PMID- 10512666 TI - On the origins of congenic MATalpha and MATa strains of the pathogenic yeast Cryptococcus neoformans. AB - The basidiomycetous yeast Cryptococcus neoformans infects humans and causes a meningoencephalitis that is uniformly fatal if untreated. The organism has a defined sexual cycle involving mating of haploid MATa and MATalpha strains, gene disruption by transformation and homologous recombination is now readily accomplished, and robust animal models for infection have been well established. In addition, a pair of congenic MATalpha and MATa haploid strains have been constructed that permit detailed studies on physiology and virulence by classical genetic approaches. These strains represent a valuable resource for further studies in this organism, and the genomic sequence of one of these strains, JEC21 (=B-4500), was recently chosen to be sequenced by an international consortium. Because of the importance of these strains for genetic studies in C. neoformans and the fact that the genomic sequence of one of these strains is in progress, we review here how these congenic strains were originally constructed. PMID- 10512667 TI - Soybean isoflavones trigger a calcium influx in Phytophthora sojae. AB - Both the motile zoospores and the hyphal germ tubes of Phytophthora sojae respond chemotropically to the soybean isoflavones daidzein and genistein. The role of Ca(2+) in the cellular response to these host signals was investigated by using X ray microanalysis of cells to monitor net changes in cellular levels of Ca(2+) and by quantifying the effects of exogenous Ca(2+) and daidzein on the developmental fate of encysted zoospores. Confirmation that isoflavones trigger a net influx of Ca(2+) into the cell was demonstrated by X-ray microanalysis of individual encysted zoospores. Zoospores exposed to 10 mM Ca(2+) and 1 microM daidzein at the time of encystment formed cysts that contained more Ca(2+) than zoospores exposed to Ca(2+) alone. The magnitude of internal Ca(2+) stores appears to be a determining factor affecting the developmental fate of P. sojae cysts. PMID- 10512668 TI - Characterization of a Neurospora crassa photolyase-deficient mutant generated by repeat induced point mutation of the phr gene. AB - We produced a photolyase-deficient mutant by repeat induced point mutation using the Neurospora crassa photolyase gene cloned previously. This mutation identified a new gene, phr, which was mapped on the right arm of linkage group I by both RFLP mapping and conventional mapping. To investigate the relationship between photoreactivation and dark repair processes, especially excision repair, double mutants of phr with representative repair-defective mutants of different types were constructed and tested for UV sensitivity and photoreactivation. The results show that the phr mutation has no influence on dark repair. Tests with CPD and TC(6-4) photoproduct-specific antibodies demonstrated that the phr mutant is defective in CPD photolyase and confirmed that there is no TC(6-4) photolyase activity in N. crassa. Furthermore, N. crassa photolyase is not a blue light receptor in the signal transduction that induces carotenoid biosynthesis. PMID- 10512669 TI - Dispersed polyphosphate in fungal vacuoles in Eucalyptus pilularis/Pisolithus tinctorius ectomycorrhizas. AB - Ectomycorrhizas produced between Pisolithus tinctorius and Eucalyptus pilularis under axenic conditions were rapidly frozen, freeze-substituted in tetrahydrofuran and embedded anhydrously, and dry-sectioned for X-ray microanalysis. The vacuoles of the sheath and Hartig net hyphae were rich in phosphorus and potassium. They also contained sulfur and variable amounts of chlorine. In anhydrously processed freeze-substituted mycorrhizas, dispersed electron-opaque material filled the fungal vacuoles. X-ray maps indicated that P was distributed evenly throughout the entire vacuole profile and was not concentrated in spherical bodies or subregions of the vacuole. There were no electron-opaque granules surrounded by electron-lucent areas, such as are commonly seen in chemically fixed material. The fungal vacuoles were also rich in K, which similarly gave a signal from the entire vacuolar profile. Such P-rich vacuoles occurred in both the mycorrhizal sheath and Hartig net hyphae. Stained sections of ether-acrolein freeze-substituted mycorrhizas also showed only dispersed material in the fungal vacuoles as, in most cases, did acetone-osmium freeze-substituted material. Precipitation of metachromatic granules by ethanol suggested that large amounts of polyphosphate are stored in these regions under the conditions of our experiments, as well as in the tips of actively growing hyphae of the same fungus. The higher plant vacuoles of ectomycorrhizas gave a much lower signal for K, and P was barely detectable. Much more K was located in the vacuoles of the root exodermal cells than in epidermal cells. The analysis of element distribution between the vacuole and cytoplasm in root cells agrees well with that found for other plant species using other techniques. We conclude that polyphosphate is indeed present in the vacuoles of the fungal cells of these ectomycorrhizas, but that in vivo it is in a dispersed form, not in granules. PMID- 10512670 TI - A quantitative trait locus of Agaricus bisporus resistance to Pseudomonas tolaasii is closely linked to natural cap color. AB - A quantitative trait locus (QTL) of resistance to Pseudomonas tolaasii was detected in Agaricus bisporus using a cross between a wild strain from the Sonoran desert and a cultivated strain. The resistance QTL was strongly linked with the brown color allele of PPC1. This QTL explained about 30% of the variation observed for living bacteria-induced symptoms. The use of bacterial toxin did not reproduce living bacteria symptoms but revealed the same QTL. The latter QTL was not affected by environmental variation. No relation was found between the resistance QTL and the tyrosinase gene, which is involved in the browning process. PMID- 10512671 TI - Partial MAT-2 gene structure and the influence of temperature on mating success in Gibberella circinata. AB - Genetic analysis of Gibberella circinata, the causative agent of pitch canker disease of pines, historically has been thwarted by a low frequency of mating success in the laboratory. We describe two findings that should facilitate genetic analysis of this fungus and related species. First, we determined that previously described degenerate primers could be used to amplify a portion of the MAT-2 mating type gene from G. circinata. This led to the cloning and sequencing of a fragment of the MAT-2 gene, which in turn made it possible to distinguish between G. circinata isolates of opposite mating types. Second, we discovered that of the 18 G. circinata field isolates in our collection, the 1 female fertile isolate expressed its fertility at 15 and 20 degrees C but not at 25 degrees C, the temperature used for crossing many Gibberella species. It is evident, therefore, that when sexual reproduction in other closely related species is initially being investigated, the crosses should be established at a variety of temperatures. Once we learned that female fertility in this G. circinata isolate was expressed at 20 degrees C, a high frequency of mating success was achieved. PMID- 10512672 TI - Microscopic analysis of Neurospora ropy mutants defective in nuclear distribution. AB - Movement and distribution of nuclei in fungi has been shown to be dependent on microtubules and the microtubule-associated motor cytoplasmic dynein. Neurospora crassa mutants known as ropy are defective in nuclear distribution. We have shown that three of the ro genes, ro-1, ro-3, and ro-4, encode subunits of either cytoplasmic dynein or the dynein activator complex, dynactin. Three other ro genes, ro-7, ro-10, and ro-11, are required for the integrity or localization of cytoplasmic dynein or dynactin. In this report, we describe a microscopic analysis of N. crassa ro mutants. Our results reveal that defects in germination of conidia, placement of septa, and mitochondrial morphology are typical of ro mutants. Two classes of cytoplasmic microtubules are identified in wild-type and ro mutants. One class of microtubules has no obvious association with nuclei while the other class of microtubules connects spindle pole bodies of adjacent nuclei. The possible role of internuclear microtubule tracts in the movement and distribution of nuclei is discussed. PMID- 10512673 TI - Deletion of the trichodiene synthase gene of Fusarium venenatum: two systems for repeated gene deletions. AB - The trichodiene synthase (tri5) gene of Fusarium venenatum was cloned from a genomic library. Vectors were created in which the tri5 coding sequence was replaced with the Neurospora crassa nitrate reductase (nit3) gene and with the Aspergillus nidulans acetamidase (amdS) gene flanked by direct repeats. The first vector was utilized to transform a nitrate reductase (niaD) mutant of F. venenatum to prototrophy, and the second vector was utilized to confer acetamide utilization to the wild-type strain. Several of the transformants lost the capacity to produce the trichothecene diacetoxyscirpenol and were shown by hybridization analysis to have gene replacements at the tri5 locus. The nit3 gene was removed by retransformation with a tri5 deletion fragment and selection on chlorate. The amdS gene was shown to excise spontaneously via the flanking direct repeats when spores were plated onto fluoroacetamide. PMID- 10512674 TI - Localization of the gene for autosomal recessive congenital hereditary endothelial dystrophy (CHED2) to chromosome 20 by homozygosity mapping. AB - Congenital hereditary endothelial dystrophy (CHED) is a corneal disorder that presents with diffuse bilateral corneal clouding. Vision may be severely impaired, and many patients require corneal transplantation. Both autosomal dominant (AD) and autosomal recessive (AR) forms of the disorder have been described. The gene responsible for AD CHED (HGMW-approved symbol CHED1) has been mapped to the pericentromeric region of chromosome 20. Investigating a large, consanguineous Irish pedigree with autosomal recessive CHED, we have previously excluded linkage to this AD CHED locus. We now describe a genome-wide search using homozygosity mapping and DNA pooling. Evidence of linkage to chromosome 20p was demonstrated with a maximum lod score of 9.30 at a recombination fraction of 0.0 using microsatellite marker D20S482. A region of homozygosity in all affected individuals was identified, narrowing the disease gene locus to an 8-cM region flanked by markers D20S113 and D20S882. This AR CHED (HGMW-approved symbol CHED2) disease gene locus is physically and genetically distinct from the AD CHED locus. PMID- 10512675 TI - Identification of genes (SPON2 and C20orf2) differentially expressed between cancerous and noncancerous lung cells by mRNA differential display. AB - mRNA differential display was applied to three small cell lung carcinoma (SCLC) cell lines, six non-small cell lung carcinoma (NSCLC) cell lines, and three normal lung tissues to identify genes differentially expressed between lung carcinoma cells and normal lung tissues and between SCLC cells and NSCLC cells. We isolated five differentially expressed genes, two that were novel and three that were already known. DIL-1 (differentially expressed in cancerous and noncancerous lung cells; HGMW-approved symbol SPON2) and pulmonary surfactant apoprotein A were expressed in normal lung tissues but not in lung carcinoma cell lines, whereas DIL-2 (HGMW-approved symbol C20orf2) and nm23-H1 were expressed in lung carcinoma cell lines but not in normal lung tissues. The remaining gene, Annexin II, was expressed at a lower level in SCLC than in NSCLC and normal lung tissues. These genes were also differentially expressed in primary lung cancers. One of the two novel genes, DIL-1, encodes a secreted protein homologous to the Mindin/F-spondin family. The other, DIL-2, encodes a protein with a putative ATP/GTP binding site motif. These data provide basic information necessary to understand the differences in gene expression profiles between lung carcinoma and normal lung and between SCLC and NSCLC. Further characterization of these genes will help to clarify the molecular mechanisms of human lung carcinogenesis. PMID- 10512676 TI - An olfactory receptor gene is located in the extended human beta-globin gene cluster and is expressed in erythroid cells. AB - An olfactory receptor gene was identified near the 3' breakpoint of a naturally occurring deletion (HPFH-1) in the human beta-globin gene cluster on chromosome 11p15.5. The gene encodes an amino acid sequence that is 40 to 51% identical to that of a set of olfactory receptors that have only recently been identified as a distinct family of receptors. There are two orthologous genes in the mouse that encode amino acid sequences that are 73 and 71% identical, respectively, to that encoded by the human gene. This olfactory receptor gene is expressed at the RNA level in human and murine erythroid cells at all stages of development. This aberrant expression is probably due to the location of the gene in the transcriptionally active chromatin domain of the extended beta-globin gene cluster in erythroid cells. PMID- 10512677 TI - Primate evolution of an olfactory receptor cluster: diversification by gene conversion and recent emergence of pseudogenes. AB - The olfactory receptor (OR) subgenome harbors the largest known gene family in mammals, disposed in clusters on numerous chromosomes. We have carried out a comparative evolutionary analysis of the best characterized genomic OR gene cluster, on human chromosome 17p13. Fifteen orthologs from chimpanzee (localized to chromosome 19p15), as well as key OR counterparts from other primates, have been identified and sequenced. Comparison among orthologs and paralogs revealed a multiplicity of gene conversion events, which occurred exclusively within OR subfamilies. These appear to lead to segment shuffling in the odorant binding site, an evolutionary process reminiscent of somatic combinatorial diversification in the immune system. We also demonstrate that the functional mammalian OR repertoire has undergone a rapid decline in the past 10 million years: while for the common ancestor of all great apes an intact OR cluster is inferred, in present-day humans and great apes the cluster includes nearly 40% pseudogenes. PMID- 10512678 TI - Human and mouse ISLR (immunoglobulin superfamily containing leucine-rich repeat) genes: genomic structure and tissue expression. AB - We have previously reported a transcript of the novel gene for human immunoglobulin superfamily containing leucine-rich repeat (ISLR). By additional screening of a human retina cDNA library, we isolated another type of transcript with a 5' UTR different from that of the previously reported type. Genomic sequencing of the ISLR gene revealed that these two types of transcripts, ISLR-1 and ISLR-2, originated from the same gene but are composed of different first exons. Because the entire open reading frame is contained in the second exon, these two transcripts produce the same protein. Radiation hybrid mapping linked the ISLR gene to AFM248yh1, which is in the critical region of Bardet-Biedl syndrome type 4 (BBS4) on chromosome 15. Sequence analysis of the ISLR gene in five BBS4 patients, however, showed no mutations, although a few polymorphic changes were detected. Cloning of the mouse homolog of ISLR (Islr) revealed that the predicted protein consists of 428 amino acids, 86% of which are identical to those of ISLR. The Islr gene was expressed in various mouse tissues, including retina, in which Islr mRNA was detected in the ganglion cell layer, the inner nuclear layer, and the inner segment of the photoreceptor. PMID- 10512679 TI - Identification and characterization of TESK2, a novel member of the LIMK/TESK family of protein kinases, predominantly expressed in testis. AB - In this study we present the cDNA sequence of a novel putative protein kinase, denoted TESK2. The open reading frame of TESK2 encodes a putative 555-amino-acid protein, including a protein kinase consensus sequence in the N-terminal half. The protein kinase domain of TESK2 is structurally similar to the kinase domain of the protein serine/threonine kinase TESK1 (64% identity) and to those of the LIMK1 and LIMK2 kinases (42 and 39% identity, respectively). TESK2, together with TESK1, constitutes a second subgroup of the LIMK/TESK family of protein kinases, as revealed by phylogenetic analysis of the protein kinase domains. Chromosomal localization of human TESK2 was assigned to 1p32. Expression analysis of human TESK2 revealed a single mRNA species of 3.0 kb predominantly expressed in testis and prostate and low expression in most other tissues examined. Rat testicles expressed a single species of TESK2 mRNA of approximately 3.5 kb. However, the transcript was first detectable in rat testis after day 30 of postnatal development and was predominantly expressed in round spermatids. These observations suggest that TESK2 plays an important role in spermatogenesis. PMID- 10512680 TI - The FSHD region on human chromosome 4q35 contains potential coding regions among pseudogenes and a high density of repeat elements. AB - The distal end of chromosome 4q contains the locus involved in facioscapulohumeral muscular dystrophy (FSHD1). Specific genomic deletions within a tandem DNA repeat (D4Z4) are associated with the disease status, but no causal genes have yet been discovered. In a systematic search for genes, a 161-kb stretch of genomic DNA proximal to D4Z4 was sequenced, analyzed for homologies, and subjected to gene prediction programs. A major fraction (45%) of the subtelomeric region is composed of repeat sequences attributable mainly to LINE-1 elements. Apart from the previously identified FRG1 and TUB4q sequences, several additional potential coding regions were identified by analyzing the sequence with exon prediction programs. So far, we have been unable to demonstrate transcripts by RT-PCR or cDNA library hybridization. However, several retrotransposed pseudogenes were identified. The high density of pseudogenes and repeat elements is consistent with the subtelomeric location of this region and explains why previous transcript identification studies have been problematic. PMID- 10512681 TI - Cloning and characterization of T-cell lymphoma invasion and metastasis 2 (TIAM2), a novel guanine nucleotide exchange factor related to TIAM1. AB - TIAM1 is a guanine nucleotide exchange factor that was identified in a screen for genes that increase the invasiveness of T lymphoma cell lines (Habets et al., 1994, Cell 77(4): 537-549). We have identified a gene, T-cell lymphoma invasion and metastasis 2 (HGMW-approved symbol TIAM2), with significant identity to the carboxyl-terminal region of the TIAM1 and mapped it to 6q25. TIAM2 is expressed as an approximately 3.3-kb transcript in cerebrum and as an approximately 4.4-kb transcript in the cerebellum and testis. The approximately 4. 4-kb message encodes a longer form of the approximately 3.3-kb mRNA predicted protein, and both contain homology to the Dbl-homologous region (70%) and Pleckstrin homologous (54%) regions of TIAM1. We have purified TIAM2 and shown it to have GDP-GTP exchange activity. In situ hybridizations demonstrate TIAM2 expression in the E13.5 telencephalon of mouse embryos and in the cerebral cortex, hippocampus, and ependyma of adult mouse brains. PMID- 10512682 TI - Human carbonic anhydrase XIV (CA14): cDNA cloning, mRNA expression, and mapping to chromosome 1. AB - A full-length cDNA clone of a human carbonic anhydrase XIV (HGMW-approved gene symbol CA14) was obtained and sequenced. The cDNA sequence was 1757 bp long and was predicted to encode a 337-amino-acid polypeptide with a molecular mass of 37.6 kDa. The deduced amino acid sequence of CA XIV showed an overall similarity of 29-46% to other active CA isozymes. The highest percentage similarity was with a transmembrane CA isoform, CA XII. As observed for CA XII, CA XIV has hydrophobic segments at both termini of the deduced protein for a putative signal sequence and a transmembrane domain. CA XIV showed low activity and was sensitive to acetazolamide, but not to sulfonamide. Northern blot analysis demonstrated an approximately 1.7-kb transcript in the adult human heart, brain, liver, and skeletal muscle. RNA dot-blot analysis for CA XIV mRNA expression showed a strong signal in all parts of the human brain and a weaker signal in the colon, small intestine, urinary bladder, and kidney. RT-PCR analysis showed an intense signal in the liver and spinal cord and a faint signal in the kidney. No CA XIV mRNA was seen in the salivary gland and pancreas. In contrast, CA XII mRNA was expressed in the kidney, salivary gland, and pancreas, but not in the liver or spinal cord. The CA XIV gene was localized to human chromosome 1q21. These findings indicate genetically distinct but closely related isoforms of human transmembrane CAs, CA XII and CA XIV, which have different patterns of tissue-specific expression. PMID- 10512683 TI - Genomic cloning and characterization of the human homeobox gene SIX6 reveals a cluster of SIX genes in chromosome 14 and associates SIX6 hemizygosity with bilateral anophthalmia and pituitary anomalies. AB - The Drosophila gene sine oculis (so), a nuclear homeoprotein that is required for eye development, has several homologues in vertebrates (the SIX gene family). Among them, SIX3 is considered to be the functional orthologue of so because it is strongly expressed in the developing eye. However, embryonic SIX3 expression is not limited to the eye field, and SIX3 has been found to be mutated in some patients with holoprosencephaly type 2 (HPE2), suggesting that SIX3 has wide implications in head development. We report here the cloning and characterization of SIX6, a novel human SIX gene that is the homologue of the chick Six6(Optx2) gene. SIX6 is closely related to SIX3 and is expressed in the developing and adult human retina. Data from chick and mouse suggest that the human SIX6 gene is also expressed in the hypothalamic and the pituitary regions. SIX6 spans 2567 bp of genomic DNA and is split in two exons that are transcribed into a 1393 nucleotide-long mRNA. Chromosomal mapping of SIX6 revealed that it is closely linked to SIX1 and SIX4 in human chromosome 14q22.3-q23, which provides clues about the origin and evolution of the vertebrate SIX family. Recently three independent reports have associated interstitial deletions at 14q22.3-q23 with bilateral anophthalmia and pituitary anomalies. Genomic analyses of one of these cases demonstrated SIX6 hemizygosity, strongly suggesting that SIX6 haploinsufficiency is responsible for these developmental disorders. PMID- 10512684 TI - Detection and cloning of an X-linked locus associated with a NotI site that is not methylated on mouse inactivated X chromosome by the RLGS-M method. AB - In looking for genes that escape X chromosome inactivation, we scanned the methylation status of genomic DNA from XX, X0, and XY mice using the method of restriction landmark genomic scanning using methylation-sensitive endonuclease. We detected and cloned a candidate locus and identified the Orf1 gene. Orf1 has sequence similarities to the B2 repetitive element and human CXORF4 (formerly called EXLM1), which escapes X inactivation. The B2 element spans the 3' terminus of the ORF and the 3' UTR of Orf1. The Orf1 gene encompasses 18.5 kb of genomic DNA including 11 exons and 10 introns. Taking advantage of genomic polymorphisms present between MSM and C3H/He, we showed that murine Orf1 is mapped to the proximal region of the X chromosome. Despite the unmethylation of the NotI site, Orf1 is subject to X inactivation. PMID- 10512685 TI - A missense mutation in the beta-2 integrin gene (ITGB2) causes canine leukocyte adhesion deficiency. AB - Canine leukocyte adhesion deficiency (CLAD) is a fatal immunodeficiency disease found in Irish setters. The clinical manifestations of CLAD are very similar to LAD in humans and BLAD in cattle, which are both caused by mutations in ITGB2 encoding the leukocyte integrin beta-2 subunit (CD18). Sequence analysis of the ITGB2 coding sequence from a CLAD dog and a healthy control revealed a single missense mutation, Cys36Ser. This cysteine residue is conserved among all beta integrins, and the mutation most likely disrupts a disulfide bond. The mutation showed a complete association with CLAD in Irish setters and was not found in a sample of dogs from other breeds. The causative nature of this mutation was confirmed by transduction experiments using retroviral vectors and human LAD EBV B-cells. The normal canine CD18 formed heterodimers with the human CD11 subunit, whereas gene transfer of the mutant CD18 resulted in very low levels of CD11/CD18 expression. The identification of the causative mutation for CLAD now makes it possible to identify carrier animals with a simple diagnostic DNA test, and it forms the basis for using CLAD as a large animal model for the development and evaluation of clinical treatments for human LAD. PMID- 10512686 TI - Radiation hybrid and cytogenetic mapping of SOCS1 and SOCS2 to chromosomes 16p13 and 12q, respectively. AB - Suppressor of cytokine signaling (SOCS) proteins are involved in the negative regulation of cytokine-induced STAT (signal transducers and activators of transcription) factor signaling. We cloned genomic regions of SOCS1 and SOCS2 genes and mapped these genes to chromosome 16p12-p13.1 and chromosome 12q21.3-q23 regions, respectively, by cytogenetic and radiation hybrid mapping. In addition, we mapped SOCS2 by yeast artificial chromosome (YAC) pool screening to YAC contig WC 12.5 on chromosome 12 with an unambiguous hit to CHLC.ATA19H12 and WI-5940, which is 461.5 cR from the top of the map. PMID- 10512687 TI - Hexarelin, a growth hormone - releasing peptide, counteracts bone loss in gonadectomized male rats. AB - The age-related decline in growth hormone (GH) secretion has been implicated in the pathogenesis of involutional bone loss. Whether restoration of GH secretion might be helpful in maintaining and/or improving bone mass during aging is still unsettled. The aim of the present study was to examine the effects of 30-day treatment with hexarelin (HEXA, 50 microg/kg subcutaneously b.i.d.), a highly effective GH-releasing compound, on bone metabolism and bone mineral density (BMD) in intact and osteopenic gonadectomized (GDX) mature male rats. Serum total alkaline phosphatase (ALP, bone formation marker) and bone resorption markers (lysylpyridinoline, LP and hydroxylysylpyridinoline, HP) were measured before and 7, 14 and 30 days after treatment. BMD was measured by dual-energy X-ray absorptiometry at lumbar vertebrae, femoral metaphysis and diaphysis before and at the end of the experiment. In intact rats, HEXA significantly (P<0.05) decreased LP (-36.3%) and HP (-22.8%) excretion at day 7, whereas it did not change serum ALP activity and BMDs. In GDX rats, HEXA completely prevented the significant (P<0. 01) increase in urinary excretion of both LP (+143.8%) and HP (+119. 4%), the early decrease in ALP activity (-26.5%) and the significant (P<0.05) decrease in BMDs in the femoral metaphysis (-7.9%) and lumbar vertebrae (-6.8%) caused by androgen deficiency. The bone-protective effects of HEXA could be attributed, at least in part, to its GH-releasing activity since chronic treated rats maintained the GH response to an acute challenge with HEXA. The evidence that HEXA, unlike GH, inhibits bone resorption indicates that other mechanisms contribute to the bone sparing effect of HEXA. PMID- 10512688 TI - Maximum anaerobic performance of childhood-onset GH-deficient adults. AB - To date, physical capacity of adults with GH deficiency (GHD) has been studied in terms of muscle strength, contractile properties and aerobic performance. As a result, scanty data are available regarding the maximum anaerobic performance of these patients with reference to healthy controls. Therefore, the objective of this study was to evaluate maximum anaerobic power of adults with GHD and of age matched controls by two methods, one testing lactacid power (w.;(c)) through a 15 s-maximal bout on a bicycle ergometer, the other testing alactic power (w.;(j)) through a vertical jump on a force platform. Absolute w.;(c)and w.;(j)values were both found to be 35% lower(P<0.04) in GHD patients than in controls. Similarly, peak pedalling velocity (V(peak)) was 21% lower (P<0.04) in patients. When w.;(c)and w.;(j)were respectively normalized for thigh and lower limb muscle plus bone volumes and V(peak)for muscle length, differences between patients and controls were no longer significant. Furthermore, the rate of power loss during the cycling bout was approximately 35% in both groups. This observation was in line with similar delta (peak minus baseline) lactate capillary blood concentrations, being 6.3 mM/l in patients and 7.5 mM/l in controls (NS). Lactacid capacity, which represents the energy extracted from lactate metabolism, normalized for body mass was similar in the two groups. In conclusion, the maximum anaerobic power that can be developed by short-statured childhood-onset GHD adults is significantly lower in terms of absolute values, but not different from that of controls once appropriately normalized. Therefore, the changes in maximum anaerobic power of GH deficient patients seem to be a consequence of their smaller muscle mass. PMID- 10512689 TI - IGF-I treatment increases motility and improves morphology of immature spermatozoa in the GH-deficient dwarf (dw/dw) rat. AB - It has recently been shown that short-term growth hormone (GH) treatment can increase the motility of spermatozoa in the GH-deficient dw/dw rat. To examine whether the effects of GH on motility of immature spermatozoa are mediated by an increase in plasma concentrations of IGF-I, we treated GH-deficient dw/dw rats with 2 microg/g/day of IGF-I using osmotic minipumps. Body weight (saline 227+/-5 g, IGF-I 253+/-4 g) and IGF-I concentrations in blood plasma (saline 472+/-19.9 ng/ml, IGF-I 986+/-43.6 ng/ml) and seminal vesicle fluid (saline 30.9+/-1.7 ng/ml, IGF-I 47.9+/-2.9 ng/ml) were significantly increased with IGF-I treatment (P<0.001), similar to the observed responses to GH therapy in our earlier study. While epididymal fluid IGF-I concentrations were not changed, IGF-I treatment significantly increased the number of immature motile spermatozoa (saline 14.4+/ 3.5%, IGF-I 28.3+/-4.1%, P<0.05) and the number of spermatozoa with normal morphology (control 65.7+/-3.3%, IGF-I 75+/-1.9%, P<0.05). These data suggest that increasing the circulating concentrations of IGF-I in the GH-deficient rat can improve the motility and morphology of immature spermatozoa and thus mimic, at least in part, the effects of GH. PMID- 10512690 TI - Inflammation-related neutrophil proteases, cathepsin G and elastase, function as insulin-like growth factor binding protein proteases. AB - Over the past few years, several proteolytic enzymes have been identified as insulin-like growth factor binding protein (IGFBP) proteases. It has been suggested that proteolytic cleavage of IGFBPs is associated with regulation of the proliferative effects of IGFs on their target cells. In this study, we have demonstrated that two neutrophil proteases, cathepsin G and elastase, effectively cleave IGFBPs in vitro and in vivo at concentrations lower than previous described IGFBP proteases. Purified leukocyte cathepsin G and elastase cleaved all six well-characterized IGFBPs into distinct fragments in a concentration dependent manner. Under similar experimental conditions, cathepsin G preferentially cleaved IGFBP-5, followed by BP-2, BP-3, BP-4, BP-1, and BP-6. In comparison, elastase equally preferred IGFBP-3 and IGFBP-4, followed by BP-1, BP 5, BP-6, and BP-2. Proteolysis of rh(125)I-IGFBP-3 by cathepsin G was blocked by alpha(1)-antichymotrypsin, while elastase proteolytic activity was blocked by alpha(1)-proteinase inhibitor as expected. Elastase, but not cathepsin G, cleaved free IGF-I into a smaller molecular weight fragment in vitro, possibly designating unique functions for each protease within the IGF axis. Sequence analysis of IGFBP-3 fragments produced by cathepsin G and elastase demonstrated that each protease cleaved IGFBP-3 at unique sites within its midregion. More importantly, extracts from purified neutrophils have demonstrated significant proteolytic cleavage of IGFBP-3 that resembles elastase proteolysis of IGFBP-3. Recent studies using a monocyte-like cell model have also shown significant cleavage of IGFBP-3. These in vitro and in vivo data suggest that the neutrophil proteases, cathepsin G and elastase, in addition to their previously described functions as extracellular matrix-degrading enzymes, may potentially act as IGFBP proteases involved in regulation of IGFs and IGFBPs during inflammation and wound healing. PMID- 10512691 TI - Strength of colonic anastomoses and skin incisional wounds in old rats - influence by diabetes and growth hormone. AB - The influence of advanced age on the mechanical strength of colonic anastomoses and skin incisional wounds in diabetic rats was investigated after 0 (suture binding capacity) and after 7 days of healing. Furthermore, the effects of growth hormone (GH) injections to old diabetic rats were investigated. Diabetes in old rats did not influence the strength of colonic anastomoses after 0 and 7 days. However, in these diabetic animals, the strength of skin incisional wounds was reduced by 27% after 7 days of healing (P< 0.01). GH injections administered to old diabetic rats doubled the mortality compared with that of saline-injected old diabetic rats (P< 0.01). GH injections did not influence the strength formation of either colonic anastomoses or skin incisional wounds in old normal rats. In conclusion, the healing of colonic anastomoses in diabetic rats was not compromised by old age, while the strength of skin wounds was decreased. PMID- 10512692 TI - Intrinsic cardiac muscle function, calcium handling and beta -adrenergic responsiveness is impaired in rats with growth hormone deficiency. AB - To evaluate whether growth hormone (GH) is required for normal cardiac muscle function, we studied left ventricular papillary muscles of mutant GH-deficient rats. Developed tension normalized by cross-sectional area (DT), intracellular [Ca(2+)](i)(aequorin method) and beta-adrenergic responsiveness were assessed with or without 3 weeks GH replacement therapy and compared to normal controls. Steady-state force-Ca(2+)relationship was determined in tetanized ryanodine treated muscles. beta-adrenergic responsiveness was tested during graded isoproterenol stimulation. [Ca(2+)](i)at baseline and the EC(50)of the force Ca(2+)relationship were similar in all groups. In dwarf rats, DT at baseline was reduced by 43% compared to controls, due to a decreased maximal Ca(2+)-activated force. beta-adrenergic responsiveness of systolic Ca(2+)-release and mechanical function were depressed in dwarf rats. GH treatment caused at least partial improvement of the depressed parameters. These data support the hypothesis that GH is required for normal intrinsic function of cardiac muscle by maintaining Ca(2+)- and beta-adrenergic responsiveness. PMID- 10512693 TI - Normal growth despite GH, IGF-I and IGF-II deficiency. AB - A 6.5-year-old male with normal linear growth, despite septo-optic dysplasia, panhypopituitarism and a deficient GH/IGF axis, is presented. In addition to measuring IGF-I, IGF-II and IGFBP-3, serum IGFBP-1, -2, -4 and -5 were measured. A human osteosarcoma cell line was used to assess growth-promoting activity in the patient's serum. The role of leptin in linear growth in this case was investigated. There was no evidence for hyperinsulinism or hyperandrogenism. GH was undetectable upon multiple stimulation. GHBP was elevated. Serum IGF-I (25 microg/l), IGF-II (194 microg/l), IGFBP-3 (0.4 mg/l), and IGFBP-5 (87 microg/l) levels were low compared to age-matched prepubertal children. Serum IGFBP-4 level was normal. Molecular size of IGF-II in the patient's serum was normal, suggesting normal IGF-II bioavailability. Human osteosarcoma cell proliferation in response to the patient's serum was similar to sera from age-matched normal controls. Leptin levels were markedly elevated. Osteoblast cell proliferation was not stimulated by leptin. The data demonstrate that normal growth and osteoblast cell proliferation in this patient is not mediated by GH, total IGFs, insulin, or leptin, and suggest the presence of a yet unidentified growth factor or mechanism. The case offers a detailed picture of binding proteins in a case of growth without GH. It introduces osteoblast cell proliferation as a method of assessing serum growth-promoting activity in such cases. It adds IGF-II and leptin to the list of excluded growth-promoting candidates in GH-independent growth, and further demonstrates our incomplete understanding of the phenomenon of growth. PMID- 10512694 TI - Yeast polypeptide fusion surface display levels predict thermal stability and soluble secretion efficiency. AB - Efficiency of yeast cell surface display can serve as a proxy screening variable for enhanced thermal stability and soluble secretion efficiency of mutant proteins. Several single-chain T cell receptor (scTCR) single-site mutants that enable yeast surface display, along with their double and triple mutant combinations, were analyzed for soluble secretion from the yeast Saccharomyces cerevisiae. While secretion of the wild-type scTCR was not detected, each of the single, double, and triple mutants were produced in yeast supernatants, with increased expression resulting from the double and triple mutants. Soluble secretion levels were strongly correlated with the quantity of active scTCR displayed as a fusion to Aga2p on the surface of yeast. Thermal stability of the scTCR mutants correlated directly with the secreted and surface levels of scTCR, with evidence suggesting that intracellular proteolysis by the endoplasmic reticulum quality control apparatus dictates display efficiency. Thus, yeast display is a directed evolution scaffold that can be used for the identification of mutant eucaryotic proteins with significantly enhanced stability and secretion properties. PMID- 10512695 TI - The N-terminal domain of MDM2 resembles calmodulin and its relatives. AB - It is shown here that the N-terminal domain of MDM2, which is not thought to bind calcium ions, otherwise bears a striking resemblance to a cluster of four EF-hand modules like those found in the calmodulin family. There are similarities in module arrangement, supersecondary structure and the main-chain to main-chain hydrogen-bonding pattern, especially in the vicinity of the short antiparallel beta-sheet, the two strands of which lie between the two E and F helices of tandem modules. Some conserved amino acid residues are identified that are associated with short side-chain to main-chain hydrogen-bonded motifs. Also, both types of domain bind a short, functionally important hydrophobic alpha-helix from another protein in a cavity between the two pairs of EF-hand, or EF-hand-like, modules. PMID- 10512696 TI - Single molecular observation of the interaction of GroEL with substrate proteins. AB - To understand the mechanism of GroEL-assisted protein folding, we observed the interaction of fluorescence-labeled GroEL with fluorescence-labeled substrate proteins at the single molecule level by total internal reflection fluorescence microscopy. GroEL with a A133C mutation in the equatorial domain was labeled with a fluorescent dye, tetramethylrhodamine. As substrate proteins, we used the largely denatured and partly denatured forms of bovine beta-lactoglobulin, both labeled with another fluorescent dye, Cy5. The complexes formed by GroEL with these substrates were characterized by size-exclusion gel chromatography. The recovered complexes were then observed by fluorescence microscopy. For both substrates, agreement of the fluorescent spots for tetramethylrhodamine and Cy5 indicated formation of the complex at the single molecule level. Similar observation of macroscopic binding by size-exclusion chromatography and microscopic binding by the fluorescence microscopy was done for the folding intermediate of Cy5-labeled bovine rhodanese. The fluorescence microscopy opens a new avenue for studying the interaction of GroEL with substrate proteins. PMID- 10512697 TI - Separate contributions of UhpA and CAP to activation of transcription of the uhpT promoter of Escherichia coli. AB - Activation of promoters by multiple transcription factors might occur through favorable contacts of the activators with themselves or RNA polymerase, or by changes in DNA geometry that enhance formation of the transcription complex. Transcription of the Escherichia coli uhpT gene, encoding the organophosphate transporter, requires the response regulator UhpA and is stimulated by the global regulator protein CAP. CAP binds to the uhpT promoter at a single site, centered at -103.5 bp relative to the start of transcription, and UhpA binds to multiple sites between positions -80 and -32. Overexpression of UhpA did not reduce the degree of CAP stimulation of uhpT-lacZ expression, showing that CAP action is more complex than enhancement of the binding of UhpA. Footprinting experiments demonstrated that UhpA and CAP modestly stimulated each other's binding to the uhpT promoter, but did not affect the positioning of the binding sites. An in vitro transcription system was used to examine the contribution of each transcription factor at the uhpT promoter. Action of UhpA and CAP proteins was not affected by template supercoiling. Kinetic analyses of productive and abortive initiation showed that CAP acted both to stabilize by fivefold the open promoter complexes formed in the presence of UhpA and to enhance by twofold the rate of their formation. These results indicate that open complex formation requires UhpA and that CAP stabilizes the open complex. PMID- 10512698 TI - The maturase encoded by a group I intron from Aspergillus nidulans stabilizes RNA tertiary structure and promotes rapid splicing. AB - The AnCOB group I intron from Aspergillus nidulans self-splices, providing the Mg2+ concentration is >/= 15 mM. The splicing reaction is greatly stimulated by a maturase protein encoded within the intron itself. An initial structural and biochemical analysis of the splicing reaction has now been performed. The maturase bound rapidly to the precursor RNA (kon approximately 3 x 10(9) M(-1) min(-1)) and remained tightly bound (koff /= 150 mM) was tenfold slower, in part because of the existence of an equilibrium between folded and partially folded RNA. In contrast, the maturase very effectively stabilized tertiary structure in 5 mM Mg2+, a noticeable example being an interaction between the P8 helix and a GNRA sequence that constitutes the L2 terminal loop of the P2 helix. Formation of the 5' splice-site recognition helix was assisted by either the maturase or high concentrations of Mg2+. The maturase was required during splicing so it is not a true chaperone. However, RNase protection assays and kinetic studies suggest that the maturase recognizes and facilitates folding of an intron with limited tertiary structure and even incomplete secondary structure. PMID- 10512699 TI - High-level expression in Escherichia coli of selenocysteine-containing rat thioredoxin reductase utilizing gene fusions with engineered bacterial-type SECIS elements and co-expression with the selA, selB and selC genes. AB - Mammalian thioredoxin reductase (TrxR) catalyzes reduction of thioredoxin and many other substrates, and is a central enzyme for cell proliferation and thiol redox control. The enzyme is a selenoprotein and can therefore, like all other mammalian selenoproteins, not be directly expressed in Escherichia coli, since selenocysteine-containing proteins are synthesized by a highly species-specific translation machinery. This machinery involves a secondary structure, SECIS element, in the selenoprotein-encoding mRNA, directing selenocysteine insertion at the position of an opal (UGA) codon, normally conferring termination of translation. It is species-specific structural features and positions in the selenoprotein mRNA of the SECIS elements that hitherto have hampered heterologous production of recombinant selenoproteins. We have discovered, however, that rat TrxR can be expressed in E. coli by fusing its open reading frame with the SECIS element of the bacterial selenoprotein formate dehydrogenase H. A variant of the SECIS element designed to encode the conserved carboxyterminal end of the enzyme (-Sec-Gly-COOH) and positioning parts of the SECIS element in the 3'-untranslated region was also functional. This finding revealed that the SECIS element in bacteria does not need to be translated for full function and it enabled expression of enzymatically active mammalian TrxR. The recombinant selenocysteine containing TrxR was produced at dramatically higher levels than formate dehydrogenase O, the only endogenous selenoprotein expressed in E. coli under the conditions utilized, demonstrating a surprisingly high reserve capacity of the bacterial selenoprotein synthesis machinery under aerobic conditions. Co expression with the selA, selB and selC genes (encoding selenocysteine synthase, SELB and tRNA(Sec), respectively) further increased the efficiency of the selenoprotein production and thereby also increased the specific activity of the recombinant TrxR to about 25 % of the native enzyme, with as much as 20 mg produced per liter of culture. These results show that with the strategy utilized here, the capacity of selenoprotein synthesis in E. coli is more than sufficient for making possible the use of the bacteria for production of recombinant selenoproteins. PMID- 10512700 TI - The influence of helix morphology on co-operative polyamide backbone conformational flexibility in peptide nucleic acid complexes. AB - A systematic analysis of peptide nucleic acid (PNA) complexes deposited in the Protein Data Bank has been carried out using a set of contiguous atom torsion angle definitions. The analysis is complemented by molecular mechanics adiabatic potential energy calculations on hybrid PNA-nucleic acid model systems. Hitherto unobserved correlations in the values of the (alpha and epsilon) dihedral angles flanking the backbone secondary amide bond are found. This dihedral coupling forms the basis of a PNA backbone conformation classification scheme. Six conformations are thus characterised in experimental structures. Helix morphology is found to exert a significant influence on backbone conformation and flexibility: Watson-Crick PNA strands in complexes with DNA and RNA, that possess A-like base-pair stacking, adopt backbone conformations distinct from those in PNA.DNA-PNA triplex and PNA-PNA duplex P-helix forms. Solvation effects on Watson Crick PNA backbone conformation in heterotriplexes are discussed and the possible involvement of inter-conformational transitions and dihedral angle uncoupling in asymmetric heteroduplex base-pair breathing is suggested. PMID- 10512701 TI - Conservation of structure and function among histidine-containing phosphotransfer (HPt) domains as revealed by the crystal structure of YPD1. AB - In Saccharomyces cerevisiae, the SLN1-YPD1-SSK1 phosphorelay system controls a downstream mitogen-activated protein (MAP) kinase in response to hyperosmotic stress. YPD1 functions as a phospho-histidine protein intermediate which is required for phosphoryl group transfer from the sensor kinase SLN1 to the response regulator SSK1. In addition, YPD1 mediates phosphoryl transfer from SLN1 to SKN7, the only other response regulator protein in yeast which plays a role in response to oxidative stress and cell wall biosynthesis. The X-ray structure of YPD1 was solved at a resolution of 2.7 A by conventional multiple isomorphous replacement with anomalous scattering. The tertiary structure of YPD1 consists of six alpha-helices and a short 310-helix. A four-helix bundle comprises the central core of the molecule and contains the histidine residue that is phosphorylated. Structure-based comparisons of YPD1 to other proteins having a similar function, such as the Escherichia coli ArcB histidine-containing phosphotransfer (HPt) domain and the P1 domain of the CheA kinase, revealed that the helical bundle and several structural features around the active-site histidine residue are conserved between the prokaryotic and eukaryotic kingdoms. Despite limited amino acid sequence homology among HPt domains, our analysis of YPD1 as a prototypical family member, indicates that these phosphotransfer domains are likely to share a similar fold and common features with regard to response regulator binding and mechanism for histidine-aspartate phosphoryl transfer. PMID- 10512702 TI - Dissecting the stability of a beta-hairpin peptide that folds in water: NMR and molecular dynamics analysis of the beta-turn and beta-strand contributions to folding. AB - NMR studies of the folding and conformational properties of a beta-hairpin peptide, several peptide fragments of the hairpin, and sequence-modified analogues, have enabled the various contributions to beta-hairpin stability in water to be dissected. Temperature and pH-induced unfolding studies indicate that the folding-unfolding equilibrium approximates to a two-state model. The hairpin is highly resistant to denaturation and is still significantly folded in 7 M urea at 298 K. Thermodynamic analysis shows the hairpin to fold in water with a significant change in heat capacity, however, DeltaCp degrees in 7 M urea is reduced. V/Y-->A mutations on one strand of the hairpin reduce folding to <10 %, consistent with a hydrophobic stabilisation model. We show that in a truncated peptide (residues 6-16) lacking the hydrophobic residues on one beta-strand, the type I' Asn-Gly turn in the sequence SINGKK is significantly populated in water in the absence of interstrand hydrophobic contacts. Unrestrained molecular dynamics simulations of unfolding, using an explicit solvation model, show that the conformation of the NG turn persists for longer than the AG analogue, which has a much lower propensity for type I' turn formation from a data base analysis of preferred turns. The origin of the high stability of the Asn-Gly turn is not entirely clear; data base analysis of 66 NG turns, together with molecular dynamics simulations, reveals no participation of the Asn side-chain in turn stabilising interactions with the peptide backbone. However, hydration analysis of the molecular dynamics simulations reveals a pocket of "high density" water bridging between the Asn side-chain and peptide main-chain that suggests solvent mediated interactions may play an important role in modulating phi,psi propensities in the NG turn region. PMID- 10512703 TI - Solution structure of the ribosomal protein S19 from Thermus thermophilus. AB - Ribosomal protein S19 is a 10.6 kDa protein in the small subunit of the prokaryotic ribosome. We have determined a high-resolution solution structure of S19 from Thermus thermophilus. Structures were calculated using 1160 distance and dihedral angle restraints derived from (1)H, (15)N and (13)C NMR spectra. The structures show that S19 is a mixed alpha/beta protein with long disordered tails. The folding topology is not homologous to that of any other known protein structure. Potential rRNA and protein binding sites have been identified on the S19 surface. PMID- 10512704 TI - Backbone dynamics of a short PU.1 ETS domain. AB - The resonance assignments, secondary structure and backbone dynamics of the ETS domain of the transcription factor PU.1 have been determined for the free protein in solution by NMR spectroscopy. The secondary structure for the free ETS domain is similar to that observed in the crystal structure of the PU.1 protein complexed with DNA, except that helix alpha2 and recognition helix alpha3 are shorter for the free protein in solution. Backbone dynamics of the protein have been examined using amide hydrogen-deuterium exchange and (15)N laboratory-frame spin relaxation measurements. A significant probability of local unfolding of helix alpha2, which precedes the loop-helix-loop DNA recognition domain, is inferred from the very fast hydrogen-deuterium exchange for amide protons in this helix. The (15)N relaxation measurements indicate that the protein is partially oligomerized at a concentration of 2.5 mM, but monomeric at a concentration of 0.3 mM. The (15)N relaxation data for the low concentration sample were interpreted, using the model-free formalism, to provide insight into protein dynamics on picosecond-nanosecond and microsecond-millisecond time scales. High flexibility of the protein backbone is observed for the residues in the loop between alpha2 and alpha3. This loop is variable in length and in structure within the class of winged helix proteins and is partially responsible for binding to DNA. The dynamic properties observed for alpha2, alpha3 and the intervening loop may indicate a correlation between protein plasticity in particular structural elements and recognition of specific DNA sequences. PMID- 10512705 TI - Solution structure of the DNA-binding domain of NtrC with three alanine substitutions. AB - The structure of the 20 kDa C-terminal DNA-binding domain of NtrC from Salmonella typhimurium (residues Asp380-Glu469) with alanine replacing Arg456, Asn457, and Arg461, was determined by NMR spectroscopy. NtrC is a homodimeric enhancer binding protein that activates the transcription of genes whose products are required for nitrogen metabolism. The 91-residue C-terminal domain contains the determinants necessary for dimerization and DNA-binding of the full length protein. The mutant protein does not bind to DNA but retains many characteristics of the wild-type protein, and the mutant domain expresses at high yield (20 mg/l) in minimal medium. Three-dimensional (1)H/(13)C/(15)N triple-resonance, (1)H (13)C-(13)C-(1)H correlation and (15)N-separated nuclear Overhauser effect (NOE) spectroscopy experiments were used to make backbone and side-chain (1)H,(15)N, and (13)C assignments. The structures were calculated using a total of 1580 intra and inter-monomer distance and hydrogen bond restraints (88 hydrogen bonds; 44 hydrogen bond restraints), and 88 phi dihedral restraints for residues Asp400 through Glu469 in both monomers. A total of 54 ambiguous restraints (intra or inter-monomer) involving residues close to the 2-fold symmetry axis were also included. Each monomer consists of four helical segments. Helices A (Trp402 Leu414) and B (Leu421-His440) join with those of another monomer to form an antiparallel four-helix bundle. Helices C (Gln446-Leu451) and D (Ala456-Met468) of each monomer adopt a classic helix-turn-helix DNA-binding fold at either end of the protein. The backbone rms deviation for the 28 best of 40 starting structures is 0.6 (+/-0.2) A. Structural differences between the C-terminal domain of NtrC and the homologous Factor for Inversion Stimulation are discussed. PMID- 10512706 TI - Structural analysis of a non-contiguous second-site revertant in T4 lysozyme shows that increasing the rigidity of a protein can enhance its stability. AB - The mutation Glu108-->Val (E108V) in T4 lysozyme was previously isolated as a second-site revertant that specifically compensated for the loss of function associated with the destabilizing substitution Leu99-->Gly (L99G). Surprisingly, the two sites are 11 A apart, with Leu99 in the core and Glu108 on the surface of the protein. In order to better understand this result we have carried out a detailed thermodynamic, enzymatic and structural analysis of these mutant lysozymes as well as a related variant with the substitution Leu99-->Ala. It was found that E108V does increase the stability of L99G, but it also increases the stability of both the wild-type protein and L99A by essentially equal amounts. The effects of E108V on enzymatic activity are more complicated. The mutation slightly reduces the maximal rate of cell wall hydrolysis of wild-type, L99G and L99A. At the same time, L99G is an unstable protein and rapidly loses activity during the course of the assay, especially at temperatures above 20 degrees C. Thus, even though the double mutant L99G/E108V has a slightly lower maximal rate than L99G, over a period of 20-30 minutes it hydrolyzes more substrate. This decrease in the rate of thermal inactivation appears to be the basis of the action of E108V as a second-site revertant of L99G. Mutant L99A creates a cavity of volume 149 A(3). Instead of enlarging this cavity, mutant L99G results in a 4 5 A displacement of part of helix F (residues 108-113), creating a solvent accessible declivity. In the double mutant, L99G/E108V, this helix returns to a position akin to wild-type, resulting in a cavity of volume 203 A(3). Whether the mutation Glu108-->Val is incorporated into either wild-type lysozyme, or L99A or L99G, it results in a decrease in crystallographic thermal factors, especially in the helices that include residues 99 and 108. This increase in rigidity, which appears to be due to a combination of increased hydrophobic stabilization plus a restriction of conformational fluctuation, provides a structural basis for the increase in thermostability. PMID- 10512707 TI - Magnesium(II) and zinc(II)-protoporphyrin IX's stabilize the lowest oxygen affinity state of human hemoglobin even more strongly than deoxyheme. AB - Studies of oxygen equilibrium properties of Mg(II)-Fe(II) and Zn(II)-Fe(II) hybrid hemoglobins (i.e. alpha2(Fe)beta2(M) and alpha2(M)beta2(Fe); M=Mg(II), Zn(II) (neither of these closed-shell metal ions binds oxygen or carbon monoxide)) are reported along with the X-ray crystal structures of alpha2(Fe)beta2(Mg) with and without CO bound. We found that Mg(II)-Fe(II) hybrids resemble Zn(II)-Fe(II) hybrids very closely in oxygen equilibrium properties. The Fe(II)-subunits in these hybrids bind oxygen with very low affinities, and the effect of allosteric effectors, such as proton and/or inositol hexaphosphate, is relatively small. We also found a striking similarity in spectrophotometric properties between Mg(II)-Fe(II) and Zn(II)-Fe(II) hybrids, particularly, the large spectral changes that occur specifically in the metal containing beta subunits upon the R-T transition of the hybrids. In crystals, both alpha2(Fe)beta2(Mg) and alpha2(Fe-CO)beta2(Mg) adopt the quaternary structure of deoxyhemoglobin. These results, combined with the re-evaluation of the oxygen equilibrium properties of normal hemoglobin, low-affinity mutants, and metal substituted hybrids, point to a general tendency of human hemoglobin that when the association equilibrium constant of hemoglobin for the first binding oxygen molecule (K1) approaches 0.004 mmHg(-1), the cooperativity as well as the effect of allosteric effectors is virtually abolished. This is indicative of the existence of a distinct thermodynamic state which determines the lowest oxygen affinity of human hemoglobin. Moreover, excellent agreement between the reported oxygen affinity of deoxyhemoglobin in crystals and the lowest affinity in solution leads us to propose that the classical T structure of deoxyhemoglobin in the crystals represents the lowest affinity state in solution. We also survey the oxygen equilibrium properties of various metal-substituted hybrid hemoglobins studied over the past 20 years in our laboratory. The bulk of these data are consistent with the Perutz's trigger mechanism, in that the affinity of a metal hybrid is determined by the ionic radius of the metal, and also by the steric effect of the distal ligand, if present. However, there remains a fundamental contradiction among the oxygen equilibrium properties of the beta substituted hybrid hemoglobins. PMID- 10512708 TI - The molten globule state of a chimera of human alpha-lactalbumin and equine lysozyme. AB - The molten globule state of equine lysozyme is more stable than that of alpha lactalbumin and is stabilized by non-specific hydrophobic interactions and native like hydrophobic interactions. We constructed a chimeric protein which is produced by replacing the flexible loop (residues 105-110) in human alpha lactalbumin with the helix D (residues 109-114) in equine lysozyme to investigate the possible role of the helix D for the high stability and native-like packing interaction in the molten globule state of equine lysozyme. The stability of the molten globule state formed by the chimeric protein to guanidine hydrochloride induced unfolding is the same as that of equine lysozyme and is substantially greater than that of human alpha-lactalbumin, although only six residues come from equine lysozyme. Our results also suggest that the non-native interaction in the molten globule state of alpha-lactalbumin changes to the native-like packing interaction due to helix substitution. The solvent-accessibility of the Trp residues in the molten globule state of the chimeric protein is similar to that in the molten globule state of equine lysozyme in which packing interaction around the Trp residues in the native state is partially preserved. Therefore, the helix D in equine lysozyme is one of the contributing factors to the high stability and native-like packing interaction in the molten globule state of equine lysozyme. Our results indicate that the native-like packing interaction can stabilize the rudimentary intermediate which is stabilized by the non specific hydrophobic interactions. PMID- 10512709 TI - Large-scale effects of transcriptional DNA supercoiling in vivo. AB - The scale of negative DNA supercoiling generated by transcription in Top(+) Escherichia coli cells was assessed from the efficiency of cruciform formation upstream of a regulated promoter. An increase in negative supercoiling upon promoter induction led to cruciform formation, which was quantitatively measured by chemical probing of intracellular DNA. By placing a cruciform-forming sequence at varying distances from the promoter, we found that the half-dissociation length of transcription supercoiling wave is approximately 800 bp. This is the first proof that transcription can affect DNA structure on such a remarkably large scale in vivo. Moreover, cooperative binding of the cI repressor to the upstream promoter DNA did not preclude efficient diffusion of transcriptional supercoiling. Finally, our plasmids appeared to contain discrete domains of DNA supercoiling, defined by the features and relative orientation of different promoters. PMID- 10512710 TI - Isolation of kasugamycin resistant mutants in the 16 S ribosomal RNA of Escherichia coli. AB - Three ribosomal RNA mutations conferring resistance to the antibiotic kasugamycin were isolated using a strain of Escherichia coli in which all of the rRNA is transcribed from a plasmid-encoded rrn operon. The mutations, A794G, G926A, and A1519C, mapped to universally conserved sites in the 16 S RNA gene. Site-directed mutagenesis studies showed that virtually all mutations at these three sites conferred kasugamycin resistance and had very slight effects on cell growth. It has been known for many years that the absence of post-transcriptional modification at A1519 and the adjacent A1518 in strains lacking a functional KsgA methylase produces a kasugamycin resistance phenotype. Mutations at A1519 conferred kasugamycin resistance and had minor effects on cell growth, whereas mutations at 1518 did not confer resistance and increased the doubling time of the cells dramatically. Expression of mutations at A1518/A1519 in a methylase deficient ksgA(-)strain had divergent effects on the phenotype of the rRNA mutants, suggesting that the base identity at either position does not affect methylation at the adjacent site. Residues A794 and G926 are protected from chemical modification by kasugamycin and tRNA, and have been implicated in the initiation of protein synthesis. Despite the universal conservation and functional importance of these residues, the results presented here show that the identity of the bases is not critical for ribosomal function. PMID- 10512711 TI - Active site dynamics of the HhaI methyltransferase: insights from computer simulation. AB - A molecular dynamics study was performed on the DNA methyltransferase M. Hha I in a ternary complex with DNA and AdoMet in solution. Methylation involves addition of the Cys81 sulfhydryl anion to the 6-position of Cyt18, followed by a nucleophilic attack of the resultant carbanion at C5 on the AdoMet methyl group. It was found in this simulation that the distances between the sulfhydryl group (SG) of Cys81 to the C6 of Cyt18 (SG-C6) and methyl carbon (CH3) of AdoMet to the C5 of cytosine (CH3-C5) are dependent on the dihedral angle chi (O4'-C1'-N1-C2) of the nucleotide. When the chi angle of Cyt18 is low (< -80 degrees), the SG-C6 and CH3-C5 distances are large. A high chi angle (> -80 degrees) for the target cytosine residue reduces the distances for both SG-C6 and CH3-C5, and the angles formed between the cytosine ring and AdoMet correspond well to values for the transition state structures formed during methylation of cytosine from ab initio calculations. Two possible proton sources for protonation of N3 of the cytosine residue upon formation of the covalent intermediate were found in the simulation. The protonated amine group of AdoMet could provide a proton via a water bridge, or Arg163 could also be the source of the proton for N3 via a water bridge. The simulation provides insights into how the H5 of cytosine could go from the active site into solvent. Conserved residues Asn304 and Gln82 stabilize a water network within the active site of M. Hha I which provides a route for H5 to diffuse into bulk solvent. An initially distant water molecule was able to diffuse into the active site of the enzyme and replace a position of a crystallographic water molecule in close proximity to the C5 of cytosine. The movement of this water molecule showed that a channel exists between Gln82 and the AdoMet in M. Hha I which allows both water and protons to easily gain access to the active site of the enzyme. PMID- 10512712 TI - Site-specific in vivo cleavages by DNA topoisomerase I in the regulatory regions of the 35 S rRNA in Saccharomyces cerevisiae are transcription independent. AB - Eukaryotic type I DNA topoisomerase controls DNA topology by transiently breaking and resealing one strand of DNA at a time. During transcription and replication its action reduces the torsional stress derived from these activities. The association of DNA topoisomerase I with the nucleolus has been reported and this enzyme was shown to be involved in yeast rDNA metabolism. Here, we have investigated the in vivo presence of DNA topoisomerase I cleavage sites in the non-transcribed spacer of the rDNA cluster. We show a specific profile of highly localized cleavage in relevant areas of this region. The sites are detected in the promoter and in the enhancer regions of the 35 S gene. The analysis of mutants in which transcription is prevented and/or reduced, namely a strain lacking the 43 kDa subunit of RNA polymerase I, a second one that does note transcribe, lacking a subunit of the core factor and another member of the RNA polymerase I transcription factors lacking one of the UAF component which transcribes at very low level, show that DNA topoisomerase I cleavage sites are not related to transcription by RNA polymerase I. These findings point to a role for DNA topoisomerase I that is additional to the commonly recognized function in removing the transcription-induced topological stress. PMID- 10512713 TI - Cloning and characterization of hurpin (protease inhibitor 13): A new skin specific, UV-repressible serine proteinase inhibitor of the ovalbumin serpin family. AB - Epidermal keratinocytes are the primary target of the midrange ultraviolet part (UVB, 280-320 nm) of terrestrial sunlight. Analysis of the resulting UV response at the transcriptional level by differential display PCR identified a formerly unrecognized large group of repressed genes. Among those UV-repressible genes, a novel serine proteinase inhibitor (serpin) termed hurpin (HaCaT UV-repressible serpin) has been identified. The isolated full-length cDNAs harbour a 1176 bp open reading frame encoding a potential protein with 391 amino acid residues and a predicted molecular mass of approximately 44 kDa. The novel serpin has nearly 59 % amino acid identity with the squamous cell carcinoma antigen 1 (SCCA1) and squamous cell carcinoma antigen 2 (SCCA2). In addition, it displays all of the structural features unique to the ovalbumin family of serpins (ov-serpins). The amino acid sequence of the hinge region in the reactive site loop suggests that hurpin has the potential for protease inhibition. The putative reactive center P1 P1'residues were identified as Thr356-Ser357 by alignment with other ov-serpins. The physiological target protease is unknown and the in vitro translated hurpin does not form SDS-stable complexes with a variety of known serine proteases. Expression of hurpin is restricted to epidermal cells where two distinct transcripts of 3.0 and 3.4 kb are detectable. Furthermore, expression of hurpin appears to be related to the activation or proliferation state of keratinocytes, since hurpin transcripts are more abundant in immortalized keratinocytes (HaCaT) and in cultured normal human keratinocytes, compared to the expression in normal skin. Moreover, in psoriasis, a skin disease characterized by hyperproliferation of keratinocytes and responsive to therapeutic UV irradiation, overexpression of hurpin is noted in psoriatic skin lesions compared to non-lesional skin. PMID- 10512714 TI - Bispecific tandem diabody for tumor therapy with improved antigen binding and pharmacokinetics. AB - To increase the valency, stability and therapeutic potential of bispecific antibodies, we designed a novel recombinant molecule that is bispecific and tetravalent. It was constructed by linking four antibody variable domains (VHand VL) with specificities for human CD3 (T cell antigen) or CD19 (B cell marker) into a single chain construct. After expression in Escherichia coli, intramolecularly folded bivalent bispecific antibodies with a mass of 57 kDa (single chain diabodies) and tetravalent bispecific dimers with a molecular mass of 114 kDa (tandem diabodies) could be isolated from the soluble periplasmic extracts. The relative amount of tandem diabodies proved to be dependent on the length of the linker in the middle of the chain and bacterial growth conditions. Compared to a previously constructed heterodimeric CD3xCD19 diabody, the tandem diabodies exhibited a higher apparent affinity and slower dissociation from both CD3(+)and CD19(+)cells. They were also more effective than diabodies in inducing T cell proliferation in the presence of tumor cells and in inducing the lysis of CD19(+)cells in the presence of activated human PBL. Incubated in human serum at 37 degrees C, the tandem diabody retained 90 % of its antigen binding activity after 24 hours and 40 % after one week. In vivo experiments indicated a higher stability and longer blood retention of tandem diabodies compared to single chain Fv fragments and diabodies, properties that are particularly important for potential clinical applications. PMID- 10512715 TI - Ribulose bisphosphate carboxylase/oxygenase from the hyperthermophilic archaeon Pyrococcus kodakaraensis KOD1 is composed solely of large subunits and forms a pentagonal structure. AB - We previously reported the presence of a highly active, carboxylase-specific ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) in a hyperthermophilic archaeon, Pyrococcus kodakaraensis KOD1. In this study, structural analysis of Pk -Rubisco has been performed. Phylogenetic analysis of Rubiscos indicated that archaeal Rubiscos, including Pk -Rubisco, were distinct from previously reported type I and type II enzymes in terms of primary structure. In order to investigate the existence of small subunits in native Pk -Rubisco, immunoprecipitation and native-PAGE experiments were performed. No specific protein other than the expected large subunit of Pk -Rubisco was detected when the cell-free extracts of P. kodakaraensis KOD1 were immunoprecipitated with polyclonal antibodies against the recombinant enzyme. Furthermore, native and recombinant Pk -Rubiscos exhibited identical mobilities on native-PAGE. These results indicated that native Pk -Rubisco consisted solely of large subunits. Electron micrographs of purified recombinant Pk -Rubisco displayed pentagonal ring-like assemblies of the molecules. Crystals of Pk -Rubisco obtained from ammonium sulfate solutions diffracted X-rays beyond 2.8 A resolution. The self-rotation function of the diffraction data showed the existence of 5-fold and 2-fold axes, which are located perpendicularly to each other. These results, along with the molecular mass of Pk -Rubisco estimated from gel filtration, strongly suggest that Pk Rubisco is a decamer composed only of large subunits, with pentagonal ring-like structure. This is the first report of a decameric assembly of Rubisco, which is thought to belong to neither type I nor type II Rubiscos. PMID- 10512716 TI - Folding and association of the antibody domain CH3: prolyl isomerization preceeds dimerization. AB - The simplest naturally occurring model system for studying immunoglobulin folding and assembly is the non-covalent homodimer formed by the C-terminal domains (CH3) of the heavy chains of IgG. Here, we describe the structure of recombinant CH3 dimer as determined by X-ray crystallography and an analysis of the folding pathway of this protein. Under conditions where prolyl isomerization does not contribute to the folding kinetics, formation of the beta-sandwich structure is the rate-limiting step. beta-Sheet formation of CH3 is a slow process, even compared to other antibody domains, while the subsequent association of the folded monomers is fast. After long-time denaturation, the majority of the unfolded CH3 molecules reaches the native state in two serial reactions, involving the re-isomerization of the Pro35-peptide bond to the cis configuration. The species with the wrong isomer accumulate as a monomeric intermediate. Importantly, the isomerization to the correct cis configuration is the prerequisite for dimerization of the CH3 domain. In contrast, in the Fab fragment of the same antibody, prolyl isomerization occurs after dimerization demonstrating that within one protein, comprised of highly homologous domains, both the kinetics of beta-sandwich formation and the stage at which prolyl isomerization occurs during the folding process can be completely different. PMID- 10512717 TI - Crystal structure of an Fab fragment in complex with a meningococcal serosubtype antigen and a protein G domain. AB - Many pathogens present highly variable surface proteins to their host as a means of evading immune responses. The structure of a peptide antigen corresponding to the subtype P1.7 variant of the porin PorA from the human pathogen Neisseria meningitidis was determined by solution of the X-ray crystal structure of the ternary complex of the peptide (ANGGASGQVK) in complex with a Fab fragment and a domain from streptococcal protein G to 1.95 A resolution. The peptide adopted a beta-hairpin structure with a type I beta-turn between residues Gly4P and Gly7P, the conformation of the peptide being further stabilised by a pair of hydrogen bonds from the side-chain of Asn2P to main-chain atoms in Val9P. The antigen binding site within the Fab formed a distinct crevice lined by a high proportion of apolar amino acids. Recognition was supplemented by hydrogen bonds from heavy chain residues Thr50H, Asp95H, Leu97H and Tyr100H to main-chain and side-chain atoms in the peptide. Complementarity-determining region (CDR) 3 of the heavy chain was responsible for approximately 50 % of the buried surface area formed by peptide-Fab binding, with the remainder made up from CDRs 1 and 3 of the light chain and CDRs 1 and 2 of the heavy chain. Knowledge of the structures of variable surface antigens such as PorA is an essential prerequisite to a molecular understanding of antigenic variation and its implications for vaccine design. PMID- 10512718 TI - Structure of the complex of the antistasin-type inhibitor bdellastasin with trypsin and modelling of the bdellastasin-microplasmin system. AB - The serine proteinase plasmin is, together with tissue-type plasminogen activator (tPA) and urokinase-type plasminogen activator (uPA), involved in the dissolution of blood clots in a fibrin-dependent manner. Moreover, plasmin plays a key role in a variety of other activation cascades such as the activation of metalloproteinases, and has also been implicated in wound healing, pathogen invasion, cancer invasion and metastasis. The leech-derived (Hirudo medicinalis) antistasin-type inhibitor bdellastasin represents a specific inhibitor of trypsin and plasmin and thus offers a unique opportunity to evaluate the concept of plasmin inhibition. The complexes formed between bdellastasin and bovine as well as porcine beta-trypsin have been crystallised in a monoclinic and a tetragonal crystal form, containing six molecules and one molecule per asymmetric unit, respectively. Both structures have been solved and refined to 3.3 A and 2.8 A resolution. Bdellastasin turns out to have an antistasin-like fold exhibiting a bis-domainal structure like the tissue kallikrein inhibitor hirustasin. The interaction between bdellastasin and trypsin is restricted to the C-terminal subdomain of bdellastasin, particularly to its primary binding loop, comprising residues Asp30-Glu38. The reactive site of bdellastasin differs from other antistasin-type inhibitors of trypsin-like proteinases, exhibiting a lysine residue instead of an arginine residue at P1. A model of the bdellastasin microplasmin complex has been created based on the X-ray structures. Our modelling studies indicate that both trypsin and microplasmin recognise bdellastasin by interactions which are characteristic for canonically binding proteinase inhibitors. On the basis of our three-dimensional structures, and in comparison with the tissue-kallikrein-bound and free hirustasin and the antistasin structures, we postulate that the binding of the inhibitors toward trypsin and plasmin is accompanied by a switch of the primary binding loop segment P5-P3. Moreover, in the factor Xa inhibitor antistasin, the core of the molecule would prevent an equivalent rotation of the P3 residue, making exosite interactions of antistasin with factor Xa imperative. Furthermore, Arg32 of antistasin would clash with Arg175 of plasmin, thus impairing a favourable antistasin-plasmin interaction and explaining its specificity. PMID- 10512719 TI - Residual dipolar coupling derived orientational constraints on ligand geometry in a 53 kDa protein-ligand complex. AB - The geometric relationships between ligands and the functional groups that bind ligands in soluble ligand-protein complexes have traditionally been deduced from distance constraints between pairs of NMR active nuclei spanning the ligand protein interface. Frequently, the steep inverse distance dependence of the nuclear Overhauser effect (NOE), from which the distance constraints are derived, makes identification of sufficient numbers of constraints difficult. In these cases the ability to supplement NOE-derived information with distance-independent angular information can be very important. Here, the observation of residual dipolar couplings from alpha-methyl mannose bound to mannose binding-protein in a dilute liquid crystalline medium has allowed the determination of a bound ligand's average orientation. The 3-fold rotational symmetry of mannose-binding protein defines its orientational tensor and obviates the need to determine experimentally the protein's average orientation. Through superimposition of ligand and protein orientational tensors we describe the binding geometry of alpha-methyl mannose bound to mannose-binding protein. This new method is of general applicability to the study of ligands bound to proteins, and it is of particular interest when neither X-ray crystallography nor NOE techniques can provide sufficient information to describe binding geometries. PMID- 10512720 TI - Calorimetric analysis of the Ca(2+)-binding betagamma-crystallin homolog protein S from Myxococcus xanthus: intrinsic stability and mutual stabilization of domains. AB - The betagamma-crystallin superfamily consists of a class of homologous two-domain proteins with Greek-key fold. Protein S, a Ca(2+)-binding spore-coat protein from the soil bacterium Myxococcus xanthus exhibits a high degree of sequential and structural homology with gammaB-crystallin from the vertebrate eye lens. In contrast to gammaB-crystallin, which undergoes irreversible aggregation upon thermal unfolding, protein S folds reversibly and may therefore serve as a model in the investigation of the thermodynamic stability of the eye-lens crystallins. The thermal denaturation of recombinant protein S (PS) and its isolated domains was studied by differential scanning calorimetry in the absence and in the presence of Ca(2+) at varying pH. Ca(2+)-binding leads to a stabilization of PS and its domains and increases the cooperativity of their equilibrium unfolding transitions. The isolated N-terminal and C-terminal domains (NPS and CPS) obey the two-state model, independent of the pH and Ca(2+)-binding; in the case of PS, under all conditions, an equilibrium intermediate is populated. The first transition of PS may be assigned to the denaturation of the C-terminal domain and the loss of domain interactions, whereas the second one coincides with the denaturation of the isolated N-terminal domain. At pH 7.0, in the presence of Ca(2+), where PS exhibits maximal stability, the domain interactions at 20 degrees C contribute 20 kJ/mol to the overall stability of the intact protein. PMID- 10512721 TI - Chaperonin-affected refolding of alpha-lactalbumin: effects of nucleotides and the co-chaperonin GroES. AB - We have studied how nucleotides (ADP, AMP-PNP, and ATP) and the co-chaperonin GroES influence the GroEL-affected refolding of apo-alpha-lactalbumin. The refolding reactions induced by stopped-flow pH jumps were monitored by alpha lactalbumin tryptophan fluorescence. The simple single-exponential character of the free-refolding kinetics of the protein allowed us to quantitatively analyze the kinetic traces of the GroEL-affected refolding with the aid of computer simulations, and to obtain the best-fit parameters for binding between GroEL and the refolding intermediate of alpha-lactalbumin by the non-linear least-squares method. When GroES was absent, the interaction between GroEL and alpha lactalbumin could be well represented by a "cooperative-binding" model in which GroEL has two binding sites for alpha-lactalbumin with the affinity of the second site being tenfold weaker than that of the first, so that there is negative cooperativity between the two sites. The affinity between GroEL and alpha lactalbumin was significantly reduced when ATP was present, while ADP and AMP-PNP did not alter the affinity. A comparison of this result with those reported previously for other target proteins suggests a remarkable adjustability of the GroEL 14-mer with respect to the nucleotide-induced reduction of affinity. When GroES was present, ATP as well as ADP and AMP-PNP were effective in reducing the affinity between GroEL and the refolding intermediate of alpha-lactalbumin. The affinity at a saturating concentration of ADP or AMP-PNP was about ten times lower than with GroEL alone. The ADP concentration at which the acceleration of the GroEL/ES-affected refolding of alphaLA was observed, was higher than the concentration at which the nucleotide-induced formation of the GroEL/ES complex took place. These results indicate that GroEL/ES complex formation itself is not enough to reduce the affinity for alpha-lactalbumin, and that further binding of the nucleotide to the GroEL/ES complex is required to reduce the affinity. PMID- 10512722 TI - How NF-kappaB can be attracted by its cognate DNA. AB - NF-kappaB is involved in the transcriptional regulation of a large number of genes, in particular those of human immunodeficiency virus (HIV). Recently, we used NMR spectroscopy and molecular modelling to study the solution structure of a native duplex related to the HIV-1 kappaB site, together with a mutated duplex for which a three base-pair change abolishes NF-kappaB binding. The native duplex shows unusual dynamics of the four steps surrounding the kappaB site. Here, we explore the intrinsic properties of the NMR-refined structures of both duplexes in order to understand why the native sequence is recognised by NF-kappaB among other DNA sequences. We establish that only the native kappaB site can adopt a conformation where its structure (curvature and base displacement), the accessibility and the electrostatic potentials of key atoms become very favourable for binding the large loops of NF-kappaB, in contrast to the mutated duplex. Finally, we show that the neutralisation of phosphate groups contacted by NF-kappaB favours a more canonical DNA structure. These findings lead to a new hypothesis for specific recognition through the phosphodiester backbone dynamics of the sequences flanking a binding site. Such unusual behaviour confers upon the overall duplex properties that can be used by NF-kappaB to select its binding site. Thus, the selectivity determinants for NF-kappaB binding appear to depend on deformability of an "extended" consensus sequence. PMID- 10512723 TI - A census of protein repeats. AB - In this study, we analyzed all known protein sequences for repeating amino acid segments. Although duplicated sequence segments occur in 14 % of all proteins, eukaryotic proteins are three times more likely to have internal repeats than prokaryotic proteins. After clustering the repetitive sequence segments into families, we find repeats from eukaryotic proteins have little similarity with prokaryotic repeats, suggesting most repeats arose after the prokaryotic and eukaryotic lineages diverged. Consequently, protein classes with the highest incidence of repetitive sequences perform functions unique to eukaryotes. The frequency distribution of the repeating units shows only weak length dependence, implicating recombination rather than duplex melting or DNA hairpin formation as the limiting mechanism underlying repeat formation. The mechanism favors additional repeats once an initial duplication has been incorporated. Finally, we show that repetitive sequences are favored that contain small and relatively water-soluble residues. We propose that error-prone repeat expansion allows repetitive proteins to evolve more quickly than non-repeat-containing proteins. PMID- 10512724 TI - Tenascin is expressed in the mesenchyme of the embryonic lung and down-regulated by dexamethasone in early organogenesis. AB - Tenascin (TN) is a hexameric extracellular matrix glycoprotein that is temporally and spatially restricted during lung development. This study examines the expression and regulation of TN in early lung organogenesis. Two TN isoforms were detected in total RNA isolated from embryonic day 14 rat lung tissues by reverse transcriptase polymerase chain reaction. The localization of TN in embryonic day 14 rat lung tissues was investigated by using in situ hybridization performed with an antisense RNA probe. TN mRNA was expressed exclusively by the mesenchyme but not by the epithelium of embryonic rat lungs. The intense expression of TN was observed in the mesenchyme that immediately surrounds the growing epithelial cells of the developing bronchi. The effect of the synthetic glucocorticoid hormone dexamethasone on the regulation of TN expression was examined by in vitro lung explant culture. Two TN polypeptides, the larger (M(r) 230 kDa, TN230) polypeptide and the smaller (M(r) 180 kDa, TN180) isoform, were detected in embryonic day 21 rat lungs by immunoblot analysis with anti-TN antibody. Dexamethasone inhibited both TN230 and TN180 biosynthesis. The study demonstrates the expression of TN at the early stage of lung organogenesis and presents evidence of hormonal regulation of TN in lung development, suggesting a potential role of TN in the communication between the epithelial and mesenchymal cells during lung branching morphogenesis. PMID- 10512725 TI - Differential expression of protein kinase C isoform transcripts in human hematopoietic progenitors undergoing differentiation. AB - Protein kinase C (PKC), a key component of the signaling pathways leading to proliferation and differentiation, consists of a family closely related serine/threonine protein kinases. The mRNA expression of these PKC isoforms has been characterized during hematopoietic differentiation. Using the reverse transcriptase polymerase chain reaction technique, we have analyzed the levels of isoform transcripts in bone marrow CD34(+) hematopoietic progenitors and their progeny differentiated along erythroid, megakaryocyte, or granulocyte/monocyte lineages, upon exposure to growth factors. In contrast with isoforms alpha, beta(I), beta(II), delta, and epsilon, ubiquitously expressed, isoforms theta, eta/L, zeta, and iota/lambda exhibited a lineage-restricted expression. These qualitative changes, which allow to distinguish the erythroid and megakaryocyte phenotypes from the granulocyte/monocyte phenotype, include zeta exclusively upregulated in granulocytes/monocytes and theta, eta/L, and iota/lambda exclusively expressed in megakaryocytes and erythroblasts. In contrast, erythroblasts and megakaryocytes, which supposedly share a common bipotential progenitor, displayed only quantitative changes. These results evidence the selective expression of PKC isoforms at transcriptional and/or posttranscriptional levels in hematopoietic progenitors induced to differentiate, which may suggest a differential contribution of individual isoforms to cellular signaling. PMID- 10512726 TI - Isolation and characterization of a novel proteinase inhibitor from the snake serum of Taiwan habu (Trimeresurus mucrosquamatus). AB - A proteinase inhibitor (designated as TMI) was isolated and purified from the snake serum of Taiwan habu (Trimeresurus mucrosquamatus) by using successive chromatographies which included Sephadex G-100, DEAE-Sephacel chromatographies, and C(4) reverse-phase HPLC. The purified inhibitor was shown to be a homogeneous protein with a molecular mass of about 47 or 36 kDa in the presence or absence of a reducing agent, beta-mercaptoethanol. The inhibitor decreases in molecular mass by about 23% with N-linked neuraminidase treatment, suggesting that it is a glycoprotein. Further enzymatic analyses indicated that this inhibitor possesses strong inhibitory activities toward three zinc-dependent metalloproteinases and not fibrinogenolytic serine proteases previously isolated from the venom of the same snake species with an IC(50) of about 0.2-1.1 microM. Its IC(50) value was approximately three orders of magnitude more effective than those of the tripeptide inhibitors we previously purified from the crude venom of the same snake (Biochem. Biophys. Res. Commun. 248, 562-568 (1998)). The purified inhibitor showed stronger inhibitory action against caseinolytic activities of crude venoms from closely related species of Taiwan habu than those from unrelated species. N-terminal sequence analysis showed that its sequence is distinctly different from sequences of those serum inhibitors reported for other snake species in the literature. Based on inhibition susceptibility and primary structures of various snake protease inhibitors, it is suggested that this novel inhibitor isolated from the serum of Taiwan habu may be a unique self-defense protein factor mainly for protection against envenomation from snakes of the same genus. PMID- 10512727 TI - Wild-type enzyme as a reporter of inhibitor binding by catalytically impaired mutant enzymes. AB - A method for the determination of inhibition constants for catalytically debilitated mutant enzymes is described. The inhibitor is partitioned between the mutant and wild-type enzymes. Catalytic rates of the wild-type enzyme are used as the signal of inhibitor binding to the mutant enzyme. The method is validated with scytalone dehydratase, the Y50F mutant, and a potent inhibitor. The K(i) value for Y50F determined by this method is 0.49 +/- 0.10 nM. The K(i) value determined using the Y50F catalytic report for inhibitor binding in the absence of wild-type enzyme is 0.20 +/- 0.030 nM. The wild-type enzyme binds the inhibitor ten-fold less tightly, thus indicating that the hydrogen-bonding interaction between the Y50 hydroxyl group and the inhibitor (suggested by X-ray crystallography) is weak. The method is most useful when the catalytic activity of the wild-type enzyme is the most sensitive report of inhibitor binding and the mutant enzyme is greatly crippled in catalytic activity. PMID- 10512728 TI - Heparin differentially regulates the interaction of fibroblast growth factor-4 with FGF receptors 1 and 2. AB - Fibroblast growth factor-4 (FGF4), like other FGFs, shares a high affinity for the anionic glycosaminoglycans heparin and heparan sulfate (HS), which in turn enhance FGF-receptor (FGFR) binding and activation. Here we demonstrate using a cell free system that, at low concentrations of heparin, FGF4 binds only to FGFR 2, while much higher heparin levels are required for binding to FGFR-1. Chemical crosslinking of radiolabeled FGF4 to the soluble FGF receptors confirms the preferential formation of FGF4-FGFR-2 complexes under restricted heparin availability, with maximal ligand-receptor interactions at almost 20-fold lower heparin concentrations then those required for the affinity labeling of FGFR-1. In accordance, HS-deficient cells expressing FGFR-2 proliferate in response to FGF4 at extremely low exogenous heparin concentrations, while FGFR-1 expressing cells are completely unresponsive under the same conditions. We suggest that FGFR 2 is the preferred receptor for FGF4 under restricted HS conditions and that the bioavailability of structurally distinct HS motifs may differentially control receptor specificity of FGF4 in vivo. PMID- 10512729 TI - Hepatocyte isolation stimulates formation of interferon stimulatory response element DNA-protein complexes. AB - We have examined the relationship between intracellular signalling pathways and loss of differentiated function during hepatocyte isolation and culture. We have shown that isolation induces the activation of the interferon stimulatory response element (ISRE). This activation was transient and peaked at 3 h before it returned to basal by 24 h of culture. Interferon regulatory factor-1 (IRF-1) was shown to be important for generation of ISRE complexes by electromobility shift assays and supershift intervention. IRF-1 was translocated to the nucleus in parallel with changes to ISRE complex formation. The p38 kinase inhibitor, SB 203580, diminished the formation of ISRE binding complexes. Hence p38 kinase may be involved in the activation and binding of IRF-1 or related proteins to the ISRE motif. Changes in ISRE activation levels in cultured hepatocytes may have important implications in primary hepatocyte differentiation and loss of function. PMID- 10512730 TI - Sulfation of iodothyronines by recombinant human liver steroid sulfotransferases. AB - Sulfation is an important pathway in the metabolism of thyroid hormones. Sulfated iodothyronines are elevated in nonthyroidal illnesses and in the normal human fetal circulation. We assayed and characterized COS-1 cell expressed recombinant human liver dehydroepiandrosterone sulfotransferase (DHEA ST or SULT2A1) and estrogen sulfotransferase (EST or SULT1E1) activities for the first time with triiodothyronine (T(3)) as the substrate. Several biochemical properties that included apparent K(m) values, thermal stabilities, and responses to the inhibitors 2, 6-dichloro-4-nitrophenol and NaCl were tested. SULT2A1, a member of the hydroxysteroid sulfotransferase family, used 3,3'-T(2) more readily than T(3) and 3,5-T(2) as substrates, but had the lowest apparent K(m) value for T(3) of any reported human SULT. SULT1E1, a member of the phenol sulfotransferase family, used 3,3'-T(2) and rT(3) more readily than T(3), and also displayed the greatest specificity for T(4) among human SULTs. SULT2A1 may contribute more to iodothyronine sulfation than previously suspected. Potential roles of both steroid sulfotransferases in the enhanced sulfation of nonthyroidal illnesses and fetal development invite further investigation. PMID- 10512731 TI - Xylanase II from an alkaliphilic thermophilic Bacillus with a distinctly different structure from other xylanases: evolutionary relationship to alkaliphilic xylanases. AB - A 1.0 kilobase gene fragment from the genomic DNA of an alkaliphilic thermophilic Bacillus was found to code for a functional xylanase (XynII). The complete nucleotide sequence including the structural gene and the 5' and 3' flanking sequences of the xylanase gene have been determined. An open reading frame starting from ATG initiator codon comprising 402 nucleotides gave a preprotein of 133 amino acids of calculated molecular mass 14.090 kDa. The occurrence of three potential N-glycosylation sites in XynII gene is a unique feature for a gene of bacterial origin. The stop codon was followed by hairpin loop structures indicating the presence of transcription termination signals. The secondary structure analysis of XynII predicted that the polypeptide was primarily formed of beta-sheets. XynII appeared to be a member of family G/11 of xylanases based on its molecular weight and basic pI (8.0). However, sequence homology revealed similar identity with families 10 and 11 of xylanases. The conserved triad (Val Val-Xaa, where Xaa is Asn or Asp) was identified only in the xylanases from alkaliphilic organisms. Our results implicate for the first time the concept of convergent evolution for XynII and provide a basis for research in evolutionary relationship among the xylanases from alkaliphilic and neutrophilic organisms. PMID- 10512732 TI - Isolation and characterization of a novel antifungal peptide from Aspergillus niger. AB - A novel antifungal peptide (termed as Anafp) was isolated from the culture supernatant of the filamentous fungi, Aspergillus niger. The whole amino acid sequence of Anafp was determined and the peptide was found to be composed of a single polypeptide chain with 58 amino acids including six cysteine residues. The peptide shows some degree of sequence homology to a cysteine-rich antifungal peptides reported from the seeds of Sinapis alba and Arabidopsis thaliana or the extracellular media of Aspergillus giganteus and Penicillium chrysogenumsome. Cysteine-spacing pattern of Anafp was similar to that of the antifungal peptide from Penicillium chrysogenum. The Anafp exhibited potent growth inhibitory activities against yeast strains as well as filamentous fungi at a range from 4 to 15 microM. In contrast, Anafp did not show antibacterial activity against Escherichia coli and Bacillus subtilis even at 50 microM. PMID- 10512733 TI - Expression and mutagenesis studies of cobrotoxin from Taiwan cobra. AB - The cDNA encoding cobrotoxin was constructed from the cellular RNA isolated from the venom glands of Naja naja atra (Taiwan cobra). The cDNA was subcloned into the expression vector pET20b(+) and transformed into BL21(DE3) Escherichia coli strain. Expressed cobrotoxin was isolated from inclusion bodies of E. coli and subjected to refolding into its folded structure. The refolded cobrotoxin was purified by high-performance liquid chromatography and exhibited a neurotoxicity in inhibiting acetylcholine-induced muscle contractions. Recombinant cobrotoxin showed a tendency to isomerize its disulfide bonds as that observed with native cobrotoxin. An appreciable decrease in the rate of isomerization reaction was observed when Glu-38 was replaced with Gln-38 or Lys-47 was replaced with Glu-47 or Gln-47. These results reflect that the element in controlling the disulfide isomerization of cobrotoxin is closely associated with the charged side chains in the cobrotoxin molecule. PMID- 10512734 TI - Localization of the linker domain of Ca2+/calmodulin-dependent protein kinase II. AB - Electron micrographs of rotary shadowed replicas of alpha-Ca2+/calmodulin dependent protein kinase II reveal a flower-shaped multimeric molecule with a central particle surrounded by 8-10 smaller peripheral particles. Peripheral particles are attached to the central particle by thin arms or "linkers." Movement of peripheral particles to contact each other for autophosphorylation is likely to involve these linkers. It has generally been accepted that the segment 317-328 of the alpha-subunit constitutes the linker domain. In the present study we test this assumption by generating a mutant lacking the proposed sequence. The mutant has biochemical and morphological properties similar to those of the wild type, and a thin linker is occasionally observed in replicas from either type. The results indicate that the deleted sequence does not correspond to the linker domain. This conclusion, combined with observations from two recent studies which identify the C-terminal domain involved in oligomerization, narrows down the location of the linker domain within the sequence 330-354. PMID- 10512735 TI - Biochemical characterization of a truncated form of CYP27A purified from rabbit liver mitochondria. AB - During purification of CYP27A from rabbit liver mitochondria, a cytochrome P450 of different molecular size was co-isolated. The latter enzyme has an apparent M(r) 51,000 which is slightly lower than that of CYP27A. The 51,000-M(r) protein was found to be present in mitochondria from liver, small intestine, kidney, and spleen but not in lung, testis, heart, or brain mitochondria. Determination of the N-terminal sequence revealed that the 51,000-M(r) protein is a truncated form of CYP27A lacking the first 12 residues. The truncated enzyme was less efficient than the full-length CYP27A in the 27-hydroxylation of C(27)-sterols and much less efficient in the 25-hydroxylation of 1alpha-hydroxyvitamin D(3). The K(m) values for cholesterol and 5beta-cholestane-3alpha,7alpha,12alpha-triol were about the same with both enzymes whereas the K(m) for 1alpha-hydroxyvitamin D(3) was much higher with the truncated CYP27A. The results strongly indicate that the 51,000-M(r) protein is formed via proteolytic processing of CYP27A by endogenous protease(s) in some of the tissues examined. The truncation at the N terminus markedly impairs the ability of CYP27A to use 1alpha-hydroxyvitamin D(3) as substrate and to catalyze 25-hydroxylation in the bioactivation of vitamin D(3). PMID- 10512736 TI - HRC (histidine-rich Ca2+ binding protein) resides in the lumen of sarcoplasmic reticulum as a multimer. AB - HRC (histidine-rich Ca2+ binding protein) has been identified from skeletal and cardiac muscle and shown to bind Ca2+ with low affinity and high capacity that is reminiscent of calsequestrin. The physiological role of HRC is largely unknown. In this study, we show that HRC exists as a multimeric complex (probably larger than a pentamer) under physiological conditions. At higher Ca2+ concentrations, the complex appeared to dissociate into dimers or trimers that form a more relaxed structure. This is in striking contrast to the characteristics of calsequestrin. An earlier immuno-electron microscopic study showed that HRC resides in the lumen of the sarcoplasmic reticulum (SR), but this conclusion has been challenged by other data. By tryptic digestion and biotinylation of SR vesicles, we provide compelling evidence showing that HRC is indeed present in the lumen of the SR. PMID- 10512737 TI - Copper-adenine catalyst for O(2) production from H(2)O(2). AB - In solutions of CuCl2 and adenine copper can be bound to adenine. Two Cu(adenine)(2) complexes [Cu(C(5)H(5)N(5))(2)]2+/Cu(C(5)H(4)N(5))(2)] are in equilibrium with free adenine. Copper-adenine complexes present a catalytic activity (e.g., H(2)O(2) disproportionation into O(2) and water) but depending on complex concentration H(2)O(2) also strongly oxidizes the adenine within the complexes. Raman spectroscopy quantifies copper-adenine complex formation and H(2)O(2) consumption; polarography quantifies O(2) production. As for C(40) catalase, optimal catalytic capacities depend on physiological conditions, such as pH and temperature. The comparative analysis of kinetic parameters shows that the affinity for H(2)O(2) of Cu(adenine)(2) is 37-fold lower than that of C(40) catalase and that the molar activity for O(2) production is 200-fold weaker for Cu(adenine)(2) than for the enzyme. In the 10(-6)-10(-3) M range, the strong decrease of activity with raising complex concentration is explained by aggregation or stacking, which protects Cu(adenine)(2) entities from H(2)O(2) oxidation, but also decreases O(2) production. PMID- 10512738 TI - N-terminal truncated cytochrome P450 2B4: catalytic activities and reduction with alternative electron sources. AB - It was shown that riboflavin binds to the truncated cytochrome P450 2B4 and forms a complex with the K(d) = 26 microM. Noncovalent complex of truncated (Delta2-27) cytochrome P450 2B4 with riboflavin was essential for electron transfer realization and catalyzed the NADH-dependent and hydrogen peroxide-supported monooxygenase reactions of aminopyrine N-demethylation and aniline p hydroxylation. Flavocytochrome molecular maquette was capable of supporting photoactivatable electron transfer and could be photoreduced and electroreduced quantitatively in the absence of pyridine nucleotides. PMID- 10512739 TI - Oxidation of tetrahydrobiopterin by peroxynitrite: implications for vascular endothelial function. AB - Subsaturating levels of tetrahydrobiopterin (BH(4)), an essential cofactor for nitric oxide synthase (NOS), can lead to endothelial dysfunction as a result of decreased production of nitric oxide. Furthermore, insufficient BH(4) can also result in NOS-uncoupled production of reactive oxygen intermediates, such as superoxide anion and hydrogen peroxide. Nitric oxide and superoxide react rapidly to form peroxynitrite, which may be the reactive species responsible for many of the toxic effects of nitric oxide. Here we show that BH(4) is a primary target for peroxynitrite-catalyzed oxidation because at pH 7.4, physiologically relevant concentrations of BH(4) are oxidized rapidly by low concentrations of peroxynitrite. Peroxynitrite oxidizes BH(4) to quinonoid 5,6-dihydrobiopterin and a large proportion of the quinonoid isomer readily loses its side chain to form 7,8-dihydropterin which is not a cofactor for nitric oxide synthase. Thus, abnormally low levels of BH(4) can promote a cycle of its own destruction mediated by nitric oxide synthase-dependent formation of peroxynitrite. This mechanism might contribute to vascular endothelial dysfunction induced by oxidative stress. PMID- 10512740 TI - Pharmacological and signaling analysis of human chemokine receptor CCR-7 stably expressed in HEK-293 cells: high-affinity binding of recombinant ligands MIP 3beta and SLC stimulates multiple signaling cascades. AB - The chemokine receptor CCR-7 is expressed in T, NK, and dendritic cells in a time ordered and stimulus-dependent manner. Thorough analyses of the pharmacological profiles of the recombinant ligands for CCR-7, MIP-3beta/ELC/CK-beta 11, and SLC/Exodus-2/TCA4/6C-kine, using CCR-7-expressing HEK-293E transfectants determine that ligands both bind with a K(d) in the 100 pM range-10- to 100-fold greater affinities than published K(d) values. High-affinity binding of each ligand is associated with rapid mobilization of intracellular calcium and cell migration as predicted for chemokine GPCRs, and in keeping with more recent evidence, robust activation of mitogen-activated protein kinase (MAPK). PMID- 10512741 TI - The role of positively charged residues in CXCR4 recognition probed with synthetic peptides. AB - A high positive charge is the common characteristic shared by the beta-sheet region of stromal cell-derived factor-1 (SDF-1) and CXCR4 antagonists such as ALX40-4C consisting of nine D-arginines. This raises the question that the positively charged residues may play a role in recognition of CXCR4. To test this hypothesis, two studies were carried out using synthetic peptides. In the first study, peptide analogs possessing amino acid sequences from both the N-terminus and the beta-sheet region of SDF-1 were used as models to study the functional role of the beta-sheet region of SDF-1. The attachment of positively charged residues to the N-terminal peptide sequence of SDF-1 was found to enhance the ability of the peptides in CXCR4 binding and inhibiting CXCR4-mediated T-tropic HIV-1 entry. In the second study, two peptides containing nine arginines and the N-terminal signal sequence of SDF-1 were used as models to study the receptor binding mechanism of CXCR4 antagonists of high positive charges such as ALX40-4C. One peptide did not show signaling activity as indicated by the lack of calcium influx while another peptide induced unusual calcium influx distinct from that induced by the SDF-1 N-terminal peptide. In addition, the signal induced by the SDF-1 N-terminal peptide was inhibited by ALX40-4C. Therefore, the first study provides experimental support for the role of the highly positive beta-sheet region of SDF-1 in CXCR4 binding. The second study suggests that the binding site of ALX40-4C in CXCR4 may partially overlap with that of the SDF-1 N-terminal peptide. Both findings should be valuable for the design of SDF-1 agonists and antagonists. PMID- 10512742 TI - Tissue distribution of the human soluble guanylate cyclases. AB - Soluble guanylate cyclase (sGC) is an important component of the NO signaling pathway. Human sGC isoforms alpha(1), alpha(2), and beta(1) show differential mRNA tissue distributions. alpha(1) and beta(1) are expressed in most tissues; however, the alpha(2) isoform shows a more restricted expression pattern with high levels in brain, placenta, spleen, and uterus only. Both alpha subunits exist as multiple transcripts whereas beta(1) exists as a single message. This study reports for the first time the tissue distribution of human sGC message and demonstrates that sGC isoforms are nonuniformly expressed which may be useful if the enzyme is to be exploited as a therapeutic target. PMID- 10512743 TI - IL1HY1: A novel interleukin-1 receptor antagonist gene. AB - Interleukin-1 is a potent mediator of inflammation, involved in regulating a wide variety of physiological and cellular events. We have identified and characterized a novel member of the human interleukin-1 gene family (IL1HY1). The encoded protein demonstrates significant amino acid homology to the receptor antagonist (IL-1ra) at 52%. The gene was mapped to the long arm of chromosome 2, in close proximity to the IL-1 locus. IL1HY1 message is tightly regulated being most predominantly expressed in the skin, but also detected in the spleen, brain leukocyte, and macrophage cell types. Furthermore, the message can be induced in THP-1 cells by phorbol ester (PMA) and lipopolysaccharide (LPS) treatment. PMID- 10512744 TI - Minimal promoter components of the human growth/differentiation factor-5 gene. AB - Growth/differentiation factor-5 (GDF-5) is a new member of the BMP family supposed to be involved in chondrogenesis. We cloned the human GDF-5 gene from lambda phage library and sequenced its 3.5-kb 5'-flanking region. The transcription start site was mapped by 5'-RACE to the sequence that coincides with the initiator element. Electrophoresis mobility shift assays (EMSA) demonstrated a zinc finger transcription factor, YY1, to bind to the sequence surrounding the transcription start site. To localize positive and negative regulatory elements in the GDF-5 5'-upstream region, we constructed a series of progressively deleted promoter-reporter plasmids. The transient transfection assay with human osteoblastic Hos cells indicated that a minimal enhancer element resides within -117 to -27 relative to the transcription initiation site. Since the GDF-5 promoter was active even in fibroblastic L cells, a mechanism governing its chondrocyte-restrictive expression needs to be explored. PMID- 10512745 TI - Transmembrane and water-soluble helix bundles display reverse patterns of surface roughness. AB - Amino acid exposure and surface roughness were calculated for 12 helices from three transmembrane alpha-helix bundles and 13 helices from seven water-soluble alpha-helix bundles. Transmembrane helix bundles have relatively rough surfaces exposed to the lipid bilayer hydrocarbon chains and relatively smooth surfaces along helix-helix interfaces. This pattern is the reverse of what occurs in water soluble helix bundles, where relatively rough surfaces are at the helix-helix interfaces and relatively smooth surfaces are exposed to water. The relatively rough exposed surfaces and buried smooth surfaces of transmembrane helices are likely to contribute to the stability of transmembrane helical bundles in a phospholipid environment. PMID- 10512746 TI - PPARgamma agonists enhance human vascular endothelial adhesiveness by increasing ICAM-1 expression. AB - Early atherosclerotic lesions are characterized by increased monocyte adhesion to the overlying endothelium. Oxidized LDL (oxLDL) stimulates the adhesion of human monocytes to endothelial cells, in part, by increasing expression of ICAM-1. However, the cellular role of oxLDL in endothelial adhesiveness is not well understood. The peroxisome proliferator-activated receptor gamma (PPARgamma), a member of the nuclear receptor superfamily, is expressed in vascular endothelial cells. Whether it can be activated by a synthetic ligand, troglitazone, as well as by natural ligands, oxLDL and its lipid components (i.e., 9- and 13-HODE), has not yet been explored. This study was undertaken to determine whether PPARgamma is expressed in ECV304 human vascular endothelial cells and if so to define the biological effects of its activation by these agonists. Our results demonstrate that PPARgamma mRNA is expressed in ECV304 cells, and transfected cells with a PPARE luciferase construct respond to these agonists. In addition, ligand dependent PPARgamma activation increased ICAM-1 protein expression and enhanced adherence of monocytes to ECV304 cells by two- to threefold. These findings suggest that the PPARgamma signaling pathway might contribute to the atherogenicity of oxLDL in vascular endothelial cells. PMID- 10512747 TI - Molecular cloning and sequence analysis of aggretin, a collagen-like platelet aggregation inducer. AB - A cDNA library derived from the Malayan-pit-viper (Calloselasma rhodostoma) venom gland was constructed in the phagemid vector. Using the information of the N terminal amino acid sequences of two subunits of aggretin, synthetic mixed-base oligonucleotides were employed as a screening probe for colony hybridization. Separate cDNA clones encoding for the alpha and beta chains of aggretin were isolated and sequenced. The results revealed that mature alpha and beta chains contain 136 and 123 amino acid residues, respectively. Aggretin subunits show high degrees of identity with respective subunits (50-60% for alpha, 49-58% for beta) of C-type lectin-like snake venoms. The identity to rattlesnake lectin is relatively lower (i.e., 39 and 30%). All cysteine residues in each chain of aggretin are well conserved and located at the positions corresponding to those of C-type lectins. Thus, three intracatenary disulfide bridges and an interchain disulfide bond between Cys83(alpha) and Cys75(beta) may be allocated. This is the first report regarding the entire sequence of venom GPIa/IIa agonist. According to the alignment of amino acid sequences, hypervariable regions among these C type lectin-like proteins were revealed. These hypervariable regions are proposed to be the counterparts directly interacting with different receptors or different domains of a receptor on the surface of platelet. PMID- 10512748 TI - Genomic organization and structure of the 3' region of human MUC3: alternative splicing predicts membrane-bound and soluble forms of the mucin. AB - The MUC3 gene encodes a large, glycosylated mucin produced by intestinal epithelial cells to form a protective barrier against the external environment. Recently published cDNA sequences for the carboxyl-terminal region of MUC3 polypeptide indicated that rodent Muc3 possesses two epidermal growth factor (EGF)-like domains, and putative transmembrane and cytoplasmic domains, whereas the sequence of human MUC3 predicted termination after the first EGF-like domain. Here we describe the complete genomic sequence encompassing the carboxyl terminal region of human MUC3, revealing the boundaries of 11 exons. RT-PCR and cDNA library cloning experiments indicate that the gene is alternatively spliced, yielding a major membrane-bound form as well as multiple soluble forms. Thus, this work indicates that both membrane-bound and soluble MUC3 mucin proteins are produced by alternative splicing of a single gene. A potentially important polymorphism involving a Tyr residue with a proposed role in signalling is described as well. PMID- 10512749 TI - Novel related cDNAs (C184L, C184M, and C184S) from developing mouse brain encoding two apparently unrelated proteins. AB - Three related cDNAs (C184L, C184M, and C184S) were isolated from a developing mouse brain cDNA library. C184S is the 5'-end portion and C184M is the 3'-end portion, respectively, of C184L. C184S and C184M have open reading frames of 199 amino acids (ORF1) and 189 amino acids (ORF2), respectively; C184L has both ORF1 and ORF2 (dicistronic structure), but seems to translate only ORF1. Southern blot analysis suggests that all of the three related mRNAs are transcribed from the same single gene. The intervening region of C184L cDNA between ORF1 and ORF2 contained a promoter sequence for C184M mRNA, which is transcribed from the corresponding genomic sequence. Very recently, a cDNA encoding human homologue of ORF1 (human autoantigen p27) and a cDNA encoding a different mouse isoform of ORF2 (mammary tumor virus receptor) were reported. Our results indicate that the mRNAs encoding these apparently unrelated proteins are transcribed within an adjacent or overlapping area on the genome, suggesting the same origin of the two transcription units. PMID- 10512750 TI - Protein kinase CKII interacts with and phosphorylates the SAG protein containing ring-H2 finger motif. AB - To investigate the biological function of CKII, we have identified proteins that interact with the subunits of CKII using the yeast two-hybrid system. Here we report that SAG, an antioxidant protein containing Ring-H2 finger motif, is a cellular partner associating with the beta subunit of CKII. SAG does not interact with the alpha subunit of CKII. Analysis of SAG deletion mutants indicates that the Ring-H2 motif of SAG is necessary and sufficient for its binding to the beta subunit of CKII. Recombinant SAG can be phosphorylated by CKII in vitro, providing evidence that the beta subunit mediates the interaction of CKII enzyme with substrate proteins. Overlay experiment shows that SAG and the beta subunit of CKII associate directly in vitro and that CKII-mediated phosphorylation of SAG does not affect the interaction between SAG and the beta subunit of CKII. Northern blot analysis indicates that both SAG and the beta subunit of CKII were relatively rich in human heart, liver, skeletal muscle, and pancreas, but were detected in only trace amounts in brain, placenta, and lung. Our present results suggest that CKII may play a role in the regulation of SAG function. PMID- 10512751 TI - Peroxisome proliferator-activated receptor gamma1 expression in porcine white blood cells: dynamic regulation with acute endotoxemia. AB - Peroxisome proliferator-activated receptor gamma (PPARgamma), a primary regulator of adipocyte differentiation, has been implicated in the regulation of monocyte and macrophage function in vitro. We report that PPARgamma protein is expressed in porcine peripheral white blood cells (WBC), and that PPARgamma1 but not gamma2 mRNA predominates. Additionally, we provide the first evidence that in vivo lipopolysaccharide challenge (LPS, 25 microg/kg BW) causes a dynamic increase in PPARgamma protein expression in peripheral WBC (P < 0.05). PPARgamma expression was increased 2-fold over basal (1 hr post-LPS), was maximal by 4 hr (3-fold), and was normalized to control by 8 hr post-LPS. Changes in PPARgamma expression coincided with or closely followed LPS-induced changes in plasma cortisol, TNF alpha, insulin, IGF-1, glucose, and free fatty acids. These data suggest that induction of PPARgamma expression in WBC may play a role in host response to acute inflammatory challenge and may prove to be an important target of anti inflammatory therapies. PMID- 10512752 TI - Nitrogen-containing bisphosphonates as carbocation transition state analogs for isoprenoid biosynthesis. AB - Nitrogen-containing bisphosphonates are potent bone antiresorptive agents as well as having herbicidal and antiparasitic activity, and are thought to act by inhibiting enzymes of the mevalonate pathway. Using molecular modeling and ab initio quantum chemical calculations, we show that bisphosphonates can act as aza isoprenoid transition state analogs, thereby inhibiting isoprenoid biosynthesis. The two phosphonate groups of the 1,1-bisphosphonates readily dock into the diphosphate-Mg(2+) binding site in farnesyl diphosphate synthase, while the charged ammonium (or pyridinium or imidazolium) groups act as carbocation transition state analogs, whose binding is stabilized by a cluster of oxygen atoms in the active site cleft, and an overall negative electrostatic potential in this region. Enhanced activity is shown to correlate with increasing van der Waals stabilization due to N-alkylation, or the presence of a charged, planar (sp(2)-hybridized) aromatic residue in the carbocation binding site. These results are of general interest since they suggest a rational approach to bisphosphonate drug design. PMID- 10512753 TI - Critical amino acid substitutions in the Src SH3 domain that convert c-Src to be oncogenic. AB - The Src homology 3 (SH3) domain, originally identified in v-Crk, plays an important role in signal transduction. The comparative study with c-src has revealed that v-src oncogene of Schmidt-Ruppin strain of Rous sarcoma virus has three point mutations in its SH3 domain and one in the upstream of SH3. To assess the role of these mutations, each of the single mutations was introduced into c Src by oligonucleotide-directed mutagenesis and its effect on cell transformation was examined. While variant Src proteins that carry each one of single mutations could not transform cells, double mutation at positions 95 and 117 converted c Src to be oncogenic and active in kinase. An additional mutation at position 124 together with one at 95 and 117 further activated Src kinase. By use of GST fusion forms of v-Src SH3 and c-Src SH3, we found that these mutations in SH3 suppressed the binding of SH3 with c-Src protein, possibly with a linker region, while v-SrcSH3 retained the ability to bind a subset of cellular protein to the level similar to those of c-SrcSH3. Taken together, our results suggest that point mutations accumulated in SH3 region can activate, in concert, Src kinase by relaxing the interaction between SH3 and the linker region and subsequently convert Src to be oncogenic. PMID- 10512754 TI - Dissociation of RalA from synaptic membranes by Ca2+/calmodulin. AB - Ras-related small GTP-binding proteins execute many cellular functions, such as cell growth, differentiation, cytoskeletal reorganization, membrane trafficking, and membrane fusion. RalA belongs to the superfamily of Ras-related small GTP binding proteins. Synaptic vesicles (SV) contain small GTP-binding proteins, where RalA, Rab3A, and Rab5A are the major GTP-binding proteins. It has been postulated that a cycling of these proteins between membrane-bound and soluble states is required for regulating cellular functions. Calmodulin (CaM) was found to dissociate Rab3A from SV membranes by forming a 1:1 complex with Ca2+/CaM. RalA was also found to be a Ca2+/CaM-binding protein. Therefore, we examined if Ca2+/CaM can also cause the RalA to dissociate from SV membranes. In this study, we identified that Ca2+/CaM dissociates RalA as well as Rab3A from synaptic vesicles. PMID- 10512755 TI - Induction of human beta-defensin-2 mRNA expression by Helicobacter pylori in human gastric cell line MKN45 cells on cag pathogenicity island. AB - Helicobacter pylori is an etiological agent of gastritis, peptic ulcer, and gastric cancer. Human beta-defensin-2 (hBD-2) is an antimicrobial peptide which belongs to one of the most important host defense systems against bacterial infection in several epithelial tissues. We studied the effect of H. pylori on the expression of hBD-2 mRNA in MKN45 gastric mucosal cells. H. pylori, but not culture filtrate, increased the hBD-2 mRNA level in MKN45 cells; the inductive effect of H. pylori was not detected with Intestine 407 cells. Among H. pylori strains, strain OHPC0002, which lacks a cag Pathogenicity Island (PAI), did not induce hBD-2 mRNA in MKN45 cells. These results suggested that H. pylori cag PAI is critical for the induction of hBD-2 mRNA in MKN45 cells. Exposure of MKN45 cells to Salmonella typhimurium, S. enteritidis, S. typhi, and S. dublin, but not Escherichia coli ML35, also resulted in induction of hBD-2 mRNA. PMID- 10512756 TI - ATP-Binding site of annexin VI characterized by photochemical release of nucleotide and infrared difference spectroscopy. AB - Structural changes induced by nucleotide binding to porcine liver annexin VI (AnxVI) were probed by reaction-induced difference spectroscopy (RIDS). Photorelease of the nucleotide from ATP[Et(PhNO2)] produced RIDS of AnxVI characterized by reproducible changes in the amide I region. The magnitude of the infrared change was comparable to RIDS of other ATP-binding proteins, such as Ca(2+)-ATPase and creatine and arginine kinases. Analysis of RIDS revealed the existence of ATP-binding site(s) (K(d) < 1 microM) within the AnxVI molecule, comprising five to six amino acid residues located in the C-terminal portion of the protein molecule. The binding stoichiometry of ATP:AnxVI was determined as 1:1 (mol/mol). ATP, in the presence of Ca2+, induced changes in protein secondary structure reflected by a 5% decrease in alpha-helix content of the protein in favor of unordered structure. Such changes may influence the affinity of AnxVI for Ca2+ and modulate its interaction with membranes. PMID- 10512757 TI - Formation of PI 3-kinase products in platelets by thrombin, but not collagen, is dependent on synergistic autocrine stimulation, particularly through secreted ADP. AB - Platelet activation by thrombin or collagen results in secretion and synthesis of several platelet agonists that enhance the responses to the primary agonists (autocrine stimulation). To disclose the effects of thrombin and collagen on the phosphorylation of 3-phosphoinositides per se we incubated platelets with five inhibitors of platelet autocrine stimulation (IAS) that act extracellularly. We found that IAS almost totally blocked thrombin-induced production of phosphatidylinositol 3,4-bisphosphate [PtdIns(3,4)P(2)] and phosphatidylinositol 3,4,5-trisphosphate [PtdIns(3,4,5)P(3)]. In contrast, collagen induced massive production of PtdIns(3,4)P(2) and PtdIns(3,4,5)P(3) in the presence of IAS. When testing the effect of each inhibitor individually we found the strongest inhibition of thrombin-induced PtdIns(3,4)P(2) production with the ADP scavenger system CP/CPK. Furthermore, we found a strong synergistic effect between exogenously added ADP and thrombin on production of PtdIns(3,4)P(2). In contrast to the results from 3-phosphorylated phosphoinositides, CP/CPK had little effect on thrombin-induced protein tyrosine phosphorylation. Our results show the importance of autocrine stimulation in thrombin-induced accumulation of 3 phosphorylated phosphoinositides and raise the question as to whether thrombin by itself is capable of inducing PI 3-K activation. In marked contrast to thrombin, collagen per se appears to be able to trigger increased production of PtdIns(3,4)P(2) and PtdIns(3,4,5)P(3). PMID- 10512758 TI - Mitogenic and antiapoptotic effects of insulin-like growth factor binding protein 6 in the human osteoblastic osteosarcoma cell line Saos-2/B-10. AB - Insulin-like growth factor (IGF) I is a potent mitogen for human osteosarcoma cells such as the Saos-2/B-10 cell line. IGF binding proteins (IGFBPs) prevent stimulation of DNA synthesis by IGFs. In contrast to recombinant human (rh) IGFBP 2, -3, -4, and -5, 10-100 nM rhIGFBP-6 stimulated [(3)H]thymidine incorporation into DNA and multiplication of Saos-2/B-10 cells. Upon withdrawal of serum, 30 nM IGFBP-6 also decreased apoptosis (within 4 h) and increased protein content and sodium-dependent phosphate uptake (within 24 h), but less potently than IGF I. (125)I-labeled rhIGFBP-6 did not bind to the cells, and cold IGFBP-6 did not affect (125)I-labeled IGF I binding. Production of IGF I, IGF II, and IGFBP-6 by the cells or significant degradation of rhIGFBP-6 could not be detected within 24 h of incubation. Thus, among the rhIGFBPs tested, rhIGFBP-6 is unique in stimulating osteosarcoma cell growth. Furthermore, it has an antiapoptotic effect. PMID- 10512759 TI - The putative morphogen, DIF-1, of Dictyostelium discoideum activates Akt/PKB in human leukemia K562 cells. AB - The differentiation-inducing factor-1 (DIF-1) is a putative morphogen that induces stalk-cell formation in the lower eukaryote Dictyostelium discoideum. This molecule has been shown to inhibit cell growth and induce erythroid differentiation in human leukemia K562 cells. In the present study, to clarify the mechanism of the actions of DIF-1, we examined the effect of DIF-1 on Akt/protein kinase B (PKB) in K562 cells. Akt/PKB is a serine/threonine kinase that plays a pivotal role in the regulation of cell survival and differentiation in a variety of cells. A nonphosphorylated (inactive) form of Akt/PKB was ordinarily expressed in K562 cells. However, Akt/PKB was phosphorylated and potently activated within several hours of incubation with 5-30 microM DIF-1, and this activation was inhibited by wortmannin, an inhibitor of phosphatidylinositol 3-kinase (PI3-kinase). Calcium-increasing agents thapsigargin and A23187 also activated Akt/PKB slightly, which was inhibited by wortmannin. By contrast, calcium-reducing agents TMB-8 and EGTA together with A23187 inhibited the DIF-1 induced activation of Akt/PKB. PMA (PKC activator) also activated Akt/PKB but this activation was not inhibited by wortmannin. DIF-1 exhibited no marked effect on the activation of PKCalpha, beta, and gamma, which were activated by PMA. These results indicate that DIF-1 activates Akt/PKB possibly via cytosolic calcium and subsequent activation of PI3-kinase and also that PMA activates Akt/PKB in a PI3-kinase-independent manner. PMID- 10512760 TI - The cytoplasmic domain of C-CAM1 tumor suppressor is necessary and sufficient for suppressing the tumorigenicity of prostate cancer cells. AB - We have previously shown that C-CAM1 cell adhesion molecule can suppress the growth of prostate cancer cells in vivo. In this study, we determined the minimal domain of C-CAM1 that is required for its tumor-suppressive activity. DU145 prostate cancer cells were infected with recombinant adenoviruses containing various C-CAM1 mutant genes, and the effects of the mutant C-CAM1 proteins on the growth of DU145 cells were assessed in a nude-mice xenograft model. Deletion of C CAM1's cytoplasmic domain, which is not required for its adhesion activity, abolished the growth-suppressive activity, whereas deletion of the adhesion domain did not. This observation suggests that C-CAM1's extracellular domain may be not essential for its tumor suppressive activity. Indeed, we found that expression of the C-CAM1 cytoplasmic domain alone led to growth suppression of DU145 cells. These results suggest that the cytoplasmic domain of C-CAM1 is necessary and sufficient for its growth-suppressive function. PMID- 10512761 TI - Oxidized low-density lipoprotein induces macrophage respiratory burst via its protein moiety: A novel pathway in atherogenesis? AB - Oxidized low-density lipoproteins (oxLDL) play a crucial role in atherogenesis mainly via their capacity to bind and to activate macrophages. However, the role of the protein LDL moiety in this process is not yet established. In this study, human LDL were exposed to hypochlorous acid (HOCl), a selective protein oxidant, or copper sulfate (CuSO(4)), a major lipid oxidant, and tested for their capacity to activate the NADPH-oxidase of human THP-1- and U937-derived macrophages as measured by lucigenin chemiluminescence (CL). Compared to native LDL which had no effect, HOCl-oxLDL triggered potent CL responses in both U937 and THP-1 cells but only when these were fully differentiated into macrophages by phorbol myristate acetate. In contrast, Cu-oxLDL only triggered a moderate CL response of U937 cells and had little effect on THP-1 cells. While delipidation did not affect HOCl-oxLDL-induced CL response it abolished that induced by Cu-oxLDL. Interestingly, U937 cells showed higher CL responses to both types of oxLDL than THP-1 cells, a finding which could be related to their higher expression of the scavenger receptor CD36. Taken together these results strongly support the role of the protein moiety in oxLDL-induced macrophage activation. PMID- 10512762 TI - Isolation of two novel metalloproteinase-disintegrin (ADAM) cDNAs that show testis-specific gene expression. AB - Metalloproteinase-disintegrins (ADAMs) are type 1 transmembrane proteins that contain a unique domain structure including a zinc-binding metalloproteinase domain. We have isolated cDNAs encoding two novel members of this family, ADAM29 and ADAM30 which show testis-specific expression. Three forms of ADAM29 were found that encode proteins of 820, 786 and 767 amino acids. All of the amino acid differences are located in the cytoplasmic domain. Two forms of ADAM30 were isolated that encode proteins of 790 and 781 amino acids, with the difference in the coding region occurring in the cytoplasmic domain. ADAM29 and ADAM30 map to human chromosome 4q34 and 1p11-13, respectively. An ancestral analysis of all known mammalian ADAMs indicates that the zinc-binding motif in the catalytic domain arose once in a common ancestor and was subsequently lost by those members lacking this motif. PMID- 10512763 TI - Role of Arg182 in the second extracellular loop of angiotensin II receptor AT2 in ligand binding. AB - The phenolic side chain of Tyr(4) present in Ang II is proposed to interact with the side chain of Arg 167 of the AT1 receptor. To determine the contribution of the analogous Arg182 in the ligand-binding properties of the AT2, we replaced the Arg182 with Glu and Ala, and analyzed the ligand-binding properties. Our results suggest that replacing Arg182 with either Glu or Ala abolished the ability of the AT2 receptor to bind the nonspecific peptidic ligands, (125)I-Ang II and [(125)I Sar(1)-Ile(8)]Ang II, as well as the AT2 receptor-specific peptidic ligand (125)I CGP42112A. We have shown previously that replacing the positively charged side chain of Lys215 with the negatively charged side chain of Glu in the fifth TMD did not alter the high affinity binding of (125)I-CGP42112A to the AT2 receptor. However, ligand-binding properties of the Arg182Glu mutant suggest that positively charged side chain of Arg182 located in the junction of second ECL and the fourth TMD is critical for high affinity binding of all three peptidic ligands to the AT2 receptor. PMID- 10512764 TI - Spectral properties of Trp182, Trp194, and Trp250 on the alpha subunit of bacterial luciferase. AB - The phosphorescence and fluorescence properties of bacterial luciferase (alphabeta) mutants from Xenorhabdus luminescens were investigated. All tryptophans in the alpha and beta subunits were replaced with tyrosines except for one or two tryptophans in the alpha subunit. Because one luciferase mutant (W250) retained only a single tryptophan in the alpha subunit while two other mutants (W182/250 and W194/250) each contained two tryptophans in the alpha subunit, it was possible to deduce the spectral properties of these specific tryptophans (Trp182, Trp194, Trp250). Analyses of the phosphorescence properties were particularly revealing as only a single phosphorescence emission peak at 411 414 nm was observed for the W250 and W194/250 mutants while peaks at 409 and 414 nm could be clearly observed for the W182/250 mutant. Coupled with intrinsic fluorescence quenching experiments, these results show that alphaTrp182 is in a distinctly polar environment while alphaTrp250 is in a hydrophobic region and illustrate the advantages of using phosphorescence to recognize different microenvironments for tryptophan residues. PMID- 10512765 TI - Pyridinylimidazole compound SB 203580 inhibits the activity but not the activation of p38 mitogen-activated protein kinase. AB - p38 MAPK is a Ser/Thr protein kinase activated by various inflammatory cytokines and a variety of stress stimuli. It is involved in many physiological processes, including the production of inflammatory cytokines. We have previously reported the design and synthesis of a series of pyridinylimidazole compounds that are selective inhibitors of p38 MAPK. These compounds, exemplified by SB 203580, are exceptionally effective in cell-based assays, including the inhibition of inflammatory cytokine production. SB 203580 is widely used as a tool to dissect the role of p38 MAPK in various physiological processes. It has previously been established that SB 203580 acts primarily to block the catalytic activity of p38 MAPK. However, it has been suggested that in cells, the compounds could also inhibit p38 MAPK activation by virtue of their ability to bind to the inactive enzyme. We undertook careful studies to definitively demonstrate that treatment with SB 203580 had no effect on Thr(180) and Tyr(182) phosphorylation, and hence activation of p38 in vivo. SB 203580, however, potently inhibited the activity of p38 MAPK as demonstrated by the inhibition of the activation of MAPKAP K2, a specific physiological substrate of p38 MAPK. This was observed regardless of stimuli or cell type. Identical results were obtained when the p38 MAPK cascade was partially reconstituted in vitro. Thus, our data clearly indicate that SB 203580 specifically inhibits the activity of p38 MAPK but not its activation by upstream MAPKK. PMID- 10512766 TI - Identification and cloning of odorant binding proteins from the scarab beetle Phyllopertha diversa. AB - Wehave identified, cloned, and characterized two odorant binding proteins from the pale brown chafer, Phyllopertha diversa. One of the proteins (OBP1, 116 amino acids long) showed high amino acid identity (>90%) to two previously identified PBPs from scarab beetles. The second protein (OBP2) showed limited sequence similarity to lepidopteran and dipteran OBPs, but contained only 133 amino acids. Both proteins showed the occurrence of six highly conserved cysteines; electrospray mass spectral data suggested they are all bound in three disulfide bonds. During purification, OBP2 separated into several isoforms; N-terminal amino acid sequencing and electrospray ionization mass spectrometry demonstrated that they are different conformations of the same protein. In the native gel electrophoresis binding experiments, none of the OBPs bound 1, 3-dimethyl-2,4 (1H,3H)-quinazolinedione but different isoforms showed different binding affinities for (R)-japonilure, a pheromone from related scarab beetles, and bombykol, the pheromone from the silkworm moth, Bombyx mori. OBP1 also bound (R) japonilure. PMID- 10512767 TI - Cytochrome P450105D1 (CYP105D1) from Streptomyces griseus: heterologous expression, activity, and activation effects of multiple xenobiotics. AB - The open reading frame of CYP105D1, a soluble cytochrome P450 from Streptomyces griseus, was cloned behind the tac promoter of the bacterial expression vector pSPg1910L and expressed in Escherichia coli. The recombinant protein retained normal spectral characteristics having a Soret peak at 448 nm in the reduced carbon monoxide difference spectrum. CYP105D1 was active, obtaining reducing equivalents from endogenous E. coli ferredoxin and ferredoxin reductase redox partners present in E. coli. In vitro activity studies revealed CYP105D1 to catalyse the NADH- and NADPH-dependent oxidation of the xenobiotic substrates benzo[a]pyrene, erythromycin, warfarin, and testosterone. Furthermore, this activity could be stimulated in the presence of either alpha-benzoflavone or beta benzoflavone in an analogous manner to that reported for mammalian P450 forms including human liver cytochrome P4503A4 (CYP3A4). The system produces an alternative to whole-cell biotransformation of xenobiotic for the production of drug metabolites and an experimental system for probing the structural features of a cytochrome P450 with a broad substrate range. PMID- 10512768 TI - Congenic diabetes-prone BB.Sa and BB.Xs rats differ from their progenitor strain BB/OK in frequency and severity of insulin-dependent diabetes mellitus. AB - Two newly established congenic diabetes-prone BB rat strains designated BB.Sa and BB.Xs carrying a region of chromosome 1 (Sa-Lsn-Secr-Igf2-Tnt, 16 cM) and a region of chromosome X (DXMgh3-Mycs/Pfkb1-Ar, 36 cM) of the SHR rats, respectively, were studied to determine whether the transferred chromosomal regions influence diabetes frequency, age at onset, and clinical picture. Therefore, 4 complete litters of BB/OK (n = 43), BB.Sa (n = 45), and BB.Xs (n = 41) were observed for diabetes occurrence up to the age of 30 weeks. From these litters 6 diabetic males of each strain manifesting in an interval of 1 week were chosen to study body weight, blood glucose, insulin requirement to survive, and several diabetes-related serum constituents at onset of diabetes and after a diabetes duration of 150 days. The diabetes frequency was significantly lower in BB.Xs than in rats of the parental strain BB/OK, whereas comparable frequencies were found between BB/OK and BB.Sa rats. Obvious differences were observed 150 days after diabetes onset between BB/OK and both BB.Sa and BB.Xs rats. BB/OK rats were significantly heavier and needed significantly more insulin/100 g body weight than BB.Sa and BB.Xs rats. Comparisons of the serum constituents as lipids, proteins, and minerals revealed significant differences between diabetic BB/OK rats and their diabetic congenic derivatives in several traits studied at onset and after 150 days of insulin treatment. These results not only show the power of congenic lines in diabetes research, but indicate for the first time that there are genetic factors on chromosomes 1 and X influencing frequency and severity of diabetes in the BB/OK rat. PMID- 10512770 TI - Lennart gram : A personal tribute PMID- 10512769 TI - Characterization of a novel cDNA sequence encoding invertebrate tachykinin related peptides isolated from the echiuroid worm, Urechis unicinctus. AB - Tachykinin is one of the most well-known bioactive peptides found in vertebrates, and tachykinin-related peptides have also been isolated from various invertebrate species. Urechistachykinin I (Leu-Arg-Gln-Ser-Gln-Phe-Val-Gly-Ser-Arg-NH(2)) and II (Ala-Ala-Gly-Met-Gly-Phe-Phe-Gly-Ala-Arg-NH(2)) were purified from the ventral nerve cords of echiuroid worm, Urechis unicinctus. In the present study, we described the characterization of a novel cDNA encoding the urechistachykinin precursor. Amino acid sequence analysis of the deduced polypeptide revealed that the urechistachykinin precursor included seven structurally related peptides, unlike mammalian tachykinin precursors which encode only one or two tachykinin peptides. This is the first identification of an invertebrate tachykinin-related peptide cDNA. PMID- 10512771 TI - Proactive pre-conception counselling for women with epilepsy-is it effective? AB - We describe the development of a proactive pre-conception counselling service for women with epilepsy based on complete re-investigation of the woman's epilepsy, a policy of withdrawing antiepileptic drugs (AEDs) thought to carry an increased risk of foetal abnormality (and substitution, where indicated, of AEDs thought to carry a lesser risk) and the exhibition of folic acid 5 mg daily plus fulfilment of the woman's educational needs and exploration of her and her partner's wishes. The outcome of the assessment of 90 such women is compared with the outcome of 59 women presenting to our service already pregnant. An audit of the outcomes in the two groups suggests that re-investigation of women pre-conceptually is worthwhile (some women turn out not to have epilepsy or have cerebral lesions best managed before pregnancy) and that foetal morbidity may be reduced by judicious rationalization of medication: folic acid taken before conception may also be protective for the foetus. Proactive pre-conception counselling, however, only works if the woman is prepared to wait (sometimes up to a year) for necessary drug changes to be instituted and is using reliable contraception. PMID- 10512772 TI - Long-term results of vagus nerve stimulation in refractory epilepsy. AB - Vagus nerve stimulation (VNS) is an adjunctive antiepileptic treatment for patients with refractory epilepsy. Limited information on long-term treatment with VNS is available. The purpose of this paper is to present our experience with VNS with a follow-up of up to 4 years. Twenty-five patients (13 females and 12 males) with refractory partial epilepsy were treated with VNS. The first 15 patients with a mean age of 30 years and a mean duration of epilepsy of 17.5 years have sufficient follow-up for analysis. Mean post-implantation follow-up was 29 months and mean stimulation output 2.25 mA. There was a mean seizure frequency reduction from 14 complex partial seizures (CPS) per month before implantation to 8 CPS per month after implantation (P = 0.0016; Wilcoxon signed rank rest (WSRT)). The mean maximum CPS-free interval changed from 9 to 312 days (P = 0.0007; WSRT). Six patients were free of CPS for at least one year. In one patient, one antiepileptic drug (AED) was tapered; in 10 patients, AEDs remained unchanged; in four, one adjunctive AED was administered. Side effects occurred in six patients, three of whom required a temporary reduction of output current. Nine patients reported no side effects at all. Treatment with VNS remains effective in the long-term. In this series 4 / 15 (27%) patients with highly refractory epilepsy experienced entirely seizure-free intervals of 12 months or more. PMID- 10512773 TI - Status epilepticus on the paediatric intensive care unit-the role of EEG monitoring. AB - A pilot study was undertaken of the feasibility of continuous EEG monitoring of patients admitted to a Paediatric Intensive Care Unit (PICU) for management of status epilepticus or its immediate sequelae. Eight children were studied and seizure activity was recorded in four patients. Additional information influencing management was obtained: the bedside nurse considered decerebrate posturing in one patient to be a seizure: there were no epileptiform EEG changes. Another patient was considered to have seizures (clonic movements of both upper limbs) following cardiac arrest; the EEG showed electrocerebral silence, and thiopentone treatment was discontinued. In another patient, continuing epileptiform activity on EEG gave intensivists the confidence to use higher than usual doses of thiopentone. The problems encountered were delays in monitoring, once for a CT scan and once because of two admissions within hours of each other. We conclude that EEG monitoring on a PICU is feasible and provides clinically useful information. PMID- 10512774 TI - Bone density and antiepileptic drugs: a case-controlled study. AB - This case-controlled study explored the relationship between bone mineral density (BMD) and long-term treatment with antiepileptic drugs (AEDs) in older adults with epilepsy. Seventy-eight patients (47 post-menopausal females, 31 males, aged 47-76 years) with epilepsy participated in the study. Each had only ever received treatment with either enzyme-inducing (n = 52) or non-inducing (n = 26) AEDs. Individuals were matched for age, sex, height and weight with a drug-naive control. All patients underwent bone densitometry at the lumbar spine and femoral neck and had blood sampling and urine collected for a range of bone markers. Male patients had lower BMD than controls at the lumbar spine (P < 0.01) and neck of the femur (P < 0.005). Female patients had significantly reduced bone density at the femoral neck (P < 0.05) only. AED usage was independently associated with an overall reduction in bone density at femoral sites and contributed to just over 5% of the variance at the femoral neck. Duration of treatment and type of AED were not independent factors for reduction in BMD. This case-controlled study supports the hypothesis that long-term AED therapy is an independent risk factor for reduced BMD in epileptic patients. Adults receiving treatment for epilepsy are at higher risk of osteoporosis and should be offered bone densitometry. PMID- 10512775 TI - A comparison of post-ictal headache between patients with occipital lobe epilepsy and temporal lobe epilepsy. AB - We investigated post-ictal headaches (PIH) using a questionnaire to ascertain their characteristics and compare them among different types of epilepsy. The subjects consisted of 34 patients with occipital lobe epilepsy (OLE) and 75 patients with temporal lobe epilepsy (TLE). PIH occurred in 62% of OLE and 23% of TLE (P < 0.05). The quality of pain in PIH was 'steady' in 71% of OLE and 29% of TLE (P < 0.05) as opposed to 'pounding'. Other factors, such as frequency, severity, duration, and accompanying symptoms showed no significant differences. We found very few patients with migraine-like headaches. Analyses of clinical factors, such as age at onset, duration of epilepsy, seizure frequency, family history of headache, and interictal headache did not reveal any relationship to PIH, although generalized tonic-clonic seizures are associated with PIH in TLE (P < 0.05). These results suggest that the nature of PIH may be different between OLE and TLE, and that the region of epileptic focus or spreading area of epileptic discharge may have a close relation to the induction of PIH. An association with migraine, which has been reported previously, was unclear in our study. PMID- 10512776 TI - SUDEP: overview of definitions and review of incidence data. AB - The classification, occurrence, and predictors of sudden unexpected and unexplained death in individuals with epilepsy (SUDEP) have received considerable attention over the last few years. Specific criteria for the classification of definite, probable, possible, and not SUDEP implemented in United States epidemiologic studies are presented. The incidence of SUDEP in different epilepsy populations is presented. SUDEP is a real phenomenon, because the occurrence of such deaths, especially at relatively young ages, among individuals with epilepsy is far greater (perhaps 40-fold) than among those without epilepsy. SUDEP incidence rates are lower in population-based studies, higher in referral populations and clinical trials of adjunct drugs for complex partial epilepsy, and highest for surgical series. Seizure severity appears to be the strongest risk factor for SUDEP because higher rates are reported from studies of individuals with intractable epilepsy. Other potential risk factors, including sex, seizure etiology, younger age at onset, and partial-onset seizures, are unresolved. PMID- 10512777 TI - Head-up tilting is a useful provocative test for psychogenic non-epileptic seizures. AB - Differentiating psychogenic non-epileptic attack disorder (NEAD) from true epilepsy is difficult. This often results in a misdiagnosis and unnecessary and ineffective treatment. Prolonged EEG/video recording is the most sensitive tool for differentiating NEAD from epilepsy, but is costly and therefore limited in availability. Provocative tests, particularly the use of saline injection, can reduce the length of monitoring but give rise to ethical dilemmas. This study assesses the value of head-up tilt testing as a provocative test for NEAD. Twenty one patients (17 female, mean age 34.6 +/- 11.5 years) with recurrent seizure like episodes and a clinical diagnosis of NEAD were studied. Patients were tilted to 80( composite function )on an electric tilt table with footplate support for up to 45 minutes during continuous ECG, EEG and blood pressure monitoring. Seventeen patients (81%) experienced typical symptoms (non-epileptiform limb shaking in 15 patients, absence in one patient, myoclonic jerking in one patient) during head-up tilt without significant EEG abnormalities or haemodynamic changes. The mean time to onset of seizure-like activity was 13.2 +/- 11 minutes (range 0-31 minutes). No patients suffered injury or any other significant side effect. Provocative testing using suggestion and head-up tilt is a sensitive tool for diagnosing NEAD and represents a safe, simple and inexpensive outpatient technique for investigating patients with suspected NEAD. PMID- 10512778 TI - Home-video observation of seizures in children with epilepsy-impact on quality of family life. PMID- 10512779 TI - Psychological management of intractable seizures in an adolescent with a learning disability. AB - Psychological interventions aimed at seizure management are described with a 14 year-old boy with a learning disability and intractable epilepsy. Baseline records suggested that a majority of tonic seizures and 'drop attacks' were associated with going off to sleep and by environmental 'startles'. Psychological formulation implicated sudden changes in arousal levels as an underlying mechanism of action. Cognitive-behavioural countermeasures were employed to alter arousal levels and processes in different ways in different 'at-risk' situations. A multiple baseline design was used to control for non-specific effects of interventions on non-targeted seizures. Results suggested significant declines in the number of sleep onset and startle-response seizures were attained by these methods. Gains were maintained at 2-month follow-up. PMID- 10512780 TI - Lamotrigine for startle-induced seizures. AB - Startle-induced seizures are reflex seizures precipitated by a sudden, surprising stimulus, usually auditory. Aetiologies, electroencephalographic correlates, and brain structural abnormalities are variable. Because of the frequent tonic component at onset, falling is a major clinical problem. There is no established drug of choice, and therapy is often unsatisfactory. Adjunctive lamotrigine therapy was used in four consecutive patients with this syndrome seen in a referral epilepsy practice. All four had been refractory to virtually every other drug, but responded dramatically to lamotrigine with elimination of falls from seizures. This observation may serve as pilot data for trials of lamotrigine for startle-induced or other varieties of reflex epilepsies, as adjunctive or monotherapy. PMID- 10512781 TI - Ictal catatonia as a manifestation of de novo absence status epilepticus following benzodiazepine withdrawal. AB - To describe ictal catatonia as a manifestation of de novo absence status epilepticus following benzodiazepine withdrawal. Ictal catatonia was documented by concurrent EEG recordings. A catatonic syndrome, first diagnosed as a psychogenic reaction, was found to be an ictal event by EEG recording. De novo absence status and benzodiazepine withdrawal should be considered when a catatonic syndrome suddenly occurs in elderly patients. PMID- 10512782 TI - How much risk does a woman with active epilepsy pose to her newborn child in the puerperium? A pilot study. AB - Much attention in the literature has recently been paid to women's issues in epilepsy but most of the literature stops in the delivery room or at the first moment of suckling. Although it is commonly supposed that a woman who continues to have active epilepsy during the puerperium will pose a risk to her child, little assessment of how great a risk this is has been carried out. We present an audit of the puerperal experiences of 187 women with epilepsy counselled before birth in our women's clinic and contrast this with a number of women with epilepsy seen for the first time in the puerperium (and therefore uncounselled). The audit suggests that in counselled women the risk is very low (women with Juvenile Myoclonic Epilepsy may be particularly at risk, as may women with tonic clonic seizures that occur without warning, plus those with automatisms or who have prolonged post-ictal confusion). Some women with controlled epilepsy prior to conception may lose that control during the puerperium so even women with well controlled epilepsy should adopt precautions in the puerperium. The only baby to die (or be seriously injured) in the puerperium born to a woman with epilepsy was killed in the mother's first seizure. PMID- 10512783 TI - Abstracts of posters presented at 'Epilepsy at the turning point' annual scientific meeting of the british branch of the international league against epilepsy birmingham, april 1999 PMID- 10512784 TI - Prevalence of photoparoxysmal response and the significance of sex steroid hormones in epilepsy. PMID- 10512785 TI - Announcements PMID- 10512786 TI - Molecular 'pharming'. PMID- 10512787 TI - Towards molecular farming in the future: moving from diagnostic protein and antibody production in microbes to plants. AB - Molecular farming of pharmaceuticals in plants has the potential to provide almost unlimited amounts of recombinant proteins for use in disease diagnosis and therapy. Transgenic plants are attracting interest as bioreactors for the inexpensive production of large amounts of safe, functional, recombinant macromolecules, such as blood substitutes, vaccines and antibodies. In some cases, the function of expressed recombinant proteins can be rapidly analysed by expression in microbes or by transient expression in intact or virally infected plants. Protein production can be increased by upscaling production in fermenters, using yeast- or plant-suspension cells or by using transient expression systems. Stable transgenic plants can be used to produce leaves or seeds rich in the recombinant protein for long-term storage or direct processing. This demonstrates the promise for using plants as bioreactors for the molecular farming of recombinant therapeutics, diagnostics, blood substitutes and antibodies. We anticipate that this technology has the potential to greatly benefit human health by making safe recombinant pharmaceuticals widely available. PMID- 10512788 TI - Towards molecular farming in the future: using plant-cell-suspension cultures as bioreactors. AB - Plant-suspension cells are an in vitro system that can be used for recombinant protein production under carefully controlled certified conditions. Plant suspension cells can be grown in shake flasks or fermenters to produce secondary metabolites, like vincristine and vinblastine, and to produce recombinant proteins after transformation. This review article focuses on discussing the generation of transformed suspension-cell lines expressing recombinant proteins, like antibodies, and recombinant-protein downstream processing and purification. PMID- 10512789 TI - Towards molecular farming in the future: transient protein expression in plants. AB - Molecular farming in plants can be achieved by stable or transient expression of a recombinant protein. Transient expression of recombinant proteins in plants can rapidly provide large amounts of the proteins for detailed characterization. It is fast, flexible and can be carried out at field scale using viral vectors, but it lacks the increases in production volume that can be achieved easily with stable transgenic crops. This review article focuses on discussing the applications of transient expression using viral vectors, biolistic methods or agroinfiltration. PMID- 10512790 TI - Towards molecular farming in the future: pichia pastoris-based production of single-chain antibody fragments. AB - This review article focuses on the use of the methylotrophic yeast Pichia pastoris as a recombinant protein-expression system. P. pastoris is a useful system for the expression of milligram-to-gram quantities of a protein, which can be scaled up to fermentation to meet greater demands. Compared with mammalian cells, Pichia do not require a complex growth medium or culture conditions, they are as easy to manipulate genetically as Escherichia coli and have a eukaryotic protein-synthesis pathway. They seem suited to laboratory-scale production of recombinant proteins for in-house use or, in some cases, molecular farming of recombinant products. This review article focuses on the use of P. pastoris, describes a fermentation production run of a single-chain antibody fragment and includes a discussion of fermentation as a production strategy. PMID- 10512791 TI - Performance comparison of protein A affinity-chromatography sorbents for purifying recombinant monoclonal antibodies. AB - We describe the performance characteristics of five Protein A affinity chromatography sorbents (Sepharose Fast Flow, Poros 50, Poros LP, Prosep and Streamline) for purifying a recombinant humanized monoclonal antibody from clarified Chinese hamster ovary cell culture fluid. We measured the dynamic capacity at varying flow rates, maximum capacity, pressure drop and production rate. For purified antibody, we measured yield and purity (by SDS/PAGE, the amount of DNA, the amount of host-cell proteins and the amount of Protein A). We found that, whereas all sorbents provided significant and essentially equivalent antibody purification, there were differences in capacity and pressure drop, which affected the production rate and had implications for process applications. PMID- 10512792 TI - Dermal and transdermal delivery of protein pharmaceuticals: lipid-based delivery systems for interferon alpha. AB - The dermal and transdermal delivery of protein pharmaceuticals faces enormous challenges, and at the same time has very significant potential for the non invasive treatment of both localized and systemic diseases. In this article we review the various approaches used to enhance and control the delivery of protein therapeutic agents through the dermal barrier. We show results of the delivery of interferon (IFN) alpha, an antiviral agent used in the treatment of condylomata acuminata (genital warts), using lipid-based delivery systems (LBDS). In the general category of LBDS, we investigated the use of liposomes and fatty acylation as ways to increase IFNalpha delivery into human skin. PMID- 10512793 TI - Design and synthesis of a morphine-6-succinyl-bovine serum albumin hapten for vaccine development. AB - A morphine-6-succinyl-BSA (M-6-S-BSA) hapten was designed in an effort to obtain a potent, long-lasting anti-morphine immune response for the treatment of morphine abuse. The analogue incorporated a succinic anhydride linker moiety in place of hydroxy group at C-6 of the morphine framework. Then morphine 6 hemisuccinate was conjugated to BSA in aqueous solution in the presence of water soluble carbodi-imide. M-6-S-BSA was synthesized in three chemical steps starting from morphine sulphate, and the extent of conjugation was determined by base hydrolysis of the conjugate, extraction and measurement of free morphine. An average of 6.5 molecules of morphine were conjugated to each molecule of protein. Six male mice, Swiss White strain, immunized with various doses of the conjugate, were found to be producing antibody 8 weeks later, as determined by a modification of the (NH(4))(2)SO(4) method, which measures primary binding of antigen by antibody. PMID- 10512794 TI - Selectivity modification of chymotryptic hydrolysis of haemoglobin by its adsorption on a solid phase. AB - A change of selectivity of the chymotryptic hydrolysis of haemoglobin was evidenced when the protein was adsorbed on to a negatively charged hydrophobic support. The hydrolysis in heterogeneous phase improved the obtaining of positively charged and hydrophobic peptides as carriers of water-insoluble molecules. Haemoglobin adsorption on Amberlyst 15Wet was carried out in 0.1 M Tris/HCl buffer at pH 6.0. Chymotryptic hydrolysis was performed for 72 h at 37 degrees C in the same buffer. In solution, the presence of SDS was necessary to achieve the complete hydrolysis of haemoglobin chains, whereas it was not needed when haemoglobin was previously adsorbed on to the resin. The hydrolysis proceeded more slowly in heterogeneous phase than in homogeneous solution because of the diffusional restrictions but, at the end of the hydrolysis, the peptide populations were very different, as shown by reversed-phase HPLC. Moreover their functional properties were different too, since the haemoglobin hydrolysate obtained by heterogeneous catalysis had a better solubilizing ability towards the water-insoluble molecule, protoporphyrin IX, a photosensitizer for photodynamic therapy. A time-course study of the hydrolysis was performed to follow the evolution of a marker peptide (1-14alpha), which allowed us to explain the change in the selectivity of the chymotryptic reaction. This change could be due to a slowing down of the cut-off of some sites interacting with the support. PMID- 10512795 TI - A novel efficient enzyme-immobilization reaction on NH2 polymers by means of L ascorbic acid. AB - A new enzyme-immobilization reaction by means of L-ascorbic acid (ASA) is described using NH(2) polymers based on cellulose or poly(vinyl alcohol) with the example of oxidoreductase enzymes. In this way, enzyme proteins such as glucose oxidase (GOD), glutamate oxidase, lactate oxidase, urate oxidase and peroxidase can be covalently fixed with a high surface loading to ultrathin and transparent NH(2)-polymer films if their surfaces are previously treated with an ASA solution, in, for example, N,N-dimethyl acetamide, DMSO or methanol. ASA then obviously reacts like a diketo compound with amino groups of the NH(2)-polymer film and enzyme protein, forming dehydroascorbic acid derivatives with neighbouring Schiff's-base structures. In a subsequent fragmentation reaction, the latter presumably form stable oxalic acid diamide derivatives as coupling structures between enzyme protein and NH(2)-polymer film, as suggested by results from investigations of the ASA reaction with n-butylamine. The immobilized enzymes can be stored at 4 degrees C in bidistilled water for at least 1 month without becoming detached from the NH(2)-polymer film and without diminished enzyme activity. The apparent K(m) values of the immobilized enzymes are in part clearly smaller than those of the dissolved enzymes or those found in other immobilization processes such as the diazo coupling or the bifunctional glutardialdehyde reaction. For example, the K(m) value of the immobilized GOD with different NH(2) polymers as the matrix structure is smaller by a factor of approx. 20 than that of the dissolved enzyme. PMID- 10512796 TI - Evaluation of refolding conditions for a recombinant human interleukin-3 variant (daniplestim). AB - The refolding of daniplestim, a human interleukin-3 variant (SC-55494) from Escherichia coli inclusion bodies, was optimized using a reversed-phase HPLC method developed to permit quantification of the reduced and oxidized forms of daniplestim. The presence of cysteine or dithiothreitol accelerated refolding of daniplestim from E. coli inclusion body slurries dissolved in urea or guanidine solutions and was complete in 4-6 h. Regardless of the dissolution and refolding protocol used to renature daniplestim, equivalently bioactive protein was produced. Under refolding conditions, no covalent modification of daniplestim by cysteine or cyanate was observed. The folding process was characterized further by following the unfolding of purified daniplestim by far-UV CD and fluorescence spectroscopies under both oxidizing and reducing conditions at pH values between 7 and 11. Formation of the single disulphide bond had a large stabilizing effect on daniplestim structure ( approximately 4-5 kCal at pH 9.5). This thermodynamic stabilization drove the refolding process towards the native form, even under conditions where the reduced protein was largely unfolded. From these data, scaleable refolding conditions for daniplestim were established. PMID- 10512797 TI - Conformation of engineered proteins. PMID- 10512798 TI - The use of differential scanning calorimetry (DSC) to determine the correctness of folding of cloned proteins. AB - The advantages and limitations of use of the differential scanning calorimetry technique to determine whether cloned and expressed mutant proteins are folded correctly are explained and discussed. Examination of stabilization of these proteins to thermal denaturation by specific ligand binding can provide the required information. PMID- 10512799 TI - Remarkable thermostability of bioelectrodes based on enzymes immobilized within hydrophobic semi-solid matrices. AB - An enhanced resistance to thermal denaturation was investigated for enzymes immobilized within hydrophobic semi-solid matrices compared with both free enzymes and polymer-entrapped enzymes. The bioelectrodes based on the immobilization of glucose oxidase, lactate oxidase, alcohol oxidase, polyphenol oxidase, peroxidase and L-amino acid oxidase within a carbon-paste matrix were constructed to examine their thermal stabilitiy at 60 degrees C or 80 degrees C. The rhodium/glucose oxidase-containing carbon-paste electrode was found to offer a remarkable stability when incubated at 60 degrees C over a long period of 4 months, with only a decrease of approx. 15% in activity. The comparative studies suggest that thermal stabilization established by this enzyme-immobilization procedure varies with the enzyme's inherent stability, the incubation temperature and the immobilizing reagent, such as pasting liquid. PMID- 10512800 TI - Stabilization of the T-state of ferrous human adult haemoglobin by chlorpromazine and trifluoperazine. AB - In the present study, the effect of the neuroleptics chlorpromazine (2-chloro-N,N dimethyl-10H-phenothiazine-10-propanamine) and trifluoperazine {10-[3-(4 methylpiperazin-l-yl)-propyl]-2-trifluoromethyl)-10-H- [phenothiazine}-10H phenothiazine? on the EPR-spectroscopic properties of ferrous human adult nitrosylated haemoglobin (HbNO) is reported. Addition of the two drugs to HbNO shifted the conformational equilibrium from the high- to the low-affinity form of the ligated tetramer, as observed for 2,3-D-glycerate bisphosphate, the physiological modulator of haemoglobin action. The effect of chlorpromazine and trifluoperazine on the EPR-spectroscopic properties of HbNO was enhanced by inositol hexakisphosphate. The binding of neuroleptics to ferrous human adult haemoglobin may represent an important undesirable side effect. In fact, oxygen affinity for ferrous human adult haemoglobin decreases on increasing chlorpromazine and trifluoperazine concentration. In addition, red blood cells may act as neuroleptic scavengers. PMID- 10512801 TI - Steered molecular dynamics simulation of the Rieske subunit motion in the cytochrome bc(1) complex. AB - Crystallographic structures of the mitochondrial ubiquinol/cytochrome c oxidoreductase (cytochrome bc(1) complex) suggest that the mechanism of quinol oxidation by the bc(1) complex involves a substantial movement of the soluble head of the Rieske iron-sulfur protein (ISP) between reaction domains in cytochrome b and cytochrome c(1) subunits. In this paper we report the results of steered molecular dynamics simulations inducing, through an applied torque within 1 ns, a 56 degrees rotation of the soluble domain of ISP. For this purpose, a solvated structure of the bc(1) complex in a phospholipid bilayer (a total of 206,720 atoms) was constructed. A subset of 91,061 atoms was actually simulated with 45,131 moving atoms. Point charge distributions for the force field parametrization of heme groups and the Fe(2)S(2) cluster of the Rieske protein included in the simulated complex were determined. The simulations showed that rotation of the soluble domain of ISP is actually feasible. Several metastable conformations of the ISP during its rotation were identified and the interactions stabilizing the initial, final, and intermediate positions of the soluble head of the ISP domain were characterized. A pathway for proton conduction from the Q(o) site to the solvent via a water channel has been identified. PMID- 10512802 TI - Sodium and chlorine ions as part of the DNA solvation shell. AB - The distribution of sodium and chlorine ions around DNA is presented from two molecular dynamics simulations of the DNA fragment d(C(5)T(5)). (A(5)G(5)) in explicit solvent with 0.8 M additional NaCl salt. One simulation was carried out for 10 ns with the CHARMM force field that keeps the DNA structure close to A DNA, the other for 12 ns with the AMBER force field that preferentially stabilizes B-DNA conformations (, Biophys. J. 75:134-149). From radial distributions of sodium and chlorine ions a primary ion shell is defined. The ion counts and residence times of ions within this shell are compared between conformations and with experiment. Ordered sodium ion sites were found in minor and major grooves around both A and B-DNA conformations. Changes in the surrounding hydration structure are analyzed and implications for the stabilization of A-DNA and B-DNA conformations are discussed. PMID- 10512803 TI - Structure, interaction, dynamics and solvent effects on the DNA-EcoRI complex in aqueous solution from molecular dynamics simulation. AB - A 0.7-ns molecular dynamics simulation of the DNA-EcoRI complex in a 7.0-A solvent shell indicated a stable behavior of the system. No significant evaporation or smearing of the solvent's outer boundary occurred. The structure and the intermolecular interactions were found to be well maintained during the simulation. The interaction pattern in the simulation was found to be very similar to that in the crystal structure. Most of the specific interactions between the DNA and the protein were found to be enhanced in the simulation compared to that in the crystal structure as a result of improved interaction geometry. The nonspecific interactions were found to be stronger than the specific ones. The specific interactions between the N7 atoms of Gua(4) or Ade(5) or Ade(6) and the protein were found to be present over almost the entire time of the simulation, whereas hydrogen bonds involving the amino groups of the Ade(5) and Ade(6) with the protein were found to be relatively weaker, with lower probability and shorter lifetime. The time evolution of the root mean square deviations of the DNA and the protein were highly correlated even at the later part of the simulation, showing the tight binding between them. Several long lived water bridges were found between the DNA backbone atoms and the protein and also between the two protein monomers, which increased the overall stability of the complex. The two protein monomers were found to interact strongly with each other. The energy of the DNA kink deformation was estimated as approximately 31 kcal/mol. PMID- 10512804 TI - Free energy calculations and molecular dynamics simulations of wild-type and variants of the DNA-EcoRI complex. AB - Molecular dynamics simulations and free energy calculations of the wild-type EcoRI-DNA complex and several variants have been performed in aqueous solvent. In general, he theoretical estimations of the free energy differences (DeltaDeltaA) qualitatively agree well with the corresponding experimental data. The modifications which were experimentally found unfavorable compared to the wild type complex were also found to be so in theoretical estimates. The mutant where the amino group of the base Ade(6) was replaced by a hydrogen atom eliminating one H-bond between the DNA and the protein, was experimentally found to be more stable than the wild-type complex. It was speculated that the modification also caused a structural relaxation in the DNA making DeltaDeltaA favorable. Our theoretical estimate yields a positive DeltaDeltaA in this case, but the difference is small, and no significant local structural relaxation was observed. The major H-bonds between the DNA and the protein in the wild-type complex are found to be maintained in the different mutants although the specific and non specific interaction energies between the interacting the DNA bases and the protein residues are different in different mutants. The interaction pattern of the other nearby nucleotides are significantly influenced by each modification. Thus, the alteration of the non-specific interactions may also play an indirect role in determining the specificity of the complex. The interaction of the Gua(4) of the DNA with the protein is found to be most sensitive to any alteration in the recognition site. Because Gua(4) is the nucleotide closest to the scissile bond, this extra sensitivity seems to play an important role in altering the functional activity of the complex. PMID- 10512805 TI - Does Ca2+ reach millimolar concentrations after single photon absorption in Drosophila photoreceptor microvilli? AB - The quantum bump, the elementary event of fly phototransduction induced by the absorption of a single photon, is a small, transient current due to the opening of cation-channels permeable to Ca2+. These channels are located in small, tube like protrusions of the cell membrane, the microvilli. Using a modeling approach, we calculate the changes of free Ca2+ concentration inside the microvilli, taking into account influx and diffusion of Ca2+. Independent of permeability ratios and Ca2+ buffering, we find that the free Ca2+ concentrations rise to millimolar values, as long as we assume that all activated channels are located in a single microvillus. When we assume that as much as 25 microvilli participate in a single bump, the free Ca2+ concentration still reaches values higher than 80 microM. These very high concentrations show that the microvilli of fly photoreceptors are unique structures in which the Ca2+ signaling is even more extreme than in calcium concentration microdomains very close to Ca2+ channels. PMID- 10512806 TI - Driven polymer translocation through a narrow pore. AB - Motivated by experiments in which a polynucleotide is driven through a proteinaceous pore by an electric field, we study the diffusive motion of a polymer threaded through a narrow channel with which it may have strong interactions. We show that there is a range of polymer lengths in which the system is approximately translationally invariant, and we develop a coarse grained description of this regime. From this description, general features of the distribution of times for the polymer to pass through the pore may be deduced. We also introduce a more microscopic model. This model provides a physically reasonable scenario in which, as in experiments, the polymer's speed depends sensitively on its chemical composition, and even on its orientation in the channel. Finally, we point out that the experimental distribution of times for the polymer to pass through the pore is much broader than expected from simple estimates, and speculate on why this might be. PMID- 10512807 TI - Molecular basis for pH sensitivity and proton transfer in green fluorescent protein: protonation and conformational substates from electrostatic calculations. AB - We performed a theoretical study to elucidate the coupling between protonation states and orientation of protein dipoles and buried water molecules in green fluorescent protein, a versatile biosensor for protein targeting. It is shown that the ionization equilibria of the wild-type green fluorescent protein fluorophore and the internal proton-binding site E222 are mutually interdependent. Two acid-base transitions of the fluorophore occur in the presence of neutral (physiologic pH) and ionized (pH > 12) E222, respectively. In the pH-range from approximately 8 to approximately 11 ionized and neutral sites are present in constant ratio, linked by internal proton transfer. The results indicate that modulation of the internal proton sharing by structural fluctuations or chemical variations of aligning residues T203 and S65 cause drastic changes of the neutral/anionic ratio-despite similar physiologic fluorophore pK(a) s. Moreover, we find that dipolar heterogeneities in the internal hydrogen-bond network lead to distributed driving forces for excited state proton transfer. A molecular model for the unrelaxed surrounding after deprotonation is discussed in relation to pathways providing fast ground-state recovery or slow stabilization of the anion. The calculated total free energy for excited-state deprotonation ( approximately 19 k(B)T) and ground-state reprotonation ( approximately 2 k(B)T) is in accordance with absorption and emission data (/= Li(+) > Tl(+) > K(+) > Rb(+) > Cs(+). Current kinetics was little affected by the permeant cation species and concentrations tested (95% of the offspring, a phenomenon known as transmission ratio distortion. The LC2 outer dynein arm light chain of Chlamydomonas reinhardtii is a homologue of the mouse protein Tctex2. We have identified Chlamydomonas insertional mutants with deletions in the gene encoding LC2 and demonstrate that the LC2 gene is the same as the ODA12 gene, the product of which had not been identified previously. Complete deletion of the LC2/ODA12 gene causes loss of all outer arms and a slow jerky swimming phenotype. Transformation of the deletion mutant with the cloned LC2/ODA12 gene restores the outer arms and rescues the motility phenotype. Therefore, LC2 is required for outer arm assembly. The fact that LC2 is an essential subunit of flagellar outer dynein arms allows us to propose a detailed mechanism whereby transmission ratio distortion is explained by the differential binding of mutant (t haplotype encoded) and wild-type dyneins to the axonemal microtubules of t-bearing or wild-type sperm, with resulting differences in their motility. PMID- 10512885 TI - Chronic asthma therapy. PMID- 10512886 TI - New advances in childhood urinary tract infections. PMID- 10512884 TI - The Saccharomyces cerevisiae homologue of human Wiskott-Aldrich syndrome protein Las17p interacts with the Arp2/3 complex. AB - Yeast Las17 protein is homologous to the Wiskott-Aldrich Syndrome protein, which is implicated in severe immunodeficiency. Las17p/Bee1p has been shown to be important for actin patch assembly and actin polymerization. Here we show that Las17p interacts with the Arp2/3 complex. LAS17 is an allele-specific multicopy suppressor of ARP2 and ARP3 mutations; overexpression restores both actin patch organization and endocytosis defects in ARP2 temperature-sensitive (ts) cells. Six of seven ARP2 ts mutants and at least one ARP3 ts mutant are synthetically lethal with las17Delta ts confirming functional interaction with the Arp2/3 complex. Further characterization of las17Delta cells showed that receptor mediated internalization of alpha factor by the Ste2 receptor is severely defective. The polarity of normal bipolar bud site selection is lost. Las17-gfp remains localized in cortical patches in vivo independently of polymerized actin and is required for the polarized localization of Arp2/3 as well as actin. Coimmunoprecipitation of Arp2p with Las17p indicates that Las17p interacts directly with the complex. Two hybrid results also suggest that Las17p interacts with actin, verprolin, Rvs167p and several other proteins including Src homology 3 (SH3) domain proteins, suggesting that Las17p may integrate signals from different regulatory cascades destined for the Arp2/3p complex and the actin cytoskeleton. PMID- 10512887 TI - A synopsis of the American Academy of Pediatrics' practice parameter on the diagnosis, treatment, and evaluation of the initial urinary tract infection in febrile infants and young children. PMID- 10512888 TI - A modern pediatric fable: two for the price of one. PMID- 10512890 TI - Earning CME credit-completing the PIR quiz PMID- 10512889 TI - Consultation with the specialist: nonrespiratory cyanosis. PMID- 10512891 TI - Index of suspicion. Case 1. Osteoporosis congenita. PMID- 10512892 TI - Chloramphenicol. PMID- 10512893 TI - Iron-fortified infant formulas. PMID- 10512894 TI - Congenital diaphragmatic hernia: the perinatalogist's perspective. PMID- 10512895 TI - Congenital diaphragmatic hernia: the surgeon's perspective. PMID- 10512896 TI - Congenital diaphragmatic hernia: the neonatologist's perspective. PMID- 10512897 TI - Reports of randomized trials in acute stroke, 1955 to 1995. What proportions were commercially sponsored? AB - BACKGROUND AND PURPOSE: Research in acute stroke has expanded rapidly. Many potentially important interventions lack commercial potential (eg, admission to a stroke unit). We therefore wished to examine the frequency of reports of randomized trials of interventions for acute stroke over the past 40 years, the source of support for such trials, the reporting of the commercial involvement, and whether the proportion of commercially supported trials had changed. METHODS: Eligible trials were identified from the Cochrane Stroke Group's specialized register of controlled clinical trials. We included all randomized trials in patients with acute stroke which published a full text report, in English, between 1955 and 1995. Two reviewers independently extracted data on the involvement of the pharmaceutical industry in all eligible trials. RESULTS: There was a substantial increase in the number of acute stroke trials published per year between 1955 and 1995. The description of pharmaceutical industry involvement in each trial report was poor. Only a minority of supported trials made explicit statements about the role of the sponsoring company. The proportion of trials apparently supported by the pharmaceutical industry has increased substantially. CONCLUSIONS: The increasingly important role of the pharmaceutical industry in evaluating new treatments gives substantial scope for bias and may not be in the interests of public health. Poor reporting of the sponsor's involvement suggests that modifications to the guidelines for the reporting of randomized controlled trials to include more details of the sponsor's involvement in the design, conduct, management, analysis, and reporting of the trial are justified. PMID- 10512898 TI - Risk factors and 20-year stroke mortality in men and women in the Renfrew/Paisley study in Scotland. AB - BACKGROUND AND PURPOSE: The aim of this study was to relate risk factors in middle-aged men and women to stroke mortality over a long follow-up period. METHODS: In the early to mid 1970s, 7052 men and 8354 women from the Renfrew/Paisley prospective cohort study in Scotland were screened when aged 45 to 64 years. Risk factors measured included blood pressure, blood cholesterol and glucose, respiratory function, cardiothoracic ratio, smoking habit, height, body mass index, age, preexisting coronary heart disease, and diabetes. These were related to stroke mortality over 20 years of follow-up. RESULTS: Women's stroke mortality rates were similar to men's, unlike coronary heart disease mortality, in which case women's rates were lower than men's. Diastolic and systolic blood pressure, smoking, cardiothoracic ratio, preexisting coronary heart disease, and diabetes were positively related to stroke mortality for men and women, while adjusted forced expiratory volume in 1 second and height were negatively related. Cholesterol and body mass index were not related to stroke mortality. Glucose in nondiabetics was positively related to stroke mortality for women but not men, and there was evidence of a threshold effect at the highest levels of glucose. Former smokers had mortality rates that were similar to those of never-smokers. In sex-specific multivariate models, most variables retained a statistically significant association with stroke mortality, illustrating the multifactorial etiology of stroke. CONCLUSIONS: Overall, findings for women were similar to those for men. Control of risk factors for reduction of stroke mortality should be targeted at men and women in a similar fashion, particularly with reference to smoking cessation and blood pressure control. PMID- 10512899 TI - What determines good recovery in patients with the most severe strokes? The Copenhagen Stroke Study. AB - BACKGROUND AND PURPOSE: Even patients with the most severe strokes sometimes experience a remarkably good recovery. We evaluated possible predictors of a good outcome to search for new therapeutic strategies. METHODS: We included the 223 patients (19%) with the most severe strokes (Scandinavian Stroke Scale score <15 points) from the 1197 unselected patients in the Copenhagen Stroke Study. Of these, 139 (62%) died in the hospital and were excluded. The 26 survivors (31%) with a good functional outcome (Barthel Index >/=50 points) were compared with the 58 survivors (69%) with a poor functional outcome (Barthel Index <50 points). The predictive value of the following factors was examined in a multivariate logistic regression model: age; sex; a spouse; work; home care before stroke; initial stroke severity; blood pressure, blood glucose, and body temperature on admission; stroke subtype; neurological impairment 1 week after onset; diabetes; hypertension; atrial fibrillation; ischemic heart disease; previous stroke; and other disabling disease. RESULTS: Decreasing age (odds ratio [OR], 0.50 per 10 year decrease; 95% CI, 0.25 to 0.99; P=0.04), a spouse (OR, 3.1; 95% CI, 1.1 to 8. 8; P=0.03), decreasing body temperature on admission (OR, 1.8 per 1 degrees C decrease; 95% CI, 1.1 to 3.1; P=0.01), and neurological recovery after 1 week (OR, 3.2 per 10-point increase in Scandinavian Stroke Scale score; 95% CI, 1.1 to 7.8; P=0.01) were all independent predictors of good functional outcome. CONCLUSIONS: Patients with the most severe strokes who achieve a good functional outcome are generally characterized by younger age, the presence of a spouse at home, and early neurological recovery. Body temperature was a strong predictor of good functional outcome and the only potentially modifiable factor. We suggest that a randomized controlled trial be undertaken to evaluate whether active reduction of body temperature can improve the generally poor prognosis of patients with the most severe strokes. PMID- 10512900 TI - Chlamydia pneumoniae antibodies and high lipoprotein(a) levels do not predict ischemic cerebral infarctions. Results from a nested case-control study in Northern Sweden. AB - BACKGROUND AND PURPOSE: An association between high lipoprotein(a) [Lp(a)] levels and positive Chlamydia pneumoniae IgG titers in coronary artery disease has been described. The possibility of predicting ischemic stroke by measurements of plasma Lp(a) and C pneumoniae antibodies was investigated. METHODS: This incident case-control study included 101 case subjects (cases) who had suffered ischemic cerebral infarctions and 201 matched control subjects (controls). The study population was nested within the Vasterbotten Intervention Program or the WHO MONICA project. Plasma samples were measured for C pneumoniae-specific IgG and IgA antibodies and Lp(a). RESULTS: A significantly higher mean Lp(a) level was found in female cases than in female controls. However, plasma Lp(a) was unable to predict ischemic cerebral infarctions in either women or men. The proportion of individuals with positive C pneumoniae-specific IgG or IgA titers did not differ between cases and controls. Antibody titers were unable to predict a future stroke. The proportion of individuals with a positive C pneumoniae IgG titer in combination with a high Lp(a) level did not differ significantly between cases and controls. CONCLUSIONS: These data suggest that there is no association between baseline plasma Lp(a) levels, presence of C pneumoniae antibodies, and future ischemic cerebral infarctions. Furthermore, no evidence of an interactive effect between high Lp(a) levels and C pneumoniae IgG titers was found. However, selection bias and a recent C pneumoniae epidemic may have influenced the results. PMID- 10512901 TI - Left atrial size and the risk of ischemic stroke in an ethnically mixed population. AB - BACKGROUND AND PURPOSE: The association between left atrial size and ischemic stroke is controversial and has been suggested to exist only in men and to be mediated by left ventricular mass. Data are available almost exclusively for white patients. The purpose of this study was to evaluate the association between left atrial size and ischemic stroke in a multiethnic population. METHODS: A population-based case-control study was conducted in 352 patients aged >39 years with first ischemic stroke and in 369 age-, gender-, and race-ethnicity-matched community controls. Left atrial diameter was measured by 2-dimensional transthoracic echocardiography and indexed by body surface area. Conditional logistic regression analysis was performed to assess the risk of stroke associated with left atrial index in the overall group and in the age, gender, and race-ethnic strata after adjustment for the presence of other stroke risk factors. RESULTS: Left atrial index was associated with ischemic stroke in the overall group (adjusted OR 1.47 per 10 mm/1.7 m(2) of body surface area; 95% CI 1.03 to 2.11). The association was present in men (adjusted OR 2.81, 95% CI 1.42 to 5.57) but not in women (adjusted OR 1.08, 95% CI 0.70 to 1.66), and in patients aged <60 years (adjusted OR 3.78, 95% CI 1.36 to 10.54) but not >60 years (adjusted OR 1.23, 95% CI 0.84 to 1.81). Subgroup analyses showed the risk to be present in men across all age subgroups. In women, the lack of association between left atrial index and stroke was most strongly influenced by left ventricular hypertrophy. A trend toward an association between left atrial index and stroke was observed in whites (adjusted OR 1.81, 95% CI 0.81 to 4.09) and Hispanics (adjusted OR 1.61, 95% CI 0.98 to 2.65) but was less evident in blacks (adjusted OR 1.25, 95% CI 0.74 to 2.14). CONCLUSIONS: Left atrial enlargement is associated with an increased risk of ischemic stroke after adjustment for other stroke risk factors, including left ventricular hypertrophy. The association is observed in men of all ages, whereas in women it is attenuated by other factors, especially left ventricular hypertrophy. Interracial differences in the stroke risk may exist that need further investigation. PMID- 10512902 TI - Extravasation of radiographic contrast is an independent predictor of death in primary intracerebral hemorrhage. AB - BACKGROUND AND PURPOSE: Hematomas that enlarge following presentation with primary intracerebral hemorrhage (ICH) are associated with increased mortality, but the mechanisms of hematoma enlargement are poorly understood. We interpreted the presence of contrast extravasation into the hematoma after CT angiography (CTA) as evidence of ongoing hemorrhage and sought to identify the clinical significance of contrast extravasation as well as factors associated with the risk of extravasation. METHODS: We reviewed the clinical records and radiographic studies of all patients with intracranial hemorrhage undergoing CTA from 1994 to 1997. Only patients with primary ICH were included in this study. Univariate and multivariate logistic regression analyses were performed to determine the associations between clinical and radiological variables and the risk of hospital death or contrast extravasation. RESULTS: Data were available for 113 patients. Contrast extravasation was seen in 46% of patients at the time of CTA, and the presence of contrast extravasation was associated with increased fatality: 63.5% versus 16.4% in patients without extravasation (P=0.011). There was a trend toward a shorter time (median+/-SD) from symptom onset to CTA in patients with extravasation (4.6+/-19 hours) than in patients with no evidence of extravasation (6.6+/-28 hours; P=0.065). Multivariate analysis revealed that hematoma size (P=0.022), Glasgow Coma Scale (GCS) score (P=0.016), extravasation of contrast (P=0.006), infratentorial ICH (P=0.014), and lack of surgery (P<0.001) were independently associated with hospital death. Variables independently associated with contrast extravasation were hematoma size (P=0.024), MABP >120 mm Hg (P=0.012), and GCS score of /=40, 82% in the oxygen group and 91% in the control group survived (OR 0. 45; 95% CI 0.23 to 0.90; P=0.023). For patients with SSS scores of <40, 53% in the oxygen group and 48% in the control group survived (OR 1.26; 95% CI 0.76 to 2.09; P=0.54). CONCLUSIONS: Supplemental oxygen should not routinely be given to nonhypoxic stroke victims with minor or moderate strokes. Further research is needed to give conclusive advice concerning oxygen supplementation for patients with severe strokes. PMID- 10512904 TI - Safety and tolerability study of aptiganel hydrochloride in patients with an acute ischemic stroke. AB - BACKGROUND AND PURPOSE: Aptiganel (CNS 1102) is a selective, noncompetitive antagonist that acts on the ion channel associated with the N-methyl-D-aspartate (NMDA) receptor and is neuroprotective in experimental focal cerebral ischemia models at a plasma concentration of 10 ng/mL. In human volunteers, dose-limiting effects of aptiganel are blood pressure increases and central nervous system (CNS) excitation or depression. This study assessed the safety and tolerability of non-weight-adjusted doses of aptiganel in patients with acute ischemic stroke. METHODS: This was a double-blind, randomized, placebo-controlled multicenter study in patients presenting within 24 hours of acute ischemic stroke. Ascending single intravenous bolus doses of aptiganel (3, 4.5, 6, and 7.5 mg) were assessed in 21 patients with a 3:1 active drug:placebo randomization schedule. In 15 subsequent patients, selected bolus doses were followed by constant intravenous infusion for 6 to 12 hours (6 mg plus 1 mg/h, n=10; then 4.5 mg plus 0.75 mg/h, n=15) in a 4:1 randomization schedule. Prospectively collected pharmacokinetic data guided selection of infusion rates. Neurological and functional status were recorded at entry and after 1 week, although the study was not designed to test efficacy. RESULTS: Forty-six patients were randomized from 4 centers (3 in the United States and 1 in the United Kingdom): 36 received aptiganel HCl, and 10 were given placebo. Hypertension and CNS events were commonly reported after a bolus dose of 7.5 mg and after a 6-mg bolus followed by an infusion of 1 mg/h. The lower regimen of 4.5-mg bolus followed by infusion of 0.75 mg/h achieved plasma aptiganel concentrations of >10 ng/mL and was well tolerated by patients but still raised systolic blood pressure by approximately 30 mm Hg over baseline. CONCLUSIONS: A 4.5-mg intravenous bolus of aptiganel HCl followed by infusion of 0.75 mg/h for 12 hours is a tolerable dose that can produce plasma drug concentrations shown to be neuroprotective in animal models. However, increases in systolic blood pressure and an excess of CNS effects were both observed at this dose. PMID- 10512905 TI - Pathophysiological topography of acute ischemia by combined diffusion-weighted and perfusion MRI. AB - BACKGROUND AND PURPOSE: Combined echoplanar MRI diffusion-weighted imaging (DWI), perfusion imaging (PI), and magnetic resonance angiography (MRA) can be used to visualize acute brain ischemia and predict lesion evolution and functional outcome. The appearance of a larger lesion by PI than by DWI quantitatively defines a mismatch of potential clinical importance. Qualitative lesion variations exist in the topographic concordance of this mismatch. We examined both the topographic heterogeneity and relative frequency of mismatched patterns in acute stroke using these MRI techniques. METHODS: Acute DWI, PI, and MRA studies of 34 prospectively recruited patients with supratentorial ischemic lesions scanned within 24 hours of stroke onset (range 2.5 to 23.3 hours, 12 patients <6 hours) were analyzed. RESULTS: Ischemic lesions were predominantly in the middle cerebral artery (MCA) territory (94%), with DWI lesions most commonly affecting the insular region. Mismatched patterns with PI lesion larger than DWI lesion occurred in 21 patients (62% overall), in all 4 patients imaged within 3 hours, and in 44% of patients imaged after 18 hours. A patient with a large PI but no DWI lesion and severe clinical deficit at 2.5 hours after stroke onset recovered completely. Regional variations in DWI and PI lesion loci were found, inferring site of proximal MCA occlusion, embolic pathogenesis, and regional arterial reperfusion. CONCLUSIONS: Analysis of the topographic concordance of PI and DWI lesions in acute stroke reveals regional PI lesions without concomitant DWI lesions, which do not necessarily progress to infarction but may suggest stroke pathogenesis and site of current arterial occlusion. Location of DWI lesions may suggest an earlier site of arterial occlusion and regions of maximal perfusion deficit. PMID- 10512906 TI - Magnetic resonance imaging white matter hyperintensities and mechanism of ischemic stroke. AB - BACKGROUND AND PURPOSE: We sought to determine the relations between infarct subtype and white matter hyperintensities (WMHIs) on MRI. MATERIALS AND METHODS: We studied 395 ischemic stroke patients with 1. 0-T MRI. The number of lacunar, border-zone, and cortical infarcts was registered. WMHIs were analyzed in 6 areas. Univariate and multivariate statistical analyses were used to find the risk factors for different infarct subtypes and to study the connections between WMHIs and brain infarcts. RESULTS: Lacunar infarcts were associated with hypertension (odds ratio [OR], 1.79; 95% CI, 1.17 to 2.73), alcohol consumption (OR, 1.96; 95% CI, 1.17 to 3.28), and age (OR, 1. 03; 95% CI, 1.00 to 1.06). Border-zone infarcts were associated with carotid atherosclerosis (OR, 2.20; 95% CI, 1.15 to 4.19). Atrial fibrillation (OR, 3.02; 95% CI, 1.66 to 5.50) and carotid atherosclerosis (OR, 1.94; 95% CI, 1.12 to 3.36) were independent positive predictors, and history of hyperlipidemia (OR, 0.44; 95% CI, 0.26 to 0.75) and migraine (OR, 0.48; 95% CI, 0.25 to 0.93) were negative predictors for cortical infarcts. Patients with lacunar infarcts had more severe WMHIs than patients with nonlacunar infarcts in all WM areas (P33% of the middle cerebral artery territory (European Cooperative Acute Stroke Study [ECASS] criteria). Diffusion weighted imaging (DWI) is more sensitive than CT in detecting acute ischemia, and the combination of DWI, MR perfusion imaging, and MR angiography provides additional information from a single examination. We sought to determine whether DWI could identify the presence and extent of major ischemia as well as CT in hyperacute stroke patients. METHODS: Seventeen suspected hemispheric stroke patients were studied with both CT and DWI within 6 hours of symptom onset. None received thrombolytic therapy. The scans were examined separately by 2 neuroradiologists in a blinded fashion for ischemic change and cerebral edema, graded as normal, <33%, or >33% of the MCA territory. Final diagnosis of stroke was determined with the use of standard clinical criteria and T2-weighted imaging at day 90. RESULTS: Sixteen of 17 patients had a final diagnosis of stroke. Acute ischemic changes were seen in all 16 on DWI (100% sensitivity) and in 12 of 16 on CT (75% sensitivity). DWI identified all 6 patients with major ischemia on CT, with excellent agreement between the 2 imaging techniques (kappa=0.88). One patient eligible for thrombolysis on the ECASS CT criteria had major ischemia on DWI. CONCLUSIONS: DWI is more sensitive than CT in the identification of acute ischemia and can visualize major ischemia more easily than CT. PMID- 10512908 TI - Diffusion-weighted MRI in acute lacunar syndromes. A clinical-radiological correlation study. AB - BACKGROUND AND PURPOSE: Clinical-radiological correlation studies in lacunar syndromes have been handicapped by the low sensitivity of CT and standard MRI for acute small-vessel infarction and their difficulty in differentiating between acute and chronic lesions. METHODS: We prospectively studied 43 patients presenting with a classic lacunar syndrome using diffusion-weighted MRI, a technique with a high sensitivity and specificity for acute small-vessel infarction. RESULTS: All patients were scanned within 6 days of stroke onset. An acute infarction was identified in all patients. Pure motor stroke was associated with lesions in the posterior limb of the internal capsule (PLIC), pons, corona radiata, and medial medulla; ataxic hemipareses with lesions in the PLIC, corona radiata, pons, and insular cortex; sensorimotor stroke with lesions in the PLIC and lateral medulla; dysarthria-clumsy hand syndrome with lesions in the PLIC and caudate nucleus; and pure sensory stroke with a lesion in the thalamus. Supratentorial lesions extended into neighboring anatomic structures in 48% of the patients. CONCLUSIONS: Lacunar syndromes can be caused by lesions in a variety of locations, and specific locations can cause a variety of lacunar syndromes. Extension of lesions into neighboring structures in patients with lacunar syndromes appears to be more frequent than previously described in studies using CT and standard MRI. PMID- 10512909 TI - Lack of evidence of acute ischemic tissue change in transient global amnesia on single-shot echo-planar diffusion-weighted MRI. AB - BACKGROUND AND PURPOSE: There is uncertainty concerning the etiology of transient global amnesia (TGA). Previous CT and MRI studies have indicated that permanent structural abnormality is rare in TGA. Diffusion-weighted (DW) MRI is very sensitive to early ischemic parenchymal changes and has recently demonstrated embolic infarction in the posterior cerebral artery territory in 2 TGA patients. We report the findings of DW MRI in 8 patients in acute stages of TGA. METHODS: Conventional and echo-planar DW MRI was performed in 2 patients in the active phase and 6 patients in the recovery phase (1 to 8 hours after cessation of anterograde memory dysfunction) of spontaneously occurring TGA. RESULTS: None of the patients showed signs of hyperintensity on DW images or hypointensity on quantitative apparent diffusion coefficient (ADC) maps to suggest regional decreases of water mobility or acute T2 changes on transverse or coronal slices. CONCLUSIONS: We were unable to detect ADC or acute T2 changes with echo-planar DW MRI in patients with TGA, which suggests that mechanisms other than ischemic infarction may cause TGA. We did not identify spreading depression-associated changes of the ADC. Further refinement of MRI sequences may be necessary to detect subtle or transient signal change in brain parenchyma. PMID- 10512910 TI - Treatment of posterior circulation ischemia with extracranial percutaneous balloon angioplasty and stent placement. AB - BACKGROUND AND PURPOSE: Vertebrobasilar territory ischemia (VBI) leads to disabling neurological symptoms and poses a risk for stroke by an embolic or flow related mechanism. We present our clinical experience in the endovascular treatment of patients with symptomatic VBI from severe atherosclerosis or dissection of the vertebral and subclavian arteries that was unresponsive to medical therapy. METHODS: Twenty-one patients (9 female, 12 male) with a mean age of 65.7 years (range 47 to 81 years) underwent treatment with percutaneous endovascular balloon angioplasty and stent placement. Sixteen patients (76.2%) had evidence of contralateral involvement, and 9 (42.8%) demonstrated severe anterior-circulation atherosclerosis. Nine patients had a previous infarct in the occipital lobe, cerebellum, or pons before treatment. Follow-up was available for all patients. RESULTS: Balloon angioplasty with intravascular stent placement was performed in 13 vertebral artery lesions (10 at the origin, 3 in the cervical segment) and in 8 subclavian lesions. The prestenting stenosis was 75% (50% to 100%) and was reduced to 4.5% (0% to 20%) after stenting. Six of the patients with proximal subclavian stenosis demonstrated angiographic evidence of subclavian steal, which resolved in all cases after treatment. All patients showed improvement in symptoms after the procedure except for 1 who developed a hemispheric stroke after thrombotic occlusion of an untreated cavernous carotid artery stenosis (rate of major stroke and mortality=4.8%). One patient (4.8%) had a periprocedural transient ischemic attack (TIA), and none had minor stroke. At long-term follow-up (mean=20.7+/-3.6 months) of the surviving 20 patients, 12 (57.1%) remained symptom-free, 4 (19%) had at most 1 TIA over a 3-month period, 2 (9.5%) had at most 1 TIA per month, and 2 (9.5%) had persistent symptoms. There were no clinically evident infarcts during the follow-up period. CONCLUSIONS: Endovascular treatment using balloon angioplasty with intravascular stent placement for symptomatic stenotic lesions resulting in VBI that is unresponsive to medical therapy appears to be of benefit in this high-risk subset of patients with poor collateral flow. PMID- 10512911 TI - Frequency and determinants of postprocedural hemodynamic instability after carotid angioplasty and stenting. AB - BACKGROUND AND PURPOSE: Hemodynamic instability can occur acutely after carotid angioplasty and stent placement (CAS). We performed this study to determine the frequency of hemodynamic instability in a series of patients who underwent CAS and to analyze factors associated with development of postprocedural hemodynamic events. METHODS: We reviewed medical records and angiograms in a series of 51 patients (mean age 68.3+/-8.9 years) who underwent CAS for symptomatic (n=29) or asymptomatic (n=22) carotid artery stenosis. Any episodes of hypotension (systolic blood pressure <90 mm Hg), hypertension (systolic blood pressure >160 mm Hg), or bradycardia (heart rate <60 bpm) that occurred in the acute postprocedural period were recorded. The effect of demographic, clinical, intraprocedural, and angiographic factors on subsequent development of hemodynamic instability was analyzed by logistic regression. RESULTS: The frequency of postprocedural hemodynamic complications in our patient series was as follows: hypotension, 22.4%; hypertension, 38.8%; and bradycardia, 27.5%. Intraprocedural hypotension (odds ratio [OR] 14.6, P=0.024) and history of myocardial infarction (OR 14.1, P=0.04) independently predicted postprocedural hypotension. Postprocedural hypertension was predicted by intraprocedural hypertension (OR 7.6, P=0.01) and previous ipsilateral carotid endarterectomy (OR 7.6, P=0.02). Postprocedural bradycardia was associated with intraprocedural hypotension (OR 74, P=0.001) and intraprocedural bradycardia (OR 12, P=0.008). All events had resolved at the conclusion of the intensive care unit monitoring period (mean 25.7 hours, range 18 to 43 hours). CONCLUSIONS: Postprocedural hemodynamic instability is frequent after CAS and supports the need for monitoring in settings suited to expeditious management of cardiovascular emergencies. Patients who have evidence of hemodynamic instability during the procedure are at highest risk. PMID- 10512912 TI - Predictors of clinical improvement, angiographic recanalization, and intracranial hemorrhage after intra-arterial thrombolysis for acute ischemic stroke. AB - BACKGROUND AND PURPOSE: We sought to evaluate predictors of clinical outcome, angiographic success, and adverse effects after intra-arterial administration of urokinase for acute ischemic stroke. METHODS: We designed a Brain Attack program at University Hospitals of Cleveland for diagnosis and treatment of patients presenting within 6 hours of onset of neurological deficit. Patients with ischemia referable to the carotid circulation were treated with intra-arterial urokinase. Angiographic recanalization was assessed at the end of medication infusion. Intracerebral hemorrhage was investigated immediately after and 24 hours after treatment. Stroke severity was determined, followed by long-term outcome. RESULTS: Fifty-four patients were treated. There was improvement of >/=4 points on the National Institutes of Health Stroke Scale from presentation to 24 hours after onset in 43% of the treated patients, and this was related to the severity of the initial deficit. Forty-eight percent of patients had a Barthel Index score of 95 to 100 at 90 days, and total mortality was 24%. Cranial CT scans revealed intracerebral hemorrhage in 17% of patients in the first 24 hours, and these patients had more severe deficits at presentation. Eighty-seven percent of patients received intravenous heparin after thrombolysis, and 9% of them developed a hemorrhage into infarction. Angiographic recanalization was the rule in complete occlusions of the horizontal portion of the middle cerebral artery, but distal carotid occlusions responded less well to thrombolysis. CONCLUSIONS: The intra-arterial route for thrombolysis allows for greater diagnostic precision and achievement of a higher concentration of the thrombolytic agent in the vicinity of the clot. Disadvantages of this therapy lie in the cost and delay. Severity of stroke and site of angiographic occlusion may be important predictors of successful treatment. PMID- 10512913 TI - Changes in coagulation and fibrinolysis markers in acute ischemic stroke treated with recombinant tissue plasminogen activator. AB - BACKGROUND AND PURPOSE: Shifts of the balance between coagulation and fibrinolysis play a crucial role in pathogenesis and treatment of cerebral ischemia. In this study, we characterized the kinetics of hemostatic abnormalities induced by acute ischemic stroke and its thrombolytic (recombinant tissue plasminogen activator [rtPA]) or anticoagulant (heparin) treatment. METHODS: Systemic generation of molecular markers of hemostasis (fibrin monomer, D-dimer, thrombin-antithrombin complex, and fibrinopeptide 1.2) was monitored in acute ischemic stroke, and the effects of thrombolytic and anticoagulant treatments were analyzed. RESULTS: Thrombolysis with rtPA induced a massive response of markers of coagulation activation and fibrin formation that peaked after 1 to 3 hours and persisted for up to 72 hours. In contrast, only minor hemostatic changes were induced by acute ischemic stroke itself. Administration of heparin did not significantly affect these hemostatic abnormalities. CONCLUSIONS: This first characterization of the coagulation activation induced by rtPA treatment for acute ischemic stroke and the failure to abolish such hemostatic abnormalities by heparin may be of value for further refinement of the currently discussed thrombolytic therapy and the controversial adjunctive anticoagulant prophylaxis in stroke patients. PMID- 10512914 TI - Trends in the incidence, severity, and short-term outcome of stroke in perth, Western Australia. AB - BACKGROUND AND PURPOSE: This report describes trends in the key indices of cerebrovascular disease over 6 years from the end of the 1980s in a geographically defined segment of the city of Perth, Western Australia. METHODS: Identical methods were used to find and assess all cases of suspected stroke in a population of approximately 134 000 residents in a triangular area of the northern suburbs of Perth. Case fatality was measured as vital status at 28 days after the onset of symptoms. Data for first-ever strokes and for all strokes for equivalent periods of 12 months in 1989-1990 and 1995-1996 were compared by age standardized rates and proportions and Poisson regression. RESULTS: There were 355 strokes in 328 patients and 251 first-ever strokes (71%) for 1989-1990 and 290 events in 281 patients and 213 first-ever strokes (73%) for 1995-1996. In Poisson models including age and period, overall trends in the incidence of both first-ever strokes (rate ratio=0.75; 95% confidence limits, 0.63, 0.90) and all strokes (rate ratio=0.73; 95% confidence limits, 0.62, 0.85) were obviously significant, but only the changes in men were independently significant. Case fatality did not change, and the balance between hemorrhagic and occlusive strokes in 1995-1996 was almost indistinguishable from that observed in 1989 1990. CONCLUSIONS: Our results, which are the only longitudinal population-based data available for Australia for key indices of stroke, suggest that it is a change in the frequency of stroke, rather than its outcome, that is chiefly responsible nationally for the fall in mortality from cerebrovascular disease. PMID- 10512915 TI - "Task-oriented" exercise improves hamstring strength and spastic reflexes in chronic stroke patients. AB - BACKGROUND AND PURPOSE: Despite the belief that after cerebral infarction only limited functional gains are possible beyond the subacute period, we tested the hypothesis that a 12-week program of "task-oriented" treadmill exercise would increase muscle strength and decrease spastic reflexes in chronic hemiparetic patients. METHODS: Fourteen subjects, aged 66+/-3 (mean+/-SEM) years, with residual gait deviations due to remote stroke (>6 months), underwent repeated measures of reflexive and volitional (concentric and eccentric) torque with use of isokinetic dynamometry on the hamstring musculature bilaterally. Torque output was measured at 4 angular velocities (30(o), 60(o), 90(o), and 120(o)/s). RESULTS: After 3 months of 3 times/wk low-intensity aerobic exercise, there were significant main effects (2 legs [P<0.01]x2 times [P<0. 01]x4 angular velocities [P<0.05]) for concentric torque production. Torque/time production in the concentric mode also improved significantly in the paretic (50%, P<0.01) and nonparetic hamstrings (31%, P<0.01). Eccentric torque/time production increased by 21% (P<0.01) and 22% (P<0.01) in the paretic and nonparetic hamstrings, respectively. Passive (reflexive) torque/time generation in the paretic hamstrings decreased by 11% (P<0.027). Reflexive torque/time was unchanged in the nonparetic hamstrings (P=0.45). CONCLUSIONS: These findings provide evidence that progressive treadmill aerobic exercise training improves volitional torque and torque/time generation and reduces reflexive torque/time production in the hemiparetic limb. Strength changes associated with improved functional mobility in chronic hemiparetic stroke survivors after treadmill training will be reported in future articles. PMID- 10512917 TI - Correlation between functional and electrophysiological recovery in acute ischemic stroke. AB - BACKGROUND AND PURPOSE: There is still controversy about the prognostic value of motor evoked potentials (MEPs) in the assessment of hemiplegia. The aims of this study are to determine the relationship between functional and electrophysiological recovery and thus the value of MEP as a prognostic indicator of clinical outcome in acute ischemic stroke. METHODS: Seventeen healthy subjects and 38 stroke patients were included in this study. Functional recovery was assessed with the Modified Canadian Neurological Scale (MCNS), the Barthel Activities of Daily Living Index (BI), and the Rankin scale. Transcranial magnetic stimulation was used to determine the change in central motor conduction time (CMCT). Stroke outcome was assessed at the end of 2 weeks. One-way ANOVA with post hoc comparisons using the Scheffe procedure as well as t tests were used to assess the significance of the results in this study. RESULTS: Unpaired t test showed significantly higher mean scores of the MCNS (2P=0.001), BI (2P=0.002), and Rankin scale (P<0.001) at day 14 in the group of patients with recordable MEP at day 1. A better clinical improvement with a higher mean score of the MCNS (2P<0.001), BI (2P<0.001), and the Rankin scale (2P<0.001) was also observed in the patients in whom the CMCT improved. CONCLUSIONS: These data show that there is a close relationship between clinical and electrophysiological improvement and that MEP is a useful prognostic indicator of clinical outcome. PMID- 10512916 TI - Effects of spontaneous recanalization on functional and electrophysiological recovery in acute ischemic stroke. AB - BACKGROUND AND PURPOSE: Transcranial Doppler ultrasound (TCD) studies have shown that spontaneous recanalization results in a better clinical improvement after the onset of stroke. However, its effect on electrophysiological recovery is still unknown. The aim of this study was to determine the effects of spontaneous recanalization on the change in central motor conduction time (CMCT) in acute ischemic stroke. METHODS: Seventeen healthy subjects and 38 consecutive patients with a first acute ischemic stroke involving the middle cerebral artery territory were included. TCD was used to detect spontaneous recanalization. Transcranial magnetic stimulation was used to determine the change in CMCT on days 1 and 14. Improvement of the CMCT at day 14 was indicated if it decreased in comparison with previous data recorded at day 1 or when a nonrecordable motor response at day 1 reappeared at day 14. No CMCT improvement was indicated when there was no recordable motor response at day 1 and day 14 or the CMCT at day 14 worsened, becoming absent or more delayed. The Pearson chi(2) test was used to assess the statistical significance of the results in this study. RESULTS: Spontaneous recanalization was observed in 62% of the patients: 24% before 24 hours and 38% after this period. No recanalization was observed in 14 patients. The CMCT improved in 87% of the patients who had recanalized before 24 hours and 62% in the recanalized after 24 hours group (P=0.005). In contrast, CMCT improved in only 17% of the patients in the non-recanalized group CONCLUSIONS: These data show that spontaneous recanalization results in a better recovery of the central motor pathway leading to a better CMCT improvement in acute ischemic stroke. PMID- 10512918 TI - The stroke impact scale version 2.0. Evaluation of reliability, validity, and sensitivity to change. AB - BACKGROUND AND PURPOSE: To be useful for clinical research, an outcome measure must be feasible to administer and have sound psychometric attributes, including reliability, validity, and sensitivity to change. This study characterizes the psychometric properties of the Stroke Impact Scale (SIS) Version 2.0. METHODS: Version 2.0 of the SIS is a self-report measure that includes 64 items and assesses 8 domains (strength, hand function, ADL/IADL, mobility, communication, emotion, memory and thinking, and participation). Subjects with mild and moderate strokes completed the SIS at 1 month (n=91), at 3 months (n=80), and at 6 months after stroke (n=69). Twenty-five subjects had a replicate administration of the SIS 1 week after the 3-month or 6-month test. We evaluated internal consistency and test-retest reliability. The validity of the SIS domains was examined by comparing the SIS to existing stroke measures and by comparing differences in SIS scores across Rankin scale levels. The mixed model procedure was used to evaluate responsiveness of the SIS domain scores to change. RESULTS: Each of the 8 domains met or approached the standard of 0.9 alpha-coefficient for comparing the same patients across time. The intraclass correlation coefficients for test-retest reliability of SIS domains ranged from 0.70 to 0.92, except for the emotion domain (0.57). When the domains were compared with established outcome measures, the correlations were moderate to strong (0.44 to 0.84). The participation domain was most strongly associated with SF-36 social role function. SIS domain scores discriminated across 4 Rankin levels. SIS domains are responsive to change due to ongoing recovery. Responsiveness to change is affected by stroke severity and time since stroke. CONCLUSIONS: This new, stroke-specific outcome measure is reliable, valid, and sensitive to change. We are optimistic about the utility of measure. More studies are required to evaluate the SIS in larger and more heterogeneous populations and to evaluate the feasibility and validity of proxy responses for the most severely impaired patients. PMID- 10512919 TI - Role for telemedicine in acute stroke. Feasibility and reliability of remote administration of the NIH stroke scale. AB - BACKGROUND AND PURPOSE: Immediate access to physicians experienced in acute stroke treatment may improve clinical outcomes in patients with acute stroke. Interactive telemedicine can make stroke specialists available to assist in the evaluation of patients at multiple urban or remote rural facilities. We tested whether interrater agreement for the NIH Stroke Scale (NIHSS), a critical component of acute stroke assessment, would persist if performed over a telemedicine link. METHODS: One bedside and 1 remote NIHSS score were independently obtained on each of 20 patients with ischemic stroke. The bedside examination was performed by a stroke neurologist at the patient's bedside. The remote examination was performed by a second stroke neurologist through an interactive high-speed audio-video link, assisted by a nurse at the patient's bedside. Kappa coefficients were calculated for concordance between bedside and remote scores. RESULTS: Remote assessments took slightly longer than bedside assessments (mean 9.70 versus 6.55 minutes, P<0. 001). NIHSS scores ranged from 1 through 24. Based on weighted kappa coefficients, 4 items (orientation, motor arm, motor leg, and neglect) displayed excellent agreement, 6 items (language, dysarthria, sensation, visual fields, facial palsy, and gaze) displayed good agreement, and 2 items (commands and ataxia) displayed poor agreement. Total NIHSS scores obtained by bedside and remote methods were strongly correlated (r=0.97, P<0.001). CONCLUSIONS: The NIH Stroke Scale remains a swift and reliable clinical instrument when used over interactive video. Application of this technology can bring stroke expertise to the bedside, regardless of patient location. PMID- 10512920 TI - How do scores on the EuroQol relate to scores on the SF-36 after stroke? AB - BACKGROUND AND PURPOSE: The EuroQol and Medical Outcome Survey 36-item short-form health survey (SF-36) questionnaires have both been validated for the assessment of health-related quality of life after stroke. However, the relationship between these instruments has not been studied after stroke. We therefore sought to compare the responses of a group of stroke patients to both instruments. METHODS: A total of 2253 patients with stroke entered by United Kingdom hospitals in the International Stroke Trial were randomized to follow-up with either the EuroQol or SF-36 instruments. We randomly selected one third of patients who had responded to the EuroQol for follow-up, again using the SF-36, and two thirds of patients who had responded to the SF-36 for follow-up, again using the EuroQol. We assessed the patients' mean score for each domain of the SF-36 categorized by their response to the corresponding EuroQol domain and the correlation between the domains of the 2 instruments. RESULTS: The domains for both instruments, which assessed physical functioning, social functioning, bodily pain, and overall health-related quality of life, correlated closely. The mental health domain of the SF-36 correlated only poorly with the psychological functioning domain of the EuroQol. CONCLUSIONS: Both the EuroQol and SF-36 measure broadly similar domains of health. The weak relationship between the assessments of mental health may reflect a difference in content or more fundamental problems with the validity or reliability of the items in one of the instruments with respect to this domain. This study has provided the first empirical qualitative evidence by which the data on the SF-36 after stroke may be interpreted. PMID- 10512921 TI - Lateralization of cerebral blood flow velocity changes during cognitive tasks. A simultaneous bilateral transcranial Doppler study. AB - BACKGROUND AND PURPOSE: Transcranial Doppler ultrasonography (TCD) permits the assessment of cognitively induced cerebral blood flow velocity (BFV) changes. We sought to investigate the lateralization of BFV acceleration induced by a variety of cognitive tasks and to determine the influence of age, gender, IQ, and quality of the performance on the relative BFV changes. METHODS: Simultaneous bilateral TCD monitoring of BFV in the middle cerebral arteries (MCAs) was performed in 90 normal right-handed volunteers during 13 verbal and visuospatial tasks and their preceding rest periods. RESULTS: All tasks induced a significant bilateral BFV increase in the MCAs compared with the preceding rest periods. Five verbal tasks showed a significant left-hemispheric BFV acceleration. Linguistic tasks that required active or creative processing of the verbal stimuli, such as sentence construction or word fluency, elicited the most asymmetric response. Five visuospatial tasks revealed a significant right-hemispheric BFV shift. Paradigms that combined visuospatial attention and visuomotor manipulation showed the most lateralized acceleration. Older volunteers (aged >50 years) showed higher relative BFV changes, but lateralization was not influenced by age. Gender, IQ, and performance quality did not reveal significant effects on BFV change. CONCLUSIONS: Bilateral TCD is a noninvasive technique that has the potential to connect the particular change in flow pattern of the MCA distribution with selective cognitive activity and thus offers specific functional information of scientific and clinical value. PMID- 10512922 TI - Cerebrovascular disease and depression symptoms in the cardiovascular health study. AB - BACKGROUND AND PURPOSE: Evidence is mounting linking cerebrovascular disease with depressive symptoms in the elderly. Lesions in both white and gray matter have been associated with depressive symptoms and major depression. We sought to investigate the relationship between depressive symptoms and white and gray matter lesions in subjects participating in the Cardiovascular Health Study. METHODS: In a sample of 3660 men and women who underwent a standardized interview, physical examination, and MRI scan, we examined the association between number of white and gray matter lesions and white matter grade (a measure of severity) and reported depressive symptoms using a modified version of the Centers for Epidemiologic Studies Depression (CES-D) scale. We controlled for a variety of demographic and medical variables as well as functional status and Modified Mini-Mental State Examination score. RESULTS: The number of small (<3 mm) basal ganglia lesions was significantly associated with reported depressive symptoms, but white matter grade was not. In subsequent logistic regression models, number of basal ganglia lesions remained a significant predictor after controlling for non-MRI variables and severity of white matter lesions. CONCLUSIONS: Our findings extend previous reports that linked cerebrovascular changes to depressive symptoms in clinical populations to a large community-based population. This report provides further evidence of the importance of basal ganglia lesions in geriatric depression. PMID- 10512923 TI - Neuropsychological impairment in stroke, carotid stenosis, and peripheral vascular disease, A comparison with healthy community residents. AB - BACKGROUND AND PURPOSE: An increasing body of literature suggests a role for clinically "silent" cerebrovascular disease in the pathogenesis of cognitive impairment. Such pathology commonly occurs in the absence of stroke. The main aim of the study was to examine neuropsychological impairment associated with cerebrovascular and peripheral vascular disease (PVD) and to compare cognitive deficits with a nonvascular control group. The main hypothesis was that older people with both transient ischemic attack (TIA) and PVD would demonstrate greater cognitive impairment than controls. METHODS: A battery of neuropsychological tests was administered to 4 groups of community residents older than 65 years. The groups comprised 25 patients with carotid stenosis and TIA, 25 nonamputees with PVD, 25 patients with stroke, and 25 matched (with the stroke group) controls. RESULTS: Stroke patients showed greater impairment than controls in all tests. PVD patients did not perform significantly worse (P<0.05 after Bonferroni correction) than control subjects on any of the neuropsychological tests. However, 25% of PVD patients had scores lying within the bottom 5% of control group scores for attention, calculation, and 1 test of frontal lobe function. TIA patients were more impaired in general intellectual impairment and frontal lobe function than controls. Frontal lobe impairment, suicidal thinking, and age were all independent predictors of global cognitive impairment in the TIA group. Frontal lobe impairment was the only predictor of global cognitive impairment in the PVD group. CONCLUSIONS: TIA and PVD patients showed similar patterns of neuropsychological impairment, but TIA may result in more prolonged cognitive impairment, particularly in frontal lobe function. This group may be at increased risk of vascular dementia as well as impulsivity and suicide. PMID- 10512924 TI - Increased IL-1beta, IL-8, and IL-17 mRNA expression in blood mononuclear cells observed in a prospective ischemic stroke study. AB - BACKGROUND AND PURPOSE: Ischemic brain injury secondary to arterial occlusion is characterized by acute local inflammation, which involves accumulation of polymorphonuclear neutrophils (PMN). Factors that influence the recruitment of PMN could represent new therapeutic targets in acute stroke. In this prospective study we evaluated numbers of peripheral blood mononuclear cells (PBMC) expressing mRNA for interleukin (IL)-1beta, IL-8, and IL-17 and macrophage inflammatory protein-1alpha (MIP-1alpha) after ischemic stroke. METHODS: Peripheral blood was obtained on days 1 to 3, 4 to 10, and 20 to 31 after onset of symptoms. In situ hybridization with radiolabeled synthetic oligonucleotide probes was adopted to measure cytokine mRNA expression in PBMC. Plasma and cerebrospinal fluid levels of IL-8 were measured by an enzyme-linked immunosorbent assay. RESULTS: Most patients with ischemic stroke had clearly elevated numbers of IL-1beta, IL-8, and IL-17 mRNA expressing PBMC 1 to 3 days after onset of symptoms compared with healthy individuals (P<0. 0001 for all comparisons). At follow-up after 20 to 31 days, numbers of IL-8 mRNA expressing PBMC were lower than during the acute stage (P<0.001), but only IL-1beta and IL 17 mRNA expression had returned to the level of the healthy individuals. Numbers of MIP-1alpha mRNA expressing PBMC did not differ between patients with ischemic stroke and healthy individuals at any time point. A correlation was observed between numbers of IL-1beta, IL-8, and IL-17 mRNA expressing PBMC and the degree of neurological impairment as measured by the Scandinavian Stroke Scale 1 to 3 days after onset of symptoms (r=0.5; P<0.01 for all correlations). CONCLUSIONS: A longitudinal study of patients with ischemic stroke revealed systemic increases of levels of IL-1beta, IL-8, and IL-17 that correlated with Scandinavian Stroke Scale scores. IL-8 levels were further increased in cerebrospinal fluid. PMID- 10512925 TI - The A677V methylenetetrahydrofolate reductase gene polymorphism and carotid atherosclerosis. AB - BACKGROUND AND PURPOSE: The alanine/valine (A/V) polymorphism at codon 677 of the 5,10 methylenetetrahydrofolate reductase (MTHFR) gene correlates with elevated levels of plasma homocysteine and with an increased risk of atherosclerotic cardiovascular disease. Our study was designed to assess the frequency of the A and V alleles in patients with asymptomatic severe carotid artery stenosis (CAS) assessed by extracranial duplex examination in comparison with age- and sex matched subjects without carotid atherosclerosis. METHODS: Consecutive patients (n=48; 28 men, mean+/-SD age 67.1+/-11. 4 years) with asymptomatic severe (>75%) CAS were compared with subjects without CAS (n=26; 15 men, aged 61.2+/-11.5). The MTHFR genotype was analyzed by polymerase chain reaction followed by HinfI digestion. The chi(2) analysis and t test were used to compare the groups. RESULTS: The frequency of V alleles was significantly higher in the CAS group (0.47) compared with control subjects (0.27, chi(2) test; OR 2.4 [95% CI 1.1 to 5.3]; P<0.02). CONCLUSIONS: Our results indicate that the MTHFR A677V allele is significantly associated with severe CAS. PMID- 10512926 TI - Intracerebral calcification in systemic sclerosis. AB - BACKGROUND AND PURPOSE: Advanced cerebrovascular wall calcification was recently observed at autopsy in 2 patients with systemic sclerosis. To further investigate this issue, we conducted a prospective CT study of scleroderma patients to detect intracerebral calcification. METHODS: Thirty-seven consecutive patients with systemic sclerosis underwent unenhanced brain CT. Images were blindly interpreted, together with those of 2 age-matched (+/-1 year) and sex-matched control subjects per patient. RESULTS: Intracerebral calcification was found in 12 patients (32.4%) and 7 controls (9.5%) (P=0.006). Among the patients, intracerebral calcification was associated with the duration of Raynaud's phenomenon (P=0.005) and not with age (P=0.086). CONCLUSIONS: Intracerebral calcification is closely associated with scleroderma, which suggests that scleroderma causes primary cerebrovascular changes. PMID- 10512927 TI - Chronic estrogen treatment increases levels of endothelial nitric oxide synthase protein in rat cerebral microvessels. AB - BACKGROUND AND PURPOSE: A number of studies indicate that the female gonadal hormone, estrogen, confers protection against cerebrovascular disorders such as stroke. One postulated mechanism for these effects of estrogen is an action on the enzyme endothelial nitric oxide synthase (eNOS), which produces the vasodilatory molecule NO. We have investigated the hypothesis that estrogen increases expression of eNOS in cerebral microvessels of male and female rats. METHODS: We measured levels of eNOS protein by Western blot in cerebral microvessels isolated from 7 groups of animals: females, ovariectomized females, ovariectomized females treated with estrogen, males, castrated males, castrated males treated with estrogen, and castrated males treated with testosterone. RESULTS: Ovariectomized female rats treated with estrogen had 17. 4-fold greater levels of eNOS protein in cerebral microvessels than ovariectomized females, and intact females had 16.6-fold greater levels than ovariectomized females (P<0.01). In intact females, cerebral microvessel eNOS protein levels were 9.2-fold higher than those of intact males (P<0.05). Levels of eNOS protein in castrated males, castrated males treated with testosterone, and males were not different from each other. Estrogen treatment of castrated animals resulted in an 18.8-fold increase in cerebral microvessel eNOS protein (P<0.05). CONCLUSIONS: Chronic estrogen treatment increases levels of eNOS protein in cerebral microvessels of male and female rats. This increase in eNOS protein correlates with our previous functional findings indicating that estrogen exposure increases NO modulation of cerebrovascular reactivity in both male and female animals. Upregulation of eNOS expression may contribute to the neuroprotective effect of estrogen. PMID- 10512928 TI - Vascular effects of lipopolysaccharide are enhanced in interleukin-10-deficient mice. AB - BACKGROUND AND PURPOSE: The role in blood vessels of interleukin-10 (IL-10), a potent anti-inflammatory cytokine, is not known. Using mice with targeted deletion of the gene for IL-10 (IL-10(-/-)), we examined the hypothesis that IL 10 is a major modulator of the vascular effects of lipopolysaccharide (LPS). Methods-We examined in vitro responses of carotid arteries obtained from wild type (129/SvEv or C57BL/6; IL-10(+/+)) and IL-10-deficient mice 6 hours after injection of a relatively low dose of LPS (10 microgram). RESULTS: Contraction of the carotid artery in response to U46619 was impaired in IL-10-deficient mice treated with LPS compared with LPS-treated controls. After LPS, U46619 (0.03 and 0.1 microgram/mL) contracted the carotid artery by 0.11+/-0.02 (mean+/-SEM) and 0.38+/-0.03 g in wild-type (n=10) and 0.03+/-0.01 and 0.19+/-0.03 g in IL-10 deficient (n=8) mice (P<0.05 versus control). Aminoguanidine, an inhibitor of inducible nitric oxide synthase (iNOS), had no significant effect on contraction of the carotid artery from LPS-treated control mice but restored contraction of the carotid artery in response to U46619 in IL-10-deficient mice to levels seen in wild-type mice. Similar findings were obtained when phenylephrine was used as a vasoconstricting agent. These findings indicate that LPS produces much greater impairment of contractile responses of the carotid artery in IL-10-deficient mice than in control mice. Impaired contractile function was eliminated by aminoguanidine, suggesting that expression of iNOS is enhanced in arteries from IL-10-deficient mice. In carotid arteries from animals injected with LPS, reverse transcription-polymerase chain reaction (RT-PCR) products for iNOS were found more frequently in IL-10-deficient mice than in wild-type mice. RT-PCR products for iNOS were not present in arteries from vehicle-treated animals (IL-10 deficient or wild-type mice). CONCLUSIONS: This is the first evidence that endogenous IL-10 is a major determinant of the effects of LPS on vascular tone. The results suggest that impaired constrictor responses of the carotid artery after LPS in IL-10-deficient mice are mediated by enhanced expression of iNOS. PMID- 10512929 TI - Cerebrovascular dynamics of autoregulation and hypoperfusion. An MRI study of CBF and changes in total and microvascular cerebral blood volume during hemorrhagic hypotension. AB - BACKGROUND AND PURPOSE: To determine how cerebral blood flow (CBF), total and microvascular cerebral blood volume (CBV), and blood oxygenation level-dependent (BOLD) contrast change during autoregulation and hypotension using hemodynamic MRI. METHODS: Using arterial spin labeling and steady-state susceptibility contrast, we measured CBF and changes in both total and microvascular CBV during hemorrhagic hypotension in the rat (n=9). RESULTS: We observed CBF autoregulation for mean arterial blood pressure (MABP) between 50 and 140 mm Hg, at which average CBF was 1.27+/-0.44 mL. g(-1). min(-1) (mean+/-SD). During autoregulation, total and microvascular CBV changes were small and not significantly different from CBF changes. Consistent with this, no significant BOLD changes were observed. For MABP between 10 and 40 mm Hg, total CBV in the striatum increased slightly (+7+/-12%, P<0.05) whereas microvascular CBV decreased (-15+/-17%, P<0.01); on the cortical surface, total CBV increases were larger (+21+/-18%, P<0.01) and microvascular CBV was unchanged (3+/-22%, P>0.05). With severe hypotension, both total and microvascular CBV decreased significantly. Over the entire range of graded global hypoperfusion, there were increases in the CBV/CBF ratio. CONCLUSIONS: Parenchymal CBV changes are smaller than those of previous reports but are consistent with the small arteriolar fraction of total blood volume. Such measurements allow a framework for understanding effective compensatory vasodilation during autoregulation and volume-flow relationships during hypoperfusion. PMID- 10512930 TI - Effects of some guanidino compounds on human cerebral arteries. AB - BACKGROUND AND PURPOSE: Accumulation of endogenous guanidino-substituted analogues of L-arginine in chronic renal failure might contribute to some of the vascular and neurological disorders of this pathology. We tested the hypothesis that in human cerebral arteries, some guanidino compounds may increase vascular tone, through nitric oxide (NO) synthase inhibition, and impair endothelium dependent relaxation. METHODS: Rings of human middle cerebral artery were obtained during autopsy of 26 patients who had died 3 to 12 hours before. The rings were suspended in organ baths for isometric recording of tension. We then studied the responses to N(G)-monomethyl-L-arginine (L-NMMA), N(G),N(G)-dimethyl L-arginine (asymmetrical dimethylarginine; ADMA), aminoguanidine (AG), and methylguanidine (MG). RESULTS: L-NMMA (10(-6) to 3x10(-4) mol/L) and ADMA (10(-6) to 3x10(-4) mol/L) caused concentration- and endothelium-dependent contractions (median effective concentrations [EC(50)]=1.1x10(-5) and 1.6x10(-5) mol/L, respectively; E(max)=35. 5+/-7.9% and 43.9+/-5.9% of the response to 100 mmol/L KCl). AG (10(-5) to 3x10(-3) mol/L) and MG (10(-5) to 3x10(-3) mol/L) produced endothelium-independent contractions (E(max)=44.3+/-8.8% and 45.7+/-5.8% of the response to 100 mmol/L KCl, respectively). L-Arginine (10(-3) mol/L) prevented the contractions by L-NMMA and ADMA but did not change contractions induced by AG and MG. L-NMMA and ADMA inhibited endothelium-dependent relaxation induced by acetylcholine in a concentration-dependent manner; AG and MG were without effect. CONCLUSIONS: The results suggest that the contractions induced by L-NMMA and ADMA are due to inhibition of endothelial NO synthase activity, whereas AG and MG do not affect the synthesis of NO. An increase in the plasma concentration of L-NMMA and ADMA associated with uremia is likely to represent a diminished release or effect of NO, and consequently, an increased cerebrovascular tone in uremic patients is highly conceivable. PMID- 10512931 TI - Rapid monitoring of diffusion, DC potential, and blood oxygenation changes during global ischemia. Effects of hypoglycemia, hyperglycemia, and TTX. AB - BACKGROUND AND PURPOSE: The increasing interest in diffusion-weighted MRI (MRI) for diagnosis and monitoring of acute stroke in humans calls for a sound understanding of the underlying mechanisms of this image contrast in acute cerebral ischemia. The present study aimed to show that a rapid decrease in brain water apparent diffusion coefficient (ADC) occurs coincident with anoxic depolarization and that this change is delayed by hyperglycemia and sodium channel blockade but accelerated by hypoglycemia. METHODS: Rats were divided into groups: normoglycemic, hypoglycemic, and hyperglycemic, and those given local tetrodotoxin (TTX) application. Cardiac arrest was effected by intravenous KCl injection during serial high-speed diffusion and blood oxygenation-sensitive gradient-recalled echo MRI. Brain DC potential was recorded simultaneously. Serial ADC maps were calculated from the diffusion-weighted data and fitted to a model function to measure the delay between cardiac arrest and rapid ADC decrease. RESULTS: The time of anoxic depolarization indicated by DC change agreed well with the rapid drop in ADC in all groups; both were accelerated with hypoglycemia and delayed by hyperglycemia. A more gradual ADC decline occurred before anoxic depolarization, which was more pronounced in hyperglycemic animals and less pronounced in hypoglycemic animals. Rapid drop in ADC was also delayed by local TTX application. Changes in gradient-recalled echo image intensity were not significantly different among groups. CONCLUSIONS: While much of the ADC decrease in ischemia occurs during anoxic depolarization, significant but gradual ADC changes occur earlier that may not be due to a massive loss in ion homeostasis. PMID- 10512932 TI - Analysis of the perception of and reactivity to pain and heat in patients with wallenberg syndrome and severe spinothalamic tract dysfunction. AB - BACKGROUND: The aim of the study was to assess the consequences of severe spinothalamic tract lesions resulting from lateral medullary infarct and to show that a specific pain perception can be elicited by strong thermal stimulation. CASE DESCRIPTIONS: Both patients examined presented with severe thermoalgic dissociation of the limbs contralateral to the lesion, with normal discriminative somatosensory perception and motor strength. They reported pain perception when touching very warm (>50 degrees C to 60 degrees C) objects and a brisk, occasionally uncontrolled withdrawal reaction of the arm and hand under the same conditions, without any perception of the heat nature of the stimulus. Warm stimulation, <45 degrees C, elicited no thermal perception or discrimination. Pain perception could be elicited in both patients by increasing the temperature, with a reproducible threshold of 47 degrees C to 49 degrees C. Pain always occurred after a prolonged delay of 8 to 10 seconds in response to threshold heat, and was described as deep and osseous, and clearly different from that perceived on the nonaffected side. The delay was much shorter when the temperature was increased by 4 degrees C to 5 degrees C. Cold stimulation elicited similar pain perception in one patient. Analysis of subjective perception of laser stimulation showed a much higher pain threshold on the affected hand. There were no laser-evoked potentials on this side, which suggested major spinothalamic injury. Assessment of the RIII noxious reflex revealed persistent response withdrawal reactions, with an increased threshold on the affected side, and partial consciousness of the noxious nature of the stimulus. CONCLUSIONS: To our knowledge, this is the first description of the appearance of pain perception of high temperatures in patients with severe spinothalamic injury who are suffering from a complete loss of temperature perception. This implies that noxious thermal stimulation can still be perceived via extra spinothalamic pathways (which are slow and multisynaptic), such as the spinoreticulothalamic tract. Patients with Wallenberg syndrome should be informed and made aware of their residual perception of and reactions to noxious stimulation. PMID- 10512933 TI - Expanding the window for thrombolytic therapy in acute stroke. The potential role of acute MRI for patient selection. AB - BACKGROUND: Effective therapy was not available for treatment of acute stroke until 1995, when tissue plasminogen activator (tPA) was shown to improve neurological and functional outcome in stroke patients who were treated within 3 hours of symptom onset. SUMMARY OF REVIEW: Currently, many patients do not qualify for tPA therapy because they present for evaluation beyond 3 hours after stroke onset. Attempts to expand the treatment window to 6 hours, using CT to select patients, have failed. Use of early MR imaging may provide significant advantages over CT for identification of patients who are likely to benefit from thrombolytic therapy because (1) the early perfusion-weighted imaging (PWI) lesion estimates the region of acute dysfunctional brain tissue, whereas the acute diffusion-weighted imaging (DWI) lesion appears to correspond to the core of the early infarction; (2) the mismatch between the acute PWI lesion and the smaller DWI lesion represents potentially salvageable brain tissue (an estimate of the ischemic penumbra); and (3) in patients with a PWI/DWI mismatch, early reperfusion is often associated with substantial clinical improvement and reversal or reduction of DWI lesion growth. CONCLUSIONS: Clinical trials that use new MRI techniques to screen patients may be able to identify a subset of acute stroke patients who are ideal candidates for thrombolytic therapy even beyond 3 hours after stroke onset. PMID- 10512934 TI - Heart rate variability following ischemic stroke. PMID- 10512936 TI - The following is a list of major ongoing studies about stroke. Information about other multicenter studies that might be included in this list should be submitted to the Stroke Editorial Office by the Principal Investigator. The list will appear in the February, June, and October issues of Stroke. PMID- 10512935 TI - Abstracts of Literature. PMID- 10512937 TI - Tumor seeding following laparoscopy: international survey. AB - The aim of the study was to determine if tumor seeding during laparoscopic surgery for cancer is a rare event or a typical complication of this procedure. Laparoscopic staging and treatment of intraabdominal tumors is increasing in gastroenterology, gynecology, and general surgery. A total of 1052 questionnaires were mailed to surgical department chairmen, members of the German Society of Surgery, Swiss Association for Laparoscopic and Thoracoscopic Surgery, and Austrian Society of Minimal Invasive Surgery asking them to list their department's experience with tumor seeding after laparoscopy for nonapparent or known malignancy. There were 607 (57.7%) surgeons who reported a total of 117,840 laparoscopic cholecystectomies, 409 incidental gallbladder carcinomas, and 412 laparoscopies on patients with colorectal carcinoma. Altogether 109 patients who developed tumor recurrence in connection with laparoscopic surgery have been reported. Port-site recurrence was identified in 70 of 409 patients (17.1%) with a median of 180 days following laparoscopic cholecystectomy for nonapparent gallbladder carcinoma. In 8 cases (11.5%) a protective plastic bag had been used for gallbladder retrieval. Six patients without port-site metastases were found to have a diffuse peritoneal carcinomatosis a median of 120 days after cholecystectomy. Of 412 laparoscopies for colorectal cancer, 19 cases (4.6%) of tumor seeding have been reported, 16 of which (3.9%) had documented port-site and scar recurrences a median of 196 days after laparoscopy. The tumor specimen was intact, and a plastic bag was used for extraction in seven cases. In 14 patients trocar-site metastases have been reported a median of 70 days after laparoscopy for different nonapparent or known malignancies. The probability of developing abdominal wall metastasis is higher after laparoscopy for cancer than after open surgery. An intact surgical specimen and the use of a plastic retrieval bag do not exclude the risk of port-site recurrences. These facts and the early appearance of peritoneal carcinosis in a few cases of intraabdominal malignancies seem to confirm a specific laparoscopic risk for intraperitoneal tumor cell seeding and implantation. PMID- 10512938 TI - Laparoscopy and laparoscopic ultrasonography for staging pancreatic cancer: critical appraisal. AB - A pilot study was designed to elucidate the role of staging laparoscopy for determining resectability in patients with pancreatic cancer. The additional value of laparoscopic contact ultrasonography (LCU) was also evaluated with specific regard to its ability to detect hepatic metastases and assess vascular infiltration of the portomesenteric trunk. A consecutive sample of 50 patients referred for operation of a suspected pancreatic cancer were submitted to preoperative contrast-enhanced high-resolution computed tomography (CT) and staging laparoscopy combined with LCU at a university hospital. For those progressing to exploratory laparotomy, the intraoperative findings relating to tumor diffusion and vascular infiltration were compared to CT, laparoscopic, and LCU data. Analytical description of the laparoscopic findings is given. Row data of predicted versus observed vascular infiltration were tabulated for CT and LCU. The sensitivity, specificity, and overall accuracy of each diagnostic test were calculated for comparative analysis. Laparoscopy alone prevented unnecessary laparotomy in 20% of cases. A complete procedure could not be achieved in 28% of patients. Three false-negative staging results occurred. LCU identified small (benign) hepatic nodules not seen by CT in 8% of patients. Sensitivity, specificity, and overall accuracy for assessing vascular infiltration were 82%, 53%, and 69% for CT and 94%, 80%, and 87% for LCU. Laparoscopy was confirmed to be safe and effective for staging pancreatic cancer. Because of its unique capabilities to detect even small peritoneal tumor deposits a quick exploration immediately before laparotomy is advised in all patients. The additional benefit of a more extensive procedure is not supported by our results. Although LCU appears to define the vascular involvement more accurately than conventional CT, the limitation of getting clinically useful ultrasound data in all the patients suggests its adoption in only a selected population. PMID- 10512939 TI - Tension-free laparoscopic and open hernia repair: randomized controlled trial of early results. AB - The aim of the study was prospectively to compare the early results and outcome in 105 patients randomly allocated to undergo tension-free laparoscopic hernia repair (LHR) with transabdominal preperitoneal technique (53 patients) or open hernia repair (OHR) with mesh apposition (52 patients). The mean (SD) operation time was longer in the LHR group than in the OHR group: 49.6 (5.4) versus 33. 9 (6.2) minutes; p < 0.001. One laparoscopic case was converted to open repair to deal with a hemorrhage from an aberrant obturatory artery at the level of Cooper's ligament. Groin discomfort or pain was the most common complication after both procedures. The patients requiring none, one, two, or more than two doses of intramuscular diclofenac were, respectively, 40.4%, 40.4%, 15.4%, and 3.8% after LHR and 50.0%, 30.8%, 17.3%, and 1.9% after OHR (p = 0.69; NS). The mean +/- SEM (range) postoperative visual analog scale score, ranging from 0 (no pain) to 10 (worst pain imaginable), was 3.1 +/- 0.2 (1-7) in the LHR subset and 2.7 +/- 0.2 (1-5) in the OHR group (p = 0.14; NS); on the second postoperative day the score was 2.3 +/- 0.2 (1-6) and 1.8 +/- 0.1 (1-4), respectively (p < 0.03). The time +/- SEM (range) of resumption of pain-free normal activities and work was faster in OHR group: 6.1 +/- 0.2 (4-8) weeks versus 6.5 +/- 0.1 (4-8) weeks; p < 0.03. Our results showed that tension-free open hernia repair is superior to LHR in terms of postoperative pain with no important differences in recovery. 0.05). In addition, the simultaneous presence of these receptors did not influence survival. Our findings indicate that in esophageal cancer the presence of EGF-R, c-erbB-2, and c-erbB-3 alone or in combination seems to have no major influence on patient prognosis and does not alter tumor growth behavior significantly. PMID- 10512941 TI - Interleukin-6 and tumor necrosis factor-alpha levels following hepatic cryotherapy: association with volume and duration of freezing. AB - Although morbidity following cryotherapy is usually minor, a syndrome of multiorgan failure and disseminated intravascular coagulation (DIC) has been described and referred to as the cryoshock phenomenon. We hypothesized that mediators similar to those in septic shock may be involved in this syndrome. In this study we aimed to assess the plasma concentrations of the cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) following hepatic cryotherapy and to relate them to the duration and volume of freezing and to hepatocellular injury. Between April and December 1997 blood samples were taken preoperatively and at different times postoperatively from patients undergoing hepatic artery catheter-insertion (HAC) (n = 15), cryotherapy (n = 5), liver resection (n = 9), liver resection and edge cryotherapy (n = 7), or liver resection and cryotherapy of additional lesions (n = 9). They were analyzed for serum aspartate transaminase (AST) and plasma TNF-alpha and IL-6 levels. There was a significant association (Pearson correlation) of serum AST levels 1 hour postoperatively with plasma TNF-alpha and IL-6 levels at the end of the procedure. In patients undergoing cryotherapy or resection with cryotherapy of additional lesions (n = 14), the volume and duration of hepatic freezing were significantly associated with postoperative serum AST and plasma TNF-alpha and IL 6 levels at various postoperative times. Hepatic cryotherapy is followed by cytokine release, with postoperative plasma TNF-alpha and IL-6 levels associated with the degree of hepatic cryotrauma. These mediators may be involved in the occurrence of cryoshock following large-volume hepatic freezing. PMID- 10512942 TI - Effect of protease inhibitor on ischemia-reperfusion injury to rat liver. AB - Liver failure due to ischemia-reperfusion injury, believed to be closely related to the generation of oxygen-free radicals, is a serious problem during liver surgery. Gabexate mesilate, a synthetic protease inhibitor, suppresses the extracellular release of oxygen-free radicals in the microvascular endothelium. To determine its effects on ischemia-reperfusion injury to the liver, we performed experiments with rats. We divided the animals into two ischemia reperfusion groups: an experimental group, which underwent ischemic injury for 30 minutes, along with the infusion of gabexate mesilate, and a control group, which underwent injury only. Each group was then divided into four subgroups: ischemic injury only and 60-, 120-, and 180-minute reperfusion injury. The test parameters were tumor necrosis factor alpha (TNFalpha) and interleukin-6 (IL-6) in serum and superoxide dismutase (SOD), catalase, and malondialdehyde (MDA) in liver and lung tissues. The experimental group had a significantly higher liver SOD and catalase levels and a significantly lower level of liver and lung MDA than the control groups. TNFalpha levels in the experimental groups were significantly lower during the early phase, but a comparison of IL-6 levels between the two groups yielded no differences. Levels of lung catalase and SOD were not significantly different between the two groups. We concluded that protease inhibitor suppressed liver ischemia-reperfusion injury, and that it was due to an increase of antioxidant or suppression of oxygen-free radicals. The roles of TNFalpha and IL 6 in liver reperfusion injury were not clear, though TNFalpha might have had an effect during the early phase. With liver ischemia-reperfusion injury, the mechanism of lung involvement might be different from that of liver involvement. PMID- 10512943 TI - New simple technique for hepatic parenchymal resection using a Cavitron Ultrasonic Surgical Aspirator and bipolar cautery equipped with a channel for water dripping. AB - We have developed a new technique to resect hepatic parenchyma without inflow occlusion by using the Cavitron Ultrasonic Surgical Aspirator (CUSA) and bipolar cautery with a saline irrigation system. The significance of this method in hepatectomy was analyzed in comparison with historical control of hepatectomy using Pringle's maneuver. An ordinary bipolar cautery was remodeled with an infusion line to bring saline droplets down the inner surface of one arm of the tweezers through an opening about 1.5 cm proximal to its tip. The optimal flow rate of saline was approximately one drop per second. The power of bipolar cautery was adjusted to 50 watts. When the tweezer blades were approximated to 1 or 2 mm, saline droplets were directed to the tip of tweezers and could be immediately evaporated. After sonicating parenchymal cells, the tissue of small branches of Glisson's tree or small tributaries of the hepatic vein were coagulated by bipolar cautery. The coagulated cords were then easily cut by scissors. The impact of this technique on ordinary liver resections was evaluated by analyzing the postoperative clinical course in relation to the hepatic functional reserve necessary for major hepatectomy, duration of hepatectomy, and intraoperative blood loss. Hepatic resection without vascular occlusion using this technique could decrease the morbidity in patients who have less hepatic functional reserve. It could also decrease intraoperative blood loss. This new technique effectively decreased the surgical load of the remnant liver during parenchymal resection by avoiding ischemic stress. Consequently it extends the safety limits of major hepatectomy. PMID- 10512944 TI - Treatment strategy for mucin-producing intrahepatic cholangiocarcinoma: value of percutaneous transhepatic biliary drainage and cholangioscopy. AB - Intrahepatic cholangiocarcinomas that secrete macroscopically excessive mucin into the biliary system are rare, and few of the previously reported cases have achieved a curative resection. We defined these tumors as "mucin-producing intrahepatic cholangiocarcinomas" and clarify the optimal preoperative and surgical management for them. Eleven patients with mucin-producing intrahepatic cholangiocarcinomas underwent surgical resection in our department. The clinical, radiologic, surgical, and pathologic findings were studied. The clinical presentation of the 11 patients included repeated abdominal pain, jaundice, and fever. Conventional cholangiographies, such as percutaneous transhepatic cholangiography or endoscopic retrograde cholangiography, could not offer precise information about tumor location and extension because of abundant mucin in the biliary system. Using percutaneous transhepatic biliary drainage (PTBD) and percutaneous transhepatic cholangioscopy (PTCS), we were able to drain the mucin and determine precisely the cancer extension into intrahepatic segmental bile ducts. Based on these findings, various types of liver resection with or without extrahepatic bile duct resection were planned, and 10 patients obtained curative resection. The cumulative 5-year survival rate after curative resection was 78%. In patients with mucin-producing intrahepatic cholangiocarcinoma, PTBD and PTCS are important for evaluating the cancer extension. Rational surgery based on accurate preoperative diagnosis improved the prognosis of patients with this disease. PMID- 10512945 TI - Pancreatic solid-cystic-papillary tumor: clinicopathologic features in eight patients from Hong Kong and review of the literature. AB - Solid-cystic-papillary tumors (SCPTs) of the pancreas are rare. The clinicopathologic features and pathogenesis of these tumors have attracted a number of investigations, but the results remain unclear. We investigated the clinicopathologic data, immunohistochemical expression of the pan-endocrine markers, hormones, steroid receptors, and p53 overexpression in pancreatic SCPTs from eight Chinese patients (seven women, one man) collected over a 24-year period. They accounted for 2.5% of the primary pancreatic tumors. The tumors were seen in young women (mean age 27 years). They were large (mean size of resected tumor was 8.4 cm), benign, had solid and cystic areas, and were evenly distributed in the pancreas. The main differential diagnosis was pancreatic endocrine tumor. The tumors were negative for pan-endocrine markers, hormones, estrogen receptor, progesterone receptor, and p53. To date, 452 pancreatic SCPTs have been documented in the English literature. They occurred in patients of different ethnic groups, particularly in non-Caucasians. The tumors were frequently noted in young females. Uncommon cases of malignant pancreatic SCPTs were often found in older men and had indolent behavior. It was concluded that pancreatic SCPTs have distinct clinicopathologic characteristics. The present observations, together with a review of the literature suggests that overexpression of p53 or estrogen receptor is not important in the pathogenesis of pancreatic SCPTs. PMID- 10512946 TI - Oral urografin in postoperative small bowel obstruction. AB - The aim of our study was to determine whether Urografin has the potential to offer surgeons a way of differentiating complete from partial small bowel obstruction and whether partial small bowel obstruction can be treated nonoperatively. Altogether 116 patients who had postoperative small bowel obstructions without any toxic signs underwent Urografin studies. Urografin (40 ml) mixed with 40 ml of distilled water was administrated either orally or via nasogastric tube to each patient. Serial plain abdominal radiographs were taken 2, 4, and 8 hours later. A total of 74 patients (63.8%) whose contrast medium reached the colon within the first 8 hours were considered to have partial obstruction and were successfully treated with intravenous hydration and nasogastric decompression. The remaining 42 patients (36.2%) in whom the contrast medium failed to reach the colon within the first 8 hours were regarded as having complete obstruction, and 34 of those patients (81.0%) underwent surgery; 8 (19.0%) received conservative treatment. Adhesion bands with complete bowel obstruction were observed in all 34 patients (100.0%) during laparotomy. Regardless of the presence of an air-fluid level on a plain abdominal radiograph or abdominal pain, a liquid diet followed by a soft diet could be given to those patients whose Urografin emptied into the colon. All the patients with partial bowel obstruction were treated successfully with nonoperative methods. The presence of Urografin in the colon within 8 hours of ingestion as an indicator for nonoperative treatment had a sensitivity of 90.2%, a specificity of 100%, and an accuracy of 93. 1%. Urografin, a safe and reliable water-soluble contrast medium, can be used to differentiate partial intestinal obstruction from complete intestinal obstruction. Early oral intake was found to be a major advantage of Urografin use in this study, and the potential of Urografin use to shorten the period of conservative treatment for postoperative small bowel obstruction needs further investigation. PMID- 10512947 TI - Strategy for surgical management of ileocolonic anastomotic recurrence in Crohn's disease. AB - After resection for ileocecal or ileocolonic Crohn's disease anastomotic recurrence is common, and many patients require further surgery. This study reviews our overall experience of surgery for ileocolonic anastomotic recurrence of Crohn's disease so we can propose a strategy for management. A series of 109 patients who underwent surgery for anastomotic recurrence after ileocecal or ileocolonic resection for Crohn's disease between 1984 and 1997 were reviewed. Ileocolonic recurrence was treated by strictureplasty in 39 patients and resection in 70 (with sutured end-to-end anastomosis, 48; stapled side-to-side anastomosis, 22). Stapled anastomosis has been frequently used between 1995 and 1997. Short recurrence was mainly treated by strictureplasty, and long or perforating disease was resected. Coexisting small bowel disease was more common in the patients having strictureplasty. Septic complications (leak/fistula/abscess) related to the ileocolonic procedure occurred in 1 of 39 patients (3%) after strictureplasty, in 6 of 48 (13%) after resection with sutured anastomosis, and in none of 22 after resection with stapled anastomosis. The median duration of follow-up was 90 months after strictureplasty, 105 months after resection with sutured anastomosis, and 22 months after resection with stapled anastomosis. Altogether 18 of 39 patients (46%) after strictureplasty, 22 of 48 (46%) after resection with sutured anastomosis, and none of 22 after resection with stapled anastomosis required further surgery for suture line recurrence. In conclusion, strictureplasty is useful for short ileocolonic recurrence in patients with multifocal small bowel disease or previous extensive resection. Stapled side-to-side anastomosis was associated with a low incidence of complications, and early recurrence was not observed, although the duration of follow-up was short. PMID- 10512948 TI - Nerve-sparing surgery for advanced rectal cancer patients: special reference to Dukes C patients. AB - Several nerve-sparing operations for advanced rectal cancer that aim to preserve genitourinary function without compromising tumor clearance have been developed in Japan. The aim of this study was to evaluate the survival and local recurrence of these procedures in Dukes B and C patients. A total of 177 patients with advanced rectal cancer underwent curative nerve-sparing surgery (NSS) over the last 11 years; 52 were Dukes B patients and 54 were Dukes C. Altogether 36 had Dukes C1 and 18 had Dukes C2 tumors, 13 with lateral lymph node metastases, designated lateral LN(+). The 5-year survival rate was 92% for Dukes B, 67% for Dukes C1, and 39% for Dukes C2 patients: 11% for Dukes C2 patients with lateral LN(+). The local recurrence rate was 6% for Dukes B, 11% for Dukes C1, and 33% for Dukes C2 patients: 20% for the lateral LN(-) group and 39% for the lateral LN(+) group. Almost all of the patients undergoing NSS could micturate spontaneously, but preservation of sexual function was not as successful. Although there is no guarantee of preserving satisfactory sexual function, our NSS is an acceptable procedure for Dukes B, C1, and C2 patients without lateral lymph node metastases. PMID- 10512949 TI - Prognostic factors in patients with locally advanced rectal adenocarcinoma treated with preoperative radiotherapy and surgery. AB - Preoperative radiation therapy (PRT) prior to potential curative resection for rectal adenocarcinoma is not widely accepted. This report evaluates the prognostic factors affecting local recurrence and 5-year survival. This is a retrospective study of 214 patients with primary rectal adenocarcinoma treated from January 1986 to December 1994. A PRT dosage of 45 Gy in 20 fractions was administered to patients with clinically tethered or fixed tumors, and 4 to 8 weeks later surgery was performed (group I). Patients with clinically mobile tumors were treated by surgery alone (group II). There were 130 men and 84 women. The median age was 58 years (range 19-85 years). There were 111 patients in group I: 7 patients had no microscopic residual tumor, 80 had Dukes' A and B, and 24 had Dukes' C. There were 103 patients in group II: 70 patients were classified as Dukes' A and B and 33 as Dukes' C. The mean follow-up of the entire cohort was 62 months (range 2-132 months). Local recurrence was seen in 17% of patients in group I and 35% in group II (p = 0.002). Distant recurrence in patients with metastatic lymph nodes was seen in 79% of group I and in 34% of group II (p = 0.001). The favorable prognostic factors for local control were the administration of PRT and well differentiated cancer. The favorable prognostic factors for survival were age < 50 years and the absence of lymph node metastasis. The administration of PRT diminishes the risk of local recurrence. The presence of metastatic lymph nodes in the postirradiated specimen is an ominous prognostic factor for survival. Therefore such patients should be considered for adjuvant chemotherapy. PMID- 10512950 TI - Value of routine computed tomography in the preoperative assessment of abdominal aneurysm replacement. AB - This study compares the findings of conventional computed tomography (CT) and ultrasound (US) imaging in the preoperative evaluation of patients with abdominal aortic aneurysm (AAA). It also assesses the impact of preoperative CT-derived information on operative strategy. A prospective study was conducted of 96 patients who were considered for aortic aneurysm surgery, and the operative notes and US and CT reports were analyzed to assess correlation of findings and influence of CT on operative tactics. Agreement between CT and US in sizing the aneurysm was generally good. CT was more accurate than US for defining the upper and lower extent of the aneurysm (75% and 83%, respectively, with CT, compared to 47% and 41% with US), although it had a high false positive rate (48%) for juxtarenal disease. Its advantage over US in regard to showing other intraabdominal pathology was only marginal, and it predicted an inflammatory reaction in only two of the five cases. Its influence on operative strategy was minimal: Of the 25 cases where a juxtarenal aneurysm was predicted by CT, only 2 patients did not have surgery as a result. CT is a relatively expensive and time consuming procedure, and its ionizing radiation, however small, and potential side effects from contrast material hypersensitivity cannot be ignored. In light of the above findings, we suggest that CT scanning need not be routinely employed in the preoperative workup of elective aortic aneurysm repair but should be used only in selected cases. PMID- 10512951 TI - Female genital mutilation: A global bug that should not cross the millennium bridge. AB - Female genital mutilation (FGM) has been practiced worldwide, clothed under the tradocultural term "circumcision." Indications for its practice include ensuring virginity, securing fertility, securing the economic and social future of daughters, preventing the clitoris from growing long like the penis, and purely as a "tradition." Outlawed only in the United Kingdom, Sweden, and Belgium, no law forbids it in most other countries. Classified into four identified types, the current perpetrators are mainly quacks, but trained medical personnel still connive at and encourage FGM. Early complications include hemorrhage, urinary tract infection, septicemia, and tetanus. Late complications include infertility, apareunia, clitoral neuromas, and vesicovaginal fistula. Reasons for the ritual persisting include fear that legislation would force it underground and it will be performed in unsterile conditions, belief that it is racist to speak out against FGM, "tolerance" by health professionals, continued use of the term "female circumcision," lack of awareness of the culture of immigrants by the physicians in areas where FGM is not culturally practiced, and sporadic or uncommitted eradication efforts. We believe there is no reason for the continued practice of FGM. It should incur global abolition, the same way slave trade or Victorian chastity belts have done. We advocate that in medical communications the term "female genital mutilation" be used in place of "female circumcision." World leaders should include unacceptable cultural practices such as FGM in the "world summit" agenda. The year 1999 should be declared the year for global eradication of FGM. PMID- 10512952 TI - Historical evolution of limb amputation. AB - The amputation of a limb is one of the oldest surgical procedures. In the course of medical history operative techniques and surgical instruments have been improved continuously. As early as the first century Celsus described an amputation. A major step in the development of the operative technique was the introduction of an artery forceps by Pare during the sixteenth century. Nevertheless, due to a lack of analgesics and narcotics the operation had to take only a few minutes. Therefore the amputation was completed in one cut (i.e., detachment of the skin, muscles, and bone at the same level). This technique, known as "classic circular cut," was modified several times in the following period: to reduce suture tension Petit recommended that we transect the skin first and the muscles and bone more proximally ("two-stage circular cut," 1718), and Bromfield approved that the skin be cut first, the muscles more proximally and the bone most proximal ("three-stage circular cut," 1773). Lowdham (1679), Verduyn (1696), and Langenbeck (1810) changed the operative technique in that they used a soft-tissue flap to cover the bone without tension ("flap amputation"). PMID- 10512953 TI - Maintenance of gingival contour during prosthodontic procedures: a clinical report. PMID- 10512955 TI - Temporary nasal prosthesis rehabilitation: a clinical report. PMID- 10512954 TI - Dental implant placement and restoration in a mandibular ridge previously restored with hydroxyapatite augmentation and a dermal graft: a clinical report. PMID- 10512956 TI - Procedure for construction of a custom tracheostomal obturator: a clinical report. PMID- 10512957 TI - Two-year clinical evaluation of direct and indirect composite restorations in posterior teeth. AB - STATEMENT OF PROBLEM: Few long-term clinical studies have reported data of modern posterior composites as direct and indirect restorations. PURPOSE: This prospective, long-term clinical trial (1) evaluated direct and indirect composite restorations for clinical acceptability as posterior restoratives in single or multisurface carious teeth and (2) provided a survey on the 2-year results. MATERIAL AND METHODS: Nine dental students placed 88 composite restorations (Tetric, blend-a-lux, Pertac-Hybrid Unifil), 43 direct composite restorations and 45 indirect inlays, under the supervision of an experienced dentist. The first clinical evaluation was performed 11 to 13 months after placement by 2 other experienced dentists, using modified USPHS criteria. A second follow-up of 60 restorations took place within 20 to 26 months after placement. RESULTS: A total of 93% of indirect and 90% of direct composite restorations were assessed to be clinically excellent or acceptable. Two restorations (1 indirect composite inlay and 1 margin of a direct composite restoration) failed during the second year because of fracture. Indirect inlays demonstrated a significantly better "anatomic form of the surface" than direct composite restorations. Premolars revealed a significantly better margin integrity and postoperative symptoms than molars. CONCLUSION: Posterior composite restorations provided a satisfactory clinical performance over a 2-year period when placed by relatively inexperienced but supervised students. PMID- 10512958 TI - Comparison of resin-bonded prosthesis groove parallelism with the use of four tooth preparation methods. AB - STATEMENT OF PROBLEM: A precise preparation is required to develop resistance form resulting in mechanical stability of the framework for resin-bonded prostheses (RBPs). PURPOSE: The effects of 4 methods of tooth preparation (freehand, guiding pin, extraoral parallelometer, and intraoral parallelometer) on the deviation of proximal grooves from a preestablished path of insertion (guide planes) were investigated under clinical conditions. MATERIAL AND METHODS: Tooth preparation of proximal grooves was performed by 32 dentists on resin substitutes of posterior segments intraorally with a single test patient. A Latin square randomized cross-over design was selected as the experimental design. RESULTS: The significant least angular deviation of proximal grooves from path of insertion was achieved with an intraoral parallelometer (mean +/- SD 3.15 +/- 1.67 degrees). Compared with freehand tooth preparations (4.37 +/- 2. 11 degrees), neither use of a guiding pin (4.10 +/- 1.62 degrees) nor an extraoral parallelometer (5.06 +/- 2.33 degrees) improved the results. CONCLUSION: Divergence of guiding grooves from path of insertion was reduced with the use of an intra-oral parallelometer. This should improve mechanical stability of posterior RBPs. PMID- 10512959 TI - Prevalence of dental occlusal variables and intraarticular temporomandibular disorders: molar relationship, lateral guidance, and nonworking side contacts. AB - STATEMENT OF PROBLEM: The association between dental occlusion and the development of intraarticular temporomandibular disorders remains unclear. PURPOSE: This study evaluated the prevalence of molar relationship, lateral guidance and nonworking side contacts and intraarticular temporomandibular disorders. MATERIAL AND METHODS: Eighty-two asymptomatic volunteers and 263 symptomatic temporomandibular disorder (TMD) patients completed a subjective questionnaire that documented the absence of jaw pain, joint noise, locking, and a positive history for TMD. Participants also underwent clinical and dental examination for signs and symptoms commonly associated with TMD or internal derangements. RESULTS: The most prevalent molar relationship was Class I. Symptomatic patients had a higher prevalence of Class II, Division 1 relationships on the left side compared with the asymptomatic volunteers with normal joints. There was a higher prevalence of canine guidance (52.04%; P <.005) on the right side in the symptomatic patients with disk displacement (DD). Volunteers with normal joints had a higher prevalence of 1 or more nonworking side contacts compared with symptomatic patients with normal joints (P <.001) and symptomatic patients with DD (P <. 001). CONCLUSION: This study suggests there are no systematic dental occlusal differences that clearly separate symptomatic from asymptomatic patients. Results indicate that it is unclear as to the relationship of the 3 analyzed factors and of intraarticular TMDs. PMID- 10512961 TI - Clinical and radiographic evaluation of early loaded free-standing dental implants with various coatings in beagle dogs. AB - STATEMENT OF PROBLEM: Immediate loading of implants may be a predictable treatment alternative when cross-arch stabilization with a fixed provisional is observed. PURPOSE: This study investigated the effect of immediate masticatory loading on the stability of single-standing dental implants with 4 different surfaces. MATERIAL AND METHODS: A total of 40 solid screw implants (diameter 3.3 mm, length 8 mm) were placed in the mandibles of 4 beagle dogs. Test groups included 3 hydroxyapatite (HA) coatings of titanium plasma-sprayed (TPS) implants. Implants with TPS alone served as control. Gold crowns were inserted 2 days after implant placement and the dogs were immediately put on a hard food diet. Implants were followed for 6 months after loading. Clinical and radiographic assessments of implants were performed at time of crown insertion (baseline) and after 1, 3, and 6 months of loading. The Periotest instrument was used for mobility measurements and radiographs were obtained for evaluation of peri-implant radiolucency and measurement of crestal bone changes. RESULTS: Of 40 implants, 39 displayed no discernible mobility, corresponding to successful clinical function. Peri-implant radiolucencies were absent for all but the 1 mobile implant. The reduction in crestal bone levels adjacent to the implants between baseline and 6 months was statistically significant (P <.0001). No statistically significant differences in crestal bone level changes over time were found between the various coatings demonstrating the absence of a treatment effect initiated by the surface coatings. CONCLUSION: In this study in beagle dogs, immediate masticatory loading of single-standing dental implants did not jeopardize tissue integration, provided the implants had excellent primary stability. PMID- 10512962 TI - Influence of hex geometry and prosthetic table width on static and fatigue strength of dental implants. AB - STATEMENT OF PROBLEM: Component fracture and screw loosening are prevalent concerns of contemporary dental implants. PURPOSE: This laboratory investigation examined the influence of design factors such as the platform diameter and the hex height on the mechanical strength and quality of fit of the implant-abutment interface. MATERIAL AND METHODS: Static and cyclic compressive bending tests were conducted on 4 and 5 mm diameter bone density-based implants. SEM evaluation of the implant-abutment interface was also conducted to assess quality of fit between the mating components.Results. The 5 mm diameter implant was stronger in both static and fatigue conditions than the 4 mm diameter implants. A comparison of the results to published literature indicated that both implants were equal to or superior to alternative prosthetic connections in an identical testing configuration. CONCLUSION: Test results demonstrated the validity of wide diameter implants to reduce the likelihood of component fracture in contemporary dental implant systems. PMID- 10512960 TI - Randomized clinical trial comparing the efficacy of mandibular implant-supported overdentures and conventional dentures in diabetic patients. Part III: comparisons of patient satisfaction. AB - STATEMENT OF PROBLEM: There is insufficient evidence to indicate the functional superiority of mandibular implant-supported overdentures to justify their use in edentulous patients. PURPOSE: This study compared the benefits perceived by patients who received a new maxillary denture and a mandibular conventional denture (CD) and an implant-supported overdenture (IOD). METHOD: New maxillary and mandibular dentures were delivered to 89 diabetic denture wearers with clinically acceptable metabolic control who treated their diabetes either with insulin (IT) or without insulin (NIT). Of the 89 patients, 37 received maxillary and mandibular CDs and 52 received a maxillary CD and an IOD. Two questionnaires with categorical responses were used; the first contained 13 questions to ascertain a patient's absolute assessments of original dentures at entry and study dentures at 6- and 24-months after treatment completion; the second questionnaire had 11 questions that assessed the relative change perceived by patients with study dentures. Of the 78 patients who completed the posttreatment (PT) assessments at 6 months, 68 patients provided longitudinal data for questionnaire I and cross-sectional data for questionnaire II. In addition, 46 patients (18 CD and 28 IOD) also provided PT assessments at 24 months. RESULTS: Both mean scores and percentage distributions of longitudinal data for questionnaire I showed perceptual improvements with both types of study dentures. Improvements were higher in the IOD than in the CD group. Mean scores failed to show any significant differences between the 2 treatment groups. The only significant difference was found in the change in percentage distributions for perceptual chewing ability in favor of the IOD group. Even this advantage was lost at 24 months. With the comparative questionnaire, a higher percentage of patients in the IOD group than in the CD group perceived improvements with study dentures from their original dentures in chewing ability, chewing comfort, and denture security. However, mean differences were statistically significant in favor of the IOD group only for chewing ability and less difficulty to chew hard foods. CONCLUSION: The mandibular implant-supported overdenture offers same advantage in terms of perceived chewing function over the conventional denture. PMID- 10512963 TI - Ultraviolet radiation-induced color shifts occurring in oil-pigmented maxillofacial elastomers. AB - STATEMENT OF PROBLEM: Oil-based pigments are added to a maxillofacial prosthesis either as base colorants present within the elastomer or as surface tints that are painted on with an adhesive. Color stability of the pigments and pigmented prosthetic materials on exposure to ultraviolet radiation are unknown. PURPOSE: This study measured DeltaE* color changes caused by ultraviolet radiation for materials colored with 5 oil pigments, applied either as base colorants (intrinsic) or surface tints (extrinsic) to a silicone elastomer. MATERIAL AND METHODS: One of 5 oil pigments was added to polydimethyl siloxane disks to serve as a base colorant (0.2 weight percent present throughout a 2 mm thick disk) or as a concentrated surface tint (2.0 weight percent concentrated in upper 0.3 mm thickness). Pigmented disks, along with pigment-only and elastomer-only control disks, were exposed to ultraviolet radiation for 400, 600 and 1800 hours. DeltaE* color changes were measured at baseline and for each time interval. RESULTS: Control samples underwent minimum color changes after 1800 hours (DeltaE* 2.0), that ranged from 2.13 to 3.98. Silskin 2000 showed greater color differences (P <.05) compared with Elastosil M3500 and Ideal, which showed similar behavior (P >.05). CONCLUSION: Outdoor exposure of silicone facial elastomers for 1 year resulted in visually detectable color differences. Duration of exposure and silicone elastomer were significant factors that affected color stability. Silskin 2000 was significantly less color stable than Elastosil M3500 and Ideal, which were not statistically significantly different from each other. PMID- 10512965 TI - Topographical characteristics and shear bond strength of tooth surfaces cut with a laser-powered hydrokinetic system. AB - STATEMENT OF PROBLEM: Erbium lasers, specifically Er;YAG and Er, Cr;YSGG that emit in the near red wavelengths, cut both enamel and dentine. Bonding to these cut surfaces with composites has not been assessed for all laser systems. PURPOSE: This investigation assessed the shear bond strength of composite bonded to tooth structure treated with an Er,Cr;YSGG-powered hydrokinetic system (HKS, Millennium system) and then was compared with surfaces treated with a carbide bur. MATERIAL AND METHODS: Extracted human molars were cut into enamel and dentin with both systems. Nonetched and acid-etched subgroups were evaluated. Shear bond strength was measured with an Instron test machine with a knife-edge loading head. In addition, SEMs were evaluated. RESULTS: There were no significant differences in shear bond strength between etched bur cut (23.3 +/- 2.5 MPa), etched laser-cut enamel (23.7 +/- 4.5 MPa), and nonetched laser-cut enamel (20.5 +/- 2.8 MPa). For nonetched enamel, bond strength values for laser-cut surfaces were significantly higher than the bur-cut surfaces (8.7 +/- 4.3 MPa). Bond strength differences for dentin between bur (14.3 +/- 1.7 MPa) and laser cuts (11.5 +/- 4.3 MPa) were not significant (P =.03). SEM revealed that laser cutting of enamel did not cause formation of a smear layer. CONCLUSION: There were no significant differences in shear bond strength between etched bur-cut, etched laser-cut, and nonetched laser-cut enamel. With nonetched enamel, bond strength values for nonetched laser-cut surfaces were significantly higher than for the bur-cut surfaces. No bond strength differences between bur and laser cutting existed for dentin. Similar topography was observed for bur and laser prepared surfaces of etched enamel and nonetched dentin. PMID- 10512966 TI - Compressibility of two polyvinyl siloxane interocclusal record materials and its effect on mounted cast relationships. AB - STATEMENT OF PROBLEM: Addition silicones (polyvinyl siloxanes) are universally accepted as accurate and stable impression materials. They have also gained popularity as interocclusal record materials. However, it has not been defined if it is possible to work with polyvinyl siloxanes without changing the recorded maxillomandibular relations. PURPOSE: This study examined the compressibility of 2 addition silicones as interocclusal record materials, analyzing the changes of maxillomandibular relations at the condyle region when different compressive forces are used to stabilize articulated casts. MATERIAL AND METHODS: Sixteen interocclusal records, obtained from the same patient (8 of each polyvinyl siloxane, Blu-Mousse, Fast Set), were interposed between the patient casts in a new measuring system obtaining 48 curves of load versus maxillomandibular positional changes in 3 axes (x, y, z). These curves were compared with curves obtained with the casts in maximum intercuspation without interocclusal records (reference curves). Analysis of variance was used to compare maxillomandibular positional changes among the 3 groups (n = 48 each): Blu-Mousse, Fast Set, and control group or maximum intercuspation without interocclusal record. RESULTS: There was no significant change in maxillomandibular relations when forces up to 1 kgf were applied to stabilize the casts related by means of Blu-Mousse and Fast Set addition silicone interocclusal records. CONCLUSION: It is possible to use these polyvinyl siloxanes as interocclusal record materials without changing the recorded maxillomandibular relations. PMID- 10512967 TI - Effect of sandblasting and silicoating on bond strength of polymer-glass composite to cast titanium. AB - STATEMENT OF PROBLEM: There is little information regarding the mechanical and chemical retention of polymer-glass composite to cast titanium. PURPOSE: This study examined whether sandblasting in conjunction with silicoating improves the bond strength of the polymer-glass composite to cast titanium. MATERIALS AND METHODS: Disk patterns (10 mm in diameter, 2.5 mm thick) were cast with commercially pure titanium (CP Ti) and Type IV gold alloy. Three pretreatments were applied: 50 microm Al(2)O(3) sandblasting (50 SB), 250 microm Al(2)O(3) sandblasting (250 SB), and 600-grit SiC paper polishing (600 SiC). After surface preparation, the Siloc system (silicoating) was applied on the disks. The 50 SB specimens without Siloc system were also prepared as controls. Then sticky tape with a circular hole (4.76 mm diameter) was placed onto the disk to define the bonding area. Artglass (polymer-glass) opaque, dentin, and enamel composites were applied using Teflon matrices and then light-polymerized. Shear bond strength (n = 8) was determined at a crosshead speed of 5 mm/min. Results were analyzed statistically with 2-way ANOVA and the Tukey-Kramer test (alpha=.05). RESULTS: The Siloc system significantly (P <.05) improved the mean shear bond strength of Artglass to both metals in the 50 SB specimens. Statistical differences (P <.05) in shear bond strength were found among surface treatments for the silicoated CP Ti specimens, in which 250 SB specimens yielded the greatest bond strength. The Type IV specimens treated with Siloc system showed no significant differences in shear bond strength between the 50 SB and the 250 SB specimens. CONCLUSION: Sandblasting with coarser alumina particles in conjunction with silicoating significantly enhanced bond strength of polymer-glass composite to cast titanium. PMID- 10512968 TI - Effects of surface finish and fatigue testing on the fracture strength of CAD-CAM and pressed-ceramic crowns. AB - STATEMENT OF PROBLEM: All-ceramic molar crowns can be fabricated with CAD-CAM or laboratory methods with different materials, and a polished or oven-glazed surface. PURPOSE: This in vitro study determined the fracture strength of various all-ceramic crowns, with and without prior cyclic loading. MATERIAL AND METHODS: Standardized molar crowns were fabricated with a CAD-CAM machine (Cerec 2), software with machinable ceramic materials (Vita Mark II and ProCAD), and also conventional heat-pressed IPS Empress crowns fabricated at 2 dental laboratories. Groups of 40 crowns of each material were manufactured with either a polished or an oven-glazed surface finish. Cyclic loading that simulated oral conditions were performed on half of each group. Afterward, all crowns were loaded until catastrophic failure.Results. Fracture loads of the polished ProCAD crowns without prior cyclic loading was 2120 +/- 231 N, significantly higher than that of the polished Vita Mark II crowns (1905 +/- 235 N), but was not significantly different from the strength of 2 laboratory-fabricated Empress crowns. Oven glazing of ProCAD crowns improved the fracture strength significantly, up to 2254 +/- 186 N. Prior cyclic loading decreased the strength of all tested crowns significantly, but the reduction was less for the Cerec crowns than the Empress crowns. CONCLUSION: Cerec ProCAD crowns demonstrated significantly greater strength than the Vita Mark II crowns, better resistance to cyclic loading and lower failure probability than the laboratory-fabricated IPS Empress crowns. Prior cyclic loading significantly reduced the strength of all-ceramic crowns, but had less effect on Cerec crowns than on the IPS Empress crowns. Oven-glazing of ProCAD crowns resulted in significantly higher strength and higher resistance to cyclic loading than surface polishing. PMID- 10512969 TI - Three-body wear associated with three ceramics and enamel. AB - STATEMENT OF PROBLEM: Wear of human enamel is a clinical concern whenever opposing teeth need to be restored using ceramic restorations. PURPOSE: This in vitro study investigated wear of human enamel and 3 dental ceramics: a conventional porcelain (Vitadur Alpha), a low-fusing hydrothermal ceramic (Duceram-LFC), and a machinable ceramic (Vita Mark II) in a 3-body wear test. MATERIAL AND METHODS: Thirty pairs of tooth-ceramic specimens were tested in a dental wear machine, under a standard load (40 N), rate (80 cycles/minute), and for 25,000 cycles in a simulated food slurry medium. Amount of wear was determined by measuring the height loss of the tooth and depth of wear track of the ceramic materials. Analysis of variance (ANOVA) was used to analyze the data, followed by Bonferroni multiple comparisons method to produce sets of simultaneous 95% confidence intervals. RESULTS: ANOVA revealed significant differences between the groups for both enamel wear (P =. 002) and ceramic wear (P <.001). Further comparisons (95% CI significance level) revealed that the difference in enamel wear produced by Alpha porcelain and Duceram-LFC ceramic material was not statistically significant, whereas that produced by Vita Mark II ceramic was significantly less. Vita Mark II ceramic was significantly more resistant to wear than Alpha porcelain and Duceram-LFC ceramic. Furthermore, Alpha porcelain was significantly more resistant to wear than Duceram-LFC ceramic. CONCLUSION: The abrasiveness of Alpha porcelain and Duceram-LFC ceramic was similar, yet both were significantly more abrasive than Vita Mark II ceramic. In addition, Vita Mark II was the most wear-resistant ceramic and Duceram-LFC ceramic the least resistant. PMID- 10512970 TI - Fabricating auricular prostheses using three-dimensional soft tissue models. AB - This article describes a method for fabricating an auricular prosthesis. This procedure uses the contours of the soft tissue surface from computerized tomography scans to fabricate a computer-generated, side-inverted 3-dimensional soft tissue model from a solid block of polyurethane using an Endoplan milling machine. The resultant 3-dimensional soft tissue model can then be used as the basis for a wax sculpture. This procedure facilitates the planning of the prosthesis; symmetrical modeling, especially for large, hemifacial defects; and the impression, which can be made on the model itself. PMID- 10512971 TI - Fabrication of a stable record base for severe soft tissue undercuts in the edentulous patient. PMID- 10512972 TI - The CrescoTi Precision method: description of a simplified method to fabricate titanium superstructures with passive fit to osseointegrated implants. AB - Casting distortion and inadequate handling in the dental laboratory are 2 factors that can cause misfit between a cast framework and the implant analogs in the master cast. This article describes the CrescoTi Precision procedure, which has been developed for correction of misfit between cast titanium frameworks and supporting dental implants. The simplicity and accuracy of the procedure appear to demonstrate technical progress toward obtaining optimal precision of fit of the superstructure components, thereby eliminating stress forces that may be transformed to the peri-implant bone. PMID- 10512973 TI - Replacement of a clip in one session. AB - Fracture of clips in implant-retained overdenture therapy is one of the most reported problems. This article describes an indirect method for repair that is reliable and can be carried out in 1 session. PMID- 10512974 TI - Method of placing personal identification on acetate mouth guards. PMID- 10512975 TI - Crustacean frequenins: molecular cloning and differential localization at neuromuscular junctions. AB - Crustacean muscles are innervated by phasic and tonic motor neurons that display differential physiology and have morphologically distinct synaptic terminals. Phasic motor neurons release much more transmitter per impulse and have filiform terminals, whereas tonic motor neurons release less transmitter and have larger terminals with prominent varicosities. Using an antibody raised against Drosophila frequenin (frq), a calcium-binding protein that enhances transmitter release in Drosophila synaptic terminals, we found that frq-like immunoreactivity is prominent in many of the phasic, but not tonic nerve endings of crayfish motor neurons. In contrast, synapsin- and dynamin-like immunoreactivities are strongly expressed in both types of terminal. The immunocytochemical findings strongly suggested the presence of an frq-like molecule in crayfish, and its differential expression indicated a possible modulatory role in transmitter release. Therefore, we cloned the cDNA sequences for the crayfish and lobster homologues of Drosophila frq. Crustacean frequenins are very similar in sequence to their Drosophila counterpart, and calcium-binding regions (EF hands) are conserved. The widespread occurrence of frq-like molecules and their differential localization in crayfish motor neurons indicate a significant role in physiology or development of these neurons. PMID- 10512976 TI - Androgen-metabolizing enzymes show region-specific changes across the breeding season in the brain of a wild songbird. AB - The Lapland longspur (Calcarius lapponicus) is an arctic-breeding songbird that shows rapid behavioral changes during a short breeding season. Changes in plasma testosterone (T) in the spring are correlated with singing but not territorial aggression in males. Also, T treatment increases song but not aggression in this species. In contrast, in temperate-zone breeders, song and aggression are highly correlated, and both increase after T treatment. We asked whether regional or temporal differences in androgen-metabolizing enzymes in the longspur brain explain hormone-behavior patterns in this species. We measured the activities of aromatase, 5alpha-reductase and 5beta-reductase in free-living longspur males. Aromatase and 5alpha-reductase convert T into the active steroids 17beta estradiol (E(2)) and 5alpha-dihydrotestosterone (5alpha-DHT), respectively. 5beta Reductase deactivates T via conversion to 5beta-DHT, an inactive steroid. We examined seven brain regions at three stages in the breeding season. Overall, aromatase activity was high in the hypothalamus, hippocampus, and ventromedial telencephalon (containing nucleus taeniae, the avian homologue to the amygdala). 5beta-Reductase activity was high throughout the telencephalon. Activities of all three enzymes changed over time in a region-specific manner. In particular, aromatase activity in the rostral hypothalamus was decreased late in the breeding season, which may explain why T treatment at this time does not increase aggression. Changes in 5beta-reductase do not explain the effects of plasma T on aggressive behavior. PMID- 10512977 TI - Genetic analysis of the Drosophila ellipsoid body neuropil: organization and development of the central complex. AB - The central complex is an important center for higher-order brain function in insects. It is an intricate neuropil composed of four substructures. Each substructure contains repeated neuronal elements which are connected by processes such that topography is maintained. Although the neuronal architecture has been described in several insects and the behavioral role investigated in various experiments, the exact function of this neuropil has proven elusive. To describe the architecture of the central complex, we study 15 enhancer-trap lines that label various ellipsoid body neuron types. We find evidence for restriction of gene expression that is correlated with specific neuronal types: such correlations suggest functional classifications as well. We show that some enhancer-trap patterns reveal a single ellipsoid body neuron type, while others label multiple types. We describe the development of the ellipsoid body neuropil in wild-type animals and propose developmental mechanisms based on animals displaying structural mutations of this neuropil. The experiments performed here demonstrate the degree of resolution possible from the analysis of enhancer-trap lines and form a useful library of tools for future structure/function studies of the ellipsoid body. PMID- 10512978 TI - Activity-dependent regulation of axonal growth: posttranscriptional control of the GAP-43 gene by the NMDA receptor in developing hippocampus. AB - The intricate circuitry of the nervous system has been shown to be refined by activity-dependent processes often involving the glutamate N-methyl-D-aspartate (NMDA) receptor. NMDA receptor activity has been directly associated with axonal growth during development and in adult models of synaptic plasticity. The axonal growth-associated protein GAP-43 has been involved in the same processes as the NMDA receptor, but a direct link between the two has never been demonstrated in vivo. It is attractive to think that the NMDA receptor may regulate axonal growth through GAP-43. We tested this idea in outgrowing axons of hippocampal granule cells, the mossy fibers. Granule cells normally only express GAP-43 in an organized outside-in manner during a restricted period in postnatal development paralleling the pattern of axonal extension. Here, we show that during postnatal development in a transgenic mouse bearing a GAP-43 promoter/lacZ reporter construct, granule cells also display an outside-in pattern of promoter activation as indexed by transgene expression (PATE). In fact, PATE precedes axonal outgrowth with temporospatial fidelity. Since PATE deactivates on growth termination, the promoter may function as a switch for an intrinsic program of regulated axonal growth. The NMDA receptor antagonist MK-801 administered within a restricted time frame (4-8 days) results in a decrease in the extent and intensity of mossy fiber staining. While levels of GAP-43 mRNA are significantly reduced in granule cells, GAP-43 PATE is not. The level of GAP-43 expression and axonal growth during development appears to be dually controlled by a transcriptional program that is activity-independent and by a posttranscriptional mechanism that is activity-dependent and NMDA mediated. PMID- 10512979 TI - Prostaglandin E(2) stimulates glutamate receptor-dependent astrocyte neuromodulation in cultured hippocampal cells. AB - Recent Ca(2+) imaging studies in cell culture and in situ have shown that Ca(2+) elevations in astrocytes stimulate glutamate release and increase neuronal Ca(2+) levels, and that this astrocyte-neuron signaling can be stimulated by prostaglandin E(2) (PGE(2)). We investigated the electrophysiological consequences of the PGE(2)-mediated astrocyte-neuron signaling using whole-cell recordings on cultured rat hippocampal cells. Focal application of PGE(2) to astrocytes evoked a Ca(2+) elevation in the stimulated cell by mobilizing internal Ca(2+) stores, which further propagated as a Ca(2+) wave to neighboring astrocytes. Whole-cell recordings from neurons revealed that PGE(2) evoked a slow inward current in neurons adjacent to astrocytes. This neuronal response required the presence of an astrocyte Ca(2+) wave and was mediated through both N-methyl-D aspartate (NMDA) and non-NMDA glutamate receptors. Taken together with previous studies, these data demonstrate that PGE(2)-evoked Ca(2+) elevations in astrocyte cause the release of glutamate which activates neuronal ionotropic receptors. PMID- 10512980 TI - Theoretical analysis of gradient detection by growth cones. AB - Gradients of diffusible and substrate-bound molecules play an important role in guiding axons to appropriate targets in the developing nervous system. Although some of the molecules involved have recently been identified, little is known about the physical mechanisms by which growth cones sense gradients. This article applies the seminal Berg and Purcell (1977) model of gradient sensing to this problem. The model provides estimates for the statistical fluctuations in the measurement of concentration by a small sensing device. By assuming that gradient detection consists of the comparison of concentrations at two spatially or temporally separated points, the model therefore provides an estimate for the steepness of gradient that can be detected as a function of physiological parameters. The model makes the following specific predictions. (a) It is more likely that growth cones use a spatial rather than temporal sensing strategy. (b) Growth cone sensitivity increases with the concentration of ligand, the speed of ligand diffusion, the size of the growth cone, and the time over which it averages the gradient signal. (c) The minimum detectable gradient steepness for growth cones is roughly in the range 1-10%. (d) This value varies depending on whether a bound or freely diffusing ligand is being sensed, and on whether the sensing occurs in three or two dimensions. The model also makes predictions concerning the role of filopodia in gradient detection. PMID- 10512981 TI - En passant synaptic varicosities form directly from growth cones by transient cessation of growth cone advance but not of actin-based motility. AB - Formation of terminal synapses at sites such as the neuromuscular junction involves transformation of the motile growth cone into the nonmotile synaptic terminal. However, transformation does not need to be the mechanism when a neurite forms multiple widely spaced synaptic varicosities along a target in an en passant configuration. Synaptic varicosities could form here by specialization of the neurite after the growth cone has advanced past the site. We examined this issue by using cocultures of identified sensory (SN) and motor (L7) neurons from Aplysia. Living SNs were labeled with fluorescent dye and their neurites were observed at high resolution every few minutes growing along the axon of L7, allowing a fine-grained analysis of the behavior of the growth cone at the sites of synapse formation. All varicosities whose formation was observed indeed developed from the growth cone. Sensory varicosities were shown by electron microscopy to contain features characteristic of active zones for transmitter release within a day of their formation on the motor axon. Growth cone advance slowed or stopped transiently during varicosity formation, but the motile activity of the peripheral region of the growth cone (veils and filopodia) was maintained. These results suggest that target "stop signals" involved in the formation of synapses, at least of the en passant variety, may be of a different type from the growth inhibitory molecules, such as the collapsins, which guide axons to their targets. PMID- 10512982 TI - Sex differences in cell migration in the preoptic area/anterior hypothalamus of mice. AB - The preoptic area/anterior hypothalamus (POA/AH) sits as a boundary region rostral to the classical diencephalic hypothalamus and ventral to the telencephalic septal region. Numerous studies have pointed to the region's importance for sex-dependent functions. Previous studies suggested that migratory guidance cues within this region might be particularly unique in their diversity. To better understand the early development and differentiation of the POA/AH, cytoarchitectural, birthdate, immunocytochemical, and cell migration studies were conducted in vivo and in vitro using embryonic C57BL/6J mice. A medial preoptic nucleus became discernible using Nissl stain in males and females between embryonic days (E) E15 and E17. Cells containing immunoreactive estrogen receptor alpha were detected in the POA/AH by E13, and increased in number with age in both sexes. From E15 to E17, examination of the radial glial fiber pattern by immunocytochemistry confirmed the presence of dual orientations for migratory guidance ventral to the anterior commissure (medial-lateral and dorsal-ventral) and uniform orientation more caudally (medial-lateral). Video microscopy studies followed the migration of DiI-labeled cells in coronal 250-microm brain slices from E15 mice maintained in serum-free media for 1-3 days. Analyses showed significant migration along a dorsal-ventral orientation in addition to medial lateral. The video analyses showed significantly more medial-lateral migration in males than females in the caudal POA/AH. In vivo, changes in the distribution of cells labeled by the mitotic indicator bromodeoxyuridine (BrdU) suggested their progressive migration through the POA/AH. BrdU analyses also indicated significant movement from dorsal to ventral regions ventral to the anterior commissure. The significant dorsal-ventral migration of cells in the POA/AH provides additional support for the notion that the region integrates developmental information from both telencephalic and diencephalic compartments. The sex difference in the orientation of migration of cells in the caudal POA/AH suggests one locus for the influence of gonadal steroids in the embryonic mouse forebrain. PMID- 10512983 TI - Age-dependent failure of axon regeneration in organotypic culture of gerbil auditory midbrain. AB - Inferior colliculus (IC) slice cultures from postnatal (P) day 6-8 gerbils exhibit axonal regeneration across a lesion site, and these regrowing processes can form synapses. To determine whether regenerative capacity is lost in older tissue, as occurs in vivo, slices from P12-21-day animals were grown under similar conditions. While these cultures displayed a near complete loss of neurons over 6 days in vitro, glutamate receptor antagonists (AP5 and/or CNQX) significantly enhanced survival, particularly at P12-15. In contrast, several growth factors or high potassium did not improve neuron survival. Therefore, axonal regeneration was assessed following complete transection of the commissure in AP5/CNQX-treated IC cultures from P12 animals. Neurofilament staining revealed that transected commissural axons survived for 6 days in vitro, but only a few processes crossed the lesion site and these axons did not extend into the contralateral lobe. In contrast, there was robust axonal sprouting and growth within one lobe of the IC, remote from the lesion site. When P6 and P12 tissue was explanted onto a coated substrate, the P6 axons grew onto the substrate, but the P12 axons were seemingly prevented from reaching the substrate by a veil of nonneuronal cells. Coculture of the IC and one of its afferent populations, the lateral superior olive, provided a similar finding, indicating that failure to regenerate was a general property at the age examined. These data show that neuron survival is not sufficient to permit axon regeneration at P12, and suggest that P12 lesion sites manufacture a prohibitive substrate since process outgrowth is blocked specifically at the commissure transection. PMID- 10512984 TI - Pax-2 expression defines a subset of GABAergic interneurons and their precursors in the developing murine cerebellum. AB - Pax-2 is a paired box transcription factor expressed in several regions of the developing mammalian central nervous system. First found in the midbrain/hindbrain region, Pax-2 expression is later found in the cerebellum, hindbrain, and spinal cord. We have examined the expression pattern of Pax-2 from embryonic day 12 (E12) through postnatal day 35 (P35) using immunohistochemistry and in situ hybridization. Expression of Pax-2 is found in scattered cells of the cerebellar ventricular zone at E13. Pax-2-expressing cells migrate away from this germinative center to positions in the deep cerebellar nuclei (DCN), internal granule cell layer, molecular layer, and folial white-matter tracts of the cerebellum. Immunocytochemistry of both tissue sections and primary dissociated cultures demonstrates that Pax-2 is expressed by cells of a neuronal lineage, but not by cells of either an astrocytic or oligodendrocytic lineage. Specifically, the presence of Pax-2 identifies the entire population of gamma-aminobutyric acid (GABA)ergic interneurons in the cerebellar cortex (Golgi II, basket and stellate cells) and in the DCN. Bromodeoxyuridase labeling and 4',6-diamino-2-phenylindole (DAPI) staining of cells in M-phase reveals that Pax-2-expressing cells in the folial white-matter tracts of the cerebellum constitute an actively dividing population. We propose that these cells are migratory precursors of the molecular layer interneurons (basket and stellate cells). Our data suggest that the role of Pax-2 in cerebellar development changes after E12, shifting from the specification of an anatomical field to the marking of a specific class of cells. Our findings also suggest a previously uncharacterized relationship among GABAergic interneurons found posterior to the midbrain. Finally, our data support the hypothesis that the basket and stellate cells arise from neuronally restricted, migratory precursors located in the early postnatal cerebellar white matter. PMID- 10512985 TI - Neurotrophic factor regulation of developing avian oculomotor neurons: differential effects of BDNF and GDNF. AB - Neurotrophic factors support the development of motoneurons by several possible mechanisms. Neurotrophins may act as target-derived factors or as afferent factors derived from the central nervous system (CNS) or sensory ganglia. We tested whether brain-derived neurotrophic factor (BDNF), neurotrophin 3 (NT-3), neurotrophin 4 (NT-4), and glial cell line-derived neurotrophic factor (GDNF) may be target-derived factors for neurons in the oculomotor (MIII) or trochlear (MIV) nucleus in chick embryos. Radio-iodinated BDNF, NT-3, NT-4, and GDNF accumulated in oculomotor neurons via retrograde axonal transport when the trophic factors were applied to the target. Systemic GDNF rescued oculomotor neurons from developmental cell death, while BDNF and NT-3 had no effect. BDNF enhanced neurite outgrowth from explants of MIII and MIV nuclei (identified by retrograde labeling in ovo with the fluorescent tracer DiI), while GDNF, NT-3, and NT-4 had no effect. The oculomotor neurons were immunoreactive for BDNF and the BDNF receptors p75(NTR) and trkB. To determine whether BDNF may be derived from its target or may act as an autocrine or paracrine factor, in situ hybridization and deprivation studies were performed. BDNF mRNA expression was detected in eye muscles, but not in CNS sources of afferent innervation to MIII, or the oculomotor complex itself. Injection of trkB fusion proteins in the eye muscle reduced BDNF immunoreactivity in the innervating motoneurons. These data indicate that BDNF trophic support for the oculomotor neurons was derived from their target. PMID- 10512986 TI - In memoriam: Gordon Sydney Anderson Birnie Stewart (1951-1999). PMID- 10512987 TI - Effect of linear polarized near-infrared ray irradiation on the chemiluminescence of human neutrophils and serum opsonic activity. AB - The purpose of this study was to investigate the in vitro effects of linear polarized near-infrared ray irradiation on neutrophil chemiluminescence (CL) and serum opsonic activity. We used luminol- and lucigenin-dependent CL to detect the affected reactive oxygen species production process of human neutrophils and measured serum opsonic activity based on luminol-dependent CL. The linear polarized near-infrared ray irradiation suppressed a maximum light emission (peak height) of luminol- and lucigenin-dependent CL in a dose-dependent manner. The findings suggested that the linear polarized near-infrared ray irradiation suppressed the superoxide anion and hypochlorite production of human neutrophils. The serum opsonic activity was decreased by linear polarized near-infrared ray irradiation, and this suppressive effect might be caused by inhibiting the activation of the classical and alternative complement pathway. Therefore, it is suggested that near-infrared ray irradiation may have an inhibitory effect against chronic pain via reduction of reactive oxygen species production and opsonic activity. PMID- 10512988 TI - Low-level chemiluminescence of N-beta-alanyl-L-histidine (L-carnosine). AB - Oxidized N-beta-Ala-L-His (L-carnosine) emitted low-level CL. The CL specificity was shown by experiments with L-carnosine from six separate vendors, several L carnosine-like compounds, and nine different oxidizers. Purity of L-carnosine samples was analysed by RP-HPLC-MS, (1)H-NMR, MALDI-TOF-MS and ESI-MS. L Carnosine CL magnitude varied with source; consequently, detection sensitivity was 5-100 nmol. CL of L-anserine (N-beta-Ala-1-methyl-L-His) was equal to or less than L-carnosine, depending upon oxidizer. H(5)IO(6) (2 mmol/L) in 11 mmol/L NaOH or 20 mmol/L K(3)Fe(CN)(6) + 10 mmol/L H(2)O(2) in 100 mmol/L NaOH were oxidizers of choice. Scavengers of (.)OH(-) radical quenched CL. Kinetic studies revealed a bi-phasic CL comprising a short-lived (<1 s) 'flash' and then prolonged ( approximately 2000 s) 'glow'. A structural basis and mechanism of L-carnosine CL are discussed. L-Carnosine CL could be useful for monitoring its level in biological samples. PMID- 10512989 TI - Improved extraction and assay of mycobacterial ATP for rapid drug susceptibility testing. AB - The firefly luciferase assay of ATP is a rapid and convenient technique for monitoring growth of mycobacteria. The time needed to obtain a drug susceptibility pattern can be reduced to less than 1 week as compared to 4 weeks with conventional methods. The ATP assay is simple and reliable. However, the extraction of bacterial ATP is not a trivial problem. Lysing the cells will immediately activate ATP-degrading enzyme systems. The extractant must therefore lyse the cells and simultaneously inactivate ATP-degrading enzyme systems. Only by comparing the ATP yields obtained with different extractants we will know something about the intracellular ATP level. In the present study various extractants were compared for the extraction of ATP from Mycobacterium bovis (BCG) cultures. Dodecyl trimethyl ammonium bromide (DTAB) in Tris-buffer with EDTA resulted at 100 degrees C in an ATP yield that was approximately twice as high as the same buffer without DTAB. The optimum temperature was 80-100 degrees C. With the optimized extraction procedure the coefficient of variation for the entire assay of ATP in BCG cultures was 5%. The analytical interference from DTAB with the firefly reaction was obviated by neutralization with alpha-cyclodextrin, making it possible to increase the sensitivity by assaying 0.5 mL rather than 0.01 mL extract. PMID- 10512990 TI - Analysis of river water by bioluminescent biotests. AB - The bacterial bioluminescence has high sensitivity to the action of various inhibitors of biological activity. The lyophilized luminous bacteria Photobacterium phosphoreum (Microbiosensor B17 677F) and luminous strain Escherichia coli (Microbiosensor EC) from the Culture Collection IBSO were used to create bioluminescent biotests. They have been applied in ecological monitoring to determine the overall toxicity of the Yenisei and Angara Rivers and some water sources of Altai Territory. As a rule the heaviest pollution of water in studied rivers was registered near cities and settlements. The luminous bacteria biotests are simple and convenient in work, standardized and quantitative, have rapid response to actions of different substances and high sensitivity to environmental pollutants. It takes less than 30 min to do the biotest (the other biotests take 48--96 h). PMID- 10512991 TI - Transfer of xenobiotics through cell membranes of luminous bacteria. AB - The influence of some chemical substances on luminous bacteria was studied to elucidate the interrelation between the xenobiotics action on bacterial luminescence and cell ultrastructure. Such substances as quinones, phenols, chlorides of heavy metals (in concentrations of substances inhibiting luminescence by 50%) resulted in damaging effects upon bacteria: a lot of cells had damage of membranes due to changes in their permeability. It was found that the high concentration of EDTA and toluene decreased the luminescence and caused the condensation of DNA-fibrils and the cell damage after long-term and short term action. The low concentration of EDTA and toluene did not decrease the bacterial luminescence; the noticeable damage of cell membranes did not take place during short-term treatment. However, the long action of these substances changed the membrane permeability resulting in increased sensitivity of bacterial luminescence to some toxic substances. PMID- 10512992 TI - Assays using digital fluorescence: 1985-1998 AB - Luminescence continues to provide comprehensive literature surveys which will be published in most issues. These are a continuation of the literature surveys begun in 1986 in the Journal of Bioluminescence and Chemiluminescence which, up until 1998, encompassed more than 6000 references cited by year or specialized topic. With this newly named journal these searches are expanding to reflect the journal's wider scope. In future we will cover all fundamental and applied aspects of biological and chemical luminescence and include not only bioluminescence and chemiluminescence but also fluorescence, time resolved fluorescence, electrochemiluminescence, phosphorescence, sonoluminescence, lyoluminescence and triboluminescence. The compilers would be pleased to receive any comments from the readership. Contact by e-mail: L.J. Kricka: larry_kricka@path1a.med.upenn.edu or P.E. Stanley: Stanley@LUMIWEB.COM Copyright 1999 John Wiley & Sons, Ltd. PMID- 10512993 TI - The natural history of CADASIL. PMID- 10512994 TI - Small but quantifiable patient preference for MRA versus catheter angiography. PMID- 10512995 TI - Poststroke sexual dysfunction and quality of life. PMID- 10512996 TI - Risk assessment and anticoagulation in atrial fibrillation in the elderly: malpractice or accuracy? PMID- 10512997 TI - Usefulness of transcranial color-coded sonography in the diagnosis of cerebral vasospasm. PMID- 10512998 TI - Should we screen for familial intracranial aneurysm? PMID- 10512999 TI - Increased intensity of physiotherapy after stroke. PMID- 10513000 TI - Prevalence of carotid disease in patients with coronary artery stenosis. PMID- 10513001 TI - Plasma homocyst(e)ine concentrations in cerebrovascular disease. PMID- 10513002 TI - Antiplatelet therapy in acute cerebral ischaemia. PMID- 10513003 TI - Poststroke sexual function. PMID- 10513004 TI - Liquid chromatographic determination of fenbendazole residues in pig tissues after treatment with medicated feed. AB - Fenbendazole (FBZ) is an anthelmintic widely used in farm animals to treat parasitic infestations. In pigs, it is administered in the food. The aim of this study was to validate an analytical method for the determination of FBZ and its metabolites in pig tissues. This method is based on oxidation of FBZ and its sulfoxide metabolite to the sulfone metabolite (FBZSO2). The limit of quantitation for this method is 20 ng FBZSO2/g for all tissues. The maximum residue limits (MRLs) established for FBZ and its metabolites in pig tissues are 50 ng/g for muscle, fat, and kidney and 500 ng/g for liver. This method is based on a liquid-liquid extraction followed by an oxidation with peracetic acid and a cleanup procedure based on 2 liquid-liquid extractions. Determination is achieved by high-performance liquid chromatography with ultraviolet detection. The present method is adjusted to the MRL established for FBZ and its metabolite residues. The analysis of the residues shows that after 72 h posttreatment, no FBZSO2 was detected in muscle, fat, and kidney and that liver levels were below the MRL. PMID- 10513005 TI - Chromatographic methods for analysis of aminoglycoside antibiotics. AB - Aminoglycosides are antimicrobial agents used frequently in treatment of human and animal diseases caused by aerobic, gram-negative bacteria. Because of the toxicity of these compounds, considerable effort has been attributed to analysis of aminoglycoside content in drug preparations, in serum and urine specimen in therapeutic drug monitoring, and in edible animal tissues in residue control. The present review emphasizes the analytical problems associated with aminoglycoside analysis. Screening methods based on microbiological and immunological procedures were briefly discussed. Gas chromatography and especially high-performance liquid chromatography appeared the most widely used chemical methods for the analysis of these compounds. Due to lack of volatility, chromophore, and hydrophility of aminoglycosides, most methods applied derivatization for enhancement of their chromatographic characteristics. The applicability and advantages of the various derivatization procedures were discussed in detail. A wide variety of detection methods, including mass spectrometry have been used. Packed column separation was generally used for gas chromatographic separation. In liquid chromatography, reversed phase, ion pair, ion exchange, and normal phase separation has been employed. Mass spectrometry, as a detection method, was discussed in detail. Extraction procedures from body fluids and tissues were emphasized. The performance and the operational conditions of the methods were described and detailed information of the data was provided also in table format. PMID- 10513006 TI - Multiresidue chromatographic method for the determination of macrolide residues in muscle by high-performance liquid chromatography with UV detection. AB - A high-performance liquid chromatographic (HPLC) method for the simultaneous determination of tilmicosin, tylosin, spiramycin, and its major metabolite neospiramycin was developed that is suitable for porcine, bovine, and poultry muscles. Macrolide residues were extracted from muscle with acetonitrile, fat was removed by liquid-liquid extraction with isooctane, and the extract was then cleaned on Bond Elut C18 cartridges. The HPLC separation was performed on an Inertsil ODS3 C18 column (150 x 4 mm) with 0.05% trifluoroacetic acid acetonitrile in a gradient mode. Two different chromatographic gradients were used for tilmicosin-tylosin and spiramycin-neospiramycin, and the detection wavelengths were 287 and 232 nm, respectively. The method was validated from 1/2 the maximum residue limit (MRL) to 4 times the MRL with pork muscle samples. Mean recoveries were 60, 63.5, 51, and 42% for tilmicosin, tylosin, spiramycin, and neospiramycin, respectively. The detection limits are 15 micrograms/kg for tilmicosin and tylosin, 30 micrograms/kg for spiramycin, and 25 micrograms/kg for neospiramycin. Linearity, precision, and accuracy of the method were also tested. PMID- 10513007 TI - Simultaneous determination of chlorthalidone and spironolactone with univariate and multivariate calibration: wavelength range selection. AB - Chlorthalidone and spironolactone were determined simultaneously with the aid of univariate and multivariate calibration methods. Univariate calibration was performed by the zero-crossing and derivative ratio spectrum methods. Extensive spectral overlap and the scarcity of wavelengths in derivative spectra allowing one analyte to be distinguished and quantitated in the presence of the other gave rise to poor results that called for multivariate calibration. Partial least squares regression was used in combination with a suitable method for selecting the optimum wavelength range and number of factors for analysis. The ensuing method was applied to simultaneous determination of chlorthalidone and spironolactone in a commercially available pharmaceutical preparation. The results were validated by high-performance liquid chromatography. PMID- 10513008 TI - Liquid chromatographic determination of para-toluenesulfonamide in edible fillet tissues from three species of fish. AB - Chloramine-T (N-sodium-N-chloro-p-toluene-sulfonamide) is a candidate therapeutic drug for treating bacterial gill disease, a predominant disease of a variety of fish species. Research has been initiated to obtain the U.S. Food and Drug Administration's (FDA) approval for the use of chloramine-T on a variety of fish species. An attribute of a therapeutic aquaculture drug that must be characterized before the FDA approves its use is depletion of the drug's marker residue (the drug's parent compound or metabolite of highest concentration in an edible tissue). para-Toluenesulfonamide (p-TSA) is the primary degradation product and marker residue for chloramine-T in rainbow trout. To conduct residue depletion studies for chloramine-T in fish, a robust analytical method sensitive and specific for p-TSA residues in edible fillet tissue from a variety of fish was required. Homogenized fillet tissues from rainbow trout (Oncorhynchus mykiss), walleye (Stizostedion vitreum), and channel catfish (Ictalurus punctatus) were fortified at nominal p-TSA concentrations of 17, 67, 200, 333, and 1000 ng/g. Samples were analyzed by isocratic reversed-phase liquid chromatography (LC) with absorbance detection at 226 nm. Mean recoveries of p-TSA ranged from 77 to 93.17%; relative standard deviations ranged from 1.5 to 14%; method quantitation limits ranged from 13 to 18 ng/g; and method detection limits ranged from 3.8 to 5.2 ng/g. The LC parameters produced p-TSA peaks without coelution of endogenous compounds and excluded chromatographic interference from at least 20 chemicals and drugs of potential use in aquaculture. PMID- 10513009 TI - Confirmatory test results on milk from commercial sources that tested positive by beta-lactam antibiotics screening tests. AB - Fifty-four milk samples from commercial sources that tested positive for beta lactam antibiotics were analyzed by a multiresidue liquid chromatographic (LC) procedure based on LC fractionation. Penicillin G and cephapirin were the beta lactam antibiotics found most frequently. Some samples did not contain detectable beta-lactam antibiotics. In a few, the presence of a beta-lactam antibiotic was suspected because certain LC fractions tested positive for antimicrobial activity, which was no longer present in a replicate treated with beta-lactamase. However, the unknowns could not be identified by LC analysis. PMID- 10513010 TI - Multiresidue analytical method for the determination of eight penicillin antibiotics in muscle tissue by ion-pair reversed-phase HPLC after precolumn derivatization. AB - A high-performance liquid chromatographic multiresidue method was developed for the determination of 8 penicillin compounds (benzylpenicillin, phenoxymethylpenicillin, ampicillin, amoxicillin, nafcillin, oxacillin, cloxacillin, and dicloxacillin) at trace levels in muscle tissue. This method involves extraction of the penicillins with phosphate buffer pH 9 followed by cleanup and concentration on a C18 solid-phase extraction column and reaction with benzoic anhydride at 50 degrees C for 5 min and with 1,2,4-triazole and mercury(II) chloride solution pH 9 at 65 degrees C for 10 min. The derivatized compounds are eluted on a C8 column with a mobile phase containing acetonitrile and phosphate buffer (pH 6; 0.1 mol/L) loaded with sodium thiosulfate and ion pairing tetrabutylammonium hydrogenosulphate. The method detection limit is approximately 3-11 micrograms/kg and the limit of determination was evaluated down to 25 micrograms/kg in line with the criteria of the EU decision No. 93/256/EEC. PMID- 10513011 TI - A test strip for diamines in tuna. AB - This study describes the production of a solid-phase assay (test strip/dipstick test) for putrescine and cadaverine in tuna based on the coupling of an amine oxidase to a peroxidase/dye system. The assay was linear to 75 microM in phosphate buffer, and the minimum detectable concentration was 0.5 microM (< 0.1 ppm), corresponding to 0.01 mg% in spiked extracts. Intra- and interassay precisions were < 20%. Test strips were stable at 4 degrees C for at least 12 months. Lysine, ornithine, and histidine did not react with the assay, and histamine reacted only minimally. Sixteen fish samples were tested by test strip and the standard AOAC protocol, and results were in good agreement. PMID- 10513012 TI - Solid-phase extraction method for patulin in apple juice and unfiltered apple juice. AB - Patulin, a mold metabolite, is commonly found in rotting apples. Some countries regulate patulin at levels ranging from 30 to 50 micrograms/L. Most analytical methods for patulin in apple juice include liquid-liquid partitions. A solid phase extraction method has been developed for apple juice and unfiltered apple juice in the United States. A portion of the test sample (5 mL) was passed through a macroporous copolymer cartridge and was washed with 1 mL 1% sodium bicarbonate and then with 1 mL 1% acetic acid. Patulin was eluted with 3 mL 2% acetonitrile in anhydrous ethyl ether and was determined by reversed-phase liquid chromatography with UV detection at 276 nm. Recoveries ranged from 93 to 104% in test samples spiked at 20-100 micrograms/L. PMID- 10513013 TI - Determination of lipids in infant formula powder by direct extraction methylation of lipids and fatty acid methyl esters (FAME) analysis by gas chromatography. AB - Fatty acid methyl esters (FAMEs) of pure triglyceride standards, oils, and fat from dry matrixes were formed by transesterification using sodium methoxide in methanol-hexane. FAMEs were produced by direct addition of sodium methoxide hexane to samples and heating to simultaneously extract and transesterify acyl lipids. FAMEs were quantitated by capillary gas chromatography (GC) over a fatty acid concentration range of 0 to 1.7 mg/mL (r > or = 0.9997). Total fat was calculated as the sum of individual fatty acids expressed as triglyceride equivalents, in accordance with nutrition labeling guidelines. Saturated, polyunsaturated, and monounsaturated fats were calculated as sums of individual free fatty acids. Absolute recoveries determined from individual fatty acids in test samples ranged from 69.7 to 106%. Recoveries (relative to the C13:0 internal standard) for individual fatty acids in test samples ranged from 95 to 106%. Reproducibility was constant at each fatty acid level in the reaction mixture (n = 5, coefficient of variation [CV] < 2%). Absolute recovery determined from the sum of total fatty acids in standard reference material (SRM) 1846 (powdered infant formula) was 96.4%. Analysis of SRM 1846 gave results that agreed closely with the certified fat and fatty acid values. Analysis of commercial infant formula gave results that were comparable to those obtained with AOAC Method 996.01. The direct extraction methylation procedure is rapid, and the transesterification of acyl lipids to form FAMEs is complete within 15 min. Classical saponification and refluxing are not required. This method provides FAMEs free of interferences and easily quantitated by GC or confirmed by GC/mass spectrometry (MS). Unambiguous MS identification of individual FAMEs derived from pure standards, SRM 1846, and powdered infant formula product was obtained. PMID- 10513014 TI - Liquid chromatographic analysis of vitamin K1 in milk-based infant formula with matrix solid-phase dispersion. AB - A liquid chromatographic method for vitamin K1 in milk-based infant formula is described. The vitamins are extracted from infant formula by matrix solid-phase dispersion and quantitated by reversed-phase chromatography with fluorescence detection. Vitamin K1 is converted to the fluorescent hydroquinone with a postcolumn zinc reductive reactor. The limit of detection is 12 pg, and the limit of quantitation is 38 pg on-column. Linear responses were obtained in the range 0.55-22.1 ng/ml (r2 = 0.9998). Recoveries of vitamin K1 from an analyte-fortified blank material for milk-based infant formula averaged 91.7% (n = 25). The method provides a rapid, specific, and easily controlled assay for vitamin K1 in fortified infant formula. PMID- 10513015 TI - Determination of choline in infant formula by ion chromatography. AB - Choline was determined in infant formula by ion chromatography with suppressed conductivity detection. Samples were digested with 1M hydrochloric acid, filtered, diluted, and injected into the chromatographic system. Choline and the alkali and alkaline earth metals were separated on a high-resolution cation exchange column and detected by suppressed conductivity. The method was linear between 2 and 200 mg/L (r2 = 0.9999), the concentration range of the diluted samples. This method accurately determined choline in powdered, concentrated, and ready-to-feed infant formulas. Recoveries of choline spikes into powdered infant formula at approximately 1, 0.8, 0.5, and 0.2 times the labeled value ranged from 85 to 114%. This method had good agreement for 8 blind duplicates. The values determined for these samples, which were used in an AOAC collaborative study of an enzymatic method, were consistent with the values determined by the enzymatic method. PMID- 10513016 TI - An immunoassay method for rapid detection of Staphylococcus aureus in cosmetics, pharmaceutical products, and raw materials. AB - The TECRA Staphylococcus aureus Visual Immunoassay allows a presumptive positive or negative result for the presence of S. aureus to be obtained within 26 h, in contrast to 4-5 days by traditional cultural methods. Presumptive positive immunoassay results are confirmed by streaking the enrichment broth onto conventional agar media. A validation study was undertaken to compare the TECRA assay with a cultural reference method based on the Bacteriological Analytical Manual (8th Ed.), which is also consistent with U.S. Pharmacopoeia requirements. The products tested included a range of cosmetics (toothpaste, shampoos, conditioners, sunscreens, moisturizers, lip and eye creams) and pharmaceuticals (cough mixtures, laxatives, ulcer treatments, infant formulae, antiseptic cream), as well as some pharmaceutical ingredients. Samples were inoculated with S. aureus at 10-20 cfu/g, and then enriched for 24 h at 35 degrees-37 degrees C at a product-to-sample ratio of 1:100. Two different enrichment broths were used for the study: Tryptone Soya Broth with 4% Tween 80 and Modified Letheen Broth. For both enrichment broths, results of the immunoassay and the reference method showed close correlation. The TECRA S. aureus Visual Immunoassay provides a rapid and convenient alternative to cultural methods and provides advantages to industry, such as greater speed of product and ingredient release and faster tracing of contamination problems. Because the immunoassay may be read either visually or with the aid of a plate reader, there is no need for an initial outlay on capital equipment. However, the assay can be automated if required. PMID- 10513017 TI - Improved signals ratio resolution method by optimization of resolution function- simultaneous determination of Cu(II) and Cd(II) in water samples. AB - From an ecological and economical point of view, it is important to design analytical procedures for monitoring heavy metals in the environment and industrial processes in a way to minimize the use of hazardous reagents and reduce the analysis time. In this paper, a well-known dithizonate extraction based method for the determination of many metal ions was improved by using chemometrical selectivity of the strongly overlapped spectra of copper and cadmium dithizonates in CCl4 for their simultaneous determination from a single extraction at pH 10. The individual absorption spectra, having absorption maxima difference of only 20 nm, were separated, and the metal ions were quantified by using an improved procedure for optimizing the resolving function in a recently proposed signals ratio method. The procedure consists of using many different resolving functions and plotting the difference of the mean of absolute and nonabsolute mean values of pseudosignals [PDMMV (PS)] against analyte concentrations obtained with each of the resolving functions, thus obtaining 2 straight lines having intersections that give a unique and reliable value of the unknown concentration of the individual analyte in mixture giving strongly overlapped spectra. In this way, the main drawback of the signals ratio resolution method, that is, the visual estimation of optimal resolving function, is eliminated. The proposed parameter, PDMMV (PS), was tested by using both simulated and experimental spectra. Copper was determined in the mixture with ca 20-fold excess of cadmium, and cadmium was determined in ca 10-fold excess of copper at submicromolar concentration levels. PMID- 10513018 TI - Determination of organochlorine pesticide residues in medicinal plants sold in Coimbra, Portugal. AB - Levels of 14 organochlorine pesticide residues--1,1,1-trichloro-2,2-bis(p chlorophenyl)ethane (DDT) group (p,p'-DDT, o,p'-DDT, p,p'-DDD, and p,p'-DDE), HCH isomers (alpha-, beta-, and gamma-HCH), hexachlorobenzene (HCB), aldrin, dieldrin, endrin, heptachlor, alpha-endosulfan, and endosulfan sulfate--were evaluated in 127 samples of medicinal plants collected in pharmacies (78 samples) and herb stores (49 samples) in 1996. Samples were divided between 15 national brands and 7 foreign brands. Most samples sold in pharmacies contained residues of gamma-HCH (51.3%). All residues were detected in analyzed samples, with exception of endrin in herb store samples. Detection frequency varied between 51.3% for gamma-HCH and 1.3% for endrin in pharmacy samples, and between 34.7% for HCB and 4.1% for endosulfan sulfate in herb store samples. Maximum residue levels were exceeded in 38 (48.7%) pharmacy samples and in 26 (53.1%) medicinal herb store samples. PMID- 10513019 TI - Determination of inorganic arsenic in marine food samples by hydrochloric acid distillation and flow-injection hydride-generation atomic absorption spectrometry. AB - A simple, rapid, and reliable method was developed for determination of inorganic As in biological samples such as fish fillet. Inorganic AS was distilled from the sample as AsCl3 with HCl. The separated inorganic AS was determined by flow injection hydride-generation atomic absorption spectrometry after prereduction with KI and HCl. The influences of various concentrations of KI, ascorbic acid, and HCl in the prereduction stage; NaBH4 as the reductant; and HCl as the carrier solution on analytical results were studied. Digestion was performed in a Kjeldahl digestion system for 75 min with 4 mL nitric acid and 1 mL sulfuric acid at 380 degrees C. The concentrations of inorganic As in samples were less than 0.1 mg/kg dry weight for fish fillet and somewhat higher for crustaceans and bivalve molluscs. The total and inorganic As contents of various marine biological samples and certified reference materials were determined. PMID- 10513020 TI - Determination of residues of azamethiphos in salmon tissue by liquid chromatography with fluorescence detection. AB - A liquid chromatographic (LC) method with fluorescence detection (FLD) is described for determining residues of the pesticide azamethiphos (AZA) in salmon tissue. The sample is extracted with ethyl acetate, centrifuged, dehydrated with anhydrous sodium sulfate, evaporated, reconstituted in water, and defatted with hexane. The aqueous phase is passed through a C18 solid-phase extraction (SPE) column. The SPE column is eluted with methanol, and the eluate is evaporated to dryness and then taken up in 10% acetonitrile (ACN) in water. The analyte is determined by LC using a C18 column, ACN-H2O (32 + 68) mobile phase, and FLD with excitation at 230 nm and emission at 345 nm. Composited salmon tissues were fortified with AZA at 5, 10, 21, 42, and 83 ng/g or ppb (target level, X = 10 ng/g). Overall recoveries were 86%, with between-day variability of 5.3%. The method detection limit was calculated as 1.2 ppb AZA based on a 5 g sample. The limit of quantitation as determined empirically by this method is the lower limit of the standard curve, approximately 5 ppb. PMID- 10513021 TI - Multiresidue determination of 19 fungicides in processed fruits and vegetables by capillary gas chromatography after gel permeation chromatography. AB - A gas chromatographic (GC) method was developed for simultaneous determination of 19 fungicides (chlorothalonil, vinclozolin, dichlofuanid, triadimefon, penconazole, chlozolinate, captan, procymidone, triadimenol, folpet, hexaconazole, myclobutanil, cyproconazole, propiconazole, nuarimol, captafol, iprodione, fenarimol, and bitertanol) and the acaricide tetradifon in tomato puree, peach nectar, orange juice, and canned peas. Samples were extracted with acetone, partitioned with ethyl acetate-cyclohexane (50 + 50, v/v), and cleaned using gel permeation chromatography with ethyl acetate-cyclohexane (50 + 50, v/v) as eluant. The final extract was analyzed by GC with ion trap mass spectrometry (ITMS) using a DB5 capillary column. Recoveries from fortified samples ranged from 74.6 to 99.3%, except for triadimenol and bitertanol. Quantitation limits for most analytes were between 0.005 and 0.050 mg/kg. The purified extracts were analyzed further by GC with electron capture and nitrogen phosphorus detection, and the results were compared with those obtained by ITMS. PMID- 10513022 TI - A pilot study for the statistical treatment of nutritional parameters in the quality control of canned fishing goods. AB - This pilot study was derived as a consequence of European Directives 496/90 and 493/91 in which a regulation on the labeling of canned fishing goods was established. The study was intended primarily to assess whether different Spanish canned fishing goods might be differentiated by their basic nutritional composition (i.e., ash, chlorine as NaCl, fat, humidity, total proteins, and dry residue) and, second, to study each particular type of good. Accordingly, a univariate nonparametric statistical test and 2 multivariate chemometric techniques (factor and cluster analyses) were used. The pilot study revealed that (1) the basic nutritional variables did not allow a clear distinction among canned goods when different commodities were considered, but they seemed useful for obtaining information for only one type of good; and (2) the variables that gave the most useful information to visualize the appearance of groups in the data sets were humidity, dry residue, fat, and proteins, although their particular usefulness was found to be different when different species were considered. PMID- 10513023 TI - Functional communication training using assistive devices: recruiting natural communities of reinforcement. AB - We evaluated the effectiveness of functional communication training (FCT) as an intervention for the problem behavior exhibited by 5 students with severe disabilities both in school and in the community. Following an assessment of the function of their problem behavior, the students were taught to use assistive communication devices in school to request the objects and activities that presumably were maintaining their behavior. Multiple baseline data collected across the students indicated that not only did the students use their devices successfully, but the intervention also reduced their problem behavior. In addition, data from community settings showed generalization to untrained community members. These results replicate other successful efforts to use FCT with individuals having limited communication skills, and demonstrate the value of teaching skills to recruit natural communities of reinforcement in order to generalize intervention effects to meaningful nontraining environments. PMID- 10513024 TI - Further analysis of problem behavior in response class hierarchies. AB - A functional analysis identified the reinforcers for 3 participants' problem behavior, but only relatively mild problem behaviors (e.g., screaming, disruption) were observed when all topographies produced tested consequences. We then conducted an extinction analysis in which specific topographies produced a reinforcer while all other topographies were on extinction. The extinction analysis confirmed that the same reinforcer identified in the initial functional analysis maintained more severe topographies of problem behavior (e.g., aggression). In addition, results of the extinction analysis indicated that 2 of the participants displayed patterns of responding consistent with a response class hierarchy hypothesis, in which less severe problem behavior frequently occurred prior to more severe topographies. The 3rd participant displayed a response pattern indicative of differential reinforcement effects. PMID- 10513025 TI - Competition between positive and negative reinforcement in the treatment of escape behavior. AB - We compared the effects of reinforcing compliance with either positive reinforcement (edible items) or negative reinforcement (a break) on 5 participants' escape-maintained problem behavior. Both procedures were assessed with or without extinction. Results showed that compliance was higher and problem behavior was lower for all participants when compliance produced an edible item rather than a break. Treatment gains were achieved without the use of extinction. Results are discussed regarding the use of positive reinforcement to treat escape behavior. PMID- 10513026 TI - Multicomponent assessment and treatment of cigarette pica. AB - We conducted a multicomponent assessment and treatment for 4 individuals who engaged in cigarette pica. During Phase 1, three stimulus preference assessments were conducted to identify (a) the reinforcing component of the cigarette, (b) potential alternative reinforcers that may be used during treatment, and (c) whether the alternative reinforcer would compete effectively with cigarettes. Results were successful in identifying the reinforcing component of the cigarette and suggested the feasibility of using alternative reinforcers during treatment to eliminate cigarette pica. During Phase 2, the effects of two treatment procedures were evaluated. Noncontingent reinforcement (NCR) with the alternative edible reinforcer reduced the pica of 2 of the participants, but effects were not maintained when the initial dense schedule of NCR was thinned. Subsequently, differential reinforcement of alternative behavior with the alternative edible reinforcer was effective in reducing pica for 3 participants. An evaluation of nine treatment procedures failed to identify an effective intervention for the remaining participant; consequently, preventive measures were designed to minimize occurrences of cigarette pica. PMID- 10513027 TI - Effects of reinforcement for alternative behavior during punishment of self injury. AB - A number of variables influence the effectiveness of punishment and may determine the extent to which less intrusive forms of punishment may be used as alternatives to more intrusive interventions. For example, it has been suggested that response suppression during punishment may be facilitated if reinforcement is concurrently available for an alternative response. However, results of basic research demonstrating this finding have not been replicated with interventions more commonly prescribed as treatments for problem behavior. We evaluated the effects of relatively benign punishment procedures (reprimands or brief manual restraint) on the self-injurious behavior of 4 individuals who had been diagnosed with mental retardation, when access to reinforcement for alternative behavior (manipulation of leisure materials) was and was not available. In all cases, punishment produced greater response suppression when reinforcement for an alternative response was available. PMID- 10513028 TI - A functional analysis of hair pulling. AB - We experimentally assessed the functions of hair pulling and hair manipulation of a 19-year-old woman (Kris) with moderate mental retardation and cerebral palsy. In Phase 1 a functional analysis revealed that Kris pulled and manipulated hair for the greatest amount of time in the alone condition, suggesting that the behaviors were maintained by some form of automatic reinforcement (Vaughan & Michael, 1982). In Phase 2 we assessed the nature of the sensory stimulation that maintained hair pulling by providing continuous access to previously pulled or cut hair and, thereafter, by having Kris wear a rubber glove. The results suggested that hair pulling was maintained by digital-tactile stimulation (automatic positive reinforcement). These findings are discussed, and recommendations for further analyses of automatically reinforced habit behaviors are provided. PMID- 10513029 TI - Augmenting simplified habit reversal in the treatment of oral-digital habits exhibited by individuals with mental retardation. AB - We investigated whether a simplified habit reversal treatment eliminates fingernail biting and related oral-digital habits exhibited by individuals with mild to moderate mental retardation. Although simplified habit reversal did little to decrease the target behaviors for 3 of 4 participants, simplified habit reversal plus additional treatment procedures decreased the behavior to near-zero levels for all participants. These procedures included remote prompting, remote contingencies involving differential reinforcement plus response cost, and differential reinforcement of nail growth. Limitations of habit reversal for individuals with mental retardation along with directions for future research involving therapist-mediated treatment procedures, particularly those involving remote prompting and remote contingencies, are discussed. PMID- 10513030 TI - Effects of presession attention on the frequency of attention-maintained behavior. AB - The effect of prior attention was systematically manipulated to study its influence on rates of yelling and head hitting, both maintained by positive reinforcement in the form of attention. Higher levels of head hitting occurred in analogue attention conditions when the person was deprived of attention (no social interactions for 1 hr) prior to the analysis in comparison to when the person received high levels of attention (attention delivered on a fixed-time 30 s schedule for 1 hr) prior to the analysis. PMID- 10513031 TI - Reducing wandering by persons with dementia using differential reinforcement. AB - Wandering behavior of 4 geriatric patients with dementia residing in a long-term care facility was assessed using direct behavioral observations. The consequences identified during the observations as maintaining wandering for each patient were then applied for the absence of wandering using differential reinforcement of other behavior (DRO). The effectiveness of the DRO procedure was evaluated using an ABAB design. Results indicated significant reductions in wandering during treatment. PMID- 10513032 TI - Training and generalization of sexual abuse prevention skills for women with mental retardation. AB - Previous research has shown that behavioral skills training to teach sexual abuse prevention skills to women with mental retardation results in skill acquisition but poor generalization. In this investigation we evaluated procedures for enhancing generalization following training. Five women with mental retardation received 10 behavioral skills training sessions followed by in situ training when the skills did not fully generalize. Behavioral skills training resulted in skill acquisition and in situ training produced generalized responding during naturalistic assessments. PMID- 10513033 TI - [Predisposition to in vitro oxidation of LDL isolated from hypercholesterolemic patients. Interaction with arterial proteoglycans]. AB - It is very well documented that cholesterol of the atherosclerotic plaques comes from LDL plasma particles and also a positive correlation between LDL plasma level and intensity of atherosclerotic lesions has been shown. Recent findings have revealed a direct relationship among arterial tissue atherogenic response and polyunsaturated fatty acids oxidation. The oxidized fatty acids within the low density lipoproteins makes these particles more atherogenic. In the present study the susceptibility of low density lipoprotein isolated from hypercholesterolemic patients to become oxidatively modified in vitro was investigated. The results show that the predisposition of hypercholesterolemic patients LDL to oxidation is fourfold increased, expressed in TBARS content (p < 0.0001) as compared to control LDL. Furthermore, the interactions of LDL with arterial proteoglycans apparently contribute to the accumulation of apo-B lipoproteins in atherogenesis. The hypercholesterolemic patients LDL shows higher values of proteoglycans-insolubilized LDL than LDL from controls. PMID- 10513034 TI - [DNA analysis for paternity testing and forensic identification]. AB - Non coding DNA of human genome contains an important part of tandem repetitive DNA which last years has been extensively applied to solve paternity disputes and individual identification cases. A class of this DNA are minisatellites and microsatellites loci. Such loci are highly polimorphics because they have a multiple of different alleles to make them ideals genetic markers for these purposes. This article is a review about these loci, methods of their analysis and practical aspects about their applications to determine biological paternity or individual identification in the forensic genetic field are considered. PMID- 10513035 TI - [Reaction time to dichotic visual stimulation and its relationship to cerebral hemispheric specialization]. AB - There is a renewed interest in the study of the cerebral hemispheric specialization given its physiological, physiopathological, clinical and educational relevancy. According to this, a stimulus is better processed in one of the cerebral hemispheres. The visual dichotic stimulation allows to present a stimulus selectively to one of the hemispheres, and this was used in the present study for the measurement of the reaction time (TR) and for the establishment of its correlation with an objective and validated measure of the cerebral hemispheric specialization. Ninety eight persons with an average age of 22.2 +/- 0.7 (X +/- EE) years were studied by means of the Oldfield protocol for the determination of the Cerebral Hemispheric Lateralization Index (IL). Additionally, the age, the sex and the date of last menstruation in the case of females, were registered. The dichotic stimulation and the determination of the visual TR was accomplished by computer with a time resolution of 100 microseconds. The stimulus consisted of the random presentation of a white square on a black background in a computer monitor, to which the subject had to respond by pressing a key with his best hand. The reaction time was registered 10 times in each visual field for a total of 20 records per eye. The IL of the sample was predominantly right (66.32 +/- 4.64), and significantly smaller in males than in females (p < 0.05). The TR to stimuli in the left visual field was significantly greater (p < 0.05) than in the right visual field for both eyes. When analyzing the global results of each cerebral hemisphere, a TR significantly smaller for the left hemisphere (304.33 +/- 4.1 ms) than for the right (312.35 +/- 4.5 ms, p < 0.05) was observed. These results suggest that the cerebral hemispheric specialization, expressed as a smaller response time, cannot alone be only the product of a better intrinsic hemispheric processing of the information, but also of the anatomical relationships of the proximity of the hemisphere that receives the stimulus and the center or area that will generate the response, as is the case of the motor center located in the left hemisphere in the immense majority of the right-handed subjects in the sample and in the general population. PMID- 10513036 TI - [Obesity, insulin resistance and skeletal muscle characteristics]. AB - Insulin resistance seems to be a metabolic aberration associated with obesity. Impaired insulin action is also central to a cluster of diseases including non insulin dependent diabetes, hypertension, dyslipidemias and atherosclerosis. Body fat distribution, especially upper body segment obesity is related to insulin resistance. Glucose uptake is insulin dependent in skeletal muscle and adipose tissue. From a quantitative standpoint, skeletal muscle has the greater impact on whole body glucose economy, therefore the cause of altered insulin sensitivity has been looked for in this tissue. The skeletal muscle is composed of different types of fibers with specific metabolic and circulatory characteristics; type IIB fibers are less insulin-sensitive and their proportion has been related to obesity and insulin resistance. The different factors that may impair insulin action and alter glucose uptake in skeletal muscle are: lower blood flow to muscle, produced by either decreased vasodilation or by increased sympathetic nerve activity; augmented diffusion distance from capillaries to muscle due to a decrease in capillary number or to enlarged muscle cells; decrease of insulin receptors; change in the fatty acid profile of major membrane structural phospholipids; decrease in glucose transporters (GLUT 4) and/or hexokinase; impairment in metabolic routes of glucose in muscle as reduction in glycogen synthase. Also, the high rate of lipolysis present in obesity and in insulin resistance could lead to an impaired glucose oxidation in muscle. PMID- 10513037 TI - [Effectiveness educational programs for school dental health. Experimental trial]. AB - This paper is a limited-scope experimental study to evaluate the effectiveness of the Dental Health Educational Programs implemented in Venezuelan schools by the Dentistry Division of the Ministry of Health and Social Welfare. Two hundred and ninety-six six-to-twelve year olds were randomly selected from two schools, one private, the other public, located in the Municipality of Maracaibo, State of Zulia, Venezuela. The children assigned to each school were randomly distributed among the experimental and control groups. A general hypothesis was set up, stating that reinforcement and motivation generate changes in the gingival health of the children under study, but with different results for each, depending on their psycho-social characteristics. Two approaches were used: instruction/supervision and instruction/supervision/reinforcement/stimulation. These two approaches were then measured against a "psycho-social profile" which is based on material conditions and the developmental stages of the children. Instructions were given to the experimental group on how to brush their teeth. They were supervised, reinforced and stimulated for a period of six months, with a post-test assessment. The control group was only given instructions on how to brush their teeth and supervised. A scale based on Massler's classification was used to determine the developmental stages. Standardized indices were used to evaluate the dependent variable: Plaque Index and Gingival Index. The results showed that: the material living conditions and developmental stage of the child are elements that influence the assimilation of pre-established behavior, the basis of preventive programs. PMID- 10513038 TI - [Effect of amiloride and its derivative dichlorobenzamil on guinea pig atria: interaction with other inotropic mechanisms]. AB - The inotropic and chronotropic effects of Amiloride (AMI) and Dichloro-benzamil Amiloride (DBC-AMI) were studied on the guinea pig isolated atria, also, the interaction between these drugs and Beta-methyl-Digoxin (BM-DIGO), epinephrine and low extracellular potassium (1 mM). AMI (10(-3) M) has a negative chronotropic and positive inotropic effects, not dependent on the autonomic system. DCB-AMI has a bimodal effect on the contractile force: increases it at low concentrations but causes a decrease at concentrations higher than 10(-6) M. The effect of AMI on the sinus frequency is unchanged by BM-DIGO. AMI (10(-3) M) decreases the inotropic effect of BM-DIGO and increases the toxic concentration of this drug on isolated tissues. The dose-response curve to epinephrine was not changed by AMI. Similar results were obtained using DCB-AMI (2 x 10(-7) M). The positive inotropic effect obtained by low extracellular potassium (1 mM) was not altered by AMI. The activity of the Mg(++)-dependent, Na+/K+ ATPase measured in the microsomal fraction obtained from guinea pig heart was diminished (10%) by AMI (10(-3) M). The drug did not affect the inhibition of the enzyme induced by ouabain. In conclusion, our experiments show multiple effects of AMI and DCB-AMI on the guinea pig heart. The inhibition of the Na+/Ca++ exchange explains them only partially. A slow channel blocking effect appears fundamental to interpret our results. PMID- 10513039 TI - Chagas' heart disease. PMID- 10513040 TI - Why do we need randomized and epidemiological studies on cardiovascular disease? Evidence-based cardiology VII. AB - Over the last two decades the results of randomized clinical studies, which are powerful aids for correctly assessing therapeutical strategies, have consolidated cardiological practice. In addition, scientifically interesting hypotheses have been generated through the results of epidemiological studies. Properly conducted randomized studies without systematic errors and with statistical power adequate for demonstrating moderate and reasonable benefits in relevant clinical outcomes have provided reliable and strong results altering clinical practice, thus providing adequate treatment for patients with cardiovascular disease (CVD). The dissemination and use of evidence-based medicine in treating coronary artery disease (CAD), heart failure (HF), and in prevention will prevent hundreds of thousands of deaths annually in developed and developing countries. CVD is responsible for approximately 12 million deaths annually throughout the world, and approximately 60% of these deaths occur in developing countries. During recent years, an increase in mortality and morbidity rates due to CVD has occurred in developing countries. This increase is an indication that an epidemiological (demographic, economical, and health-related) transition is taking place in developing countries and this transition implies a global epidemic of CVD, which will require wide-ranging and globally effective strategies for prevention. The identification of conventional and emerging risk factors for CVD, as well as their management in high-risk individuals, has contributed to the decrease in the mortality rate due to CVD. Through a national collaboration, several multi-center and multinational randomized and epidemiological studies have been carried out throughout Brazil, thus contributing not only to a generalized scientific growth in different Brazilian hospitals but also to the consolidation of an increasingly evidence-based clinical practice. PMID- 10513041 TI - Cholinergic stimulation with pyridostigmine, hemodynamic and echocardiographic analysis in healthy subjects. AB - OBJECTIVE: Growing evidence suggests that sudden death after an acute myocardial infarction (AMI) correlates with autonomic nervous system imbalance. Parasympathomimetic drugs have been tested to reverse these changes. However, their effects on ventricular function need specific evaluation. Our objective was to analyze pyridostigmine's (PYR) effect on hemodynamic and echocardiographic variables of ventricular function. METHODS: Twenty healthy volunteers underwent Doppler echocardiographic evaluations, blood pressure (BP), and heart rate (HR) assessment at rest, before and 120 min after ingestion of 30 mg PYR or placebo, according to a double-blind, placebo-controlled, crossed and randomized protocol, on different days. RESULTS: PYR was well tolerated and did not cause alterations in BP or in ventricular systolic function. A reduction in HR of 10.9 +/- 1.3% occurred (p < 0.00001). There was an A wave reduction in the mitral flow (p < 0.01) and an E/A ratio increase (p < 0.001) without changes in the other diastolic function parameters (p > 0.05). CONCLUSION: PYR reduces HR and increases E/A ratio, without hemodynamic impairment or ventricular function change. PMID- 10513042 TI - Anomalous origin of the left coronary artery from the pulmonary trunk. Clinical features and midterm results after surgical treatment. AB - OBJECTIVE: To report the authors' experience with the anomalous origin of the left coronary artery (AOLCA) from the pulmonary trunk, emphasizing preoperative data, surgical aspects and midterm results of the follow-up. METHODS: Retrospective analysis of 11 patients operated upon at the Royal Brompton Hospital from October, 84 to April, 97. RESULTS: Nine infants had heart failure (HF) and two other children presented with dyspnea and chest pain. All had ECG changes. The echocardio-gram identified the anomalous origin of the coronary artery in 7 (64%) patients and hemodynamic studies were performed in 7 patients. All infants were operated upon between the 2nd and 10th month of life. Six patients were treated with aortic reimplantation of the left coronary artery, whereas five were operated upon according to the Takeuchi technique. All patients are alive, with clear improvement of the ECG changes and ventricular function, as evaluated by echocardiography. Two patients operated upon according to the Takeuchi technique required additional surgery due to severe supravalvular pulmonary stenosis. CONCLUSION: AOLCA is a rare disease. Most patients show early signs of severe HF associated with ECG findings. Surgical therapy must be instituted early in the disease, preferentially through aortic implantation of the anomalous coronary artery, with a high possibility of success. Shortly after surgery, clinical and ECG improvement, as well as normalization of left ventricular function, should be expected. PMID- 10513043 TI - Therapeutic approach to patients complaining of high blood pressure in a cardiological emergency room. AB - OBJECTIVE: To evaluate the management of patients complaining of high blood pressure (BP) in a cardiological emergency room. METHODS: Patients referred to the cardiological emergency room with the main complaint of high blood pressure were consecutively selected. The prescriptions and the choice of antihypertensive drugs were assessed. The classification of these patients as hypertensive emergencies or pseudoemergencies, according to the physician who provided initial care, was recorded. RESULTS: From a total of 858 patients presenting to the emergency room, 80 (9.3%) complained of high BP, and 61 (76.3%) received antihypertensive drugs. Sublingual nifedipine was the most commonly used drug (59%). One patient received intravenous medication, one patient was hospitalized and 6 patients (7.5%) were classified as hypertensive emergencies or pseudoemergencies. CONCLUSION: High BP could seldom be classified as a hypertensive emergency or pseudoemergency, even though it was a frequent complaint (9.3% of visits). Currently, the therapeutic approach is not recommended, even in specialized clinics. PMID- 10513044 TI - Mital valve disease with rheumatic appearance in the presence of left ventricular endomyocardial fibrosis. AB - This is a report of a nine-year-old boy with both mitral stenosis and regurgitation and extensive endomyocardial fibrosis of the left ventricle. Focus is given to the singularity of the fibrotic process, with an emphasis on the etiopatho-genic aspects. PMID- 10513045 TI - Atrial infarction is a unique and often unrecognized clinical entity. AB - A patient with heart failure and acute atrial fibrillation received the final diagnosis of atrial infarction associated with ventricular infarction based on clinical findings of ischemia in association with atrial fibrillation and heart failure (mechanisms probably involved: contractile dysfunction and loss of atrial contribution). Although a transesophageal echocardiography, which could refine the diagnosis of anatomic abnormalities, was not performed, all evidence led to the diagnosis of atrial involvement. Electrocardiographic findings were consistent with Liu's major criterion 3. Therapy with digitalis, quinidine and angiotensin-converting enzyme inhibitors was chosen, as the patient had acute pulmonary edema. The use of beta-blockers and verapamil was res-tricted. No other complications, such as thrombo-embolism or atrial rupture, were noted. PMID- 10513046 TI - Prognostic factors in patients with congestive heart failure. PMID- 10513047 TI - Cardiopulmonary resuscitation: update, controversies and new advances. AB - Cardiopulmonary arrest is a medical emergency in which the lapse of time between event onset and the initiation of measures of basic and advanced support, as well as the correct care based on specific protocols for each clinical situation, constitute decisive factors for a successful therapy. Cardiopulmonary arrest care cannot be restricted to the hospital setting because of its fulminant nature. This necessitates the creation of new concepts, strategies and structures, such as the concept of life chain, cardiopulmonary resuscitation courses for professionals who work in emergency medical services, the automated external defibrillator, the implantable cardioverter-defibrillator, and mobile intensive care units, among others. New concepts, strategies and structures motivated by new advances have also modified the treatment and improved the results of cardiopulmonary resuscitation in the hospital setting. Among them, we can cite the concept of cerebral resuscitation, the application of the life chain, the creation of the universal life support algorithm, the adjust-ment of drug doses, new techniques--measure of the end-tidal carbon dioxide levels and of the coronary perfusion pressure--and new drugs under research. PMID- 10513048 TI - Continuing medical education in cardiology. Proposal for a national program. Committee of Medical Residency and Training in Cardiology--SBC/FUNCOR. PMID- 10513049 TI - Preliminary sketch of medical residency in cardiology. Committee of Medical Residency and Training in Cardiology SBC/FUNCOR. PMID- 10513050 TI - [Ultrastructural features of cardiac muscle fibers of albino rats during the pregnancy and puerperal cycle]. AB - PURPOSE: In the present study we evaluated the morphology of left ventricular cardiomyocytes of albino rats during pregnancy and puerperium by means of transmission electron microscopy. METHODS: Once pregnancy was confirmed, 77 rats were randomly divided in two groups, respectively, gestation (G) and puerperium (P). The animals of the gestation group were divided into four subgroups, according to gestational age: 1st (G-A), 7th (G-B), 14th (G-C) and 21st (G-D) days of pregnancy. The group defined as puerperium was divided into three subgroups: 7th (P-A), 14th (P-B) and 21st (P-C) days of puerperium. In the end of each established period, the animals were sacrificed and fragments of the medium third of the left ventricle were resected and routinely prepared for electron microscopy analysis. RESULTS: The results obtained with transmission electron microscopy analysis revealed a gradative increase of the cardiomyocytes during pregnancy (increase of myofibrils, which are permeated by numerous mitochondria at the end of gestation). The blood capillary wall showed progressive thinning, with an increase of pynocytotic vesicles in endothelial cells, and intense sarcolemal folding at T-tubule level (capillary tunnels). In the puerperium group, there is a progressive regression in these alterations returning to pre gestational level at the end of the puerperium cycle. These findings indicate the occurrence of hypertrophy of cardiomyocytes during pregnancy. CONCLUSION: The findings indicate the occurrence of hypertrophy of cardiomyocytes during pregnancy. PMID- 10513051 TI - [Vasoactive substances and the modulation of the hepatic microvascular system]. AB - PURPOSE: To review some aspects of the hepatic phylogeny and ontogeny, the hepatic microvascular system, and the modulation of the tonus on this vascular system by different vasoactive substances. METHOD: Text books and articles from MEDLINE-indexed journals were consulted. RESULTS: Fifty-two articles, that were published between 1949 and 1997, were selected. They provided us with information concerning hepatic phylogeny and ontogeny, the hepatic microvascular system, and the modulation of the tonus in this vascular system. CONCLUSION: The architecture of the hepatic microvascular system, a unique and complex vascular system, is well suited to the functions of the organ. Different factors, including endothelial vasoactive substances, participate in the modulation of the vascular resistance through the liver adequating the liver perfusion to the homeostatic needs. The liver is eminently a maintainer of internal stability. PMID- 10513052 TI - [Implantation of a program of terminal evaluation of graduating medicalstudent performance to estimate the efficacy of the curriculum in the ++Ribeirao Preto Medical School]. AB - PURPOSE: To describe the main steps in the implementation of such assessment, as well as to present the measures taken in order to overcome the difficulties that have been found. METHODS: The proposed two-step assessment consists of a test of cognitive aspects based on Multiple Choice Questions (MCQ) and practical exams of clinical skills in each of 5 terminal areas. Students enrolled as volunteer for either step or both. The examinations were developed and carried out by faculty members nominated by the involved Departments under the supervision of an institutional work group. The rates of student enrollment for the summative assessment has been recorded and the responses of both students and faculty members were evaluated by means of specific questionnaires. RESULTS: In the first two years, adhesion of students to the MCQ test was consistently high, but volunteering to the practical exams was persistently low. Moreover, there was a sharp decrease in faculty involvement, from the first to the second year. To overcome these difficulties, a number of measures were implemented aiming at increasing student adhesion and faculty involvement, as well as to improve assessment methods. As a consequence, a remarkable increase in both student adhesion and faculty involvement was recorded. CONCLUSION: The measures taken resulted in sharp increases in both acceptance of the proposed assessment and the quality of the examination methods, which have allowed a more reliable characterization of the strenghts and weaknesses of the local curriculum. PMID- 10513053 TI - Persistence of Leishmania antigen in C57Bl/6j inbred mice infected with Leishmania (Leishmania) amazonensis. AB - PURPOSE: To develop an animal model for studying mucocutaneous leishmaniasis. METHODS: The hind footpad of C57Bl/6j inbred mice was experimentally infected with 10(7) Leishmania (Leishmania) amazonensis promastigote and the skin was studied through light and electron transmission microscopy and immunohistochemistry (PAP) techniques. RESULTS: There were morphological evidences of cellular immune mechanisms and hypersensitivity reaction after eight weeks of infection and metastasis and well shaped parasites at ultrastructural level by fifty-one weeks post infection. Relapse of infection with mucosa lesions occurred around the 50th week after inoculation. CONCLUSION: The use of this animal model in long term follow up could be an useful experimental model for human mucocutaneous leishmaniasis. PMID- 10513054 TI - [Clinical response to inhalation and oral antibiotics in patients with bronchiectasis]. AB - INTRODUCTION: The aim of this technique was to achieve high concentration of the antibiotic that excedeed the minimal inhibitory concentration (MIC) of the microbial load present in the sputum. METHOD: To evaluate the bronchiectasis patients with pictures of infectious exacerbations response to the treatment with antibiotic by oral way (roxitromicin 300 mg/day for 21 days) and in cases of failure of this schema the use of antibiotic by inhalatory way (gentamicin 80 mg/2 times day for 21 days), 28 patients were evaluated a special ambulatory, some signs and respiratory symptoms according to the "Cotes modified scale" (sputum, cough, bronchospasm and dyspnea). STATISTICAL ANALYSIS: We used: Kappa concordant test and McNemar test for discordation in the evaluation of the degree of signal and respiratory symptoms, Wilcoxon test for periods without infection, Fisher test for the collaterals effects presented, G of Cochran test for the personal history analysis. RESULTS: The personal history did not influence the evolution of bronchopulmonary infeccion, the evaluated signs and symptoms had significant improvement except dyspnea that stayed the same in 80% of the cases. DISCUSSION: The group that had used the antibiotics schema by inhalatory way after oral scheme failure had a significant longer period without bronchopulmonary infection but with superior collateral effects without clinicals repercussions. CONCLUSION: The use of inhalatory antibiotic in infectious exacerbations in patients with bronchiectasis was better than the oral way. PMID- 10513055 TI - [Otorhinolaryngological effects of cocaine and/or crack abuse in drug addicts]. AB - PURPOSE: To assess if there are typical otorhinolaryngological findings in patients addicted to cocaine and/or crack. METHODS: Eighteen patients who were addicted to cocaine and/or crack, were studied. Their clinical history was obtained and were submitted to an otorhinolaryngological examination. RESULTS: It was impossible to find out a correlation between the time the patient is addicted to the drug, the amount of drug the patient is used to inhale, the frequency or the last time he used it and the nose and throat symptoms, as well as those factors and the otorhinolaryngological examination. CONCLUSION: Owing to the unspecificity on clinical findings during the otorhinolaryngological examination, it is necessary that the physician suspects the abuse of drugs when he is performing the clinical history; otherwise, he will not reach this important diagnosis. PMID- 10513056 TI - [Histomorphometric study of cardiomyocytes of the left ventricle of pregnant albino rats]. AB - PURPOSE: In the present study we evaluated, by light microscopy, and throughout morphometry, whether hypertrophy of cardiac striated muscular fibers of left ventricular occur in albino rat, during pregnancy. METHODS: After maiting, 12 nuliparous rats were divided into four groups with three animals for each group. The female rats corresponding to each group were killed at 1st, 7th, 14th and 21st days of pregnancy. RESULTS: Observation, on light microscopy (H.E) had at one view, did not display any alterations during pregnancy. However, the morphometry revealed that nuclei of cardiomyocytes are augmented in volume at 14th day of pregnancy and statistically significant data show the hypertrophy. CONCLUSION: The obtained data show the existence of a dinamic and reversible process of ventricular remodelling in consequence of adaptation physiological alterations of the cardiovascular normal pregnancy. PMID- 10513057 TI - [Evidence-based medicine]. AB - The growing array of diagnostic and therapeutic options available to the clinician has created the necessity of techniques, such as the randomized clinical trial, to evaluate their effectiveness. Recently, with the increasing penetration of the methods of clinical epidemiology into medical practice, the concept of an evidence-based medicine has arisen. Emphasizing the necessity of solid clinical evidence for clinical decision-making, evidence-based medicine provides a framework for the integration of research results into clinical practice. The evidence is graded, principally on the basis of study design, and norms are established as to what constitutes adequate evidence for clinical decision-making. The combination of this new paradigm of medical practice with the power of modern telecommunications is causing a revolution in the way medicine is practiced. Its integration into clinical practice is being facilitated by quantitative overviews of the literature, and the creation of clinical guidelines based on these reviews. Shortcomings of the traditional sources of evidence have been documented. New sources of evidence, such as the ACP Journal Club, and the Cochrane Collaboration, diminish dramatically the time required of clinicians to obtain the best available evidence. All health professionals should familiarize themselves with evidence-based medicine. PMID- 10513058 TI - [Mineral waters from several Brazilian natural sources]. AB - PURPOSE: To divulge information on the chemical composition and physical-chemical features of some mineral waters from Brazilian natural sources that will be of useful protocol investigation and patient advice. METHODS: The survey was based on bottle labels of non-gaseous mineral waters commercially available in the city of Rio de Janeiro. The ion concentration of each mineral was calculated from the salt content. RESULTS: 36 springs were enralled from different states of the country. The pH (25 degrees C), 4.1 to 9.3, varied on dependence of the source and it was linearey correlated with the cations calcium, magnesium and sodium and the anion bicarbonate. It was atributed to high alkalinity (about 70% of bicarbonate in the molecula-gram) of these salts. The calcium (0.3 to 42 mg/l), magnesium (0.0 to 18 mg/l) and bicarbonate (4 to 228 mg/l) contents are relatively low. CONCLUSION: The mineral content of the Brazilian springs enrolled in this survey is low; about 70% of the sources having calcium and magnesium less than 10 mg/l and 1.0 mg/l, respectively, similar to local tap water. PMID- 10513059 TI - [Transfusion-associated graft-versus-host disease guideline on gamma irradiation of blood components]. AB - Transfusion-associated graft-versus-host disease (TA-GVHD) is a rare and usually fatal syndrome. Clinical manifestations are fever, maculopapular skin rash, nausea, vomiting, diarrhea, hepatitis and pancytopenia owing to bone marrow hypoplasia. It can occur in recipients with severe immunosuppression and in immunocompetent recipients after transfusion of cellular components from HLA homozygous donor to recipients heterozygous for that HLA haplotype. The diagnosis is made by clinical manifestation and skin biopsy. Antithymocyte globulin and high dose systemic corticosteroids are both the most used therapy. The back of knowledge about this syndrome, the rapid evolution and the absence of treatment response are related to patients bad evolution. Gamma irradiation of blood products has been the mainstay of TA-GVHD prevention. Dose of 2500 cGy is required to completely inactivate T cells. Irradiation damage red cells membrane and the red celis units can not be storage for long time after irradiation. High potassium levels is the mainly change in red cells units. White cell-reduction filters do not prevent TA-GVHD and gamma irradiation does not prevent alloimmunization or blood reactions. Only cellular components like whole blood, red cells, platelets and granulocytes need be irradiated. Ali blood components should be irradiated to: first or second-degree relatives, patients need HLA matched platelets, recipients of allogeneic or autologous bone marrow transplantation, patients with Hodgkin's disease, patients treated with purine analogue drugs, intrauterine transfusion, pre-term infants and when congenital immunodeficiency states is suspected. It is recommended irrradiated blood to patients with neoplastic disease when they receive intensive chemotherapy. PMID- 10513060 TI - [Snoring and obstructive sleep apnea]. PMID- 10513061 TI - [Detection and treatment of chronic complications of diabetes mellitus: Consensus of the Brazilian Diabetes Society and the Brazilian Ophthalmology Council]. PMID- 10513062 TI - [Systemic talc granulomatosis in a HIV-negative intravenous drug addict]. AB - Anatomo-pathological correlation in a case of systemic talc granulomatosis affecting lungs, pleura, liver, spleen and mesenteric lymph nodes resulting in pulmonary arterial hypertension and cor pulmonale is described. The patient, a 26 year-old male HIV-negative intravenous drug addict had no lymphopenia or any histopathologic findings at necroscopy compatible with AIDS, despite of a chronic high-risk behavior favoring this illness. PMID- 10513063 TI - Isokinetic assessment of the flexor-extensor balance of the knee in athletes with total rupture of the anterior cruciate ligament. AB - The purpose of this study was to assess the flexor-extensor group of muscles of the knee in young athletes diagnosed with a total rupture of the anterior cruciate ligament (ACL). Eighteen knees of 18 athletes (14 men and 4 women) with an average age of 21.6 years (range 16-32 years) were assessed with a Cybex 6000 model isokinetic apparatus. The average internal between occurrence of the injury and assessment was 10.2 months (range 2-48 months). There was an associated meniscal injury in eight of the knees. Athletes with any other kind of associated injury, limitation, or blockage of the movement of the joint, significant pain during the exam, or interval between injury and exam of less than two months were excluded from the study. The parameters studied were the peak torque-velocity and flexor-extensor relationships at the constant angular velocities of 60 degrees/sec and 240 degrees/sec. Previous warming-up was done by means of an ergometric bicycle and adaptation with 3 submaximal repetitions. The contra lateral side, which presented no injury, was used as control. Peak torque (PT) at the constant velocity of 60 degrees/sec was greater than that at 240 degrees/sec for knees with and without injuries. However, there was no significant difference between the injured and uninjured sides at 60 degrees/sec or at 240 degrees/sec. The average value for the flexor-extensor relationship at 60 degrees/sec on the injured was 60% ((6), compared to 57% ((10) on the contra-lateral side. At 240 degrees/sec, the average value was 75% ((10) on the injured side, and 65% ((12) on the contra-lateral side. In conclusion, despite the complete rupture of the ACL of one knee, the average values for the flexor-extensor relationship were similar on the injured and uninjured sides at the velocity of 60 degrees/sec. As the velocity increased, an increase in the values for the flexor-extensor relationship of the knee also occurred, indicating a tendency of the performance of the flexor muscle group to approach that of the extensor muscle group, and this tendency was more pronounced on the side of the injury. PMID- 10513064 TI - Sodium serum levels in hypoalbuminemic adults at general medical wards. AB - Hypoalbuminemia may cause interstitial edema and hemodilution, which we hypothesized may influence serum sodium levels. Our purpose was to compare serum sodium levels of hospitalized adults with or without hypoalbuminemia. All sodium and albumin serum levels of 142 adults hospitalized at general medical wards over a six-month period were searched at a University Hospital mainframe computer. Relevant laboratory data and clinical details were also registered. Hypoalbuminemia was defined by serum albumin concentration < 3.3 g/dl Fisher, Mann-Whitney, and Student's t tests were applied to compare groups with or without hypoalbuminemia. Ninety-nine patients, classified as hypoalbuminemic, had lower blood hemoglobin (10.68 +/- 2.62 vs. 13.54 +/- 2.41), and sodium (135.1 +/- 6.44 vs. 139.9 +/- 4.76 mEq/l) and albumin (2.74 +/- 0.35 vs. 3.58 +/- 0.28 g/dl) serum levels than non-hypoalbuminemic (n = 43). Pearson's coefficient showed a significant direct correlation between albumin and sodium serum levels (r = 0.40) and between serum albumin and blood hemoglobin concentration (r = 0.46). Our results suggest that hypoalbuminemic adults have lower serum sodium levels than those without hypoalbuminemia, a phenomenon that may be at least partially attributed to body water retention associated with acute phase response syndrome. PMID- 10513066 TI - Determinants of mortality in oncology patients colonized or infected with Staphylococcus aureus. AB - Oxacillin-resistant Staphylococcus aureus (ORSA) infection is an important cause of hospital morbidity and mortality. The objective of this study was to identify the main factors associated with death in patients colonized or infected with Staphylococcus aureus in a cancer center. A matched-pair case-control study enrolled all patients infected or colonized with ORSA (cases) admitted to the Hospital do Cancer in Rio de Janeiro from 01/01/1992 to 12/31/1994. A control was defined as a patient hospitalized during the same period as the case-patients and colonized or infected with oxacillin-susceptible Staphylococcus aureus (OSSA). The study enrolled 95 cases and 95 controls. Patient distribution was similar for the two groups (p > or = 0.05) with respect to gender, underlying diseases, hospital transfer, prior infection, age, temperature, heart and respiratory rates, neutrophil count, and duration of hospitalization. Univariate analysis of putative risk factors associated with mortality showed the following significant variables: admission to the intensive care unit (ICU), presence of bacteremia, use of central venous catheter (CVC), ORSA colonization or infection, pneumonia, use of urinary catheter, primary lung infection, prior use of antibiotics, mucositis, and absence of cutaneous abscesses. Multivariate analysis showed a strong association between mortality and the following independent variables: admission to ICU (OR [odds ratio] = 7.2), presence of Staphylococcus bacteremia (OR = 6.8), presence of CVC (OR = 5.3), and isolation of ORSA (OR = 2.7). The study suggests a higher virulence of ORSA in comparison to OSSA in cancer patients. PMID- 10513065 TI - Megabladder in experimental Chagas disease: pathological features of the bladder wall. AB - Mega-organs, primarily in the digestive tract, are well known to occur in chronic Chagas disease. Acute experimental infection with Trypanosoma cruzi results in parasitism of a wide range of cells, tissues, and organs, including the urinary bladder. Infection of BALB/c mice with 100,000 bloodstream forms of the Y strain of T. cruzi induced acute infection with intense parasitism of all layers of the urinary bladder. Parasites were found in the mucosa, lamina propria, muscular, adventitial connective, and fat tissue. Desquamate epithelial cells with amastigotes in the bladder lumen were also found. After 60 days of infection, mice inoculated with 50 bloodstream forms developed dilated, thin-walled bladders that had inflammatory infiltrates and foci of fibrosis replacing areas of damaged muscular layer. These lesions result from direct damage to the muscle fibers by the T. cruzi, leading to myosites, muscle damage, and scarring. Direct damage of paraganglia cells secondary to parasitism, leading to dilatation, damage of muscle fibers, and scarring with replacement of muscular tissue with connective tissue, should also be considered as a cause of functional disturbance of the urinary bladder. PMID- 10513067 TI - Indicators of inflammation and cellular damage in chronic asymptomatic or oligosymptomatic alcoholics: correlation with alteration of bilirubin and hepatic and pancreatic enzymes. AB - Biochemical and hematimetric indicators of inflammation and cell damage were correlated with bilirubin and hepatic and pancreatic enzymes in 30 chronic male alcoholics admitted into psychiatric hospital for detoxification and treatment of alcoholism. Aspartate aminotransferase, alanine aminotransferase, gamma glutamyltransferase, alkaline phosphatase, and total bilirubin were altered, respectively, in 90%, 63%, 87%, 23% and 23% of the cases. None of the indicators of inflammation (lactic dehydrogenase, altered in 16% of the cases; alpha-1 globulin, 24%; alpha-2 globulin, 88%; leucocyte counts, 28%) was correlated with alterations of bilirubin or liver enzymes. Lactic dehydrogenase was poorly sensitive for detection of hepatocytic or muscular damage. Alterations of alpha globulins seemed to have been due more to alcohol metabolism-induced increase of lipoproteins than to inflammation. Among indicators of cell damage, serum iron, increased in 40% of the cases, seemed to be related to liver damage while creatine phosphokinase, increased in 84% of the cases, related to muscle damage. Hyperamylasemia was found in 20% of the cases and significantly correlated with levels of bilirubin, alkaline phosphatase and gamma-glutamyltransferase. It was indicated that injuries of liver, pancreas, salivary glands, and muscle occurred in asymptomatic or oligosymptomatic chronic alcoholics. PMID- 10513068 TI - Reflux esophagitis and gastroesophageal reflux disease: a cross-sectional study of gastroesophageal reflux disease patients by age group. AB - The purpose of this study was to explore the relationship between the intensity of acid reflux and severity of esophageal tissue damage in a cross-sectional study of patients with gastroesophageal reflux disease (GERD). Seventy-eight patients with were selected in accordance with the strict 24-hour ambulatory esophageal pHmetry (24h-pHM) criteria and distributed into three age groups: Group A: 14-24 years of age. Group B: 25-54; and Group C: 55-64. The 24h-pHM was carried out in accordance with DeMeester standardization, and the Savary-Miller classification for the diagnosis of reflux esophagitis was used. The groups were similar in 24h-pHM parameters (p > 0.05), having above normal values. For the study group as a whole, there was no correlation between age group and intensity of acid reflux, and there was no correlation between intensity of acid reflux and severity of esophageal tissue damage. However, when the same patients were sub grouped in accordance with the depth of their epithelial injury and then distributed into age groups, there was a significant difference in esophagitis without epithelial discontinuity. Younger patients had less epithelial damage than older patients. Additionally, although there was a significant progression from the least severe to the moderate stages of epithelial damage among the age groups, there was no apparent difference among the age groups in the distribution between the moderate stages and most severe stages. The findings support the conclusion that the protective response of individuals to acid reflux varies widely. Continued aggression by acid reflux appears to lead to the exhaustion of individual mechanisms of epithelial protection in some patients, but not others, regardless of age or duration of the disease. Therefore, the diagnosis and follow up of GERD should include both measurements of the quantity of refluxed acid and an assessment of the damage to the esophageal epithelium. PMID- 10513069 TI - Clinical and radiological aspects in Melnick-Needles syndrome. AB - Melnick-Needles syndrome is an X-linked dominant bone dysplasia, lethal in males, characterized by a typical facies and characteristic radiological findings: including sclerosis of skull base and mastoids. S-shaped appearance of tibia; cortical irregularities with a ribbon appearance of the ribs. About 48 well documented cases have been reported, most of them were sporadic. Parental transmission has been published in only 11 kindreds. We are presenting the first Brazilian family with mother-daughter transmission. The proposita presented the typical clinical and radiological features with characteristic facies, severe thoracic cage restriction and pulmonary hypertension. Her mother was more mildly affected. PMID- 10513070 TI - [Current life expectancy in Chile]. AB - An increased life expectancy is associated to a nation's progress. Life expectancy at birth is 78 years in developed countries, 72 in Latin America, 70 in China, 63 in Eastern Europe and 51 in Africa. Chile is the Latin American country with the higher increment in life expectancy, reaching 75 years at birth, during the period 1990-95. A higher life expectancy is not always an advantage. People over 60 years old may survive with a burden of chronic diseases or physical and mental disability. The higher health care needs of the increasing population of elderly people is raising the costs of health care in modern nations. PMID- 10513071 TI - [Transcranial cytokine gradient and intestinal permeability in acute severe brain injury]. AB - BACKGROUND: Acute brain injury is associated with a bimodal hypermetabolic state probably caused by cytokine secretion and high hormone and catecholamine concentrations. In a first stage, the brain would produce these substances and afterwards, another production source, most probably the splanchnic territory, would perpetuate the hypermetabolic state. AIM: To investigate the cytokine production source and to assess intestinal permeability in acute brain injury in the absence of cerebral ischemia and systemic oxygen deficit. PATIENTS AND METHODS: Arterial systemic and cerebral venous bulbar interleukin 1 beta and interleukin 6 levels were measured during the first seven days of evolution in 15 patients with acute brain injury. Serum lactate, the oxygen/lactate ratio, gastric intramucosal pH and intestinal permeability using the lactulose/mannitol test were also assessed in the same period. RESULTS: High arterial and venous interleukin 1 beta and interleukin 6 levels were detected. A positive gradient for interleukin 6 levels was detected throughout the study period with normal intramucosal pH, lactate and oxygen/lactate ratio. There was also an early impairment of intestinal permeability in these patients. CONCLUSIONS: High arterial and venous cytokine concentrations were detected in patients with acute brain injury. The positive gradient for interleukin 6 suggests a brain origin for this cytokine. Intestinal permeability is also altered in these patients. PMID- 10513072 TI - [Non-invasive mechanical ventilation in patients with severe stable COPD]. AB - BACKGROUND: The benefits of non-invasive mechanical ventilation (NIMV) in hypercapnic patients with severe stable COPD remain controversial mainly due to their unknown mechanisms. AIM: To assess the clinical and physiological benefits of a 3 weeks period of intermittent NIMV and their underlying mechanisms in COPD patients. PATIENTS AND METHODS: Twelve patients (10 male) prospectively recruited (age 65 +/- 3 years, FEV1 27 +/- 2% predicted, PaO2 46 +/- 2 mmHg, PaCO2 55 +/- 2 mmHg) were submitted to NIMV using a commercially available system (BiPAP) 3 h a day, 5 days a week for 3 weeks. Arterial blood gases, 6 min walking distance, dyspnea (Mahler's scale), breathing pattern, PIMax, ventilatory drive (P0.1) and the impedance of the respiratory system (P0.1/V1/T1) were measured before and after NIMV. RESULTS: A significant improvement in PaO2, PaCO2, PIMax, dyspnea and exercise capacity was observed in addition to a trend for VT to increase and for respiratory rate (RR) to decrease. The impedance of the respiratory system showed a significant reduction. Ventilatory drive, normalized for PaCO2 levels, did not change. Improvement in PaCO2 was related to an increase in Vp whereas a significant association between the reduction in RR and the fall in respiratory system impedance was also found. CONCLUSIONS: Our study supports previous data demonstrating that NIMV improves clinical and physiologic parameters in advanced stable COPD and suggest that the underlying mechanism is a reduction in the inspiratory load. A randomized clinical trial is needed to confirm that this mechanism is operative. PMID- 10513073 TI - [Esophageal atresia and associated malformations]. AB - BACKGROUND: Survival of newborns with esophageal atresia and tracheoesophageal fistula has increased in the last years. AIM: To assess the prevalence of esophageal atresia and describe associated malformations in Chilean newborns. MATERIAL AND METHODS: All births occurring between January 1983 and June 1998 were studied. All malformed children were registered and the next non malformed born child was considered as control. RESULTS: During the study period, 50,965 births occurred and 3,336 malformed children were born. Eighteen (3 stillborn) had esophageal atresia with a rate of 3,53 per 10,000 born alive. Overall survival was 73%. Survival among children classified in Waterson groups A and B was 100% and 50% among those classified in group C. Seventy two percent had associated malformations, being congenital cardiopathies and skeletal malformations the most frequent. VACTERL association was found in 44% of children. All stillborn children had other severe malformations. When compared to controls, malformed children had a lower weight, a lower gestational age, their mothers had a higher age, a higher frequency of relatives with malformations and a higher frequency of maternal diseases during the first trimester of pregnancy. CONCLUSIONS: The rate of esophageal atresia found in this study is similar to that reported in other Chilean obstetrical units as part of the Latin American Study of Congenital Malformations (ECLAMC). PMID- 10513074 TI - [Acute effect of dobutamine and amrinone on hemodynamics and splanchnic perfusion in septic shock patients]. AB - BACKGROUND: Vasoactive drugs used in the reanimation of septic patients, can modify splanchnic perfusion. AIM: To compare the effects of dobutamine and amrinone on gastric intramucosal pH (pHi), lactate levels and hemodynamics in surgical patients with compensated septic shock. PATIENTS AND METHODS: Fourteen postoperative patients with abdominal sepsis and compensated septic shock (pHi < 7.32 or lactate > 2.5 mmol/l) were studied in a prospective, randomized, unblinded study. Patients were randomized to receive (Group 1, n = 7) dobutamine at 5 micrograms/Kg/min or (Group 2, n = 7) amrinone at 5 micrograms/Kg/min. Hemodynamic data, arterial lactate and pHi were measured before and 30, 60 and 120 minutes after starting drug infusion. RESULTS: Both drugs were associated with a decrease in lactate levels. Dobutamine infusion, but not amrinone, increased gastric pHi, as well as cardiac index and oxygen delivery. CONCLUSIONS: An improvement in gastric pHi associated with an increase in oxygen delivery, was observed with dobutamine. Amrinone showed no effect at the fixed, low dose used in the study. PMID- 10513075 TI - [Detection of thyroglobulin autoantibodies and potential interference with serum thyroglobulin measurement]. AB - BACKGROUND: Thyroglobulin measurement is useful for the follow up of patients subjected to total thyroidectomy for differentiated thyroid carcinoma. Thyroglobulin autoantibodies may interfere with its determination. AIM: To measure thyroglobulin autoantibodies and their interference with thyroglobulin determination. MATERIAL AND METHODS: The presence of thyroglobulin autoantibodies was investigated in 801 serum samples sent to the laboratory for measurement of thyroglobulin levels. A serum was considered positive for these autoantibodies when radioactivity corresponding to 125I-thyroglobulin bound to thyroglobulin autoantibodies, precipitated with human gamma globulin, exceeded in 1.4 times that of a negative sera pool. In positive sera, thyroglobulin autoantibody concentration was measured and its interference with thyroglobulin radioimmunoassay was assessed through a recuperation test using exogenous thyroglobulin. RESULTS: Thyroglobulin autoantibodies were detected in 149 sera (18.6%). Of these, 65 had a recuperation that fluctuated between 1 and 80%. Thyroglobulin autoantibody concentration was negatively correlated with recuperation percentages (r = -0.64; p < 0.001) but not with thyroglobulin concentrations (r = 0.08). Thyroglobulin was higher in positive sera with a recuperation over 80% than in sera with a recuperation of less than 80% (12.7 +/- 1.7 and 5.9 +/- 0.6 ng/ml, respectively; p < 0.001). CONCLUSIONS: Thyroglobulin autoantibodies interfere with thyroglobulin measurement by radioimmunoassay, sequestering variable amounts of thyroglobulin. The presence of these autoantibodies must be investigated prior to thyroglobulin determination. PMID- 10513076 TI - [Factors which determine the psychological adjustment to permanent colostomies. An empirical study in Santiago, Chile]. AB - BACKGROUND: The acceptance of a colostomy by a patient requires a great amount of psychological resources and social support. AIM: To identify factors that influence the postoperative adaptation to a colostomy. PATIENTS AND METHODS: Sixty patients subjected to a colostomy in 5 hospitals in Santiago were interviewed. The adaptation to the procedure was assessed using the Olsbrisch Ostomy Adjustment Scale. Three regression equations were elaborated to determine the main predictors associated to adaptation. Control variables such as sex, age and the lapse between the interview and the surgical procedure, were included in the model. RESULTS: The model used identified the simultaneous and combined effects of socioeconomic variables such as education and income on the adaptation to colostomy. A generalized detriment of the self image was detected. Patients valued the social support given by the family and friends. Multiple regression analysis determined that the main predictor of the patient's adaptation to colostomy was the level of self care developed. CONCLUSIONS: The better adaptation to a colostomy could be achieved by training the patient for an adequate self care and providing him with psychological and social support. PMID- 10513077 TI - [Esophagogastric varix hemorrhage. Experience with cyanpoacrylate and polidocanol in 68 patients with active hemorrhage]. AB - BACKGROUND: Sclerosis, injection of cianoacrylate and rubber band ligation are the most commonly used endoscopic techniques for the treatment of bleeding esophageal varices. AIM: To assess the effectiveness of cianoacrylate and polidocanol in the treatment of bleeding esophageal varices. PATIENTS AND METHODS: Sixty eight patients with active variceal bleeding were studied. Bleeding varices were classified as thin, thick or gastric. Bleeding from thin varices was treated with polidocanol. Bleeding from thick or gastric varices was treated with cianoacrylate. Variceal eradication was done with polidocanol. RESULTS: Bleeding came from thin esophageal varices in 23% of patients and endoscopic treatment stopped bleeding in 95% of them, from thick esophageal varices in 62% and endoscopic treatment was successful in 94% of these, and from gastric varices in 12% and treatment stopped bleeding in 87% of these (in 3% bleeding was considered subcardial). Twenty-five percent of patients bled again during variceal eradication, 12% died due to uncontrollable bleeding and 20% died due to liver failure. During variceal eradication 59% of patients classified as Child Pugh C, died. CONCLUSIONS: Treatment of bleeding esophageal varices with cianoacrylate or polidocanol is effective. Patients classified as Child Pugh C have a had prognosis. PMID- 10513078 TI - [Vascular accesses for chronic hemodialysis in children]. AB - BACKGROUND: The success of a chronic hemodialysis program depends on a good vascular access. AIM: To evaluate the experience with vascular accesses for chronic hemodialysis in pediatric patients. PATIENTS AND METHODS: One hundred fifty-one vascular accesses used in 60 pediatric patients (33 female) coming from 2 hemodialysis (HD) centers were analyzed. RESULTS: The average age of admission to the hemodialysis program was 10 years old (range 1.8-15). Forty percent of accesses were internal arterio-venous fistulae (AVF), 58% were central venous catheters and 2% were grafts. Twenty four patients required a central venous catheter from the beginning since they required immediate dialysis. Twenty patients began dialysis with a permeable AVF and never required another vascular access. Eight small children used a central catheter as the only vascular access and 32 patients required both types of vascular accesses. Eighty-seven catheters were used in 34 patients, of which 77 were temporary and 10 permanent. Seventeen patients needed only one catheter and one girl required 15 catheters. The average life span for AVF was 524 days (20-1277), for temporary catheters 34 days (1-76) and for permanent catheters 73 days (9-147). Two years survival of AVF was 95%. One month survival for subclavian and jugular catheters was 50%. Fifty-six percent of AVF had no complications, 12 failed due to insufficient flow and 24% had a complication. Of the 87 catheters placed, 75% had complications and 22 were electively removed. CONCLUSIONS: Arteriovenous fistula is the vascular access of choice for hemodialysis in pediatric patients. Central venous catheters can become an essential access, specially in small children. PMID- 10513080 TI - [Colonic perforation secondary to ileocecal tuberculosis. Report of one case]. AB - We report a 44 years old male, presenting with an eight months history of right lower quadrant pain, diarrhea and weight loss. Colonoscopy showed a proliferative and ulcerated lesion in the cecum, with necrotic areas. Barium enema showed an extensive irregular stenosis with rigidity of cecum and ascending colon. The endoscopic biopsy showed numerous granulomas with giant multinucleated cells of Langhans type. A right colectomy was performed with a good postoperative evolution. Anti tuberculosis treatment was started two weeks later and was well tolerated. The patient is currently asymptomatic after two years of followup. PMID- 10513079 TI - [Experience with laparoscopic surgery for adnexal masses at the Regional Hospital of Temuco, Chile]. AB - BACKGROUND: Laparoscopic surgery has clear advantages over open surgical procedures. In gynecology, laparoscopic surgery for adnexal masses in pre or post menopausal women has been used for several years. AIM: To report the experience with gynecologic laparoscopic surgery at the Temuco Regional Hospital. PATIENTS AND METHODS: Between 1996 and 1998, laparoscopic surgery was done in 96 patients aged 16 to 56 years and open surgery in 56 patients aged 15 to 74 years, with a clinical or ultrasound diagnosis of adnexal masses or ovarian dermoid cysts. RESULTS: The most frequent tumors excised were epithelial and germinal cell. Laparoscopic surgery required a mean operative time of 69.9 min and it had a 3.1% of complications. Women subjected to this type of surgery had a mean hospital stay of 3.1 days and the mean postoperative stay was 2 days. Open surgery required an operative time of 69 min and it had no postoperative complications. The mean hospital stay for women subjected to this type of procedure was 9.5 days. CONCLUSIONS: Women subjected to laparoscopic surgical procedures for adnexal masses had a shorter hospital stay than women subjected to open surgical procedures. PMID- 10513081 TI - [Retraction of lower limbs in a female with hyperthyroidism]. AB - We report a 47 years old woman with hyperthyroidism that had a severe tendinous retraction of hips and knees that subsided with propylthiouracil treatment. Electrodiagnosis showed myopathic alterations and muscle strength was moderately reduced. The authors did not find references of a similar condition in patients with hyperthyroidism. PMID- 10513082 TI - [Rhinocerebral mucormycosis. Report of four cases]. AB - We report four diabetic patients (one male) aged 64, 63, 61 and 77 years old with rhinocerebral mucormycosis. Three had an acute and one a chronic form of the disease. The chronic form was diagnosed with a biopsy of the nasal cavity. Three patients received amphotericin but all died. The diagnosis of the disease must be suspected in diabetics or patients with some type of immune depression. PMID- 10513083 TI - [Effect of catecholamines on splanchnic perfusion in sepsis]. AB - Splanchnic ischemia is frequent in sepsis and septic shock and is related to impairment in intestinal permeability, derangement in mucosal barrier functions and translocation of proinflammatory mediators. These changes can contribute to the pathogenesis of multiple organ failure. Vasoactive drugs such as dobutamine and dopexamine can improve splanchnic perfusion and gastric intramucosal pH during sepsis. However, contradictory results have been obtained with dopamine and norepinephrine. On the other hand, epinephrine further impairs splanchnic perfusion. In view of the contradictory effects of different vasoactive drugs, gastric tonometry must be measured during their use, to find the optimal drug combination that optimizes splanchnic blood flow. PMID- 10513084 TI - [Prevalence of hypertension in Valparaiso. Results of the base survey of the CARMEN project (Set of measures for the multifactorial reduction of non transmissible diseases)]. AB - BACKGROUND: There is little information about the real prevalence of hypertension in Chile. AIM: To assess the adjusted prevalence of hypertension and its main therapeutic measures among adults living in Valparaiso, Chile. MATERIALS AND METHODS: A random sample of dwellings in Valparaiso was chosen. Among these, an individual of 25 to 64 years old was randomly surveyed for risk factors for chronic diseases and sociodemographic parameters. Blood pressure, weight, height, oral glucose tolerance test, fasting cholesterol and triglycerides were also measured. Prevalence was pondered according to age, sex, and probability of selection in the dwelling interior. RESULTS: Three thousand one hundred twelve individuals were studied. The adjusted prevalence of hypertension was 11.4% (11.6% among females and 10.6% among men). The prevalence increased along with age from 3 and 1.9% in men and women of 25 to 34 years old respectively, to 18.2 and 27.4% among men and women of 55 to 64 years old (p < 0.01). People of low socioeconomic level had a higher prevalence of hypertension than those of high socioeconomic level (14.2 and 9.3% respectively, P < 0.05). Diabetes, obesity and hypercholesterolemia were significantly more frequent in subjects with hypertension than in the general population. Forty-four percent of diagnosed hypertensives were receiving medications (angiotensin converting enzyme inhibitors 40%, calcium antagonists 34%, beta blockers 22%). Twenty five percent of patients were treated with a combination of medications. Of those treated, only 22% had normal blood pressure levels at the moment of examination. CONCLUSIONS: High blood pressure is an important public health problem that requires more efficient detection and treatment programs. PMID- 10513085 TI - [Advances in diagnosis and treatment of pancreatic carcinoma]. PMID- 10513086 TI - [The physician and death]. AB - This was a lecture given by Monsignor Pinera, Honorary Member of the Chilean Academy of Medicine who is a Bishop of the Catholic Church and also an MD. The author addresses the problem of the dying patient in the different positions held by a churchman and by the physician. The religious minister, whether Jewish, Catholic or from any other Christian nomination, helps the dying patient to understand his her situation as an inevitable step towards Sheol (in the Old Testament) or Heaven (New Testament). The physician confronts the problem of a terminal disease or a fatal event. But in both positions, hope must be sustained to the very end and due respect to the patient's will as a human being has to be maintained, helping him her to die with dignity and the best comfort available. Unnecessary or unjustified measures to resuscitate or to prolong life are criticised. PMID- 10513087 TI - [Development of a scale for quality of care management process. A Delphi survey and studies on reliability and validity]. AB - The purpose of this study was to develop a measurement tool for assessing quality in care management and to test its reliability and validity. A Delphi survey was initially administered on 96 experienced community health nurses, to improve the content validity of a questionnaire that was developed after three repeated rounds of data collection and content analysis. A total of 353 community health nurses, from 121 cities and towns across Japan, completed the mailed questionnaire. Cronbach's alpha value was more than 0.8, respectively, for all items in questionnaires, and for each factor, indicating internal consistency in reliability. Five factors were identified through factor analysis using a principal factor method with varimax rotation. These factors were good reflections of components classified by some researchers, indicating construct validity. In addition, care managers were grouped according to such criteria as their age and work experience. The QCM-P (Quality of Care Management-Process Measurement) score of each group was compared, as theoretically differences are expected. The questionnaire's validity was evidenced by significant differences in the QCM-P score among each group. Further studies on criterion-related validity and stability which relates to reliability are required. Thus, although further work is needed, QCM-P was found to have both reliability and validity at a permissible level as a scale for measuring the quality of care management process. PMID- 10513088 TI - [Economic evaluation of health care program for hepatitis C virus antibody screening]. AB - We made a trial of introducing a health care program for hepatitis C virus (HCV) antibody screening in Saga prefecture, where mortality rate of hepatoma is one of the highest in Japan. The program started in 1992 and covered nearly the entire population older than 30 years in this prefecture. This was the first implementation in Japan. In the present study, we performed a cost-benefit analysis of this program. It included three steps; early detection of hepatitis C cases in the free-living population; implementation of interferon (IFN) therapy for detected chronic active hepatitis cases; follow-up of the cases who underwent IFN therapy. We counted, as cost, expenditure for primary screening, that for subsequent clinical examinations, that for IFN therapy, that for follow-up of detected HCV carriers, and loss of earned income during a leave of absence from work for the IFN therapy. On the other hand, we counted as benefit the medical expenditures saved, and gained earned income by reduction of hepatoma, cirrhosis and hepatitis due to medical intervention, both of which would have sustained losses had the health care program not been implemented. The employed model assumed age and gender specific natural histories for hepatitis C. The benefit/cost ratio was found to be in 1.71 to 2.32, suggesting economic validity of the proposed health care program. We further evaluated the economic validity by sensitivity analysis by changing rates of HCV carriers, discounting rates, rates of "complete responder" of chronic active hepatitis cases to IFN therapy, and detection rate of otherwise submerged chronic active hepatitis cases. Benefit/cost ratios were found to be greater than unity, given that the population rate of HCV carriers is higher than 1%. PMID- 10513089 TI - [Knowledge about AIDS and risk behaviors among hill tribes in northern Thailand]. AB - Two hundred eighty-one Hmong, 240 Mien, and 210 Lahu hill tribe people (ages between 17-39) in northern Thailand were interviewed from March to May 1996 about their knowledge, beliefs, attitudes regarding AIDS, STDs and condoms, and sexual behavior, to guide future tribal AIDS education and care programs. Premarital sex for both men and women and extramarital sex for men seems to be still culturally permissive and wide-spread among the Hmong and Mien people. The Lahu reported lower cultural acceptance, but this group had higher levels of premarital and extramarital sex than the other two groups, excepting extramarital sex for Hmong men. Many men among the three groups knew about condoms, but they do not like to wear condoms because, "it is not natural." Lahu people had the least education, were the least literate, and had the least family income. In addition, Lahu had the lowest level of knowledge of AIDS and STDs and they seem to have the most risky behaviors among the three groups. On the other hand, although Mien people had the highest education, were the most literate, had the most family income, and had the highest level of knowledge of AIDS and STDs, they seem to have high risk sexual behavior, also. There were increasing numbers of hill tribe people migrating to the cities and becoming involved to a greater degree in seasonal or permanent wage labor, including the commercial sex industry among Lahu and Mien groups. Hmong seem to have the lowest number of workers migrating to the cities. The results indicate the need to inform and educate tribal people about AIDS in a socio-cultural specific context. PMID- 10513090 TI - [Implication of health checkups of students from developing countries in Japan]. AB - We investigated the prevalence of peripheral blood abnormalities, parasitic infestation, and hepatitis virus infection, by using the results of the primary screening health checkups for 423 students (male: 317, female: 106, average age +/- SD: 34.2 +/- 5.5 year-old) from abroad. Most of them were from Southeast Asia, Africa, Central and South America, and other developing countries in tropical or subtropical areas. Thalassemia-like hematological disorders, showing microcytic peripheral red blood cells without any anemia, were seen in 7.6 percent of the students, and intestinal parasites were revealed in 12.7 percent of them. The positive rate for anti-hepatitis A virus antibodies (84.3%) and the exposure rate of hepatitis B viruses (35.3%) were similar to previous reports. Compared with the positive rate for anti-hepatitis C virus antibodies (anti HCVAb) of students from other regions (1.5%), a significantly high seropositivity for anti-HCVAb was encountered in Egyptian participants (21.1%). In recent years, population shifts and rapid transportation have facilitated the spread of certain infectious diseases from endemic to non-endemic areas. International preventive strategies, education of people regarding infectious diseases, and sufficient medical staffs for this purpose are urgently recommended. PMID- 10513091 TI - [A survey of smoking behavior among junior high school students and smoking prevention education developed using the survey results]. AB - In June 1996 and 1997, a self-administered anonymous questionnaire survey of smoking behavior was conducted on students (118 boys and 135 girls) of two junior high schools in Kami-gun, Kochi Prefecture. Following the surveys, smoking prevention education classes were held using the survey results as material for education. The survey results were as follows; 1. The percentage of ever-smokers was higher in boys than in girls, 37.9% and 22.8% respectively. The percentage of those who smoked at least one cigarette during the past one month was 2.9%, and the rate increased with school grade. 2. Most commonly, first experience of smoking was between the 4th to 6th grades in elementary school, and the main motive for first smoking was curiosity. 3. The association between smoking and lung cancer was the most widely known, and that between smoking and intrauterine growth retardation was relatively well-known among students. More than 90% of students clearly understood the harmfulness of passive smoking. 4. The results of logistic regression analyses revealed that friends' and parents' smoking were significant correlates of smoking experience in boys. Among girls no significant relationship was found. The content of smoking prevention education class consisted of quizzes about smoking, an explanation of the survey results, an exhibition of a cross-sectional model of the lung, an experiment of smoking using a small doll, an analysis of expired air carbon monoxide concentration, and an explanation of the harmfulness of smoking. The reports of the students after the class showed that not only the explanation of the harmfulness of smoking, but also the experiment or explanation of the survey results appeared to be effective educational content. PMID- 10513092 TI - [Evidence-based medicine and 'The Cochrane Collaboration']. AB - In Evidence-Based Medicine (EBM), a clinical decision is based neither on pathophysiological theories nor personal experience but on the results derived from scientifically designed clinical epidemiological studies (i.e., evidence). EBM is used in various clinical applications, such as therapy, diagnosis, and prognosis prediction. The process includes (1) asking a clinical question consisting of the three elements of "patient", "exposure", and "outcome"; (2) searching for the best evidence using MEDLINE or Cochrane Library; (3) appraising critically the validity of the method and the magnitude and probability of the result; and finally (4) applying the evidence of the patient. In actual clinical practice, clinical expertise and patient preferences should be as much regarded as research evidence. 'The Cochrane Collaboration' supplies systematic reviews of clinical trials carried out all over the world to its consumers. Its fruit, 'The Cochrane Library (CD-ROM),' is a highly valuable resource. 'The Cochrane Collaboration' serves as the infrastructure for EBM. EBM, which was originally developed for the individual patient care, can also be applicable to community- or workplace-healthcare and policy making by governments. Thus, EBM is both a philosophy and a method to provide people with the most appropriate medical practice. PMID- 10513093 TI - The use of acupuncture-like electrical stimulation for wound healing of lesions unresponsive to conventional treatment. AB - Based on previous experimental evidence suggesting improved healing of wounds treated with electrical stimulation, we conducted a clinical trial with patients seeking alternative medicine after unsuccessful conventional medical treatment. Electricity was delivered in two forms: (1) For wounds with extensive loss of tissue and/or those that had failed to heal spontaneously, electrical stimulation was delivered via subcutaneously inserted needles surrounding the wound edges and applying a dose charge of 0.6 coulombs/cm2/day; (2) in second degree burn injuries, lesions were covered with gauze soaked in a 10% (w/v) sterile saline solution and the same dose of electricity was applied as for (1). Forty-four patients were treated with electrical stimulation of the skin; 34 in group (1) and 10 in group (2). Following electrostimulation in all patients in both groups healing proceeded in a thoroughly organized manner, almost regardless of the severity of the type of wound or burn treated. Advantages and limitations of this technique are discussed. PMID- 10513094 TI - The treatment of chronic inflammatory demyelinating polyradiculoneuropathy with acupuncture: a clinical case study. AB - Chronic inflammatory demyelinating polyradiculoneuropathy (CIDP), also referred to as chronic relapsing Guillain-Barre syndrome, is a rare neurological disease characterized by progressive symmetrical motor and sensory loss. Current biomedical treatments for this disease are often only short term in nature and have limitations with respect to side-effects and cost. The authors review CIDP from the perspectives of Western biomedicine and traditional Chinese medicine. A case report of a 17-year-old female diagnosed with CIDP and treated successfully with acupuncture is reviewed. PMID- 10513095 TI - Case report: successful treatment of varicose veins with acupuncture. AB - This case report describes the treatment of varicose veins of the lower leg and their complete resolution by acupuncture. According to the author, there is no description of this condition and no guidelines for diagnosis and treatment in the literature of Chinese medicine and acupuncture. PMID- 10513096 TI - A comparative study on the treatment of migraine headache with combined distant and local acupuncture points versus conventional drug therapy. AB - According to Chinese medicine, the differential diagnosis of migraine headache may be classified based on the state of the viscera, channels and collaterals. In this study, acupuncture treatment prescriptions combining distant and local acupoints were selected according to the differential diagnosis. Sixty-four patients were divided into two equal groups: one group received acupuncture, and the other group underwent conventional drug treatment. RESULTS: The efficacy rates in the acupuncture and control groups were as follows, respectively: Cure: 75% versus 34.4%, marked improvement: 18.8% versus 28.1%, no effect: 6.3% versus 37.5%. The overall effective rates for the acupuncture and control groups were 93.8% and 62.5%, respectively, indicating a significantly greater effect in the acupuncture group (P < 0.01, x = 13.475). PMID- 10513097 TI - Primary Parkinson's disease: the use of Tuina and acupuncture in accord with an evolving hypothesis of its cause from the perspective of Chinese traditional medicine--Part 2. AB - In Part II of this series, the author continues the discussion of a hypothesis based on acupuncture channel theory regarding the cause of Parkinson's disease (PD). A system of Yin-type Tuina, termed FSR (forceless spontaneous response) evolved from this theory; its clinical application has resulted in various degrees of relief from PD symptoms, (e.g., tremor, rigidity, decreased dyskinesia; improved balance, circulation) regardless of the stage of the disease, and in several cases enabled a reduction of conventional medication. This installment includes basic instruction in the technique, and a discussion of the dyskinesia which occurs during restoration of Qi to the Large Intestine and Stomach channels during treatment. A case study chronicles the weekly changes in symptoms typically experienced by patients during FSR therapy. PMID- 10513098 TI - Energetics and transformation: insights on the paradoxical opportunity presented by chronic illness and pain--Part III. AB - Following the author's own experiences with chronic pain due to a motorcycle accident during medical school, and the pain's unresponsiveness to conventional medicine, he sought other solutions. His journey led him to a new understanding of health and illness. With these insights and through the use of acupuncture, bodywork and various breathing techniques, he began to see phenomena--emotional release, myoclonic shaking, and regression--and healing that could not be explained in terms of a rational or structural framework. He posits that such phenomena represent different forms of de-stressing which together serve to release "blocked feeling," which he suggests is the "energy block" described by acupuncture theory as "stagnant Qi." The third in a series, this installment explores the issue of "disempowerment" as one of the fundamental energetic imbalances of illness. PMID- 10513099 TI - Acupoint energetics of nutritional supplement intolerance: patient self prescribing may impair clinical progress. AB - Chronically ill patients failing to obtain effective treatment with conventional medicine often self-prescribe multiple nutritional supplements. In some cases, supplements can exacerbate pathology or block therapeutic progress because of negative effects on gastrointestinal, immunological, neurotransmitter, and hormonal functions. Nutritional supplements may also contribute to the growth of bacterial, viral, and fungal pathogens. Case studies involving evaluation of supplement energetics through acupoint biophoton coherence measurements at Pericardium-6 reflect the potential for harm from patient self medication. PMID- 10513100 TI - Colored light therapy: overview of its history, theory, recent developments and clinical applications combined with acupuncture. AB - Light therapy has a long history, dating from ancient Egypt to the contemporary treatment of seasonal affective disorder. In the early half of this century, Dinshah Ghadiali, MD PhD, refined a sophisticated system of color therapy. Influenced by a strong background in mathematics and physics, he determined specific "attributes" of the colors of the spectrum, i.e., their specific effects on human physiology. Later research has confirmed many of his concepts and spawned evolution of new systems for application of light therapy including irradiation of acupuncture points. According to the author, his system dovetails nicely with traditional Oriental medicine theory, relating colors to the internal organs and meridian system. Of particular note is recent Russian research which has shown that light is conducted within the body along the acupuncture meridians leading the authors to ponder: Do acupuncture meridians function as a light (photon) transferal system within the body, not unlike optical fiber? Case studies provide support for the clinical benefits of light therapy. The emerging contemporary color therapy systems of Mandel (Colorpuncture) and McWilliams (Chromo-pressure) are discussed, and a newly patented device is introduced. PMID- 10513102 TI - Issues in acupuncture research: the failure of quantitative methodologies and the possibilities for viable, alternative solutions. AB - Thirty years of active acupuncture research have failed to unequivocally demonstrate its clinical efficacy. Certain characteristics of acupuncture are difficult to fit into an experimental study. Many researchers mention selection of appropriate controls, single- or double-blind research design, and application of relevant outcome measures as areas causing most difficulties. Also cited are the variability of acupuncture techniques, difficulty of standardizing acupuncture treatments, inadequate population size, significant variability of response to treatments, the use of a distinctive terminology, and importance of practitioner's experience. Acupuncture and Chinese traditional medicine are based on a unique philosophical model, and the instruments of biomedical research may be inadequate and inappropriate. In contrast to the quantitative experimental method, introspective self-observation and qualitative observation are offered as a means of studying the effectiveness of acupuncture. PMID- 10513101 TI - A review of recent research studies on the efficacy of Esogetic Colorpuncture Therapy--A wholistic acu-light system. AB - This article reviews recent studies conducted in Europe which sought to evaluate the effectiveness of Peter Mandel's Esogetic Colorpuncture Therapy (ECT). These investigations addressed the use of specific ECT therapies for treating a variety of difficult health problems: migraines, childhood insomnia, bronchitis, ADD or learning disorders, and uterine fibroids. Limitations in research design and sample size necessitate that these studies be viewed as pilot or preliminary research. However, in all the studies, the findings showed dramatic improvement of symptoms after ECT treatments. This suggests that ECT may offer fast, economical, non-invasive and non-toxic methods for treating the selected health problems and that ECT continues to show promise as a powerful new method of wholistic healing. PMID- 10513103 TI - Toward improving the reliability of clinical acupuncture trials: arguments against the validity of "sham acupuncture" as controls. AB - In refining the design of single-blind acupuncture research trials, "sham acupuncture" has been proposed and used in trials as a useful "inactive" control, yet the degree of inactivity with sham acupuncture points remains largely undetermined. With neural and dermal effects occurring whenever a needle is inserted into the skin, regardless of depth or location, the estimated effect upon Qi when needling sham acupuncture points remains a matter of conjecture. This paper examines the statistical consequences of utilizing sham acupuncture as a control when its energetic and biomedical effects are unknown. It argues the need for a more careful consideration of "assumptions" in clinical research design. PMID- 10513104 TI - Off white. PMID- 10513105 TI - Ending the misery of child dental decay. AB - Dental caries in children's teeth has drastically declined in the last 30 years. We should be proud of this achievement but there is much still to do. One of the greatest stumbling blocks to further progress is, I believe, the current payment system for NHS children's dentistry which has failed to promote effective prevention amongst the most caries prone. To remedy this I propose some simple, low cost changes which could make decay in a child's tooth a rarity in our practices. PMID- 10513106 TI - Recurrent Kawasaki disease. PMID- 10513107 TI - Sucking cup caries. PMID- 10513108 TI - Varying standards of oral surgery. PMID- 10513109 TI - Futuristic forensics. PMID- 10513110 TI - Sialadenitis and sialolithiasis. PMID- 10513111 TI - Teeth and implants. AB - An osseointegrated implant restoration may closely resemble a natural tooth. However, the absence of a periodontal ligament and connective tissue attachment via cementum, results in fundamental differences in the adaptation of the implant to occlusal forces, and the structure of the gingival cuff. PMID- 10513112 TI - The safety-in-use of 10% carbamide peroxide (Opalescence) for bleaching teeth under the supervision of a dentist. AB - Fears that the dentist-supervised use of a product that contains carbamide peroxide and that emits hydrogen peroxide may not be safe from the viewpoints of toxicity and cancer risk were engendered by unrealistic animal tests. These fears prompted the UK Government Departments of Trade and Industry (DTI) and of Health (DOH) to try to prohibit the marketing of Opalescence (manufactured by Ultradent Inc.). Faced with the fact that Opalescence had already been awarded a CE mark under the EC Medical Devices Directive, the DTI and DOH attempted to bring about its prohibition by reclassifying Opalescence as falling under the EC Cosmetics Directive, according to which the marketing of products containing more than 0.1% hydrogen peroxide is not permitted. PMID- 10513113 TI - Barriers to accessing dental care: dental health professional factors. AB - This article discusses the ways that the dental health professional can, sometimes unwittingly, discourage the patient from seeking dental treatment. PMID- 10513114 TI - A confocal microscopic study relating the autofluorescence of carious dentine to its microhardness. AB - OBJECTIVE: An in-vitro study to examine the correlation between the distribution of the autofluorescent signal emitted from carious dentine (detected using confocal laser scanning microscopy) and its microhardness, within the depths of human dentine lesions. MATERIALS AND METHODS: Twelve carious teeth were sectioned longitudinally, the cut faces marked with a grid reference line system and colour photomicrographs taken. The same samples were imaged using confocal laser scanning microscopy for autofluorescence and then subjected to microhardness testing using a Knoop microhardness indenter. Adjacent sound dentine was used as a control reference. Digital image superimposition allowed direct comparisons to be made between the colour, autofluorescence and microhardness of each lesion. RESULTS: Sound enamel and dentine did not autofluorescence. Autofluorescence distribution from carious dentine correlated with the highly softened tissue (detected using the Knoop indenter) and terminated at a level superficial to the translucent zone. This zone was still pigmented. Normal, sound dentine hardness levels were found deep to the translucent zone. CONCLUSIONS: A correlation existed between the zone of autofluorescence and carious dentine that was markedly softened by the carious process. These findings highlighted a possibility that the autofluorescence might be used as an in-vitro, objective histological marker for the softened, carious dentine requiring clinical excavation. PMID- 10513115 TI - A closer look at General Dental Service orthodontics in England and Wales. I: Factors influencing effectiveness. AB - OBJECTIVE: To evaluate factors influencing effectiveness in General Dental Service (GDS) orthodontics. DESIGN: Retrospective analysis of systematic 2% sample of GDS (England and Wales) cases. METHOD: Records of cases were collected during 1991. Assessment involved occlusal indices and data from National Health Service forms for 1,411 cases. Multivariate analyses were used with Peer Assessment Rating Index (PAR) score at Finish as the outcome indicator. RESULTS: Dual arch fixed appliances: achieved lower Finish PAR scores than other appliances; only 1.5% of the variance was explained, by treatment time and Dental Health Component of the Index of Orthodontic Treatment Need (DHC). Finish PAR was unaffected by Starting PAR. All other appliances: the model explained 25% of the variance for Finish PAR, which varied with Starting PAR and DHC scores. Social class had effects of little clinical significance, but the data suggested availability of orthodontic treatment was poorer in 'manual class' areas. Orthodontic qualifications, number of arches treated and mixed dentition starts had no significant effects when submitted to multivariate analysis. CONCLUSIONS: The importance of appliance selection is reinforced: dual arch fixed appliances are generally more consistent. Lower social class areas may be poorly provided with orthodontic services. PMID- 10513116 TI - Teaching and assessing ethics and law in the dental curriculum. AB - The General Dental Council's recommendations on dental education places a new emphasis on the importance of ethics and law in the dental curriculum, stating that students should have an awareness of moral and ethical responsibilities involved in the provision of care to individual patients and to populations. The duties of care to protect a patient's life and health at all times, to respect their autonomy to make informed choices about what happens to them, and to do this fairly and without prejudice, are widely accepted as the fundamental ethical principles governing all health care. The specifics of these duties of care are detailed in Maintaining Standards: guidance to dentists on professional and personal conduct, published by the GDC. PMID- 10513117 TI - Adverse health events associated with the 1998 ice storm: report of hospital surveillance of the eastern Ontario Health Unit region. PMID- 10513118 TI - Pneumococcal vaccines: World Health Organization position paper. AB - Pneumococcal diseases are a major public-health problem all over the world. The etiological agent, Streptococcus pneumoniae (the pneumococcus) in surrounded by a polysaccharide capsule. Differences in the composition of this capsule permit the serological differentiation between about 90 capsular types, some of which are frequently associated with pneumococcal disease, others rarely. Invasive pneumococcal infections include pneumonia, meningitis, and febrile bacteremia; among the common non-invasive manifestations are otitis media, sinusitis, and bronchitis. At least one million children die of pneumococcal disease every year, most of these being young children in developing countries. In the developed world, elderly persons carry the major disease burden. Conditions associated with increased risk of serious pneumococcal disease include HIV infection, sickle-cell anaemia, and a variety of chronic organ failures. Vaccination is the only available tool to prevent pneumococcal disease. The recent development of widespread microbial resistance to essential antibiotics underlines the urgent need for more efficient pneumococcal vaccines. Immunity following pneumococcal disease is directed primarily against the capsular serotype involved. The currently licensed pneumococcal vaccine is based on the 23 most common serotypes, against which the vaccine has an overall protective efficacy of about 60% to 70%. Children aged < 2 years, and persons suffering from various states of immunodeficiency, for example HIV infection, do not consistently develop immunity following vaccination, thus reducing the protective value of the vaccine in some major target groups for pneumococcal disease. However, in the healthy elderly population, the polysaccharide vaccine provides relatively efficient protection against invasive pneumococcal disease. Extensive clinical trials are now under way with a new generation of pneumococcal vaccines. These protein-polysaccharide combinations, known as conjugate vaccines, contain 7-11 selected polysaccharides bound to a protein carrier, and induce a T-cell dependent immune response. These vaccines are likely to be protective even in children < 2 years of age, and may reduce pneumococcal transmission through a herd effect. PMID- 10513119 TI - Information fusion: application to data and model fusion for ultrasound image segmentation. AB - Nowadays, information fusion constitutes a challenging research topic. Our study proposes to achieve the fusion of several knowledge sources. This, in order to detect the esophagus inner wall from ultrasound medical images. After a brief description of information fusion concepts, we propose a system architecture including both model and data fusion. The data fusion is accomplished using fuzzy modeling, which can be seen as a monosensor/multiple sources data fusion system. The model fusion is performed using a full-adapted snake theory, which projects the fuzzy decision into the binary decision space. PMID- 10513120 TI - Fusion of angiography and intravascular ultrasound in vivo: establishing the absolute 3-D frame orientation. AB - Data fusion of biplane angiography and intravascular ultrasound (IVUS) facilitates geometrically correct reconstruction of coronary vessels. The locations of IVUS frames along the catheter pullback trajectory can be identified, however the IVUS image orientations remain ambiguous. An automated approach to determination of correct IVUS image orientation in three-dimensional space is reported. Analytical calculation of the catheter twist is followed by statistical optimization determining the absolute IVUS image orientation. The fusion method was applied to data acquired in patients undergoing routine coronary intervention, demonstrating the feasibility and good performance of our approach. PMID- 10513121 TI - Improving clinical decision support through case-based data fusion. AB - This paper presents an information fusion technique based on a knowledge discovery model, and the case-based reasoning decision framework. Using signal data and database records from the heart disease risk estimation domain, three data fusion methods are discussed. Two of these methods combine information at the retrieval-outcome level, and one method merges data at the discovery-input level. The result of these three models are compared and evaluated against the performance of single-source models. It is shown that the methods that fuse information at the retrieval-outcome level are significantly superior. PMID- 10513122 TI - Multisensor fusion for atrial and ventricular activity detection in coronary care monitoring. AB - Information management for critical care monitoring is still a very difficult task. Medical staff is often overwhelmed by the amount of data provided by the increased number of specific monitoring devices and instrumentation, and the lack of an effective automated system. Specifically, a basic task such as arrhythmia detection still produce an important amount of undesirable alarms, due in part to the mechanistic approach of current monitoring systems. In this work, multisensor and multisource data fusion schemes to improve atrial and ventricular activity detection in critical care environments are presented. Applications of these schemes are quantitatively evaluated and compared with current methods, showing the potential advantages of data fusion techniques for event detection in noise corrupted signals. PMID- 10513123 TI - Dynamics of the disparity vergence step response: a model-based analysis. AB - A new method to analyze the dynamics of vergence eye movements was developed based on a reconstruction of the presumed motor command signal. A model was used to construct equivalent motor command signals and transform an associated vergence transient response into an equivalent set of motor commands. This model represented only the motor components of the vergence system and consisted of signal generators representing the neural burst and tonic cells and a plant representing the ocular musculature and dynamics of the orbit. Through highly accurate simulations, dynamic vergence responses could be reduced to a set of five model parameters, each relating to a specific feature of the internal motor command. This dynamic analysis tool was applied to the analysis of inter-movement variability in vergence step responses. Model parameters obtained from a large number of response simulations showed that the width of the command pulse was tightly controlled while its amplitude, rising slope, and falling slope were less tightly regulated. Variation in the latter three parameters accounted for the most of the movement-to-movement variability seen in vergence step responses. Unlike version movements, pulse width did not increase with increased stimulus amplitude, although the other command signal parameters were substantially influenced by stimulus amplitude. PMID- 10513124 TI - Numerical computation of differential-algebraic equations for nonlinear dynamics of multibody android systems in automobile crash simulation. AB - The principle of virtual work is used to derive the Euler-Lagrange equations of motion in order to describe the dynamics of multibody android systems. The constrained variational equations are in fact differential-algebraic equations of high index and are cast as ordinary differential equations through differentiation of the constraint equations. The integration routine LSODAR and the fourth-order Runge-Kutta method are used to compute the generalized coordinates, their time derivatives and the body forces of two android models. The graphs of the constraint forces reveal the whiplash effect on the neck and that the stiffness of both multibody systems is due to large magnitude impulsive forces experienced by many bodies simultaneously. PMID- 10513125 TI - Sol-gel-based biosensor for use in stroke treatment. AB - A fiber optic biosensor for the detection of fibrinolytic products produced during lysis of "soft" blood clots is described. The biosensor was constructed to be selective toward D dimer antigens, which form from the dissolution of cross linked fibrin clots. The presence of D dimer antigens above a threshold level is a clinical diagnostic used to determine the presence of such occlusions following a stroke. Fluorescein-labeled D dimer antibodies are immobilized on the tip of an optical fiber by dip coating from a silica sol-gel solution. When D dimer antigens combine with the antibodies, fluorescence intensity decreases. The response of the sensor was examined in phosphate buffered saline, human plasma, and blood. Calibration plots for the sensor were obtained in the clinically significant D dimer concentration range from 0.54 microgram/ml to 6 micrograms/ml. Changes in spectroscopic properties as the sol-gel encapsulated tagged antibodies aged were examined; a decrease in fluorescence intensity with age was noted. The D dimer antibodies remain viable for at least 4 weeks while encapsulated in the sol-gel network when stored at 4 degrees C in PBS solution. This novel sensor is being developed for use with other catheter-based microtools to treat stroke resulting from occlusion in the vascular system. PMID- 10513126 TI - Force controlled and teleoperated endoscopic grasper for minimally invasive surgery--experimental performance evaluation. AB - Minimally invasive surgery generates new user interfaces which create visual and haptic distortion when compared to traditional surgery. In order to regain the tactile and kinesthetic information that is lost, a computerized force feedback endoscopic surgical grasper (FREG) was developed with computer control and a haptic user interface. The system uses standard unmodified grasper shafts and tips. The FREG can control grasping forces either by surgeon teleoperation control, or under software control. The FREG performance was evaluated using an automated palpation function (programmed series of compressions) in which the grasper measures mechanical properties of the grasped materials. The material parameters obtained from measurements showed the ability of the FREG to discriminate between different types of normal soft tissues (small bowel, lung, spleen, liver, colon, and stomach) and different kinds of artificial soft tissue replication materials (latex/silicone) for simulation purposes. In addition, subjective tests of ranking stiffness of silicone materials using the FREG teleoperation mode showed significant improvement in the performance compared to the standard endoscopic grasper. Moreover, the FREG performance was closer to the performance of the human hand than the standard endoscopic grasper. The FREG as a tool incorporating the force feedback teleoperation technology may provide the basis for application in telesurgery, clinical endoscopic surgery, surgical training, and research. PMID- 10513127 TI - Use of the electrohysterogram signal for characterization of contractions during pregnancy. AB - This article proposes a method to evaluate the ability of the electrohysterogram signal to characterize the contractions during pregnancy, in a population with high risk of preterm deliveries. This study constitutes a first stage of a project intended to develop a monitoring system for the early diagnosis of preterm deliveries. After a proper signal denoising, we calculate some parameters characteristic of the extracted contractions. These contractions are then divided into classes of different physiological terms. Classical techniques of data analysis, such as principal component analysis and discriminant analysis, permit us to show an evolution of the contractions during pregnancy, which is different between the groups of preterm deliveries and that of deliveries at term. We show that, in an early term of pregnancy, we can separate the two populations: women delivering at term from women delivering preterm. We then show that these two kinds of pregnancy are of different evolutions. These results are encouraging, because they would permit, in a follow-up medical study, to diagnose a possible preterm delivery, as well as the proximity of the delivery. PMID- 10513128 TI - Wavelet analysis of oscillations in the peripheral blood circulation measured by laser Doppler technique. AB - The wavelet transform technique, a time-frequency method with logarithmic frequency resolution, was used to analyze oscillations in human peripheral blood flow measured by laser Doppler flowmetry. The oscillations extended over a wide frequency scale and their periods varied in time. Within the frequency range studied, 0.0095-1.6 Hz, five characteristic oscillations were revealed, arising from both local and central regulatory mechanisms. After the insertion of endothelium-dependent and endothelium-independent vasodilators the spectra of blood flow markedly differed in the frequency interval 0.0095-0.02 Hz. In this way it was demonstrated that endothelial activity is a rhythmic process that contributes to oscillations in blood flow with a characteristic frequency of around 0.01 Hz. The study illustrates the potential of laser Doppler flowmetry combined with dynamical systems analysis for studies of both the micro- and macroscopic mechanisms of blood flow regulation in vivo. PMID- 10513129 TI - On statistical properties of whole nerve cuff recordings. AB - Whole nerve cuff electrodes can record an electric signal generated by the superposition of single fiber action potentials (AP's). Using a simple stochastic model for the superposition of AP's, the statistical properties of nerve cuff signals are mathematically derived in this study. Consequences of common signal processing methods like rectification and time-averaging are also explained. The nerve cuff signals are found to be approximately identically, independently distributed Gaussian signals with zero mean and varying variance. The spectral properties of the cuff signals generated by single AP shape or different AP shapes are also addressed and investigated by examining the properties of the autocorrelation functions of the nerve cuff signals. The theoretical results were found to be in accordance with computer simulations and processing of actual recorded data. PMID- 10513130 TI - Quantitative laser scanning confocal autofluorescence microscopy of normal, premalignant, and malignant colonic tissues. AB - Laser scanning confocal autofluorescence microscopy (LSCAM) using 351- to 364-nm excitation light was used to quantitatively compare fluorescent spectral emission of unstained, frozen histological sections of normal, premalignant, and malignant colonic tissues. To identify the spatial origins of fluorescent signals accurately, the same frozen section slides used for microscopy were fixed and histochemically stained immediately following LSCAM imaging. Tissue fluorescence emission was quantified in terms of the intrinsic fluorescence coefficient beta (lambda), defined as the fluorescence power per unit tissue volume per unit wavelength (centered at lambda) divided by the incident light irradiance. Over all emission wavelengths, colonic tissues emitted autofluorescence ranging from beta (lambda) approximately 10(-1.5) to 10(-3.0) cm-1. In the 530- to 610-nm spectral region, markedly increased autofluorescence (beta up to 10(-2.5)) was observed in the dysplastic cells of adenomatous polyps, as compared to normal epithelial cells. Compared to adenomatous polyps, decreased dysplastic cell autofluorescence was observed in adenocarcinoma. The brightest fluorescence in the lamina propria, which was attributed to eosinophils (beta approximately 10( 2.5)) in previous studies, was also observed in other granular structures (beta up to 10(-1.4)). LSCAM reveals quantitative significant differences in fluorescence emission between normal and diseased colonic tissues. PMID- 10513131 TI - Virtual reality-based training for the diagnosis of prostate cancer. AB - Prostate malignancies are the second leading cause of cancer deaths among men. The most common method of detecting this disease is digital rectal examination (DRE). Current DRE training is inadequate, since the number of patients that students can practice on is limited. Furthermore, allied care personnel do not train in screening for prostate cancer. Finally, there is no objective way to follow the improvement in DRE skills for medical personnel. This paper presents a virtual reality-based simulator that addresses the above problems. The prototype consists of a PHANToM haptic interface which provides feedback to the trainee's index finger, a motion restricting board, and an SGI workstation, which renders the patient's anatomy. Four types of prostates were modeled--normal, enlarged with no tumor, incipient malignancy (single tumor), and advanced malignancy (tumor cluster). Human factors studies were conducted on both nonmedical students and urology residents in order to quantify the system usefulness. After only five minutes of training, nonmedical students had a 67% correct diagnosis rate of malignant versus nonmalignant cases. This compared with 56% for urology residents in the same trials. Subjective evaluation by the residents pointed out the need to improve the virtual prostate model realism. A control group formed of urology residents performed the same trials on a modified Merck Procar simulator. The control group scored significantly better (96% correct diagnosis of malignancies). We conclude that the virtual prostate palpation simulator, while promising, needs significant improvement in both model realism and haptic interface hardware. PMID- 10513132 TI - Estimation of distortion product otoacoustic emissions. AB - Distortion product otoacoustic emissions (DPOAE's) are acoustic signals generated from the cochlea when stimulated by dual tone stimulus. The measurement of the DPOAE's is useful in objectively assessing the hearing functionality of humans. The parameter estimation of the DPOAE's has been based on the fast Fourier transform (FFT) technique, which may not guarantee optimum performance in general. In this paper, we investigate the optimal maximum-likelihood estimator (MLE) for the DPOAE's. Experiments showed that the performance of the MLE is superior to those of the conventional FFT estimators. Furthermore, it is proven that the unwindowed FFT estimator is essentially the MLE when the stimulus tones are constrained to exhibit complete cycle within the data frame. PMID- 10513133 TI - Estimating cortical potentials from scalp EEG's in a realistically shaped inhomogeneous head model by means of the boundary element method. AB - Cortical potentials are estimated from scalp potentials using a realistically shaped inhomogeneous head model, by means of the boundary element method (BEM). A new adaptive algorithm has been developed to achieve high accuracy to link directly the cortical potentials to the scalp potentials in a realistically shaped inhomogeneous head model including the thin low-conductivity skull layer. Computer simulations using a concentric three-spheres head model have tested this approach. The present study demonstrates that the cortical potentials can be directly estimated from the scalp potentials using the BEM in a realistically shaped inhomogeneous head model. PMID- 10513134 TI - Assessment of extensive surgery for locally advanced lung cancer. Safety and efficacy of induction therapy. AB - OBJECTIVE: Locally advanced lung cancer has a poor prognosis, despite extensive surgery conducted in an effort to improve survival. We evaluated the safety and efficacy of induction therapy prior to extensive surgery for locally advanced lung cancer. METHODS: Primary resection for lung cancer was done in 549 consecutive patients divided into three groups; 446 undergoing standard pulmonary resection (no extensive surgery), 87 undergoing extensive surgery without induction therapy, and 16 undergoing surgery after induction therapy. RESULTS: Morbidity was 23.5%, 28.6%, and 43.8%, respectively. The rate was significantly higher in the induction group compared with the no extensive surgery group (P < 0.05). Surgical mortality was 0.67%, 3.4%, and 6.3%, respectively. The difference was statistically significant between the no extensive surgery and extensive surgery groups (P < 0.02), and between the no extensive surgery and induction groups (P < 0.02). Hospital mortality was 2.2%, 9.2%, and 6.3%, respectively. The rates were significantly higher in the extensive surgery (P < 0.01) and induction (P < 0.05) groups compared to the no extensive surgery group. Five-year survival was 50.3% for the patients who received induction therapy, and 14.7% for the patients who did not receive induction therapy. CONCLUSIONS: Survival differences between the induction and non induction groups were not significant, but some patients with T3 or T4 disease may benefit from induction therapy. The high morbidity of induction treatment should be recognized, and strict candidate selection and careful postoperative care used to help prevent increased mortality. PMID- 10513135 TI - Simultaneous repair of arch and abdominal aortic aneurysms. A simple new technique using a temporary bypass graft. AB - OBJECTIVE: Atherosclerotic aneurysms in the aortic arch are associated with abdominal aortic aneurysms in up to 37% of cases. We have developed a single stage approach to the repair of both aneurysms using a temporary bypass. SUBJECTS: Since November 1996, 5 patients underwent simultaneous repair of aneurysms in the aortic arch and in the infrarenal abdominal aorta, using a new temporary bypass graft technique. Entire arch replacement with simultaneous abdominal aortic aneurysmectomy was performed in one patient. The other 4 patients underwent distal hemi-arch replacement distal from the orifice of the brachiocephalic artery with simultaneous repair of the abdominal aortic aneurysm. METHOD: For the entire arch replacement procedure, blood flow to all major branches of the aortic arch was established using a bifurcated graft. This graft anastomosed to the ascending aorta was used as the proximal inflow of the temporary bypass graft. For the hemi-arch replacement procedure, the proximal inflow segment of the temporary bypass graft was anastomosed to the brachiocephalic artery. In both cases, the distal outflow segment of the temporary bypass graft was the graft used for repair of the abdominal aortic aneurysm. In order to prevent any clamp injury, Teflon felt was tightly wrapped around the aorta before the clamp was applied. RESULTS: Evaluation of the hemodynamic parameters measured during cross-clamping of the aortic arch revealed stable distal perfusion to the visceral organs and no excessive increase in cardiac afterload. All patients had an uneventful postoperative course and were discharged within 1 month of surgery. CONCLUSION: Our temporary bypass method is recommended for simultaneous replacement of aneurysms in the aortic arch and the abdominal aorta. PMID- 10513136 TI - Cardioprotective effect of orally administered angiotensin-converting enzyme inhibitor against ischemia. Reperfusion injury in the isolated rat heart. AB - Angiotensin-converting enzyme inhibitors are reported to be cardioprotective against ischemia/reperfusion injury. Few studies have been made, however, on the cardioprotectiveness of orally administered angiotensin-conrerting enzyne inhibitors. Wistar rats were pretreated with oral delapril--30 mg/kg/day in the low-dose group and 90 mg/kg/day in the high-dose group--for one week. Cardiac function recovery was assessed after ischemia/reperfusion in the isolated working heart model. Rat hearts in the high-dose group were also reperfused with a solution containing nitro-L-arginine methyl ester, a nitric-oxide synthase inhibitor. Oral pretreatment of delapril did not affect baseline cardiac function. The percentage of cardiac output recovery for controls was 22 +/- 4.5%, for the low-dose group 44 +/- 6.5% (P < 0.05 versus controls), and for the high dose group 76 +/- 5.3% (P < 0.001 versus controls and low-dose). Although coronary vascular resistance at the end of reperfusion showed no difference, mean coronary vascular resistance early after reperfusion was significantly lower (P < 0.0001) in both delapril groups than in control. In the high-dose group, reperfusion with L-NAME significantly increased coronary vascular resistance after ischemia/reperfusion and attenuated the cardioprotectiveness of delapril (P < 0.05 versus without nitro-L-arginine methyl ester). We thus found that oral administration of delapril was cardioprotective dose-dependently against ischemia/reperfusion injury. Nitric oxide appeared to be involved in the mechanism behind this cardioprotective effect. PMID- 10513137 TI - Long-term results of the fenestrated Fontan operation. Progress of patients with patent fenestrations. AB - The fenestrated Fontan operation was introduced as a modification of the "completed" Fontan operation for patients with high risk factors, and low operative mortality has frequently been reported. However, use of the umbrella device is now restricted, and this procedure should be performed without subsequent closure. In this paper, we review our clinical experience with this procedure and discuss ongoing problems. Sixteen patients (4 tricuspid atresia and 12 other cardiac anomalies including 5 cases of univentricular heart) underwent the fenestrated Fontan operation (7 atriopulmonary and 9 total cavopulmonary connection). All of them have some risk factors for a completed Fontan operation. There were three early deaths of the 16. Two experienced an anticipated thromboembolic accident, one of which involved the pulmonary aspect while the other involved the arterial aspect. Patients who survived the operation have progressed well and have a clinical status of New York Heart Association class I, with the exception of one late death due to congestive heart failure. There have been no thromboembolic accidents in this group during the late follow-up period. Spontaneous closures of the fenestrations were noted in two patients. The late mean Qp/Qs value in patients with patent fenestrations was 0.80 +/- 0.1, SaO2 was 88.8 +/- 5.6%, and right atrial pressure was 9.7 +/- 3.8 mmHg. No major problems have been encountered in patients with a patent fenestration over extended periods. A modified Fontan operation to fit a permanently open fenestration may be considered as a final surgical option for certain high-risk patients. PMID- 10513138 TI - Partial median sternotomy as a minimal access for the closure of subarterial ventricular septal defect. Feasibility of transpulmonary approach. AB - BACKGROUND: Minimally invasive techniques in congenital heart surgery have evolved steadily over the past few years, but documentation in the literature is rare. The majority of reported techniques involve thoracoscopic approach and partial sternotomy. We have employed a lower partial sternotomy as a minimal access procedure for the closure of subarterial ventricular septal defect, for situation where this approach would be unsuitable for adequate exposure of the pulmonary artery. The purpose of this study is to demonstrate the feasibility and safety of this technique and report its superior cosmetic result. SUBJECTS AND METHODS: Beginning in 1997, we began approaching the closure of subarterial ventricular septal defect through a lower sternal split incision using a 6 to 10 cm skin opening, associated with a reversed L incision at the left second intercostal space. A total of consecutive 12 patients (6 male and 6 female) have been operated on using this approach. The patients ranged in age from 6 to 21 years (mean, 12.8 +/- 5.0 years). The straight cannula with stylet was used for aortic cannulation. RESULTS: There was no mortality or morbidity, except for late pericardial effusion in 4 cases. The durations of cardiopulmonary bypass and aortic cross-clamping ranged from 94 to 206 (mean, 131 +/- 33) minutes and from 40 to 122 (mean, 70 +/- 26) minutes, respectively. Ten of 12 patients were extubated in the operating room, and no patient required blood transfusion. The postoperative hospital stay ranged from 8 to 21 (mean, 13.4 +/- 4.2) days. No patient developed deterioration of aortic regurgitation or residual ventricular septal defect. CONCLUSIONS: Our experience demonstrates that the lower partial sternotomy for the closure of subarterial ventricular septal defect is technically feasible and can be used with excellent cosmetic results and safety. Although experience is limited and follow-up is relatively short, this less invasive surgical technique may become a beneficial option for the management of subarterial ventricular septal defect. PMID- 10513139 TI - Lung volume reduction surgery results in pulmonary emphysema. Changes in pulmonary function. AB - From December 1993 to August 1998, we conducted 57 lung volume reduction surgeries on patients with severe pulmonary emphysema but without giant bullae. Of these, 26 underwent unilateral lung volume reduction surgery and 31 bilateral surgery. We analyzed the results of thoracoscopic lung volume reduction surgery (unilateral: 25; bilateral: 16) and volume reduction surgery by median sternotomy (unilateral: 1; bilateral: 15). Bilateral surgery via thoracoscope and median sternotomy significantly improved symptoms and pulmonary functions; mean improvement in forced expiratory volume in 1 second was 42.4% for bilateral thoracoscopic volume reduction surgery and 60.0% for median sternotomy. Unilateral volume reduction surgery produced a mean improvement in forced expiratory volume in 1 second of 28.9%. No significant complications were seen with either procedure. Reevaluation at 1 and 2 years after lung volume reduction surgery showed improvement to be well maintained. PMID- 10513140 TI - Left atrial myxoma associated with acute myocardial infarction. AB - We describe a patient with left atrial myxoma associated with acute myocardial infarction. Since hemodynamics were impaired even with the support of an intra aortic balloon pump, the patient underwent removal of the tumor concomitant with coronary artery bypass grafting to the right coronary artery on the fifth day from infarction onset. In circumstances of life-threatening of myxoma associated with acute myocardial infarction, removal of myxoma with coronary artery bypass should be performed in an acute phase of myocardial infarction. PMID- 10513141 TI - A long-term survival case of thymic squamous cell carcinoma, performed complete extirpation with vascular reconstruction of the superior vena cava. AB - A 59-year-old woman who complained of anterior chest pain exhibited an abnormal shadow on chest X ray and was admitted to our hospital. The chest X ray showed a demarcated tumor at the anterosuperior mediastinum and she was diagnosed as having a mediastinal tumor. After the midsternotomy was performed, the mediastinal tumor derived from the thymic tissues was discovered to have invaded the right upper lung, pericardium and superior vena cava. After excising the tumor, anastomosis between the right brachiocephalic vein and superior vena cava, followed by that between the left brachiocephalic vein and right cardiac auricle was performed using expanded polytetrafluoroethylene-ringed vascular grafts (phi 10 mm) for reconstruction. The tumor was diagnosed as a thymic carcinoma (squamous cell carcinoma) pathologically. After surgery, she was treated by cobalt irradiation. One month and again 3 months after the operation, venography showed patency. The patient has not demonstrated recurrence for 9 years and 6 months. PMID- 10513142 TI - Axillofemoral bypass for recurrent atypical coarctation of the thoracic aorta. Woman in childbearing age. AB - Various surgical techniques for recurrent atypical coarctation have been described, and extra-anatomic bypass with a thoracotomy or a sternotomy approach has been widely recommended. We report a case where axillofemoral bypass has been used to treat a 28-year-old woman with recurrent atypical coarctation. Ordinarily, she had not suffered greatly from hypertension, but she experienced repeated miscarriages most probably owing to uncontrolled hypertension over 200 mmHg during pregnancy. We chose an axillofemoral bypass for extra-anatomic bypass to manage intractable hypertension during pregnancy. Postoperatively, her hemodynamics improved substantially, particularly during pregnancy, and two children were successfully delivered. The patient remains in excellent condition 74 months after operation. We suggest that an axillofemoral bypass will become an option among surgical techniques for recurrent coarctation under individual circumstances. PMID- 10513143 TI - Type I acute aortic dissection accompanied by ischemic enterocolitis due to blood flow insufficiency in the superior mesenteric artery. AB - We report a case of acute type I aortic dissection with ischemic enterocolitis due to blood flow insufficiency in the superior mesenteric artery. The patient was a 52-year-old man who visited the hospital with major complaints of sudden low back pain and melena. Mesenteric ischemia was suspected, and angiography revealed type I aortic dissection with accompanying blood flow insufficiency in the superior mesenteric artery. Because catheterization during angiography improved the blood flow disorder and prevented intestinal necrosis, it was possible to replace the ascending aorta with a prosthetic graft. Arterial pulsation in the mesentery was recovered by the operation and the patient's life was saved without bowel resection. This case demonstrates that prompt surgical or percutaneous relief of ischemia in major organs is important to save lives in the cases of acute aortic dissection with ischemic complications. PMID- 10513144 TI - Perceived racial discrimination, depression, and coping: a study of Southeast Asian refugees in Canada. AB - Using data obtained from personal interviews with 647 Southeast Asian refugees in Canada, this study tests hypotheses regarding both the association between perceived racial discrimination and depression, and the roles of coping and ethnic identity in conditioning the nature of the discrimination-depression relation. Refugees who reported that they had experienced racial discrimination had higher depression levels than their counterparts who reported no such experiences. Responding to discrimination through confrontation was not significantly associated with depression. Study findings support the effectiveness of forbearance in diminishing the strength of the association between discrimination and depression. The moderating effect of forbearance was conditioned by the level of ethnic identity: The beneficial effect of forbearance was significantly greater among those holding stronger ethnic identification. Cultural and situational interpretations of the findings are presented. PMID- 10513145 TI - The prevalence, distribution, and mental health correlates of perceived discrimination in the United States. AB - The survey data presented here are on the national prevalences of major life-time perceived discrimination and day-to-day perceived discrimination; the associations between perceived discrimination and mental health; and the extent to which differential exposure and differential emotional reactivity to perceived discrimination account for the well-known associations between disadvantaged social status and mental health. Although more prevalent among people with disadvantaged social status, results show that perceived discrimination is common in the total population, with 33.5 percent of respondents in the total sample reporting exposure to major lifetime discrimination and 60.9 percent reporting exposure to day-to-day discrimination. The associations of perceived discrimination with mental health are comparable in magnitude to those of other more commonly studied stressors, and these associations do not vary consistently across subsamples defined on the basis of social status. Even though perceived discrimination explains only a small part of the observed associations between disadvantaged social status and mental health, given its high prevalence, wide distribution, and strong associations with mental health, perceived discrimination needs to be treated much more seriously than in the past in future studies of stress and mental health. PMID- 10513146 TI - Unwanted childbearing, health, and mother-child relationships. AB - This paper investigates the relationships among unwanted childbearing, health, and mother-child relationships. We hypothesize that unwanted childbearing affects mother-child relationships in part because of the physical and mental health consequences of unwanted childbearing. Impaired mental health hampers women's interaction with their infants, and these poor neonatal relationships translate into poor mother-adult child relationships. Using the Intergenerational Panel Study of Mothers and Children--a 31-year longitudinal survey of a probability sample of 1,113 mother-child pairs begun in 1961--we demonstrate that mothers with unwanted births have lower quality relationships with their children from late adolescence (age 18) throughout early adulthood (ages 23 and 31). Furthermore, these lower quality relationships are not limited to the child born as a result of the unwanted pregnancy; all the children in the family suffer. Using the 1987-88 wave of the National Survey of Families and Households, a survey of a national probability sample of U.S. households, we show that mothers with unwanted births suffer from higher levels of depression and lower levels of happiness. We also demonstrate that they spank their young children more and spend less leisure time with them. We conclude that experiencing unwanted childbearing reduces the time and attention that mothers give their young children and that these early mother-child interactions set the stage for long term, lower quality relationships. PMID- 10513147 TI - The intergenerational transmission of health-risk behaviors: adolescent lifestyles and gender moderating effects. AB - The present longitudinal study of 330 adolescents used structural equation models to investigate whether (1) health-risk lifestyles exist among adults and adolescents, (2) parents' health-risk behaviors influence adolescents' health risk behaviors, and (3) intergenerational transmission occurs by way of a health risk lifestyle, by direct transmission of specific behaviors, or through both mechanisms. To address these questions, we estimated several models. The findings were generally supportive of the expectations. Results of single factor measurement models provided modest evidence for the existence of an underlying health-risk lifestyle factor among parents and adolescents. Results of structural equation models also demonstrated that parents' health-risk behaviors were transmitted to adolescents both at the lifestyle factor level and the unique component level. These associations prevailed even after controlling for family social status. However, parents' health-risk lifestyles mediated the effect of family social status on adolescents' lifestyles, net of the direct effect of family social status on adolescents' lifestyle. In these two-parent families, the effects of parents' health-risk lifestyles on adolescents seems to have gender symmetry. The findings of the separate models for boys and girls demonstrated that (1) fathers' health-risk lifestyle affected only boys' health-risk lifestyle, whereas (2) mother's health-risk lifestyle affected only girls' health risk lifestyle. A similar gender moderating effect was not found for specific health-risk behaviors. Implications of these findings for future research and theoretical development are discussed. PMID- 10513148 TI - Age and anger. AB - Are older people less angry? I propose that age differences in roles, personal and social circumstances, the sense of control, health, and socio-emotional outlook explain the association. I use data from a 1981 representative sample of 951 physically disabled individuals from Southwestern Ontario, Canada and a 1996 national probability sample of 1,450 U.S. respondents--the General Social Survey (GSS). Both surveys show a negative association between age and anger. In the Ontario sample older people are more likely to occupy widowhood and retirement roles, live with fewer people, have less interpersonal estrangement, and have fewer life events; these characteristics explain their lower anger. Also, were it not for their lower control and worse health older people in the Ontario sample would report even lower anger. In the GSS sample, age differences in household composition, satisfaction with family life and financial circumstances, perceived time pressures in daily life, religious involvement, and socio-emotional outlook contribute to the lower anger among older adults. Collectively, my findings show that the psychosocial and structural environment--experienced differently by age- influences the risk of anger. PMID- 10513149 TI - Job complexity and employee substance use: the moderating effects of cognitive ability. AB - This study examines the extent to which individual's general cognitive ability influences relations between the complexity of their jobs and their use of four different substances: cigarettes, alcohol, marijuana, and cocaine. We tested this possibility using 1992 and 1982 data sets from the National Longitudinal Survey of Youth (Center for Human Resource Research 1993). The 1992 data set included 7,112 individuals and measures of all four substances. The 1982 set included 8,548 individuals and a measure of alcohol use only. Our results showed that for three of the substances (cigarettes, alcohol, and marijuana), individuals responded differently to job complexity as a function of their cognitive ability. Specifically, for individuals low in cognitive ability, the more complex their jobs, the greater their use of cigarettes, alcohol, and marijuana. On the other hand, for those with high cognitive ability, the more complex their jobs, the lower their use of these substances. Results also showed that cognitive ability had no significant impact on the association between job complexity and cocaine use. In sum, our findings suggest that providing employees with jobs that are compatible with their general cognitive ability may result in lower levels of licit and illicit substance use. PMID- 10513150 TI - Employee exposure to coworker substance use and negative consequences: the moderating effects of work group membership. AB - The current study assesses: (1) whether the relationship between individual exposure to coworker substance use and negative consequences resulting from exposure depends on work group membership, and (2) whether group-level characteristics moderate the relationship between exposure and consequences. At the group-level, we assessed occupations involving safety risk or high mobility and social factors of drinking climate and group cohesiveness. We conducted Hierarchical Linear Modeling (HLM) across two samples of municipal employees (ns = 650, 878; n of groups = 50, 49). Our results revealed that groups with higher proportions of jobs involving risk (e.g., machine work) and, to a lesser extent, groups with a higher level of drinking climate were those most vulnerable to consequences under conditions of exposure. Importantly, our findings controlled for individual risk factors (e.g., personal drinking, job stress). Our discussion examines the implications of this study for theory and policy related to workplace substance abuse. PMID- 10513151 TI - [The pattern of mediastinal nodal involvement in lung cancer according to tumor located lobe]. AB - To clarify the pathway of the metastases from each pulmonary lobe to mediastinal nodes, we examined the pattern of mediastinal nodal involvement in 462 resected pN2 non-small cell lung cancer. Carcinomas of the right upper lobe frequently involved #3 (78/133) and #4 (70/133) nodes, whereas those of the right middle or lower lobe frequently metastasized to #7 nodes (18/23 and 86/113, respectively). On the other hand, carcinoma of left upper lobe frequently involved #5 nodes (81/118), whereas those of the left lower lobe most frequently metastasized to #7 nodes (50/75). Of 462 pN2 patients, 95 (20.6%) had skip metastases to the mediastinal nodes. Skip metastasis was observed more frequently in carcinomas of right upper and middle lobe. One of the reasons of skip metastasis may be the direct lymph drainage through subpleural space to mediastinum. PMID- 10513152 TI - [Surgical results on N2 lung cancer with special reference to correlation between tumor size and extension of lymph node metastases]. AB - Systematic lymph node dissection was performed for every patients undergoing surgical intervention. Since 1981, there were 218 stage IIIA-N2 patients who underwent resection with two operative mortality. The five-year survival rate of whole cases was 22.6%, and that of 152 completely resected cases was 30.0%. Favorable factors on long-term survival of pN2 patients were cN0, T1-2 N2M0, single mediastinal node involvement, and tumor less than 20 mm or less. The five year survival rates of stage IIIA-N2 patients with tumor diameter of < or = 20 mm, 21-30 mm, 31-50 mm, and > or = 51 mm were 48.1%, 27.7%, 31.2%, and 16.7%, respectively. When micrometastases to lymph node in the p-stage I patients (diagnosed by H-E staining) were examined by immunohistochemical staining, 36 patients (27%) out of 132 verified micrometastases in the lymph nodes. PMID- 10513153 TI - [Surgical treatment of N2 involved non-small cell lung cancer--the systematic extended lymph node dissection based on the regional lymphatic drainage]. AB - From the study on regional lymphtic drainage, we have decided the extent of lymphadenectomy as follows; a) For the left lung cancer and the right upper lobe primary, systematic bilateral mediastinal dissection (R3 alpha) through a median sternotomy, b) For the cases with the highest mediastinal node involvement, lower half of radical neck dissection (R3 gamma) through a cervical collar incision in addition to a). The cN diagnosis by CT interpretation and pN diagnosis were compared. The under estimated rates of N were 32% of 137 patients with the left lung primary. 46 patients with pN2(+) included 14 patients (31%) with pN3 disease. As for the right upper lobe primary, 17 patients with pN2(+) included 13 patients (76%) with pN3 disease. Postoperative survival rates calculated with Kaplan-Meiermethod; 1) The five-year survival rates were 43% of 46 patients with pT1-3 N2-3 of the left lung primary. 2) As for the right upper lobe primary, the two-year survival rates were 51% of 17 patients with pT1-4 N2-3. 3) The three year survival rates of 26 patients with pN3 gamma diagnosed as cN0-3 alpha preoperatively were 41%. These systematic extended dissection (R3 alpha, R3 gamma) would bring better prognosis in the patients with pN2-3 disease. PMID- 10513154 TI - [A clinical study for improving the survival rate of pN2 lung cancer through our case of 53 patients]. AB - BACKGROUND: In management of non-small cell lung cancer, the evaluation and treatment of N2 disease has a lot of controversy. MATERIALS AND METHODS: Between 1983 and 1998, 53 patients of pN2 non-small cell lung cancer were operated by standard lymph node dissection method (R2) using CUSA system. We studied the sensitivity of the diagnosis of preoperative N factor, survival rate, and analysed the relationship between the postoperative mediastinal lymph node metastasis and the site of recurrence. RESULTS: Three-year and five-year survival rates for 53 cases were 46.8% and 33.4% respectivery. Preoperative sensitivity of CT scan for N factors were only 45% in squamous cell carcinoma and 24.2% in adenocarcinoma. Even with intraoperative findings, the sensitivity was not better. In a follow up survey, ipsilateral mediastinal lymph node recurrence was not detected, contralateral mediastinal lymph node recurrences were rare and the distant metastases were common cause of death. CONCLUSION: It is more important to accomplish the standard lymph node dissection completely in all cN cases than to evaluate the preoperative node stage aggressively using invasive methods. PMID- 10513155 TI - [Analysis of the long-term survived patients with P-N2 non-small cell lung cancer]. AB - We had 308 patients with P-N2 non-small cell lung cancer who were treated with pulmonary resection therapy from June 1975 to March 1994. We analysed 9 patients who could survive for 5 years and more after the operation. All of them were male and the mean age was 61.1 years. Histologically, 2 patients had adenocarcinoma and 7 patients had squamous cell carcinoma. The clinical staging was T2N1M0 in 2 patients, T1N2M0 in one, T2N2M0 in 2, T3N2M0 in 2, and T4N2M0 in 2 patients. The histopathological staging was T1 in one patient, T2 in 4, T3 in 3, and T4 in one patient. Three patients who had absolute non-curative operation (residual carcinoma on bronchial stump in 2 patients and residual mediastinal lymph nodes in one patient) were treated with radiation therapy after having operation. The one level mediastinal lymph node metastasis was found in 7 patients and multi level one in 2 patients. Four patients out of nine are still alive and free from the cancer and 2 patients died of other unrelated disease. The patient who even have P-N2 non-small cell lung cancer with mediastinal lymph node involvement will live long after curative operation. PMID- 10513156 TI - [Effect of preoperative chemotherapy for bulky N2 non-small-cell lung cancer]. AB - The effect of chemotherapy as the adjuvant therapy to bulky N2 disease (stage IIIA) non-small-cell lung cancer was examined. From January 1992 to December 1996, 464 patients with non-small-cell lung cancer underwent surgery. Seven patients (1.5%) with N2 disease (stage IIIA) received two cycles of preresectional cisplatin and vindesin chemotherapy, followed by standardized surgical resection (Group A). 46 patients (9.9%) had pathological N2 disease (T1 3, M0) after surgery (Group B). In Group A a complete resection was accomplished in two patients (28.6%), and five patients had incomplete resection with a deseased margin, followed thoracic irradiation 60 to 75 Gy. In three patients in Group A the N2 disease was pathologically downstaged to N1 or N0 disease. Overall survival at five years in Group A and in Group B was 48% and 39%, and median survival time was 49 months and 38 months. Although complete resection rate was lower in Group A (28.6%) than in Group B (78.2%), there was no significant difference between five year survival and median survival time in Group A and Group B. These data may be thought to suggest that induction chemotherapy in Group A was effective on occult micrometastatic disease. PMID- 10513157 TI - [The case report of minimal access CABG for triple-vessel disease]. AB - A 55-year-old male patient underwent CABG for triple-vessel disease using the minimal access approach. The procedure was performed through a limited (10 cm) left para-sternal thoracotomy using extracorporeal circulation established with a usual aortic cannula, and pulmonary arterial and right atrial drainage. The myocardium was protected by antegrade administration of cold cardioplegic solution while the aorta was being cross-clampled. The saphenous vein graft was connected sequentially to the 4 PD and OM branches, and the left internal thoracic artery was grafted to the LAD. The postoperative course was uneventful and coronary angiography showed that all three grafts were patent. The patient was discharged one week postoperatively. PMID- 10513158 TI - [Surgical treatment of a ruptured saccular aneurysm associated with bilateral coronary arteries-pulmonary artery fistulas: a case report]. AB - We reported a successful operative case of ruptured coronary artery saccular aneurysm associated with bilateral coronary arteries-pulmonary artery fistulas. A 57-year-old woman, had been treated with hemodialysis due to chronic renal failure, suffered from acute heart failure with chest pain suddenly. Echocardiograph showed moderate pericardial effusion. A saccular coronary artery aneurysm with bilateral coronary arteries-pulmonary artery fistulas revealed by coronary angiogram. Ligation of coronary artery fistula, closure of orifice of draining artery to pulmonary artery and aneurysmorrhaphy were performed emergently. Post operative angiogram revealed complete disappearance of the fistulas. PMID- 10513159 TI - [A case report of total aortic reconstruction and choice of staged operation in Marfan syndrome]. AB - A 37-year-old man with Marfan syndrome underwent four operations for extensive cardiovascular disease. He was diagnosed as having AAE, AR and DeBakey type I aortic dissection. First, Bentall operation using Piehler procedure and total aortic arch replacement using retrograde cerebral perfusion and profound hypothermia at 18 degrees C were performed on May 11, 1994. Second, repair of leakage of the right coronary artery anastomosis and grafting for the descending thoracic aortic aneurysm were performed on December 3, 1994. Y-type grafting for the AAA was performed on December 21, 1996. Last, grafting for TAAA was performed under hypothermia at a rectal temperature of 20 degrees C on November 17, 1997. This surgical strategy of staged operation for extensive cardiovascular disease in Marfan syndrome is an effective method. Regular follow-up by CT is necessary for deciding the time and method of reoperation. PMID- 10513160 TI - [Removal of a giant left atrial myxoma by superior transseptal approach and division of the ascending aorta: a case report]. AB - A 61-year-old male with the complaint of chest oppression and dyspnea was diagnosed as having giant left atrial tumor by echocardiography and MRI. Removal of the tumor was performed by division of superior transseptal approach and the ascending aorta. Excellent exposure and complete resection were obtained using this approach. PMID- 10513161 TI - [Regrowth of bullae around the staple-line is one of the causes of postoperative recurrence in thoracoscopic surgery for spontaneous pneumothorax]. AB - Since August 1994, we have performed 65 thoracoscopic surgeries for spontaneous pneumothorax. Our operative method is partial resection of the lung with bullae using endoscopic autosuture. Three patients underwent reoperation because of persistent air leakage in one case and postoperative recurrences in two cases. In addition, we performed a second operation in one case that had undergone thoracoscopic surgery for spontaneous pneumothorax in another hospital. At the second operation, regrowth of bullae was found at the edge of staple-line in three of the 4 cases, and in one of these cases, such regrown bullae seemed to be a cause of recurrence of pneumothorax. The regrowth of the bullae was caused by incomplete resection at the initial surgery in one case. Such incomplete resection and oversight were not noticed in the review of the videotape of the initial surgeries in two cases. Reinforcement of the staple-line is necessary for prevention of recurrence of pneumothorax. PMID- 10513162 TI - [Dose the administration of low-dose aprotinin contribute to an anti-inflammatory effect in coronary artery bypass grafting?]. AB - A study was conducted to determine whether the administration of low-dose aprotinin contributed to an anti-inflammatory effect in coronary artery bypass grafting. Levels of the inflammatory cytokines; IL-6, IL-8, and GEL, were measured before and after cardiopulmonary bypass, then 1, 3 and 6 days after coronary artery bypass grafting, in a group of patients given aprotinin (n = 7) and a control group (n = 15). A comparison of the levels of all these inflammatory cytokines between the two groups revealed no significant difference at any time point. This indicates, that low-dose aprotinin did not contribute to an anti-inflammatory effect in coronary artery bypass grafting. PMID- 10513164 TI - [A case who needed additional Daggett's procedure for residual shunt after infarction exclusion technique for post-infarction ventricular septal perforation]. AB - A 71-year-old woman had the surgical repair of post-infarction ventricular septal perforation with infarction exclusion technique. Three days after operation, residual shunt was observed by echocardiogram and she developed cardiac failure. Pulmonary to systemic flow ratio was 2.1, and pulmonary artery pressure was 42/23 (33) mmHg. Additional surgery for residual shunt was performed 22 days after the first operation. The part of Xenomedica patch was found loosely floating in the LV cavity. The infarcted myocardium was firm enough to closed directly, so a double Hemashield cardiovascular fabric was sutured on the left side of the perforated septum around VSP (Daggett's Procedure). The ventriculotomy was closed including the fabric with two felt strips. The postoperative course was uneventful. Though infarction exclusion technique has the advantage in many cases, much attention must be paid to prevent residual shunt. PMID- 10513163 TI - [A case of simple coarctation]. AB - We report a successful surgical repair of the simple coarctation of a 80-day-old girl by extended end-to-end aortic arch reconstruction. She was admitted to our hospital at the age of 4 days because of poor pulsation of femoral arteries. The systolic blood pressure gradient between the arm and the leg was 30 mmHg. Echocardiography on admission revealed a simple coarctation and patent foramen ovale, with the mildly impaired left ventricular contraction (left ventricular fractional shortening was 23%). Although aortography demonstrated an isolated interrupted segment at the aortic isthmus with collaterals (type A classification of Celoria-Patton), the tubular connection between the distal arch and the descending aorta, of which intralumen was obstructed with abundant ductal tissues, was found at operation. The obstruction of the lumen of aortic isthmus in our case, which was originally patent, might be caused by ductal closure and present as a simple coarctation. PMID- 10513165 TI - [Tricuspid valve replacement in a patient with idiopathic tricuspid regurgitation]. AB - Tricuspid regurgitation is often seen but is rarely categorized as "idiopathic". A 66-year-old man suspected of idiopathic tricuspid regurgitation underwent tricuspid valve replacement. With right heart failure over twenty years, his CTR increased to eighty six (86) per cent. During operation, no abnormal findings were observed except for the dilated tricuspid valve ring. Then a stented porcine xenograft (Carpentier-Edwards 33 mm) was inserted in the tricuspid valve annulus. Seven years after replacement, he died. Autopsy demonstrated myocardial hypertrophy of the right ventricle and dilatation of the right heart, supporting the diagnosis of idiopathic tricuspid regurgitation. PMID- 10513166 TI - [Two cases of the bronchial cyst located in the anterior mediastinum]. AB - Bronchial cysts are common cystic tumors around the tracheobronchial tree in the middle and posterior mediastinum and rarely locate in the anterior mediastinum. We reported two cases of the bronchial cyst located in the anterior mediastinum. One case was a 57 year-old-female. A thymic cyst was suspected and the extended total thymectomy was performed through the mediansternotomy. The microscopic examination showed bronchial epithelium and cartilage in the cystic wall. The another case was 71 year-old-male operated by thoracoscopic surgery for the cystic tumor in the anterior mediastinum. Microscopic examination showed bronchial epithelium and gland in the cystic wall. PMID- 10513167 TI - [A case of lung large cell carcinoma that survived for 8 years after resection of a solitary metastatic lymph node]. AB - A 54-year-old man who underwent Miles operation for primary rectal cancer, had undergone right upper lobectomy for large cell carcinoma of the right lung 2 years before. In the Miles operation, the large lymph node was palpable in the mesenterium near the small intestine. Small bowel resection with lymphadenectomy was performed. Histological examination showed lymph node metastatic large cell carcinoma of the lung. The patient has survived for 8 years after the second operation without recurrence. Large cell carcinoma of the lung tends to metastasize to digestive organs, so careful follow-up is necessary after surgery. Surgical treatment for a solitary metastatic lesion of lung cancer is considered to be effective and to have a satisfactory prognosis. PMID- 10513168 TI - [A case of spontaneous hemopneumothorax associated with uncontrolled massive bleeding after inserting thorathic drain]. AB - A 31-year-old man admitted to our hospital complaining of right chest pain. Chest X-ray on admission revealed a collapsed lung and an air fluid line in the right thorax. A chest tube drainage was carried out, but hemorrhagic pleural fluid was drainaged. Forty minutes later, an anemia developed and chest X-ray showed increased massive right pleural collection. Therefore, emergent surgery was performed. An operation under thoracoscopic guidance was converted into thoracotomy because of massive blood clots and fresh bleeding. A bleeding originating from the branch of 1st intercostal artery and a bulla on upper lobes were noted. The artery was coagulated with electrocoutary and ligated using Endo loop. This artery is not congenital abnormal one but collateral expanded one of which the elastic lamina is thickened. Spontaneous hemopneumothorax is life threatening, emergent operation should be undergone. PMID- 10513169 TI - [A case of lung cancer (small cell carcinoma) occurring esophago-pericardial fistula and purulent pericarditis]. AB - A 72-year-old woman who had had an endoscopic sclerotherapy for esophageal varices presented with high fever and severe cough. Chest X-ray and CT demonstrated a pneumopericardium and pericardial effusion. Esophagoscopy and esophagography revealed an esophageal perforation into the pericardial cavity and into the lung. Consequently, drainage and irrigation of the pericardial cavity and mediastinum were done for MRSA infection. However, these procedures failed to reduce the inflammation, and she expired because of liver failure soon after placing a covered stent in the esophagus. Postmortem examination revealed small cell carcinoma in the left lung invading into the esophagus and pericardium. PMID- 10513170 TI - [The effect of sevoflurane and isoflurane on striatal dopamine of awake freely moving rats observed in an in vivo microdialysis study]. AB - We investigated the effect of sevoflurane and isoflurane on the level of interstitial dopamine of in vivo awake, free moving rats brain striatum using microdialysis techniques. Rats were implanted with a microdialysis probe to the right striatum of the brain and administered with 1.2 MAC of each volatile anesthetics for 1 hour, and dialysates from the probe were determined every 20 minutes. Both anesthetics reduced the amount of dopamine derived from dialysate, and increased the efflux of dopamine with pretreatment of nomifensine 10mg. kg-1 i.p. The change of metabolites of dopamine during anesthesia was increased. No significant difference was found between sevoflurane and isoflurane. We hypothesized that these anesthetics might have special actions on interactions between metabolism and re-uptake of dopamine in rats striatum during anesthesia. PMID- 10513171 TI - [The study of the anesthetic action of halothane on the rat spinal cord by fos immunoreactivity]. AB - This study was performed to examine the anesthetic action of halothane on the spinal cord of rats by using fos immunoreactivity, a marker for neuronal activity following noxious stimulation. Sprague-Dawley rats were injected with 150 microliters of 5% formalin subcutaneously into the left hindpaw. Control group (n = 5) received 100% oxygen for 3 hours after injection. 1 MAC group (n = 5) and 1.5 MAC group (n = 5) of rats were anesthetized with 1 MAC or 1.5 MAC halothane, respectively, for 3 hours after injection. The number of fos immunoreactive cells was counted in the lumbar spinal cord of each rat. All rats showed escape reactions against noxious stimulation in the control group. In the 1 MAC and 1.5 MAC groups, two of five rats showed response to noxious stimulation and the another three showed no response, respectively. There was profound relation between the responding rats and the expression of fos immunoreactivity in the spinal dorsal horn. The number of fos immunoreactive cells decreased in the cord of rats that showed no response to noxious stimulation by halothane 1 or 1.5 MAC. These findings suggest that halothane has analgesic action on spinal nociceptive neurons in the rats on the condition that its reactions to noxious stimulation are suppressed by halothane of any concentration. PMID- 10513172 TI - [Comparison of the effect of rapid infusion of lactated and that of acetated Ringer's solutions on maternal and fetal metabolism and acid-base balance]. AB - The maternal and neonatal metabolism and acid-base balance were investigated in 20 parturients undergoing combined spinal and epidural anesthesia for cesarean delivery. Patients received intravenous infusion at a rate of either 25 ml.kg-1.h 1 of lactated (LR group, n = 10) or acetated (AR group, n = 10) Ringer's solution before anesthesia, to prevent hypotension during anesthesia. We obtained venous blood samples as follows; maternal control before anesthesia, maternal sample A after the infusion, umbilical sample B, and neonatal pedal sample C 5 h after birth, and determined lactate, pyruvate, bicarbonate, and base excess concentrations, and pH in each sample. In sample A, the lactate level was significantly higher and base excess level was significantly lower in the LR group than in the AR group. The pH of sample A and B was significantly higher in the AR group than in the LR group. However, no differences in all parameters of sample C between the two groups were observed. These results demonstrated that acetated Ringer's solution is better than lactated Ringer's solution in rapid infusion before cesarean section because of the correction of neonatal lactic acidosis. PMID- 10513173 TI - [Assessment of postoperative pain using face scale judged by nurses: comparison between hepatectomy and esophagectomy]. AB - The agreement between scores for observer-reported face scale (FS) and the self reported visual analog scale (VAS) in postoperative pain assessment has not been compared for different types of surgery and for different times in the postoperative course. Five grade FS (1-5) judged by a nurse was compared with VAS (0-100 mm) reported by patients who had undergone hepatectomy (group H, n = 60) or esophageal cancer surgery by a thoracoabdominal procedure (group E, n = 50). Postoperative analgesia was mainly achieved by epidural morphine administration combined with lidocaine or bupivacaine in both groups. Pain measurement was performed at admission to the ICU, 1, 2, 6, and 10 hours later, and the following morning in group H, and 0.5, 1, 2, 6, 10, and 14 hours after tracheal extubation in group E. VAS scores (means +/- SD) were respectively 46 +/- 29 and 31 +/- 25 at ICU admission and one hour later in group H, and 41 +/- 36 and 36 +/- 33 thirty minutes and one hour after tracheal extubation in group E. FS values (means +/- SD) were respectively 2.6 +/- 1.2 and 2.1 +/- 1.1 at ICU admission and one hour later in group H, and 2.4 +/- 1.3 and 2.2 +/- 1.2 thirty minutes and one hour after tracheal extubation in group E. VAS and FS decreased in both groups over time postoperatively. A fair degree of agreement was found between VAS and FS scores in group H at ICU admission and one hour later (weighted kappa values = 0.29 and 0.28, respectively); on the other hand, good agreement between these two scores was found in group E thirty minutes and one hour after tracheal extubation (weighted kappa values = 0.67 and 0.62, respectively). Weighted kappa values decreased thereafter in group E, but did not change in group H over the postoperative course. We conclude that postoperative pain assessment based on facial expression is more useful early after extubation for patients who have undergone esophagectomy than for those who have undergone hepatectomy. PMID- 10513174 TI - [Total intravenous anesthesia with propofol, fentanyl and ketamine for carotid endarterectomy under somatosensory evoked potential monitoring--combination with intraoperative hypothermia]. AB - We reported anesthetic management combined with hypothermia for carotid endarterectomy under somatosensory evoked potential monitoring. Anesthesia was induced by propofol, fentanyl and ketamine, and maintained by infusion of propofol and ketamine and intermittent injections of fentanyl. Perioperative hypothermia was induced by gradually reducing the temperature of a circulating water mattress underneath the body to 15 degrees C. Additionally, somatosensory evoked potential monitoring was performed and recordings were obtained immediately after induction of anesthesia, and before as well as during cross clamping of the internal carotid artery. Rectal temperature was reduced to 33.7 degrees C when cross-clamping of carotid artery was carried out, but major changes between before and during the procedure was not observed. All procedures were done uneventfully and gradual rewarming was accomplished by electric blanket. No neurological deficits were observed following recovery from anesthesia. Total intravenous anesthesia with propofol, fentanyl and ketamine may be useful for carotid endarterectomy under hypothermia and somatosensory evoked potential monitoring. This method may provide neuronal protection against ischemia injuries induced by cross-clamping of the carotid artery. PMID- 10513175 TI - [The effect of intraoperative mild hypothermia on the development of perioperative cardiac ischemia]. AB - We evaluated the risk of perioperative cardiac ischemia associated with mild hypothermia as adjunct management for neurosurgical procedures. Forty-seven elective neurosurgical patients were randomly assigned to either hypothermic group (H, n = 24) or normothermic one (N, n = 23). Patients in group H were cooled to and maintained at 34.5 degrees C (tympanic membrane temperature) and rewarmed after main neurosurgical manipulation, while those in group N were kept in normothermic state. Cardiac ischemia was diagnosed with Holter electrocardiogram monitored for 24 hours after admission to the operating room. No differences were observed in demographic data including age, gender, weight, height, rate of having cardiac disease, anesthesia methods, and contents of surgery. Temperature was significantly lower in group H than N both at the lowest point (34.6 +/- 0.5 vs 35.9 +/- 0.6, mean +/- SD) and at the conclusion of anesthesia (35.9 +/- 0.9 vs 36.5 +/- 0.5). Electrocardiographic ST segmental depression was observed in 3 (group H) and 5 cases (group N), and postoperative shivering occurred in 3 (group H) and 2 cases (group N), respectively. Those incidences were not statistically significant (chi-square test, P was set at 0.05). We concluded that intraoperative mild hypothermia might not increase the risk of perioperative cardiac ischemia in patients for neurosurgical procedures. PMID- 10513176 TI - [Perioperative considerations for a holoprosencephaly patient]. AB - We had twelve anesthetic experiences of seven holoprosencephaly patients for the past thirteen years. We classified these seven patients and compared the difficulty in perioperative control in the patients. Seven patients were divided according to two different holoprosencephaly classifications, one is that by DeMyer and the other by Osaka. We studied them retrospectively about airway management, the control of convulsion or instability of body temperature, and so on. Case of abortive type had nothing particular to be considered about, but cases of semilober type which had more severe brain anomaly, required careful respiratory management, due to their immature nasopharyngeal function. Concerning the convulsion or body temperature, cases of semilober type were difficult to control. When we anesthetize these holoprosencephaly patients, we must consider the classification of the anomaly and clinical conditions in each case. Especially, in cases of semilober types, prudent treatment is needed thoughout the perioperative period. PMID- 10513177 TI - [A patient with pulmonary alveolar proteinosis who underwent whole lung lavage under total intravenous anesthesia with propofol]. AB - A 32-year-old male suffering from pulmonary alveolar proteinosis underwent whole lung lavage under one-lung ventilation and total intravenous anesthesia with propofol and fentanyl. The lung lavage was initially performed for the right lung, and for the left lung two weeks later. Spo2 and Pao2 declined at the time of lung degassing and drainage of lavage fluid (minimum values; Spo2 = 83%, Pao2 = 55.9 mmHg), and recovered on filling with the lavage fluid. Although pulmonary arterial pressure and central venous pressure were slightly increased at the time of unilateral lung filling, there was no marked change in arterial pressure, heart rate or cardiac output for lavage of either lung. % VC, DLCO and PaO2 markedly improved after whole lung lavage, from 26.0%, 5.84 ml.min 1.mmHg-1, and 61.9 mmHg in room air to 41.6%, 9.96 ml.min-1.mmHg-1, and 89.3 mmHg in room air, respectively. Since propofol does not inhibit hypoxic pulmonary vasoconstriction and is able to maintain a stable level of anesthesia, it is a useful anesthetic agent for whole lung lavage. PMID- 10513178 TI - [Urinary retention as a transient neurologic symptom after accidental total spinal anesthesia with mepivacaine hydrochloride]. AB - We report a patient in whom urinary retention as a transient neurologic symptoms (TNS) developed after accidental total spinal anesthesia with mepivacaine hydrochloride. Mepivacaine, an amide local anesthetic, has been used for spinal anesthesia and considered one of the best for spinal anesthesia for its low incidence of TNS. However, we suggest that TNS associated with mepivacaine might not be a rare complication in spinal anesthesia. PMID- 10513179 TI - [Anuria due to bilateral renal artery spasm during hysterectomy and oophorectomy]. AB - A decrease in urinary volume during surgery is often encountered. Usually it can be treated with intravenous fluid or diuretics. We here report a rare case of intraoperative anuria in which renal blood flow ceased totally. The patient was 36 year old female (166 cm 50 kg), who was admitted for a investigations of long term severe hypertension of unknown origin. Radiographic examination showed no adrenal tumor but a right ovarian cyst was found and suspected to be malignant, for which oophorectomy was indicated. After epidural catheterization, general anesthesia was induced by intravenous propofol and vecuronium, and maintained with epidural lidocaine and the inhalation of isoflurane and nitrous oxide mixed with oxygen. During surgery, urinary outflow decreased gradually leading to total anuria, which was resistant to intravenous fluid and furosemide. Intraoperative pyelography was performed and both kidneys and urinary tracts were not visualized. After the surgery, when the patient returned to the ward, urine began to flow. Postoperative pathological examination of the removed ovary showed a presence of renin excreting tumor cells. The anuria was considered to be the result of transient spastic obstruction of bilateral renal arteries, presumably in response to a high level of plasma renin. PMID- 10513180 TI - [Repeated postoperative hypoglycemia in a woman with multiple pheochromocytomas]. AB - A case of repeated postoperative hypoglycemia following removal of multiple pheochromocytomas in a non-diabetic 23-year-old woman is presented. Although the hypoglycemia did not occur after the first operation, it was recognized after the second and third operations. The blood sugar levels were 54 mg.dl-1 and 25 mg.dl 1, respectively, and continuous intravenous glucose infusion was necessary for about 15 hours postoperatively. This complication may be related to sudden withdrawal of catecholamines. Frequent monitoring of blood sugar level as well as cardiovascular system is important for perioperative management of pheochromocytoma. PMID- 10513181 TI - [The anesthetic management of a patient with a congenital mediastinal cyst undergoing video-assisted thoracoscopic surgery (VATS)]. AB - We recently conducted video-assisted thoracoscopic surgery (VATS) on an 8-day-old boy in whom a congenital mediastinal cyst had caused obstruction of the left main bronchus. After the surgery, the cyst recurred and caused reobstruction of the airway. When he was 51-day-old, we conducted VATS again. We could not perform cystectomy by VATS because the left lung was inflated, and we conducted cystectomy by thoracotomy. PMID- 10513182 TI - [Circulatory disaster following infiltration of epinephrine contained in local anesthetic]. AB - There are many case reports regarding unexpected circulatory responses such as hypertension, dysarrythmias, pulmonary edema and catecholamine myopathy, following infiltration of epinephrine into surgical fields. These cases occurred under general anesthesia, whereas we report a case which occurred with local anesthetic use only, with manifested severe circulatory system response to a small dose of epinephrine administered with lidocaine. The case is a 53 year old female with tongue cancer. A skinflap of the formerly operated tongue was infiltrated with 4 ml of 0.5% lidocaine and 0.0005% epinephrine for local anesthesia and hemostasis prophylaxis. Soon thereafter, the patient developed ventricular tachycardia, severe hypertension and pulmonary edema, but was treated successfully. It is important to recognize the risk of severe circulatory system response which may be elicited by a small dose of epinephrine contained in local anesthetics whether under general anesthesia or not. PMID- 10513183 TI - [Anesthetic management of a patient with postthymectomy myasthenia gravis]. AB - A 56-year-old male who had received total thymectomy for treatment of myasthenia gravis was scheduled for sigmoidectomy under general anesthesia. Since his symptoms had become worse after the thymectomy along with increased anti acetylcholine receptor antibody titer, preoperatively we could not estimate his sensitivity to non-depolarizing muscle relaxants. We initially tried tracheal intubation without using a non-depolarizing muscle relaxant immediately after intravenous injection of propofol 2 mg.kg-1 and fentanyl 4 micrograms.kg-1. Since the intubation was unsuccessful, however, vecuronium 0.01 mg.kg-1 was repeatedly administered until TOF ratio reached 0%. Successful intubation was performed with 3.5 mg of vecuronium. We conclude that the initial trial of tracheal intubation should be performed without a non-depolarizing muscle relaxant in patients with myasthenia gravis whose symptoms have become worse after thymectomy. If first attempt is unsuccessful, the tracheal intubation should be performed with a smaller dose of vecuronium using an electrical nerve stimulator. PMID- 10513184 TI - [Anesthetic management of a patient with Coffin-Lowry syndrome]. AB - Coffin-Lowry syndrome (CLS) is characterized by mental retardation, a peculiar face and deformities of the thorax and spine. A 33-year-old female with Coffin Lowry syndrome (CLS), further complicated with atrial septal defect and ventricular tachycardia, underwent elective surgery for anterior cervical cyst. As difficult intubation had been anticipated, anesthesia was induced with continuous administration of propofol. After confirming that she could be ventilated by mask, vecuronium bromide, midazolam and fentanyl were given. The operation and anesthesia were conducted uneventfully. No complications occurred postoperatively. The use of propofol for slow induction of anesthesia was advantageous for hemodynamic stability in this case. PMID- 10513185 TI - [Propofol anesthesia for a patient with congenital myotonic dystrophy]. AB - We report the anesthetic management for a five year old boy with congenital myotonic dystrophy. The patient was scheduled for bilateral orchiopexy under general anesthesia. Anesthesia was induced with fentanyl 50 micrograms, vecuronium 0.6 mg and propofol 40 mg intravenously to facilitate tracheal intubation. During operation, we monitored train of four ratio (TOF) to confirm effect of muscle relaxation. Anesthesia was maintained with propofol (2 mg.kg 1.hr-1), nitrous oxide and caudal block. At the end of the operation, the patient recovered smoothly from anesthesia and post-operative course was uneventful. Congenital myotonic dystrophy presents many problems for the management of general anesthesia, because of respiratory or circulatory complications. In this case, we were careful not to use drugs which may cause respiratory or circulatory depression. We have demonstrated that anesthesia with propofol is a safe method for the anesthetic management of a patient with this disease. PMID- 10513186 TI - [Is local anesthesia necessary for spinal needle insertion?]. AB - It is a routine procedure for anesthesiologists to use local anesthesia (LA) before spinal needle insertion (SNI), but LA itself produces pain on injection. We evaluated the necessity of LA before spinal block using a 25-gauge needle by questioning whether LA makes SNI painless and easy. Sixty patients without lumbar abnormality for spinal block were allocated to 3 groups: Group A, LA with 2 ml of 1% lidocaine using a 24-gauge needle; Group B, LA with 0.5 ml of 1% lidocaine using a 27-gauge needle; Group C, without LA. Visual analog pain scales of LA and SNI were obtained respectively. Though LA reduced the pain associated with SNI, total pain scales in Group A and Group B were significantly larger than those in Group C. The pain scale of LA was significantly lower in Group B than Group A, but there were no significant differences in the pain on SNI between the Groups. The times needed for SNI were not significantly different among the three Groups. In conclusion, LA with 2 ml of 1% lidocaine using a 24-gauge needle is not useful for pain relief on spinal block using a 25-gauge needle. Intradermal anesthesia using a 27-gauge needle is preferable to reduce the pain on SNI, if LA is necessary. PMID- 10513187 TI - [The evaluation of preoperative anesthetic visit in our hospital]. AB - The preoperative visit by an anesthetist has been thought to be important for the assessment of patient and to communicate with them. However, there are few reports on the visit in Japan until now. The effect of preoperative anesthetic visit in our hospital was estimated by interviewing patients just before surgery who had received a visit by their anesthetist. The answers from sequential 346 scheduled surgical patients for 8 weeks were obtained. Only 72 patients were able to recall the name of their anesthetists. About half of patients believed that anesthetist just had put the patient to sleep and relieved them from pain. We measured the number of treatments each patient could remember that had been explained by the anesthetist on the visit, and found it was unexpectedly small at the interview. These data suggest that our preoperative visit may not be satisfying in view of making good relationship between patients and anesthetists, and educating patients for recent anesthesia. We should make an effort to educate the patients about up-to-date and reasonable anesthesia. PMID- 10513188 TI - [The importance of occupational standards regulation for the environmental factors in the maintenance of public health]. PMID- 10513189 TI - [Dust as a factor in crystalline silicon production]. AB - Up-to-date crystalline silicon production is characterized by such leading occupational hazards as higher level of dust with complicated chemical composition, aerosols that vary with technology and derive from disintegration of silica, crystalline silicon or condensation of silica. Experiments comparing prevalence of diseases caused by silica and crystalline silicon dusts in acute and chronic intratracheal administration and inhalation identified crystalline silicon as moderately fibrogenic and specified its MAC at 4 mg/m3 in the air of workplace. PMID- 10513190 TI - [Metabolism of serum phospholipids in coal miners]. AB - The studies covered metabolism of serum phospholipids during lipids peroxidation and hydrolysis by phospholipase A2. Metabolism of serum phospholipids appeared to depend on duration of exposure to mine dust and on coal pneumoconiosis stage. Lipids peroxidation becomes activated after 20 and more years of service, intensifies with anthracosilicosis development on background of higher catalase activity that is low on early stages of the disease. Activity of phospholi pase A2 increases with pulmonary fibrosis progression. PMID- 10513191 TI - [Parameters of oxidative metabolism, neuro-humoral and hormonal regulation in the condensed exhaled air in early stages of anthracosilicosis]. AB - For early diagnosis of anthracosilicosis, the authors studied oxidation parameters, neuromediators and hormones in expired air condensate in coal miners. Premorbid stage of anthracosilicosis is characterized by activated lipids peroxidation, high levels of neuromediators (acetylcholine, catecholamines), tissue hormones (histamine, serotonine) and lower contents of adaptive hormones (cortisol, triiodothyronine, thyroxine). PMID- 10513192 TI - [Principles and methods of the evaluation of occupational risks in the complex exposure to pesticides]. AB - The authors suggested integral criterion for complex evaluation of pesticides variety and dose loads specific for the occupation. Therefore, pesticides could be classified according to toxicologic criteria and application conditions. Recommendations include management to minimize risk associated with pesticides. PMID- 10513193 TI - [The main pathogenetical mechanisms of occupational lung diseases of dust-related etiology]. PMID- 10513194 TI - [Specific aspects in the classification of pneumoconioses in 1996]. PMID- 10513195 TI - [On the new classifications of pneumoconioses]. PMID- 10513196 TI - [Organizational and structural principles of the construction of territorial centers of occupational prophylaxis]. AB - In organization-normative period of Russian occupational pathology service establishing expediently to foresee some different levels for territorial occupational pathology institutions with corresponding differentiation for three category. The appointing a category to the occupational pathology centre must be in competence of accreditive-licencive commission, which follows "On system of medical practice licencing in the part concerning occupational pathology". PMID- 10513197 TI - [The use of pharmacological tests for the diagnosis of the nature of bronchial obstruction in coal miners with premorbid stage of chronic dust-related bronchitis]. AB - Bronchospasm in premorbid chronic dust bronchitis in coal miners is caused mostly by dystonic effects of sympathetic and parasympathetic nervous system, and minimally by hypersecretion and swelling of respiratory tract mucosa. Pathogenetic features of obstruction in premorbid chronic dust bronchitis are lower sympathetic tone (major feature) and higher parasympathetic tone (minor one). Those items should be considered in therapeutic and prophylactic tactics. PMID- 10513198 TI - [The treatment of chronic bronchitis caused by mineral and organic dust]. AB - Stage I of chronic bronchitis due to occupational dust exposure is characterized by pathologic changes of distal pulmonary area without clinical obstruction--that evidence should be considered in occupational examinations and placement. Early and continuous rational therapy for chronic bronchitis patients, better work conditions led to lower occurrence of deteriorated chronic bronchitis with marked restrictive and obstructive changes. PMID- 10513199 TI - [Aspects of clinico-social rehabilitation of patients with occupational allergic dermatoses]. PMID- 10513200 TI - [The development and the implementation of screening research for the diagnosis of chronic respiratory diseases in textile industry workers]. PMID- 10513201 TI - [Immunotherapy in the eye]. PMID- 10513202 TI - [Localization of extracellular matrix proteins after holmium YAG laser radiation in rat cornea]. AB - PURPOSE: To understand corneal responses to holmium YAG (Ho: YAG) laser radiation, we used immunofluorescent microscopy to examine changes in the localization of extracellular matrix proteins. METHODS: Rats were radiated with an Ho: YAG laser. On days 1, 3, and 7 after radiation, the eyes were enucleated and frozen. The cryosections were stained by immunofluorescent microscopy using antibodies against type I collagen, fibronectin, type IV collagen, and laminin. RESULTS: One day after Ho: YAG laser radiation, contraction of the stromal collagen fibrils was observed. Keratocytes could not be observed at the radiated stromal region on day 1 after radiation. One week after radiation, keratocytes returned to the radiated area. Although the stromal collagen fibrils were contracted, they were stained by an antibody against type I collagen. Dense fluorescence for fibronectin was observed at the margin of the stromal acellular zone. Both laminin and type IV collagen were observed at the basement membrane under the corneal epithelium regardless of whether the corneas were radiated or not. CONCLUSIONS: These results suggest that Ho: YAG laser radiation might be useful for collagen contraction of the stroma, without causing serious damage to the corneal epithelium or the basement membrane. PMID- 10513203 TI - [The effect of substance P with insulin-like growth factor-1 on corneal epithelial cell proliferation]. AB - PURPOSE: To elucidate the pathogenesis of neuroparalytic keratopathy, we examined the effect of substance P (SP) on corneal epithelial cell proliferation in comparison with that of insulin-like growth factor-1 (IGF-1) in vivo and in vitro. METHODS: In the in vivo study, the epithelium of rabbit cornea was removed mechanically and treated with eye drops containing SP, IGF-1, SP + IGF-1, or physiological saline as a control. Corneas were excised 48 hours after the removal of the epithelium, labeled with 3H-thymidine, and subjected to autoradiography. An in vitro study was also performed by culturing rabbit corneal epithelial cells in culture medium containing SP with or without IGF-1 and labeling cells with 3H-thymidine for a subsequent autoradiographical study. FINDINGS: In the in vivo study, SP in combination with IGF-1 enhanced the healing of rabbit corneal epithelial defect. 3H-thymidine autoradiography revealed that in both in vivo and in vitro studies, SP + IGF-1 stimulated corneal epithelial cell proliferation, but SP or IGF-1 alone did not. CONCLUSION: These results indicate that SP enhances the stimulatory effect of IGF-1 on corneal epithelial cell proliferation and accelerates corneal epithelial healing. PMID- 10513204 TI - [Changes in cytoskeletal proteins in childhood cataract lenses]. AB - PURPOSE: Approximately 50% of congenital and childhood cataract seen in the clinic is of undetermined origin. Biochemical analysis of the cataracts is rare. This study analyzes lens proteins to determine the mechanism of congenital and childhood cataracts. METHOD: We analyzed the lens proteins from 10 young patients after cataract operations, using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), densitometry analysis, and western immunoblotting. RESULTS: Densitometry of separated proteins showed a decrease in the high molecular protein bands of posterior subcapsular cataract (PSC). Specifically, spectrin (235 kDa), filensin (100 kDa), and vimentin (57 kDa) were absent from the SDS-PAGE of PSC. Increases in filensin and vimentin were observed in a Christmas tree cataract and lamellar cataracts. Western immunoblots confirmed the densitometry of SDS-PAGE. CONCLUSION: These results suggest that changes in cytoskeletal proteins may contribute to congenital and childhood cataract. PMID- 10513205 TI - [Epiretinal membranous tissue in retinal detachment due to macular holes in highly myopic eyes]. AB - OBJECTIVE: To determine the frequency of occurrence of epiretinal membranous tissue in retinal detachment due to macular hole and to evaluate the efficacy of vitreous surgery including removal of the membrane. DESIGN: Noncomparative case series. PARTICIPANTS: Eighteen highly myopic eyes with retinal detachment due to macular holes. RESULTS: Epiretinal membranous tissue was removed from 7 eyes (39%) and retinal reattachment was obtained in 14 eyes (78%) after the initial vitreous surgery. Of 4 eyes that required reoperation, 2 eyes achieved reattachment after a long term of internal tamponade. Histological examination of an epiretinal membrane from one case revealed that it was composed of glial cells, connective tissue, and fragments of inner limiting membrane. An intraoperative use of autologous serum as a tissue adhesive for macular holes had no effect on surgical success. CONCLUSION: The first therapeutic choice for this disease should be vitreous surgery with removal of the epiretinal membrane when practical, though the membrane can not always be identified in every case. PMID- 10513206 TI - [A case of chronic iridocyclitis in young girls--surgery for complicated cataract and effect of cyclosporin A]. AB - PURPOSE: To report a case of chronic iridocyclitis in young girls and the effect of systemic cyclosporin A for inflammation following surgery for complicated cataract. CASE: A 6-year-old girl presented with white uveitis, complicated cataract, and band keratopathy in both eyes. We diagnosed her as having chronic iridocyclitis in young girls. Eight months later, the anterior chamber in the left eye became flat. Ultrasound biomicroscopy (UBM) showed the presence of cyclitic membrane and adhesion of the ciliary body to the lens capsule. Phacoemulsification with pars plana vitrectomy was used to remove the lens, anterior vitreous, and cyclitic membrane. Severe inflammation and hypotony developed after surgery and persisted after systemic and topical corticosteroids. Systemic cyclosporin A induced rapid resolution of inflammation. CONCLUSION: Systemic cyclosporin A was useful for postoperative inflammation in this case of uveitis of young girls. PMID- 10513207 TI - [Impression on the 11th Specialist Licensure examination by the Japanese Ophthalmological Society]. PMID- 10513208 TI - [Pathophysiology of established urinary incontinence (UI) in the elderly and an ameliorative method for disuse syndrome including UI seen in the bed-ridden elderly--using prone position and its variations]. AB - Multipathology is the physical characteristics of the elderly, and their established urinary incontinence (UI) is usually based on multiple causal diseases and types of UI. Decubitus voiding (urination and defecation) inevitably causes UI and fecal incontinence (FI). Difficulty in controlling UI and FI seen in bed-ridden elderly results in long-term use of diapers and indwelling catheters, which eventually leads to the progression of disuse syndrome and decline of ADL (activity of daily living). Most elderly UI cases have only a few major causal diseases. Arrangement of relationship between the diseases and the existing types of the UI; recognition of gender gap in urination; determination and execution of various kinds of treatment in a methodical way; and the maintenance of the proper medication dosage are the four keys to the effective and safe control of the UI. For bed-ridden elderly, passively provided suitable positions on urination and defeca-tion has become a prerequisite condition. These are not supine or Fowler (semi-reclining) positions, but normal sitting, or prone and its variation forward-tilting positions. The latter two positions, which ameliorate UI and FI in the bed-ridden elderly, have been found to improve all of the pathologies including the disuse syndrome as well. The Seikatsudai (Life rack), which provides a forward-tilting position, not only has such an effect, but also offers the possibility to make the bed-ridden elderly more independent in their lives. PMID- 10513209 TI - [Mechanism of insulin resistance]. PMID- 10513210 TI - [Quality of life in elderly patients with cerebral vascular disease and Parkinson's disease]. AB - We developed a questionnaire for the study of background factors and quality of life (QOL) in elderly patients with cerebral vascular disease (CVD) and Parkinson's disease (PD). The questionnaire covered the background factors and four sections such as physical, functional, psychological and social health sections. Each section had 15 questions and disease-specific questions for CVD or PD were included in the physical health section. We analyzed 107 patients with CVD (76 elderly patients, aged 65 or more, 31 non-elderly patients under 65) and 136 patients with PD (91 elderly, 45 non-elderly). In the background section, of a total of 243 patients with CVD and PD, the elderly patients needed the assistance of their spouse and their sons wives more frequently than non-elderly patients. With regard to rehabilitation, non-elderly CVD patients had rehabilitation more frequently than the elderly CVD patients, while a higher percentage of elderly patients with PD had rehabilitation training more frequently than the non-elderly PD patients. In the QOL section, there was no difference between elderly and non-elderly CVD patients, while elderly PD patients were statistically more significantly disabled physically and weak minded psychologically. The physical disabilities of the elderly PD patients in this statistical investigation included slow motion, stooped posture, frozen gait, difficulty in turning and standing up, constipation and dysuria. The psychological problems of elder PD patients included forgetfulness and a feeling of aging. These patients had significantly fewer consultations by family and relatives than the non-elderly PD patients. The overall tendency of QOL in patients with CVD and PD was similar to that of PD patients. PMID- 10513211 TI - [Qualitative change in the elastin from the calcified portion of human artery]. AB - To examine the qualitative changes of elastin and the aorta related to calcification of human arteries, biochemical properties were measured, including calcium (Ca), phosphorus (P) and magnesium (Mg) contents in the aorta or in the elastin fraction in calcification, cholesterol content in atherosclerosis, desmosine content of cross-link, free thiol contents (free SH/total SH) and hydrophobic properties in the elastin fraction from the calcified portion, adjacent sites and another normal artery. The results from different sites of the calcified abdominal artery are as follows: The contents of Ca, P and Mg in aorta and the elastin fraction from the calcification site were higher than those at other sites. Moreover, Ca in the aorta and elastin fraction correlated positively with P and Mg. The content of cholesterol in the calcification site was the same as at other sites and did not correlate with Ca, P or Mg. The content of desmosine in the calcification site was significantly lower than that in different sites. In addition, its content was negatively associated with Ca and P in the elastin fraction and with the aortic Mg. The content of free thiol in the calcification site was similar to the other sites and correlated negatively with Ca and P in the aorta. The hydrophobicity in the calcification was similar to that at other sites, and was negatively associated with Ca and Mg in the elastin fraction. PMID- 10513212 TI - [Comparison of interviews and review of patient chart in evaluating falls among geriatric patients]. AB - This study compared the reliability of interviewing and reviewing medical records to determine falls experienced by elderly patient admitted to a geriatric hospital. Subjects were 130 consecutive patients (mean age 76 years) who received physical therapy. They were divided into two different age groups: those aged below 75 years and those aged 75 years and over. In reviewing medical records, it was difficult for researchers to find out about falls based on information by patients and families, if no injury occurred. This tendency remarkable in patients aged below 75 years. Because of it, the rate of falls was significantly lower and that of injuries was higher than in the interviews. We conclude that review of medical records involves problems concerning the accuracy of information about falls. Hip fractures due to falls were seen only in patients aged 75 years and over, so we recognized that it is important to prevent falls especially in this group. PMID- 10513213 TI - [COP-BLAM therapy for a Hodgkin's disease in the elderly]. AB - Hodgkin's disease (HD) is a disorder with a better prognosis than non-Hodgkin's lymphoma and it predominantly affects young persons. In association with the aging of the population, however, HD has been increasing among persons aged 65 years and over in recent years. We used the COP-BLAM regimen to treat elderly patients with HD, and responses and adverse reactions were investigated. A total of 14 patients with HD treated at our department between April 1987 and December 1997 were included in this study. The patients were 8 men and 6 women aged 65 years or older, with a median age of 68 years. Five patients with clinical stage I or II disease, who had factors indicating a poor prognosis, received 3 courses of the COP-BLAM regimen with additional regional therapy of the involved field (IF). Six courses of COP-BLAM were administered to 9 patients with stage III or IV disease. The treatment was evaluable in all patients. Treatment achieved a complete remission (CR) in 12 (85.7%) of the 14 patients and a partial remission in 2 (14.3%). The CR rate was 100% for stage I or II and 77.8% for stage III or IV. The overall 5-year survival rate was 76.2% and overall disease-free 5-year survival rate was 75.7%. Adverse reactions included grade 3 or higher leukopenia in 35.7% and grade 3 or higher thrombocytopenia in 7.1%. Grade 3 or higher non hematological toxicity included stomatitis and peripheral neuropathy in one patient each. From these results, we concluded that the COP-BLAM regimen was safe for elderly patients with HD and could achieve prolongation of survival. PMID- 10513214 TI - [Large cell carcinoma of the lung with metastasis of the gastric submucosa]. AB - A 66-year-old man was admitted with dyspnea on exertion and an abnormal shadow on chest roentgenogram. Transbronchial biopsy yielded a diagnosis of large cell carcinoma of the lung. His dyspnea improved following irradiation and corticosteroid treatment and one month later, he was admitted again for chemotherapy. Because occult blood in stool was detected, upper gastrointestinal endoscopy was performed. Gastric submucosal large cell carcinoma was diagnosed, and this was considered to be metastatic from the lung. Such cases diagnosed prior to death are rare. PMID- 10513215 TI - [An autopsy case of pulmonary embolism due to renal angiomyolipoma in an elderly woman]. AB - An 86-year-old woman with a 13-year history of hypertension was admitted because of consciousness disturbance, hypotension, tachycardia, and cyanosis at her extremities. Enhanced computed tomography showed a thrombus in the truncus pulmonalis and right pulmonary artery, and also showed a left renal mass and a right renal cyst. Under a diagnosis of pulmonary embolism we started anticoagulant therapy, but the patient died five days after admission. At autopsy, a saddle-like thrombus was found in the truncus pulmonalis and bilateral trunks of pulmonary arteries. Microscopic examination showed smooth muscle cells in the thrombus. We could not find any other thrombus in the inferior vena cava, intrapelvic veins, nor in veins of lower extremities by milking. We also found tumors in both kidneys. Microscopically all tumors were diagnosed as angiomyolipoma. There were many fibrin thrombi in the sinuses of the tumors but there was no evidence of malignancy. We finally diagnosed pulmonary embolism due to renal angiomyolipoma because there was no other thrombus origin and microscopically the same smooth muscle cells were found both in the renal tumor and the pulmonary thrombus. There is only one case report concerning pulmonary embolism due to renal angiomyolipoma which happened during operative treatment. The treatment method of renal angiomyolipoma is determined by tumor size and symptoms, and usually intensive treatment is not performed in cases without symptoms. Our patient had no symptoms until the onset of severe complication of pulmonary embolism, suggesting that radical treatment is necessary for renal angiomyolipoma with a thrombus even when there are no symptoms. PMID- 10513216 TI - [Molecular evolution of hepatitis viruses]. PMID- 10513217 TI - [Imported intestinal infectious and parasitic diseases]. PMID- 10513218 TI - [The relationship between the histopathological atypia and expression of beta catenin in colonic neoplasms resected by endoscopy; comparison with that of p53 protein]. AB - Previous studies have reported predominantly nuclear localization of beta-catenin as a role for colorectal carcinogenesis. In this study, we examined the immunohistochemical expression of beta-catenin and p53 protein in 90 colonic neoplasms {33 carcinomas in adenoma (CIA), 28 high grade adenomas and 29 low grade adenomas}, resected by colonic endoscopy. Out of 33 CIAs. 28 (84.8%) cases showed predominantly nuclear localization of beta-catenin, and that was significantly higher than those of both high grade (46.4%) and low grade (13.8%) adenomas. The positiveness of p53 expression in CIAs was 51.5% (17/33), while 17.9% in high grade and 3.4% in low grade adenomas. However, there was no correlation between both protein expressions (p = 0.3472, chi 2 test). The results suggests that nuclear localization and accumulation of beta-catenin is earlier event than that of p53 mutation in adenoma-carcinoma sequence, and is useful as a marker in histopathological diagnosis for malignant conversion as well as p53. PMID- 10513219 TI - [A case of neuroendocrine cell carcinoma of the rectum]. PMID- 10513220 TI - [A case of neuroendocrine cell carcinoma of the ascending colon]. PMID- 10513221 TI - [A case of diffuse infiltrating ileal cancer with metachronous and synchronous malignancies]. PMID- 10513222 TI - [A case of mixed type hepatoma with sarcomatous change]. PMID- 10513223 TI - [A case of ectopic ACTH syndrome associated with liver metastasis of pancreas head gastrinoma]. PMID- 10513224 TI - [Abdominal wall hernia diagnosed by ultrasonography]. PMID- 10513225 TI - Maternal immunization. PMID- 10513226 TI - Screening contacts of children with tuberculosis: an important and worthwhile part of case management. AB - The outcome of screening the household contacts of 49 newly diagnosed tuberculous children as currently practised in the Paediatric Unit of the Port Moresby General Hospital is described. The screening program generated 182 chest X-rays and 67 Mantoux tests. 32 (39%) of 83 child contacts and 11 (11%) of 99 adults were commenced on antituberculous therapy, and 2 children aged 6 months were started on INAH chemoprophylaxis. Adult contacts were identified in 11 (22%) of the 49 families screened. Such a program is an extremely important part of the case management of children with newly diagnosed tuberculosis and their families. PMID- 10513227 TI - Parasitological response of Plasmodium falciparum infection to chloroquine treatment in malaria patients in Port Moresby. AB - A 7-day in vivo test system was applied to assess the parasitological response to chloroquine treatment in patients with falciparum malaria in the Central Province and National Capital District of Papua New Guinea. 30 patients were investigated but only 23 took a full course of chloroquine and were completely followed up. Of the 23 patients, 13 (57%) were negative for malaria parasites on day 2, 4 (17%) had significantly reduced parasitaemia by day 2 and cleared parasites by day 7, and 1 (4%) showed a partial response (R2). In 5 (22%) of the patients resistance at the R3 level was observed. The indication from this study is that chloroquine should continue to be the first-line drug for the treatment of uncomplicated falciparum malaria. However, judicious use of chloroquine in uncomplicated falciparum malaria is required to halt the spread of chloroquine-resistant strains of Plasmodium falciparum. PMID- 10513228 TI - Ward Six Psychiatric Unit at the Port Moresby General Hospital: a historical review and admission statistics from 1980 to 1989. AB - OBJECTIVE: The objective of this study was to document the acute psychiatric service offered by the Ward Six Psychiatric Unit at the Port Moresby General Hospital by means of admission statistics. METHODS: The study was designed to cover the period 1980 to 1989, for which reliable medical records were available. Data were collected on the total number of psychiatric admissions per year, diagnostic classification, occupation, province of origin of the patients, age and sex. A brief history of Psychiatric Ward Six is added. RESULTS: The results showed that the total number of admissions to Ward Six from 1980 to 1989 was 725. There were 462 (64%) male and 263 (36%) female patients. The ratio of male to female patients was 1.8 to 1.0. Diagnostic classification of the patients was done by the International Classification of Diseases (Ninth Edition). The most common diagnosis was schizophrenia with 358 patients (49%). The majority (63%) of the patients were unemployed. A large number of the patients, 295 (41%), were from Central Province. The young age group 21-30 years accounted for 267 (37%) of the patients. The mean annual incidence for the ten-year period of the study was 5.4 patients per 10,000 population. There was an increase in the annual incidence from 3.6 per 10,000 population in 1983 to 7.9 per 10,000 population in 1989. CONCLUSION: In developing countries, including Papua New Guinea, hospital utilization studies and statistics provide an initial source of information. These may be followed later with community surveys and field surveys when more resources including funding become available. PMID- 10513229 TI - Reference ranges for serum creatinine and urea in elderly coastal Melanesians. AB - Mean values and reference ranges are presented for serum creatinine and serum urea in Melanesian men and women aged over 50 years from coastal Papua. The values are presented separately for three age groups, 51-60, 61-70 and 71-85 years, but there was no significant difference between them. The values for women were lower than for men in all age groups. PMID- 10513230 TI - A case of factor V deficiency presenting as menorrhagia. AB - Factor V deficiency is a rare hereditary disorder. We report a patient with factor V deficiency who presented with menorrhagia and pelvic haematoma. The Haematology Department at the Royal Brisbane Hospital performed the definitive factor assays leading to the diagnosis. The challenges of her management were obtaining adequate supplies of factor V and her socioeconomic circumstances. The main future challenge will be the supervision of her pregnancies. PMID- 10513231 TI - Prevalence of malaria species observed at Port Moresby General Hospital from 1988 to 1996. PMID- 10513232 TI - "Loss prevention". PMID- 10513233 TI - "Loss prevention". PMID- 10513235 TI - "Loss prevention". PMID- 10513234 TI - "Loss prevention". PMID- 10513236 TI - All aboard for physician unions: collective bargaining efforts for doctors are picking up steam. PMID- 10513237 TI - Domestic violence: raising awareness all over again. PMID- 10513238 TI - Loss prevention case of the month. The same lesson again and again. PMID- 10513239 TI - An unusual peripheral vascular response to dopamine in a neonate. AB - We report a case of a neonate who developed hypotension immediately after birth, and needed dopamine infusion to sustain his blood pressure and tissue perfusion. He developed cyanosis of his extremity immediately after dopamine was started via peripheral line and improved spontaneously after dopamine was stopped. This happened repeatedly at various sites and at lower concentrations of dopamine. Subsequently, dopamine was replaced by dobutamine and the patient did well. We conclude that some neonates can show heightened alpha-adrenergic response to dopamine and this can lead to ischemic vascular events. Dopamine infusion in neonates should be started at a low-dose via central line. PMID- 10513240 TI - Neuroleptic malignant syndrome. PMID- 10513241 TI - Not just another piece of paper. PMID- 10513242 TI - The use of controlled subatmospheric pressure to promote wound healing in preparation for split-thickness skin grafting in a fourth degree burn. AB - Subatmospheric pressure application to acute and chronic wounds has been shown to increase local wound blood flow, increase the rate of formation of granulation tissue, and enhance bacterial clearance. The mechanical forces applied to the wound enhance the rate of granulation tissue formation by the increase intracellular messengers regulating protein production and turnover. This method of wound care is particularly useful for larger wounds that could not be readily closed by local methods, but may also be useful for chronic wounds in debilitated patients who may not be ideal surgical candidates. Further basic scientific research is needed to discern the exact mechanisms of action of subatmospheric pressure, and more clinical experience is needed to establish guidelines for its application. Further clinical studies with larger subject populations in a randomized prospective study would lend support to the utility of this wound management protocol. PMID- 10513243 TI - [Magnetic resonance angiography in diagnosis of head and neck vessels diseases]. AB - Aiming to study diagnostic potential of MRA 286 patients have bean observed with pathology of the brain vessels. High efficiency of MRA has been confirmed: sensitivity and specificity with vascular malformations were 98% and 96%, with arterial aneurysm it was 97% and 82%, with stenosis of more than 50% it was 98% and 46%, with stenosis of less than 50% it was 96% and 84%, with thrombosis--100% and 100%. It was noted that TOF method is the most informative nature to estimate the arterial bed. We have managed to study with the PC method venous component of malformation, residual flow in huge thrombosed aneurysm, condition of the external carotid arteries and their branches. PMID- 10513244 TI - [Disseminated pulmonary tuberculosis: significance of high-resolution computerized tomography]. AB - Clinical and X-Ray studies were performed in 85 patients with disseminated pulmonary tuberculosis. All the patients underwent routine computerized tomography (CT) and high-resolution CT. According to the pathogenetic process, the authors identified hematogenic (n = 38), lymphogenic (n = 19), bronchogenic (n = 18) and mixed (n = 10) disseminations. High-resolution CT was found to have great advantages in detecting various types of tuberculous disseminations and in assessing the pattern of pulmonary abnormalities. Disseminated tuberculosis was revealed in 7 patients who had no pathological changes on routine lung X-ray films. The specific signs of hematogenic, lymphogenic disseminations and bronchgenic inoculations were identified in other forms of pulmonary tuberculosis. CT symptomatology is shown to be determined by the pathogenetic variant of its development and the stage of the process. Small focal changes in the lung were prevalent in patients with acute and subacute hematogenic forms of the disease. Infiltrates with decay cavities, thin-wall caverns, emphysema and bronchoectases were detected over the chronic course. Lymphogenic disseminations were characterized by the predominance of interstitial changes along with multiple minor foci. High-resolution CT had advantages in identifying decay cavities, signs of fibrosis and in evaluating mediastinal lymph nodes. CT data are of great significance for differential diagnosis of disseminated tuberculosis with lung metastases and diffuse interstitial diseases. PMID- 10513245 TI - [Diagnosis of rheumatoid arthritis of knee joints using magnetic resonance imaging]. AB - A combination of clinical, X-Ray and magnetic resonance tomographic studies for 129 knee joints was made in 85 patients with rheumatoid arthritis of knee joints. MRI symptoms of rheumatoid arthritis of knee joints, including fluid accumulation in the articular cavity, degeneration of the articular cartilage, meniscus, ligaments, proliferation of the synovial membrane, destructive changes in osseous epiphysis were defined. Comparative analysis of the X-Ray and MRI imaging findings has shown that MRI has advantages structures of joints in rheumatoid arthritis. PMID- 10513246 TI - [New aspects of echography using during delivery]. AB - The paper presents some aspects of the use of echography in obstetrics for the prediction and early diagnosis of labour abnormalities just before delivery and its first period. Prediction and early diagnosis of the nature of labour are made on the studies of changes in the myometrial thickness in the first period of delivery and myometrial echo structural features before and during delivery. Proceeding from the findings, labour abnormalities were corrected and the efficiency of drug therapy was evaluated. PMID- 10513247 TI - [Age-related changes of vertebral bone mineral density in Russian population]. AB - Vertebral trabecular bone mineral density (BMD) was measured by quantitative computed tomography (QCT) in 1061 subjects (610 females and 451 males aged from 7 to 91 and from 12 to 89, respectively) with known history of diseases or taking medicines affecting bone metabolism. Peak BMD values in our patients were observed at the age of 20-29 years with further gradual decrement in men and more steep in women. Negative relationship between BMD and age was r = -0.991 for men and r = -0.968 for women. Analyzing BMD changes by decades we observed the largest decrement in men after 60 (13.1% for 60-69 and 14.1% for 70-79 years of age) and in women after 50 (22.5% for 50-59, 22.1% for 60-69 and 20.8% for 70-79 years) which was most probably due to decline in sex hormones production that is known to significantly influence bone metabolism. This was confirmed by BMD values three-phase approximation in women showing the lowest rate of calcium loss by trabecular bone in reproductive period (1.9 mg/cm3/yr) and the highest in perimenopause (3.98 mg/cm3/yr). Annual calcium loss in postmenopause was 2.22 mg/cm3. PMID- 10513248 TI - [Radiation diagnosis of involutional osteopenia]. AB - 160 patients with involutive osteopaenia were investigated. The diagnostic possibilities of roentgenography, magnetic resonance imaging and computed tomography were evaluated. There was determined that roentgenography gives the possibility to determine the common localisation and the expressiveness of the pathologic process. MRI and CT give a valuable additional information on early stages of osteopaenia and it's complications. PMID- 10513249 TI - [First MRI detected metastasis to the fifth cervical vertebrae as a sign of thyroid tumor]. PMID- 10513251 TI - [Radiation diagnosis of endogenous allergic alveolitis]. PMID- 10513250 TI - [X-ray study in inflammatory changes of the jaw]. PMID- 10513252 TI - New hope for heart failure. PMID- 10513253 TI - The long and short of allergy shots. PMID- 10513254 TI - HDL on the rise. PMID- 10513255 TI - Watch out for eye infections. PMID- 10513256 TI - Microwaves help restore urine flow. PMID- 10513257 TI - Celebrex mix-up. PMID- 10513259 TI - How to walk well, not wounded. PMID- 10513258 TI - What really triggers your heartburn? PMID- 10513260 TI - Miscarriage: uncertainties remain. PMID- 10513261 TI - More could benefit from drug to forestall kidney failure. PMID- 10513262 TI - Weighing the options for hernia repair. PMID- 10513263 TI - Transcatheter Cardiovascular Therapeutics 11th annual symposium. Washington D.C., USA. September 22-26, 1996. Abstracts. PMID- 10513264 TI - European Helicobacter pylori Study Group XII International Workshop on Gastroduodenal Pathology and Helicobacter pylori. Helsinki, Finland, 2-4 September 1999. Abstracts. PMID- 10513265 TI - 17th European Congress of Pathology and XIX Spanish Congress of Pathology. Barcelona, Spain, September 18-23, 1999. Abstracts. PMID- 10513266 TI - Rough seas in US managed care. PMID- 10513267 TI - European ban on bovine growth hormones should continue: expert. PMID- 10513268 TI - Hepatitis outbreak in UK blamed on alternative doctor. PMID- 10513269 TI - A morally irrelevant distinction on euthanasia. PMID- 10513270 TI - Following the rules in marketing. PMID- 10513271 TI - Unconventional therapies and cancer. PMID- 10513272 TI - Driving for safety on our roads. PMID- 10513273 TI - Moon over Surrey. PMID- 10513274 TI - Hepatitis B virus infection among street youths in Montreal. AB - BACKGROUND: Street youths are at high risk for many health problems, including sexually transmitted diseases and bloodborne infections. The authors conducted a cross-sectional anonymous study from December 1995 to September 1996 involving street youths in Montreal to estimate the prevalence of risk behaviours for hepatitis B virus (HBV) infection and of markers of past and present HBV infection. METHODS: Participants were 437 youths aged 14 to 25 meeting specific criteria for itinerancy who were recruited in collaboration with the 20 major street youth agencies in Montreal. Sociodemographic and lifetime risk factor data were obtained during a structured interview, and a blood sample was taken to test for HBV markers (hepatitis B surface antigen and antibodies to the hepatitis B core antigen). Univariate analyses and multivariate logistic regressions were conducted. RESULTS: The mean age of the subjects was 19.5 years; 69.3% (303/437) were males. Many subjects had high-risk behaviours: 45.8% (200/437) had injected drugs, 24.5% (107/436) had engaged in prostitution, and 8.7% (38/437) reported having a sexual partner with a history of unspecified hepatitis. The prevalence rate for one or both HBV markers was 9.2% (40/434) (95% confidence interval [CI] 6.7%-12.3%). Multivariate logistic regression analysis showed that being over 18 years of age (adjusted odds ratio [OR] 4.5, 95% CI 1.8-11.7), having injected drugs (adjusted OR 3.5, 95% CI 1.5-8.3) and having had a sexual partner who had unspecified hepatitis (adjusted OR 3.2, 95% CI 1.3-7.5) were all associated with HBV infection. INTERPRETATION: Street youths are at high risk for HBV infection. Early and complete HBV vaccination among this vulnerable population is urgently needed. PMID- 10513275 TI - Hormone replacement therapy: a survey of Ontario physicians' prescribing practices. AB - BACKGROUND: Although much has been written about hormone replacement therapy (HRT), there are few clearcut recommendations on its use. The purpose of this study was to determine Ontario physicians' patterns of and reasons for prescribing HRT, their use of pretreatment investigations and their surveillance of HRT users, and to determine whether physicians' reported practice is consistent with existing recommendations. METHODS: A self-administered questionnaire was mailed to a nonproportional stratified sample of 327 Ontario physicians (23.9% gynecologists, 76.1% general practitioners/family physicians [GP/FPs]). Outcome measures were ranking of reasons for prescribing HRT, nature of preliminary testing, regimens prescribed, duration of HRT and frequency of follow-up. RESULTS: The response rate was 60.9% overall (70.9% of the gynecologists, 58.3% of the GP/FPs). Prevention of osteoporosis was reported by 97.4% as an important or very important reason for prescribing HRT; prevention of coronary artery disease was important or very important for 89.3%. When considering whether or not to prescribe HRT, 97.3% stated that breast cancer was an important or very important factor. When presented with hypothetical cases, 97.0% stated that they would prescribe combined estrogen-progestin for a symptomatic woman with an intact uterus; 13.6% stated that they would do so for a woman with no uterus. Most reported that they would prescribe HRT for 12 or more years (73.3%) and would follow up patients every 1 to 2 years (70.6%). INTERPRETATION: Despite controversy about HRT in the published literature, the Ontario physicians surveyed reported similar reasons and patterns of prescribing, pretreatment investigations, and surveillance of postmenopausal women using HRT. These results suggest that Ontario physicians' knowledge about HRT is consistent with recommendations in the published literature. PMID- 10513276 TI - Trends in the prevalence and treatment of hypertension in Halifax County from 1985 to 1995. AB - BACKGROUND: The objective of this study was to document changes in the prevalence and treatment of hypertension in Halifax County from 1985 to 1995 in an effort to observe, at the population level, the consequences of the availability of new antihypertensive medications. METHODS: The study population comprised a random sample of Halifax County residents, aged 25-64 years, who responded to the 1985 and 1995 surveys of the Halifax County MONICA Project and residents who responded to the Nova Scotia Health Survey conducted in 1995. Data from the two 1995 surveys were pooled. Information on hypertension awareness and use of medication were obtained through questionnaires, and blood pressure was measured according to a standard protocol, using phase I and V of Korotkoff sounds as respective markers for systolic and diastolic pressures. Uncontrolled hypertension was defined as a systolic pressure of 140 mm Hg or greater and a diastolic pressure of 90 mm Hg or greater. Changes in the prevalence of hypertension, prescribing trends and medication costs were examined, and the association between the type of antihypertensive treatment and characteristics of the respondents with self reported hypertension was investigated by multivariate logistic regression. RESULTS: Of the 917 people interviewed in 1985 and the 1338 in 1995, 274 (29.9%) and 356 (26.6%), respectively, reported a history of hypertension. When age was controlled for, the proportion of respondents reporting hypertension did not differ between survey years or between men and women. The proportion of treated respondents who had uncontrolled hypertension increased between 1985 and 1995, from 32.6% to 57.4% among men and from 38.0% to 42.6% among women. An increase was seen in the use of calcium-channel blockers (from 2.1% to 19.7%) and angiotensin-converting-enzyme inhibitors (from 5.2% to 25.4%); the proportion of patients receiving combination therapy or diuretics decreased (from 39.6% to 15.6% and from 31.3% to 17.2% respectively). These changes were associated with an increase in the average daily cost of medication from $0.48 to $0.85 per patient. INTERPRETATION: The shift to new antihypertensive drugs was not associated with improved blood pressure control, but it was associated with an increase in average medication costs per patient. Uncontrolled hypertension remains a public health problem. PMID- 10513277 TI - Attitudes of Canadian women toward birthing centres and midwife care for childbirth. PMID- 10513278 TI - The treatment of hypertension in Canada: are we making progress? PMID- 10513280 TI - Effectiveness of the Quick Medical Reference as a diagnostic tool. AB - A number of computer-based systems with diagnostic capabilities have been developed for internal medicine. Quick Medical Reference (QMR) is one such program. The authors describe key features of QMR and report on their study of its effectiveness as a diagnostic tool. They investigated how frequently the correct diagnosis would appear among the 5 highest ranked diagnoses generated by QMR. The charts of 1144 consecutive patients admitted to a teaching unit were retrospectively screened. Eligible cases included those referred for investigation of an undiagnosed illness with an objectively proven final diagnosis (n = 154). Two physicians familiar with, but not experts in, the use of QMR entered clinical information abstracted from the patients' charts into the program. Physician A obtained the correct diagnosis in 62 (40%) of the 154 cases, and physician B was successful in 56 (36%) of the cases. The authors use study cases to illustrate QMR's strengths and weaknesses. PMID- 10513281 TI - An interdisciplinary approach to a day-long palliative care course for undergraduate students. AB - Although it is desirable that students in the health sciences be educated together to prepare them for interdisciplinary practice, many educational programs remain discipline specific. An undergraduate course in palliative care, originally designed for medical students at McMaster University, Hamilton, Ont., was expanded in 1993 to include students from various health sciences programs in the region. The course introduces students to the components of palliative care and its interdisciplinary nature in a problem-based way and directs students to additional educational resources. The authors describe the planning, content and evaluation of the course material. The observed decline in attendance by medical students, which coincided with the introduction of the interdisciplinary format, warrants further investigation. Future directions of the course are discussed. PMID- 10513279 TI - Tuberculosis: 10. Prevention. PMID- 10513282 TI - It's wise to immunize, regardless of what the Web says. PMID- 10513283 TI - Physicians may have to "sell" benefits of immunization to sceptical parents. PMID- 10513284 TI - Health Canada leery as US "nutraceutical" movement prepares to move north. PMID- 10513285 TI - Swissair disaster taught medical examiners a lesson in logistical challenges. PMID- 10513286 TI - Breast cancer screening, diagnosis, and treatment. AB - The incidence of breast cancer in US women remains disturbingly high, and unfortunately primary care physicians still frequently encounter patients in whom the disease is suspected or, even worse, confirmed. Fortunately, however, the body of knowledge surrounding the disease has grown dramatically during the past decade, and major advances have been made in the understanding of breast cancer risk, prevention, diagnosis, and treatment. Controversies persist, particularly those concerning the screening of younger women, but consensus now exists regarding many clinical issues relevant to primary care practice. Although multidisciplinary subspecialty expertise must be made available to all women with known or suspected breast cancer, the primary care physician has an important role to play when dealing with patients with this condition. The following article focuses on what primary care practitioners need to know to expertly contribute to the diagnosis, counseling, and initial treatment of women with this disease. PMID- 10513287 TI - Antiserum generated by DNA vaccine binds to hepatitis E virus (HEV) as determined by PCR and immune electron microscopy (IEM): application for HEV detection by affinity-capture RT-PCR. AB - Previously, we have described that injection of an expression vector containing hepatitis E virus (HEV) open reading frame 2 (HEV-ORF-2) generated a strong antibody response in mice. To characterize the reaction of this antiserum with native HEV and to evaluate its potential diagnostic application, we tested the antiserum's ability to bind HEV using immune electron microscope (IEM) and affinity-capture reverse transcription polymerase chain reaction (RT-PCR) amplification. Antiserum to ORF-2 aggregated HEV virions as seen by electron microscopy, providing direct evidence that ORF-2 encodes a structural protein. Antiserum also captured HEV for RT-PCR amplification. This antiserum bound HEV from diverse origins (Asia, Africa, Mexico) at virus concentrations found in patient fecal specimens and bile from inoculated non-human primates. The specificity of the affinity binding was demonstrated when pre-immune sera or sera collected from mice injected with control DNA vector (lacking the HEV ORF-2 gene) failed to bind HEV for RT-PCR amplification and IEM. Specific RT-PCR amplification was confirmed by restriction enzyme digestion of PCR products. The sensitivity of the binding was evaluated by RT-PCR amplification of serially diluted bile containing a genetically divergent HEV, Mexico'86. HEV was detected in a 10(-8) dilution of this bile. This is the first report that antibodies elicited by a DNA vaccine recognize native HEV. Our results indicate that ORF-2 encodes a structural protein and that antiserum to this protein enables simple, sensitive, and specific HEV detection by affinity-capture RT-PCR. PMID- 10513288 TI - Localization of foot-and-mouth disease virus RNA by in situ hybridization within bovine tissues. AB - Foot-and-mouth disease is a highly contagious disease of cloven hooved animals. In cattle, both acute and long-term persistent infections occur. Foot-and-mouth disease virus (FMDV), a picornavirus, has been shown, using virus isolation procedures, to replicate in the pharynx and soft palate of cattle. In this study, in situ hybridization has been used to detect FMDV RNA within the cells of tissues removed from infected bovines. A digoxigenin-labelled anti-sense RNA probe was prepared corresponding to a region of the FMDV genome encoding part of the RNA-dependent RNA polymerase (3D). The efficacy and specificity of this probe for in situ hybridisation was determined using virus-infected cells in tissue culture. Strong cytoplasmic staining was only detected in FMDV-infected cells. Various tissue samples were collected from FMDV-infected cattle between 5 and 17 days post-infection. Viral RNA was detected by in situ hybridisation within cells of the soft palate, tonsil and pharynx up to 17 days post-infection. This technique is useful for the study of FMDV localization in cattle both during and after the acute clinical phase of disease and may assist in identifying specific sites of virus persistence. PMID- 10513289 TI - Influence of different cellular transcription factors on the regulation of varicella-zoster virus glycoproteins E (gE) and I (gI) UTR's activity. AB - Varicella-zoster virus (VZV) glycoproteins E (ORF 68) and I (ORF 67) are members of late genes. They belong to the major components of the virion envelope and can be found on the host cell surface as well. To get further insights in the regulation of gE and gI expression, which are known to be activated by IE4 and IE62, we analysed the intergenic regions of ORF 66/67 and ORF 67/68, containing the putative promoters of gI and gE. We have mapped the transcriptional start site of gE and have identified an extensive set of eucaryotic cis-elements: typical TATA- and CAAT-motifs and further regulatory sequences to facilitate interaction with eucaryotic transcription factors. Reporter constructs have been made using the intergenic regions of ORF 66/67 and ORF 67/68 as promoter elements. In cis-trans interaction studies, an influence on the regulation of transcription and reporter gene expression of overexpressed transfactors, LAP/LIP, Sp1, YY1 and NF-E2 has become measurable. In addition, protein-DNA binding assays using both gE- and gI-intergenic regions and cellular extracts from different VZV-permissive cells have suggested a binding of a 32 and 18 kD protein. In conclusion, these data indicate an involvement of common cellular transcription factors in the regulation of VZV late gene expression. PMID- 10513291 TI - XXI Congress of the European Society of Cardiology. Barcelona, Spain, August 28 September 1, 1999. Abstracts. PMID- 10513292 TI - 7th Conference on Cell Biology. Krakow, Poland, 9-11 September 1999. Abstracts. PMID- 10513290 TI - Genotypic and antigenic characterization of hemagglutinin proteins of African measles virus isolates. AB - A comprehensive phylogenetic study based on the hemagglutinin (H) protein of all known African measles virus (MV) isolates is presented. The study includes 64 new H gene sequences from Ghana. Nigeria and South Africa as well as viruses from Zambia and The Gambia for which only incomplete sequencing data were available and that have previously not been genotyped. The results provide further support to the tentative assignment of the Nigerian and Ghanaian viruses to a new genotype B3 within clade B. A distinct geographic distribution pattern emerged with clade B viruses circulating exclusively in African countries north of the equator. All MV strains from southern Africa grouped in clades A and D with the majority of viruses belonging to genotype D4. The viruses considerably differed by their sensitivity to neutralization by monoclonal antibodies (mAb), but three selected antibodies were sufficient to distinguish between African MVs representing four different genotypes. PMID- 10513293 TI - Society for the Study of Inborn Errors of Metabolism (SSIEM) 37th annual symposium. Genova, 7-10 September 1999. Abstracts. PMID- 10513294 TI - International Society for Interferon and Cytokine Research (ISICR) annual meeting. Paris, France, September 5-9, 1999. Abstracts. PMID- 10513295 TI - European best practice guidelines for the management of anaemia in patients with chronic renal failure. Working Party for European Best Practice Guidelines for the Management of Anaemia in Patients with Chronic Renal Failure. PMID- 10513296 TI - 12th International Workshop on Multiple Pregnancy: from Curiosity to Epidemic. Assisi, Italy, June 24-27, 1999. Abstracts. PMID- 10513297 TI - One-third and two-thirds of all "naturally" fertilized human eggs never develop into anything most people would recognize as human. PMID- 10513298 TI - First choice. PMID- 10513299 TI - Abortion and assent. PMID- 10513300 TI - Thomson's violinist and conjoined twins. PMID- 10513301 TI - Abortion, relationship, and property in labor: a clinical case study. PMID- 10513302 TI - After feticide: coping with late-term abortion in Israel, western Europe, and the United States. PMID- 10513303 TI - The problem of coerced abortion in China and related ethical issues. PMID- 10513304 TI - The horns of the dilemma are sharp. PMID- 10513305 TI - The need for more physicians trained in abortion: raising future physicians' awareness. PMID- 10513306 TI - Abortion: three rival versions of suffering. PMID- 10513307 TI - CQ sources/bibliography. PMID- 10513308 TI - Protecting communities in research: philosophical and pragmatic challenges. PMID- 10513309 TI - 60 minutes sets the record straight. PMID- 10513310 TI - Navigating turbulent and uncharted waters. PMID- 10513311 TI - Mediating disputes about medical futility. PMID- 10513312 TI - Providing comfort or prolonging death for a baby with "dead gut syndrome"? PMID- 10513313 TI - The ethical and religious challenges of reproductive technology. PMID- 10513314 TI - Does the "sanctity of human life" doctrine sanctify humanness, or life? PMID- 10513315 TI - Health care without harm: cleaning up healthcare's act. An interview with Michael Lerner. Interview by Steve Heilig. PMID- 10513316 TI - Kate Christensen speaks with Pat Matheny, a recipient of lethal medication under Oregon's Death with Dignity Act. Interview by Kate Christensen. PMID- 10513317 TI - EEG and epilepsy in the elderly compared to a younger group. AB - EEG in epilepsy in the elderly (onset > 60 yrs) was investigated (161 patients- 302 EEGs) and was compared to a group of younger patients (onset 20-59 yrs) matched for sex ratio (also 161 patients--296 EEGs). In the elderly, the frequency of the background rhythm was decreased, as was the rhythmicity and amplitude. Complex partial attacks were twice as often seen in the older group, but generalized tonic-clonic seizures were more than three times as often in the younger group. The major etiology in the elderly was cerebrovascular disease, but brain tumors were found in nearly one quarter, while head injury, drug abuse, and AVM were more often seen in the younger patients. The first EEG was positive for epileptiform activity in 83%, later at 92-93% in both groups, emphasizing adequate sleep records. Although temporal lobe abnormalities were most often seen, frontal lobe discharges and slow waves were significantly more often noted in the elderly. As an example of the prominence of epileptiform activity in some elderly, PLEDs and very many discharges were seen mainly in the older group, which less often showed only rare discharges. PMID- 10513318 TI - Creutzfeldt-Jakob disease diagnosable by EEG and cerebrospinal fluid analysis without brain biopsy: a case report. AB - This case illustrates a classic example of CJD in its clinical presentation and course and the EEG. It also shows dramatically the utility of a newly developed protein assay in the diagnosis of this disease. This assay has the potential of eliminating the need for brain biopsy in most cases, thus providing a safer diagnostic method for both staff and patients. In addition, the case points out that anatomical structural studies such as CT and MRI do not replace the utility of EEG in the comprehensive evaluation of rapid onset dementia, but rather complement the usefulness of EEG. PMID- 10513319 TI - Clinical aspects of the "third rhythm" of the temporal lobe. AB - We studied clinical aspects of the "third rhythm," which was first described by Niedermeyer as alpha-like activity of the temporal lobe. By scalp EEG, temporal alpha-like activity was recorded in 15 (0.30%) of 4929 patients over 20 years of age. The temporal alpha-like activity was observed in 5 patients who had clinical and brain imaging findings indicating the presence of a cerebrovascular disorder. The alpha-like rhythm of these patients was left-sided, and wicket spikes appeared in the same region as the alpha-like rhythms in 4 of the 5 patients. In 8 of the 15, the temporal alpha-like rhythm was recorded over the defective bone or replacement bone after intracranial surgery. The alpha-like rhythm of these patients was similar to breach rhythm in the temporal region. The remaining 2 patients had not undergone intracranial surgery nor did they show symptoms of cerebrovascular disorders. The temporal alpha-like rhythms in one of these 2 might be a physiological third rhythm detected by scalp EEG through congenital bone thinning. Our observation supports the existence of intrinsic activity of the temporal lobe (the third rhythm). The third rhythm can be recorded by routine scalp EEG in some clinical conditions, and it is important to recognize this rhythm when one encounters temporal rhythmic activity independent of alpha rhythm of the occipital lobe. PMID- 10513320 TI - Detecting symmetric patterns in EEG data: a new method of analysis. AB - Theoretical models of higher cognitive function predict that cortical activity will exhibit families of spatial-temporal patterns of activity whose individual members are related to each other by specific symmetry transformations. In the trion model, it is suggested that these inherent symmetries play a vital role in how we think and reason. We have developed a method of analysis (SYMMETRIC analysis), which detects families of patterns in EEG data, and characterizes the symmetry relationships between members of those pattern families. Using this analysis, significant symmetry families have been found in EEG and single unit spike train data. If symmetry is a crucial aspect of brain function, it is possible that different pathologies are associated with specific types of symmetry relationships in brain activity that could be detected in EEG data by a SYMMETRIC analysis. PMID- 10513321 TI - Music enhances spatial-temporal reasoning: towards a neurophysiological basis using EEG. AB - Motivated by predictions from the structured trion model of the cortex, based on Mountcastle's columnar organizational principle, behavioral experiments have demonstrated a causal short-term enhancement of spatial-temporal reasoning in college students following listening to a Mozart Sonata (K.448) but not in control conditions. An EEG coherence study reported presence of right frontal and left temporoparietal activity induced by listening to the Mozart Sonata, which carried over into the spatial-temporal tasks in three of the seven subjects. In this paper, we present further predictions from the trion model and discuss how the new SYMMETRIC analysis method can be used in EEG recordings to help determine the neurophysiological basis of specific music enhancing spatial-temporal reasoning. We conclude with potential clinical applications of major significance. PMID- 10513322 TI - Consciousness as a definition of death: its appeal and complexity. AB - A new formulation of death proposed in this study is based on the basic physiopathological mechanisms of consciousness generation in human beings. Two physiological components control conscious behavior: arousal and awareness (content of consciousness). We cannot simply differentiate and locate arousal as a function of the ascending reticular activating system and awareness as a function of the cerebral cortex. Substantial interconnections among the brainstem, subcortical structures and the neocortex, are essential for subserving and integrating both components of human consciousness. Therefore, consciousness does not bear a simple one-to-one relationship with higher or lower brain structures, because the physical substratum for consciousness is based on anatomy and physiology throughout the brain. This new account of human death is based on the irreversible loss of consciousness because it provides the key human attributes and the highest level of control in the hierarchy of integrating functions within the organism. The notion of consciousness as the ultimate integrative function is more consistent with the biologically-based systems than the more philosophically-based notions of personhood. PMID- 10513323 TI - Quantitative EEG findings associated with chronic stimulant and cannabis abuse and ADHD in an adult male substance use disorder population. AB - QEEG was studied in a population of chronic male PSUD/ADHD (psychoactive substance use disorder/attention deficit hyperactivity disorder) subjects vs. a matched sample of non-ADHD subjects with PSUD. Our first interest in conducting this study was to determine if the Thatcher University of Maryland database and complex demodulation method could replicate the specific QEEG findings reported for cocaine and cannabis using the John-NYU database and Fourier Transform method. The effects of cannabis and stimulants were also studied both separately and together to see if there were interactions and to see if the QEEG changes associated with chronic stimulant dependence were predicted by childhood ADHD status. Eyes-closed QEEGs were obtained and two independent artifacted 60 second samples were compared for reliability. The Thatcher database was used to analyze QEEG data from 56 subjects with mixed substance use disorder. Results showed that the Thatcher database replicates the John database for chronic stimulant dependence findings. Because of confounding variables of alcohol and polysubstance abuse, the findings related to cannabis and stimulant interaction were difficult to assess. Cannabis and stimulant dependence together produced more QEEG changes than either alone. More right temporal abnormalities were observed with stimulant dependence. In the absence of stimulant use, the QEEG effects of cannabis were relatively small; however, sample selection and methods used precluded comparison to previous studies. The persistent QEEG abnormalities associated with chronic stimulant dependence were independent of ADHD status in this sample using the methods of this study. Further research is needed to clarify the relationship of stimulant dependence with QEEG changes and ADHD status, and to clarify the interactions of chronic stimulant and cannabis abuse on QEEG. PMID- 10513324 TI - Quantitative EEG during early recovery from hypoxic-ischemic injury in immature piglets: burst occurrence and duration. AB - This study examined the course of EEG recovery in an animal model of hypoxic ischemic injury. The model used periods of hypoxia, room air and asphyxia to induce cardiac arrest. One-week-old piglets (n = 16) were exposed to a period of hypoxia, room air and complete asphyxia for 7 minutes. After cardiac arrest and resuscitation, two EEG features were evaluated as prognostic indicators of behavioral outcome as assessed by a neuroscore at 24 hours after insult. A prominent EEG feature was the number and duration of bursts evident during recovery. Episodes of bursting were detected through the thresholds on sustained periods of elevated power. After the animal was resuscitated, the EEG was monitored continuously for 4 hours. To assess outcome in the recovering animal, a behavioral testing scale was used to test the animal's neurological capabilities. Trends of EEG burst counts were measured through thresholds on sustained power changes. Bursts are energy transients in the EEG record. High degrees of bursting were characteristic of animals having good neurological condition whereas piglets having low burst counts had poor 24 hr neuroscores. At 100 min the average burst rate of the good neuroscore outcome group was more than 8 per min and was significantly different from the poor outcome group's level of 2.7 (p < or = 0.05). When these counts were weighted by their total duration, differences between groups increased (p < or = 0.02). This study showed that the QEEG measure of burst counts and duration together provided a strong prognostic indication of the 24 hour outcome after asphyxic injury in a neonatal animal model. The critical determinant of the bursting character was the time when bursting occurred. Bursting occurring early in recovery was a good gauge of outcome. We conclude that quantitative EEG analysis and interpretation can be an important tool for the outcome determination during recovery from cerebral injury states. PMID- 10513325 TI - [The antiparkinson activity of a new adamantane derivative]. AB - The activity of the new original compound--hydrochloride N-2-(adamantyl) hexamethylenimine (code A-7) synthesized at the Institute of Pharmacology, Russian Academy of Medical Sciences, was studied on models of akinetic-rigid and tremor manifestations of the parkinsonian syndrome. A-7 completely relieves the akinetic-rigid manifestations of the syndrome and surpasses mydantan and L-dopa in antagonism with haloperidol and triftazine. Just as L-dopa, cyclodol, and mydantan, A-7 alleviates reserpine-induced oligokinesis and restores body temperature which is lowered by reserpine. A-7 possesses evident antitremor activity which advantageously distinguishes it from mydantan which failed to demonstrate antagonism with arecoline. A-7 is promising for further study as a potential agent for the treatment of parkinsonism. PMID- 10513326 TI - [The effect of selective m-cholinopositive substances on the antiparkinson activity of atropine]. AB - The effect of the selectivity of cholinopositive compounds on atropin antiparkinsonian activity was compared in experiments on rats. The selectivity of the effect of two new synthesized cholinopositive compounds of the phenylcarbamate group was evaluated in preliminary experiments. It was determined by means of these compounds that stimulation of m1-cholinoceptors reduces while stimulation of m2-cholinoceptors has no effect on the ability of atropin to prevent haloperidol-induced catalepsy. PMID- 10513327 TI - [The effect of bemitil on conditioned-reflex memory in normal rats and under stress exposures]. AB - The authors studied the effect of the actoprotector bemitil on the conditional reflex memory of rats and its functional disorder by disturbance of the cause effect (abatement of the avoidance reaction) or space relations. Intraperitoneal injections of 1.8 mg/kg of bemitil were given daily 30 min before the experiment. The training of animals improved authentically from experiment to experiment. Thus, the drug, possesses the nootropic properties. Exposure to stressogenic factors of various depth demonstrated the stress-protective effect of bemitil. PMID- 10513328 TI - [The effects of L-arginine in chronic pain syndromes]. AB - The effect of L-arginine in long-term parenteral administration was studied on a model of neuropathic pain syndrome and adjuvant arthritis. L-arginine produced a preventive and therapeutic effect in the neuropathic pain syndrome. It weakened the development of adjuvant arthritis for the period of its administration. The factor underlying the effect of L-arginine is its double action as a precursor of nitrous oxide and kiotorphin, an endogenous antinociceptive dipeptide. PMID- 10513329 TI - [The effect of tropoxin on the neurogenic and vasogenic inflammation of the dura mater vessels]. AB - Subchronic administration of tropoxin (in doses of 7.5 and 10 mg/kg) caused dose dependent blocking of 131I-albumin plasma transudation from the dura mater vessels, induced by electrical stimulation of the trigeminal ganglion and intravenous infusion of the agonist of 5-HT2B/2C receptors metachlorophenylpiperazine. The antimigraine agent metisergid produced a similar effect. A single injection of metisergid and tropoxin did not block albumin transudation. A 3 mg/kg dose of mianserin prevented the blocking effect of tropoxin and metisergid on plasma exudation into the dura mater. It is suggested that the mechanism of the tropoxin antimigraine effect is realized through the presynaptic 5-HT1 receptors of afferent endings of the trigeminal nerve and the postsynaptic 5-HT2B/2C receptors of the dura mater vessels. PMID- 10513330 TI - [The action of a number of cardiac glycosides on an isolated system of myocardial contractile proteins in heart failure due to toxic-allergic myocarditis. The molecular mechanism]. AB - Experiments conducted on an isolated contractile apparatus, myocardial fibers (MF) in cardiac insufficiency (CI) caused by toxico-allergic myocarditis of 10 days duration (TAM10dd) showed that cardiac glycosides (CG), beta-acetyldigoxin (beta AD), beta-methyldigoxin, and strophanthin K (SK) increase the capacity of the actomyosin ensemble (AME) for generation of force, hydrolization, and economic use of the free energy of ATP hydrolysis. The mechanism of the effect of these CG in the phase of contraction differs from that of their effect on the AE of a normal myocardium. For instance, in severe CI induced by TAM10dd beta AD, in distinction from its action on the AME of a normal myocardium, can increase contractility economy, particularly in the phase of highest energy capacity, the phase of force generation, and exceed the level encountered in normal conditions, it also increases significantly the rate and reduces the time of MF relaxation as in the case of MF of a normal heart. PMID- 10513332 TI - [Isothiorbamin possesses stress-protective action]. AB - It was shown on a model of long-term stress induced by chronic electrostimulation of the ventromedial hypothalamus of rabbits that administration of isothiorbamin in the active periods of the experiment prevents hyperactivation of lipid peroxidation in the blood and myocardium by maintaining the activity of antioxidant enzymes. Isothiorbamin also causes an antiatherogenic effect on the blood lipid spectrum, raises the efficacy of the work of the heart and decreases the stressogenic ischemic disorders of the myocardium. PMID- 10513331 TI - [The anti-arrhythmia activity of new dicyclohexylamide derivatives of N substituted alpha-aminocarboxylic acids]. AB - Experiments on arrhythmia models showed a high antiarrhythmic activity of new derivatives of dicyclohexylamides of N-replaced alpha-aminocarbonic acids. The new compounds surpassed in intensity and duration of the antiarrhythmic effect the standard agents with classes I and III antiarrhythmic activity. In doing so they raise myocardial electrical stability and prevent sudden development of ventricular fibrillation. According to the mechanism of the antiarrhythmic activity, the new compounds may be related to antiarrhythmic agents possessing the properties of classes I and III. PMID- 10513333 TI - [The effect of perftoran on the morphofunctional status of the digestive system organs in experimental duodenal ulcer]. AB - The authors studied the effect of perftoran (PF) on the morpho-functional parameters of alimentary system organs (ASO) (liver, pancreas, stomach, duodenum, and small intestine) in rats with experimental chronic acetate duodenal ulcer. Inflammatory-infiltrative and dystrophic changes were found in the ASO tissues which led to activation of the enzymes superoxide dismutase and catalase in their homogenates. PF pharmacotherapy reduces the inflammatory-infiltrative changes in the ASO tissues and normalizes the enzyme systems of antioxidant protection. PMID- 10513334 TI - [The mechanisms of the anti-ulcer action of plant drug agents]. AB - The mechanisms of the gastroprotective effect of well-known drugs (plantaglucid, befungin, plantain juice), extractions of birch bark (Betula pendula Roth,) and burdock seeds (Arctium tomentosum Mill.) on various links of gastric ulcer pathogenesis were compared. Phytopreparations were found to have a variety of effects on gastric secretion in rats. Differences in their action on the smooth muscles of the stomach and intestine were detected. PMID- 10513335 TI - [The effect of bromantane on the erythro- and leukocytic profile of the peripheral blood in rats]. AB - The specific effect of bromantan on blood tissue is demonstrated in rats. In chronic injection in a dose of 30 mg/kg bromantan raised the level of hemoglobin and leukocytes. In doses of 150 and 600 mg/kg it increased at first (3 months) and then reduced the level of erythrocytes, hemoglobin, and leukocytes. Reversible poikilocytosis, granulocytosis, and agranulocytosis were encountered. Hyperchromatosis and hypertrophy of the hepatocytes were found in the tissues of the liver and hemosiderosis was discovered in the spleen. It is suggested that the blood tissue is a "target" in the toxic effect of bromantan. PMID- 10513336 TI - [The immunoregulatory properties of recombinant human tumor necrosis factor-beta in mice opposite-reacting to antigen]. AB - The effect of 10(3)-10(5) E/20 g doses of the recombinant factor of human beta tumor necrosis (rFNT-beta) on formation of the immune response and macrophage functional activity was studied in CBA and C57Bl/6 mice that differ in genetically determined level of the immune response to an antigen (sheep erythrocytes). The rFNT-beta was found to cause a modulating effect on the cell and humoral links of the immune response. The effect of the agent depended on the dose and the genotype of the experimental animals. It is suggested that the interlinear differences in the intensity of the humoral immune response in rFNT beta administration may be connected with the different sensitivity to the agent of the peritoneal macrophages of mice of the used lines. PMID- 10513337 TI - [The antimutagenic activity of aspartame]. AB - The method of chromosome aberration count in the bone marrow cells of C57B1/6 mice was used to study the influence of aspartame on the cytogenetic effects of dioxydin and cyclophosphan. Aspartame (0.4-40 mg/kg) was found to possess antimutagenic properties in relation to the listed mutagens. The discovered antimutagenic activity of aspartame was manifested more when it was injected for 5 days before the administration of a mutagen, whereas in joint administration of aspartame with the mutagens, the substitute for sugar did not change the clastogenic effect of dioxydin and cyclophosphan. PMID- 10513338 TI - [The dehydration action of ketamine on tumorous and normal glandular breast tissues in vitro]. AB - The authors studied the effect of the ketamine anesthetic on hydration of tumoral and normal glandular cells and on the binding of labelled ouabain with the tissues in order to reveal changes in the number of active Na-K-ATPase molecules on the cell membrane. Hydration of tumoral and normal cells diminished in one hour incubation in a solution of narcotic and subnarcotic doses of ketamine based on Tyrode's solution. The binding of ouabain by 3H also sharply reduced in a concentration of 10(-8) M in 30 min incubation, after incubation of pieces of tissues in ketamine, which was also evidence of reduction of the cell volume. This proves that the content of water in the tumoral cells increases and that ketamine in subnarcotic doses exerts a selective effect on tumoral cells. PMID- 10513340 TI - [The effect of altan on the functional activity of the liver mitochondria and microsomes from rats with toxic hepatitis]. AB - In in vitro experiments althan had no effect on the respiration of intact mitochondria in state 4 according to Chance and produced a high antioxidant effect in fermentative and ascorbate-dependent lipid peroxidation in intact microsomes isolated from the rat liver. In ethanol-induced toxic hepatitis althan restored the functional activity of mitochondria to the level of that in intact animals and increased microsome hydroxylase activity in CCl4-hepatitis. PMID- 10513339 TI - [The role of the blockade of separate subtypes of muscarinic receptors in realizing the antidotal effect of m-cholinolytics in dimethyl dichlorovinyl phosphate poisoning]. AB - On comparison of the values of receptor selectivity of a series of m cholinoblockers in vitro with those of the selectivity of their effect in in vivo experiments, pharmacological tests characterizing the interaction of ligands with m1, m2, and m3-subtypes of muscarine receptors were determined. Analysis of the protective effect of m-cholinoblockers in poisoning with organophosphorus compounds (OPC) depending on their activity in the determined tests showed that blocking of the m1-cholinoceptors is responsible for the antidotal effect of the antagonists, whereas block ing of m2-cholinoceptors prevents it. It is suggested that the negative effect of m2-cholinoceptor blocking on the protective effect of the drugs in OPC poisoning is mediate d by increased excretion of the mediator into the synaptic cleft as a result of interaction of the ligands with the presynaptic autochol inoceptors. PMID- 10513341 TI - [The effect of tactivin and levamisole on the development of toxic brain edema swelling]. AB - Edema-swelling of the brain (ESB) is among the urgent problems of modern medicine. The purpose of the research was the study of the effect of immunotropic agents on formation of the pathologic process. It was demonstrated on a model of toxic ESB that taktivin and levamisole possess antiedemic activity. The effect is dose-dependent and is determined by the structural characteristics of the drugs under study. Unlike taktivin, large doses of levamisole demonstrate synergism with the effect of the edematous factor. PMID- 10513342 TI - [The effect of polikatan on the ototoxic action of kanamycin]. AB - Experiments on guinea pigs demonstrated that preliminary injection of polycatan (standardized magnesium solution containing the mineral bischofite) into the parotid region by means of electrophoresis reduces the ototoxic effect of the aminoglycoside antibiotic kanamycin. Polycatan prevents kanamycin-induced degenerative changes of the hair cells found in the labyrinth of the internal ear and improves the local blood flow. PMID- 10513343 TI - [GABA-A receptor subunits and their reactions to neuropharmacological substances]. AB - The structural-functional foundations of the heterogeneity of GABA(A)-receptors are discussed. The heterogeneity of the receptors was revealed by clone-formation methods. The existence of 5 classes of subunits (alpha, beta, gamma, delta, rho) was proved, each of them possessing isoforms (alpha 1-6, beta 1-4, gamma 1-4, delta, rho 1-2). Studies with expression of receptors showed the efficacy of neuroactive drugs of various groups (agonists, antagonists, reversible agonists of benzodiazepine receptors, barbiturates) to be dependent on the subunit composition of the complexes. Because of this, study of the properties of various recombinant receptors is very important for the advancement of neurobiology, neuropathology, neurochemistry, toxicology, and pharmacology. PMID- 10513344 TI - [The molecular mechanisms of the action of antiprogestins]. PMID- 10513345 TI - [The outlook for the use of the nonpeptide angiotensin II-receptor antagonist losartan in treating heart failure]. PMID- 10513346 TI - Antiinflammatory effects of oxatomide. AB - There is increasing evidence that histamine may have wider proinflammatory and immunomodulatory activities than previously reported. It may influence several functions of lymphocytes, monocytes, basophils and macrophages, modulating the release of inflammatory mediators and cytokines. These observations have aroused interest in the pharmacology and clinical applications of histamine H1 receptor antagonists and have led to the identification of novel antiinflammatory properties for this class of drugs. Oxatomide, initially characterized as an H1 antagonist, inhibits the secretion of several mediators of inflammation from human basophils and mast cells. In vitro oxatomide inhibits the release of both preformed (histamine and tryptase) and de novo synthesized mediators (leukotriene C4 and prostaglandin D2). The inhibitory effect is not restricted to basophils and mast cells but is also evident on other inflammatory cells such as the neutrophils. In this cell, oxatomide inhibits arachidonic acid mobilization, and leukotriene B4 and platelet-activating factor synthesis, presumably by reducing the activity of cytosolic phospholipase A2. These observations extend the pharmacological activities of oxatomide beyond H1 receptor antagonism and suggest that this drug influences a variety of biochemical events in human inflammatory cells. These antiinflammatory activities help to explain its beneficial effect in various allergic and inflammatory disorders, including urticaria, allergic rhinitis and bronchial asthma. PMID- 10513347 TI - Allergy to betalactams: relationship between chemical structure and antigenicity of molecules. AB - As betalactams (penicillins, cephalosporins, carbapenems, monobactams, betalactamase inhibitors) are the most widely used antibiotic drugs worldwide, allergy to betalactams is a frequent problem encountered in clinical practice, concerning from 0.7-8% of treated patients. It is now clear that the antigenicity of these drugs is strictly related to the chemical structure of the various molecules and of their constituent parts. In this paper we first describe the chemical structure of betalactams and then try to establish a correlation between this structure and their antigenicity. PMID- 10513348 TI - Atopy parameters in asthmatic infants. AB - We evaluated various atopy parameters in 44 asthmatic infants aged 1-3 years: nonspecific parameters (total IgE and eosinophilia), and specific parameters relative to 21 allergens (specific IgE and skin tests), together with tests of leukotriene release by blood leukocytes (cellular allergen stimulation test; CAST) in the presence of a mixture of 21 allergens. Thus far 17 infants have displayed no sign of atopy, but 27 met at least one criterion. Eight met nonspecific criteria, and the others single or multiple criteria. Of the 21 allergens, 20 gave rise to at least one sensitization in this population. The specific IgE was more frequently positive than the skin tests. Dissociations between the two types of specific tests were practically systematic. Different phenotypes based on the chosen parameters were individualized, demonstrating heterogeneity in the expression of atopy in these young infants. The polyvalent CAST was positive only when other criteria of atopy were also positive (specific IgE and/or skin tests), and the range of intensity of the responses obtained supports reactivity rather than sensitization. PMID- 10513349 TI - Airborne pollens and prevalence of pollenosis in western Liguria: a 10-year study. AB - During the last 15 years aerobiology has become a relevant branch of allergy, making possible the partial clarification of the relationships between clinical diseases and environment. We performed a 10-year survey of pollen counts and pollen sensitization in a confined area on the western Ligurian coast of Italy in order to evaluate possible changes in aerobiological pattern and to correlate them with the prevalence of sensitization. Pollen counts for the area surrounding Bordighera in the period from 1988-1997 were analyzed; the occurrence of skin sensitization in outpatients were also studied during the same period. We considered the following allergens: Parietaria, grasses, Compositae, Cupressaceae, olive and birch. We also examined the possible differences between patients living on the seaside and those living inland. Over the 10-year period a significant increase in the pollen counts was seen for birch and Compositae (p = 0.001); this was accompanied by a parallel significant increase in the rate of sensitization (p = 0.004 and p = 0.01, respectively). Conversely, an increase in sensitization to Cupressaceae (p = 0.001) and olive (p = 0.03) was also seen, although no change in the pollen counts was detectable. Finally, the prevalence of sensitization to Cupressaceae and Compositae was higher in the patients living in the coastal region than those residing inland. These data suggest that a positive correlation between the pollen counts and the rate of sensitization exists for certain pollens. Nevertheless, for other species such a correlation was not apparent, and additional environmental factors maybe involved in the increased prevalence of sensitization. PMID- 10513350 TI - Proteinase and gelatinolytic activities of house dust mite and cockroach extracts. AB - The presence of gelatinolytic activity in dust mite and Periplaneta americana allergenic crude extracts were studied. The former presented major activity in a broad band between 45 and 66 kDa and minor activity at 32 kDa, while the latter showed a more complex pattern with gelatinolytic activity at 90, 78, 65, 34, 32 and 24 kDa. When the proteolytic activity patterns of dust mites and cockroach crude extracts were analyzed at three different pH levels, the proteases in both cases were optimally active at pH 6, showed no activity at pH 3.5 and little activity at pH 8.5. The susceptibility of both extracts to a set of well-known protease inhibitors suggested that they are composed of cysteine and serine proteinases, the latter probably being a trypsin-like type. When immunochemical properties were studied, dust mite bands of about 200, 110, 65, 60 and 43 kDa showed immunoreactivity against a polyclonal human anti-dust mite serum, with the band of approximately 200 kDa presenting the highest antigenicity. A similar analysis was applied to the cockroach extract, which exhibited immunoreactive bands at 90, 78, 65 and 34 kDa when incubated with a polyclonal rabbit anti Blatta serum. Only those of 90, 78 and 65 kDa reacted against a polyclonal human anti-Blatta serum. These results suggested a correlation between some proteases with gelatinolytic activity and the allergenicity of both extracts. PMID- 10513351 TI - Effect of inhaled beclomethasone dipropionate and budesonide dry powder on pulmonary function and serum eosinophil cationic protein in adult asthmatics. AB - The aim of this study was to determine whether the effects of inhaled beclomethasone dipropionate and budesonide dry powder on pulmonary function correlate with those on measurement of serum eosinophil cationic protein (sECP). Thirty-two asthmatic adults in a stable phase, treated daily with 1,000 micrograms beclomethasone dipropionate metered-dose inhaler, completed a 2-week wash-out period and were then randomized to receive a 200 micrograms/dose q.i.d. of either drug, over an 8-week period. Pulmonary function tests (FEV1, FVC, PEFR, FEF25-75% and MEF50) were measured at study entry, before and after every 2 weeks of treatment, while PEFR (morning and evening), symptom scores and salbutamol use PRN were recorded daily on a dairy card. sECP was measured at baseline and after 4 and 8 weeks of treatment. Safety variables included adverse reactions, morning serum cortisol and vital signs (heart rate and blood pressure). FEV1, FVC, PEFR, FEF25-75%, MEF50 and morning PEFR significantly increased (p < 0.05) over baseline in the beclomethasone group, while only FEV1 at week 6 and evening PEFR significantly increased (p < 0.05) in the budesonide group; no significant differences between groups were reported. sECP significantly decreased (p < 0.01) in the beclomethasone group at week 4 and 8 (p < 0.05 between groups). Evidence of statistically significant negative correlation between the FEV1 percent predicted and sECP was assessed at baseline (correlation coefficient r = -0.60, p < 0.05) in the total patient sample, and in the results, expressed as percent change over baseline, obtained at both week 4 and 8 (p < 0.01). A significant decrease in salbutamol use PRN, symptom score and number of daily bronchospasm attacks was also reported in the beclomethasone group (p < 0.05). No adverse reactions or relevant changes in morning cortisol and vital signs were reported in either group. It was concluded that sECP proved to be a reliable marker for monitoring inflammatory events in asthma; inhaled beclomethasone dipropionate dry powder was at least as effective as budesonide in improving lung function and the underlying asthma inflammation. PMID- 10513352 TI - Elevated CD154 (CD40 ligand) synthesis in T-cells from allergic patients after nonspecific stimulation in vitro. AB - The interaction of the CD154 molecule (CD40 ligand, gp39) on activated T-cells with the CD40 antigen on B-cells seems to play a key role in immunoglobulin class switching. We aimed to compare the capacity of intracellular CD154 expression after nonspecific stimulation with phorbol-12-myristate-13-acetate and ionomycin on separated T-cells from allergic patients and healthy donors. We analyzed blood from 104 patients allergic to grass pollen, house dust mites or birch pollen, and from 44 healthy donors. Lymphocytes were isolated using a density gradient and B cells were extracted by magnet-activated cell separation (MACS) using anti-CD19 microbeads. Cells were nonspecifically stimulated for 5 h, permeabilized and stained with anti-CD154 for fluorescence-activated cell sorter analysis. It was found that stimulation induced a 1.4% increase of intracellular CD154+ T-cells; a 4.6% increase of mean channel fluorescence of all T-cells from healthy donors; a 6.1% increase in intracellular CD154+ T-cells; and a 28.1% increase of mean channel fluorescence of all T-cells from allergic patients. The data demonstrated an elevated capability of B-cell independent CD154 synthesis in T-cells from allergic patients when compared to healthy individuals. It is possible that the enhanced IgE production of B-cells from allergic patients might be partly due to the phenomena described. PMID- 10513354 TI - Effect of fluticasone propionate on interleukin-12 and interferon-gamma production in patients affected by allergic bronchial asthma. AB - Fluticasone propionate is a very effective topical treatment for asthma. Interleukin-12, which is produced by monocyte-macrophage and B lymphocytic cell lines, plays an important role in the induction of Th1 cells; whereas interferon gamma (IFN-gamma), mainly produced by Th1 cells, exhibits a crucial effect on macrophage and natural killer cell activation. Since previous studies have demonstrated a reduced production of IL-12 and IFN-gamma in patients with allergic asthma, we examined whether fluticasone propionate therapy could influence the release of these cytokines in asthma. We selected two groups of subjects (15 per group): nonatopic healthy donors (group A) and asthmatic patients (group B). Cytokine release was assessed in sera as well as in supernatants of whole blood cultures after IFN-gamma priming and lipopolysaccharide stimulation. Venous blood and spirometric tests from patients in group B were obtained before and after treatment. Fluticasone propionate therapy significantly increased interleukin-12 levels in both supernatants of blood cultures (1,152.12 +/- 225.57 vs. 540.87 +/- 130.07 pg/ml; p < 0.05) or in sera (72.24 +/- 15.76 vs. 10.83 +/- 2.70 pg/ml; p < 0.05). Patients treated with fluticasone propionate also displayed increased levels of IFN-gamma in either blood culture supernatants (59.12 +/- 16.88 vs. 18.87 +/- 7.53 IU/ml; p < 0.05) or in sera (5.60 +/- 2.87 vs. < 1 IU/ml; p < 0.05). In addition, we observed a significant increase in FEV1 values in asthmatic patients after fluticasone propionate therapy (51.86 +/- 5.31 vs. 71.46 +/- 10.37% predicted). We propose that fluticasone propionate may exert at least in part of its antiinflammatory activity through the modulation of the cytokine output. PMID- 10513353 TI - Effects of sampling height and climatic conditions in aerobiological studies. AB - This study examined the effect of sampling height on the measurement of airborne particles (pollen grains) common in the sampling area in the outskirts of the city of Cordoba, Spain. The effect of certain meteorological parameters on variations in concentration at different heights were also examined. The study was carried out throughout 1991 and 1992 using two Hirst samplers placed at two different heights (1.5 and 15 m) at the Faculty of Science at the University of Cordoba. The statistical results indicated that there were significant differences in the concentrations obtained at different heights, the values at 1.5 m being generally higher with the exception of pollen belonging to the Urticaceae family. The pollen counts of this type were greater at the higher elevation, probably due to the small size of the pollen, especially in the Urtica membranacea species, and to the convective phenomena in this climatic zone in spring, the season in which this species blooms. When these height comparison studies were conducted, the importance of the effect of placing the sampler in relation to a nearby building was also observed. Higher pollen concentrations were detected when the lower sampler was located on the leeward side. The meteorological parameters studied had some influence on the vertical dispersion of the pollen, although the percentage of variation according to height was very small, probably due to the short duration of the study. However, a certain relation between the differences in concentration per height and the degree of atmospheric stability was observed. PMID- 10513355 TI - Beer-induced anaphylaxis due to barley sensitization: two case reports. AB - Despite the large worldwide beer consumption, allergic reactions have very rarely been reported. We describe two cases of severe systemic reactions due to beer ingestion: one case of anaphylaxis requiring emergency care and one of generalized urticaria and angioedema. The two patients underwent a detailed diagnostic procedure involving skin testing, oral challenge with additives, detection of specific IgE to the components of beer, and oral challenge with beer in one patient. The results of the tests, in the particular skin tests and IgE assay, allowed us to detect barley as the specific ingredient responsible for the observed allergic reactions to beer. Therefore, such a sensitization should always be taken into account in the case of suspected reactions following beer ingestion. PMID- 10513356 TI - Osteochondritis dissecans: a multicenter study of the European Pediatric Orthopedic Society. AB - To assess of the value of conservative and operative treatment of osteochondritis dissecans of the knee, a multicenter study was performed. In 12 European countries, 798 cases of osteochondritis of the knee have been collected from 44 hospitals. Results were based on 452 patients with 509 affected knees with minimum follow-up was 1 year (mean follow-up, 3 years and 11 months) and sufficient data for evaluation: 61% were male patients; 39% female patients; 318 affected knees were found in juvenile patients; 191 affected knees were in adult or premature patients. The localization was the medial femoral condyle on the lateral side in 51% (typical site) of patients. Various other sites were involved. Of the 509 affected knees, 154 were treated conservatively, 355 were treated surgically (many with multiple operations). For evaluation, the initial situation (at the time of the diagnosis) was favorable in 198 patients (no effusion, diameter of the lesion < 20 mm and no gross dissection on imaging) and unfavorable (one of the parameters did not meet these prerequisites) in 311 patients. The results were better in young patients than in adult patients. However, in the adolescent group, 22% of patients had abnormal knee at follow-up. The classical localization has a better prognosis than an unusual one. Patients with a favorable situation at diagnosis have significantly better results after conservative treatment than those who have undergone operation. When there are signs of dissection, the results are better after operative than after conservative treatment. PMID- 10513357 TI - Clubfoot surgical treatment: preliminary results of a prospective comparative study of two techniques. AB - Twenty patients with 30 idiopathic resistant clubfeet were operated on by the same surgeon. The mean patient age was 7.7 months (range, 3.5-19 months). Two different surgical techniques (15 posteromedial release and 15 complete circumferential subtalar release) were used during a prospective randomized study. Average follow-up was 2 years 3 months. None of the children had received previous conservative treatment. Radiologic assessment on lateral and anteroposterior radiographs included preoperative and follow-up measurements of tibiotalar, tibiocalcaneal, talocalcaneal, talo-I meta, and calcaneo-V meta angles, as well as physis morphology, talocalcaneal divergence, and location of the navicular. Before surgery, both groups were statistically similar as assessed by the Student t test. Follow-up results were also statistically similar between the groups. Functional assessment, according to Magone's score, showed global average excellent and good results in 23 feet (76.7%), with a slight but not significant difference (P = 0.77) between the two techniques. At short-term follow-up, no significant differences were found in radiologic and functional results between the two surgical procedures for idiopathic clubfoot. PMID- 10513358 TI - Treatment of severe residual clubfoot deformity by trans-midtarsal osteotomy. AB - This study reviews the preliminary results of transmidtarsal osteotomy performed on 11 patients (12 feet) who previously underwent surgery for resistant clubfoot and needed further surgery for severe residual deformities. Opening wedge medial cuneiform osteotomy, closing wedge cuboid osteotomy, and truncated wedge middle and lateral cuneiform osteotomy were performed. The procedure was performed initially on normal cadaver feet. The average improvement of anteroposterior talo first metatarsal angle was 20 degrees and lateral calcaneo-first metatarsal angle was 16 degrees. The authors conclude that with this simple procedure, angular and rotational correction in three planes can be obtained simultaneously in severe residual clubfoot deformity without the need for extensive soft tissue release. PMID- 10513359 TI - Calcaneocuboid malalignment in clubfoot. AB - In a series of 179 clubfeet treated surgically with a follow-up of 3 to 14 years, the clinical significance of calcaneocuboid malalignment was assessed on the basis of a standardized anteroposterior radiograph. The revision rate was 15% and the clinical requirement for a further soft tissue release was related to the talocalcaneal and calcaneocuboid angles. Calcaneocuboid malalignment does not have an adverse effect on the good prognosis of an otherwise well-corrected foot and does not alter the surgery needed to improve a clearly uncorrected foot. When talocalcaneal correction is doubtful, calcaneocuboid malalignment should tilt the balance toward a revision and is of value when the navicular has yet to ossify. Surgical release of the calcaneocuboid joint is unnecessary, particularly the lateral dissection, provided that the medial and subtalar dissection is complete. PMID- 10513361 TI - Concordance between hip ultrasonography and hip arthrography in the assessment of developmental dysplasia of the hip. AB - Controversy exists regarding the possibility of predicting hip reducibility in the congenitally dislocated hip, with arthrography still regarded as the gold standard in this situation. This study aims at assessing the degree of concordance between ultrasonography and arthrography in the detection of anatomic elements obstructing hip reduction. Forty-nine hips were studied both by ultrasonography and arthrography. Three anatomic sources of obstruction to reduction were assessed in each hip: ligamentum teres hypertrophy, inverted labrum, and the presence of soft tissue in the acetabulum. For each variable, congruence between ultrasound and arthrography was measured by kappa analysis. Values > 0.40 expressed sufficient concordance, and they were detected with regard to inverted labrum and the presence of soft tissue in the acetabulum. The results of this study suggest that ultrasonography may be considered a reliable technique for the prediction of the main causes of obstruction in the congenitally dislocated hip, such as inverted labrum and soft tissue in the acetabulum. PMID- 10513360 TI - Normal collagen structure in the posterior ankle capsule in different types of clubfeet. AB - In 21 clubfeet (17 children) of different types (idiopathic, spina bifida, syndromal), the authors analyzed modifications and crosslinks in collagen and their possible relations with clinical stiffness of clubfoot deformity, as measured by the Dimeglio/Bensahel method. In a biopsy of the capsule of the ankle joint, the number of hydroxylysine residues (mean +/- standard deviation; 27.1 +/ 3.4) and the number of pyridinium crosslinks per collagen molecule (0.41 +/- 0.12) were normal. This indicates normal processing of collagen molecules and normal alignment of collagen molecules within fibrils despite the variety of clubfeet, and even in scar tissue. PMID- 10513362 TI - Acetabulum-head index in children with normal hips: a radiographic study of 154 hips. AB - The acetabulum-head index (AHI), which is used to assess femoral head coverage on plain radiographs, was measured in 77 children (154 hips) with normal hips aged 2 to 14 years. The mean AHI value was 94 (range, 79-114). Both the intraobserver and the interobserver reproducibility of the measurements was high. The AHI values tended to decrease with increasing age. The mean AHI minus 2 standard deviations, which was used to define the border value for subluxation of the femoral head, was 80. The authors propose that an AHI < or = 80 can be used as a guideline to reveal abnormal lateral displacement of the femoral head in children. PMID- 10513363 TI - Femoral neck lengthening osteotomy after growth disturbance of the proximal femur. AB - Treatment of congenital dislocation of the hip, Perthes disease, bacterial coxitis, or fractures in childhood may be complicated by vascular insufficiency and subsequent growth disturbance of the proximal femur. The resulting deformity, with a high-standing greater trochanter and a short femoral neck, causes leg length shortening and insufficiency of the hip abductors with a positive Trendelenburg sign and limp. Normal anatomy and biomechanics of the hip joint can be restored by lengthening the femoral neck after two parallel osteotomies of the femur at the the upper and lower border of the femoral neck, followed by distalization the greater trochanter. This femoral neck lengthening osteotomy was first described by the senior author (EM) in 1980. This retrospective study of 37 operated patients with a mean follow-up of 8 years shows good results in 32 patients with little or no preexisting osteoarthritis. Four of five patients with marked degenerative changes underwent a total hip replacement within 1 to 9 years after the osteotomy. PMID- 10513364 TI - Ilizarov fixator for treatment of Legg-Calve-Perthes disease. AB - Eleven hips of 11 patients (8 boys, 3 girls; mean age, 7.5 years) with a diagnosis of Perthes disease underwent distraction using an Ilizarov external fixator. All patients had one or more Catterall signs of poor prognosis, with four hips classified as Herring class B and seven as class C. Patients were followed for a mean of 36 months. Average time for wearing the fixator was 99 days (range, 40-150 days). After fixator removal, containment was lost in two more patients and was sustained in only four patients. The most common complication was pin track infection, which occurred in eight patients. The low rate of success found does not justify the routine use of this technique. PMID- 10513365 TI - Correction of knee deformity in patients with Ellis-van Creveld syndrome. AB - Six knees in three patients with Ellis-van Creveld syndrome were treated with lateral soft tissue release and corrective osteotomy of the tibia at 10 years of age on average. The main feature was valgus deformity with lateral dislocation of the patella. All patellae were reduced. The valgus deformity improved from 35 degrees (range, 48 degrees-20 degrees) to 17 degrees (range, 35 degrees-5 degrees) of the femorotibial angle (FTA) on average, although the FTA in five of six knees was < 5 degrees after surgery. There was one recurrent case and one transient peroneal nerve palsy. The reason for undercorrection was a depression of the lateral tibial plateau. The deformity of the articular surface is the most important problem in correcting the valgus deformity of the knee in this syndrome. PMID- 10513366 TI - Definition, quantification, and correction of translation deformities using long leg, frontal plane radiography. AB - The surgical realignment of mechanical axis deviation is necessary to prevent early joint degeneration. Modern types of external fixation systems allow alignment of the mechanical axis to exact degrees. Predominately, these are corrections of angulation deformities. In some cases, the analysis of the mechanical axis deviation does not show any angulation deformity, but rather a parallel staggering of the mechanical axis lines of a bone. Such parallel staggering of the mechanical axis lines is defined as a translation deformity of the bone. In combined deformities with angulation and translation, the center of deformity can be established proximal or distal to the limit of the bone. In translation deformities, the realignment of the mechanical axis requires a parallel restaggering made by a translation-osteotomy or by a counterangulated double osteotomy. In complex deformities with angulation and translation, the translation requires separate corrective planning. In frontal plane radiographs of the standing leg, the components of angulation and translation can be established graphically or by simple trigonometric formulas. The analysis and surgical procedure to realign translation deformities or a combination of translation and angulation deformities using an unilateral fixator device are discussed. PMID- 10513367 TI - Relationship between radiologic morphology of the bone lengthening formation and its complications. AB - The objective was to study the different types of lengthened bone regeneration and their development during the various phases of the process to correlate them with patient factors and the surgical technique used, and to establish a possible relation between the development of the bone lengthening formation and the problems or complications. The authors studied the radiographs of a random group of 55 patients taken at three points during the course of treatment. The callus was classified with regard to its transverse diameter and the presence or absence of hypodense areas. The overall callus type was significantly influenced by the etiology, the osteotomy site, and the percentage lengthened. The percentage by which the limb was lengthened at the beginning of the process influences the overall morphology of the callus. Poor callus had been lengthened the most, atrophic callus the least. There was a correlation between the morphology of the overall callus at the end of treatment and the percentage lengthened, and between the percentage lengthened and the presence of bands at the end of treatment. The authors also found a significant correlation between age and the appearance of bands at the end of distraction. A central band was found among younger patients. The type of osteotomy affected the overall callus at the end of distraction and at the end of treatment and also influenced the transverse diameter. All the elongations with poor bone formation at the end of treatment were found to have undergone a diaphyseal osteotomy. The most common complication at the first follow-up and at the end of distraction was angulation. The diameter of the callus and the presence of bands at the end of treatment were significantly related to the complications. Fracture occurred in the first 2 weeks after removal of the external fixator in 88% of cases and in the third and fourth week in the rest. However, the segment had no significant influence on the appearance of complications. Lengthened callus with incomplete trabecular formations and hypodense areas at the end of the treatment has a high risk of fracture at the end of treatment. Callus with axial deviation, hypodense areas, or an insufficient transverse diameter during the lengthening procedure must be manipulated so that it reaches the maturing phase in better condition. PMID- 10513368 TI - Congenital pseudarthrosis of the clavicle and thoracic outlet syndrome in adolescence. AB - A 15-year-old girl with thoracic outlet syndrome associated with congenital pseudarthrosis of the clavicle was examined. The indication for treatment of congenital pseudarthrosis of the clavicle is discussed. PMID- 10513369 TI - Epidemiologic, bacteriologic, and long-term follow-up data of children with acute hematogenous osteomyelitis and septic arthritis: a ten-year review. AB - A total of 69 children with acute hematogenous osteomyelitis and 48 with septic arthritis admitted in the period 1978 through 1987 were included in a retrospective review. Epidemiologic and bacteriologic data were analyzed and compared with those of an earlier study (1965 through 1975), confined to the same geographic area. Long-term outcome was evaluated by a questionnaire and clinical and radiographic follow-up. A significant increase in the admission rate for both disorders was observed. The long-term outcome was favorable: major sequelae were found in three patients (3%), minor sequelae in two patients (2%). The benign long-term outcome may well be related to rapid hospital admission and appropriate long-lasting antibiotic treatment. PMID- 10513370 TI - Herniation of calcified intervertebral disk in a lumbar vertebral body presenting as acute scoliosis in a child. A case report and literature review. AB - Herniation of a calcified intervertebral disk into the cervical and thoracic spine is a well-documented entity, but herniation of a calcified intervertebral disk into the lumbar spine presenting as acute scoliosis has not been previously reported. PMID- 10513371 TI - The proximal end of the tube. PMID- 10513372 TI - Multifetal pregnancies. AB - Parturients with multiple gestation challenge all members of the perinatal team. Satisfactory maternal and fetal outcome require the experience and expertise of the obstetrician, anesthesiologist and neonatologist, all of whom must be well informed in terms of the total care and management plan. PMID- 10513373 TI - Modification of an anesthesia machine for MRI suite. AB - The modification of a wall mounted anesthesia machine, rendering it nonmagnetic for use in the magnetic resonance imaging suite, is described. The modified anesthesia machine functioned properly at 1.8 m distance from 1.5 tesla electromagnet without degrading its imaging. PMID- 10513374 TI - Congenital heart disease at a tertiary care center in Lebanon. AB - OBJECTIVE AND METHODS: To study the epidemiology of congenital heart disease (CHD) at the American University of Beirut-Medical Center, we reviewed the medical records of all cardiac patients seen at our outpatient cardiology clinic (OPD) between 1980 and 1995. The charts of all patients with CHD seen as inpatients and/or outpatients at our center during the year 1995 were also reviewed. A cardiologist evaluated all patients and the diagnosis was confirmed at least by echocardiography. The frequency of CHD was reported among three groups: 1980-1995 OPD groups (Group A); the group with CHD seen during the year 1995 (Group B); and (Group C), a subgroup of group B, included all newborns with CHD born at our hospital during the year 1995. Stillbirth and premature infants with the diagnosis of patent ductus arteriosus were excluded from the study. RESULTS: Group A included 883 patients. 344 patients were evaluated in Group B, with a mean age of 3.8 years. The incidence of CHD was 11.5/1,000 live births at our center. There was a relatively low prevalence of complex lesions (i.e., hypoplastic left heart syndrome, transposition of the great arteries) and a relatively high prevalence of the simpler cardiac malformation (i.e., ventricular and atrial septal defects, pulmonary stenosis) in Groups A and B. CONCLUSION: The relatively low prevalence of complex cardiac lesions in our study is probably related to the age of the studied patients, and reflects the high mortality of these complex lesions in our country early in life. The incidence of CHD of 11.5/1,000 live births at our center is higher than that reported in the literature, with evidence of more frequent ventricular septal defects and pulmonary atresia lesions. This may be related to high rate of consanguinity in our population. This review underscores the need for a national cardiac registry center for children in a developing country like Lebanon. Such a database will allow referral and care of complex cardiac lesions. PMID- 10513375 TI - Difficult airway in obstetrics. PMID- 10513376 TI - Alfentanil decreases myoclonus caused by etomidate. PMID- 10513377 TI - Death within 24 hours of anesthesia is not always an anesthetic death. AB - PURPOSE: We report an anesthetic death of a young lady after uneventful anesthetics. CLINICAL FEATURES: A 25-year-old female who had undergone emergency incision and drainage of an abscess in her right breast. Her condition, however, deteriorated few hours after anesthesia and died in less than 24 hours from causes unrelated to anesthesia. The computerized tomography (CT) scan showed a large meningioma in frontal lobe. CONCLUSION: This case illustrates that the risks of anesthesia are increased significantly in patients with silent meningionmas. PMID- 10513378 TI - Progression of first degree atrioventricular block to second degree Mobitz type I block during spinal anesthesia associated with induced hypertension. AB - A case is presented in which an elderly patient with preexisting first degree atrioventricular (AV) block progressed to second degree Mobitz Type I AV block during spinal anesthesia and associated with hypertension induced by a pure alpha 1 agonist. Second degree AV block caused by increased vagal tone was transient, which resolved as the blood pressure normalized. Hypotension due to spinal anesthesia treated with pure alpha 1 agonist can increase AV block in patient with pre-existing first degree heart block. PMID- 10513379 TI - The modified Guedel airway for fiberoptic intubation. PMID- 10513380 TI - [Academician Evgenii Mikhailovich Kreps (on the centenary of his birth)]. PMID- 10513381 TI - [The antioxidant prevention of disorders in calcium ion metabolism under the action of glutamate on the synaptosomes of the rat cerebral cortex]. AB - An increase of intracellular calcium ion concentration and of the 45Ca2+ entry, a decrease in Na+,K(+)-ATPase activity, and activation of Na+/Ca2+ exchange were shown to be initiated by glutamate in the rat brain cortex synaptosomes. These effects could be prevented with antagonists and blocking agents of the NMDA receptors. Pre-incubation of the synaptosomes with alpha-tocopherol, superoxide dismutase, and ganglioside GM1 was shown to normalise [45Ca2+], the rate of 45Ca2+ entry, and the activity of Na+,K(+)-ATPase in the synaptosomes. The data obtained suggest that calcium ions entering the brain cortex neurones via the NMDA receptors in presence of excessive glutamate, trigger activation of free radical reactions damaging the neurones in ischemia, cerebral lesions, and other pathological conditions. PMID- 10513382 TI - [The participation of glutaminergic transmission in the mechanism of movement disorders induced by reverse rotation in white mice]. AB - Administration of MK-801 or IEM-1754 prevented akinesia in mice induced by reversing rotation, not less effectively than scopolamine. Quaternary adamantane derivative IEM-1857 was ineffective. IEM-1925 enhanced the locomotor disturbance induced by reversing rotation due, probably, to different spectrum of its antiglutamate action. The data obtained suggest involvement of glutamate synaptic transmission in development of locomotor disturbances of a vestibular origin. PMID- 10513383 TI - [Age-related changes in the activity of free-radical processes in rat tissues and blood serum]. AB - Activity of Cu,Zn-superoxide dismutase (SOD) in old rats' brain was found to be decreased by 46.8% as compared to young animals. The brain concentration of Schiff bases (SB) was decreased by 13.6% in old rats, whereas concentration of diene conjugates (DC), protein peroxidation (PP), and total antioxidative activity (TAA) was the same in old as well as in young rats. The liver level of the DC and TAA was also the same. The serum level of the PP, SB, and DC was increased whereas the activity of the SOD and TAA was decreased in old rats. The findings suggest occurrence of considerable age-related changes in free radical processes as well as the organ specifics of these changes in rats. PMID- 10513385 TI - [The colocalization of neurotransmitters in the presynaptic boutons of inhibitory synapses in the lamprey spinal cord]. AB - Using the electron microscopy immunocytochemistry, the GABA and glycine immunoreactivity was studied in presynaptic axon terminals of the spinal cord central gray in the lamprey Lampetra fluviatilis. All immunopositive presynaptic terminals contacting motoneurones or non-identified post-synaptic profiles were divided into only GABA- (44%), only glycine-immunopositive terminals (26%), and both GABA- and glycine-containing terminals (30%). The glycine-immunopositive axon terminals contained flattened synaptic vesicles. Large dense core vesicles were co-localised with conventional synaptic vesicles in 74% of GABA-containing presynaptic terminals. PMID- 10513384 TI - [The action of ADP ribose on the mechanical and bioelectrical activity of the frog heart]. AB - In the frog isolated heart, cyclic perfusion of ADP-ribose induced a dose dependent decrease in the heart rate and the contraction force, a decrease in the AP duration as well as in the rate of rise in the sinus node. It also shortened the atrial AP and exerted no significant effect upon multicellular ventricular preparations. In conditions of systemic administration in unanesthetised frogs, the ADP-ribose induced a reversible increase in the heart rate due, probably, to a sympathetic effect. PMID- 10513386 TI - [A new class of N-methyl-D-aspartic acid receptor agonists and antagonists- derivatives of imidazole-4,5- and pyrazole-3,4-dicarboxylic acids]. AB - New agonists and antagonists of the N-methyl-D-aspartic acid (NMDA) receptors were found among the derivatives of 1- or 2-alkyl-substituted imidazole-4,5- and pyrazole-3,4-dicarboxylic acids. Lipophilic surrounding of the nitrogen atom in these compounds was found to determine their ability to be either agonists or antagonists, while the distance between the terminal acidic functions was the same. An increase in the lipophilicity can also cause loss of selective action upon the NMDA receptors and occurrence of non- NMDA antagonistic activity. PMID- 10513387 TI - [The effect of oxidative stress on brain nitric oxide synthase activity in vivo and in vitro]. AB - Within 8 days of a 10-min cardiac arrest, accumulation of material reacting with 2-thiobarbituric acid was revealed in the hippocampus (74%) and cerebellum (47%) of male Wistar rats. Oxidative stress was accompanied by a twofold decrease of the nitric oxide synthase activity in the brain tissue. In vitro experiments showed a dose-dependent decrease of nitric oxide synthase activity in the brain homogenates as a result of the oxidative stress induced by hydrogen peroxide or sodium hypochlorite. The findings suggest that the oxidative stress may decrease nitric oxide synthase activity as a result of direct effects of the active oxygen on the enzyme. PMID- 10513388 TI - [The importance of the ion-transport systems of the sarcolemma and sarcoplasmic reticulum in changing rat cardiac contractile function under a hypersodium medium]. AB - Following a reduced pressure in the left ventricle, elevated concentrations of sodium ions enhanced by half the contraction force of the rat isolated heart. This effect was shown to be independent of the Na-channels blockers or Na/H exchange of caffeine but quite susceptible to sodium channel blockers, caffeine, and the blocking agent for Na-Ca exchange Ni2+. A decrease in potassium concentration amplified, and elevation of K+ level attenuated the positive inotropic effect of the elevated concentration of sodium ions. The effect was preserved even after heart arrest induced by verapamil. The findings suggest that elevated concentration of sodium ions may affect the Na+/Ca2+ exchange and provoke Ca2+ release from sarcoplasmic reticulum by means of changing the sodium gradient. These data corroborate the Leblanc and Hume hypothesis of the sodium induced calcium ions release from sarcoplasmic reticulum. PMID- 10513389 TI - [The dynamics of the chronotropic effect of a single vagal stimulus in cats under the action of met-enkephalin and neurotensin]. AB - Following a burst of pulses applied to the vagus nerve with progressively incremental delay after the P wave of the ECG, the narrow zone of the cardiac cycle was identified where even a small shift of the vagal burst position evoked an abrupt alteration of the chronotropic effect magnitude. Met-enkephalin potentiated the phase-dependent vagal chronotropic effect, whereas neurotensin moved its limits toward the initial part of the P-P interval. PMID- 10513390 TI - [The effect of stimulation of the adrenergic receptors on the frequency depression of the temporal parameters of the action potential in the right atrium of rats]. AB - At a stimulation rate of 1 Hz, activation of alpha-adrenoreceptors prolonged the AP duration at 10%, 50%, and 90% repolarisation at 10(-7), 10(-6) M in the rat isolated right atria, but shortened it at a higher concentration of 10(-5) M. The frequency-induced depression of the AP duration became more evident in cardiomyocytes stimulated by 10(-7), 10(-6) M and less obvious at 10(-5) M of alpha-adrenoagonist. Activation of alpha-adrenoreceptors by isoprenalin shortened the AP duration and enhanced the stimulation-rate-dependent changes in the atrial AP configuration. PMID- 10513391 TI - [The role of adrenergic stimulation of the arteries in their developing rhythmic contractions in arterial normo- and hypertension in rats]. AB - In anaesthetised Wistar and SHR rats, rhythmic contractile responses of mesenteric arteries to noradrenaline as well as sympathetic nerve stimulation, were studied. Under equal experimental conditions, different parameters of vasomotion in normo- and hypertensive rats were revealed. PMID- 10513392 TI - [The characteristics of the cholinoreceptors on the identified TAN neuron of the giant African snail Achatina fulica]. AB - Acetylcholine, nicotine, a selective agonist of N-cholinoreceptors suberildicholine dibromide, as well as a selective agonist of M-cholinoreceptors 5-methylfurmethide inhibited spike discharges in a dose-dependent manner up to a complete ceasing of the firing in cholinoreceptors situated on the identified neurone TAN of African giant snail Achatina fulica. M-cholinoblocker metamizylum completely prevented the inhibitory effect of methylfurmethide. Central cholinoblocker aetherophen completely prevented the inhibitory effect of suberildicholine dibromide. Metamizylum or aetherophen used alone were only able to decrease the inhibitory effect of acetylcholine, whereas a mixture of these agents suppressed completely the acetylcholine-induced inhibition. The findings suggest that, on the TAN membrane, nicotinic and muscarinic cholinoreceptors co exist and function in one and the same direction. PMID- 10513393 TI - [The effect of the phosphorylation of the Na+,K(+)-ATPase alpha subunit in rat kidneys by protein kinase C on the activity of the enzyme]. AB - Phosphorylation was shown to lead to a change in the conformational equilibrium toward E1 form associated with a decrease in apparent affinity for the K+ in alpha-1 subunit of the rat kidney Na+, K(+)-ATPase. Rate of transition from E2 to E1 was apparently unaffected by phosphorylation. ATP hydrolysis by the protein kinase C-phosphorylated Na+, K(+)-ATPase shows a decrease in the Vmax and Km for K+. PMID- 10513394 TI - [The evolution of osmoregulation in vertebrates (on the centenary of the birth of E. M. Kreps)]. AB - The paper substantiates the point of view that the first step in the evolution of osmoregulation in vertebrates involved development of water-impermeable epithelium which depended on specific distribution of aquaporins in plasma membranes, secretion of autacoids, formation of water-impermeable cell contacts with a high electrical resistance. Migration of vertebrates to the land was associated with appearance of the ability to regulate osmotic permeability of epithelium in a number of organs involving vasotocin and other pituitary hormones, insertion of aquaporins into the luminal plasma membrane. Enhancement of the renal role in the water-salt balance, intensification of the kidney function was due to a rise in renal circulation, glomerular filtration rate, and tubular reabsorption which, despite the increase in the energy expenditure, provided for an obvious enhancement of the renal work efficiency in maintaining the constancy of the internal medium. Colonisation of desert regions became possible in birds and mammals due to development of the system of urine osmotic concentration. Unlike the accepted point of view that this depended on the appearance of Henle's loops, the statement is substantiated that the leading role in this process is played by differentiation of the kidney into the cortex and medulla, as at our Laboratory nephron loops similar to Henle's loop have been revealed in the lamprey. In the vertebrates, the evolution shifted from the polyorgan to mono-organ type of osmotic homeostasis maintenance, with the leading role of the kidney in maintaining physico-chemical constants of the internal medium fluids. PMID- 10513395 TI - [The participation of the striatum in the central regulation of gonadal hormonal function]. AB - Intrastriatal corticotrophin-releasing hormone (CRH) was shown to induce a decrease in the plasma tectosterone concentration. Besides, the 6-OHDA pre treatment completely prevented the suppression of plasma testosterone in response to intrastriatal CRH administration. The findings suggest that the striatum is involved in the extrahypothalamic regulation of the gonadal endocrine function. PMID- 10513396 TI - The role of phonology in the activation of word meanings during reading: evidence from proofreading and eye movements. AB - Six experiments explored the role of phonology in the activation of word meanings when words were embedded in meaningful texts. Specifically, the studies examined whether participants detected the substitution of a homophone mate for a contextually appropriate homophone. The frequency of the incorrect homophone, the frequency of the correct homophone, and the predictability of the correct homophone were manipulated. Also, the impact of reading skill was examined. When correct homophones were not predictable and participants had a range of reading abilities, the evidence indicated that phonology plays a role in activating the meanings of low-frequency words only. When the performance of good and poor readers was examined separately, the evidence indicated that good readers primarily activate the meanings of words using the direct route, whereas poor readers primarily activate the meanings of words using the phonological route. PMID- 10513397 TI - Global-local interference modulated by communication between the hemispheres. AB - Three experiments examined whether interhemispheric interaction modulates selective attention in a same-different version of D. Navon's (1977) global-local paradigm. In Experiments 1 and 2, interhemispheric interaction reduced interstimulus interference produced when two stimuli matched at a preassigned level (e.g., local) but differed at the irrelevant level (e.g., global). This effect was greater for stimuli made of a few large elements than for those made of many small elements. Experiment 3 demonstrated that (a) the ability of interhemispheric interaction to reduce interstimulus interference is not constrained by hemispheric differences for global and local processing and (b) interhemispheric interaction does not strongly modulate intrastimulus interference produced when the forms at the preassigned (e.g., local) and irrelevant (e.g., global) levels differ within an individual stimulus. These findings indicate that interaction between the hemispheres is a neural mechanism that may aid selective attention. PMID- 10513398 TI - Working memory, short-term memory, and general fluid intelligence: a latent variable approach. AB - A study was conducted in which 133 participants performed 11 memory tasks (some thought to reflect working memory and some thought to reflect short-term memory), 2 tests of general fluid intelligence, and the Verbal and Quantitative Scholastic Aptitude Tests. Structural equation modeling suggested that short-term and working memories reflect separate but highly related constructs and that many of the tasks used in the literature as working memory tasks reflect a common construct. Working memory shows a strong connection to fluid intelligence, but short-term memory does not. A theory of working memory capacity and general fluid intelligence is proposed: The authors argue that working memory capacity and fluid intelligence reflect the ability to keep a representation active, particularly in the face of interference and distraction. The authors also discuss the relationship of this capability to controlled attention, and the functions of the prefrontal cortex. PMID- 10513399 TI - Degradation of Bowman-Birk protease inhibitor mRNA with a cell-free extract. AB - A cell-free mRNA degradation system consisting of the polysomal and postpolysomal fractions was obtained from cultured soybean cotyledons for studying Bowman-Birk protease inhibitor (BBPI) mRNA stability. This in vitro system reflected the shorter in vivo half-life of BBPI mRNA of cotyledons cultured in basal-medium as compared to cotyledons cultured in methionine-supplemented medium. Most of the BBPI mRNA degradative activity was found to be present in the postpolysomal supernatant fraction. The higher rate of BBPI mRNA degradation in basal medium cultured cotyledons was due to an increased destabilizing activity specific to BBPI mRNA in the postpolysomal fraction from basal-medium cultured cotyledons. The specificity was absent when purified RNA was used as the substrate. Degradation of the mRNA was not divalent cation-dependent and was inhibited in the presence of higher concentrations of monovalent and divalent cations. PMID- 10513400 TI - Purification and characterization of a protein phosphatase from winged bean. AB - A protein phosphatase (WbPP) has been purified from the soluble fraction of the winged bean (Psophocarpus tetragonolobus) shoot extract. The preparation is essentially homogenous as shown by the constant specific activity of the enzyme across the peak fractions, eluted from the thiophosphorylated histone-Sepharose affinity column, the last step of purification and by single protein bands on polyacrylamide gel electrophoresis (PAGE) in the presence as well as absence of denaturating agents. The monomeric nature of WbPP is revealed by an M(r) of 92,000 and 85,000, respectively, as estimated by SDS-PAGE and gel permeation chromatography under non-denaturating conditions. Autophosphorylated calmodulin like domain protein kinase (P-WbCDPKI) [Saha, P., & Singh, M. (1995). Biochem. J., 305, 205] and phosphohistone H1 (P-hisH1), prepared by using the other homologous CDPK, i.e. WbCDPKII [Ganguly, S., & Singh, M. (1998). Phytochemistry, 48(1), 61], are good substrates of the purified enzyme, while P-hisH1 and phosphocasein prepared by using heterologous cAMP-dependent protein kinase, are respectively very poor and totally inactive as substrate. WbPP is adjudged to be a protein phosphoserine phosphatase since phosphoserine is the only phosphorylated amino acid residue detected in our earlier analysis of P-WbCDPKI and P-hisH1. The enzyme is strongly stimulated by a combination of Mg2+ and Ca2+, without being dependent on either of them and is also unaffected by calmodulin and fluphenazine. Orthovanadate strongly inhibits the enzyme while okadaic acid is a poor inhibitor. PMID- 10513401 TI - Two 6-substituted 5,6-dihydro-alpha-pyrones from Ravensara anisata. AB - The leaves and bark dichloromethane extracts of Ravensara anisata showed antifungal activity against the yeast Candida albicans and the phytopathogenic fungus Cladosporium cucumerinum in bioautographic TLC assays. Activity-guided fractionation afforded two new alpha-pyrones: 6R*-(4R*-acetoxy-2S*-hydroxy- 8 phenyloctyl)-5,6-dihydro-2-H-pyran-2-one and 6R*-(2S*-acetoxy-4R*- hydroxy-8 phenyloctyl)-5,6-dihydro-2-H-pyran-2-one. Their structures have been established by NMR spectroscopy, chemical methods and X-ray crystallographic analysis. The antifungal activity against C. albicans and C. cucumerinum was determined for both compounds. PMID- 10513402 TI - Microbial hydroxylation of natural drimenic lactones. AB - Incubation of confertifolin and isodrimenin with Mucor plumbeus, Aspergillus niger or Rhizopus arrhizus gave in good yields the corresponding 3 beta-hydroxy derivatives. From isodrimenin, the known natural 7 alpha-hydroxy derivative (futronolide) was also obtained and its structure was definitely established by X ray crystallographic study of its acetate derivative. PMID- 10513403 TI - Antioxidants from Lespedeza homoloba. (I). AB - The stems of Lespedeza homoloba yielded eight new and three known phenolic compounds. Their structures have been elucidated on the basis of their spectral data. These compounds had strong antioxidative activity against lipid peroxidation in the rat brain homogenate test. 3,9-Dihydroxypterocarp-6a-en and lespedezol A2 showed significant antiallergic activity in allergic (type I) mice. PMID- 10513404 TI - Antioxidants from Lespedeza homoloba (II). AB - The stems of Lespedeza homoloba yielded fifteen new isoflavonoids and a new stilbenoid having antioxidative activity. Their structures were determined by analysis of spectroscopic evidence. PMID- 10513405 TI - The nutrient-toxin dosage continuum in human evolution and modern health. AB - Recent findings support the long-recognized principle that nutritive and toxic effects of an ingested material depend not only on its nature but very much on its quantity. The well known observation that essential nutrients can be toxic at high dosages suggests that the same reversal of effect may be true of many substances that could be beneficial but not essential at low dosages (the phenomenon of hormesis). This has been demonstrated for many well known toxins. We suggest a mathematical model that describes these dosage effects as an expected result of the evolution of human metabolic and dietary adaptations for maximizing benefits and minimizing costs of the ingestion or other intake of any substance. Evolved mechanisms for achieving benefits may be unrelated to those for reducing costs. These evolutionary considerations suggest important consequences demonstrable by experimental or epidemiological studies. They also suggest ways in which our evolved dietary adaptations may be currently maladaptive, and individual development of taste preferences poorly calibrated by early experience in modern environments. The apparent reality of hormesis raises the possibility of counterproductive effects of current dosage recommendations and limits for nutrients and pollutants. We propose that some conceptual and factual problems are urgently in need of resolution. Fundamental to evolutionary biology is the tendency for organisms to become increasingly adapted to those environments to which they are most commonly exposed (Parsons 1990). PMID- 10513406 TI - [Remedies and quackeries]. AB - Several unorthodox drugs for healing cancer recommended widely in the USA and Hungary are discussed. Credulity plays a decisive role in the application of the drugs of doubtful effect and/or "remedies" that are undoubtedly without any effect. Typical examples are Krebiozen, Laetrile, Celladam and water with decreased deuterium concentration. In connection with Avemar, the most recent Hungarian remedy with so far clinically unproven physiological effect it is surprising why an expensive seminatural material is sold when the agen, in fact, is a well-known and synthetically easily available compound. PMID- 10513407 TI - [Unusual coordination behavior of D-fructose towards dimethyltin(IV)2+: metal promoted deprotonation of alcoholic hydroxyl groups in aqueous solutions at low pH]. AB - To study the effect of the conformation of sugar hydroxy groups on metal complexation processes, complex formation of eight saccharides (D-fructose, L sorbose, L-arabinose, D-arabinose, D-glucose, D-sorbitol, 2-deoxy-D-glucose and D saccharose) with dimethyltin(IV)2+ cations was investigated in aqueous solution by potentiometric equilibrium measurements, 13C NMR, polarimetric and Mossbauer spectroscopic methods. The experimental results proved that deprotonation of D fructose and L-sorbose is caused by the coordination of dimethyltin(IV)2+ in the unusual low pH interval 4-6 in contrast to the other saccharides deprotonated in analogous way at pH > 8. Increasing the pH of the solution resulted in the formation of further complexes. Stability and composition of the species was determined by potentiometric studies. 13C NMR measurements led to the assignment of the sugar OH groups participating in the processes. Mossbauer investigations in the quick-frozen solutions permitted the determination of the stereochemistry of tin(IV) in the complexes. PMID- 10513408 TI - [Stereochemistry of proton catalyzed dimerization of 3-alkyl-indoles]. AB - Two dimers (I and II) having 3,3'-disubstituted 2-(2'-indolil)-indoline skeleton were prepared by trifluoroacaetic acid treatment of N-acetyl-L-tryptophan methyl ester. Their CD spectra suggest enantiomeric configuration in 2- and 3-positions of indoline ring. The (-)-enantiomer of skatole dimer has nearly the same CD spectrum as the dimer I. X-ray crystal structure determination of (-)-skatole dimer shows, that it has 2S,3S-trans configuration in the indoline ring suggesting the same configuration of dimer I and opposite, 2R,3R-trans configuration of dimer II. PMID- 10513409 TI - [Pseudopolymorphism of NO-SPA, 1-(3,4-diethoxy-benzyl)-6,7-diethoxy-3,4-dihydro isoquinoli ne hydrochloride]. AB - The spasmolytic agent No-Spa, 1-(3,4-diethoxybenzyl)-6,7-diethoxy-3,4 dihydroisoquinoli ne hydrochloride (1) is a synthetic analogue of the naturally occurring alkaloid papaverine. (1) is prone to form stoichiometric crystalline solvates with a number of solvents causing technological and stability problem. Present paper describes the X-ray structure determination of the hemiethanol, hemibenzene and hemihydrochloride structures being the most important and interesting from both practical and theoretical point of view. The solvate formation is facilitated by the presence of the flexible ethoxy group encapsulating the solvent and by the quasi perpendicular position of the isoquinoline and phenyl group. PMID- 10513410 TI - [Stereochemical aspects of narcotic and psychotropic drug action]. AB - Medicinal drugs are predominantly, whereas narcotic and psychotropic drugs are exclusively exogenous compounds. Three further fundamental properties of the narcotic/psychotropic compounds of significant abuse potential is that they all are of natural (plant or microbial) origin, they contain a large number of chiral atoms, and they influence the neurotransmission processes in the central nervous system. For some of them, the existence of corresponding endogenous ligands have recently been reported. Since exogenous compounds and their endogenous ligands are assumed to bind the same target moiety of the receptor, several fundamental questions arise: To what extent can stereochemical relationships be established between the exogenous compound and its endogenous counterpart? Do they have superimposable moieties? Are the corresponding chiral atoms of the same configuration? Is there a chiral-genetic relationship between the exogenous and endogenous compound? Theoretical aspects and answers to all these questions are sought for morphine, cocaine, LSD, tetrahydrocannabinol, amphetamine and related molecules. PMID- 10513411 TI - [Comparative pharmacokinetics of 800 mg of acyclovir-containing Telviran and Zovirax tablets in healthy volunteers]. AB - The authors performed a detailed comparative clinical and pharmacokinetic study with two 800 mg acyclovir containing tablets, namely the Telviran (EGIS Pharmaceuticals Ltd.) and the Zovirax (Wellcome Foundation Ltd.). The determination of the detailed pharmacokinetic parameters and the relative bioavailability was carried out on 24 healthy male volunteers in a two way, open, randomised, cross-over design study after single dose administration. The plasma concentration of acyclovir was determined by a newly developed and validated highly sensitive (LLOQ = 10 ng/ml) HPLC-UV bioanalytical method after sample preparation with RP-18 solid phase extraction method (SPE). The individual pharmacokinetic parameters calculated from the time-plasma concentration curve (tmax, Cmax, AUC0-infinity, AUC0-16, AUC0-t, AUC0-z, AUCRest, t beta 1/2, MRT, Cmax/AUC0-infinity) were compared with statistical methods (ANOVA, ANOVAlog, Confidence interval, Schuirmann, Wilcoxon tests). On the basis of the results of the statistical evaluation and the clinical study, there was no significant difference found between the two acyclovir containing preparations. The comparative pharmacokinetic study demonstrated, that the relative bioavailability of the 800 mg Telviran and Zovirax tablets are equivalent and the two products are bioequivalent. PMID- 10513412 TI - [Transfer possibilities of the mobile phases between different liquid chromatographic techniques for the analysis and isolation of compounds of biological matrices]. AB - After the survey and characterisation of the solid/liquid chromatographic methods, the author summarized the features of overpressure layer chromatography; the disturbing zone and the multi-front effect as well as the elimination of their influence. In light of these effects, the strategy of the mobile phase transfer possibilities is demonstrated between the various analytical and preparative liquid chromatographic methods, with the OPLC playing a central role. The main point of this strategy is that the examination of biological matrices is always begun with unsaturated TLC chamber, in which the compounds to be separated are placed between the Rf values of 0.3 and 0.8. The optimized TLC mobile phase is transferred without changes to the OPLC technique where a prerun is applied. For separation of nonpolar compounds, the prerun can be performed with hexane; for separation of polar substances the prerun can be performed with any component of the mobile phase in which the components are unable to migrate. The selection of this solvent might be considered during optimization of the mobile phase. Using HPTLC chromatoplate and analytical OPLC technique, highly effective separation can be achieved. The scaling-up for the various preparative chromatographic systems can be performed on basis of the applied chromatographic circumstances. The dry-filled preparative (FC, LPLC, MPLC) columns can be equilibrated with the solvent used for the prerun in analytical OPLC, while in case of filling with slurry technique, the slurry has to be prepared using the same solvent as was used for the prerun of OPLC. The air bubbles can be eliminated in both cases by pumping over the appropriate quantity of the solvent used for prerun, afterwards the preparative separation can be started with the optimized unsaturated TLC mobile phase. The author deals separately with the mobile phase transfer possibilities between the different analytical and preparative planar (OPLC and RPC with various chamber types) chromatographic techniques, where the saturation of the vapour phase is the determinant. The strategy is demonstrated through various plant extracts. The mobile phase transfer possibilities between the different solid-liquid chromatographic methods are summarized in comprehensive figures. Figure 13 demonstrates the direct transfer possibilities, different lines show those applicable with various methods: dotted line stands for off-line and thin line for on-line methods, while the ones marked with thick line ensure the transfer of the optimized mobile phase without change between different solid-liquid planar and column chromatographic techniques in case of both--off-line and on-line--procedures. PMID- 10513413 TI - [Pavlov's Koltushi: a scientific center of world importance]. PMID- 10513414 TI - [Probability organization of the lexicon in mental disorders: association with features of hemispheric interaction]. PMID- 10513415 TI - [Evoked potentials in the hemispheres of the cerebral cortex during recognition of facial expression in the right and left visual half-fields]. PMID- 10513416 TI - [Biological principles of individual differences of children in the second half year of life. Communication I. Factor structure of temperament assessment]. PMID- 10513417 TI - [Computer classification of age groups of schoolchildren by the features of the temporal organization of the wave structure of the EEG pattern]. PMID- 10513418 TI - [Features of cerebral organization of the visual recognition process in adult subjects and children, aged 8-10 years. Communication II. Spatio-temporal organization of alpha-components of EEG at different stages of recognition process]. PMID- 10513419 TI - [Vasomotor reactions in children of early postnatal age]. PMID- 10513421 TI - [The personality typology approach to evaluation of populational psychophysiologic potential]. PMID- 10513420 TI - [Clinical and physiologic characterization of therapeutic effects of intrathecal infusion of autologous blood serum in Parkinson's disease]. PMID- 10513422 TI - [Content of somatotropin and other regulators of muscle metabolism in human blood during long-term space flights and hypokinesia]. PMID- 10513424 TI - [Features of anaerobic and aerobic processes in rowers of high sports class]. PMID- 10513423 TI - [Mechanisms of regulation of human posture in the frontal plane while standing]. PMID- 10513425 TI - [Experimental substantiation for the potential for implantation of embryonal cardiomyocytes in the comprehensive treatment of myocardial failure]. PMID- 10513426 TI - [Biochemical determinants and developmental mechanisms of extreme hypoxic hypoxia]. PMID- 10513428 TI - [Erythrocyte deformability in athletes]. PMID- 10513427 TI - [The concept of "laser histochemical surgery" as a nontraditional trend in medicine]. PMID- 10513429 TI - [Eating to nutrition]. PMID- 10513430 TI - Midwestern regional meeting. Chicago, Illinois, USA. September 16-18, 1999. Abstracts. PMID- 10513431 TI - New instructional strategies needed. PMID- 10513432 TI - An introductory course on study skills forming a bridge between traditional and problem based learning (PBL). AB - OBJECTIVE: To introduce entrants in a medical university coming from traditional system of education to student-centered, problem based learning (PBL) and acquaint them with small group dynamics in order to make them life long learners. SETTING: Ziauddin Medical University, Karachi, Pakistan. SUBJECTS: First Year Students of Medical University. DESIGN: The course was of ten hours, divided in six session. In the first session, there were exercises for interaction in small groups, followed by sessions on principles of learning, learning methodologies, PBL, assessment and lastly importance of feedback. RESULT: Evaluation of the course revealed that 63% of the students agreed that the course achieved its objectives, 25% were not sure while 12% disagreed. Seventy one percent agreed that the course promoted student to student interaction, 24% were not sure while 5% disagreed. CONCLUSION: The course made the process of transition from traditional system of learning to more innovative methods of learning, smoother for the students. The general feeling of the faculty members was that these students were able to perform better than the students who had not taken this course. It is too early to determine the success of this course. A follow-up study however is needed to evaluate the effectiveness of this course by determining the performance of students in small group session of Problem-Based Learning. PMID- 10513433 TI - Lecture as a mode of instruction in undergraduate medical education. AB - OBJECTIVE: This study was carried out to determine the place of lecture as a mode of instruction in undergraduate medical education by examining the views of the students about some selected aspects of the lecturing sessions and their ability to recall the varying percentages of the content of the delivered lecture after a specified period of time. METHODS: An observational cross sectional study was carried out in which the final year students of Fatima Jinnah Medical College for Women, Lahore belonging to sessions 1995-1996 and 1996-1997 participated. The study comprised of two parts; first was administration of a questionnaire containing the questions regarding students' opinions about the lectures attended, duration of lecturing sessions and comparison between lecture and clinical classes. The second part consisted of two surprise tests examining the ability of the students to recall the subject taught in a particular lecture after one week and one month intervals. RESULTS: According to 74% students lecturing sessions were not beneficial. Only 12% students remained attentive throughout the lecture their reason being ineffective and boring presentations and 55% participants wanted the existing duration of the lecture period to be halved. Clinical sessions were considered to be superior by 79% respondents. About 34% to 35% students could recall more than 50% of the content of the delivered lecture after one week and 20% were able to do so after one month. CONCLUSION: Consideration should be given to the feedback received and follow up studies should be carried out so that appropriate changes in the curriculum could be induced in order to ensure the effectiveness of the lecture and to avoid wastage of resources and time and to justify the place of lecture as a mode of instruction in undergraduate medical education. PMID- 10513434 TI - Foreign body aspiration in children--a persistent problem. AB - OBJECTIVE: Cough and respiratory distress due to foreign body inhalation in children is a common problem in our society. This study was planned to identify the criteria for early diagnosis and management in suspected cases of foreign body inhalation. SETTING: The study was carried out on indoor patients of Paediatrics Department, Rawalpindi General Hospital (RGH) affiliated with Rawalpindi Medical College (RMC), from January, 1995 to December, 1996. Paediatrics Department has 50-bedded general paediatrics ward, 20-bedded neonatal unit (NNU) and 150-200 daily attendance of outpatient department (OPD). It offers primary to tertiary level care to its patients from whole of Rawalpindi Division. METHODS: This prospective study encompasses profile and immediate outcome of 20 suspected cases of foreign body inhalation. Selection of study subjects and subjection to bronchoscopy was based on symptomatology, clinical and radiological findings and response to treatment. The data with outcome was recorded on a proforma. RESULTS: In 18 cases foreign body was removed successfully by rigid bronchoscope with immediate relief of symptoms and normal chest roentgenograms. Most susceptable age for foreign body inhalation was 1-3 years (n = 14) and male children were at higher risk than females (13 vs 7). Valuable clinical features were sudden onset with respiratory distress, cough, choking, localized poor air entry, crepitations or rhonchi in descending order of frequency. Most common finding in chest roentgenogram was consolidation-collapse and/or emphysema in 50% cases. 30% cases (n = 6) with persistent respiratory tract infection inspite of adequate treatment of recurrent episodes of respiratory distress with wheeze turned out as foreign body inhalation and therefore, such cases need re evaluation. Betelnut was the most common foreign body removed (n = 7) followed by peanut (n = 6). Most frequent site involved was right main bronchus (n = 7) followed by left main bronchus (n = 5). CONCLUSION: Public awareness through mass media needs attention to prevent foreign body inhalation. High index of clinical suspicion is mandatory for early diagnosis and management to prevent fatal outcome and long term morbidity. PMID- 10513435 TI - Carcinoma cervix: a retrospective study. PMID- 10513436 TI - Atherosclerosis and plaque rupture: an update. PMID- 10513437 TI - Association of Hb-D trait with nephrotic syndrome: a fact or coincidence? PMID- 10513438 TI - View box--case 2. Pulmonary cryptococcosis. PMID- 10513439 TI - Dealing with irritable bowel syndrome. PMID- 10513440 TI - Intrauterine growth restriction: a perspective for Pakistan. AB - BACKGROUND: Intrauterine growth restriction (IUGR) is the second leading contributor to the prevailing perinatal mortality and morbidity. It affects 23.8% of newborns around the world and 75% of these affected infants are born in Asia. In Pakistan the incidence of IUGR is around 25%, more than the WHO criteria for triggering a public health action. INTRODUCTION: IUGR is implicated with profound long-term impacts in adult life; like coronary heart disease, NIDDM and abnormal cortisol levels. The effects of the short and long term sequelae are reviewed. ETIOLOGY: IUGR is associated with a wide variety of etiological factors. But the factor unique in its importance to Pakistan is maternal malnutrition. The fetal gene expression is under the influence of nutrition. Growth projection curves showing the interaction between the genetic and environmental factors are discussed. SURVEILLANCE: Identification of IUGR baby in a primary care setting and the options in diagnosis in secondary and tertiary care settings are overviewed. CONCLUSION: The roots of this problem, with multi factorial causes are in the socioeconomic infrastructure. This calls for a synergistic approach of reducing this public health issue. Women empowerment can help us to get out of this intergenerational cycle of growth failure. Availability of resources aside, it is also a matter of political will to change things for the better. PMID- 10513441 TI - Manual therapy cults. PMID- 10513442 TI - The development of hip osteoarthritis: implications for conservative management. AB - Hip osteoarthritis (OA) can be the result of a multitude of causes. There is some evidence to suggest that a dysfunction in the neuromuscular system may contribute to the development of hip OA. This article reviews the forces and loads at the hip joint and the different presentations of hip OA in order to present a hypothesis that the neuromuscular system can play a role in the development of hip OA. Changes in the neuromuscular system include alterations in gait patterns and changes in muscular synergies. An improved understanding of the role these factors may play in the development of hip OA provides a perspective to explore treatment rationales and to provide more effective conservative management. PMID- 10513443 TI - Assessment and treatment of hip osteoarthritis. PMID- 10513444 TI - Plinth padding confounds measures of posteroanterior spinal stiffness. AB - This study investigated whether the presence of plinth padding influences measured posteroanterior spinal stiffness. Two measures of posteroanterior stiffness, the slope of the loading curve (K) and the displacement at 30 N (D30) were made at three vertebral levels: L3, T12 and T6, on a rigid and a padded plinth in 24 asymptomatic subjects. Analysis of variance demonstrated a significant reduction in K and increase in D30 when measured on the padded compared to the rigid surface and an interaction effect for both the K and D30 measures, indicating that the effect of the padding depends upon the vertebral level tested. The correlations between the padded and unpadded stiffness measures ranged from 0.70 to 0.87. The data from this study suggest that the type of plinth surface needs to be standardised when evaluating PA stiffness. PMID- 10513445 TI - Manipulative physiotherapists can reliably palpate nominated lumbar spinal levels. AB - Palpating a nominated spinal level is a prerequisite to more complex tasks such as palpating the level most likely to be the source of the patient's symptoms. The aim of this study was to investigate the reliability of physiotherapists with a post-graduate qualification in manipulation (manipulative physiotherapists) in palpating the lumbar spines of patients in a clinical setting. Three pairs of manipulative physiotherapists palpated the randomly-nominated lumbar spinal levels of 20 patients presenting to their practices for treatment of low-back pain. Each therapist marked the skin overlying the spinous process of the nominated spinal level with an ultraviolet pen and these marks were transcribed onto transparencies for analysis. The therapists obtained an overall weighted kappa of 0.92 indicating almost perfect agreement for locating the nominated spinal level. The results of this study indicate that manipulative physiotherapists can reliably palpate nominated lumbar spinal levels, suggesting further training in spinal therapy enhances the palpatory skills of physiotherapists in palpating nominated lumbar spinal levels. PMID- 10513446 TI - Comparison of ribcage and posteroanterior thoracic spine stiffness: an investigation of the normal response. AB - Evaluation of the movement response to posteroanterior (PA) loads applied to the spinous processes is a recognized part of the physical examination of the thoracic spine. During this clinical procedure the thoracic spine is supported by the ribcage which may contribute to the movement response. However, the contribution of ribcage stiffness to the PA stiffness of the thoracic spine has not been established. The purpose of this study was to measure the PA stiffness of the thoracic spine and compare this to the stiffness of the ribcage under anteroposterior load. Using force-displacement analysis, this study measured the PA stiffness of the thoracic spine at T4, T7 and T10 in 20 asymptomatic individuals, and compared this to the ribcage stiffness measured through sternal compression. The mean PA stiffness at T7 (10.7 N/mm) was significantly greater than at T4 (9.1 N/mm, P < 0.001), and there was a non-significant increase between T7 and T10 (11.4 N/mm, P = 0.08). The stiffness of the ribcage measured via sternal compression (7.6 N/mm) was significantly lower than the thoracic PA stiffness at all levels (P < 0.01). A significant proportion (33%) of the thoracic spine PA stiffness was accounted for by the stiffness of the ribcage (P < 0.01). In young, asymptomatic subjects, the PA stiffness of the thoracic spine is significantly greater than the stiffness of the ribcage. This suggests that the response to PA load application in the thoracic spine can be attributed to factors relating to the spine as well as the ribcage. Defining consistent patterns of PA stiffness in the thoracic spine may assist in the interpretation of clinical measurements of patients with mechanical dysfunction of the thoracic spine. PMID- 10513447 TI - The case of a non-traumatic sprained ankle. PMID- 10513448 TI - Pain memory. PMID- 10513449 TI - [Anesthesia ambulatory one-day surgery]. AB - The spreading of the cost-benefit attitude is a considerable help in the progress of the one day surgery. The patient selection, the preoperative patient preparations and the preoperative examination has done in the anaestesiologic ambulance. Aims of ambulatory anaesthesia are to achieve sedation, hypnosis, analgesia, amnesia and muscle relaxation during the operation, to preserve preoperative mental and physiologic state, analgesia and to make early postoperative nourishing possible. Besides personnel and equipment of anaesthesia and reanimation, monitoring of circulation, respiration and neuromuscular transmission is needed. Anaesthetic methods: local, regional and general anaesthesia or sedation. Ways of general anaesthesia are intravenous, inhalation or combined. Intravenous anaesthetic drugs (barbiturates, ketamine, etomidate, midazolam, propofol and eltanolon) can be used in monotherapy or in combination with each other or opioids (morphine, alfentanil, fentanyl, sufentanil, remifentanil). Among inhalatic agents N2O isoflurane, desflurane, sevoflurane are advisable. Recommended non depolarising muscle relaxants are the short-acting atracurium, mivacurium, vecuronium and rocuronium. Methods for loco-regional anaesthesia are infiltration, peripheral nerve blockade, epidural and intradural anaesthesia which can be used with additional vigil sedation. Blockades with local analgetics, intraoperative opioids, non-steroid anti-inflammatory drugs, sedatives, pre-emptiv analgesia and patient controlled analgesia can be used for postoperative pain relief. Besides the patient and intervention type selection the adequate perioperative anaesthesiologic work and the prudent specifications of leaving conditions is the most important terms of the safety of one-day surgery and anaesthesia. PMID- 10513450 TI - [Hysteroscopic diagnosis and treatment of endometrial polyps]. AB - The aim of the retrospective study was to analyse the frequency of polyps, anamnestic data and the result of histological examination overviewing 163 polyps which was find during 1900 hysteroscopy. In the history of patients dilatation and curettage was find in 22% one time and in 6.6% two times, but the polyps were removed by hysteroscopy in the end. The indication of hysteroscopy was haemorrhage in 55%, abnormal ultrasonographic results in 25% and sterility in 15%. During 153 polypectomy were two perforations (0.89%), no other major complication. The result of histological examination of 153 removed polyps was surprising, because in 22 cases proliferative endometrium, in 17 cases hyperplasia, in 6 cases secretory endometrium, in 5-5 cases fibroid and dyshormonal effect and in 1-1 case endometritis, adenomyosis, atrophy and malignant transformation was the histological diagnosis. Despite of relatively high false positive hysteroscopic results, the endoscopic procedure is useful method for treatment of endometrial polyps if the aim is the minimal invasivity and organ saving surgery. PMID- 10513451 TI - [Percutaneous drainage in the treatment of accidentally discovered hydatid cyst in patients with polycystic liver]. AB - This article report on a not very well-known therapeutical procedure used among only fairly diagnosed cases based on international and Hungarian experiences. In Hungary echinococcus cyst of the liver is mostly treated surgically (the average mortality rate of liver resection is 0-6.3%). In many places the percutan puncture and drainage are contraindicated in case of echinococcus cysts. The authors assert that the percutaneous treatment of echinococcus cysts in proper technical and methodic circumstances in safe regarding the previous studies and their own data obtained of a few cases, and describe the technique and raise some question in them. Drainage treatment is very well tolerated by the patients it reduces the duration (of time) and the cost of the hospitalization. PMID- 10513452 TI - [In vitro findings of antioxidant properties of Hungarian red wines]. AB - In the present study the antioxidant activity of some selected Hungarian red wines produced in traditional wine-growing regions of the country was investigated in different chemical systems. All the samples exhibited strong hydrogen-donating ability, showed significant reducing power and copper-chelating ability. The samples could retard the autooxidation of linoleic acid during a 10 days incubation period at 40 degrees C. All the investigated properties depended on the total polyphenol content of the wines. The highest polyphenol content and antioxidant activity was in Pinot Noir and in Merlot from Villany. Results of this research and other literary data indicate that polyphenols in the concentrations in wines consumed frequently, and moderately together with meal can give an antioxidative contribution to the human health. PMID- 10513453 TI - [Three unusual cases of mastocytosis]. AB - A unique opportunity arose to introduce the rare disease called mastocytosis as we had three patients with radically different clinical signs and disease progression. The authors would like to draw attention to the diagnostic problems that may emerge with this disease, as well as the diagnostic procedures are detailed. These problems, however, are dwarfed by the therapeutic difficulties faced by the clinicians. Complete remission with the present treatment opportunities may not be achieved, nevertheless, there are options to improve quality of life and to alleviate the symptoms that cause suffering to patients. PMID- 10513454 TI - Treatment of depression--newer pharmacotherapies. AB - OBJECTIVES: Depressive disorders are persistent, recurring illnesses that impose enormous personal suffering on individuals and their families. Major depression alone is estimated as the fourth most important cause of worldwide loss in disability-adjusted life years and is likely to become the second most important within 20 years. A continued quest for more effective treatments has spawned newer antidepressants and herbal treatments, which have contributed to explosive growth in antidepressant prescribing, increasing pharmacy costs, and wider but sometimes confusing choices for clinicians and patients. This evidence report provides a comprehensive evaluation of the benefits and adverse effects of newer pharmacotherapies and herbal treatments for depressive disorders in adults and children. SEARCH STRATEGY: Pertinent literature from 1980 to January 1998 was identified from a specialized registry of controlled trials, meta-analyses, and experts. The registry contained trials addressing depression that had been identified from multiple electronic bibliographic databases, hand searches of journals, and pharmaceutical companies. The search, which yielded 1,277 records, combined terms "depression," "depressive disorder," or "dysthymic disorder" with a list of 32 specific "newer" antidepressant and herbal treatments. SELECTION CRITERIA: Randomized controlled trials were reviewed if they (1) were at least 6 weeks in duration; (2) compared a "newer" antidepressant with another antidepressant (newer or older), placebo, or psychosocial intervention; (3) involved participants with depressive disorders; and (4) had a clinical outcome. Two or more independent reviewers identified 315 trials that met these criteria. DATA COLLECTION AND ANALYSIS: Two persons independently abstracted data from each trial. Data were synthesized descriptively, paying attention to participant and diagnostic descriptors, intervention characteristics, study designs and clinical outcomes. Some data were analyzed quantitatively using an empirical Bayes random effects estimator method. Primary outcomes were response rate, total discontinuation rates (dropouts), and discontinuation rates due to adverse events. Response rates were defined as a 50 percent or greater improvement in symptoms as assessed by a depression symptoms rating scale or a rating of much or very much improved as assessed by a global assessment method. MAIN RESULTS: There were 264 trials that evaluated antidepressants in patients (adults and children) with major depression. Of these, 81 compared newer agents with placebo, 150 newer with older agents, 32 newer agents with newer agents, and 1 newer agent with psychotherapy. There were 14 trials evaluating hypericum (St. John's wort), 27 trials each in primary care patients and older adults, 10 trials limited to patients with specific concomitant illnesses, 9 trials in patients with dysthymia, 3 trials each in patients with mixed anxiety depression and subsyndromal depression, 2 trials in adolescents, and 1 in the postpartum setting. Most trials were conducted in outpatients and examined only acute phase treatment of less than 12 weeks' duration. Newer antidepressants were more effective than placebo in treating major depression (risk ratio 1.6, 95% CI 1.5 to 1.7) and dysthymia (risk ratio 1.7, 95% CI 1.3 to 2.3). They were effective among older adults and in primary care patients. In general, there were no significant differences in efficacy among individual newer agents or between newer and older agents. Hypericum (St. John's wort) was more effective than placebo in treating mild to moderately severe depressive disorders (risk ratio 1.9, 95% CI 1.2 to 2.8). Whether hypericum (St. John's wort) is as effective as standard antidepressant agents given in adequate doses was not established. No significant differences were found between newer and older antidepressants in overall discontinuation rates. Selective serotonin reuptake inhibitors (SSRIs), reversi PMID- 10513455 TI - High frequency of CYP2D6 poor and "intermediate" metabolizers in black populations: a review and preliminary data. AB - There is little and conflicting information concerning polymorphism of CYP2D6 in populations of Africans and African descent. Estimations of the prevalence of poor metabolizers (PMs) in Black populations have ranged from 0 to 19 percent, and unlike Caucasian and Asian populations, there seems to be a poor correlation in metabolic ratios (MRs) between commonly used CYP2D6 probe drugs. A novel mutant allele, CYP2D6*17, which is associated with reduced metabolic rates, has been determined to occur in high frequencies in African and African American populations. In the present pilot study, there was a high frequency of CYP2D6*17, and about one-third of the African-American participants showed a reduced capacity to metabolize dextromethorphan, a CYP2D6 probe drug. The CYP2D6*17 allele and other variants may possibly play a role in the inconsistent variation of phenotypes in Black populations. PMID- 10513456 TI - A strategy to evaluate a covariate by group interaction in an analysis of covariance. AB - Analysis of covariance (ANCOVA) is commonly used in psychiatric research to statistically adjust for illness severity when making group comparisons. A fundamental assumption of the procedure is that the groups' differences in outcome are constant across all levels of baseline severity. This is the classic assumption of equal linear slopes. The plausibility of this assumption is rarely reported in manuscripts that use an ANCOVA. A statistical model is discussed that can be used to test the assumption, and if the assumption is not fulfilled, the model can incorporate a covariate by group interaction. An application of this model, the multiple linear regression approach to analysis of covariance, is provided using data from a large clinical trial for patients with panic disorder. The authors recommend that manuscripts reporting results from an ANCOVA also report the results of the test of the covariate by group interaction and account for the interaction when the slopes are heterogeneous. PMID- 10513457 TI - A bridging study of fananserin in schizophrenic patients. AB - Fananserin is a potential antipsychotic compound with high affinity for both D4 and 5-HT2A receptors, and negligible affinity for D2 receptors. Because the tolerance for antipsychotic compounds often differs between schizophrenic patients and healthy subjects, this bridging study was designed to evaluate the tolerability of fananserin, to define the slow titration maximum tolerated dose in the target population, and to identify the most rapid well-tolerated rate of titration for this compound. Three rates of titration regimens were examined in a total of 26 schizophrenic patients in a parallel group design, following a 3-day placebo washout period from previous antipsychotic medication. The slow rate of titration (reaching the maximum dose of 600 mg/day in 16 days with 100-mg increases every 3 days) was well tolerated, but a rapid titration schedule (reaching 600 mg/day in 8 days with 200-mg increases every 3 days) resulted in hypotension in 3 of 6 patients and termination of the group on Day 10. An intermediate rate of titration (reaching 600 mg/day in 10 days with 100-mg increases every 2 days) was then examined and was well tolerated, with transient episodes of mild hypotension reported in 2 of 10 patients. Thus, although hypotension was identified as the dose-limiting adverse event in this study, a reduction in the titration rate was effective in reducing the incidence and severity of this side effect. In this study, schizophrenic patients administered multiple doses of fananserin tolerated doses 400 percent greater than the maximum tolerated single dose in healthy volunteers. PMID- 10513458 TI - Olanzapine therapy in elderly patients with schizophrenia. AB - Compared to young adults, elderly individuals with schizophrenia may have a six fold increase in the prevalence of tardive dyskinesia. The atypical antipsychotic, olanzapine, may offer particular benefit for this population. This is a prospective, open-label trial of olanzapine therapy in elderly schizophrenic patients. Individuals aged 65 years or older with DSM-IV schizophrenia and a history of neuroleptic responsiveness were given olanzapine as an add-on therapy to their existing medication regimen. Other antipsychotic medication was gradually discontinued. Psychopathology was assessed using the Brief Psychiatric Rating Scale (BPRS). Abnormal movements were assessed with the Simpson-Angus Neurological Rating Scale (SA), the Barnes Akathisia Scale (BA), and the Abnormal Involuntary Movement Scale (AIMS). Cognitive status was assessed with the Mini Mental State Evaluation (MMSE). Twenty-seven individuals received a mean dosage of 8.4 (+/- 4.2) mg/day. Mean age of the group was 70.6 (+/- 4.1) with a range of 65 to 80 years. Patients had a mean of 1.6 (+/- 1.4) significant comorbid medical illnesses. Change in BPRS scores were not significant for the group as a whole, whereas SA score change was substantial, with a pre-treatment mean of 13.7 (+/- 10.3), compared with a mean of 4.8 (+/- 4.1) for those treated with olanzapine (p < .0002). Changes in AIMS and BA score were also significant on olanzapine therapy. MMSE score change was not statistically significant. Comorbid medical illnesses were not adversely affected. Olanzapine is an effective antipsychotic medication in older adults with schizophrenia, and is associated with significant improvement in extrapyramidal side effects. Implications for effect on cognitive status should be explored in larger, long-term trials. PMID- 10513459 TI - Evaluation of individual subjects in the analog classroom setting: I. Examples of graphical and statistical procedures for within-subject ranking of responses to different delivery patterns of methylphenidate. AB - In this article, we describe graphical and statistical methods developed to evaluate the response patterns of individual children with attention deficit hyperactivity disorder (ADHD) to different conditions of treatment with stimulant medication. We used data from an investigation of drug delivery patterns to demonstrate these methods. Thirty-one children with ADHD participated in a double blind crossover study of four conditions (three patterns of delivery of methylphenidate and a placebo control). In each condition, the children were evaluated across an 11-hour (7:00 a.m. to 6:00 p.m.) laboratory school day, and ratings of classroom behavior were obtained at regular intervals across the day. Graphical procedures were developed to display, for each individual, time courses of multiple measures of behavior taken across each double-blind test day. Expert clinicians judged these graphs and used this information to rank-order the test days from best to worst. A within-subject variant of Kendall's W was used to evaluate, for each subject, whether the rankings of these multidimensional graphs were reliable (concordant) across judges. A generalized kappa statistic was used to evaluate, for each condition, the reliability of the judges' rankings across subjects. Friedman's analysis of variance of ranks was used to evaluate, for the study, whether the conditions differed in terms of the average (consensus) rank assigned by the judges. PMID- 10513460 TI - Evaluation of individual subjects in the analog classroom setting: II. Effects of dose of amphetamine (Adderall). AB - Multiple dependent variables were graphed for 29 subjects who participated in a double-blind evaluation of 4 doses of Adderall, plus positive (methylphenidate) and placebo control conditions. Five judges ranked the conditions for each subject, and analyses of individual subjects indicated that these rankings were concordant (reliable) across judges. Consensus rankings were assigned to each subject, and an analysis of these ranks showed that the conditions differed significantly. The choice of best conditions were judged to be across 3 doses of Adderall (10, 15, and 20 mg). This confirms the clinical impression of individual differences in optimal dose of stimulant medication. The methodological, graphical, and statistical methods presented in this article provide a systematic, reliable procedure for evaluating relative response of individuals to different doses of stimulant medication. PMID- 10513461 TI - [Manual therapy in the rehabilitation of patients with ischemic heart disease in the early period following myocardial revascularization]. AB - Therapeutic complexes eliminating anginal and nonanginal pain syndromes including physiotherapy and manual therapy have been developed and tried in 57 patients with coronary heart disease early after surgical treatment. The addition of manual therapy in the rehabilitation complex shortens duration of rehabilitation, corrects post-operative angina and reflex pain syndrome in the chest and shoulders. PMID- 10513462 TI - [Impulse barotraining in the regimen of daily atmospheric pressure fluctuation in the treatment of the meteoropathic reactions of patients with ischemic heart disease]. PMID- 10513463 TI - [The therapeutic effects of a dry sodium chloride aerosol in bronchial asthma patients]. PMID- 10513464 TI - [Low-intensity infrared laser radiation in the treatment of bronchial asthma patients]. PMID- 10513465 TI - [The hemodynamic effects of transcerebral electro- and electromagnetotherapy in stroke patients]. AB - The results of a 10-year prospective trial of physiotherapeutic factors in 350 convalescents after cerebral stroke are reviewed. Nonspecific aspects of mechanisms of therapeutic action of transcerebral physiotherapy (development of collateral circulation and cerebral hemodynamic reserve) and specific ones seen primarily in extracranial parts of major head arteries are shown. PMID- 10513466 TI - [The effect of laser radiation on the blood somatomedin activity in patients with spinal osteochondrosis]. PMID- 10513467 TI - [The use of laser radiation and sinusoidal modulated currents in the therapy of patients with chronic calculous pyelonephritis]. AB - The authors present a technique of treating chronic calculous pyelonephritis with laser radiation and sinusoidal modulated currents which promotes a complete elimination of the calculus fragments in 100, 70 and 50% of the patients in the stone size 0.2-0.5 cm, 0.5-0.7 cm and > 7 cm, respectively. This combined therapy had also antiinflammatory, and immunity-stimulating effects. PMID- 10513468 TI - [An efficacy study of the treatment of patients with chronic pyelonephritis and urolithiasis using sulfate-bicarbonate calcium-magnesium mineral water]. AB - Clinical effects of spa treatment on renal function in middle-aged and elderly male and female patients with chronic pyelonephritis and urolithiasis was studied. Combined sanatorium treatment included a course intake of low-mineral sulphate-hydrocarbonate calcium-magnesium mineral water Kazanskaia. Diuresis, especially daytime, was activated in all the patients. Maximum diuresis was observed in cool seasons in the elderly patients. To the end of the treatment proteinuria, oxaluria and uraturia diminished. A course of drinking mineral water Kazanskaia proved effective and is recommended for patients with chronic pyelonephritis and urolithiasis. PMID- 10513469 TI - [The blood antioxidant system under the influence of biocontrolled laser and antioxidant therapy in patients with urethral strictures]. AB - Superoxide dismutase activity in blood red cells of patients with urethral strictures reflects the disease severity and the response to relevant treatment. Antioxidant therapy with dibunol on the first postoperative days, administration of vitamin E before and after the operation and laser biocontrolled chronotherapy for 2 weeks before and after the operation with repetition in 1-2 months are recommended in addition to conventional treatment to reduce probability of complications and recurrences after surgical treatment of urethral strictures. PMID- 10513470 TI - [An experimental validation of the use of the ultraphonophoresis of the mud preparation Eplir in inflammation of the uterine adnexa]. AB - Eplir ultraphonophoresis started on day 5 of experimental inflammation of the uterine appendages in the presence of antibacterial therapy reduces the severity of exudative processes, hemodynamic disorders, leukocytic infiltration, follicular atresia, prevents growth of connective tissue stroma. PMID- 10513471 TI - [The physiological validation of the use of iodinol in physiotherapy]. AB - Experiments on 43 thyroidectomized Wistar male rats with experimental hypothyroidism have shown a favourable effect of iodinole electrophoresis but not galvanization on the disease. The effect was assessed by organ weight and intraorgan water concentration. PMID- 10513472 TI - [The status of and outlook for the creation of Russian physiotherapeutic equipment]. PMID- 10513473 TI - [The development of bromine sodium chloride brines in the Moscow area]. PMID- 10513474 TI - [The use of a complex of biological indices in the balneological assessment of peloids]. PMID- 10513475 TI - [The organizational problems of the physiotherapy service and of sanatorium health resort care]. PMID- 10513476 TI - [The structure of erythrocyte membranes in patients with chronic obstructive bronchitis and its alteration during EHF therapy]. PMID- 10513477 TI - [The efficacy of the physical rehabilitation of patients with spinal osteochondrosis and scoliosis]. PMID- 10513478 TI - [The clinical effect of low-intensity laser radiation as a result of the development of body adaptation]. PMID- 10513479 TI - [Apparatus and devices for local cryotherapy in joint diseases]. PMID- 10513480 TI - [In Process Citation] PMID- 10513481 TI - [In Process Citation] PMID- 10513482 TI - Removal of low concentrations of carbon tetrachloride in compost-based biofilters operated under methanogenic conditions. AB - Research was performed to demonstrate the removal of carbon tetrachloride (CT) using compost biofilters operated under methanogenic conditions. Biofilters were operated at an empty-bed residence time of 2.8 minutes using nitrogen as the atmosphere. Hydrogen and carbon dioxide were supplied as an electron donor and carbon source, respectively, during acclimation of the bed medium microbes. Once methanogenesis was demonstrated, CT flow to the biofilter was established. Biofilters were operated over a CT concentration range from 20 to 700 ppbv for 6 months. Bed medium microbes were able to remove up to 75% of the inlet CT. At excessively high CT concentrations (> 500 ppmv), methane production and hydrogen utilization by the bed medium microbes appeared to be inhibited. CT removal by the biofilter decreased when the hydrogen supply was removed from the biofilter inlet, indicating that hydrogen acted as the electron donor for reductive dechlorination. The removal efficiency and relatively low empty bed residence times demonstrated by these laboratory-scale biofilters indicate that anaerobic biofiltration of CT may be a feasible full-scale process. PMID- 10513483 TI - Whiplash-associated disorder: WAD is it? PMID- 10513484 TI - The relationship of anxiety and depression with self-reported knee pain in the community: data from the Baltimore Longitudinal Study of Aging. AB - OBJECTIVE: To investigate the relationship between anxiety and depression and reporting of knee pain in the community. METHODS: Subjects (n = 374) were community volunteers aged 40 years and above who are participants in the Baltimore Longitudinal Study of Aging, a prospective multidisciplinary research study of normative aging. Knee pain was defined by the First National Health and Nutrition Examination Survey question "have you ever had pain in or around your knee on most days for at least one month?"; anxiety and depression were measured by the relevant subscales of the Arthritis Impact Measurement Scales questionnaire. All subjects had standing anteroposterior radiographs, read for Kellgren and Lawrence (K + L) grade. RESULTS: After adjustment for age, women reporting "ever" knee pain had significantly higher anxiety scores than those reporting "never" pain (3.06 +/- 0.26 versus 2.35 +/- 0.17; P = 0.025). Pain reporting was related neither to anxiety scores in men, nor to depression in either sex. Analysis stratified by radiographic severity, adjusted for age and gender, showed that differences in anxiety were confined to those reporting knee pain in the absence of radiographic change (i.e., K + L grade 0). CONCLUSIONS: In the community, women reporting knee pain in the absence of radiographic osteoarthritis have higher anxiety scores than those without pain. Depression was not significantly related to knee pain in this population. Psychosocial factors may explain some of the discrepancy between reported knee pain and structural change as seen on x-ray. PMID- 10513485 TI - Determinants of hip and knee flexion range: results from the San Antonio Longitudinal Study of Aging. AB - OBJECTIVE: We analyzed data from the San Antonio Longitudinal Study of Aging, a neighborhood-based study of community-dwelling elderly people, to identify factors that determine the flexion range (FR) of hips and knees. METHODS: The FR of hips and knees was measured in a cohort of 687 subjects aged 65 to 79 years. We used multivariate models to examine the associations among the FR of hips and knees, and between these and age, gender, ethnicity, body mass index (BMI), pain and its location, self-reported arthritis, and diabetes mellitus. The functional relevance of hip and knee FR was tested by measuring its association with 50-foot walking velocity. RESULTS: More than 90 degrees of flexion in both hips and both knees was observed in 619 subjects (90.1%). Correlations among the FR of hips and knees ranged from 0.54 to 0.80 (P < 0.001 for Spearman r values). Multivariate analysis revealed a pattern of significant associations between each of the joints and its contralateral mate and ipsilateral partner joints that was consistent for both hips and both knees. Using each individual joint as the unit of analysis, the following variables were independently associated with hip or knee FR in multivariate models: rising BMI and female sex with reduced FR of both hips and knees, a Mexican American ethnic background with decreased hip FR, and knee pain with decreased knee FR. The functional importance of the FR of these two important joints was supported by its significant association with walking velocity in a model that adjusted for age, gender, ethnic background, BMI, and hip or knee pain. CONCLUSIONS: Most community-dwelling elderly people have a FR of hips and knees that can be considered functional. The ipsilateral and contralateral hip or knee are significant independent determinants of the FR of each of these joints. Obesity, a health problem potentially amenable to preventive and therapeutic interventions, is a factor significantly associated with decreased FR of hips and knees. PMID- 10513486 TI - Internal consistency and validity of an observational method for assessing disability in mobility in patients with osteoarthritis. AB - OBJECTIVE: To establish the internal consistency and validity of an observational method for assessing disability in mobility in patients with osteoarthritis (OA). METHODS: Data were obtained from 198 patients with OA of the hip or knee. Results of the observational method were compared with results of self-report methods (questionnaires) on disability in mobility. RESULTS: Both Cronbach's alpha and Mokken Scale Analysis indicate that the method is internally consistent. Using factor analysis, observed and self-reported disability in mobility were found to be closely associated and could not be differentiated. CONCLUSIONS: The observational method is internally consistent and indeed measures disability in mobility (high convergent validity), but observations and self-report seem to yield largely equivalent information (low divergent validity). This raises questions regarding the simultaneous use of both observational and self-report methods in the assessment of disability in mobility in OA patients. PMID- 10513487 TI - Assessment of nutritional status in patients with rheumatoid arthritis and osteoarthritis undergoing joint replacement surgery. AB - OBJECTIVE: To evaluate pre- and postoperative nutritional status in patients with rheumatoid arthritis (RA) and osteoarthritis (OA). METHODS: Preoperative dietary intake was assessed by a food frequency questionnaire, and postoperative dietary intake by food records. Anthropometric and laboratory measurements were assessed 1 day before and 10 days after surgery. Disease activity and acute response to surgery were assessed by erythrocyte sedimentation rate and G-reactive protein. RESULTS: The dietary intake was similar in the two groups preoperatively. Energy, protein, and fluid intake was significantly higher in the RA group postoperatively. There was a significant reduction in the concentration of hemoglobin, albumin, total protein, and ferritin in the OA group after surgery, whereas only hemoglobin concentration was reduced in the RA group. CONCLUSION: Preoperative nutritional status in the RA group was reduced as compared with preoperative nutritional status in the OA group. However, nutritional status in the RA group was less affected after joint replacement surgery compared with nutritional status in the OA group. PMID- 10513488 TI - A five-year followup of hand function and activities of daily living in rheumatoid arthritis patients. AB - OBJECTIVE: To follow hand function and activity of daily living (ADL) capacity prospectively during a 5-year period in a cohort of outpatients with rheumatoid arthritis (RA). METHODS: Forty-three patients (28 women, 15 men), mean age 53.7 years and mean disease duration 7.5 years, were included. The Grip Ability Test (GAT), grip strength, the Keitel Function Test (KFT), the Health Assessment Questionnaire (HAQ), self-estimated hand function, and pain scales were used. Need of personal assistance in the HAQ components was recorded as ADL dependence. RESULTS: After 5 years, the GAT, the KFT, and 3 HAQ components were significantly worse in women. Improved GAT was the only significant change in men. An additional 12 patients needed personal ADL assistance, bringing the total to 21 patients (49%). CONCLUSIONS: Hand function deteriorated during a 5-year period in female RA patients. Hand disability (GAT) improved in the male RA group, although hand impairment (grip strength, KFT) was unchanged. Over one-fourth of each gender group had developed a new handicap (dependence). PMID- 10513490 TI - Position statement of the American College of Rheumatology regarding referral of children and adolescents to pediatric rheumatologists. Executive Committee of the American College of Rheumatology Pediatric Section. PMID- 10513489 TI - The Fibromyalgia Impact Questionnaire: a useful tool in evaluating patients with post-Lyme disease syndrome. AB - OBJECTIVE: To determine the reliability and validity of a modified version of the Fibromyalgia Impact Questionnaire (FIQ) in evaluating patients with post-Lyme disease syndrome (PLDS). METHODS: In this cross-sectional analysis 13 PLDS, 18 fibromyalgia (FM), and 16 healthy controls (n = 47) completed a modified FIQ containing items to evaluate physical impairment, symptom severity, and global well-being. Comparisons between groups were done using analysis of variance with a significance level set at 0.05. RESULTS: PLDS patients demonstrated statistically significantly greater levels of impairment than controls in physical functioning, FIQ total score, global well-being, joint pain, fatigue, depression, ability to perform activities of daily living, and memory/concentration. FM patients demonstrated a statistically significantly greater level of impairment than the control group in all categories, and the scores were significantly higher than the PLDS group in the measurement of physical impairment, FIQ total score, muscle pain, and joint pain. Overall, the instrument possesses good reliability and validity, although adequacy of this instrument to measure impairment in the male PLDS population needs further elucidation. CONCLUSION: The results of this study suggest that the modified FIQ may be a useful tool in evaluating PLDS patients. The findings suggest that there may be some differences in the etiopathology of the symptoms experienced by PLDS and FM patients. PMID- 10513491 TI - Effects of an eight-week physical conditioning program on disease signs and symptoms in children with chronic arthritis. AB - OBJECTIVE: To investigate the effects of an 8-week, 24-session weight-bearing physical conditioning program on disease signs and symptoms in children with chronic arthritis. METHODS: In a within-subjects, repeated measures design, 25 subjects, ages 8-17 years, with chronic polyarticular juvenile rheumatoid arthritis (JRA), were assessed at study entry, after an 8-week control period, and after an 8-week exercise period, for 1) disease status, based on joint count (JC) and articular severity index (ASI) (sum of scores for joint swelling, pain on motion, tenderness, and limitations of motion); 2) worst pain during the past week, using a 10-cm visual analog scale (VAS), and 3) aerobic endurance, using the 9-minute run-walk test of the Health Related Physical Fitness Test battery. The 60-minute conditioning program included warm-up (10 minutes), low-impact aerobics (25 minutes), strengthening (15 minutes), and cool-down and flexibility exercises (10 minutes). Subjects exercised twice a week at their rheumatology center and once a week at home, using a commercial exercise video-tape supplied by the investigator. RESULTS: Significant improvement was found in the ASI (Friedman analysis of variance [ANOVA]), JC, and 9-minute run-walk test (repeated measures ANOVA) from the pre- to post-exercise tests. Mean VAS pain scores decreased 16% from study entry to the post-exercise test. Statistically significant improvement (reliable change index > 1.96) occurred in 80% of subjects on the ASI and 72% on the JC. CONCLUSION: Children and adolescents with chronic polyarticular JRA can improve their aerobic endurance through participation in weight-bearing physical conditioning programs without disease exacerbation or increased pain, and may achieve decreased joint signs and symptoms through increased physical activity. PMID- 10513492 TI - The utility of trained arthritis patient educators in the evaluation and improvement of musculoskeletal examination skills of physicians in training. AB - OBJECTIVE: To determine the level of examination skills of internal medicine residents and to assess whether an intervention by trained persons with arthritis could have a greater impact on their examination skills than participation in an ambulatory care training experience. METHODS: Twenty-seven residents attended a 6 week ambulatory care rotation that included didactic teaching as well as attendance at an outpatient arthritis clinic with supervision by rheumatologists. Sixteen residents were randomly assigned to have a training encounter with an arthritis educator along with the standard experience in the arthritis clinic, whereas 11 residents received training in the arthritis clinic only. Arthritis educators evaluated the musculoskeletal examination skills of each resident during the first week of the rotation. The 16 residents in the intervention group received instruction on joint examination techniques by the arthritis educator immediately following their evaluation. At the end of the 6-week rotation, the groups were re-evaluated by a different arthritis educator. A group of 21 rheumatologists was also asked to perform a comprehensive musculoskeletal examination on individual arthritis educators. The arthritis educators assessed the examination of the rheumatologists using the same evaluation instrument that was used to assess the residents. RESULTS: Initially, internal medicine residents carried out the musculoskeletal examination poorly (34.2 +/- 0.09% correct, n = 27). By contrast, the rheumatologists carried out a significantly greater amount of the examination correctly (54.5 +/- 0.05%). The musculoskeletal examination skills of the residents who received additional training from an arthritis educator were significantly greater at the end of the rotation than the group who did not receive this intervention (50.5 +/- 0.10% versus 41.9 +/- 0.14% correct, P = 2.15 x 10(-5). CONCLUSION: Internal medicine residents carried out the musculoskeletal examination poorly. However, an intervention by arthritis educators improved the musculoskeletal examination skills of internal medicine residents significantly and more effectively than the standard clinical teaching in a rheumatology outpatient clinic. The impact of the arthritis educator intervention persisted for at least 5 weeks. PMID- 10513493 TI - Development of persistent neurologic symptoms in patients with simple neck sprain. PMID- 10513494 TI - The National Arthritis Action Plan: a public health strategy for a looming epidemic. PMID- 10513495 TI - What constitutes a fibromyalgia expert? PMID- 10513496 TI - The needs of patients with arthritis: the patient's perspective. AB - OBJECTIVE: To identify concerns and learning interests of patients with arthritis. METHODS: A questionnaire was developed, pilot tested, and then used to evaluate 197 patients with arthritis, including osteoarthritis (OA) (n = 41), rheumatoid arthritis (RA) (n = 57), back disease (n = 55), systemic lupus erythematosus (n = 27), and systemic sclerosis (SSc) (n = 17). Twenty concerns and 12 learning interests were rated. Questionnaires were also administered to assess physical disability (Health Assessment Questionnaire), psychological disability (Arthritis Impact Measurement Scales 2), and pain (visual analog scale). Participants addressed accessibility of health services, satisfaction with their physician, psychosocial needs, use of self-help groups, and behavioral strategies used to assist coping. Patients with RA, OA, and back disease, at both a community and a hospital center, were tested to assess whether concerns and learning interests differed based on site of treatment. Analytic methods included analysis of variance, factor analysis, and multiple linear regression. RESULTS: There were no differences in concerns or learning interests based on treatment site. Between diagnostic groups, patients with SSc were more interested in learning about self-help groups. The most frequently reported concern was worsening of the illness. The majority of respondents were interested in learning more about topics that were illness specific. The physician was chosen as the preferred source of information, and the preferred format was in writing. On factor analysis, the 20 concerns were reduced to 5 factors: psychological, coping, medication, social, and financial. Three factors were identified for learning interests: the illness, traditional health management topics, and nontraditional health management topics. Stepwise multiple linear regression revealed predictors for the 5 concern and 3 learning interest factors. The concerns were best predicted by self-reported disease severity, physical disability, and psychological distress, while learning interests were best predicted by self-reported disease severity, pain, and self-help group membership. CONCLUSION: Concerns and learning interests of persons with arthritis did not differ based on the center of treatment or the diagnosis, but can be predicted by the level of pain and simple measures of disability. Better understanding of the relationship between health status and patient-perceived needs will result in improved patient-centered care. PMID- 10513497 TI - Development and validation of a patient satisfaction scale for musculoskeletal care. AB - OBJECTIVE: To test the hypothesis that 3 distinct domains of patient satisfaction with musculoskeletal care--satisfaction with the office environment, provider patient interaction, and treatment outcomes--can be measured reliably and, when considered separately, are more valid indicators of satisfaction than global measures. METHODS: Three hundred ninety-nine outpatients who presented with knee or shoulder pain were enrolled in a prospective cohort study. We measured patient satisfaction with musculoskeletal care by adapting a widely used generic satisfaction survey. RESULTS: Each domain of the scale was internally consistent, with Cronbach's alphas for satisfaction with the office environment, provider patient interaction, and treatment outcome subscales of 0.68, 0.95, and 0.93, respectively. Validity correlations demonstrated the greater specificity of the subscales than global measures for particular aspects of musculoskeletal care. CONCLUSIONS: The musculoskeletal-specific satisfaction scale has excellent reliability and good discriminant validity. From a policy perspective, the distinct subscale structure is critical because problems within each domain may have different remedies. PMID- 10513498 TI - Spouse-assisted coping skills training in the management of knee pain in osteoarthritis: long-term followup results. AB - OBJECTIVE: To evaluate the long-term effects of a spouse-assisted coping skills intervention in patients with osteoarthritis (OA) of the knees, and to evaluate how pre- to posttreatment changes in marital adjustment and self-efficacy relate to long-term improvements in pain, psychological disability, physical disability, pain coping, and pain behavior. METHODS: A followup study was conducted with 88 OA patients who had been randomly assigned to 1 of 3 treatment conditions: 1) spouse-assisted coping skills training (spouse-assisted CST), 2) a conventional CST intervention with no spouse involvement, and 3) an arthritis education spousal support (AE-SS) control condition. To evaluate long-term outcome, comprehensive measures of self-efficacy, marital adjustment, pain, psychological disability, physical disability, pain coping, and pain behavior were collected from these individuals at 6 and 12 months posttreatment. RESULTS: Data analysis revealed that, at 6-month followup, patients in the spouse-assisted CST condition scored higher on measures of coping and self-efficacy than those in the AE-SS control group. At 6-month followup, patients who received CST without spouse involvement showed a significantly higher frequency of coping attempts and reported higher levels of marital adjustment than those in the AE-SS control group. At 12-month followup, patients in the spouse-assisted CST condition had significantly higher overall self-efficacy than those in the AE-SS control condition. In addition, patients in both the spouse-assisted CST and CST only conditions tended to show improvements in physical disability at the 12-month followup. Individual differences in outcome were noted at the 12-month followup. Patients in the spouse-assisted CST condition who reported initial (pre- to posttreatment) increases in marital adjustment had lower levels of psychological disability, physical disability, and pain behavior at 12-month followup. However, for patients in the conventional CST and AE-SS control conditions, increases in marital adjustment occurring over the initial phase of treatment were related to increases in pain and decreases in scores on the Pain Control in Rational Thinking factor of the Coping Strategies Questionnaire. Finally, patients in the spouse-assisted CST condition who showed pre- to posttreatment increases in self efficacy were more likely to show decreases in pain, psychological disability, and physical disability at 12-month followup. CONCLUSIONS: These findings suggest that spouse-assisted CST can enhance self-efficacy and improve the coping abilities of OA patients in the long term. Individual differences in the long term outcome of spouse-assisted CST were noted, with some patients (those showing increases in marital satisfaction and self-efficacy) showing much better outcomes than others. PMID- 10513499 TI - The role of stress in functional disability among women with systemic lupus erythematosus: a prospective study. AB - OBJECTIVE: In the last decade, the biopsychosocial approach has been applied to systemic lupus erythematosus (SLE) to understand the multiple factors involved in the disease course. This study examined the link between stress and changes in functional disability as assessed by the Stanford Health Assessment Questionnaire (HAQ) in women with SLE. METHODS: Forty-two women with SLE were assessed at baseline and 8 months later. Major stress (Life Events), minor stressors (Hassles), depression (Beck Depression Inventory), disease activity (Systemic Lupus Activity Measure), and functional disability were collected at both time points, while demographic and disease damage variables (Systemic Lupus International Collaborating Clinics/American College of Rheumatology Damage Index) were collected once at baseline. RESULTS: Mean HAQ scores at baseline (0.52) and followup (0.46) indicated mild disability and remained fairly stable, although individual variation was observed (mean change -0.07; range -1.25-0.5). Demographic (age, education) and disease (duration, activity, damage) variables were not related to 8-month changes on the HAQ. Of the baseline stress measures, greater negative life events in the preceding 6 months was correlated with reduced functional ability (r = 0.42) 8 months later. Individual changes in depressed mood over the 8-month period were correlated (r = 0.33) with changes in functional ability. Hierarchical multiple regression revealed that after controlling for baseline HAQ scores and changes in depressed mood, baseline negative life events remained a significant predictor of changes in functional ability. CONCLUSION: We found that the major short-term determinants of functional disability were not demographic- or disease-related factors, but rather stress caused by negative life events. Comprehensive treatment of SLE requires management of life stress. PMID- 10513500 TI - A randomized, controlled trial of exercise and education for individuals with fibromyalgia. AB - OBJECTIVE: To evaluate the efficacy of a 6-week exercise and educational program for patients with fibromyalgia. METHODS: Forty-one subjects were randomly assigned to the program or served as waiting list controls. Program outcome was assessed with a 6-minute walk test, the Fibromyalgia Impact Questionnaire, a Self Efficacy Scale, and a "knowledge" questionnaire (based on information provided during the educational sessions). Waiting list control subjects subsequently completed the program. Program outcome was reassessed 3 or 6 months post-program. RESULTS: The program produced significant improvements in 6-minute walk distance, well-being, fatigue, self-efficacy (for controlling pain and other symptoms), and knowledge. At followup, immediate gains in walk distance, well-being, and self efficacy were maintained, but gains in fatigue and knowledge were lost. CONCLUSION: Short-term exercise and educational programs can produce immediate and sustained benefits for patients with fibromyalgia. The benefits of our program may be due to exercise or education since both interventions were given. PMID- 10513501 TI - On the nature of rheumatism. PMID- 10513502 TI - Joint deformity and dysfunction: a basic review of underlying mechanisms. AB - Five common examples of joint deformity or dysfunction were presented. In each case, abnormal biomechanics were caused by or associated with arthritis or arthrosis due to advanced age or overused joints. The principles described for each example can be applied to essentially all synovial joints in the body. Understanding the underlying pathologic and biomechanical mechanisms that cause joint dysfunction can enhance treatment and patient education programs. PMID- 10513503 TI - Why is methotrexate the first selection in disease-modifying antirheumatic drug therapy? Comment on the article by Wolfe et al. PMID- 10513504 TI - Systemic lupus erythematosus, hypercoagulable states, and antiphospholipid antibodies. PMID- 10513505 TI - Quality of life in ankylosing spondylitis: validation of the ankylosing spondylitis Arthritis Impact Measurement Scales 2, a modified Arthritis Impact Measurement Scales Questionnaire. AB - OBJECTIVE: To develop a questionnaire measuring quality of life in ankylosing spondylitis (AS). METHODS: A focus group technique contributed to the generation of an AS-specific questionnaire using the Arthritis Impact Measurement Scales 2 (AIMS2) (12 dimensions), supplemented with items from the Health Assessment Questionnaire for the Spondylar-thropathies, as well as original new items. Patients from a self-help group provided data to assess the validity (n = 146) and reproducibility (n = 43) of the AS-AIMS2. RESULTS: The focus group generated a thirteenth dimension including 6 items labeled "spine mobility." Principal component analyses on AS-AIMS2 showed satisfactory dimensionality (74.1% of variance explained), high internal consistency (Cronbach's alpha = 0.78-0.91), significant convergent validity of the new dimension with spine pain and mobility, and good reproducibility in the 13 dimensions and 5 components (intraclass correlation coefficient = 0.7-0.9). The scores in each component of the AS-AIMS2 showed a marked deterioration of quality of life in these young patients. CONCLUSIONS: The disease-specific AS-AIMS2 questionnaire should help improve care management in AS. PMID- 10513506 TI - Short-form Arthritis Impact Measurement Scales 2: tests of reliability and validity among patients with osteoarthritis. AB - OBJECTIVE: To validate a short-form Arthritis Impact Measurement Scales 2 (AIMS2 SF) among 147 patients with osteoarthritis (OA). METHODS: We used factor analysis to identify domains of functional health associated with OA. Multitrait scaling analysis was used to evaluate the reliability and validity of the AIMS2-SF. RESULTS: The results suggested that upper body limitations should be distinguished from lower body limitations in the physical function scale. The AIMS2-SF was psychometrically sound, with all 5 scales having high item discriminant validity and Cronbach's alpha reliability above the 0.70 criterion (except 0.67 for the social function scale). CONCLUSION: The AIMS2-SF is a reliable and valid instrument among patients with OA. Because of its simplicity and ease of application, it may be useful in routine evaluation of health status, in clinical research, and in predicting use of medical services among OA patients. PMID- 10513507 TI - Comparison of the responsiveness and relative effect size of the western Ontario and McMaster Universities Osteoarthritis Index and the short-form Medical Outcomes Study Survey in a randomized, clinical trial of osteoarthritis patients. AB - OBJECTIVE: This study compares the responsiveness and relative effect sizes of the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) with the Medical Outcomes Study Short Form Health Survey (SF-36) in a randomized clinical trial for treatment of osteoarthritis (OA). METHODS: Patients with OA of the knee or hip were randomized to receive either placebo or 2,400 mg/day of ibuprofen for 28 days. Patients completed the WOMAC and SF-36 at baseline and days 7, 14, and 28 of the trial. RESULTS: Patients receiving ibuprofen showed significant improvement in WOMAC pain, physical functioning, and the total score, while improvement was detected only for bodily pain on the SF-36. The WOMAC detected significant differences between ibuprofen and placebo for pain and physical functioning, whereas the SF-36 detected differences for the bodily pain subscale. CONCLUSION: These results suggest the WOMAC has greater power to detect treatment differences than the SF-36, with respect to pain and physical functioning, in OA clinical trials. PMID- 10513508 TI - Rheumatologist-patient communication about exercise and physical therapy in the management of rheumatoid arthritis. AB - OBJECTIVE: Little is known about the features and role of exercise discussions between rheumatologists and patients. The goals of this study were to: 1) describe rheumatologists' and patients' attitudes and beliefs regarding exercise and physical therapy for rheumatoid arthritis (RA); 2) describe frequency and length of exercise discussions; 3) determine the accuracy of recall for exercise discussions; and 4) assess the influence of attitudes regarding exercise on communication about exercise. METHODS: Goals 1-3 were addressed with analysis of baseline questionnaires and audiotaped encounters. The influence of attitudes and beliefs regarding exercise on the frequency and length of exercise discussions was assessed prospectively. Patients and rheumatologists were enrolled from a large tertiary care institution. Clinical encounters were audiotaped, transcribed, coded, and analyzed to identify specific characteristics of the exercise discussions. RESULTS: One hundred thirty-two patients and 25 rheumatologists participated in the study. Rheumatologists and patients discussed exercise in 53% of the encounters. Rheumatologists' beliefs regarding the usefulness of exercise for RA varied, with the least positive beliefs being reported for aerobic exercise. Exercise discussions were more likely to occur if the patient was currently exercising, odds ratio (OR) = 2.4; 95% confidence interval (CI) (1.2-4.9), and when the rheumatologist believed aerobic exercises were useful in managing RA, OR = 1.4; 95% CI (1.1-1.9). Current exercise behavior was associated with patients' positive attitude toward exercise (chi 2 1 = 8.4; P = 0.004) and perceived social support for exercise (chi 2 1 = 4.5; P = 0.04). When rheumatologists initiated exercise discussions, there was nearly twice as much discussion (beta = -8.4; P = 0.001). CONCLUSIONS: Exercise talk was influenced by patients' and rheumatologists' beliefs and attitudes regarding the effectiveness of exercise and physical therapy in managing RA, patient experience with exercise, and by characteristics of the rheumatologist. PMID- 10513509 TI - Tests of functional limitations in fibromyalgia syndrome: a reliability study. AB - OBJECTIVE: To evaluate the reliability and discriminative ability of a test battery consisting of 7 tests designed for the assessment of functional limitations in patients with fibromyalgia syndrome (FMS). METHODS: The intrarater reliability of the test battery was evaluated for 15 women with FMS. Interrater reliability was calculated on 4 tests separately. Fifteen healthy women constituted a reference group. RESULTS: The intrarater coefficient of variation was < 8% for the shoulder range of motion tests, chair test, and 6-minute walk test, and < 21% for the shoulder endurance test, with correlation coefficients above 0.80 for all tests. Kappa was 0.70-0.80 for the hand-to-scapula tests. The interrater coefficient of variation was < 5% for shoulder range of motion. The performances of the FMS patients were significantly decreased in comparison with healthy subjects in all the tests except for the hand-to-scapula movement. CONCLUSIONS: All but 1 of the selected 7 tests were considered to possess acceptable intrarater reliability for use in FMS in clinical physical therapy practice. PMID- 10513510 TI - Development of the Joint Protection Behavior Assessment. AB - OBJECTIVE: To develop an observational assessment of the use of joint protection (JP) methods by people with rheumatoid arthritis (RA). METHODS: Subjects with and without RA were videotaped performing a kitchen activity to identify the range of JP and non-JP methods used. Behavior codes were developed for these. Seven rheumatology occupational therapists reviewed and scored behaviors as correct, partially correct, or incorrect JP methods. Test-retest and interrater agreement studies were conducted. RESULTS: The Joint Protection Behavior Assessment (JPBA) demonstrated good content validity (kappa = 0.46 to 1.00), test-retest reliability (P < 0.0001), and interobserver agreement (kappa = 0.68 to 0.88). Construct validity was supported by significant correlations with hand impairment and function variables. CONCLUSION: The JPBA is a reliable and valid assessment of the use of JP methods by people with mild to moderate hand and upper limb joint involvement, which can be used to evaluate the effectiveness of JP education programs. PMID- 10513511 TI - Six-minute walk test: a potential outcome measure for hydrotherapy. PMID- 10513512 TI - Thrombosis in systemic lupus erythematosus: the role of antiphospholipid antibody. PMID- 10513513 TI - Quantitative ultrasound: is it a useful test in osteoporosis? PMID- 10513514 TI - Mixed connective tissue disease, or should it be Sharp's syndrome? Comment on the article by Farhey and Hess. PMID- 10513516 TI - It's getting there. PMID- 10513515 TI - You reap what you sow. PMID- 10513517 TI - The changing face of district nursing services. PMID- 10513518 TI - Reducing the health risks in a diabetic pregnancy. PMID- 10513519 TI - Incontinence: patients do not need to suffer in silence. PMID- 10513520 TI - When good compliance equals good asthma control. PMID- 10513521 TI - Know how. Treating isolated systolic hypertension. PMID- 10513522 TI - Wet wrapping in eczema care. PMID- 10513523 TI - Contraceptive choices: a look at intrauterine devices. PMID- 10513524 TI - An alternative approach to menopausal symptoms. PMID- 10513525 TI - Tuberculosis: a need for a UK-wide vaccination policy? PMID- 10513526 TI - Crown Review: a framework and policy for prescribing. PMID- 10513528 TI - Developing an assessment tool for acute wounds. PMID- 10513527 TI - Wet, sloughy and necrotic wound management. PMID- 10513529 TI - Practice nurses = quality service. PMID- 10513530 TI - A weighty issue. PMID- 10513531 TI - Role of nutrition in promoting healthy bones. PMID- 10513533 TI - A multidisciplinary approach to psoriatic arthropathy. PMID- 10513534 TI - A nurse-led service for leg ulcer care. PMID- 10513532 TI - Psychological adjustment when diabetes is diagnosed. PMID- 10513535 TI - How to dress awkward places. PMID- 10513536 TI - Promoting continence care and creating awareness. PMID- 10513537 TI - Using sexual health clinics to reduce teenage pregnancy. PMID- 10513538 TI - Nursing care of minor burns. PMID- 10513539 TI - Do leg ulcer guidelines work? An evaluation project. PMID- 10513540 TI - How the Drug Tariff works: its role in effective practice. PMID- 10513541 TI - Confronting the devil within. PMID- 10513542 TI - A strategic plan. PMID- 10513543 TI - The Family Circle project. PMID- 10513544 TI - Targeting unmet needs. PMID- 10513545 TI - Tackling the threat of TB. PMID- 10513546 TI - Taking a look at new advances in asthma therapy. PMID- 10513547 TI - Goals in the management of type 2 diabetes. PMID- 10513548 TI - How to take cervical smears. PMID- 10513549 TI - Health initiatives to help men fight the flab. PMID- 10513550 TI - Taking the stress out of travelling with a stoma. PMID- 10513551 TI - Role of poor glycaemic control in foot ulceration. PMID- 10513552 TI - The process of granulation and its role in wound healing. PMID- 10513553 TI - Gamma-vinyl gamma-aminobutyric acid attenuates the synergistic elevations of nucleus accumbens dopamine produced by a cocaine/heroin (speedball) challenge. AB - In the present study, we examined the effect of an acute administration of the selective suicide inhibitor of gamma-aminobutyric acid (GABA)-transaminase, gamma vinyl GABA on increases in nucleus accumbens dopamine produced by a cocaine/heroin challenge in freely moving animals. Cocaine (20 mg/kg, i.p.) produced an elevation in extracellular nucleus accumbens dopamine of approximately 380% above baseline, while heroin produced only a moderate increase of 70%. Coadministration of these two drugs, however, produced a synergistic elevation in nucleus accumbens dopamine of 1000%. This response was reduced by 50% in animals pretreated with gamma-vinyl GABA (300 mg/kg, i.p.) 2.5 h prior to challenge. This same dose of gamma-vinyl GABA inhibited cocaine-induced increases in nucleus accumbens dopamine by 25% and completely abolished heroin-induced increase. These findings indicate that gamma-vinyl GABA can interfere with the synergistic effects produced by the combination of an indirect dopamine releaser (heroin) and a dopamine reuptake blocker (cocaine). PMID- 10513554 TI - Dissociation of inhibitory effects of guanethidine on adrenergic and on purinergic transmission in isolated canine splenic artery. AB - The aim of this study was both to investigate the effects of progressive inhibition of adrenergic neurons by increasing concentrations of guanethidine (0.1-10 microM) on the double-peaked vasoconstrictor responses to electrical periarterial nerve stimulation in the isolated and perfused canine splenic artery, and to clarify whether release of noradrenaline is presynaptically separate from release of adenosine 5'-triphosphate (ATP). Double-peaked vasoconstrictions (biphases of vasoconstrictions) were consistently observed under the conditions of 30-s trains of pulses at 1-10 Hz frequencies. Guanethidine, at a lower concentration (0.1 microM) did not modify the first (1st) phase vasoconstriction at low frequencies (1-2 Hz), but markedly inhibited the second (2nd) responses. On the other hand, it slightly but significantly inhibited the double-peaked vasoconstrictor responses at high frequencies (6-10 Hz). Furthermore, a 10-fold increase of concentration of guanethidine (1 microM) almost completely inhibited the 2nd phase responses at any frequencies used but did not completely inhibit the 1st phase response. A further increased concentration of guanethidine (10 microM) failed to enhance the 1 microM guanethidine-induced inhibition. The 1 microM guanethidine-resistant 1st phase responses at any frequencies used (1-10 Hz) were sensitive to tetrodotoxin (30 nM). Treatment with 0.1 microM prazosin did not modify the 1st phase response at any frequencies used in the 1 microM guanethidine-treated preparation. The responses remaining after 1 microM guanethidine and 0.1 microM prazosin were completely suppressed by a subsequent application of 1 microM alpha,beta methylene ATP at any frequencies used. The results indicated that guanethidine, an adrenergic neuron blocker, may exert a dominant inhibitory effect on adrenergic rather than on purinergic components of sympathetic nerve co transmission, indicating that guanethidine-sensitive mechanisms may mainly contribute to determine noradrenaline secretion from neurosecretory vesicles rather than ATP secretion. PMID- 10513555 TI - Effects of the insurmountable angiotensin AT1 receptor antagonist candesartan and the surmountable antagonist losartan on ischemia/reperfusion injury in rat hearts. AB - Two angiotensin AT1 receptor antagonists with different receptor binding characteristics, candesartan (insurmountable antagonism) and losartan (surmountable antagonism), were compared as regards their effects on angiotensin II-induced vasoconstriction and on myocardial ischemia/reperfusion injury. In isolated rat hearts perfused under constant flow, it was found that at equipotent concentrations candesartan (10 nM) and losartan (3 microM) almost completely inhibited the angiotensin II-induced increase in coronary perfusion pressure. However, if a washout period was introduced before the angiotensin II challenge, the effect of losartan quickly vanished, while that of candesartan remained. In hearts subjected to 25 min of global ischemia and 45 min of reperfusion, pre treatment with candesartan (10 nM) or losartan (3 microM) immediately prior to ischemia improved the recovery of left ventricular developed pressure as compared to the effect of vehicle (69 +/- 3.2 and 64 +/- 2.3 vs. 44 +/- 6.2%, respectively; mean +/- S.E.M, P < 0.05). When ischemia was initiated following 30 min of washout after drug administration, the recovery of left ventricular developed pressure was higher in the candesartan group (73 +/- 3.2%, P < 0.05), but not in the losartan group (63 +/- 2.8%), than in the vehicle group (58 +/- 4.8%). The cumulative creatine kinase release during the first 30 min of reperfusion in the washout experiments was lower in the candesartan group (28.5 +/- 2.30 U, P < 0.05), but not in the losartan (40.8 +/- 6.73 U) group, than in the vehicle group (48.1 +/- 4.35 U). No significant difference between groups in left ventricular end-diastolic pressure and coronary perfusion pressure was found. The present results demonstrate that angiotensin AT1 receptor antagonists at equipotent concentrations could differ in their cardioprotective effects in hearts subjected to ischemia/reperfusion. It is suggested that the insurmountable AT1 receptor characteristics of candesartan could provide more persistent cardioprotection than the surmountable receptor characteristics of losartan. PMID- 10513556 TI - Impaired endothelium-dependent relaxation by adrenomedullin in monocrotaline treated rat arteries. AB - The effects of adrenomedullin were evaluated in isolated vascular rings from rats treated with monocrotaline (60 mg/kg, s.c.) causing pulmonary hypertension and right ventricular hypertrophy within 3 to 4 weeks. Sham animals (NaCl-treated rats) were used for comparison. The relaxing effects of adrenomedullin (10(-8) M) and acetylcholine (10(-6) M) were determined in thoracic aorta and pulmonary artery rings precontracted with phenylephrine (10(-7) M). In sham animals, adrenomedullin caused significant vasorelaxation of aorta and pulmonary artery although of different amplitude (24 +/- 3% and 40 +/- 2%, respectively). A greater relaxation was observed in response to acetylcholine. Monocrotaline treated rats exhibited a reduction in adrenomedullin relaxation in pulmonary artery (54 and 68% loss of effect, at 3 and 4 weeks, respectively, P < 0.01 vs. sham) and comparable reductions in acetylcholine responses. The decrease in adrenomedullin relaxing effect was less pronounced in aorta than in pulmonary artery, suggesting a distinct tissue sensitivity to monocrotaline. In contrast, the relaxing effect of acetylcholine on aorta was decreased at 4 weeks (36% reduction, P < 0.01 vs. sham). In this model, the adrenomedullin-induced relaxation of the pulmonary artery was impaired due to a severe endothelial dysfunction which may contribute partly to the evolving pathophysiological process. PMID- 10513557 TI - Role of calcitonin gene-related peptide and capsaicin-sensitive afferents in central thyrotropin-releasing hormone-induced hepatic hyperemia. AB - The involvement of capsaicin-sensitive afferent neurons and calcitonin gene related peptide (CGRP) in the central thyrotropin-releasing hormone (TRH)-induced hepatic hyperemia was investigated in urethane anesthetized rats. Both systemic capsaicin pretreatment and intravenous administration of CGRP receptor antagonist, human CGRP-(8-37), completely abolished the stimulatory effect of hepatic blood flow induced by intracisternal injection of TRH analog (RX-77368; p Glu-His-(3,3'-dimethyl)-Pro-NH2, 100 ng), assessed by the hydrogen gas clearance method. These data demonstrate the involvement of capsaicin-sensitive afferent neurons and CGRP in the central TRH-induced stimulation of hepatic blood flow. PMID- 10513558 TI - Protective role of nitric oxide synthase against ischemia-reperfusion injury in guinea pig myocardial mitochondria. AB - In guinea-pig myocardial mitochondria preparation, lowering the Ca2+ concentration or pH level in the perfusate rapidly elevated the fura-2 Ca2+ signal ([Ca2+]m). Pretreatment with 10(-4) M L-Arg inhibited the rapid [Ca2+]m influx, whereas administration of 10(-4) M L-NAME did not, suggesting some association between nitric oxide (NO*) synthase (NOS) activation and Ca2+ kinetics in mitochondria. Immunoblotting analysis showed that endothelial (e)-NOS was present in mitochondria, but not inducible (i)-NOS or brain (b)-NOS. Electron microscopy observations revealed that the e-NOS antibody-reactive site in the mitochondria was the inner cristae. The production of reactive oxygen species and NO* in isolated mitochondria was detected by the spin trapping technique with electron paramagnetic resonance (EPR) spectrometry. Pretreatment with 10(-5) M S nitroso-N-acetyl-DL-penicillamine (SNAP) and 10(-5) M 3-[2-Hydroxy-1-(1 methylethyl)-2-nitrosohydrazino]-1-propananin e (NOC 5), which spontaneously generate NO*, completely inhibited the [Ca2+]m uptake. In addition, N-morpholino sydnonimine hydrochloride (SIN-1) (10(-5) M), which simultaneously generates NO* as well as *O2- and peroxynitrite anion (ONOO-), inhibited the increase in [Ca2+]m. ONOO- (3 x 10(-4) M) itself also inhibited this increase. Pretreatment with the *O2(-)-scavenger manganese superoxide dismutase or catalase (200 units/ml) completely inhibited the increase in [Ca2+]m caused by lowering of either the Ca2+ concentration or the pH in the perfusate. These results suggested that the formation of reactive oxygen species promoted the [Ca2+]m influx. The agents that inhibited the [Ca2+]m influx improved contractility even in Langendorff preparations after ischemia. Based on these findings, we concluded that e-NOS exists in mitochondria and that NO* may play an important protective role in reperfusion cardiac injury after ischemia, by inhibiting the Ca2+ influx into mitochondria which are otherwise damaged by *O2-. PMID- 10513559 TI - Antagonism between the metabolic responses induced by epinephrine and piroxicam on isolated rat hepatocytes. AB - Nonsteroidal anti-inflammatory drugs (NSAIDs) are one of the most employed therapeutic agents. They have a wide spectrum of biological effects, some of which are independent of cyclooxygenase inhibition, such as the alterations on the components of signal transduction systems. In particular, previous data from our laboratory suggested an antagonism between epinephrine and piroxicam, one of the most prescribed NSAIDs. Thus, this study deals with the epinephrine-piroxicam antagonism recorded for metabolic responses in isolated rat hepatocytes. The obtained results show that epinephrine stimulates lactate and ethanol consumption, stimulates glucose release from lactate only, and has no effect on cellular triacylglycerides content. Otherwise, in a dose-dependent basis, piroxicam stimulates lactate and ethanol consumption accompanied by an increase in triacylglycerides content, without changes in glucose release by hepatocytes. Piroxicam blocks the epinephrine-induced stimulation of glucose release from lactate, and epinephrine blocks the piroxicam-mediated increase in triacylglycerides content from lactate or ethanol. In contrast, the effects of epinephrine and piroxicam, promoting the consumption of lactate and ethanol, are not antagonized or added after the simultaneous administration of both compounds. This last result is probably related to the ability of both compounds to stimulate oxygen consumption. On isolated rat liver mitochondria, micromolar doses of piroxicam partially uncouple oxidative phosphorylation, and paradoxically stimulates an ATP-dependent mitochondrial function as citrullinogenesis. These results show for first time, on isolated rat hepatocytes, an antagonism between the metabolic responses of epinephrine and piroxicam, at the concentration found in plasma after its therapeutical administration. PMID- 10513560 TI - Valproate and carbamazepine increase prefrontal dopamine release by 5-HT1A receptor activation. AB - The anticonvulsant mood stabilizers valproic acid (250, 500 but not 50 mg/kg) and carbamazepine (6, 12.5 but not 3 mg/kg) were found to increase extracellular dopamine levels in rat medial prefrontal cortex, but not nucleus accumbens. Increased prefrontal dopamine was completely abolished by the selective 5-HT1A receptor antagonist N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-2 pyridinylcyclohexa necarboxamide (WAY100635, 0.05 mg/kg). Anticonvulsants and clozapine may share a common mood stabilizing mechanism since clozapine is reported to have mood stabilizing effects and increase prefrontal dopamine by 5 HT1A receptor activation. PMID- 10513561 TI - 9-(Aminomethyl)-9,10-dihydroanthracene is a novel and unlikely 5-HT2A receptor antagonist. AB - Structural elaboration of phenylethylamine to 9-(aminomethyl)-9,10 dihydroanthracene (AMDA) produces an agent with high affinity (Ki = 9.5-21 nM) at 5-HT2A receptors. It was shown that AMDA acts as a 5-HT2A receptor antagonist. The structure and molecular geometry of AMDA are not consistent with existing pharmacophore models for 5-HT2A receptor antagonist activity. Thus, AMDA may be a structurally novel parent of a new class of 5-HT2A receptor antagonists that binds to the receptor in a unique fashion that is distinct from the binding topology of existing 5-HT2A receptor antagonists. PMID- 10513562 TI - (2S,3R)TMT-L-Tic-OH is a potent inverse agonist at the human delta-opioid receptor. AB - We examined the pharmacologic effect of beta-methyl-2',6'-dimethyltyrosine-L tetrahydroisoquinone-3- carboxylic acid ((2S,3R)TMT-L-Tic-OH) on G protein activation in membranes prepared from Chinese Hamster Ovary cells transfected with cDNA of the human delta-opioid receptor. (2S,3R)TMT-L-Tic-OH inhibited G protein activation to 58% of basal with an EC50 of 0.72 nM as determined by [35S]GTPgammaS binding. These findings suggest that (2S,3R)TMT-L-Tic-OH is a highly potent inverse agonist at the human delta-opioid receptor. PMID- 10513563 TI - Metrifonate improves working but not reference memory performance in a spatial cone field task. AB - The effects of metrifonate (3, 10 and 30 mg/kg, p.o.) on the working and reference memory performance of the rat were assessed in a spatial cone field task. The highest dose of metrifonate (30 mg/kg) improved the working memory performance, whereas none of the doses affected the reference memory performance. Other parameters of spatial discrimination performance were not affected by metrifonate treatment. The present results suggest that metrifonate has cognition enhancing properties which are likely to be related to aspects of (spatial) working memory. PMID- 10513564 TI - Influence of LY 300164, an antagonist of AMPA/kainate receptors, on the anticonvulsant activity of clonazepam. AB - LY 300164 [7-acetyl-5-(4-aminophenyl)-8,9-dihydro-8-methyl-7H-1,3-dioxolo(4, 5H) 2,3-benzodiazepine], an antagonist of AMPA/kainate receptors, at 5 mg/kg exerted a significant anticonvulsant effect, as regards seizure and afterdischarge durations in amygdala-kindled seizures in rats. At lower doses, LY 300164 did not exert anticonvulsant activity. Clonazepam alone (0.003-0.1 mg/kg) significantly diminished seizure severity, seizure and afterdischarge durations. Coadministration of LY 300164 (2 mg/kg) with clonazepam (0.001 mg/kg) resulted in the significant anticonvulsant activity. Seizure severity score, seizure and afterdischarge durations were reduced from 5 to 4, from 32.6 s to 12.3 s, and 42.7 s to 23.2 s. LY 300164 (2 mg/kg), clonazepam (0.001-0.1 mg/kg) and the combination of clonazepam (0.001 mg/kg) with LY 300164 (2 mg/kg) did not affect long-term memory evaluated in the passive avoidance task in rats. LY 300164 (at the subprotective dose of 2 mg/kg) significantly potentiated the anticonvulsant action of clonazepam against maximal electroshock but not against pentylenetetrazol-induced convulsions in mice. The results indicate that blockade of glutamate-mediated events at AMPA/kainate receptors may differently affect the protection offered by clonazepam, which seems dependent upon the model of experimental seizures. PMID- 10513566 TI - Activation of the reticulothalamic cholinergic pathway by the major metabolites of aniracetam. AB - The aim of the study was to further investigate the effects of aniracetam, a cognition enhancer, and its metabolites on the brain cholinergic system. We measured choline acetyltransferase activity and acetylcholine release using in vivo brain microdialysis in stroke-prone spontaneously hypertensive rats (SHRSP). The enzyme activity in the pons-midbrain and hippocampus, and basal acetylcholine release in the nucleus reticularis thalami were lower in SHRSP than in age matched Wistar Kyoto rats, indicating central cholinergic deficits in SHRSP. Repeated treatment of aniracetam (50 mg/kg p.o. x 11 for 6 days) preferentially increased the enzyme activity in the thalamus, whereas decreased it in the striatum. Among the metabolites of aniracetam, local perfusion of N-anisoyl-gamma aminobutyric acid (GABA, 0.1 and/or 1 microM) and p-anisic acid (1 microM) into the nucleus reticularis thalami, dorsal hippocampus and prefrontal cortex of SHRSP produced a significant but delayed increase of acetylcholine release. We failed, however, to find any effect of aniracetam itself. A direct injection of N anisoyl-GABA (1 nmol) into the pedunculopontine tegmental nucleus of SHRSP enhanced the release in the nucleus reticularis thalami. Thus, these data prove that aniracetam can facilitate central cholinergic neurotransmission via both metabolites. Based on its pharmacokinetic profile, N-anisoyl-GABA may contribute to the clinical effects of aniracetam, mainly by acting on the reticulothalamic cholinergic pathway. PMID- 10513565 TI - Antinociceptive properties of FR140423 mediated through spinal delta-, but not mu and kappa-, opioid receptors. AB - We investigated the antinociceptive effect of FR140423, 3-(difluoromethyl)-1-(4 methoxyphenyl)-5-[4-(methylsulfinyl)phenyl] pyrazole, in the tail-pinch test in mice, and evaluated the mechanism of action using various opioid receptor antagonists. P.o. and i.t. injection of FR140423 exerted dose-dependent antinociceptive activities with ED50 values of 21 mg/kg and 3.1 microg/mouse, respectively. However, i.c.v. injection of FR140423 did not show an antinociceptive effect. The antinociceptive effects of FR140423 were completely abolished by naloxone and naltrindole but not by naloxonazine, beta funaltrexamine and nor-binaltorphimine. FR140423 did not affect any opioid receptor binding in mouse spinal membranes at concentrations up to 100 microM in vitro. Naloxone-induced jumping and diarrhea tests for morphine-like physical dependence of FR140423 gave negative results. These results suggest that FR140423 can induce antinociception by acting on the spinal but not the supraspinal site, and that spinal delta-opioid systems indirectly play a role in the antinociception produced by FR140423 in mice. PMID- 10513567 TI - NMDA receptor antagonism, but not AMPA receptor antagonism attenuates induced ischaemic tolerance in the gerbil hippocampus. AB - Recent studies have shown that a brief 'pre-conditioning' ischaemic insult reduces the hippocampal cell death caused by a subsequent more severe test insult. In the present studies, we have examined the effects of the non competitive NMDA receptor antagonist ((5R,10S)-(+)-5-methyl-10,11-dihydro-5H dibenzo[a,d]cyclohepten-5, 10-imine, MK-801) a competitive NMDA receptor antagonist, LY202157, AMPA receptor antagonist ((3S,4aR,6R,8aR)-6-[2-(1(2)H tetrazole-5-yl)]decahydroiso quinoline-3-carboxylic acid, LY293558), a non competitive AMPA receptor antagonist ((-)-1-(4-amino-phenyl)-4-methyl-7,8 methylenedioxy-4,5-dihydro-3-acetyl -2,3-benzodiazepine, LY300164), and a mixed NMDA/AMPA receptor antagonist, LY246492, in a gerbil model of ischaemic tolerance. Ischaemic tolerance was induced by subjecting gerbils to a 2-min 'pre conditioning' ischaemia (bilateral carotid occlusion) 2 days prior to a 3-min test ischaemia. The effects of MK-801 (2 mg/kg i.p.), LY293558 (20 mg/kg i.p., followed by 4 x 10 mg/kg at 3 h intervals), LY300164 (4 x 10 mg/kg i.p. at 1 h intervals), LY246492 (40 mg/kg i.p., followed by 4 x 20 mg/kg i.p. at 3 h intervals) and LY202157 (30 mg/kg i.p., followed by 4 x 15 mg/kg i.p. at 2 h intervals) were then examined in this model. Initial dosing commenced 30 min prior to the 2-min 'pre-conditioning' ischaemia. Results indicated that a 2-min 'pre-conditioning' ischaemia produced ischaemic tolerance in all cases. The non competitive NMDA receptor antagonist, MK-801, produced a significant (P < 0.01) reduction in the induced tolerance, while the competitive NMDA receptor antagonist, LY202157, also attenuated (P < 0.05) the induction of tolerance. In contrast, two AMPA receptor antagonists (LY293558 and LY300164) and a mixed NMDA/AMPA receptor antagonist (LY246492) had no effect on the induction of tolerance. These results suggest that NMDA receptor activation, but not AMPA receptor activation is involved in the phenomenon of ischaemic tolerance. PMID- 10513568 TI - Effect of donepezil hydrochloride (E2020) on basal concentration of extracellular acetylcholine in the hippocampus of rats. AB - The effects of oral administration of the centrally acting acetylcholinesterase (AChE) inhibitors, donepezil hydrochloride (donepezil: E2020: (+/-)-2-[(1 benzylpiperidin-4-yl)methyl]-5,6-dimethoxy-indan-1-one monohydrochloride), tacrine (9-amino-1,2,3,4-tetrahydroacridine hydrochloride) and ENA-713 (rivastigmine: (S)-N-ethyl-3-[(1-dimethyl-amino)ethyl]-N-methyl-phenylcarbamate hydrogentartrate), which have been developed for the treatment of Alzheimer's disease, on the extracellular acetylcholine concentration in the hippocampus of rats were evaluated by using a microdialysis technique without adding cholinesterase inhibitor to the perfusion solution. We also compared the inhibition of brain AChE and the brain concentrations of these drugs. Donepezil at 2.5 mg/kg and tacrine at 5 mg/kg showed significant effects for more than 6 h. At these doses, the maximum increases were observed at about 1.5 h after administration of donepezil, and at about 2 h with tacrine, and were 499% and 422% of the pre-level, respectively. ENA-713 produced significant effects at doses of 0.625, 1.25 and 2.5 mg/kg, which lasted for about 1, 2 and 4 h, respectively. The maximum increases produced by these doses at about 0.5 h after administration were 190, 346 and 458% of the pre-level, respectively. The time courses of brain AChE inhibition with donepezil at 2.5 mg/kg, tacrine at 10 mg/kg and ENA-713 at 2.5 mg/kg were mirror images of the extracellular acetylcholine increasing action at the same doses. The time courses of the brain concentrations of drugs after oral administration of donepezil at 2.5 mg/kg and tacrine at 10 mg/kg were consistent with those of brain AChE inhibition at the same doses, and there was a linear relation between these parameters. Brain concentration of ENA 713 at 2.5 mg/kg was below the limit of quantification at all time points measured. These results suggest that oral administration of donepezil, tacrine and ENA-713 increases acetylcholine concentration in the synaptic cleft of the hippocampus mostly through AChE inhibition, and that donepezil has a more potent activity than tacrine and a longer-lasting effect than ENA-713 on the central cholinergic system. PMID- 10513569 TI - Gamma-Hydroxybutyrate inhibits excitatory postsynaptic potentials in rat hippocampal slices. AB - Gamma-Hydroxybutyrate (GHB) has been shown to mimic different central actions of ethanol, to suppress alcohol withdrawal syndrome, and to reduce alcohol consumption both in rats and in humans. The aim of the present study was to determine if GHB shared with alcohol the ability to inhibit glutamate action at both NMDA and AMPA/kainate receptors. The NMDA or the AMPA/kainate receptors mediated postsynaptic potentials were evoked in CA1 pyramidal neurons by stimulation of Schaffer-collateral commissural fibers in the presence of CGP 35348, bicuculline to block the GABA(B) and GABA(A) receptors, and 10 microM 6,7 dinitroquinoxaline-2,3-dione (DNQX) or 30 microM DL-2-amino-5-phosphonovalerate (d-APV) to block AMPA/kainate or NMDA receptors, respectively. GHB (600 microM) produced a depression of both NMDA and AMPA/kainate receptors-mediated excitatory postsynaptic potentials with recovery on washout. The GHB receptors antagonist, NCS-382, at the concentration of 500 microM had no effect per se on these responses but prevented the depressant effect of GHB (600 microM) on the NMDA and AMPA/kainate-mediated responses. In the paired-pulse experiments, GHB (600 microM) depressed the amplitude of the first and the second evoked AMPA/kainate excitatory postsynaptic potentials, and significantly increased the paired-pulse facilitation (PPF). These results suggest that GHB inhibits excitatory synaptic transmission at Schaffer-collateral commissural-pyramidal neurons synapses by decreasing the probability of release of glutamate. PMID- 10513570 TI - 7-Nitroindazole reduces nitric oxide concentration in rat hippocampus after transient forebrain ischemia. AB - We investigated the effects of 7-nitroindazole, a specific inhibitor of neuronal nitric oxide (NO) synthase, on NO concentration and on blood flow in rat hippocampus after transient forebrain ischemia which was induced by 4-vessel occlusion for 10 min. NO concentration was measured directly by an NO-selective electrode method. Hippocampal blood flow was also estimated by laser Doppler flowmetry. 7-Nitroindazole [0 (vehicle), 12.5, 25, 50 or 100 mg/kg] was administered intraperitoneally 20 min before ischemia. 7-Nitroindazole at any dose used did not affect basal NO levels before ischemia. 7-Nitroindazole (25, 50 and 100 mg/kg) reduced the NO concentration significantly during post-ischemic early reperfusion. Before 10 min of ischemia and during post-ischemic early reperfusion, there were no significant differences in hippocampal basal blood flow and reactive hyperemia between vehicle- and 7-nitroindazole-treated groups. These results demonstrate that the neuronal NO synthase inhibitor, 7 nitroindazole, can effectively inhibit NO synthesis in rat hippocampus during post-ischemic early reperfusion. PMID- 10513572 TI - Type I and II metabotropic glutamate receptors mediate depressor and bradycardic actions in the nucleus of the solitary tract of anaesthetized rats. AB - The potential role of metabotropic glutamate (mGlu) receptors in cardiovascular function in the nucleus of the solitary tract was examined following the microinjection of a number of selective mGlu receptor compounds into this site of anaesthetized rats. The prototypic mGlu receptor selective agonist 1S,3R-1-amino cyclopentane dicarboxylate elicited depressor and bradycardic actions following microinjection into the nucleus tractus solitarius, which were similar to those produced by L-glutamate. Similarly, decreases in blood pressure and heart rate were observed upon administration of the type I and II selective mGlu receptor agonists, (R,S)-3,5-dihydroxyphenylglycine (DHPG) and 2R,4R-4-aminopyrrolidine 2,4-dicarboxylate (APDC), respectively. These actions of DHPG were selectively attenuated by (+/-)-1-aminoindane-1,5-dicarboxylate, a type I mGlu receptor antagonist, whilst cardiovascular responses to APDC were unaffected by this compound. Interestingly, the proposed type II antagonist, (2S,4S)-2-amino-4-(4,4 diphenylbut-1-yl)-pentane-1,5-doic acid, reduced the cardiovascular responses to intra-nucleus tractus solitarius administration of both APDC and DHPG. The type III mGlu receptor agonist, L-2-amino-4-phosphonobutyrate, however, failed to elicit any cardiovascular actions when microinjected into the nucleus tractus solitarius. These studies provide new evidence for functional type I and II mGlu receptors in modulating cardiovascular responses in the nucleus tractus solitarius. PMID- 10513571 TI - Cardiac inotropic vs. chronotropic selectivity of isradipine, nifedipine and clevidipine, a new ultrashort-acting dihydropyridine. AB - Cardiac effects of clevidipine, a new ultrashort-acting dihydropyridine Ca2+ channel antagonist were investigated in Langendorff-perfused rat hearts and compared to those of nifedipine and isradipine. The aim was to determine and compare the negative inotropic vs. chronotropic potency of these drugs. The hearts were perfused with oxygenated Krebs-Henseleit buffer at a perfusion pressure of 90 cm H2O. After stabilization, one concentration of each drug was administered for 45 min followed by a higher concentration for an additional 45 min. The concentrations of each drug in this study were 10(-9), 3 x 10(-9), 10( 8), 10(-7), 10(-6.5) and 10(-6) M for clevidipine and nifedipine, and 10(-10), 3 x 10(-10), 10(-9), 10(-8), 10(-7.5) and 10(-7) M for isradipine. Each concentration of each drug was tested in six hearts. Coronary flow, left ventricular dP/dt max, left ventricular systolic pressure and heart rate were recorded when the hearts were beating spontaneously and during pacing at a constant rate for 1 min. Spontaneous heart rate and atrio-ventricular conduction were not affected by clevidipine at any of the concentrations studied, while nifedipine and isradipine caused a concentration-dependent decrease. These two drugs caused atrio-ventricular block at high concentrations. All three compounds reduced cardiac contractility in a concentration-dependent manner. When isradipine was administered, at a given concentration, heart rate and contractility decreased proportionately. When clevidipine or nifedipine was given, at a given concentration, the proportionate reduction in left ventricular dP/dt max was greater than that in heart rate, resulting in a high inotropic vs. chronotropic selectivity. It is concluded that in contrast to nifedipine and isradipine, clevidipine does not impair atrio-ventricular conduction. Like nifedipine, clevidipine is selective for inotropic vs. chronotropic cardiac effects. PMID- 10513573 TI - An immunohistochemical and pharmacological study of tachykinins in the rat and guinea-pig prostate glands. AB - This study investigated the presence and effects of tachykinin peptides within the rat and guinea-pig prostate glands. Immunohistochemical studies demonstrated the presence of substance P and neurokinin A immunoreactive nerve fibres, sparsely distributed throughout the prostatic fibromuscular stroma in both species. In organ bath experiments, nerve terminals within rat and guinea-pig prostatic tissues were electrically field stimulated (60 V, 0.5 ms, 10 Hz, 20 pulses every 50 s). In rat preparations, the exogenous application of substance P, neurokinin A and the tachykinin NK3 receptor agonist senktide (1 nM-1 microM) had no effect on contractile responses. In contrast, substance P and neurokinin A (1 nM-3 microM) concentration-dependently enhanced electrically-evoked contractile responses in the guinea-pig prostate. Senktide was without effect. The potentiation of electrical field stimulation-induced contractions by substance P and neurokinin A in the guinea-pig prostate was competitively antagonized by ((S)1-[2-[3-(3,4-dichlorophenyl)-1-(3-isopropoxyphenylacetyl) piperidin-3-yl]ethyl]-4-phenyl-1-azonia-bicyclo[2.2.2]octane , chloride) (SR 140333) at 10 nM, a tachykinin NK1 receptor antagonist. The tachykinin NK2 receptor antagonist (S)-N-methyl-N[4-(4-acetylamino-4-phenylpiperidino)-2-(3,4 dichloropheny l)butyl]benzamide (SR 48968) was without effect at 10 nM, suggesting that neuromodulation of electrically-evoked contractions in the guinea pig prostate occurs through activation of a tachykinin NK1 receptor subtype. PMID- 10513574 TI - Luteolytic effects of DL111-IT in pregnant rats. AB - The present studies were conducted to evaluate the effects of DL111-IT [3-(2 ethyl phenyl)-5-(3-methoxy phenyl)-1H-1,2,4 triazol] on ovaries of pregnant rats. Pregnant rats were i.m. treated with DL111-IT 2.5 mg kg(-1) day(-1) or camellia oleum (vehicle control) 0.2 ml/day from day 6 of pregnancy for 1, 3 or 5 days. Blood and ovaries were collected 24 h after the last injection. Ovarian fresh weight and protein contents, activities of the 3beta-hydroxysteroid dehydrogenase (3beta-HSD) and 20alpha-hydroxysteroid dehydrogenase (20alpha-HSD) in ovaries, and cell apoptosis of corpus luteum (including hematoxylin-eosine stain, in situ 3'-end labeling and nucleosomal banding) were estimated. Compared with that in the control group, ovarian fresh weight declined 11% and 22% after DL111-IT-3 days and -5 days; protein content dropped 29% after 5-day administration. DL111 IT for 3 days provoked a marked decrease of serum progesterone, by 31% of the control; the activity of 3beta-HSD decreased 34.4% after i.m. DL111-IT for 5 days, while that of 20alpha-HSD increased dramatically after only one injection of DL111-IT (P < 0.01). Histological analysis and in situ 3'-end DNA labeling indicated that DL111-IT induced the pyknosis of cells and the formations of apoptotic bodies and intense oligonucleosomes in luteal cells of pregnant rats. The cell apoptosis induced by DL111-IT was further confirmed by evaluation of nucleosomal DNA fragmentation by agarose gel electrophoresis in cultured luteal cells exposed to DL111-IT for 24 h. In conclusion, all results, including shrunken luteal cells, decreased concentration of protein content and serum progesterone, changed activities of 3beta-HSD and 20alpha-HSD and formation of DNA fragments in luteal cells, showed the luteolytic effect of DL111-IT in pregnant rats. PMID- 10513576 TI - 5-hydroxytryptamine induces transient Ca2+ influx through Ni2+-insensitive Ca2+ channels in rat vascular smooth muscle cells. AB - The effects of Ni2+, a non-selective cation channel inhibitor, on 5 hydroxytryptamine (5-HT)- and angiotensin II (Ang II)-induced intracellular Ca2+ dynamics in rat aortic smooth muscle cells were investigated. Ni2+ (1 mM) significantly inhibited the transient increase in intracellular Ca2+ concentration ([Ca2+]i) induced by Ang II (100 nM) in aortic smooth muscle cells, as measured using fura-2. However, Ni2+ did not suppress the transient increase in Ca2+ influx induced by 5-HT (10 microM), while significantly suppressed the sustained increase. Ca2+ influx evoked by high KCl (80 mM), thapsigargin (TG) (1 microM) or depletion of intracellular Ca2+ store was almost completely suppressed by Ni2+. Ni2+ had no effect on 5-HT-induced inositol triphosphate production and Ca2+ release from the intracellular store(s). These results suggest that 5-HT, but not Ang II, induces transient Ca2+ influx through Ni2+-insensitive Ca2+ channels, which are distinguishable from the voltage-dependent or store-operated Ca2+ channels. PMID- 10513575 TI - Resolution, absolute stereochemistry and molecular pharmacology of the enantiomers of ATPA. AB - (RS)-2-Amino-3-(5-tert-butyl-3-hydroxy-4-isoxazolyl)propionic acid (ATPA), an analogue of (RS)-2-amino-3-(3-hydroxy-5-methyl-4-isoxazolyl)propionic acid (AMPA). has previously been shown to be a relatively weak AMPA receptor agonist and a very potent agonist at the GluR5 subtype of kainic acid-preferring (S) glutamic acid ((S)-Glu) receptors. We report here the separation of (+)- and (-) ATPA, obtained at high enantiomeric purity (enantiomeric excess values of 99.8% and > 99.8%, respectively) using chiral chromatography, and the unequivocal assignment of the stereochemistry of (S)-(+)-ATPA and (R)-(-)-ATPA. (S)- and (R) ATPA were characterized in receptor binding studies using rat brain membranes, and electrophysiologically using the rat cortical wedge preparation and cloned AMPA-preferring (GluR1, GluR3, and GluR4) and kainic acid-preferring (GluR5, GluR6, and GluR6 + KA2) receptors expressed in Xenopus oocytes. In the cortical wedge, (S)-ATPA showed AMPA receptor agonist effects (EC50 = 23 microM) approximately twice as potent as those of ATPA. (R)-ATPA antagonized depolarizations induced by AMPA (Ki = 253 microM) and by (S)-ATPA (Ki = 376 microM), and (R)-ATPA antagonized the biphasic depolarizing effects induced by kainic acid (Ki = 301 microM and 1115 microM). At cloned AMPA receptors, (S)-ATPA showed agonist effects at GluR3 and GluR4 with EC50 values of approximately 8 microM and at GluR1 (EC50 = 22 microM), producing maximal steady state currents only 5.4-33% of those evoked by kainic acid. (R)-ATPA antagonized currents evoked by kainic acid at cloned AMPA receptor subtypes with Ki values of 33-75 microM. (S)-ATPA produced potent agonist effects at GluR5 (EC50 = 0.48 microM). Due to desensitization of GluR5 receptors, which could not be fully prevented by treatment with concanavalin A, (S)-ATPA-induced agonist effects were normalized to those of kainic acid. Under these circumstances, maximal currents produced by (S)-ATPA and kainic acid were not significantly different. (R)-ATPA did not attenuate currents produced by kainic acid at GluR5, and neither (S)- nor (R) ATPA showed significant effects at GluR6. (S)-ATPA as well as AMPA showed weak agonist effects at heteromeric GluR6 + KA2 receptors, whereas (R)-ATPA was inactive. Thus, (S)- and (R)-ATPA may be useful tools for mechanistic studies of ionotropic non-NMDA (S)-Glu receptors, and lead structures for the design of new subtype-selective ligands for such receptors. PMID- 10513577 TI - Identification of an amino acid residue important for binding of methiothepin and sumatriptan to the human 5-HT(1B) receptor. AB - Site-directed mutagenesis of the human 5-HT1B receptor was performed to investigate the role of the amino acid residues cysteine 326 and tryptophan 327 in transmembrane region VI and aspartic acid 352 in transmembrane region VII in ligand binding. Binding studies were performed with the antagonist radioligand [3H]GR125743 on mutant and wild-type receptors stably expressed in Chinese hamster ovary cells (CHO)-K1 cells. Substitution of tryptophan 327 by alanine resulted in decreased affinities of all ligands tested. The most prominent changes in affinity were observed for the antagonist methiothepin and the antimigraine drug sumatriptan, which were reduced approximately 300- and 60-fold, respectively. Nevertheless, the affinity of 5-HT remained the same. Replacement of the aspartic acid 352 by alanine reduced high-affinity binding of 5-HT. Substitution of cysteine 326 by alanine had minor effects on ligand binding. Some of these results agree with the results from mutagenesis studies of the corresponding amino acids in other receptors. However, some notable differences also emerge showing that functional roles of individual amino acid residues must be tested experimentally in each receptor subtype. PMID- 10513578 TI - Site-directed mutagenesis of the putative human muscarinic M2 receptor binding site. AB - Experimental probing of the model of the muscarinic M2 receptor binding site proposed by Hibert et al. [Hibert, M.F., Trumpp-Kallmeyer, S., Bruinsvels, A., Hoflak, K., 1991. Three-dimensional models of neurotransmitter G-binding protein coupled receptors. Mol. Pharmacol. 40, 8-15.] was achieved by mutating each amino acid proposed to interact with muscarinic ligands. Pharmacological analysis of the different mutant receptors transiently expressed in human embryonic kidney (HEK/293) cells was performed with a variety of agonists and antagonists. D103A, Y403A and N404A mutations prevented binding of [3H] N-methylscopolamine and [3H] quinuclidinyl benzilate with a reduction in affinity greater than 100-fold, indicating essential contributions of these residues to the binding site for the radioligands. W400A and W155A mutations had very large effects on the binding of [3H] N-methylscopolamine (150-fold, 960-fold) but modest effects on the binding of [3H] quinuclidinyl benzilate (4-fold, 17-fold). In addition, binding of oxotremorine-M, oxotremorine, arecoline and pilocarpine to W155A resulted in a greater than 100-fold decrease in affinity. Threonine mutations (T187A and T190A) alter binding of most agonists but not of antagonists. W99 makes little contribution (< 10-fold) to the binding site of the M2 receptor. D103, W155, W400, Y403 and N404 are likely to be part of the binding site for N methylscopolamine and also to contribute to the binding site for quinuclidinyl benzilate. Some of the predicted residues do not seem to be part of the M2 receptor binding site but W155 is important for proper ligand binding on the muscarinic M2 receptor, as predicted by the proposed model. PMID- 10513579 TI - Autoradiographic comparison of the potency of several structurally unrelated adenosine receptor antagonists at adenosine A1 and A(2A) receptors. AB - We have examined the potency of several adenosine receptor antagonists at adenosine A1 and A2A receptors using quantitative autoradiography and have compared the results with those of previous studies using the same radioligands in membrane preparations. The agonists [3H]cyclohexyladenosine and [3H]2-[p-(2 carbonylethyl)-phenylethylamino]-5'-N-ethylcarbo xamido adenosine ([3H]CGS 21680) were used as radioligands for the two receptors. The results show that 1,3 dipropyl-8-cyclopentyl xanthine (DPCPX) is almost 1000-fold and 8-chloro-4 cyclohexyl-amino-1-(trifluoromethyl)[1,2,4]triazolo[4,3-a] quinoxaline (CP 68,247) about 300-fold more potent at adenosine A1 receptors in cortex and striatum than at striatal adenosine A2A receptors. Conversely, 5-amino-7-(2 phenylethyl)-2-(2-furyl)-pyrazolo-[4,3-e]-1,2,4-triazolo [1,5-c]pyrimidine (SCH 58261) is approximately 1000-fold and 4-(2-[7-amino-2-(2-furyl) [1,2,4] triazolo[2,3-a][1,3,5]triazin-5-yl amino]ethyl)phenol (ZM 241,385) about 400-fold more potent at adenosine A2A than at A1 receptors. Caffeine and its metabolites did not show any selectivity. Other studied antagonists were non-selective or showed a modest (20- to 40-fold) adenosine A2A receptor selectivity. Thus, only a few of the antagonists show such high selectivity that it is not offset by differences in drug distribution and levels of receptor subtype expression. PMID- 10513581 TI - The hippocampus and mechanisms of declarative memory. AB - The hippocampus is critical to declarative memory in humans and spatial memory in rodents. This review attempts to bridge between these two characterizations of hippocampal dependent memory, and in doing so reveal fundamental cognitive and neural coding mechanisms that are common to both. Evidence is presented that the hippocampus and its connections are critical to the establishment of a systematic organization of memories and to flexible expression of memory outside repetition of the training experience. In addition, evidence is presented that hippocampal neurons encode a broad range of experience and that these codings may be organized as representations of episodes in memory. It is suggested that these episodic codings are linked by common elements to construct an organized representation of acquired knowledge. PMID- 10513580 TI - Characterization of the cloned guinea pig leukotriene B4 receptor: comparison to its human orthologue. AB - A cDNA clone coding for the guinea pig leukotriene B4 (BLT) receptor has been isolated from a lung cDNA library. The guinea pig BLT receptor has an open reading frame corresponding to 348 amino acids and shares 73% and 70% identity with human and mouse BLT receptors, respectively. Scatchard analysis of membranes prepared from guinea pig and human BLT receptor-transfected human embryonic kidney (HEK) 293 EBNA (Epstein-Bar Virus Nuclear Antigen) cells showed that both receptors displayed high affinity for leukotriene B4 (Kd value of approximately 0.4 nM) and were expressed at high levels (Bmax values ranging from 9 to 12 pmol/mg protein). The rank order of potency for leukotrienes and related analogs in competition for [3H]leukotriene B4 specific binding at the recombinant guinea pig BLT receptor is leukotriene B4 > 20-OH-leukotriene B4 > 12(R)-HETE ((5Z,8Z,10E,12(R)14Z)-12-hydroxyeicosatetraen -1-oic acid) > 12(S)-HETE ((5Z,8Z,10E,12(S)14Z)-12-Hydroxyeicosatetraen -1-oic acid) > 20-COOH-leukotriene B4 > U75302 (6-(6-(3-hydroxy-1E,5Z-undecadienyl)-2-pyridinyl)-1,5-hexane diol) >> leukotriene C4 = leukotriene D4 = leukotriene E4. For the human receptor the rank order of 12(S)-HETE, 20-COOH-leukotriene B4 and U75302 was reversed. Xenopus melanophore and HEK aequorin-based reporter gene assays were used to demonstrate that the guinea pig and human BLT receptors can couple to both the cAMP inhibitory and intracellular Ca2+ mobilization signaling pathways. However, in the case of the aequorin-expressing HEK cells (designated AEQ17-293) transfected with either the guinea pig or human BLT receptor, expression of Galpha16 was required to achieve a robust Ca2+ driven response. Leukotriene B4 was a potent agonist in functional assays of both the guinea pig and human BLT receptors. U 75302 a leukotriene B4 analogue which possesses both agonistic and antagonistic properties behaved as a full agonist of the guinea pig and human BLT receptors in AEQ17-293 cells and not as an antagonist. The recombinant guinea pig BLT receptor will permit the comparison of the intrinsic potencies of leukotriene B4 receptor antagonists used in guinea pig in vivo models of allergic and inflammatory disorders. PMID- 10513582 TI - Monocular horizontal OKN in observers with early- and late-onset strabismus. AB - Several reports on monocular optokinetic nystagmus (OKN) in observers with strabismus have found that asymmetry of OKN tends to occur in both eyes of observers with an early onset of strabismus but only in the deviating eye of those with a later onset of strabismus. Our objective was to quantify and compare the magnitude of the OKN asymmetry in each eye as a function of observer's age at onset of strabismus. We studied monocular OKN in ten observers with early-onset (up to 24 months of age), seven observers with late-onset (after 24 months of age) unilateral strabismus, and 12 normally sighted control observers. In the deviating eye, observers with early-onset strabismus showed large OKN asymmetries in favour of nasalward motion while observers with late-onset strabismus showed smaller OKN asymmetries in that eye. The majority of early- and late-onset observers showed near normal OKN in the non-deviating eye although the early onset observers showed bilateral asymmetries more often. These findings may be due to both age at onset of strabismus and chronological age and are discussed in terms of the issue of plasticity or recovery of function. PMID- 10513583 TI - Progressive and gender-dependent cognitive impairment in the APP(SW) transgenic mouse model for Alzheimer's disease. AB - To determine if early cognitive sensorimotor deficits exist in APP(SW) transgenic mice overexpressing human amyloid precursor protein (APP). Tg+ and Tg- animals at both 3 and 9 months of age (3M and 9M, respectively) were evaluated in a comprehensive battery of measures. The performance of all Tg+ mice at both ages was no different from all Tg- controls in Y-maze alternations, water maze acquisition, passive avoidance, and active avoidance testing. By contrast, results from other tasks revealed substantive cognitive deficits in Tg+ mice that were usually gender-dependent and sometimes progressive in nature. Between 3M and 9M, a progressive impairment was observed in circular platform performance by Tg+ males, as was a progressive deficit in visible platform testing for all Tg+ animals. Other transgenic effects included both impaired water maze retention and circular platform performance in 3M Tg+ females; this later effect was responsible for an overall (males + females) Tg+ deficit in circular platform performance at 3M. Sensorimotor testing revealed several Tg+ effects, most notably an increased activity of Tg+ males in both open field and Y-maze at 3M. Significant correlations between a number of behavioral measures were observed, although factor analysis suggests that each task measured components of sensorimotor/cognitive function not measured by other tasks. Finally, Tg+ mice had lower survivability than Tg- animals through 9M (85 vs. 96%). In summary, these results demonstrate the presence of gender-related and progressive cognitive deficits in APP(SW) transgenic mice at a relatively early age (i.e., prior to overt, beta-amyloid deposition in the brain), suggesting a pathophysiologic role for elevated levels of 'soluble' beta-amyloid in such impairments. PMID- 10513584 TI - Variations in CS associability and multiple unit hippocampal activity in the rabbit. AB - Hippocampal multiple unit activity was recorded in rabbits during each of four preacquisition treatments and during subsequent classical conditioning of the nictitating membrane response. Three preexposure conditions were employed: CS alone presentations, presentations of the CS paired with a second, neutral stimulus, or unpaired presentations of the CS and second stimulus. It was predicted that (a) CS alone preexposures would produce a decrease in hippocampal activity and a retarded rate of subsequent conditioned response (CR) acquisition and (b) the magnitude of both effects would be attenuated by preexposures of the CS paired with a second stimulus. The results partially supported both predictions. Hippocampal activity was inhibited during CS alone preexposures and that inhibition was attenuated by pairing the CS with a second, neutral stimulus. Behaviorally, all of the preexposure groups showed equivalent, retarded rates of acquisition compared to a nonpreexposed control group. Hippocampal activity throughout acquisition was significantly greater in the nonpreexposed group compared to the group preexposed to the CS alone. Hippocampal activity of the other two groups was intermediate between the nonpreexposed and the CS alone groups. It is suggested that alterations in the magnitude of hippocampal activity may provide a reliable, neuronal correlate of CS associability changes. PMID- 10513585 TI - Lesions of midline midbrain structures leave medial forebrain bundle self stimulation intact. AB - Previous work with psychophysically-based collision methods and pharmacological manipulation suggests a role in medial forebrain bundle (MFB) self-stimulation for neurons lying along the midline between the cerebral hemispheres, in the mid- and/or hindbrain. Also, recently-proposed models of the anatomical substrate for medial forebrain bundle stimulation reward suggest that at least part of the directly-activated axons of this substrate arise from mid- and/or hindbrain somata, bifurcate, and send bilateral projections to the MFB of each hemisphere. Branches of these axons are thought to cross the midline at some point near the ventral tegmental area. This study examines the effects on MFB stimulation reward of lesioning midbrain structures that lie along the midline between hemispheres. In 13 rats, lesions of the median raphe, the decussation of the superior cerebellar peduncle, or the interpeduncular nucleus were all ineffective in altering the stimulation frequency required to maintain half-maximal levels of operant responding for stimulation reward. These results are discussed in terms of implications for recent models of the anatomical substrate for brain stimulation reward. PMID- 10513587 TI - Interaction between acoustic and electric sensitization of the acoustic startle response in rats. AB - Sensitization is the general increase of responsiveness observed after aversive stimulation. Usually footshocks are used as aversive stimuli. According to the 'Dual Process Theory' by Groves and Thompson. Psychol. Rev. 1970;77:419-450, not only additional aversive stimuli but also the response-eliciting stimuli themselves have a sensitizing effect, the degree of sensitization depending upon the stimulus intensity. We tested this suggestion in the footshock sensitization paradigm of the acoustic startle response (ASR): (1) High SPL (sound pressure level) acoustic stimuli (119 dB SPL) presented instead of footshocks also elicited strong sensitization. (2) While footshocks presented after startle stimuli with low SPL (95 dB) were able to produce a strong further sensitization of the ASR, footshocks presented after startle stimuli with high SPL (110 dB) only caused a minor sensitization of the ASR. (3) Diazepam (3 mg/kg i.p.) decreased ASR to high SPL (115 dB) stimuli. In this case footshocks elicited significant sensitization of the ASR despite intense startle stimuli. The present results support the 'Dual Process Theory'. Furthermore we could show that acoustic and footshock sensitization interact. We therefore suggest that both, acoustic and footshock sensitization, are mediated partly via the same neural circuitry. PMID- 10513586 TI - Effects of menstrual cycle phase and oral contraceptives on alertness, cognitive performance, and circadian rhythms during sleep deprivation. AB - The influence of menstrual cycle phase and oral contraceptive use on neurobehavioral function and circadian rhythms were studied in healthy young women (n = 25) using a modified constant routine procedure during 24 h of sleep deprivation. Alertness and performance worsened across sleep deprivation and also varied with circadian phase. Entrained circadian rhythms of melatonin and body temperature were evident in women regardless of menstrual phase or oral contraceptive use. No significant difference in melatonin levels, duration, or phase was observed between women in the luteal and follicular phases, whereas oral contraceptives appeared to increase melatonin levels. Temperature levels were higher in the luteal phase and in oral contraceptive users compared to women in the follicular phase. Alertness on the maintenance of wakefulness test and some tests of cognitive performance were poorest for women in the follicular phase especially near the circadian trough of body temperature. These observations suggest that hormonal changes associated with the menstrual cycle and the use of oral contraceptives contribute to changes in nighttime waking neurobehavioral function and temperature level whereas these factors do not appear to affect circadian phase. PMID- 10513588 TI - Environmental modulation of amphetamine-induced c-fos expression in D1 versus D2 striatal neurons. AB - We have reported previously that exposure to environmental novelty enhances the behavioral activating effects of amphetamine and its ability to induce the immediate early gene c-fos in the striatum and in other brain regions. In the present study, we used double in situ hybridization histochemistry to study the effect of amphetamine and/or novelty on c-fos expression in two populations of striatal neurons that preferentially express either D1 or D2 dopamine receptor mRNA. When given intraperitoneally to rats in their home cage, amphetamine (2.0 mg/kg) increased c-fos expression only in D1 neurons. In contrast, when the same dose of amphetamine was administered to rats in a novel environment, c-fos was increased in both D1 and D2 neurons. We conclude that the neural populations engaged by amphetamine vary as a function of the circumstances surrounding its administration. PMID- 10513589 TI - Transfer of learned information between ganglia in the insect ventral nerve cord. AB - A yoked control training procedure was used on the decapitated cockroach, L. maderae. The right prothoracic leg was trained to lift in order to avoid a shock. It was found that this information transferred via the two interganglionic connectives from the first or prothoracic ganglion (T1) to the second or mesothoracic ganglion (T2) so that now the right mesothoracic leg lifted to avoid shock even though it was not directly trained. If both connectives were cut before training T1, no transfer to T2 was seen, i.e. the mesothoracic leg did not lift and avoid shock. However, if both connectives were cut immediately after training T1, the information had already transferred and was available for use by T2. There was redundancy in the transfer in that either connective alone could carry the same information from T1 to T2. Either mesothoracic leg could tap into this information. Using a reversible cold block on the connectives, it was found that if it was applied before training T1 it did not interfere with T1 learning but no transfer to T2 was seen after the cold block wore off. That the block was transitory and did not permanently impair the connectives was shown by the fact that if it was applied and then allowed to wear off before training began there was normal learning in T1 and transfer to T2. The transection and cold-block studies were consistent in demonstrating that the transfer of the information was 'on-line' and only occurred during T1 learning. If transfer was blocked during T1 learning the information could not be transferred or tapped into by T2 at a later time even though it was stored in T1 and available for later use by T1. The transfer occurred so quickly it most likely occurred via nerve impulses. Because no primary sensory or motor neurons are in the connectives, the information must have been coded onto interneurones for transfer from the first (T1) to the second (T2) ganglion. PMID- 10513590 TI - A within-subjects microdialysis/behavioural study of the role of striatal acetylcholine in D1-dependent turning. AB - In rats lesioned with 6-hydroxydopamine (6-OHDA) the effect of the noncompetitive N-methyl D-aspartate (NMDA) receptor antagonist, MK-801, the dopamine (DA) D2 receptor agonist quinpirole and the A2A adenosine antagonist SCH 58261 was studied on acetylcholine (ACh) release in the lesioned striatum and contralateral turning behaviour stimulated by the administration of the DA D1 receptor agonist CY 208-243. Administration of CY 208-243 (75, 100 and 200 microg/kg) to 6-OHDA lesioned rats dose-dependently stimulated ACh release and induced contralateral turning. MK-801 (50 and 100 microg/kg) reduced basal ACh release (max 22%) and did not elicit any turning. MK-801 (50 and 100 microg/kg) potentiated the contralateral turning, but failed to modify the stimulation of ACh release elicited by 100 and 200 microg/kg of CY 208-243. MK-801 (100 microg/kg) prevented the increase in striatal ACh release evoked by the lower dose of CY 208-243 (75 microg/kg) but contralateral turning was not observed. The D2 receptor agonist quinpirole (30 and 60 microg/kg) elicited low-intensity contralateral turning and decreased basal ACh release. Quinpirole potentiated the D1-mediated contralateral turning behaviour elicited by CY 208-243 (100 microg/kg), but failed to affect the increase in ACh release elicited by the D1 agonist. The adenosine A2A receptor antagonist SCH 58261 (1 microg/kg i.v.) failed per se to elicit contralateral turning behaviour. SCH 58261 potentiated the contraversive turning induced by CY 208-243 but failed to affect the increase of ACh release. The results of the present study indicate that blockade of NMDA receptors by MK-801. stimulation of DA D2 receptors by quinpirole and blockade of adenosine A2A receptors by SCH 58261 potentiate the D1-mediated contralateral turning behaviour in DA denervated rats without affecting the action of the D1 agonist on ACh release. These observations do not support the hypothesis that the potentiation of D1-dependent contralateral turning by MK-801, quinpirole or SCH 58261 is mediated by changes in D1-stimulated release of ACh in the striatum. PMID- 10513591 TI - What causes lateralization of detour behavior in fish? Evidence for asymmetries in eye use. AB - A consistent population bias to detour a vertical-bar barrier preferentially leftwise during approach to inspect a dummy predator was demonstrated in the poeciliid fish Girardinus falcatus. The asymmetry seems to be due to a preferential use of the lateral visual field of the right eye during fixation of biologically relevant stimuli such as a predator. Viewing tests revealed in fact that fish which tended to detour the barrier on the left side used the right eye to scrutiny a dummy predator and the left eye to scrutiny a neutral stimulus, whereas fish which tended to detour the barrier on the right side showed the reverse pattern of eye use; fish that did not show any consistent bias in the detour test did not reveal any significant preference in the viewing test. PMID- 10513592 TI - Statistical evaluation of clusters derived by nonlinear mapping of EEG spatial patterns. AB - New methods were devised to improve the discrimination of EEG spatial amplitude patterns recorded from arrays of 64 electrodes placed on visual, auditory or somatic cortex. The 64 traces shared a spatially coherent, aperiodic carrier wave with a spatial pattern of amplitude modulation (AM). Previous observations on AM patterns from rabbits trained to discriminate conditioned stimuli with reinforcement (CS+) and without (CS-) had revealed epochs between the CS and the CR in which AM patterns on CS+ trials could be distinguished from AM patterns on CS- trials. The AM patterns were expressed by points in 64-space that formed clusters. Levels of CS-/CS+ pattern separation were quantified by a pair-wise Euclidean distance method with cross-validation. The present study documents use of the technique for nonlinear mapping (NLM) to project the 64-dimensional structure onto a plane while preserving the relative distances between all points. The goodness of classification by the Euclidean distance measure was the same or improved after projection. Whereas the Euclidean distance measure only gave pair-wise classifications, the planar displays showed the patterns for multiple clusters simultaneously. These NLM-based methods revealed previously unrecognized structures within distributions of AM patterns in sensory cortices in the time period between the CS and CR. PMID- 10513593 TI - A 6-dof device to measure head movements in active vision experiments: geometric modeling and metric accuracy. AB - This work describes a technique for measuring human head movements in 3D space. Rotations and translations of the head are tracked using a light helmet fastened to a multi-joint mechanical structure. This apparatus has been designed to be used in a series of psycho-physiological experiments in the field of active vision, where position and orientation of the head need to be measured in real time with high accuracy, high reliability and minimal interference with subject movements. A geometric model is developed to recover the position information and its parameters are identified through a calibration procedure. The expected accuracy, derived on the basis of the pure geometric model and the sensor resolution, is compared with the real accuracy, obtained by performing repetitive measurements on a calibration fixture. The outcome of the comparison confirms the validity of the proposed solution which turns out to be effective in providing measurement of head position with an overall accuracy of 0.6 mm and sampling frequency above 1 kHz. PMID- 10513594 TI - An in vitro rabbit retina model to study electrophysiologic and metabolic function during and following ischemia. AB - Most in vitro studies involving neuronal ischemia use biochemical measures and/or cell counting to assess cellular death. We describe an in vitro rabbit retina model in which we measured glucose utilization, lactate production, and light evoked compound action potentials (CAPs) to assess metabolic and functional recovery following ischemia. Under control conditions, retinal glucose utilization and lactate production (n = 7), as well as CAPs (n = 8) remained quite constant for 6-8 h. During ischemia (glucose reduced from 6 to 1 mM and oxygen from 95 to 15%), glucose utilization and lactate production fell to 50%. CAPs fell to 50% in 3-4 min, and to 0% in 8-10 min. Recovery during 3-4 h of 'return-to-control' was dependent upon the length of ischemia. Glucose utilization recovered to 63% after 1 h (n = 4) and to 18% after 2 h of ischemia (n = 6, P < 0.001). Lactate production recovered to 77% after 1 h (n = 4) and to 54% after 2 h of ischemia (n = 6, P < 0.001). CAPs returned to 51, 15, and 0.13% of the control responses after 0.5 h (n = 7), 1 h (n = 8), and 2 h (n = 5) of ischemia, respectively (P < 0.001). This avascular, blood-brain barrier-free preparation provides an opportunity to use both metabolic and functional criteria to test protection against neuronal ischemia. PMID- 10513595 TI - Slice blotting: a method for detecting the release of immunoreactive substances from living brain tissue. AB - The intricate circuitry of the CNS forms highly organized structures containing a multitude of transmitters, modulators and other chemical signals expressed in specific patterns and pathways. Anatomical studies with immunohistochemical and molecular biological techniques have mapped in fine detail the distribution of these substances in fixed tissue. However, the release of neuroactive substances is often under precise spatial control and is regulated by numerous physiological factors. Understanding such complex intercellular signaling systems will require the development of new spatially resolved methods for detecting secretion in living systems. A simple but powerful method is described here for visualizing and quantifying the time-integrated spatial pattern of release of chemical signals from living neural tissue. The method combines the in vitro brain slice preparation with immunostaining protocols used for antigen detection on Western blots. It has widespread potential application in biological research because it can map in vitro patterns of release of cytokines, growth factors, chemoattractants, chemorepellents, morphogens, enzymes, and other paracrine signals in spatially organized systems, subject to a variety of stimuli and conditions. PMID- 10513596 TI - An ultrastructural analysis of tissue surrounding a microdialysis probe. AB - Microdialysis is a widely used in vivo sampling technique commonly used to monitor extracellular levels of a variety of molecules including neurotransmitters and metabolites. To facilitate interpretation of microdialysis results, this study critically examines changes in synaptic morphology induced by microdialysis. Tissue surrounding microdialysis probes was examined using light and electron microscopy at three distances from the probe tract. Microdialysis probes were implanted into rat striatum, and after 40 h of post-operative recovery were perfused with a modified Ringer's solution. Light microscope analysis revealed tissue disruption up to 1.4 mm from the probe site. Axonal damage indicative of non-excitotoxic insult was also seen as far away from the probe as was examined. The presence of dark-degenerating neurons was also noted and estimates of neuronal densities revealed loss up to 400 microm from the probe tract. This study, the first qualitative ultrastructural investigation of neuropil surrounding the probe site, indicated swollen processes up to 1.4 mm from the probe tract. Swollen mitochondria and bloated endoplasmic reticulum suggest intracellular chemical disruption. Tissue damage resulting in synaptic and neuronal disruption may affect neurotransmitter efflux or extracellular concentrations of metabolites. PMID- 10513597 TI - High-resolution functional labeling of vertebrate and invertebrate olfactory receptor neurons using agmatine, a channel-permeant cation. AB - Methods are described for odor-stimulated labeling of olfactory receptor neurons (ORNs) of the freshwater zebrafish Danio rerio and the marine spiny lobster Panulirus argus. Permeation of a cationic molecule, 1-amino-4-guanidobutane ( = agmatine, AGB), through ion channels following odor stimulation, and its detection by an anti-AGB antibody, allow labeling of odor-stimulated ORNs. Parameters adjusted to optimize activity-dependent labeling included labeling medium ionic composition, stimulation times, and AGB concentration. For lobsters, 7% of ORNs were labeled by a complex odor, oyster mixture, under optimal conditions, which was stimulation for 5 s per min for 60 min with 20 mM AGB in artificial seawater with reduced sodium and calcium concentrations. AGB was a weak odorant for lobsters; it elicited only a small electrophysiological response from ORNs and labeled < 1% of the ORNs during stimulation with AGB in the absence of odors. For the zebrafish, stimulation for 10 s per min for 10 min with 5 mM AGB plus odorant (L-glutamine) in fish Ringer's solution was the optimal labeling condition, resulting in labeling of 17% of the olfactory epithelial area. Approximately 6% of the olfactory epithelium was labeled during stimulation with a control stimulus, AGB alone. This labeling by AGB alone suggests it is an olfactory stimulus for zebrafish; a conclusion supported by electrophysiological recordings. We used electrophysiological assays and channel blockers to examine, for each species, potential ion channels for entry of AGB into ORNs. These results show that AGB can be used as an activity-dependent label for chemoreceptor neurons of diverse phyla living in a range of environmental conditions. PMID- 10513598 TI - Digitized analysis of handwriting and drawing movements in healthy subjects: methods, results and perspectives. AB - The diagnosis of movement disorders and the distinction between their possible generation by drug-treatment or illness can be done more objectively by using digitized analyses of hand movements. The aim of this study was to define this method, that is to identify its reliability and the influence of several covariables upon measurements, in healthy subjects. Simple writing and drawing tests were administered, using a digitizing tablet, transmitting signals to a computer for processing. The kinematic parameters identified in this way provided objective, reliable and valid measures for the dynamics and the degree of automation of hand movements. Analysis of the data showed that younger subjects write faster and with a higher degree of automation than older subjects. Other moderating variables, such as verbal intelligence and customary motor activity in everyday life (motoric practice) could be identified, whereas personality and gender were found to have little influence. There were no significant differences between left-handers and right-handers in hand movements. The movement parameters had a high test retest stability. The results of this study in healthy subjects indicate that age, verbal intelligence and motor practice should be considered when evaluating the effects of drug-treatment or psychiatric illness upon hand movement in patients. PMID- 10513599 TI - Difference in the in vivo influence of serotonin1A autoreceptors on serotonin release in prefrontal cortex and dorsal hippocampus of the same rat treated with fluoxetine. AB - Recent studies have demonstrated that antagonism of somatodendritic serotonin1A (5-HT1A) autoreceptors can potentiate the increase of extracellular 5-HT concentrations induced by selective serotonin reuptake inhibitors including fluoxetine. The present study was conducted to uncover any functional difference between the 5-HT1A autoreceptors located on the cell bodies of 5-HT neurons in the dorsal (DRN) and median (MRN) raphe nuclei. The investigational approach used in the present study was to detect extracellular 5-HT concentrations in two terminal areas, prefrontal cortex (Pfc) and dorsal hippocampus (Dhp), which are mainly innervated by the 5-HT neurons located in the DRN and MRN respectively. To avoid possible variation between individual animals a dual-probe microdialysis procedure was applied to determine 5-HT concentrations in both brain areas of the same rat. Fluoxetine (10 mg/kg, s.c.) alone produced a smaller increase in the extracellular 5-HT concentration in the Pfc than Dhp of the same rat (maximal 5 HT concentrations were 183% and 223% of the baseline values in Pfc and Dhp respectively). However, an antagonist of 5-HT1A receptors, WAY100635, subsequently injected (s.c.) at 1 mg/kg brought the 5-HT concentrations to similar levels in the Pfc (332%) and Dhp (308%). Since the 5-HT concentrations immediately before the injection of WAY100635 were lower in the Pfc (102%) than Dhp (186%), WAY100635 induced a larger 5-HT net increase in the Pfc (332% 102%=230%) than Dhp (308%-186%=122%). On the other hand, WAY100635 alone did not significantly change the extracellular 5-HT concentrations in both areas. Furthermore, extracellular concentrations of dopamine (DA) and two DA metabolites, 3,4-dihydroxyphenylacetic acid and homovanillic acid, in both areas were not altered by the administrations of fluoxetine and WAY100635. In conclusion, the present study demonstrated that the antagonist of 5-HT1A receptors, WAY100635, produced a more robust potentiation in the fluoxetine induced 5-HT increases in the Pfc than Dhp. Since Pfc and Dhp are predominately innervated by 5-HT neurons located in the DRN and MRN respectively, this result may indicate a functional difference between the 5-HT1A autoreceptors located on the cell bodies of 5-HT neurons in the DRN and MRN. PMID- 10513600 TI - Chronic exercise enhances vascular responses to clonidine in rats by increasing endothelial alpha2-adrenergic receptor affinity. AB - Chronic exercise increases endothelium-dependent vasodilating responses. To investigate whether endothelial alpha2-adrenergic receptor upregulation is involved in the enhancement of clonidine-induced vasorelaxation by chronic exercise, 4-week-old male Wistar rats were used. They were divided into control and exercise groups. The trained animals ran on a treadmill at a moderate intensity for 60 min per day, 5 days per week for 10 weeks in total. Resting heart rates were measured by a tail-cuff method to confirm training effects. After training, rings of the thoracic aorta were prepared to evaluate vasodilating responses to clonidine, an alpha2 agonist. Released endothelium derived relaxing factors were pharmacologically identified by treatment of N(omega)-nitro-L-arginine, a nitric oxide (NO) synthase inhibitor, or tetraethylammonium chloride, an endothelium-derived hyperpolarization factor (EDHF) inhibitor. Receptor binding assays were performed by using 3H-labeled clonidine as a tracer. We found that chronic exercise enhanced vascular responses to clonidine by stimulating the release of both NO and EDHF. It also increased the binding affinity of endothelial cell alpha2 receptor without changing the number of binding sites. Therefore, the elevated vasorelaxing responses to clonidine after chronic exercise may be partially resulted from an increase in endothelial alpha2 receptor binding affinity. PMID- 10513601 TI - Activation of gonadotropin-releasing hormone receptors produces neuronal excitation in the rat hippocampus. AB - Whole-cell patch-clamp recordings of evoked action potentials were made in CA1 and CA3 pyramidal neurons of rat hippocampal slices. Previously we have demonstrated that activation of gonadotropin-releasing hormone (GnRH) receptors induces a long-lasting enhancement of synaptic transmission mediated by ionotropic glutamate receptors in CA1 pyramidal neurons of rat hippocampal slices. Here, we further studied whether activation of GnRH receptors could modulate intrinsic neuronal excitability in CA1 and CA3 pyramidal neurons of rat hippocampal slices. The use of a specific GnRH analog, leuprolide (10(-8) M), elicited a relatively long-term increase in evoked action potentials in CA1 and CA3 pyramidal neurons, respectively. The GnRH receptor-induced increase in evoked action potentials in both CA1 and CA3 pyramidal neurons could be abolished by a potent GnRH receptor antagonist, [acetyl-3,4-dehydro-Pro1,D-p-F-Phe2,D-Trp(3,6)] LHRH (10(-8) M). The present study suggests that activation of GnRH receptors can lead to an increase of intrinsic neuronal excitability of both CA1 and CA3 pyramidal neurons in the rat hippocampus, an important integrative region for reproductive process, both endocrinologically and behaviorally. PMID- 10513602 TI - Corticotropin-releasing factor enhances brain-derived neurotrophic factor gene expression to facilitate memory retention in rats. AB - In the present study, we investigated the effects of corticotropin-releasing factor (CRF) injected into the dentate gyrus (DG) of the hippocampus on brain derived neurotrophic factor (BDNF) mRNA expression and studied whether N-methyl-D aspartate (NMDA) receptor mediates the effects of CRF on BDNF mRNA expression in the DG. Since both CRF and BDNF gene expressions are involved in memory processing in rats, we further investigated whether CRF facilitates memory retention through enhanced BDNF mRNA expression in the hippocampus. Effect of direct BDNF injection to the DG on retention performance in rats was also assessed. Results indicated that intra-DG CRF injection produced a dose-dependent (0.1 microg, 1.0 microg and 10 microg) increase in BDNF mRNA level, while intra DG MK801 injection produced a dose-dependent (0.08 microg, 0.2 microg and 2.0 microg) decrease in BDNF mRNA expression in the DG. MK801, at a dose having no significant effect alone (0.08 microg), significantly antagonized the effect of CRF on BDNF mRNA expression. On the other hand, CRF (1.0 microg) consistently and markedly improved retention performance in rats in an inhibitory avoidance learning task. BDNF antisense oligonucleotide treatment, at a concentration which did not affect retention performance alone (0.5 mM), blocked the memory-enhancing effect of CRF. However, direct and chronic BDNF injection to the DG did not improve memory performance in rats. These results together suggest that at least one of the mechanisms responsible for the memory-facilitating effect of CRF is mediated through enhanced BDNF mRNA expression in the hippocampus. The lack of an effect of intra-DG BDNF injection on memory retention is also discussed. PMID- 10513603 TI - Nicotine-induced hyperlocomotion is not modified by the estrous cycle, ovariectomy and estradiol replacement at physiological level. AB - The present study was designed to investigate whether nicotine's effect on locomotion might be modulated by the ovarian hormone at physiological level. Rats at normal cycling of estrus and diestrus were selected for the comparison of nicotine-induced hyperlocomotion based on the document that the release of striatal dopamine was greatest at the estrous phase. Ovariectomized rats primed with or without estrogen at physiological level were also selected for comparison. Increase in spontaneous locomotion by nicotine was statistically significant at the doses of 0.15 and 0.3 mg/kg (p < 0.001). The stimulating effect of nicotine led the locomotor response to almost the same magnitude in all hormonal groups studied. Nicotine-induced hyperlocomotion appeared to be mediated by central nicotinic receptor because it was blocked by mecamylamine (0.5 and 1.0 mg/kg, i.p.). Also it was blocked by haloperidol (0.04 and 0.08 mg/kg, i.p.) indicating the involvement of dopaminergic neurotransmission. These effects were similar in all groups regardless of the estrous cycle or ovariectomy. The observed data provided behavioral evidence to suggest that the effect of nicotine on locomotion-related dopaminergic neurons might not be modified by the physiological action of estrogen. PMID- 10513604 TI - Effects of thyroid hormones on the release of calcitonin gene-related peptide (CGRP) by rat prostate glands in vitro. AB - It has been well known that calcitonin (CT) and calcitonin gene-related peptide (CGRP) are derived from the CT/CGRP gene which is localized in chromosome 11. CGRP is a 37-amino acid neuropeptide expressed predominantly in the nervous system and is one of the most potent endogenous vasodilatory peptides that have been found. Only few reports described the distribution of CGRP in reproductive organs. Moreover, the hormonal regulation of CGRP secretion is still not clear. The present study was designed to examine the presence of CGRP in rat prostates and the direct effect of thyroxine (T4) on the release of CGRP by rat prostate glands in vitro. Male rats were thyroidectomized (Tx) or sham Tx for two weeks before decapitation. The ventral prostate glands were either extracted by phosphate buffer saline or bisected and preincubated with Locke's solution containing 10 mM glucose, 0.03% bacitracin, and 0.05% Hepes at 37 degrees C for 90 min. The hemi-prostate tissues were then incubated with Locke's medium containing T4 (0 to approximately 10(-7) M) for 1 hr. After incubation, the medium was collected, and the prostate tissues were weighed. The concentration of CGRP in both medium and prostate tissue extracts were measured by a specific radioimmunoassay (RIA) developed in our laboratory. Incubation of T4 at 10(-9) M was effective to increase the release of CGRP in rat prostate glands. Incubation of rat prostate glands with T4 at 10(-7) M resulted in a maximal release of CGRP (270% of the basal). These results suggest that thyroid hormones increase CGRP release by acting directly on rat prostate glands. PMID- 10513605 TI - Non-NMDA receptors mediate both pressor and depressor actions of the cardiovascular-reactive areas in the brainstem of cats. AB - L-glutamate (Glu), an important excitatory transmitter in the central nervous system, is mainly mediated via two kinds of ionotropic Glu receptors: N-methyl-D aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (AMPA)/kainate (non-NMDA) receptors. Microinjection of Glu (0.1 M, 30 nL) into gigantocellular tegmental field (FTG), dorsomedial medulla (DM) and rostral ventrolateral medulla (RVLM) induced increases of the systemic arterial pressure (SAP) and the sympathetic vertebral nerve activities (VNA), while its microinjection into caudal ventrolateral medulla (CVLM) induced decreases of SAP and VNA. In this study, the 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), a non NMDA antagonist, was used to examine the effects of non-NMDA receptors on Glu induced cardiovascular responses. Cats were anesthetized intraperitoneally with a mixture of urethane (400 mg/kg) and alpha-chloralose (40 mg/kg) and paralyzed with gallamine triethiodide (4 mg/kg, i.v. per hour). CNQX blocked the Glu induced pressor responses in FTG, DM and RVLM but potentiated the depressor responses in CVLM. These results suggest that non-NMDA receptors modulate the central pressor and depressor responses in an opposite direction. On the other hand, activation of DM and RVLM neurons by application of AMPA (5 mM, 30 nL) evoked pressor responses. These AMPA-induced responses were significantly blocked by CNQX. Interestingly, CNQX itself induced pressor responses in many stimulated points of the pressor areas (FTG: 6/9; DM: 13/24; RVLM: 6/13), indicating a tonic release of Glu mediating depressor effects. In conclusion, non-NMDA receptors within the pressor (FTG, DM and RVLM) and depressor (CVLM) areas may play different modulatory roles in cardiovascular integration. The depressor mechanism mediated by non-NMDA receptors is tonically activated by the release of endogenous Glu in these pressor and depressor areas. PMID- 10513606 TI - Cyclic strain induces redox changes in endothelial cells. AB - Our previous studies have shown that cyclic strain to endothelial cells (ECs) increases reactive oxygen species (ROS) that act as second messengers. The potential impact of these enhanced ROS levels on ECs was examined by studying the antioxidant activities and heme oxygenase-1 (HO-1) expression in strained ECs. Cyclic strain to ECs increased lipid peroxidation and augmented oxidation of low density lipoproteins. ECs subjected to strain increased their superoxide dismutase activities. Concomitantly, glutathione peroxidase activities increased in 3 to 6 hr and returned to basal level 24 hr after continuous cyclic strain treatment. A decrease of glutathione (GSH) was accompanied with an increase of oxidized glutathione (GSSH) level in ECs 3 to 6 hr after strain treatment. This was followed with a return of both GSH and GSSH to basal levels in 24 hr. Consistently, H2O2 treatment of ECs decreased the GSH/GSSG ratio. ECs pretreated with catalase abolished the strain-induced change in GSH/GSSG. Strain treatment, similar to H2O2 exposure, induced HO-1 expression in a time-dependent manner. This induction was inhibited after treating ECs with catalase or free radical scavenger. ECs treated with N-acetyl-cysteine abolished HO-1 gene induction. Our results suggest that cyclic strain-induced ROS cause a transient increase of glutathione peroxidase activity that results in a decrease of GSH level in ECs and that this decrease is crucial to HO-1 induction. PMID- 10513607 TI - Primary structure of rat spermidine synthase: an example of refining the cDNA derived amino acid sequence. AB - The primary structure of rat spermidine synthase having the N-terminal acetylated methionine and 98.7% homology with that of the mouse enzyme is presented using a limited amount of the homogeneous enzyme. The study strategy was principally to compare the molecular masses of liberated peptides determined by three specific cleavage methods with those expected from known cDNA-derived amino acid sequences of mouse and human enzymes using matrix-assisted laser desorption/ionization time of-flight mass spectrometry (MALDI TOF-MS). The cleavage methods involved two enzymatic methods using lysylendopeptidase and arginylendopeptidase, and a chemical method for cleaving at the cysteine residue using 2-nitro-5 thiocyanobenzoic acid. Their usefulness was clearly demonstrated. Column switching semimicro reversed-phase HPLC, which permits application of the entire reaction mixture, was useful for collecting a small amount of peptides containing the N-terminal amino acid, to confirm acetylation of the N-terminal methionine by MALDI TOF-MS. It was necessary in this approach to examine the amino acid sequence of certain peptides. The Edman method was used for the sequence analysis, and this will be replaced by an improved MALDI TOF-MS now available in a few laboratories. PMID- 10513608 TI - Detection of anaphylactic reaction in the percutaneously sensitized mouse using the AW method. AB - Anaphylactic reactions of mice sensitized percutaneously with 2,4 dinitrofluorobenzene (DNFB) were investigated by the AW method assay, which is a mouse anaphylactic model using the abdominal wall as the site for induction with either 2,4-dinitrophenyl (DNP)-human serum albumin or anti-mouse IgE antibody and then estimation of the response. DNP-specific and IgE-dependent anaphylactic reaction after contact sensitization with DNFB could be induced and detected by the abdominal wall (AW) method assay in both groups with and without previous ear challenge with DNFB. Thus, the anaphylactic reaction in the group of twice contact with 0.5% DNFB was observed on the 9th day from the sensitization (5th day from the ear challenge), and the reaction in the group of a single contact with 0.5% DNFB was observed 10 d after sensitization. The DNP-specific anaphylactic reaction was observed earlier than the 10th day with higher doses of DNFB. As for the mice of the former twice-contact group, the first and second characteristic ear swelling responses appeared within 1-6 h and 2 d of the ear challenge, respectively, and small swelling was observed 7 d after the challenge. It is suggested that Th1 and Th2 cells are activated at the almost same time, in other words, the preparation for both cell-mediated and humoral immunity could be accomplished to function, in vivo by a single percutaneous sensitization with DNFB. PMID- 10513609 TI - Increase in thioredoxin activity of intestinal epithelial cells mediated by oxidative stress. AB - Hydrogen peroxide was cytotoxic to the small intestine epithelial cell line, IEC 6, as judged from an MTT assay and the release of lactate dehydrogenase. The glutathione S-transferase and thioredoxin reductase activities and SH content decreased dose-dependently with H2O2, but thioredoxin activity increased at low H2O2 concentrations. In addition, the increase in thioredoxin activity was time dependent during the initial stages of oxidative stress. A reverse transcription polymerase chain reaction (RT-PCR) amplification also showed that the mRNA content in IEC-6 cells increased time-dependently at 0.25 mM H2O2. These results indicate that cellular oxidative shock causes an increase in the activity of thioredoxin, which is involved in the defense mechanism against oxidative stress. PMID- 10513610 TI - Identification of a high-copy-number plasmid suppressor of a lethal phenotype caused by mutant DnaA protein which has decreased intrinsic ATPase activity. AB - The induction of a mutant DnaA protein (DnaA E204Q) with decreased intrinsic ATPase activity in cells causes a lethal phenotype. Based on our results that the decreased ATPase activity of DnaA E204Q is activated to the level of a wild-type protein in the presence of partially purified stimulating factors for DnaA ATPase, we tested the hypothesis that genes encoding the stimulating factors can be cloned as high-copy-number plasmid suppressors for the lethality caused by DnaA E204Q. We isolated a number of high copy-number plasmids that suppress the lethal phenotype. The genes responsible for the suppression of the lethal phenotype were revealed to be dnaN, relA, pcnB and cyaA by subcloning, and a site directed mutational analysis. The mechanisms by which these genes suppressed the lethal phenotype were examined in vivo and in vitro. PMID- 10513611 TI - DNA sequence and expression of a defective mer operon from Pseudomonas K-62 plasmid pMR26. AB - pMRB01 cloned from Pseudomonas K-62 plasmid pMR26 conferred bacterial hypersensitivity to organomercurials. DNA sequence analysis of a 2.3-kb SacI Aor51HI fragment encompassing the whole region required for expression of the hypersensitive phenotype, revealed three open reading frames. The DNA sequence of these frames had 82.5%, 99.2% and 97.0% homology with the pDU1358 merR, merB and merD, respectively. The pMRB01 mer operon differs from the already known mer operon by the absence of the merT, merP and merA genes in this plasmid. An inverted repeat-like sequence upstream from the predicted merR was observed suggesting that this defective mer operon could be part of a transposon-like structure. Induction experiments and maxicell analysis of the mer-polypeptide showed that the lyase enzyme encoded by pMRB01 merB gene is mercurial-inducible and regulated by the transacting product of the merR gene. These results suggest that the hypersensitivity to organomercurials resulted from the expression of lyase activity encoded by the defective mer operon in the absence of reductase activity. The lyase enzyme encoded by pMRB01 merB catalyzes the protonolysis of the C-Hg bond of both arylmercury and alkylmercury compounds. PMID- 10513612 TI - Analysis of the disruption mutant of the oscillin homolog gene of Dictyostelium discoideum. AB - A homolog of oscillin, the Ca2+ oscillation-inducing factor of the hamster, was identified from the cellular slime mold Dictyostelium discoideum and designated Dd-oscillin. In the developmental stages of D. discoideum, the gene is expressed at the prestalk region which contains a higher concentration of cytosolic Ca2+ than the prespore region. The Dd-oscillin null strain aggregated but did not develop further when the cells were plated on non-nutrient agar at a density of 1.5x10(6) cells/cm2, showing that the Dd-oscillin gene is important for development. Since the null cells carrying the hamster oscillin gene formed fruiting body, the hamster oscillin was the homolog of Dd-oscillin as far as function is concerned. In addition, the null cells formed fruiting body in the presence of 2,5-di(tert-butyl)-1,4-hydroquinone (BHQ: a specific inhibitor of Ca2+-ATPase activity in the endoplasmic reticulum). These results suggest that Dd oscillin will increase cytosolic Ca2+ in the cells and promote further development. PMID- 10513613 TI - Suppressive effect of norzoanthamine hydrochloride on experimental osteoporosis in ovariectomized mice. AB - Norzoanthamine is an alkaloid isolated from a colonial zoanthid. We examined the effects of norzoanthamine hydrochloride on bone weight, strength and morphology in ovariectomized mice, a postmenopausal osteoporosis model. Norzoanthamine hydrochloride significantly suppressed the decrease in femoral weight and bone biomechanical parameters caused by ovariectomy without an increase in uterine weight. This means that norzoanthamine hydrochloride does not have an estrogen like effect on reproductive organs. Morphological observations of longitudinally ground sections of the humeri showed that norzoanthamine hydrochloride administration (2 mg/kg/d, p.o.) completely suppressed the loss of trabecular bone. Furthermore, norzoanthamine hydrochloride thickened the cortical bone. Based on these results, norzoanthamine hydrochloride may act as both a suppressor of bone resorption and an enhancer of bone formation in vivo. PMID- 10513614 TI - The role of thunberginol A, an isocoumarin constituent of Hydrangeae Dulcis Folium, on the signal transmission pathway for rat mast cell degranulation. AB - The role of the signal transmission pathway of thunberginol A (TA) in mast cell degranulation was examined using rat peritoneal mast cells. First of all, we investigated the cellular distribution of TA using fluorescent microscopy. This indicated that TA is immediately incorporated into cells and distributes in cytosol around the nucleus. We then investigated the effect of TA on mast cell protein tyrosine phosphorylation, which is part of the signal transduction cascade for degranulation. TA non-specifically inhibited the tyrosine phosphorylation induced by compound 48/80 (Co. 48/80), at 10 to 100 microM, and orthovanadate/hydrogen peroxide at more than 50 microM in a dose-dependent manner. As far as the intracellular Ca2+ change in fluo-3-loaded cells was concerned, TA (10 microM) suppressed the rise in Ca2+ induced by antigens, ionomycin and thapsigargin, while TA did not suppress the rise induced by Co. 48/80. This evidence suggests that TA mainly inhibits extracellular Ca2+ influx, but TA does not act on the intracellular Ca2+ mobilization from the endoplasmic reticulum. We also investigated the influence of TA on the cytoskeleton and membrane changes using mast cells and erythrocytes. TA (10 microM) inhibited the cytoskeletal assembly formation in dicyanovinyl julolidin-loaded mast cells induced by Co. 48/80. Moreover, TA suppressed the hypotonic hemolysis of erythrocytes, from 3 to 1000 microM, in a dose-dependent manner. These results suggest that inhibition of protein tyrosine phosphorylation, extracellular Ca2+ influx and cytoskeletal assembly formation, and membrane stabilization are involved in the inhibitory effect of TA in mast cell degranulation. PMID- 10513615 TI - Phagocytic activity of ethyl alcohol fraction of deer antler in murine peritoneal macrophage. AB - The mechanism of phagocytic activity of the ethyl alcohol fraction of Cervus nippon (CN-E) was investigated in vivo. The administration of CN-E (100 mg/kg, p.o.) enhanced lucigenin chemiluminescence and the engulfment of fluorescein conjugated E. coli particles in murine peritoneal macrophages. Phagocytic activity was suppressed by the treatment of S-nitrosoglutathione (GSNO) which is an exogenous nitric oxide donor depending on the concentration of dose. CN-E suppressed the production of nitric oxide and enhanced the concentration in [Ca2+]i. The enhancement in [Ca2+]i was diminished by the treatment of EGTA. These results indicate that CN-E enhances the phagocytic activity of murine peritoneal macrophage via a suppression of nitric oxide production and an increase in [Ca2+]i. PMID- 10513616 TI - Dipeptides containing L-arginine analogs: new isozyme-selective inhibitors of nitric oxide synthase. AB - Several L-arginine analogs are known as potent inhibitors of nitric oxide synthase (NOS). We recently synthesized dipeptides containing such amino acids, and found that they are potent and isozyme-selective NOS inhibitors. For example, S-methyl-L-isothiocitrullinyl-L-phenylalanine showed 66-fold selectivity for iNOS over nNOS, while S-methyl-L-isothiocitrullinyl-L-leucine and N(G)-nitro-L argininyl-L-phenylalanine showed 20- and 14-fold selectivity, respectively. Interestingly, S-methyl-L-isothiocitrullinyl-D-phenylalanine showed no selectivity, and S-methyl-L-isothiocitrullinyl-L-phenylalanine showed competitive inhibition. These results suggest that each NOS isozyme has a cavity of different size near the C-terminal of the L-arginine binding site, and that the selectivity of inhibitors is due to the differences in the size of the cavity. PMID- 10513617 TI - Changes in lactate dehydrogenase and 3-hydroxyacetyl-CoA dehydrogenase activities in rat skeletal muscle by the administration of Eucommia ulmoides OLIVER leaf with spontaneous running-training. AB - We examined the effect of Eucommia ulmoides OLIVER leaf on rat skeletal muscles together with spontaneous running-training in terms of the isozyme profile and specific activity of lactate dehydrogenase (LDH; EC 1.1.1.27) and 3-hydroxyacetyl CoA dehydrogenase (HAD; EC 1.1.1.35). On the twenty-ninth day of the experimental period, a mandatory endurance running exercise (treadmill, 7 degrees grade) was conducted. Twenty-four hours later, the rats were sacrificed and the skeletal muscles and other organs were dissected. Due to the training, the HAD specific activity in the skeletal muscles had increased and a more oxidative metabolism had developed, which was further enhanced by the administration of the leaf. In soleus (SOL) muscle in the Eucommia leaf treated running-training group (ET), the LDH specific activity in the skeletal muscle was significantly higher than in the sedentary control group (SC). The isozyme profile of the group ET was significantly different when compared with the group SC. The changes in the LDH isozyme profile were larger in the SOL than that in extensor digitorum longus (EDL) muscle. The results show that mechanical training and the use of the leaf cooperatively increase the ability to avoid lactate accumulation in skeletal muscle. This effect is supported by the group where 67% of rats accomplished the endurance running exercise. Theses results suggest that the administration of Eucommia ulmoides OLIVER leaf along with light intensity training enhances the ability of a muscle to resist fatigue. PMID- 10513618 TI - Molecular cloning and characterization of two cDNAs for Glycyrrhiza glabra squalene synthase. AB - Two cDNAs (GgSQS1 and GgSQS2) encoding squalene synthase have been isolated from licorice, Glycyrrhiza glabra L., and characterized. The deduced amino acid sequence of GgSQS1 was 88%, 81%, 78%, 45-44%, and 45-41% identical to those of GgSQS2, Nicotiana, Arabidopsis, mammal and yeast squalene synthases, respectively. Squalene synthase activity was found in the cell-free extracts of Escherichia coli transformed with the recombinant plasmids for GgSQS1 and GgSQS2, respectively. Genomic Southern blot hybridization indicated that there are three squalene synthase genes in the licorice genome. Northern blot analysis showed that GgSQS2 mRNA is mainly expressed during the exponential growth phase of the cultured licorice cells. PMID- 10513619 TI - Glycyrrhizin inhibits TNF-induced, but not Fas-mediated, apoptosis in the human hepatoblastoma line HepG2. AB - To determine the transaminase-lowering action of glycyrrhizin (GL) immunologically, the effect of GL on tumor necrosis factor (TNF)-alpha- and Fas mediated apoptosis was assessed using a human hepatoblastoma line, HepG2 cells. The HepG2 cells were resistant to TNF-alpha and anti-Fas antibody, but were rendered susceptible to TNF-alpha and anti-Fas antibody in the presence of actinomycin D (Act D), an inhibitor of RNA synthesis. The cytotoxicity induced by TNF-alpha/Act D or anti-Fas/Act D was accompanied by DNA fragmentation, indicating apoptotic death of HepG2 cells. GL partially prevented the apoptosis of HepG2 cells induced by TNF-alpha/Act D in a GL-dose dependent fashion. However, this protective effect of GL was not observed in the cytotoxicity of HepG2 caused by anti-Fas/Act D. Although the protection mechanism of GL, observed in a limited fashion against TNF-alpha-mediated apoptosis, is unclear, the present results provide an immunological explanation for the transaminase lowering action of GL in the GL treatment of chronic liver diseases involving apoptotic hepatocyte death in their pathogenesis. PMID- 10513620 TI - Drug absorption behavior after periocular injections. AB - The purpose of this study was to investigate the absorption behavior of an ophthalmic drug injected in rabbit periocular tissues. After intracapsular, retrobulbar and palpebral conjunctival injections of 150 microl and 50 microl fluorescein isothiocyanate dextran (FITC-dextran, average molecular weight 11000), leakage of the dye into the tear fluid was dependent on the injection route and volume. After periocular injections (50 microl) of tilisolol, as a model beta-blocker, the concentrations in the tear fluid, blood, aqueous humor and vitreous body were determined by HPLC. Slight drug leakage was observed in the tear fluid after injections. The periocular injections showed a faster absorption and a higher area under the concentration-time curve (AUC) in the plasma and a lower AUC in the aqueous humor than those observed in instillation. They also showed a higher ratio of AUC of tilisolol in the vitreous body to AUC in the aqueous humor than that observed in the instillation. Among the periocular injections, retrobulbar injection showed the highest concentrations in the plasma and the lowest in the aqueous humor and vitreous body, while intracapsular injection showed the lowest in the plasma and the highest in the aqueous humor and vitreous body. Although the periocular injections showed a rapid systemic absorption of drug by a rich topical vasculature, it might be an effective approach to deliver the drug to the periocular tissues and vitreous body. PMID- 10513621 TI - Topical delivery system of ophthalmic drugs by periocular injection with viscous solution. AB - The purpose of this study is to evaluate periocular injections with viscous solution as a topical delivery system of ophthalmic drugs. Tilisolol and carboxymethylcellulose (CMC) were used as a model beta-blocker and a viscous polymer, respectively. After intracapsular, retrobulbar and palpebral conjunctival injections (50 microl) of tilisolol with 3% CMC into rabbits, drug concentrations in the tear fluid, blood, aqueous humor and vitreous body were determined by HPLC. Periocular injection (50 microl) of tilisolol with 3% CMC showed slight leakage of the drug in the tear fluid from the injection site. The viscous vehicle decreased the absorption rate constant of the drug from the injection site to systemic circulation compared with the buffer solution. It suggests that the viscous solution improved the retention of drug at both the injection site and in periocular tissues. Although the periocular injections with viscous vehicle (3% CMC) showed lower AUC in the aqueous humor than that observed in instillation, they showed comparable AUC in the vitreous humor. Compared to the results after the periocular injections with buffer solution, CMC increased the AUCs in the vitreous body 3.1-fold with retrobulbar injection and 1.4-fold with palpebral conjunctival injection, respectively. As a result, periocular injections with 3% CMC showed higher delivery of tilisolol to the vitreous body against the aqueous humor than the instillation and periocular injections with buffer solution. PMID- 10513622 TI - Structural features and hypoglycemic activity of a polysaccharide (CS-F10) from the cultured mycelium of Cordyceps sinensis. AB - A polysaccharide (CS-F10) purified from a hot water extract of the cultured mycelium of Cordyceps sinensis was composed of galactose, glucose and mannose in a molar ratio of 43:33:24; its molecular weight was estimated to be about 15000. The results of chemical and spectroscopic investigations suggest that CS-F10 has a comb-type structure, and has alpha-D-glucopyranosyl residues on the terminal of the side-chains and characteristic sugar residues of C. sinensis i.e., 1,5-linked beta-D-galactofuranosyl residues. CS-F10 significantly lowered the plasma glucose level in normal, streptozotocin (STZ)-induced diabetic and epinephrine-induced hyperglycemic mice after intraperitoneal administration (50 mg/kg). Administration of CS-F10 to STZ-induced diabetic mice significantly increased the activity of hepatic glucokinase. A significant reduction in the hepatic glucose output was observed following the infusion of CS-F10 using the perfused rat liver. CS-F10 also significantly decreased protein content of facilitative glucose transporter isoform 2 (GLUT2) from rat liver following i.p. administration. These effects presumably contribute to the hypoglycemic activity. PMID- 10513623 TI - In vitro inhibitory effects of the optical isomers and metabolites of fluvastatin on copper ion-induced LDL oxidation. AB - Fluvastatin is a synthetic hypolipidemic drug which inhibits 3-hydroxy-3 methylglutaryl coenzyme A (HMG-CoA) reductase. We compared in vitro the antioxidative effects of two enantiomers (3R, 5S and 3S, 5R) of fluvastatin, which is clinically used as a racemic mixture, on copper ion-induced oxidation of human low-density lipoprotein (LDL). Although 3R,5S-enantiomer of fluvastatin has 30-fold stronger inhibitory activity on HMG-CoA reductase than its optical counterpart, the antioxidative effects of these enantiomers on copper ion-induced LDL oxidation were similar. The antioxidative effects of the metabolites of fluvastatin (M2, M3, M4 and M7) on the copper ion-induced LDL oxidation were also investigated. All the metabolites tested showed an inhibitory effect on this system. Among them, the effects of M2 and M3, which have a phenolic hydroxyl group in each indole moiety, were strong and their potencies were 30-50 times greater than that of fluvastatin. We conclude that not only 3R,5S-enantiomer of fluvastatin but also its optical counterpart and the metabolites also have a potential to show the anti-atherosclerotic effect through their antioxidative activities on lipid peroxidation. PMID- 10513624 TI - Determination of valproic acid in human serum by high-performance liquid chromatography with fluorescence detection. AB - A simple and highly sensitive high-performance liquid chromatographic method is described for the determination of valproic acid in human serum. The method is based on the direct derivatization of serum sample with 6,7-methylenedioxy-1 methyl-2-oxo-1,2-dihydroquinoxaline-3-ylpr opionohydrazide. The derivatization reaction proceeds in aqueous solution in the presence of pyridine and 1-ethyl-3 (3-dimethylaminopropyl)carbodiimide at 37 degrees C. The resulting derivatives were separated on a reversed-phase column (YMC Pack ODS-A) with isocratic elution and were fluorimetrically detected at 440 nm with excitation at 365 nm. The detection limit (signal-to-noise ratio=3) for valproic acid added to serum sample was 0.1 microg (700 fmol)/ml serum sample (2.3 fmol on column). The method was applied to determine the unbound- and total-valproic acid levels in the serum obtained from three healthy volunteers after oral administration of the drug (600 mg). PMID- 10513625 TI - Application of calcein-loaded liposomes for the determination of membrane channel size. AB - We found that calcein-loaded liposomes can be used to evaluate the sizes of channels in membranes by measuring changes in calcein release when the molecular size of the solute added to the outer suspension medium is changed. If the solute added to the outer medium can enter the inner aqueous phase through the channel, the osmotic pressure of the inner phase increases, causing bursting of the liposomes and the release of calcein. Thus, the size of the channel formed in the liposomal membrane can be determined by examining whether the solute added to the outer medium induces calcein release. We used a series of sugars as solutes and estimated the channel size formed by polyene antibiotics. The results agreed well with those conducted in a similar manner using erythrocytes, demonstrating that this method should be useful for examining channel sizes in membranes. PMID- 10513626 TI - Effects of development on acetoacetyl-CoA synthetase biosynthesis in rat liver. AB - In order to investigate the physiological role of acetoacetyl-CoA synthetase (acetoacetate-CoA ligase, EC 6.2.1.16), a cytosolic acetoacetate-activating enzyme, the effects of animal development on the activity and content of the enzyme were examined in rat liver. In male rats, the enzyme specific activity increased 21-fold at 4 weeks of age from that at 2 weeks of age, and then gradually decreased, while in female rats, it increased similarly to that of male rats, but further increased, reaching a maximum about 3-fold higher than that of male rats, at 6 weeks of age. The developmental patterns of the enzyme content correlated with that of the enzyme specific activity. These results indicate that changes in this enzyme activity and content during the developmental process might influence the rate of ketone body utilization for the formation of physiologically important lipidic substances in rat liver. PMID- 10513627 TI - Relaxant effects of pyranocoumarin compounds isolated from a Chinese medical plant, Bai-Hua Qian-Hu, on isolated rabbit tracheas and pulmonary arteries. AB - Qian-Hu is a well-known traditional Chinese medicine used for the treatment of respiratory diseases and pulmonary hypertension. We compared the relaxant effects of pyranocoumarin compounds, including (+)-praeruptorin A (Pra-C), Pd-Ia (=(+/-) praeruptorin A), pteryxin, peucedanocoumarin II (P-II) and 8-methoxy-psoralen (8 MOP) purified from Bai-Hua Qian-Hu (BQ) in isolated rabbit tracheas and pulmonary arteries. Pra-C, pteryxin and Pd-Ia produced significant relaxant effects in tracheal preparations constricted with 40 mM KCl or 10 microM acetylcholine. The relaxant response to Pra-C, pteryxin or Pd-Ia in preparations constricted with KCl was significantly more potent than that in preparations constricted with acetylcholine. Pra-C, pteryxin or Pd-Ia at a concentration of 30 microM completely relaxed tracheas constricted with 40 mM KCl, whereas P-II at the same concentration showed only partial relaxation. In pulmonary arterial preparations, 8-MOP produced a significant relaxant effect on contractions by 10 microM phenylephrine, without any effect on the contraction by 40 mM KCl. These results suggest that Pd-Ia, pteryxin and Pra-C for their calcium antagonistic action, and 8-MOP for its inhibitory effect on contraction induced by phenylephrine, may be the active principles of BQ for relaxing the smooth muscle of tracheas and pulmonary arteries, and the principle may produce a synergistic effect. PMID- 10513628 TI - New model of progressive non-insulin-dependent diabetes mellitus in mice induced by streptozotocin. AB - This study was designed to clarify the relationship between streptozotocin (STZ) dosage (100, 150 and 200 mg/kg i.p.) and the resulting diabetogenic response in mice (8-week-old male ICR). In this experiment, we found that a single i.p. injection of 100 mg/kg STZ is able to induce progressive diabetes mellitus, in which non-fasting serum glucose levels begin to rise from 3 weeks and continue to rise throughout the experimental period until 9 weeks. The non-fasting serum insulin levels of 100 mg/kg STZ-treated mice were normal during the experimental period. In addition, the population of insulin-immunoreactive cells (beta cells) in the islets of pancreata was slightly less than in normal mice at 9 weeks. In 200 mg/kg STZ-treated mice, on the other hand, the insulin levels were below measurable values and insulin-immunoreactive cells were not observed. It is concluded from these results that the progressive diabetic mouse model induced by a single i.p. injection of 100 mg/kg STZ, unlike 200 mg/kg STZ-induced diabetes which is insulin-dependent, is non-insulin-dependent. PMID- 10513629 TI - Antimicrobial activity and metalloprotease inhibition of hinokitiol-related compounds, the constituents of Thujopsis dolabrata S. and Z. hondai MAK. AB - Gamma-thujaplicin, beta-dolabrin and hinokitiol(beta-thujaplicin), hinokitiol related compounds isolated from the wood of Thujopsis dolabrata S. and Z. hondai MAK have antimicrobial activity. In particular, strong antibacterial activity of hinokitiol and beta-dolabrin on Staphylococcus epidermidis IFO-12993 was found, with a minimum inhibitory concentration (MIC) of 0.2 microg/ml. This activity was higher than that of gentamicin, used as a positive control, and so the strong antibacterial activity of both compounds on this bacterium is of considerable interest. Of the three compounds, gamma-thujaplicin showed the strongest antifungal activity and its MIC was found to be around 1.5 microg/ml. The three compounds also inhibited metalloproteases. The inhibitory activity of hinokitiol on carboxypeptidase A was especially strong, its 50%-inhibitory concentration (IC50) being 2.76x10(-6) M. Considering that metalloproteases are involved in inflammation, the strong inhibitory activity of hinokitiol could be important. On the other hand, hinokitiol-acetate did not show any antimicrobial activity and metalloprotease inhibition, suggesting that at least part of the activity is due to metal chelation between the carbonyl group at C-1 and the hydroxyl group at C 2 in the tropolone skeleton. PMID- 10513630 TI - Repression of acyl-CoA:cholesterol acyltransferase by a traditional herbal medicine (Kampo medicine), Ogi-Keishi-Gomotsu-To-Ka-Kojin. AB - Ogi-Keishi-Gomotsu-To-Ka-Kojin (OKGK) is a traditional herbal medicine (Kampo medicine) which has been found to ameliorate hypercholesterolemia and hypertriglyceridemia in rats and rabbits. In the present study, the effect of OKGK on acyl-CoA:cholesterol acyltransferase (ACAT) was studied in order to elucidate the mechanism of its antihypercholesterolemic action. Oral administration of OKGK to rats fed a cholesterol-enriched diet for 4 weeks markedly repressed the increase in ACAT activity in the small intestine. In contrast, OKGK did not influence hepatic ACAT activity. These results indicate that OKGK selectively inhibits ACAT activity in the small intestine relative to that in the liver, resulting in a reduction of cholesterol absorption, followed by a decrease in serum cholesterol. PMID- 10513631 TI - The marked inhibition of the bitter taste of Polymyxin B sulfate and trimethoprim x sulfamethoxazole by flavored BMI-60 in pediatric patients. AB - Taste acceptability of ground Polymyxin B sulfate and Bactramin C tablets was examined when flavored BMI-60, a food additive, was added. Both adult and child volunteers found the bitter taste of the two drugs markedly inhibited, making it clinically useful. Noncompliance, due to this bitterness, was improved using flavored BMI-60. The most striking characteristic of flavored BMI-60 is the ease of preparation compared with the manufacture of other hospital pharmaceuticals such as jelly, gummi and candy done to mask bitterness. PMID- 10513632 TI - Immobilization of tripeptide growth factor glycyl-L-histidyl-L-lysine on poly(vinylalcohol)-quarternized stilbazole (PVA-SbQ) and its use as a ligand for hepatocyte attachment. AB - A tripeptide growth factor, glycyl-L-histidyl-L-lysine (GHK), was immobilized on the surface of poly(vinylalcohol)-quarternized stilbazole (PVA-SbQ) gel. The photoreactive substance, 4-(3-trifluoromethylazirino)benzoyl-N-hydroxysuccinimide (TDBA-OSu), was employed to link the gel and ligand GHK. The density of immobilized GHK was 70 nmol/cm2. Isolated rat hepatocytes were inoculated on the GHK-immobilized PVA-SbQ gel and cultured for 5 d. About 24 h after inoculation, hepatocytes started to aggregate and formed multicellular spheroids while almost no cells attached to GHK-non-immobilized PVA-SbQ gel. The formed spheroids attached firmly to the surface of PVA-SbQ gel for 5 d. GHK was, thus, shown to be an effective ligand for hepatocyte attachment. Dodecamethylenediamine was used to extend the length between the gel surface and GHK. Extension of the length significantly increased the number of attached hepatocytes. PMID- 10513633 TI - The effects of interleukin (IL)-4 and IL-10 on macrophage growth-stimulating activities of oxidized low density lipoprotein (LDL), acetylated LDL and macrophage colony-stimulating factor: the activity of oxidized LDL is refractory to the inhibitory cytokines. AB - We have previously reported that ligands of scavenger receptor such as acetylated low density lipoprotein (acetyl-LDL) and oxidized LDL induced growth of peripheral macrophages in vitro. This suggests the possibility that in addition to foam cell formation, modified or oxidized LDLs induce macrophage proliferation in atherosclerotic lesions. To learn further the physiological regulation of macrophage growth, we comparatively examined the effect of interleukin (IL)-4 and IL-10 which have been reported to be suppressive to various macrophage functions on macrophage growth-stimulating activities of the acetyl-LDL, oxidized LDL and macrophage colony-stimulating factor (M-CSF). An in vitro study showed that the activity of M-CSF-containing L-cell-conditioned medium was the most sensitive to the suppressive effects of these cytokines. The growth-inducing activity of acetyl-LDL was significantly inhibited by both IL-4 and IL-10. On the other hand, the activity of oxidized LDL was not attenuated by IL-4 or IL-10. These data indicate that macrophage growth-stimulating activity of oxidized LDL, in contrast to that of M-CSF or acetyl-LDL, is refractory to these suppressive cytokines. Oxidized LDL may act as a potent macrophage growth-stimulating factor in atherosclerotic or other inflammatory sites, even when these cytokines are produced by inflammatory and immunological reactions in situ. PMID- 10513634 TI - Arginyl-tRNA-protein transferase activities in crude supernatants of rat tissues. AB - A fluorescent HPLC method for the assay of arginyl-tRNA-protein transferase (R Transferase) activity was applied to obtain quantitative data of the enzyme activity in rat tissues for the first time. In this assay, the major problem was a significant hydrolysis of the substrate, N-aspartyl-N'-dansylamido-1,4 butanediamine, and the product, N-arginylaspartyl-N'-dansylamido-1,4 butanediamine (ArgAsp(4)DNS) by aminopeptidases in crude samples such as 105000g supernatants (105S) of tissue homogenates. As bestatin inhibited the hydrolysis of ArgAsp(4)DNS, a standard-addition method in the presence of bestatin, using a partially purified R-Transferase preparation from hog kidney as a standard, made it possible to measure directly R-Transferase activities in 105S with a short incubation time and sufficient reliability. It was found by the established method that of 14 tissues examined, stomach was rich in the R-Transferase activity with the highest specific activity, suggesting a target tissue for the future studies on R-Transferase to elucidate its physiological significance. PMID- 10513635 TI - Potent homophthalimide-type inhibitors of B16F10/L5 mouse melanoma cell invasion. AB - Recently, we developed a series of novel and potent aminopeptidase inhibitors with a homophthalimide skeleton. Among them, N-(2,6-diethylphenyl)homophthalimide (PIQ-22) possesses a specific aminopeptidase-inhibiting activity more potent than that of bestatin or actinonin, as assayed in terms of hydrolysis of L-alanine 4 methylcoumaryl-7-amide (Ala-AMC) by human acute lymphoblastic leukemia MOLT-4 cells. We show here that PIQ-22 and its 2,6-dimethylphenyl derivative (PIQ-11) are more potent inhibitors of tumor cell invasion than bestatin and actinonin in a Matrigel assay using mouse melanoma B16F10/L5 cells. PMID- 10513636 TI - Chemotherapy with hybrid liposomes for melanomatosis. AB - The inhibitory effects of the hybrid liposomes on the growth of B-16 melanoma cells in vitro and in vivo were examined. The 50% inhibitory concentration of the hybrid liposomes composed of 90 mol% dimyristoylphosphatidylcholine (DMPC) and 10 mol% polyoxyethylenedodecyl ether (C12(EO)10) was one-twelfth of that of DMPC liposomes. It was noteworthy that for the first time significantly prolonged survival was obtained using a mouce model of carcinoma after the administration of the hybrid liposomes of 90 mol% DMPC/10 mol% C12(EO)n (n=10 or 23) without antitumor drugs. PMID- 10513638 TI - Effectiveness of methadone treatment in reducing HIV risk behavior and HIV seroconversion among injecting drug users. PMID- 10513637 TI - The steady-state plasma pharmacokinetics of indinavir alone and in combination with a low dose of ritonavir in twice daily dosing regimens in HIV-1-infected individuals. AB - OBJECTIVE: To explore the steady-state plasma pharmacokinetics of indinavir in twice daily dosing regimens with and without the co-administration of 100 mg ritonavir. DESIGN: Observational pharmacokinetic study. PATIENTS: HIV-1-infected individuals who use indinavir alone (1200 mg twice daily, n = 6), or the combination of 100 mg ritonavir twice daily plus either 800 mg (n = 6), or 1200 mg indinavir twice daily (n = 2). METHODS: Steady-state pharmacokinetics of indinavir and ritonavir were assessed by drawing 12 blood samples during an 8-h period after ingestion of the medication. RESULTS: Significant differences were observed for indinavir pharmacokinetics between the dosing regimens indinavir 1200 mg twice daily alone and indinavir/ ritonavir 800/100 mg twice daily with respect to the mean trough concentration (0.21 and 0.99 microg/ml, respectively, P = 0.002), the mean maximum concentration (13.79 and 8.74 microg/ml, respectively, P = 0.028), and for the mean plasma elimination half-life (1.6 and 3.2 h, respectively, P = 0.001). The combination indinavir/ritonavir 1200/100 mg twice daily led to very high exposure to indinavir and was not well tolerated. However, the combination indinavir/ritonavir 800/100 mg twice daily was well tolerated and resulted in therapeutic concentrations of indinavir with improved trough concentrations and similar maximum concentrations as observed with the licensed dosage of 800 mg three times daily. CONCLUSION: Combination of indinavir and 100 mg ritonavir in twice daily dosing regimens significantly affects the pharmacokinetic profile of indinavir. The results of this observational study provide a pharmacologic basis for the combination of indinavir (800 mg) and ritonavir (100 mg) in twice daily dosing regimens. PMID- 10513639 TI - Characterization of subtype A HIV-1 from Africa by full genome sequencing. AB - OBJECTIVE: To improve our understanding of the genetic complexity of HIV-1 subtype A by increasing the number of subtype A isolates that have been sequenced in their entirety. METHODS: Nine HIV-1-seropositive patients from Africa living in Sweden contributed peripheral blood mononuclear cells (PBMC) for this study. Sequencing of the C2-V3 region of env had shown them to be subtype A. DNA from virus cultures was used for the amplification of virtually full-length proviral sequences, and the resulting fragment was sequenced. RESULTS: Six of the nine viral isolates were subtype A throughout the genome, or non-recombinant, and all of these were from east Africa. One virus from the Ivory Coast had the AG(IbNG) genetic form, a recombinant form common in west Africa. Two of the isolates were novel recombinants: one was an A/C recombinant and the other was A/D. Analysis of gag reveals three subclusters within the A subtype: one containing the AG(IbNG) subtype viruses, one containing the AE(CM240) viruses and one containing the non recombinant A viruses. These genetic clusters have different geographical distributions in Africa. CONCLUSION: The prevailing view of HIV-1 subtype A forming a uniform band across the center of sub-Saharan Africa needs revision. In all probability, the most common subtype in west Africa and west central Africa is the AG recombinant, AG(IbNG), whereas in east central Africa it is the non recombinant subtype A. PMID- 10513640 TI - Both necrosis and apoptosis contribute to HIV-1-induced killing of CD4 cells. AB - BACKGROUND: Data currently available on HIV-1-induced cytopathology is unclear regarding the mechanism of cell killing. OBJECTIVE: To clarify the extent to which apoptosis or necrosis is involved in HIV-1-induced cell death in view of conflicting existing data. METHODS: T lymphoblastoid cells or peripheral blood mononuclear cells were infected by various strains of HIV-1 and the numbers of apoptotic or necrotic cells were quantified at various times after infection using video-image analysis techniques; the results were compared with the amount of fragmented DNA using a quantitative method. Measurement of mitochondrial transmembrane potential (deltapsi(m)) and intracellular calcium concentrations [Ca2+]i was performed with fluorescent probes and fluorescence concentration analysis (FCA). RESULTS: Although lymphoblastoid and monocytoid cells acutely infected by HIV-1 had increased levels of fragmented DNA, a marker of apoptotic cell death, few (<12%) had condensed chromatin and fragmented nuclei, the morphological features of apoptosis. The predominant alterations in acutely infected cells were distended endoplasmic reticulum and abnormal mitochondria; these ultrastructural changes are consistent with necrosis, although some infected cells simultaneously displayed features of both necrosis and apoptosis. Viability of cells persistently infected by HIV-1 was only minimally reduced from that of uninfected cells. This reduction was accounted for by an increased propensity of the persistently infected cells to die by apoptosis. Alterations in [Ca2+]i and deltapsi(m) occurred in both acutely and persistently infected cells. CONCLUSION: Both necrosis and apoptosis contribute to HIV-1-induced killing of CD4 cells. PMID- 10513641 TI - Strong correlation between the complement-mediated antibody-dependent enhancement of HIV-1 infection and plasma viral load. AB - OBJECTIVE: We have previously demonstrated that complement-mediated antibody dependent enhancement (C-ADE) of HIV-1 infection correlates with accelerated immunosuppression and disease progression in HIV-1-infected individuals. In the present work the relationship between C-ADE and plasma HIV-1 RNA concentrations was studied to determine the effect of C-ADE on viral replication. METHODS: Three studies were performed: (a) C-ADE and HIV-1 RNA concentrations were determined in the serum and plasma aliquots taken at the same time from 98 HIV patients, mostly in the advanced stage of the disease; (b) the above two parameters as well as HIV enzyme-linked immunosorbent assay (ELISA)-reactive antibodies (Abbott HIV 1/2 test), and p24 antigen levels (Abbott antigen test; Abbott, Delkenheim, Germany) were determined in four seroconversion panels purchased from the Boston Biomedica firm; (c) changes of HIV-1 RNA concentration and C-ADE during a 17 month follow up period were determined in 18 HIV-infected patients. C-ADE was measured by the method previously established in our laboratories. The results were expressed by an enhancement/neutralization index (E/NI). HIV-1 RNA levels were determined with the Amplicor monitor kit (Roche, Basel, Switzerland), and in some experiments with the nucleic acid sequence based amplification (Organon Teknika, Turnhout, Belgium) kits. RESULTS: (a) We found a highly significant (P<0.0001) positive correlation between E/NI values reflecting the extent of HIV-1 infection enhancement and plasma HIV-1 RNA levels. Both E/NI and HIV-1 RNA levels negatively correlated to the CD4 cell counts. (b) C-ADE was first detected just before, or concomitantly with, seroconversion in 4/4 seroconversion panels. (c) Both E/NI values and HIV-1 RNA levels significantly (P<0.001) increased during a 17 month observation period in 18 HIV-infected patients. CONCLUSION: We found strong association between the extent of the complement-mediated antibody dependent enhancement of HIV-1 infection and the plasma viral load in HIV patients. On the basis of these findings, C-ADE correlates with HIV replication in vivo, and potentially contributes to the progression of HIV disease. PMID- 10513642 TI - Human herpesvirus 8-encoded interleukin 6 activates HIV-1 in the U1 monocytic cell line. AB - OBJECTIVE: Human herpesvirus 8 (HHV-8) encodes a viral interleukin 6 (vIL-6) which is structurally and functionally similar to human interleukin 6 (hIL-6). Since hIL-6 has been shown to upregulate the expression of HIV-1, the objectives of this study were to examine the ability of vIL-6 to upregulate HIV-1, and to determine the interactions of this virokine (viral cytokine) with the components of the interleukin 6 (IL-6) receptor complex. DESIGN AND METHODS: Recombinant HHV 8 vIL-6 (rvIL-6) was assayed for bioactivity in the IL-6-dependent cell line MH60.BSF2. HIV-1 p24 production by the U1 monocytic and ACH-2 T-cell lines, which are chronically infected with HIV-1, was used to assess the ability of vIL-6 to affect HIV-1 expression. hIL-6 and vIL-6 receptor utilization was determined by quantifying HIV-1 p24 production after neutralization of components of the IL-6 receptor complex, CD126'IL-6R' and CD130'gp130', on U1 cells with blocking antibodies. RESULTS: HHV-8 rvIL-6 was seen to have IL-6-like bioactivity in MH60.BSF2 cells, and readily upregulated HIV-1 p24 production in U1 monocytic cells, but not in ACH-2 T cells. The vIL-6 appeared to utilize the IL-6-specific component of the IL-6 signaling complex, CD126'IL-6R', in U1 cells. CONCLUSIONS: HHV-8 vIL-6 clearly has the potential to upregulate HIV-1 expression in monocytic cells, and therefore may play a role in AIDS pathogenesis in individuals infected with both viruses. PMID- 10513643 TI - HAART in HIV-infected patients: restoration of antigen-specific CD4 T-cell responses in vitro is correlated with CD4 memory T-cell reconstitution, whereas improvement in delayed type hypersensitivity is related to a decrease in viraemia. AB - OBJECTIVE: To analyse prospectively the effect of highly active antiretroviral treatment (HAART) on CD4 T-cell responses in vitro and in vivo in HIV-infected patients. DESIGN: Prospective study with 49 protease inhibitor-naive adult patients. Data were collected at baseline and after 3 and 6 months of HAART. METHODS: In vitro CD4 T-cell reactivity was analysed by stimulation of peripheral blood mononuclear cells with several antigens. In vivo CD4 T-cell reactivity (delayed type hypersensitivity) was assessed by Multitest Merieux. Both measurements were correlated to CD4 (memory) T-cell count and HIV-1 viraemia. RESULTS: Restoration of specific CD4 T-cell proliferation was observed in most patients. The in vitro T-cell response was restored more frequently against antigens to which the immune system is constantly exposed (Candida albicans, Mycobacterium tuberculosis, M. avium) as compared with a low-exposure antigen (tetanus toxoid). Overall, delayed type hypersensitivity detection rate increased under HAART. Multivariate analysis showed improvement of antigen-specific T-cell proliferation to be significantly associated with an increase in memory CD4 T cells, whereas improvement of the delayed type hypersensitivity response was associated with a decrease in plasma HIV-1 RNA. CONCLUSIONS: HAART for 6 months restored antigen-specific CD4 T-cell response to several antigens. In vitro immune reconstitution was closely correlated with an increase in memory CD4 cells. Restoration of delayed type hypersensitivity was associated with suppression of viraemia. It appears that in addition to expansion of memory CD4 cells, suppression of viraemia following HAART may allow an improved inflammatory reaction, thus providing even stronger immune reconstitution. PMID- 10513644 TI - Baseline HIV drug resistance profile predicts response to ritonavir-saquinavir protease inhibitor therapy in a community setting. AB - OBJECTIVE: To determine whether baseline drug resistance assays could help to predict treatment failure with the protease inhibitor combination ritonavir saquinavir. METHODS: Baseline HIV-1 drug resistance was determined for 76 consecutive patients who started treatment with the dual protease inhibitor combination ritonavir-saquinavir between September 1996 and June 1997 either alone or in combination with other antiviral agents. Resistance to 10 different antiviral agents was assessed by both phenotype (Virco Antivirogram) and genotype (Vircogen). RESULTS: Resistance inferred from viral genotype was similar to measured phenotypic resistance for both ritonavir and saquinavir (P<0.01). Baseline drug resistance phenotype was predictive of poor virological response to this dual protease inhibitor combination, despite the confounding effects of other antivirals. Patients were at least four times less likely to achieve a 0.5 log10 decrease in plasma HIV RNA viral load if their viral isolates were resistant to ritonavir or saquinavir. Patients classified as resistant to either drug using either method had median decreases in plasma viral load of 0.05 log10 HIV RNA copies/ml or less, compared to >0.8 log10 for those with sensitive virus. Patients resistant to both drugs never achieved plasma viral loads <100000 copies/ml. As little as fourfold increases in baseline resistance appeared to be sufficient to compromise even dual protease inhibitor therapy. CONCLUSION: Baseline resistance to ritonavir or saquinavir or both was associated with a poor antiviral response. Our data suggest that the measurement of drug resistance may assist in optimizing antiretroviral therapy in the clinic. PMID- 10513645 TI - Predictors of optimal virological response to potent antiretroviral therapy. AB - BACKGROUND: Current potent antiretroviral therapy (using a protease inhibitor and two nucleoside reverse transcriptase inhibitors) produces a durable suppression of HIV replication in less than 75% of treated patients. Identification of predictors of successful therapy might allow the development of improved strategies to increase response rates. METHODS: We analyzed retrospectively the results from a multicenter, randomized, double-blind Phase III study of combination anti-HIV therapy with nelfinavir, zidovudine, and lamivudine to evaluate the relationship between virological response over 48 weeks of treatment to variables which could potentially serve as early predictors of long-term response. The goal was to produce long-term suppression of viral load to sensitive (<400 copies HIV RNA/ml) and ultrasensitive (<50 copies HIV RNA/ml) limits of quantification with the Amplicor PCR assay. FINDINGS: Baseline viral load, the change in viral load over the first 4 weeks of treatment, the 2 h post dose nelfinavir levels and the time to respond to HIV RNA levels of <400 copies/ml and <50 copies/ml have the best predictive value in determining response and response duration. Patients who achieved very low viral nadirs (<50 copies HIV RNA/ml) had significantly longer responses than those who achieved nadirs of 50-400 copies HIV RNA/ml. INTERPRETATION: Parameters that can be measured easily at baseline or early after therapy is started can identify patients at high risk of failure with standard treatment. Such patients may be candidates for more aggressive therapy or for alternative strategies designed to improve outcome. In addition, these results support the use of ultra-sensitive HIV RNA assays to predict long-term outcome of anti-HIV therapy. PMID- 10513646 TI - Survival improvement of AIDS-related progressive multifocal leukoencephalopathy in the era of protease inhibitors. Clinical Epidemiology Group. French Hospital Database on HIV. AB - OBJECTIVE: To estimate the change in survival of patients with AIDS-related progressive multifocal leukoencephalopathy (PML), in relation to the introduction of protease inhibitors (PI). DESIGN: The French Hospital Database on HIV (FHDH) is a prospective cohort of 70 224 HIV-infected subjects. This study included the patients diagnosed with PML between 1 July 1995 and 30 June 1997. PML diagnosis was both presumptive and confirmed. We compared the survival probability according to the diagnosis period (period 1 or 2, before or after introduction of PI in France on 1 April 1996). Cox's model was used to calculate the relative hazards of death according to the antiretroviral regimen. RESULTS: The study included 246 patients, 109 diagnosed during period 1 and 137 during period 2. In all, 131 patients received an antiretroviral combination that included PI. By 31 December 1997, a total of 131 deaths had been reported. The probability of survival at 6 months for patients from period 2 was nearly twice as high as for patients from period 1 (60.5 versus 34.5%). In comparison with patients receiving no treatment, the risk of death in patients on combination therapy not including PI was reduced by 38% [relative hazard (RH) 0.62, 95% confidence interval (CI) (0.41; 0.95), P = 0.026] and in patients on combination therapy with PI, by 63% [RH 0.37, 95% CI (0.22; 0.64), P = 0.0004]. CONCLUSION: This study of a large cohort of patients diagnosed with PML (n = 246), provides evidence that a combination antiretroviral regimen, especially one including PI, confers marked survival benefits. PMID- 10513647 TI - Positive and sustained effects of highly active antiretroviral therapy on HIV-1 associated neurocognitive impairment. AB - OBJECTIVES: To determine whether highly active antiretroviral therapy (HAART) is effective in HIV-associated neurocognitive impairment. DESIGN: An open label, prospective, observational study. METHODS: Since April 1996, 116 patients with advanced HIV infection, reverse transcriptase inhibitor (nRTI) experienced but protease inhibitor (PI) naive, were screened for the presence of neurocognitive impairment. Ninety patients with confounding neurological illness, opportunistic infections or drug abuse were excluded. The remaining 26 patients underwent comprehensive neuropsychological testing, and laboratory measures before, after 6 and after 15 months of treatment with one PI plus two nRTI. RESULTS: The prevalence of neurocognitive impairment decreased from 80.8% (baseline) to 50.0% (P<0.05) (sixth month) and to 21.7% (P<0.001) (15th month). Among the functions explored, the impairment of concentration and speed of mental processing decreased from 65.4 to 21.7% (P<0.01) and of memory impairment from 50 to 8.7% (P<0.01). Comparing baseline with the sixth and 15th month raw scores, a statistically significant improvement was seen in measures exploring concentration and speed of mental processing (P<0.05), mental flexibility (P<0.05), memory (P<0.05), fine motor functions (P<0.05) and visuospatial and constructional abilities (P<0.01). After 6 months of HAART patients with a normal neuropsychological examination had lower mean plasma viraemia (2.95 versus 3.97 log copies/ml, P<0.05) and greater mean log plasma HIV RNA changes from baseline (-1.84 versus -0.83 log copies/ml, P<0.05) than neuropsychologically impaired subjects. CONCLUSION: HAART produces a positive and sustained effect on neurocognitive impairment in HIV-infected patients. A reduction of plasma viral load was associated with the regression of neuropsychological test abnormalities. PMID- 10513648 TI - Relapse rates after short-course (6-month) treatment of tuberculosis in HIV infected and uninfected persons. AB - OBJECTIVE: To determine the rate of tuberculosis relapse among HIV-seropositive and -seronegative persons treated for active tuberculosis with short-course (6 month) therapy. DESIGN: Consecutive cohort study. SETTING: City of Baltimore tuberculosis clinic. PATIENTS: Tuberculosis patients treated between 1 January 1993 and 31 December 1996. INTERVENTION: Patients received 2 months of isoniazid, rifampin, pyrazinamide and ethambutol followed by 4 months of isoniazid and rifampin. MAIN OUTCOME MEASURE: Passive follow-up for tuberculosis relapse was performed through September 30, 1998. RESULTS: There were 423 cases of tuberculosis during the study period; 280 patients completed a 6-month course of therapy. Therapy was directly-observed for 94% of patients. Of those who completed therapy, 47 (17%) were HIV-seropositive, 127 (45%) were HIV seronegative, and 106 (38%) had unknown HIV status. HIV-infected patients required more time to complete therapy (median 225 versus 205 days; P = 0.04) but converted sputum culture to negative within the same time period (median 77 versus 72 days; P = 0.43) as HIV-seronegative or unknown patients. Relapse occurred in three out of 47 (6.4%) HIV-infected patients compared to seven out of 127 (5.5%) HIV-seronegative patients (P = 1.0). Relapse rates also did not differ when HIV-seropositive patients were compared with HIV-seronegative and patients with unknown HIV status (6.4% versus 3.0%; P = 0.38). Of the 10 patients with tuberculosis relapse, restriction fragment length polymorphism data were available for five; all five relapse isolates matched the initial isolate. CONCLUSIONS: These results support current recommendations to treat tuberculosis in HIV-infected patients with short-course (6-month) therapy. PMID- 10513649 TI - Association of indicators of bacterial vaginosis with a female genital tract factor that induces expression of HIV-1. AB - OBJECTIVE: The aim of this study was to determine the relationship of bacterial vaginosis and bacterial vaginosis-associated microorganisms with an HIV-inducing factor (HIF) found in cervicovaginal lavage. DESIGN: A total of 26 cervicovaginal lavage specimens collected from 17 women were used in this study to determine if HIF was significantly associated with features consistent with bacterial vaginosis. METHODS: Patients were evaluated for various clinical features including age, HIV status and stage, CD4 cell counts, clinical diagnosis of gynecological infections, vaginal pH, Gram stains of vaginal fluid, phase of menstruation, and presence of cervical dysplasia. Cervicovaginal lavage specimens were analyzed for the presence of HIF by U1 bioassay. The presence of Gardnerella vaginalis, and general Mycoplasmataceae, and specifically Mycoplasma hominis, Ureaplasma urealyticum, M. fermentans, M. genitalium in cervicovaginal lavage were determined by semiquantitative PCR. RESULTS: Eleven cervicovaginal lavage samples from seven women were HIF-positive and 15 cervicovaginal lavage samples from 11 women were HIF-negative (patient No. 8 had two HIF-negative cervicovaginal lavage and one HIF-positive cervicovaginal lavage). The following parameters were significantly associated with HIF: abnormal vaginal fluid pH (>4.5) (P = 0.006), Gram stains indicative of bacterial vaginosis (P = 0.007), normal menstrual cycle (P = 0.0007) and PCR detection and relative quantity of M. hominis (P = 0.0003, P = 0.002). CONCLUSIONS: This study indicates that HIF is closely associated with features of bacterial vaginosis. PMID- 10513650 TI - Impaired liver function and retroviral activity are risk factors contributing to HIV-associated thrombocytopenia. Swiss HIV Cohort Study. AB - OBJECTIVE: To investigate the relationship between thrombopoetin (TPO) serum levels and HIV-associated thrombocytopenia. DESIGN AND METHODS: The relationship between TPO levels and severity of HIV-associated thrombocytopenia was investigated. Thirty-eight patients (19 patients with 30-96x10(9) platelets/l and 19 patients with <10x10(9) platelets/l) were matched with 38 HIV-positive non thrombocytopenic patients (>150x10(9) platelets/l). RESULTS: HIV-positive patients with normal platelet counts had a median TPO serum level of 137 pg/ml. Patients with 30-96x10(9) platelets/l had decreased TPO levels with a median of 90 pg/ml (P = 0.016), and were more likely to have elevated serum aspartate transferase levels (P<0.001) and hepatomegaly by palpation or ultrasound imaging (P = 0.005). The median TPO serum level of HIV-infected patients with severe thrombocytopenia was 110 pg/ml (non-significant). All patients with severe thrombocytopenia were positive for antibodies against hepatitis B virus core antigen, compared with 80% of HIV-infected persons without thrombocytopenia. Patients with severe thrombocytopenia were more likely to have high HIV replication compared to patients with normal platelet counts (P = 0.02), and reduction of plasma HIV-1 RNA levels was associated with increasing platelet counts. Severe thrombocytopenia was not associated with liver disease. CONCLUSIONS: Liver disease predisposes for low TPO serum levels and mild thrombocytopenia. High retroviral activity predisposes for severe, immune thrombocytopenic purpura-like thrombocytopenia. At least two distinct categories of severe HIV-associated thrombocytopenia exist, one responsive to antiretroviral treatment and one non-responsive to antiretroviral treatment. PMID- 10513651 TI - HIV viral load and response to antileishmanial chemotherapy in co-infected patients. AB - OBJECTIVE: To investigate whether clearance of Leishmania parasites from tissue aspirate smears in patients with HIV and visceral leishmaniasis (VL) co-infection treated with pentavalent antimonials is influenced by initial HIV viral load and to assess the effect of active VL on HIV viral load and replication in vivo. METHODS: Leishmania parasites were identified in Giemsa-stained smears prepared from tissue aspirates. Parasite index was determined by quantifying Leishmania donovani bodies in smears. HIV-1 RNA was quantitated by using the nucleic acid sequence-based amplification technique with a limit of detection of 500 copies/ml. All patients were treated with pentavalent antimonials at 20 mg pentavalent antimony (Sb(V))/kg daily for a total of 28 days. None of the patients received specific anti-retroviral therapy. RESULTS: Seventeen patients (73.9%) showed good initial response to anti-leishmanial treatment and the remaining six (26.1%) had very poor response. Among the good responders, 11 (64.7%) had no demonstrable Leishmania donovani bodies in post-therapy tissue aspirate smear preparations, and in the remaining six (35.3%) their parasite loads were reduced to very low levels. Patients with poor response had persistently high parasite index despite completion of anti-leishmanial chemotherapy. Poor responders had pre-treatment median HIV viral load that was >160-fold higher than responders to anti-leishmanial chemotherapy; [410000 copies/ml (quartile range, 33000-530000) and 2500 copies/ml (quartile range 500 297500), respectively]. Furthermore, compared with pre-treatment viral concentrations, patients with good response showed marked reduction in post treatment viral load. In contrast, post-treatment HIV viral concentrations were markedly increased among patients with poor response to anti-leishmanial therapy. CONCLUSIONS: The results suggest that pre-treatment HIV viral load influences response to anti-leishmanial chemotherapy and active VL is associated with increased viral replication in vivo, supporting the notion that dual infection plays an important role in the pathogenesis and disease progression of either infection. PMID- 10513652 TI - Maternal viral load and vertical transmission of HIV-1 in mid-trimester gestation. AB - BACKGROUND: It is now accepted that the majority of HIV-1 vertical transmissions occur in late gestation and at the time of delivery. However, there is wide variation in the prevalence rate of mid-trimester vertical transmission. We assessed the maternal HIV-1 RNA viral load and in utero transmission during mid trimester gestation. METHODS: Patients were enrolled when they decided to have their pregnancies terminated between 17 and 24 weeks of gestation. Prostaglandin induced abortion with PGE1 analogue vaginal administration was carried out in all patients. Maternal plasma HIV-1 RNA viral load and plasma HIV-1 RNA (qualitative) from abortus heart blood were assessed. RESULTS: Amongst 41 HIV-1 seropositive pregnant women not receiving antiretroviral therapy plasma HIV-1 RNA was detected in the abortus heart blood from two women (4.9%; 95% confidence interval (CI), 0.6-16.5). Transmission occurred in one out of nine (11.1%; 95% CI, 0.3-48.2) with maternal viral load > or =100000 copies/ml versus one out of 32 (3.1%; 95% CI, 0.1-16.2) of those with <100000 copies/ml (P = 0.39). CONCLUSIONS: The frequency of HIV-1 vertical transmission during mid-trimester was approximately 5% as detected by plasma HIV-1 RNA (qualitative) method in the fetuses aborted from the prostaglandin termination of pregnancy. During mid-trimester gestation there was no correlation between high maternal viral load and vertical transmission. PMID- 10513653 TI - Natural history of HIV infection in the era of combination antiretroviral therapy. AB - BACKGROUND: The use of protease inhibitor-containing (PI) combination antiretroviral therapy has led to a reduction in the incidence of opportunistic illness and mortality (events) in HIV infection. We wished to quantify the changing incidence of these events in our clinical practice and delineate the relationship between CD4, HIV-1 RNA, and development of events in patients receiving PI combination therapy. METHODS: We assessed HIV-infected patients with CD4 counts < or =500 cells x10(6)/l. We calculated the incidence of events from 1994 through 1998 and analyzed the association of temporal changes in event incidence and use of antiretroviral therapy. In patients on PI combination therapy, we determined the probability of achieving and maintaining an undetectable HIV-1 RNA response and determined the association of CD4, HIV-1 RNA, and developing an event. RESULTS: The incidence of opportunistic illness declined from 23.7 events/100 person-years in 1994 to 14.0 events/100 person-years in 1998 (P<0.001). Mortality declined from 20.2 deaths/100 person-years in 1994 to 8.4 deaths/ 100 person-years in 1998 (P<0.001). Use of PI combination therapy was associated with a relative rate of opportunistic illness or death of 0.66 [95% confidence interval (CI), 0.51-0.85; P<0.001]. The relative incidence of each of 16 opportunistic illnesses was approximately the same in 1998 as in 1994 except for lymphoma, cervical cancer and wasting syndrome which do not appeared to have declined in incidence. Approximately 60% of patients who received PI therapy achieved an undetectable HIV-1 RNA, and 65% of these patients maintained durable suppression of HIV-1 RNA. Achieving an undetectable HIV-1 RNA was associated with a decreased risk of an event, and was the variable most strongly associated with an increase in CD4 level. By multivariate analysis, the concurrent CD4 level was most strongly associated with developing an event. CONCLUSIONS: We observed a significant decline in the incidence of opportunistic illness and death from 1994 through 1998 associated with combination antiretroviral therapy. Patients who develop events while being treated with PI combination therapy were not likely to have achieved an undetectable HIV-1 RNA and are likely to have a low concurrent CD4 level. PMID- 10513654 TI - The practical significance of potential biases in estimates of the AIDS incubation period distribution in the UK register of HIV seroconverters. AB - OBJECTIVES: To assess the practical significance of the following sources of bias for estimates of the AIDS incubation period in a large seroconverter cohort: estimation of the time of seroconversion; presentation with an HIV-related illness; preferential inclusion of survivors; loss to follow-up and analysis cut off date; the inclusion of Kaposi's sarcoma as an AIDS event; death without an AIDS diagnosis; and representativeness of the HIV population. METHODS: Standard non-parametric survival methods were used to estimate the AIDS incubation period distribution. The practical importance of each type of bias was assessed using various sensitivity analyses. RESULTS: The potential sources of bias of most practical importance in this study were the right-censoring strategy and that due to lack of documentation of a negative HIV antibody test. Five different right censoring strategies gave estimates of the median time to AIDS ranging from 8.1 to 10.8 years for the 1202 individuals enrolled in the UK Register of HIV Seroconverters. HIV-infected persons with a history of a previous antibody negative test which could not be verified appeared to progress to AIDS more rapidly than persons with such verification (Relative risk = 1.8, 95% confidence intervals = 1.3-2.3). CONCLUSIONS: As a number of possible causes of bias can impact on results, care must be taken to document them and control for them wherever possible. In our study, this was particularly relevant in relation to the documentation of a previous HIV antibody negative test and the choice of analysis cut-off dates. As methods may differ between cohorts, comparison of the published results from one cohort with those of another may be misleading. PMID- 10513655 TI - HIV prevalence, sexual risk behaviour and sexual mixing patterns among migrants in Amsterdam, The Netherlands. AB - OBJECTIVE: To study (1) HIV prevalence; (2) sexual risk behaviour; (3) sexual mixing patterns; (4) determinants of disassortative (between-group) mixing among migrant groups in Amsterdam, the Netherlands and to gain insight into the potential for heterosexual spread of HIV/sexually transmitted diseases. DESIGN: Cross-sectional study among 1660 Surinamese, Antilleans and sub-Saharan Africans, mainly recruited on the streets. METHODS: Saliva was tested for HIV and questions were asked about sociodemographic characteristics, sexual behaviour and the ethnicity of sexual partners. Multivariate logistic regression analysis was used to find predictors for disassortative mixing. RESULTS: HIV prevalence was 1.1% (95% confidence interval: 0.6-1.7). Compared with the Dutch population in general, our study group reported having multiple partners, concurrent partnerships and a history of sexually transmitted diseases much more frequently. Sex in the country of origin during a visit occurred frequently and there was a considerable degree of sexual mixing between different ethnic groups in the Netherlands. For men, disassortative mixing was associated with hard drug use, recent immigration, a high number of partners, being from Nigerian or Hindu Surinamese origin, a recent sexually transmitted disease and, for steady relationships, consistent condom use. For women, determinants included: hard drug use, low income, being a-religious and, for Antillean and Ghanaian women, consistent condom use. CONCLUSION: Our data suggest a potential for heterosexual spread of sexually transmitted diseases within and between ethnic groups in the Netherlands. The potential for HIV spread is however limited by the low HIV prevalence at present among these groups. This situation may change when HIV prevalence increases in the countries of origin, as bridges exist between those countries and the Netherlands. Culturally appropriate AIDS prevention programmes remain important for these groups. PMID- 10513656 TI - Clinical disease associated with HIV-1 subtype B' and E infection among 2104 patients in Thailand. AB - BACKGROUND: Two HIV-1 envelope subtypes have accounted for virtually all infections in Thailand: subtype B' (Thai B), found mainly in injection drug users (IDU), and subtype E, found in over 90% of sexually infected persons and an increasing proportion of IDU in recent years. It remains unclear whether there are differences in pathogenesis associated with these HIV-1 subtypes. METHODS: From November 1993 to June 1996, demographic, risk, clinical, and laboratory data were collected by enhanced surveillance from HIV-infected inpatients (> or =14 years) at an infectious disease hospital near Bangkok. HIV-1 subtype was determined by V3-loop peptide enzyme immunoassay (EIA). Because of confounding, multivariate analyses were stratified by risk category and controlled for sex and age. RESULTS: The infecting HIV-1 subtype was determined for 2104 (94.9%) of 2217 HIV-infected patients with complete data: 284 (13.5%) were subtype B', 1820 (86.5%) were E. Specimens from 113 (5.1%) patients were non-reactive or dually reactive on peptide EIA and were excluded. Among IDU, 199 (67.2%) had subtype B', and 97 (32.7%) had E. IDU accounted for 70.1% (199/284) of patients with subtype B' and 5.3% (97/1820) of those with E. Patients infected with HIV-1 subtypes B' or E had similar spectrums of opportunistic infections (OI), levels of immunosuppression, and in-hospital mortality rates. Of patients who did not inject drugs, more patients infected with subtype E had extrapulmonary cryptococcosis than those with subtype B' (adjusted odds ratio, 2.6; 95% confidence interval, 1.28-5.37). CONCLUSION: HIV-1 subtypes B' and E seem to be associated with similar degrees of immunosuppression and, with one exception, with similar OI patterns. These data do not suggest an association between HIV-1 subtype and differences in pathogenicity. PMID- 10513657 TI - Pneumonia in HIV-infected patients: a case-control survey of factors involved in risk and prevention. AB - OBJECTIVE: To assess the factors that increase or decrease the risk of pneumonia with particular attention to immunization with pneumococcal and influenza vaccines in a group of HIV-infected persons. DESIGN: A retrospective, case control study based on information entered into a standard database and the medical record. SETTING: Patients attending a referral clinic specializing in AIDS/HIV care at a public hospital. PATIENTS: Among over 2000 subjects entered into a database in 8 years, 127 incidents of pneumonia were identified from the record. These cases were matched with 127 CD4 cell count matched, concurrent controls. INTERVENTIONS: None. MAIN OUTCOME MEASURE: The principal hypothesis was that chart review would find a decreased frequency of pneumococcal immunization in the pneumonia cases compared with matched controls. RESULTS: Pneumococcal immunization was associated with a reduction of the risk of pneumonia by nearly 70%. The effect was seen even when immunization was given with a CD4 cell count of less than 100/mm3. Injection drug users and African-Americans had a twofold increased risk of pneumonia. CONCLUSION: The study provides data to support the current recommendation for pneumococcal immunization of all HIV-infected persons. Although this conclusion could lead to renewed enthusiasm for increasing pneumococcal immunization rates in HIV-infected persons, it must be recognized that the study is observational and ascertainment bias cannot be excluded. PMID- 10513658 TI - Comparison of three commercial assays for the quantification of plasma HIV-1 RNA from individuals with low viral loads. PMID- 10513659 TI - Intralesional or topical cidofovir (HPMPC, VISTIDE) for the treatment of recurrent genital warts in HIV-1-infected patients. PMID- 10513660 TI - Increase in oral sex and pharyngeal gonorrhoea: an unintended effect of a successful condom promotion programme for vaginal sex. PMID- 10513661 TI - Knowledge, attitudes and practices of university students regarding HIV infection, in Phnom Penh, Cambodia, 1999. PMID- 10513663 TI - Overdose of atovaquone in a patient with AIDS. PMID- 10513662 TI - Renin-angiotensin system inhibition in a patient having an overdose of HIV protease inhibitor. PMID- 10513664 TI - Impact of Kaposi's sarcoma in occurrence of opportunistic central nervous system disease. Clinical Epidemiology Group. PMID- 10513665 TI - Diabetic ketoacidosis in an HIV patient: a new mechanism of HIV protease inhibitor-induced glucose intolerance. PMID- 10513667 TI - HLA-B*5703 independently associated with slower HIV-1 disease progression in Rwandan women. PMID- 10513666 TI - Pharmacokinetics of nelfinavir in an HIV patient with renal insufficiency. PMID- 10513668 TI - Isolation of HIV-1-specific cytotoxic T lymphocytes using human leukocyte antigen coated beads. PMID- 10513669 TI - Sudden cardiac death in a patient on 2 years of highly active antiretroviral treatment: a case report. PMID- 10513670 TI - Differential effect of ritonavir and indinavir on immune response to hepatitis C virus in HIV-1 infected patients. PMID- 10513671 TI - Reply. Lest we forget: neuropsychiatry and the new generation anti-HIV drugs. PMID- 10513672 TI - Avascular necrosis in HIV infection. PMID- 10513673 TI - Spanish consensus conference on drug resistance testing in clinical practice. HIV drug resistance Spanish panel. PMID- 10513674 TI - Local lymph node assay: use in hazard and risk assessment. AB - The murine local lymph node assay (LLNA) was developed as an alternative method for the identification of chemicals that have the ability to cause skin sensitization and allergic contact dermatitis. The assay now has been evaluated extensively in the context of both national and international inter-laboratory collaborative trials and has been the subject of detailed comparisons with guinea pig test methods and human skin sensitization data. On the basis of these evaluations the LLNA has been endorsed recently by the US Interagency Coordinating Committee on the Validation of Alternative Methods (ICCVAM) as a stand-alone method for skin sensitization testing. The assay offers a number of important benefits compared with conventional guinea pig test methods, among these being provision of an objective and quantitative endpoint. Moreover, the LLNA provides advantages in the context of animal welfare; compared with guinea pig tests, fewer animals are required and these animals are subject to less trauma. It is important now that the validation status of the LLNA is recognized and the method applied widely so that these advantages may be realized. Hazard identification represents only the first step in the risk assessment process. A full toxicological evaluation of skin sensitization activity requires an understanding of relative potency. Guinea pig methods do not lend themselves readily to assessment of potency, and interest recently has focused on the utility of the LLNA for this purpose. Contained within this review article are brief descriptions of the history of the LLNA and the immunobiological basis for the method, together with detailed accounts of the conduct and interpretation of the assay. Procedural modifications to, and alternative endpoints for, the LLNA are considered also. Finally, the current regulatory status of the LLNA is summarized and the application of the method for the purposes of defining relative potency and developing risk assessments is reviewed. PMID- 10513675 TI - Characterization of the Asialofetuin microtitre plate-binding assay for evaluating inhibitors of ricin lectin activity. AB - Optimum conditions for the binding of ricin to the glycoprotein asialofetuin immobilized on microtitre plates were investigated for the purpose of evaluating inhibitors of ricin B-chain lectin activity. Such inhibitors are of potential value in the use of immunotoxins based on ricin. This assay was first reported in 1986, but has not been characterized fully. Maximum binding of asialofetuin to the plate was observed at a concentration of ca. 4 microg ml(-1). Binding increased with time of incubation (1-24 h), pH (7.4-9.9) and temperature (2-37 degrees C). The pH effects were more marked at lower temperatures. Saturable binding of ricin to immobilized asialofetuin was observed, and at least 80% of maximum binding was observed by 10 min of incubation time. The binding was found to be very tight, such that an appreciable proportion of ricin added to the wells was bound at low concentrations, and binding was only partially reversible by addition of free galactose. Consequently, only estimates of the ricin asialofetuin and ricin-galactose dissociation constants could be determined: 1.9 nM and 83 microM, respectively. Binding of ricin A- and B-chains was found to be 47% (at a 200-fold higher concentration) and 26% (at a twofold higher concentration) of that of the whole ricin molecule, respectively. The assay permits qualitative comparison of inhibitors of ricin B-chain lectin activity. PMID- 10513676 TI - Immunohistochemical characterization of the basement membrane epitopes in bis(2 chloroethyl) sulfide-induced toxicity in mouse ear skin. AB - Sulfur mustard (bis(2-chloroethyl) sulfide (HD)), a potent cutaneous vesicant and bifunctional alkylating agent, produces significant time-dependent histopathological changes in the skin of the mouse. The right ears of male CD1 mice were exposed topically to 5.0 microl of 195 mM (0.16 mg) HD in dichloromethane and harvested at 6, 12, 18 and 24 h. The left ear control was dosed with 5.0 microl of dichloromethane. In all controls and HD-treated mouse ear, moderate immunofluorescence staining was seen at the epidermal-dermal junction with bullous pemphigoid (BP), epidermolysis bullosa acquisita (EBA) and laminin (Lam), and light staining was observed with bullous pemphigoid 180 (BP180), fibronectin (Fn) and type IV collagen (Coll IV). Mouse anti-human monoclonal antibodies for GB3, L3d and 19-DEJ-1 (Uncein) did not cross-react. In microvesicles, BP, BP180 and Fn showed areas of light focal epidermal staining and homogeneous dermal staining, and EBA, Lam and Coll IV showed moderate dermal staining. Both BP and Fn exhibited weak, inconsistent staining with time. Immunoelectron microscopy (IEM) revealed similar results, with an increase in cell damage from 6 to 24 h, which corresponded to a decrease in staining intensity. Cell proliferation, expressed as the growth fraction of proliferating cell nuclear antigen (PCNA), showed an increase in cell damage. The growth fraction was lower in the inner ear and showed time-dependent differences. The immunofluorescence and IEM results indicate that HD causes an undulating inconsistent separation in the uppermost lamina lucida with focal cleavage into the lower portion of the basal keratinocytes just above the plasma membrane. Although this pattern of separation differs from other in vivo models in which the split occurs exclusively within the lamina lucida, this should not preclude its role as a screening model to study the effects and development of specific prophylactic and therapeutic strategies. PMID- 10513677 TI - L-lactate reduces in vitro the inhibition of butyrylcholinesterase (BChE) by paraoxon (E 600). AB - Intoxication with the organophosphorus compound paraoxon (POX), an inhibitor of serine hydrolases, is frequent. Oximes are the only enzyme reactivators clinically available. Serendipitous observation led us to the hypothesis that lactate might attenuate some of the POX effects. In vitro effects of lactate on the inhibition of butyrylcholinesterase (BChE) by POX were assessed in plasma of 12 healthy human volunteers. The determinations were repeated using different lactate and different POX concentrations. The BChE activity determinations were performed in the following settings: (i) baseline untreated plasma (BL); (ii) after addition of POX to plasma (pl+POX); (iii) after POX and plasma were incubated and then lactate was added (pl+POX/lact); (iv) after addition of lactate to plasma (pl+lact); (v) after lactate and plasma were incubated and then POX was added (pl+lact/POX); (vi) after lactate and POX were incubated and then added to plasma (lact+POX/pl). In the micro- and millimolar ranges, lactate is able to abolish in vitro the inhibition of BChE by POX in human plasma when added to plasma prior to POX or when incubated with POX prior to addition to plasma. Lactate added to plasma after POX has no protective effect. In a second set of experiments, the effect of lactate on BChE activity was determined. At high millimolar concentrations, lactate itself inhibits BChE to an extent comparable to POX. Lactate is a mixed inhibitor of BChE, being able to interfere with the enzyme-substrate complex (inhibition constant for the enzyme-inhibitor-substrate complex K'I(EIS) = 81 mM) and the enzyme (inhibition constant for the enzyme inhibitor complex K(I) (EI) = 26 mM). PMID- 10513678 TI - Effects of inorganic divalent cations on the renal basolateral transport system for organic anions. AB - Recent work demonstrated that the heavy metal ion Cd2+ increases the transport of p-aminohippuric acid (PAH) across the basolateral membrane of microdissected non perfused rabbit kidney S2 proximal tubule segments. Usually, such ions induce damage of various renal transport systems, therefore the effects of divalent metal ions Zn2+, Co2+ and Ni2+ on this transporter were investigated. Addition of Ni2+ or Zn2+ to the bathing solution leads to a significant reduction of basolateral PAH transport, with IC50 values of 2 x 10(-5) and 10(-6) M, respectively, whereas Co2+ failed to inhibit PAH accumulation. Simultaneous incubation with thrombin (10(-9)M), which is known to increase [Ca2+]i, abolished the effects of the divalent ions. Our results indicate that Ni2+ and Zn2+ reduce cellular PAH uptake. Because Ni2+ and Zn2+ are calcium channel blockers, these effects are probably due to a reduction of [Ca2+]i by an interaction of these metals with binding sites in the calcium channel, whereas Co2+ does not affect these binding sites. This finding is supported by the fact that thrombin abolished the cation effects. PMID- 10513679 TI - Influence of the viscosity of iodixanol on medullary and cortical blood flow in the rat kidney: a potential cause of Nephrotoxicity. AB - The aim of the present study was to investigate the effects of four iodinated contrast media on cortical, inner medullary and outer medullary blood flow in the rat kidney by using laser-Doppler flowmetry. The high-osmolar contrast medium diatrizoate did not significantly modify medullary perfusion but moderately decreased the cortical blood flow when injected at a dose of 1600 mg iodine kg( 1). Similar effects were obtained with the low-osmolar contrast media ioxaglate and iobitridol. In contrast, the new iso-osmolar contrast medium iodixanol induced a dose-dependent reduction of perfusion in all regions tested. This effect was accompanied by concomitant hypotension. The reduction of inner medullary and cortical blood flow induced by iodixanol was partially alleviated by heating the solution prior to injection and subsequently reducing its viscosity. In the outer medulla, however, this procedure did not improve blood flow. These results suggest that lowering the viscosity may palliate the harmful effects of iodixanol on the inner medulla and cortex, but may not protect the outer medulla from hypoxic injury. PMID- 10513681 TI - Behavioural and histological effects of atrazine on freshwater molluscs (Physa acuta Drap. and Ancylus fluviatilis Mull. Gastropoda). AB - This study examines the direct and indirect effects of atrazine on two grazer species--Physa acuta and Ancylus fluviatilis--as assessed by changes in mortality rates, biomass, searching behaviour and histological properties. No direct effects were observed in the acute toxicity test (48 h) with 0.02, 0.2, 2, 10 and 20 mg l(-1) of atrazine. A chronic toxicity test (18 days) performed in six experimental channels with 15 microg l(-1) of atrazine showed significant changes in grazer behaviour, increased searching velocity and different movement patterns in animals exposed to herbicide. No significant effects were observed in rates of mortality and biomass. Kidney cells of Physa acuta displayed an important cell lysis when animals were exposed to 0.1 mg l(-1) of atrazine for 10 days, and this effect was not reversed after a decontamination process. These results provide evidence of behavioural and structural changes in freshwater molluscs due to a subacute atrazine concentration. PMID- 10513682 TI - Developmental effects of plasticizer butyl benzyl phthalate after a single administration in rats. AB - The objective of this study was to determine the susceptible day for the developmental toxicity of butyl benzyl phthalate (BBP) by a single administration on one of the days during organogenesis. Pregnant rats were given a single dose of BBP by gastric intubation at a dose of 1000 mg kg(-1) on one of days 13-15 of pregnancy and at 1500 mg kg(-1) on one of days 6-16 of pregnancy. Post implantation embryolethality was found in pregnant rats given on one of days 6 16, except for day 7. Teratogenicity was noted after a single dosing of BBP on one of days 6, 7, 9, 10, 12, 14 and 15. Deformity of the cervical vertebrae frequently was observed after administration of BBP on day 7. Cleft palate and fusion of the sternebrae were found exclusively, after administration of BBP on day 15. It can be concluded that the manifestation of deviant development induced by BBP varies with the developmental stage at the time of administration and that BBP induces two discrete responses from embryos to teratogenicity during early and late organogenesis. PMID- 10513680 TI - Effects of lead(II) nitrate and a dithiocarbamate fungicide on the rat lung. AB - The pulmonary toxicity of two potential environmental pollutants was studied in rats 1, 7 and 30 days after a single intratracheal instillation of lead nitrate and Dithane M-45 (mancoceb), either individually or in various combinations. The cell count, protein, phospholipids and lactate dehydrogenase level were determined in the bronchoalveolar lavage fluid, as were the protein, phospholipids and acid phosphatase contents in the lung tissue. Lead nitrate and Dithane M-45 induced acute inflammation reactions with different features. The effects of mixtures of lead nitrate and Dithane M-45 were found to be different from those of the individual components. PMID- 10513684 TI - Carbon monoxide. PMID- 10513683 TI - Effects of p-nonylphenol (NP) and diethylstilboestrol (DES) on the Alderley Park (Alpk) rat: comparison of mammary gland and uterus sensitivity following oral gavage or implanted mini-pumps. AB - An earlier report by Colerangle and Roy indicated that administration of p nonylphenol (NP) to Noble rats, via subcutaneously implanted mini-pumps at estimated doses of 53.2 and 0.073 mg kg(-1) day(-1) for 11 days, led to proliferation of the mammary gland. Those results indicated a ca. 600-fold enhancement in assay sensitivity to NP over that of the standard 3-day rat uterotrophic assay. The potential importance of these observations led us to repeat the experiments in the Noble rat, as described earlier. Although our earlier results confirmed the reported effects of diethylstilboestrol (DES) on the mammary gland of Noble rats, we found no effects with NP. The present report extends our investigations of the effects of NP and DES on the mammary gland and uterus of other rat strains using both oral dosing and exposure via mini-pumps. The 3-day oral uterotrophic assay responses to NP were similar for immature Alderly Park (Alpk; Wistar-derived) and immature Sprague-Dawley rats. Likewise, oral administration of NP to ovariectomized Alpk rats for 11 days gave responses of a similar magnitude to those seen in the 3-day immature assays and in earlier 3-and 11-day oral assays conducted using Noble rats. Administration of NP via mini-pumps to ovariectomized Alpk rats, at the implant doses employed by Colerangle and Roy, gave a negative uterotrophic response. The highest achieved dose levels of NP in the implant experiment (27 mg kg(-1) day(-1)) were lower than in the above assays and the negative response was therefore consistent with the previously defined minimum detection level for NP in the uterotrophic assay of ca. 40 mg kg(-1) day(-1) day(-1). It is concluded that the uterotrophic activity of NP is independent of the strain of rat, the duration of dosing and the route of exposure. Two mammary gland studies were conducted on NP and DES in the Alpk rat. In the first study (a repeat of the techniques used in earlier studies with the Noble rat), NP was administered via mini-pumps (achieved doses of 0.052 and 37.4 mg kg(-1) day(-3) NP) and produced no effect on mammary gland development, whereas DES gave the expected trophic response. In the second mammary gland study, NP was administered orally to Alpk rats at 100 mg kg(-1) day(-1) for 11 days (a dose that produced a positive uterotrophic response in ovariectomized rats). In this experiment, DES, and to a lesser extent NP, increased mammary gland differentiation and cell proliferation. The present studies have demonstrated that the rat mammary gland responds predictably to oestrogenic stimulation but does not show increased sensitivity to oestrogens when compared to the rat uterus. It is also concluded that the minimum detection level for oestrogenic responses of NP in rodents, following oral, dietary and implant routes of exposure, is ca. 40 mg kg(-1) day(-1). PMID- 10513685 TI - Prenatal and perinatal striatal injury: a hypothetical cause of attention-deficit hyperactivity disorder? AB - Experimental data indicate a particular vulnerability of striatal neurons in the developing brain, and together with the idea that the striatum is important for context recognition and behavior, these data have led the author to search for subtle striatal lesions, in the form of biochemical changes, in children who have suffered perinatal adverse events. Evidence is presented to demonstrate that the composition of metabolites in the striatum is altered, primarily in the form of an elevated level of lactate, in human neonates who have suffered various perinatal disorders, such as germinal matrix hemorrhage, intrauterine growth retardation, and asphyxia. An elevated level of lactate suggests tissue hypoxia, which may interfere with the formation of frontostriatal circuits and may play a role in the pathogenesis of the behavioral disturbances observed in a proportion of children with a history of perinatal adverse events. PMID- 10513686 TI - Platelet count and function in children receiving sodium valproate. AB - To evaluate whether valproic acid (VPA) can cause thrombocytopenia and impaired platelet function, the authors prospectively studied 20 children (12 female and eight male; age range 4.9-9.2 years) before and after 6 months of VPA monotherapy. Fifteen healthy sex- and age-matched children served as control subjects. VPA was prescribed at normal dosages (19.7 +/- 9.9 mg/kg), and plasma levels were within the therapeutic range (60.1 +/- 16.5 microg/mL). At the first evaluation, no significant difference between patients and control subjects was observed for platelet count and function. At the second evaluation the platelet counts were significantly lower in the patients (194,200 +/- 37,800/microL; range 157,700-222,400) than in the control subjects (291,100 +/- 41,300/microL; range 261,000-332,500; P < 0.01). Significant differences occurred between patients and control subjects in the release of adenosine triphosphate (ATP) after collagen and adenosine diphosphate (ADP) stimuli and in aggregation after stimulation with collagen, ADP, and arachidonic acid. Significant correlations between platelet count, aggregation, and ATP release and VPA dosage and plasma concentration were also observed. VPA can cause a decreased platelet count and aggregation and ATP release impairment. These side effects can appear after a few months of therapy and with plasma valproate levels within the normal range. They do not seem to be associated with clinical symptoms, and drug discontinuation is not necessary. PMID- 10513687 TI - Peripheral lymphocyte subsets and other immune aspects in Rett syndrome. AB - A possible role of the immune system in the pathogenesis of some neurologic disorders, including infantile autism, was recently postulated. This observation prompted the authors to investigate some immunologic aspects in a group of patients with Rett syndrome, a disorder still not completely clarified but with some points of commonality with infantile autism. Humoral and cell-mediated immunity were investigated in 20 females with Rett syndrome. Peripheral lymphocyte subsets revealed a reduced percentage of CD8+ suppressor-cytotoxic cells in all of the patients with Rett syndrome, resulting in an increased CD4+/CD8+ ratio. In addition, 15 (75%) of the 20 patients had low levels of natural killer cells. Soluble interleukin-2 receptor was elevated in the youngest patients. Antineuronal and antimyelin ganglioside antibodies were absent, as were antinuclear antibodies, antistriated muscle antibodies, and antismooth muscle antibodies. Immunoglobulin fractions and complement were normal for age in all of the patients. PMID- 10513689 TI - Life expectancy and median survival time in the permanent vegetative state. AB - The authors studied life expectancy and risk factors for mortality of persons in the vegetative state (VS). The study participants were 1,021 California patients in the VS during 1981-1996. Because of the large sample size, the authors were able to use multivariate methods to assess the effect of several risk factors on mortality. The authors found a strong secular trend in infant mortality, with rates in the mid-1990s being only one third of those in the early 1980s (P < 0.01). A smaller secular trend was observed for children aged 2-10 years and none for older patients. The mortality risk for older patients fell by approximately 8% for each year since the onset of the VS. The need for gastrostomy feeding was associated with a substantially higher risk, especially for infants and older patients (P < 0.01). Ventilator dependence also appeared to be a risk factor. On the basis of recent mortality rates, life expectancy in the VS is frequently higher than has generally been thought. For example, it is 10.5 additional years (+/- 2 years) for a 15-year-old patient who has been in the VS for 1 year, and 12.2 years for a 15-year-old patient who has been in the VS for 4 years. PMID- 10513688 TI - Diagnostic value of short duration outpatient video electroencephalographic monitoring. AB - There is little published on the diagnostic value of short duration outpatient video electroencephalographic (VEEG) monitoring in children. The authors performed a prospective study on 37 patients (mean age = 10.4 years), with daily paroxysmal events who underwent short duration (mean = 3.2 hours) outpatient VEEG monitoring. Events were detected in 23 patients (62.2%), and a change in management as a result of outpatient VEEG monitoring was documented in 25 patients (67.6%). Despite the short duration of the outpatient VEEG in this study, the detection rate was comparable with the previously reported studies with longer duration monitoring. The authors found it convenient for the patient and less costly. The study demonstrated that short duration outpatient VEEG monitoring was able to differentiate between seizures and nonseizures in 11 patients (78.6%) and resulted in changing seizure classification in five patients (62.5%), and in selecting epilepsy surgery candidates in nine patients (60%). Short duration outpatient VEEG is useful as a diagnostic tool in patients with daily paroxysmal events, particularly in identifying nonepileptic events. PMID- 10513690 TI - Methylphenidate effects on global and complex measures of EEG. AB - Methylphenidate (MPH) effects on global and complex measures of electroencephalography were examined in boys with attention-deficit-hyperactivity disorder between the ages of 9 and 11 years. Electroencephalogram (EEG) data were collected separately from the administration of a continuous performance task and were evaluated for changes in overall frequency, coherence, phase, and asymmetry and against a referential database. MPH did not produce a clear change in EEG frequency measures compared with the task condition, although it did induce regional changes in the EEG and produced an improvement in task performance. In comparison against the referential database, MPH appeared to lessen the impact of abnormalities in EEG coherence, EEG phase, and EEG asymmetry on performance measures. PMID- 10513691 TI - Re-evaluation of the hypoxia theory as the mechanism of hyperventilation-induced EEG slowing. AB - To determine whether the well-accepted hypoxia theory accounts for hyperventilation-induced electroencephalogram (EEG) slowing, the authors monitored changes in cerebral oxygenation and end-tidal concentrations of carbon dioxide in 67 patients with epilepsy (age range = 5-12 years) during the hyperventilation activation test in a routine EEG examination. Relative concentration changes in cerebral oxygenated, deoxygenated, total hemoglobin, and oxidized cytochrome oxidase were measured by near-infrared spectroscopy in the frontal region. In all patients, except one who demonstrated EEG slowing, total and oxygenated hemoglobin decreased, and cytochrome oxidase was not reduced. EEG slowing occurred intermittently in 22 patients and was not synchronous with changes in either the cerebral oxygenation or end-tidal concentration of carbon dioxide. The degree of EEG slowing was diminished or the slow waves disappeared abruptly within 1 second after the cessation of hyperventilation in 22 patients when both the cerebral oxygenation and end-tidal concentration of carbon dioxide were still at low levels. The findings during the recovery periods do not confirm the hypoxia theory. It is thus supposed that more subtle mechanisms are the cause of EEG slowing. PMID- 10513693 TI - Power Doppler ultrasound imaging in neonatal cerebral venous sinus thrombosis. AB - Neonatal sinus thrombosis is a rare occurrence in sick neonates. Because of its nonspecific manifestations, the incidence is underestimated. This disease may not be demonstrated by conventional color Doppler and is diagnosed by computed tomography or magnetic resonance imaging. The authors report a neonate with neonatal sinus thrombosis diagnosed by power Doppler and suggest that the technique may be used as a less expensive and more available screening and follow up method in high-risk neonates. PMID- 10513692 TI - Newborn radial nerve palsy: report of four cases and review of published reports. AB - Four newborns presented with isolated radial nerve palsy during the first 2 days of life. In three, there was a history of failure of progression of labor, which may have resulted in prolonged radial nerve compression. Furthermore, three infants had fat necrosis of the upper arm above the elbow, suggestive of compression of the radial nerve in the region of the spiral groove. Significant recovery of function was evident within 1 month in all four infants. The authors review published reports about the rare condition of isolated radial nerve palsy in the newborn. PMID- 10513694 TI - Congenital fiber type disproportion: severe form with marked improvement. AB - A 30-month-old male exhibited marked hypotonia at birth accompanied by respiratory distress necessitating ventilator support. He subsequently demonstrated marked improvement in muscle power. He became independent of the respirator at 21 days of age and was able to sit without support at 11 months and walked alone at 24 months. Histopathologic analysis of the quadriceps femoris muscle confirmed the diagnosis of congenital fiber type of disproportion at 11 months of age. No other studies have described a patient with a severe neonatal form of congenital fiber type of disproportion who demonstrated such clear improvement. Physicians should be aware of this possibility when they interact with such patients and their families. PMID- 10513695 TI - Fetal brain infection with human parvovirus B19. AB - Intrauterine parvovirus B19 infection is known to be one of the causes of hydrops fetalis. However, there are few reports of the pathologic changes in the central nervous system. Postmortem examination of a fetus revealed multinucleated giant cells of macrophage/microglia lineage and many small calcifications around the vessels, predominantly in the cerebral white matter. Parvovirus B19 genome DNA was detected in the nucleus of the multinucleated giant cells and solitary endothelial cells by polymerase chain reaction amplification and in situ polymerase chain reaction methods. Capsid antigen was also demonstrated in the cytoplasm of the endothelial cells by immunofluorescent assay. Thus, intrauterine B19 parvovirus infection could be associated with marked neuropathologic changes in the fetal brain at the midembryonal period. Neurologic follow-up of complications may be necessary for children who survive the intrauterine infection. PMID- 10513696 TI - Epileptic negative myoclonus induced by carbamazepine in a child with BECTS. Benign childhood epilepsy with centrotemporal spikes. AB - A 7-year-old female with benign childhood epilepsy with centrotemporal spikes developed epileptic negative myoclonus (ENM) seizures during carbamazepine (CBZ) treatment. She had experienced nocturnal partial seizures since 5 years of age. Interictal electroencephalography demonstrated typical rolandic discharges. Valproate was first initiated at 6 years of age, but the seizures were uncontrollable. Carbamazepine was added and valproate withdrawn. The frequency of partial seizures did not decrease. Moreover, she had brief episodes of tone loss in each or both arms and eye blinking several weeks after CBZ introduction. Unilateral loss of arm tone corresponded to spike-and-wave discharges in the contralateral centrotemporal region, and a loss of tone in arms was associated with bilateral synchronous discharges. Eye blinking was also related to bilateral synchronous discharges and classified as a myoclonic seizure. The ENM and myoclonic seizures disappeared soon after CBZ withdrawal. Therefore the authors concluded that CBZ induced the ENM and myoclonic seizures in this patient. CBZ sometimes induces generalized seizures in the treatment of partial epilepsy and generalized epilepsy. CBZ-induced ENM seizures should be considered when a brief lapse of tone appears during CBZ treatment. PMID- 10513697 TI - Influenza A encephalitis with movement disorder. AB - Influenza A is an uncommon but well-recognized cause of viral encephalitis in childhood, occurring most commonly during community influenza outbreaks. The authors report four cases of influenza A encephalitis that occurred during an Australian epidemic in 1997-1998. Choreoathetosis during the acute phase of infection or basal ganglia involvement on neuroimaging was observed in three of the four patients. These findings in pediatric encephalitis are suggestive of influenza A infection and may guide investigation and early diagnosis. PMID- 10513698 TI - Hydrocephalus associated with glycogen storage disease type II (Pompe's disease). AB - The authors describe a case of hydrocephalus in an 8-month, 2-week-old infant who had been previously diagnosed with glycogen storage disease type II. Cranial imaging revealed no evidence of obstruction within the ventricular system. This case adds to the central nervous system complications associated with this disorder. Several possible mechanisms for the hydrocephalus observed in this infant are discussed. PMID- 10513699 TI - Patenting the human genome. PMID- 10513700 TI - Reduction of organ-retrieval damage and organ-discard rates. PMID- 10513701 TI - PANDAS and immunomodulatory therapy. PMID- 10513702 TI - Time to register randomised trials. PMID- 10513703 TI - Reappraisal of dermatoses of pregnancy. PMID- 10513704 TI - Epstein-Barr-virus infection and persistence: a B-cell marriage in sickness and in health. PMID- 10513705 TI - Treatment of children with malnutrition and diarrhoea. PMID- 10513706 TI - Kidney damage during organ retrieval: data from UK National Transplant Database. Kidney Advisory Group. AB - BACKGROUND: Kidney damage at organ retrieval is believed to be an increasing problem that is under reported. We aimed to identify the true rate of such damage and assess the effects on transplant survival. METHODS: Data from the UK National Transplant Database were analysed on all cadaveric kidneys donated over a 5-year period in the UK. Records indicated whether kidneys had been retrieved by a liver or renal surgical team and whether damage was noted at the time of retrieval or at the transplant procedure. Multivariate Cox's regression models were fitted to 1-year and 3-year transplant-survival data in those kidneys that were transplanted between 1992 and 1994. FINDINGS: Of 9014 kidneys retrieved, 96 could not be transplanted because of damage sustained at retrieval. Damage was reported in 1726 (19%) kidneys although by both donor and recipient centres in only 270 (3%). 1070 (62%) of the damaged organs were from donors aged 40 years or older. Reported kidney damage was more likely for retrievals of kidney only by a renal team (503 [26%]) than for multiorgan retrieval (454 [21%]), the proportion was lower when a liver team retrieved both liver and kidneys (415 [17%]). 794 (14%) kidneys retrieved and retained locally were reported as damaged, compared with 932 (29%) kidneys which had been exported. Donors' age had a significant effect on both 1-year and 3-year transplant survival (p<0.01 for both) but kidney damage did not (1 year p=0.40; 3 year p=0.81). INTERPRETATION: Despite the high rate of damage to kidneys at retrieval, most of the organs can be transplanted with no adverse effect on transplant survival. Kidney damage is least likely to occur with kidneys from young donors, and when liver teams or centres undertaking more than 50 retrievals per year do the retrieval. PMID- 10513707 TI - Analysis of factors that affect outcome of primary cadaveric renal transplantation in the UK. HLA Task Force of the Kidney Advisory Group of the United Kingdom Transplant Support Service Authority (UKTSSA) AB - BACKGROUND: In the UK, kidneys are exchanged between centres on the basis of matching for HLA. We analysed various factors that might affect graft outcome to establish whether exchange of kidneys on this basis remains valid. METHODS: 6363 primary cadaveric renal transplants carried out in 23 centres in the UK between 1986 and 1993 were used in the analysis. 6338 (99.6%) patients who underwent transplantation were followed up at 1 year. 5-year follow-up data were available for 2907 (97.8%) of the 2972 patients who survived to 5 years. We made random checks to validate the data. A multifactorial analysis with Cox's proportional hazards models was used to analyse factors that had a possible effect on graft outcome. To ensure that the analysis of matching was constant during the 8-year study, our analysis was based on the HLA antigens used for organ exchange (11 A locus antigens, 27 B locus antigens, and 12 DR locus antigens). We assessed overall outcome at 5 years and during three periods after transplantation at: 0-3 months, 3-36 months, and after 36 months. FINDINGS: The following factors were significantly associated with graft outcome in the multifactorial analysis: year of graft, age of donor, age of recipient, whether the recipient had diabetes, cause of donor's death, cold ischaemic time, transport of kidneys, transplant centre, and matching for HLA. The best outcome was achieved with kidneys that had no mismatches at HLA-A, HLA-B, and HLA-DR loci (000 mismatches). The next most favourable outcome was achieved with one mismatch at either A or B loci or one mismatch at both the A and B , but no mismatch at the DR locus (100, 010, or 110 mismatches). Age of the donor and recipient had a significant effect on transplant outcome: older age was associated with increased risk of graft failure. INTERPRETATION: Various factors affect the outcome of primary cadaveric renal transplantation, particularly the age of the donor and the recipient. However, the effect of matching for HLA remains a strong one and fully justifies the continuing policy in the UK of exchanging kidneys on the basis of HLA matching, especially to recipients when there is a 000 mismatch for HLA between donor and recipient. On the basis of this analysis, a new allocation scheme for kidneys was introduced in the UK in 1998. During the first 9 months of the scheme, there has been a doubling of the number of HLA-000 mismatched kidneys transplanted. PMID- 10513708 TI - Therapeutic plasma exchange and intravenous immunoglobulin for obsessive compulsive disorder and tic disorders in childhood. AB - BACKGROUND: In children, exacerbations of tics and obsessive symptoms may occur after infection with group A beta-haemolytic streptococci. If post-streptococcal autoimmunity is the cause of the exacerbations, then children might respond to immunomodulatory treatments such as plasma exchange or intravenous immunoglobulin (IVIG). We studied whether plasma exchange or IVIG would be better than placebo (sham IVIG) in reducing severity of neuropsychiatric symptoms. METHODS: Children with severe, infection-triggered exacerbations of obsessive-compulsive disorder (OCD) or tic disorders, including Tourette syndrome, were randomly assigned treatment with plasma exchange (five single-volume exchanges over 2 weeks), IVIG (1 g/kg daily on 2 consecutive days), or placebo (saline solution given in the same manner as IVIG). Symptom severity was rated at baseline, and at 1 month and 12 months after treatment by use of standard assessment scales for OCD, tics, anxiety, depression, and global function. FINDINGS: 30 children entered the study and 29 completed the trial. Ten received plasma exchange, nine IVIG, and ten placebo. At 1 month, the IVIG and plasma exchange groups showed striking improvements in obsessive-compulsive symptoms (mean improvement on children's Yale-Brown obsessive compulsive scale score of 12 [45%] and 13 [58%], respectively), anxiety (2.1 [31%] and 3.0 [47%] improvement on National Institute of Mental Health anxiety scale), and overall functioning (2.9 [33%] and 2.8 [35%] improvement on National Institute of Mental Health global scale). Tic symptoms were also significantly improved by plasma exchange (mean change on Tourette syndrome unified rating scale of 49%). Treatment gains were maintained at 1 year, with 14 (82%) of 17 children "much" or "very much" improved over baseline (seven of eight for plasma exchange, seven of nine for IVIG). INTERPRETATION: Plasma exchange and IVIG were both effective in lessening of symptom severity for children with infection-triggered OCD and tic disorders. Further studies are needed to determine the active mechanism of these interventions, and to determine which children with OCD and tic disorders will benefit from immunomodulatory therapies. PMID- 10513709 TI - Unrecognised Mycobacterium tuberculosis bacteraemia among hospital inpatients in less developed countries. AB - BACKGROUND: Nosocomial transmission of Mycobacterium tuberculosis is a global public-health concern. Although early clinical recognition of M. tuberculosis in hospital inpatients is critical for effective infection control, such recognition may be difficult in patients with HIV infection. To find out whether M. tuberculosis bacteraemia frequently goes unrecognised, we did a prospective blood culture survey in an infectious-diseases hospital in Thailand and a general hospital in Malawi. METHODS: Consecutive febrile (> or = 37.5 degrees C axillary or > or = 38.0 degrees C orally) hospital inpatients (aged > or = 18 years) were enrolled; blood was obtained for mycobacterial culture and HIV testing. Simple diagnostic tests, such as chest radiographs and sputum smears, were ordered by clinicians as deemed necessary, and were carried out with existing local resources. FINDINGS: Of 344 patients enrolled, 255 (74%) were HIV infected, the median age was 33 years (range 18-87), and 208 (61%) were male. 34 (10%) patients had M. tuberculosis bacteraemia; five of these patients were already on antituberculosis therapy. Only HIV-infected patients had M. tuberculosis bacteraemia. Of the 29 patients with M. tuberculosis bacteraemia who were not already receiving antituberculosis therapy, 13 (45%) had an abnormal chest radiograph or a positive sputum smear. 16 (55%) patients had no additional diagnostic test results to indicate M. tuberculosis infection; 18 (81%) of these had a cough. INTERPRETATION: In less developed countries where both M. tuberculosis and HIV infections are prevalent, M. tuberculosis bacteraemia may frequently go unrecognised among febrile hospital inpatients. PMID- 10513710 TI - Moderate hypothermia for uncontrolled intracranial hypertension in acute liver failure. AB - BACKGROUND: Increased intracranial pressure as a complication of acute liver failure has a mortality of about 90% in patients who do not respond to treatment with mannitol and ultrafiltration. We investigated the safety and efficacy of moderate hypothermia for uncontrolled increase in intracranial pressure in patients with acute liver failure. METHODS: We studied seven consecutive patients aged 16-46 years (five women, four candidates for orthotopic liver transplantation [OLT]) with acute liver failure who fulfilled criteria for poor prognosis liver failure and had increased intracranial pressure that was unresponsive to two treatments with mannitol and ultrafiltration. We used cooling blankets to lower the patients' core temperature to 32-33 degrees C. Patients who were not suitable candidates for OLT (patients 1-3) were cooled for 8 h and then gradually rewarmed over 1 h to a baseline temperature of 37 degrees C. Patients who were suitable candidates for OLT (patients 4-7) were cooled before and during the OLT procedure. We measured cerebral blood flow and metabolic indices before and after cooling. FINDINGS: The four patients who were candidates for OLT were successfully maintained until transplantation with 13 (range 10-14) h of hypothermia. The three patients who were unsuitable candidates for OLT died after rewarming. Intracranial pressure before cooling was 45 (25-49) mm Hg and was reduced in all patients to 16 (13-17) mm Hg (p<0.05). Cerebral blood flow decreased from 103 (25-134) mL 100 g(-1) min(-1) before cooling to 44 (24-75) mL 100 g(-1) min(-1) after cooling (p<0.05). The corresponding changes for cerebral perfusion pressure was an increase from 45 (37-56) mm Hg to 70 (60-78) mm Hg (p<0.05) and for cardiac index a decrease from 9.8 (7-13) to 5.1 (4.3-6.1) L per min per m2 of body surface area. During hypothermia there was no significant relapse of increased intracranial pressure. Arterial ammonia and cerebral uptake of ammonia were significantly reduced with cooling. No adverse effects of hypothermia were observed. INTERPRETATION: Moderate hypothermia is useful in the treatment of uncontrolled increase in intracranial pressure in patients with acute liver failure and may serve as a bridge to OLT. PMID- 10513711 TI - Association of Rickettsia helvetica with chronic perimyocarditis in sudden cardiac death. AB - BACKGROUND: Rickettsia helvetica is the only non-imported rickettsia found in Scandinavia. It was first detected in Ixodes ricinus ticks, but has never been linked to human disease. We studied two young Swedish men who died of sudden cardiac failure during exercise, and who showed signs of perimyocarditis similar to those described in rickettsial disease. METHODS: Samples from the heart and other organs were analysed by PCR and DNA sequencing. May-Grunwald-Giemsa, Grocott, and acridine-orange stains were used for histopathological examinations. Staining of R. helvetica grown on shell-vials in vero cells, and the early descriptions of R. rickettsii by H T Ricketts and S B Wohlbach served as controls. Immunohistochemistry was done with Proteus OX-19 rabbit antisera as the primary antibody. The structure of rickettsia-like organisms was investigated by transmission electron microscopy. Serological analyses were carried out by indirect immunofluorescence with R. helvetica as the antigen. FINDINGS: By use of a semi-nested PCR, with primers specific for the 16S rRNA and 17-kDa outer membrane-protein genes, and sequence analysis of the amplified products, genetic material from R. helvetica was detected in the pericardium and in a lymph node from the pulmonary hilum in case 1, and in a coronary artery and the heart muscle in case 2. A serological response in case 1 revealed an endpoint titre for R. helvetica of 1/320 (1/256 with R. rickettsii as the antigen). Examination of PCR positive tissue showed chronic interstitial inflammation and the presence of rickettsia-like organisms predominantly located in the endothelium. These organisms reacted with Proteus OX-19 antisera, and their size and form were consistent with rickettsia. Electron microscopy confirmed that the appearance of the organisms was similar to that described for spotted-fever rickettsia. INTERPRETATION: R. helvetica, transmitted by I. ricinus ticks, may be an important pathogen in the aetiology of perimyocarditis, which can result in sudden unexpected cardiac death in young people. PMID- 10513712 TI - A "little cough" for 40 years. PMID- 10513713 TI - Evidence of AIDS-related mortality in Mumbai, India. AB - From an estimated 85,200 HIV-infected individuals in Mumbai in 1997, at least 4120 excess deaths attributed to AIDS occurred among 15-54-year-olds. To prevent repetition of this excess in other parts of India, priority intervention programmes should be instituted quickly because the window of opportunity is closing quickly. PMID- 10513714 TI - Intoxication due to negative canrenone interference in digoxin drug monitoring. AB - Canrenone and spironolactone caused falsely low readings in a common assay for digoxin (AxSym MEIA) due to negative cross-reactivity. Misleading subtarget concentrations were repeatedly reported, and falsely guided drug dosing resulted in a case of digoxin intoxication. PMID- 10513715 TI - Adverse effects of contact isolation. AB - Health-care workers are half as likely to enter the rooms of patients in contact isolation, but are more likely to wash their hands after caring for them than after caring for patients not in isolation. PMID- 10513716 TI - Tropheryma whippelii DNA in saliva of healthy people. AB - In a random sample of 40 healthy people, 35% showed evidence of Tropheryma whippelii DNA in their saliva. Consistent detection of T. whippelii DNA on repeated sampling suggests that this organism can be an oral commensal. PMID- 10513717 TI - Incidence of paediatric Crohn's disease in Stockholm, Sweden. AB - We report an increase in the incidence of Crohn's disease in northern Stockholm, Sweden. PMID- 10513718 TI - Oxaliplatin-induced haemolytic anaemia. AB - A case of haemolytic anaemia after therapy with oxaliplatin, an anticancer chemotherapeutic agent, was investigated. Haemolytic anaemia has been associated with cisplatin and carboplatin, two related drugs, but not with oxaliplatin. PMID- 10513719 TI - Clozapine-induced acute interstitial nephritis. AB - Drug hypersensitivity reactions commonly cause acute interstitial nephritis (AIN). Clozapine, a new antipsychotic, can cause fatal bone-marrow toxicity. We report clozapine-induced AIN as another serious adverse drug reaction. PMID- 10513720 TI - Taxane-induced glaucoma. AB - We report a case of glaucoma induced by doxetaxel therapy for metastatic breast cancer. The disorder recurred during treatment with paclitaxel. PMID- 10513721 TI - Hyperkalaemia in a Thai man. AB - Blood specimens from a Thai man showed greatly increased time-dependent and temperature-dependent efflux of potassium from cells into serum. No in-vivo correlate was found, and no similar results were obtained in screening 131 other Thai men. Findings may be related to decreased Na,K-ATPase density or activity. PMID- 10513722 TI - Phyto-oestrogens get more attention at menopause meeting. PMID- 10513723 TI - David Hill: behavioural insights into cancer control. PMID- 10513724 TI - The looming threat of AIDS and HIV in Latin America. PMID- 10513725 TI - In fighting over HIV-positive personnel in the Indian navy. PMID- 10513726 TI - Leishmaniasis. AB - In 1903, Leishman and Donovan separately described the protozoan now called Leishmania donovani in splenic tissue from patients in India with the life threatening disease now called visceral leishmaniasis. Almost a century later, many features of leishmaniasis and its major syndromes (ie, visceral, cutaneous, and mucosal) have remained the same; but also much has changed. As before, epidemics of this sandfly-borne disease occur periodically in India and elsewhere; but leishmaniasis has also emerged in new regions and settings, for example, as an AIDS-associated opportunistic infection. Diagnosis still typically relies on classic microbiological methods, but molecular-based approaches are being tested. Pentavalent antimony compounds have been the mainstay of antileishmanial therapy for half a century, but lipid formulations of amphotericin B (though expensive and administered parenterally) represent a major advance for treating visceral leishmaniasis. A pressing need is for the technological advances in the understanding of the immune response to leishmania and the pathogenesis of leishmaniasis to be translated into field-applicable and affordable methods for diagnosis, treatment, and prevention of this disease. PMID- 10513727 TI - Three cases of bone and joint surgery in the 14th century. PMID- 10513728 TI - Are drugs within a class interchangeable? PMID- 10513729 TI - Early Lung Cancer Action Project. PMID- 10513730 TI - Early Lung Cancer Action Project. PMID- 10513731 TI - Early Lung Cancer Action Project. PMID- 10513732 TI - Early Lung Cancer Action Project. PMID- 10513733 TI - Early Lung Cancer Action Project. PMID- 10513734 TI - Varicella zoster virus immunisation. PMID- 10513735 TI - Lowering of LDL cholesterol. PMID- 10513736 TI - Implication of fibrate therapy for homocysteine. PMID- 10513737 TI - Implication of fibrate therapy for homocysteine. PMID- 10513738 TI - Implication of fibrate therapy for homocysteine. PMID- 10513739 TI - Diagnosis of Helicobacter pylori infection by HpSA test. PMID- 10513740 TI - Diagnosis of Helicobacter pylori infection by HpSA test. PMID- 10513741 TI - Diagnosis of Helicobacter pylori infection by HpSA test. PMID- 10513742 TI - Tamoxifen for intraductal cancer. PMID- 10513743 TI - Obstructive sleep apnoea. PMID- 10513744 TI - Obstructive sleep apnoea. PMID- 10513745 TI - Obstructive sleep apnoea. PMID- 10513746 TI - Charles Dickens' work to help establish Great Ormond Street Hospital. PMID- 10513747 TI - Charles Dickens' work to help establish Great Ormond Street Hospital. PMID- 10513748 TI - Medical indemnities in Ireland. PMID- 10513749 TI - Myth, magic, and the history of modern medicine. PMID- 10513750 TI - The Nobel chronicles. 1974: Albert Claude (1899-1983), George Emil Palade (b 1912), and Christian Rene de Duve (b 1917). PMID- 10513751 TI - P.A. Schleisner: a pioneer in epidemiology. AB - In 1847 Schleisner (b. 1818) was sent from Copenhagen to the Vestmanna Islands in Iceland to study the epidemic of tetanus neonatorum. The neonatal mortality in those islands at that time was 600-740 per 1000 children. He built a small hospital and introduced treatment with Peru balsam of the umbilicus. Schleisner probably assumed that the infection was caused by airborne infection, contact infection or poor hygiene. The neonatal mortality fell to about 50 per 1000. Schleisner published his results in 1849. Semmelweiss published his observations in 1850 and Snow successfully fought the cholera epidemic in London in 1854. Schleisner deserves recognition as a pioneer in the field of epidemiology. PMID- 10513752 TI - Problems with using capture-recapture in epidemiology: an example of a measles epidemic. AB - Capture-recapture is becoming widely used in epidemiology to estimate disease prevalence or sizes of population at risk. When such estimates are obtained from uncontrolled observation of existing lists, a huge act of faith is required, usually without any scientific justification. Fitting of loglinear models appears to offer some hope but contains major problems of analysis and interpretation. These are illustrated by reanalysis of data on a measles epidemic-see McGilchrist et al. 1996 [J Clin Epidemiol 49, pp. 293-296]--for which the wrong model was selected. It is argued that the measles lists contained so few overlaps that no reliable information is provided by that study about the size of the epidemic. PMID- 10513753 TI - Model selection and population size using capture-recapture methods. PMID- 10513754 TI - Recommendations for presentation and evaluation of capture-recapture estimates in epidemiology. AB - We propose 15 recommendations for approaches to capture-recapture analysis in epidemiology. We apply them to a report of such an analysis of a measles epidemic [McGilchrist et al., J Clinical Epidemiol 1996, 49: 293-296] and to comments thereon by R. C. Cormack [J Clinical Epidemiol 1999; 52: 909-914]. The latter challenged the utility of the data on the measles outbreak for any reliable capture-recapture estimates. We suggest that, adopting the perspective of W. Edwards Deming, one can only make judgments as to the reliability of capture recapture data, methods, and derived estimates in the light of (i.e., conditional upon) their eventual intended use. Capture-recapture approaches "unreliable" from one perspective may be "reliable," and/or more appropriately, "useful" from another. We consider the utility of ancillary and ad hoc information that may be available or worth seeking to supplement a capture-recapture analysis. We use information within the study of McGilchrist et al. to illustrate how, with such ancillary information, one may overcome the main thrust of the objections of Cormack in situations in which one observes apparently anomalous or hard to understand data structures. Making certain simple assumptions we regard as plausible, we estimate the number of affected in the measles epidemic as between about 700-1300. We derive this from data on 502 cases in a Register, an ad hoc sample of 91 cases in one age group in the general population, and the report of 41 cases in both of these. Our result is only 15-30% the total implied by the estimates McGilchrist et al. derived with more complex methods and many assumptions in addition to our own. We discuss various approaches to evaluating "reliability" of our estimate conditional upon intended uses by policy makers. PMID- 10513755 TI - The importance of source selection and pilot study in the capture-recapture application. PMID- 10513756 TI - Stepwise selection in small data sets: a simulation study of bias in logistic regression analysis. AB - Stepwise selection methods are widely applied to identify covariables for inclusion in regression models. One of the problems of stepwise selection is biased estimation of the regression coefficients. We illustrate this "selection bias" with logistic regression in the GUSTO-I trial (40,830 patients with an acute myocardial infarction). Random samples were drawn that included 3, 5, 10, 20, or 40 events per variable (EPV). Backward stepwise selection was applied in models containing 8 or 16 pre-specified predictors of 30-day mortality. We found a considerable overestimation of regression coefficients of selected covariables. The selection bias decreased with increasing EPV. For EPV 3, 10, or 40, the bias exceeded 25% for 7, 3, and 1 in the 8-predictor model respectively, when a conventional selection criterion was used (alpha = 0.05). For these EPV values, the bias was less than 20% for all covariables when no selection was applied. We conclude that stepwise selection may result in a substantial bias of estimated regression coefficients. PMID- 10513757 TI - Meta-analysis of diagnostic tests with imperfect reference standards. AB - We present a method to estimate the summary receiver operating characteristic (SROC) curve for combining information on a diagnostic test from several different studies. Unlike previous methods that assume the reference standard to be error free, our approach allows for the possibility of errors in the reference standard, through use of a latent class model. The model provides estimates of the sensitivity and specificity of the diagnostic test and the case prevalence in each study; these parameters can then be used in a meta-analysis, for example, using the regression method proposed by Moses et al., of a measure of test discrimination on a measure of the diagnostic threshold, to fit the SROC. The method is illustrated with an example on Pap smears that shows how adjusting for imperfection in the reference standard typically reduces the scatter of data in the SROC plot, and tends to indicate better performance of the test than otherwise. PMID- 10513758 TI - Cervical screening in Africa: discordant diagnosis in a double independent reading. DYSCER-CI Group. AB - Interobserver variation in the cytological diagnosis of cervical lesions poses a problem for public health screening programs. This study assessed the frequency of discordant diagnoses between two independent cytopathologists in the screening of African women. In Abidjan, Cote d'Ivoire, 2157 women were recruited from three outpatient gynecology clinics and screened for cervical abnormalities and genital and human immunodeficiency virus (HIV) infections. The degree of agreement between the cytopathologists was assessed by kappa statistics. The overall agreement was poor (kappa = 0.33); however, the degree of agreement increased with the severity of the lesions and was fairly good (kappa = 0.53) for high grade and invasive lesions requiring curative treatment. Discordance was associated with HIV infection but not with genital infections. For a prevention program of cervical cancer in this African context, strategies must be developed to minimize errors in cervical screening. Particularly, HIV-infected women require a systematic rereading to reduce false-negative results. PMID- 10513759 TI - Validation of the Agency for Health Care Policy and Research (AHCPR) classification for managing unstable angina. AB - To validate the AHCPR classification for the prognosis of unstable angina, 225 consecutive patients were recruited with a suspected diagnosis of that condition attending a tertiary hospital from November 1994 through April 1995 and followed for one year. One-hundred fifty-six (69.3%) patients were considered at high risk, 37 (16.5%) at intermediate, and 32 (14.2%) at low risk of cardiac complications. All of the patients with major in-hospital cardiac complications (8 patients) had at least one of the features of the high risk group. The high to intermediate-low hazard ratio (HR) for one-year cardiac complications after the onset of unstable angina was 4.03. Predictors of major complications (myocardial infarction or death) after the follow-up were age > 65 (HR, 5.69); diabetes (HR, 4.94); heart failure (HR, 2.65); and prolonged angina (HR, 2.55). AHCPR classification correctly identified patients with risk of severe outcomes at the hospital. Also, the classification predicted outcomes one year after hospitalization, diabetes being an important determinant of adverse cardiac events. PMID- 10513760 TI - The level of alcohol consumption at which all-cause mortality is least. AB - Moderate consumers of alcohol have lower mortality than either nondrinkers or heavy drinkers. This systematic review aimed to quantify the level of alcohol consumption (termed the nadir) at which the lowest mortality occurs. Twenty cohort studies reported analyses of all-cause mortality for at least three categories of alcohol consumption, giving a total of 60,224 deaths among men and 74,824 deaths among women. The nadir in each study was estimated for men and women separately in units per week, where 1 unit is 9 g of alcohol. The estimated nadirs varied substantially between countries. Combined nadirs were estimated for U.S. men (overall nadir 7.7 units per week, 95% confidence interval [CI] 6.4 9.1), U.K. men (12.9 units per week, 95% CI 10.8-15.1), and U.S. women (2.9 units per week, 95% CI 2.0-4.0). The nadirs were not found to be increased in studies of older persons and apply for ages 50 to 80 years. PMID- 10513761 TI - Therapeutic drug use during pregnancy: a comparison in four European countries. OECM Working Group. Occupational Exposures and Congenital Anomalies. AB - A drug utilization study was performed using data of the OECM study on Occupational Exposures and Congenital Malformations, which was conducted in six European Registries of Congenital Anomalies (two in France, two in Italy, one in Great Britain, and one in The Netherlands): the mothers were interviewed after delivery for exposures during pregnancy, including use of therapeutic drugs. The analysis of drug use considered only the 1134 control mothers of healthy newborns, and focused on the first trimester of pregnancy: 36.2% of the interviewed mothers used at least one drug (excluding vitamins and minerals) during the first trimester. This rate varied from 22.5% in Glasgow to 50.3% and 44.2% in the French centers. Anti-infectives were the most frequent drugs (12.3% of mothers), then antinauseants (10.6%), and treatments for threatened abortion (5.5%). Important variations between countries were observed, reflecting different medical attitudes towards drug use during pregnancy. PMID- 10513762 TI - Stress, social relations, and old age mortality in Taiwan. AB - The research analyzed the relationships among stress, social relations, and mortality in a probability sample of 4,049 Taiwanese adults, aged 60 and over. The baseline survey was conducted in 1989 and the survival status of the respondents was ascertained during the subsequent 4 years. Death of a spouse or a child was found to increase the risk of dying directly and indirectly, whereas major financial difficulty during the past 5 years and current financial strain influenced mortality indirectly through their effects on self-rated health disability. In addition to their direct effect on mortality, martial status and work status lowered the probability of dying through decreased disability and subjective ill health. Finally, no buffering effects of social support were substantiated. PMID- 10513763 TI - Comparison of open and closed questionnaire formats in obtaining demographic information from Canadian general internists. AB - The objective of this study was to compare the impact of closed- versus open ended question formats on the completeness and accuracy of demographic data collected in a mailed survey questionnaire. We surveyed general internists in five Canadian provinces to determine their career satisfaction. We randomized respondents to receive versions of the questionnaire in which 16 demographic questions were presented in a closed-ended or open-ended format. Two questions required respondents to make a relatively simple computation (ensuring that three or four categories of response added to 100%). The response rate was 1007/1192 physicians (80.0%). The proportion of respondents with no missing data for all 16 questions was 44.7% for open-ended and 67.0% for closed-ended formats (P < 0.001). The odds of having missing items remained higher for open-ended response options after adjusting for a number of respondent characteristics (2.67, 95% confidence interval 2.01 to 3.55). For the two questions requiring computations focused on professional activity and income, there were more missing data (P = 0.02, 0.02, respectively) but fewer inaccurate responses (P = 0.009, 0.20, respectively) for the open-ended compared to the closed-ended format. Investigators can achieve higher response rates for demographic items using closed format response options, but at the risk of increasing inaccuracy in response to questions requiring computation. PMID- 10513764 TI - Consumption of soft drinks with phosphoric acid as a risk factor for the development of hypocalcemia in postmenopausal women. AB - The objective of this study was to determine the relationship between the consumption of phosphoric acid-containing soft drinks and hypocalcemia in postmenopausal women. A case control study was designed to include 21 cases and 64 controls, matched by age and menopausal duration with similar family income, scholarship, and daily dietary intakes. Clinical and dietetic conditions that may produce hypocalcemia were considered as exclusion criteria. Cases were defined as a serum Ca level < or = 8.8 mg/dl, and controls as a serum CA level > 8.8 mg/dl. Women in the case group had a higher consumption of phosphoric acid-containing soft drink, and showed increased serum levels of PTH and hyperphosphaturia, than those in the control group without significant differences in 1,25(OH)2D3. In the multivariate regression analysis consumption of one or more bottles per day of cola soft drinks showed association with hypocalcemia (1.28, CI 95% 1.06-1.53). The consumption of soft drinks with phosphoric acid should be considered as an independent risk factor for hypocalcemia in postmenopausal women. PMID- 10513765 TI - Reflections on the science of therapeutics. PMID- 10513766 TI - Thrombophilias: diagnosis and treatment of thrombophilia relating to contraception and pregnancy. AB - The thrombogenicity of oral contraceptives (OCs) has been known since the 1960s. The thrombotic risk remains increased even with the third-generation preparations containing low-dose estrogen and a progesterone. Thromboembolic disorders are a leading cause of maternal morbidity and mortality during pregnancy and puerperium in Western countries. Women with thrombophilic conditions are even more prone to hypercoagulability when using hormone therapy, or during pregnancy and puerperium. At present, it is not recommended that all OC users or pregnant women be routinely screened for thrombophilic abnormalities. The usefulness of testing for these disorders in the presence of multiple thrombotic risk factors should be considered on an individual basis. Anticoagulant treatment may be advised for selected patient populations. PMID- 10513767 TI - Systemic causes of excessive uterine bleeding. AB - In assessing a patient with excessive uterine bleeding, the clinician should consider systemic causes in the differential diagnosis. Both hereditary and acquired conditions can result in mucous membrane bleeding, including menorrhagia, epistaxis, and gum bleeding, as well as excessive bruising. Among hereditary conditions, von Willebrand disease (vWD) is by far the most common, affecting an estimated 1% of the population worldwide. It is important to consider the possibility of vWD, and to establish the proper diagnosis (including subtype), as safe, effective, and easy-to-use treatment is available for most persons with this disorder. This review also covers a number of other systemic conditions that can be manifested by excessive uterine bleeding, including congenital deficiency of factor XI, idiopathic thrombocytopenic purpura and other acquired platelet disorders, acquired autoantibodies against factor VIII (FVIII), and vitamin K deficiency states. PMID- 10513768 TI - Women and inherited bleeding disorders: menstrual issues. AB - Von Willebrand, who described a bleeder family from Aland in 1926 in whom the index patient died at the fourth menses, stated, "the trait seems to be seen among women." Menorrhagia is defined objectively as a menstrual blood loss of at least 80 mL. However, even though 5% of women aged 30 to 49 years consult their general practitioner and 12% of gynecologic referrals are for menorrhagia, the diagnosis is difficult. A pictorial bleeding assessment chart (PBAC) has a specificity and sensitivity of more than 80%, with a score of > or = 100 being equivalent to more than 80 mL of blood loss. Using this chart to screen 150 women with menorrhagia who attended a gynecology clinic, an inherited bleeding disorder was diagnosed in 17%. Menorrhagia with onset at the menarche was predictive of an inherited bleeding disorder in 65% of von Willebrand's disease (vWD) and 67% of factor XI (FXI)-deficient patients. A retrospective survey in patients with inherited bleeding disorders using the PBAC showed menorrhagia in 74% of patients with vWD, 57% of carriers of hemophilia A or B, and 59% of FXI-deficient patients, compared with 29% of control individuals. Menorrhagia was the most common symptom in 60% of patients with FVII deficiency studied. Menorrhagia in women with bleeding disorders can be controlled with tranexamic acid with good effect. More recently, a desmopressin acetate (DDAVP) spray has been shown to achieve good FVIII and von Willebrand factor (vWF) levels and is efficacious for women with these deficiencies. The oral contraceptive pill may be useful. Since bleeding disorders are found in a substantial number of women with menorrhagia, it is important that such patients are investigated for these disorders before invasive procedures are done; hysteroscopy and hysterectomy in these patients are associated with a high rate of postoperative bleeding. PMID- 10513769 TI - Women and inherited bleeding disorders: pregnancy and delivery. AB - The most common inherited bleeding disorders in women are von Willebrand disease (vWD), carriership of hemophilia A and B, and factor XI (FXI) deficiency. Pregnancy and delivery are associated with major concerns and particular risks in women with these disorders. An increased awareness among clinicians of these disorders and their obstetric complications, a multidisciplinary approach to management, close collaboration between obstetricians and hemophilia centers, and the availability of management guidelines are essential to minimize maternal and neonatal complications. The special aspects of obstetric management include prenatal diagnosis, and antenatal, intrapartum, postpartum, and neonatal care. The issue of prenatal diagnosis is primarily considered in carriers of hemophilia because of severity of the disease in their male offspring and knowledge of genetic defects in most of affected families. The uptake of prenatal diagnosis, methods used, and technical aspects of invasive procedures are discussed. Hemostatic response to pregnancy is variable in different types of inherited bleeding disorders. Monitoring of coagulation status and appropriate prophylaxis, when indicated, is essential for safe pregnancy and delivery. Invasive intrapartum monitoring techniques and instrumental deliveries should be avoided. Delivery should be achieved by the least traumatic method to minimize the risk of postpartum hemorrhage and neonatal hemorrhagic complications. PMID- 10513770 TI - Improvement in three-month angiographic outcome suggested after primary angioplasty for acute myocardial infarction (Zwolle trial) compared with successful thrombolysis (APRICOT trial). Antithrombotics in the Prevention of Reocclusion In COronary Thrombolysis. AB - It has been shown that primary percutaneous transluminal coronary angioplasty (PTCA) for acute myocardial infarction results in higher patency rates than thrombolytic therapy. However, no data are available on differences in long-term angiographic outcome after successful primary PTCA compared with successful thrombolysis. Therefore, we compared angiographic data of the Antithrombotics in the Prevention of Reocclusion In COronary Thrombolysis (APRICOT) trial and the Zwolle primary PTCA trial. In the APRICOT trial 248 patients underwent coronary angiography at a mean of 24 hours after thrombolysis and had a patent infarct related vessel (Thrombolysis In Myocardial Infarction-3 trial flow) when entering the study. Reocclusion rates were assessed at a second angiography after 3 months. In the Zwolle trial 136 patients had a successful primary PTCA. At 3 months 131 patients underwent a second angiography. Quantitative coronary angiography showed a significant lower mean diameter stenosis of the infarct related vessel after primary PTCA (27 +/- 12% vs 57 +/-12%; p = 0.00001). At 3 months this difference was sustained (35 +/- 22% vs 63 +/- 26%; p = 0.00001). After thrombolysis the reocclusion rate at 3 months was 29% compared with 5% after primary PTCA (p = 0.0001). Results show that compared with successful thrombolytic therapy, primary PTCA for acute myocardial infarction results in an improved infarct-related vessel status not only short term but also long term, with a low reocclusion rate. PMID- 10513771 TI - Relation between systemic hypertension and blood lipids on the risk of myocardial infarction. AB - We sought to evaluate the potential interactions between systemic hypertension and blood lipids on the risk of myocardial infarction (MI). Recent evidence suggests that hypertension may interact with other risk factors such as dyslipidemia in the development of coronary heart disease. However, the precise nature of that interrelation remains unclear. We selected 340 cases of first MI and an equal number of age-, sex-, and community-matched controls. Data were collected on a large number of coronary risk factors, and fasting blood samples were obtained. Logistic regression was used to calculate the odds ratio (OR) of nonfatal MI. The age- and sex-adjusted OR of MI was 1.61 (95% confidence interval [CI] 1.15 to 2.25) among treated hypertensives compared with nonhypertensives. Further adjustment for coronary risk factors did not materially alter the results (OR 1.67, 95% CI 1.16 to 2.41). To explore the interrelations among hypertension, lipids, and risk of MI, each lipoprotein parameter was individually added to the risk factor-adjusted multivariate model. The apparent risk associated with hypertension was substantially attenuated by the addition of either high-density lipoprotein cholesterol (OR 1.25, 95% CI 0.82 to 1.90) or triglycerides (OR 1.37, 95% CI 0.91 to 2.05). No significant interactions were found between hypertension and any lipoprotein parameter. These data indicate that the risk of MI associated with treated hypertension may have a lipid mechanism involving high-density lipoprotein cholesterol and/or triglycerides. PMID- 10513772 TI - Cardiac angiotensin-converting enzyme activity in myocardial infarction. AB - The cardiac renin-angiotensin system is regarded as an important modulator in the infarct heart. Little is known about their presence and regulation in human hearts. We measured angiotensin-converting enzyme (ACE) and renin activities at the aortic root and anterior interventricular vein (AIV) in 51 patients with previous myocardial infarction (MI): anterior wall MI in 31 and inferior wall MI in 20 and 33 control subjects. In the anterior wall MI group, the serum ACE activity was increased significantly in the AIV than in the aortic root (16.2 +/- 5.3 vs 15.3 +/- 5.0 nmol/min/ml, p <0.001), whereas the activity was not different between the aortic root and AIV in the control (14.4 +/- 3.7 vs 14.4 +/ 3.7 nmol/min/ ml) and in the inferior wall MI (16.5 +/- 4.8 vs. 17.0 +/-5.2 nmol/min/ml) groups. On the other hand, there was no significant difference in plasma renin activity between the AIV and aortic root in the 3 groups (control group, 1.0 +/- 0.5 vs 1.0 +/- 0.5 pg/ml/hour; anterior wall MI group, 1.3 +/- 0.8 vs 1.3 +/- 0.8 pg/ml/hour; inferior wall MI group, 1.2 +/- 0.7 vs 1.3 +/- 0.8 pg/ml/ hour). The difference in serum ACE activity between the AIV and aortic root had a significant positive linear correlation with pulmonary capillary wedge pressure (r = 0.606, p <0.001), and had a significant negative linear correlation with left ventricular ejection fraction (r = -0.620, p <0.001) in the anterior wall MI group. Serum ACE activity from the infarct region of the left ventricle was augmented in patients with MI, and the activity was increased in proportion to the severity of left ventricular dysfunction. PMID- 10513773 TI - Effectiveness of early coronary angioplasty and abciximab for failed thrombolysis (reteplase or alteplase) during acute myocardial infarction (results from the GUSTO-III trial). Global Use of Strategies To Open occluded coronary arteries. AB - We evaluated the effects of abciximab treatment during early angioplasty after clinically failed thrombolysis for acute myocardial infarction. In the Global Use of Strategies To Open occluded coronary arteries (GUSTO-III) trial of reteplase versus alteplase for acute infarction (n = 15,059), 392 patients underwent angioplasty a median of 3.5 hours after thrombolysis and had complete procedural data. We compared 30-day mortality and in-hospital outcomes between patients who received abciximab (n = 83) and those who did not (n = 309), and (among patients given abciximab) between those randomized to alteplase versus reteplase. Patients given abciximab had anterior infarction less often, but were more often in Killip classes III or IV. The 30-day mortality rate tended to be lower with abciximab (3.6% vs 9.7%, p = 0.076), more so after adjustment for baseline differences (p = 0.042). The composite of death, stroke, or reinfarction did not differ significantly with abciximab treatment (12% vs 14%, p = 0.7), but it occurred less often among abciximab-treated patients who had been randomized to reteplase (n = 55) versus alteplase (n = 28) (7% vs 21%, p = 0.08). Severe bleeding was increased among abciximab-treated patients (3.6% vs 1.0%, p = 0.08), despite less heparin use. No intracranial hemorrhages occurred with abciximab. The use of abciximab for early angioplasty after clinically failed thrombolysis resulted in trends toward lower 30-day mortality and increased bleeding. PMID- 10513774 TI - Operative results of "repair" of ventricular septal rupture after acute myocardial infraction. AB - Fifty-four consecutive patients with postinfarction ventricular septal defect were reviewed. The rupture was closed with a patch and the left ventricle remodeled in all patients. Coronary artery bypass surgery was performed in 28 patients (52%). Fourteen patients (26%) died after operation and 19 during follow up (mean 42 months). Cumulative survival (including operative deaths) was 78%, 65%, and 40% at 1, 5, and 10 years, respectively. A short interval between septal rupture and operation was a risk factor for early mortality (p = 0.03). Treated associated coronary artery disease had no effect. A residual septal shunt, detected in 10 patients (18%), warranted reoperation in 7 and contributed to 2 early and 1 late death. The location and morphology of the septal rupture were not associated with increased risk of residual shunt. Thus, patch closure of the ventricular septal rupture, remodeling of the left ventricle to improve stroke volume and reduce wall stress, and selective myocardial revascularization provided acceptable results. PMID- 10513775 TI - Baseline clinical and angiographic variables associated with long-term outcome after successful intracoronary stent implantation. AB - Although randomized studies have demonstrated improved outcomes with stents over balloon angioplasty in straightforward coronary narrowings in low-risk patients, this advantage is less clear for complex lesions and high-risk patients. This study was designed to identify clinical and angiographic variables that are associated with long-term outcome after stent implantation. We identified 1,709 patients undergoing successful stent placement without in-hospital major adverse events. We analyzed clinical, lesional, and procedural variables to determine their correlation with outcome. Mean duration of follow-up was 1.6 +/- 1.4 years. Cox proportional-hazards models and stepwise methods were used to assess which covariates were potentially related to each end point. The occurrence of death/myocardial infarction (MI) was associated with any history of congestive heart failure (relative risk [RR] 3.3, 95% confidence interval [CI] 2.3 to 4.7, p <0.0001), procedure within 24 hours of MI (RR 2.3, CI 1.3 to 4.1, p = 0.0048), vein graft intervention (RR 1.8, CI 1.3 to 2.6, p = 0.0007), and prior MI (RR 1.8, CI 1.2 to 2.6, p = 0.004). Repeat revascularization was associated with multivessel stent placement (RR 1.8, CI 1.2 to 2.8, p = 0.006) and stent for abrupt closure (RR 1.7, CF 1.1 to 2.7, p = 0.03), but was less frequent with de novo lesions and right coronary artery lesions (RR 0.6, CI 0.5 to 0.8, p = 0.0007, and RR 0.8, CI 0.6 to 1.0, p = 0.05, respectively). The cumulative end point of death/MI/repeat revascularization was associated with congestive heart failure (RR 1.7, CI 1.3 to 2.2, p <0.0001), multivessel stent placement (RR 1.6, Cl 1.1 to 2.3, p = 0.03), warfarin therapy (RR 1.4, CI 1.2 to 1.8, p = 0.001), and procedure within 24 hours of MI (RR 1.5, CI 1.1 to 2.1, p = 0.02), but was less frequent with complete revascularization and right coronary artery intervention (RR 0.8, CI 0.7 to 0.99, p = 0.04, and RR 0.7, CI 0.6 to 0.9, p = 0.009, respectively). Thus, this study demonstrates that there are readily identifiable characteristics in patients treated successfully with stents that are associated with long-term outcome. PMID- 10513776 TI - Assessment of left internal mammary artery graft patency and flow reserve after minimally invasive direct coronary artery bypass. AB - Despite its merits, minimally invasive direct coronary artery bypass surgery (MIDCAB) has been criticized for variable left internal mammary artery (LIMA) graft patency rates, prompting the frequent use of postoperative LIMA angiography. Noninvasive transthoracic Doppler interrogation of LIMA grafts has recently been shown to have utility for assessing patency and flow reserve after conventional bypass surgery, but data after MIDCAB has been limited. The objective of this study was to assess LIMA graft anatomy and physiology in 54 patients after MIDCAB using angiography and noninvasive LIMA Doppler imaging. The right internal mammary artery (RIMA) was studied as a control. LIMA flow reserve in response to adenosine was evaluated in a subgroup of 18 randomly chosen patients with patent grafts. LIMA angiographic patency was 93%. Forty-four patients (81%) had obtainable LIMA Doppler data. Patent grafts had a diastolic dominant flow pattern with a peak diastolic/systolic velocity ratio of 1.3 +/- 0.6 and a percent diastolic time-velocity integral (TVI) of 70 +/- 11%. These data were significantly different than the RIMA control values of 0.2 +/- 0.1 and 30 +/- 10%, respectively (p <0.05). Occluded grafts had absent flow or a systolic dominant pattern. Adenosine-induced increases in LIMA peak diastolic velocity from 48 +/- 20 to 105 +/-28 cm/s (p <0.05 vs baseline) and diastolic TVI from 21 +/- 10 to 37 +/- 19 cm (p <0.05 vs baseline), yielding adenosine/baseline ratios of 2.4 +/- 0.9 and 2.0 +/- 0.7, respectively, which was consistent with normal flow reserve. The diastolic flow velocity reserve response was inversely related to baseline diastolic flow (r = -0.69). In conclusion, MIDCAB can be associated with a high rate of LIMA potency and favorable physiologic Doppler flow patterns. Correlation of these findings to long-term patient outcome after MIDCAB is warranted. PMID- 10513777 TI - Alcohol consumption, coronary calcium, and coronary heart disease events. AB - This study was performed to determine if alcohol intake was associated with reduced coronary risk in a high-risk asymptomatic population, and whether this effect was independent of coronary risk factors and coronary calcium. In 1,196 asymptomatic subjects with coronary risk factors, we assessed alcohol consumption history, performed risk factor measurements, and quantified coronary calcium with electron beam computed tomography. These subjects were then followed for a mean of 41 months, and coronary events (myocardial infarction or coronary death) were noted. Significant inverse predictors of coronary events included alcohol use and serum high-density lipoprotein cholesterol level. Direct predictors of events were history of systemic hypertension, smoking, diabetes mellitus, serum cholesterol, and coronary calcium score. Subjects with coronary calcium were 3.1 times more likely to suffer a coronary event than those without calcium (95% confidence interval [CI] limits 1.3 to 7.2). Subjects who drank alcohol had a relative risk of 0.3 (95% CI limits 0.2 to 0.6) for developing coronary events. After controlling for age, gender, and other risk factors with logistic regression, these differences in relative risk persisted (relative risk 0.58; 95% CI limits 0.41 to 0.82). Alcohol consumption is a significant inverse predictor of coronary events, comparable in magnitude to standard risk factors and to radiographically measured coronary calcium. This effect is independent of coronary risk factors and coronary calcium. PMID- 10513778 TI - Effect of dipyridamole on QT dispersion in vasospastic angina pectoris. AB - Life-threatening ventricular arrhythmias have frequently been documented in patients with vasospastic angina. Moreover, the incidence of ventricular arrhythmias has been closely associated with increased QT dispersion. However, the underlying mechanism responsible for this arrhythmogenesis has not been clarified. The effects of dipyridamole and subsequent aminophylline administration on QT dispersion were examined in 35 patients with vasospastic angina and 30 patients with atypical chest pain. None of the patients enrolled in this study revealed any significant stenosis in coronary angiography. QT dispersion during dipyridamole followed by aminophylline administration was compared between the 2 groups. The baseline QT dispersion was similar in both groups (vasospastic angina: 27 +/- 8 ms; atypical chest pain: 28 +/- 7 ms). No significant changes in QT dispersion were observed in patients with atypical chest pain by dipyridamole (23 +/- 9 ms) and subsequent aminophylline administration (23 +/- 5 ms). However, the QT dispersion in patients with vasospastic angina increased significantly by dipyridamole administration (53 +/- 14 ms, p <0.0001) and returned to baseline by subsequent aminophylline administration (26 +/- 10 ms). Our data suggest that the disparity of ventricular repolarization in vasospastic angina may be mediated by increased endogenous adenosine. PMID- 10513779 TI - Concomitant use of cytochrome P450 3A4 inhibitors and simvastatin. AB - The long-term safety profile of simvastatin, established over 10 years of clinical use, is excellent. The principal adverse effect of all inhibitors of hydroxymethylglutarate co-enzyme A (HMG-CoA) reductase, myopathy, is infrequent. Simvastatin is a substrate for cytochrome P450 3A4 (CYP3A4). CYP3A4 inhibitors can elevate the plasma concentration of HMG-CoA reductase inhibitory activity derived from simvastatin. Clinical experience has shown that concomitant use of potent inhibitors of CYP3A4 increase the risk for myopathy. Evaluation of data from clinical trials and postmarketing surveillance allows assessment of whether concomitant use of weaker CYP3A4 inhibitors, as represented by calcium channel blockers, has any effect on the risk of myopathy. Cases of myopathy in long-term clinical megatrials and in analyses of postmarketing adverse event reports have been surveyed. In megatrials with simvastatin, the overall incidence of myopathy was 0.025%. The proportion of patients developing myopathy who were taking a calcium channel blocker with simvastatin (1 of 3) was similar to the proportion of patients taking a calcium channel blocker overall. Among marketed-use adverse event reports, concomitant medication with a potent CYP3A4 inhibitor was more frequent among reports of myopathy than among reports of nonmusculoskeletal adverse events. No excess use of calcium channel blockers among myopathy reports was observed. We conclude that the overall risk of myopathy during treatment with simvastatin is very low. Potent CYP3A4 inhibitors, especially cyclosporine, significantly increase the risk. There is no evidence that weaker CYP3A4 inhibitors such as calcium channel blockers increase the risk. PMID- 10513780 TI - Effect of batroxobin on spontaneous echo contrast and hemorheology in left atrial appendage in atrial fibrillation assessed by transesophageal echocardiograpy. AB - Controversy exists regarding the effect of defibrination on spontaneous echo contrast and flow dynamics in left atrial appendage (LAA) in atrial fibrillation. We aimed to investigate the effect of batroxobin, which decreases plasma fibrinogen level, on the echo intensity of spontaneous echo contrast in LAA. In 36 patients with atrial fibrillation (duration 7 +/- 4 years), transesophageal echocardiography was performed at baseline and 24 hours after batroxobin administration (0.2 U/kg). At the orifice of the LAA, integrated backscatter of echo contrast and peak velocity of LAA emptying flow were measured. Plasma fibrinogen and whole blood viscosity were also measured. Fibrinogen and viscosity were significantly lower after batroxobin administration (96 +/-38 mg/dl and 4.35 +/- 0.56 cp) than those at baseline (320 +/- 61 mg/dl and 4.71 +/- 0.61 cp, both p <0.001). A significant positive correlation between changes in plasma fibrinogen and whole blood viscosity (r = 0.49, p = 0.002) was shown. The integrated backscatter significantly decreased from 14 +/- 3 to 12 +/- 3 decibels after batroxobin (p <0.001), and the changes in integrated backscatter and plasma fibrinogen was significantly correlated. Therefore, batroxobin administration improved blood rheology and decreased blood cell aggregation, which are effective in preventing left atrial thrombus formation. PMID- 10513781 TI - Outcome of pregnancy in patients with congenitally corrected transposition of the great arteries. AB - To assess maternal and fetal outcome of pregnancy in patients with congenitally corrected transposition of the great arteries, we reviewed 19 patients (18 retrospectively) who had 45 pregnancies. Their ages ranged from 18 to 40 years (mean 27) at the time of pregnancy. Thirty-six percent of the pregnancies were undertaken while patients were cyanosed, 7% in patients with unpaced complete heart block, and 16% were undertaken after surgical repair of the associated anomalies. Change in functional class and maternal cardiovascular complications during pregnancy were analyzed as well as number of live births, miscarriages, elective termination of pregnancies, timing of delivery, and incidence of cardiac defects in the live offspring. Five patients (26% of patients) developed cardiovascular complications during pregnancy, namely congestive heart failure (3 patients), worsening cyanosis (1 patient), and cerebrovascular accident (1 patient). No maternal deaths occurred. There were 27 live births (60%), 12 miscarriages (27%), and 6 elective terminations of pregnancy (13%). Cyanosis was a significant risk factor for miscarriage. One live offspring had congenital heart disease. Close follow-up of these patients during pregnancy, by a team of experienced physicians, is recommended. PMID- 10513782 TI - Lack of association between seropositivity to Chlamydia pneumoniae and carotid atherosclerosis. AB - Since the Chlamydia pneumoniae (C. pneumoniae)-specific antibody was shown to be associated with acute myocardial infarction and chronic coronary heart disease, the role of C. pneumoniae in the etiology of cardiovascular disease has been studied by a number of groups. We investigated the association between the C. pneumoniae-specific antibody, measured by microimmunofluorescence, risk factors for cardiovascular disease, and atherosclerosis in a randomly selected urban population. Overall, immunoglobulin-G (IgG) seroprevalence to C. pneumoniae in this sample of 1,034 subjects was 58%, whereas IgA seroprevalence was 32%. There was a decline in seropositivity with age for IgG but not IgA. Men were more likely than women to be IgG (66% vs 51%, chi-square p = 0.001) and IgA seropositive (36% vs 28%, chi-square p = 0.005). Current smokers had higher IgA seropositivity than nonsmokers (43% vs 30%). Those patients with a family history of cerebrovascular disease were more likely to have IgG antibody than those without (75% vs 57%, chi-square p= 0.007). Neither IgG nor IgA seropositivity was associated with the standard risk factors of hypertension, hyperlipidemia, or family history of ischemic heart disease, nor was seropositivity associated with carotid intima medial thickening (IMT) or atherosclerotic plaque as measured by carotid B-mode ultrasound. There was no difference between those participants who were IgG or IgA seropositive and seronegative in measurements of mean IMT, prevalence of abnormal IMT, and percentage with atherosclerotic plaque. In conclusion, although C. pneumoniae was associated with several risk factors for cardiovascular disease in a large cross-sectional population, we found no independent association between seroprevalence to C. pneumoniae and carotid atherosclerosis as measured by carotid IMT. PMID- 10513783 TI - Best method in clinical practice and in research studies to determine left atrial size. AB - Although the anteroposterior dimension of the left atrium is universally used in clinical practice and research, we hypothesized that it may be an inaccurate surrogate for volume because its use is based on the unlikely assumption that there is a constant relation among atrial dimensions. The following measurements of the left atrium were made at end ventricular systole: (1) M-mode-derived anteroposterior linear dimension from the parasternal long-axis view; (2) digitized planimetry of the left atrial (LA) cavity from the apical 4-chamber view; and (3) digitized planimetry of the LA cavity from the apical 2-chamber view. The following volume calculations were obtained from these digital measurements: (1) volume derived from the M-mode dimension assuming a spherical shape; (2) volume derived from the single plane area-length of apical 4-chamber view, which assumes that LA geometry can be generalized from a single 2 dimensional plane; and (3) volume derived from the biplane method of discs. The correlation coefficient between the M-mode and biplane methods of determining LA volume was r = 0.76. The mean difference (+/-2 SDs) between these methods is -25 +/- 33 ml. The correlation coefficient between the single plane apical 4-chamber and biplane methods of determining LA volume is r = 0.97. The mean difference (+/ 2 SDs) between these methods was -5.0 +/- 12 ml, indicating good agreement. The M mode measure of the left atrium is an inaccurate representation of its size. Two dimensional-derived LA volumes provide a more accurate measure of the true size of the left atrium and are more sensitive to changes in LA size. When an echocardiographic measure of LA size is made either in an individual patient or as a variable in a research study, the M-mode measure should be avoided. PMID- 10513784 TI - Bertram Pitt, MD: a conversation with the editor. Interview by William Clifford Roberts. PMID- 10513785 TI - Frequency of left atrial thrombus and spontaneous echocardiographic contrast in acute myocardial infarction. AB - Left ventricular systolic dysfunction may precipitate blood stasis as well as thrombus formation in the left atrial appendage of patients with acute myocardial infarction, even in the presence of sinus rhythm. Thus, left atrial thrombi may be an alternative source for systemic embolism in acute myocardial infarction. PMID- 10513786 TI - Intravascular ultrasound volumetric assessment of intimal hyperplasia in stents treated with intracoronary radiation. AB - Iridium-192 (gamma)-radiation is effective in preventing recurrent in-stent restenosis by reducing neointimal hyperplasia as illustrated by intravascular ultrasound study and plaque area-length plot. This analytic technique will further our understanding of vessel behavior to radiant energy source both inside and outside the stented coronary artery segments. PMID- 10513787 TI - Atenolol versus amlodipine versus isosorbide-5-mononitrate on anginal symptoms in syndrome X. AB - The effects of a beta blocker (atenolol), a calcium antagonist (amlodipine), and a nitrate (isosorbide-5-mononitrate) on anginal symptoms in 10 patients with syndrome X were assessed in a crossover, double-blind, randomized trial. Only atenolol was found to significantly improve chest pain episodes, suggesting that it should be the preferred drug when starting pharmacologic treatment of patients with syndrome X. PMID- 10513788 TI - Influence of warfarin therapy on left atrial spontaneous echo contrast in nonvalvular atrial fibrillation. AB - To examine whether warfarin therapy had any influence on left atrial spontaneous echo contrast, we performed serial transesophageal echocardiography with integrated backscatter analysis in 12 patients with non-valvular atrial fibrillation. We found that the integrated backscatter intensity of the left atrial cavity did not change after 1 to 2 months of warfarin therapy, and concluded that this therapy does not influence spontaneous echo contrast. PMID- 10513789 TI - Use of telemonitoring to decrease the rate of hospitalization in patients with severe congestive heart failure. AB - We investigated whether the application of a home-based telemonitoring system could decrease hospital admissions and emergency room visits in 30 patients with severe congestive heart failure compared with 51 patients with congestive heart failure who received standard therapy. Emergency room visits (1 vs 11) and hospitalizations (13 vs 36) were decreased in the telemonitored group (both p <0.05). PMID- 10513790 TI - Prothrombotic state and elevated levels of plasminogen activator inhibitor-1 in mitral stenosis with and without atrial fibrillation. AB - Patients with mitral stenosis in sinus rhythm are in a prothrombotic state and have fibrinolytic dysfunction, shown by an increase in levels of the inhibitor of tissue plasminogen activator, D-dimer, and modified antithrombin III. This state may be observed even in patients without dilated left atria (diameter < or =45 mm). PMID- 10513791 TI - Guidelines for referral and management of systemic lupus erythematosus in adults. American College of Rheumatology Ad Hoc Committee on Systemic Lupus Erythematosus Guidelines. AB - SLE is a complex disorder with variable presentations, course, and prognosis. Since its prevalence is only 1/1,000, most primary care physicians and general internists will not have sufficient experience in the management of moderate-to severe life-threatening disease. The major tasks of the primary care physician in the diagnosis and management of patients with SLE include early diagnosis, appropriate referral, monitoring patients with mild, stable disease, and collaboration with a specialist in the management of severe disease. Guidelines for the initial evaluation, reasons for referral, and management of mild and severe SLE are provided. PMID- 10513792 TI - Listening to the patient: a practical guide to self-report questionnaires in clinical care. PMID- 10513793 TI - Lyme arthritis: lessons learned and to be learned. PMID- 10513794 TI - Association of antibiotic treatment-resistant Lyme arthritis with T cell responses to dominant epitopes of outer surface protein A of Borrelia burgdorferi. AB - OBJECTIVE: To explore further the association of antibiotic treatment-resistant Lyme arthritis and T cell reactivity with outer surface protein A (OspA) of Borrelia burgdorferi, including the identification of T cell epitopes associated with this treatment-resistant course. METHODS: The responses of peripheral blood and, if available, synovial fluid lymphocytes to B burgdorferi proteins, fragments, and synthetic peptides, as determined by proliferation assay and interferon-gamma production, were compared in 16 patients with treatment responsive and 16 with treatment-resistant Lyme arthritis. RESULTS: The maximum severity of joint swelling correlated directly with the response to OspA. Moreover, the only significant difference between patients with treatment resistant and treatment-responsive arthritis was in reactivity with N-terminal and C-terminal fragments of OspA, OspA1 (amino acids [aa] 16-106), and OspA3 (aa 168-273). Epitope mapping showed that 14 of the 16 patients with treatment resistant arthritis had responses to OspA peptides (usually 4 or 5 epitopes), whereas only 5 of the 16 patients with treatment-responsive arthritis had reactivity with these peptides (usually 1 or 2 epitopes) (P = 0.003). Patients with HLA-DRB1 alleles associated with treatment-resistant arthritis were more likely to react with peptide 15 (aa 154-173) and, to a lesser degree, with peptide 21 (aa 214-233) than patients with other alleles, whereas the responses to other epitopes were similar in both groups. CONCLUSION: The maximum severity of joint swelling and the duration of Lyme arthritis after antibiotic treatment are associated with T cell responses to specific epitopes of OspA. PMID- 10513795 TI - Fcgamma receptor polymorphisms in Wegener's granulomatosis: risk factors for disease relapse. AB - OBJECTIVE: Several studies have recently identified polymorphisms of receptors for the Fc fragment of IgG (FcgammaR) as genetic factors influencing susceptibility to multiple autoimmune and infectious diseases. This genetic predisposition could also influence the expression of Wegener's granulomatosis (WG), a systemic autoimmune disease with chronic nasal carriage of Staphylococcus aureus as an important risk factor for disease relapses. Therefore, we analyzed 3 functional FcgammaR polymorphisms from 91 patients with WG and 154 controls for a possible relationship with disease expression and occurrence of relapses. METHODS: FcgammaR phenotypes were determined using amplification of FcgammaR genomic regions in allotype-specific polymerase chain reactions. Of particular interest in the analysis were 2 allotypic forms of FcgammaRIIa (R131 or H131) and 2 allotypic forms of FcgammaRIIIa (V158 or F158), all of which are functionally different. RESULTS: Analysis of FcgammaR phenotypes demonstrated that patients with WG were more prone to disease relapse in the first 5 years after diagnosis if they were homozygous for both the R131 form of FcgammaRIIa and the F158 form of FcgammaRIIIa (relative risk 3.3, 95% confidence interval 1.6-6.8). These polymorphisms are both associated with decreased FcR-mediated clearance, which may be relevant to the chronic nasal carriage of S aureus. CONCLUSION: Both the R/H131 polymorphism of FcgammaRIIa and the V/F158 polymorphism of FcgammaRIIIa represent heritable risk factors for the development of disease relapses in WG. PMID- 10513796 TI - Overrepresentation of the Fcgamma receptor type IIA R131/R131 genotype in caucasoid systemic lupus erythematosus patients with autoantibodies to C1q and glomerulonephritis. AB - OBJECTIVE: To test the hypothesis that there is an association between the Fcgamma receptor type IIA (FcgammaRIIA)-H/R131 polymorphism and autoantibodies to the collagenous region (CLR) of C1q in patients with systemic lupus erythematosus (SLE). METHODS: One hundred ninety-five Caucasoid lupus patients were studied. Anti-C1q(CLR) antibodies in serum were measured by enzyme-linked immunosorbent assay (ELISA) and FcgammaRIIA genotype analysis was performed by polymerase chain reaction. Immunoglobulin subclass of the autoantibodies was measured by ELISA. RESULTS: Fifty-six patients were anti-C1q antibody positive, and Ig subclass analysis indicated a predominance of IgG2 anti-C1q antibodies. Analysis of the SLE population as a whole revealed no significant difference in the allele frequencies of R131 and H131 compared with controls. There was, however, a significantly increased frequency of the R131 allele both in the anti-C1q positive subgroup of patients (chi2 = 7.66, P<0.01) and in the 71 patients with nephritis (chi2 = 7.76, P< 0.01), compared with controls. CONCLUSION: FcgammaRIIA R131 constitutes a heritable susceptibility factor for the development of SLE with manifestations in the kidney in Caucasoid patients. The close associations demonstrated between this FcgammaRII variant, antibodies to C1q(CLR), and glomerulonephritis may be due to a failure of clearance of the potentially pathogenic IgG2 autoantibody. PMID- 10513797 TI - The spectrum of apoptotic defects and clinical manifestations, including systemic lupus erythematosus, in humans with CD95 (Fas/APO-1) mutations. AB - OBJECTIVE: To determine the clinical spectrum of disease in humans with mutations in the CD95 (Fas/ APO-1) receptor and to obtain mechanistic insight into the different clinical phenotypes observed. METHODS: Clinical information for each of the index cases, first-degree relatives, and any family members reported to have Canale-Smith syndrome (or another autoimmune disease) was gathered by direct interview, chart review, and verification of data by the physician or pathologist concerned. Apoptosis of activated T or B lymphocytes was induced by agonistic anti-CD95 antibodies and quantified by a cell death assay (propidium iodide staining in the subdiploid peak) or cell viability assay (alamar blue or 3H thymidine incorporation). RESULTS: Evaluation of an additional 8 probands with novel heterozygous CD95 mutations revealed hypergammaglobulinemia and immune mediated cytopenias in all patients, as well as urticarial rash, oral ulceration, lymphopenia, and peripheral neuropathy in some individuals. One patient (P4) had systemic lupus erythematosus (SLE) characterized by a World Health Organization class V lupus nephropathy, a recurrent, reversible multifocal central nervous system disorder, high-titer antiphospholipid autoantibodies, and autoimmune cytopenias. In the P4 pedigree, the father had reduced T and B cell apoptosis associated with a CD95 mutation, whereas an independent B cell apoptotic defect was demonstrated in maternal family members who did not have a CD95 mutation. Three cases of B cell lymphoma occurred in carriers of the CD95 mutation. CONCLUSIONS: CD95 mutations are associated with loss of regulation of B lymphocytes, which predisposes to systemic autoimmunity including SLE. The P4 family provides a model of the complex genetic and functional interactions that are required for the development of a lupus-like syndrome. PMID- 10513798 TI - Age-specific effects of juvenile rheumatoid arthritis-associated HLA alleles. AB - OBJECTIVE: To define the onset and duration of effect of the HLA alleles that are associated with disease susceptibility and protection in juvenile rheumatoid arthritis (JRA) and 2 of its subtypes. METHODS: We typed 680 patients with JRA and 254 ethnically matched unrelated controls for HLA class I and II genes. The frequency of each allele was calculated for each of the age-at-onset, onset type, and sex categories and plotted against the allele frequency in the control population. Survival analysis (with onset of disease as the terminating event) was used to calculate the age by which 50% (St0.5) and 80% (St0.2) of the children with particular alleles and combinations of alleles develop disease. This allele-specific survival analysis also allowed for the comparison of the overall survival functions for the various JRA subtype and sex categories. RESULTS: Certain alleles are strongly associated with early susceptibility to pauciarticular JRA, including HLA-A2, DR8, DR5, and DPB1*0201. Fifty percent of the children carrying at least 1 of these alleles had disease onset prior to their third birthday. Among children who carried HLA-A2 and any 2 HLA-DR alleles (DR3, DR5, DR6, or DR8), the median age at the onset of pauciarticular disease was 2.7 years. Combinations of A2 and DPB1*0201 and one DR allele narrowed the window further to a median age at onset of 2.4 years. B27 and DR4 were associated with protection early in life but with increased risk later in childhood, with St0.5 values of 7.3 and 6.6 years, respectively, for pauciarticular JRA and St0.5 values of 10.2 and 10.7 years, respectively, for polyarticular JRA. Sex strongly influenced the age at which many of the alleles have their effect. CONCLUSION: These data define at what age and for how long various HLA alleles influence susceptibility and protection (window-of-effect) in patients with JRA. In addition, these data establish more clearly the boundaries of ages-at-onset for 2 of the subtypes of the disease. PMID- 10513799 TI - Long-term course and outcome of functional capacity in rheumatoid arthritis: the effect of disease activity and radiologic damage over time. AB - OBJECTIVE: To investigate the evolution of functional capacity, disease activity, and joint destruction over time in a 12-year prospective cohort of rheumatoid arthritis (RA) patients, and to study the relative contribution of disease activity and joint destruction to the loss of functional capacity. METHODS: One hundred thirty-two female patients with recent-onset RA were assessed at 0, 3, 6, and 12 years of followup for functional capacity (Health Assessment Questionnaire [HAQ] score), disease activity (Disease Activity Score [DAS]), and joint destruction (Sharp score of radiologic damage). RESULTS: The Sharp score deteriorated steadily over time, while the HAQ score and DAS showed a variable course. The DAS correlated strongly with the HAQ score throughout the disease course. The correlation between the Sharp score and the HAQ score was weak at study start, but became strong after 12 years. After 12 years of followup, disease activity was the main determinant of the HAQ score when entered in a multivariate analysis. CONCLUSION: Functional capacity is strongly influenced by disease activity throughout the course of RA. Even in longstanding RA, disease activity proves to be the main determinant of the HAQ score for functional capacity. PMID- 10513800 TI - Serum levels of hyaluronan, antigenic keratan sulfate, matrix metalloproteinase 3, and tissue inhibitor of metalloproteinases 1 change predictably in rheumatoid arthritis patients who have begun activity after a night of bed rest. AB - OBJECTIVE: To evaluate whether and how moderate physical activity following a night of rest influences serum levels of matrix metalloproteinase 3 (MMP-3), tissue inhibitor of metalloproteinases 1 (TIMP-1), antigenic keratan sulfate (Ag KS), and hyaluronan (HA) in 10 normal subjects and 38 patients with rheumatoid arthritis (RA). METHODS: Blood was obtained from 20 RA patients before they arose from a night's sleep, and again 1 and 4 hours after they had begun to perform moderate physical activity. Another 18 RA patients remained in bed and blood was sampled at the same time periods. Serum levels of MMP-3, TIMP-1, Ag KS, and HA were measured by enzyme-linked immunosorbent assay. Clinical activity was evaluated by the Lansbury index. RESULTS: Both in normal subjects and in RA patients who did not remain in bed throughout the period of blood sampling, levels of HA, Ag KS, and MMP-3 increased significantly during the first hour after the subjects arose: the increase in HA and Ag KS correlated with the Lansbury index in the RA group. Three hours later, levels of Ag KS had dropped to baseline values in both groups of subjects. Levels of HA remained significantly and moderately elevated in the RA group but not in the control group, while levels of MMP-3 did not drop significantly in either group. In contrast, levels of HA, Ag KS, and MMP-3 did not change significantly in RA patients who had remained in bed. Unlike the other markers, the levels of TIMP-1 remained unchanged at the different time periods in all 3 groups studied. CONCLUSION: Significant changes in serum levels of some metabolic markers occur during the first hour after one arises from a night of sleep, especially in patients with RA. Measurement of the magnitude of these changes at different times in individual patients provides very different information about metabolic changes occurring in joint tissue than does measurement of the level of the markers at a single time point, as is usually currently reported. PMID- 10513801 TI - Function and health-related quality of life: results from a randomized controlled trial of leflunomide versus methotrexate or placebo in patients with active rheumatoid arthritis. Leflunomide Rheumatoid Arthritis Investigators Group. AB - OBJECTIVE: To assess the efficacy of leflunomide or methotrexate compared with placebo in improving function and health-related quality of life in patients with active rheumatoid arthritis (RA), and to examine correlations between response status (as defined by the American College of Rheumatology [ACR] response criteria) and improvement in these measures. METHODS: This 52-week, multicenter, doubleblind, controlled trial compared responses to the Health Assessment Questionnaire (HAQ), modified Health Assessment Questionnaire (MHAQ), Problem Elicitation Technique (PET), Medical Outcomes Study Short Form 36 (SF-36), and questions regarding work productivity among 3 treatment groups (leflunomide, methotrexate, and placebo). Improvement in the PET top 5 and SF-36 scales and component scores were compared with ACR response rates. RESULTS: Clinically meaningful and statistically significant (P<0.0001) improvement in measures of function and heath-related quality of life (MHAQ scores, all scales and disability index of the HAQ, weighted top 5 score of the PET, 5 of 8 scales and physical component score of the SF-36, and work productivity) was seen during treatment with leflunomide in comparison with placebo. Methotrexate administration resulted in significant improvements (P<0.05) in comparison with placebo in the MHAQ scores, HAQ disability index, weighted top 5 score of the PET, physical component score of the SF-36, and bodily pain scale. Compared with methotrexate, leflunomide administration resulted in significantly (P<0.01) more improvement in the MHAQ scores, 5 of 8 scales and disability index of the HAQ, weighted top 5 score of the PET, and 2 of 8 scales and physical component score of the SF-36. Improvements in the PET score, SF-36 physical component score, and work productivity correlated with the ACR responder rates of > or =20% and > or =50% improvement. CONCLUSION: Significant improvements in function and health related quality of life occurred in patients with RA during treatment with leflunomide or methotrexate. These findings were clinically meaningful and correlated with the ACR response status. PMID- 10513802 TI - Global statistical tests for comparing multiple outcomes in rheumatoid arthritis trials. MIRA Trial Group. AB - OBJECTIVE: To evaluate global statistical tests (GSTs) of treatment effectiveness for rheumatoid arthritis (RA) trials measuring multiple outcomes. METHODS: Using outcome measures from American College of Rheumatology (ACR) core set variables available in 3 RA trials, GSTs were calculated using the O'Brien ranking procedure and a procedure for binary data. GSTs take correlations among outcomes into account. Power calculations using 1 trial data set provide comparisons of GSTs and ACR criteria for improvement. RESULTS: Spearman correlations among outcomes ranged from 0.21 to 0.73. Erythrocyte sedimentation rate had the lowest correlation with other outcomes in all 3 trials. Within a trial, joint swelling and joint tenderness or patient and physician assessment had the highest correlations, depending on the trial. Results were consistent with results using the ACR criteria, although the GST was more powerful. CONCLUSION: GSTs are a useful tool for comparing treatment effects across multiple clinically meaningful outcome measures. The GST allows easy inclusion of validated, reliable new measures that are not a part of ACR criteria, such as quality of life, and can be computed with or without selecting a cutoff point defining patient improvement. GSTs should be considered for rheumatic disease treatment trials. PMID- 10513803 TI - Lower prevalence of Chlamydia pneumoniae DNA compared with Chlamydia trachomatis DNA in synovial tissue of arthritis patients. AB - OBJECTIVE: To assess the presence of Chlamydia pneumoniae DNA in the joints of patients with reactive arthritis (ReA) and other arthritides. METHODS: DNA was prepared from synovial tissue (ST) and several synovial fluid (SF) samples from 188 patients with either ReA, undifferentiated oligoarthritis, or other forms of arthritis, and from 24 normal (non-arthritis) individuals. Preparations were screened using polymerase chain reaction (PCR) assays that independently targeted the C. pneumoniae 16S ribosomal RNA and major outer membrane protein genes. RESULTS: Twenty-seven of 212 ST samples (12.7%) were PCR positive for C. pneumoniae DNA; 10 SF samples from these 27 patients were similarly positive. Among the PCR-positive patients, 3 had ReA, 2 had Reiter's syndrome, 7 had undifferentiated oligoarthritis, 4 had undifferentiated monarthritis, 6 had rheumatoid arthritis, and 5 had other forms of arthritis. No samples from normal control individuals were PCR positive. CONCLUSION: DNA of C pneumoniae is present in synovial specimens from some arthritis patients. The prevalence of this organism in the joints was lower than that of C trachomatis, and synovial presence of the organism was not associated with any distinct clinical syndrome. Widely disseminated nucleic acids such as those of C. pneumoniae might have some role in the pathogenesis of several arthritides, since the organism was not found in the ST from normal control individuals. PMID- 10513804 TI - Ciprofloxacin treatment does not influence course or relapse rate of reactive arthritis and anterior uveitis. AB - OBJECTIVE: To assess the efficacy of ciprofloxacin in the treatment of reactive arthritis (ReA) and anterior uveitis (AU) in a double-blind, randomized, placebo controlled trial. METHODS: Seventy-two patients participated in this study, 56 with ReA and 42 with AU (26 patients had both ReA and AU). Ciprofloxacin (750 mg twice a day) was administered for 12 months with a 12-month followup. End points of the study included time to disease relapse and measures of disease severity. RESULTS: There was no difference between groups in time to disease relapse, joint inflammation, number of joints and enthesis involved in patients with ReA, or signs and symptoms of AU. CONCLUSION: Long-term treatment of ReA and AU with ciprofloxacin made no statistically significant difference to the natural history of these diseases or their severity. PMID- 10513805 TI - Hepatitis C virus but not GB virus C/hepatitis G virus has a role in type II cryoglobulinemia. AB - OBJECTIVE: Hepatitis C virus (HCV) infection is associated with type II cryoglobulinemia. HCV is specifically concentrated in type II cryoglobulins and has been implicated in the cutaneous vasculitis associated with the disease. In contrast to HCV, a role for hepatitis G virus (HGV) in type II cryoglobulinemia has not been defined, although prevalences as high as 43% of HGV infections in type II cryoglobulinemia have also been reported. METHODS: We studied 34 patients with type II and 29 patients with type III cryoglobulinemia associated with HCV infection, 6 patients with essential mixed cryoglobulinemia (EMC; all with type II), 50 hospital control patients, and 125 normal individuals. Serum HCV and HGV RNA were detected by reverse transcription-polymerase chain reaction (RT-PCR). In coinfected sera, HCV and HGV were quantitated by competitive RT-PCR assays. One coinfected patient was studied longitudinally for 6 years. RESULTS: Two (5.9%) of 34 patients with HCV-infected type II cryoglobulinemia, none of 29 patients with type III cryoglobulinemia, and none of 6 patients with EMC were positive for HGV RNA, for an overall prevalence of 3.0% in mixed cryoglobulinemia. None of the control populations were positive for HGV. No statistical difference was seen between the prevalence in patients with type II cryoglobulinemia and the other populations studied. In coinfected sera, HCV, but not HGV, was concentrated in cryoglobulins, and HCV, but not HGV, correlated with cryoglobulinemia in a longitudinal study. CONCLUSION: There is a low prevalence of coinfection with HGV in patients with mixed cryoglobulinemia and HCV infection in the United States. HCV is selectively precipitated by type II cryoglobulins in coinfected sera. HGV infection does not appear to have a role in mixed cryoglobulinemia. PMID- 10513806 TI - Confirmation of genetic linkage between human systemic lupus erythematosus and chromosome 1q41. AB - OBJECTIVE: Genetic susceptibility to systemic lupus erythematosus (SLE) is undoubtedly complex and, presumably, involves multiple loci. Linkage of SLE to D1S229 at chromosome 1q41 has been previously reported in a cohort of 52 affected sibpairs. The present study sought to confirm this reported linkage in an independent cohort of 127 extended multiplex SLE pedigrees containing 107 affected sibpairs. METHODS: Genotype data were collected for D1S229 and 18 flanking microsatellite markers spanning chromosome 1q32-1q42. Analyses of genotype data included a model-based logarithm of odds (LOD) score approach, affected sibpair analyses, and transmission disequilibrium tests. RESULTS: A maximum LOD score of 1.46 was found with D1S229 in a subgroup of 78 European American pedigrees, with additional support from multiple markers clustered around D1S229. Increased allele sharing in affected siblings was most significant at D1S2616, particularly in European Americans (P = 0.0005), followed by D1S229 (P = 0.002), D1S490 (P = 0.028), and D1S1605 (P = 0.037). Although linkage in a subgroup of 40 African American pedigrees was not suggested by the analyses of any marker tested in the chromosomal region surrounding D1S229, a maximum LOD score of 3.03 was found with D1S3462, mapped 15 centimorgans distal to D1S229. CONCLUSION: Our linkage analysis results in European Americans at D1S229 are remarkably similar to those previously reported. That at least 1 genetic effect near this locus is important for susceptibility to lupus should now be generally accepted, and efforts to identify the gene are thereby justified. PMID- 10513807 TI - Diminished levels of T cell receptor zeta chains in peripheral blood T lymphocytes from patients with systemic lupus erythematosus. AB - OBJECTIVE: To examine the expression of molecules known to participate in early T cell receptor (TCR)/CD3 signaling in peripheral blood (PB) T lymphocytes from patients with systemic lupus erythematosus (SLE). METHODS: Signaling molecules were analyzed by immunoprecipitation and Western blotting of unstimulated PB T lymphocyte cell lysates from SLE patients, non-SLE disease controls, and healthy controls. Flow cytometry was used for analysis of the expression of membrane markers in intact cells. RESULTS: PB T lymphocytes from SLE patients showed diminished levels of TCRzeta chains. This was not due to trapping of these molecules in the cytoskeleton, nor was it dependent on the presence of monocyte/macrophages. There was normal expression of CD3epsilon chains and normal assembly of TCR/CD3 complexes in membranes. We observed a lack of expression of TCRzeta chains in in vitro cultures of SLE T cells, and reversal of the defective expression in some patients by culturing T cells in the presence of NH4Cl. CONCLUSION: Blood lymphocytes from SLE patients have a diminished expression of TCRzeta chains that may be related to enhanced degradation in the lysosomal compartment. The defective expression of these molecules may alter signal transduction via the CD3 pathway and contribute to abnormal T cell responses in T lymphocytes from SLE patients. PMID- 10513808 TI - Differential regulation of rheumatoid synovial cell interleukin-12 production by tumor necrosis factor alpha and CD40 signals. AB - OBJECTIVE: To investigate the roles of tumor necrosis factor alpha(TNFalpha) and the CD40-CD154 interaction in interleukin-12 (IL-12) production by rheumatoid synovial cells (SC). METHODS: Levels of IL-12 (p40 and p70) in synovial tissue and culture supernatants of SC from patients with rheumatoid arthritis (RA), osteoarthritis (OA), and ankylosing spondylitis (AS) were assayed by enzyme linked immunosorbent assay. Effects of anti-CD154 and anti-TNFalpha antibody on spontaneous and lipopolysaccharide (LPS)-stimulated IL-12 production by SC were examined. Effects of immobilized anti-CD3 treatment and depletion of CD4+ T cells on IL-12 production were also tested. CD154 expression by synovial T cells and intracellular IL-12 production during culture were analyzed by flow cytometry. RESULTS: IL-12 p40 and p70 levels in RA synovial tissue and spontaneous IL-12 p40 production by SC from RA patients were significantly higher than the levels in OA and AS patients. Spontaneous IL-12 production by SC from RA patients significantly decreased after depletion of CD4+ T cells from SC or after application of anti-CD154 antibody, but not by treatment with anti-TNFalpha antibody. Anti-CD3 antibody stimulation increased spontaneous IL-12 p40 production and CD154 expression by synovial T cells. The increment of IL-12 p40 production by anti-CD3 was abrogated by anti-CD154 antibody. IL-12 p40 production was also increased by LPS stimulation. LPS-stimulated IL-12 production was inhibited by anti-TNFalpha antibody, but not by T cell depletion and anti-CD154 antibody treatment. The TNFalpha inhibitor rolipram inhibited LPS-stimulated IL 12 p40 production by RA SC more strongly than spontaneous production. TNFalpha restored LPS-stimulated IL-12 production that had been inhibited by rolipram. CONCLUSION: IL-12 production in RA is regulated by 2 different pathways. One pathway is T cell dependent, predominantly through a CD40-CD154 interaction, while the other is T cell independent, mediated through TNFalpha. Inhibition of IL-12 production by interference with CD40-CD154 interaction and TNFalpha production may be a potential therapeutic strategy for treating RA. PMID- 10513809 TI - Treatment with sulfasalazine or sulfapyridine, but not 5-aminosalicyclic acid, inhibits basic fibroblast growth factor-induced endothelial cell chemotaxis. AB - OBJECTIVE: Rheumatoid arthritis (RA) is characterized by leukocyte recruitment and angiogenesis. We investigated the effects of sulfasalazine (SSZ) and its metabolites, sulfapyridine (SP) and 5-aminosalicylic acid (5-ASA), on components of angiogenesis, namely, endothelial cell (EC) chemotaxis and proliferation, as well as on EC chemokine and soluble adhesion molecule expression. METHODS: SSZ, SP, and 5-ASA were assayed for their effects on basic fibroblast growth factor (bFGF)-induced human dermal microvascular endothelial cell (HMVEC) chemotaxis and proliferation. EC were plated on Matrigel to assess the effect of SSZ on EC tube formation. Enzyme-linked immunosorbent assays were performed to determine changes in HMVEC production of interleukin-8 (IL-8), monocyte chemoattractant protein-1 (MCP-1), growth-related oncogene alpha (GROalpha), epithelial neutrophil activating peptide 78 (ENA-78), soluble E-selectin (sE-selectin), and soluble intercellular adhesion molecule 1 (sICAM-1) upon treatment with SSZ or its metabolites. RESULTS: HMVEC incubated with SSZ or SP exhibited reduced bFGF induced chemotaxis (59%, [n = 7] and 22%, [n = 3], respectively) (P<0.05). SSZ and SP decreased basal HMVEC proliferation, while 5-ASA increased proliferation (P<0.05; [n = 5]). SSZ decreased bFGF-induced HMVEC proliferation (P<0.05 [n = 5]). SSZ inhibited phorbol 12-myristate 13-acetate-induced HMVEC tube formation (P<0.05; [minimum n = 5]). Tumor necrosis factor alpha-stimulated HMVEC shedding of sICAM-1 was reduced by incubation with either SSZ (19%) or 5-ASA (23%) (P<0.05; [n = 6]). SP inhibited cytokine-stimulated HMVEC expression of IL-8 and MCP-1 (P<0.05; [n = 4]). Neither SSZ nor its metabolites had any effect on HMVEC production of sE-selectin, GROalpha, or ENA-78. CONCLUSION: These results demonstrate that SSZ and its metabolite SP may affect the pathogenesis of RA by inhibiting EC chemotaxis, proliferation, tube formation, and expression of sICAM 1, IL-8, and MCP-1. PMID- 10513810 TI - Induction of alpha1-antitrypsin synthesis in human articular chondrocytes by interleukin-6-type cytokines: evidence for a local acute-phase response in the joint. AB - OBJECTIVE: We have previously shown that human articular chondrocytes synthesize large amounts of interleukin-6 (IL-6) upon stimulation with proinflammatory cytokines and that they express the IL-6 receptor. The present study was undertaken to analyze whether different IL-6-type cytokines can induce synthesis of the acute-phase protein alpha1-antitrypsin in human articular chondrocytes. METHODS: Chondrocytes from human articular cartilage, cultured in agarose, were stimulated with IL-6-type cytokines. Total RNA was isolated and analyzed by Northern blotting. Levels of alpha1-antitrypsin protein were determined by enzyme immunoassay. RESULTS: Stimulation of chondrocytes with oncostatin M (OSM) and IL 6 led to a 5-10-fold increase in alpha1-antitrypsin synthesis. This increase was dose and time dependent. Furthermore, OSM and IL-6 induced IL-6 synthesis in chondrocytes, resulting in an autocrine amplification loop. CONCLUSION: Our data strongly suggest the existence of a local acute-phase response in the joint. Synthesis of the acute-phase protein alpha1-antitrypsin, a major inhibitor of serine proteinases, may be an important protective mechanism of articular chondrocytes to prevent cartilage damage in inflammatory joint diseases. PMID- 10513811 TI - Up-regulation of MDC15 (metargidin) messenger RNA in human osteoarthritic cartilage. AB - OBJECTIVE: The aim of the study was to investigate the messenger RNA (mRNA) expression of the disintegrin metalloproteinase MDC15 (metargidin, or ADAM-15) in normal and osteoarthritic (OA) articular cartilage. METHODS: In situ hybridization experiments and reverse transcription-polymerase chain reaction (RT PCR) were performed on tissue samples of adult normal and OA articular cartilage. RESULTS: MDC15 mRNA could be detected in normal articular cartilage by RT-PCR using tissue-extracted total RNA as a template. However, the mRNA level remained below the sensitivity of in situ hybridization. In contrast, in situ hybridizations of OA cartilage revealed an intense staining with the MDC15 specific riboprobes. The extension of the analysis to chondrosarcomas showed a strong up-regulation of MDC15 mRNA in these malignant transformed cells. CONCLUSION: Our results demonstrate a markedly strong up-regulation of MDC15 in adult OA and neoplastic cartilage compared with adult normal articular cartilage, indicating a potential role of the disintegrin metalloproteinase in cartilage remodeling. PMID- 10513812 TI - Evidence for neurogenic transmission inducing degenerative cartilage damage distant from local inflammation. AB - OBJECTIVE: To investigate involvement of the nervous system in ipsilateral and contralateral joint inflammation. METHODS: Freund's complete adjuvant (CFA; 1 mg or 1 microg) was injected unilaterally and the messages (a) from the hind paw to the ipsilateral and contralateral knees and (b) from one knee to the contralateral knee were analyzed. The degenerative impact of the local injury on distant cartilage was assessed using patellar proteoglycan synthesis as an indicator. Neurogenic mechanisms were blocked either by spinal cord compression or by injection of neurokinin 1 (NK-1) antagonist, or they were mimicked by intraarticular injection of substance P. The data were compared with those gathered in a model of systemic inflammation, characterized by fever and serum interleukin-6 production. RESULTS: After unilateral subcutaneous injection of CFA, proteoglycan anabolism decreased bilaterally. Spinal cord compression and administration of the NK-1 antagonist inhibited the response in the contralateral limb. Following 1 mg CFA subcutaneous injection, the ipsilateral response implicated both neurogenic and systemic mechanisms, whereas the nervous system alone was implicated after 1 microg subcutaneous CFA injection. The 1 microg CFA intraarticular injection induced a degenerative contralateral signal, which was abolished by spinal cord compression and by pretreatment with the NK-1 antagonist. Intraarticular injection of 1 microg CFA also induced an ipsilateral increase of anabolism, which was enhanced by spinal cord compression. Similar results were obtained after intraarticular injections of substance P. These effects were not reproduced with turpentine treatment, a systemic model, in which spinal cord compression had no effect. CONCLUSION: A unilateral inflammation can induce, by neurogenic mechanisms, distal bilateral degeneration of articular cartilage, implicating involvement of neuropeptides. PMID- 10513813 TI - Association analysis between the MIC-A and HLA-B alleles in Japanese patients with Behcet's disease. AB - OBJECTIVE: Behcet's disease is known to be strongly associated with HLA-B51 in many different ethnic groups. Recently, by association analysis using refined microsatellite mapping, the critical region for Behcet's disease was identified as a 46-kb segment centromeric to the HLA-B gene. No expressed gene has been detected in this segment to date except the MIC-A (major histocompatibility complex class I chain-related gene A) and HLA-B genes. The present study was undertaken to analyze allelic distribution of the MIC-A gene among Japanese patients with Behcet's disease. METHODS: Ninety-five Japanese patients with Behcet's disease and 116 ethnically matched healthy controls were enrolled in this study. MIC-A genotyping was performed by direct sequencing of polymerase chain reaction products from exons 2, 3, and 4 of the MIC-A gene, using an automated DNA sequencer. RESULTS: The MIC-A009 allele was significantly more frequent in the patient group (69.5%) compared with the healthy controls (31.0%) (relative risk 5.06, corrected P = 0.00000024). In stratification analysis on the confounding effect of MIC-A009 on HLA-B*51 association and vice versa, Behcet's disease was distinctively associated only with HLA-B*51. Further, MIC-A009 was found to be strongly associated not only with HLA-B51, but also with HLA-B52, which was not increased in the patient group to any degree. CONCLUSION: These results imply that the real disease susceptibility gene involved in the development of Behcet's disease is the HLA-B*51 allele itself and that the significant increase of the MIC-A009 allele in the patient group results secondarily from a strong linkage disequilibrium with HLA-B*51. PMID- 10513814 TI - Th1 polarization of the immune response in Behcet's disease: a putative pathogenetic role of interleukin-12. AB - OBJECTIVE: To investigate whether immunologic abnormalities in patients with Behcet's disease (BD) are related to abnormalities of the Th1/Th2 ratio. METHODS: Th1/Th2 cytokine production by peripheral blood lymphocytes (PBL) from 31 patients with BD, 11 patients with inflammatory arthritis, and 10 healthy blood donors was evaluated by intracellular immunofluorescence staining. Serum interleukin-12 (IL-12) levels were measured using an enzyme amplified-sensitivity immunoassay. The effect of recombinant IL-12 (rIL-12) on spontaneous and Fas mediated apoptosis of phytohemagglutinin (PHA)-stimulated PBL was evaluated by flow cytometry using propidium iodide (PI) staining and a bromodeoxyuridine (BrdU)/PI procedure. RESULTS: Intracellular immunofluorescence staining of IL-2, IL-4, and interferon-gamma (IFNgamma) in CD3+ lymphocytes from BD patients demonstrated a strong polarization of the immune response toward the Th1 pathway that correlated with the progression of BD. Peripheral Th1 cells were significantly increased in patients with active disease (n = 14) as compared with those in patients in complete remission (n = 17), patients with inflammatory arthritis, and normal donors. In addition, serum IL-12 levels were correlated with peripheral Th1 lymphocytes and disease progression. Apoptotic analysis revealed that PHA-activated PBL from patients with active disease were highly sensitive to spontaneous and Fas-mediated activation-induced cell death. However, addition of rIL-12 to complete medium prevented this spontaneous and Fas-induced apoptosis and enhanced the proliferation of Th1 lymphocytes. CONCLUSION: Taken together, these results indicate that a strong Th1 immune response occurs in active BD and suggest that IL-12 plays a substantial part in the pathogenesis of BD. By preventing spontaneous and Fas-induced cell death, in fact, it results in an abnormal growth of autoreactive Th1 lymphocytes that could contribute to the prolonged inflammatory autoimmune condition of BD. PMID- 10513815 TI - Modulation at multiple anchor positions of the peptide specificity of HLA-B27 subtypes differentially associated with ankylosing spondylitis. AB - OBJECTIVE: To investigate the rules governing peptide binding to HLA-B*2705, and to B*2704 and B*2706, which are 2 subtypes differentially associated with ankylosing spondylitis. METHODS: Poly-Ala analogs carrying the HLA-B27 motif Arg 2, and substitutions at anchor positions P1, P3, or Pomega, were used to determine a binding score for each residue at each position. Binding was assessed in a quantitative epitope stabilization assay, where the cell surface expression of HLA-B27 was measured by flow cytometry as a function of peptide concentration. RESULTS: Peptide anchor residues contributed additively to B*2705 binding. About 15% of the natural B*2705 ligands used a deficient P3 or Pomega anchor, but never both, indicating that detrimental anchoring at one of these positions is always compensated by a good anchor at the other one. About 50% of the B*2705 ligands used suboptimal P1 residues. However, this was compensated with optimal P3 and/or Pomega anchoring. Peptides that were longer than decamers used good anchor residues at the 3 positions, suggesting more stringent binding requirements. B*2704 and B*2706 differed in their residue specificity at P1, P3, and Pomega. The rules derived for B*2705 also applied to the known ligands of these 2 subtypes. CONCLUSION: The B*2705, B*2704, and B*2706 peptide repertoires are limited by the allowed residue combinations described in this study. The differential association of B*2704 and B*2706 with spondylarthropathy correlates with differences in their peptide specificity at multiple anchor positions. However, it is now possible to predict the peptide features that determine this differential binding to both subtypes. PMID- 10513816 TI - Differential mechanisms of inorganic pyrophosphate production by plasma cell membrane glycoprotein-1 and B10 in chondrocytes. AB - OBJECTIVE: Increased nucleoside triphosphate pyrophosphohydrolase (NTPPPH) activity in chondrocytes is associated with cartilage matrix inorganic pyrophosphate (PPi) supersaturation in chondrocalcinosis. This study compared the roles of the transforming growth factor beta (TGFbeta)-inducible plasma cell membrane glycoprotein-1 (PC-1) and the closely related B10 NTPPPH activities in chondrocyte PPi metabolism. METHODS: NTPPPH expression was studied using reverse transcriptase-polymerase chain reaction and Western blotting. Transmembrane PC-1 (tmPC-1), water-soluble secretory PC-1 (secPC-1), and transmembrane B10 were expressed by adenoviral gene transfer or plasmid transfection, and expression of PPi was assessed in cultured articular chondrocytes and immortalized NTPPPH deficient costal chondrocytes (TC28 cells). RESULTS: PC-1 and B10 messenger RNA were demonstrated in articular cartilages in situ, in untreated cultured normal articular chondrocytes, and in TC28 cells. Expression of tmPC-1 and secPC-1, but not B10, rendered the NTPPPH-deficient TC28 cells able to increase expression of extracellular PPi, with or without addition of TGFbeta (10 ng/ml) to the media. More plasma membrane NTPPPH activity was detected in cells transfected with tmPC 1 than in cells transfected with B10. Furthermore, confocal microscopy with immunofluorescent staining of articular chondrocytes confirmed preferential plasma membrane localization of PC-1, relative to B10. Finally, both PC-1 and B10 increased the levels of intracellular PPi, but PC-1 and B10 appeared to act principally in different intracellular compartments (Golgi and post-Golgi versus pre-Golgi, respectively). CONCLUSION: PC-1 and B10 NTPPPH activities were not redundant in chondrocytes. Although increased PC-1 and B10 expression caused elevations in intracellular PPi, the major effects of PC-1 and B10 were exerted in distinct subcellular compartments. Moreover, PC-1 (transmembrane and secreted), but not B10, increased the levels of extracellular PPi. Differential expression of PC-1 and B10 could modulate cartilage mineralization in degenerative joint diseases. PMID- 10513817 TI - The relationship between pityriasis rubra pilaris and inflammatory arthritis: case report and response of the arthritis to anti-tumor necrosis factor immunotherapy. AB - Pityriasis rubra pilaris (PRP) refers to a group of erythematous, scaling dermatologic conditions that have been associated with seronegative arthritis. We report a case of polyarthritis in a young man with PRP in which magnetic resonance imaging suggested an entheseal-based pathology for the joint disease. The arthritis, but not the skin condition, demonstrated dramatic response to anti tumor necrosis factor immunotherapy. PMID- 10513818 TI - Hypertrophic osteoarthropathy can indicate recurrence of Whipple's disease. AB - We report the case of a patient with Whipple's disease (WD) who developed hypertrophic osteoarthropathy (HOA) characterized by digital clubbing, periostosis of the tubular bones, and polysynovitis. The HOA disclosed the recurrence of the patient's WD, since polymerase chain reaction (PCR) analysis clearly demonstrated the presence of Tropheryma whippelii in the synovial fluid from the patient's left knee. Initiation of appropriate antibiotic therapy resulted in complete healing of all clinical rheumatologic manifestations within 2 months and in disappearance of radiographic bone changes at 7-month followup. We suggest that HOA be included within the spectrum of rheumatologic manifestations of WD, and that an evaluation for WD should be considered in patients, especially middle-aged men, presenting with HOA even without gastrointestinal symptoms. PCR analysis may be useful in accurate diagnosis and management of early WD with unusual clinical manifestations, and may contribute to decreased morbidity and mortality. PMID- 10513819 TI - Mixed cryoglobulinemia secondary to visceral Leishmaniasis. AB - We describe a case of type II mixed cryoglobulinemia, with monoclonal IgMkappa rheumatoid factor, associated with visceral leishmaniasis caused by Leishmania infantum. Involvement of Leishmania antigen(s) in the formation of cryoprecipitable immune complexes was suggested by the fact that cryoglobulinemic vasculitis subsided after antiparasite therapy and that anti-Leishmania antibodies, as well as rheumatoid factor, were enriched in the cryoprecipitate. We observed 2 additional patients with visceral leishmaniasis and cryoglobulinemic vasculitis. All 3 patients had seemingly contracted leishmaniasis in Italy, were hepatitis C virus negative, and were initially diagnosed as having autoimmune disorders. These findings indicate that Leishmania can be an etiologic agent of type II mixed cryoglobulinemia. This parasitosis should be taken into consideration in the differential diagnosis of vasculitides in endemic areas. PMID- 10513820 TI - Clinical Image: acute gout in a young man with osteochondromatosis. PMID- 10513821 TI - Fluorosis and osteomalacia. PMID- 10513822 TI - Increased expression of multidrug resistance of P-glycoprotein on Th1 cells correlates with drug resistance in rheumatoid arthritis. PMID- 10513823 TI - The HLA-DRB1 QR/KRAA sequence cannot alone explain the rheumatoid arthritis susceptibility in the Corsican population. PMID- 10513824 TI - Single nucleotide polymorphic haplotypes of the interleukin-10 5' flanking region are not associated with renal disease or serology in Caucasian patients with systemic lupus erythematosus. PMID- 10513825 TI - Intravenous pulse versus oral administration of cyclophosphamide: comment on the article by Haubitz et al. PMID- 10513826 TI - Sensitivity and specificity of tests for autoantibodies: comment on the article by Tan et al. PMID- 10513827 TI - What constitutes a mortality study? Comment on the article by Uramoto et al. PMID- 10513828 TI - Airpuff startle stress elicited fos expression in brain cardiovascular areas of young SHR and WKY rats. AB - Prior studies comparing Fos expression in adult Wistar Kyoto (WKY) and Spontaneously Hypertensive rats (SHR) identified more Fos-positive neurons in a subset of brain regions following two stressors: placement in a startle chamber and presentation of an airpuff startle stimulus. The present study assessed Fos expression in five week old SHR and WKY rats in those same brain areas. Like adults, young SHR expressed more Fos-positive neurons than WKY in response to the startle chamber alone. Unlike adults, in the SHR only the locus coeruleus showed a increases in Fos expression following addition of the airpuff. Otherwise, startle chamber and airpuff startle treatments induced roughly equivalent Fos expression in the SHR, possibly reflecting a ceiling effect. Young WKY exhibited predominant airpuff-induced elevations. The present results demonstrate that certain brain regions are strain-differentially activated by stressors prior to overt hypertension and that differential Fos expression is an early developmental feature of these strains. PMID- 10513829 TI - The role of the parathyroid glands in adrenocorticotrophin-induced hypertension in the rat. AB - The aim of this study was to determine whether parathyroidectomy (PTx) would modify hypertension secondary to adrenocorticotrophin (ACTH) administration. Male Sprague Dawley (SD) rats were randomly assigned to one of five groups; (i) sham (saline) treatment (NaCl 0.9% s/c 0.5 ml/kg/day), (ii) ACTH treatment (Synacthen Depot 0.5 mg/kg/day), (iii) saline/PTx/1% CaCl2 in water, (iv) ACTH/PTx/1% CaCl2 in water and (v) ACTH/1% CaCl2 in water. Tail cuff systolic blood pressure (SBP) and metabolic parameters were measured on alternate days for 4 control (C) and 11 treatment days (T0-T10). There was no change in SBP in the sham and saline/PTx/CaCl2 groups over T0-10. SBP increased in the ACTH treated groups. PTx did not modify ACTH-induced increases in SBP or metabolic effects. These results do not support a role for the parathyroids in the genesis of ACTH-induced hypertension in the rat. PMID- 10513830 TI - Efficacy and safety of an oral fixed low-dose perindopril 2 MG/indapamide 0.625 MG combination: a randomized, double-blind, placebo-controlled study in elderly patients with mild to moderate hypertension. AB - The efficacy and safety of 12 weeks treatment with an oral fixed low-dose perindopril 2 mg + indapamide 0.625 mg (Per/Ind) combination in elderly and very elderly patients (65-85 years) with mild to moderate systolic and diastolic hypertension (SDH) or isolated systolic hypertension (ISH) were investigated vs placebo. This trial was a multinational randomized double-blind study with doubling of active drug dosage in nonresponders. Intention to treat analysis was performed in 383 patients (age 72.4 years; ISH 32%). 58.5% remained on their initial dosage. Per/Ind decreased supine diastolic and systolic blood pressure (sDBP/sSBP) by 13.2+/-8.0 mm Hg and 22.5+/-13.9 mm Hg (P <.0001) versus placebo 7.3+/-9.0 mm Hg and -12.3+/-15.2 mm Hg, respectively. In ISH (n = 123), Per/Ind decreased sSBP by 23.0+/-11.8 mm Hg (P <.0001). Overall response and normotension rates was 81.3% with Per/Ind (P <.0001). Adverse event rates (including hypokalemia) were similarly low in both groups. Analysis in the over-75 year subgroup showed similar safety and efficacy results. Fixed low-dose Per/Ind is a safe and effective treatment of hypertension including isolated systolic hypertension in the elderly. PMID- 10513831 TI - Effect of blockade of nitric oxide synthesis on renin secretion in human subjects. AB - Nitric oxide (NO) has been implicated in the control of renin secretion in experimental animals but little information is available concerning its role in humans. The aim of the present study was to investigate the effects of inhibition of NO synthesis on resting renin secretion and on the renin secretory responses to activation of the macula densa and sympathetic neural mechanisms controlling renin secretion. In eight healthy subjects, injection of furosemide increased plasma renin activity (PRA) with little or no change in blood pressure or heart rate. Injection of the NO synthase inhibitor L-NMMA increased blood pressure and decreased heart rate and PRA, but failed to alter the PRA response to furosemide. In another ten subjects, standing increased PRA. L-NMMA again decreased PRA but failed to alter the PRA response to standing. These results suggest that NO participates in the regulation of resting renin secretion in humans, and provide preliminary evidence that NO does not contribute significantly to the renin responses to activation of the macula densa or sympathetic mechanisms controlling renin secretion. PMID- 10513832 TI - Increased plasma atrial natriuretic factor in catecholamine-producing tumor patients. AB - The aim of this study was to evaluate plasma levels of ANF in patients with catecholamine-secreting tumors with and without hypertension and to relate ANF secretion to levels of plasma and urinary catecholamines and blood pressure. Twenty-one pheochromocytoma (15 with sustained, 6 with paroxysmal hypertension), 6 neuroblastoma (1 hypertensive) patients and 28 aged-matched controls were studied in basal conditions. Plasma and urinary norepinephrine (NE),epinephrine (E), dopamine (DA) and DOPA were determined by HPLC-ED and plasma ANF by RIA. Both neuroblastoma and pheochromocytoma patients had significantly higher plasma ANF levels than controls. Neuroblastomas showed higher ANF concentration than pheochromocytomas. No differences were found in plasma ANF between hypertensive and normotensive patients. Pheochromocytomas with ANF levels within the normal range had plasma and urinary NE and urinary DA and DOPA levels significantly higher than patients with high ANF. Plasma ANF levels were unrelated to systolic or diastolic blood pressure or heart rate. A negative correlation between plasma ANF and urinary DA was found only in the patients groups. In conclusion, plasma ANF was increased in pheochromocytoma and neuroblastoma patients. Our data suggest that the excessive catecholamine secretion is not responsible for the increased ANF secretion in these patients. The significance of the relationships among plasma ANF and urinary and plasma catecholamines requires further investigation. PMID- 10513833 TI - Human tissue kallikrein attenuates hypertension and secretes into circulation and urine after intramuscular gene delivery in hypertensive rats. AB - Systemic delivery of the human tissue kallikrein transgene has been shown to markedly delay the increase of blood pressure in hypertensive rat models. To demonstrate potential hypotensive effects of kallikrein via local delivery, adenovirus carrying the human tissue kallikrein gene was inoculated into quadriceps of spontaneously hypertensive rats (SHR). A single intramuscular injection of the kallikrein gene caused a significant delay of blood pressure increase for 5 weeks. The expression of human tissue kallikrein and its mRNA was identified solely in injected muscle. Immunoreactive human tissue kallikrein was detected in the muscle as well as in the circulation and urine of adult and newborn rats. Urinary kinin and cGMP levels increased significantly in rats receiving kallikrein gene delivery as compared with rats receiving control virus containing the LacZ gene. The detection of human tissue kallikrein in rat urine after local gene delivery into the muscle provides direct evidence that circulatory kallikrein can be secreted into the urine. These findings indicated that a continuous supply of human tissue kallikrein in the circulation is sufficient to reduce blood pressure and kallikrein gene delivery via the intramuscular route may have significant implications in therapeutic applications. PMID- 10513834 TI - A sibling-pair analysis of fasting lipids and anthropometric measurements and their relationship to hypertension. AB - Fifty non-diabetic, young Chinese hypertensives were compared to their normotensive siblings with respect to body fat distribution and fasting lipid and glucose (FPG) profiles. Sitting BP in hypertensives met conventional hypertension criteria after a 4-week washout period on placebo. Hypertensives had greater body mass index (BMI), subscapular skin-fold thickness (SFT), waist circumference (W), waist-to-height (WHtR) and waist-to-hip ratio (WHR), p<0.0001 for all. Higher triglycerides (p=0.004) and lower HDL-cholesterol (p=0.015) concentrations were also observed. Weight, W, WHtR and BMI were higher in hypertensives in both male (n=9) and female gender-concordant sibling pairs (n=21). Higher WHR, hip circumference and subscapular SFT were only seen in the male hypertensives. Hypertension is associated with central adiposity and adverse lipid profiles in this group of young hypertensives, supporting the hypothesis that obesity, particularly central, is closely associated with hypertension in Chinese as in other ethnic groups. PMID- 10513835 TI - Effects of nilvadipine on cytokine-levels and soluble factors in collagen disease complicated with essential hypertension. AB - We examined some immunological parameters, particularly cytokines and soluble factors in collagen diseases complicated with essential hypertension. We also investigated the effects of Nilvadipine on immunological parameters after treatment with this drug for six months. The frequency of helper/inducer T cells (CD4+ CD8- cells, CD4+ CD45RA- cells) decreased in the peripheral blood on a 6 month treatment with nilvadipine. There was a significant decrease of suppressor/inducer T cells (CD4+ 45RA+ cells), and an insignificant decrease of activated T cells (CD3+ HLA-DR+ cells) and memory T cells (CD45RA- CD45RO+ cells) after treatment. Before treatment with Nilvadipine, interleukin-1beta, tumor necrosis factor-a, and interleukin-6 levels increased higher in the patients than in healthy volunteers. However, interleukin-1beta and interleukin-6 concentrations tended to decrease after treatment with Nilvadipine. Besides, tumor necrosis factor-alpha decreased significantly after treatment. The soluble interleukin-2 receptor concentrations also showed a decreased tendency after treatment, although high concentrations were found in the patients before treatment. In contrast, soluble human leukocyte antigen-1 and soluble thrombomodulin levels showed no significant change after treatment. These results suggest that Nilvadipine inhibits the generation of cytokines derived from activated T lymphocytes. Nilvadipine, calcium antagonist, may be useful for inhibition of vascular complication in collagen diseases. PMID- 10513836 TI - Daily response of blood pressure to day-to-day variation of urinary sodium to potassium ratio. AB - The relationship between blood pressure and urinary sodium-to-potassium (Na/K) ratio was assessed in eight healthy men, none of whom used antihypertensive medications. Blood pressure and urinary sodium and potassium concentrations were measured for 11 to 33 days without any dietary restriction. For two of the eight subjects, the urinary Na/K ratio significantly correlated with systolic blood pressure (r=0.70 and 0.45, respectively), and in one of the two subjects, the urinary ratio also positively correlated with diastolic blood pressure (r=0.72). In the others, no relationship between the ratio and blood pressure was observed (r=-0.24 to 0.26). The results indicate that, in some individuals, the daily variation of urinary Na/K ratio is closely correlated with day-to-day changes in blood pressure level, and suggest that the urinary Na/K ratio is useful in the management of the daily sodium and potassium intake balance of hypertensive patients who need to restrict salt intake. PMID- 10513837 TI - Arterial relaxation mediated by endothelium-derived hyperpolarizing factor in hypertension induced by chronic inhibition of nitric oxide synthesis. AB - The aim of this study was to evaluate arterial relaxation mediated by endothelium derived hyperpolarizing factor (EDHF) during chronic inhibition of nitric oxide (NO) synthase. We measured the isometric tension of isolated mesenteric arteries of Wistar rats administered Nomega-nitro-L-arginine methyl ester (L-NAME, 100 mg/Kg/day) for 3 weeks. Relaxation to acetylcholine (ACh) was reduced in L-NAME treated rats (maximum relaxation, 52% versus 79% ). After acute superfusion of 1x10(-4) M L-NAME, half the relaxation was inhibited in controls, while the relaxation was not changed in L-NAME treated rats. In contrast, relaxation to nitroprusside was normal in L-NAME treated rats. Superfusion of 1x10(-6) M apamin, which inhibits the effects of EDHF, reduced the relaxation. The relaxation inhibited by apamin was not significantly different between the two groups. These findings suggested that in endothelial cells, the synthesis of EDHF is unchanged during a chronic deficiency of relaxation influence of NO. PMID- 10513838 TI - Evaluation of high through-to-peak ratio of perindopril in SHR. AB - The present study was designed to evaluate trough-to-peak ratio (T/P) of ACE inhibitors in spontaneously hypertensive rats (SHR) by a continuous monitoring of ambulatory blood pressure for 24 hours with a biotelemetric system. Blood pressure was recorded uninterruptedly with a battery-operated transmitter connected to a sensor catheter. Perindopril (3 mg/kg), trandolapril (1 mg/kg), quinapril (10 mg/kg) and enalapril (6 mg/kg) were given once a day for 7 days. On the first day of the treatment these ACE inhibitors equally decreased blood pressure by 20 mmHg at each peak. The peak and trough blood pressure decreased steadily until day 4, and then they were constant until the end of experiment (day 7). T/P for each inhibitor also increased until day 4, and the ratios in systolic blood pressure at the end of experiments (day 7) were as follows, perindopril: 0.63, trandolapril: 0.62, quinapril: 0.41, enalapril: 0.27. The T/P of perindopril was significantly higher than that of enalapril. The results of the present studies testing four ACE inhibitors are well consistent with those in clinical trials. Thus, the measurement of T/P in SHR would provide a meaningful information for the evaluation of antihypertensive agents like ACE inhibitors. PMID- 10513840 TI - Antiviral and hemolytic activities of surfactin isoforms and their methyl ester derivatives. AB - Inactivation of enveloped viruses (VSV, SFV, and SHV-1) by surfactin lipopeptides was dependent on the hydrophobicity, i.e. the number of carbon atoms of the fatty acid, and on the charge of the peptide moiety as well as on the virus species. Surfactins with fatty acid chains of 13 carbon atoms showed very low antiviral activity in comparison to C14 and C15 isoforms. C15 surfactin monomethyl ester also inactivated SFV which was resistant to the mixture of surfactin isoforms as produced by Bacillus subtilis. In contrast, the dimethyl ester showed no virus inactivation capacity. Disintegration of viral structures as determined by electron microscopy after inactivation of VSV and SFV was comparable to the titer reduction. The effect of the surfactin isoforms and methyl esters on erythrocyte hemolysis correlated with the virus-inactivation capacity. Surfactins with a fatty acid chain moiety of 15 carbon atoms and one negative charge showed the highest antiviral activity. PMID- 10513841 TI - Bioconversion of milbemycin-related compounds: isolation and utilization of non producer, strain RNBC-5-51. AB - A non-producing strain, the so-called strain RNBC-5-51 SANK 60198, was isolated during a screening program of strain improvement in the milbemycin production. Strain RNBC-5-51 indicated almost the same characteristics as those in the parent strain, that is, the abundant spore formation on agar media and the good growth in liquid media. But it does not produce any kind of milbemycins. In addition, strain RNBC-5-51 accumulated precursor-like compounds of milbemycin-polyketide, the production of which were inhibited by the addition of cerulenin. In the bioconversion experiments, strain RNBC-5-51 converted milbemycin beta6 and A4 to milbemycin alpha14, and milbemycin beta7 and A3 to milbemycin alpha11, respectively. This strain also converted milbemycin D and avermectin B1a, to 26 (3-methyl-2-butenoyloxy)milbemycin D and 26-(3-methyl-2-butenoyloxy)avermectin B1a, respectively. These results suggest that milbemycin alpha11 is biosynthesized through the same route as milbemycin alpha14, and the mutated step in strain RNBC-5-51 might be in the polyketide synthetic pathway of milbemycins. Strain RNBC-5-51 loses the ability for de novo synthesis of milbemycins, but it retains the ability to bioconvert the milbemycin skeleton. This strain might be useful for C-26 modification of milbemycin-related compounds. PMID- 10513839 TI - TMC-66, a new endothelin converting enzyme inhibitor produced by Streptomyces sp. A5008. AB - A new endothelin converting enzyme (ECE) inhibitor, TMC-66 was isolated from the fermentation broth of Streptomyces sp. A5008. The structure of TMC-66 was elucidated by spectroscopic analyses to be a new member of benzo[a]naphthacenequinone class of antibiotics. TMC-66 had a highly selective inhibitory activity for ECE with an IC50 value of 2.9 microM. Taxonomy of the producing strain is also described. PMID- 10513842 TI - Mode of action of anhydrofulvic acid against Candida utilis ATCC 42402 under acidic condition. AB - The mode of action of anhydrofulvic acid against Candida utilis ATCC 42402 was investigated under acidic conditions. Anhydrofulvic acid inhibited the incorporation of radioactive precursors into DNA, RNA, protein and lipid fractions. Although it did not induce leakage of intracellular materials from the treated cells, it had inhibitory effects on both endogenous and exogenous cellular respiration. Moreover, it inhibited mitochondrial respiration of Candida utilis ATCC 42402 using both succinate and cytochrome c as respiratory substrates, but not using NADH. Unexpectedly, the inhibition against isolated mitochondria was observed at pH 7.0. These results suggested that the action site against the respiratory inhibition of anhydrofulvic acid might be involved in succinate dehydrogenase, complex II in the mitochondrial electron transport chain of the yeast cells. Judging from the inhibitory effect of anhydrofulvic acid on mitochondria detected at pH 7.0, it was postulated that the antifungal activity at a low pH level might depend on the elevation of drug permeability to the cell membrane under acidic conditions. PMID- 10513843 TI - Cineromycins, gamma-butyrolactones and ansamycins by analysis of the secondary metabolite pattern created by a single strain of Streptomyces. AB - The antifungal and antibacterial properties of the crude extract from Streptomyces sp. (strain Go 40/10) encouraged us to perform a detailed analysis of its secondary metabolite pattern by chemical screening, which revealed the presence of at least 30 different compounds. Ten of the isolated 18 metabolites proved to be new. Remarkable are hydrogenated (3, 4) and glucosylated (5, 6, 7) 14-membered macrolides derived from cineromycin B, two gamma-butyrolactones (8, 9), the so far unknown naphthomycin K (14) and the collinolactones A and B. The constitution and relative stereochemistry of these metabolites were deduced from spectroscopic data. Finally the concept of our one strain/many compounds (OSMAC) method for exploring new microbial secondary metabolites is discussed. PMID- 10513844 TI - Convergent syntheses of oral THF 1beta-methylcarbapenems. AB - Convergent syntheses of oral THF 1beta-methylcarbapenems 4 (OCA-983) and 5 starting from M2-phosphate 1 were developed. Reaction of the M2-phosphate 1 with THF thiols containing a requisite prodrug side chain, 9 and 10, gave the desired oral THF 1beta-methylcarbapenems 4 and 5, respectively, in 46% and 42% overall yields. PMID- 10513845 TI - In vitro antibacterial activity of FK041, a new orally active cephalosporin. AB - The in vitro activity of FK041, a new orally active cephem antibiotic, against a wide variety of clinical isolates of bacteria was investigated and compared with those of cefdinir (CFDN) and cefditoren (CDTR). FK041 exhibited broad spectrum activity against reference strains of Gram-positive and Gram-negative aerobes and anaerobes. FK041 was active against clinical isolates of Gram-positive organisms except Enterococcus faecalis with MIC90s less than 1.56 microg/ml. FK041 was more active than CFDN and CDTR against Staphylococcus aureus, Staphylococcus epidermidis, and Streptococcus agalactiae and was comparable to CFDN and CDTR against Streptococcus pyogenes and Streptococcus pneumoniae. FK041 had no activity against methicillin-resistant staphylococci, like CFDN and CDTR. FK041 showed moderate activity against penicillin-resistant S. pneumoniae with an MIC range of 0.05 approximately 3.13 microg/ml, and was superior to CFDN but inferior to CDTR. Against clinical isolates of many Gram-negative organisms such as Neisseria gonorrhoeae, Escherichia coli, Klebsiella pneumoniae, and Proteus mirabilis, FK041 had good activity comparable or superior to those of CFDN and CDTR. However, it was inferior to CDTR in activity against Moraxella catarrhalis, Haemophilus influenzae, Morganella morganii, and Serratia marcescens, and was inactive against Pseudomonas aeruginosa. With FK041 a small difference between MIC and MBC against S. aureus, E. coli, K. pneumoniae, and H. influenzae was found, indicating that its action is bactericidal against these species. FK041 was stable to group 2beta-lactamase hydrolysis but was unstable to group 1beta lactamase hydrolysis. The stability of FK041 to these enzymes was similar to those of CFDN and CDTR. FK041 showed high affinity for the main penicillin binding proteins (PBPs) of S. aureus (PBP 3, 2, and 1) and E. coli (PBP 3, 4, lbs, 2, and 1a). PMID- 10513846 TI - In vivo antibacterial activity of FK041, a new orally active cephalosporin. AB - The therapeutic activities of orally administered FK041 were evaluated in mouse models of systemic and local infections with a variety of bacteria and were compared with those of cefdinir (CFDN) and cefditoren pivoxil (CDTR-PI). FK041 exhibited potent therapeutic activity against lethal systemic infections induced by intraperitoneally inoculated Staphylococcus aureus, Escherichia coli, and Klebsiella pneumoniae with 50% effective doses (ED50) in the range of 0.20 to 0.36 mg/kg and was more active than CFDN and CDTR-PI. This result correlated well with its in vitro activity. The therapeutic effects of FK041 and reference drugs on murine local infections were evaluated in an in vivo pharmacokinetic model simulating human plasma concentrations for oral administration of 50 mg, 100 mg, and 200 mg. Against murine subcutaneous abscess induced by S. aureus, FK041 was as effective as CFDN and significantly more effective than CDTR-PI in reducing the number of recoverable viable bacteria in the skin at the infection sites. The efficacy of FK041 against murine pneumonia with H. influenzae was comparable to that of CDTR-PI and was superior to that of CFDN in reducing viable bacteria activity in the lungs. These results strongly suggest that FK041 has potential for clinical use against various bacterial infections. PMID- 10513847 TI - Isolation and structural elucidation of new peptaibols, bergofungins B, C and D, from Emericellopsis donezkii HKI 0059. PMID- 10513848 TI - Macquarimicin A inhibits membrane-bound neutral sphingomyelinase from rat brain. PMID- 10513850 TI - Introduction: treatment of depression in long-term care patients. PMID- 10513849 TI - FK463, a novel water-soluble echinocandin lipopeptide: synthesis and antifungal activity. PMID- 10513851 TI - Adverse reactions of antidepressants in elderly patients. AB - As patients age, adverse drug reactions increase dramatically in frequency and severity. Although the population of elderly patients in controlled studies of antidepressants is small, key factors have been identified that may influence proper dosing, including the different pharmacokinetic properties of antidepressants in elderly compared with younger patients and individual patient characteristics. In elderly patients, antidepressant side effects of concern include orthostatic hypotension, anticholinergic effects, extrapyramidal symptoms, and syndrome of inappropriate antidiuretic hormone secretion. A routine treatment procedure that includes risk-factor identification, a thorough drug history, patient and caregiver education on compliance, and use of the lowest effective dose can help clinicians effectively manage elderly patients and limit the risk of adverse drug reactions. PMID- 10513852 TI - Epidemiology and diagnosis of depression in late life. AB - Depression is a significant concern in elderly patients. Reported prevalence rates differ greatly depending on the definition of depression and the population of interest, with increases reported in settings where comorbid physical illnesses are more common. In community-dwelling elderly patients, prevalences of depressive symptoms and major depressive disorder are 15% and 1% to 3%, respectively. Factors associated with depression in the elderly include female gender, alcohol and substance abuse, pharmaceuticals, family history, and medical conditions such as stroke, Alzheimer's disease, cancer, and heart disease. Recognition of depression is complex because patients often deny their depression, present with somatic complaints, or may have comorbid anxiety or cognitive impairment. Depression is underrecognized and undertreated in the elderly, despite evidence that the benefits of treatment outweigh potential risks. PMID- 10513854 TI - Continuation and maintenance pharmacotherapy in geriatric depression: an open trial comparison of paroxetine and nortriptyline in patients older than 70 years. AB - We present preliminary data on the efficacy of paroxetine, as compared with nortriptyline, in preventing or delaying relapse and recurrence of major depression in elderly patients. Following double-blind, acute-phase pharmacotherapy, 25 patients (mean age = 72.5 years) began open-trial continuation treatment with paroxetine (mean dose = 24.5 mg/day), and 15 patients (mean age = 77.5 years) received nortriptyline (mean dose = 51.3 mg/day; mean blood level = 85.5 ng/mL). Over an 18-month period, paroxetine and nortriptyline have shown comparable efficacy in preventing or delaying relapse and recurrence, with 80% to 90% of patients remaining well. These data suggest that paroxetine holds promise for long-term maintenance treatment in patients in their 70s and older with depression; however, further controlled evaluation is necessary. PMID- 10513853 TI - A double-blind randomized comparison of nortriptyline and paroxetine in the treatment of late-life depression: 6-week outcome. AB - BACKGROUND: Some studies have suggested that selective serotonin reuptake inhibitors may be less efficacious than tricyclic antidepressants in the treatment of severe depression in older patients. The objective of this study was to compare the 6-week outcome of treatment with nortriptyline and paroxetine in older patients with a major depressive episode. METHOD: A double-blind randomized comparison of nortriptyline and paroxetine was conducted in 80 elderly (mean +/- SD age = 75.0 +/- 7.4 years) psychiatric inpatients and outpatients who presented with a major depressive episode. Dropout and response rates were compared in patients who began or completed treatment. Rates of response of inpatients and patients with melancholic depression were also compared. RESULTS: Over 6 weeks, there were no significant differences in dropout rates due to side effects (nortriptyline, 14% vs. paroxetine, 19%) or for any reason (27% vs. 33%). Similarly, there were no significant differences between the rates of favorable response to nortriptyline or paroxetine (intent-to-treat analysis, 57% vs. 44%; completer analysis, 78% vs. 66%). Analyses restricted to inpatients or to patients with melancholic depression yielded similar results. CONCLUSION: Nortriptyline and paroxetine appear to have similar efficacy and tolerability in the acute (6-week) treatment of older depressed patients, including hospitalized patients and those with melancholic features. PMID- 10513855 TI - Cognitive effects of paroxetine in older depressed patients. AB - This study examined whether paroxetine produces cognitive toxicity in elderly patients suffering from a major depressive episode. Twenty-nine depressed patients with a wide range of cognitive functioning were treated with paroxetine. At baseline and during 6 weeks of treatment, patients were asked to complete various measures of cognitive function and had blood drawn to determine serum anticholinergicity. Measures of attention and cognitive speed showed significant improvement with treatment, while the memory performance remained unchanged. A similar pattern of results was found in both cognitively impaired and intact patients. The slight increase in serum anticholinergicity seen in some elderly patients did not significantly impair cognitive function, even in patients with a preexisting cognitive impairment. PMID- 10513856 TI - Practical considerations for the treatment of depression in elderly and very elderly long-term care patients. AB - Depression in the elderly and very elderly, especially those in long-term care facilities, often is more difficult to treat than depression in young or middle aged adults. Because this population may be more sensitive to the common adverse effects of many antidepressant drugs, particularly the anticholinergic side effects, administration of pharmacologic therapy for depression in the elderly requires careful consideration of the side effect profiles of the various classes of antidepressant medication. PMID- 10513857 TI - Treatment of depression in patients with heart disease. AB - Patients with depression are more likely than patients without depression to develop ischemic heart disease and suffer cardiac-related death. Recent evidence suggests that the association between depression and increased cardiac mortality may in part be due to an increase in platelet activity and an imbalance in sympathetic and parasympathetic activity that makes the patient more susceptible to ventricular fibrillation. Available data suggest that the tricyclic antidepressants (TCAs) may increase the risk of mortality in patients with ischemic heart disease. Three studies with the selective serotonin reuptake inhibitors (SSRIs), including a double-blind, randomized comparison of paroxetine with nortriptyline, support the conclusion that the SSRIs have a relatively benign cardiovascular profile. Therefore, they are preferable to the TCAs for treatment of depression in patients at risk for cardiac events. Additional studies are needed to definitively establish the cardiovascular safety of the SSRIs. PMID- 10513859 TI - Primary care issues related to the treatment of depression in elderly patients. AB - Late-life depression is a serious public health problem and a concern for the primary care physician. Illnesses that often occur with aging may present in association with depression, which can interfere with patient compliance and recovery and worsen disease outcomes. Late-life depression is also associated with disproportionately high rates of completed suicide and high mortality rates independent of suicide. A shared therapeutic nihilism exists between many patients and physicians, who inappropriately accept major depression as normal and inevitable during advanced age and with related chronic disease states. Thus, the older depressed patient is too often not diagnosed and not treated. Furthermore, symptom overlap between depression, anxiety, and many chronic medical illnesses may confuse proper diagnosis. Therefore, screening for and diagnosing depression using an inclusive approach is highly recommended in the primary care setting and long-term care facility. Because of their improved safety, tolerability, and ease of dosing, newer generation antidepressants, such as the selective serotonin reuptake inhibitors, should be the first choice of treatment. Collaboration between primary and specialty providers is recommended, and referral to psychiatry is advised for patients with complex medical illnesses, comorbid psychiatric illness, suicidal ideation or intent, complicated medication regimens, and poor or no response to antidepressant therapy. PMID- 10513860 TI - The gift of authorship. PMID- 10513858 TI - Pharmacokinetic considerations of antidepressant use in the elderly. AB - The elderly can be more difficult to treat with antidepressants than are young adults. Older patients take more medications, which increases the potential for drug interactions. Their altered physiology results in widely variable plasma drug concentrations from standard doses. Pharmacokinetic and pharmacodynamic changes in this population may predispose patients to experience an increased number of adverse events. The newer antidepressants have been studied for pharmacokinetic changes in the elderly compared with younger volunteers. Most antidepressants show some changes, including slower metabolic clearance and an extended elimination half-life, although the changes described are not uniform between drugs. Recommended initial doses are lower for the elderly for all antidepressants, although optimal doses may not differ from those for younger patients once dosing is individualized. PMID- 10513861 TI - The effect of posture and anesthesia on the occlusal relationship in orthognathic surgery. AB - PURPOSE: This study investigated the differences in centric occlusion (intercuspal-CO) and centric relation contact position (CR) in patients in an upright and supine posture when conscious and when supine under general anesthesia to rationalize planning for complex bimaxillary osteotomies. PATIENTS AND METHODS: Twenty-four men and 37 women, with a mean age of 24.7 +/- 4.5 years were included in this study. Occlusal records on each patient included 1 in CO and 3 in CR in each of the following postures: conscious upright, conscious supine, and anesthetized supine. Casts were mounted in the Denar Mark II articulator and transferred to the Denar Vericheck (Denar Corp, Anaheim, CA) recording tool for measurements. The centric relation displacement vectors (CO CR) were projected in various axes by 3D graphics. RESULTS: The upright conscious CO-CR (mm) vectors were significantly different from both the supine conscious and anesthetized positions in the anteroposterior plane. The difference in the vertical plane was only significant in Class II division 1 patients. CONCLUSION: The posterior change in centric relation explains the apparent occlusal discrepancies created by rigid fixation of the mandible applied in the supine anesthetized position. By taking the CR records in the supine conscious position, the "loss" of maxillary advancement can be avoided. PMID- 10513862 TI - Osseous remodeling after inferior border osteotomy for chin augmentation: an indication for early surgery. AB - PURPOSE: This study investigated the relationship of age at surgery and type of fixation to the pattern and extent of bone remodeling associated with inferior border osteotomy for chin augmentation. PATIENTS AND METHODS: Four groups of patients with similar chin advancement were established by age at the time of surgery: younger than 15, 15 to 19, 20 to 24, and older than 39 years. Cephalometric radiographs for immediate preoperative, immediate postoperative, and at least 9 months postoperative times were traced, digitized, and superimposed. RESULTS: The pattern of osseous remodeling was similar for all age groups. This consisted of resorption of the superior-buccal aspect of the distal segment, bone apposition on the buccal surface of the proximal segment, and modest resorption at pogonion (mean change, 1 mm or less). There was no significant difference in stability of the chin advancement between wire and rigid (screw) fixation. There was a marked difference in the symphysis thickness regeneration of the youngest group (92% of the original symphysis thickness) compared with the rest of the groups (< or =66%, P < .001). CONCLUSIONS: Minimal remodeling at pogonion occurs in all age-groups with both wire and rigid fixation. Regeneration of symphysis thickness is much more complete in patients younger than 15 years at the time of surgery. This is potentially important for early treatment of severe chin deficiency, because it permits additional advancement of the chin later in life, if necessary. PMID- 10513863 TI - Differences in soft tissue profile changes following mandibular setback in Chinese men and women. AB - PURPOSE: This study investigated the soft tissue profile changes after surgical correction of mandibular prognathism in Chinese patients and evaluated the sex differences in the ratios of soft tissue to hard tissue change. PATIENTS AND METHODS: Forty-three adult Chinese patients (18 men and 25 women) with mandibular prognathism were treated by intraoral oblique or vertical ramus osteotomy. Lateral cephalometric radiographs were taken presurgically and between 6 to 12 months postsurgically, and specific soft and hard tissue points were digitized and analyzed with a computer-aided system. RESULTS: As a result of surgical setback of the mandible, the soft tissue to hard tissue change ratios were: Li:Ii = 0.71:1, Si:B = 0.90:1, Pg':Pg = 0.94:1 for the males; and Li:Ii = 0.82:1, Si:B = 0.92:1, Pg':Pg = 1.06:1 for the females. CONCLUSION: The changes in soft tissue were closely correlated with the hard tissue movement after surgical setback of mandible. The average ratios of soft tissue to hard tissue change in horizontal direction appear to show a gender difference, which suggests the need for different ratios when predicting the results of orthognathic surgery. PMID- 10513864 TI - Relationship between joint effusion, joint pain, and protein levels in joint lavage fluid of patients with internal derangement and osteoarthritis of the temporomandibular joint. AB - PURPOSE: The purpose of this study was to investigate the relationship between the presence of joint effusion, joint pain, and protein levels in joint lavage fluid (JL) of patients with internal derangement (ID) and osteoarthritis (OA) of the temporomandibular joint (TMJ). PATIENTS AND METHODS: Thirty-eight joints in 26 patients with ID and OA of the TMJ were studied. Magnetic resonance imaging (MRI) evidence of joint effusion was evaluated in T2-weighted images. Samples of JL were collected from the superior joint space during pumping manipulation, and the protein concentration was measured. The presence of pain was based on joint tenderness or a complaint of pain in the preauricular region during mouth opening or closing. RESULTS: Joint effusion was demonstrated in 20 of 25 (80%) painful joints; a significantly higher incidence than in pain-free joints (5 of 13, 38.5%). The mean protein concentration (2.15 mg/mL) in JL from painful joints was significantly higher than in pain-free joints (1.22 mg/mL) (P < .05). Furthermore, the mean protein concentration (2.12 mg/mL) in JL from joints with effusion was significantly higher than in joints without joint effusion (1.27 mg/mL) (P < .05). CONCLUSIONS: These data demonstrate that painful joints are more likely to show joint effusion on MRI, and the protein levels in JL recovered from these joints is higher than in pain-free joints. These data also suggested that joint effusion may be related to the inflammatory changes seen in patients with ID and OA. PMID- 10513865 TI - The effect of preoperative ondansetron on the incidence of postoperative nausea and vomiting in patients undergoing outpatient dentoalveolar surgery and general anesthesia. AB - PURPOSE: The purpose of this investigation was to evaluate the efficacy of ondansetron in controlling postoperative nausea and vomiting (PONV) when used prophylactically in patients undergoing routine dentoalveolar surgery performed under general anesthesia. PATIENT AND METHODS: This was a prospective, double blind, randomized, placebo-controlled evaluation. Fifty adult ASA I or II patients, requiring routine dentoalveolar surgery performed under general anesthesia, without a prior history of PONV, were randomly assigned to the experimental or control groups. Ondansetron (2.0 mL = 4.0 mg) or normal saline (2.0 mL) were administered intravenously before surgery. Age, gender, type of surgery, duration of surgery, anesthetic dosages, and PONV were evaluated. PONV was evaluated at time 0 (end of anesthesia) and at 30 and 60 minutes postoperatively. Nausea was evaluated using a visual analog scale (1, not nauseous; 5, about to vomit). Vomiting was assessed as a yes or no response. At 20 to 28 hours postoperatively, PONV was evaluated via a telephone call as a yes or no response, along with the number of episodes of nausea, vomiting, or both. Means and standard deviations were calculated for age, surgery, and anesthetics, and differences were assessed using an independent samples t-test. Differences for gender between the control and experimental groups were tested by a nonparametric chi-squared test. Differences between groups for nausea and vomiting were tested with a continuity correction chi-squared test. Differences were considered significant for a P < .05. RESULTS: No significant differences (P < .05) were found between the PONV groups for gender, duration of procedure, or anesthetic dosages. Statistically significant differences were noted in age and the type of surgical procedures performed. No statistically significant differences (P < .05) were noted between groups for nausea or vomiting. CONCLUSION: Based on the results of this study, PONV occurred in approximately 20% of patients (20% for nausea, 8% for vomiting). With the types of anesthetic agents and techniques used in this investigation, there were no significant differences between ondansetron and placebo for prophylaxis against PONV. PMID- 10513866 TI - Determination of the anti-inflammatory effects of methylprednisolone on the sequelae of third molar surgery. AB - PURPOSE: The anti-inflammatory effect and adrenal suppressive side effect of methylprednisolone sodium succinate (MP) on the postoperative sequelae of third molar surgery were evaluated using objective methods in a double-blind, crossover study. PATIENTS AND METHODS: Twenty patients who were to undergo surgical removal of bilateral, symmetrically placed lower third molars were studied. Each patient was given 125 mg MP intravenously before surgery on one side, and a placebo before surgery on the opposite side on a random basis. Ultrasonographic and computed tomographic examinations were performed to determine the amount of facial edema. Trismus was evaluated by measuring maximal interincisal opening, and pain was evaluated by recording the number of standard analgesic tablets used on the day of surgery and the first postoperative day. Hypothalamic-pituitary adrenal (HPA) axis function was tested by measuring basal plasma cortisol (hydrocortisone) levels preoperatively and postoperatively. The adrenocorticotropic hormone (ACTH) stimulation test also was performed before and after administration of MP, to evaluate adrenal function. RESULTS: Statistical analysis of the data indicated a significant decrease in edema, trismus, and pain in the MP group. Plasma cortisol levels showed a nonsignificant decrease in both the MP- and placebo-treated groups. The ACTH stimulation test indicated normal HPA axis function before and after MP administration. No clinically apparent infection, disturbance of wound healing, or other corticosteroid-related complications were noted. Eighteen patients (90%) indicated a preference for the overall postoperative course when MP was used. CONCLUSION: In the absence of contraindications for corticosteroid administration, preoperative use of MP appears to be a safe and effective method of reducing postoperative complications in third molar surgery. PMID- 10513867 TI - Computed tomographic evaluation of bone formation after secondary bone grafting of alveolar clefts. AB - PURPOSE: The purpose of this article was to evaluate the initial volume of alveolar clefts and the bone bridges formed in patients who had undergone secondary bone grafting. PATIENTS AND METHODS: Fifteen cleft patients were studied. Computed tomography (CT) scans were made at 2-mm slice sections immediately before operation and at 3 months and 1 year postoperatively. The volume of the alveolar clefts and bone bridges was calculated from stored images by using an image scanner and a personal computer. RESULTS: The mean cleft volume of the patients was 1.1 +/- 0.3 cm3. The corresponding volume of the bone bridges 3 months and 1 year postoperatively were 1.2 +/- 0.6 cm3 and 1.1 +/- 0.5 cm3, respectively. The value at 1 year was significantly decreased compared with that at 3 months. CONCLUSIONS: Adequate bone bridging was maintained for 1 year. However, it is suggested that the grafted site should be restored with a functioning tooth or a dental implant to place it under physiologic stress, because the volume of the bone bridges tends to decrease from 3 months to 1 year. PMID- 10513868 TI - The effect of sagittal orientation of the distractor on the biomechanics of mandibular lengthening. AB - PURPOSE: It has been previously demonstrated that distractors placed parallel to the mandible in the transverse plane, without regard to the vector of distraction create lateral displacement tendencies at the appliance-bone interface, leading to potential clinical problems and complications. The purpose of this study was to evaluate the biomechanical effects of linear distractor orientation in the sagittal plane relative to the anatomic axis of the mandible (mandibular plane) and the maxillary occlusal plane. MATERIALS AND METHODS: A 2-dimensional model of the human mandible was generated for computer simulation of osteodistraction. Positional changes of the distal mandibular segment were then analyzed during 10 mm of incremental lengthening based on distractor orientation relative to the maxillary occlusal plane. RESULTS: Distractors placed parallel to the inferior border of the mandible without regard to the maxillary occlusal plane created a vertical translation of the distal bone segment resulting in an anterior open bite. The magnitude of the anterior open bite was proportional to the angle between the vector of distraction and the maxillary occlusal plane, and to the amount of distraction. Placement of the distractors parallel to the maxillary occlusal plane eliminated the tendency for an anterior open bite. CONCLUSIONS: The orientation of the distractors relative to the maxillary occlusal plane is one of the important biomechanical parameters that must be included in preoperative planning for mandibular osteodistraction. PMID- 10513869 TI - Radiodensitometric study of resorption in the site of mandibular fractures. AB - PURPOSE: The area of bone resorption resulting from a fracture varies widely. This study was performed to determine the magnitude of this area in mandibular fractures by means of quantitative radiodensitometry and to determine the influence of the kind of treatment applied, location of fracture, and the age and sex of the patient on this zone. PATIENTS AND METHODS: In successive panoramic radiographs after mandibular fracture (postoperatively and at 15, 30, 60, and 90 days), optical density changes occurring in the region of the fracture were analyzed. An area of 2 cm on each side of the fracture was studied, divided into 31 concentric regions of equal size (4 pixels thick). RESULTS: The magnitude of the area of necrosis in mandibular fractures is 7.7 mm in cases treated by maxillomandibular fixation and 5.8 mm in those cases treated with rigid internal fixation. CONCLUSIONS: The area of resorption in mandibular fractures is determined, in part, by the type of treatment used and the location of the fracture. PMID- 10513870 TI - Aesthetic uses of botulinum toxin A. PMID- 10513871 TI - Anterior maxillary lesion. PMID- 10513872 TI - Practice differences between male and female oral and maxillofacial surgeons: survey results and analysis. AB - PURPOSE: This study describes the personal and practice characteristics of oral and maxillofacial surgeons, with an emphasis on gender differences. Potential explanations for differences found are offered. MATERIALS AND METHODS: A 39-item questionnaire was designed to address areas of suspected differences between male and female oral and maxillofacial surgeons. It included items regarding training, certification, practice type and location, time spent working, practice composition, and personal characteristics. Identical questionnaires were sent to all of the 130 female oral and maxillofacial surgeons (OMSs) registered with AAOMS, as well as to 264 randomly sampled male OMSs. RESULTS: Of the 192 responses received, 109 were from male surgeons, and 83 were from females, 56.8% versus 43.2% of the total. For men, the response rate was 41.3%, whereas the rate was 68.3% for the women; overall, 48.7% of the 394 oral surgeons responded. Profiles of the "average" male and female OMS showed similarities with regard to race, training, and reasons for choosing an OMS career. Age, marital status, number of children, physical characteristics, and use of time outside of work indicated some significant personal differences, as did practice characteristics, including income, number of years in practice, hours spent working, and number of patients seen. Clear gender-based trends were found regarding opinions of physical and mechanical disadvantages, with women more strongly denying such problems. A variety of both positive and negative viewpoints were elicited by the free comment section. CONCLUSIONS: Although male and female OMSs are similar with regard to most variables investigated, some significant differences do exist. PMID- 10513873 TI - Herpes zoster infection of the maxilla: case report. PMID- 10513874 TI - Simultaneous occurrence of segmental odontomaxillary dysplasia and Becker's nevus. PMID- 10513875 TI - Simultaneous metastatic ameloblastoma and thyroid carcinoma in the cervical region: report of a case. PMID- 10513876 TI - Combined central odontogenic fibroma and giant cell granuloma-like lesion of the mandible: report of a case and review of the literature. PMID- 10513877 TI - Respiratory mucocele formation after augmentation genioplasty with nasal osteocartilaginous graft. PMID- 10513878 TI - Odontogenic ghost cell carcinoma: report of a case. PMID- 10513879 TI - Chondroblastoma of the temporomandibular region. PMID- 10513880 TI - Proper management of postsurgical epistaxis. PMID- 10513881 TI - Nocturnal asthma: role of circadian rhythms in its mechanisms and therapy. PMID- 10513882 TI - Clinical chronobiology and chronotherapeutics with applications to asthma. AB - The concept of homeostasis (i.e., constancy of the milieu interne) has long dominated the teaching and practice of medicine. Concepts and findings from chronobiology, the scientific study of biological rhythms, challenge this construct. Biological processes and functions are not at all constant; rather, they are organized in time as rhythms with period lengths that range in duration from as short as a second or less to as long as a year. It is the body's circadian (24 h) rhythms that have been researched most intensely. The peak and trough of these rhythms are ordered rather precisely in time to support the biological requirements of activity during the day and sleep at night. The timing of the peak and trough plus the magnitude of variation (amplitude) of physiological and biochemical functions during the 24 h give rise to predictable in-time, day-night patterns in the manifestation and exacerbation of many common medical conditions. Circadian rhythms also can influence the response of patients to diagnostic tests and therapeutic interventions according to their timing with reference to body rhythms. Rhythms in the pathophysiology of medical conditions and patient tolerance to medications constitute the basis for chronotherapeutics, the timing of treatment in relation to biological rhythm determinants as a means of optimizing beneficial effects and safety. The article discusses recent advances in medical chronobiology and chronotherapeutics and their relevance to clinical medicine in general and the management of asthma in particular. Indeed, since asthma is a disease that exhibits rather profound circadian rhythmicity, investigation of its pathophysiology and therapy necessitates a chronobiologic approach. PMID- 10513883 TI - Sleep, respiratory physiology, and nocturnal asthma. AB - The nocturnal worsening of asthma is a common feature of this disease that recently has received extensive investigation. Most recent efforts have focused on the role of circadian biorhythms that could promote a nocturnal increase in airway inflammation, leading to a subsequent increase in airflow obstruction and asthma symptoms. However, definitive studies remain lacking. As discussed in this review, there is also substantial evidence that sleep itself may play a direct role in the nocturnal worsening of asthma. Potential mechanisms for such a sleep related effect could include the supine posture, alterations in sympathetic and parasympathetic "balance," sleep-associated reductions in lung volume, intrapulmonary pooling of blood, and sleep-associated upper airway narrowing, both with and without snoring and obstructive sleep apnea (OSA). These potential contributors to this troublesome phenomenon deserve further consideration when investigating mechanisms of nocturnal asthma. PMID- 10513884 TI - Biologic rhythms in the immune system. AB - In all of its components, the immune system shows regularly recurring, rhythmic variations in numerous frequencies; the circadian (about 24 h) rhythms are the best explored. The circadian variations in immunocompetent cells circulating in the peripheral blood are of a magnitude to require attention in medical diagnostics. Both the humoral arm and the delayed (cellular) arm of the immune system function in a rhythmic manner. The response of the immune system to introduction of an antigen and to challenge of the sensitized organism varies in extent in the circadian frequency range and also in lower frequencies, for example, of about a week (circaseptan) or seasonally (circannual). The medical application of the biologic rhythms of the immune system extends to diagnostic measures, as well as treatment. PMID- 10513885 TI - Location of airway inflammation in asthma and the relationship to circadian change in lung function. AB - Nocturnal asthma is an important part of asthma as the majority of patients with asthma have nocturnal worsening in lung function. The etiology of this process is multifactorial and interactive. There are many naturally occurring circadian rhythms, which for the normal individual have only a minor effect on lung function. However, in the asthmatic patient, these day-to-night alterations produce increased airway inflammation and worsening of asthma. Although asthma is considered an airway disease, the location of the inflammatory response may be greater in the alveolar tissue area. If correct, this could alter the therapeutic approach to this disease. PMID- 10513886 TI - Circadian variation in allergen and nonspecific bronchial responsiveness in asthma. AB - In a majority of patients, exacerbations of asthma occur more frequently during the night than day. Many hypotheses have been proposed to explain such variation in asthma. The airways of asthmatic persons are characterized by an abnormal degree of inflammation and bronchial hyperresponsiveness to both nonspecific and specific challenges. Studies of both children and adults with asthma document marked circadian rhythmicity in the response of airways to bronchial challenges with histamine, methacholine, acetylcholine, saline, and house dust mite. Taken together, the findings of these investigations indicate that the hyperreactivity of airways to these agents is more profound and prolonged following evening and overnight tests compared to tests conducted in the midday and afternoon. The temporal pattern in bronchial response to the hyperventilation of cold dry air is different. The hyperresponsiveness of airways to this challenge is greatest in the afternoon. The amplitude of the circadian rhythm in airway hyperreactivity seems to be correlated to the amplitude of the circadian rhythm of pulmonary function; the greater the day-night difference in bronchial reactivity is, the greater is the day-night difference in flow rates. PMID- 10513887 TI - Nocturnal asthma: role of nocturnal gastroesophageal reflux. AB - Gastroesophageal reflux (GER) is common in those with asthma, with 77% of asthmatics complaining of heartburn, with 41% experiencing reflux-associated respiratory symptoms. Likewise, 24% of those with asthma that is difficult to control have "clinically silent" GER. There are no studies examining nocturnal reflux symptoms in asthmatics. Esophageal dysmotility is also common, and abnormal esophageal acid contact times on 24h esophageal pH tests were found in 82% of asthmatics examined consecutively. Most asthmatics with GER also have abnormal esophageal acid contact times while in the supine position, reflecting sleep time. Endoscopic evidence of esophagitis was found in 43% of asthmatics. Two mechanisms of bronchoconstriction induced by esophageal acid have been proposed: a vagally mediated reflex, by which esophageal acid in the distal esophagus causes reflex bronchoconstriction, and microaspiration. Although there is conflicting evidence, distal esophageal acid causes a decrease in peak expiratory flow rates, an increase in respiratory resistance, and an increase in minute ventilation. If microaspiration is present, there is further augmentation of this airway response. Although only a few studies have been performed in those with nocturnal asthma with GER, one study in a pediatric population showed that esophageal acid infusions caused more airway responses at 04:00 than at 24:00. Also, asthmatic children with nocturnal asthma symptoms have a higher reflux score, with a positive correlation between reflux score and nighttime-associated wheezing. Despite these findings in children, a study performed in sleeping adults with nocturnal asthma noted no alterations in airflow resistance with esophageal acid, concluding that GER contributed little to the nocturnal worsening of asthma. There are also gastroesophageal circadian issues that may influence GER in asthmatics. Gastric acid secretion peaks at approximately 21:00, and gastric emptying is delayed when a meal is given at 20:00 versus 08:00. Esophageal acid clearance is delayed significantly during sleep, and acid clearance occurs during arousals. Upper esophageal sphincter (UES) pressure also decreases with sleep onset, which may predispose to microaspiration. Further research is needed to clarify what role nocturnal reflux has on nocturnal asthma and airway inflammation and whether circadian rhythm factors alter airway responses to esophageal acid. PMID- 10513888 TI - Circadian rhythms in the pharmacokinetics and clinical effects of beta-agonist, theophylline, and anticholinergic medications in the treatment of nocturnal asthma. AB - Published asthma consensus reports now acknowledge that asthma is a nocturnal disease in as many as 75% of those afflicted by this medical condition. Nonetheless, the treatment of this chronic obstructive pulmonary disease in the clinic continues to be based primarily on homeostatic considerations in that it relies on long-acting bronchodilator and other therapies formulated and scheduled to ensure constant or near-constant levels of medication during the 24h. The need of asthma patients prone to nighttime attacks is not the same during the day and night; the therapeutic requirements of patients who experience nocturnal asthma, especially ones with the more severe forms of the disease, are often not satisfied by conventional medications. The therapeutic response and patient tolerance to bronchodilator medications can be improved markedly when the medications are proportioned during the 24h as a chronotherapy, that is, when more medication is delivered during nighttime sleep than daytime activity, as verified by numerous studies. This article reviews how the body's circadian rhythms influence the pharmacokinetics and effects of commonly prescribed asthma therapies and addresses why and how they must be taken into consideration to increase the effectiveness of asthma treatment. PMID- 10513889 TI - Corticosteroids and leukotrienes: chronobiology and chronotherapy. AB - Corticosteroids and leukotrienes play opposite roles in asthma. Corticosteroids, both endogenously secreted and exogenously administered, are antiinflammatory and are very effective in the treatment of asthma. They have also been evaluated chronotherapeutically and have been found to be very effective in reducing the enhanced airway inflammation and decrement in lung function associated with nocturnal worsening of asthma. Leukotrienes are potent proinflammatory and spasmogenic mediators that have been shown to be increased at night in patients with nocturnal asthma (NA). Leukotriene modifiers, a new class of medications to treat asthma, improve, but do not abolish, the symptoms and decrement in lung function associated with nocturnal asthma. However, they have not been evaluated chronotherapeutically. This article addresses the roles of corticosteroids and leukotrienes in nocturnal asthma and their position as therapeutic agents or targets for therapy. PMID- 10513890 TI - Neuropsychological outcomes of nocturnal asthma. AB - In spite of frequent reports that nocturnal asthma results in fatigue and impaired cognitive performance, there exists little objective evidence as to the daytime consequences of this disorder. Treatment studies have established that the symptoms of nocturnal asthma improve with medication intervention, but performance does not. Studies of obstructive sleep apnea (OSA), a source of generally more severe sleep fragmentation, have demonstrated that measurement of sleep-deprivation effects is limited to tasks requiring heightened alertness and rapid information processing, and that the degree of score change is related to the degree of sleep disruption. Studies of normal, but sleep-deprived, subjects indicate that (1) utilization of repetitive measures sustained for long duration can potentiate motivation to overcome the effects of fatigue in the laboratory, and (2) even when average scores do not change significantly, performance becomes more irregular. These collective findings about the measurement of performance impairment secondary to sleep deprivation can be used to guide new studies of nocturnal asthma. Finally, children must be included in future investigations because they may be at even greater risk for daytime consequences of nocturnal asthma than adults. PMID- 10513891 TI - Discovery of some of the biological effects of nitric oxide and its role in cell signaling. AB - The role of nitric oxide in cellular signaling in the past 22 years has become one of the most rapidly growing areas in biology with more than 20,000 publications to date. Nitric oxide is a gas and free radical with an unshared electron that can regulate an ever-growing list of biological processes. In many instances nitric oxide mediates its biological effects by activating guanylyl cyclase and increasing cyclic GMP synthesis from GTP. However, the list of effects of nitric oxide that are independent of cyclic GMP is also growing at a rapid rate. For example, nitric oxide can interact with transition metals such as iron, thiol groups, other free radicals, oxygen, superoxide anion, unsaturated fatty acids and other molecules. Some of these reactions result in the oxidation of nitric oxide to nitrite and nitrate to terminate its effect, while other reactions can lead to altered protein structure, function, and/or catalytic capacity. These diverse effects of nitric oxide that are either cyclic GMP dependent or independent can alter and regulate important physiological and biochemical events in cell regulation and function. Nitric oxide can function as an intracellular messenger, an autacoid, a paracrine substance, a neurotransmitter, or as a hormone that can be carried to distant sites for effects. Thus, it is a unique simple molecule with an array of signaling functions. However, as with any messenger molecule, there can be too little or too much of the substance and pathological events result. Some of the methods to regulate either nitric oxide formation, metabolism, or function have been in clinical use for more than a century as with the use of organic nitrates and nitroglycerin in angina pectoris that was initiated in the 1870's. Current and future research with nitric oxide and cyclic GMP will undoubtedly expand the clinicians' therapeutic armamentarium to manage a number of important diseases by perturbing nitric oxide and cyclic GMP formation and metabolism. Such promise and expectations have obviously fueled the interests in these signaling molecules for a growing list of potential therapeutic applications. PMID- 10513892 TI - Lysosomal alpha-D-mannosidase. AB - Alpha-mannosidosis in the human is an autosomal recessive lysosomal storage disease caused by a deficiency of lysosomal alpha-D-mannosidasea activity. Lysosomal alpha-D-mannosidase is involved in the catabolism of N-linked glycoproteins through the sequential degradation of high-mannose, hybrid and complex oligosaccharides. This review is focused on human, mouse, bovine and feline genes coding for lysosomal alpha-D-mannosidase. In particular the exon intron structure of the genes, their promoters, and the identification of mutations causing the disease have been examined. The construction, by homologous recombination, of a mouse model of alpha-mannosidosis is reported. PMID- 10513893 TI - The structural basis for the susceptibility of gangliosides to enzymatic degradation. AB - The conformational properties of GM2, GalNacbeta-4(Neu5Acalpha-3) Galbeta 4Glcbeta-1Cer have been compared to those of 6'-GM2, in which the linkage between the GalNAc and Gal was altered from GalNacbeta-4Galbeta- to GalNacbeta-6Galbeta-, and to those of GD1a, Neu5Acalpha-3Galbeta-3GalNAcbeta-4(Neu5Acalpha-3 )Galbeta 4Glcbeta-1Cer, and GalNAc-GD1a. Our results revealed that unlike the compact and rigid oligosaccharide head group found in GM2, where the Neu5Ac and the GalNAc residues interact, the sugar chain of 6'-GM2 is in an open spatial arrangement, with the Neu5Ac no longer interacting with GalNAc, freely accessible to external interactions. The structure of GD1a can be regarded as that of GM2 with an extension of the terminal Neu5Acalpha-3Galbeta-disaccharide. The inner portion of GD1a is that of GM2 comprising the very rigid GalNAc-[Neu5Ac-]Gal trisaccharide. The terminal Neu5Ac-Gal linkage is flexible and fluctuates between two limiting conformations. In GalNAc-GD1a the outer sialic acid gains conformational rigidity due to the presence of the outer GalNAc in position 4 of galactose. This ganglioside has two 'core' GalNAc-[Neu5Ac-]Gal trisaccharide linked in tandem. PMID- 10513894 TI - Mannosylphosphodolichol synthase activity is associated with a 32 kDa phosphoprotein. AB - Failure of actinomycin D to block the activation of Dol-P-Man synthase in isoproterenol-treated capillary endothelial cells, supported that isoproterenol effect was not mediated by active transcription of the Dol-P-Man synthase gene during a short-term beta-adrenoreceptor stimulation. Instead, it was a net effect of protein phosphorylation by cAMP-dependent protein kinase. Using antibody as a probe we have now demonstrated that Dol-P-Man synthase activity is associated with a 32 kDa ER phosphoprotein. PMID- 10513895 TI - Guanylyl cyclase C receptor: regulation of catalytic activity by ATP. AB - Guanylyl cyclase C (GCC), a member of the family of membrane bound guanylyl cyclases is the receptor for the heat-stable enterotoxin (ST) peptides and the guanylin family of endogenous peptides. GCC is activated upon ligand binding to increase intracellular cGMP levels, which in turn activates other downstream signalling events in the cell. GCC is also activated in vitro by nonionic detergents. We have used the T84 cell line as a model system to investigate the regulation of GCC activity by ATP. Ligand-stimulated GCC activity is potentiated in the presence of ATP, whereas detergent-stimulated activity is inhibited. The potentiation of GCC activity by ATP is dependent on the presence of Mg2+ ions, and is probably brought about by a direct binding of Mg-ATP to GCC. The protein kinase-like domain of GCC, which has earlier been shown to play a critical role in the regulation of GCC activity, may be a possible site for the binding of Mg ATP to GCC. PMID- 10513896 TI - Interaction of newly synthesized apolipoprotein B with calnexin and calreticulin requires glucose trimming in the endoplasmic reticulum. AB - Apolipoprotein B (ApoB) is the only protein component of the low density lipoproteins (LDL) in plasma. It is a glycoprotein with a molecular mass of about 550 kDa (4536 amino acids) containing 16 N-glycans. We have studied the interaction of ApoB with two lectin-like chaperones of the Endoplasmic Reticulum (ER)--Calnexin (CN) and Calreticulin (CR). Using a co-immunoprecipitation approach we observed that newly synthesized ApoB associates with CN and CR. The interaction was transient; within 30-60 min after synthesis bulk of newly formed ApoB dissociated. Using McA Rh7777 cells expressing an N-terminal fragment of ApoB we found that inhibition of glucosidases in the ER prevented the association of CN and CR to newly synthesized ApoB. The results showed that like for association with other glycoprotein substrates, trimming of glucose residues was essential for ApoB binding to CN and CR. PMID- 10513898 TI - Molecular basis of the polydispersity of mucins: implications for the generation of saccharide diversity. AB - Secreted epithelial mucins are large macromolecules which exhibit extreme polydispersity, the molecular basis of which is not fully understood. We have obtained partial sequences of two genes (BSM1 and BSM2) coding for two distinct molecules. This is the first time that such closely-related genes have been identified for any mucin from an animal. We propose that a combination of multiple homologous genes, alternative splicing, differential glycosylation, and additional post-translational processing all contribute to the extreme polydispersity of mucins. The multiple domain structure and non-identical tandem repeats are also very important for the generation of the saccharide diversities of mucins. PMID- 10513897 TI - Glycosphingolipid domains on cell plasma membrane. AB - In this study we analyzed by immunofluorescence, laser confocal microscopy, immunoelectron microscopy and label fracture technique the ganglioside distribution on the plasma membrane of several different cell types: human peripheral blood lymphocytes (PBL), Molt-4 lymphoid cells, and NIH 3T3 fibroblasts, which mainly express monosialoganglioside GM3, and murine NS20Y neuroblastoma cells, which have been shown to express a high amount of monosialoganglioside GM2. Our observations showed an uneven distribution of both GM3 and GM2 on the plasma membrane of all cells, confirming the existence of ganglioside-enriched microdomains on the cell surface. Interestingly, in lymphoid cells the clustered immunolabeling appeared localized over both the microvillous and the nonvillous portions of the membrane. Similarly, in cells growing in monolayer, the clusters were distributed on both central and peripheral regions of the cell surface. Therefore, glycosphingolipid clusters do not appear confined to specific areas of the plasma membrane, implying general functions of these domains, which, as structural components of a cell membrane multimolecular signaling complex, may be involved in cell activation and adhesion, signal transduction and, when associated to caveolae, in endocytosis of specific molecules. PMID- 10513899 TI - Carbohydrate analysis of bradyrhizobial (NC92) lipopolysaccharides by high performance-anion exchange chromatography with pulsed amperometric detection. AB - Composition analysis of monosaccharides of Sepharose 4B purified NC 92 LPS and the polysaccharides fractions from Sephadex G-50 chromatography was performed by high performance anion exchange chromatography using pulsed amperometric detection. Rhamnose, mannose, galactose and glucose are present in a substantial amount in the purified LPS (Pk I). High molecular weight purified polysaccharides (PS I) obtained after sephadex gel filtration of the purified LPS (Pk I) acid hydrolysate showed an increase in glucose:galactose ratio. This indicates the presence of the peanut root lectin (PRA II) specific sugar in higher proportion on the O-antigen part of the LPS molecule, which may aid in the critical recognition reaction. PMID- 10513900 TI - Expression, purification and characterization of peanut (Arachis hypogaea) agglutinin (PNA) from baculovirus infected insect cells. AB - Peanut (Arachis hypogaea) seed lectin, PNA is widely used to identify tumor specific antigen (T-antigen), Galbeta1-3GalNAc on the eukaryotic cell surface. The functional amino acid coding region of a cDNA clone, pBSH-PN was PCR amplified and cloned downstream of the polyhedrin promoter in the Autographa californica nucleopolyhedrovirus (AcNPV) based transfer vector pVL1393. Co transfection of Spodoptera frugiperda cells (Sf9) with the transfer vector, pAcPNA and AcRP6 (a recombinant AcNPV having B-gal downstream of the polyhedrin promoter) DNAs produced a recombinant virus, AcPNA which expresses PNA. Infection of suspension culture of Sf9 cells with plaque purified AcPNA produced as much as 9.8 mg PNA per liter (2.0 x 10(6) cells/ml) of serum-free medium. Intracellularly expressed protein (re-PNA) was purified to apparent homogeneity by affinity chromatography using ECD-Sepharose. Polyclonal antibodies against natural PNA (n PNA) cross-reacted with re-PNA. The subunit molecular weight (30 kDa), hemagglutination activity, and carbohydrate specificity of re-PNA were found to be identical to that of n-PNA, thus confirming the abundant production of a functionally active protein in the baculovirus expression system. PMID- 10513901 TI - Outcomes of brachial plexus reconstruction in 204 patients with devastating paralysis. AB - Thus far, devastating injuries of the adult brachial plexus have had a poor prognosis. This article presents the possible outcomes of aggressive microsurgical reconstruction in the largest series of patients in North America to date. It should change the pessimistic outlook that has surrounded these lesions. In this study, the outcomes of surgery were analyzed in relation to the type and level of injury, the age of the patient, and the denervation time; stronger donors for neurotization in relation to the various targets were delineated. The results were analyzed in 204 patients with adequate follow-up from a total of 263 patients who were operated on between 1978 and 1996. The mean age of the patients was 25.9 years, and the injuries were caused by high-velocity motor accidents involving avulsion in 55 percent of the patients. Nerve reconstruction included 577 nerve repairs (140 direct neurotizations and 437 cases of nerve grafting). Microneurolysis was performed in 89 cases. Vascularized nerve grafts were used in 120 repairs. Muscle transfers (29 pedicled and 78 free) were used to enhance function. The results were good or excellent in 75 percent of suprascapular nerve reconstructions, 40 percent of deltoid reconstructions, 48 percent of biceps reconstructions, 30 percent of triceps reconstructions, 35 percent of finger-flexion reconstructions, and 15 percent of finger-extension reconstructions. The majority of the patients had protective sensation and pain relief postoperatively. PMID- 10513902 TI - Psychological complications in 281 plastic surgery practices. AB - Surgery is a high-stakes stressor with possible consequences that include death, pain, disfigurement, economic losses, and alterations in social roles. Often, the most disturbing complications to surgeons and patients are psychological rather than physical. Ineffective management of psychological complications of surgery can have profound consequences, resulting in delayed recuperative times, delayed return to work, poor patient compliance, dissatisfaction with the surgical outcome, hostility toward surgeons, and anxiety. The purpose of this study was to investigate in a large randomized group of plastic surgery practices the relative incidence of negative psychological outcomes and to compare these with the incidence of adverse physical outcomes to gain a greater appreciation of the relative magnitude of each type of perioperative complication. The study design was a descriptive, correlational survey that assessed psychological complications reported by plastic surgeons. The Plastic Surgery Questionnaire was sent to 702 randomly selected board-certified plastic surgeons. The sample consisted of 281 board-certified plastic surgeons (40 percent response rate). The study instrument was found to be highly reliable, with inter-item Cronbach's alpha r = 0.85. The demographics were representative of the specialty as a whole. It was found in general that psychological complications were much more prevalent than physical problems such as hematoma or infection. Anxiety reactions were commonly encountered by 95.4 percent of surgeons; disappointment (96.8 percent), depression (95.0 percent), nonspecific physical complaints (92.2 percent), and sleep disorders (88.5 percent) were the next most commonly reported complications. Most surgeons (75.8 percent) reported that screening for depression was important, but only 18.8 percent identified screening for post traumatic stress disorder as important, even though 86 percent had diagnosed post traumatic stress disorder in their postoperative patients. Psychological complications occur at rates equal to or greater than those of physical complications in the plastic surgery practice. Patients who experience physical complications are much more likely to simultaneously experience psychological complications. Patients with preexisting psychological conditions are more at risk for postoperative psychological complications. Disappointment, anxiety, and depression were the most frequently seen psychological complications. Nursing personnel are perceived by plastic surgeons to have the primary role in screening patients for pertinent psychological history. Directed research should be undertaken to determine which treatment regimens are most effective in reducing preoperative psychological complications. Controlled clinical trials of pharmaceuticals and alternative therapies must be designed and carried out in a prospective manner to establish the optimum treatment for alleviation of adverse emotional consequences of surgery. The next frontier for the specialty is to actively and consciously investigate and improve our patients' emotional and psychological results from surgery. PMID- 10513903 TI - The principle of rotation advancement for repair of unilateral complete cleft lip and nasal deformity: technical variations and analysis of results. AB - This is an assessment of one surgeon's 15-year experience (1981-1995) using the Millard rotation-advancement principle for repair of unilateral complete cleft lip and nasal deformity. All infants underwent a prior labio-nasal adhesion. Since 1991, dentofacial orthopedics with a pin-retained (Latham) appliance was used for infants with a cleft of the lip and palate. Technical variations are described, including modifications in sequence of closure. A high rotation and releasing incision in the columella lengthens the medial labial element and produces a symmetric prolabium with minimal transgression of the upper philtral column by the advancement flap. Orbicularis oris muscle is everted, from caudad to cephalad, to form the philtral ridge. A minor variation of unilimb Z-plasty is used to level the cleft side of Cupid's bow handle, and cutaneous closure proceeds superiorly from this junction. The dislocated alar cartilage is visualized though a nostril rim incision and suspended to the ipsilateral upper lateral cartilage. Symmetry of the alar base is addressed in three dimensions, including maneuvers to position the deviated anterior-caudal septum, configure the sill, and efface the lateral vestibular web. Secondary procedures were analyzed in 105 consecutive patients, both revised (n = 30) and unrevised (n = .75). The possible need for revision in the latter group was determined by panel assessment of six indicators of nasolabial asymmetry, documented by frontal and submental photographs. In the entire study period, a total of 80 percent of children required or will need nasal revision, and a total of 42 percent required or will require labial revision. In the last 5 years, as compared with the earlier decade, there was a significantly diminished incidence of patients requiring labial revision (54 percent to 21 percent) and alar suspension (63 percent to 32 percent). These improvements are attributable to technical refinements and experience, although dentofacial orthopedics may also have played a role. PMID- 10513904 TI - Children with repaired bilateral cleft lip/palate: effect of age at premaxillary osteotomy on facial growth. AB - This study compared facial growth in three groups of patients with bilateral complete cleft lip/palate: those who had (1) no premaxillary osteotomy, (2) premaxillary osteotomy before age 8 years, and (3) premaxillary osteotomy after age 8 years. Of 24 children with bilateral complete cleft lip/palate, 7 had early premaxillary osteotomy (mean age, 6.1; range, 3.7 to 7.6 years), 10 had late osteotomy (mean age, 11.2; range, 8.3 to 20.7 years), and 7 did not require premaxillary repositioning and served as controls (mean age, 12.4; range, 6.4 to 17.8 years). Presurgical and postsurgical lateral cephalograms were digitized using the Dentofacial Planner software; most current lateral cephalograms comprised the control group. Forty-one bony and 25 soft-tissue landmarks were digitized, and 8 angles were measured: SNA, (sella-nasion-A point), SNPg (sella nasion-pogonion), ANB (A point-nasion-B point), NAPg (nasion-A point-pogonion), ST convexity (glabella-subnasale-soft-tissue pogonion), Sn-G vertical (line perpendicular to the horizontal plane dropped from glabella and distance measured from subnasale to this vertical), Cm-Sn-Ls (columella-subnasale-abial superioris), and Sn-Gn-C (subnasale-soft-tissue gnathion-chin point). Statistical difference in mean preoperative and postoperative values were measured with analysis of variance. Tests of significance were adjusted for multiple comparisons using the Bonferroni correction. Mean age at follow-up for early, late, and control groups was 11.8, 14.0, and 12.4 years, respectively. Mean follow-up for early and late groups was 5.7 and 2.8 years. There was a significant preoperative difference among the three groups for mean SNA (p < 0.01), ANB (p < 0.01), and NAPg (p < 0.01). Bonferroni analyses revealed that the early group had significantly greater SNA, ANB, and NAPg angles than the late (p < 0.01) and control groups (p < 0.05). There was a significant postoperative difference among groups for ANB (p < 0.05); Bonferroni analyses also showed that the control group had a significantly greater ANB than the late group (p < 0.05). The t test for equity of means established postoperative change for SNA (p < 0.01), ANB (p< 0.01), NAPg (p < 0.01), and ST convexity (p < 0.01) for the early group was significantly greater than for the late group. Children who required early premaxillary positioning had more significant preoperative deformity; however, this group's postoperative profile was not, on average, significantly different from either the late or control groups. Our findings that the early group had more significant change with premaxillary osteotomy than the late group suggest that premaxillary positioning can be done before completion of facial growth without compromise. PMID- 10513906 TI - Distribution of the superficial temporal artery in the Chinese adult. AB - The superficial temporal artery is important in head and neck surgery. Ethnologic variation may affect surgical procedure. In this study, we evaluated the variations of the artery in Chinese adults. We measured its bifurcating location, the diameter of its vessels, and its relationship to nearby structures. A total of 26 cadavers with 52 superficial temporal arteries were examined in 3 consecutive years. The superficial temporal artery ran 1.14 cm anteriorly to the bony external auditory canal. The average diameters of the superficial temporal artery, frontal branch, and parietal branch were 2.14, 1.61, and 1.68 mm, respectively. In 45 of 52 cases (86.5 percent), bifurcation of the artery occurred well above the zygomatic arch. The present study thus demonstrated that the superficial temporal artery in the Chinese adult differs from that in the Caucasian and has provided a detailed anatomic distribution analysis of the superficial temporal artery in Chinese adults, which should benefit the clinician in dealing with operation procedures related to this artery. PMID- 10513905 TI - The use of hydroxyapatite cement in secondary craniofacial reconstruction. AB - Sixty-one patients underwent secondary craniofacial reconstruction for contour defects using hydroxyapatite cement over a 3-year period (20-month mean follow up). There were 56 children, aged 2.2 to 18 years (mean, 10.7 years), 21 boys and 35 girls. This is the first series of pediatric patients in whom the use of hydroxyapatite cement has been reported. There were five adults aged 21 to 46 years (mean, 32 years), 3 men and 2 women. Thirty-one patients underwent reconstruction for secondary orbitocranial defects after surgery for synostosis, 7 after surgery for hypertelorism, 10 for posttraumatic skull defects, and 13 for a variety of other facial skeletal defects. There were seven complications (11 percent), ranging from a retained drain to postoperative seromas, all of which required reoperation without loss of the contour correction. All of the complications occurred in the first 18 months of our study. There has been excellent retention of implant volume with no recurrence of contour defects to date. We have not found any visible evidence of interference with craniofacial growth over the study period. We conclude that hydroxyapatite cement is a versatile and safe biomaterial when used for the correction of secondary craniofacial contour defects in children and adults. The coupling of antibiotics with this biomaterial may have applications in the treatment of osteomyelitis. PMID- 10513907 TI - The reverse auricular flap: a new flap for nose reconstruction. AB - In the present article, the authors describe a new chondrocutaneous island flap from the ear helix for nose reconstruction. Anatomic studies showed that helix vascularization depends mainly on the superficial temporal vessels. The presence of vascular communications between the anterior frontal branch of the superficial temporal system and the supraorbital and supratrochlear arterial systems allows this flap to be used in a reverse vascular flow fashion. This new flap has been used successfully in seven cases for reconstructing composite defects of the nasal tip and ala. The donor-site defect is repaired with an advancement and rotation flap from the helical rim, leaving an inconspicuous scar and giving an acceptable cosmetic result of the donor area. PMID- 10513908 TI - The aesthetic unit dorsal nasal flap: rationale for avoiding a glabellar incision. AB - The dorsal nasal flap was first introduced by Rieger in 1967. Since that time, it has proven reliable in the coverage of dorsal nasal soft-tissue defects; however, the glabellar component of the flap can leave a conspicuous scar and/or a contracture band. The authors present their experience with 48 patients who had an aesthetic unit dorsal nasal flap and their technique of incisional interface resurfacing, which obviates the need for the glabellar component. Objective independent assessment of the outcomes revealed overall excellent results, with no flap loss, hematoma, or dehiscence. Two cases of contour deformity were noted in male patients with preexisting rhinophyma in the area of the reconstruction. These design modifications enhance the aesthetic result and simplify the use of this flap in dorsal nasal reconstruction. PMID- 10513909 TI - An analysis of 123 temporoparietal fascial flaps: anatomic and clinical considerations in total auricular reconstruction. AB - This article presents an updated review of our experience with 122 temporoparietal fascial flaps, which were used for coverage of fabricated autogenous cartilage frameworks in total auricular reconstructions. Our indications for use of the temporoparietal fascial flap are presented. Partial flap necrosis occurred in 5 cases, total necrosis in 2 of 14 microsurgically transplanted cases, cartilage infection in 2 cases, and paralysis of the frontal branch of facial nerve in 1 case. Prospective observations of vascular anatomy were carried out in the last 93 temporoparietal fascial flaps during flap elevations. Only 59 flaps (63.4 percent) showed a typical pattern, distributed mainly by the superficial temporal artery and vein. Others (36.6 percent) were distributed mainly by various combinations of the posterior auricular artery or vein, occipital artery or vein, diploic vein, and the superficial temporal artery or vein. At the upper margin of the imaginary reconstructed auricle, the mean diameters of the artery and vein were 1.7 mm and 2.2 mm, respectively. There were no significant differences of vascular patterns and their diameters between the temporoparietal fascial flap of microtia sides and of nonmicrotia sides (sides with acquired ear deformities or free-flap donor sides). We are presenting our technical evolution in using the temporoparietal fascial flap for total auricular reconstruction with the goal of reducing surgical complications and improving aesthetic results. PMID- 10513910 TI - Comparison of innervated and noninnervated free flaps in oral reconstruction. AB - Thirteen patients who had undergone ablative surgery for advanced squamous cell carcinoma in which more than half of the tongue had been resected underwent reconstruction in which the cutaneous nerve of a free flap was anastomosed to the stump of the transected lingual nerve. Eight of the patients underwent reconstruction with an innervated anterolateral thigh flap and five patients underwent reconstruction with an innervated rectus abdominis musculocutaneous flap. Sensory recovery of the flap at least 6 months postoperatively was compared in these 13 patients and in 16 additional patients who received noninnervated versions of the same flaps for the same defect. The degree of sensory recovery of innervated thigh flaps was significantly greater than that of noninnervated ones in all modalities and that of innervated rectus abdominis flaps was also greater than that of noninnervated flaps, except for hot and cold perception. These results indicate that sensory regrowth occurs in most areas through the surgically created pathways. However, results of Semmes-Weinstein testing showed that recovery did not reach the level of protective sensation in either type of innervated flap. Although these findings must be followed by additional objective and functional tests and the need for sensory reeducation should be considered, this simple operative procedure can improve postoperative intraoral function and should be attempted whenever possible after ablative surgery. PMID- 10513911 TI - Reconstruction of the mandible with osseous free flaps: a 10-year experience with 150 consecutive patients. AB - Osseous free flaps have become the preferred method for reconstructing segmental mandibular defects. Of 457 head and neck free flaps, 150 osseous mandible reconstructions were performed over a 10-year period. This experience was retrospectively reviewed to establish an approach to osseous free flap mandible reconstruction. There were 94 male and 56 female patients (mean age, 50 years; range 3 to 79 years); 43 percent had hemimandibular defects, and the rest had central, lateral, or a combination defect. Donor sites included the fibula (90 percent), radius (4 percent), scapula (4 percent), and ilium (2 percent). Rigid fixation (up to five osteotomy sites) was used in 98 percent of patients. Aesthetic and functional results were evaluated a minimum of 6 months postoperatively. The free flap success rate was 100 percent, and bony union was achieved in 97 percent of the osteotomy sites. Osseointegrated dental implants were placed in 20 patients. A return to an unrestricted diet was achieved in 45 percent of patients; 45 percent returned to a soft diet, and 5 percent were on a liquid diet. Five percent of patients required enteral feeding to maintain weight. Speech was assessed as normal (36 percent), near normal (27 percent), intelligible (28 percent), or unintelligible (9 percent). Aesthetic outcome was judged as excellent (32 percent), good (27 percent), fair (27 percent), or poor (14 percent). This study demonstrates a very high success rate, with good-to excellent functional and aesthetic results using osseous free flaps for primary mandible reconstruction. The fibula donor site should be the first choice for most cases, particularly those with anterior or large bony defects requiring multiple osteotomies. Use of alternative donor sites (i.e., radius and scapula) is best reserved for cases with large soft-tissue and minimal bone requirements. The ilium is recommended only when other options are unavailable. Thoughtful flap selection and design should supplant the need for multiple, simultaneous free flaps and vein grafting in most cases. PMID- 10513912 TI - Long-term predictable nipple projection following reconstruction. AB - The creation of the nipple-areola complex is often the final step in the surgical treatment of breast cancer patients, and it consequently has important symbolic and aesthetic implications. Patient expectations and the need for symmetry make nipple projection a crucial aesthetic determinant of nipple reconstruction. We hypothesize that long-term nipple projection and shape can be achieved in a predictable fashion using the modified star dermal fat flap technique. Prospectively, 93 nipples were reconstructed by a single surgeon using a modified star dermal fat flap technique in 44 implant and 49 TRAM flap breast reconstructions. Flap dimensions (base diameter and flap length) were designed according to patient desire or to the base diameter and projection of the opposite breast nipple. A standardized, 3-month postoperative care regimen was observed in all patients. Nipple projection was assessed by the same observer at each follow-up examination. The average length of follow-up was 730 days (745 for TRAM reconstructions and 713 for implants). Consistently, an average of 41 percent of the intraoperative projection remained intact in both groups at final evaluation (SD 12 percent). The total flap length was strongly predictive of intraoperative and long-term projection (r = 0.64 and 0.86, p < 0.0001). Flap lengths ranged from 5.5 to 9.0 cm, and in a linear correlation, resulted in intraoperative projection of 1.0 to 2.1 cm, respectively, and long-term projection of 0.4 to 0.83 cm, respectively. Based on the linear relationship, every 1-cm increase in flap length could be expected to result in a 0.16-cm increase in projection. When controlled for flap length and intraoperative projection, there was no difference between TRAM and implant nipple reconstruction in predicting postoperative nipple projection. Intraoperative planning and execution are critical to achieve predictable nipple shape, size, and projection. The dimensions of the star dermal fat flap can be strategically modified to allow the surgeon predictable projection with a consistent 41-percent preservation of intraoperative nipple projection in both TRAM and implant patients at 2 years. PMID- 10513913 TI - Surgical options for the early-stage breast cancer: factors associated with patient choice and postoperative quality of life. AB - Patients with early-stage breast cancer have three surgical options: lumpectomy with radiotherapy, mastectomy alone, and mastectomy with breast reconstruction. Our objective was to compare women in these three groups with respect to demographics, preoperative counseling, postoperative body image, and quality of life. Women having undergone surgery for stage 1 or 2 breast cancer between 1990 and 1995 were selected by random sampling of hospital tumor registries and were mailed a self-administered questionnaire, which included the Medical Outcomes Survey Short Form 36. Patients were stratified into three mutually exclusive groups: lumpectomy with axillary node dissection and radiotherapy, modified radical mastectomy, and modified radical mastectomy with breast reconstruction. In total, 267 of 525 surveys were returned (50.9 percent). Compared with mastectomy patients, breast reconstruction patients were younger (p < 0.001), better educated (p = 0.001), and more likely Caucasian (p = 0.02). Among mastectomy patients, 54.9 percent recalled that lumpectomy had been discussed preoperatively and 39.7 percent recalled discussion of breast reconstruction. Post-operative comfort with appearance was significantly lower for mastectomy patients. The relationship between type of surgery and postoperative quality of life varied with age. Under 55, quality of life was lowest for mastectomy patients on all but two Medical Outcomes Survey Short Form 36 subscales. Over 55, quality of life was lowest for lumpectomy patients on all subscales (p < 0.05 for all subscales except social functioning and role-emotional). Treatment choice may be related to age, race, education, and preoperative counseling. Whereas the effect of breast cancer on a woman's life is complex and individual, the type of surgery performed is a significant variable, whose impact may be related to patient age. PMID- 10513914 TI - Factors predictive of quality of life after silicone-implant explanation. AB - 'Medical records of 180 patients who underwent silicone gel-filled breast implant explanation were retrospectively reviewed. The goal of this study was to determine if any patient variable(s) had predictive value for positive quality of life after explanation. The medical complaints, symptoms, and established diagnoses were considered equally and were referred to as self-reported medical problems. The study revealed that no single problem or pairing of problems was associated with or predictive of outcome. The results show, however, that the number of medical problems was significantly predictive of patient perception of quality of life. A total of 50 explanation patients completed quality-of-life surveys. Specifically, those patients who reported five or fewer medical problems that predated explantation were significantly more likely to perceive an increased quality of life after surgery than those who reported nine or more medical problems (p < 0.04). In conclusion, it is difficult to correlate subjective patient symptoms with postoperative improvements in quality of life after explantation. PMID- 10513915 TI - Optimization of conscious sedation in plastic surgery. AB - The administration of conscious sedation by the plastic surgeon must be safe, efficient, and consistent. In the proper setting, with trained staff and appropriate backup, conscious sedation can allow optimal patient satisfaction with expedient recovery in addition to cost containment. The highly effective local anesthesia afforded by dilute, high-volume ("tumescent") infiltration extends the use of conscious sedation to cases previously performed under general anesthesia or deep sedation. The purpose of this analysis was to identify variables in conscious sedation that affect traditional outcome parameters in ambulatory surgery, particularly the duration of recovery and adverse events such as nausea and emesis. All perioperative and operative records of 300 consecutive patients having plastic surgical procedures under conscious sedation were carefully reviewed. Patients were ASA class I or II by requisite. Conscious sedation followed a standardized administration protocol, using incremental doses of two agents: midazolam (0.25 to 1 mg) and fentanyl (12.5 to 50 mcg). A subset of patients received preoperative oral sedation. Multivariate statistical analysis was conducted using SPSS 8.0 for Windows (SPSS Inc., Chicago, Ill.). Of the 300 patients, same-day discharge was intended for 281. Eight procedure categories were defined. No anesthetic complications occurred. As expected, recovery time was significantly correlated with the duration and type of procedure (p < 0.001) and the total dosage of both intraoperative sedative agents (p < 0.001). Interestingly, a negative correlation with advancing age existed (p < 0.001), likely reflecting the significantly higher intraoperative sedative dosing in younger patients (p < 0.001). When controlled for the effects of procedure duration and intraoperative sedative dosing, two other variables-use of preoperative oral sedation and postoperative nausea/emesis-significantly lengthened recovery time (p = 0.0001 for each). Fifteen unintended admissions occurred secondary to nausea, prolonged drowsiness, or pain control needs. Conscious sedation is an effective anesthetic choice for routine plastic surgical procedures, many of which would commonly be performed under general anesthesia. In our experience with a carefully structured and controlled conscious sedation protocol, the technique has proven to be safe and effective. This analysis of outcome parameters identified two important and potentially avoidable causes of recovery delay following conscious sedation-oral premedication and nausea/emesis. Nausea and emesis were particularly problematic in that they were responsible for 11 of 15 (73 percent) unintended admissions. Preoperative sedation is valuable in certain circumstances, and its use is not discouraged; however, its benefits must be weighed against its unwanted effects, which can include a prolongation of recovery. PMID- 10513916 TI - Treatment of pyogenic granuloma by shave excision and laser photocoagulation. AB - We present herein our technique for the management of pyogenic granulomas. Twenty such lesions were treated in 13 patients by shave excision followed by laser photocoagulation of the base. Recurrence was noted in just one case and was successfully treated by repeated laser treatment. The cosmetic results have been uniformly excellent. Shave excision followed by laser photocoagulation is an effective therapeutic alternative to excision and linear closure for the treatment of pyogenic granuloma, one that minimizes scar formation while preserving the ability to confirm the diagnosis. PMID- 10513917 TI - Fist position for skin grafting on the dorsal hand: I. Analysis of length of the dorsal hand surgery in hand positions. AB - In skin grafting for reconstruction of burns and contracture deformities of the dorsal hand, the hand is kept in a proper position to provide the greatest amount of skin and to avoid the secondary functional deformity. The safe position has been commonly used for immobilizing the hand, but this is to protect the hand function rather than to provide maximal surface for skin grafting. Split thickness skin graft contracts up to 30 to 50 percent of the original size owing to secondary contraction. If insufficient skin is grafted, contracture deformity of the dorsal hand may occur. To graft the greatest amount of skin on the dorsal hand, the hand should be kept preoperatively in a position flexing all joints of the wrist, metacarpophalangeal joints, and interphalangeal joints and maximally stretching the dorsal hand (a fist position). We studied the surface length of the dorsal hand between the wrist, the metacarpophalangeal joint, and the eponychium in the anatomic, safe, and fist positions of the right hand in 60 adults. Difference of total length between the anatomic and safe positions was not statistically significant (p > 0.05). The total length in a fist position was significantly increased in comparison with the other two positions (p < 0.05). In a fist position compared with the safe position, the increase in length of the dorsal surface of the proximal hand was 11 to 20 percent except in the thumb, and the increase in length of the dorsal surface of the finger was 12 to 17 percent. The increase in total length of a fist position was about 9 mm (7 to 8 percent) in the thumb and 20 to 32 mm (14 to 18 percent) in the index to little fingers. It suggests that the safe position fails to provide an increased dorsal hand surface area for skin grafting compared with the anatomic position. The greatest amount of skin can be grafted in a fist position. Hand immobilization in a fist position for 7 to 9 days after skin grafting has not resulted in irrevocable joint stiffness in our experience. If injury of the deep structures is not present, the hand should be immobilized in a fist position before skin grafting on the dorsal hand. PMID- 10513918 TI - The fetal cleft palate: II. Scarless healing after in utero repair of a congenital model. AB - The role of fetal surgery in the treatment of non-life-threatening congenital anomalies remains a source of much debate. Before such undertakings can be justified, models must be established that closely resemble the respective human anomalies, and the feasibility and safety of these in utero procedures must be demonstrated. The authors recently described and characterized a congenital model of cleft palate in the goat. The present work demonstrates the methodology they developed to successfully repair these congenital cleft palates in utero, and it shows palatal healing and development after repair. A surgically created cleft model was developed for comparative purposes. Palatal shelf closure normally occurs at approximately day 38 of gestation in the caprine species. Six pregnant goats were gavaged twice daily during gestational days 32 to 41 (term, 145 days) with a plant slurry of Nicotiana glauca containing the piperidine alkaloid anabasine; the 12 fetuses had complete congenital clefts of the secondary palate. Repair of the congenital clefts was performed at 85 days of gestation using a modified von Langenbeck technique employing lateral relaxing incisions with elevation and midline approximation of full-thickness, bilateral, mucoperiosteal palatal flaps followed by single-layer closure. Six congenitally clefted fetuses underwent in utero repair, six remained as unrepaired controls. Twelve normal fetuses underwent surgical cleft creation by excision of a 20 x 3 mm full thickness midline section of the secondary palate extending from the alveolus to the uvula, at 85 days of gestation. Six surgically clefted fetuses underwent concurrent repair of the cleft at that time; six clefted fetuses remained as unrepaired controls. At 2 weeks of age, no congenitally or surgically created clefts repaired in utero demonstrated gross or histologic evidence of scar formation. A slight indentation at the site of repair was the only remaining evidence of a cleft. At 6 months of age, normal palatal architecture, including that of mucosal, muscular, and glandular elements, was seen grossly and histologically. Cross-section through the mid-portion of the repaired congenitally clefted palates demonstrated reconstitution of a bilaminar palate, with distinct oral and nasal mucosal layers, after single-layer repair. In utero cleft palate repair is technically feasible and results in scarless healing of the mucoperiosteum and velum. The present work represents the first in utero repair of a congenital cleft palate model in any species. The use of a congenital cleft palate model that can be consistently reproduced with high predictability and little variation represents the ideal experimental situation. It provides an opportunity to manipulate specific variables, assess the influence of each change on the outcome and, subsequently, extrapolate such findings to the clinical arena with a greater degree of relevance. PMID- 10513919 TI - Composite tissue allografts in rats: IV. Graft-versus-host disease in recipients of vascularized bone marrow transplants. AB - This laboratory has used a composite tissue allograft model as a vehicle for studies on a new type of bone marrow transplant, the vascularized bone marrow transplant. The model consists of a rat hind limb transplant that incorporates integumentary musculoskeletal, and lymphopoietic tissues. These transplants, in comparison with conventional marrow transplants, have the advantage of providing a syngeneic microenvironment and immediate engraftment of both mature and progenitor hemopoietic cells at the time of transplantation. The characteristics of graft-versus-host disease were studied in this model. Lewis X Brown Norway F1 (LBN RT-1(1+n)) rats received hind limbs from Lewis (LEW RT-1(1)) donors (n = 19). Animals were observed daily for signs of graft-versus-host disease. Necropsies were performed. A minority of animals developed lethal disease (7 of 19 recipients) and demonstrated cachexia with concomitant histopathologic changes of the disease. Acute and chronic groups emerged with distinct clinical courses, which are similar to other models of this disease. Recipients of vascularized bone marrow transplants (limb transplants) showed clinical and histopathologic changes of the disease. The transplants may be used as a model of graft-versus host disease in humans. Most interestingly, the transplant has a lower incidence of disease compared with other methods of bone marrow transplantation and represents an alternative to conventional bone marrow transplantation, which deserves further exploration. It may be possible to develop a new technique for bone marrow transplantation based on this surgical approach. It is proposed that the transfer of vascularized blocks of bone/marrow into prospective recipients as opposed to cellular bone marrow transplants may be preferable. PMID- 10513921 TI - Effects of nerve growth factor on nerve regeneration through a vein graft across a gap. AB - The limited availability of donor sites for nerve grafts and their inherent associated morbidity continue to stimulate research toward finding suitable alternatives. In the following study, the effect of direct administration of nerve growth factor (NGF) into a nerve conduit across a gap was tested in a rat sciatic nerve model. A 1-cm segment of the right sciatic nerve in Sprague-Dawley rats was resected, and the gap was then bridged using one of three methods: group I (NGF-treated group, n = 12), a vein graft filled with NGF (100 ng in 0.3-ml phosphate buffered saline); group II (control group, n = 12), a vein graft filled with phosphate buffered saline only; group III (standard nerve graft, n = 11), a resected segment of the sciatic nerve. All animals were evaluated at 3 and 5 weeks by behavioral testing and at 5 weeks by electrophysiologic testing. At 3 weeks, sensory testing showed that the latency to a noxious stimulus in group I animals (8.0 +/- 5.4 sec, mean +/- SD) was significantly lower than that of group II animals (13.2 +/- 6.5 sec), indicating that sensory recovery was superior in the animals receiving NGF. The mean latency of animals in group III was 12.9 +/- 6.5 sec, but the difference between the latencies of group I and group III did not reach statistical significance. At 5 weeks, there was no difference in sensory testing between groups. Motor function in groups I and III as measured by walk pattern analysis was superior to that of group II at 5 weeks (toe spread ratios 0.66 +/- 0.09, 0.48 +/- 0.07, and 0.69 +/- 0.09 for groups I, II, and III, respectively). Mean motor conduction velocities across the 1-cm gap were 8.6 +/- 4.7 m/sec, 2.5 +/- 0.7 m/sec, and 6.9 +/- 2.9 m/sec in groups I, II, and III respectively. The difference between groups I and III was not statistically significant, but the motor conduction velocity of group II was significantly slower than that of either group I or III (p < 0.002). The positive effects of NGF on regeneration of nerves across a gap seen in this study suggest that it may be useful for treating peripheral nerve injuries in combination with autogenous vein grafts. PMID- 10513920 TI - Effects of 8-Gy radiation on the microcirculation of muscle flaps in the rat. AB - Combination of radical excision and radiation has been used as a treatment modality for cancer patients. As a result, in reconstructive surgery there is often a need to harvest flaps in the vicinity of previously irradiated tissues. Radiation has been shown to cause progressive injury to the macrocirculation and microcirculation, often jeopardizing flap survival. The purpose of this study was to examine whether radiation significantly affects the sequence of leukocyte endothelial interactions or the hemodynamics of the muscle flap in both acute and chronic situations. Male Sprague-Dawley rats (n = 42) were divided into seven groups of six rats each. Rats in group I were not irradiated. Groups II through VII received 8-Gy radiation to the right groin and scrotum. Groups II, III, and IV were examined at 4, 24 and 72 hours, respectively, and groups V, VI, and VII were examined at 1, 2 and 12 weeks. For intravital microscopy, the cremaster muscle was dissected on its neurovascular pedicle. Vessel diameters and red blood cell velocities were measured in the central cremasteric branches and branch arterioles. Capillary perfusion was evaluated in 27 visual fields of each flap. Leukocyte-endothelial interactions were evaluated by numbers of rolling, adhering, and transmigrating leukocytes in post-capillary venules. In the same postcapillary venule, we measured the endothelial edema index (constriction index). The hemodynamics of irradiated flaps did not differ significantly from those of controls. Diameter and red blood cell velocity were increased in the first- and second-order arterioles and were highest at 72 hours and 1 week. After irradiation, third-order arterioles were constricted. Radiation reduced capillary perfusion by 4.3, percent. None of the differences were statistically significant. Neither leukocyte behavior nor the constriction indices differed among the groups. In conclusion, low-dose radiation of 8 Gy does not affect hemodynamics or leukocyte-endothelial interactions of muscle flaps in the rat. Muscle tissue with intact microvasculature can be harvested for reconstructive procedures after low-dose radiation. PMID- 10513922 TI - A comparative study of nerve healing in adult, neonatal, and fetal rabbits. AB - This experiment quantitatively compared the human equivalent of a nerve repair following surgical division in the fetal, adult, and early childhood period of development using a rabbit as an experimental animal model. Twelve time-dated pregnant New Zealand White rabbits at 24 days' gestation (term = 31 days) underwent hysterotomy; one hind limb was delivered through the uterine opening. The sciatic nerve was divided and repaired by primary neurorrhaphy using two 11-0 epineural sutures. Sciatic nerve repair was also performed in 10 neonatal and 10 adult New Zealand White rabbits. Following repair, each group was assessed using electromyography examination, measuring distal motor latency and amplitude at 1, 2, 3, and 4 months postrepair. There was no difference in any of the groups in distal motor latency. The amplitude rose incrementally in all groups, and the fetal group had significantly higher amplitudes (p < 0.02) at 1, 2, 3, and 4 months in comparison with the adult group. There was no statistically significant difference between fetal and neonatal nerve repairs at any of the time periods. At the completion of the study, the nerve repair sites were harvested for histologic estimation of mean myelinated fiber density and fiber diameter distribution distal and proximal to the repair site. A greater percentage of myelinated axons crossed the repair site in the fetal group (83 percent) in comparison with the adult group (63 percent) (p < 0.03). Our study also demonstrated significant increases in the number of larger myelinated fibers crossing the repair site in comparison with the neonatal and adult groups (p < 0.04). This study found that fetal nerve healing following surgical repair is superior to that found in adult animals and results in a higher number of larger myelinated fibers crossing the repair site in comparison with adult and neonatal repairs. PMID- 10513924 TI - Differences between scar and dermal cultured fibroblasts derived from a patient with recurrent abdominal incision wound herniation. AB - Fibroblasts were derived from dermis and scar of a 47-year-old white man with a recurrent incisional hernia as a result of fractured ribs. The scar was thin and stretched, suggesting a defect in the maturation of granulation tissue. After surgical repair, biopsy specimens of discarded scar and skin were used to generate fibroblast cell lines. Fibroblasts maintained in medium containing 10% fetal bovine serum and antibiotic were studied between their third and eighth passage. By phase contrast microscopy, no structural differences were obvious, but it was noted that to pass scar fibroblasts, a more aggressive trypsin regimen was required. Immunohistologic and Western blot analysis of patient scar fibroblasts showed (1) more a smooth muscle actin within stress fibers, (2) increased expression of the vitronectin integrin receptor alpha(v) (CD 51), and (3) reduced expression of the collagen integrin receptor alpha2 (CD 49b). The expression of vinculin from focal adhesions or a tubulin from microtubules was the same among cell lines. Contractions of scar and dermal fibroblast-populated collagen lattice were compared. At 24 hours, contractions were 69 percent with newborn fibroblasts (normal); 68 percent for patient dermal fibroblasts; and only 48 percent for patient scar fibroblasts. The retarded contraction of scar fibroblast-populated collagen lattice was significant (p > or = 0.002). Myosin ATPase activity, critical for lattice contraction, and cell migration were equivalent among all cell lines. A plausible mechanism for the retardation of scar lattice contraction is disruption of fibroblasts and collagen interactions, for which the attachment of cells to collagen is altered. It is proposed that either the decrease in the expression of collagen integrin receptor alpha2 (CD 49b), an increase in the expression of the vitronectin receptor alpha(v) (CD 51), or a combination of both is responsible for disruption of collagen fibroblast interactions. PMID- 10513923 TI - Role of the thromboxane A2 receptor in the vasoactive response to ischemia reperfusion injury. AB - Neutrophil-endothelial adhesion in venules and progressive vasoconstriction in arterioles seem to be important microcirculatory events contributing to the low flow state associated with ischemia-reperfusion injury of skeletal muscle. Although the neutrophil CD-18 adherence function has been shown to be a prerequisite to the vasoconstrictive response, the vasoactive substances involved remain unknown. The purpose of this study was to evaluate the role of thromboxane A2 receptor in the arteriole vasoactive response to ischemia-reperfusion injury. An in vivo microscopy preparation of transilluminated gracilis muscle in male Wistar rats (175 +/- 9 g) (n = 12) was used for this experiment. Three experimental groups were evaluated in this study: (1) sham, flap raised, no ischemia (20 venules, 20 arterioles), (2) 4 hours of global ischemia only (19 venules, 22 arterioles), and (3) 4 hours of global ischemia + thromboxane A2 receptor antagonist (ONO-3708) (17 venules, 20 arterioles). ONO-3708 (5 mg/kg), a specific competitive antagonist of thromboxane A2 receptor, was infused at a rate of 0.04 ml/minute into the contralateral femoral vein 30 minutes before reperfusion. Mean arterial blood pressure was not changed at this dose of ONO 3708 (88 +/- 6 mmHg before infusion, 81 +/- 4 mmHg after infusion, n = 3). The number of leukocytes rolling and adherent to endothelium (15-sec observation) were counted in 100-microm venular segments, and arteriole diameters were measured at 5, 15, 30, 60, and 120 minutes of reperfusion. Leukocyte counts and arteriole diameters were analyzed with two-way factorial analysis of variance for repeated measures and Duncan's post hoc mean comparison. Statistical significance was indicated by a p < or = 0.05. The ischemia-reperfusion-induced vasoconstriction was significantly reduced by the thromboxane A2 receptor antagonist (ONO-3708). The mean arteriole diameters at 30, 60, and 120 minutes reperfusion were significantly greater in the treated animals than in the ischemia-reperfusion controls. Despite a significant increase in treated mean arteriole diameters, 30 percent of arterioles still demonstrated vasoconstriction. Neutrophil-endothelial adherence was not reduced by ONO-3708. Thromboxane A2 receptor blockade significantly reduces but does not eliminate ischemia-reperfusion-induced vasoconstriction in this model. This finding suggests that additional and perhaps more important vasoactive mediators contribute to vasoconstriction. Furthermore, thromboxane A2 receptor blockade has no effect on polymorphonuclear endothelial adherence. PMID- 10513925 TI - Survival of free tissue transfer following internal jugular venous thrombosis. PMID- 10513926 TI - A composite forearm free flap for the secondary repair of the ruptured Achilles tendon. PMID- 10513928 TI - The transverse orbicularis oculi myocutaneous flap: its use as nasal lining. PMID- 10513927 TI - Prefabricated implants or grafts with reverse models of three-dimensional mirror image templates for reconstruction of craniofacial abnormalities. PMID- 10513929 TI - Island dorsalis pedis skin flap in combination with toe or toe segment transfer based on the same vascular pedicle. PMID- 10513930 TI - Circle-to-W flap. PMID- 10513931 TI - On the nature of hypertrophic scars and keloids: a review. PMID- 10513932 TI - The role of open rhinoplasty in the management of nasal dermoid cysts. AB - The nasal dermal sinus cyst is one of many midline nasal masses that often pose diagnostic and treatment dilemmas for the plastic and reconstructive surgeon. The differential diagnosis of the midline nasal mass includes both congenital and acquired processes. A thorough understanding of its cause is crucial to treatment. A comprehensive discussion of the pathogenesis, diagnosis, sequelae, and surgical management, and a representative case analysis, of the nasal dermal sinus cyst is presented to delineate the role of open rhinoplasty in optimizing the care of this congenital nasal deformity. PMID- 10513933 TI - If it sounds too good to be true...evaluating consumer health and cosmetic surgery claims. PMID- 10513935 TI - Vertical preperiosteal rejuvenation of the frame of the eyelids and midface. AB - Aging usually shows first on the eyelids, the periocular frame, and the midface, which is the anterior mobile area of facial expression. The author, therefore, recommends performing a blepharoplasty and rhytidectomy of the upper two-thirds of the face in patients from their mid-30s on. Instead of a subcutaneous approach, correction can be performed at the deep level, with either a lateral or a vertical approach to the midface. The deep lateral approach to the midface was initiated by Skoog in 1974, the vertical subperiosteal approach by Tessier in 1979, and the vertical preperiosteal midface approach by Hinderer in 1985, followed by de la Plaza in 1988. The author's technique is analyzed according to the level of undermining, the transection of the retaining ligaments, the vector of tissue elevation, the stabilization of the superficial musculoaponeurotic system, and the safety of the procedure, mainly regarding the facial nerve. The related anatomy is described. The technique is reliable. No damage to the facial nerve occurred in a consecutive series of 535 patients who were operated on from 1983 through 1998; this was because the author respected the pathway of the nerve, which was determined using cadaver dissections when the technique was first developed. Long-term follow-up, some lasting 15 years, has shown that the results are long-lasting; this is because of the stabilization of the vertically elevated soft tissues, which is done by suturing the temporoparietalis fascia to the temporalis fascia and by using the author's orbicularis suspension technique. A short-scar variation instead of the coronal approach can also be used; the latter is indicated for patients with high foreheads or large forehead ptosis. The author prefers to use the preperiosteal approach, which elevates the soft tissues themselves, after transecting the retaining ligaments, rather than the vertical approach. PMID- 10513934 TI - The role of sclerotherapy in abnormal varicose hand veins. AB - Large, tortuous veins involving the dorsa of the hands often become more prominent with the passage of time and are a common source of patient dissatisfaction. The objective was to evaluate the results of sclerotherapy in the management of unsightly varicose veins of the dorsum of the hand. From January of 1987 to August of 1998, 100 healthy, ambulatory female patients with a mean age of 56.5 years (range, 35 to 78 years) underwent sclerotherapy treatment for abnormally dilated veins on the dorsum of the hands. Patients were divided into two groups: group A consisted of 20 patients treated with 0.5% sodium tetradecyl sulfate (Sotradecol Elkins-Sin, Inc., Cherry Hill, N.J.) or 1.5% polidocanol (Aethoxysklerol Kreussler, Chemische-Fabrik, Wiesbaden, Germany). Group B consisted of 80 patients treated with 3% polidocanol. Postsclerotherapy compression was utilized in all cases. Failure was observed in 16 patients (80 percent) in group A. Successful elimination of varicose veins was obtained in 76 of 80 patients (95 percent) in group B. The diameter of treated vessels ranged from 1 to 6 mm (mean, 3 mm). Adverse events observed included pain, ecchymosis, various degrees of edema, and thrombosis of the treated veins. One patient (1 percent) developed transient neuropraxia of the superficial branch of the radial nerve following treatment of vessels located on the thenar web. Eleven of the 76 patients (14.5 percent) treated successfully with higher concentration developed microscopic neovascularization (matting). In conclusion, (1) low concentrations of sclerosing agents were associated with a high incidence of failure (80 percent); (2) the use of higher concentrations of polidocanol (3%) produced good results in 95 percent of treated patients; (3) adverse events common to sclerotherapy were observed in 90 percent of the treated patients-there were no serious adverse events; and (4) when appropriate sclerosant concentrations were employed, compression sclerotherapy proved to be an effective method of treatment for varicose veins involving the dorsum of the hand. PMID- 10513936 TI - Aging nasolabial fold and treatment by direct excision. AB - The anatomy, terminology, and management of the aging nasolabial fold represent a subject of some controversy in the plastic surgery literature. A variety of rhytidectomy techniques and adjuncts, and directed surgical procedures, have been designed to improve the aesthetics of the aging nasolabial region. Experience with direct excision of the nasolabial fold in more than 30 patients is presented, and the anatomy and terminology of the nasolabial region are discussed. Direct excision has proven to be an effective therapy for the management of the aging nasolabial fold in selected patients, primarily men with sun-damaged skin. PMID- 10513937 TI - Periauricular face lift incisions and the auricular anchor. AB - There is one problematic part of the facialplasty procedure that has been addressed with relative infrequency in the plastic surgery literature. This is the aesthetic management of the periauricular incisions and the resultant wound. The following problems associated with the design and closure of the periauricular incisions are recognized: (1) wide, hypertropic, or hypopigmented scars; (2) excessively elevated temporal hairline and a postauricular hairline "step" deformity; (3) "hidden" or "buried" tragus; (4) low, visible mastoid skin scars; and (5) deformed ear lobule or "pixie ear" deformity. Prevention of these undesirable results of facialplasty and cosmetic enhancement of the periauricular area for the facialplasty patient is the surgical challenge about which this article is concerned. The author's approach to the management of the periauricular area has been used successfully on 770 patients since 1985. This approach includes a unique incision design as well as a technique for internally splinting the periauricular incision at the time of wound closure. The incision design can eliminate the problems of elevated temporal hairline, disruption of the postauricular hair-line, and visible mastoid skin scarring. The incorporated internal splinting technique reduces skin level closure tension to improve the quality of the skin scar formation. This approach further allows for retaining a natural-appearing shape of both the tragus and the lobule of the ear. The biomechanical basis for the internal splinting technique was examined by measuring the forces required to transpose facialplasty flaps to the periauricular area. The auricular cartilage was demonstrated to be a remarkably stable structure to which these flaps could be anchored. PMID- 10513938 TI - Combined gluteoplasty: liposuction and lipoinjection. AB - From April of 1995 to August of 1998, 62 female and four male patients had gluteoplasties. To improve the gluteal region, two techniques that create excellent results in other parts of the body, liposuction and lipoinjection, were combined. The ages of the patients ranged from 18 to 52 years (mean, 31 years). Liposuction was done with a tumescent technique in the lumbosacral, trochanteric, and subgluteal region to improve gluteal shape. The amount of fat aspirated was only that necessary to obtain the desired contour. In all cases, liposuction was also performed in other areas. Lipoinjection was done with round-tip cannulas in different planes of the gluteal region, and the fat was applied in small strips. The quantity of fat infiltrated varied from 120 to 280 cc per gluteus, with a mean of 210 cc. The results were evaluated by the patients and the surgical team with preoperative and postoperative photographs. Follow-up ranged from 3 months to 3 years and 5 months, with a mean of 17 months. No patient was dissatisfied with the results, and more than 90 percent considered their results good or excellent. Liposuction complications consisted of four seromas, six visible irregularities, and two palpable irregularities. Lipoinjection complications occurred in 16 gluteus regions (12 percent); all had gluteal temporal hyperemia and erythema, which resolved with conservative treatment except in one case (4 cc of sterile material corresponding to fat necrosis was drained in that patient). No irregularities or depressions occurred in the gluteus. One case of probable fat embolism syndrome had a satisfactory evolution. This gluteoplasty technique is simple and low in cost, with minimal morbidity and very good results. It is important to note that a good result does not depend on a great amount of fat infiltration but rather on a harmonious way of combining both surgical procedures: fat elimination by liposuction and gluteus augmentation by lipoinjection. PMID- 10513939 TI - Acute hearing loss after liposuction. PMID- 10513940 TI - Simaplast inflatable breast implants: evaluation after 23 years in situ. PMID- 10513942 TI - My modest proposal. PMID- 10513941 TI - Liposuction with standing technique: the true lipo "sculpture". PMID- 10513943 TI - Guidelines for a successful microsurgery training center and research fellowship. PMID- 10513944 TI - Sedation and analgesia in ambulatory settings. American Society of Plastic and Reconstructive Surgeons. Task Force on Sedation and Analgesia in Ambulatory Settings. PMID- 10513945 TI - Operation smile and its help in the development of plastic and reconstructive surgery in Russia. PMID- 10513946 TI - Cranial bone grafts for dorsal nasal augmentation. PMID- 10513947 TI - Anatomic basis and clinical implications for nasal tip support in open and closed rhinoplasty. PMID- 10513948 TI - Simple tie-over: the herniotomy approach. PMID- 10513949 TI - Upper eyelid orbicularis oculi flap with tarsoconjunctival island for reconstruction of full-thickness lower lid defects. PMID- 10513950 TI - Complete excision with staged reconstruction in the treatment of earlobe keloid after ear piercing. PMID- 10513951 TI - Correction of the constricted ear with a boomerang-shaped incision. PMID- 10513952 TI - A complication of a retromolar mucosal flap for cleft palate repair. PMID- 10513953 TI - The sternalis muscle in head and neck reconstruction. PMID- 10513954 TI - Lymphatic malformation or lymphovenous malformation. PMID- 10513955 TI - Treatment of axillary bromhidrosis with superficial liposuction. PMID- 10513956 TI - The distally based forearm island flap in hand reconstruction. PMID- 10513957 TI - Finger splints from syringes. PMID- 10513958 TI - Periareolar marking in Lejour's mammaplasty. PMID- 10513959 TI - Urinary incontinence was improved after abdominoplasty using a very low incision. PMID- 10513960 TI - Endermologie. PMID- 10513961 TI - The future of liposuction and fat. PMID- 10513962 TI - A garment idea for body contour photography. PMID- 10513963 TI - What's wrong with Dermabond? PMID- 10513965 TI - Who's getting old? PMID- 10513964 TI - Telomeres and telomerase and their possible future in plastic surgery. PMID- 10513966 TI - Legislation in Florida regarding the overnight stays for plastic surgery patients. PMID- 10513967 TI - A prospective, randomized clinical trial comparing tibial nailing using fracture table traction versus manual traction. AB - OBJECTIVE: We sought to determine the effectiveness of intramedullary tibial nailing using manual traction with the leg draped free versus standard fracture table positioning and traction. STUDY DESIGN: Prospective, randomized clinical trial. METHODS: Eighty-five tibial shaft fractures (in seventy-nine patients) treated by intramedullary nailing were randomized either to manual traction with the leg draped free or to standard fracture table traction applied through a boot attachment. RESULTS: We found that manual traction provided results, in terms of intraoperative parameters and quality of fracture reduction, similar to those with standard fracture table traction. Manual traction significantly reduced positioning time (twelve minutes versus twenty-five minutes, p = 0.002) and also allowed for multiple simultaneous or sequential procedures in polytrauma patients without the need for re-positioning or re-draping. This saved a further thirty two minutes (mean) in 37 percent of cases treated by manual traction. CONCLUSION: Manual traction for intramedullary nailing of the tibia is an effective technique that can save a significant amount of time without sacrificing the quality of reduction or fixation of tibial shaft fractures. It is especially useful in polytrauma patients with multiple lower-extremity injuries. PMID- 10513968 TI - Influence of the design for fixation implants on local infection: experimental study of dynamic compression plates versus point contact fixators in rabbits. AB - OBJECTIVES: Comparison of infection resistance after local bacterial challenge associated with two different designs for fixation implants: the conventional dynamic compression plate (DCP) and the point contact fixator (PC-Fix). DESIGN: Randomized, prospective study in experimental animals. Grouped sequential experimental procedure. Observation time was twenty-eight days, with twenty animals per group. SETTING: Following surgery, animals were kept without restrictions in individual hutches. ANIMALS: Forty White New Zealand rabbits. Thirty-eight animals, nineteen per group, were included in the final evaluation. INTERVENTION: Under sterile conditions, specially manufactured titanium DCP or PC Fix of identical dimensions were fixed to rabbit tibiae. After wound closure, different concentrations of Staphylococcus aureus, between 2 x 10(4) and 2 x 10(8) colony-forming units (CFU), were inoculated percutaneously at the implant site. MAIN OUTCOME MEASUREMENTS: Implants, underlying bone, and surrounding soft tissues were removed under sterile conditions and quantitatively evaluated for bacterial growth. Infection was defined as positive bacterial growth at the bone implant interface. RESULTS: The overall infection rate was 45 percent. The infection dose of 50 percent (ID50) was 7.08 x 10(5) CFU for the DCP group and 8.51 x 10(6) CFU for the PC-Fix group. The infection rate was 63 percent (twelve of nineteen animals) for the DCP group and 26 percent (five of nineteen animals) for the PC-Fix group. This difference was statistically significant (p = 0.022). CONCLUSIONS: After local bacterial challenge, we found a statistically significant difference in the infection rates depending on the implant design. The higher infection resistance associated with the PC-Fix design seems to be related to the reduced contact area at the bone-implant interface. PMID- 10513969 TI - The effect of surface material and roughness on bone screw stability. AB - OBJECTIVE: To evaluate the effects of osteointegration ability and surface texture on bone screw interface stability in three different groups of titanium screws. DESIGN: Sixty titanium tapered screws were used: twenty screws were polished, uncoated (Group A); twenty screws were rough, coated with titanium (Group B); and twenty screws were rough, coated with fluor-hydroxyapatite (Group C). Thirty screws, ten per group, were implanted in the femurs and tibiae of two sheep, which were euthanized one month after surgery. The remaining thirty screws, ten per group, were implanted in the femurs and tibiae of another two sheep, which were euthanized three months after surgery. RESULTS: At one month, extraction torque of Group C was higher than that of Group A (p = 0.042). At three months, extraction torque of Group C was higher than that of Group A (p < 0.0001) and Group B (p < 0.0001). At three months, extraction torque of Group C was higher compared with the corresponding insertion torque (p < 0.0001) and compared with the corresponding extraction torque at one month (p < 0.0001). At one and three months, a high percentage of bone-screw contact was observed histologically in Groups B and C. A continuous gap with fibrous tissue encapsulation was observed in Group A. CONCLUSIONS: This study shows that the osteointegration ability provided by the type of coating is a very important parameter for optimizing the bone-screw stability. Surface texture is also important. By using screws with optimal osteointegration ability, very positive clinical consequences can be expected. PMID- 10513971 TI - Early versus delayed stabilization of pediatric femur fractures: analysis of 387 patients. AB - OBJECTIVES: To assess the effect of timing of femur fracture stabilization on pulmonary complication rates in pediatric trauma patients. DESIGN: Retrospective review. SETTING: Level I trauma center. PATIENTS: Three hundred eighty-seven previously healthy patients from zero to fifteen years of age with traumatic diaphyseal femur fractures. INTERVENTION: Femur fracture stabilization: early (less than twenty-four hours after injury) in 213 patients and late in 174 patients. MAIN OUTCOME MEASUREMENTS: Age, sex, GCS (Glasgow Coma Score), AIS/ISS (Abbreviated Injury Score/Injury Severity Score), timing of fracture stabilization, duration of mechanical ventilation, intensive care unit stay, and hospital stay were recorded. Pulmonary complications, including pneumonia, respiratory distress syndrome, and pulmonary embolus, were recorded. RESULTS: Thirteen patients developed pulmonary complications. Twelve of these had severe head injuries (GCS < or = 8). One had sustained an upper cervical spine fracture that resulted in quadriplegia. Statistical analysis revealed GCS, GCS < or = 8, ISS, and head and neck AIS to be significant predictors of pulmonary complications. Early stabilization of femur fractures had no apparent effect on the pulmonary complication rate. CONCLUSIONS: Pulmonary complications are rare in pediatric femur fracture patients. Patients with severe head injuries (GCS < or = 8) or cervical spinal cord injuries are at high risk for pulmonary complications. The timing of femur fracture stabilization does not appear to affect the prevalence of pulmonary complications in these patients. PMID- 10513970 TI - Biomechanical study of atlantoaxial arthrodesis: transarticular screw fixation versus modified Brooks posterior wiring. AB - OBJECTIVE: The purpose of the present study was to compare the biomechanical stability of C1 and C2 vertebrae after treatment of ligamentous instability by either modified Brooks posterior wiring (MB) or transarticular screw (TAS) techniques. We hypothesized that the TAS technique would be more stable because of direct fixation through the facet joints. STUDY DESIGN: We studied the in vitro stability (arthrodesis) of TAS fixation of C1 and C2 versus that of MB. TAS fixation involves placing screws across the facets from posteriorly at C2 to the anterior surface of C1, plus a bone graft and posterior wiring of C1 and C2. METHODS: Cervical spines from nine individuals with an average age of sixty-two years (range 51 to 71 years) were harvested from cadavers (six male, three female). C1 and the segment from C2 to C5 were potted to allow motion only at the C1-C2 articulation. The specimens were destabilized by cutting the transverse ligament on both sides of the odontoid and the tectorial membrane between C1 and C2. The MB and TAS techniques were performed by methods similar to those described in the literature. The stiffness of the C1-C2 articulation of each specimen was tested under rotation, lateral bending, flexion, and anterior translation in random order. Intact and destabilized specimens fixed with either MB or TAS were tested in sequence. RESULTS: Significantly higher stiffness values in the elastic zone were obtained with the TAS technique than with the MB technique for all modes of testing (p < 0.002, t test). Values for the neutral zone (the region where minimal loads produce displacement) were not significantly different between the MB and TAS techniques (p > 0.1, t test). CONCLUSION: We conclude that stability is significantly enhanced by use of the TAS construct for treatment of ligamentous instability at the atlantoaxial joint for all motions tested in the present study. PMID- 10513973 TI - Use of the Hoffman 2 compact external fixator in the treatment of redisplaced unstable distal radial fractures. AB - The aim of this study was to examine the use of a new joint sparing external fixation device for unstable redisplaced fractures of the distal radius. Participants were twenty consecutive patients with unstable redisplaced fractures of the distal radius with sufficient space in the distal fragment to allow use of a nonbridging technique. The patients had to be capable of cooperating with functional outcome measures. All patients underwent closed nonbridging external fixation of the distal radius using the Hoffman 2 compact external fixator. The main outcome measures were radiological determinations of dorsal angle, radial shortening, and carpal alignment; measurements of mass grip strength and range of movement; and rate of complications. Volar tilt was successfully regained and maintained (mean 4 degrees) at final review. Radial shortening was a mean of one millimeter at final review. Nineteen of twenty patients regained normal carpal alignment. Grip strength returned to a mean of 74 percent of the opposite (normal) side in the whole group and 88 percent in those who completed review for the longest periods. Ranges of movement were restored to around 80 percent, except flexion (66 percent). The rate of major complications was 15 percent. We conclude that nonbridging external fixation using the Hoffman 2 compact device reliably restores and maintains volar tilt and radial length after re-reduction of unstable fractures of the distal radius. Functional outcome and complications are comparable with findings in previous reports. PMID- 10513972 TI - Treatment of ununited femoral shaft fractures associated with locked nail breakage: comparison between closed and open revision techniques. AB - OBJECTIVES: To investigate and compare closed and open revision techniques in the treatment of ununited femoral shaft fractures associated with locked nail breakage. DESIGN: Retrospective. SETTING: University hospital. METHODS: Ununited femoral shaft fractures associated with locked nail breakage were treated with either closed or open revision (nine or eighteen cases, respectively). The closed technique entailed closed removal of the broken nail and reinsertion of a stable intramedullary nail after reaming the marrow cavity. The open technique included open removal of the broken nail, reinsertion of a stable intramedullary nail or plate, and cancellous bone graft supplementation. Union rate, union period, perioperative course, and complications were compared. RESULTS: Eight closed and fifteen open technique cases were followed for at least one year (median two years). Cases treated with the closed technique had a union rate of 100 percent, a union period of 4.4+/-0.9 months, an operating time of 1.5+/-0.4 hours, no blood transfusion, and no complications. Open technique cases demonstrated a union rate of 100 percent, a union period of 5.7+/-1.5 months (p = 0.033), an operating time of 2.4+/-0.4 hours (p < 0.001), blood transfusion of 1,000+/-500 milliliters (p < 0.001), and no complications. CONCLUSIONS: We recommend the closed revision technique because its union period and operating time are shorter, and it does not require a blood transfusion. Because there is no local wound dissection, infection rates should also be lower. However, the procedure is technically demanding. If it cannot be completed successfully, using the open technique can still achieve a satisfactory outcome. PMID- 10513974 TI - Removal of the deeply inserted proximal interlocking screw in the greater trochanter after femoral nailing. AB - Removal of the proximal interlocking screw (PIS), especially one that is deeply inserted in the greater trochanter or covered by heterotopic ossification, is troublesome and requires considerable effort because the search for its head usually requires removing bone from around the insertion site. We introduce a simple tip to remove this complicated PIS with the aid of a proximal drill guide, drill bit, and AO countersink. PMID- 10513975 TI - Broken Ender nails after fixation of concomitant ipsilateral fractures of the femoral neck and shaft. AB - Concomitant ipsilateral fractures of the upper part and shaft of the femur are uncommon injuries. Because of the small number of cases reported and the diversity of devices used, no single type of internal fixation stands out as the best treatment for both of these fractures. This report describes a case of concomitant ipsilateral fractures of the femoral neck and shaft that is further distinguished by a rare complication: breakage of Ender nails after fixation. PMID- 10513976 TI - Anterior tibial aneurysm following inversion injury to the ankle. AB - Arterial injuries in nonpenetrating low-energy injuries to the extremities are rare but can occur when joint injuries put the vessels in traction against their immobile attachments to the long bones. The most common injuries are to the popliteal artery (because of its tethered nature proximal to the popliteal fossa) and the brachial artery (because it is tethered to the humerus proximal to the elbow). The second reported case of an aneurysm of the anterior tibial artery resulting from an inversion injury to the ankle is described. PMID- 10513977 TI - Nonunion of fracture of the neck of the radius: a report of three cases. AB - Nonunion of a radial neck fracture is uncommon. Our report of three cases aims to highlight the fact that this complication is possible following such a fracture in adults. Appropriate clinical and radiologic follow-up is necessary to make sure such nonunion not missed. Surgical fixation (when nonunion of radial neck fracture is suspected) or excision of the radial head may be necessary if the complication is symptomatic. When associated with an ulna fracture, the threshold for internal fixation of both fractures must be lower. PMID- 10513978 TI - Emerging technology: remote analysis of traumatic musculoskeletal radiographs transmitted by electronic mail. AB - OBJECTIVES: To determine whether remote analysis of radiographs via electronic mail (e-mail) had an impact on treatment decision-making. DESIGN: Prospective. SETTING: Level I trauma center. PATIENTS/PARTICIPANTS: Twenty-five cases randomly selected from previous emergency room consultation. INTERVENTION: Textual descriptions obtained from emergency medicine physicians were compared with computer-digitized images of radiographs sent via e-mail and with the actual radiographs. Four board-certified orthopaedic surgeons reviewed all three forms of data to determine fracture diagnosis and treatment plans. MAIN OUTCOME ASSESSMENT: Diagnosis and treatment plans were obtained via written questionnaire after review of each group of data (textual, digitized image, and actual radiograph). Results were then compared across groups to determine whether digitized images were better than textual descriptions and equivalent to actual radiographs. RESULTS: Statistical analysis revealed a significant improvement in the frequency of correct diagnosis and treatment planning when digitized images were used (91 percent) compared with textual descriptions alone (48 percent) (p < 0.001). The difference in correct diagnosis and treatment plans between digitized images and actual radiographs was not statistically significant (p = 0.27). CONCLUSION: Digitized radiographs sent via e-mail can significantly improve accuracy of diagnosis and treatment compared with a simple verbal description. PMID- 10513979 TI - Regeneration of bone in critical defects. PMID- 10513980 TI - MRI and avascular necrosis. PMID- 10513981 TI - Stress protein/peptide complexes derived from autologous tumor tissue as tumor vaccines. AB - Vaccination of inbred mice with tumor-derived stress proteins hsp70, hsp90, and gp96/grp94 elicits a protective immunity to the tumor from which the vaccine was purified. There is now comprehensive experimental evidence that the antigenicity of tumor-derived hsp70, hsp90, and gp96 preparations results from diverse arrays of endogenous peptide antigens complexed with these stress proteins. Vaccination with tumor-derived stress protein/peptide complexes leads to their uptake and processing by professional antigen-presenting cells and to presentation of associated tumor peptide antigens to cytotoxic T cells. This induces a tumor specific cytotoxic T cell response. The attractiveness of the concept of using tumor-derived stress proteins as vaccines is derived from two observations: (i) tumor stress protein vaccines mirror the individual antigenicity of a tumor, which results from random mutations due to genetic instability; and (ii) stress proteins represent powerful adjuvants for the peptide antigens complexed to them. PMID- 10513983 TI - Role of HSP90 in mediating cross-talk between the estrogen receptor and the Ah receptor signal transduction pathways. AB - Tetrachlorodibenzo-p-dioxin (TCDD)-mediated gene transactivation via the Ah receptor (AhR) has been shown to be dependent upon estrogen receptor (ER) expression in human breast cancer cells. We have investigated the 90-kDa heat shock protein (HSP90) as a mediator of cross-talk between the AhR and the ER signal transduction pathways. The effect of HSP90 overexpression on receptor activity was determined by transient transfection assays using a HSP90 expression vector. Ligand-inducible gene expression was inhibited when the HSP90 expression vector was cotransfected with a TCDD-responsive reporter plasmid. However, overexpression of HSP90 did not block induction of an estrogen-responsive reporter plasmid. To determine whether ER facilitates AhR signaling through its ability to squelch HSP90, two vectors expressing protein products that bind HSP90 were transfected into MDA-MB-231 cells. Introduction of (i) He11, an ER deletion mutant that does not bind DNA, and (ii) the ligand-binding domain of human AhR, both led to increased basal and TCDD-inducible CYP1A1 expression. Finally, the subcellular distribution of HSP90 was investigated in human breast cancer cell lines. These studies showed HSP90 to be primarily cytoplasmic in ER-positive cell lines, whereas in matched ER-negative cell lines HSP90 was distributed equally between the cytoplasm and nucleus. Taken together, these results demonstrate that HSP90 can regulate AhR activity in vivo, and that Ah-responsiveness is dependent upon cellular ER content through a mechanism that involves HSP90. PMID- 10513982 TI - Mitochondrial peripheral-type benzodiazepine receptor expression. Correlation with gonadotropin-releasing hormone (GnRH) agonist-induced apoptosis in the corpus luteum. AB - We have demonstrated that continuous administration of a gonadotropin-releasing hormone agonist (GnRH-Ag) decreases the expression of the mitochondrial peripheral-type benzodiazepine receptor (PBR) and increases the rate of DNA degradation in a time-dependent manner in the corpora lutea of pregnant rats. In the present study, we show in situ the GnRH-Ag-induced DNA fragmentation and correlate the increase of the rate of DNA degradation with the decrease in mitochondrial PBR ligand binding (r = 0.89). The GnRH-Ag-induced decrease in the 18-kDa PBR protein also correlated with the reduction in the Bcl-X(L), but not Bcl-2 (cell survival), gene product levels and the increase in the Bax (cell death) gene product expression in the luteal mitochondrial preparations. Considering the function of PBR in cholesterol uptake and intramitochondrial movement, we propose that decreased PBR expression may lead to reduced levels of mitochondrial membrane cholesterol, which, together with the ability of Bcl-X(L) and Bax to form ion channels, produces breaks in the outer membranes allowing the exit of cytochrome c, thus triggering apoptosis. Alternatively, PBR may exert an as yet unidentified anti-apoptotic function. PMID- 10513984 TI - In vitro and in vivo antitumor activity of a novel immunomodulatory drug, leflunomide: mechanisms of action. AB - Leflunomide, a novel immunomodulatory drug, has two biochemical activities: inhibition of tyrosine phosphorylation and inhibition of pyrimidine nucleotide synthesis. In the present study, we first showed that A77 1726 [N-(4 trifluoromethylphenyl-2-cyano-3-hydroxycrotoamide)], the active metabolite of leflunomide, was more effective at inhibiting the tyrosine kinase activity of platelet-derived growth factor (PDGF) receptor than that of epidermal growth factor (EGF) receptor, and had no effect on the tyrosine kinase activity of the fibroblast growth factor receptor. In the presence of exogenous uridine, A77 1726 was more effective at inhibiting the PDGF-stimulated proliferation of PDGF receptor-overexpressing C6 glioma than the EGF-stimulated proliferation of EGF receptor-overexpressing A431 cells. In vivo studies demonstrated that leflunomide treatment strongly inhibited the growth of the C6 glioma but had only a modest effect on the growth of the A431 tumor. Uridine co-administered with leflunomide did not reverse the antitumor activity of leflunomide on C6 and A431 tumors significantly. Quantitation of nucleotide levels in the tumor tissue revealed that leflunomide treatment significantly reduced pyrimidine nucleotide levels in the fast-growing C6 glioma but had no effect on the relatively slow-growing A431 tumor. Whereas uridine co-administration normalized pyrimidine nucleotide levels, it had minimal effects on the antitumor activity of leflunomide in both tumor models. Immunohistochemical analysis revealed that leflunomide treatment significantly reduced the number of proliferating cell nuclear antigen-positive cells in C6 glioma, and that uridine only partially reversed this inhibition. These results collectively suggest that the in vivo antitumor effect of leflunomide is largely independent of its inhibitory effect on pyrimidine nucleotide synthesis. The possibility that leflunomide exerts its antitumor activity by inhibition of tyrosine phosphorylation or by a yet unidentified mode of action is discussed. PMID- 10513985 TI - Cholesterol 7-hydroperoxides in rat skin as a marker for lipid peroxidation. AB - Concentrations of cholesterol 7alpha- and 7beta-hydroperoxides (Ch 7-OOHs) in the skin of rats were determined by HPLC with a chemiluminescence detector. We demonstrated that (a) the concentrations of Ch 7-OOHs in rat skin were highly correlated with rat age (r = 0.929; N = 51, 1 to 55 weeks old), (b) the concentrations of Ch 7-OOHs in the skin of rats in an ambient light room were not significantly different from those found in rats kept in a dark room for 12 weeks, and (c) lipid peroxidation in vitro induced by ADP-Fe2+ caused an increase in the concentrations of Ch 7-OOHs in homogenates of rat skin. These results indicated that levels of Ch 7-OOHs in skin might be a good marker for aging of rats and might be independent of housing illumination, thus a good marker for endogenous lipid peroxidation. Furthermore, we observed that ultraviolet light B (UVB) irradiation markedly enhanced the concentrations of Ch 7-OOHs in the skin of rats in vivo depending on the duration of the irradiation, and the increases in Ch 7-OOHs were inhibited by radical scavengers, i.e. tocopherols. Therefore, it was suggested that the levels of Ch 7-OOHs in the skin could also be a good marker for UVB-dependent lipid peroxidation. PMID- 10513986 TI - Homing of negatively charged albumins to the lymphatic system: general implications for drug targeting to peripheral tissues and viral reservoirs. AB - The present study shows the lymphatic distribution of the negatively charged anti HIV-1 agents succinylated or aconytilated human serum albumins (HSAs) in rats. Quantitation of blood and lymphatic concentrations of these proteins was performed through fluorescence detection of the fluorescein isothiocyanate (FITC) labeled proteins. At several time points after i.v. injection, samples were taken from the cannulated thoracic duct and the carotid artery. Distribution of the negatively charged albumins (NCAs) to lymph was much more rapid than that of albumin itself and was dependent on the total net negative charge added to the protein: the half-life times of lymphatic equilibration were 15, 30, and 120 min for FITC-labeled aconytilated HSA, FITC-labeled succinylated HSA, and FITC labeled HSA, respectively. Lymph to blood concentration ratios of the studied compounds obtained at steady state approached unity. In addition, the fluorescence in both body fluids was shown to represent unchanged labeled proteins. It was therefore inferred that the NCAs efficiently passed the endothelial barrier from blood to the interstitial compartment. Subsequently, we studied whether a specialized process was involved in the endothelial passage of the NCAs to the lymph. The following observations supported such a mechanism: a) preinjection of the scavenger receptor blockers polyinosinic- and formaldehyde treated HSA reduced the transport from blood to the lymphatic compartment of FITC labeled aconytilated HSA by more than 90%; b) the rate of lymphatic distribution was largely reduced when the body temperature of the rat was lowered to 28 degrees; and c) pre-administration of chloroquine resulted in a significant reduction in the lymphatic distribution of the NCAs. These data collectively indicate that a scavenger receptor-mediated process is involved in the transendothelial transport of NCAs. In situ localization in lymph nodes of the rat showed that FITC-labeled aconytilated and succinylated HSA are mainly present in the germinal center and parafollicular zones. The efficient distribution of these anionized proteins to the lymphatic system is of particular interest for HIV therapy, taking into account that replication of HIV mainly takes place in the lymphoid system. The observation that macromolecules, through charge modification, can extravasate through a receptor-mediated transcytotic process is potentially of major importance for the delivery of drugs with macromolecular carriers to cells not directly in contact with the blood. PMID- 10513987 TI - Alpha1-adrenoceptor-mediated formation of glycerophosphoinositol 4-phosphate in rat heart: possible role in the positive inotropic response. AB - In the present study, we investigated whether phospholipase A2 (PLA2)/lysophospholipase activity producing glycerophosphoinositols from phosphoinositides was operating in rat heart and could be stimulated by alpha1 adrenergic agonists. PLA2/lysophospholipase activity was found in homogenates from rat right ventricles. The stimulation of PLA2/lysophospholipase activity by noradrenaline (NA) was prevented either by the alpha1-adrenergic antagonist prazosin or arachidonyl trifluoromethyl ketone, a selective inhibitor of the 85 110 kDa, sn-2-arachidonyl-specific cytosolic PLA2. The selective alpha1 adrenergic agonist phenylephrine induced a concentration- and time-dependent increase in glycerophosphoinositol (GroPIns) and glycerophosphoinositol 4 phosphate (GroPIns4P) in rat right ventricle slices prelabelled with D-myo [3H]inositol. In electrically driven strips of rat right ventricles, prelabelled with D-myo-[3H]inositol, the positive inotropic effect induced by 20 microM NA in the presence of propranolol was accompanied by the formation of GroPIns and GroPIns4P. The concentration of the formed GroPIns4P (1.33+/-0.12 microM, N = 6) was similar to that previously reported to inhibit the Na+/Ca2+ exchanger in cardiac sarcolemmal vesicles (Luciani S, Antolini M, Bova S, Cargnelli G, Cusinato F, Debetto P, Trevisi L and Varotto R, Biochem Biophys Res Commun 206: 674-680, 1995). These findings show that the stimulation of alpha1-adrenoceptors in rat heart is followed by an increase in the formation of GroPIns4P, which may contribute to the positive inotropic effect of alpha1-adrenergic agonists by inhibition of the Na+/Ca2+ exchanger. PMID- 10513988 TI - Structure-activity relationship studies of propafenone analogs based on P glycoprotein ATPase activity measurements. AB - Propafenone analogs (PAs) were previously identified as potent inhibitors of P glycoprotein (Pgp)-mediated toxin efflux. For this as well as other classes of Pgp inhibitors, lipophilicity as well as hydrogen bond acceptor strength are important determinants of biological activity. The question as to whether a direct interaction between PA-type modulators and Pgp takes place was addressed by means of Pgp ATPase measurements and transport studies. Propafenone-type modulators stimulated ATPase activity up to 2-fold over basal activity in a concentration-dependent biphasic manner. Within a series of structural homologs, Ka values of ATPase stimulation strongly correlated with lipophilicity. Analogs containing a quaternary nitrogen stimulated Pgp ATPase activity with lesser efficacy, while Ka values were somewhat higher when compared to corresponding tertiary analogs. Transport studies performed in inside-out plasma membrane (I/O) vesicles demonstrated that analogs containing a tertiary nitrogen rapidly associated with the biomembrane. Quaternary analogs, which are restricted by a permanent positive charge in transiting the plasma membrane by diffusion, accumulated in Pgp containing I/O vesicles in an ATP-dependent and cyclosporin A inhibitable manner, which identified them as Pgp substrates. Identical structure activity relationships were found in either Pgp ATPase stimulation experiments in I/O vesicles or in toxin efflux inhibition studies using intact cells. Therefore, differences in membrane transit are not responsible for the observed structure activity relationships. PMID- 10513989 TI - Pyrimidine nucleobase ligands of orotate phosphoribosyltransferase from Toxoplasma gondii. AB - Sixty-seven pyrimidine nucleobase analogues were evaluated as ligands of Toxoplasma gondii orotate phosphoribosyltransferase (OPRTase, EC 2.4.2.10) by measuring their ability to inhibit this enzyme in vitro. Apparent Ki values were determined for compounds that inhibited T. gondii OPRTase by greater than 20% at a concentration of 400 microM. 1-Deazaorotic acid (0.47 microM) and 5-azaorotic acid (2.1 microM) were found to bind better (8.3- and 1.9-fold, respectively) to T. gondii OPRTase than orotic acid, the natural substrate of the enzyme. Based on these results, a structure-activity relationship of ligand binding to OPRTase was formulated using uracil, barbituric acid, and orotic acid as reference compounds. It was concluded that the following structural features of pyrimidine nucleobase analogues were required or strongly preferred for binding: (i) an endocyclic pyridine-type nitrogen or methine at the 1-position; (ii) exocyclic oxo groups at the 2- and 4-positions; (iii) a protonated endocyclic pyridine-type nitrogen at the 3-position; (iv) an endocyclic pyridine-type nitrogen or methine at the 5 position; (v) an exocyclic hydrogen or fluorine at the 5-position; (vi) an endocyclic pyridine-type nitrogen or methine at the 6-position; and (vii) an exocyclic negatively charged or electron-withdrawing group at the 6-position. A comparison of the results from the present study with those from a previous study on mammalian OPRTase [Niedzwicki et al., Biochem Pharmacol 33: 2383-2395, 1984] identified four compounds (6-chlorouracil, 5-azaorotic acid, 1-deazaorotic acid, and 6-iodouracil) that may bind selectively to T. gondii OPRTase. PMID- 10513990 TI - S-methyl N-butylthiocarbamate sulfoxide: selective carbamoylating agent for mouse mitochondrial aldehyde dehydrogenase. AB - Liver mitochondrial low-Km aldehyde dehydrogenase (ALDH2, EC 1.2.1.3), the isoform responsible for the conversion of acetaldehyde to acetate, is inhibited by the sulfoxide bioactivation products of Et2NC(O)SMe (from the alcohol aversion drug disulfiram), Pr2NC(O)SEt (the herbicide S-ethyl N,N-dipropylthiocarbamate), and BuNHC(O)SMe (from the fungicide benomyl). This study tested the hypothesis that bioactivated BuNHC(O)SMe, the most potent of these thiocarbamates, is a selective carbamoylating agent for ALDH2 of mouse liver in vivo and in vitro. [14C]BuNHC(O)SMe administered i.p. to mice labeled one principal mitochondrial protein, which cochromatographed with ALDH activity by in-gel assay after isoelectric focusing. The labeled protein was isolated by isoelectric focusing (pI 6.1) and SDS-PAGE (54 kDa) and identified as ALDH2 by sequencing of peptides from a tryptic digest. In vivo at 1.5 mg/kg, enzyme inhibition was 80%, and ALDH2 was the only mitochondrial protein labeled extensively, illustrating the outstanding potency and specificity. ALDH2 also was labeled upon incubation of mouse liver mitochondria with [14C]BuNH-C(O)SMe in the presence of microsomes (P450) and NADPH. In contrast, under similar conditions, [14C]Pr2NC(O)SEt sulfoxide labeled primarily two other proteins at approximately 58 and approximately 61 kDa, establishing a very different selectivity for the two sulfoxides. These findings are of interest relative to selective inhibitors and carbamoylating agents for ALDH2 and to alcohol aversion upon exposure to herbicides and fungicides. PMID- 10513991 TI - Cellular and molecular mechanisms involved in insulin's potentiation of glycogen synthase activity by metformin. AB - By taking advantage of the Xenopus oocyte model, we recently confirmed the in vitro enhancing effect of metformin (MET) on glycogen synthase (GS) activity when induced by insulin (INS). We now investigated some mechanistic aspects of its modulatory role upon the hormonal regulation of this rate-limiting enzyme. The action of 20 microM MET (approximately 3.3 microg/mL) was measurable at early steps in the intracellular metabolic pathway: the amount of adenosine 3',5' cyclic monophosphate (cAMP) was markedly decreased in the presence of the biguanide plus 50 nM INS (to about 60% of control vs 25% with INS alone). The injection of tyrphostin B46, a potent inhibitor of insulin receptor (IR) associated tyrosine kinase activity, led to a drastic reduction in MET-stimulated GS activity in the presence of INS. MET failed to increase the activity of type 2 protein phosphatases whether INS was present or not. However, a specific inhibitor of type 1 phosphatases, when microinjected, blocked both the hormonal effect on GS and its potentiation by MET. The salient feature of this study was that there was almost no accumulation of radiolabeled MET in oocytes: less than 0.1% was found in the cytosol of cells which had been exposed to MET at a therapeutic dose (10 microM) for up to 16 hr. Moreover, a lack of detectable intracellular MET after a 60-min incubation nevertheless correlated with its sustained action on INS-regulated GS activity. From these results, it could be inferred that the major site of MET action may reside within some membrane components of a signaling complex most likely linked to the IR, but in any case located upstream of the branching of reactions which tightly control GS activity. PMID- 10513992 TI - Possible Contribution of Prostaglandin E2 to the antiproliferative effect of phosphodiesterase 4 inhibitors in human mononuclear cells. AB - Phosphodiesterase (PDE) 4, mixed PDE3/4, and non-selective PDE inhibitors have been shown to inhibit the proliferation of human peripheral blood mononuclear cells (HPBM). The aim of the present study was to examine whether endogenous prostaglandins, in particular prostaglandin E2 (PGE2), are involved in mediating the antiproliferative actions of PDE inhibitors, by comparing their effects with drugs which elevate or mimic adenosine 3',5'-cyclic monophosphate (cAMP) through mechanisms other than PDE inhibition. Indomethacin significantly reduced the antiproliferative effects of the PDE4 inhibitors rolipram and CDP840 and the mixed PDE3/4 inhibitor zardaverine, increasing the IC50 values from 2.51 microM to >10 microM, 0.81 microM to 2.82 microM, and 1.58 microM to 4.82 microM, respectively (P < 0.05), but did not alter the effects of theophylline. Forskolin, PGE2, and dibutyryl cAMP also inhibited HPBM proliferation, and in the presence of indomethacin the effects of forskolin and dibutyryl cAMP were reduced (although this was not significant), whereas PGE2 was not affected. Rolipram, CDP840, zardaverine, and dibutyryl cAMP all produced a concentration-related increase in PGE2 production (P < 0.05, ANOVA), but theophylline significantly increased PGE2 production only at the highest concentration examined, 1000 microM. The ability of indomethacin to reduce the antiproliferative effects of rolipram, CDP840, and zardaverine, together with the fact that these drugs can stimulate PGE2 production, suggests that their antiproliferative actions may be mediated in part by stimulation of endogenous PGE2 production. In contrast, it appears that endogenous PGE2 is not critical for the antiproliferative actions of theophylline, forskolin, and dibutyryl cAMP in HPBM. These results establish the importance of co-ordinated regulation of the cAMP phosphodiesterase and cyclooxygenase-PGE2 systems for the regulation of lymphocyte function in man, and have clinical implications for therapeutic approaches to diseases associated with lymphocyte dysregulation. PMID- 10513993 TI - Translocation of G-protein beta3 subunit from the cytosol pool to the membrane pool by beta1-adrenergic receptor stimulation in perfused rat hearts. AB - To elucidate the intracellular function and localization of the heterotrimeric G protein beta3 subunit (Gbeta3) in the heart, we studied the effects of subtype specific beta-adrenergic receptor (beta-AR) stimulation on Gbeta3 localization using isoform-specific antibodies. The amount of Gbeta3 in the cytosol dramatically decreased in hearts perfused with isoproterenol (ISO) alone or ISO with ICI 118551, a beta2-AR antagonist. Propranolol or CGP 20712A, a beta1-AR antagonist, blocked the ISO-induced decrease in the Gbeta3 content of the cytosol. In contrast, Gbeta3 content of the membrane fraction significantly increased in hearts perfused with ISO alone or ISO with ICI 118551. We conclude that stimulation of the beta1-AR induces isoform-specific translocation of Gbeta3 from the cytosol to the membrane fraction in rat hearts. PMID- 10513994 TI - Role of oxidative stress in nickel chloride-induced cell injury in rat renal cortical slices. AB - Nickel chloride (NiCl2) induced lactate dehydrogenase (LDH) release and lipid peroxidation (LPO) in rat renal cortical slices in vitro in a concentration- (0-2 mM) and time- (0-4 hr) dependent manner, with initial significant LDH release occurring as early as 1 hr, whereas significant increase in LPO started 3 hr after exposure, suggesting that LPO results from renal cell injury. Both NiCl2 induced LDH release and LPO were prevented significantly by glutathione and dithiothreitol, suggesting that NiCl2-induced renal cell injury is dependent on thiols. However, such injury is not dependent solely on thiols, because (a) these thiols failed to inhibit completely the uptake of Ni2+ by the renal cortex, and (b) diethylmaleate pretreatment failed to increase NiCl2-induced cell injury further. Superoxide dismutase partially reduced the NiCl2-induced LDH release without affecting LPO and glutathione, whereas catalase did not affect such LDH release and LPO. Dimethylthiourea and DMSO completely prevented NiCl2-induced LPO, but only partially reduced LDH release. Deferoxamine prevented NiCl2-induced renal cell injury without affecting LPO and without significantly reducing Ni2+ uptake by the renal cortex, suggesting that nickel chelation is not important in such prevention of injury. NiCl2-induced inhibition of para-aminohippurate uptake was prevented significantly by thiols, deferoxamine, and dimethylthiourea. NiCl2 induced loss of cellular glutathione content was prevented significantly by thiols and deferoxamine, but not by superoxide dismutase and dimethylthiourea. These results suggest that LPO was not related to NiCl2-induced lethal renal cell injury, whereas such injury may be caused by the induction of the Fenton reaction, generating hydroxyl radicals. PMID- 10513995 TI - Acetaldehyde increases the activity and gene expression of urokinase type plasminogen activator in a hepatic stellate cell line. AB - The aim of this study was to investigate the effect of acetaldehyde on the activity and expression of urokinase type plasminogen activator gene in a clone of hepatic stellate cells. CFSC-2G cells showed typical morphological changes of the stellate cell activation, which were accompanied by an increase in the amount of collagen with all doses of acetaldehyde used. The treatment of the cells with doses of 100 and 175 micromol/l acetaldehyde, produced an increase in the urokinase type plasminogen activator activity not only in the cell extract, but also in conditioned medium. However, the use of higher doses of acetaldehyde (250 and 350 micromol/l) produced an inhibitory effect on the urokinase type plasminogen activator activity. In contrast, the higher urokinase type plasminogen activator gene expression was observed with doses of 175, 250, and 350 micromol/l. Our results shown that acetaldehyde induced changes in synthesis, release, and expression of urokinase type plasminogen activator in CFSC-2G cells. Those findings suggest that the alterations in the synthesis and expression of the urokinase type plasminogen activator might be another event associated to the activation of hepatic stellate cell after exposure to hepatotoxic agents like acetaldehyde. The role of urokinase type plasminogen activator in fibrogenesis was analyzed. PMID- 10513996 TI - Cadmium-induced testes oxidative damage in rats can be influenced by dietary zinc intake. AB - We tested the hypothesis that zinc deficient animals would be characterized by an increased sensitivity to cadmium-induced oxidative damage to the testes. Weanling male rats were given free access to either a control (25 microg Zn/g) or a zinc deficient (0.5 microg Zn/g) diet; or restricted access to the 25 microg Zn/g diet at a level of intake similar to that of rats fed the 0.5 microg Zn/g diet. After 14 days on the diets, animals were injected s.c. with either saline or CdCl2 (2 mg Cd/kg body weight) solutions, and killed 24 h later. In the zinc-deficient group, testes weight and testes/body weight were higher in the cadmium-injected rats than in the saline-injected rats. The extent of hemorrhages, as indicated by high hemoglobin and testes iron concentrations was higher in the cadmium-treated zinc deficient group than in the cadmium-injected controls. In the zinc-deficient group, cadmium injection was associated with higher levels of lipid peroxidation (33% higher TBARS content) and protein oxidation (17% lower glutamine synthetase activity). Cadmium injection did not influence these parameters in the zinc adequate groups. Extracellular superoxide dismutase activity was lower in the zinc-deficient group than in the zinc-sufficient groups; there was a trend (P < 0.06) for a lower activity in the cadmium- versus the saline-injected rats. These results support the concept that zinc deficiency increases the susceptibility of testes to cadmium-mediated free radical damage. PMID- 10513997 TI - Effect of cisplatin and 6-bromo-6-deoxy-L-ascorbic acid on some biochemical and functional parameters in mice. AB - The results of the present study demonstrate that 6-bromo-6-deoxy-L-ascorbic acid (6-BrAA), an antioxidative derivative of ascorbic acid, is capable of lowering the toxicity of cisplatin, cis-diaminedichloroplatinum (cis-DDP), an anticancerogenic drug. The biological aspects and pharmacological significance of a combined treatment of these two substances were investigated in a mouse model. The results indicate that the effectiveness of 6-BrAA on biological response(s) is strongly dependent on the dose of cis-DDP. Injection of 10 mg/kg body weight (bw) of cis-DDP following pretreatment with 6-BrAA (480 mg/kg bw) enhances the tissue-protecting effect of 6-BrAA and reduces, to some extent, the ensuing nephro-, liver and spleen toxicity. On the other hand, 6-BrAA in animals treated with a higher dose of cis-DDP (15 mg/kg bw) leads to exacerbation of the toxic cis-DDP effect and concurrent loss of the protective potential of 6-BrAA with respect to tissue damage. The exact mechanism(s) of 6-BrAA protection and exacerbation of the toxic cis-DDP effect is unclear, although scavenging or generating of free radicals may play an important role. The results obtained may be of importance in planning the rational use of cis-DDP and 6-BrAA administration in the potential treatment of cancer. PMID- 10513998 TI - Acute immunotoxicity of p-chloronitrobenzene in mice: II. Effect of p chloronitrobenzene on the immunophenotype of murine splenocytes determined by flow cytometry. AB - To evaluate the immunotoxicity of p-chloronitrobenzene (p-CNB), we investigated its effect on the immunophenotype of murine splenocytes. BDF1 male mice were randomly divided into exposed and control groups: the exposed group received p CNB at 300 mg/kg dissolved in olive oil, while the control group received only olive oil, by a single intraperitoneal (i.p.) or subcutaneous (s.c.) injection. On days 3, 5, 7, and 10 after the injection, splenocytes were harvested from both groups, and the following cell phenotypes were quantified by flow cytometry: (1) B cells (CD45R/B220); (2) T cells (CD3e); (3) T-cell subsets (CD4 and CD8a); (4) natural killer (NK) cells (NK-1.1); (5) macrophages (CD11b; Mac-1); (6) nucleated erythrocytes (Ter-119); and (7) dead cells with propidium iodide (PI). The percentages and numbers of B, T, subsets of T (CD4 and CD8), and NK cells in the exposed mice significantly decreased as compared with the respective control. On the other hand, macrophages (Mac-1+ cells), nucleated erythrocytes (Ter-119+ cells), and dead cells in the exposed mice markedly increased as compared with the respective control after i.p. injection of p-CNB. The above findings indicate that p-CNB has an immunotoxic effect on mice. PMID- 10514000 TI - Multivariate statistical analysis of organ weights in toxicity studies. AB - Toxicity studies with drugs in animals are performed to establish the toxicity profile including the no-toxic-effect-level in the animals. One of the responses used in the search for effects is organ weight. Since organ weight is related to body weight of the animal, the analysis of organ weight has to be adjusted. A traditional analysis of covariance with body weight as covariate is not always appropriate since the body weight itself can be affected by the test compound. Hence, a multivariate analysis of variance is suggested, where the correlation between organ weight and body weight is taken into account in testing an overall treatment effect. Two dimensional plots of the contour curves are used to visualize the treatment effect on organ weight and body weight simultaneously. The test procedure suggested for comparing differences between control and treated groups of animals is: start by testing hypotheses of equality of covariance matrices for the different treatment groups and then, depending on the results from the first test, test equality of the mean vectors. PMID- 10513999 TI - Comparison of detection sensitivity of immuno- and genotoxicological effects of subacute cypermethrin and permethrin exposure in rats. AB - Immuno- and genotoxicological effects of a 28-day oral treatment by equitoxic (1/10, 1/25, 1/50 LD50) doses of cypermethrin (55.4, 22.2, and 11.1 mg/kg) and permethrin (125.7, 50.3, and 12.6 mg/kg) were compared on male Wistar rats. Humoral and cell-mediated immunity were investigated by PFC assay and delayed type hypersensitivity (DTH) reaction (footpad swelling assay), and the genotoxic effects were studied by structural and numerical chromosome aberrations in bone marrow cells. The experimental system also involved certain general toxicological (body weight gain, organ weights) and haematological [white blood cell (WBC), red blood cell (RBC), haematocrit (Ht) and cell content of the femoral bone marrow] investigations. Among the immune function assays, only DTH reaction decreased at the two higher cypermethrin (CY) doses. These doses also increased the number of numerical chromosome aberrations of the bone marrow cells but did not change the number of structural aberrations. All CY doses decreased the mean cell volume (MCV) of RBCs and the Ht value. The two higher doses also reduced the WBC count in the peripheral blood. Permethrin (PE), in the applied dose range, had no effect on the examined immune function parameters, but all three doses increased the number of numerical chromosome aberrations. A dose-dependent increase in the liver weight, decreased MCV value, and elevated cell content of the femoral bone marrow were also observed. Under these experimental conditions, examination of chromosome aberrations proved to be less sensitive in detection of exposure by cypermethrin than applied immune function assays did. Permethrin, on the contrary, increased the number of numeric aberrations at all dose levels but had no effect on the immune function parameters examined. PMID- 10514002 TI - A physiological model for ligand-induced accumulation of alpha 2u globulin in male rat kidney: roles of protein synthesis and lysosomal degradation in the renal dosimetry of 2,4,4-trimethyl-2-pentanol. AB - A physiologically based pharmacokinetic (PBPK) model was constructed for the disposition of 2,4,4-trimethyl-2-pentanol (TMP-2-OH) in male rats and its induction of accumulation of renal alpha2u-globulin (alpha2u). The model included diffusion-restricted delivery of TMP-2-OH to compartments representing liver, lung, fat, kidney, GI tract, aggregated rapidly perfused tissues, and aggregated slowly perfused tissues. Metabolism by oxidation and glucuronidation was included for liver and kidneys. Rates of hepatic alpha2u production and resorption by renal proximal tubules were taken from the literature. Degradation of liganded alpha2u by renal lysosomal cathepsins was modeled with a Km value corresponding to the measured 30% reduction in proteolytic efficiency and with free and bound forms of alpha2u competing for access to the enzymes. Increased pinocytotic uptake of alpha2u into the kidney induces cathepsin activity. A model that ascribed renal alpha2u accumulation solely to reduced lysosomal proteolysis failed to reproduce the observed accumulation. The model could reproduce experimental observations if a transient increase in hepatic synthesis of alpha2u, stimulated by the presence of liganded alpha2u in the blood, and accelerated secretion of the protein from the liver were assumed. This model reproduces time course data of blood and kidney TMP-2-OH and renal alpha2u concentrations, suggesting that renal accumulation of alpha2u is not simply a consequence of reduced proteolytic degradation but may also involve a transient increase in hepatic alpha2u production. The model predicts increased delivery of TMP-2-OH to the kidney and consequent increased renal production of potentially toxic TMP-2-OH metabolites than would be the case if no alpha2u were present. Induced lysosomal activity and increased production of toxic metabolites may both contribute to the nephrotoxicity observed in male rats exposed to an alpha2u ligand or its precursor. PMID- 10514001 TI - Distribution of gamma-tubulin in multipolar spindles and multinucleated cells induced by dimethylarsinic acid, a methylated derivative of inorganic arsenics, in Chinese hamster V79 cells. AB - Localization of gamma-tubulin, a well-characterized component of microtubule organizing centers (MTOCs), was investigated because of interest in the mechanism of the induction of aberrant mitotic spindles in Chinese hamster V79 cells exposed to dimethylarsinic acid (DMAA). In control cultures, gamma-tubulin in interphase cells was located as a perinuclear dot on which the microtubules were nucleated. In metaphase cells, the location of gamma-tubulin coincided with that of the mitotic spindle poles. DMAA caused mitotic delay and aberrant spindles, such as tripolar- and quadripolar spindles, in the mitotic cells. Gamma-tubulin was co-localized with the aberrant spindles induced by DMAA. The incidence of gamma-tubulin in the mitotic cells coincided with that of the aberrant spindles and rose with an increasing concentration of DMAA. By contrast, DMAA did not influence the number and location of gamma-tubulin signals in interphase cells. These results suggest that multiple microtubule nucleation sites were induced by DMAA during transition from interphase to mitotic phase. DMAA-induced multiple signals of gamma-tubulin were integrated into one signal at the center of multinucleated cells, surrounded by multiple nuclei as the cell cycle progressed to the next interphase, suggesting the presence of a self-integration mechanism of centrosomal MTOCs during the cell cycle. PMID- 10514003 TI - PCB congener 126-induced ultrastructural alterations in the rat liver: a stereological study. AB - Hepatocyte cytoplasmic alterations were morphometrically determined in male and female Sprague-Dawley rats fed PCB congener 126 (3,3',4,4',5-pentachlorobiphenyl) in concentrations of 0.1, 1.0, 10, 100 ppb or corn oil in diets for 13 weeks. A dose-dependent increase (P < 0.05) in the volume fraction of smooth endoplasmic reticulum (SER) and mitochondria was measured in the hepatocytes of the females. However, these cells of the male rats contained a significantly greater baseline volume fraction of SER compared to that in the females. Statistical differences were not detected in the volume fractions of rough endoplasmic reticulum, peroxisomes or lipid droplets of the hepatocytes in either the males or females. We conclude the increase in mitochondrial volume was a necessary cellular adaptation to meet the heightened energy demands by the SER to produce the necessary enzymes to detoxify the PCB. Morphometric analysis rather than a descriptive methodology allowed for a more accurate determination of the liver pathology induced by PCB 126. PMID- 10514004 TI - The makings of a tumor rejection antigen. PMID- 10514005 TI - Interleukin-2 expression by a subpopulation of primary T cells is linked to enhanced memory/effector function. AB - Single cell studies have identified intraclonal heterogeneity of cytokine production by activated T cells. To investigate implications of cytokine heterogeneity for cell fate, an interleukin (IL)-2 promoter-green fluorescent protein (GFP) reporter transgenic model was developed to track IL-2+ and IL-2- T cells during differentiation from naive precursors. Antigen-activated IL-2+ and IL-2- cells had comparable proliferative capacities in primary responses. However, T cells that expressed IL-2 in primary responses demonstrated enhanced antigenic sensitivity and increased expression of effector cytokines in secondary responses in vitro and in vivo. Thus, heterogeneity of activation during a primary response translates into heterogeneous secondary responses, in which enhanced memory/effector function is linked to cells that previously exceeded an activation threshold associated with IL-2 gene transcription. PMID- 10514006 TI - Uncoupling IL-2 signals that regulate T cell proliferation, survival, and Fas mediated activation-induced cell death. AB - IL-2 is an important growth and survival factor for T lymphocytes but also sensitizes these cells to Fas-mediated activation-induced cell death (AICD). The molecular basis of these different effects of IL-2 was studied by introducing wild-type and mutant forms of the IL-2 receptor beta (IL-2Rbeta) chain that lacked specific signaling capacities into receptor-deficient T cells by retroviral gene transfer. Activation of Stat5 by IL-2 was found to be involved in T cell proliferation and promoted Fas ligand (FasL) expression and AICD. T cell survival was dependent on a receptor region that activated Akt and the expression of Bcl-2. Thus, distinct IL-2Rbeta chain signaling modules regulate T cell fate by stimulating growth and survival or by promoting apoptosis. PMID- 10514007 TI - Cross-antagonism of a T cell clone expressing two distinct T cell receptors. AB - Inhibition of T cell activation can be mediated by analogs of the original antigenic peptide (TCR antagonists). Here, a T cell clone expressing two distinct TCR was used to investigate whether such inhibition involves an active mechanism by examining whether an antagonist for one TCR could influence responses stimulated by the other TCR engaging its agonist. Our results demonstrate functional cross-inhibition under these conditions involving the ability of antagonist: TCR interactions to diminish Lck enzymatic activity associated with the agonist-recognizing second TCR, apparently through enhancement of SHP-1 association with these receptors. Our findings reveal that inhibition of cellular responses by antagonists arises at least in part from active negative regulation of proximal TCR signaling and identify elements of the biochemical process. PMID- 10514008 TI - Notch1 expression in early lymphopoiesis influences B versus T lineage determination. AB - Notch receptors regulate fate decisions in many cells. One outcome of Notch signaling is differentiation of bipotential precursors into one cell type versus another. To investigate consequences of Notch1 expression in hematolymphoid progenitors, mice were reconstituted with bone marrow (BM) transduced with retroviruses encoding a constitutively active form of Notch1. Although neither granulocyte or monocyte differentiation were appreciably affected, lymphopoiesis was dramatically altered. As early as 3 weeks following transplantation, mice receiving activated Notch1-transduced BM contained immature CD4+ CD8+ T cells in the BM and exhibited a simultaneous block in early B cell lymphopoiesis. These results suggest that Notch1 provides a key regulatory signal in determining T lymphoid versus B lymphoid lineage decisions, possibly by influencing lineage commitment from a common lymphoid progenitor cell. PMID- 10514009 TI - Four of five RAG-expressing JCkappa-/- small pre-BII cells have no L chain gene rearrangements: detection by high-efficiency single cell PCR. AB - Single cell PCR assays have been further developed that detect over 80% of all VkappaJkappa, VkappaRS, and VlambdaJlambda rearrangements at efficiencies between 70% and 90%. These IgL chain gene rearrangement assays were used with small pre BII cells that develop in comparably high numbers in the bone marrow of wild type, Ckappa-deficient, and JCkappa-deficient homozygous and heterozygous mice. In all of these mice, only 15%-25% of all small pre-BII cells carry VlambdaJlambda rearrangements. These results confirm that lambdaL chain gene rearrangements occur independently of kappaL chain gene rearrangement and expression. They also show that a large part of the small pre-BII cells that express the rearrangement machinery can develop without IgL chain gene rearrangements. PMID- 10514010 TI - Frequencies of multiple IgL chain gene rearrangements in single normal or kappaL chain-deficient B lineage cells. AB - PCR analyses of the kappaL chain locus in single B-lineage cells of wild-type, Ckappa-, or JCkappa-deficient homozygous or heterozygous mice often detect multiple in- and out-of-frame rearrangements at the kappaL and lambdaL loci. They are most frequent in small pre-BII cells and equally so in wild-type and kappaL chain-deficient cells. Hence, kappaL chain production appears not to inhibit secondary rearrangements. Around 20% of all small preBII cells express IgL chains in their cytoplasm. Cells with a first productive rearrangement on one allele are favored to enter the immature B cell compartment. Thus, allelic exclusion might be secured by control of accessibility of IgL chain loci for rearrangement and by rapid selection of cells with a fitting over those with a nonfitting IgL chain. PMID- 10514011 TI - Junctional amino acids determine the maturation pathway of an antibody. AB - We found that two distinct antibody maturation pathways exist in the immune response of C57BL/6 mice to (4-hydroxy-3-nitrophenyl)acetyl and that the junctional amino acid introduced by a process far preceding somatic hypermutation determined the pathway of affinity maturation. Antibodies belonging to each pathway clearly separated into two separate branches of a phylogenic tree. We also constructed a three-dimensional fitness landscape for antibody evolution by introducing the association constants of the antibodies into the phylogenic tree as the third axis, allowing us to comprehend the significance of junctional diversity in the "evolvability" of antibodies. Thermodynamic analyses of the antigen-antibody interactions suggested that a high conformational versatility in the antigen-combining site allows for the enhanced evolvability of antibodies. PMID- 10514013 TI - Autoimmune intestinal pathology induced by hsp60-specific CD8 T cells. AB - Due to their ubiquitous distribution and high degree of structural similarity, heat shock proteins (hsp) are potential target antigens in autoimmune diseases. Here, we describe induction of intestinal inflammation following transfer of hsp60-reactive CD8 T cells into mice. Inflammatory reactions were MHC class I dependent and developed primarily in the small intestine. IFN gamma and TNF alpha, as well as gut-derived hsp60, were elevated at sites of T cell infiltration. Intestinal lesions were drastically reduced in mice lacking receptors for TNF alpha. Pathology also developed in germ-free mice, indicating recognition of host-derived hsp60 by CD8 T cells. This report demonstrates that CD8 T cells with defined antigen specificity cause intestinal inflammation, emphasizing a link between infection and autoimmune disease. PMID- 10514012 TI - Ku in the cytoplasm associates with CD40 in human B cells and translocates into the nucleus following incubation with IL-4 and anti-CD40 mAb. AB - CD40 plays a critical role in survival, growth, differentiation, and class switching of B lymphocytes. Although Ku is required for immunoglobulin class switching, how CD40 signal transduction is coupled to Ku is still unknown. Here, we show that CD40 directly interacts with Ku through the membrane-proximal region of cytoplasmic CD40. Ku was confined to the cytoplasm in human primary B cells, and the engagement of CD40 on the B cells cultured in the presence of IL-4 resulted in the dissociation of Ku from CD40, translocation of Ku into the nucleus, and increase in the activity of DNA-dependent protein kinase. These findings indicate that Ku is involved in the CD40 signal transduction pathway and may play an important role in the CD40-mediated events. PMID- 10514014 TI - Caspase-1-independent, Fas/Fas ligand-mediated IL-18 secretion from macrophages causes acute liver injury in mice. AB - IL-18, produced as a biologically inactive precursor, is processed by caspase-1 in LPS-activated macrophages. Here, we investigated caspase-1-independent processing of IL-18 in Fas ligand (FasL)-stimulated macrophages and its involvement in liver injury. Administration of Propionibacterium acnes (P. acnes) upregulated functional Fas expression on macrophages in an IFNgamma-dependent manner, and these macrophages became competent to secrete mature IL-18 upon stimulation with FasL. This was also the case for caspase-1-deficient mice. Administration of recombinant soluble FasL (rFasL) after P. acnes priming induced comparable elevation of serum IL-18 in parallel with elevated serum liver enzyme levels. However, liver injury was not induced in IL-18-deficient mice after rFasL administration. These results indicate a caspase-1-independent pathway of IL-18 secretion from FasL-stimulated macrophages and its critical involvement in FasL induced liver injury. PMID- 10514015 TI - PSGL-1-mediated adhesion of human hematopoietic progenitors to P-selectin results in suppression of hematopoiesis. AB - Cellular interactions are critical for the regulation of hematopoiesis. The sialomucin PSGL-1/CD162 mediates the attachment of mature leukocytes to P selectin. We now show that PSGL-1 also functions as the sole receptor for P selectin on primitive human CD34+ hematopoietic progenitor cells (HPC). More importantly, ligation of PSGL-1 by immobilized or soluble ligand or anti-PSGL-1 antibody results in a profound suppression of HPC proliferation stimulated by potent combinations of early acting hematopoietic growth factors. These data demonstrate an unanticipated but extremely marked growth-inhibitory effect of P selectin on hematopoiesis and provide direct evidence that PSGL-1, in addition to its well-documented role as an adhesion molecule on mature leukocytes, is a potent negative regulator of human hematopoietic progenitors. PMID- 10514016 TI - TRAF2 deficiency results in hyperactivity of certain TNFR1 signals and impairment of CD40-mediated responses. AB - Tumor necrosis factor (TNF) receptor-associated factor 2 (TRAF2) can interact with various members of the TNF receptor family. Previously, we reported that TRAF2-deficient mice die prematurely and have elevated serum TNF levels. In this study, we demonstrate that TRAF2-deficient macrophages produce increased amounts of nitric oxide (NO) and TNF in response to TNF stimulation. Furthermore, we could enhance the survival of TRAF2-deficient mice by eliminating either TNF or TNFR1. Using these double-knockout mice, we show that in the absence of TRAF2, the T helper-dependent antibody response, CD40-mediated proliferation, and NF kappaB activation are defective. These data demonstrate two important roles of TRAF2, one as a negative regulator of certain TNFR1 signals and the other as a positive mediator of CD40 signaling. PMID- 10514017 TI - Leukotriene modifiers and Churg-Strauss syndrome: adverse effect or response to corticosteroid withdrawal? AB - Zafirlukast, montelukast and pranlukast are all cysteinyl leukotriene receptor antagonists that have recently been approved for the treatment of asthma. Within 6 months of zafirlukast being made available on the market, 8 patients who received the agent for moderate to severe asthma developed eosinophilia, pulmonary infiltrates, cardiomyopathy and other signs of vasculitis; the syndrome that these patients developed was characteristic of the Churg-Strauss syndrome. All of the patients had discontinued systemic corticosteroid use within 3 months of presentation and all developed the syndrome within 4 months of zafirlukast initiation. The syndrome dramatically improved in each patient upon reinitiation of corticosteroid therapy. Since the initial report, there have been multiple similar cases reported to the relevant pharmaceutical companies and to federal drug regulatory agencies in association with zafirlukast as well as with pranlukast, montelukast, and with use of high doses of inhaled corticosteroids, thus leading to an increased incidence rate of the Churg-Strauss syndrome. Many potential mechanisms for the association between these drugs and the Churg Strauss syndrome have been postulated including: increased syndrome reporting due to bias; potential for allergic drug reaction; and leukotriene imbalance resulting from leukotriene receptor blockade. However, careful analysis of all reported cases suggests that the Churg-Strauss syndrome develops primarily in those patients taking these asthma medications who had an underlying eosinophilic disorder that was being masked by corticosteroid treatment and unmasked by novel asthma medication-mediated corticosteroid withdrawal, similar to the forme fruste of the Churg-Strauss syndrome. It remains unclear what the exact mechanism for this syndrome is and whether this represents an absolute increase in cases of vasculitis, but it appears that none of the asthma medications implicated in leading to the development of Churg-Strauss syndrome was directly causative of the syndrome. These agents remain well tolerated and effective medications for the treatment of asthma, although physicians must be wary for the signs and symptoms of the Churg-Strauss syndrome, particularly in patients with moderate to severe asthma in whom corticosteroids are tapered. PMID- 10514019 TI - Zanamivir: a review of clinical safety. AB - Preclinical and clinical studies have clearly demonstrated that zanamivir, a potent and highly selective inhibitor of the influenza A and B virus neuraminidase, has an impressive safety profile. This report describes the safety and tolerability findings from the clinical studies completed up to the 17 July 1998 involving over 6000 adult and adolescent patients from North America, Europe and the Southern Hemisphere. Serious adverse events from an ongoing Japanese clinical programme are also reported. Zanamivir was administered in various dose forms and frequencies and was found to have a comparable safety profile with placebo when given for both the treatment and prophylaxis of influenza-like illness. These findings were independent of age and underlying medical condition. 4152 patients received zanamivir and the most commonly reported adverse events were consistent with the signs and symptoms of influenza-like illness. Most of the adverse events were mild and did not result in patient withdrawal from the studies. Less than 1% of zanamivir and placebo recipients reported a serious adverse event. In addition, 490 healthy volunteers received zanamivir in clinical pharmacology studies. It was well tolerated and the incidence of adverse events was similar in zanamivir and placebo recipients. In addition, no clinically significant laboratory abnormalities were detected. Results from in vitro and in vivo animal studies suggest that zanamivir has low acute toxicity and no significant systemic toxicity or respiratory tract irritancy at plasma exposures more than 100-fold higher than those anticipated following clinical use. Neither genotoxic nor reproductive types of toxicity have been observed in toxicology studies at doses equal to 17 to 197 times the current therapeutic dose (20 mg/day). The characteristics of the molecule and the low systemic exposure indicate a very low potential for drug interactions with the inhaled route. Furthermore, repeated 600mg intravenous doses were well tolerated in healthy volunteers. The observed safety profile of zanamivir compares favourably with currently available agents with anti-influenza virus activity, such as rimantadine and amantadine, as well as GS4104, a neuraminidase inhibitor currently in phase III development. This may be attributed to the low systemic bioavailability of zanamivir, which is given by oral inhalation, direct to the primary site of viral replication. The potential advantages of this include a reduced risk of drug-drug interactions, other nontarget organ toxicities (e.g. brain) and drug clearance issues from both kidney and liver. Therefore, the safety profile of zanamivir supports its use in the management of influenza. PMID- 10514018 TI - Antioxidant vitamin supplements: update of their potential benefits and possible risks. AB - Oxidative damage to biological structures has been implicated in the pathophysiology of cardiovascular disease and cancer, the 2 most common causes of death in developed countries. This has stimulated interest in the possible role of natural antioxidant vitamins in preventing the development of these diseases. Epidemiological studies have offered support for the notion that high blood concentrations or dietary intake of antioxidant vitamins may have a protective effect. On the basis of these findings and powerful marketing strategies, many healthy members of the population are now voluntarily consuming antioxidant supplements. A number of long term, prospective, randomised, placebo-controlled trials examining the protective effect of antioxidant supplements have now been completed. Their results have been generally disappointing and have provided little evidence of efficacy. Of greater concern, they have unexpectedly raised concerns that antioxidants, notably betacarotene, might increase the rate of development of cancers in high risk individuals. For this reason regular consumption of antioxidant vitamins supplements cannot yet be advocated as a healthy lifestyle trait. PMID- 10514020 TI - Risk-benefit assessment of opioids in chronic noncancer pain. AB - Opioids have been accepted as appropriate treatment for acute and cancer pain, but their role in the management of chronic nonmalignant pain is the subject of much debate, mainly due to concerns about waning efficacy, the potential for neuropsychological impairment and the development of drug addiction. Controlled clinical trials demonstrated that opioids may be effective in both nociceptive and neuropathic noncancer pain, although the former responded more consistently than the latter. Gastrointestinal and CNS adverse effects were frequent in most studies. Observational studies have generated contradictory findings regarding efficacy and safety as well as the risk of drug addiction in patients with chronic noncancer pain receiving long term opioid therapy. However, they suggest that opioids may be effective in individual cases, whichever the pathophysiological mechanism of pain. Taken together, the available data indicate that the outcomes associated with opioid therapy vary markedly across patients experiencing chronic nonmalignant pain. The main consensus is that a subset of these patients may gain substantial benefit from opioid analgesics without requiring rapidly escalating doses or developing intolerable adverse effects or drug addiction. Prescribing guidelines have been developed to assist practitioners in selecting the appropriate patients and ensuring an acceptable risk : benefit ratio of opioid therapy. Finally, it must be emphasised that chronic pain is a complex entity wherein analgesics, including opioids, are only part of the treatment. PMID- 10514021 TI - Risks and benefits of aromatase inhibitors in postmenopausal breast cancer. AB - Aromatase inhibitors were first reported in the early 1970s and have been used to treat breast cancer since that time. Until recently, essentially the only agent available in this class was aminoglutethimide, a nonspecific inhibitor with multiple adverse effects and drug interactions. Selective and potent aromatase inhibitors are now available (formestane, exemestane, fadrozole, anastrozole and letrozole), and we review the risks and benefits of these agents in order to assist clinicians in making treatment decisions. Formestane is an injectable steroidal aromatase inhibitor with significant activity against metastatic breast cancer. It has been shown to have similar efficacy and superior tolerability compared with megestrol, and is similar to tamoxifen in the metastatic setting. Exemestane is an oral steroidal aromatase inhibitor. It has been shown to be effective third-line therapy after tamoxifen and megestrol in postmenopausal patients with metastatic breast cancer. All the nonsteroidal (imidazole/triazole) aromatase inhibitors are orally available. Fadrozole has similar activity to megestrol and tamoxifen in the setting of metastasis, but has been shown in phase II trials to inhibit cortisol and aldosterone production. Anastrozole and letrozole have similar toxicity profiles. Compared with megestrol, anastrozole improves overall survival and has superior tolerability. Letrozole is superior to megestrol and aminoglutethimide in terms of overall survival and time to progression, and is also better tolerated. Although there is a strong rationale for using these agents in the treatment of breast cancer, the information presently available is insufficient to recommend any one agent over another. Direct comparative studies are lacking, and comparing agents across studies is limited by many biases and may not be valid. Formestane is only available as an injection and exemestane is not commercially available in many countries, making these agents more difficult to recommend over the other 3 agents. Fadrozole is less potent and less selective in inhibiting aromatase than letrozole. The efficacies of fadrozole, megestrol and tamoxifen appear to be similar; however, comparative data show no advantage of fadrozole over letrozole. Anastrozole and letrozole are generally considered to be similar agents. The clinical future of the selective aromatase inhibitors is promising, and these agents may change the way postmenopausal breast cancer is treated at all stages of the disease. PMID- 10514023 TI - Comparative safety and tolerability of clopidogrel and aspirin: results from CAPRIE. CAPRIE Steering Committee and Investigators. Clopidogrel versus aspirin in patients at risk of ischaemic events. AB - OBJECTIVE: The objective of this study was to provide a comprehensive comparison of the long term safety and tolerability of clopidogrel, a new adenosine diphosphate (ADP) receptor antagonist that inhibits platelet activation induced by ADP, and aspirin (acetylsalicylic acid). PATIENTS AND METHODS: The study population comprised 19,185 patients with symptomatic atherosclerosis manifested as recent ischaemic stroke, recent myocardial infarction or symptomatic peripheral arterial disease. Patients were randomised to receive clopidogrel 75 mg/day or aspirin 325 mg/day for a minimum of 1 year and a maximum of 3 years. RESULTS: Compared with aspirin, clopidogrel reduced the combined risk of ischaemic stroke, myocardial infarction or vascular death by 8.7% (p = 0.043). The incidence of early permanent discontinuations of the study drug due to adverse events was almost identical in both treatment groups (11.94% for clopidogrel vs 11.92% for aspirin). Reported neutropenia was similar in the clopidogrel and aspirin groups (0.10 vs 0.17%, respectively) with corresponding rates (0.05 vs 0.04%, respectively) for severe neutropenia. Thrombocytopenia was identical in the clopidogrel and aspirin groups (0.26%), with the rates of severe thrombocytopenia being 0.19 vs 0.10%, respectively. None of these observed differences was statistically significant. The overall incidence of haemorrhagic events did not differ statistically significantly between treatment groups (9.27% for clopidogrel vs 9.28% for aspirin; p = 0.98). There was a trend towards a lower incidence of intracranial haemorrhage in the clopidogrel group (0.31%) compared with the aspirin group (0.42%). Any reported gastrointestinal haemorrhage was significantly less frequent with clopidogrel (1.99%) than with aspirin (2.66%) [p < 0.002]. The corresponding data for severe gastrointestinal bleeding were 0.49 vs 0.71%; p < 0.05. Overall, there were significantly fewer gastrointestinal adverse events with clopidogrel than with aspirin (27.1 vs 29.8%; p < 0.001), with less abdominal pain, dyspepsia, constipation, or peptic, gastric, or duodenal ulceration with clopidogrel. Diarrhoea was significantly more common in the clopidogrel group (4.46 vs 3.36%; p < 0.001), although the incidence of severe diarrhoea (0.23 vs 0.11%) was low and was not significantly different between groups. There were significantly more patients with rash in the clopidogrel group (6.0%) compared with the aspirin group (4.6%) [p < 0.001]. However, these events were generally mild and transient in nature. CONCLUSION: Given the favourable benefit/risk ratio, clopidogrel represents a clinically important advance in the treatment of patients with manifest atherosclerotic disease. PMID- 10514024 TI - Postmarketing studies: the work of the Drug Safety Research Unit. PMID- 10514025 TI - Effect of lycopene on lipid peroxidation and glutathione-dependent enzymes induced by T-2 toxin in vivo. AB - Lycopene, obtained from fresh tomatoes, was incorporated into the chicks diet. The treatments were: (1) Control, (2) 1.5 mg T-2 toxin/kg body weight/day; (3) 25 mg lycopene/kg body weight/day, (4) 1.5 mg T-2 toxin plus 25 mg lycopene/kg body weight/day. Male broiler chicks, 7-28 days of age, were provided with feed and water ad libitum. Every 7 days, malondialdehyde (MDA) and glutathione (GSH) levels, and enzymatic activities of glutathione-S-transferase (GST), gamma glutamyltransferase (GGT) and glutathione peroxidase (GP) were evaluated in liver homogenates. Compared to the controls after 7 days of treatment, T-2 toxin increased hepatic MDA concentration (128%). A significant consumption of endogenous antioxidant GSH (45%) was induced as well as a marked increase in hepatic enzymatic activities of GST, GGT, and GP (312, 187, and 324%, respectively). Addition of T-2 plus lycopene, at an approximate ratio of 1:17 in the diet, diminished some parameters measured (P < 0.05). Apparently lycopene participated as an antioxidant agent and also protecting the cellular level of GSH. PMID- 10514026 TI - Co-administration of toluene and xylene antagonized the testicular toxicity but not the hematopoietic toxicity caused by ethylene glycol monoethyl ether in Sprague-Dawley rats. AB - Occupational painters are exposed to ethylene glycol monoethyl ether (EGEE), a widely used emulsifying solvent known to cause testicular degeneration and bone marrow depression, together with toluene (TOL) and xylene (XYL) as a mixture. In the previous study (Chung et al., Tox. Lett. 104:143, 1999), testicular atrophy caused by EGEE (200 mg/kg) was shown to be antagonized by co-administration of TOL (250 mg/kg) and XYL (500 mg/kg). This study was conducted to provide histological support for the previously observed antagonistic protective effect of TOL + XYL on EGEE inducible testicular toxicity and to determine whether a similar antagonistic effect can be demonstrated against the EGEE derived hematopoietic toxicity. Compared to the extent of seminiferous tubule degeneration caused by EGEE (150 mg/kg, six times per week for 4 weeks), testes of rats given co-administration of TOL (250 mg/kg) + XYL (500 mg/kg) showed dramatically reduced tubular degeneration. Hyperplasia of Leydig cells in the interstitium was observed in both EGEE and EGEE + TOL + XYL-treated rats. Although a minimal dose of EGEE causing testicular atrophy was used, WBC and platelet counts were decreased significantly. In the TOL + XYL-treated control group, the WBC and platelet counts were not decreased. However, the bone marrow depression caused by EGEE was not reversed by the combined administration of TOL + XYL. In all experimental groups (EGEE alone, TOL + XYL, EGEE + TOL + XYL), plasma levels of creatinine and alkaline phosphatase were significantly decreased. In addition to the marked testicular atrophy, EGEE also decreased the weights of adrenal glands and epididymis. In conclusion, while the testicular degeneration caused by EGEE was antagonized by TOL + XYL, the EGEE derived hematopoietic suppression was not reversed. PMID- 10514022 TI - Treating inflammatory bowel disease during pregnancy: risks and safety of drug therapy. AB - The safety of drug therapy for inflammatory bowel disease during pregnancy is an important clinical concern. Current available information is largely derived from animal studies and clinical experience among patients with inflammatory bowel disease and autoimmune disorders and organ transplant recipients. However, these data are confounded by various factors including difficulty projecting the results of animal studies to humans, methodological deficiencies of some studies, insufficient experience with certain agents, difficulty distinguishing the fetal effects of underlying disease from drug therapy and a need to consider the impact of background rates of adverse fetal outcomes which apply to all pregnancies. In inflammatory bowel disease, the effects of active inflammation on the fetus are believed to be more harmful than those of drug treatment, and therapy is often justified to induce or maintain remission during pregnancy. The choice of appropriate treatment is determined by the severity of the disease and the potential for drug toxicity. No causal relationship has been established between exposure to sulfasalazine or other 5-aminosalicylic acid drugs and the development of congenital malformations. These drugs may be used with relative safety during pregnancy and lactation. Considerable experience with corticosteroids have shown them to pose very small risk to the developing fetus. Current evidence indicates that maternal use of azathioprine is not associated with an increased risk of congenital malformations, though impaired fetal immunity, growth retardation or prematurity is occasionally observed. Preliminary evidence derived from patients with inflammatory bowel disease show no significant fetal toxicity following first trimester exposure to mercaptopurine, though its elective use in pregnancy is controversial. Cyclosporin is not teratogenic, but may be associated with growth retardation and prematurity. Pregnancy should be avoided in women treated with methotrexate because of its known abortifacient effects and risk of causing typical malformations. Although treatment with metronidazole or ciprofloxacin for short durations appear to be devoid of adverse fetal reactions, the effect of prolonged exposure as required in Crohn's disease remains unknown. PMID- 10514027 TI - Evidence that nicotine acetylcholine receptors are not the main targets of cotinine toxicity. AB - The toxicity of nicotine, cotinine and their mixtures was studied in Mus musculus mice as well their effects on growth after repetitive administration to young mice. The affinity constants of the two alkaloids for the nicotinic acetylcholine receptors (nAChRs) of Torpedo and rat brain membranes were determined. The administration of these alkaloids produced distinct symptoms of intoxication. Nicotine was 100-fold more toxic than cotinine and 10-fold more rapid than cotinine at producing respiratory arrest. The affinity of nicotine for both subtypes of nAChRs was > 100-fold higher than that of cotinine. Repetitive administrations of nicotine caused weight loss, whereas that of cotinine caused weight gain (P < 0.01). The administration of the two alkaloids as mixtures to mice caused significantly (P < 0.01) higher mortality than theoretically expected. Furthermore, hexamethonium pretreatment reduced by 2-fold (P < 0.01) the toxicity of nicotine but enhanced by 1.6-fold (P < 0.01) that of cotinine and was without effects on toxicity of mixtures. We suggest that nAChRs are not the main targets of cotinine toxicity. PMID- 10514028 TI - Correlation between cell survival, micronuclei-induction, and LDH activity in V79 cells treated with teniposide (VM-26) before exposure to different doses of gamma radiation. AB - The influence of teniposide (VM-26) treatment was studied on the radiation induced alterations in cell survival, micronuclei (MN) formation and lactate dehydrogenase (LDH) release in V79 cells. Treatment of V79 cells with 10 nM teniposide before exposure to different doses of gamma radiation resulted in a significant decline in the cell survival when compared with the PBS + irradiation group. The decline in cell survival was dose related. The cell proliferation indices also declined in a dose-dependent manner in both PBS + irradiation and VM 26 + irradiation groups. The decline was higher in the VM-26 + irradiation group in comparison with the PBS + irradiation group. In contrast, the frequency of micronuclei increased in a dose-related manner in both PBS + irradiation and VM 26 + irradiation groups. However, the frequency of micronuclei was significantly greater in the latter group when compared with the former group at all the post irradiation time periods studied. The LDH contents increased in a dose-dependent manner in both PBS + irradiation and VM-26 + irradiation groups at all the post irradiation time periods evaluated. This elevation in LDH contents was significantly greater in the VM-26 + irradiation group in comparison with the PBS + irradiation group. PMID- 10514029 TI - Changes in neurotransmitter receptors and neurobehavioral variables in rats co exposed to lead and ethanol. AB - The influence of lead (10 mg/kg) and ethanol (10% v/v, in drinking water) administration, either alone or in combination for 8 weeks was investigated on the uptake of lead in tissues, dopamine, benzodiazepine and cholinergic (muscarinic) receptors, motor activity, number of fighting episodes and several selected lead-sensitive biochemical indices in young rats. Lead or ethanol treatment did not elicit any appreciable influence on body weight gain, tissue weight, and food intake. However, a decreased amount of mean water intake was noticed in animals ingesting ethanol plus lead. Both lead and ethanol when administered individually exacerbate the decrease of blood delta-aminolevulinic acid dehydratase (ALAD) activity while lead and ethanol administered together exhibited a pronounced inhibition of ALAD activity. A significant increase in hepatic GSH levels, hepatic and brain MDA levels was also observed in animals co exposed to lead and ethanol compared to lead alone treated rats. Simultaneous exposure to lead and ethanol also resulted in a significantly more pronounced decrease in binding of [3H]fluintrazepam in membranes prepared from the fronto cortical region compared to the corresponding controls. The binding of [3H]spiroperidol to striatal and [3H]quinuclidinyl benzylate (QNB) to cerebellar membranes remained unaltered in all the exposed animals. Spontaneous locomotor activity and aggressive behaviour increased significantly in the group treated with both lead and ethanol compared to the control group. The lead concentrations in blood, liver, kidney and brain were significantly higher in rats exposed simultaneously to lead and ethanol compared to lead alone exposed group. The results indicate more pronounced neurotoxic and neurobehavioural changes in animals co-exposed to lead and ethanol; however, the exact mechanism or site of its action is still not clear. PMID- 10514030 TI - Tumor necrosis factor is involved in chlorpyrifos--induced changes in core temperature in the female rat. AB - Chlorpyrifos (CHP), an OP-based pesticide, induces hypothermia in the rat followed by a fever that persists for several days. The cytokine, tumor necrosis factor-alpha (TNF), is induced by lipopolysaccharide (LPS) and released during fever and has both pyrogenic and cryogenic (i.e. antipyretic) properties. Administering antibodies to TNF (anti-TNF) is known to disrupt fever from infection. Thus, the purpose of this study was to examine whether anti-TNF also disrupts CHP-induced changes in body temperature of the female Long-Evans rat. A positive effect would suggest a role of TNF in the etiology of OP toxicity. In study one, rats were given either saline or anti-TNF (50,000 units, i.p.). Three hours later, animals were given corn oil (CO) or 25 mg/kg CHP by oral gavage in the morning. In study two, rats were given anti-TNF followed by CO or 10 mg/kg CHP in the afternoon. Core temperature and motor activity were monitored continuously by telemetry. In study one, anti-TNF (50,000 units) had no effect on the hypothermic response to 25 mg/kg CHP. However, anti-TNF treated animals maintained higher fevers 3 days (48-96 h post-injection) after CHP treatment. In study two, anti-TNF attenuated the hypothermic response induced by 10 mg/kg CHP but had no effect on the magnitude of the delayed fever. Overall, 25 mg/kg CHP elicited a longer period of hypothermia and delayed fever compared to 10 mg/kg CHP. Anti-TNF pretreatment attenuated the hypothermic response at the lower CHP dose and exacerbated the fever at the higher CHP dose. Anti-TNF also attenuated the hypothermic effect of high doses of LPS and exacerbated LPS-induced fever. These data indicate that endogenously produced TNF is involved in the etiology of CHP mediated hypothermia and fever. PMID- 10514031 TI - Toxicity of low levels of methylglyoxal: depletion of blood glutathione and adverse effect on glucose tolerance in mice. AB - Methylglyoxal, a metabolic by-product of glycolysis is also formed during food processing and has serious toxicological effects when in excess. In this study, ddY mice were exposed to low levels of methylglyoxal (1% v/v) via drinking water while in utero continuing until 2 months of age when investigations on blood GSH status and selected GSH dependent functions in the blood, and glucose tolerance were carried out. The results showed that GSH content was significantly decreased in the blood of methylglyoxal exposed mice when compared with controls (mean, 0.756 mmol/l vs. 1.090 mmol/l, p < 0.001). The data showed significant (p < 0.001) decreases in blood GSH-S-transferase activity and red blood cell (rbc) capacity to refract oxidative stress. Impaired glucose tolerance was 5.3 times more prevalent in the methylglyoxal exposed mice when compared with the controls. The results indicate that chronic intake of methylglyoxal, at levels that could be attained in food, is toxic by depletion of blood GSH and could have adverse effect on some GSH dependent functions in vivo. PMID- 10514032 TI - Proliferation of hepatic peroxisomes in rats following the intake of green or black tea. AB - Rats maintained on green, black or decaffeinated black tea (2.5%, w/v) as their sole drinking fluid displayed higher hepatic CN- insensitive palmitoyl CoA oxidase activity than controls; the extent of increase was similar with the three types of tea. Morphological examination of the liver using electron microscopy revealed an increase in the number of peroxisomes in the tea-treated animals. The same treatment of the animals with green and black tea resulted in a similar rise in hepatic microsomal lauric acid hydroxylation. Analysis by HPLC of the aqueous tea extracts employed in the current study showed that the total flavanol content of the green variety was much higher than the black varieties, and confirmed the absence of caffeine in the decaffeinated black tea. It may be concluded from the present studies that neither caffeine nor flavanoids are likely to be responsible for the proliferation of peroxisomes observed in rats treated with tea. PMID- 10514033 TI - Infiltration of lymphocytes in the limbic brain following stimulation of subclinical cellular immunity and low dosages of lithium and a cholinergic agent. AB - This experiment was designed to investigate the hypothesis that single small dosages of lithium (1.5 mEq/kg), the muscarinic agent pilocarpine (15 mg/kg) and spinal cord emulsion encourage perivascular infiltration of lymphocytes into the brain even when overt symptoms of experimental allergic encephalomyelitis are not apparent. The brains of rats that had received this small dosage of lithium and pilocarpine exhibited discernable infiltrations of lymphocytes within limbic tracts but no discernable neuronal loss. Although the brains of the rats that displayed overt seizures following larger dosages of lithium (3 mEq/kg) and pilocarpine (30 mg/kg) exhibited the usual pattern of neuronal loss within multiple thalamic and limbic structures and conspicuous foci of lymphocytic infiltration (particularly within the hippocampal formation) the correlation between the numbers of foci and the proportions of neuronal damage in these structures was not significant statistically. These results indicate that infiltrations of lymphocytes into brain parenchyma are not simple artifacts of the amount of neuronal damage and may be sensitive toxicological markers for subclinical interactions between drugs and immune responses. PMID- 10514034 TI - The effect of curcumin on glutathione-linked enzymes in K562 human leukemia cells. AB - Curcumin, an antioxidant present in the spice turmeric (Curcuma longa), has been shown to inhibit chemical carcinogenesis in animal models and has been shown to be an anti-inflammatory agent. While mechanisms of its biological activities are not understood, previous studies have shown that it modulates glutathione (GSH) linked detoxification mechanisms in rats. In the present studies, we have examined the effects of curcumin on GSH-linked enzymes in K562 human leukemia cells. One micromolar curcumin in medium (16 h) did not cause any noticeable change in glutathione peroxidase (GPx), glutathione reductase, and glucose-6 phosphate dehydrogenase activities. Gamma-glutamyl-cysteinyl synthetase activity was induced 1.6-fold accompanied by a 1.2-fold increase in GSH levels. GSH S transferase (GST) activities towards 1-chloro-2,4-dinitrobenzene, and 4 hydroxynonenal (4HNE) were increased in curcumin-treated cells 1.3- and 1.6-fold, respectively (P = 0.05). The GST isozyme composition of K562 cells was determined as follows: 66% of GST Pl-1, 31% of Mu class GST(s), and 3% of an anionic Alpha class isozyme hGST 5.8, which was immunologically similar to mouse GSTA4-4 and displayed substrate preference for 4HNE. The isozyme hGST 5.8 appeared to be preferentially induced by curcumin, as indicated by a relatively greater increase in activity toward 4HNE. Immunoprecipitation showed that GPx activity expressed by GST 5.8 contributed significantly (approximately 50%) to the total cytosolic GPx activity of K562 cells to lipid hydroperoxides. Taken together, these results suggest that GSTs play a major role in detoxification of lipid peroxidation products in K562 cells, and that these enzymes are modulated by curcumin. PMID- 10514035 TI - Effect of a single or repeated dose of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on T cell subpopulations in the Long-Evans rat. AB - 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) exposure results in adverse effects on the immune system of experimental animals. The purpose of this study was to compare the effects of a single and repeated dosing of TCDD on splenic T-cell subpopulations in Long Evans rats 9 days post-exposure to TCDD. A single dose (25 microg/kg body weight) of TCDD resulted in reduced body weight. The percentage and total number of CD4+ or CD8+ subsets and percentage of CD4+ or CD8+ cell cycling in the S and G2M phases were similar in the single dosed (25 microg/kg body weight) TCDD group compared with the vehicle control. A repeated dose (5 microg/kg/day for 5 days) of TCDD also resulted in a significant reduction in body weight. However, multiple doses of TCDD significantly decreased the percentage of the CD4+ subset and the percentage of CD4+ cells cycling in the S and G2M phases. No significant change occurred in the CD8+ cell subpopulation after single or multiple dosing with TCDD. These results demonstrated that repeated dosing of TCDD decreased the total percentage of CD4+ cells and the percentage of CD4+ cells cycling 9 days post-exposure, while an analogous single dose of TCDD failed to affect the CD4+ cell subpopulation. The difference in biological responses to a single versus 'equivalent' multiple (cumulative) dose of TCDD is discussed. PMID- 10514036 TI - Rat liver in vivo replicative DNA synthesis test for short-term prediction of nongenotoxic (Ames-negative) hepatocarcinogenicity: a collaborative study of the Nongenotoxic Carcinogen Study Group of Japan. AB - A collaborative study was conducted to evaluate whether a replicative DNA synthesis (RDS) test using the rat liver can detect nongenotoxic (Ames-negative) hepatocarcinogens with three or seven daily administrations at dose-levels effective in long-term bioassays. The assay methods were well-validated by the 14 participants. Of six compounds tested, carbon tetrachloride (50 and 100 mg/kg), clofibrate (125 and 250 mg/kg), diethylstilbestrol (0.125 and 0.25 mg/kg) and urethane (100 mg/kg) gave positive results, methyl carbamate (200 and 400 mg/kg) exerted equivocal effects, and D,L-ethionine (125 mg/kg) failed to elevate RDS. These findings suggest that the RDS test can detect many nongenotoxic rat hepatocarcinogens with short-term administration at dose-levels used in long-term bioassays. PMID- 10514037 TI - Hepatic Ah receptor binding affinity for 2,3,7,8-tetrachlorodibenzo-p-dioxin: similarity between beagle dog and cynomolgus monkey. AB - Hepatic AhR binding affinity for [3H]-2,3,7,8-tetrachlorodibenzo-p-dioxin ([3H]TCDD) was compared between two species widely used as laboratory animals: beagle dog and cynomolgus monkey (Macaca fascicularis). The enriched 9S fractions from both species were obtained by sucrose gradient sedimentation. After incubation with [3H]TCDD, dextran-coat charcoal treatment (10 mg/ml) revealed that dog and monkey possess an AhR with a low binding affinity for [3H]TCDD. Saturation experiments were then achieved according to the method developed in experiments on human samples. The binding characteristics were determined after analysis of the data by Scatchard and Woolf plots. Receptor concentrations were quite similar in dog and monkey liver (26.6 and 14.4 pmol/mg, respectively) as well as the affinity (Kd) for [3H]TCDD (17.1 and 16.5 nM, respectively). The low binding affinity of dog and monkey AhRs appeared to be similar to those observed in human. PMID- 10514038 TI - Flow in carotid bifurcations: effect of the superior thyroid artery. AB - The superior thyroid artery was ignored in previous fluid dynamics studies of carotid bifurcations. However, it is not clear to what extent the flow patterns within the carotid might be influenced by the presence of this outflow tract in reality. In this study, quantitative effects of the superior thyroid artery upon the flow patterns and wall shear stress in the carotid bifurcation were investigated in detail by a numerical simulation method. Comparisons of the maximum reversed flow velocities, flow patterns and wall shear stress were made between models with and without the superior thyroid artery. Results demonstrate that this small artery has only a marginal effect on the overall flow characteristics within the carotid sinus. However, it does have significant effects on flow patterns in the common-external side branch. An alternative approach is proposed to compensate for the absence of this small artery in numerical calculations. PMID- 10514039 TI - Measurement of airborne ultrasonic slow waves in calcaneal cancellous bone. AB - Measurements of an airborne ultrasonic wave were made in defatted cancellous bone from the human calcaneus using standard ultrasonic equipment. The wave propagating under these conditions was consistent with a decoupled Biot slow wave travelling in the air alone, as previously reported in gas-saturated foams. Reproducible measurements of phase velocity and attenuation coefficient were possible, and an estimate of the tortuosity of the trabecular framework was derived from the high frequency limit of the phase velocity. Thus the method offers a new approach to the acoustic characterisation of bone in vitro which, in contrast to existing techniques, has the potential to yield information directly characterising the trabecular structure. PMID- 10514040 TI - Real time monitoring of muscular fatigue from dynamic surface myoelectric signals using a complex covariance approach. AB - A method aimed at the real-time monitoring of muscular fatigue was implemented and optimized. The method is based on an estimate of the complex covariance function in order to evaluate, in real time, the mean frequency of the myoelectric signal spectrum. Real-time implementation is guaranteed by a recursive computation of the complex covariance and then of the mean frequency. The results show good performance on both synthetic and experimental non stationary myoelectric signals recorded during fatiguing dynamic protocols. Performance in the presence of noise is highly satisfactory on both deterministic signals and stochastic processes, even when there are strong non-stationarities. Moreover, the computational complexity is highly reduced with respect to that offered by traditional methods based on short time Fourier transform. PMID- 10514041 TI - Inter- and intra-individual variability of ground reaction forces during sit-to stand with principal component analysis. AB - Variable reduction is an important issue in biomechanics, because the definition of a non-redundant set of variables necessary for a complete description of a given motor act provides information about the motor strategy. A systematic tool for dealing with variable reduction problems is Principal Component Analysis. In this paper, as an example of an application of this technique, the set of Ground Reaction Forces (GRFs) provided by a six-component force plate, gained during standing up in a heterogeneous population of 82 normal individuals, was reduced to a set of fewer variables. Each subject was required to stand up from a chair five times at different, randomly self selected, speeds, obtaining a data set of 410 trials. Principal Components (PCs) of GRFs were computed for each trial. On average, over the ensemble of trials, first and second PCs (PC1 and PC2) explained together 90% of PCs. Inter- and intra-individual repeatability of the first two PCs was investigated by examining the correlation coefficient between PC waveforms obtained from the whole set of trials and within the set of trials performed by the same subject, respectively. While the PC1 exhibited repeatable patterns, the second one, although repeatable within the group of trials performed by the same subject, displayed marked inter-individual variability. Therefore, PC1 was related to intrinsic aspects of the motor task and PC2 to inter-subject features. PMID- 10514042 TI - Fluorescent strip-lights as a source of error in tissue reflectance spectroscopy. AB - Tissue reflectance spectroscopy is a widely used technique for measuring the blood content of the skin. Fluorescent strip-lighting of a type in widespread use (Osram L36 W/23 Gelbweiss-white) is a potential source of misleading artefacts in tissue reflectance spectroscopy. The fluorescent light signal has successive peaks and troughs in the 520-580 nm range which closely emulate the inverse of the reflected signal of white light modulated by blood. The presence of fluorescent light in the reflected signal cancels some of the blood signature. Conversely, the presence of fluorescent light in the white reference augments the true blood reflected signal. The marked similarity of the resultant artefact blood spectrum to the true blood spectrum is a hazard, as the presence of the artefact may go undetected. PMID- 10514043 TI - Normalisation of EMG amplitude: an evaluation and comparison of old and new methods. AB - The purpose of this study was to evaluate and compare four different methods of normalising the amplitude of electromyograms (EMGs), from the biceps brachii. Five males performed isotonic contractions of the elbow flexors with an external force of 50 N, 100 N, 150 N and 200 N. These were followed by a single isometric maximal voluntary contraction (MVC) and ten isokinetic MVCs at 0.35 rad s(-1) intervals between 0.35 rad s(-1) and 3.50 rad s(-1). The processed EMGs recorded from the isotonic contractions were normalised by expressing them as a percentage of: (i) the mean (Dynamic Mean Method) and (ii) the peak EMG from the same contraction (Dynamic Peak Method), (iii) the EMG from the isometric MVC (Isometric MVC Method), and (iv) the EMG from an isokinetic MVC at the same elbow angle and angular velocity (Isokinetic MVC Method). The root mean square difference (RMSD) between the outputs of the Isokinetic MVC and Dynamic Mean methods was significantly greater (P<0.05) than between the Isokinetic MVC method and the Dynamic Peak and the Isometric MVC methods. The small (10%) difference between the Isokinetic MVC and the Isometric MVC Methods was a consequence, firstly, of the lack of difference in EMG recorded from the isometric and isokinetic MVCs and, secondly, the consistency in EMG over the range of motion and at different angular velocities of isokinetic MVC. We conclude that only the Isometric and Isokinetic MVC methods should be used to normalise the amplitude of EMGs from the biceps brachii. PMID- 10514044 TI - Measurement of posture by triangulation using potentiometers. AB - To measure the posture of a paraplegic while standing up, an economical method has been developed. Markers, glued over the joints, hook onto strings which run to rotary potentiometers mounted on a fixed frame. Springs maintain a near constant small tension in the strings, and the potentiometers rotate as the strings wind onto pulleys. The positions are calculated by triangulation, using two potentiometers per joint, assuming that body segment lengths are constant, or three potentiometers without this assumption. Using a second-order polynomial fit, the random error in length measurement for each potentiometer is less than +/-2 mm for the string length from 0 mm to 1100 mm, or less than +/-1 mm in actual range from 600 mm to 1000 mm. With two potentiometers per joint, using a second-order polynomial fit and assuming the ankle position is known exactly, an estimate of the resulting errors in the knee and hip marker positions are 4 and 8.5 mm, respectively. PMID- 10514045 TI - A new system for the measurement of displacements of the human body with widespread applications in human movement studies. AB - This paper reports the development, construction and use of a new system for the measurement of linear kinematics in one, two or three dimensions. The system uses a series of rotary shaft encoders and inelastic tensioned strings to measure the linear displacement of key anatomical points in space. The system is simple, inexpensive, portable, accurate and flexible. It is therefore suitable for inclusion in a variety of motion analysis studies. Details of the construction, calibration and interfacing of the device to an IBM PC computer are given as is a full mathematical description of the appropriate measurement theory for one, two and three dimensions. Examples of the results obtained from the device during gait, running, rising to stand, sitting down and pointing with the upper limb are given. Finally it is proposed that, provided the constraints of the system are considered, this method has the potential to measure a variety of functional human movements simply and inexpensively and may therefore be a valuable addition to the methods available to the motion scientist. PMID- 10514046 TI - Estimation of survival rates in haematophagous insects. AB - Estimation of the survival rate through a gonotrophic cycle is an important factor in determining the vectorial capacity of a population of haematophagous insects in a disease cycle. Most methods used to calculate survival rates make stringent assumptions which may not be valid for all species. Birley and colleagues used a time series analysis of samples collected over several consecutive days, the lagged parous rate. Here, we use a simulation model to investigate (i) the length of data series needed and (ii) the consequences of failures in the assumptions of this method for the estimated survival rate. The accuracy of the estimated survival rate per cycle was high with sample periods of 10-100 days. The standard deviation (a measure of precision) decreased with the length of the sample period. When random sampling efficiency was included, the accuracy remained high but the estimates were less precise (larger standard deviations). If the sampling was biased in favour of either nulliparous or parous females, estimates of the survival rate were not accurate. The relationship between estimated survival rate, bias in collection, and true survival rate was non-linear. Thus, correction for the bias requires (i) prior knowledge of the direction and the severity of the bias and (ii) an independent estimate of the survival rate. This method of estimating survival rates is less accurate when the collection method is biased for or against parous females, although robust to other assumptions. PMID- 10514047 TI - Laboratory testing of a lethal ovitrap for Aedes aegypti. AB - Laboratory tests were conducted to determine the feasibility of making the mosquito ovitrap lethal to Aedes aegypti (L.) when they attempt to oviposit in the trap. Heavy-weight velour paper strips (2.54 x 11 cm) were used as an alternative to the wooden paddle normally provided as a substrate for mosquito oviposition. The paper strips were pretreated with insecticide solutions and allowed to dry before being used in oviposition cups of 473 ml capacity, filled with water initially to within 2.5 cm of the brim. Insecticides chosen for their quick knock-down efficacy were bendiocarb 76% WP (1.06 mg a.i./strip) and four pyrethroids: permethrin 25% WP (0.16 mg a.i./strip), deltamethin 4.75% SC (0.87 mg a.i./strip), cypermethrin 40% WP (2.81 mg a.i./strip), and cyfluthrin 20% WP (0.57 mg a.i./ strip). For experimental evaluation, two oviposition cups (one with an insecticide-treated strip and one with an untreated strip) were placed in cages (cubic 30 cm) with gravid female Ae. aegypti mosquitoes (aged 6-8 days) from a susceptible laboratory strain. Mortality-rates of female mosquitoes were 45% for bendiocarb, 47% for permethrin, 98% for deltamethrin, 100% for cypermethrin, and 100% for cyfluthrin. Young instar larvae added to the treated cups died within 2h. After water evaporation from the cups for 38 days, fresh mosquito females had access to previously submerged portions of the velour paper paddle, and mortality rates of 59% or more occurred. Cups that had water (360 ml) dripped into them, to simulate rain, produced female mosquito mortality rates of > 50% and all larvae died within 3 h of being added. These tests demonstrate that the ovitrap can be made lethal to both adults and larvae by insecticidal treatment of the ovistrip. Field efficacy trials are underway in Brazil to access the impact of this simple, low-cost, environmentally benign approach on populations of the dengue vector Ae. aegypti. PMID- 10514048 TI - Elevated oxidase and esterase levels associated with permethrin tolerance in Anopheles gambiae from Kenyan villages using permethrin-impregnated nets. AB - The permethrin tolerance (PT) of a population of the mosquito Anopheles gambiae (Diptera: Culicidae) increased following the introduction of permethrin impregnated nets for malaria control in certain villages near Kisumu, western Kenya. Using a biochemical test that indirectly measures oxidases associated with permethrin resistance, we found that this population had higher oxidase levels than a comparison population from villages without impregnated nets. Mosquitoes from a colony of An. gambiae selected for PT, the RSP (reduced susceptibility to permethrin) strain, were exposed to permethrin with or without the oxidase inhibitor piperonyl butoxide (PB). Significantly higher mortality rates occurred when permethrin was synergized by PB, presumably by suppression of oxidases responsible for PT. An unselected (UNS) colony of An. gambiae that was more susceptible than RSP in a permethrin-susceptibility bioassay (i.e. LT50 22 min for UNS, vs. 42min for RSP) was compared with the RSP colony for levels of oxidases and esterases. The levels of both enzymes were very significantly higher in the RSP strain (P<0.0001). We speculate that use of impregnated nets selected for higher oxidase and esterase levels in An. gambiae to metabolize permethrin acquired from the nets. Both oxidase and esterase mechanisms could confer cross resistance to other pyrethroids. PMID- 10514049 TI - Seasonal and spatial changes in blowfly production from small and large carcasses at Durham in lowland northeast England. AB - Colonization by blowflies (Diptera: Calliphoridae) of mouse carcasses exposed in open agricultural land near Durham (54 45'N) changed from early spring monopolization by Calliphora vicina R.-D. to a summer pattern of multiple species exploitation by this species together with Lucilia caesar L., L. illustris Mg., L. silvarum Mg., L. sericata Mg. and L. richardsi Collin. In a garden at the edge of Durham, mouse carcasses were dominated by C. vicina from spring to autumn. Difference in mouse colonization between the agricultural and garden sites seemed to reflect differences in the blowfly species present, as measured by baited trap catches at the sites. In sets of C. vicina reared from mice under conditions of competition for larval food, it was found that resulting females were significantly larger than males, size being measured as mean wing length. Blowfly production from three sheep carcasses exposed successively at the agricultural site was dominated by C. vomitoria L. and L. caesar, but also produced other Lucilia species in small numbers, including L. sericata. These L. sericata females from sheep that had died from causes other than myiasis included full sized specimens, in contrast to those produced from mouse carcasses that were all undersized individuals. As L. sericata females trapped on sheep pastures are predominantly full-sized, this suggests that large carcasses may, in part, be a source of the L. sericata population that attacks sheep as a myiasis agent. The nature of large carcasses as possible sources of L. sericata in lowland Britain is discussed. PMID- 10514050 TI - Abundance of house dust mites in relation to climate in contrasting agricultural settlements in Israel. AB - The correlation between climatic conditions and mite numbers in houses from rural areas was studied in 13 agricultural communities (kibbutzim and moshavim) in nine geo-climatic subregions of Israel. Mites were present in 97% of the dust samples. The average number of mites per gram of dust in the different localities ranged between 84 and 2053. The maximum number of mites (7440/g dust) was found in a carpet from a house in Geva Carmel in the northern coastal region. The most prevalent species of mites were Dermatophagoides pteronyssinus and Dermatophagoides farinae, which were found in 85.6% and 71.3% of the samples, respectively. The house dust mites D. pteronyssinus, D. farinae and Euroglyphus maynei constituted 94.8% of the mites. Most of the mites were isolated from the carpets and sofas (37.0% and 33.7%, respectively), and a smaller number from beds (29.3%). The smallest number of mites (< or = 250/g dust) were found at a minimum relative humidity (RH) of 30% and lower, with a maximum temperature of 32 degrees C and higher, i.e. in the Jordan valley and Negev mountains. A greater number of mites (250-500/g dust) were found at a minimum ambient RH of 35-40% and a maximum temperature of 32 degrees C and higher, i.e. the Hula valley. A large number of mites (500-1000/g dust) were found at a minimum RH of 35-40% with a maximum temperature of 30 degrees C and lower, i.e. in the Judean and Samarian range, as well as in upper Galilee. The largest number of mites (1000-2000/g dust) was found at a minimum RH of 45% and higher, with a maximum temperature ranging between 30 and 32 degrees C. These conditions occur in the coastal strip, the coastal plain and in the Judean and Samarian foothills. A monthly examination of two houses in Zova, a kibbutz in the Judean hills next to Jerusalem, and two houses from Palmachim, a kibbutz in the coastal region, revealed that the highest prevalence of mites was found in the months April-November and May-November, respectively. In Zova, the highest number of mites were found during the months of June and July while the highest concentrations of D. pteronyssinus-antigen (Der p I) were measured during the month of September. A positive correlation between mite numbers and the quantity of Der p I in house dust was found. PMID- 10514051 TI - Distribution of domestic Psocoptera in Madrid apartments. AB - Domestic Psocoptera were sampled in Madrid throughout a complete year (1991-92) by means of sticky traps in bathrooms, kitchens and inside windows. A total of 409 individuals comprising ten species of psocids was caught from 4056 traps inspected fortnightly. The predominant species were Liposcelis decolor, Li. brunnea, Psyllipsocus ramburi, Li. bostrychophila, Li. pearmani, Dorypteryx domestica and Lachesilla pedicularia, in descending order of abundance. Apartments were classified according to eight quantitative variables: age and area of building, level of apartment (i.e. storey), numbers of windows, inhabitants, their pets (birds and mammals), ornamental plants indoors; and two qualitative factors: floor material and presence or absence of trees adjacent to the house. Psocid communities in human habitations were assessed by the island biogeography theory: diversity increased with apartment size and the number of species decreased with building date. Psocid populations were most abundant in bathrooms, which presumably act as sources for dispersal to other parts of the premises. No other significant correlations were found with the other housing factors investigated. Whereas psocids were active indoors during all months of the year, the peak prevalence was in summer, July and August, one or two months ahead of the local seasonal pattern for non-domestic psocids. Although psocids are usually not seriously unhygienic, causing only minor problems of contamination and allergies, this information should help towards their environmental control. PMID- 10514052 TI - PCR-based methods for identification of species of the Anopheles minimus group: allele-specific amplification and single-strand conformation polymorphism. AB - We report two polymerase chain reaction (PCR)-based methods for distinguishing morphologically similar species based on amplification of a variable region of the 28S gene of ribosomal DNA. The four species we investigated are mosquitoes of the Anopheles minimus group: An. aconitus, An. varuna and An. minimus species A and C. The formally named species are vectors of human malaria parasites in south east Asia but are difficult to distinguish with certainty on the basis of morphology. Allele-specific amplification was used to differentiate An. minimus A from An. minimus C. This technique has been widely used for the diagnosis of species. Single-strand conformation polymorphisms (SSCPs) were used to separate all four species. This technique, which has seldom been used for species identification, has many advantages: it does not require sequence information beyond that needed for amplification; it is ideally suited for the detection of heterozygotes; it utilizes more of the information in the PCR product than allele specific amplification; it distinguishes all four species considered here and could easily be extended to other species; previously unknown intraspecific variation and additional species are likely to be detected. Thus, SSCPs provide valuable population genetic information which allele-specific amplification does not. PMID- 10514053 TI - Molecular and kinetic evidence for allelic variants of esterase Estbeta1 in the mosquito Culex quinquefasciatus. AB - Elevated esterase Estbeta1 was purified from larvae of newly isolated strains of the mosquito Culex quinquefasciatus from Colombia (COL) and Trinidad (TRI) with resistance to organophosphate (OP) insecticides. Insecticide interactions were compared with those of elevated Estbeta1(2) from the OP-resistant Habana strain and the non-elevated Estbeta1(3) from the susceptible PelSS strain. On the basis of insecticide binding efficiency, all elevated Estbeta1 esterases were readily distinguishable. Differences between the EcoRI restriction fragment patterns of the amplified estbeta1 gene in COL and TRI strains compared with each other, and between amplified estbeta1(1), estbeta1(2) and the non-amplified estbeta1(3), suggest differences in their nucleotide sequence. Considering their variable insecticide binding efficiencies, these genetic differences would imply that, in contrast to estalpha2 and estbeta2, amplification of estbeta1 has occurred several times independently. Generally, the elevated Estbeta1s were more reactive with insecticides than the non-elevated Estbeta1(3). This supports the hypothesis that the elevated esterase-based mechanism confers resistance through amplification of alleles coding for esterases which have a greater specificity for the insecticides they sequester than the esterases coded by their non amplified counterparts. PMID- 10514054 TI - Identification of bloodmeals in haematophagous Diptera by cytochrome B heteroduplex analysis. AB - We developed a DNA assay for bloodmeal identification in haematophagous insects. Specific host cytochrome B gene sequences were amplified by PCR and classified on the basis of their mobility in a heteroduplex assay. In the blackfly Simulium damnosum s.l. (Diptera: Simuliidae), human cytochrome B DNA sequences were identifiable up to 3 days following ingestion of the bloodmeal. In the tsetse Glossina palpalis (Diptera: Glossinidae) collected from tsetse traps in Ivory Coast, bloodmeals were identified as taken from domestic pigs on the basis of their heteroduplex pattern and DNA sequence. Evidently the cytochrome B sequence shows sufficient interspecific variation to distinguish between mammalian host samples, while exhibiting minimal intraspecific variation. The stability of DNA in bloodmeals, for several days post-ingestion by haematophagous insects, allows PCR-HDA assays to be used reliably for host identification. PMID- 10514055 TI - Scanning electron microscopy of the second-instar larva of Gasterophilus nasalis. AB - The second-instar larva of the bot fly Gasterophilus nasalis (L.) (Diptera: Gasterophilidae) is described for the first time, based on scanning electron microscope (SEM) studies. On the pseudocephalum the larva bears an antenomaxillary sensory complex formed by the antenna (coeloconic sensilla) and the maxillary palp with a set of six coeloconic sensilla and four basiconica sensilla. The oral opening is latero-posteriorly limited by small spines, and exhibits strongly ornamented maxillae and mandibles. The thoracic and abdominal segments are circled by two bands, each with two rows (except the last segment that has one row) of backwardly pointed spines, and have cuticular depressions. Trichoid and campaniform sensilla surround the larval segments. The anterior spiracular opening is a small aperture. The terminal end of the eighth abdominal segment shows a spiracular cavity, lateral tubercles, eight basiconic and two trichoid sensilla. Each spiracular plate has two slightly curved slits, each with a serrated rima. There is a probable ecdysial scar. The findings of this ultrastructural study are compared with those other of larval flies. PMID- 10514056 TI - Colonization and bionomics of the sandfly Phlebotomus kazeruni from Sinai, Egypt. AB - Phlebotomuius kazeruni (Diptera: Psychodidae) females were collected by light trap in southern Sinai, Egypt, and this sandfly species was colonized for the first time as a laboratory strain, maintained by the procedures of Modi & Tesh (1983). Laboratory-reared females did not lay eggs autogenously; they blood-fed more readily (P=0.02) on a hamster (37%) than a human (22%) during 1 h exposure. Fecundity of hamster-fed females was significantly greater than for those fed on human blood: 69.4 +/- 5.8 vs. 45.2 +/- 8.1 eggs/female from the first gonotrophic cycle. Pre-oviposition and egg incubation periods were significantly less for females fed on hamster compared with human blood, but the larval development and pupation periods were not affected by this difference in bloodmeal source. Egg to adult survival was equivalent (38%) for progeny of females blood-fed on hamster or human. The mean generation time of progeny from females fed on hamster (51.9 +/- 1.0 days) or human (53.3 +/- 1.7 days) was not significantly different. The sex ratio of adult male:female progeny was similar (P=0.2) for both hosts: 42:58% from hamster, 46:54% from human blood-fed female parents. Evidently P. kazeruni from Sinai is sufficiently anthropophagic to be a potential vector of Leishmania from rodents to humans. PMID- 10514057 TI - Experimental comparison of anthropophily between geographically dispersed populations of Lutzomyia whitmani (Diptera: Psychodidae). AB - Lutzomyia whitmani, a major vector of cutaneous leishmaniasis in Brazil, occupies diverse habitats from the Amazon forest canopy to suburban animal pens. Three mitochondrial lineages of Lu. whitmani ('Amazonian', 'North-South' and 'North east') have parapatric distributions coinciding with different ecological zones. We assessed the host preferences of populations representing the three lineages in standardized field experiments, and found that Lu. whitmani in all sites were significantly more attracted to humans than to dogs or chickens. Females from a southerly population of the North-South lineage showed the greatest degree of anthropophily. Lu. whitmani from Amazonia were also strongly attracted to human baits, contradicting previously published accounts. Intraspecific comparisons in non-Amazonian sites suggest that Lu. whitmani is less anthropophilic than Lu. intermedia but more so than Lu. longipalpis. No significant difference was detected in anthropophily between Lu. whitmani in the Amazon and either Lu. dendrophyla or Lu. gomezi. Anthropophilic behaviour was demonstrated in the same site for Lu. complexa, Lu. flaviscutellata and Lu. brachyphalla, but not for Lu. infraspinosa. PMID- 10514058 TI - Protective efficacy of lambdacyhalothrin-impregnated bednets against Phlebotomus orientalis, the vector of visceral leishmaniasis in Sudan. AB - Field investigations on the sandfly Phlebotomus orientalis (Diptera: Psychodidae), the vector of Leishmania donovani causing visceral leishmaniasis (VL) in Sudan, were undertaken in two villages (Bellow and Elgamel) and Dinder National Park, to determine the protective value of bednets (polyester, 100 denier) impregnated with lambda-cyhalothrin 10 mg a.i./m2 pyrethroid insecticide. After exposure to treated netting for 30 s, P. orientalis females all died within 1 h. When field-tested in Acacia woodland, treated bednets provided complete protection from bites of the vector. Numbers of P. orientalis females landing on human collectors without bednets or using untreated bednets averaged 32.0 +/- 8.3 or 6.9 +/- 2.7 per man-night, respectively, whereas collectors using treated bednets experienced no sandfly bites during the same period (18.00-06.00 hours, 12 nights in June 1995). Socio-behavioural observations on the bed-time of people living in both study villages indicated that the use of impregnated bednets against P. orientalis would give more potential protection for women and children than for male adults. Overall the proportions of people and their durations of exposure to the risk of sandfly bites (i.e. after sunset until they went to bed) were 40% unprotected for< 1h, 50% for 1-2h and >10% for > or = 2h. Because visceral leishmaniasis in Sudan occurs mainly in children, the use of impregnated bednets (outdoors as well as indoors), and going to bed early could provide a high degree of personal protection against this zoonotic infection. PMID- 10514059 TI - The efficacy of various pyrethroid insecticides for use on odour-baited targets to control tsetse. AB - The efficacy of various pyrethroid insecticides for use on odour-baited targets to control tsetse was compared in Zimbabwe. Formulations were applied to cotton cloth and polyester net and, at various intervals, the materials were bioassayed by exposing fed female Glossina pallidipes (Austen) (Diptera: Glossinidae) to cloth for 45 s or by inducing them to collide briefly with net. Trial formulations were compared with deltamethrin suspension concentrate (s.c.), the insecticide currently used in tsetse control operations in Zimbabwe. Applying 0.8% suspension of alphacypermethrin to cloth or net produced high mortalities for 9 months which was similar in performance to 0.4% suspension of deltamethrin s.c. Deltamethrin s.c. and beta-cyfluthrin s.c. applied to cloth as 0.1% suspensions were equally effective, producing high mortalities for 2 months during the wet season, and 0.8% suspension of beta-cyfluthrin was effective for 12 months. Suspensions of 0.1% lambdacyhalothrin capsule suspension or 0.1% lambdacyhalothrin wettable powder were significantly less effective than 0.1% deltamethrin s.c. Chemical analyses showed that increasing the concentration of insecticide applied to material increased the initial amount of insecticide on the material and decreased the subsequent rate of loss; 0.1% suspension of beta cyfluthrin s.c. applied to cloth produced an initial concentration of approximately 280 mg/m2 which declined by 94% in 12 months whereas 0.8% suspension showed no significant decrease in concentration (mean= 1304 mg/m2) over the same period. For controlling tsetse by means of pyrethroid-treated targets, it is suggested that beta-cyfluthrin s.c. is as effective as deltamethrin s.c. but that alphacypermethrin s.c. should be used at twice the concentration of deltamethrin s.c. to obtain the same performance. PMID- 10514060 TI - Skin lesions and cattle hide damage from Haematobia irritans infestations. AB - The horn fly Haematobia irritans L. (Diptera: Muscidae) has recently spread to Argentina and Uruguay and is believed to cause damage to cattle hides. Four groups of ten Holstein steers each were maintained for 58 weeks under different infestation levels with H. irritans to determine if it was the cause of this problem. Hides (chrome tanned) from steers maintained under minimum infestation level had 4.7 +/- 3.8% of the area damaged. Maintaining the steers under low H. irritans level for the last 44 days of the trial using insecticidal ear-tags, resulted in 29.5 +/- 15.8% of hide area being damaged. Steers that were treated with 5% cypermethrin pour-on, when the H. irritans population was close to 50 flies, showed that 31.3 +/- 16.6% of hide area was injured, and 46.6 +/- 12.8% of damaged hide area was found in hides from non-treated steers. Significant differences were found between mean hide damage from steers maintained continuously under low H. irritans infestation levels and all other groups. Hyperaemia was significantly lower in the skin of steers under low H. irritans infestation level than in the skins of non-treated steers and steers maintained under low-level infestations for the final 44 days. Eosinophil and mononuclear cell infiltration was significantly lower when the population of H. irritans was less than six per steer than when the population was more than 100 flies per steer. Low numbers of Stomoxys calcitrans were found in all groups, but most hide damage was presumed due to H. irritans. PMID- 10514061 TI - Host influence on adaptation of Trypanosoma congolense metacyclics to vertebrate hosts. PMID- 10514062 TI - Heterogeneity in the risk of sleeping sickness in coffee and cocoa commercial plantations in Ivory Coast. PMID- 10514063 TI - Self-curing membranes of chitosan/PAA IPNs obtained by radical polymerization: preparation, characterization and interpolymer complexation. AB - Chitosan/polyacrylic acid I PN's were prepared by radical polymerization of acrylic acid, AA, activated at low temperature, in an aqueous/alcoholic chitosan dispersion. AA monomer to polymer conversion and membrane compositions were determined by elemental analysis and FTIR. Evidences of interpolyelectrolite complex formation were found from the FTIR spectra as well. The gravimetric measurements and the elemental analysis after some exhaustive PAA extraction support the existence of some PAA grafting on the reactive amine group of the chitosan. Swelling degree of the membranes is highly dependent on pH and composition, showing a higher swelling in membranes richer in AA and increased pH due to the breaking of interpolyelectrolite salt bonds. PMID- 10514064 TI - Photoimmobilisation of poly(N-vinylpyrrolidinone) as a means to improve haemocompatibility of polyurethane biomaterials. AB - A novel method to improve the haemocompatibility of polymeric biomaterials (in particular: polyurethane elastomers) is reported. The new approach essentially rests upon photochemical immobilisation of the highly biocompatible polymer poly(N-vinylpyrrolidinone) (poly(NVP)) onto the biomaterial's surface. One of the key steps in the surface modification procedure is the preparation of a copolymer of NVP and the photoreactive building block 4-[4'-azidobenzoyl]-oxo-n butylmethacrylate (1). This copolymer is first dissolved in a volatile solvent, then sprayed onto the biomaterial's surface, and subsequently immobilised via irradiation with ultraviolet light. The paper describes: (i) preparation of 1, (ii) preparation of the copolymer (NVP + 1), (iii) physico-chemical characterisation of the modified surfaces, and (iv) results of two in vitro haemocompatibility assays (i.e. thrombin generation and adhesion of blood platelets from recalcified human platelet-rich plasma). Furthermore, the surface modification was performed with a microporous polyurethane vascular graft (Chronoflex), which is already in clinical use. The in vitro experiments revealed that significant improvement of the haemocompatibility of polyurethanes can be achieved through this method. PMID- 10514065 TI - F+ ion implantation induced cell attachment on intraocular lens. AB - Cell attachment on the polymethylmethacrylate (PMMA) intraocular lens (IOL) was studied by ion implantation. F+ ion implantation was performed at an energy of 80 keV with fluences ranging from 5 x 10(12) to 5 x 10(15) ions/cm2 at room temperature. The cell attachment tests gave interesting results in that the number of the platelets, the neutral granulocytes, and the macrophages adhering on the surface of the IOLs was reduced significantly after F+ ion implantation. The optimal fluence was about 3 x 10(14) to 4 x 10(14) ions/cm2. The hydrophobicity imparted to the surface was also monitored. At the same time, no appreciable change in the tensile strength and the optical transmittance of the implanted samples was observed. X-ray photoelectron spectroscopy (XPS) and fourier transfer infrared (FTIR) analysis showed that F+ ion implantation caused the cleavage of some pendants, the oxidation of the surface, and the formation of some new F-containing groups. These results were responsible for the cell attachment changes. PMID- 10514066 TI - Effect of filler content on the profile of released biodegradation products in micro-filled bis-GMA/TEGDMA dental composite resins. AB - This study assesses the effect of the filler content, in a micro-filled composite (0.04 microm), on the liberation of biodegradation products derived from two model composite systems. The materials were based on bis-phenyl glycidyl dimethacrylate (bis-GMA) and triethylenene glycol dimethacrylate (TEGDMA) monomers. The composites were produced using silica filler concentrations of 20 and 40%) by weight. Samples were incubated with either cholesterol esterase (CE) or phosphate buffer solutions (PBS) for 8, 16 and 32 days. Products were isolated by high-performance liquid chromatography (HPLC) and identified by mass spectrometry. The identified products included TEGDMA, 2,2-bis[4(2,3 hydroxypropoxy)-phenyl]propane (bis-HPPP) and triethylene glycol methacrylate (TEGMA). Bis-HPPP was only produced in the presence of enzyme. The amount of isolated TEGMA, in both composite systems, was shown to be significantly higher for materials incubated with enzyme than their buffer counterparts (P < 0.05). Between 0 and 8 days incubation with enzyme, significantly higher amounts of Bis HPPP and TEGMA were generated with the lower filler model material (composite-20) than the higher filled composite (composite-40), while the opposite effect was observed between 8 and 16 days. The data indicate that biodegradation product release profiles are dependent on the filler/resin ratios, and suggests that this parameter should be considered when assessing product release for biocompatibility issues pertaining to dental composite systems. PMID- 10514067 TI - Osteogenesis with coral is increased by BMP and BMC in a rat cranioplasty. AB - Autologous bone marrow cells (BMC), bone morphogenetic protein (BMP) and natural coral exoskeleton (CC) were used to enhance the repair of large skull bone defects in a craniotomy model. Nine millimeter calvarial defects were created in adult rats and were either left empty (control defects) or implanted with CC alone, CC-BMC, CC-BMP, or CC-BMC-BMP. After 1 or 2 months, osteogenesis was insufficient to allow union when defects were left empty or filled with CC. Addition of BMC alone to CC had no positive influence on osteogenesis at any time and increased CC resorption at 2 months (0.1 +/- 0.1 mm2 versus 0.5 +/- 0.3 mm2). In contrast addition of BM P or BM P/BMC to CC led to a significant increase in osteogenesis and allowed bone union after 1 month. At 2 months, the combination of CC-BM P-BMC was the most potent activator of osteogenesis. Filling a defect with CC-BMP-BMC resulted in significantly increased bone surface area (11 +/- 2.7 mm2) in comparison to filling a defect with CC-BMP (7.0 +/- 1.4 mm2), CC-BMC (3.5 +/- 1.1 mm2) or CC (4.5 +/- 0.4 mm2). CC resorption was significantly decreased in the presence of BMP with or without BMC at both times. These data are in accordance with the presence of progenitor cells in bone marrow that are inducible by BMP to the osteogenic pathway in a cranial site. The increase in material resorption in defects filled with CC-BMC could suggest that cells from the granulocyte-macrophage lineage survived the grafting procedure and were still active after 2 months. PMID- 10514068 TI - Bupivacaine-loaded comatrix formed by albumin microspheres included in a poly(lactide-co-glycolide) film: in vivo biocompatibility and drug release studies. AB - Bupivacaine-loaded comatrix, formed by bupivacaine-loaded microspheres included in a poly(lactide-co-glycolide) film, was assayed for the controlled release of the drug 'in vivo'. The comatrix, with 66.37 microg of bupivacaine, signifying a dose of 265.5 microg/kg, was subcutaneously implanted in the back of rats. Maximum plasma bupivacaine concentration was 147.6 +/- 5.0 ng/ml 95 h after the device implantation, and the drug was detected in plasma for 17 days. The half life time of bupivacaine improves by more than 50 times with regard to that of the drug administered in a solution by intraperitoneal injection. After 15 days of implantation the comatrix was included in a thin fibrous capsule and degradation of the microspheres was observed. The histological studies show good biocompatibility of this comatrix. After 50 days the comatrix was degraded and its remains were almost indistinguishable from the surrounding tissue. Small number of microspheres was observed and they were surrounded by conjunctive tissue. Nerve packets and small blood vessels were also observed in the periphery of the implant. PMID- 10514069 TI - Mixed-oxides prosthetic ceramic ball heads. Part 2: effect of the ZrO2 fraction on the wear of ceramic on ceramic joints. AB - Three different types of mixed-oxides ceramic ball heads have been investigated for their wear behaviour against acetabular cups of the same materials in a hip joint simulator. Mixed-oxides ceramics have been indicated in literature as a promising compromise between strength and wear but no reports are available on the influence of a percentage of zirconia in a ceramic femoral head when sliding against itself. Mixed-oxides ceramic acetabular cups and femoral heads were tested on a simulator apparatus with a sinusoidal load in presence of bovine calf serum. The experimental results did not show any significant difference between the experimental and commercial ceramic material couplings. These results were found to be in accord with those developed in Part 1. PMID- 10514070 TI - Microspheres of hydroxyapatite/reconstituted collagen as supports for osteoblast cell growth. AB - Microspheres comprised of particulate hydroxyapatites dispersed in fibrous collagen matrices were prepared. The procedure involved the droplet formation of hydroxyapatite/collagen mixture emulsified in olive oil, followed by the reconstitution of collagen in the presence of hydroxyapatite particles at 37 degrees C. Various sizes of microspheres could be obtained by controlling the stirring speed of the emulsified mixture. By increasing the stirring speed from 200 to 350 and 500 rpm, the average diameter of the microspheres decreased from 1038 to 513 and 226 microm, respectively. The sizes of the microspheres reduced substantially to a range of 141 microm when 2%. Span 85 was present in the emulsion mixture stirring at 200 rpm. The microspheres thus obtained can be used as carriers to support the growth of osteoblast cells. Osteoblast cells derived from calvaria proliferated from 1.5 x 10(5) to 4.5 x 10(5) cells/ml in 7 days. Correspondingly, the alkaline phosphatase activity increased 6 fold during this period. These results suggested that the hydroxyapatite/collagen microspheres could be used as the filling materials for bone defect. PMID- 10514071 TI - Effects of chitosan on rat knee cartilages. AB - The aim of this work is to study the effects of chitosan on rat knee cartilages. 0.2 ml of 0.1% chitosan solution pH = 6.9 were injected inside rat knees articular cavity. One, three and six weeks after injection, histological and histomorphometric studies were performed on undecalcified samples embedded in polymethylmetacrylate. Results show that after 1 and 6 weeks: (i) chitosan slows significantly (P < 0.005) the decrease in epiphyseal cartilage thicknesses and (ii) increases significantly articular cartilage chondrocyte densities (P < 0.002). However chitosan solution induces a proliferation of fibrous tissue with abundant fibroblasts, fibrocytes and monocytes inside the joint and this proliferation is still present after 6 weeks. This study suggests that chitosan could act on the growth of epiphyseal cartilage and wound healing of articular cartilage. PMID- 10514072 TI - Presence and expression of collagen adhesin gene (cna) and slime production in Staphylococcus aureus strains from orthopaedic prosthesis infections. AB - Prosthesis-associated infections still represent one of the most serious complications in the clinical use of biomaterials. The most frequent causes are Staphylococcus aureus and Staphylococcus epidermidis. Several studies have been devoted to identify adhesion mechanisms for these bacteria. Slime in particular has been extensively investigated. Recently, in Staphylococcus aureus species, considerable attention has been given to the host protein receptors that have been shown in in vitro assays to serve as substrates for bacterial adhesion. Collagen-rich tissues, as bone and cartilage, that are the preferential sites of staphylococcal infections, are also the tissues that harbour orthopaedic implants. These can be easily coated in vivo by collagen and thus become prone to adhesion of Staphylococci strains which carry the collagen adhesin gene (cna). In this study the frequency of cna was determined within a collection of 35 Staphylococcus aureus strains from orthopaedic prosthesis infections by a PCR method. Also the collagen-binding ability and slime forming capacity was evaluated. 29% of the strains were cna-positive and also able to bind collagen in vitro. 83% of the strains were slime forming. The results indicate that in the examined bacterial population slime-positive strains predominate over the cna positive strains, with a striking association of the two adhesion mechanisms in cna-positive strains. PMID- 10514073 TI - Fluorescence labeling to study platelet and leucocyte deposition onto vascular grafts in vitro. AB - Platelets and leucocytes are important participants in the response of the body to small diameter vascular grafts implanted into the arterial circulation. A sensitive and quick method for measuring platelet and leucocyte deposition contributes to material evaluation. With a newly developed fluorescence labeling method we examined the deposition of platelets and leucocytes onto vascular grafts in vitro. Human platelets and leucocytes were isolated and labeled with the fluorescence label Europium trichloride (EuCl3). After reconstitution of the labeled cells in plasma their functionality appeared intact and competitive with unlabeled cells. Eu-labeled platelets or leucocytes were then incubated with expanded polytetrafluoroethylene (ePTFE), Dacron and polyurethane (PU) vascular grafts in autologous plasma. Beta-thromboglobin and thromboxane release from platelets and beta-glucuronidase release from leucocytes during the incubation experiments were measured. Platelets and leucocytes deposited significantly less onto ePTFE compared to Dacron and polyurethane (P < 0.01). Our results are in accordance with results of in vivo studies using radio-active labeling to study platelet and leucocyte deposition. However, a new finding was that this reduced cell deposition may in part be due to possible toxic effects of ePTFE, shown by increased haemolysis and beta-thromboglobin release. PMID- 10514075 TI - Biological performance of a three-dimensional fabric as artificial cartilage in the repair of large osteochondral defects in rabbit. AB - A new artificial cartilage [A three-dimensional fabric (3-DF) comprising ultra high molecular weight polyethylene fiber with a triaxial three-dimensional structure and coated with hydroxyapatite] was used to repair large osteochondral defects in rabbit knees. The knees with the 3-DF implanted in the osteochondral defects were evaluated 2-24 weeks after the operation macroscopically and microscopically. The 3-DF coated with hydroxyapatite was fixed firmly to the subchondral bone with the newly formed bone into and around the 3-DF. Hyaline like cartilage was formed to a certain extent on the surface of the 3-DF, and the surface damage of the patella opposed to the 3-DF was minimal. In addition, the 3 DF caused no adverse effects such as particle disease, infection or severe synovitis during the experimental period. Consequently, the 3-DF seems to have successful biocompatibility in this rabbit experiment. PMID- 10514074 TI - Dissociation between complement activation, integrin expression and neutropenia during hemodialysis. AB - Complement activation, neutrophil stimulation, increased cellular adhesiveness, transient leukocyte margination and pulmonary leukostasis take place during hemodialysis with cellulosic dialysis membranes. Several investigators have hypothesized that complement activation is primarily responsible for the acute neutropenia occurring during the early phase of bio-incompatible hemodialysis. We have investigated the relationship between complement activation, levels of expression of CD11b and CD61 integrins on neutrophils and platelets, neutrophil counts and blood gas measurements in patients dialyzed with three types of membranes, known to activate the complement system to a different extent. Polysulfone, cellulose acetate and cuprophane membranes were used subsequently in six patients in a prospective cross-over trial design to reduce inter-individual variability. Increased levels of CD61 and CD11b, as well as neutropenia, were detected regardless of the type of membrane used. We observed a high inter individual variation with regard to complement activation suggesting varying susceptibility to dialysis membranes. We also report that the kinetics of anaphylatoxin generation were dissociated from those of the upregulation of adhesion molecules, early neutrophil margination and decrease in PaO2 during the first 30 min of hemodialysis. Similar results were obtained with all three types of dialysis membranes. The data strengthen the hypothesis that factors other than complement are involved in the induction of dialysis-related neutropenia and hypoxemia. PMID- 10514076 TI - Insertional tagging of at least two loci associated with resistance to adenine arabinoside in Toxoplasma gondii, and cloning of the adenosine kinase locus. AB - A genetic approach has been exploited to investigate adenylate salvage pathways in the protozoan parasite Toxoplasma gondii, a purine auxotroph. Using a new insertional mutagenesis vector designed to facilitate the rescue of tagged loci even when multiple plasmids integrate as a tandem array, 15 independent clonal lines resistant to the toxic nucleoside analog adenine arabinoside (AraA) were generated. Approximately two-thirds of these clones lack adenosine kinase (AK) activity. Parallel studies identified an expressed sequence tag (EST) exhibiting a small region of weak similarity to human AK, and this locus was tagged in several AK-deficient insertional mutants. Library screening yielded full-length cDNA and genomic clones. The T. gondii AK gene contains five exons spanning a approximately 3 kb locus, and the predicted coding sequence was employed to identify additional AK genes and cDNAs in the GenBank and dbEST databases. A genomic construct lacking essential coding sequence was used to create defined genetic knock-outs at the T. gondii AK locus, and AK activity was restored using a cDNA-derived minigene. Hybridization analysis of DNA from 13 AraA-resistant insertional mutants reveals three distinct classes: (i) AK-mutants tagged at the AK locus; (ii) AK- mutants not tagged at the AK locus, suggesting the possibility that another locus may be involved in regulating AK expression; and (iii) mutants with normal AK activity (potential transport mutants). PMID- 10514077 TI - Recombinant expression, purification, and characterization of Toxoplasma gondii adenosine kinase. AB - Toxoplasma gondii lacks the capacity to synthesize purines de novo, and adenosine kinase (AK)-mediated phosphorylation of salvaged adenosine provides the major route of purine acquisition by this parasite. T. gondii AK thus represents a promising target for rational design of antiparasitic compounds. In order to further our understanding of this therapeutically relevant enzyme, an AK cDNA from T. gondii was overexpressed in E. coli using the pBAce expression system, and the recombinant protein was purified to apparent homogeneity using conventional protein purification techniques. Kinetic analysis of TgAK revealed Km values of 1.9 microM for adenosine and 54.4 microM for ATP, with a k(cat) of 26.1 min(-1). Other naturally occurring purine nucleosides, nucleobases, and ribose did not significantly inhibit adenosine phosphorylation, but inhibition was observed using certain purine nucleoside analogs. Adenine arabinoside (AraA), 4-nitrobenzylthioinosine (NBMPR), and 7-deazaadenosine (tubercidin) were all shown to be substrates of T. gondii AK. Transgenic AK knock-out parasites were resistant to these compounds in cell culture assays, consistent with their proposed action as subversive substrates in vivo. PMID- 10514078 TI - Molecular cloning and characterization of a novel Schistosoma japonicum "irradiated vaccine-specific" antigen, Sj14-3-3. AB - A Schistosoma japonicum cDNA coding for a full length S. japonicum 14-3-3 protein was obtained by antibody screening of an adult worm cDNA library using sera taken from mice vaccinated with UV-attenuated cercariae, which are capable of transferring high levels of passive immunity to this parasite. The deduced amino acid sequence consists of 254 amino acids and is highly homologous with 14-3-3 family of proteins from a variety of species (55-69% identity). The recombinant S. japonicun 14-3-3 protein (rSj14-3-3) was expressed and purified in pGEX/E. coli, and in Western blotting was strongly recognised by sera from mice, rats and bovines vaccinated with irradiated S. japonicum cercariae. Analysis of mRNA showed that Sj14-3-3 is expressed in sporocysts and adult worms, but not in cercariae, however mouse antisera against rSj14-3-3 recognised a 29 kDa native antigen in antigen preparations made from eggs, cercariae, schistosomula and adult worms of S. japonicum indicating that this antigen is present in all life cycle stages. The presence of the native antigen in detergent extracts of intact schistosomula suggests that it is also present in the schistosomular tegument which is the most vulnerable target for immune attack. However, antisera against rSj14-3-3 did not recognise a similar band in S. mansoni or S. haematobium antigens, indicating that, like the UV-attenuated vaccines, this protein induced species-specific immune responses. Southern blot analysis suggested that there may exist more than one gene copy and/or polymorphism for Sj14-3-3. Immunoelectron microscopy confirmed that the native antigen is present throughout the body of adult worms including the tegument, but is less abundant in the muscles. The potential of rSj14-3-3 as a vaccine is now under further investigation. PMID- 10514079 TI - Conditional expression of glycosylphosphatidylinositol phospholipase C in Trypanosoma brucei. AB - Trypanosoma brucei glycosylphosphatidylinositol phospholipase C (GPIPLC) is expressed in the bloodstream stage of the life cycle, but not in the procyclic form. It is capable of hydrolyzing GPI-anchored proteins and phosphatidylinositol (PI) in vitro. Several roles have been proposed for GPIPLC in vivo, in the release of variant surface glycoprotein during differentiation or in the regulation of GPI and PI levels, but none has been substantiated. To explore GPIPLC function in vivo, tetracycline-inducible GPIPLC gene (GPIPLC) conditional knock-out bloodstream form and tetracycline-inducible GPIPLC-expressing procyclic cell lines were constructed. We were unable to generate GPIPLC null mutants. Cleavage of GPI-anchored proteins was abolished in extracts from uninduced conditional knock-outs and was restored upon induction. Despite the barely detectable level of GPIPLC activity in uninduced conditional knock-out bloodstream forms, their growth was not affected. GPI-protein cleavage activity could be induced in procyclic cell extracts, up to wild-type bloodstream levels. Myo-[3H]inositol incorporation into [3H]inositol monophosphate was about 14-fold lower in GPIPLC conditional knock-out bloodstream forms than in the wild type. Procyclic cells expressing GPIPLC showed a 28-fold increase in myo-[3H]inositol incorporation into [3H]inositol monophosphate and a 1.5-fold increase in [3H]inositol trisphosphate levels, suggesting that GPIPLC may regulate levels of inositol phosphates, by cleavage of PI and phosphatidylinositol 4,5-bisphosphate. PMID- 10514080 TI - The Leishmania mexicana proteasome. AB - As a start to understanding the importance of intracellular proteolysis in the protozoon Leishmania mexicana, the parasite proteasome has been purified and characterised. The L. mexicana proteasome is similar to proteasomes from other eukaryotes. It is soluble, and the 20S form has a mass of around 670 kDa, composed of at least 10 distinct subunits in the 22 to 32 kDa size range. The molecular mass of the L. mexicana proteasome increases to 1200 kDa in the presence of adenosine-5'-triphosphate, consistent with there being a 26S proteasome in the parasite. The purified 20S proteasome has activity towards substrates with hydrophobic, basic and acidic P, residues, and is sensitive to a range of peptide aldehyde inhibitors, as well as the proteasome-specific inhibitor lactacystin. The peptide aldehydes are able to arrest parasite growth in vitro with the same relative effectiveness as against the purified proteasome activity. The parasite population arrests with an increased 4N DNA content, indicating that, in part, the essential nature of the proteasome for L. mexicana proliferation is due to a role in the parasite cell cycle. Surprisingly, lactacystin is a relatively inefficient inhibitor of L. mexicana growth in vitro. PMID- 10514081 TI - Entamoeba histolytica lacks trypanothione metabolism. AB - Entamoeba histolytica lacks glutathione reductase activity and the ability to synthesise glutathione de novo. However, a recent report suggested that exogenous glutathione can be taken up and conjugated to spermidine to form trypanothione, a metabolite found so far only in trypanosomatids. Given the therapeutic implications of this observation, we have carefully analysed E. histolytica for evidence of trypanothione metabolism. Using a sensitive fluorescence-based HPLC detection system we could confirm previous reports that cysteine and hydrogen sulphide are the principal low molecular mass thiols. However, we were unable to detect trypanothione or its precursor N1-glutathionylspermidine [ < 0.01 nmol (10(6) cells)(-1) or < 1.7 microM]. In contrast, Trypanosoma cruzi epimastigotes (grown in a polyamine-supplemented medium) and Leishmania donovani promastigotes contained intracellular concentrations of trypanothione two to three orders of magnitude greater than the limits of detection. Likewise, trypanothione reductase activity was not detectable in E. histolytica [ < 0.003 U (mg protein)(-1)] and therefore at least 100-fold less than trypanosomatids. Moreover, although E. histolytica were found to contain trace amounts of glutathione (approximately 20 microM), glutathione reductase activity was below the limits of detection [ < 0.005 U (mg protein)(-1)]. These findings argue against the existence of trypanothione metabolism in E. histolytica. PMID- 10514082 TI - Intra and inter-specific microsatellite variation in the Leishmania subgenus Viannia. AB - Leishmania species of the subgenus Viannia are responsible for a large proportion of New World leishmaniasis. Here we report the development of a set of microsatellite markers which are able to discriminate between all species within the subgenus Viannia, including the closely related species pairs: Leishmania (V.) braziliensis and Leishmania (V.) peruviana; Leishmania (V.) panamensis and Leishmania (V.) guyanensis. Potential species hybrids were uncovered in the analysis. These markers are sufficiently polymorphic such that within-species epidemiological, population and genetic studies are theoretically possible for all species analyzed. PMID- 10514083 TI - Analysis of the gene expression profile of Schistosoma mansoni cercariae using the expressed sequence tag approach. AB - ESTs constitute rapid and informative tools with which to study gene-expression profiles of the diverse stages of the schistosome life cycle. Following a comprehensive EST study of adult worms, analysis has now targeted the cercaria, the parasite larval form responsible for infection of the vertebrate host. Two Schistosoma mansoni cercarial cDNA libraries were examined and partial sequence obtained from 957 randomly selected clones. On the basis of database searches, 551 (57.6%) ESTs generated had no homologs in the public databases whilst 308 (32.2%) were putatively identified, totaling 859 informative ESTs. The remaining 98 (10.2%) were uninformative ESTs (ribosomal RNA and non-coding mitochondrial sequences). By clustering analysis we have identified 453 different genes. The most common sequences in both libraries represented Sm8 calcium binding protein (8% of ESTs), fructose-1,6-bisphosphate aldolase, glyceraldehyde-3-phosphate dehydrogenase, cytochrome oxidase subunit 1, ATP guanidine kinase and triose phosphate isomerase. One hundred and nineteen identified genes were sorted into 11 functional categories, with genes associated with energy metabolism being the most abundant (13%) and diverse. The diversity and abundance of genes associated with the transcription/translation machinery and with regulatory/signaling functions were also marked. A paramyosin transcript was identified, indicating that this gene is not exclusively expressed in adult worms and sporocysts (as had been suggested previously). The possible physiological relevance to cercariae of the presence of transcripts with homology to calcium binding proteins of the EF hand superfamily, Gq-coupled rhodopsin photoreceptor, rod phosphodiesterase 8 subunit and peripheral-type benzodiazepine receptor is discussed. PMID- 10514084 TI - An unambiguous nomenclature for the major surface glycoproteins of the procyclic form of Trypanosoma brucei. PMID- 10514085 TI - Kinetics of release of amino acids by Leishmania major. PMID- 10514086 TI - Leishmania donovani proton translocating P-type adenosine triphosphatases LDH1A and LDH1B: trans-splicing and polyadenylation of transcripts in amastigotes and promastigotes. PMID- 10514087 TI - A beta-tubulin gene from Trichuris trichiura. PMID- 10514088 TI - Transfer of genes into Plasmodium falciparum by polyamidoamine dendrimers. PMID- 10514090 TI - Targeting T cells in myasthenia gravis. PMID- 10514089 TI - The major allelic dimorphisms in four Plasmodium falciparum merozoite proteins are not associated with alternative pathways of erythrocyte invasion. PMID- 10514091 TI - Imaging stroke in progress: magnetic resonance advances but computed tomography is poised for counterattack. PMID- 10514092 TI - Prevention of autoimmune attack by targeting specific T-cell receptors in a severe combined immunodeficiency mouse model of myasthenia gravis. AB - Myasthenia gravis (MG) is an autoimmune disease targeting the skeletal muscle acetylcholine receptor. We have previously demonstrated a selection bias of CD4+ T cells expressing the Vbeta5.1 T-cell receptor gene in the thymus of HLA-DR3 patients with MG. To evaluate the pathogenicity of these cells, severe combined immunodeficiency mice engrafted with MG thymic lymphocytes were treated with anti Vbeta5.1 antibody. Signs of pathogenicity (eg, acetylcholine receptor loss and complement deposits at the muscle end plates of chimeric mice) were prevented in anti-Vbeta5.1-treated severe combined immunodeficiency chimeras. Pathogenicity was mediated by autoantibodies against acetylcholine receptor. Thymic cells depleted of Vbeta5.1-positive cells in vitro before cell transfer were nonpathogenic, indicating that Vbeta5.1-positive cells are involved in the production of pathogenic autoantibodies. Acetylcholine receptor loss was prevented by Vbeta5.1 targeting in HLA-DR3 patients only, demonstrating specificity for HLA-DR3-peptide complexes. The action of the anti-Vbeta5.1 antibody involved both the in vivo depletion of Vbeta5.1-expressing cells and an increase in the interferon-gamma/interleukin-4 ratio, pointing to an immune deviation-based mechanism. This demonstration that a selective and specific T helper cell population is involved in controlling pathogenic autoantibodies in MG holds promise for the treatment of MG. PMID- 10514093 TI - Longitudinal magnetic resonance imaging study of perfusion and diffusion in stroke: evolution of lesion volume and correlation with clinical outcome. AB - A prospective longitudinal diffusion-weighted and perfusion-weighted magnetic resonance imaging (DWI/PWI) study of stroke patients (n = 21) at five distinct time points was performed to evaluate lesion evolution and to assess whether DWI and PWI can accurately and objectively demonstrate the degree of ischemia-induced deficits within hours after stroke onset. Patients were scanned first within 7 hours of symptom onset and then subsequently at 3 to 6 hours, 24 to 36 hours, 5 to 7 days, and 30 days after the initial scan. Lesion evolution was dynamic during the first month after stroke. Most patients (18 of 19, 95%) showed increased lesion volume over the first week and then decreased at 1 month relative to 1 week (12 of 14, 86%). Overall, lesion growth appeared to depend on the degree of mismatch between diffusion and perfusion at the initial scan. Abnormal volumes on the acute DWI and PWI (<7 hours) correlated well with initial National Institutes of Health (NIH) stroke scale scores, outcome NIH stroke scale scores, and final lesion volume. DWI and PWI can provide an early measure of metabolic and hemodynamic insufficiency, and thus can improve our understanding of the evolution and outcome after acute ischemic stroke. PMID- 10514094 TI - Language recovery after left hemispherectomy in children with late-onset seizures. AB - We investigated the language capabilities of the isolated right hemisphere in 6 children (age, 7-14 years) after left hemidecorticectomy for treatment of Rasmussen's syndrome. Patients were right-handed before surgery and had at least 5 years of normal language development before the onset of seizures. Language testing included speech sound (phoneme) discrimination, single word and phrasal comprehension, repetition, and naming. Within 4 to 16 days after surgery, patients showed improved phoneme discrimination compared with their performance shortly before surgery. Other language functions remained severely impaired until at least 6 months after surgery. By 1 year after surgery, receptive functions were comparable with, or surpassed, patient presurgery performance. Although word repetition was intact by 1 year after surgery, naming remained impaired, and patient speech was limited largely to production of single words. These results suggest that the right hemisphere is innately capable of supporting multiple aspects of phoneme processing. Recovery of higher level receptive and, to a lesser extent, expressive language functions is attributed to plasticity of the right hemisphere, which appears to persist beyond the proposed critical period for language acquisition and lateralization. PMID- 10514095 TI - Participation of prostate apoptosis response-4 in degeneration of dopaminergic neurons in models of Parkinson's disease. AB - Dysfunction and death of midbrain dopaminergic neurons underlies the clinical features of Parkinson's disease (PD). Increasing evidence suggests roles for oxidative stress and a form of cell death called apoptosis in the pathogenesis of PD. We recently identified a 38-kd protein called prostate apoptosis response-4 (Par-4), which is rapidly induced in cultured neurons after exposure to apoptotic insults, and appears to play a necessary role in the cell death process. We now report that Par-4 levels increase dramatically in midbrain dopaminergic neurons of monkeys and mice exposed to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The increase in Par-4 levels occurs in both neuronal cell bodies in the substantia nigra and their axon terminals in the striatum, and precedes loss of tyrosine hydroxylase immunoreactivity and cell death. In the monkey model, Par-4 levels were also increased in several brain regions (red nucleus, lateral geniculate nucleus, and cerebral cortex) in which functional alterations have previously been documented in PD patients and MPTP-treated monkeys. Exposure of cultured human dopaminergic neural cells to the complex I inhibitor rotenone, or to Fe2+, resulted in Par-4 induction, mitochondrial dysfunction, and subsequent apoptosis. Blockade of Par-4 induction by antisense treatment prevented rotenone- and Fe2+-induced mitochondrial dysfunction and apoptosis demonstrating a critical role for Par-4 in the cell death process. The data suggest that Par-4 may be involved in the neurodegenerative process in PD. PMID- 10514096 TI - Evidence of active nerve cell degeneration in the substantia nigra of humans years after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine exposure. AB - This report provides the first detailed neuropathological study of 1-methyl-4 phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced parkinsonism in humans. All 3 subjects self-administered the drug under the impression it was "synthetic heroin" and subsequently developed severe and unremitting parkinsonism, which was L-dopa responsive, at least in the earlier stages of illness. Survival times ranged from 3 to 16 years. Neuropathological examination revealed moderate to severe depletion of pigmented nerve cells in the substantia nigra in each case. Lewy bodies were not present. In Patients 1 and 2, there was gliosis and clustering of microglia around nerve cells. Patient 3 had a similar picture and also showed large amounts of extraneuronal melanin. These findings are indicative of active, ongoing nerve cell loss, suggesting that a time-limited insult to the nigrostriatal system can set in motion a self-perpetuating process of neurodegeneration. Although the mechanism by which this occurs is far from clear, the precedent set by the cases could have broad implications for human neurodegenerative disease. PMID- 10514097 TI - Migraine without aura: a population-based twin study. AB - To investigate the importance of genetic and environmental factors to the etiology of migraine without aura and to compare the symptomatology of migraine without aura in monozygotic and dizygotic twins, 2,680 twin pairs were recruited from the population-based Danish Twin Registry. Monozygotic (MZ) and same-sex dizygotic (DZ) twin pairs, where at least one twin had self-reported migraine or self-reported severe headache with accompanying symptoms, were telephone interviewed by a physician. The participation rate in the telephone interview was 90%. The pairwise concordance rate was significantly higher in MZ than in DZ twin pairs (28% vs 18%). The probandwise concordance rate was 40% (95% CI, 33-48%) in MZ and 28% (95% CI, 23-33%) in DZ twin pairs. The pairwise concordance rates for the different pain characteristics and accompanying symptoms were not significantly different in MZ and DZ twin pairs. However, comparing all of the pairwise concordance rates of pain characteristics and accompanying symptoms together, MZ twin pairs were significantly more concordant than DZ twin pairs. Our data demonstrate a significant genetic factor in migraine without aura. The size of this factor is modest and the demonstration of susceptibility genes is predicted to be laborious and difficult. PMID- 10514098 TI - Linkage and association analysis of susceptibility regions on chromosomes 5 and 6 in 106 Scandinavian sibling pair families with multiple sclerosis. AB - In the genetically homogeneous Scandinavian population, we have investigated chromosome 5 and the HLA (human leukocyte antigen) region on chromosome 6p21 by applying linkage and association analyses on 106 white sibling pair families with multiple sclerosis. The importance of genes within the HLA region for the susceptibility of multiple sclerosis has previously been reported. More recently, findings have suggested importance of regions on chromosome 5. Half of chromosome 5 was analyzed by using 14 microsatellite markers and a susceptibility region with a maximum LOD score of 1.1 was identified. Chromosome 6 was analyzed by HLA DR typing and using the TNF alpha microsatellite marker. A peak maximum LOD score of 2.0 was found at the HLA-DR marker. Association studies were made for all the markers, comparing 106 probands from the sibling pairs with 100 unrelated controls. None of the markers on chromosome 5 showed significant association with multiple sclerosis, whereas strong association between multiple sclerosis and DR2 was found, with an odds ratio of 3.7 (p < 10(-5)). It is surprising that association was not seen for any of the TNF alpha alleles including the 121-bp allele, although this allele was in positive linkage disequilibrium with DR2 in both patients and controls. Our results support the existence of multiple sclerosis susceptibility loci on chromosomes 5p and 6p21. PMID- 10514100 TI - Magnetoencephalographic localization in pediatric epilepsy surgery: comparison with invasive intracranial electroencephalography. AB - The object of this study was to determine the concordance of the anatomical location of interictal magnetoencephalographic (MEG) spike foci with the location of ictal onset zones identified by invasive ictal intracranial electroencephalographic recordings in children undergoing evaluation for epilepsy surgery. MEG was performed in 11 children with intractable, nonlesional, extratemporal, localization-related epilepsy. Subsequently, chronic invasive intracranial electroencephalographic monitoring was performed by using subdural electrodes to localize the ictal onset zone and eloquent cortex. Based on the invasive monitoring data, all children had excision of, or multiple subpial transections through, ictal onset cortex and surrounding irritative zones. In 10 of 11 patients, the anatomical location of the epileptiform discharges as determined by MEG corresponded to the ictal onset zone established by ictal intracranial recordings. In all children, the anatomical location of the somatosensory hand area, determined by functional mapping through the subdural electrode array, was the same as that delineated by MEG. Nine of 11 patients became either seizure-free or had a greater than 90% reduction in seizures after surgery, with a mean follow-up of 24 months. MEG is a powerful and accurate tool in the presurgical evaluation of children with refractory nonlesional extratemporal epilepsy. PMID- 10514099 TI - Phenotypic variation in hereditary frontotemporal dementia with tau mutations. AB - Several mutations in the tau gene have been found in families with hereditary frontotemporal dementia and parkinsonism linked to chromosome 17q21-22 (FTDP-17). This study is the first attempt to correlate genotype and phenotype in six families with FTDP-17 with mutations in the tau gene (deltaK280, G272V, P301L, and R406W). We have investigated tau pathology in 1 P301L and 1 R406W patient. The R406W family showed a significantly higher age at onset (59.2 +/- 5.5 years) and longer duration of illness (12.7 +/- 1.5 years) than the families with the other mutations. The six families showed considerable variation in clinical presentation, but none of them had early parkinsonism. Mutism developed significantly later in the R406W family than in the other families. Frontotemporal atrophy on neuroimaging in the R406W family was less severe than in the P301L and deltaK280 families. The P301L brain contained many pretangles in the frontal and temporal cortex, and the dentate gyrus of hippocampus, showing three tau bands (64, 68, and 72 kd) of extracted tau from the frontal cortex. The presence of many neurofibrillary tangles, many diffuse and classic neuritic plaques in the temporal and parietal cortex, and the hippocampus of the same P301L brain correlated with the presence of four sarkosyl-insoluble (60, 64, 68, and 72 kd) tau bands. The coexistence of characteristic P301L and Alzheimer pathology in the same brain needs further explanation. The R406W brain showed abundant neurofibrillary tangles in several brain regions, and four tau bands (60, 64, 68, and 72 kd) of extracted tau from these regions. The slower progression of the disease in the R406W family might be explained by the microtubule-binding properties of the mutant protein. PMID- 10514101 TI - Hyperekplexia phenotype due to compound heterozygosity for GLRA1 gene mutations. AB - Hyperekplexia (MIM 149400), or startle disease, is a neurological disorder characterized by generalized stiffness during the neonatal period, excessive startle reflexes, and generalized stiffness related to the startle response. Linkage analysis mapped a major gene for this disorder to chromosome 5q33-35. Subsequently, mutations in the GLRA1 gene, encoding the alpha1 subunit of the glycine receptor, were found in hyperekplexia families with an autosomal dominant or recessive inheritance pattern. In the present study, we describe the genetic analysis of the GLRA1 gene of a family consisting of 2 children with hyperekplexia, 2 nonaffected children, and their healthy nonconsanguineous parents. Although the pedigree suggested the presence of a recessive mutation, haplotype construction showed that the 2 affected children shared the same haplotype combination in which the maternal haplotype differed from the paternal haplotype, suggesting the presence of compound heterozygosity. Mutation analysis revealed different missense mutations on the two haplotypes, changing an arginine to a histidine at amino acid positions 252 and 392, respectively. It is interesting that the hyperekplexia phenotype was only seen in individuals compound heterozygous for the two mutations, whereas family members carrying either one of the two mutations had no clinical signs. PMID- 10514102 TI - Novel functional epsilon-subunit polypeptide generated by a single nucleotide deletion in acetylcholine receptor deficiency congenital myasthenic syndrome. AB - Acetylcholine receptor (AChR) deficiency is a recessively inherited congenital myasthenic syndrome in which fatigable muscle weakness results from impaired neuromuscular transmission caused by reduced AChR numbers. In mature muscle, AChRs consist of alpha2 betadelta together with the adult-specific epsilon subunit. We have identified a deletion of the first nucleotide in exon 12 of the AChR epsilon-subunit gene (epsilon1267delG) and demonstrate its recessive inheritance segregates with disease in 6 unrelated cases of AChR deficiency. In addition, we found that both healthy and AChR-deficient muscle contain a population of AChR epsilon-subunit mRNA transcripts that retain intron 11. We investigated the possible consequences of combining this mutation with the alternative mRNA species through AChR expression studies in human embryonic kidney cells and Xenopus oocytes. Epsilon1267delG generates a polypeptide that lacks M4 and is not detected in surface AChR, whereas retention of intron 11 in the RNA transcript restores the reading frame, conserves M4, and generates a polypeptide that is incorporated into functional surface AChR, although at a reduced level, consistent with the disease phenotype. Our results indicate that for some AChR deficiency mutations located between M3 and M4, the retention of intron 11 in the epsilon-subunit mRNA transcripts may rescue adult AChR function. PMID- 10514103 TI - Sleep disorders in children with blindness. AB - To evaluate the frequency and type of sleep disorders seen in blind children compared with matched controls, a 42-item questionnaire was used on 156 children (77 blind children) ranging from 3 to 18 years of age. A total of 17.4% of blind children reported sleeping less than 7 hours per night on weekdays compared with 2.6% of controls, with blind children awakening much earlier. Blind children had more sleep complaints, and 13.4% of blind subjects had daily episodes of involuntary sleepiness compared with 1.3% of controls. Blindness has an impact on sleep and alertness that adds to the primary disability. PMID- 10514104 TI - Serial magnetic resonance imaging in patients with Balo's concentric sclerosis: natural history of lesion development. AB - We reviewed the magnetic resonance imaging scans from 22 serial studies of 5 patients with Balo's concentric sclerosis collected during the past 3 years. The data showed the concentric lesions did not occur simultaneously but developed step by step in a centrifugal direction. The development of lesions was preceded by an enhancing ring relatively devoid of demyelination and was followed by progressive demyelination occurring mainly at the inner aspect of the enhancement. The same process recurred on the edge of the previous enhanced zone. Thus, an appearance of concentric rings with alternating demyelinated and relatively myelin-preserved bands was formed. PMID- 10514105 TI - Irregular distribution of cytochrome c oxidase protein subunits in aging and Alzheimer's disease. AB - This study aims to investigate the cellular distribution of human cytochrome c oxidase (COX) subunit II (CII) and COX subunit IV (CIV) in Alzheimer's disease relative to control brains. The levels of CIV and CII proteins in the cerebellar Purkinje cells were reduced in age-matched controls relative to young controls and in the Alzheimer's disease group relative to both age-matched and young controls. Results suggest that these age-associated changes are more marked in Alzheimer's disease. PMID- 10514106 TI - Diffuse pachygyria with cerebellar hypoplasia: a milder form of microlissencephaly or a new genetic syndrome? AB - We report on 2 families with diffuse pachygyria and cerebellar hypoplasia, who presented hypotonia, ataxia, seizures, and developmental delay since infancy. Computed tomography (CT) and magnetic resonance imaging (MRI) revealed decreased gyral formation in the cerebral cortex and marked hypoplasia in the cerebellum. Cerebellar hypoplasia is often associated with type 2 lissencephaly; however, our cases showed no polymicrogyria, and their clinical findings were quite mild compared with those of microlissencephaly. Their characteristic phenotype suggested a new genetic syndrome, which was possibly inherited as an autosomal recessive trait. PMID- 10514107 TI - Association between Alzheimer's disease and the NOS3 gene. AB - Alzheimer's disease (AD) is the most common form of neurodegenerative disorder of later life. Genetic studies have demonstrated that the apolipoprotein E (ApoE) gene is an important susceptibility locus; however, other environmental and genetic factors operating alone or in combination with ApoE must also be involved. Among candidate genes that may contribute to this residual risk is the endothelial nitric oxide synthase (NOS3) gene. NO release from vascular endothelium accounts in large part for endothelium-derived relaxing factor bioactivity. Abnormalities of cerebral small vessels occur early in AD, and it has been demonstrated recently that beta-amyloid interacts with endothelial cells in blood vessels to produce an excess of superoxide radicals. We have genotyped 122 cases of early-onset AD (EOAD) and 317 cases of late-onset AD (LOAD) as well as 392 controls for a common structural polymorphism Glu/Asp at codon 298 in the NOS3 gene. We find a highly significant enrichment for Glu/Glu homozygotes in LOAD compared with controls. The effect appears to be independent of ApoE status. NOS3 may be a new genetic risk factor for LOAD. PMID- 10514108 TI - Quantitative near-infrared spectroscopy discriminates between mitochondrial myopathies and normal muscle. AB - Five patients with chronic progressive external ophthalmoplegia (CPEO) and 27 healthy controls were examined by near-infrared spectroscopy (NIRS) for the noninvasive and direct quantitative measurement of muscle oxygen consumption and forearm blood flow. NIRS measurements were obtained in rest and during static isometric handgrip exercise at 10% of the maximum voluntary contraction (MVC) force. A significantly decreased oxygen consumption at rest as well as during exercise was found in patients with CPEO. Our results suggest that NIRS is able to discriminate between CPEO patients and healthy controls, which makes NIRS a valuable tool in the diagnostic workup of patients suspected to have a mitochondrial myopathy. PMID- 10514109 TI - A locus for febrile seizures (FEB3) maps to chromosome 2q23-24. AB - Febrile seizures are the most common form of childhood seizures, occurring in 2% to 5% of North American children. We report a large Utah family with 21 members affected by febrile seizures inherited as an autosomal dominant trait. All had generalized tonic-clonic seizures with onset associated with fever, consistent with the consensus febrile seizure phenotype, and none had febrile seizures beyond 6 years of age. Eighteen affected individuals had recurrent febrile seizures. Eight individuals developed afebrile seizures between ages 5 and 13 years. Afebrile seizures consisted of generalized tonic-clonic, generalized tonic, generalized atonic, simple partial, and partial complex seizure types and were associated with abnormal electroencephalographic findings in 5 individuals, all of whom were intellectually normal. We undertook linkage analysis in this family, defining the disease phenotype as febrile seizures alone. Linkage analysis in epilepsy candidate gene/loci regions failed to show evidence for linkage to febrile seizures. However, a genomewide scan and subsequent fine mapping revealed significant evidence for a new febrile seizure locus (FEB3) on chromosome 2q23-24 with linkage to the marker D2S2330 (LOD score 8.08 at theta = 0.001). Haplotype analysis defined a critical 10-cM region between markers D2S141 and D2S2345 that contains the FEB3 locus. PMID- 10514110 TI - Prescribing habits in primary biliary cirrhosis: a national survey. AB - BACKGROUND: Primary biliary cirrhosis (PBC) is a chronic cholestatic liver disease of unknown aetiology. A number of drugs have been used in its treatment, but only ursodeoxycholic acid (UDCA) has been shown to improve survival. Our aims were to determine the current prescribing habits in PBC of all practising gastroenterologists in the UK. METHODS: A postal questionnaire was sent to 454 gastroenterologists in 1996, followed by a second questionnaire a month later to the non-responders. RESULTS: Of 454 doctors sent questionnaires, 379 (83%) replied. Of these, 58 were excluded from further analysis as they were not practising gastroenterologists. There are an estimated 4337 patients with PBC being seen by gastroenterologists in hospitals. Of these, only 1376 (32%) are being seen in liver units. Ninety-one per cent of gastroenterologists look after patients with PBC (median 10 patients, range 1-500). Ninety-five per cent of gastroenterologists prescribe UDCA but there is a large dose range (median 11.5 mg/kg/day, range 1.5-23.1). Of these, 93% also prescribe cholestyramine. Only 45 (14%) gastroenterologists prescribed other treatments for PBC (13 colchicine, 24 steroids, nine penicillamine, 13 immunosuppressants). Only 53 (17%) treat the symptoms/complications of PBC (37 fat-soluble vitamins, 15 calcium, six bisphosphonates, one hormone replacement therapy, 10 antihistamines, 10 rifampicin). CONCLUSIONS: UDCA is being prescribed for PBC by the majority of practising gastroenterologists but over a wide dose range. Very few gastroenterologists are using preventive treatment for osteoporosis in this high risk group. Other treatments, as yet unproven in trials, are being prescribed by a minority of gastroenterologists. PMID- 10514111 TI - Diversity of core promoter mutations in immune clearance phase of chronic HBV infection. AB - BACKGROUND/AIMS: Transcription of HBV (hepatitis B virus) pre-core and pre genomic mRNAs is controlled by core promoter. Therefore, mutations in the core promoter region might change the activity of liver diseases through an altered transcriptional level of the mRNA. The present study was carried out to determine the diversity of HBV core promoter sequences in chronic HBV carriers. METHODS: DNA sequences in the core promoter region were determined after cloning the PCR product. Two groups of chronic HBV carriers with HBeAg, including five cases of asymptomatic carriers (ASCs, 21 clones) and eight with chronic hepatitis (CH, 50 clones) were studied. RESULTS: Mutations in the core promoter were found in three out of the ASCs (11 clones), and in all eight cases in the CH group (48 clones). While mutations at nucleotide 1762 (A-->T) and 1764(G-->A) were not found in ASC, mutations at the same positions were found in all the cases of CH group (40 clones) (P=0.003). Diverse patterns of mutations in the core promoter were observed in each patient in the CH group. CONCLUSIONS: Further studies are needed to determine whether the diversity of HBV core promoter mutations has clinical significance such as the seroconversion of HBeAg to anti-HBe. PMID- 10514112 TI - Comparison of assays for N-amino terminal propeptide of type III procollagen in chronic hepatitis C by using receiver operating characteristic analysis. AB - OBJECTIVE: During the process of liver fibrosis, type III procollagen is converted to type III collagen by cleavage of its amino terminal and carboxy terminal propeptides. Serum levels of amino terminal propeptide of type III procollagen (PIIINP) are a marker of collagen turnover in liver fibrosis. Two assays for PIIINP are available, one which measures both Col 1-3 (collagen synthesis) and Col 1 (collagen degradation) peptides, and one which measures Col 1-3 only. Using receiver operating characteristic analysis, the two PIIINP assays were compared with serum ALT as markers of liver disease in chronic hepatitis C. METHODS: Serum PIIINP was measured using both assays in 33 patients with chronic hepatitis C and five healthy controls. Liver biopsies in chronic hepatitis C patients were scored using a previously described grading and staging system. RESULTS: Serum PIIINP was significantly elevated in chronic hepatitis C compared to controls using both the combined Col 1-3 and Col 1 (median 0.61 vs 0.33 U/ml, P=0.001) and Col 1 assays (median 6.5 vs 3.5 microg/l, P=0.006). Serum PIIINP measured by the combined assay was significantly related to liver fibrosis, periportal necrosis and histological activity index (P<0.05). The area under the curve for specificity and sensitivity in detecting advanced liver disease was only significant for the combined assay (P=0.017). Serum PIIINP measured by the Col 1 assay was not related to these indices of disease severity while serum ALT was only related to portal inflammation. CONCLUSION: A serum PIIINP assay which measures both Col 1-3 and Col 1 peptides instead of Col 1-3 peptide alone is more predictive of severity of liver disease and should be used in preference as a non invasive marker of liver injury in chronic hepatitis C. PMID- 10514113 TI - Fatigue does not correlate with the degree of hepatitis or the presence of autoimmune disorders in chronic hepatitis C infection. AB - BACKGROUND: Fatigue is probably the most commonly reported symptom in chronic hepatitis C virus (HCV) infection. It is unclear whether fatigue is related to the severity of underlying liver disease or other autoimmune disorders often described with chronic HCV infection. OBJECTIVE: To quantify fatigue in terms of its impact on quality of life in a homogeneous cohort and examine its relationship to the status of liver disease or associated autoimmunity. METHODS: The Fatigue Impact Scale (FIS) questionnaire (Fisk et al. Clin Infect Dis 1994; 18:S79-S83), a recently validated psychometric tool for assessing patients' perceptions of the functional limitations attributable to fatigue (40 statements; three subscales: physical, cognitive and psychological; maximum score = 160), was applied to a cohort of Irish women who were PCR-positive for HCV genotype 1b via inoculation with contaminated anti-D products in 1977. RIBA-positive, PCR negative patients (n = 20) and healthy age-matched women (n = 50) served as controls. The degree of hepatitis was assessed using the Knodell histological activity index (HAI) score on previous liver biopsies. Clinical and laboratory evidence of cryoglobulinaemia, Sjogren's syndrome, connective tissue diseases, autoimmune thyroid disease and glomerulonephritis was sought. RESULTS: The mean FIS score of the 66 PCR-positive women (mean 78+/-36; range 7-153) was significantly higher than in age-matched controls (mean 31+/-24, range 0-78, P<0.001) but not statistically different from that of the RIBA-positive, PCR negative group. The FIS score did not correlate with the HAI score (median HAI = 4; range 2-9; Pearson's correlation coefficient r=0.01, P=0.9). Significant levels of cryoglobulins were detected in 10 (15.2%). The sicca complex was diagnosed in six patients, three of whom had associated cryoglobulinaemia. Thyroid antibodies, anti-nuclear antibody, rheumatoid factor, antimitochondrial antibody and anti-smooth muscle antibody were detected in 15.2%, 6%, 4.5%, 4.5% and 1.5%, respectively. There was no significant difference in the FIS score between the groups with autoimmune diseases and those without. The FIS score of the nine patients previously treated with interferon was not statistically different from the untreated group (P=0.39). CONCLUSION: The perceived functional impact of fatigue on quality of life is significantly higher in patients with chronic HCV genotype 1b infection compared to healthy controls. However, it is unrelated to the degree of hepatitis and cannot be accounted for by the co existence of autoimmune disorders alone. PMID- 10514114 TI - Increased release of interleukin-6 by oesophageal mucosa in children with reflux oesophagitis. AB - OBJECTIVE: To evaluate the release of interleukin-6 (IL-6) by oesophageal mucosa and to establish the serum levels of IL-6 and C-reactive protein (CRP), and plasma fibrinogen in children with reflux oesophagitis. DESIGN: In a prospective study, IL-6 release by tissue fragments obtained from oesophageal biopsies was determined and serum IL-6 and CRP as well as plasma fibrinogen were analysed. METHODS: The study population comprised ten children with reflux oesophagitis, diagnosed on the basis of 24 h oesophageal pH monitoring and endoscopy with biopsies. Ten children with recurrent abdominal pain were studied for comparative purposes. Biopsy tissue fragments were processed to obtain a cell suspension and the release of IL-6 was determined in culture medium. Serum IL-6 levels were measured by ELISA, serum CRP by turbidimetry, and plasma fibrinogen by spectrophotometry. RESULTS: Oesophageal cells obtained from reflux oesophagitis patients synthesize and release in vitro a significantly higher amount of IL-6 than controls (71.26+/-19.5 versus 31.67+/-8.02 pg/10(6) cells; P<0.01). Serum IL 6, serum CRP and plasma fibrinogen levels were not statistically different between patients with reflux oesophagitis and controls. CONCLUSIONS: These results suggest a short-term action of IL-6 since its effects could be exerted only in the microenvironment of the oesophageal mucosa. PMID- 10514115 TI - Different prevalences of reflux oesophagitis and hiatus hernia among dyspeptic patients in England and Singapore. AB - OBJECTIVE: To compare the frequency of reflux oesophagitis and hiatus hernia in dyspeptic patients in England with that in Singapore. DESIGN: Demographic, clinical and endoscopic findings in consecutive dyspeptic patients seen in England and Singapore by the same clinician were compared. The association of various factors with the occurrence of hiatus hernia and oesophagitis was analysed by logistic regression. SETTING: District general hospital in England and university hospital in Singapore. PARTICIPANTS: The English series comprised 212 consecutive patients, and 173 patients were seen in Singapore. RESULTS: Reflux oesophagitis and hiatus hernia were found in 52 (25%) and 50 (49%) of the English patients, and 12 (6%) and 7 (4%) of the Singapore patients, respectively (P<0.005 in each case). Race, body mass index and age were independently associated with hiatus hernia (odds ratios 3.07, 1.08 and 1.04, respectively). The risk factors for oesophagitis were race, sex, body mass index and age (odds ratios 4.04, 2.37, 1.11 and 1.02, respectively). If hiatus hernia was included in the analysis, the risk factors were hiatus hernia, sex, race and body mass index (odds ratios 20.10, 3.07, 2.81 and 1.09, respectively). CONCLUSIONS: Reflux oesophagitis and hiatus hernia are more common in English dyspeptic patients compared to those in Singapore. The most important risk factor for both oesophagitis and hiatus hernia is race. PMID- 10514116 TI - Endoscopy for Helicobacter pylori sero-negative young dyspeptic patients: an economic evaluation based on a randomized trial. AB - BACKGROUND: A policy of withholding endoscopy in Helicobacter pylori sero negative dyspeptic patients without sinister symptoms saves up to 36% of endoscopies. However, it is unclear whether the net cost of applying this policy outweighs that of conventional management. AIM: To determine the direct (healthcare) and indirect (productivity) costs of applying a strategy of endoscopy versus no endoscopy in H. pylori sero-negative young dyspeptics in the UK. METHOD: The direct and indirect incremental costs for both strategies were calculated amongst 154 H. pylori seronegative subjects randomized to have an endoscopy or no endoscopy before subsequent management by their general practitioners. The cost per patient of each strategy was calculated using reference values in our clinical setting and sensitivity analysis was used to test different scenarios. RESULTS: The total direct cost rose for the endoscopy group (mean Pound Sterling 103, 95% CI 78 to 127) but did not change for the non endoscopy group (mean Pound Sterling 6, 95% CI -32 to 44). On average, direct (healthcare) costs for patients in the endoscopy group rose by Pound Sterling 96 (95% CI 51 to 142) more than those for non-endoscopy patients. Indirect (productivity) cost fell for the non-endoscopy group (mean -Pound Sterling 40, 95% CI -220 to 140) compared to a rise for the endoscopy group (mean Pound Sterling 180, 95% CI -60 to 420) (difference not significant). The total cost (including direct and indirect costs) fell for the non-endoscopy group (mean Pound Sterling 34, 95% CI -228 to 160) but rose for the endoscopy group (mean Pound Sterling 283, 95% CI 32 to 533)--an incremental cost of Pound Sterling 317 (95% CI 0 to 634). For all assumptions in the sensitivity analysis, the mean cost in the endoscopy group was at least Pound Sterling 200 higher than in the non endoscopy group. CONCLUSIONS: It is less expensive to manage H. pylori-negative dyspeptic patients aged under 45 without sinister symptoms by withholding endoscopy. PMID- 10514117 TI - Reducing the endoscopic workload: does serological testing for Helicobacter pylori help? AB - OBJECTIVE: Serological screening for Helicobacter pylori (H. pylori) in young (< or = 45 years) dyspeptic patients has been used to avoid oesophago-gastro duodenoscopy (OGD). We used serology to identify seronegative and seropositive patients without sinister symptoms and combined approaches of avoiding OGD in both groups. We aimed to determine the reduction of OGD in this group. DESIGN: Prospective study on the treatment of 232 patients with dyspepsia. SETTING: Six hundred and fifty bed district general hospital serving rural Northamptonshire, UK. INTERVENTIONS: Two hundred and thirty-two patients referred by local general practitioners for OGD were offered serology. Symptom severity was scored using a questionnaire. One hundred and eleven seronegative patients received symptomatic treatment, 105 seropositive patients received triple therapy for 1 week. Sixteen patients with equivocal results were offered OGD. Patients were followed up after 6 months. MAIN OUTCOME MEASURES: Severity of dyspepsia symptoms and proportion of patients returning for OGD. RESULTS: Fifteen equivocal patients underwent OGD, one refused. Forty-six patients (33 seronegative, 13 seropositive) had persisting symptoms and underwent OGD. Mean symptom severity was reduced significantly in equivocal (P<0.01), seronegative (P<0.001) and seropositive (P<0.001) patients. Fewer seronegative patients were symptom-free at follow up compared to seropositive patients (n = 15 (16%) vs n = 48 (51%); P<0.001), 171 patients avoided OGD, a 74% reduction. CONCLUSIONS: Use of H. pylori serology in the management of young dyspeptic patients without sinister symptoms can reduce the OGD workload by 74%, decreasing the length of time that older patients, who are at greater risk of malignant disease, may have to wait for OGD. PMID- 10514118 TI - A comparison of two rapid whole-blood tests and laboratory serology, in the diagnosis of Helicobacter pylori infection. AB - OBJECTIVE: To compare two rapid whole-blood serology tests for Helicobacter pylori and a laboratory serology assay against a gold standard. DESIGN: Prospective comparison of tests in 81 patients. SETTING: A hospital rapid access endoscopy clinic. PARTICIPANTS: Dyspeptic patients requiring assessment of H. pylori status. INTERVENTIONS: Measurement of H. pylori antibody status by Quickvue One-step, Helisal, and Premier H. pylori test; 13C urea breath test for H. pylori, and gastric biopsies for histology, culture and rapid urease test. MAIN OUTCOME MEASURE: Sensitivity and specificity of Quickvue One-step, Helisal and Premier tests, compared to a gold standard based on 13C urea breath test, biopsy culture, histology and urease test. RESULTS: The Quickvue assay has significantly greater sensitivity (81%) than Helisal (67%), but without appreciable loss of specificity (86% and 93%, respectively). The Premier laboratory assay is significantly more sensitive than both of the rapid blood tests (96%), with comparable specificity to the Quickvue assay. CONCLUSION: The rapid serology tests used in this study are quick and convenient to use, but do not approach the sensitivity of a laboratory assay in detecting H. pylori status in this group of dyspeptic patients attending an endoscopy clinic. PMID- 10514119 TI - Determinants of Helicobacter pylori seroprevalence among Italian blood donors. AB - BACKGROUND/AIM: Helicobacter pylori is a worldwide infection. It is estimated that approximately 50% of the general population is affected, but this percentage varies considerably between countries. To investigate the prevalence of H. pylori infection, a cross-sectional epidemiological study, based on the serological determination of the IgG antibodies against H. pylori, was carried out in healthy Italian blood donors by using a commercially available kit. METHODS: From March 1995 to March 1997, a total of 2598 consecutive volunteer blood donors were tested for the presence of antibodies against H. pylori. All patients answered a detailed questionnaire which collected sociodemographic characteristics, and smoking, alcohol drinking and dietary habits. Test-positive subjects with gastrointestinal symptoms underwent endoscopy, with biopsies taken for histological diagnosis. RESULTS: The global prevalence of H. pylori infection in our study was 1161/2598 (45%). It was directly correlated with age (67% in subjects aged > or = 50 years). The prevalence of H. pylori infection was higher in men (46.4%) than women (38.4%), and more frequent in patients with a low educational level, in the lower quintile of height and in the upper quintile of body mass index (BMI). No significant association with smoking and alcohol drinking was found. Inverse associations were found with elevated consumption of milk (chi-square for trend 5.49, P < 0.05), but not other examined food groups. Multivariate analysis selected sex, age, BMI and educational level as the variables independently related to H. pylori infection. CONCLUSION: This study confirms relatively high prevalence of H. pylori seropositivity among Italian healthy adults and points to sex, age, BMI and sociocultural class as persisting determinant features of H. pylori infection. PMID- 10514120 TI - Contribution of major histocompatibility complex genes to susceptibility and resistance in Helicobacter pylori related diseases. AB - BACKGROUND/AIM: Although there have been numerous reports concerning the virulence factors of isolates for investigating the pathogenesis of Helicobacter pylori infection, few studies have been carried out regarding the association of HLA class II genes of the host with H. pylori related diseases. Two published studies have only analysed the HLA DQ locus alone. The aim of this study was thus to determine the association of HLA class II genes (DR, DQ and DP) with H. pylori related diseases using the DNA typing method. METHODS: Fifty-eight patients with H. pylori positive gastric ulcers, 44 patients with H. pylori positive duodenal ulcers, 45 patients with H. pylori positive gastritis and 34 healthy subjects without H. pylori infection were typed for HLA class II genes by means of DNA typing with the polymerase chain reaction-sequence specific oligonucleotide probes method. RESULTS: A negative association with DRB1*1501, DQA1*01021 and DQB1*0602 alleles was noted in all three of the patient groups studied. Compared with the healthy controls, a positive association with DPA1*0201 (P= 0.032) and DPB1*0901 (P=0.005) in gastric ulcers, a positive association with DRB1*0405 (P=0.022) and DQB1*0401 (P=0.044) in duodenal ulcers, and a positive association with DPB1*0901 (P=0.016) in gastritis were observed. A haplotype analysis showed that the association of alleles with H. pylori related disease was with the haplotype rather than with either of the alleles individually. After correction for multiple comparisons, all the significant associations obtained between H. pylori related diseases and HLA class II genes disappeared. CONCLUSIONS: The interplay between host immunogenetic factors, bacterial virulence factors and environmental conditions may thus play a more important role in the outcome of H. pylori infection than immunogenetic factors alone. PMID- 10514121 TI - Does fear of serious disease predict consulting behaviour amongst patients with dyspepsia in general practice? AB - BACKGROUND: There have been relatively few studies of health care utilization amongst patients with upper gastrointestinal (GI) complaints. We postulated that health care utilization amongst patients with dyspepsia is primarily driven by fear of serious disease. METHOD: Consecutive patients presenting to primary care with dyspepsia were questioned about their health care utilization over a 12 month pre-consultation period. The patients completed a questionnaire which included validated measures of GI symptoms (including symptom frequency, duration and severity), and the shortened neuroticism scale of the Eysenck Personality Questionnaire. In total, 614 patients were recruited into the study, and 596 patients provided details of their health care utilization. Previous health care utilization was defined as one or more primary care visits for upper GI symptoms in the 12 months prior to the index visit; frequent health care utilization was defined as six or more visits over the same period. RESULTS: Previous health care utilization was reported by 80% of patients, while frequent health care utilization was reported by 26% of patients. Fear of serious illness and fear of cancer were univariately associated with previous and frequent health care utilization (both P = 0.001). However, the only independent predictors of previous health care utilization were frequent dyspepsia (odds ratio (OR) = 2.17), pain-related anxiety (OR = 2.08-4.66) and higher neuroticism scores (OR = 1.12); independent predictors of frequent health care utilization were frequent dyspepsia (OR = 3.25), pain-related anxiety (OR = 1.74-6.08), female gender (OR = 1.73) and being a non-drinker (OR = 1.72). Health care utilization was not independently associated with symptom severity or duration, or with patients' characteristics, such as age, marital status, ethnicity, smoking status or the use of non-steroidal antiinflammatory drugs. CONCLUSIONS: Consulting behaviour amongst patients with dyspepsia is driven in part by psychological factors and, in particular, by symptom-related anxiety as well as by the frequency of dyspepsia, but not primarily by fear of serious disease. Anxiety may help sustain health care utilization once the behaviour has been established. PMID- 10514122 TI - VacA seropositivity is not associated with the development of gastric cancer in a Japanese population. AB - OBJECTIVES: Infection with Helicobacter pylori strains producing vacuolating cytotoxin (VacA) is associated with enhanced gastric mucosal damage and the development of gastric mucosal atrophy. The aim of this study was to investigate whether VacA seropositivity is associated with increasing risk of gastric cancer in Japanese populations which have much higher incidence of gastric cancer than Western populations. METHODS: Serum sample was collected from 81 patients with gastric cancer and 81 sex- and age-matched endoscopically evaluated control subjects. Histologically, 62 cancers were of the intestinal type and 76 were early gastric cancer. H. pylori and VacA IgG antibodies were assayed by Western blotting using Chiron Diagnostics' Recombinant Immunoblot Assay (RIBAa). RESULTS: VacA seropositivity was 68% (55/81) in patients with gastric cancer and 70% (57/81) in controls. The odds ratio for the risk of gastric cancer in VacA seropositives was 0.89 (95% CI 0.46-1.74). In H. pylori seropositive patients and their control subjects (matched H. pylori-positive controls), VacA seropositivity was the same 80.6% (50/62). The odds ratio for the risk of gastric cancer in H. pylori-positive patients if VacA seropositive was 1.00 (95% CI 0.41-2.44). The mean relative intensity of VacA antibody titre was 3.01+/-0.18 in the 55 VacA seropositive cancer patients and 3.09+/-0.17 in the 57 VacA seropositive control subjects (difference not significant). CONCLUSION: These results suggest that VacA seropositivity is not associated with increasing risk of gastric cancer in Japanese populations. PMID- 10514123 TI - Extensive aetiological investigations in acute pancreatitis: results of a 1-year prospective study. AB - BACKGROUND: Epidemiological data on acute pancreatitis are poorly defined. AIMS: To prospectively evaluate the aetiology of acute pancreatitis and to assess the benefits of intensive investigations. METHODS: In a prospective, 1-year study all cases of acute pancreatitis in the Nice catchment area were enrolled. Subjects underwent routine (serum calcium, phosphate and triglycerides; abdominal ultrasonography and CT scan) and additional, delayed intensive investigations (ERCP with bile sampling and/or endoscopy ultrasonography). RESULTS: One hundred and twenty-one cases were included. After routine investigations, a biliary, alcoholic, miscellaneous or unknown origin was diagnosed in 43%, 31.4%, 9.9% and 15.7%, respectively. In subjects with biliary pancreatitis, 43% had no previous history of biliary disease. In the alcohol-related subgroup, pancreatitis recurred in 18.5% during 114.5 days mean follow-up. In subjects with a first episode of alcoholic pancreatitis, delayed supplemental investigations revealed underlying chronic pancreatitis in 92.8%. After routine investigations, a diagnosis of pancreatitis of unknown origin was made in 15.7% (n = 19) of subjects. Additional investigations revealed an underlying cause in 57.8% of these patients (n = 11), including malignancy (n = 3) and biliary disease (n = 4), reducing the overall rate of pancreatitis with no apparent cause to 6.6%. CONCLUSIONS: Investigative techniques, particularly ERCP, will reveal the underlying aetiology of pancreatitis in the majority of patients presenting with 'idiopathic' pancreatitis and should be considered when routine tests are negative. PMID- 10514124 TI - The effects of somatostatin and octreotide on the human sphincter of Oddi. AB - OBJECTIVE: Somatostatin acts at different sites in the human gastrointestinal tract and generally inhibits the release and effects of many gastrointestinal hormones and neuropeptides. Together with its long-acting analogue octreotide, somatostatin is widely used in the treatment of hormone-producing tumours, variceal bleeding, etc., but multi-centre trials have failed to prove a beneficial effect in the treatment of acute pancreatitis or in the prevention of post-ERCP pancreatitis (pancreatitis following endoscopic retrograde cholangiopancreatography). The aim of the present work was to study the effects of somatostatin and octreotide on the human sphincter of Oddi by means of quantitative hepatobiliary scintigraphy (QHBS). METHOD: Fifteen cholecystectomized patients were enrolled in the study, six in the somatostatin group and nine in the octreotide group. QHBS was performed initially with a standard protocol (baseline data), then repeated after 0.1 mg octreotide or a 250 microg bolus + 250 microg/h somatostatin administration. In the 60th min of QHBS, 0.5 mg glyceryl trinitrate (GTN) was administered sublingually. RESULTS: QHBS demonstrated that both somatostatin and octreotide caused a marked impairment in the bile flow: the half-time of excretion (T1/2) over the common bile duct was significantly prolonged compared with baseline data (somatostatin group: common bile duct T1/2 180 min versus 59.7+/-31 min; octreotide group: common bile duct T1/2 140.9+/-60.5 min versus 30.7+/-11.7 min). Glyceryl trinitrate administration accelerated the transpapillary bile flow, with significant decreases in the elevated T1/2 in both groups. CONCLUSION: Increased transpapillary flow induced by glyceryl trinitrate may be beneficial in the treatment of acute or post-ERCP pancreatitis. PMID- 10514125 TI - Ornithine decarboxylase in colonic mucosa from patients with moderate or severe Crohn's disease and ulcerative colitis. AB - Polyamines, as well as ornithine decarboxylase (ODC), the enzyme involved in their synthesis, were reported to be closely related to cell proliferation. In Crohn's disease and ulcerative colitis, cell destruction and proliferation increase in the active stage. The aim of the present study was to determine the ODC in both involved and uninvolved areas of the colonic mucosa of active Crohn's disease and ulcerative colitis patients. The patients were divided in two groups, owing to the different level of activity (severe or moderate), by means of clinical endoscopy, laboratory, and histology evaluations. Subjects with suspected disease, but endoscopically unconfirmed, were used as controls. In all ulcerative colitis and Crohn's disease patients the ODC values both in involved and uninvolved mucosa were significantly lower than in controls. In severe Crohn's disease ODC was significantly reduced versus moderate Crohn's disease only in affected tissues. In all ulcerative colitis patients (moderate or severe) the ODC was significantly decreased in involved mucosa compared with uninvolved mucosa. Severe ulcerative colitis showed the significantly lowest ODC. We suggest that the significant decrease of ODC in the bowel mucosa is closely related to the severity of the disease. The highest decrease of ODC in ulcerative colitis patients would be due both to the enhanced cell destruction, and to the feed-back from exogenous increased polyamine production (bowel bacteria, cell desquamation). Therefore ODC would be considered a sensitive index of the inflammatory derangement of the mucosa, especially in acute ulcerative colitis. We conclude that this behaviour may result in an enhanced risk of neoplasia. PMID- 10514126 TI - Oral microemulsion cyclosporin to reduce steroids rapidly in chronic active ulcerative colitis. AB - OBJECTIVE: The list of the therapeutic tools for chronic active ulcerative colitis lacks a potent and reliably absorbed immune modifier to help wean patients from steroids. We investigated whether oral microemulsion cyclosporin (Neoral) can achieve this goal. DESIGN: Retrospective analysis. SETTING: Tertiary care referral centre. PARTICIPANTS: Nine consecutive patients facing colectomy because of steroid-dependent chronic active ulcerative colitis were studied: the disease was left-sided in six of them and had lasted a median of 10 years. Two patients depended on prednisone 12.5 mg/day, six on 25 mg, and one on 35 mg. The median cumulative steroid dose in the month preceding the trial had been 750 mg. INTERVENTIONS: Neoral was started at a dose of 5 mg/kg/day, which was then titrated to reach a whole-blood therapeutic range of 60-240 ng/ml. During the three-month trial, previous drugs were continued but steroids were tapered according to the patient's clinical condition. PRIMARY OUTCOME MEASURES: The number of patients leaving the trial with quiescent disease and a significantly decreased need for steroids. RESULTS: Eight of the nine patients (89%) showed an initial response and commenced tapering steroids. Remission to the end of the 3rd month was observed in five cases treated with < or = 15 mg steroids daily (median cumulative dose 150 mg). Of the remaining four cases, two were partial responders and two failures, with the failure being linked to poor drug absorption in at least one patient. CONCLUSION: Neoral may help wean patients with chronic active ulcerative colitis from steroids, but this novel indication of cyclosporin must be tested in a specifically designed study. PMID- 10514127 TI - Frequency and clinical evolution of indeterminate colitis: a retrospective multi centre study in northern Italy. GSMII (Gruppo di Studio per le Malattie Infiammatorie Intestinali). AB - OBJECTIVE: To evaluate the prevalence and the clinical evolution of patients with an initial diagnosis of indeterminate colitis. DESIGN: Retrospective, observational study. SETTING: Fifteen gastrointestinal units in northern Italy. PARTICIPANTS: Patients with an initial diagnosis of indeterminate colitis seen between 1988 and 1993. INTERVENTIONS: Patients were traced through a common database and centres were requested to update their clinical follow-up. MAIN OUTCOME MEASURES: Frequency of patients with an initial diagnosis of indeterminate colitis among those with IBD; rate of patients who subsequently had a definite diagnosis of either Crohn's disease or ulcerative colitis. RESULTS: Fifty out of 1113 IBD patients (4.6%) had been diagnosed as having indeterminate colitis. During follow-up, 37 patients (72.5%) had a definite diagnosis of either Crohn's disease or ulcerative colitis. The cumulative probability of having a definite diagnosis of either ulcerative colitis or Crohn's disease was 80% 8 years after the first one (i.e. the first diagnosis). The probability of having a diagnosis of Crohn's disease was increased in patients with fever at onset, segmental endoscopic lesions or extra-intestinal complications and in current smokers. The probability of having a diagnosis of ulcerative colitis was increased in patients who had not undergone appendectomy before diagnosis. CONCLUSIONS: In our area, indeterminate colitis accounts for about 5% of initial diagnoses of IBD. In about 80% of patients, a diagnosis of either ulcerative colitis or Crohn's disease is made within 8 years. Several clinical and demographic features can help in identifying those patients more likely to have a subsequent diagnosis of Crohn's disease and those more likely to have a subsequent diagnosis of ulcerative colitis. PMID- 10514128 TI - Hepatocellular carcinoma arising in the absence of cirrhosis in genetic haemochromatosis: three case reports and review of literature. AB - Genetic haemochromatosis constitutes a high risk factor for the development of hepatocellular carcinoma. It is widely accepted that venesection prevents the evolution of cirrhosis in haemochromatosis and indirectly protects against the development of hepatocellular carcinoma. Clinical, pathological and radiological data are presented on three patients who did not conform to the 'siderosis cirrhosis-carcinoma' sequence and in whom prompt and adequate iron depletion did not prevent the development of cancer. This is the first report of hepatocellular carcinoma intervening in non-cirrhotic liver in two siblings with genetic haemochromatosis. The current literature on the subject is reviewed. The direct oncogenic role of iron remains to be elucidated. Hepatocellular carcinoma should be considered as a differential diagnosis in patients with non-cirrhotic genetic haemochromatosis who present with clinical deterioration during the course of an otherwise uneventful venesection programme. PMID- 10514129 TI - Hepatomegaly and cholestasis as primary clinical manifestations of an AL-kappa amyloidosis. AB - A 53-year-old man, who presented with weight loss over a period of 10 months, hepatomegaly, markedly raised cholestatic enzymes and Ca 19-9, was initially suspected of suffering from metastatic cholangio-carcinoma. Liver biopsy revealed depositions of AL-amyloid. Further investigations confirmed a generalized amyloidosis. Biopsies taken from the gastric, colonic, and bronchial mucosa all showed depositions of amyloid. A nephrotic syndrome was interpreted as being secondary to the renal involvement. Echocardiography identified changes which were consistent with cardiac involvement. A plasmacytoma or lymphoma was excluded. At the time of diagnosis the patient was in a good physical condition with normal renal function. Within a few weeks the renal function deteriorated and after 2 months the patient developed ascites and became jaundiced. Four months after initial presentation the patient died from cardiac failure. PMID- 10514130 TI - Increased survival in advanced primary biliary cirrhosis patients with regular albumin infusions? AB - Primary biliary cirrhosis (PBC) is a chronic cholestatic liver disease. Frequently, it can slowly progress to cirrhosis and, finally, death. Liver transplantation is the only way to avoid this endpoint. For patients not accepted for liver transplantation, only symptomatic treatment remains. In advanced PBC, one of the main concerns is a decreasing albumin level. As albumin is an important protein in the human body, we wondered whether regular albumin infusions may result in a higher quality of life and prolong survival. In this case report we describe three patients who survived for a remarkably long time on this treatment regimen before finally succumbing to the sequela of advanced liver disease. Some features of albumin are mentioned which may help explain this phenomenon. We conclude that regular albumin infusions may be a means of decreasing morbidity and prolonging survival in a subgroup of patients with advanced PBC. However, prospective evaluation of regular albumin infusions in a group of PBC patients (e.g. those rejected for liver transplantation) seems mandatory. PMID- 10514131 TI - Acute self-limiting jejunitis as the first manifestation of microscopic polyangiitis associated with Sjogren's disease: report of one case and review of the literature. AB - We report a case of acute self-limiting ulcerative jejunitis of unknown aetiology in a 72-year-old female patient in which a subsequent diagnosis of microscopic polyangiitis and Sjogren's syndrome was made. All known causes of jejunal ulceration and inflammation were excluded. Previously reported cases of acute self-limiting jejunitis are reviewed and the possibility that acute jejunitis in this patient had been the first manifestation of systemic vasculitis is discussed. PMID- 10514132 TI - Intestinal capillariasis acquired in Egypt. AB - A 34-year-old Egyptian man presented with a 4-month history of profound weight loss, diarrhoea and abdominal pain. Extensive investigations in Egypt had failed to provide a diagnosis but subsequent stool examinations revealed ova of Capillaria philippinensis. The patient made a slow but complete recovery after treatment with albendazole. The literature on intestinal capillariasis is reviewed. PMID- 10514133 TI - Phlebotomy reduces serum procollagen III peptide levels in patients with chronic hepatitis C. PMID- 10514134 TI - Combined coeliac disease and thyroid disease. PMID- 10514135 TI - Pathogenesis and medical therapy of primary sclerosing cholangitis. PMID- 10514136 TI - Contrasting views on human population growth. One wisdom justifies complacency: the other demands action now. PMID- 10514137 TI - Impediments to effective fertility reduction. Contraception should be moved out of the hands of doctors. PMID- 10514138 TI - The population policy pendulum. Needs to settle near the middle--and acknowledge the importance of numbers. PMID- 10514139 TI - Failure of an intervention to stop teenagers smoking. Not such a disappointment as it appears. PMID- 10514140 TI - Fertility after treatment for cancer. Questions remain over ways of preserving ovarian and testicular tissue. PMID- 10514141 TI - Stumbling into rationing. A national debate on values is needed to sustain the NHS. PMID- 10514142 TI - NICE to rule on influenza flu drug zanamivir. PMID- 10514143 TI - Japan's worst nuclear accident leaves two fighting for life. PMID- 10514144 TI - London professor struck off for bullying and dishonesty. PMID- 10514146 TI - In brief PMID- 10514145 TI - GP faces 15 charges of murder. PMID- 10514148 TI - Scientists who do not publish trial results are "unethical" PMID- 10514147 TI - CSM takes small step towards openness PMID- 10514149 TI - England to have more walk-in clinics PMID- 10514150 TI - Mental health services framework welcomed PMID- 10514151 TI - Cooperation could provide better hospital care. PMID- 10514152 TI - Kosovo to shift emphasis to a primary care system. PMID- 10514153 TI - Eight die in outbreak of virus spread from birds. PMID- 10514154 TI - King in a maverick style. PMID- 10514155 TI - Behavioural counselling in general practice for the promotion of healthy behaviour among adults at increased risk of coronary heart disease: randomised trial. AB - OBJECTIVE: To measure the effect of behaviourally oriented counselling in general practice on healthy behaviour and biological risk factors in patients at increased risk of coronary heart disease. DESIGN: Cluster randomised controlled trial. PARTICIPANTS: 883 men and women selected for the presence of one or more modifiable risk factors: regular cigarette smoking, high serum cholesterol concentration (6.5-9.0 mmol/l), and high body mass index (25-35) combined with low physical activity. INTERVENTION: Brief behavioural counselling, on the basis of the stage of change model, carried out by practice nurses to reduce smoking and dietary fat intake and to increase regular physical activity. MAIN OUTCOME MEASURES: Questionnaire measures of diet, exercise, and smoking habits, and blood pressure, serum total cholesterol concentration, weight, body mass index, and smoking cessation (with biochemical validation) at 4 and 12 months. RESULTS: Favourable differences were recorded in the intervention group for dietary fat intake, regular exercise, and cigarettes smoked per day at 4 and 12 months. Systolic blood pressure was reduced to a greater extent in the intervention group at 4 but not at 12 months. No differences were found between groups in changes in total serum cholesterol concentration, weight, body mass index, diastolic pressure, or smoking cessation. CONCLUSIONS: Brief behavioural counselling by practice nurses led to improvements in healthy behaviour. More extended counselling to help patients sustain and build on behaviour changes may be required before differences in biological risk factors emerge. PMID- 10514156 TI - Cluster randomised controlled trial of expert system based on the transtheoretical ("stages of change") model for smoking prevention and cessation in schools. AB - OBJECTIVES: To examine whether a year long programme based on the transtheoretical model of behaviour change, incorporating three sessions using an expert system computer program and three class lessons, could reduce the prevalence of teenage smoking. DESIGN: Cluster randomised trial comparing the intervention to a control group exposed only to health education as part of the English national curriculum. SETTING: 52 schools in the West Midlands region. PARTICIPANTS: 8352 students in year 9 (age 13-14 years) at those schools. MAIN OUTCOME MEASURES: Prevalence of teenage smoking 12 months after the start of the intervention. RESULTS: Of the 8352 students recruited, 7444 (89.1%) were followed up at 12 months. The intention to treat odds ratio for smoking in the intervention group relative to control was 1.08 (95% confidence interval 0.89 to 1.33). Sensitivity analysis for loss to follow up and adjustment for potential confounders did not alter these findings. CONCLUSIONS: The smoking prevention and cessation intervention based on the transtheoretical model, as delivered in this trial, is ineffective in schoolchildren aged 13-14. PMID- 10514158 TI - Effect of station design on death in the London underground: observational study. PMID- 10514157 TI - Relation between income inequality and mortality: empirical demonstration. AB - OBJECTIVE: To assess the extent to which observed associations at population level between income inequality and mortality are statistical artefacts. DESIGN: Indirect "what if" simulation by using observed risks of mortality at individual level as a function of income to construct hypothetical state level mortality specific for age and sex as if the statistical artefact argument were 100% correct. SETTING: Data from the 1990 census for the 50 US states plus Washington, DC, were used for population distributions by age, sex, state, and income range; data disaggregated by age, sex, and state from the Centers for Disease Control and Prevention were used for mortality; and regressions from the national longitudinal mortality study were used for the individual level relation between income and risk of mortality. RESULTS: Hypothetical mortality, while correlated with inequality (as implied by the logic of the statistical artefact argument), showed a weaker association with states' levels of income inequality than the observed mortality. CONCLUSIONS: The observed associations in the United States at the state level between income inequality and mortality cannot be entirely or substantially explained as statistical artefacts of an underlying individual level relation between income and mortality. There remains an important association between income inequality and mortality at state level over and above anything that could be accounted for by any statistical artefact. This result reinforces the need to consider a broad range of factors, including the social milieu, as fundamental determinants of health. PMID- 10514160 TI - Pragmatic randomised controlled trial of local corticosteroid injection and naproxen for treatment of lateral epicondylitis of elbow in primary care. AB - OBJECTIVE: To compare the clinical effectiveness of local corticosteroid injection, standard non-steroidal anti-inflammatory drugs, and simple analgesics for the early treatment of lateral epicondylitis in primary care. DESIGN: Multicentre pragmatic randomised controlled trial. SETTING: 23 general practices in North Staffordshire and South Cheshire. PARTICIPANTS: 164 patients aged 18-70 years presenting with a new episode of lateral epicondylitis. INTERVENTIONS: Local injection of 20 mg methylprednisolone plus lignocaine, naproxen 500 mg twice daily for two weeks, or placebo tablets. All participants received a standard advice sheet and co-codamol as required. MAIN OUTCOME MEASURES: Participants' global assessment of improvement (five point scale) at four weeks. Pain, function, and "main complaint" measured on 10 point Likert scales at 4 weeks, 6 months, and 12 months. RESULTS: Over 2 years, 53 subjects were randomised to injection, 53 to naproxen, and 58 to placebo. Prognostic variables were similar between groups at baseline. At 4 weeks, 48 patients (92%) in the injection group were completely better or improved compared with 30 (57%) in the naproxen group (P<0.001) and 28 (50%) in the placebo group (P<0.001). At 12 months, 43 patients (84%) in the injection group had pain scores 0.05). CONCLUSIONS: Early local corticosteroid injection is effective for lateral epicondylitis. Outcome at one year was good in all groups, and effective early treatment does not seem to influence this. PMID- 10514159 TI - Primary prevention of arterial thromboembolism in non-rheumatic atrial fibrillation in primary care: randomised controlled trial comparing two intensities of coumarin with aspirin. AB - OBJECTIVE: To investigate the effectiveness of aspirin and coumarin in preventing thromboembolism in patients with non-rheumatic atrial fibrillation in general practice. DESIGN: Randomised controlled trial. PARTICIPANTS: 729 patients aged >/=60 years with atrial fibrillation, recruited in general practice, who had no established indication for coumarin. Mean age was 75 years and mean follow up 2. 7 years. SETTING: Primary care in the Netherlands. INTERVENTIONS: Patients eligible for standard intensity coumarin (international normalised ratio 2.5-3.5) were randomly assigned to standard anticoagulation, very low intensity coumarin (international normalised ratio 1.1-1.6), or aspirin (150 mg/day) (stratum 1). Patients ineligible for standard anticoagulation were randomly assigned to low anticoagulation or aspirin (stratum 2). MAIN OUTCOME MEASURES: Stroke, systemic embolism, major haemorrhage, and vascular death. RESULTS: 108 primary events occurred (annual event rate 5.5%), including 13 major haemorrhages (0.7% a year). The hazard ratio was 0.91 (0.61 to 1.36) for low anticoagulation versus aspirin and 0.78 (0.34 to 1.81) for standard anticoagulation versus aspirin. Non-vascular death was less common in the low anticoagulation group than in the aspirin group (0.41, 0.20 to 0.82). There was no significant difference between the treatment groups in bleeding incidence. High systolic and low diastolic blood pressure and age were independent prognostic factors. CONCLUSION: In a general practice population (without established indications for coumarin) neither low nor standard intensity anticoagulation is better than aspirin in preventing primary outcome events. Aspirin may therefore be the first choice in patients with atrial fibrillation in general practice. PMID- 10514162 TI - That's show business PMID- 10514161 TI - Science, medicine, and the future. Contraception. PMID- 10514163 TI - ABC of complementary medicine. Acupuncture. PMID- 10514164 TI - Intractable hiccups PMID- 10514165 TI - Human numbers, environment, sustainability, and health. PMID- 10514166 TI - World population and health in transition. PMID- 10514167 TI - Paths to lower fertility. PMID- 10514168 TI - Prospects for feeding the world. PMID- 10514169 TI - China's one child family policy. PMID- 10514170 TI - India: looking ahead to one and a half billion people. PMID- 10514171 TI - The US Department of State is policing the population policy lockstep. PMID- 10514172 TI - The need to know PMID- 10514173 TI - The plague PMID- 10514174 TI - Reasons for not seeing drug representatives. They should be seen because they are a good resource. PMID- 10514175 TI - Postcodes don't indicate individuals' social class. PMID- 10514176 TI - Coronary artery disease varies seasonably in subtropics. PMID- 10514177 TI - Acute urinary retention in men. Management is more complex issue than was described. PMID- 10514178 TI - DEC methods for appraising new drugs. Are a foundation for the nice appraisal committee. PMID- 10514179 TI - Drugs arriving in Kosovo need checking. PMID- 10514181 TI - Data on babies' safety during hospital births are being ignored. PMID- 10514183 TI - Obituaries PMID- 10514180 TI - Rates of anxiety and depression in African-Caribbeans may not reflect reality. PMID- 10514184 TI - Doctors need 5% pay rise just to stand still PMID- 10514182 TI - Midwives would prefer a vaginal delivery. PMID- 10514186 TI - How the idea of profession changed the writing of medical history PMID- 10514185 TI - The baby makers PMID- 10514187 TI - Curing the incurable PMID- 10514188 TI - Demographic entrapment PMID- 10514189 TI - Both sides PMID- 10514191 TI - Benign uproar PMID- 10514190 TI - Oracles PMID- 10514192 TI - Behavioural counselling in primary care leads to improvements in health behaviour PMID- 10514193 TI - Schools smoking prevention programme based on "stages of change" model does not work PMID- 10514195 TI - Tube trains kill fewer people when there's a pit under the rails PMID- 10514194 TI - Observed association between income inequality and mortality is not a statistical artefact PMID- 10514197 TI - Steroid injection gives best relief for tennis elbow PMID- 10514196 TI - Aspirin is best for mild atrial fibrillation in primary care PMID- 10514198 TI - Popliteal artery entrapment syndrome: more common than previously recognized. AB - PURPOSE: This report summarizes our experience with the popliteal entrapment syndrome in 88 limbs (48 patients) treated during a 10-year period. METHOD: The study cohort consisted of a retrospective analysis of those patients who were seen with symptoms of claudication or severe ischemia by a single surgical group and in whom unequivocal evidence of popliteal entrapment was shown either with angiography or at the time of operation. The cases were collected prospectively in a private vascular surgical practice. RESULTS: Bilateral popliteal entrapment was found in 40 of the 48 patients. The mean age at the time of presentation was 35.0 years (SD, 11.6 years). Claudication was the most frequent presenting symptom (70 of 88 limbs). Types I, II, III, and IV popliteal entrapment were found in 58 limbs (15 arteries occluded), and 30 limbs (three occlusions) were seen with a "functional" popliteal artery entrapment (apparent absence of a developmental anatomic abnormality). Of the 18 limbs with severe ischemia and associated occlusion of the popliteal artery, 15 underwent bypass grafting with reversed saphenous vein grafts, all of which remained patent during the follow-up period (median follow-up, 4.2 years; range, 1 to 10 years). One popliteal artery occlusion that was treated with thrombectomy and vein patching occluded within 6 months and necessitated subsequent vein grafting. Two limbs with inoperable occluded popliteal arteries were not subjected to reconstruction (one necessitated amputation because of advanced ischemia, and the second had extensive thrombosis of the distal run-off). In two patients (four limbs), moderate presenting symptoms abated without surgery after the discontinuation of an extreme exercise program. The remaining limbs underwent surgical decompression (all popliteal arteries remained patent, with a median follow-up of 3.9 years). CONCLUSION: The popliteal entrapment syndrome is more prevalent than has formerly been appreciated. On the basis of observations made in this series and in the surgical literature, we advise surgical correction in all cases of types I, II, III, and IV entrapment at the time of diagnosis to avoid occlusion as a result of continued arterial wall degeneration. In contrast, decompression is only advised in those patients with "functional entrapment" if they have discrete and typical symptoms because up to 50% of the normal population may display transient popliteal artery compression with extremes of plantar flexion or dorsiflexion. On the basis of the severe histologic changes found in those popliteal arteries that had undergone occlusion at the time of presentation, it is advised that the popliteal artery should be completely replaced, ideally with a vein graft, when significant degeneration or occlusion of the popliteal artery is noted at the time of operation. PMID- 10514199 TI - The value of acetazolamide single photon emission computed tomography scans in the preoperative evaluation of asymptomatic critical carotid stenosis. AB - PURPOSE: Acetazolamide (ACZ)-enhanced single photon emission computed tomography (SPECT) scans can assess both cerebral perfusion and vascular reactivity. Patients with asymptomatic critical carotid artery stenosis were evaluated for cerebral vascular reactivity to determine the effect of extracranial occlusive disease and the effect of carotid endarterectomy (CEA) on intracerebral reactivity. METHODS: In 44 patients with asymptomatic critical carotid artery stenosis, cerebral perfusion and vascular reactivity were assessed before CEA with resting and ACZ-enhanced SPECT scans. All patients had a 70% or greater ipsilateral internal carotid artery stenosis. Preoperative ACZ-enhanced SPECT scans were obtained, usually 5 days before CEA. Postoperative ACZ-enhanced SPECT scans were obtained in 30 patients. RESULTS: Preoperative SPECT scans were asymmetric, revealing focal (n = 19) or global (n = 15) decreased reactivity in 34 patients (77%). Ten patients had symmetric or normal reactivity. After CEA, 23 patients demonstrated an improvement in reactivity ipsilateral to the side of surgery. The remaining seven patients failed to improve after surgery. CONCLUSION: Although all patients had a high-grade internal carotid stenosis, nearly a quarter of the patients had excellent intracerebral collateral flow. Only 71% of patients demonstrated improved intracerebral vasoreactivity after CEA. The lack of improvement in the other patients may have resulted from intracerebral pathology or lack of improvement in the extracranial carotid hemodynamics. PMID- 10514200 TI - Refining the indications for carotid endarterectomy in patients with symptomatic carotid stenosis: A systematic review. AB - OBJECTIVE: The purpose of this study was to summarize the existing literature on the efficacy of carotid endarterectomy in patients with ipsilateral symptomatic carotid stenosis. METHODS: Database searching, relevance assessment, methodologic quality assessments, and data extraction were all performed in duplicate with prespecified criteria. RESULTS: Twenty-three publications were identified from the North American Symptomatic Carotid Endarterectomy Trial, the European Carotid Surgery Trial, and the Veterans Affairs Cooperative Studies Program. Stenosis was reported as measured in the North American Symptomatic Carotid Endarterectomy Trial. In patients with >70% stenosis, carotid endarterectomy was associated with a pooled relative risk reduction of 48% (95% confidence interval [CI], 27% to 73%) and an absolute risk reduction of 6.7% (95% CI, 3.2% to 10%) for the outcome of death or major disability from stroke. This translates into a number needed to treat of 15 (95% CI, 10 to 31). For patients with 50% to 69% stenosis, the benefit of surgery was less and the confidence intervals were wider. A relative risk reduction of 27% (95% CI, 5% to 44%), an absolute risk reduction of 4.7% (95% CI, 0.8% to 8.7%), and a number needed to treat of 21 (95% CI, 11 to 125) were observed in this group. The patients with the lowest degrees of stenosis (<50%) were harmed by the intervention (number needed to harm, 45). Increasing degree of stenosis, increasing age, male sex, the presence of other medical risk factors, and the presence of hemispheric rather than retinal antecedent events were factors that increased the benefits from surgery. CONCLUSION: Carotid endarterectomy reduced death or major disability from stroke in patients with >50% symptomatic stenosis. To maximize the benefits of surgery, careful preoperative risk assessment and the maintenance of low rates of major perioperative complications are mandatory. PMID- 10514201 TI - Perioperative risk and late outcome of nonelective carotid endarterectomy. AB - PURPOSE: In an earlier report of our database for 1924 isolated carotid endarterectomies (CEAs) from 1989 to 1995, multivariable analysis results indicated that the urgency of operation unfavorably influenced the combined stroke and mortality rate (CSM). This study was conducted in an attempt to document the features that contribute to perioperative complications and late outcome in 314 patients for whom CEA was considered to be nonelective because of the severity of previous symptoms, carotid stenosis, or medical comorbidities. METHODS: All the hospital charts and outpatient records were reviewed retrospectively for the 209 men and 105 women who had undergone nonelective CEAs (median age, 69 years). Information regarding the clinical risk factors, the operative indications (CHAT classification), the severity and distribution of carotid disease, and the surgical management were analyzed to assess the impact on the 30-day CSM and on the long-term survival rate and neurologic events during a median follow-up period of 34 months. RESULTS: Previous symptoms had occurred in 285 patients (91%) and included cortical transient ischemic attacks in 47%, amaurosis fugax in 20%, completed strokes in 14%, unstable strokes in 2%, and nonspecific or miscellaneous symptoms in 8%. Preoperative angiography was performed in 308 patients (98%), which confirmed the presence of 80% to 99% ipsilateral carotid stenosis in 79% of the patients and >90% stenosis in 43%. The median interval between presentation and surgical treatment was 2 days, but 48% of the 314 CEAs were performed within 24 hours of presentation. The 30-day CSM was 6.7% and ranged from 3.4% for 29 patients with severe asymptomatic carotid stenosis to 14% for those patients with unstable strokes. The cardiac and pulmonary risk factors were the only variables that were related statistically to the CSM. During the follow-up period, the risk for ipsilateral stroke was significantly higher in women (risk ratio [RR], 2.38; 95% confidence interval [CI], 1.02 to 5.56; P =.04) and in patients with higher gradients of cardiac and pulmonary risk factors (RR, 2.8; 95% CI, 1.6 to 4.8 per gradient increase; P <.001). The risk was significantly lower in patients who had undergone vein patch angioplasty (RR, 0.29; 95% CI, 0.12 to 0. 71; P =.006) in comparison with synthetic patching. However, 38 of the 55 patients (69%) who underwent synthetic patching also had widespread atherosclerosis for which the saphenous veins already had been harvested for coronary bypass grafting surgery or infrainguinal revascularization. CONCLUSION: In our experience, the perioperative risk of nonelective CEA primarily is determined by incidental cardiopulmonary disease. Vein patch angioplasty appears to enhance late results, but the late stroke rate associated with synthetic patching also may have been influenced by the extent of vascular disease in our study group. PMID- 10514202 TI - The epidemiology of surgically repaired aneurysms in the United States. AB - OBJECTIVE: The purpose of this study was to determine the results of surgery for hospitalized cases of aneurysms in the United States, thereby providing a standard of comparison for new techniques proposed to treat aneurysms. METHODS: Data on hospitalized aneurysm cases were collected from the National Hospital Discharge Survey, a comprehensive database of patients hospitalized in the United States for treatment from the years 1984 to 1994. The National Hospital Discharge Survey samples non-federal, acute-care hospitals with an average length of stay of less than 30 days. All the cases had a diagnosis of or a surgical procedure for a non-cerebral aneurysm. RESULTS: In the year 1994, 51,949 non-cerebral aneurysms were repaired in the United States, and 75% of these procedures were abdominal aortic aneurysm (AAA) surgeries. The operative mortality rates for AAA were higher than previously reported from multi-institutional studies and were found to be 8.4% for elective repair and 68% for emergency AAA repair. The number of aneurysm surgeries per thousand population varied by region: surgery rates were more frequent in the Northeast and less frequent in the West. Surgical volume appeared to decrease for smaller hospitals and increase for larger hospitals for the period between 1990 and 1994. The overall mortality rates for all aneurysm surgeries diminished with hospital size. However, no significant difference was found for the rates of elective AAA repair between hospital sizes. The percentage of men with aneurysms who underwent surgery for repair was significantly higher than for women with aneurysms. In addition, the AAA repair rates increased for men from 1985 to 1994, and the number of women reported with repaired AAAs remained constant. CONCLUSION: The location of aneurysm, urgency of repair, region, sex, and hospital size are important factors related to patient treatment and outcome. These data provide a standard of comparison against which surgeons can compare their own results, and they provide a benchmark for the evaluation of interventional techniques proposed to treat aneurysms. PMID- 10514203 TI - Coagulation and fibrinolysis in patients undergoing operation for ruptured and nonruptured infrarenal abdominal aortic aneurysms. AB - PURPOSE: Hemorrhage and thrombosis predisposing to myocardial infarction, multiple organ failure, and thromboembolism account for the majority of the morbidity and mortality associated with repair of ruptured and nonruptured abdominal aortic aneurysms (AAAs). The aim of this study was to examine coagulation and fibrinolysis in patients operated on for ruptured and nonruptured infrarenal AAAs. METHODS: Ten patients operated on for ruptured and 9 patients operated on for nonruptured AAAs were studied. Tissue plasminogen activator (t PA) antigen, thrombin-antithrombin (TAT), and D-dimer were measured before induction of anesthesia. Plasminogen activator inhibitor (PAI) activity, t-PA activity, and prothrombin fragment (PF) 1+2 were measured before induction of anesthesia, immediately before aortic clamp release, and 5 minutes and 24 hours after aortic clamp release. RESULTS: Preoperatively, ruptured AAA was associated with significantly elevated t-PA antigen (median 15.7 ng/mL, range 9. 0 to 22.1 ng/mL versus nonrupture: median 6.6 ng/mL, range 4.7 to 16. 4 ng/mL; P <.01, Mann Whitney test), increased PAI activity (median 36.5 arbitrary units/mL, range 20.6 to 38.8 arbitrary units/mL versus nonrupture: median 8.2 arbitrary units/mL, range 3.2 to 21.7 arbitrary units/mL; P <.001), reduced t-PA activity (median 0.12 IU/mL, range 0.06 to 0.4 IU/mL versus nonrupture: median 0.49 IU/mL, range 0.14 to 3.2 IU/mL; P <.01), elevated TAT (median 135.5 microg/L, range 61.2 to 209.4 microg/L versus nonrupture: median 21. 6 microg/L, range 6.6 to 180.4 microg/L; P <.02) and elevated PF 1+2 (median 9.0 nmol/L, range 5.4 to 11.6 nmol/L versus nonrupture: median 2.2 nmol/L, range 0.7 to 7.1 nmol/L, P <.001). There was no significant difference in preoperative D-dimer levels (median 3460 ng/mL, range 1236 to 7860 ng/mL versus nonrupture: median 1642 ng/mL, range 728 to 5334 ng/mL; P =.07). The differences in PAI activity, t-PA activity, and PF 1+2 persisted throughout the course of surgery, but there was no significant difference between the groups at 24 hours. CONCLUSION: These novel data demonstrate that ruptured AAA repair is associated with inhibition of systemic fibrinolysis and intense thrombin generation. Similar changes are seen in nonruptured AAA but are of a lesser magnitude. This procoagulant state may contribute to the microvascular and macrovascular thrombosis that leads to myocardial infarction, multiple organ failure, and thromboembolism. PMID- 10514204 TI - Epidural anesthesia reduces length of hospitalization after endoluminal abdominal aortic aneurysm repair. AB - PURPOSE: The low invasiveness of endoluminal abdominal aneurysm repair (EAAR) appears optimal for the use of epidural anesthesia (EA). However, reported series on EAAR show that general anesthesia (GA) is generally preferred. To evaluate the feasibility and problems encountered with EA for EAAR, patients undergoing EAAR with EA and patients undergoing EAAR with GA were examined. METHODS: From April 1997 through October 1998, EAAR was performed on 119 patients at the Unit of Vascular Surgery at Policlinico Monteluce in Perugia, Italy. Four patients (3%) required conversion to open repair and were excluded from the analysis because they were not suitable candidates for evaluating the feasibility of EA. The study cohort thus comprised 115 patients undergoing abdominal aortic aneurysm (AAA) repair with the AneuRx Medtronic stent graft. The incidence of risk factors and anatomical features of the aneurysm were compared in patients selected for EA or GA on the basis of intention-to-treat analysis. Intraoperative and perioperative data were compared and analyzed on the basis of intention-to-treat and on treatment analysis. RESULTS: Sixty-one patients (54%) underwent the surgical procedure with EA (group A), and 54 (46%) underwent the surgical procedure with GA (group B). Conversion from EA to GA was required in four patients (3 of 61 patients, 5%). There were no statistically significant differences between the two study groups in demographics, clinical characteristics, and American Society of Anesthesiology classification (ASA). There was no perioperative mortality. Major morbidity occurred in 3% of patients (group B). According to intention-to treat analysis, no significant differences were observed between the two groups in mean operating time, fluoro time, blood loss, amount of contrast media used, mean units of transfused blood, need of intensive care unit, mean postoperative hospital stay, and postoperative endoleak. Conversely, significant differences were found by means of on-treatment analysis in the need of intensive care unit (0 vs 5 patients; P =.02), and length of hospitalization (2.5 vs 3.2 days; P =.04). Multivariate logistic regression analysis showed that GA and ASA 4 were positive independent predictors of prolonged (more than 2 days) postoperative hospitalization (hazard ratio, 2.5; 95% CI, 1.1 to 5.8; P =.03, and hazard ratio, 5.1; 95% CI, 1.5 to 17.9; P =.007, respectively). CONCLUSION: EA for EAAR is feasible in a high percentage of patients in whom it is attempted, and it ensures a technical outcome comparable with that of patients undergoing EAAR with GA. Successful completion of EAAR with EA is associated with a short period of hospitalization. PMID- 10514205 TI - Stent attachment site-related endoleakage after stent graft treatment: An in vitro study of the effects of graft size, stent type, and atherosclerotic wall changes. AB - OBJECTIVE: Perigraft endoleakage is a major complication of the endovascular treatment of abdominal aortic aneurysms. The factors that cause this form of endoleakage are not completely identified. The effect of sizing of the prosthesis in combination with either self-expandable or balloon-expandable stents is evaluated in this study. Further, the influence of atherosclerotic changes on endoleakage is evaluated. METHODS: Eight human abdominal aortas were assessed macroscopically at 11 sites for the presence of atherosclerotic changes with intravascular ultrasound scanning (IVUS) and with computed tomography (CT). Five aortas were placed in in vitro circulation with physiologic parameters. After the determination of the proximal and distal landing site of the stent graft, the diameter and surface measurements of the cross sections were taken. The stent graft diameters were chosen from 4-mm undersizing to 6-mm oversizing, both for Gianturco stent grafts (William Cook Europe A/S, Bjaeverskov, Denmark) and for Palmaz stent grafts (Cordis/Johnson & Johnston Co, Warren, NJ). After placement of the stent graft, the diameter and surface measurements of the aortic cross section were determined at the proximal and distal stent attachment sites. The presence and size of the folds at the stent attachment site and the interface with the aortic wall were determined with IVUS and angioscopy. Endoleakage was evaluated with angiography. After angioplasty of the stent attachment site, IVUS, angioscopy, and angiography were repeated. RESULTS: Regarding atherosclerotic changes of the aortic wall, the correlations between clinical impression and CT, clinical impression and IVUS, and CT and IVUS were high (r = 0.77, r = 0.79 and r = 0.79, respectively). For the Gianturco stent grafts, no significant relationship existed between the diameters measured before and after stent graft placement, leading to great differences in intended and achieved oversizing. The achieved oversizing was less in the case of minimal atherosclerotic changes of the aortic wall. The Gianturco stent graft followed the aortic wall closely during the heart cycle. The sizes of the folds of the fabric were clearly correlated with the achieved oversizing (r = 0.83; P =.04) and the grade of endoleakage (r = 0.88; P =.022). Angioplasty after stent graft placement had no effect on the diameter and the grade of endoleakage. Palmaz stent grafts did not follow the aortic wall during the heart cycle. A significant correlation existed between oversizing and both space between aortic wall and stent graft (r = -0.88; P =.02) and grade of endoleakage (r = 0.84; P =.036). Grade of endoleakage in the Palmaz stent graft group was less than in the Gianturco stent graft group. CONCLUSION: With the use of Gianturco stents, a great difference between intended and achieved oversizing is accomplished. The atherosclerotic changes of the aortic wall possibly affect this finding. The configuration of the Gianturco stent results in the formation of fold in the case of oversizing, which is associated with endoleakage. However, the self-expandable character of the stent leads to a close relation to the aortic wall during the heart cycle, and this may possibly accommodate future aortic neck dilation. The Palmaz stent grafts do not follow the aortic wall during the heart cycle, but they do lead to better interface between the graft and the aortic wall, which results in less endoleakage. PMID- 10514206 TI - Costing vascular surgery: A review of current reporting practice. AB - OBJECTIVE: The objective of this study was to assess the level of reporting in economic studies in the area of peripheral vascular disease. Adequate reporting of data is necessary to judge the quality of economic studies by means of critical appraisal criteria. METHODS: A systematic review of the journal literature between 1986 and the first half of 1997 was undertaken. Studies that have attempted to estimate the resource consequences of one or more vascular procedure were the focus of the review. The extent of reporting in each study was assessed by using published guidelines. RESULTS: The review identified 30 articles from nine different countries for inclusion in the study. Of these, more than half were published in the last 2(1/2) years of the search period, indicating a recent and rapid growth in economic studies in this area. When subjected to the reporting guidelines, the studies performed rather poorly overall. CONCLUSIONS: Although the vascular studies can be criticized for inadequate reporting of economic data, it appears from the limited evidence from elsewhere that inadequate reporting is a problem in other clinical areas. In view of the importance of reporting to the ability to critically assess studies-and thus separate the "good" from the "bad"-there is a need for reporting to improve future published studies. PMID- 10514207 TI - Early and long-term results of percutaneous transluminal angioplasty of the lower abdominal aorta. AB - PURPOSE: The purpose of this study was to determine the early and long-term results of percutaneous transluminal angioplasty (PTA) of atherosclerotic lower abdominal aorta stenosis. METHODS: This study was performed as a retrospective study. From 1980 to 1997, 46 patients with chronic lower limb ischemia with moderate to severe claudication as the result of isolated infrarenal disease or aortoiliac disease underwent PTA. All patients underwent angiography before and after angioplasty and Doppler ultrasound scan examination with ankle-brachial index determination. No stents were used. RESULTS: The technical success rate was 96% (44 of 46 cases). Thirty-eight patients (83%) immediately showed clinical, hemodynamic, and angiographic improvement. The initial success rate for patients with isolated infrarenal or bifurcation disease was 92%, whereas it was 71% for aortoiliac disease. Among the eight patients with no initial improvement, four had clinical deterioration and two required emergency surgical revascularization. There were no other complications. Fifty-six percent of the patient conditions (95% confidence interval [CI], 38% to 74%) remained clinically improved at the 5 year follow-up examination. Recurrence of symptoms was caused by femoropopliteal disease in most patients. The primary patency rate assumed with maintenance of hemodynamic improvements was 70% (95% CI, 52% to 88%) and 64% (95% CI, 44% to 84%) at 4 and 5 years of follow-up, respectively. The primary patency rate at 4 years for patients with isolated infrarenal or bifurcation disease was 83% (95% CI, 64% to 100%), whereas it was 55% for aortoiliac disease (95% CI, 30% to 80%; P =.06) The variables that were statistically predictive of patency failure were poor runoff (P =. 01) and presence of aortoiliac atherosclerotic disease (P =.04). CONCLUSION: Our results suggest that PTA is an excellent treatment for chronic arterial insufficiency of the lower extremities as the result of isolated atherosclerotic lower abdominal aortic occlusive lesions because of good long term patency. Aortic PTA for those patients with iliac involvement or with poor runoff gives acceptable results but carries lower patency and clinical success rates. PMID- 10514208 TI - Crossover iliofemoral bypass grafting for treatment of unilateral iliac atherosclerotic disease. AB - PURPOSE: In patients with unilateral iliac disease, a less invasive procedure than aortobifemoral bypass grafting may be desirable, especially in poor-risk patients or when sexual dysfunction is feared. In these cases, femorofemoral (FF) bypass grafting is often proposed. Compared with FF bypass grafting, iliofemoral (IF) bypass grafting avoids bilateral exposure of the groins, which may reduce the risk of infection. When the primitive iliac artery is occluded from its origin or heavily calcified, one may use the contralateral artery as inflow, after a small retroperitoneal exposure, to perform a crossover iliofemoral (CIF) bypass grafting procedure, through the Retzius space. Our 10-year experience with CIF bypass grafting in a select group of patients was studied. METHODS: Between 1986 and 1996, 36 patients underwent CIF bypass grafting for symptomatic unilateral iliac occlusion or stenosis. All patients were examined by means of Doppler ultrasound scanning and underwent bilateral multiplane angiography. Patients were considered for this procedure when the ipsilateral common iliac artery was occluded from its origin or was diffusely and heavily calcified. The decision to perform a CIF bypass grafting procedure was made when no significant disease of the contralateral common iliac artery was seen, and patients who had features of contralateral iliac disease were excluded. The main outcomes were perioperative mortality and morbidity, long-term primary and secondary patency rates, and limb salvage rate. RESULTS: The study included 31 men and five women, with a mean age of 58.8 years. Indications for bypass grafting were disabling claudication (26 of 36 patients, 72%) and limb-threatening ischemia (10 of 26 patients, 28%). Twelve procedures were performed simultaneously: endarterectomy of the recipient common femoral artery (n = 3), femoropopliteal bypass grafting (n = 4, 11.1%), profundoplasty (n = 4, 11%), and right internal carotid endarterectomy (n = 1). New postoperative erectile dysfunction did not develop in any of the patients. The survival rate was 97.3% at 1 year and 68.5% at 5 years. The primary and secondary patency rates were 94% and 100%, respectively, at 1 year and 76.7% and 95%, respectively, at 5 years. The limb salvage rate was 100% at 1 year and 87% at 3 years. CONCLUSION: The operative mortality associated with CIF is low. The long-term primary and secondary patency rates are satisfactory, and they are lower than those reported for aortobifemoral bypass grafting. This procedure does not preclude a later performance of an aortobifemoral bypass grafting procedure. CIF bypass grafting is not only suitable for poor-risk patients with a limited life expectancy who have the appropriate arterial anatomy, but also may be warranted for young patients in whom erectile dysfunction is feared. PMID- 10514210 TI - Evaluating and improving health-related quality of life in patients with varicose veins. AB - PURPOSE: We set out to assess the new Aberdeen Varicose Veins Questionnaire (Aberdeen Questionnaire) for the properties necessary for a valid measure of health outcome, to determine quality of life of patients with varicose veins, and to determine the effect of surgery on quality of life. METHODS: A prospective consecutive cohort of 137 patients undergoing varicose vein surgery completed the self-administered SF-36 and Aberdeen Questionnaire and 25 questions relating to the symptoms and concerns of patients with varicose veins. Follow-up was done by repeated questionnaires 6 weeks after surgery. The Aberdeen Questionnaire was assessed for reliability, validity, responsiveness, and practicality. Quality of life of patients with varicose veins was compared with an age- and sex-matched sample of the general population. RESULTS: Reliability estimates for the 8 scales short-form health survey (SF-36) and the Aberdeen Questionnaire were all above 0.7 (Cronbach's alpha). The Aberdeen Questionnaire had a highly significant correlation (r = 0. 74, P <.0001) with the patients' symptoms and concerns questionnaire, which is evidence of its validity. Patients with varicose veins score lower than United Kingdom norms (P <.001) in the physical domains of the SF 36, indicating worse health. After surgery, the SF-36 scores improved in all 8 domains of health, reaching significance in "Mental Health" (P <.05) and approaching significance in "General Health" (P =.066). The Health Transition Item of the SF-36 and the Aberdeen Questionnaire both showed a highly significant improvement in health (P <.001). CONCLUSION: The Aberdeen Questionnaire is a valid measure of quality of life for patients with varicose veins. Persons with varicose veins have a reduced quality of life compared with the general population, and this discrepancy is significantly improved at 6 weeks by operating on them. PMID- 10514209 TI - Systematic review of randomized controlled trials of aspirin and oral anticoagulants in the prevention of graft occlusion and ischemic events after infrainguinal bypass surgery. AB - PURPOSE: We sought to determine the efficacy of antiplatelet therapy and oral anticoagulants in maintaining graft patency and preventing ischemic complications in patients after infrainguinal bypass surgery. METHODS: We performed a meta analysis of randomized controlled trials of aspirin with or without other antiplatelet therapy and oral anticoagulants after infrainguinal bypass surgery. Outcome measures studied were graft occlusion, stroke, myocardial infarction, vascular and total mortality, and the composite outcome of stroke, myocardial infarction, and vascular mortality. RESULTS: Five trials of antiplatelet therapy versus placebo were included. The relative risk (RR) for occlusion was 0.78 (95% CI, 0.64-0.95). For prevention of stroke, myocardial infarction, and death, and for the composite outcome, no significant effect was measured. Only one trial of oral anticoagulants versus control treatment was included. The RR for occlusion was 0.55 (95% CI, 0.30-0.99), and that for amputation was 0.30 (95% CI, 0.10 0.87). The mortality rate did not differ significantly between the groups. One trial of oral anticoagulant therapy plus aspirin versus aspirin alone in high risk patients was included. The RR for occlusion was 0.38 (95% CI, 0.15-0. 95). There were no significant differences for prevention of amputation, myocardial infarction, and death between the groups. CONCLUSION: Antiplatelet therapy and oral anticoagulants reduce the risk of graft occlusion. Oral anticoagulant therapy appears to be the more effective treatment in high-risk patients. Data on the reduction of the risk of stroke, myocardial infarction, and death are inconclusive. Evidence for the beneficial effects of antiplatelet and oral anticoagulant therapy after infrainguinal bypass surgery is based on a small number of trials only. There is no proof as to which modality is the most effective in the prevention of graft occlusion and ischemic events in patients after infrainguinal bypass surgery, which is reason for a randomized comparison of aspirin with oral anticoagulants. PMID- 10514211 TI - Redone endoscopic perforator surgery: feasibility and failure analysis. AB - PURPOSE: In many hospitals and medical practices, subfascial endoscopic perforator surgery (SEPS) has become the treatment of choice in patients with incompetent perforator veins and active venous ulcers. A substantial number of surgeons consider SEPS to be an operation that can be performed only once because extensive scarring and narrowing of the subfascial space make a second endoscopic operation impossible. It is the purpose of this report to prove the feasibility, efficacy, and safety of a second SEPS procedure. METHODS: Within a period of 30 months, 105 primary SEPS procedures were performed in patients with healed or still active ulcers. In addition to these cases, within a period of 30 months, a consecutive number of 19 patients were examined and scheduled for a second SEPS procedure. All patients were in class 5 with healed ulcers or in class 6 with still active ulcers. The CEAP classification of the American Venous Forum was used to evaluate the results and to calculate the clinical, disability, and outcome scores. The redone operation was performed by using CO(2) insufflation, a dual-port technique, and subfascial balloon dissection. RESULTS: In two patients conversion to a conventional procedure was necessary. There were no major complications, but there was a 21% incidence of minor problems, such as hematoma or cellulitis. The mean total clinical score improved after surgery from 7.91 to 3.23 (P <.01), the disability score changed from 1.10 to 0.57 after surgery (P <.02), and the clinical outcome score was 1. 47 after surgery (P <.001). Cumulative ulcer healing could be achieved in 85.8% of class 6 patients. Failure analysis revealed that an incomplete subfascial dissection had been performed during the first endoscopic operation. A septum intermusculare medialis or an intact deep posterior fascia with incompetent Cockett II perforators were major factors contributing to the initial treatment failures. In addition to incompetent perforators, postthrombotic deep venous reflux was seen in eight (42.1%) patients, and four (21%) patients had a combination of secondary reflux and obstruction. CONCLUSION: Subfascial endoscopic procedures can be redone safely. In addition to exploring the superficial posterior compartment, the deep posterior compartment must be opened to prevent recurrent symptoms in patients with incompetent perforator veins. PMID- 10514212 TI - A practical approach to vascular access for hemodialysis and predictors of success. AB - PURPOSE: The long-term results and predictors of success for vascular access at The Toronto Hospital were studied. This report describes the access program and emphasizes the role of the vascular access coordinator. METHODS: A total of 384 consecutive patients underwent 466 vascular access procedures. The access program is centered around a dedicated, full-time vascular access coordinator, who is a registered nurse and is responsible for all aspects of access care, including follow-up. Outcome variables were collected prospectively. Primary, primary assisted, and secondary success was determined by means of Kaplan-Meier analysis, and the stepwise Cox proportional hazards model was used for multivariate analysis of the factors that were independently predictive of primary success. RESULTS: There were 235 autogenous arteriovenous fistulae (AVFs) and 231 arteriovenous grafts (AVGs). The cumulative primary, assisted-primary, and secondary success (patent and functional for effective dialysis) at 24 months for all 466 cases combined was 36% +/- 3%, 54% +/- 3%, and 66% +/- 3%, respectively. The primary success for AVFs and AVGs at 2 years was 54% +/- 4% and 18% +/- 4%, respectively (P <.001; log-rank test); the primary-assisted success for AVFs and AVGs at 2 years was 62% +/- 4% and 44% +/- 6%, respectively (P <.001; log-rank test); and the secondary success for AVFs and AVGs at 2 years was 70% +/- 4% and 60% +/- 5%, respectively (P =.331; log-rank test). Stratification of variables revealed significant benefit for AVFs (P =.001), the female sex (P =.014), and the absence of diabetes mellitus (P =.001). Multivariate analysis with Cox regression determined that access type (AVF vs AVG; P =.001) and diabetes mellitus (P =.024) were independently predictive of primary success. The improved clinical coordination of access patients with the initiation of the vascular access program resulted in a significant reduction in length of hospital stay before and after the program was organized (2.5 +/- 0.06 vs 1.1 +/- 0.03 days; P =.001). CONCLUSION: The organization of a vascular access program in a practical and cost-effective way for reduced length of hospital stay is streamlined through a dedicated access coordinator, who ensures an integrated, multidisciplinary approach. The results for the Cox model is useful when discussing the anticipated results of access procedures with individual patients. PMID- 10514213 TI - The proliferative capacity of neonatal skin fibroblasts is reduced after exposure to venous ulcer wound fluid: A potential mechanism for senescence in venous ulcers. AB - PURPOSE: We have previously shown that fibroblasts cultured from venous ulcers display characteristics of senescence and have reduced growth rates. Susceptibility of young fibroblasts to the microcirculatory changes associated with venous ulcers, such as macrophage trapping and activation, could explain the prevalence of senescent fibroblasts in these wounds. METHODS: We tested the in vitro effect of venous ulcer wound fluid (VUWF), as well as pro-inflammatory cytokines known to be present in VUWF (TNF-alpha, IL-1beta, and TGF-beta1), on neonatal foreskin fibroblasts (NFFs). NFF growth rates, cellular morphology, and senescence-associated beta-galactosidase (SA-beta-Gal) activity were determined in the presence or absence of VUWF and the above cytokines. VUWF TNF-alpha concentration and the effect of anti-TNF-alpha antibody on VUWF inhibitory activity were determined in samples obtained from four patients with venous ulcers. RESULTS: NFF growth rates were significantly reduced by VUWF (42,727 +/- 6301 vs 3902 +/- 2191 P =. 006). TNF-alpha also significantly reduced NFF growth rates in a dose-dependent manner (P =.01). No significant growth-inhibitory activity was seen for IL-1alpha or TGF-beta. Incubation with VUWF significantly increased the percentage of SA-beta-Gal-positive fibroblasts in vitro on culture day 12 (P =.02). TNF-alpha and TGF-beta1 had similar effects. TNF-alpha was detected in all VUWF tested, with a mean of 254 +/- 19 pg/mL. CONCLUSION: These data suggest that the venous ulcer microenvironment adversely affects young, rapidly proliferating fibroblasts such as NFFs and induces fibroblast senescence. Pro-inflammatory cytokines such as TNF-alpha and TGF-beta1 might be involved in this process. The role of other unknown inhibitory mediators, as well as pro inflammatory cytokines, in venous ulcer development and impaired healing must be considered. PMID- 10514214 TI - Dialysis access failure: A sheep model of rapid stenosis. AB - PURPOSE: Intimal hyperplasia at the venous anastomosis of dialysis access grafts causes early failure, although increased flow inhibits intimal hyperplasia in arterial grafts and after vessel injury. We designed a sheep model to study this process. METHODS: Polytetrafluoroethylene (PTFE) grafts were placed in the necks of sheep from the carotid artery to the external jugular vein. Grafts were harvested after perfusion fixation at 4, 8, and 12 weeks and submitted for histologic and immunohistochemical examination, including morphometry of neointimal lesions. RESULTS: The venous anastomoses developed thick neointima within the PTFE graft by 4 weeks. Lesions at the venous end were significantly thicker than those at the arterial end by 8 weeks (1.2 +/- 0.1 vs 0.38 +/- 0.05 mm, P <.02) and had greater cross-sectional area at both 4 (0.32 +/- 0.21 vs 3.6 +/- 0.8 mm(2), n = 7, P <.02) and 8 weeks (9.8 +/- 1.9 vs 1.1 +/- 0.7 mm(2), n = 7, P <.02). Only one of the four grafts (25%) in the 12-week group remained patent. Lesions were composed of smooth muscle cells, matrix, and thrombus of various ages. Cellular proliferation was prominent in neointima adjacent to thrombus and in granulation tissue surrounding the graft. Organizing thrombus contributed significantly to luminal narrowing. CONCLUSION: The sheep model of dialysis access reliably produces venous stenosis within 4 weeks. Lesions develop in the absence of uremia, graft puncture, or dialysis, suggesting that these factors are not necessary for graft failure. The continued presence of thrombus and high rates of cellular proliferation suggest ongoing injury is an important cause of lesion formation. This model allows study of the cellular mechanisms of dialysis failure. PMID- 10514215 TI - A specific inhibitor of apoptosis decreases tissue injury after intestinal ischemia-reperfusion in mice. AB - PURPOSE: Apoptosis is a stereotypical pathway of cell death that is orchestrated by a family of cysteine endoproteases called caspases. This study examined the effect of apoptosis inhibition with a specific caspase inhibitor on murine intestinal viability after ischemia-reperfusion (IR). METHODS: C57Bl6 X SV129 mice underwent segmental small bowel ischemia by vascular isolation of 10 cm of terminal ileum. In separate experiments, the ischemic time was varied from 30 to 130 minutes with a reperfusion interval of 6 hours. The degree of small bowel injury was quantified from 1 to 5 (increasing severity) by standardized, blinded histologic grading. The degree of apoptosis was assessed with a specific assay (terminal deoxyamcleotydil transferase-mediated deoxyuridine triphosphate nick end labeling [TUNEL]) and quantified by calculating the apoptotic index (apoptotic cells/10 high-power fields). To evaluate for activation of interleukin 1beta converting enzyme we measured tissue mature interleukin-1beta levels using a specific enzyme-linked immunosorbent assay. To evaluate the effect of apoptosis inhibition on intestinal viability after IR, mice received 3.0 mg of the caspase inhibitor ZVAD (N-benzyloxycarbonyl Val-Ala-Asp-Ome-fluoromethylketone) subcutaneously before and after IR in five divided doses (n = 11), the same dose of ZFA (N-benzyloxycarbonyl Phe-Ala fluoromethylketone), a structurally similar molecule with no anticaspase activity (n = 9), or sham operation (n = 6). RESULTS: A linear relationship existed between ischemic interval and histologic grade (r = 0.69, P <.006). Increasing the ischemic interval from 0 to 50 minutes was associated with a fivefold increase in apoptotic index (P =.05). Ischemic bowel was measured to have an average of 57.3 +/- 7.8 pg/mL whereas normal bowel had an average of 1.8 +/- 0.5 pg/mL of mature interleukin-1beta present. Mice tolerated multiple injections of ZVAD and ZFA without signs of toxicity. Animals treated with ZVAD (apoptosis inhibitor) had little injury after 50 minutes of ischemia and 6 hours of reperfusion (injury grade 1.8) compared with sham controls (injury grade 1.2, P =.7) and had significantly less injury than mice treated with ZFA (placebo) (injury grade 3.0, P <.006). CONCLUSIONS: Increasing ischemic interval in a segmental small bowel murine IR model is associated with increased histologic injury and augmented apoptosis as evidenced by increased TUNEL staining and interleukin-1beta converting enzyme activation. Inhibition of apoptosis with a specific caspase inhibitor significantly diminishes the degree of small bowel injury. PMID- 10514216 TI - Drag reducing polymers may decrease atherosclerosis by increasing shear in areas normally exposed to low shear stress. AB - PURPOSE: Drag reducing polymers (DRPs) have been shown to decrease plaque formation. Their mechanism of action is unknown. Atherosclerosis tends to develop in areas of low shear stress. This study investigates whether DRPs increase shear stress in areas normally exposed to low shear stress. METHODS: Six dogs underwent surgical plication of the left half of the aorta. A specially modified 20-MHz Doppler ultrasound probe mounted at a 45-degree angle on a micromanipulator was used to measure blood flow velocity at six 4-mm intervals along both lateral sides of the aorta starting at the aortic wall and then at subsequent 0.1-mm depths moving into the lumen before and after administering DRP. Shear rates were calculated using linear regression and then compared using the paired t test. The blood viscosity remained constant at 0.04 poise during infusions of this amount of DRP. RESULTS: The maximum shear rate occurring during the cardiac cycle on the side of the aortic stenosis (plication) was 9.96 +/- 1.52/sec before the administration of the DRP and 14.27 +/- 2.01/sec after the administration of the DRP (P =.0240). The maximum shear rate on the side of the unstenosed aortic wall was 57.25 +/- 7.93/sec before the administration of the DRP and 44.80 +/- 6.23/sec after the administration of the DRP (P =. 0081). CONCLUSION: One of the ways that DRPs inhibit the development of atherosclerosis appears to be by increasing shear stress in areas normally exposed to low shear stress. Understanding this mechanism may lead to the development of pharmaceutical agents that inhibit the development of atherosclerosis. PMID- 10514217 TI - Regarding "Coagulation and fibrinolysis in patients undergoing operation for ruptured and nonruptured infrarenal abdominal aortic aneurysms". PMID- 10514219 TI - Reply PMID- 10514218 TI - Regarding "Problems with the dissemination of up-to-date information on the results of endograft repair for abdominal aortic aneurysm". PMID- 10514220 TI - Regarding "Rupture of a nonaneurysmal atherosclerotic infrarenal aorta". PMID- 10514221 TI - Regarding "Can duplex scan arterial mapping replace contrast arteriography as the test of choice before infrainguinal revascularization?". PMID- 10514222 TI - Regarding "Endovascular treatment of renal artery thrombosis caused by umbilical artery catheterization". PMID- 10514223 TI - Reply PMID- 10514224 TI - Herbert henri jasper PMID- 10514225 TI - Bell's palsy and HSV-1 infection. PMID- 10514226 TI - Intravenous immunoglobulin in the treatment of autoimmune neuromuscular diseases: present status and practical therapeutic guidelines. AB - This review summarizes the current status of intravenous immunoglobulin (IVIg) in the treatment of autoimmune neuromuscular disorders and the possible mechanisms of action of the drug based on work in vivo, in vitro, and in animal models. Supply of idiotypic antibodies, suppression of antibody production, or acceleration of catabolism of immunoglobulin G (IgG) are relevant in explaining the efficacy of IVIg in myasthenia gravis (MG), Lambert-Eaton myasthenic syndrome (LEMS), and antibody-mediated neuropathies. Suppression of pathogenic cytokines has putative relevance in inflammatory myopathies and demyelinating neuropathies. Inhibition of complement binding and prevention of membranolytic attack complex (MAC) formation are relevant in dermatomyositis (DM), Guillain-Barre syndrome (GBS), and MG. Modulation of Fc receptors or T-cell function is relevant in chronic inflammatory demyelinating polyneuropathy (CIDP), GBS, and inflammatory myopathies. The clinical efficacy of IVIg, based on controlled clinical trials conducted in patients with GBS, CIDP, multifocal motor neuropathy (MMN), DM, MG, LEMS, paraproteinemic IgM anti-myelin-associated glycoprotein (anti-MAG) demyelinating polyneuropathies, and inclusion body myositis is summarized and practical issues related to each disorder are addressed. The present role of IVIg therapy in other disorders based on small controlled or uncontrolled trials is also summarized. Finally, safety issues, risk factors, adverse reactions, spurious results or serological tests, and practical guidelines associated with the administration of IVIg in the treatment of neuromuscular disorders are presented. PMID- 10514227 TI - Charcot-marie-tooth disease and related neuropathies: molecular basis for distinction and diagnosis. AB - Great advances have been made in understanding the molecular basis of Charcot Marie-Tooth disease (CMT) and related neuropathies, namely Dejerine-Sottas disease (DSD), hereditary neuropathy with liability to pressure palsies (HNPP) and congenital hypomyelination (CH). The number of newly uncovered mutations and identified genetic loci is rapidly increasing, and, as a consequence, the classification of these disorders is becoming more complicated. Molecular genetics, animal models, and transfected cell studies are shedding light on function and dysfunction of proteins involved in hereditary myelinopathies peripheral myelin protein 22 (PMP22), myelin protein zero (PO), connexin 32 (Cx32), and early growth response 2 (EGR2). Gene dosage effect, loss of function, gain of toxic function, and dominant negative effect are possible mechanisms whereby different gene mutations may exert their detrimental action on peripheral nerves. A tentative rational approach to clinical and molecular diagnosis based on genotype-phenotype correlation analysis is described. PMID- 10514228 TI - Effect of movement on dipolar source activities of somatosensory evoked potentials. AB - The early scalp somatosensory evoked potentials (SEPs) to median and tibial nerve stimulation were recorded at rest and during voluntary movement of the stimulated hand and foot, respectively. Both tibial and median nerve SEP distributions at rest could be explained by four-dipole models, in which one dipole was activated at the same latency as the subcortical far field and the three remaining dipolar sources were located in the perirolandic region contralateral to the stimulated side. Voluntary movement reduced all cortical dipoles in strength, while the subcortical one remained unchanged, suggesting that the effect of movement occurs above the cervicomedullary junction. In animals, cutaneous inputs are suppressed during movement and we therefore interpreted the depression of activity in the primary somatosensory cortex induced by movement as due to selective "gating" of cutaneous afferents. Because the reduction in strength of the cortical dipoles was generally lower during passive than active movement, both centrifugal and centripetal mechanisms probably contribute to the phenomenon of "gating." PMID- 10514229 TI - Modular organization of human leg withdrawal reflexes elicited by electrical stimulation of the foot sole. AB - Human withdrawal reflex receptive fields were determined for leg muscles by randomized, electrical stimulation at 16 different positions on the foot sole. Tibialis anterior, gastrocnemius medialis, peroneus longus, soleus, rectus femoris, and biceps femoris reflexes, and ankle joint angle changes were recorded from 14 subjects in sitting position. Tibialis anterior reflexes were evoked at the medial, distal foot and correlated well with ankle dorsal flexion. Gastrocnemius medialis reflexes were evoked on the heel and correlated with plantar flexion. Stimulation on the distal, medial sole resulted in inversion (correlated best with tibialis anterior activity), whereas stimulation of the distal, lateral sole evoked eversion. Biceps femoris reflexes were evoked on the entire sole followed by a small reflex in rectus femoris. A detailed withdrawal reflex organization, in which each lower leg muscle has its own receptive field, may explain the ankle joint responses. The thigh activity consisted primarily of flexor activation. PMID- 10514230 TI - Current perception threshold testing in Fabry's disease. AB - We investigated 16 patients with Fabry's disease (eight hemizygous men and eight heterozygous women) in one family. We used constant current perception threshold (CPT) testing, which evaluated three major sensory nerve fiber populations, to assess subjective complaints of pain and paresthesias. We also examined clinical and biochemical features and compared the values of CPTs and nerve conduction studies (NCS) in detecting the sensory neuropathy. Our results showed that CPT testing at low frequencies (5 and 250 Hz) was significantly more sensitive than at a higher frequency (2 kHz) and NCS in detecting sensory neuropathy in patients with Fabry's disease. However, there was no correlation between CPT testing and clinical symptom scores, duration of disease, creatinine clearance (Ccr) values or alpha-galactosidase A (AGA) activities in either hemizygous or heterozygous patients. Hemizygous patients clinically demonstrated more severe symptom scores, poorer renal function, and higher prevalence of hypohidrosis and corpora angiokeratomas than did heterozygous patients, which indicates that detailed clinical examinations can differentiate the clinical status of hemizygous men from heterozygous women. There were no associations between the biochemical levels of serum AGA activity and renal function (Ccr values) or the symptom scores (grading of acroparesthesia), indicating that biochemical parameters do not predict clinical severity. PMID- 10514231 TI - Intraoperative electrical stimulation for identification of cranial nerve nuclei. AB - The purpose of this study was to evaluate the feasibility and usefulness of cranial nerve nuclei monitoring during resection of brainstem cavernous malformations. Eleven patients with brainstem cavernous malformations underwent resection of their malformations utilizing cranial nerve nuclei monitoring. Cranial nerves V and VII were monitored by placing electrodes in muscle groups innervated by these nerves and recording manipulation-induced neurotonic discharges and triggered electromyographic (EMG) activity, after electrical stimulation of the corresponding brainstem nuclei. Seven of 11 procedures (64%) with cranial nerve nuclei monitoring were noted to have cranial nerve nuclei activity corresponding to manipulation of the nuclei. The cavernous malformation was completely resected in 5 of 7 cases with cranial nerve nuclei activity and in all 4 cases without activity. In the remaining 2 cases, the cavernous malformation was not resected due to the proximity of the monitored cranial nerve nuclei to the cavernous malformation and to increasing neurotonic activity as the cavernous malformation was approached. None of the 11 patients had new permanent postoperative deficits corresponding to the cranial nerve nuclei monitored; 1 patient had a transient partial facial palsy lasting 2 days. Preliminary results indicate that cranial nerve nuclei monitoring proves useful in preserving neurologic function and reducing surgical morbidity during resection of brainstem cavernous malformations, particularly indicating when lesion resection places these nuclei at risk. PMID- 10514232 TI - Modulation of motor evoked potentials by muscle vibration: the role of vibration frequency. AB - Augmentation of motor evoked potentials (MEPs) by muscle vibration (MV) was studied in 10 healthy subjects with regard to the vibration frequency (VF). The extensor carpi radialis muscle (ECR) was vibrated using VFs of 80, 120, and 160 Hz. Motor evoked potentials following transcranial magnetic stimulation were recorded simultaneously from the vibrated ECR and the antagonist flexor carpi radialis muscle (FCR) without MV, 0.5 s and 3 s after onset of MV and 1 s after offset. Only the VFs of 80 Hz and 120 Hz caused MEP augmentation and latency shortening in ECR, whereas depression of MEPs in FCR was induced by all VFs used. It appears that MEP augmentation and latency shortening in ECR are mediated by the primary muscle spindle endings which respond with optimal discharge rates to VFs of up to 100 Hz. Motor evoked potential depression in FCR, being well expressed also with VF 160 Hz, seems to involve other dynamic mechanoreceptors. PMID- 10514233 TI - Cardiomyopathy in duchenne, becker, and sarcoglycanopathies: a role for coronary dysfunction? AB - Dilated cardiomyopathy is a feature of Duchenne and Becker muscular dystrophies and occasionally of sarcoglycanopathies. Its pathogenesis is unknown. Patients with myotonic dystrophy have an impairment of coronary smooth muscle and this could contribute to their cardiomyopathy. We used positron emission tomography (PET) to study myocardial blood flow and coronary vasodilator reserve at baseline and during hyperemia in 7 Duchenne, 8 Becker, and 5 sarcoglycanopathy patients. The study was normal in all Becker patients. In contrast, baseline myocardial blood flow was increased and coronary vasodilator reserve blunted in Duchenne and sarcoglycanopathy patients despite normal hyperemic myocardial blood flow. The reduction of coronary vasodilator reserve was due to an increased baseline myocardial blood flow. In Duchenne dystrophy, but not in sarcoglycanopathies, correction for cardiac workload normalized the coronary vasodilator reserve. In the latter patients, abnormal baseline myocardial blood flow could be due to vascular smooth muscle dysfunction. PMID- 10514234 TI - Changes of forearm EMG and cerebral evoked potentials following sudden muscle stretch in patients with Huntington's disease. AB - Various investigators have reported that the late-reflex electromyographic (EMG) activity following muscle stretch is decreased in Huntington's disease. To explore the basis of this decreased activity, we recorded the late EMG responses together with associated cerebral responses following muscle stretch in patients with Huntington's disease. Five patients and seven controls voluntarily participated in two sets of experiments in which they grasped a handle attached to a torque motor and maintained the wrist isometrically against a constant flexor force of 2.3 newtons (N). The force was changed unpredictably (first set of experiments) or predictably (second set) to 10.4 N, causing a stretch of wrist extensors or flexors. Rectified surface EMG from the extensor and flexor carpi radialis muscles was integrated for the M2 and M3 components of the late responses. Cerebral responses were recorded from F3, F4, C3, C4, and Cz and averaged separately depending upon condition. The late muscle responses to unpredictable muscle stretch were decreased or absent in patients with Huntington's disease. The cerebral responses recorded at Cz differed markedly between patients and controls, beginning approximately 15 ms prior to the onset of the late M2 muscle response. Although the initial positivity was similar in amplitude, all other cerebral components were markedly reduced in the patient group. Both controls and patients showed a markedly attenuated cerebral response when the muscle stretch was predictable. The electrocerebral response to muscle stretch is thus altered prior to the onset of M2 in patients with Huntington's disease, suggesting that the long-latency reflex involves transcerebral pathways that are affected in this disease. PMID- 10514235 TI - The effect of differential and complete nerve block on experimental muscle pain in humans. AB - We have attempted to examine the nerve fiber population mediating experimentally induced muscle pain in humans. Two established methods induced muscle pain: continuous, intramuscular, electrical stimulation and intramuscular infusion of hypertonic saline. A progressive nerve block was achieved by a combination of compression nerve block and intravenous regional anesthesia. Regular tests of muscle pain intensity were performed during the blocking period of 60 min. At the same time, the blocking of thick and thin afferents was monitored by assessment of proprioception and cutaneous touch, pin-prick, pressure pain, and heat detection thresholds. Electrically induced muscle pain was inhibited (P < 0.0001) in parallel with proprioception, touch, and pin-prick, which were mediated by thick and thin myelinated nerve fibers. Saline-induced muscle pain was inhibited (P < 0.002) synchronously with heat detection and pressure pain, which are mediated by unmyelinated nerve fibers. Based on the present psychophysical experiments, it is suggested that: (1) myelinated afferents mediated mainly electrically induced muscle pain, and (2) unmyelinated afferents mediated mainly saline-induced muscle pain. PMID- 10514236 TI - Motor unit number estimate-based rates of progression of ALS predict patient survival. AB - We have examined, as predictors of survival in patients with amyotrophic lateral sclerosis (ALS), linear estimates of rates of disease progression (LEP), based on motor unit number estimates (MUNE). Motor unit number estimates of thenar, hypothenar, and extensor digitorum brevis muscles (according to the manual method of McComas), isometric grip and foot dorsiflexion (FD) strength, and forced vital capacity (FVC) were available in 34 patients. Linear estimates of rates of disease progression were derived. Probability of survival was calculated using the Kaplan-Meier method. Motor unit number estimates, LEP based on MUNE, and demographic characteristics were tested as risk factors within the Cox Proportional Hazards Model, using regression techniques. Individually, all MUNE based LEP were highly significant (P < 0.00005); bulbar onset attained modest significance (P = 0.044). Secondary analysis showed MUNE-based LEP were more significant than regionally concordant function-based LEP. Linear estimates of rates of disease progression based on MUNE may thus predict survival of patients with ALS better than LEP based on function. PMID- 10514238 TI - A relief maneuver in carpal tunnel syndrome. AB - We describe a maneuver that eases or abolishes paresthesias in carpal tunnel syndrome. With the affected hand palm up, the distal metacarpal heads are gently squeezed together; in some instances stretch of digits III and IV is also required. This maneuver may help in the clinical diagnosis of carpal tunnel syndrome, can be useful as a means of relieving symptoms, and provides the basis for the design of an innovative splint. PMID- 10514237 TI - Leukemia inhibitory factor, glial cell line-derived neurotrophic factor, and their receptor expressions following muscle crush injury. AB - Using in situ hybridization histochemistry, we characterized the spatiotemporal gene expression patterns of leukemia inhibitory factor (LIF) and glial cell line derived neurotrophic factor (GDNF), and their receptor components (LIFR, GFR alpha1, RET) induced in muscle cells, intramuscular nerves, and motoneurons in the regeneration processes of both muscle cells and nerves following muscle contusion. Muscle contusion induced upregulation of GDNF and GFR-alpha1 mRNAs in Schwann cell-like cells in the intramuscular nerves and of LIFR mRNA in damaged muscle cells. LIFR, GFR-alpha1, and RET mRNA expressions in motoneurons were upregulated following muscle contusion. Muscle contusion also induced more rapid, prominent transactivations of GFR-alpha1 and RET genes in motoneurons than did sciatic nerve axotomy. These findings suggest that rapid and prominent upregulation of the receptor components for LIF and GDNF in motoneurons is important for the regeneration of intramuscular motor nerves damaged by muscle contusion. PMID- 10514239 TI - The silent period after magnetic brain stimulation in generalized tetanus. AB - The cortical silent period has not previously been studied in tetanus. Transcranial magnetic brain stimulation in a patient with generalized tetanus revealed enlarged electromyographic (EMG) responses and absence or reduction of the late phase of EMG silence following the motor evoked potential in sternomastoid and biceps brachii muscles. Following clinical recovery, the silent period returned to normal. This observation is interpreted as evidence of impaired inhibitory mechanisms at multiple levels of the nervous system, including the cortex, in generalized tetanus. PMID- 10514240 TI - Atypical phenotype of charcot-marie-tooth disease type 1A. AB - Two sisters with a Charcot-Marie-Tooth disease type 1A (CMT1A) duplication, who had an unusual CMT1A clinical phenotype, are described. The 63-year-old proband presented with dysesthesia on the inner side of the right leg. Neurological examination revealed a localized sensory disturbance in the lower extremities and mild weakness in the feet and left hand. Her 61-year-old sister had experienced several episodes of acute paralysis, and neurological examination showed moderate, sensory-dominant polyneuropathy. A reduction of myelinated fibers with many onion-bulb formations were observed in the sural nerve of the proband, and electrophysiological studies showed reduced motor nerve conduction velocities in both sisters. To diagnose CMT1A, we developed a CMT1A duplication test based on detection of CMT1A-specific junction fragments using the long polymerase chain reaction (PCR) method. A 3.3-kb CMT1A-specific junction fragment was detected in both patients, and their neuropathy may therefore have been associated with CMT1A duplication. PMID- 10514241 TI - Bilateral medial pectoral neuropathy in a weight lifter. AB - Weight lifting has occasionally been associated with focal neuropathies of unknown cause. We describe a case of progressive bilateral medial pectoral neuropathy in a body builder. Whereas the medial pectoral nerve passes through pectoralis minor to reach the pectoralis major, the lateral pectoral nerve has no such intramuscular course. We postulate that weight lifting and concomitant pectoralis minor hypertrophy in our patient produced intramuscular entrapment of the medial pectoral nerves. PMID- 10514242 TI - Myasthenia gravis, thymoma, intestinal pseudo-obstruction, and neuronal nicotinic acetylcholine receptor antibody. AB - Intestinal pseudo-obstruction occurs rarely in patients with myasthenia gravis (MG) and thymoma. The etiology of the intestinal pseudo-obstruction remains to be elucidated, although an autoimmune mechanism is postulated. We present the first report of neuronal nicotinic acetylcholine receptor (AChR)-specific antibody in a patient with seropositive MG, malignant thymoma, and intestinal pseudo obstruction. This finding provides evidence that intestinal pseudo-obstruction associated with thymoma and possibly other neoplasms may be related to antibodies against the neuronal nicotinic receptors at autonomic ganglia. PMID- 10514243 TI - Complex repetitive discharges: cause or effect of neurogenic muscle hypertrophy? AB - We report a patient with adult-onset spinal muscular atrophy (SMA) of the scapulohumeral type with neurogenic muscle hypertrophy (NMH) in markedly weakened biceps muscles in association with continuous complex repetitive discharges (CRDs). This is an apparently unique case due to the bilaterality of the NMH associated with CRDs as well as the well-circumscribed symmetric upper extremity distribution of the hypertrophy. The possible mechanisms of NMH in association with spontaneous motor activity are discussed. PMID- 10514244 TI - Subacute sensory neuropathy associated with Epstein-Barr virus. AB - A 35-year-old man experienced severe sensory loss, pseudoathetosis, and areflexia during recovery from a severe viral illness. Sensory nerve action potentials were absent, motor conduction velocities were mildly slowed, and blink reflexes were normal. Magnetic resonance imaging (MRI) revealed abnormal signal within the central and dorsal aspects of the thoracic cord. Acute and convalescent Epstein Barr virus (EBV) titers suggested EBV as the etiology. Subacute sensory neuropathy, with peripheral and central nervous system involvement, is a rare complication of EBV infection. PMID- 10514246 TI - Reply PMID- 10514245 TI - AAEM news and comments PMID- 10514247 TI - Biobased Industrial Products: Bioprocess Engineering When Cost Really Counts. PMID- 10514248 TI - Biocommodity Engineering. AB - The application of biotechnology to the production of commodity products (fuels, chemicals, and materials) offering benefits in terms of sustainable resource supply and environmental quality is an emergent area of intellectual endeavor and industrial practice with great promise. Such "biocommodity engineering" is distinct from biotechnology motivated by health care at multiple levels, including economic driving forces, the importance of feedstocks and cost motivated process engineering, and the scale of application. Plant biomass represents both the dominant foreseeable source of feedstocks for biotechnological processes as well as the only foreseeable sustainable source of organic fuels, chemicals, and materials. A variety of forms of biomass, notably many cellulosic feedstocks, are potentially available at a large scale and are cost-competitive with low-cost petroleum whether considered on a mass or energy basis, and in terms of price defined on a purchase or net basis for both current and projected mature technology, and on a transfer basis for mature technology. Thus the central, and we believe surmountable, impediment to more widespread application of biocommodity engineering is the general absence of low-cost processing technology. Technological and research challenges associated with converting plant biomass into commodity products are considered relative to overcoming the recalcitrance of cellulosic biomass (converting cellulosic biomass into reactive intermediates) and product diversification (converting reactive intermediates into useful products). Advances are needed in pretreatment technology to make cellulosic materials accessible to enzymatic hydrolysis, with increased attention to the fundamental chemistry operative in pretreatment processes likely to accelerate progress. Important biotechnological challenges related to the utilization of cellulosic biomass include developing cellulase enzymes and microorganisms to produce them, fermentation of xylose and other nonglucose sugars, and "consolidated bioprocessing" in which cellulase production, cellulose hydrolysis, and fermentation of soluble carbohydrates to desired products occur in a single process step. With respect to product diversification, a distinction is made between replacement of a fossil resource derived chemical with a biomass-derived chemical of identical composition and substitution of a biomass-derived chemical with equivalent functional characteristics but distinct composition. The substitution strategy involves larger transition issues but is seen as more promising in the long term. Metabolic engineering pursuant to the production of biocommodity products requires host organisms with properties such as the ability to use low-cost substrates, high product yield, competitive fitness, and robustness in industrial environments. In many cases, it is likely to be more successful to engineer a desired pathway into an organism having useful industrial properties rather than trying to engineer such often multi-gene properties into host organisms that do not have them naturally. Identification of host organisms with useful industrial properties and development of genetic systems for these organisms is a research challenge distinctive to biocommodity engineering. Chemical catalysis and separations technologies have important roles to play in downstream processing of biocommodity products and involve a distinctive set of challenges relative to petrochemical processing. At its current nascent state of development, the definition and advancement of the biocommodity field can benefit from integration at multiple levels. These include technical issues associated with integrating unit operations with each other, integrating production of individual products into a multi-product biorefinery, and integrating biorefineries into the broader resource, economic, and environmental systems in which they function. We anticipate that coproduction of multiple products, for example, production of fuels, chemicals, power, and/or feed, is likely to be essential for economic viability. Lifecycle analysis is necessary to verify the sustainability and environmental quality benefits of a particular biocommodity product or process. We see biocommodity engineering as a legitimate focus for graduate study, which is responsive to an established personnel demand in an industry that is expected to grow in the future. Graduate study in biocommodity engineering is supported by a distinctive blend of intellectual elements, including biotechnology, process engineering, and resource and environmental systems. PMID- 10514249 TI - Process Design and Costing of Bioethanol Technology: A Tool for Determining the Status and Direction of Research and Development. AB - Bioethanol is a fuel-grade ethanol made from trees, grasses, and waste materials. It represents a sustainable substitute for gasoline in today's passenger cars. Modeling and design of processes for making bioethanol are critical tools used in the U.S. Department of Energy's bioethanol research and development program. We use such analysis to guide new directions for research and to help us understand the level at which and the time when bioethanol will achieve commercial success. This paper provides an update on our latest estimates for current and projected costs of bioethanol. These estimates are the result of very sophisticated modeling and costing efforts undertaken in the program over the past few years. Bioethanol could cost anywhere from $1.16 to $1.44 per gallon, depending on the technology and the availability of low cost feedstocks for conversion to ethanol. While this cost range opens the door to fuel blending opportunities, in which ethanol can be used, for example, to improve the octane rating of gasoline, it is not currently competitive with gasoline as a bulk fuel. Research strategies and goals described in this paper have been translated into cost savings for ethanol. Our analysis of these goals shows that the cost of ethanol could drop by 40 cents per gallon over the next ten years by taking advantage of exciting new tools in biotechnology that will improve yield and performance in the conversion process. PMID- 10514250 TI - Substrate and Enzyme Characteristics that Limit Cellulose Hydrolysis. AB - The ability and, consequently, the limitations of various microbial enzyme systems to completely hydrolyze the structural polysaccharides of plant cell walls has been the focus of an enormous amount of research over the years. As more and more of these extracellular enzymatic systems are being identified and characterized, clear similarities and differences are being elucidated. Although much has been learned concerning the structures, kinetics, catalytic action, and interactions of enzymes and their substrates, no single mechanism of total lignocellulosic saccharification has been established. The heterogeneous nature of the supramolecular structures of naturally occurring lignocellulosic matrices make it difficult to fully understand the interactions that occur between enzyme complexes and these substrates. However, it is apparent that the efficacy of enzymatic complexes to hydrolyze these substrates is inextricably linked to the innate structural characteristics of the substrate and/or the modifications that occur as saccharification proceeds. This present review is not intended to conclusively answer what factors control polysaccharide biodegradation, but to serve as an overview illustrating some of the potential enzymatic and structural limitations that invariably influence the complete hydrolysis of lignocellulosic polysaccharides. PMID- 10514251 TI - Enzymes, Energy, and the Environment: A Strategic Perspective on the U.S. Department of Energy's Research and Development Activities for Bioethanol. AB - For well over one hundred years, researchers around the world have pursued ways to make ethanol from biomass such as wood, grasses, and waste materials. To distinguish it from ethanol made from starch and sugars in traditional agricultural crops, we refer to ethanol made from biomass as "bioethanol." The effort to develop bioethanol technology gained significant momentum in the late 1970s as a result of the energy crises that occurred in that decade. This article briefly reviews the broader history of bioethanol technology development. With this as a background, we focus our attention on the strategic thinking behind the U.S. Department of Energy's Bioethanol Program, which envisions remarkable advances in cellulase enzyme research and as the basis for significant future process cost reductions. PMID- 10514253 TI - Reactor Design Issues for Synthesis-Gas Fermentations. AB - Synthesis gas is readily obtained by gasifying coal, oil, biomass, or waste organics and represents an abundant, potentially inexpensive, feedstock for bioprocessing. The primary components of synthesis gas, carbon monoxide and hydrogen, can be converted into methane, organic acids, and alcohols via anaerobic fermentations. Bioconversion of synthesis gas is an attractive alternative to catalytic processing because the biological catalysts are highly specific and often more tolerant of sulfur contaminants than inorganic catalysts. However, because the aqueous solubilities of carbon monoxide and hydrogen are low, synthesis-gas fermentations are typically limited by the rate of gas-to liquid mass transfer. Consequently, a major engineering challenge in commercial development of synthesis-gas fermentations is to provide sufficient gas mass transfer in an energy-efficient manner. This paper reviews recent progress in the development of synthesis-gas fermentations, with emphasis on efforts to increase the efficiency of gas mass transfer. Metabolic properties of several microbes able to ferment synthesis gas are described. Results of synthesis-gas fermentations conducted in various bioreactor configurations are summarized. Recent results showing enhancement of synthesis-gas fermentations using microbubble dispersions are presented, and studies of the mass-transfer and coalescence properties of microbubbles are described. PMID- 10514252 TI - Cloning and expression of Trichoderma reesei cellobiohydrolase I in Pichia pastoris. AB - Pichia pastoris was transformed with the Trichoderma reesei cbh1 gene, and the recombinant enzyme was purified and analyzed kinetically and by circular dichroism. The P. pastoris rCBH I was recognized by MoAb raised to T. reesei CBH I but was found in multiple molecular weight species on SDS-PAGE gels. Carbohydrate content determination and SDS-PAGE western analysis indicated that the recombinant protein was hyperglycosylated, although a species very similar in molecular weight to the T. reesei enzyme could be isolated chromatographically. The P. pastoris rCBH I also demonstrated activity toward soluble and insoluble substrates (i.e., pNPL and Sigmacell), although at a level significantly lower than the wild-type enzyme. More seriously, the yeast-expressed enzyme showed non wild-type secondary structure by circular dichroism. We conclude that P. pastoris may not serve as an adequate host for the site-directed mutagenesis of T. reesei CBH I. PMID- 10514254 TI - Catalytic Upgrading of Fermentation-Derived Organic Acids. AB - The production of organic acids in low-cost, high-efficiency fermentation processes makes available a new route to chemical production from biomass. Because of their multiple functional groups and high reactivity, organic acids can undergo a variety of reactions that are effectively catalyzed by inorganic heterogeneous or homogeneous catalysts. Lactic acid and succinic acid, in particular, are approaching large-scale production via fermentation and show excellent promise as feedstocks for catalytic conversion routes such as hydrogenation, dehydration, or condensation. A number of catalytic conversion pathways of organic acids are potentially competitive with petroleum-based routes in the current economic environment, particularly when integrated into existing biomass/crop processing schemes. This article reviews some of the key reaction pathways available using fermentation-derived organic acids as feedstocks and presents recent results from the authors' lab on succinate hydrogenation to 1,4 butanediol and tetrahydrofuran. By a judicious choice of support properties and reaction conditions, it is possible to achieve yields of either of these two products in excess of 80%. PMID- 10514255 TI - Enteric bacterial catalysts for fuel ethanol production. AB - The technology is available to produce fuel ethanol from renewable lignocellulosic biomass. The current challenge is to assemble the various process options into a commercial venture and begin the task of incremental improvement. Current process designs for lignocellulose are far more complex than grain to ethanol processes. This complexity results in part from the complexity of the substrate and the biological limitations of the catalyst. Our work at the University of Florida has focused primarily on the genetic engineering of Enteric bacteria using genes encoding Zymomonas mobilis pyruvate decarboxylase and alcohol dehydrogenase. These two genes have been assembled into a portable ethanol production cassette, the PET operon, and integrated into the chromosome of Escherichia coli B for use with hemicellulose-derived syrups. The resulting strain, KO11, produces ethanol efficiently from all hexose and pentose sugars present in the polymers of hemicellulose. By using the same approach, we integrated the PET operon into the chromosome of Klebsiella oxytoca to produce strain P2 for use in the simultaneous saccharification and fermentation (SSF) process for cellulose. Strain P2 has the native ability to ferment cellobiose and cellotriose, eliminating the need for one class of cellulase enzymes. Recently, the ability to produce and secrete high levels of endoglucanase has also been added to strain P2, further reducing the requirement for fungal cellulase. The general approach for the genetic engineering of new biocatalysts using the PET operon has been most successful with Enteric bacteria but was also extended to Gram positive bacteria, which have other useful traits for lignocellulose conversion. Many opportunities remain for further improvements in these biocatalysts as we proceed toward the development of single organisms that can be used for the efficient fermentation of both hemicellulosic and cellulosic substrates. PMID- 10514256 TI - Fermentations with new recombinant organisms. AB - United States fuel ethanol production in 1998 exceeded the record production of 1.4 billion gallons set in 1995. Most of this ethanol was produced from over 550 million bushels of corn. Expanding fuel ethanol production will require developing lower-cost feedstocks, and only lignocellulosic feedstocks are available in sufficient quantities to substitute for corn starch. Major technical hurdles to converting lignocellulose to ethanol include the lack of low-cost efficient enzymes for saccharification of biomass to fermentable sugars and the development of microorganisms for the fermentation of these mixed sugars. To date, the most successful research approaches to develop novel biocatalysts that will efficiently ferment mixed sugar syrups include isolation of novel yeasts that ferment xylose, genetic engineering of Escherichia coli and other gram negative bacteria for ethanol production, and genetic engineering of Saccharoymces cerevisiae and Zymomonas mobilis for pentose utilization. We have evaluated the fermentation of corn fiber hydrolyzates by the various strains developed. E. coli K011, E. coli SL40, E. coli FBR3, Zymomonas CP4 (pZB5), and Saccharomyces 1400 (pLNH32) fermented corn fiber hydrolyzates to ethanol in the range of 21-34 g/L with yields ranging from 0.41 to 0.50 g of ethanol per gram of sugar consumed. Progress with new recombinant microorganisms has been rapid and will continue with the eventual development of organisms suitable for commercial ethanol production. Each research approach holds considerable promise, with the possibility existing that different "industrially hardened" strains may find separate applications in the fermentation of specific feedstocks. PMID- 10514257 TI - Microbial synthesis of 3-dehydroshikimic acid: a comparative analysis of D xylose, L-arabinose, and D-glucose carbon sources. AB - 3-Dehydroshikimic acid is a hydroaromatic precursor to chemicals ranging from L phenylalanine to adipic acid. The concentration and yield of 3-dehydroshikimic acid microbially synthesized from various carbon sources has been examined under fed-batch fermentor conditions. Examined carbon sources included D-xylose, L arabinose, and D-glucose. A mixture consisting of a 3:3:2 molar ratio of glucose/xylose/arabinose was also evaluated as a carbon source to model the composition of pentose streams potentially resulting from the hydrolysis of corn fiber. Escherichia coli KL3/pKL4.79B, which overexpresses feedback-insensitive DAHP synthase, synthesizes higher concentrations and yields of 3-dehydroshikimic acid when either xylose, arabinose, or the glucose/xylose/arabinose mixture is used as a carbon source relative to when glucose alone is used as a carbon source. E. coli KL3/pKL4.124A, which overexpresses transketolase and feedback insensitive DAHP synthase, synthesizes higher concentrations and yields of 3 dehydroshikimic acid when the glucose/xylose/arabinose mixture is used as the carbon source relative to when either xylose or glucose is used as a carbon source. Observed high-titer, high-yielding synthesis of 3-dehydroshikimic acid from the glucose/xylose/arabinose mixture carries significant ramifications relevant to the employment of corn fiber in the microbial synthesis of value added chemicals. PMID- 10514258 TI - Novel secretion system of recombinant Saccharomyces cerevisiae using an N terminus residue of human IL-1 beta as secretion enhancer. AB - An N-terminus sequence of human interleukin 1beta (hIL-1beta) was used as a fusion expression partner for the production of two recombinant therapeutic proteins, human granulocyte-colony stimulating factor (hG-CSF) and human growth hormone (hGH), using Saccharomyces cerevisiae as a host. The expression cassette comprised the leader sequence of killer toxin of Kluyveromyces lactis, the N terminus 24 amino acids (Ser5-Ala28) of mature hIL-1beta, the KEX2 dibasic endopeptidase cleavage site, and the target protein (hG-CSF or hGH). The gene expression was controlled by the inducible UAS(gal)/MF-alpha1 promoter. With the expression vector above, both recombinant proteins were well secreted into culture medium with high secretion efficiencies, and especially, the recombinant hGH was accumulated up to around 1.3 g/L in the culture broth. This is due presumably to the significant role of fused hIL-1beta as secretion enhancer in the yeast secretory pathway. In our recent report, various immunoblotting analyses have shown that the presence of a core N-glycosylation resident in the hIL-1beta fragment is likely to be of crucial importance in the high-level secretion of hG-CSF from the recombinant S. cerevisiae. When the N-glycosylation was completely blocked with the addition of tunicamycin to the culture, the secretion of hG-CSF and hGH was decreased to a negligible level although the other host-derived proteins were well secreted to the culture broth regardless of the presence of tunicamycin. The N-terminal sequencing of the purified hG-CSF verified that the hIL-1beta fusion peptide was correctly removed by in vivo KEX2 protease upon the exit of fusion protein from Golgi complex. From the results presented in this article, it is strongly suggested that the N-terminus fusion of the hIL-1beta peptide could be utilized as a potent secretion enhancer in the expression systems designed for the secretory production of other heterologous proteins from S. cerevisiae. PMID- 10514260 TI - Structured Model-Based Analysis and Control of the Hyaluronic Acid Fermentation by Streptococcus zooepidemicus: Physiological Implications of Glucose and Complex Nitrogen-Limited Growth. AB - The hyaluronic acid (HA) fermentation by Streptococcus zooepidemicus under anaerobic and aerated conditions in glucose-complex media was well described by a structured, two-compartment model. The two-compartment model framework was found to be robust, easily adaptable, and able to predict the transient consumption of substrates and formation of products. Aerobic culture produced a substantially higher concentration of HA than an equivalent anaerobic culture; however biomass specific growth rate and yield were lower due to partial inhibition by hydrogen peroxide. The model was then used to investigate the physiological implications of glucose and complex-nitrogen-limited growth on the anaerobic production of hyaluronic acid (HA). Glucose-limited growth agreed well with model predictions, although the HA molecular weight was lower than expected even though the absolute HA concentration remained unaffected. Heterofermentative growth was also observed for growth rates below 0.1 h(-)(1). Despite a comparatively lower specific growth rate, the biomass yield was higher; however the metabolic shift did not significantly affect HA production. For complex-nitrogen-limited growth, diauxic growth on complex-nitrogen (yeast extract) was observed and explained by partitioning the array of nitrogen components into two distinct but homogeneous pools. While nitrogen-limited growth was found to increase the HA to biomass yield, like that observed under glucose-limited growth, the resulting HA molecular weight was reduced. PMID- 10514259 TI - Biosynthetic burden and plasmid burden limit expression of chromosomally integrated heterologous genes (pdc, adhB) in Escherichia coli. AB - Previous studies have shown an unexpectedly high nutrient requirement for efficient ethanol production by ethanologenic recombinants of Escherichia coli B such as LY01 which contain chromosomally integrated Zymomonas mobilis genes (pdc,adhB) encoding the ethanol pathway. The basis for this requirement has been identified as a media-dependent effect on the expression of the Z. mobilis genes rather than a nutritional limitation. Ethanol production was substantially increased without additional nutrients simply by increasing the level of pyruvate decarboxylase activity. This was accomplished by adding a multicopy plasmid containing pdc alone (but not adhB alone) to strain LY01, and by adding multicopy plasmids which express pdc and adhB from strong promoters. New strong promoters were isolated from random fragments of Z. mobilis DNA and characterized but were not used to construct integrated biocatalysts. These promoters contained regions resembling recognition sites for 3 different E. coli sigma factors: sigma(70), sigma(38), and sigma(28). The most effective plasmid-based promoters for fermentation were recognized by multiple sigma factors, expressed both pdc and adhB at high levels, and produced ethanol efficiently while allowing up to 80% reduction in complex nutrients as compared to LY01. The ability to utilize multiple sigma factors may be advantageous to maintain the high levels of PDC and ADH needed for efficient ethanol production throughout batch fermentation. From this work, we propose that the activation of biosynthetic genes in nutrient-poor media creates a biosynthetic burden that reduces the expression of chromosomal pdc and adhB by competing for transcriptional and translational machinery. This reduced expression can be viewed as analogous to the effect of plasmids (plasmid burden) on the expression of native chromosomal genes. PMID- 10514261 TI - The Effect of Rapeseed Oil Uptake on the Production of Erythromycin and Triketide Lactone by Saccharopolyspora erythraea. AB - Saccharopolyspora erythraea was grown in an oil-based process medium at two different laboratory scales. The initial concentration of rapeseed oil in the medium was shown not to affect the growth, while addition of oil significantly increased erythromycin A production. Increasing the agitation speed at the 2 L scale increased the growth of the culture and the production of erythromycin A but had little effect on the level of oil remaining in the fermentation. Maximum oil utilization of 50% (w/w) was obtained in both 2 and 7 L cultures. Gas chromatography analysis of the process medium showed that there was no accumulation of fatty acids and glycerides during the fermentations. The specific oil utilization by the recombinant strain of S. erythraea was significantly lower compared to the wild-type strain grown at the same scale and initial oil concentration; the recombinant strain also produced lower concentrations of the novel polyketide, triketide lactone. PMID- 10514262 TI - Lipase Production by Acinetobacter radioresistens in the Presence of a Nonwoven Fabric. AB - Lipase production by Acinetobacter radioresistens was performed in a 2-L tank fermentor equipped with a nonwoven fabric, which was made of nylon 6 fiber and coated with a hydrophobic acrylic resin. The fermentation medium contained 2% (v/v) n-hexadecane as the carbon source. The use of the nonwoven fabric was intended for the dispersion of hydrocarbons; thus, the contact surface for the cells to assimilate n-hexadecane can be increased without using emulsifiers. The formation of lipase was found to be growth-associated when the cells grew on n hexadecane. The use of the nonwoven fabric increased the lipase yield by 130%. Further supplementation of 0.1% (v/v) olive oil to the medium could markedly shorten the fermentation time, and thus increase the volumetric productivity. The use of the nonwoven fabric offered two additional advantages in the lipase fermentation: ease of foam control and favorable partition of lipase in the aqueous phase. PMID- 10514263 TI - Estimation of O(2) and CO(2) Solubility in Microbial Culture Media. AB - A simple model for predicting gas solubilities of O(2) and CO(2) at low pressure and near ambient temperature in solutions of salts, sugars, and organic solvents (alcohols, ketones, ethers, aldehydes, etc.) is proposed. It is derived from the Van Laar assumptions and takes account of size differences between molecules in solution by their volumetric fraction. It is a group contribution model where anions and cations are considered as groups and other molecules are treated as in the UNIFAC (UNIQUAC Functional group Activity Coefficients) procedure. Pseudo Henry's constants for groups were determined using solubility data in aqueous solutions containing only one salt, one sugar, or one organic compound at 25 degrees C. The predictive performance of the model was evaluated by comparison with experimental data using multicomponent aqueous salt-sugar mixtures. The model was used to estimate solubilities of oxygen and carbon dioxide in fermentation media. PMID- 10514265 TI - Effect of Resin Characteristics on Fluidized Bed Adsorption of Proteins. AB - Despite numerous advantages, fluidized bed adsorption is limited in commercial applications due to a poor understanding of the relationships between mass transfer, hydrodynamics, and adsorption. To obtain a better understanding of these parameters, we used two commercially available resins to compare the adsorption of lysozyme as a function of bed expansion and bulk-phase viscosity using frontal analysis. Under static conditions adsorption was well represented by a Langmuir isotherm. Dynamic capacities at breakthrough were measured and normalized relative to the equilibrium capacity of each resin to facilitate direct comparison of resin performance under each condition. To quantify the impact of resin characteristics in expanded bed adsorption, we carried out a comparison of mass-transfer effects using a macroporous resin, Streamline SP, and a hyper-diffusive resin, S-HyperD LS. Both resins are designed for fluidized bed adsorption of proteins. In this study the results of frontal analysis showed that breakthrough was due to mass-transfer limitations of the adsorbing particles at expansions of 2 times the settled bed height. At expansions of 3 and 4 times the settled bed height, axial dispersion increased significantly; however mass transfer limitations were still the dominant mechanistic feature contributing to early breakthrough at reduced dynamic capacity. Adsorption capacity under all conditions in this study was poorer for the macroporous resin than for the hyper diffusive resin due to intraparticle mass-transfer limitations. PMID- 10514264 TI - Homologous human blood protein separation using immobilized metal affinity chromatography: protein C separation from prothrombin with application to the separation of factor IX and prothrombin. AB - Protein C (PC) is a natural anticoagulant and antithrombotic present in human blood at a concentration of 4 microg/mL. Its deficiency can result in excessive clotting and thrombosis. Protein C can be obtained from human blood plasma; however, there are other coagulant proteins in blood, including prothrombin (factor II), which is present in relatively large amounts and is one of the most active components. Protein C and prothrombin are homologous proteins with similar biochemical features; therefore, immunoaffinity chromatography is used for their separation. However, this technology is very expensive, protein C recovery and activity is low, and contamination problems with mouse antibody are likely. Immobilized metal affinity chromatography (IMAC) utilizes the protein metal binding properties for protein separation. Protein C has twelve surface accessible histidines, which are the major metal-binding groups for IMAC separation. After investigating metal ion-binding properties of protein C, we used an IDA-Cu column to separate protein C and prothrombin. Following protein adsorption to the column, prothrombin was washed out using a sodium phosphate buffer containing 2 mM imidazole and protein C was recovered with 15 mM imidazole in the buffer. The mild elution condition allows a high protein C activity and a high recovery. Also, this technology introduces no immunoglobulins, and it is relatively inexpensive. IMAC could replace the immunoaffinity technology for the large-scale separation of protein C from blood plasma Cohn Fraction IV-1. In addition, this work demonstrates a significant application of this technology for the separation of factor IX from prothrombin. Prothrombin has proven to be a harmful contaminant in factor IX cocktails that have been administered to humans in the treatment of hemophilia B. PMID- 10514266 TI - Production of retroviral vectors for gene therapy with the human packaging cell line FLYRD18. AB - The development of gene therapy is hampered by the difficulty of producing large stocks of retroviral vectors at high titer. This study aimed to improve culture conditions and to intensify the production of retroviruses by FLYRD18, a packaging cell line derived from the HT1080 human fibrosarcoma line. Batch virus production proved to be feasible in unsupplemented basal medium and provided significantly higher titers and productivities than medium supplemented with 10% serum. For longer-term production, however, AIM-V complete serum-free medium and basal medium supplemented with 2% serum gave superior results. Serum supplementation should nevertheless be optimized to take into account the presence of inhibitors of viral production. In monolayer cultures with 0.2 mL/cm(2), the cell concentration was increased up to 2 x 10(6) cells/mL without loss of cell productivity. A semicontinuous production process, which enables the collection of larger amounts of viruses from the same culture, has also been successfully used. Suspension culture processes were prevented by the anchorage dependency of the FLYRD18 cell line. Microcarrier cultures were able to produce viruses but will require further investigation and optimization for their performance to become competitive with monolayer cultures. In the course of this study, more than a 10-fold increase of titer has been achieved. PMID- 10514267 TI - Simultaneous Optimization of Feeding Rate and Operation Parameters for Fed-Batch Fermentation Processes. AB - An efficient method is introduced for simultaneously determining optimal polices of feeding rate and operation parameters for a fermentation process. Such an optimization problem is converted into the finite dimensional optimization problem using the control parametrization technique. The hybrid differential evolution is introduced to solve the converted problem. The optimal production rate obtained by the simultaneous optimization approach could be significantly improved with comparison to a simplified optimization problem, which is considered the optimal feed control only, as observed from the simulation results. PMID- 10514268 TI - Optimization of sorting conditions for the selection of stable, high-producing mammalian cell lines. AB - The production of Green Fluorescent Protein in recombinant NIH3T3 mouse fibroblast cells was used as a model to determine the optimal conditions for the rapid isolation of high-producing cell lines with a fluorescence-activated cell sorter. "Bulk sorting", that is, sorting of a large number of positive cells, did not result in a stable, high-producing cell line due to overgrowth of high producing cells by low- or nonproducing cells. The production kinetics and expression of GFP during batch culture was found to differ between NIH3T3 cells and HepG2 hepatoma cells, even though the same plasmid was used for transfection. The kinetics of product formation need therefore to be determined from case to case to select the optimal timepoint for analysis and sorting. Subcloning of sorted cells into microtiter plates only resulted in high-producing subclones when 1 or 2 cells were seeded per well. Higher seeding rates again resulted in overgrowth of low- or nonproducers. By subcloning, two high-producing cells lines could be isolated. They had a 10- and 15-fold higher fluorescent signal compared to the negative control. While one of these subclones started to decrease it's GFP expression after 2 months, the other clone stably expressed GFP for 4 months. PMID- 10514270 TI - 2000-2001 predoctoral fellowships PMID- 10514269 TI - Stress Survival of a Genetically Engineered Pseudomonas in Soil Slurries: Cytochrome P-450cam-Catalyzed Dehalogenation of Chlorinated Hydrocarbons. AB - Biological treatment of hazardous chemical wastes has potential as an effective, practical, and economically viable process in above the ground treatment systems that consist of both genetically engineered microorganisms (GEMs) and bioreactors with process control instruments to create ideal conditions for biodegradation. A strain of Pseudomonas putida coexpressing cytochrome P-450cam and luciferase (lux) that provides both the reductive detoxification potential of the hemoprotein and a mechanism for its reduction in the absence of "normal" P-450 redox partners was evaluated for its ability to survive and remain metabolically competent under nutrient stress in soil slurry microcosms. More than 74% of the cells of engineered Pseudomonas were culturable after 7 days of multiple nutrient (C,N,P) starvation. The diagnostic luminescence and carbon monoxide-difference spectra for the two engineered traits could be detected in a significant fraction of the surviving population. The GEM could be revived after repeated desiccation and starvation using Luria broth, benzoate, or citrate as nutrients. Soil slurries inoculated with the GEM transformed hexachloroethane (HCE) to tetrachloroethylene (tetraCE) 8-10-fold faster than uninoculated slurries. The GEM also transformed the insecticide, gamma-HCH (gamma-3,4,5,6 hexachlorocyclhexene), to gamma-3,4,5,6-tetrachlorocyclohexene (gammatetraCH) in soil slurries under subatmospheric conditions. These results indicate that GEMs can be constructed with broad substrate range detoxification catalysts such as cytochrome P-450 for remediation. PMID- 10514271 TI - Diethylcarbamoylating/nitroxylating agents as dual action inhibitors of aldehyde dehydrogenase: a disulfiram-cyanamide merger. AB - Benzenesulfohydroxamic acid (Piloty's acid) was functionalized on the hydroxyl group with the N,N-diethylcarbamoyl group, and the hydroxylamine nitrogen was substituted with acetyl (1a), pivaloyl (1b), benzoyl (1c), and ethoxycarbonyl (1d) groups. Only compound 1d inhibited yeast aldehyde dehydrogenase (AlDH) in vitro (IC(50) 169 microM). When administered to rats, 1d significantly raised blood acetaldehyde levels following ethanol challenge, thus serving as a diethylcarbamoylating/nitroxylating, dual action inhibitor of AlDH in vivo. A more potent dual action agent was N-(N, N-diethylcarbamoyl)-O methylbenzenesulfohydroxamic acid (5c), which was postulated to release diethylcarbamoylnitroxyl (9), a highly potent diethylcarbamoylating/nitroxylating agent, following metabolic O-demethylation in vivo. The dual action inhibition of AlDH exhibited by 1d, and especially 9, constitutes a merger of the mechanism of action of the alcohol deterrent agents, disulfiram and cyanamide. PMID- 10514272 TI - High-resolution NMR and computer modeling studies of the cannabimimetic aminoalkylindole prototype WIN-55212-2. AB - Aminoalkylindoles (AAIs), although structurally dissimilar from the classical cannabinoids (CCs), are known to be capable of binding to cannabinoid receptors and of evoking cannabimimetic responses. However, their mode of binding remains unknown. In this communication, we have carried out further studies on the AAI prototype (R)-[2, 3-dihydro-5-methyl-3-[(4-morpholinyl)methyl]pyrrolo[1,2,3-de] 1, 4-benzoxazin-6-yl](1-naphthalenyl)methanone (WIN-55212-2, 1) by the combined use of high-resolution 2D NMR and computer modeling. Our results suggest that the minimum energy conformations of the molecule 1 have distinct pharmacophoric features: (i) The naphthyl ring is oriented off the plane of the benzoxazine ring by approximately 59 degrees with the carbonyl C=O group pointing toward the C2 CH(3) group. (ii) At the C10-position the axial morpholinomethyl conformation is preferred over the equatorial in order to relieve a steric interaction with the C2-methyl group. The preferred conformer as defined by the three key pharmacophores, naphthyl, morpholino, and 3-keto groups, shows that the morpholinyl ring of the molecule 1 deviates from the plane of the benzoxazine ring by about 32 degrees and orients in the left molecular quadrant. This model supports the hypothesis that a certain deviation of the morpholino group from the plane of the indole ring in compound 1 is essential for cannabimimetic activity. We postulate that such an alignment by the respective pharmacophores allows them to interact optimally with the receptor. The results should help us to better understand the pharmacophoric requirements of the AAIs and serve as a basis for future SAR studies and drug design. PMID- 10514273 TI - Design, synthesis, and evaluation of a novel sequence-selective epoxide containing DNA cross-linking agent based on the pyrrolo[2, 1 c][1,4]benzodiazepine system. AB - Synthetic routes have been investigated to prepare a novel C8-epoxide functionalized pyrrolo[2,1-c][1,4]benzodiazepine 6 as a potential sequence selective DNA cross-linking agent (Wilson et al. Tetrahedron Lett. 1995, 36, 6333 6336). A successful synthesis was accomplished via a 10-step route involving a pro-N10-Fmoc cleavage method that should have general applicability to other pyrrolobenzodiazepine (PBD) molecules containing acid- or nucleophile-sensitive groups. During the course of this work, a one-pot reductive cyclization procedure for the synthesis of PBD N10-C11 imines from nitro dimethyl acetals was also discovered, although this method results in C11a racemization which can reduce DNA binding affinity and cytotoxicity. The target epoxide 6 was shown by thermal denaturation studies to have a significantly higher DNA-binding affinity than the parent DC-81 (3) or the C8-propenoxy-PBD (15), which is structurally similar but lacks the epoxide moiety. The time course of effects upon thermal denaturation indicated a rapid initial binding phase followed by a slower phase consistent with the stepwise cross-linking of DNA observed for a difunctional agent. This was confirmed by an electrophoretic assay which demonstrated efficient induction of interstrand cross-links in plasmid DNA at concentrations >1 microM. Higher levels of interstrand cross-linking were observed at 24 h compared to 6 h incubation. A Taq polymerase stop assay indicated a preference for binding to guanine-rich sequences as predicted for bis-alkylation in the minor groove of DNA by epoxide and imine moieties. The pattern of stop sites could be partly rationalized by molecular modeling studies which suggested low-energy models to account for the observed binding behavior. The epoxide PBD 6 was shown to have significant cytotoxicity (45-60 nM) in the A2780, CH1, and CH1cis(R) human ovarian carcinoma cell lines and an IC(50) of 0.2 microM in A2780cis(R). The significant activity of 6 in the cisplatin-resistant CH1cis(R) cell line (IC(50) = 47 nM) gave a resistance factor of 0.8 compared to the parent cell line, demonstrating no cross-resistance with the major groove cross-linking agent cisplatin. PMID- 10514274 TI - Synthesis and antiviral activity of ethidium-arginine conjugates directed against the TAR RNA of HIV-1. AB - The regulatory protein Tat is essential for viral gene expression and replication of the human immunodeficiency virus type 1 (HIV-1). Tat transactivates the HIV-1 long terminal repeat (LTR) via its binding to the transactivation responsive element (TAR) and increases the viral transcription. Studies have shown that the binding of arginine and arginine derivatives induces a conformational change of the TAR RNA at the Tat-binding site. The unpaired A17 residue delimits a small cavity which constitutes a receptor site for small molecules, especially for ethidium bromide. These binding characteristics have prompted us to design a series of ethidium-arginine conjugates capable of interacting with the TAR RNA. Here we report the synthesis of six ethidium derivatives equipped with arginine side chains. These molecules were biologically evaluated, and two compounds (17 and 20) exhibited in vitro anti-HIV-1 activity at micromolar concentration, without toxicity (up to 100 microM concentration). Melting temperature studies indicated that the most active molecule (20) bound strongly to TAR in vitro. RNase protection experiments agreed with the molecular modeling studies which suggested that the ethidium moiety of 20 was inserted next to the A17 residue while the arginine side chain occupied the pyrimidine bulge. PMID- 10514275 TI - Inhibitors of the C(2)-symmetric HIV-1 protease: nonsymmetric binding of a symmetric cyclic sulfamide with ketoxime groups in the P2/P2' side chains. AB - Symmetric cyclic sulfamides, substituted in the P2/P2' position with functional groups foreseen to bind preferentially to the S2/S2' subsites of HIV-1 protease, have been prepared. Despite efforts to promote a symmetric binding, the sulfamides seemed prone to bind nonsymmetrically, as deduced from X-ray crystal structure analysis of one of the most potent inhibitors, possessing ketoxime groups in the P2/P2' side chains. Ab initio calculations suggested that the nonsymmetric conformation of the cyclic sulfamide scaffold had lower energy than the corresponding symmetric, cyclic urea-like conformation. PMID- 10514276 TI - A novel approach to predicting P450 mediated drug metabolism. CYP2D6 catalyzed N dealkylation reactions and qualitative metabolite predictions using a combined protein and pharmacophore model for CYP2D6. AB - A combined protein and pharmacophore model for cytochrome P450 2D6 (CYP2D6) has been extended with a second pharmacophore in order to explain CYP2D6 catalyzed N dealkylation reactions. A group of 14 experimentally verified N-dealkylation reactions form the basis of this second pharmacophore. The combined model can now accommodate both the usual hydroxylation and O-demethylation reactions catalyzed by CYP2D6, as well as the less common N-dealkylation reactions. The combined model now contains 72 metabolic pathways catalyzed by CYP2D6 in 51 substrates. The model was then used to predict the involvement of CYP2D6 in the metabolism of a "test set" of seven compounds. Molecular orbital calculations were used to suggest energetically favorable sites of metabolism, which were then examined using modeling techniques. The combined model correctly predicted 6 of the 8 observed metabolites. For the well-established CYP2D6 metabolic routes, the predictive value of the current combined protein and pharmacophore model is good. Except for the highly unusual metabolism of procainamide and ritonavir, the known metabolites not included in the development of the model were all predicted by the current model. Two possible metabolites have been predicted by the current model, which have not been detected experimentally. In these cases, the model may be able to guide experiments. P450 models, like the one presented here, have wide applications in the drug design process which will contribute to the prediction and elimination of polymorphic metabolism and drug-drug interactions. PMID- 10514277 TI - Bioreductive activation of a series of indolequinones by human DT-diaphorase: structure-activity relationships. AB - A series of indolequinones including derivatives of EO9 bearing various functional groups and related indole-2-carboxamides have been studied with a view to identifying molecular features which confer substrate specificity for purified human NAD(P)H:quinone oxidoreductase (DT-diaphorase), bioreductive activation to DNA-damaging species, and selectivity for DT-diaphorase-rich cells in vitro. A broad spectrum of substrate specificity exists, but minor changes to the indolequinone nucleus have a significant effect upon substrate specificity. Modifications at the 2-position are favorable in terms of substrate specificity as these positions are located at the binding site entrance as determined by molecular modeling studies. In contrast, substitutions at the (indol-3-yl)methyl position with bulky leaving groups or a group containing a chlorine atom result in compounds which are poor substrates, some of which inactivate DT-diaphorase. Modeling studies demonstrate that these groups sit close to the mechanistically important amino acids Tyr 156 and His 162 possibly resulting in either alkylation within the active site or disruption of charge-relay mechanisms. An aziridinyl group at the 5-position is essential for potency and selectivity to DT-diaphorase rich cells under aerobic conditions. The most efficient substrates induced qualitatively greater single-strand DNA breaks in cell-free assays via a redox mechanism involving the production of hydrogen peroxide (catalase inhibitable). This damage is unlikely to form a major part of their mechanism of action in cells since potency does not correlate with extent of DNA damage. In terms of hypoxia selectivity, modifications at the 3-position generate compounds which are poor substrates for DT-diaphorase but have high hypoxic cytotoxicity ratios. PMID- 10514278 TI - Antitumor agents. 196. Substituted 2-thienyl-1,8-naphthyridin-4-ones: their synthesis, cytotoxicity, and inhibition of tubulin polymerization. AB - As part of our continuing search for potential anticancer drug candidates in the 2-aryl-1,8-naphthyridin-4-one series, we have synthesized a series of substituted 2-thienyl-1, 8-naphthyridin-4-ones. Most compounds showed significant cytotoxic effects (log GI(50) < -4.0; log molar drug concentration required to cause 50% growth inhibition) against a variety of human tumor cell lines in the National Cancer Institute's in vitro screen, including cells derived from solid tumors such as non-small-cell lung, colon, central nervous system, melanoma, ovarian, prostate, and breast cancers. The most active compounds (31-33,40) demonstrated strong cytotoxic effects with ED(50) values in the micromolar or submicromolar range in most of the tumor cell lines. The most cytotoxic compounds inhibited tubulin polymerization at concentrations substoichiometric to the tubulin concentration. The most potent inhibitors of polymerization (40,42,43) had effects comparable to those of the potent antimitotic natural products podophyllotoxin and combretastatin A-4 and to that of NSC 664171, a particularly potent, structurally related analogue. Only compound 40 was a potent inhibitor of the binding of radiolabeled colchicine to tubulin, and it was both the most cytotoxic agent and the most effective inhibitor of polymerization among the newly synthesized compounds. PMID- 10514279 TI - Discovery of potent and selective SH2 inhibitors of the tyrosine kinase ZAP-70. AB - A series of 1,2,4-oxadiazole analogues has been shown to be potent and selective SH2 inhibitors of the tyrosine kinase ZAP-70, a potential therapeutic target for immune suppression. These compounds typically are 200-400-fold more potent than the native, monophosphorylated tetrapeptide sequences. When compared with the high-affinity zeta-1-ITAM peptide (Ac-NQL-pYNELNLGRREE-pYDVLD-NH(2), wherein pY refers to phosphotyrosine) some of the best 1,2, 4-oxadiazole analogues are approximately 1 order of magnitude less active. This series of compounds displays an unprecedented level of selectivity over the closely related tyrosine kinase Syk, as well as other SH2-containing proteins such as Src and Grb2. Gel shift studies using a protein construct consisting only of C-terminal ZAP-70 SH2 demonstrate that these compounds can effectively engage this particular SH2 domain. PMID- 10514280 TI - Synthesis and enantiopharmacology of new AMPA-kainate receptor agonists. AB - Regioisomeric 3-carboxyisoxazolinyl prolines [CIP-A (+/-)-6 and CIP-B (+/-)-7] and 3-hydroxyisoxazolinyl prolines [(+/-)-8 and (+/-)-9] were synthesized and assayed for glutamate receptor activity. The tests were carried out in vitro by means of receptor binding techniques, second messenger assays, and the rat cortical wedge preparation. CIP-A showed a good affinity for both 2-amino-3-(3 hydroxy-5-methylisoxazol-4-yl)propionic acid (AMPA) and kainic acid (KAIN) receptors. These results were confirmed in the cortical slice model where CIP-A displayed an EC(50) value very close to that of AMPA. The convulsant properties of all the compounds were evaluated in vivo on DBA/2 mice after icv injection. CIP-A showed a convulsant activity, measured as tonus and clonus seizures, 18-65 times higher than that produced by AMPA. It was also quite active after ip administration, since it induced seizures in mice at doses as low as 3.2 nmol/mouse. On the basis of the above-reported results we prepared and tested the enantiomers of CIP-A and CIP-B, obtained by reacting (S)-3,4-didehydroproline and (R)-3,4-didehydroproline, respectively, with ethoxycarbonylformonitrile oxide. In all the tests the S-form, CIP-AS [(-)-6], emerged as the eutomer evidencing common stereochemical requirements with the reference compounds AMPA and KAIN. Through modeling studies, carried out on CIP-A, AMPA, and KAIN, active conformations for CIP-AS and AMPA at AMPA receptors as well as for CIP-AS and KAIN at KAIN receptors are suggested. PMID- 10514281 TI - Inhibition of neuronal nitric oxide synthase by 4-amino pteridine derivatives: structure-activity relationship of antagonists of (6R)-5,6,7,8 tetrahydrobiopterin cofactor. AB - The family of nitric oxide synthases (NOS) catalyzes the conversion of L-arginine to L-citrulline and nitric oxide (NO), an important cellular messenger molecule which has been implicated in the pathophysiology of septic shock and inflammatory and neurodegenerative disease states. NOS can be maximally activated by the ubiquitous cofactor, (6R)-5,6,7,8-tetrahydrobiopterin (H(4)Bip), and antagonists of H(4)Bip may be of therapeutic importance to inhibit pathologically high NO formation. The 4-amino substituted analogue of H(4)Bip was reported to be a potent NOS inhibitor. Therefore, we developed a series of novel 4-amino pteridine derivatives, anti-pterins, to pharmacologically target the neuronal isoform of nitric oxide synthase (NOS-I). To functionally characterize the pterin/anti pterin interaction and establish a structure-activity relationship (SAR), we systematically altered the substituents in the 2-, 4-, 5-, 6-, and 7-position of the pteridine nucleus. Varying the substitution pattern in the 2-, 5-, and 7 position resulted in no significant inhibitory effect on enzyme activity. In contrast, bulky substituents in the 6-position, such as phenyl, markedly increased the inhibitory potency of the reduced 4-amino-5,6,7,8 tetrahydropteridines, possibly as a consequence of hydrophobic interactions within NOS-I. However, this was not the case for the aromatic 4-amino pteridines. Interestingly, chemical modification of the 4-amino substituent by dialkyl/diaralkylation together with 6-arylation of the aromatic 2,4-diamino pteridine resulted in potent and efficacious inhibitors of NOS-I, suggesting possible hydrophilic and hydrophobic interactions within NOS-I. This SAR agrees with (a) the recently published crystal structure of the oxygenase domain of the inducible NOS isoform (NOS-II) and (b) the comparative molecular field analysis of selected NOS-I inhibitors, which resulted in a 3D-QSAR model of the pterin binding site interactions. Further optimization should be possible when the full length structure of NOS-I becomes available. PMID- 10514282 TI - Design and synthesis of lipophilic phosphoramidate d4T-MP prodrugs expressing high potency against HIV in cell culture: structural determinants for in vitro activity and QSAR. AB - A series of new substituted-aryl phosphoramidate derivatives of the anti-HIV drug d4T were synthesized as membrane-soluble nucleotide prodrugs, to extend and quantify the SAR observed for an earlier series of related derivatives. All of the compounds were found to be significantly more potent against HIV in cell culture than the nucleoside analogue d4T, and most were also found to be significantly more potent than the parent phosphoramidate. A Hansch type QSAR analysis was applied to the combined series of 21 compounds. The results of this analysis revealed anti-HIV activity to be principally dependent on lipophilicity in a quadratic manner, with terms representing substituent steric bulk and electronic effects having a minimal significance. PMID- 10514283 TI - Protein kinase C ligands based on tetrahydrofuran templates containing a new set of phorbol ester pharmacophores. AB - A series of substituted tetrahydrofurans with an embedded glycerol backbone carrying additional tetrahydrofuranylideneacetate or tetrahydrofuranylacetate motifs were grouped into four distinct templates (I-IV) according to stereochemistry. The compounds were designed to mimic three essential pharmacophores (C(3)-C=O, C(20)-OH and C(13)-C=O) of the phorbol esters according to a new, revised model. The tetrahydrofuran ring was constructed from glycidyl 4 methoxyphenyl ether, and the structures of the isomeric templates were assigned by NMR spectroscopy, including NOE. The binding affinity for protein kinase C (PKC) was assessed in terms of the ability of the ligands to displace bound [(3)H 20]phorbol 12, 13-dibutyrate (PDBU) from a recombinant alpha isozyme of PKC. Geometric Z- and E-isomers (1 and 3, respectively) containing a tetrahydrofuranylideneacetate motif were the most potent ligands with identical K(i) values of 0.35 microM. Molecular modeling studies of the four templates showed that the rms values when fitted to a prototypical phorbol 12,13-diacetate ester correlated inversely with affinities in the following order: I approximately II > III > IV. These compounds represent the first generation of rigid glycerol templates seeking to mimic the binding of the C(13)-C=O of the phorbol esters. The binding affinities of the most potent compounds are in the same range of the diacylglycerols (DAGs) despite the lack of a phorbol ester C(9) OH pharmacophore surrogate. This finding confirms that mimicking the binding of the C(13)-C=O pharmacophore of phorbol is a useful strategy. However, since the C(9)-OH and C(13)-C=O in the phorbol esters appear to form an intramolecular hydrogen bond that functions as a combined pharmacophore, it is possible the lack of this combined motif in the target templates restricts the compounds from reaching higher binding affinities. PMID- 10514284 TI - Synthesis and structure-activity relationships of the (alkylamino)piperidine containing BHAP class of non-nucleoside reverse transcriptase inhibitors: effect of 3-alkylpyridine ring substitution. AB - Development of resistance to currently approved HIV therapies has continued to fuel research efforts to improve the metabolic stability and spectrum of activity of the (alkylamino)piperidine-containing bis(heteroaryl)piperazine (AAP-BHAP) class of non-nucleoside reverse transcriptase inhibitors (NNRTIs). The synthesis of analogues in which the usual 3-alkylamino substituent on the pyridine ring is replaced by a 3-alkyl substituent led to compounds which retained activity against recombinant P236L and wild-type (WT) reverse transcriptase (RT), while inhibition of the Y181C mutant RT was reduced relative to the activity of the 3 alkylamino-substituted congeners. Testing of representative analogues in an in vitro liver microsome assay indicated that the alkyl substituent would not appreciably improve the metabolic stability of the AAP-BHAP template. In vivo pharmacokinetic evaluation of three compounds confirmed these results in that high systemic clearances were observed. Nevertheless, one compound (13), PNU 103657, possessed oral bioavailability in rats approaching that of the structurally related NNRTI drug delavirdine which is currently on the market for the treatment of HIV infection. PMID- 10514285 TI - Urea-PETT compounds as a new class of HIV-1 reverse transcriptase inhibitors. 3. Synthesis and further structure-activity relationship studies of PETT analogues. AB - The further development of allosteric HIV-1 RT inhibitors in the urea analogue series of PETT (phenylethylthiazolylthiourea) derivatives is described here. The series includes derivatives with an ethyl linker (1-5) and racemic (6-16) and enantiomeric (17-20) cis-cyclopropane compounds. The antiviral activity was determined both at the RT level and in cell culture on both wild-type and mutant forms of HIV-1. Most compounds have anti-HIV-1 activity on the wt in the nanomolar range. Resistant HIV-1 was selected in vitro for some of the compounds, and the time for resistant HIV-1 to develop was longer for urea-PETT compounds than it was for reference compounds. Preliminary pharmacokinetics in rats showed that compound 18 is orally bioavailable and penetrates well into the brain. The three-dimensional structure of complexes between HIV-1 RT and two enantiomeric compounds (17 and 18) have been determined. The structures show similar binding in the NNI binding pocket. The propionylphenyl moieties of both inhibitors show perfect stacking to tyrosine residues 181 and 188. The cyclopropyl moiety of the (+)-enantiomer 18 exhibits optimal packing distances for the interactions with leucine residue 100 and valine residue 179. PMID- 10514286 TI - 6-Substituted 2-oxo-2H-1-benzopyran-3-carboxylic acid as a core structure for specific inhibitors of human leukocyte elastase. AB - Pyridyl esters of 6-substituted 2-oxo-2H-1-benzopyran-3-carboxylic acid were designed as mechanism-based inhibitors of human leukocyte elastase. Compounds of series 4 specifically inhibited this enzyme. Several of the tested compounds (series 2 and 3) acted as powerful time-dependent inhibitors of both human leukocyte elastase and alpha-chymotrypsin; some compounds of these series inhibited thrombin. Trypsin was not inhibited. A transient inactivation was observed for human leukocyte elastase (k(i)/K(I) = 107 000 M(-1). s(-1) for 4c) and thrombin (k(i)/K(I) = 7 200 M(-1).s(-1) for 3b) as demonstrated by spontaneous or hydroxylamine-accelerated reactivation, irrespective of the nature of the substituent at the 6-position. Conversely, alpha-chymotrypsin was irreversibly inhibited by 6-chloromethyl derivatives (k(i)/K(I) = 107 400 M(-1). s(-1) for 3b). The presence of a latent alkylating function at the 6-position (chloromethyl group) was required for leading to this inactivation. In the absence of such an alkylating function (series 4), human leukocyte elastase was specifically inhibited suggesting that this new series of human leukocyte elastase inhibitors may be of potential therapeutic interest in degradative and degenerative processes involving this enzyme. PMID- 10514287 TI - Antitumor benzothiazoles. 8. Synthesis, metabolic formation, and biological properties of the C- and N-oxidation products of antitumor 2-(4 aminophenyl)benzothiazoles. AB - 2-(4-Aminophenyl)benzothiazoles 1 and their N-acetylated forms have been converted to C- and N-hydroxylated derivatives to investigate the role of metabolic oxidation in the mode of action of this series of compounds. 2-(4-Amino 3-methylphenyl)benzothiazole (1a, DF 203, NSC 674495) is a novel and potent antitumor agent with selective growth inhibitory properties against human cancer cell lines. Very low IC(50) values (<0.1 microM) were encountered in the most sensitive breast cancer cell lines, MCF-7 and T-47D, whereas renal cell line TK 10 was weakly inhibited by 1a. Cell lines from the same tissue origin, MDA-MB-435 (breast), CAKI-1 (renal), and A498 (renal), were insensitive to 1a. Accumulation and metabolism of 1a were observed in sensitive cell lines only, with the highest rate of metabolism occurring in the most sensitive MCF-7 and T-47D cells. Thus, differential uptake and metabolism of 1a by cancer cell lines may underlie its selective profile of anticancer activity. A major metabolite in these sensitive cell lines has been identified as 2-(4-amino-3-methylphenyl)-6 hydroxybenzothiazole (6c). Hydroxylation of 1a was not detected in the homogenate of previously untreated MCF-7, T-47D, and TK-10 cells but was readily observed in homogenates of sensitive cells that were pretreated with 1a. Accumulation and covalent binding of [(14)C]1a derived radioactivity was observed in the sensitive MCF-7 cell line but not in the insensitive MDA-MB-435 cell line. The mechanism of growth inhibition by 1a, which is unknown, may be dependent on the differential metabolism of the drug to an activated form by sensitive cell lines only and its covalent binding to an intracellular protein. However, the 6-hydroxy derivative 6c is not the 'active' metabolite since, like all other C- and N-hydroxylated benzothiazoles examined in this study, it is devoid of antitumor properties in vitro. PMID- 10514288 TI - Design and synthesis of potent bradykinin agonists containing a benzothiazepine moiety. AB - A bradykinin analogue (H-Arg-Pro-Pro-Gly-Phe-Ser-D-BT-Arg-OH, 3) in which the Pro Phe dipeptide was replaced by the (3S)[amino]-5-(carbonylmethyl)-2,3-dihydro-1, 5 benzothiazepin-4(5H)-one (D-BT) moiety has been synthesized. The same modification was performed on the potent bradykinin B(2) receptor antagonist HOE 140 (H-D-Arg-Arg-Pro-Hyp-Gly-Thi-Ser-D-Tic-Oic-Arg-OH), in which the -D-Tic-Oic- moiety was replaced by D-BT to yield H-D-Arg-Arg-Pro-Hyp-Gly-Thi-Ser-D-BT-Arg-OH, 1 (JMV1116). These compounds were examined in vitro for their binding affinity toward bradykinin B(1) and B(2) receptors as well as for their ability to interfere with bradykinin-induced contraction of both human umbilical vein and rat uterus. The two compounds 3 and 1 competed with [(3)H]bradykinin binding to the human cloned B(2) receptor giving K(i) values of 13 +/- 2 and 0.7 +/- 0.1 nM, respectively. Unexpectedly, both compounds were full bradykinin B(2) receptor agonists on the human umbilical vein (pD(2) = 6.60 +/- 0.07 for 3 and 6.80 +/- 0.08 for 1) and rat uterus (pD(2) = 7.20 +/- 0.09 for 3 and 7.50 +/- 0.09 for 1) preparations with the same efficacy as bradykinin. In addition 1 induced a concentration-dependent phosphoinositide production in CHO cells expressing the human cloned B(2) receptor. These data provide evidence for a bioactive conformation of bradykinin constrained at the dipeptide Pro-Phe. PMID- 10514289 TI - Synthesis and characterization of bradykinin B(2) receptor agonists containing constrained dipeptide mimics. AB - We have previously shown that substitution of the D-Tic-Oic dipeptide by a (3S) [amino]-5-(carbonylmethyl)-2,3-dihydro-1, 5-benzothiazepin-4(5H)-one (D-BT) moiety in the bradykinin B(2) receptor antagonist HOE 140 resulted in a full potent and selective bradykinin B(2) receptor agonist (H-DArg-Arg-Pro-Hyp-Gly-Thi Ser-D-BT-Arg-OH, JMV1116) exhibiting a high affinity for the human receptor (K(i) 0.7 nM). In the present study, we have investigated the effects of replacement of the D-Tic-Oic moiety by various constrained dipeptide mimetics. The resulting compounds were tested for their binding affinity toward the cloned human B(2) receptor and for their functional interaction with the bradykinin-induced contraction of isolated human umbilical vein. Subsequently, we have designed novel bradykinin B(2) receptor agonists which are likely to be resistant to enzymatic cleavage by endopeptidases and which might represent interesting new pharmacological tools. In an attempt to increase the potency of compound JMV1116, both its N-terminal part and the D-BT moiety were modified. Substitution of the D arginine residue by a L-lysine residue led to a 10-fold more potent bradykinin B(2) ligand [compound 22 (JMV1465) (K(i) 0.07 nM)], retaining full agonist activity on human umbilical vein. Substitution of the D-BT moiety by a (3S) [amino]-5-(carbonylmethyl)-2,3-dihydro-8-methyl-1, 5-benzothiazepin-4(5H)-one [D BT(Me)] moiety led to compound 23 (JMV1609) which exhibited a higher agonist activity (pD(2) = 7.4) than JMV1116 (pD(2) = 6.8). PMID- 10514290 TI - Synthesis and antimicrobial activity of 4H-4-oxoquinolizine derivatives: consequences of structural modification at the C-8 position. AB - The antibacterial 4H-4-oxoquinolizines were introduced recently to overcome bacterial resistance to fluoroquinolones. They exhibit potent antibacterial activity against Gram-positive, Gram-negative, and anaerobic organisms and are highly active against some quinolone-resistant bacteria including quinolone resistant MRSA. Preliminary studies indicated that oxoquinolizines possess distinct activity and toxicity profiles as compared with their parent quinolones. In order to develop a potent antibacterial agent with the desired spectrum of activity, good tolerability, and balanced pharmacokinetic profile, we synthesized and evaluated a series of oxoquinolizines with various substituents at the C-8 position. Most compounds tested in this study demonstrated better activity against Gram-positive bacteria than ciprofloxacin and exhibited good susceptibility against ciprofloxacin- and methicillin-resistant S. aureus. While maintaining potent in vitro activity, several compounds showed improved in vivo efficacy over ABT-719 as indicated by the mouse protection test. As an example, the oral ED(50) values for the cis-3-amino-4-methylpiperidine analogue 3ss against S. aureus NCTC 10649M, S. pneumoniae ATCC 6303, and E. coli JUHL were 0. 8, 2.0, and 1.4 mg/kg, compared to 3.0, 10.0, and 8.3 mg/kg for ABT-719. The current study revealed that the steric and electronic environment, conformation, and absolute stereochemistry of the C-8 group are very important to the antibacterial profiles. Structural modifications of the C-8 group provide a useful means to improve the antibacterial activities, physicochemical properties, and pharmacokinetic profiles. Manipulation of the C-8 group also allows us to generate analogues with the desired spectrum of activity, such as analogues that are selective against respiratory pathogens. PMID- 10514291 TI - WB 4101-related compounds. 2. Role of the ethylene chain separating amine and phenoxy units on the affinity for alpha(1)-adrenoreceptor subtypes and 5-HT(1A) receptors. AB - WB 4101 (1)-related benzodioxanes were synthesized by replacing the ethylene chain separating the amine and the phenoxy units of 1 with a cyclopentanol moiety, a feature of 6, 7-dihydro-5-[[(cis-2-hydroxy-trans-3 phenoxycyclopentyl)amino]meth yl] -2-methylbenzo[b]thiophen-4(5H)-one that was reported to display an intriguing selectivity profile at alpha(1) adrenoreceptors. This synthesis strategy led to 4 out of 16 possible stereoisomers, which were isolated in the case of (-)-3, (+)-3, (-)-4, and (+)-4 and whose absolute configuration was assigned using a chiral building block for the synthesis of (-)-3 starting from (+)-(2R)-2, 3-dihydro-1,4-benzodioxine-2 carboxylic acid ((+)-9) and (1S,2S, 5S)-2-amino-5-phenoxycyclopentan-1-ol ((+) 10). The aim of this project was to further investigate whether it is possible to differentiate between these compounds with respect to their affinity for alpha(1) adrenoreceptor subtypes and the affinity for 5-HT(1A) receptors, as 1 binds with high affinity at both receptor systems. The biological profiles of reported compounds at alpha(1)-adrenoreceptor subtypes were assessed by functional experiments in isolated rat vas deferens (alpha(1A)), spleen (alpha(1B)), and aorta (alpha(1D)) and by binding assays in CHO and HeLa cells membranes expressing the human cloned alpha(1)-adrenoreceptor subtypes and 5-HT(1A) receptors, respectively. Furthermore, the functional activity of (-)-3, (+)-3, ( )-4, and (+)-4 toward 5-HT(1A) receptors was evaluated by determining the induced stimulation of [(35)S]GTPgammaS binding in cell membranes from HeLa cells transfected with human cloned 5-HT(1A) receptors. The configuration of the cyclopentane unit determined the affinity profile: a 1R configuration, as in (+) 3 and (-)-4, conferred higher affinity at alpha(1)-adrenoreceptors, whereas a 1S configuration, as in (-)-3 and (+)-4, produced higher affinity for 5-HT(1A) receptors. For the enantiomers (+)-4 and (-)-4 also a remarkable selectivity was achieved. Functionally, the stereoisomers displayed a similar alpha(1) selectivity profile, that is alpha(1D) > alpha(1B) > alpha(1A), which is different from that exhibited by the reference compound 1. The epimers (-)-3 and (+)-4 proved to be agonists at the 5-HT(1A) receptors, with a potency comparable to that of 5-hydroxytryptamine. PMID- 10514292 TI - Heterodimeric tacrine-based acetylcholinesterase inhibitors: investigating ligand peripheral site interactions. AB - Dimeric acetylcholinesterase (AChE) inhibitors containing a single 9-amino 1,2,3,4-tetrahydroacridine (tacrine) unit were constructed in an effort to further delineate structural requirements for optimal binding to the AChE peripheral site. Basic amines of differing hydrophobicity were selected as peripheral site ligands, and in each case, improvements in inhibitory potency and selectivity were seen relative to tacrine itself. AChE IC(50) values of the optimum dimers decrease significantly as the peripheral site ligand was permuted in the series ammonia > dimethylamine > 4-aminopyridine > 4-aminoquinoline > tacrine. Calculated desolvation free energies of the optimum dimers match the trend in IC(50) values, suggesting the importance of ligand hydrophobicity for effective cation-pi interaction with the peripheral site. PMID- 10514293 TI - Selective A(3) adenosine receptor antagonists: water-soluble 3, 5-diacyl-1,2,4 trialkylpyridinium salts and their oxidative generation from dihydropyridine precursors. AB - A(3) adenosine receptor antagonists are sought for their potential antiinflammatory, antiasthmatic, and antiischemic properties. We have found that 3,5-diacyl-1,2,4-trialkyl-6-phenylpyridinium derivatives constitute a novel class of selective A(3) adenosine receptor antagonists. The structure-activity relationships of this class of antagonists, incorporating the 3-thioester, have been explored. The most potent analogue in this group was 2, 4-diethyl-1-methyl-3 (ethylsulfanylcarbonyl)-5-ethyloxycarbonyl -6-phe nylpyridinium iodide (11), which had an equilibrium inhibition constant (K(i)) value of 219 nM at human A(3) receptors (binding of [(125)I]AB-MECA (N(6)-(4-amino-3-iodobenzyl)-5'-N methylcarbamoyladenosine)) expressed in Chinese hamster ovary (CHO) cells and >10 microM at rat brain A(1) and A(2A) receptors and at recombinant human A(2B) receptors. Compound 11 could be generated through oxidation of the corresponding 3,5-diacyl-1,2,4-trialkyl-6-phenyl-1,4-dihydropyridine, 24, with iodine or in the presence of rat brain homogenates. A 6-cyclopentyl analogue was shown to increase affinity at human A(3) receptors upon oxidation from the 1-methyl-1,4 dihydropyridine analogue, 25, to the corresponding pyridinium derivative, 23 (K(i) 695 nM), suggesting a prodrug scheme. Homologation of the N methylpyridinium derivatives to N-ethyl and N-propyl at the 1-position caused a progressive reduction in the affinity at A(3) receptors. Modifications of the alkyl groups at the 2-, 3-, 4-, and 5-positions failed to improve potency in binding at A(3) receptors. The pyridinium antagonists are not as potent as other recently reported, selective A(3) receptor antagonists; however, they display uniquely high water solubility (43 mM for 11). Compound 11 antagonized the inhibition of adenylate cyclase elicited by IB-MECA in CHO cells expressing the human A(3) adenosine receptor, with a K(B) value of 399 nM, and did not act as an agonist, demonstrating that the pyridinium salts are pure antagonists. PMID- 10514294 TI - Synthesis of a series of 4-benzyloxyaniline analogues as neuronal N-type calcium channel blockers with improved anticonvulsant and analgesic properties. AB - In this article, the rationale for the design, synthesis, and biological evaluation of a series of N-type voltage-sensitive calcium channel (VSCC) blockers is described. N-Type VSCC blockers, such as ziconotide, have shown utility in several models of stroke and pain. Modification of the previously reported lead, 1a, led to several 4-(4-benzyloxylphenyl)piperidine structures with potent in vitro and in vivo activities. In this series, the most interesting compound, (S)-2-amino-1-{4-[(4-benzyloxy-phenyl)-(3-methyl-but-2-enyl)-amino]-p iperidin-1-yl}-4-methyl-pentan-1-one (11), blocked N-type calcium channels (IC(50) = 0.67 microM in the IMR32 assay) and was efficacious in the audiogenic DBA/2 seizure mouse model (ED(50) = 6 mg/kg, iv) as well as the antiwrithing model (ED(50) = 6 mg/kg, iv). Whole-cell voltage-clamp electrophysiology experiments demonstrated that compound 11 blocked N-type Ca(2+) channels and Na(+) channels in superior cervical ganglion neurons at similar concentrations. Compound 11, which showed superior in vivo efficacy, stands out as an interesting lead for further development of neurotherapeutic agents in this series. PMID- 10514295 TI - 3-Alkyl-6-chloro-2-pyrones: selective inhibitors of pancreatic cholesterol esterase. AB - A series of 3-alkyl-6-chloro-2-pyrones with cyclohexane rings tethered to the 3 position was synthesized. The tether ranged from 0 to 4 methylene units. Inhibition of pancreatic cholesterol esterase by this series of pyrones was markedly dependent upon the length of the tether. Dissociation constants as low as 25 nM were observed for 6-chloro-3-(1-ethyl-2-cyclohexyl)-2-pyranone. This class of cholesterol esterase inhibitors functioned as simple competitive inhibitors of substrate binding rather than as suicide substrates or active site inactivators. Trypsin and chymotrypsin were not strongly inhibited by this class of pyrones. Selectivities for cholesterol esterase were greater than 10(3). This is in contrast to 3-aryl-6-chloro-2-pyrones which are nonselective, irreversible inactivators of serine hydrolases. Thus, replacement of the 3-aryl group by an appropriately tethered 3-alkyl ring can produce highly selective inhibitors of cholesterol esterase. A second series of halogen-containing esters was prepared in which cholesterol was esterified with alpha-haloacyl halides. These haloesters were simple substrates of cholesterol esterase with no evidence of irreversible inactivation. PMID- 10514296 TI - Further studies on oxygenated tryptamines with LSD-like activity incorporating a chiral pyrrolidine moiety into the side chain. AB - The enantiomers of 3-(N-methylpyrrolidin-2-ylmethyl)-5-methoxyindole, 1, and 3-(N methylpyrrolidin-2-ylmethyl)-4-hydoxyindole, 3, were prepared using an asymmetric synthesis that employed (+)- or (-)-proline. A new approach was developed that had certain advantages over the synthesis originally reported for the isomers of 1. (+/-)-3-(N-Methylpyrrolidin-3-yl)-4-hydroxyindole, 5, was also prepared as a rigid analogue of psilocin and compared with its 5-methoxy counterpart 4. Radioligand competition assays were used to assess the affinity of compounds for the 5-HT(2A) receptor labeled with the agonist ligand [(125)I]DOI and the antagonist ligand [(3)H]MDL100907. Two-lever drug discrimination assays in rats trained to discriminate either LSD or DOI from saline were employed to assess the hallucinogen-like behavioral properties of these rigid tryptamine analogues. The receptor binding assay results clearly demonstrated a stereochemical preference for the R enantiomers that did not discriminate the position of the oxygen function. The receptor is 10-20-fold more selective for the R isomers. The affinities of the R enantiomers were virtually identical for both 1 and 3 at the agonist-labeled receptor, while racemic 4 and 5 had about one-tenth the affinity. The drug discrimination data in both LSD- and DOI-trained rats paralleled the binding data using [(125)I]DOI displacement. Both (R)-1 and (R)-3 are about equipotent, comparable to DOI in activity but about 10-fold less potent than LSD. Compound 4 produced only partial substitution, even at a dose nearly 5-fold higher than for (R)-1. Based on conformational energies, it seems doubtful that these compounds bind to the 5-HT(2A) receptor in an ergoline-like conformation. The results also suggest that both 1 and 3 would possess LSD-like psychopharmacology in humans. PMID- 10514297 TI - Preparation and characterization of novel trans-[PtCl(2)(amine)(isopropylamine)] compounds: cytotoxic activity and apoptosis induction in ras-transformed cells. AB - The synthesis and chemical characterization of three new transplatinum complexes of structural formula trans-[PtCl(2)(amine)(isopropylamine)] (amine = n,n dimethylamine, propylamine, and butylamine), 1-3, are described. Cytotoxicity tests in tumor cell lines sensitive to cis-DDP (Jurkat, Hela, and Vero) and also in tumor cell lines overexpressing ras oncogenes and resistant to cis-DDP (HL-60 and Pam 212-ras) show that complexes 1 and 3 have higher cytotoxic activity than cisplatin. Moreover, these two trans-Pt(II) complexes kill Pam 212-ras cells through apoptosis induction. These results suggest that trans-PtCl(2) complexes with asymmetric aliphatic amines may be considered a new class of biologically active trans-platinum drugs. PMID- 10514298 TI - Novel partial agonists for the histamine H(3) receptor with high in vitro and in vivo activity. AB - Novel branched N-alkylcarbamates and aliphatic ethers derived from 3-(1H-imidazol 4-yl)propanol were prepared. The compounds were investigated on two functional histamine H(3)-receptor assays. Some compounds displayed partial agonist activity on synaptosomes of rat brain cortex but behaved as pure competitive antagonists on the guinea pig ileum. Under in vivo conditions after po application to mice, some of the compounds showed partial or full agonist activity. Highest in vivo potency was found for the 3,3-dimethylbutyl ether 10 (ED(50) = 0.29 mg/kg, full intrinsic activity). These novel agonists are structurally diverse from classical aminergic histamine H(3)-receptor agonists (e.g., (R)-alpha-methylhistamine, imetit) as they lack a basic moiety in the side chain, which is supposed to be important for the activation of the receptor protein. The selectivity for the histamine H(3) receptor was proven by determination of H(1)- and H(2)-receptor activity on functional assays of the guinea pig. PMID- 10514300 TI - Biologically active secondary metabolites from fungi. 12.(1) oidiolactones A-F, labdane diterpene derivatives isolated from oidiodendron truncata AB - Two known (1 and 2) and four new (3-6) diterpenes named oidiolactones A-F, respectively, and the antibiotic cladosporin were isolated from the fungus Oidiodendron truncata. The structure determination was mainly based on 1D and 2D NMR spectroscopy. The structures of compound 4, displaying an equilibrium between open-chain and cyclized form, and of cladosporin were confirmed by X-ray analysis. PMID- 10514299 TI - New cytotoxic polyacetylenes from the marine sponge Petrosia. AB - New polyacetylenic alcohols with a C(45) carbon skeleton (2) and with an enone moiety in the alkyl chain (C(46), 1) were isolated from the marine sponge Petrosia sp. The gross structures of 1 and 2 were established by spectral methods, and the absolute stereochemistry was determined by the modified Mosher's method. Compounds 1 and 2 displayed considerable cytotoxicity against a small panel of human solid tumor cell lines. Significant inhibitions on DNA replication by 1 and 2 were also observed which could be explanative of their cytotoxicity. PMID- 10514301 TI - Antimicrobial and antitumor activities of mycosporulone. AB - The conditions for optimal production of mycosporulone (1) are given. Its cytotoxic, antimicrobial, and antitumor activities are described. The biological activities of 1 were compared with those of known antibacterial, antifungal, and antitumor agents. The compound was particularly active against Pseudomonas aeruginosa and Staphylococcus aureus (resistant to penicillin). Compound 1 was not toxic to normal human cells (MRC(5)), although it exhibited cytotoxic activity against the human tumor cell lines MDA-MB 231 and PC(3) and the murine L 1210 leukemia cell line. PMID- 10514302 TI - Antioxidant principles from Ephemerantha lonchophylla. AB - One dihydrostilbene and three phenanthrene antioxidants were isolated from an ethanolic extract of the Chinese herbal Ephemerantha lonchophylla. One of these compounds, ephemeranthone (4) is a new natural product. Denbinobin (1) and 3 methylgigantol (3) have been previously isolated from this plant, and 3-ethoxy-5 hydroxy-7-methoxy-1,4-phenanthraquinone (2) is an artifact. The structures of these compounds were determined by spectroscopic analysis. The antioxidative activities for inhibiting human low density lipoprotein oxidation in vitro of compounds 1-4 were determined, and only 4 was active (5.3 times that of probucol). PMID- 10514303 TI - Highly oxygenated pseudopterane and cembranolide diterpenes from the Caribbean sea feather Pseudopterogorgia bipinnata. AB - A chemical study of the sea feather Pseudopterogorgia bipinnata from Colombia has produced four known metabolites, namely, kallolide A (1), bipinnatin A (2), bipinnatin C (3), and bipinnatin J (4), in addition to nine previously undescribed diterpenes possessing an uncommonly high level of oxygenation. One of them, bipinnapterolide A (5), is a new representative of the pseudopterane family of diterpenes possessing the rare 2,3-epoxy-1,4-dione moiety. The other metabolites, bipinnatins G-I (6-8) and bipinnatolides F-J (9-13), are highly oxygenated cembranolide diterpenes. Their chemical structures including relative stereochemistry were established by detailed analysis of the spectral data in addition to X-ray diffraction analysis and NMR spectral comparisons with known pseudopterane and cembranolide models. PMID- 10514304 TI - Nonhalogenated and halogenated phlorotannins from the brown alga carpophyllum angustifolium AB - Using HPLC, forty-five phloroglucinol derivatives were obtained from an ethanolic extract of the brown alga Carpophyllum angustifolium after peracetylation. Compounds of the fuhalol-B series are described. The structure of hydroxyheptafuhalol-B nonadecaacetate is confirmed by the addition of (13)C NMR spectral data, octafuhalol-C heneicosaacetate is described for the first time. The occurrence of four halogenated phlorotannins is reported: 2 chlorophloroglucinol triacetate, 2[D']iododiphlorethol pentaacetate, which had been isolated formerly only as a mixture, 3[A]chlorobifuhalol hexaacetate, and 3[A(4)]chlorodifucol hexaacetate. All substances were characterized by means of spectral analysis. PMID- 10514305 TI - Novel and known constituents from Buddleja species and their activity against leukocyte eicosanoid generation. AB - We have undertaken a systematic survey of the genus Buddleja used in traditional Chinese medicine for antiinflammatory and other indications by testing extracts and isolated natural products for their activity against the enzymes of the arachidonate cascade. This was done by using elicited rat peritoneal leukocytes, a physiologically relevant established whole cell system that expresses both cyclo-oxygenase (COX) and 5-lipoxygenase (5-LOX) activity. Lipophilic extracts of B. globosa roots and B. myriantha stem exhibited inhibitory activities in the 5 LOX and COX enzyme assays, whereas those of B. officinalis flowers, B. yunanesis stems, and B. asiatica stems showed inhibitory activities only against COX. The phytochemical investigation of these extracts, and consequent structure elucidation of isolated compounds using spectroscopic data, led to the isolation from B. globosa of three new terpenoid compounds named dihydrobuddledin A, buddledone A, and buddledone B and four known compounds-buddledins A, B, and C and zerumbone; 12 known compounds from B. officinalis-calceolarioside, campneoside, verbascoside, echinacoside, forsythoside B, angoroside A, crocetin monogentibiosyl ester, acacetin, acacetin-7-O-alpha-L-rhamnopyranosyl (1-6)-beta D-glucopyranoside, acacetin-7-O-alpha-L-rhamnopyranosyl (1-6)[alpha-L rhamnopyranosyl (1-2)]-beta-D-glucopyranoside, songarosaponin A, delta-amyrone; and eight known compounds fromB. yunanesis-11,14-dihydroxy-8,11, 13-abietatrien-7 one, beta-sitosterol, verbascoside, echinacoside, forsythoside B, angoroside A, methylcatapol, and sucrose. Tests on the isolated compounds for inhibition of eicosanoid synthesis showed that buddledin A, crocetin monogentibiosyl ester, and acacetin exhibited an inhibitory effect on COX with IC(50) values of 13.7 microM, 28.2 microM, and 77.5 microM, respectively, whereas buddledin A exhibited inhibitory effect on 5-LOX with an IC(50) value of 50.4 microM. PMID- 10514306 TI - Echinosulfonic acids A-C and echinosulfone A: novel bromoindole sulfonic acids and a sulfone from a southern australian marine sponge, echinodictyum AB - The crude EtOH extract of an Echinodictyum sp. collected during trawling operations in the Great Australian Bight, Australia, displayed antibacterial and antiparasitic properties. Bioassay-directed fractionation yielded three novel sulfonic acids, the echinosulfonic acids A to C (1-3), and a new sulfone, echinosulfone A (4). Structures were assigned to these compounds on the basis of detailed spectroscopic analysis. It was determined that echinosulfonic acids A-C (1-3) and echinosulfone A (4) contributed to the antibacterial but not antiparasitic activity of the crude extract. PMID- 10514308 TI - Geodin A magnesium salt: A novel nematocide from a southern australian marine sponge, geodia AB - A Geodia species collected from southern Australian waters of the Great Australian Bight has yielded a potent new in vitro nematocidal agent identified as geodin A Mg salt (1), a new macrocyclic polyketide lactam tetramic acid magnesium salt. The structure for 1 was assigned on the basis of detailed spectroscopic analysis. PMID- 10514307 TI - Annocherin and (2,4)-cis- and trans-annocherinones, monotetrahydrofuran Annonaceous acetogenins with a C-7 carbonyl group from Annona cherimolia seeds. AB - The new cytotoxic monotetrahydrofuran Annonaceous acetogenins, annocherin (1) and a mixture of (2,4)-cis- and trans-annocherinones (2 and 3), were isolated from the bioactive methanolic extract of Annonacherimolia seeds. Compounds 1-3 each possess an unusual 7-carbonyl group. Their structures were established on the basis of chemical and spectral evidence. Compounds 1-3 showed significant toxicity in the brine shrimp lethality test and cytotoxicity for six human solid tumor cell lines, with selectivity for the renal cell line (A-498) at potencies equivalent to Adriamycin. PMID- 10514309 TI - Four new tetranortriterpenoids from cedrela odorata associated with leaf rejection by exopthalmus jekelianus AB - Four new tetranortriterpenoids, 3-deoxo-3beta,8beta-epoxy-6, 14alpha-dihydroxy 8,14-dihydromexicanolide (cedrodorin, 1); 3-deoxo-3beta,8beta-epoxy-6-acetoxy 14alpha-hydroxy-8, 14-dihydromexicanolide (6-acetoxycedrodorin, 2); 3-deoxo 3beta, 8beta-epoxy-6-deoxy-9alpha,14alpha-dihydroxy-8, 14-dihydromexicanolide (6 deoxy-9alpha-hydroxycedrodorin, 3); and 3-deoxo-3beta,8beta-epoxy 6,9alpha,14alpha-trihydroxy-8, 14-dihydromexicanolide (9alpha-hydroxycedrodorin, 4), have been isolated from the leaves of Cedrela odorata by HPLC. Their molecular structures were determined by 1D and 2D NMR. Three of the compounds are associated with leaf rejection by the polyphagous, folivorous weevil, Exopthalmus jekelianus. The importance of these compounds as insect deterrents in C. odorata, and their potential value in the selection of insect-resistant clones for timber plantations is discussed. PMID- 10514310 TI - Structures and cytotoxicity relationship of isoaaptamine and aaptamine derivatives. AB - A series of 9-O-acylisoaaptamine (3-14) and 4-N-acyl-dihydroaaptamine (16-19) derivatives have been prepared and evaluated for antitumor activity against murine P-388 and human tumor cells including KB16, A549, and HT-29 cell lines. All of compounds showed significant cytotoxicity against P-388 cells. Among them, compounds 9-11 showed potent activity as isoaaptamine (1). There was an apparent lack of linear relationship between cytotoxicity and carbon number of the side chain. The structure and activity relationship for these particular compounds are discussed. PMID- 10514311 TI - Five novel taxanes from taxus canadensis AB - Five novel taxanes (1-5) have been isolated from the needles of Taxus canadensis, and their structures were elucidated on the basis of spectroscopic data. Two of these taxanes have structural features seldom found in yews, with a C-12, C-13 double bond and a hydroxymethylene at C-4. In addition, the 2D structures generated by molecular modeling of taxanes 1 and 2, differing mostly in the location of the double bond in ring A, were compared. The different conformation of ring A derived from the C-12, C-13 double bond is particularly interesting because it implies that if a Taxol (paclitaxel) side chain were attached to C-13 it would have a different orientation from that in paclitaxel. PMID- 10514312 TI - Calycopterones and calyflorenones, novel biflavonoids from Calycopteris floribunda. AB - Neocalycopterone (1) and its methyl ether (2), along with two new biflavonoids, calyflorenones A (3) and B (4), were isolated from dried leaves of Calycopteris floribunda (Combretaceae). Structures 1-4 were investigated by spectroscopic methods. The relative stereochemistry of 3 and 4 was deduced through NOE and ROESY experiments, comparative CD and optical rotation evaluations. Cytotoxicity test results of 1 and 2 are reported. The monoflavonoid penduletin was isolated as a minor component from C. floribunda for the first time. PMID- 10514313 TI - Triterpene saponins and lignans from the roots of Pulsatilla chinensis and their cytotoxic activity against HL-60 cells. AB - Two new triterpene saponins (8 and 9) and seven previously reported triterpene saponins (1-7) based upon oleanolic acid or hederagenin, along with two known lignans, (+)-pinoresinol (10) and beta-peltatin (11), were isolated from a saponin fraction prepared from the MeOH extract of the roots of Pulsatilla chinensis. The structures of the new saponins were determined by spectroscopic analysis, including 2D NMR spectroscopic techniques, and the results of hydrolytic cleavage. The isolated compounds and some derivatives were evaluated for their cytotoxic activity against HL-60 human leukemia cells. PMID- 10514315 TI - Withanolides from ajuga parviflora AB - Two new withanolides named ajugin A [14alpha,20, 28-trihydroxy-1-oxo-(20R,22R) witha-3,5,24-trienolide] (1) and ajugin B [14alpha,20,27-trihydroxy-1-oxo (20R,22R)-witha-5, 24-dienolide] (2) were isolated from the whole plant of Ajuga parviflora, and their structures were deduced by NMR and MS analyses. PMID- 10514314 TI - Stereochemistry of the roridins. Diastereomers of roridin E. AB - A careful investigation of cultures of Myrothecium verrucaria has shown that this fungus produces all four roridin E isomers (3a-d), diastereomeric at the C-6' and C-13' centers. The stereochemistries at these centers were established by a combination of X-ray crystallographic analysis, NMR spectroscopy, and chemical transformations. NMR data from these and other macrocyclic trichothecenes allows for the assignment of configurations at the C-6' and C-13' centers for most of these compounds, the exceptions being those congeners having a C-4' ketone group in the macrolide ring. PMID- 10514316 TI - Strong antinociceptive effect of incarvillateine, a novel monoterpene alkaloid from Incarvillea sinensis. AB - Incarvillea sinensis is a wild plant distributed in northern China. The dried whole plant has been traditionally used to treat rheumatism and relieve pain as an ancient Chinese crude drug. To investigate its antinociceptive activity, we evaluated several fractions derived from the methanolic extract of Incarvillea sinensis in the formalin-induced pain model in mice. Incarvillateine, a novel monoterpene alkaloid, has been found to show significant antinociceptive activity. Here we report the antinociceptive activity of incarvillateine and compare its activity with that of morphine. Additionally, we suggest that its action may be related to influence on the central opioid pathways. PMID- 10514317 TI - New bromopyrrole alkaloids from the Indopacific sponge Agelas nakamurai. AB - Two new dimeric bromopyrrole alkaloids, nakamuric acid (1) and its corresponding methyl ester (2), have been isolated from the Indopacific sponge Agelas nakamurai along with the known metabolites sceptrin (3), debromosceptrin (4), and ageliferin (5). Their structures were identified by analysis of spectral data. All compounds inhibited the growth of several Gram-positive and Gram-negative bacteria in the agar plate diffusion assay. PMID- 10514318 TI - New xanthorin glycosides from a Dermocybe species. AB - A mixture of the 8-O-beta-D-glucopyranoside of omega-hydroxyxanthorin 1-O-methyl ether (3) and the 8-O-beta-D-gentiobioside of xanthorin 1-O-methyl ether (4) was isolated from the water-soluble extractives of the Australian toadstool Dermocybe sp. WAT 22963. Compounds 3 and 4 were identified by characterization of their respective peracetyl derivatives 5 and 6. PMID- 10514319 TI - Major flavonoids from Arabidopsis thaliana leaves. AB - The three major flavonoids isolated from Arabidopsis thaliana plants grown in the greenhouse were identified by means of spectroscopic analysis (UV, NMR, MS) and chiral capillary zone electrophoresis as the novel kaempferol 3-O-beta-[beta-D glucopyranosyl(1-->6)D-glucopyranoside]-7-O-alpha-L- rhamnopyranoside (1), kaempferol 3-O-beta-D-glucopyranoside-7-O-alpha-L-rhamnopyranoside (2), and kaempferol 3-O-alpha-L-rhamnopyranoside-7-O-alpha-L-rhamnopyranoside (3). Comprehensive NMR studies including selective 1D and gradient-enhanced 2D techniques were applied in order to achieve full signal assignment and definitive proof of linkage for compound 1. PMID- 10514320 TI - Methanol adduct of puupehenone, a biologically active derivative from the marine sponge Hyrtios species. AB - A methanol adduct of puupehenone (1), 15alpha-methoxypuupehenol (2), an artifact resulting from the action of MeOH on puupehenone, was isolated during purification of the CH(2)Cl(2) extract of the New Caledonian marine sponge Hyrtios sp., as the major constituent. Its chemical structure was elucidated by 2D NMR experiments. Compound 2 displayed similar antimicrobial and antifungal activity as puupehenone and a lower cytotoxic activity toward KB cells with ED(50) values of 6 and 0.5 microg/mL, respectively. Compound 2 was slightly more active against three strains of Plasmodium falciparum than puupehenone. PMID- 10514321 TI - Longithorols A and B, novel prenylated paracyclophane- and metacyclophane-type hydroquinones from the tunicate Aplidium longithorax. AB - The tunicate Aplidium longithorax collected from Palau contained two novel prenylated paracyclophane- and metacyclophane-type hydroquinones, longithorols A (1) and B (2), in addition to longithorones A-I. Longithorols A and B were very unstable and were therefore isolated as their more stable pentaacetate forms, 3 and 4, respectively. The structures of 3 and 4 were determined by spectral data, especially 2D NMR data. PMID- 10514322 TI - Three secoiridoid glucosides from Jasminum nudiflorum. AB - Phytochemical study of the leaves and stems of Jasminum nudiflorum has led to the isolation of three secoiridoid glucosides, jasnudiflosides A-C (1-3). The structures of these compounds were elucidated on the basis of chemical and spectroscopic evidence. PMID- 10514323 TI - A new cacospongionolide derivative from the sponge Fasciospongia cavernosa. AB - Cacospongionolide F (4a), a new bioactive cacospongionolide-related sesterterpene, has been isolated from the Northern Adriatic sponge Fasciospongia cavernosa. The structure was proposed on the basis of spectroscopic data and chemical transformations. The absolute configuration was established using the modified Mosher's method. A molecular mechanics study of the dehydrodecalin ring explained the observed differences in dynamic behavior between cacospongionolide F and mamanuthaquinone, a related compound. Antimicrobial activity, brine shrimp and fish lethalities of this new compound are reported. PMID- 10514324 TI - Antibacterial diterpenoid acids from Azorella compacta. AB - The two novel diterpenoid acids mulin-12,14-dien-11-on-20-oic acid (1) and mulin 12-ene-11,14-dion-20-oic acid (2) have been isolated from Azorella compacta. Their structures have been elucidated by 1D and 2D NMR methods. In contrast to the closely related known mulinolic acid (3) and its dehydration product (4) these new natural products have been shown to exhibit antimicrobial activity. PMID- 10514325 TI - New fish feeding deterrents, including a novel sesquiterpenoid heterogorgiolide, from the brazilian gorgonian heterogorgia uatumani(Octocorallia, gorgonacea) AB - Studies of the Brazilian gorgonian octocoral Heterogorgia uatumani have resulted in the discovery of two metabolites that inhibit fish feeding under natural conditions. These are the previously reported eunicellane diterpenoid, (6E) 2alpha,9alpha-epoxyeunicella-6, 11(12)-dien-3beta-ol (1) and a new sesquiterpene lactone, heterogorgiolide (2). The structures of 1 and 2 were determined by spectroscopic methods and by comparison of spectral data with literature values. Field bioassays of the two compounds, at their natural concentrations, confirmed that they deter predation by a complex assemblage of reef fishes. This is an unusual observation showing that defenses are derived from both sesqui- and diterpenoid metabolites. PMID- 10514326 TI - Morinins H-K, four novel phenylpropanol ester lipid metabolites from morina chinensis AB - The medicinal plant, Morina chinensis, afforded four novel phenylpropanol ester lipid metabolites, named morinins H-K (1-4). Their structures were identified on the basis of spectral data interpretation. PMID- 10514327 TI - Mitorubrin derivatives on ascomata of some talaromyces species of ascomycetous fungi AB - Two groups, producers of (+)- and (-)-mitorubrin derivatives, coexist in the series Lutei of the genus Talaromyces. The optical rotations of mitorubrins from T. emodensis, T. hachijoensis, and T. wortmannii var. sublevisporus, which produced mitorubrinol acetate (5), were all positive, whereas those from T. austrocalifornicus and T. convolutus, which produced mitorubrinal (3) and mitorubrinic acid (4), were all negative. PMID- 10514328 TI - New nitrogenous bisabolene-type sesquiterpenes from a micronesian marine sponge, axinyssa species AB - Chemical investigation of an Axinyssa species of sponge collected at Yap Island afforded three new nitrogenous sesquiterpenes, 1-3, together with the known compound 3-formamidotheonellin (4). The structures of compounds 1, 3, and 4 were confirmed by spectroscopic methods, and compound 2 was tentatively identified by comparison of its (1)H and (13)C NMR data with those of 1, 3, and 4 and (1)H decoupling experiments. PMID- 10514329 TI - Tanikolide, a toxic and antifungal lactone from the marine cyanobacterium Lyngbya majuscula. AB - A new brine-shrimp toxic and antifungal compound, tanikolide 1, has been isolated from the lipid extract of a Madagascan collection of the marine cyanobacterium Lyngbya majuscula. The structure of tanikolide was determined by spectroscopic methods, relying heavily on 2D NMR spectroscopy. The absolute configuration at C 5 of tanikolide was established as R by oxidizing the primary alcohol to an acid and analyzing the corresponding (R)- and (S)-PGME amide derivatives by (1)H NMR. PMID- 10514330 TI - Swinholides and new acetylenic compounds from an undescribed species of Theonella sponge. AB - Four new acetylenic compounds, 5-8, along with four known ones, 1-4, were isolated from a sponge, Theonella sp., collected in Chuuk Atoll, Federated States of Micronesia. Three unrelated swinholide-type compounds were also isolated. The structures of the new acetylenes were determined from spectral data and chemical degradation. PMID- 10514331 TI - Two new cytotoxic carbonimidic dichlorides from the nudibranch Reticulidia fungia. AB - Two new sesquiterpenes, reticulidins A (1) and B (2), containing a rare functional group, N=CCl(2), have been isolated together with known congeners from the nudibranch Reticulidia fungia and their structures elucidated by spectroscopic data. Both 1 and 2 were cytotoxic against KB and L1210 cells. PMID- 10514332 TI - New antimycobacterial saponin from Colubrina retusa. AB - A new jujubogenin saponin was isolated from the stems of Colubrina retusa and identified as jujubogenin 3-O-alpha-L-arabinofuranosyl (1-->2)-[3-O-(trans)-p coumaroyl-beta-D-glucopyranosyl (1-->3)]-alpha-L-arabinopyranoside (4) on the basis of chemical and spectroscopic data. The antimycobacterial activity expressed as minimum inhibitory concentration (MIC) for compound 4 was 10 microg/mL. PMID- 10514333 TI - 3-methyl-4-phenylpyrrole from the ants Anochetus kempfi and Anochetus mayri. AB - The cephalic extracts of the ant Anochetus kempfi were found to contain 2,5 dimethyl-3-isoamylpyrazine (1) and 3-methyl-4-phenylpyrrole (2). The structures of these compounds were established from their spectral data and by comparison with synthetic samples. This is the first report of a phenylpyrrole found in an insect and only the third report of a pyrrole from ants. PMID- 10514334 TI - 1',2',3',4'-tetradehydrotubulosine, a cytotoxic alkaloid from Pogonopus speciosus. AB - Bioassay-guided phytochemical investigation of the stems of Pogonopus speciosus, using human oral epidermoid carcinoma (KB) cells as a monitor, led to the isolation of a novel alkaloid, 1',2', 3',4'-tetradehydrotubulosine (1), along with tubulosine (2) and psychotrine (3) as bioactive constituents. The structure of the novel compound was elucidated through 1D- and 2D-NMR spectroscopic methods. Alkaloids 1 and 3 showed weak cytotoxic activity against a panel of human cancer cell lines, with the potency of these compounds being markedly less than that of tubulosine (2). PMID- 10514336 TI - In memoriam: jeffrey L. Fox PMID- 10514335 TI - An unusual novel C(29) steroid from the soft coral lobophytum crassum PMID- 10514338 TI - Transdermal penetration enhancers: applications, limitations, and potential. PMID- 10514337 TI - Blood-brain barrier properties of human immunodeficiency virus antiretrovirals. PMID- 10514339 TI - Frequency-domain photon migration measurements for quantitative assessment of powder absorbance: A novel sensor of blend homogeneity. AB - The measurement and analysis of frequency-domain photon migration (FDPM) measurements of powder absorbance in pharmaceutical powders is described in the context of other optical techniques. FDPM consists of launching intensity modulated light into a powder and detecting the phase delay and amplitude modulation of the re-emitted light as a function of the modulation frequency. From analysis of the data using the diffusion approximation to the radiative transport equation, the absorption coefficient can be obtained. Absorption coefficient measurements of riboflavin in lactose mixtures are presented at concentrations of 0.1 to 1% (w/w) at near-infrared wavelengths where solution absorption cross sections are difficult to accurately measure using traditional transmission measurements in nonscattering solutions. FDPM measurements in powders enabled determinations of absorption coefficients that increase linearly with concentration (w/w) according to Beer-Lambert relationship. The extension of FDPM for monitoring absorbance of low-dose and ultralow-dose powder blending operations is presented. PMID- 10514340 TI - Solubilization of fluasterone. AB - Solubilization of nonpolar drugs constitutes one of the most important tasks in parenteral formulations design. This study investigates and assesses the solubility enhancement of Fluasterone by various techniques including cosolvency, micellization, and complexation. Of the solubilizing agents used, the modified beta-cyclodextrins were found to be the most effective. The solubility of Fluasterone is 1.55 x 10(-4) mM, 3.13 mM, and 4.04 mM in water, 20% sulfobutyl ether-beta-cyclodextrin (SBEbetaCD), and 20% hydroxypropyl-beta-cyclodextrin (HPbetaCD), respectively. PMID- 10514341 TI - Heptakis(2,6-di-O-methyl-3-O-acetyl)-beta-cyclodextrin: A water-soluble cyclodextrin derivative with low hemolytic activity. AB - Acetyl groups were introduced to the hydroxyl groups of heptakis(2, 6-di-O methyl)-beta-cyclodextrin (DM-beta-CyD), and the resulting heptakis(2,6-di-O methyl-3-O-acetyl)-beta-CyD (DMA-beta-CyD) was evaluated for the inclusion property and hemolytic activity. It was confirmed by means of NMR and mass spectroscopies that in the DMA-beta-CyD molecule, all seven hydroxyl groups at the 3-position were substituted by acetyl groups. Thus, it has the degree of substitution (DS) of 7, whereas DMA4-beta-CyD with the lower substitution (DS 3.8) was a mixture of components with different DS. The aqueous solubility of DMA beta-CyD was higher than those of beta-CyD, DM-beta-CyD, and heptakis(2,3,6-tri-O methyl)-beta-CyD (TM-beta-CyD). The hydrophobicity of the whole molecule, assessed from measurements of surface tension, increased in the order of DM-beta CyD < DMA-beta-CyD < TM-beta-CyD. The half-life of DMA-beta-CyD for hydrolysis in pH 9.5 and 60 degrees C was about 19 h, and there was only slight liberation of acetic acid in rabbit plasma and carboxylesterase (EC 3.1.1.1) at 37 degrees C. DMA-beta-CyD had an inclusion ability similar to that of TM-beta-CyD for p hydroxybenzoic acid esters with different alkyl chain lengths and an antiinflammatory drug, flurbiprofen, although it was inferior to that of DM-beta CyD. The hemolytic activity and rabbit muscular irritation of DMA-beta-CyDs were much weaker than those of DM-beta-CyD: no hemolysis was observed even in the presence of 0.1 M DMA-beta-CyD with DS 7. The results suggest that the water soluble CyD derivative with superior bioadaptability and inclusion ability can be prepared by properly designing substituents at the 3-position and by optimally controlling their degree of substitution. PMID- 10514342 TI - Modulation of sulfate renal transport by alterations in cell membrane fluidity. AB - Changes in membrane fluidity have been shown to alter the sodium-dependent renal transport of glucose and phosphate; however, this has not been examined for sodium/sulfate cotransport in the renal proximal tubule. Sodium/sulfate cotransport regulates the homeostasis of sulfate in mammals. The objective of this study was to investigate the influence of alterations of membrane fluidity on sodium-coupled sulfate transport in the Madin-Darby canine kidney cells, which have been stably transfected with sodium/sulfate cotransporter (NaSi-1) cDNA (MDCK-Si). Preincubation of cells with 0. 2 mM cholesterol significantly decreased the V(max) for sodium/sulfate cotransport (13.69 +/- 1.11 vs 10.15 +/- 1.17 nmol/mg protein/5 min, mean +/- SD, n = 4, p < 0.01) with no significant alteration in K(m). The addition of benzyl alcohol (20 mM) to cells increased the V(max) of sulfate uptake by 20% (11.97 +/- 0.91 vs 14. 35 +/- 0.56 nmol/mg protein/5 min, mean +/- SD, n = 3, p < 0.05) with no significant change in K(m). Membrane fluidity, as measured by the fluorescence polarization of 1,6-diphenyl 1,3,5-hexatriene (DPH), was significantly increased in MDCK-Si cells treated with 20 mM benzyl alcohol and decreased in the cells preincubated with 0.2 mM cholesterol, compared with control cells. Our results suggest that alterations in membrane fluidity that may occur as a result of disease states, aging, and pregnancy may play an important role in the modulation of renal sodium/sulfate cotransport. PMID- 10514343 TI - Physicochemistry, pharmacokinetics, and pharmacodynamics of S-nitrosocaptopril crystals, a new nitric oxide donor. AB - S-nitrosocaptopril (CapNO) has been proposed as a compound possessing capacities of both a nitric oxide (NO) donor and an inhibitor of angiotensin converting enzyme (ACE). In the present study, we characterized the physicochemical, pharmacokinetic, and pharmacological properties of the crystalline CapNO. The novel stable crystals are in a red flake form. Spectroscopic analyses of CapNO revealed its UV/visible lambda(max) and the corresponding extinction coefficients, and characteristic infrared frequencies for the N=O and S-N stretch. The NMR signals corresponding to the protons attached to the carbon (C S) and the carbon itself were remarkably shifted downfield upon S-nitrosylation. Mass and HPLC analyses, solubility, and melting point of CapNO were determined. Simultaneous on-line analyses of pharmacodynamic and pharmacokinetic profiles of CapNO in catheterized awake rats of spontaneous hypertension (SHR) showed acute decreases in mean arterial pressure (MAP), concomitant with the corresponding increases in plasma levels of CapNO after po or iv administration. The pharmacokinetic parameters for CapNO, i.e., t(1/2), T(max), C(max), V(d), AUC, and oral bioavailability were analyzed to understand the dose-dependent potency and effective period of CapNO. The highest concentrations of oral CapNO distributed in tissues were found in kidney, liver, lung, and small intestine. CapNO was excreted predominantly via urine, and second via feces in the detectable forms of thiols and nitrogen oxide although a small portion of CapNO was found in bile. The results provide the evidence of in vivo cleavage of the S N bond and biotransformation of CapNO. PMID- 10514344 TI - Role of baseline parameters in determining indirect pharmacodynamic responses. AB - Indirect Response Models account for the pharmacodynamics of numerous drugs which inhibit or stimulate the production (k(in)) or loss (k(out)) of the response variable (R). The dose and pharmacokinetics, capacity (S(max), I(max)), and potency (SC(50), IC(50)) factors of the Hill function incorporated in these models are the primary determinants of overall responsiveness. However, the initial or baseline value for the response (R(0) = k(in)/k(out)) should also be considered as an important factor for the net response. Using Indirect Response Model III (stimulation of input) as an example, the net area under the effect curve (AUEC(NET)) can be proportional to the R(0) values. Such a feature is demonstrated in this report by computer simulations, by examination of the integral of the simulated response vs time profiles, and with examples from the literature. Also shown is an adjustment of R(0) when the therapeutic agent is an endogenous substance. These analyses show that the role of R(0) and k(in) should not be overlooked as determinants of indirect responses and source of variation among subjects or patient groups. PMID- 10514345 TI - Pharmacokinetic and glucodynamic comparisons of recombinant and animal-source glucagon after IV, IM, and SC injection in healthy volunteers. AB - The structure of the hormone glucagon is identical among humans and several species of other mammals. Equivalence of recombinant glucagon (rG) to animal source glucagon (aG) was assessed in this two-part, open-label, randomized study. Part I was a four-way crossover intravenous dose-ranging study of rG (pH 2.8) involving 12 subjects. Part II was a six-way crossover study of 29 subjects comparing rG (diluent pH 2.0 and 2.8) with aG administered subcutaneously (sc) and intramuscularly (im). Maximum glucagon plasma concentrations (C(max)) and area under the glucagon concentration curve (AUC) were calculated. Additionally, maximum blood glucose concentrations (BG(max)), maximum absolute BG excursion (MAE), and area under the glucose concentration curve from time of dosing to return to baseline (AUC(rtb)) were calculated. The primary focus was equivalence of the formulation intended for marketing (rG pH 2.0) to aG. Administration of rG pH 2.0 through the im route demonstrated equivalence to aG for all pharmacokinetic and glucodynamic comparisons. Subcutaneous administration of rG pH 2.0 demonstrated standard bioequivalence for AUC (5.87 versus 6.63 ng x h/mL; NS) and near equivalence for C(max) (7.94 versus 9.12 ng/mL; p < 0.05). rG pH 2.0 showed glucodynamic equivalence to aG (BG(max), 136 versus 133 mg/dL; MAE, 50.0 versus 47.4 mg/dL, respectively) and statistically greater AUC(rtb) values (151 versus 126 mg x h/dL, p < 0. 05). rG and aG were equally safe and well tolerated. In conclusion, rG provides equivalent safety and efficacy to aG. PMID- 10514346 TI - Formulation of highly soluble poly(ethylene glycol)-peptide DNA condensates. AB - Two poly(ethylene glycol) (PEG)-peptides were synthesized and tested for their ability to bind to plasmid DNA and form soluble DNA condensates with reduced spontaneous gene expression. PEG-vinyl sulfone or PEG-orthopyridyl disulfide were reacted with the sulfhydryl of Cys-Trp-Lys(18) (CWK(18)) resulting in the formation of nonreducible (PEG-VS-CWK(18)) and reducible (PEG-SS-CWK(18)) PEG peptides. Both PEG-peptides were prepared on a micromole scale, purified by RP HPLC in >80% yield, and characterized by (1)H NMR and MALDI-TOF. PEG-peptides bound to plasmid DNA with an apparent affinity that was equivalent to alkylated (Alk)CWK(18), resulting in DNA condensates with a mean diameter of 80-90 nm and zeta (zeta) potential of +10 mV. The particle size of PEG-peptide DNA condensates was constant throughout the DNA concentration range of 0. 05-2 mg/mL, indicating these to be approximately 20-fold more soluble than AlkCWK(18) DNA condensates. The spontaneous gene transfer to HepG2 cells mediated by PEG-VS-CWK(18) DNA condensates was over two orders of magnitude lower than PEG-SS-CWK(18) DNA condensates and three orders of magnitude lower than AlkCWK(18) DNA condensates. PEG-VS-CWK(18) efficiently blocked in vitro gene transfer by reducing cell uptake. The results indicate that a high loading density of PEG on DNA is necessary to achieve highly soluble DNA condensates that reduce spontaneous in vitro gene transfer by blocking nonspecific uptake by HepG2 cells. These two properties are important for developing targeted gene delivery systems to be used in vivo. PMID- 10514347 TI - Rapidly disintegrating tablets prepared by the wet compression method: mechanism and optimization. AB - To make rapidly disintegrating tablets with sufficient mechanical integrity, tablets were prepared by compressing wet granules under low compression force and then drying the resulting wet mass in a circulating-air oven (wet compression method). Lactose with various particle sizes was used as the excipient, and water was used as a wetting agent. The effect of drying time, compression force, size of lactose particles, and moisture content of wet granules on tablet properties indicated that the formation and disintegration time of tablets were related to the effect of the formation of solid bridges between lactose particles. By optimizing compression force, size of lactose particles, and moisture content of the granules, tablets meeting tensile strength greater than 0.5 MPa and disintegration time shorter than 15 s were obtained by the wet compression method. PMID- 10514348 TI - Development and evaluation of microbicidal hydrogels containing monoglyceride as the active ingredient. AB - A number of medium-chain saturated and long-chain unsaturated fatty acids and their monoglycerides were tested against herpes simplex virus (HSV-1) to determine which lipids were most active during a short incubation time. The aim was to find which lipid would be preferable as the active ingredient in a virucidal hydrogel formulation for the purpose of preventing transmission of pathogens to mucosal membranes, particularly sexually transmitted viruses, such as herpes simplex virus and human immunodeficiency virus (HIV), and bacteria, such as Chlamydia trachomatis and Neisseria gonorrheae. The main strategy was that the formulations would be fast-acting, killing large numbers of virus or bacteria on contact in a short time, preferably causing at least a 10000-fold reduction in virus/bacteria titer in 1-5 min. Monocaprin, the 1-monoglyceride of capric acid, and lauric acid were found to be most active of all the lipids tested, causing a greater than 100000-fold reduction in virus titer in 1 min at a concentration of 20 mM. When tested at a concentration of 10 mM for 1 min, monocaprin was still fully active whereas lauric acid had no or negligible activity. It was concluded that monocaprin was most suitable as the active ingredient in a fast-acting virucidal gel formulation, and several hydrogel formulations containing monocaprin were tested. Formulations where the monoglyceride was dissolved in glycofurol were found to be active against HSV-1. The hydrogel formulations containing 20 mM monocaprin were highly virucidal in vitro and caused a greater than 100000-fold (HSV-1) inactivation of virus in human semen in 1 min. Formulations in dilution 1:10 were cytotoxic in monolayers of CV-1 cells, but they were 10-100 fold less cytotoxic than a commercial product which contains 2% nonoxynol-9. PMID- 10514349 TI - Biopharmaceutics and pharmacokinetics of 5-phenyl-1, 2-dithiole-3-thione complexed with sulfobutyl ether-7-beta-cyclodextrin in rabbits. AB - The biopharmaceutics and pharmacokinetics of 5-phenyl-1, 2-dithiole-3-thione (5PDTT) were investigated in rabbits, after administration as a complex with sulfobutyl-ether-7-beta-cyclodextrin (SBE7-beta-CD) by intravenous and oral routes and as a micronized powder by oral route. 5PDTT had a rapid and large red blood cell partitioning that was not dependent on drug concentration either in vitro or ex vivo. The blood clearance was very high (354 +/- 131 mL/min) suggesting extrahepatic metabolism and/or nonrenal elimination and a significant volume of distribution (67 +/- 76 L). The renal clearance was 0.17% of total clearance. 5-phenyl-1,2-dithiol-3-one (5PDTO) was identified as a metabolite in blood and urine. The bioavailability of 5PDTT following administration of 5PDTT/SBE7-beta-CD complex was estimated to 41% while it was close to zero when 5PDTT was given as a micronized powder. PMID- 10514350 TI - Dehydration behavior of eprosartan mesylate dihydrate. AB - Eprosartan mesylate (SKF 108566-J; EM) is an antihypertensive agent approved for marketing in the USA. EM dihydrate was prepared by three methods, one of which included suspending the anhydrous drug in an aqueous solution of 1.0 M methanesulfonic acid to form a slurry, followed by filtration. The dehydration kinetics of EM dihydrate were derived by analyzing the fit of the isothermal thermogravimetric analytical (TGA) data to numerous kinetic models. EM dihydrate undergoes dehydration in two distinct steps, each involving the loss of 1 mol of water at 25-70 degrees C and 70-120 degrees C, respectively. Recrystallization of EM occurs at approximately 120-140 degrees C after dehydration to the anhydrous phase. This explanation is supported by variable temperature powder X-ray diffractometry. The mechanism of the dehydration reaction is complex, the dependence of the reaction rate on temperature varying as a function of the particles size. For the dihydrate of sieve fraction <125 microm, the kinetics of the first and second dehydration steps are consistent with the Avrami-Erofeev equation (A3, n = 1/3) over the temperature range studied, corresponding to three dimensional growth of nuclei. In contrast, for the 125-180-microm and 180-250 microm sieve fractions, the kinetics are best described by the two-dimensional phase boundary reaction (R2) at a lower dehydration temperature (i.e., 28.3 degrees C), and by the Avrami-Erofeev equation (A3, n = 1/3) at a higher dehydration temperature (i.e., 93.7 degrees C). The activation energies (15-40 kcal/mol) and frequency factors of the dehydration of EM dihydrate were determined both by Arrhenius plots of the isothermal rates determined by TGA and by Kissinger plots of the nonisothermal differential scanning calorimetric data. Hot stage microscopy of single crystals of EM dihydrate showed random nucleation at the surface and dehydration with the growth of microcrystals along the needle a axis. Cerius(2) molecular modeling software showed the existence of water channels along the a axis and enabled the observed dehydration behavior of EM dihydrate crystals to be explained in terms of the bonding environment of water molecules in the crystal structure. PMID- 10514351 TI - Effect of transdermal iontophoresis codelivery of hydrocortisone on metoclopramide pharmacokinetics and skin-induced reactions in human subjects. AB - The effects of transdermal iontophoresis (IP) codelivery of hydrocortisone (HC) on metoclopramide hydrochloride (MCP) pharmacokinetics and on skin-induced reactions were evaluated in a randomized, crossover clinical study. MCP, an antiemetic, low molecular weight, cationic drug intended for systemic delivery, was delivered from the anode of IP systems at a constant current of 100 microA/cm(2). HC, a neutral endogenous antiinflammatory agent, was codelivered from the same electrode, primarily by electroosmotic processes. Each subject (n = 7) wore two identical IP systems (MCP alone or MCP plus HC), each supplying 500 microA, one on each upper arm for 4 h. One week later, each subject repeated the procedure with the alternate type of MCP system. HC did not change the pharmacokinetics of MCP: There were no statistically significant differences in MCP plasma concentrations, half-life, area under the curve (AUC), or rate of absorption between the two treatment groups. However, HC significantly decreased erythema and edema scores produced by the IP of MCP. In both groups, a steady state MCP flux of about 100 microg/(cm(2) x h) was achieved after only 1 h of transport, and input rate dropped dramatically immediately after removal of the system. In vitro, HC flux through human epidermis from an MCP plus HC formulation was 2.8 +/- 1.1 microg/(cm(2) x h) after 4 h transport at 100 microA/cm(2), suggesting negligible systemic exposure to hydrocortisone. These data indicate that MCP input rate and its clearance from the skin are unaltered by HC and that the codelivery of HC by IP is an effective strategy for inhibition of local reactions resulting from the transdermal delivery of drugs. PMID- 10514352 TI - Development of an in vitro dissolution method using microdialysis sampling technique for implantable drug delivery systems. AB - The major challenge faced during the development of implantable dosage forms for site-specific delivery is monitoring the local concentration of the drug at or around the site of action. The tissue concentration at the site is generally measured by either sacrificing the animal at different points in time or by determining the amount of drug left in the implants at various time intervals. Unfortunately, there are no official in vitro dissolution methods available to study the release characteristics of drugs from this drug delivery system. The objective of this investigation was to develop a simple method using microdialysis sampling technique to serve as an in vitro dissolution method for implantable drug delivery systems. Ciprofloxacin implants were prepared by compressing ciprofloxacin microcapsules in poly(lactic acid) (PLA) and poly(lactic-glycolic acid) (PLGA). A sensitive HPLC method was developed and validated for the assay of Ciprofloxacin. An in vitro dissolution method was developed to study the release characteristics of drug from these implants. The method used a microdialysis sampling technique and a small sample volume of release medium. The various advantages and disadvantages of this method over other USP methods are discussed. PMID- 10514353 TI - A spin filter method for continuous evaluation of hemolysis. AB - A novel method for quantifying hemolysis is described. This method uses a spin filter to separate the free hemoglobin from the red blood cells suspended in the test solution. This procedure enables the use of a closed loop system that continuously measures hemolysis spectrophotometrically. It is shown that hemolysis does not always stop after the solution has been quenched with normal saline. In fact, the process of hemolysis induced by chemicals such as potassium oleate is relatively slow. PMID- 10514354 TI - Pore charge distribution considerations in human epidermal membrane electroosmosis. AB - The aim of this study was to assess the extent to which a model with pores having only net negative charges would adequately describe transdermal electroosmosis in human epidermal membrane (HEM) at neutral pH. Such information would enhance the predictive value of the modified Nernst-Planck model for transdermal iontophoresis, in addition to providing insights regarding the likelihood of significant pore charge distribution in HEM. Baseline results (the control) obtained from 0.1 to 0.4 V anodal and cathodal electroosmosis experiments with synthetic polycarbonate membranes (Nuclepore membranes), using radiolabeled urea and mannitol as the model permeants, demonstrated that such a membrane system can be modeled by the electrokinetic (electroosmosis) theory with the assumption of the pores possessing only negative charges. The studies with HEM were carried out at low voltage (/approximately = 1.0 V) where significant field induced pore formation in HEM occurred. In both the low and high voltage studies, radiolabeled urea, mannitol, and water were employed as permeants in cathodal and anodal iontophoresis experiments. The results of the low voltage iontophoresis experiments suggest significant pore charge distribution in HEM (a significant deviation between the predictions from the single pore charge type assumption and the experimental data). Under the higher applied voltage conditions (>/approximately = 1.0 V), results from anodal and cathodal electroosmosis studies were consistent with the model in which the HEM has only pores that are net negatively charged. PMID- 10514355 TI - Studies of diffusional release of a dispersed solute from polymeric matrixes by finite element method. AB - This paper presents systematic analyses by the finite element method of release kinetics of a dispersed solute from various matrixes (i.e. , slab, sphere, cylinder, and convex tablet), with or without boundary-layer resistance, into a finite or an infinite external volume. In the case of sink conditions, the numerical results agree well with the existing analytical solutions. For the problems of solute release into a finite external volume, where the analytical solutions are not available, this work has provided numerical solutions of the differential equations describing the release kinetics, moving boundaries, and concentration profiles. This work has also revealed the dependence of release kinetics on the initial solute loading, the external volume, and the boundary layer thickness. The method presented here can describe the entire process of diffusional release before and after the dispersed solute has been dissolved without the pseudo steady-state assumption and it is applicable to both small and large ratio of initial solute loading to the solute solubility in the matrix. PMID- 10514356 TI - Solid dispersion of poorly water-soluble drugs: early promises, subsequent problems, and recent breakthroughs. AB - Although there was a great interest in solid dispersion systems during the past four decades to increase dissolution rate and bioavailability of poorly water soluble drugs, their commercial use has been very limited, primarily because of manufacturing difficulties and stability problems. Solid dispersions of drugs were generally produced by melt or solvent evaporation methods. The materials, which were usually semisolid and waxy in nature, were hardened by cooling to very low temperatures. They were then pulverized, sieved, mixed with relatively large amounts of excipients, and encapsulated into hard gelatin capsules or compressed into tablets. These operations were difficult to scale up for the manufacture of dosage forms. The situation has, however, been changing in recent years because of the availability of surface-active and self-emulsifying carriers and the development of technologies to encapsulate solid dispersions directly into hard gelatin capsules as melts. Solid plugs are formed inside the capsules when the melts are cooled to room temperature. Because of surface activity of carriers used, complete dissolution of drug from such solid dispersions can be obtained without the need for pulverization, sieving, mixing with excipients, etc. Equipment is available for large-scale manufacturing of such capsules. Some practical limitations of dosage form development might be the inadequate solubility of drugs in carriers and the instability of drugs and carriers at elevated temperatures necessary to manufacture capsules. PMID- 10514357 TI - Role of P-glycoprotein-mediated secretion in absorptive drug permeability: An approach using passive membrane permeability and affinity to P-glycoprotein. AB - It has been shown in vivo and in vitro that P-glycoprotein (P-gp) may be able to influence the permeability of its substrates across biological membranes. However, the quantitative contribution of the secretion process mediated by P-gp on the overall permeability of membranes has not been determined yet. In particular, observations need to be clarified in which substrates showing high affinity to P-glycoprotein, e.g., verapamil, apparently do not seem to be greatly influenced by P-gp in their permeability and consequently also with respect to their extent of GI-absorption after oral administration, whereas weaker substrates of P-gp, e.g., talinolol, have clearly shown P-gp-related absorption phenomena such as nonlinear intestinal permeability and bioavailability. Experiments with Caco-2 cell monolayers and mathematical simulations based on a mechanistic permeation model should aid in clarifying the underlying mechanism for these observations and quantifying the influence of passive membrane permeability and affinity to P-gp to the overall transmembrane drug flux. In addition, the concentration range of drug at which P-glycoprotein-mediated transport across the biological membrane is relevant should be examined. The permeability of various drugs in Caco-2 monolayers was determined experimentally and modeled using a combination of passive absorptive membrane permeability and a Michaelis-Menten-type transport process in the secretory direction. The passive permeabilities were experimentally obtained for the apical and basolateral membrane by efflux experiments using Caco-2 monolayers in the presence of a P-gp inhibitor. The Michaelis-Menten parameters were determined by a newly developed radioligand-binding assay for the quantification of drug affinity to P-gp. The model was able to accurately simulate the permeability of P-glycoprotein substrates, with differing passive membrane permeabilities and P-glycoprotein affinities. Using the outlined approach, permeability vs donor-concentration profiles were calculated, and the relative contribution of passive and active transport processes to the overall membrane permeability was evaluated. A model is presented to quantitatively describe and predict direction-dependent drug fluxes in Caco-2 monolayers by knowing the affinity of a compound to the exsorptive transporter P-gp and its passive membrane permeability. It was shown that a combination of high P-gp affinity with good passive membrane permeability, e.g., in the case of verapamil, will readily compensate for the P-gp-mediated reduction of intestinal permeability, resulting in a narrow range in which the permeability depends on the apical drug concentration. On the other hand, the permeability of compounds with low passive membrane permeability (e. g., talinolol) might be affected over a wide concentration range despite low affinity to P-gp. PMID- 10514358 TI - Chemical stability of peptides in polymers. 1. Effect of water on peptide deamidation in poly(vinyl alcohol) and poly(vinyl pyrrolidone) matrixes. AB - This paper examines the effect of water content, water activity, and glass transition temperature (T(g)) on the deamidation of an asparagine-containing hexapeptide (VYPNGA; Asn-hexapeptide) in lyophilized poly(vinyl alcohol) (PVA) and poly(vinyl pyrrolidone) (PVP) at 50 degrees C. The rate of Asn-hexapeptide deamidation increases with increasing water content or water activity and, hence, decreasing T(g). The rate of deamidation is more sensitive to changes in these parameters in PVA than in PVP. Deamidation is clearly evident in the glassy state in both formulations. In the glassy state, the peptide is more stable in PVA than in PVP formulations but is less stable in the rubbery state. No single variable (water content, water activity, or T(g)) could account for the variation in deamidation rates in PVA and PVP formulations. Deamidation rates were correlated with the degree of plasticization by water (distance of T(g) from the dry intrinsic glass transition temperature); coincident curves for the two polymers were obtained with this correlation. Deamidation in PVA and PVP was closely correlated with the extent of water-induced plasticization experienced by the formulation relative to its glass transition at 50 degrees C, suggesting that the physical state of formulations could be used to predict chemical stability. PMID- 10514359 TI - Chemical stability of peptides in polymers. 2. Discriminating between solvent and plasticizing effects of water on peptide deamidation in poly(vinylpyrrolidone). AB - The mechanistic role of water in the deamidation of a model asparagine-containing hexapeptide (Val-Tyr-Pro-Asn-Gly-Ala) in lyophilized formulations containing poly(vinylpyrrolidone) (PVP) and glycerol was investigated. Glycerol was used as a plasticizer to vary formulation glass transition temperature (T(g)) without significantly changing water content or activity. Increases in moisture and glycerol contents increased the rate of peptide deamidation. This increase was strongly correlated with T(g) at constant water content and activity, suggesting that increased matrix mobility facilitates deamidation. In rubbery systems (T > T(g)), deamidation rates appeared to be independent of water content and activity in formulations with similar T(g)s. However, in glassy formulations with similar T(g)s, deamidation increased with water content, suggesting a solvent/medium effect of water on reactivity in this regime. An increase in water content also affected the degradation product distribution; less of the cyclic imide intermediate and more of the hydrolytic products, isoAsp- and Asp-hexapeptides, were observed as water content increased. Thus, residual water appears to facilitate deamidation in these solid PVP formulations both by enhancing molecular mobility and by solvent/medium effects, and also participates as a chemical reactant in the subsequent breakdown of the cyclic imide. PMID- 10514360 TI - Mechanisms of solvent evaporation encapsulation processes: prediction of solvent evaporation rate. AB - The mechanism of organic solvent evaporation during microencapsulation and its role during microsphere hardening has been investigated. Evaporation and encapsulation studies were carried out in a jacketed beaker, filled with aqueous hardening solution, which was maintained at constant temperature and constant stirring rate in the turbulent regime. Evaporation of dissolved methylene chloride (MC), ethyl acetate (EA), and acetonitrile (ACN) was examined by the decline in organic solvent concentration in the hardening bath, which was monitored by gas chromatography. The evaporation from the bath followed first order kinetics under dilute conditions (e.g., MC < 3 mg/mL), yielding an overall permeability coefficient, P. The value of P was theoretically related to the Kolmogorov length-scale of turbulence under conditions that favor liquid-side transport control. According to theory, factors that favored liquid-phase control (as opposed to gas-phase control) were those that favored a high Henry's law constant [i.e., elevated temperature near the normal boiling point (bp) of the organic solvent] and properties of the dissolved organic solvent (i.e., low normal bp and low aqueous solubility). These theoretical hypotheses were confirmed by (1) correlating the experimentally determined P with process variables raised to the appropriate power according to theory, r(2) = 0.95 (i.e., P approximately rotational speed, omega(3/4), impeller diameter, d (5/4), volume of hardening bath, V(-1/4), and the product of kinematic viscosity and diffusion coefficient, nu(-5/12)D (2/3)), and (2) illustrating that at constant temperature, the tendency of the evaporation system to obey liquid-side transport control follows the same order of increasing Henry's law constant (i.e., MC > EA > ACN). To establish the relationship of evaporation with microsphere hardening, the decline in MC concentration was determined in both the continuous and dispersed polymer phases during microencapsulation. By applying a mass balance with respect to MC in the hardening bath, the cumulative hardening profile of the microspheres was accurately predicted from the interpolating functions of the kinetics of MC loss from the bath with and without polymer added. These results have potential use for microsphere formulation, design of encapsulation apparatus, and scale up of microsphere production. PMID- 10514361 TI - Some relationships between the physical properties of various 3-acyloxymethyl prodrugs of phenytoin to structure: potential in vivo performance implications PMID- 10514363 TI - Room-Temperature Magnetic Bistability in Organic Radical Crystals. AB - A large first-order magnetic phase transition in an organic radical, 1,3,5 trithia-2,4,6-triazapentalenyl, is described. The transition occurs with a wide thermal hysteresis loop over the temperature range 230 to 305 kelvin. The high temperature phase is paramagnetic, and its structure consists of a uniform one dimensional stacking of the radical. The low-temperature phase is diamagnetic because of strong dimerization along the stacking direction. The results may have applications in thermal sensors, switching units, and information storage media based on organic radical crystals. PMID- 10514362 TI - Structural changes in bacteriorhodopsin during ion transport at 2 angstrom resolution. AB - Crystal structures of the Asp96 to Asn mutant of the light-driven proton pump bacteriorhodopsin and its M photointermediate produced by illumination at ambient temperature have been determined to 1.8 and 2.0 angstroms resolution, respectively. The trapped photoproduct corresponds to the late M state in the transport cycle-that is, after proton transfer to Asp85 and release of a proton to the extracellular membrane surface, but before reprotonation of the deprotonated retinal Schiff base. Its density map describes displacements of side chains near the retinal induced by its photoisomerization to 13-cis,15-anti and an extensive rearrangement of the three-dimensional network of hydrogen-bonded residues and bound water that accounts for the changed pKa values (where Ka is the acid constant) of the Schiff base and Asp85. The structural changes detected suggest the means for conserving energy at the active site and for ensuring the directionality of proton translocation. PMID- 10514364 TI - Gate-Controlled Superconducting Proximity Effect in Carbon Nanotubes. AB - The superconducting proximity effect in single-walled carbon nanotubes connected to niobium electrodes was controlled with the use of nearby gates that tune the niobium-nanotube transparency. At 4.2 kelvin, when the transparency was tuned to be high, a dip in the low-bias differential resistance was observed, indicating a proximity effect mediated by Andreev reflection. When the transparency was tuned to be low, signatures of Andreev reflection disappeared and only tunneling conduction was observed. Below approximately 4 kelvin, a narrow peak in differential resistance around zero bias appeared superimposed on the Andreev dip, probably as a result of electron-electron interaction competing with the proximity effect. PMID- 10514365 TI - Evidence for One-Dimensional Charge Transport in La(2-x-y)Nd(y)Sr(x)CuO(4). AB - Doping dependences of the resistivity and the Hall coefficient are presented for neodymium-doped lanthanum strontium cuprate (La(1.4-x)Nd(0.6)Sr(x)CuO(4)) in the static spin-charge stripe ordered phase. For doping concentration x 1/8, the Hall coefficient remains relatively large in the ordered phase. The results indicate a crossover from one- to two-dimensional charge transport taking place at x = 1/8. PMID- 10514366 TI - One-Dimensional Electronic Structure and Suppression of d-Wave Node State in (La(1.28)Nd(0.6)Sr(0.12))CuO(4). AB - Angle-resolved photoemission spectroscopy was carried out on (La(1.28)Nd(0.6) Sr(0.12))CuO(4), a model system of the charge- and spin-ordered state, or stripe phase. The electronic structure contains characteristic features consistent with other cuprates, such as the flat band at low energy near the Brillouin zone face. However, the low-energy excitation near the expected d-wave node region is strongly suppressed. The frequency-integrated spectral weight is confined inside one-dimensional segments in the momentum space (defined by horizontal momenta &cjs3539;k(x)&cjs3539; = pi/4 and vertical momenta &cjs3539;k(y)&cjs3539; = pi/4), deviating strongly from the more rounded Fermi surface expected from band calculations. This departure from the two-dimensional Fermi surface persists to a very high energy scale. These results provide important information for establishing a theory to understand the charge and spin ordering in cuprates and their relation with high-temperature superconductivity. PMID- 10514367 TI - Evidence of Nonlinear Elasticity of the Crust from the Mw7.6 Manyi (Tibet) Earthquake. AB - Satellite synthetic aperture radar (SAR) interferometry shows that the magnitude 7.6 Manyi earthquake of 8 November 1997 produced a 170-kilometer-long surface break with up to 7 meters of left-lateral slip, reactivating a N76 degrees E quaternary fault in western Tibet. The radar interferometric map reveals asymmetric, along-strike displacement profiles between the two sides of the surface rupture, a pattern that cannot be explained with linear elastic theory. This observation suggests that the elastic moduli of the crust in tension and in compression are different because of the presence of cracks in the crust at shallow depth. A model indicates that a ratio of 2 between compressive and tensile elastic moduli can account for the observed asymmetry, a ratio that is consistent with laboratory and borehole measurements. PMID- 10514368 TI - Measurements of Past Ice Sheet Elevations in Interior West Antarctica. AB - A lateral moraine band on Mount Waesche, a volcanic nunatak in Marie Byrd Land, provides estimates of past ice sheet surface elevations in West Antarctica. Helium-3 and chlorine-36 surface exposure ages indicate that the proximal part of the moraine, up to 45 meters above the present ice surface, was deposited about 10,000 years ago, substantially later than the maximum ice extent in the Ross Embayment. The upper distal part of the moraine may record multiple earlier ice sheet high stands. A nonequilibrium ice sheet model predicts a delay of several thousand years in maximum ice levels at Mount Waesche relative to the maximum ice extent in the Ross Sea. The glacial geologic evidence, coupled with the ice sheet model, indicates that the contribution of the Ross Sea sector of the West Antarctic Ice Sheet to Holocene sea level rise was only about 3 meters. These results eliminate West Antarctic ice as the principle source of the large meltwater pulse during the early Holocene. PMID- 10514369 TI - Past and Future Grounding-Line Retreat of the West Antarctic Ice Sheet. AB - The history of deglaciation of the West Antarctic Ice Sheet (WAIS) gives clues about its future. Southward grounding-line migration was dated past three locations in the Ross Sea Embayment. Results indicate that most recession occurred during the middle to late Holocene in the absence of substantial sea level or climate forcing. Current grounding-line retreat may reflect ongoing ice recession that has been under way since the early Holocene. If so, the WAIS could continue to retreat even in the absence of further external forcing. PMID- 10514370 TI - Tributaries of West Antarctic Ice Streams Revealed by RADARSAT Interferometry. AB - Interferometric RADARSAT data are used to map ice motion in the source areas of four West Antarctic ice streams. The data reveal that tributaries, coincident with subglacial valleys, provide a spatially extensive transition between slow inland flow and rapid ice stream flow and that adjacent ice streams draw from shared source regions. Two tributaries flow into the stagnant ice stream C, creating an extensive region that is thickening at an average rate of 0.49 meters per year. This is one of the largest rates of thickening ever reported in Antarctica. PMID- 10514371 TI - X-ray crystallographic structure of the Norwalk virus capsid. AB - Norwalk virus, a noncultivatable human calicivirus, is the major cause of epidemic gastroenteritis in humans. The first x-ray structure of a calicivirus capsid, which consists of 180 copies of a single protein, has been determined by phase extension from a low-resolution electron microscopy structure. The capsid protein has a protruding (P) domain connected by a flexible hinge to a shell (S) domain that has a classical eight-stranded beta-sandwich motif. The structure of the P domain is unlike that of any other viral protein with a subdomain exhibiting a fold similar to that of the second domain in the eukaryotic translation elongation factor-Tu. This subdomain, located at the exterior of the capsid, has the largest sequence variation among Norwalk-like human caliciviruses and is likely to contain the determinants of strain specificity and cell binding. PMID- 10514372 TI - Crystal structure of invasin: a bacterial integrin-binding protein. AB - The Yersinia pseudotuberculosis invasin protein promotes bacterial entry by binding to host cell integrins with higher affinity than natural substrates such as fibronectin. The 2.3 angstrom crystal structure of the invasin extracellular region reveals five domains that form a 180 angstrom rod with structural similarities to tandem fibronectin type III domains. The integrin-binding surfaces of invasin and fibronectin include similarly located key residues, but in the context of different folds and surface shapes. The structures of invasin and fibronectin provide an example of convergent evolution, in which invasin presents an optimized surface for integrin binding, in comparison with host substrates. PMID- 10514373 TI - Evolutionarily conserved pathways of energetic connectivity in protein families. AB - For mapping energetic interactions in proteins, a technique was developed that uses evolutionary data for a protein family to measure statistical interactions between amino acid positions. For the PDZ domain family, this analysis predicted a set of energetically coupled positions for a binding site residue that includes unexpected long-range interactions. Mutational studies confirm these predictions, demonstrating that the statistical energy function is a good indicator of thermodynamic coupling in proteins. Sets of interacting residues form connected pathways through the protein fold that may be the basis for efficient energy conduction within proteins. PMID- 10514374 TI - Mimicry of CD40 signals by Epstein-Barr virus LMP1 in B lymphocyte responses. AB - The effect of the Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) on the activation and differentiation of normal B cells was investigated. B cells of transgenic mice expressing LMP1 under the control of immunoglobulin promoter/enhancer displayed enhanced expression of activation antigens and spontaneously proliferated and produced antibody. Humoral immune responses of LMP1 transgenic mice in CD40-deficient or normal backgrounds revealed that LMP1 mimics CD40 signals to induce extrafollicular B cell differentiation but, unlike CD40, blocks germinal center formation. Thus, these specific properties of LMP1 may determine the site of primary B cell infection and the state of infection in the natural course of EBV infection, whereas subsequent loss of LMP1 expression may affect the site of persistent latent infection. PMID- 10514375 TI - A nonpeptidyl mimic of superoxide dismutase with therapeutic activity in rats. AB - Many human diseases are associated with the overproduction of oxygen free radicals that inflict cell damage. A manganese(II) complex with a bis(cyclohexylpyridine)-substituted macrocyclic ligand (M40403) was designed to be a functional mimic of the superoxide dismutase (SOD) enzymes that normally remove these radicals. M40403 had high catalytic SOD activity and was chemically and biologically stable in vivo. Injection of M40403 into rat models of inflammation and ischemia-reperfusion injury protected the animals against tissue damage. Such mimics may result in better clinical therapies for diseases mediated by superoxide radicals. PMID- 10514376 TI - Anaerobic microbes: oxygen detoxification without superoxide dismutase. AB - Superoxide reductase from the hyperthermophilic anaerobe Pyrococcus furiosus uses electrons from reduced nicotinamide adenine dinucleotide phosphate, by way of rubredoxin and an oxidoreductase, to reduce superoxide to hydrogen peroxide, which is then reduced to water by peroxidases. Unlike superoxide dismutase, the enzyme that protects aerobes from the toxic effects of oxygen, SOR does not catalyze the production of oxygen from superoxide and therefore confers a selective advantage on anaerobes. Superoxide reductase and associated proteins are catalytically active 80 degrees C below the optimum growth temperature (100 degrees C) of P. furiosus, conditions under which the organism is likely to be exposed to oxygen. PMID- 10514377 TI - The tyrosine kinase negative regulator c-Cbl as a RING-type, E2-dependent ubiquitin-protein ligase. AB - Ubiquitination of receptor protein-tyrosine kinases (RPTKs) terminates signaling by marking active receptors for degradation. c-Cbl, an adapter protein for RPTKs, positively regulates RPTK ubiquitination in a manner dependent on its variant SRC homology 2 (SH2) and RING finger domains. Ubiquitin-protein ligases (or E3s) are the components of ubiquitination pathways that recognize target substrates and promote their ligation to ubiquitin. The c-Cbl protein acted as an E3 that can recognize tyrosine-phosphorylated substrates, such as the activated platelet derived growth factor receptor, through its SH2 domain and that recruits and allosterically activates an E2 ubiquitin-conjugating enzyme through its RING domain. These results reveal an SH2-containing protein that functions as a ubiquitin-protein ligase and thus provide a distinct mechanism for substrate targeting in the ubiquitin system. PMID- 10514378 TI - Congenital nephrotic syndrome in mice lacking CD2-associated protein. AB - CD2-associated protein (CD2AP) is an 80-kilodalton protein that is critical for stabilizing contacts between T cells and antigen-presenting cells. In CD2AP deficient mice, immune function was compromised, but the mice died at 6 to 7 weeks of age from renal failure. In the kidney, CD2AP was expressed primarily in glomerular epithelial cells. Knockout mice exhibited defects in epithelial cell foot processes, accompanied by mesangial cell hyperplasia and extracellular matrix deposition. Supporting a role for CD2AP in the specialized cell junction known as the slit diaphragm, CD2AP associated with nephrin, the primary component of the slit diaphragm. PMID- 10514379 TI - Coordinated polar localization of auxin efflux carrier PIN1 by GNOM ARF GEF. AB - The plant hormone auxin is transported in a polar manner along the shoot-root axis, which requires efflux carriers such as PIN1. Asymmetric localization of PIN1 develops from a random distribution in Arabidopsis early embryogenesis. Coordinated polar localization of PIN1 is defective in gnom embryos. GNOM is a membrane-associated guanine-nucleotide exchange factor on ADP-ribosylation factor G protein (ARF GEF). Thus, GNOM-dependent vesicle trafficking may establish cell polarity, resulting in polar auxin transport. PMID- 10514380 TI - NAB2: a transcriptional brake for activated gene expression in the vessel wall? PMID- 10514382 TI - Release of cytochrome c, Bax migration, Bid cleavage, and activation of caspases 2, 3, 6, 7, 8, and 9 during endothelial cell apoptosis. AB - Although the executioner phase of apoptosis has been well defined in many cell types, the subcellular events leading to apoptosis in endothelial cells remain undefined. In the current study, apoptosis was induced in primary human umbilical venous endothelial cells by the photosensitizer verteporfin and light. Release of mitochondrial cytochrome c into the cytosol was detectable immediately and accumulated over 2 hours after treatment while cytosolic levels of the proapoptotic Bcl-2 family member, Bax, decreased reciprocally over the same time period. Cleavage of another proapoptotic Bcl-2 family member, Bid, was observed by 2 hours after treatment. Although Bid cleavage has been shown to occur as an upstream event responsible for inducing cytochrome c release, we demonstrate that Bid cleavage can also occur after cytochrome c release. Activation of caspases 2, 3, 6, 7, 8, and 9 occurred following the release of cytochrome c, and cleavage of downstream substrates was observed. In summary, endothelial cell death involves the cellular redistribution of Bax and cytochrome c, followed by the activation of multiple caspases which manifest the apoptotic phenotype. PMID- 10514381 TI - Endothelial ligands for L-selectin: from lymphocyte recirculation to allograft rejection. PMID- 10514383 TI - Expression of Bcl-2 and amplification of c-myc are frequent in basaloid squamous cell carcinomas of the esophagus. AB - Basaloid squamous cell carcinoma (BSCC) of the esophagus is a rare, poorly differentiated variant of typical esophageal squamous cell carcinoma (SCC) characterized by high proliferative activity and frequent spontaneous apoptoses. In the present study, we investigated the expression of the apoptosis-suppressing protein Bcl-2 in 23 BSCC of the esophagus and 23 stage-matched typical esophageal SCC by means of immunohistochemistry. In addition, amplification of the apoptosis and proliferation-inducing gene c-myc was determined by means of differential polymerase chain reaction. Bcl-2 expression was found significantly more often in BSCC than in SCC (86.9% vs. 17.4%, P < 0.0001). Amplification of c-myc was nearly twice as common in BSCC as in SCC (47.8% vs. 26.1%, not significant). Bcl-2 protein expression together with c-myc amplification was detected in 43.5% of the BSCC but in none of the typical SCC (P < 0.0001). Taken together, our findings indicate that the molecular pathogenesis of esophageal BSCC differs from that of typical SCC and frequently involves coactivation of c-myc and Bcl-2. PMID- 10514384 TI - beta-catenin regulates the expression of the matrix metalloproteinase-7 in human colorectal cancer. AB - Most colorectal cancers have loss of function mutations in the adenomatosis polyposis coli (APC) tumor suppressor gene. This leads to accumulation of beta catenin, which together with the DNA binding protein TCF-4 functions as a transcriptional activator. Recently defined target genes are c-myc and cyclin D1, linking the APC gene defect to the capacity for autonomous proliferation of colon tumors. Here we report the identification of the matrix metalloproteinase MMP-7 as another target gene of beta-catenin/TCF-4. MMP-7 is overexpressed in 80% of human colorectal cancers and known to be an important factor for early tumor growth, with a potential function also for later progression steps, like invasion and metastasis. Our results explain the high percentage of MMP-7 overexpression in colon tumors. Moreover they indicate that defects in the APC tumor suppressor gene may also have an influence on later steps of colon tumor progression. PMID- 10514385 TI - Analysis of genomic alterations in sporadic adrenocortical lesions. Gain of chromosome 17 is an early event in adrenocortical tumorigenesis. AB - Genetic changes underlying the tumorigenesis of sporadic adrenocortical tumors are poorly characterized. To search for characteristic genomic imbalances involved in adrenocortical tumors, we examined 41 adrenocortical lesions (12 carcinomas, 23 adenomas, and 6 hyperplasias) by comparative genomic hybridization. Our results show that genetic alterations are more frequent in malignant than in benign lesions and that they rarely occur in hyperplastic lesions. The most frequent DNA copy number changes in adrenocortical carcinomas included losses of 1p21-31, 2q, 3p, 3q, 6q, 9p, and 11q14-qter, as well as gains and amplifications of 5q12, 9q32-qter, 12q, and 20q. The genomic aberrations prevalently occurring in adrenocortical adenomas were gains of 17q, 17p, and 9q32 qter. Gains found in 2 of 6 adrenocortical hyperplastic lesions involved chromosome 17 or 17q only. These data indicate that oncogenes determining the early tumorigenesis of adrenocortical tumors may exist on chromosome 17 and that the number of genomic alterations is closely associated with tumor behavior in adrenocortical tumors. PMID- 10514387 TI - Damage to the enteric nervous system in experimental colitis. AB - Inflammation of the intestine causes pain and altered motility, at least in part through effects on the enteric nervous system. While these changes may be reversed with healing, permanent damage may contribute to inflammatory bowel disease (IBD) and post-enteritis irritable bowel syndrome. Since little information exists, we induced colitis in male Sprague-Dawley rats with dinitrobenzene sulfonic acid and used immunocytochemistry to examine the number and distribution of enteric neurons at times up to 35 days later. Inflammation caused significant neuronal loss in the inflamed region by 24 hours, with only 49% of neurons remaining by days 4 to 6 and thereafter, when inflammation had subsided. Eosinophils were found within the myenteric plexus at only at the earliest time points, despite a general infiltration of neutrophils into the muscle wall. While the number of myenteric ganglia remained constant, there was significant decrease in the number of ganglia in the submucosal plexus. Despite reduced neuronal number and hyperplasia of smooth muscle, the density of axons among the smooth muscle cells remained unchanged during and after inflammation. Intracolonic application of the topical steroid budesonide caused a dose dependent prevention of neuronal loss, suggesting that evaluation of anti inflammatory therapy in inflammatory bowel disease should include quantitative assessment of neural components. PMID- 10514386 TI - Immunohistochemical analysis of uroplakins, urothelial specific proteins, in ovarian Brenner tumors, normal tissues, and benign and neoplastic lesions of the female genital tract. AB - Uroplakins are the characteristic integral membrane proteins in terminally differentiated, superficial urothelial asymmetric unit membrane. Brenner tumors of the ovary and Walthard cell nests of Fallopian tubes have been considered to represent urothelial differentiation in the female genital tract, but no definitive differentiation marker has been demonstrated supporting such a conclusion. An immunohistochemical analysis was performed to assess the expression of uroplakins in these lesions as well as in various benign and neoplastic lesions and normal tissues of the female genital tract. Focal expression of uroplakins was observed on the luminal surface of ovarian Brenner tumor cells forming microcysts in all 5 cases examined. In contrast, uroplakins were slightly expressed in only 1 of 12 cases of Walthard cell nests, even in the presence of microcyst formation. Uroplakins were not expressed in other benign or malignant lesions or normal tissues of the female genital tract. These results support the hypothesis that the Brenner tumor and possibly Walthard cell nests represent urothelial (transitional cell) differentiation. PMID- 10514388 TI - IL-13 activates STAT6 and inhibits liver injury induced by ischemia/reperfusion. AB - Hepatic ischemia/reperfusion injury is initiated by the activation of Kupffer cells and their subsequent release of proinflammatory mediators, including tumor necrosis factor-alpha (TNFalpha). These mediators stimulate a cascade of events including up-regulation of CXC chemokines and vascular endothelial adhesion molecules, leading to hepatic neutrophil recruitment and tissue injury. Interleukin-13 (IL-13) is a cytokine that has been shown to suppress macrophage production of proinflammatory mediators. The objective of the current study was to determine whether IL-13 could regulate the liver inflammatory injury induced by ischemia and reperfusion. C57BL/6 mice underwent 90 minutes of partial hepatic ischemia followed by reperfusion with or without intravenous administration of recombinant murine IL-13. Hepatic ischemia/reperfusion increased expression of TNFalpha and macrophage inflammatory protein-2 (MIP-2), leading to hepatic neutrophil recruitment, hepatocellular injury, and liver edema. Administration of IL-13 reduced the production of TNFalpha and MIP-2 mRNA and protein. IL-13 suppressed liver neutrophil recruitment by up to 72% and hepatocellular injury and liver edema were each reduced by >60%. Administration of IL-13 had no effect on liver NFkappaB activation, but greatly increased the activation of STAT6. The data suggest that the hepatoprotective effects of IL-13 may be a result of STAT6 activation. PMID- 10514389 TI - Hepatocellular hypoxia-induced vascular endothelial growth factor expression and angiogenesis in experimental biliary cirrhosis. AB - We tested the potential role of vascular endothelial growth factor (VEGF) and of fibroblast growth factor-2 (FGF-2) in the angiogenesis associated with experimental liver fibrogenesis induced by common bile duct ligation in Sprague Dawley rats. In normal rats, VEGF and FGF-2 immunoreactivities were restricted to less than 3% of hepatocytes. One week after bile duct ligation, hypoxia was demonstrated by the immunodetection of pimonidazole adducts unevenly distributed throughout the lobule. After 2 weeks, hypoxia and VEGF expression were detected in >95% of hepatocytes and coexisted with an increase in periportal vascular endothelial cell proliferation, as ascertained by Ki67 immunolabeling. Subsequently, at 3 weeks the density of von Willebrand-labeled vascular section in fibrotic areas significantly increased. Semiquantitative reverse transcription polymerase chain reaction showed that VEGF(120) and VEGF(164) transcripts, that correspond to secreted isoforms, increased within 2 weeks, while VEGF(188) transcripts remained unchanged. FGF-2 mainly consisting of a 22-kd isoform, according to Western blot, was identified by immunohistochemistry in 49% and 100% of hepatocytes at 3 and 7 weeks, respectively. Our data provide evidence that in biliary-type liver fibrogenesis, angiogenesis is stimulated primarily by VEGF in response to hepatocellular hypoxia while FGF-2 likely contributes to the maintenance of angiogenesis at later stages. PMID- 10514390 TI - Modulation of the gene network connected to interferon-gamma in liver regeneration from oval cells. AB - Suppression subtractive hybridization was used to clone genes associated with proliferation of oval cells in rat liver regenerating after a 70% partial hepatectomy combined with the feeding of 2-acetylaminofluorene. A subset of the identified genes comprised interferon-gamma receptor alpha subunit (IFN gammaRalpha), gp91phox, interleukin-1beta (IL-1beta), lymphocyte function associated molecule-1alpha (LFA-1), eukaryotic initiation factor-2-associated 67 kd protein (eIF-2-associated 67-kd protein), and alpha-fetoprotein, which constitute part of the cellular program modulated by IFN-gamma. Therefore, expression analysis performed by Northern blotting and immunohistochemistry were extended to include IFN-gamma, the IFN-gamma receptor beta subunit (IFN gammaRbeta), three secondary response genes induced by interaction of IFN-gamma with IFN-gamma receptor complexes, ie, IL-1beta-converting enzyme (ICE), intercellular adhesion molecule-1 (ICAM-1), and urokinase-type plasminogen activator receptor (uPAR), and a cytokine inducing IFN-gamma expression, ie, interleukin-18 (IL-18). The Northern blot analysis showed that all examined genes were modulated when progenitor-like oval cells were activated and recruited for liver regeneration. Immunohistochemistry localized the subunits of the IFN-gamma receptor complex, IFN-gammaRalpha and IFN-gammaRbeta, the secondary response genes uPAR and ICAM-1, the IFN-gamma-inducing factor IL-18, and ICE to the ductular structures of oval cells. In contrast, during liver regeneration after a 70% partial hepatectomy, only modulation of IL-1beta and ICE was observed. Our results, therefore, indicate that IFN-gamma-mediated events may be particularly important when cells in the bile ductules must respond to liver damage by production of ductular oval cells. PMID- 10514391 TI - Activation of pancreatic stellate cells in human and experimental pancreatic fibrosis. AB - The mechanisms of pancreatic fibrosis are poorly understood. In the liver, stellate cells play an important role in fibrogenesis. Similar cells have recently been isolated from the pancreas and are termed pancreatic stellate cells. The aim of this study was to determine whether pancreatic stellate cell activation occurs during experimental and human pancreatic fibrosis. Pancreatic fibrosis was induced in rats (n = 24) by infusion of trinitrobenzene sulfonic acid (TNBS) into the pancreatic duct. Surgical specimens were obtained from patients with chronic pancreatitis (n = 6). Pancreatic fibrosis was assessed using the Sirius Red stain and immunohistochemistry for collagen type I. Pancreatic stellate cell activation was assessed by staining for alpha-smooth muscle actin (alphaSMA), desmin, and platelet-derived growth factor receptor type beta (PDGFRbeta). The relationship of fibrosis to stellate cell activation was studied by staining of serial sections for alphaSMA, desmin, PDGFRbeta, and collagen, and by dual-staining for alphaSMA plus either Sirius Red or in situ hybridization for procollagen alpha(1) (I) mRNA. The cellular source of TGFbeta was examined by immunohistochemistry. The histological appearances in the TNBS model resembled those found in human chronic pancreatitis. Areas of pancreatic fibrosis stained positively for Sirius Red and collagen type I. Sirius Red staining was associated with alphaSMA-positive cells. alphaSMA staining colocalized with procollagen alpha(1) (I) mRNA expression. In the rat model, desmin staining was associated with PDGFRbeta in areas of fibrosis. TGFbeta was maximal in acinar cells adjacent to areas of fibrosis and spindle cells within fibrotic bands. Pancreatic stellate cell activation is associated with fibrosis in both human pancreas and in an animal model. These cells appear to play an important role in pancreatic fibrogenesis. PMID- 10514392 TI - Increased mucosal production of monomeric IgA1 but no IgA1 protease activity in Helicobacter pylori gastritis. AB - Immunoglobulin A and IgM are subjected to epithelial transport only when they are produced as polymers with incorporated J chain. Immunocytes containing various Ig isotypes and associated J chain in gastric mucosa, as well as IgA-degrading protease activity in Helicobacter pylori cultures, were examined. Gastric body specimens from 15 H. pylori-positive and 14 H. pylori-negative patients were studied by paired immunofluorescence for IgA, IgA1, IgA2, IgG, or IgM and concurrent cellular J chain. H. pylori isolates were incubated with IgA1 or secretory IgA and examined by immunoelectrophoresis for cleavage products. A substantial increase of Ig-producing cells occurred in chronic gastritis, particularly in the IgA1 isotype, but H. pylori was shown to possess neither IgA1 specific nor nonspecific IgA-degrading protease activity. Regardless of infection status, reduced J chain expression was observed for all immunocyte isotypes (except for IgM) in inflamed compared with normal gastric body mucosa, the median positivity for IgA1 cells being reduced to 58.7% versus 87.9% (P = 0.0002), and for IgA2 cells to 48.9% versus 87.8% (P = 0.0002). This down-regulation of the J chain suggested that a large fraction of IgA monomers is produced in gastritis. PMID- 10514394 TI - Demonstration of urokinase expression in cancer cells of colon adenocarcinomas by immunohistochemistry and in situ hybridization. AB - The question whether urokinase is expressed in human colon cancer by the cancer cells themselves or by surrounding stromal elements such as fibroblasts, macrophages, and leukocytes, which transfer the activator to the receptors of the cancer cells, has been a controversial one. In the present study 12 cases of colorectal cancer were investigated by immunohistochemical methods using three monoclonal antibodies of different specificity against urokinase. Cytoplasmic staining of strongly varying intensity was observed in all cases, with the antigen expressed most strongly in the apical and the basal regions of the cancer cells. In some cases, staining was also found in stromal elements surrounding the cancer glands. That the activator was indeed the product of the cancer cells was demonstrated by in situ hybridization using a uPA-cDNA probe, which detected the presence of uPA-mRNA in both the basal and the apical regions of the cancer cells. A monoclonal antibody against the receptor for uPA showed similar localization. These findings indicate that the activator is expressed by the cancer cells and is not recruited by them from stromal elements. PMID- 10514393 TI - Overexpression of pterin-4a-carbinolamine dehydratase/dimerization cofactor of hepatocyte nuclear factor 1 in human colon cancer. AB - Pterin-4a-carbinolamine dehydratase (PCD) is a bifunctional protein also known as DCoH (dimerization co-factor of hepatocyte nuclear factor 1 (HNF1)). PCD/DCoH modulates the DNA binding specificity of HNF1, thus acting on its transcriptional activity. In addition, it participates in the recycling of tetrahydrobiopterin (BH(4)), an essential cofactor of several metabolic reactions. We investigated colorectal tumors and colorectal tumor cell lines as compared to normal colon samples in search of a potential differential expression of PCD/DCoH. Immunohistochemistry was conducted on 20 human colorectal tumors and 20 normal samples using a specific polyclonal antibody. Immunoblotting and RT-PCR analysis for PCD/DCoH and HNF1 were also performed on both human tissues and CACO-2 and HT 29 cell lines. All of the 20 tumors and both colon cancer cell lines presented a strong and widespread immunoreactivity for PCD/DCoH, contrasting with the absence of expression in the normal epithelia. We thus report the massive overexpression of PCD/DCoH in colon tumors, which is in striking contrast with the absence of staining in normal counterparts. The sharp contrast in the expression of a modulator of transcriptional activity between tumoral and normal cells may have a physiopathological role. PCD/DCoH could potentially be a new marker of malignant colon cells in vivo. PMID- 10514395 TI - Different responses of epidermal and hair follicular cells to radiation correlate with distinct patterns of p53 and p21 induction. AB - Different parts of the skin respond to ionizing radiation with different sensitivities. To examine the mechanisms underlying these different responses, we investigated various cellular parameters in the skin after exposure of mice to 5 Gy of ionizing radiation. Epidermal cells responded to radiation by undergoing growth arrest, whereas the cells in the matrix of hair follicles underwent apoptosis but not growth arrest. These distinct responses correlated with differential increases in p53 and p21 proteins in these two populations of cells; whereas an increase in p53 protein levels was observed in both epidermis and hair follicular matrix, especially in the latter, the induction of p21 was strong in the epidermis but absent in the follicular matrical cells. Studies using p53-null and p21-null mice demonstrated that the radiation-induced apoptosis in the hair follicles was fully dependent on p53, and growth arrest in the epidermis was only partially dependent on p53 but fully dependent on p21. These results indicate that two epithelial cell types respond to radiation by different pathways that are governed in part by the differential p53- and p21-dependent responses of these cells; high-level induction of p53 in the absence of p21 induction led to apoptosis, whereas intermediate induction of both p53 and p21 led to growth arrest. PMID- 10514396 TI - Loss of cell cycle regulators p27(Kip1) and cyclin E in transitional cell carcinoma of the bladder correlates with tumor grade and patient survival. AB - The cyclin-dependent kinase inhibitor p27(Kip1) is a powerful molecular determinant of cell cycle progression. Loss of expression of p27(Kip1) has been shown to be predictive of disease progression in several human malignancies. In this study we investigated the expression of two key cell cycle regulators, p27(Kip1) and cyclin E, in the progression of transitional cell carcinoma of the bladder. An immunohistochemical analysis was conducted in a series of 50 bladder tumor specimens, including 3 metastatic lymph nodes, and 7 normal bladder specimens, using specific antibodies against the two regulators of the cell cycle, p27(Kip1) and cyclin E. The degree of immunoreactivity was correlated with the pathological tumor grade, stage, and patient survival. A uniformly intense immunoreactivity for p27(Kip1) and cyclin E was observed in epithelial cells of normal bladder tissue. Malignant bladder tissue demonstrated a heterogeneous pattern of significantly reduced p27(Kip1) and cyclin E immunoreactivity, compared with normal urothelium (P < 0.01). In addition, there was progressive loss of expression of both cell cycle proteins with increasing tumor grade and pathological stage. Expression of p27(Kip1) was significantly lower in the poorly differentiated tumors (grades III) compared to well and moderately differentiated (grades I and II) tumors (P = 0.004). Moreover, the expression of cyclin E was lower in grade III tumors compared to grade I and II lesions, although this difference failed to reach statistical significance. Most significantly, Kaplan Meier plots of patient survival show increased mortality risk associated with low levels of p27(Kip1) (P = 0.001) and cyclin E (P = 0.002) expression. This is the first evidence that loss of expression of p27(Kip1) and cyclin E in human bladder transitional cell carcinoma cells correlates with advancing histological aggressiveness and poor patient survival. These results have clinical importance, because they support a role for p27(Kip1) and cyclin E as novel predictive markers of the biological potential of bladder tumors that will enable identification of those tumors most likely to progress to muscle invasive disease and of patient survival. PMID- 10514397 TI - Topical estrogen accelerates cutaneous wound healing in aged humans associated with an altered inflammatory response. AB - The effects of intrinsic aging on the cutaneous wound healing process are profound, and the resulting acute and chronic wound morbidity imposes a substantial burden on health services. We have investigated the effects of topical estrogen on cutaneous wound healing in healthy elderly men and women, and related these effects to the inflammatory response and local elastase levels, an enzyme known to be up-regulated in impaired wound healing states. Eighteen health status-defined females (mean age, 74.4 years) and eighteen males (mean age, 70.7 years) were randomized in a double-blind study to either active estrogen patch or identical placebo patch attached for 24 hours to the upper inner arm, through which two 4-mm punch biopsies were made. The wounds were excised at either day 7 or day 80 post-wounding. Compared to placebo, estrogen treatment increased the extent of wound healing in both males and females with a decrease in wound size at day 7, increased collagen levels at both days 7 and 80, and increased day 7 fibronectin levels. In addition, estrogen enhanced the strength of day 80 wounds. Estrogen treatment was associated with a decrease in wound elastase levels secondary to reduced neutrophil numbers, and decreased fibronectin degradation. In vitro studies using isolated human neutrophils indicate that one mechanism underlying the altered inflammatory response involves both a direct inhibition of neutrophil chemotaxis by estrogen and an altered expression of neutrophil adhesion molecules. These data demonstrate that delays in wound healing in the elderly can be significantly diminished by topical estrogen in both male and female subjects. PMID- 10514398 TI - Age dependence of clinical and pathological manifestations of autoimmune demyelination. Implications for multiple sclerosis. AB - A prominent feature of the clinical spectrum of multiple sclerosis (MS) is its high incidence of onset in the third decade of life and the relative rarity of clinical manifestations during childhood and adolescence, features suggestive of age-related restriction of clinical expression. Experimental allergic encephalomyelitis (EAE), a model of central nervous system (CNS) autoimmune demyelination with many similarities to MS, has a uniform rapid onset and a high incidence of clinical and pathological disease in adult (mature) animals. Like MS, EAE is most commonly seen and studied in female adults. In this study, age related resistance to clinical EAE has been examined with the adoptive transfer model of EAE in SJL mice that received myelin basic protein-sensitized cells from animals 10 days (sucklings) to 12 weeks (young adults) of age. A variable delay before expression of clinical EAE was observed between the different age groups. The preclinical period was longest in the younger (<14 days of age) animals, and shortest in animals 6 to 8 weeks old at time of transfer. Young animals initially resistant to EAE eventually expressed well-developed clinical signs by 6 to 7 weeks of age. This was followed by a remitting, relapsing clinical course. For each age at time of sensitization, increased susceptibility of females compared to males was observed. Examination of the CNS of younger animal groups during the preclinical period showed lesions of acute EAE. Older age groups developed onset of signs coincident with acute CNS lesions. This age-related resistance to clinical EAE in developing mice is reminiscent of an age-related characteristic of MS previously difficult to study in vivo. The associated subclinical CNS pathology and age-related immune functions found in young animals may be relevant to the increasing clinical expression of MS with maturation, and may allow study of factors associated with the known occasional poor correlation of CNS inflammation and demyelination and clinical changes in this disease. PMID- 10514399 TI - A new molecular link between the fibrillar and granulovacuolar lesions of Alzheimer's disease. AB - Alzheimer's Disease (AD) is a progressive neurodegenerative disorder involving select neurons of the hippocampus, neocortex, and other regions of the brain. Markers of end stage disease include fibrillar lesions, which accumulate hyperphosphorylated tau protein polymerized into filaments, and granulovacuolar lesions, which appear primarily within the hippocampus. The mechanism by which only select populations of neurons develop these lesions as well as the relationship between them is unknown. To address these questions, we have turned to AD tissue to search for enzymes specifically involved in tau hyperphosphorylation. Recently, we showed that the principal phosphotransferases associated with AD brain-derived tau filaments are members of the casein kinase-1 (CK1) family of protein kinases. Here we report the distribution of three CK1 isoforms (Ckialpha, Ckidelta, and Ckiepsilon) in AD and control brains using immunohistochemistry and Western analysis. In addition to colocalizing with elements of the fibrillar pathology, CK1 is found within the matrix of granulovacuolar degeneration bodies. Furthermore, levels of all CK1 isoforms are elevated in the CA1 region of AD hippocampus relative to controls, with one isoform, Ckidelta, being elevated >30-fold. We propose that overexpression of this protein kinase family plays a key role in the hyperphosphorylation of tau and in the formation of AD-related pathology. PMID- 10514400 TI - Non-Abeta component of Alzheimer's disease amyloid (NAC) revisited. NAC and alpha synuclein are not associated with Abeta amyloid. AB - alpha-Synuclein (alphaSN), also termed the precursor of the non-Abeta component of Alzheimer's disease (AD) amyloid (NACP), is a major component of Lewy bodies and Lewy neurites pathognomonic of Parkinson's disease (PD) and dementia with Lewy bodies (DLB). A fragment of alphaSN termed the non-Abeta component of AD amyloid (NAC) had previously been identified as a constituent of AD amyloid plaques. To clarify the relationship of NAC and alphaSN with Abeta plaques, antibodies were raised to three domains of alphaSN. All antibodies produced punctate labeling of human cortex and strong labeling of Lewy bodies. Using antibodies to alphaSN(75-91) to label cortical and hippocampal sections of pathologically proven AD cases, we found no evidence for NAC in Abeta amyloid plaques. Double labeling of tissue sections in mixed DLB/AD cases revealed alphaSN in dystrophic neuritic processes, some of which were in close association with Abeta plaques restricted to the CA1 hippocampal region. In brain homogenates alphaSN was predominantly recovered in the cytosolic fraction as a 16-kd protein on Western analysis; however, significant amounts of aggregated and alphaSN fragments were also found in urea extracts of SDS-insoluble material from DLB and PD cases. NAC antibodies identified an endogenous fragment of 6 kd in the cytosolic and urea-soluble brain fractions. This fragment may be produced as a consequence of alphaSN aggregation or alternatively may accelerate aggregation of the full-length alphaSN. PMID- 10514401 TI - Abundant production of brain-derived neurotrophic factor by adult visceral epithelia. Implications for paracrine and target-derived Neurotrophic functions. AB - Brain-derived neurotrophic factor (BDNF) plays a crucial role for the survival of visceral sensory neurons during development. However, the physiological sources and the function of BDNF in the adult viscera are poorly described. We have investigated the cellular sources and the potential role of BDNF in adult murine viscera. We found markedly different amounts of BDNF protein in different organs. Surprisingly, BDNF levels in the urinary bladder, lung, and colon were higher than those found in the brain or skin. In situ hybridization experiments revealed that BDNF mRNA was made by visceral epithelial cells, several types of smooth muscle, and neurons of the myenteric plexus. Epithelia that expressed BDNF lacked both the high- and low-affinity receptors for BDNF, trkB and p75(NTR). In contrast, both receptors were present on neurons of the peripheral nervous system. Studies with BDNF-/-mice demonstrated that epithelial and smooth muscle cells developed normally in the absence of BDNF. These data provide evidence that visceral epithelia are a major source, but not a target, of BDNF in the adult viscera. The abundance of BDNF protein in certain internal organs suggests that this neurotrophin may regulate the function of adult visceral sensory and motor neurons. PMID- 10514402 TI - Tryptase-chymase double-positive human mast cells express the eotaxin receptor CCR3 and are attracted by CCR3-binding chemokines. AB - Eosinophils, basophils, and Th2 cells express the chemokine receptor CCR3, which binds eotaxin, RANTES, and some other chemokines. Using immunohistochemistry and flow cytometry, we demonstrate that CCR3 is also expressed by a variable proportion of human mast cells in gut, skin, and lung tissue. By contrast, with the same anti-CCR3 antibody (B711), CCR3 was poorly if at all detectable on human Th2 cells in vitro and in vivo. Eotaxin neither induced histamine release from purified human mast cells nor increased anti-IgE-stimulated histamine secretion. However, both eotaxin and RANTES elicited mast cell migration in vitro with a similar efficacy. High percentages of CCR3-expressing mast cells were present in the skin and in the intestinal submucosa; much lower percentages were found in the intestinal mucosa and in lung interstitium. Double immunostaining with anti CCR3 and anti-chymase antibody showed that the vast majority of CCR3-expressing mast cells in the various tissues examined were tryptase-chymase double-positive. Therefore, tryptase-chymase double-positive mast cells express CCR3 and are attracted by CCR3-binding chemokines, eotaxin, and RANTES. Our findings indicate that these chemokines may play an important role in the differentiation and/or migration of this mast cell subset in connective tissues, as well as in sites of allergic inflammation. PMID- 10514403 TI - The role of B lymphocytes in coxsackievirus B3 infection. AB - Coxsackieviruses are important human pathogens, frequently causing myocarditis, pancreatitis, and a variety of less severe diseases. B lymphocytes appear central to the interaction between these viruses and their mammalian hosts, because agammaglobulinemic humans, genetically incapable of antibody production, are susceptible to chronic infections by coxsackieviruses and related enteroviruses, such as poliovirus and echovirus. However, recent studies show that Type B coxsackievirus (CVB) infects B lymphocytes soon after infection, suggesting the possibility that these cells may play some role in virus dissemination and/or that the virus may be able to modulate the host immune response. We analyzed the role of B lymphocytes in CVB infection and confirmed that CVB infects B lymphocytes, and extended these findings to show that this is a productive infection involving approximately 1 to 10% of the cells; however, infectious center assays show that other splenocytes are infected at approximately the same frequency. Virus is readily detectable by in situ hybridization in the spleen of immunocompetent mice but is difficult to detect in mice deficient in B cells (BcKO mice), consistent with much of the splenic signal being the result of B cell infection. Surprisingly, given the extent of their infection, B cells express barely detectable levels of the murine coxsackievirus-adenovirus receptor (mCAR), suggesting that another means of cell entry may be used. We found no evidence of B cell depletion following CVB infection, indicating that this is not the explanation for the transient immunosuppression previously reported. Virus replication and dissemination are slightly delayed in BcKO mice, consistent with B cells' playing a role as an important early target of infection and/or a means to distribute the virus to many tissues. In addition, we show that BcKO mice recapitulate a central feature of human agammaglobulinemia: CVB establishes chronic infection in a variety of organs (heart, liver, brain, kidney, lung, pancreas, spleen). In most of these tissues the viral titers remain high (10(5) 10(8) plaque forming units (pfu) per gram of tissue) for the life of the mouse, and in several there is severe pathology, particularly severe myocardial fibrosis with ventricular dilation, reminiscent of the dilated cardiomyopathy seen in humans with chronic enteroviral myocarditis. Transfer of B and/or T cells from non-immune mice had no discernible effect, whereas equivalent transfers from immune mice often resulted in transient or permanent disappearance of detectable CVB. PMID- 10514404 TI - Neuropathogenesis of simian immunodeficiency virus in neonatal rhesus macaques. AB - Neonatal human immunodeficiency virus (HIV) infection usually occurs intrapartum or postpartum and results in a higher incidence of neurological dysfunction than is seen in adults. To explore the neuropathogenesis of neonatal HIV infection, we infected neonatal macaques with simian immunodeficiency virus (SIV) and followed the course of infection focusing on early time points. Infected neonates had decreased brain growth and mild histological changes in brain that resembled those seen in pediatric AIDS, including perivascular infiltrates of mononuclear cells, mineralization of vessels in the basal ganglia, and gliosis. The perivascular lesions and gliosis were associated with the presence of occasional infected cells that required in situ hybridization with radiolabeled riboprobes for detection. Using this technique, SIV-infected cells were detected in the brain parenchyma within 7 days of infection. These findings were confirmed by nested PCR for SIVgag DNA in brain and RT-PCR for viral RNA in cerebrospinal fluid. Together, these techniques revealed SIV infection of the CNS in 12 of 13 neonates infected with SIVmac239, 3 of 3 infected with SIVmac251, and 2 of 2 infected with SIVmac239/316. The prevalence of CNS infection was indistinguishable from that of older animals infected with the same dose and stock of virus, but neonates appeared to have fewer infected cells in the CNS and detecting them required more sensitive techniques. This observation was true regardless of inoculum and despite the fact that neonates had equal or greater viral loads in the periphery compared with older animals. These data suggest that maturation-dependent host factors have a major impact on the neuropathogenesis of pediatric AIDS. PMID- 10514406 TI - Widespread alterations of alpha-synuclein in multiple system atrophy. AB - Glial cytoplasmic inclusions (GCI) are the hallmark of multiple system atrophy (MSA), a rare movement disorder frequently associated with autonomic dysfunction. In this study of 21 cases of MSA, GCI were consistently immunoreactive for alpha synuclein and double-immunostained for ubiquitin and oligodendroglial markers, but not glial fibrillary acidic protein. No statistically significant difference was found in the density of GCI in various brain regions in the two forms of MSA, striatonigral degeneration (SND) and olivopontocerebellar atrophy (OPCA). Postmortem brain samples from 9 cases of MSA were fractionated according to solubility in buffer, Triton-X 100, sodium dodecyl sulfate (SDS), and formic acid, and alpha-synuclein immunoreactivity was measured in Western blots. Total alpha-synuclein immunoreactivity was increased in MSA compared to controls, with no statistically significant difference between SND and OPCA. Most of the increase was due to alpha-synuclein in SDS fractions. In controls this fraction had little or no immunoreactivity. In 7 cases and 4 controls correlations were investigated between quantitative neuropathology and biochemical properties of alpha-synuclein. Surprisingly, the amount of SDS-soluble alpha-synuclein correlated poorly with the number of GCI in adjacent sections. Furthermore, areas with few or no GCI unexpectedly had abundant SDS-soluble alpha-synuclein. These findings provide evidence that modifications of alpha-synuclein in MSA may be more widespread than obvious histopathology. Moreover, these alterations may constitute a biochemical signature for the synucleinopathies. PMID- 10514405 TI - Establishment of a human thymic myoid cell line. Phenotypic and functional characteristics. AB - The subset of myoid cells is a normal component of the thymic stroma. To characterize these cells, we immortalized stromal cells from human thymus by using a plasmid vector encoding the SV40 T oncogene. Among the eight cell lines obtained, one had myoid characteristics including desmin and troponin antigens. This new line was designated MITC (myoid immortalized thymic cells). These cells expressed both the fetal and adult forms of muscle acetylcholine receptor (AChR) at the mRNA level, as well as the myogenic transcription factor MyoD1. alpha Subunit AChR protein expression was detected by flow cytometry and the AChR was functional in patch-clamp studies. In addition, AChR expression was down modulated by myasthenia gravis sera or by monoclonal antibody anti-AChR on MITC line similarly to TE671 rhabdomyosarcoma cells, making the MITC line an interesting tool for AChR antigenic modulation experiments. Finally, the MITC line expressed LFA-3, produced several cytokines able to act on T cells, and protected total thymocytes from spontaneous apoptosis in vitro. These results are compatible with a role of thymic myoid cells in some steps of thymocyte development. Therefore MITC line appears to be a useful tool to investigate the physiological role of thymic myoid cells. PMID- 10514407 TI - Immunohistochemical evidence of loss of PTEN expression in primary ductal adenocarcinomas of the breast. AB - Germline mutations in PTEN, encoding a dual-specificity phosphatase on 10q23.3, cause Cowden syndrome (CS), which is characterized by a high risk of breast and thyroid cancers. Loss of heterozygosity of 10q22-24 markers and somatic PTEN mutations have been found to a greater or lesser extent in a variety of sporadic component and noncomponent cancers of CS. Among several series of sporadic breast carcinomas, the frequency of loss of flanking markers around PTEN is approximately 30 to 40%, and the somatic intragenic PTEN mutation frequency is <5%. In this study, we analyzed PTEN expression in 33 sporadic primary breast carcinoma samples using immunohistochemistry and correlated this to structural studies at the molecular level. Normal mammary tissue had a distinctive pattern of expression: myoepithelial cells uniformly showed strong PTEN expression. The PTEN protein level in mammary epithelial cells was variable. Ductal hyperplasia with and without atypia exhibited higher PTEN protein levels than normal mammary epithelial cells. Among the 33 carcinoma samples, 5 (15%) were immunohistochemically PTEN-negative; 6 (18%) had reduced staining, and the rest were PTEN-positive. In the PTEN-positive tumors as well as in normal epithelium, the protein was localized in the cytoplasm and in the nucleus (or nuclear membrane). Among the immunostain negative group, all had hemizygous PTEN deletion but no structural alteration of the remaining allele. Thus, in these cases, an epigenetic phenomenon such as hypermethylation, -ecreased protein synthesis or increased protein degradation may be involved. In the cases with reduced staining, 5 of 6 had hemizygous PTEN deletion and 1 did not have any structural abnormality. Finally, clinicopathological features were analyzed against PTEN protein expression. Three of the 5 PTEN immunostain-negative carcinomas were also both estrogen and progesterone receptor-negative, whereas only 5 of 22 of the PTEN-positive group were double receptor-negative. The significance of this last observation requires further study. PMID- 10514408 TI - The PEN5 epitope identifies an oligodendrocyte precursor cell population and pilocytic astrocytomas. AB - PEN5 is a sulfated polylactosamine carbohydrate epitope first described in a subpopulation of mature natural killer cells. Here we report that it is also expressed in a developmentally regulated fashion in human and rat central nervous systems and that its protein carrier is P-selectin glycoprotein ligand-1 (PSGL 1), a ligand for selectins. In rat neural primary cultures, PEN5 is transiently and selectively expressed by oligodendrocyte precursor cells and marks the transition from proliferative to postmitotic stages. In concordance, in human central nervous system tumors, PEN5 is observed in a subset of oligodendrogliomas and in all pilocytic astrocytomas, a class of tumor of uncertain histogenesis. These data suggest that PEN5-PSGL-1 plays a role in the differentiation of oligodendrocytes and that pilocytic astrocytomas are likely to result from a dysregulation occurring in oligodendrocyte precursor cells at the crucial stage of exit from the cell cycle. PMID- 10514409 TI - Tyrosine kinases expressed in vivo by human prostate cancer bone marrow metastases and loss of the type 1 insulin-like growth factor receptor. AB - An important biological feature of prostate cancer (PCa) is its marked preference for bone marrow as a metastatic site. To identify factors that may support the growth of PCa in bone marrow, expression of receptor and nonreceptor tyrosine kinases by androgen-independent PCa bone marrow metastases was assessed. Bone marrow biopsies largely replaced by PCa were analyzed using reverse transcriptase polymerase chain reaction amplification with degenerate primers that amplified the conserved kinase domain. Sequence analyses of the cloned products demonstrated expression of multiple kinases. Expression of the receptor and nonreceptor tyrosine kinases, alpha platelet-derived growth factor receptor and Jak 1, respectively, was confirmed by immunohistochemistry. In contrast, the type 1 insulin-like growth factor receptor, thought to play a role in PCa development, was lost in metastatic PCa. These results implicate several specific growth factors and signaling pathways in metastatic androgen-independent PCa and indicate that loss of the type 1 insulin-like growth factor receptor contributes to PCa progression. PMID- 10514410 TI - Augmented expression of cyclooxygenase-2 in human atherosclerotic lesions. AB - Cyclooxygenase-1 (Cox-1) and Cox-2 convert arachidonic acid to prostaglandin H(2), the precursor of other prostaglandins and thromboxanes, eicosanoids important in vascular pathophysiology. However, knowledge of the expression of cyclooxygenases within atherosclerotic lesions is scant. This study tested the hypothesis that human atheroma and nonatherosclerotic arteries express the two Cox isoforms differentially. Cox-1 mRNA and protein localized on endothelial and medial smooth muscle cells of normal arteries (n = 5), whereas Cox-2 expression was not detectable. In contrast, atheromatous (n = 7) lesions contained both Cox 1 and Cox-2, colocalizing mainly with macrophages of the shoulder region and lipid core periphery, whereas smooth muscle cells showed lower levels, as demonstrated by immunohistochemical and in situ hybridization analysis. Furthermore, microvascular endothelium in plaques showed notable staining for both isoforms. In accord with immunohistochemical studies, Western blot analysis of protein extracts from normal arteries revealed constitutive Cox-1, but not Cox 2, expression. Extracts of atheromatous lesions, however, contained both Cox-1 and Cox-2 protein, detected as two immunoreactive proteins of approximately 70 and 50 kd. Macrophages expressed the short form of Cox-1/-2 constitutively after several days of in vitro culture, rather than the 70-kd protein. These results shed new light on the inflammatory pathways that operate in human atheroma. In particular, the expression of Cox-2 in atheromatous, but not in unaffected, arteries has therapeutic implications, given the advent of selective Cox-2 inhibitors. PMID- 10514411 TI - C6 produced by macrophages contributes to cardiac allograft rejection. AB - The terminal components of complement C5b-C9 can cause significant injury to cardiac allografts. Using C6-deficient rats, we have found that the rejection of major histocompatibility (MHC) class I-incompatible PVG.R8 (RT1.A(a)B(u)) cardiac allografts by PVG.1U (RT1.A(u)B(u)) recipients is particularly dependent on C6. This model was selected to determine whether tissue injury results from C6 produced by macrophages, which are a conspicuous component of infiltrates in rejecting transplants. We demonstrated that high levels of C6 mRNA are expressed in isolated populations of macrophages. The relevance of macrophage-produced C6 to cardiac allograft injury was investigated by transplanting hearts from PVG. R8 (C6-) donors to PVG.1U (C6-) rats which had been reconstituted with bone marrow from PVG.1U (C6+) rats as the sole source of C6. Hearts grafted to hosts after C6 reconstitution by bone marrow transplantation underwent rejection characterized by deposition of IgG and complement on the vascular endothelium together with extensive intravascular aggregates of P-selectin-positive platelets. At the time of acute rejection, the cardiac allografts contained extensive perivascular and interstitial macrophage infiltrates. RT-PCR and in situ hybridization demonstrated high levels of C6 mRNA in the macrophage-laden transplants. C6 protein levels were also increased in the circulation during rejection. To determine the relative contribution to cardiac allograft rejection of the low levels of circulating C6 produced systemically by macrophages, C6 containing serum was passively transferred to PVG.1U (C6-) recipients of PVG.R8 (C6-) hearts. This reconstituted the C6 levels to about 3 to 6% of normal values, but failed to induce allograft rejection. In control PVG.1U (C6-) recipients that were reconstituted with bone marrow from PVG.1U (C6-) donors, C6 levels remained undetectable and PVG.R8 cardiac allografts were not rejected. These results indicate that C6 produced by macrophages can cause significant tissue damage. PMID- 10514412 TI - Endothelial L-selectin ligands are likely to recruit lymphocytes into rejecting human heart transplants. AB - L-selectin-dependent lymphocyte extravasation is a hallmark of acute heart allograft rejection in rats. On screening over 600 endomyocardial biopsies (EMBs), taken at different time points after heart transplantation in man, we identified 91 samples with histological signs of acute rejection. Rejection and nonrejection EMBs were analyzed for the presence of properly glycosylated, ie, sulfated sialyl Lewis-x (sLex) decorated L-selectin ligands. Two anti-sLex (2F3 and HECA-452) and one anti-6- or 6'-sulfated and/or 6, 6'-bisulfation (MECA-79) monoclonal antibodies were used. Nonrejecting heart endothelium did not express, or expressed only weakly, sulfated and or sLex decorations of L-selectin ligands. On the contrary, these epitopes were readily detectable on endothelium of capillaries and venules at the onset and during acute rejection episodes. The more intense the sulfated sLex expression was, the more severe the rejection episode was in histological grading. The endothelial expression of L-selectin ligands decreased to background levels as the rejection resolved. Our data demonstrate a complete correlation between the level of expression of the sulfated sLex-decorated ligands on the one hand and the histological severity of acute heart allograft rejection on the other hand. These data suggest that functionally active endothelial L-selectin ligands are instrumental in lymphocyte extravasation into the human heart allografts at the onset and during acute rejection episodes. PMID- 10514413 TI - Vascular smooth muscle cells express the transcriptional corepressor NAB2 in response to injury. AB - The early growth response 1 (Egr-1 or NGFI-A) gene product is a zinc finger protein transcription factor which has been implicated in the regulation of genes differentially expressed during the development of vascular disease. Egr-1 activity is regulated by alterations in the amount of protein, as well as protein protein interactions with positive and negative transcriptional cofactors. NGFI-A binding protein 2 (NAB2) is an example of a negative transcriptional cofactor capable of binding directly to Egr-1 and repressing Egr-1-mediated transcription. In this study, we show that NAB2 is rapidly and transiently expressed in vascular smooth muscle cells (VSMC) in response to the model agonist phorbol 12-myristate 13-acetate (PMA). This induction occurs at the protein as well as mRNA level, and the time course of induction trails closely behind that of Egr-1. NAB2 expression in VSMC is capable of inhibiting Egr-1 dependent gene expression in response to either PMA or fibroblastic growth factor-2 (FGF-2). In an in vivo model of mechanical arterial injury NAB2 levels also increase transiently in VSMC at a time when Egr-1 is elevated. It is possible that NAB2 is part of a negative feedback mechanism which serves to down-regulate Egr-1-mediated gene transcription in injured VSMC. PMID- 10514414 TI - Mouse senile amyloid deposition is suppressed by adenovirus-mediated overexpression of amyloid-resistant apolipoprotein A-II. AB - Apolipoprotein A-II (apoA-II), the second most abundant apolipoprotein of serum high density lipoprotein, deposits as an amyloid fibril (AApoAII) in old mice. Mouse strains with a high incidence of senile amyloidosis have the type C apoA-II gene (Apoa2(c)), whereas the strains with a low incidence of amyloidosis have the type B apoA-II gene (Apoa2(b)). In this study, to investigate whether the type B apoA-II protein inhibits the extension of amyloid fibrils, we constructed an adenovirus vector bearing the Apoa2(b) cDNA (Adex1CATApoa2(b)), which is expressed under the control of a hepatocyte-specific promoter. The mice were infected with Adex1CATApoa2(b) before induction of amyloidosis by the injection of AApoAII amyloid fibril seeds. Compared with the mice infected with the control virus, amyloid deposition was suppressed significantly in the mice infected with Adex1CATApoa2(b). Fluorometry using thioflavine T also revealed that AApoAII fibril extension was inhibited by the addition of type B apoA-II in vitro. Thus, we propose that Apoa2(b) contributes as an active inhibitor of amyloid fibril extension and overexpression of amyloid-resistant gene variant may be an attractive therapeutic target in amyloidosis. PMID- 10514415 TI - Comparative analysis of different methodological approaches to the in vitro study of drug-induced apoptosis. AB - Apoptosis is a dynamic process in which a characteristic morphological or biochemical event used in an assay as a specific marker of apoptosis may be observed over a limited period of time. Asynchronous involvement of cells in apoptosis results in different proportions of apoptotic cells with blebbed membrane, broken nuclei, modified mitochondrial units or fragmented DNA coexisting in the culture at any single moment. Thus, depending on the method used, the extent of apoptosis determined in the same cell population may vary. In the present study, a microculture kinetic (MiCK) assay was used to monitor apoptosis in HL-60 cells exposed to 1, 2.5, 5, 10, and 20 micromol/L etoposide and cisplatin. Both the extent and timing of apoptotic responses were dependent on the drug and drug concentration. Time-lapse video microscopy (TLVM), flow cytometry analysis of the light scattering properties of cells, morphological studies of Giemsa-stained cells, annexin V binding, and DNA fragmentation assays were performed at multiple times of cell exposure to 10 micromol/L etoposide and 5 micromol/L cisplatin. Steep linear increases in optical density, indicating apoptosis in the MiCK assay, correlated with both linear increases in the proportion of cells with plasma membrane blebbing in TLVM and with increased side scattering properties of apoptotic cells in flow cytometry. During a 24-hour culture period, the MiCK assay and TLVM provided multiple consecutive appraisals of nondisturbed cell microcultures at intervals of 5 and 2.5 minutes, respectively, and thus could be considered as real time kinetic assays. With the three endpoint assays, each of which was applied 12 times at 2-hour intervals, maximum apoptotic responses varied from 22.5 to 72% in etoposide-treated cells and from 30 to 57% in cisplatin-treated cells. With the annexin V binding assay, maximum apoptosis could always be detected 4 to 5 hours earlier than it was seen in Giemsa-stained preparations and 8 hours earlier than it was detected by measuring of DNA fragmentation. Values of the maximum extent of apoptosis varied, being the lowest with annexin V and the greatest with DNA fragmentation assays. The best correlations of both extent and timing of apoptosis were observed between the MiCK, TLVM, and morphological assays. In conclusion, both a maximum apoptotic response and the time at which it was achieved are the obligatory requirements for determining the apoptosis-inducing potency of an agent and for comparing results of studies performed in different laboratories. PMID- 10514416 TI - The role of p53 in bleomycin-induced DNA damage in the lung. A comparative study with the small intestine. AB - To elucidate the role of p53 and apoptosis in the pathogenesis of lung injury, we examined histological changes, expressions of p53 and p21waf1/cip1 (p21), apoptosis, DNA double strand breaks, cell kinetics, and DNA synthesis in C57/BL6 mice (p53+/+) and mice deficient for p53 (p53-/-) at 2 hours to 7 days after a single intravenous administration of bleomycin. We also compared these parameters between the lung cells and small intestinal epithelial cells to explore potential differences in their response to DNA damage. Bleomycin induced p21 expression in a p53-dependent manner in p53+/+ mice but neither p53 nor p21 expression in p53-/ mice. In the lung of both groups of mice, focal inflammation followed by fibrosis was observed, but there was no evidence of apoptosis. Cells with DNA breaks and those undergoing DNA synthesis were unequivocally increased, but the cycling cell fraction remained unchanged, suggesting that the DNA synthesis detected in the lung reflected unscheduled DNA synthesis for repair of damaged DNA. DNA breaks and unscheduled DNA synthesis were prolonged in p53-/- mice compared to p53+/+ mice. By contrast, in the small intestine, marked cell cycle arrest and extensive apoptosis were evoked in the cycling crypt cells of both groups of mice, but these changes were milder and DNA breaks remained detectable for a longer time in p53-/- mice than in p53+/+ mice. Among the resting enterocytes in the villi, apoptosis was observed almost equally in both groups, but repair of DNA breaks was significantly delayed in the p53-/- mice. These observations imply that apoptosis is mediated largely by the p53-dependent pathway in the crypts but exclusively by the p53-independent pathway in the villi, that this pathway is particularly important in DNA repair in the villi, and that despite this difference in the significance of apoptosis, p53 plays an important role in DNA repair in both the crypts and villi. Our results suggest that the lung cells and small intestinal cells respond to the bleomycin treatment in different ways in terms of the induction of apoptosis and that p53 carries out an essential role in the early response to and repair of DNA damage by a non apoptotic mechanism which appears to be crucial in the noncycling lung cells and enterocytes. Importantly, the p53-p21 pathway and apoptosis are unlikely to be essential for bleomycin-induced tissue injury in the lung. PMID- 10514417 TI - A murine xenograft model for human CD30+ anaplastic large cell lymphoma. Successful growth inhibition with an anti-CD30 antibody (HeFi-1). AB - To develop a model for the biology and treatment of CD30+ anaplastic large cell lymphoma (ALCL), we transplanted leukemic tumor cells from a 22-month-old girl with multiple relapsed ALCL. Tumor cells were inoculated intraperitoneally into a 4-week-old SCID/bg mouse and produced a disseminated tumor within 8 weeks; this tumor was serially transplanted by subcutaneous injections to other mice. Morphology, immunohistochemistry, and molecular genetics which demonstrated the NPM-ALK fusion protein, resulting from the t(2;5)(p23;q35), confirmed the identity of the xenograft with the original tumor. The tumor produced transcripts for interleukin-1alpha, tumor necrosis factor-alpha, and interferon-gamma which could explain the patient's B-symptoms. Treatment of mice with monoclonal antibody (HeFi-1) which activates CD30 antigen administered on day 1 after tumor transplantation prevented tumor growth. Treatment with HeFi-1 after tumors had reached a 0.2 cm(3) volume caused tumor growth arrest and prevention of tumor dissemination. We conclude that transplantation of CD30+ ALCL to SCID/bg mice may provide a valuable model for the study of the biology and design of treatment modalities for CD30+ ALCL. PMID- 10514418 TI - Megalin knockout mice as an animal model of low molecular weight proteinuria. AB - Megalin is an endocytic receptor expressed on the luminal surface of the renal proximal tubules. The receptor is believed to play an important role in the tubular uptake of macromolecules filtered through the glomerulus. To elucidate the role of megalin in vivo and to identify its endogenous ligands, we analyzed the proximal tubular function in mice genetically deficient for the receptor. We demonstrate that megalin-deficient mice exhibit a tubular resorption deficiency and excrete low molecular weight plasma proteins in the urine (low molecular weight proteinuria). Proteins excreted include small plasma proteins that carry lipophilic compounds including vitamin D-binding protein, retinol-binding protein, alpha(1)-microglobulin and odorant-binding protein. Megalin binds these proteins and mediates their cellular uptake. Urinary loss of carrier proteins in megalin-deficient mice results in concomitant loss of lipophilic vitamins bound to the carriers. Similar to megalin knockout mice, patients with low molecular weight proteinuria as in Fanconi syndrome are also shown to excrete vitamin/carrier complexes. Thus, these results identify a crucial role of the proximal tubule in retrieval of filtered vitamin/carrier complexes and the central role played by megalin in this process. PMID- 10514419 TI - A rat model of progressive chronic renal failure produced by microembolism. AB - We report a new model of chronic progressive renal failure in rats, produced by a single injection of microspheres (20 to 30 micrometer in diameter) into the left renal artery after right nephrectomy. Significant proteinuria appeared after 4 weeks, followed by hypoalbuminemia and hypercholesterolemia, in rats that received approximately 5 x 10(5) microspheres (0.8 mg). Renal function partially recovered by 4 weeks after nephrectomy and injection from postoperative dysfunction, but deteriorated again 12 weeks after operation. In the early stage, histologic examination showed tubules with cuff-like thickening of basement membranes scattered among apparently intact tubules. Many epithelial cells in the atrophic tubuli were immunoreactive for proliferating cell nuclear antigen (PCNA). Dilated tubules became apparent several weeks after development of tubular atrophy, most likely representing distal tubules. Dilated tubuli were mostly negative for the proliferation marker. These results showed similarity to findings in human chronic renal failure and strongly suggested that tubular atrophy and dilation in chronic tubulointerstitial lesions differ in pathogenesis. This new model of renal failure induced by microembolism should be useful for studying the interaction between normal and diseased tissue elements in histologically heterogenous lesions as well as the pathogenesis of interstitial fibrosis in disturbance of microcirculation. PMID- 10514420 TI - Nitric oxide synthase inhibition by N(G)-nitro-L-arginine methyl ester inhibits tumor-induced angiogenesis in mammary tumors. AB - Using a murine breast cancer model, we earlier found a positive correlation between the expression of nitric oxide synthase (NOS) and tumor progression; treatment with inhibitors of NOS, N(G)-methyl-L-arginine (NMMA) and N(G)-nitro-L arginine methyl ester (L-NAME), had antitumor and antimetastatic effects that were partly attributed to reduced tumor cell invasiveness. In the present study, we used a novel in vivo model of tumor angiogenesis using subcutaneous implants of tumor cells suspended in growth factor-reduced Matrigel to examine the angiogenic role of NO in a highly metastatic murine mammary adenocarcinoma cell line. This cell line, C3L5, expresses endothelial (e) NOS in vitro and in vivo, and inducible (i) NOS in vitro on stimulation with lipopolysaccharide and interferon-gamma. Female C3H/HeJ mice received subcutaneous implants of growth factor-reduced Matrigel inclusive of C3L5 cells on one side, and on the contralateral side, Matrigel alone; L-NAME and D-NAME (inactive enantiomer) were subsequently administered for 14 days using osmotic minipumps. Immediately after sacrifice, implants were removed and processed for immunolocalization of eNOS and iNOS proteins, and measurement of angiogenesis. Neovascularization was quantified in sections stained with Masson's trichrome or immunostained for the endothelial cell specific CD31 antigen. While most tumor cells and endothelial cells expressed immunoreactive eNOS protein, iNOS was localized in endothelial cells and some macrophages within the tumor-inclusive implants. Measurable angiogenesis occurred only in implants containing tumor cells. Irrespective of the method of quantification used, tumor-induced neovascularization was significantly reduced in L-NAME-treated mice relative to those treated with D-NAME. The quantity of stromal tissue was lower, but the quantity of necrotic tissue higher in L-NAME relative to D-NAME-treated animals. The total mass of viable tissue (ie, stroma and tumor cells) was lower in L-NAME relative to D-NAME-treated animals. These data suggest that NO is a key mediator of C3L5 tumor-induced angiogenesis, and that the antitumor effects of L-NAME are partly mediated by reduced tumor angiogenesis. PMID- 10514422 TI - Context-dependent transcriptional regulation. PMID- 10514421 TI - Expression of E-selectin, P-selectin, and intercellular adhesion molecule-1 during experimental murine listeriosis. AB - The expression of adhesion molecules E-selectin, P-selectin, and intercellular adhesion molecule-1 (ICAM-1) was immunohistochemically investigated during the course of experimental murine listeriosis. Infection was monitored by microbiological count of blood, liver, and spleen. After an early generalized expression of P-selectin and ICAM-1, a later regulation occurred specifically to areas of inflammation. Expression of E-selectin was faint and inconstantly detected in all of the studied organs. In the liver, typical lesions of murine listeriosis were related to the expression of ICAM-1 on sinusoidal endothelial cells and the biliary system and to the de novo expression of P-selectin in hepatic portal vessels. Inflammation in the spleen was related to the expression of ICAM-1 on red pulp sinusoidal cells, especially in the marginal sinus. High endothelial venules of inflamed lymph nodes also expressed P-selectin and ICAM-1. Lesions in the central nervous system appeared on day 3 after infection as a pyogranulomatous leptomeningitis associated with an intense expression of P selectin and ICAM-1 in meningeal vessels, especially those in the hippocampal sulcus, suggesting a way through which inflammation initially reach the central nervous system during experimental murine listeriosis. Leptomeningitis was followed by the presence of ventriculitis, which was related to the up-regulation of ICAM-1 on choroid plexus epithelial cells, periventricular vessels and ependymal cells. Up-regulation of P-selectin and ICAM-1 during experimental murine listeriosis could play an important role in the recruitment of leukocytes, especially to the liver, lymphoid organs, and central nervous system. PMID- 10514423 TI - Processing of the human heparanase precursor and evidence that the active enzyme is a heterodimer. AB - Human platelet heparanase has been purified to homogeneity and shown to consist of two, non-covalently associated polypeptide chains of molecular masses 50 and 8 kDa. Protein sequencing provided the basis for determination of the full-length cDNA for this novel protein. Based upon this information and results from protein analysis and mass spectrometry, we propose a scheme to define the structural organization of heparanase in relation to its precursor forms, proheparanase and pre-proheparanase. The 8- and 50-kDa chains which make up the active enzyme reside, respectively, at the NH(2)- and COOH-terminal regions of the inactive precursor, proheparanase. The heparanase heterodimer is produced by excision and loss of an internal linking segment. This paper is the first to suggest that human heparanase is a two-chain enzyme. PMID- 10514425 TI - Stress-induced JNK activation is independent of Gadd45 induction. AB - DNA damage and environmental stress activate signaling and induce genes involved in cell cycle and cell death. Expression of the Gadd45 protein is induced following DNA damage and other stress. Gadd45 is believed to play a role in growth arrest and possibly in cell death. The JNK signaling pathway is also activated by some DNA-damaging agents. This activation leads to phosphorylation and activation of transcription factors, such as c-Jun/AP-1 and ATF2, which mediate immediate early gene induction. Recently Gadd45 was suggested to be involved in JNK activation. However, as this suggestion relied on in vitro experiments and ectopic overexpression of Gadd45 protein, we examined whether physiological levels of Gadd45 that are induced following exposure to DNA damaging agents and stress can lead to JNK induction. We found that JNK activation by UV irradiation and anisomycin treatment precedes the induction of gadd45 mRNA by these agents. Gadd45 protein induction by methyl methanesulfonate also lagged behind JNK activation. The use of protein synthesis inhibitors suggested that newly synthesized proteins, including the stress-induced Gadd45, make only a marginal contribution to JNK activation. We also found that stresses such as gamma irradiation induce Gadd45 and do not activate JNK in mouse fibroblasts. Therefore, stress-induced JNK does not depend on Gadd45 induction. PMID- 10514424 TI - beta-TrCP mediates the signal-induced ubiquitination of IkappaBbeta. AB - We have examined the role of beta-TrCP (beta-transducin repeat-containing protein) in the ubiquitination and degradation of IkappaBbeta, one of the two major IkappaB isoforms in mammalian cells. We demonstrate that beta-TrCP interacts specifically with IkappaBbeta, and such interaction is dependent on prior phosphorylation of IkappaBbeta on serines 19 and 23. Interaction with beta TrCP is also necessary for ubiquitination of IkappaBbeta upon stimulation of cells, and deletion of the F-box in beta-TrCP abolishes its ability to ubiquitinate IkappaBbeta. Therefore, these results indicate that beta-TrCP plays a critical role in the activation of NF-kappaB by assembling the ubiquitin ligase complex for both phosphorylated IkappaBalpha and IkappaBbeta. PMID- 10514426 TI - gadd45 is not required for activation of c-Jun N-terminal kinase or p38 during acute stress. AB - Cells respond to environmental stress with activation of c-Jun N-terminal kinase (JNK) and p38. Recent studies have implicated Gadd45 and two related proteins, MyD118/Gadd45beta and CR6/Gadd45gamma, as initiators of JNK/p38 signaling via their interaction with an upstream kinase MTK1. It was proposed that stress induced expression of the Gadd45-related proteins leads to MTK1 activation and subsequent JNK/p38 activation. Using embryo fibroblasts from gadd45-null mice, we have addressed the requirement for Gadd45 in mediating JNK/p38 activation during acute stress. Comparison of JNK/p38 activities in response to methyl methanesulfonate, hydrogen peroxide, UVC irradiation, sorbitol, and anisomycin treatment of gadd45(+/+) and gadd45(-/-) fibroblasts revealed no deficiency in JNK/p38 activation in gadd45(-/-) fibroblasts. In addition, in wild type cells, JNK and p38 activation significantly preceded gadd45 induction with all stresses. Examination of myd118/gadd45beta and cr6/gadd45gamma expression in gadd45(+/+) and gadd45(-/-) fibroblasts revealed similar induction patterns in the two cell types, which, like gadd45 expression, was delayed relative to JNK/p38 activation. We conclude that gadd45 expression is not required for activation of JNK/p38 by environmental stresses, nor are stress-induced increases in myd118/gadd45beta and cr6/gadd45gamma expression necessary for kinase activation in response to such insults. PMID- 10514427 TI - Spontaneous human B2 bradykinin receptor activity determines the action of partial agonists as agonists or inverse agonists. Effect of basal desensitization. AB - In this report, we show that desensitization regulates ligand-independent, spontaneous activity of the human B2 bradykinin (BK) receptor, and the level of spontaneous receptor activity determines the action of the BK antagonists and partial receptor agonists NPC17731 and HOE140 as agonists or inverse agonists. Spontaneous receptor activity was monitored by measuring basal cellular phosphoinositide (PI) hydrolysis as a function of the density of the receptor in transiently transfected HEK293 cells. Minimal spontaneous activity of the wild type B2 receptor was detected in these cells. Mutating a cluster of serines and threonines within the fourth intracellular domain of the receptor, which is critical for agonist-promoted desensitization, significantly increased the spontaneous receptor activity. BK, the natural B2 receptor ligand and, consequently, a full agonist, stimulated PI hydrolysis at high and low levels of spontaneous receptor activity. On the other hand, the partial agonists NPC17731 and HOE140 were stimulatory, or agonists, at the lower level of receptor activity but inhibitory, or inverse agonists, at the higher level of activity. These results show that receptors are desensitized in response to their spontaneous activity. Furthermore, these results, which refute traditional theories, show that the capacity of a drug to modulate a receptor response is not intrinsic to the drug but is also dependent on the cellular environment in which the drug acts. PMID- 10514428 TI - Porin is present in the plasma membrane where it is concentrated in caveolae and caveolae-related domains. AB - Mitochondrial porin, or voltage-dependent anion channel, is a pore-forming protein first discovered in the outer mitochondrial membrane. Later investigations have provided indications for its presence also in other cellular membranes, including the plasma membrane, and in caveolae. This extra mitochondrial localization is debated and no clear-cut conclusion has been reached up to now. In this work, we used biochemical and electrophysiological techniques to detect and characterize porin within isolated caveolae and caveolae like domains (low density Triton-insoluble fractions). A new procedure was used to isolate porin from plasma membrane. The outer surface of cultured CEM cells was biotinylated by an impermeable reagent. Low density Triton-insoluble fractions were prepared from the labeled cells and used as starting material to purify a biotinylated protein with the same electrophoretic mobility and immunoreactivity of mitochondrial porin. In planar bilayers, the porin from these sources formed slightly anion-selective pores with properties indistinguishable from those of mitochondrial porin. This work thus provides a strong indication of the presence of porin in the plasma membrane, and specifically in caveolae and caveolae-like domains. PMID- 10514429 TI - Carotenoid-binding sites of the major light-harvesting complex II of higher plants. AB - Recombinant light-harvesting complex II (LHCII) proteins with modified carotenoid composition have been obtained by in vitro reconstitution of the Lhcb1 protein overexpressed in bacteria. The monomeric protein possesses three xanthophyll binding sites. The L1 and L2 sites, localized by electron crystallography in the helix A/helix B cross, have the highest affinity for lutein, but also bind violaxanthin and zeaxanthin with lower affinity. The latter xanthophyll causes disruption of excitation energy transfer. The occupancy of at least one of these sites, probably L1, is essential for protein folding. Neoxanthin is bound to a distinct site (N1) that is highly selective for this species and whose occupancy is not essential for protein folding. Whereas xanthophylls in the L1 and L2 sites interact mainly with chlorophyll a, neoxanthin shows strong interaction with chlorophyll b, inducing the hyperchromic effect of the 652 nm absorption band. This observation explains the recent results of energy transfer from carotenoids to chlorophyll b obtained by femtosecond absorption spectroscopy. Whereas xanthophylls in the L1 and L2 sites are active in photoprotection through chlorophyll-triplet quenching, neoxanthin seems to act mainly in (1)O(2)(*) scavenging. PMID- 10514430 TI - Surface plasmon resonance studies resolve the enigmatic endotoxin neutralizing activity of polymyxin B. AB - Polymyxin B (PMB), a cyclic cationic peptide antibiotic, despite its severe side effects continues to occupy a premiere position for treating endotoxicosis. Its mode of neutralization of endotoxin has remained elusive for the last three decades. Several synthetic peptide mimics of PMB, capable of binding endotoxin, have been made. However, the binding ability alone appears to be a deceptive indicator of endotoxin neutralizing activity as molecules with similar binding propensities could either sequester or opsonize the toxin. Hence identification of additional physical parameters which describe adequately the outcome of PMB endotoxin interaction become imperative. Surface plasmon resonance (SPR) studies reported here show that several mimics of PMB despite exhibiting lipopolysaccharide binding affinities comparable with it but, unlike it, do not sequester the endotoxin. These studies thus provide a striking illustration of the difference in the behavior of PMB, vis a vis its mimics toward the endotoxin lamellae, and define further, in chemical terms, mechanism of the action of PMB and allow us to posit that the design of molecules as effective antidotes for sepsis should incorporate the ability to sequester endotoxin specifically. PMID- 10514431 TI - Three-state unfolding and self-association of maspin, a tumor-suppressing serpin. AB - Maspin is a tumor suppressor protein expressed by normal human mammary epithelium but not by many breast tumor cell lines. Recombinant human maspin (rMaspin) inhibits tumor cell motility, invasion, and metastasis and thus has potential value as an anti-cancer therapeutic. Maspin is a member of the serpin family and, although the molecular mechanism by which maspin acts is unknown, recent work suggests that tissue plasminogen activator is a potential target. A puzzling observation in previous cell culture studies was loss of rMaspin activity at higher protein concentrations. One hypothesis to explain these results is self association of rMaspin at the higher concentrations, which would be consistent with the tendency of serpins to form noncovalent polymers. This hypothesis is addressed by examining the relationship between rMaspin stability and self association. Urea denaturation of rMaspin at pH 7 and 25 degrees C and at protein concentrations ranging from 0.01 to 0.2 mg/ml has been monitored by circular dichroism and intrinsic tryptophan fluorescence. Denaturation profiles show a protein concentration dependence and indicate the presence of at least one unfolding intermediate. The results suggest that destabilization of native monomeric rMaspin leads to partial unfolding and formation of an intermediate which can self-associate. PMID- 10514432 TI - Differential binding of histidine-rich glycoprotein (HRG) to human IgG subclasses and IgG molecules containing kappa and lambda light chains. AB - In previous studies we showed that the plasma protein histidine-rich glycoprotein (HRG) binds strongly to pooled human IgG. In the present work myeloma proteins consisting of different human IgG subclasses were examined for their ability to interact with human HRG. Using an IAsys optical biosensor we found initially that IgG subclasses differ substantially in their affinity of interaction with HRG. However, the most striking finding was the observation that the kinetics of the HRG interaction was dramatically affected by whether the IgG subclasses contained the kappa or lambda light (L)-chains. Thus, the on-rate for the binding of HRG to the kappa L-chain containing IgG1 and IgG2 (IgG1kappa and IgG2kappa) was approximately 4- and approximately 10-fold faster than that for the binding of HRG to lambda L-chain containing IgG1 and IgG2 (IgG1lambda and IgG2lambda), respectively, with the dissociation constants (K(d)) in the range 3-5 nM and 112 189 nM for the kappa and lambda isoforms, respectively. In contrast, the on-rate for the binding of HRG to IgG3kappa and IgG4kappa was found to be 9- and 20-fold slower than that for the binding of HRG to IgG3lambda and IgG4lambda, respectively, with the K(d) in the range 147-268 nM and 96-109 nM for the kappa and lambda isoforms, respectively. The binding of HRG to immunoglobulins containing the kappa L-chain (particularly IgG1kappa) was generally potentiated in the presence of a physiological concentration (20 microM) of Zn(2+) (K(d) decreased to 0.60 +/- 0.01 for IgG1kappa), but Zn(2+) had no effect or slightly inhibited the binding of HRG to immobilized IgG subclasses possessing the lambda L-chain. Interestingly, HRG also bound differentially to Bence Jones (BJ) proteins containing kappa and lambda L-chains, with HRG having a 14-fold lower K(d) for BJkappa than for BJlambda when 20 microM Zn(2+) was present. HRG also bound to IgM (IgMkappa), but the affinity of this interaction (K(d) approximately 1.99 +/- 0.05 microM) was markedly lower than the interaction with IgG, and the affinity was actually decreased 4-fold in the presence of Zn(2+). The results demonstrate that both the heavy (H)- and L-chain type have a profound effect on the binding of HRG to different IgG subclasses and provide the first evidence of a functional difference between the kappa and lambda L-chains of immunoglobulins. PMID- 10514433 TI - Molecular dissection of the interactions among IkappaBalpha, FWD1, and Skp1 required for ubiquitin-mediated proteolysis of IkappaBalpha. AB - The SCF complex containing Skp1, Cul1, and the F-box protein FWD1 (the mouse homologue of Drosophila Slimb and Xenopus beta-TrCP) functions as the ubiquitin ligase for IkappaBalpha. FWD1 associates with Skp1 through the F-box domain and also recognizes the conserved DSGXXS motif of IkappaBalpha. The structural requirements for the interactions of FWD1 with IkappaBalpha and with Skp1 have now been investigated further. The D31A mutation (but not the G33A mutation) in the DSGXXS motif of IkappaBalpha abolished the binding of IkappaBalpha to FWD1 and its subsequent ubiquitination without affecting the phosphorylation of IkappaBalpha. The IkappaBalpha mutant D31E still exhibited binding to FWD1 and underwent ubiquitination. These results suggest that, in addition to site specific phosphorylation at Ser(32) and Ser(36), an acidic amino acid at position 31 is required for FWD1-mediated ubiquitination of IkappaBalpha. Deletion analysis of Skp1 revealed that residues 61-143 of this protein are required for binding to FWD1. On the other hand, the highly conserved residues Pro(149), Ile(160), and Leu(164) in the F-box domain of FWD1 were dispensable for binding to Skp1. Together, these data delineate the structural requirements for the interactions among IkappaBalpha, FWD1, and Skp1 that underlie substrate recognition by the SCF ubiquitin ligase complex. PMID- 10514434 TI - Localization of the PAK1-, WASP-, and IQGAP1-specifying regions of Cdc42. AB - The Rho family small GTPase Cdc42 transmits divergent intracellular signals through multiple effector proteins to elicit cellular responses such as cytoskeletal reorganization. Potential effectors of Cdc42 implicated in mediating its cytoskeletal effect in mammalian cells include PAK1, WASP, and IQGAP1. To investigate the determinants of Cdc42-effector specificity, we utilized recombinant Cdc42 mutants and chimeras made between Cdc42 and RhoA to map the regions of Cdc42 contributing to specific effector p21-binding domain (PBD) interaction. Site-directed mutants of the switch I domain and neighboring regions of Cdc42 demonstrated differential binding patterns toward the PBDs of PAK1, WASP, and IQGAP1, suggesting that switch I provides essential determinants for the effector binding, but recognition of each effector by Cdc42 involves a distinct mechanism. Differing from Rac1, the switch I domain and the surrounding region (amino acids 29 to 55) of Cdc42 appeared to be sufficient for specific binding to PAK1, whereas determinants outside the switch I domain, residues 157 191 and 84-120 in particular, were necessary and sufficient to confer specificity to WASP and IQGAP1, respectively. In addition, IQGAP1, but not PAK1 nor WASP, required the unique "insert region," residues 122-134, of Cdc42 to achieve high affinity binding. Microinjection of the constitutively active Cdc42/RhoA chimeras into serum-starved Swiss 3T3 cells showed that although preserving PAK1- and WASP binding activity could retain the peripheral actin microspike (PAM)-inducing activity of Cdc42, interaction with PAK1 or WASP was not required for this activity. Moreover, IQGAP1-binding alone by Cdc42 was insufficient for PAM induction. Thus, Cdc42 utilizes multiple distinct structural determinants to specify different effector recognition and to elicit PAM-inducing effect. PMID- 10514435 TI - Regulation of metallothionein gene transcription. Identification of upstream regulatory elements and transcription factors responsible for cell-specific expression of the metallothionein genes from Caenorhabditis elegans. AB - Metallothioneins are small, cysteine-rich proteins that function in metal detoxification and homeostasis. Metallothionein transcription is controlled by cell-specific factors, as well as developmentally modulated and metal-responsive pathways. By using the nematode Caenorhabditis elegans as a model system, the mechanism that controls cell-specific metallothionein transcription in vivo was investigated. The inducible expression of the C. elegans metallothionein genes, mtl-1 and mtl-2, occurs exclusively in intestinal cells. Sequence comparisons of these genes with other C. elegans intestinal cell-specific genes identified multiple repeats of GATA transcription factor-binding sites (i.e. GATA elements). In vivo deletion and site-directed mutation analyses confirm that one GATA element in mtl-1 and two in mtl-2 are required for transcription. Electrophoretic mobility shift assays show that the C. elegans GATA transcription factor ELT-2 specifically binds to these elements. Ectopic expression of ELT-2 in non intestinal cells of C. elegans activates mtl-2 transcription in these cells. Likewise, mtl-2 is not expressed in nematodes in which elt-2 has been disrupted. These results indicate that cell-specific transcription of the C. elegans metallothionein genes is regulated by the binding of ELT-2 to GATA elements in these promoters. Furthermore, a model is proposed where ELT-2 constitutively activates metallothionein expression; however, a second metal-responsive factor prevents transcription in the absence of metals. PMID- 10514436 TI - Mouse semaphorin H induces PC12 cell neurite outgrowth activating Ras-mitogen activated protein kinase signaling pathway via Ca(2+) influx. AB - We recently showed that mouse semaphorin H (MSH), a secreted semaphorin molecule, acts as a chemorepulsive factor on sensory neurites. In this study, we found for the first time that MSH induces neurite outgrowth in PC12 cells in a dose dependent manner. Comparison of Ras-mitogen-activated protein kinase (MAPK) signaling pathways between MSH and nerve growth factor (NGF) revealed that these pathways are crucial for MSH action as well as NGF. K-252a, an inhibitor of tyrosine autophosphorylation of tyrosine kinase receptors (Trks), did not inhibit the action of MSH, suggesting that MSH action occurs via a different receptor than NGF. L- and N-types of voltage-dependent Ca(2+) channel blockers, diltiazem and omega-conotoxin, inhibited MSH-induced neurite outgrowth and MAPK phosphorylation in a Ca(2+)-dependent manner. A transient elevation in intracellular Ca(2+) level was observed upon MSH stimulation. These findings suggest that extracellular Ca(2+) influx, followed by activation of the Ras-MAPK signaling pathway, is required for MSH induced PC12 cell neurite outgrowth. PMID- 10514437 TI - JNK activation is required for JB6 cell transformation induced by tumor necrosis factor-alpha but not by 12-O-tetradecanoylphorbol-13-acetate. AB - Signal transduction via mitogen-activated protein kinase pathways plays a key role in a variety of cellular responses, including cell proliferation, differentiation, tumor promotion, and cell death. c-Jun N-terminal kinases (JNKs) are identified as members of the mitogen-activated protein kinase family and are known to phosphorylate and activate several transcription factors, including c Jun, ATF, and Elk-1. However, the role of JNK activation in tumor promotion is not yet defined. Because previous studies have indicated that exposure of JB6 Cl 41 cells to either 12-O-tetradecanoylphorbol-13-acetate (TPA) or tumor necrosis factor-alpha (TNF-alpha) results in cell transformation, we investigated the role of JNKs in this biological process by using dominant negative JNK(1) and the cell transformation model JB6 Cl 41 cells. Incubation of Cl 41 cells with TNF-alpha led to cell transformation and activation of JNKs. Introduction of the dominant negative mutant of JNK(1) into JB6 Cl 41 cells specifically inhibited TNF-alpha induced activation of JNKs, but not Erks and p38 kinases. Most importantly, expressing dominant negative mutant JNK(1) inhibited TNF-alpha-induced cell transformation but not TPA-induced cell transformation. Our results directly demonstrated for the first time that JNK activation is required for TNF-alpha- but not TPA-induced cell transformation. PMID- 10514438 TI - p53 negatively regulates cdc2 transcription via the CCAAT-binding NF-Y transcription factor. AB - The p53 tumor suppressor protein regulates the transcription of regulatory genes involved in cell cycle arrest and apoptosis. We have reported previously that inducible expression of the p53 gene leads to the cell cycle arrest both at G(1) and G(2)/M in association with induction of p21 and reduction of mitotic cyclins (cyclin A and B) and cdc2 mRNA. In this study, we investigated the mechanism by which p53 regulates transcription of the cdc2 gene. Transient transfection analysis showed that wild type p53 represses whereas various dominant negative mutants of p53 increase cdc2 transcription. The cdc2 promoter activity is not repressed in cells transfected with a transactivation mutant, p53(22/23). An adenovirus oncoprotein, E1B-55K inhibits the p53-mediated repression of the cdc2 promoter, while E1B-19K does not. Since the cdc2 promoter does not contain a TATA sequence, we performed deletion and point mutation analyses and identified the inverted CCAAT sequence located at -76 as a cis-acting element for the p53 mediated regulation. We found that a specific DNA-protein complex is formed at the CCAAT sequence and that this complex contains the NF-Y transcription factor. Consistently, a dominant negative mutant of the NF-YA subunit, NF-YAm29, decreases the cdc2 promoter, and p53 does not further decrease the promoter activity in the presence of NF-YAm29. These results suggest that p53 negatively regulates cdc2 transcription and that the NF-Y transcription factor is required for the p53-mediated regulation. PMID- 10514439 TI - Molecular distinction of phosphatidylcholine synthesis between the CDP-choline pathway and phosphatidylethanolamine methylation pathway. AB - In addition to the CDP-choline pathway for phosphatidylcholine (PC) synthesis, the liver has a unique phosphatidylethanolamine (PE) methyltransferase activity for PC synthesis via three methylations of the ethanolamine moiety of PE. Previous studies indicate that the two pathways are functionally different and not interchangeable even though PC is the common product of both pathways. This study was designed to test the hypothesis that these two pathways produce different profiles of PC species. The PC species from these two pathways were labeled with specific stable isotope precursors, D9-choline and D4-ethanolamine, and analyzed by electrospray tandem mass spectrometry. Our studies revealed a profound distinction in PC profiles between the CDP-choline pathway and the PE methylation pathway. PC molecules produced from the CDP-choline pathway were mainly comprised of medium chain, saturated (e.g. 16:0/18:0) species. On the other hand, PC molecules from the PE methylation pathway were much more diverse and were comprised of significantly more long chain, polyunsaturated (e.g. 18:0/20:4) species. PC species from the methylation pathway contained a higher percentage of arachidonate and were more diverse than those from the CDP-choline pathway. This profound distinction of PC profiles may contribute to the different functions of these two pathways in the liver. PMID- 10514440 TI - Muscarinic receptor stimulation increases regulators of G-protein signaling 2 mRNA levels through a protein kinase C-dependent mechanism. AB - RGS2, a member of the Regulators of G-protein Signaling family, modulates the activity of G-proteins coupled to the phosphoinositide signal transduction system, but little is known about what regulates RGS2. In human neuroblastoma SH SY5Y cells stimulation of muscarinic receptors by carbachol activates phosphoinositide signaling and also caused a rapid, large, and long lasting increase in RGS2 mRNA levels. Direct activation of protein kinase C also rapidly increased RGS2 mRNA levels. Inhibition of protein kinase C with Ro31-8220, GF109203x, or Go6976 or down-regulation of protein kinase C inhibited increases in RGS2 mRNA levels induced by carbachol or by the activation of protein kinase C. Blockade of calcium signaling did not alter carbachol-induced increases in RGS2 mRNA levels. Neither activation of epidermal growth factor receptors nor stimulation of cyclic AMP production with forskolin increased RGS2 mRNA levels. Pretreatment with actinomycin D blocked increases in RGS2 mRNA levels but caused a surprisingly small, although statistically significant, partial blockade of protein kinase C-mediated feedback inhibition of carbachol-induced phosphoinositide hydrolysis. Thus, RGS2 mRNA levels are increased by activation of muscarinic receptors coupled to the phosphoinositide signal transduction system through a protein kinase C-dependent mechanism. This action may contribute to negative feedback control of this signaling cascade, but because the small contribution to negative feedback contrasts with the large and prolonged elevations in RGS2 mRNA levels, we speculate that its primary role may be in modulating other signaling components. PMID- 10514441 TI - Thermodynamic studies of saccharide binding to artocarpin, a B-cell mitogen, reveals the extended nature of its interaction with mannotriose [3,6-Di-O-(alpha D-mannopyranosyl)-D-mannose]. AB - The thermodynamics of binding of various saccharides to artocarpin, from Artocarpus integrifolia seeds, a homotetrameric lectin (M(r) 65, 000) with one binding site per subunit, was determined by isothermal titration calorimetry measurements at 280 and 293 K. The binding enthalpies, DeltaH(b), are the same at both temperatures, and the values range from -10.94 to -47.11 kJ mol(-1). The affinities of artocarpin as obtained from isothermal titration calorimetry are in reasonable agreement with the results obtained by enzyme-linked lectin absorbent essay, which is based on the minimum amount of ligand required to inhibit horseradish peroxidase binding to artocarpin in enzyme-linked lectin absorbent essay (Misquith, S., Rani, P. G., and Surolia, A. (1994) J. Biol. Chem. 269, 30393-30401). The interactions are mainly enthalpically driven and exhibit enthalpy-entropy compensation. The order of binding affinity of artocarpin is as follows: mannotriose>Manalpha3Man>GlcNAc(2)Man(3)>MealphaMan>Man>M analpha6Man> Manalpha2Man>MealphaGlc>Glc, i.e. 7>4>2>1.4>1>0.4>0.3>0.24>0.11. The DeltaH for the interaction of Manalpha3Man, Manalpha6Man, and MealphaMan are similar and 20 kJ mol(-1) lower than that of mannotriose. This indicates that, while Manalpha3Man and Manalpha6Man interact with the lectin exclusively through their nonreducing end monosaccharide with the subsites specific for the alpha1,3 and alpha1,6 arms, the mannotriose interacts with the lectin simultaneously through all three of its mannopyranosyl residues. This study thus underscores the distinction in the recognition of this common oligosaccharide motif in comparison with that displayed by other lectins with related specificity. PMID- 10514442 TI - Hereditary pancreatitis-associated mutation asn(21) --> ile stabilizes rat trypsinogen in vitro. AB - Mutations Arg(117) --> His and Asn(21) --> Ile in human trypsinogen-I have been recently associated with hereditary pancreatitis (HP). The Arg(117) --> His substitution is believed to cause pancreatitis by stabilizing trypsin against autolytic degradation, while the mechanism of action of Asn(21) --> Ile has been unknown. In an effort to understand the effect(s) of this mutation, Thr(21) in the highly homologous rat trypsinogen-II was replaced with Asn or Ile, and the recombinant zymogens and their active trypsin forms were studied. Kinetic parameters of all three trypsins were comparable, and the active enzymes suffered autolysis at similar rates, indicating that neither catalytic properties nor proteolytic stability of trypsin are influenced by mutations at position 21. When incubated at pH 8.0, 37 degrees C, pure zymogens underwent autoactivation with concomitant trypsinolytic degradation in a Ca(2+)-dependent fashion. Thus, in the presence of 5 mM Ca(2+), autoactivation and digestion of the zymogens after Arg(117) and Lys(188) were observed, while in the presence of 1 mM EDTA autoactivation and cleavage at Lys(188) were reduced, and zymogenolysis at the Arg(117) site was enhanced. Overall rates of zymogen degradation in [Asn(21)]- and [Ile(21)]trypsinogens were higher in Ca(2+) than in EDTA, while [Thr(21)]trypsinogen demonstrated inverse characteristics. Remarkably, both in the presence and absence of Ca(2+), [Ile(21)]trypsinogen exhibited significantly higher stability against autoactivation and proteolysis than zymogens with Asn(21) or Thr(21). The observations suggest that autocatalytic trypsinogen degradation may be an important defense mechanism against excessive trypsin generation in the pancreas, and trypsinogen stabilization by the Asn(21) --> Ile mutation plays a role in the pathogenesis of HP. PMID- 10514443 TI - Transmembrane segment (TMS) VIII of the Na(+)/Citrate transporter CitS requires downstream TMS IX for insertion in the Escherichia coli membrane. AB - The amino acid sequence of the sodium ion-dependent citrate transporter CitS of K. pneumoniae contains 12 hydrophobic stretches that could form membrane-spanning segments. A previous analysis of the membrane topology in Escherichia coli using the PhoA gene fusion technique indicated that only nine of these hydrophobic segments span the membrane, while three segments, Vb, VIII and IX, were predicted to have a periplasmic location (Van Geest, M., and Lolkema, J. S. (1996) J. Biol. Chem. 271, 25582-25589). A topology study of C-terminally truncated CitS molecules in dog pancreas microsomes revealed that the protein traverses the endoplasmic reticulum membrane 11 times. In agreement with the PhoA fusion data, segment Vb was predicted to have a periplasmic location, but, in contrast, segments VIII and IX were found to be membrane-spanning (Van Geest, M., Nilsson, I., von Heijne, G., and Lolkema, J. S. (1999) J. Biol. Chem. 274, 2816-2823). In the present study, using site-directed Cys labeling, the topology of segments VIII and IX in the full-length CitS protein was determined in the E. coli membrane. Engineered cysteine residues in the loop between the two segments were accessible to a membrane-impermeable thiol reagent exclusively from the cytoplasmic side of the membrane, demonstrating that transmembrane segments (TMSs) VIII and IX are both membrane-spanning. It follows that the folding of CitS in the E. coli and endoplasmic reticulum membrane is the same. Cysteine accessibility studies of CitS-PhoA fusion molecules demonstrated that in the E. coli membrane segment VIII is exported to the periplasm in the absence of the C terminal CitS sequences, thus explaining why the PhoA fusions do not correctly predict the topology. An engineered cysteine residue downstream of TMS VIII moved from a periplasmic to a cytoplasmic location when the fusion protein containing TMSs I-VIII was extended with segment IX. Thus, downstream segment IX is both essential and sufficient for the insertion of segment VIII of CitS in the E. coli membrane. PMID- 10514444 TI - A novel subtype of class II alcohol dehydrogenase in rodents. Unique Pro(47) and Ser(182) modulates hydride transfer in the mouse enzyme. AB - Mice and rats were found to possess class II alcohol dehydrogenases with novel enzymatic and structural properties. A cDNA was isolated from mouse liver and the encoded alcohol dehydrogenase showed high identity (93.1%) with the rat class II alcohol dehydrogenase which stands in contrast to the pronounced overall variability of the class II line. The two heterologously expressed rodent class II enzymes exhibited over 100-fold lower catalytic efficiency (k(cat)/K(m)) for oxidation of alcohols as compared with other alcohol dehydrogenases and were not saturated with ethanol. Hydride transfer limited the rate of octanol oxidation as indicated by a deuterium isotope effect of 4.8. The mutation P47H improved hydride transfer and turnover rates were increased to the same level as for the human class II enzyme. Michaelis constants for alcohols and aldehydes were decreased while they were increased for the coenzyme. The rodent class II enzymes catalyzed reduction of p-benzoquinone with about the same maximal turnover as for the human form. This activity was not affected by the P47H mutation while a S182T mutation increased the K(m) value for benzoquinone 10-fold. omega-Hydroxy fatty acids were catalyzed extremely slow but functioned as potent inhibitors by binding to the enzyme-NAD(+) complex. All these data indicate that the mammalian class II alcohol dehydrogenase line is divided into two structurally and functionally distinct subgroups. PMID- 10514445 TI - Expression and functional interaction of the catalytic and regulatory subunits of human methionine adenosyltransferase in mammalian cells. AB - Methionine adenosyltransferase (MAT) catalyzes the synthesis of S adenosylmethionine (AdoMet). The mammalian MAT II isozyme consists of catalytic alpha(2) and regulatory beta subunits. The aim of this study was to investigate the interaction and kinetic behavior of the human MAT II subunit proteins in mammalian cells. COS-1 cells were transiently transfected with pTargeT vector harboring full-length cDNA that encodes for the MAT II alpha(2) or beta subunits. Expression of the His-tagged recombinant alpha(2) (ralpha(2)) subunit in COS-1 cells markedly increased MAT II activity and resulted in a shift in the K(m) for L-methionine (L-Met) from 15 microM (endogenous MAT II) to 75 microM (ralpha(2)), and with the apparent existence of two kinetic forms of MAT in the transfected COS-1 cell extracts. By contrast, expression of the recombinant beta (rbeta) subunit had no effect on the K(m) for L-Met of the endogenous MAT II, while it did cause an increase in both the V(max) and the specific activity of endogenous MAT. Co-expression of both ralpha(2) and rbeta subunits resulted in a significant increase of MAT specific activity with the appearance of a single kinetic form of MAT (K(m) = 20 microM). The recombinant MAT II alpha(2) and rbeta subunit associated spontaneously either in cell-free system or in COS-1 cells co expressing both subunits. Analysis of nickel-agarose-purified His-tagged ralpha(2) subunit from COS-1 cell extracts showed that the beta subunit co purified with the alpha(2) subunit. Furthermore, the alpha(2) and beta subunits co-migrated in native polyacrylamide gels. Together, the data provide evidence for alpha(2) and beta MAT subunit association. In addition, the beta subunit regulated MAT II activity by reducing its K(m) for L-Met and by rendering the enzyme more susceptible to feedback inhibition by AdoMet. We believe that the previously described differential expression of MAT II beta subunit may be an important mechanism by which MAT activity can be modulated to provide different levels of AdoMet that may be required at different stages of cell growth and differentiation. PMID- 10514446 TI - Peroxynitrite inactivates tryptophan hydroxylase via sulfhydryl oxidation. Coincident nitration of enzyme tyrosyl residues has minimal impact on catalytic activity. AB - Tryptophan hydroxylase, the initial and rate-limiting enzyme in serotonin biosynthesis, is inactivated by peroxynitrite in a concentration-dependent manner. This effect is prevented by molecules that react directly with peroxynitrite such as dithiothreitol, cysteine, glutathione, methionine, tryptophan, and uric acid but not by scavengers of superoxide (superoxide dismutase), hydroxyl radical (Me(2)SO, mannitol), and hydrogen peroxide (catalase). Assuming simple competition kinetics between peroxynitrite scavengers and the enzyme, a second-order rate constant of 3.4 x 10(4) M(-1) s(-1) at 25 degrees C and pH 7.4 was estimated. The peroxynitrite-induced loss of enzyme activity was accompanied by a concentration-dependent oxidation of protein sulfhydryl groups. Peroxynitrite-modified tryptophan hydroxylase was resistant to reduction by arsenite, borohydride, and dithiothreitol, suggesting that sulfhydryls were oxidized beyond sulfenic acid. Peroxynitrite also caused the nitration of tyrosyl residues in tryptophan hydroxylase, with a maximal modification of 3.8 tyrosines/monomer. Sodium bicarbonate protected tryptophan hydroxylase from peroxynitrite-induced inactivation and lessened the extent of sulfhydryl oxidation while causing a 2-fold increase in tyrosine nitration. Tetranitromethane, which oxidizes sulfhydryls at pH 6 or 8, but which nitrates tyrosyl residues at pH 8 only, inhibited tryptophan hydroxylase equally at either pH. Acetylation of tyrosyl residues with N-acetylimidazole did not alter tryptophan hydroxylase activity. These data suggest that peroxynitrite inactivates tryptophan hydroxylase via sulfhydryl oxidation. Modification of tyrosyl residues by peroxynitrite plays a relatively minor role in the inhibition of tryptophan hydroxylase catalytic activity. PMID- 10514447 TI - Scavenger receptor class B, type I, mediates selective uptake of low density lipoprotein cholesteryl ester. AB - Scavenger receptor, class B, type I (SR-BI) is a cell-surface glycoprotein that mediates selective uptake of high density lipoprotein cholesteryl ester (CE) without the concomitant uptake and degradation of the particle. We have investigated the endocytic and selective uptake of low density lipoprotein (LDL) CE by SR-BI using COS-7 cells transiently transfected with mouse SR-BI. Analysis of lipoprotein uptake data showed a concentration-dependent LDL-CE-selective uptake when doubly labeled LDL particles were incubated with SR-BI-expressing COS 7 cells. In contrast to vector-transfected cells, SR-BI-expressing COS-7 cells showed marked increases in LDL cell association and CE uptake by the selective uptake pathway, but only a modest increase in CE uptake by the endocytic pathway. SR-BI-mediated LDL-CE-selective uptake exceeded LDL endocytic uptake by 50-100 fold. SR-BI-mediated LDL-CE-selective uptake was not inhibited by the proteoglycan synthesis inhibitor, p-nitrophenyl-beta-D-xylopyranoside or by the sulfation inhibitor sodium chlorate, indicating that SR-BI-mediated LDL-CE uptake occurs independently of LDL interaction with cell-surface proteoglycan. Analyses with subclones of Y1 adrenocortical cells showed that LDL-CE-selective uptake was proportional to the level of SR-BI expression. Furthermore, antibody directed to the extracellular domain of SR-BI blocked LDL-CE-selective uptake in adrenocortical cells. Thus, in cells that normally express SR-BI and in transfected COS-7 cells SR-BI mediates the efficient uptake of LDL-CE via the selective uptake mechanism. These results suggest that SR-BI may influence the metabolism of apoB-containing lipoproteins in vivo by mediating LDL-CE uptake into SR-BI-expressing cells. PMID- 10514448 TI - Cytosine arabinoside lesions are position-specific topoisomerase II poisons and stimulate DNA cleavage mediated by the human type II enzymes. AB - Cytosine arabinoside (araC) is an important drug used for the treatment of human leukemias. In order to exert its cytotoxic effects, araC must be incorporated into chromosomal DNA. Although specific DNA lesions that involve base loss or modification stimulate nucleic acid cleavage mediated by type II topoisomerases, the effects of deoxyribose sugar ring modification on enzyme activity have not been examined. Therefore, the effects of incorporated araC residues on the DNA cleavage/religation equilibrium of human topoisomerase IIalpha and beta were characterized. AraC lesions were position-specific topoisomerase II poisons and stimulated DNA scission mediated by both human type II enzymes. However, the positional specificity of araC residues differed from that previously reported for other cleavage-enhancing DNA lesions. Finally, additive or synergistic increases in DNA cleavage were observed in the presence of araC lesions and etoposide. These findings broaden the range of DNA lesions known to alter topoisomerase II function and raise the possibility that this enzyme may mediate some of the cellular effects of araC. PMID- 10514449 TI - The diabetes-associated 3243 mutation in the mitochondrial tRNA(Leu(UUR)) gene causes severe mitochondrial dysfunction without a strong decrease in protein synthesis rate. AB - Cells harboring patient-derived mitochondria with an A-to-G transition at nucleotide position 3243 of their mitochondrial DNA display severe loss of respiration when compared with cells containing the wild-type adenine but otherwise identical mitochondrial DNA sequence. The amount and degree of leucylation of tRNA(Leu(UUR)) were both found to be highly reduced in mutant cells. Despite the low level of leucyl-tRNA(Leu(UUR)), the rate of mitochondrial translation was not seriously affected by this mutation. Therefore, decrease of mitochondrial protein synthesis as such does not appear to be a necessary prerequisite for loss of respiration. Rather, the mitochondrially encoded proteins seem subject to elevated degradation, leading to a severe reduction in their steady state levels. Our results favor a scheme in which the 3243 mutation causes loss of respiration through accelerated protein degradation, leading to a disequilibrium between the levels of mitochondrial and nuclear encoded respiratory chain subunits and thereby a reduction of functional respiratory chain complexes. The possible mechanisms underlying the pathogenesis of mitochondrial diabetes is discussed. PMID- 10514450 TI - Identification of a bile acid-responsive element in the human ileal bile acid binding protein gene. Involvement of the farnesoid X receptor/9-cis-retinoic acid receptor heterodimer. AB - Intestinal bile acid-binding protein (I-BABP) is a cytosolic protein that binds bile acids (BAs) with a high affinity. In the small intestine, its expression is restricted to the ileum where it is involved in the enterohepatic circulation of BAs. Using the human enterocyte-like Caco-2 cell line, we have recently shown that BAs increased I-BABP gene expression. To determine whether this regulation occurs in vivo, the effect of BA depletion or supplementation was studied in mice. A dramatic drop in I-BABP mRNA levels was observed in mice treated with the BA-binding resin cholestyramine, whereas an increase was found in animals fed with taurocholic acid. BAs are physiological ligands for the nuclear farnesoid X receptor (FXR). Both FXR and I-BABP are co-expressed along the small intestine and in Caco-2 cells. To determine the role of FXR in the regulation of I-BABP expression, the promoter of the human I-BABP gene was cloned. In Caco-2 cells, cotransfection of FXR and RXRalpha is required to obtain the full transactivation of the I-BABP promoter by BAs. Deletion and mutation analyses demonstrate that the FXR/RXRalpha heterodimer activates transcription through an inverted repeat bile acid responsive element located in position -160/-148 of the human I-BABP promoter. In conclusion, we show that FXR is a physiological BA sensor that is likely to play an essential role in BA homeostasis through the regulation of genes involved in their enterohepatic circulation. PMID- 10514451 TI - Reactivity of tetrahydrobiopterin bound to nitric-oxide synthase. AB - Levels of tetrahydrobiopterin (BH(4)) bound to nitric-oxide synthase (NOS) were examined during multiple turnovers of the enzyme in the presence of an NADPH regenerating system. Our findings show that NOS-bound BH(4) does not remain in a static state but undergoes redox reactions. Under these experimental conditions, the redox state of BH(4) was determined by the balance between calcium/calmodulin (Ca(2+)/CaM)-dependent oxidation of BH(4) mediated by the uncoupled formation of superoxide/hydrogen peroxide on the one hand and by reductive regeneration of BH(4) on the other hand. BH(4) oxidation was appreciably increased in the presence of arginine. Levels of NOS-bound BH(4) were also examined under single turnover conditions in the absence of an NADPH-regenerating system and in the presence of added superoxide dismutase and catalase to suppress the accumulation of superoxide and hydrogen peroxide. BH(4) oxidation was again dependent on Ca(2+)/CaM. The insensitivity to superoxide dismutase and catalase suggested that the single turnover oxidation of BH(4) did not proceed through superoxide/peroxide, although the involvement of these oxidants could not be definitively excluded. The amount of BH(4) oxidized was highest in the presence of arginine, and this oxidation significantly exceeded that in the presence of N(G)-hydroxy-L-arginine. The findings that single turnover oxidation of BH(4) is stimulated by arginine in the presence of Ca(2+)/CaM and that BH(4) is regenerated are consistent with a role for the pterin as an electron donor in product formation; this role remains to be defined. PMID- 10514452 TI - The procyclin repertoire of Trypanosoma brucei. Identification and structural characterization of the Glu-Pro-rich polypeptides. AB - The surface of the insect stages of the protozoan parasite Trypanosoma brucei is covered by abundant glycosyl phosphatidylinositol (GPI)-anchored glycoproteins known as procyclins. One type of procyclin, the EP isoform, is predicted to have 22-30 Glu-Pro (EP) repeats in its C-terminal domain and is encoded by multiple genes. Because of the similarity of the EP isoform sequences and the heterogeneity of their GPI anchors, it has been impossible to separate and characterize these polypeptides by standard protein fractionation techniques. To facilitate their structural and functional characterization, we used a combination of matrix-assisted laser desorption ionization and electrospray mass spectrometry to analyze the entire procyclin repertoire expressed on the trypanosome cell. This analysis, which required removal of the GPI anchors by aqueous hydrofluoric acid treatment and cleavage at aspartate-proline bonds by mild acid hydrolysis, provided precise information about the glycosylation state and the number of Glu-Pro repeats in these proteins. Using this methodology we detected in a T. brucei clone the glycosylated products of the EP3 gene and two different products of the EP1 gene (EP1-1 and EP1-2). Furthermore, only low amounts of the nonglycosylated products of the GPEET and EP2 genes were detected. Because all procyclin genes are transcribed polycistronically, the latter finding indicates that the expression of the GPEET and EP2 genes is post-transcriptionaly regulated. This is the first time that the whole procyclin repertoire from procyclic trypanosomes has been characterized at the protein level. PMID- 10514453 TI - Rubredoxin from the green sulfur bacterium Chlorobium tepidum functions as an electron acceptor for pyruvate ferredoxin oxidoreductase. AB - Rubredoxin (Rd) from the moderately thermophilic green sulfur bacterium Chlorobium tepidum was found to function as an electron acceptor for pyruvate ferredoxin oxidoreductase (PFOR). This enzyme, which catalyzes the conversion of pyruvate to acetyl-CoA and CO(2), exhibited an absolute dependence upon the presence of Rd. However, Rd was incapable of participating in the pyruvate synthase or CO(2) fixation reaction of C. tepidum PFOR, for which two different reduced ferredoxins are employed as electron donors. These results suggest a specific functional role for Rd in pyruvate oxidation and provide the initial indication that the two important physiological reactions catalyzed by PFOR/pyruvate synthase are dependent on different electron carriers in the cell. The UV-visible spectrum of oxidized Rd, with a monomer molecular weight of 6500, gave a molar absorption coefficient at 492 nm of 6.89 mM(-1) cm(-1) with an A(492)/A(280) ratio of 0.343 and contained one iron atom/molecule. Further spectroscopic studies indicated that the CD spectrum of oxidized C. tepidum Rd exhibited a unique absorption maximum at 385 nm and a shoulder at 420 nm. The EPR spectrum of oxidized Rd also exhibited unusual anisotropic resonances at g = 9.675 and g = 4.322, which is composed of a narrow central feature with broader shoulders to high and low field. The midpoint reduction potential of C. tepidum Rd was determined to be -87 mV, which is the most electronegative value reported for Rd from any source. PMID- 10514454 TI - Differential effect of retinoic acid on growth regulation by phorbol ester in human cancer cell lines. AB - Phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) and all-trans-retinoic acid (trans-RA) are potent regulators of growth of cancer cells. In this study, we investigated the effect of TPA and trans-RA alone or their combination on proliferation of human breast cancer ZR75-1 and T47D and lung cancer H460 and H292 cell lines. trans-RA caused various degrees of growth inhibition of these cell lines. However, TPA showed inhibition of proliferation of H460 and H292 cells and induction of ZR75-1 cell growth. Although trans-RA did not significantly regulate the growth inhibitory effect of TPA, it completely prevented its growth stimulating function. The divergent effects of TPA were associated with specific disruption of cell cycle events, an induction of G(0)/G(1) arrest in H460 and H292 cells and inhibition of G(0)/G(1) arrest with increase of S phase in ZR75-1 cells. Induction of G(0)/G(1) arrest was accompanied by induction of p21(WAF1) and ERK activity, whereas inhibition of G(0)/G(1) arrest was associated with enhanced activity of JNK and AP-1 but not ERK. trans-RA did not affect TPA-induced p21(WAF1) expression. However, it inhibited TPA-induced AP-1 activity in ZR75-1 cells and the constitutive AP-1 activity in H460 and H292 cells. Thus, trans-RA modulates TPA activity through its interaction through TPA-induced JNK/AP-1 pathway but not TPA-induced ERK/p21(WAF1) pathway. PMID- 10514455 TI - Phosphatidic acid synthesis in mitochondria. Topography of formation and transmembrane migration. AB - The topography of formation and migration of phosphatidic acid (PA) in the transverse plane of rat liver mitochondrial outer membrane (MOM) were investigated. Isolated mitochondria and microsomes, incubated with sn-glycerol 3 phosphate and an immobilized substrate palmitoyl-CoA-agarose, synthesized both lyso-PA and PA. The mitochondrial and microsomal acylation of glycerophosphate with palmitoyl-CoA-agarose was 80-100% of the values obtained in the presence of free palmitoyl-CoA. In another series of experiments, both free polymyxin B and polymyxin B-agarose stimulated mitochondrial glycerophosphate acyltransferase activity approximately 2-fold. When PA loaded mitochondria were treated with liver fatty acid binding protein, a fifth of the phospholipid left the mitochondria. The amount of exportable PA reduced with the increase in the time of incubation. In another approach, PA-loaded mitochondria were treated with phospholipase A(2). The amount of phospholipase A(2)-sensitive PA reduced when the incubation time was increased. Taken together, the results suggest that lysophosphatidic acid (LPA) and PA are synthesized on the outer surface of the MOM and that PA moves to the inner membrane presumably for cardiolipin formation. PMID- 10514456 TI - Differential phosphorylation paradigms dictate desensitization and internalization of the N-formyl peptide receptor. AB - Following activation by ligand, most G protein-coupled receptors undergo rapid phosphorylation. This is accompanied by a drastic decrease in the efficacy of continued or repeated stimulation, due to receptor uncoupling from G protein and receptor internalization. Such processing steps have been shown to be absolutely dependent on receptor phosphorylation in the case of the N-formyl peptide receptor (FPR). In this study, we report results that indicate that the mechanisms responsible for desensitization and internalization are distinct. Using site-directed mutagenesis of the serine and threonine residues of the FPR carboxyl terminus, we have characterized regions that differentially regulate these two processes. Whereas substitution of all 11 Ser/Thr residues in the carboxyl terminus prevents both desensitization and internalization, substitution of four Ser/Thr residues between 328-332 blocks desensitization but has no effect on internalization. Similarly, substitution of four Ser/Thr residues between positions 334 and 339 results in a deficit in desensitization but again no decrease in internalization, suggesting that phosphorylation at either site evokes receptor internalization, whereas maximal desensitization requires phosphorylation at both sites. These results also indicate that receptor internalization is not involved in the process of desensitization. Further analysis of the residues between 328-332 revealed that restoration either of Ser(328) and Thr(329) or of Thr(331) and Ser(332) was sufficient to restore desensitization, suggesting that phosphorylation within either of these two sites, in addition to sites between residues 334 and 339, is sufficient to produce desensitization. Taken together, these results indicate that the mechanisms involved in FPR processing (uncoupling from G proteins and internalization) are regulated differentially by phosphorylation at distinct sites within the carboxyl terminus of the FPR. The relevance of this paradigm to other G protein-coupled receptors is discussed. PMID- 10514457 TI - Calcium influx through L-type channels is required for selective activation of extracellular signal-regulated kinase by gonadotropin-releasing hormone. AB - The hypothalamic decapeptide gonadotropin-releasing hormone stimulates mobilization of two discrete pools of calcium in clonal (alphaT3-1) and primary pituitary gonadotropes. A multidisciplinary approach was implemented to investigate the effects of discrete calcium fluctuations on the signaling pathways linking the gonadotropin-releasing hormone receptor to activation of mitogen-activated protein kinases and immediate early genes. Blockade of calcium influx through nifedipine-sensitive voltage-gated calcium channels reduced buserelin-induced activation of extracellular signal-regulated kinase (ERK) and c Fos while activation of c-Jun N-terminal kinase and c-Jun was unaffected. Inhibition of buserelin-stimulated ERK activity by nifedipine was also observed in rat pituitary cells in primary culture. Direct activation of alphaT3-1 cell L type calcium channels with the agonist Bay-K 8644 resulted in phosphorylation of ERK and induction of c-Fos. However, simple voltage-induced channel activation did not produce a sufficient calcium signal, since depolarization with 35 mM KCl failed to induce activation of ERK. Depletion of intracellular calcium stores with thapsigargin did not affect buserelin-induced ERK activation. An inhibitor of protein kinase C decreased calcium influx through nifedipine-sensitive calcium channels and phosphorylation of ERK induced by buserelin. Pharmacological inhibition of protein kinase C did not block Bay-K 8644-induced ERK activation. These observations suggest that calcium influx through L-type channels is required for GnRH-induced activation of ERK and c-Fos and that the influence of calcium lies downstream of protein kinase C. PMID- 10514458 TI - Novel protein-disulfide isomerases from the early-diverging protist Giardia lamblia. AB - Protein-disulfide isomerase is essential for formation and reshuffling of disulfide bonds during nascent protein folding in the endoplasmic reticulum. The two thioredoxin-like active sites catalyze a variety of thiol-disulfide exchange reactions. We have characterized three novel protein-disulfide isomerases from the primitive eukaryote Giardia lamblia. Unlike other protein-disulfide isomerases, the giardial enzymes have only one active site. The active-site sequence motif in the giardial proteins (CGHC) is characteristic of eukaryotic protein-disulfide isomerases, and not other members of the thioredoxin superfamily that have one active site, such as thioredoxin and Dsb proteins from Gram-negative bacteria. The three giardial proteins have very different amino acid sequences and molecular masses (26, 50, and 13 kDa). All three enzymes were capable of rearranging disulfide bonds, and giardial protein-disulfide isomerase 2 also displayed oxidant and reductant activities. Surprisingly, the three giardial proteins also had Ca(2+)-dependent transglutaminase activity. This is the first report of protein-disulfide isomerases with a single active site that have diverse roles in protein cross-linking. This study may provide clues to the evolution of key functions of the endoplasmic reticulum in eukaryotic cells, protein disulfide formation, and isomerization. PMID- 10514459 TI - Identification of a bipartite nuclear localization sequence necessary for nuclear import of 5-lipoxygenase. AB - 5-Lipoxygenase catalyzes the synthesis of leukotrienes from arachidonic acid. This enzyme can reside either in the cytoplasm or the nucleus; its subcellular distribution is influenced by extracellular factors, and its nuclear import correlates with changes in leukotriene synthetic capacity. To identify sequences responsible for the nuclear import of 5-lipoxygenase, we transfected NIH 3T3 cells and RAW 264.7 macrophages with expression vectors encoding various 5 lipoxygenase constructs fused to green fluorescent protein. Overexpression of wild type 5-lipoxygenase with or without fusion to green fluorescent protein resulted in a predominantly intranuclear pattern of fluorescence, similar to the distribution of native 5-lipoxygenase in primary alveolar macrophages. Within the 5-lipoxygenase protein is a sequence (Arg(638)-Lys(655)) that closely resembles a bipartite nuclear localization signal. Studies using deletion mutants indicated that this region was necessary for nuclear import of 5-lipoxygenase. Analysis of mutants containing specific amino acid substitutions within this sequence confirmed that it was this sequence that was necessary for nuclear import of 5 lipoxygenase and that a specific arginine residue was critical for this function. As nuclear import of 5-lipoxygenase may regulate leukotriene production, natural or induced mutations in this bipartite nuclear localization sequence may also be important in affecting leukotriene synthesis. PMID- 10514460 TI - Expression of functionally distinct variants of the beta(4)A integrin subunit in relation to the differentiation state in human intestinal cells. AB - Integrins are important mediators of cell-laminin interactions. In the small intestinal epithelium, which consists of spatially separated proliferative and differentiated cell populations located, respectively, in the crypt and on the villus, laminins and laminin-binding integrins are differentially expressed along the crypt-villus axis. One exception to this is the integrin alpha(6)beta(4), which is thought to be ubiquitously expressed by intestinal cells. However, in this study, a re-evaluation of the beta(4) subunit expression with different antibodies revealed that two forms of beta(4) exist in the human intestinal epithelium. Furthermore, we show that differentiated enterocytes express a full length 205-kDa beta(4)A subunit, whereas undifferentiated crypt cells express a novel beta(4)A subunit that does not contain the COOH-terminal segment of the cytoplasmic domain (beta(4)A(ctd-)). This new form was not found to arise from alternative beta(4) mRNA splicing. Moreover, we found that these two beta(4)A forms can associate into alpha(6)beta(4)A complexes; however, the beta(4)A(ctd-) integrin expressed by the undifferentiated crypt cells is not functional for adhesion to laminin-5. Hence, these studies identify a novel alpha(6)beta(4)A(ctd ) integrin expressed in undifferentiated intestinal crypt cells that is functionally distinct. PMID- 10514461 TI - Human inhibitor antibodies specific for the factor VIII A2 domain disrupt the interaction between the subunit and factor IXa. AB - Factor VIIIa, a heterotrimer of the A1, A2, and A3-C1-C2 subunits, increases the catalytic efficiency for factor IXa-catalyzed activation of factor X. A significant fraction of naturally occurring, anti-factor VIII inhibitor antibodies reacts with the A2 domain. Utilizing the capacity for isolated A2 subunit to stimulate factor IXa activity, we show that a panel of these inhibitors block this activity. Inhibition of activity parallels the antibody potency as measured in the Bethesda assay. These antibodies also block the A2 dependent increases in fluorescence anisotropy of fluorescein-Phe-Phe-Arg factor IXa. Similar to the IgG fractions, a peptide representing the sequence of the inhibitor epitope (A2 residues 484-509) blocked the A2-dependent stimulation of factor IXa. These results indicate that antibodies possessing this specificity directly inhibit the interaction of A2 subunit with factor IXa, thus abrogating the contribution of this subunit to cofactor activity. Furthermore, these results also suggest that factor VIII residues 484-509 contribute to a factor IXa interactive site. PMID- 10514462 TI - Bcl-2 and mitochondrial oxygen radicals. New approaches with reactive oxygen species-sensitive probes. AB - Investigations into the capacity of the Bcl-2 protein to prevent apoptosis have targeted mitochondria as key sites of the preventative action accorded by Bcl-2 to cells. Using novel approaches with fluorescence probes and autofluorescence detection of endogenous NAD(P)H, we have examined the effects of expressing Bcl-2 in the Bcl-2 negative Burkitt's lymphoma cell line Daudi. We evaluated for the first time the effect of Bcl-2 expression on the intracellular distribution and production of hydrogen peroxide, under basal conditions and after treatment with apoptosis inducing agents, ceramide analogs and tumor necrosis factor (TNF) alpha. Increased availability of mitochondrial NAD(P)H was detected in Bcl-2 expressing cells and was correlated with an increased constitutive mitochondrial production of hydrogen peroxide. Although production of hydrogen peroxide was increased by either C(6)-ceramide or TNF-alpha in Bcl-2 negative Daudi cells commensurate with the early phases of apoptosis, this increase did not occur in Bcl-2-expressing cells. Thus, Bcl-2 appears to allow cells to adapt to an increased state of oxidative stress, fortifying the cellular anti-oxidant defenses and counteracting the radical overproduction imposed by different cell death stimuli. Furthermore, we report altered cytological features of mitochondria during the early phases of apoptosis induced by C(6)-ceramide and TNF-alpha. In particular, mitochondria changed in appearance, clustering in the perinuclear region and Bcl-2 expression prevented these changes from occurring. PMID- 10514463 TI - The relative strengths of SR protein-mediated associations of alternative and constitutive exons can influence alternative splicing. AB - We have characterized the functional role of SR protein-mediated exon/exon associations in the alternative splicing of exon 5 of chicken cardiac troponin T (cTnT). We have previously shown that SR proteins can promote the association of the alternative exon 5 with the flanking constitutive exon 6 of this pre-mRNA. In this study, we have shown that when exons 2, 3, and 4 of the cTnT pre-mRNA are spliced together, the composite exon 2/3/4 contains an additional SR protein binding site. Furthermore, we have found that SR proteins can also promote interactions between the pairs of exons 2/3/4-5 and 2/3/4-6. We then asked whether the SR protein binding sites in these exons play a role in cTnT alternative splicing in vivo. We found that the SR protein binding sites in exons 2/3/4 and 6 promote exon 5 skipping, and it has previously been shown that the SR protein binding site in exon 5 promotes exon 5 inclusion. Consistent with these results, we find that the SR protein-mediated association of exon 2/3/4 with 6 is preferred over associations involving exon 5, in that exons 2/3/4 and 6 are more efficient than exon 5 in competing an SR protein-mediated exon/exon association. We suggest that the relative strengths of SR protein-mediated associations of alternative and constitutive exons play a role in determining alternative splicing patterns. PMID- 10514464 TI - The apical Na(+)/H(+) exchanger isoform NHE3 is regulated by the actin cytoskeleton. AB - The epithelial isoform of the Na(+)/H(+) exchanger, NHE3, associates with at least two related regulatory factors called NHERF1/EBP50 and NHERF2/TKA-1/E3KARP. These factors in addition interact with the cytoskeletal protein ezrin, which in turn binds to actin. The possible linkage of NHE3 with the cytoskeleton prompted us to test the effect of actin-modifying agents on NHE3 activity. Cytochalasins B and D and latrunculin B, which interfere with actin polymerization, induced a profound inhibition of NHE3 activity. The effect was isoform-specific inasmuch as the "housekeeping" exchanger NHE1 was virtually unaffected. Cytoskeletal disorganization was associated with a subcellular redistribution of NHE3, which accumulated at sites where actin aggregated, suggesting a physical interaction of exchangers with the cytoskeleton. An interaction was further suggested by the co sedimentation of a detergent-insoluble fraction of NHE3 with the actin cytoskeleton. Inhibition of transport was not due to diminution in the number of transporters at the plasmalemma. Functional analyses of NHE1/NHE3 chimeras revealed that the cytoplasmic domain of NHE3 conferred sensitivity to cytochalasin B. Progressive carboxyl-terminal and internal deletions of the cytoplasmic region of NHE3 indicated that the region between residues 650 and 684 is critical for this response. This region overlaps with the domain reported to interact with NHERF and also contains a putative ezrin-binding site; hence, it likely plays a role in interactions with the cytoskeleton. PMID- 10514465 TI - BiP-binding sequences in HIV gp160. Implications for the binding specificity of bip. AB - BiP, a resident endoplasmic reticulum member of the HSP70 family of molecular chaperones, associates transiently with a wide variety of newly synthesized exocytotic proteins. In addition to immunoglobulin heavy and light chains, the first natural substrates identified for BiP, a number of viral polypeptides including the human immunodeficiency virus type 1 envelope glycoprotein gp160 interact with BiP during their passage through the endoplasmic reticulum. We have used a computer algorithm developed to predict BiP-binding sites within protein primary sequences to identify sites within gp160 that might mediate its association with BiP. Analysis of the ability of 22 synthetic heptapeptides corresponding to predicted binding sites to stimulate the ATPase activity of BiP or to compete with an unfolded polypeptide for binding to BiP indicated that about half of them are indeed recognized by the chaperone. All of the confirmed binding sites are localized within conserved regions of gp160, suggesting a conserved role for BiP in the folding of gp160. Information on the characteristics of confirmed BiP-binding peptides gained in this and previous studies has been utilized to improve the predictive power of the BiP Score algorithm and to investigate the differences in peptide binding specificities of HSP70 family members. PMID- 10514467 TI - Male-specific modification of human CD52. AB - CD52 is an unusually short, bipolar glycopeptide bearing a highly charged N linked carbohydrate moiety and a glycosylphosphatidylinositol membrane anchor. It is exclusively expressed on lymphocytes and in the male genital tract where it is shed into the seminal plasma and inserts into the sperm membrane. The sperm surface molecule has potential significance as a target for antibodies that inhibit sperm function and gamete interaction. Western blot analyses suggested cell type-specific modifications of the antigen. It was purified from seminal plasma and a detailed structural analysis performed. The majority of anchor structures in male genital tract CD52 showed 2-inositol palmitoylation, rendering molecules insensitive toward phospholipase C, and a sn-1-alkyl-2-lyso-glycerol structure in place of the diacylated anchor described by Treumann et al. (Treumann, A., Lifely, M. R., Schneider, P., and Ferguson, M. A. (1995) J. Biol. Chem. 270, 6088-6099). N-Glycans of the male genital tract product were based on bi-, tri-, and tetraantennary structures of highly charged (up to -7), terminally sialylated complex-type sugars. A substantial proportion carried varying numbers of lactosamine repeats of which nearly 30% were branched. Different from lymphocytes, 10-15% of all N-glycans of the male genital tract antigen also contained peripheral fucose. These data confirm that male genital tract CD52 is distinct from the lymphocyte form by both N-linked glycans and COOH-terminal attached lipid anchor. PMID- 10514466 TI - Muscle as the primary site of urea cycle enzyme activity in an alkaline lake adapted tilapia, Oreochromis alcalicus grahami. AB - The tilapia fish Oreochromis alcalicus grahami from Kenya has adapted to living in waters at pH 10.5 by excreting the end product of nitrogen metabolism as urea rather than as ammonia directly across the gills as occurs in most fish. The level of activity in liver of the first enzyme in the urea cycle pathway, carbamoyl-phosphate synthetase III (CPSase III), is too low to account for the observed high rates of urea excretion. We report here the surprising finding that CPSase III and all other urea cycle enzyme activities are present in muscle of this species at levels more than sufficient to account for the rate of urea excretion; in addition, the basic kinetic properties of the CPSase III appear to be different from those of other known type III CPSases. The sequence of the CPSase III cDNA is reported as well as the finding that glutamine synthetase activity is present in liver but not in muscle. This unusual form of adaptation may have occurred because of the apparent impossibility of packaging the needed amount of urea cycle enzymes in liver. PMID- 10514469 TI - SecA is not required for signal recognition particle-mediated targeting and initial membrane insertion of a nascent inner membrane protein. AB - In Escherichia coli, signal recognition particle (SRP)-dependent targeting of inner membrane proteins has been described. In vitro cross-linking studies have demonstrated that short nascent chains exposing a highly hydrophobic targeting signal interact with the SRP. This SRP, assisted by its receptor, FtsY, mediates the transfer to a common translocation site in the inner membrane that contains SecA, SecG, and SecY. Here we describe a further in vitro reconstitution of SRP mediated membrane insertion in which purified ribosome-nascent chain-SRP complexes are targeted to the purified SecYEG complex contained in proteoliposomes in a process that requires the SRP-receptor FtsY and GTP. We found that in this system SecA and ATP are dispensable for both the transfer of the nascent inner membrane protein FtsQ to SecY and its stable membrane insertion. Release of the SRP from nascent FtsQ also occurred in the absence of SecYEG complex indicating a functional interaction of FtsY with lipids. These data suggest that SRP/FtsY and SecB/SecA constitute distinct targeting routes. PMID- 10514470 TI - Structural basis for recognition of phosphorylated high mannose oligosaccharides by the cation-dependent mannose 6-phosphate receptor. AB - Mannose 6-phosphate receptors (MPRs) play an important role in the targeting of newly synthesized soluble acid hydrolases to the lysosome in higher eukaryotic cells. These acid hydrolases carry mannose 6-phosphate recognition markers on their N-linked oligosaccharides that are recognized by two distinct MPRs: the cation-dependent mannose 6-phosphate receptor and the insulin-like growth factor II/cation-independent mannose 6-phosphate receptor. Although much has been learned about the MPRs, it is unclear how these receptors interact with the highly diverse population of lysosomal enzymes. It is known that the terminal mannose 6-phosphate is essential for receptor binding. However, the results from several studies using synthetic oligosaccharides indicate that the binding site encompasses at least two sugars of the oligosaccharide. We now report the structure of the soluble extracytoplasmic domain of a glycosylation-deficient form of the bovine cation-dependent mannose 6-phosphate receptor complexed to pentamannosyl phosphate. This construct consists of the amino-terminal 154 amino acids (excluding the signal sequence) with glutamine substituted for asparagine at positions 31, 57, 68, and 87. The binding site of the receptor encompasses the phosphate group plus three of the five mannose rings of pentamannosyl phosphate. Receptor specificity for mannose arises from protein contacts with the 2-hydroxyl on the terminal mannose ring adjacent to the phosphate group. Glycosidic linkage preference originates from the minimization of unfavorable interactions between the ligand and receptor. PMID- 10514468 TI - STAT5b down-regulates peroxisome proliferator-activated receptor alpha transcription by inhibition of ligand-independent activation function region-1 trans-activation domain. AB - Growth hormone-activated STAT5b inhibits by up to 80% the transcriptional activity of peroxisome proliferator-activated receptor (PPAR) alpha, a nuclear receptor activated by diverse environmental chemicals and hypolipidemic drugs classified as peroxisome proliferators. This inhibitory cross-talk between STAT5b and PPAR is now reported for PPAR forms gamma and delta and for thyroid hormone receptor, indicating a more general potential for inhibitory cross-talk between JAK/STAT and nuclear receptor signaling pathways. Further investigations revealed that SOCS-3, a growth hormone-inducible negative regulator of cytokine signaling to STAT5b, abolished the STAT5b inhibitory response. A constitutively active STAT5b mutant failed to inhibit PPARalpha activity, indicating that STAT5b does not induce synthesis of a more proximal PPARalpha inhibitor. STAT5b inhibition was not reversed by overexpression of the heterodimerization partner of PPAR (retinoid X receptor) or the nuclear receptor coactivators P300 and SRC-1, suggesting that STAT5b does not inhibit PPARalpha by competing for these limiting cellular cofactors. STAT5b did not inhibit a chimeric receptor comprised of yeast GAL4 DNA-binding domain linked to the ligand binding/AF-2 trans-activation domain of PPARalpha, indicating that the COOH-terminal AF-2 domain of PPAR is not the target of STAT5b inhibition. Rather, STAT5b inhibited transcription driven by the NH(2)-terminal ligand-independent AF-1 trans-activation domain of PPARalpha in a GAL4-linked chimera by approximately 80%. The conservation of this AF-1 trans activation function in many nuclear receptors suggests that AF-1 may serve as an important target for inhibitory cross-talk between STAT transcription factors and nuclear receptors in a variety of signaling pathways. PMID- 10514472 TI - Cytochrome c-mediated apoptosis in cells lacking mitochondrial DNA. Signaling pathway involving release and caspase 3 activation is conserved. AB - Mitochondria serve as a pivotal component of the apoptotic cell death machinery. However, cells that lack mitochondrial DNA (rho(0) cells) retain apparently normal apoptotic signaling. In the present study, we examined mitochondrial mechanisms of apoptosis in rho(0) osteosarcoma cells treated with staurosporine. Immunohistochemistry revealed that rho(0) cells maintained a normal cytochrome c distribution in mitochondria even though these cells were deficient in respiration. Upon staurosporine treatment, cytochrome c was released concomitantly with activation of caspase 3 and loss of mitochondrial membrane potential (Deltapsi(m)). After mitochondrial loss of cytochrome c, rho(0) cells underwent little change in glutathione (GSH) redox potential whereas a dramatic oxidation in GSH/glutathione disulfide (GSSG) pool occurred in parental rho(+) cells. These results show that mitochondrial signaling of apoptosis via cytochrome c release was preserved in cells lacking mtDNA. However, intracellular oxidation that normally accompanies apoptosis was lost, indicating that the mitochondrial respiratory chain provides the major source of redox signaling in apoptosis. PMID- 10514471 TI - Characterization of human and murine PMP20 peroxisomal proteins that exhibit antioxidant activity in vitro. AB - We have isolated the cDNAs encoding human and mouse homologues of a yeast protein, termed peroxisomal membrane protein 20 (PMP20). Comparison of the amino acid sequences of human (HsPMP20) and mouse (MmPMP20) PMP20 proteins revealed a high degree of identity (93%), whereas resemblance to the yeast Candida boidinii PMP20A and PMP20B (CbPMP20A and CbPMP20B) was less (30% identity). Both HsPMP20 and MmPMP20 lack transmembrane regions, as do CbPMP20A and CbPMP20B. HsPMP20 mRNA expression was low in human fetal tissues, especially in the brain. In adult tissues, HsPMP20 mRNA was expressed in the majority of tissues tested. HsPMP20 and MmPMP20 contained the C-terminal tripeptide sequence Ser-Gln-Leu (SQL), which is similar to the peroxisomal targeting signal 1 utilized for protein import into peroxisomes. HsPMP20 bound directly to the human peroxisomal targeting signal 1 receptor, HsPEX5. Mutagenesis analysis showed that the C-terminal tripeptide sequence, SQL, of HsPMP20 is necessary for its binding to HsPEX5. Subcellular fractionation of HeLa cells, expressing epitope-tagged PMP20, revealed that HsPMP20 is localized in the cytoplasm and in a particulate fraction containing peroxisomes. Double-staining immunofluorescence studies showed colocalization of HsPMP20 and thiolase, a bona fide peroxisomal protein. The amino acid sequence alignment of HsPMP20, MmPMP20, CbPMP20A, and CbPMP20B displayed high similarity to thiol-specific antioxidant proteins. HsPMP20 exerted an inhibitory effect on the inactivation of glutamine synthetase in the thiol metal-catalyzed oxidation system but not in the nonthiol metal-catalyzed oxidation system, suggesting that HsPMP20 possesses thiol-specific antioxidant activity. In addition, HsPMP20 removed hydrogen peroxide by its thiol-peroxidase activity. These results indicate that HsPMP20 is imported into the peroxisomal matrix via PEX5p and may work to protect peroxisomal proteins against oxidative stress. Because some portion of PMP20 might also be present in the cytosol, HsPMP20 may also have a protective effect in the cytoplasm. PMID- 10514473 TI - An atypical mitogen-activated protein kinase (MAPK) homologue expressed in gametocytes of the human malaria parasite Plasmodium falciparum. Identification of a MAPK signature. AB - The cDNA encoding Pfmap-2, an enzyme of the human malaria parasite Plasmodium falciparum, was cloned, sequenced, and expressed in Escherichia coli. The open reading frame carried by the Pfmap-2 cDNA encodes a 508-amino acid polypeptide of 59.2 kDa with maximal homology to mitogen-activated protein kinases (MAPKs) from various organisms. The purified recombinant enzyme displayed functional characteristics of MAPKs such as (i) ability to undergo autophosphorylation, (ii) ability to phosphorylate myelin basic protein, a classical MAPK substrate, (iii) regulation of kinase activity by a MAPK-specific phosphatase, and (iv) ability to be activated by component(s) present in cell extracts. Mutational analysis of the recombinant protein allowed the identification of residues that are important for enzymatic activity. Northern blot analysis and immunofluorescence assays indicated that Pfmap-2 is expressed specifically in gametocytes, the form that is responsible for transmission of the parasite to the mosquito vector. Gametocyte extracts activated recombinant Pfmap-2 more efficiently than extracts from asexual parasites, which is consistent with this stage specificity. Despite its overall high level of homology to MAPKs, Pfmap-2 presents the peculiarity of not possessing the conserved threonine-X-tyrosine activation motif usually found in enzymes of this family; instead, it has a threonine-serine-histidine at the same location. This atypical feature formed the basis for a detailed analysis of the primary structure of MAPKs, allowing us to define an operational MAPK signature, which is shared by Pfmap-2. The fact that no MAPK from vertebrates diverge in the activation motif suggests that the fine mechanisms of Pfmap-2 regulation may offer an opportunity for antimalarial drug targeting. PMID- 10514474 TI - Oligodendrocyte programmed cell death and central myelination deficiency induced in transgenic mice by synergism between c-Myc and Oct-6. AB - The basic helix-loop-helix transcription factor c-Myc is a potent trigger of programmed cell death when overexpressed during late oligodendrocyte development in transgenic mice. Here we provide evidence that c-Myc can act synergistically with the Pit, Oct, Unc homeodomain transcription factor Oct-6 to produce myelin disease pathogenesis in transgenic mice. More than 70% of c-myc/Oct-6 bitransgenic mice, obtained from crosses between phenotypically normal heterozygous mice of various My (c-Myc) and Oc (Oct-6) transgenic strains that express c-myc and oct-6 transgenes under transcriptional control of the myelin basic protein gene, developed severe neurological disturbances characterized by action tremors, recurrent seizures, and premature death. Affected bitransgenic mice exhibited multiple hypomyelinated lesions in the white matter that did not stain with myelin-specific antibodies against myelin basic protein, proteolipid protein, CNPase, and myelin-associated glycoprotein. The mice also exhibited a larger number of terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end-labeling positive cells in the white matter as well as ultrastructural evidence of glial cell death and astrogliosis. These observations indicate that the myelin lesions observed in the c-myc/oct-6 bitransgenic mice result from the untimely programmed cell death of oligodendroglia and that the c-myc and oct-6 transgenes act synergistically in producing the lesions. PMID- 10514475 TI - Functional association of type IIA secretory phospholipase A(2) with the glycosylphosphatidylinositol-anchored heparan sulfate proteoglycan in the cyclooxygenase-2-mediated delayed prostanoid-biosynthetic pathway. AB - An emerging body of evidence suggests that type IIA secretory phospholipase A(2) (sPLA(2)-IIA) participates in the amplification of the stimulus-induced cyclooxygenase (COX)-2-dependent delayed prostaglandin (PG)-biosynthetic response in several cell types. However, the biological importance of the ability of sPLA(2)-IIA to bind to heparan sulfate proteoglycan (HSPG) on cell surfaces has remained controversial. Here we show that glypican, a glycosylphosphatidylinositol (GPI)-anchored HSPG, acts as a physical and functional adaptor for sPLA(2)-IIA. sPLA(2)-IIA-dependent PGE(2) generation by interleukin-1-stimulated cells was markedly attenuated by treatment of the cells with heparin, heparinase or GPI-specific phospholipase C, which solubilized the cell surface-associated sPLA(2)-IIA. Overexpression of glypican-1 increased the association of sPLA(2)-IIA with the cell membrane, and glypican-1 was coimmunoprecipitated by the antibody against sPLA(2)-IIA. Glypican-1 overexpression led to marked augmentation of sPLA(2)-IIA-mediated arachidonic acid release, PGE(2) generation, and COX-2 induction in interleukin-1-stimulated cells, particularly when the sPLA(2)-IIA expression level was suboptimal. Immunofluorescent microscopic analyses of cytokine-stimulated cells revealed that sPLA(2)-IIA was present in the caveolae, a microdomain in which GPI-anchored proteins reside, and also appeared in the perinuclear area in proximity to COX-2. We therefore propose that a GPI-anchored HSPG glypican facilitates the trafficking of sPLA(2)-IIA into particular subcellular compartments, and arachidonic acid thus released from the compartments may link efficiently to the downstream COX-2-mediated PG biosynthesis. PMID- 10514476 TI - Interaction of macrophage-stimulating protein with its receptor. Residues critical for beta chain binding and evidence for independent alpha chain binding. AB - Macrophage-stimulating protein (MSP) and hepatocyte growth factor/scatter factor (HGF/SF) are plasminogen-related growth and motility factors that interact with cell-surface protein tyrosine kinase receptors. Each one is a heterodimeric protein comprising a disulfide-linked alpha chain and a serine protease-like beta chain. Despite structural similarities between MSP and HGF, the primary receptor binding site is located on the alpha chain of HGF/SF but on the beta chain of MSP. To obtain insight into the structural basis for MSP beta chain binding, beta chain structure was modeled from coordinates of an existing model of the HGF beta chain. The model revealed that the region corresponding to the S1 specificity pocket in trypsin is filled by the Asn(682)/Glu(648) interacting pair, leaving a shallow cavity for possible beta chain interaction with the receptor. Mutants in this region were created, and their binding characteristics were determined. A double mutation of Asn(682)/Glu(648) caused diminished binding of the beta chain to the MSP receptor, and a single mutation of neighboring Arg(683) completely abolished binding. Thus, this region of the molecule is critical for binding. We also found that at equimolar concentrations of free alpha and beta chains, alpha chain binding to receptor was detectable, at levels considerably lower than beta chain binding. The EC(50) values determined by quantitative enzyme-linked immunosorbent assay are 0.25 and 16.9 nM for beta and alpha chain, respectively. The data suggest that MSP has two independent binding sites with high and low affinities located in beta and alpha chain, respectively, and that the two sites together mediate receptor dimerization and subsequent activation. PMID- 10514478 TI - Characterization of the sequence of interactions of the fusion domain of the simian immunodeficiency virus with membranes. Role of the membrane dipole potential. AB - The simian immunodeficiency virus fusion peptide constitutes a 12-residue N terminal segment of the gp32 protein that is involved in the fusion between the viral and cellular membranes, facilitating the penetration of the virus in the host cell. Simian immunodeficiency virus fusion peptide is a hydrophobic peptide that in Me(2)SO forms aggregates that contain beta-sheet pleated structures. When added to aqueous media the peptide forms large colloidal aggregates. In the presence of lipidic membranes, however, the peptide interacts with the membranes and causes small changes of the membrane electrostatic potential as shown by fluorescein phosphatidylethanolamine fluorescence. Thioflavin T fluorescence and Fourier transformed infrared spectroscopy measurements reveal that the interaction of the peptide with the membrane bilayer results in complete disassembly of the aggregates originating from an Me(2)SO stock solution. Above a lipid/peptide ratio of about 5, the membrane disaggregation and water precipitation processes become dependent on the absolute peptide concentration rather than on the lipid/peptide ratio. A schematic mechanism is proposed, which sheds light on how peptide-peptide interactions can be favored with respect to peptide-lipid interactions at various lipid/peptide ratios. These studies are augmented by the use of the fluorescent dye 1-(3-sulfonatopropyl)-4-[beta[2-(di-n octylamino)-6-naphthyl]vinyl ] pyridinium betaine that shows the interaction of the peptide with the membranes has a clear effect on the magnitude of the so called dipole potential that arises from dipolar groups located on the lipid molecules and oriented water molecules at the membrane-water interface. It is shown that the variation of the membrane dipole potential affects the extent of the membrane fusion caused by the peptide and implicates the dipolar properties of membranes in their fusion. PMID- 10514479 TI - cdk6 can shorten G(1) phase dependent upon the N-terminal INK4 interaction domain. AB - Deregulated activity of cdk4 or cdk6 can lead to inappropriate cellular proliferation and tumorigenesis accompanied by unchecked inactivation of the retinoblastoma tumor suppressor protein. Certain tumor types preferentially activate either cdk4 or cdk6, suggesting that these kinases may not be equivalently oncogenic in all cell types. Although it is clear that cdk4 can act as an oncogene at least in part by evading inhibition by p16(INK4a), the role of cdk6 in tumorigenesis is less well understood. To investigate the consequences of aberrant expression of cdk6, the requirements for proliferation caused by cdk6 overexpression were studied. cdk6-transfected U2OS cells displayed an accelerated progression through G(1) phase that was dependent on kinase activity and that did not correlate with p27 binding. Furthermore, a mutation that prevents cdk6 interaction with INK4 proteins (cdk6R31C) was found to inactivate the proliferative effect of cdk6 and increase cytoplasmic localization, despite the fact that this mutant could phosphorylate the retinoblastoma protein in vitro. Together, these data suggest a role for the cdk6 INK4 interaction domain in the generation of functional, nuclear cdk6 complexes and demonstrate the importance of elevated cdk6 kinase activity in G(1) acceleration. PMID- 10514477 TI - Mutants of interleukin 13 with altered reactivity toward interleukin 13 receptors. AB - Interleukin 13 (IL13) belongs to a family of cytokines whose members exhibit structural homology, despite amino acid sequence dissimilarity. For example, while of limited sequence homology, IL13 and IL4 share a signaling receptor, IL13/4 receptor, on a variety of human normal cells. However, a subclass of IL4 independent IL13 receptors is overexpressed on certain transformed cells, including human malignant gliomas. We introduced mutations into human (h) IL13 to determine the site(s) involved in interaction with the shared receptor and/or the glioma-associated receptor. This analysis identified at least three protein regions that are needed for signaling through the shared receptor. These regions were localized to alpha-helices A, C, and D and were mainly separate from the region(s) needed to interact with the glioma-associated receptor. Glutamic acids at positions 13 and 16 in hIL13 alpha-helix A, arginine and serine at positions 66 and 69 in helix C, and arginine at position 109 in helix D were found to be important in inducing biological signaling since their specific mutation resulted in loss and/or gain of function phenomena. We demonstrate that the molecular requirements of hIL13 to interact with its respective receptors are generally distinct and can be controlled by mutagenesis of the cytokine. PMID- 10514481 TI - Oncogenic Ras sensitizes cells to apoptosis by Par-4. AB - Certain mutations in the mammalian ras gene are oncogenic and are often detected in human cancers. Oncogenic Ras induces the transcription activity of NF-kappaB that confers cell survival. Oncogenic Ras also down-modulates the expression of Par-4, a transcriptional repressor protein, that is essential but not sufficient on its own to induce apoptosis. Here we show that reintroduction of Par-4 by transient transfection leads to apoptosis in cells expressing oncogenic Ras but not in those that lack oncogenic Ras expression. Par-4 abrogates oncogenic Ras inducible NF-kappaB transcription activity but does not interfere with cytoplasmic activation, or the DNA binding activity, of NF-kappaB. Because abrogation of NF-kappaB transcription activity is sufficient to cause apoptosis in cells expressing oncogenic Ras, our findings identify Par-4 as a novel example of a pro-apoptotic protein that selectively inhibits oncogenic Ras-dependent NF kappaB function at the transcription level and suggest a mechanism by which Par-4 expression may selectively induce apoptosis in oncogenic Ras-expressing cells. PMID- 10514480 TI - Endocytosis of ligand-human parathyroid hormone receptor 1 complexes is protein kinase C-dependent and involves beta-arrestin2. Real-time monitoring by fluorescence microscopy. AB - Endocytosis and intracellular trafficking of the human parathyroid hormone receptor subtype 1 (hPTH1-Rc) and its ligands was monitored independently by real time fluorescence microscopy in stably transfected HEK-293 cells. Complexes of fluorescence-labeled parathyroid hormone (PTH)-(1-34) agonist bound to the hPTH1 Rc internalized rapidly at 37 degrees C via clathrin-coated vesicles, whereas fluorescent PTH-(7-34) antagonist-hPTH1Rc complexes did not. A functional C terminus epitope-tagged receptor (C-Tag-hPTH1-Rc) was immunolocalized to the cell membrane and, to a lesser extent, the cytoplasm. PTH and PTH-related protein agonists stimulated C-Tag-hPTH1-Rc internalization. Relocalization to the cell membrane occurred 1 h after removal of the ligand. Endocytosis of fluorescent PTH agonist-hPTH1-Rc complexes was blocked by the protein kinase C (PKC) inhibitor staurosporine but not by the specific protein kinase A inhibitor N-(2 (methylamino)ethyl)-5-isoquinoline-sulfonamide. Fluorescent PTH antagonist-hPTH1 Rc complexes were rapidly internalized after PKC activation by phorbol 12 myristate 13-acetate or thrombin, but not after stimulation of the cAMP/protein kinase A pathway by forskolin. In cells co-expressing the hPTH1-Rc and a green fluorescent protein-beta-arrestin2 fusion protein (beta-Arr2-GFP), PTH agonists stimulated beta-Arr2-GFP mobilization to the cell membrane. Subsequently, fluorescent PTH-(1-34)-hPTH1Rc complexes and beta-Arr2-GFP co-localized intracellularly. In conclusion, agonist-activated hPTH1-Rc internalization involves beta-arrestin mobilization and targeting to clathrin-coated vesicles. Our results also indicate that receptor occupancy, rather than receptor-mediated signaling, is necessary, although not sufficient, for endocytosis of the hPTH1 Rc. Activation of PKC, however, is absolutely required. PMID- 10514482 TI - YY1 binds five cis-elements and trans-activates the myeloid cell-restricted gp91(phox) promoter. AB - Four transcriptional activating cis-elements within the gp91(phox) promoter bind a protein complex of similar mobility and binding specificity, denoted BID (binding increased during differentiation). The intensity of BID complexes increases upon myeloid cell differentiation, coincident with induction of gp91(phox) expression, and BID competes with the transcriptional repressor CDP for binding to each of these promoter elements. To determine the identity of BID, an expression library was ligand screened with the BID-binding site that surrounds the -145-base pair (bp) region of the gp91(phox) promoter. One recovered factor that exhibits the expected binding specificity is YY1, a ubiquitous multifunctional transcription factor. BID complexes that form with the four binding sites within the gp91(phox) promoter are disrupted by YY1 antiserum, and a fifth YY1-binding site was detected in the -412-bp promoter region. Overexpression of YY1 in transient co-transfection assays trans-activates a minimal promoter containing two copies of the -145-bp binding site from the gp91(phox) promoter. Neither the level of YY1 protein nor DNA binding activity increases during myeloid cell differentiation. These studies identify a target gene of YY1 function in mature myeloid cells, and demonstrate that YY1 function can be controlled during myeloid development by the modulation of a competing DNA binding factor. PMID- 10514483 TI - Mutational analysis of the antagonist-binding site of the histamine H(1) receptor. AB - We combined in a previously derived three-dimensional model of the histamine H(1) receptor (Ter Laak, A. M., Timmerman, H., Leurs, H., Nederkoorn, P. H. J., Smit, M. J., and Donne-Op den Kelder, G. M. (1995) J. Comp. Aid. Mol. Design. 9, 319 330) a pharmacophore for the H(1) antagonist binding site (Ter Laak, A. M., Venhorst, J., Timmerman, H., and Donne-Op de Kelder, G. M. (1994) J. Med. Chem. 38, 3351-3360) with the known interacting amino acid residue Asp(116) (in transmembrane domain III) of the H(1) receptor and verified the predicted receptor-ligand interactions by site-directed mutagenesis. This resulted in the identification of the aromatic amino acids Trp(167), Phe(433), and Phe(436) in transmembrane domains IV and VI of the H(1) receptor as probable interaction points for the trans-aromatic ring of the H(1) antagonists. Subsequently, a specific interaction of carboxylate moieties of two therapeutically important, zwitterionic H(1) antagonists with Lys(200) in transmembrane domain V was predicted. A Lys(200) --> Ala mutation results in a 50- (acrivastine) to 8-fold (d-cetirizine) loss of affinity of these zwitterionic antagonists. In contrast, the affinities of structural analogs of acrivastine and cetirizine lacking the carboxylate group, triprolidine and meclozine, respectively, are unaffected by the Lys(200) --> Ala mutation. These data strongly suggest that Lys(200), unique for the H(1) receptor, acts as a specific anchor point for these "second generation" H(1) antagonists. PMID- 10514484 TI - Unique lipoproteins secreted by primary astrocytes from wild type, apoE (-/-), and human apoE transgenic mice. AB - Composition of central nervous system lipoproteins affects the metabolism of lipoprotein constituents within the brain. The epsilon4 allele of apolipoprotein E (apoE) is a risk factor for Alzheimer's disease via an unknown mechanism(s). As glia are the primary central nervous system cell type that synthesize apoE, we characterized lipoproteins secreted by astrocytes from wild type (WT), apoE (-/ ), and apoE transgenic mice expressing human apoE3 or apoE4 in a mouse apoE (-/-) background. Nondenaturing size exclusion chromatography demonstrates that WT, apoE3, and apoE4 astrocytes secrete particles the size of plasma high density lipoprotein (HDL) composed of phospholipid, free cholesterol, and protein, primarily apoE and apoJ. However, the lipid:apoE ratio of particles containing human apoE is significantly lower than WT. ApoE localizes across HDL-like particle sizes. ApoJ localizes to the smallest HDL-like particles. ApoE (-/-) astrocytes secrete little phospholipid or free cholesterol despite comparable apoJ expression, suggesting that apoE is required for normal secretion of astrocyte lipoproteins. Further, particles were not detected in apoE (-/-) samples by electron microscopy. Nondenaturing immunoprecipitation experiments indicate that apoE and apoJ reside predominantly on distinct particles. These studies suggest that apoE expression influences the unique structure of astrocyte lipoproteins, a process further modified by apoE species. PMID- 10514485 TI - Prenylated prelamin A interacts with Narf, a novel nuclear protein. AB - Prelamin A is farnesylated and methylated on the cysteine residue of a carboxyl terminal CaaX motif. In the nucleus, prelamin A is processed to lamin A by endoproteolytic removal of the final 18 amino acids, including the farnesylated cysteine residue. Using the yeast two-hybrid assay, we isolated a novel human protein, Narf, that binds the carboxyl-terminal tail of prelamin A. Narf has limited homology to iron-only bacterial hydrogenases and eukaryotic proteins of unknown function. Narf is encoded by a 2-kilobase mRNA expressed in all human cell lines and tissues examined. The protein is detected in the nuclear fraction of HeLa cell lysates on Western blots and can be extracted from nuclear envelopes with 0.5 M NaCl. When a FLAG epitope-tagged Narf is expressed in HeLa cells, it is exclusively nuclear and partially co-localizes with the nuclear lamina. The farnesylation status of prelamin A determines its ability to bind to Narf. Inhibition of farnesyltransferase and mutation or deletion of the CaaX motif from the prelamin A tail domain inhibits Narf binding in yeast two-hybrid and in vitro binding assays. The prenyl-dependent binding of Narf to prelamin A is an important first step in understanding the functional significance of the lamin A precursor. PMID- 10514486 TI - Substrate gating confers steroid specificity to estrogen sulfotransferase. AB - Estrogen sulfotransferase (EST) exhibits a high substrate specificity and catalytic efficiency toward estrogens such as estradiol (E2) but insignificant ability to sulfate hydroxysteroids such as dehydroepiandrosterone (DHEA). To provide the structural basis for this estrogen specificity, we mutated amino acid residues that constitute the substrate-binding site of EST. Among these mutants, only Tyr-81 decreased E2 and increased DHEA sulfotransferase activities. Substitution for Tyr-81 by smaller hydrophobic residues increased K(m(E2)) for E2 activity, whereas the k(cat(E2)) remained relatively constant. The Y81L mutant exhibited the same DHEA activity as wild-type hydroxysteroid sulfotransferase, for which K(m(DHEA)) remained relatively constant, and k(cat(DHEA)) was markedly increased. The side chain of Tyr-81 is directed at the A-ring of the E2 molecule in the substrate-binding pocket of EST, constituting a steric gate with Phe-142 sandwiching E2 from the opposite side. The present mutagenesis study indicates that the 3beta-hydroxyl group of the DHEA molecule is excluded from the catalytic site of EST through steric hindrance of Tyr-81 with the C-19 methyl group of DHEA. Thus, this stricture-like gating caused by steric hindrance appears to be a structural principle for conferring estrogen specificity to EST. PMID- 10514487 TI - Fatty acylation of phospholipase D1 on cysteine residues 240 and 241 determines localization on intracellular membranes. AB - We have reported previously that phospholipase D1 (PLD1) is labeled specifically with [(3)H]palmitate following transient expression and immunoprecipitation and that this modification appeared important both for membrane localization and catalytic activity. In this work we identify by mutagenesis that the acylation sites on PLD1 are cysteine residues 240 and 241, with the cysteine at position 241 accounting for most but not all of the modification. Replacement of both cysteine residues with either serines or alanines resulted in a mutant protein that contained undetectable [(3)H]palmitate. In comparison with the wild type protein, the double mutant showed reduced catalytic activity in vivo, whereas its activity in vitro was unchanged. In addition, the localization of the double mutant was altered in comparison with the wild type protein, whereas wild type PLD1 is primarily on intracellular membranes and on punctate structures, the double mutant was on plasma membrane. Because cysteines 240 and 241 lie within a putative pleckstrin homology domain of PLD1, it is likely that fatty acylation on these residues modulates the function of the PLD1 pleckstrin homology domain. PMID- 10514488 TI - Increased levels of nuclear SREBP-1c associated with fatty livers in two mouse models of diabetes mellitus. AB - Hepatic steatosis is common in non-insulin-dependent diabetes and can be associated with fibrosis and cirrhosis in a subset of individuals. Increased rates of fatty acid synthesis have been reported in livers from rodent models of diabetes and may contribute to the development of steatosis. Sterol regulatory element-binding proteins (SREBPs) are a family of regulated transcription factors that stimulate lipid synthesis in liver. In the current studies, we measured the content of SREBPs in livers from two mouse models of diabetes, obese ob/ob mice and transgenic aP2-SREBP-1c436 (aP2-SREBP-1c) mice that overexpress nuclear SREBP 1c only in adipose tissue. The aP2-SREBP-1c mice exhibit a syndrome that resembles congenital generalized lipodystrophy in humans. Both lines of mice develop hyperinsulinemia, hyperglycemia, and hepatic steatosis. Nuclear SREBP-1c protein levels were significantly elevated in livers from ob/ob and aP2-SREBP-1c mice compared with wild-type mice. Increased nuclear SREBP-1c protein was associated with elevated mRNA levels for known SREBP target genes involved in fatty acid biosynthesis, which led to significantly higher rates of hepatic fatty acid synthesis in vivo. These studies suggest that increased levels of nuclear SREBP-1c contribute to the elevated rates of hepatic fatty acid synthesis that leads to steatosis in diabetic mice. PMID- 10514489 TI - Purification, molecular cloning, and expression of a human stratum corneum trypsin-like serine protease with possible function in desquamation. AB - A new human 33-kDa serine protease was purified from human epidermis, and its cDNA was cloned from a keratinocyte library, from mRNA from a human keratinocyte line (HaCat) and from mRNA from human skin. Polyclonal antibodies specific for the new protein detected three groups of proteins in partially purified extracts of cornified eptihelium of human plantar skin. The three components are proposed to correspond to proenzyme, active enzyme, and proteolytically modified active enzyme. After N-deglycosylation, there was a decrease in apparent molecular mass of all detected components. Expression of the cloned cDNA in a eukaryotic virus derived system yielded a recombinant protein that could be converted to an active protease by treatment with trypsin. Polymerase chain reaction analyses of cDNA from a number of human tissues showed high expression of the new enzyme in the skin and low expression in brain, placenta, and kidney. Homology searches yielded the highest score for porcine enamel matrix protease (55% amino acid sequence homology). High scores were also obtained for human and mouse neuropsin and for human stratum corneum chymotryptic enzyme. The function of this new protease, tentatively named stratum corneum tryptic enzyme, may be related to stratum corneum turnover and desquamation in the epidermis. PMID- 10514490 TI - Respiratory epithelial cell expression of vascular cell adhesion molecule-1 and its up-regulation by rhinovirus infection via NF-kappaB and GATA transcription factors. AB - Virus infections, the majority of which are rhinovirus infections, are the major cause of asthma exacerbations. Asthma now affects one-fifth of the population, yet treatment of exacerbations is unsatisfactory, and the pathogenesis is unclear. Intraepithelial lymphocyte and eosinophil infiltration and activation are strongly implicated, but the mechanisms regulating these processes are unknown. We hypothesized that lower airway epithelial expression of vascular cell adhesion molecule-1 (VCAM-1) may be important in intraepithelial inflammation and that expression would be induced by pro-inflammatory stimuli and rhinovirus infection. We investigated respiratory epithelial cell VCAM-1 expression and its regulation to identify new targets for treatment of virus-induced asthma exacerbations. We observed constitutive respiratory epithelial cell VCAM-1 expression and that rhinovirus infection, but no other pro-inflammatory stimuli tested increased VCAM-1 cell surface expression in respiratory epithelial cell lines and primary bronchial epithelial cells. We then observed rhinovirus induction of VCAM-1 mRNA expression, promoter activity, and mRNA transcription. Rhinovirus induction of VCAM-1 promoter activity was critically dependent on up regulation of proteins binding to the -254/-251 and -239/-236 GATA-binding sites and to the -72/-63 and -57/-48 NF-kappaB-binding sites in the VCAM-1 promoter. These studies identify VCAM-1 and the NF-kappaB and GATA transcription factor families as new targets for development of therapeutic interventions for virus induced asthma exacerbations. PMID- 10514491 TI - Phospholipase C binds to the receptor-like GPR1 protein and controls pseudohyphal differentiation in Saccharomyces cerevisiae. AB - The hormone receptor-like protein Gpr1p physically interacts with phosphatidylinositol-specific phospholipase C (Plc1p) and with the Galpha protein Gpa2p, as shown by two-hybrid assays and co-immune precipitation of epitope tagged proteins. Plc1p binds to Gpr1p in either the presence or absence of Gpa2, whereas the Gpr1p/Gpa2p association depends on the presence of Plc1p. Genetic interactions between the null mutations plc1Delta, gpr1Delta, gpa2Delta, and ras2Delta suggest that Plc1p acts together with Gpr1p and Gpa2p in a growth control pathway operating in parallel to the Ras2p function. Diploid cells lacking Gpr1p, Plc1p, or Gpa2p fail to form pseudohyphae upon nitrogen depletion, and the filamentation defect of gpr1Delta and plc1Delta strains is rescued by activating a mitogen-activated protein kinase pathway via STE11-4 or by activating a cAMP pathway via overexpressed Tpk2p. Plc1p is also required for efficient expression of the FG(TyA)::lacZ reporter gene under nitrogen depletion. In conclusion, we have identified two physically interacting proteins, Gpr1p and Plc1p, as novel components of a nitrogen signaling pathway controlling the developmental switch from yeast-like to pseudohyphal growth. Our data suggest that phospholipase C modulates the interaction of the putative nutrient sensor Gpr1p with the Galpha protein Gpa2p as a downstream effector of filamentation control. PMID- 10514492 TI - The role of SOCS-3 in leptin signaling and leptin resistance. AB - We earlier demonstrated that leptin induces expression of SOCS-3 mRNA in the hypothalamus. Furthermore, transfection data suggest that SOCS-3 is an inhibitor of leptin signaling. However, little is known about the regulation of SOCS-3 expression by leptin and the mechanism by which SOCS-3 inhibits leptin action. We here show that in CHO cells stably expressing the long form of the leptin receptor (CHO-OBRl), leptin induces transient expression of endogenous SOCS-3 mRNA but not of CIS, SOCS-1, or SOCS-2 mRNA. SOCS-3 protein levels were maximal after 2-3 h of leptin treatment and remained elevated at 20 h. Furthermore, in leptin-pretreated CHO-OBRl cells, proximal leptin signaling was blocked for more than 20 h after pretreatment, thus correlating with increased SOCS-3 expression. Leptin pretreatment did not affect cell surface expression of leptin receptors as measured by (125)I-leptin binding assays. In transfected COS cells, forced expression of SOCS-3 results in inhibition of leptin-induced tyrosine phosphorylation of JAK2. Finally, JAK2 co-immunoprecipitates with SOCS-3 in lysates from leptin-treated COS cells. These results suggest that SOCS-3 is a leptin-regulated inhibitor of proximal leptin signaling in vivo. Excessive SOCS-3 activity in leptin-responsive cells is therefore a potential mechanism for leptin resistance, a characteristic feature in human obesity. PMID- 10514493 TI - Sequences required for induction of neurotensin receptor gene expression during neuronal differentiation of N1E-115 neuroblastoma cells. AB - The promoter region of the mouse high affinity neurotensin receptor (Ntr-1) gene was characterized, and sequences required for expression in neuroblastoma cell lines that express high affinity NT-binding sites were characterized. Me(2)SO induced neuronal differentiation of N1E-115 neuroblastoma cells increased both the expression of the endogenous Ntr-1 gene and reporter genes driven by NTR-1 promoter sequences by 3-4-fold. Deletion analysis revealed that an 83-base pair promoter region containing the transcriptional start site is required for Me(2)SO activation. Detailed mutational analysis of this region revealed that a CACCC box and the central region of a large GC-rich palindrome are the crucial cis regulatory elements required for Me(2)SO induction. The CACCC box is bound by at least one factor that is induced upon Me(2)SO treatment of N1E-115 cells. The Me(2)SO effect was found to be both selective and cell type-restricted. Basal expression in the neuroblastoma cell lines required a distinct set of sequences, including an Sp1-like sequence, and a sequence resembling an NGFI-A-binding site; however, a more distal 5' sequence was found to repress basal activity in N1E-115 cells. These results provide evidence that Ntr-1 gene regulation involves both positive and negative regulatory elements located in the 5'-flanking region and that Ntr-1 gene activation involves the coordinate activation or induction of several factors, including a CACCC box binding complex. PMID- 10514494 TI - Direct interaction of the trans-Golgi network membrane protein, TGN38, with the F actin binding protein, neurabin. AB - TGN38 is a type I integral membrane protein that constitutively cycles between the trans-Golgi network (TGN) and plasma membrane. The cytosolic domain of TGN38 interacts with AP2 clathrin adaptor complexes via the tyrosine-containing motif ( SDYQRL-) to direct internalization from the plasma membrane. This motif has previously been shown to direct both internalization and subsequent TGN targeting of TGN38. We have used the cytosolic domain of TGN38 in a two-hybrid screen, and we have identified the brain-specific F-actin binding protein neurabin-I as a TGN38-binding protein. We demonstrate a direct interaction between TGN38 and the ubiquitous homologue of neurabin-I, neurabin-II (also called spinophilin). We have used a combination of yeast two-hybrid and in vitro protein interaction assays to show that this interaction is dependent on the serine (but not tyrosine) residue of the known TGN38 trafficking motif. We show that TGN38 interacts with the coiled coil region of neurabin in vitro and binds preferentially with the dimeric form of neurabin. TGN38 and neurabin also interact in vivo as demonstrated by coimmunoprecipitation from stably transfected PC12 cells. These data suggest that neurabin provides a direct physical link between TGN38-containing membranes and the actin cytoskeleton. PMID- 10514495 TI - Collagenase-3 binds to a specific receptor and requires the low density lipoprotein receptor-related protein for internalization. AB - We have previously identified a specific receptor for collagenase-3 that mediates the binding, internalization, and degradation of this ligand in UMR 106-01 rat osteoblastic osteosarcoma cells. In the present study, we show that collagenase-3 binding is calcium-dependent and occurs in a variety of cell types, including osteoblastic and fibroblastic cells. We also present evidence supporting a two step mechanism of collagenase-3 binding and internalization involving both a specific collagenase-3 receptor and the low density lipoprotein receptor-related protein. Ligand blot analysis shows that (125)I-collagenase-3 binds specifically to two proteins ( approximately 170 kDa and approximately 600 kDa) present in UMR 106-01 cells. Western blotting identified the 600-kDa protein as the low density lipoprotein receptor-related protein. Our data suggest that the 170-kDa protein is a specific collagenase-3 receptor. Low density lipoprotein receptor-related protein-null mouse embryo fibroblasts bind but fail to internalize collagenase-3, whereas UMR 106-01 and wild-type mouse embryo fibroblasts bind and internalize collagenase-3. Internalization, but not binding, is inhibited by the 39-kDa receptor-associated protein. We conclude that the internalization of collagenase 3 requires the participation of the low density lipoprotein receptor-related protein and propose a model in which the cell surface interaction of this ligand requires a sequential contribution from two receptors, with the collagenase-3 receptor acting as a high affinity primary binding site and the low density lipoprotein receptor-related protein mediating internalization. PMID- 10514496 TI - Exonuclease X of Escherichia coli. A novel 3'-5' DNase and Dnaq superfamily member involved in DNA repair. AB - DNA exonucleases are critical for DNA replication, repair, and recombination. In the bacterium Escherichia coli there are 14 DNA exonucleases including exonucleases I-IX (including the two DNA polymerase I exonucleases), RecJ exonuclease, SbcCD exonuclease, RNase T, and the exonuclease domains of DNA polymerase II and III. Here we report the discovery and characterization of a new E. coli exonuclease, exonuclease X. Exonuclease X is a member of a superfamily of proteins that have homology to the 3'-5' exonuclease proofreading subunit (DnaQ) of E. coli DNA polymerase III. We have engineered and purified a (His)(6) exonuclease X fusion protein and characterized its activity. Exonuclease X is a potent distributive exonuclease, capable of degrading both single-stranded and duplex DNA with 3'-5' polarity. Its high affinity for single-strand DNA and its rapid catalytic rate are similar to the processive exonucleases RecJ and exonuclease I. Deletion of the exoX gene exacerbated the UV sensitivity of a strain lacking RecJ, exonuclease I, and exonuclease VII. When overexpressed, exonuclease X is capable of substituting for exonuclease I in UV repair. As we have proposed for the other single-strand DNA exonucleases, exonuclease X may facilitate recombinational repair by pre-synaptic and/or post-synaptic DNA degradation. PMID- 10514497 TI - Identification of flow-dependent endothelial nitric-oxide synthase phosphorylation sites by mass spectrometry and regulation of phosphorylation and nitric oxide production by the phosphatidylinositol 3-kinase inhibitor LY294002. AB - Endothelial cells release nitric oxide (NO) acutely in response to increased laminar fluid shear stress, and the increase is correlated with enhanced phosphorylation of endothelial nitric-oxide synthase (eNOS). Phosphoamino acid analysis of eNOS from bovine aortic endothelial cells labeled with [(32)P]orthophosphate demonstrated that only phosphoserine was present in eNOS under both static and flow conditions. Fluid shear stress induced phosphate incorporation into two specific eNOS tryptic peptides as early as 30 s after initiation of flow. The flow-induced tryptic phosphopeptides were enriched, separated by capillary electrophoresis with intermittent voltage drops, also known as "peak parking," and analyzed by collision-induced dissociation in a tandem mass spectrometer. Two phosphopeptide sequences determined by tandem mass spectrometry, TQpSFSLQER and KLQTRPpSPGPPPAEQLLSQAR, were confirmed as the two flow-dependent phosphopeptides by co-migration with synthetic phosphopeptides. Because the sequence (RIR)TQpSFSLQER contains a consensus substrate site for protein kinase B (PKB or Akt), we demonstrated that LY294002, an inhibitor of the upstream activator of PKB, phosphatidylinositol 3-kinase, inhibited flow-induced eNOS phosphorylation by 97% and NO production by 68%. Finally, PKB phosphorylated eNOS in vitro at the same site phosphorylated in the cell and increased eNOS enzymatic activity by 15-20-fold. PMID- 10514498 TI - von Hippel-Lindau protein induces hypoxia-regulated arrest of tyrosine hydroxylase transcript elongation in pheochromocytoma cells. AB - Rat pheochromocytoma (PC12) cells were stably transfected with either wild type or mutated human von Hippel-Lindau tumor suppressor protein (hpVHL). These proteins have opposing effects on regulating expression of the gene encoding tyrosine hydroxylase (TH), the rate-limiting enzyme in catecholamine synthesis. Whereas wild type hpVHL represses levels of TH mRNA and protein 5-fold, a truncated pVHL mutant, pVHL(1-115), induces accumulation of TH mRNA and protein 3 fold. hpVHL-induced inhibition of TH gene expression does not involve either a decrease in TH mRNA stability or repression of TH promoter activity. However, repression results from inhibition of RNA elongation at a downstream region of the TH gene. This elongation pause is accompanied by hpVHL sequestration in the nuclear extracts of elongins B and C, regulatory components of the transcription elongation heterotrimer SIII (elongin A/B/C). Hypoxia, a physiological stimulus for TH gene expression, alleviates the elongation block. A truncated pVHL mutant, pVHL(1-115), stimulates TH gene expression by increasing the efficiency of TH transcript elongation. This is the first report showing pVHL-dependent regulation of specific transcript elongation in vivo, as well as dominant negative activity of pVHL mutants in pheochromocytoma cells. PMID- 10514500 TI - Role of the N-terminal region of the regulatory light chain in the dephosphorylation of myosin by myosin light chain phosphatase. AB - Myosin regulatory light chain (RLC) is phosphorylated at various sites at its N terminal region, and heterotrimeric myosin light chain phosphatase (MLCP) has been assigned as a physiological phosphatase that dephosphorylates myosin in vivo. Specificity of MLCP toward the various phosphorylation sites of RLC was studied, as well as the role of the N-terminal region of RLC in the dephosphorylation of myosin by MLCP. MLCP dephosphorylated phosphoserine 19, phosphothreonine 18, and phosphothreonine 9 efficiently with almost identical rates, whereas it failed to dephosphorylate phosphorylated serine 1/serine 2. Deletion of the N-terminal seven amino acid residues of RLC markedly decreased the dephosphorylation rate of phosphoserine 19 of RLC incorporated in the myosin molecule, whereas this deletion did not significantly affect the dephosphorylation rate of isolated RLC. On the other hand, deletion of only four N-terminal amino acid residues showed no effect on dephosphorylation of phosphoserine 19 of incorporated RLC. The inhibition of dephosphorylation by deletion of the seven N-terminal residues was also found with the catalytic subunit of MLCP. Phosphorylation at serine 1/serine 2 and threonine 9 did not influence the dephosphorylation rate of serine 19 and threonine 18 by MLCP. These results suggest that the N-terminal region of RLC plays an important role in substrate recognition of MLCP. PMID- 10514501 TI - Activation of the p38 mitogen-activated protein kinase by type I interferons. AB - The p38 mitogen-activated protein (Map) kinase plays a critical role in the generation of signals in response to stress stimuli, but its role in interferon (IFN) signaling and its potential regulatory role in the activation of Jak-signal transducer and activator of transcription (Stat) pathway are not known. In the present study, we provide evidence that the p38 Map kinase is rapidly phosphorylated and activated during treatment of cells with Type I interferons (IFNalpha and IFNbeta). Furthermore, the Type I IFN-dependent activation of p38 regulates induction of the catalytic domains of MapKap kinase-2 and MapKap kinase 3, strongly suggesting the existence of an IFNalpha signaling cascade activated downstream of the p38 kinase. The engagement of this pathway in interferon signaling plays a critical role in interferon-dependent transcriptional regulation, as evidenced by the fact that inhibition of p38 activation results in abrogation of interferon-dependent gene transcription via interferon-stimulated response elements. Interestingly, inhibition of the kinase activity of the p38 blocks IFNalpha-induced gene transcription without inhibiting DNA binding or tyrosine phosphorylation of Stat proteins, suggesting that the p38 pathway acts in cooperation with the Stat pathway. Thus, the p38 kinase signaling cascade is activated by the Type I interferon receptor and plays a critical role in interferon signaling and interferon-dependent transcriptional regulation. PMID- 10514499 TI - Mammal-specific, ERK-dependent, caldesmon phosphorylation in smooth muscle. Quantitation using novel anti-phosphopeptide antibodies. AB - Extracellular signal-regulated kinases (ERKs) phosphorylate the high molecular mass isoform of the actin-binding protein caldesmon (h-CaD) at two sites (Ser(759) and Ser(789)) during smooth muscle stimulation. To investigate the role of phosphorylation at these sites, antibodies were generated against phosphopeptides analogous to the sequences around Ser(759) and Ser(789). Affinity purified antibodies were phosho- and sequence-specific. The major site of phosphorylation in h-CaD in porcine carotid arterial muscle strips was at Ser(789); however, the amount of phosphate did not vary appreciably with either KCl or phorbol ester stimulation. Phosphorylation at Ser(759) of h-CaD was almost undetectable (<0.005 mol of phosphate/mol of protein). Moreover, phosphorylation of the low molecular mass isoform of the protein (l-CaD) at the site analogous to Ser(789) was greater in serum-stimulated cultured smooth muscle cells than in serum-starved cells. Serum-stimulated l-CaD phosphorylation was attenuated by the protein kinase inhibitor PD98059. These data 1) identify Ser(789) of h-CaD as the major site of ERK-dependent phosphorylation in carotid arteries; 2) show that the level of phosphorylation at Ser(789) is relatively constant following carotid arterial muscle stimulation, despite an increase in total protein phosphate content; and 3) suggest a functional role for ERK-dependent l-CaD phosphorylation in cell division. PMID- 10514502 TI - TEL is a sequence-specific transcriptional repressor. AB - TEL is a gene frequently involved in specific chromosomal translocations in human leukemia and sarcoma that encodes a member of the ETS family of transcriptional regulators. TEL is unusual among other ETS proteins by its ability to self associate in vivo, a property that is essential to the oncogenic activation of TEL-derived fusion proteins. We show here that TEL is a sequence-specific transcriptional repressor of ETS-binding site-driven transcription of model and natural promoters. Deletion of the oligomerization domain of TEL or its substitution by the homologous region of monomeric ETS1 impaired the ability of TEL to repress. In contrast, substitution of the oligomerization domain of TEL by unrelated oligomerization domains resulted in an active repressor, showing that the ability of TEL to repress depends on its ability to self-associate. The study of the properties of TEL fusions to the heterologous DNA binding domain of Gal4 identified two autonomous repression domains in TEL, distinct from its oligomerization domain, that are essential to the ability of TEL to repress ETS binding site-containing promoters. These results have implications for the normal function of TEL, its relation to other ETS proteins, and its role in leukemogenesis. PMID- 10514503 TI - Transforming growth factor-beta overrides the adhesion requirement for surface expression of alpha(5)beta(1) integrin in normal rat kidney fibroblasts. A necessary effect for induction of anchorage-independent growth. AB - We have previously shown that the expression of alpha(5)beta(1) integrin on the cell surface is dependent upon cell adhesion to the extracellular matrix, and we report here that transforming growth factor-beta (TGF-beta) overcomes this requirement in normal rat kidney (NRK) fibroblasts. Thus, suspended NRK cells treated with TGF-beta show levels of surface alpha(5)beta(1) integrin that are equivalent to those seen in adherent cells. Moreover, several experiments showed that this effect is necessary for the induction of anchorage-independent growth by TGF-beta. First, a kinetic analysis showed that surface expression of alpha(5)beta(1) integrin was restored in TGF-beta-treated NRK cells prior to the induction of anchorage-independent growth. Second, NRK cell mutants that have lost their TGF-beta requirement for surface expression of alpha(5)beta(1) integrin were anchorage-independent in the absence of TGF-beta. Third, an antisense oligonucleotide to the beta(1) integrin subunit or, fourth, stable expression of an alpha(5)-antisense cDNA blocked the ability of TGF-beta to stimulate anchorage-independent growth. Thus, TGF-beta overrides the adhesion requirement for surface expression of alpha(5)beta(1) integrin in NRK cells, and this effect is necessary for the induction of anchorage-independent growth. PMID- 10514504 TI - Lack of a C-terminal tail in the mammalian gonadotropin-releasing hormone receptor confers resistance to agonist-dependent phosphorylation and rapid desensitization. AB - The mammalian gonadotropin-releasing hormone receptor (GnRH-R) is, at present, the only G-protein-coupled receptor that activates phospholipase C and lacks a C terminal tail. We have previously demonstrated that this unique structural feature is associated with resistance to rapid desensitization of phosphoinositide signaling in COS-7 and HEK-293 cells (Heding, A., Vrecl, M., Bogerd, J., McGregor, A., Sellar, R., Taylor, P. L., and Eidne, K. A. (1998) J. Biol. Chem. 273, 11472-11477). Using receptors tagged with a nonapeptide of the influenza hemagglutinin protein to enable immunoprecipitation, we now demonstrate that the mammalian GnRH-R is not phosphorylated in an agonist-dependent manner. In contrast, the mammalian thyrotropin-releasing hormone receptor and the African catfish GnRH-R, both of which have a C-terminal tail, are phosphorylated in response to agonist challenge. Furthermore, chimeras of the mammalian GnRH-R with the C-terminal tail of either the mammalian thyrotropin-releasing hormone receptor or the catfish GnRH-R are also phosphorylated in an agonist-dependent manner. Only those receptors having C-terminal tails showed desensitization of phosphoinositide responses within 5-10 min of agonist challenge. We also show that the internalization of all these receptors when expressed transiently in COS 7 cells is similar. This dissociates receptor internalization from rapid desensitization and demonstrates that the lack of a C-terminal tail in the mammalian GnRH-R results in an inability of the receptor to undergo agonist dependent phosphorylation and that this results directly in a resistance to rapid desensitization. PMID- 10514505 TI - CrkL mediates Ras-dependent activation of the Raf/ERK pathway through the guanine nucleotide exchange factor C3G in hematopoietic cells stimulated with erythropoietin or interleukin-3. AB - CrkL is an SH2 and SH3 domain-containing adaptor protein implicated in pathogenesis of chronic myelogenous leukemia. Here, we demonstrate that overexpression of CrkL enhances the erythropoietin (Epo)- or interleukin (IL)-3 induced activation of Elk-1 and the c-fos gene promoter activity in 32D/EpoR-Wt cells. Moreover, the Epo-induced activation of ERK1 and ERK2 was augmented and prolonged in cells inducibly overexpressing CrkL. A moderate increase in Epo induced activation of JNK was also observed in cells overexpressing CrkL. Overexpression of C3G enhanced the Elk-1 activation synergistically with CrkL, while a C3G mutant lacking the guanine nucleotide exchange domain showed an inhibitory effect. Studies using a dominant negative Ha-Ras mutant demonstrated that the Elk-1 and ERK2 activation enhanced by CrkL and C3G was dependent on Ras. Consistent with this, the Epo-induced activation of Ras was augmented in cells inducibly overexpressing CrkL. Most importantly, a CrkL mutant defective in the SH2 or N-terminal SH3 domain showed an inhibitory effect on the Epo-induced activation of ERK2. These data indicate that the CrkL-C3G complex plays a role in Epo- or IL-3-induced, Ras-dependent activation of the Raf/ERK pathway leading to the activation of Elk-1 and the c-fos gene transcription. PMID- 10514506 TI - Increased myosin light chain phosphorylation is not required for growth factor stimulation of collagen matrix contraction. AB - Previous research suggested the possibility that contraction of floating collagen matrices by human fibroblasts required increased myosin light chain (MLC) phosphorylation. In the current studies, we show that increased MLC phosphorylation was neither necessary for platelet-derived growth factor (PDGF) dependent matrix contraction nor sufficient for lysophosphatidic acid (LPA) dependent contraction. In contrast, increased MLC phosphorylation did appear to be coupled to the formation of stress fibers by cells spreading in monolayer culture. Signal transduction pathways required for PDGF- and LPA-dependent matrix contraction involved phosphatidylinositol 3-kinase and the G(i) class of heterotrimeric G proteins, respectively. Our results indicate that PDGF- and LPA dependent contraction of floating collagen matrices can be uncoupled from an increase in MLC phosphorylation. PMID- 10514507 TI - Quantification of short term signaling by the epidermal growth factor receptor. AB - During the past decade, our knowledge of molecular mechanisms involved in growth factor signaling has proliferated almost explosively. However, the kinetics and control of information transfer through signaling networks remain poorly understood. This paper combines experimental kinetic analysis and computational modeling of the short term pattern of cellular responses to epidermal growth factor (EGF) in isolated hepatocytes. The experimental data show transient tyrosine phosphorylation of the EGF receptor (EGFR) and transient or sustained response patterns in multiple signaling proteins targeted by EGFR. Transient responses exhibit pronounced maxima, reached within 15-30 s of EGF stimulation and followed by a decline to relatively low (quasi-steady-state) levels. In contrast to earlier suggestions, we demonstrate that the experimentally observed transients can be accounted for without requiring receptor-mediated activation of specific tyrosine phosphatases, following EGF stimulation. The kinetic model predicts how the cellular response is controlled by the relative levels and activity states of signaling proteins and under what conditions activation patterns are transient or sustained. EGFR signaling patterns appear to be robust with respect to variations in many elemental rate constants within the range of experimentally measured values. On the other hand, we specify which changes in the kinetic scheme, rate constants, and total amounts of molecular factors involved are incompatible with the experimentally observed kinetics of signal transfer. Quantitation of signaling network responses to growth factors allows us to assess how cells process information controlling their growth and differentiation. PMID- 10514508 TI - Characterization of Osf1, an osteoblast-specific transcription factor binding to a critical cis-acting element in the mouse Osteocalcin promoters. AB - To elucidate the mechanisms of osteoblast-specific gene expression we are studying the regulation of osteocalcin, the most osteoblast-specific gene. Previous studies of OG2, one of the two mouse osteocalcin genes, identified two osteoblast-specific cis-acting elements, OSE1 and OSE2, the latter being the binding site of Cbfa1, the only osteoblast-specific transcription factor known to date. Here we show that OSE1 is a cis-acting element as important as OSE2 for the osteoblast-specific expression of OG2 in cell culture and transgenic mice. We also show that OSE1 is present in the promoter of several osteoblast-specific genes including Cbfa1 itself. These biological features demonstrate the importance of OSE1 and led us to further characterize this site and the factor binding to it, provisionally termed Osf1. We first defined the core OSE1 sequence, 5'-TTACATCA-3', which is necessary and sufficient for Osf1 binding to DNA. This sequence has no strong homology to any known transcription factor binding sites. As a first step in identifying Osf1, we performed an analytical purification of this protein using nuclear extracts from two different osteoblastic cell lines. We purified Osf1 to homogeneity through a five-step procedure including a renaturation experiment and found that its apparent molecular mass is 40 kDa. In conclusion, this study indicates the existence of multiple osteoblast-specific cis-acting elements of equal importance in controlling OG2 promoter activity and provides the first biochemical characterization of Osf1, a novel osteoblast-specific transcription factor. PMID- 10514509 TI - ATP and the core "alpha-Crystallin" domain of the small heat-shock protein alphaB crystallin. AB - Electrospray ionization mass spectrometry (ESI-LC/MS) of tryptic digests of human alphaB-crystallin in the presence and absence of ATP identified four residues located within the core "alpha-crystallin" domain, Lys(82), Lys(103), Arg(116), and Arg(123), that were shielded from the action of trypsin in the presence of ATP. In control experiments, chymotrypsin was used in place of trypsin. The chymotryptic fragments of human alphaB-crystallin produced in the presence and absence of ATP were analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Seven chymotryptic cleavage sites, Trp(60), Phe(61), Phe(75), Phe(84), Phe(113), Phe(118), and Tyr(122), located near or within the core alpha crystallin domain, were shielded from the action of chymotrypsin in the presence of ATP. Chemically similar analogs of ATP were less protective than ATP against proteolysis by trypsin or chymotrypsin. ATP had no effect on the enzymatic activity of trypsin and the K(m) for trypsin was 0.031 mM in the presence of ATP and 0.029 mM in the absence of ATP. The results demonstrated an ATP-dependent structural modification in the core alpha-crystallin domain conserved in nearly all identified small heat-shock proteins that act as molecular chaperones. PMID- 10514510 TI - Angiopoietin-1 and -2 coiled coil domains mediate distinct homo-oligomerization patterns, but fibrinogen-like domains mediate ligand activity. AB - Activity of endothelial Tie2 receptor tyrosine kinase is modulated by two naturally occurring, secreted ligands, angiopoietin-1 and -2, which have opposing effects on its phosphorylation. Receptor tyrosine kinase activation requires receptor dimerization/multimerization, which, for many receptors, is mediated by homo-oligomeric ligands binding to and bridging receptor molecules. We show here that angiopoietin-1 and -2 form distinct arrays of disulfide-linked homo oligomeric complexes. Their mobilities on nonreducing gels suggest that angiopoietin-2 exists predominantly as a homodimer but also forms higher order multimers. In contrast, angiopoietin-1 forms some homotrimers, but predominantly exists in higher order multimers. These two structurally related, 60% homologous ligands are predominantly composed of an amino-terminal coiled coil domain and a carboxyl-terminal fibrinogen-like domain. We show that their distinct oligomerization patterns are determined by their coiled coil domains and, furthermore, that their coiled coil domains, but not their fibrinogen-like domains, are sufficient to mediate formation of disulfide-linked homo-oligomers. In contrast, the differential effects of these ligands on endothelial Tie2 phosphorylation is mediated by their fibrinogen-like domains. We conclude from these studies that the coiled coil and fibrinogen-like domains of the angiopoietins have distinct functions with the coiled coil domain mediating ligand homo-oligomerization and the fibrinogen-like domain mediating ligand activity. PMID- 10514511 TI - TRAF family proteins interact with the common neurotrophin receptor and modulate apoptosis induction. AB - The common neurotrophin receptor, p75(NTR), has been shown to signal in the absence of Trk tyrosine kinase receptors, including induction of neural apoptosis and activation of NF-kappaB. However, the mechanisms by which p75(NTR) initiates these intracellular signal transduction pathways are unknown. Here we report interactions between p75(NTR) and the six members of TRAF (tumor necrosis factor receptor-associated factors) family proteins. The binding of different TRAF proteins to p75(NTR) was mapped to distinct regions in p75(NTR). Furthermore, TRAF4 interacted with dimeric p75(NTR), whereas TRAF2 interacted preferentially with monomeric p75(NTR). TRAF2-p75(NTR), TRAF4-p75(NTR), and TRAF6-p75(NTR) interactions modulated p75(NTR)-induced cell death and NF-kappaB activation with contrasting effects. Coexpression of TRAF2 with p75(NTR) enhanced cell death, whereas coexpression of TRAF6 was cytoprotective. Furthermore, overexpression of TRAF4 abrogated the ability of dimerization to prevent the induction of apoptosis normally mediated by monomeric p75(NTR). TRAF4 also inhibited the NF-kappaB response, whereas TRAF2 and TRAF6 enhanced p75(NTR)-induced NF-kappaB activation. These results demonstrate that TRAF family proteins interact with p75(NTR) and differentially modulate its NF-kappaB activation and cell death induction. PMID- 10514512 TI - Post-transcriptional regulation of H-ferritin mRNA. Identification of a pyrimidine-rich sequence in the 3'-untranslated region associated with message stability in human monocytic THP-1 cells. AB - We have previously demonstrated that phorbol myristate acetate (PMA) up-regulates H-ferritin gene expression in myeloid cells by stabilization of its message. In the present report, we showed that insertion of the 3'-untranslated region (3' UTR) of H-ferritin mRNA at the 3'-end of luciferase coding sequence significantly reduced the stability of luciferase mRNA in human monocytic THP-1 cells. However, the half-life of the chimeric transcript was markedly prolonged after PMA treatment. A cytosolic protein factor from THP-1 cells was found to specifically bind to H-ferritin 3'-UTR. PMA treatment of THP-1 cells resulted in the reduction of the RNA binding activity in a time-dependent manner. Deletion analysis and RNase T1 mapping revealed a pyrimidine-rich sequence within the 3'-UTR which interacts with the protein factor. Competition experiments with homoribopolymers further demonstrated the importance of uridines for the binding activity. Point mutations in uridines of the pyrimidine-rich sequence reduced the protein binding to 3'-UTR, while increasing the stability of the chimeric luciferase transcript. Together, these results demonstrate that the pyrimidine-rich sequence in the 3' UTR is involved in post-transcriptional regulation of H-ferritin gene expression in myeloid cells. PMID- 10514513 TI - Insulin-like growth factor-binding protein-3 binds fibrinogen and fibrin. AB - Following tissue injury, a fibrin network formed at the wound site serves as a scaffold supporting the early migration of stromal cells needed for wound healing. Growth factors such as insulin-like growth factor-I (IGF-I) concentrate in wounds to stimulate stromal cell function and proliferation. The ability of IGF-binding proteins (IGFBPs) such as IGFBP-3 to reduce the rate of IGF-I clearance from wounds suggests that IGFBP-3 might bind directly to fibrinogen/fibrin. Studies presented here show that IGFBP-3 does indeed bind to fibrinogen and fibrin immobilized on immunocapture plates, with K(d) values = 0.67 and 0.70 nM, respectively, and competitive binding studies suggest that the IGFBP-3 heparin binding domain may participate in this binding. IGF-I does not compete for IGFBP-3 binding; instead, IGF-I binds immobilized IGFBP-3.fibrinogen and IGFBP-3.fibrin complexes with affinity similar to that of IGF-I for the type I IGF receptor. In the presence of plasminogen, most IGFBP-3 binds directly to fibrinogen, although 35-40% of the IGFBP-3 binds to fibrinogen-bound plasminogen. IGFBP-3 also binds specifically to native fibrin clots, and addition of exogenous IGFBP-3 increases IGF-I binding. These studies suggest that IGF-I can concentrate at wound sites by binding to fibrin-immobilized IGFBP-3, and that the lower IGF affinity of fibrin-bound IGFBP-3 allows IGF-I release to type I IGF receptors of stromal cells migrating into the fibrin clot. PMID- 10514514 TI - The mitogen-activated protein (MAP) kinase p38 and its upstream activator MAP kinase kinase 6 are involved in the activation of signal transducer and activator of transcription by hyperosmolarity. AB - Environmental stress (e.g. aniso-osmolarity and UV light), hypoxia/reoxygenation, and reactive oxygen species activate intracellular signaling cascades such as the "stress-responsive" mitogen-activated protein kinases and nuclear factor kappaB. We have recently shown that the Janus tyrosine kinase/signal transducer and activator of transcription (Jak/STAT) pathway is ligand-independently activated by hyperosmotic shock. In the present study, we show that besides STAT1 also the tyrosine phosphatase SHP2 became tyrosine-phosphorylated upon hyperosmolarity. SB 202190 and SB 203580 (specific inhibitors of p38) inhibited both STAT activation and tyrosine phosphorylation of SHP2 induced by hyperosmotic stress. Overexpression of wild-type p38 mitogen-activated protein kinase and its upstream activator mitogen-activated protein kinase kinase 6 (MKK6) resulted in an enhanced STAT1 tyrosine phosphorylation upon osmotic shock. Accordingly, overexpression of dominant negative mutants of p38 and MKK6 largely decreased hyperosmotic STAT1 activation and tyrosine phosphorylation of SHP2. Furthermore, we provide evidence that a genistein-sensitive tyrosine kinase different from Jak1 is involved in stress-activation of STAT1 and tyrosine phosphorylation of SHP2. These results strongly suggest that hyperosmotic shock activates STAT1 and SHP2 via p38 and its upstream activator MKK6. PMID- 10514515 TI - At physiological expression levels the Kidd blood group/urea transporter protein is not a water channel. AB - The Kidd (JK) blood group locus encodes a urea transporter that is expressed on human red cells and on endothelial cells of the vasa recta in the kidney. Here, we report the identification in human erythroblasts of a novel cDNA, designated HUT11A, which encodes a protein identical to the previously reported erythroid HUT11 urea transporter, except for a Lys(44) --> Glu substitution and a Val-Gly dipeptide deletion after proline 227, which leads to a polypeptide of 389 residues versus 391 in HUT11. Genomic typing by polymerase chain reaction and transcript analysis by ribonuclease protection assay demonstrated that HUT11A encodes the true Kidd blood group/urea transporter protein, which carries only 2 Val-Gly motifs. Upon expression at high levels in Xenopus oocytes, the physiological Kidd/urea transporter HUT11A conferred a rapid transfer of urea (which was insensitive to p-chloromercuribenzene sulfonate or phloretin), a high water permeability, and a selective uptake of small solutes including amides and diols, but not glycerol and meso-erythritol. However, at plasma membrane expression levels close to the level observed in the red cell membrane, HUT11A mediated water transport and small solutes uptake were absent and the urea transport was poorly inhibited by p-chloromercuribenzene sulfonate, but strongly inhibited by phloretin. These findings show that, at physiological expression levels, the HUT11A transporter confers urea permeability but not water permeability, and that the observed water permeability is a feature of the red cell urea transporter when expressed at unphysiological high levels. PMID- 10514516 TI - Expression of a dominant negative SHP-2 in transgenic mice induces insulin resistance. AB - To elucidate the roles of SHP-2, we generated transgenic (Tg) mice expressing a dominant negative mutant lacking protein tyrosine phosphatase domain (DeltaPTP). On examining two lines of Tg mice identified by Southern blot, the transgene product was expressed in skeletal muscle, liver, and adipose tissues, and insulin induced association of insulin receptor substrate 1 with endogenous SHP-2 was inhibited, confirming that DeltaPTP has a dominant negative property. The intraperitoneal glucose loading test demonstrated an increase in blood glucose levels in Tg mice. Plasma insulin levels in Tg mice after 4 h fasting were 3 times greater with comparable blood glucose levels. To estimate insulin sensitivity by a constant glucose, insulin, and somatostatin infusion, steady state blood glucose levels were higher, suggesting the presence of insulin resistance. Furthermore, we observed the impairment of insulin-stimulated glucose uptake in muscle and adipocytes in the presence of physiological concentrations of insulin. Moreover, tyrosine phosphorylation of insulin receptor substrate-1 and stimulation of phosphatidylinositol 3-kinase and Akt kinase activities by insulin were attenuated in muscle and liver. These results indicate that the inhibition of endogenous SHP-2 function by the overexpression of a dominant negative mutant may lead to impaired insulin sensitivity of glucose metabolism, and thus SHP-2 may function to modulate insulin signaling in target tissues. PMID- 10514517 TI - Cloning and functional expression of a gene encoding a P1 type nucleoside transporter from Trypanosoma brucei. AB - Nucleoside transporters are likely to play a central role in the biochemistry of the parasite Trypanosoma brucei, since these protozoa are unable to synthesize purines de novo and must salvage them from their hosts. Furthermore, nucleoside transporters have been implicated in the uptake of antiparasitic and experimental drugs in these and other parasites. We have cloned the gene for a T. brucei nucleoside transporter, TbNT2, and shown that this permease is related in sequence to mammalian equilibrative nucleoside transporters. Expression of the TbNT2 gene in Xenopus oocytes reveals that the permease transports adenosine, inosine, and guanosine and hence has the substrate specificity of the P1 type nucleoside transporters that have been previously characterized by uptake assays in intact parasites. TbNT2 mRNA is expressed in bloodstream form (mammalian host stage) parasites but not in procyclic form (insect stage) parasites, indicating that the gene is developmentally regulated during the parasite life cycle. Genomic Southern blots suggest that there are multiple genes related in sequence to TbNT2, implying the existence of a family of nucleoside transporter genes in these parasites. PMID- 10514518 TI - An inducible nitric-oxide synthase (NOS)-associated protein inhibits NOS dimerization and activity. AB - A variety of transcriptional and post-transcriptional mechanisms regulate the expression of the inducible nitric-oxide synthase (iNOS, or NOS2). Although neurons and endothelial cells express proteins that interact with and inhibit neuronal NOS and endothelial NOS, macrophage proteins that inhibit NOS2 have not been identified. We show that murine macrophages express a 110-kDa protein that interacts with NOS2, which we call NOS-associated protein-110 kDa (NAP110). NAP110 directly interacts with the amino terminus of NOS2, and inhibits NOS catalytic activity by preventing formation of NOS2 homodimers. Expression of NAP110 may be a mechanism by which macrophages expressing NOS2 protect themselves from cytotoxic levels of nitric oxide. PMID- 10514519 TI - Neuronal Ca(2+) sensor 1. Characterization of the myristoylated protein, its cellular effects in permeabilized adrenal chromaffin cells, Ca(2+)-independent membrane association, and interaction with binding proteins, suggesting a role in rapid Ca(2+) signal transduction. AB - Overexpression of frequenin and its orthologue neuronal Ca(2+) sensor 1 (NCS-1) has been shown to increase evoked exocytosis in neurons and neuroendocrine cells. The site of action of NCS-1 and its biochemical targets that affect exocytosis are unknown. To allow further investigation of NCS-1 function, we have demonstrated that NCS-1 is a substrate for N-myristoyltransferase and generated recombinant myristoylated NCS-1. The bacterially expressed NCS-1 shows Ca(2+) induced conformational changes. The possibility that NCS-1 directly interacts with the exocytotic machinery to enhance exocytosis was tested using digitonin permeabilized chromaffin cells. Exogenous NCS-1 was retained in permeabilized cells but had no effect on Ca(2+)-dependent release of catecholamine. In addition, exogenous NCS-1 did not regulate cyclic nucleotide levels in this system. These data suggest that the effects of NCS-1 seen in intact cells are likely to be due to an action on the early steps of stimulus-secretion coupling or on Ca(2+) homeostasis. Myristoylated NCS-1 bound to membranes in the absence of Ca(2+) and endogenous NCS-1 was tightly membrane-associated. Using biotinylated NCS-1, a series of specific binding proteins were detected in cytosol, chromaffin granule membrane, and microsome fractions of adrenal medulla. These included proteins distinct from those detected by biotinylated calmodulin, demonstrating the presence of multiple specific Ca(2+)-independent and Ca(2+) dependent binding proteins as putative targets for NCS-1 action. A model for NCS 1 function, from these data, indicates a constitutive membrane association independent of Ca(2+). This differs from the Ca(2+) myristoyl switch model for the closely related recoverin and suggests a possible action in rapid Ca(2+) signal transduction in response to local Ca(2+) signals. PMID- 10514520 TI - CIS associates with the interleukin-2 receptor beta chain and inhibits interleukin-2-dependent signaling. AB - CIS is a cytokine-induced SH2-containing protein that was originally cloned as an interleukin (IL)-3-inducible gene. CIS is known to associate with the IL-3 receptor beta chain and erythropoietin receptor and to inhibit signaling mediated by IL-3 and erythropoietin. We now demonstrate that CIS also interacts with the IL-2 receptor beta chain (IL-2Rbeta). This interaction requires the A region of IL-2Rbeta (residues 313-382), which also mediates the association of IL-2Rbeta with Lck and Jak3. Correspondingly, CIS inhibits functions associated with both of these kinases: Lck-mediated phosphorylation of IL-2Rbeta and IL-2-mediated activation of Stat5. Thus, we demonstrate that CIS can negatively control at least two independent IL-2 signaling pathways. Although a functional SH2 binding domain of CIS was not required for its interaction with IL-2Rbeta in vitro, its phosphotyrosine binding capability was essential for the inhibitory action of CIS. On this basis, we have generated a mutant form of CIS protein with an altered SH2 domain that acts as a dominant negative and should prove useful in further understanding CIS action. PMID- 10514521 TI - The Raf-1/MEK/ERK pathway regulates the expression of the p21(Cip1/Waf1) gene in chondrocytes. AB - The gene encoding the cyclin-dependent kinase inhibitor p21(Cip1/Waf1) is up regulated in many differentiating cells, including maturing chondrocytes. Since strict control of chondrocyte proliferation is essential for proper bone formation and since p21 is likely involved in this control, we initiated analyses of the mechanisms regulating expression of p21 in chondrocytes. p21 expression and promoter activity was strongly increased during the differentiation of chondrogenic MCT cells. We have identified a 68-base pair fragment conferring transcriptional up-regulation of the p21 gene in chondrocytes. The activity of this fragment required active Raf-1 in MCT cells as well as in primary mouse chondrocytes. Inhibition of downstream factors of Raf-1 (MEK1/2, ERK1/2, and Ets2) also repressed the activity of the 68-base pair fragment in MCT cells. The chemical MEK1/2 inhibitor PD98059 reduced protein levels of p21 in MCTs and primary mouse chondrocytes. These data suggest that signaling through the Raf-1 pathway is necessary for the optimal expression of p21 in chondrocytes and may play an important role in the control of bone formation. PMID- 10514522 TI - The A-kinase anchor protein MAP2B and cAMP-dependent protein kinase are associated with class C L-type calcium channels in neurons. AB - Phosphorylation by cAMP-dependent protein kinase (PKA) increases the activity of class C L-type Ca(2+) channels which are clustered at postsynaptic sites and are important regulators of neuronal functions. We investigated a possible mechanism that could ensure rapid and efficient phosphorylation of these channels by PKA upon stimulation of cAMP-mediated signaling pathways. A kinase anchor proteins (AKAPs) bind to the regulatory R subunits of PKA and target the holoenzyme to defined subcellular compartments and substrates. Class C channels isolated from rat brain extracts by immunoprecipitation contain an endogenous kinase that phosphorylates kemptide, a classic PKA substrate peptide, and also the main phosphorylation site for PKA in the pore-forming alpha(1) subunit of the class C channel complex, serine 1928. The kinase activity is inhibited by the PKA inhibitory peptide PKI(5-24) and stimulated by cAMP. Physical association of the catalytic C subunit of PKA with the immunoisolated class C channel complex was confirmed by immunoblotting. A direct protein overlay binding assay performed with (32)P-labeled RIIbeta revealed a prominent AKAP with an M(r) of 280,000 in class C channel complexes. The protein was identified by immunoblotting as the microtubule-associated protein MAP2B, a well established AKAP. Class C channels did not contain tubulin and MAP2B association was not disrupted by dilution or addition of nocodazole, two treatments that cause dissociation of microtubules. In vitro experiments show that MAP2B can directly bind to the alpha(1) subunit of the class C channel. Our findings indicate that PKA is an integral part of neuronal class C L-type Ca(2+) channels and suggest that the AKAP MAP2B may mediate this interaction. Neither PKA nor MAP2B were detected in immunoprecipitates of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid type glutamate receptors or class B N-type Ca(2+) channels. Accordingly, MAP2B docked at class C Ca(2+) channels may be important for recruiting PKA to postsynaptic sites. PMID- 10514523 TI - Stimulatory function of paired immunoglobulin-like receptor-A in mast cell line by associating with subunits common to Fc receptors. AB - Paired Ig-like receptors (PIR) are polymorphic type I transmembrane proteins belonging to an Ig superfamily encoded by multiple isotypic genes. They are expressed on immune cells such as mast cells, macrophages, and B lymphocytes. Two subtypes of PIR have been classified according to the difference in the primary structure of the PIR transmembrane and cytoplasmic regions. These subtypes are designated as PIR-A and PIR-B. In this study, the transmembrane and cytoplasmic regions of the PIR-A subtype were shown to mediate activation signal events such as cytoplasmic calcium mobilization, protein tyrosine phosphorylations, and degranulation in rat mast cell line RBL-2H3. The association of the Fc receptor gamma and beta subunits with PIR-A was shown to be responsible for PIR-A function but not required for membrane expression of PIR-A on COS-7 cells. We further revealed the role of two charged amino acid residues in the transmembrane region, namely arginine and glutamic acid, in PIR-A function and its association with the above subunits. In contrast to the inhibitory nature of the PIR-B subtype, present findings reveal that PIR-A potentially acts as a stimulatory receptor in mast cells, suggesting a mechanism for regulation of mast cell functions by the PIR family. PMID- 10514524 TI - Mutations in a GTP-binding motif of eukaryotic elongation factor 1A reduce both translational fidelity and the requirement for nucleotide exchange. AB - A series of mutations in the highly conserved N(153)KMD(156)GTP-binding motif of the Saccharomyces cerevisiae translation elongation factor 1A (eEF1A) affect the GTP-dependent functions of the protein and increase misincorporation of amino acids in vitro. Two critical regulatory processes of translation elongation, guanine nucleotide exchange and translational fidelity, were analyzed in strains with the N153T, D156N, and N153T/D156E mutations. These strains are omnipotent suppressors of nonsense mutations, indicating reduced A site fidelity, which correlates with changes either in total translation rates in vivo or in GTPase activity in vitro. All three mutant proteins also show an increase in the K(m) for GTP. An in vivo system lacking the guanine nucleotide exchange factor eukaryotic elongation factor 1Balpha (eEF1Balpha) and supported for growth by excess eEF1A was used to show the two mutations with the highest K(m) for GTP restore most but not all growth defects found in these eEF1Balpha deficient strains to near wild type. An increase in K(m) alone, however, is not sufficient for suppression and may indicate eEF1Balpha performs additional functions. Additionally, eEF1A mutations that suppress the requirement for guanine nucleotide exchange may not effectively perform all the functions of eEF1A in vivo. PMID- 10514525 TI - The linker region between the helicase and primase domains of the bacteriophage T7 gene 4 protein is critical for hexamer formation. AB - The gene 4 protein of bacteriophage T7, a functional hexamer, comprises DNA helicase and primase activities. Both activities depend on the unidirectional movement of the protein along single-stranded DNA in a reaction coupled to the hydrolysis of dTTP. We have characterized dTTPase activity and hexamer formation for the full-length gene 4 protein (gp4) as well as for three carboxyl-terminal fragments starting at residues 219 (gp4-C219), 241 (gp4-C241), and 272 (gp4 C272). The region between residues 242 and 271, residing between the primase and helicase domains, is critical for oligomerization of the gene 4 protein. A functional TPase active site is dependent on oligomerization. During native gel electrophoresis, gp4, gp4-C219, and gp4-C241 migrate as oligomers, whereas gp4 C272 is monomeric. The steady-state k(cat) for dTTPase activity of gp4-C272 increases sharply with protein concentration, indicating that it forms oligomers only at high concentrations. gp4-C219 and gp4-C241 both form a stable complex with gp4, whereas gp4-C272 interacts only weakly with gp4. Measurements of surface plasmon resonance indicate that a monomer of T7 DNA polymerase binds to a dimer of gp4, gp4-C219, or gp4-C241 but to a monomer of gp4-C272. Like the homologous RecA and F(1)-ATPase proteins, the oligomerization domain of the gene 4 protein is adjacent to the amino terminus of the NTP-binding domain. PMID- 10514526 TI - Stimulation of DNA replication in Saccharomyces cerevisiae by a glutamine- and proline-rich transcriptional activation domain. AB - Glutamine-rich Sp1 and proline-rich CTF1, two extensively studied mammalian transcription factors, bind to origins of replication in DNA tumor viruses and stimulate viral DNA replication in mammalian cells. Here it is shown that, when tethered to a plasmid-borne cellular origin of replication, the activation domains of both proteins can enhance origin function in Saccharomyces cerevisiae. Hydrophobic patches in Sp1 and CTF1 that mediate transcriptional activation in higher eukaryotes are also important for activation of replication in yeast. However, only the activation domain of CTF1 can enhance initiation of replication from a chromosomally embedded origin. This correlates with the ability of CTF1 to alter the local chromatin structure around the chromosomal origin of replication. The CTF1-induced chromatin remodeling occurs at multiple stages of the cell cycle. These findings strongly suggest a high degree of conservation in the mechanisms used by various types of transcription factors to stimulate viral and cellular DNA replication in eukaryotes. PMID- 10514528 TI - Activation of the L voltage-sensitive calcium channel by mitogen-activated protein (MAP) kinase following exposure of neuronal cells to beta-amyloid. MAP kinase mediates beta-amyloid-induced neurodegeneration. AB - Neuronal degeneration in Alzheimer's disease (AD) has been variously attributed to increases in cytosolic calcium, reactive oxygen species, and phosphorylated forms of the microtubule-associated protein tau. beta-Amyloid (betaA), which accumulates extracellularly in AD brain, induces calcium influx in culture via the L voltage-sensitive calcium channel. Since this channel is normally activated by protein kinase A-mediated phosphorylation, we examined kinase activities recruited following betaA treatment of cortical neurons and SH-SY-5Y neuroblastoma. betaA increased channel phosphorylation; this increase was unaffected by the protein kinase A inhibitor H89 but was reduced by the mitogen activated protein (MAP) kinase inhibitor PD98059. Pharmacological and antisense oligonucleotide-mediated reduction of MAP kinase activity also reduced betaA induced accumulation of calcium, reactive oxygen species, phospho-tau immunoreactivity, and apoptosis. These findings indicate that MAP kinase mediates multiple aspects of betaA-induced neurotoxicity and indicates that calcium influx initiates neurodegeneration in AD. betaA increased MAP kinase-mediated phosphorylation of membrane-associated proteins and reduced phosphorylation of cytosolic proteins without increasing overall MAP kinase activity. Increasing MAP kinase activity with epidermal growth factor did not increase channel phosphorylation. These findings indicate that redirection, rather than increased activation, of MAP kinase activity mediates betaA-induced neurotoxicity. PMID- 10514527 TI - Targeted down-regulation of caveolin-3 is sufficient to inhibit myotube formation in differentiating C2C12 myoblasts. Transient activation of p38 mitogen-activated protein kinase is required for induction of caveolin-3 expression and subsequent myotube formation. AB - Caveolin-3 is the principal structural protein of caveolae membrane domains in striated muscle cells. Caveolin-3 mRNA and protein expression are dramatically induced during the differentiation of C2C12 skeletal myoblasts, coincident with myoblast fusion. In these myotubes, caveolin-3 localizes to the sarcolemma (muscle cell plasma membrane), where it associates with the dystrophin glycoprotein complex. However, it remains unknown what role caveolin-3 plays in myoblast differentiation and myotube formation. Here, we employ an antisense approach to derive stable C2C12 myoblasts that fail to express the caveolin-3 protein. We show that C2C12 cells harboring caveolin-3 antisense undergo differentiation and express normal amounts of four muscle-specific marker proteins. However, C2C12 cells harboring caveolin-3 antisense fail to undergo myoblast fusion and, therefore, do not form myotubes. Interestingly, treatment with specific p38 mitogen-activated protein kinase inhibitors blocks both myotube formation and caveolin-3 expression, but does not affect the expression of other muscle-specific proteins. In addition, we find that three human rhabdomyosarcoma cell lines do not express caveolin-3 and fail to undergo myoblast fusion. Taken together, these results support the idea that caveolin-3 expression is required for myoblast fusion and myotube formation, and suggest that p38 is an upstream regulator of caveolin-3 expression. PMID- 10514529 TI - The cytoplasmic domain of human FcgammaRIa alters the functional properties of the FcgammaRI.gamma-chain receptor complex. AB - The gamma/zeta-chain family of proteins mediate cell activation for multiple immunoglobulin receptors. However, the recognition that these receptors may have distinct biologic functions suggests that additional signaling elements may contribute to functional diversity. We hypothesized that the cytoplasmic domain (CY) of the ligand binding alpha-chain alters the biological properties of the receptor complex. Using macrophage FcgammaRIa as a model system, we created stable transfectants expressing a full-length or a CY deletion mutant of human FcgammaRIa. Both receptors functionally associate with the endogenous murine gamma-chain. However, we have established that the CY of FcgammaRIa directly contributes to the functional properties of the receptor complex. Deletion of the FcgammaRIa CY leads to slower kinetics of receptor-specific phagocytosis and endocytosis as well as lower total phagocytosis despite identical levels of receptor expression. Deletion of the CY also converts the phenotype of calcium independent FcgammaRIa-specific phagocytosis to a calcium-dependent phenotype. Finally, deletion of the CY abrogates FcgammaRIa-specific secretion of interleukin-6 but does not affect production of interleukin-1beta. These results demonstrate a functional role for the CY of FcgammaRIa and provide a general model for understanding how multiple receptors that utilize the gamma-chain can generate diversity in function. PMID- 10514530 TI - [Mapping the gene responsible for Smith-Fineman-Myers syndrome to Xq25]. AB - OBJECTIVE: To map and eventually identify the gene responsible for Smith-Fineman Myers syndrome. METHODS: The short tandem repeat markers(STRs) distributed on X chromosome at 8-10cM interval were used in the initial mapping to look for the candidate region for Smith-Fineman-Myers syndrome locus and the linked marker. The additional STRs flanking the linked marker were tested to confirm the candidate region and decide the interval of disease gene. RESULTS: Thirteen DNA samples from a Chinese family with Smith-Fineman-Myers syndrome were genotyped using 20 polymorphic STRs which cover the whole X chromosome. Of 20 STRs, DXS1001 on Xq25 suggested linkage and yielded a lod score of 3.01 at straight theta = 0 additional STRs flanking DXS1001 were tested. Fourteen polymorphic STRs out of 27 confirmed that Smith-Fineman-Myers syndrome locus is linked to several markers on Xq25. Haplotype analysis placed the disease locus within a 14.6cM interval bounded by DXS8064 and DXS8050. CONCLUSION: The gene responsible for Smith Fineman-Myers syndrome is mapped to a 14.6cM interval between DXS8064 and DXS8050 on Xq25. This result will be helpful for the identification of disease gene. PMID- 10514531 TI - [CAG trinucleotide mutation detection in patients with hereditary spinocerebellar ataxia]. AB - OBJECTIVE: To assess the frequency of the SCA1, SCA2,SCA3/MJD, SCA6, SCA7 and DRPLA CAG trinucleotide repeat expansions(CAG)n among individuals diagnosed with hereditary spinocerebellar ataxia(SCA) from Chinese families. METHODS: The SCA1, SCA2, SCA3/MJD, SCA6, SCA7 and DRPLA(CAG)n mutations were detected by polymerase chain reaction (PCR), denaturing polyacrylamide gel electrophoresis and silver staining technique in 167 patients with autosomal dominant SCA from 85 Chinese families and 37 sporadic SCA patients. RESULTS: Among 85 families, four families(4.70%) had seven SCA1 patients with the CAG repeat expanded to 53 to 62 repeats, five (5.88%) had twelve SCA2 patients with the CAG repeat expanded to 43 to 47 repeats, and 41 (48.23%) had 83 SCA3/MJD patients with the CAG repeat expanded to 68 to 83 repeats. Analysis of the mutation in these families showed a strong negative correlation between the size of the expanded CAG repeat and the age of disease onset. None of the SCA patients were positive for SCA6, SCA7, or DRPLA. Nor was any of the sporadic SCA patients positive for the CAG repeat expansion in the SCA1, SCA2, SCA3/MJD, SCA6, SCA7, or DRPLA gene. CONCLUSION: The frequency of SCA3/MJD is substantially higher than that of SCA1 and SCA2 in the autosomal dominant SCA from Chinese families. Chinese SCA3/MJD patients are non Portuguese. Clinical expressions of the various SCAs overlap one another and hence can make the phenotype-based diagnostic classification inaccurate in many instances. It is important for SCA clinical studies to make an SCA gene diagnosis and genotype analysis. PMID- 10514532 TI - [Genetic complementation group analysis of xeroderma pigmentosum patients in China]. AB - OBJECTIVE: To establish skin fibroblast cell strains derived from Chinese xeroderma pigmentosum(XP) patients and to analyze the genetic complementation group and distribution. METHODS: From four XP patients, skin fibroblast cell strains were derived and used for the complementation group analysis by autoradiography and cell-fusion technique. RESULTS: Of the four XP patients from apparently independent families, three were assigned to group C, and one belonged to group E. This is the first report on XP-E patient in China. CONCLUSION: Based on the data from the previous 7 cases and the present 4 cases, the distribution of complementation group in Chinese XP patients known at present is: nine XP-C, one XP-F and one XP-E. Although the number of cases is still limited, XP-C appears to be more frequent in China. This forms a marked contrast to Japanese XP patients, who are dominated by XP-A with infrequent presence of XP-C. PMID- 10514533 TI - [Analysis of fusion points in hybrid genes and gene deletion for congenital red green color vision defects]. AB - OBJECTIVE: To investigate gene deletion and the fusion points of hybrid genes in congenital red-green color vision defects. METHODS: Genomic DNA was collected from 11 protans, 19 deutans and 5 normal controls. Promoter and exons 2-5 of the red and green pigment genes in these subjects were analyzed by using PCR Heteroduplex-SSCP analysis. The origin and component of each individual gene were determined by comparison with the patterns of known sequence of the red and green visual pigment genes. RESULTS: Fourteen out of the 30 patients with red-green color vision defects were found to have hybrid gene. The fusion points of the hybrid gene were located in exon 1-intron 1(4 cases), introns 2-3(5 cases) and intron 4 (5 cases). CONCLUSION: The fusion point of a hybrid gene may occur in exon 1-intron 1 and intron 4 as well as in introns 2-3(including exon 3). PMID- 10514534 TI - [Mutation analysis of p16 gene in non-small cell lung carcinomas]. AB - OBJECTIVE: To investigate the role of p16 gene in the tumorigenesis of non-small cell lung carcinomas (NSCLC). METHODS: Homozygous deletion and mutation of exon 2 of p16 genes in 40 NSCLC tissues were analyzed using PCR and dsDNA direct sequencing technique. RESULTS: In 2 NSCLC tissues, homozygous deletions of p16 gene were detected; in 14 NSCLC tissues, 19 point mutations and a frameshift were detected. CONCLUSION: Point mutations of p16 gene were frequent and might play a role in the initiation and progression of NSCLC. Nt380 in this gene was a hot spot of mutation. PMID- 10514535 TI - [The genotype-based haplotype relative risk and transmission disequilibrium test analyses of familial febrile convulsions]. AB - OBJECTIVE: To confirm the linkage of familial febrile convulsions to the short arm of chromosome 6(6p) or the long arm of chromosome 8(8q). METHODS: The authors finished genotyping of Pst I locus on the coding region of heat shock protein (HSP) 70, 5'untranslated region of HSP70-1, 3' untranslated region of HSP70-2, D8S84 and D8S85. The data were processed by the genotype-based haplotype relative risk(GHRR) and transmission disequilibrium test(TDT) methods in PPAP. RESULTS: Some signs of association and disequilibrium between D8S85 and FC were shown by GHRR and TDT. CONCLUSION: A suspect linkage of familial febrile convulsions to the long arm of chromosome 8 has been proposed. PMID- 10514536 TI - [Relationships of angiotensinogen gene M235T variant with diabetic nephropathy in Chinese type 2 diabetes mellitus]. AB - OBJECTIVE: To investigate whether angiotensinogen(AGT) gene M235T variant is associated with type 2 diabetes mellitus without nephropathy (DM) and/or diabetic nephropathy (DN) in Chinese type 2 diabetes. METHODS: AGT genotypes of 84 cases with DM, 96 with DN and 98 controls were determined by polymerase chain reaction (PCR) amplification and restriction fragment length polymorphism analysis of the region of the variant. RESULTS: Increased frequencies of T allele (0.82) and TT genotype (0.70) were observed in 96 subjects with DN, compared with those observed in 98 control subjects (0.63 and 0. 43 respectively, P = 0.003, P = 0.0004). The odds ratio associated with TT genotype was 3.47 (95% CI 1.51-7.94, P = 0.0033) for DN in analysis adjusted for several DN risk factors. There was no difference in allele distribution between 84 DM patients and the controls. CONCLUSION: TT genotype of the AGT gene might be an independent risk factor of diabetic nephropathy in Chinese patients with type 2 diabetes mellitus. PMID- 10514537 TI - [Gene expression profiles in squamous esophageal cancer tissues and adjacent almost normal tissues]. AB - OBJECTIVE: To describe an esophageal cancer-specific expression profile and to identify genes that showed altered expression in squamous esophageal cancer tissues and their adjacent almost normal tissues. METHODS: The cDNA probes were synthesized from polyA(+)RNA of cancer and adjacent almost normal tissues and were differentially hybridized with two identical Atlas human cDNA expression arrays membranes containing 588 known genes. RESULTS: Autoradiographic analysis showed that of the 588 genes analyzed, 61 were found up-regulated in cancer, including cdc25B, Notch1, MMP, MET etc, and 22 down regulated in cancer, including cytokeratin4, BAD, IL-1 RECEPTOR ANTAGONIST, IL-6, etc. Expression levels of genes that associated with the regulation of cell proliferation, apoptosis, differentiation and metastasis altered most. CONCLUSION: The results for the first time provide an esophageal cancer-specific expression profile, showing that complex alterations of gene expression underlie the development of malignant phenotype of esophageal cancer cells. In addition, this line of research can lead to the identification of EC-specific genes which may be helpful for the development of diagnostic and prognostic biomarkers or therapeutic targets. The differential hybridization technique of Atlas Human cDNA expression array can be a useful method for describing the expression profiles of a tissue of cell interested. PMID- 10514538 TI - [A preliminary study on the genetic susceptibility of asthma in a Chinese population]. AB - OBJECTIVE: To study the asthma genetic susceptibility by way of ccandidate region so as to accumulate data on the related loci in Chinese population and determine whether genetic susceptibility to asthma is linked to the chromosome region. METHODS: One hundred ninety-two samples from 32 families were collected from Zhaoan county in Fujian province. Two Rsa I polymorphic sites within uncoded region of the high-affinity IgE receptor beta chain gene (Fc(epsilon) RI-beta) located on chromosome 11q13 were detected by PCR/Rsa I restriction endonuclease digestion. The polymorphic markers D5S436 and D5S393 within chromosome 5q31-33 were amplified by PCR incorporated with radioactive isotope. The asthma family samples were compared with unrelated samples randomly selected from general population. RESULTS: Genotypes containing A allele within intron 2 polymorphic site of Fc(epsilon) RI-beta gene were associated with asthma (P<0.05,OR = 2.039). Significant difference in the level of total serum IgE was noted among three genotypes of intron 2 polymorphic site of Fc(epsilon) RI-beta gene (P<0.05). The association between A allele and elevated total serum IgE was also significant (P<0.05, RR = 1.361). No relationship between exon 7 polymorphic site of Fc epsilonRI-beta gene and asthma was shown. By sib-pair analysis, significant evidence for linkage to asthma was observed with D5S436 (P<0.05) but not with D5S393. No significant linkage to total serum IgE was identified with both of these markers. CONCLUSION: The above-accumulated data suggested that both Fc epsilonRI-beta gene and chromosome 5q31-33 could be very attractive candidate gene and region for further studies in Chinese population, and to confirm these preliminary results, more markers within these regions merit testing in additional population samples. PMID- 10514539 TI - [Identification of mutations of SRD5A2 gene and SRY gene in patients with hypospadias]. AB - OBJECTIVE: To identify possible molecular mechanism underlined hypospadias and any relationship of the mutations of SRD5A2 gene and SRY gene to hypospadias. METHODS: Twenty-three blood samples from the patients with hypospadias were obtained from Aug.1996 to Jan. 1998. DNA was extracted from blood leukocytes. Exons 1 to 5 of the SRD5A2 gene and exon of the SRY gene were amplified by PCR. Mutation detection was performed using PCR-SSCP/silver staining and direct DNA sequencing. RESULTS: In 3 cases (named 5R2-China-1, 5R2-China-2, and 5R2-China 3), DNA sequencing revealed that a homozygous change from nucleotide G to A occurred in 5R2-China-1 and 2, leading to a substitution of glutamine to arginine in the codon 227(Arg 227 to Gln). In the third patient (5R2-China-3), DNA sequencing revealed two different heterozygous mutations(Arg 227 to Gln, Phe 186 to Leu) in exon 4 of the SRD5A2 gene. No mutation of SRY gene was found in all patients. CONCLUSION: The mutation of SRD5A2 gene affects the differentiation of the external genitalia and may play a role in the etiology of hypospadias. Since no mutation of SRY gene was found, this study might suggest that the mutation of SRY gene is not an important event in hypospadias. Codon 227 is a hotspot site of mutation within the gene. The mutation of codon 186(Phe 186Leu) represents a new form of SRD5A2 mutation. PMID- 10514540 TI - [Homozygous deletion of CDKN2/p16 gene in lung cancers]. AB - OBJECTIVE: To study the relationship between CDKN2/p16 gene homozygous deletion and lung cancer progression. METHODS: Improved multiplex PCR technique was applied to detect deletion of CDKN2/p16 gene exon1 and exon2 in 89 cases of lung cancers. RESULTS: Gene deletion rate was increased by the improved PCR technique. Exon 1 and exon 2 deletion rates were 19.1% (17/89) and 22.5% (20/89) respectively, with total rate of exon1 and/or exon2 deletion 25.8% (23/89). Deletion of CDKN2/p16 gene occurred in non-small cell lung carcinoma (NSCLC) and was related to metastasis and progressive stage. CONCLUSION: Abnormality of CDKN2/p16 gene is a genetic factor for NSCLC susceptibility, and may play a role to some extent in NSCLC malignant progression. PMID- 10514541 TI - [Linkage analysis of chromosome 5 and asthma in a Chinese population]. AB - OBJECTIVE: To investigate the linkage between asthma and 5q31-33 in a Chinese population. METHODS: The linkage between microsatellite markers in 5q and asthma and allergy was tested by lod score analysis. RESULTS: The linkage between asthma and 5q31-33 was not confirmed. CONCLUSION: The genes at 5q31-33 are not likely to contribute to inheritance of asthma in this Chinese population. PMID- 10514542 TI - [An extensive matrilineal nonsyndromic sensorineural deafness family and mtDNA 12SrRNA gene mutation]. AB - OBJECTIVE: To investigate the possible cause and molecular genetic mechanism of matrilineal nonsyndromic sensorineural deafness, the authors analyzed an extensive matrilineal nonsyndromic sensorineural deafness family. METHODS: PCR amplification of the nt1555 and nt7445 of the mitochondrial DNA, combined with PCR-SSCP, PCR-RFLP and sequence to analyze the family. RESULTS: The authors found a homoplasmic A to G transition at position 1555(A1555G) of the mitochondrial 12SrRNA gene from all the patients, and four matrilineal relatives of this family, but the mutation was not found in the normal spouses of the family and controls (100 normal persons). CONCLUSION: The A1555G mutation may be one of the major factors that cause deafness in this family. PMID- 10514543 TI - [Cloning and expression analyses of down-regulated cDNA C6-2A in human esophageal cancer]. AB - OBJECTIVE: To clone genes associated with the genesis of human esophageal cancer. METHODS: Identifying missing or low expressing cDNAs in human esophageal cancer tissues by mRNA differential display and examining its mRNA expression in 4 human cancer cell lines, 9 fetal tissues and other matched esophageal cancer tissues by Northern blot, dot blot and RT-PCR. RESULTS: One cDNA fragment named C6-2A, was cloned and sequenced. There was no identical sequence with C6-2A in BLASTN database; but in querying Genbank EST, the authors found that C6-2A was identical with ne27b03.s1NCI-CGAP-C03 humans sapiens cDNA clone IMAGE:898541 3' and zv30g07.rl Soares ovary tumor NbHOT homo sapiens cDNA clone 755196. 6/6 esophageal cancer tissues in Northern blot and 7/8 in dot blot did not or slightly express C6-2A. RT-PCR analysis showed that C6-2A was expressed much lower in 17/20 esophageal cancer tissues than adjacent microscopically normal mucosa, highly expressed in fetal esophageal mucosa, skin, cerebrum, placenta; moderately expressed in fetal stomach and liver, but not detected in fetal heart, small intestine and kidney. CONCLUSION: The high frequency of deletion of decreased expression of C6-2A in esophageal cell lines and human esophageal cancer tissues suggested that C6-2A might be involved in the carcinogenesis of esophagus. PMID- 10514544 TI - [A new explanation for shadow bands on denaturing PAGE for products of STRS--the difference between sense and antisense strands in electrophoresis]. AB - OBJECTIVE: To explore the mechanism of shadow bands on denaturing PAGE for PCR products of STR sequences and try to eliminate them. METHODS: PCR and asymmetric PCR were employed. The amplified fragments were separated on denaturing PAGE and visualized by silver stain. RESULTS: By asymmetric PCR, half of shadow bands of dinucleotide repeats were eliminated and there was only one band for each allele for tetranucleotide sequence. A different migration between the sense strand and antisense strand was identified. CONCLUSION: The difference between sense and antisense strands in electrophoresis is one of the causes in shadow bands, and these shadow bands can be eliminated by asymmetric PCR partly or totally. PMID- 10514545 TI - [Double-strand-primered polymerase chain reaction]. AB - OBJECTIVE: To develop a simple and reliable method for DNA amplification. METHODS: Double-strand-primers of fragile X mental retardation 1 gene (FMR1) were generated and used to amplify DNAs that were roughly extracted from human peripheral blood. RESULTS: All products from 15 DNA samples by double-strand primered polymerase chain reaction (DSP-PCR) were more specific than those amplified by routine PCR. CONCLUSION: The double-strand-primers reported here may increase the specificity of PCR amplification. PMID- 10514546 TI - [Detection of methylation status of tumor suppressor gene p16 by methylation specific polymerase chain reaction]. AB - OBJECTIVE: To introduce a new method for studies on methylation status of CpG island: methylation specific polymerase chain reaction (MSP) with some modifications. METHODS: Target DNA was modified by sodium bisulfite, converting all unmethylated, but not methylated, cytodines to uracil, and subsequent amplification with primers specific for methylated versus unmethylated DNA. The status of 5'CpG island of tumor suppressor gene p16 in maxilliofacial squamous cell carcinoma (MSCC) was analyzed by modified MSP assay. RESULTS: To get single strand DNA, the authors denatured the target DNA by heating instead of alkaline treatment. DNA purification kit was also replaced by dialysis. A cheaper restriction enzyme Taq I was used in place of BstU I to distinguish methylated fragment from unmethylated fragment. The 5'CpG island of p16 gene was methylated in some MSCC tumors. CONCLUSION: MSP is a simple, sensitive, and specific method for detecting the methylation status of any CpG-rich region and the modifications make it more simple to perform. PMID- 10514547 TI - Origin of selective inhibition of mitochondrial complex I by pyridinium-type inhibitor MP-24. AB - Positively charged pyridiniums are unique inhibitors to probe the structural and functional properties of the ubiquinone reduction site of bovine heart mitochondrial complex I. In this study, we synthesized a series of neutral as well as pyridinium analogues of MP-24 (N-methyl-4-[2-methyl-2-(p-tert butylbenzyl)propyl]pyridinium), a selective inhibitor of one of the two proposed binding sites of these pyridinium-type inhibitors of complex I (H. Miyoshi et al., J. Biol. Chem. 273 (1998) 17368-17374), to elucidate the origin of its selectivity. Inhibitory potencies of all neutral and pyridinium analogues with tetraphenylboron (TPB(-)), which forms an ion-pair with pyridiniums, were comparable, although the degrees of selective inhibition by pyridiniums without TPB(-) were entirely different. In contrast to MP-24, the dose-response curves of nonselective pyridiniums and all neutral analogues were not affected by incubation conditions. These results strongly suggested that the process of the inhibitor passage to the binding sites is responsible for the selective inhibition. PMID- 10514548 TI - H(2)O(2) detection from intact mitochondria as a measure for one-electron reduction of dioxygen requires a non-invasive assay system. AB - Evaluation of the existence of superoxide radicals (O*-(2)), the site of generation and conditions required for one-e(-) transfer to oxygen from biological redox systems is a prerequisite for the understanding of the deregulation of O(2) homeostasis leading to oxidative stress. Mitochondria are increasingly considered the major O*-(2) source in a great variety of diseases and the aging process. Contradictory reports on mitochondrial O*-(2) release prompted us to critically investigate frequently used O*-(2) detection methods for their suitability. Due to the impermeability of the external mitochondrial membrane for most constituents of O*-(2) detection systems we decided to follow the stable dismutation product H(2)O(2). This metabolite was earlier shown to readily permeate into the cytosol. With the exception of tetramethylbenzidine none of the chemical reactants indicating the presence of H(2)O(2) by horseradish peroxidase-catalyzed absorbance change were suited due to solubility problems or low extinction coefficients. Tetramethylbenzidine-dependent H(2)O(2) detection was counteracted by rereduction of the dye through e(-) carriers of the respiratory chain. Although the fluorescent dyes scopoletin and homovanillic acid were found to be suited for the detection of mitochondrial H(2)O(2) release, fluorescence change was strongly affected by mitochondrial protein constituents. The present study has resolved this problem by separating the detection system from H(2)O(2)-producing mitochondria. PMID- 10514549 TI - Evidence for the stabilization of NADPH relative to NADP(+) on the dIII components of proton-translocating transhydrogenases from Homo sapiens and from Rhodospirillum rubrum by measurement of tryptophan fluorescence. AB - A unique Trp residue in the recombinant dIII component of transhydrogenase from human heart mitochondria (hsdIII), and an equivalent Trp engineered into the dIII component of Rhodospirillum rubrum transhydrogenase (rrdIII.D155W), are more fluorescent when NADP(+) is bound to the proteins, than when NADPH is bound. We have used this to determine the occupancy of the binding site during transhydrogenation reactions catalysed by mixtures of recombinant dI from the R. rubrum enzyme and either hsdIII or rrdIII.D155W. The standard redox potential of NADP(+)/NADPH bound to the dIII proteins is some 60-70 mV higher than that in free solution. This results in favoured reduction of NADP(+) by NADH at the catalytic site, and supports the view that changes in affinity at the nucleotide binding site of dIII are central to the mechanism by which transhydrogenase is coupled to proton translocation across the membrane. PMID- 10514550 TI - Extinction coefficients and midpoint potentials of cytochrome c(6) from the cyanobacteria Arthrospira maxima, Microcystis aeruginosa, and Synechocystis 6803. AB - Cytochrome c(6) is a soluble heme protein that serves as a photosynthetic electron transport component in cyanobacteria and algae, carrying electrons from the cytochrome bf complex to photosystem I. The rapid accumulation of cytochrome c(6) sequence data from a wide range of species, combined with significant advances in determining high resolution three-dimensional structures, provides a powerful database for investigating the relationship between structure and function. The fact that the gene encoding cytochrome c(6) can be readily modified in a number of species adds to the usefulness of cytochrome c(6) as a tool for comparative analysis. Efforts to relate cytochrome c(6) sequence information to structure, and structural information to function depend on knowledge of the physical and thermodynamic properties of the cytochrome from different species. To this end we have determined the optical extinction coefficient, the oxidation/reduction midpoint potential, and the pH dependence of the midpoint potential of cytochrome c(6) isolated from three cyanobacteria, Arthrospira maxima, Microcystis aeruginosa, and Synechocystis 6803. PMID- 10514551 TI - In vitro expression of long and short ovine prolactin receptors: activation of Jak2/STAT5 pathway is not sufficient to account for prolactin signal transduction to the ovine beta-lactoglobulin gene promoter. AB - The recent finding that sheep had long (l-oPRLR) and short (s-oPRLR) prolactin receptors provided new tools to further explore prolactin signaling to target genes. Here we used CHO cells transfected with l-oPRLR or s-oPRLR cDNAs to compare the activation of known key steps of prolactin signaling by the two receptors. We found that prolactin stimulated l-oPRLR tyrosine phosphorylation, although it lacked the last tyrosine residue found in other long prolactin receptors. In addition, l-oPRLR and s-oPRLR both responded to prolactin stimulation by (1) Janus kinase 2 (Jak2) tyrosine phosphorylation, (2) DNA binding activation of signal transducer and activator of transcription 5 (STAT5), (3) stimulation of transcription from a promoter made of six repeats of STAT5 responsive sequence. However, although it contains STAT5-binding consensus sequences, the ovine beta-lactoglobulin promoter (-4000 to +40) was transactivated by l-oPRLR, but not by s-oPRLR. Taken together, our results indicate that activation of Jak2/STAT5 pathway alone is not sufficient to account for prolactin-induced transcription of this milk protein gene, and that sequences of its promoter, other than STAT5-specific sequences, account for the opposite transcriptional activation capabilities of l-oPRLR and s-oPRLR. PMID- 10514552 TI - The upregulation of messenger ribonucleic acids during 17alpha, 20beta-dihydroxy 4-pregnen-3-one-induced ovulation in the perch ovary. AB - While progestins appear to be involved in the local ovarian regulation of vertebrate ovulation, their specific role is unclear. In yellow perch (Perca flavescens) the progestin, 17alpha, 20beta-dihydroxy-4-pregnen-3-one (17,20-P), stimulates ovulation in vitro and this induction requires gene activation. Therefore, the perch model was used to isolate progestin-upregulated mRNAs. Perch ovaries were incubated for 32 h with or without 17,20-P (0.1 microg/ml). Messenger ribonucleic acids were isolated from the tissue and used for differential display PCR (DDPCR). From DDPCR, 5 bands were eventually obtained that were verified by Northern analysis to be consistently upregulated by 17,20-P at 32 h. Using these bands, full-length cDNAs were obtained by library screening and completely sequenced. Based on similarity to known sequences, four of the cDNAs presumably encode for perch forms of (1) neprilysin (PNEP-1; 63% identical); (2) a lysyl oxidase-type protein (PLO-2; 43.2% identical); (3) calmodulin (PCAL-1; 100% identical); and (4) a microtubule aggregate-like protein (PMAP-1; 29.6% identical). The fifth cDNA obtained from DDPCR most likely encodes for an egg protein and will be reported separately. Each of the cDNAs was used to probe Northern blots of ovarian mRNA taken at 0, 12, 24, 32 and 42 h of incubation with 17,20-P. This temporal Northern analysis verified that all four were upregulated by 32 h. In addition, PNEP and PMAP transcripts began to increase by 12 h, while PCAL and PLO transcripts remained elevated through 42 h. On Northern blots of RNA from other perch tissues, calmodulin was found in all tissues, PLO mRNA was ovarian specific, and PMAP mRNA was also present in the gills and liver. Multiple transcripts were observed for PNEP, but the ovarian form induced by 17,20-P was only found in high abundance in the heart. To our knowledge, this is the first report that specifically characterizes progestin upregulated mRNAs in the vertebrate ovary at ovulation. PMID- 10514553 TI - Oestrogens regulate pituitary alpha2,3-sialyltransferase messenger ribonucleic acid levels in the female rat. AB - Follicle-stimulating hormone (FSH) is synthesized by the anterior pituitary gland in multiple molecular forms. Increased acidic/sialylated FSH charge isoforms are associated with conditions characterized by a low oestrogen output. In the present study, we analysed the dynamics of the changes in mRNA levels of the enzyme Galbeta1,3[4]GlcNAc alpha2,3-sialyltransferase (2,3-STase) (one of the enzymes that incorporate sialic acid residues into the FSH molecule) in intact and ovariectomized rats. The anterior pituitaries of 4-day regularly cyclic adult female Wistar rats were obtained at 1000 h on the days of pro-oestrus (P), oestrus (O), dioestrus 1 (D1) and dioestrus 2 (D2), at 0200 h, 1400 h, 1800 h and 2200 h on D1, at 1800 h on day of O and at 1000 h after 7, 14, 21, 28 and 45 days of oophorectomy performed on the morning of P. Total RNA was isolated from each gland and the 2,3-STase levels were measured by Northern blot hybridization analysis employing a 346-base pair cDNA probe encoding for a non-conserved amino acid sequence of the catalytic domain of the enzyme. Maximal levels of the enzyme mRNA were detected at 1000 h on D1; thereafter, they progressively decreased by 60% during the ensuing 24 h, reaching the lowest concentration values (26% of the maximally observed level on D1) at 1000 h on day of P and remaining unchanged during the morning of O. Administration of the potent oestradiol receptor antagonist ICI 182,780 at 1000 h on D1 completely reverted the time-dependent decrease in 2,3-STase mRNA levels observed during the afternoon of D1, whereas oestradiol benzoate administered at 1000 h on day of O significantly reduced the enzyme mRNA levels (to 21% of the levels detected in vehicle-treated controls). In ovariectomized rats, the alpha2,3-STase mRNA progressively increased from day 21 to day 45 post castration. Administration of oestradiol benzoate on day 28 after oophorectomy significantly reduced the 2,3-STase mRNA levels (to 36% of the levels detected in vehicle-injected controls); ICI 182,780 partially counteracted this oestradiol-mediated effect. The dynamics of these changes in 2,3-STase mRNA levels partially correlated with changes in the relative abundance of the FSH charge isoforms separated by preparative chromatofocusing of anterior pituitary extracts, particularly in glands obtained during the morning of P and O. These data demonstrate for the first time that pituitary 2,3-STase is a hormonally regulated enzyme and that the changes in transcription and/or stability of its mRNA may be involved, in part, in the post-translational processing of the FSH molecule during certain physiological conditions. PMID- 10514554 TI - Suppression of P450 aromatase gene expression in sex-reversed males produced by rearing genetically female larvae at a high water temperature during a period of sex differentiation in the Japanese flounder (Paralichthys olivaceus). AB - The phenotypic sex of many teleost fishes including flounders can be experimentally altered by treating embryos or larvae with varied temperatures or sex-steroid hormones. To analyse the sex determination mechanism, especially the role of cytochrome P450 aromatase (P450arom), an enzyme that catalyses the conversion of androgens to estrogens, in temperature-dependent gonadal sex differentiation in the Japanese flounder, we generated two populations of larvae, both having XX (genetic females) but each growing up to display all phenotypic females or males, by rearing the larvae at normal (18 degrees C) or high (27 degrees C) water temperatures from days 30 to 100 after hatching respectively. The larvae (XX) were produced artificially by mating normal females (XX) with gynogenetic diploid males (XX) which had been sex-reversed to phenotypic males by 17alpha-methyltestosterone. To study the role of P450arom in sex determination in the flounder, we first isolated a P450arom cDNA containing the complete open reading frame from the ovary. RT-PCR showed that P450arom mRNA was highly expressed in the ovary and spleen but weakly in the testis and brain. Semi quantitative analyses of P450arom mRNA in gonads during sex differentiation showed that there was no difference in the levels of P450arom mRNA between the female and male groups when the gonad was sexually indifferent (day 50 after hatching). However, after the initiation of sex differentiation (day 60), the mRNA levels increased rapidly in the female group, whereas they decreased slightly in the male group. Similarly, estradiol-17beta levels rose remarkably in the female group, yet remained constant in the male group. These results suggest that induction of sex reversal of genetically female larvae to phenotypic males by rearing them at a high water temperature caused a suppression of P450arom gene expression. Furthermore, we suggest that the maintenance of P450arom mRNA at very low levels is a prerequisite for testicular differentiation, while the increased levels are indispensable for ovarian differentiation. PMID- 10514555 TI - Blue gourami (Trichogaster trichopterus) gonadotropic beta subunits (I and II) cDNA sequences and expression during oogenesis. AB - We have cloned two cDNAs from the pituitary gland of blue gourami (Trichogaster trichopterus), coding for the beta subunits of the gonadotropin hormones GtH-I and GtH-II. The two cDNAs were sequenced and subjected to sequence analysis. We have found that the deduced amino acid sequences of both cDNAs were most similar to their striped bass counterparts. The beta GtH-I subunits of blue gourami and striped bass shared 73% of their residues, and the beta GtH-II subunits 84%. The cloning of the cDNAs of beta GtH-I and beta GtH-II has enabled us to measure the expression of their respective mRNAs in the pituitaries of female blue gourami at different stages of the reproductive cycle. The highest levels of beta GtH-I and beta GtH-II mRNA were found in specimens classified as high vitellogenic and in females that were at the final stages of oocyte maturation. PMID- 10514556 TI - Changes in the levels of mRNAs for GH/prolactin/somatolactin family and Pit-1/GHF 1 in the pituitaries of pre-spawning chum salmon. AB - Changes in the levels of pituitary mRNAs encoding GH, prolactin (PRL) and somatolactin (SL) were determined in pre-spawning chum salmon (Oncorhynchus keta) caught at a few key points along their homing pathway in 1994 and 1995. Furthermore, we analyzed relationships between expression of pituitary-specific POU homeodomain transcription factor (Pit-1/GHF-1) and GH/PRL/SL family genes. In 1994, seawater (SW) fish and matured fresh-water (FW) fish were sequentially captured at two points along their homing pathway, the coast and the hatchery. In addition to these two points, maturing FW fish were captured at the intermediate of the two points in 1995. The levels of hormonal mRNAs were determined by a quantitative dot blot analysis using single-stranded sense DNA as the standard. Relative levels of Pit-1/GHF-1 mRNAs were estimated by Northern blot analysis. In 1994, the levels of GH/PRL/SL family mRNAs except for PRL mRNA in the male FW fish were 1.8-4 times higher than those in the SW fish. In 1995, the level of PRL mRNA was somewhat sharply elevated in the maturing FW fish soon after entry into the FW environment, while that of SL mRNA was gradually increased during upstream migration from the coast to the hatchery. The levels of 3 kb Pit-1/GHF-1 mRNA in the FW fish were higher than those in the SW fish in both 1994 and 1995. The present results indicate that expression of genes for the GH/PRL/SL family and Pit-1/GHF-1 is coincidentally enhanced in homing chum salmon. Moreover, the present study suggests that expression of the SL gene is elevated with sexual maturation, whereas that of PRL gene is elevated with osmotic change during the final stages of spawning migration. PMID- 10514557 TI - Clusterin is expressed in the anterior and intermediate lobes, but not in the posterior pituitary of sheep. AB - We have examined the expression of the ovine clusterin gene in the sheep pituitary gland, with the aim of determining its site of synthesis in this tissue. Northern blotting analysis of extracted polyadenylated RNA, using a (32)P labelled rat clusterin cDNA probe, detected the greatest amounts of clusterin mRNA in the anterior part of dissected pituitary glands. In situ hybridisation studies showed clusterin mRNA in anterior and intermediate pituitary cells, with lower amounts in vascular endothelium and posterior pituicytes. Clusterin protein, detected by immunohistochemistry, was observed in some single secretory cells, within the capillary lumen and in cells around capillaries in the anterior and intermediate lobes, but no immunoreactivity was observed in posterior pituitary tissue. The pattern of clusterin expression in anterior and intermediate pituitary cells suggests possible roles for the protein in secretory cell turnover and/or hormone secretion or lipid uptake. Clusterin does not appear to be involved in ovine posterior pituitary hormone neurosecretion. PMID- 10514558 TI - Testosterone effect on growth and growth mediators of the GH-IGF-I axis in the liver and epiphyseal growth plate of juvenile rats. AB - Several studies have suggested that testosterone may have a direct, GH independent effect on growth. In order to assess possible mechanism(s) whereby testosterone exerts its growth-promoting effect, we evaluated its effect on growth mediators of the GH-IGF-I axis, in both the liver and the epiphyseal growth plate (EGP). Testosterone was administered to peripubertal rats and the responses of mRNA of GH receptor, IGF-I, IGF-I receptor and IGF-binding proteins 1 and -3 (IGFBP-1 and IGFBP-3) as well as circulating IGF-I were evaluated in two time-related models: over 12 h after a single injection (short-term study) and 10 days after continuous administration (long-term study). Rats in the short-term study were castrated and were killed 1, 4, 6 and 12 h post injection. Rats in the long-term study were divided into two groups: castrated vs castrated and hypophysectomized, in order to assess the effect of testosterone in the presence and absence of GH. mRNA levels were determined by RNase protection assay, and serum IGF-I by RIA. Testosterone enhanced weight gain in the rats treated for 10 days, a change that was similar in the presence or absence of GH. This effect was relatively small, however, by comparison with the total weight gained without testosterone. Testosterone had no effect on hepatic IGF-I mRNA abundance but induced a reduction in circulating IGF-I levels, in both the short- and long-term study. Testosterone had no effect on hepatic GH receptor and IGFBP-3 mRNA levels but resulted in a transient, short-term elevation in IGFBP-1 mRNA levels that was maximal 4 h post injection. In the EGP, neither testosterone administration nor hypophysectomy had any effect on IGF-I and IGF-I receptor mRNA levels. However, testosterone increased GH receptor mRNA abundance after 10 days of continuous administration in hypophysectomized rats only. These data suggest that the effect of testosterone on growth (as assessed by weight gain) is small and is not mediated by changes in hepatic gene expression of IGF-I, IGF-I receptor, IGFBP-1, IGFBP-3 or circulating IGF-I. At the EGP, the testosterone effect on linear growth is not mediated through changes in mRNA abundance of IGF-I and IGF-I receptor. The small but significant elevation of GH receptor mRNA levels in hypophysectomized rats may suggest a testosterone-mediated augmentation of a GH effect at the target organ. PMID- 10514559 TI - Resolution and identification of Gq/G11alpha and Gialpha/Goalpha proteins in brown adipose tissue: effect of cold acclimation. AB - Levels of guanine nucleotide-binding proteins G(q)alpha and G(11)alpha, which produce receptor regulation of phospholipase C, were measured immunologically in purified plasma membrane fractions of hamster brown adipose tissue (BAT). This was achieved by immunoblotting with antisera (CQ series) that identify these two polypeptides equally, following separation of the plasma membranes using SDS-PAGE in the presence of 6 M urea, i.e. conditions that can resolve G(q)alpha and G(11)alpha. The ratio of levels of G(q)alpha to G(11)alpha was 1:1. A similar approach was used for resolution and identification of G(o)1alpha and G(o)2alpha, the latter representing the prevailing form of G(o)alpha proteins in this tissue. Although clearly recognized in brain microsomes, which were used as positive controls, no detected levels of G(o)*alpha protein were noted. Using specific anti-peptide antibodies directed against the carboxy-terminal decapeptide of G(i)3alpha, this G protein was also found to be expressed in BAT tissue. Cold acclimation resulted in reduction of the plasma membrane levels of all these Galpha proteins. PMID- 10514560 TI - Creation of a fully active, cytosolic form of human type I 3beta-hydroxysteroid dehydrogenase/isomerase by the deletion of a membrane-spanning domain. AB - Human 3beta-hydroxysteroid dehydrogenase/steroid Delta(5)-Delta(4)-isomerase (3beta-HSD/isomerase) is a bifunctional, single enzyme protein that is membrane bound in the endoplasmic reticulum (microsomes) and mitochondria of cells in the placenta (type I) and in the adrenals and gonads (type II). Two membrane-binding domains (residues 72-89 and 283-310) have been predicted by analyses of hydrophobicity in the type I and II isoenzymes (90% regional homology). These putative membrane domains were deleted in the cDNA by PCR-based mutagenesis, and the two mutant enzymes were expressed by baculovirus in insect Sf9 cells. Differential centrifugation of the Sf9 cell homogenate containing the 283-310 deletion mutant revealed that 94% of the 3beta-HSD and isomerase activities were in the cell cytosol, 6% of the activities were in the microsomes, and no activity was in the mitochondria. This is the opposite of the subcellular distribution of the wild-type enzyme with 94% of the activities in the microsomes and mitochondria and only 6% activity in the cytosol. The organelle distribution of the 72-89 deletion mutant lies between these two extremes with 72% of the enzyme activity in the cytosol and 28% in the microsomes/mitochondria. The integrity of the subcellular organelle preparations was confirmed by electron microscopy. Western immunoblots confirmed the presence of the 283-310 deletion mutant enzyme and the absence of the wild-type enzyme in the insect cell cytosol. The unpurified, cytosolic 383-310 deletion mutant exhibited 3beta-HSD (22 nmol/min per mg) and isomerase (33 nmol/min per mg) specific activities that were comparable with those of the membrane-bound, wild-type enzyme. The isomerase reaction of the cytosolic 283-311 deletion mutant requires activation by NADH just like the isomerase of the microsomal or mitochondrial wild-type enzyme. In contrast, the 72-89 deletion mutant had low 3beta-HSD and isomerase specific activities that were only 12% of the wild-type levels. This innovative study identifies the 283-310 region as the critical membrane domain of 3beta HSD/isomerase that can be deleted without compromising enzyme function. The shorter 72-89 region is also a membrane domain, but deletion of this NH(2) terminal region markedly diminishes the enzyme activities. Purification of the active, cytosolic 283-310 deletion mutant will produce a valuable tool for crystallographic studies that may ultimately determine the tertiary/quaternary structure of this key steroidogenic enzyme. PMID- 10514562 TI - The proteolytic system of the yeast Kluyveromyces lactis. AB - Major proteolytic activities were characterized in the yeast K. lactis NRRL 1118, grown in chemostat cultures. This yeast expressed proteolytic activities similar to those found in S. cerevisiae. This fact was particularly evident in the case of proteases such as PrA, PrB and CpY with regard to substrate specificity, activation at pH 5. 0 and inhibition patterns. The presence of a CpS activity could not be detected in either fresh or activated cell-free extracts by using the dipeptide N-Cbz-Gly-Leu, even in the presence of Zn(+2). On the other hand, K. lactis exhibits at least two major intracellular Ap activities different from those reported in other yeasts, and these seem to be carried out by closely related proteins. These activities corresponded to molecular masses of about 60 kDa, close pI values, and a similar behaviour in non-denaturing polyacrylamide electrophoresis. Both activities were enhanced by Co(+2) and inhibited by EDTA. Among different aminoacyl-p-NAs, they preferentially hydrolysed Lys-p-NA. No increase of Ap activity was obtained by incubation of extracts at acid pH. The maximum PrA and PrB activities detected in N-limited cultures were six-fold higher than those expressed under C- or P-limitation. The effect of culture conditions on the Cp and Ap expression was much less pronounced in comparison with PrA and PrB activities, Ap levels even being slightly higher in C-limited cells. This fact suggests that hydrolysis of protein to peptides might be the limiting step in the pathway of general protein degradation in the vacuole. PMID- 10514561 TI - Differential expression of thyroid hormone receptor isoforms is strikingly related to cardiac and skeletal muscle phenotype during postnatal development. AB - The genomic actions of thyroid hormones (THs) are mediated by receptors (TRs) that are encoded by two protooncogenes, c-erbA-alpha and c-erbA-beta. The precise functions of the TR isoforms are unclear and this study focuses on the potential roles of the TRalpha and TRbeta isoforms in mammalian striated muscles postnatally. The porcine TRalpha1, TRalpha2 and TRbeta1 cDNAs were first cloned, sequenced and characterised by Northern blotting. A quantitative analysis of TR isoform expression was then undertaken, using RNase protection analysis with novel riboprobes designed to detect relative expression levels of TRalpha1, TRalpha2, TRbeta1 and TRbeta2, in functionally distinct muscles from 7-week-old pigs kept under controlled conditions of nutrition and thermal environment. We found a striking muscle-specific pattern of TRalpha isoform distribution: in heart the mRNA level of TRalpha2 (non-TH binding) was markedly greater (P<0.01) than that of TRalpha1 (TH binding); in longissimus dorsi the opposite pattern of expression occurred (TRalpha1>TRalpha2, P<0.001); in soleus, diaphragm and rhomboideus there were no differences between the two isoforms. The overall abundance of TRbeta was very much lower than that of TRalpha, and TRbeta1 was expressed at a higher level than TRbeta2 in all muscles. Together with recent data from TR gene inactivation studies and the established role of TH in determining myosin heavy chain isoform expression and muscle phenotype, these results suggest a role for differential expression of TR isoforms in acquisition and maintenance of optimal cardiac and skeletal muscle function. PMID- 10514563 TI - Role of the sulphate assimilation pathway in utilization of glutathione as a sulphur source by Saccharomyces cerevisiae. AB - Mutants unable to grow on medium containing glutathione as a sole source of sulphur (GSH medium) were isolated from Saccharomyces cerevisiae strains carrying met17(deficiency of O-acetylserine and O-acetylhomoserine sulphydrylase). They were defective in the high-affinity glutathione transport system, GSH-P1. Newly acquired mutations belonged to the same complementation group, gsh11. However, it became apparent that gsh11 conferred the mutant phenotype not by itself but in collaboration with met17. Moreover, mutations conferring the defect in sulphate assimilation made the cell unable to grow on GSH medium in collaboration with gsh11. From this finding, we propose that the sulphate assimilation pathway acts as a sulphur-recycling system and that this function is especially vital to the cell when the supply of glutathione is limited. PMID- 10514564 TI - The yeast PRS3 gene is required for cell integrity, cell cycle arrest upon nutrient deprivation, ion homeostasis and the proper organization of the actin cytoskeleton. AB - PRS3 is one of a family of five genes encoding phosphoribosylpyrophosphate synthetase, an enzyme which catalyses the first step in a variety of biosynthetic pathways, including purine and pyrimidine biosynthesis. We report here that prs3Delta mutants have a number of phenotypes that suggest an unexpected role for PRS3 in linking nutrient availability to cell cycle progression, cell integrity and the actin cytoskeleton. Upon nutrient limitation, prs3Delta mutants fail to arrest in G(1)-cells remain budded and a significant fraction have a G(2) DNA content. Furthermore, in such conditions, prs3Delta mutants have a disorganized actin cytoskeleton: actin accumulates in one or two intensely staining clumps per cell. Prs3Delta mutants also show defects in ion homeostasis and cell integrity. They fail to grow on medium containing 1.0 M NaCl, 5 mM caffeine or when incubated at 37 degrees C. The caffeine and temperature sensitivity are rescued by supplementing the growth medium with 1.0 M sorbitol. These phenotypes resemble those of whi2Delta mutations and indeed, a prs3 allele was recovered in a colony sectoring screen for mutations that are co-lethal with whi2Delta. However, further investigation showed that the prs3Delta whi2Delta double mutant was viable, with no obvious growth defect compared to either single mutant. In the same colony-sectoring assay, an mpk1 allele was also recovered. Multicopy PRS3 rescued the caffeine sensitivity of this mpk1 allele. PMID- 10514565 TI - STRE- and cAMP-independent transcriptional induction of Saccharomyces cerevisiae GSY2 encoding glycogen synthase during diauxic growth on glucose. AB - It has been shown that the so-called stationary phase GSY2 gene encoding glycogen synthase was induced as the cells left the exponential phase of growth, while glucose and all other nutrients were still plentiful in the medium (Parrou et al., 1999). Since this effect was essentially controlled at the transcriptional level, we looked for the cis- and trans-acting elements required for this specific growth-related genetic event. We demonstrated that mutations of the HAP2/3/4 binding site and of the two STress-Responsive cis-Elements (STRE) did not abolish the early induction of GSY2, although the latter mutation led to a 20 fold drop in the transcriptional activity of the promoter, as determined from lacZ gene fusions. Insertion of a DNA fragment (from -390 to -167 bp, relative to the ATG) of the promoter lacking the two STREs, upstream to the TATA box of a CYC1-lacZ fusion gene, allowed this reporter gene to be induced with a kinetic similar to that of GSY2-lacZ. Mutations in BCY1, which results in a hyperactive protein kinase A, did not alleviate the early induction, while causing a five- to 10-fold reduction in the transcriptional activity of GSY2. In addition, the repressive effect of protein kinase A was quantitatively conserved when both STREs were mutated in GSY2 promoter, indicating that the negative control of gene expression by the RAS-cAMP signalling pathway does not act solely through STREs. Taken together, these results are indicative of an active process that couples growth control to dynamic glucose consumption. PMID- 10514566 TI - A novel vacuolar protein encoded by SSU21 / MCD4 is involved in cell wall integrity in yeast. AB - Using a screening procedure for obtaining yeast strains with enhanced ability to secrete heterologous protein, we have isolated a mutant with alteration of the cell wall structure. This mutant displayed strong decrease in cell wall mannoprotein content, which was not accompanied by decreased glycosylation of secreted proteins. The mutation defines a gene, designated SSU21(identical to previously characterized MCD4), which encodes a novel vacuolar protein. SSU21 is probably connected to the cell integrity protein kinase C-mediated pathway, since ssu21 and pkc1Delta double mutant is synthetic lethal. To our knowledge, this is the first example of a yeast vacuolar protein whose alteration results in a cell wall defect. PMID- 10514567 TI - Allelism of Saccharomyces cerevisiae genes PSO6, involved in survival after 3 CPs+UVA induced damage, and ERG3, encoding the enzyme sterol C-5 desaturase. AB - Sequencing of the yeast gene that complemented the sensitivity to the photoactivated monofunctional 3-carbethoxypsoralen of the pso6-1 mutant strain revealed that the ERG3 locus, encoding sterol C-5 desaturase involved in biosynthesis of ergosterol, is allelic to PSO6. Disruption of the ERG3 gene yielded an erg3Delta mutant viable in ergosterol-containing YEPD media with the same pleiotropic mutant phenotype known for pso6-1 and erg3 mutants, including sensitivity to hydrogen peroxide and paraquat. Thus, the erg3/pso6 yeast mutant seems to be more sensitive than the WT to 3-CPs+UVA because of the oxidative damage contributed by this treatment and not because of an impaired repair of the furocoumarin-thymine monoadducts formed in the DNA. We found a significant increase of petites amongst erg3Delta and pso6-1 yeast mutant strains grown in conditions where respiration was mandatory. Mutant pso6-1, with its lowered content of ergosterol, exhibited enhanced synthesis of chitin that was maldistributed and not confined to the bud scars. Chitin overproduction in pso6/erg3 mutants resulted in hypersensitivity to Calcofluor White. PMID- 10514568 TI - Replicative ageing in the fission yeast Schizosaccharomyces pombe. AB - Saccharomyces cerevisiae has been widely used as a model organism in studies of replicative ageing and senescence. The relevance of these studies to ageing in other organisms has, however, been questioned, since this yeast divides by budding rather than fission, the more common pattern in higher organisms. Here we report that, contrary to popular belief, the fission yeast Schizosaccharomyces pombe also undergoes replicative senescence and in a manner superficially analogous to budding yeast. These experiments provide the first evidence of age asymmetry in cell fission and are consistent with the hypothesis of Jazwinski, that asymmetric division underlies culture immortality. Given their evolutionary divergence, comparison of the ageing determinants in fission and budding yeasts may help identify common mechanisms of the ageing process. PMID- 10514569 TI - Sequence analysis of SLA2 of the dimorphic yeasts Candida albicans and Yarrowia lipolytica. AB - We report the complete nucleotide sequence of SLA2 of the dimorphic yeasts Candida albicans and Yarrowia lipolytica. In Saccharomyces cerevisiae, SLA2 codes for an actin binding protein. The deduced amino acid (aa) sequences of C. albicans CaSla2p and Y. lipolytica YlSla2p consist of 1063 and 1054 aa, respectively. The alignment of the deduced proteins of Saccharomyces cerevisiae, Y. lipolytica and C. albicans shows regions of identity in the N-terminal part of the proteins, which are essential for growth at 37 degrees C, endocytosis and actin organization in S. cerevisiae. The Sla2p proteins have also several conserved regions in the C-terminal moiety, the I/LWEQ boxes, displaying homology to the talin protein of mouse, Dictyostelium discoideum, Caenorhabditis elegans and to human huntingtin interacting protein (Hip 1p). The sequence data of C. albicans SLA2 are registered in the EMBL database (AJ009556), and for the Y. lipolytica gene in GenBank (U65409). PMID- 10514570 TI - Disruption and functional analysis of six ORFs on chromosome XV: YOL117w, YOL115w ( TRF4), YOL114c, YOL112w ( MSB4), YOL111c and YOL072w. AB - We have carried out the systematic disruption of six ORFs on chromosome XV, of Saccharomyces cerevisiae using the long flanking homology technique to replace each with the KanMX cassette; we have also constructed plasmids containing replacement cassettes and cognate clones for each ORF. Disruption of three of the ORFs-YOL117w, YOL114c, and YOL112w (also known as MSB4)-does not result in any noteworthy phenotype with respect to temperature or nutritional requirements, but yol112w mutants with an additional disruption of YNL293w, which encodes a protein similar to Yol112w, exhibit a slow growth phenotype. The protein specified by YOL114c shares similarity with the human DS-1 protein. Disruption of YOL115w confers slow growth, cold sensitivity and poor sporulation; this ORF has been described elsewhere as TRF4, which encodes a topoisomerase I-related protein. Cells with disruptions of YOL111c, whose product is weakly similar to the human ubiquitin-like protein GdX, are slightly impaired in mating. Mutants disrupted for YOL072w, the predicted product of which is unrelated to any protein of known function, grow slowly, are cold-sensitive and sporulate with reduced efficiency. PMID- 10514571 TI - Three new dominant drug resistance cassettes for gene disruption in Saccharomyces cerevisiae. AB - Disruption-deletion cassettes are powerful tools used to study gene function in many organisms, including Saccharomyces cerevisiae. Perhaps the most widely useful of these are the heterologous dominant drug resistance cassettes, which use antibiotic resistance genes from bacteria and fungi as selectable markers. We have created three new dominant drug resistance cassettes by replacing the kanamycin resistance (kan(r)) open reading frame from the kanMX3 and kanMX4 disruption-deletion cassettes (Wach et al., 1994) with open reading frames conferring resistance to the antibiotics hygromycin B (hph), nourseothricin (nat) and bialaphos (pat). The new cassettes, pAG25 (natMX4), pAG29 (patMX4), pAG31 (patMX3), pAG32 (hphMX4), pAG34 (hphMX3) and pAG35 (natMX3), are cloned into pFA6, and so are in all other respects identical to pFA6-kanMX3 and pFA6-kanMX4. Most tools and techniques used with the kanMX plasmids can also be used with the hph, nat and patMX containing plasmids. These new heterologous dominant drug resistance cassettes have unique antibiotic resistance phenotypes and do not affect growth when inserted into the ho locus. These attributes make the cassettes ideally suited for creating S. cerevisiae strains with multiple mutations within a single strain. PMID- 10514572 TI - Lipid composition of subcellular membranes of an FY1679-derived haploid yeast wild-type strain grown on different carbon sources. AB - The aim of the project EUROFAN (European Functional Analysis Network) is to elucidate the function of unknown genes of the yeast Saccharomyces cerevisiae at a large scale. Functional analysis is based on general and specific tests with yeast deletion strains. A prerequisite for these studies is a profound knowledge of the biochemistry and cell biology of the corresponding wild-type strain FY1679. As a contribution from our laboratory we present here a systematic lipid analysis of the major organelles isolated from FY1679 grown in the presence of different carbon sources. Phospholipid, sterol and fatty acid composition are characteristic for each organelle. Moreover, growth of the yeast on glucose, ethanol or lactate causes in some cases marked changes of the organelle lipid pattern. As the most prominent example, cultivation of the yeast on non fermentable carbon sources results in an increase of mitochondrial cardiolipin. As another example, the ratio of unsaturated to saturated fatty acids is enhanced in cells grown on ethanol or lactate as compared to glucose. Thus, the lipid composition of yeast subcellular membranes reflects in a significant way the nutrient conditions caused by variation of the carbon source. PMID- 10514573 TI - Transcranial doppler in neurological disorders. PMID- 10514574 TI - Ischaemic stroke: new frontiers. AB - Lot of advancement has taken place, not only in the management but also in the pathophysiology and imaging modalities in patients of stroke. Indolent chronic infections, particularly those due to H. pylori, have been identified as one of the risk factors. The mechanism of inflammation in inducing a precoagulant state has also been worked out. SPECT studies have detected ischaemic areas before appearance of CT abnormalities. CT angiography identifies abnormalities in the 'circle of willis' in posterior circulation strokes much better, and helps weigh the risk versus benefit of thrombolysis. With experiance in use of r-TPA, the list of contra indications and precautions has become longer than its indications. Newer drugs like lubeluzole and edselen have also been recommended. Various other drugs e.g. aptiganel hydrochloride, MDL 28170, 'basic fibroblast growth factor' and 'superoxide dismutase' are at an experimental stage. The concept of a 'stroke cocktail' may be in vogue soon. Controversies still exit regarding the exact indication of prophylactic anticoagulant and the 'international normalized ratio' (INR) to be achieved. Guidelines have been laid down for the approach to patients with asymptomatic carotid artery stenosis. However, the paramount message in stroke care is dissipation of the concept of 'brain attack', amongst the primary care medical and para-medical personnel. PMID- 10514575 TI - Ultrastructural changes in medullomyoblastoma. Similarities with foetal rhabdomyoma. AB - The light and electronmicroscopic changes are described in two cases of medullomyoblastoma, and compared with the changes seen in a case of foetal rhabdomyoma. The medullomyoblastomas in two children aged 8 and 5 years, consisted predominantly of classical type of medulloblastoma cells, along with few to many 'strap cells' or 'myoid cells' which, on closer examination, showed clear cross striations, consistent with muscle fibres or myofibrils. The primitive myoid cells were similar to those encountered in larger numbers in a post-auricular rhabdomyoma, possibly of foetal origin in a 40 day old infant. The four pathogenetic mechanisms i.e. (i) an embryonal stage of myofibrillar differentiation; (ii) a malformative factor; (iii) a teratoid factor on account of the presence of mesenchyme derived striated muscle tissue in the obviously predominant ectodermal medulloblastoma; and (iv) metaplasia of the vascular smooth muscle cells in the medullomyoblastoma, are discussed. PMID- 10514576 TI - Central nervous system toxoplasmosis in acquired immunodeficiency syndrome: An emerging disease in India. AB - With the incidence of patients infected with human immuno-deficiency virus (HIV) increasing in India, the central nervous system (CNS) manifestations of the disease will be seen more frequently. The CNS may be primarily afflicted by the virus or by opportunistic infections and neoplasms secondary to the immune suppression caused by the virus. In India, although mycobacterium tuberculosis has been reported to be the most common opportunistic infection, toxoplasmosis may become as common owing to the ubiquitous nature of the protozoan. Since an empirical trial of medical therapy without histopathological diagnosis is recommended, the true incidence of this condition may remain under estimated. The role of ancillary tests such as radiology and serology in the initial diagnosis of this condition remain crucial. This report highlights two patients who were diagnosed to have acquired immuno-deficiency syndrome (AIDS) only after the biopsy of the intracranial lesion was reported as toxoplasmosis. Presently all patients for elective neurosurgery are tested for HIV antigen. The management protocol to be followed in a known patient with AIDS presenting with CNS symptoms is discussed in detail. The value of ancillary tests is also reviewed. PMID- 10514577 TI - Transoral decompression for craniovertebral osseous anomalies: perioperative management dilemmas. AB - The surgical outcome of 74 patients, who underwent transoral decompression (TOD) for ventral irreducible craniovertebral junction anomalies between January 1989 to September 1997, was studied to evaluate the perioperative complications and problems encountered. The indications for TOD included irreducible atlantoaxial dislocation (n=24), basilar invagination (n=16), and a combination of both (n=35). Following TOD, occipitocervical stabilization using Jain's technique was carried out in 50 (67.5%) and atlantoaxial fusion using Brooks' construct in 18 (24.3%) patients. The pre- and postoperative radiology was compared to assess the adequacy of decompression and stability. The major morbidity included pharyngeal wound sepsis leading to dehiscence (20.3%) and haemorrhage (4%), valopharyngeal insufficiency (8.1%), CSF leak (6.7%) and inadequate decompression (6.7%). Neurological deterioration occurred transiently in 17 (22.9%) and was sustained in 7 (9.4%) patients. The mortality in six cases was due to operative trauma, exanguination from pharyngeal wound (one each), postoperative instability and inability to be weaned off from the ventilator (two each). Of the 47 (63.5%) patients available at follow up ranging from 3 months to 2 years, 26 (55.3%) showed improvement from their preoperative status while 14 (29.8%) demonstrated stabilization of their neurological deficits. Seven (14.9%) of them deteriorated. Though TOD is logical and effective in relieving ventral compression due to craniovertebral junction anomalies, it carries the formidable risks of instability, incomplete decompression, neurological deterioration, CSF leak, infection and palatopharyngeal dysfunction. PMID- 10514578 TI - Pathology of temporal lobe epilepsy: An analysis of 100 consecutive surgical specimens from patients with medically refractory epilepsy. AB - The neuropathological features of temporal lobe epilepsy were studied utilising 100 consecutive surgical specimens from patients with medically refractory complex partial seizures. A wide spectrum of neuropathological changes was recorded in 98 specimens. Fifty-eight specimens showed features of Ammon's horn sclerosis. Diffuse accumulation of corpora amylacea were demonstrated in the resected temporal lobes from 54 patients. Six patients had neoplastic lesions of temporal lobe. One unique case of dysembryoplastic neuroepithelial tumour showed a melanotic component within the tumour. The neuropathological features were regarded as nonspecific in 31% of cases. Our results indicate that a majority of patients with medically intractable epilepsy of temporal lobe origin reveal significant neuropathological features. Careful documentation of the neuropathological features and its correlation with radiological, electrophysiological and pre- and post-surgical clinical features will help in predicting the seizure outcome after temporal lobectomy for medically refractory epilepsy. PMID- 10514579 TI - Pyogenic brain abscess managed by repeated elective aspiration. AB - 37 cases of capsular stage of brain abscess based upon CT scan staging were treated by repeated elective aspiration through a burr hole and intracavitary application of antibiotics on alternate days, till two consecutive negative aspirations were obtained. A combination of furosemide and antibiotics in multiple doses were also given. The mortality rate was 2.7% and the morbidity rate 8.3%. Corticosteroids were not used in the management of brain abscess. Thus, repeated elective aspiration was found to be an effective mode of surgical management of brain abscess. PMID- 10514580 TI - Botulinum toxin treatment of hemifacial spasm and blepharospasm: objective response evaluation. AB - Twenty seven patients with hemifacial spasm (HFS) and sixteen patients with blepharospasm (BS) having mean Jankovic disability rating scale score of 2.56+0.58 SD and 2.81+0.54 SD, respectively, were treated with botulinum toxin A (BTX-A) injections. The total number of injection sessions were ninety one with relief response in 98.91%. The mean improvement in function scale score was 3.78+0.64 SD and 3.29+1.07 SD respectively, in HFS and BS groups. The clinical benefit induced by botulinum toxin lasted for a mean of 4.46+3.11 SD (range 2 to 13) months in HFS group and 2.66+1.37 SD (range 1 to 6) months, in BS groups. Transient ptosis was seen in 4.39% of total ninety one injection sessions. These findings show that local botulinum toxin treatment provides effective, safe and long lasting relief of spasms. PMID- 10514581 TI - Factors of error involved in the diagnosis of juvenile myoclonic epilepsy: A study from South India. AB - The study was aimed at finding possible factors for delay in the diagnosis of juvenile myoclonic epilepsy (JME) in a developing country. Data was analyzed retrospectively through the medical records and prospectively through a re evaluation of the history and EEGs of patients with JME registered in a university hospital in south India. Of the 131 patients, 23 (17.5%) patients were seen by neurologists before registration in the clinic. Diagnosis of JME was established in 118 patients at the time of registration and in 13 (10%) patients during follow-up in the clinic. The mean interval between onset of disease and the diagnosis was 6.8 + 6.3 years. In 20 patients the diagnosis was established 10 years after the onset. The mean interval between the first evaluation and diagnosis was 24. 2 months in the 13 patients in whom the diagnosis was established during follow-up in the clinic. Lack of familiarity with the clinical syndrome was probably the factor for delay in the diagnosis in 108 patients seen by practising physicians. The factors for delay in the diagnosis in patients seen by neurologists included failure to ask about myoclonic jerks resulting in misinterpretation of EEGs in 28 patients, misinterpretation of absences and/or unilateral jerks in 4 patients, and failure to ask about myoclonic jerks and misinterpretation of focal EEG abnormalities in 4 patients. This study suggests that the possible factors of error in the diagnosis of JME among the neurologists were similar to the observations reported from the developed countries; whereas the factor for delay in the diagnosis of JME among practising physicians was lack of familiarity with the epileptic syndrome. PMID- 10514582 TI - Intracranial hydatid cyst: a report of five cases and review of literature. AB - The authors present five cases of intracranial hydatid cysts managed at the department of Neurosurgery, King Edward Memorial Hospital, Mumbai, between 1984 1997. The mean age of presentation was 13.4 years. Four patients (80%) were in the first decade of life. All patients presented with focal neurological deficit and clinical features of raised intracranial pressure. Radiological investigations included computerised tomography (CT) scan in three cases, CT and magnetic resonance (MR) scan in one case and accidental cystogram in one case. Two patients had multiple intracranial cysts. One patient had a solitary cyst in the lateral ventricle. Commonest location was in the parietal lobe (3 cases). Total excision of the cyst was done in all five cases. Recurrence was seen in two cases, probably as a result of rupture of the cyst during first surgery. The features of this rare disease are retrospectively analyzed in this presentation and the literature is reviewed. PMID- 10514583 TI - Genetic polymorphism in muscle biopsies of Duchenne and Becker muscular dystrophy patients. AB - Duchenne muscular dystrophy (DMD), with an incidence of one in 3500 male new borns, and its milder variant, Becker muscular dystrophy (BMD), are allelic X linked recessive disorders, caused by mutations in the gene coding for dystrophin, a 427 kD cytoskeleton protein. There are no available molecular markers to differentiate these two. The purpose of this study was to study genetic polymorphism in muscular dystrophy and explore its potential in discriminating these two allelic forms of the disease. The results revealed unambiguously the presence of three transcripts : 598bp, 849bp and 1583bp long which are selectively expressed in the muscles afflicted with muscular dystrophy as compared to the normal muscle. 1583bp gene transcript was conspicuously present in the muscle tissues of both DMD and BMD patients whereas 598bp and 849bp long transcripts were exclusively present in DMD but not in BMD patients or normal human subjects. These gene transcripts had no sequence homology with dystrophin gene and these were also present in the families belonging to DMD and BMD patients. These results point to the fact that based upon the selective expression of these three gene transcripts, one could not only differentiate between DMD and BMD diseases at the molecular level, but also between normal and dystrophic muscle. Further, these findings also reveal that apart from dystrophin gene, these gene transcripts may also be responsible for the differential progression of DMD/BMD phenotype. PMID- 10514584 TI - Management of intramedullary spinal cord tumours: review of 68 patients. AB - 68 consecutive patients admitted with intramedullary spinal cord tumours and operated at Vellore during a six year period from January 1990 are discussed. 41 tumours were radically resected, 11 partially excised while 14 had only a biopsy. Radiation therapy was advised post operatively to those patients for whom a partial excision or biopsy was done. There was no postoperative mortality. Two patients developed wound infection and one developed postoperative hydrocephalus. Postoperative clinical assessment between four to eight weeks after surgery showed that 25 out of 68 patients improved, 29 remained unchanged, while 14 had worsening of deficits. Immediate post operative assessment, however, was less encouraging. Evaluation of these patients was done using a functional scoring system and Karnofsky rating. The follow up period ranged from 2 weeks to 64 months after discharge from hospital with a mean of 14.6 months. The indicators of radical excision were good tumour-cord interface, cranially located tumours, presence of syringomyelia and histology of ependymoma. Two patients had recurrence of tumour. PMID- 10514586 TI - Solitary plasmacytoma presenting as peripheral neuropathy: a case report. AB - We report a 26 year old woman presenting with a chronic predominantly motor radiculoneuropathy. A biopsy proven solitary lytic plasmacytoma was detected in the left sixth rib. Serum immunoelectrophoresis revealed 'M' band (IgG, l light chain). Excision of the plasmacytoma resulted in gradual but steady recovery. We suggest that all patients with 'cryptogenic' polyneuropathy be investigated for this potentially treatable condition. A high index of suspicion assists the organization of appropriate diagnostic investigations. PMID- 10514585 TI - A study of transcranial magnetic stimulation in older (>3 years) patients of malnutrition. AB - Transcranial magnetic stimulation was performed in 40 subjects. Twenty patients in the age group of 3 to 8 years and having different grades of malnutrition were included in the 'study group' whereas 20 normal children having no complaints comprised the 'control group'. The coil of the magnetic stimulator was applied tangentially over the vertex to stimulate the cortex. The motor evoked potential (MEP) was obtained using root stimulation by applying the coil at the cervical and lumbosacral spines. Recordings were made from the abductor pollicis brevis (APB) and extensor digitorum brevis (EDB) muscles of both sides. Cortical threshold, latency and amplitude of motor evoked potential and central conduction time were recorded. Malnourished children showed significantly increased cortical threshold, prolonged cortical latency and central conduction time and reduction in amplitude of MEP. Observed delay in central motor conduction in malnourished children suggests asymptomatic involvement of corticospinal pathways. PMID- 10514587 TI - Tuberculoma in the Meckel's cave: a case report. AB - A case of an intracranial tuberculoma located within the confines of the Meckel's cave is presented. The patient was young, non-immunocompromised and otherwise in good health. The granuloma mingled with the fibres of the trigeminal nerve. The lesion mimicked a trigeminal neurinoma in its clinical presentation, preoperative investigations and intraoperative consistency and vascularity. The rarity of the location and possible mode of transmission of infection to this site is discussed. The literature on this subject is briefly reviewed. PMID- 10514588 TI - Facial myokymia as the presenting symptom of a pontine glioma. PMID- 10514589 TI - Nocardial abscess of spinal cord. PMID- 10514590 TI - Idiopathic hypoparathyroidism presenting as epilepsy in a 40 years female. PMID- 10514591 TI - Basilar artery occlusion in cryptococcal meningitis. PMID- 10514593 TI - Glioma mimicking a tuberculoma. PMID- 10514592 TI - Melanoma secondaries presenting as stroke. PMID- 10514594 TI - Cavernous sinus haemangioma with hereditary multiple exostoses. PMID- 10514595 TI - Transcranial Doppler studies of middle cerebral artery blood flow following different test conditions. PMID- 10514596 TI - Acquired cauda equina epidermoid cyst. PMID- 10514598 TI - Social determinants of health, the physical environment and public health nursing. PMID- 10514597 TI - Conus medullaris teratoma presenting as myokymia. PMID- 10514599 TI - Once upon a time ... narratives and research. AB - Research incorporating the use of narratives is not new, yet it has been subject to a revival, as if new, in more recent times. Whilst the many disciplines that use narratives may emphasise different foci, there is agreement with the fundamental premise that people convey the meanings of their everyday life though the use of narratives. The purpose of this paper is to explore what narratives are and how they are used in research by reviewing the current literature and weaving the story of my learning journey through the paper. PMID- 10514600 TI - The evening tea break ritual--a case study. AB - Many nursing scholars, such as Carter (1990), Ford and Walsh (1994), Huey (1986), James (1990), Thomlinson (1990), Larson (1987), Rush et al (1990), Spiller (1992), Street (1990a,b; 1992a,b,c,d), Lawrence (1996), Walker (1967), Walsh and Ford (1989 a,b,c,d; 1991 a,b), have examined the effects of rituals in nursing practice, and have argued for a need to scrutinise these ritual practices because of their negative effect on patients. Other scholars, such as Holland (1993) and Biley and Wright (1997), have argued that these rituals have meanings for nurses. Street (1992c) has argued that rituals are resilient in nursing practice. In order to identify the factors that contribute to rituals' resilience, there is a need to understand their meanings and effects on nurses, and how nurses keep them alive. This study has explored the meanings of the 'evening tea break' ritual to a group of nurses in a medical ward. The study has employed an ethnographic approach, and has found that the evening tea break ritual had affective effects on the participating nurses. It provides the time, space, the process and the environment for nurses to ventilate their feelings, to gain support from each other. PMID- 10514601 TI - Breaking the silence: marginalisation of registered nurses employed in nursing homes. AB - This paper reports on a study which employed auto/biography as a research method to expose the marginalisation and discrimination experienced by Registered Nurses (RN) employed in the nursing home sector. By virtue of their area of practice, these nurses are frequently perceived by the wider nursing community and society at large to be deskilled and not competent. A critical incident from my story is recounted and thereafter discussed from a personal perspective and within a wider context of nursing and related literature. I argue that the complexity of clinical practice in a nursing home means that RNs are no less skilled or competent than RNs in other clinical settings. However, marginalisation and discrimination occur because nurses and nursing are not isolated from the values and beliefs of society. This is a society which under values caring and nurturing and places a higher value on complex technologies with an emphasis on cure being the dominant ideology. The style of reporting the research is congruent with an auto/biographical approach. PMID- 10514602 TI - Nurses re-entering the workforce: a special needs group. AB - Registered Nurses wishing to re-enter the paid workforce have been shown to experience low self-esteem and high levels of anxiety. An intensive twelve-week re-entry program was designed to address these needs through a process of critical pedagogy. In this paper the student cohort since 1995 is described and the program, its structure, style, physical environment, teaching, and course content are introduced. Evaluation of student outcomes was ascertained by a convenience survey in which a questionnaire was posted to successful students. The results included employment rates of 97.4% and high levels of work and personal satisfaction. Both the benefits and the limitations of the program are discussed in light of the questionnaire responses. PMID- 10514603 TI - The lost children. AB - Women who have lost children to perinatal complications, are subjected to pain and grief continuously; however their agony increases on remembrance days such as birthdate, or on Mother's Day. Fathers, siblings and grandparents suffer too. Common disorders of pregnancy, such as pregnancy-induced hypertension (PIH), or the more serious pre-eclampsia, HELLP syndrome, or eclampsia, can lead to devastating effects such as miscarriage, stillbirth, or neonatal death; or at the very least, a sick infant. With many of these consequences, the loss of the dreams, hopes and plans that parents have made is imminent. The investigation of the psychosocial aspects of 'high-risk' pregnancy has never been fully addressed. However, the threat of loss, or the actual experience, may provoke the onset of a potential psychological crisis during the perinatal period. Therefore, it is important that these issues be addressed by the nurse in order to aid the development of coping mechanisms to enable women and their families to deal with what may happen. This may be done by predicting the stages of the bereavement process experienced by these women and their family members, as outlined in the Kubler-Ross model of bereavement (1969), which is indicative of many types of grief reactions. Other issues including the restriction in activity, uncertainty of pregnancy outcomes, disruption in work or career activities, financial strains, and reduced labour and birthing options, become concerns for high-risk pregnant women. The way women deal with these issues and the pathways nurses can take to help these women develop effective coping strategies, will be addressed also. PMID- 10514604 TI - Leadership: a means of empowering the nursing profession. AB - For too long now, nurses have assumed an obsequious role in their working relationship with other health professionals. The nursing profession holds the majority membership in the field of health care professions yet nurses continue to be cast into subordinate roles within professional relationships, failing to be recognised as peers. This is despite many nurses possessing unique but analogous knowledge, credentials and professionalism. It is time nurses were agitated to reflect upon their position and be reminded nursing is also a profession. Some suggestions and motivation are offered. PMID- 10514605 TI - Insulin infusion pilot study. AB - To help resolve debate as to the relative efficacy of two different insulin delivery media, we set out to examine differences between Haemaccel and Normal Saline with reference to our hospital's current infusion procedures. After analysing the data, we found that there was a statistically significant difference between Haemaccel and Normal Saline and that, for solutions containing one unit of soluble insulin per ten mls of carrier solution, Haemaccel was the carrier of choice and clearly indicated as the preferable medium in acute diabetic situations like DKA and HHNKC. PMID- 10514607 TI - Enhancing safety in a pediatric unit. AB - This descriptive study demonstrates how a group of nurses in the clinical setting can identify and implement a safe sharps disposal strategy to enhance safety. The aim of this small study was to determine a safer method of disposing used sharp items. The findings indicated that many factors can contribute to inappropriate disposal of used sharp items. Changes in practice were implemented following the findings of this study and resulted in enhanced safety in this paediatric unit. In addition, results of the study were utilised to contribute to evidence based nursing practice. PMID- 10514606 TI - Performance of urinary catheterisation by mothers of children with spina bifida. AB - This paper describes a review of the practices related to the performance of catheterisation for obtaining residual volume of urine by mothers of children with spina bifida. Mothers of children, up to five years of age and attending an outpatient Specialist Clinic were requested to perform catheterisation once a month to obtain urine residuals. The procedure for catheterisation was taught to mothers in the Outpatient's Clinic. A review of clinical practice resulted in discontinuation of this procedure. Mothers (n = 16) reported varied reasons for feeling relieved that the procedure was discontinued. PMID- 10514608 TI - Clinical experience in undergraduate education: issues and challenges. PMID- 10514609 TI - Community mental health nursing. PMID- 10514610 TI - A primary health care service in a general practitioner's practice. PMID- 10514611 TI - Saying the last goodbye consciously. PMID- 10514612 TI - The nurses pay award. PMID- 10514613 TI - Drumcree. Ready for peace, preparing for disaster. PMID- 10514614 TI - Reflective practice. AB - Christine Nash exemplifies reflective practice as a frequently used but infrequently defined concept in nursing (Jarvis 1992), with intuition seldom granted legitimacy as a sound approach to clinical judgement (Benner and Tanner 1987). She follows with a personal account of how she used reflective practice and intuitive skills in reassessing an A&E attender. PMID- 10514615 TI - Ocular burns. PMID- 10514616 TI - Shoulder injury: a case study. PMID- 10514617 TI - The airway and the A&E nurse. PMID- 10514618 TI - Essence of emergency nursing. PMID- 10514619 TI - The effects of systemic illness on the eye and vision. PMID- 10514620 TI - Interpersonal approaches to managing violence and aggression. PMID- 10514621 TI - Acute poisoning: antituberculosis drugs--clinical features and management. PMID- 10514622 TI - Managing major incidents. A dynamic educational initiative. PMID- 10514623 TI - Shaping the role of the clinical placement coordinator in an Irish A&E. PMID- 10514624 TI - Paediatric resuscitation. PMID- 10514625 TI - FDA and NIOSH public health advisory: explosions and fires in aluminum oxygen regulators. PMID- 10514626 TI - The power of hidden numbers. PMID- 10514627 TI - Evaluation of potential closed head injury in patients with known coagulopathy. AB - A case report is provided of an elderly "minor" trauma patient in whom an acute subdural hematoma developed. The division of traumatology and surgical critical care worked with other hospital departments to develop and implement a trauma care guideline, promote ongoing communication between departments, and monitor compliance with the guideline. PMID- 10514629 TI - Practice basics: quick neurovascular examination of the hand. PMID- 10514630 TI - You've got mail! PMID- 10514633 TI - Esophageal detector device: changes for trauma care. PMID- 10514631 TI - The human factors implications of peritoneal dialysis: cycler overfill incident reports. AB - Serious errors can occur with many medical devices (e.g., infusion pumps, ventilators, anesthesia machines), and even highly trained professionals can make critical errors. The FDA now requires that manufacturers consider the user when designing medical equipment and has issued a guidance document describing human factor (HF) problems and the HF design process. The Association for Advancement of Medical Instrumentation has also published guidelines, and the American Medical Association's National Patient Safety Foundation has selected reduction of medical errors as their mission. It is important that the medical community reinforce the patient-safety initiative by evaluating user-interface design and instructional manuals when purchasing such equipment. The payoff will be fewer incidents and less time required in device training. The FDA actively solicits help in identifying and reporting adverse events associated with medical devices. Health care practitioners employed by facilities subject to the FDA's user facility reporting requirements should follow the user-reporting procedures established by their facility. Practitioners and users may want to directly report an incident to the MedWatch Program, the FDA's voluntary Medical Products Reporting Program. PMID- 10514634 TI - Glass capillary tubes: safety advisory. PMID- 10514635 TI - "Tough as nails, heart of gold"--nurses in the refugee camps of Macedonia. Interview by Judith Stoner Halpern. PMID- 10514636 TI - Trauma nursing in a safe community. PMID- 10514637 TI - Follow-up chest radiographs after traumatic pneumothorax or hemothorax in the outpatient setting: a retrospective review. AB - PURPOSE: To evaluate the need for obtaining postdischarge chest radiographs for trauma patients who were treated with a thoracostomy tube. METHODS: A retrospective medical record review was conducted for all patients treated with a thoracostomy tube while admitted to the trauma service at Saint Louis University Hospital over a 12-month period. Patients who died during their hospital stay were excluded. RESULTS: During the 12-month study period, 155 trauma patients who were treated with a thoracostomy tube were discharged from the hospital. The indications for the thoracostomy tube were pneumothorax (n = 79, 51% of study population), hemopneumothorax (n = 34, 22%), hemothorax (n = 28, 18%), diaphragmatic rupture/laceration (n = 8, 5%), post thoracotomy (n = 4, 3%), and iatrogenic pneumothorax (n = 2, 1%). A follow-up clinic visit was scheduled for 1 to 2 weeks after discharge. Forty patients (26%) were lost to follow-up. Two patients called to report they had no symptoms and canceled their appointments. A total of 113 patients returned for follow-up appointments. Fifty-two patients had a predischarge chest radiograph that was negative for pneumothorax or hemothorax, had no symptoms, had normal results of a physical examination at the time of their clinic visit, and did not have a postdischarge chest radiograph. A total of 61 (54%) had postdischarge chest radiographs. Of that number, 56 (92%) were negative for pneumothorax. Three patients (5%) had a small pneumothorax, and 2 patients (3%) were noted to have a resolving hemothorax. All 5 patients were without symptoms and were released from the trauma service. CONCLUSION: A postdischarge chest radiograph is not indicated for an asymptomatic trauma patient who was treated with a tube thoracostomy and had a predischarge chest radiograph that was negative for pneumothorax or hemothorax. PMID- 10514639 TI - Practice basics: quick neurovascular examination of the leg and foot. PMID- 10514638 TI - Helping the bereaved at the emergency department: a study at the Brussels University Hospital. AB - An evaluation was conducted of the emergency staff who cared for relatives of patients who had died suddenly. Physicians and nurses completed questionnaires and participated in follow-up meetings to discuss the results of the survey and to recommend actions to improve care for both bereaved families and staff. An informational brochure was developed that provides standard and customized information for families. PMID- 10514640 TI - Thermoregulation in pediatric trauma: an overview. PMID- 10514641 TI - Spotlight on forensic nursing. PMID- 10514642 TI - Nurses play a pivotal role in adverse event problem identification. PMID- 10514643 TI - Common causes for medication errors identified. PMID- 10514644 TI - Parents', physicians' and nurse practitioners' perceptions of behaviors associated with Attention Deficit Hyperactivity Disorder. PMID- 10514645 TI - Planning end of life care. PMID- 10514647 TI - Health care services for adolescents: are we meeting developmental needs? PMID- 10514646 TI - Developing a community based screening program: commitment to the underserved. PMID- 10514648 TI - Managing cardiovascular risk in noninsulin dependent diabetes mellitus. PMID- 10514649 TI - [Holistic care concept]. PMID- 10514650 TI - [The handicapped child in the hospital]. PMID- 10514651 TI - [Socio-pediatric aspects in children and adolescents with multiple abnormalities]. PMID- 10514652 TI - [Organ transplantation--nursing aspects of care]. PMID- 10514653 TI - [Kidney transplantation in children]. PMID- 10514654 TI - [Care of children with poisonings]. PMID- 10514655 TI - [Handling drugs so as to prevent suicides in psychiatry]. PMID- 10514656 TI - [Mixing of infusions]. PMID- 10514657 TI - [Difficult birth]. PMID- 10514658 TI - [Disease concepts and their management by chronically ill children--chronic arthritis]. PMID- 10514659 TI - [Project: holistic neonatal nursing--accessible to nursing students?]. PMID- 10514660 TI - [First care with subsequent preoperative and postoperative care in coccygeal teratoma]. PMID- 10514662 TI - [Drugs during pregnancy]. PMID- 10514661 TI - [Incubator care versus "open care" in the warming bed for very small premature infants]. PMID- 10514663 TI - [Mentoring of students in pediatric nursing]. PMID- 10514664 TI - [Employee resignation and its protection]. PMID- 10514665 TI - [Additional leave for coworkers who nurse and treat patients with infectious diseases]. PMID- 10514666 TI - [What is moving us]. PMID- 10514667 TI - [Monitoring on the premature ward]. PMID- 10514668 TI - [Monitoring of oxygen therapy in the newborn]. PMID- 10514669 TI - [Air transportation of children and newborns with heart disease]. PMID- 10514670 TI - [Types of albinism and their ocular and cutaneous manifestations]. PMID- 10514671 TI - [Disorders during pregnancy and lactation]. PMID- 10514672 TI - [Motivation and its stimulation]. PMID- 10514673 TI - [Children and travel]. PMID- 10514674 TI - [Toxic and less toxic plants. 42]. PMID- 10514675 TI - [Concerning the article: "The handicapped child in the hospital]. PMID- 10514676 TI - [Touch--food for the newborn]. PMID- 10514677 TI - [Physiotherapeutic measures on the premature-intensive care unit]. PMID- 10514678 TI - [Sexual abuse of children and adolescents. Pedophilia--does it still exist?]. PMID- 10514679 TI - [The concept of pain in children and adolescents]. PMID- 10514680 TI - [Sports and movement during pregnancy and lactation]. PMID- 10514681 TI - [Toxic and less toxic plants. 43]. PMID- 10514682 TI - [Looks, aura and presence in nursing: there is no second chance for a first impression]. PMID- 10514683 TI - [Learning with all one's senses. Neurolinguistic programming in the teaching of pediatric nursing]. PMID- 10514684 TI - [Indian baby massage]. PMID- 10514685 TI - [Nosocomial infections at intensive care units. Don't give germs a chance!]. PMID- 10514686 TI - [Correct handling of venous catheters]. PMID- 10514687 TI - [Hand hygiene. Impressive potential for prevention]. PMID- 10514688 TI - [Everything changes. Entering an old age home]. PMID- 10514689 TI - [Short stories. As long as there is life...]. PMID- 10514690 TI - [We found an approach]. PMID- 10514691 TI - [The importance of communication]. PMID- 10514692 TI - [Ways of communication: so that discussions promote contact]. PMID- 10514693 TI - [Advantages of oil in the bath]. PMID- 10514694 TI - [Understanding autism]. PMID- 10514695 TI - [A way of "traveling along"]. PMID- 10514696 TI - [A nursing concept: caring--helping the other to grow]. PMID- 10514697 TI - [No caring without conscience. What to do about disgust?]. PMID- 10514699 TI - [Labor market. Independent women]. PMID- 10514698 TI - [Everyday ethics. Is silence always golden?]. PMID- 10514700 TI - Nurses must insist that a clean hospital is not only desirable but essential. PMID- 10514701 TI - From latex, with glove. PMID- 10514702 TI - Bottling out over formula feed. PMID- 10514703 TI - Off-the-shelf child care. PMID- 10514704 TI - Staying the course. Interview by Adrian O'Dowd. PMID- 10514705 TI - Head start or handicap? PMID- 10514706 TI - What is a local health care cooperative? PMID- 10514707 TI - Can nurses come off the fence and welcome support workers as part of the family? PMID- 10514708 TI - Hello to fresh ideas about PREP. PMID- 10514709 TI - Tropical troubles. PMID- 10514710 TI - Bon voyage. PMID- 10514711 TI - Should relatives be allowed into the resuscitation room? PMID- 10514712 TI - Compulsory treatment. The chase is on. PMID- 10514713 TI - Elderly care. Partners in care. PMID- 10514714 TI - Property prices. PMID- 10514715 TI - So you want to be a ... herbalist. PMID- 10514716 TI - Admission criteria to short-stay units. PMID- 10514717 TI - Gastrointestinal bleeding. PMID- 10514718 TI - The advantages of using suppositories. AB - Suppositories offer a broad spectrum of clinical benefits but are often not used because of patient prejudice. Do patients need to be educated to overcome the psychological barriers to their use or should nurses undergo further training first? PMID- 10514719 TI - The hospital care of patients with diabetes. PMID- 10514720 TI - Respiratory care. Clear path ahead for specialists. PMID- 10514721 TI - Respiratory care. Out of breath, into rehab. PMID- 10514722 TI - Respiratory care. Under pressure. PMID- 10514723 TI - [Chosing between death and the devil] [In Process Citation] PMID- 10514724 TI - [The market is booming but nurses and patients suffer]. PMID- 10514725 TI - [Board for Aid to the Aged: limiting grievances in care. At least 60% specialists are necessary]. PMID- 10514726 TI - [Alarm at the nurses association: ever more nurses fall ill]. PMID- 10514728 TI - [New topics at the Baden-Baden summer academy: from grievance management to discussions with graduate scholars] [In Process Citation] PMID- 10514727 TI - [Not enough young people in geriatric nursing. Lack of interest in social professions in the young]. PMID- 10514729 TI - [Congress on "Multimedia in Nursing": nursing schools have to catch up]. PMID- 10514730 TI - [Foreigners in nursing: "We want to create orderly conditions for ourselves. Interview by Elisabeth Malleier]. PMID- 10514731 TI - [Palliative work in a community hospital: hospice rooms promote discussions about the meaning of death]. PMID- 10514732 TI - [Alternative care for prematures: oils, music and Bach flowers for the "preemie"]. PMID- 10514733 TI - [Giving a subcutaneous injection: the nurses are responsible]. PMID- 10514734 TI - [Indemnity legislation: employee's liability was restricted]. PMID- 10514735 TI - [Interviewing patients in the pediatric ward: how children and their parents experience the hospital]. PMID- 10514736 TI - [Teamwork in nursing management: an unreal idea or a working necessity?]. PMID- 10514737 TI - [Quality circles in the hospital: external counsellors help to carry things through]. PMID- 10514738 TI - [Masters studies at Edinburgh University: study course with international flair]. PMID- 10514739 TI - [Doubts about modern treatment methods but: moist wound treatment is cheaper]. PMID- 10514740 TI - [Patient instructions--a contribution to the formation of the last phase of life? Considerations on the ethical aspects of the patients' autonomy]. PMID- 10514741 TI - [Personal qualities of nurses]. PMID- 10514742 TI - [A dialectic view of the efficacy of nursing interventions. Connections between nursing interventions and effects of psychotherapy]. AB - The following essay deals with the question whether there are psychotherapeutic effects within nursing interventions. The first part describes the framework for nursing interventions according to the nursing theories of I. King considering particularly the process of goal-achievement. Humanistic thoughts and views of Patterson, Zderad and Watson have also been taken into consideration. In order to look at the question concerning the effectiveness of psychotherapeutic intervention the scientific studies of K. Grawe, R. Donati und F. Bernauer as well as those of V. Tschuschke und D. Czogalik are discussed. Particular attention has been paid to the so-called non-specific factors with regard to the effectiveness of the interventions which was the basis for the formulation of the question. The second part combines the non-specific factors relating to the effectiveness of psychotherapy and the way in which nursing interventions work which is the basis for a model relevant to the different levels of nursing intervention. From this synthesis it can be said that nursing interventions do process psychotherapeutic effectiveness. To come to a conclusion the necessary consequences regarding training and continuing education in nursing management are discussed. PMID- 10514743 TI - [Fatigue in healthy and cancer patients. 1. A qualitative study on conceptual analysis]. AB - Interest in fatigue-research has grown since the finding that fatigue/tiredness is the most frequently reported symptom of cancer and its treatment. But even though several authors have tried to conceptualise fatigue, its mechanisms are still poorly understood. The aim of this study was twofold: 1) to explore fatigue in cancer patients, inductively, and 2) to compare experiences of fatigue/tiredness of healthy individuals with that of cancer patients to identify cancer-specific fatigue/tiredness and related concepts. PATIENTS AND METHODS: A qualitative research strategy was adopted using a grounded theory approach. The prospective study took place in the Oncology Department of the Kantonsspital St. Gallen (Switzerland) with samples of 20 cancer patients and 20 healthy individuals. Unstructured, tape recorded interviews were conducted to collect data. The transcripts of the interviews were analysed using content analysis and constant comparison. RESULTS: Different themes emerged between the two groups although both fitted a classification system, which categorised fatigue into physical, affective and cognitive expressions of fatigue/tiredness. Physical signs were more frequent than affective and cognitive signs in both groups. In the cancer patients, fatigue involved decreased physical performance, extreme, unusual tiredness, weakness and unusual need for rest, which was distinctly different for healthy persons. Affective and cognitive distress was also more prominent in cancer patients. Interestingly, the concept of malaise was not identified by either sample and not understood as an expression of fatigue by this German speaking population. Linguistic differences in the description of fatigue/tiredness between healthy and ill individuals revealed different perceptions of the phenomenon. A step-like theory, explaining the production of fatigue/tiredness was tentatively put forward involving nociception, perception and expression of tiredness. CONCLUSIONS: The emerging concepts break tiredness/fatigue into expression of physical, affective and cognitive tiredness/fatigue. The experience is different between healthy individuals and cancer patients. Generalisability of data needs precaution but the results of the study identifies and clarifies ideas that might form an important basis for further, controlled studies. PMID- 10514744 TI - [The role of nursing in the rehabilitation of acute stroke patients. Towards a unified theoretical perspective]. AB - A review of existing literature indicates an uncertainty about the specific therapeutic role of nurses in the rehabilitation of stroke patients. Two different conceptualizations of the nursing role exist, but neither is related to specific rehabilitation goals and patient outcomes. A beginning theoretical account of the specific role of nursing in stroke recovery is offered as a structure to integrate the therapeutic aspects of the coordinating, maintenance and training functions of the nurse. Existing research literature is reviewed to substantiate the account. Further research is needed to develop the specific content and focus of nursing in stroke recovery. PMID- 10514745 TI - [Interviews in historical nursing research]. AB - In this article the method of interviewing is discussed in its context with historical nursing research. In the first part the development of the academic discipline of history and historical nursing research are outlined briefly. A discussion on the research method interview follows and the concept of oral history is described. Lastly, examples in historical nursing research using the method interview are examined. PMID- 10514746 TI - [Nursing science and natural sciences. Critical notes on a difficult relationship as a starting point for its new definition]. AB - This article deals with the question of how nursing, supported by its own science, can gain benefit from natural science disciplines, if it can be presupposed that such disciplines represent one of the most important bases of nursing science. In the context of this question it is necessary to ascertain the condition, which must be fulfilled by the natural sciences in order to be able to occupy the honorable position of a cognate discipline of nursing science. This position is elucidated on the background of following aspects: The fundamental relationship between nursing science and the natural science with regard to their traditional perspectives and scientific objectives is discussed and illuminated on the background of differing concepts of body and person. In a further elaboration of this theme the particular meaning of the concept of person for nursing action is taken up and on its bases an extension of nursing issues which should be further developed is devised. In conclusion some propositions are brought up for discussion. PMID- 10514749 TI - [Guaranty of information: the case of nosocomial infections] PMID- 10514747 TI - [Effects and importance of higher nursing education (Part 1). On the further development and work satisfaction of students. Presentation and discussion of qualitative questioning of graduated students]. AB - This article shows the outcome of a quality questionnaire from students who have completed a post-diploma course in advanced nursing at the Cantonal Hospital Basle. The results show particularly the relevance of this education/course for the practical work. The final discussion and interpretation refers to the whole outcome. PMID- 10514748 TI - [Qualitative methods in the planning and designing of teaching units in the nursing profession]. AB - New teaching methods for the continuing education need to be developed in order to further professionalize nursing. Favoring qualitative procedures in teaching units of the planning and structure of nursing help make subjective theories that influence happenings in nursing more transparent. The aim of practical-oriented teaching units in nursing is to influence nursing according to the new nursing paradigm. PMID- 10514750 TI - [Uncertainty, agency relationships and miscommunication]. AB - This author finds it curious that for a long time, whenever I have tried to explain what an agent relationship is, everyone understands the concept with ease. "It's common sense," they usually respond. Nonetheless, it is curious to note to what point agent relationships are ignored by those persons having managerial responsibilities in the majority of businesses in any industrial or service field. We use this statement by the author to introduce the fundamental purpose of this article. Probably, once having read it, any professional would have a clearer idea about such important concepts (in order to realize how costs function and how they are generated inside a health organization) as uncertainty, agent relationships or asymmetrical information. Using a British Airways flight as a comparison, it will be difficult not to understand these terms in their practical applications to the health field. PMID- 10514751 TI - [Home enteral nutrition]. AB - Enteral nutrition in the home is applied to stabilized patients who do not require hospitalization or to chronically ill patients who can stay in their homes. However, ensuring the correct administration of this treatment requires a coordinated, expert multidisciplinary team. This article reviews the conditions for use of enteral nutrition in the home, the means of access, the nutritional formulas, the administrative technique, and the complications enteral nutrition in the home may present. Furthermore, the composition and characteristics of the multidisciplinary team which will be in charge of carrying out this treatment is discussed. PMID- 10514752 TI - [Administrative perspectives at Sarasota Memorial Hospital]. AB - Sarasota Memorial's Health VISION project goal is to develop an electronic patient record. This requires total reengineering of health care delivery in Sarasota county to provide optimal exchange of aggregate, educational, cost and patient information to hospitals, clinics, offices and homes. PMID- 10514753 TI - [Work and maternity]. PMID- 10514755 TI - [Interior of the shoulder joint. Lateral aspect]. PMID- 10514754 TI - [Necessity of special education in social abilities and communication techniques during nursing education]. PMID- 10514756 TI - [Effect of music on children with cancer]. AB - This study looked for a relationship between immunity and one's spirits while investigating the effect musical therapy produces on Immunoglobulina A (IgA) found in saliva and a Patient's Opinion on the Likert Scale (OPEL). There were 30 children as patients, 15 in a control group and 15 in an experimental group. They were 5 or 16 years of age and checked into the Sant Joan de Deu Hospital in Barcelona due to neoplastic illness. PMID- 10514757 TI - [The patient's privacy and the treatment of intimacy: an ethical reflection]. AB - Although no one today doubts that speaking of a nurse's patient care presupposes the nurse will carry out her/his role as a professional; nonetheless, this has been related to the patient's intimacy very few times. This article concerns this last perspective since the author maintains as her hypothesis that the sharing of myths and images people hold, as an essential element of each person, requires nurse's care. For this reason, the author presents the concept of the intimacy of the patient from an ethical point of view and defines the principles which compose the treatment of intimacy. PMID- 10514758 TI - [Dorothea E. Orem: thoughts on her theory]. AB - Dorothea E. Orem's theory of nursing, based on the key concept of self-care, carries a particular way of viewing the reality of nursing treatments. According to this author, every individual adult has the capacity for self-care; however, when a health problem arises it is possible that this capacity is insufficient to confront the situation, making it then necessary to receive help from other persons who compensate for this deficit. PMID- 10514759 TI - [Retinopathy in the newborn. Nursing care]. AB - The use of oxygen as treatment for cardiorespiratory pathology in newborns is an accepted, regular practice. The prolonged use and high concentration of oxygen in premature newborn infants has been associated with an increase in cases of Proliferate Retinopathy in Premature Infants. This article publishes the results obtained over the past two years from the application of a treatment procedure which followed the recommendations made by the Standards Commission of the Spanish Association of Pediatricians regarding the measurement, administration and monitoring of oxygen to at risk newborn infants while evaluating the number of cases of retinopathy and their severity. Included in this study are all the newborns attended to at the Neonatal Intensive Care Unit of the "Arquitecto Marcide-Novoa Santos" Clinical Hospital in Ferrol from March 1995 to February 1997 which weighed less than 1500 grams or were born before the 36th week of gestation and received oxygen at a concentration above 50%. We encountered one case of level 3 retinopathy. In this case, we noticed a significant relationship among the point of gestation completed at birth, weight at birth and the severity of Proliferate Retinopathy in Premature Infants. We also proved that a painstaking control of oxygen allows a medical team to lower the degree of retinopathy in premature infants. PMID- 10514760 TI - [Great challenges for the editorial world: where went our pleasure in reading? May we photocopy indiscriminately?]. PMID- 10514761 TI - [Uncertainty, relationship with agencies and asymmetrical information (II)]. AB - In the second part of this article, the author relates the "knightian" uncertainty to consumer, i.e. patient, preferences and to types of choice, considering that choice in a health organization usually is made based on uncertain, or high risk, expectations whose determination is often difficult. The author reviews the concepts of 1) "ethical risk"--or potencial overuse of a form of treatment when the consumer does not bear its cost directly--, 2) "asymmetrical information", and 3) "adverse choice"; all of which are related to relationships with an agency and therefore, have the corresponding costs derived from these agencies themselves. Finally, the author reviews the "agency theory" and, in relationships to the agency itself, evaluates the "agency costs", costs which are not always evaluated in a just manner. The author make use of the same flight comparison, British Airways flight number 1, as he did in the first part of this article to bring these concepts into health field and to provide a clearer description of these concepts. PMID- 10514762 TI - [Cardiopulmonary resuscitation. Support of circulation]. AB - Whether the cause is primary or secondary, cardiac arrest requires cardiopulmonary resuscitation techniques in order to avoid brain lesions. Cardiac compressions combined with the control of the air passageway and artificial respiration have made it possible to reverse clinical death status in a patient. In this article, the authors review various proven methods including heart pump, thoracic pump, asynchronous respiration, abdominal contrapulsation, cardiopulmonary resuscitation by means of coughing, MAST, ADC, pneumatic vest, etc. which are currently in use. PMID- 10514763 TI - [The World Day of Tobacco: "the decision has been made, I reliquish tobacco". The World Health Organization promotes the World Day Without Tobacco on the 31. May since 1998]. PMID- 10514765 TI - [Aging as a novelty]. PMID- 10514764 TI - [Addicted patients. Attitude of nursing personnel]. AB - Inside the realm of therapeutic relationships, one recognizes the importance of the nurse's attitude towards patients as one of those elements bearing influence on the objectives desired by the nursing profession. This study compares the attitudes towards via parenteral drug addicts held by nursing students and non specialized nurses. By means of a pilot study which used a selfadministered Litcker scale questionnaire developed by the study authors, two samples were selected: a student sample (n = 40) and a non specialized nurse sample (n = 40). These variables were evaluated in the two samples: age, professional experience, and the frequency in sporadic care for via parenteral drug addicts. The conclusion drawn from this study is that the attitude of nursing students is significantly more positive towards via parenteral drug addicts than the attitude held by non specialized nurses. The adoption of a more negative attitude towards these patients occurs at the start of a nurse's professional career. PMID- 10514766 TI - [Detection of tuberculosis in HIV-positive patients]. PMID- 10514767 TI - [Goals and objectives in environmental health]. PMID- 10514768 TI - [Ethical and legal questions in psychiatric nursing]. PMID- 10514769 TI - [Culture and nutrition. Dietetics in hospital nursing during the 17 century]. AB - Starting from the important role which feeding and dietetics play in our culture and specifically in activities such as medicine and nursing, this article analyzes aspects like 1., the place of dietetics and feeding found in two manuals, "Instructions for nurses to apply remedies to all kinds of illnesses" and "To attend to many accidents which in the absence of Doctors"; 2., the importance of diets in hospitals throughout the 17th century; 3., the work by nurses related to diets and alimentation during that epoch and the professional training received by these professionals in this regard. PMID- 10514770 TI - [Enteral nutrition in the hospital]. AB - The author presents an interesting historical journey documenting the search for solutions to feed patients who were not capable of feeding themselves by conventional means. Patients deemed at risk nutritionally are analyzed, along with the means of detecting them. The characteristics of enteral nutrition plus its most important indications and counterindications are discussed. Mention is also made of the important role of nurses in hospital care, in the types of feeding patients receive, and in the form of administering this feeding. PMID- 10514771 TI - [Radial and cubital deep arteries]. PMID- 10514772 TI - [Anxiety and success in nursing exams]. AB - This article presents an evaluation about anxiety and its influence on the degree of success nursing students have on their exams. The influence of anxiety on cognitive processes under situations perceived as threatening, as well as the perception of physiological symptoms, are analyzed. This study may be considered to be a replica of one carried out by Sarason and Stoops in 1978. PMID- 10514773 TI - [Elections and health]. PMID- 10514774 TI - [Uncertainty, relations with agencies and asymmetric information (III)]. AB - In this the third part of the article on uncertainty, agent relationships and asymmetrical information, Doctor Lara passes through Reykiavik and London on her to her final description in Seville. She takes advantage of this analogy to conceptualize the problem of principal and agent, analyzing generic agency relationships pertaining directly with health organizations. When dealing with the interesting question if the problem of principal and agent in health organizations can be lessened, a few equally interesting answers were formulated: allow the professionals to assume the mission and the values of the organization; a well thought out leadership policy; and improvement in information provided; an incentive policy; compatibility of interests between the principal and the agent; ethics and competitive advantage; quality management.... PMID- 10514775 TI - [Pain and respiratory physical therapy in patients after heart surgery]. AB - Between the months of March and June 1997, an extended observational study was carried out on 32 patients hospitalized in the intensive care unit of "Hospital General Universitario" in Alicante for postoperative care after undergoing heart surgery which included extracorporeal circulation. The variables analyzed were of a socio-demographical nature and related to the type of operation in order to homogenize the samples. The main variables under study were pain while resting and pain while carrying out these exercises: coughing, deep breathing, stimulated inspirometer, and clapping; the number of balls the patient could raise; collaboration with the physical therapist and the kind of analgesic administered. The pain measuring instrument was the visual analogous scale. The patients with the highest pain levels when performing the protocolary extubation exercises raised fewer balls in the stimulated inspirometer exercise and did not collaborate with the physical therapist. The patients who receive more doses of sedatives collaborate with the respiratory physical therapist. All those patients who receive an analgesic at a set hour raise one or two balls in the inspirometer exercise and all collaborated with the physical therapist. PMID- 10514776 TI - [Calculation of body fat by skinfold thickness]. AB - This paper describes the method of determining the sizes of the subcutaneous fat folds found in the biceps, triceps, subscapular, suprailiac and submandibular. One is shown how to make the calculation of body fat and the areas of the section of the arm as related to the sizes of the folds and, finally, the author provides some recommendations so that one may correctly evaluate the data obtained. PMID- 10514777 TI - [Administration of propofol. Addition of lidocaine eases the pain]. AB - This article corresponds to the final paper accepted as passing as a requirement to accede to the level of 500 hours in the "IDER" Masters program "Methodology of Research and Investigation in Healthcare" as part of a correspondence course designed by the University of Alcala with backing from the University of Antioquia in Colombia. The pain which occurs as a response to an intravenous injection of Propofol is one of the main obstacles to its use as an anesthetic. The wide range of uses of propofol increases the desire to minimize this inconvenience. This study compares the occurrences of pain and the injection of propofol in three groups: the first group was administered propofol without any additives; the second group was administered propofol along with 20 mg of lidocaine; the third group had 40 mg of lidocaine added to the calculated doses of propofol. This study concluded that the addition of lidocaine significantly reduced the pain felt in the circulatory tract selected for the injection of propofol. Administering 20 or 40 mg of lidocaine did not produce any differences in the analgesic effect of this anesthetic. PMID- 10514778 TI - [IDER improves education about severe burns. Barcelona was the scene of a course on "Nursing care for the burn patient"]. PMID- 10514779 TI - [Midwives and conscientious objection. The case at the Son Dureta Hospital in Mallorca]. PMID- 10514780 TI - [Ligaments of the left wrist and metacarpals (palmar aspect)]. PMID- 10514781 TI - [Role of the family in the care of an elderly person with cerebrovascular disease]. AB - This study tries to identify the roles family members have in the care of an elderly person experiencing the effects of cerebral-vascular disease in the home and the main strategies to employ in the treatment of such a patient. The theoretical methodological procedure was based on a psychological project, a historical drawing about a theme, in which five subjects who were caretakers drew two achromatic pictures narrating corresponding stories individually in their homes. This form of expression allowed the project designers to have an interpretation of the daily situations involved in the care of these patients. In the first analysis phase, we drew the conclusion that caretakers construct their own particular knowledge system regarding the care of the elderly patient in their home. We verified that this construction started with a network relating common, daily elements which are interwoven in the treatment process occurring within a specific social milieu which as a result generates strategies that rely heavily on common sense knowledge in the manner of carrying out this social function. PMID- 10514782 TI - [Transportation of newborns in critical condition]. AB - The transportation of a newborn baby in critical condition bears a set of important peculiarities, characteristics and problems which noticeably distinguish it from the transport of adults. The health teams which transport newborns in critical condition from hospital to hospital must possess adequate knowledge, be experts in the care of newborns, and have mobile assistance units equipped with all the materials necessary to be able to provide their pediatric patient with a quality care similar to that in an intensive care unit. PMID- 10514783 TI - [Evaluation of clinical competence. The immediate future of nursing]. AB - In this article, the authors analyze the concept of professional competence and they describe some of the methods destined to be used in the evaluation of said competence in nursing professionals. The authors describe different levels of professional performance to which an evaluation may be carried out according to the conceptual model developed by Miller. Special emphasis is placed on the third level of evaluation which is based on simulations of professional practice whose protagonists are the so called simulated or role-playing/standardized sick. One type of very novel evaluation test is the Objective Structured Clinical Examination (OSCE) which makes use of, besides the simulated or role playing/standardized sick, other mediums such as manikins, standardized tests, open-ended short answer questions, etc. This is one of the most complete clinical competence evaluation methods currently available, although one should not forget that this evaluation is carried out in a simulated, laboratory, setting. PMID- 10514784 TI - [Preparing for Utopia]. PMID- 10514785 TI - [Uncertainty, relationships with agencies and asymmetric information (IV)]. AB - This last section ends Dr. Ruiz's article in which he conceptualized terms such as uncertainty, principal and agents, information cost, superior and inferior assets, induced demand, consumer preferences,... He has also presented agent relationships inside health organizations and the tolls suggested to lessen the problem of principal and of agent. This last section is dedicated to a deep analysis of one of the aforementioned tools: improving information. At the same time, an appendix which summarizes the concepts discussed throughout the four sections of this article is provided. PMID- 10514786 TI - [Self care in adolescents]. AB - The purpose of this work is to offer an analysis on the adolescence as stage of the life with some specific characteristics due to the transformations that happen so much at biological level as cognitive and psychosocial. During this period, the adolescent develops their autonomy and by so much starts their independence of the parents, some and other will experience situations that them will make be felt insecure in their/its performances until each one assume their new paper in the familiar environment and before itself. Parents and adolescent can need support and advice to obtain that this stage, in conflicting occasions, is developed normally, so that the adolescent become a capable adult about caring whether same and about others, with a life style that favor the integrated operation and the continued development. The family nurse can lend them the support and advice that need, since occupies a privileged place in the health equipment to guarantee the continuity of the assistance to the familiar group from the birth until the maturity. Basing us on the theory of the self-care developed by D. Orem, we will check the specific requirements of self-care of this stage of the vital cycle, of great transcendency for the step to the adult age. PMID- 10514787 TI - [The patients' silence]. AB - We cannot speak about care nor cure without realizing that the concept of health care is designed to aid all the needs a patient has. And surely, only by placing ourselves in the patient's spot, trying to live inside his/her skin for a few minutes, we can comprehend the complexity of the situation which surrounds him/her. This is the way to understand, from all angles, that all efforts must be coordinated to attend to each patient's physical and emotional needs. PMID- 10514788 TI - [Cutaneous nerves of the right upper extremity]. PMID- 10514789 TI - [Modified states of consciousness: the profound reconciliation with life]. PMID- 10514790 TI - [Treatment of addicts with various substances]. PMID- 10514791 TI - [Coloplast presents a new dressing for colostomies]. PMID- 10514792 TI - [Clinical evaluation of a hydro-cellular dressing for the treatment of venous leg ulcers]. AB - INTRODUCTION: Varicose ulcers (UV) are a serious health problem which produce tremendous pain for those who suffer from them; furthermore, they consume a large amount of health care system resources. Hydro-cellular dressings provide a moist environment cure option as a local treatment of varicose ulcers. MATERIALS AND METHODS: With the purpose of evaluating the use of an Allevyn non-adhesive hydro cellular dressing under real clinical conditions, an open, observational multi centric study was carried out. The study group was composed of outpatient care patients or patients hospitalized due to having varicose vein ulcers or having a mixed etiology with an ankle/arm index less than or equal to 0.8. MAIN RESULTS: 24 patients were initially part of this study; two of these abandoned this study group without a justified reason. Of the 22 who finished this study, 16 or 72.7% were women while 6 or 27.3% were men. The mean age in the women patients was 73.07 +/- 9.31 (DS), while among the men patients, the mean age was 67.23 +/- 18.17 (DS). 17 or 77.3% of the ulcers studied were varicose, 1 or 4.5% had a mixed etiology, 1 or 4.5% was due to amputation, and in three cases, the kind of ulcer was not specified. 22 patients finished this study with their lesions healed or in the process of healing; 2 abandoned this study without justification. The patient study group passed from an average initial sore area (IC 95%) or 9.41 or 19.42 cm2 to a final average sore area (IC 95%) of 4.45 or 10.5 cm2 with a mean percent reduction of (IC 95%) 57.22 or 86.22%. In 7 cases or 31.8% of the patients, their lesions healed completely during the course of the study. Inside the group of patients whose lesion did not fully heal during this study, the average initial sore area (IC 95%) was 15.66 +/- 12.75 cm2, a mean of 8.6 or 22.2 cm2 and the medium equaled 9 cm2. The information regarding comfort, absorption quality, ease in application and removal, and its fine results in the presence of pain and in the creation of new lesions once the dressing has been removed as well as the absolute lack of adverse reactions indicates we have discovered an excellent therapeutic option. PMID- 10514793 TI - [The concept of nursing. A systematic analysis]. AB - Nursing is a field which employs numerous people whose formation and professional roles are regulated by a wide set of rules and norms, and one which benefits innumerable patients. A paradox exists when, upon consulting various dictionaries, one of these the Royal Academy Dictionary (Diccionario de la Real Academia), the term nursing is defined as a physical space/set of nurses without any reference whatsoever to nurses' role in the health and educational systems. The definition we find in a dictionary should correspond to the concept the general public holds and reflect the true meaning of the profession. During a three year period, academic years 1994-96, on the first day of class, we asked each student to define or describe what he/she understood nursing to be. We consider that their responses, in a large sense, should correspond to the idea which the general public holds about nursing. An analysis of the content of these definitions allowed us to establish that there is a clear confrontation between what dictionaries state and what students, as a sample of society as a whole, think. Our results permit us to offer a definition for nursing which may be incorporated into new dictionary editions for the purpose of completing the already existing ones. PMID- 10514794 TI - [Genetic maternal-child identification with DNA]. AB - BACKGROUND: Classical methods for newborn identification cannot establish a true biological nexus between mother and newborn, and hence they have been widely criticized. Therefore, a pilot study on a mother-infant genetic identification program (PROIGMI) has been started in order to ensure the determination of a biological relationship between mother and newborn in cases of vaginal delivery, caesarean birth or fetal autopsies. MATERIAL AND METHOD: In the delivery room and after informed consent, a total of 100 blood samples from mother/newborn couples were taken and deposited on clean and sterile paper supports. DNA from a total of 20 mother/newborn couples was studied by PCR techniques, being able to unequivocally establish the biological relationship in all cases, even when using minimal amounts of DNA. RESULTS: Blood samples collection does not show differences regarding the kind of birth (delivery, cesarean). The protocol used is easy and fast, and does not employ materials not known for health care professionals. Minimal amounts of blood yield enough DNA to obtain conclusive inclusion probabilities. CONCLUSIONS: The use of DNA allows to stablish the so called biological truth in cases of doubt or where necessary; with the use of medical protocols these studies can be completed in 6 to 8 hours using small amounts of DNA (5 microliters). PMID- 10514797 TI - Calling 'em like he sees 'em. Interview by Chris Serb. PMID- 10514795 TI - [Nursing diagnosis. Keys for its development]. AB - The authors carried out a study of the taxonomy used at the 1990 NANDA Conference. A few of the authors findings were some diagnostic labels were found to be imprecise; some of the diagnostic labels relate to causes which would only allow nurses to work in collaboration with other medical team professionals; other labels described symptoms which were hard to define objectively or were non verifiable, imprecise and vague defining the problem. As a conclusion to this study, the authors contend that in order to develop nursing diagnostics it is necessary to validate the existing labels, develop new labels, make diagnostic tests serve as an appraisal and detection instrument, and to develop the critical symptoms necessary for identifying diagnostic labels. This study is based on a paper presented at the "First International Symposium on Nursin Diagnostics" which was celebrated in Barcelona. PMID- 10514798 TI - New disasters, new plans. PMID- 10514799 TI - 1999 salary survey. Is this the future of your bonus? AB - As pay trends go, one executive's bonus can be another's bane. Asked what they like about their compensation programs, health care execs and trustees say they're satisfied with the base salaries paid to their top people, but not with their bonus plans, especially for meeting long-term goals like stronger credit ratings and asset positions. PMID- 10514800 TI - A sharper point on using safer needles. AB - National legislation is just one sign of changing attitudes about safety needles. Fifteen states are considering similar bills; five already have passed such laws. OSHA also says it will put needlestick injuries on its agenda this fall. Yet if needlesticks and efforts to prevent them are nothing new, what's causing this change of heart? Two things: improved technology and media attention. PMID- 10514801 TI - Deep & Wide. Health care boards look beyond their communities for new trustees. AB - Trend watchers stop short of calling it a groundswell, but broader board membership is gaining support. "Traditionally, governance on behalf of a community has meant governance by the community," says one consultant. "Changing circumstances are challenging that convention." Driving the trend are changes often initiated by trustees themselves. PMID- 10514802 TI - Deselected! A California case could forever change how HMOs and others hire and fire doctors. AB - Deselection--a bit of health insurance parlance capable of giving doctors a case of the shivers--may never be the same. An obscure California lawsuit, barely known outside legal circles, threatens to change the way physicians are hired and fired from provider panels. And that's making HMOs and others nervous. PMID- 10514803 TI - Questions before I go. PMID- 10514804 TI - New waves of research in the epilepsies: crossing into the third millennium. PMID- 10514805 TI - Genetic heterogeneity and epidemiology of the epilepsies. PMID- 10514806 TI - Neural development, genes and the epilepsies: introduction. PMID- 10514807 TI - Homeobox genes in development. PMID- 10514808 TI - Cortical dysplasias, genetics, and epileptogenesis. PMID- 10514809 TI - The relevance of slight migrational disturbances (microdysgenesis) to the etiology of the epilepsies. PMID- 10514810 TI - The role of neural activity in synaptic development and its implications for adult brain function. AB - In much of the developing nervous system, electrical activity guides the formation of neural connections, with lasting effects on adult brain function. Epilepsy, a defect in neuronal excitability, might result from abnormal patterns of activity in the young brain. Many connections are organized by selective stabilization of synapses when they are activated simultaneously on the same postsynaptic cell during a sensitive period in early life. This process often involves calcium entry through the N-methyl-D-aspartate (NMDA) subtype of glutamate receptor. The magnitude of the current passed by this receptor depends on its subunit composition, which varies with age and brain region. Although receptor configurations that admit large calcium currents are permissive of synaptic plasticity, they also increase neural vulnerability to excitotoxic cell death. In most regions of developing brain, activity that can drive NMDA receptors initially is low and increases with maturation. Thus, the replacement of NMDA receptors that flux large calcium currents during early periods of synaptic organization with NMDA receptor subtypes that flux less calcium as synapses become more active, more effective, and less plastic allows maturing neurons to maintain optimal levels of intracellular calcium in the face of drastic developmental changes in their inputs. We have proposed that this transition in NMDA receptors from high to low calcium permeabilities is itself activity dependent. This idea is supported by data showing that many synaptic proteins, including receptor subunits, can be regulated by activity. Cultured cerebellar granule neurons require NMDA receptor stimulation for survival and differentiation, which may replicate the activation provided by the arrival of mossy fiber innervation in vivo. In these cultures, chronic depolarization and glutamate or NMDA treatment induces more mature NMDA receptor subunit expression patterns and function and also increases the expression of several gamma aminobutyric acid type A (GABAA) receptor subunits, changing that receptor's function. In addition, evidence from in vivo studies indicates that synaptic maturation itself may depend on NMDA receptor activity. During the formation of topographic connections between the retina and superior colliculus (SC) of young rats, chronic local application of the competitive NMDA receptor antagonist +2 amino-5-phosphonovalerate (D-APV) blocks the normal developmental up-regulation of NMDA receptor subunit 1 (NR1) mRNA and nitric oxide synthase activity, as well as maturation of calcium and calmodulin-dependent kinase distribution, activity, and substrate phosphorylation. Together, these recent molecular findings suggest that chronic seizure disorders could result from any of a variety of early developmental events. Any disturbance that locally perturbs regulation of NMDA receptors or the temporal correlations in synaptic activity that drive these receptors has the potential to alter the normal development of local circuitry and the critical balance of inhibition and excitation required to contain seizure activity. PMID- 10514811 TI - Ontogeny of channels, transmitters and epileptogenesis. PMID- 10514812 TI - Age-dependent vulnerability to seizures. AB - Seizure disorders frequently occur early in life. Seizures are classified as reactive, symptomatic, or idiopathic depending on whether their cause can be identified. Reactive seizures are the result of acute environmental perturbations. Early in life, many stressors can produce seizures and the ultimate outcome may depend on the particular precipitating factor and its intensity. Febrile convulsions are the most common reactive seizures, although they must be differentiated from symptomatic seizures precipitated by fever. Symptomatic seizures are often associated with varying degrees of central nervous system (CNS) insults, including congenital malformations and metabolic storage diseases of the gray matter. These seizures may have age-specific characteristics and may at times be difficult to treat with conventional antiepileptic treatments. To develop a better understanding of the pathophysiology of seizures early in life, we have extensively used animal models of epilepsy. In this chapter, we report our findings with a rat model of developmental cortical dysplasias produced by intrauterine injections of methylazoxymethanol acetate. These rats are more susceptible to kainic acid, flurothyl, and hyperthermic seizures than normal rats. Rats with severe cortical dysplasia are most susceptible to seizures. We have also studied the mechanisms involved in the control of seizures during development because status epilepticus is more prevalent in infants than in adults. Our data suggest that the substantia nigra may play a crucial role in status epilepticus as a function of age. In the adult substantia nigra two regions mediate opposing effects on seizures following infusions of gamma-aminobutyric acid type A (GABAA) agents. One region is located in the anterior substantia nigra, and muscimol infusions in this region mediate anticonvulsant effects. The second region is in the posterior substantia nigra, and here muscimol infusions produce proconvulsant effects. In situ hybridization data demonstrate that, at the cellular level, neurons in the two substantia nigra regions differ in the amount of hybridization grains for GABAA receptor alpha 1 and gamma 2L subunit mRNAs. In developing male rats, only the "proconvulsant" region is present up to the age of 21 days. The transition from the immature to mature substantia nigra mediated seizure control occurs between age 25 and 30 days. The identification of age-dependent functional networks involved in the containment of seizures may lead to possible new pharmacologic strategies to control seizures, thus aiding the development of age-appropriate treatments of seizure disorders. PMID- 10514813 TI - Nicotinic receptor: a prototype of allosteric ligand-gated ion channels and its possible implications in epilepsy. PMID- 10514814 TI - GABA is the principal fast-acting excitatory transmitter in the neonatal brain. AB - gamma-aminobutyric acid (GABA) is the principal neurotransmitter of inhibition in the adult mammalian brain. However, at early stages of development, including the embryonic period and first week of postnatal life, GABA plays the role of main neurotransmitter of excitation. The paradoxical excitatory effect of GABA is caused by an inverted chloride gradient and, therefore, a depolarizing direction of GABA type A (GABAA) receptor mediated responses. In addition, another type of GABAergic inhibition mediated by postsynaptic GABA type B (GABAB) receptors is not functional at early stage of life. In the neonatal rat hippocampus, GABA, acting via GABAA receptors, activates voltage-gated sodium and calcium channels and potentiates the activity of N-methyl-D-aspartate (NMDA) receptors by reducing their voltage-dependent Mg2+ block. The temporal window when GABA exerts excitatory actions coincides with a particular pattern of activity of hippocampal neuronal network that is characterized by periodical giant depolarizing potentials (GDPs) reminiscent of interictal-like epileptiform discharges. Recent studies have shown that GDPs result from the synchronous discharge of GABAergic interneurons and principal glutamatergic pyramidal cells, and they are mediated by the synergistic excitatory actions of GABAA and glutamate receptors. GDPs provide synchronous intracellular Ca2+ oscillations and may, therefore, be implicated in hebbian modulation of developing synapses and activity-dependent formation of the hippocampal network. PMID- 10514816 TI - Introduction to the idiopathic epilepsies. PMID- 10514815 TI - Developmental neuroplasticity: roles in early life seizures and chronic epilepsy. AB - Both clinical and experimental studies suggest that the immature nervous system is unusually susceptible to seizures during critical periods in postnatal life. A late onset of gamma-aminobutyric acid (GABA)-mediated synaptic inhibition could conceivably play a contributing role in this phenomenon. Numerous studies have shown that neural systems that use GABA in the neonatal brain are different than those of adulthood. GABA is an excitatory neurotransmitter that likely plays a neurotrophic role in neuronal differentiation. Other reports suggest that unique, possibly transient, GABAergic interneuron populations exist in the embryonic and neonatal nervous system. At these early times in development, the immature nervous system is remarkably resistant to seizure generation. However, as the hippocampus and neocortex enter the critical period of enhanced seizure susceptibility, inhibitory GABA systems mature rapidly. At this time, blockade of GABA type A (GABAA) receptors produce unusually severe seizure discharges. In hippocampus, concurrent exuberant outgrowth of recurrent excitatory axon collaterals and synapses appear to play a role in the generation of these seizures. As the hippocampus matures, these axons are morphologically remodeled and nearly 50% of branches within arbors are pruned. This pruning of axon branches corresponds in time with the decrease in seizure susceptibility that characterizes adulthood. Developmental remodeling of neuronal connectivity is a common feature of most areas of the central nervous system. Results from an audiogenic seizure model of early onset epilepsy suggest that prevention of axon arbor remodeling by transient sensory deprivation can lead to a permanent overinnervation of target nuclei and chronic seizure susceptibility. Early life seizures may have a similar effect. Recent results in one model have shown that repeated seizures induced by intrahippocampal injections of tetanus toxin during a critical period results in a chronic epilepsy. Future studies should attempt to determine if the synchronized discharging of early-life seizures prevents the remodeling of neuronal connectivity that normally takes place during postnatal development and results in an overinnervated and chronically hyperexcitable hippocampus. PMID- 10514817 TI - Single-gene models of epilepsy. AB - Single-gene models of epilepsy present valuable opportunities to isolate and experimentally reproduce gene mutations for human seizure disorders, to test molecular mechanisms of epileptogenesis, and to explore strategies to correct early hyperexcitability defects in the developing brain. Although not all inherited epilepsies are monogenic, analysis of epileptic phenotypes in spontaneous and transgenic mouse mutants is beginning to define the kinds of molecular defects favoring inherited aberrant synchronization in central neurons. The range of genes identified shows that rather than arising from a few superfamilies that regulate membrane excitability, the gene products are drawn from many categories involved in widely diverse functions of the cell. Although some primary defects directly alter membrane electrogenesis and neurotransmitter signaling at synapses, others are too far removed from these processes to allow one to visualize the steps by which they promote epileptogenesis. There is now clear evidence that several and probably most epilepsy genes entrain specific patterns of secondary cellular plasticity during brain development. It can be predicted that these downstream rearrangements may partially account for the delayed temporal onset and other progressive features of epilepsy syndromes. Experimental alterations that target the mutant gene product and patterns of secondary network plasticity provide a basis for future strategies to reverse the epileptogenic process. PMID- 10514818 TI - Studies of the lethargic (lh/lh) mouse model of absence seizures: regulatory mechanisms and identification of the lh gene. AB - To understand the cellular and molecular mechanisms that underlie generalized absence seizures sufficiently well to design rational, efficacious new therapies for patients, it is necessary to turn to animal models to gain insights into these mechanisms. The lethargic (lh/lh) mutant mouse expresses spontaneous absence seizures that share behavioral, electrographic, and anticonvulsant profiles with absence seizures in patients. This validates its use to study the mechanisms that underlie absence seizures. This chapter discusses two scientific approaches that involve the use of lh/lh mice. The first part of the chapter discusses neurobiologic approaches used to investigate critical mechanisms that regulate the synchronized burst firing within the thalamocortical network that generates absence seizures. Two of these critical mechanisms have been studied in detail with lh/lh mice. The first critical mechanism involves the required activation of gamma-aminobutyric acid B (GABAB) receptors to generate absence seizures. Because the numbers of GABAB receptors are increased in thalamocortical populations among lh/lh mice compared with littermates without epilepsy, these receptors appear to play a pathophysiologic role in the expression of absence seizures among lh/lh mice. Moreover, there may be a role for GABAB receptors in the generation of absence seizures among humans, because administration of compounds that activate GABAB receptors can produce absence seizures among humans. These findings suggest that GABAB receptor antagonists may represent a new class of antiabsence compounds that will be efficacious against absence seizures among patients. A second critical mechanism that regulates generation of absence seizures involves GABAA receptors in the nucleus reticularis thalami (NRT), a nucleus that sends GABA-ergic afferents to thalamic relay nuclei. Activation of GABAA receptors in the NRT appears to suppress the generation of absence seizures among lh/lh mice and in other models. Moreover, clonazepam may exert its antiabsence actions through this mechanism. Together, these findings suggest that compounds that selectively activate GABAA receptor isoforms expressed in NRT may represent a class of antiabsence drugs that could have fewer side effects than compounds currently used to treat patients. The second part of the chapter discusses a molecular genetic approach to delineation of the mechanisms that underlie absence seizures. Absence seizures among lh/lh mice are caused by a single-gene defect on chromosome 2. If positional cloning and gene isolation techniques are successful, it will be possible to identify the lh disease gene. Subsequent studies of the lh gene product should greatly increase not only our understanding of the pathophysiologic basis for absence seizures among lh/lh mice but also our ability to seek similar mutations in homologous genes in human families that express absence seizures. Accordingly, strategies and progress in cloning and identifying the lh disease gene are presented. PMID- 10514819 TI - Absence epilepsy: advances in experimental animal models. PMID- 10514820 TI - Experimental models of multifactorial epilepsies: the EL mouse and mice susceptible to audiogenic seizures. AB - This chapter reviews two well-characterized mouse epilepsy models with a multifactorial etiology, the epileptic EL mouse and mice susceptible to audiogenic seizures (AGS). Multifactorial disorders are quantitative traits where the action of more than one gene together with environmental factors contributes to the disease phenotype. The EL (epilepsy) mouse has been studied extensively as a genetic model for idiopathic complex partial seizures in humans. EL seizures are associated with an intense hippocampal gliosis in the absence of obvious neuronal loss and an elevated calcium-dependent release of aspartate that is present both before and after seizure onset. The inheritance of epilepsy is complex and several seizure frequency quantitative trait loci (QTL) have been mapped. Much of this genetic complexity may arise from the influence of environmental factors, including the seizure testing procedure, seizure history, and age. AGS, which are violent sound-induced convulsions, are considered a genetic model for generalized brainstem or reflex epilepsies. AGS susceptibility can arise as an inherited trait in some mouse strains or can be induced in genetically resistant strains from environmental factors (e.g., prior acoustic stimulation). AGS susceptibility and long-term potentiation (LTP) may also share common mechanisms. Several Asp genes have been mapped that influence AGS susceptibility. The expression of one of these can be modified by genomic imprinting and another has been identified as the X-linked 5-HT2e serotonin receptor. The genetic dissection of convulsive behavior in EL and AGS susceptible mice could help identify candidate genes for human multifactorial epilepsies. PMID- 10514821 TI - Transgenic approaches to epilepsy. AB - Transgenic animal models can reconstruct cellular, biochemical, and molecular alterations associated with human diseases and may help identify key disease mechanisms. Gene disruption, which results in the loss of the functional gene product, is the most common genetic alteration generated by transgenic approaches. Gene disruption can be introduced by random transgene integration or by gene targeting. Both of these approaches have resulted in mice that display recurrent seizures reminiscent of epilepsy. Here, we briefly describe the techniques of random transgene insertion and gene targeting and discuss how these methods were used to generate the epileptic jerky and 5-hydroxytryptamine (5 HT2C) receptor deficient mice. We also analyze how these mutants can contribute to our understanding of the molecular and cellular mechanisms underlying epileptic seizures. PMID- 10514822 TI - Intrinsic properties of reticular thalamic neurons relevant to genetically determined spike-wave generation. PMID- 10514823 TI - Neuronal networks in the genetically epilepsy-prone rat. AB - It is now possible to develop a dynamic neuronal network model for generalized convulsive seizures because of in vivo data recently obtained in a naturally occurring epilepsy model--the genetically epilepsy-prone rats (GEPR-9s). GEPR-9s exhibit audiogenic seizures (AGS) that consist of a sequence of discrete behavioral phases (i.e., wild running, clonus-tonus, and post-ictal depression). The neuronal firing changes in most nuclei implicated in the network during each phase of AGS in behaving GEPR-9s have been examined. The inferior colliculus is critical in AGS initiation, because extensive firing increases in inferior colliculus are observed preceding seizure initiation. The deep layers of superior colliculus (DLSC) are crucial to wild running, based on the emergence of tonic firing of DLSC neurons just preceding this phase. The pontine reticular nucleus (PRF) and periaqueductal gray (PAG) are critical to the clonic-tonic phase, because tonic firing patterns appear in these neurons just prior to this phase. During post-ictal depression all areas except the PRF are quiescent. These temporal relationships suggest that each nucleus plays a specific hierarchic role in each discrete convulsive behavior. Generalized tonic-clonic seizure behavior observed in human epilepsy, in GEPR-9s, and in other seizure models is likely to involve similar neuronal network components. The neurotransmitter mechanisms subserving the abnormal neuronal responses in the GEPR-9 neuronal network involve an increased availability of glutamate and a decrease in the effectiveness of gamma-aminobutyric acid (GABA) in many brain regions. Focal modification of the effects of GABA, glutamate, norepinephrine, or serotonin also modulates the nuclei of the network differentially. Together, these data reveal the anatomic, neurotransmitter, and neurophysiologic mechanisms of the neuronal network hierarchy in GEPR-9s, which is currently the most completely developed of any generalized convulsive model. Differential effects of anticonvulsants on the AGS phases and concomitant differential modifications of neuronal firing are observed on neurons in these network nuclei. With nearly complete identification of the network nuclei, the differential effects of these anticonvulsant drugs on different aspects of neuronal firing in different brain sites indicate that this experimental approach can likely identify the most sensitive therapeutic target for these agents. This concept is potentially vital to developing the most selective treatment of different convulsive behaviors occurring in human epilepsy. The neuronal network for AGS does not require brain structures rostral to the midbrain for seizure expression. However, the forebrain is recruited into an expanded seizure network through AGS repetition ("kindling"), resulting in prolonged AGS, post-tonic clonus, and epileptiform electrographic cortical abnormalities. AGS kindling produces network expansion into medial geniculate body (MGB) and amygdala and involves neuronal firing increases in MGB. PMID- 10514824 TI - Genetic epidemiology and the search for epilepsy genes. PMID- 10514825 TI - Genes for rare idiopathic generalized epilepsies: BFNC. PMID- 10514826 TI - Mapping and positional cloning of common idiopathic generalized epilepsies: juvenile myoclonus epilepsy and childhood absence epilepsy. AB - Among the 40 to 100 million persons with epilepsy worldwide and the 2 to 2.5 million persons with epilepsies in the United States, approximately 50% have generalized epilepsies. Among all epilepsies, the most common are juvenile myoclonus epilepsy (JME) with 10% to 30% of cases, childhood absence epilepsy (CAE) with 5% to 15% of cases, and pure grand mal on awakening with 22% to 37% of cases. In the last decade, six different chromosomal loci for common generalized epilepsies have been identified. These include two separate loci for JME in chromosomes 6p and 15q. The epilepsy locus in chromosome 6p expresses the phenotypes of classic JME, pure grand mal on awakening, and possibly JME mixed with absences. Two separate loci also are present for pyknoleptic CAE, namely, CAE that evolves to JME in chromosome 1p and CAE with grand mal in chromosome 8q24. Pandolfo et al. from the Italian League Against Epilepsy have reported two other putative susceptibility loci for idiopathic generalized epilepsies, namely, grand mal and generalized spike waves 35l in chromosome 3p and generalized epilepsies with febrile convulsions, grand mal, JME, absences, and electroencephalographic spike waves in 8q24. This chapter reports on the debate concerning whether there may be two separate epilepsy loci in chromosome 6p, one in the HLA region and one below HLA. The chapter then discusses the progress made in our laboratories as a result of the Genetic Epilepsy Studies (GENES) International Consortium. We discuss (a) the 2 to 6 cM critical region for classic JME located some 20 cM below HLA in chromosome 6p, (b) the 7-cM area for pyknoleptic CAE that evolves to JME in chromosome 1p, and (c) the 3.2 cM area for pyknoleptic CAE with grand mal and irregular 3 to 4 Hz spike waves in chromosome 8q24. We discusses efforts underway to refine the genetic map of JME in chromosome 6p11 and the advances in physical mapping and positioning of candidate genes, such as the gamma-aminobutyric acid receptor gene, the potassium channel gene of the long-QT family (KvLQT), named KCNQ3, and the human homologue of the mouse jerky gene for CAE in chromosome 8q24 and JME in chromosome 6p11. PMID- 10514827 TI - Genetics of partial epilepsies. PMID- 10514828 TI - The molecular genetic bases of the progressive myoclonus epilepsies. AB - Among the epilepsies, the progressive myoclonus epilepsies (PMEs) form a heterogeneous group of rare diseases characterized by myoclonus, epilepsy, and progressive neurologic deterioration, particularly dementia and ataxia. The success of the Human Genome Project and the fact that most PMEs are inherited through a mendelian or mitochondrial mode have resulted in important advances in the definition of the molecular basis of PME. The gene defects for the most common forms of PME (Unverricht-Lundborg disease, the neuronal ceroid lipofuscinoses, Lafora disease, type I sialidosis, and myoclonus epilepsy with ragged-red fibers) have been either identified or mapped to specific chromosome sites. Unverricht-Lundborg disease has been shown to be caused by mutations in the gene that codes for cystatin B, an inhibitor of cysteine protease. The most common mutation in Unverricht-Lundborg disease is an expansion of a dodecamer repeat located in a noncoding region upstream of the transcription start site of the cystatin B gene, making it the first human disease associated with instability of a dodecamer repeat. Juvenile neuronal ceroid lipofuscinosis is caused by mutations in the CLN3 gene, a gene of unknown function that encodes a 438-amino-acid protein of possible mitochondrial location. Other forms of neuronal ceroid lipofuscinosis that occur as PME and Lafora disease have been mapped by means of linkage analysis, but the corresponding gene defects remain unknown. Sialidosis has been shown to be caused by mutations in the sialidase gene, and myoclonus epilepsy with ragged-red fibers is well known to be caused by mutations in the mitochondrial gene that codes for tRNA(Lys). How the different PME gene defects described produce the various PME phenotypes, including epileptic seizures, remains unknown. The development of animal models that bear these mutations is needed to increase our knowledge of the basic mechanisms involved in the PMEs. This knowledge should lead to the development of new and effective forms of therapy, which are especially lacking for the PMEs. PMID- 10514829 TI - Dentatorubral-pallidoluysian atrophy (DRPLA): clinical features and molecular genetics. PMID- 10514830 TI - Mitochondrial genes for generalized epilepsies. PMID- 10514831 TI - Parental imprinting and Angelman syndrome. AB - Angelman syndrome is an inherited disorder that includes severe mental retardation and epilepsy. Patients have no speech, puppet-like gait with jerky movements, hyperactivity, disturbed sleep, bouts of inappropriate laughter, a pronounced jaw, and widely spaced teeth. The syndrome results from deletion or mutation within maternal chromosome 15q11-q13. Considerable evidence suggests that the gene or genes responsible for Angelman syndrome are expressed only from the maternal chromosome 15, a situation known as parental imprinting. This epigenetic marking of certain regions of the parental genomes is characterized by parent-of-origin-specific allelic DNA methylation, allele-specific DNA replication timing, and physical pairing of the two chromosome 15 homologues. Imprinting is important for normal development, and its disregulation causes several human disorders. The epilepsy of Angelman syndrome has been studied and indicates a rather typical electroencephalographic abnormality with slowing and notched wave and spikes. Various types of seizures occur, usually including myoclonus and atypical absence. Variable severity among patients suggests potential molecular diversity in the genetic mechanism, possibly the involvement of more than one gene. Angelman syndrome can arise from the following molecular genetic defects: a deletion in 15q11-q13 that covers the Angelman gene or genes, mutations that alter imprinting, and paternal uni-parental disomy for the region. Another 20% or so of patients with clinical symptoms of Angelman syndrome have none of these three defects but are believed to have mutations in one or more genes in the region, and this may be familial. The UBE3A gene, which codes for the enzyme ubiquitin protein ligase involved in protein degradation and processing, has been found to be mutated in many but not all of patients with Angelman syndrome and can be considered a major Angelman candidate gene. Other potential candidate genes in the region include a cluster of three GABAA receptor subunits, which are involved in inhibitory synaptic transmission in the brain. The GABRB3 gene, which codes for the beta 3 subunit, is deleted in most persons with Angelman syndrome. The absence of this gene in mice causes craniofacial abnormalities and neurologic impairment with seizures. The exact role of UBE3A and GABRB3 in the syndrome and their imprinting status are under investigation. PMID- 10514832 TI - Symptomatic lesional epilepsies: introduction. PMID- 10514833 TI - Neuronal channels, receptors and transporters: molecular structure, gating, and pharmacology. Introduction. PMID- 10514834 TI - Molecular properties of brain sodium channels: an important target for anticonvulsant drugs. AB - The voltage-gated sodium channels that are responsible for action potential generation in central neurons are important targets for the actions of antiepileptic drugs. These channels consist of a complex of three glycoprotein subunits: a pore-forming alpha subunit of 260 kd associated noncovalently with a beta 1 subunit of 36 kd and disulfide-linked to a beta 2 subunit of 33 kd. The alpha subunit forms a functional voltage-gated sodium channel by itself, whereas the beta 1 and beta 2 subunits modulate channel gating. The beta 1 and beta 2 subunits also have immunoglobulin-like folds in their extracellular domains that are predicted to interact with extracellular proteins. The alpha subunit is comprised of four homologous domains containing six transmembrane alpha helices (S1 through S6) and additional membrane-associated segments (SS1/SS2). The S4 segments in each domain function as voltage sensors for voltage-dependent activation of the sodium channel. The S5 and S6 segments in each domain and the short SS1/SS2 segments between them form the pore of the channel. The intracellular loop between domains III and IV forms the inactivation gate, which folds into the pore and occludes it within 1 msec of channel opening. The activity of brain sodium channels in modulated by protein phosphorylation G proteins. Activation of muscarinic acetylcholine receptors in hippocampal neurons slows the inactivation of sodium channels and reduces peak sodium currents through activation of protein kinase C (PKC) phosphorylation of sites in the inactivation gate and the intracellular loop between domains I and II of the alpha subunit. Other neurotransmitters that activate the PKC pathway are likely to have similar effects. Activation of D1-like dopamine receptors in hippocampal neurons reduces peak sodium currents through activation of cyclic adenosine monophosphate (cAMP)-dependent protein kinase phosphorylation of sites in the intracellular loop between domains I and II. Modulation by PKC and cAMP-dependent protein kinase is convergent--phosphorylation of the inactivation gate by PKC is required before phosphorylation of sites in the intracellular loop between domains I and II can reduce peak sodium currents. Brain sodium channels are also modulated by G proteins. Activation of endogenous G protein-coupled receptors causes negative shifts in the voltage dependence of sodium channel activation and inactivation. Overexpression of G protein beta gamma subunits induces persistent sodium currents. Regulation of sodium channel function by these multiple pathways can produce a flexible tuning of electrical excitability of central neurons in response to neurotransmitters, hormones, and second messengers. The antiepileptic drugs phenytoin, carbamazepine, and lamotrigine inhibit brain sodium channels substantially at clinically relevant concentrations. Their inhibition of sodium channels is increased by depolarization because they bind preferentially to the inactivated state of the channels. This effect increases the inhibition of sodium channels in depolarized tissue at the center of an epileptic focus. Local anesthetics also inhibit sodium channels by preferential binding to the inactivated state. Site-directed mutagenesis experiments show that antiepileptic drugs and local anesthetics bind to a common receptor site in the pore of the channel that is formed in part by three critical amino acid residues in transmembrane segment S6 in domain IV. Mutations in these amino acid residues prevents preferential binding to the inactivated state and thereby greatly reduces the affinity for inhibition of sodium channels by these drugs. Knowledge of the structure-function relationships for drug binding at this receptor site may open the way to development of novel classes of antiepileptic drugs. PMID- 10514835 TI - Chemical modulation of sodium channels and GABAA receptor channel. PMID- 10514836 TI - Voltage-dependent activation of ion channels. PMID- 10514837 TI - Membrane properties and epilepsy. PMID- 10514838 TI - GABA receptor function and epilepsy. PMID- 10514840 TI - Glutamate receptor channels: a possible link between RNA editing in the brain and epilepsy. PMID- 10514839 TI - Ligand-gated channel: postsynaptic receptors and drug targets. AB - Rapid synaptic transmission depends on the activation of ligand-gated channels by neurotransmitters. The molecular characterization of these membrane proteins has provided a valuable framework for investigating basic synaptic mechanisms and for developing pharmacologic strategies for the manipulation of neural function. This chapter begins with a summary of present molecular knowledge about different classes of ligand-gated channels, including the acetylcholine-gamma-aminobutyric acid (GABA) receptor superfamily, the excitatory amino acid receptor superfamily, and purine receptors. The activation mechanisms of ligand-gated channels are discussed in terms of detailed allosteric models that involve conformational transitions coupled with the occupation of binding sites by ligands. In addition to providing a quantitative understanding of the postsynaptic aspects of synaptic transmission, these allosteric models have been used to clarify the action of drugs that serve as valuable tools for the investigation and management of epilepsy. Relating these physical theories of receptor function to molecular structure represents a growing field of research on the nervous system. PMID- 10514841 TI - Antibodies to glutamate receptors: a role in excitatory dysregulation of the central nervous system? PMID- 10514842 TI - Evidence for glutamate receptor autoimmunity in the pathogenesis of Rasmussen encephalitis. PMID- 10514843 TI - GABA and glutamate transporters: therapeutic and etiologic implications for epilepsy. PMID- 10514844 TI - Glial and epilepsy. Introduction. PMID- 10514846 TI - Modulation of neuronal excitability by astrocytes. PMID- 10514845 TI - Physiology of glial cells. PMID- 10514847 TI - Contribution of astrocytes to seizure activity. PMID- 10514848 TI - The molecular neuron-glia couple and epileptogenesis. PMID- 10514849 TI - Initiation, synchronization, and spread of epileptic discharges. Introduction. PMID- 10514850 TI - Mechanisms of epileptogenesis. PMID- 10514851 TI - Synaptic plasticity in kindling. AB - Kindling is an experimental model of epilepsy resulting from progressive activity dependent changes in neuronal structure and function. During kindling, pathologic changes occur at various levels of organization of the nervous system, ranging from altered gene-expression in individual neurons to the loss of specific neuronal populations and the rearrangement of synaptic connections. This chapter summarizes recent findings about the long-lasting (plastic) nature of the alterations at the level of single neurons with emphasis on the role of altered excitatory and inhibitory amino acid receptors. The modified synaptic ligand gated ion channels (i.e., "epileptic receptors") may ultimately be responsible for the kindling epileptogenesis. PMID- 10514852 TI - Presynaptic and postsynaptic mechanisms of long-term potentiation. PMID- 10514853 TI - Second messengers, long-term potentiation, gene expression and epileptogenesis. PMID- 10514854 TI - Epileptogenic neurons and circuits. PMID- 10514855 TI - Role of metabotropic glutamate receptors in epilepsy. AB - Considerable information is available regarding the role of ionotropic glutamate receptors in the generation of interictal spikes. Progress in the study of metabotropic glutamate receptors (mGluRs) makes clear that activation of these receptors can contribute greatly to seizure discharges and epileptogenesis. The effects of activation of the different mGluR subgroups on neuronal hypersynchrony and the initiation and propagation of seizure discharges in hippocampal slices are discussed herein. To help one understand the mechanisms that underlie these effects, information regarding the action of mGluRs on cellular and synaptic properties is summarized. The data bring to the forefront the critical role of mGluRs in epilepsy and emphasize the anticonvulsant effects of group II and III mGluR activation as opposed to the convulsant action of group 1, which elicits seizure discharges and epileptogenesis in experimental models. PMID- 10514856 TI - Mechanisms of neuronal synchronization during epileptiform activity. PMID- 10514857 TI - Functionally relevant and functionally disruptive (epileptic) synchronized oscillations in brain slices. AB - Focal seizurelike events can be induced in experimental preparations by means of a number of distinct manipulations that differ in synaptic mechanisms. Nevertheless, the form of the seizurelike events can be explained with common principles, including long-lasting excitation of pyramidal cell dendrites and recurrent excitation between pyramidal cells that provides synchronization. One means of induction of seizurelike events, tetanic stimulation, induces a more physiologic type of activity before seizures are elicited, that is, gamma frequency (> 20 Hz) oscillations. Such oscillations, called 40-Hz oscillations, are believed to be important for cognition in vivo. Experimental gamma oscillations depend critically on synaptic inhibition between interneurons, from interneurons to pyramidal cells, and on a tonic drive to pyramidal cells and interneurons by metabotropic glutamate receptors. The function of gamma oscillations appears to be imposition of a precise temporal structure on the firing patterns of pyramidal cells while still allowing the pyramidal cells to influence each other and be influenced by afferents selectively. We suggest that a relative loss of synaptic inhibition, occurring by any of a number of mechanisms, prevents the occurrence of gamma activity, allows recurrent pyramidal cell-pyramidal cell excitation to predominate, and thereby allows neuronal networks to generate functionally disruptive seizures. PMID- 10514858 TI - Basic mechanisms of status epilepticus. AB - This chapter reviews two main aspects of the basic mechanisms of status epilepticus--acute factors, which are important in inducing status epilepticus in an in vitro brain slice model of status epilepticus, and the acute and chronic epileptogenic consequences of status epilepticus. Status epilepticus is difficult to produce in vitro in normal extracellular medium. This suggests that seizure terminating mechanisms are normally quite robust. To produce long- duration, self sustained epileptic discharges in vitro, we have found it necessary to include reciprocally connected entorhinal cortex with our hippocampal slices. Doing so closes the normal excitatory limbic loop in the brain. We found incorporation of the full loop in our brain-slice preparations necessary to bring about epileptic discharges of long duration that fit the definition of status epilepticus. Reentrant activation from distant sites may be necessary for maintenance of status epilepticus-like activity of long duration. Similar requirements may exist for generalized tonic-clonic status epilepticus discharges, but as yet no data support or refute this hypothesis. There are both acute and chronic consequences of an episode of status epilepticus. Acute consequences are alterations in membrane potential and membrane properties of hippocampal pyramidal cells accompanied by alterations in neurotransmitter-activated conductances and receptor expression. Some of these acute alterations in receptor and transmembrane iongradient associated with status epilepticus may be critically involved in the development of drug resistance during the late stages of status epilepticus. Long-term consequences of status epilepticus in the limbic system include alterations in patterns of expression of neurotransmitter receptors and in the function of excitatory and inhibitory synapses, cell loss, and circuit rearrangements within the limbic system. An episode of status epilepticus that involves the limbic system clearly elicits brain damage, at least among adult animals. This brain damage can contribute to the development of epilepsy, or a condition of recurrent, spontaneous seizures. Conversely, development of an epileptic condition enhances the susceptibility of the limbic system to trigger status epilepticus discharges. PMID- 10514859 TI - Epileptic cell damage and epileptogenesis. Introduction. PMID- 10514860 TI - Neurocytology of a primate model of human temporal lobe epilepsy. PMID- 10514861 TI - Neuronal loss and synaptic reorganization in temporal lobe epilepsy. PMID- 10514862 TI - Synaptic reorganizations in human and rat hippocampal epilepsy. PMID- 10514863 TI - Physiological and anatomical correlates of the human dentate gyrus: consequences or causes of epilepsy. AB - The presence of paroxysmal discharges in the epileptic human dentate gyrus provides a physiologic basis for hyperexcitability that may initiate seizure discharges during the development of epilepsy. Although these responses can occur with single orthodromic stimulation, data obtained under conditions that weaken synaptic inhibition (e.g., 1 Hz stimulation or bicuculline disinhibition) suggest that paroxysmal discharges may be a more common feature of tissue from temporal lobe epileptic patients than has been reported previously. Hilar cell loss and weakened synaptic inhibition may provide conditions favorable for the activation of N-methyl-D-aspartate acid (NMDA) receptors that would allow triggering of paroxysmal discharges that normally never are evoked in dentate granule cells in nonepileptic humans. As the dentate gyrus in normal animal tissue is not susceptible to intrinsic bursting behavior and is characterized by a relatively short duration excitatory postsynaptic potential even under pharmacologic disinhibition, paroxysmal discharges in the epileptic human dentate gyrus may provide an important clue to understanding the prerequisite conditions for seizure discharge. PMID- 10514864 TI - Excitability changes in epileptic human dentate gyrus and temporal neocortex. PMID- 10514865 TI - The role of nonprincipal cells in dentate gyrus excitability and its relevance to animal models of epilepsy and temporal lobe epilepsy. PMID- 10514866 TI - Excitation and inhibition in temporal lobe epilepsy: a close encounter. PMID- 10514867 TI - Seizure-induced hippocampal damage and chronic epilepsy: a hebbian theory of epileptogenesis. PMID- 10514868 TI - Epilepsy and the blood brain barrier: endothelial cell responses to seizures. PMID- 10514869 TI - Frontiers in brain imaging and therapeutics. Introduction. PMID- 10514870 TI - Cerebral blood flow and glucose metabolism in human epilepsy. PMID- 10514871 TI - Basic mechanisms of childhood epilepsies: studies with positron emission tomography. AB - Although functional imaging with positron emission tomography (PET) and single photon emission computed tomography are useful in the clinical evaluation of intractable epilepsy, these techniques have not been widely applied to understanding the basic mechanisms of the epilepsies. Among patients with infantile spasms, PET studies with 2-deoxy-2[18F]fluoro-D-glucose (FDG) suggest that the spasms are the result of secondary generalization from cortical foci and that maturational factors result in the recruitment of basal ganglia and brainstem serotonin mechanisms that lead to secondary generalization and the unique semiology of the spasms. Attempts to develop an animal model of infantile spasms have not been successful. Glucose utilization studies in the Lennox Gastaut syndrome also indicate cortical lesions and further suggest that the electroencephalographic pattern of 1 to 2.5 Hz spike-wave activity (slow spike wave pattern) is an interictal phenomenon. There is a remarkable consistency between 14C-2-deoxyglucose autoradiographic findings and PET observations of glucose utilization performed for patients in the ictal, interictal, and postictal states. Although three patterns of ictal glucose hypermetabolism have been described, hypermetabolism also can be seen in the postictal and interictal clinical states and in various animal models. Preliminary studies of benzodiazepine receptor binding with PET have found that the cortical epileptic region of decreased binding is smaller than the region of hypometabolism on glucose utilization studies, but detailed electrophysiologic comparisons have not been made. Development of new PET methods for the study of presynaptic and postsynaptic neurotransmitter functions will offer unique opportunities in the study of epileptic mechanisms. PMID- 10514872 TI - Positron emission tomography receptor studies. AB - Several PET receptor ligands have been used to investigate the neurochemical basis of the epilepsies. 11C-Flumazenil binds to the central benzodiazepine receptor (cBZR)-gamma-aminobutyric acid (GABA) A receptor complex; 11C diprenorphine, 18F-cyclofoxy, and 11C-carfentanil to opiate receptors; and 11C Deprenyl to monoamine oxidase B. These studies should be considered alongside high-quality magnetic resonance imaging that demonstrates the structural basis of the condition. The results should be correlated with those of quantitative in vitro neuropathologic and autoradiographic studies. Idiopathic generalized epilepsy has been studied with 11C-flumazenil and 11C-diprenorphine. There is no evidence of any interictal overall abnormality of opioid receptors in idiopathic generalized epilepsy, but typical absences have been found to displace 11C diprenorphine from the association areas of the neocortex. This finding implies that release of endogenous opioids has a role in the pathophysiologic mechanisms of typical absences in humans. In contrast, binding of 11C-flumazenil to cBZRs has been shown not to be affected by serial absences. Studies of interictal 11C flumazenil binding in idiopathic generalized epilepsy have not given uniform results. In one investigation a slight reduction was reported in the neocortex of patients with idiopathic generalized epilepsy in comparison with patients with partial seizures. Also observed was increased benzodiazepine receptor density in the cerebellar nuclei and decreased density in the thalamus. Widespread increases in cBZRs also have been reported in cerebral neocortex, thalamus, and cerebellar cortex. In unilateral hippocampal sclerosis, reduction of binding of 11C flumazenil has been shown to be confined to the hippocampus and to be over and above that caused by neuron loss and hippocampal atrophy. In malformations of cortical development, abnormalities of cBZRs, as demonstrated with 11C-flumazenil PET, are more extensive than the structural abnormality revealed with magnetic resonance imaging. There often are areas of increased cBZRs, a pattern that appears unique to malformations of cortical development and that may reflect both functional and structural anomalies. In patients with mesial temporal lobe epilepsy, upregulation of mu opioid receptors has been found in lateral neocortex without an overall increase of opioid receptor binding. The pathophysiologic explanation for this finding is not clear. Possibilities include up-regulation of mu receptors in response to epileptic activity and down-regulation or occupation of kappa opioid receptors. Important future developments in this field that will increase understanding of the processes that underlie the epilepsies will come from the development of further ligands, particularly tracers that are specific for excitatory amino acid receptors, the subtypes of the opioid receptors, and the GABAB receptor. PMID- 10514873 TI - Positron emission tomography: the contribution of cognitive activation paradigms to the understanding of the epilepsies. AB - Cognitive activation paradigms coupled with positron emission tomographic techniques may aid in the identification of functional and epileptogenic zones for presurgical evaluation. More work is needed to determine the most clinically efficacious paradigms for different seizure types. The real strength of activation positron emission tomography lies in the ability to study shifts in cognitive circuitry that accompany a fixed neuropathologic entity for both groups of similar subjects and individuals. These techniques are enhancing our understanding of the fundamentals of brain plasticity and may be used in the future to predict precise surgical risks. PMID- 10514874 TI - New NMR measurements in epilepsy. General introduction, functional magnetic resonance imaging, magnetic resonance spectroscopy, and diffusion-weighted imaging. PMID- 10514875 TI - New NMR measurements in epilepsy. Diffusion-weighted magnetic resonance imaging of spreading depression. PMID- 10514876 TI - New NMR measurements in epilepsy. T2 relaxometry and magnetic resonance spectroscopy. PMID- 10514877 TI - New NMR measurements in epilepsy. Measuring brain GABA in patients with complex partial seizures. PMID- 10514878 TI - AMPA receptors in epilepsy and as targets for antiepileptic drugs. AB - alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors are key mediators of seizure spread in the nervous system and represent promising targets for antiepileptic drugs. There is emerging evidence that AMPA receptors may play a role in epileptogenesis and in seizure-induced brain damage. This evidence suggests that AMPA receptor antagonists could have broad utility in epilepsy therapy. Regional, developmental, and disease-associated variations in AMPA receptors produced by differential expression of AMPA receptor subunits and variations in posttranscriptional processing, including alternative splicing and pre-mRNA editing, provide a diversity of functionally distinct AMPA receptor isoforms that allow opportunities for selective drug targeting. Four types of AMPA receptor antagonist are discussed in this chapter: (a) competitive AMPA recognition site antagonists, including those of the quinoxalinedione and newer nonquinoxalinedione classes, (b) 2,3-benzodiazepine noncompetitive (allosteric) antagonists, (c) desensitization enhancing antagonists, exemplified by SCN-, and (d) antagonists of Ca(2+)-permeable AMPA receptors, including polyamine amide arthropod toxins and their synthetic analogues. Competitive and noncompetitive AMPA receptor antagonists are broad-spectrum anticonvulsants in animal seizure models. Their effectiveness and safety for humans remain to be determined. There is evidence that these antagonists can potentiate the antiseizure activity of N methyl-D-aspartate (NMDA) receptor antagonists and conventional antiepileptic drugs. This evidence suggests that the preferred use of AMPA receptor antagonists may be in combination therapies. Agents that enhance desensitization may have advantages in comparison with other AMPA receptor antagonists to the extent that they preferentially block high-frequency synaptic signaling and avoid depressing AMPA receptors on interneurons, which would lead to disinhibition and enhanced excitability. Evidence has accumulated that Ca(2+)-permeable AMPA receptors (those lacking the edited GluR2 subunit) may play a role in epileptogenesis and the brain damage occurring with prolonged seizures. Because Ca(2+)-permeable AMPA receptors are predominately expressed in gamma-aminobutyric acid (GABA)ergic interneurons, it is hypothesized that some forms of epilepsy might be caused by reduced GABA inhibition resulting from Ca(2+)-permeable AMPA receptor-mediated excitotoxic death of interneurons. It is further proposed that drugs that selectively target Ca(2+)-permeable AMPA receptors might have antiepileptogenic and neuroprotective properties. Certain polyamine toxins and their analogues are channel-blocking AMPA receptor antagonists that selectively inhibit Ca(2+) permeable AMPA receptors. These substances might give clues to the development of such antagonists. PMID- 10514879 TI - Excitatory amino acid receptors and antiepileptic drug development. PMID- 10514880 TI - Pharmacological properties of recombinant and hippocampal dentate granule cell GABAA receptors. PMID- 10514881 TI - Neuronal circuitry of thalamocortical epilepsy and mechanisms of antiabsence drug action. AB - Powerful mechanisms exist within the thalamus that lead to the promotion of synchronous and phasic 3 Hz neuronal activity. These mechanisms include robust burst-firing capability of thalamic neurons, recurrent excitatory and inhibitory synaptic connectivity, and long-lasting and powerful inhibitory synaptic responses arising from activity in thalamic reticular neurons and mediated by gamma-aminobutyric acid (GABA) receptors. The 3 Hz thalamic synchronization appears to arise from a perturbation of a physiologic, higher frequency spindle oscillation. Two currently available antiabsence medications interact with this circuitry with the net result of decreased synchronization, largely through reduction in inhibitory output from the thalamic reticular nucleus. Ethosuximide blocks T-type calcium channels and thus reduces the ability of thalamic neurons to fire bursts of spikes, thereby reducing inhibitory (and excitatory) output within the circuit. By contrast, clonazepam enhances recurrent inhibitory strength within the reticular nucleus. This results in a decreased ability of neighboring inhibitory neurons to fire synchronously and produce the powerful inhibitory synaptic responses that are required for network synchronization. PMID- 10514882 TI - Adenosine and suppression of seizures. PMID- 10514884 TI - Genetic analysis of neuronal physiology with defective herpes simplex virus vectors. PMID- 10514883 TI - Mechanisms of new antiepileptic drugs. PMID- 10514885 TI - Perspectives on priorities, time management, and patient care. PMID- 10514886 TI - Home HIV tests prove highly inaccurate. PMID- 10514888 TI - Creating outcomes with redesign efforts. AB - Integrating principles from a variety of theories, managers have developed a conceptual framework for reengineering processes in an endoscopy unit to improve the value of services provided to customers. A major goal of this redesign was to enhance or maintain quality of care, increase efficiency, and maintain or reduce costs. This was accomplished by analyzing data and outcome measures related to patient, physician, and staff member satisfaction, as well as resource allocation. The departmental results were tangible, positive, and visible almost immediately. With the right team and the right techniques, tools, methodologies, and decision-making processes, redesign projects can and do lead to dramatic improvements in productivity, service, customer and staff member satisfaction, cost control, and innovation. PMID- 10514887 TI - Lost gallstones--a small but real complication. AB - Retained gallstones are considered a complication of cholecystectomies. If stones can be removed using minimally invasive procedures (i.e., laparoscopically), patients' recovery time may decrease and satisfaction may increase. Correct and immediate diagnosis of this complication at the time of the procedure often is the determining factor between patients' full recovery and their potential chronic illness. This article describes diagnostic tests and techniques used to surgically correct this complication and presents two case studies depicting patients' experiences from a few months to nine years postoperatively. PMID- 10514889 TI - Entrepreneurship--transforming a perioperative problem into a product solution. AB - No one is more capable of identifying problems related to health care than health care practitioners. Identifying a problem is the first step to every new product or invention. This article provides step-by-step information to help health care practitioners identify problems and possible solutions, look for a creative company to bring the solution to the marketplace, and present the product idea to potential industry partners. The author also discusses the legal aspects of product development and the advantages and disadvantages of licensing a new product or starting a business to develop a product. PMID- 10514890 TI - Issues surrounding xenotransplantation. AB - Currently there is a shortage of cadaver organs that can be transplanted from one human being to another. In response to this shortage, scientists and medical researchers have developed techniques for transplanting animal organs into humans, a procedure known as xenotransplantation. This may address the shortage of organs for patients in need; however, it raises other concerns related to cross-species disease transmission, consent issues, ethical issues of sacrificing animals for their organs, psychological issues of receiving organs from an animal, religious concerns, and economic factors. These medical, ethical, and philosophical issues need to be thoroughly addressed before xenotransplantation becomes a reality. PMID- 10514891 TI - Etiology and incidence of pressure ulcers in surgical patients. AB - This experimental study was designed to identify the etiology of pressure ulcers in a surgical sample and to evaluate a special OR mattress overlay in preventing pressure ulcer development. Surgical patients (N = 413) were randomized to receive "usual perioperative care" or the new mattress overlay. Over six postoperative days, 89 patients (21.5%) developed pressure ulcers, primarily stage I. Only 2% developed stage II or IV ulcers. Patients with ulcers were statistically older, had diabetes, were smaller in body mass, had lower Braden Scale scores on admission, and used the new mattress overlay (P < .02). Pressure ulcers that presented as "burns" or ecchymosis did not deteriorate to stage III or IV ulcers during the study. The mattress overlay was not effective in preventing pressure ulcer development. PMID- 10514892 TI - The RN first assistant's role in a laparoscopic cholecystectomy. AB - Laparoscopic cholecystectomy is one of the most frequently performed laparoscopic procedures today and involves the same surgical principles as an open cholecystectomy, except that access to the abdomen is gained transabdominally using video laparoscopic techniques and instrumentation. Registered nurse first assistants (RNFAs) provide retraction and tissue exposure for the surgeon during this procedure and should be aware of safe laparoscopic surgical techniques and potential injuries that can occur from manipulation of surrounding structures. The experience and expertise of the RNFA when assisting in a laparoscopic cholecystectomy are key to the successful completion of the surgery. PMID- 10514893 TI - Postlaryngospasm pulmonary edema in adults. AB - Laryngospasm is a potential serious complication of intubation. Pulmonary edema can develop after laryngospasm and can affect any patient who has been intubated. Postlaryngospasm pulmonary edema is potentially life threatening and can result in reintubation, mechanical ventilation, admission to an intensive care unit, and a prolonged hospitalization for the patient. Perioperative nurses play a significant role in the prompt detection, diagnosis, and treatment of this syndrome. PMID- 10514894 TI - The legal implications of Y2K. PMID- 10514895 TI - New direction for Medicare reimbursement. PMID- 10514897 TI - Internet resources for pressure ulcer prevention and treatment PMID- 10514896 TI - When research cannot answer a question--standards of care. PMID- 10514898 TI - Antioxidative activity of 1-methyl-1,2,3,4-tetrahydro-beta-carboline-3-carboxylic acid. AB - The (-)-(1S, 3S) isomer of 1-methyl-1,2,3,4-tetrahydro-beta-carboline-3 carboxylic acid was synthesised by Pictet-Spengler condensation of tryptophan with acetaldehyde. It was evaluated for its antioxidative activity in the linoleic acid autooxidation system by the ferric thiocynate method. Butylated hydroxy anisole (BHA), butylated hydroxy toluene (BHT), and alpha-tocopherol were used as the reference standards. The compound showed moderate antioxidative activity and also synergistic effect with the reference standards. The synergistic effect was in the increasing order of BHT, alpha-tocopherol and BHA. The synergistic effect was higher at higher concentrations studied. PMID- 10514899 TI - Cerebral blood flow effects of the nitric oxide donor, nitroglycerin and its drug combinations in the non-human primate model. AB - Nitroglycerin (CAS 55-63-0, Nitrocene) has successfully been used in the management of angina during the last several decades. Although important information on the pharmacological actions and efficacy of nitroglycerin have been extracted, to date, limited research has been conducted on its effects on cerebral blood flow. In recent years, with the aid of SPECT (single photon emission computed tomography) and PET (positron emission tomography) it has been shown that marked cerebral blood flow changes occur under treatment of a wide variet of drugs. Illucidation of the pharmacological mode of action of nitroglycerin has gained momentum with the discovery of nitric oxide (NO) as an endogenous mediator and with the knowledge that nitroglycerin acts as a NO donor. The present study investigated the effects of nitroglycerin (0.25 microgram/kg/min over 10 min) on the cerebral blood flow, using 99mTc-HMPAO (hexamethylpropylene amine oxime) and SPECT, in an anaesthetised primate model, as well as the effects of its drug interactions with therapeutic agents that influence cerebrovascular dynamics, e.g. sumatriptan, nimodipine and acetazolamide. The present study with nitroglycerin indicates that the response time to measure cerebral blood flow effects seems to be present and an important factor as the transient is relatively short. The current treatment regime with nitroglycerin indicates a slight increase, when compared with control control results, although not significant, except for regional significant increases in particular the occipital regions of the brain. Drug interaction between nitroglycerin and nimodipine may occur as a reduction of 20% in cerebral blood flow from the control control was observed in this case. The results for the combination of nitroglycerin with sumatriptan showed a further increase of the cerebral blood flow to near significance, when compared with the control results and is significantly increased (+27%) when compared with sumatriptan treatment alone. Effective treatment with sumatriptan may therefore be compromised with simultaneous administration of nitroglycerin or NO donor drugs. The combination of nitroglycerin and acetazolamide suggested that the increase in cerebral blood flow is primarily attributed to the influence of acetazolamide. The cerebral blood flow effects of these drugs and possible interactions during an angina attack need to be investigated. PMID- 10514900 TI - Biological activities of novel thienothiazine derivatives on heart and smooth muscle preparations of guinea pigs. AB - New thienothiazine derivatives which differ in their side chains on the nitrogen atom of the thienothiazine ring were investigated regarding their biological activity and compared with calcium antagonistic drugs. Isometric contraction force was measured in guinea-pig papillary muscles, aortic strips and terminal ilea. Chronotropic activity was studied in right atria of guinea pigs. MS 69 with a ethylpyridylcarbonyl side chain had the most potent negative inotropic effect on isolated papillary muscles, followed by MS 74, which has an ethylmethylpiperazinylthiocarbonyl side chain. The negative inotropic effect could be antagonized by increasing the extracellular calcium concentration. MS 87 with an ethylpyridylcarboxamide side chain had the most potent relaxing effect on aortic strips and MS 48 with an ethylbenzylpiperazinylcarbonyl side chain was most potent on terminal ilea. Compounds with a more hydrophilic side chain had less activity. It is concluded that increasing the lipophilicity by substitution of an oxygen atom by a sulfur atom (MS 74) or addition of a pyridine ring (MS 69) results in a higher biological activity. PMID- 10514901 TI - In vitro absorption studies with carvedilol using a new model with porcine intestine called BM-RIMO (Boehringer-Mannheim ring model). AB - Carvedilol (CAS 72956-09-3, Dilatrend) is a beta-blocker with additional vasodilating, antiproliferative and antioxidative properties. It is indicated for the treatment of high tension (HT), coronary artery disease (CAD) and congestive heart failure (CHF). Carvedilol has been investigated in numerous in vivo studies and thus comparisons of in vitro results to in vivo observations are possible. In this publication the results of studies on the in vitro absorption of carvedilol in a new model called BM-RIMO (Boehringer-Mannheim ring model) using porcine intestine are reported. In particular the influence of absorption enhancers and of pH and the existence of absorption windows were examined and the relevance of the obtained data for in vivo conditions was estimated. The main route of carvedilol absorption seemed to be transcellular. In vitro as well as in vivo absorption decreased within the intestine in the following order: jejunum > ileum > colon. The highest amount of in vitro absorption of carvedilol was achieved in the jejunum at a neutral pH. Enhancers such as bile and the mucoadhesive agent chitosan had opposite effects on the absorption of the compound. The results indicate that BM-RIMO is a simple, cheap and fast tool for the investigation of the influence of absorption enhancers, pH and different parts of intestine on absorption. For carvedilol the in vitro model tends to overestimate absorption in the colon, possibly because of the lack of faeces. PMID- 10514902 TI - Pharmacokinetics of pentoxifylline after oral administration of a sustained release tablet at two different times of the day. AB - The pharmacokinetics of pentoxifylline (CAS-6493-05-6) was studied in healthy subjects by orally administering a 400 mg sustained release tablet (Trental) at two different times (10:00 or 22:00 h) of the day in a crossover design. Pentoxifylline concentrations in serum samples were estimated by using high performance liquid chromatography. The mean values of Cmax (326.38 +/- 39.77 vs 266.35 +/- 36.0 ng/ml, p < 0.01, n = 8), AUC0-t (2424 +/- 382 vs 2141 +/- 300 ng/ml/h, p < 0.05, n = 8) were significantly higher and Vss/f (16537 +/- 2869 vs 20136 +/- 5006 ml/kg), Vd/f (11807 +/- 2704 vs 15801 +/- 5960 ml/kg) were significantly (p < 0.05, n = 8) lower following morning (10:00 h) administration than in the night (22:00 h). These variations should be considered while designing sustained release dosage forms. PMID- 10514903 TI - Synthesis of some new pyridyl-ethylated benzoxazolinones with analgesic and anti inflammatory activities. AB - Twelve new 3-[2-(2- and/or 4-pyridyl)ethyl]benzoxazolinone derivatives have been synthesized by reacting 2- and/or 4-vinylpyridine and appropriate benzoxazolinones. Their chemical structures have been proven by IR, 1H-NMR and elemental analysis. Analgesic activities of these compounds were investigated by a modified Koster and constant temperature hot-plate test. Test results revealed that most of the compounds at 100 mg/kg dose level showed significant analgesic activities when compared to acetylsalicylic acid and morphine. Therefore the compounds were screened for their anti-inflammatory activities using the carrageenan hind paw edema test. Compounds 3-8 were found more active than indometacin. In gastric ulceration studies, any of the compounds showed no gastrointestinal bleeding at 100 mg/kg dose level. PMID- 10514904 TI - Synthesis and antichagasic properties of new 1,2,6-thiadiazin-3,5-dione 1,1 dioxides and related compounds. AB - In a search for antichagasic drugs, 14 new 4-(nitroarylidene)-1,2,6-thiadiazin 3,5-dione 1,1-dioxide derivatives were synthesized and tested in vitro against the epimastigote form of Trypanosoma cruzi and some of them showed important antiprotozoan activity. Attempts to synthesize new 4-(nitroarylidene)-3,5-diamino 1,2,6-thiadiazine 1,1-dioxides were unsuccessful. The antichagasic properties of nitroarylidene-malononitrile and nitroarylidene-cyanoacetamide derivatives, thus obtained, were also tested. The cytotoxic properties against Vero cells of compounds which showed remarkable in vitro antichagasic activity were evaluated. All compounds tested exhibited high toxicity percentages at 100 micrograms/ml. However, compound 3c showed in vitro antichagasic and cytotoxic properties such as nifurtimox at the dose of 10 micrograms/ml. PMID- 10514905 TI - Synthesis and antiprotozoan properties of new 3,5-disubstituted-tetrahydro-2H 1,3,5-thiadiazine-2-thione derivatives. AB - In a search for antiprotozoan compounds, 34 new 3,5-disubstituted-tetrahydro-2H 1,3,5-thiadiazine-2-thione derivatives were synthesized and tested in vitro against Trypanosoma cruzi and Trichomonas vaginalis. Some of them showed important antiprotozoan activity. In vivo assays of compounds which showed remarkable in vitro activity against T. vaginalis were carried out. PMID- 10514906 TI - Cytotoxic activity of Acinetobacter baumannii after treatment with aminoglycosides. AB - The effect of three aminoglycoside antibiotics--netilmicin (CAS 56391-57-2), gentamicin (CAS 1403-66-3) and amikacin (CAS 37517-28-5)--at subinhibitory concentrations (sub-MICs; 1/4 or 1/16 of the MICs) on the cytotoxic activity of Acinetobacter baumannii was studied. The cytotoxic factor was manifested by reduction of the number of isolated rat lung type II cells evaluated by alkaline phosphatase determination. All aminoglycosides at both concentrations studied (with the exception of 1/4 of the MIC of amikacin) did not modify cytotoxic activity of A. baumannii. This activity was partially inhibited only after treatment with amikacin at 1/4 of the MIC. In this case the number of type II cells after incubation with antibiotic treated A. baumannii was higher (71%) as compared to the number of type II cells cultured with untreated A. baumannii (47%). PMID- 10514907 TI - Effect of Sch 55700, a humanized monoclonal antibody to human interleukin-5, on eosinophilic responses and bronchial hyperreactivity. AB - This report describes the development and the biology of Sch 55700, a humanized monoclonal antibody to human IL-5 (hIL-5). Sch 55700 was synthesized using CDR (complementarity determining regions) grafting technology by incorporating the antigen recognition sites for hIL-5 onto consensus regions of a human IgG4 framework. In vitro, Sch 55700 displays high affinity (Kd = 20 pmol/l) binding to hIL-5, inhibits the binding of hIL-5 to Ba/F3 cells (IC50 = 0.5 nmol/l) and blocks IL-5 mediated proliferation of human erythroleukemic TF-1 cells. In allergic mice, Sch 55700 (0.1-10 mg/kg, i.p. or i.m.) inhibits the influx of eosinophils in the lungs, demonstrates long duration of activity and the anti inflammatory activity of this compound is additive with oral prednisolone. In allergic guinea pigs, Sch 55700 (0.03-30 mg/kg i.p.) inhibits both the pulmonary eosinophilia and airway hyperresponsiveness and at 30 mg/kg, i.p. inhibited allergic, but not histamine-induced bronchoconstriction. In allergic rabbits, Sch 55700 blocks cutaneous eosinophilia. Sch 55700 (0.1-1 mg/kg i.p.) also blocks the pulmonary eosinophilia and neutrophilia caused by tracheal injection of hIL-5 in guinea pigs. In allergic cynomolgus monkeys, a single dose of Sch 55700 (0.3 mg/kg i.v.) blocks the pulmonary eosinophilia caused by antigen challenge for up to six months. Sch 55700 is, therefore, a potent antibody against IL-5 in vitro and in a variety of species in vivo that could be used to establish the role of IL-5 in human eosinophilic diseases such as asthma. PMID- 10514908 TI - Studies on activities of invariant chain peptides on releasing or exchanging of antigenic peptides at human leukocyte antigen-DR1. AB - An invariant chain peptide (Ii77-92; YRMKLPKSAKPVSQMR; 'Ii-Key') enhances 10-50 times baseline levels, the presentation of synthetic antigenic peptides to murine T cell hybridomas, by an exchange mechanism at cell surface MHC class II molecules. Two differing activities, to promote the release of antigenic peptide in the presence or absence in solution of a second antigenic peptide, were characterized with truncation homologs through assays for release or binding of human myelin basic protein biotinylated (*) peptide 90-102 on purified HLA-DR1: 1) release of bound hMBP *peptide from DR1 in the presence or absence of free hMBP peptide in solution, 2) exchange of hMBP *peptide from solution with hMBP peptide on DR1, and 3) binding of hMBP *peptide to 'empty' DR1. Peptides such as Ii81-88, LPKSAKPV, released prebound hMBP *peptide from DR1 without free hMBP peptide in solution. They also exchanged hMBP *peptide from solution for prebound hMBP peptide. Peptides including hIi77-83, LRMKLPK, released hMBP *peptide only when free hMBP peptide was in solution. Nevertheless, hIi77-85, LRMKLPKPP, released hMBP peptide without hMBP peptide in solution. Either type of peptide accelerated hMBP *peptide binding to 'empty' DR1. Competitive binding assays with hMBP *peptide or several *Ii-Key truncation homologs, with respective not biotinylated forms, demonstrated that the Ii77-83, LRMKLPK, binding site was distinct from the HLA-DR1 antigenic peptide binding site. PMID- 10514909 TI - Peroral polypeptide delivery. A comparative in vitro study of mucolytic agents. AB - Besides the absorption and enzymatic barrier, the diffusion barrier based on the mucus gel layer covering gastrointestinal (GI) epithelia represents the third major factor being responsible for a very poor bioavailability of orally administered (poly)peptide drugs. In order to overcome the latter ones, the mucus liquefying action of various types of mucolytic agents and their influence on the model polypeptide drug insulin has been evaluated in a comparative in vitro study. The results demonstrated that the proteases pronase (EC 3.4.24.31) and papain (EC 3.4.22.2) cause a relative reduction in mucus viscosity of 75.9 +/- 6.8% and 51.1 +/- 3.3% (n = 3) after 6 h of incubation at pH 5.0, respectively, whereas the mucolytic effect of the detergents Triton X-100 (octoxinol) and Tween 20 (polysorbate 20) was markedly lower. Mucolytic proteases and well established mucolytic sulfhydryl compounds, however, caused a rapid degradation of insulin. Therefore these two types of mucolytic agents can only be used if the (poly)peptide drug proves stable towards degradation. The results indicate the strict need of novel mucolytic agents for the peroral administration of therapeutic (poly)peptides. PMID- 10514910 TI - Diagnostic and surgical features of Klatskin tumors. AB - AIM OF STUDY: Klatskin tumors are rare and their prognosis is poor. Long term survival can be expected only after a surgical resection, the treatment of choice. The aim of this study is to report our single centre experience and, by literature analysis, to define the role of surgery in the treatment of hilar cholangiocarcinoma. MATERIALS AND METHODS: Between 1990 and 1998, 27 patients affected by Klatskin's tumor were observed. Eight women and seven men (mean age 59 years) underwent surgical resection. Thirteen patients (86%) had curative resection (7 hilar resection (HR), 4 HR combined with partial hepatectomy (PH) and 2 HR + PH with portal vein resection). Two patients (13%) had palliative biliary resection and surgical drainage. RESULTS: One in-hospital death was recorded after a right hepatectomy with portal vein resection (6.6%). Postoperative morbidity rate was 40%. Patients were regularly followed up to date or to death. Ten patients died and 5 survived. The 1-, 2- and 3-year survival rate after curative resection was 84%, 54% and 34%. The median survival was 28.5 months. Lymph node involvement did not show a statistically significant difference on median survival between the positive group and the negative group (26.2 vs 29.8 months), nor did perineural invasion, because of the small number of patients. The 1-, 2-, 3- and 5-year survival rate after isolated hilar resection was 100%, 57.1%, 28.6% and 0. Four out of 6 patients who underwent hilar resection combined with partial hepatectomy are still alive 1, 23, 29, 38 months after resection. Hepatectomy increased mortality (16% vs 0). Palliative biliary resection and surgical drainage were successfully performed in 2 patients with satisfactory results. CONCLUSION: Aggressive surgical treatment of Klatskin tumors can improve patients' survival. Careful preoperative management has to be carried out by a multidisciplinary approach including surgeons, gastroenterologists, radiologists and pathologists. Hepatic resection involving the caudate lobe is often performed in order to obtain microscopic tumor-free margins and curative resection (R0). Biliary drainage and treatment of cholangitis is mandatory before surgery in order to improve surgical outcome. Surgical treatment is characterized by high technical difficulties and better results can be achieved by hepatobiliary surgical teams. PMID- 10514911 TI - Liver transplantation. A summary of our surgical practice. AB - At the turn of the new century, liver transplant procedures can finally be considered an efficient treatment option. Technology has helped transplant intervention become a preferred treatment for patients with progressive and irreversible liver failure. New immuno-suppressive drugs have been introduced which reduce the patient's immunological reaction to the implanted organ, entail minimal side effects and improve practical applications of liver transplantation. As a result of these technological advanced and proper disease management, liver transplant procedures are no longer thought of as an elite therapy, reserved for selected patients with end stage liver disease. In our opinion, it is now a sound and valid surgical option with strictly defined characteristics, indications and well-understood limits. Throughout the past decade, we have studied and applied this type of intervention and have come to terms with its rapid expansion at both the theoretical and practical levels. The most significant obstacle remaining today is the discrepancy between the ever increasing demand and limited supply of organs. The future of liver transplant lies in overcoming this obstacle. Liver transplant practice began at our Institute on 23 November 1990 with the first surgical intervention to replace an organ. In the past 10 years, we have exceeded 200 liver transplant procedures. PMID- 10514912 TI - Pre and intra-operative methods of staging gastric cancer. AB - The aim of this paper was to examine the indications and limits of pre- and intra operative instrumental diagnosis of gastric cancer. In order to achieve this effectively, the authors emphasized the importance of proper staging methods and obtained a detailed description of tumor diffusion. The most important diagnostic instruments considered were magnetic resonance, endosonography, intra-operative echography, pre- and intra-operative immunoscintography and a cytological examination of peritoneal lavage fluid. The authors concluded that pre- and intra operative staging of gastric cancer is important for two major reasons: it results in the most accurate definition possible of disease evolution, enabling a proper therapeutic program; and it involves a combination of three complementary metasurgical treatments. PMID- 10514913 TI - The use of prosthesis in abdominal and thoracic wall defect, 15 year experience: evaluation of tissue reactions and complications. AB - Synthetic prosthesis (Polypropylene, Dacron and expanded Polyterafluoroethylene) is now widely used in abdominal and thoracic wall reconstructive surgery. Many surgeons have reported great success with various types of prosthetic implants but tissue reactions and other complications have never been well defined. The aim of this study was to determine which molecules react upon tissue contact, which synthetic materials result in less complications and whether some non specialized prosthetics are correlated with certain types of complications. We studied 54 patients from 1982-1997 who had each been re-operated on for prosthetic complications. Our clinical data was then compared to data collected from animal models. Twenty-one pigs received one or more prosthetic implants: 14 of these pigs received their implants with a "proper surgical technique" while 8 underwent "improper surgical technique". The results from both the clinical and animal study were significantly similar. From a microscopic point of view, we can conclude that different tissues react in the same way with the same or similar types of prostheses. The reactions begin to differ when the thickness and rigidity of the material is considered. A PTFE-polyporpylene combination (Composix Mesh) seems to be the most effective solution, especially in abdominal defect repair which involves peritoneal organ contact. We would also like to emphasize that prosthetic complications can be quite serious and this type of procedure should only be performed by experienced and qualified surgeons. PMID- 10514914 TI - Predictive value of a pathophysiological score in the surgical treatment of perforated diverticular disease. AB - Resection is the preferred method of perforated diverticular disease treatment compared to conservative treatment. However, the immediate or deferred timing of bowel continuity restoration for advanced degrees of peritoneal contamination is debatable. This is a retrospective study designed to identify operative mortality predictors and guidelines for safe primary anastomosis. A pathophysiological score (acute physiology and chronic health evaluation, APACHE II) was applied to 135 consecutive patients who had undergone surgery for acute inflammatory complication of diverticular disease. A multivariate analysis was used to identify prognostic factors such as age, chronic diseases, neoplastic cancer, Acute Physiology Score (APS), Hinchey's classification and APACHE II scores. Seventy patients underwent primary resection and anastomosis, 35 underwent Hartmann's procedure and 15 conservative treatment. There was a significant correlation between operative mortality and increasing disease severity based on Hinchey's classification, APS and APACHE II scores. The multivariate analysis proved APACHE II scores to be the only prognostic factor of operative mortality. Both single and multivariate analysis of variance failed to identify a factor significantly associated with surgical and/or medical postoperative complications. APACHE II scores were the best predictor for operative mortality in patients with diverticular disease complications, but none of the classification criteria used was effective in predicting postoperative complication. Patients with phlegmonous sigmoiditis can be safely treated with primary resection and anastomosis. Conservative treatment should not be considered an effective method for diverticular disease. A prospective trial comparing resection with and without colostomy should be done for local and diffuse purulent peritonitis treatment. Hartmann's procedure is seen to be the only indicator for faecal peritonitis. PMID- 10514915 TI - [Liver transplantation from elderly donors]. AB - This retrospective, case-controlled study compared the outcome of 17 OLTs (group A) using livers donated by subjects over 70 years of age with 17 OLTs (group B) with livers from donors under 40. Clinical data were used form the period 1996 1998. The following variables were considered in the analysis: donor clinical and laboratory parameters, cold ischemic periods, intra-operative blood and plasma replacement, 30-day mortality rate, incidence of primary graft dysfunction, acute rejection and arterial complication and long term survival. The main post operative parameters were also included. Liver biopsy, performed in 9/17 of group A, revealed minimal steatosis. There were 2 post-operative deaths in group A and 1 in group B (p = NS). Two arterial complications were observed in group A (p = NS) and only one patient required retransplantation (p = NS). The only other difference found among clinical variables was the amount of total bilirubin at post-operative days 8 and 10, aPTT at days 6 and 13 and albumin at days 5 and 6. A two-year follow-up showed survival rates to be 88.2% and 94.1% for groups A and B, respectively (p = NS). Candidates over 70 years of age should be excluded as liver donors. In such cases, greater care needs to be placed on pathological vascular conditions related to advanced stage atherosclerosis such as calcified plaques on the hepatic artery, a possible factor in severe postoperative complications. PMID- 10514916 TI - [Usefulness of chromogranin A determination in the diagnosis of neuroendocrine neoplasia]. AB - Neuroendocrine gastroenteropancreatic tumor diagnosis is a very difficult and expensive procedure. This study compared Chromogranin A (CgA) to Neuron-specific enolase (NSE) in 55 patients affected by neuroendocrine tumors. Advanced local or metastatic neoplasia was found in 43 patients. Radical operation was performed in 12 patients. Seventeen cases of lung microcystoma, 23 cases of other intestinal tumors and 19 patients affected by irritable bowel syndrome were used as controls. CgA sampling demonstrated sensitivity of 73% and specificity of 66%, a positive predictive value of 77% and a negative predictive value of 61% while NSE sampling showed sensitivity of 100%, specificity of 36%, a positive predictive value of 15% and a negative predictive value of 100%. CgA values demonstrated a statistically significant difference between patients with neuroendocrine tumors and tumor-free resected patients (p = 0.0015), microcystoma patients (p = 0.0087), other types of neoplasia (p = 0.01) and irritable bowel syndrome patients (p = 0.0004). No significant difference was found among the same groups when NSE values were analyzed. The high diagnostic accuracy of CgA sampling renders it very useful in early neoplastic detection, even in cases of nonfunctioning neoplasms or absence of liver metastases. In addition, CgA sampling may be an effective screening test in patients with irritable bowel syndrome or with liver or lung metastases when there is no evidence of the primitive tumor. PMID- 10514917 TI - [Treatment of chronic critical ischemia of the lower limbs with spinal cord electrostimulation]. AB - The treatment of Fontaine's third and fourth stage chronic critical lower limb ischemia can be considered a medical, social and economic problem. One current form of therapeutic intervention in some cases is medullary electrostimulation (SCS: spinal cord stimulator). This study looks at the period from January 1998 to December 1997 in which patients were selected for an etiological and symptomatological examination. The criteria established at the European Consensus Conference on Critical Leg Ischemia were employed to perform medullary electrostimulation. The entire procedure included a trial period with a temporary implant and if considered tolerable and effective, a permanent implant. One hundred sixty-four patients (117 male and 46 female, aged 41-93) affected by peripheral obstructive arteriopathy were examined. Etiological causes included atherosclerosis (70.7%), diabetes or other atherosclerotic diseases (25%), inflammatory arteriopathy (1.8%) and chronic renal failure under dialysis treatment (2.4%). The procedure was successful in 103 patients (62.8%) while unsuccessful in 61 (37.2%). The best results were obtained in patients at the Fontaine's 3rd stage in which the limb was saved in 72.2% of the patients and at the beginning of the 4th stage with a success rate of 62.7%. The advanced 4th stage had a success rate of only 42.4%. From an etiological point of view, the rate of limb preservation for atherosclerosis patients was 68.1%, in diabetics 56% and inflammatory diseases 33%. However, no positive results were obtained in patients with chronic renal failure. PMID- 10514918 TI - [Thyroid carcinoma: evaluation of prognostic factors]. AB - Between 1990 and 1997, 36 thyroid cancer patients were observed at the III Department of Surgical Oncology, Regina Elena Cancer Institute in Rome. There were 31 (86.1%) women and 5 (13.9%) men, with a mean age of 47.7 years (range 17 72). Diagnostic data consisted of: papillary carcinoma in 25 patients, follicular carcinoma in 9 and Hurthle cell carcinoma in 1. Twenty patients were less than 45 years old (15 papillary and 5 follicular carcinoma) and 16 were more than 45 years old (10 papillary and 4 follicular carcinoma, 1 Hurthle cell carcinoma and 1 with undifferentiated cancer). We performed total thyroidectomy in 32 cases and isthmectomy in 3 (2 papillary carcinoma, T1, < 45 years, 1 follicular carcinoma with minimal invasion). At the time of diagnosis, 6 patients with papillary carcinoma showed signs of local metastasis. No patients exhibited distant diffusion. A follow-up was performed at mean 41 months (2-84 months). Two patients with follicular carcinomas had been treated with radioiodine and showed disease progression with distant metastasis. Our results coincide with the literature on this topic. Total thyroidectomy is preferred, in low-risk patients as well, because subsequent radioiodine treatment and disease relapse monitoring are facilitated. Lateral cervical lymphadenectomy was performed only in cases when there was clinical evidence of metastasis and recurrent disease at this level has never been observed. PMID- 10514919 TI - [Perforation of the esophagus during pneumatic dilatation in achalasia]. AB - Esophageal perforation is a serious complication of pneumatic dilatation. We studied the cases of 4 patients (2 men and 2 women, mean age 58 years, range 56 62) who had surgical treatment for achalasia, two of which had had previous dilatation. The main symptoms were pain and dyspnea. Pneumomediastinum was present in all patients, pleural effusion in 2 and cervical emphysema in 1. Esophagographic results showed evidence of perforation in all four cases and gastric patches were surgically placed on the esophageal tear within 12 hours. Three patients received enteral nutrition for an average of 13 days. Mean hospital stay was 14 days. No post-operative complications were exhibited although one patient did develop gastroesophageal reflux 3 months later and underwent surgery to repair a hernia in the thorax 5 years later. Early and aggressive treatment is considered the best therapy and the gastric patch, in our opinion, is an effective and reliable technique for esophageal perforation repair in achalasia patients. PMID- 10514920 TI - [Preoperative diagnosis using color Doppler flowmetry in focal and diffuse thyroid pathology]. AB - Color-doppler was first used in the study and classification of specific pathologies in 1992. One hundred and eighteen patients with focal and diffuse thyroid pathologies underwent color-doppler, flowmetry analysis and peak systolic velocity measurement (CD-FM-PSV). The PSV results allowed us to identify two subclasses a and b in class three and four (a: = < or = 30 cm/sec., b: = > 30 cm/sec.). Class 3a and 3b lesions are the most likely to represent neoplastic nodules. Based on our results, we assigned 58 patients to type 2 (follicular hyperplasia), 20 patients to type 3a (follicular adenoma and carcinomas), 16 patients to type 3b (carcinomas and Plummer's adenoma), 15 patients to type 4a (autoimmune thyroiditis and hypothyroidism) and 12 patients to type 4b (Graves' disease). Preliminary results were compared with FNAB, intra-operative and post operative histological data. The specificity of CD-FM-PSV in diagnosis is 86%. We have concluded that CD-FM-PSV is an effective imaging technique for pre-operative diagnosis of thyroid pathologies and along with FNAB, a adequate predictive tool for thyroid nodules. PMID- 10514921 TI - Multivisceral cluster transplantation: a preliminary experience. PMID- 10514922 TI - [2 cases of choledochal calculosis associated with juxtapapillary diverticulum in patients with gastric resection: submesocolon approach]. AB - Although laparoscopy and endoscopy have reduced the need for laparotomies in biliary tract surgery, open surgery is sometimes still needed. One case in particular is when previous operations have significantly distorted normal upper abdomen anatomy. We chose an inframesocolic entrance to the posterior peritoneum in two patients with bile duct stones, juxtapapillary duodenal diverticulum and a history of cholecystectomy and partial gastric resectioning. The duodenum was reached at the junction between the second and third section by entering the posterior peritoneum through the inferior sheet of the mesocolon, a relatively avascular area. The diverticulum was incised, the sphincter and papilla operation was performed and the bile duct stones removed. The diverticulum was then resected. Our conclusion is that in certain cases, an inframesocolic entrance can significantly reduce technical difficulties involved in re-operating through dense adhesions, minimize surgical time and blood loss and, when operating through the open diverticulum, spare an unnecessary duodenotomy. PMID- 10514923 TI - [First-line chemo-radiotherapy neoadjuvant treatment in locally advanced (T4) epidermoid carcinoma of the esophagus]. AB - In the period 1993-1997 we performed two phase II pilot studies of first-line chemo-radiotherapy in patients with locally advanced (T4) SCC of the esophagus. The first protocol (3 cycles of DDP-VP16 + 45 Gy) was used in 37 patients: toxicity was not negligible; a clinical tumor downstaging was obtained in 54% of cases; an R0 resection surgery was performed in 40% of patients. The overall median survival of the whole group of 37 patients was 11 months, while it was > 36 months for patients undergoing R0 resection. The second protocol (4 cycles of DDP-5FU + 45 Gy) was used in 25 patients: a clinical tumor downstaging was obtained in 55% of cases, and R0 resection surgery was performed in 45% of patients. The overall median survival of the whole group was 11 months. To date, all patients but one (who died after 13 months) are alive with a median follow up of 13 months. The prognosis of both groups of patients was improved compared to patients with T4 SCC of the esophagus who did not undergo chemo and/or radiotherapy. The survival advantage was especially evident for those who were able to undergo an R0 resection. First line chemo-radiotherapy should be considered the standard treatment for locally advanced esophageal SCC. PMID- 10514924 TI - [Current trends in the surgical treatment of lesions caused by caustic ingestion]. AB - During a twenty-six year period from 1973 to 1999 a total of 78 patients with severe esophago-gastric lesions after caustic ingestion were referred to and managed at our unit. Hydrochloric acid was the most frequently involved caustic agent and the lesions were located in the esophagus (52.6%), in the stomach (19.2%) and in both the esophagus and stomach (28.2%). Thirty-seven patients were managed by endoscopic dilation of esophageal stricture that gave permanent relief of dysphagia in 13 cases only. Twelve patients are still managed with endoscopic dilation and in another 12 we performed bypass surgery. Two perforations developed and spontaneously sealed after T.P.N. for 6-8 weeks. No death occurred. Fifty-three patients were operated on in emergency or during survey period or for stenotic lesions. Surgical procedures were: in esophageal strictures a substernal esophago or pharyngocologastroplasty without esophagectomy; in esophago-gastric strictures, after an esophago-gastrectomy, an esophago or pharyngocolojejunoplasty in one or two steps; in total gastric stenosis without esophageal involvement a total gastrectomy followed by a Roux-en-Y esophago jejunal anastomosis and in antropyloric strictures a Billroth I or II partial gastrectomy. The morbidity rate was 32% with a 5.6% mortality rate. PMID- 10514925 TI - [Pulmonary carcinoids. Analysis of 53 cases]. AB - We have performed a retrospective analysis of 53 cases of bronchial carcinoids using our own patient data from more than 4700 lung tumors and 1632 resections. The male/female ratio was 1:12 (28/25) and the age range 13 to 75 years (mean 52.2). Fifty-three tumors resections of varying extent were performed, including one radical pneumonectomy in a patient who had previously undergone a lobectomy, and one limited resection of the main left bronchus; there was no intraoperative mortality. After histological examination, 44 tumors (83%) were found to be typical carcinoids and nine (17%) atypical carcinoids. The median follow-up period was 4.56 years, with a range from 1 to 10 years. Only one patient with an atypical carcinoid tumor had a relapse and died three years after, while another patient underwent surgery of the contralateral lung for a second primary lung cancer (SPLC). On the basis of these observations we would like to underline the importance of an accurate histopathological classification for both therapeutic and prognostic purposes; given the higher aggressiveness of atypical carcinoids, these tumors would be eligible for a therapeutic approach analogous to that adopted for bronchogenic carcinoma. PMID- 10514926 TI - Small solitary pulmonary nodules: assessment of enhancement and enhancement patterns in benign and malignant tumours by high resolution computed tomography. AB - Our aim was to determine the accuracy of quantitative and qualitative findings of contrast-enhanced computed tomography (CT) by means of differential analysis of small uncalcified solitary pulmonary nodules and to compare the CT diagnosis with the results of transthoracic needle biopsies (TTNB). We assessed a consecutive series of 109 patients with 66 malignant or 45 benign pulmonary nodules before TTNB and surgery with contrast and high resolution computed tomography (HRCT). Pulmonary nodules were classified as small when equal to or smaller than 15 mm and large when larger than 15 mm. Diagnostic accuracy of CT qualitative evaluation was 95% for large nodules and 92% for small nodules. Specificity was 92% for small nodules, 80% for large nodules. Enhancing regular septa were observed in 28 hamartomas (80%) while except for two cases (3%), inner septa were absent or irregular in malignant tumours. TTNB accuracy was 70% for small nodules and 94% for large ones. Low-enhancing hamartomas are more frequent in Italy than in the US where the prevalence of high-enhancing granulomas in benign nodules reduces the specificity of quantitative CT analysis. We propose that certain geographic areas would benefit from enhanced-CT in place of TTNB in managing lung nodules equal to or less than 1.5 cm. PMID- 10514927 TI - [Laparoscopic treatment of gastroesophageal reflux]. AB - Gastroesophageal reflux disease (GERD) is a frequent illness, sometimes causing disabling symptoms and/or permanent oesophageal lesions. Etiology is multifactorial and not completely defined. Therapy is medical at first step, surgical indication is reserved to those patients with less compliance for medical therapy, unsuccessful medical therapy or reflux related complications. Different surgical techniques have been suggested for treatment of GERD, like Nissen, Rossetti or Toupet fundoplication. During the last decade laparoscopy has been proposed as a less invasive approach when surgery is indicated. From 1995 to the first months of 1999, 42 pts (28 females, 14 males, mean age 53.7 years), were operated on. Diagnosis and surgical indication were confirmed preoperatively by barium X-rays, endoscopy and 24 hrs-Ph-manometry. Hiatal hernia was demonstrated in 37 cases (88%), I or II grade esophagitis in 16 and III grade in 2; 1 patient had Barrett oesophagus. 37 pts were operated on by laparoscopic Nissen fundoplication, 5 patients had a Toupet operation. Mortality and conversion rate were 0. Complications occurred in 3 patients: 1 intraoperative pneumothorax, 1 acute cardiac ischemia in a patient with known hypertension, 1 permanent dysphagia successfully treated by endoscopic dilatation. Mean postoperative hospital stay was 6.1 days. Mean follow up was 9 months (3-48) in 100% of cases. Despite the fact that few patients were operated on by using this new less invasive approach, results are encouraging with no mortality, less morbidity and great advantages for patients. PMID- 10514928 TI - [Aortic dissection: role of the general surgeon in diagnostic-therapeutic procedure]. AB - Aortic dissection occurs when there is a tear or separation of the aortic intima from the media; flow of blood into the intima-media space allows the tear to develop into a dissecting hematoma. Aortic dissection is a rare condition which represents an acute cardiovascular emergency for which the appropriate therapy is immediate surgical correction. Patients with aortic dissection show a heterogeneous constellation of symptoms; hence, clinical suspicion is often difficult. Only a minority of patients has "classic" symptomatology, the electrocardiogram is often misleading and the chest radiogram is almost always non-specific. In Italy, the usual "hospital routine practice" assigns the key role in the emergency diagnostic procedure for these patients to the general surgeon. In view of the necessity of immediate cardiac surgery and the overwhelming likelihood of adverse events when surgery is delayed, techniques for diagnosis must be accurate, widely available and easily and quickly used. The present study consists of four case-reports of aortic dissection thoroughly examined and confronted with the literature. Management strategy based on emergency echocardiography is a reliable, feasible and successful technique for evaluating patients with aortic dissection. It allows a rapid accurate diagnosis with a single examination that can also be performed in the emergency room and provides information of sufficient diagnostic value to allow immediate cardiovascular surgery. CT scan and/or MRI are also valid tools for the emergency diagnosis of aortic dissection. However, a major problem still exists: the range of symptoms is sufficiently broad that a high index of "personal suspicion" of aortic dissection is required on the part of the general surgeon when he performs the role of "emergency-team leader". PMID- 10514929 TI - [Treatment of supra-aortic trunk aneurysms: report on 16 cases]. AB - The Authors report their own experience in treating 16 patients suffering from aneurysms of the supra-aortic trunks, 6 of the innominate artery, 4 of the common carotid artery, 3 of the intrathoracic subclavian artery and 3 of the extrathoracic subclavian artery. Eleven cases were of atherosclerotic nature, 3 from Takayasu's, 1 dysplasic and 1 mycotic. To aneurysms of the innominate artery with diameter less than 2 cm and one of the subclavian artery, in a patient with on carcinoma of the lung, was treated non surgically; all other patients were operated. The mortality was 0, and morbidity rate very low. With a mean follow-up of three years, only a patient, with Takayasu's disease, developed a recurrence and one, with mycotic aneurysm, died from persistence of the sepsis. PMID- 10514930 TI - [Vascular clamping in cirrhotic liver surgery]. AB - Different vascular clamp methods in liver surgery have led to less complications. The aim of this study was to evaluate the results after hepatic resection involving different vascular clamping methods and liver function outcome. Our study examined 46 patients who underwent surgery for liver lesions, developed on cirrhotic and noncirrhotic livers, applying the technique of selective clamping and pedicular clamping. There was one death (1/17; 5.9%) due to postoperative liver failure which occurred in a cirrhotic liver patient who underwent left hepatectomy with pedicular clamping. Complication rate was higher, but not significant (4/7; 57.1%) in the group with selective clamping compared to those with pedicular clamping (3/10; 30%). Hemorrhagic complications were observed in a higher rate among patients with selective clamping (3/7; 42.9%) compared to those with pedicular clamping (1/10; 10%). Selective clamping seems to find major indications in patients with chronic liver disease undergoing minimal hepatic resections. Intermittent pedicular clamping seems to be more effective in regards to blood loss and postoperative hepatic function. PMID- 10514931 TI - [Routine use of open laparoscopy: technique and results in 1006 consecutive cases]. AB - Blind insertion of the Veress needle and/or the first trocar is one of the most frequent causes of laparoscopic surgery complications. Nevertheless, the closed technique is still more preferred than the open one. The Authors retrospectively analyzed 1006 consecutive laparoscopic procedures in which Hasson's technique was routinely utilized. The overall complication rate was 2.2%, but the vast majority of complications occurred during the learning curve (6% vs. 1.9%). The Authors conclude that after the first 50 cases the open technique is a quick and safe procedure. PMID- 10514932 TI - [Surgical problems in the aged patients]. AB - Authors, considering the increasing of the middle duration of life and the increasing of geriatric population, examine various surgical problems in the elderly. They took in consideration not only the age, but all the other markers of surgical risk related to the surgical illness, to the associate illness and to the type of operation. After reporting their experience in the treatment of geriatric patients, they conclude that a scrupulous surgical preparation, a correct indication to the operation and an accurate overseeing after surgery are necessary to do that the elderly patient faces surgical intervention with the same capability of success of the young. PMID- 10514933 TI - [Tissue expression of carbohydrate antigen 19-9 (CA 19-9) in adenomatous lesions of the colorectum]. AB - Serum levels of tumor markers almost not detectable in precancerous states or early cancer, the behavior of the tumor associated antigen CA 19-9 in colorectal carcinogenesis was analysed. In order to investigate the adenoma-carcinoma sequence, tissue antigenic expression was measured in adenomas, characterized by different size and histology, and compared with that found in normal colonic mucosa. Tissue content of CA 19-9 was determined by an immunoperoxidase technique (ABC-POD) in 88 colonic polyps, and correlated to the degree of histological dysplasia, and dimension of adenoma. Tissue content of CA 19-9 was also evaluated in non-adenomatous mucosa obtained by endoscopic biopsy. Among the 88 polyps, 50 showed the tubular histological type, while 31 resulted tubulovillous and 7 villous. High degree of dysplasia was present in 7 adenomas (7.9%) and focal carcinoma was observed in 6 (6.8%). Positivity for CA 19-9 was registered in 60.2% of adenomas. No correlation was found between tissue-CA 19-9 and degree of dysplasia, size of adenoma and villous component. However, a statistically significant correlation was observed between expression and cellular distribution of the antigen (chi 2 = 98.07, p < 0.00001). Our data confirmed CA 19-9 expression in adenomas, but it is unlikely this tissue antigen proves to be a reliable marker of adenoma-carcinoma sequence. PMID- 10514934 TI - [Spontaneous rupture of the excretory tract following renal colic in kidney malformation]. AB - This paper reports a group's experience in treating a case of excretory tract rupture caused by a renal colic. After careful analysis and description of this case as well as related literature analysis, they have hypothesised the dynamic physiopathological events involved in this case. In addition, they have offered clinical considerations in preventing and detecting the development of such a lesion. PMID- 10514936 TI - Cardiac disease in the adult: MR evaluation. AB - Cardiac MR imaging to date has provided detailed information regarding cardiac structure and anatomy. Recent developments in imaging speed have broadened the applications of cardiac MR from the research laboratory to a wide variety of clinical applications involving assessment of both cardiac function and perfusion. In this review, we present methods used for MR evaluation of acquired cardiac disease in the adult. Applications to ischemic heart disease, cardiomyopathy, coronary imaging, myocardial perfusion, and pericardial disease are discussed. PMID- 10514935 TI - [Choledochojejunostomy using a mechanical stapler]. AB - Authors performed the mechanical termino-lateral bilio-digestive anastomosis, adopting a circular 21 mm stapler device. The patient was affected by non neoplastic papillary stenosis producing a common biliary tract dilatation so large to allow the stapler's head introduction. A 12 month follow-up was performed using ultrasonography and MRI cholangiopancreatography, that obtained a good demonstration of both biliary tract and bilio-digestive anastomosis. PMID- 10514937 TI - Imaging features of Pneumocystis carinii pneumonia. AB - Despite a declining prevalence secondary to improved prophylaxis, Pneumocystis carinii remains an important pulmonary pathogen in the immunocompromised host. Because the radiologist is often the first to suggest the diagnosis of PCP, an awareness of the entire spectrum of imaging features associated with this organism is important. The classic presentation of PCP is a bilateral interstitial pattern, which may be characterized as finely granular, reticular, or ground-glass opacities. When chest radiographic findings are normal or equivocal, high-resolution CT may be helpful, because it is more sensitive than chest radiographs for detecting PCP. The classic CT finding is extensive ground glass attenuation. Increasingly recognized characteristic patterns of PCP in AIDS patients include cystic lung disease, spontaneous pneumothorax, and an upper lobe distribution of parenchymal opacities. Although the radiographic findings in PCP are similar for AIDS and non-AIDS immunosuppressed patients, cystic lung disease has not been described in the latter patient population. PMID- 10514938 TI - Proinsulin in subjects with hyperglycaemia and in first degree relatives of patients with IDDM. PMID- 10514939 TI - Haemophilus influenzae type b. Epidemiology of invasive diseases, antimicrobial resistance of clinical isolates, and response to a conjugate vaccine in selected risk groups. PMID- 10514940 TI - Magnetic resonance imaging in rheumatoid arthritis. Quantitative methods for assessment of the inflammatory process in peripheral joints. PMID- 10514941 TI - Active life in old age. Combining measures of functional ability and social participation. AB - This paper describes a new measure of Active Life Expectancy, called Active Life Classification (ALC) in which the criterion for successful aging is a combination of good functional ability and high social participation. OBJECTIVES: 1) to describe the distribution of ALC among 75-year-old men and women, 2) to investigate the association between ALC and life satisfaction and 3) to describe how ALC is determined by socio-demographic, psycho-social, and health factors. DESIGN: A cross-sectional population survey. SETTING: Eleven municipalities in the Western part of Copenhagen County in 1989. SUBJECTS: A random sample of 75 year-old people who were invited to participate in the study (participation rate: 89, n = 477). MAIN OUTCOME MEASURE: ALC is a combination of two dichotomized variables: functional ability (dependent vs not dependent of help) and social participation (low vs. high). RESULTS: For both men and women an active life (measured by ALC) was significantly associated with life satisfaction. For men only good self-rated health was related to ALC in the multivariate analysis. Among women high income, many social contacts, good self-rated health, good memory and lack of chronic diseases were associated with ALC. CONCLUSIONS: It is an advantage to combine functional ability and social participation in the description of quality of life in old age, as 1) a high social participation may compensate for a poor functional ability, and vice versa, 2) the combined measure is meaningful for both sexes, and 3) it gives more information than the two concepts used as separate outcome measures. PMID- 10514942 TI - Empirical treatment of bacteraemic urinary tract infection. Evaluation of a decision support system. AB - INTRODUCTION: In a Danish county with a low prevalence of antibiotic resistance to most antibiotics, we have constructed and evaluated a decision support system (DSS) for guidance of empirical antibiotic therapy in patients with bacteraemia originating from the urinary tract. METHODOLOGY: The DSS was based upon a causal probabilistic network, and a decision theoretic approach was used to balance the costs of antibiotic therapy against the therapeutic benefit. The costs included direct cost of purchasing antibiotics, side effects, and the risk of development of antibiotic resistance. The therapeutic benefit was defined as the increase in life-expectancy caused by antibiotic therapy. Life-years were chosen as the common currency unit. Four hundred and ninety-one bacteraemias seen during 1992 1994 were used to construct the DSS (derivation set), and 426 bacteraemias during 1995-1996 were used for evaluation (validation set). The cases were identified in a regional register of bacteraemias. The study was non-interventional. RESULTS: The DSS suggested antibiotics which would provide coverage in 377 of the 426 episodes (88.5%) compared to 259 episodes (60.8%) for which empirical therapy actually provided coverage (p < 0.01, McNemar-test). The regimens suggested by the DSS included mecillinam as monotherapy in 240 episodes (56.3%), gentamicin as monotherapy in 81 (19.0%), and a combination of gentamicin and ampicillin in 80 (18.8%). CONCLUSION: A decision theoretic approach shows promise of improving empirical antibiotic treatment, and may be a measure to support an antibiotic policy. PMID- 10514943 TI - The Danish registers of causes of death. AB - In 1875 registration of causes of death in Denmark was established by the National Board of Health, and annual statistics of death have since been published. Until 1970 the national statistics were based upon punched cards with data collected from the death certificates. Since then the register has been fully computerized and includes individual based data of all deaths occurring among all residents in Denmark dying in Denmark. Furthermore, a microfilm of all death certificates from 1943 and onward is kept in the National Board of Health. The Danish Institute for Clinical Epidemiology (DICE) has established a computerized register of individual records of deaths in Denmark from 1943 and onwards. No other country covers computerized individual based data of death registration for such a long period, now 54 years. This paper describes the history of the registers, the data sources and access to data, and the research based upon the registers, presenting some examples of research activities. PMID- 10514945 TI - Applying the reality principle to health care policy. PMID- 10514944 TI - The Danish in vitro fertilisation (IVF) register. AB - The Danish IVF Register was established in 1994 and covers all treatments with in vitro fertilisation (IVF), intracytoplasmatic sperm injection (ICSI), frozen embryo replacements (FER) and egg donations (ED). Since data are recorded with personal identification numbers, they provide the starting point for cohort studies of treated women and offspring. It is obligatory for each clinic to report each treatment cycle to the register, by means of special treatment report forms that contain clinical as well as laboratory data. The pregnancy outcome is reported on special forms no later than two months after birth. The personal identification number (CPR) allows cross-linkage of the data from the register, with several other national Danish registers, such as the National Hospital Register the Abortion Register, the Danish Register of Causes of Death, the Cytogenetic Central Register and the Cancer Register. In 1998 a total of 7131 IVF and ICSI cycles were performed in Denmark. This corresponds to around 6500 cycles per 1 million women in the reproductive age, which is among the highest number per capita in the world. The coverage of the register is believed to be very close to 100% for the treatment reports, but less for the pregnancy outcome forms, at least during the first two years after the register was established. The main importance of the register is quality control aspects of assisted reproductive techniques and research in relation to follow-up on maternal and infant health. PMID- 10514946 TI - Treatment trials in real world settings. Methodological issues and measurement of disability and costs. PMID- 10514947 TI - Captive patients, captive doctors: clinical dilemmas and interventions in caring for patients in managed health care. AB - This article explores common clinical dynamics resulting from the denial of choice that many patients experience in managed health care and proposes clinical adaptations for the treating or consulting psychiatrist. Patients who feel they have been denied the right to choose their health plan, treatment setting, or personal physician commonly go through a subjective experience analogous to that of being held captive. This sense of captivity can exacerbate the feelings of helplessness and hopelessness brought on by serious illness. It can also intensify the patient's feelings of alienation and betrayal when managed care constrains patient-physician decision making by limiting treatment options. These dynamics can lead to identifiable transference reactions and, in turn, to physician countertransference. Psychiatrists can do much to ameliorate these potentially destructive dynamics both as treating therapists and as consultants to general physicians. Indications for consultation or intervention are analyzed and specific clinical strategies to enhance the patient's decision-making capacity throughout the introductory, ongoing, and termination phases of the treatment alliance are reviewed. PMID- 10514948 TI - Neurasthenia: cross-cultural and conceptual issues in relation to chronic fatigue syndrome. AB - The purpose of this study was to examine several conceptual and cross-cultural issues in neurasthenia, particularly in terms of their relationship to chronic fatigue syndrome. A review of this relationship led to the conclusion that these conditions are much more alike in Western countries than in countries such as China, where neurasthenia could almost be regarded as a "culture-bound syndrome." This may be a consequence of factors such as the heterogeneous nature of neurasthenia and different diagnostic practices in different countries, despite the ICD-10 definition of neurasthenia, intended for worldwide use. Likewise, there is no consensus on what the "core" characteristics of neurasthenia are, because its clinical presentation and key features in different countries are very different. Despite the finding of relatively low comorbidity rates between neurasthenia and other mental disorders, clinical experience suggests that features of neurasthenia frequently overlap with those of depression, chronic anxiety, and somatoform disorders. There is no convincing evidence that in cases of overlap or comorbidity, other diagnoses should automatically have "primacy" over neurasthenia nor should the diagnosis of neurasthenia thereby be excluded. Although some aspects of its validity have improved recently, especially its descriptive validity, the overall validity of the diagnosis of neurasthenia is still not satisfactory. Suggestions for further research, aimed at improving the diagnostic validity of neurasthenia, are offered in this paper. PMID- 10514949 TI - Is managed care ethical? AB - The author identifies ethical dilemmas inherent in fee-for-service medicine (grounded in patients' autonomous decision making) and managed care (grounded in the principle of efficiency). He argues that the moral legitimacy of the latter system of health care is more in question, and suggests a methodology for assessing its ethical status. PMID- 10514950 TI - The treatment effectiveness project. A comparison of the effectiveness of paroxetine, problem-solving therapy, and placebo in the treatment of minor depression and dysthymia in primary care patients: background and research plan. AB - This report describes the background, rationale, and research plan for a comparative treatment trial of the effectiveness of paroxetine, problem-solving therapy (PST-PC), and placebo in the treatment of minor depression and dysthymia in primary care patients. Patients were recruited from a variety of primary care practice settings in four separate geographic locations (Hanover, New Hampshire; Pittsburgh, Pennsylvania, San Antonio, Texas; and Seattle, Washington). Patients were randomly assigned to each of the three intervention conditions the medication/placebo conditions were double-blinded. The treatment trial was 11 weeks, with independent assessments of patient clinical status at baseline, 6 weeks, and 11 weeks. There was a follow-up at 25 weeks. Since there are relatively few placebo-controlled trials in primary care settings on patients with these disorders, the background of this project and a description of it are presented at this time, prior to the availability of outcome data, to provide methodological detail and to increase awareness in the research community of this treatment trial, with results to appear subsequently. PMID- 10514951 TI - Effectiveness research and implications for study design: sample size and statistical power. AB - Most clinical trials have started to incorporate more broadly defined outcome measures, such as health-related quality of life, to complement clinical status measures as well as direct costs and cost-effectiveness analyses. Contrasting a broad range of outcome and cost measures, we analyze the implications for sample sizes and study design using data from prior mental health and primary care studies that span a wide range of practice settings, patient populations, and geographic areas. While meaningful clinical symptomatic differences are often detectable with sample sizes of well under 100 per cell, detecting even large changes in health-related quality of life generally requires several hundred observations per cell. Reasonable precision in cost estimates usually requires sample sizes in the thousands. Very few clinical trials or observational effectiveness studies that incorporate quality of life or cost measures have such sample sizes, resulting in many (unreported) null findings and, due to publication biases favoring significant results, scientific publications that exaggerate true effects. It raises issues for the general direction of clinical trials and effectiveness studies, as well as for how cost and health-related quality of life results based on small studies should be dealt with in publications. PMID- 10514952 TI - Qualitative outcome assessment of a medical ethics program for clinical clerkships: a pilot study. AB - This study assessed the usefulness of an open-ended case analysis test instrument for evaluating the effects of a 1-year ethics course on medical students' decision-making skills. Through case-oriented seminars in gynecology, internal medicine, obstetrics, pediatrics, psychiatry, and surgery, third-year medical students were taught a structured analytic framework for analyzing clinical ethical problems stressing the interactive relationships among medical indications, patient preferences, quality of life, and contextual (social, legal, economic) matters. At precourse, the students were given a test case and asked to provide a line of reasoning for their clinical decisions. At postcourse, the students were given the same case. Content analysis of pre- and postcourse responses of a random student sample revealed increases in student awareness in the following areas: 1) consideration of informed consent, 2) professional liability, 3) physician-assisted suicide, and 4) resource utilization. With some modifications, open-ended case analysis holds promise for evaluating medical ethics courses. The authors make recommendations for future research in evaluating the true impact of clinical ethics courses in medical education. PMID- 10514953 TI - Neuropsychiatric and neuropsychological manifestations of central pontine myelinolysis. AB - A patient with central pontine myelinolysis (CPM) underwent neurological and mental status examination, as well as neuropsychological testing, during the acute stage of the disease. After correction of the hyponatremia, a gross change in his neuropsychiatric status was observed. The patient underwent extensive neurological, psychiatric, and neuropsychological testing during the acute phase of the disease and at follow-up 4 months later. All major neurological and neuropsychiatric symptoms present at onset were fully reversible. Neuropsychological examination revealed deficits in the domains of attention and concentration, short-term memory and memory consolidation, visual motor and fine motor speeds, and learning ability. Although improved, neuropsychological testing still revealed remarkable deficits at follow-up. We conclude that neuropsychological deficits can accompany CPM, and that these deficits do not necessarily diminish simultaneously with the radiological or clinical neurological findings but may persist for a longer period of time, or even become permanent. In his recovery the patient started to manifest new neurological symptoms consisting of a mild resting tremor of both hands and slow choreoathetotic movements of the trunk and the head, which we considered to be late neurological sequelae of CPM. The significance of CPM in the differential diagnosis of acute behavioral changes after correction of hyponatremia is stressed, even if correction is achieved slowly and carefully. PMID- 10514954 TI - Help-seeking behaviors in relatives of schizophrenics in Taiwan. AB - Despite its merits, the Western-style psychiatric community rehabilitation model is not well accepted by caregivers in Taiwan. We examined factors affecting the utilization of community rehabilitation programs in Taiwan. Our stepwise logistic regression revealed that psychoeducation regarding the biological cause of schizophrenia emerged as the major factor for increasing utilization treatment modality. Eighty-nine pairs of schizophrenic patients (who had been recommended for rehabilitation) and their relatives were divided into two groups, the rehabilitation group and the nonrehabilitation group. Both groups were surveyed on help-seeking behavior scales and mental function measurements. The results showed no significant differences in patients' psychopathology, though the rehabilitation group had higher employment rates. As for caregivers, the rehabilitation group scored significantly better on some cognitive appraisals, whereas the nonrehabilitation group was more inclined to institutionalize the patients for life. No significant differences were noticed on rejection attitude, subjective care burden, or expressed emotion measures. Improving caregiver's knowledge about the disease, providing activities that lend emotional, physical, and financial support and thereby reduce the burden and increase the satisfaction of caregivers may be useful. Besides making the Western-style psychiatric community rehabilitation model more effective and accessible for patients and caregivers in Taiwan, cultural adaptation is also needed. PMID- 10514956 TI - Controversial aspects of hormone replacement therapy upon cardiovascular mortality and morbidity of women in menopause. PMID- 10514955 TI - A multicenter investigation of consultation-liaison psychiatry in Italy. Italian C-L Group. AB - In order to evaluate the extent and quality of consultation-liaison (C-L) activity in Italy, a multicenter investigation was conducted in 17 general hospitals. All of the hospitalized patients referred to C-L psychiatry during a 1 year period were assessed by means of a specific instrument (Patient Registration Form, PRF-SF). Of 518,212 patients, 4182 were referred to C-L services (referral rate = 0.72%). Typical consultations were for female patients (60.1%), admitted to medical wards (71.5%), aged 55-75 years. Most interventions were carried out within 2 days; a minority (22%) were urgent requests. Gastrointestinal and cardiovascular disorders, and unexplained medical symptoms were the most frequent ICD-9 somatic diagnoses at admission. One-third of the patients were not informed of having been referred to C-L and half of them had a lifetime history of psychiatric disturbances. Most frequent ICD-10 psychiatric diagnoses were neurotic, stress-related, and somatoform syndromes (33.1%), affective syndromes (19.4%), and organic mental syndromes (10.7%). Two-thirds of the patients were given only one consultation whereas the reminder received two to four follow-up visits. The rate of transfer to psychiatric wards was low (2.1%). Psychopharmacological treatment was suggested in 65% of cases, and 75.5% of the patients were referred to community psychiatric care at discharge. The implications of the findings are discussed. PMID- 10514957 TI - [The use of coronary risk charts]. PMID- 10514958 TI - Beneficial impact of isoflurane during coronary bypass surgery on troponin I release. AB - BACKGROUND: Experimental studies indicate that isoflurane, a commonly used volatile anesthetic, mimics the cardioprotective effects of ischemic preconditioning, probably through ATP-sensitive K+ (KATP) channel activation. The aim of this study was to evaluate the impact of isoflurane during coronary bypass surgery (CABG) on troponin I release. MATERIAL AND METHODS: Forty consecutive patients with chronic stable angina and multivessel disease undergoing isolated CABG were randomized to a control (16 men and 4 women, aged 51 to 73 years, mean 62) or isoflurane (15 men and 5 women, aged 51 to 77 years, mean 65) group before aortic cross-clamping and cardioplegia. Serum levels of troponin I and creatine kinase (CK)-MB, as markers of ischemic injury, were obtained at 24 hours after CABG. Regional wall motion score and left ventricular ejection fraction (LVEF) at transthoracic echocardiography were assessed 5 days postoperatively. Comparisons between groups were performed in the entire population and, subsequently, in those patients with preoperative LVEF < 50%. RESULTS: There were no significant differences between isoflurane-treated patients and controls in cross-clamp time (49 +/- 14 vs 51 +/- 13 min, p = ns), peak values of troponin I (0.9 +/- 0.7 vs 1.4 +/- 1.3 ng/ml, p = ns) and CK-MB (62 +/- 27 vs 64 +/- 27 U/l, p = ns), or postoperative echocardiographic score (26 +/- 7 vs 22 +/- 5, p = ns) and LVEF (53 +/- 10 vs 55 +/- 7%, p = ns). When the comparisons were restricted to those patients with preoperative LVEF < 50%, at 24 hours the isoflurane-treated patients exhibited a smaller release of troponin I and of CK-MB than controls (1.1 +/- 0.7 vs 2.3 +/- 1.3 ng/ml, p = 0.03, and 39 +/- 10 vs 57 +/- 22 U/l, p = 0.04, respectively). CONCLUSIONS: Isoflurane reduces myocardial injury in patients with impaired left ventricular function undergoing CABG; thus, it can be safely used as an additional cardioprotective tool during routine CABG in high risk patients with poor left ventricular function. PMID- 10514959 TI - Factors influencing outcome after emergency surgical repair of acute type A aortic dissection. AB - OBJECTIVE: We retrospectively reviewed our more recent experience with acute type A aortic dissection in order to identify possible risk factors influencing current surgical results. METHODS: Between January 1990 and January 1998, 122 patients (86 males and 36 females; mean age 60 +/- 12 years) underwent emergency repair of acute type A aortic dissection using a standard surgical approach. Seventy-four (61%) patients required isolated replacement of the dissected ascending aorta, 27 (22%) required additional replacement of the aortic arch and 21 (17%) required total aortic root replacement. Surgical outcome was evaluated in terms of operative mortality and morbidity. Results of patients presenting with preoperative complications (Group C) (i.e. cardiac tamponade, cerebral stroke, cardiogenic shock, acute myocardial infarction, anuria or visceral ischemia) were compared with those of uncomplications cases (Group U) and with a calculated risk of expected operative mortality (EOM-rate) based on an analysis of each patient set of preoperative risk factors. Sixteen preoperative and 18 perioperative variables were also analyzed to identify conditions influencing morbidity and mortality. RESULTS: Fifty-seven patients (47%) presented with preoperative complications (Group C) and 65 (53%) did not (Group U). Overall operative mortality was 22% (27 patients). Mortality within subgroups was 40 and 6% for complicated and uncomplications cases, respectively (p < 0.001). The 85% of the overall mortality occurred in Group C patients. During the experience, the operative mortality rate actually observed ranged from 0 to 38% and was similar to the calculated expected risk, thus proving a direct relationship with the amount of complicated cases operated on each year. Multivariate analysis revealed that older age and hemopericardium significantly increased the risk of operative death, while male gender, preoperative complications, postoperative bleeding, duration of circulatory arrest and aortic cross-clamp time significantly predicted morbidity (p = 0.02). CONCLUSIONS: Current results of emergency repair of acute type A aortic dissection are strictly dependent on the number of complicated cases referred for operation. Earlier diagnosis and prompt referral before development of preoperative complications appear essential to improve surgical results. PMID- 10514960 TI - Percutaneous transmyocardial revascularization with holmium laser in patients with refractory angina: a pilot feasibility study. AB - BACKGROUND: Percutaneous transluminal myocardial revascularization (PTMR) is a new procedure to improve perfusion of the ventricular wall for patients with intractable angina that is untreatable by surgery or conventional catheter-based intervention. PTMR allows the creation of myocardial channels through the controlled delivery of holmium laser energy from the ventricular chamber. Preliminary studies in animals and human subject have yielded promising results. We now report the feasibility study of PTMR using a laser delivered through a novel Eclipse system, and we present the results of this sole therapy in patients with severe coronary disease and angina refractory to maximal medical treatment angina (III-IV CCS). METHODS: Percutaneous vascular access for PTMR treatment was obtained via the femoral artery. A 9F directional catheter carrying flexible fiber optics was used with a holmium laser (Eclipse system) and was placed across the aortic valve into the left ventricle cavity to create channels with a depth of 5 mm from the endocardial surface into the myocardial tissue. From April to November 1998, 15 patients underwent PTMR with Eclipse system. Two patients were female; the mean age was 66 +/- 8 (range 59-74). Five patients had a severe LV dysfunction (FE < 30%). Preoperative angina class was III in 10 patients and IV in 5 and previous myocardial procedures had been performed in all patients. RESULTS: The procedure was well tolerated and procedural success was obtained in 14 of 15 patients. There was one myocardial perforation because of guiding catheter manipulation (pericardial drainage in fourth day). We performed a mean of 13 +/- 4 channels in a mean fluoro time of 23 +/- 11 min. Upon release and during follow-up (5.3 months +/- 4.2, range 2-10), angina class had significantly improved in 14 of 14 patients with complete PTMR treatment, with 4 asymptomatic patients, 6 patients in CCS I, 3 in CCS II, 2 in CCS III and only one patient hospitalized due to angina. CONCLUSION: This pilot study confirmed the safety and technical feasibility of PTMR. Immediate and short-term results confirm that a clinical improvement is obtained in most patients. Although these are early clinical benefits, the true efficacy of this approach will necessarily be defined by a randomized trials with prospectively-defined endpoints and with PTMR compared with medical therapy. PMID- 10514961 TI - Cardiac papillary fibroelastoma: report of two cases. AB - We report 2 cases of cardiac papillary fibroelastoma in aortic position that were successfully treated by complete surgical excision, without damage to the aortic valve. This rare cardiac tumor can be associated with serious embolic or hemodynamic complications and therefore, prompt surgical intervention is required. PMID- 10514962 TI - Radiofrequency ablation in two patients with typical and atypical atrioventricular nodal reentrant tachycardias, associated with atrioventricular reentrant tachycardias. AB - The incidence of dual atrioventricular nodal physiology in patients with Wolff Parkinson-White syndrome is quite frequent, but arrhythmia related to an accessory pathway and atrioventricular nodal reentrant tachycardia (AVNRT) in a single patient is less common. Two of our cases (patients aged 24 and 19 yrs) presented the rare evidence of both typical and atypical AVNRTs, associated in the first case with two other atrioventricular reentrant tachycardias (AVRTs), and in the second case with a single AVRT. Both underwent radiofrequency catheter ablation of the slow nodal pathway and of the accessory pathways in a single session, without any complications. After a 3-month follow-up, they were free from symptoms suggestive of tachycardia, without any antiarrhythmic treatment. PMID- 10514963 TI - [Intravascular echocardiography (ICUS) diagnosis of post-traumatic coronary dissection involving the common trunk. A case report and review of the literature]. AB - A case is reported in which a 31-years-old man experienced coronary artery dissection with an acute anterior myocardial infarction following blunt chest trauma in a car accident. Due to ECG signs of acute myocardial infarction a coronary angiography was performed showing an abrupt occlusion of the mid part of the left anterior descending artery and a linear filling defect in the proximal portion of the vessel. Additional detailed intravascular ultrasound was performed, revealing a long intimal tear involving the left anterior descending artery and the left main. The patient underwent immediate coronary artery bypass surgery. Two vein grafts were made from aorta to the left anterior descending artery and the circumflex artery, respectively. Repeat angiography was performed early after the operation; dissection of the left main and the left anterior descending artery was still visible and the grafts to the left descending artery and the circumflex were patent. PMID- 10514964 TI - [Evidence-based medicine and the problem of freedom]. PMID- 10514965 TI - [What is evident is not always true. Epistemological reflections on evidence based medicine]. PMID- 10514966 TI - [One medicine]. PMID- 10514967 TI - [Effects on vasomotor tone and hemostatic function of radiologic contrast media used during invasive cardiological procedures]. AB - There are several types of radiologic contrast media which can be used in invasive cardiology: 1) ionic media with high osmolality (2000 mOsm/kg, about 6 times that of plasma), the prototype of which is diatrizoate; 2) ionic media with low osmolality (600-900 mOsm/kg), the prototype of which is ioxaglate; 3) non ionic monomeric, low-osmolality media, such as iopromide, iopamidol and iohexol; and 4) non-ionic dimeric media, iso-osmolal compared to plasma (290-300 mOsm/kg), among which the most used is iodixanol. Non-ionic media--by far the most expensive--have a generally better tolerability profile for the patient, since they clearly induce a less gastro-intestinal, renal, hemodynamic, electrophysiological and pseudo-allergic side effects. They interfere much less with the physiology of vascular and circulating blood cells, and have lesser negative interference on the hemostatic function. Whether these lesser anti hemostatic properties become a possible downside in situations at high thrombotic risk, such as in some interventional procedures, has been so far the object of isolated reports, usually with limited numbers of patients, and is presently being verified in adequate clinical trials. Radiologic contrast media also have disparate vasoactive properties on epicardial and myocardial resistance vessels. The knowledge and awareness of such effects is of potential importance for the performance of studies requiring the accurate quantitative evaluation of coronary diameters or of myocardial blood flow. PMID- 10514968 TI - [Concerning guidelines]. PMID- 10514970 TI - [Standards and VRQ for ergometry laboratories. Ad hoc commission. National Association of Hospital Cardiologists (ANMCO). Italian Society of Cardiology (SIC). Italian Group of Functional Evaluation and Rehabilitation of Cardiopathies (GIVFRC)]. PMID- 10514969 TI - [ANMCO-SIC-GIVFRC guidelines on cardiological rehabilitation]. PMID- 10514971 TI - [Standards and VRQ in cardiological rehabilitation. Ad hoc commission. National Association of Hospital Cardiologists (ANMCO). Italian Society of Cardiology (SIC). Italian Group of Functional Evaluation and Rehabilitation of Cardiopathies (GIVFRC)]. PMID- 10514973 TI - On the possibility of shear-driven chromatography: a theoretical performance analysis. AB - The use of shear forces for the generation of the mobile phase flow in chromatographic separations is proposed. This novel chromatographic operating principle, referred to as shear-driven chromatography (SDC), completely circumvents the pressure-drop limitation of conventional pressure-driven GC and LC without affecting the operational flexibility (choice of mobile and stationary phases, possibility of solvent and/or temperature programming, etc.). In the present paper, the expression for the height equivalent to a theoretical plate in SDC in a channel with a flat rectangular cross-section is established and is used to demonstrate the large gain in analysis speed under LC, GC and supercritical fluid chromatography conditions. PMID- 10514972 TI - Retention models for ions in chromatography. AB - Since chromatography of ions is a widely used technique in analytical chemistry a basic understanding of the retention mechanism is important. The principles of the different retention models that have been proposed are examined in this paper. The focus is on those models that are derived from the physical chemistry of charged surfaces immersed in an electrolyte solution. In the first two sections the theory for the electrical double layer and the Donnan potential are presented together with experimental results from surface and colloid chemistry. In Section 3 a comparison between stoichiometric and non-stoichiometric models is made. In this section the physical meaning of the retention factor is also examined. The Donnan model and the different double layer models developed for ion exchange chromatography of small ions are discussed in Section 4. The next section presents the corresponding models that have been developed for ion pair chromatography and compares them with the experimental findings. The theoretical modifications needed when going from small ions to ionic macromolecules are discussed in the last section and the developed models are compared with the experimental results. PMID- 10514974 TI - Optimization of throughput and desorbent consumption in simulated moving-bed chromatography for paclitaxel purification. AB - In simulated moving-bed (SMB) applications, throughput and desorbent consumption are two key factors that control process cost. For a given adsorbent volume and product purity requirements, throughput and desorbent consumption depend on desorbent composition, column configuration, column length to diameter ratio, and adsorbent particle size. In this study, these design parameters are systematically examined for paclitaxel purification. The results show that if adsorbent particle size, column dimensions and column configuration are fixed, the higher the product purity required, the lower the throughput. If product purity and yield are fixed, the larger the solute migration speed ratio, the higher the throughput, and the lower the desorbent consumption. If total bed volume and product purities are fixed, the longer the separation zones, the higher the throughput, but the higher the desorbent flow-rate. An intermediate configuration gives the minimum desorbent consumption. If there are no limits on pressure drop or zone flow-rate, the larger the column length to diameter ratio, the smaller the adsorbent particle size, the higher the throughput, and the lower the desorbent consumption. If the maximum zone flow-rate is controlled by the pressure drop limit and not by the standing waves requirement, the longer the columns, the lower the zone flow-rates and the lower the throughput. For 150 microns adsorbent particles and a maximum zone flow-rate of 300 ml/min, a design with optimal throughput and desorbent consumption is found for paclitaxel purification. PMID- 10514975 TI - Application of experimental design for the characterisation of a novel elution system for high-capacity anion chromatography with suppressed conductivity detection. AB - A novel elution system for the application of high-capacity anion exchangers with suppressed conductivity detection in ion chromatography is presented. The ternary elution system is based on perchloric acid, sodium hydroxide and sodium carbonate. The novel elution system was applied to a self-made high-capacity anion-exchange column (Q = 453 mu equiv. Cl-). A central composite design with 20 experiments was used to investigate the influence of the eluent compounds, which varied from 0.2 to 1.0 mM HClO4, 20 to 100 mM NaOH and 0 to 20 mM Na2CO3, on the retention factor k' of seven common anions. A quadratic model including interactions was postulated. The model equations were used to estimate retention factors at known eluent compositions. No significant differences of calculated and experimental retention factors were found. Further statistical analysis was done by analysis of variance. The results showed that the three eluent compounds have completely different effects on the retention behaviour of the anions investigated. The elution of soft and weakly hydrated anions like bromide and nitrate is strongly influenced by the content of perchloric acid, whereas strongly hydrated anions like fluoride and sulphate are mainly affected by the hydroxide or carbonate content of the eluent. The results can qualitatively be explained comparing the relation of ionic radii to charge for eluent and analyte anions. As a consequence, the retention of the anions investigated can easily be manipulated by varying the contents of the eluent compounds. The usefulness of the novel elution system and high-capacity anion chromatography is demonstrated by the determination of trace anions in phosphate and fluoride matrices. PMID- 10514976 TI - Eriochrome Black T as a post-column reagent for the ion chromatographic determination of rare earths. AB - The use of Eriochrome Black T in an alkaline, 40% methanol solution was found to be appropriate as post-column reagent for the determination of rare earths by ion chromatography. Detection of individual lanthanides and lanthanum was carried out at 512 nm and 650 nm after separation by dynamic cation exchange chromatography with gradient elution on C18 column and employing a solution containing alpha hydroxyisobutiric acid/sodium octanesulfonate at pH 3.8 as eluent. The effect of the presence of micelles in the post-column reagent was studied. Sensitivities obtained by the addition of the cationic surfactants cetylpyridinium chloride (CPC) and hexadecyltrimethylammonium bromide (CTAB) were lower than those measured without surfactant addition. In some cases, the signal was totally suppressed. No change in sensitivity was observed with non-ionic (Triton X-100) or anionic (sodium dodecylsulphate, SDS) surfactants but a slight improvement in the baseline noise was observed with the SDS. An evaluation of the influence of chemical and operational variables on the post column reaction (PCR) reagent was carried out either by spectrophotometric tests or by chromatographic experiments. A comparison was performed between three PCR reagents: Eriochrome Black T and xylenol orange in the presence of a cationic surfactant and arsenazo III. Calibration response was linear up to an analyte concentration of 5.0 micrograms ml-1. Absolute detection limits lower than 7 and 17 ng were obtained at the detection wavelengths of 650 nm and 512 nm respectively, for all the natural lanthanides and lanthanum. PMID- 10514977 TI - Comparative study of divalent metals and amines as silanol-blocking agents in reversed-phase liquid chromatography. AB - In this work we compare the silanol-blocking ability of different alkaline earth metal cations, including calcium, magnesium and barium, and strong amine silanol blockers, such as triethylamine and octylamine, using six basic probe solutes at pH 7 on a conventional octadecylsilane phase. Some amines are better blocking agents than the metal cations but this varies with the amine and analyte structure. Among the metals, barium is the best blocker. For certain solutes barium is as effective at blocking silanols as some of the amine blockers. It produces short retention times and good peak shapes with satisfactory peak symmetry factors. However, amines with long alkyl chains, such as octylamine, are better blocking agents than barium. Peak symmetry is still poor for some solutes even in the presence of the strongest blocking agent in the eluent. PMID- 10514978 TI - Macroporous poly(glycidyl methacrylate-triallyl isocyanurate-divinylbenzene) matrix as an anion-exchange resin for protein adsorption. AB - A novel macroporous poly(glycidyl methacrylate-triallyl isocyanurate divinylbenzene) matrix was prepared by a radical suspension copolymerization. The matrix contained epoxy groups, so diethylaminohydroxypropyl groups were coupled to the matrix, leading to an anion-exchange resin. We studied the components, surface and pore structures of the anion-exchange resin by Fourier transform infared spectroscopy and scanning electron microscopy (SEM). SEM observations showed that the resin abounded in macropores as large as 3 to 8 microns both in the surface and the interior. The back-pressure of the column packed with the resin was modest even at a high flow-rate (60.2 cm/min). Then, bovine serum albumin (BSA) was used as a model protein to examine the adsorption properties of the anion-exchange resin. The results showed that under optimum conditions the resin had a capacity as high as 22.8 mg BSA/g wet resin, or 68.7 mg/g dry resin. The adsorbed protein could be desorbed by increasing the liquid phase ionic strength. Most importantly, the matrix had little nonspecific adsorption for BSA before introducing the ion-exchange groups. PMID- 10514979 TI - Capillary electrochromatography with a silica column with a dynamically modified cationic surfactant. AB - A novel mode of capillary electrochromatography (CEC), called dynamically modified silica-capillary electrochromatography, is described in this paper. The column packed with bare silica was dynamically modified with long chain quaternary ammonium salt, cetyltrimethylammonium bromide (CTAB), which was added into the mobile phase. CTAB ions were adsorbed onto the surface of bare silica, and the resulted hydrophobic layer on the silica gel was used as the stationary phase. Using the dynamically modified silica column, neutral solutes were separated by CEC. The highest number of theoretical plates obtained was about 71,500/m and the relative standard deviations for t0 and capacity factor of toluene were 4.7% and 4.9% for 20 consecutive runs, respectively. The separation mechanism of neutral solutes and the influence of mobile phase composition on the separation was investigated. The separation of nitrogen-containing solutes was carried out with this mode and the peak tailing of basic solute was effectively eliminated because the adsorption of basic solute on silica was blocked by the preferred adsorption of CTAB. PMID- 10514980 TI - Separation of oxidized and deamidated human growth hormone variants by isocratic reversed-phase high-performance liquid chromatography. AB - Reversed-phase high-performance liquid chromatography (RP-HPLC) was utilized for the separation of recombinant human growth hormone (hGH) variants on a C18 silica column at 55 degrees C using an isocratic mobile phase which contained 27% 1 propanol in a 25 mM potassium phosphate buffer, pH 6.5. Three of the obtained peaks were characterized by tryptic mapping and mass spectrometry; two of the peaks were found to contain oxidized hGH (dioxy Met14/Met125 and Met125 sulfoxide) while the third contained a deamidated form (Asn149-->Asp149 or Asn152 ->Asp152). Compared to the European Pharmacopoeia RP-HPLC method of hGH analysis, this new method gives two additional peaks and a 50% reduction in the analysis time. PMID- 10514981 TI - Reproducibility of the separation of astaxanthin stereoisomers on Pirkle covalent L-leucine and D-phenylglycine columns. AB - The reproducibility of the separation of astaxanthin stereoisomers on columns packed with Pirkle covalent L-leucine chiral stationary phase (CSP) was examined by comparing six columns purchased from the same manufacturer. Differences were found even for columns packed with CSP from the same lot. The reproducibility of columns packed with Pirkle covalent D-phenylglycine CSP was also examined by comparing columns purchased from the same manufacturer as well as from different manufacturers. Significant differences were found for columns packed by different manufacturers. Chiral column-to-column reproducibility for complex stereoisomeric separations should therefore not be taken for granted. PMID- 10514982 TI - Determination of polyphenols by high-performance liquid chromatography with inhibited chemiluminescence detection. AB - A chemiluminescence reaction detector was developed for the detection of polyphenols separated by HPLC based on the inhibition of chemiluminescence from the luminol-potassium hexacyanoferrate(III) reaction by polyphenols. The separation was carried out on a RP-C18 column at 37 degrees C by using stepwise gradient elutions. The detection limits are in the range of 6.8 x 10(-7)-2.0 x 10(-9) g/ml for catechol, protocatechuic acid, chlorogenic acid, rutin, resorcinol, hydroquinone and p-tert.butylpyrocatechol. The method is sensitive, selective, fast and simple. It has been successfully applied to the determination of chlorogenic acid and rutin in real tobacco samples. PMID- 10514983 TI - Fractionation of apple procyanidins by size-exclusion chromatography. AB - Oligomeric constituents of apple procyanidins were fractionated by size-exclusion chromatography using a TSKgel Toyopearl HW-40F column. The best separation was obtained using a mobile phase of acetone-8 M urea (6:4; adjusted to pH 2) at a flow-rate of 1.0 ml/min. In this chromatographic system, the use of 8 M urea in the mobile phase resulted in a molecular sieve effect without any surface affinity interaction between the gel beads and the procyanidin molecules. Each fraction obtained was examined by reversed-phase high-performance liquid chromatography and time-of-flight mass spectrometry. The order of elution of the procyanidins from the column was coincident with their degree of polymerization. PMID- 10514984 TI - Determination of 20 underivatized proteinic amino acids by ion-pairing chromatography and pneumatically assisted electrospray mass spectrometry. AB - A qualitative determination of 20 underivatized proteinic amino acids by LC-MS is reported. The need for chromatographic separation before mass spectrometry determination is demonstrated based on the study of several amino acid pairs which have some similar characteristics. Two suitable LC-MS systems are proposed for amino acid analysis. A preliminary optimization of these systems has been investigated using evaporative light scattering detection as these two detection modes have the same chromatographic requirements. The amino acid separation was achieved on a Purospher RP-18e or a Supelcosil ABZ+Plus column with tridecafluoroheptanoic acid or pentadecafluorooctanoic acid as volatile ion pairing reagent in an acetonitrile-water mobile phase. In order to elute the most retained amino acids, an elution gradient based on simultaneously increasing the concentration of acetonitrile and decreasing the concentration of the ion-pairing reagent was used. The detection limits of the present work (without specialized optimization) varied from 0.5 to 1 mg 1(-1). PMID- 10514985 TI - Renaturation of heterodimeric platelet-derived growth factor from inclusion bodies of recombinant Escherichia coli using size-exclusion chromatography. AB - A procedure for renaturation of heterodimeric platelet-derived growth factor (PDGF-AB) from inclusion bodies of recombinant Escherichia coli using size exclusion chromatography is described. Either prepurified or crude PDGF-AB inclusion bodies solubilized with guanidinium hydrochloride were subjected to buffer exchange from denaturing to renaturing conditions during chromatography. Renaturation of PDGF-AB involves folding of the solubilized and unfolded molecules into dimerization competent monomers during size-exclusion chromatography and subsequent dimerization of folded monomers into the biologically active heterodimeric growth factor. Optimized conditions result in an overall yield of 75% active PDGF-AB with respect to size-exclusion chromatography and subsequent dimerization. The described approach allows renaturation at high protein concentrations and circumvents aggregation which is observed when refolding is carried out by dilution. PMID- 10514986 TI - Comparison of capillary electrochromatography with high-performance liquid chromatography for the analysis of pirimicarb and related compounds. AB - The applicability of capillary electrochromatography (CEC) to the analysis of pirimicarb and structurally related pyrimidines has been investigated. Methods were developed to improve the separation of closely related compounds. Resolution was achieved both by the use of running buffers containing a mixture of two organic modifiers to increase selectivity and reduce retention times. Solvent composition step gradients were used to separate compounds of widely differing retention factors. A comparison has been made between HPLC and CEC using identical separation parameters and the same stationary phase, from which two important conclusions are drawn. First, it has been shown that values of k' for the compounds analyzed were the same in both techniques. Secondly, although it is evident that CEC produces higher efficiencies than HPLC when running buffers with high organic solvent content are used, as the aqueous content of the running buffer is increased the efficiencies achieved in CEC and HPLC converge until they become equivalent. This is contrary to the theoretical model which predicts efficiencies are inherently higher using electrically rather than pressure driven flow. Disadvantages of the limited control of flow-rate in CEC in comparison with HPLC, are shown. PMID- 10514987 TI - Determination of indole-3-acetic acid, tryptophan and other indoles in must and wine by high-performance liquid chromatography with fluorescence detection. AB - The development of a robust method to analyse the content of tryptophan and of indole-3-acetic acid at the microgram per litre level in must and wine is necessary in order to study the formation of 2-aminoacetophenone and of other indole compounds causing the 'untypical ageing off-flavour'. The present paper discusses the development and validation of a reversed-phase high-performance liquid chromatography method with fluorescence detection for the analysis of indole-3-acetic acid, tryptophan, tryptophol, indole and skatole in must and white wine. The required selectivity and sensitivity was gained through the solid phase extraction on a polystyrene-based polymer column. PMID- 10514988 TI - Determination of folate vitamers in food and in Italian reference diet by high performance liquid chromatography. AB - A trienzyme treatment (conjugase, alpha-amylase, protease) followed by affinity chromatography and reversed-phase HPLC with UV and fluorescence detection was performed for the quantification of folate vitamers in legumes (chickpea and beans), processed meats (salami Milano and Parma ham) and in an Italian reference diet. This method allowed a good separation of six folate vitamers: 5 methyltetrahydrofolate, 5-formyltetrahydrofolate, folic acid, 10-formylfolic acid, 10-formyldihydrofolate and tetrahydrofolate within 30 min. Recovery, reproducibility and limits of detection of the method are reported. HPLC results were 24-52% lower than the microbiological assay findings. PMID- 10514989 TI - Application of ion-exchange cartridge clean-up in food analysis. II. Determination of benzylpenicillin, phenoxymethylpenicillin, oxacillin, cloxacillin, nafcillin and dicloxacillin in meat using liquid chromatography with ultraviolet detection. AB - A multiresidue analytical method was developed for the simultaneous determination of benzylpenicillin (PCG), phenoxymethylpenicillin (PCV), oxacillin (MPIPC), cloxacillin (MCIPC), nafcillin (NFPC) and dicloxacillin (MDIPC) in meat. The method involves the use of an ion-exchange cartridge for sample clean-up followed by ion-pair high-performance liquid chromatography with ultraviolet detection. The recoveries of PCG, PCV, MPIPC, MCIPC, NFPC and MDIPC from pork muscle spiked at levels of 0.5, 0.1 and 0.05 mg/kg were in the range of 77-90, 73-95 and 80-93% with coefficients of variation of 0.5-1.7, 1.6-4.4 and 3.2-6.6%, respectively. For beef muscle spiked at levels of 0.5, 0.1 and 0.05 mg/kg, the recoveries of these compounds were 83-92, 71-86 and 77-90% with coefficients of variation of 1.7-4.4, 2.6-7.0 and 3.9-6.4%, respectively. The detection limits for each penicillin were 0.02 mg/kg in meat. PMID- 10514991 TI - Determination of inorganic anions by ion chromatography using a graphitized carbon column dynamically coated with cetyltrimethylammonium ions. AB - The simultaneous determination of inorganic anions by ion chromatography using a dynamically coated graphitized carbon column with cetyltrimethylammonium (CTA) ions was investigated with suppressed conductivity detection. Column preparations with CTA and sodium carbonate-sodium hydrogencarbonate concentration in the eluent were examined to optimize the separation of seven common anions (F-, Cl-, NO2-, Br-, NO3-, HPO(4)2- and SO(4)2-). Calibration curves were linear from 0.5 to 5 micrograms/ml for F-, from 1.0 to 10 micrograms/ml for Cl-, from 1.5 to 15 micrograms/ml for NO2-, from 2.0 to 20 micrograms/ml for Br- and NO3-, from 5.0 to 50 micrograms/ml for HPO(4)2- and from 3.0 to 30 micrograms/ml for SO(4)2- with correlation coefficients (r) of 0.999 or better. The relative standard deviations of peak areas were between 0.3 and 0.9% for 10 repeated measurements. The application of this newly developed method was demonstrated by the determination of inorganic anions in the water for pharmaceutical purposes. Using CTA-Br as the coating agent, a permanently coated ion-exchange column was obtained, which allowed efficient separations of seven anions without adding any coating agent to the eluent. PMID- 10514990 TI - Liquid chromatographic determination of benzocaine and N-acetylbenzocaine in the edible fillet tissue from rainbow trout. AB - A method was developed for determining benzocaine and N-acetylbenzocaine concentrations in fillet tissue of rainbow trout. The method involves extracting the analytes with acetonitrile, removing lipids or hydrophobic compounds from the extract with hexane, and providing additional clean-up with solid-phase extraction techniques. Analyte concentrations are determined using reversed-phase high-performance liquid chromatographic techniques with an isocratic mobile phase and UV detection. The accuracy (range, 92 to 121%), precision (R.S.D., < 14%), and sensitivity (method quantitation limit, < 24 ng/g) for each analyte indicate the usefulness of this method for studies characterizing the depletion of benzocaine residues from fish exposed to benzocaine. PMID- 10514992 TI - Ion chromatographic determination of cyanate in saline gold processing samples. AB - An ion chromatographic method was developed for the determination of cyanate (CNO ) in saline gold processing samples. The method is based on the use of a very weak-eluting buffer (5 mM sodium borate) and a Dionex AS4A-SC anion-exchange column. This weak-eluting buffer facilitates the wide chromatographic separation of chloride (Cl-) from CNO-. After CNO- has been eluted, the switch to 1.8 mM Na2CO3-1.7 mM NaHCO3 buffer allows the fast elution of other major inorganic and organic anions. Validation of this method, including identification of interferences, has shown that this method is reliable, accurate, sensitive (detection limit, 0.1 mg/l CNO-) and reproducible. PMID- 10514993 TI - Capillary columns with in situ formed porous monolithic packing for micro high performance liquid chromatography and capillary electrochromatography. AB - Capillary columns with monolithic stationary phase were prepared from silanized fused-silica capillaries of 75 microns I.D. by in situ copolymerization of divinylbenzene either with styrene or vinylbenzyl chloride in the presence of a suitable porogen. The porous monolithic support in this study was used either directly or upon functionalization of the surface to obtain a stationary phase that was appropriate for the separation of peptides by capillary electrochromatography (CEC). The main advantages of monolithic columns are as follows. They do not need retaining frits, they do not have charged particles that can get dislodged in high electric field, and they have relatively high permeability and stability. Whereas such columns are designed especially for CEC, they find application in micro high-performance liquid chromatography (mu-HPLC) as well. Five different porogens were employed to prepare the monolithic columns that were examined for permeability and porosity. The flexibility of fused-silica capillaries was not adversely affected by the monolithic packing and the longevity of the columns was satisfactory. This may also be due to the polymerization technique, which resulted in a fluid-impervious outer layer of the monolith that precluded contact between the fused-silica surface and the liquid mobile phase. For the most promising columns, the conductivity ratios and the parameters of the simplified van Deemter equation, both in mu-HPLC and CEC, were evaluated. It was found that the efficiency of the monolithic columns in CEC was significantly higher than in mu-HPLC in the same way as observed with capillary columns having conventional particulate packing. This is attributed to the relaxation of band-broadening with electroosmotic flow (EOF) with respect to that with viscous flow. It follows then that the requirement of high packing uniformity to obtain high efficiency may also be relaxed in CEC. Angiotensin-type peptides were separated by CEC with columns packed with a monolithic stationary phase having fixed n-octyl chains and quaternary ammonium groups at the surface. Plate heights of about 8 microns were routinely obtained. The mechanism of the separation is based on the interplay between EOF, chromatographic retention and electrophoretic migration of the positively charged peptides. The results of the complex migration process, with highly nonlinear dependence of the migration times on the organic modifier and the salt concentration, cannot be interpreted within the framework of classical chromatography or electrophoresis. PMID- 10514994 TI - Understanding of peak deterioration in hyphenated speciation systems due to gas liquid separation in the hydride generation interface. AB - In hyphenated speciation systems with a hydride generation interface one of the processes influencing peak deterioration is gas-liquid separation. A mathematical model was developed to calculate attenuation, signal tailing and resolution loss of HPLC peaks due to gas-liquid separation. It was shown experimentally--using an HPLC-hydride generation-atomic fluorescence spectrometry system for arsenic speciation--that the mathematical model predicts peak deterioration well. This allowed us to study the parameters influencing the deterioration, viz. gas-liquid separation parameters (gas-liquid separator head space volume and purge gas volume flow-rate) and HPLC peak parameters [width (ratio) and resolution] theoretically, simulating HPLC peaks with gaussian functions. PMID- 10514995 TI - Adaptive numerical morphological filter for identifying chromatographic signals. AB - A morphological algorithm has been proposed to filter away impulsive noises confounded in the chromatographic signal. Compared with the conventional median filtering method, the results showed that the proposed method has the advantages of a better filtering effect and less distortion. In particular, the morphological filter with adaptive scale gives very good results. PMID- 10514996 TI - Capillary column supercritical fluid chromatography-atmospheric pressure ionisation mass spectrometry interface performance of atmospheric pressure chemical ionisation and electrospray ionisation. AB - A supercritical fluid chromatography interface probe for atmospheric pressure ionisation mass spectrometry (API-MS) with the advantage of convenient switch between ionisation modes [atmospheric pressure chemical ionisation (APCI) and electrospray ionisation (ESI)] has recently been reported [P.J.R. Sjoberg, K.E. Markides, J. Chromatogr. A, 785 (1997) 101]. In order to obtain a stable ion signal and a low minimum detectable quantity, the design of the spray devise has to be optimised. For easy optimisation in the APCI mode, the corona needle was mounted directly on the interface probe. To compensate for the adiabatic cooling of the expanding mobile phase in the APCI mode, a heated region around the restrictor tip was used. In comparison, ESI required no additional heat, which might also prevent fragmentation for thermolabile compounds. As the mobile phase used was neat CO2, a low flow of make-up liquid was utilised in the ESI mode for transfer of the analytes from the expanding CO2 gas to the liquid phase before ionisation. The low make-up liquid flow in the ESI mode was sufficient for preventing the restrictor from becoming blocked. Factors that influence the ion signal intensity and stability have been studied. In APCI mode, corona needle position, nebuliser gas flow and gas additives were studied and in ESI mode, spray capillary assembly dimension and position, liquid flow-rate and composition were studied. The achievable detection limits were in the 50-0.1 pg (i.e., 290 fmol-140 amol) range. The detection limit in APCI mode was improved by a factor of about 20-25 compared to an earlier design [L.N. Tyrefors, R.X. Moulder, K.E. Markides, Anal. Chem. 65 (1993) 2835]. PMID- 10514998 TI - A simple assay for formate dehydrogenase activity by gas chromatography-mass spectrometry. AB - A new method using GC-MS was devised for the convenient measurement of formate dehydrogenase (FDH) activity in crude tissue samples. FDH activity was detected by measuring headspace 13CO2, which was enzymically converted from [13C]formic acid. This method proved to be sensitive and simple for the estimation of FDH activity without complicated pretreatment. PMID- 10514997 TI - Capillary electrophoresis separation and permanganate chemiluminescence on-line detection of some alkaloids with beta-cyclodextrin as an additive. AB - Capillary electrophoresis has been used in combination with on-line permanganate chemiluminescence detection for the simultaneous determination of morphine, 6 monoacetylmorphine and heroin. It was found that beta-cyclodextrins could improve the separation efficiency and enhance the chemiluminescence signal. Improved sensitivity over capillary electrophoresis with UV detection was obtained. The procedure has detection limits of 23, 66 and 115 fmol for morphine, 6 monoacetylmorphine and heroin, respectively. PMID- 10514999 TI - Identification of 2,3-butanedione monoxime hydrogenation products by gas chromatography-mass spectrometry in an ion trap mass spectrometer. AB - The reduced products of 2,3-butanedinone monoxime by reaction with hydrogen in the presence of homogeneous catalysts were identified by gas chromatography coupled to an ion trap mass spectrometer operating either in the electron impact or chemical ionization mode. The major hydrogenation products were found to be several heterocyclic nitrogen-containing compounds: tetramethylpyrazine, 2,4 dimethyl-3-ethylpyrrole, 3,4,5-trimethylpyrazole, 2,5-dimethyl-1-propylpyrrole, 3 acetyl-2,4-dimethylpyrrole, 3,5-dimethyl-4-allypyrazole and tetramethylpyrazine N monoxide. PMID- 10515000 TI - Visual signalling by asymmetry: a review of perceptual processes. AB - Individual levels of asymmetry in traits that display fluctuating asymmetry could be used as visual signals of phenotypic (and perhaps genotypic) quality, as asymmetry can often be negatively related to fitness parameters. There are some data to support this hypothesis but the experimental protocols employed have commonly resulted in asymmetries far larger than those observed in nature. To date, there has been little consideration of the ability of animals to accurately discriminate small asymmetries (of the magnitude observed in the wild) from perfect symmetry. This is key to assessing the plausibility of the asymmetry signalling hypothesis. Here, I review the perceptual processes that may lead to the discrimination of asymmetry and discuss a number of ecologically relevant factors that may influence asymmetry signalling. These include: signal orientation, distance of trait elements from the axis of symmetry, trait complexity, trait contrast and colour, and the behaviour of both signaller and receiver. I also discuss the evolution of symmetry preferences and make suggestions as to where researchers should focus attention to examine the generality of asymmetry-signalling theory. In highly developmentally stable signalling systems the magnitude of asymmetry may be too small to be detected accurately and reliably, hence asymmetry signalling is unlikely to have evolved in these situations. PMID- 10515001 TI - Prospects for the clinical application of neural transplantation with the use of conditionally immortalized neuroepithelial stem cells. AB - Although neural transplantation has made a relatively successful transition from the animal laboratory to human neurosurgery for the treatment of Parkinson's disease, the use of human embryonic brain tissue as the source of transplants raises difficult ethical and practical problems. These are likely to impede the widespread use of this otherwise promising therapy across the range of types of brain damage to which the results of animal experiments suggest its potential applicability. Various alternative approaches are reviewed briefly, aimed at developing sources of tissue for transplantation that can be maintained in vitro until needed, so obviating the requirement for fresh embryonic tissue at each occasion of surgery. Particularly promising are conditionally immortalized neuroepithelial stem cell lines in which the immortalizing gene is downregulated upon transplantation into a host brain. We describe experiments from our laboratory with the use of cells of this kind, the multipotent MHP clonal cell lines, derived from the developing hippocampus of a transgenic mouse harbouring a temperature-sensitive oncogene. Implanted into the hippocampus of rats and marmosets with damage to the CA1 cell field, the MHP36 line gave rise to healthy surviving grafts and to essentially complete recovery of cognitive function. Postmortem study of the implanted rat brains indicated that MHP36 cells migrate to the region of damage, adopt both neuronal (pyramidal) and glial phenotypes in vivo, and reconstitute the normal laminated appearance of the CA1 cell field. We have previously shown that, when primary differentiated foetal tissue is used as the source of grafts in rats with CA1 damage, there is a stringent requirement for replacement with homotypic CA1 cells. We interpret our results as showing that the MHP36 cell line responds to putative signals associated with damage to the hippocampus and takes up a phenotype appropriate for the repair of this damage; they therefore open the way to the development of a novel strategy with widespread applicability to the treatment of the diseased or damaged human brain. PMID- 10515002 TI - A brief history of human autosomes. AB - Comparative gene mapping and chromosome painting permit the tentative reconstruction of ancestral karyotypes. The modern human karyotype is proposed to differ from that of the most recent common ancestor of catarrhine primates by two major rearrangements. The first was the fission of an ancestral chromosome to produce the homologues of human chromosomes 14 and 15. This fission occurred before the divergence of gibbons from humans and other apes. The second was the fusion of two ancestral chromosomes to form human chromosome 2. This fusion occurred after the divergence of humans and chimpanzees. Moving further back in time, homologues of human chromosomes 3 and 21 were formed by the fission of an ancestral linkage group that combined loci of both human chromosomes, whereas homologues of human chromosomes 12 and 22 were formed by a reciprocal translocation between two ancestral chromosomes. Both events occurred at some time after our most recent common ancestor with lemurs. Less direct evidence suggests that the short and long arms of human chromosomes 8, 16 and 19 were unlinked in this ancestor. Finally, the most recent common ancestor of primates and artiodactyls is proposed to have possessed a chromosome that combined loci from human chromosomes 4 and 8p, a chromosome that combined loci from human chromosomes 16q and 19q, and a chromosome that combined loci from human chromosomes 2p and 20. PMID- 10515003 TI - The Croonian Lecture 1999. Intracellular membrane traffic: getting proteins sorted. AB - The secretory and endocytic pathways within higher cells consist of multiple membrane-bound compartments, each with a characteristic composition, through which proteins move on their way to or from the cell surface. Sorting of proteins within this system is achieved by their selective incorporation into budding vesicles and the specific fusion of these with an appropriate target membrane. Cytosolic coat proteins help to select vesicle contents, while fusion is mediated by membrane proteins termed SNAREs present in both vesicles and target membranes. SNAREs are not the sole determinants of target specificity, but they lie at the heart of the fusion process. The complete set of SNAREs is known in yeast, and analysis of their locations, interactions and functions in vivo gives a comprehensive picture of the traffic routes and the ways in which organelles such as the Golgi apparatus are formed. The principles of protein and lipid sorting revealed by this analysis are likely to apply to a wide variety of eukaryotic cells. PMID- 10515004 TI - [Is there progress in the chemotherapy of small cell lung cancer?]. AB - Small cell lung cancer (SCLC) is a frequent neoplastic disease which has shown an increasing incidence during recent years, particularly in women. Because of the high sensitivity of chemotherapy, remission is usually achieved with standard regimens. However, relapses are common and 5-year survival is < 10%. Over the last few years intensive chemotherapy with or without stem cell support has been developed and has brought about an improvement of overall survival in SCLC patients. This therapeutic approach is reviewed. PMID- 10515005 TI - [Diagnostic and prognostic evaluations of sterility and the Sierre Hospital. 389 cases]. AB - AIM OF THE STUDY: The management of infertile couples at regional hospital level is rarely reported and is generally less known, whereas the activity of university infertility clinics is usually the reference. We evaluate the diagnostic approach to infertile couples at the Regional Hospital of Sierre. METHOD: The medical charts of 389 couples presenting with infertility between 1989 and 1996 were reviewed. The investigations and interpretation of results were always performed by the same investigator. RESULTS: A lower pregnancy rate was observed in the case of a pathological postcoital test and when proximal tubal occlusion or phimosis were diagnosed by hysterosalpingography or laparoscopy. In our population, age of the female partner > 40 years, duration of infertility of more than 36 months and multifactorial, male and female, aetiology of infertility were significantly associated with a poor prognosis (p < 0.001). For the assessment of tubal patency, hysterosalpingography and laparoscopy with chromopertubation showed an acceptable level of agreement (kappa coefficient = 0.58). Hysterosalpingography does not provide a precise diagnosis in peritoneal and ovarian pathology (72.9% sensitivity, 55.7% specificity). A third of pregnancies were obtained without treatment during the diagnostic study of the menstrual cycle, or after hysterosalpingography and laparoscopy with chromopertubation. CONCLUSION: Most diagnostic procedures for infertility can be performed in a regional hospital by an infertility specialist collaborating closely with a university centre. When infertile couples are transferred to a university centre their evaluation is simplified, as they have already completed an initial systematic investigation and treatment plan at the regional hospital. This two-step approach may be beneficial in selecting couples who need specific treatment not available in a regional hospital. PMID- 10515006 TI - [A female patient with splenomegaly, interstitial pneumopathy and giant foam cells in bone marrow]. AB - We describe a case of Niemann-Pick disease type B. A 13-year-old female adolescent of Turkish origin suffered from abdominal pain for several months, finally leading to hospitalisation. The investigations revealed splenomegaly and interstitial pneumopathy. The bone marrow contained giant foam cells typical of Niemann-Pick disease. Enzymatic analysis of a fibroblast culture confirmed the diagnosis of Niemann-Pick disease type B, with reduced activity of acid sphingomyelinase. Niemann-Pick disease is an inherited autosomal recessive lysosomal storage disorder of sphingolipids, resulting in an accumulation of sphingomyelin in the cells of the reticulo-histiocytic system due to an enzymatic defect. In Niemann-Pick disease type B the spleen and lung are the main organs affected. At present no treatment exists for this disorder. PMID- 10515007 TI - [New aspects of interdisciplinary therapy for malignant gliomas in adults]. AB - Malignant gliomas are the most frequent primary brain tumours in adults. Their histopathological grade (WHO) and histological subtype (astrocytoma, oligodendroglioma, mixed tumour) is associated with biological behaviour and clinical outcome. Throughout the course of glioma an interdisciplinary approach is mandatory. Surgery and radiation therapy are standard procedures. Oligodendroglial tumours are chemosensitive and molecular biological markers can predict this sensitivity. Prediction of chemosensitivity in astrocytic tumors is not available yet. We propose a multimodal therapeutic approach and refer to ongoing clinical trials. PMID- 10515008 TI - [Yellow nail syndrome]. PMID- 10515010 TI - The influence of lumbar disc height and cross-sectional area on the mechanical response of the disc to physiologic loading. AB - STUDY DESIGN: The influence of lumbar disc height and cross-sectional area on the mechanical response of the disc to physiologic loading was determined using a finite element model. OBJECTIVES: To identify which geometric characteristics are potentially related to motion segment mechanical response to applied load, such as flexibility, fiber stress, disc bulge, and nucleus pressure. SUMMARY OF BACKGROUND DATA: The height and area of the lumbar disc varies within the disc itself, between disc levels, between people, between men and women, with aging, and during the day. Mechanical theory dictates that the height and area influence the mechanical response of the disc to loading. This could have important consequences in risk of injury. METHODS: Three-dimensional finite-element models representing three disc heights (5.5 mm, 8.5 mm, and 10.5 mm) and three disc areas (1060 mm2, 1512 mm2, and 1885 mm2) were generated. The effect of disc geometry on the mechanical properties of the disc were studied for four moment loads (magnitude, 7.5 Nm) with compressive preload (400 N) and for three different direct forces. Commercially available finite-element software was used. RESULTS: Discs with a ratio of small disc area to disc height were more prone to larger motion, higher anular fiber stresses, and larger disc bulge. When the disc height alone was increased by a factor, its flexibility also increased, either by the same amount or by a much larger ratio. CONCLUSIONS: Discs with the most height and smallest area are exposed to much higher risk of failure than other combinations of disc height and geometry. PMID- 10515009 TI - Experimental spinal fusion using sintered bovine bone coated with type I collagen and recombinant human bone morphogenetic protein-2. AB - STUDY DESIGN: Posterolateral lumbar transverse process fusion using recombinant human bone morphogenetic protein (rhBMP)-2 carried by sintered bovine bone and Type I collagen complex was compared with fusion achieved using autogeneous bone graft or sintered bovine bone alone. OBJECTIVES: This study examined the efficacy of sintered bovine bone coated with Type I collagen as a carrier of rhBMP-2 for lumbar intertransverse process arthrodesis. SUMMARY OF BACKGROUND DATA: Posterolateral intertransverse process arthrodesis using osteoinductive growth factors is performed experimentally in the lumbar spine. The previous studies revealed the efficacy of osteoinductive factors applied to carriers having no bony structures, such as collagen sheet or polylactic acid polymer, for the spinal fusion. However, in their studies, a large amount of osteoinductive proteins have been applied for the spinal fusion. We used the sintered bovine bone "True Bone Ceramics" (TBC; Koken Co., Tokyo, Japan) coated with type I collagen as the carrier. True Bone Ceramics is the only biomaterial possessing a natural trabecular structure and an organized crystal of bone minerals. METHODS: Twenty-two adult rabbits underwent bilateral lumbar intertransverse process arthrodesis at L4-L5. The animals were divided into four groups and had materials implanted as follows: autologous bone group, grafted autologous corticocancellous bone harvested from the posterior iliac crest; implanted TBC group; TBC collagen group, implanted TBC coated with Type I collagen infiltrating into the porous space; and BMP group, implanted sintered bovine bone coated with Type I collagen infiltrated with 100 micrograms of rhBMP-2. Spinal fusion was evaluated by radiographic analysis, manual palpation, biomechanical testing, and histologic examination 6 weeks after surgery. RESULTS: Two rabbits were killed because of infection and lumbar plexus palsy. Radiographs of the BMP group showed a homogeneous fusion mass at the intertransverse area, and stability was confirmed by dynamic radiographs at 3 and 6 weeks after surgery. In the BMP group, a bony mass in the intertransverse area was more prominent than in the other groups. The BMP group had a higher fusion rate based on manual palpation than the-other groups, and BMP fusions showed significantly higher tensile strength and stiffer fusion. The histologic findings in the BMP group demonstrated membranous bone and endochondral bone formations between the transverse process and the fusion mass. In the other groups, continuous trabecular bone formation was observed in the area surrounding the transverse process, but gaps between grafted fragments and less mature bone formation were present in the intertransverse area. CONCLUSIONS: Sintered bovine bone coated with Type I collagen and rhBMP-2 resulted in a higher fusion rate than the autograft and can be used as a carrier for rhBMP-2 in spinal fusion. PMID- 10515011 TI - Anterior shear of spinal motion segments. Kinematics, kinetics, and resultant injuries observed in a porcine model. AB - STUDY DESIGN: A basic study of 56 porcine specimens in anterior shear loading. OBJECTIVES: To determine some modulators of the biomechanics of spinal motion segments exposed to acute shear loading and to identify the resultant injuries. SUMMARY OF BACKGROUND DATA: Most research on spinal injury mechanisms has focused on compressive loading, leaving a void in understanding of the effect of shear loading on origin of injury. METHODS: Cervical spines (n = 56) of domestic pigs (6 months old) were loaded to failure in a specially designed jig that restricted their motion to primarily the shear plane. The specimens were tested at load rates of 100 N/sec or 10,810 N/sec and either in a flexed or neutral posture. In addition, the function of the individual structures of the motion segment were determined by serial dissection forming three groups: whole specimens, specimens with no posterior ligaments, and specimens with no posterior ligaments or facet joints. Load-deformation curves were collected using analog-to-digital sampling rates of 50 and 100 Hz. The mode of failure was then documented through systematic dissection of the specimen and/or radiology techniques. Modeling approaches were then used to gain insight into the failure mechanisms. RESULTS: Dynamic loading (10,810 N/sec) and flexion of the specimens were found to increase the ultimate load at failure when compared with quasistatic loading (100 N/sec) and neutral postures. The disc resisted up to 70% of an applied load, with the pars interarticularis responsible for only 30% of the load. Nonetheless, the pars was the primary site of failure. Furthermore, higher load rates also caused endplate avulsion, specifically in the lateral borders of the anulus. CONCLUSIONS: The porcine model appears to reproduce injuries found in the data available on human lumbar material. Fractures in the pars interarticularis may not greatly weaken the joint, given the dominant role of the disc, but compromise its normal kinematics. Clinically, this may explain the occurrence of pars fractures, without total disability. PMID- 10515012 TI - Use of percutaneous transpedicular external fixation pins to measure intervertebral motion. AB - STUDY DESIGN: Direct measurement of intervertebral motion was compared to motion determined by measuring the position of the exposed ends of the external fixation pins. OBJECTIVES: To verify the accuracy of this technique, so that this protocol can be used to study intervertebral motion in the clinical setting. SUMMARY OF BACKGROUND DATA: The transpedicular external fixation test has been shown to be a test that can predict the outcome of spinal fusion. In patients who are candidates for this test, intervertebral motion can be calculated from motion at the external ends of these pins. METHODS: Six fresh cadaveric spinal segments from L2 to L5 were instrumented with titanium Schanz screws. Reflective markers were placed on the tips of the pins, and intervertebral motion was measured using a noncontacting camera system. Computed tomography data were used to determine the position of the vertebra relative to the reflective markers. Intervertebral distances were calculated and compared with direct measurements obtained using a three-dimensional digitizing arm. RESULTS: There was an excellent correlation (r2 = 0.931) between the directly measured intervertebral motions and those that were indirectly calculated from measurements of motion at the end of the Schanz screws. CONCLUSIONS: Intervertebral motion can be measured by monitoring motion of the ends of transpedicular external fixation pins. Motion of anatomic landmarks on the vertebrae can be calculated from the pin end's motion if computed tomography data are used to determine the geometric relation between the vertebrae and the external fixation pins. This validation study supports the use of this method in clinical investigations of intervertebral motion in patients with low back pain and external fixation. PMID- 10515013 TI - Differential biomechanical effects of injury and wiring at C1-C2. AB - STUDY DESIGN: An in vitro study compared the biomechanics of the upper cervical spine among three groups of cadaveric specimens, each with a different source of instability: transverse-alar-apical ligament disruptions, odontoid fractures, or odontoidectomies. The responses of the three groups were again compared after a uniform posterior cable and graft fixation was applied to the specimens. OBJECTIVES: To quantify and compare the effects of different injuries on atlantoaxial stability and to determine whether a single fixation technique effectively treats each injury. SUMMARY OF BACKGROUND DATA: Previous biomechanical studies of atlantoaxial instability have been focused on mechanisms of injury or on comparison among fixation types. METHODS: Cables and pulleys applied torques to human cadaveric C0-C6 specimens quasistatically while an optical system tracked three-dimensional angular and translational motion at C0 C1 and C1-C2. Specimens were tested immediately after injury, after posterior cable and graft fixation, and after 6000 cycles of fatigue. RESULTS: Odontoidectomies increased C1-C2 angular and translational range of motion significantly more than odontoid fractures or ligament disruptions, especially during flexion-extension. Odontoid fractures produced a slightly larger increase in C1-C2 angular range of motion than ligament disruptions but a smaller increase in C0-C1 range of motion. The different injuries affected the lax zone and the position of C1-C2 axis of rotation differently. Restabilization by posterior cable and graft reduced motion only moderately for each injury type. All three fixated injuries were susceptible to loosening from fatigue. CONCLUSION: The three different injuries produce different spinal biomechanical responses. To best promote fusion, posterior cable and graft fixation should be used with an adjunctive stabilizing technique to treat all three injuries. PMID- 10515014 TI - Impact of the type of brace on the quality of life of Adolescents with Spine Deformities. AB - STUDY DESIGN: A group of 102 brace-treated adolescents, aged 10-19 years with spine deformities participated in a cross-sectional study. OBJECTIVES: To determine the effect of various types of orthoses on self-perceived health status. SUMMARY OF BACKGROUND DATA: Spinal orthosis is an effective means of controlling progressive scoliosis, but bracing has shown a considerable impact on several aspects of adolescent functioning. METHODS: Skeletally immature patients with spine deformities (75% with idiopathic scoliosis) who visited consecutively for routine biannual follow-up evaluations of orthotic treatment were studied. Twenty-five patients used the Milwaukee brace, 30 the Boston brace, 13 the thoracolumbosacral orthosis (TLSO), and 34 the Charleston bending orthosis. Patients completed the Quality of Life Profile for Spine Deformities (QLPSD), a specific instrument that measures quality of life in five areas labeled psychosocial functioning, sleep disturbances, back pain, body image, and back flexibility. Higher QLPSD scores mean a high level of impairment of quality of life. RESULTS: Milwaukee brace-treated patients scored significantly higher than Boston brace-treated and TLSO-braced patients and patients with Charleston bending orthosis in the overall QLPSD score (mean +/- SD, 53.60 +/- 13.26 vs. 45.65 +/- 12.81 and 42.79 +/- 12.99, respectively) and in back flexibility and psychosocial functioning. Other quality-of-life-related variables selected in multivariate analysis were the Risser sign, clinical diagnosis, duration of brace treatment, and degrees of correction. CONCLUSION: In cases of different orthoses of proven similar effectiveness in controlling the scoliotic curves, the use of bracing with the lowest impact on the quality of life should be recommended. PMID- 10515015 TI - Late-developing infection in instrumented idiopathic scoliosis. AB - STUDY DESIGN: This is a retrospective review of all patients requiring either Cotrel-Dubousset or Moss Miami rod removal. All initial spinal instrumentations were for adolescent idiopathic scoliosis from 1985 through 1994. Twenty-two patients who underwent rod removal for late-developing infection constitute the study group. OBJECTIVES: To determine the bacteriology and treatment of patients with late-developing infection after posterior spinal instrumentation for scoliosis. SUMMARY OF BACKGROUND DATA: There have been conflicting reports regarding this entity, some reporting a high percentage of positive cultures and others a low yield. The latter have attributed the entity to fretting corrosion. Much literature describes late appearance of infection with large foreign bodies (implants). Glycocalyx, a membrane that surrounds bacteria adjacent to surgical implants, results in poor antibiotic penetration, poor macrophage action, and difficulty in culturing bacteria. METHODS: One thousand two hundred forty-seven patients who underwent posterior instrumentation from 1985 through 1994 were reviewed. Those requiring implant removal were further studied. Those with late developing infection (more than 1 year after the initial procedure) were further reviewed. Culture reports, presence of pseudarthrosis, and antibiotic regimen after implant removal were the primary parameters studied. RESULTS: Twenty-two patients (1.7%) experienced development of late infection a mean of 3.1 years after the initial procedure. In specimens from these patients cultured only 72 hours, only 1 of 10 was positive. Of those cultured for 7-10 days (the last 12) 11 were positive, usually for low-virulence skin organisms. After surgery, patients received antibiotics parenterally for 48 hours and orally for 7 days. All wounds were closed primarily. Four patients had pseudarthroses, two underwent revised procedures with titanium implants without signs of infection at more than 2 years' follow-up. CONCLUSIONS: Late-appearing infection with spinal instrumentation can be treated with device removal, primary skin closure, and short-term oral antibiotics. The infections affect soft tissue, not the bone. PMID- 10515016 TI - The clinical significance of the high-intensity zone on lumbar spine magnetic resonance imaging. AB - STUDY DESIGN: Prospective observational study of anular tears, diagnosed by a high-intensity zone within the anulus on lumbar spine magnetic resonance imaging, and correlation with the clinical features. OBJECTIVES: To assess the prevalence of high-intensity zones in patients who are investigated for back and leg pain and to determine whether there are clinical features that can be used for diagnosis of the presence of a high-intensity zone. SUMMARY OF BACKGROUND DATA: Results in previous studies have shown that the presence of a high-intensity zone is associated with reproduction of a patient's pain on stress discography. Neural compression on magnetic resonance imaging has been shown to be associated with back pain, but to date, no study has correlated the presence of a high-intensity zone with the clinical features. METHODS: The lumbar spine magnetic resonance images in 156 patients in whom back and leg pain were investigated were analyzed for the presence and appearances of high-intensity zones. The clinical features of those patients with a high-intensity zone but with no evidence of neural compression on magnetic resonance imaging were analyzed by t test and X2 test. RESULTS: A high-intensity zone occurred in patients at a prevalence of 45.5% and usually occurred posteriorly (77%) and posterolaterally (22%) within the anulus. There were no features within the history, functional disability questionnaire, or physical examination that aided in a clinical diagnosis of those patients with a high-intensity zone. CONCLUSIONS: A high-intensity zone is a common finding in patients in whom low back and leg pain are investigated, but the presence of a high-intensity zone does not define a group of patients with particular clinical features. PMID- 10515017 TI - Spinal curvatures and quality of life in women with vertebral fractures secondary to osteoporosis. AB - STUDY DESIGN: A prospective cross-sectional case-control study. OBJECTIVES: To compare spinal curvatures in women with osteoporosis and control subjects with a new instrument, the curviscope. SUMMARY OF BACKGROUND DATA: Few instruments are available for measuring spinal curvatures in the sagittal plane. Most of them have poor reproducibility, and they have been poorly investigated in osteoporosis. METHODS: Ninety-eight postmenopausal women were evaluated. They were divided into two groups, according to their bone status: women with osteoporosis with at least one vertebral fracture (n = 51) and control subjects (n = 47). Women with osteoporosis were divided into two subgroups, according to the delay since the last vertebral fracture had occurred (i.e., more or less than 3 months). Quality of life was assessed by using a generic instrument, the Nottingham Health Profile, in patients with osteoporosis only. RESULTS: Reproducibility of the curviscope was satisfactory. For kyphosis measurements, the coefficients of variation were 2.8% and 2.4% in control subjects and women with osteoporosis, respectively. Kyphosis values were significantly higher in women with osteoporosis than in age-matched control subjects (63 degrees +/- 13 degrees vs. 52 degrees +/- 11 degrees, respectively; P < 0.005). Nottingham Health Profile scores were significantly different (P < 0.05) in women with osteoporosis with a recently diagnosed vertebral fracture, compared with other women with osteoporosis in two aspects, physical mobility and energy. Kyphosis measurements were significantly correlated with age in the whole group (r = 0.26; P < 0.05). In the Nottingham Health Profile, physical mobility was significantly correlated with kyphosis (r = 0.35; P < 0.05). CONCLUSIONS: The curviscope is a reliable tool, particularly useful in the assessment of osteoporosis. Moreover, kyphosis angles measured with the curviscope are markedly increased in women with osteoporosis, compared with control subjects. Finally, an increase of kyphosis angles is associated with decreased physical mobility. PMID- 10515018 TI - Evaluation of pedicle screw position in thoracic and lumbar spine fixation using plain radiographs and computed tomography. A prospective study of 35 patients. AB - STUDY DESIGN: This was a prospective study of 35 consecutive patients in whom pedicle screw position was assessed after surgery, using lateral radiographs and computed tomography. OBJECTIVE: To evaluate the accuracy of plain radiographs and computed tomography in assessment of pedicle screw position. SUMMARY OF BACKGROUND DATA: Imaging techniques, such as postoperative anteroposterior and lateral plain radiographs and computed tomography, are currently the primary means of assessing pedicle screw placement. METHODS: Postoperative radiographs and computed tomographic scans were used to evaluate the position of 220 pedicle screws inserted in the spines of 35 consecutive patients who underwent thoracic and lumbar spine fusion and instrumentation. No recognized neurologic complication resulted from pedicle screw placement. Screw position was graded as in, out, or questionable. All observations were performed independently by three observers. The authors also analyzed the position of the screws according to the underlying spinal disease. RESULTS: More misplaced screws were clearly seen on computed tomographic scans than on plain radiographs; however, this difference was not statistically significant. Interobserver differences were not statistically significant. Intraobserver differences approached statistical significance when the results of the two test were compared. CONCLUSIONS: Although the accuracy of computed tomographic imaging is better than that of plain radiographs, the difference does not reach statistical significance. Postoperative use of plain radiographs remains a reliable method for evaluation of pedicle screw insertion in the absence of neurologic deficit. PMID- 10515019 TI - Predictors of time loss after back injury in nurses. AB - STUDY DESIGN: A 2-year prospective inception cohort study of back injury in nurses. OBJECTIVES: To determine the extent to which characteristics of nurses, of the injury, and of the workplace predict occurrence and duration of time loss from work after back injury. SUMMARY OF BACKGROUND DATA: During 2 years, 320 nurses incurred 416 back injuries at a large teaching hospital in Winnipeg, Canada. Nurses injured on preselected wards were targeted for early intervention, including provision of modified work, whereas nurses injured on other wards received the usual care. METHODS: Time loss attributable to the back injury during the 6 months after injury was analyzed. Three statistical models were used to examine occurrence of time loss (logistic regression), duration of time loss (Tobit regression), and duration of time loss once an injury incurring time loss had been documented (least-squares regression). RESULTS: In 218 of the 416 injuries, the injured nurse consented to interview. Whereas perceived disability was related to whether a time loss injury would ensue, self-reported pain was strongly related to the duration of time loss once an injury had become a time loss injury. Duration of time loss was reduced by participation in the return-to work program. Mechanism of injury, specifically injury occurring while lifting patients, resulted in greater time loss. CONCLUSIONS: Focusing on reducing the perception of disability at the time of injury is critical to preventing time loss, but once time loss has occurred, offer of modified work and attention to pain reduction are warranted. The findings add to the evidence that workplace based intervention programs can be effective in reducing the morbidity resulting from back injury. PMID- 10515020 TI - Randomized trial of radiofrequency lumbar facet denervation for chronic low back pain. AB - STUDY DESIGN: A prospective double-blind randomized trial in 31 patients. OBJECTIVES: To assess the clinical efficacy of percutaneous radiofrequency denervation of the lumbar zygapophysial joints in reducing pain, functional disability, and physical impairment in patients with back pain originating from the lumbar zygapophysial joints. SUMMARY OF BACKGROUND DATA: Chronic low back pain is a major health problem in the industrialized world. A treatment option is percutaneous radiofrequency denervation of the lumbar zygapophysial joints. Its clinical efficacy has never been formally tested in a controlled trial. METHODS: Thirty-one patients with a history of at least 1 year of chronic low back pain were selected on the basis of a positive response to a diagnostic nerve blockade and subsequently randomly assigned to one of two treatment groups. Each patient in the radiofrequency treatment group (15 patients) received an 80 C radiofrequency lesion of the dorsal ramus of the segmental nerve roots L3, L4, and L5. Patients in the control group (n = 16) underwent an the same procedure but without use of a radiofrequency current. Both the treating physician and the patients were blinded to the group assignment. Before treatment, physical impairment, rating of pain, the degree of disability, and quality of life were assessed by a blinded investigator. RESULTS: Eight weeks after treatment, there were 10 success patients in the radiofrequency group (n = 15) and 6 in the sham group (n = 16). The unadjusted odds ratio was 3.3 (P = 0.05, not significant), and the adjusted odds ratio was 4.8 (P < 0.05, significant). The differences in effect on the visual analog scale scores, global perceived effect, and the Oswestry disability scale were statistically significant. Three, 6, and 12 months after treatment, there were significantly more success patients in the radiofrequency group compared with the sham group. CONCLUSIONS: Radiofrequency lumbar zygapophysial joint denervation results in a significant alleviation of pain and functional disability in a select group of patients with chronic low back pain, both on a short-term and a long-term basis. PMID- 10515021 TI - Complication, survival rates, and risk factors of surgery for metastatic disease of the spine. AB - STUDY DESIGN: The risk factors for complications and complication and survival rates in patients with metastatic disease of the spine were reviewed. A retrospective study was performed. OBJECTIVES: To determine the surgical complication and survival rates of patients with metastatic disease of the spine and risk factors for complication occurrence. SUMMARY OF BACKGROUND DATA: The role of surgical intervention for patients with metastatic disease of the spine has been controversial. Several risk factors for surgical complications have been identified. Short survival times and high complication rates have failed to justify surgical intervention in many cases. METHODS: Patients (n = 80) undergoing surgical treatment for metastatic disease of the spine were reviewed. Surgical indications included progressive neurologic deficit, neurologic deficit failing to respond to, or progressing after, radiation treatment; intractable pain; radioresistant tumors; or the need for histologic diagnosis. Patients underwent anterior, posterior, or combined decompression and stabilization procedures. Neurologic examination was recorded before surgery, postoperative period, and at least follow-up. Complication and survival rates were calculated. Several variables were examined for risk of complication. RESULTS: The mean age at time of surgery was 55.6 years (range, 20-84 years). Mean survival time after the diagnosis of spinal metastasis was 26.0 months (range, 1-107.25 months). Mean survival time after surgery was 15.9 months (range, 0.25-55.5 months). Sixty-five patients showed no change in Frankel grade, 19 improved one Frankel grade, and 1 deteriorated one Frankel grade; 1 patient had paraplegia. Thirty-five complications occurred in 20 patients (25.0%). Ten patients (12.5%) had multiple complications accounting for 23 of the 35 postoperative problems (65.7%). Sixty patients had no surgical complications (75%). There were no intraoperative deaths. CONCLUSIONS: The likelihood that a complication occurred was significantly related to Harrington classifications demonstrating significant neurologic deficits and the use of preoperative radiation therapy. In general, Harrington classifications with neurologic deficits and lower Frankel grades before and after surgery were associated with an increased risk of complication. Overall, the major complication rate was relatively low, and minor complications were successfully treated with minimal morbidity. The relatively long survival time after spinal surgery in this group of patients justifies surgical treatment for metastatic disease. Most complications occurred in a small percentage of patients. To minimize complications, patients must be carefully selected based on expected length of survival, the use of radiation therapy, presence of neurologic deficit, and impending spinal instability or collapse caused by bone destruction. PMID- 10515022 TI - Surgical treatment of far lateral lumbar disc herniation. Identification of compressed root and discectomy by lateral approach. AB - STUDY DESIGN: A new method is described of compressed root identification and discectomy for extraforaminal disc herniation, by a lateral intertransversalis approach. OBJECTIVES: To describe a safe surgical approach that does not require resection of adjacent bone structures during extraforaminal discectomy. SUMMARY OF BACKGROUND DATA: Most earlier series have reported approaches that damaged bordering bone structures with wide laminoarthrectomy. This is an attempt at a safer, simpler surgical approach. METHODS: Thirteen patients with lateral hernia have undergone this surgical procedure since 1995. Herniectomy was performed after identification of the compressed root within the iliopsoas muscle. RESULTS: All the patients resumed the upright position with the aid of semirigid brace 24 hours after surgery. Upon awakening from the anesthesia, no patient reported peripheral pain. Motor deficits resolved after physical rehabilitation in all but one patient. At a mean follow-up of 14 months, there was no report of back pain. CONCLUSION: The procedure described in this article offers a simple alternative to the valid procedures presently at hand. It offers the advantage of no bone resection and of minimizing nerve structures manipulation. PMID- 10515023 TI - Postoperative lumbar microdiscectomy pain. Minimalization by irrigation and cooling. AB - STUDY DESIGN: Seventy patients undergoing de novo lumbar microdiscectomy were prospectively randomized into a control group and a group in which cold intraoperative wound irrigation along with postoperative wound cooling was used. Postoperative analgesia requirements and length of hospital stay were analyzed and correlated. OBJECTIVES: To evaluate the role of intraoperative cold irrigation and postsurgical cooling in minimizing postoperative lumbar discectomy pain. SUMMARY OF BACKGROUND DATA: Regulated hypothermia has been used frequently in pain reduction; however, the efficacy of such a strategy in lumbar disc procedures has not been established. METHODS: Seventy patients (43 men and 27 women), operated on the first time for lumbar disk herniation were prospectively randomized into two groups. A standard microdiscectomy was performed on all patients. In cohort A the wound site was irrigated with a cold (18 C) 5% bacitracin solution for 5 minutes. Additionally, a cooling microtemperature pump was placed on the wound site for 24 hours after surgery. The patients in the control group (cohort B) were treated in a standard fashion without additional hypothermic therapy. All patients received postoperative analgesia through a self administered morphine pump. The amount of postoperative analgesia received was calculated in morphine equivalents per kilogram. The length of hospital stay was also noted. RESULTS: The total amount of pain medication was significantly smaller in cohort A than in the control group (cohort B). For the statistical analysis of the results, covariate analyses for both the length of hospital stay and the morphine dose were used, demonstrating a statistically significant difference with P = 0.0001. No postoperative wound infection was noted in either group. CONCLUSIONS: Intraoperative and postoperative wound site cooling is a safe, inexpensive, and efficient therapeutic method. It reduces the patients' postoperative pain, promotes earlier ambulation and decreases the length of hospital stay. PMID- 10515024 TI - Complete reduction of retro-odontoid soft tissue mass in os odontoideum following the posterior C1-C2 tranarticular screw fixation. AB - STUDY DESIGN: A case report of os odontoideum with retro-odontoid soft tissue hypertrophy treated by the transarticular screw fixation. OBJECTIVES: To present a case of os odontoideum that showed complete reduction of retro-odontoid soft tissue mass caused by atlantoaxial subluxation after the C1-C2 transarticular screw fixation. SUMMARY OF BACKGROUND DATA: Hypertrophy of the periodontoid soft tissue has been reported to be associated with chronic atlantoaxial subluxation and progressive myelopathy. While the rheumatoid pannus has been reported to become reduced of disappear after fixation of the unstable segment, the reduction of the hypertrophied soft tissue mass has never been reported in atlantoaxial subluxation of nonrheumatoid origin, especially in the case of os odontoideum. METHODS: Posterior C1-C2 transarticular screw fixation was performed in a patient with os odontoideum, who showed signs of progressive myelopathy by the compression of retro-odontoid soft tissue mass and atlantoaxial subluxation. RESULTS: The fixation of atlantoaxial subluxation achieved not only the complete reduction of the retro-odontoid soft tissue mass, but also clinical improvement of the myelopathy. CONCLUSIONS: Posterior atlantoaxial fixation is worth trying in slow progressing myelopathy by the compression of hypertrophy of the soft tissue even in nonrheumatoid atlantoaxial subluxation, thereby obviating the need for direct removal of the mass via the transoral route. PMID- 10515025 TI - [Atherosclerosis: current status and prospects of prevention and treatment]. PMID- 10515026 TI - [Effect of mildronate on life quality of patients with chronic heart failure]. AB - AIM: To study quality of life (QL) of patients with chronic heart failure (CHF) and QL changes resultant from mildronate therapy. MATERIALS AND METHODS: QL was studied in 30 IHD patients with CHF of NYHA class II-IV, ejection fraction > 45%; 40 IHD patients without CHF and 30 healthy subjects. CHF patients were treated for 30 days with oral mildronate (250 mg 4 times a day). QL was assessed according to the method SF-36 Health Status Survey. RESULTS: QL in CHF patients is much lower than that of the controls. Objective severity of the disease and subjective satisfaction with life do not always coincide. Mildronate in a dose 1 g/day per os may be beneficial for LQ of CHF patients. CONCLUSION: It is thought desirable to include QL in analysis of efficiency of managing patients with CHF. SF-36 is a tool able to follow up changes in QL of CHF patients within a short term treatment period. PMID- 10515027 TI - [Cardiac failure treatment with berlipril: effects on hemodynamics, neurohumoral status and activity of free radical lipid peroxidation]. AB - AIM: To test the ability of ACE inhibitor berlipril to control neurohumoral hyperactivation and reestablish balance between oxidant and antioxidant systems in patients with ischemic heart disease (IHD) associated with heart failure (HF). MATERIALS AND METHODS: 145 patients (mean age 51.5 +/- 3.94 years) with IHD class II-III associated with circulatory insufficiency NYHA class II-III received berlipril for 6 weeks. RESULTS: Berlipril treated patients exhibited decreased class of HF, improved left ventricular conductivity, attenuated neurohumoral stimulation, intensity of cell membrane peroxidation, increased plasmic pool of antioxidant enzyme systems. CONCLUSION: 6-week berlipril treatment promoted a pronounced improvement of neurohormonal profile of plasm and recovery of free radical lipid peroxidation which resulted in reduction of HF. PMID- 10515028 TI - [Effect of brerlipril in patients with non-insulin dependent diabetes mellitus and mild or moderate arterial hypertension as indicated by 24-h blood pressure monitoring]. AB - AIM: To study a hypotensive effect of berlipril, an ACE inhibitor, using 24-h BP monitoring in patients with non-insulin-dependent diabetes mellitus (NIDDM) combined with stable mild or moderate arterial hypertension (AH). MATERIALS AND METHODS: 22 NIDDM patients with mild or moderate AH were treated with berlipril. 24-h monitoring of BP was made in all the patients before and 3 months after the treatment. RESULTS: A stable hypotensive effect of berlipril was achieved at its therapeutic dose 1.5 tablets a day once a day or divided into two doses a day. On treatment week 3-4 a hypotensive effect of berlipril enhanced in 5 patients. This may be due to ACE inhibitors action on the tissue component of the reninangiotensin system. CONCLUSION: Berlipril produces a high hypotensive effect and brings about a positive response of insulin-resistance in NIDDM associated with mild and moderate AH. PMID- 10515029 TI - [Ednit influence on oxidation resistance of LDL lipoproteins in hypertensive subjects]. AB - AIM: Evaluation of resistance of low density lipoproteins (LDLP) of the serum to oxidation in patients with essential hypertension (EH) before and after course treatment with ednit. MATERIALS AND METHODS: 10 patients with mild or moderate EH aged 38-58 years were given ednit for 1 or 2 months. 10 healthy males aged 35-48 years served control. RESULTS: EH patients had high baseline levels of lipid peroxidation products in LDLP, while antioxidant resistance of LDPL was diminished by 67% (p < 0.05) compared to controls. Ednit treatment for 1 and 2 months enhanced this resistance in mild and moderate EH (p < 0.05). CONCLUSION: Ednit raises resistance of LDLP to oxidation and as ACE inhibitor lowers arterial pressure. Therefore it can be used in EH. PMID- 10515030 TI - [Lipid and non-lipid effects of enduracin in patients with arterial hypertension]. AB - AIM: To analyze lipid and non-lipid effects of 6-month administration of enduracine in patients with marked dislipoproteinemia suffering from arterial hypertension with ischemic heart disease or without it. MATERIALS AND METHODS: 40 hypertensive patients (27 males and 13 females, mean age 52.43 +/- 1.68 years) entered the study of enduracine effects. Most of them received enduracine for 6 months in a dose 1500 mg/day. Lipids levels were measured in all the patients. Blood flow along major brain arteries was determined at transcranial dopplerography in 23 patients. RESULTS: A 6-month course of enduracine in a dose 1500 mg/day promoted normalization of serum lipid spectrum, vascular tonicity and reactivity of cerebral arteries, produced a mild hypotensive effect. CONCLUSION: Endurance (a long-acting form of nicotinic acid) has favourable lipid and non lipid effects in patients with dislipoproteinemias and arterial hypertension in the presence or absence of ischemic heart disease. PMID- 10515031 TI - [Hypotensive and anti-ischemic efficiency of estulic in course treatment of myocardial infarction complicated by essential hypertension]. AB - AIM: To assess hypotensive and antiischemic activity of estulik (Sandos, Switzerland) in patients with myocardial infarction (MI) complicated by essential hypertension (EH). MATERIALS AND METHODS: 23 MI patients with moderate EH (diastolic blood pressure up to 110 mm Hg) and 25 patients with manifest EH (diastolic blood pressure 115-120 mm Hg) of group 1 and 2, respectively, have received single doses and a 10-day course of estulik in a dose 0.5-1.0 mg/day. Hemodynamic and antiischemic activities were evaluated at echocardiography, coupled veloergometry, Holter monitoring, myocardial scintigraphy, polarography. In 31% of the patients selective coronarography was made. RESULTS: Clinical condition and hemodynamics of group 1 patients have improved by 30-40%, microcirculation has improved by 25-35%. In group 2 patients the drug was found ineffective. CONCLUSION: Estulik is a drug of choice in moderate EH with manifest angina pectoris. PMID- 10515032 TI - [Cerivastatin-a new synthetic 3-hydroxy-3-methylglutaryl (HMG) inhibitor: effect of 0,2 mg dose in patients with primary hyperlipidemias]. AB - AIM: To elucidate efficacy, safety and tolerance of lipobay (cerivastatin), a new HMG-CoA-reductase inhibitor (0.2 mg/day) in patients with primary hyperlipidemia (PHL). MATERIALS AND METHODS: The trial enrolled 15 men aged 21-64 years with PHL of type 2a and 2b. After 1 and 3 months of treatment all the patients underwent a general clinical examination with measurements of blood lipids (total cholesterol, triglycerides, high density lipoprotein cholesterol--HDL-C), apolipoproteins (apo A-1 and apo B). RESULTS: After 3 months of treatment total cholesterol, LDL-C and apolipoprotein B decreased by 24.94 +/- 2.87%, 28.94 +/- 3.08%, 19.32 +/- 2.43% (p = 0.0001), respectively, while LDL-C levels were < 3.4 mmol/l in 4 patients. Triglycerides dropped by 17.84 +/- 6.41%, while HDL-C rose by 5.01 +/- 4.47%, but the changes were not significant. Lipobay in doses 0.2 mg/day was well tolerated. One patient stopped taking the drug because of severe abdominal pain. CONCLUSION: Lipobay is a novel, effective and well-tolerated drug for treatment of patients with primary hyperlipidemia of type 2a and 2b. PMID- 10515033 TI - [Dipyridamole (curantil) treatment of anginal patients: results of treatment for many years]. AB - AIM: To assess effect of curantil optimal doses on clinical condition, coronary reserves, systemic microcirculation and peripheral hemodynamics in patients with stable angina pectoris (AP) given a course or continuous treatment. MATERIALS AND METHODS: A 2-month treatment with curantil with a gradual increase of the daily dose to 0.45 g was given to 261 patients with AP. After that the treatment was continued to 3 years in 38 patients. The coronary reserves were examined with bicycle exercise tests, peripheral hemodynamics and microcirculation were investigated with tachooscillography and conjunctival biomicroscopy. RESULTS: The 2-month curantil treatment was effective in 81% of the patients with AP functional class II-III and 60% of those with AP class 4. Exercise tolerance increased in AP of functional class II-III and IV by 50 and 24%, respectively. Ischemic depression of ECG ST segment induced by the exercise shortened. Prolongation of the treatment to 1 year resulted in further regression of AP and provided a 57% increase in muscular performance. The 3-year treatment maintained the 1-year effectiveness which was associated with lowering of postload and improvement of microcirculation, in particular, a 2-fold decrease in manifestations of sludge. CONCLUSION: The antianginal effect of optimal daily doses of curantil is more potent in angina of effort, grows step-by-step and reaches maximum if continued for a year. A hemodynamic effect of curantil manifests with lowering of postload and improvement of microcirculation. This allows to recommend curantil in combined treatment of chronic coronary heart disease. PMID- 10515034 TI - [Therapeutic potential of glucocorticoids inhalation in bronchial asthma]. AB - AIM: To assess effectiveness and safety of inhalation glucocorticoids (budesonide, beklometasone dipropionate) in bronchial asthma (BA). MATERIALS AND METHODS: 19 BA patients (8 males and 11 females) inhaled glucocorticoids for 9 months. RESULTS: The treatment reduced the need in inhalation of short-acting beta-2-adrenomimetics, episodes of asphyxia occurred less frequently. A significant improvement of bronchial permeability was achieved only after 3 months of therapy. Blood levels of hydrocortisone, concentrations of magnesium, calcium and potassium in the serum, red cells and 24-h urine, serum osteocalcin, lumbar vertebral tissue density 3, 6 and 9 months after the treatment start were almost similar to the baseline. CONCLUSION: Glucocorticoids for inhalation are clinically effective in BA. They should be used for not less than 3 months. Topical steroids in mean therapeutic doses had no negative effects on the levels of hydrocortisone, mineral metabolism and bone tissue. PMID- 10515035 TI - [Importance of microcirculatory disorders in combined treatment of acute pleural empyema and pyopneumothorax]. AB - AIM: To investigate responses to application of quick-frozen or cryosupernatant plasma, heparin and proteinases inhibitors of DIC-syndrome in patients with pleural empyema and pyopneumothorax. MATERIALS AND METHODS: Transfusions of quick frozen or cryosupernatant plasma, injections of heparin and inhibitors of proteinases were used as an adjuvant to conventional treatment in 548 patients with acute pleural empyema and pyopneumothorax. RESULTS: The addition of cryoplasm-antienzyme complex to conventional treatment of acute pleural empyema resulted in reduced number of lethal outcomes compared to the conventional treatment alone (5.9 vs 14.4%, respectively). CONCLUSION: Transfusion of quick frozen or cryosupernatant plasma, injection of heparin and inhibitors of proteinases improve microcirculation around the inflammatory focus. In addition to antibiotic therapy and evacuation of the pus from the pleural cavity, the above methods contribute to better outcomes of pleural empyema and pyopneumothorax. PMID- 10515036 TI - [Nicotinamide in combined treatment of chronic pancreatitis]. AB - AIM: To evaluate effects of nicotinamide on insulin secretion in glucose tolerance test and on blood clotting in patients with chronic pancreatitis (CP). MATERIALS AND METHODS: 30 patients with CP of alcoholic etiology received combined treatment with enzyme medicines. In addition, some of them were administered nicotinamide as 2.5% solution for 2 weeks (1 ml twice a day). Before the treatment and during its course, measurements were made of fasting and post glucose test values of insulin secretion and thromboelastogram. RESULTS: Both basal and stimulated insulin secretion in CP patients was low compared to control subjects. Nicotinamide significantly increased basal secretion of insulin and insignificantly aroused its glucose-stimulated secretion. Nicotinamide promoted reduction of hypercoagulation and time to remission. CONCLUSION: Nicotinamide administration is thought valid for correction of endocrine pancreatic function and hemocoagulation in patients with alcoholic CP. PMID- 10515037 TI - [Choice of anti-helicobacter treatment of gastroduodenal ulcer associated with helicobater pylori]. AB - AIM: Choice of optimal antihelicobacter (AH) treatment of ulcer. MATERIALS AND METHODS: 249 outpatients with duodenal ulcer associated with Helicobacter pylori entered a blind multicenter controlled trial. The patients were given one of the following drugs: metronidazole, tonidazol, amoxicilline, clarithromycin, rovamycin, omeprazole, azitromycin. RESULTS: Of all the drugs used, rovamycin appeared preferable as it has high AH activity, is safe and cost-effective. CONCLUSION: Rovamycin has a high AH activity and can be applied for treatment of diseases associated with Helicobacter pylori as a drug of choice. PMID- 10515038 TI - [Treatment of idiopathic thrombocytopenic purpura in adults: efficacy of domestic intravenous immunoglobulin in immune thrombocytopenia]. AB - AIM: The study of effectiveness of intravenous immunoglobulin (IVIG) in therapy of idiopathic thrombocytopenic purpura (ITP) in adults. MATERIALS AND METHODS: High doses of IVIG (0.2-0.4 g/kg b.w. for 4-6 days) were given to 6 female patients aged 20 to 59 years (median--39 years) with immune thrombocytopenia. 4 patients had primary ITP resistant to glucocorticosteroids (GCS) and 1 female had chronic ITP treated by splenectomy without effect. 1 patient with rheumatoid arthritis developed severe thrombocytopenia combined with agranulocytosis when treated with nonsteroid antiinflammatory drugs. RESULTS: The response was observed in 5 of 6 patients (in 4 patients resistant to GCS and 1 RA patient). In 3 of them the effect was rated as excellent (platelets level > 150,000 per cubic millimeter), in 2 patients it was good (platelets count from 50,000 to 150,000 per cubic millimeter). Splenectomy was performed in 4 cases with ITP on day 8-14 after IVIG therapy. The 2- and 6-month follow-up evidenced for good results of the surgical treatment. The RA patient showed a stable rise of the blood count. Serious side effects of IVIG therapy were not registered. CONCLUSION: IVIG of Russian produce is effective in the treatment of drug-induced thrombocytopenia and ITP resistant to GCS. PMID- 10515039 TI - [Garlic effectiveness in rheumatoid arthritis]. AB - AIM: To perform of clinical trial of alisate--a garlic preparation produced in Russia. MATERIALS AND METHODS: An open controlled trial of alisate enrolled 30 patients with rheumatoid arthritis (RA). 15 patients with RA of varying clinical form, stage and activity were given alisate in a dose 300 mg (1 tablet) twice a day for 4-6 weeks. 15 control RA patients received conventional antirheumatic therapy. RESULTS: The alisate group achieved a good and partial response in 86.5% of cases. The drug was well tolerated and had no side effects. In control group, some parameters changed for the worse. CONCLUSION: Alisate can be recommended for treatment of RA patients in combined and monotherapy. PMID- 10515040 TI - [Treatment of bone disorders in renal diseases]. AB - AIM: To study possible correction of bone disorders (osteopenia, Ca/P-imbalance, bone pain, limited volume of indolent movements) which are still a serious complication associated with renal diseases and pathogenic therapy (steroids). MATERIALS AND METHODS: The bone disorders were treated in 10 uremic hemodialyzed patients (8 men, 2 women; group 1) with vitamin D3 (calcitriol made in Russia) + rhEPO (recormon; Boehringer Mannheim), in 15 patients (15 women, 0 men) with lupus-nephritis (group 2) with vitamin D3 (n = 5, group 2a) or miscalcic (Sandoz) (n = 10, group 2b), in 2 patients (2 men, 0 women) with glomerulonephritis (group 3) with vitamin D3 + miacalcic. Additionally all the patients received Ca salts. In groups 2 and 3 renal function was normal. The duration of the treatment was 3 6 months. RESULTS: In all the groups we obtained an analgetic effect (attenuation of bone pain and more indolent movements), improvement of life quality, diminished need in analgetics, elevation of serum Ca level (p > 0.05). CONCLUSION: Treatment of renal patients with bone affection with vitamin D3 and miacalcic has an analgetic effect, improves life quality. PMID- 10515041 TI - [Role of IL-8 and defensins in pathogenesis of chronic glomerulonephritis and pyelonephritis]. AB - AIM: The study of interleukine-8 (IL-8) and defensines contribution to pathogenesis of chronic glomerulonephritis (CGN) and pyelonephritis (PN). MATERIALS AND METHODS: 122 patients were assigned to three groups: 42 CGN patients with isolated urinary syndrome (group 1); 60 patients with chronic pyelonephritis (CP) with normal nitrogen-excretory function (group 2); 20 patients with CGN and chronic renal failure (CRF) (group 3). 24 healthy volunteers served control. IL-8 and defensines were measured in urine and plasm of all the patients and controls using enzyme immunoassay. RESULTS: IL-8 in urine and plasm of controls was not found, in plasm of patients was found in 45%, the differences between the groups being insignificant. The highest IL-8 urine concentration was found in group 2. It was significantly higher than in group 1 and 3 (p < 0.001). Mean IL-8 urine concentration in patients with secondary pyelonephritis was significantly higher than in those with primary pyelonephritis (p < 0.05). In primary pyelonephritis, urinary IL-8 was higher than in patients of groups 1 and 3 (p < 0.001). Mean urinary IL-8 was significantly higher in patients of group 3 than 1 (p < 0.005). IL-8 urinary concentrations of group 3 patients tended to an increase with growing severity of renal failure. Plasm defensines were present in 46.5% of patients without marked differences between the groups. Urine defensines were the highest in group 2 being significantly higher than in group 1, 3 and controls (p < 0.001). Urine defensines in group 1 were slightly higher than in controls and significantly reduced vs group 3 (p < 0.005). Urine defensines concentrations rose with CGN stage. A strong positive correlation was established between IL-8 and defensines in urine (r = 0.62), leukocyturia and IL-8 in urine (r = 0.52). A weak positive correlation (r = 0.38) existed between proteinuria and urine IL-8 only in group 3 patients. A week later concentrations of IL-8 and defensines were low in group 2. In group 1 they rose, fell or remained unchanged. CONCLUSION: IL-8 and defensines may be of the same pathogenetic importance both in infectious and non-infectious inflammation in the kidneys. Cytotoxic action of defensines can be related to location of the inflammation in the kidney. IL-8 urine tests can be used in monitoring of inflammation activity and diagnosis of latent glomerulonephritis and pyelonephritis. PMID- 10515042 TI - ["Sicilian gambit": pathophysiological approach to drug therapy of arrhythmia (lecture]]. PMID- 10515043 TI - [Combined antihypertensive therapy: review of multicenter studies conducted in foreign countries]. PMID- 10515044 TI - [Current aspects of sandostatin use in gastroenterology and surgery (review of literature for 1993-1998)]. PMID- 10515045 TI - Resonance energy transfer study of hemoglobin complexes with model phospholipid membranes. AB - By examining the resonance energy transfer between fluorescent probes, embedded in the lipid bilayer (4-(dimethylaminostyryl)-1-methylpiridine, 4 (dimethylaminostyryl)-1-dodecylpiridine, N,N'-bishexamethylenrhodamine, rhodamine 6G) as donors, and the heme group of hemoglobin as acceptor, the structure of the protein complexes with the model membranes composed of phosphatidylcholine and cardiolipin was characterized. Quantitative interpretation of the experimental data was performed in terms of the model of energy transfer in two-dimensional systems, using a set of parameters including the distance of closest approach between donor and acceptor, the vertical separation of donor planes, the acceptor distance from the donor plane and the orientation factor. The limits for the heme distance from the lipid bilayer center and the depth of the protein penetration in the membrane interior were estimated. The results obtained suggest that the depth of hemoglobin insertion into liposomal membranes decreases upon increasing CL content in the lipid bilayer. PMID- 10515046 TI - Supramolecular interactions of solid human serum albumin with binary mixtures of solvent vapors. AB - Sorption isotherms of organic compounds on solid human serum albumin (HSA) from binary vapor mixtures were determined by gas chromatographic headspace analysis. The shape of sorption isotherms depends on molecular structure of studied sorbates. The 'active' compounds capable to sorb effectively on dry HSA increase the sorption of 'passive' compounds unable to be sorbed by dry HSA in absence of the third component. The critical hydration of HSA is required for sorption activation of 'passive' sorbates if water is taken as 'active' component. Ethanol and acetonitrile exhibit such activation effect without threshold. 'Passive' sorbates are able to produce cooperative activation effect on the sorption of 'active' component. Hydration history effect is observed for sorption on prehydrated HSA and HSA hydrated in situ. Obtained results were interpreted in terms of clathrate formation by 'passive' sorbate (substrate) and 'active' component inside the HSA (receptor) binding centers. PMID- 10515047 TI - Molecular dynamics of a tetrasaccharide subunit of chondroitin 4-sulfate in water. AB - Molecular dynamics (MD) simulations on a tetrasaccharide subunit of chondroitin 4 sulfate (CS4) in aqueous solution were carried out to study its interactions with water. Pair distribution functions and diffusion coefficients were calculated from a 4 ns trajectory and the hydration of different molecular groups was analysed. The average values of the interglycosidic torsion angles found in the simulations are phi 13 = -10 degrees, psi 13 = -85 degrees and phi 13 = 80 degrees, psi 13 = 90 degrees for the beta-(1-->3) linkage, and phi 14 = -10 degrees, psi 14 = -70 degrees for the beta-(1-->4) linkage. Hydrophobic patches formed by sugar ring CH groups were found. The diffusion coefficients of the water molecules vary from 1.4 x 10(-9) to 2.3 x 10(-9) m2 s-1 depending on the distances between the water molecules and the atoms of the CS4 molecule and the type of CS4 atoms, respectively. Reorientation correlation times of the water molecules in the vicinity of different CS4 atoms were estimated to be about 1 ps at a polymer concentration of 4 wt.% CS4. The number of hydrogen bonds between the water molecules and the acceptor atoms of CS4 was determined to be about 20 per disaccharide unit, indicating a higher hydration ability of chondroitin sulfate in comparison with non-sulfated oligosaccharides. Substructures, where water molecules are involved in hydrogen bonds to different sugar rings, were found, which may be important for the stabilisation of the secondary structure of the CS4 molecule. PMID- 10515048 TI - Solvent-dependent conformational behaviour of lipochitoligosaccharides related to Nod factors. AB - The solution conformation of two lipooligosaccharides related to Nod factors or lipochitoligosaccharides have been analysed by 1D and 2D 1H and 13C NMR spectroscopy, molecular mechanics and dynamics calculations. The obtained data indicate that the glycosidic torsion angles have restricted fluctuations, but may adopt a variety of shapes. Remarkably, the relative orientation of the fatty acid chain towards the oligosaccharide backbone is solvent dependent. In water solution, the acyl residue and the oligosaccharide adopt a quasi-parallel orientation for a significant amount of time. PMID- 10515049 TI - Control of enzymatic degradation of hyaluronan by divalent cations. AB - Enzymatic degradation of hyaluronan (HA) by testicular hyaluronidase (HAase, hyaluronate 4-glucanohydrolase) requires inclusion of mono- or divalent cations in the reaction mixture. Most divalent cations activated HAase with equal potency; however, Cu2+ suppressed degradation, and Ca2+ showed a concentration dependent regulation of size of the oligosaccharide products. Careful selection of HAase assay parameters is critical for discovery of novel HAase inhibitors and for preparation of controlled-size oligosaccharide fragments. PMID- 10515050 TI - Active-latent glycosylation strategy toward Lewis X pentasaccharide in a form suitable for neoglycoconjugate syntheses. AB - Glycosylation of 4-nitrophenyl 2-acetamido-6-O-tert-butyldiphenylsilyl-2-deoxy-1 thio-beta-D-gluc opyranoside with phenyl 2,3,4,6-tetra-O-benzoyl-1-thio-beta-D galactopyranoside in the presence of NIS and TfOH as catalyst gave the lactosamine derivative regiospecifically in high yield. Further 3-O-fucosylation with phenyl 2,3,4-tri-O-benzyl-1-thio-beta-L-fucopyranoside using DMTST as promoter afforded the Lex trisaccharide intermediate. The latent glycosyl donor was transformed into its active form (p-acetamidothiophenyl) by reduction with zinc in acetic acid and N-acetylation. Glycosidation with p-nitrothiophenyl lactoside acceptor in the presence of NIS/TfOH as catalyst gave the Lex pentasaccharide in 71% yield. PMID- 10515051 TI - Scope and limitation of the application of the (methoxydimethyl)methyl group in the synthesis of 2'-O-, 6'-O- and 2',6'-di-O-(alpha-L-arabinofuranosyl)-beta-D galactopyranosyl- (1-->6)-D-galactoses. AB - For the characterisation of the anticipated epitope of an arabinogalactan, isolated from the extract of Echinacea purpurea, the trisaccharide alpha-L-Araf (1-->2)-beta-D-Galp- (1-->6)-D-Gal was synthesized. The easily available 3,4-O isopropylidene-6-O-(methoxydimethyl)methyl-beta-D-galactopyr anosyl- (1-->6) 1,2:3,4-di-O-isopropylidene-alpha-D-galactopyranose having the OH-2' free served as aglycone upon direct arabinosylation at the 2' position under Helferich conditions. The formed HgBr2 cleaved the acid sensitive protecting group at position 6', but under basic conditions the desired, fully protected trisaccharide, or its symmetrical dimerization derivative linked 6'- to 6' position via an isopropylidene bridge, could be obtained. An alternative route involved arabinosylation of a hepta-O-acetylated digalactose with free OH-2'. Two other oligosaccharides, namely, alpha-L-Araf-(1-->6)-beta-D-Galp-(1-->6)-D-Gal and (alpha-L-Araf)2-(1-->2,6)-beta-D-Galp-(1-->6)-D-Gal were also synthesized and characterised. PMID- 10515052 TI - Occurrence of an unusual lactose sulfate in dog milk. AB - The milk of a beagle dog (Canis familiaris) was extracted and fractionated to yield, inter alia, beta-D-Galp3S-(1-->4)-D-Glc (lactose 3'-sulfate), which does not appear to have previously been isolated from milk or other natural sources. The structure was established by 2D NMR spectroscopy and mass spectrometry. By contrast with the milk of some closely related Carnivora, the major constituent of the dog milk was lactose, with minor amounts of 2'-fucosyllactose and sialyl oligosaccharides. PMID- 10515053 TI - Structure of a new acidic O-antigen of Proteus vulgaris O22 containing O acetylated 3-acetamido-3,6-dideoxy-D-glucose. AB - The acidic O-specific polysaccharide of Proteus vulgaris O22 was studied using 1H and 13C NMR spectroscopy, including 2D COSY, TOCSY, NOESY, and H-detected 1H, 13C heteronuclear multiple-quantum coherence (HMQC) experiments, and the following structure for the branched pentasaccharide repeating unit was established: [sequence: see text] where Qui3NAc is 3-acetamido-3,6-dideoxyglucose, O acetylation of QuiNAc at position 4 is stoichiometric and at position 2 nonstoichiometric. Serological relationships of P. vulgaris O22 with some other Proteus strains were substantiated on the level of the O-antigen structures. PMID- 10515054 TI - Synthesis of 3-C-(methyl beta-D-xylofuranosid-3-yl)-5-phenyl-1,2,4-oxadiazole. AB - Nucleophilic addition of KCN to 5-O-benzoyl-1,2-O-isopropylidene-alpha-D-erythro pentofuranos-3-++ +ulose gave the xylo cyanohydrin stereoselectively. Several xylos-3-yl-1,2,4-oxadiazole derivatives were synthesized from this cyanohydrin and were converted into 3-C-(methyl beta-D-xylofuranosid-3-yl)-5-phenyl-1,2,4 oxadiazole. PMID- 10515055 TI - An improved synthesis of an umbelliferyl 5-thioxylopyranoside, precursor of the antithrombotic drug Iliparcil. AB - 4-Ethyl-2-oxo-2H-1-benzopyran-7-yl 2,3,4-tri-O-acetyl-5-thio-beta-D xylopyranoside, a synthetic intermediate of the orally active antithrombotic compound Iliparcil, has been prepared in 44-47% isolated yield. Different conditions were used for the glycosylation of 4-ethyl-2H-7-hydroxy-1-benzopyran-2 one 6 applying 2,3,4-tri-O-acetyl-5-thio-D-xylopyranosyl bromide (2), the analogous beta-chloride 3 or the alpha-trichloroacetimidate 5 as donors. With halides 2 and 3, the reaction was carried out in the presence of ZnO-ZnCl2 or ZnO alone. Both promoters are cheap, safe and therefore compatible with large-scale industrial processes. PMID- 10515056 TI - Synthesis of 5-hydroxy-2-(beta-D-ribofuranosyl)pyran-4-one from a pyranulose glycoside. AB - The synthesis of 5-hydroxy-2-(beta-D-ribofuranosyl)pyran-4-one (9) is described. Treatment of pyranulose glycoside with bromine in carbon tetrachloride afforded brompyranulose glycoside in 90% yield. The reaction of (6S)- and (6R)-4-bromo-6 hydroxy-6-(2,3,5-tri-O-benzoyl-beta-D-ribofuranosyl)-6H- pyran-3-one (2) in acidic media was examined with the following results: the reaction of 2 with trifluoroacetic acid (TFA) in dioxane afforded a mixture of 5-hydroxy-2-(2,3,5 tri-O-benzoyl-beta-D-ribofuranosyl)pyran-4-one (3) and its furan derivative 5 hydroxy-2-{5-(benzoyloxy)methyl]furan-2-yl}pyran-4-one (4), but the use of hydrochloric acid formed the bromofurfural, 3-bromo-5-(2,3,5-tri-O-benzoyl-beta-D ribofuranosyl)-2-furancarboxyal dehyde only. Acetylation of a mixture (3 and 4) with acetic anhydride facilitated product separation to give the corresponding acetates 5-acetoxy-2-(2,3,5-tri-O-benzoyl-beta-D-ribofuranosyl)pyran-4-one (5) and 5-acetoxy-2-{5-[(benzoyloxy)methyl]furan-2-yl}pyran-4-one (6). Treatment of 5 with hydrazine afforded 3-hydroxymethyl-6-(beta-D-ribofuranosyl)-1H-pyridazin-4 one in 43% yield. Debenzoylation of 5 with aq ammonia gave 9 in 50% yield. PMID- 10515057 TI - Structure and cross-reactivity of the O-antigen of Proteus vulgaris O8. AB - A high-molecular-mass O-specific polysaccharide was obtained by mild acid degradation of Proteus vulgaris O8 lipopolysaccharide followed by gel permeation chromatography. Studies of the polysaccharide by sugar and methylation analyses and 1H and 13C NMR spectroscopy, including 2D COSY, TOCSY, NOESY, and H-detected 1H, 13C heteronuclear multiple-quantum coherence (HMQC) experiments, demonstrated the presence of a tetrasaccharide repeating unit having the following structure: [sequence: see text] The role of an epitope associated with the alpha-L-FucpNAc (1-->3)-D-GlcpNAc disaccharide in serological cross-reactivity of P. vulgaris O8 is discussed. PMID- 10515058 TI - Interaction between calcofluor white and carbohydrates of alpha 1-acid glycoprotein. AB - Interactions between the fluorescent probe, calcofluor white, and human serum albumin (HSA) and alpha 1-acid glycoprotein (orosomucoid) are compared. The two proteins have comparable isoelectric points, but alpha 1-acid glycoprotein is highly glycosylated (40% of glycans by weight), while the serum albumin is not. Binding of calcofluor to the proteins induces an increase in both the fluorescence anisotropy and the fluorescence intensity of the fluorophore. Also, we found that the calcofluor exhibits a fluorescence emission with a maximum located at 432, 415 or 445 nm, respectively, in the absence of proteins, in the presence of HSA, and in the presence of alpha 1-acid glycoprotein. The stoichiometries of the calcofluor-serum albumin and calcofluor-alpha 1-acid glycoprotein complexes are 2:1 and 1:1, respectively. The association constants are 0.04 and 0.15 microM-1, respectively. The calcofluor does not interact with Lens culinaris agglutinin (LCA), although the protein has a hydrophobic site. Nevertheless, one cannot exclude that the binding of the fluorophore to the HSA is nonspecific. Our results, when compared with those obtained with calcofluor dissolved in the hydrophobic solvent isobutanol, and with the fluorescent probe, potassium 6-(p-toluidino)-2-naphthalenesulfonate (TNS), bound to alpha 1-acid glycoprotein, indicate that the emission of calcofluor bound to HSA occurs from a hydrophobic state, while that of calcofluor bound to alpha 1-acid glycoprotein occurs from a hydrophilic state. The fluorescence intensity of calcofluor decreases in the presence of carbohydrates isolated from alpha 1-acid glycoprotein, while it increases in the presence of alpha 1-cellulose. Thus, calcofluor interacts mainly with the glycan moiety of alpha 1-acid glycoprotein, and its fluorescence is sensitive to the secondary structure of the glycans. PMID- 10515059 TI - Synthesis of a tetrasaccharide fragment of cobra venom factor oligosaccharide. AB - Synthesis of a tetrasaccharide fragment, alpha-L-Fuc-(1-->3)-beta-D-GlcNAc-(1- >2)-alpha-D-Man-(1-->6)-alpha-D-Man- OMe of the cobra venom factor (CVF) oligosaccharide is described. PMID- 10515060 TI - Synthesis and antitumor activity of 5'-demethyl-5'-trifluoromethyl-daunorubicin and -doxorubicin. AB - The title compounds were prepared by coupling phenyl 4-O-acetyl-2,3,6-trideoxy 6,6,6-trifluoro-1-thio-3-trifluoroacetamido -alpha-L- and beta-L-lyxo hexopyranoside with daunomycinone. The key step of this synthesis is the C trifluoromethylation of a 1-protected 2,3-dideoxy-3-azido-D-erythro pentodialdose, prepared from 2-deoxy-D-erythro-pentose, to give a 6,6,6-trifluoro L-lyxo-hexose derivative. The synthetic products showed higher cytotoxicity than doxorubicin against most of the human tumor cell lines tested in vitro, possibly by the effect of the CF3 group at C-5'. PMID- 10515061 TI - Chemical synthesis of 6"'-alpha-maltotriosyl-maltohexaose as substrate for enzymes in starch biosynthesis and degradation. AB - A branched nonasaccharide 6"'-alpha-maltotriosyl-maltohexaose was synthesised in 40 steps from D-glucose and maltose. Phenyl O-(2,3,4,6-tetra-O-benzyl-alpha-D glucopyranosyl)-(1-->4)-O- (2,3,6-tri-O-benzyl-alpha-D-glucopyranosyl)-(1-->4) 2,3-di-O-benzyl-1-th io- beta-D-glucopyranoside and O-(2,3,4,6-tetra-O-benzyl alpha-D-glucopyranosyl)-(1-->4)-O-(2,3,6-tri- O-benzyl-alpha-D-glucopyranosyl)-(1 ->4)-2,3,6-tri-O-benzyl-alpha, beta-D-glucopyranosyl trichloroacetimidate were coupled by a general condensation reaction to form the per-O-benzylated branched hexasaccharide phenyl thioglycoside. The phenylthio group of this compound was converted into a trichloroacetimidate, which was coupled with phenyl O-(2,3,6-tri O-benzyl-alpha-D-glucopyranosyl)-(1-->4)-O-(2,3,6-tri-O- benzyl-alpha-D glucopyranosyl)-(1-->4)-2,3,6-tri-O-benzyl-1-thio-beta-D- glucopyranoside to afford the per-O-benzylated branched nonasaccharide phenyl thioglycoside. Replacement of the phenylthio group with a free OH-group followed by hydrogenolysis gave the desired product. The synthons reported for this synthesis constitute a versatile tool for the chemical synthesis of other complex carbohydrates. PMID- 10515062 TI - Separation and structural analysis of saponins in a bark extract from Quillaja saponaria Molina. AB - Six major saponins were isolated from a bark extract from Quillaja saponaria Molina. Solid-phase extraction, followed by a two-step reversed-phase HPLC separation procedure with phosphate and ammonium acetate buffers of different pH values, was used. The compounds were characterised using NMR spectroscopy, mass spectrometry and chemical methods. PMID- 10515063 TI - (2-O-alpha-D-galactopyranosyl-4-O-methyl-alpha-D-glucurono)-D-xylan from Eucalyptus globulus Labill. AB - An unusual heteroxylan composed of galactosyl, 4-O-methyl-glucuronosyl and xylosyl residues with molar ratio 1:3:30 was isolated from the wood of Eucalyptus globulus Labill. The results of linkage analysis, supported by data of 1H, 2D 1H 1H COSY and 13C NMR spectroscopy, revealed that the polysaccharide is a (2-O alpha-D-galactopyranosyl-4-O-methyl-alpha-D- glucurono)-D-xylan with a (1-->4) linked beta-D-xylopyranosyl backbone branched at O-2 by short side chains composed of terminal 4-O-methyl-alpha-D-glucuronic acid and of 4-O-methyl-alpha-D glucuronic acid substituted at O-2 with alpha-D-galactose. PMID- 10515064 TI - A study of fucoidan from the brown seaweed Chorda filum. AB - Fucoidan fractions from the brown seaweed Chorda filum were studied using solvolytic desulfation. Methylation analysis and NMR spectroscopy were applied for native and desulfated polysaccharides. Homofucan sulfate from C. filum was shown to contain poly-alpha-(1-->3)-fucopyranoside backbone with a high degree of branching, mainly of alpha-(1-->2)-linked single units. Some fucopyranose residues are sulfated at O-4 (mainly) and O-2 positions. Some alpha-(1-->3) linked fucose residues were shown by NMR to be 2-O-acetylated. The 1H and 13C NMR spectra of desulfated, deacetylated fucan were completely assigned. The spectral data obtained correspond to a quasiregular polysaccharide structure with a branched hexasaccharide repeating unit. Other fucoidan fractions from C. filum have more complex carbohydrate composition and give rather complex methylation patterns. [formula: see text] PMID- 10515065 TI - Health and educational outcomes of children who experienced severe neonatal medical complications. AB - To determine the long-term developmental and educational outcomes of a sample of low birthweight infants with intraventricular hemorrhage (IVH), the authors conducted developmental assessments and interviews 8 years after the initiation of an early intervention project. At the time of the follow-up, 62% of the children were experiencing some developmental or behavior problems, with visual impairments, cerebral palsy, and attention deficits occurring most frequently. Grade of IVH and the number of days spent in the neonatal intensive care unit were the best predictors of later developmental delays. The sample also scored below average on school achievement; approximately 30% of those in school were eligible for special education services. These findings corroborate results from investigations with similar populations whose birth characteristics put them at risk for subsequent developmental delay. PMID- 10515066 TI - Infants' processing of causal and noncausal events at 3.5 months of age. AB - Infants' processing of causal (direct launching) and noncausal (delayed reaction, launching without collision) events was investigated with 30 infants who were 3.5 months old. The habituation-dishabituation technique was used. Results showed that the infants did not process direct launching as causal. However, their pattern of responding revealed a tendency to key on the spatial and temporal properties of the events. Those findings are discussed in terms of their compatibility with a modular, an information-processing, and a Piagetian framework. PMID- 10515067 TI - Social support and subjective well-being among Hong Kong Chinese young adults. AB - The associations between social support and 3 measures of subjective well-being- depressive symptomatology, negative affect, and positive affect--were studied among Hong Kong Chinese young adults (N = 475) between 16 and 19 years old. Significant bivariate relationships were found between positive affect and all dimensions of social support (including social network size, social contact frequency, satisfaction with social support, instrumental support, and helping others) except composition of social network. Helping others variables and relationship satisfaction variables were negatively related to both depressive symptoms and negative affect. Multiple regression models revealed that satisfaction with relationships with family members and friends was consistently associated with all measures of subjective well-being, and number of friends felt close to was positively related to positive affect. PMID- 10515069 TI - Gender differences in preschoolers' help-eliciting communication. AB - Gender differences in help-eliciting communication and the relationship of such utterances with ability were explored. A sample of 71 preschoolers (38 boys, 33 girls; mean age 4 years 3 months) were videotaped as they solved a difficult puzzle. Spontaneous talk was analyzed for orientation (to whom or to what an utterance referred) and for the frequency of utterances coded as help eliciting. Significant main effects for gender were observed, with more frequent help eliciting utterances (HEUs) made by the girls than by the boys, particularly HEUs about themselves (self-disclosing). Although the girls' HEUs were not predictive of ability on the puzzle, the boys' were. No gender differences in puzzle-solving ability were observed. Findings are discussed with regard to problem solving and social/linguistic development. PMID- 10515068 TI - Sex differences in neonates' cuddliness. AB - The authors undertook the present study to determine whether under ecologically valid, low-stress conditions, female and male neonates could be differentiated on cuddliness. Sixteen female and 15 male neonates were videotaped interacting briefly with both a female and a male adult who were blind to the sex of the neonate. Raters coded degree of cuddliness and activity level. Results showed that raters could discriminate the sex of the neonate on the basis of degree of cuddliness. Discussion focuses on the importance of theoretical and methodological considerations in assessing sex differences in behavioral characteristics of neonates. PMID- 10515070 TI - A test of the Piagetian water-level task with Chinese students. AB - A sample of 486 children in Beijing, China, were tested on the water-level task (WLT; J. Piaget & B. Inhelder, 1948/1956). The participants were 256 boys and 230 girls from 4th, 5th, 6th, 8th, and 11th grades. Three levels of mastery of the WLT were found. Level 1 consisted of 4th and 5th graders, who averaged about 71% correct. Level 2 consisted of 6th and 8th graders, who averaged about 83% correct. Level 3 consisted of 11th graders, who averaged 97% correct. The results provide partial support for the Piagetian theory of age-related developmental differences in performance on the WLT. The findings depict an interactive relationship of maturation with culture and education in the development of the ability to solve the WLT. PMID- 10515071 TI - Sex role identity, attitudes toward the opposite sex and same sex, and defense style. PMID- 10515072 TI - Photodynamic effects induced by meso-tetrakis[4-(carboxymethyleneoxy)phenyl] porphyrin on isolated Sarcoma 180 ascites mitochondria. AB - Using mitochondria isolated from Sarcoma 180 ascites tumour in Swiss mice as a model system, we have evaluated the ability of a novel porphyrin, meso-tetrakis[4 (carboxymethyleneoxy)phenyl]porphyrin (H2T4CPP), to induce damage on photosensitization. Oxidative damage to mitochondria, one of the primary and crucial targets of the photodynamic effect, is assessed by measuring products of lipid peroxidation such as thiobarbituric acid reactive substances (TBARS) and lipid hydroperoxides (LOOH), besides the loss of activity of the mitochondrial marker enzyme succinate dehydrogenase (SDH). Analysis of product formation, the effect of deuteration and selective inhibition by scavengers of reactive oxygen species (ROS) show that the damage observed is due mainly to singlet oxygen (1O2) and to a minor extent to hydroxyl radicals (.OH). The 1O2 generation and triplet lifetime of this porphyrin have also been estimated. Fluorescence spectroscopy, used to ascertain the binding of this porphyrin to the mitochondrial proteins, shows a rapid association within 0-2 h and a decline thereafter. Confocal microscopy reveals intracellular localisation of this porphyrin in cells in vitro. Our overall results suggest that the porphyrin H2T4CPP, due to its ability to bind to mitochondrial protein components and to generate ROS upon photoexcitation, may have potential applications in photodynamic therapy. PMID- 10515073 TI - White-light toxicity, resulting from systemically administered 5-aminolevulinic acid, under normal operating conditions. AB - This study has investigated damage to the intraperitoneal organs of the rat after systemic (intraperitoneal and intravenous) administration of low doses of 5 aminolevulinic acid (ALA) and illumination with a standard white-light operating room (o.r.) lamp. The study has been done within the framework of a larger study in which the possibility of using ALA for localization of small-volume macroscopically nonvisible peritoneal metastasis of ovarian tumors is being investigated. Fluorescence diagnostics are done in addition to the standard staging and localization procedures, either through a laparoscope or during laparotomy. In these circumstances, fluorescence diagnostics involve some risk of photosensitization of critical organs since a broad-band (o.r.) light source is used during the surgical procedures for illumination of the operating area. The drug dose and the time interval between administration of ALA and illumination are varied and normal tissues are examined both macroscopically and microscopically for damage. A relationship is demonstrated between the maximum tolerable dose (MTD) of ALA (defined as the dose that does not cause any tissue damage) and the time interval between administration and illumination. The white light that is used for illumination of the operating area is sufficient to induce damage to the peritoneal organs at relatively low ALA doses. The MDTs for 2, 6 and 16 h intervals are found to be respectively 1, 10 and 100 mg kg-1. The results are similar for both intraperitoneal and intravenous administration. PMID- 10515075 TI - Activation of macrophage tumoricidal activity by photodynamic treatment in vitro- indirect activation of macrophages by photodynamically killed tumor cells. AB - Macrophages constitute a major part of natural tumor defense by their capacity to destroy selectively a broad range of tumor types upon specific activation. In the last couple of years, these cells have also been implicated as effector cells in the destruction of tumors by photodynamic therapy. In the present work, the potential role of macrophage-mediated tumor cytotoxicity after photodynamic treatment in vitro has been investigated with respect to photodynamic activation of macrophages for tumoricidal effector functions. Our data show that photodynamic treatment of highly pure murine bone-marrow-derived macrophages with the hematoporphyrin derivative Photosan-3 does not result in activation of these cells for cytotoxicity against YAC-1 tumor cells or secretion of tumor necrosis factor and nitric oxide, irrespective of co-stimulation with interferon-gamma, a potent priming agent for macrophage antitumoral activity. On the contrary, treatment with higher photosensitizer doses is found to reduce markedly the viability of the macrophage effector cells. Thus, these results do not lend any support to the hypothesis of direct macrophage activation by photodynamic treatment. However, macrophages are found to be activated for tumoricidal effector functions indirectly by photodynamically killed tumor cells, in a way reminiscent of phagocytosis-inducing stimuli. It is thus suggested that recognition and phagocytosis of photodynamically destroyed tumor cells constitutes the major signal for local activation of macrophages in photodynamically treated tumor tissues, which may be crucial for final, specific eradication by the immune system of tumor cells surviving photodynamic treatment. PMID- 10515074 TI - The effect of glucose and temperature on the in vivo efficiency of photochemotherapy with meso-tetra-hydroxyphenyl-chlorin. AB - Balb/c athymic nude mice bearing WiDr human colon adenocarcinoma have been employed to investigate the effect of glucose administration, cooling or slight heating on the anti-tumor activity of photochemotherapy (PCT) with meso-tetra hydroxyphenyl-chlorin (mTHPC). An apparent delay in the tumor growth is found by combining PCT with either single or multiple injections of glucose. The anti tumor effect of PCT is slightly enhanced by cooling the tumor to 5 degrees C. Cooling also enhances the efficiency of PCT and glucose injection combined. Heating the tumor to 37 degrees C has no significant effect on either PCT alone or on the combination of PCT and glucose injection. Furthermore, the kinetics of the accumulation of mTHPC in tissue have been studied. Single or multiple injections of glucose have an enhancing effect on the accumulation of mTHPC in the tumor. PMID- 10515077 TI - The feasibility of using fluorescence spectroscopy as a rapid, non-invasive method for evaluating sunscreen performance. AB - We have carried out ex vivo studies to examine the feasibility of using fluorescence spectroscopy as an in vivo quantitative technique to assess sunscreen substantivity in terms of skin surface thickness and/or photoprotection. We found that the majority of sunscreens produced insufficient natural fluorescence and so we have attempted to increase the fluorescent signal by adding various fluorescing agents to the sunscreens. However, none of these substances is ideal; either they do not bind sufficiently strongly to sunscreen products, or their fluorescence is quenched by the active ingredients contained within sunscreens. The feasibility of using fluorescence spectroscopy for in vivo quantitative assessments of sunscreen substantivity therefore remains unproved and is dependent on a suitable fluorescent agent being found. Such an agent would have to be non-toxic, mix readily with sunscreens and be excited by visible wavelengths. PMID- 10515076 TI - Effect of delivery system on the pharmacokinetic and phototherapeutic properties of bis(methyloxyethyleneoxy) silicon-phthalocyanine in tumor-bearing mice. AB - A Si(IV)-phthalocyanine bearing two methoxyethyleneglycol axial ligands bound to the central metal ion (SiPc) has been prepared by chemical synthesis and analyzed for its phototherapeutic activity after administration in a Cremophor or liposome formulation to C57B1/6 mice bearing a subcutaneously transplanted Lewis lung carcinoma (LLC). The maximum drug accumulation in the tumor is found at 24 h after intraperitoneal injection, independent of the delivery system. However, the tumor concentration of SiPc in the Cremophor formulation is about two-fold higher, while the drug concentration in liver and skin shows similar trends with the two delivery systems. The drug accumulation and retention in the brain is much larger when using Cremophor emulsion. Photodynamic therapy (672 nm, 370 mW m 2, 360 J cm-2) at 24 h after the injection of Cremophor emulsion- or DPPC liposome-formulated SiPc causes a very efficient and similar response for the LLC (approximately 8 versus 22 mm mean tumor diameter for the control groups at 21 days after phototreatment). These very promising effects, obtained both at higher and lower tumor drug concentrations, clearly demonstrate the potential phototherapeutical activity of the newly synthesized SiPc. PMID- 10515078 TI - Quantitative studies on the respiratory burst generated in peritoneal macrophages. AB - The luminol-dependent chemiluminescence of macrophages during the zymosan stimulated respiratory burst has been studied both in the absence and in the presence of the radical inhibitor 3,5 di-tert-butyl-4-hydroxyphenyl propionic acid. In addition, the consumption of luminol and of the inhibitor has been followed analytically. Based on the rates of the consumption of the inhibitor, an iteration procedure yields a value of 2.2 x 10(-7) M for the steady-state concentration of radicals generated by cells at the maximum of the chemiluminescence in the presence of inhibitor. Approximate calculations have indicated that under the experimental conditions applied, additional formation of superoxide anion radicals by the oxidation of luminol is negligible. By assuming that in an inhibitor-free system the disappearance of radicals takes place via their combination process as well as by their interaction with luminol and/or with luminol-derived species, numerical integration yields a calculated curve of radical concentration versus time in fair agreement with experimental data and a rate-constant value for the combination of radicals of approximately 10(6) M-1 s 1, supporting literature findings according to which primarily superoxide anion radicals are formed. PMID- 10515079 TI - Spectrally resolved fluorescence imaging of human colonic adenomas. AB - Native fluorescence (autofluorescence) of human tissues can be a valuable source of diagnostic information for detecting premalignant and malignant lesions in the human body. Digital imaging of autofluorescence may be useful for localization of such lesions during endoscopic examinations. Tissue fluorescence of 31 adenomatous polyps obtained from 16 patients has been excited in vitro using the 325 nm line of a He-Cd laser. Digital images of the autofluorescence are recorded in six spectral bands. This study provides new data about the spatial distributions of autofluorescence intensities emitted in different spectral bands by colonic adenomatous lesions and normal colonic mucosa. Areas characterized by autofluorescence intensity lower than in normal mucosa are found for a majority of the polyps under study. The observed patterns of spatial distribution differ for the different spectral bands and for different polypoid lesions. No inverse correlation is found between the emission intensity and the thickness of colonic mucosa. The results indicate the spectral bands most useful for diagnostic applications and demonstrate the complexity of the optical processes involved in shaping both the spectra and intensities of the autofluorescence. PMID- 10515080 TI - Development and characterization of a DNA solar dosimeter. AB - In this paper, we report the development and characterization of a solar ultraviolet (UV) dosimetry system that can be used as a film badge for radiation monitoring. DNA molecules are coated on a thin nylon membrane as a UV dosimeter. The membrane is sealed in a polyethylene filter envelope with silica gel to keep the humidity low. After exposure to UV or solar light, induced DNA damage is measured by an immunochemical reaction. The intensity of color developed during the immunological reaction can be correlated linearly with the irradiated UV dose delivered by an Oriel solar simulator within a limited dose range. We observe no effects of temperature on the level of damage induction. The membrane is proficient for measuring DNA damage for more than 21 days when stored at either 37 or 4 degrees C. The induced damage remains stable on the membrane for at least 22 days at both 37 and 4 degrees C. In addition to these indoor experiments, we report measurements of solar UV dose in outdoor experiments. PMID- 10515081 TI - Hormone-refractory prostate cancer? Anti-androgen withdrawal and intermittent hormone therapy. AB - OBJECTIVE: To separate hormone-dependent from hormonally relapsed prostate cancers, a D3 category has been proposed. The term has become synonymous with a hormone-refractory state with the implication that any further hormonal treatment would not be beneficial. In this review we examine some data on androgen receptor expression, anti-androgen withdrawal syndrome and intermittent androgen deprivation (IAD) in patients with advanced prostate cancer. MATERIALS: The literature on the mechanism of the withdrawal phenomenon in patients with prostate cancer submitted to hormone therapy was reviewed. Experimental and clinical data are reported. RESULTS: The progression of prostate cancer in patients treated with combined androgen blockade (CAB) is associated with the activation of previously androgen-repressed genes, some of which may code for autocrine and paracrine growth factors that substitute for androgens in maintaining the viability of the tumorigenic stem cells. If androgens are replaced before progression begins, the surviving stem cells should give rise to an androgen-dependent tumor, which would be amenable to retreatment by CAB. This theory provides the rationale for intermittent androgen deprivation. We suggest that IAD could be more effective in patients with initial prostate-specific antigen (PSA) progression after radical prostatectomy. CONCLUSIONS: Response to withdrawal therapies or second-line treatments is an example of a "hormonal" therapy that may benefit a proportion of patients with hormone-refractory disease, suggesting that the tumor is still androgen-dependent. Whether IAD enhances progression-free survival or overall survival must be verified in randomized clinical trials. Until further studies have been completed, the therapeutic concept of IAD should be treated as experimental. PMID- 10515082 TI - Three-year follow-up after transurethral microwave thermotherapy (TUMT) for benign prostatic hyperplasia using the PRIMUS U + R device. AB - OBJECTIVE: We report long-term (3 years) follow-up data of transurethral microwave thermotherapy (TUMT) for benign prostatic hyperplasia (BPH) using a lower-power treatment protocol. MATERIAL AND METHODS: Ninety-one patients were treated in a 1-h session with the PRIMUS U + R device. RESULTS: Forty-five of the patients were still on TUMT monotherapy at 3-year follow-up, while 32 received additional therapy for their lower urinary tract symptoms. In patients with monotherapy there was a 45% decrease in international prostate symptom score (IPSS) when compared to pretreatment values. The moderate increase in peak uroflow seen early after TUMT could not be observed after 3 years. No serious side-effects were seen. CONCLUSION: Three years after lower-power TUMT, 49% of patients treated were on TUMT monotherapy, while 35% received additional therapy for their voiding symptoms. Symptom score decreased 45% in patients with TUMT monotherapy concomitant with unchanged uroflow. PMID- 10515083 TI - Blood ammonia levels after intravenous infusion of glycine solution with and without ethanol. AB - OBJECTIVE: Absorption of glycine 1.5% during transurethral resection of the prostate may increase blood ammonia levels, but hyperammonaemia has not been described when the fluid also contained ethanol 1%. The aim of this experimental study was to evaluate whether ethanol 1% reduces glycine-induced hyperammonaemia. MATERIAL AND METHODS: Two intravenous infusions of glycine solution with and without ethanol 1% added were given on different occasions to 20 male volunteers (mean age 30 years). Half of them received 22 g of glycine over 50 min and the others approximately 18 g over 30 min. Blood ammonia was measured before and 30 min after the infusion. The serum levels of free amino acids were measured on 7 occasions during 10 of the experiments. RESULTS: The glycine infusions increased blood ammonia levels from 37 micromol/l (median, 10th and 90th percentile limits 34-53) to 57 micromol/l (27-110; p < 0.001). The change was greater after the larger glycine dose, regardless of whether the fluid contained ethanol (p < 0.05). The only amino acid concentration correlating with blood ammonia was glycine, which showed higher levels in those who had a rise in blood ammonia of 50% or more. CONCLUSIONS: Ethanol 1% did not reduce the increase in blood ammonia concentration after the administration of glycine solution. PMID- 10515084 TI - Voiding and sexual dysfunctions after pelvic fracture urethral injuries treated with either initial cystostomy and delayed urethroplasty or immediate primary urethral realignment. AB - OBJECTIVE: The aim of this study is to evaluate the effects of the different immediate treatment modalities on the sexual and voiding functions in pelvic fracture urethral injuries. METHODS: The records of 38 male patients with traumatic posterior urethral injuries were reviewed, 18 of whom were treated by initial suprapubic cystostomy and delayed repair (Group 1), and 20 by primary urethral realignment (Group 2). Types of pelvic fractures and urethral injuries were classified according to surgical and radiological findings. Long-term voiding functions were determined by the patient questionnaire, residual urine and uroflow. Sexual functions were also determined by the patient questionnaire and a penile duplex ultrasound study. RESULTS: Mean follow-ups of Groups 1 and 2 were 37 and 39 months, respectively. Membranous urethral disruption extending to the urogenital diaphragm was the most frequent urethral injury (type 3), with incidences of 66.7% and 77.7%, respectively. There were no statistically significant differences in mean age, incidence of pelvic fracture types and urethral injury types between groups (p > 0.05). After the immediate treatments, 16.7% and 55% of the patients regained normal urination, and stricture developed in 83.3% and 45% of the patients, respectively. In 44.4% of the patients in Group 1 and 10% in Group 2, urethral strictures required open urethroplasty (p < 0.05). Erectile impotence before urethroplasty in 17.6% and 20%, anejaculation after urethroplasty in 17.6% and 15% and incontinence in 5.6% and 10% of the patients were found in Groups 1 and 2, respectively (p > 0.05). However, 88.8% and 90% of patients eventually achieved normal urination with complete continence. CONCLUSION: Sexual and voiding dysfunction after pelvic fracture posterior urethral injury seem to be the result of the injury itself, not of the immediate treatment modalities. In urethral disruption injuries, primary urethral realignment seems more favourable than suprapubic cystostomy and delayed repair. PMID- 10515085 TI - Specificity of human complement factor H-related protein test (Bard BTA stat Test). AB - OBJECTIVE: The Bard BTA stat Test is a new, rapid, non-invasive qualitative test that detects bladder tumour-associated antigen (human Complement Factor H-related protein) in urine. The objective of this study was to evaluate the specificity of the Bard BTA start Test in a healthy population. MATERIALS AND METHODS: Voided urine samples from 100 healthy volunteers were collected and tested using the BTA stat Test. In the case of a positive BTA stat result, i.v. urography, cystoscopy, ultrasound and-re-testing were performed and the BTA stat Test was repeated three months later. RESULTS: Two subjects tested positive by the BTA stat Test, but no bladder cancer was found. The specificity of the BTA stat Test in the studied healthy population was 98%. CONCLUSIONS: The BTA stat Test has very high specificity with a true test-dependent false-positive rate occurring in only 2% of the healthy population. These results justify further studies with the BTA stat Test for bladder cancer diagnosis, monitoring, and screening. PMID- 10515086 TI - Estrogen receptors in the human male prostatic urethra and prostate in prostatic cancer and benign prostatic hyperplasia. AB - Estrogen receptors (ERs) in the prostate and prostatic urethra were examined in 33 men with benign prostatic hyperplasia (BPH) and in 11 with prostate cancer (PC). The Abbot monoclonal ER-ICA assay was used for immunohistochemical investigation. In the BPH group, ERs were revealed in the prostatic stroma in eight cases and in the glandular epithelium in one. In four cases ERs were seen in the prostatic stroma and in the glandular epithelium. In the prostatic urethra, ERs were found in 19 cases located in the urothelium, lamina propria and/or periurethral glands. In the PC group, ERs were demonstrated in the prostatic stroma and/or prostatic urethra in 6 out of 11 cases. In both BPH and PC patients, immunoreactivity was weak and confined to few cells, indicating low ER content in the prostate as well as in the prostatic urethra. Dextran-coated charcoal (DCC) analysis was used for detection and quanticization of cytosolic and nuclear ERs. In the BPH group, ERs were detected once in the prostate and prostatic urethra in the nuclear and cytosol, and additionally in the prostatic urethra in the cytosol fraction in three cases. In all cases, ER content was low, ranging from 10-15 fmol/mg protein. In the PC group, ERs were detected in the prostatic urethra and/or prostate in the cytosol fraction from two patients. The contents were low, ranging from 10-13 fmol/mg protein. We conclude that in human BPH and PC, ERs can be present in the prostate and prostatic urethra. In the prostate, ERs are mainly located in the stroma, but in BPH specimens they can also be found in the glandular epithelium. Biochemically, the use of the DCC analysis is of limited value, since ER content in the human prostate and prostatic urethra is at the limit of detection with this method. PMID- 10515087 TI - Localization of transforming growth factors beta1 and beta2 and epidermal growth factor in IgA nephropathy. AB - OBJECTIVE: The localization of transforming growth factor (TGF)-beta1. TGF-beta2 and epidermal growth factor (EGF) was investigated in IgA nephropathy, and was compared with the severity of histological damage (including tubulointerstitial lesions). MATERIALS AND METHODS: The enzyme antibody method was used to stain paraffin-embedded sections of renal tissue from 42 patients with IgA nephropathy (19 males and 23 females). RESULTS: There was a significant correlation between glomerular positivity for TGF-beta1 and TGF-beta2 and the severity of histological damage. There was also a significant correlation between positivity for TGF-beta1 and TGF-beta2 in the tubular epithelium and tubulointerstitial lesions. In contrast, there was no relationship between glomerular positivity for EGF and histological damage, although there was a significant correlation between positivity for EGF in the tubular epithelium and tubulointerstitial lesions. CONCLUSIONS: These findings suggest that TGF-beta1 and TGF-beta2 may be important in the progression of IgA nephropathy, and that the distribution of EGF may also be a useful marker for the progression of renal damage, including tubulointerstitial lesions. PMID- 10515088 TI - Effect of pH and glucose on cultured human peritoneal mesothelial cells. AB - OBJECTIVE: We investigated the effects of various pH and glucose concentrations on the growth of human peritoneal mesothelial cells and on coagulation and fibrinolytic factors. MATERIALS AND METHODS: Cells were cultured at various pH values in Ham's F-12 medium containing 1.0% foetal calf serum and supplemented with D-glucose or D-mannitol at various concentrations. After 4-48 h, cell proliferation and 3H-thymidine incorporation were determined. Coagulation and fibrinolytic factors were measured after 48 h. RESULTS: Glucose caused concentration-dependent inhibition of cell growth at all pH values, but the deleterious effect of low pH on cell proliferation was faster and stronger than that of high glucose. At a similar osmolality, mannitol caused less inhibition of cell proliferation than glucose. There was a glucose concentration-dependent increase of thrombin-antithrombin III complex production at all pH values. At pH 5.2, tissue-type plasminogen activator production was far lower than at higher pH values, and production of the plasminogen activator inhibitor showed a glucose concentration-dependent increase. At pH 6.5 or 7.3, however, the plasminogen activator inhibitor production decreased and tissue-type plasminogen activator production increased in a glucose concentration-dependent manner. CONCLUSIONS: Low pH and/or high glucose culture medium had an inhibitory effect on peritoneal mesothelial cells, with the effect of high glucose being partially related to hyperosmolality. These cells may modulate peritoneal coagulant and fibrinolytic activity, with the balance between coagulation and fibrinolysis being disturbed by low pH and/or high glucose. PMID- 10515089 TI - Cephalad renal ectopia, duplication of pelvicalyceal system and patent ductus arteriosus in an adult female. AB - An unusual cause for a shadow in the plain CX-ray of a female with uncomplicated patent ductus arteriosus is presented herein. The chest CT scan and IVU revealed the presence of a high (cephalad) right kidney in an eventrated hemidiaphragm. A bifid ureter and duplication of the pelvis were found in the contralateral kidney. This constellation of anomalies is exceedingly rare. PMID- 10515091 TI - Hypercalciuria preceding IgA nephropathy in a child with haematuria. AB - We describe a child with isolated haematuria who was diagnosed and successfully treated for idiopathic hypercalciuria for 6 months, after which IgA nephropathy was demonstrated on renal biopsy performed due to the relapse of haematuria in spite of low calciuria levels. To our knowledge, this is the first case evaluated systematically in the literature shown to have IgA nephropathy while being followed up for idiopathic hypercalciuria. PMID- 10515090 TI - Rhabdomyolysis and acute renal failure associated with influenza virus type A. AB - Two patients with rhabdomyolysis-induced acute renal failure due to influenza A virus infection are presented. Both had influenza symptoms, with high fever and severe muscular pain leading to walking problems. In addition, they were dehydrated due to vomiting and diarrhoea. Both had evidence of an ongoing influenza infection according to serological tests. Muscle injury due to the viral infection gave rise to rhabdomyolysis with efflux of myoglobin from the muscles, causing renal failure. In conclusion, influenza A virus infection can cause rhabdomyolysis accompanied by reversible acute renal failure. PMID- 10515093 TI - Hypothesis: from epidermal barrier dysfunction to atopic disorders. AB - The role of a genetically-impaired epidermal barrier as the primary cause of the rapid increase in prevalence of atopic dermatitis and respiratory atopy is proposed, based on available clinical and experimental data. The subsequently increased exposure to irritants and allergen postnatally in predisposed individuals would lead in a subset of these to a specific TH2 cell activation favouring the development of IgE responses to atopens. Other routes of sensitization are probably important, but skin offers a good target to implement prevention strategies, so far completely ignored in the prophylactic recommendations given to high-risk families. Candidate genes for skin-barrier impairment are possibly those associated with ichthyosis vulgaris and X-linked hypohidrotic ectodermal dysplasia. PMID- 10515092 TI - A case of tuberculosis of the prostate. AB - Tuberculosis of the prostate is uncommon. However, the number of patients with tuberculosis has once again recently been gradually increasing in Japan. The number of immunocompromised hosts, such as those with AIDS, is also increasing, suggesting that this rare infectious disease may increase in frequency in the near future. We present a case of tuberculosis of the prostate. PMID- 10515094 TI - Allergic contact dermatitis from a perinone-type dye C.I. Solvent Orange 60 in spectacle frames. AB - This is the 1st case report of allergic contact dermatitis from a perinone-type plastic dye, C.I. Solvent Orange 60, used in the earpieces of spectacle frames. Sensitization of this dye was confirmed by patch tests and chemical analysis of the causative earpieces and coloring agents. Solvent Orange 60 is suspected of being the contact allergen in at least 2 other Japanese cases of spectacle earpiece dermatitis, and provoked strong reactions on sensitized individuals. Its use in products that are applied on human skin for a prolonged period of time, such as spectacle frames or hearing aids, would best be avoided. PMID- 10515095 TI - Topical provocation in 27 cases of cotrimoxazole-induced fixed drug eruption. AB - The purpose of this study was to investigate the usefulness of topical provocation in the diagnosis of cotrimoxazole-induced fixed-drug eruption (FDE). 27 patients with established cotrimoxazole-induced FDE by oral provocation and 20 healthy controls were tested with drugs at increasing concentrations in white petrolatum and dimethyl sulfoxide (DMSO) both on previously involved and uninvolved skin sites. Tape-stripping occlusive patch testing in petrolatum remained negative in 19 tested patients. Open testing with drug preparations in DMSO revealed positive results in 25 of 27 tested patients. 1 patient showed an additional positive reaction on previously uninvolved skin. Lesions on male genitalia and on face reacted to testing once with 10% or 20% of the suspected drug, whereas repeated testing with concentrations up to 50% was necessary in lesions on trunk & extremities. Open testing with drug preparations in DMSO at concentrations of 10%, 20% and 50% and pure DMSO remained negative in 20 healthy controls. The present study shows that repeated open testing with graded concentrations of the drugs up to 50% in DMSO is a reliable test method in sulfamethoxazole/trimethoprim-induced FDE. Patients and physicians should be aware of the transient irritant reaction to DMSO that is not infrequent, so as to avoid false-positive interpretations. PMID- 10515096 TI - Putative skin-protective formulations in preventing and/or inhibiting experimentally-produced irritant and allergic contact dermatitis. AB - The effectiveness of skin protective formulations was evaluated in a previously described in vivo human model. All formulations failed to inhibit ammonium hydroxide and urea irritation. Only paraffin wax in cetyl alcohol statistically (p<0.01) reduced Rhus allergic contact dermatitis. 3 commercial formulations markedly (p<0.001) suppressed sodium lauryl sulfate irritation. Paraffin wax in cetyl alcohol was quantitatively the most effective formulation. These results suggest that some formulations may provide protective effects against certain, but not all, irritants or allergens. PMID- 10515097 TI - Suspected fragrance allergy requires extended patch testing to individual fragrance allergens. AB - This study has been performed to evaluate the efficacy of fragrance mix (FM) as a screen for fragrance allergy. Patients were included if they had had positive allergic reactions to FM, to 1 of the 8 ingredients of FM, to 1 of 14 other fragrance materials, or to their own perfume. 91 patients were studied. There were 65 women and 23 men (in 3, their sex was not recorded) allergic to FM on patch testing. The mean (+/-SD) age was 48.4+/-18.6 years. 22 patients gave a past history of atopic eczema. Dermatitis of the hands (31%) and face (26%) were the most common presenting complaints. 85 patients (93%) had a positive allergic patch test reaction to FM. 22 of the 40 tested to the extended fragrance series were positive to other perfumes as well, and of these, there were 14 reactions (in 9 patients) to allergens not in the FM. In addition, 6 patients were positive only to separately tested fragrance constituents and not to the FM. In conclusion, FM is an accurate screen for fragrance contact sensitivity. However, patch testing to an extended series is needed if there is clinical suspicion of perfume allergy, as otherwise about 7% of patients allergic to fragrances will be missed. PMID- 10515098 TI - Dose-response studies of contact allergens using 3 guinea pigs models. AB - A multi-dose-response induction protocol for the guinea pig maximization test (GPMT), including a statistical computer program, has earlier been developed to improve the power of predictive tests for identification of contact allergens. This dose-response protocol, with 2 modifications (i.e., increased number of animals in each group and increased number of challenge concentrations) was evaluated in the GPMT, the cumulative contact enhancement test (CCET) and the Freund's complete adjuvant test (FCAT), using potassium dichromate and hydroxycitronellal as model contact allergens. Application of the dose-response protocol on the CCET and the FCAT resulted in either monotone or non-monotone curves with significant dose-response. However, application of the dose-response protocol on the GPMT gave curves with no significant dose-response. The protocol makes it possible to obtain an EC50 value, thus improving the possibility of ranking contact allergens, which is of substantial use for risk assessments. The dose-response protocol could benefit from a few adjustments: a wider span in the induction doses; change to simultaneous increase in intradermal and topical induction doses to obtain a proper dose-response for the GPMT; the addition of further challenge concentrations. In addition the computer program should allow calculation of threshold concentration for sensitization and EC50 value for a non monotone curve. PMID- 10515099 TI - Histological distinction between early allergic and irritant patch test reactions: follicular spongiosis may be characteristic of early allergic contact dermatitis. AB - Comparative light microscopic studies have revealed subtle differences between allergic and irritant reactions in the skin. In the search for specific differences, we focussed on the early inflammatory response. This pilot study was conducted to test the hypothesis that follicular spongiosis can differentiate between early allergic and irritant patch test reactions. 8 patients with known contact allergy to either colophony or quarternium-15 participated in the study. In each patient, allergic and irritant patch tests reactions were elicited, and 4 mm punch biopsies were taken after 6 8 h from clinically equipotent reactions. Paired sets of slides were assessed blindly by 2 pathologists. 1 patient showing a pityrosporum folliculitis was excluded from the study. All biopsies from allergic patch tests were characterized by follicular spongiosis, while biopsies from irritant patch tests showed no recognizable changes except a slight follicular spongiosis in 1 patient. The 2 pathologists agreed independently on the correct classification in 6 out of 7 cases (p=0.0156). We tested an optimized model, selecting non-irritant allergens and a well-known irritant. Further investigations are needed to elucidate the diagnostic significance of the histological classification of allergic and irritant cutaneous reactions in punch biopsies. PMID- 10515100 TI - Assessment of budesonide patch tests. AB - Patch-test technique for budesonide needs improvement. 20 subjects with positive or questionable patch-test responses to budesonide were retested for 3 to 96 h (4 days [D]) with polyester patches coated with budesonide in serial doses (150 to 0.074 microg/cm2). Multiple readings were taken visually and with a laser Doppler perfusion imaging technique up to 264 h (day [D]11). Additionally, all subjects were tested with 0.1% budesonide in petrolatum in Finn Chambers for 48 h (2D) with readings taken at 72 (D3), 96 (D4) and 168 h (D7). Different dose levels and application times affected unpredictably highest assessments of reactions. No clear suppression of reactivity was observed at high doses. Time points of highest assessments of reactions varied between subjects but were generally the same for each subject with both reading methods regardless of dose levels or application times. Positive and negative subjects during the study were easily distinguished with all serial doses, regardless of assessment technique. At 2.0 microg/cm2, the lowest dose level tested on all subjects, longer applications than 24 h (1D) were required to detect all positive subjects. 48-h (2-D) applications required 2 readings, optimally at 96 (D4) and 216 h (D9). The only test technique with Finn Chambers used here did not make such distinction possible. PMID- 10515101 TI - Allergic contact dermatitis due to bendazac and alclometasone dipropionate. PMID- 10515102 TI - Airborne occupational hypersensitivity to isothiazolinones in a papermaking technician. PMID- 10515103 TI - Delayed reaction to oral treatment with clindamycin. PMID- 10515104 TI - Contact allergy to 'caines' caused by anti-hemorrhoidal ointments. PMID- 10515105 TI - Drug eruption due to tribenoside. PMID- 10515106 TI - IgG latex RAST is not a specific marker for latex Type I hypersensitivity. PMID- 10515107 TI - Clinical relevance of positive patch test reactions to preservatives and fragrances. PMID- 10515108 TI - Occupational allergic contact dermatitis from epoxy resin used to restore window frames. PMID- 10515110 TI - Dermatitis induced by a spinal cord stimulator implant. PMID- 10515109 TI - Acute contact dermatitis from Naphthol AS. PMID- 10515111 TI - Allergic contact dermatitis from myrrh. PMID- 10515112 TI - Lack of patch test reactivity to 3-dimethylaminopropylamine in German hairdressers. PMID- 10515114 TI - Benzoyl peroxide as a contact allergen in adhesive tape. PMID- 10515113 TI - Phytophotodermatitis due to Ruta graveolens applied as protection against evil spells. PMID- 10515115 TI - Contact allergy to isopropanolamine in Traxam gel. PMID- 10515116 TI - Contact urticaria syndrome caused by patch testing with cefotiam hydrochloride. PMID- 10515117 TI - Occupational allergic contact dermatitis from epoxy resins based on bisphenol F. PMID- 10515118 TI - Allergic contact dermatitis from cobalt in a beauty product. PMID- 10515119 TI - Acute anaphylaxis resulting from routine patch testing with latex. PMID- 10515120 TI - An unusual case of allergic contact dermatitis from corticosteroids. PMID- 10515121 TI - Erythematous skin reaction to subcutaneous injection of ribonucleic acid. PMID- 10515122 TI - Genetic and epigenetic contributions to colorectal cancer. AB - Both genetic and epigenetic factors contribute to the development of colorectal cancer. Specific genetic changes in proto-oncogenes, tumor suppressor genes, and DNA mismatch repair genes have led to a genetic model of colorectal tumorigenesis. Recent data highlight the importance of the TGF-beta signaling pathway in regulating the progression of colorectal cancer. The loss of the tumor suppressor activity of this pathway as well as the potentially cooperative genetic aberrations involving APC, K-ras, and p53 are reviewed in the context of the multi-step adenoma-carcinoma sequence that characterizes the development of colorectal tumorigenesis. In addition, contributing epigenetic factors including age, diet, angiogenesis, and immune response are also discussed. Combining our knowledge of the genetic and epigenetic events implicated in this disease may allow a broader understanding of the pathogenesis of colorectal cancer and hence the design of better anti-tumor interventions. PMID- 10515123 TI - Anti-miracidial effect of recombinant glutathione S-transferase 26 and soluble egg antigen on immune responses in murine schistosomiasis mansoni. AB - The anti-miracidial potential of recombinant Schistosoma mansoni glutathione S transferase 26 (rSmGST26) or native crude soluble egg antigens (SEA) was assessed. The associated dynamics of granuloma formation and immune responses were evaluated. Naive C57BL/6 mice were injected intravenously with multiple doses of either SEA (SEA-group) or rSmGST26 (GST-group) 7 days before cercarial infection. The immunized groups and the respective controls were sacrificed 6, 8 and 16 weeks postinfection (p.i.). Acceleration of ova destruction and reduction of granuloma diameter were greater in the GST-group than the SEA-group, mainly at 8 weeks p.i. However, the amelioration of hepatic pathology and function was more evident in the SEA-group. Concurrently, serum-specific IgG1 levels were elevated throughout the course of infection in the immunized groups compared to the infected controls. Initial rise of all splenic cytokines and serum anti-SEA IgE levels at 6 weeks p.i. was observed, followed by a dramatic drop in the levels of the proinflammatory cytokines IL-2, IFNgamma, IL-4 and TNF-alpha and IgE at 8 weeks of infection. IL-10 level was lower at 8 weeks p.i. than at 6 weeks, but was higher in immunized groups than in infected controls. Several responses may be implicated as an outcome of the present immunization protocol, such as increased levels of blocking antibody (IgG1) and IL-10 with decreased levels of proinflammatory cytokines and IgE. PMID- 10515124 TI - Moraxella catarrhalis-induced purulent otitis media in the rat middle ear. Structure, protection, and serum antibodies. AB - To study the effects of viable and heat-killed Moraxella catarrhalis bacteria on the middle ear mucosa and to evaluate the protection after whole-cell immunizations, Sprague-Dawley rats were challenged and rechallenged with four different M. catarrhalis strains. The animals were monitored by clinical observations, bacterial and histological samples from middle ears, and serum IgG levels. Only viable bacteria at a high concentration induced purulent otitis media, which was culture positive in 58% of the cases on day 4. The infection was characterized by a mild acute reaction lasting otomicroscopically about 8 days, together with quantitative and qualitative changes of the goblet cells. Structurally the mucosal effects of the heat-killed bacteria were less pronounced in the early phase compared to the viable bacteria, but similar at the end of the experiment at 6 months. The intrabullar and subcutaneous immunizations evoked an IgG antibody response in all animals, and the protection rate after immunization was 50% or more. The induced protection was not strain-specific. The study showed the rat to be a possible alternative for the study of different aspects of M. catarrhalis otitis media, an infection that is clinically and structurally different from that elicited by Streptococcus pneumoniae and Haemophilus influenzae in the rat. PMID- 10515125 TI - Minute oxidative stress is sufficient to induce apoptotic death of NIT-1 insulinoma cells. AB - When cultured NIT-1 cells were subjected to a low level of oxidative stress (30 microM hydrogen peroxide for 15 min at 37 degrees C) several of their lysosomes ruptured, as demonstrated by intravital staining with the lysosomotropic weak base acridine orange. Such rupture is due to intralysosomal, iron-catalyzed oxidative reactions, since it was largely prevented by previous endocytotic uptake of desferrioxamine. The resultant limited leakage of lysosomal hydrolytic enzymes into the cytosol could be important for an apoptotic-type degradation/fragmentation process within initially intact plasma membranes. In contrast, extensive lysosomal rupture leads to necrosis. The development of the damage process was followed by light- and electron microscopy; and by the TUNEL reaction. As a result of the applied oxidative stress, which is comparable to that expected to occur within the microenvironment surrounding activated macrophages under oxidative burst (e.g. during autoimmune insulitis), about 90% of the cells eventually died due to post-apoptotic secondary necrosis. The few surviving cells phagocytosed the debris from their fragmented neighbours and began to divide about 24 h after the insult. Thus the sensitivity to oxidative stress varies, perhaps as a consequence of varying amounts of intralysosomal redox-active iron, as we have found to be the case in several other cellular systems. Since the NIT-1 cells are highly differentiated, and in many ways like beta cells, we consider our result to be of value for the understanding of beta cell death during the development of insulin-dependent (Type I) diabetes mellitus (IDDM). PMID- 10515126 TI - Basaloid type adenoid cystic carcinoma of the breast. AB - We report a case of adenoid cystic carcinoma (ACC) of the breast with a prominent basaloid feature. The patient was a 62-year-old Japanese woman with a right breast mass, measuring 1.5 cm in diameter. Histologically, the tumor was composed of basal cell-like tumor cells, and it was originally diagnosed as invasive ductal carcinoma. The presence of PAS-positive basement membrane material around the tumor cell nests may be a diagnostic clue to ACC. The prognosis of ACC of the breast is considered to be favorable. However, basaloid type ACC may represent a poor prognosis, since our case revealed an aggressive behavior in spite of its small size. PMID- 10515127 TI - Expression of fibroblast growth factor-2 transcripts in the healing of acetic acid-induced gastric ulcers. AB - Basic fibroblast growth factor-2 (FGF-2) has an important role in angiogenesis, and has been demonstrated to promote ulcer healing. However, the actual part played by FGF-2 in the process of ulcer healing is not well understood. In this study, we investigated expression of FGF-2 transcripts at each stage of gastric ulcer healing, using an acetic acid model in rats. We made ulcers in the rat stomach by direct application of acetic acid, and 3 days and 1, 2, 3, 4, 8 weeks after treatment, we examined expression of FGF-2 transcripts by in situ hybridization. On day 3, FGF-2 transcripts were detected in mononuclear cells infiltrating the submucosal layer around the ulcer. After 1 and 2 weeks, expression of FGF-2 transcripts was prominent in spindle-shaped mesenchymal cells and endothelial cells, which proliferated in the ulcerative region. Some of the spindle-shaped cells which expressed FGF-2 transcripts also showed immunoreactivity for alpha-smooth muscle actin. After 3 and 4 weeks, FGF-2 expression was seen mainly in endothelial cells of vessels. These results suggest that different cells produce FGF-2 during the process of gastric ulcer healing, and some of the spindle-shaped cells expressing FGF-2 transcripts in the early phase are myofibroblasts. PMID- 10515128 TI - The role of volume-weighted mean nuclear volume in predicting disease outcome in patients with prostate cancer treated with radical prostatectomy. AB - BACKGROUND: Estimates of volume-weighted mean nuclear volume (MNV) are the only means by which unbiased estimates of three-dimensional parameters can be obtained from single two-dimensional sections without any assumptions. We have reported that for prostate cancer estimates of MNV are prognostically equal or superior to morphological grading of malignancy, such as Gleason score (GS), and in particular, that MNV proved to be a meaningful predictor of prognosis for patients with clinically localized tumors. However, all previous studies were conducted on patients treated conservatively, and no authors have tested whether estimates of MNV can predict the prognosis of patients treated with radical prostatectomy. MATERIALS AND METHODS: A retrospective prognostic study of 52 patients with clinically localized prostate cancer diagnosed at three Hospitals in Shizuoka Prefecture, Japan (Shizuoka City Hospital, Shizuoka Prefectural Hospital and Shimada Municipal Hospital) and treated by radical prostatectomy was performed. Twenty of these patients were treated with hormone therapy before radical prostatectomy. Unbiased estimates of MNV were compared with clinical stage, histological grading according to GS and neo-adjuvant hormone therapy with regard to the prognostic value. RESULTS: MNV was significantly correlated with pathological T stage, but was not significantly correlated with the presence or absence of lymph node metastasis. Univariate analysis revealed that MNV correlated significantly with progression-free survival (p = 0.0116). Multivariate analysis revealed that MNV (p = 0.0115) and GS (p = 0.0275) were two significant independent predictors of progression-free survival. CONCLUSIONS: The results of the present study suggest that MNV and GS are powerful independent predictors of prognosis for prostate cancer treated with radical prostatectomy. We recommend estimates of MNV as a supportive method to the histological grading for patients with prostate cancer. PMID- 10515129 TI - Blood-group-related carbohydrates are expressed in organotypic cultures of human skin and oral mucosa. AB - Cellular maturation and migration are usually associated with changes in cell surface carbohydrates, but the relationship between these changes and cell behaviour is at present largely unknown. To investigate whether an organotypic culture system can be used as an in vitro model to study the function of cell surface carbohydrates, we established organotypic cultures of skin and buccal mucosa. In these cultures, keratinocytes are grown at the air-liquid interface on a supporting matrix consisting of homologous fibroblasts embedded in a collagen type I gel. We examined the expression of blood-group-related carbohydrate structures, including Lewis x, sialylated Lewis x, Lewis y, Lewis a, and Lewis b, on the surface of epithelial cells in the cultures. We compared the results with the expression of more well-established markers, including cytokeratins, integrins, bullous pemphigoid antigen and laminin, in the same cultures. The organotypic skin and oral mucosa cultures showed a histological differentiation pattern analogous to that of normal skin and buccal mucosa, and a tissue-specific expression of carbohydrate structures and cytokeratins. However, both types of organotypic cultures also expressed markers which are normally seen during wound healing, including Lewis y, cytokeratin 16, and cytokeratin 19. We conclude that the organotypic cultures of oral mucosa and skin are suitable models for future studies of the function of cell-surface carbohydrates, although the expression of wound healing markers has to be taken into consideration. PMID- 10515130 TI - Unbiased estimation of total beta-cell number and mean beta-cell volume in rodent pancreas. AB - We describe a method for unbiased assumption-free estimation of the total number of beta-cells and the mean beta-cell volume in mouse and rat pancreas based on light microscopy. Such a method, which takes advantage of one of the most recent developments in stereology, the fractionator, has not previously been described. It relies on repeated fractionation of the tissue using systematic uniform random sampling combined with an unbiased counting principle. The method was applied to eight BALB/cBom male mice (56 days) and six Lewis/MOL male rats (47 days). In mice, the total number of beta-cells was 1.06 +/- 0.07 x 10(6) (mean +/- SEM) per pancreas with a mean beta-cell volume of 1280 +/- 17 microm3, while in rats the total beta-cell number was 2.76 +/- 0.42 x 10(6) per pancreas with a mean beta cell volume of 1170 +/- 65 microm3. Furthermore, the results showed that in both species the biological variability in the total beta-cell volume is due to differences in the number of beta-cells rather than variability of the mean beta cell volume. The method can be used to give a precise description of number and volume of beta-cells at different ages, and will make it possible to estimate the contributions of hyper/hypotrophia and hyper/hypoplasia to a given induced or spontaneous change in the total beta-cell mass. PMID- 10515131 TI - A novel and efficient regimen for producing chronic relapsing experimental autoimmune encephalomyelitis (CR-EAE) in SJL mice. AB - We report that SJL mice developed chronic relapsing experimental autoimmune encephalomyelitis (CR-EAE) when injected with a mixture of mouse spinal cord homogenate (MSCH), killed mycobacteria tuberculosis (M. tb), and mycobacteria butyricum (M. b) in PBS 2 months before a conventional acute experimental autoimmune encephalomyelitis (EAE) induction injection. The altered progression of the disease involved an accelerated but less severe acute attack and development of a chronic course with relapsing-remitting episodes. Histological examination revealed inflammatory cell infiltration and demyelination in the brain. The dose of neuroantigen as well as the anatomical sites of injections were found to be crucial for the development of the disease. PMID- 10515132 TI - The expanded critique concept for enlarging the reviewer's influence regarding original research articles. PMID- 10515133 TI - Clinical research. Studying relationships between occlusion and TM disorders. PMID- 10515134 TI - Value of CPI, MPI. PMID- 10515135 TI - Measurements must be interval, not ordinal. PMID- 10515136 TI - Stability of implants as anchorage for orthopedic traction. AB - The aim of this animal study was to investigate the stability of osseointegrated fixtures when used as anchorage for orthopedic traction with extreme force magnitude. Three Branemark fixtures were placed in the left zygomatic arch and three in the right of five adult dogs. An orthopedic nonaxial force of 5 N was applied using an intraoral coil system. The initial displacement immediately after force application was measured by means of speckle interferometry. After 2 months of continuous loading, bone adaptation and mineralization around all implants were analyzed. All the loaded implants were immobile. Significant marginal bone loss at the abutment-fixture interface (<1 mm) was observed around each loaded fixture implant. Bone remodeling was significantly more pronounced at the tension side of the implants, irrespective of fixture length. Radiographical and histological analyses showed bone with normal trabecular pattern around the implants. PMID- 10515137 TI - Cephalometric characteristics of nonobese patients with severe OSA. AB - The purpose of this study was to determine the facial characteristics of nonobese patients with obstructive sleep apnea (OSA). Observational data on a cohort of patients was analyzed retrospectively. The subjects were classified into four groups: nonobese mild, obese mild, nonobese severe, and obese severe. The nonobese mild group included patients with a body mass index (BMI = kilogram/meter2) <25 and an apnea-hypopnea index (AHI) >5 and <15; the obese mild patients had a BMI >35 and an AHI >5 and <15; the nonobese severe patients had a BMI <25 and an AHI >40; the obese severe group had a BMI >35 and AHI >40. Thirty three male patients referred for overnight polysomnography and lateral cephalometry who met the selection criteria were included. Between-group differences were examined pairwise by analysis of variance (ANOVA) with Bonferroni correction. Only two variables--lower facial height and overbite--were significantly different at p<0.05 between the nonobese severe group and the obese mild group. A discriminant analysis on the cephalometric measurements revealed that patients in the nonobese severe group could be distinguished from patients in other groups by their facial characteristics. OSA patients do not have a homogenous bony structure of the face. In particular, OSA severity in nonobese severe patients may be associated with a vertical skeletal disharmony. PMID- 10515138 TI - A longitudinal study of nasal airway size from age 9 to age 13. AB - In order to define nasal breathing for diagnostic purposes, reference values of normal nasal airway size in children are needed. The purpose of this study was to examine longitudinally changes in nasal airway size that occur with age. Minimum nasal cross-sectional areas of 82 children were measured by the pressure-flow technique at 1-year intervals, from age 9 through age 13. A mixed factorial ANOVA showed that the effect of age on nasal airway size was statistically significant (p<0.001) and the effect of gender was nonsignificant. Although the mean nasal size increased from 0.4 cm2 to 0.5 cm2, it also decreased at some point between 9 and 13 years. The results suggest that the adult nasal size may be reached earlier than previously reported in cross-sectional studies. PMID- 10515139 TI - Relationship of the functional oropharynx to craniofacial morphology. AB - The association between the functional oropharyngeal airway (defined as the minimal sagittal dimension at right angles to the airstream) and craniofacial morphology was investigated using 16 craniofacial variables taken from lateral cephalometric radiographs. The sample consisted of 70 subjects (31 males and 39 females) 10 to 13 years of age. There was no difference in ages between males and females, and no correlation with age except upper face height. Oropharyngeal airway was positively correlated with length of the mandible (Gon-Men), the distance between the third cervical vertebra and the hyoid bone (C3-Hy), and cranial base angle (NSBa). Although short mandibular length is a characteristic finding in patients with obstructive sleep apnea, none of the subjects in this study had this diagnosis. PMID- 10515140 TI - Incisal changes and orthodontic stability. AB - Lateral cephalograms and study casts of 55 patients were evaluated to determine if any relationships exist among incisal positions and angulations, changes in positions and angulations, and long-term occlusal stability. No significant relationships could be found between long-term changes occurring in a number of commonly used incisal measurements and end-of-treatment incisal positions, changes in incisal positions during treatment, or long-term changes in the facial axis angle, ANB angle, or weighted PAR score. Long-term incisal changes occurring in individual patients were not necessarily associated with negative occlusal changes. Since incisal positions usually change in the long-term, it is suggested that the use of published norms or recommended absolute goals for end-of treatment incisal positions be used more as general functional and esthetic clinical guides, rather than as predictors of stability. PMID- 10515141 TI - An investigation into the effects of polishing on surface hardness and corrosion of orthodontic archwires. AB - The purpose of this study was to investigate the effect of surface roughness on the relative corrosion rates of wires of four alloys-stainless steel, nickel titanium, cobalt chromium, and beta titanium. Batches of wire were divided into two groups. Wires in one group were industrially polished to provide a uniform surface finish; wires in the other group were left for comparison "as received." Wire diameter, hardness, and relative corrosion rates were compared within groups before and after polishing. Comparisons were also made across the four groups of alloys. The samples of as-received wires showed variations in surface finish, with beta titanium having the roughest appearance and cobalt chromium the smoothest. Nickel titanium and stainless steel surfaces were similar. Polishing provided a more uniform finish, but significantly reduced the diameter of the wires. Microhardness testing of wire surfaces of each alloy indicated that no significant work-hardening occurred as a result of polishing. The relative corrosion rates (expressed in terms of corrosion current density) in a 0.9% sodium chloride solution were estimated using the electrochemical technique of polarization resistance. Nickel titanium wires exhibited the greatest corrosion current density in the as-received state. Polishing significantly reduced the corrosion rate of nickel titanium, such that comparison between the four alloys in the polished state revealed no significant difference in their relative corrosion rate/corrosion current density. PMID- 10515142 TI - Stress-relaxing composite ligature wires: formulations and characteristics. AB - A stress-relaxing composite ligature was developed that has both mechanical and esthetic characteristics that make it attractive for use in orthodontics. The neutrally-colored polymer-polymer composite was created by encasing ultra-high molecular weight poly(ethylene) fibers in a poly(n-butyl methacrylate) polymer, which was formulated from a polysol and an optimal benzoin ethyl-ether concentration. The resulting composite ligature exhibited a tensile strength more than twice that of dead-soft stainless steel ligature, and a stress-relaxation decay significantly greater than stainless steel ligature. With these characteristics, the material could be used as an orthodontic ligature when tooth movement with negligible friction due to ligation is desired. A Maxwell-Weichert model predicted the load-decay profiles that ultimately resulted in the general loss of frictional forces with time. PMID- 10515143 TI - Effect of contamination and etching on enamel bond strength of new light-cured glass ionomer cements. AB - The effect of water and saliva contamination on the bond strength of metal orthodontic brackets cemented to etched (10% polyacrylic acid) and unetched human premolar enamel was investigated. Two bonding agents were used: one commercially available product (LC) and one experimental (EX) light-cured glass ionomer. Shear bond strength was measured after aging for 5 minutes, 15 minutes, and 24 hours. The results were compared by ANOVA and Scheffe's tests at p = 0.05. For LC, the bond strength of brackets bonded to etched enamel, with and without contamination, was statistically higher than that of brackets bonded to unetched enamel for all aging times. An exception was the bond strength to unetched enamel with saliva contamination after 24 hours; for EX, this value was statistically higher than that measured on unetched enamel with water contamination. Contamination by saliva did not reduce bond strength to unetched enamel. For both etched and unetched enamel, there was no significant difference between LC and EX after 24 hours for all contamination conditions. PMID- 10515144 TI - Comparison of shear bond strength of three bonding agents with metal and ceramic brackets. AB - Shear bond strengths of a light-cured composite resin, a light-cured glass ionomer cement, and a light-cured compomer used with metal and ceramic brackets were compared, and ARI scores were evaluated. Ceramic brackets showed statistically higher shear bond strengths than metal brackets when bonded with all test materials (p<0.001). When used with metal brackets, the light-cured glass ionomer cement (LCGIC) and compomer materials demonstrated statistically lower shear bond strengths than the light-cured composite (p<0.01 and p<0.001, respectively). When used with ceramic brackets, LCGIC was found to have significantly lower shear bond strength than the composite material (p<0.001). Despite its relatively low shear bond strength, LCGIC demonstrated optimal bonding values (8.39+/-3.24 MPa) with ceramic brackets. Bond failures within the LCGIC groups occurred at the adhesive-tooth interface, whereas in the compomer and composite groups, failures were detected at the adhesive-bracket interface. In the metal bracket group, clinically acceptable shear bond strength was obtained only with the composite resin (7.06+/-1.65 MPa). Compomer and LCGIC demonstrated values well below the accepted standard for metal brackets (4.32+/ 1.75 MPa and 4.45+/-1.06, respectively), while in the ceramic bracket group, values for composite and compomer were above the desired level (14.40+/-5.88 MPa and 12.31+/-6.09, respectively). LCGIC showed reasonably good bond strength with ceramic brackets, suggesting that this material may be considered suitable for use with ceramic brackets in clinical situations where moisture cannot be controlled. PMID- 10515145 TI - Comparison of bracket debonding force between two conventional resin adhesives and a resin-reinforced glass ionomer cement: an in vitro and in vivo study. AB - The purpose of this study was to compare the debonding force of orthodontic brackets bonded with two conventional resin adhesives (Resilience L3 and Light Bond) and a resin-reinforced glass ionomer cement (Fuji Ortho LC). For the in vitro part of the study, 80 extracted premolars were randomly divided into four groups. In groups A and B, brackets were bonded to unetched enamel using Fuji Ortho LC cement in wet and dry conditions, respectively. In groups C and D, brackets were bonded to etched enamel using Resilience L3 and Light Bond, respectively. Debonding force was determined using a servohydraulic testing machine at a crosshead speed of 1 mm/min. Data was analyzed using the ANOVA and Tukey-Kramer multiple comparison test at p<0.05. A significant difference was found in debonding force between unetched Fuji Ortho LC and the two conventional resins. There was no significant difference between the two conventional resins or between unetched resin-reinforced glass ionomer in the wet and dry conditions. For the in vivo part of the study, 30 patients were randomly assigned to one of the three bonding material groups. Bracket survival rates and distributions were obtained by following these patients for 1.2 years. Data was analyzed using the Kaplan-Meier product-limit estimates of survivorship function. Bond failure interface was determined using a modified adhesive remnant index (ARI). These results showed no significant difference between survival rates and distributions among the three bonding materials with respect to the type of malocclusion, type of orthodontic treatment, or location of bracket. There were significant differences between survival distributions of males and females in the unetched Fuji Ortho LC group and among type of teeth in the conventional resin groups. The predominant mode of bracket failure for the unetched Fuji Ortho LC cement was at the enamel-adhesive interface, and for conventional resins, the enamel-adhesive interface and the bracket-adhesive interface. These results suggest that resin reinforced glass ionomer cement can withstand occlusal and orthodontic forces despite having a bond strength lower than that of conventional resin adhesives. PMID- 10515146 TI - Orthodontic treatment of openbite and deepbite high-angle malocclusions. AB - The aim of the investigation was to assess the effect of orthodontic treatment on dentoskeletal morphology in children with openbite and deepbite high-angle malocclusion. Subjects (n = 54) in the mixed dentition with a hyperdivergent mandibular plane angle (high-angle, NSL/ML > or =40 degrees) were surveyed. Pre- and posttreatment lateral roentgenographic cephalograms were analyzed. Subjects were divided into three subgroups according to the amount of pretreatment overbite: < 0 mm = insufficient/no compensation (openbite); 0-4 mm = acceptable compensation (normal overbite); >4 mm = overcompensation (deepbite). Pretreatment, 20% of the high-angle cases exhibited insufficient dentoskeletal compensation (overbite <0 mm), and 35% displayed overcompensation (overbite >4 mm). Influences of habits such as lip sucking and tongue-thrust swallowing were more common in the openbite group. No major difference in treatment approach could be found between subgroups. In 82% of the openbite group and 90% of the deepbite group, overbite was corrected by orthodontic treatment. The mandibular plane angle was unaffected in both groups. The mechanisms of overbite correction differed between groups. The openbite group exhibited a significant decrease in interjaw-base angle. Increases in anterior and posterior dentoalveolar heights were comparable. The deepbite group showed no significant changes in skeletal morphology. The increase in dentoalveolar height was approximately twice as large posteriorly as anteriorly. The majority of children (80%) with high-angle morphology had a positive pretreatment overbite, thus exhibiting compensation of jaw-base hyperdivergency. Orthodontic treatment of high-angle malocclusions did not influence the mandibular plane angle in openbite or deepbite cases. Overbite correction was accomplished by tipping the maxilla downward anteriorly in openbite subjects, and by controlling incisor eruption in deepbite subjects. PMID- 10515148 TI - Hydroxyurea and didanosine long-term treatment prevents HIV breakthrough and normalizes immune parameters. AB - Hydroxyurea and didanosine treatment suppressed HIV replication for more than 2 years, in the absence of viral breakthrough, in chronically infected patients. The profile of viral load reduction was unusual for a two-drug combination, since a continuous gradual decrease in viremia persisted despite residual viral replication. The increase in CD4+ T cell counts was not robust. However, unlike those of patients treated by other therapies, CD4+ T lymphocytes were functionally competent against HIV, mediating a vigorous HIV-specific helper T cell response in half of these patients. In addition, the percentages of naive CD4+ and CD8+ T lymphocytes were not different from those in uninfected individuals. These results demonstrate that prolonged antiretroviral therapy with a simple, well-tolerated combination of two affordable drugs can lead to sustained control of HIV, normalization of immune parameters, and specific anti HIV immune response. PMID- 10515147 TI - Intrapatient variability of HIV type 1 group O ANT70 during a 10-year follow-up. AB - HIV-1 ANT70 is the first HIV-1 group O virus isolate obtained from a 25-year-old Cameroonian woman, who seroconverted in March 1987. This individual has remained asymptomatic and clinically healthy (clinical stage WHO 1, CDC II) even though she did not receive any antiretroviral therapy for HIV-1 before 97 months post seroconversion. CD4+ T cell counts declined steadily to 200/microl at 70 months postseroconversion. The HIV-1 ANT70 nucleotide and amino acid sequence diversity of the V3C3-encoding env fragment within this individual was followed over a 10 year period. RT-PCR, cloning, sequencing, and genetic analyses were performed on eight plasma follow-up samples. Extensive increasing intra- and intersample variation was observed. This is the first long-term (>10 years) follow-up of the genetic variability of an HIV-1 group O-infected individual. As the course of the disease in the HIV-1 ANT70-infected woman was similar in many aspects to that of group M-infected individuals, it remains to be elucidated whether the changes observed in the V3 loop are critical for disease progression. PMID- 10515150 TI - Spontaneous activation of human immunodeficiency virus type 1 in an immunotoxin resistant variant T cell line. AB - We have previously used immunotoxins to select for HIV-1 variant cells that transcribe HIV at extremely low levels and fail to produce HIV proteins. Further observation of one of these variants (E9) demonstrated spontaneous in vitro activation of HIV production. The mechanism of activation is different from that of U1 and ACH-2, two other cell lines that can be activated to express HIV in vitro. PMID- 10515149 TI - The benign cystic lymphoepithelial lesion of the parotid gland is a viral reservoir in HIV type 1-infected patients. AB - The presence of HIV-1 in cystic fluid aspirates from six cases of benign cystic lymphoepithelial lesion (BLL) of the parotid gland, a rare disorder affecting HIV 1-infected patients, has been investigated. HIV-1 p24 protein was present at a concentration ranging from 3 to 15 ng/ml, while it was undetectable in the peripheral blood of the same patients. The number of RNA copies of HIV-1 in the cystic fluids was high, ranging from 0.5 x 10(7) to 7.2 x 10(7) RNA copies/ml. BLL cystic fluid aspirates, despite the high level of HIV-1 RNA, were found to contain only a few infectious virions. The low infectivity correlated with the infrequent detection by electron microscopy of complete HIV-1 particles. The pathogenic mechanism leading to virus accumulation in the cystic fluid was studied by immunohistochemistry of tissue sections. p24 protein was associated with DRC-1+/S-100+ follicular dendritic reticulum cells, which were also present within the cystic cavities. Our findings are consistent with the possibility that the large amounts of virus present in the fluid derive from continuous shedding of HIV-1-infected cells from the surrounding lymphoid tissue. PMID- 10515151 TI - Influence of Rex and intronic sequences on expression of spliced mRNAs produced by human T cell leukemia virus type I. AB - Expression of the incompletely spliced HTLV-I mRNAs relies on the viral posttranscriptional activator Rex, whose interaction with the Rex-responsive element (RXRE) overcomes effects of cis-acting repressive sequences (CRSs). Studies based on heterologous reporter plasmids identified an intronic CRS in the 5' LTR and a CRS that overlaps with the RXRE. The present study investigated the effects of these elements in the context of spliced viral mRNAs encoding p21Rex (mRNA 1-3), Tax/Rex (mRNA 1-2-3), and Tof (mRNA 1-2-B). All three mRNAs were inefficiently expressed when transcribed in their mature intronless form, with the p21Rex mRNA showing the weakest expression. In contrast, efficient expression of p21Rex was obtained from a plasmid containing the 5' LTR and 3' portion of the genome that encoded a spliceable RNA. The defective expression of the intronless mRNAs reflected the inhibitory activity of the RXRE and the lack of 5' intronic sequences. Insertion of an intronic 5' LTR segment located upstream of the 5' CRS overcame Rex dependence conferred by the RXRE. The activity of this segment was mapped to the major splice donor and sequences overlapping with, but functionally distinct from, a previously described transcriptional enhancer. The three mRNAs responded differently to Rex and to insertion of the constitutive transport element of simian retrovirus type 1. Taken together, these results suggest that expression of the spliced mRNAs is controlled by the relative influence of positive and negative sequences present on the primary transcript as well as the Rex-RXRE interaction. PMID- 10515152 TI - Comparison of the distribution of IgG and IgA antibodies in serum and various mucosal fluids of HIV type 1-infected subjects. AB - We compared IgG and IgA distribution in serum, three different salivary samples, two different rectal secretion samples, cervicovaginal secretions, and seminal secretions from asymptomatic CDC stage II/III HIV-1-infected subjects (n = 44) and from HIV-1-seronegative volunteers (n = 52). In-house ELISAs were used to measure total IgG and total IgA levels, as well as HIV-specific anti-gp120 MN and anti-p24 LAI IgG and IgA. Human serum albumin was titrated in parallel to calculate the relative coefficient of excretion (RCE). In spite of substantial interindividual variability, total IgG concentrations in all fluids were found to be significantly greater in the HIV-1-infected group than in the seronegative subjects. Calculation of RCE values revealed three different types of mucosal secretion: secretions with no local Ig production, such as sperm; secretions with local production of IgA and transudative origin of IgG, such as salivary and rectal samples; and secretions with local production of both IgG and IgA, such as in cervicovaginal secretions. For all mucosal specimens from HIV-1-infected subjects, the response to HIV-1 was predominantly IgG, with highest titers observed in cervicovaginal secretions (although these were lower than serum levels). In contrast, the specific IgA response appeared weaker in the mucosa than in serum. PMID- 10515153 TI - Investigation of recombinant human insulin-like growth factor type I in thymus regeneration in the acute stage of experimental FIV infection in juvenile cats. AB - Thymus involvement and the development of thymic lesions in HIV-1 infection is hypothesized to suppress thymus function and limit T cell maturation and replenishment of the peripheral lymphoid pool. Therapeutic modulation to protect or enhance thymus function may therefore ameliorate peripheral lymphocytopenia and retard disease progression. Thymotrophic agents, such as insulin-like growth factor type I (IGF-I), may therefore represent adjunctive but important methods of treatment to protect or promote thymus function. The assessment of rhIGF-I in lentiviral infection and its impact on the thymus was performed using the feline immunodeficiency virus (FIV) model. Regeneration of the thymus in juvenile cats and amelioration of the thymic lesion after FIV infection was assessed by multiple measurements including thymic weight, stereologic analysis of the thymus cortex and medulla, histologic and immunohistologic analysis, quantitation of thymocyte and peripheral lymphocyte subsets, and quantitative competitive RT-PCR. Evidence of thymic cortical regeneration was observed in FIV-inoculated cats after 12 and 20 weeks of rhIGF-I treatment. Inflammation in the thymus was reduced during this period of treatment in this group of rhIGF-I/FIV-inoculated cats as evidenced by the reduced numbers of B cells detected. Viral replication rates in peripheral lymph nodes were not altered by rhIGF-I treatment and were decreased by 1 log in the thymus after 20 weeks of treatment. Peripheral blood CD4+ T cell counts also increased after 14 weeks of treatment. This suggests that rhIGF-I treatment can enhance thymus function and replenishment of the peripheral T cell pool. PMID- 10515154 TI - Simian AIDS-associated lymphoma in rhesus and cynomolgus monkeys recapitulates the primary pathobiological features of AIDS-associated non-Hodgkin's lymphoma. AB - Non-Hodgkin's lymphomas occur with increased frequency (3-6%) in HIV-infected individuals. These AIDS-associated lymphomas (AALs) exhibit characteristics that distinguish them from lymphomas in the general population. A proposed model for the pathogenesis of AAL includes the following: (1) Tumorigenesis is multistep; (2) tumors occur in long-term survivors; (3) tumors are of clonal B cell origin; (4) HIV acts early and is an indirect effector; (5) tumor cells are infected with EBV; and (6) specific genetic lesions occur in tumor cells. Many aspects of this process remain to be tested in an animal model system. Since 1984, necropsy examinations have been performed on more than 1000 SIV-infected rhesus and cynomolgus monkeys at the Tulane Regional Primate Research Center. Lymphoid malignancies were detected in a proportion of SIV-infected animals. These SAIDS associated lymphomas (SALs) have been studied to determine the extent to which their pathological features recapitulate a working model for the pathogenesis of AAL. The results show that lymphomas occur in SIV-infected rhesus macaques at 4% incidence, similar to that of AAL, and that the incidence of SAL in cynomolgus macaques is eightfold higher. Analysis of SAL from both species of macaques demonstrated significant similarity to the hallmark pathobiological features of AAL. These findings indicate that the HIV-infected human and the SIV-infected macaque share a common pathobiology and mechanism of lymphomagenesis. PMID- 10515156 TI - Increase in activated CD4+ lymphocytes with CCR5 and CXCR4 in HIV type 1-infected persons. PMID- 10515155 TI - Alterations in the V1/V2 domain of HIV-2CBL24 glycoprotein 105 correlate with an extended cell tropism. PMID- 10515157 TI - Genetic basis of the human epilepsies. AB - Genetic factors contribute to aetiology in up to 40% of patients with epilepsy. Over 100 single gene Mendelian disorders include epilepsy as one component of what is usually a complex neurological phenotype, but the majority of idiopathic or primary epilepsies display a 'complex' non-Mendelian pattern of inheritance. There have been significant recent advances in understanding the genetic basis of inherited epilepsies at a molecular level. Epilepsy genes fall into several distinct categories including those in which mutations cause abnormal brain development, progressive neurodegeneration, disturbed energy metabolism and abnormal function of ion channels. Ion channel genes involved include those encoding neuronal nicotinic acetylcholine receptor subunits and voltage-gated potassium and sodium channels. PMID- 10515158 TI - Detecting genes in new and old mouse models for epilepsy: a prospectus through the magnifying glass. AB - Various spontaneous mutants and natural strain variants for either generalized tonic-clonic seizures, or non-convulsive absence seizures have been described in mice and rats over the years. Convulsive seizure models are usually ascertained by mere visual observation, while finding the less noticeable seizures of absence models often requires proactive screening of existing mutants with other phenotypes. To date, molecular cloning technologies has elucidated the primary basis of most of the known single locus epilepsy mutants. Together with the 20 or so mouse knockouts with seizure-related phenotypes described to date, the frequency at which the mutants appear and diversity of the proteins involved would suggest that 1000 or more genes can be mutated to give rise to influence epilepsy phenotypes. As many of these genes will cluster into molecular, cellular and developmental pathways, their identification may be very important for better understanding epileptic mechanisms. With this perspective, the approaches taken towards positional cloning of mouse epilepsy mutations is illustrated by comparing and contrasting the different stages of gene identification in three different models with which this author has been fortunate enough to be intimately involved: slow-wave epilepsy (common gene symbol: swe, Chr 4); tottering (tg, Chr 8); and stargazer (stg, Chr 15). The comparatively sobering outlook for positional cloning of the more common genetically 'complex' epilepsies will also be discussed, as will more efficient new strategies for model screening and identifying the remaining 985 (or so) genes. PMID- 10515159 TI - Single gene defects in mice: the role of voltage-dependent calcium channels in absence models. AB - Nineteen genes encoding alpha1, beta, gamma, or alpha2delta voltage-dependent calcium channel subunits have been identified to date. Recent studies have found that three of these genes are mutated in mice with generalised cortical spike wave discharges (models of human absence epilepsy), emphasising the importance of calcium channels in regulating the expression of this inherited seizure phenotype. The tottering (tg) locus encodes the calcium channel alpha1 subunit gene Cacna1a, lethargic (lh) encodes the beta subunit gene Cacnb4, and stargazer (stg) encodes the gamma subunit gene Cacng2. These calcium channel mutants should provide important insights into the basic mechanisms of neuronal synchronisation, and the genes may be considered candidates for involvement in similar human disorders. The mutant models offer an important opportunity to elucidate the molecular, developmental, and physiological mechanisms underlying one subtype of absence epilepsy. Since calcium channels are involved in numerous cellular functions, including proliferation and differentiation, membrane excitability, neurite outgrowth and synaptogenesis, signal transduction, and gene expression, their role in generating the absence epilepsy phenotype may be complex. A comparative analysis of channel function and neural excitability patterns in tottering, lethargic, and stargazer brain should be useful in identifying the common elements of calcium channel involvement in these absence models. PMID- 10515160 TI - GABA and epileptogenesis: comparing gabrb3 gene-deficient mice with Angelman syndrome in man. AB - The GABAergic system has long been implicated in epilepsy with defects in GABA neurotransmission being linked to epilepsy in both experimental animal models and human syndromes (Olsen and Avoli, 1997). However, to date no human epileptic syndrome has been directly attributed to an altered GABAergic system. The observed defects in GABA neurotransmission in human epileptic syndromes may be the indirect result of a brain besieged by seizures. The use of animal models of epilepsy has sought to address these matters. The advent of gene targeting methodologies in mice now allows for a more direct assessment of GABA's involvement in epileptogenesis. To date several genes associated with the GABAergic system have been disrupted. These include the genes for glutamic acid decarboxylase, both the 65- and 67-kDa isoforms (GAD65 and GAD67), the tissue non specific alkaline phosphatase gene (TNAP) and genes for the GABA(A) receptor subunits alpha6, beta3, gamma2, and delta (gabra6, gabrb3, gabrg2, and gabrd respectively). Gene disruptions of either GAD67 or gabrg2 result in neonatal lethality, while others, GAD65, TNAP, and gabrb3 exhibit increased mortality and spontaneous seizures. GABA receptor expression has been found to be both regionally and developmentally regulated. Thus in addition to their obvious role in controlling excitability in adult brain, a deficit in GABAergic function during development could be expected to elicit pleiotropic neurodevelopmental abnormalities perhaps including epilepsy. The GABA(A) receptor beta3 subunit gene, gabrb3/GABRB3 (mouse/human), is of particular interest because of its expression early in development and its possible role in the neurodevelopmental disorder Angelman syndrome. Individuals with this syndrome exhibit severe mental retardation and epilepsy. Mice with the gabrb3 gene disrupted likewise exhibit electroencephalograph (EEG) abnormalities, seizures, and behavioral characteristics typically associated with Angelman syndrome. These gabrb3 gene knockout mice provide direct evidence that a reduction of a specific subunit of the GABA(A) receptor system can result in epilepsy and support a GABAergic role in the pathophysiology of Angelman syndrome. PMID- 10515161 TI - Neuronal migration disorders in humans and in mouse models--an overview. AB - The spectrum of neuronal migration disorders (NMD) in humans encompasses developmental brain defects with a range of clinical and pathological features. A simple classification distinguishes agyria/pachygyria, heterotopia, polymicrogyria and cortical dysplasia as distinct clinico-pathological entities. Many of these conditions are associated with intractable epilepsy. When considering the pathogenesis of NMD, a critical developmental process is the migration of neuroblasts along the processes of radial glia during the formation of the layered structure of the cerebral cortex. In addition, faulty cytodifferentiation and programmed cell death play important roles in the generation of dysplasias and heterotopias respectively. A number of genes have been identified that participate in the regulation of neuronal migration. Mouse models, in which these genes are mutated, provide insight into the developmental pathways that underlie normal and abnormal neuronal migration. PMID- 10515162 TI - Genes that regulate neuronal migration in the cerebral cortex. AB - Malformations of cortical development are increasingly recognized as causes of mental retardation and epilepsy. However, little is known about the molecular and biochemical signals that control the proliferation, migration, and organization of the cells involved in normal cerebral cortical development. Analysis of genes required for cortical development will help elucidate the pathogenesis of some epilepsies. In humans, two striking examples of abnormal cortical development, with varying degrees of epilepsy and mental retardation, are 'double cortex' and lissencephaly. Double cortex (DC), also known as subcortical band heterotopia, shows an abnormal band of neurons in the white matter underlying a relatively normal cortex. In pedigrees, DC often occurs in females, whereas affected males show more severe lissencephaly (XLIS), i.e. an abnormally thick cortex with decreased or absent surface convolutions. We and others have identified a novel brain specific gene, doublecortin, that is mutated in Double Cortex/X-linked lissencephaly (DC/XLIS) patients. Although the cellular function of doublecortin (DCX) is unknown, sequence analysis reveals a cytoplasmic protein with potential MAP kinase phosphorylation sites, as well as a site that is likely to be phosphorylated by c-Abl, suggesting that doublecortin functions as an intracellular signaling molecule critical for the migration of developing neurons. Interestingly, the scrambler mouse mutant demonstrates abnormal lamination with some similarity to lissencephaly and reflects a mutation in the murine homolog of the Drosophila disabled gene, mdab1, which binds c-Abl. Although a direct interaction between doublecortin and mDab1 has not been demonstrated, it is plausible that these two proteins may be part of a common signaling pathway. Therefore, abnormalities in signal transduction may be an underlying mechanism for the neuronal migration defects in DC/XLIS and the scrambler mouse, but further research is necessary to determine how such abnormalities give rise to cortical malformations and epilepsy. PMID- 10515163 TI - Focal cortical dysplasia: a neuropathological and developmental perspective. AB - Focal cortical dysplasia (FCD) is a rare, sporadic disorder which is a recognised cause of chronic epilepsy. It is proposed to result from disordered neuronal migration and differentiation and has characteristic histological features which include disturbed cortical lamination, large abnormal neurons and the presence of large balloon cells with glassy eosinophilic cytoplasm and pleomorphic eccentric nuclei. These latter express both glial and neuronal markers indicative of abnormal neuroglial differentiation. In this paper we review the current literature on the neuropathology of FCD and discuss potential mechanisms. We focus on growth factors, signalling pathways and candidate genes with known roles in Drosophila and vertebrate brain development that could be responsible for the developmental brain changes seen in FCD. At issue are the factors that influence cell fate and differentiation and which regulate neural migration. Some of the molecular pathways, such as those involving the Notch and the Wnt pathways have particularly important roles in neuroglial differentiation in vertebrates, and these are proposed as potential candidates. PMID- 10515165 TI - Glutamate receptors and transporters in genetic and acquired models of epilepsy. AB - Glutamate, the principal excitatory neurotransmitter in the brain, acts on three families of ionotropic receptor--AMPA (alpha-amino-3-hydroxy-5-methyl-isoxazole-4 propionic acid), kainate and NMDA (N-methyl-D-aspartate) receptors and three families of metabotropic receptor (Group I: mGlu1 and mGlu5; Group II: mGlu2 and mGlu3; Group III: mGlu4, mGlu6, mGlu7 and mGlu8). Glutamate is removed from the synaptic cleft and the extracellular space by Na+-dependent transporters (GLAST/EAAT1, GLT/EAAT2, EAAC/EAAT3, EAAT4, EAAT5). In rodents, genetic manipulations relating to the expression or function of glutamate receptor proteins can induce epilepsy syndromes or raise seizure threshold. Decreased expression of glutamate transporters (EAAC knockdown, GLT knockout) can lead to seizures. In acquired epilepsy syndromes, a wide variety of changes in receptors and transporters have been described. Electrically-induced kindling in the rat is associated with functional potentiation of NMDA receptor-mediated responses at various limbic sites. Group I metabotropic responses are enhanced in the amygdala. To date, no genetic epilepsy in man has been identified in which the primary genetic defect involves glutamate receptors or transporters. Changes are found in some acquired syndromes, including enhanced NMDA receptor responses in dentate granule cells in patients with hippocampal sclerosis. PMID- 10515164 TI - Mechanisms underlying epileptogenesis in cortical malformations. AB - The presence of developmental cortical malformations is associated with epileptogenesis and other neurological disorders. In recent years, animal models specific to certain malformations have been developed to study the underlying epileptogenic mechanisms. Teratogens (chemical, thermal or radiation) applied during cortical neuroblast division and migration result in lissencephaly and focal cortical dysplasia. Animals with these malformations have a lowered seizure threshold as well as histopathologies typical of those found in human dysgenic brains. Alterations that may promote epileptogenesis have been identified in lissencephalic brains, such as increased numbers of bursting types of neurons, and abnormal connections between hippocampus, subcortical heterotopia, and neocortex. A distinct set of pathological properties is present in animal models of 4-layered microgyria, induced with cortical lesions made during late stages of cortical neuroblast migration. Hyperexcitability has been demonstrated in cortex adjacent to the microgyrus (paramicrogyral zone) in in vitro slice preparations. A number of observations suggest that cellular differentiation is delayed in microgyric brains. Other studies show increases in postsynaptic glutamate receptors and decreases in GABA(A) receptors in microgyric cortex. These alterations could promote epileptogenesis, depending on which cell types have the altered receptors. The microgyrus lacks thalamic afferents from sensory relay nuclei, that instead appear to project to the paramicrogyral region, thereby increasing excitatory connectivity within this epileptogenic zone. These studies have provided a necessary first step in understanding molecular and cellular mechanisms of epileptogenesis associated with cortical malformations. PMID- 10515166 TI - Molecular neuropathology of human mesial temporal lobe epilepsy. AB - With the recent progress in surgical treatment modalities, human brain tissue from patients with intractable focal epilepsies will increasingly become available for studies on the molecular pathology, electrophysiological changes and pathogenesis of human focal epilepsies. An inherent problem for studies on human temporal lobe epilepsy (TLE) is the lack of suitable controls. Strategies to alleviate this obstacle include the use of human post mortem samples, hippocampus from experimental animals and, in particular, the comparative analysis of surgical specimens from patients with Ammon's horn sclerosis (AHS) and with focal temporal lesions but anatomically preserved hippocampal structures. In this review we focus on selected aspects of the molecular neuropathology of TLE: (1) the potential impact of persisting calretinin immunoreactive neurons with Cajal-Retzius cell morphology, (2) astrocytic tenascin-C induction and redistribution as potential regulator of aberrant axonal sprouting and (3) alterations of Ca2+ -mediated hippocampal signalling pathways. The diverse and complex changes described so far in human TLE specimens require a systematic interdisciplinary approach to distinguish primary, epileptogenic alterations and secondary, compensatory mechanisms in the pathogenesis of human temporal lobe epilepsies. PMID- 10515167 TI - Nuclear calcium-activated gene expression: possible roles in neuronal plasticity and epileptogenesis. AB - Nuclear calcium signals associated with electrical activation of neurons are critical regulators of gene expression and may cause changes in neuronal structure and function. Recent studies have identified a key component of the transcriptional machinery, the coactivator CREB binding protein (CBP), as a target for a nuclear calcium signalling pathway. Because the regulation of many genes involves transcription factors that function through their interaction with CBP, this mechanism, termed 'the coactivator control model', may modulate the expression of a large number of genes. During normal working of the brain, nuclear calcium increases may be transient and initiate transcriptional responses that are important for learning and memory. However, more intense or sustained stimulations of neurons (for example those used in the kindling model) may overactivate nuclear calcium-regulated processes. This may initiate inappropriate gene expression responses and could lead to the formation of epileptic neuronal circuits and disorders of neuronal excitability. PMID- 10515168 TI - Effect of long-term spontaneous recurrent seizures or reinduction of status epilepticus on the development of supragranular mossy fiber sprouting. AB - In a recent report we have shown that a protein synthesis inhibitor, cycloheximide (CHX), is able to block the mossy fiber sprouting (MFS) that would otherwise be triggered by pilocarpine (Pilo)-induced status epilepticus (SE), and also gives relative protection against hippocampal neuronal death. Under this condition animals still showed spontaneous recurrent seizures (SRS) which led us to question the role played by sprouted mossy fibers in generating those seizures. In both patients and animal models of epilepsy the relative contribution of SE (when present) and/or SRS for the development of MFS is not known. In the present study we investigated the relationship between MFS, SE and SRS, and evaluated whether the CHX-induced blockade of MFS was transient or permanent in nature. We performed a chronic study which included animals subject to Pilo-induced SE in the presence of CHX and sacrificed between 8 and 10 months later, and animals that were subject to Pilo-induced SE in the presence of CHX and underwent a reinduction of SE with Pilo, 45 days after the first induction, but this time in the absence of CHX. Re-induction of SE or a long period of chronic seizures, were able to trigger supragranular MFS even in animals where the first (or only) SE event was triggered in the presence of CHX. MFS did not show any association with the frequency of SRS, and thus seemed to depend more critically on time. Our current findings allow us to suggest that MFS are neither the cause nor the consequence of SRS in the pilocarpine model. PMID- 10515169 TI - Expression of mRNAs encoding flip isoforms of GluR1 and GluR2 glutamate receptors is increased in rat hippocampus in epilepsy induced by tetanus toxin. AB - The messenger RNAs encoding the flip and flop isoforms of the glutamate receptor subunits GluR1 and GluR2 were detected and quantified by in situ hybridization in the hippocampal formation of rats following intrahippocampal injection of tetanus on one side. The mRNAs encoding the flip isoforms of both GluR1 and GluR2 were significantly increased 4 weeks after injection. The GluR1 flip mRNA was significantly elevated only in the dentate gyrus, whereas significant increases in the GluR2 flip mRNA were seen in all hippocampal subfields examined. There were no significant changes in the mRNA encoding the flop isoforms of either GluR1 or GluR2. The significant changes in flip isoform mRNAs occurred on both sides. PMID- 10515170 TI - Differences in evoked potential characteristics between DRPLA patients and patients with progressive myoclonic epilepsy: preliminary findings indicating usefulness for differential diagnosis. AB - The characteristics of evoked potentials in patients with dentatorubral pallidoluysian atrophy (DRPLA) were investigated. Twelve patients with DRPLA and three patients with progressive myoclonic epilepsy (PME) attributable to other causes participated in the study. In 11 out of the 12 patients, the diagnosis of DRPLA was genetically confirmed, based on a 56-75 CAG triplet repeat expansion on chromosome 12p; in the remaining patient, the diagnosis was not genetically confirmed but the patient was clinically diagnosed as having DRPLA and was within the same pedigree as one of the 11 genetically confirmed patients. Two out of the three patients with PME, who had been tested for dodecamer repeat expansion in the cystatin B gene, were genetically confirmed as having Unverricht-Lundborg disease (UL); the remaining patient was also clinically diagnosed as having UL, but the patient did not have the aforementioned genetic abnormality. Somatosensory evoked potentials (SEPs) and brainstem auditory evoked responses (BAERs) were recorded. The amplitudes of the SEPs were determined as the peak-to peak amplitudes between P2 and N2 deflections. The results revealed that high amplitude SEPs were not evoked in any of the DRPLA patients; on the other hand, high-amplitude SEPs were evoked in all the patients with UL. Moreover, BAERs were absent in seven out of the 12 patients with DRPLA; on the other hand, all UL patients showed BAERs in which all peaks, from I to V, were distinguishable. These results suggest differences in pathophysiology between DRPLA, which predominantly affects the brainstem and subcortical regions, and PME, characterized by cortical hyperexcitability. Thus, evoked potential measurements may be useful to differentiate DRPLA patients from those with progressive myoclonic epilepsy. PMID- 10515171 TI - The relationship between memory performance, perceived cognitive function, and mood in patients with epilepsy. AB - OBJECTIVE: The low correlations between memory performance and subjective memory may be attributable to disparities between tasks in neuropsychological tests and cognitive experiences of day-to-day living. This study evaluated the relationship between everyday memory performance, perceived cognitive functioning, and mood among patients with epilepsy. METHODS: From three epilepsy centers in the USA, 138 patients were recruited. Everyday memory performance was measured using the Rivermead Behavioural Memory Test (RBMT). Questionnaires assessed perceived cognitive function (cognitive domain, Quality of Life in Epilepsy Inventory, QOLIE-89) and mood (Profile of Mood States, POMS). RESULTS: Memory performance scores were weakly correlated with perceived cognitive functioning (r =0.22, P < 0.01). Perceived cognitive functioning was strongly correlated with mood (r = - 0.75, P < 0.0001). Multiple regression analysis indicated memory performance (RBMT) and mood (POMS) were independent predictors of perceived cognitive functioning (P < 0.02); however, the explained variance for RBMT and POMS combined (R2 = 0.58) is only slightly higher than the predictive value for the POMS score alone (R2 = 0.56). CONCLUSIONS: Memory performance tests provide qualitatively different information than patients' self-reported cognitive difficulties, thus it is important to assess memory performance, perceived cognitive function, and mood separately because the constructs are related but not redundant. PMID- 10515172 TI - Activity-dependent enhancement of hyperpolarizing and depolarizing gamma aminobutyric acid (GABA) synaptic responses following inhibition of GABA uptake by tiagabine. AB - The effects of the 7-aminobutyric acid (GABA) uptake blocker tiagabine on isolated inhibitory postsynaptic potentials (IPSPs) were examined in CA1 pyramidal cells of the rat hippocampal slice preparation. The IPSPs were elicited by either single stimuli or by high frequency (100 Hz, 200 ms) stimulation (HFS) of inhibitory interneurons. Bath applied tiagabine (20 microM) produced little or no increase in the amplitude of IPSPs evoked by low (30-50 microA) or high (200 400 microA) intensity single stimuli. Only the duration of IPSPs evoked by high intensity stimuli was substantially prolonged by tiagabine, the time integral of the hyperpolarizing response being increased 3.2-fold. HFS elicited much larger fast and slow IPSPs than a single stimulus. In addition, with increments in the intensity (80-550 microA) of HFS, a GABA(A) receptor-mediated depolarizing response of progressively larger amplitude appeared between, and overlapped with, the fast and slow hyperpolarizing components of the IPSP. Tiagabine application markedly increased the GABA-mediated responses evoked by both low and high intensity HFS. Increasing the intensity of HFS enhanced the drug effect. Thus, measurements of the time integral of evoked responses showed that with weak (60 microA) HFS, tiagabine caused a 3.6-fold increase in the area of hyperpolarization while, in contrast, with strong (530 microA) HFS, tiagabine produced a 13.5-fold increase in the depolarizing actions of GABA. Our results suggest that tiagabine, a therapeutically effective anticonvulsant, may paradoxically increase, through a GABA(A) receptor-mediated mechanism, neuronal depolarization during the high frequency discharge of neurons involved in epileptiform activity. PMID- 10515173 TI - Anticonvulsant action of 2-chloroadenosine injected focally into the perirhinal cortex in amygdaloid kindled rats. AB - Possible anticonvulsant effects of 2-chloroadenosine injected focally into the perirhinal cortex of amygdala kindled rats were investigated over a 2 h period. Animals were microinfused (1 microl) with 2-chloroadenosine (2-CLA; 5, 10, 15, 25 and 100 nM) or artificial cerebrospinal fluid applied through a cannula located in the perirhinal cortex. At the doses employed, 2-CLA significantly reduced afterdischarge duration and stage 5 seizure duration. The latency to stage 4 seizure was increased only at the highest dose of 2-CLA (100 nM), while even at this dose no significant change in seizure stage could be seen. The maximum effect of 2-CLA was obtained 30 min after microinfusion of the drug. Pre treatment (intraperirhinal cortex) of animals with the nonselective adenosine antagonist, caffeine (50 microM; 1 microl), blocked the anticonvulsant activity of 2-CLA. These results suggest that adenosine receptors located in the perirhinal cortex may play an important role in the suppression of seizure activity elicited from the amygdala. PMID- 10515174 TI - Effect of physical exercise on seizure occurrence in a model of temporal lobe epilepsy in rats. AB - Although the favorable effect of physical fitness on general health is unquestionable, physical exercise and fitness programs in patients with epilepsy are still a matter of controversy. Little objective evidence regarding the effect of exercise on seizure frequency and severity has been reported. One sought to clarify the relationship between exercise and epilepsy in an animal model of temporal lobe epilepsy (the pilocarpine model of epilepsy). To evaluate the effect of an aerobic physical program on seizure frequency, 29 epileptic animals were continuously monitored during 24 h for 135 days after the first spontaneous recurrent seizure (SRS) and divided into three groups. The first group (N = 14) was submitted to an aerobic exercise program (training group). The second group (N = 7) was maintained in the treadmill for the same time as the training group without being submitted to physical exercise (sham group). The third group (N = 8) served as control. The behavioral observation was divided in three periods of 45 days. The first period was used to determine the number of seizures before physical training program. The second period was utilized to determine the number of seizures during the physical training program. The third period was used to analyze the frequency of seizures after the physical training program. The mean frequency of seizures in the control and sham groups increased significantly from period 1 to period 2 and from period 1 to period 3. However, in the training group, the frequency of seizures did not change significantly between the three periods of 45 days of observation. When the same periods of the three groups were analyzed together, a significant reduction in seizure frequency was observed comparing the training group with the control and sham groups during the period of physical training. The data presented in this study suggest that physical exercise is not a seizure-inducing factor in this experimental model of epilepsy. PMID- 10515175 TI - Amygdala sclerosis in sudden and unexpected death in epilepsy. AB - Sclerosis of the amygdala is a not uncommon finding in patients with chronic epilepsy. The amygdala has efferent connections, via the central nuclei, to cardioregulatory centres in the medulla. Experimental studies have suggested that damage to the central nucleus may be of functional significance in patients with sudden and unexpected death in epilepsy (SUDEP) in particular with regard to their susceptibility to cardiac arrhythmias. We investigated this possibility by carrying out a quantitative immunohistochemical analysis of the patterns of neuronal loss and gliosis in three amygdala subnuclei (central, basal and lateral) in post mortem material from 15 SUDEP cases and seven normal controls. We identified significant neuronal loss in the medial division of the lateral amygdaloid nucleus in SUDEP cases but not in central or basal nuclei. These patterns of cell loss in the amygdala do not differ from previous studies in both humans and animal models of chronic epilepsy suggesting that there is not a specific pattern of amygdaloid sclerosis in SUDEP patients which could implicate a functional role for this nucleus in the mechanism of the sudden death. PMID- 10515176 TI - Reduced density of parvalbumin- and calbindin D28-immunoreactive neurons in experimental cortical dysplasia. AB - Cortical dysplasia (CD) is a congenital brain malformation in humans that is closely associated with intractable epilepsy. This study utilized an animal model of CD, in utero irradiation in rats, to determine if experimental dysplastic cortex demonstrates a reduction in the density of inhibitory interneurons. Fetal rats were exposed to external irradiation on gestational day 17 to produce diffuse CD and heterotopic grey matter. As adults, these rats were processed for immunohistochemistry using primary antibodies for parvalbumin (PA), calbindin D28k (CA), the 67 kD subunit of glutamic acid decarboxylase (GAD67), and cresyl violet staining. Quantitative methods were used to determine the density of immunoreactive neurons and cresyl violet-stained neurons in the neocortex at the rostro-caudal level of the anterior commissure. Compared to control values, the density of PA- and CA-immunoreactive neurons was reduced in dysplastic cortex. Density of glutamic acic decarboxylase-immunoreactive neurons was not different between control and dysplastic cortex. Overall neuronal density, measured in cresyl violet-stained sections, was not significantly different between control and dysplastic cortex. These data suggest a selective reduction in inhibitory interneurons in experimental CD cortex or an impaired ability for these neurons to produce PA and CA. PMID- 10515177 TI - Morbidity and infection in combined subdural grid and strip electrode investigation for intractable epilepsy. AB - The coverage of large surface areas of the brain for electrographic monitoring purposes necessitates a craniotomy to achieve comprehensive sampling. We undertook a review and prospective analysis over 3 years of 38 patients undergoing craniotomy for electrode implantation. The indication for invasive monitoring was to determine candidacy for resective surgery in patients whose seizure focus was not well localized by scalp electroencephalography and other noninvasive testing. Prophylactic cultures from the epidural space were obtained at electrode removal. There were five positive epidural cultures. All five patients went on to seizure-free status. Two positive cultures occurred in patients without obvious infection and who were not treated with antibiotics. Other complications included individual cases of atrial fibrillation, pulmonary edema, postoperative fever, and epidural hematoma. There was no mortality or permanent neurologic morbidity related to craniotomy for electrode placement. There was a 7.9% rate of clinical infection per patient and a 5.7% rate per craniotomy side. This study has identified several factors that significantly correlate with positive epidural culture results: > 100 electrodes, more than ten cables, more than 14 days of implantation, and more than one cable exit site. PMID- 10515178 TI - Multicentre, double-blind, randomised comparison between lamotrigine and carbamazepine in elderly patients with newly diagnosed epilepsy. The UK Lamotrigine Elderly Study Group. AB - In a multicentre, double-blind trial 150 elderly patients (mean age 77 years) with newly diagnosed epilepsy were randomised in a 2:1 ratio to treatment with lamotrigine (LTG) or carbamazepine (CBZ). Following a short titration period, the dosage was individualised for each patient while maintaining the blind over the next 24 weeks. The main difference between the groups was the rate of drop-out due to adverse events (LTG 18% versus CBZ 42%). This was in part a consequence of the lower rash rate with LTG (LTG 3%, CBZ 19%; 95% CI 7-25%). LTG-treated patients also complained less frequently of somnolence (LTG 12%, CBZ 29%; 95% CI 4-30%). Although there was no difference between the drugs in time to first seizure, a greater percentage of LTG-treated patients remained seizure-free during the last 16 weeks of treatment (LTG 39%, CBZ 21%; P = 0.027). Overall, more patients continued on treatment with LTG than CBZ (LTG 71%, CBZ 42%; P < 0.001) for the duration of the study. The hazard ratio for withdrawal was 2.4 (95% CI 1.4-4.0) indicating that a patient treated with CBZ was more than twice as likely to come off medication than one taking LTG. In conclusion, LTG can be regarded as an acceptable choice as initial treatment for elderly patients with newly diagnosed epilepsy. PMID- 10515179 TI - Heme oxygenase-2 expression at rat neuromuscular junctions. AB - The neuromuscular junction is specialized for rapid transmission of electrical signals. Nitric oxide synthase (NOS) is concentrated at the junction, and NO modulates transmission and could influence signaling pathways. Increasing evidence suggests that carbon monoxide (CO) serves as a neurotransmitter, and heme oxygenase (HO), the enzyme that catalyzes the formation of CO, is often colocalized with NOS. Immunoreactivity for HO-2 was present at rat neuromuscular junctions of leg muscles and persisted in denervated muscle indicating the localization of the enzyme to the postsynaptic surface. In contrast, HO-2 immunoreactivity was absent from the en grappe and orbital en plaque endplates of extraocular muscle (EOM), while only the global en plaque endplates possessed HO 2 immunoreactivity. The difference between EOM and leg endplates may arise from EOM's unique physiology. The presence of HO-2 at neuromuscular junctions suggests CO could serve as a pre- and post-synaptic messenger. PMID- 10515180 TI - Differential effects of treatment with typical and atypical antipsychotic drugs on adenylyl cyclase and G proteins. AB - We examined the effects of chronic in vivo antipsychotic drug treatments on G protein function and regulation. Mice were treated with typical antipsychotic haloperidol (6 mg/kg per day) and atypical agent olanzapine (20 mg/kg per day) for 14 days via mini-osmotic pumps. G protein-activated adenylyl cyclase activity in brain tissues was measured in the presence of guanine nucleotide analogue guanosine-5'-O(3-thiotriphosphate) tetralithium salt, or GTPgammaS. In frontal cortex, haloperidol treatment produced 21% increases in the GTPgammaS -mediated adenylyl cyclase Emax value (vs. vehicle controls) while olanzapine produced 20% reductions in this value (vs. controls); these effects were significant. In striatum, olanzapine treatment produced significant 31 and 27% decreases in Emax values compared with vehicle and haloperidol treatment, respectively. Chronic haloperidol treatment produced significant 24% reductions in the immunoreactivity of cortical, but not striatal, Gialpha1,2 subunits. There were no effects of chronic olanzapine treatment on G(i)alpha1,2 levels and no effects of either antipsychotic on G(s)alpha, levels. Chronic haloperidol and olanzapine treatments differentially regulate G protein-mediated adenylyl cyclase responses in brain regions possibly relating to their unique effects on G protein-coupled receptors. PMID- 10515181 TI - Dopamine receptor D2 intronic polymorphism in patients with Parkinson's disease. AB - An association between the intronic allele 3 of the dopamine receptor D2 (DRD2) gene and European Parkinson's disease (PD) cases has been reported recently. We initiated the present work in order to determine whether this association between the DRD2 locus and PD is also present in our population from Spain. The DRD2 gene polymorphism has been genotyped in 154 patients and in 125 controls. The allele 3 is present in 60.3% of the patients and in 55.2% of the controls. The genotype 3/3 is present in 36.3% of the patients and in 34.4% of the controls. No statistical differences in the genotype and allelic frequencies between the two groups have been found. No differences were also found when the patients were classified according to different criteria such as onset, family history, gender or clinical presentation. Thus our results do not support a role for the DRD2 locus to develop PD. PMID- 10515182 TI - Altered seizure susceptibility after high-frequency transcranial magnetic stimulation in rats. AB - The long-term effect of repetitive transcranial magnetic stimulation (rTMS) on the susceptibility of amygdala kindling was studied. Two weeks after a single high-frequency rTMS train (120 A/micros, 20 Hz for 3 s), the rats had a 55% higher threshold for induction of epileptic afterdischarges compared with sham treated or control rats. However, subsequent kindling revealed no difference between rTMS-treated and control rats. Our data suggest that a single rTMS train has long-term effects on the neuronal excitability. These effects may be anticonvulsant and therefore support the safety of rTMS in clinical use. PMID- 10515183 TI - Non-invasive long-term recordings of cortical 'direct current' (DC-) activity in humans using magnetoencephalography. AB - Recently, biomagnetic fields below 0.1 Hz arising from nerve or muscle injury currents have been measured non-invasively using superconducting quantum interference devices (SQUIDs). Here we report first long-term recordings of cortical direct current (DC) fields in humans based on a horizontal modulation (0.4 Hz) of the body and, respectively, head position beneath the sensor array: near-DC fields with amplitudes between 90 and 540 fT were detected in 5/5 subjects over the auditory cortex throughout prolonged stimulation periods (here: 30 s) during which subjects were listening to concert music. These results prove the feasibility to record non-invasively low amplitude near-DC magnetic fields of the human brain and open the perspective for studies on DC-phenomena in stroke, such as anoxic depolarization or periinfarct depolarization, and in migraine patients. PMID- 10515184 TI - Cytidine diphosphate choline administration activates brain cytidine triphosphate: phosphocholine cytidylytransferase in aged rats. AB - Beneficial effects of cytidine (5') diphosphocholine (CDP-choline) administration on several diseases including brain aging, ischemia and stroke are based on an increase in membrane phospholipid turnover. We have studied the possible involvement of CTP:phosphocholine cytidylyltransferase (CT) in this mechanism by measuring its gene expression and enzyme activity in the brains of young and aged rats treated with 500 mg/kg per day of CDP-choline. Older animals showed higher (57%) of total CT activity in particulate (active) fraction than younger animals (46%). Treatment of aged animals for 8, 16, or 60 days had no effect on the CT gene expression but increased activation of the CT by translocation to membranes. The particulate fraction rose from 57% of total activity to more than 65% after 2 months of treatment. This may explain the long-term repairing effects of CDP choline on damaged membranes of aged animals. PMID- 10515185 TI - Characterization of a putative calcitonin receptor in IMR 32 human neuroblastoma cells. AB - In this study we characterized calcitonin (CT) receptors in human neuroblastoma IMR 32 cells. Saturation binding assays indicated that [125I]-human CT bound with high affinity to IMR 32 cell membranes (K(d) = 253.6 pM; Bmax = 3.84 fmol/ mg protein). In competition binding studies, human adrenomedullin displayed high affinity for these sites (IC50 = 30 nM) whereas human alpha calcitonin-gene related peptide (alphaCGRP; IC50 = 145 nM) and human amylin (IC50 = 415 nM) showed lower affinity. These peptides increased cAMP levels in viable cells; the relative potencies were: human CT > human adrenomedullin > human cCGRP > or = human amylin. The expression of mRNA coding for the published sequences of the human calcitonin receptor and of the human calcitonin receptor-like receptorwas evaluated by reverse transcriptase-polymerase chain reaction. Electrophoretic analysis did not confirm the occurrence of mRNA coding for the above mentioned receptors in these cells. This study suggests the presence of a novel, putative CT receptor in IMR 32 cells. PMID- 10515186 TI - Inhibition of MK801 binding in adult rat brain sections by conantokin-G and conantokin-T. AB - The functional interactions of conantokins with anatomical sites in rat brain have been assessed through displacement of the non-competitive N-methyl-D aspartate receptor (NMDAR) antagonist, dizocilpine (MK801). The binding of (+)-3 [125I]-iodo-MK801 (1 nM) to coronal sections from adult rat brain was inhibited in a dose-dependent manner by conantokin-T (con-T) and conantokin-G (con-G). Quantitative densitometry was used to determine IC50 values for conantokin inhibition of [125I]-MK801 binding in the cortex, thalamus and hippocampus. Con-T completely inhibited [125I]-MK801 specific binding in all brain regions at a saturating concentration of 100 microM. Con-G was able to completely displace [125I]-MK801 in the cortex and thalamus, but only inhibited this same binding up to approximately 90% in the hippocampus. Both peptides maintained their inhibitory properties in the presence of 1 mM EDTA, suggesting that divalent cations are not required for their action in this regard. The added presence of spermine (150 microM) resulted in a two-fold increase in [125I]-MK801 binding and a two-fold decrease in the IC50 values for both peptides. The data obtained in this investigation further demonstrate that [125I]-MK801 is a useful probe for the indirect determination of functional NMDAR ligand binding sites in rat brain sections. PMID- 10515187 TI - Calcitonin reverts pertussis toxin blockade of the opioid analgesia in mice. AB - The aim of this paper is to study the influence of salmon calcitonin (SCT) on opioid analgesia when opioid transduction pathways are functionally uncoupled from Gi/o proteins by treatment with pertussis toxin (PTX). The antinociceptive effect of morphine and three selective opioid agonists, [D-Ala2,N-Me-Phe2,Gly5 ol]enkephalin (DAMGO) (OP(3-mu receptor agonist), [D-Pen2.5]-enkephalin (OP-1 delta receptor agonist) and trans-( +/- )-3,4-dichloro-N-methyl-N-[2-1( pyrrolidinyl)-cyclohexyl]-benzene-acetam ide methane sulfonate (U-50, 488H) (OP1 kappareceptor agonist) was evaluated, using the tail flick test, in mice treated with PTX or with PTX and SCT. PTX blocked the antinociceptive effect of the opioids, being the antinociception similar in control animals and in mice treated with PTX and SCT. Thus, SCT prevents the effect of the blockade of Gi/o-proteins. From this it could be suggested that calcitonin activates alternative antinociceptive mechanisms that are not dependent on Gi/o-proteins. PMID- 10515188 TI - Mechanical transduction by rat dorsal root ganglion neurons in vitro. AB - Although it is generally presumed that mechanical sensitivity of somatosensory nerve fibers results from the activation of mechanosensitive ion channels, a mechanically-gated whole-cell current has never been demonstrated in dorsal root ganglion (DRG) neurons. We performed patch clamp experiments on rat DRG neurons in culture, and report the first mechanically-activated current in somatosensory neurons (I(mech)). This whole-cell current is observed in most dorsal root ganglion neurons but not in non-sensory sympathetic ganglion neurons. The current voltage relation of I(mech) indicates that it is a non-selective cation current. Sensitivity of I(mech) to block by gadolinium suggests that it may be mediated by a member of a family of mechanosensitive non-selective cation channels observed in many cell types. Sensitivity to benzamil supports this idea, and further suggests that the current might be mediated by a member of the degenerin/ epithelial sodium channel (DEG/ENaC) family. PMID- 10515190 TI - Reciprocal changes in the expression of neurotrophin mRNAs in target tissues and peripheral nerves of aged rats. AB - trk receptors are downregulated in both dorsal root ganglion (DRG) and spinal motoneurons of aged rats with behavioral sensorimotor deficits. Here we provide evidence, using reverse transcription-polymerase chain reaction (RT-PCR), of decreased levels of neurotrophin (nerve growth factor, NGF; brain-derived neurotrophic factor, BDNF; neurotrophin-3, NT-3; and neurotrophin-4, NT-4) mRNAs in target muscles. Moreover, the degree of neurotrophin mRNA decrease in target muscles seems to co-vary with the extent of sensorimotor disturbances. In contrast, the peripheral nerve of aged rats showed a reciprocal regulation of neurotrophins, with increased levels of NGF, BDNF, and NT-4 mRNAs. Taken together, evidence suggest an aging-related attenuation of neurotrophin signaling between target tissues, on one hand, and DRG and motoneurons, on the other, and, furthermore, that target-derived neurotrophins regulate the expression levels of trk mRNAs in both DRG and motoneurons. PMID- 10515189 TI - Site-specific splice variation of the human P2X4 receptor. AB - P2X4 receptors are expressed in specific brain areas. We now describe site specific splice variations of the human P2X4 receptor subunit, occurring at residue [YVIG / WVFV(W)] near the end of the first predicted transmembrane domain. p2X4(b) is formed by the insertion of an additional 16 amino acids. p2X4(C) is formed by deleting a cassette of 130 amino acids, including six of the 10 conserved extracellular cysteine residues. Transfection of P2X4(a), but not p2x4(c), formed functional channels in Xenopus oocytes and human 1321N1 cells. After transfection of p2X4(b) small, inconsistent ATP-evoked responses were detected only in the human cells, but when co-expressed, p2x4(b) may alter the function of P2X4(a) in oocytes. The distribution of splice variant RNA within human brain suggests regionally-dependent expression. These data indicate that the functions of the human P2X4 receptor may be altered by alternative splicing. PMID- 10515191 TI - LY379268, a potent and selective Group II metabotropic glutamate receptor agonist, is neuroprotective in gerbil global, but not focal, cerebral ischaemia. AB - The neuroprotective effects of a selective Group II metabotropic glutamate receptor (mGluR) agonist, LY379268, have been evaluated against global and focal cerebral ischaemia. Loss of CA1 hippocampal neurones following 5 min bilateral occlusion of the carotid artery (BCAO) in the gerbil was almost completely prevented by LY379268 (10 mg/kg, i.p.) given 30 min post-occlusion (P < 0.001); 10 mg/kg 1 h after and 20 mg/kg 2 h after BCAO also produced significant neuroprotection (P < 0.05). Similarly the BCAO-induced increase in TUNEL positive cells at 5 days post-occlusion was reduced by LY379268. By contrast the size of the infarct following middle cerebral artery occlusion (MCAO) induced by endothelin-1 infusion in the rat was unaffected by either 10 or 20 mg/kg i.p. of LY379268. This contrast between the results from these two animal models with LY379268, agrees with previous data on a less potent but similarly selective mGluR2/3 agonist, LY354740. It further suggests that mGluR Group II agonists are likely to have more utility in global, than in focal, cerebral ischaemia. PMID- 10515192 TI - A functional magnetic resonance imaging study of mental subtraction in human subjects. AB - The neuronal network involved in a precise type of calculation procedure, mental subtraction, was investigated by means of functional magnetic resonance imaging. Two tasks were used requiring covert production of numbers: (1) with calculation; (2) without calculation. During the first task, activation was observed in the left dorsolateral prefrontal and premotor cortices, in Broca's area and bilaterally in the inferior parietal cortex. During the second task, activation was mainly observed in Broca's area and to a less extent in the left prefrontal and premotor cortices. Statistical comparison of data in the two situations revealed that the procedure of mental subtraction is mediated by a distributed system which includes predominantly the left dorsolateral prefrontal cortex and the inferior parietal cortex bilaterally. PMID- 10515193 TI - Muscarinic control of intracortical connections to layer II in rat entorhinal cortex slice. AB - The cholinergic system is critically involved in oscillatory network activity and synaptic plasticity in the entorhinal cortex (EC) hippocampal formation. Here we demonstrate robust inhibition of field potentials in layer II of the medial EC evoked by stimulation in the deep EC or in the lateral layer II by carbachol (CCh, 0.1-100 microM, K(D) approximately 1 microM). This effect appears not to be mediated by suppression of presynaptic Ca(2+)-signals since paired pulse facilitation was increased by CCh. Blockade of the effect by the muscarinic antagonists atropine and pirenzepine demonstrates mediation by muscarinic receptors, most likely of the M1 subtype. The effect is characterized by absence of desensitization and should be important for laminar shaping of oscillatory activity and synaptic plasticity during acetylcholine-dependent theta-rhythmic activity. PMID- 10515194 TI - Cerebrospinal fluid levels of amyloid beta-peptide1-42, but not tau have positive correlation with brain glucose metabolism in humans. AB - To address the question of whether assay for cerebrospinal fluid (CSF) levels of amyloid beta-peptide 1-42 (A(beta)1-42) and tau allow us to monitor the neurodegenerative processes that lead to a progressive and massive death of neurons in Alzheimer's disease (AD) and non-AD patients, cerebral glucose metabolism using 2-[18F] fluoro-2-deoxy-glucose was quantified by positron emission tomography in fifteen AD patients and nine non-AD patients with defined levels of CSF-A(beta)1-42 and CSF-tau. The CSF-A(beta)1-42 levels, but not the CSF-tau levels, in both AD and non-AD patients consistently and significantly correlated with global and, in particular, temporal lobe glucose metabolism. Results from our study suggest that the CSF-A(beta)1-42 levels may reflect residual brain function and help monitoring progression of dementing disorders. PMID- 10515195 TI - A supernumerary tooth in a 1.7 million-year-old Australopithecus robustus from Swartkrans, South Africa. AB - The maxillary dental arch of a partial cranium of an adult specimen of Australopithecus robustus shows the presence of a supernumerary tooth between the right first and second incisors. The fossil specimen, SK 83, is from Swartkrans Member 1 sediments, considered, on the basis of associated fauna, to be approximately 1.7 million yr old. The specimen was analyzed macroscopically, stereomacroscopically and by X-ray computed tomography. The anatomic position and the morphology of the tooth are consistent with a diagnosis of supernumerary lateral incisor rather than a mesiodens or a retained deciduous tooth. This is the first description of a supernumerary tooth found in Plio-Pleistocene hominid fossils from South African cave deposits. PMID- 10515196 TI - Reliability and validity of the Dutch version of the Social Attributes of Dental Anxiety Scale. AB - The aim of the present study was to assess the reliability and (factorial) validity of the Dutch version of the Social Attributes of Dental Anxiety Scale (SADAS). A factor analysis using the English version of the SADAS revealed two separate scales. The first eight items involved unwanted psychological upsets when patients encounter dental care directly; the four remaining items were about social inhibitions or restrictions due to the perceived state of oral health. Psychometric properties of the Dutch version were assessed using a sample of 170 highly anxious dental patients of a dental fear clinic in Amsterdam, The Netherlands. The Dental Anxiety Scale (DAS) and the Short version of the Dental Anxiety Inventory (S-DAI) were used as measures of dental anxiety. Factor analysis revealed that four factors explained 72.8% of the variance, and two forcedly extracted factors explained 53.4% of the variance. Correlations indicated that the SADAS does measure a different concept than dental anxiety. In addition, t-tests indicated that the SADAS was able to discriminate between a group of non-anxious individuals and the present group of patients. In conclusion, the SADAS is a promising new questionnaire with moderate factorial, but with more than sufficient reliability, as well as construct and discriminant validity. PMID- 10515197 TI - Chitinase in whole and glandular human salivas and in whole saliva of patients with periodontal inflammation. AB - In recent studies the existence of a chitinase in various mammals, like man, was described. The aim of the present study was to find out whether salivas of periodontally healthy and inflamed humans also contain chitinase activity. Chitinase activity, assayed with the substrate 4-methylumbelliferyl-beta-D N,N',N"-triacetylchitotrioside, was shown to be present in human whole saliva, with an activity level and apparent molecular mass (35 kDa) that were comparable with those of the human serum enzyme. Both lysozyme and beta-N acetylhexosaminidase could be separated from chitinase by means of Bio-Gel P-100 gel filtration chromatography. The enzyme was also present in glandular saliva of parotid, palatine, submandibular and sublingual glands. The chitinase activity was not of oral epithelial, bacterial or plaque bacterial origin and was not correlated with the activity of salivary amylase. A comparative study of whole salivas of periodontally healthy controls and gingivitis and periodontitis subjects showed that only in the case of periodontitis there was a significant increase of the specific chitinase activity. The latter enzyme showed a gel filtration pattern that was comparable with that of the enzyme from controls. The measured albumin levels in saliva and the absence of correlation between the chitinase activity levels in plasma and saliva from periodontitis patients indicated that the (increased) chitinase activities did not originate from blood leakage to the oral cavity. PMID- 10515198 TI - Chemosensitivity testing of oral cancer cells treated with a p185neu-specific agent. AB - The amplification and overexpression of the erbB-2 oncogene and its involvement in tumorigenesis makes this receptor an appropriate target for specific agents directed towards tumor cells. The purpose of this study was to evaluate the in vitro effect of the bacterially produced recombinant immunotoxin scFv(FRP5)-ETA on the protein synthesis and adenosine triphosphate (ATP) reduction in oral squamous cell carcinoma (OSCC) cells. This agent recognizes the erbB-2 receptor and inhibits protein synthesis in receptor-overexpressing cells. OSCC cells were selected for this study, and amplification and expression levels of the erbB-2 receptor were determined. Cell suspensions were cultured for 6 d with various concentrations of scFv(FRP5)-ETA (1-1000 ng/ml). A431 and MDA-MB468 cell lines were used as controls. Chemosensibility of tumor cells was measured by [3H]leucine incorporation assay and by an ATP luminescence assay. In OSCC cells with amplification and overexpression of erbB-2 inhibition, up to 92% of protein synthesis and 90% of ATP reduction was observed when cells were exposed to 1,000 ng/ml immunotoxin. In OSCC cells showing a deletion of erbB-2 and in erbB-2 negative MDA-MB468 cells, protein synthesis was inhibited by 22% and 8%, respectively. These results indicate that the effectiveness of a recombinant immunotoxin targeting erbB-2 receptors in OSCC cells depends on the level of erbB 2 amplification and expression, that it is highly specific for tumor cells expressing these receptors, and that a dose-dependency can be observed. PMID- 10515199 TI - Palatal mucoperiosteal wound healing in the rat. AB - The objective of this study was to analyze the changes in tissue architecture and matrix composition during healing of palatal wounds of immature rats, and to compare this with rats of the same age that did not receive mucoperiosteal wounds. Wounds were made in the mucoperiosteum of the palate of 35-d-old rats. Samples were evaluated histologically at numerous points in time after wounding. The DNA, hydroxyproline and sulphated glycosaminoglycan contents were determined at 8, 15, 30, and 60 d post-wounding. Eight-d-old granulation tissue contained 43% less hydroxyproline, and 100% more glycosaminoglycans and cells than unwounded palatal tissue of 43-d-old rats. Sixty-d-old wounds contained 100% more DNA and 39% more hydroxyproline than unwounded tissue of 95-d-old rats. At the same time, densely packed and transversely aligned collagen fibres were present. It is concluded that palatal mucoperiosteal wounds made in 35-d-old rats heal with distinct scar tissue formation. The scar contains more collagen than non wounded palatal tissue of rats of the same age. Therefore, this model may be of use for the development of therapies aiming to reduce palatal scarring. PMID- 10515200 TI - Activation of recombinant bovine matrix metalloproteinase-20 and its hydrolysis of two amelogenin oligopeptides. AB - Enamelysin is a matrix metalloproteinase (MMP-20) secreted by ameloblasts, previously shown to hydrolyze recombinant amelogenin. The purpose of this study was to use recombinant MMP-20 to further investigate the specific hydrolysis of peptide fragments containing cleavage sites for tyrosine-rich amelogenin peptide (TRAP) and leucine-rich amelogenin peptide (LRAP). MMP-20 cDNA was isolated from a subtracted bovine cDNA library, reconstructed into pRSET A vector, and overexpressed in BL21 Escherichia coli. The recombinant MMP-20 was purified using Mono-S ion exchange and nickel affinity chromatography. The proteinase was renatured by dialysis against buffer containing 50 microM zinc and 5 mM calcium and autolysed to form several active fragments. The varying sizes and activities of the activated enzyme fragments appeared to be due to sequential autolysis at different location of the carboxyl terminus of the intact enzyme. Two synthetic peptides corresponding to amelogenin amino acid sequences 36-49 and 181-188 were hydrolyzed by the activated rMMP-20. Mass spectrometry and amino acid composition analysis showed that the cleavage sites were between the tryptophan and leucine (45 and 46) for TRAP and between proline and alanine (186-187) for LRAP. These results indicate that MMP-20 can be autoactivated, and activated MMP-20 has a functional role in the initial cleavage of amelogenin. PMID- 10515201 TI - Response of dental follicular cells to the exposure of denuded enamel matrix in rat molars. AB - Recent studies have indicated that cementum formation can be induced when dental follicular cells are exposed to enamel matrix. The purpose of the present investigation was to study this cementum formation and the appearance of the cells involved, including their expression of collagen types alpha1(I), alpha1(II) and alpha1(III) mRNAs, during this process by means of light microscopy and in situ hybridisation. The mandibular first molars of 5-d-old rats were surgically taken out, their enamel epithelium was removed, and then the crowns were re-inserted with the occlusal surface downwards in their crypts to allow the follicular cells to come in contact with the denuded enamel matrix. After observation periods of 2-14 d, the teeth were prepared for light microscopic examination and in situ hybridisation. A monolayer of follicular cells in contact with the exposed enamel matrix changed their morphology and increased their expression of collagen type I mRNA as early as 2-4 d after exposure to the enamel matrix. A cementum-like tissue was formed at the surface of the enamel matrix. Collagen type II mRNA was never expressed in the tissues studied, whereas collagen type III mRNA was weakly expressed in the follicular cells throughout the experiment. PMID- 10515202 TI - Immunolocalization of Bone Morphogenetic Protein-2 and -3 and Osteogenic Protein 1 during murine tooth root morphogenesis and in other craniofacial structures. AB - The distribution of Bone Morphogenetic Protein-2, and -3 (BMP-2 and BMP-3) and Osteogenic Protein-1 (OP-1, also known as BMP-7) during root morphogenesis and in other craniofacial structures was examined in sections of 12- to 18-d-old mouse heads using polyclonal and monoclonal antibodies. BMP-3 and OP-1 were localized in alveolar bone, cementum, and periodontal ligament, whereas BMP-2 was only localized in the alveolar bone of periodontium. All three BMPs were localized in predentine, dentine, odontoblasts, osteoblasts, osteocytes, osteoid, cartilage, chondrocytes and spiral limbus. BMP-2 and OP-1 were also localized in spiral ligament and interdentate cells of the cochlea, whilst BMP-3 was restricted to the spiral ganglion. BMP-3 was also localized in ducts of submandibular and sublingual salivary glands, acini of the lacrimal gland, Purkinje cells in the cerebellum, nerve fibres of the cerebellum and brain, afferent cells of the dorsal root ganglia, inferior alveolar nerve, and peripheral processes of the vestibulocochlear nerve. OP-1 was also localized in hair and whisker follicles, sclera of the eye and in ameloblasts. The demonstration of BMP-3 in the nervous system suggests that this protein may be neurotrophic during development and maintenance of the nervous system. The composite expression of BMPs/OPs during periodontal tissue morphogenesis suggests that optimal therapeutic regeneration may entail the combined use of different BMPs/OPs. PMID- 10515203 TI - Organic leachables from polymer-based dental filling materials. AB - Results are reported from a study on the in vitro separation and identification of leachables from three different polymer-based dental filling materials by using a combined method of gas chromatography and mass spectrometry. The median number of separable organic leachables in these materials was between 14 and 22. Of these organic leachables the following were identified and quantified: DL camphorquinone, 4-dimethylaminobenzoic acid ethyl ester (DMABEE), drometrizole, 1,7,7-trimethylbicyclo[2,2,1]heptane, 2,2-dimethoxy[1,2] diphenyletanone (DMBZ), ethyleneglycol dimethacrylate (EGDMA), and triethyleneglycol dimethacrylate (TEGDMA). Three of the leachables have previously been shown to provoke allergy. The range of log P(ow) values (representing the lipophilicity of these compounds) varied between 1.09 and 4.20. By multivariate data analysis, selected leachables from the tested materials were shown to separate into characteristic patterns. The results contribute to a characterization of potential hazardous compounds in polymer-based dental filling materials. PMID- 10515204 TI - Surface energy of non-corroded and corroded dental ceramic materials before and after contact with salivary proteins. AB - After sintering and autoglazing, the surface free energy of seven different dental ceramic materials was determined by contact angle measurements using a Dynamic Adsorption Tester and three different standard probe liquids before and after in vitro corrosion and also before and after incubation with human whole saliva. The surface free energy was calculated from the Young-Dupre equation, and the polar and non-polar components were determined. The total surface free energy (range 50 +/- 5 mN/m) did not differ significantly between the different ceramic materials or between corroded and non-corroded specimens. The basicity of the surface persisted after corrosion, although reduced to a lower level, which probably reflects ion exchange processes at the exposed surfaces. After saliva incubation, the basic (gamma-) component of the surface free energy increased, which may be interpreted as the result of salivary proteins binding to the predominantly basic (gamma-) surface with their anionic groups, orienting their cationic sites towards the bulk phase. Selective adsorption of proteins to biosurfaces will no doubt influence both microbiological colonisation and cell adhesion, although the exact mechanisms are not known. PMID- 10515205 TI - Resin-modified glass-ionomers: effect of dentin primer application on the development of bond strength. AB - The purpose of this study was to investigate the rate of development of dentin bond strengths of resin-modified glass-ionomer cements (RMGIC) with use of dentin primers. The prepared dentin surface was treated according to each manufacturer's instruction or the dentin primer. Cements were condensed into a vinyl mold and light activated. The shear bond strengths were measured at a crosshead speed of 1.0 mm/min after 1, 5, 10, 30, 60 min, 2, 5, and 24 h storage in water at 37 degrees C. Presence of a significant difference between the mean bond strength at 1 min and each of the other test times was analyzed. The first time at which there was a significant increase in bond strength was defined as the "initial increasing time". As compared with the manufacturer's suggested dentin treatments, the bond strengths increased significantly when the dentin primers were used. The initial increasing times when the specimens were made following each manufacturer's instructions were 10 approximately 60 min. When dentin primer was used, the initial increasing time shortened to 5 approximately 10 min. It was concluded that the use of dentin primers for RMGIC restorations resulted in reduction of the initial increasing time. PMID- 10515206 TI - In vitro dentin pretreatment: surface roughness and adhesive shear bond strength. AB - The aim was to define the morphology and roughness of dentin from different tooth areas after various pretreatments to identify the effect of hybrid layer, resin tags, and mineralised dentin surface on shear bond strength. Thirty-eight extracted molars were used, each providing two sections of cervical (c) and lateral (l) dentin. Five pretreatments were performed: A) 0.2% EDTA; B) abrasion with Al2O3 particles, 0.2% EDTA; C) 10% H3PO4; D) 10% H3PO4 and immersion in a collagenase solution; E) control: no treatment. Z100 composite resin cylinders were bonded to the specimens with All Bond 2 bonding resin and tested for shear bond strength. Twelve other specimens from each group were analysed with an optical profilometer and an atomic force microscope, and four were further examined by scanning electron microscopy (SEM). Mean shear strength values in MPa were: Ac: 8.36 +/- 4.23; Al: 8.77 +/- 3.68; Bc: 6.05 +/- 3.62; Bl: 8.39 +/- 4.60; Cc: 6.87 +/- 3.45; Cl: 9.00 +/- 5.62; Dc: 13.30 +/- 5.45; Dl: 8.44 +/- 4.47; Ec: 4.10 +/- 1.54; El: 6.09 +/- 4.34. No statistically significant difference for cervical versus lateral dentin was found within treatments except for group D. Treatments performed on lateral dentin did not differ significantly. In cervical dentin, A differed from E; C from E; and D from A, B, C and E. An increased surface roughness was found in group D. Shear bond strength to dentin did not seem to depend on a hybrid layer formation, but on the direct contact of the adhesive with the mineralised dentinal surface and partly on the orientation of the dentinal tubules. PMID- 10515207 TI - Somatic pairing between subtelomeric chromosome regions: implications for human genetic disease? AB - Fluorescence in-situ hybridization (FISH) has been used to study the spatial orientation of subtelomeric chromosome regions in the interphase nucleus. Compared to interstitial chromosomal sites, subtelomeres showed an increased number of somatic pairings. However, pairing frequency also depended on the specific regions involved and varied both between different subtelomeres and between different interstitial regions. An increased incidence of somatic pairing may play at least some role in the frequent involvement of the subtelomeres in cytogenetically cryptic chromosome rearrangements. In patients suffering from facioscapulohumeral muscular dystrophy (FSHD), which is associated with a deletion of subtelomeric repeats, the FSHD region on 4qter showed a changed pairing behavior, which could be indicative of a position effect and/or trans sensing effect as a cause for disease. PMID- 10515208 TI - Chromosomal homologies between Drosophila melanogaster and D. funebris determined by in-situ hybridization. AB - Seventeen biotin-labeled DNA sequences were hybridized to polytene chromosomes of Drosophila melanogaster and D. funebris in order to establish chromosomal homologies between these species. Ten probes correspond to cloned DNA sequences from D. melanogaster (RpII 215, MHC, H3-H4, Tor, hsp 68, hsp 28/23, hsp 83, PP1alpha, RpII 140, and ey), four are clones isolated from a D. subobscura genomic library (Xdh, lambdaDsubS3, lambdaDsubG3, and lambdaDsubG4), two are clones from D. funebris (F2 and Adh) and one from D. virilis (ci). The probes were chosen in order to cover all the autosomes, since X-chromosome homologies have already been studied by linkage analysis of morphological mutants. Most probes gave a unique hybridization signal; consequently, our results allow unambiguous inferences about chromosomal homologies. The results show extensive gene rearrangement within all chromosomal elements, probably due to paracentric inversions, but are consistent with Muller's proposal that chromosomal elements have conserved their genetic content during the evolution of Drosophila. PMID- 10515209 TI - Purification and initial characterization of primate satellite chromatin. AB - Nucleoprotein hybridization, a method for the purification of specific DNA sequences as chromatin, was employed to fractionate primate centromeric alpha satellite chromatin as a first step in the identification and analysis of novel centromere-enriched proteins. In order to optimize the amount of material available for further study, cultured African green monkey cells were employed because satellite DNA represents approximately 25% of the genome. Two chromatin preparations were compared for the yield and total protein content of purified material. Regardless of the preparation, alpha satellite sequences were enriched to near purity. Since intact satellite chromatin is relatively refractile to the enzymatic digestion steps in the method, the total amount of solubilized material available for purification is rather low. In contrast, nuclei treated with acidic washes to extract histone H1 provided solubilized material enriched in satellite sequences. In addition, this material is more efficiently utilized in an affinity chromatography step. However, the extraction of many non-histones at low pH resulted in very low yields of protein in the purified fraction. Two-dimensional gel comparisons of proteins associated with H1-containing satellite chromatin after iodination of total chromatin proteins revealed a number of polypeptides enriched to varying degrees in the purified fraction. The electrophoretic mobilities of a few enriched polypeptides corresponded to previously identified heterochromatin-associated proteins while many others appear to be novel. The work presented validates nucleoprotein hybridization as a purification method for highly repeated sequences as chromatin in analytical amounts. The fact that a number of the enriched proteins are visible in stained gels of bulk chromatin proteins suggests that further biochemical analysis can be carried out on these polypeptides directly. PMID- 10515210 TI - An easy and reliable procedure of microdissection technique for the analysis of chromosomal breakpoints and marker chromosomes. AB - Microdissection in combination with reverse painting fluorescence in-situ hybridization (FISH) is a very effective method to identify breakpoints and rearrangements of derived chromosomes and reveal the chromosomal origin of marker chromosomes. We describe an innovation that allows a convenient, fast and safe isolation of microdissected fragments as currently available protocols. The microdissected chromosomes are harvested in a collection drop located in a movable micropipette adjusted to a second micromanipulator under microscopic observation. We used this technique to analyze several cytogenetic aberrations. In order to evaluate the efficiency of our microdissection procedure, we compared the results obtained with microdissection probes made from only one fragment with those obtained with more than six microdissected fragments. In all cases, the single-fragment microdissections were sufficient to provide probes. PMID- 10515211 TI - Chromosomal localization of 5S rDNA genes in Leporinus fish (Anostomidae, Characiformes). AB - The large 45S rDNA chromosome sites have often been analyzed in fish. In contrast, little is known about the 5S genes in this animal group. In the genus Leporinus, the NOR chromosomal location has been shown to be very diverse. In the present work, chromosome mapping of 5S rDNA in three anostomids, Leporinus elongatus, L. obtusidens and L. friderici, is investigated using fluorescence in situ hybridization (FISH) with PCR-obtained 5S probes and primed in-situ labeling (PRINS). Major 5S rDNA chromosomal sites were found to be subterminally located in a small metacentric pair, while minor ones were detected near the centromeric region of a medium-sized submetacentric pair in all studied species. The 5S rDNA genes were not associated with the NORs or sex chromosomes. A highly conserved chromosomal location of these genes appears to characterize the karyotype evolution of this fish group. PMID- 10515212 TI - Bivalent 15 regularly associates with the sex vesicle in normal male meiosis. AB - Using fluorescent in-situ hybridization, we investigated the positioning of different human bivalents at the pachytene stage of normal male meiosis. We showed that, in about 35% of nuclei, the pericentromeric region of bivalent 15 is closely associated with the sex vesicle (SV). This behaviour may be linked to the presence of three domains in the pericentromeric region of chromosome 15: a large imprinted domain, a nucleolar organizing region (NOR), and a heterochromatic block. In order to define the domains of chromosome 15 involved in this association, we analysed the meiotic behaviour of other bivalents with similar domains: human bivalent 11 and mouse bivalent 7, bearing imprinted domains, other human acrocentric bivalents bearing a NOR, and the human bivalents 1, 9 and 16 containing a heterochromatic region. None of these bivalents were as frequently associated with the SV as the human bivalent 15. Nevertheless, we suggest that the bivalent 15 heterochromatin may be responsible for the association because of two properties: its telomeric location on chromosome 15 and its strong sequence homology with the Yq heterochromatin. This phenomenon could explain the high frequency of translocations between the chromosome 15 and the X or Y chromosomes. PMID- 10515214 TI - The 5S rRNA gene clusters have a defined orientation toward the nucleolus in Petunia hybrida and Crepis capillaris. AB - The 3D localization of the 5S ribosomal RNA genes was studied in cells of the cortex zone of roots in the plant species Petunia hybrida inbred line V26 and in Crepis capillaris. The analysis was carried out on interphase nuclei (both species) and on prophase nuclei (C. capillaris). The 5S ribosomal RNA genes were detected by fluorescence in-situ hybridization and 3D images were obtained by confocal scanning laser microscopy. In both plant species, the 5S ribosomal genes were localized at the short arm of chromosome 2, which, in both plants, also possesses a satellite at its end. Statistical and visual analysis of interphase nuclei showed that: (1) there is a preference for an association of the 5S rRNA gene clusters of the two homologous chromosomes, and (2) the 5S rRNA gene clusters in both species had a preserved spatial position within the interphase nucleus and they tended to be polarized with respect to their neighbouring cells (i.e. a relic telophase orientation). Moreover, tracing of the chromosomal segment between the 5S loci and the active NOR revealed that the homologous chromosomes during early/mid prophase were aligned and that they entered the nucleolus side by side, at least for these chromosome segments. We interpret our data to mean that location of 5S rRNA near the nucleolus favours their functioning in ribosome biogenesis. PMID- 10515213 TI - Identification of the gene-richest bands in human prometaphase chromosomes. AB - The human genome is a mosaic of long, compositionally homogeneous DNA segments, the isochores, that can be partitioned into five families, two GC-poor families (L1 and L2), representing 63% of the genome, and three GC-rich families (H1, H2 and H3), representing 24%, 7.5% and 4-5% of the genome, respectively. Gene concentration increases with increasing GC levels, reaching a level 20-fold higher in H3 compared with L isochores. In-situ hybridization of DNA from different isochore families provides, therefore, information on the chromosomal distribution of genes. Using this approach, three subsets of reverse or Giemsa negative bands, H3+, H3* and H3-, containing large, moderate, and no detectable amounts, respectively, of the gene-richest H3 isochores were identified at a resolution of 400 bands. H3+ bands largely coincide with the most heat denaturation-resistant bands, the chromomycin-A3-positive, DAPI-negative bands, the bands with the highest CpG island concentrations, and the earliest replicating bands. Here, we have defined the H3+ bands at a 850-band resolution, and have thus identified the human genome regions, having an average size of 4 Mb, that are endowed with the highest gene density. PMID- 10515215 TI - The DAPI banded karyotype of the domestic dog (Canis familiaris) generated using chromosome-specific paint probes. AB - The domestic dog (Canis familiaris) is widely used as a model in the study of human disease. However, many of the 78 chromosomes comprising the canine karyotype are extremely difficult to identify reliably by classical cytogenetics. This has been a major hindrance to molecular cytogenetic studies of this species. The Animal Health Trust and the Sanger Centre have developed a set of canine whole chromosome-specific fluorescence in situ hybridisation (FISH) probes (chromosome paints). We have used these chromosome paints to identify unequivocally each chromosome in a metaphase spread. An increasing number of laboratories are making use of cooled CCD cameras and sophisticated software for FISH mapping. Consequently, there is a major trend towards the use of DAPI banding for concurrent chromosome identification during FISH analyses in a range of species. Here we present, for the first time, a complete DAPI banded karyotype of the dog in which each chromosome has been accurately placed, together with a 460-band DAPI ideogram. These data will facilitate the accurate assignment of FISH-mapped loci to all chromosomes comprising the karyotype and form the basis for an agreed standard of the dog karyotype. PMID- 10515216 TI - An alternate mechanism of glucocorticoid anti-proliferative effect: promotion of a Th2 cytokine-secreting profile. AB - Glucocorticoids (GCs) are used as immunosuppressive and anti-inflammatory agents in organ transplantation and in treating autoimmune diseases and inflammatory disorders and they exert their effects by several mechanisms, the most significant of which is inhibition of cytokine production and action. Recent reports suggested that GCs inhibit cytokine expression indirectly through promotion of a T helper cell type 2 (Th2) cytokine-secreting profile, thereby resulting in preferential blockade of pro-inflammatory monokine and T helper cell type 1 (Th1) cytokine expression. The target of GCs appeared to be monocytes macrophages, whereby altered regulation of interleukin (IL)-1/IL-1 receptor antagonist (IL-1ra), coupled with profound blockade of IL-12 synthesis and inhibition of interferon (IFN)-gamma-induced major histocompatibility complex (MHC) class II expression, lead to a preferential cognate stimulation of Th2 cells at the expense of Th1 cells. It is possible that this may have involved the expansion of a Th2-cell pool or, in addition, frank stimulation of uncommitted naive CD4 + T cells toward the Th2 lineage. In addition, GCs may have blocked Th1 cytokine expression, thereby inhibiting ongoing Th1 cytokine secretion, and consequently provided for the unimpeded production of Th2 cytokines. Collectively, this indicates that, in exerting their anti-proliferative effects, GCs act indirectly by altering Th1/Th2 cytokine balance, blocking the (pro inflammatory) Th1 program and favoring the (anti-inflammatory) Th2 program. PMID- 10515217 TI - Influence of hyperglycemia and hyperuricemia on long-term transplant survival in kidney transplant recipients. AB - Long-term prognosis in kidney transplant recipients depends on multiple factors. The purpose of this study was to quantify the influence of hyperuricemia and hyperglycemia (elements of the so-called 'syndrome X', i.e., a combination of metabolic disorders like hyperuricemia, diabetes mellitus, hyperlipidemia, and hypertension) on organ function in 350 kidney transplant recipients who had received 375 kidney transplants up to 1990 and in whom sex, age of recipient and donor, nephrologic disease, duration of dialysis, human leukocyte antigen (HLA) classification, and duration of transplant ischemia had been well matched. We found the influence of hyperuricemia on graft survival to be statistically significant (p < or = 0.05), while a statistically significant correlation between hyperglycemia and graft survival could not be detected in the present study. The transplant survival rates 2, 4, and 5 yr post-kidney-transplantation were 96.7, 80.7, and 78.7 in normogylcemic patients vs. 96.9, 85, and 82.7% in hyperglycemic ( > 100 mg,dL) kidney transplant recipients (p > 0.05). Transplant survival in hyperuricemic patients (male, > 8 mg dL; female, > 6.2 mg/dL) 2, 4, and 5 yr post-transplantation was significantly reduced (92.2, 70.6, and 68.8% vs. 98.1, 85.6, and 83.3%), as compared to normouricemic recipients. A combined presence of both hyperuricemia and hyperglycemia probably influencing the prognosis post-kidney-transplantation failed to reach the level of statistical significance. We found a significant correlation between age of recipients and plasma glucose (p < or = 0.01) and between serum uric acid concentrations and diuretic therapy (p < or = 0.05) and gender (p < or = 0.(5). In conclusion, hyperuricemia after kidney transplantation seems to reduce graft survival, whereas an influence of the carbohydrate metabolism has to be denied. PMID- 10515218 TI - Functional, life-threatening disorders and splenectomy following liver transplantation. AB - Splenectomy (SPL) in cirrhotic patients undergoing liver transplantation (LTx) may resolve specific problems related to the procedure itself, in case of functional and life-threatening clinical situations often occurring as a result of liver cirrhosis and portal hypertension. METHOD: A single-center experience of ten splenectomies in a series of 180 consecutive adult liver transplant patients over a period of 6 yr is reported. The mean patient age was 46.8 +/- 9.5 yr (range 25 57 yr). Indications for SPL were post-operative massive ascitic fluid loss (n = 3), severe thrombocytopenia (n = 3), acute intra-abdominal hemorrhage (n = 2), infarction of the spleen (n = 1), and multiple splenic artery aneurysms (n = 1). RESULTS: Extreme ascites production due to functional graft congestion disappeared post-SPL, with an improvement of the hepatic and renal functions. SPL was also effective in cases of thrombocytopenia persistence post-LTx, leading to an increase in the platelet count after about 1 wk. Bleeding episodes related to left-sided portal hypertension or trauma were also resolved. The rejection rate during hospitalization was 0%, and no other episodes were recorded in the course of the long-term follow-up. However, sepsis with a fatal outcome occurred in 4 patients, i.e. between 2 and 3 wk post-SPL in three cases and 1 yr after the procedure as a result of pneumococcal infection in the last case. Fatal traumatic cranial injury occurred 3 yr post-LTx in another case. Five patients (50%) are still alive and asymptomatic after a median follow-up period of 36 months. CONCLUSION: The lowering of the portal flow appears to resolve unexplained post operative ascitic fluid loss as a result of functional graft congestion following LTx. However, because of the enhanced risk of SPL-related sepsis, a partial splenic embolization (PSE) or a spleno-renal shunt could be used as an alternative procedure because it allows us to preserve the immunological function of the spleen. SPL is indicated in case of post-transplant bleeding due to left sided portal hypertension and trauma, spleen infarction, and to enable prevention of hemorrhage in liver transplant patients with multiple splenic artery aneurysms. Severe and persistent thrombocytopenia could be treated with PSE. Because the occurrence of fatal sepsis post-SPL is a major complication in LTx, functional disorders, such as ascites and thrombocytopenia, should be treated with a more conservative approach. PMID- 10515219 TI - Comparison of rejection rate and functional outcome of small bowel transplantation alone or in conjunction with the ileocecal valve versus combined small and large bowel transplantation. AB - Preservation of the ileocecal valve improves absorptive function and decreases the amount of small bowel needed for survival in patients with short gut syndrome. We compared the results of small and large bowel transplant (SLBTx), small bowel transplant only (SBTx), and SBTx with the ileocecal valve (ICVTx) in a porcine model. Total enterectomy was performed on 18 Yorkshire-Landrace pigs followed by orthotopic SBLTx (n = 6), SBTx (n = 6), and ICVTx (n = 6). A jejunostomy and an ileostomy were constructed for biopsies. Overall mean survival was 17 d with no statistically significant difference between groups. Rejection was seen in 6/6 SLBTx, 4/6 SBTx, and 4/6 ICVTx recipients. Acute rejection was seen in 84.3% of SLBTx, 52.3% of SBTx, and 42.5% of the ICVTx mucosal biopsy samples. Two cases of intra-abdominal infection were in the ICVTx group only. Weight loss was 147 g/d in the SLBTx group, 643 g/d in the SBTx group, and 393 g/d in the ICVTx group. While the functional outcome after SLBTx and ICVTx was noticeably better than the SBTx group, the increased rejection and intra abdominal infection rates make transplanting the large bowel or the ileocecal valve a less attractive clinical option. PMID- 10515220 TI - Immunoreactive trypsinogen levels in pediatric patients with intestinal failure awaiting intestinal transplantation. AB - The aim of this study was to evaluate pancreatic function in total parenteral nutrition (TPN)-dependent children with permanent intestinal failure by measuring immunoreactive trypsinogen (IRT) levels. Between 1992 and 1996, 105 pediatric patients with permanent intestinal failure were referred to the Children's Hospital of Pittsburgh for small intestinal transplant evaluation. Serum samples were available from 55 of them. Ten suffered from intestinal pseudo-obstruction or microvillus inclusion disease, while 45 had short bowel syndrome (SBS). IRT levels were significantly higher (p < 0.001) in SBS patients (89.4 +/- 9.2 ng mL) compared to controls (43.4 +/- 5.6 ng/ nL) without liver, gastrointestinal, or kidney disease. IRT levels did not correlate with liver injury, length of bowel, or the cause of SBS. Five of 20 patients who underwent intestinal transplantation developed pancreatitis during a median post-operative follow up 15.4 months later. IRT levels failed to predict who would develop pancreatitis post transplant. The data suggest that elevated plasma IRT levels are common among children with intestinal failure, but fail to identify patients at risk for pancreatitis post-transplant. PMID- 10515221 TI - Differences in Type 1 and Type 2 intracytoplasmic cytokines, detected by flow cytometry, according to immunosuppression (cyclosporine A vs. tacrolimus) in stable renal allograft recipients. AB - Recent multicenter, randomized clinical trials have shown that in renal transplant patients tacrolimus (FK506) was more efficient than cyclosporine A (CsA) at preventing acute rejection. In order to try and evaluate whether this difference was related to a different in vivo T-cell suppression we assessed, in a prospective study, the frequencies of interleukin (IL)-2-, IL-4-, IL-5-, IL-6-, IL-10-, interferon-gamma (IFN-gamma)- and double-positive IL-2/IFN-gamma producing whole T cells, CD4 + and CD8 + T-cell subsets by means of cytokine flow cytometry. This was performed after in vitro stimulation of peripheral blood mononuclear cells (PBMCs) with phorbol myristate acetate (PMA) and ionomycin, in the presence of monensin, in 14 healthy volunteers (controls) and in 14 renal transplant patients. The immunosuppression of the latter was based either on CsA (n = 7) or on FK506 (n = 7). Cytokine-expressing T-cell frequencies were assessed immediately pretransplantation (DO), and subsequently 3 months (M3) and 6 months (M6) afterwards in fasting patients prior to the morning intake of the immunosuppressive drug. We found that at DO the frequencies of IL-2-(22 +/- 2% vs. 22.2 +/- 2%), IFN-gamma-(26 +/- 3% vs. 29 + 3.4%) and IL-4-(0.8 +/- 0.2% vs. 1.4 +/- 0.2%)-expressing T lymphocytes were not significantly different between the controls and the patients, respectively. Conversely, the frequency of IL 2/IFN-gamma double positive cells was higher in the latter (9.3 +/- 1.6%) than in the controls (5.6 +/- 0.8); p = 0.06. Finally, on D0 the frequencies of IL-5-, IL 6-, and IL-10-producing T lymphocytes were lower than 1%, in both groups, as well as after grafting, i.e. on M3 and M6. As compared to baseline (DO): (a) chronic immunosuppression significantly decreased the frequencies of IL-2-, IL-4- and IL 2/IFN-gamma-expressing T cells, whereas those of IFN-gamma, IL-5, IL-6, and IL-10 were not significantly affected; (b) the frequencies of cytokine-expressing T cells were not statistically different between M3 and M6; (c) the decrease in the frequencies of IL-2- and IL-2/IFN-gamma-expressing T cells affected CD4 + and CD8 + cells equally; (d) there was a marginal decrease in the frequency of IFN-gamma expressing cells only in the CD4 + subset but not in the CD8 population; and (e) for CsA, but not for FK506, the frequency of the IL-2-expressing T cells was negatively correlated with the whole blood trough levels. When we compared the frequencies of cytokine-expressing cells in FK506- and CsA-treated patients, we found that the frequency of IL-2-expressing T cells was significantly lower with FK506 (10.9+/-1.61%) than with CsA (16.3 +/- 1.8%; p = 0.03), whereas the frequencies of the other cytokine-expressing cells were not statistically different between the two groups. In conclusion, our study clearly demonstrated that studied ex vivo, FK506 and CsA decrease the frequencies of cells expressing IL-2, IL-4 and IL-2/IFN-gamma in vivo but do not affect those expressing IFN gamma. Meanwhile, the frequency of IL-2-producing T cells was more affected with FK506 than with CsA and was negatively correlated with the CsA trough level. Finally, our results regarding IL-2 might explain to some extent the higher efficiency of FK506 in vivo than CsA. PMID- 10515222 TI - Exercise blod flow and microvascualr distensibility in skeletal muscle normalize after heart transplantation. AB - This study investigated the effect of heart transplantation (HTX) on reduced exercise blood flow and microvascular stiffness in patients with congestive heart failure (CHF). Skeletal muscle blood flow (SMBF) during graded maximal supine bicycle exercise and microvascular distensibility (i.e., stiffness) were measured in musculus tibialis anterior by the isotope washout method. Measurements were performed in a cross-sectional study with 31 CHF patients and 28 patients, mean 9 months after HTX, and in a longitudinal study in 12 CHF patients before, 3 months, and 14 months after HTX, and in 31 healthy controls. Maximal SMBF: In the cross-sectional study, maximal SMBF was reduced in severe CHF patients (3.6 +/- 2.5 mL (100 g min)(-1)) and increased after HTX (7.7 +/- 4.8; p < 0.01 versus controls (11 +/- 4.1). Maximal SMBF was reduced in CHF patients (5.8 +/- 4.0) and reversed to normalization 3 months after HTX (10.3 +/- 4.4) in the longitudinal study. Microvascular distensibility: The distensibility was reduced (severe CHF, 12 +/- 8%; moderate CHF, 23 +/- 14%) in the cross-sectional study and increased after HTX towards normalization (38 +/-20%; controls: 44 +/- 17). In the longitudinal study, distensibility in CHF patients (14 +/- 6%) increased gradually to 32 +/- 12% (p < 0.005) at 3 months and normalized 14 months after HTX (46 +/- 17%). HTX gradually reversed the reduced SMBF and microvascular distensibility in CHF patients towards normalization. PMID- 10515224 TI - Carvedilol improves functional class in patients with severe left ventricular dysfunction referred for heart transplantation. AB - PURPOSE: Limited options are available to improve the functional class of patients awaiting cardiac transplantation. We assessed the effect of carvedilol on New York Heart Association (NYHA) class, heart rate (HR), blood pressure (BP), jugular venous pressure (JVP), electrolytes and renal function in patients with markedly decreased left ventricular (LV) function referred for cardiac transplantation assessment. METHODS: Sixty-one patients (age = 52 +/- 12 yr, EF = 23 +/- 7%, VO2 max = 16 +/- 5 mL/kg/min) referred to the cardiac transplant clinic were reviewed before and after the addition of carvedilol (starting dose 3.125 mg twice daily to target dose of 25 mg twice daily) to usual heart-failure therapy. Over a 1-yr period, at each visit prior to initiation, at baseline initiation visit and at each follow-up visit, NYHA class, BP, HR, JVP, electrolytes, and renal function were obtained. Statistical analysis was performed using random effects regression approach. A multiple logistic regression analysis was performed on 52/61 patients to determine possible associations between NYHA improvement and the following patient characteristics: sex, etiology of cardiomyopathy, initial NYHA class, and dose of carvedilol. RESULTS: Three patients died (2 after stopping carvedilol) and 3 were transplanted. Median follow-up was 100 d. Sixteen patients reached the target dose after a mean of 137 d (2.75 visits). Estimated time-to-target dose is 8 months (5.6 visits). BP tended to increase (p = 0.07 for change in trend) with no change in JVP, electrolytes or renal function. HR decreased 6 +/- 3 b.p.m. (p = 0.03). Of 14 patients NYHA class I/II at baseline, none were class III/IV at last follow-up visit. Of 47 patients NYHA class III/IV at baseline, 25 were class I/II, and 22 were class III/IV at last follow-up (p < 0.001). Multiple logistic regression analysis did not demonstrate any factor predictive of subsequent NYHA class improvement. CONCLUSIONS: Despite less than target doses in most patients, a favorable effect of carvedilol on functional class in patients with severe congestive heart failure (CHF) referred for transplant was observed. Those with good functional status remained stable and those with poor functional status improved or remained stable. No baseline patient characteristic predicted improvement. The shortage of donor organ requires optimal use of medical regimens which may improve functional class while awaiting transplantation and which may delay the necessity for heart transplantation. Therefore, addition of carvedilol to usual medical therapy may be beneficial even in patients with severe LV dysfunction and poor NYHA classification. PMID- 10515223 TI - Cold preservation of the human colon and ileum with University of Wisconsin solution. AB - The inclusion of the colon in the intestinal graft resulted in worsening patient and graft outcome and increased the incidence of infection and rejection. In this study, we examine the role of ischemia on the barrier function of the epithelium during cold ischemia. Samples were collected from 15 harvested and transplanted human donor grafts (colon, 10; ileum, 6), which were immersed in University of Wisconsin (UW) solution. Ischemia (6, 12, 24, and 48 h) and reoxygenation were performed to evaluate the mucosal electrical status using the Ussing chamber technique. The functions of enterocytes and crypt cells were tested by glucose and theophylline challenge. Modified Park's classification was applied to evaluate the severity of mucosal damage under light microscopy. The colon had higher levels of baseline potential difference, short-circuit current, and resistance than the ileum during 6 48 h of ischemia. Colonic epithelial cells responded well to theophylline stimulation at 24 h of ischemia, while there was no ileal response. The colonic mucosa was histopathologically well preserved in UW solution for 48 h, and mucosal damage induced by reoxygenation was less than in the ileum. In conclusion, electrophysiologically and histopathologically, the colon is less susceptible to cold preservation damage than the ileum during storage with UW solution. PMID- 10515225 TI - Adhesion molecules in patients after lung transplantation. AB - Leukocyte adhesion molecules, such as intercellular adhesion molecule (ICAM)-1 and its ligands, are involved in inflammatory processes of the lung. For ICAM-1, differential expression during different kinds of complications after transplantation has been proposed. We analyzed the role of ICAM-1, CD18, CD11a, CD11b, and CD11c during episodes of rejection or infection in patients after lung transplantation and compared the results to episodes without apparent complication. A total of 98 bronchoalveolar lavage (BAL) samples and 90 serum samples were analyzed. ICAM-1, CD18, CD11a, CD11b, and CD11c expressions were detected immunocytochemically on alveolar macrophages. Soluble ICAM-1 was quantified in serum and BAL. In the control group, 49.8 +/- 18% of macrophages stained positive for CD11b. During rejection, the mean of cells showing CD11b on the surface was significantly higher (64.6 +/-11.4%) with no difference compared to episodes of infection (59.7 +/-22.7). All other epitopes were not expressed differently with regard to a normal clinical course or episodes of infection and rejections. In summary, assessment of ICAM-1 and corresponding ligands did not allow for a reliable discrimination between episodes of rejection or infection in lung transplantation. PMID- 10515226 TI - The extracellular matrix of gliomas: modulation of cell function. AB - The ECM of astrocytic tumors promotes and modulates a variety of cell functions, such as cell attachment, migration, proliferation, survival, and signaling. Recent studies indicate that there are extensive and complex interactions among ECM molecules and that these can modify the function of the participating molecules, interactions between the proteoglycan, phosphacan, and the ECM protein, tenascin, being an example (63). In addition, on nonastrocytic cell types it has been shown that an integrin receptor and the cell surface proteoglycan CD44 recognize the same ECM ligand osteopontin, and thus modulate each others function (77, 86). Thus, interacting components in the ECM and cell surface receptors likely cooperate in regulating cell function and tumor invasion (59, 77, 80, 85-87, 95). As tumor cells are capable of remodeling their ECM through synthesis of ECM proteins and proteoglycans, as well as upregulating integrin receptors and proteoglycans on their cell surface, tumor cells are capable of controlling their own destiny. ECM molecules which are concentrated in blood vessels of malignant astrocytomas, such as tenascin-C and the 250-kDa CSPG (NG2), are potentially therapeutic targets. PMID- 10515227 TI - Identification of two distinct deleted regions on the short arm of chromosome 1 and rare mutation of the CDKN2C gene from 1p32 in oligodendroglial tumors. AB - Oligodendroglial tumors frequently show allelic losses on the short arm of chromosome 1. To narrow down the putative tumor suppressor gene site(s) on 1p, we have investigated 35 oligodendrogliomas and 10 mixed gliomas (oligoastrocytomas) for loss of heterozygosity (LOH) at 21 highly polymorphic loci on chromosome 1 (19 loci on 1p and 2 loci on 1q). LOH at loci on 1p was found in 30 of the 45 tumors (67%). Two distinct regions of common allelic loss were identified: a distal region between D1S76 and D1S253 at 1p36.3, and a proximal region between D1S482 and D1S2743 at 1p34-p35. We also analyzed our tumor series for genetic alterations and expression of the cyclin dependent kinase inhibitor gene CDKN2C (p18INK4c) from 1p32. We found 1 recurrent anaplastic oligodendroglioma that carried a somatic CDKN2C mutation at codon 113 (GAA ==> TAA: Glu ==> Stop). The remaining 44 tumors of our series showed neither coding sequence mutations nor homozygous deletions of CDKN2C. Investigation of 35 tumors by differential reverse transcription-PCR revealed expression of CDKN2C transcripts in all instances. Our data thus provide evidence for more than a single oligodendroglioma-associated tumor suppressor gene on 1p and implicate CDKN2C as a candidate tumor suppressor gene altered in a low fraction of oligodendroglial tumors. PMID- 10515228 TI - Upregulation of vascular endothelial growth factor is associated with radiation induced blood-spinal cord barrier breakdown. AB - The pathogenesis of radiation-induced injury to the central nervous system (CNS) remains unclear. Dysfunction of the blood-brain barrier (BBB) is associated with radiation-induced white matter lesions. The aim of this study was to determine if vascular endothelial growth factor (VEGF) is implicated in radiation-induced BBB disruption. Adult rats were irradiated with a single dose of 8 or 22 Gy to the spinal cord from C2 to T2. At various times up to 20 weeks following irradiation, blood-spinal cord barrier (BSCB) permeability was assessed using immunohistochemistry with anti-albumin antibody. Cell proliferation was assessed using bromodeoxyuridine (BrdU), and endothelial cell identity was assessed morphologically and using immunostaining for factor VIII-related antigen. Expression of VEGF protein and message was assessed using immunohistochemistry and in situ hybridization respectively. In the unirradiated rat spinal cord, there was no evidence of albumin immunoreactivity and little evidence of VEGF expression. After a dose of 22 Gy, focal albumin staining in white matter was observed at 16 weeks. Diffuse staining was seen at 20 weeks and was associated with necrosis and demyelination in white matter. This was associated with a significant increase in white matter glial cells that showed immunoreactivity and in situ hybridization signal for VEGE VEGF expressing cells showed dual immunoreactivity for glial fibrillary acidic protein. No increase in VEGF positive cells was observed in gray matter after 22 Gy. After a dose of 8 Gy, there was no increase in VEGF expression or albumin immunostaining in either white or gray matter. Microvessel endothelial cell density showed a trend towards a decrease with time after 22 Gy as compared with 8 Gy or unirradiated controls. BrdU immunostaining provided no evidence for endothelial cell proliferation in control or in the irradiated spinal cord. It is concluded that radiation-induced BSCB dysfunction is associated with upregulation of VEGF in astrocytes without associated endothelial proliferation. The temporal and spatial association of VEGF upregulation with the white matter lesions suggests a role of VEGF in radiation-induced late CNS injury. PMID- 10515229 TI - Pediatric astrocytomas with monomorphous pilomyxoid features and a less favorable outcome. AB - Among tumors classified as pilocytic astrocytoma (PA) in the Johns Hopkins Hospital Department of Pathology files, we identified 18 cases with a distinctive monomorphous pilomyxoid histological pattern and a higher recurrence rate than that of PA with classical histological features (classical PA). The majority of the tumors occurred in infants and young children and involved the hypothalamic/chiasmatic region. The tumors were histologically similar to PA, but they were more monomorphous and more myxoid. Rosenthal fibers were not seen and only 1 of 18 tumors had eosinophilic granular bodies. At the end of the follow-up period, 6 patients were dead and 12 were alive with evidence of disease. Progression free survival (PFS) at 1 year was 38.7%. In comparison, we identified a control group of 13 classical PAs in the same age range and location as the study group. In this group, PFS at 1 year was 69.2%, which was significantly better than that for pilomyxoid tumors (p = 0.04). There was no CSF dissemination or death due to tumor progression among patients with classical PA. Eight of these patients are alive with recurrent disease, and 4 have no evidence of disease. While the monomorphous pilomyxoid tumors have some resemblance to classical PA, our results suggest that the former is a more aggressive variant or a separate entity that needs to be recognized for prognostic purposes. PMID- 10515230 TI - Primary brain lymphoma cell turnover differs in patients with and without AIDS: relationships to bcl-2 expression and host cell reaction. AB - Primary central nervous system lymphomas (PCNSLs) are more resistant to radiotherapy and chemotherapy in AIDS (A-PCNSLs) than in non-AIDS patients (NA PCNSLs). We investigated 23 A-PCNSLs and 24 NA-PCNSLs. Lymphoma cell kinetics (i.e. proliferation [mitotic index, MIB-1 and PCNA labeling indices], apoptosis and turnover) were determined and compared with bcl-2 and LMP-1 expression, and to the percentage of tumor-infiltrating T-lymphocytes (T-TILs) and macrophages. A PCNSLs showed lower proliferation (p < 0.005), less apoptosis (p < 0.0001) and slower cell-turnover (p < 0.0001) than NA-PCNSLs. LMP-1 was detected in 90% of A PCNSLs and 5% of NA-PCNSLs, a finding correlating positively with bcl-2 expression (p < 0.0007). In contrast, T-TIL counts and CD4/CD8 T-TIL ratios were similar in A-PCNSLs and NA-PCNSLs. T-TIL counts correlated negatively with proliferation indices (from p < 0.05 to p < 0.0005) in NA-PCNSLs, but not in A PCNSLs. Macrophage counts correlated positively with apoptosis in both groups. We concluded the following: (i) A-PCNSLs are characterized by accumulation of slow cycling, long-lived cells that might be protected from apoptosis by LMP-1 induced bcl-2 expression, and independently from the host response; (ii) NA-PCNSLs are characterized by a faster cell turnover associated with an insufficient antiproliferative host response; and (iii) A-PCNSLs and NA-PCNSLs constitute 2 entities with distinctive morphology and different kinetic profiles that could account for different responses to therapy. PMID- 10515231 TI - Microglial activation by components of gram-positive and -negative bacteria: distinct and common routes to the induction of ion channels and cytokines. AB - Gram-positive Streptococcus pneumoniae is the major pathogen causing lethal meningitis in adults. We used pneumococcal cell walls (PCW) to investigate microglial consequences of a bacterial challenge and to determine the role of serum in the activation process. PCW caused the characteristic induction of an outwardly rectifying K+ channel (IK+(OR)), together with a concomitant suppression of the constitutively expressed inward rectifier K+ current, and evoked the release of tumor necrosis factor-alpha (TNF alpha), interleukin-6 (IL 6), IL-12, KC, macrophage inflammatory protein (MIP) 1alpha and MIP-2. Serum presence strongly facilitated the PCW effects, similarly as observed for lipopolysaccharide (LPS) from gram-negative Escherichia coli. The inflammatory cytokine, interferon-gamma (IFNgamma) induced the same electrophysiological changes, but independent of serum. Recombinant LPS binding protein (LBP) could partially replace serum activity in LPS stimulations. In contrast, neither LBP nor an antibody-mediated blockade of the LPS receptor, CD14 had significant influences on PCW-inducible changes. Cell surface interactions and cofactor involvement in microglial activation by gram-positive bacteria are thus distinct from the mechanisms employed by LPS. Moreover, tyrphostin AG126, a protein kinase inhibitor that prevents activation of the mitogen-activated protein kinase, p42MAPK (ERK2), potently blocked the PCW-stimulated cytokine release while having only a limited effect on LPS-inducible cytokines. In contrast, AG126 did not influence IK+(OR) inductions. This indicates that PCW recruits more than 1 intracellular signaling pathway to trigger the various responses and that different bacterial agents signal through both common and individual routes during microglial activation. PMID- 10515232 TI - Demonstration by fluorescence resonance energy transfer of a close association between activated MAP kinase and neurofibrillary tangles: implications for MAP kinase activation in Alzheimer disease. AB - Indirect evidence suggests that activation of the mitogen activated protein kinase (MAPK) cascade contributes to the hyperphosphorylation of tau found in paired helical filaments in Alzheimer disease (AD). We report colocalization of the activated form of MAPK with Ser 199/202 and Ser 396/404 phosphotau immunoreactive neurofibrillary tangles and neuritic plaques in the Alzheimer brain. Fluorescence resonance energy transfer studies (FRET) demonstrate a tight intermolecular association of activated MAPK with these phosphotau epitopes. These data support the hypothesis that activation of MAPK contributes directly to phosphorylation of tau in AD. Moreover, the stable nature of this association in postmortem human brain may suggest a stable interaction in which activated MAPK becomes tightly linked to neurofibrillary tangles. PMID- 10515233 TI - Extra neurofilament NF-L subunits rescue motor neuron disease caused by overexpression of the human NF-H gene in mice. AB - Previous studies demonstrated that transgenic mice overexpressing human neurofilament heavy (hNF-H) protein develop a progressive motor neuron disease characterized by the perikaryal accumulations of neurofilaments resembling those found in amyotrophic lateral sclerosis (ALS). To further investigate this neurofilament-induced pathology, we generated transgenic mice expressing, solely or concomitantly, the hNF-H and the human neurofilament light (hNF-L) proteins. We report here that the motor neuron disease caused by excess hNF-H proteins can be rescued by overexpression of hNF-L in a dosage-dependent fashion. In hNF-H transgenic mice, the additional hNF-L led to reduction of perikaryal swellings, relief of axonal transport defect and restoration of axonal radial growth. A gene delivery approach based on recombinant adenoviruses bearing the hNF-L gene also demonstrated the possibility to reduce perikaryal swellings after their formation in adult mice. The finding that extra NF-L can protect against NF-H-mediated pathogenesis is of potential importance for ALS, particularly for cases with NF-H abnormalities. PMID- 10515234 TI - Synaptophysin in choroid plexus epithelial cells: no useful aid in differential diagnosis. PMID- 10515235 TI - Cutaneous signs of selected systemic diseases. AB - In this work we will discuss three cutaneous markers of systemic disease. Acanthosis nigricans is the most widely known in this category. Cutaneous angiokeratomas are usually associated with enzyme defect disorders, such as Fabry's disease, and are linked in this work as a reactive pattern in hepatorenal failure. Mast cell disease may occur in a number of forms, often with systemic involvement. PMID- 10515236 TI - Unusual premature ovarian failure with hypogonadotropic hyperprolactinemia and 46, XX, 13ph+. AB - A 34-year-old woman with secondary amenorrhea is reported. She had a 30-day regular cycle menstruation from 12 to 24 years old. Her hormonal examination showed hypogonadotropic, secondary ovarian functional defect, and hyperprolactinemia. Her chromosomal arrangement was 46, XX, 13ph+ which is the elongation of 13p. PMID- 10515237 TI - Serum thrombomodulin as a prognostic marker of disseminated intravascular coagulation. AB - Disseminated intra-vascular coagulation (DIC) is associated with severe bleeding tendency and organ failure, the extent of which is thought to be related to the prognosis of DIC patients. Thrombomodulin (TM) is a high-affinity thrombin receptor on vascular endothelial cells. Clinical importance of soluble TM is still controversy as a diagnostic and prognostic indicator in patients with disseminated intravascular coagulation (DIC). We compared plasma levels of TM with fibrin degradation product (FDP) in patients with DIC through the clinical course. The significant elevation of circulating TM in nonsurvivors with DIC compared with survived patients with DIC(TM 3.1+/-1.52 vs 8.1+/-3.89 FU/ml), as well as FDP (12.9+/-12.12 vs 49.8+/-55.42 microg/ml) but the levels of FDP were not different between the two groups. The measurement of circulating TM was a relatively good prognostic marker of patients with DIC. PMID- 10515238 TI - Effects of physical therapy on cytokines and two color analysis-lymphocyte subsets in patients with cerebrovascular diseases. AB - The effects of physical therapy on immunological parameters were evaluated in 12 patients (8 males and 4 females, 69.2 +/- 9.0 years) with cerebrovascular diseases in a stable situation two to three months after the onset of stroke who entered in our hospital between 1994 and 1997. After a two-month physical therapy program, the proportions of helper-inducer T (Thi) cells and suppressor-inducer T (Tsi) cells were increased significantly and that of cytotoxic T (Tc) cells was decreased, although those of HLA-DR+, suppressor T (Ts) and activated T (Tac) cells were not changed. The antibody dependent cellular cytotoxicity (ADCC) was significantly increased, although natural killer (NK) cell activity was not changed. The serum levels of interleukin-2 receptor was significantly increased but those of interleukin-2, interleukin-6 and interleukin-12 were not changed. The serum levels of interleukin-10, interleukin-12 and tumor necrosis factor alpha were not detectable, while interleukin-1beta was decreased in 2 patients and interleukin-10 was increased in 2 patients. These findings suggest that daily physical exercise may activate the immune system possibly through the cytokine network in patients with cerebrovascular diseases (CVD). PMID- 10515239 TI - Hypercalcemia-induced renal insufficiency during therapy with dihydrotachysterol. AB - During vitamin-D therapy drug accumulation and intoxication should be considered. In the present study we report on five patients with renal insufficiency during therapy with dihydrotachysterol or calcitriol. Four patients received dihydrotachysterol for 29 (7-44) years and one patient received calcitriol for 4 years to treat hypoparathyroidism after thyroid surgery. As confirmed by renal biopsy impairment of renal function was due to calcifications as a consequence of prolonged hypercalcemia. The effective duration of dihydrotachysterol is ten days as compared with five days for calcitriol. Severe hypercalcemic episodes with dihydrotachysterol are longer-lasting than those with the shorter acting vitamin D derivatives. Further, they occur with higher incidence as was shown by our own observations and previously published data by other workers. Hence, impairment of renal function during therapy with dihydrotachysterol should be considered as being due to hypercalcemia and hypercalciuria. PMID- 10515240 TI - Serum and synovial fluid concentrations of endothelin-1 in patients with rheumatoid arthritis. AB - To determine the endothelin-1 (ET-1) concentrations in synovial fluid and serum of rheumatoid arthritis (RA) patients, this study was designed to examine if serum ET-1 concentration of control subjects has any correlation either with the ET-1 concentration of synovial fluid or ET-1 concentration of serum from RA patients. Twenty-eight patients were studied of whom eight males and twenty females with confirmed rheumatoid arthritis. Twenty-eight healthy volunteers were also included as controls. The immunoreactive concentration of ET-1 was measured using commercially available radioimmunoassay (RIA) kits (Peninsula Laboratories, Belmont CA) specific for ET-1. All the samples were performed in duplicate and after plotting % B/Bo for each standard directly on Y axis and endothelin concentrations on the X axis, the "best fit" curve was drawn and the amount of ET 1 was calculated. Mean ET-1 level in synovial fluid was 15.53 +/- 2.82 pg/me. In serum samples from RA patients, the mean ET-1 level was detected as 16.42 +/- 3.07 pg/ml (n = 28). Sera from twenty-eight healthy volunteers were analyzed as controls and mean ET-1 concentration was 8.68 +/- 1.96 pg/ml. A significant difference (P < 0.001) was found between ET-1 level of sera from RA patients and ET-1 levels from control sera. Highly significant difference (P < 0.001) was also detected between synovial fluid ET-1 and control ET-1 levels. However, no significant difference was found between ET-1 levels of synovial fluid and serum ET-1 levels of RA patients. Results of this study confirmed the presence of elevated levels of ET-1 concentration in synovial fluid and serum samples of patients with RA. The clinical significance and physiological role of endothelin in synovial fluids and sera of patients suffering from a variety of pathophysiological conditions of arthritis deserves further studies. PMID- 10515241 TI - Clinical analysis of breathing exercise during immersion in 38 degrees C water for obstructive and constrictive pulmonary diseases. AB - Breathing exercises during immersion in 38 degrees C water were performed in 22 patients with bronchial asthma, pulmonary emphysema and constrictive pulmonary diseases. The patients entered a pool filled with 38 degrees C water to shoulder level. While standing, they breathed in deeply and breathed out slowly through the mouth into water while sinking the nose below the water level. This breathing method was repeated for 20 min. with a 5-min. rest out of water and this cycle was performed twice a day for two months. Respiratory function test and arterial blood gas analysis were examined before and after the two-month exercise program. FEV1.0% was significantly increased in patients with asthma and emphysema (p = 0.042 and 0.032, respectively) but did not change in patients with constrictive pulmonary diseases. %FVC, PF and Vmf25 did not change in any of the diseases. PaO2 was significantly increased in emphysematous patients (p = 0.0002) and PaCO2 was significantly decreased in asthmatic and emphysematous patients (p = 0.034 and 0.046, respectively). These results suggest that our breathing exercise by immersion is useful in patients with asthma and emphysema but is less effective in patients with constrictive pulmonary diseases. PMID- 10515242 TI - Steroid therapy is effective in a young patient with an inflammatory abdominal aortic aneurysm. AB - We report a successful resection of an inflammatory aneurysm following treatment with steroids in a 23-year-old man. Suffering from fever and severe lumbago, he was admitted to our hospital. An ultrasound and computed tomography of the abdomen revealed an infrarenal abdominal aortic aneurysm surrounded by dense perianeurysmal fibrous tissue. We diagnosed it as an inflammatory abdominal aortic aneurysm based on a symptomatic inflammatory reaction and the findings of ultrasound and computed tomography. Since the aneurysmal wall strongly adhered to the surrounding tissues and surgery was ruled out when it proved impossible to expose the vessels sufficiently to obtain vascular control, steroid therapy was started to control fever and severe lumbago. Five months later, we undertook surgery. Our conclusion is that steroid therapy was very effective against a young patient with inflammatory abdominal aortic aneurysm. PMID- 10515243 TI - Comparative accuracy of automated computer analysis versus physicans in training in the interpretation of electrocardiograms. AB - We compared the interpretation of an electrocardiogram (ECG) made by computer with that made by physicians in training, as well as with the diagnosis made by cardiologists. ECGs were collected from 1058 Japanese adults (812 men and 246 women, mean age 49 +/- 19 years). With the computer program, the incidence of false-negative reports was 10.5% while that of false-positive reports was 16.5%, when compared with the physician's diagnosis. The incidence of a false-positive diagnosis with the computer was 18 times higher than that found by the physicians in training. The results show the advantages and the limitations in the use of computers for analysis of ECGs. PMID- 10515244 TI - A combination of autoimmune hepatitis, sensory-dominant peripheral neuropathy, and primary Sjogren's syndrome in the same patient: a rare association. AB - Although autoimmune hepatitis and sensory-dominant neuropathy have been known to independently accompany primary Sjogren's syndrome, the combination of all these conditions in the same patient has not been described. We report the case of a woman who initially suffered from autoimmune hepatitis and later was diagnosed with primary Sjogren's syndrome upon the development of sensory-dominant neuropathy. In this patient, autoimmune hepatitis preceded neuropathy by one year. All of the diagnoses were confirmed by histological examinations of the liver, sural nerve, and minor salivary gland. Her autoimmune hepatitis was relieved with conservative treatment, and her sensory-dominant neuropathy was alleviated by prednisolone therapy. Our case indicated that the multiple organ involvement, especially that in the liver and peripheral nerves, should be taken into account in the course of primary Sjogren's syndrome. PMID- 10515245 TI - Pulmonary nodule mimicking lung cancer in a human immunodeficiency virus type-1 infected patient. AB - We reported a human immunodeficiency virus type 1-infected patient with a small solitary pulmonary nodule mimicking adenocarcinoma, who was treated successfully with antituberculosis therapy. We believe that high-resolution CT scans of thorax are important examinations to detect pulmonary inflammatory findings, such as ectasis of the bronchi leading to the nodules and calcifications in the nodules, and also as follow-up tests for evaluating effectiveness of treatment on pulmonary inflammatory nodules in human immunodeficiency virus type 1-infected patients. PMID- 10515246 TI - Successful low-dose cytosine arabinoside treatment in a patient with acute leukemia from idiopathic myelofibrosis. AB - We report a 63-year-old male with acute leukemia developed from idiopathic myelofibrosis. Cytogenetic analysis revealed chromosomal abnormalities; 46,XY,t(3;21)(q21;q22) del(20q). Intensive chemotherapies [combination of adriamycine/vincristine/prednisolone, or idarubicine/cytosine arabinoside (Ara C)] were unsuccessful. The patient was then treated with continuous intravenous low-dose Ara-C because of his poor physical condition and showed marked hematological improvement. Leukemic cells disappeared from the peripheral blood and the segmented neutrophil count recovered. The dysplastic morphology observed in the segmented neutrophils suggests that the recovery of the neutrophils has resulted mainly from the differentiation activity of Ara-C. PMID- 10515248 TI - Aortic reconstruction in the HIV infected patient. AB - Major vascular reconstruction using prosthetic implantation is uncommon in the HIV infected patient. We describe a 52-year-old man with HIV infection and HIV induced thrombocytopenia, who underwent successful aorto-bifemoral bypass for lower extremity ischemic symptoms. Postoperative progressive thrombocytopenia and subsequent hemorrhage were successfully treated with intravenous steroid replacement, platelet transfusion, alpha-globulin administration and expectant management. This report serves to illustrate that major vascular reconstruction can be successfully accomplished in the HIV infected population, even in the presence of significant hematologic dysfunction. PMID- 10515247 TI - Circulating levels of soluble CD30 and other markers in colorectal cancer patients. AB - In a search for new biologic serum tumor markers with prognostic value we evaluated the soluble form of the CD30 (sCD30), a marker of cells producing T helper 2 (Th2)-type cytokines, interleukin-1 receptor antagonist (IL-1ra), soluble interleukin-2 receptor (sIL-2R), soluble tumor necrosis factor -type I, type II (sTNF-R55, -75) and immunosuppressive acidic protein (IAP) in patients, with advanced colorectal cancer. The data showed that abnormal levels of sCD30 were detected in eight (80.0%) out of ten patients. In contrast, sCD30 levels were not detected in healthy volunteers. The relationship between sCD30, sIL-2R and IAP were positively correlated. In contrast, sCD30 and IL-1ra were negatively correlated. These results suggested that IL-1ra may play a role, at least in part, to inhibit CD30 release, and sCD30 appears to be a new biologic serum tumor marker of possible use in the clinical setting of cancer patients. PMID- 10515249 TI - Merits of medical history. PMID- 10515250 TI - Evolution and analysis of model CPGs for walking: I. Dynamical modules. AB - Can one develop an abstract description of the dynamics of pattern generators that provides quantitative insight into their operation? We explored this question by examining the dynamics of a model central pattern generator that was created using an evolutionary algorithm. We propose an abstract description based on the concept of a dynamical module, a set of neurons that simultaneously make their transitions from one quasistable state to another while the synaptic inputs that they receive from other neurons remain essentially constant, thus temporarily reducing the dimensionality of the circuit dynamics. Using the mathematical tools of dynamical systems theory, we describe a method for identifying dynamical modules and demonstrate that this concept can be used to quantitatively characterize constraints on neural architecture, account for phase durations, and predict the effects of parameter changes. Moreover, this abstract description reveals coordinated parameter changes that leave the overall circuit dynamics essentially unchanged. In a companion article we employ this abstract description to examine the relationship between general principles and individual variability in large populations of evolved model pattern generators. PMID- 10515251 TI - Evolution and analysis of model CPGs for walking: II. General principles and individual variability. AB - Are there general principles for pattern generation? We examined this question by analyzing the operation of large populations of evolved model central pattern generators (CPGs) for walking. Three populations of model CPGs were evolved, containing three, four, or five neurons. We identified six general principles. First, locomotion performance increased with the number of interneurons. Second, the top 10 three-, four-, and five-neuron CPGs could be decomposed into dynamical modules, an abstract description developed in a companion article. Third, these dynamical modules were multistable: they could be switched between multiple stable output configurations. Fourth, the rhythmic pattern generated by a CPG could be understood as a closed chain of successive destabilizations of one dynamical module by another. A combinatorial analysis enumerated the possible dynamical modular structures. Fifth, one-dimensional modules were frequently observed and, in some cases, could be assigned specific functional roles. Finally, dynamic dynamical modules, in which the modular structure itself changed over one cycle, were frequently observed. The existence of these general principles despite significant variability in both patterns of connectivity and neural parameters was explained by degeneracy in the maps from neural parameters to neural dynamics to behavior to fitness. An analysis of the biomechanical properties of the model body was essential for relating neural activity to behavior. Our studies of evolved model circuits suggest that, in the absence of other constraints, there is no compelling reason to expect neural circuits to be functionally decomposable as the number of interneurons increase. Analyzing idealized model pattern generators may be an effective methodology for gaining insights into the operation of biological pattern generators. PMID- 10515252 TI - A comparative survey of automated parameter-search methods for compartmental neural models. AB - One of the most difficult and time-consuming aspects of building compartmental models of single neurons is assigning values to free parameters to make models match experimental data. Automated parameter-search methods potentially represent a more rapid and less labor-intensive alternative to choosing parameters manually. Here we compare the performance of four different parameter-search methods on several single-neuron models. The methods compared are conjugate gradient descent, genetic algorithms, simulated annealing, and stochastic search. Each method has been tested on five different neuronal models ranging from simple models with between 3 and 15 parameters to a realistic pyramidal cell model with 23 parameters. The results demonstrate that genetic algorithms and simulated annealing are generally the most effective methods. Simulated annealing was overwhelmingly the most effective method for simple models with small numbers of parameters, but the genetic algorithm method was equally effective for more complex models with larger numbers of parameters. The discussion considers possible explanations for these results and makes several specific recommendations for the use of parameter searches on neuronal models. PMID- 10515253 TI - Direction selectivity and spatiotemporal separability in simple cortical cells. AB - Simple cells in mammalian visual cortex are quasi-linear mechanisms whose behavior departs from true linearity in a very consistent manner. Empirical research on direction selectivity (DS) clearly illustrates these characteristics. A linear DS cell will be DS for all stimuli, whereas a linear non-DS cell will not be DS for any stimuli. However, many simple cells have opposite preferred directions for stimuli of reversed polarity, and some cells are DS for some stimuli (e.g., moving bars) but not for others (e.g., drifting gratings). Also, linear non-DS cells must have separable spatiotemporal receptive fields (RFs), and linear DS cells must have inseparable RFs. Yet many actual DS cells have separable RFs. Here we present a nonlinear model of simple-cell behavior that reproduces all of these empirical behaviors. The model is a variant of the current linear model, amended to include an interleaved nonlinearity (half-wave rectification) that allows it to mimic the (im)balance of push-pull mechanisms. We present simulation results showing that balanced push-pull mechanisms result in linear behavior, while imbalanced push-pull arrangements produce all of the incongruent DS-related behaviors that have been reported for simple cells. PMID- 10515254 TI - Deprived of habitual running, rats downregulate BDNF and TrkB messages in the brain. AB - To study possible effects of physical training on the expression of neurotrophic factors and their receptors in the brain, we used a rat strain (spontaneously hypertensive rat, SHR), known to spontaneously run up to 20 km/night. We show that such long-distance running affects the brain-derived neurotrophic factor (BDNF) and TrkB system in hippocampus, and in particular that abrupt deprivation of habitual running leads to long-lasting decreases of BDNF/TrkB expression in hippocampus. Quantitative in situ hybridization demonstrates that running increases the expression of mRNA coding for BDNF and its high affinity receptor TrkB in hippocampus in a running length dependent manner. In addition, we show that an abrupt interruption of prolonged spontaneous exercise decrease expression of mRNA encoding BDNF and TrkB in certain hippocampal areas and that this decrease lasts at least 10 days. This down-regulation was most prominent in medial cornu ammonis 3 (CA3M). Several other trophic factors and receptors were investigated, including NGF, NT3, GDNF, trkC and p75. For these other probes investigated, no robust changes in mRNA expression were noted. Areas examined included sensorimotor cortex and hippocampus. For RET, p75, NT3, TrkB and BDNF we also examined the spinal cord without detecting any robust changes. We conclude that spontaneous running as well as its abrupt termination, leads to area specific and trophic factor-specific changes in hippocampus. PMID- 10515255 TI - Recovery by NO of the iron-attenuated dopamine dynamics in nigrostriatal system of rat brain. AB - In vivo electrochemical detection was employed to study the chronic effect of nitric oxide on iron-induced alterations in dopamine dynamics, including K+ evoked dopamine overflows and the clearance of exogenous dopamine in rat striatum. Intranigral infusion of fresh S-nitroso-N-acetylpenicillamine (SNAP), a nitric oxide donor, did not alter either dopamine dynamics in the striatum or the lipid peroxidation in substantia nigra 7 days after the infusion, indicating that nitric oxide is not neurodestructive. By contrast, infusion of iron in substantia nigra chronically degenerated the nigrostriatal dopaminergic system. Co-infusion of iron and fresh SNAP, but not aged SNAP, prevented the iron-induced reductions in K+ -evoked dopamine overflows. Furthermore, the clearance of exogenous dopamine was attenuated in the striatum ipsilateral to the substantia nigra infused with iron. An improvement by fresh SNAP of iron-induced reduction in dopamine clearance was observed in rats co-infused with fresh SNAP and iron mixture compared to iron-lesioned group. Taken together, our in vivo electrochemical study suggests that nitric oxide does not alter dopamine dynamics in nigrostriatal dopaminergic system. Rather, nitric oxide appears to protect dopamine dynamics from iron-induced oxidative stress in rat brain. PMID- 10515256 TI - Purkinje cell inhibitory responses to 3-APPA (3-aminopropylphosphinic acid) in RAT cerebellar slices. AB - 3-APPA is considered to be a GABA(B) agonist more potent than baclofen. We report here the results obtained by applying this agonist to Purkinje cells (PCs) recorded in current clamp mode on cerebellar slices. The responses were compared to those obtained with other GABA agonists and antagonists. The drugs were delivered either in the perfusion solution or by pressure to the molecular layer near the recorded cell. When applied to the PCs either in the bathing medium or by pressure, 3-APPA evoked a potent inhibitory response which was however different from that obtained with baclofen. The response was complex and similar to that evoked by application of GABA, the endogenous neurotransmitter. In fact it showed: (1) very sensitive dose-response not affected by TTX in the bath; (2) an equilibrium potential compatible with Cl-channel conductance; (3) a massive reduction with the competitive GABA(A) antagonist bicuculline; (4) a small reduction, if any, with the potent competitive GABA(B) antagonist CGP55845A; (5) persistence of the responses under 4-AP (4-aminopyridine), the potassium channel blocker, and inhibition of the 4-AP-induced calcium bursts of spikes. The conclusion was reached that the inhibitory response of PCs to 3-APPA is induced, like GABA inhibition, by binding to both GABA(A) and GABA(B) postsynaptic receptors. PMID- 10515258 TI - The clustering of NMDA receptor NR1 subunit is regulated by the interaction between the C-terminal exon cassettes and the cytoskeleton. AB - The clustering of neurotransmitter receptors at postsynaptic sites is considered to play an important role in modulating synaptic efficacy. To investigate the mechanisms underlying neurotransmitter receptor clustering, we expressed the NMDA (N-methyl-D-aspartate) receptor NR1A subunit in human embryonic kidney (HEK) 293 cells. As previously shown, the cells exhibited subcellular clusters of the receptor protein with a mean diameter of approximately 0.7 microm. To examine the involvement of cytoskeletal structures on this clustering, we disrupted actin filaments or microtubules by treating the cells with alkaloids. In the actin filament-disrupted cells, the receptor protein shifted from the cellular membrane to the cytoplasm where it formed macroclusters (approximate diameter 3 microm). In the microtubule-disrupted cells, the subcellular clusters of NR1A could not be detected and the protein was diffusely distributed throughout the cytoplasm. Similar results were obtained by coexpression of the receptor protein with fusion proteins harboring various C-terminal exon cassettes. These results suggest that subcellular clustering of the NR1 subunit of the NMDA receptors is regulated by the interaction of its C-terminus with cytoskeletal components, where differentially spliced cassettes interact separately with actin filaments or microtubules. Modulation of the interaction between the neurotransmitter receptors and the cytoskeleton leads to the rearrangement of the receptor clusters and may contribute to certain types of synaptic plasticity. PMID- 10515257 TI - Unique behavioural phenotypes of recombinant-inbred CXBK mice: partial deficiency of sensitivity to mu- and kappa-agonists. AB - Recombinant-inbred CXBK mice have been used for various studies as putative mu opioid-receptor deficient mice. However, CXBK mice have never been compared with gene-targeting mice lacking the mu-opioid receptor (muKO) and the K-opioid receptor (kappaKO). Here we report that CXBK mice show distinct behavioural phenotype in opioid-induced analgesia and sedation. Intraperitoneal (i.p.) administration of morphine (3 and 10 mg kg(-1)) induced significantly lower levels of analgesia in CXBK mice than in the control C57BL/6 mice, while higher doses of morphine (30 and 100 mg kg(-1)) induced marked analgesia in CXBK mice. CXBK mice also showed lower analgesia and sedation levels than did C57 mice after i.p. administration of U-50488 (10 and 30 mg kg(-1)). The partial deficiency of sensitivity to morphine and U-50488 of CXBK mice is in sharp contrast to the complete lack of sensitivity to morphine and U-50488 in muKO and kappaKO mice, respectively. Furthermore, CXBK mice showed a lower threshold for nociceptive stimuli when they were not given an opioid, suggesting that CXBK mice could have alterations in the genes related to the nociceptive threshold. These unique behavioural phenotypes of CXBK mice suggest unique genetic alterations in CXBK mice. PMID- 10515259 TI - Down-regulated expression of glutamate transporter GLAST in Purkinje cell associated astrocytes of reeler and weaver mutant cerebella. AB - The glutamate transporter plays an important role in rapid removal of glutamate from the synaptic cleft. Glutamate transporter GLAST is highly expressed in the Bergmann glia (BG), a unipolar cerebellar astrocyte associated structurally and functionally with Purkinje cells (PCs). Here we investigated the expression and localization in the reeler and weaver mutant cerebella with disorganized cytoarchitecture and disrupted synaptic circuitry. In the cortex of both cerebella, GLAST-expressing cells were astrocytes associating PCs; they were located around PC somata and primary dendrites, and extended glial fibrillary acidic protein (GFAP)-immunopositive processes surrounding PC somata and dendrites. Additional signals were detected in astrocytes of the reeler subcortex; they were dispersed among ectopic PCs and had GFAP-positive processes apposing to PC somata and stunted dendrites. Therefore, GLAST expression in PC associated astrocytes was conserved in these mutants. Compared to the wild-type BG, however, the transcription level in individual mutant astrocytes was significantly reduced to about one-third level in the reeler and weaver cortex or one-sixth level in the reeler subcortex. Taking previous results on remarkable up regulation during dendritogenic/synaptogenic stages and down-regulation following experimental glutamatergic denervation, it is suggested that GLAST expression in cerebellar astrocytes is regulated correlatively with cytological and/or synaptic differentiation of neighboring PCs. PMID- 10515260 TI - The site of origin of calcitonin gene-related peptide-like immunoreactive afferents to the inferior olivary complex of the mouse. AB - The intent of the present study is to define the brainstem nuclei which give rise to CGRP-immunolabeled afferents to the inferior olivary complex of the mouse. A technique which combines retrograde transport of fluorescent microspheres with immunohistochemistry was used to address this question. In the present study, intensely labeled CGRP neurons were localized within several cranial nerve nuclei including the hypoglossal, facial, oculomotor, motor nucleus of the trigeminal nerve and nucleus ambiguus, as well as in the parabrachial nucleus, locus coeruleus and medullary and pontine reticular formation. In addition, lightly labeled CGRP neurons were identified within the deep cerebellar nuclei, the inferior olivary complex, lateral reticular nucleus, medial and lateral vestibular nuclei, nucleus Darkschewitsch, interstitial nucleus of Cajal, the central gray area adjacent to the third ventricle, and the zona incerta. The origin of the projection to the inferior olivary complex primarily arises from the deep cerebellar nuclei, the locus coeruleus, and the central gray matter of the mesodiencephalic area. In addition, a small CGRP input is derived from the superior and lateral vestibular nuclei as well as the zona incerta. In conclusion, we have identified several extrinsic sources of CGRP to the inferior olivary complex and have localized it within afferents that have been shown to have either excitatory (mesodiencephalic nuclei) or inhibitory (cerebellar nuclei) effects on olivary circuits. The presence of CGRP in these functionally diverse brainstem and cerebellar afferents suggests that the peptide may act as a co-transmitter to modulate the activity of olivary neurons. PMID- 10515261 TI - Long-term potentiation of Ca2+ signal in the rat auditory cortex. AB - The Ca2+ signal in supragranular layers of the rat auditory cortex (AC) was studied in slice preparations using rhod-2, a Ca2+ indicator. White matter stimulation elicited an increase in the Ca2+ signal, which was maximal in the image taken 34 ms after stimulation. This peak time was the same as that of the Ca2+ signal in pyramidal neurons injected with rhod-2. The intensity of the Ca2+ signal was proportional to the amplitude of the field potentials in supragranular layers. The Ca2+ signal was inhibited almost completely by 200 microM Ni2+ , but only slightly by 50 microM D-2-amino-5-phosphonovalerate (APV), an NMDA-receptor antagonist. Tetanic stimulation of the white matter or supragranular layers elicited long-term potentiation (LTP) of the Ca2+ signal in AC slices, but the potentiation was not clear in slices of the visual cortex (VC). The induction of LTP of the field potentials in AC slices was blocked by 50 microM APV or 50 microM Ni2+. These results indicate that Ca2+ influx through Ni2+ -sensitive Ca2+ channels in pyramidal neurons is potentiated by tetanic stimulation in parallel with LTP of neural activities and might be important for the induction of LTP in AC slices. PMID- 10515262 TI - Morbillivirus infections, with special emphasis on morbilliviruses of carnivores. AB - Morbilliviruses infections cause significant mortality in human beings and animals. Measles virus is responsible for up to two million childhood deaths annually in the developing world, while rinderpest and peste des petits ruminants cause severe epizootics in domestic and wild ruminants in areas of the world where they remain endemic. Canine distemper virus (CDV) is a cause of fatal disease in many species of carnivores. Distemper is controlled by vaccination in domestic dogs and farmed mink, but it may be impossible to eradicate the virus because of its global distribution and wide variety of susceptible host species, which includes both freshwater and marine seals. Research is currently under way to develop new recombinant vaccines, since the currently available live attenuated vaccines for CDV are not safe for use for all species and many valuable zoo animals need to be protected from CDV. New morbilliviruses with potentially disastrous ecological consequences for marine mammals have been discovered in the past decade; phocid distemper virus (PDV) in seals and the cetacean morbillivirus (CMV) has been found in dolphins, whales and porpoises. Reverse transcription, coupled with the polymerase chain reaction (RT/PCR) and nucleic acid sequencing, has been used to characterise the morbilliviruses and has given insights into the evolution of this virus genus. PMID- 10515263 TI - Analysis of the H gene, the central untranslated region and the proximal coding part of the F gene of wild-type and vaccine canine distemper viruses. AB - This paper summarizes the results of the genetic analysis of several parts of the genome of canine distemper virus (CDV) field isolates and vaccine viruses. The haemagglutinin (H) gene analysis showed that recent viruses did not differ significantly from vaccine strains. The analysis of the long untranslated region between the matrix (M) and fusion (F) gene revealed distinct genetic heterogeneity. The putative F protein start codon AUG461 of vaccine strain Onderstepoort was found to be mutated in all wild-type isolates and in another vaccine strain. The proximal coding part of the F gene was well conserved. Phylogenetic analysis of this segment showed the presence of several cocirculating CDV genotypes. PMID- 10515264 TI - Morbilliviruses in Mediterranean monk seals. AB - Two morbilliviruses were isolated from Mediterranean monk seals (Monachus monachus), one from a stranded animal in Greece and the other one from carcasses washed ashore during a mass die-off in Mauritania. From both viruses N and P gene fragments were sequenced and compared to those of other known morbilliviruses. The monk seal morbilliviruses most closely resembled previously identified cetacean morbilliviruses, indicating that interspecies transmission from cetaceans to pinnipeds has occurred. PMID- 10515265 TI - Restricted virus protein translation in canine distemper virus inclusion body polioencephalitis. AB - In this study, inclusion body polioencephalitis, an uncommon form of canine distemper virus (CDV)-induced encephalitis, was investigated for viral protein and mRNA expression by immunohistochemistry (IH) and in situ hybridization and, in addition, infiltrating cells were characterized by IH. Lesions were predominantly found in the grey matter of the brain stem and the immune response, dominated by T cells, was associated with a strong MHC II upregulation. Abundant expression of all viral protein mRNAs and reduced or lacking protein translation, especially of the matrix protein were the most important findings, indicating that restricted virus infection in the grey matter might represent a mechanism for viral persistence in distemper polioencephalitis. PMID- 10515266 TI - Host range relationships and the evolution of canine parvovirus. AB - Canine parvovirus (CPV) is an example of an unusual class of emerging virus-those that gain an altered host range through genetic variation and subsequently become widespread pathogens of their new and previously resistant host species. CPV was first detected in 1978 as the cause of new diseases in dogs throughout the world, when it rapidly spread throughout domestic populations, as well as becoming widespread in wild dogs. CPV was soon shown to be a variant of the long recognized feline panleukopenia virus (FPV), from which it differed in less than 1% at the nucleotide sequence level. Genetic analysis showed that virtually all of the biological differences between CPV and FPV, including the canine host range, were determined by three or four sequence differences in the viral capsid protein gene. Analysis of the atomic structures of the CPV and FPV capsids showed that the differences controlling host range were located within two different structural regions and were exposed on the capsid surface. The CPV which first emerged in 1978 appeared to be derived from a single ancestral sequence, which has allowed the ready analysis of the subsequent evolution of the virus in nature. Sequence analysis has also revealed that CPV strains have undergone a series of evolutionary selections in nature which have resulted in the global distribution of new virus variants. This was first seen in the global replacement between 1979 and 1981 of the original (1978) strain of the virus by a genetically and antigenically variant strain, and the subsequent widespread selection of other variants which have also become globally distributed. The genetic and antigenic variation in the virus strains was also correlated with changes in the host range of the virus, in particular in the ability to replicate in cats, and in canine host range differences seen in tissue culture cells. PMID- 10515267 TI - Efficacy of feline panleucopenia vaccine to prevent infection with an isolate of CPV2b obtained from a cat. AB - Cats vaccinated against FPLV were protected against infection with a feline isolate of CPV2b. Nonvaccinated cats developed a lymphopenia and excreted virus which infected susceptible in-contact cats. PMID- 10515268 TI - Emergence and recent evolution of canine parvovirus. AB - This review summarizes the current knowledge about the emergence of canine parvovirus from an ancestor virus similar to feline panleukopenia virus most likely from a wild carnivore host. The recent evolution of CPV, namely the emergence of new antigenic types, their biological properties and global distribution are also discussed. PMID- 10515270 TI - Caliciviruses: an overview. AB - Several caliciviruses are known to cause diseases in animal and man. Amongst them are vesicular exanthema virus of swine (VESV), feline calicivirus, rabbit hemorrhagic disease virus (RHDV), and Norwalk and Sapporo-like viruses; the latter viruses cause gastroenteritis in man. In this paper, an overview of caliciviruses is presented, with particular emphasis on the molecular aspects. The unique properties as compared to other related families of positive-stranded RNA viruses will be discussed. Many findings about the molecular biology of caliciviruses came from systems that are difficult to work with, such as RHDV, which can thus far only be propagated either in animals or in primary rabbit hepatocyte cultures. Therefore, the review will concentrate on RHDV. PMID- 10515269 TI - Feline interferon-omega treatment on canine parvovirus infection. AB - Recombinant feline interferon-omega preparation (rFeIFN-omega, trade name: INTERCAT) showed good clinical efficacy on canine parvovirus infection both in an experimental trial with beagles, and in field trials. PMID- 10515271 TI - Feline calicivirus capsid protein expression and self-assembly in cultured feline cells. AB - Feline calicivirus (FCV) capsid protein was expressed in feline cells employing the vaccinia virus MVA/T7 RNA polymerase system. The precursor protein was processed to a mature size protein that assembled to virus like particles (VLPs). PMID- 10515272 TI - Quasispecies evolution of a hypervariable region of the feline calicivirus capsid gene in cell culture and persistently infected cats. AB - The study determines the sequence evolution of feline calicivirus both in cell culture and in persistently infected cats and relates this to changes in virus neutralisation. PMID- 10515273 TI - Genetic analysis of a canine calicivirus: evidence for a new clade of animal caliciviruses. AB - This paper describes the relationship of a canine calicivirus, named No.48, to other human and animal caliciviruses, based on phylogeny of the 3' half of its genome. It was found that No.48 constitutes a unique lineage, most closely related but distinct from feline and San Miguel sea lion caliciviruses. PMID- 10515274 TI - Herpesviruses of carnivores. AB - This review focuses on felid herpesvirus 1 (FHV-1), the most studied of the carnivore herpesviruses. Canid herpesvirus (CHV-1) and phocid (seal) herpesvirus 1 (PhHV-1) are also included where information is available. FHV-1 is a member of the Varicellovirus genus of the Alphaherpesvirinae, which appears to be closely related phylogenetically to both CHV-1 and PhHV-1. FHV-1 infects both domestic and some wild Felidae, such as cheetahs, and is predominantly a respiratory pathogen of cats. As in other herpesviruses, infection with FHV-1 is characterised by a latent carrier state, during which intermittent shedding of infectious virus may occur. Typical of an alphaherpesvirus, the primary site of FHV-1 latency is neurological tissue (trigeminal ganglion), though recent studies using the polymerase chain reaction have suggested that some latency may occur in non-neurological sites. Latently infected carriers are epidemiologically important as sources of infection for susceptible animals. Though conventional modified live and inactivated vaccines have been available for a number of years, they do not protect against infection nor the development of latency. Recently, work has focused on molecular characterisation of FHV-1, detecting genes such as glycoproteins or regulatory genes. Such work will enable better understanding of the interaction of FHV-1 with the natural host. Deletion mutants of some of these genes may also have potential as vaccine strains. PMID- 10515275 TI - Structure-function analysis of the feline herpesvirus virulence factors gE and gI. AB - The biosynthesis and function of the gE-gI complex of feline herpesvirus (FHV) was studied by heterologous expression of the cloned genes in the vaccinia virus based vTF7-3 expression system and by analysis of FHV recombinants. This work has led to the identification of domains in gI involved in complex formation with gE, and to a model for the disulfide-bonded structure of gI. The effects of mutations in gI on gE-gI-dependent cell-to-cell transmission are discussed. PMID- 10515277 TI - Strategies of retrovirus survival in the cat. AB - Retroviruses establish persistent infections in their hosts which often lead to serious and fatal diseases after a long incubation period. The molecular basis of this persistence is the integration of a copy of the viral genome into cellular chromosomal DNA. At the level of the whole animal, however, each retrovirus genus has evolved a different strategy to ensure its survival. This variety is well illustrated in the cat. Feline leukaemia virus, an oncovirus, has a simple genomic structure and survives in its host by suppressing the immune response to the virus. As a result, this virus is antigenically highly conserved. By contrast, feline immunodeficiency virus and feline foamy virus, representatives of the lentiviruses and spumaviruses, respectively, have more complex genomes which include genes responsible for maintaining the virus in a latent state thereby avoiding elimination in the face of a powerful antiviral immune response. In the lentiviruses, this response drives the selection of viruses exhibiting variation in antigenicity and pathogenicity. PMID- 10515276 TI - Molecular phylogenetic analysis of felid herpesvirus 1. AB - The position of felid herpesvirus 1 within the alphaherpesvirus subfamily was investigated using molecular phylogenetic techniques applied to multiple sequence alignments of recently reported FHV-1 gene homologs (glycoprotein B, ribonucleotide reductase and DNA polymerase). FHV-1 was most closely related to other carnivore alphaherpesviruses, (phocid herpesvirus 1 and canid herpesvirus 1) and to the equid herpesviruses 1 and 4. PMID- 10515278 TI - Vaccination of cats against feline immunodeficiency virus (FIV): a matter of challenge. AB - So far, most apparently successful immunodeficiency virus vaccines have only been tested against challenge with cell culture-adapted virus. However, even live priming of cats with feline herpesvirus (FHV) vectors expressing the FIV gag and env gene followed by inactivated booster vaccination with fixed FIX infected cell vaccine proved non-protective against infection with a primary FIV isolate. An effective FIV vaccine for field applications therefore is still not underway. PMID- 10515279 TI - Treatment of feline leukemia virus (FeLV) infection. AB - FeLV infection is still considered to account for most disease-related deaths in pet cats. Different treatment attempts with various drugs were performed in the past but none resulted in healing or complete virus elimination. Therefore, it caused a sensation when Horber and Mayr [Horber, D., Mayr, B., 1991. Prax. 19, 311-314; Horber, D., Schnabl, W., Mayr, B., 1992. Tierarztl. Umschau 47, 556-560; Mayr, B., Horber, D., 1992. Kleintierprax. 37, 515-518] published that they were able to cure 80 to 100% FeLV-infected cats from viremia by using an immunomodulating compound. Articles in cat breeder and cat owner journals appeared assuming that obviously there is a rescue for FeLV-infected cats suffering from this deadly infection. The immunomodulator [Buttner, M., 1993. Comp. Immun. Microbiol. Infect. Dis. 18, 1-10] used in those studies was the so called 'paramunity inducer' PIND-ORF (Baypamun, Bayer, Leverkusen, Germany) consisting of inactivated parapox ovis virus. Since that time, Baypamun is the most commonly used drug for treatment of FeLV infection in Germany and other European countries. Four placebo-controlled double-blind trials were performed to determine the therapeutic efficacy of Baypamun and other compounds in naturally FeLV-infected cats under controlled conditions. PMID- 10515280 TI - The feline immunodeficiency virus envelope protein precursor: functional analysis of a leader deletion mutant. PMID- 10515281 TI - The molecular dynamics of feline coronaviruses. AB - Feline coronaviruses are widespread and come in different flavors. There are two main serotypes both of which occur in two pathotypes, the avirulent enteric viruses and the virulent, usually fatal peritonitis viruses, the latter in turn occurring either in a 'wet' or exudative form or in a 'dry' or proliferative form. In this paper a concise overview is given of the molecular features of these viruses. Special attention is given to the genetic dynamics of the viruses as these now allow us to begin to understand the origin of the different phenotypes, in particular the genesis of virulence during persistent infection. As discussed, the surprising new insights obtained over the last few years call for a critical reevaluation of strategies for protection. PMID- 10515282 TI - Detection of FcoV quasispecies using denaturing gradient gel electrophoresis. PMID- 10515283 TI - Histopathological alterations of lymphatic tissues in cats without feline infectious peritonitis after long-term exposure to FIP virus. PMID- 10515284 TI - Genetic drift and genetic shift during feline coronavirus evolution. PMID- 10515285 TI - Relationship between platelet activation and cytokines in systemic inflammatory response syndrome patients with hematological malignancies. AB - We investigated the significance of platelet activation and platelet-derived microparticles (PMP) in 14 patients with systemic inflammatory response syndrome (SIRS) and hematological malignancies. In the phenotypic analysis of lymphocytes, there was a significant decrease of total and activated T cells after panipenem/betamipron (PAPM/BP) treatment (p<0.05). The percentages of helper/inducer T cells and suppressor/cytotoxic T cells were insignificantly decreased after PAPM/BP treatment. The number of natural killer (NK) cells of potent activity was significantly decreased after treatment (p<0.05). The levels of the cytokines interleukin (IL)-1beta, IL-6, and IL-8 in the patients were increased before treatment. IL-1beta concentrations were not changed after treatment. In contrast, the IL-6 and IL-8 levels were significantly decreased (p<0.05) after treatment, while tumor necrosis factor (TNF)-alpha and interferon gamma remained almost normal. We found an increase of soluble IL-2 receptor (sIL 2R) and soluble vascular cell adhesion molecule-1 (sVCAM-1) levels in the patients before treatment. After treatment, the sIL-2R concentrations tended to be decreased and sVCAM-1 levels showed a significant decrease (p<0.01). In contrast, soluble thrombomodulin (sTM) level did not change. Regarding the platelet activation markers, CD62P, CD63, and PMP levels in the patients were increased before treatment. CD62P and CD63 tended to be decreased after treatment, whereas PMP levels were significantly reduced from 1,056+/-103 to 762+/-64/10(4) platelets (p<0.05). Furthermore, CD62P, CD63, and PMP correlated with the levels of IL-6 and IL-8. These results suggest that activated platelets and PMP may be predictive markers in pre-disseminated intravascular coagulation and hypercytokine conditions related to SIRS. PMID- 10515287 TI - Reduction of platelet activity markers in type II hypercholesterolemic patients by a HMG-CoA-reductase inhibitor. AB - We have investigated the effects of a potent lipid-lowering therapy on the activity of platelets as measured ex vivo by the surface activation markers CD62 (PADGEM, P-selectin, GMP 140) and CD63 (GP53) in a double-blind, randomized, placebo-controlled study. Treatment with the HMG-CoA-reductase inhibitor fluvastatin (40 mg) significantly reduced the serum low density lipoprotein cholesterol concentration by 30% (p<0.01) and total cholesterol by 25% (p<0.01). The platelet membrane activation markers CD62 (PADGEM, P-selectin, GMP140) and 63 (GP53) significantly decreased by 22 and 13% (in terms of the relative fluorescence intensity) under the treatment with fluvastatin (p<0.05), respectively. The cholesterol-lowering effect is accompanied by a significant reduction of the platelet membrane activation markers CD62 and CD63 reflecting a reduced platelet activity that may contribute to the vasoprotective profile of fluvasatin. PMID- 10515286 TI - Studies on the dual effects on platelets of a monoclonal antibody to CD9, and on the properties of platelet CD9. AB - The article describes effects on human platelets of a murine monoclonal antibody of the IgG2a subtype (clone FN99) directed against the membrane glycoprotein CD9. This antibody exerts a dual action on human platelets in plasma depending on whether the complement system can be activated or not, resulting either in membrane permeabilization or a true platelet aggregation. Secretion from the alpha-granules during permeabilisation was not observed in the sense that the granule-located protein thrombospondin was retained in the platelets, as opposed to what was seen with platelets that had undergone an antibody-induced aggregation. Only a small fraction of P-selectin was found on the surface of the permeabilised platelets. The cytoskeletal protein actin-binding protein (filamin) was profoundly degraded during membrane permeabilisation, however, and scanning electron microscopy showed platelets that were swollen with only a few pseudopodia. Preincubation of platelets with three different antibodies to CD9 showed strong inhibition of a subsequent binding of FITC-labelled Fab fragment of FN99 indicating that antibodies tend to bind in the same area of the CD9 molecule. No association of CD9 to the platelet actin-based cytoskeleton was observed. CD9 was present on the surface of microvesicles derived from calcium ionophore-treated platelets. PMID- 10515288 TI - p-Aminobenzoic acid and its metabolite p-acetamidobenzoic acid inhibit agonist induced aggregation and arachidonic acid-induced [Ca2+]i transients in human platelets. AB - We have previously found that the naturally occurring amine p-aminobenzoic acid (PABA) inhibits the thrombin-induced thromboxane B2 production in human platelets. In this report we show that PABA and its acetylated metabolite p acetamidobenzoic acid (PACBA) inhibit platelet aggregation induced by agonists such as adenosine diphosphate (ADP) and arachidonic acid (AA). Both substances were equipotent to acetylsalicylic acid regarding inhibition of ADP-induced aggregation and approximately 50% as potent as acetylsalicylic acid regarding arachidonic acid-induced aggregation. Although not significantly inhibiting collagen aggregation, PABA and PACBA reduced the concomitant adenosine triphosphate (ATP) secretion by approximately 30 and 20%, respectively. The antiaggregatory effect does not seem to be mediated through cyclic adenosine monophosphate (cAMP) increase because in our experiments PABA and PACBA did not significantly affect cAMP levels. However, we have found that PABA and PACBA inhibit the intracellular aequorin indicated Ca2+ transient upon arachidonic acid stimulation. Our results describe a hitherto unknown effect of PABA and PACBA on platelet aggregation. PMID- 10515289 TI - Reversible exposure of human platelet fibrinogen receptors by antiplatelet tetraspanin monoclonal antibodies via induction of a conformational change in membrane glycoprotein IIb/IIIa complex. AB - Antihuman platelet tetraspanin (CD9 antigen) monoclonal antibodies, HI117 and SJ9A4, can induce human platelet aggregation and secretion. As platelet aggregation is mediated by fibrinogen binding to its receptors exposed on platelet glycoprotein IIb/IIIa complex, we, therefore, investigated the induction of platelet fibrinogen receptors by HI117 and SJ9A4. It was found that HI117 and SJ9A4 induced specific fibrinogen binding to human platelets, suggesting that the two monoclonal antibodies evoked obvious exposure of fibrinogen receptors on human platelets. But in the absence of fibrinogen, the monoclonal antibody exposed fibrinogen receptors gradually lost their capacity to bind fibrinogen and closed. Our results also showed that HI117 and SJ9A4, when activating platelets, caused a conformational change in glycoprotein IIb/IIIa complex, which must contribute to the exposure of functional fibrinogen receptors on this integrin. The effect of HI117 and SJ9A4 on glycoprotein IIb/IIIa complex seems, however, to be indirect, because the HI1117 and SJ9A4-induced fibrinogen binding was reduced by pretreatment of platelets with sphingosine, aspirin, apyrase, and/or PGI2. Taken together, we conclude that the antihuman platelet tetraspanin monoclonal antibodies, HI117 and SJ9A4, reversibly expose platelet fibrinogen receptors via inducing a conformational change in glycoprotein IIb/IlIa complex. Three signaling pathways, that is, thromboxane, secreted ADP, and cAMP pathways may be involved in this process, while protein kinase C activation seems to be the final common step of the three pathways. PMID- 10515290 TI - The promoter of human tissue factor pathway inhibitor gene: identification of potential regulatory elements. AB - Tissue factor pathway inhibitor is the major potent physiologic inhibitor of tissue factor-induced coagulation. Several potential binding sites for transcription factors have been described in the 750 bp of the 5' flanking region of the human tissue factor pathway inhibitor gene reported earlier. To identify elements that regulate the expression of tissue factor pathway inhibitor in endothelial, hepatocyte, and monocyte cells, the sequence of an additional 770 bp of tissue factor pathway inhibitor was determined. Comparison of this new sequence as well as that reported earlier with consensus sequences for transcription factor binding sites provided matches for GATA-2, SP1, and c-Myc sequences. Moreover, plasmids containing deletion mutants of the 5' tissue factor pathway inhibitor promoter region and the luciferase reporter gene were transfected into HepG2, ECV304, and THP1 cells. Three negative regulatory elements were localized between -548 to -390, - 390 to -75, and -1158 to -796 relative to the transcriptional start, respectively, in HepG2, ECV304 and THP-1 cells. PMID- 10515292 TI - Discriminative stimulus properties of antipsychotics. AB - Drug discrimination methodology has been used in a number of ways to analyze the actions of novel and putative novel antipsychotics in vivo. Recent studies suggest (a) in contrast to earlier theorizing, antagonism of the low-dose d amphetamine stimulus in rats may not be an effective screen for novel antipsychotics; (b) dopamine D2-like agonists and antagonists, some of which are putative antipsychotics, can be studied in vivo as discriminative cues, although there is a pressing need for more selective drugs that differentiate the various members of the D2 family. (c) antagonism of the cue induced by the noncompetitive NMDA antagonist MK-801, which has been proposed as a possible screen for clozapine-like compounds, may be an unreliable assay; and (d) the clozapine stimulus is probably a compound cue (a drug "mixture"), which can be used to screen for novel clozapine-like antipsychotics, although the precise receptor mechanisms involved in mediating the clozapine stimulus, and its direct relevance to the antipsychotic action of clozapine remains to be proven conclusively. PMID- 10515291 TI - Plasma D-dimer testing improves the management of thromboembolic disease in hospitalized patients. PMID- 10515293 TI - Discriminative and affective stimulus effects of dihydropyridine calcium channel modulators: relationship to antialcohol effects. AB - Voltage-operated calcium channels (VOCCs) have been implicated in alcoholism. Thus, dihydropyridine (DHP) VOCC antagonists, such as nimodipine, reduce ethanol (EtOH) intake and preference in a variety of animal models of alcoholism. Paradoxically, the DHP VOCC agonist BAY k 8644 also demonstrates antialcohol effects in such models. The antialcohol effects of BAY k 8644 are stereoselective [the "agonistic" (-)-enantiomer being more potent than the "antagonistic" (+) enantiomer], and are not blocked by pretreatment with nimodipine. The present review summarizes studies on the effects of DHPs in drug discrimination (DD), conditioned taste aversion (CTA), and conditioned place preference (CPP) paradigms, and discusses the possibility that the apparent antialcohol effect of these compounds is related to their discriminative and/or affective stimulus effects. In rats trained to discriminate nimodipine from vehicle, (-)-BAY k 8644 completely generalizes to the nimodipine cue; whereas, in rats trained to discriminate (-)-BAY k 8644, nimodipine completely generalizes to, and is unable to block, the (-)-BAY k 8644 cue. The same stereoselectivity is obtained for BAY k 8644 in DD paradigms and models of alcoholism. The apparent similarity of these profiles of activity suggests that a common neurobiological mechanism underlies the discriminative stimulus and antialcohol effects of DHPs. It appears unlikely, however, that the antialcohol effects of DHPs are based on substitution for, or blockade of, the EtOH cue, as these compounds were not found to generalize to, or block, the EtOH cue. Comparison of the effects of DHPs in CTA and CPP paradigms suggests that the affective stimulus effects of these compounds are dissimilar, and that the mechanism underlying the latter effects is probably not related to the mechanism underlying the antialcohol effects of DHP VOCC modulators. PMID- 10515294 TI - Stimulus properties of the selective 5-HT reuptake inhibitor fluvoxamine in conditioned taste aversion procedures. AB - Previous attempts to train pigeons and rats to discriminate between the antidepressant fluvoxamine and its vehicle as assessed in a drug discrimination paradigm have been without success. The present experiments were, therefore, designed to assess in a conditioned taste aversion procedure (CTA) whether or not fluvoxamine possesses stimulus properties. Rats were exposed to a conditioned taste aversion (CTA) procedure. In Experiment I, subjects were given 15 mg/kg fluvoxamine p.o. or vehicle after drinking a novel tasting saccharin solution. In Experiment II, a comparison was made between the effects of 15 mg/kg fluvoxamine i.p., 30 mg/kg fluvoxamine i.p., NaCl, and lithium chloride (LiCl). In Experiment III, subjects were treated with either 10 mg/kg fluoxetine i.p., 30 mg/kg fluvoxamine i.p., or LiCl. CTA was observed after treatment with LiCl, but never after treatment with fluvoxamine or fluoxetine, suggesting that fluvoxamine does not have clear stimulus properties, which can serve as a discriminative stimulus in operant procedures. In a crossfamiliarization CTA procedure in mice, however, fluvoxamine elicited a reliable CTA, suggesting that under certain conditions (species, dose?) selective serotonin reuptake inhibitors (SSRIs) may lead to certain discriminable effects. It is as yet unclear why SSRIs apparently produce such weak and species or situation-dependent discriminable effects. PMID- 10515296 TI - Discriminative effects of compound drug stimuli: a focus on attention. AB - Discrimination research has increasingly used compound stimuli emerging from drugs acting through multiple neurotransmitter systems or from injections of drug mixtures that approximate "street-wise" drug-taking behaviors. Accompanying this trend has been an interest in the role of cognitive factors in drug discrimination learning. Accounts of multidimensional drug stimuli have focused mainly on specific neuronal mechanisms, and have largely ignored the contribution of stimulus information to the perception of internal events or to the selection processes, heretofore called attention mechanisms, which may underlie the observer's idiosyncratic response to drug administration. It is argued here that research in drug discrimination may benefit from a more detailed consideration of the processes by which an observer interacts with the emergent stimulus properties of drug administration. Therapeutic intervention initiatives may critically depend on knowing the interactions between the specific attributes of the drug experience that capture the attention of the individual and that may later acquire stimulus control over complex drug-taking behaviors. PMID- 10515295 TI - Drug mixtures and ethanol as compound internal stimuli. AB - Drug discrimination methods that entail training with mixtures of drugs may shed light on polydrug abuse and on the actions of single drugs that interact with more than one receptor. In AND-discrimination procedures (drug A + drug B vs. vehicle), mixtures are discriminated primarily on the basis of their component drugs: these discriminations may be useful for testing interactions between component drugs in mixtures. The role of training dose, overshadowing and associative blocking in AND-discriminations have been investigated. For example, after prior training with midazolam, it was possible to demonstrate associative blocking of the nicotine element of the mixture stimulus, and vice versa. Using the AND-OR discriminations (drug A + drug B vs. drug A or drug B) increased pharmacological specificity considerably, and these procedures may be valuable for determining whether the effects of a novel mixture are similar to the combined effects of the training drugs. Ethanol is an example of a single drug that may produce a compound cue; rats trained to discriminate ethanol from water generalize (asymmetrically) to GABA(A) enhancers such as chlordiazepoxide (CDP) or pentobarbitone, to NMDA antagonists such as dizocilpine (MK-801), and to some serotonin agonists, such as trifluoromethylphenylpiperazine (5-HT(1B/2C)). In addition, rats trained to discriminate mixtures of either CDP or pentobarbitone plus MK-801 generalize to ethanol. A previous history of training with MK-801 or CDP (prior to ethanol discrimination training) enhanced the MK-801-like and CDP like effects of ethanol respectively, but associative blocking of proposed elements in the ethanol stimulus was not seen. These studies provide some support for the multielement concept of ethanol discrimination but also suggest that rules governing three-component stimuli (such as those putatively produced by ethanol) may differ from those for the two-component mixtures of drugs studied previously. PMID- 10515297 TI - High-dose discrimination training with midazolam: context determines generalization profile. AB - In previous work, greater differentiation among ligands for the benzodiazepine site was found in rats trained to discriminate among vehicle, 0.32, and 3.2 mg/kg midazolam than in animals trained to discriminate a single midazolam dose from vehicle (i.e., virtually all test drugs occasioned low-dose midazolam-appropriate responding, but most did not occasion high-dose midazolam-appropriate responding even at high test doses). A possibility was that merely training with 3.2 mg/kg midazolam (not previously studied) would result in greater selectivity than training with lower midazolam doses. In the present study, rats were trained to discriminate 3.2 mg/kg i.p. midazolam from no drug under a two-lever, food maintained, procedure; and drugs from the previous three-lever studies were tested. Triazolam, bretazenil, clonazepam, lorazepam, midazolam, zolpidem, chlordiazepoxide, pentobarbital, and flurazepam all dose-dependently occasioned >80% responding on the midazolam-appropriate lever in roughly that order of potency. Only triazolam had occasioned midazolam 3.2 mg/kg-appropriate responding in the previous work. The greater differentiation among these drugs in the dose vs.-dose procedure likely was due to a training dose context rather than to the high training dose per se. PMID- 10515298 TI - The novel-response procedure in humans. AB - The novel-response drug discrimination procedure is one of several three-choice procedures developed to address interpretational difficulties that can occur under standard two-response procedures. The novel-response procedure is unique among three-choice discrimination procedures by using instructions, rather than explicit training procedures. With the novel-response procedure, participants are trained under a standard two-response (drug vs. placebo) discrimination, and then instructed that in the presence of a drug stimulus unlike either of the training drugs, responses should be made on the novel-response alternative. Several studies have assessed the utility of the novel-response procedure by comparing effects under a standard two-response and the novel-response procedure in participants trained to discriminate triazolam from placebo. Results indicate that the novel-response procedure can increase the selectivity of both placebo- and drug-appropriate responding, and in this way, allows for finer distinctions to be made among sedatives than a standard two-response procedure. PMID- 10515299 TI - Arylalkylamine drugs of abuse: an overview of drug discrimination studies. AB - Abused arylalkylamines fall into two major categories: the indolealkylamines, and the phenylalkylamines: These agents can be further subclassified on the basis of chemical structure. Examples of these agents possess hallucinogenic, stimulant, and other actions. Drug-discrimination techniques have been used to classify and investigate this large family of agents. Such studies have allowed the formulation of structure-activity relationships and investigations of mechanisms of action. Arylalkylamine designer drugs also possess the same or a combination of actions, and are being investigated by the same methods. PMID- 10515300 TI - Cannabis: discrimination of "internal bliss"? AB - The recent discovery of arachidonylethanolamide (anandamide), an endogenous ligand for cannabinoid receptors, and the synthesis of SR141716A, a cannabinoid antagonist selective for brain cannabinoid (CB1) receptors, have provided new tools to explore the mechanisms underlying cannabis abuse and dependence. Drug discrimination is the animal model with the most predictive validity and specificity for investigation of the psychoactive effects of cannabinoids related to their abuse potential, because, unlike many other drugs of abuse, delta9 tetrahydrocannabinol (delta9-THC), the major psychoactive ingredient of marijuana, is not self-administered by animals. Results of delta9-THC discrimination studies have revealed that the subjective effects of cannabis intoxication are pharmacologically selective for centrally active cannabinoid compounds, and that cannabis action at CB1 receptors is involved in medication of these effects. Less clear is the role of endogenous cannabinoid system(s) in cannabis intoxication. Anandamide, named for a Sanskrit word for "internal bliss," unreliably substitutes for delta9-THC. Further, substitution, when it is observed, occurs only at doses that also significantly decrease response rates. In contrast, delta9-THC and other structurally diverse cannabinoids fully substitute for delta9-THC at doses that do not substantially affect response rates. Attempts to train animals to discriminate anandamide (or SR141716A) have so far been unsuccessful. Preliminary evidence from drug discrimination studies with more metabolically stable anandamide analogs have suggested that these differences in the discriminative stimulus effects of delta9-THC and anandamide like cannabinoids are not entirely due to pharmacokinetic factors, but the exact role of "internal bliss" in cannabis intoxication and dependence is still not completely understood. PMID- 10515301 TI - Strategies for understanding the pharmacological effects of ethanol with drug discrimination procedures. AB - Ethanol appears to produce a stimulus complex, or compound cue, composed of distinct components that are mediated by different receptor systems. In ethanol vs. water discriminations, it appears that ethanol produces a redundant stimulus complex such that separate, receptor-mediated activity can serve as the basis for the discrimination. These discriminations have been termed redundant, because multiple features of the cue could serve as the basis of the discrimination. In ethanol vs. water discriminations, one common feature is the asymmetrical generalizations between components of the ethanol cue and ethanol. There is also evidence for overshadowing of one component by other components of the ethanol stimulus complex. It appears possible to transfer the basis of the ethanol cue from a redundant cue to a conditional cue with specific training procedures. When the discriminative stimulus effects of ethanol are juxtoposed with those of one component of the ethanol complex, as in ethanol vs. water vs. pentobarbital discriminations, the ethanol discrimination shifts to a conditional basis. The ability to antagonize an ethanol discrimination may be dependent upon whether the discrimination is based on redundant component stimuli or conditional presence of all component stimuli. PMID- 10515302 TI - Discriminative stimulus effects of drugs acting at GABA(A) receptors: differential profiles and receptor selectivity. AB - The GABA(A) receptor complex contains a number of binding sites at which a variety of psychotropic drugs, including benzodiazepines, barbiturates, and some neurosteroids, act to potentiate or inhibit the effect of the transmitter. Many studies have reported that these drugs can produce discriminative stimulus actions, but the cueing effects of compounds acting at different sites to enhance the effects of GABA are not identical. The discriminative stimulus effects of benzodiazepines have been analyzed in detail, and there is also a great deal of information available on the effects of nonbenzodiazepine compounds acting at BZ(omega) recognition sites, which form part of the GABA(A) receptor complex. Of particular interest are compounds with selectivity for the BZ1(omega1) receptor subtype including zolpidem, zaleplon, and CI 218,872. BZ1(omega1)-selective drugs substitute for the discriminative stimulus produced by chlordiazepoxide only partially and at sedative doses. This is consistent with the view that sedative effects of BZ(omega) receptor agonists are mediated by the BZ1(omega1) receptor subtype, whereas the discriminative stimulus produced by chlordiazepoxide may be produced by activity at the BZ2(omega2) subtype. Analysis of this hypothesis is complicated by the variety of levels of intrinsic activity shown by different drugs. PMID- 10515303 TI - N-methyl-D-aspartate antagonists and drug discrimination. AB - Excitatory amino acids (EAA), such as glutamate, are thought to be involved in various disorders (e.g., ischemic brain damage, epilepsy, Parkinson's disease), and EAA antagonists have been suggested as potential treatments for these disorders. Phencyclidine (PCP), with produces psychotomimetic effects in humans, has antagonist properties at the N-methyl-D-aspartate (NMDA) subtype of glutamate receptors that have been suggested to underlie some of its actions. This suggestion, and concern about possible psychotomimetic activity, has stimulated research aimed at examining to what extent the behavioral profile of other NMDA antagonists resembles that of PCP. Drug discrimination (DD) is prominent among the procedures used to carry out such comparisons. The results of clinical studies with NMDA antagonists provide feedback about the predictive validity of the DD procedures used to characterize their preclinical behavioral profile. Further, DD is used also to examine the ability of compounds to attenuate the discriminative stimulus (DS) effects of PCP-type drugs, and results of such studies have been suggested to provide evidence of antipsychotic potential. Finally, although many instances of intermediate responding in DD can be explained by low efficacy at the receptors that mediate the DS effects of the training drug, certain outcomes produced by PCP-type drugs do not offer valid measures of efficacy, and require more detailed behavioral analyzes. PMID- 10515304 TI - Serotonergic receptor subtypes and hallucinogen-induced stimulus control. AB - More than a quarter century has passed since the demonstration that indoleamine and phenethylamine hallucinogens can function as discriminative stimuli in the rat, and that serotonergic systems are critically involved. During that period our knowledge of the physiology, pharmacology, biochemistry, and molecular biology of serotonergic receptors has increased exponentially; with each advance it has been necessary to reexamine our assumptions regarding hallucinogen-induced stimulus control. Of particular interest is the hypothesis that a drug may act, at a molecular level, upon multiple receptors to produce, at a behavioral level, a compound discriminative stimulus. The salience of the individual elements of such compound stimuli may be influenced by a variety of experimental factors including training dose, pretreatment time, the state of sensitization of the systems being acted upon, and the nature of the drugs chosen for tests of generalization. This article provides examples of experimental approaches to these complexities using selective agonists and antagonists, depletion-induced sensitization, and antagonist correlation analysis. PMID- 10515305 TI - Nicotine discrimination in men and women. AB - Nicotine is the primary compound that maintains tobacco smoking behavior, and nicotine reinforcement may be related to its discriminative stimulus effects. Nicotine in novel form, isolated from tobacco smoke, is often reinforcing in men but not in women, and clinical trials with nicotine replacement via gum or patch have often shown less efficacy in women vs. men trying to quit smoking. We hypothesize that this sex difference in nicotine reinforcement or clinical efficacy may be related to reduced intensity of nicotine's discriminative stimulus effects in women. Using formal drug discrimination procedures, we have found in several studies that discrimination responding across nasal spray nicotine doses tends to be flatter for women than men (i.e., sex x dose interaction), suggesting reduced sensitivity to changes in dose. Results from the field of psychophysiology, involving detection of physiological changes, are generally consistent with our findings, and suggest that the environmental context accompanying physiological change is important in understanding this sex difference. The implications of this sex difference for smoking cessation treatment and future research directions are presented. PMID- 10515306 TI - Relationship of components of an alcohol interoceptive stimulus to induction of desire for alcohol in social drinkers. AB - The ability of a low (0.2 g/kg) oral dose of ethanol to provide a drug discriminative stimulus was studied in young healthy human volunteers, who were social drinkers. Seventeen of 24 subjects acquired the discrimination following 10 trials in which they received aliquots of ethanol or of placebo drink (tonic water mixed with Tabasco sauce). In generalization studies, in which the dose of ethanol was varied, discrimination performance was dose dependent; doses greater than 0.05 g/kg gave rise to significant ethanol-appropriate responding. Concurrent estimates of the subjective effects of doses administered as discriminative stimuli revealed that two factors--taste and light-headedness- were associated with discrimination: at the training dose, 0.2 g/kg, although both the factors taste and light-headedness were significantly increased, only taste predicted discrimination performance. At lower doses, taste did not contribute to discrimination, but the subjective rating light-headedness correlated significantly with discrimination accuracy. Post hoc analyses of the influence of the amount of alcohol regularly drunk by the volunteers, on discrimination performance suggested light-headedness correlated with discriminative performance only in social drinkers drinking more than 20 units per week. In a second experiment, groups of "high" (mean 40 units per week) and "low" (mean 10 units per week) social drinkers were prospectively identified. Discrimination performance of 0.2 g/kg ethanol in orange juice vs. orange juice vehicle indicated that both groups were able to perform the discrimination following a single training trial, and that generalization curves over the range 0.05-0.2 g/kg were dose dependent, and not different between the groups. At the lowest dose, discrimination performance was predicted by taste, stimulation, and light-headedness in the "high" group, but not in the "low" group. The ability of these ethanol doses to induce feelings of craving for ethanol were assessed in parallel, using the Desire for Alcohol Questionnaire (DAQ). "High" drinkers showed higher desire for ethanol on all factors of the DAQ except the "positive negative reinforcement" factor, and sampling ethanol tended to increase desire in these measures. However, at each dose, the induction of feelings of desire for ethanol showed a negative correlation with discrimination performance. These findings are discussed in the context of the ability of animals and humans to use several components of drug-induced stimuli in the performance of drug discrimination, and the role of such discriminative stimuli in priming of ethanol drinking. PMID- 10515307 TI - Hormones of the hypothalamo-pituitary-gonadal axis in drug discrimination learning. AB - More than 30 years ago, T-maze studies with progesterone indicated that sex hormones have the potential to act as a discriminative stimulus in rats. Despite these early positive findings, the interest in discriminative stimulus properties of sex hormones remained low; few studies were dedicated to the investigation of discriminative stimulus properties of hypothalamo-pituitary-gonadal axis hormones (i.e., LHRH, LH/FSH, sex steroids). Nevertheless, the few studies that were published showed some interesting, and often sex-dependent results. Applying various methodologies (T-, or Y-maze, two-lever drug discrimination, taste aversion procedures), it was found that not only progesterone but also the two other principal sex steroids estradiol and testosterone can serve as discriminative stimuli in rodents. In addition to these gonadal hormones, the hypothalamic peptide LHRH (having a key role in the neuroendocrine regulation of steroid release from the gonads) appears to generate discriminative stimulus properties. Interestingly, recent (but preliminary) studies in postmenopausal women suggest that estradiol (and possibly progesterone) may also function as a discriminative stimulus in human subjects. PMID- 10515308 TI - Effects of training dose on the relationship between discriminative-stimulus and self-reported drug effects of d-amphetamine in humans. AB - The aim of the present experiment was to examine the relationship between the discriminative-stimulus and self-reported effects of drugs in humans. To accomplish this aim, nine healthy adult volunteers (four females, five males) were trained to discriminate between placebo and 10 mg d-amphetamine (low-dose group) or 20 mg d-amphetamine (high-dose group). After acquiring the placebo amphetamine discrimination, a range of doses of d-amphetamine (1.25-20 mg) was tested to determine if they shared discriminative stimulus effects with the training dose. Participants in the low-dose group exhibited a significant leftward shift in the dose-response function for discrimination performance, which is concordant with previous preclinical and human drug discrimination studies that assessed the effects of training dose. Consistent with the drug discrimination findings, participants in the low-dose group exhibited a significant leftward shift in the dose-response function for several self reported drug effects (e.g., Like the Drug and Stimulated). However, several other self-reported drug effect items were not significantly influenced by training condition (e.g., Anxious/Nervous and Bad Effects). These results suggest that the discriminative-stimulus and self-reported drug effects of d-amphetamine overlap, but are not isomorphic. Furthermore, these results illustrate that behavioral history significantly influences subsequent drug effects in humans. PMID- 10515309 TI - Pharmacological and environmental determinants of relapse to cocaine-seeking behavior. AB - Animal models have been developed that simulate relevant features of relapse to cocaine-seeking behavior in humans. These models have provided valuable information about pharmacological and environmental factors that precipitate reinstatement of extinguished cocaine-seeking in rats and monkeys, as well as new insights about potential pharmacotherapies for relapse prevention. Reinstatement of cocaine-seeking behavior in animals can be induced by cocaine priming or by cocaine-paired environmental stimuli: however, maximum reinstatement of drug seeking appears to be induced when cocaine priming and cocaine-paired stimuli are combined. Drugs that share cocaine's indirect dopamine agonist properties or that act as direct agonists at D2-like dopamine receptors also induce reinstatement of cocaine-seeking behavior, whereas with some exceptions (e.g., caffeine, morphine) drugs from other pharmacological classes do not. D1-like receptor agonists block rather than mimic the priming effects of cocaine, suggesting different roles for D1- and D2-like receptor mechanisms in cocaine relapse. Although considerable overlap exists, drugs that exhibit cocaine-like discriminative stimulus and/ or reinforcing effects in other situations do not invariably induce cocaine-like reinstatement of drug-seeking and vice versa, implying that these effects are not simply different behavioral expressions of a unitary neurobiological process. Finally, recent findings with D1-like receptor agonists, partial agonists, and antagonists suggest that some of these drugs may be viable candidates for development as antirelapse pharmacotherapies. PMID- 10515310 TI - Drug discrimination in neurobiology. AB - Areas of neurobiological interest are identified towards which drug discrimination (DD) studies have made important contributions. DD allows ligand actions to be analyzed at the whole organism level, with a neurobiological specificity that is exquisite and often unrivalled. DD analyses have thus been made of a vast array of CNS agents acting on receptors, enzymes, or ion channels, including most drugs of abuse. DD uniquely offers access to the study of subjective drug effects in animals, using a methodology that also is transposable to humans and has generated unprecedented models of pathology (e.g., chronic pain, opiate addiction). Parametric studies of such independent variables as training dose and reinforcement provide refined insights into the dynamic psychophysiological mechanisms of both drug effects and behavior. Three different mechanisms have been identified by which discriminative, and perhaps other behaviors, can come about. DD also is superbly sensitive to small, partial activation of molecular substrates; this has enabled DD analyses to pioneer the unravelling of molecular mechanisms of drug action (attributing, f.ex., LSD's particular subjective effects to an unusual, partial activation of 5-HT, and perhaps other receptors). DD has both oriented and served as a tool to conduct drug discovery research (e.g., pirenperone-risperidone, loperamide). The DD response arguably constitutes a quantal, rather than graded, variable, and as such allows a comprehension of molecular, pharmacological, and behavioral mechanisms that would have been otherwise inaccessible. Perhaps most important are the following further contributions. One is the notion that particular, different levels of receptor activation are associated with qualities of neurobiological actions that also differ and are unique, this notion arguably constituting the most significant addition to affinity and intrinsic activity since the earliest theoretical conceptions of molecular pharmacology. Another contribution consists of studies that render redundant the notion of tolerance and identify fundamental mechanisms of signal transduction; these mechanisms account for apparent tolerance, dependence, addiction, and sensitization, and appear to operate ubiquitously in a bewildering array of biological systems. PMID- 10515311 TI - Creation and first 20 years of the society for the stimulus properties of drugs (SSPD). AB - Clinical observations and novels in the 19th century recognized that memory of some events can be retrieved only under the influence of the same drug condition that was present during the event. This dissociative effect of drugs probably reflects the same drug effects that were later called the discriminative stimulus effects of drugs. The Society for Stimulus Properties of Drugs (SSPD) was founded in 1978 as a forum for communications and periodic meetings on this drug effect. During its early years many of its members were psychologists, but subsequent to that time the most frequent research application has been for the pharmacological purpose of identifying new drugs that have the same discriminative stimulus attributes as a prototype training drug. The majority of members have been in the United States, but several major international meetings have been in Europe. The methods used by the society's members involve both neuropharmacological and psychological processes, allowing them to make unique contributions to the study of both mind and brain. PMID- 10515312 TI - Increasing the selectivity of drug discrimination procedures. AB - In an attempt to increase the selectivity of drug discrimination, rats were trained to discriminate LSD (0.08 mg/kg) from a group of "other" compounds consisting of cocaine (10 mg/kg), pentobarbital (5 mg/kg), and saline. Acquisition of this LSD-other discrimination was rapid (31 days) in chambers equipped with retractable levers and did not differ significantly from that of a group of animals trained to discriminate LSD from saline (26 days). In substitution (generalization) tests, hallucinogens such as LSD, DMT, and DOM mimicked LSD in a dose-dependent manner in both groups. The designer drug (+/ )MDMA substituted for LSD in the LSD-other group (ED50 = 1.38) but did not substitute for the training drug in the LSD-ND group; neither (+) MDMA nor PCP mimicked LSD in either group. Most importantly, lisuride, quipazine, and yohimbine, drugs that have been described as "false positives," substituted for LSD in animals trained to discriminate LSD from saline (ED50s = 0.012, 1.662, 2.344, respectively), but not in animals trained to discriminate LSD from other drugs. Thus, the LSD-other training procedure can be described as more selective than the standard drug-ND procedure. PMID- 10515313 TI - Comparison of antipsychotic activity and discriminative stimulus effects of the novel acylprolyltyrosine containing compound, GZR-123, and sulpiride. AB - The present experiment has been performed to determine the pharmacological profile of a newly synthesized systematically active prolyltyrosine containing compound, caproyl-L-Pro-L-Tyr methyl ester (GZR-123), and to compare it with that of the standard atypical benzamide neuroleptic, sulpiride. GZR-123 demonstrated antagonistic activity on apomorphine-induced climbing and on L-DOPA-dependent extrapolatory behavior in dose ranging between 0.4-4.0 mg/kg i.p.. It did not provoke a cataleptogenic effect or lethality, even at doses much higher than those causing antidopamine effects (more than 500 mg/kg). The effective doses of sulpiride in the above-listed antidopamine tests were shown to be 17.5 and 16.0 mg/kg i.p. correspondingly. Although these doses of sulpiride did not demonstrate cataleptogenic effects, an increase of the dose level to 120 mg/kg induced pronounced catalepsy. Both GZR-123 (6 mg/kg) and sulpiride (40 and 60 mg/kg) were investigated by training rats to discriminate each of them from saline in a two lever, water-reinforcement operant procedure. Both GZR-123 and sulpiride demonstrated weak discriminability in this task. GZR-123 increased drug associated lever selection when administered in doses of 2 and 6 mg/kg, for sulpirirde these doses were demonstrated to be 25-60 mg/kg. In contrast to GZR 123, which did not provoke a sedative effect, sulpiride in higher discriminable doses caused sedation, which stems probably from the motivational, but not from the motor deficit. PMID- 10515314 TI - Contribution of GABA(A) and GABA(B) receptors to the discriminative stimulus produced by gamma-hydroxybutyric acid. AB - The present study examined the involvement of GABA(A) and GABA(B) receptors in the discriminative stimulus effects of gamma-hydroxybutyric acid (GHB). Rats were trained to discriminate either 300 or 700 mg/kg GHB IG from water using a T-maze, food-reinforced drug-discrimination procedure. The direct GABA(B) agonist, baclofen, substituted completely for both training doses of GHB; its potency to substitute for GHB increased moderately as the training dose of GHB was increased. The positive GABA(A) modulator, diazepam, substituted partially for 300 mg/kg GHB, but failed to elicit GHB-appropriate responding in rats trained with the higher GHB dose. Finally, the GABA(B) antagonist, CGP 35348, completely blocked the discriminative stimulus effects of the high training dose of GHB, but only partially antagonized the effects of the low training dose. These results suggest that (a) GHB produces a compound stimulus, and (b) both GABA(B)- and GABA(A)-mediated cues are prominent components of this compound stimulus; the contribution of each component, however, appears to vary as the training dose of GHB is increased. PMID- 10515315 TI - Effects of a novel fentanyl derivative on drug discrimination and learning in rhesus monkeys. AB - Three monkeys discriminated 1.78 mg/kg of mirfentanil while responding under a fixed-ratio 5 schedule of stimulus-shock termination. Two mirfentanil derivatives, OHM3295 and OHM10579, substituted for mirfentanil in all subjects. However, other drugs produced variable effects among monkeys; for example, mu and kappa opioid agonists and clonidine substituted for mirfentanil on some occasions in two monkeys. Cocaine, amphetamine, and ketamine did not substitute in any subject. Opioid antagonists did not attenuate the effects of mirfentanil. In monkeys responding under a repeated acquisition and performance procedure, errors increased only during the acquisition phase at doses of mirfentanil that decreased response rates. Thus, unlike fentanyl, the discriminative stimulus effects of mirfentanil do not appear to be mediated exclusively through opioid receptors. Finally, mirfentanil does not appear to disrupt complex behavioral processes. PMID- 10515317 TI - The influence of menstrual cycle phase on sensitivity to ethanol-like discriminative stimulus effects of GABA(A)-positive modulators. AB - Previous studies showed that sensitivity to the ethanol-like discriminative stimulus effects of allopregnanolone and ethanol are enhanced during the luteal phase of the menstrual cycle when progesterone levels peak in monkeys trained to discriminate 1.0 g/kg ethanol. The present study further explored the influence of the menstrual cycle phase on the discriminative stimulus effects of ethanol, allopregnanolone, and midazolam. Female adult cynomolgus monkeys (Macaca fascicularis) were trained to discriminate 1.0 g/kg ethanol (n = 3) or 2.0 g/kg ethanol (n = 4) (20% w/v; i.g.) from water (i.g.). A cumulative dosing procedure was used to test discriminative stimulus effects of ethanol (0.5-2.5 g/kg; i.g.) and the ethanol-like discriminative stimulus effects of allopregnanolone (0.1-1.0 mg/kg; i.v.) or midazolam (1.0-17 mg/kg; i.g.) during the follicular vs. luteal phase of the menstrual cycle. In the 2.0-g/kg group, sensitivity to the ethanol like effects of allopregnanolone was increased during the luteal vs. follicular phase in two of three monkeys. In contrast, average sensitivity to ethanol was not different in the luteal compared to the follicular phase in the 2.0-g/kg group. Finally, there was no difference in sensitivity to midazolam between the follicular and luteal phases in monkeys trained with either 2.0 g/kg or 1.0 g/kg ethanol. Overall, the ethanol-like discriminative stimulus effects of midazolam are not sensitive to the menstrual cycle phase. In addition, there was less influence of the menstrual cycle phase on allopregnanolone and ethanol sensitivity in a 2.0-g/kg compared to a 1.0-g/kg ethanol training dose. PMID- 10515318 TI - Discriminative stimulus properties of mCPP and alprazolam are not mediated by anxiety. AB - We investigated whether the interoceptive cues mediated by the anxiolytic benzodiazepine receptor agonist alprazolam and the anxiogenic serotonin (5 HT)(1B/2C) receptor agonist 1-(3-chlorophenyl)piperazine (mCPP) in rats are related to anxiety. mCPP-induced anxiety in humans can be blocked with alprazolam, and if mCPP drug discrimination is to be used as a model of anxiety, mCPP's stimulus should be blocked by alprazolam. Therefore, two groups of rats were trained to discriminate either alprazolam (2 mg/kg, p.o.) or mCPP (2 mg/kg, p.o.) from vehicle in a two-level operant drug discrimination procedure. Cross antagonism tests were performed with alprazolam and mCPP. mCPP did not antagonize alprazolam's stimulus to any extent, but disrupted responding severely. Low and intermediate doses of alprazolam (1.0-4.0 mg/kg, p.o.) did not antagonize the mCPP discriminative stimulus; only a high dose of 8.0 mg/kg (p.o.) partially antagonized mCPP but disrupted responding in most of the animals. We conclude that, at best, there is only weak evidence to suggest that the interoceptive cues of alprazolam and mCPP are mediated by modulation of anxiety processes, and that the mCPP drug discrimination as a model for anxiety is unreliable. PMID- 10515319 TI - The 5-HT1A receptor agonist flesinoxan shares discriminative stimulus properties with some 5-HT2 receptor antagonists. AB - Ten homing pigeons were trained to discriminate the selective 5-HT1A receptor agonist flesinoxan (0.25 mg/kg p.o.) from its vehicle in a fixed-ratio (FR) 30 two-key operant drug discrimination procedure. The 5-HT2 receptor antagonist mianserin (ED50 = 4.8 mg/kg) fully substituted for flesinoxan, whereas ketanserin, ritanserin, mesulergine, and SB200646A substituted only partially, suggesting an interaction between 5-HT1A and 5-HT2 receptors. However, the 5-HT2 receptor agonists [DOI (0.6 mg/kg), TFMPP (10 mg/kg), mCPP (4 mg/kg)] were unable to antagonize the flesinoxan cue. The 5-HT1A receptor antagonists DU125530 (0.5 13 mg/kg) and WAY100,635 (0.1-1 mg/kg) partially antagonized the generalization of mianserin to flesinoxan. Taken together, these results are in accordance with the hypothesis that 5-HT1A receptor activation exerts an inhibitory effect on activation of 5-HT2 receptors. These results are in broad agreement with existing theories on 5-HT1A and 5-HT2 receptor interaction. Furthermore, it is argued that the discriminative stimulus properties of a drug may undergo qualitative changes with prolonged training. PMID- 10515316 TI - Analysis of D2 and D3 receptor-selective ligands in rats trained to discriminate cocaine from saline. AB - This study examined the role of dopamine D3 receptors in the stimulus generalization produced by 7-OH-DPAT and PD 128907 in rats trained to discriminate cocaine from saline. Twelve male Sprague-Dawley rats were trained to discriminate cocaine (10 mg/kg) from saline in a two-choice operant procedure using a FR20 schedule of water reinforcement. Stimulus generalization tests were administered with the D3-preferring agonists (+/-)-7-OH-DPAT (0.01-0.3 mg/kg), (+)-7-OH-DPAT (0.01-0.3 mg/kg), and PD 128907 (0.01-0.3 mg/kg), and the selective D2 agonist PNU-39156 (0.01-0.3 mg/kg). Complete generalization to cocaine was observed with (+/-)-7-OH-DPAT at doses that markedly suppressed response rate. Only partial stimulus generalization was observed with (+)-7-OH-DPAT and PD 128907 when these compounds were administered intraperitoneally, although subcutaneous injections of these compounds produced complete substitution. Response rate was also significantly reduced by these compounds. The selective D2 agonist, PNU-91356 also fully substituted for the cocaine cue and suppressed response rate in a dose-dependent manner. To ascertain the importance of D3 receptor actions in the stimulus generalization produced by (+/-)-7-OH-DPAT (0.1 mg/kg) and PD-128907 (0.3 mg/kg), the fairly selective D3 antagonist, PNU-99194A (2.5-20 mg/kg) was also tested in combination with these compounds. Although PNU 99194A partially attenuated the stimulus generalization produced by (+/-)-7-OH DPAT, it failed to block PD-128907 substitution for cocaine. These results indicate at least some involvement of D3 receptors in the stimulus effects of (+/ )-7-OH-DPAT, although further investigations are clearly warranted. The present results also suggest that the cue properties of cocaine may be dissociated from the locomotor activating effects of this drug, because D3/D2 receptor agonists suppress locomotor activity but produce stimulus generalization to cocaine. PMID- 10515320 TI - Discriminative effects of phenazepam and gidazepam in rats: comparison with other GABA-related drugs. AB - The present study assessed the discriminative stimulus effects of phenazepam (PHZ) (2 mg/kg, i.p.), gidazepam (GDZ) (10 mg/kg, i.p.), pentobarbital (PB) (10 mg/kg, i.p.), and buspirone (B) (5 mg/kg, i.p.) by testing GABA-related drugs in the two-lever liquid reinforced operant discrimination procedure in rats. Diazepam (5-30 mg/kg, i.p.) dose dependently and completely substituted in GDZ trained rats and in only 40% PHZ-trained rats. Following phenobarbital (40-100 mg/kg, i.p.) injections the mean percentages of PHZ- and GDZ-lever responding generally were a monotonically increasing function of dose, but peaked at 39.3 and 52.9%, respectively. The PB discriminative cue was generalized completely to PHZ, GDZ, and phenobarbital. Picrotoxin (2 mg/kg, s.c.) did not inhibit the PHZ and GDZ discriminations, while it antagonized the PB (10 mg/kg, i.p.) cue. Calcium valproate (200 mg/kg, i.p.) failed to produce PHZ effects, and partially substituted for GDZ. B failed to substitute for the discriminative effects of PHZ, GDZ, or PB, producing a maximum 9.3, 18.0, and 33.3% drug lever responding, respectively. These results suggest that the discriminative stimuli of PHZ and GDZ are similar to those of other benzodiazepine agonists. However, the PHZ cue is more selective than that of GDZ. PMID- 10515321 TI - Discriminated taste aversion and context: a progress report. AB - The research described here concerns the interaction between the environment (context), the organism, and the effects of opiates, focusing on how conditioning and contextual cues affect drug controlled behaviors. This analysis applies the powerful tool of drug discrimination to a respondent conditioning procedure (discriminated taste aversion, DTA). Data show that the use of DTA is feasible in that it is sensitive to morphine dose and saccharin concentration. Swifter control over DTA was achieved by increasing the LiCl dose (UCS magnitude). It is also clear that morphine alone can serve as a discriminative stimulus not requiring saccharin as a contextual element (which has been the case for most DTA studies to date), or saccharin being part of a compounded stimulus. Pharmacological specificity was demonstrated in substitution tests with delta-9 tetrahydrocannabinol. This research continues a systematic experimental analysis of the interaction between drug-controlled behavior and context. PMID- 10515322 TI - Influence of training paradigm on specificity of drug mixture discriminations. AB - Generalization to different drugs and drug mixtures has been examined in rats trained to discriminate a mixture of amphetamine (0.4 mg/kg) plus pentobarbitone (10 mg/kg) from saline (AND discrimination, n = 8) or to discriminate the same mixture from its component drugs alone (AND-OR discrimination, n = 9). The studies used two-lever operant procedures with a tandem variable interval 1-min fixed-ratio 10 schedule of food reinforcement. There was partial generalization to nicotine and midazolam and no generalization to cocaine, caffeine, or ethanol under AND-discrimination conditions and no generalization to any of these drugs in the AND-OR discrimination. Nicotine or midazolam coadministered with the training doses of pentobarbitone and amphetamine, respectively, produced full generalization in the AND discrimination and partial generalization under AND-OR conditions. Cocaine coadministered with pentobarbitone generalized fully under both procedures, but at larger doses in the AND-OR than in the AND discrimination. Mixtures of either nicotine plus midazolam or caffeine plus ethanol produced very marked generalization under AND-discrimination conditions, but were without significant effect in the AND-OR procedure. The results consistently supported the hypothesis that the AND-OR discrimination procedure increases the specificity of discriminations based on drug mixtures. PMID- 10515323 TI - Differences between alcohol-preferring and alcohol-nonpreferring rats in ethanol generalization. AB - Alcohol-preferring (P rats) and alcohol-nonpreferring rats (NP rats) were trained to discriminate intraperitoneal injections of 0.5 g/kg ethanol, or subcutaneous injections of 0.6 mg/kg nicotine from saline. P rats learned the ethanol discrimination more rapidly and made a higher percentage (88%) of their responses on the ethanol lever after ethanol and a lower percentage (7%) after saline than NP rats (78 and 15%, respectively). In substitution tests, increasing doses of ethanol produced increases in the percentage of responses on the ethanol lever with similar ED50s (0.43 and 0.44 g/kg) in P and NP rats. P rats trained to discriminate ethanol from saline made more responses on the ethanol lever after nicotine (80%) and d-amphetamine (63%) than NP rats (33 and 40%). The ethanol stimulus did not generalize to morphine in either P or NP rats. NP rats trained to discriminate ethanol from saline responded more on the ethanol lever after bupropion (77%) than P rats (49%). In rats trained to discriminate nicotine from saline, the nicotine discriminative stimulus did not generalize to ethanol in either P or NP rats, suggesting that the genetic difference in the stimulus generalization of ethanol was not symmetrical. PMID- 10515324 TI - Discriminative stimulus properties of eltoprazine in the pigeon. AB - Twelve pigeons were successfully (ED50 = 2.4 mg/kg p.o.) trained to discriminate the 5-HT(1A/B) receptor agonist eltoprazine (5.0 mg/kg p.o.) from its vehicle in a fixed-ratio (FR)30 two-key operant drug discrimination procedure. Tests for generalization and antagonism showed that 5-HT1A receptor agonists, such as 8-OH DPAT (8-hydroxy-2-(di-n-propylamino)tetralin) (66.7%), flesinoxan (72.7%), buspirone (58.3%), and ipsapirone (36.4%) only partially substituted for the eltoprazine cue. Compounds with mixed agonistic action at 5-HT1 receptors, completely (> or = 80%) [(eltoprazine; TFMPP (1-(3-trifluoromethylphenyl) piperazine (ED50 = 7.68 mg/kg) and RU 24969 (5-methoxy-3-(1,2,3,6 tetrahydropyridin-4-yl-1H-indole) (ED50 = 15.8 mg/kg)] substituted for eltoprazine; whereas m-CPP (1-(3-chlorophenyl)piperazine) did not. The selective 5-HT reuptake inhibitor fluvoxamine partially (44%) substituted for the eltoprazine cue. The 5-HT1A receptor antagonist NAN-190 (1-(2-methoxyphenyl)-4-[4 (2-phtalimido)butyl]piperazine) fully blocked the eltoprazine cue. Both (+/-) pindolol and (+/-)-propranolol showed partial antagonism of the eltoprazine cue (66.7 and 50.0%, respectively). (+/-)-Pindolol also showed partial substitution (50%) for the eltoprazine cue, but NAN-190 and (+/-)propranolol did not. It is concluded that the discriminatory stimulus properties of eltoprazine in the pigeon are mediated by 5-HT1A and 5-HT1B receptors. PMID- 10515325 TI - Does MK-801 discrimination constitute an animal model of schizophrenia useful for detecting atypical antipsychotics? AB - Two groups of female Wistar rats were trained to discriminate two doses (0.075 and 0.0375 mg/kg) of the noncompetitive NMDA antagonist MK-801 (dizocilpine) in a food-rewarded operant FR30 drug discrimination task. The atypical neuroleptic clozapine (2-6 mg/kg) produced only minimal antagonism (max. 32%) of the MK-801 cue at either training dose, and the "antagonist" effects were not clearly dose related. Furthermore, in the 0.075 mg/kg trained animals clozapine at 3 mg/kg failed to shift the MK-801 dose-response curve to the right. The alpha1 adrenoceptor antagonist prazosin (1-8 mg/kg) was also tested for antagonism of the 0.0375 mg/kg MK-801 cue, and again, only partial antagonism was seen (maximum 36%). Recently, it was suggested [4] that as the discriminative stimulus produced by MK-801 (0.075 mg/kg) was fully antagonized by clozapine at 3 mg/kg, but not by the typical neuroleptic haloperidol, this assay may be a useful screen for detecting atypical neuroleptics. It would seem, however, that this is not necessarily the case, and that the MK-801 discriminative cue may not be psychotomimetic. However, as this was a food rewarded rather than an avoidance paradigm that was used in the prior study [4], it may be that the drug discrimination procedure itself is a critical factor, although this hypothesis requires empirical testing. PMID- 10515326 TI - Inability of antipsychotics to antagonize the cueing properties of cocaine in rats. AB - In this study the possible antagonistic effects of five different antipsychotics on the discriminative stimulus properties of 10 mg/kg cocaine were evaluated by use of a two-lever food-reinforced drug discrimination procedure in rats. To do so, rats were treated with several doses of haloperidol, risperidone, seroquel, sertindole, and olanzapine, either at 60 or 120 min prior to testing. With all compounds tested, no substantial antagonism of the cocaine cue was observed. Only with haloperidol (maximum 60%), risperidone (maximal 20%), and olanzapine (maximal 20%) a partial antagonism without clearcut dose-response was observed. Clozapine, seroquel, and sertindole did not influence the discriminative stimulus properties of cocaine. These results indicate that antipsychotics with different pharmacological profiles are unable to antagonize more than partially the cueing properties of 10 mg/kg cocaine in rats, pointing to the unique underlying stimulus properties of this stimulant. PMID- 10515327 TI - GTS-21, a mixed nicotinic receptor agonist/antagonist, does not affect the nicotine cue. AB - Identification of nicotinic receptor subtypes involved in nicotine dependence is required for guiding the design of more selective antagonists capable of blocking the nicotine cue and nicotine self-administration. Due to the multiplicity of nicotinic receptors in the mammalian brain, selective agonists and antagonists are needed to assess the functional involvement of a particular subtype in vivo. Only recently have a few nicotinic receptor subtype-selective antagonists and agonists been identified. GTS-21 (also known as DMBX-anabaseine) is the only agent so far reported that selectively stimulates the alpha7 nicotinic receptor. Here GTS-21 was used to assess the possible mediation of the nicotine cue by this receptor subtype. Long-Evans rats were trained to discriminate between presession administration of 0.10 or 0.40 mg/kg (-)-nicotine bitartrate and its vehicle. GTS 21 did not substitute for nicotine, as all subjects consistently chose the vehicle lever after GTS-21 substitution. In another experiment, different doses of GTS-21 were administered prior to nicotine administration to investigate whether GTS-21 would antagonize the nicotine cue. Such was not the case. The lack of effect of GTS-21 upon the nicotine cue is consistent with the notion that the cue is mediated by nicotinic receptors other than the alpha7 receptor. PMID- 10515328 TI - Discriminative stimulus effects of nalbuphine in nontreated and morphine-treated pigeons. AB - In the present study, the stimulus effects of the low efficacy agonist nalbuphine were examined under two conditions: nontreated and morphine treated. In the first experiment, five pigeons were trained to discriminate among 3.2 mg/kg morphine, 5.6 mg/kg nalbuphine, and saline. Nalbuphine produced nalbuphine-like responding. Low doses of morphine produced nalbuphine-like responding, whereas high doses produced morphine-like responding. Naltrexone produced saline-like responding and reversed the stimulus effects produced by the training doses of morphine and nalbuphine. Five different pigeons were treated daily with 10 mg/kg morphine (i.m.) and trained 6 h later to discriminate among 10 mg/kg morphine, 1.0 mg/kg nalbuphine and saline. In these pigeons, morphine produced morphine-like responding and nalbuphine produced nalbuphine-like responding. Morphine abstinence produced nalbuphine-like responding that was reversed by morphine. Additionally, naltrexone produced nalbuphine-like responding. These data suggest that the discrimination between morphine and nalbuphine in the nontreated and morphine-treated pigeons may be based on the relative efficacy differences between morphine, a higher efficacy mu-agonist, and nalbuphine a lower efficacy mu-agonist. PMID- 10515329 TI - Stimulus properties of PMMA: effect of optical isomers and conformational restriction. AB - para-Methoxymethamphetamine (PMMA), a structural hybrid of two central stimulants, lacks stimulant properties but behaves in a manner similar to that of MDMA [N-methyl-1-(3,4-methylenedioxyphenyl)-2-aminopropane]. PMMA has been established as a training drug in drug discrimination studies, and in the present investigation we sought to determine which optical isomer of PMMA is primarily responsible for its stimulus effects. Because PMMA is a conformationally flexible molecule, it was also of interest to determine what conformation is most important for its actions. Accordingly, we prepared and examined S(+)PMMA, R( )PMMA, and conformationally restricted forms of PMMA: PMMA-AT, TIQ-1, and TIQ-2. S(+)PMMA (ED50 = 0.32 mg/kg) was found to be at least as potent as PMMA (ED50 = 0.41 mg/kg), whereas R(-)PMMA failed to result in complete stimulus generalization. An aminotetralin-like conformation, as found in PMMA-AT (ED50 = 0.29 mg/kg), seems to better account for the actions of PMMA than a tetrahydroisoquinoline-like conformation because TIQ-1 and TIQ-2 failed to result in stimulus generalization. The results of the present study further support the concept that PMMA and MDMA share considerable similarity with respect to their stimulus properties in animals except that PMMA lacks the amphetaminergic stimulant component of action associated with MDMA. PMID- 10515330 TI - Adjuvant therapy for rectal cancer can no longer be ignored. PMID- 10515331 TI - Pre-operative chemoradiotherapy and total mesorectal excision surgery of rectal cancer: towards the eradication of pelvic failures? PMID- 10515333 TI - Pre-operative localization and tissue uptake study in parathyroid imaging with technetium-99m-sestamibi. AB - BACKGROUND: The diagnostic ability of 99mTc-sestamibi was compared with other techniques and the mechanism of parathyroid uptake was investigated. METHODS: Double-phase 99mTc-sestamibi scanning was performed in 52 primary and 28 renal hyperparathyroidism patients. Gene expressions of mdr1 and mrp were examined by reverse transcriptase polymerase chain reaction in parathyroid tissue. RESULTS: The sensitivity of 99Tc-sestamibi in primary and renal hyperparathyroidism was 91% and 75%, respectively, higher than for ultrasonography, T1/Tc subtraction scintigraphy, or computed tomography. Early 99mTc-sestamibi uptake was washed out in delayed images in 7% and 30% of glands in primary and renal hyperparathyroidism, respectively. Expression of mdr1 and mrp mRNA was found in 5 of 23 and 16 of 31 glands, respectively, and their expression correlated with washout in delayed images. CONCLUSION: 99mTc-sestamibi was the best localization test. mdr1 and mrp were associated with 99Tc-sestamibi washout, but their role in the parathyroid remains unclear. PMID- 10515332 TI - Application of prognostic scoring systems in differentiated thyroid carcinoma. AB - BACKGROUND: Total thyroidectomy is widely practised in Australasia for papillary and follicular thyroid carcinoma. Data from large overseas series have demonstrated that patients with these cancers may be separated into risk groups based on clinicopathological prognostic factors. Furthermore, evidence suggests that low-risk patients may be safely treated with less than total thyroidectomy. The aim of the present paper was to determine what proportion of our patients with papillary and follicular thyroid cancer were in the low-risk group in order to select candidates for less aggressive treatment. METHODS: A prospectively documented series of 175 previously untreated patients with papillary and follicular thyroid carcinoma, treated principally by total thyroidectomy over a 10-year period, was divided into risk groups using the Mayo Clinic, Lahey Clinic and Memorial Hospital prognostic scoring systems. Complication rates for 103 patients treated by total thyroidectomy were also studied and reported. RESULTS: Women outnumbered men by 2.3:1. There were 128 papillary carcinomas (73%) and 47 follicular cancers (27%). These tumours were < 4 cm in diameter in 81% of patients, and 41% of patients were 40 years of age or younger. Low-risk patients accounted for 75, 81 and 45% of the study group, respectively, when the three prognostic scoring systems were applied to our patient population. The rates for recurrent laryngeal nerve palsy and permanent hypoparathyroidism for patients having total thyroidectomy were 1 and 1.9%, respectively. In the low-risk group there were no permanent complications. CONCLUSION: Most patients treated at Royal Prince Alfred Hospital during the past 10 years were low-risk patients who may have been eligible for less aggressive surgical treatment. PMID- 10515334 TI - Voice changes and thyroid surgery: is pre-operative indirect laryngoscopy necessary? AB - BACKGROUND: Indirect laryngoscopy (IDL) is often performed prior to thyroid surgery to detect pre-existing vocal cord pathology. METHODS: A retrospective chart review of 201 patients undergoing thyroid surgery at the Prince of Wales Hospital was undertaken in order to study the patterns of pre-operative and postoperative voice changes and IDL findings. RESULTS: A total of 9% of patients had pre-operative voice symptoms, and 22% of this group had abnormalities detected on pre-operative IDL. Of 160 documented IDL, 4% revealed vocal cord pathology in asymptomatic patients, including an asymptomatic recurrent laryngeal nerve palsy. CONCLUSIONS: Indirect laryngoscopy remains a useful but flawed pre operative screening tool for patients with voice symptoms, but the literature suggests that more advanced phoniatric tests will provide superior diagnostic sensitivity. The role of routine pre-operative laryngoscopy for asymptomatic patients is of debatable value. PMID- 10515335 TI - Cystosarcoma phyllodes: the Western Australian experience. AB - BACKGROUND: Cystosarcoma phyllodes is a rare breast tumour whose behaviour is not well understood by many clinicians. METHODS: In 1998 a retrospective study was undertaken of women diagnosed with phyllodes tumour of the breast who had their initial surgery between 1983 and 1994 in Western Australian public hospitals. RESULTS: Forty women were diagnosed and treated over this period; however, only 28 received ongoing follow-up (70%). Follow-up was obtained on 26 of these. The median age at diagnosis in this group was 46 years. Cases were predominantly Caucasian (85.5%). Postmenopausal women were affected in 26.9% of cases. Four patients had recurrences after surgery (one malignant and three benign). No patient had a recurrence with primary tumours with a diameter of < or = 2 cm. Mean time to recurrence was 35.8 months. CONCLUSIONS: Breast-conserving surgery was used in almost all cases (96.2% of first operations). PMID- 10515336 TI - Predictors of breast cancer in women recalled following screening. AB - BACKGROUND: Established risk factors are associated with between 25 and 56% of breast cancer cases, but the relative importance and relevance to different age groups is unclear. METHODS: This case-control study examines established risk factors in 298 women with breast cancer and 1926 women without breast cancer aged 40-87 who were recalled for assessment following routine mammography. RESULTS: The cancer group were significantly older than the non-cancer group (F1,222 = 107.6; P < 0.0001). Postmenopausal obesity increased the odds of developing breast cancer (OR: 2.35; CI: 1.33-4.16). The breast cancer group were more likely to have used oral contraceptives (OR: 1.50; CI: 1.09-2.05), and women who used contraceptives for more than 10 years in total were at the highest risk (OR: 1.73; CI: 1.13-2.65). Daily consumption of alcohol was also associated with increased risk of developing breast cancer (OR: 1.62; CI: 1.13-2.33). Reproductive factors and a family history of breast cancer did not affect the odds of developing breast cancer and the reasons for these findings are explored. CONCLUSIONS: Results suggest that the effects of weight reduction in reducing postmenopausal breast cancer risk should be assessed. PMID- 10515337 TI - Nitric oxide biosynthesis and malondialdehyde levels in advanced breast cancer. AB - BACKGROUND: Nitric oxide (NO) is a free oxygen radical studied in many tissues. Its tumour killing structure is shown especially by macrophages. The end products of NO are nitrite and nitrate. Their plasma levels are used biochemically to determine nitric oxide synthase (NOS) activity. The proliferative capacity of cancer cells accompanies the alteration in oxidant-anti-oxidant status. The risk of breast cancer is decreased in association with an increased level of polyunsaturated fatty acids in the erythrocyte membranes. The more the anti oxidant capacity increases, the more the transformed cells grow. Malondialdehyde (MDA) is a lipid peroxidation marker, and low plasma levels of MDA are associated with advanced stages of breast cancer. METHODS: In the present study, the alteration of serum plasma levels of nitrate, nitrite and MDA were determined in patients with stage IIIB breast cancer and controls. RESULTS: It was found that products of NO biosynthesis were higher and plasma MDA levels were lower in patients with breast cancer. CONCLUSIONS: It can be stated that in advanced breast cancer, the NO radical production is increased while the lipid metabolism is altered, and these changes can be related to an alteration in oxidant-anti oxidant status. PMID- 10515338 TI - The cost of elective and emergency repair of AAA in patients under and over the age of 80. AB - BACKGROUND: As Australia's population ages, the number of elderly patients presenting for surgery of abdominal aortic aneurysms (AAA), both elective and ruptured, will increase. The aim of the present study was to compare the costs of treatment of patients with AAA, under and over the age of 80, in the elective and emergency settings in a hospital with a divisional structure in which the true costs can be accurately obtained. METHODS: A total of 40 patients were selected at random from a series of 267 patients treated with open surgery for AAA between January 1987 and December 1994, 10 in each of four groups: group A, elective repair in patients aged < 80 (171/267); group B, elective AAA repair in patients aged > 80 (25/267); group C, emergency AAA repair in patients aged < 80 (50/267); and group D, emergency AAA repair in patients aged > 80 (11/267). A retrospective analysis of the hospital costs of treatment of these patients at St George Hospital was conducted. These true costs were then compared to Australian National Diagnostic Related Group (AN-DRG) costs. RESULTS: Group A and B had no mortality. In Group C and D the mortality was 20 and 60%, respectively. The emergency treatment groups also had longer lengths of stay. A statistically significant difference in cost of AAA repair between elective and emergency groups in both age groups was seen; that is, group A cost less than group C and group B cost less than group D. Costs per survivor, however, showed a dramatic difference between the cost of group C patients ($30000) and group D patients ($60000). In comparison with AN-DRG calculated costs, the true costs of groups A and B were equivalent to AN-DRG costs. In the emergency groups, however, there were marked discrepancies between the true cost ($61000) and that calculated by the DRG ($25000) in group D, with similar differences seen in group C to a lesser extent. CONCLUSION: Emergency repair of AAA is significantly more expensive and has a high mortality in the over-80 age group. Also, there is a substantial shortfall between the true costs of treating these patients and the funds allocated for treatment in this group. PMID- 10515339 TI - Keloids and hypertrophic scars: results with intraoperative and serial postoperative corticosteroid injection therapy. AB - BACKGROUND: The management of keloids and hypertrophic scars continues to be controversial. Experience of treating 58 such lesions, 58.62% of which were recurrent, is presented. METHODS: Each lesion was subjected to surgical excision with intra-operative local injection of triamcinolone acetonide, followed by repeat injection of the same drug at weekly intervals for 2-5 weeks depending on the symptomatic relief, and then monthly injections for 4-6 months. RESULTS: Complete symptomatic relief was achieved in all patients within 5 weeks of surgery. Objective response in terms of no recurrence was noted in 91.9% of patients with keloids, and 95.24% of patients with hypertrophic scars at a mean follow-up of 30.5 months. Local or systemic complications were insignificant. CONCLUSION: Because of promising results, further use and evaluation of this method of treatment is recommended for large, recurrent and complicated keloids and hypertrophic scars. PMID- 10515340 TI - Phenoxybenzamine in the management of neuropathic bladder following spinal cord injury. AB - BACKGROUND: The present study aims to show the clinical and urodynamic effects of phenoxybenzamine on the neuropathic bladder of spinal cord-injured patients who failed to be free of catheter by attaining satisfactory voiding function, despite initial bladder training. METHODS: Forty-six spinal cord-injured patients were subjected to pharmacological manipulation with phenoxybenzamine. It was used as an adjunct in the management of neuropathic bladder dysfunction that caused failure of the bladder to empty, by tapping or crede to achieve satisfactory residual urine volume of < 100 mL. Phenoxybenzamine was started with a dose of 10 mg daily, increased by 10 mg every 3 days to a dose of 30 mg daily; this was maintained from 3 weeks to 6 months (mean: 39 days). The pre-treatment residual urine volume ranged between 100 and 1050 mL (mean: 360 mL). Follow-up periods ranged between 12 and 36 months (mean: 16 months). RESULTS: Five patients (11%) were excluded due to either inadequate treatment or inadequate follow-up. Nineteen patients (41%) with reflex (upper motor neurone) bladders showed improvement of bladder evacuation. There was a reduction of the maximum urethral closure pressure, which ranged between 10 and 32 cm of water (mean: 22 cm). Twenty-two patients (48%) did not respond, requiring other measures to be taken which included transurethral surgery (n = 19). Nine of the failures involved areflex (lower motor neurone) bladders, and seven failures involved reflex bladders with an extremely tight outlet and urethral closure pressure of > 50 cm of water. Six failures involved reflex bladders that were lacking strong enough detrusor contractions to attain a balanced bladder responsive to abdominal tapping; response was achieved by administration of a parasympatheticomimetic drug. Neuropathic bladders with uninhibited detrusor contractions responded well to phenoxybenzamine. CONCLUSIONS: Phenoxybenzamine proved useful in reducing bladder outlet resistance after spinal cord injury, provided that detrusor bladder contractions were present. It is useful in controlling detrusor-sphincter dyssynergia and autonomic hyperreflexia. It was not useful in areflex bladders, perhaps due to the development of spasticity of the striated muscle component of the external sphincter. The presence of bladder neck (internal sphincter) dysfunction may modify or abolish its effect. PMID- 10515341 TI - Computed tomography in the diagnosis of equivocal appendicitis. AB - BACKGROUND: The clinically obscure right iliac fossa (RIF) pain remains a diagnostic problem. The present study examines the use of computed tomography (CT) in improving the accuracy of clinical assessment in these difficult surgical cases. METHODS: The trial design was a retrospective review of all patients admitted under one surgeon with suspected acute appendicitis, between 1 January 1995 and 30 June 1997. The study setting was a district hospital (Calvary Hospital) that received patients from both an urban and rural environment. The patient cohort was identified from the Unit Registry and an International Classification of Diseases-based review of medical records. Twenty-one prospective data points were obtained from patient records. Those patients admitted with RIF pain and equivocal symptoms and signs subsequently underwent a CT and/or ultrasound (US) examination, conducted by the attending radiologist. For those patients who proceeded to appendicectomy, the histopathological findings were correlated with the imaging report. Those patients who were discharged after imaging without proceeding to operation were not readmitted to any regional hospital during the course of the study. RESULTS: A total of 84 patients were identified. Thirty-three patients (39%) underwent appendicectomy without imaging and were excluded from further analysis. A total of 51 patients (61%) underwent 61 imaging procedures. The CT scan was correct in 35/36 patients (97%), while US was correct in 17/25 patients (68%). CONCLUSIONS: The present study suggests that CT can be used to improve the accuracy of diagnosis of obscure RIF pain. As a pilot study, it supports the development of a randomized controlled trial in a multicentre regional study. PMID- 10515343 TI - Deriving opera from operation. AB - Male castration has been practised for centuries in many parts of the world. In Italy in the 16th to the 19th centuries it was performed for the purpose of preserving the soprano voice of boys. The castrati performed in the church choirs, but in the field of opera they achieved the popularity and status of the modern day rock singers. An outline of the operative procedure is given with its physiological consequences, and mention is made of some of the singers who achieved fame at that time. Recently an attempt has been made to reproduce the sound of the castrato voice using the facilities of modern electronic technology. PMID- 10515342 TI - Self-expandable metal stents for malignant dysphagia. AB - BACKGROUND: The use of self-expandable metal stents in relieving dysphagia for patients with incurable malignant oesophageal strictures was retrospectively evaluated. METHODS: Between September 1993 and August 1996, 66 male and 16 female patients with a median age of 72 years received self-expandable metal stents for malignant dysphagia. Six patients had concurrent tracheo-oesophageal fistulas. All patients were stented under sedation and stent insertion was performed under fluoroscopic guidance. RESULTS: Stent placement was successful in 80 patients (98%). There were seven early complications (inaccurate positioning (n = 3), migration (n = 1), incomplete expansion (n = 1), intractable pain (n = 1), and perforation (n = 1)). Two complications were lethal and three were treated endoscopically. Mean dysphagia grade improved from 3.2+/-0.7 to 1.8+/-0.9 (P < 0.05) after implantation. All tracheo-esophageal fistulas were successfully occluded. Upon a median follow-up of 8 weeks (range: 2-20 weeks), 30 complications developed in 21 patients (tumour overgrowth (n = 15), food bolus obstruction (n = 7), tumour ingrowth (n = 2), buckling of stent (n = 2), tracheo esophageal fistula (n = 2), bleeding (n = 1), and gastric wall herniation through metal coils (n = 1)). Median survival was 13 weeks (range: 1-82 weeks). CONCLUSION: Self-expandable metal stents provide useful palliation in patients with incurable malignant dysphagia. PMID- 10515344 TI - How large should a skin trephine be for an end stoma? AB - BACKGROUND: Although much has been written about techniques for making a good stoma, little has been described regarding how to excise a trephine that will perfectly fit a given end stoma. METHODS: A reproducible technique for making a stomal trephine to a precise fit for each stoma is described. RESULTS: More than 20 stomas have been made with good results. CONCLUSION: The skin trephine should have a diameter approximately two-thirds of the width of the crushed bowel end. PMID- 10515345 TI - Barriers to participation in randomized clinical trials for early breast cancer among Australian cancer specialists: comment. PMID- 10515346 TI - Basingstoke: comment. PMID- 10515347 TI - Multiple laparotomies for severe intra-abdominal infection: comment. PMID- 10515349 TI - Neutropenic proctosigmoiditis complicating breast cancer chemotherapy. PMID- 10515348 TI - Neutropenic enterocolitis due to dipyrone use. PMID- 10515350 TI - Colonic intussusception in Crohn's disease. PMID- 10515351 TI - Spontaneous mid-trimester uterine rupture. PMID- 10515352 TI - Effect of divergent selection for total plasma phosphorus on plasma and yolk very low density lipoproteins and plasma concentrations of selected hormones in laying Japanese quail. AB - Japanese quail lines were divergently selected over 32 generations for laying hen plasma yolk precursor, as measured by total plasma phosphorus (TPP). The high (HP) and low (LP) lines were developed from a randombred control population (R1) that was maintained without conscious selection. The purpose of the present study was to characterize the composition of very low density lipoproteins (VLDL) in laying Japanese quail hens (VLDLy) and the concentration of selected hormones in laying hens from the HP, LP, and R1 lines. The changes in TPP because of genetic selection in the Japanese quail lines were associated with large alterations in plasma VLDLy concentration (HP > R1 > LP), but only minor changes in lipid composition and size (HP > LP = R1; P< or =0.01) of plasma VLDLy particles. Basal plasma levels of hormones associated with reproduction and lipid metabolism were also different among lines, with luteinizing hormone (LH) ranking HP >R1 = LP and triiodothyronine (T3), thyroxine (T4), and 17beta-estradiol ranking HP > R1 > LP (P< or =0.05). The results suggest possible increased rates of hepatic lipogenesis, hepatic VLDLy assembly and secretion, and plasma VLDLy concentration in association with increases in concentrations of plasma LH, T3, T4, and 17beta estradiol. Concentrations of total lipids in yolk VLDL were not different among lines, and only minor line differences in the concentration of different classes of yolk VLDL neutral lipids were detected. The data indicate a preferential uptake of a specific plasma VLDLy subpopulation into rapidly growing ovarian follicles, resulting in a constant composition of yolk VLDL of laid eggs among lines of Japanese quail with large differences in plasma VLDLy concentration. PMID- 10515353 TI - Heterosis following long-term bidirectional selection for mating frequency in male Japanese quail. AB - Reciprocal crosses (sire line shown first and dam line second) among high (H) and low (L) selected lines and the randombred control line (C), which was the base population for the selected lines, were made after 40 generations of bidirectional selection for mating frequency of male Japanese quail. Significant heterosis for the selected trait was found only in crosses between Lines C and L, being 62 and 92% for LC and CL, respectively. Heterosis for percentage of maters was present in all crosses, ranging from 8% for HC and CH to 46% for HL. Three (HC, LH, and CL) of the six crosses had significant heterosis for both 4- and 8 wk BW. Heterosis for 4- and 8-wk BW was also significant for the HL and CH crosses, respectively. For area of the cloacal gland, heterosis was significant in five crosses. Although crosses tended to exhibit higher relative aggressiveness than their respective midparent means, heterosis for this trait was not significant. Reciprocal effects, although not important for most traits, were present for BW in crosses between Lines C and L and Lines H and C. In general, long-term selection for mating frequency of males changed the genetic basis of selected and correlated traits with considerable nonadditive genetic effects observed for most traits in specific crosses. PMID- 10515354 TI - Factors affecting ostrich egg hatchability. AB - Ostrich eggs often have low hatchability (HATCH) rates because they do not lose sufficient weight during incubation. Because egg size, eggshell porosity and thickness (THICK), and length of preincubation egg storage are known to affect egg weight loss during incubation (EWL) and HATCH of chicken eggs, these factors were examined using ostrich eggs. The effects of eggshell porosity (number of large pores per cm2 of shell; LP); and THICK on EWL and HATCH were assessed by categorizing the eggs as having either low, intermediate, or high LP or low, intermediate, or high THICK. Mean EWL was higher (P<0.05) in eggs of the high LP group when compared with eggs in either the low or intermediate LP groups that lost similar amounts of weight during incubation. Mean HATCH was also higher (more than 25%; P<0.10) in eggs with high LP when compared with the HATCH found in eggs having low LP. Eggs from the intermediate LP group had an intermediate HATCH response. Moreover, numbers of LP were positively correlated to both EWL (r2 = 0.64; P<0.0001) and HATCH (r2 = 0.25; P<0.03). Inverse relationships existed between THICK and EWL and between THICK and HATCH according to the order (P< 0.05): eggs of low THICK, highest mean EWL and HATCH > eggs of intermediate THICK, intermediate mean EWL and HATCH > eggs of highest THICK, lowest mean EWL and HATCH. Shell thickness was not correlated to either EWL or HATCH. The influence of egg size on mean LP, THICK, EWL, HATCH, and chick weight (CWT) was assessed. Although THICK was unaffected by egg size, higher LP (P<0.10), EWL (P<0.05), and HATCH (P<0.10) were found in medium-sized eggs when compared with either small or large eggs. The CWT was associated with egg size (P<0.05) according to the order: large eggs, highest CWT > medium eggs, intermediate CWT > small eggs, lowest CWT. Neither EWL nor HATCH was affected by length of preincubation egg storage. Collectively, our findings suggest that 1) ostrich eggs that possess low LP and increased THICK hatched poorly, 2) intermediate sized eggs hatch best, 3) large eggs produced large chicks, and 4) ostrich eggs can be stored under conditions typically used in the poultry industry for a minimum of 10 d without negatively impacting HATCH. PMID- 10515355 TI - Influence of body weight restriction in a body-weight-selected line of turkeys on response to challenge with Pasteurella multocida. AB - Previous research has shown that a line (F) of turkeys selected long-term for increased 16-wk BW was more susceptible to challenge with washed Pasteurella multocida (PM) than a randombred control line (RBC2), the base population of the F line. Published research indicated that the mortality of the F line following challenge with PM was similar to that of two commercial sire lines. The purpose of the present study was to determine the influence of reducing BW of the F line to that of the RBC2 line by nutrient restriction on resistance to PM. Four challenge trials were conducted over a 2-yr period. The BW of a group of F line birds was restricted to that of the RBC2 line by limiting access to feed from 1 to 6 wk of age. The F line restricted birds and full-fed RBC2 and F line birds were challenged with a field isolate of washed PM (1.2x10(7) organisms/bird of capsular serogroup A and somatic serotype 3, 4) at 6 wk of age. Birds were checked twice daily for 14 d. Resistance to PM was measured by days to death of those that died and percentage mortality. The BW of the restricted group of the F line did not differ from full-fed RBC2 birds for males or females. In males, the restricted F line birds had similar mortality (48.0%) to the full-fed RBC2 line birds (44.3%), and the mortalities in both groups were significantly lower than that observed for the full-fed F line birds (81.3%) following challenge with PM. The mortality following challenge in females did not differ significantly among groups, even though mortality of the full-fed F line birds (64.1%) and restricted F line birds (63.3%) was more than 9% higher than that (54.2%) observed for the full-fed RBC2 line birds. Days to death was not a sensitive indicator of resistance to PM, as no differences among the three groups of birds were observed for either sex. PMID- 10515356 TI - Effect of feed withdrawal or challenge with Pasteurella multocida on growth, blood metabolites, circulating growth hormone, and insulin-like growth factor-I concentrations in eight-week-old turkeys. AB - The daily effects of feed withdrawal or a bacterial disease (Pasteurella multocida; PM) challenge was studied in a slow-growing line of turkeys. The following groups (n = 6 birds/group) were sampled for up to 13 d: untreated control (CON), 4-d feed withdrawal followed by refeeding (FAST), a group that succumbed within the first 2 to 3 d after PM challenge (E-DEAD), a group that succumbed 8 to 9 d after PM challenge (L-DEAD), a group that survived the PM challenge (SUR), and a group treated with both PM challenge and 4-d feed withdrawal followed by refeeding (FAST/CHAL). Daily feed intake and BW gains were markedly reduced in the E-DEAD and L-DEAD groups immediately and 3 d after PM challenge, respectively. Feed intake and BW gain between CON and SUR groups of turkeys were not different throughout the trial. The turkeys in the FAST group followed the expected feed withdrawal and refeeding patterns for feed intake and BW loss or gain. The FAST/CHAL turkeys consumed the minimal amount of feed to maintain BW after refeeding. Plasma uric acid sharply increased 1 d prior to death in both E-DEAD and L-DEAD groups of turkeys. Plasma uric acid also increased each consecutive day during fasting in the FAST and FAST/CHAL groups of turkeys. Plasma growth hormone was measured in only the CON and FAST groups and increased from about 40 to 85 ng/mL in the FAST group during fasting but returned to control levels within 1 d of refeeding. Circulating plasma insulin-like growth factor-I (IGF-I) decreased from about 17 to 5 ng/mL in the PM-challenged (E-DEAD, L-DEAD, and FAST/CHAL groups) and FAST groups. The concentration of IGF-I returned to prefeed withdrawal levels within 3 d of refeeding the FAST group of turkeys. It was concluded that 1) turkey poults that were not susceptible to the PM challenge generally maintained physiological functions at control bird levels, 2) susceptible turkey poults generally exhibited depressed feed intake and BW gains, and 3) poults challenged with both feed withdrawal and PM treatment responded differently than poults challenged with either feed withdrawal or challenge with PM. The depletion of energy intake and mobilization of energy stores in susceptible poults might have contributed to the rate at which PM caused the poults to die. PMID- 10515357 TI - Effects of fumonisin B1 on selected immune responses in broiler chicks. AB - Three experiments were conducted to evaluate immune responses in chicks fed fumonisin B1 (FB1). Day-old male chicks were randomly allotted to dietary treatments: 0, 50, 100, or 200 mg FB1/kg diet. In Experiment 1, chicks were fed diets for 3 wk and were injected intravenously with 4.6x10(6) Escherichia coli on Day 21. Blood samples were collected at 60, 120, and 180 min postinjection, and liver, spleen, and lung were collected after 180 min. Chicks fed 200 mg FB1/kg diet had significantly higher numbers of bacterial colonies in blood, spleen, and liver (P<0.05) than control chicks. In Experiment 2, chicks were placed on the diets for 4 wk and were injected with 0.5 mL inactivated Newcastle Disease virus vaccine on Weeks 2 and 3 of the experiment, and primary and secondary antibody titers were measured 7 d after each injection. The secondary antibody response in chicks fed 200 mg FB1/kg diet was significantly lower (P<0.05) than that of control chicks. In Experiment 3, lymphocyte proliferation in chicks exposed to FB1 in vivo or in vitro was determined. Results of the in vivo study showed that cell proliferation in response to mitogens was lower (P<0.05) in chicks fed 200 mg FB1/kg diet than in control chicks. For the in vitro study, cell proliferation was lower (P<0.05) when cells were exposed to > or = 2.5 microg FB1/mL. Data of the current study suggested that FB1 is immunosuppressive in chicks when present in the ration at 200 mg FB1/kg diet. PMID- 10515358 TI - Tryptophan requirement of the commercial hen. AB - An experiment was conducted to determine the Trp requirement of the commercial laying hen. A corn-soybean meal diet was used that contained gelatin, which is low in Trp. Tryptophan was the first-limiting amino acid in this diet. Methionine, Lys, Arg, Thr, Val, and Ile were added to ensure that they were not deficient. Experimental diets containing 0.0, 0.02, 0.04, 0.06, 0.08, 0.10, and 0.12% Trp were fed, and a corn-soybean meal diet served as the positive control. Egg production (EP), egg mass (EM), and egg content (EC) were significantly increased by the addition of supplemental Trp. Broken-line regression indicated that the Trp requirement for EP and EC for this experiment was 136.0 (R2 = 0.81) and 136.5 (R2 = 0.82) mg per hen per d, respectively. These hens produced 43.5 g of EC, resulting in a Trp requirement of 3.14 mg/g of EC. Therefore, assuming an output of 50 g of EC, we suggest 157 mg per hen per d as the requirement for maximum production of EC. PMID- 10515359 TI - Lipoprotein hydrolysis and fat accumulation in chicken adipose tissues are reduced by chronic administration of lipoprotein lipase monoclonal antibodies. AB - The lipoprotein lipase (LPL) catalyzed hydrolysis of plasma lipoproteins is a rate-limiting step in the lipid transport into peripheral tissues. The aim of the present study was to isolate monoclonal antibodies against chicken adipose LPL and to investigate whether chronic infusion of the LPL monoclonal antibodies inhibits adipose LPL activity and consequently reduces fat accumulation in broiler chickens. The LPL catalyzed very low density lipoprotein (VLDL) hydrolysis was completely inhibited by the addition of 100 microg/mL of monoclonal antibodies (CLP10, CLP14, CLP16) in the in vitro incubation with plasma VLDL and LPL. A single injection of CLP10 and CLP16 into chickens fed or starved for 24 h elevated plasma triacylglycerol concentrations for 24 h, whereas that of CLP14 was ineffective. Intravenous injection every other day and continuous infusion by osmotic minipump with CLP16 maintained higher plasma triacylglycerol concentration for 5 d than that of the control group and extensively reduced LPL activity in adipose tissues and abdominal fat pad weight. Lipoprotein lipase mRNA and protein levels in adipose tissue were not modified by chronic administration of anti-LPL antibody. The results indicate that chronic administration of anti-LPL antibodies is effective in retarding fatness in broiler chickens, and the antibodies are a proper subject for studies of lipoprotein metabolism. PMID- 10515360 TI - Effects of dietary fat type and xylanase supplementation to rye-based broiler diets on selected bacterial groups adhering to the intestinal epithelium. on transit time of feed, and on nutrient digestibility. AB - Two experiments were conducted to examine the effects of different fat types, i.e., soybean oil (S) and beef tallow (T), in rye-based broiler diets, either unsupplemented (-) or supplemented (+) with xylanase (Avizyme 1300 at 1 g/kg diet), on selected bacterial groups adhering to the epithelium of crop, duodenum, jejunum, and ileum (Experiment 1, 16 d of age), on mean retention time (MRT) of digesta, and on digestibility of N and dry matter in successive segments of the digestive tract (Experiment 2, 24 d of age). Live weight of enzyme-treated and S fed chickens was significantly higher than that for unsupplemented or T-fed birds, respectively, in both experiments. In Experiment 1, a reduction in bacterial colonization from crop to duodenum was followed by a continuous increase as far as the ileum. Xylanase supplementation significantly reduced enterobacteria and total anaerobe microbes with a similar trend for Gram-positive cocci and enterococci. The latter two groups were significantly enhanced in birds fed T. In Experiment 2, xylanase supplementation resulted in a decrease in MRT in several segments of the digestive tract. This effect was most pronounced in the small intestine, where MRT of 268, 217, 241, and 209 min in groups S-, S+, T-, and T+, respectively, were measured. Apparent digestibility of N and dry matter was slightly improved by xylanase supplementation in the jejunum and ileum. Nitrogen digestibility by the terminal ileum was 80.3, 83.7, 79.4, and 82.2% for the S-, S+, T-, and T+ groups, respectively, and dry matter digestibility amounted to 61.2, 65.5, 62.1, and 64.0%, respectively. PMID- 10515361 TI - Evaluation of sodium bicarbonate, chloride, or sulfate with a coccidiostat in corn-soy or corn-soy-meat diets for broiler chickens. AB - During the period from January to June, combined-sex broiler chickens were inoculated with coccidia via drinking water at 14 d of age. In a completely randomized design (eight replicate pens; 88 chicks per pen) using built-up litter, experimental diets contained monensin plus 0.20% dietary sodium bicarbonate (SBC), which provided 0.054% sodium and 0.144% bicarbonate. Treatment with SBC significantly improved coccidial lesion score, 45-d body weight, and feed efficiency compared with monensin alone. In a 2 x 5 factorial trial using built-up litter pens (eight replicate pens; 88 chicks per pen) vs. each ionophore alone, 0.20% dietary SBC with monensin significantly improved body weight, uniformity, and feed efficiency; 0.20% SBC with halifuginone, lasalocid, monensin, or salinomycin significantly reduced mortality; and 0.20% SBC with lasalocid, monensin, or salinomycin significantly increased breast meat yield. In a 2x4 factorial trial (12 replicate pens; 88 chicks per pen) on built-up litter, corn-soy and corn-soy-meat diets (higher potassium, lower chloride) with monensin were evaluated using 0.054% sodium from SBC, NaCl, or sodium sulfate decahydrate (SSD). With both diet types, SBC (0.20%) or NaCl (0.139% extra) significantly improved weight uniformity, feed efficiency, mortality, and breast meat yield; however, the SSD results were closer to controls. In a 21-d battery brooder test using similar diets and design (2x4 factorial; 4 replicate pens; 10 chicks per pen), SBC and NaCl significantly reduced coccidial lesion scores; SSD produced a significant, but weaker effect. Extra NaCl significantly increased water intake (approximately 37%), water excretion (approximately 27%), and litter moisture (approximately 22%) with both diet types. The SSD did not affect water intake. PMID- 10515362 TI - Effects of line, dietary protein, sex, age, and feed withdrawal on insulin-like growth factor-I in White Pekin ducks. AB - Three experiments were conducted to determine and characterize plasma insulin like growth factor-I (IGF-I) concentrations in Pekin ducks. Plasma IGF-I in Pekin ducks was assayed in a heterologous radioimmunoassay for human IGF-I. When treated with acid, the response dose of duck IGF-I was parallel to that of human recombinant (r) IGF-I. The effect of line (greater breast muscle thickness vs. control) was determined in Experiment 1 in female ducks. The ducks with greater breast muscle thickness had higher (P<0.05) plasma IGF-I concentrations than the control ducks. In Experiment 2, the effects of dietary protein, sex, and age were examined from 42 to 49 d of age. Three dietary programs that differ in dietary crude protein were used in this experiment. Ducks on the high protein program had (P<0.05) higher plasma IGF-I concentrations than ducks on either medium or low protein programs. Males exhibited higher (P<0.05) IGF-I than females. Plasma IGF I concentrations decreased with age from 42 to 49 d. In Experiment 3, the effects of selection criterion (high or low breast muscle thickness to total breast thickness ratio) and the feed-deprived or fed state were studied in female Pekin ducks. The high ratio ducks were more affected by feed deprivation. These ducks had similar plasma IGF-I concentrations to low ratio ducks during feed withdrawal, but had higher (P<0.05) concentrations when fed. These data contribute to an understanding of the influence of IGF-I on metabolism and will be of value to the improvement of lean Pekin duck production. PMID- 10515363 TI - Hydrogen gas production of broiler chicks in response to soybean meal and alpha galactoside free, ethanol-extracted soybean meal. AB - In order to measure broiler chick hydrogen gas production, a sealed atmosphere chamber was constructed and chicks were intubated with soybean meal (SBM), alpha galactoside free, ethanol-extracted soybean meal (ESBM), and ESBM with alpha galactosides added to the levels of SBM (ESBMG). Six male broiler chicks averaging 156 g of weight were deprived of feed for 12 h prior to intubation with 6 g of the test soybean meals. Two chicks were used for each treatment. Following intubation, chicks were placed in the sealed atmosphere chamber for 20 min at 2-h intervals for 28 h. At the end of this 20-min period, a sample of the chamber atmosphere was collected with a gas-tight syringe and analyzed for hydrogen gas by gas-solid chromatography. The hydrogen production of the two chicks intubated with SBM peaked 7 h postintubation at 127 ppm. The ESBM produced a peak at approximately 17 h postintubation at 26 ppm. Intubation with ESBMG resulted in peak hydrogen production at approximately 12 h postintubation at an average of 67 ppm. Results indicate that chicks intubated with SBM produced 3.2 times the amount of total hydrogen gas than those chicks intubated with ESBM. Chicks intubated with ESBMG produced 2.2 times the amount of total hydrogen gas than chicks intubated with ESBM. The research indicates the alpha-galactoside oligosaccharides are a major cause of hydrogen gas production from SBM in poultry. PMID- 10515364 TI - Effect of phytase enzyme on dietary nitrogen-corrected apparent metabolizable energy and the ileal digestibility of nitrogen and amino acids in broiler chicks. AB - The effect of phytase on the performance, AMEn, and the ileal digestibility of N and amino acids was investigated in a 15-d trial using day-old male broilers with diets that were low in Ca (0.9% for control and 0.79% for phytase treatment) and available P (AP; 0.45% for control and 0.35% for phytase treatment). The assayed dietary phytase activity of crumble diet was 1,149 phytase unit (FTU)/kg. Chromic oxide was added to the diets to estimate ileal digestibility of N and amino acid. Excreta were collected from Day 12 to 15 to estimate AMEn. Weight gain, feed intake, and feed:gain of chicks fed phytase using diets with low Ca and AP were comparable with those observed for chicks fed more normal levels of Ca and AP. The diet with supplemental phytase had a higher AMEn (P< or =0.01) compared with the control diet. Chicks fed phytase had higher digestibilities for Val, Ile, nonessential amino acids (P< or =0.05), and total amino acids (P< or =0.01). PMID- 10515365 TI - In ovo peptide YY administration improves growth and feed conversion ratios in week-old broiler chicks. AB - The effects of in ovo Peptide YY (PYY) administration on growth and feed conversion ratios in a commercial broiler line were investigated. Six hundred Ross male x Cobb female eggs were administered either 0.9% saline (control) or 600 microg/kg egg weight PYY in ovo at Day 18 of incubation. On day of hatching, 210 birds from each treatment group were randomly placed by sex into pens. Body weights at placement were not different between treatment groups. Average chick body weight and adjusted pen feed conversion ratios were improved by PYY in ovo treatment at 7 d posthatch (165.7 vs. 170.2 g, P<0.02; and 1.55 vs. 1.49, P<0.04, respectively). No significant differences between treatments were noted for these parameters at 21 or 42 d of age. These results suggest that in ovo treatment of broiler chicken eggs with gastrointestinal hormones that increase intestinal nutrient absorption, such as PYY, may enhance chick performance. PMID- 10515366 TI - Broiler breast meat color variation, pH, and texture. AB - Four experiments were conducted to determine the range of breast meat color variation observed in commercial processing plants and its relationship to muscle pH and texture. Boneless, skinless breast fillets were collected weekly from the deboning lines of five commercial processing plants over a 4-wk period. Plant personnel selected breast fillets based on their appearance as being "lighter than normal," "normal," or "darker than normal" in color. Five representative fillets of each appearance classification were transported to a central laboratory for analyses of visual score (1 to 5 with 1 = light, 3 = normal, or 5 = dark), instrumental color, muscle pH, and Allo-Kramer shear (n = 300). Visual scores, lightness (L*), redness (a*), and pH were different (P<0.05) among the three appearance groups, with no significant differences in yellowness (b*). Visual scores averaged 2.4, 3.0, and 3.6, L* was 48.8, 45.6, and 43.1, and pH was 5.63, 5.70, and 5.81 for the lighter, normal, and darker fillets, respectively. There were no significant effects of color group on breast meat texture. Correlations between the color values and pH, however, were all highly significant. These results not only indicate that there are wide variations in breast meat color in commercial production, but they also demonstrate a strong relationship between breast meat color and muscle pH. PMID- 10515367 TI - Age-related changes in meat tenderness and tissue pentosidine: effect of diet restriction and aminoguanidine in broiler breeder hens. AB - The nonenzymatic glycosylation of tissue protein contributes to the formation of crosslinks that leads to structural and functional deterioration in the long lived tissue protein, collagen. The accumulation of these crosslinks thus contributes to the objectionable toughness of meat from aged animals, decreases its economic value, and limits its use in whole muscle foods. The objectives of this study were to determine the effectiveness of diet restriction and the crosslinking inhibitor, aminoguanidine (AG), on reducing the accumulation of crosslinks, thereby improving meat tenderness in broiler breeder hens. The glycoxidation product, pentosidine, was also measured in skin (Ps) to determine whether changes in its concentrations correlated with the changes in shear value (SV). Chicks (n = 450) were randomly assigned to four treatment groups from 8 to 125 wk after hatch: ad libitum (AL), diet restricted (DR), AL and DR groups supplemented with 400 ppm AG each (AL+AG and DR+AG, respectively). Shear value was measured with an Instron Universal Mechanical Machine. Skin pentosidine was isolated by reverse phase HPLC. There was an age-related, linear increase in SV (P<0.0001, r = 0.96), which correlated (r = 0.86) with the age-related increase in Ps in AL hens. Diet restriction retarded SV (P<0.0001) over the sampling period. In general, SV values for AL+AG were similar to those measured in DR, whereas no additive effect was observed for AG in DR birds. It was concluded that there was a linear increase in meat toughness (SV) with age that correlates with the accumulation of Ps, and that the decline in meat tenderness can be retarded by DR or AG. Secondly, the effect of DR on accumulation of Ps was so pronounced that AG supplementation did not further enhance this effect. PMID- 10515368 TI - A comparison of texture and quality of breast fillets from broilers stunned by electricity and carbon dioxide on a shackle line or killed with carbon dioxide. AB - To compare the broiler breast muscle quality resulting from three different slaughter methods, 36 broilers in each of two replicates were randomly divided into three groups receiving CO2 stunning, electrical stunning (ES), or CO2 killing. Carbon dioxide stunning was accomplished in a tunnel with a gradient from 40 to 60% CO2 by allowing the broilers on shackles to pass through the tunnel for 25 s. Electrical stunning was done by passing the bird's head through a charged 1% brine solution (35 mA, 7 s). For CO2 killing, the birds were killed by asphyxiation in an atmosphere of less than 2% oxygen (air displaced by CO2) for 2.5 min. Following slaughter, all breast fillets were harvested at 1.25 h postmortem and analyzed for pH, R value, shear value (SV), expressible moisture, and color (lightness and redness at 1.25 and 24 h postmortem). There were no differences (P<0.05) between treatments in pH, R value, SV, 1.25-h color values, or expressible moisture. There was an increase (P<0.05) in lightness between 1.25 and 24 h postmortem in all treatments, with the CO2 stun exhibiting the greatest increase and resulting in a significantly greater L* value at 24 h postmortem than the CO2 killing treatment. These results suggest that the postmortem metabolism or characteristics of the meat from animals processed with these stunning or killing methods does not differ to a large extent. PMID- 10515369 TI - Chronic allograft rejection. Do the Th2 cells preferentially induced by indirect alloantigen recognition play a dominant role? AB - Chronic rejection has been the major obstacle to the long-term allograft survival in the clinic. Although the etiology of this rejection reaction is multifactorial, alloantigen-specific immune activation plays the most critical role. We herein hypothesize that CD4+ Th2 cells that are preferentially induced by the indirect recognition of allogeneic histocompatibility antigens late in transplantation may play the most critical role in the initiation and/or maintenance of chronic allograft rejection. Immunosuppression used to prevent acute rejection and the nature of antigen-presenting cells and alloligands in the graft may all contribute to immune deviation to the Th2 response. This response may be further perpetuated by type 2 cytokines conceivably produced by activated macrophages, NK cells, and CD8+ T cells in the graft. Cytokines and growth factors induced by this type 2 response, in turn, allow for activation of B, endothelial, and smooth muscle cells that collectively contribute to the pathogenesis of chronic allograft rejection by producing alloantibodies and growth hormones required for interstitial fibrosis, extracellular matrix deposition, and vascular neointimal hyperplasia. PMID- 10515370 TI - Accessory function for NK1.1+ natural killer cells producing interferon-gamma in xenospecific cytotoxic T lymphocyte differentiation. Whither the natural killer cell. PMID- 10515371 TI - Prevention of chronic renal allograft rejection in rats with an oral endothelin A receptor antagonist. Relax-relax. PMID- 10515372 TI - A new, unique and simple method for ureteral implantation in kidney recipients with small, defunctionalized bladders. AB - BACKGROUND: Major, almost insurmountable, deterrents exist to the use of the small capacity, defunctionalized, nonneurogenic urinary bladder in renal transplantation, namely, the technical difficulty in performing a satisfactory ureteral implantation with conventional methods and the potential secondary problems with high grade ureteral reflux and obstruction. Alternatives are less than ideal and include transplantation into a bowel-augmented urinary bladder with intermittent self-catheterization, ileal conduit urinary diversion, or avoidance of transplantation and relegating the patient to life-long dialysis. METHODS: Eight consecutive patients (ages 13 months to 29 years) with small, defunctionalized urinary bladders underwent a new method of intravesical implantation of the transplant ureter. The mean capacity of these bladders was 18.5+/-13.1 ml (range 6 to 45 ml), with the bladders defunctionalized for a mean 81.6+/-24.3% of the patients' total lifetime. The technique involved placement of the transplant ureter into a shallow, mucosa-denuded, rectangular trough extending from a superiorly placed ureteral hiatus distally to the trigone. We hypothesized that the mucosal margins on the two lateral aspects of the rectangular trough would grow over the anterior surface of the ureter until they met the advancing mucosal edges from the contralateral side to form a natural neosubmucosal tunnel. RESULTS: Posttransplantation cystoscopic examination demonstrated bladder mucosal regeneration and growth over the ureter, confirming the spontaneous development of a good length neosubmucosal tunnel. All patients demonstrated no evidence of ureteral reflux or ureteral obstruction, whereas an immediate prior cohort of four consecutive patients with bladder capacities < or =30 ml showed that three of four had ureteral reflux (P=0.02) and four of four developed hydronephrosis (P=0.002). All urinary bladders in the present cohort enlarged to expected normal or nearnormal capacities. Serum creatinines were stable throughout the entire follow-up period, with the exception of one patient who had rejection episodes. Two patients had urinary tract infections posttransplantation, but there were no episodes of acute pyelonephritis. CONCLUSIONS: This novel technique for ureteral implantation successfully capitalizes on the regenerative potential of the bladder mucosa, resulting in a physiological, anatomically natural, and very effective neosubmucosal tunnel. It appears to guarantee success against both ureteral reflux and obstruction, no matter how small the urinary bladder, and offers no hindrance to enlarging the bladder to near normal capacity posttransplantation. The implantation technique is simple and safe, and its use should eliminate the reluctance to use these bladders. Moreover, this procedure offers a major incentive for the successful rehabilitation of small, defunctionalized, nonneurogenic bladders after kidney transplantation. PMID- 10515373 TI - Prevention of chronic renal allograft rejection in rats with an oral endothelin A receptor antagonist. AB - BACKGROUND: Chronic rejection is the most common cause of graft loss in renal transplantation. The pathomechanisms underlying chronic rejection are poorly understood, and no treatment has yet successfully been established. We hypothesized that, in analogy to models of reduced renal mass, the administration of a selective endothelin (ET) A receptor antagonist could improve the course of chronic rejection in renal allografts. METHODS: Experiments were performed in the Fisher-to-Lewis rat model of chronic rejection. Lewis-->Lewis isografts served as controls. Animals were treated with either the oral selective ET-A receptor antagonist LU135252 (50 mg/kg/day) or vehicle. Animal survival, blood pressure, creatinine clearance, proteinuria, and urinary ET excretion were investigated for 24 weeks. Kidneys were removed for light microscopical evaluation, determination of ET mRNA expression and tissue protein concentration, and immunohistochemical assessment of cell surface markers. RESULTS: Rats with chronic rejection showed an increase in renal ET mRNA synthesis and ET protein content. Treatment with LU135252 resulted in a significant improvement in survival after 24 weeks (0.92 vs. 0.38, P<0.01 by log-rank test). Creatinine clearance was higher in animals treated with the selective ET-A receptor antagonist (P<0.05). LU135252 had no influence on blood pressure and proteinuria. Selective ET-A blockade was associated with significantly less morphological changes and a significant reduction of expression of cell surface markers for macrophages (ED1), T cells (R73), and MHC II (F17-23-2). CONCLUSION: The renal ET-A system plays an important role in the pathomechanisms underlying chronic renal allograft rejection, because the treatment with a selective ET-A receptor antagonist dramatically improves the course of chronic renal failure after allograft transplantation. These results offer a novel therapeutical option for treatment of chronic renal allograft rejection, for which so far no therapy is known. PMID- 10515375 TI - Prolonged cardiac allograft survival in rats systemically injected adenoviral vectors containing CTLA4Ig-gene. AB - BACKGROUND: CTLA4Ig, a soluble recombinant fusion protein that contains the extracellular domain of the CTLA4 and Fc portion of IgG1, strongly adheres to the B7 molecule to block CD28-mediated costimulatory signals and inhibits in vitro and in vivo immune responses. In vivo gene transfer using adenovirus vector achieves a high transfection rate into organ cells that usually contain adenoviral receptors. In this study, we investigated expression levels of the transfected gene and the survival times of the allografts in cardiac recipients systemically administered adenoviral vectors containing CTLA4Ig. METHODS: Hearts from DA rats (RT-1a) were transplanted into a cervical location in LEW recipients (RT1(1)). The adenoviral vectors containing CTLA4Ig was injected via a recipient vein immediately after grafting. RESULTS: The serum level of CTLA4Ig reached to maximum at 51-93 microg/ml 3 to 7 days after gene-transfection and declined after 14 days, although detectable levels were observed up to 49 days. The median survival time of the allografts in the gene-transfected group were significantly prolonged (27 days) in compared to the control group (6 days). In addition, down regulation of IL-2 and IFN-gamma mRNAs and persistence of IL-4 and IL-10 transcripts were observed in the graft infiltrating cells. CONCLUSION: The adenovirous-mediated CTLA4Ig gene transfer into a recipient liver by systemic administration resulted in remarkable prolongation of cardiac allograft survival. Its action mechanisms may be mediated by inhibition of CD28-associated signal transduction, reduction of Th1-type cytokine production, and continuous expression of Th2-type cytokines in the activating lymphocytes. PMID- 10515376 TI - A rat small bowel transplant model of chronic rejection: histopathologic characteristics. AB - BACKGROUND: The major impediment to long-term success in solid organ transplantation is the development of chronic rejection (CR). The vascular lesion of CR, transplant vascular sclerosis (TVS) is characterized by neointimal smooth muscle cell proliferation, and is driven by both immune- and nonimmune-mediated mechanisms. Although the features of chronic heart and kidney allograft rejection have been well characterized, the more immunogenic small bowel allograft has not received similar study. METHODS: F344 small bowel (SB) was transplanted heterotopically into Lewis recipients that were treated with low-dose Cyclosporine A for 15 days. Lewis recipients of F344 or Lewis SB grafts without immunosuppression, served as controls. Grafts were assessed histologically when recipients showed clinical signs of rejection or at predetermined time points. The immunological components involved in the chronic rejection process were evaluated by immunohistochemical staining. RESULTS: All SB allografts (100%) developed histologic evidence of CR Cyclosporine A. TVS was seen in 36 of the 46 (78%) of these allografts. The median time to develop TVS was 45 days. Immunohistochemical staining of chronically rejected grafts showed infiltration predominantly by CD4+ cells and macrophages, uniform up-regulation of class II MHC molecule expression, moderate to intense ICAM-1 staining in grafts harvested at postoperative day 45, and uniform neointimal cell staining for smooth muscle cell alpha-actin in the TVS lesions. CONCLUSIONS: This F344 to Lewis SB transplant model is a useful model that reproduces significant features of CR. The highly immunogenic nature of the SB allografts allows this model to serve as a stringent test for protocols designed to prevent CR. PMID- 10515374 TI - Increased apoptosis of immunoreactive host cells and augmented donor leukocyte chimerism, not sustained inhibition of B7 molecule expression are associated with prolonged cardiac allograft survival in mice preconditioned with immature donor dendritic cells plus anti-CD40L mAb. AB - BACKGROUND: We previously reported the association among donor leukocyte chimerism, apoptosis of presumedly IL-2-deficient graft-infiltrating host cells, and the spontaneous donor-specific tolerance induced by liver but not heart allografts in mice. Survival of the rejection-prone heart allografts in the same strain combination is modestly prolonged by the pretransplant infusion of immature, costimulatory molecule-(CM) deficient donor dendritic cells (DC), an effect that is markedly potentiated by concomitant CM blockade with anti-CD40L (CD154) monoclonal antibody (mAb). We investigated whether the long survival of the heart allografts in the pretreated mice was associated with donor leukocyte chimerism and apoptosis of graft-infiltrating cells, if these end points were similar to those in the spontaneously tolerant liver transplant model, and whether the pretreatment effect was dependent on sustained inhibition of CM expression of the infused immature donor DC. In addition, apoptosis was assessed in the host spleen and lymph nodes, a critical determination not reported in previous studies of either spontaneous or "treatment-aided" organ tolerance models. METHODS: Seven days before transplantation of hearts from B10 (H-2b) donors, 2x10(6) donor-derived immature DC were infused i.v. into C3H (H-2k) recipient mice with or without a concomitant i.p. injection of anti-CD40L mAb. Donor cells were detected posttransplantation by immunohistochemical staining for major histocompatibility complex class II (I-Ab) in the cells of recipient lymphoid tissue. CM expression was determined by two-color labeling. Host responses to donor alloantigen were quantified by mixed leukocyte reaction, and cytotoxic T lymphocyte (CTL) assays. Apoptotic death in graft-infiltrating cells and in areas of T-dependent lymphoid tissue was visualized by terminal deoxynucleotidyltransferase-catalyzed dUTP-digoxigenin nick-end labeling and quantitative spectrofluorometry. Interleukin-2 production and localization were estimated by immunohistochemistry. RESULTS: Compared with control heart transplantation or heart transplantation after only DC administration, concomitant pretreatment with immature donor DC and anti-CD40L mAb caused sustained elevation of donor (I-Ab+) cells (microchimerism) in the spleen including T cell areas. More than 80% of the I-Ab+ cells in combined treatment animals also were CD86+, reflecting failure of the mAb to inhibit CD40/ CD80/CD86 up-regulation on immature DC in vitro after their interaction with host T cells. Donor-specific CTL activity in graft-infiltrating cells and spleen cell populations of these animals was present on day 8, but decreased strikingly to normal control levels by day 14. The decrease was associated with enhanced apoptosis of graft-infiltrating cells and of cells in the spleen where interleukin-2 production was inhibited. The highest levels of splenic microchimerism were found in mice with long surviving grafts (>100 days). In contrast, CTL activity was persistently elevated in control heart graft recipients with comparatively low levels of apoptotic activity and high levels of interleukin-2. CONCLUSION: The donor-specific acceptance of rejection-prone heart allografts by recipients pretreated with immature donor DC and anti-CD40L mAb is not dependent on sustained inhibition of donor DC CM (CD86) expression. Instead, the pretreatment facilitates a tolerogenic cascade similar to that in spontaneously tolerant liver recipients that involves: (1) chimerism-driven immune activation, succeeded by deletion of host immune responder cells by apoptosis in the spleen and allograft that is linked to interleukin-2 deficiency in both locations and (2) persistence of comparatively large numbers of donor derived leukocytes. These tolerogenic mechanisms are thought to be generic, explaining the tolerance induced by allografts spontaneously, or with the aid of various kinds of immunosuppression. PMID- 10515377 TI - The effect of graded steatosis on flow in the hepatic parenchymal microcirculation. AB - BACKGROUND: Steatosis is a major cause of microcirculatory impairment and graft dysfunction after liver transplantation. The mechanism of this circulatory compromise is unclear. The aim of this study was to evaluate in vivo the effect of steatosis on parenchymal microcirculation and on total hepatic blood flow in an animal model. METHODS: Four groups of New Zealand White rabbits (n=24) were investigated. Group 1 were fed on normal diet (controls). In groups 2, 3, and 4 graded steatosis was induced by feeding on a high cholesterol diet (1.5%) for 4, 8, and 12 weeks, respectively. After laparotomy and exposure of the liver, total hepatic blood flow (THBF) and the hepatic parenchymal microcirculation (HPM) were measured. These parameters were correlated with the degree of histological fat infiltration classified as mild (<30%), moderate (30-60%), or severe (>60%). RESULTS: The 4-, 8-, and 12-week cholesterol diets resulted in mild, moderate, and severe steatosis, respectively. There was an inverse correlation between the degree of fat infiltration and both HPM (Spearman r=-0.967, P<0.0001) and THBF (r=-0.893, P<0.0001). THBF was 137+/-6 ml/min in controls, which reduced to 121+/ 3, 99+/-5, and 63+/-5 ml/min in steatotic livers of groups 2, 3, and 4, respectively. HPM was 226+/-5 flux units in the controls and 197+/-7, 119+/-8, and 37+/-9 flux units in steatotic livers of groups 2, 3, and 4 respectively. Comparing with controls using analysis of covariance, the fall in HPM and THBF was found to be significant (P<0.002) in the moderate and severe groups, but not significant (P>0.050) in the mild group. Parenchymal perfusion was reduced to a greater extent than total liver blood flow in moderate and severe grades of steatosis. CONCLUSIONS: Fatty infiltration reduces hepatic blood flow and parenchymal microcirculation. The latter is more markedly reduced with severe steatosis. This may explain the development of microcirculatory impairment and graft failure after transplantation of fatty livers despite adequate liver blood flow. PMID- 10515378 TI - Transforming growth factor-beta levels in human allograft chronic fibrosis correlate with rate of decline in renal function. AB - BACKGROUND: Long-term renal transplant function is limited primarily by a progressive scarring process loosely termed "chronic rejection, chronic allograft nephropathy, or allograft fibrosis." Although the etiology of transplant fibrosis is uncertain, several possible factors including chronic cyclosporin A (CsA) exposure may contribute to its pathogenesis. CsA stimulates renal fibrosis perhaps through the induction of the potent pro-sclerotic growth factor, transforming growth factor beta (TGFbeta). Previously, we demonstrated that, in human transplant biopsies, acute CsA toxicity but not acute tubular necrosis is associated with elevated levels of renal TGFbeta protein. We now examine whether long-term CsA treatment (>1 year) is associated with elevated levels of intra allograft TGFbeta and whether heightened expression of TGFbeta is clinically significant. METHODS: Using immunohistochemical techniques, we determined the relative level of expression of intrarenal TGFbeta protein in transplant biopsies. We studied biopsies obtained from 40 CsA-treated patients that were diagnosed as having chronic allograft fibrosis. Biopsies were scored as having minimal or high levels of TGFbeta. RESULTS: Seventy-two percent of patients expressed high levels of intra-allograft TGFbeta. This group of patients lost renal function at an average rate of -19.5+/-17.3 ml/min/year. In contrast, patients with minimal or no TGFbeta expression experienced a decline of only 6.2+/-4.1 ml/min/year (P=0.01). CONCLUSIONS: These results suggest that the majority of CsA-treated patients with biopsy proven chronic fibrosis have elevated levels of intra-graft TGFbeta that correlates with an increased rate of decline in renal function. PMID- 10515379 TI - Plasma cell-rich acute renal allograft rejection. AB - BACKGROUND: Acute renal allograft rejection is usually seen within the first 3 months posttransplant, and is characterized by an intense infiltrate of T cells. Some acute rejections, however, contain many plasma cells and/or appear late posttransplant. METHODS: We have investigated 27 cases of intensely plasma cell rich acute rejections (PCAR) from 1987 to 1997 and have compared them to 21 control cases (CAR) of typical acute rejection. Each group was divided into early (<6 months) and late (>6 months) subgroups. PCAR and CAR cases were matched for histological features of chronic allograft nephropathy. In all four groups, most cases had Banff '97 type IB and IIA acute rejection. RESULTS: A significantly greater number of PCAR cases experienced graft failure due to chronic allograft nephropathy or complications of acute rejection (P<0.05). There was no significant difference between PCAR and CAR in HLA matching, occurrence of posttransplant acute tubular necrosis, presence versus absence of previous allografts, number of previous or subsequent acute rejection episodes, Banff '97 sum scores for acute rejection, cyclosporine A or FK506 levels, or percent change from baseline creatinine at time of biopsy. Plasma cells in PCAR cases showed IgG predominance whereas those in CAR had comparable staining for IgG and IgA. Kappa and lambda light chain immunostaining of all PCAR cases revealed polyclonality. Three of 18 PCAR cases studied for the presence of Epstein-Barr virus RNA showed scattered positivity in 2-7% of lymphoid cells, although the remainder was negative. None of the PCAR cases developed post-transpland lymphoproliferative disorder. CONCLUSIONS: We conclude that PCAR can occur from 1 month to many years posttransplant, is associated with poor graft survival, and is not a manifestation of concomitant chronic allograft nephropathy or viral infection, including posttransplant lymphoproliferative disorder. PMID- 10515380 TI - Right lobe living donor liver transplantation. AB - BACKGROUND: The shortage of livers for transplantation has prompted transplant centers to seek alternatives to conventional cadaveric liver transplantation. Left lateral segmentectomy from living donors has proven to be a safe operation for the donor with excellent results in the pediatric population. Left lobectomy, conceived to supply more tissue, still provides insufficient liver mass for an average size adult patient. Right lobectomy could supply a graft of adequate size. METHODS: Donors were considered only after recipients were listed according to United Network for Organ Sharing (UNOS) criteria. Donor evaluation included liver biopsy, magnetic resonance imaging, and celiac and mesenteric angiography. The donor operation consisted of a right lobectomy uniformly performed throughout the series as described herein. RESULTS: Twenty-five right lobe living donor liver transplants were performed between adults, with no significant complications in donors. Recipient and graft survival was 88%, with three recipient deaths secondary to uncontrolled sepsis in patients at high risk for liver transplant; all three had functioning grafts. CONCLUSIONS: Right lobe living donor liver transplantation poses challenges that require a meticulous surgical technique to minimize morbidity in the recipient. Right lobectomies for living donation can be performed safely with minimal risk to both donor and recipient although providing adequate liver mass for an average size adult patient. PMID- 10515381 TI - Beneficial effects of inducible nitric oxide synthase inhibitor on reperfusion injury in the pig liver. AB - BACKGROUND: Although inhibition of endothelial nitric oxide synthase (eNOS) has been reported to aggravate hepatic ischemia-reperfusion (I/R) injury, the role of inducible nitric oxide synthase (iNOS) has been still unknown. We investigated the role of NO produced by iNOS, and evaluated the effect of an iNOS inhibitor on prolonged warm I/R injury in the pig liver. METHODS: Pigs were subjected to 120 min of hepatic warm I/R under the extracorporeal circulation. We investigated the time course of changes in serum and hepatic microdialysate NO2- + NO3- (NOx) and the cellular distribution of eNOS and iNOS by immunohistochemistry, including a double-immunofluorescence technique in combination with confocal laser scanning microscopy. The effect of iNOS inhibitor was also investigated. RESULTS: Hepatic I/R induced new nitric oxide production in serum and hepatic microdialysate NOx after reperfusion and severe hepatic damage in the centrilobular region where nitrotyrosine was strongly expressed. Diffuse eNOS expression in sinusoidal endothelium did not differ before and after reperfusion. In contrast, strong iNOS expression in Kupffer cells and neutrophils appeared strongly in the centrilobular region after reperfusion. Pigs with intraportal administration of N(G)-nitro-L-arginine (10 mg/kg) died during the period of ischemia or early in the period of reperfusion with a high mortality rate (80.0%). Intraportal administration of aminoguanidine hemisulfate (10 mg/kg) significantly suppressed nitric oxide production and serum aspartate aminotransferase after reperfusion, inhibited nitrotyrosine expression, and attenuated hepatic damage. CONCLUSIONS: These results indicate that hepatic I/R injury is triggered by centrilobular iNOS expression; and attenuated by inhibition of iNOS. PMID- 10515382 TI - Novel mechanism of inhibition of cytomegalovirus by the experimental immunosuppressive agent leflunomide. AB - BACKGROUND: Despite progress in antiviral chemotherapy, cytomegalovirus (CMV) remains a major cause of morbidity and mortality among pharmacologically immunosuppressed organ transplant recipients, frequently engaging the clinician in a struggle to balance graft preservation with control of CMV disease. Leflunomide, an inhibitor of protein kinase activity and pyrimidine synthesis, is an experimental immunosuppressive agent effective against acute and chronic allograft rejection in animal models. Because a number of CMV proteins are known to be phosphorylated, we tested the hypothesis that this agent might exert inhibitory activity against CMV. METHODS AND RESULTS: Plaque assays demonstrated dramatic dose-dependent attenuation of production of multiple clinical CMV isolates in leflunomide-treated human fibroblasts and endothelial cells, common targets for CMV infection in vivo. As shown by Northern blot analysis and immunohistochemical staining, leflunomide neither interferes with transcription of immediate early or late viral genes, nor with expression of corresponding proteins. CMV-specific DNA dot blots and biochemical enzyme assays indicated that, in contrast to currently approved anti-CMV drugs, leflunomide exerts no inhibitory effect on the accumulation of viral DNA in infected cells, or on viral DNA polymerase activity. Rather, as visualized by transmission electron microscopy, this agent appears to act at a late stage in virion assembly by preventing tegument acquisition by viral nucleocapsids. Finally we have demonstrated equivalent inhibitory activity of leflunomide against multi-drug resistant CMV isolates. CONCLUSIONS: These findings imply that leflunomide, an effective immunosuppressive agent, shows potential to concurrently attenuate a major complication of immunosuppression, CMV disease, by a novel mechanism of antiviral activity. PMID- 10515383 TI - Antisense oligodeoxynucleotides prevent acute cardiac allograft rejection via a novel, nontoxic, highly efficient transfection method. AB - BACKGROUND: We hypothesized that ex vivo donor allograft transfection with antisense oligodeoxynucleotide (AS ODN) would inhibit the expression of intercellular adhesion molecule (ICAM)-1, an important mediator of T-cell adhesion and costimulation, and therefore suppress acute cardiac rejection. METHODS: Hearts were transfected ex vivo with AS, reverse AS ODN, or saline by applying 3 atm pressure for 45 min at 4 degrees C. Grafts were then transplanted into allogenic recipients +/- treatment with leukocyte function-associated antigen (LFA)-1 monoclonal antibody (mAb) (1.5 mg/kg intravenously), cyclosporine (2.5 mg/ kg/day p.o.), or rapamycin (0.025 mg/kg/day intraperitoneally). Reperfusion injury was assessed in grafts harvested at early time points using the myeloperoxidase, %wet weight, and %contraction band necrosis assays; transfection efficiency was assessed using fluorescent microscopy; and efficacy of ICAM-1 blockade was assessed using immunohistochemistry. Other grafts were followed until rejection with donor/third-party skin grafting, adoptive transfer, and interleukin 2 infusion studies in selected recipients. RESULTS: Transfection was highly efficient (fluorescein isothiocyanate-ODN in 48+/-5% of total myocardial nuclei), nontoxic, and reduced the ICAM-1-positive area to 53+/-14% versus having no effect on MHC class I expression (n=4). The incidence of survival >60 days after AS ODN + LFA-1 monoclonal antibody was 75%, significantly higher than other regimens. CONCLUSION: AS ODN hyperbaric transfection proved highly efficient, effective at ICAM-1 blockade, and induced cardiac allograft tolerance when combined with LFA-1 monoclonal antibody. This highly targeted alteration of allograft immunogenicity may have an important role in future immunosuppressive strategies. PMID- 10515385 TI - Accessory function for NK1.1+ natural killer cells producing interferon-gamma in xenospecific cytotoxic T lymphocyte differentiation. AB - BACKGROUND: We have previously demonstrated that xenospecific cytotoxic T lymphocyte (CTL) differentiation requires accessory function by NK1.1+ cells, yet the mechanism by which NK1.1+ cells support CTL generation had not been elucidated. METHODS: An established model in which mice generate a strong local popliteal lymph node CTL response to footpad immunizations with human tumor cells was used. Mice depleted of NK1.1+ cells fail to mount a maximal xenospecific CD8+ CTL response. The xenospecific CTL response in anti-NK1.1 monoclonal antibody depleted mice could be completely restored if mice were coinoculated with human tumor cells (the xenoantigen) and xenoantigen-stimulated syngeneic natural killer (NK) cells from wild-type or perforin-deficient mice. By contrast, NK1.1+ cells from interferon-gamma-deficient mice did not restore the maturation of xenospecific CTL in anti-NK1.1 monoclonal antibody-treated mice. Depletion of NK1.1+ cells in vivo from both wild-type and Jalpha281-deficient mice (which lack Valpha14 NK1.1+ T cells) abrogated the generation of xenospecific CTL, however, untreated Jalpha281-deficient mice mounted a normal xenogeneic response. CONCLUSIONS: These data indicate that local NK cell production of interferon gamma at the site of challenge is an important stimulus for generating xenospecific CTL in local draining lymph nodes and that Valpha14 NK T cells play little or no regulatory function in this response. PMID- 10515384 TI - Regulated inhibition of coagulation by porcine endothelial cells expressing P selectin-tagged hirudin and tissue factor pathway inhibitor fusion proteins. AB - BACKGROUND: Thrombotic vascular occlusion resulting in infarction occurs during hyperacute rejection of allografts transplanted into sensitized patients and remains a major problem in experimental xenotransplantation. A similar process is also found in disorders of diverse etiology including atherosclerosis, vasculitis, and disseminated intravascular coagulation. METHODS: We have previously constructed two membrane-tethered anticoagulant fusion proteins based on human tissue factor pathway inhibitor and the leech anticoagulant hirudin and demonstrated their functional efficacy in vitro. These constructs have now been modified by the addition of a P-selectin sequence to the cytoplasmic tail to localize them in Weibel-Palade bodies. They have been transfected into Weibel Palade body-positive endothelial cells isolated from the inferior vena cava of normal pigs. RESULTS: In resting endothelial cells, fusion protein expression colocalized with P-selectin and was confined to Weibel-Palade bodies. These cells had a procoagulant phenotype in recalcified human plasma. However, after activation with phorbol ester the anticoagulant proteins were rapidly relocated to the cell surface where they specifically inhibited the clotting of human plasma. CONCLUSIONS: Novel anticoagulant molecules may prove useful therapeutic agents for gene therapy in thrombotic disease and postangioplasty or for transgenic expression in animals whose organs may be used for clinical xenotransplantation. Expression in vascular endothelial cells may be regulated by inclusion of P-selectin cytoplasmic sequence, to restrict cell surface expression to activated endothelium. PMID- 10515386 TI - Suppression of delayed xenograft rejection by specific depletion of elicited antibodies of the IgM isotype. AB - BACKGROUND: Hamster hearts transplanted into untreated rats undergo delayed xenograft rejection (DXR). This acute inflammatory response is associated with the deposition of anti-graft antibodies of the immunoglobulin (Ig)M isotype in the vasculature. We have previously shown that these antibodies are generated in a T cell-independent manner. In this study, we tested whether the generation of anti-graft IgM antibodies is involved in the pathogenesis of DXR. In addition, we tested whether the suppression of this antibody response would overcome DXR. METHODS: Hamster hearts were transplanted into rats treated with an anti-mu monoclonal antibodies (mAb) to deplete circulating IgM or with an isotype-matched control mAb recognizing the dinitrophenyl epitope. T cell immunosuppression was achieved with cyclosporin A (CsA). RESULTS: Depletion of circulating IgM by anti mu mAb inhibited DXR, whereas the control mAb had no effect on DXR. In anti-mu treated rats, xenografts were rejected 5-7 days after transplantation through a T cell-dependent mechanism associated with the generation of antibodies of the IgG isotype. Combination of anti-mu with CsA suppressed the anti-graft IgM and IgG response and resulted in long-term xenograft survival (> 50 days). Xenograft long term survival occurred despite the return of anti-graft IgM antibodies to the circulation, a phenomenon referred to as accommodation. CONCLUSION: This study demonstrates that the pathogenesis of DXR can be initiated by anti-graft antibodies of the IgM isotype, which are generated in a T-cell independent manner. In addition, we show that under T cell immunosuppression, specific depletion of this IgM response by anti-mu mAb administration results in xenograft long-term survival and accommodation. PMID- 10515387 TI - In vivo T cell depletion in miniature swine using the swine CD3 immunotoxin, pCD3 CRM9. AB - BACKGROUND: Partially inbred miniature swine developed in this laboratory provide a unique preclinical large animal model for studying transplant tolerance. The importance of in vivo T cell depletion for establishing stable mixed hematopoietic cell chimerism using a clinically relevant sublethal regimen has been well documented in murine studies (1). Until now, the lack of an effective in vivo T cell-depleting reagent in swine has limited the progress of studies involving hematopoietic cell transplants. METHODS: The swine CD3 immunotoxin, pCD3-CRM9, was prepared by conjugating our porcine-specific CD3 monoclonal antibody 898H2-6-15 to the diphtheria toxin derivative, CRM9. The resultant immunotoxin was administered i.v. to several miniature swine at doses ranging from 0.15-0.2 mg/kg either in a single dose or two doses 2 days apart. T-cell depletion was monitored in the peripheral blood, mesenteric lymph node, and thymus by flow cytometric analysis and histological examination. RESULTS: T cells were depleted to less than 1% of their pretreatment levels based on absolute numbers in the peripheral blood. Fluorescence activated cell sorter analysis and histological examination of serial lymph node biopsies confirmed depletion of the CD3+ T cells rather than down modulation or masking of the surface CD3 expression. Depletion of the CD3 bright medullary thymocytes could also be detected by flow cytometry and histological examination after immunotoxin treatment. CONCLUSIONS: Administration of the immunotoxin i.v. drastically depletes mature T cells from the peripheral blood, lymph node, and thymus compartments of the pig. This first description of an effective in vivo T-cell depleting reagent for the pig provides a valuable tool for studies of transplant tolerance in this large animal model. It also makes possible preclinical studies of T cell depletion with anti-CD3 immunotoxin in this large animal model. PMID- 10515388 TI - Alpha-galactosyl-mediated activation of porcine endothelial cells: studies on CD31 and VE-cadherin in adhesion and signaling. AB - BACKGROUND: Ligation of alpha-galactosyl epitopes on endothelial cells by naturally occurring human antibodies causes hyperacute rejection in porcine-to human xenotransplantation. The alpha-galactosyl-specific lectin Bandeiraea simplicifolia isolectin B4 (IB4) has been reported to trigger endothelial "gap" formation and tyrosine phosphorylation of an unidentified 130-kDa protein. We have studied two 130-kDa junctional adhesion molecules, CD31 and VE-cadherin, in porcine aortic endothelial cells (PAECs) during IB4-mediated activation. The cellular distribution of these molecules, their susceptibility to tyrosine phosphorylation, and their capacity to bind IB4 or natural human antibodies have been determined. METHODS: Porcine CD31 and VE-cadherin were cloned. Recombinant proteins and monoclonal antibodies were prepared. The distribution and phosphorylation of CD31 and VE-cadherin in confluent PAECs activated with IB4 or human serum were studied by confocal microscopy and Western blotting, respectively. RESULTS: IB4 caused rapid redistribution of CD31 and VE-cadherin away from cell junctions and tyrosine-phosphorylation of CD31 but not VE cadherin. A monoclonal antibody to CD31 also triggered tyrosine phosphorylation of this molecule, but brief exposure of PAECs to normal human serum did not. Tyrosine-phosphorylated CD31 complexed with SHP2 and other unidentified phosphoproteins. Both IB4 and natural human antibodies bound to porcine CD31 but not to VE-cadherin. Cell adhesion tests showed that porcine and human CD31 are functionally incompatible. CONCLUSIONS: Endothelial cell retraction during IB4 mediated activation of PAECs is associated with rapid loss of CD31 and VE cadherin from cell junctions. CD31 becomes strongly tyrosine-phosphorylated and forms a cell signaling complex, which may have a significant role in the response of the xenograft vascular endothelium. PMID- 10515389 TI - The control of epidermal stem cells (holoclones) in the treatment of massive full thickness burns with autologous keratinocytes cultured on fibrin. AB - BACKGROUND: Cell therapy is an emerging therapeutic strategy aimed at replacing or repairing severely damaged tissues with cultured cells. Epidermal regeneration obtained with autologous cultured keratinocytes (cultured autografts) can be life saving for patients suffering from massive full-thickness burns. However, the widespread use of cultured autografts has been hampered by poor clinical results that have been consistently reported by different burn units, even when cells were applied on properly prepared wound beds. This might arise from the depletion of epidermal stem cells (holoclones) in culture. Depletion of holoclones can occur because of (i) incorrect culture conditions, (ii) environmental damage of the exposed basal layer of cultured grafts, or (iii) use of new substrates or culture technologies not pretested for holoclone preservation. The aim of this study was to show that, if new keratinocyte culture technologies and/or "delivery systems" are proposed, a careful evaluation of epidermal stem cell preservation is essential for the clinical performance of this life-saving technology. METHODS: Fibrin was chosen as a potential substrate for keratinocyte cultivation. Stem cells were monitored by clonal analysis using the culture system originally described by Rheinwald and Green as a reference. Massive full-thickness burns were treated with the composite allodermis/cultured autograft technique. RESULTS: We show that: (i) the relative percentage of holoclones, meroclones, and paraclones is maintained when keratinocytes are cultivated on fibrin, proving that fibrin does not induce clonal conversion and consequent loss of epidermal stem cells; (ii) the clonogenic ability, growth rate, and long-term proliferative potential are not affected by the new culture system; (iii) when fibrin-cultured autografts bearing stem cells are applied on massive full-thickness burns, the "take" of keratinocytes is high, reproducible, and permanent; and (iv) fibrin allows a significant reduction of the cost of cultured autografts and eliminates problems related to their handling and transportation. CONCLUSION: Our data demonstrate that: (i) cultured autografts bearing stem cells can indeed rapidly and permanently cover a large body surface; and (ii) fibrin is a suitable substrate for keratinocyte cultivation and transplantation. These data lend strength to the concept that the success of cell therapy at a clinical level requires cultivation and transplantation of stem cells. We therefore suggest that the proposal of a culture system aimed at the replacement of any severely damaged self-renewing tissue should be preceded by a careful evaluation of its stem cell population. PMID- 10515390 TI - Perfusion of kidneys with unformulated "naked" intercellular adhesion molecule-1 antisense oligodeoxynucleotides prevents ischemic/reperfusion injury. AB - BACKGROUND: We have previously shown that phosphorothioate intercellular adhesion molecule (ICAM)-1 antisense oligodeoxynucleotide (oligo) IP-9125 blocks the expression of rat ICAM-1 mRNA in rat L2 cells. A single ex situ perfusion of grafts with unformulated IP-9125, suspended in Euro-Collins solution, prolonged the survival of kidney allografts in rats. The present experiments examined whether perfusion of kidneys with unformulated IP-9125 prevents ischemic/reperfusion injury. METHODS: Kidneys were perfused ex situ with 2 ml of Euro-Collins solution without or with IP-9125 and exposed to 30-min cold (4 degrees C storage time) and 30-min warm (anastomosis time) ischemia. Kidneys were then transplanted to syngeneic nephrectomized recipients. RESULTS: Within 24 hr after transplantation, the glomerular filtration rate values were reduced by almost 60% to 0.49+/-0.14 ml/min from 1.20+/-0.27 ml/min in normal kidneys (P<0.001). Kidney perfusion with 10 mg of either IP-12140 (0.41+/-0.07 ml/min) or IP-13944 (0.47+/-0.07 ml/min) control oligo was ineffective. In contrast, perfusion with 10 mg of IP-9125 significantly improved kidney function (0.8+/ 0.18 ml/min; P<0.005), whereas the lower doses of 2 mg (0.47+/-0.13 ml/min; NS) or 4 mg (0.54+/-0.04 ml/min; NS) had no significant effect. The glomerular filtration rate results were confirmed by measurements of blood creatinine (CR) levels at 24 hr after grafting: untreated recipients had a twofold higher CR value (0.70+/-0.14 mg/dl) compared with normal controls (0.65+/-0.07 mg/dl; P<0.001). Although perfusion with 10 mg of control IP-12140 (0.80+/-0.14 mg/dl) or IP-13944 (0.65+/-0.07 mg/dl) did not affect CR levels, perfusion with 10 mg of IP-9125 (0.45+/-0.07 mg/dl) lowered CR levels. The Western blots or reverse transcription-polymerase chain reaction experiments performed in kidney transplants within 24 hr after grafting showed that 10 mg of IP-9125 (but not control IP-12140) reduced the expression of ICAM-1 protein and ICAM-1 mRNA, respectively. CONCLUSIONS: Perfusion of grafts with unformulated ICAM-1 antisense oligo specifically reduces intragraft ICAM-1 protein expression and prevents ischemic/reperfusion injury. PMID- 10515391 TI - Resistance of established porcine islet xenografts to humoral rejection by hyperimmune sera. AB - BACKGROUND: Although preformed natural antibodies cause hyperacute rejection of primarily vascularized xenografts, tissue grafts such as skin or islets are revascularized by in-growth of host capillaries and therefore might be resistant to circulating antibodies. We examined the effect of hyperimmune serum and primed T cells on the survival of long-term porcine islet xenografts in diabetic nude mice. METHODS: Porcine islets were transplanted beneath the kidney capsule of streptozotocin-induced diabetic BALB/c athymic mice. Hyperimmune serum and sensitized splenocytes were prepared by repeated immunization of BALB/c mice with porcine lymph node cells. Splenic T cells were enriched by nylon wool column separation. Tissues were examined by immunohistology using murine- and porcine specific monoclonal antibodies. RESULTS: Porcine islets survived in nude mice for > 100 days with high levels of circulating porcine C-peptide and maintenance of normoglycemia. Injection of the hyperimmune sera (IgG) into normoglycemic nude mice bearing porcine islets for > 70 days failed to induce rejection despite the continued presence of circulating anti-porcine cytotoxic antibody. Injection of sensitized T cells caused acute rejection of long-term (>140 days) porcine islets, whereas injection of naive T cells had no effect. Histologically, porcine islets removed from mice treated with hyperimmune serum showed no staining for IgG. Long-surviving porcine islet grafts showed strong staining for interleukin (IL)-10 and a lesser amount of IL-4 but no staining for IL-2 or interferon-gamma. Although fresh porcine islets were positive for swine leukocyte antigen class 1 antigen and intercellular adhesion molecule (ICAM)-1 but negative for mouse platelet endothelial cell adhesion molecule and ICAM-2, long-surviving porcine islets showed positive endothelial staining for mouse platelet endothelial cell adhesion molecule and ICAM-2. CONCLUSIONS: Established islet xenografts are resistant to hyperimmune serum as a result of a lack of target endothelial antigens, whereas they remain susceptible to rejection caused by primed T cells. Local production of Th2 cytokines may explain the inability of long-surviving islet xenografts to activate injected naive T cells. PMID- 10515392 TI - Complement deposition in early cardiac transplant biopsies is associated with ischemic injury and subsequent rejection episodes. AB - BACKGROUND: Prolonged warm or cold ischemia is associated with poor survival of cardiac transplants, and ischemic changes in early posttransplantation endomyocardial biopsies correlate with the later development of chronic rejection. In animal models, tissue ischemia has been shown to activate complement. METHODS: To determine whether ischemic changes in endomyocardial biopsies were associated with complement deposition, biopsies obtained 1-3 weeks after transplantation from 33 patients were evaluated immunohistologically for C4d and C3d deposition as well as for IgM, IgG, and IgA. The histological changes associated with ischemic injury were scored independently, using previously reported criteria without knowledge of the immunohistochemical results. RESULTS: Diffuse capillary and pericapillary deposition of C4d or C3d were detected in endomyocardial biopsies of 14 of the 33 patients. The majority of biopsies (79%) with C4d or C3d deposits had histological evidence of ischemic injury, including eight of the nine biopsies containing both C4d and C3d deposition. In contrast, only 8 of 18 (45%) of the biopsies without C4d or C3d deposition had ischemic injury. Only trace amounts of IgM and no IgG or IgA were demonstrable in the biopsies. Only 2 of the 14 biopsies with C4d or C3d deposition had evidence of moderate acute rejection, whereas 5 of the 18 biopsies without C4d or C3d deposition had moderate acute rejection. However, C4d and C3d deposition did correlate with repeated acute rejection episodes on subsequent biopsies. CONCLUSIONS: Thus, ischemic changes are associated with the activation of complement. Complement activation may in turn promote tissue injury and provide a potential target for future treatment. PMID- 10515393 TI - A nonhealing ulcer diagnosed as extramedullary plasmocytoma of the limb eight years after cardiac transplantation. AB - A 63-year-old man was hospitalized for a nonhealing ulcer of the left lower leg that appeared 8 years after orthotopic cardiac transplantation under immunosuppressive therapy including cyclosporine. Serum protein electrophoresis, immunofixation, and urinalysis revealed a monoclonal gammopathy IgG kappa. The final diagnosis of an extramedullary plasmocytoma was made by biopsy of the ulcer, which showed formations of plasmablastic cells. We report a rare case of extramedullary plasmocytoma as a posttransplantational malignancy. PMID- 10515394 TI - Cell-mediated graft rejection observed in two lines of human histocompatibility leukocyte antigen class I transgenic mice. AB - BACKGROUND: Human histocompatibility leukocyte antigen (HLA) class I molecules are essential for graft rejection. However, to determine the specific role of these molecules in clinical situations is difficult. We investigated the applicability of HLA class I transgenic mice (C3H.B35 and C3H.B51) for elucidation of the role of HLA class I molecules. METHODS: Skin or heart grafts were transplanted. Cytotoxic T cells (CTL) of C3H.B51 against C3H.B35 were generated and their cytotoxicity against various transfectant cell lines was determined. RESULTS: C3H.B35 skin and heart grafted to C3H.B51 were rejected within 17 and 28 days, respectively. Cytotoxic T cells generated from C3H.B51 showed cytotoxicity against a HLA-B*3501-transfectant cell line that did not express H-2 molecule, which indicates that these cytotoxic T cells recognize HLA B35 molecules directly without H-2 restriction. CONCLUSION: Our results suggest that C3H.B51 recognize C3H.B35 grafts as allo-MHC class I-incompatible grafts, and these mice are valuable to elucidate the role of HLA class I molecules in transplantation. PMID- 10515395 TI - Report of ERS Task Force: guidelines for measurement of acellular components and standardization of BAL. PMID- 10515396 TI - Long acting inhaled beta2-agonists: anti-inflammatory effects not evident during treatment of day to day asthma. PMID- 10515397 TI - Different cytokine profiles in cryptogenic fibrosing alveolitis and fibrosing alveolitis associated with systemic sclerosis: a quantitative study of open lung biopsies. AB - Differences in the inflammatory response and prognosis of cryptogenic fibrosing alveolitis (CFA) and that associated with systemic sclerosis (FASSc) are beginning to emerge. It is hypothesized that these differences may be reflected in a distinct pattern of T-helper (Th)-1 and Th-2-type cytokines. Open lung biopsies were obtained from clinically well-documented cases of CFA and FASSc and, as a control, compared with grossly and histologically normal parenchyma obtained from smokers whose lungs were resected for cancer (n=5 in each group). In situ hybridization (ISH) was applied to the samples using anti-sense and sense 35S-labelled riboprobes to detect messenger ribonucleic acid (mRNA) for interleukins (IL)-2, IL-4, IL-5 and interferon (IFN)-gamma. Between 52-91% of cells expressing the cytokines studied were present in the alveolar interstitium rather than in lumenal cells or the alveolar epithelial lining. The highest values for all four cytokines were present in the patients with FASSc, i.e., 22 39 ISH positive cells x mm(-2) alveolar tissue compared with 1-19 cells x mm(-2) and 4-5 cell x mm(-2) in CFA and control subjects, respectively. Whereas the proportions of the four cytokines in FASSc were similar to the control subjects, IL-4 and IL-5 predominated significantly in CFA (p<0.001). For example, the ratio of IL-5 to IFN-gamma was 22:1 in CFA, significantly higher than in the cases with FASSc (2:1) or the control subjects (4:1) (p<0.05). In conclusion, cryptogenic fibrosing alveolitis is an inflammatory condition which is characterized, like asthma, by a predominance of gene expression for T-helper-2-type regulatory cytokines, whereas cryptogenic fibrosing alveolitis associated with systemic sclerosis appears to have a distinct mixed T-helper-1/T-helper-2 functional phenotype and a greater number of cells expressing each of these pro-inflammatory cytokines. PMID- 10515398 TI - Interval versus continuous training in patients with severe COPD: a randomized clinical trial. AB - Limited information is available regarding the physiological responses to different types of exercise training in patients with severe chronic obstructive pulmonary disease (COPD). The aim of this study was two fold: firstly, to investigate the physiological response to training at 60% of achieved peak load in patients with severe COPD; and secondly to study the effects of interval (I) versus continuous (C) training in these patients. Twenty-one patients with COPD (mean+/-SD forced expiratory volume in one second: 37+/-15% of predicted, normoxaemic at rest) were evaluated at baseline and after 8 weeks' training. Patients were randomly allocated to either I or C training. The training was performed on a cycle ergometer, 5 days a week, 30 min daily. The total work load was the same for both training programmes. C training resulted in a significant increase in oxygen consumption (V'O2) (17%, p<0.05) and a decrease in minute ventilation (V'E)/V'O2 (p<0.01) and V'E/carbon dioxide production (V'CO2) (p<0.05) at peak exercise capacity, while no changes in these measures were observed after interval training. During submaximal exercise a significant decrease was observed in lactic acid production, being most pronounced in the C trained group (-31%, p<0.01 versus -20%, p<0.05). Only in the I-trained group did a significant increase in peak work load (17%, p<0.05) and a decrease in leg pain (p<0.05) occur. Training did not result in a significant improvement in lung function, but maximal inspiratory mouth pressure increased in both groups by 10% (C: p<0.05) and 23% (I: p<0.01). The present study shows a different physiological response pattern to interval or continuous training in chronic obstruction pulmonary disease, which might be a reflection of specific training effects in either oxidative or glycolytic muscle metabolic pathways. Further work is required to determine the role of the different exercise programmes and the particular category of patients for whom this might be beneficial. PMID- 10515399 TI - Noninvasive measurement of respiratory muscle performance after exhaustive endurance exercise. AB - The use of noninvasive techniques to measure respiratory muscle performance after different types of endurance exercise has not been entirely successful, as the results have not consistently indicated diminished performance for similar types of exercise. The aim of the present study was 1) to compare different, noninvasive methods to assess respiratory muscle performance before and after an exhaustive cycling endurance test (which has previously been shown to induce diaphragmatic fatigue) and 2) to determine which of the tests best reflect published results of measurements of diaphragmatic fatigue. Twelve healthy subjects participated in the study and performed three different test series in a random order on three different days. These tests were performed before, and 5, 40 and 75 min after an exhausting task (a cycling endurance run at 85% of maximal oxygen uptake (V'O2,max)). The tests of the three test series were 1) breathing against a constant inspiratory resistance to task failure, 2) determination of 12 min sustained ventilatory capacity, and 3) spirometric and maximal inspiratory and expiratory mouth pressure measurements. The only measurement that was affected by exhaustive cycling was the time to task failure breathing against inspiratory resistance. It was significantly reduced from (mean+/-sD) 364+/-88 s before exercise to 219+/-122 s at 5 min after cessation of exercise. It is concluded that the constant-load resistive breathing test to task failure is the only noninvasive respiratory muscle performance test evaluated in this study which shows a decrease in respiratory muscle performance after exhaustive endurance exercise. PMID- 10515400 TI - Six minute walking distance in healthy elderly subjects. AB - The six minute walking distance (6MWD) test is a commonly used test to estimate functional exercise capacity in patients with chronic diseases including chronic obstructive lung disease. Surprisingly, no attempt has been made to establish normal values for the 6MWD. The aim of this study, therefore, was to evaluate the 6MWD in healthy elderly volunteers and to evaluate its determining factors. Fifty one healthy subjects aged 50-85 yrs volunteered to participate in the trial. All subjects were free of diseases that could interfere with performance in a walking test. Tests were performed in a quiet 50-m long hospital corridor. Patients were encouraged every 30 s to continue walking as quickly as possible. Walking distance averaged 631+/-93 m and was 84 m greater in the male compared to female subjects (p<0.001). The 6MWD showed significant correlations with age (r=-0.51, p<0.01) and height (r=0.54, p<0.01). Stepwise multiple regression analysis showed that age, height, sex and weight were independent contributors to the 6MWD in healthy subjects, thus explaining 66% of the variability. It is concluded that the six minute walking distance can be predicted adequately using a clinically useful model in healthy elderly subjects. Its variability is explained largely by age, sex, height and weight. Results of the six minute walking distance may be interpreted more adequately if expressed as a percentage of the predicted value. PMID- 10515401 TI - The long-acting beta2-agonist salmeterol xinafoate: effects on airway inflammation in asthma. AB - Salmeterol xinafoate is an inhaled long-acting beta2-adrenoceptor agonist recently introduced for the treatment of asthma. Both in vitro and animal studies suggest that it may have anti-inflammatory activities of benefit in this disease. To assess this directly, the effects of 6 weeks' treatment with salmeterol on indices of clinical activity, airway dysfunction and inflammation in subjects with stable atopic asthma were investigated. In a double blind study, asthmatic patients were randomized to 6 weeks' treatment with either salmeterol 50 microg twice daily (n=14) or placebo (n=12). They underwent bronchoscopy with bronchoalveolar lavage (BAL) and bronchial biopsy immediately before starting treatment and again after 6 weeks. Treatment with salmeterol improved clinical indices of asthma activity, but there were no changes in BAL differential cell counts or mediator levels, and no change in T-cell numbers or activation status. In the biopsy specimens there were no changes in numbers of inflammatory cells, sub-basement membrane collagen deposition or mast cell degranulation. Regular treatment with salmeterol improves clinical indices of asthma but has no effect on the underlying inflammatory process. These findings strengthen guideline recommendations that long-acting beta2-agonists should not be prescribed as sole antiasthma medication. PMID- 10515402 TI - Tolerance to beta-agonists during acute bronchoconstriction. AB - Previous reports suggest that regular use of beta-agonists does not lead to tolerance to their bronchodilator effects. However, most studies have been conducted in stable asthma. This study investigates whether bronchodilator tolerance can be demonstrated during acute bronchoconstriction. Thirty-four asthmatic subjects were treated with 6 weeks inhaled terbutaline (1 mg q.i.d.), budesonide (400 microg, b.i.d.), both drugs or placebo in a randomized, double blind, cross-over study. After each treatment methacholine was administered to induce a 20% fall in the forced expiratory volume in one second (FEV1). The response to inhaled salbutamol 100, 100, 200 microg at 5 min intervals) was then measured. Dose-response curves were compared using an analysis of covariance. Pre methacholine FEV1, the highest pre-methacholine FEV1, the fall in FEV1 induced by methacholine and the logarithm of the provocative dose of methacholine required to induce the 20% fall in FEV1 (PD20) were used as covariates. There was a significantly reduced response to salbutamol after 6 weeks terbutaline treatment: the mean (95% confidence intervals (CI)) area under the dose-response curve was reduced by 36% (24, 47) compared to placebo (p<0.0001). The reduction in bronchodilator response was not affected by concomitant treatment with budesonide. Significant tolerance to the bronchodilator effect of inhaled beta agonists may be demonstrated when tested during acute bronchoconstriction. Continuous treatment with inhaled beta-agonists may lead to a reduced response to emergency beta-agonist treatment during asthma exacerbations. PMID- 10515403 TI - International variations in asthma treatment compliance: the results of the European Community Respiratory Health Survey (ECRHS). AB - Noncompliance to medication is a major barrier to effective asthma management. Its real extent and geographical variation throughout the world are not yet known. The data on compliance, collected in the framework of the European Community Respiratory Health Survey (ECRHS) on 1771 subjects (aged 20-44 yrs) with current asthma identified in 14 countries, offer a unique opportunity to assess the extent of noncompliance and its variation across countries. The median percentage of current asthmatics who had received a medical prescription at least once was 95%. The compliance of those patients who had received a medical prescription was found to be low in all countries (median 67%) but with wide variations, the rate ranging from 40% (USA) to 78% (Iceland). During exacerbations patients' rate of compliance increased to 72%. Age was the only variable which influenced compliance to treatment. A significant, although weak, negative correlation was found between patients' compliance and rate of hospital casualty department or emergency room admissions. This study documents that compliance to the treatment of asthma is poor worldwide and that there are large variations between countries. These results emphasize the necessity for further efforts to improve patients' education and to promulgate the international guidelines. PMID- 10515404 TI - Dynamic hyperinflation and flow limitation during methacholine-induced bronchoconstriction in asthma. AB - Although persistent activation of the inspiratory muscles and narrowing of the glottic aperture during expiration have been indicated as relevant mechanisms leading to dynamic hyperinflation in acute asthma, expiratory flow limitation (EFL) has recently been proposed as a possible triggering factor for increasing endexpiratory lung volume (EELV). To establish whether the attainment of maximal flow rate during tidal expiration could elicit dynamic elevation of EELV, breathing pattern, change in EELV by measuring inspiratory capacity (IC) and occurrence of EFL by the negative expiratory pressure (NEP) method were monitored in 10 stable asthmatic subjects during methacholine-induced, progressive bronchoconstriction in seated position. Change in dyspnoea was scored using the Borg scale. At maximum response forced expiratory volume in one second (FEV1) fell on average by 45+/-2% (p<0.001 versus control), while IC decreased 29+/-2%, (by 0.89+/-0.07 L, (p<0.01 versus control)). Only 2 subjects exhibited EFL at the end of methacholine challenge. In 7 subjects EELV started to increase before the occurrence of EFL. Dyspnoea, which increased from 0.2+/-0.1 to 5.5+/-1.0 (Borg scale) at maximum response (p<0.001), was significantly related to the level of bronchoconstriction as assessed by change in (delta)FEV1 (r=0.72; p<0.001) and to dynamic hyperinflation as measured by deltaIC (r=0.50; p<0.001). However, for both deltaFEV1 and deltaIC the slope of the relationship with increasing dyspnoea was highly variable among the subjects. It is concluded that in acute methacholine-induced bronchoconstriction, dynamic hyperinflation may occur in the absence of expiratory flow limitation and that expiratory flow limitation does not represent the triggering factor to generate dynamic hyperinflation. In these circumstances, dyspnoea appears to be related to the increase in end-expiratory lung volume and not to the onset of expiratory flow limitation. PMID- 10515405 TI - Development of wheezing in patients with cough variant asthma during an increase in airway responsiveness. AB - Two theories explaining the mechanism for the manifestation of cough without wheeze in patients with cough variant asthma (CVA) are either a higher wheezing threshold or a milder degree of airway hyperresponsiveness. A significant proportion of patients diagnosed as having CVA eventually develop wheezing. The aim of this study was to investigate whether this change in the manifestation of asthma was associated with a decrease in wheezing threshold and/or an increase in airway hyperresponsiveness. Thirty-six children (7-15 yrs) with CVA were prospectively studied for 4 yrs. Bronchial provocation tests with methacholine using the stepwise increasing concentration technique were performed annually to measure the provocative cumulative dose producing a 20% fall in forced expiratory volume in one second (PD20). Wheezing thresholds were additionally determined at the initiation of and the end of the study (development of wheezing, or after 4 yrs). Sixteen (Group 1) of 29 patients available for the follow-up developed clinical wheezing during the period; 13 patients (Group 2) stayed as CVA or their cough resolved. There was no significant change in wheezing thresholds from the initiation to the end of the study (Group 1: 40.9+/-8.2% versus 40.2+/-8.3%; Group 2: 41.4+/-7.1% versus 40.1+/-7.3%). Methacholine PD20 (geometric mean, range of 1 SD), expressed as breath unit (BU), significantly decreased in Group 1 patients as they developed wheezing (initial versus wheezing year: 60.8 BU, 29.2 126.5 versus 32.8 BU, 11.5-93.3; p<0.01), whereas the value did not change in Group 2 patients (initial versus after 4 yrs: 85.3 BU, 45.2-161.1 versus 84.3 BU, 39.7-179.1; NS). The results suggest that an increase in airway hyperresponsiveness, but not a decrease in wheezing threshold, may have a pathogenetic role in the development of wheezing during the course of cough variant asthma in childhood. PMID- 10515406 TI - Interleukin-10 level in sputum is reduced in bronchial asthma, COPD and in smokers. AB - Interleukin (IL)-10 is a potent regulatory cytokine that decreases inflammatory responses. This study investigated whether IL-10 levels in the airway are decreased in chronic airway inflammation associated with asthma or chronic obstructive pulmonary disease (COPD). Sputum was obtained from 12 healthy nonsmokers, 10 healthy smokers, 16 asthmatic patients and seven patients with COPD by means of the sputum-induction method. The IL-10 level was measured via enzyme-linked immunosorbent assay and immunocytochemical analysis. The IL-10 level in sputum was significantly lower in asthma and COPD patients and healthy smokers compared with that in healthy nonsmokers (nonsmokers, 68.0+/-11.3; smokers, 45.3+/-7.8; asthma, 26.7+/-4.0; COPD, 18.0+/-2.3 pg x mL(-1); p<0.05 for nonsmokers versus the other groups). The percentage of IL-10-positive cells in the sputum was also significantly lower in asthma and COPD and in smokers (nonsmokers, 13.2+/-1.7; smokers, 6.4+/-1.8; asthma, 5.4+/-3.5; COPD, 3.5+/-1.6%; p<0.05 for nonsmokers versus the other groups). The IL-10-positive cell appeared morphologically to be the macrophage. These data suggest that the reduced level of interleukin-10 within the airways plays a role in the pathogenesis of chronic airway inflammation in asthma and chronic obstructive pulmonary disease. PMID- 10515407 TI - Passive sensitization of human airways increases responsiveness to leukotriene C4. AB - Passive sensitization of human airways in vitro causes increased responsiveness to histamine and induces specific immunoglobulin (Ig)E-dependent contractile responsiveness to allergen. Leukotrienes (LTs) and, to a lesser extent, histamine are the major mediators of allergen-induced contraction. Since it is unclear whether passively sensitized airways are also hyperresponsive to cysteinyl leukotrienes, this study investigated the effect of passive sensitization on LTC4 , in addition to histamine- and allergen-induced contractions in vitro. Bronchial rings from nine nonatopic patients were sensitized overnight with serum containing high levels of total IgE (>250 U x mL(-1)) and allergen-specific IgE against Dermatophagoides farinae (fluorescence allergosorbent test) (FAST class > or =3). The potency (-log10 of the mediator concentration causing a half maximal response (pEC50) of histamine was significantly increased in serum-sensitized tissues compared to nonsensitized controls ((mean+/-SEM) pEC50 5.20+/-0.27 versus 5.64+/-0.18; p=0.02) and maximal contractions were enhanced (877+/-47 versus 543+/-51 mg; p<0.0001). Similarly, the potency of LTC4 was significantly increased in sensitized compared to nonsensitized bronchial rings (pEC50 9.37+/ 0.20 versus 8.66+/-0.26; p=0.004); maximal contractions were also enhanced (811+/ 57 versus 361+/-86 mg; p<0.0001). These data demonstrate that passive sensitization of human airways induces an increase not only in histamine but also in leukotriene responsiveness. Therefore, it might be speculated that allergen responses in sensitized airways are effected through a combination of increased mediator release from inflammatory cells and increased responsiveness of airway smooth muscle. PMID- 10515408 TI - Sensory nerve activation in airway microvascular permeability in guinea-pig late allergic response. AB - Because both bradykinin and tachykinins have a potent inflammatory action, these molecules may be involved in the late allergic response. The role of these molecules in airway microvascular permeability during the late allergic response in sensitized guinea-pigs was investigated. Three weeks after ovalbumin sensitization, the animals were pretreated with bradykinin B2 receptor antagonist HOE 140, neurokinin 1 receptor antagonist CP 96,345 or vehicle, 30 min before the ovalbumin inhalation challenge. The occurrence of the late allergic response was determined by a two-fold increase in the transpulmonary pressure from the baseline values. The microvascular permeability in the trachea was assessed by an index defined as the ratio of the area of vasculature labelled by Monastral blue dye (area density %). Significant microvascular permeability and eosinophil accumulation were observed during the late allergic response. Both the bradykinin and substance P concentrations in the bronchoalveolar lavage fluid were increased during the late allergic response. Pretreatment with HOE 140 suppressed the substance P elevation. Both HOE 140 and CP 96,345 also inhibited the airway microvascular permeability during the late allergic response without affecting the eosinophil accumulation in the airways. These findings suggest that bradykinin-mediated sensory nerve activation may play a role in microvascular permeability during the late allergic response in guinea-pigs. PMID- 10515409 TI - Quantification of the dose of inhaled flour: relation with nonspecific bronchial and immunological reactivities. AB - The aim of this study was to investigate the relationship between specific bronchial reactivity and respective nonspecific bronchial and immunological reactivities. Twenty-one patients underwent bronchial challenges with lactose and flour. The aerosol of particles was generated by a computer-controlled aerosolizer. Specific bronchial challenge results were expressed as the provocative dose of flour (PDf) that caused a 20% or 15% decrease in the forced expiratory volume in one second (FEV1). For each subject, the decrease in FEV1 observed during the challenge with flour was compared with the calculated lower limit of the 99.7% confidence interval for the lactose challenge. The subjects also underwent a nonspecific challenge with methacholine and a measurement of the specific immunoglobulin E against wheat. The inhalation of lactose did not significantly affect FEV1. Nine subjects had high reactivity to wheat flour with a PDf20 <400 microg. Five subjects had intermediate reactivity: FEV1 fell by <20% but by significantly more than that in the test with lactose. For 7 subjects, there was no significant change in FEVI for inhaled doses of flour over 1390 microg. The results for specific bronchial challenge were significantly correlated with those for the methacholine test (p<0.02). Positive skin tests and specific immunoglobulin E against wheat were observed more frequently in the high reactivity group. Specific bronchial challenge can be performed safely to establish precise dose-response curves. The provocative dose of flour causing a 20% decrease in forced expiratory volume in one second is useful for evaluating the degree of specific reactivity but is not suitable in cases of intermediate reactivity in which comparison with the lactose test is necessary. Specific reactivity is probably a function of immunological and nonspecific bronchial reactivities. PMID- 10515410 TI - Bronchial hyperresponsiveness and adult onset wheeze: the influence of atopy. AB - The commonly held belief that adult onset wheezing illness is primarily nonatopic in nature suggests that the role of atopy in the pathophysiology of bronchial hyperresponsiveness (BHR) in adult onset wheeze may be minimal. This study examined risk factors for BHR (BHR: provocative dose causing a 20% fall in forced expiratory volume in one second PD20 < or =16.38 micromol methacholine) among 82 subjects with adult onset wheeze and among 191 subjects who had never wheezed. Subjects were identified from a cohort of subjects aged 39-45 yrs who were known to have had no childhood wheeze and who were involved in a 30 yr follow-up survey. Risk factors for BHR were examined among all subjects with BHR and among subjects with BHR stratified according to whether or not they had ever wheezed. The prevalence of BHR was 40% (33/82) among the subjects with adult onset wheeze and 11% (21/191) among the subjects who had never wheezed. Lower baseline lung function (odds ratio (OR) = 0.94; 95% confidence interval (CI) = 0.92-0.97 per unit forced expiratory volume (FEV1)% predicted) and atopy (OR = 7.23; CI = 2.53 20.64 for all three measures of atopic compared to nonatopic) were associated with BHR, while smoking and family history showed no statistically significant relation to BHR. This pattern was also apparent in analyses stratified by symptom status. A family history of atopy increased the risk that BHR was accompanied by wheezing symptoms (OR = 4.75; CI = 1.53-14.72 for more than one affected relative compared to no affected relatives). These findings suggest that atopy is associated with bronchial hyperresponsiveness in adults known to have had no childhood wheeze. A familial factor reflecting genetic influences and/or shared environmental factors may influence whether bronchial hyperresponsiveness is associated with symptoms. PMID- 10515411 TI - Distinct sputum cytokine profiles in cystic fibrosis and other chronic inflammatory airway disease. AB - The dominant role of inflammation in airways disease progression in cystic fibrosis (CF) is now well established and, based on recent findings, the possibility of an inappropriate inflammatory response in the lung of patients with CF has emerged. In order to characterize this response, the aim of the present work was to evaluate the levels of a number of pro- and anti-inflammatory cytokines in the sputum of CF children and to compare these levels to those observed in the sputum from non-CF children with diffuse bronchiectasis (DB). Three groups of patients were investigated: a group of 25 CF children (mean age: 12.2 yrs), a group of 10 non-CF children with DB (mean age 11.5 yrs), and a group of five healthy young adults (mean age 24 yrs). Elevated concentrations of pro inflammatory cytokines, tumour necrosis factor (TNF)-alpha, interleukin (IL) 1beta and IL-8 were found in children with CF and in non-CF children with DB, with significantly higher concentrations of IL-1beta in CF children. Analysis of the natural anti-inflammatory cytokine IL-1 receptor antagonist (IL-1ra) and type II TNF soluble receptor (sTNFRII) concentrations showed distinct patterns, with elevated levels of both inhibitors in CF patients, whereas only sTNFRII was found to be increased in non-CF children with DB. IL-10 data indicated low concentrations in the CF group. In all CF children, the concentrations of IL-6 in the airways were extremely low, independent of the clinical, bacteriological or functional status. By contrast, significantly increased IL-6 levels were found in non-CF children with DB. These results document distinct cytokine profiles in cystic fibrosis patients and noncystic fibrosis patients. They also suggest that impairment of interleukin-6 expression may represent an important component of the excessive inflammatory response observed in cystic fibrosis. PMID- 10515412 TI - Therapeutic drug monitoring in patients with cystic fibrosis and mycobacterial disease. AB - Cystic fibrosis (CF) patients require higher dosages of many antibiotics. The relapse of tuberculosis in one CF patient, and the repeated growth of Mycobacterium avium-intracellulare in another, despite conventional therapy, raised the question of whether the serum levels of the antimycobacterial drugs were adequate. Antimycobacterial drug serum concentrations were assayed in 10 CF patients with pulmonary mycobacterial disease. Serum levels below the proposed target range were seen 2 h after drug intake in the initial four patients treated: for rifampicin in 2/3, ethambutol in 3/4 and for clarithromycin in 2/3 patients, despite standard dosages. Reassays after dose adjustment and assays in six other patients showed that adequate levels were not achieved 4 h after clarithromycin in 3/5, ethambutol in 1/5, ciproflaxacin in 1/2 and ofloxacin in 2/2 patients. The patient with relapse of tuberculosis and the patient with continuous growth of M. avium-intracellulare improved and became culture negative after dose adjustment. Low drug serum levels is one reason for therapy failure in cystic fibrosis patients with mycobacterial disease. Therapeutic drug monitoring is recommended. PMID- 10515413 TI - Allergy to laboratory animals in children of parents occupationally exposed to mice, rats and hamsters. AB - Sensitization to laboratory animals (LA) has a high prevalence among laboratory workers. It is unknown whether transportation of LA allergens can be a risk factor for sensitization of subjects outside the laboratory environment. The aim of the study was to investigate the prevalence of sensitization to LA among children whose parents were and were not occupationally exposed to LA. The first group consisted of 50 children (age 12.3+/-4.3 yrs) whose parents were occupationally exposed to mice, rats and hamsters. The second group consisted of 40 children (age (mean+/-SD) 10.8+/-3.0 yrs) whose parents were not occupationally exposed to LA. Children having LA at home were eliminated from the study. All children responded to a questionnaire, underwent spirometry and were also tested with skin prick tests with the use of common allergens and prick tests with hair extracts from mouse, hamster and rat. Total immunoglobulin (Ig)E levels and the presence of specific IgE against LA were also estimated. Children of parents occupationally exposed to LA presented significantly more positive skin prick tests against allergens from the hair of laboratory animals compared to children of nonexposed parents. Five children from the first group were also found to have specific IgE against LA, with three of these five children complaining of rhinitis and cough while visiting their parents' workplace. It is concluded that the observed increased sensitization to laboratory animals among children of occupationally exposed parents could be the result of poor hygienic conditions at their parents' workplace. Hence, parents' job seems to be an additional risk factor of sensitization and should be taken into consideration when recording an allergic history. PMID- 10515414 TI - Paralysis of ventilated newborn babies does not influence resistance of the total respiratory system. AB - Paralysis with pancuronium bromide is used in newborn infants to facilitate ventilatory support during respiratory failure. Changes in lung mechanics have been attributed to paralysis. The aim of this study was to examine whether or not paralysis per se has an influence on the passive respiratory mechanics, resistance (Rrs) and compliance (Crs) of the respiratory system in newborn infants. In 30 infants with acute respiratory failure, Rrs was measured during paralysis with pancuronium bromide and after stopping pancuronium bromide (group A). Rrs was also measured in an additional 10 ventilated infants in a reversed fashion (group B): Rrs was measured first in nonparalysed infants and then they were paralysed, mainly for diagnostic procedures, and the Rrs measurement repeated. As Rrs is highly dependent on lung volume, several parameters, that depend directly on lung volume were recorded: inspiratory oxygen fraction (FI,O2), arterial oxygen tension/alveolar oxygen tension (a/A) ratio and volume above functional residual capacity (FRC). In group A, the Rrs was not different during (0.236+/-0.09 cmH2O x s x mL(-1)) and after (0.237+/-0.07 cmH2O x s x mL( 1)) paralysis. Also, in group B, Rrs did not change (0.207+/-0.046 versus 0.221+/ 0.046 cm x s x mL(-1) without versus with pancuronium bromide). FI,O2, a/A ratio and volume above FRC remained constant during paralysis. These data demonstrate that paralysis does not influence the resistance of the total respiratory system in ventilated term and preterm infants when measured at comparable lung volumes. PMID- 10515415 TI - Effect of fluticasone propionate and salmeterol on Pseudomonas aeruginosa infection of the respiratory mucosa in vitro. AB - The purpose of this study was to investigate the effect of the corticosteroid, fluticasone propionate (FP), on Pseudomonas aeruginosa infection of the respiratory mucosa of an organ culture model in vitro. Organ cultures infected with P. aeruginosa had significantly (p< or =0.05) elevated levels of mucosal damage and significantly (p< or =0.05) less ciliated cells compared to controls. Preincubation of tissue with FP (10(-6) or 10(-5) but not 10(-7) M) prior to P. aeruginosa infection significantly (p< or =0.05) reduced the bacterially induced mucosal damage in a concentration-dependent manner. FP (10(-5) M) also significantly (p< or =0.05) prevented loss of ciliated cells. FP did not alter the density of bacteria adherent to the different mucosal features of the organ cultures, but did reduce total bacterial numbers due to the reduced amount of damaged tissue, which is a preferred site of P. aeruginosa adherence. It has previously been shown that the long-acting beta2-agonist salmeterol (4 x 10(-7)M) also reduces the mucosal damage caused by P. aeruginosa infection, probably via elevation of intracellular cyclic adenosine monophosphate concentrations. Preincubation of tissue with both 10(-7)M FP and 10(-7)M salmeterol, concentrations at which they did not by themselves influence the effect of P. aeruginosa infection, significantly (p< or =0.05) reduced P. aeruginosa-induced loss of cilia. However, there was no additional benefit from adding 4 x 10(-7)M salmeterol to 10(-6)M FP. In conclusion fluticasone propionate reduced mucosal damage caused by P. aeruginosa infection in vitro and preserved ciliated cells. There was a synergistic action with salmeterol in the preservation of ciliated cells. PMID- 10515416 TI - Comparative validation of prognostic rules for community-acquired pneumonia in an elderly population. AB - The aim of the study was to validate the prediction rule of M.J. Fine and coworkers for clinical outcome variables and three prognostic rules for the individual outcome of community-acquired pneumonia in an elderly population (rule 1: respiratory frequency > or =30 breaths x min(-1), diastolic blood pressure < or =60 mmHg, blood urea nitrogen >7 mM; rule 2: respiratory frequency > or =30 breaths x min(-1), diastolic blood pressure < or =60 mmHg, mental confusion; and rule 3: systolic blood pressure < or =80 mmHg, cardiac frequency > or =90 beats x min(-1), lactate dehydrogenase activity > or =260 IU x L(-1); death was predicted in the presence of at least two of three parameters). Overall 168 consecutive episodes of community-acquired pneumonia in patients aged > or =65 yrs and hospitalized in a primary care hospital were studied prospectively. Fine's rule was tested for its ability to predict length of hospital stay, requirement for intensive care unit (ICU) admission and death. For the three prognostic rules of individual outcome, performance regarding predicting death was determined. Mortality was 17/168 (10%). Fine's rule accurately predicted length of stay, the requirement for ICU admission and the risk of death from pneumonia as compared to the original derivation and validation cohorts. All three rules achieved moderate to-high specificity (73%, 88% and 80%, respectively) and high negative predictive values (95%, 94% and 93%, respectively) but had a low sensitivity (65%, 47% and 47%, respectively). Rule 2 most closely reflected the risk of death from pneumonia when Fine's classification was used as reference. Fine's rule proved to give valid estimations regarding clinical outcome variables of community-acquired pneumonia in the elderly. The prognostic rules may be useful in determining individual patients at lower risk of death caused by pneumonia. PMID- 10515417 TI - Clinical significance of MCP-1 levels in BALF and serum in patients with interstitial lung diseases. AB - It has previously been reported that the expression of monocyte chemoattractant protein-1 (MCP-1) in the lung tissues of patients with idiopathic pulmonary fibrosis (IPF) was different from that in the tissues of patients with other interstitial lung diseases (ILDs). The aim of this study was to determine whether this difference reflects the amount of MCP-1 in the bronchoalveolar lavage fluid (BALF) or serum of patients with ILD, and whether such a correlation, if it exists, is clinically useful. MCP-1 concentrations in the BALF and sera were evaluated in 86 patients with ILDs including IPF, acute interstitial pneumonia, interstitial pneumonia with collagen vascular disease (IP-CVD), chronic interstitial pneumonia (CIP), bronchiolitis obliterans-organizing pneumonia, sarcoidosis, hypersensitivity pneumonitis, and in 10 normal healthy volunteers who were controls (NC). BALF MCP-1 levels were significantly elevated in the IPF, IP-CVD, CIP and sarcoidosis groups compared with the NC group. The level in the IPF group was significantly higher than that in any other patient group. Serum MCP-1 levels in the IPF, IP-CVD, CIP and sarcoidosis groups were significantly higher than the NC group. No statistical difference was found in serum MCP-1 levels between the IPF, IP-CVD and CIP groups. BALF MCP-1 levels were significantly higher than serum MCP-1 levels in the IPF group and lower than in the IP-CVD and CIP groups. Serum MCP-1 levels correlated with the clinical course of ILD treated with corticosteroid therapy. These results show that measurement of monocyte chemoattractant protein-1 levels in both bronchoalveolar lavage fluid and serum may be helpful in discriminating idiopathic pulmonary fibrosis from other types of interstitial lung disease and that monitoring of serum monocyte chemoattractant protein-1 may be useful for predicting the clinical course of interstitial lung diseases. PMID- 10515418 TI - Elevated bronchoalveolar concentrations of MCP-1 in patients with pulmonary alveolar proteinosis. AB - Pulmonary alveolar proteinosis (PAP) is a rare disease of unknown aetiology characterized by accumulations of lipoproteinaceous material within the alveoli. The alveolar macrophages become increasingly foamy, and are thought to have a role in the pathogenesis of PAP. However, the mechanisms of macrophage recruitment are unclear. In the bronchoalveolar lavage fluid (BALF) of four patients with PAP and 20 normal control subjects, the following were examined: the monocyte chemotactic activity due to the chemokine monocyte chemoattractant protein (MCP)-1 with the use of a chemotactic chamber assay, the levels of MCP-1 by enzyme-linked immunosorbent assay, and the MCP-1 expression on lavage cells by immunocytochemistry and in situ hybridization. The monocyte chemotactic activity in the BALF of the PAP patients was markedly elevated, and the activity was completely absorbed by treatment with anti-MCP-1. The MCP-1 levels in the BALF were surprisingly high in the PAP group (25,100+/-472 pg x mL(-1)), whereas low levels of MCP-1 were detected in the normal control subjects (mean: never smokers 4.8; smokers 10.4 pg x mL(-1)). MCP-1 protein and messenger ribonucleic acid were expressed by macrophages from the PAP patients, and the expression was reduced according to foaming of the cells; there were monocyte-like macrophages with strong expression, small foamy cells with moderate expression, large foamy cells with a faint expression of MCP-1, and ghost cells with no expression. However, the increase of macrophage number in the PAP BALF was relatively small. These data suggest that monocyte chemoattractant protein(-1) expression by alveolar macrophages represents an amplification mechanism for the recruitment of additional macrophages to the alveoli in pulmonary alveolar proteinosis. It is possible that an ingestion of an excess of alveolar materials in pulmonary alveolar proteinosis may impair the macrophage function and the survival, resulting in the lack of a prominent increase in the macrophage number in bronchoalveolar lavage fluid. PMID- 10515419 TI - M1/MUC5AC mucin released by human airways in vitro. AB - A series of monoclonal antibodies which bind to a mucin known as M1 (anti-M1 MAbs) have also been shown to detect the product of the human gene MUC5AC. The aim of this investigation was to determine the concentration of the M1 mucin in the surface epithelium of human bronchial preparations by means of immunohistochemistry and in the bronchial fluid derived from human airways by means of an immunoradiometric assay. Human bronchial ring preparations from the resection material of 20 patients were challenged with methacholine, leukotriene D4, or anti-immunoglobulin E. Experiments were performed in preparations with an intact epithelium as well as in tissues in which the epithelium had been mechanically removed. The anti-M1 MAbs stained the goblet cells in the epithelium intensely and there was also light and less uniform staining in the submucosa. The M1/MUC5AC mucin in the fluids secreted by the bronchial preparations was not modified during either the experimental protocol or stimulation with the different secretagogues. However, in preparations in which the epithelium had been removed, there was a significant reduction in the amount of M1/MUC5AC mucin detected. These data suggest that the M1/MUC5AC mucin detected in the biological fluids produced by human airways in vitro may be released constantly, and principally from the goblet cells in the epithelial layer. PMID- 10515420 TI - Phospholipase A2 augments contraction and intracellular calcium mobilization through thromboxane A2 in bovine tracheal smooth muscle. AB - Phospholipase A2 (PLA2) induces hyper-sensitivity to muscarinic agonists in airway smooth muscle in vitro. The precise mechanism of this is unknown, but might involve altered calcium homeostasis. In order to elucidate the effects of PLA2, on bovine tracheal smooth muscle contraction, isometric tension and intracellular calcium concentration ([Ca2+]i) were simultaneously measured in fura 2-loaded muscle strips. A high concentration of PLA2 (0.5 microg x mL(-1)) caused the muscle strips to contract, and this contractile response was significantly attenuated by pretreatment with indomethacin (IND; 10 microM), but not by nordihydroguaiaretic acid (NDGA; 10 microM). A low concentration of PLA2 (0.02 microg x mL(-1)) did not directly contract muscle strips. However a low concentration PLA2 significantly enhanced the threshold of the contractile response and that of the [Ca2+]i response to acetylcholine (ACh), but not that of the response to a high K+ concentration. These augmented responses to ACh returned to control levels after pretreatment with IND, a thromboxane (TX) synthetase inhibitor (OKY-046; 10 microM) or a TXA2 receptor antagonist (ONO 3708; 10 microM), but not after NDGA pretreatment. These results suggest that a low concentration of phospholipase A2 enhances smooth muscle responsiveness to acetylcholine by agonist-mediated Ca2+ mobilization facilitated by thromboxane A2. It is concluded that phospholipase A2 plays an important role in bronchial hypersensitivity involving thromboxane A2. It remains to be examined whether similar abnormalities in calcium homeostasis and muscarinic receptor function or coupling are involved in the pathogenesis of asthma. PMID- 10515421 TI - Epidermal growth factor receptor ligands are chemoattractants for normal human mesothelial cells. AB - Signalling through epidermal growth factor (EGF) receptor leads to several cellular responses including cell division and cell migration. Since EGF receptors are expressed on normal mesothelial cells, this study investigated whether EGF receptor ligands act as chemoattractants on these cells. The study used Boyden chambers fitted with filters coated with the adhesive matrix proteins fibronectin, laminin, collagen type IV and the nonmatrix adhesive molecule poly-L lysine, for the migration studies. Normal mesothelial cells migrated to EGF receptor ligands such as EGF, transforming growth factor (TGF)-alpha and heparin binding epidermal growth factor (HB-EGF) at concentrations ranging 0.024-100 ng x mL(-1) (with a peak stimulation at 6.25 ng x mL(-1)), if matrix proteins were present as adhesive substrates. This migration was integrin-dependent, since the same cells failed to migrate in the absence of extracellular matrix molecules or when the Boyden chamber assay was performed in the presence of anti-beta1 integrin monoclonal antibodies. These findings describe for the first time epidermal growth factor receptor ligands acting as chemoattractants on normal mesothelial cells, and that signalling through epidermal growth factor receptors leading to mesothelial cell migration also requires the activation of integrins. PMID- 10515422 TI - Expression and localization of cyclo-oxygenase isoforms in non-small cell lung cancer. AB - The beneficial effects of cyclo-oxygenase (COX) inhibitors in both colon cancer and adenomatous polyps suggest a role for the prostanoid pathway in epithelial malignancy. Although variable prostanoid synthesis in non-small cell lung cancer (NSCLC) has been demonstrated in freshly obtained tissue, COX messenger ribonucleic acid (mRNA) and protein localization in such tumours had not been investigated ex vivo. Thirty-four cases of primary NSCLC were examined for both constitutive (COX-1) and inducible COX (COX-2) by means of in situ hybridization and immunohistochemistry. COX-1 mRNA expression was absent or below the level of detection via in situ hybridization. COX-1 immunohistochemistry demonstrated uniform faint cytoplasmic staining in tumour cells and stromal inflammatory cells. Semiquantitative analysis of COX-2 expression in NSCLC demonstrated the highest levels of both mRNA and protein in adenocarcinoma cells (n=10, p<0.005 compared with large cell and squamous cell carcinoma), intermediate and variable expression in large cell carcinoma (n=11) and low or absent expression in squamous cell tumours (n=13). Levels of COX-2 expression in infiltrating inflammatory cells was the same in all tumour types. In conclusion, tumour cell cyclo-oxygenase-2 rather than cyclo-oxygenase-1 expression may account for the variable prostanoid production seen in non-small cell lung cancer, and primary lung adenocarcinoma expresses the highest levels of cyclooxygenase-2. Assessment of cyclo-oxygenase-2 expression ex vivo should be performed in studies examining the potential therapeutic effects of cyclo-oxygenase inhibitors in non-small cell lung cancer. PMID- 10515423 TI - Influence of age on operative mortality and long-term survival after lung resection for bronchogenic carcinoma. AB - The proportion of elderly patients presenting with bronchogenic carcinoma is increasing. To study the impact of age on clinical presentation, management and outcome of patients, the authors have reviewed their clinical experience over the last 20 yrs. Between 1977 and 1996, 1,079 patients underwent thoracotomy for primary lung carcinoma in the authors' institution. Patients were grouped by age at the time of surgery as <60 yrs, 60-69 yrs and > or =70 yrs. Although the mode of clinical presentation was similar between all age groups, patients <60 yrs were more prone to have advanced stage carcinoma at the time of diagnosis. The rates of exploratory thoracotomy and pneumonectomy were higher in patients <70 yrs, whereas lobectomies and lesser resections largely predominated in patients > or =70 yrs. The mortality rate following lobectomy and lesser resection increased from 1.3% in patients <60 yrs to 5.5% in patients > or =60 yrs (p=0.04) and the mortality rate following pneumonectomy increased from 6.5% in patients <60 yrs to 13.7% in patients > or =70 yrs (p=0.24). The specific long-term survival, which included only the patients who died from primary lung carcinoma, was similar in all age groups. Operative mortality and survival rates are acceptable in patients > or =70 yrs. Therefore, age in itself should not constitute a contraindication to surgical lung resection for primary lung carcinoma as long as a careful preoperative assessment is performed to appropriately select surgical candidates. PMID- 10515424 TI - Lymphoma of pulmonary mucosa-associated lymphoid tissue: CT scan findings and pathological correlations. AB - The aim of this study was to describe the lesions of pulmonary mucosa-associated lymphoid tissue (MALT) lymphoma observed by means of computed tomography (CT) and to determine the value of CT in the management of this disease. Sixty six CT scans were performed in 13 consecutive cases of histologically proven pulmonary MALT lymphoma at the time of diagnosis (n=13) or periodically (n=53). They were reviewed separately with chest radiographs by consensus between two observers. Pulmonary abnormalities were described and compared to histopathological findings on surgical specimens from seven patients. At the time of diagnosis, elementary lesions observed by means of CT consisted of nodular areas of attenuation (12 of 13), linear areas of attenuation (eight of 13) and consolidations (six of 13). All these lesions were centred on airways that appeared dilated in seven cases and were more often bilateral and multiple on CT than on the chest radiographs. CT abnormalities correlated with gross pathological appearance and were related to a lymphomatous infiltration with a peribronchovascular distribution. Pathological examination also confirmed the presence of dilated airways within lymphomatous lesions and showed that the bronchial wall was respected. During follow-up, in patients on chemotherapy, response, relapse or progression were easily identified by means of plain radiography. In the initial evaluation, computed tomography contributed to the choice of therapeutic strategy, avoiding unnecessary surgical resection in one-third of patients. By contrast, it is unclear whether computed tomography is useful for post-treatment follow-up of mucosa-associated lymphoid tissue. PMID- 10515425 TI - A critical evaluation of the Mefar dosimeter. AB - Multicentre studies of airway responsiveness (AR) are increasingly important tools in asthma epidemiology. Because comparisons of AR are made between centres it is essential that measurement techniques are accurate and standard. This study investigated the Mefar dosimeter which is currently used in the 35 centre European Community Respiratory Health Survey (ECRHS) with the next phase currently being planned. Significant differences were found in driving pressures and aerosol outputs between the three Mefar dosimeters in the laboratory. A linear relationship was also found between driving pressure and aerosol output (R2=0.96). These differences are important as they may lead to variations between centres of < or =35% in the drug dose delivered in AR measurement, which could potentially diminish the power of individual study centres to accurately detect national differences in AR. Dosimeter driving pressure and nebulizer output should be standardized in future studies of airway responsiveness. With relatively simple quality control measures in place it is believed that the Mefar dosimeter can produce reliable between-centre longitudinal data with an increase in the accuracy of these important studies. PMID- 10515426 TI - Cystic fibrosis: inflammatory response to infection with Burkholderia cepacia and Pseudomonas aeruginosa. AB - Pulmonary colonization by Burkholderia cepacia in cystic fibrosis (CF) may be associated with enhanced deterioration of pulmonary function. This may be due to a more florid host inflammatory response than in colonization by Pseudomonas aeruginosa, leading to greater lung injury. Circulating markers of inflammation were determined during infective exacerbations and periods of clinical stability in an 18 month prospective study in adults with CF colonized by P. aeruginosa (n=41). B. cepacia (n=13) and in adults who intermittently grew B. cepacia (n=6). There were no differences between the levels of the inflammation markers measured in the three groups (P. aeruginosa, B. cepacia, B. cepacia intermittent) at any of the assessment points. When clinically stable, levels of inflammatory markers in all groups were elevated compared to a matched non-CF population, indicating, continuous inflammation and the potential for lung damage between infective exacerbations. This study does not support the hypothesis that pulmonary colonization with Burkholderia cepacia is associated with a heightened inflammatory response compared with Pseudomonas aeruginosa colonization. PMID- 10515427 TI - Endothelin-1 potentiates cholinergic nerve-mediated contraction in human isolated bronchus. AB - That endothelin-1(ET-1) plays a mediator role in asthma is consistent with reports of ET-1-induced potentiation of cholinergic nerve-mediated contraction in airways from various animal species. This study examined the effect of ET-1 on cholinergic contractions in human isolated bronchus. Macroscopically nondiseased human bronchial tissue was obtained from 23 patients with respiratory tumours. An electrical field stimulation (EFS) frequency that produced one third of the contraction at 30 Hz (EFS30) was estimated. The effect of ET-1 on these EFS evoked contractions was assessed. EFS-evoked contractions were frequency dependent and abolished by either atropine or tetrodotoxin. Thus, EFS-induced contractions were mediated by acetylcholine from cholinergic nerves. ET-1 (3 nM) potentiated EFS-evoked contractions by 10+/-2% EFS30 (p<0.05) without any significant effect on contractions induced by exogenous acetylcholine. Neither the ET(A) receptor-selective antagonist BQ-123 (3 microM) nor the ET(B) receptor selective antagonist BQ-788 (10 microM) alone significantly altered ET-1-induced potentiation of EFS-evoked contractions. However, in the combined presence of both BQ-123 and BQ-788, ET-1-induced potentiation of EFS-evoked contractions was abolished. Thus, prejunctional endothelinA and endothelinB receptors appear to mediate endothelin-1-induced potentiation of electrical field stimulation-evoked cholinergic contractions in human bronchus. This suggests another potentially important mechanism through which endothelin-1 could increase bronchial tone in asthma. PMID- 10515428 TI - Bronchodilation by pituitary adenylate cyclase-activating peptide and related peptides. AB - Pituitary adenylate cyclase-activating peptide (PACAP) is present in nerves in the vicinity of bronchial and vascular smooth muscle in the airways. At least one endogenous form of PACAP, PACAP 1-27, has high affinity binding sites in the lung, probably including cholinergic nerve terminals, bronchial smooth muscle, epithelial and mononuclear inflammatory cells. The mechanism of action for PACAP 1-27 and 1-38 in vivo involves endogenous catecholamines, peptidases and nitric oxide, depending on tissue type. Intracellularly, cyclic adenosine monophosphate (cAMP) as well as calcium and sodium mobilization is probably involved. PACAP 1 27 and 1-38 inhibit airway smooth muscle tone in vitro and in vivo. The inhibitory effect of PACAP 1-38 is more sustained than that of PACAP 1-27, in vitro as well as in vivo. PACAP 1-38 also causes more sustained inhibition of bronchoconstriction after inhalation in vivo, than does vasoactive intestinal peptide (VIP). PACAP 1-27 given intravenously virtually abolishes allergen induced bronchoconstriction in vivo. Novel synthetic analogues of PACAP 1-27 cause more sustained inhibition of airway smooth muscle tone in vitro and in vivo than do PACAP 1-27 or 1-38. Both PACAP 1-27 and 1-38 inhibit arterial smooth muscle tone but, administration of PACAP 1-27, 1-38 or a structural analogue of PACAP 1-27 in the airways, induces no cardiovascular side effects at doses inhibiting bronchoconstriction. PACAP 1-38 enhances phagocytosis in macrophages and inhibits the release of the pro-inflammatory cytokine interleukin-2 in lymphocytes, suggesting antiinflammatory effects. It is concluded that pituitary adenylate cyclase-activating peptide 1-27 and 1-38, or structurally related molecules, may be useful as bronchodilators but their effect on human bronchial smooth muscle and on human inflammatory cells is in need of evaluation. PMID- 10515429 TI - Effects of drugs on mucus clearance. AB - Mucociliary clearance (MCC), the process in which airway mucus together with substances trapped within are moved out of the lungs, is an important defence mechanism of the human body. Drugs may alter this process, such that it is necessary to know the effect of the drugs on MCC. Indeed, agents stimulating MCC may be used therapeutically in respiratory medicine, especially in patients suspected of having an impairment of their mucociliary transport system. In contrast, caution should be taken with drugs depressing MCC as an undesired side effect, independently of their therapeutic indication. Since cough clearance (CC) serves as a back-up system when MCC fails, the influence of drugs must be examined not only on MCC but also on CC. Ultimately, the clinical repercussions of alterations in mucus transport induced by drug administration must be studied. Tertiary ammonium compounds (anticholinergics), aspirin, anaesthetic agents and benzodiazepines have been shown to be capable of depressing the mucociliary transport system. Cholinergics, methylxanthines, sodium cromoglycate, hypertonic saline, saline as well as water aerosol have been shown to increase MCC. Adrenergic antagonists, guaifenesin, S-carboxymethylcysteine, sodium 2-mercapto ethane sulphonate and frusemide have been reported not to alter the mucociliary transport significantly. Amiloride, uridine 5'-triphosphate (UTP), quaternary ammonium compounds (anticholinergics), adrenergic agonists, corticosteroids, recombinant human deoxyribonuclease (rhDNase), N-acetylcysteine, bromhexine and ambroxol have been reported either not to change or to augment MCC. Indirect data suggest that surfactant as well as antibiotics may improve the mucociliary transport system. As for the influence of drugs on CC, amiloride and rhDNase have been demonstrated to increase the effectiveness of cough. A trend towards an improved CC was noted after treatment with adrenergic agonists. The anticholinergic agent ipratropium bromide, which is a quaternary ammonium compound, has been suggested to decrease CC significantly. Bromhexine, ambroxol and neutral saline seemed not to alter CC, either positively or negatively. Finally, treatment with either amiloride, recombinant human deoxyribonuclease, bromhexine, ambroxol, N-acetylcysteine, S-carboxymethylcysteine or hypertonic saline has been suggested as a possible cause of clinical improvement in patients, such as the experience of dyspnoea, the case of expectoration or the frequency of infective exacerbations. Other agents did not show a clinical benefit. PMID- 10515430 TI - A man with upper respiratory tract infections, general weakness and fever. PMID- 10515431 TI - T-cell chronic lymphocytic leukaemia with pulmonary involvement and relapsing BOOP. AB - We report on a case of T-cell chronic lymphocytic leukaemia involving the lung, with clinical, radiological and histological evidence of relapsing bronchiolitis obliterans-organizing pneumonia in a 70-yr-old female. Pulmonary disease was the major clinical manifestation of this chronic lymphocytic leukaemia. The first two episodes of the patient's pulmonary disorder resolved without treatment, and the third episode was treated with cytotoxic agents as part of the leukaemia treatment regimen. Two additional episodes of the pulmonary disorder occurred; both responded to prednisone. PMID- 10515432 TI - Left bronchial isomerism, normal atrial arrangement and bronchomalacia mimicking asthma: a new syndrome? AB - Three children who presented with steroid-resistant airflow obstruction are described. They all had bronchomalacia in the setting of a rare visceral arrangement, namely left bronchial isomerism with normal atrial arrangement. Imaging and, in two cases, a normal residual volume in the face of severe airflow obstruction were diagnostic pointers to a nonasthmatic cause of wheeze. Although the association of these abnormalities may be coincidental, together they may constitute a new clinical syndrome. PMID- 10515433 TI - Hip fracture and bone histomorphometry in a young adult with cystic fibrosis. AB - A 25-yr-old male with cystic fibrosis sustained a fragility fracture of the left femoral neck, which required surgical correction. He had several risk factors for the development of low bone density and despite treatment with an oral bisphosphonate, his bone mineral density reduced further. The patient died 2 yrs after sustaining the fracture. Bone specimens obtained at post mortem demonstrated severe cortical and trabecular osteopenia, but the histological features were not typical of osteoporosis or osteomalacia. Osteoporosis is thought to be a common complication of cystic fibrosis. The novel histomorphometric appearances reported here suggest that the bone disease of cystic fibrosis may be more complex and possibly unique. Labelled bone biopsies are required to clarify the bone defect leading to low bone density in cystic fibrosis patients so that appropriate therapeutic strategies can be developed. PMID- 10515434 TI - Malignant mesothelioma and erionite exposure. PMID- 10515435 TI - "Cross-talk" among a multiplicity of pro-inflammatory agents: main cause of tissue damage in pulmonary inflammation? PMID- 10515436 TI - Sarcoidosis and cancer revisited. PMID- 10515437 TI - Hepatocyte growth factor (HGF) as a potential index of severity of hypertension. AB - Hepatocyte Growth Factor (HGF) is a mesenchyme-derived pleiotropic factor that regulates cell growth, cell motility, and morphogenesis of various cells, and is thus considered a humoral mediator of epithelial-mesenchymal interactions. We previously identified HGF as a novel member of the family of endothelium-specific growth factors. Moreover, the presence of a local HGF system (HGF and its specific receptor, c-met) has been demonstrated in vascular cells both in vitro and in vivo. HGF might contribute to the protection and/or repair of vascular endothelial cells injured by high blood pressure. If so, serum HGF level might be elevated in response to endothelial cell damage. To test this hypothesis, we measured serum levels of HGF in hypertensive and normotensive patients. Serum HGF concentration in hypertensive patients without any complications was significantly higher than that in normal subjects. Interestingly, serum HGF concentration in hypertensive patients with complications was significantly higher than that in either hypertensive patients without complications or normotensive subjects. Of importance, hypertensive patients treated with antihypertensive drugs showed the same level of serum HGF concentration as normotensive subjects. In contrast, serum HGF concentration in diabetic patients without hypertension was significantly lower than that in normal subjects, whereas serum HGF concentration in diabetic patients with hypertension was significantly higher than that in normal subjects. Moreover, serum HGF concentration in diabetic patients with hypertensive complications was even higher than that in diabetics without complications. This review discusses the possibility that HGF may be considered as a new index of the severity of hypertension. PMID- 10515438 TI - Sexual differences in relationships between birth weight or current body weight and blood pressure or cholesterol in young Japanese students. AB - This study was designed to examine the relationships between birth weight or current body weight and blood pressure (BP) or cholesterol in 178 Japanese high school students (98 male, 80 female, age 15-16 yr). All subjects were born after a full-term pregnancy (gestational age > or = 38 wk) with a birth weight > or = 2,500 g; these data were obtained from routine obstetrical records. At a health check-up, nurses used an automatic device to perform two consecutive BP measurements with each subject in a sitting position after resting for at least 5 min. Serum total and high-density lipoprotein (HDL) cholesterol levels were measured. Birth weight was not related to BP, but was inversely related to serum total cholesterol in both males (r= -0.241, p < 0.05) and females (r= -0.351, p < 0.01). Current body weight was significantly related to systolic BP (r=0.369, p<0.01), diastolic BP (r=0.216, p<0.05), and HDL cholesterol level (r= -0.224, p < 0.05) in males, but not in females. Although no relationship was demonstrated between birth weight and BP level in young Japanese students without intrauterine growth retardation, an inverse relationship between birth weight and serum total cholesterol level was found. There was a gender difference in the relationship between current body weight and either BP or HDL cholesterol in these subjects. PMID- 10515439 TI - Inhibition by transforming growth factor-beta1 of the cellular action of arginine vasopressin in cultured rat glomerular mesangial cells. AB - The present study was undertaken to determine whether transforming growth factor (TGF)-beta1 modulates the cellular actions of arginine vasopressin (AVP) in cultured rat glomerular mesangial cells. AVP increased cytosolic free calcium ([Ca2+]i), and TGF-beta1 dose-dependently reduced the AVP-mobilized [Ca2+]i. Such an inhibition by exogenous TGF-beta1 was abolished by liposomal transfection of antisense oligodeoxynucleotide for the TGF-beta type II receptor. AVP activated mitogen-activated protein (MAP) kinase, which was significantly reduced by 1 ng/ml TGF-beta1. AVP increased [3H]thymidine incorporation into mesangial cells in a dose-dependent manner, and 1 ng/ml TGF-beta1 significantly reduced the AVP stimulated [3H]thymidine incorporation. However, 10 microM antisense oligodeoxynucleotide for the TGF-beta type II receptor seemed to attenuate the inhibition by TGF-beta1. 1 X 10(-7) M AVP significantly increased inositol 1,4,5 trisphosphate (IP3) production by 1.8-fold, but this production was totally blunted by 1 ng/ml TGF-beta1. TGF-beta1 did not affect [3H]AVP receptor binding. 1 X 10(-6) M AVP concentration stimulated TGF-beta1 production in mesangial cells by 4-fold. These results indicate that TGF-beta1 inhibits the cellular signaling of AVP at steps beyond the AVP receptors and prior to the phospholipase C activation, and that TGF-beta1 may participate in a negative feedback regulation on the cellular action of AVP in glomerular mesangial cells. PMID- 10515440 TI - Effects of salt-loading on membrane potentials in mesenteric arteries of spontaneously hypertensive rats. AB - Although salt intake and blood pressure are correlated, with hypertensives tending to exhibit higher blood pressure sensitivity to salt than normotensives, the precise mechanisms underlying this relationship remain unclear. This study aimed to determine whether salt-loading affects arterial membrane properties of spontaneously hypertensive rats (SHR). SHR and age-matched Wistar Kyoto rats (WKY) received either an 8% high salt diet or standard rat chow from 6 to 16 wk of age. Systolic blood pressure was significantly higher in salt-loaded SHR than in control SHR (267+/-7 vs. 235 +/- 5 mmHg, p < 0.05). The membrane potential of isolated conduit and resistance arteries of the superior mesenteric vascular bed, measured with microelectrodes, was less negative in salt-loaded SHR than in control SHR or salt-loaded WKY (conduit arteries, -39.9 +/- 0.3 vs. -44.5 +/- 0.4 or -47.4 +/- 0.4 mV, respectively, p < 0.05 for each; resistance arteries, -55.5 +/- 0.5 vs. -62.5 +/- 0.5 or -67.0 +/- 0.5 mV, respectively, p < 0.05 for each). Furthermore, conduit arteries of salt-loaded SHR exhibited spontaneous electrical activity (4-13 mV, 1-3/min), which was sensitive to ONO-3708, a thromboxane A2/prostaglandin H2 receptor antagonist. These findings suggest that salt-loading in SHR leads to a membrane depolarization in both conduit and resistance arteries, as well as to spontaneous electrical activity, presumably mediated by eicosanoids, in conduit arteries. These alterations in membrane properties might contribute to the exacerbation of hypertension and/or the target organ damage after salt loading in SHR. PMID- 10515441 TI - The unique 5-flanking region of the human basic calponin gene. AB - Calponin has been implicated in the regulation of smooth muscle contraction. Basic calponin, one of the calponin isoforms, is expressed exclusively in smooth muscle cell (SMC)-rich tissues, and is considered to be a phenotypic marker of differentiated SMC. To define the molecular mechanism of SMC-specific gene transcription in humans, we isolated and characterized the 5'-flanking region of this gene. Sequence analysis revealed that several putative cis-acting elements were clustered within a 500-bp sequence upstream of the transcription start site. However, the 1.9-kb promoter region obtained herein lacked a completely matched consensus sequence of the CArG box that is commonly identified in the promoter region of other SMC-specific genes. A luciferase assay demonstrated that the 1.9 kb promoter region was sufficient to drive a basal transcriptional activity not only in human vascular smooth muscle cells (VSMC) but also in HeLa cells. In particular, the sequence between positions -1,906 and -867 had a significantly higher transcriptional activity in VSMC than in HeLa cells. In contrast, the promoter activity was drastically decreased between positions -327 and -257 in both types of cells. These results indicate that the sequence spanning from position -327 to -257 contains an essential domain involved in the basal transcriptional activity of the human basic calponin gene, and that the distal region of the 1.9-kb 5'-flanking sequence presented herein may play a pivotal role in the phenotypic modulation of VSMC. PMID- 10515442 TI - Difference in the incidence of cough induced by angiotensin converting enzyme inhibitors: a comparative study using imidapril hydrochloride and enalapril maleate. AB - To compare the incidence of cough between two angiotensin converting enzyme (ACE) inhibitors, imidapril and enalapril, comparative crossover study was performed in 489 patients (228 men and 261 females) with essential or renal parenchymal hypertension. Patients were randomly assigned to one of two treatment groups, a group receiving imidapril for 12 wk (Period I) followed by enalapril for 12 wk (Period II), and a group in which the order of drugs was reversed. The occurrence of cough during treatment was monitored by questionnaire in all cases. There were no differences in background characteristics between the two groups. The incidence of cough during Period I was 15.2% (32/210) in the group initially treated with imidapril (Group IE) and 38.6% (85/220) in the group initially treated with enalapril (Group EI), the difference being statistically significant (p < 0.001). During Period I, decrease in blood pressure was observed in 63.9% (115/180) of Group IE and 64.6% (115/178) of Group EI patients. In approximately half of the patients in Group EI who developed cough during Period I and in whom the treatment was subsequently switched to imidapril, cough subsequently disappeared. It was concluded that the incidence of cough was significantly less under imidapril than under enalapril treatment, while there was no difference in the antihypertensive effects of the two ACE inhibitors. PMID- 10515443 TI - Low wall shear stress contributes to atherosclerosis of the carotid artery in hypertensive patients. AB - Numerous in vitro studies have indicated that low shear stress may contribute to intimal thickening and development of atherosclerosis. In the present study, we investigated wall shear stress in hypertensive patients and its relevance to atherosclerosis in the carotid arteries by means of a non-invasive technique. Fifty-five hypertensive patients and 23 normotensive controls were investigated. Intima-media thickness, number of plaques, internal dimension and blood flow velocity of the carotid artery were evaluated. Wall shear stress was calculated using the Poiseuillean parabolic model of velocity distribution as follows: shear stress = 4 X blood viscosity X central line flow velocity/internal dimension. Hypertensive patients showed increased intima-media thickness and dilated common carotid arterial dimension relative to normotensive controls. There was no difference in blood viscosity between the two groups. Both the mean shear stress and systolic peak shear stress were significantly lower in hypertensive patients than normotensive controls. Further, wall shear stress at both mean and peak velocity was significantly and negatively related to intima-media thickness and number of plaques in hypertensive patients, as well as in the total study population. These findings indicate that structural and functional alterations in the common carotid artery of hypertensive patients further precipitates atherosclerosis through low shear stress. PMID- 10515444 TI - Cardiovascular risk factors emerging in Chinese populations undergoing urbanization. AB - In this assessment of cardiovascular risk factors, we examined the association between dietary habits and blood pressure (BP) according to the World Health Organization (WHO) CARDIAC Study protocols in three Chinese populations aged 47 57 in Guangzhou prefecture (GZ group; 141 males, 158 females), Guiyang prefecture (GY group; 101 males, 103 females) and Taiwan (TW group; 102 males, 98 females). The same survey was repeated 10 yr later in the GY group to follow-up the past trends (MONALISA study). The observed systolic BP (SBP), diastolic BP (DBP), and body mass index (BMI), as well as the rates of hypertension, obesity and antihypertensive medication use were significantly higher in both genders in the TW group compared to the groups GZ and GY. There was no significant difference in SBP or DBP in either gender between groups GZ and GY. Blood analyses revealed that the levels of serum total cholesterol (T-CHO), and HbA1c, and the rates of hypercholesterolemia and high HbA1c were significantly higher in both genders in the TW than in the GZ and GY groups. No significant difference among the populations was observed in 24-h urinary sodium or magnesium excretion in either gender. In the combined total populations of men and women, however, significant positive correlations were observed between BMI and each of SBP, DBP, T-CHO, and glycohemoglobin in both genders. A food frequency analysis revealed significantly greater meat consumption and significantly less tea consumption and vegetable intake in the TW than in the GY and GZ groups. Both SBP and DBP have increased significantly over the past 10 yr in the GY group in both genders, and T-CHO as well as the rate of hypercholesterolemia increased over the same period in both genders. In conclusion, cardiovascular risk factors leading to hypertension, such as obesity, hypercholesterolemia and diabetes mellitus, are emerging in urbanized Taiwan and developing Guiyang due to the loss of traditional dietary habits. PMID- 10515445 TI - The role of the renin-angiotensin and cardiac sympathetic nervous systems in the development of hypertension and left ventricular hypertrophy in spontaneously hypertensive rats. AB - To elucidate the relationship between the development of left ventricular hypertrophy (LVH) in hypertension and the development of both the cardiac sympathetic nervous and renin-angiotensin systems, as measured by norepinephrine and angiotensin II levels, respectively. In this longitudinal study, we compared blood pressure (BP), left ventricular weight, and norepinephrine (NE) and angiotensin II (Ang II) concentrations, in Spontaneously Hypertensive Rats (SHR) and age-matched Wistar-Kyoto (WKY) rats at 5, 10, 15, 20, and 28 wk of age. Blood pressure, plasma and ventricular Ang II and tissue NE were measured by the tail cuff method, radioimmunoassay, and high-performance liquid chromatography (HPLC), respectively. At 5 wk, systolic blood pressure was the same in both strains. But the left ventricular plus septum weight to body weight (LVSW/BW) ratio was higher in SHR than in WKY rats (p < 0.01), which finding may have been related to the increased cardiac tissue NE concentration, and this increase tended to parallel the rise in blood pressure. Both left ventricle and forelimb muscle NE concentrations were significantly higher in SHR than in WKY rats at 5, 10, and 15 wk of age (p < 0.01, respectively), and were similar at 20 and 28 wk of age. The heart and plasma Ang II levels decreased with age, which results were in keeping with the known developmental tendencies of the biological aging progress. There was no significant difference in plasma Ang II levels between the two strains from 5 to 20 wk, whereas these levels were remarkably higher in WKY than in SHR rats at 28 wk (p< 0.01). Otherwise, the left ventricular tissue Ang II concentrations were significantly higher in SHR than in WKY rats at the late stage (from 15 to 28 wk), which may have contributed to the late-stage cardiac hypertrophy. These results suggested that the sympathetic nervous system (SNS) and the renin-angiotensin-system (RAS) in SHR may contribute to the pathogenesis of hypertension and LVH at the early and late stages, respectively. PMID- 10515446 TI - Renal protective effects of blocking the intrarenal renin-angiotensin system. AB - It has been well demonstrated that angiotensin-converting enzyme inhibitors (ACEIs) can retard the progression of renal failure and kidney sclerosis in patients and animal models with glomerular diseases. The aim of this study was to observe the influences of ACEI on intrarenal Ang II and TGFbeta1 local formation and their relation to renal protective effects. Experimental glomerulosclerosis with nephrotic syndrome was induced in unilateral nephrectomized rats with repeated injections of adriamycin. Rats were randomly divided into three groups: 1) a sham-operated control group (n=8); 2) an NS group treated with ACEI (benazepril 4 mg/kg/d) (n=10), and 3) an NS group not treated (n=10). After 8 wk, serum, urine and renal tissue were collected for study. ACE activity and Ang II concentration in renal tissue were measured by colorimetry and radioimmunoassay, respectively. Immunohistochemistry staining was employed for transforming growth factor-beta1 (TGFbeta1) and extracellular matrix (ECM) examination. TGFbeta1 mRNA was assessed by in situ hybridization. Compared with those of non-treated nephropathy rats, ACE activity (13.39+/-5.02 vs. 49.13+/-12.92 U/ml, p< 0.01) and Ang II (402.61+/-80.22 vs. 751.63+/-137.45 pg/mg/pr p < 0.01) in renal tissue were significantly inhibited in the rats treated with ACEI. At the same time, proteinuria was significantly reduced (155.06+/-103.56 vs. 421.11+/-148.45 mg/24 h, p < 0.01) and renal function improved (Scr 76.3+/-33.1 vs. 107.1+/-71.0, p < 0.05), concomitant with a reduction in the glomerular sclerosis index (30.6+/ 19.5 vs. 120.3+/-61.9, p < 0.01) and a reduction in ECM accumulation such as Col IV, III, LN and FN (29.2+/-9.8 vs. 76.8+/-12.4; 29.5+/-12.4 vs. 85.9+/-11.5; 26.0+/-5.1 vs. 69.6+/-1.73; 32.4+/-12.4 vs. 70.5+/-13.5; p< 0.01 in all cases). In the ACEI treated group, these histologic benefits coincided with a reduced expression of TGFbeta1 in both tubular cells and sclerosed glomeruli in protein as well as mRNA level. These findings provide further evidence that ACEI (benazepril) can prevent the progression of renal damage in both the function and morphologic changes which associated with a down-regulation of intrarenal Ang II level through the relative inhibition of renal ACE activity. The blocking of the intrarenal renin angiotensin system (RAS) might contribute to the inhibition of TGFbeta1 local formation and the TGFbeta1-mediated ECM accumulation that are related to the renal protective effects of ACEI. PMID- 10515447 TI - Improved artificial death switches based on caspases and FADD. AB - A number of "suicide genes" have been developed as safety switches for gene therapy vectors or as potential inducible cytotoxic agents for hyperproliferative disorders, such as cancer or restenosis. However, most of these approaches have relied on foreign proteins, such as HSV thymidine kinase, that primarily target rapidly dividing cells. In contrast, novel artificial death switches based on chemical inducers of dimerization (CIDs) and endogenous proapoptotic molecules function efficiently in both dividing and nondividing cells. In this approach, lipid-permeable, nontoxic CIDs are used to conditionally cross-link target proteins that are fused to CID-binding domains (CBDs), thus activating signaling cascades leading to apoptosis. In previous reports, CID-regulated Fas and caspases 1, 3, 8, and 9 were described. Since the maximum efficacy of these artificial death switches requires low basal and high specific activity, we have optimized these death switches for three parameters: (1) extent of oligomerization, (2) spacing between CBDs and target proteins, and (3) intracellular localization. We describe improved conditional Fas and caspase 1, 3, 8, and 9 alleles that function at subnanomolar levels of the CID AP1903 to trigger apoptosis. Further, we demonstrate for the first time that oligomerization of the death effector domain of the Fas-associated protein, FADD, is sufficient to trigger apoptosis, suggesting that the primary function of FADD, like that of Apaf-1, is oligomerization of associated caspases. Finally, we demonstrate that nuclear-targeted caspases 1, 3, and 8 can trigger apoptosis efficiently, implying that the cleavage of nuclear targets is sufficient for apoptosis. PMID- 10515448 TI - Perflubron enhances adenovirus-mediated gene expression in lungs of transgenic mice with chronic alveolar filling. AB - Perfluorochemical (PFC) liquids have both low surface tension and a high capacity to dissolve O2 and CO2, and have been shown to improve gas exchange and lung compliance in animal models of lung injury. We have previously demonstrated that perflubron and other PFC liquids enhance transgene expression in lungs of spontaneously breathing normal rodents after intratracheal instillation of either adenoviral or liposomal vectors followed by a single instillation of PFC liquid. We reasoned that PFC liquids may also be useful for enhancing transgene expression in abnormal lungs. GM-CSF knockout mice develop chronic accumulation of surfactant lipids and proteinaceous material in alveolar spaces and serve as a useful model of chronic alveolar filling. Intratracheal instillation of the adenoviral vector Adlac-Z resulted in patchy in situ distribution of beta-Gal activity, predominantly in larger proximal airways. In contrast, in mice instilled with Adlac-Z followed by instillation of a single dose of perflubron (10 ml/kg body weight), increased expression was observed in distal airway and alveolar epithelial cells. In particular, expression was observed in epithelial cells of debris-filled alveoli. Spectrophotometric measure of quantitative beta Gal activity in lung homogenates demonstrated increased activity in lungs of mice receiving Adlac-Z plus perflubron compared with lungs of animals receiving Adlac Z alone. These studies demonstrate that use of perflubron enhances transgene expression in lungs of animals with a chronic alveolar filling process. This approach may be applicable for gene delivery in diseases marked by chronic airway or alveolar filling such as cystic fibrosis. PMID- 10515449 TI - Enhanced expression of transgenes from adeno-associated virus vectors with the woodchuck hepatitis virus posttranscriptional regulatory element: implications for gene therapy. AB - The woodchuck hepatitis virus posttranscriptional regulatory element (WPRE) evolved to stimulate the expression of intronless viral messages. To determine whether this ability to enhance expression could be useful in nonviral and heterologous viral gene delivery systems, we analyzed the ability of the WPRE to elevate the expression of a cDNA encoding the green fluorescent protein (GFP) in these contexts. We find that the WPRE can stimulate the expression of GFP when the gene is delivered by transfection or transduction with recombinant adeno associated virus (AAV). Enhancement occurred both during transient expression and when the gene is stably incorporated into the genome of target cells. This enhancement required that the WPRE be located in cis within the GFP message, and was observed in both transformed cell lines and primary human fibroblasts. These results demonstrate that the WPRE will be an effective tool for increasing the long-term expression of transgenes in gene therapy. PMID- 10515450 TI - Intraoperative multiplane transesophageal echocardiography for guiding direct myocardial gene transfer of vascular endothelial growth factor in patients with refractory angina pectoris. AB - Gene transfer for therapeutic angiogenesis represents a novel treatment for patients with chronic angina refractory to standard medical therapy and not amenable to conventional revascularization. We sought to assess the role of intraoperative multiplane transesophageal echocardiography (MPTEE) in guiding injection of naked DNA encoding vascular endothelial growth factor (VEGF) into the left ventricular (LV) myocardium of patients with refractory angina. After exposing the LV myocardium via a limited lateral thoracotomy, each of 17 patients in this series received 4 separate injections of VEGF DNA into different myocardial sites. Initial injections in the first patient produced intracavitary microbubbles, indicating injection of DNA into the LV chamber. Subsequently, each injection was preceded by a test injection of agitated saline. The absence of microbubbles while visualizing the LV cavity during the test injection verified that the ensuing injection of DNA would not be inadvertently squandered in the LV chamber itself. Intracavitary LV microbubbles were observed by MPTEE in 13 of 64 (20.3%) saline test injections and in 8 of 16 (50.0%) patients in which saline test injection was used, leading to adjustments in needle position. MPTEE imaging detected a previously unknown large, apical left ventricular thrombus in one patient, thereby preventing inadvertent injection of VEGF DNA through the myocardium into the thrombus. Imaging during and after injection verified no deleterious impact on LV function. We conclude that MPTEE is a useful tool for ensuring that myocardial gene therapy performed by direct needle injection results in gene transfer to the LV myocardium. PMID- 10515451 TI - Treatment of bleomycin-induced pulmonary fibrosis by transfer of urokinase-type plasminogen activator genes. AB - During acute and chronic inflammatory lung diseases, the normal fibrinolytic activity in the alveolar space is inhibited by increased levels of plasminogen activator inhibitor 1 (PAI-1). Transgenic mice having increased fibrinolytic activity due to genetic deficiency of PAI-1 develop less fibrosis after bleomycin induced lung inflammation. These observations led us to hypothesize that pulmonary fibrosis could be limited through enhancement of alveolar fibrinolytic activity by adenovirus-mediated transfer of the urokinase-type plasminogen activator (uPA) gene to the lung. To investigate this hypothesis, 0.075 U of bleomycin was introduced intratracheally into mice. Twenty-one days later, the mice were treated intratracheally with phosphate-buffered saline (PBS), a control adenovirus, or adenoviruses containing murine or human uPA cDNAs. On day 28, the mice were sacrificed, and lung fibrosis was quantitated by measuring hydroxyproline content. As expected, bleomycin caused a doubling in lung hydroxyproline to 345.6+/-28.2 microg/lung (SEM) compared with mice receiving PBS (170.2+/-4.0 microg/lung). Treatment of the bleomycin-injured mice with the control adenovirus on day 21 had no impact on lung fibrosis (338.4+/-17.2 microg/lung). Importantly, the human uPA adenovirus significantly reduced (p<0.05) lung hydroxyproline (281.2+/-22.8 microg/lung), thus attenuating by 38% the bleomycin-induced increase in lung collagen. The improvement in bleomycin induced lung fibrosis resulting from treatment with the human uPA adenovirus further supports the importance of the fibrinolytic system during inflammatory lung injury and repair. PMID- 10515453 TI - Engraftment of hematopoietic progenitor cells transduced with the Fanconi anemia group C gene (FANCC). AB - Fanconi anemia (FA) is an autosomal recessive disorder that leads to aplastic anemia. Mutations in the FANCC gene account for 10-15% of cases. FA cells are abnormally sensitive to DNA-damaging agents such as mitomycin C (MMC). Transfection of normal FANCC into mutant cells corrects this hypersensitivity and improves their viability in vitro. Four FA patients, representing the three major FANCC mutation subgroups, were entered into a clinical trial of gene transduction aimed at correction of the hematopoietic defect. Three patients received three or four cycles of gene transfer, each consisting of one or two infusions of autologous hematopoietic progenitor cells that had been transduced ex vivo with a retroviral vector carrying the normal FANCC gene. Prior to infusion, the FANCC transgene was demonstrated in transduced CD34-enriched progenitor cells. After infusion, FANCC was also present transiently in peripheral blood (PB) and bone marrow (BM) cells. Function of the normal FANCC transgene was suggested by a marked increase in hematopoietic colonies measured by in vitro cultures, including colonies grown in the presence of MMC, after successive gene therapy cycles in all patients. Transient improvement in BM cellularity coincided with this expansion of hematopoietic progenitors. A fourth patient, who received a single infusion of transduced CD34-enriched BM cells, was given radiation therapy for a concurrent gynecologic malignancy. The FANCC transgene was detected in her PB and BM cells only after recovery from radiation-induced aplasia, suggesting that FANCC gene transduction confers a selective engraftment advantage. These experiments highlight both the potential and difficulties in applying gene therapy to FA. PMID- 10515452 TI - A phase 1-2 clinical trial of gene therapy for recurrent glioblastoma multiforme by tumor transduction with the herpes simplex thymidine kinase gene followed by ganciclovir. GLI328 European-Canadian Study Group. AB - This study has investigated the effects of herpes simplex thymidine kinase gene (HSV-tk) transfer followed by ganciclovir treatment as adjuvant gene therapy to surgical resection in patients with recurrent glioblastoma multiforme (GBM). The study was open and single-arm, and aimed at assessing the feasibility and safety of the technique and indications of antitumor activity. In 48 patients a suspension of retroviral vector-producing cells (VPCs) was administered by intracerebral injection immediately after tumor resection. Intravenous ganciclovir was infused daily 14 to 27 days after surgery. Patients were monitored for adverse events and for life by regular biosafety assaying. Tumor changes were monitored by magnetic resonance imaging (MRI). Reflux during injection was a frequent occurrence but serious adverse events during the treatment period (days 1-27) were few and of a nature not unexpected in this population. One patient experienced transient neurological disorders associated with postganciclovir MRI enhancement. There was no evidence of replication competent retrovirus in peripheral blood leukocytes or in tissue samples of reresection or autopsy. Vector DNA was shown in the leukocytes of some patients but not in autopsy gonadal samples. The median survival time was 8.6 months, and the 12-month survival rate was 13 of 48 (27%). On MRI studies, tumor recurrence was absent in seven patients for at least 6 months and for at least 12 months in two patients, one of whom remains recurrence free at more than 24 months. Treatment-characteristic images of injection tracks and intracavity hemoglobin were apparent. In conclusion, the gene therapy is feasible and appears to be satisfactorily safe as an adjuvant to the surgical resection of recurrent GBM, but any benefit appears to be marginal. Investigation of the precise effectiveness of this gene therapy requires prospective, controlled studies. PMID- 10515454 TI - Distribution of recombinant adenovirus in the cerebrospinal fluid of nonhuman primates. AB - Gene therapy by administration of vectors into the cerebrospinal fluid (CSF) may be used in treatment of leptomeningeal metastases (cancer gene therapy) as well as in treatment of neurodegenerative disorders, traumatic injury, and chronic pain. Recombinant adenoviruses are attractive vectors for intra-CSF administration because they can efficiently transfer genes into the nonreplicating cells of the central nervous system (CNS). In addition, they can be produced in high titers and, because no producers cells are introduced, the risk of CSF obstruction by clustering cells is circumvented. However, successful application requires favorable distribution dynamics, high transduction efficiency, and long-lasting transgene expression. In this study we examined the distribution of a recombinant adenovirus containing the lacZ gene after administration into the CSF of nonhuman primates. After intraventricular and suboccipital administration, homogeneous distribution of the vector along the meninges covering the brain and spinal cord was obtained, as demonstrated by extensive and intense blue staining of cells, predominantly in the arachnoid and pia mater. In one animal we also found beta-galactosidase activity in the cervical paraspinal fat and in one of the deep cervical lymph nodes, indicating drainage of the vector or vector products with CSF into cervical lymph. This route of vector clearance from the CNS may result in antigenic presentation and an effective immune response and may explain the sixfold higher serum antibody titers after intrathecal injection of adenovirus as compared with intranasal application in Fischer rats. We conclude that distribution dynamics of recombinant adenovirus after intra-CSF administration are excellent. However, because of the immune response elicited by the virus, even after administration to the CNS, development of immunomodulating strategies remains a challenge. PMID- 10515455 TI - A pressure-mediated nonviral method for efficient arterial gene and oligonucleotide transfer. AB - In this study, we report a method of controlled pressure-mediated delivery of "naked" DNA that achieves efficient and safe arterial gene and oligonucleotide transfer. We demonstrated a pressure-dependent uptake of fluorescein-labeled (FITC) oligonucleotide (ODN) in rabbit carotid arteries with preexisting neointimal hyperplasia, using nondistending intravascular delivery pressures ranging from 0 to 760 mm Hg. At an infusion pressure of 50 mm Hg, 10.5+/-5% of neointimal cell nuclei were positive for nuclear uptake of FITC-ODN 4 days after transfection. With an infusion pressure of 760 mm Hg, the transfection efficiency increased to 84.2+/-5.3% of neointimal cells, and to 64.5+/-11.6 and 92.4+/-5.5% of medial and adventitial cells, respectively. Similar patterns of FITC-ODN uptake were seen in atherosclerotic injured arteries. We also investigated the pressure-mediated delivery of plasmid DNA. Transfection of a luciferase expression plasmid, using an infusion pressure of 760 mm Hg, yielded luciferase expression of 816.6+/-108.6 fg/mg protein in normal rabbit carotid arteries, as compared with 38.9+/-23.7 fg/mg protein at 100 mm Hg. Luciferase expression was significantly higher in pressure-transfected injured atherosclerotic arteries (5467.3+/-1047.6 fg/mg protein at 760 mm Hg). Transfection of beta-galactosidase indicated that significant transgene expression occurred in the neointima and media. These data indicate that this pressure-mediated transfection method yields efficient oligonucleotide delivery and enhances transduction with plasmid DNA in normal as well as injured nonatherosclerotic or atherosclerotic arteries. PMID- 10515456 TI - Adenovirus-mediated suicide gene transduction: feasibility in lens epithelium and in prevention of posterior capsule opacification in rabbits. AB - The most common complication of cataract surgery is the development of posterior capsule opacification (PCO). Hyperplasia of the lens epithelium is one of the main cellular events following phacoemulsification, and has been found to be an important feature contributing to opacification of the posterior capsule. Adenoviral vector-mediated transfer is a suitable method for transducing the herpes simplex virus thymidine kinase gene (HSV-tk) into proliferating cells, allowing for the selective killing of these cells by ganciclovir (GCV) treatment. To determine the potential of gene transduction for lens epithelial cells, we studied the transduction of rabbit lens epithelial cells with adenoviral vectors containing either the Escherichia coli beta-galactosidase (lacZ) gene or the HSV tk gene in vitro and in vivo in an experimental model of PCO. The efficiency of lacZ gene transfer in rabbit lens epithelial cells was at least 95% both in vitro and in vivo. In vivo transduction with HSV-tk adenoviral vector followed by GCV treatment significantly inhibited the development of PCO (p<0.001). These results suggest that adenoviral vector-mediated transfer of HSV-tk into the proliferating lens epithelial cells is feasible and may provide a novel therapeutic strategy for PCO. PMID- 10515457 TI - In vivo transfer of bacterial marker genes results in differing levels of gene expression and tumor progression in immunocompetent and immunodeficient mice. AB - To optimize gene delivery for the treatment of malignant mesothelioma, expression of the beta-galactosidase marker gene was examined in a murine model of intraperitoneal malignant mesothelioma. The beta-galactosidase gene was delivered to the peritoneal cavity of tumor-bearing mice by various plasmid-liposome complexes or by replication-incompetent retrovirus, used alone or complexed to liposomes. In tumor samples from immunodeficient nude mice, moderate levels of gene expression were achieved by liposome-complexed plasmids. Retroviral gene delivery was more effective, and was increased nearly 10-fold by complexing the retrovirus to liposomes. In contrast, in tumor samples from immunocompetent CBA mice treated with the same vectors, no marker gene expression was detected. In immunodeficient mice, tumor growth was not affected by beta-galactosidase gene transfer. However, immunocompetent mice showed a significant decrease in tumor size and increase in survival time after beta-galactosidase delivery. Induction of cytotoxic T cells capable of lysing beta-Gal-transfected tumor cells suggests that tumor cells transduced with the bacterial beta-galactosidase gene may be eliminated in immunocompetent hosts. Our findings also indicate that plasmid liposome complexes, which achieve a low level of gene expression, and retrovirus liposome complexes, which result in nearly 100 times higher levels of gene expression in tumor cells in vivo, are similarly effective in inducing an antitumor immune response. PMID- 10515458 TI - Conferral of an antiviral state to CD4+ cells by a zipper motif envelope mutant of the human immunodeficiency virus type 1 transmembrane protein gp41. AB - We showed in a transient coexpression study that a single proline substitution for any of the five conserved leucine or isoleucine residues located in the envelope (Env) transmembrane protein gp41 zipper motif of the human immunodeficiency virus type 1 dominantly interferes with wild-type Env-mediated viral infectivity. In the present study, we intended to explore the feasibility of developing a genetic anti-HIV strategy targeting the zipper motif. Stable HeLa CD4-LTR-beta-gal clones that harbored silent copies of Tat-regulated expression cassettes encoding the zipper motif Env mutants were first generated. Expression of any of the five Env mutants in transfectants interfered with exogenously expressed homologous HXB2 Env-mediated cytopathic effects. Mutant transfectants 566, 573, and 580 were further examined. Viral transmission mediated by the laboratory-adapted T cell-tropic HXB2 and NL4-3 viruses was greatly reduced in these transfectants compared with that observed in the env-defective control deltaKS and wt env transfectants. Moreover, viral replication mediated by the NL4 3 virus and a macrophage-tropic ADA-GG virus was delayed or reduced in human T cells harboring the mutant 566 or 580 env construct as opposed to those observed in cells harboring the control deltaKS or mutant 573 env construct. The wt and mutant Env proteins formed a hetero-oligomer when they were coexpressed. These results demonstrate that zipper motif Env mutants 566 and 580 confer an anti-HIV state to the host CD4+ cells, which indicates that dominant inhibitory mutants targeting the gp41 zipper motif might function as genetic anti-HIV agents to combat HIV-1 infection. PMID- 10515459 TI - MHC class II alpha/beta heterodimeric cell surface molecules expressed from a single proviral genome. AB - Transplantation tolerance to renal allografts can be induced in large animal preclinical models if the donor and recipient have identical major histocompatibility complex (MHC) class II loci. Such class II matching is, however, not clinically achievable owing to the extreme diversity of class II sequences. With the ultimate goal of creating a somatic class II match in the bone marrow of an allograft recipient, the aim of the study is to develop a double-copy retrovirus construct to express both chains of the MHC class II DQ glycoprotein on a single transduced cell. Analysis of the expression patterns of the retroviral DQ transgenes in both virus producer and transduced fibroblasts revealed correct transcription and stable surface expression of the DQ heterodimers. In addition, we demonstrate that both the DQA and DQB sequences are functional within the same proviral copy, a prerequisite for efficient induction of transplantation tolerance following transduction of bone marrow precursor cells. The DQ double-copy retrovirus vector showed efficient expression of the transferred class II cDNA in murine colony-forming units for the granulocyte monocyte lineage (CFU-GM), indicating that it is suitable for gene therapy of multimeric proteins in hematopoietic cells. PMID- 10515461 TI - Recombinant adenovirus gene transfer in adults with partial ornithine transcarbamylase deficiency (OTCD). PMID- 10515460 TI - Reciprocal enhancement of gene transfer by combinatorial adenovirus transduction and plasmid DNA transfection in vitro and in vivo. AB - The addition of replication-defective recombinant adenovirus to plasmid transfection (termed here "adenofection") has been shown to increase plasmid transgene expression in limited studies. Similarly, the addition of cationic liposomes to adenovirus increases adenovirus-mediated gene transduction (termed here "lipoduction"). Here we demonstrate that adenofection was effective at enhancing transgene expression when used in conjunction with a variety of different transfection reagents, including a monocationic liposome, a polycationic liposome, an activated dendrimer, a large multilamellar liposomal vesicle, and a protein/amphipathic polyamine complex. The effect was seen regardless of the cellular expression of the adenovirus receptor, CAR, in three different human cancer cell lines derived from rhabdomyosarcomas (Rh18 and RD, CAR-) and cervical carcinoma (HeLa, CAR+). The protein/amphipathic polyamine complex showed an adenofection effect but did not show a lipoduction effect, consistent with different mechanisms of action for adenofection and lipoduction. Using dual-color flow cytometric analysis of cells transfected with a plasmid expressing the enhanced blue fluorescent protein (pEBFP) and a recombinant adenovirus expressing the green fluorescent protein (Ad5-GFP), we demonstrate that adenofection works primarily by increasing gene expression within a cell, whereas lipoduction increases the percentage of cells expressing the transgene. In addition, these studies show that both adenofection and lipoduction can occur simultaneously, further increasing gene transfer. The combination of lipofection and adenovirus transduction also prolonged the duration of transient gene expression and was generally no more toxic than lipofection alone. The enhancement of gene transfer was also seen after injection of complexes directly into subcutaneous human xenograft tumors. Therefore, more effective gene transfer in vitro and in vivo of either plasmid DNA, adenovirus DNA, or both can be achieved by combining liposomal transfection with adenoviral transduction. PMID- 10515462 TI - Changes in forebrain function from waking to REM sleep in depression: preliminary analyses of [18F]FDG PET studies. AB - Based on recent functional brain imaging studies of healthy human REM sleep, we hypothesized that alterations in REM sleep in mood disorder patients reflect a functional dysregulation within limbic and paralimbic forebrain structures during that sleep state. Six unipolar depressed subjects and eight healthy subjects underwent separate [18F]2-fluoro-2-deoxy-D-glucose ([18F]FDG) PET scans during waking and during their first REM period of sleep. Statistical parametric mapping contrasts were performed to detect changes in relative regional cerebral glucose metabolism (rCMRglu) from waking to REM sleep in each group as well as interactions in patterns of change between groups. Clinical and EEG sleep comparisons from an undisturbed night of sleep were also performed. In contrast to healthy control subjects, depressed patients did not show increases in rCMRglu in anterior paralimbic structures in REM sleep compared to waking. Depressed subjects showed greater increases from waking to REM sleep in rCMRglu in the tectal area and a series of left hemispheric areas including sensorimotor cortex, inferior temporal cortex, uncal gyrus-amygdala, and subicular complex than did the control subjects. These observations indicate that changes in limbic and paralimbic function from waking to REM sleep differ significantly from normal in depressed patients. PMID- 10515464 TI - Brain gray and white matter transverse relaxation time in schizophrenia. AB - Recent in vivo diffusion brain imaging studies of schizophrenic patients have revealed microstructural abnormalities, with low diffusion anisotropy present throughout much of cortical white matter. Brain anisotropy is produced when proton movement reflects physically restricted water movement, for example, by myelin sheaths. Conditions that increase self-diffusion, such as edema, may also alter the longitudinal and transverse relaxation time of protons, and it is possible that such changes could explain the observed anisotropy diminution seen in schizophrenia. To test this possibility, we calculated pixel-by-pixel transverse relaxation time (T2) and proton density (PD) maps for gray matter and white matter across eight 5-mm-thick axial slices of fast spin echo MRI in 10 control men (age 30-57 years) and 10 men with schizophrenia (age 32-64 years). Schizophrenics had significantly longer mean white matter T2 (84.0 vs. 81.9 ms, P<0.03) and gray matter T2 (95.1 vs. 92.2, P = 0.003); their mean white and gray matter PD values were not significantly different from those of controls. Correlations were not significant between anisotropy and T2 in either grey or white matter but were significant between anisotropy and PD in white matter. T2 relaxation times are longer in schizophrenics than in controls in both gray and white matter whereas anisotropy reduction is restricted to white matter. Taken together, these results suggest that the process producing prolonged T2 does not fully account for the abnormally low anisotropy observed selectively in white matter in this group of schizophrenic patients. PMID- 10515463 TI - Quantitation of benzodiazepine receptor binding with PET [11C]iomazenil and SPECT [123I]iomazenil: preliminary results of a direct comparison in healthy human subjects. AB - Although positron emission tomography (PET) and single photon emission computed tomography (SPECT) are increasingly used for quantitation of neuroreceptor binding, almost no studies to date have involved a direct comparison of the two. One study found a high level of agreement between the two techniques, although there was a systematic 30% increase in measures of benzodiazepine receptor binding in SPECT compared with PET. The purpose of the current study was to directly compare quantitation of benzodiazepine receptor binding in the same human subjects using PET and SPECT with high specific activity [11C]iomazenil and [123I]iomazenil, respectively. All subjects were administered a single bolus of high specific activity iomazenil labeled with 11C or 123I followed by dynamic PET or SPECT imaging of the brain. Arterial blood samples were obtained for measurement of metabolite-corrected radioligand in plasma. Compartmental modeling was used to fit values for kinetic rate constants of transfer of radioligand between plasma and brain compartments. These values were used for calculation of binding potential (BP = Bmax/Kd) and product of BP and the fraction of free non protein-bound parent compound (V3'). Mean values for V3' in PET and SPECT were as follows: temporal cortex 23+/-5 and 22+/-3 ml/g, frontal cortex23+/-6 and 22+/-3 ml/g, occipital cortex 28+/-3 and 31+/-5 ml/g, and striatum 4+/-4 and 7+/-4 ml/g. These preliminary findings indicate that PET and SPECT provide comparable results in quantitation of neuroreceptor binding in the human brain. PMID- 10515465 TI - Age of onset of depression and quantitative neuroanatomic measures: absence of specific correlates. AB - The purpose of our study was to examine the relationship between volumetric neuroanatomic measures and age of onset of illness in subjects with late-life major depression. Our sample was composed of 51 elderly subjects with Major Depressive Disorder who were scanned using a 1.5-tesla GE magnetic resonance imaging scanner with head coil. Absolute total and focal brain volumes were obtained together with quantified estimates of high intensity lesions. The relationship of these measures to onset age was examined using a regression while adjusting for subjects' current age and total intracranial volume. There was no significant linear relationship between age of onset of the first episode and any of the neuroanatomic measures examined. These data do not support the notion that neuroanatomical contributions to depression increase with a later age of onset of illness. PMID- 10515466 TI - First-episode schizophrenics show normal duration and topography of quasistationary EEG segments as compared to controls, during rest as well as during active tasks. AB - In patients suffering from chronic schizophrenia, both altered temporospatial structure of the EEG and impaired activation during cognitive tasks have repeatedly been demonstrated. The present study evaluates whether similar abnormalities are present in drug-naive first-episode schizophrenics. The EEGs of 32 schizophrenic patients and of 52 healthy controls were recorded during a simple and a complicated motor task, a simple and a complicated auditory stimulus, and during resting periods between the tasks. The temporospatial characteristics were evaluated by adaptive segmentation of EEG, which decomposes an EEG into temporal segments of quasistationary activity. No differences in the temporal and topographic aspects of the EEGs were found between the first-episode schizophrenic patients and the controls, neither during the resting EEGs nor during active tasks. Moreover, the dynamic course of the EEGs, defined as the alternation between task-related changes of temporospatial patterns and the reappearance of resting patterns, was identical in patients and controls. The present findings suggest that while abnormal EEG power spectra seem a consistent finding in treated as well as in never-treated schizophrenics, altered temporospatial patterns and reduced task-related EEG changes are inconsistent signs. PMID- 10515467 TI - Restorative neurosurgery: opportunities for restoration of function in acquired, degenerative, and idiopathic neurological diseases. AB - Historically, neurosurgery has improved the environment of the nervous system to promote maximal spontaneous recovery of function. The population of patients whom we treat at present is a small portion of those who suffer from disabling neurological illnesses. Based on a combination of new technology, and advances in neuroscience, restorative neurosurgery is advancing the frontiers of our specialty, and providing the potential to restore lost function. Significant advancements in gene therapy, the discovery and delivery of neurotrophic factors, and cell transplantation now require neurosurgeons to broaden the scope of our practice so that it includes the restoration of function in an enormous number of patients with acquired, degenerative and idiopathic neurological diseases. In order to meet the present challenge, neurosurgeons must broaden our vision, our role, and our future educational goals. In this review, we summarize the landmark advances in the basic and clinical neurosciences and the results of clinical trials that are driving our evolution from passive reaction to disease to active attempts to restore lost central nervous system function. PMID- 10515468 TI - Dose reduction improves hearing preservation rates after intracanalicular acoustic tumor radiosurgery. AB - OBJECTIVE: To assess the potential for long-term serviceable hearing preservation in intracanalicular acoustic tumor patients who underwent stereotactic radiosurgery. METHODS: Between August 1987 and December 1997, 29 patients with intracanalicular acoustic tumors underwent stereotactic radiosurgery at our center using the Leksell gamma knife (Elekta Instruments, Inc., Atlanta, GA). Fifteen assessable patients had serviceable preradiosurgery hearing (pure tone average, < or = 50 dB; speech discrimination score, > or = 50%). We retrospectively analyzed our hearing results and compared hearing preservation in patients who received a minimal tumor dose of 14 Gy or less versus those who received more than 14 Gy to the tumor margin. RESULTS: No perioperative patient morbidity or mortality was observed. Serviceable hearing was preserved in 11 (73%) of 15 assessable patients (actuarial rate, 65%). Long-term follow-up demonstrated serviceable hearing preservation in 10 (100%) of 10 patients who received marginal tumor doses of 14 Gy or less but in only one of five patients who received more than 14 Gy. Preradiosurgery Gardner-Robertson class was preserved in 49%, and testable hearing was present in 68% of patients who had any testable hearing at presentation. Five patients demonstrated improvement in hearing (three had serviceable and two had nonserviceable hearing before radiosurgery). No patient developed a facial or trigeminal neuropathy. Seven of 13 patients with preoperative tinnitus continued to experience tinnitus in follow up. Episodic vertigo continued in 3 of the 11 patients who presented with vertigo. CONCLUSION: Gamma knife radiosurgery (using conformal dose planning, small-beam geometry, and < or = 14 Gy to the margin) prevents tumor growth and achieves excellent hearing preservation rates. PMID- 10515469 TI - Spontaneous cerebrospinal fluid fistulae associated with empty sellae: surgical treatment and long-term results. AB - OBJECTIVE: To evaluate the accuracy of different diagnostic tools for cerebrospinal fluid fistulae (CFFs) associated with empty sellae and to assess the long-term results after surgical treatment. METHODS: Records were retrospectively reviewed for 38 of 44 patients who were treated for CFFs associated with empty sellae between 1972 and 1996. Glucose and radioisotope analyses of nasal fluids were used to establish the CFF diagnosis. Computed tomography, magnetic resonance imaging, and cisternography were used to localize the sites of the CFFs. Treatment was performed using three different approaches, i.e., subfrontal approach, transsphenoidal approach, and lumboperitoneal shunt placement. RESULTS: Glucose and radioisotope analyses of nasal fluids confirmed CFFs in 97 and 100% of the cases, respectively. For localization of the sites, a leak in the sella was suggested in 32% of the cases using gammagraphy, in 50% using computed tomography or magnetic resonance imaging, and in 75% using computed tomographic contrast cisternography. After surgery, rhinorrhea ceased for 34 patients (89%), but 14 (41%) patients experienced recurrences of the rhinorrhea 6 months to 20 years after surgery (mean, 4.2 yr). Cure rates for the subfrontal approach, transsphenoidal approach, and shunt placement were 94, 85, and 86%, and the recurrence rates were 43, 33, and 50%, with mean times of 7.8, 4.2, and 0.8 years, respectively. CONCLUSION: In this series, confirmation of CFFs was easy, although localization of the sites remained difficult. Recurrences were more numerous and occurred sooner after treatment with shunts; packing of the sella through a craniotomy achieved better and more stable results, although differences were not statistically significant. PMID- 10515470 TI - Peculiarities of intracranial arachnoid cysts: location, sidedness, and sex distribution in 126 consecutive patients. AB - OBJECTIVE: To study the distribution of intracranial arachnoid cysts in a large and nonbiased patient population. METHODS: One hundred twenty-six patients with 132 arachnoid cysts were studied. Patients were consecutively referred to our department during a 10-year period from a well-defined geographical area with a stable population. RESULTS: The cysts had a strong predilection for the middle cranial fossa; 86 patients (65.2%) had cysts in this location. Of 106 cysts with clearly unilateral distribution, 64 were located on the left side and 42 on the right side. This significant difference resulted solely from the marked preponderance of middle fossa cysts for the left (left-to-right ratio, 2.1:1). There were significantly more males than females (92 males/34 females). This difference was exclusively due to male preponderance of unilateral middle fossa cysts (66 males/14 females; ratio, 4.7:1). For all other cyst locations, there was no difference between the two sexes (26 males/20 females) or the two sides (10 left, 16 right). The marked left-sidedness for middle fossa cysts was found only in males. Females had an even distribution between the two sides. CONCLUSION: Arachnoid cysts have a strong predilection for the middle cranial fossa that may be explained by a meningeal maldevelopment theory: the arachnoid coverings of the temporal and frontal lobes fail to merge when the sylvian fissure is formed in early fetal life, thereby creating a noncommunicating fluid compartment entirely surrounded by arachnoid membranes. Why males develop more middle fossa cysts on the left side remains a mystery. PMID- 10515471 TI - Cost-effectiveness analysis of nimodipine treatment after aneurysmal subarachnoid hemorrhage and surgery. AB - OBJECTIVE: To assess the cost-effectiveness ratio of nimodipine administration after aneurysmal subarachnoid hemorrhage (SAH) and surgery. METHODS: One hundred twenty-seven patients of both sexes who had a ruptured aneurysm (verified using angiography), who presented with Hunt and Hess Grades I to III on admission, who underwent an operation within the first week after SAH, and who had participated in a randomized prospective clinical trial of nimodipine medication were enrolled in the study. The efficiency (cost-effectiveness) of nimodipine treatment was evaluated by incremental cost-effectiveness analysis. The cost-effectiveness ratio was evaluated for two groups: patients treated with nimodipine and patients given placebo. The cost was estimated as direct hospitalization costs, and the patient outcome was measured as life years gained. RESULTS: The incremental cost effectiveness ratio for nimodipine treatment was $223 per life year gained on the basis of 1996 monetary values and contemporary management of SAH. Patients in the nimodipine group had an average of 3.46 years longer life expectancy (incremental effectiveness) than those in the placebo group. There was a significant difference in 3-month follow-up mortality and a slight difference in sickness pensions during the 10 years after SAH. Nimodipine treatment was associated with a significant decrease in mortality. There were no statistically significant differences between the treatment groups in the length of hospital stay. There were no statistically significant differences between the treatment groups in sickness pensions. CONCLUSION: Nimodipine is cost-effective. Therefore, its use in the management of patients with SAH seems economically justified because it increases patient life years at very low incremental cost. PMID- 10515472 TI - Clinical outcomes for patients at high risk who underwent carotid endarterectomy with regional anesthesia. AB - OBJECTIVE: To compare the clinical outcomes for patients with carotid artery stenosis with advanced age, diabetes mellitus, atherosclerotic coronary vascular disease, and contralateral internal carotid artery occlusion who underwent carotid endarterectomy (CEA), using regional anesthesia, with the outcomes for patients without these risk factors. METHODS: A prospective series of 600 CEAs performed using regional anesthesia was analyzed. All patients were surgically treated under the direction of one neurosurgeon, in an academic medical center. Clinical outcome measures were any stroke, death, or cardiac morbidity within 30 days after surgery. All patients were monitored until a clinical end point was reached and/or 6 weeks had elapsed after surgery. The incidence of adverse clinical outcomes among the suspected high-risk patients was compared with the incidence for the entire series using contingency-table analysis (chi2 and Fisher's exact tests). RESULTS: Fifteen strokes (2.5%), three cardiac complications (0.5%), and two deaths (0.3%) occurred within 30 days after CEA. None of the suspected risk factors was associated with a significantly (P < 0.05) increased risk of perioperative morbidity or death. CONCLUSION: CEA using regional anesthesia can be performed for patients with advanced age, diabetes mellitus, atherosclerotic coronary vascular disease, and contralateral internal carotid artery occlusion, with acceptably low perioperative morbidity rates. PMID- 10515473 TI - Clinical and angiographic results of endosaccular coiling treatment of giant and very large intracranial aneurysms: a 7-year, single-center experience. AB - OBJECTIVE: To evaluate whether the objectives of surgical treatment, i.e., prevention of aneurysmal rebleeding, relief of aneurysmal mass effect, and prevention of embolic complications, are met by endosaccular coiling treatment applied to giant and very large wide-necked aneurysms. METHODS: Thirty patients with 31 giant or very large aneurysms were considered to show unacceptable risk/benefit ratios for open surgery and were treated using the Guglielmi detachable coil (GDC) method between 1992 and 1998. RESULTS: With endosaccular GDC treatment, 73.3% of the population experienced excellent to good recoveries (Glasgow Outcome Scale scores of 4 or 5), with a 13.3% procedure-related morbidity rate and a 6.7% procedure-related mortality rate. Two hemorrhaging episodes occurred after GDC treatment (annual bleeding rate, 2.5%; 2 hemorrhaging episodes/79.2 patient-yr). Symptoms related to aneurysmal mass effect were improved for 45.5% of the patients presenting with signs of neural compression. Among 23 patients with 24 aneurysms who were available for angiographic follow-up assessment, complete or nearly complete occlusion was observed for 17 aneurysms (71%; angiographic follow-up period, 24.3 +/- 19.6 mo, mean +/- standard deviation). A single total embolization served as definitive treatment for only 12.5% of the giant aneurysms and 31% of the very large aneurysms. CONCLUSION: Endosaccular GDC treatment of giant and very large aneurysms was accomplished with procedure-related morbidity and mortality rates comparable to those for open surgery performed by experts. However, because coil stability was unsatisfactory, we suggest that the GDC method should currently be reserved for individuals who are considered poor candidates for open surgery. PMID- 10515474 TI - Ethmoidal dural arteriovenous fistulae: an assessment of surgical and endovascular management. AB - OBJECTIVE: Endovascular treatment of ethmoidal dural arteriovenous fistulae (DAVFs) has become technically feasible, but its relative risks and benefits have not justified its use. We present a series of patients with ethmoidal DAVFs treated almost exclusively with surgery at an institution where expert endovascular therapy was available. Surgical risks, treatment efficacy, and patient outcomes were determined for comparison with published endovascular data. METHODS: Sixteen patients with ethmoidal DAVFs were treated during a 17-year period from 1982 to 1999. In three patients, feeding arteries from the internal maxillary artery were embolized; no ophthalmic artery embolizations were performed. A low bifrontal surgical approach was used in most patients to expose, coagulate, and divide the fistulous site. RESULTS: Ethmoidal DAVFs were occluded grossly and angiographically in all 16 patients. There was no treatment associated neurological morbidity, and clinical outcomes were good in all but one patient who was comatose initially. CONCLUSION: Review of our surgical experience with ethmoidal DAVFs as well as published endovascular results for these lesions suggests that endovascular management of ethmoidal DAVFs has a small but clinically significant risk to vision, is rarely effective in curing the fistula, and does not eliminate the need for surgery. In contrast, surgical management has no associated risk to vision, is highly effective at obliterating the fistula, and can contribute to good clinical outcomes in most patients. For these reasons, surgical management of ethmoidal DAVFs remains the treatment of choice. PMID- 10515476 TI - The pterional approach for the microsurgical removal of olfactory groove meningiomas. AB - OBJECTIVE: Currently, the surgical approach to olfactory meningiomas can vary depending on the size and expansion of the tumor, although surgical treatment still relies on the anterior bilateral craniotomy. Since 1989, we have use the pterional approach as a standard procedure in the treatment of 37 consecutive cases. We present our results in an attempt to contribute an alternative and valid surgical strategy for the treatment of these tumors. METHODS: Between 1989 and 1996, a series of 37 consecutive patients underwent microsurgical tumor resection using the unilateral pterional approach; all patients except one underwent operations on the right side. In 23 patients (62%), the tumor diameter measured approximately 6 cm, and the size was less than 4 cm in only 5 patients. The clinical presentation included mental dysfunction in 27 patients and visual impairment in 16 patients. The advantages of this approach are the early recognition of the posterior cerebrovascular complex, followed by a safe, rapid, and complete devascularization of the tumor and later by a favorable dissection of the capsular area from the frontal vascular branches and parenchyma. RESULTS: Total removal was achieved in all cases. There was one death unrelated to surgery. All patients presenting with mental dysfunction or with preoperative visual deficits recovered or improved. Postoperative magnetic resonance imaging confirmed complete tumor removal and demonstrated the brain parenchyma to be preserved and intact, primarily on the side opposite from the craniotomy. CONCLUSION: Our experience with the pterional approach suggests a greater role for this procedure in the treatment of olfactory groove meningiomas. PMID- 10515475 TI - Management of acute odontoid fractures with single-screw anterior fixation. AB - OBJECTIVE: Accepted management strategies for odontoid fractures include external immobilization and surgical stabilization using anterior or posterior approaches. Displaced Type II fractures and rostral Type III fractures are at high risk for nonunion. Anterior fixation of odontoid fractures with a single cortical lag screw is a relatively new technique that combines rigid internal stabilization with preservation of intrinsic C1-C2 motion. We retrospectively reviewed our series of 26 consecutive patients who underwent odontoid screw fixation, to further define the safety and efficacy of the technique. METHODS: During a 5-year period, 26 patients presented with acute traumatic Type II odontoid fractures. Ten patients were female and 16 were male, with a mean age of 35 years. All patients underwent anterior odontoid screw fixation by the senior surgeon (RWH), within a mean of 3 days after injury. All patients were postoperatively maintained in external orthoses, for a mean of 7.2 weeks, and were monitored with serial clinical and radiographic examinations. RESULTS: With a mean follow-up period of 30 months, radiographic fusion was documented for 25 of 26 patients (96%). No complications related to the surgical approach were identified, and all patients remained in neurologically stable condition. Two complications (8%) were related to the instrumentation; one patient required external immobilization because of suboptimal screw placement, and one patient required posterior atlantoaxial arthrodesis because of inadequate fracture reduction. CONCLUSION: Single-screw anterior odontoid fixation was associated with a relatively low complication rate and a high fusion rate in this study. We think that this should be the preferred treatment method for acute Type II odontoid fractures. PMID- 10515477 TI - Does lamina terminalis fenestration reduce the incidence of chronic hydrocephalus after subarachnoid hemorrhage? AB - OBJECTIVE: The incidence of chronic hydrocephalus requiring cerebrospinal fluid shunting was analyzed for a prospective series of 52 consecutive patients with ruptured cerebral aneurysms who underwent fenestration of the lamina terminalis during early microsurgical aneurysm repair. We hypothesized that, by creating an anterior ventriculocisternostomy, fenestration of the lamina terminalis would facilitate cerebrospinal fluid dynamics and decrease the risk of subsequent hydrocephalus. METHODS: Patients were enrolled according to the following criteria: 1) age more than 40 years; 2) admission Hunt and Hess Grade 2 to 4; 3) initial subarachnoid hemorrhage severity of Fisher Grade 3 or 4; and 4) early microsurgical repair of an anterior circulation aneurysm. RESULTS: The mortality rate in this series was 9.6%. Of the 47 surviving patients, 32 (68%) were discharged with a Glasgow Outcome Scale score of 5, 10 (21%) with a Glasgow Outcome Scale score of 4, and 5 (11%) with a Glasgow Outcome Scale score of 3. The follow-up period ranged from 12 to 60 months. Chronic hydrocephalus was radiographically and clinically evident in 3.8% of the total population. Shunt surgery was performed for two patients who exhibited symptoms resulting from hydrocephalus, corresponding to 4.2% of the surviving patients. CONCLUSION: Estimates from the most recently published studies indicate that an incidence of chronic post-subarachnoid hemorrhage hydrocephalus (requiring shunt surgery) of 15 to 20% is representative for an average contemporary population of patients with aneurysmal subarachnoid hemorrhage. The lower incidence of chronic hydrocephalus observed in this series possibly reflects the favorable effect of lamina terminalis fenestration on cerebrospinal fluid dynamics. PMID- 10515478 TI - Midthoracic catheter tip placement for intrathecal baclofen administration in children with quadriparetic spasticity. AB - OBJECTIVE: In an effort to increase the effect of intrathecal baclofen on upper extremity spasticity, the tip of the intrathecal catheter was placed at the T6-T7 level rather than at the traditional T11-T12 level in children with spastic quadriparesis. METHODS: Twelve children with spastic quadriparesis from varying causes had significant reductions in spasticity after a test dose of intrathecal baclofen and subsequently underwent placement of a programmable pump and intrathecal catheter tip placed at the T6-T7 level with fluoroscopic guidance. With the use of Ashworth scores for four muscle groups in both the upper and lower extremities, degrees of spasticity were determined by a physiatrist preoperatively and at 1, 3, 6, and 12 months postoperatively. Mean changes in upper- and lower-extremity Ashworth scores and baclofen dosages for the entire cohort were compared with published results in which the catheter tip had been placed at the T11-T12 level. RESULTS: Spasticity was significantly reduced in all muscle groups (P < 0.001). The lower-extremity reduction in spasticity of 1.6 points at 3 and 12 months was greater than published reductions of 1.1 points at 3 and 12 months. The upper-extremity reduction in spasticity was noticeably greater at 3 and 12 months (1.7 and 2.0 points, respectively) than published results at 3 and 12 months (0.4 and 0.6 points, respectively). At 3, 6, and 12 months, our mean baclofen dosage remained below the dosages administered at the T11-T12 level. There were no complications related either to the positioning of the catheter higher in the spinal canal or to the administration of baclofen at the T6-T7 level. CONCLUSION: Compared with published results, placement of the tip of the intrathecal catheter at the T6-T7 level was associated with greater relief of upper-extremity spasticity without loss of effect on the lower extremities. The mean dosages of baclofen in our study group were lower compared with mean dosages administered at the T11-T12 level. There was no morbidity related to the more rostral location of the catheter. PMID- 10515479 TI - Anticoagulation in neurosurgical patients. AB - OBJECTIVE: Few recommendations have been outlined in the neurosurgical literature regarding when it is safe to initiate postoperative or posthemorrhage anticoagulation (AC), or for what duration it is safe to discontinue AC therapy in patients with clear indications for AC therapy. Our objective was to formulate guidelines for managing AC in neurosurgical patients, based on patients' needs for AC and the risk of complications. METHODS: We conducted a systematic review of the neurosurgical and general surgical literature pertaining directly to postoperative or posthemorrhage management of AC. In addition, we surveyed the general medical, cardiology, cardiothoracic surgery, general surgery, vascular surgery, and neurology literature to determine the risk of thromboembolic complications when AC is stopped in specific patient groups. RESULTS: Postoperative bleeding complications occurred more frequently when correction of coagulation abnormalities was inadequate in the preoperative period, when AC was reinstituted in the early (24-48 h) postoperative period, and when AC was supratherapeutic in the postoperative period. Risk of significant thromboembolic complications while off AC varied significantly depending on the primary disease process necessitating AC. CONCLUSION: Adequate preoperative correction of coagulation abnormalities and strict regulation of coagulation to avoid supratherapeutic AC is essential. Reintroduction of AC after an intracranial hemorrhage treated without surgery, or after a neurosurgical procedure, particularly an intracranial procedure, can be guided by determining whether the patient is at high, moderate, or low risk for thromboembolic complications. On the basis of experimental studies, the patient's thromboembolic risk, and the experience of other surgeons, we propose therapeutic options for use of AC in neurosurgical patients undergoing intracranial procedures. PMID- 10515480 TI - Embryonic central nervous system transplants mediate adult dorsal root regeneration into host spinal cord. AB - OBJECTIVE: The aim of this study was to determine whether embryonic central nervous system transplants assisted cut dorsal root axons of adult rats to regenerate into the spinal cord. METHODS: Rats received transplants of embryonic spinal cord, hippocampus, or neocortex into dorsal quadrant cavities aspirated in the lumbar enlargement. The transected L5 dorsal root stump was secured between the transplant and the spinal cord. Regenerated dorsal roots were subsequently labeled by using immunohistochemical methods to detect calcitonin gene-related peptide. RESULTS: Calcitonin gene-related peptide-immunoreactive axons extended into all host spinal cords examined, but the patterns of regrowth differed in rats that had received embryonic spinal cord and brain transplants. In rats with embryonic spinal cord transplants, regenerated axons traversed the dorsal root/spinal cord interface, entered the spinal cord, and frequently formed plexuses with arborizations in motoneuron pools; some of these axons established synapses on spinal cord neurons. In rats with embryonic brain transplants, regenerated axons were diffusely distributed in the spinal cord but did not form plexuses. Few axons regenerated into the spinal cords of lesion-only animals. The results of quantitative analyses confirmed these findings. CONCLUSION: These findings suggest that transplants of embryonic spinal cord and brain supply cues that enable cut dorsal roots to regenerate into the host spinal cord and that the cues provided by spinal cord transplants favor more extensive growth than do those provided by brain transplants. These cues are likely to depend in part on neurotrophic effects of embryonic central nervous system tissues. Therefore, embryonic central nervous system transplants, especially spinal cord grafts, may contribute to techniques for restoring interrupted spinal reflex arcs. PMID- 10515481 TI - Effects of epidermal growth factor and dibutyryl cyclic adenosine monophosphate on the migration pattern of astrocytes grafted into adult rat brain. AB - OBJECTIVE: Neonatal rat astrocytes transplanted into host brains migrate in specific patterns, which are determined by the developmental stage of the host brain and the region of implantation. We hypothesized that the differentiation state of the implanted astrocytes could also affect astrocyte migration. METHODS: Astrocytes derived from neonatal rats (1-4 d) were placed in culture and exposed to growth- or differentiation-promoting agents (e.g., epidermal growth factor or dibutyryl cyclic adenosine monophosphate). Treated cells were then injected into different regions of the adult rat brain. At 3, 6, and 9 days after implantation, the extent and pattern of astrocyte migration after injection into the cortex, hippocampus, and corpus callosum were assessed. RESULTS: Astrocytes pretreated with either factor did not migrate during the first 3 days after implantation into the host cortex and hippocampus, whereas untreated cells migrated extensively by Day 3. After 9 days, implanted cells that had been pretreated with dibutyryl cyclic adenosine monophosphate began to demonstrate migratory activity, whereas those exposed to epidermal growth factor remained at the site of implantation. These findings corresponded to the effects of these agents in culture. On the other hand, cells implanted into the corpus callosum migrated in spite of pretreatment. CONCLUSION: Epidermal growth factor and dibutyryl cyclic adenosine monophosphate each altered the cells in culture such that they were inhibited from migrating after transplantation into the host cortex and hippocampus. This finding suggests that the activation of either growth or differentiation cascades partially inhibits the migratory ability in these cells either through effects on their internal migratory potentials or their responsiveness to external migratory signals. In contrast, cells implanted into the corpus callosum migrated in spite of pretreatment, suggesting that this structure may present migratory cues sufficient to override the effects of treatment. PMID- 10515482 TI - Prolongation of survival of mice with glioma treated with semiallogeneic fibroblasts secreting interleukin-2. AB - OBJECT: The current prognosis for patients with malignant brain tumors remains poor, and new therapeutic options are urgently needed. We previously have shown that prolongation of survival can be achieved in C57BL/6 mice (H-2b) with a syngeneic intracerebral or subcutaneous glioma when treated with allogeneic mouse fibroblasts (H-2k) genetically engineered to secrete interleukin-2 (IL-2). Like other antigens, tumor-associated antigens are recognized by cytotoxic T lymphocytes in the context of determinants specified by the major histocompatibility complex class I locus. Because the rejection of allogeneic major histocompatibility complex determinants has the property of an immune adjuvant, immunotherapy of glioma with a cellular immunogen that combines the expression of both syngeneic class I determinants and allogeneic antigens could have advantages over an immunogen that expresses syngeneic or allogeneic determinants alone. METHODS: To investigate this question in a mouse glioma model, we further modified allogeneic mouse fibroblasts (H-2k), not only for IL-2 secretion, but also for the expression of H-2Kb class I determinants. We tested the immunotherapeutic properties of these semiallogeneic cells (LM-IL-2/H-2Kb) in C57BL/6 mice with Gl261 glioma in both subcutaneous and intracerebral glioma models. RESULTS: C57BL/6 mice with either a subcutaneous or intracerebral glioma treated solely by injection of these IL-2-secreting semiallogeneic cells had significantly prolonged survival rates compared with untreated mice or mice treated with cells secreting only IL-2 or cells lacking the H-2Kb determinants. In some instances, the mice treated with the semiallogeneic cells survived indefinitely, suggesting total eradication of the glioma. When a 51Cr release assay was used, the specific immunocytotoxicity measured by release of isotopes from labeled Gl261 glioma cells coincubated with spleen cells from mice immunized with the semiallogeneic IL-2-secreting cells was significantly higher than that of spleen cells from nonimmunized mice or mice immunized with allogeneic cells lacking syngeneic major histocompatibility complex determinants. In addition, antibody depletion studies using monoclonal antibodies against CD8+ and natural killer/lymphokine-activated killer cells demonstrated a specific CD8+ immunocytotoxic response in animals immunized with the semiallogeneic IL-2 secreting cells compared with only a natural killer/lymphokine-activated killer response in mice immunized with the allogeneic IL-2-secreting cells. CONCLUSION: The augmented immune response against glioma in mice treated with the semiallogeneic IL-2-secreting cells points toward a new form of immunotherapy, "immuno-gene therapy," for patients with malignant intracerebral glioma. PMID- 10515484 TI - The development of anatomic art and sciences: the ceroplastica anatomic models of La Specola. AB - Anatomic models are important heuristic aids for surgeons in training. They are uniquely able to convey the three-dimensional relationships of anatomic structures with a physical immediacy not allowed by any other media. We examine the conceptual development of the anatomic model in light of the history of neuroanatomic understanding and coexistent artistic movements. The teaching anatomic model traces its ancestry to the work of Gaetano Zumbo in the late 17th century, on the heels of important anatomic discoveries made in the preceding 100 years of investigation. The anatomic model reached its peak expression in the late 18th century with the founding of the ceroplastica laboratory in Florence. We discuss the technological, artistic, and scientific origins of the anatomic wax model and the conditions that allowed it to flourish in the late 18th century. PMID- 10515483 TI - Dural closure with nonpenetrating clips prevents meningoneural adhesions: an experimental study in dogs. AB - OBJECTIVE: Meningospinal and cranial dural adhesions were compared in a canine model, after duraplasty using nonpenetrating clips or penetrating needles and sutures. METHODS: Fourteen dogs underwent bilateral craniotomies and duraplasties, with implantation of dural prostheses (DuraGuard; Biovascular Corp., Minneapolis, MN), using either 6-0 silk sutures or titanium clips (DuraClose; Surgical Dynamics, Norwalk, CT). Fourteen other dogs underwent L3-L4 laminectomies; three longitudinal dural incisions were closed with 6-0 silk sutures, 6-0 polyglactin 910 (Vicryl) sutures, or clips. Groups of eight dogs (four cranially treated and four spinally treated) were killed 6, 12, 24, and 52 weeks after surgery, and specimens were collected for study after perfusion and fixation (two cranial and two spinal dural reconstructions at 52 wk). Evaluations included assessment of the appearance of approximated dural margins and responses to clips, sutures, and dural prostheses (inflammation, foreign body reaction, fibrosis, and severity of meningospinal/meningocerebral adhesions). Data were evaluated using the Wilcoxon signed-rank and McNemar tests. RESULTS: Duraplasties with clips displayed significantly less extensive acute and chronic inflammation, foreign body reaction, and meningoneural adhesions than did repairs with needles and sutures. CONCLUSION: This report is the first long-term experimental study comparing two fundamentally different methods for dural repair in a relevant animal model. PMID- 10515485 TI - Integration of a variable action suction adapter into ultrasonic aspirators. AB - PURPOSE: Use of ultrasonic aspirators has become a mainstay in the neurosurgical armamentarium. The handpiece design and ultrasonic parameters have evolved to maximize its safe and efficacious use. Despite these modifications, continuous suction through the tip of the aspirator can result in neurovascular damage, especially when the aspirator is working in the cavernous sinus region or cerebellopontine angle. CONCEPT: We describe the integration of a variable suction adapter into the existing handpiece of an ultrasonic aspirator to minimize potential injury from the continuous forceful suction normally associated with the use of these devices. RATIONALE: The integration of such an adapter can reduce the potential for suction injury to cranial nerves or microvascular structures or smaller-caliber arteries and veins. DISCUSSION: This variable action suction adapter can decrease suction injuries to cranial nerves or the microvasculature. PMID- 10515486 TI - Technology and neurosurgery in developing countries: experience and present situation in Morocco. AB - OBJECTIVE: The high cost of technology is considered to be the determining factor slowing the expansion of modern neurosurgery in many developing countries. The literature dedicated to this topic rarely proposes internal solutions whereby affected countries can overcome this economic impediment. Certain articles cite inevitable obstacles, and the neurosurgeons of these countries can become disheartened when these articles conclude with calls for foreign help as the only approach to the development of neurosurgery. METHODS: Morocco is presented as an example of a developing country in which neurosurgery has become well established in the past 30 years, using a program based on four guidelines, as follows: 1) encouraging the local training of young neurosurgeons, 2) organizing and promoting neurosurgery, 3) integrating the development of neurosurgery into the health care pyramid system, and 4) stimulating research on local pathological conditions. RESULTS: Because of the internal planning efforts stimulated by the first national neurosurgeons, Morocco has progressed from 2 underequipped neurosurgical services and 5 neurosurgeons in 1968 to 12 well-equipped services and 80 neurosurgeons in 1998. The main benefits of this progress are discussed. CONCLUSION: Neurosurgery in developing countries can be promoted if the first working neurosurgeons take up their responsibilities as pioneers. This role requires that they initiate the training of young neurosurgeons as soon as possible and that they find in the local conditions the necessary factors to promote neurosurgery and to integrate it into the health care development of their country. PMID- 10515487 TI - Neurosurgery at the University of Lausanne. AB - The University of Lausanne was founded in 1537. The faculty of medicine was created in 1890, and the service of surgery was directed by Cesar Roux. Roux, a well-known surgeon, was visited by Harvey Cushing during 1900-1901. In the early 1930s, Jean Rossier from Lausanne trained with Cushing, but Rossier passed away in 1942. Eric Zander created the division of neurosurgery in 1959; it became an independent service in 1967. Nicolas de Tribolet served as chairman from 1984 until 1994, when he was asked to take charge of the merger of the university services of Geneva and Lausanne. In October 1997, Jean-Guy Villemure joined him in the newly merged department, becoming chairman in Lausanne, while de Tribolet is chairman in Geneva and head of the department comprising both services. PMID- 10515488 TI - Vascular compression of the medulla oblongata by the vertebral artery: report of two cases. AB - OBJECTIVE AND IMPORTANCE: Compression of the medulla oblongata by a tortuous vertebral artery is rare. We report two patients with this lesion who were treated with vascular decompression of the vertebral artery. CLINICAL PRESENTATION: A 36-year-old man developed right hemiparesis with lower cranial nerve deficits, and a 47-year-old man developed left lower cranial nerve deficits and left cerebellar dysfunction. In both patients, magnetic resonance imaging revealed a tortuous vertebral artery compressing the medulla oblongata. INTERVENTION: In both patients, the compressed medulla oblongata was treated by detaching the vertebral artery from the medulla oblongata, shifting it, and anchoring it to the nearby dura mater. Postoperatively, both patients are asymptomatic and have returned to their previous jobs. CONCLUSION: Although compression of the medulla oblongata by a tortuous vertebral artery is rare, it can cause brainstem dysfunction. Magnetic resonance imaging clearly revealed the vascular compression in these patients. Surgical treatment was effective. The symptoms related to a tortuous vertebral artery and some techniques for surgical treatment are discussed. Awareness of this rare lesion is necessary to ensure appropriate treatment. PMID- 10515489 TI - Anterior thalamoperforating artery aneurysm associated with internal carotid artery occlusion: case report. AB - OBJECTIVE AND IMPORTANCE: We describe a rare case of a ruptured distal anterior thalamoperforating artery aneurysm associated with right internal carotid artery occlusion. CLINICAL PRESENTATION: A 59-year-old woman experienced sudden occipital headache, vomiting, and subsequent coma as a result of massive intraventricular hemorrhage. An initial angiogram revealed only an occlusion of the right internal carotid artery just distal to the posterior communicating artery. Repeat angiography 1 month later, however, revealed a saccular aneurysm at a distal anterior thalamoperforating artery in addition to the occlusion of the internal carotid artery. INTERVENTION: We approached this aneurysm through the right temporal horn after opening the ambient cistern. The aneurysm, which was located in the brain parenchyma just medial to the temporal horn, was successfully resected. CONCLUSION: This rare aneurysm probably developed as a result of hemodynamic stress on the anterior thalamoperforating artery after occlusion of the internal carotid artery and/or secondary to chronic hypertension. PMID- 10515490 TI - Ectopic corticotroph adenoma in the cavernous sinus: case report. AB - OBJECTIVE AND IMPORTANCE: Adrenocorticotropin (ACTH)-secreting pituitary adenomas causing Cushing's disease are often difficult to identify because of their variable locations and their small size. This report presents histological evidence of an ectopic ACTH-secreting adenoma located entirely within the cavernous sinus. CLINICAL PRESENTATION: A 62-year-old woman presented with central obesity, hypertension, and osteoporosis. Endocrinological evaluation suggested the presence of an ACTH-secreting pituitary adenoma; however, imaging studies, including dynamic magnetic resonance imaging, did not reveal any visible lesions in the pituitary gland. Bilateral cavernous sinus sampling demonstrated a large central/peripheral ACTH gradient, with a right/left ACTH gradient. The patient was treated as having pituitary-dependent Cushing's disease, until she died suddenly as a result of acute respiratory failure. INTERVENTION: In a postmortem histological examination, an ACTH-secreting adenoma was found in the right cavernous sinus, which was completely surrounded by dura mater and had no direct connection with the pituitary gland. CONCLUSION: Although they are rare, such adenomas located in the cavernous sinus should be recognized as one of the reasons for inaccurate cavernous sinus sampling and the failure of transsphenoidal surgery for patients with ACTH-dependent Cushing's syndrome. PMID- 10515491 TI - Tandem intracranial stent deployment for treatment of an iatrogenic, flow limiting, basilar artery dissection: technical case report. AB - OBJECTIVE AND IMPORTANCE: Intimal dissection constitutes one of the complications associated with angioplasty of intracranial vessels. We present a case of iatrogenic dissection of the entire basilar artery, which was induced by angioplasty and stenting of symptomatic, focal, intracranial vertebral artery stenosis, and its successful treatment with tandem deployment of a downstream stent. CLINICAL PRESENTATION: A 61-year-old, hypertensive, renal transplant recipient presented with orthostatic vertebrobasilar insufficiency that was refractory to medical management, including anticoagulation therapy. Angiography revealed an occluded right vertebral artery and focal, high-grade, left intracranial vertebral artery stenosis. Magnetic resonance imaging showed multiple posterior fossa infarctions. The left intracranial vertebral artery stenosis was successfully treated with primary stent deployment and balloon angioplasty, with symptom resolution. On postprocedure Day 2, the patient noted worsening right hemiparesis. INTERVENTION: Subsequent angiography revealed a flow limiting, windsock-type, basilar artery dissection beginning at the distal end of the left vertebral artery stent and extending to the origin of the left posterior cerebral artery. A tandem stent was navigated intracranially and deployed past the first one, successfully sealing the dissection inflow zone and reconstituting normal flow to the top of the basilar artery. A clinical follow-up examination at 3 months revealed no further orthostatic symptoms and only mild residual right sided weakness. CONCLUSION: This is the first description of iatrogenic stent induced dissection of the entire basilar artery that was successfully treated by inflow zone control via tandem intracranial stent deployment. PMID- 10515492 TI - Percutaneous transluminal angioplasty and stenting of midbasilar stenoses: three technical case reports and literature review. AB - OBJECTIVE AND IMPORTANCE: Symptomatic basilar artery stenosis is a highly morbid disease process. Recent technological and pharmaceutical advances make endovascular treatment of this disease process possible. CLINICAL PRESENTATION: We report three cases of patients with a symptomatic basilar artery stenosis despite anticoagulation. INTERVENTION: All patients were successfully treated with a flexible coronary stent and perioperative antiplatelet medications without incident. Poststenting angiography demonstrated a normal-caliber artery with patent perforators. In one case, a poststenting cerebral blood flow study revealed improved perfusion. CONCLUSION: A new generation of stents and balloons makes access to intracranial intradural arterial pathological abnormalities possible. Such devices may well revolutionize the management of ischemic and hemorrhagic intracranial cerebrovascular disease. PMID- 10515493 TI - Embolization of a spinal arteriovenous malformation: correlation between motor evoked potentials and angiographic findings: technical case report. AB - OBJECTIVE AND IMPORTANCE: Endovascular procedures for the treatment of spinal arteriovenous malformations place the spinal cord at risk of ischemia. This report illustrates the usefulness of motor evoked potentials (MEPs) in detecting functional changes within the spinal cord motor pathways during embolization of a spinal arteriovenous malformation under general anesthesia. CLINICAL PRESENTATION: A 28-year-old man presented with a history of progressive lower extremity numbness and weakness followed by bladder dysfunction. Magnetic resonance imaging and angiography disclosed a T11-T12 spinal arteriovenous malformation. INTERVENTION: During the endovascular procedure, before injection of particles, the disappearance of MEPs from the tibialis anterior muscle led to prompt angiographic reevaluation, which disclosed the arrest of spinal blood flow secondary to radiculomedullary artery occlusion by the catheter. Embolization and catheter withdrawal were followed by temporary recovery of spinal blood flow and MEPs. A second arrest of spinal cord blood flow, caused by severe vasospasm of the feeding radiculomedullary artery, was documented by a control angiogram, and its functional relevance was revealed by a second disappearance of MEPs. The therapeutic effect of papaverine infusion and induced moderate hypertension was confirmed angiographically by complete reopacification of the anterior spinal artery and confirmed neurophysiologically by the complete recovery of MEPs. At the end of the procedure, no additional neurological deficits were noted. CONCLUSION: During spinal cord embolization, MEPs may play a critical role in early detection of spinal cord dysfunction by aiding in the prevention of damage to the spinal cord as well as by predicting the clinical outcome. PMID- 10515495 TI - Library: historical perspective. Norman M. Dott (1897-1973). PMID- 10515494 TI - New aneurysm clip and applier for narrow spaces: technical note. AB - OBJECTIVE: Aneurysm surgery carries considerable risk to the patient, in part because the surgical field can be so constricted that accurate placement of the aneurysm clip is impeded. In fact, most aneurysm clip appliers are so bulky that they can impair the surgeon's view, if the field is tight. We describe a system of aneurysm clips and appliers that have a very low profile and consequently allow better visualization of the operative field at critical moments during surgery. INSTRUMENTATION: A new system of clips for aneurysm surgery was developed to overcome some of the deficiencies of conventional sets. The new clip blades are opened from inside the spring, so the clip blades are clearly seen as they are applied across the base of the aneurysm. In addition, these clips can be angled in any desired direction in the applier, which makes their application easier. A third advantage is that the clip applier is used as a clip remover should adjustment or reapplication of the clip be necessary. RESULTS: Thirty aneurysms were obliterated with the new clips without any complications attributable to the clips. The new instruments allowed the safe application of clips to aneurysms in deep, confined, and anatomically complex spaces, such as aneurysms of the basilar tip and anterior communicating artery. CONCLUSION: The newly described system of aneurysm clips and appliers appears to have significant advantages over other currently available systems. PMID- 10515496 TI - Microvascular decompression of the left lateral medulla oblongata for severe refractory neurogenic hypertension. PMID- 10515497 TI - Preservation of the olfactory tract in bifrontal craniotomy for various lesions of the anterior cranial fossa. PMID- 10515498 TI - Emergency magnetic resonance imaging of cervical spinal cord injuries: clinical correlation and prognosis. PMID- 10515499 TI - Endoscopic third ventriculostomy: outcome analysis of 100 consecutive procedures. PMID- 10515500 TI - Arctic and Antarctic exploration including the contributions of physicians and effects of disease in the polar regions. PMID- 10515501 TI - Treatment of an intracranial aneurysm using a new three-dimensional-shape Guglielmi detachable coil: technical case report. PMID- 10515502 TI - Retroviral vector-mediated transfer and expression of human tissue plasminogen activator cDNA in bovine brain endothelial cells. AB - OBJECTIVES: Gene transfer of thrombolytic enzymes to vascular endothelial cells may influence the kinetics of intravascular thrombosis. This study defines the potential for gene transfer of tissue plasminogen activator (tPA) into bovine brain endothelial cells (BBEC). METHODS: The retroviral vectors derived from murine leukemia virus (MuLV) were used to transfer human tPA cDNA to BBEC. The tPA activity, tPA antigen and tPA inhibitor 1 (PAI-1) antigen were determined in the supernatant of transduced (BBEC/tPA) cell cultures by an immunoassay. RESULTS: The tPA antigen and enzymatic activity in cell culture supernatants of BBEC/tPA transduced cells were 75 ng/ml and 14 IU/ml after 4 days, that was 25 and 28-fold higher compared to the respective values in control cells. The PAI-1 antigen was not affected by tPA cDNA transfer. The Western blot assay of cell lysates confirmed that the majority of tPA in BBEC/tPA transduced cells was in the form of free tPA. While the maximal transduction efficiency of BBEC with an amphotropic MuLV vector was about 15%, a MuLV pseudotyped with vesicular stomatitis virus G glycoprotein envelope achieved high > 90% maximal transduction efficiency. CONCLUSIONS: The fibrinolytic activity of brain endothelial cells can be enhanced by transferring human tPA cDNA. These findings provide an initial step in implementation of future studies that investigate the use of this technology as an adjunctive treatment for cerebrovascular disease. PMID- 10515503 TI - Acoustic reflex detection using wide-band acoustic reflectance, admittance, and power measurements. AB - The measurement of the acoustic reflex threshold is a basic component of the diagnostic audiological test battery that may subject patients to potentially harmful sound pressures. A wide-band acoustic impedance and reflectance system (D. H. Keefe, R. Ling, & J. C. Bulen, 1992) was investigated as a means of obtaining reflex thresholds at a reduced level and as a means of providing a more complete characterization of the reflex than current clinical methods provide. Reflex thresholds obtained by measuring changes in wide-band admittance, reflectance, and power were at least 8 dB lower than those obtained with the standard clinical technique. These reflex-induced changes were accounted for by a simple oscillator model of the middle ear, assuming that the acoustic reflex results in an increase in stiffness. The results support further investigation of reflectance-based measures of the acoustic reflex as a clinical tool and as a means of studying the reflex mechanism. PMID- 10515504 TI - Speech-sound discrimination in school-age children: psychophysical and neurophysiologic measures. AB - This study measured behavioral speech-sound discrimination and a neurophysiologic correlate of discrimination in normal school-age children (ages 6 to 15) to determine if developmental effects exist. Just noticeable differences (JNDs) and mismatch responses (MMNs) were assessed for synthetic syllables that differed in third-formant onset frequency (/da-ga/) and formant transition duration (/ba wa/). These stimuli were selected because children with learning problems often find it difficult to discriminate rapid spectrotemporal changes like /da-ga/, whereas the ability to distinguish /ba-wa/ is relatively unimpaired. Results indicate that JNDs for /da-ga/ show no developmental effects and that JNDs for /ba-wa/ decrease slightly with age (although likely for task-related reasons). MMNs elicited by two /da-ga/ stimulus pairs (onset frequency differences = 20 Hz, 280 Hz) and three /ba-wa/ stimulus pairs (transition duration differences = 3, 5, 15 ms) showed no systematic or significant differences for onset latency, duration, or area as a function of age. Normative JND and MMN data are provided. These norms provide a metric against which children with suspected central auditory processing difficulties or auditory-based language disorders can be compared. PMID- 10515505 TI - Brief-tone frequency discrimination by children. AB - This study investigated maturational changes in children's ability to discriminate the frequency of short-duration tone pulses. Frequency difference limens (DLs) were measured for digitally generated 1000-Hz tones with pulse durations of 200, 50, and 20 ms using a two-alternative, two-interval, forced choice procedure. Participants were 16 5-year-old children; 10 children each in the age categories of 7, 9, and 11 years; and a control group of 10 young adults. Eleven of the 5-year-old children were unable to learn the experimental task. All children in the three older groups and the adults successfully completed the study. The five 5-year-old children who completed the task performed similarly to the 7-year-old children. All groups of participants showed an inverse relationship between duration of the signal and the size of the DL. The DLs at all three tone durations were significantly larger for the 7-year-old children than they were for the older children and adults. There were no significant differences in DL size among the 9-year-old, 11-year-old, and adult subjects at any tone duration. These findings suggest that the sensory and/or cognitive skills required to discriminate the frequency of brief-duration tones may not reach maturity until after age 7 years. PMID- 10515506 TI - Speech perception and verbal memory in children with and without histories of otitis media. AB - Two groups of children, with (n = 7) and without (n = 7) first-year histories of otitis media, were participants in a longitudinal study that included periodic audiological and medical evaluations during the first year of life. At age 9, these children were tested on a series of speech perception and verbal short-term memory tasks using stimuli of varying degrees of phonetic contrast. Although the otitis-positive group performed less accurately than the otitis-free group, the pattern of errors was the same for the two groups. The performances of the children with and without positive histories of otitis media were negatively affected by an increase in phonetic similarity of the stimulus items. The two groups, however, did not differ on identification or on temporal-order recall when the speech sounds were differentiated by multiple features. These findings provide evidence of subtle, long-term effects of early episodes of otitis media on phonological representations and on working memory. PMID- 10515507 TI - An acoustic analysis of the development of CV coarticulation: a case study. AB - This study analyzed stop consonant-vowel productions from babbling to meaningful speech in a single female child spanning the period from age 7 months to age 40 months. A total of 7,888 utterances (3,103 [bV], 3,236 [dV], and 1,549 [gV]) were analyzed to obtain frequencies at F2 onset and F2 at vocalic center for each utterance. A linear regression line ("locus equation") was fit to the cluster of F2 coordinates per stop place category produced during each month. The slope of the regression lines provided a numerical index of vowel-induced coarticulation on consonant productions. Labial, alveolar, and velar CV productions followed distinct articulatory paths toward adult-like norms of coarticulation. Inferences about the gradual emergence of segmental independence of the consonant and vowel in the three stop place environments were made from locus equation scatterplots and mean F2 onset and F2 midvowel frequencies obtained across babbling, early words, and natural speech. PMID- 10515508 TI - Early childhood stuttering I: persistency and recovery rates. AB - The divergent developmental course of stuttering with its two major paths, persistency and spontaneous (unaided) recovery, has been a focus of scientific attention because of its critical theoretical, research, and clinical perspectives. Issues concerning factors underlying persistency and recovery and their implications for early intervention have stirred considerable controversy among scientists. In light of the intense interest, the scarcity of direct essential epidemiological data concerning the magnitude of the two paths and the timing of recovery is problematic. Most past studies have used retrospective methodologies. The few longitudinal studies have been severely limited in scope or objective data. The purpose of the investigation reported herein is to study the pathognomonic course of stuttering during its first several years in early childhood with special reference to the occurrence of persistent and spontaneously recovered forms of the disorder. Employing longitudinal methodology with thorough, frequent periodic follow-up observations, multiple testing, and recording of extensive speech samples, 147 preschool children who stutter have been closely followed for several years from near the onset of stuttering. In this, the first of three related articles, we present findings regarding the current stuttering status of 84 of these children, who have been followed for a minimum of 4 years after their onset of stuttering. The data indicate continuous diminution in the frequency and severity of stuttering over time as many children progressed toward recovery. Our findings lead to conservative estimates of 74% overall recovery and 26% persistency rates. The process of reaching complete recovery varied in length among the children and was distributed over a period of 4 years after onset. Detailed analyses of phonological and language skills pertaining to differentiation of the developmental paths of children who persist and those who recover are presented in the two other articles in the series (E. P. Paden et al., 1999, and R. V. Watkins et al., 1999). PMID- 10515509 TI - Early childhood stuttering II: initial status of phonological abilities. AB - Research on the relation between stuttering and phonological/articulation deficits has been reported in the literature over several decades. Yet virtually none of these investigations has taken into account that "children who stutter" includes a large number who spontaneously recover within a few months or years after onset. Thus, little attention has been given to differences between the phonological abilities of children whose stuttering persists and those who recover. This investigation compares these two groups soon after stuttering onset, before it was possible to classify them as members of either group, on a number of phonological characteristics, including mean percentage of error, relative levels of severity of phonological impairment, error on specific phonological patterns, progress in development of key patterns, and the children's strategies for coping with unmastered patterns. The results indicate that the children whose stuttering would be persistent had poorer mean scores on each of our measures than did the children who would recover from stuttering. Both groups, however, showed progression in phonological development that followed the expected order, and they used typical strategies when patterns had not yet been acquired. The persistent group was moving more slowly, however, so phonological development was more delayed than in the children who would recover from stuttering. Our findings support the assumption that most previous studies probably have compared children with persistent stuttering to normally fluent children, and that those who recovered early were not considered differentially. PMID- 10515510 TI - Early childhood stuttering III: initial status of expressive language abilities. AB - This investigation evaluated the expressive language abilities of 84 preschool age children who stuttered, 62 who recovered from stuttering, and 22 who persisted in stuttering. The participants were identical to those identified in E. Yairi and N. G. Ambrose (1999) and E. Paden, E. Yairi, and N. G. Ambrose (1999). A range of lexical, morphological, and syntactic measures--calculated from spontaneous language samples of approximately 250-300 utterances in length collected relatively near stuttering onset--were used to examine the children's expressive language skills. For the purpose of analysis and comparison to normative data, children were grouped into three age intervals, in terms of the age at which they entered the study (2- to 3-year-olds, 3- to 4-year-olds, and 4- to 5-year-olds). Findings revealed similarity in the expressive language abilities of children whose stuttering persisted as opposed to abated at all age intervals. In addition, persistent and recovered stutterers displayed expressive language abilities near or above developmental expectations, based on comparison with normative data, at all age intervals. Children who entered the study at the youngest age level consistently demonstrated expressive language abilities well above normative expectations; this pattern was found for both persistent and recovered groups. These findings provide relatively limited information to assist in the early differentiation of persistence in or recovery from stuttering, but they do shed light on theoretical issues regarding the nature and character of early stuttering and potential associations with language learning. PMID- 10515512 TI - The effects of a flattened fundamental frequency on intelligibility at the sentence level. AB - The purpose of this preliminary experiment was to evaluate the effect of a flattened fundamental frequency (F0) contour on sentence intelligibility. The perceptual dimension monotone pitch is frequently used to describe the speech of persons with dysarthria, and relatively flat F0 contours have been noted in several acoustic studies of dysarthria. To determine the independent effect of a flattened F0 contour on sentence intelligibility a resynthesis technique was used that held timing and spectral characteristics of utterances constant while allowing parametric control over successive pitch periods. Two male speakers produced low-probability utterances selected from the SPIN test, which were then resynthesized with a flattened F0 contour. Speech intelligibility was assessed using two measures: one involving word transcription and the other interval scaling. These measures were collected from 10 listeners. The results showed that both measures were significantly lower when the F0 contour was flattened, as compared with naturally varying contours. Several different explanations are proposed for this effect, which can and should be explored in greater detail using the resynthesis technique given the prominence of this characteristic in dysarthria. PMID- 10515511 TI - Estimation of alveolar pressure during speech using direct measures of tracheal pressure. AB - The pressure in the alveoli of the lungs, created by the elastic recoil of the lungs and respiratory muscle activity, is referred to as alveolar pressure (Pa). The extent to which tracheal pressure (Pt) approximates Pa depends on the resistance to airflow offered by structures above and below the point at which tracheal pressure is measured. An understanding of the relationship among Pa, Pt, and upper and lower airway resistance, and how these values fluctuate during speech, could aid in interpretation and modeling of speech aerodynamics. The purpose of this study was to (a) obtain values for lower airway resistance (Rlow), (b) use these Rlow values to estimate Pa during speech, and (c) quantify the degree to which Pt approximates Pa during production of voiced and voiceless sounds, in comparison to inhalation. In addition, the results were discussed in terms of the degree to which the respiratory system functions as a pressure source. Tracheal pressure (obtained with tracheal puncture) and airflow were measured during sentence production in 6 subjects. Using a technique introduced in this paper, Rlow was determined from measures of tracheal pressure and flow obtained during a sudden change in upper airway resistance because of release of a voiceless plosive. Mean Rlow values ranged from 0.14 to 0.32 kPa/(l/s). Each subject's mean Rlow was used to derive a time-varying measure of Pa during speech from continuous measures of tracheal pressure and airflow. Pt was approximately 95% of Pa during phonation (i.e., when the vocal folds were adducted), 75% of Pa during release of the voiceless stop consonant /p/, and 55% of Pa during inhalation (i.e., when the vocal folds were abducted). Therefore, the degree to which the respiratory system functioned as an ideal pressure source varied during speech. The ability to estimate Pa provides a measure of the pressure produced by the respiratory system that is not influenced by laryngeal activity. PMID- 10515513 TI - Speech and oral motor learning in individuals with cerebellar atrophy. AB - The purpose of this study was to determine whether cerebellar pathology interferes with motor learning for either speech or novel tasks. Practice effects were contrasted between persons with cerebellar cortical atrophy (CCA) and control participants on previously learned real speech, nonsense speech, and novel nonspeech oral-movement tasks. Studies of limb motor learning suggested that control participants would evidence reduced variability, increased speed of movement, and reduced movement amplitude with practice as compared with the CCA group. No significant differences were found between the real- and nonsense speech tasks. For both speech tasks, although neither group reduced their movement variability with practice, both groups significantly reduced jaw closing displacement and velocity with practice. For the novel nonspeech oral-movement task, no change with practice was observed in either group in terms of variability, amplitude, or peak velocity. No effects of cerebellar pathology were seen in either the speech- or oral-movement tasks. These results demonstrated that with practice of speech tasks, a previously learned motor skill, movement speed and displacement decreased in both groups. Therefore, the effects of practice differed between previously learned speech tasks and the novel oral movement task regardless of cerebellar pathology. PMID- 10515514 TI - Voice and speech characteristics of persons with Parkinson's disease pre- and post-pallidotomy surgery: preliminary findings. AB - Pallidotomy surgery, lesioning the globus pallidus internal, has been performed to alleviate Parkinsonian symptoms and drug-induced dyskinesias. Improvements in limb motor function have been reported in recent years following pallidotomy surgery. The purpose of this preliminary study was to determine the effect of pallidotomy surgery on select voice and speech characteristics of 6 patients with Parkinson's disease. Acoustic measures were analyzed pre-pallidotomy surgery and again at 3 months following surgery. Preliminary findings indicated that all participants demonstrated positive changes in at least one acoustic measure; 2 of the participants consistently demonstrated positive changes in phonatory and articulatory measures, whereas 3 participants did not consistently demonstrate positive changes postsurgery. The results are discussed relative to the differential effects observed across participants. PMID- 10515515 TI - Classification of children with specific language impairment: longitudinal considerations. AB - This paper reports on the longitudinal results of a large project involving 242 seven-year-old children attending language units in England. Following our work outlining 6 subgroups of children with language impairment (Conti-Ramsden, Crutchley, & Botting, 1997), we examine the stability of the 6 subgroups of children with specific language impairment already identified, using data collected from the same children at age 8 years. The findings suggest there is considerable stability in the patterns of difficulties delineated by the classification system involving 6 subgroups. Poorer stability was evident in the classification of the children across time with 45% of children moving across subgroups. The membership stability of the proposed classification system was very similar to that found when the children were classified into 3 subgroups following another well-known system (Rapin, 1996). The findings are discussed with particular reference to issues surrounding the classification of children with SLI. PMID- 10515516 TI - Testing the generalized slowing hypothesis in specific language impairment. AB - This study investigated the proposition that children with specific language impairment (SLI) show a generalized slowing of response time (RT) across tasks compared to chronological-age (CA) peers. Three different theoretical models consistent with the hypothesis of generalized slowing--a proportional, linear, and nonlinear model--were examined using regression analyses of group RT data. Each model was an excellent fit with the RT data. The most parsimonious model indicated that the SLI group was proportionally slower than the CA group. Mean RTs of the SLI group were about one fifth slower across tasks than the CA group's mean RTs. Less slowing was evident for a subgroup of young children with expressive SLI than for children with mixed (expressive and receptive) SLI. Although the mean RT data reflected many individual SLI children's RT performance, not all SLI children showed generalized slowing. PMID- 10515517 TI - Guiding language development: how African American mothers and their infants structure play interactions. AB - This investigation explored how African American mothers and their infants at the single-word stage of development structured their play and communicated with one another. Six mother-child dyads of low socioeconomic status (SES) and six of middle SES were observed at play. Few group differences were found, with the majority of the differences involving language behaviors. The middle-SES dyads included language goals more often in their play. Middle-SES infants initiated play verbally more frequently and produced over twice as many vocalizations as their low-SES peers. In addition, middle-SES mothers used a wider variety of words when playing with their children than their low-SES counterparts. A range of play styles was found within both groups. These were categorized into three general play styles: mothers and children actively involved in play; mothers' involvement varied; and children actively engaged and mothers attentive. PMID- 10515518 TI - Effects of treatment on linguistic and social skills in toddlers with delayed language development. AB - This study investigated the effects of early language intervention on various linguistic and social skills of late-talking toddlers. The 21 children who participated in the investigation were randomly assigned to an experimental group (n = 11) or a control (delayed-treatment) group (n = 10). The experimental group participated in a 12-week clinician-implemented language intervention program. Groups were compared at pretest and posttest on five linguistic variables: Mean Length of Utterance, Total Number of Words, Number of Different Words, Lexical Repertoire, and Percentage of Intelligible Utterances, as well as on Socialization and Parental Stress. Significant group differences were found for each of the variables, indicating facilitative effects of the treatment. Notably, increases were observed in areas that were not specifically targeted by the intervention. Implications of these results are discussed with respect to considerations regarding clinical management decisions for toddlers with delayed language development. PMID- 10515519 TI - An examination of verbal working memory capacity in children with specific language impairment. AB - This study investigated verbal working memory capacity in children with specific language impairment (SLI). The task employed in this study was the Competing Language Processing Task (CLPT) developed by Gaulin and Campbell (1994). A total of 40 school-age children participated in this investigation, including 20 with SLI and 20 normal language (NL) age-matched controls. Results indicated that the SLI and NL groups performed similarly in terms of true/false comprehension items, but that the children with SLI evidenced significantly poorer word recall than the NL controls, even when differences in nonverbal cognitive scores were statistically controlled. Distinct patterns of word-recall errors were observed for the SLI and NL groups, as well as different patterns of associations between CLPT word recall and performance on nonverbal cognitive and language measures. The findings are interpreted within the framework of a limited-capacity model of language processing. PMID- 10515520 TI - Children with SLI use argument structure cues to learn verbs. AB - Across two tasks, children's use of argument structure cues to learn verbs was tested. In Task 1, we examined children's use of cues to interpret novel verbs while viewing single action scenes. In Task 2, we examined the role of cues for novel verb interpretation and retention through a story viewing task. The participants were 20 6-year-olds who were diagnosed as specifically language impaired (SLI) and 40 normally developing children who served as either age matched or language-matched controls. Across tasks, the children with SLI demonstrated an ability to use cues to interpret verb meaning. For Task 1, scores of the children with SLI were not significantly different from those of either control group; for Task 2, their scores exceeded chance and were not found to be different from those obtained by the language-matched controls. When verb retention was examined, scores of the children with SLI were lower than those of both control groups, and they also did not exceed chance even after repeated exposure to the stimuli and additional testing. Patterns within the data ruled out inattention and an inability to follow the narrative as contributing to the children's low scores. Additionally, poor verb retention was not found to be related to a limitation in the perception and encoding of the cue content. Specific deficits with the storage and retrieval of grammatical information within the lexicon, general working memory/capacity limitations, or both are posited as plausible, but unconfirmed, explanations for the verb retention difficulties of the children with SLI. PMID- 10515521 TI - Present and future possibilities for defining a phenotype for specific language impairment. AB - This brief report summarizes a workshop that was held at the National Institutes of Health in April 1998. The goal of the workshop was to further the development of a definition for the phenotype of specific language impairment (SLI). The report includes a discussion of research recommendations that will refine our current views of the definition of the SLI phenotype and sets out priority areas that are in need of further study to help advance understanding of this complex language-based disorder. PMID- 10515522 TI - Facing up to the diagnostic uncertainty and management of onychomycoses. PMID- 10515523 TI - The scope of onychomycosis: epidemiology and clinical features. PMID- 10515524 TI - Quality of life for patients with onychomycosis. PMID- 10515525 TI - Onychomycosis in a special patient population: focus on the diabetic. PMID- 10515526 TI - Pharmacokinetics of orally administered antifungals in onychomycosis. PMID- 10515527 TI - The use of intermittent itraconazole therapy for superficial mycotic infections: a review and update on the 'one week' approach. PMID- 10515528 TI - Safety review of the oral antifungal agents used to treat superficial mycoses. PMID- 10515529 TI - Pharmacoeconomic analysis of the new oral antifungal agents used to treat toenail onychomycosis in the USA. PMID- 10515530 TI - New developments in antifungals. PMID- 10515531 TI - Gastroduodenostomy after gastric resection for cancer. AB - Gastrojejunostomy after resection for gastric cancer has been associated with a variety of complications, including bile reflux gastritis, marginal ulcers, and afferent loop syndrome. Gastroduodenostomy, although more physiologic, has not been recommended because of the fear of obstruction due to tumor recurrence. A review of 62 patients with gastric adenocarcinoma who underwent gastric resection from 1986 to 1996 was performed. Of the 62 patients, 52 (83%) underwent subtotal gastric resection and 10 (17%) underwent total gastrectomy with Roux-en-Y reconstruction due to tumor location. Forty-seven (90%) of the 52 patients underwent gastroduodenostomy, and 5 (10%) of these patients underwent gastrojejunostomy due to operative findings of excessive tumor burden or the inability to create a safe tension-free anastomosis. Patients who underwent gastroduodenostomies were followed for a period of 6 months to 5 years and had a median survival of 2.5 years. Two (4.2%) of the 47 patients obstructed due to tumor recurrence at 2 and 4 years postoperatively. One patient (2%) had symptomatic bile reflux gastritis, which was treated conservatively without reoperation. There were no incidences of marginal ulcers. In conclusion, gastroduodenostomy should be considered for patients undergoing resection for gastric cancer due to its physiological benefits and acceptable rate of obstruction. PMID- 10515532 TI - Simplifying the Waterston's stratification of infants with tracheoesophageal fistula. AB - The survival of infants with tracheoesophageal fistula was stratified by David J. Waterston et al. in 1962. This classification has been used as a guide to direct the timing of operative intervention in these infants. This study examines the current applicability of this classification system. The hospital records of 64 infants with esophageal atresia and/or tracheoesophageal fistula were reviewed. The survival rate was analyzed as a function of the infants' risk stratification, birth weight, and additional anomalies. Twenty-three infants were in Waterston Group A, 20 infants in Group B, and 21 infants in Group C. The survival of all infants was 81 per cent. Six infants died after recognition of severe anomalies and withdrawal of care, four infants died of cardiopulmonary arrest, and two infants died of sepsis. The survival of infants in both Groups A and B was 100 per cent, in contrast to 43 per cent survival in Group C. Only infants who weighed <1800 g or had severe additional anomalies were at risk of dying. Therefore, the classification of infants with esophageal atresia and/or tracheoesophageal fistula may be simplified by combining Waterston's Groups A and B into a single risk stratum. PMID- 10515533 TI - Abnormal esophageal body function: radiographic-manometric correlation. AB - Stationary manometry is the gold standard for the evaluation of patients with suspected esophageal motility disorders. Comparison of videoesophagram in the evaluation of esophageal motility disorders with stationary motility has not been objectively studied. Two hundred two patients with foregut symptoms underwent stationary motility and videoesophagram. Radiographic assessment of esophageal motility was done by video recording of five 10-cc swallows of barium. Abnormal esophageal body function was defined by stasis of barium in the middle third of the esophagus on at least four swallows or stasis on at least three swallows in the distal third. Stationary manometry was performed using a five-channel water perfused system. Contraction amplitudes <25 mm Hg in any of the last two channels or the presence of simultaneous or interrupted waves in 10 per cent or more were considered to be abnormal. Sixty-two patients had abnormal manometry. Thirty-four patients also demonstrated abnormal videoesophagrams for an overall sensitivity of 55 per cent. The positive predictive value was 53 per cent; specificity was 79 per cent; and negative predictive value was 80 per cent. Sensitivity was greatest in patients with achalasia (94%) and scleroderma (100%) and in patients presenting with dysphagia (89%). Sensitivity was poor for nonspecific esophageal motility disorders. A videoesophagram is relatively insensitive in detecting motility disorders. It seems most useful in the detection of patients with esophageal dysfunction, for which surgical treatment is beneficial, and in those patients presenting with dysphagia. PMID- 10515535 TI - The utility and reliability of computed tomography scan in the diagnosis of small bowel obstruction. AB - Accurate diagnosis and treatment of small bowel obstruction (SBO) is critical to prevent complications and reduce costs. The purpose of the current study was to evaluate the evolving role of CT scan for patients with suspected SBO and compare its sensitivity and specificity with plain abdominal films. We identified 108 patients over a 1-year period with an admission or discharge diagnosis of SBO. Five patients treated on a clinical basis without radiographic imaging and those with indeterminate films were excluded from analysis. CT and radiograph reports were compared with operative findings or clinical course in 103 patients. By the clinical course, 66 of the patients had a partial or complete SBO. Plain film radiography correctly established the diagnosis of SBO in 50 of 66 patients (sensitivity, 75%). Nine of 17 plain films were true negatives (specificity, 53%). CT was able to correctly identify partial or complete SBO in 35 of 38 patients (sensitivity, 92%) and correctly identified the lack of obstruction in 8 of 13 patients (specificity, 71%). These data document that plain films are a less sensitive and less specific method of diagnosing SBO and confirm that the use of CT scan is a valuable modality. CT scan should be considered for use as the primary test for patients with suspected SBO. PMID- 10515534 TI - Signet ring cell histology is associated with unique clinical features but does not affect gastric cancer survival. AB - Signet ring cell histology is found in 3 to 39 per cent of gastric cancer cases and has been reported to be a feature of poor prognosis, although this issue has not been rigorously examined. The objective of this study was to determine those demographic and clinical variables associated with signet ring cell histology and to determine the effect of signet ring cell histology on survival using multivariate analyses. We studied a historical cohort of consecutive cases of gastric cancer reported to the population-based California Cancer Registries of Orange, San Diego, and Imperial Counties from 1984 through 1994. Factors associated with signet ring cell histology were assessed using chi2 and logistic regression. Life tables were constructed to assess unadjusted survival and survival differences in patient subgroups. Multivariate survival was determined using a Cox proportional hazards model. Of 3020 patients, 464 (15%) had signet ring cell histology. Patients with signet ring cell histology were more likely to be younger than 50 years (odds ratio (OR) = 2.4; 95% confidence interval (CI) = 1.6-3.5), less likely to be male (OR = 0.49; 95% CI = 0.37-0.66), and more likely to have tumors of the distal stomach (OR = 2.0; 95% CI = 1.4-3.0). Signet ring cell histology did not adversely affect unadjusted overall survival, race stratified survival, or stage-stratified survival. Multivariate analysis indicated that patients with signet ring cell histology had an insignificant increased risk of dying (relative risk = 1.027; P>0.10) in comparison with patients without signet ring cell histology. Patients with signet ring cell histology were more likely to be young women and to have tumors of the distal stomach. Signet ring cell histology did not impact survival in our group of largely advanced gastric cancer cases. PMID- 10515536 TI - Same admission colon resection with primary anastomosis for acute diverticulitis. AB - Current standard of care for complicated diverticulitis includes urgent resection with colostomy versus antibiotic treatment, followed by delayed resection with primary anastomosis at a second admission. In certain circumstances, it is possible to perform resection and anastomosis on the same admission for acute diverticulitis. A retrospective review was completed for patients undergoing surgery for diverticulitis from 1991 to 1998. Groups included: 1) sigmoid resection with primary anastomosis on same admission (n = 18); 2) resection with protective end colostomy (n = 16); and 3) in-patient antibiotic treatment alone, followed by a second admission for resection with primary anastomosis (n = 5). Four patients initially treated with antibiotics worsened symptomatically or developed radiographic evidence of perforation and required resection with colostomy. Five patients in Group 1 had abscesses or contained perforations based on radiographic studies. Findings on CT scans did not predict treatment. Group 1 patients had uneventful recoveries and few minor complications (wound infections and an incisional hernia). One anastomotic leak occurred in Group 2 after colostomy closure. Although there will continue to be a role for emergent operation for diverticulitis, same admission sigmoid resection with primary anastomosis after antibiotic treatment is safe, uses a shorter course of antibiotics, and has a low complication rate. PMID- 10515537 TI - Primary resection and anastomosis for perforated left colon lesions. AB - The records of 33 patients with perforated left colon lesions over a 6-year period from 1992 to 1998 were examined retrospectively for clinical course and complications. All patients had a free perforation, feculent or purulent peritonitis, and/or a large inflammatory mass. All patients had primary resection and anastomosis without a protective colostomy or ileostomy. Indications included diverticulitis in 28 patients, obstructing colorectal carcinoma in 3, and iatrogenic perforations in 2. Complications occurred in 10 patients, including atelectasis in 1, urinary retention in 2, urinary tract infections in 2, wound infections in 3, line catheter sepsis in 1, and acute tubular necrosis in 1. There was one anastomotic dehiscence, which was successfully converted to a Hartmann procedure. Patients were discharged an average of 7 days after surgery. Two patients required surgery after discharge: one had a fascial dehiscence and the other an incisional ventral hernia. There was no mortality. Primary resection and anastomosis of selected perforated left colon lesions can be performed with a morbidity and mortality rate lower than that usually reported for the Hartmann procedure. PMID- 10515538 TI - Clinical use of a bioartificial liver in the treatment of acetaminophen-induced fulminant hepatic failure. AB - Patients with acetaminophen-induced fulminant hepatic failure (FHF) who meet the King's College Hospital criteria have a high mortality risk (>90%) if they do not undergo liver transplantation. We have developed a treatment strategy for these patients based on the use of an extracorporeal bioartificial liver (BAL) support system. In this study, we report the results of the clinical application of BAL support in patients with acetaminophen-induced FHF. All patients were admitted to a dedicated surgical intensive care unit. They were evaluated for urgent liver transplantation and received the standard medical measures, including N acetylcysteine administration and intracranial pressure monitoring. Moreover, they underwent daily 6-hour BAL treatments. Eight patients were treated. Three patients were bridged to liver transplantation, and five patients recovered without a transplant. All patients experienced neurological and metabolic improvement after treatments with the BAL support system. The BAL support system seems to improve the outcome of high-risk patients with acetaminophen-induced FHF, even in the absence of liver transplantation. Avoiding liver transplantation is particularly important in an era of organ shortage and high cost of transplants. PMID- 10515539 TI - Current management of recurrent pyogenic cholangitis. AB - Recurrent pyogenic cholangitis (RPC) is a chronic disease with multiple exacerbations requiring repeated biliary dilatation and stone removal. Even after adequate biliary drainage, most patients will have progression of intrahepatic disease. Management of patients with RPC is a multidisciplinary challenge for endoscopists, interventional radiologists, and surgeons because of the frequency and inaccessibility of strictures and stones. Complete stone clearance at any one operation is difficult. Hepaticojejunostomy with a subcutaneous afferent limb is a safe and effective way to provide access to the biliary tree for the management of patients with RPC. In our experience, trans-stomal cholangioscopic stricture dilatation followed by stone removal remains the basis of therapy in patients with RPC. By diligent surveillance, we should be able to eliminate or decrease the number of stones and prevent cholangitis and its sequelae. PMID- 10515540 TI - Iatrogenic gallbladder perforation during laparoscopic cholecystectomy: etiology and sequelae. AB - Iatrogenic perforation of the gallbladder (PGB) during laparoscopic cholecystectomy (LC) leads to spillage of bile and gallstones into the peritoneal cavity, which can result in serious postoperative infection. The objective of this study is to prospectively evaluate with long-term follow-up the risk factors, mechanisms, and complications associated with PGB in patients undergoing LC. Data from 1412 patients undergoing LC were collected prospectively between 1989 and 1995. Patients with and without iatrogenic gallbladder perforation were compared. Long-term follow-up was obtained using mailed questionnaires and telephone interviews, when needed. Of the 1412 patients, 512 (36%) sustained a PGB. Male sex, weight, gallbladder inflammation, thickening of the gallbladder, presence of adhesions, and a difficult hilar dissection were all associated with an increased incidence of PGB. The most common mechanisms of PGB were laceration due to grasper traction (55%) and electrocautery dissection (40%). Both the operating time and length of hospital stay were significantly longer in the PGB group. No difference was observed in the rate of wound infections between PGB and non-PGB patients (1.6% versus 1.8%). Only one patient (with an inflamed and perforated gallbladder) developed an early postoperative intra-abdominal abscess. Long-term follow-up averaging 48 months was achieved with a response rate of 44 per cent. No late intra-abdominal abscesses or complications attributable to retained gallstones were discovered. PMID- 10515541 TI - A mortality-free decade of pancreatoduodenectomy: is quality independent of quantity? AB - Pancreatoduodenectomy (PD) for periampullary cancer is a procedure of high morbidity and poor long-term survival. Superior clinical outcome has been described in high-volume institutions or for surgeons with a high case load. All patients undergoing pancreatectomy at the City of Hope National Medical Center (Duarte, CA) between 1987 and 1998 were analyzed retrospectively for postoperative outcome, and correlating or predictive clinicopathological factors were identified. Fifty-four patients underwent pancreatectomy [PD, n = 43; pylorus-preserving PD, n = 8; total pancreatectomy, n = 3]. There were 26 males and 28 females, with a median age of 63 years (range, 19-86). Fifty patients had a malignant diagnosis, and four patients had a benign diagnosis. Nine surgical oncologists performed an average of six pancreatectomies (range, 2-8). There was no perioperative death. Postoperative complications occurred in 30 patients, and infections predominated (n = 17). The median hospital stay was 16.5 days. The median postoperative actuarial survival by cancer site was 56 months (ampullary/ bile duct), 32.5 months (duodenal), 22.5 months (pancreatic), and 23.2 months (others). In this 11-year single institutional experience, PD and total pancreatectomy have been performed without lethal complication. In the setting of an exclusive oncology practice, operative mortality rates and survival outcome can be generated that compare favorably to large center experiences. Quality of outcome after pancreatectomy can be independent of quantity. PMID- 10515542 TI - Unresectable pancreatic carcinoma: correlating length of survival with choice of palliative bypass. AB - The preferred method of biliary bypass and the need for prophylactic gastroenterostomy in unresectable pancreatic carcinoma are dependent on the length of survival of the patient. From 1980 through 1996, 60 patients with biopsy-proven pancreatic cancer were found to be unresectable at exploration. The reasons for unresectability included major vascular involvement in 21 patients (35%), liver metastases in 16 (26.7%), celiac or portal lymph node metastases in 13 (21.7%), carcinomatosis in 5 (8.3%), and advanced age and/or comorbid medical condition in 4 patients (6.7%). One patient refused pancreaticoduodenectomy. Nine patients (15%) underwent Roux-en-Y choledochojejunostomy, and 51 (85%) underwent choledochoduodenostomy. Prophylactic gastroenterostomy was not performed routinely; however, in 9 patients (15%), gastrojejunostomy was performed for impending duodenal obstruction. Late biliary obstruction did not occur. Late gastric obstruction occurred in 6 of 51 patients (11.7%), at a median of 13.5 months after initial operation (range, 5-26 months). However, late gastric obstruction primarily occurred in 5 of 31 patients (16%) with locally advanced disease (major vessel involvement or lymph node metastases). The median survival was 12.0 months (range, 3.5-62 months) for patients with major vessel involvement, 11.5 months (range, 3-42 months) for patients with lymph node metastases, 4.5 months (range 0.5-24 months) for patients with liver metastases, 5.0 months (range, 4-7 months) for patients with carcinomatosis, and 9.0 months (range 2-27 months) for patients with significant comorbid medical illness and/or advanced age. Patients with liver metastases and carcinomatosis do not survive long enough to develop late obstruction. On the other hand, patients with locally advanced pancreatic carcinoma have a longer median survival and could be considered for prophylactic gastroenterostomy to avoid late gastric obstruction. Choledochoduodenostomy offers effective palliation for biliary obstruction. PMID- 10515543 TI - Computed tomography scanning for the diagnosis of perforated appendicitis. AB - The optimal initial treatment for perforated appendicitis may be nonoperative. For this reason it is important to be able to reliably distinguish between acute and perforated appendicitis. CT scanning has been shown to be highly accurate for the diagnosis of appendicitis, but it has not been specifically evaluated for perforated appendicitis. Our objective was to evaluate CT for the diagnosis of perforated appendicitis. Our study population comprised 84 patients who underwent appendectomy between 1993 and 1997 and who had CT scanning performed preoperatively. Medical records were reviewed retrospectively. CT scans were reviewed in a blinded fashion. CT findings were correlated with pathologic and clinical factors. Sixteen patients with acute appendicitis, 59 patients with gangrenous or perforated appendicitis, and 9 patients with normal appendices or other diagnoses were evaluated. For patients with pathologic documentation of appendicitis, CT findings that independently predict perforation or gangrene included abscess (P<0.001), phlegmon (P<0.001), extraluminal gas (P = 0.01), and terminal ileal wall thickening (P = 0.03). CT findings of an abscess, extraluminal gas, or phlegmon have a sensitivity of 92 per cent, specificity of 88 per cent, positive predictive value of 96 per cent, and negative predictive value of 74 per cent for perforated or gangrenous appendicitis. We conclude that CT can reliably distinguish between acute and perforated appendicitis. PMID- 10515544 TI - Perforated appendicitis is not a contraindication to laparoscopy. AB - Recent studies have reported an increased risk of intra-abdominal abscess formation following laparoscopic operation for perforated appendicitis. We undertook this study to compare laparoscopic versus open appendectomy in the treatment of perforated appendicitis. Records of all patients undergoing an appendectomy between January 1994 and June 1997 were reviewed, classifying appendicitis as acute, gangrenous, or perforated based on the intraoperative findings. Operative procedures were categorized as open, laparoscopic converted to open, or laparoscopic. The study group included 690 patients; four hundred fourteen (60%) were acute, 77 (11%) were gangrenous, and 199 (29%) were perforated. Although mean length of stay was shorter for all patients undergoing laparoscopic appendectomy, patients with perforated appendicitis had similar length of stay between treatment groups. Mean operative time for open appendectomy was significantly shorter than for converted or laparoscopic appendectomy regardless of diagnosis (P<0.01). Ten patients (1.4%) developed an intra-abdominal abscess: six after open appendectomy (1.7%), one after converted appendectomy (3.7%), and three after laparoscopic appendectomy (1%). There was no significant difference in rate of abscess formation in patients with perforated appendicitis undergoing open, converted, or laparoscopic appendectomy. We conclude that laparoscopic appendectomy for perforated appendicitis is not associated with an increased rate of intra-abdominal abscess formation. PMID- 10515546 TI - Complex repair for the management of duodenal injuries. AB - The management of duodenal injuries is a subject of ongoing debate. In this study we attempt to describe duodenum-related morbidity (DRM) after primary repair or complex repair (CR) and to identify risk factors for development of complications. The medical records of 145 consecutive patients admitted to Los Angeles County + University of Southern California Medical Center with duodenal injuries between January 1991 and December 1997 were reviewed. Fifty-four (37%) died within 24 hours of admission because of associated injuries. The remaining 91 were subjected to univariate and multivariate analysis. Of them, 66 (72.5%) developed complications and 3 (3%) died. CR was used in 32 (35%) patients and with increasing frequency as the grade of duodenal injury increased. DRM rate was overall low (9%) and not different between low-grade and high-grade duodenal injuries. This occurred despite a significant increase in Injury Severity Score and abdominal Abbreviated Injury Score in patients with more severe duodenal injuries. Patients with overall complications had higher Injury Severity Scores, higher abdominal Abbreviated Injury Scores, and more severe duodenal injuries. We conclude that duodenal injuries are frequently associated with other highly lethal injuries. Liberal use of CR in patients with more severe duodenal injuries prevents DRM. PMID- 10515545 TI - Abdominal computed tomography scan in pediatric blunt abdominal trauma. AB - The purpose of this study was to evaluate the role of abdominal CT scans in pediatric patients and correlate the findings with the clinical examination. A 2 year retrospective review of 88 patients with an abdominal CT scan after blunt trauma was performed. Seventy-two patients were identified with complete clinical examination data available. In its ability to predict the need for surgery, the CT scan had a sensitivity of 67 per cent and a negative predictive value of 98.7 per cent. The combination of the clinical examination and the CT scan findings did not miss any significant injuries. No patient with a soft, nontender abdomen and a negative CT scan required an abdominal operation. We conclude that the CT scan alone may miss clinically significant injuries. In blunt abdominal trauma in the pediatric population, the CT scan findings should be coupled with the clinical examination to ensure that no significant abdominal injuries are missed. PMID- 10515547 TI - Determinants of survival after inferior vena cava trauma. AB - Inferior vena cava (IVC) injuries continue to be associated with mortality rates of 21 to 66 per cent despite advances in prehospital, surgical, and critical care. The purpose of this study was to evaluate outcome of patients with IVC injury after treatment at a major urban trauma center and to identify factors predictive of survival. Between 1989 and 1995, 158 patients presented to the Los Angeles County + University of Southern California Medical Center with IVC injuries. One hundred thirty-six patient records were available for review, and 69 data points were collected and analyzed. Mean age was 26 years (range, 6-54), and 122 (90%) patients were male. Mechanism of injury included gunshot in 88 (65%) patients, stab in 23 (17%) patients, shotgun in 7 (5%) patients, and blunt trauma in 18 (13%) patients. The mean Injury Severity Score was 25. Seventy (52%) patients were hypotensive. Eleven (8%) patients died before surgical intervention, and 25 (18%) patients died before operative repair. Repair (79), ligation (20), or observation (1) was accomplished in 100 (74%) patients. Overall survival was 48 per cent and 65 per cent in the 100 patients surviving to operative repair, including 5 of 20 patients requiring IVC ligation. Significant differences (P<0.001) between survivors and nonsurvivors included Injury Severity Score, Glasgow Coma Score, hematocrit, hypotension, emergent thoracotomy, blood loss, level of injury, tamponade, and associated aortic injury. Logistic regression analysis identified hypotension, anatomic level of injury, and associated aortic injury as significant predictors of outcome (P = 0.001). Survival is predominantly determined by severity and anatomic accessibility of the IVC injury and by the absence of associated major vascular injuries. Ligation may control otherwise exsanguinating injuries and should be considered early in the management of complex injuries. PMID- 10515548 TI - Percutaneous dilatational tracheostomy: still a surgical procedure. AB - Although percutaneous dilatational tracheostomy (PDT) has been shown to be a cost effective bedside alternative to open tracheostomy (OT), prior reports of the complications of the procedure are contradictory. Reported complications range from minor events to fatal ones, in varying percentages. This prospective study was designed to identify the type and severity of complications accompanying the introduction of PDT to a tertiary medical center. Surgical and medical intensive care unit (ICU) patients requiring elective tracheostomy were identified as appropriate for PDT using approved institutional criteria. All procedures were performed at an ICU bedside in the presence of a surgeon privileged to perform OT. Demographic data, procedural information, and patient outcome (including minor and major complications, length of stay, and survival) were collected. PDT was performed in 96 ICU patients, with complete data available for 95 patients. PDT was performed in an average of 13.1+/-1.0 minutes. Twenty-three major and minor complications occurred, including two perioperative deaths, in 15 patients (15.8%). A total of 37 PDT patients (38.9%) died in the hospital, indicative of the severity of illness of patients requiring tracheostomy. Based on the experience to date, Cedars-Sinai Medical Center (Los Angeles, CA) continues to require a surgeon privileged to perform OT to participate in all PDT procedures. PMID- 10515549 TI - Nosocomial infections in the surgical intensive care unit: a difference between trauma and surgical patients. AB - In 1970, the Centers for Disease Control and Prevention (CDC) established the National Nosocomial Infection Surveillance System to assist institutions with infection surveillance, data collection, and processing. This facilitates interinstitutional comparison for nosocomial infection rates. Nosocomial infection rates in the surgical intensive care unit have been shown to be different from the medical intensive care unit. Whether there exists a difference in infection rates between trauma and surgical patients in the intensive care unit has not been established. Our objective was to determine whether there is a difference in rates of nosocomial infections between trauma and surgical patients in the surgical intensive care unit. From January 1995 through December 1997, we reviewed 3715 admissions to the surgical intensive care unit and separated them into trauma (1272) or surgical (2443) cases. We documented all nosocomial pneumonias, urinary tract infections, bloodstream infections, and surgical site infections. From these data we determined infection rates per 100 admissions. We also identified all device-related nosocomial infections and calculated infection rate by current CDC standards using number of device infections divided by number of device-days times 1000. We found that the overall trauma patient infection rate was 11.64 per cent compared with 6.43 per cent for surgical patients (P<.001). Using conventional infection rate criteria, trauma patients had higher frequency in the rate of ventilator-associated pneumonia (6.13% vs. 2.50%; P<0.001), urinary tract infection (2.36 versus 1.76; P<0.2), and bloodstream infection (2.52% versus 1.27%; P<0.01). However, when using the CDC guidelines, which correct for the number of device-days for infections, only the difference in rate of pneumonia between the two groups reached statistical significance (23.9 rate for trauma patients vs. 16.7 for the surgery group; P<0.005). We conclude that trauma patients are at higher risk for nosocomial infections than routine surgical patients. Because of this difference, centers should collect and report data separately for trauma and surgical patients in the intensive care unit. Specific attention should be focused on the causes and prevention of increased rates of nosocomial pneumonia in trauma patients. PMID- 10515550 TI - Axillary dissection after unsuccessful sentinel lymphadenectomy for breast cancer. AB - Intraoperative lymphatic mapping and sentinel lymphadenectomy (LM/SL) has been demonstrated to provide sensitive axillary staging for breast cancer. LM/SL has a steep learning curve, and factors associated with unsuccessful LM/SL are not well known. Two hundred sixty patients with breast carcinoma and clinically negative axillae underwent injection of about 5 cm3 of isosulfan blue dye (Lymphazurin, US Surgical Corp, Norwalk, CT) into breast tissue surrounding a cancer or biopsy site. After 5 minutes of breast compression, blue-stained lymph nodes were sought. In 47 patients, no blue nodes were detected; a standard axillary dissection was performed. All 47 patients were women with a mean age of 56 years (range, 34-80). Ductal carcinoma was most common (91.5%). Mean tumor size was 1.99 cm. Axillary dissection yielded a mean of 15.8 lymph nodes (range, 6-35). Sixteen patients (34%) had positive lymph nodes (mean, 7.6; median, 6; range, 1 24). Factors associated with LM/SL difficulty include surgeon inexperience, medial hemisphere primary location, extensive axillary metastases, and extranodal invasion. Inability to identify a sentinel node in a clinically negative axilla is a risk factor for extensive axillary tumor burden. Axillary dissection should be performed for patients with unsuccessful LM/SL, particularly those with lateral hemisphere primaries. PMID- 10515551 TI - Objective assessment of axillary morbidity in breast cancer treatment. AB - Historically, axillary lymph node dissection (ALND) was a critical aspect of the operative management of breast cancer. Recently, the role of ALND has been questioned, with postoperative morbidity possibly overshadowing patient benefit. Our objective was to quantitatively assess the long-term morbidity of ALND in patients with breast cancer. We conducted a cross-sectional study of patients being followed by the Breast Surgery Clinic at a university-affiliated urban hospital. Ninety-five patients with unilateral breast cancer who had undergone ALND were evaluated at routine follow-up visits in the latter half of 1998. A questionnaire was used to quantify the degree of subjective findings, including arm swelling, chest wall pain, decreased mobility, and weakness. Upper extremity strength, active range of motion, and circumference were measured. Overall, 70 per cent of patients had at least one complaint, with 18 per cent having moderate to severe symptoms. Twenty-one per cent had notable decrements in strength or range of motion, 9.3 per cent of patients required chronic compression garments for lymphedema, and 6.4 per cent changed their vocational status because of surgical morbidity. We conclude that adverse effects from ALND occur commonly. Objective findings are less common, perhaps causing clinicians to underappreciate postoperative morbidity. A significant subset of patients had enduring disability. PMID- 10515552 TI - Effects of electrophysiologic-guided therapy with Class IA antiarrhythmic drugs on the long-term outcome of patients with idiopathic ventricular fibrillation with or without the Brugada syndrome. AB - INTRODUCTION: Implantation of a implantable cardioverter defibrillator (ICD) is viewed universally as the "gold standard" therapy for patients with idiopathic ventricular fibrillation (VF). We sought to study the long-term value of electrophysiologic (EP)-guided therapy with Class IA antiarrhythmic drugs in patients with idiopathic VF with or without the Brugada syndrome. METHODS AND RESULTS: We performed EP studies in 34 consecutive patients who had idiopathic VF with (n = 5) or without (n = 29) the Brugada syndrome. All patients with inducible sustained polymorphic ventricular tachycardia (SPVT) or VF underwent repeated EP evaluation after oral administration of a Class IA antiarrhythmic drug (mainly quinidine). Patients rendered noninducible received this therapy on a long-term basis. SPVT/VF were induced in 27 (79.4%) patients at baseline studies. Class IA drugs effectively prevented induction of SPVT/VF in 26 (96%) patients. Of the 23 patients treated with these medications, no patient died or had a sustained ventricular arrhythmia during a mean follow-up period of 9.1 +/- 5.6 years (7 to 20 years in 15 patients). Two deaths occurred in patients without inducible SPVT/VF at baseline studies who had been treated empirically. CONCLUSION: Our results suggest that EP-guided therapy with Class IA agents is a reasonable, safe, and effective approach for the long-term management of patients with idiopathic VF. A randomized prospective study of EP-guided Class IA therapy in patients with ICDs seems warranted. PMID- 10515553 TI - Idiopathic ventricular fibrillation: is there a role for electrophysiologic guided antiarrhythmic drug therapy? PMID- 10515554 TI - Ischemia-induced changes of the signal-averaged electrocardiogram: experimental investigation during percutaneous transluminal coronary angioplasty balloon occluded coronary artery. AB - INTRODUCTION: The influence of myocardial ischemia on the detection of an arrhythmogenic substrate with the signal-averaged ECG is unclear. METHODS AND RESULTS: In 80 patients with single vessel coronary artery disease and a critical stenosis of the left anterior descending vessel selected after coronary angiography in whom percutaneous transluminal coronary angioplasty (PTCA) was planned, we retrospectively investigated the signal-averaged ECGs in the time domain before, during, and after occlusion of the coronary artery by the PTCA balloon. Forty patients were resuscitated from ventricular fibrillation (VF group), and 40 patients had no ventricular arrhythmia (non-VF group). Late potentials were seen at rest in 26 of 40 patients in the VF group. During ischemia, the duration of the filtered QRS complex and the duration of low amplitude signals < 40 microV increased significantly. In another 14 patients in the VF group, late potentials were observed only during ischemia. In 4 of 26 patients in the VF group without prior infarction but with severe ischemia present at rest, successful PTCA eliminated preexistent late potentials. In the non-VF group, one patient had late potentials present at rest. In two patients with prior infarction, late potentials were provokable only during transmural ischemia. CONCLUSION: Myocardial ischemia is able to modify detection of an arrhythmogenic substrate with the signal-averaged ECG. PMID- 10515555 TI - Ischemia-induced ventricular tachyarrhythmias: should we focus on conduction delay, dispersion of repolarization, or both? PMID- 10515556 TI - Dual morphology of idiopathic ventricular tachycardia. AB - INTRODUCTION: Idiopathic ventricular tachycardia (VT) typically has a single morphology originating either in the right ventricular outflow tract (RVOT) or near the posterior fascicle of the left ventricle (LV) in most instances. We present our observations in six patients with idiopathic VT in whom two morphologies were present. METHODS AND RESULTS: Of 55 patients with idiopathic VT who underwent radiofrequency (RF) ablation, 44 had LV "fascicular" tachycardia, whereas 11 had RVOT tachycardia. During RF energy delivery, there was a change in VT morphology in two patients with idiopathic LV tachycardia. This second morphology was not ablated initially, recurred at follow-up, and was reablated successfully. In two additional patients with idiopathic LV tachycardia, a second VT was inducible after ablation of the "clinical" VT. This second morphology recurred at follow-up and was ablated successfully in one patient. The site where the second VT was ablated in all the three patients was remote from that of the first VT. In two patients with RVOT tachycardia, a second VT, originating from a different area of the RVOT, was induced after RF ablation of the "clinical" VT. This second VT recurred at follow-up and was reablated successfully in one patient. CONCLUSION: Idiopathic VT is a more heterogenous entity than hitherto believed. A second VT was seen in 11% of patients during or after RF ablation of the "clinical" VT. The appearance of a second VT suggests either a different exit site of the same circuit or another site of origin. PMID- 10515557 TI - Amiodarone reduces the prevalence of T wave alternans in a population with ventricular tachyarrhythmias. AB - INTRODUCTION: Testing for the presence of microvolt T wave alternans (TWA) is useful for arrhythmic risk stratification. Whether antiarrhythmic pharmacotherapy affects the presence of TWA is unknown. We tested whether patients with known ventricular tachyarrhythmias who were receiving amiodarone were less likely to manifest TWA as compared with those not receiving amiodarone. METHODS AND RESULTS: Forty-four patients with a history of ventricular tachyarrhythmias and an implantable cardioverter defibrillator (ICD) implanted at least 1 month earlier underwent TWA testing. In this group, 14 patients were receiving amiodarone and 30 were not. Indeterminate test results occurred in 13 patients without a significant difference in those receiving or not receiving amiodarone. In the 31 patients with determinate TWA testing, a positive test was less likely in those receiving amiodarone (1 of 9 [11%]) as compared with those not receiving amiodarone (14 of 22 [64%]; P = 0.04). During a follow-up period averaging 0.9 +/ 0.2 years, the presence of TWA (P = 0.04) and decreased left ventricular ejection fraction (P = 0.05) predicted appropriate ICD therapy for ventricular tachyarrhythmias. CONCLUSION: The prevalence of TWA was decreased in a chronic ventricular tachyarrhythmic population receiving amiodarone as compared with a population not receiving amiodarone. PMID- 10515558 TI - Accuracy and limitations of published algorithms using the twelve-lead electrocardiogram to localize overt atrioventricular accessory pathways. AB - INTRODUCTION: The purpose of this study was to evaluate the accuracy and limitations of published algorithms using the 12-lead ECG to localize AV accessory pathways (APs). METHODS AND RESULTS: The 11 relevant algorithms found in the literature (MEDLINE database and major scientific sessions) were tested on a series of 266 consecutive patients who successfully underwent radiofrequency catheter ablation of a single overt AV AP. The positive predictive values (PPV) of the algorithms in applicable patients were significantly lower for algorithms with > 6 accessory location sites (40.6% +/- 10.9% vs 61.2% +/- 8.0%; P < 0.03) and show a tendency for algorithms not relying on delta wave polarity but on QRS polarity only (36.6% +/- 11.2% vs 52.3% +/- 13.1%; P = 0.09). The PPV in applicable patients is related to the AP location (P < 0.001) and ranked from the highest to the lowest as follows: left lateral (mean PPV = 86.3%), posteroseptal (mean PPV = 65.2%), right anteroseptal (mean PPV = 45.2%), and right posterolateral (mean PPV = 23.4%). CONCLUSION: Our study suggests that the accuracy of algorithms relying on the 12-lead ECG depends on AP locations as defined in the algorithms and on the number of AP sites. The accuracy tends to be lower when delta wave polarity is not included in the algorithm's architecture. This should be considered when using these algorithms or when building new ones. PMID- 10515559 TI - No evidence of chaos in the heart rate variability of normal and cardiac transplant human subjects. AB - INTRODUCTION: The variability observed in the heart rate may reflect fundamental aspects of cardiac activity. It has been under discussion whether heart rate variability (HRV) is due to noise or chaos, which is irregular behavior occurring in deterministic nonlinear systems. METHODS AND RESULTS: Using chaos analysis techniques, we analyzed HRV of five normal and five human cardiac transplant subjects at rest. HRV is studied using the beat-to-beat RR interval time series extracted from the ECG. The cardiac transplant subjects exhibited a much smaller HRV than the normal subjects because of heart denervation. We present the map and correlation dimension estimation for the RR time series. To test for nonlinear correlations in the dynamics, we built surrogate time series that have the same power spectra as the experimental time series, but also have randomized phases. The experimental and the surrogate data were compared using the correlation integral. No correlation dimension was found for the RR time series of either the normal or the cardiac transplant subjects. Nevertheless, nonlinear correlations were detected in the HRV of the normal subjects but not in HRV of the cardiac transplant subjects. For the latter, no significant changes were observed in the correlation integral as a function of time after transplantation. CONCLUSION: We found no evidence of low-dimensional chaos in the HRV of normal and cardiac transplant subjects. However, some nonlinear correlations were detected in the HRV of the normal subjects, which may be associated with autonomic nervous system influence. PMID- 10515560 TI - Chaos and heart rate variability. PMID- 10515561 TI - Characterization of conduction in the ventricles of normal and heterozygous Cx43 knockout mice using optical mapping. AB - INTRODUCTION: Gap junction channels are important determinants of conduction in the heart and may play a central role in the development of lethal cardiac arrhythmias. The recent development of a Cx43-deficient mouse has raised fundamental questions about the role of specific connexin isoforms in intercellular communication in the heart. Although a homozygous null mutation of the Cx43 gene (Cx43-/-) is lethal, the heterozygous (Cx43+/-) animals survive to adulthood. Reports on the cardiac electrophysiologic phenotype of the Cx43+/- mice are contradictory. Thus, the effects of a null mutation of a single Cx43 allele require reevaluation. METHODS AND RESULTS: High-resolution video mapping techniques were used to study propagation in hearts from Cx43+/- and littermate control (Cx43+/+) mice. Local conduction velocities (CVs) and conduction patterns were quantitatively measured by determining conduction vectors. We undertook the characterization of ECG parameters and epicardial CVs of normal and Cx43+/- mouse hearts. ECG measurements obtained from 12 Cx43+/+ and 6 Cx43+/- age matched mice did not show differences in any parameter, including QRS duration (14.5 +/- 0.9 and 15.7 +/- 2.3 msec for Cx43+/+ and Cx43+/-, respectively). In addition, using a sensitive method of detecting changes in local CV, video images of epicardial wave propagation revealed similar activation patterns and velocities in both groups of mice. CONCLUSION: A sensitive method that accurately measures local CVs throughout the ventricles revealed no changes in Cx43+/- mice, which is consistent with the demonstration that ECG parameter values in the heterozygous mice are the same as those in wild-type mice. PMID- 10515562 TI - A gap in understanding the connection between connexins and cardiac conduction. PMID- 10515563 TI - Cardiac conduction abnormalities in mice lacking the gap junction protein connexin40. AB - INTRODUCTION: The gap junction protein connexin40 (Cx40) normally is expressed in the murine atrial myocardium and ventricular conduction system. In mice lacking Cx40, several changes in the surface ECG have been described. In this study, we analyzed cardiac conduction in more detail. METHODS AND RESULTS: In open chest mice under urethane anesthesia, epicardial electrodes were used to determine a number of atrial and ventricular pacing parameters. The corrected sinus node recovery time was significantly longer in Cx40-/- mice than in Cx40+/+ mice (44.4 +/- 7.2 msec vs 35.5 +/- 8.0 msec). In addition, the Wenckebach period was longer in Cx40-/- mice compared with the wild type (84.6 +/- 5.4 msec vs 78.8 +/- 3.6 msec), with the AV node probably limiting AV conduction in both cases. Whereas arrhythmias could not be induced by ventricular burst pacing in any of the mice, atrial burst pacing induced atrial tachyarrhythmias in 5 of 10 Cx40-/- mice, but not in any of 9 Cx40+/+ mice. Conduction velocities were measured in vivo using an array of unipolar recording electrodes. Ventricular conduction velocity did not differ between the groups, but atrial conduction velocity was reduced by 30% in Cx40-/- mice compared with the wild type. Heterozygous Cx40+/- mice did not differ significantly from the wild type in any respect. CONCLUSION: These findings indicate that in the atria and the AV conduction system, Cx40 is an important determinant of conduction. PMID- 10515565 TI - Inspiration induced by phrenic nerve stimulation increases internal defibrillation energy requirements. AB - INTRODUCTION: The purpose of this study was to systematically evaluate the effects of active inspiration induced by phrenic nerve stimulation on the energy required for 50% successful defibrillation (E50). METHODS AND RESULTS: Shocks (95 microF biphasic waveform) were delivered after 10 seconds of ventricular fibrillation between a right ventricular coil and left pectoral test can in ten anesthetized pigs (25 to 37 kg). Using a 1-J step size, the E50 was determined with an up/down, three-reversal method. Positive-pressure ventilation was halted just before fibrillation, and shocks were delivered during expiration or at the end of 2 seconds of bilateral phrenic stimulation (50 Hz, 0.3 msec, 5 to 6 V). Phrenic stimulation produced inspiratory volumes that were 15.3 +/- 1.7 mL/kg (mean +/- SD). The E50 was 9.8 +/- 1.9 J during expiration and increased to 13.0 +/- 1.7 during inspiration (P = 0.001). The leading-edge voltage at the E50 was 451 +/- 46 V during expiration and 519 +/- 33 V during inspiration (P = 0.001). The leading-edge current at the E50 was 9.7 +/- 1.0 A during expiration and increased to 11.3 +/- 1.4 A during inspiration (P = 0.002). The average impedance was 47.8 +/- 2.7 omega during expiration and 47.3 +/- 3.3 omega during inspiration (P = 0.12). CONCLUSION: Inspiration induced by phrenic stimulation results in a 31% increase in the E50 compared with expiration. The decrease in shock efficacy occurs in the absence of a change in impedance. Active inspiration may alter the distribution of the electrical field leading to a decrease in shock efficacy. PMID- 10515564 TI - Voltage-gated Na+ channel activity and connexin expression in Cx43-deficient cardiac myocytes. AB - INTRODUCTION: Dynamic interplay between active and passive electrical properties of cardiac myocytes is based on interrelationships between various channels responsible for depolarizing and repolarizing ionic currents and intercellular conductances. Mice with targeted disruption of the connexin43 (Cx43) gene have hearts completely devoid of Cx43, the principal gap junctional protein expressed in mammalian hearts. METHODS AND RESULTS: To determine whether cardiac myocytes that develop in an abnormal environment of reduced intercellular coupling have altered active membrane properties, we studied whole cell action potentials, Na+ channel currents, and Na+ channel expression and distribution via immunoblotting and confocal immunofluorescence in neonatal ventricular myocytes isolated from Cx43 wild-type, heterozygous, and homozygous null hearts. Action potential morphology, peak Na+ current, activation and inactivation kinetics, and Na+ channel protein expression and distribution were not different among myocytes isolated from wild-type, heterozygous, or null hearts. Active membrane properties and Na+ channel activity were completely normal in Cx43-deficient myocytes isolated from hearts that have been shown to exhibit markedly reduced Cx43 expression, gap junction number, and epicardial conduction delay. CONCLUSION: Despite a genetic inability to produce Cx43 and a developmental history that culminates in marked gross cardiac morphologic abnormalities, premature death, and myocardial inexcitability ex vivo, cardiac Na+ channel distribution and function appear to be normal in Cx43 null hearts. Although intimate structural and functional interrelationships have been described between ion channels and gap junction channels, expression and function of Na+ channels is not affected by the absence of Cx43. PMID- 10515566 TI - Cell coupling and impulse propagation in the failing heart. AB - Cell coupling and impulse propagation were investigated in the ventricle of cardiomyopathic hamsters at an advanced stage of heart failure. An appreciable decline in junctional conductance was found, a phenomenon in part related to activation of the plasma and cardiac renin-angiotensin systems. Decreased expression of connexin43 or an alteration of junctional proteins also might be implicated in the decreased cell coupling. Morphologic abnormalities such as fibrosis, necrosis, and rupture of cell contacts contribute to the decline of conduction velocity or to the blockade of impulse propagation in some areas of the ventricle, creating the conditions for anisotropic conduction and cardiac arrhythmias. The decrease in membrane potential found in myopathic cells is related in part to depression of Na-KATPase activity, and the lack of action of beta-adrenergic agonists on junctional conductance is explained by down regulation of beta receptors and an abnormality of adenyl cyclase. PMID- 10515567 TI - Modulation of gap junction properties in failing hearts. PMID- 10515568 TI - Heart failure: the electrophysiologic connection. AB - Heart failure often is complicated by lethal arrhythmias. This article presents an overview of current thinking about the mechanisms of sudden death in heart failure, with an emphasis on the hypothesis that cardiomyopathy is an acquired form of the long QT syndrome. PMID- 10515569 TI - A narrow QRS complex tachycardia: what is the mechanism of tachycardia? PMID- 10515570 TI - Mechanism of induction of atrial flutter. PMID- 10515571 TI - Elevated ovarian and thymic cell apoptosis in wild cotton rats inhabiting petrochemical-contaminated terrestrial ecosystems. AB - The objective of this study was to determine the rates of apoptotic cell death in ovary and thymus collected from wild female cotton rats (Sigmodon hispidus) inhabiting five petrochemical-contaminated and five ecologically matched reference sites in Oklahoma. Overall comparison of reference and contaminated sites, using individual sites as replicates, revealed a significantly increased rate of ovarian cell apoptosis in cotton rats inhabiting contaminated sites. In comparison to rats from reference sites, the number of uterine scars was lower in rats collected from the contaminated sites. There were no significant differences in the percentage of atretic follicles among animals collected from reference and contaminated sites. The rate of thymocyte apoptosis was elevated at one of five contaminated sites, although the overall rate of thymocyte apoptosis was not significantly different when comparing all sites. To our knowledge, this is the first study documenting elevated rates of ovarian and thymic cell apoptosis in wild mammals exposed chronically to environmental toxicants. PMID- 10515572 TI - A study of the effect of chrysotile fiber surface composition on genotoxicity in vitro. AB - Chrysotile fibers (NIEHS intermediate length) were treated with ultrapure HCl to alter the fiber surface chemistry without substantially changing fiber morphology or dimensions. The objective of the study was to determine whether fiber surface chemistry is an important variable in fiber genotoxicity in vitro. The modified fibers, along with native chrysotile fibers, were used to challenge Chinese hamster lung fibroblasts (V79) in vitro using the micronucleus induction genotoxicity assay. Fiber dimensions were assessed using scanning electron microscopy by measuring the distribution of fiber lengths in 3 length ranges: less than 3 microm, 3-10 microm, and greater than 10 microm. For both treated and native fiber samples, 500 fibers were examined. Results indicate that acid treated fibers were about 20% shorter than untreated chrysotile. Surface chemistry alterations were verified by zeta-potential reversal, x-ray photoelectron spectroscopy (XPS), and scanning electron microscopy/energy dispersive x-ray spectroscopy (SEM-EDS) elemental analysis. Scanning Auger spectrometry indicated the presence of Mg, O, and Si in both treated and native chrysotile samples, which confirmed the surface purity of both fiber samples. Both XPS and SEM-EDS analysis demonstrated substantial depletion of Mg from fiber surfaces. Results of the micronucleus assay showed a positive concentration related response for both samples, with toxicity evident only at the highest concentration. No significant difference was found for the treated and untreated chrysotile samples. These results indicate that the surface chemistry is not an important variable in the in vitro genotoxicity of chrysotile asbestos in V79 cells as detected by the micronucleus assay under the conditions used in this study, and support a model of chemically nonspecific chromosomal and spindle damage effects. PMID- 10515573 TI - Lung injury from intratracheal and inhalation exposures to residual oil fly ash in a rat model of monocrotaline-induced pulmonary hypertension. AB - A rat model of monocrotaline (MCT)-induced pulmonary injury/hypertension has been recently used in particulate matter (PM) health effects studies, however, results have been equivocal. Neither the mechanism by which mortality occurs in this model nor the variation in response due to differences in PM exposure protocols (i.e., a bolus dose delivered intratracheally versus a similar cumulative dose inhaled over three days) have been fully investigated. Sprague Dawley rats (SD, 60 d old; 250-300 g) were injected with either saline (healthy) or MCT, 60 mg/kg, i.p. (to induce pulmonary injury/hypertension). Ten days later they were exposed to residual oil fly ash (ROFA), either intratracheally (IT; saline, 0.83 or 3.33 mg/kg) or by nose-only inhalation (15 mg/m3 x 6 h/d x 3 d). Lung histology, pulmonary cytokine gene expression (0 and 18 h postinhalation), and bronchoalveolar lavage fluid (BALF) markers of injury were analyzed (24 and 96 h post-IT; or 18 h post-inhalation). Data comparisons examined three primary aspects, 1) ROFA IT versus inhalation effects in healthy rats; 2) pulmonary injury caused by MCT; and 3) exacerbation of ROFA effects in MCT rats. In the first aspect, pulmonary histological lesions following ROFA inhalation in healthy rats were characterized by edema, inflammatory cell infiltration, and thickening of alveolar walls. Increases in BALF markers of lung injury and inflammation were apparent in ROFA-IT or nose-only exposed healthy rats. Increased IL-6, and MIP-2 expression were also apparent in healthy rats following ROFA inhalation. In regards to the second aspect, MCT rats exposed to saline or air showed perivascular inflammatory cell infiltrates, increased presence of large macrophages, and alveolar thickening. Consistently, BALF protein, and inflammatory markers (macrophage and neutrophil counts) were elevated indicating pulmonary injury. In regards to the third aspect, 58% of MCT rats exposed to ROFA IT died within 96 h regardless of the dose. No mortality was observed using the inhalation protocol. ROFA inhalation in MCT rats caused exacerbation of lung lesions such as increased edema, alveolar wall thickening, and inflammatory cell infiltration. This exacerbation was also evident in terms of additive or more than additive increases in BALF neutrophils, macrophages and eosinophils. IL-6 but not MIP-2 expression was more than additive in MCT rats, and persisted over 18 h following ROFA. IL-10 and cellular fibronectin expression was only increased in MCT rats exposed to ROFA. In summary, only the bolus IT ROFA caused mortality in the rat model of lung injury/hypertension. Exacerbation of histological lesions and cytokine mRNA expression were most reflective of increased ROFA susceptibility in this model. PMID- 10515574 TI - Utilization of electrochemically generated ozone in the degradation and detoxication of benzo[a]pyrene. AB - The ability of electrochemically generated ozone (O3) to degrade and detoxify the polycyclic aromatic hydrocarbon (PAH) benzo[a]pyrene (BaP) was assessed utilizing the chick embryotoxicity screening test (CHEST) and Hydra attenuata bioassays. Aqueous solutions containing 10 microg/ml BaP and 0.5% (v/v) acetonitrile were subjected to ozonolysis for 0 to 30 min. Rapid degradation of BaP was evident by both gas chromatography/mass spectroscopy (GC/MS) and high-performance liquid chromatography (HPLC) analysis. HPLC fluorescence detection revealed no BaP shortly after 5 min of ozonolysis, while HPLC with PDA detection demonstrated continued reactions with ozone over the 30-min time course. As little as 2 min of O3 treatment afforded protection from BaP-induced mortality and toxicity (embryolethality and liver discoloration) in the chicken embryos. In the hydra bioassay, no toxicity was observed in the adult hydra until the ozonolysis products were reconstituted 100-fold from their initial post-ozonolysis concentrations. The results obtained from this study clearly demonstrate the potential application of electrochemically generated O3 for the detoxication and prevention of toxicity of BaP. Both CHEST and hydra assays predict that the ozonolysis products of BaP are less toxic than the parent compound. PMID- 10515575 TI - High-temperature effects on antioxidant systems and toxic product formation in nutritional oils. AB - In this study, effects of high-temperature heating on antioxidant defense potential (AOP) and malondialdehyde (MDA) levels were investigated in several types of oils ingested by humans. Natural olive oil, refined olive oil, sunflower oil, and soy oil were examined. High-temperature heating to 180 degrees C significantly decreased AOP. This was accompanied by a significant increase in MDA levels. The observed changes were quantitatively greater in soy and sunflower oil compared to olive oil. The loss in antioxidant defense potential and elevation in peroxidation products may be associated with enhanced disease processes. PMID- 10515577 TI - Glutathione status in retinopathy of prematurity. AB - This study examines the glutathione status of red blood cells in patients with retinopathy of prematurity (ROP) both in vivo and after an in vitro oxidative challenge. Fifty ROP patients of different ages (between 6 weeks and 6 years), born prematurely (gestational age: 28.7 +/- 1.3 weeks; birth weight: 1210 +/- 313 g; mean +/- SD) suffering either from active ROP (<3 months old; n = 12) or from a visual handicap due to preceding ROP (3 months-6 years; n = 38) as well as control patients of similar age and maturity (n = 56) were included. Infants with active disease have the lowest levels of reduced glutathione (GSH), the highest levels of oxidized form (GSSG), the highest GSSG/GSH ratios and the greatest fall in GSH after an in vitro oxidative challenge. After an in vitro oxidative stress, defective glutathione recycling was found in patients with preceding ROP and was suggested as a factor predisposing to oxidative hemolysis. The glutathione redox ratio was warranted as a biochemical screen for active ROP in premature infants. PMID- 10515576 TI - Age-dependent increase of collagenase expression can be reduced by alpha tocopherol via protein kinase C inhibition. AB - Total protein kinase C (PKC) activity in human skin fibroblasts increases during in vivo aging as a function of the donor's age. During in vitro aging protein kinase C activity is also increased, as a function of cell passage number. Using PKC isoform specific antibodies, we demonstrate that the increase in total PKC activity is mainly due to the PKC a isoform. PKC alpha protein expression increased up to 8 fold during in vivo aging. Collagenase (MMP-1) gene transcription and protein expression also increased with age, concomitant with the increase in protein kinase C alpha. Furthermore, alpha-tocopherol, which inhibits protein kinase C activity, is able to diminish collagenase gene transcription without altering the level of its natural inhibitor, tissue inhibitor of metalloproteinase, TIMP-1. We propose that an aging program leads to increased protein kinase C alpha expression and activity. This event would induce collagenase overexpression followed by increased collagen degradation. Our in vitro experiments with skin fibroblasts suggest that alpha-tocopherol may protect against skin aging by decreasing the level of collagenase expression, which is induced by environmental insults and by aging. PMID- 10515578 TI - Diphenylacetaldehyde-generated excited states promote damage to isolated rat liver mitochondrial DNA, phospholipids, and proteins. AB - This work studies damage to rat liver mitochondrial protein, lipid, and DNA caused by electronically excited states generated by cytochrome c-catalyzed diphenylacetaldehyde enol oxidation to triplet benzophenone. The extension of lipid peroxidation was estimated by production of thiobarbituric acid-reactive substances and by formation of Schiff bases with membrane proteins, evaluated by SDS-polyacrylamide gel electrophoresis. Concomitant with DPAA-driven mitochondrial permeabilization, extensive mtDNA fragmentation occurred and DNA adducts with aldehydes-products of fatty acid oxidation-were observed. The degree of lipid peroxidation and mtDNA alterations were significantly decreased by butylated hydroxytoluene, a potent peroxidation chain breaker. The lipid peroxidation process was also partially inhibited by the bioflavonoid rutin and urate totally prevented the mitochondrial transmembrane potential collapse. In all cases, the mitochondrial damage was dependent on the presence of phosphate ions, a putative bifunctional catalyst of carbonyl enolization. These data are consistent with the notion that triplet ketones may act like alkoxyl radicals as deleterious reactive oxygen species on biologic structures. Involvement of singlet dioxygen formed by triplet-triplet energy transfer from benzophenone in the model reaction with DPAA/cytochrome c in the presence of DCP liposomes was suggested by quenching of the accompanying chemiluminescence upon addition of histidine and lycopene. PMID- 10515579 TI - Generation of reactive oxygen intermediates, activation of NF-kappaB, and induction of apoptosis in human endothelial cells by glucose: role of nitric oxide synthase? AB - Exposure to high glucose causes characteristic dysfunction and morphologic changes of the endothelium. To study the underlying mechanisms of glucotoxicity, human endothelial cells (HUVECs) were isolated from umbilical veins and cultivated under hyperglycemic conditions (10-30 mM) for up to 72 h. The generation of reactive oxygen intermediates (ROIs) was determined by histochemical staining of the cells by dichlorodihydrofluorescein. Activation of the transcription factor nuclear factor-kappa B (NF-KB) family was analyzed by the electromobility shift assay and by histochemical staining of the cells with rhodamine-labelled consensus sequences of activated NF-KB. Apoptotic cells were identified by morphologic analysis and DNA fragmentation. Incubation of HUVECs with high glucose led to rapid increase in the generation of ROIs. After an incubation of 2 to 6 h, NF-KB became activated, with the maximum at 4 h. Exposure of HUVECs to high glucose for up to 72 h caused a significant induction of apoptosis in HUVECs. The increased generation of ROIs, activation of apoptosis, and induction of apoptosis were also observed in cells incubated with 3-O-methyl D-glucose, a glucose derivative that is taken up by the cells but not metabolized. Generation of ROIs, activation of NF-kappaB, and induction of apoptosis were not only prevented by antioxidants (thioctic acid, tocopherol, superoxide dysmutase-mimetic), but also by L-nitroarginine. These observations indicate that high glucose leads to an increase in generation of ROIs, an activation of NF-KB, and an induction of apoptosis by a glucose-specific and NO synthase-dependent mechanism. Our data suggest that peroxynitrite, which is rapidly formed from nitric oxide and superoxide anions, is the mediator of the cytotoxic effects of high glucose on endothelial cells. Because the induction of apoptosis by glucose was prevented by an antisense nucleotide to the p65NF-kappaB binding site, we assume that the ROI-mediated activation of NF-kappaB plays an important role for induction of apoptosis by glucose. PMID- 10515580 TI - Possible role of RAC-GTPase-activating protein in the termination of superoxide production in phagocytic cells. AB - The mechanism leading to the termination of superoxide production of phagocytes is poorly understood. The aim of the present study was to investigate the involvement of the active (GTP-bound) form of the GTP-binding proteins in maintaining continuous electron transport through the reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex. Activation of the enzyme was carried out under in vitro conditions and a shift from the active to the inactive form of the GTP-binding protein was attained (i) by addition of an excess of GDP to the assembled enzyme complex or (ii) by variation of the Rac GTPase activating (Rac-GAP) capacity of the constituents of the cell-free system. Significant inhibition of O2*- production was observed when guanine dinucleotides were added after the assembly of the active enzyme complex. The effect was specific for GDP and GDP,S whereas ADP, CDP and UDP were ineffective. GTP was significantly less efficient in inducing superoxide production in a cell-free system containing endogenous GAP activity than in a system devoid of GAP activity. It is suggested that the active, GTP-bound form of Rac is required for sustained catalytic function and Rac-GAP proteins are involved in the downregulation of the oxidase. PMID- 10515581 TI - Influence of nitric oxide on the intracellular reduced glutathione pool: different cellular capacities and strategies to encounter nitric oxide-mediated stress. AB - Different cell types exhibit huge differences towards the cytotoxic action of NO. In search for an explanation, we used subtoxic concentrations of the NO-donors S nitrosocysteine (SNOC) for short-term challenge and of (Z)-1-[N-(2-aminoethyl)-N (2-ammonioethyl)amino]diazen-1- ium-1,2-diolate (DETA/NO) for longer periods of exposure, respectively, and subtoxic concentrations of the oxidant H2O2 to determine the impact on intracellular reduced glutathione (GSH) concentrations. We find that GSH concentrations are always decreased, but that different cell types show different responses. Incubation of the relatively NO-sensitive murine lymphocytes with both NO-donors, but not with H2O2, resulted in a nearly complete loss of intracellular GSH. Short-term NO-treatment of P815 mastocytoma cells, also sensitive to NO-mediated cell death, decreased GSH to a similar extent only if either glutathione reductase (GSHR) activity or y-glutamylcysteine synthetase (gammaGCS) activity were inhibited concomitantly by specific inhibitors. Long term NO-treatment of P815 cells, however, resulted in a significant decrease of GSH that could be further enhanced by inhibiting gammaGCS activity. In contrast, neither short-term nor long-term NO-exposure nor H2O2-treatment affected intracellular GSH levels of L929 fibroblasts, which were previously shown to be extremely resistant towards NO, whereas concomitant gammaGCS inhibition, but not GSHR inhibition, completely decreased GSH concentrations. These results show that different cell types use different pathways trying to maintain glutathione concentrations to cope with nitrosative stress, and the overall capability to maintain a critical amount of GSH correlates with susceptibility to NO-induced cell death. PMID- 10515582 TI - Increased expression of inducible nitric oxide synthase and peroxynitrite in Helicobacter pylori gastric ulcer. AB - The role of nitric oxide in ulcer formation remains unknown. Accordingly, we assessed local expression of inducible nitric oxide synthase (NOS) and nitration of tyrosine as an indicator of peroxynitrite formation in patients with Helicobacter pylori (HP)-associated gastric ulcers compared with HP-negative ulcers. Biopsy specimens were taken from the ulcer margin and from an area remote from the ulcer portion. Inducible NOS, nitrotyrosine, and macrophage immunoreactivity were assessed immunohistochemically using a labeled streptavidin biotin method. In HP-positive gastric ulcers, inducible NOS and nitrotyrosine immunoreactivity was frequently observed at active ulcer margins, sometimes in surface epithelial cells as well as in the lamina propria. Occasionally, inducible NOS and nitrotyrosine reactivity were found in areas remote from the lesion in cases of HP-positive ulcer and HP-related gastritis. Macrophages accumulated significantly in the margin of HP-positive ulcers. In HP-negative gastric ulcers, inducible NOS and nitrotyrosine immunoreactivity also were frequent at the ulcer margin, but no significant immunoreactivity was observed at a distance. HP eradication caused significant attenuation in inducible NOS and macrophage immunoreactivity. In conclusion, nitric oxide and peroxynitrite formation is increased in HP-infected gastric mucosa, suggesting that HP promotes nitric oxide stress. PMID- 10515583 TI - Enhanced alloxan-induced beta-cell damage and delayed recovery from hyperglycemia in mice lacking extracellular-superoxide dismutase. AB - Alloxan is a diabetogenic agent which apparently acts through formation of superoxide radicals formed by redox cycling. Superoxide radicals are also formed by a variety of mechanisms in hyperglycemia. We exposed extracellular-superoxide dismutase (EC-SOD) null mutant and wild-type mice to alloxan, and followed up both the initial diabetes induction and the long-term course of the hyperglycemia. The null mutant mice responded with a modestly enhanced hyperglycemia compared to the wild type controls. In the long-term follow-up all mice eventually regained glycemic control, although it took longer for individuals with higher initial hyperglycemia. This delaying effect of the hyperglycemia was much more pronounced in the null mutant mice. These data suggest that the difference in initial diabetes induction between the groups is due to interception by EC-SOD of extracellular superoxide radicals produced by alloxan. The delayed recovery in the null mutant mice suggests that superoxide radicals released as a result of hyperglycemia impair beta-cell regeneration and that EC-SOD provides some protection. Mouse islets were found to contain little EC-SOD, whereas the content of the cytosolic Cu- and Zn-containing SOD was very high. This low EC-SOD activity may contribute to the high alloxan susceptibility of beta-cells, and may also cause a high susceptibility to superoxide radicals produced by activated inflammatory leukocytes and in hyperglycemia. PMID- 10515584 TI - Role of distinct subpopulations of peritoneal macrophages in the regulation of reactive oxygen species release. AB - It has been reported in vitro that during the respiratory burst of phagocytic cells the superoxide anion production per cell shows a negative relation with the cell density. This process has been described as autoregulation. The aim of this work was to analyze the superoxide anion production in thioglycollate-elicited peritoneal macrophage exudates to evaluate the importance of the peritoneal cavity environment in the autoregulation process. 12-O-tetradecanoylphorbol-13 acetate (PMA) was used to stimulate the respiratory burst and superoxide anion production was measured evaluating the intracellular formazan deposits that precipitate as a result of nitro blue tetrazolium (NBT) reduction. We have demonstrated a negative correlation between superoxide anion production and cell density in the peritoneal cavity in macrophages challenged with PMA. The response of individual cells was analyzed by means of an image analyzer, measuring the amount of formazan per cell and cell-size changes during the process of activation. The results revealed that the decrease in individual cell response as a function of higher cell densities were due to a significant increase in the amount of basal reaction macrophages. Concomitantly, the number of reactive cells remained unchanged irrespective of the cell density of the population. A direct correlation between cell size and superoxide anion production was observed. This phenomenon was demonstrated in SENCAR and Balb/c strains. However, macrophages from SENCAR mice showed greater superoxide anion production than those from Balb/c. The differences between strains could be associated to the increased sensitivity to PMA tumor promotion of SENCAR mice. Based on this property, macrophages from SENCAR mice were stimulated with opsonized zymosan, a particulate stimulus that reflects the interaction macrophage-microorganism during the phagocytic process. This data will contribute to the knowledge of infection control. We conclude that variations in basal reaction cells modulates the macrophage activation response when excess macrophages are recruited to the peritoneum. This is demonstrated using different stimuli, thus suggesting that this response may be applied to a wide variety of stimuli-macrophage interactions. The differences between strains may be associated to the increased sensitivity to PMA tumor promotion of SENCAR mice. PMID- 10515585 TI - Effects of reactive oxygen species on brain synaptic plasma membrane Ca(2+) ATPase. AB - The regulation of free intracellular calcium [Ca2+]i is altered in neurons from the aged brain, possibly due to reductions in the activity of Ca2+ transporters. The plasma membrane Ca(2+)-ATPase (PMCA) plays a critical role in Ca2+ homeostasis, and its kinetic properties change in aged rat brain. These changes could be due to oxidative modification of PMCA as a result of age-related chronic oxidative stresses. The present studies were undertaken to determine the sensitivity of the neuronal PMCA to in vitro exposure of synaptic plasma membranes (SPMs) to reactive oxygen species (ROS). We examined the effects of three oxidants including peroxyl radicals generated by azo-initiators, 2,2' Azobis 2-amidinopropane dihydrochloride (AAPH) and 4,4'-Azobis 14-cyanovaleric acid (ACVA), hydrogen peroxide (H2O2), and peroxynitrite (ONOO-). Synaptic plasma membranes briefly exposed to these oxidants were analyzed for functional and structural alterations in PMCA. Although all three oxidants led to significant loss of PMCA activity, the effect of ONOO- was the most potent, followed by peroxyl radicals and H2O2. Kinetic analysis of PMCA activity after oxidant treatment showed decreases in Vmax without significant changes in K(act). Immunoblots revealed oxidant-induced cross-linking of PMCA molecules that were partially reversed under reducing conditions and completely reversed with addition of urea. The PMCA appears to be very sensitive to inhibition by ROS and hence may be a target of oxidative stress in the aging brain. Reduction in its activity may contribute to age-related alterations in neuronal [Ca2+]i regulation. PMID- 10515586 TI - Interaction of hydrated electron with dietary flavonoids and phenolic acids: rate constants and transient spectra studied by pulse radiolysis. AB - The reaction rate constants and transient spectra of 11 flavonoids and 4 phenolic acids reacting with e(aq)- at neutral pH were measured. Absorption bands of the transients of e(aq)- reacting with the above compounds all located at a wavelength shorter than 400 nm. The e(aq)- scavenging abilities were divided into three groups: (+)catechin ((1.2 +/-0.1) x 10(8) M(-1)s(-1)) < 4-chromanol ((4.4 +/- 0.4) x 10(8) M(-1)s(-1)) < genistein ((6.2+/-0.4) x 10(9) M (-1) s(-1) approximately genistin ((8 +/- 1) x 10(9) M(-1)s(-1)) approximately rutin ((7.6 +/- 0.4) x M(-1)s(-1) approximately caffeic acid ((8.3 +/- 0.5) x 10(9)M(-1)s( 1)) < transcinnamic acid((1.1 +/- 0.1) x 10(10) M(-1)s(-1)) approximately p coumaric acid ((1.1 +/- 0.1) x 10(10) M(-1)s(-1) approximately 2,4,6 trihydroxylbenzoic acid((1.1 +/- 0.1) x 10(10) M(-1)s(-1)) approximately baicalein ((1.1 +/- 0.5) x 10(10) M(-1)s(-1)) approximately baicalin((1.3 + 0.1) X 10(10) M(-1)s(-1)) approximately naringenin ((1.2 +/- 0.1) x 10(10) M(-1)s(-1)) approximately naringin ((1.0 +/- 0.1) x 10(10) M(-1)s(-1)) approximately gossypin((1.2 +/- 0.1) x 10(10) M(-1)s(-1)) approximately quercetin((1.3 +/- 0.5) x 10(10) M(-1)s(-1)). These results suggested that C4 keto group is the active site for e(aq)- to attack on flavonoids and phenolic acids, whereas the o dihydroxy structure in B ring, the C2,3 double bond, the C3-OH group, and glucosylation, which are key structures that influence the antioxidant activities of flavonoids and phenolic acids, have little effects on the e(aq)- scavenging activities. PMID- 10515587 TI - Overexpression of catalase provides partial protection to transgenic mouse beta cells. AB - Pancreatic beta cells are sensitive to reactive oxygen species and this may play an important role in type 1 diabetes and during transplantation. Beta cells contain low levels of enzyme systems that protect against reactive oxygen species. The weakest link in their protection system is a deficiency in the ability to detoxify hydrogen peroxide by the enzymes glutathione peroxidase and catalase. We hypothesize that the deficit in the ability to dispose of reactive oxygen species is responsible for the unusual sensitivity of beta cells and that increasing protection will result in more resistant beta cells. To test these hypotheses we have produced transgenic mice with increased beta cell levels of catalase. Seven lines of catalase transgenic mice were produced using the insulin promoter to direct pancreatic beta cell specific expression. Catalase activity in islets from these mice was increased by as much as 50-fold. Northern blot analysis of several tissues indicated that overexpression was specific to the pancreatic islet. Catalase overexpression had no detrimental effects on islet function. To test whether increased catalase activity could protect the transgenic islets we exposed them to hydrogen peroxide, streptozocin, and interleukin-1beta. Fifty-fold overexpression of catalase produced marked protection of islet insulin secretion against hydrogen peroxide and significantly reduced the diabetogenic effect of streptozocin in vivo. However, catalase overexpression did not provide protection against interleukin-1beta toxicity and did not alter the effects of syngeneic and allogenic transplantation on islet insulin content. Our results indicate that in the pancreatic beta cell overexpression of catalase is protective against some beta cell toxins and is compatible with normal function. PMID- 10515588 TI - Melatonin induces gamma-glutamylcysteine synthetase mediated by activator protein 1 in human vascular endothelial cells. AB - In the present study, we show that melatonin induces the expression of gamma glutamylcysteine synthetase (gamma-GCS), the rate-limiting enzyme of glutathione (GSH) synthesis, in ECV304 human vascular endothelial cells. One micromolar melatonin induced the expression of gamma-GCS mRNA followed by an increase in the concentration of GSH with a peak at 24 h. An electrophoretic mobility shift assay showed that melatonin stimulates the DNA-binding activity of activator protein-1 (AP-1) as well as retinoid Z receptor/retinoid receptor-related orphan receptor alpha (RZR/RORalpha). ECV304 cells transiently transfected with a plasmid containing the gamma-GCS promoter-luciferase construct showed increased luciferase activity when treated with melatonin. The melatonin-dependent luciferase activity was found in the gamma-GCS promoter containing AP-1 site. The luciferase activity mediated by AP-1 was repressed in the promoter containing RZR/RORalpha site. In addition, cell cycle analysis showed that melatonin increases the number of cells in the G0/G1 phase; however, treatment of the cells with buthionine sulfoximine, a specific inhibitor of gamma-GCS, abolished the effect of melatonin on the cell cycle, suggesting induction of cell arrest by melatonin requires GSH. As conclusion, induction of GSH synthesis by melatonin protects cells against oxidative stress and regulates cell proliferation. PMID- 10515589 TI - Thermolabile 8-hydroxyguanine DNA glycosylase with low activity in senescence accelerated mice due to a single-base mutation. AB - 8-hydroxyguanine (8-oxoguanine; oh8Gua) DNA glycosylase (OGG1) repairs oh8Gua, a highly mutagenic oxidative DNA damage. In the present study, we compared two strains of senescence-accelerated mouse (SAM) expressing senescence-prone phenotypes, SAMP1 and SAMP8, with one strain of SAM expressing senescence resistant phenotype, SAMR1. We found three distinct characteristics of OGG1 in SAMPs: (i) low activity (10-40% of the SAMRI enzyme in all organs and ages observed), (ii) thermolability, and (iii) mutation from Arg (CGG) in SAMR1 to Trp (TGG) at codon 304. There was no difference in the levels of mRNA and protein. As expected, oh8Gua level in tissues was higher in the SAMPs. In contrast, O6 methylguanine-DNA methyltransferase, which repairs alkylated DNA, showed no difference in its activity. The impairment of oh8Gua repair activity caused by the 304 mutation in OGG1 may be one of the factors contributing to the high somatic mutation rate and the accelerated senescence observed in these strains. PMID- 10515590 TI - Scavenging mechanisms of (-)-epigallocatechin gallate and (-)-epicatechin gallate on peroxyl radicals and formation of superoxide during the inhibitory action. AB - The scavenging effects of (-)-epigallocatechin gallate (EGCG) and (-)-epicatechin gallate (ECG) on peroxyl radicals and their mechanisms were studied by investigating the products formed during the first stages by 2,2'-azobis(2 aminopropane) hydrochloride (AAPH)-induced oxidation, without any isolation, using LC/MS, spectrophotometry, chemiluminescence analyses, and semiempirical molecular orbital (MO) calculations. The results show that EGCG can be converted to an anthocyaninlike compound followed by cleavage of the gallate moiety by oxidation. On the other hand, ECG can be converted to an anthocyaninlike compound after cleavage of the gallate moiety. The calculated C-H bond dissociation enthalpies (BDEs) for EGCG and ECG at the C-2 position were quite low (62.7 and 66.8 kcal/mol, respectively) compared with O-H BDEs at the phenolic sites (ca. 70 kcal/mol), suggesting that the C-2 hydrogen can be abstracted by free radicals. The addition of superoxide dismutase (SOD) decreased the chemiluminescence in EGCG by one-half during the inhibitory action. Active oxygen including superoxide (O2-) would be produced in EGCG, but not in ECG. The authors proposed the antioxidative mechanisms of EGCG and ECG depending on the experimental results and theoretical calculations. PMID- 10515591 TI - Reassignment of organic peroxyl radical adducts. AB - The study of the important role of peroxyl radicals in biological systems is limited by their difficult detection with direct electron spin resonance (ESR). Many ESR spectra were assigned to 5,5-dimethyl-1-pyrroline N-oxide (DMPO)/peroxyl radical adducts based only on the close similarity of their ESR spectra to that of DMPO/superoxide radical adduct in conjunction with their insensitivity to superoxide dismutase, which distinguishes the radical adduct from DMPO/superoxide radical adduct. Later, the spin-trapping literature reported that DMPO/peroxyl radical adducts have virtually the same hyperfine coupling constants as synthesized alkoxyl radical adducts, raising the issue of the correct assignment of peroxyl radical adducts. However, using 17O-isotope labelling, the methylperoxyl and methoxyl radical adducts should be distinguishable. We have reinvestigated the spin trapping of the methylperoxyl radical. The methylperoxyl radical was generated in aerobic solution with 17O-molecular oxygen either in a Fenton system with dimethylsulfoxide or in a chloroperoxidase system with tert butyl hydroperoxide. Two different spin traps, DMPO and 2,2,4-trimethyl-2H imidazole-1-oxide (TMIO), were used to trap methylperoxyl radical. 17O-labelled methanol was used to synthesize methoxyl radical adducts by nucleophylic addition. It was shown that the 17O hyperfine coupling constants of radical adducts formed in methylperoxyl radical-generating systems are identical to that of the methoxyl radical adduct. Therefore, methylperoxyl radical-producing systems form detectable methoxyl radical adduct, but not detectable methylperoxyl radical adducts at room temperature. One of the possible mechanisms is the decomposition of peroxyl radical adduct with the formation of secondary alkoxyl radical adduct. These results allow us to reinterpret previously published data reporting detection of peroxyl radical adducts. We suggest that detection of 17O alkoxyl radical adduct from 17O-labelled molecular oxygen can be used as indirect evidence for peroxyl radical generation. PMID- 10515592 TI - Evidence for free radical formation during the oxidation of 2'-7' dichlorofluorescin to the fluorescent dye 2'-7'-dichlorofluorescein by horseradish peroxidase: possible implications for oxidative stress measurements. AB - The oxidation of 2'-7'-dichlorofluorescin (DCFH) to the fluorescent 2'-7' dichlorofluorescein (DCF) by horseradish peroxidase (HRP) was investigated by fluorescence, absorption, and electron spin resonance spectroscopy (ESR). As has been previously reported, HRP/H2O2 oxidized DCFH to the highly fluorescent DCF. However, DCF fluorescence was still observed when H2O2 was omitted, although its intensity was reduced by 50%. Surprisingly, the fluorescence increase, in the absence of exogenous H2O2, was still strongly inhibited by catalase, demonstrating that H2O2 was present and necessary for DCF formation. H2O2 was apparently formed during either chemical or enzymatic deacetylation of 2'-7' dichlorofluorescin diacetate (DCFH-DA), probably by auto-oxidation. Spectrophotometric measurements clearly showed that DCFH could be oxidized either by HRP-compound I or HRP-compound II with the obligate generation of the DCF semiquinone free radical (DCF*-). Oxidation of DCF*- to DCF by oxygen would yield superoxide (O2*-). ESR spectroscopy in conjunction with the spin trap 5,5 dimethyl-1-pyrroline N-oxide (DMPO) revealed the presence of both superoxide and hydroxyl radicals in the DCFH/H2O2/HRP system. Both radicals were also detected in the absence of added H2O2, although the intensities of the resultant adducts were decreased. This work demonstrates that DCF fluorescence cannot be used reliably to measure O2*- in cells because O2*- itself is formed during the conversion of DCFH to DCF by peroxidases. The disproportionation of superoxide forms H2O2 which, in the presence of peroxidase activity, will oxidize more DCFH to DCF with self-amplification of the fluorescence. Because the deacetylation of DCFH-DA, even by esterases, can produce H2O2, the use of this probe to measure H2O2 production in cells is problematic. PMID- 10515593 TI - HDL is the major source of vitamin E for type II pneumocytes. AB - The alveolar surfactant is a prime target of reactive oxygen species present in air. Alveolar surfactant is supplemented with vitamin E during its assembly in type II pneumocytes. However, it is unknown which of the lipoproteins supply type II pneumocytes with vitamin E. The measurement of the uptake kinetics indicates that HDL might be the primary source of the vitamin E uptake by type II pneumocytes. Vitamin E depletion of rats caused an increase of vitamin E uptake by isolated type II pneumocytes from HDL but not from LDL or VLDL. We demonstrated that type II pneumocytes express the scavenger receptor class B type 1 (SR-B1), an HDL-specific receptor. Vitamin E depletion caused an increased expression of SR-B1 by a post-transcriptional mechanism. The increased vitamin E uptake from HDL and the increased expression of the SR-B1 were reversed by refeeding the vitamin. We propose that HDL is the primary source of vitamin E for type II pneumocytes. The rate of uptake of vitamin E by this cell type might be regulated by the expression of SR-B1. PMID- 10515594 TI - Ethanol stimulates the production of reactive oxygen species at mitochondrial complexes I and III. AB - The aim of this study was to investigate the hepatocellular site of reactive oxygen species generation during acute ethanol metabolism. Reactive oxygen species production was detected using the 2',7'-dichlorofluorescein fluorescence assay and cell injury was determined by lactate dehydrogenase release. Incubation with 1 and 10 mM ethanol increased the production of reactive oxygen species by 72% and 151%, respectively, which was associated with mild decreases in cell viability. Antimycin, a mitochondrial complex III inhibitor, elicited a 17-fold increase in the levels of reactive oxygen species and markedly decreased hepatocyte viability and ATP levels. Ethanol increased reactive oxygen species production and the cytosolic NADH/NAD+ ratio in antimycin-treated cells. Rotenone, a mitochondrial complex I inhibitor that allows electron flow through the flavin mononucleotide (FMN), but prevents electron flow to complex III, significantly increased reactive oxygen species production in untreated cells, but decreased reactive oxygen species production in antimycin plus ethanol treated cells. Diphenyliodonium, a mitochondrial complex I inhibitor that inhibits electron flow through FMN, attenuated reactive oxygen species generation in all groups. Fructose prevented cytotoxicity in all treatment groups. Though they do not eliminate the participation of other intracellular compartments, these results indicate that the NADH dehydrogenase complex, as well as complex III of mitochondria, are involved in ethanol-related production of reactive oxygen species. PMID- 10515595 TI - Thyroid status modulates glycoxidative and lipoxidative modification of tissue proteins. AB - Steady state protein modification by carbonyl compounds is related to the rate of carbonyl adduct formation and the half-life of the protein. Thyroid hormones are physiologic modulators of both tissue oxidative stress and protein degradation. The levels of the glycation product N(epsilon)-fructoselysine (FL) and those of the oxidation products, N(epsilon)-(carboxymethyl)lysine (CML) and malondialdehyde-lysine (MDA-lys), identified by GC/MS in liver proteins, decreased significantly in hyperthyroid rats, as well as (less acutely) in hypothyroid animals. Immunoblotting of liver proteins for advanced glycation end products (AGE) is in agreement with the results obtained by GC/MS. Cytosolic proteolytic activity against carboxymethylated foreign proteins measured in vitro was significantly increased in hypo- and hyperthyroidism. Oxidative damage to DNA, estimated as 8-oxo-7,8-dihydro-2'-deoxyguanosine (8oxodG), did not show significant differences between groups. The results suggests that the steady state levels of these markers depend on the levels of thyroid hormones, presumably through their combined effects on the rates of protein degradation and oxidative stress, whereas DNA is more protected from oxidative damage. PMID- 10515596 TI - Primate herpesviral oncogenes. AB - Gammaherpesviruses are the most rapidly growing members of the herpesviridae family. Gamma herpesviruses share similarity in their genome organizations and in early and late lytic genes that are required for viral replication. A distinct characteristic of gamma herpesviruses is their ability to establish latent infection in lymphoid cells, and some of these viruses are closely associated with abnormal proliferation and cancer in primates. The first open reading frame of the primate gamma herpesviruses has been shown to directly contribute to virus associated pathogenesis. This open reading frame encodes latent membrane protein 1 (LMP1) in Epstein-Barr virus, Saimiri transformation protein (STP) in Herpesvirus Saimiri, K1 in Kaposi's sarcoma-associated herpesvirus, and R1 in Rhesus monkey Rhadinovirus. All of these gene products are capable of eliciting cellular signal transduction events, resulting in cell growth transformation. This review briefly summarizes the current view on the transforming mechanisms utilized by primate herpesviral oncogenes. PMID- 10515597 TI - Z-curve: a computer program calculating DNA helical axis coordinates for three dimensional graphic presentation of curvature. AB - In order to predict curvature of DNA fragments, we previously developed a computer program for simply calculating a vectorial sum of all individual roll, tilt and twist wedge angles between the nearest base pairs for a given DNA fragment [Lee et al., (1991)]. Now, a new program, called Z-curve, was developed to calculate three-dimensional coordinates of the helical center of each base pair along the DNA, using helical axis deviations from B-form DNA by wedge angles. The output file of the new program was designed to become an input file for a graphics program, Insight II. Thus, we were able to obtain three dimensional graphic presentations of DNA helical axis curvatures of any length. It visualized spatial details of the DNA curvature, where and how much it curves, and to which direction. It also allowed calculation of the three-dimensional distance between two ends of a DNA fragment, which could provide a measure of its curvature. Here, three DNA fragments, both curved and straight, were subjected to the Z-curve and Insight II programs. The results showed that their curvature details could be visualized to the level of the base pair, whether the DNA fragments contained an oligo(A) track or not. Their estimated curvatures were consistent with the experimental results of permutation gel mobility assay. PMID- 10515598 TI - A new autoantigen reactive with prediabetic nonobese diabetic mice sera. AB - The identification and characterization of new autoantigens would widen the knowledge of the pathogenic mechanism of insulin dependent diabetes mellitus. Screening of lambda gt11 mouse insulinoma (MIN6N8a) cell cDNA library with prediabetic nonobese diabetic (NOD) mice sera resulted in the isolation of a strong positive clone, named the clone 3-5, of 1579 nucleotides without a poly A region. After 5'-rapid amplification of the cDNA end (RACE), complete nucleotide sequence of the clone 3-5 gene consisting of 2231 nucleotides showed that the 3-5 gene had the theoretical open reading frame of 634 amino acids. However, the real antigenic protein of the clone 3-5 was only 21 amino acids long encoded by only 63 nucleotides. The 21 amino acids were expressed as a fusion protein in E. coli and purified by affinity chromatography. The purified 3-5 recombinant protein was examined for its reactivity with prediabetic NOD mice sera by immunoblotting. The only non-denatured form of the 3-5 protein showed a binding reactivity with NOD mice sera, demonstrating that the conformational epitope of 3-5 protein was important for antibody recognition. The prevalence of autoantibody reactive to the 3-5 protein was about 78% (14/18) and 46% (11/24) in prediabetic and acute diabetic NOD mice sera, respectively. However, the sera from other mouse strains such as BALB/c, ICR, C57BL/6, SJL/J, and NOD/SCID did not show a positive reactivity to the 3-5 protein, which indicated that immune reactivity against the 3-5 protein was autoimmune diabetic mouse-specific. PMID- 10515599 TI - Distribution of ganglioside GM3 in the rat ovary after gonadotropin stimulation. AB - Gangliosides are ubiquitous membrane components in mammalian cells and are suggested to play important roles in various cell functions, such as cell-cell recognition, differentiation and transmembrane signalling. Ovaries have been shown to contain GM3 as a major ganglioside. To study GM3 distribution during gonadotropin stimulation in the hypophysectomized rat ovary, ovarian sections and cultured granulosa cells were stained with specific monoclonal antibody against GM3. Interstitial cells of follicles of immature hypophysectomized rat ovary expressed ganglioside GM3. Theca cells of early antral follicles but not primary follicles expressed GM3. No granulosa cells of these follicles expressed GM3. When a surge dose of FSH/LH was injected, Graafian follicles were formed and GM3 expression was detected in granulosa cells of these follicles. After ovulation, cumulus cells kept expressing GM3 in the ampulla region of ovulated oviduct. The follicles did not show GM3 expression in their granulosa cells after an ovulatory dose of FSH/LH. At 48 h after in vitro culture with FSH/LH of granulosa cells from preantral follicles, GM3 was expressed to a detectable extent on the outer part of the granulosa layer. Finally, at 72 h after culture, all granulosa cells became positive to anti-GM3 antibody. These data suggest that the expression of ganglioside GM3 in the hypophysectomized rat ovary is spatiotemporally regulated by FSH/LH during follicular development and ovulation. PMID- 10515600 TI - Gene silencing-mediated resistance in transgenic tobacco plants carrying potato virus Y coat protein gene. AB - Unlike other pathogens, plant viruses are hardly controlled by chemical agents. Potato virus Y (PVY) is distributed around the world, and causes a great loss economically. In an attempt to minimize the damage by viruses, the PVY coat protein (CP) gene was introduced into tobacco by Agrobacterium-mediated transformation. A significant proportion of the transgenic plants displayed resistance to PVY and showed substantially decreased CP transgene expression at both protein and steady-state mRNA levels compared to susceptible transgenic or nontransgenic plants. A resistant plant was selected and self-fertilized for several generations until T4 progenitor lines were obtained. Most of these T4 plants accumulated extremely low levels of CP protein and steady-state mRNA, and exhibited almost complete resistance to PVY. DNA gel blot analysis revealed that the transgenic plants typically had two or three copies of the transgene. These results are characteristic of pathogen-derived resistance, in which the resistance against virus is the consequence of post-transcriptional gene silencing directed by homologous transgenes. To uncover factors that may play roles in gene silencing, sequences in the 3' part of the transcribed region of the CP gene were transcribed in vitro and the RNA fragments were incubated with cell extracts from transgenic plants. A ribonuclease activity was detected that appeared to be specific for this transcript in the PVY-resistant transgenic plants. PMID- 10515601 TI - Effective immunotherapy of cancer by DNA vaccination. AB - Direct injection of naked plasmid DNA either intramuscularly or intradermally induces strong, long-lived cell-mediated and humoral immune responses to the antigen encoded by the gene vaccine. In the present study, we used gene vaccination with naked plasmid DNA to induce prophylactic immune responses to tumor associated antigens. MAGE-1 (melanoma antigen 1) is an ideal candidate for cancer vaccines because it belongs to a family of genes that are expressed in a number of human tumors of various histological types but not in normal adult tissues except for the testis, and because both humoral and cell-mediated immune responses against MAGE-1 antigen were detected in tumor patients. Intradermal administration of plasmid DNA encoding MAGE-1 (pcMAGE1) induced anti-MAGE-1 specific antibody in BALB/c mice. In contrast, no detectable level of anti-MAGE-1 antibody was induced by intramuscular injection of pcMAGE1. Also, intradermal injection of pcMAGE1 was capable of generating CTLs reactive with MAGE-1 transfected murine tumor cells, M-MSV-MAGE1. Most of the mice (8 out of 10) immunized with pcMAGE1 rejected the challenge of M-MSV-MAGE1 tumor cells, compared with control animals most of which developed tumors. This suggests that intradermal DNA vaccination could provide a novel immunotherapy of cancer. PMID- 10515602 TI - Isolation and characterization of a cDNA encoding the cysteine proteinase inhibitor, induced upon flower maturation in carnation using suppression subtractive hybridization. AB - The suppression subtractive hybridization (SSH) method was used to isolate differentially expressed genes during carnation flower maturation. Five cDNA clones, designated as carnation flower maturation-induced (CFMI), were verified as flower maturation-induced cDNAs. Sequence analysis of five CFMI (CFMI-5, CFMI 6, CFMI-7, CFMI-9, and CFMI-10) clones revealed that one of the clones, CFMI-5, showed high sequence similarity to the cysteine proteinase inhibitor gene, predicted to be involved in flower maturation. The full length cDNA clone CFMI-5 was 531 nucleotides (nts) long and consisted of an open reading frame of 294 nucleotides, encoding a 98 amino acid protein, 12 nucleotides of 5'-untranslated region and 3'-untranslated region (225 nts) with a poly(A)+ tail. The predicted CFMI-5 amino acid sequence had a conserved sequence Gln-Val-Val-Ala-Gly, which corresponds to the active site of proteinase inhibition. Northern blot analysis revealed tissue-specific expression of CFMI-5 transcripts, as the transcripts were expressed preferentially in petals and styles. A PCR-based cDNA subtraction method, termed suppression subtractive hybridization, was identified as a rapid method to screen differentially expressed genes in a short time. PMID- 10515603 TI - Expanded polyglutamine tract itself induces cell death in cultured cells. AB - Several neurodegenerative diseases including Huntington disease, Machado-Joseph disease and spinocerebellar ataxias type 1 are caused by expansion of a polyglutamine tract within their respective gene products. In order to assess the role of the tract, 293T cells were transfected with plasmids that contain various lengths of CAG repeat encoding polyglutamine without the repeat disorder proteins: (CAG)27, (CAG)40, (CAG)80, (CAG)130, and (CAG)180. Except for (CAG)27, and (CAG)40, 293T cells showed a common set of morphological alterations such as shrinkage, rounding and surface blebbing when the expanded stretch was expressed. In addition, nuclear staining experiments showed chromatin condensation in COS-7 cells transfected with the vectors containing expanded CAG repeats. These results indicate that expanded polyglutamine itself is able to induce cell death, suggesting existence of a common molecular mechanism in the etiology of neurodegenerative polyglutamine diseases. PMID- 10515604 TI - Chromosomal localization of ESTs obtained from human fetal liver via BAC-mediated FISH mapping. AB - A total of 55 expressed sequence tags (ESTs) randomly chosen from our collection of fetal liver ESTs were mapped to chromosomes by fluorescence in situ hybridization (FISH) mapping techniques. To generate FISH mapping probes, the genomic DNAs for each EST were selected by screening an arrayed human bacterial artificial chromosome (BAC) library. In total, 73 BACs were used for mapping of the 55 ESTs. Among them, 70 BACs representing 52 ESTs unequivocally mapped to single chromosomal regions. The remaining 3 BACs representing 3 ESTs were localized to multiple regions, suggesting that BACs may have very low chimerism. Our mapping results were compared with EST mapping databases deposited in NCBI. Thirty-six of 55 ESTs corresponded to previously mapped positions of ESTs, 2 ESTs mapped to different positions from previously determined ones, and it was found that 17 ESTs have been mapped on new locations from this study. These mapping data may be used for completing the framework of the human physical map, and also for providing a good starting point for searching disease-related genes. PMID- 10515605 TI - Antifungal activities of recombinant antifungal protein by conjugation with polyethylene glycol. AB - Tenecin 3, an antifungal protein isolated from coleopteran insect Tenebrio molitor larvae, inhibited growth of the fungus Candida albicans. We have previously reported that tenecin 3 has a propensity of random structure with very loose turn-like elements by circular dichroism (CD) analysis and 2D nuclear overhauser effect spectroscopy [Lee et al. (1999)]. However, the antifungal mechanism of tenecin-3 has not yet been studied due to its very low availability from natural sources. As an initial step to study the antifungal mechanism of tenecin 3, recombinant tenecin 3 (RT-3) obtained from an expression system in Escherichia coli showed antifungal activity against C. albicans as did natural tenecin 3. To elucidate the antifungal mechanism of RT-3 and to explore the possibility of preparing polyethylene glycol (PEG) conjugated derivative, we synthesized PEG conjugated RT-3 (RT-3-PEG) and examined its antifungal activity against C. albicans in vitro. RT-3-PEG showed greater antifungal activity against C. albicans than RT-3 alone at the same dose. When C. albicans was treated with RT-3-PEG in vitro, K+ in the C. albicans cell was leaked out rapidly compared to the C. albicans treated with RT-3 alone. When the morphological change of RT-3 PEG treated C. albicans was examined by scanning electron microscopy, string-like substances, which may have been derived from the fungus, were stacked around the cell whose wall was damaged. Also, no appreciable hemolysis of mouse erythrocytes was detected under conditions in which 1% melittin caused 100% hemolysis. These results suggested that the RT-3-PEG derivative probably does not interact with mammalian cell appreciably, although it has antifungal activity. PMID- 10515607 TI - Fetal mouse selenophosphate synthetase 2 (SPS2): biological activities of mutant forms in Escherichia coli. AB - A novel gene, sps2, detected in mouse embryo at the early stages of development has been identified as an analog of the E. coli selenophosphate synthetase gene. Unlike the E. coli enzyme, the presence of selenocysteine in the mouse enzyme is indicated by a TGA codon in the open reading frame of the cDNA. Using an N-FLAG monoclonal antibody, it was shown that the full length N-FLAG-sps2 gene product was expressed in COS-7 cells. To investigate the biological activity of the sps2 gene product in vivo, the mutated sps2 gene, which contains cysteine in the place of the TGA encoded selenocysteine in the wild type, was expressed in the E. coli selD deficient mutant, MB08. Like the E. coli wild type selD gene, the mutant sps2 gene complemented the selD mutation. However, replacement of Cys with either Ala, Ser, or Thr resulted in a loss of ability to complement the selD mutation. The SPS2-CYS protein expressed in E. coli was purified and its catalytic activity was determined. The Km value for ATP was 0.75 mM and Vmax was 9.23 nmole/min/mg protein. These results confirm that the mouse embryonic sps2 gene encodes an eukaryotic selenophosphate synthetase, and that availability of selenophosphate as a selenium donor compound is widespread. PMID- 10515606 TI - Localization of dopamine D1 and D2 receptor mRNAs in the rat systemic and pulmonary vasculatures. AB - The present study was designed to evaluate the expression of dopamine D1 and D2 receptor mRNAs in systemic and pulmonary vasculatures. Using specific antisense riboprobes for dopamine D1 and D2 receptor cDNAs, in situ hybridization histochemistry was performed in the aorta, common carotid artery, vertebral artery, pulmonary artery, and superior vena cava of the adult male Sprague Dawley rat. In the case of the aorta, common carotid artery, and vertebral artery, dopamine D1 receptor mRNAs localized mainly in the smooth muscle cells of the tunica media. However, the signals of dopamine D2 receptor mRNAs were found in the endothelium and subendothelial layer of tunica intima, and interstitial cells of tunica adventitia. In the case of the pulmonary artery, signals of dopamine D1 receptor mRNAs were detected within the tunica intima, media, and adventitia. Expression of D2 receptor mRNAs was detected in the walls of small blood vessels within the tunica adventitia of the pulmonary artery. There were no detectable signals of dopamine D1 and D2 receptor mRNAs in the vein. The uneven distribution of dopamine D1 and D2 receptor mRNAs in the rat systemic vasculatures and pulmonary artery suggests that dopamine differentially regulates the vasodilation of the systemic and pulmonary arteries through the differential stimulation of dopamine D1 and D2 receptor. PMID- 10515609 TI - Metaphase chromosome accumulation and flow karyotypes in rice (Oryza sativa L.) root tip meristem cells. AB - Highly efficient cell synchronization and metaphase chromosome accumulation in rice root tip cells were achieved. Flow cytometric analysis was performed for obtaining optimal parameters to synchronize the cell cycles. High mitotic indices (about 57.6% in root tip meristemic area) were obtained by treating seedlings with 0.5 cm length using 0.5 mM hydroxyurea at 30 degrees C for 4 h, incubating in a hydroxyurea-free solution for 30 min, and then treating with 0.3 microM trifluralin for 3 h. After trifluralin treatment, incubation in distilled water for 15 min reduced chromosome clumping on metaphase spread. Uniformity of seed germination at the time of treatment is a critical parameter for obtaining high metaphase index. Isolated rice chromosomes were suitable for flow cytometric analysis and chromosome sorting. The morphology of flow sorted metaphase chromosomes was intact. PMID- 10515608 TI - Alteration of the NAD+/NADH ratio in CHO cells by stable transfection with human cytosolic glycerol-3-phosphate dehydrogenase: resistance to oxidative stress. AB - The intracellular level of the NAD+/NADH ratio plays a vital role in sustaining and coordinating the catabolic reaction of the cell, and reflects the redox state of cytosol. Antioxidants play a role to protect cytosol and membrane from free radicals. This role of antioxidants involves sustaining cell viability and the procedure is thought to be regulated by the equilibrium of the redox state of the cell. However, there is very little known about how the NAD+/NADH level is set and changed. To alter the ratio, human NAD-dependent glycerol-3-phosphate dehydrogenase (cGPDH) cDNA was transfected stably in CHO dhfr- cells. When compared to parental CHO cells, cGPDH activities of the transfected cells were increased 8-12 fold, but the NAD+/NADH ratio was decreased. Specific growth rate of the transfected cells was similar to or slight lower than that of wild type CHO cells. Cell viability of the stable transformants against H2O2 was increased without change of either catalase or glutathione peroxidase activity. However, the increase of cell viability was correlated with the decrease of NAD+/NADH ratio in transfectants. From these results, it is suggested that the overexpression of cGPDH changes the NAD+/NADH ratio toward a decrease, and by this change in the redox state the cell confers more resistance against H2O2. PMID- 10515610 TI - Characterization of ephrin-A1 and ephrin-A4 as ligands for the EphA8 receptor protein tyrosine kinase. AB - The Eph receptors are the largest known family of receptor protein tyrosine kinases, which play important roles with their ligands called ephrin in the neural development, angiogenesis, and vascular network assembly. It was previously shown that ephrin-A2, -A3 and -A5 bind to, and activate the EphA8 receptor tyrosine kinase, respectively. In this study, we have examined if there are other additional ephrin ligands interacting with the EphA8 receptor tyrosine kinase expressed in NIH3T3 fibroblasts. For this purpose, we have constructed chimeric ephrin-A1, -A4, -B1, -B2 or -B3 ligands consisting of the Fc portion of human IgG fused to their carboxyl-terminus. Both ephrin-A1 and ephrin-A4 chimeric ligands efficiently bound to the EphA8 receptor expressed in NIH3T3 fibroblasts, whereas the transmembrane ligands including ephrin-B1, -B2 and -B3 did not. Additionally we have demonstrated that both the EphA8-TrkB chimeric receptor and the EphA8 receptor expressed in NIH3T3 fibroblasts are efficiently tyrosine phosphorylated upon stimulating with epthin-A1 or -A4 but none of transmembrane ephrin-B proteins. These results strongly indicate that the EphA8 receptor functions exclusively as an glycosyl phosphatidylinositol (GPI)-linked ephrin ligand-dependent receptor protein tyrosine kinase. PMID- 10515611 TI - Analysis of integration activity of human immunodeficiency virus type-1 integrase. AB - The integration activity of human immunodeficiency virus type-1 (HIV-1) integrase was characterized in vitro by using pre-processed oligonucleotide substrates. The highest level of integration activity was found at pH 6.5 to 7.0, while the endonucleolytic activity was highest at pH 7.4 to 8.0. Although the endonucleolytic and integration reactions are consecutive in retroviral integration, our result indicates that the optimal conditions of the two reactions are quite different. In addition, it is suggested that the endonucleolytic and integration steps can be separated by control of the cellular physiological state in retroviral therapy. Strong integration was detected in the presence of 0.5-10 mM Mn2+ ion, but weak integration at around 10 mM Mg2+ ion. This observation explains that the Mn2+ ion is preferred to the Mg2+ ion as a cofactor in the integration reaction. Although there was no sequence-specificity in the integration site of the target DNA, integration was found to frequently occur at particular regions of the target DNA. Furthermore, the mutant integrases such as Asp116, Ser147, and Glu152, which had been reported previously, were shown to lose integration activity completely, indicating that these residues are critically involved in catalytic action. PMID- 10515612 TI - Identification of phage-displayed peptides which bind to the human HnRNPA1 protein-specific monoclonal antibody. AB - We have screened a peptide phage display library to examine if monoclonal antibody-binding phages could be isolated from the library and thereby predict the antigenic epitopes of the antibodies from the isolated phages. The library was screened for high-avidity binding to monoclonal antibodies by an affinity purification technique called biopanning. Among the monoclonal antibodies examined, the human hnRNPA1 protein-specific monoclonal antibody 9H10 showed selective binding of phages. After two rounds of the biopanning, twelve clones of high-avidity-binding phages were chosen and their inserts were sequenced. Nucleotide sequence comparison of the 12 clones showed that there were 5 different species, with two species containing four members, implying that they were predominantly selected by the biopanning. The amino acid sequences of the inserts of the 12 clones were compared with that of the human hnRNPA1 protein in order to find the putative epitope of the human hnRNPA1 protein for 9H10. The C terminal region of the human hnRNPA1 protein shows significant homology with the peptide sequences of the selected phage clones. These results show that this peptide phage display library can be useful in defining the epitope of some monoclonal antibodies. PMID- 10515614 TI - CAP reform: the government sets out the options. PMID- 10515613 TI - A method of microinjection: delivering monoclonal antibody 1223 into sea urchin embryos. AB - In this paper, a simpler method of microinjecting sea urchin embryos without using the conventional microinjection chamber designed by Kiehart is reported. A trough was made on a surface of 0.6% agarose gel dissolved in artificial sea water. Approximately fifty hatched embryos could be loaded in the trough and, consequently, swimming embryos were trapped in the trough. Monoclonal antibody (mAB) 1223 which blocks spiculogenesis in vitro was delivered into the blastocoels of sea urchin embryos to test whether this antibody inhibits spiculogenesis in vivo and also, whether this new technique is effective for the microinjection of the sea urchin embryos. The embryos were injected with mAB1223 at the hatched blastula, early mesenchyme blastula and early gastrula stages, and 63%, 90% and 97% of the embryos did not form spicules at the late gastrula stage, respectively. Therefore, mAB1223 was shown to also block spiculogenesis in vivo. From the fact that spiculogenesis occurred at a lower rate when mAB1223 was injected at the hatched blastula stage than at later stages, it may be speculated that endogenous proteases degraded the injected antibodies. Using this technique, extracellular events in the blastocoel or the function of certain molecules expressed in blastocoel can be easily investigated in vivo. PMID- 10515615 TI - Demographic characteristics of the equine population of northern Britain. AB - The size, composition and distribution of the equine population of Scotland and the five northernmost counties in England were estimated through a series of mailed questionnaire surveys of sentinel veterinary practices and horse owners. An estimated 96,622 equine animals were kept by an estimated 26,114 owners. The mean (sd) age of the population was 11.0 (7.5) years (range one month to 37 years). Thoroughbred or thoroughbred-cross animals were the most numerous, constituting 30 per cent (95 per cent confidence interval 27 to 33 per cent) of the total population. The ratio of males:females was 1:1. PMID- 10515616 TI - Cowdria ruminantium infection in ticks in the Kruger National Park. AB - Adult Amblyomma hebraeum ticks, the principle vector of heartwater (cowdriosis) of domestic ruminants in southern Africa, were collected in pheromone traps placed in Kruger National Park, an exclusively wildlife sanctuary in South Africa. These ticks transmitted Cowdria ruminantium, the rickettsial agent causing heartwater, to a susceptible goat, resulting in acute, fatal disease. C ruminantium was isolated in bovine endothelial cell culture from the plasma of this animal during the febrile stage of the disease and transmitted to susceptible goats, causing fatal heartwater. The prevalence of C ruminantium infection in 292 ticks was determined by polymerase chain reaction (PCR) analysis to be 1.7 per cent (95 per cent confidence interval 0.71 to 4.0 per cent). A DNA probe analysis, which is less sensitive than PCR, detected infection in three of the five PCR-positive ticks. The remaining infections were below the detection limit of the DNA probe, which is approximately 70,000 organisms. This is the first evidence that a vector-wildlife cycle of transmission of C ruminantium can be maintained independently of domestic ruminants. PMID- 10515617 TI - Clinical, haematological and biochemical findings in milk-fed calves with chronic indigestion. AB - The principal clinical signs in 59 milk-fed calves with chronic indigestion were general malaise and depression, poor appetite, poor body condition, dehydration, a dull and scaly hair coat, alopecia and clay-like faeces. All the calves had metabolic acidosis, which was associated with an inability to stand up in more than half of them. There were significant differences in the severity of acidosis between the calves that could stand and those that could not. Other signs in some of the calves were dehydration, leucocytosis, and increased activities of liver enzymes. PMID- 10515618 TI - Experimental cross-infections with Ehrlichia phagocytophila and human granulocytic ehrlichia-like agent in cows and horses. AB - Four cows and four horses were infected experimentally with Ehrlichia phagocytophila, the cause of tickborne fever in ruminants, and with human granulocytic ehrlichia-like agent, a recently discovered species that infects people, horses and dogs in the USA and Europe. They were infected in either order, 30 days apart, to investigate serological cross-reactivity within the Ephagocytophila genogroup. The course of infection was assessed by routine clinical, haematological, serological and polymerase chain reaction (PCR) examinations. Two of the cows infected with Ephagocytophila and two of the horses infected with granulocytic ehrlichia-like agent, developed characteristic signs of ehrlichiosis. When the same animals were infected with their heterologous ehrlichial isolate 30 days later, they did not develop clinical signs of disease. The infection of the other two cows with human granulocytic ehrlichia-like agent and the other two horses with Ephagocytophila, resulted in asymptomatic seroconversion. When the same animals were infected with their homologous ehrlichial isolate 30 days later, they remained asymptomatic and had normal haematological results and negative PCRS until the end of the monitoring period, 60 days after the first infection. In these animals, there was an increase in antibody titre after the second infection which was interpreted as a specific immune response, and as a reactivation of the immune response to the first infection. PMID- 10515619 TI - Reduction of bacterial contamination of bovine semen by filtration. PMID- 10515620 TI - Chronic and fatal fascioliasis in llamas in the UK. PMID- 10515621 TI - Isolation of Campylobacter sputorum from lesions of papillomatous digital dermatitis in dairy cattle. PMID- 10515622 TI - Serum electrolyte concentrations in bitches with eclampsia. PMID- 10515623 TI - VPHA visit to Spain. Veterinary Public Health Association. PMID- 10515624 TI - Takeover of VETCPD. Veterinary Education Trust for Continuing Professional Development. PMID- 10515625 TI - Breeding and sale of dogs (welfare) act. PMID- 10515626 TI - Avian malaria in gannets. PMID- 10515627 TI - Cat collars PMID- 10515628 TI - Empowerment of women for health promotion: a meta-analysis. AB - The objective of this paper is to identify conditions, factors and methods, which empower women and mothers (WAM) for social action and health promotion movements. WAM are the primary caregivers in almost all cultures; they have demonstrated bold leadership under extreme adversity. Consequently, when empowered and involved, WAM can be effective partners in health promotion programs. The methodology includes a meta-analysis of 40 exemplary case studies from across the world, which meet predetermined criteria, to draw implications for social action and health promotion. Cases were selected from industrialized and less industrialized nations and from four problem domains affecting quality of life and health: (1) human rights, (2) women's equal rights, (3) economic enhancement and (4) health promotion. Content analysis extracted data from all cases on six dimensions: (1) problem, (2) impetus/leadership, (3) macro-environment, (4) methods used, (5) partners/opponents and (6) impact. Analysis identified seven methods frequently used to EMPOWER (acronym): empowerment education and training, media use and advocacy, public education and participation, organizing associations and unions, work training and micro-enterprise, enabling services and support, and rights protection and promotion. Cochran's Q test confirmed significant differences in the frequencies of methods used. The seven EMPOWER methods were used in this order: enabling services, rights protection/promotion, public education, media use/advocacy, and organizing associations/unions, empowerment education, and work training and micro-enterprise. Media and public education were more frequently used by industrialized than non-industrialized societies (X2 tests). While frequencies of methods used varied in all other comparisons, these differences were not statistically significant, suggesting the importance of these methods across problem domains and levels of industrialization. The paper integrates key findings into an empowerment model consisting of five stages: motivation for action, empowerment support, initial individual action, empowerment program, and institutionalization and replication. Implications for policy and health promotion programs are discussed. PMID- 10515629 TI - Women, poverty and common mental disorders in four restructuring societies. AB - BACKGROUND: Poverty and female gender have been found to be associated with depression and anxiety in developed countries. The rationale behind this paper was to bring together five epidemiological data sets from four low to middle income countries to examine whether key economic and development indicators such as income and poor education, and female gender, were associated with common mental disorders. METHOD: The paper is based on five datasets: three based on primary care attenders in Goa, India; Harare, Zimbabwe and Santiago, Chile; and two based on community samples in Pelotas, Brazil and Olinda, Brazil. All five studies estimated prevalence of common mental disorders along with variables to measure economic deprivation and education. FINDINGS: In all five studies, female gender, low education and poverty were strongly associated with common mental disorders. When income was divided into tertiles, with the lowest tertile as a reference value, there was a significant trend for reduced morbidity for the lower two tertiles. DISCUSSION: These findings have considerable implications since the rapid economic changes in all four societies have been associated with rising income disparity and economic inequality. Examples of population based prevention strategies based on increasing the proportion of those who complete schooling and on high-risk strategies such as providing loan facilities to the impoverished are potential outcomes of these findings. Development agencies who focus on women as a priority group have failed to recognize their unique vulnerability to common mental disorders and need to reorient their priorities accordingly. PMID- 10515630 TI - Referral revisited: community financing schemes and emergency transport in rural Africa. AB - Referral between first and second levels of care in rural African health systems is an extremely complex problem. Problems that have plagued the process of referral include poor service quality, low availability of trained personnel, inadequate supplies of drugs and medical diagnostic equipment and inadequate communication infrastructure. In this paper, the authors analyse the role of transport costs in the utilization of referral and how community health insurance schemes can help reduce the economic burden of transport costs, thereby improving referral utilization and health outcomes. Following the introduction, the authors provide a conceptual framework of the individual-, household- and community-level factors that affect referral in the rural African context, with particular emphasis on the role of the time and monetary costs of transport and the potential role of community risk-sharing schemes. The paper then presents a detailed case study from Kenya where a community has been experimenting with a health insurance scheme which provides emergency transport for emergency referral. Data from the past eight years of experience in northern Kenya suggests that support for the insurance scheme has depended on the reliability of the health system, as well as the seasons and various external problems, such as political interference, drought and insecurity. Conclusions drawn support the idea of community financing schemes for transport, not merely as a life-saving strategy in remote and resource-poor health infrastructures, but also as a means to help build trust in the health system itself and thus improve sustainability through local institutional support. PMID- 10515631 TI - Purchasing a quick fix from private pharmacies in the Gaza Strip. AB - Increasingly, it is recognised by health planners and social scientists that self medication with drugs bought over the counter in private pharmacies is extremely widespread. Some anthropologists see this trend as an aspect of the 'commodification of health'. In this study, group interviews with health service users and providers in Gaza revealed many health service users reporting an inadequate supply of drugs resulting in the purchasing of drugs in private pharmacies. As a result, a survey of the pattern of utilization of three private pharmacies in three contrasting urban areas within the Gaza Strip was undertaken. Using a questionnaire, data were collected from all customers buying drugs. The results show that variations in the patterns of health seeking behaviour were associated with socioeconomic status. Adult males were the most frequent customers of all three pharmacies. They were buying medicines for members of their nuclear family more often than for themselves. Overall, pain and influenza were the most commonly reported conditions. The drugs purchased most frequently for women were for reproductive health problems, particularly infertility. Customers of the pharmacy in the relatively prosperous area more commonly purchased drugs which were prescribed by a private doctor. PMID- 10515632 TI - Online commentary in acute medical visits: a method of shaping patient expectations. AB - This paper conceptualizes a type of physician communication, termed 'online commentary'. Online commentary is talk that describes what the physician is seeing, feeling or hearing during physical examination of the patient. Some dimensions of online commentary are described, and its functions in routine and acute medical consultations are distinguished. Using a case study method, the paper focuses on the role of online commentary in pre-empting patient resistance to upcoming 'no problem' diagnostic evaluations which could delegitimize patients' decisions to seek medical assistance, or deprive them of anticipated medical benefits. It is hypothesized that this role for online commentary may be associated with successful physician resistance to implicit or explicit patient demands for inappropriate antibiotic medication. PMID- 10515633 TI - Treatment issues for HIV+ Africans in London. AB - Black Africans are the second largest group of HIV/AIDS service users in London, UK. They are distinguished from other patient groups by their delay in access to services and appear to have a lower uptake of antiretroviral therapies. This study explores the treatment issues concerning black Africans which may affect their uptake of therapies. Issues raised included questions about if and when to start treatment, fears of side-effects both short and long term, awareness of the current uncertainties surrounding combination therapies and concerns about how to achieve compliance. The social circumstances of HIV positive black Africans living in London together with differences in cultural beliefs and experience of health care in the UK give rise to particular treatment concerns. These concerns include the fear of being experimented upon, lack of confidence in drugs tested only on Caucasians, distrust of the medical profession and fears of discrimination. Efforts to encourage the uptake of antiretroviral therapies by black Africans in Britain must take into account the particular experiences, fears and concerns of this patient group. PMID- 10515634 TI - Attitudes to health care prioritisation methods and criteria among nurses, doctors, politicians and the general public. AB - The aim of this postal questionnaire study was to measure attitudes to health care prioritisation criteria among the Finnish general public (n = 1156), politicians (n = 1096), doctors (n = 803) and nurses (n = 667), altogether 3722 subjects. The questionnaire consisted of questions on background data, a list of seven alternative prioritisation methods and a list of 11 possible criteria for health care prioritisation. The most acceptable prioritisation methods were increased treatment fees and restricting expensive treatments and examinations. Only a few supported administrative prioritisation decisions. One third of the general public indicated that they did not accept any limitations in health care, whereas only 5% of doctors agreed with them. More doctors accepted prioritisation methods than respondents in other groups. Patient is a child, patient is an elderly person, severity of the disease and prognosis of the disease were the most accepted prioritisation criteria. Politicians and the general public also accepted self-induced nature of disease and patient's wealth as prioritisation crieteria. Logistic regression analysis of the general public respondents demonstrated that male gender, higher education and higher personal income were associated with acceptance of most prioritisation criteria. Similarly, older age of the respondent was associated with acceptance of self-induced nature of disease and patient's wealth as prioritisation criteria. PMID- 10515635 TI - The psychosocial burden of caring for some Nigerian women with breast cancer and cervical cancer. AB - In Nigeria, the rising incidence of cancer and the paucity of institutional facilities and specialist man-power implies that the burden of care rests largely on relatives. We assessed the severity of indices of psycho-social and economic burden among relatives of women with breast and cervical cancer; and its relationship with patients' psychosocial distress. Using a burden questionnaire, relatives of 73 women with cancer (41 cervical and 32 breast, mean age of caregivers 35.6 years) were interviewed, in out-patient clinics. While the caregivers admitted high frequency of all indices of 'objective' burden, emotional ties at home and social relationships in the neighbourhood seemed intact, indicating tolerance and lack of social stigma. The financial burden was more problematic than the effect of caring on family routines; and these two factors significantly predicted global rating of burden. The severity of patient's worries and psychopathological symptoms were not significantly correlated with care-giver global rating of burden. The tolerance shown by this group of relatives implies that they have strong potentials for playing useful roles in community care of patients. PMID- 10515636 TI - Assessment of ambulance response performance using a geographic information system. AB - The accessibility, distribution and utilisation of emergency medical services are important components of health care delivery. The impact of these services on well-being is heightened by the fact that ambulance resources must respond in a reliable and timely manner to emergency calls from demand areas. However, many factors, such as the unavailability of an ambulance at a center closest to a call, can adversely influence response time. This paper discusses the design and implementation of a framework developed in a Geographic Information System for assessing ambulance response performance. A case study of ambulance response in three communities in Southern Ontario, Canada is presented that allows easy and rapid identification of anomalous calls that may adversely affect overall operating performance evaluation. Extensions of the framework into a fully fledged service deployment and planning decision support system are discussed. PMID- 10515637 TI - Metabolic bone disease in patients with inflammatory bowel disease. PMID- 10515638 TI - Gene transfer as therapy for rheumatoid arthritis: why, what and how? PMID- 10515639 TI - Reliability and validity of the EuroQol in patients with osteoarthritis of the knee. AB - OBJECTIVE: To assess the reliability and validity of the EuroQol (EQ-5D) for osteoarthritis of the knee (OA knee). METHODS: Eighty-two patients with OA knee were asked to complete on two occasions, separated by 1 week, the EQ-5D, the Western Ontario and McMaster Universities (WOMAC) Osteoarthritis Index and the 36 item short form of the Medical Outcomes Study (SF-36). RESULTS: In this patient population, < 10% of the 243 EQ-5D health states were active. The EQ-5D demonstrated a non-Gaussian distribution. Reliability [intraclass correlation coefficient (ICC) = 0.70] is acceptable for aggregate level data. There were significant rank correlations with both the WOMAC and SF-36. CONCLUSIONS: This study provides some evidence of EQ-5D construct validity and reliability. However, the restricted and non-normal distribution of scores, the marked difference between patients' self evaluation and derived societal utility tariffs, as well as the lack of discriminative ability for patients with 'moderate' morbidity within each of the five EQ-5D dimensions, are of concern. PMID- 10515640 TI - Zidovudine in primary Sjogren's syndrome. AB - OBJECTIVE: To evaluate the efficacy of the administration of zidovudine (AZT), an antiretroviral drug, in patients with primary Sjogren's syndrome (SS). METHODS: Seven female patients (age 57 +/- 8.6 yr) with primary SS were enrolled in an open, uncontrolled trial of AZT (250 mg b.i.d.) for the treatment of primary SS. The efficacy variables were oral and ocular dryness symptoms, fatigue, tender points, physician's and patient's global assessments (GA), ocular function tests (fluorescein tear break-up time, Schirmer's test, Rose Bengal staining) and laboratory parameters [erythrocyte sedimentation rate (ESR), serum IgG, IgA and IgM]. RESULTS: A significant improvement was observed in all subjective manifestations, as well as the objective parameters of ocular dryness. The treatment was well tolerated, except for mild and transitory gastrointestinal disturbances in 6/7 patients. Laboratory parameters did not change significantly. The clinical benefit persisted in 5/7 patients 1 month after the end of therapy. CONCLUSION: AZT seems to be effective and well tolerated in patients with primary SS. PMID- 10515641 TI - Liposomal clodronate eliminates synovial macrophages, reduces inflammation and ameliorates joint destruction in antigen-induced arthritis. AB - OBJECTIVES: To investigate the efficacy of a single i.v. dose of clodronate encapsulated within small unilamellar vesicles in suppressing joint inflammation and the histological progression of rat antigen-induced arthritis (AIA). METHODS: Rats with AIA received a single i.v. injection of 20 mg of clodronate encapsulated within small unilamellar vesicles (SUVc) or larger multilamellar vesicles (MLVc) 7 days post-arthritis induction. Free clodronate or saline were used as negative controls. RESULTS: SUVc was shown to be more effective than MLVc, sustaining a significant reduction in knee swelling for up to 7 days after the initial systemic administration. Knee swelling in free clodronate-treated animals was not significantly affected. The increased efficacy of SUVc in reducing inflammation and joint destruction was associated with a significant depletion of resident ED1+, ED2+ and ED3+ macrophages from the synovial membrane (SM). CONCLUSIONS: SUVc is more efficient than MLVc in reducing the severity of inflammation and joint destruction in rat AIA, and is associated with the specific elimination of macrophage subpopulations from the SM. PMID- 10515642 TI - Microvascular abnormalities in Sjogren's syndrome: nailfold capillaroscopy. AB - OBJECTIVE: To describe microvascular abnormalities by nailfold capillaroscopy in patients with primary Sjogren's syndrome (SS) with or without Raynaud's phenomenon (RP) and those with anticentromere antibodies (ACA). METHODS: Forty patients with SS (14 without RP, 16 with RP, 10 with ACA), 20 patients with scleroderma (SSc) (10 with limited and 10 with diffuse disease) (disease control group) and 40 healthy controls (control group) were evaluated by nailfold capillaroscopy. RESULTS: Capillaroscopic abnormalities in SS ranged from non specific findings (crossed capillaries) to more specific findings (confluent haemorrhages and pericapillary haemorrhages) or scleroderma-type findings. SS patients with RP presented capillary abnormalities in higher frequency than patients without RP. The majority of SS patients with ACA (80%) presented scleroderma-type findings. CONCLUSION: Nailfold capillaroscopy can be used as a simple non-invasive method to evaluate the microvascular abnormalities in SS patients, especially in those with RP and those with ACA. PMID- 10515643 TI - Studying patients with inflammatory back pain and arthritis of the lower limbs clinically and by magnetic resonance imaging: many, but not all patients with sacroiliitis have spondyloarthropathy. AB - OBJECTIVE: Clinical and magnetic resonance imaging (MRI) data of 170 consecutive patients with inflammatory back pain (IBP) and/or oligoarthritis of the lower limbs were evaluated in a retrospective study. The aim was to determine the frequency of sacroiliitis and spondyloarthropathy (SpA) in this population, and to assess the significance of HLA B27 measurements for diagnosis in early disease. METHODS: Pelvic X-rays were performed in all IBP patients and dynamic MRI of the sacroiliac joints in patients with IBP who had indefinite results on sacroiliac X-rays (n = 32). RESULTS: European Spondyloarthropathy Study Group criteria for SpA were fulfilled by 106/170 patients (62.4%); eight additional patients had symptoms suggestive of SpA (4.7%). The most frequent SpA subset was undifferentiated SpA (uSpA), diagnosed in 46/106 patients (43.4%). Sacroiliitis was detected by MRI in 21/32 patients with IBP and unclear X-rays (65.6%). Of those, 14 were diagnosed as SpA and seven females with moderate unilateral sacroiliitis, but no features of SpA, also not on follow-up (at least 1 yr), were classified as undifferentiated sacroiliitis (US). Ten of the 14 SpA (71.4%) and none of the seven US patients were HLA B27 positive. CONCLUSION: HLA B27 positivity in IBP patients with MRI-proven sacroiliitis positively predicts SpA. uSpA is a frequent SpA subset. There are HLA B27-negative non-SpA patients with moderate unilateral sacroiliitis whom we propose to be classified as US. PMID- 10515644 TI - Changes in serum chondroitin sulphate epitopes 3-B-3 and 7-D-4 in early rheumatoid arthritis. AB - OBJECTIVES: The aims of the present rheumatoid arthritis (RA) study were (1) to examine the levels of serum 3-B-3 and 7-D-4 to find out whether they are different from controls, (2) to find out whether the concentrations of these epitopes change with disease duration in early RA and (3) whether the serum concentrations of 3-B-3 and 7-D-4 in early RA are prognostic for subsequent disease progression. METHODS: The concentrations of 3-B-3 and 7-D-4 in sera were quantitated by immunoassays. RESULTS: The levels of 3-B-3 and 7-D-4 were significantly lower in RA than in controls (3- to 30-fold, P < 0.001). Changes in 3-B-3 and 7-D-4 were apparent with disease duration. At first presentation, the 3 B-3 concentration was lowest and increased at 12 months (3-fold, P < 0.001). This increase was transient since by 24 and 36 months the concentrations were not different to those at first presentation. The level of 7-D-4 was also lowest when the patients first presented at clinic and increased with time at 6 months (2 fold, P < 0.001). The increase was more prolonged for 7-D-4, remaining elevated at 12, 24 and 36 months. The lack of correlations of serum 3-B-3 and 7-D-4 with clinical measurements showed that these markers were not prognostic for disease severity. CONCLUSIONS: The levels of 3-B-3 and 7-D-4 differed between RA and control sera, and changed with disease duration. These markers were not prognostic in predicting disease outcome. PMID- 10515646 TI - Splenectomy for refractory thrombocytopenia in the antiphospholipid syndrome. AB - OBJECTIVE: Thrombocytopenia, usually mild, is one of the clinical criteria of the antiphospholipid syndrome (APS). Rarely, this disorder requires treatment and, due to the shared characteristics with idiopathic thrombocytopenic purpura (ITP), similar rules are followed. We report our experience in patients who required splenectomy after being refractory to steroids and immunosuppressive therapy. METHODS: Fifty-five APS patients with a platelet count of < 100 x 10(9)/l at least twice were analysed retrospectively. Therapeutic response or remission was considered when the platelet count was > 100 x 10(9)/l after 1 month and with no relapse on stopping or tapering the steroid dose. No response or refractory disease was defined as an absence of increase in platelet count, a total count that never exceeded 50 x 10(9)/l during treatment or when the dose requirements were such that the patient developed serious side-effects. RESULTS: Fifty patients were classified as having secondary APS associated with systemic lupus erythematosus (SLE) and five were identified as primary APS (PAPS). Splenectomy was performed in 11 cases (20%), two PAPS and nine SLE-APS, with an average time of 28 +/- 9 months after the development of thrombocytopenia. Eight patients were initially characterized as ITP (six SLE-APS, two PAPS) with an average time of 4.4 +/- 1.1 yr until the APS diagnosis. All but two were responsive to splenectomy. CONCLUSION: Splenectomy was required in 11 (20%) of the patients with APS-associated thrombocytopenia. There was a high rate of good and long-term response. PMID- 10515645 TI - Relationship between soluble markers of immune activation and bone turnover in post-menopausal women with rheumatoid arthritis. AB - OBJECTIVE: Regarding interactions between pro-inflammatory cytokines and bone metabolism in rheumatoid arthritis (RA), we investigated the relationship between the serum levels of interleukin (IL)-1beta, IL-6, soluble interleukin-2 receptor (sIL-2r), C-reactive protein (CRP), the vitamin D metabolites 25-hydroxyvitamin D3 (25OHD3) and 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], intact parathyroid hormone (iPTH) as well as serum and urinary parameters of bone turnover in 74 post-menopausal women with RA. RESULTS: SIL-2r correlated negatively with 1,25(OH)2D3 (P < 0.01), whereas IL-6 showed a positive correlation with urinary excretion of deoxypyridinoline collagen cross-links (P < 0.01). 1,25(OH)2D3 (P < 0.01) and iPTH (P < 0.01) were negatively related to CRP, whereas the urinary excretion of pyridinoline (P < 0.01) and deoxypyridinoline (P < 0.01)-collagen cross-links showed a positive correlation with CRP. 1,25(OH)2D3 (P < 0.01) and iPTH (P < 0.05) were positively related to bone alkaline phosphatase as a marker of osteoblast function. CONCLUSION: Our data indicate that IL-6 is a critical determinant of increased bone resorption in post-menopausal RA women with high disease activity and that serum levels of 1,25(OH)2D3 are inversely related to T cell activation. PMID- 10515647 TI - Establishment of nurse-like stromal cells from bone marrow of patients with rheumatoid arthritis: indication of characteristic bone marrow microenvironment in patients with rheumatoid arthritis. AB - OBJECTIVE: To investigate the microenvironment of bone marrow (BM) of patients with rheumatoid arthritis (RA). METHODS: Nurse cell-like BM stromal cell lines were established from BM mononuclear cells of patients with RA. We examined the various characteristics of these cell lines, including morphology, pseudoemperipolesis activity, cell surface markers, cytokine production and hyaluronan (HA) production. RESULTS: These RA BM nurse cell-like lines (RA-BMNC) were of mesenchymal origin and positive for CD44, CD54 and HLA-DR. They were defined as nurse cells because of pseudoemperipolesis activity that allowed lymphocytes to migrate underneath. RA-BMNC lines produced HA and multiple cytokines including interleukin (IL)-6, IL-7, IL-8 and granulocyte-macrophage colony-stimulating factor (GM-CSF). HA production by BM stromal cells was correlated with pseudoemperipolesis activity. RA-BMNC produced significantly higher levels of IL-6, IL-8 and GM-CSF by co-culture with lymphocytes. The cells also produced IL-1beta, G-CSF and tumour necrosis factor only when co-cultured with lymphocytes. The RA-BMNC maintained the growth of CD14+ myeloid cells unique to severe RA. CONCLUSION: The present results both indicate that RA-BMNC are nurse cells and suggest that they may play an important role in the pathogenesis of RA. PMID- 10515648 TI - The effect of acupuncture on patients with rheumatoid arthritis: a randomized, placebo-controlled cross-over study. AB - OBJECTIVE: Acupuncture is commonly used by patients with chronic painful musculoskeletal disorders. There are, however, few well-designed studies of its efficacy. This paper describes a randomized placebo-controlled cross-over design to evaluate acupuncture as a useful treatment adjunct in the management of patients with rheumatoid arthritis (RA). METHODS: Sixty-four patients were centrally randomized from a hospital-based rheumatology out-patient clinic. Fifty six patients were suitable for study, all were on second-line therapy and aged 18 75 yr. There had been no change in therapy for the preceding 3 months. Patients who had previous acupuncture, anticoagulation, fear of needles or infection were excluded. Single-point (Liver 3) acupuncture or placebo was given with an intervening 6 week wash-out period. The acupuncturist, patient and statistician were blinded as far as possible. The outcome measures included the inflammatory markers (erythrocyte sedimentation rate and C-reactive protein), visual analogue scale of pain, global patient assessment, 28 swollen and tender joint count, and a general health questionnaire. RESULTS: The results demonstrated no significant effect of treatment or period and no significant interaction between treatment and period for any outcome variable. No adverse effects were reported. CONCLUSION: Acupuncture of this type cannot be considered as a useful adjunct to therapy in patients with RA. Possible reasons why this is the case are discussed. PMID- 10515649 TI - Generic and condition-specific outcome measures for people with osteoarthritis of the knee. AB - OBJECTIVES: The aims of this study were to evaluate two condition-specific and two generic health status questionnaires for measuring health-related quality of life in patients with osteoarthritis (OA) of the knee, and to offer guidance to clinicians and researchers in choosing between them. METHODS: Patients were recruited from two settings: 118 from knee surgery waiting lists and 112 from rheumatology clinics. Four self-completion questionnaires [Western Ontario and McMaster University Osteoarthritis Index (WOMAC), Health Assessment Questionnaire (HAQ), Short Form-36 (SF-36) and Euroqol] were sent to subjects on two occasions 6 months apart. Construct validity, convergent validity, internal consistency and responsiveness were examined using primarily non-parametric methods. RESULTS: All instruments proved satisfactory in terms of ease of use, acceptability to patients, internal consistency and reliability. In the surgical group, the OA specific WOMAC performed better than the HAQ and the generic measures in terms of validity and responsiveness to change, whereas in the rheumatology group the SF 36 was more responsive. CONCLUSION: WOMAC is the instrument of choice for evaluating the outcome of knee replacement surgery in OA. The SF-36 provides a more general insight into patients' health and may be more responsive to change than the WOMAC in a heterogeneous rheumatology clinic population. Researchers wishing to undertake an economic evaluation might consider the EQ-5D for a surgical, but not a rheumatology clinic group. PMID- 10515650 TI - Defining disease activity in ankylosing spondylitis: is a combination of variables (Bath Ankylosing Spondylitis Disease Activity Index) an appropriate instrument? AB - OBJECTIVE: Disease activity has been defined using a self-administered instrument, focusing on fatigue, axial pain, peripheral pain, enthesopathy and morning stiffness [Bath Ankylosing Spondylitis Disease Activity Index (BASDAI)]. This validated instrument is simple and takes 40 s to complete, but whether the index is an accurate expression of the component parts, or whether additional weighting would enhance its efficacy, is unclear. METHODS: Four hundred and seventy-three patients with ankylosing spondylitis received placebo or active non steroidal anti-inflammatory drug (NSAID) for 6 weeks, and changes between entry and completion were captured by BASDAI and the individual components. Principle component analysis (PCA) was used to explore the best combinations of variables in decreasing order of explained total dispersion and to assess whether a single sum (or algebraic expression) best defined disease activity status. RESULTS: At entry, the correlation between BASDAI and the first axis was 0.99, 0.11 with the second, and zero thereafter. Data at study end and relating to change revealed a 100% correlation (R = 1) between the first axis and the sum, with zero for the remainder. CONCLUSIONS: The data support BASDAI as being a valid and appropriate composite to define disease activity in ankylosing spondylitis. Developed as a simple sum of its components, BASDAI has excellent content validity. PMID- 10515651 TI - A Fas promoter polymorphism at position -670 in the enhancer region does not confer susceptibility to Felty's and large granular lymphocyte syndromes. AB - OBJECTIVE: We examined whether there are associations between a polymorphism in the Fas promoter, recently found to be associated with rheumatoid arthritis (RA), and Felty's syndrome or large granular lymphocyte (LGL) leukaemia. METHODS: Thirty-five patients with Felty's were studied, along with 18 patients with LGL syndrome and arthritis, 17 patients with LGL syndrome but no arthritis, and 128 controls. The polymorphism was typed by polymerase chain reaction followed by digestion with the restriction enzyme MvaI. RESULTS: No significant difference was found in genotype or allele frequencies between the groups. CONCLUSION: This promoter polymorphism is not a significant risk factor responsible for the LGL expansions seen in Felty's and LGL syndromes. Abnormal, constitutive expression of Fas ligand may be more relevant to the aetiology of these diseases. PMID- 10515652 TI - Directions in rheumatology: past, present and future. Report of a conference to celebrate the 50th anniversary of the Oliver Bird Fund held at Churchill College Cambridge, 24-26 September 1998. PMID- 10515653 TI - Imaging of pelvic fracture in a patient with psoriatic arthritis. PMID- 10515654 TI - Gorham-Stout disease (phantom bone) of the shoulder girdle. PMID- 10515655 TI - The ARC steroid trial and its effect on clinical practice. PMID- 10515656 TI - Histoplasmosis of the wrist: a case report. PMID- 10515657 TI - Future strategies for corticosteroid therapy. PMID- 10515658 TI - Angio-neurotic oedema associated with methotrexate treatment in rheumatoid arthritis. PMID- 10515659 TI - Endurance training and bone metabolism in middle-aged rats. AB - This study was performed to observe the influence of moderate treadmill running on bone of middle-aged male rats. Seventy 15-month-old Wistar rats were used. Ten initial controls (IC) were killed on day 0. Among the 60 others, three groups of ten exercised rats (E) run 1 h/day, 6 days/week at 60% of their maximum aerobic capacity. On days 30, 60 and 90 of the training period, 20 rats, ten E and ten R (resting animals), were killed. Femoral failure stress never varied and was never different in E and R during the experiment. On day 90 whole body mineral content and mineral density were higher in E than R. Simultaneously, total, diaphyseal and metaphyseal femoral densities were lower in R than IC or than in E. No difference was observed between IC and E. In resting rats, urinary deoxypyridinoline excretion (a marker of bone resorption) increased between days 0 and 90, while it did not change in runners. These results indicate that in middle-aged rats, moderate running prevents decrease in bone mineral density, probably by inhibiting bone resorption. PMID- 10515660 TI - Age-related defects in Th1 and Th2 cytokine production by human T cells can be dissociated from altered frequencies of CD45RA+ and CD45RO+ T cell subsets. AB - The present study investigated whether age-related changes in the production of Th1 and Th2 cytokines by human T cells might be linked to altered frequencies of naive (CD45RA+) and memory (CD45RO+) T cell subsets. T cells from healthy elderly humans (n = 32) stimulated with anti-CD3epsilon monoclonal antibody OKT3 plus PMA produced significantly lower levels of IL-2 and IFNgamma (Th1 type) and of IL-4 (Th2 type) cytokines compared with T cells from young subjects. Although considerable heterogeneity was observed in the levels of cytokines produced by activated T cells from elderly individuals, linear regression analysis failed to demonstrate any significant shift in Th1 to Th2 type cytokine profiles of human T cells during aging. Sufficient T cells were available from eighteen elderly subjects to quantitate the levels of cytokine production in parallel with flow cytometry analysis of the frequencies of CD45RA+ naive and CD45RO+ memory T cells. Compared with the group of young subjects, the elderly group exhibited significant decreases in the frequencies of naive T cells with reciprocal increases in memory T cells. However, defects in Th1 and Th2 cytokine production were not significantly correlated with altered frequencies of naive/memory T cells among elderly individuals. In addition, those elderly individuals with normal frequencies of naive/memory T cells exhibited decreases in cytokine production comparable to the reductions observed for elderly donors with alterations in the frequencies of naive/memory T cells. These findings suggest that age-related defects in Th1 and Th2 cytokine production cannot be attributed entirely to alterations in the frequencies of naive/memory T cell subsets and point toward intrinsic aberrancies within human T cell cytokine networks during aging. PMID- 10515661 TI - Does pH 6 beta-galactosidase activity indicate cell senescence? AB - Apparently two forms of beta-galactosidase (beta-GAL) in cells or tissue sections can be detected by enzyme histochemical staining (X-GAL). Using a sensitive and specific HPLC method we have determined the pH dependent activity of beta-GAL in cell lines of lung carcinoma (A549), colon carcinoma (Caco2-TC7), promyelocytic leukemia (HL60), hepatoma (HepG2) and human liver homogenates. The HPLC method has been validated and the influence of pH and substrate concentration was studied. There was a good linear correlation between HPLC and quantitative enzyme histochemistry (pH 4.5: r = 0.985; pH 6.0: r = 0.967). Both, pH 4.5 beta-GAL and pH 6 beta-GAL could be demonstrated in all biological material tested and pH 6 beta-GAL activity was always lower (25-50%) than pH 4.5 activity. In Caco2-TC7 cells both activities increased by a factor of 10 from day 3 to day 17 after seeding. In addition, since the beta-GAL activity decreased with increase in pH both in human liver homogenates (independent of the age of the donor) as well as in tumor cell lysates in a similar fashion we believe that the activity at pH 6 can hardly be considered as an exclusive 'senescence marker'. In addition, the more sensitive HPLC method could demonstrate activity in cells that showed negative reaction with X-GAL. PMID- 10515662 TI - Comparison of erythropoietic response to androgen in young and old senescence accelerated mice. AB - In this study, to clarify whether the functional capacity of hemopoietic progenitor cells and the micro-environment of aged mice are identical with those of the young, we investigated the changes in the number of hemopoietic progenitor cells and the production of regulatory cytokines from splenic cells as well as changes in the serum levels of cytokine in senescence-accelerated mice (SAM) after administration of 19-nandrolone decanoate (19-ND), a synthetic androgenic anabolic steroid. 19-ND induced an increase in erythroid colony-forming units (CFU-E), erythroid burst-forming units (BFU-E), and granulocytic-macrophage committed progenitor cells (CFU-GM) in bone marrow and spleen; especially remarkable increases were observed in the splenic CFU-E in both young and old mice. Antigen expression analysis of hemopoietic organs revealed that total TER 119+ cells per spleen of young and old mice with androgen treatment rose 2.6- and 3.2-fold over their respective control values. The responsiveness of hemopoietic progenitor cells to androgen did not change with age. Injection of 19-ND into young and old mice markedly enhanced the erythropoietin levels but not IL3 and GM CSF levels in the serum of both groups. Cytokine production assessed by pokeweed mitogen-stimulated spleen condition medium showed an age-related decline. Androgen treatment could not influence IL-3 and GM-CSF production of spleen. These findings suggest that the spleen of both old and young mice served as the major site of regenerative repopulation of hemopoietic progenitors, especially the late erythroid progenitors in 19-ND-treated mice. The proliferative reserve of erythropoiesis with androgen treatment in aged mice was not reduced more than that in treated-young mice. PMID- 10515664 TI - Replicating Huntington's disease phenotype in experimental animals. AB - Huntington's disease (HD) is an inherited, autosomal dominant, neurodegenerative disorder characterized by involuntary choreiform movements, cognitive decline and a progressive neuronal degeneration primarily affecting the striatum. There is at present no effective therapy against this disorder. The gene responsible for the disease (IT15) has been cloned and the molecular defect identified as an expanded polyglutamine tract in the N-terminal region of a protein of unknown function, named huntingtin (The Huntington's Disease Collaborative Research Group, 1993. Cell 72, 971-983). An intense, search for the cell pathology attached to this molecular defect is currently under way [see Sharp and Ross (1996, Neurobiol. Dis. 3, 3-15) for review]. Huntingtin interacts with a number of proteins, some of which have well identified functions, and it has thus been suggested that alterations in glycolysis, vesicle trafficking or apoptosis play a role in the physiopathology of HD. On the other hand data derived from positron emission tomography (PET), magnetic resonance spectroscopy and post-mortem biochemical evidence for a defect in succinate oxidation have suggested the implication of a primary impairment of mitochondrial energy metabolism. All these hypotheses are not necessarily to be opposed and recent findings indicate that the HD mutation could possibly directly alter mitochondrial functions which would in turn activate apoptotic pathways. To test this mitochondrial hypothesis, we studied the effects in rodents and non-human primates of a chronic blockade of succinate oxidation by systemic administration of the mitochondrial toxin 3-nitropropionic acid (3NP). Extensive behavioural and neuropathological evaluations showed that a partial but prolonged energy impairment induced by 3NP is sufficient to replicate most of the clinical and pathophysiological hallmarks of HD, including spontaneous choreiform and dystonic movements, frontal-type cognitive deficits, and progressive heterogeneous striatal degeneration at least partially by apoptosis. 3NP produces the preferential degeneration of the medium-sized spiny GABAergic neurons with a relative sparing of interneurons and afferents, as was observed in HD striatum. The present manuscript reviews the different aspects of this neurotoxic treatment in rodents and non-human primates, and its interest as a phenotypic model of HD to understand the degenerative process of HD and test new therapeutic strategies. PMID- 10515663 TI - Age-dependent changes in DNA polymerase fidelity and proofreading activity during cellular aging. AB - DNA polymerase alpha and the 3'-->5' exonuclease involved in the proofreading of DNA synthesis were isolated from human diploid fetal lung fibroblast (TIG-1) cells at various population doubling levels (PDL). The final PDL of the TIG-1 cells used in these experiments was 70. The fidelity of DNA polymerase alpha remained high until late passage and fell suddenly just before the end of the life span between 65 and 69 PDL. The activities of the 3'-->5' exonuclease related to proofreading remained unchanged from 21 to 61 PDL, but the activity decreased rapidly in more aged cells. The 3'-->5' exonuclease activity at 69 PDL was about 50% of that in TIG cells at 21 PDL. In vitro DNA synthesis by DNA polymerase alpha from TIG-1 cells harvested at 69 PDL showed the amount of non complementary nucleotides incorporated to be decreased by the addition of the 3'- >5' exonuclease from the same cells. However, not all errors were edited out since the ratio of DNA polymerase activity to 3'-->5' exonuclease activity was adjusted to reflect that in vivo and the infidelity of DNA synthesis by error prone DNA polymerase alpha from aged cells was improved by the addition of the highly active 3'-->5' exonuclease from cells at 41 PDL. These results suggested that the mutation frequency rises just before the end of the life span of TIG-1 cells. PMID- 10515665 TI - Neurophysiological support of consciousness during waking and sleep. AB - The aim of this review was to try to establish the current neurophysiological knowledge capable of explaining the differences of mental functioning during the different stages of sleep and waking. The analysis focused on the cortical state. Waking is characterized by electrophysiological activities (low voltage and gamma range EEG field patterns, unitary activities) and cerebral blood flow reflecting an activated state. On the contrary, neurochemical influences are marked by inhibitory afferent processes since dopamine, noradrenaline, serotonin and histamine tend, for the most part, to inhibit cortical neurons by diffuse release at the level of varicosities. During slow wave sleep, all these brain stem influences sustaining the cortical state decrease and transiently disappear just prior to onset of REM sleep. During REM sleep, pontine and mesopontine ascending activating influences invade the cortex in their turn while neurochemical inhibitory influences disappear with the exception of dopaminergic ones. We hypothesize that the activating influences acting on the cortex allow the latter to function, just as petrol makes an engine run, but that the diffuse inhibitory influences somehow regulate cortex functioning. Therefore, it is understandable that, during waking, mental activity is reflective and rational, and that psychological content is less intense during slow wave sleep. During REM sleep, the activated and mostly disinhibited state might induce the characteristic dream activity which appear to be rather ill-considered and illogical. Persistent dopaminergic input combined with the absence of noradrenergic input may induce psychological activities somewhat similar to those related to psychotic syndromes. Deactivation of part of the prefrontal cortex could contribute to this unusual mental activity. PMID- 10515666 TI - From experimentation to the surgical treatment of Parkinson's disease: prelude or suite in basal ganglia research? AB - Parkinson's disease remains one of the greatest challenges facing those who work in the field of neurological research. Although the development of levodopa treatment revolutionised management of this debilitating diseases, no effective long-term therapy has yet been found. With recent advances in the understanding of basal ganglia physiopathology and the experimental demonstration of the efficacy of certain surgical procedures, there is a renewed interest in the surgical approach. This paper provides a chronological overview of the history of parkinsonian surgery and discusses the principal surgical options at our disposal today. These take three main forms: ablation (thalamotomy, pallidotomy and subthalamotomy); cell graft and gene therapy (mainly in the striatum); and deep brain stimulation (of the thalamus, globus pallidus pars internalis and the subthalamic nucleus). Our knowledge of basal ganglia function and our conception of how motor information is processed by this network have evolved parallel to the development of surgical techniques. Recent results from both clinical and experimental studies underline the complexity of the physiopathological mechanisms which generate parkinsonian symptomatology and lead us to question our assumption that each class of clinical signs (tremor, akinesia, rigidity, levodopa-induced dyskinesias...) is produced by a specific and separate mechanism. In the same way, comparison of the electrophysiological and biochemical effects of the different techniques induced in brain function vary considerably. This complex world of interaction and interconnection is a labyrinth that we are still far from comprehending in its entirety. All the more reason, in consequence, for extending experimental investigation into the impact of any new therapy before proposing its clinical application. PMID- 10515667 TI - Octopamine in invertebrates. AB - Octopamine (OA), a biogenic monoamine structurally related to noradrenaline, acts as a neurohormone, a neuromodulator and a neurotransmitter in invertebrates. It is present in relatively high concentrations in neuronal as well as in non neuronal tissues of most invertebrate species studied. It functions as a model for the study of modulation in general. OA modulates almost every physiological process in invertebrates studied so far. Among the targets are peripheral organs, sense organs, and processes within the central nervous system. The known actions of OA in the central nervous system include desensitization of sensory inputs, influence on learning and memory, or regulation of the 'mood' of the animal. Together with tyramine, OA it is the only neuroactive non-peptide transmitter whose physiological role is restricted to invertebrates. This focussed the interest on the corresponding OA receptors. They are believed to be good targets for highly specific insecticides as they are not found in vertebrates. All octopamine receptors belong to the family of G-protein coupled receptors. Four of them could be distinguished using pharmacological tools. They show different coupling to second messenger systems including activation and inhibition of adenylyl cyclase, activation of phospholipase C and coupling to a chloride channel. Recently, octopamine receptors from molluscs and insects have been cloned. Further studies of all aspects of octopaminergic neurotransmission should give deeper insights into modulation of peripheral and sense organs and within the central nervous system in general. PMID- 10515668 TI - Postmenopausal hormone therapy and the risk of breast cancer. PMID- 10515669 TI - HRT, breast and endometrial cancers: strategies and intervention options. AB - The demand for hormone replacement therapy (HRT) by women who enter the menopause is rapidly increasing in all developed countries. The concern that HRT may enhance morbidity and mortality from malignant diseases still limits the widespread adoption of hormonal treatments. Overall, epidemiological data on cancer incidence and HRT are reassuring, although long-term or inappropriate therapies may slightly increase the risk of developing malignant diseases. Many commercial hormonal compounds are currently available and the safest HRT regimen with regard to cancer risk must be identified. It is equally important that the best strategies for breast and endometrial surveillance in women commencing HRT be outlined, bearing in mind that the diffusion of hormonal therapies may be halted by unnecessary medical interventions. PMID- 10515670 TI - Declining socioeconomic differences in the use of menopausal and postmenopausal hormone therapy in Finland. AB - OBJECTIVE: Sales figures for the use of menopausal and postmenopausal hormone therapy in Finland show a rapid increase during the 1980s continued into the first half of the 1990s. Hormone therapy use became very common in Finland compared to many other Western countries. The aim of our study was to investigate the sociodemographic distribution of hormone therapy among Finnish women aged 45 64 years. METHODS: The study is based on population-based surveys conducted in 1989 and 1996 (response rates 87% and 78%). RESULTS: Between 1989 and 1996 the current use of hormone therapy increased from 22% to 27%; in 1989 it was most common in the age group 50-54 years, but in 1996 among 55-59-year-olds. In 1989 it was significantly more common among women with longer education than other women in every age group, but in 1996 this difference was significant only in those 55 years and older. In 1989 the use was more common in the capital area than elsewhere and this difference decreased but remained significant in 1996. CONCLUSION: Our results suggest that hormone therapy has become a routine treatment during the menopause in all educational groups and throughout the country. The lack of socioeconomic differences indicates that among women under 55 year of age the saturation point in short-term hormone use was reached in 1996. However, the persistence of socioeconomic differences among older women suggests that the use of long-term postmenopausal hormone therapy will continue to increase for some time. PMID- 10515671 TI - Beneficial effect of hormone replacement therapy on weight loss in obese menopausal women. AB - OBJECTIVE: At the onset of menopause, weight-gain and the aggravation of certain cardiovascular risk factors are frequently observed. The aim of this study was to examine the metabolic effects of combined hormone replacement therapy (17beta oestradiol transdermic 50 microg for 21 days and oral medroxyprogesterone acetate 5 mg from day 10 to 21) using, in particular, indirect calorimetry. METHODS: Patients (21; 12 substituted and nine controls) were studied twice (3 months apart) during an oral glucose load (75 g). RESULTS: Total body weight was unaltered after 3 months in the control group, whereas a fat-loss of 2.1+/-0.2 kg and a decrease of the waist:hip ratio were observed in the substituted group. In the latter group, a significant increase in lipid oxidation was observed (0.58+/ 0.06 mg/kg/min before and 0.75+/-0.04 mg/kg/min after substitution P<0.05), whilst total energy expenditure and thermogenesis were also increased. Glucose, lipid and protein oxidation remained stable during three months in the control group. The insulin response to an oral glucose load diminished by 30% with hormone replacement therapy (102.3+/-32.8 mmicro/l versus 71.4+/-20.0 mmicro/l). Total and LDL-cholesterol improved after hormone replacement therapy whereas plasma triglycerides were not altered. CONCLUSIONS: Combined hormone replacement therapy not only prevented weight-gain, but favored weight-loss by significantly increasing lipid oxidation after 3 months of treatment. It also favourably influenced the insulin response, plasma lipids and energy expenditure. PMID- 10515672 TI - Histological patterns in endometrial samples from perimenopausal women. AB - OBJECTIVE: Evaluation of endometrial histology patterns in perimenopausal women. METHODS: Endometrial biopsies (202) taken from perimenopausal women by suction curette were assessed by light microscopy. RESULTS: Out of 142 adequate specimens a total of 82 (57.7%) specimens could not be classified in the well-defined categories of the fertile period because of mixed histological patterns. Of the 142 specimens, 59 (41.5%) showed abnormal secretory endometrium, three (2%) disordered proliferative endometrium and 20 (14.1%) a mixture of non-secretory and secretory endometrium. CONCLUSION: The often used histological classification for endometrium, with well-defined regular patterns based on normal cyclic changes, often does not apply to endometrial tissue obtained from perimenopausal women due to a mixed pattern within one biopsy. PMID- 10515673 TI - Adverse endometrial effects during long cycle hormone replacement therapy. Scandinavian Long Cycle Study Group. AB - OBJECTIVES: Treatment with unopposed estrogen is known to increase the risk of endometrial hyperplasia, atypia, and carcinoma, and therefore the administration of a progestin during hormone replacement therapy (HRT) is recommended. The addition of a progestin may cause unwanted side effects. Progestin administration of various durations are therefore used in HRT. STUDY DESIGN: Data were obtained about endometrial histopathology, bleeding interval and compliance in 240 early postmenopausal women receiving HRT with a progestin administered for 10 days during 12 week or 4 week cycles of estrogen administration. These regimens were studied for as long as 4 years. The daily estrogen given was 17beta-estradiol 2 mg per day which was reduced to 1 mg day during the last 6 days of each cycle. The progestin used was norethindrone acetate, given at a dose of 1 mg per day. RESULTS: The incidence of endometrial hyperplastic changes, i.e. simple or complex hyperplasia, atypia or cancer, was significantly higher in the 12 weeks cycle than in the monthly cycle group (P = 0.003), with an overall annual incidence of 5.6% in the 12 weeks cycle group and 1% in the monthly cycle group. One case of atypical hyperplasia and one case of endometrial adenocarcinoma was observed in the long cycle group. Long cycle treatment produced more irregular bleeding pattern. Accordingly, the rate of drop-out due to bleeding was significantly higher in the long cycle group (P<0.01). CONCLUSION: We conclude that the long cycle HRT modality investigated did not improve compliance and may increase the risk of endometrial hyperplasia and eventually cancer compared to conventional HRT with a monthly cycle. Caution using long cycle HRT regimens is advisable, and careful monitoring of the endometrium during treatment is recommended. PMID- 10515674 TI - The prevalence of osteoporosis in the Hong Kong Chinese female population. AB - OBJECTIVES: This paper aims to present population-based age-related bone mass values in the Hong Kong Chinese female population, and to assess the number and proportion of Chinese women considered osteoporotic according to the WHO diagnostic guidelines. METHODS: A total of 769 community-based female subjects were recruited. Social demographic characteristics of these subjects were similar to the Hong Kong general population. All bone mass measurements were performed by means of a dual energy X-ray densitometry (Norland XR 26) at two sites: lumbar vertebrae L2-L4 and left hip. These values were expressed as T-scores, with reference to the mean bone mineral density (BMD) values of the group aged 21-40 years. RESULTS: The study revealed that, in women aged 60 years and above, their mean BMD values are 30% lower than the young normal mean. The prevalence of osteoporosis at the spine increased dramatically from about 10% in the age group 50-59 to 45% in the group aged 60-69. In women aged 70 onwards, over half have osteoporosis at the hip. The prevalence of osteoporosis at the spine is relatively stable in the age groups above 60, while that for osteoporosis at the hip increased exponentially with age. CONCLUSIONS: The prevalence of osteoporosis in Hong Kong women is comparable to that found in Caucasian populations. Prevention of osteoporosis, involving both immediate and long-term measures, and targeting at different age groups, are required to combat this serious public health problem in Hong Kong. PMID- 10515675 TI - Does the presence of postmenopausal symptoms influence susceptibility to vertebral deformity? European Vertebral Osteoporosis Study (EVOS) Group. AB - BACKGROUND: Previous reports suggest a possible increased risk of osteoporosis in those with postmenopausal symptoms. There are, however, no data from population samples, exploring the relationship between postmenopausal symptoms and vertebral osteoporosis. AIM: To determine if there is an association between self-reported postmenopausal symptoms and radiographic vertebral deformity. METHODS: Women aged 50 years and over were recruited from population registers in 30 European centres and invited to attend for an interviewer administered questionnaire and lateral thoracic and lumbar spine radiographs. The questionnaire sought information about aspects of lifestyle, personal, medical and gynaecological history, including postmenopausal symptoms: flushing, sleep disturbance and 'other' symptoms. Radiographs were taken according to a standard protocol and evaluated morphometrically. Vertebral deformity was defined according to the McCloskey Kanis method. Bone mineral density data were obtained in a subsample of women at both the spine and femoral neck. RESULTS: A total of 4023 postmenopausal women, aged 50-64 years, were studied: 73% reported a history of flushing, 45% sleep disturbance and 23% 'other' symptoms, at or around their menopause. The prevalence of vertebral deformity was 8.2%. Those with postmenopausal symptoms were slightly younger and more likely to have ever taken hormone replacement therapy (HRT) than those without symptoms. After adjusting for potential confounders (age, centre, body mass index, cigarette smoking and HRT) there was no association between deformity and any of the postmenopausal symptoms: flushing (odds ratio (OR) 1.0; 95% confidence intervals (CI) 0.8, 1.3), sleep disturbance (OR 1.0; 95% CI 0.8, 1.2), 'other' symptoms (OR 0.9; 95% CI 0.7, 1.3). Amongst women who had ever taken HRT, however, those with vertebral deformity were more likely to report a history of flushing (OR 2.1; 95% CI 0.9,4.8). CONCLUSION: A history of postmenopausal symptoms per se does not appear to be associated with increased susceptibility to vertebral osteoporosis. However, women with more severe symptoms (as suggested by their use of HRT) may be at increased risk. PMID- 10515676 TI - Comparison of transdermal estradiol and tibolone for the treatment of oophorectomized women with deep residual endometriosis. AB - STUDY OBJECTIVE: To compare the effect of HRT with transdermal estradiol and that of treatment with tibolone in post-menopausal women with residual endometriosis. MATERIALS AND METHODS: 21 women with residual pelvic endometriosis after bilateral oophorectomy with or without hysterectomy were enrolled in the study and were randomized to HRT with transdermal estradiol 50 mg twice weekly (n = 10) associated with cyclic medroxyprogesterone acetate 10 mg daily in women who preserved uterus, and to treatment with tibolone 2.5 mg administered orally once a day (n = 11). The duration of both treatments was scheduled to last at least 12 months. Residual endometriosis was located in the bowel wall in four patients, in the rectovaginal septum in six and deeply in the retroperitoneal pelvic space in six. All women were symptomatic before oophorectomy. RESULTS: All the women were followed for 12 months. No patient suspended therapy because of side effects. Four patients of the estradiol group experienced moderate pelvic pain during treatment compared with only one patient in the tibolone group. One patient in the estradiol group reported severe dyspareunia. CONCLUSION: Although our series is very small, it seems that tibolone may be a safe hormonal treatment for post menopausal women with residual endometriosis. PMID- 10515677 TI - Hormone replacement therapy in perimenopausal women and 2-year change of carotid intima-media thickness. AB - OBJECTIVES: To assess the 2-year effects of a combined regimen of oral 17beta estradiol and desogestrel (17betaE-D) and a sequential combination of conjugated equine estrogens and norgestrel (CEE-N) on common carotid intima-media thickness and end-diastolic lumen diameter in comparison to placebo in perimenopausal women. METHODS: The study was a single center, randomized, group-comparative, double-blind study with respect to the 17betaE-D and placebo groups and open with respect to CEE-N. After cycle 6, the blind was broken and the trial was continued as an open trial for another 18 months for the active study arms. The study included 121 perimenopausal women recruited from the general population. Common carotid intima-media thickness and end-diastolic lumen diameter were measured at baseline and cycle 24 with B-mode ultrasonography. RESULTS: At cycle 24 small changes in intima-media thickness and lumen diameter were observed. Relative to placebo, changes in intima-media thickness were -0.009 mm [95% CI -0.045; 0.027] for 17betaE-D and -0.016 mm [95% CI -0.055; 0.024] for CEE-N. For end-diastolic lumen diameter the changes were -0.091 mm [95% CI -0.236; 0.055] and -0.125 mm [95% CI -0.820; 0.032] for 17betaE-D and CEE-N, respectively. CONCLUSIONS: In this study among perimenopausal women a significant effect of 17betaE-D and CEE-N on common carotid intima-media thickness and lumen diameter could not be demonstrated. Although the sample size of the present trial is too limited to provide definite conclusions, the direction of the effect is in agreement with evidence from earlier studies on the effects of hormone replacement therapy in postmenopausal women. PMID- 10515678 TI - Gender differences in the recognition of depression in old age. AB - OBJECTIVE: The study should answer the question of whether identical symptom presentations of depression in male and female patients leads to similar recognition rates in primary care. METHOD: We performed a survey in primary care. Two written case vignettes were presented to 170 family physicians in a face-to face interview which took place in their practices. The case vignettes described either a mildly depressed otherwise healthy old patient (case 1) or a severely depressed patient with somatic comorbidity (case 2). For each case different versions with regard to patients' gender were used: in case 1 only the gender of the patient varied; in case 2 both the gender and the anamnesis (stroke/hypothyroidism) varied. Afterwards the interviewers asked standardised open questions. The physicians were not aware of the mental health focus and the gender focus of the study. RESULTS: The study is representative with a response rate of 77.6%. For primary diagnosis, the female versions were given the diagnosis of depression more often. There was a non-significant trend that female physicians considered depression more often. CONCLUSION: The results show that gender-related experience and stereotypes on the physicians' side influence the diagnosis of (old age) depression in primary care. Further studies should elucidate the influence of the physicians' gender on the management of psychiatric disorders. PMID- 10515679 TI - CD30L-ETA': a new recombinant immunotoxin based on the CD30 ligand for possible use against human lymphoma. AB - Recombinant DNA technology makes it possible to genetically fuse V genes or cytokines to toxin domains, resulting in immunotherapeutics for selective destruction of tumor cells. Since recombinant immunotoxins can be easily manipulated in terms of affinity or cytotoxic potency and produced in large quantities, we have developed a new CD30 ligand-based fusion toxin (CD30L-ETA'). Human CD30L cDNA was ligated into a pET-based expression plasmid and thereby fused to a modified Pseudomonas aeruginosa exotoxin A (ETA') lacking its cell binding domain I. After IPTG-indiced expression in E. coli strain BL21(DE3), the 60 kDa His-tagged fusion protein (CD30L-ETA') was isolated from inclusion bodies. Denatured protein was renatured in the presence of 0.4 M arginine and a glutathione redox system. Refolded protein was purified and concentrated by ion exchange chromatography on a HiTrap Q column. The binding properties of CD30L ETA' were evaluated by competitive ELISA, immunohistochemical staining, and FACS analysis on CD30-expressing cells. The in vitro toxicity of the fusion protein was then tested on the CD30+ Hodgkin-derived cell line L540cy and the Burkitt's lymphoma cell line BL38. CD30L-ETA' exhibited specific cytotoxicity against L540cy cells (IC50 = 24 ng/ml) as determined by [3H]leucine uptake assays. This is the first report on the specificity and cytotoxic potency of a chimeric CD30L fusion toxin against Hodgkin's disease-derived cells. PMID- 10515680 TI - Molecular analysis of resistance to interferon in patients with laryngeal papillomatosis. AB - Although interferon (IFN)-alpha has been used successfully as an adjuvant therapy in laryngeal papillomatosis, some patients are resistant to this treatment. In order to know which patients will benefit from the therapy, we have tried to find a relationship between the IFN response and the viral and host parameters in the lesion. Detection of viral type and copy numbers by polymerase chain reaction (PCR) showed that all patients infected with human papillomavirus (HPV)-11 were sensitive to the treatment, in contrast to those infected with HPV-6. These differences could be explained in part by the inability of HPV-11 E7 to inhibit the induction of an IFN-responsive element, whereas HPV-6 E7 almost completely inhibited the activity of this promoter in transient transfection experiments. Local immune status in the lesion showed that all HPV-11-infected patients had detectable levels of interleukin (IL)-15 and IFN-gamma mRNA, in contrast to HPV-6 infected patients, in whom mRNA for these cytokines was almost absent. Viral copy numbers and levels of IL-4 mRNA could not be correlated with IFN response. Only one patient resistant to recombinant IFN-alpha2b and negative for HPV DNA presented high titers of neutralizing anti-IFN-alpha2b antibodies. This patient became sensitive when natural IFN-alpha was administered. These results suggest that response to IFN may be a complex phenomenon resulting from the interaction between viral and host elements. PMID- 10515681 TI - Interleukin-3 in hematology and oncology: current state of knowledge and future directions. AB - Interleukin (IL)-3 is a multipotent hematopoietic growth factor produced by activated T cells, monocytes/macrophages and stroma cells. The human IL-3 gene is located on chromosome 5 near segment 5q31. The high-affinity receptor for human IL-3 is composed of alpha and beta subunits. IL-3 shares a common beta subunit with granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-5; this subunit has been mapped to chromosome 22q13.1. The biological effects of IL-3 have been studied in human and murine hematopoietic cell lines and normal human marrow cells. Addition of IL-3 to the culture medium induces proliferation, maturation and probably self-renewal of pluripotent hematopoietic stem cells and cells of myeloid, erythroid and megakaryocytic lineages. Human IL-3 was cloned in 1986, and since then various clinical trials have assessed the in vivo potential of recombinant human (rhIL-3). Initial results of phase I/II studies of IL-3 at a dose of 5-10 microg/kg subcutaneously daily for 5-10 days in patients with relapsed lymphomas, small-cell lung cancer, breast cancer and ovarian cancer showed that post-chemotherapy application of IL-3 reduces chemotherapy delays and induces faster regeneration of granulocytes and platelets. However, these results were not confirmed in phase III studies. The role of IL-3 alone in the treatment of myelodysplastic syndromes (MDS), aplastic anemia (AA) and other bone marrow failure disorders have also been disappointing. However, preliminary studies of IL-3 in combination with chemotherapeutic agents and immunosuppression have demonstrated encouraging results in patients with MDS and AA respectively. The therapeutic potential of IL-3 in peripheral blood stem cell (PBSC) harvesting and priming of stem cells before harvest is beginning to be identified. Initial results of IL-3 combination with GM-CSF or later-acting growth factors such as granulocyte colony-stimulating factor (G-CSF) have yielded larger amounts of PBSC during harvesting. In recent years, the availability of synthetic IL-3 receptor (IL-3R) agonists and similar chimeric molecules with greater in vitro biological activity and fewer inflammatory side-effects has extended our options to employ and compare these molecules and rhIL-3 for the prevention of chemotherapy-induced myelosuppression. The role of IL-3 and IL-3R agonists in ex vivo expansion of stem cells, dendritic cell development and gene transfer requires further evaluation. It appears that future application of IL-3 in combination with other cytokines is an attractive way forward in the prevention of treatment-related mortality and morbidity in oncology patients. It also shows prospects for the development of new therapeutic strategies for dose escalation and immune modulation for cancer patients with relapsed and resistant disease. PMID- 10515682 TI - The therapeutic potential of erythropoietin receptor transgenes. AB - Allogeneic bone marrow transplantation is an effective curative therapy for both malignant and heritable diseases. The use of genetically altered autologous hematopoietic stem cells (HSC) is being increasingly investigated as a treatment for a variety of non-malignant but significantly morbid diseases, including hemoglobinopathies, immunodeficiencies and autoimmune diseases. Other hematopoietic cells capable of proliferation, such as antigen-specific T cells and dendritic cells, have also been used for adoptive immunotherapy. Genetic procedures to modify these various therapeutic cells so that they can be selectively amplified either in vitro or in vivo could enhance their efficacy. For example, HSC that contain a gene that confers a survival, selection or growth advantage may enhance their engraftment. Such enhancement could be expected to reduce graft failures and the intensity of the required conditioning regimen, thereby decreasing the toxicities of transplantation. In this review, the functions of cytokine receptor transgenes coding for erythropoietin receptors (EpoR) are analyzed. The characteristics of these transgenic cells and animals are discussed with regard to the possible therapeutic use of EpoR transgenes in the transplantation of hematopoietic cells. PMID- 10515683 TI - Adoptive T-cell immunotherapy of cancer. AB - Adoptive T-cell therapy involves the passive transfer of antigen-reactive T cells to a tumor-bearing host in order to initiate tumor rejection. Based upon animal models, effector T cells with tumor-specific reactivity are superior to non specific effector T cells in mediating tumor regression in vivo. Both CD4+ and CD8+ T cells are capable of initiating tumor rejection after adoptive transfer. Several different culture methods have been reported that permit in vitro expansion of immune T cells while retaining tumor specificity. The ability to generate human tumor-specific effector T cells capable of mediating tumor rejection in vivo has provided tools to identify tumor-associated antigens. Future directions in this field involve the selective isolation and expansion of subpopulations of T cells critical to initiating tumor rejection, and the use of molecular techniques to generate effector T cells. PMID- 10515684 TI - Generation of autologous immunity to acute myeloid leukaemia and maintenance of complete remission following interferon-alpha treatment. AB - Interferon-alpha (IFN-alpha) is established as part of the treatment for chronic myeloid leukaemia, although its precise mode of action remains largely unknown. Its use in acute myeloid leukaemia (AML) has been limited. We have previously documented autologous cytolytic activity against AML blasts in patients after autologous bone marrow transplantation. Here we present a patient with poor-risk AML who relapsed from first complete remission (CR) and was unwilling to undergo high-dose chemotherapy with stem cell rescue. In second chemotherapy-induced CR, the patient had no evidence of antileukaemia cytolytic activity in an in vitro assay, and she commenced IFN-alpha (Roferon). She subsequently developed high levels of leukaemia-specific cytotoxicity, and has remained in second CR for two years. These findings support the use of IFN-alpha in patients with poor-risk AML, and suggest that one mechanism of action may be immunological. PMID- 10515685 TI - A re-evaluation of the molecular mass of earthworm extracellular hemoglobin from meniscus depletion sedimentation equilibrium. Nature of the 10 S dissociation species. AB - Previous calculations from meniscus depletion sedimentation equilibrium earthworm hemoglobin from Lumbricus terrestris (E.J. Wood et al., Biochem. J. 153 (1976) 589-96) and from the related species Lumbricus sp. (L. sp.) (M.M. David and E. D Mol. Biol. 87 (1974) 89--101) were made on the assumption that the solutions behaved ideally. Re-examination of their results reveals, however, a dependence of the apparent molecular mass on concentration. Taking this effect into consideration, we have nowrecalculated from their data molecular masses of 4.4- 4.5 MDa for the hemoglobin of both L. terrestris and L. sp. On the basis of the new determinations, we propose for the polypeptide chain composition of L. terrestris hemoglobin a model [(abcd )4L1L2L3]12 where a,b,c,d are the four globin and L1,L2,L3 are the three major linker chain constituents of the protein. The model is consistent with the D6 symmetry of the molecule. A 10 S intermediate product in the alkaline dissociation Lumbricus hemoglobin is viewed as a binary mixture of products resulting from a disproportionation reaction involving the structural unit. The present interpretation is shown to be consistent with observed relations between molecular masses and SDS gel electrophoretic band patterns of 10 S species and intact hemoglobin. PMID- 10515686 TI - NMR study of the binding of all-trans-retinoic acid to type II human cellular retinoic acid binding protein. AB - Cellular RA binding proteins are thought to play important roles in the (RA), a hormonally active metabolite of vitamin A that has profound effects on cell growth, + differentiation and morphogenesis. Binding of RA to type II human cellular RA binding proteins (CRABPII) has been investigated by NMR spectroscopy. The sequential resonance assignments of +CRABPII in the presence of RA were established by heteronuclear three-dimensional NMR at pH 7.3. The resonance assignments of the bound RA were achieved by homonucl NMR. The secondary structures of holo-CRABPII determined by NMR were ess as revealed by the crystal structure of holo-CRABPII. Most of the nuclear Overhauser effects (NOEs) between CRABPII and the bound RA were consistent with those predicted crystal structure of holo-CRABPII. The results suggested that the conformations in solution and in the crystalline state are highly similar. Compared to the ligand binding pocket, especially the ligand entrance, was stabilize Ser12-Leu18, one of the structure elements that constitute the ligand binding pocket, became more mobile upon binding of RA. Intramolecular NOEs between protons of the bo the carboxylate end of the bound RA is well fixed but the β-ionone PMID- 10515687 TI - Binding studies of tear lipocalin: the role of the conserved tryptophan in maintaining structure, stability and ligand affinity. AB - The principal lipid binding protein in tears, tear lipocalin (TL), binds acid and the fluorescent fatty acid analogs, DAUDA and 16-AP at one site TL compete for this binding site. A fluorescent competitive binding assay revealed that apo-TL has a high affinity for phospholipids and stearic acid (Ki) of 1.2 microM and 1.3 microM, respectively, and much less affinity for cholesterol (Ki) of 15.9 of the hydrocarbon chain. TL binds most strongly the least soluble lipids permitting these lipids to exceed their maximum solubility in aqueous solution. These data implicate TL in solubilizing and transporting lipids in the tear film. Phenylalanine, tyrosine and cysteine+ were substituted for TRP 17, the only invariant residue throughout the lipocalin superfamily. Cysteine substitution resulted in some loss os secondary structure, relaxation of aromatic side chain rigidity, decreased binding affinity for DAUDA and destabilization of structure. Mutants of TL, W17Y, and W17F showed a higher binding affinity for DAUDA than wild-type TL. Comparison of the results of the tryptophan 17 substitution in lipocalin with those of tryptophan 19 substitution in beta-lactoglobulin revealed important differences in binding characteristics that reflect the functional heterogeneity within the lipocalin family. PMID- 10515688 TI - [Modification of nutritional septic risk in perioperative transfusion]. PMID- 10515689 TI - The spatial properties of a model neuron increase its coding range. AB - The coding properties of one-compartment and two-compartment model neurons are compared. The membrane depolarization in both models is described as a deterministic leaky integrator. Interspike intervals are identified with the periods between reset of the depolarization after firing and consecutive crossing of a fixed firing threshold. The two-point model has an input in the dendritic compartment and an output in the trigger-zone compartment. it is shown that the sensitivity threshold for the two-point model is shifted to the larger values of the input intensity with respect to the sensitivity threshold of its single-point counterpart. Further, its coding range is substantially larger than the coding range of the single-point model. PMID- 10515690 TI - Right-left discrimination in a biologically oriented model of the cockroach escape system. AB - We present a biologically oriented model that accounts for left-right discrimination in the cockroach's escape behavior. The model includes the main groups of neurons found to be involved in the escape response. Each one of the included neurons is described by the actual processes taking place in an individual neuron (formation of an action potential, transmitter release, conductance changes, etc.). Furthermore, realistic chemical synapses (excitatory or inhibitory and able to undergo different types of modulation) connect the various neurons. With this model, we ere able to achieve, for a wide range of inputs representing different wind directions, behavior which resembles that found experimentally. The model indicates that several synaptic properties, in particular postsynaptic inhibition and presynaptic facilitation, play a key role in the discrimination of wind direction. PMID- 10515691 TI - Identification of plasticizers in medical products by a combined direct thermodesorption--cooled injection system and gas chromatography--mass spectrometry. AB - The combination of a new thermodesorption module with a cooled injection system now provides a powerful system for direct analysis of volatile trace compounds in gaseous, liquid and solid samples by gas chromatography-mass spectrometry (GC MS). As a cooled injection system is used for the cryofocusing of the desorbed volatiles the GC-MC system still can be used for the regular analysis of liquid samples. Although plasticizers usually are analyzed by GC-MS after solvent extraction, contaminated solvents and glassware are very well known problems. Analysis of plasticizers in plastic materials by direct thermodesorption instead saves time and avoids cross contaminations. Many medical products are made of plasticized polyvinyl chloride. Extraction of the common plasticizer di(2 ethylhexyl) phthalate (DEHP) into blood will occur, and harmful effects of DEHP in the human body have been suggested. We therefore analyzed 21 different plastic devices which are used for various invasive techniques in medicine by direct thermodesorption GC-MS. In some of the plastics up to 30 different components were identified. By far the most common plasticizer found was DEHP, followed by diethyl and dibutyl phthalates. PMID- 10515692 TI - Commotio cordis: early observation. PMID- 10515694 TI - Effect of repetitive episodes of exercise induced myocardial ischaemia on left ventricular function in patients with chronic stable angina. PMID- 10515693 TI - Self administration of metolzaone reduces readmissions with decompensated congestive cardiac failure. PMID- 10515695 TI - Beta-blockers and heart failure. PMID- 10515696 TI - Spreading the word. PMID- 10515697 TI - Tile automation: a model for an architecture of a living system. AB - To understand an architecture of a living system, "Tile Automaton" is introduced as an abstract model of chemical reaction of molecules scattered over a space. The model consists of tiles of various shapes that stand for molecules. The chemical reaction, induced by the collisions of tiles, is represented by the change of the tile shapes. The rules for reaction are deterministic, and the evolution of the system strongly depends on mutual spatial relationship among tiles. The evolution often leads to self-organization of a "factory," a set of tiles that produces tiles continuously and keeps its structure. Several interesting phenomena, such as a deformation or a division of a factory, are also observed. It is proposed that the formation of the factory is due to the interference between different aspects of tiles - the shape and the motion. The concept of "entanglement" is introduced as a mechanism of living systems. PMID- 10515698 TI - Polyhydroxylated pyrrolidine and pyrrolizidine alkaloids from Hyancinthoides non scripta and Scilla campanulata. AB - Aqueous ethanol extracts from the immature fruits and stalks of bluebell (Hyacinthoides non-scripta) were subjected to various ion-exchange column chromatographic steps to give 1,4-dideoxy-1,4-imino-D-arabinitol (1),2(R),5(R) bis(hydroxymethyl)-3(R),4(R)-dihydroxypyrrolidine (DMDP) (2), 6-deoxy-6-C-(2,5 dihydroxyhexyl)-DMDP (3),2,5-dideoxy-2,5-imino-DL-glycero-D-manno-heptitol (homoDMDP)(4),homoDMDP-7-O-apioside (5), homoDMDP-7-O-beta-D-xylopyranoside (6), (1S*,2R*,3R*,5R*,7aR*)-1,2-dihydroxy-3,5- dihydroxymethylpyrrolizidine (7), and (1S*,2R*,3R*,5R*,6R*,7R*,7aR*)-3-hydroxymethyl-5-methyl-1,2,6,7 tetrahydroxypyrrolizidine (8). Bulbs of Scilla campanulata (Hyacinthaceae) yielded (1S*,2R*,3R*,5S*,7aR*)-1,2-dihydroxy-3,5-dihydroxy-methylpyrrol izidine (9) in addition to compounds 1-7. Compounds 3,6,7,8, and 9 are new natural products. Compound 4 is a potent competitive inhibitor with K(i) values of 1.5 microM for Caldocellum saccharolyticum beta-glucosidase and 2.2 microM for bovine liver beta-galactosidase. The 7-O-beta-D xyloside 6 was a stronger competitive inhibitor than 4 of C saccharolyticum beta-glucosidase and rat intestinal lactase, with K(i) values of 0.06 and 0.07 microM, respectively, but a weaker inhibitor of bovine liver beta-galactosidase. Furthermore, compound 4 is also a competitive inhibitor (K(i) = 1.8 microM) of porcine kidney trehalase, but 6 was inactive against this enzyme. PMID- 10515699 TI - [Pelvic radiation with concurrent chemotherapy compared with pelvi and para aortic radiation for high-risk cervical cancer. Concurrent cisplatin-based radiotherapy and chemotherapy for locally advanced cervical cancer. Cisplatin, radiation, and adjuvant hysterectomy for bulky stage Ib cervical carcinoma]. PMID- 10515700 TI - Frontal dementias. Etiological, clinical, therapeutical and pathological aspects. Preface. PMID- 10515701 TI - E-publishing on the Web: promises, pitfalls, and payoffs for bioinformatics. PMID- 10515702 TI - Addiction and mental health. PMID- 10515703 TI - Clearance of apoptotic thymocytes is decreased by inhibitors of eicosanoid synthesis. PMID- 10515704 TI - A novel assay for apoptotic body formation and membrane release during apoptosis. PMID- 10515705 TI - Use of controlled randomized trials to evaluate new technologies and new operative procedures in surgery. PMID- 10515706 TI - Useful understanding of postoperative atrial fibrillation. PMID- 10515707 TI - Comments on "Estrogenicity of resin-based composites and sealants used in dentistry". PMID- 10515709 TI - Kids swallowing pennies. PMID- 10515710 TI - Allergy receptor pictured. PMID- 10515708 TI - Error in DEHP background concentration. PMID- 10515711 TI - Safer drugs for children. PMID- 10515712 TI - Analysis of breast milk to assess exposure to chlorinated contaminants in Kazakhstan: sources of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposures in an agricultural region of southern Kazakhstan. AB - High levels of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; up to 208 pg/g fat) were measured in samples of breast milk collected in 1997 from 64 donors [41 first-time mothers (primiparae)] living on state farms in southern Kazakhstan. TCDD was the major contributor (70%) to the toxic equivalents, matching the congener patterns found in breast milk and serum samples collected in 1994 and 1996 from donors in nearby villages. The highest TCDD levels were found in state farms adjacent to a reservoir (zone A), which receives agricultural runoff from cotton fields. TCDD levels in zone A were significantly higher than levels in a region more distant (zone B; > 10 miles) from the reservoir (zone A: mean 53 pg/g, n = 17; zone B: mean 21 pg/g, n = 24; p = 0.0017). Levels of TCDD in breast milk and animal-derived foodstuffs were 10 times U.S. levels. Body burden and dietary data suggest that exposures to TCDD are chronic, environmental, and long term and may be related to the use of chemicals in cotton agriculture. The data suggest that the most likely source is the use of cotton defoliants contaminated with TCDD, and the most likely pathway for human exposure is via the consumption of contaminated foodstuffs. PMID- 10515713 TI - Sentinel lymph node detection and imaging. PMID- 10515714 TI - The value of renal scintigraphy during controlled diuresis. PMID- 10515715 TI - Impressions and clinical consequences of a multidisciplinary congress: neurology and nuclear medicine. 10th seminar of Serre Chevalier, France, 24-30 January, 1999 under the auspices of the French Society of Biophysics and Nuclear Medicine. PMID- 10515716 TI - Serotonin 5-HT1A receptor imaging in the human brain with PET. Co-ordination of the standardisation and dissemination of methodology. Workshop on [carbonyl 11C]WAY-100635 radiochemistry and metabolite analysis 30-31 March 1999. MRC Cyclotron Unit, London, UK. PMID- 10515717 TI - Workshop on 5-HT1A receptor imaging--modelling of [carbonyl-11C]WAY-100635 in the human brain. 6 May 1999, PET Centre, San Raffaele, Milan, Italy. PMID- 10515718 TI - 13th Meeting of the International Research Group on Immunoscintigraphy and Immunotherapy (IRIST). 7-8 May 1999, Gottingen, Germany. PMID- 10515719 TI - Is colonoscopy indicated for small adenomas found by screening flexible sigmoidoscopy? PMID- 10515720 TI - Differences in the diagnostic criteria used by Japanese and Western pathologists to diagnose colorectal carcinoma. PMID- 10515721 TI - Endoscopic resection fo large sessile colorectal polyps using a submucosal saline injection technique. PMID- 10515722 TI - [History of the development of public hygiene in Russia]. PMID- 10515724 TI - Recent publications in hematologic oncology. PMID- 10515723 TI - [Impact of damaged genes: effects on replication and transcription]. PMID- 10515725 TI - TRendys in Biochemistry-98. PMID- 10515727 TI - [The new tax status of private health insurance plans and the Spanish National Health System]. PMID- 10515726 TI - [Prevention levels of occupational risks]. PMID- 10515728 TI - [Group health insurance and tax deductions]. PMID- 10515729 TI - [Presentation: Medline in Internet]. PMID- 10515730 TI - [Omission of variables in regression models with high multi-colinearity]. PMID- 10515731 TI - [Assumption of maximal indetermination, the sample size and other considerations relating to the sample]. PMID- 10515733 TI - National commission to target economics of veterinary profession. PMID- 10515732 TI - Imports banned in wake of Belgian food scandal. PMID- 10515734 TI - Allocation of attention in dichotic listening: effects on the detection and localization of targets within lists. AB - In each of two dichotic listening experiments, 48 normal right-handed adults were instructed to attend selectively to the left and right ears and to divide attention equally between ears. Participants listened for specified targets and reported the ear of entry when the target was heard. Stimuli consisted of lists of digit names in Experiment I and lists of words in Experiment 2. Shifts of attention altered ear asymmetry for localizing but not for detecting digit names. For words, attention shifts altered both detection asymmetry and localization asymmetry, but the effect of attention on detection seemed to reflect differential retrieval from short-term memory rather than differential perception. In both experiments, shifting attention toward either ear resulted in a reporting bias such that signals were attributed to the attended ear more often than to the unattended ear. The results confirm our previous findings, for single pairs of stimuli, that volitional shifts of attention alter response selection rather than perception. PMID- 10515735 TI - Latent structure of the Children's Category Test at two age levels in the standardization sample. AB - The latent structure of the Children's Category Test (CCT; Boll, 1993) was evaluated in the standardization sample, with separate principal factor analyses for the younger subgroup (ages 5-8 years; CCT-1) and the older subgroup (ages 9 16 years; CCT-2). At both age levels, a two-factor solution was found. It is concluded that the CCT is a multifactorial instrument, and that clinical interpretation should consider the diversity of dimensions assessed by this test. PMID- 10515736 TI - Further considerations of null hypothesis testing. PMID- 10515737 TI - Considerations in evaluation of the applicability of DNA fingerprinting techniques for species differentiation. PMID- 10515738 TI - Detection of Epstein-Barr virus DNA in plasma from patients with lymphoproliferative disease after allogeneic bone marrow or peripheral blood stem cell transplantation. PMID- 10515739 TI - Comparison of the MB/BacT and BACTEC 460 TB systems. PMID- 10515741 TI - Abolition of swarming of Proteus. PMID- 10515740 TI - Rapid PCR-based detection of Streptococcus pneumoniae DNA in cerebrospinal fluid. PMID- 10515742 TI - The 1999 ASCI Presidential Address: Executive Summary of the Nerflex Commission Report PMID- 10515743 TI - The 1999 ASCI Award. PMID- 10515744 TI - Acceptance of the Kober Medal for 1999. PMID- 10515745 TI - [A continuing medical education committee at the Physicians' Order... Why and how?]. PMID- 10515746 TI - British guidelines on hypertension management issued. PMID- 10515747 TI - Innovations in teaching anatomy. PMID- 10515748 TI - Communication skills: how do we teach them to medical students? PMID- 10515749 TI - Symmetrical erythematous butterfly rash. PMID- 10515750 TI - Liposomal amphotericin B for fever and neutropenia. PMID- 10515751 TI - Liposomal amphotericin B for fever and neutropenia. PMID- 10515752 TI - Liposomal amphotericin B for fever and neutropenia. PMID- 10515753 TI - Liposomal amphotericin B for fever and neutropenia. PMID- 10515754 TI - Liposomal amphotericin B for fever and neutropenia. PMID- 10515756 TI - Use of alternative medicine by women with breast cancer. PMID- 10515755 TI - Liposomal amphotericin B for fever and neutropenia. PMID- 10515757 TI - Use of alternative medicine by women with breast cancer. PMID- 10515758 TI - Use of alternative medicine by women with breast cancer. PMID- 10515759 TI - Use of alternative medicine by women with breast cancer. PMID- 10515760 TI - Use of alternative medicine by women with breast cancer. PMID- 10515761 TI - Use of alternative medicine by women with breast cancer. PMID- 10515762 TI - Use of alternative medicine by women with breast cancer. PMID- 10515763 TI - Lack of efficacy of transdermal nicotine in smoking cessation. PMID- 10515764 TI - Reduction of serum testosterone in men by licorice. PMID- 10515765 TI - Commentary on Roebuck DJ: "Risk and benefit in paediatric radiology". PMID- 10515766 TI - Toxic effects of indoor molds. PMID- 10515767 TI - When is there enough evidence to stop worrying about pertussis vaccine? PMID- 10515768 TI - Read this--it's important. PMID- 10515769 TI - Observer bias in acellular pertussis vaccine trials. PMID- 10515770 TI - Too little water intake causing nephrolithiasis, revealed by too much water! PMID- 10515771 TI - Myoclonic movements in very low birth weight premature infants associated with midazolam intravenous bolus administration. PMID- 10515772 TI - Comparison of Canadian education students' and teachers' estimates of the prevalence of learning 'disabilities'. PMID- 10515773 TI - Localized form of dyschromatosis universalis hereditaria in a 14-year-old girl. PMID- 10515775 TI - Interferon for tufted angioma. PMID- 10515774 TI - Zinc aspartate, biotin, and clobetasol propionate in the treatment of alopecia areata in childhood. PMID- 10515776 TI - Epilepsies: from new concepts to new images. Proceedings of the International Meeting of the French Society of Neurology. Paris, 4-5 June 1998. PMID- 10515777 TI - [Excessive watery diarrhea and pronounced fatigue due to Cyclospora cayetanensis infection in an HIV infected traveler returning from the tropics]. AB - We report the case of a 46-year-old HIV-1 infected patient who acquired a Cyclopsora cayetanensis infection during travel to southeast Asia. He developed excessive watery diarrhoea and pronounced fatigue, which resolved after treatment with cotrimoxazole. The term Cyclospora cayetanensis was first suggested in 1993 for the infectious agent of diarrhoea in tropical and subtropical regions. The infection is usually self-limiting. However, in HIV-1 infected persons the symptoms are often more severe and protracted. Microbiological diagnosis is performed by light microscopy and at X400 magnification of faeces fixed in SAF medium or 10% formalin preservatives. PMID- 10515778 TI - [A dental assistant for the 3rd millennium]. PMID- 10515779 TI - [The influence of developments in Europe on dentistry]. PMID- 10515780 TI - [A tooth puller and his crew. Constantin Guys]. PMID- 10515781 TI - [Education of the patient]. PMID- 10515782 TI - [The fluoride content in children's toothpastes]. PMID- 10515783 TI - FDA weighs using tumor cell lines for vaccine development. PMID- 10515784 TI - Wellcome seeks new home for business park. PMID- 10515785 TI - Fending off furtive strategists. PMID- 10515786 TI - Kansas evolution ruling. PMID- 10515787 TI - NIH-study-section scoring. PMID- 10515788 TI - Reducing natural disasters. PMID- 10515789 TI - Regulation of "nutraceuticals". PMID- 10515790 TI - Beating the odds with Big K. PMID- 10515791 TI - How serpins are shaping up. PMID- 10515792 TI - Complex grabbing. PMID- 10515793 TI - In memoriam, Per Wallen, 1927-1999. PMID- 10515794 TI - Calcium homeostasis in dystrophic muscle. PMID- 10515795 TI - COPD: new developments and therapeutic opportunities. PMID- 10515796 TI - [Laparoscopic surgery in the treatment of perforating gastroduodenal ulcers]. PMID- 10515798 TI - Epidemiology of acquisition and clearance of cervical human papillomavirus infection in women from a high-risk area for cervical cancer. AB - Acquisition and clearance of cervical human papillomavirus (HPV) infection were analyzed among 1425 low-income women attending a maternal and child health program in Sao Paulo, Brazil. Specimens collected every 4 months were tested by a polymerase chain reaction protocol (MY09/11). In all, 357 subjects were positive at least once. There were 1.3% new infections per month, with 38% cumulative positivity after 18 months. Of 177 positive subjects at enrollment, only 35% remained infected after 12 months. The monthly clearance rate was higher for nononcogenic types (12.2%; 95% confidence interval [CI], 9.6-15.4) than for oncogenic HPV infections (9.5%; 95% CI, 7.5-11.9). Median retention times were 8.1 months (95% CI, 7.8-8.3) for oncogenic types and 4.8 months (95% CI, 3.9-5.6) for nononcogenic HPV infections. The mean infection durations were 8.2 and 13.5 months for nononcogenic and oncogenic types, respectively. Although a woman's age did not affect mean duration for oncogenic types (13-14 months), nononcogenic type infections lasted longer (10. 2 months) among younger (<35 years old) than in older women (5.6 months). PMID- 10515797 TI - Cyclic-imide-hydrolyzing activity of D-hydantoinase from Blastobacter sp. strain A17p-4. AB - The cyclic-imide-hydrolyzing activity of a prokaryotic cyclic-ureide-hydrolyzing enzyme, D-hydantoinase, was investigated. The enzyme hydrolyzed cyclic imides with bulky substituents such as 2-methylsuccinimide, 2-phenylsuccinimide, phthalimide, and 3,4-pyridine dicarboximide to the corresponding half-amides. However, simple cyclic imides without substituents, which are substrates of imidase (ie.g., succinimide, glutarimide, and sulfur-containing cyclic imides such as 2,4-thiazolidinedione and rhodanine), were not hydrolyzed. The combined catalytic actions of bacterial D-hydantoinase and imidase can cover the function of a single mammalian enzyme, dihydropyrimidinase. Prokaryotic D-hydantoinase also catalyzed the dehyrative cyclization of the half-amide phthalamidic acid to the corresponding cyclic imide, phthalimide. The reversible hydrolysis of cyclic imides shown by prokaryotic D-hydantoinase suggested that, in addition to pyrimidine metabolism, it may also function in cyclic-imide metabolism. PMID- 10515799 TI - Seroreactivity to human papillomavirus types 16, 18, 31, and 45 virus-like particles in a case-control study of cervical squamous intraepithelial lesions. AB - Serum IgG antibodies to human papillomavirus (HPV) types 16, 18, 31, and 45 virus like particles were measured in a nested case-control study of cervical squamous intraepithelial lesions. HPV-16 seroreactivity was strongly associated with HPV 16 DNA detection (odds ratio, 9.0; 95% confidence interval, 4.4-19.4), and similar type specificity was observed for HPV-31 and -45. In contrast, seroreactivity to any type was associated with elevated seroreactivity to all others. Among cases and controls, HPV-16 showed the highest seroprevalence, with 23.8% of 80 cases and 10.5% of 258 controls seroreactive to HPV-16 alone, and another 27.5% and 5.4%, respectively, seroreactive to HPV-16 plus other types. Overall, 24 (30.0%) cases and 17 (6.6%) controls were seroreactive to multiple types. These data suggest that seroreactivity to a given type reflects mainly type-specific HPV infection as measured by DNA detection and may also signal past exposure to other types that are now only serologically detected. PMID- 10515800 TI - Early CD69 expression on peripheral blood lymphocytes from children with dengue hemorrhagic fever. AB - Recent reports have demonstrated immune activation in dengue hemorrhagic fever (DHF) by cytokine and soluble receptor detection in blood. The goal of this study was to determine which cell types are activated and likely to be responsible for cytokine production. Whole blood specimens from 51 Thai children presenting within 72 h of fever onset and with detectable plasma dengue viral RNA were studied by flow cytometry. Absolute CD4 T cell, CD8 T cell, NK cell, and gammadelta T cell counts were decreased in children with DHF compared with those with dengue fever (DF) early in the course of illness. The percent of cells expressing CD69 was increased on CD8 T cells and NK cells in children who developed DHF more than in those with DF. These data directly demonstrate that cellular immune activation is present early in acute dengue and is related to disease severity. PMID- 10515801 TI - O'nyong-nyong fever in south-central Uganda, 1996-1997: description of the epidemic and results of a household-based seroprevalence survey. AB - O'nyong-nyong (ONN) fever, an acute, nonfatal illness characterized by polyarthralgia, is caused by infection with a mosquito-borne central African alphavirus. During 1996-1997, south-central Uganda experienced the second ONN fever epidemic ever recognized. During January and early February 1997, active case-finding and a household cluster serosurvey were conducted in two affected and two comparison areas. A confirmed case was defined as an acute febrile illness with polyarthralgia occurring within the previous 9 months plus serologic confirmation or isolation of ONN virus from blood. In affected (n=129) and comparison (n=115) areas, the estimated infection rates were 45% and 3%, respectively, and the estimated attack rates were 29% and 0%, respectively, for an apparent:inapparent infection ratio of nearly 2 in affected areas. In villages sampled near Lake Kijanebalola, Rakai District, the estimated infection and attack rates were 68% and 41%, respectively, and 55% of sampled households had >/=1 case of ONN fever. In conclusion, this epidemic was focused near lakes and swamps, where it was associated with high infection and attack rates. PMID- 10515802 TI - Prevalence and significance of naturally occurring mutations in the surface and polymerase genes of hepatitis B virus. AB - The prevalence and clinical significance of naturally occurring mutations in the full-length surface and overlapping polymerase genes of hepatitis B virus (HBV) were analyzed in 42 patients with chronic hepatitis. Mutations were observed in 10 patients (24%) in the a determinant region, which is the neutralizing epitope within the major hydrophilic region of the surface gene. A high proportion of these mutations (17/18; 94%) occurred in the first loop, unlike mutations induced by immunization. The presence of serum antibody to hepatitis B surface antigen was significantly associated with these mutations. No other region of the surface gene contained any cluster of mutations. These results suggest that escape mutations commonly contribute to persistency in the natural course of HBV infection. In contrast, mutations affecting the major catalytic domains of the polymerase gene, which could alter susceptibility to antiviral nucleoside analogues, were not detected at all. PMID- 10515803 TI - Associations between cellular immune effector function, iron metabolism, and disease activity in patients with chronic hepatitis C virus infection. AB - We studied the associations of macrophage activity, T-helper cell types 1 and 2 (Th-1/Th-2) responses, and iron status in 55 patients with hepatitis C virus (HCV)-related liver disease and 28 control patients with noninfectious liver disease. Serum concentrations of soluble tumor necrosis factor receptor type II (sTNFrec 75), a macrophage activation marker, were higher in cirrhotic than in noncirrhotic patients (P<.001) regardless of their HCV status, whereas levels of neopterin, interleukin (IL)-4 and IL-10 did not differ significantly. In HCV positive patients, sTNFrec 75 levels and transferrin saturation (TfS) correlated positively with levels of aspartate transaminase (P<.001 for sTNFrec 75 and P=.028 for TfS) and alanine transaminase (P=.003 for sTNFrec 75 and P=.039 for TfS). Increased TfS correlated significantly with both advanced liver disease and a predominant Th-2 pattern in HCV patients. Our data suggest that an association exists between macrophage activation and hepatic dysfunction, and that iron status may affect the clinical course of HCV infection by modulating Th-1/Th-2 responses in vivo. PMID- 10515804 TI - Prevalence and incidence of herpes simplex virus type 2 infection among male Zimbabwean factory workers. AB - Stored sera from a cohort of 2397 male factory workers in Harare, Zimbabwe, were screened for herpes simplex virus type 2 (HSV-2)-specific antibodies, to estimate the prevalence and incidence of genital herpes infection and to assess the relation between HSV-2 and human immunodeficiency virus (HIV) acquisition. The prevalence of HSV-2 at enrollment was 39.8%. Correlates of HSV-2 seropositivity were HIV seropositivity, marital status, history of sexually transmitted disease (STD), older age, and higher income. The incidence of HSV-2 seroconversion during follow-up was 6.2/100 person-years. Correlates of HSV-2 seroconversion were enrollment while HIV-positive or seroconversion during follow-up, reported genital ulcer, history of STD, and number of sex partners. No evidence was found that HSV-2 infection was more likely to precede HIV or vice versa. HSV-2 and HIV seropositivity are strong markers for high-risk sexual behavior. Improved interventions targeted to populations in which the incidence of either viral infection is high are needed. PMID- 10515805 TI - Molecular polymorphism of Kaposi's sarcoma-associated herpesvirus (Human herpesvirus 8) latent nuclear antigen: evidence for a large repertoire of viral genotypes and dual infection with different viral genotypes. AB - Molecular polymorphism was found in Kaposi's sarcoma-associated herpesvirus (KSHV) latent nuclear antigen (LNA), mapped to the internal repeat domain of the encoding orf73 gene, and used to develop a novel genotyping technique, KSHV LNA genotyping (KVNAtyping). KVNAtype was stable during latent and lytic viral replication in cell culture and in humans. Diverse KVNAtypes were identified in 43 specimens: 6 KSHV cell lines and 6 Kaposi's sarcoma (KS) and 4 primary effusion lymphoma (PEL) tumor samples from the United States, 15 KS tumor samples from Italy, and 12 KS tumor samples from Zambia. A single KVNAtype was detected in each of 41 specimens, and 2 KVNAtypes were detected in each of 2 KS specimens. Multifocal KS from 3 patients showed the same single KVNAtype at all sites in each patient. These results demonstrate a large repertoire of KSHV genotypes and suggest that the development of most KSs and PELs is associated with a single viral genotype. PMID- 10515806 TI - Human interleukin-12 enhances interferon-gamma-producing influenza-specific memory CD8+ cytotoxic T lymphocytes. AB - Interferon (IFN)-gamma synthesis of CD45RO+ (memory) and CD45RA+ (naive) CD8+ cytotoxic T lymphocytes (CTLs) and the role of interleukin (IL)-12 in the regulation of human CTL functions in virus-specific immunity were investigated. After culture with influenza virus, CD45RO+ CD8+ T cells from human peripheral blood mononuclear cells increased in frequency and exhibited significant major histocompatibility complex class I-mediated CTL activity, whereas CD45RA+ CD8+ T cells did not. Influenza virus-stimulated CD45RO+ CD8+ T cells contained significantly higher levels of IFN-gamma-producing cells and IFN-gamma-specific mRNA than did CD45RA+ CD8+ T cells. Recombinant human IL-12 further enhanced CTL activity and IFN-gamma production by CD45RO+ CD8+ T cells. These data clearly show that human virus-specific CTL activity and coproduction of IFN-gamma are associated with the CD45RO+ CD8+ T cells that are modulated by the cell-mediated, immunity-inducible cytokine IL-12 in humans. PMID- 10515808 TI - Co-receptor usage of BOB/GPR15 in addition to CCR5 has no significant effect on replication of simian immunodeficiency virus in vivo. AB - Human immunodeficiency virus type 2 (HIV-2) and the closely related simian immunodeficiency viruses (SIVs) frequently use the orphan receptor BOB/GPR15 in addition to the chemokine receptor CCR5 for efficient entry and replication. However, the role of BOB/GPR15 in replication and pathogenesis of HIV-2 and SIV in vivo is unclear. This study shows that a single amino acid substitution in the V3 loop of the pathogenic SIVmac239 clone, 321P-->S, impaired the ability to use BOB/GPR15 for entry and replication but had little effect on the ability to use CCR5. This envelope variant replicated with an efficiency comparable with the parental SIVmac239 isolate in rhesus macaques. Furthermore, the mutant genotype and phenotype remained stable even after the onset of immunodeficiency. These results suggest that this cofactor plays only a minor role for the pathogenicity of the HIV-2/SIVmac/SIVsm group of primate lentiviruses. PMID- 10515807 TI - Quantitative proviral DNA and antibody levels in the natural history of HTLV-I infection. AB - The pathogenesis of human T-cell lymphotropic virus type I (HTLV-I) in adult T cell leukemia/lymphoma (ATL) and HTLV-I associated myelopathy/tropical spastic paraparesis (HAM/TSP) is poorly understood. We prospectively followed up and evaluated the virologic correlates of infection in transfusion recipients after seroconversion, in asymptomatic carriers, and in ATL and HAM/TSP patients. Proviral DNA levels (copies/105 lymphocytes) were determined by real-time automated polymerase chain reaction and antibody titers by end-point dilution by use of an HTLV-I enzyme-linked immunoassay. In early infection, proviral load was initially elevated (median, 212 copies/105 lymphocytes at time 1) and later decreased (median, 99 copies at time 2, and 27 copies at time 3). Corresponding antibody titers were low at time 1 (1:2154), had significantly increased by time 2 (1:12312), and were stable by time 3 (1:4694). These viral markers were significantly lower in asymptomatic carriers than in HAM/TSP or ATL patients. Therefore, proviral load and antibody titers may be useful as predictive markers of disease among carriers. PMID- 10515809 TI - Progressive expansion of an L-selectin-negative CD8 cell with anti-feline immunodeficiency virus (FIV) suppressor function in the circulation of FIV infected cats. AB - The acute stage of feline immunodeficiency virus (FIV) infection is characterized by the appearance of a major CD8 subpopulation with reduced expression of the CD8 beta chain (CD8alpha+betalo). CD8 antiviral activity was subsequently shown to be mediated by the CD8alpha+betalo phenotype, which is the dominant CD8 phenotype in long-term infected cats. Two- and three-color flow cytometric analysis demonstrated that the CD8alpha+betalo subset is L-selectin negative (CD62L-) and has increased expression of CD44, CD49d, and CD18, consistent with an activation phenotype. The CD8alpha+betaloCD62L- cells but not the CD8alpha+betahiCD62L+ cells demonstrated strong antiviral activity in the FIV acute-infection assay. The progressive expansion of the CD8alpha+betaloCD62L- effector subset cells in FIV-infected cats parallels that seen in human immunodeficiency virus (HIV) infected patients, suggesting that failure in homeostatic mechanisms regulating lymphocyte activation or trafficking (or both) may be a consequence of both HIV and FIV infections. PMID- 10515810 TI - Evaluation of human immunodeficiency virus (HIV) type 1 load, CD4 T cell level, and clinical class as time-fixed and time-varying markers of disease progression in HIV-1-infected children. AB - Human immunodeficiency virus (HIV) type 1 RNA load, CD4 T cell level, and Centers for Disease Control and Prevention (CDC) clinical class history were measured as potential correlates of a CDC class C diagnosis or death in 165 HIV-1-infected children followed from birth. These covariates were assessed at fixed "landmark" ages from 6 to 24 months and were also assessed as time-varying values. Virus load was associated with progression in all analyses, even after adjusting for immunologic and clinical status. This confirms its importance for monitoring pediatric disease progression. CD4 T cell level was associated with disease progression in time-varying but not in adjusted landmark analysis, suggesting that CD4 cells reflects immediate risk more than long-term risk. The distinction between clinical class B and lower classes is prognostic during the first 18 months of life; class C versus classes N/A/B becomes more important as the patient ages. Virologic, immunologic, and clinical status all provide information regarding disease progression risk. PMID- 10515811 TI - Variants of the human NRAMP1 gene and altered human immunodeficiency virus infection susceptibility. AB - In a population-based case-control study, 182 human immunodeficiency virus (HIV) positive persons and 135 healthy control subjects were enrolled from the metropolitan area of Medellin, Colombia. Four genotypes of the natural resistance associated macrophage protein l (NRAMP1) gene (5' GT repeat, 274C/T, 469+14G/T, and 823C/T) were associated with altered risk of HIV infection (P=.013,.015,.020, and. 035, respectively). Three of these markers (5' [GT]n, 274C/T, 469+14G/T) are in strong linkage disequilibrium, and genotypes of these markers are associated with reduced risk of HIV infection with relative risks (RRs) of 0.35 (95% confidence interval [CI], 0.14-0.91), 0.31 (CI, 0.10-0.93), and 0.24 (CI, 0.08 0.72), respectively. Conversely, heterozygosity at the fourth independent marker (823C/T) was associated with increased risk of HIV infection (RR, 2.29; CI, 1.06 4.92). These findings suggest that NRAMP1 modifies risk of HIV infection. PMID- 10515812 TI - Quantitative and qualitative differences in the serum antibody profiles of human immunodeficiency virus-infected persons with and without Cryptococcus neoformans meningitis. AB - The importance of humoral immunity for resistance to Cryptococcus neoformans is uncertain. A case-controlled study of the human antibody response to C. neoformans comparing the serum antibody profiles of human immunodeficiency virus (HIV)-infected persons who did (HIV+/CM+) or did not (HIV-infected controls) develop cryptococcal meningitis (CM) and HIV-uninfected persons with samples obtained from the Multicenter AIDS Cohort Study was performed. Total immunoglobulin concentrations were determined, and the specificity, isotype, and idiotype expression of antibodies to C. neoformans capsular glucuronoxylomannan were analyzed by ELISA. Compared with the HIV+/CM+ group, the HIV-infected control group had significantly lower levels of total IgM, IgA, and antibodies expressing a certain VH3 determinant. The HIV-infected control group manifested an increase in immunoglobulin levels with a decrease in CD4 lymphocytes. The findings suggest a possible association between reduced expression of certain immunoglobulin subsets and HIV-associated CM. PMID- 10515813 TI - Pharmacokinetics of didanosine in antepartum and postpartum human immunodeficiency virus--infected pregnant women and their neonates: an AIDS clinical trials group study. AB - Didanosine (ddI) pharmacokinetics in antepartum and postpartum human immunodeficiency virus (HIV)-infected women and their neonates were studied. HIV infected pregnant women received an intravenous (iv) ddI infusion (1.6 mg/kg/h) or an oral dose (200 mg bid or 125 mg bid) at 31 weeks antepartum and 6 weeks postpartum. Blood samples were obtained regularly up to 6 or 8 h after drug administration. The same oral dose of ddI (bid) was administered until labor began. Then, ddI was infused iv until delivery. An oral pharmacokinetic study (60 mg/m2) was conducted in infants at day 1 and at week 6 after birth. Plasma concentrations of ddI were measured by radioimmunoassay. After iv ddI administration, only the maternal plasma clearance was found to be significantly increased antepartum (1028+/-231 mL/min) versus postpartum (707+/-213 mL/min). No pharmacokinetic parameters after oral administration were significantly affected by pregnancy. The pharmacokinetics of ddI in the neonates were highly variable. We conclude that the oral ddI dose need not be adjusted during pregnancy. PMID- 10515814 TI - Neutrophils from patients with advanced human immunodeficiency virus infection have impaired complement receptor function and preserved Fcgamma receptor function. AB - Interleukin (IL)-8 production by human polymorphonuclear leukocytes (PMNL) to Cryptococcus neoformans is related to complement activation. Generation of the bioactive fragments C3a and C5a is responsible for IL-8 release. IL-8 production was analyzed in response to C. neoformans by PMNL from persons with early- and late-stage (>400 and <200 CD4 cells/mm3, respectively) human immunodeficiency virus (HIV) infection who were at high risk for cryptococcosis. IL-8 release by PMNL from persons with early-stage infection and from healthy donors was similar; however, PMNL from persons with late-stage HIV infection had significantly impaired IL-8 production, which correlated with reduced IL-8 response to C3a and C5a proteins and decreased CD88 expression. Addition of murine monoclonal antibody (MAb) 18B7 promoted phagocytosis and restored IL-8 release consistent with integrity of FcgammaRIII. These results provide evidence for a selective defect in CD88 expression on PMNL from persons with late-stage HIV infection. However, Fcgamma receptor expression in PMNL appears to be intact and allows MAb to glucuronoxylomannan to positively influence PMNL function. PMID- 10515815 TI - Neutrophil A2A adenosine receptor inhibits inflammation in a rat model of meningitis: synergy with the type IV phosphodiesterase inhibitor, rolipram. AB - Bacterial meningitis is a disease worsened by neutrophil-induced damage in the subarachnoid space. In this study, the A2A adenosine receptors on human neutrophils were characterized, and the role of A2A receptors on the trafficking of leukocytes to the cerebrospinal fluid and on blood-brain barrier permeability (BBBP) was assessed in a rat meningitis model. Neutrophils bind the A2A selective antagonist, 125I-ZM241385 (Bmax=843 receptors/neutrophil; KD=0.125 nM). A selective A2A receptor agonist, WRC-0470 (2-cyclohexylmethylidene hydrazinoadenosine; 0.03-1 microM), alone and synergistically with the type IV phosphodiesterase inhibitor, rolipram, increased neutrophil [cAMP]i and reduced cytokine-enhanced neutrophil adherence, superoxide release, and degranulation. These effects of WRC-0470 were reversed by ZM241385 (100 nM). In a lipopolysaccharide-induced rat meningitis model, WRC-0470 (0-0.9 microgram/kg/h), with or without rolipram (0-0.01 microgram/kg/h), inhibited pleocytosis and reduced the lipopolysaccharide-induced increase in BBBP, indicative of decreased neutrophil-induced damage. PMID- 10515816 TI - Influence of in vitro susceptibility phenotype against thrombin-induced platelet microbicidal protein on treatment and prophylaxis outcomes of experimental Staphylococcus aureus endocarditis. AB - Thrombin-induced platelet microbicidal protein-1 (tPMP-1) is a small, cationic staphylocidal peptide from rabbit platelets. In the current study, the outcomes of vancomycin treatment and prophylaxis were compared in experimental infective endocarditis (IE) caused by an isogenic Staphylococcus aureus strain pair differing in tPMP-1 susceptibility (tPMPS) or resistance (tPMPR) in vitro (ISP479C and ISP479R, respectively). Vancomycin therapy (selected for its intrinsically slow bactericidal activity) reduced ISP479C (but not ISP479R) densities in vegetations compared with controls (P<.01). In contrast, prophylactic administration of vancomycin yielded no differences in efficacies for the 2 challenge strains. These data suggest that the tPMPR phenotype in vitro has a negative effect on the antimicrobial therapy (but not the prophylaxis) of experimental S. aureus IE. These disparate results may be explained in part by the requirement for microbicidal effects in the treatment of established IE, whereas prophylactic efficacy depends more on growth inhibitory and antiadhesion effects. PMID- 10515817 TI - Immunity to cross-reactive serotypes induced by pneumococcal conjugate vaccines in infants. AB - Infants were immunized with 1 of the 3 experimental pneumococcal conjugate vaccines that contain 6B and 19F but not 6A or 19A serotypes. Their sera were studied for the capacity to opsonize Streptococcus pneumoniae 6A, 6B, 19A, and 19F serotypes and the level of IgG antibody to the 4 serotypes. Significant increases were observed in the number of infants with detectable opsonophagocytic titers with 3 conjugate vaccines for 6B (vaccine) serotype but with only 2 vaccines for 6A (cross-reactive) serotype. Significant increases were observed with 2 conjugate vaccines for 19F serotype but with only 1 vaccine for 19A serotype. Thus, some conjugate vaccines may elicit cross-protection better than others. In addition, correlations between opsonophagocytic titers and IgG antibody levels by ELISA were high for 6B and 19F serotypes but low for 6A and 19A serotypes. Thus, ELISA may be an inadequate surrogate assay of vaccine response for cross-reactive serotypes. PMID- 10515819 TI - Relationship between plasma levels of lipopolysaccharide (LPS) and LPS-binding protein in patients with severe sepsis and septic shock. AB - Plasma endotoxin and lipopolysaccharide-binding protein (LBP) levels were measured in a group of 253 patients at the onset of severe sepsis and/or septic shock. Endotoxin levels were significantly greater than control levels (n=33; mean +/- SD, 5.1+/-7.3 pg/mL) in 78.3% of patients. Median endotoxin levels in patients with sepsis were 300 pg/mL (25%-75% interquartile range, 110-726 pg/mL). LBP levels were elevated in 97% of patients compared with normal control values of 4.1+/-1.65 microgram/mL. Median LBP levels in patients with sepsis were 31.2 microgram/mL (interquartile range, 22.5-47.7 microgram/mL). Median endotoxin levels at study entry were more highly elevated (515 vs. 230 pg/mL; P<.01), and LBP levels were less highly elevated (28.0 vs. 33.2 microgram/mL; P<.05) in nonsurvivors than survivors over the 28-day study period. No correlation was found between endotoxin and LBP levels. The quantitative level of both endotoxin and LBP may have prognostic significance in patients with severe sepsis. PMID- 10515818 TI - Characterization of a multidrug-resistant clone of invasive Streptococcus pneumoniae serotype 6B in Alaska by using pulsed-field gel electrophoresis and PspA serotyping. AB - Antimicrobial susceptibility, pneumococcal surface protein A (PspA) serotyping, and pulsed-field gel electrophoresis (PFGE) were used to evaluate clonal relatedness among 66 invasive isolates of Streptococcus pneumoniae serotype 6B collected during 1982-1996 from patients in Alaska. Thirty-seven (56%) of the isolates had penicillin minimal inhibitory concentration values >/=0.125 microgram/mL and were resistant to at least 1 other antibiotic. Fourteen PspA serotypes were observed; PspA 16 was the most common (35%). Forty-five (68%) of the 66 isolates shared common and highly related PFGE patterns using 3 enzymes. Twenty-six (58%) of the isolates with common PFGE patterns were from Native Alaskan children /=1:128 during CTX therapy in 12 episodes and were measurable in 7 patients after the last dose. All patients were cured. The present study provides scientific rationale to the clinical studies that suggest treating SBP episodes with lower doses of antibiotics and shorter treatment duration. PMID- 10515822 TI - Neutrophil chemotaxis on silicone and polyurethane surfaces. AB - Silicone vascular catheters have a greater risk of infection and produce greater inflammation in vivo and greater complement activation in vitro than other vascular catheter polymer materials. This study investigated whether polymorphonuclear leukocyte (PMNL) chemotaxis under agarose on silicone surfaces is different than on polyurethane (PU). Glass slides were coated with silicone and PU by use of a constant-speed dipping apparatus. Chemotaxis (3 h) in response to (10-7 mL) FMLP, zymosan-activated serum, and fresh serum (100%) was greater on silicone than on PU (P<.05). Polyclonal antibody to C5a blocked >50% of the movement toward serum (P<.05). Serum in the PMNL well significantly decreased chemotaxis toward FMLP on silicone (P<.05) but not on PU. These findings suggest that excessive complement activation by silicone may interfere with chemotaxis, but further work is necessary to determine whether this is relevant to an increased risk of catheter-related infection. PMID- 10515823 TI - Transmission of a multidrug-resistant Mycobacterium tuberculosis strain resembling "strain W" among noninstitutionalized, human immunodeficiency virus seronegative patients. AB - Since 1990, several outbreaks of multidrug-resistant tuberculosis (MDR-TB) have been described among institutionalized patients infected with human immunodeficiency virus (HIV). We describe a community MDR-TB outbreak among HIV seronegative patients in Cape Town, South Africa. Isolates were characterized by restriction fragment length polymorphism (RFLP) analysis and dot-blot hybridization analysis of mutations conferring resistance for isoniazid, rifampin, streptomycin, and ethambutol. All isolates were identical on RFLP analysis. In 2 patients, RFLP analysis showed exogenous reinfection during or after treatment for drug-susceptible TB. Mutation analysis confirmed the genotypic identity of the isolates. The infecting strain was genotypically related to strain W, which is responsible for the majority of MDR-TB outbreaks in New York City. Transmission of MDR-TB is thus not limited to HIV-seropositive patients in an institutional setting but occurs within a community. PMID- 10515824 TI - Mycobacterium-induced transmesothelial migration of monocytes into pleural space: role of intercellular adhesion molecule-1 in tuberculous pleurisy. AB - The pleural mesothelium is a dynamic cellular membrane with multiple key functions. It plays a pivotal role in pleural inflammation through its release of several cytokines and the expression of cell-surface molecules. The expression of intercellular adhesion molecule (ICAM)-1 in the pleural mesothelium of patients with active pleural tuberculosis and the role of ICAM-1 in monocyte transmigration across pleural mesothelium during tuberculous inflammation was investigated. Results indicate pleural mesothelial cells (PMCs) express ICAM-1 in tuberculous pleuritis. When PMCs were stimulated with bacille Calmette-Guerin (BCG) in vitro, they expressed ICAM-1 in a time-dependent manner. Monocyte transmigration was higher across PMC monolayers that had been stimulated with BCG. Blocking ICAM-1 on BCG-activated PMC monolayers inhibited monocyte transmigration against chemotactic gradient generated by macrophage inflammatory protein 1-alpha or monocyte chemotactic protein-1. These results indicate that ICAM-1 expression in PMCs facilitates monocyte transmigration during tuberculous pleural inflammation. PMID- 10515825 TI - High prevalence and incidence of sexually transmitted diseases in urban adolescent females despite moderate risk behaviors. AB - To better understand the prevalence, incidence, and risk factors for sexually transmitted diseases (STDs) among female adolescents, a prospective 6-month cohort study was conducted at four teen clinics in a southeastern city. At enrollment, 260 (40%) of 650 sexually active females ages 14-19 years had an STD: chlamydia, 27%; herpes simplex virus type 2 (HSV-2), 14%; gonorrhea, 6%; trichomoniasis, 3%; and hepatitis B, 2%. At follow-up, 112 (23%) of 501 participants had an incident infection: chlamydia, 18%; HSV-2, 4%; gonorrhea, 4%; and trichomoniasis, 3%. At either enrollment or follow-up, 53% had >/=1 STD; of those with 1 lifetime partner, 30% had an STD. Having a new partner (odds ratio [OR], 2.2; 95% confidence interval [CI], 1. 1-4.2) or friends who sell cocaine (OR, 1.6; CI, 1.0-2.6) was independently associated with incident infection. STD incidence and prevalence were extremely high in this population, even in teenagers with only 1 lifetime partner. Individual risk behaviors appeared less important for STD risk than population factors. PMID- 10515826 TI - Correlation of behaviors with microbiological changes in vaginal flora. AB - Bacterial vaginosis (BV) is characterized by dramatic changes in the vaginal ecosystem. Women without evidence of vaginal infection may exhibit transient changes in their flora. We prospectively followed up women by using diaries and self-obtained vaginal smears to correlate behaviors with changes in flora. The majority of women (38/51, 78%) had significant, although transient, changes. Behaviors associated with unstable flora were a history of BV, a greater number of partners, and more frequent episodes of receptive oral sex. Only the latter remained significantly associated in the multivariate analysis. Variables that were associated with day-to-day variability in the flora included use of vaginal medication, menses, greater number of partners, spermicide use, more frequent vaginal intercourse, and less frequent use of condoms. Only a minority of women (11/51, 22%) maintained a "normal" lactobacillus-predominant flora. Factors associated with instability of the flora are similar to those epidemiologically associated with BV. PMID- 10515827 TI - Enhanced sensitivity of tumor necrosis factor/lymphotoxin-alpha-deficient mice to Cryptococcus neoformans infection despite increased levels of nitrite/nitrate, interferon-gamma, and interleukin-12. AB - The cytokine network and infection severity were characterized during disseminated cryptococcosis in tumor necrosis factor (TNF)- and lymphotoxin (Lt) alpha-deficient mice. On day 16, the fungus burden was higher and median survival time was reduced, as was polymorphonuclear leukocyte infiltrate in the brains of knockout mice. TNF/Lt-alpha-deficient mice had lower levels of interleukin (IL)-6 in lungs and brains, IL-1beta, and the chemokine KC in brain and spleen and of the chemokine monocyte chemoattractant protein (MCP)-1 in spleen than control animals. In contrast, higher levels of IL-6, IL-10, and MCP-1 in plasma and higher levels of IL-12, interferon (IFN)-gamma, and nitrite/nitrate were found in all compartments of TNF/Lt-alpha-deficient mice. These data confirm that TNF or Lt-alpha is a key cytokine for the anticryptococcal response and demonstrate its major role for the induction of IL-1beta, IL-6, and KC in the brain; however, its presence is not a prerequisite for IL-12, IFN-gamma, and nitrite/nitrate production. PMID- 10515828 TI - Fas-FasL interactions modulate host defense against systemic Candida albicans infection. AB - Fas-FasL costimulation modulates the production of proinflammatory cytokines, and MRL/lpr mice, which lack a functional Fas molecule, produce more proinflammatory cytokines. This study found that Fas-FasL interactions are involved in host defense against lethal infection with Candida albicans. Macrophages of MRL/lpr mice produced significantly more tumor necrosis factor and interleukin-1 after stimulation with C. albicans than did control MRL+/+ macrophages. Mortality of Fas-deficient mice with disseminated candidiasis was significantly lower than control animals' because of decreased fungal load and inhibition of the formation of invasive hyphae in their organs. Increased recruitment of neutrophils at the infection site appeared to be responsible for these effects. In contrast, phagocytosis and killing of C. albicans by neutrophils of MRL/lpr and MRL+/+ mice was similar. Absence of Fas-FasL interactions leads to increased cytokine production after C. albicans stimulation, protecting mice against disseminated candidiasis. PMID- 10515829 TI - Potent induction of focused Th1-type cellular and humoral immune responses by RTS,S/SBAS2, a recombinant Plasmodium falciparum malaria vaccine. AB - The RTS,S/SBAS2 vaccine confers sterile protection against Plasmodium falciparum sporozoite challenge. The mechanisms underlying this are of great interest, yet little is known about the immune effector mechanisms induced by this vaccine. The immune responses induced by RTS,S/SBAS2 were characterized in 10 malaria-naive volunteers. Several epitopes in the circumsporozoite protein (CSP) were identified as targets of cultured interferon (IFN)-gamma-secreting CD4+ T cells. RTS,S-specific IFN-gamma-secreting effector T cells were induced in 8 subjects; this ex vivo response mapped to a single peptide in Th2R. CSP-specific CD8+ cytotoxic T lymphocytes were not detected. RTS, S-specific IFN-gamma production was universal, whereas interleukin-4 and -5 production was rare. RTS,S-specific lymphoproliferative responses and antibodies to CSP were strongly induced in all volunteers. Responses waned with time but were boostable. Thus, RTS, S/SBAS2 is a potent inducer of Th1-type cellular and humoral immunity. These results highlight possible immune mechanisms of protection and have important implications for vaccine design in general. PMID- 10515831 TI - Antibody-dependent reductions in mouse hookworm burden after vaccination with Ancylostoma caninum secreted protein 1. AB - Vaccination of mice with either third-stage Ancylostoma caninum infective hookworm larvae (L3) or alum-precipitated recombinant Ancylostoma secreted protein 1 from A. caninum (Ac-ASP-1) results in protection against hookworm challenge infections. Vaccine protection is manifested by reductions in lung hookworm burdens at 48 h postchallenge. Mice actively immunized 4 times with Ac ASP-1 also exhibited reductions in hookworm burden in the muscles. Hookworm burden reductions from Ac-ASP-1 immunization were associated with elevations in all immunoglobulin subclasses, with the greatest rise observed in host IgG1 and IgG2b. The addition of a fourth immunization resulted in even higher levels of IgG and IgE. In contrast, L3-vaccinated mice exhibited marked elevations in IgG1 and IgM, including anti-Ac-ASP-1 IgM antibody. Passive immunization with pooled sera from recombinant Ac-ASP-1-vaccinated mice also resulted in lung hookworm burden reductions. It is hypothesized that recombinant Ac-ASP-1 vaccinations elicit antibody that interferes with parasite larval migration. PMID- 10515830 TI - Familial resemblance in humoral immune response to defined and crude Schistosoma mansoni antigens in an endemic area in Brazil. AB - This study addressed whether the humoral immune response to crude and defined Schistosoma mansoni antigens aggregates within families. The sample included 155 siblings from 42 nuclear families in Brazil. Sera examined by ELISA for antibody isotypes reactive to defined schistosome antigens and crude schistosome antigens (soluble adult worm antigen preparation and soluble egg antigen) demonstrated that there was a difference in sibling-pair correlations between defined and crude S. mansoni antigens. In contrast to the finding with crude antigens, egg positive sibling pairs showed significant familial resemblance for all IgG subclasses and IgE to adult-stage antigens Smp20.8 and Smp50. Only the IgE and IgG4 isotypes showed familial resemblance to the egg-stage antigen, Smp40. Egg negative sibling pairs showed significant familial resemblance only for IgE and IgG4 to Smp40. That both the IgE and IgG4 response to defined S. mansoni antigens showed familial resemblance is interesting in light of the converging evidence for the role of IgE and IgG4 in human susceptibility and resistance to reinfection. PMID- 10515832 TI - Human herpesvirus 8 cellular immune responses in homosexual men. AB - Little is known about cellular immunity to human herpesvirus 8 (HHV-8), the virus associated with Kaposi's sarcoma (KS). T cell proliferative responses to purified HHV-8 were measured in homosexual men, a group with elevated HHV-8 seroprevalence and high risk of KS. None of 20 blood donor controls had T cell responses to HHV 8. Among human immunodeficiency virus (HIV)-negative homosexual men, 8 (42%) of 19 HHV-8 seropositive men responded as did 4 (16%) of 25 HHV-8 seronegative men. Among HIV-positive homosexual men, however, none of 21 HHV-8 seropositives had T cell responses to HHV-8, even though most responded to common recall antigens, and 10 had >/=400 CD4 cells/mm3. The results suggest that HHV-8 T cell proliferative responses are common in HIV-negative homosexual men and that HIV infection may be associated with diminished HHV-8 cellular immunity, possibly before there is substantial depletion of CD4 cells. If correct, this could explain why KS occurs relatively early in HIV infection/AIDS. PMID- 10515833 TI - Molecular characterization of a nosocomial outbreak of human respiratory syncytial virus on an adult leukemia/lymphoma ward. AB - Although nosocomial transmission of human respiratory syncytial virus (HRSV) and its effect on morbidity and mortality among immunocompromised adults are well recognized, few studies have applied molecular techniques to differentiate nosocomial from community-acquired infections. Between January and April 1997, an outbreak of HRSV occurred among adult patients in a leukemia/lymphoma ward. Among 45 hospitalized patients undergoing bronchoscopy for investigation of acute respiratory illness, 8 were identified with HRSV infection. One infected patient developed symptoms before admission and was thought to be the index case. However, subsequent sequencing of 7 HRSV isolates identified 2 distinct genotypes, GA5 (1 case) and GB3 (6 cases). The 6 GB3 isolates could be further differentiated into 2 strains with identical nucleotide sequences that differed from each other and from 14 community HRSV isolates. Instead of a single nosocomial outbreak of HRSV, multiple introductions of HRSV likely occurred with distinct lines of nosocomial transmission. PMID- 10515834 TI - Quantification of JC virus DNA in the cerebrospinal fluid of patients with human immunodeficiency virus-associated progressive multifocal leukoencephalopathy--a longitudinal study. AB - In progressive multifocal leukoencephalopathy (PML) the JC virus (JCV) load in the cerebrospinal fluid (CSF) is discussed as a parameter for disease progression. To investigate the evolution of viral shedding into the CSF, the JCV DNA concentration was quantified by competitive polymerase chain reaction (PCR) in multiple CSF samples from prior to and during an unsuccessful intrathecal salvage therapy in 2 human immunodeficiency virus-infected patients with biopsy proven PML. With continuous clinical progression the virus load varied considerably intra- and interindividually, ranging from nondetectable to 1.2x108 genome equivalents/10 microliter CSF. Whereas an overall increase during progressive disease was confirmed, the virus burden was either constant or fluctuated irregularly during the intermediate stage of disease. This shows a variability of viral shedding during active disease that must be taken into account when the JCV load is measured by quantitative PCR for both the diagnosis of PML and monitoring under investigational treatment. PMID- 10515835 TI - Borna disease virus in human brains with a rare form of hippocampal degeneration but not in brains of patients with common neuropsychiatric disorders. AB - To estimate the frequency of persistent Borna disease virus (BDV) infections of the human central nervous system and to determine which neuropsychiatric disorders might be associated with this viral infection, reverse transcription nested polymerase chain reaction was used to screen a large collection of autopsy brain samples for the presence of BDV-specific nucleic acids. The presence of BDV RNA was found in 3 brains of persons with psychiatric symptoms and prominent hippocampal degeneration previously reported to be positive by others. However, no BDV RNA was detected in 86 randomly collected brains from persons with various psychiatric disorders, including schizophrenia, affective disorders, and Alzheimer's disease, or from suicide victims or in 52 brains from healthy controls. Furthermore, no BDV-RNA was detected in 16 surgical brain samples from persons with epilepsy-associated hippocampal sclerosis. These results indicate that life-long persistent BDV infections are rare in humans and that such infections may be associated with certain forms of hippocampal degeneration. PMID- 10515837 TI - Intrahepatic expression of interleukin-1beta and tumor necrosis factor-alpha in chronic hepatitis C. AB - The intrahepatic expression of interleukin (IL)-1beta and tumor necrosis factor (TNF)-alpha was studied in liver specimens from patients with chronic hepatitis C (n=29) and primary biliary cirrhosis (PBC; n=12) and from normal controls (n=19). IL-1beta and TNF-alpha immunoreactivity was predominantly localized in sinusoidal cells, with IL-1beta immunoreactivity being weaker in chronic hepatitis C samples than in PBC or control samples, whereas no difference in staining intensity could be observed for TNF-alpha. On semiquantitation by reverse transcription/competitive polymerase chain reaction, IL-1beta mRNA levels were significantly lower in chronic hepatitis C than in PBC or control samples (chronic hepatitis C, 0.87+/-0.77; PBC, 7.96+/-3.32; control, 3.78+/-2.56 amole IL-1beta mRNA/fmole beta-actin mRNA; P<.001). In contrast, no significant differences in TNF-alpha mRNA levels were observed between the groups. The data suggest insufficient IL-1beta production by sinusoidal cells in chronic hepatitis C, which might facilitate viral persistence. PMID- 10515836 TI - Effects of antigen dose and immunization regimens on antibody responses to a cytomegalovirus glycoprotein B subunit vaccine. AB - The purpose of this phase I study was to evaluate the safety and immunogenicity of 2 doses of cytomegalovirus glycoprotein B (CMV gB)/MF59 vaccine at 3 different immunization schedules. Ninety-five volunteers were randomized to 6 groups. Antibodies to gB represent the majority of the CMV-specific neutralizing response. Three groups received 5 microgram of gB antigen combined with MF59 (a proprietary adjuvant) and 3 groups received a 30-microgram dose at 0, 1, and 2 months; 0, 1, and 4 months; or 0, 1, and 6 months. The vaccine was well tolerated, and there was no significant difference in antibody production between the 2 doses. The vaccine induced highest antibody titers when given at 0, 1, and 6 months. A low dose of CMV gB/MF59 may be the preferred dose for future studies. PMID- 10515838 TI - Effect of small bowel bacterial overgrowth on the immunogenicity of single-dose live oral cholera vaccine CVD 103-HgR. AB - Several live oral vaccines (polio, bovine rotavirus, CVD 103-HgR cholera) are less immunogenic in developing than in industrialized countries. It was hypothesized that proximal small bowel bacterial overgrowth (common in children in less developed countries but rare in industrialized settings) diminishes the vibriocidal antibody response to CVD 103-HgR. In total, 202 fasting Santiago schoolchildren aged 5-9 years had lactulose breath H2 tests to detect proximal small bowel bacteria 1 day before ingesting CVD 103-HgR. Florid small bowel overgrowth was observed in 10 (5.6%) of 178 analyzable children. In children with florid overgrowth, vibriocidal seroconversion differed little from other children (60% vs. 67%), but the geometric mean titer was lower (160 vs. 368; P=.25). By logistic regression, increased peak breath H2 at small bowel time points was associated with diminished seroconversion (P=.04), as was the interaction of H2 value and weight (children >25 kg had lower seroconversion rates among subjects with heaviest overgrowth). PMID- 10515839 TI - Effect of nitric oxide on Helicobacter pylori morphology. AB - Helicobacter pylori causes chronic gastritis punctuated with fluctuating episodes of acute distress that can lead to peptic ulcer disease. Several factors produced by the bacterium have been shown to initiate the inflammatory response, but mechanisms potentially involved in the down-regulation of inflammation have not been described. We show that nitric oxide (NO) released from synthetic NO generators causes a rapid and dose-dependent morphologic conversion of H. pylori from the replicating spiral form to the nonreplicating, but viable, coccoid form. Because only spiral organisms-and not coccoid forms-are capable of inducing interleukin-8 secretion by epithelial cells, this conversion could result in down regulation of the inflammatory response. These data suggest that the increase in NO synthase activity observed during gastritis results in morphologic conversion to a potentially dormant but viable H. pylori. PMID- 10515840 TI - Tumor necrosis factor-alpha and interleukin-1alpha decrease the adherence of Streptococcus pyogenes to cultured keratinocytes. AB - We hypothesized that the primary epidermal cytokines, tumor necrosis factor (TNF) alpha and interleukin (IL)-1alpha, which are produced after skin injury, modulate bacterial adherence and the initiation of group A streptococcal skin infections. Streptococcus pyogenes binds preferentially to highly differentiated keratinocytes in vitro, simulating the superficial human skin infection, impetigo, and providing a model system for testing this hypothesis. Exposure of keratinocytes to 10 ng/mL TNF-alpha for 20 h decreased adherence to undifferentiated and differentiated keratinocytes by 33% and 38%, respectively. Treatment with 1 ng/mL IL-1alpha decreased adherence to undifferentiated and differentiated keratinocytes by 23% and 18%, respectively. Exposure to both cytokines simultaneously produced an additive 50% reduction in adherence. These data suggest that TNF-alpha and IL-1alpha may play a role in cutaneous host defense by impeding streptococcal adherence and decreasing its ability to form a nidus of infection in the skin. PMID- 10515841 TI - YKL-40, a matrix protein of specific granules in neutrophils, is elevated in serum of patients with community-acquired pneumonia requiring hospitalization. AB - The serum concentration of YKL-40, a matrix protein of specific granules in neutrophils, was determined by RIA in 90 patients hospitalized with pneumonia of suspected bacterial origin. Of these, 64 were followed prospectively during antibiotic treatment with blood samples taken on day 0 (on admission and the start of treatment) and on days 1, 3, 5, 7, 10, and 21. Serum YKL-40 at admission was increased in patients with Streptococcus pneumoniae pneumonia (median, 893 microgram/L; 95% confidence interval [CI], 704-1560), compared with healthy subjects (median, 102 microgram/L; 95% CI, 64-247 microgram/L; P<.001) and in patients with pneumonia of unknown etiology (median, 448 microgram/L; 95% CI, 334 700; P<.05). Peak YKL-40 serum values were observed on day 1 and thereafter declined steeply to almost normal by day 3. During the first 10 days, there was a close relation between serum YKL-40 and markers of specific granules of neutrophils (serum lactoferrin and neutrophil gelatinase-associated lipocalin), which suggests that serum YKL-40 reflects exocytosis of specific granules of neutrophils in persons with acute bacterial pneumonia. PMID- 10515842 TI - Identification of Trichomonas vaginalis alpha-actinin as the most common immunogen recognized by sera of women exposed to the parasite. AB - A study on presence of antibodies to Trichomonis vaginalis in serum was done on a group of 500 pregnant, asymptomatic Angolan women. A serologic screening, done by ELISA, revealed that 41% of the women had IgG and IgM against the parasite. Analysis of sera by immunoblotting revealed that 94.4% of sera with anti-T. vaginalis IgG class antibodies were reactive against a common immunogenic protein of 115 kDa. The common immunogen was identified as the protozoan alpha-actinin. All sera recognizing the 115-kDa antigen were reactive against both native and recombinant T. vaginalis alpha-actinin and nonreactive against human alpha actinin. The findings presented in this work offer a new tool for epidemiologic studies and open new perspectives for vaccination. PMID- 10515843 TI - Down-regulation of Th1 type of response in early human American cutaneous leishmaniasis. AB - This study examined the T cell responses in the early phase of Leishmania braziliensis infection. Cytokine profiles, lymphoproliferative responses, and skin test results in 25 patients with early cutaneous leishmaniasis (ECL; illness duration <60 days) were compared with those in persons with late cutaneous leishmaniasis (LCL; illness duration >2 months). Absent or low lymphoproliferative responses were observed in 8 (32%) of 25 patients and an absence of interferon (IFN)-gamma production in 9 (41%) of 22 patients prior to therapy. IFN-gamma production in ECL (mean +/- SD) was lower than in LCL (293+/ 346 vs. 747+/-377 pg/mL, respectively; P<.01). In contrast, interleukin (IL)-10 production in ECL (mean +/- SD) was higher than in LCL (246+/-56 vs. 50+/-41 pg/mL, respectively; P<.01). Restoration of lymphoproliferative responses and IFN gamma production was achieved when monoclonal antibody to IL-10 or IL-12 was added to the cultures. These results show that T cell responses during early phase infection are down-regulated by IL-10 and may facilitate parasite multiplication. PMID- 10515844 TI - Randomized, double-blind study of stibogluconate plus human granulocyte macrophage colony-stimulating factor versus stibogluconate alone in the treatment of cutaneous Leishmaniasis. AB - The response to recombinant human granulocyte macrophage colony-stimulating factor (GM-CSF) in the treatment of cutaneous leishmaniasis was evaluated. Twenty patients with cutaneous leishmaniasis who had lesions for 60 days were enrolled in a double-blind placebo trial of GM-CSF with standard parenteral sodium stibogluconate (20 mg/kg-1/day-1) for 20 days. Ten patients were randomized to receive intralesionally injected GM-CSF (200 microgram) at enrollment and 1 week after, and 10 patients received saline as placebo. GM-CSF- and antimony-treated patients healed faster than patients who received antimony alone (49+/-32.8 vs. 110+/-61.6 days, P<.05). Seven of 10 patients were healed of their lesions before 40 days after therapy in the GM-CSF group, compared with only 1 of 10 patients in the placebo group (relative risk, 7; 95% confidence interval, 1.04-47.00). Thus, GM-CSF plus antimony significantly increased the chance of lesion healing in 40 days. PMID- 10515845 TI - Chloroquine-resistant Plasmodium falciparum cerebral malaria in a chloroquine susceptible area. AB - Chloroquine-resistant Plasmodium falciparum is endemic in many areas. Saudi Arabia was considered to have chloroquine-susceptible P. falciparum. During the 1997-1998 season, an outbreak of malaria occurred in the southwestern region. Over a 4-month period, 32 cases (6.2%) of 520 malaria admissions met the World Health Organization criteria for cerebral malaria. The mean patient age was 28 years. Thirteen male and 19 female patients were admitted in coma. The mean duration of coma was 4.3 days; the case fatality rate was 41%. Compared with those who recovered, patients who died had a lower mean admission diastolic blood pressure and hemoglobin level, higher mean blood urea nitrogen and blood glucose levels, and thrombocytopenia. Logistic regression analysis identified treatment with quinine rather than chloroquine to be associated with survival. These findings show the potential of P. falciparum to emerge as chloroquine resistant in previously susceptible areas, resulting in significant morbidity and mortality in spite of sophisticated medical care. PMID- 10515846 TI - Cytokine expression in the brain in human cerebral malaria. AB - Evidence from clinical studies and murine models supports a role for cytokines in the pathogenesis of human cerebral malaria (CM). In this study, reverse transcriptase-polymerase chain reaction (RT-PCR) was used to investigate expression of mRNA for transforming growth factor (TGF)-beta, interleukin (IL) 1beta, and tumor necrosis factor (TNF)-alpha in human postmortem tissue. Immunohistochemistry was used to examine the distribution of cytokine protein. TGF-beta was expressed in normal brain, in CM, and in meningitis and encephalitis. IL-1beta was absent from normal brain but was detected in CM and other cerebral infections. TNF-alpha mRNA was expressed only in CM, although TNF alpha protein was also seen in meningitis. Cytokine mRNA expression in the brain did not correlate with the density of parasitized erythrocytes detected using RT PCR for major surface protein-2. This report of RT-PCR on postmortem human tissues infected with CM demonstrates induction of the proinflammatory cytokines TNF-alpha and IL-1beta in the brain. PMID- 10515847 TI - Neutralizing antibody response against human cytomegalovirus in allogeneic bone marrow-transplant recipients. PMID- 10515849 TI - Influence of primers on the detection of TT virus DNA by polymerase chain reaction. PMID- 10515850 TI - Mutations in genes associated with drug resistance in Mycobacterium tuberculosis isolates from Italy. PMID- 10515852 TI - The cytokine balance in severe malarial anemia. PMID- 10515853 TI - Reply PMID- 10515855 TI - Introduction: Focus on hematology. CD34(+) or CD34(-): does it really matter? PMID- 10515857 TI - Posttranslational processing of the thrombopoietin receptor is impaired in polycythemia vera. AB - Recently, we demonstrated a marked reduction in the expression of the thrombopoietin receptor, Mpl, in polycythemia vera (PV) platelets and megakaryocytes using an antiserum against the Mpl extracellular domain. To further examine this abnormality, we raised an antibody to the Mpl C-terminus. Immunologic analysis of PV platelets with this antiserum confirmed the reduction in Mpl expression. However, the C-terminal antiserum detected 2 forms of Mpl in PV platelets in contrast to normal platelets, in which a single form of Mpl was detected by both the extracellular domain and C-terminal antisera. Two dimensional gel electrophoresis studies with isoelectric focusing in the first dimension identified normal platelet Mpl as an 85 to 92 kD protein with an isoelectric point (pI) of 5.5. PV platelets contained an additional 80 to 82 kD Mpl Mpl isoform with a pI of 6.5. Analysis of Mpl expressed by the human megakaryocytic cell line, Dami, showed 2 isoforms similar to those found in PV platelets suggesting a precursor-product relationship. Digestion of Dami cell and normal platelet lysates with neuraminidase converted the more acidic Mpl isoform to the more basic one, indicating that the 2 isoforms differed with respect to posttranslational glycosylation. Furthermore, in contrast to normal platelet Mpl, PV platelet Mpl was susceptible to endoglycosidase H digestion, indicating defective Mpl processing by PV megakaryocytes. The glycosylation defect was specific for Mpl, as 2 other platelet membrane glycoproteins, glycoprotein IIb and multimerin, were processed normally. Importantly, the extent of the PV platelet Mpl glycosylation defect correlated with disease duration and extramedullary hematopoiesis. PMID- 10515856 TI - Reversible expression of CD34 by murine hematopoietic stem cells. AB - We used a mouse transplantation model to address the recent controversy about CD34 expression by hematopoietic stem cells. Cells from Ly-5.1 C57BL/6 mice were used as donor cells and Ly-5.2 mice were the recipients. The test cells were transplanted together with compromised marrow cells of Ly-5.2 mice. First, we confirmed that the majority of the stem cells with long-term engraftment capabilities of normal adult mice are CD34(-). We then observed that, after the injection of 150 mg/kg 5-fluorouracil (5-FU), stem cells may be found in both CD34(-) and CD34(+) cell populations. These results indicated that activated stem cells express CD34. We tested this hypothesis also by using in vitro expansion with interleukin-11 and steel factor of lineage(-) c-kit(+) Sca-1(+) CD34(-) bone marrow cells of normal mice. When the cells expanded for 1 week were separated into CD34(-) and CD34(+) cell populations and tested for their engraftment capabilities, only CD34(+) cells were capable of 2 to 5 months of engraftment. Finally, we tested reversion of CD34(+) stem cells to CD34(-) state. We transplanted Ly-5.1 CD34(+) post-5-FU marrow cells into Ly-5.2 primary recipients and, after the marrow achieved steady state, tested the Ly-5.1 cells of the primary recipients for their engraftment capabilities in Ly-5.2 secondary recipients. The majority of the Ly-5.1 stem cells with long-term engraftment capability were in the CD34(-) cell fraction, indicating the reversion of CD34(+) to CD34(-) stem cells. These observations clearly demonstrated that CD34 expression reflects the activation state of hematopoietic stem cells and that this is reversible. PMID- 10515858 TI - A bispecific diabody that mediates natural killer cell cytotoxicity against xenotransplantated human Hodgkin's tumors. AB - CD16/CD30 bispecific monoclonal antibodies can induce remissions of Hodgkin's disease refractory to chemo- and radiotherapy. However, the development of human antimouse immunoglobulin antibodies and allergic reactions precludes repeated applications of the antibody. Moreover, problems of producing and purifying sufficient amounts of material limit the clinical practicability of this novel treatment approach. To overcome these obstacles, we have constructed a bispecific antibody in a diabody form that only employs the variable domains of the CD16/CD30 hybrid hybridoma. The diabody compared favorably with the parent CD16/CD30 bispecific antibody in its ability to activate and target natural killer cells in vitro. Its administration to mice bearing xenografted Hodgkin's lymphoma resulted in a marked regression of tumor growth, thus proving for the first time the capability of a diabody for immune recruitment in vivo. The CD16/CD30 diabody is a novel reagent that should considerably facilitate the immunotherapy of patients with refractory Hodgkin's lymphoma. PMID- 10515859 TI - Uremic bleeding: closing the circle after 30 years of controversies? PMID- 10515860 TI - Lymphoproliferative syndrome with autoimmunity: A possible genetic basis for dominant expression of the clinical manifestations. AB - Fas (CD95/Apo-1) mutations were previously reported as the genetic defect responsible for human lymphoproliferative syndrome associated with autoimmune manifestations (also known as autoimmune lymphoproliferative syndrome or Canale Smith syndrome). We have identified 14 new heterozygous Fas mutations. Analysis of patients and families allow us to further dissect this syndrome with regards to the relationship between Fas mutations, inheritance pattern, and phenotype as observed on long-term follow-up. In vitro studies show that lymphocytes from all Fas mutant carriers exhibit a Fas-antibody-induced apoptosis defect. However, among the 8 inherited mutations, 4 of 4 Fas missense mutations were associated with high clinical penetrance, whereas 3 of 4 mutations leading to a truncated Fas product were associated with variable clinical penetrance. This suggests that a second defect, in another yet undefined factor involved in apoptosis and/or lymphoproliferation control, is necessary to induce full clinical expression of the disease. These results also indicate that the currently available antibody mediated in vitro apoptosis assay does not necessarily reflect the in vivo ability of abnormal Fas molecules to trigger lymphocyte death. In addition, we found that lymphoproliferative manifestations resolved with age, whereas immunological disorders [ie, hypergammaglobulinemia and detection of TcR alphabeta(+) CD4(-) CD8(-) lymphocytes] persisted. This observation suggests that Fas-mediated apoptosis plays a more important role in lymphocyte homeostasis in early childhood than later on in life. PMID- 10515861 TI - Monosomy 13 is associated with the transition of monoclonal gammopathy of undetermined significance to multiple myeloma. Intergroupe Francophone du Myelome. AB - Chromosomal abnormalities are present in most (if not all) patients with multiple myeloma (MM) and primary plasma cell leukemia (PCL). Furthermore, recent data have shown that numerical chromosomal changes are present in most individuals with monoclonal gammopathy of undetermined significance (MGUS). Epidemiological studies have shown that up to one third of MM may emerge from pre-existing MGUS. To clarify further possible stepwise chromosomal aberrations on a pathway between MGUS and MM, we have analyzed 158 patients with either MM or primary PCL and 19 individuals with MGUS using fluorescence in situ hybridization (FISH). Our FISH analyses were designed to detect illegitimate IGH rearrangements at 14q32 or monosomy 13. Whereas translocations involving the 14q32 region were observed with a similar incidence (60%) in both conditions, a significant difference was found in the incidence of monosomy 13 in MGUS versus MM or primary PCL. It was present in 40% of MM/PCL patients, but in only 4 of 19 MGUS individuals. Moreover, whereas monosomy 13 was found in the majority of plasma cells in MM, it was observed only in cell subpopulations in MGUS. It is noteworthy that, in a group of 20 patients with MM and a previous MGUS history, incidence of monosomy 13 was 70% versus 31% in MM patients without a known history of MGUS (P =.002). Thus, this study highlights monosomy 13 as correlated with the transformation of MGUS to overt MM and may define 2 groups of MM with possible different natural history and outcome, ie, post-MGUS MM with a very high incidence of monosomy 13 and de novo MM in which other genetic events might be involved. Serial analyses of individuals with MGUS will be needed to validate this model. PMID- 10515862 TI - Gene-gene and gene-environment interactions determine risk of thrombosis in families with inherited antithrombin deficiency. AB - To analyze inherited antithrombin deficiency as a risk factor for venous thromboembolism in various conditions with regard to the presence or absence of additional genetic or acquired risk factors, we compared 48 antithrombin deficient individuals with 44 nondeficient individuals of 14 selected families with inherited antithrombin deficiency. The incidence of venous thromboembolism for antithrombin deficient individuals was 20 times higher than among nondeficient individuals (1.1% v 0.05% per year). At the age of 50 years, greater than 50% of antithrombin-deficient individuals had experienced thrombosis compared with 5% of nondeficient individuals. Additional genetic risk factors, Factor V Leiden and PT20210A, were found in more than half of these selected families. The effect of exposure to 2 genetic defects was a 5-fold increased incidence (4.6% per year; 95% confidence interval [CI], 1.9% to 11.1%). Acquired risk factors were often present, determining the onset of thrombosis. The incidence among those with exposure to antithrombin deficiency and an acquired risk factor was increased 20-fold (20.3% per year; 95% CI, 12.0% to 34.3%). In conclusion, in these thrombophilia families, the genetic and environmental factors interact to bring about venous thrombosis. Inherited antithrombin deficiency proves to be a prominent risk factor for venous thromboembolism. The increased risks among those with exposure to acquired risk factors should be considered and adequate prophylactic anticoagulant therapy in high-risk situations seems indicated in selected families with inherited antithrombin deficiency. PMID- 10515863 TI - Symmetry of initial cell divisions among primitive hematopoietic progenitors is independent of ontogenic age and regulatory molecules. AB - We have developed a time-lapse camera system to follow the replication history and the fate of hematopoietic stem cells (HSC) at a single-cell level. Combined with single-cell culture, we correlated the early replication behavior with colony development after 14 days. The membrane dye PKH26 was used to monitor cell division. In addition to multiple, synchronous, and symmetric divisions, single sorted CD34(+)/CD38(-) cells derived from fetal liver (FLV) also gave rise to a daughter cell that remained quiescent for up to 8 days, whereas the other daughter cell proliferated exponentially. Upon separation and replating as single cells onto medium containing a cytokine cocktail, 60.6% +/- 9.8% of the initially quiescent cells (PKH26 bright) gave rise again to colonies and 15.8% +/- 7.8% to blast colonies that could be replated. We have then determined the effects of various regulatory molecules on symmetry of initial cell divisions. After single cell sorting, the CD34(+)/CD38(-) cells derived from FLV were exposed to flt3 ligand, thrombopoietin, stem cell factor (SCF), or medium containing a cytokine cocktail (with SCF, interleukin-3, interleukin-6, granulocyte-macrophage colony stimulating factor, and erythropoietin). Whereas mitotic rate, colony efficiency, and asymmetric divisions could be altered using various regulatory molecules, the asymmetric division index, defined as the number of asymmetric divisions versus the number of dividing cells, was not altered significantly. This observation suggests that, although lineage commitment and cell proliferation can be skewed by extrinsic signaling, symmetry of early divisions is probably under the control of intrinsic factors. PMID- 10515864 TI - Enhanced in vivo regenerative potential of HOXB4-transduced hematopoietic stem cells with regulation of their pool size. AB - After bone marrow transplantation (BMT), there is a rapid regeneration to normal pretransplantation levels in the number of hematopoietic progenitors and mature end cells, whereas hematopoietic stem cell (HSC) numbers recover to only 5% to 10% of normal levels. This suggests that HSC are significantly restricted in their self-renewal behavior and hence in their ability to repopulate the host stem cell compartment. Previously, we have reported that HSC engineered to overexpress the homeobox transcription factor HOXB4 have a large repopulation advantage over untransduced cells as assessed at 4 months in a murine transplantation model (Sauvageau et al, Genes Dev 9:1753, 1995). This phenomenon has now been examined in detail for periods extending to 12 months in cohorts of mice transplanted with various numbers of HOXB4-transduced HSC. In all mice analyzed, HOXB4-transduced HSC were capable of fully reconstituting the HSC compartment, resulting, on average, in some 14-fold greater numbers of HSC than observed when transplanting control, non-HOXB4-transduced bone marrow cells. These data indicate that HOXB4 is a limiting factor in the regeneration of HSC to normal levels after BMT. Furthermore, we show that HOXB4-transduced HSC did not expand above levels normally observed in unmanipulated mice, indicating that its overexpression does not override the regulatory mechanisms that maintain the HSC pool size within normal limits. PMID- 10515865 TI - Increased fetal and extramedullary hematopoiesis in Fas-deficient C57BL/6-lpr/lpr mice. AB - In this study, we examined the consequences of Fas deficiency on hematopoiesis in C57BL/6-lpr/lpr mice. We found a striking extramedullary increase in hematopoietic progenitor cells, comprising erythroid and nonerythroid lineages alike. These modifications preceded the lymphadenopathy, because early progenitors (colony-forming units-spleen [CFU-S] day 8) were already augmented in day-18 fetal livers of the lpr phenotype. Three weeks after birth, CFU-S increased in peripheral blood and spleen and colony-forming cells (CFU-C) began to accumulate 1 to 3 weeks later. Extramedullary myelopoiesis augmented progressively in Fas-deficient mice, reaching a maximum within 6 months. By then, mature and immature myeloid cells had infiltrated the spleen, the liver, and the peritoneal cavity. Similar changes occurred in C57BL/6-gld/gld mice, indicating that they resulted from Fas/FasL interactions. Medullary hematopoiesis was not significantly modified in adult mice of either strain. Yet, the incidence of CFU S decreased after Fas cross-linking on normal bone marrow cells in the presence of interferon gamma, consistent with a regulatory function of Fas/FasL interactions in early progenitor cell development. These data provide evidence that Fas deficiency can affect hematopoiesis both during adult and fetal life and that these modifications occur independently from other pathologies associated with the lpr phenotype. PMID- 10515866 TI - Gene duplication of zebrafish JAK2 homologs is accompanied by divergent embryonic expression patterns: only jak2a is expressed during erythropoiesis. AB - Members of the JAK family of protein tyrosine kinase (PTK) proteins are required for the transmission of signals from a variety of cell surface receptors, particularly those of the cytokine receptor family. JAK function has been implicated in hematopoiesis and regulation of the immune system, and recent data suggest that the vertebrate JAK2 gene may play a role in leukemia. We have isolated and characterized jak cDNAs from the zebrafish Danio rerio. The zebrafish genome possesses 2 jak2 genes that occupy paralogous chromosome segments in the zebrafish genome, and these segments conserve syntenic relationships with orthologous genes in mammalian genomes, suggesting an ancient duplication in the zebrafish lineage. The jak2a gene is expressed at high levels in erythroid precursors of primitive and definitive waves and at a lower level in early central nervous system and developing fin buds. jak2b is expressed in the developing lens and nephritic ducts, but not in hematopoietic tissue. The expression of jak2a was examined in hematopoietic mutants and found to be disrupted in cloche and spadetail, suggesting an early role in hematopoiesis. Taken together with recent gene knockout data in the mouse, we suggest that jak2a may be functionally equivalent to mammalian Jak2, with a role in early erythropoiesis. PMID- 10515867 TI - Genetic modification of human B-cell development: B-cell development is inhibited by the dominant negative helix loop helix factor Id3. AB - Transgenic and gene targeted mice have contributed greatly to our understanding of the mechanisms underlying B-cell development. We describe here a model system that allows us to apply molecular genetic techniques to the analysis of human B cell development. We constructed a retroviral vector with a multiple cloning site connected to a gene encoding green fluorescent protein by an internal ribosomal entry site. Human CD34(+)CD38(-) fetal liver cells, cultured overnight in a combination of stem cell factor and interleukin-7 (IL-7), could be transduced with 30% efficiency. We ligated the gene encoding the dominant negative helix loop helix (HLH) factor Id3 that inhibits many enhancing basic HLH transcription factors into this vector. CD34(+)CD38(-) FL cells were transduced with Id3-IRES GFP and cultured with the murine stromal cell line S17. In addition, we cultured the transduced cells in a reaggregate culture system with an SV-transformed human fibroblast cell line (SV19). It was observed that overexpression of Id3 inhibited development of B cells in both culture systems. B-cell development was arrested at a stage before expression of the IL-7Ralpha. The development of CD34(+)CD38(-) cells into CD14(+) myeloid cells in the S17 system was not inhibited by overexpression of Id3. Moreover, Id3(+) cells, although inhibited in their B-cell development, were still able to develop into natural killer (NK) cells when cultured in a combination of Flt-3L, IL-7, and IL-15. These findings confirm the essential role of bHLH factors in B-cell development and demonstrate the feasibility of retrovirus-mediated gene transfer as a tool to genetically modify human B-cell development. PMID- 10515868 TI - Expression of pTalpha mRNA in a committed dendritic cell precursor in the human thymus. AB - We have characterized dendritic cell precursors (pre-DC) in the human thymus. These CD1a(-)CD3(-)CD4(+)CD8(-) cells express high levels of interleukin-3Ralpha (IL-3Ralpha) on the membrane and are able to develop into mature DC upon culture with IL-3 and CD40 ligation. The DC precursors are predominantly located in the thymic medulla. Interestingly, the pre-DC express pTalpha mRNA, which is also present in CD1a(+)CD3(-)CD4(+)CD8(-) pre-T cells. Yet, the pre-DC lack expression of recombination activating gene-1 mRNA and fail to develop into T cells in appropriate assays. The thymic pre-DC are very similar to the recently characterized pre-DC found in the T cell areas of the tonsil, and it is suggested that these pre-DC populations are of lymphoid origin. PMID- 10515869 TI - The W(sh), W(57), and Ph Kit expression mutations define tissue-specific control elements located between -23 and -154 kb upstream of Kit. AB - The Kit and PDGFRa receptor tyrosine kinases are encoded in close proximity at the murine white spotting (W) and patch (Ph) loci. Whereas W mutations affect hematopoiesis, melanogenesis, and gametogenesis, the Ph mutation affects melanogenesis and causes early lethality in homozygotes. The W(sh), W(57), and Ph mutations diminish Kit expression in certain cell types such as mast cells and enhance it in others. The W(sh), W(57), and Ph mutations arose from deletions and inversions affecting sequences in between the Kit and PDGFRa genes. We have determined the precise location of the breakpoint of the W(sh) inversion and the endpoints of the W(57) deletion upstream of the Kit transcription start site and examined the effect of these mutations on Kit expression in mast cells and hematopoietic stem cells and lineage progenitors. Our results indicate that positive elements controlling Kit expression in mast cells mapping in between -23 and -154 kb from the transcription start site can be dissociated from negative elements controlling Kit misexpression during embryonic development in the vicinity of the PDGFRa gene. In addition, we have identified two clusters of hypersensitive sites in mast cells at -23 -28 kb and -147 -154 kb from the Kit gene transcription start site. Analysis of these hypersensitive sites in mutant mast cells indicates a role for HS4-6 in Kit expression in mast cells. These findings provide a molecular basis for the phenotype of these Kit expression mutations and they provide insight into the complex mechanisms governing the regulation of Kit expression. PMID- 10515870 TI - Activation of the erythropoietin receptor is not required for internalization of bound erythropoietin. AB - Erythropoietin (EPO) is required for the survival and expansion of red blood cell progenitor cells and supports continued differentiation of these committed progenitors to mature red blood cells. After binding to its cognate receptor, EPO promotes receptor homodimerization, activation of receptor-associated JAK2, subsequent receptor tyrosine phosphorylation, and transduction of signal. EPO is also internalized and degraded in lysosomes. The contribution of EPO-induced receptor internalization to modulation of EPO signals has not been determined. To examine this question, we generated a panel of hematopoietic cell lines containing progressively truncated isoforms of the erythropoietin receptor (EPO R) and determined the rate and extent of EPO internalization and receptor downregulation. We demonstrated that a membrane-proximal domain of the cytoplasmic tail of the EPO-R was the minimal region required for EPO-induced receptor internalization. This cytoplasmic domain is also the minimal domain required for activation of JAK2, a cytosolic tyrosine kinase essential for the function of the EPO-R. However, neither EPO activation of cytosolic JAK2 tyrosine kinase activity nor tyrosine phosphorylation of the EPO-R cytoplasmic tail was required for EPO-induced receptor downregulation. Both functional and nonfunctional cell surface receptor isoforms were internalized equally. These results suggest that, for downregulation of cell surface ligand occupied EPO-R and possibly for signaling receptors of the cytokine receptor superfamily in general, internalization of cell surface ligand occupied receptors may follow a pathway distinct from signaling receptors of the receptor tyrosine kinase (RTK) family. PMID- 10515871 TI - A thrombopoietin receptor mutant deficient in Jak-STAT activation mediates proliferation but not differentiation in UT-7 cells. AB - Thrombopoietin (TPO) stimulates proliferation and differentiation of cells of the megakaryocytic lineage. It exerts its function by binding and activating c-mpl, a member of the hematopoietic receptor superfamily. Upon binding of TPO to its receptor, numerous signaling events are triggered. These include activation of the Jak-STAT (signal transducers and activators of transcription) pathway, mitogen-activated protein kinase (MAPK), Tec, and phospatidylinositol (PI) 3 kinase and phosphorylation of Shc and Vav. The contribution of different signaling pathways to the induction of specific cellular processes such as proliferation and differentiation is incompletely understood. We have previously described a mutant of c-mpl that fails to activate the Jak-STAT pathway but nevertheless retains its ability to mediate proliferation and activation of most signaling events in the murine hematopoietic precursor cell lines BAF/3 and 32D. We confirm here the ability of this mutant to mediate proliferation in the absence of Jak-STAT activation in the human cell line UT-7 and further show that this mutant fails to mediate TPO-induced megakaryocytic differentiation. Comparison of the signaling capacity of this mutant in UT-7 and BAF/3 cells shows considerable cell-type-specific differences. Whereas in BAF/3 cells the mutant still mediates activation of Shc, MAPK, Vav, and PI 3-kinase at levels comparable to the wild-type receptor, these events are strongly diminished in UT-7 cells expressing the mutant. Furthermore, we show that the C-terminal 25 amino acid residues of the receptor mutant are crucial for the mitogenic response in UT-7 cells. PMID- 10515872 TI - Unique differentiation programs of human fetal liver stem cells shown both in vitro and in vivo in NOD/SCID mice. AB - Comparative measurements of different types of hematopoietic progenitors present in human fetal liver, cord blood, and adult marrow showed a large (up to 250 fold), stage-specific, but lineage-unrestricted, amplification of the colony forming cell (CFC) compartment in the fetal liver, with a higher ratio of all types of CFC to long-term culture-initiating cells (LTC-IC) and a lower ratio of total (mature) cells to CFC. Human fetal liver LTC-IC were also found to produce more CFC in LTC than cord blood or adult marrow LTC-IC, and more of the fetal liver LTC-IC-derived CFC were erythroid. Human fetal liver cells regenerated human multilineage hematopoiesis in NOD/SCID mice with the same kinetics as human cord blood and adult marrow cells, but sustained a high level of terminal erythropoiesis not seen in adult marrow-engrafted mice unless exogenous human erythropoietin (Epo) was injected. This may be due to a demonstrated 10-fold lower activity of murine versus human Epo on human cells, sufficient to distinguish between a differential Epo sensitivity of fetal and adult erythroid precursors. Examination of human LTC-IC, CFC, and erythroblasts generated either in NOD/SCID mice and/or in LTC showed the types of cells and hemoglobins produced also to reflect their ontological origin, regardless of the environment in which the erythroid precursors were generated. We suggest that ontogeny may affect the behavior of cells at many stages of hematopoietic cell differentiation through key changes in shared signaling pathways. PMID- 10515873 TI - Targeting of a heterologous protein to a regulated secretion pathway in cultured endothelial cells. AB - The stimulation of regulated exocytosis in vascular endothelial cells (EC) by a variety of naturally occurring agonists contributes to the interrelated processes of inflammation, thrombosis, and fibrinolysis. The Weibel-Palade body (WPB) is a well-described secretory granule in EC that contains both von Willebrand factor (vWF) and P-selectin, but the mechanisms responsible for the targeting of these proteins into this organelle remain poorly understood. Through adenoviral transduction, we have expressed human growth hormone (GH) as a model of regulated secretory protein sorting in EC. Immunofluorescence microscopy of EC infected with GH-containing recombinant adenovirus (GHrAd) demonstrated a granular distribution of GH that colocalized with vWF. In contrast, EC infected with an rAd expressing the IgG(1) heavy chain (IG), a constitutively secreted protein, did not demonstrate colocalization of IG and vWF. In response to phorbol ester, GH as well as endogenously synthesized vWF were rapidly released from GHrAd infected EC. By immunofluorescence microscopy, granular colocalization of GH with endogenous tissue-type plasminogen activator (tPA) was also demonstrated, and most of the tPA colocalized with vWF. These data indicate that EC are capable of selectively targeting heterologous proteins, such as GH, to the regulated secretory pathway, which suggests that EC and neuroendocrine cells share common protein targeting recognition signals or receptors. PMID- 10515874 TI - Influence of fibrillar collagen structure on the mechanisms of platelet thrombus formation under flow. AB - We have used real-time video microscopy to study the mechanisms of platelet adhesion to type I collagen fibrils of distinct structure exposed to flowing blood. Electron microscopy analysis by surface replication demonstrated morphological differences between acid-insoluble fibrils, displaying a regularly repeating striated pattern (banded collagen), and acid-soluble fibrils generated by pepsin treatment of insoluble collagen, smaller in size with a helical configuration (nonbanded collagen). These structural differences proved to be related to the role of platelet integrin alpha(2)beta(1) in stabilizing adhesion to collagen under a variety of flow conditions. Blocking alpha(2)beta(1) function with a monoclonal antibody had no effect on platelet adhesion to insoluble type I collagen coated at high density on a glass surface, whereas there was an absolute dependence of alpha(2)beta(1) function for the initial permanent arrest of platelets and subsequent thrombus formation on pepsin-solubilized type I collagen under the same conditions. In contrast, reconstituted, banded fibrils prepared from pepsin-solubilized type I collagen supported platelet adhesion and thrombus development even when platelet alpha(2)beta(1) function was blocked, a process that was greatly accelerated by pre-exposure of this substrate to autologous plasma under flow. These results implicate a collagen receptor(s) on platelets other than alpha(2)beta(1) that can selectively engage domains in banded, but not nonbanded type I collagen when alpha(2)beta(1) function is blocked. In addition, collagen structure may regulate the extent and affinity of the binding under flow of plasma components such as von Willebrand factor and/or other alpha(IIb)beta(3) ligands. PMID- 10515875 TI - Neutrophil adhesion on polyurethanes preadsorbed with high molecular weight kininogen. AB - Interaction of biomaterials with blood components including neutrophils is responsible for some of the clinical complications that have occurred in cardiopulmonary bypass, hemodialysis, and ventricular assist procedures. The possibility of inhibiting the initial adhesion of neutrophils to biomaterials has been studied extensively, but the problem remains unsolved. In this study, we investigated the effect of HK adsorption on polyurethane, a widely used component of extracorporeal and intracorporeal devices. HK and HKa were allowed to adsorb on 4 different charged polyurethanes: noncharged (PU), cationic (NR(4)), anionic (SO(3)), and zwitterionic (GPC) polyurethanes. The effect of kininogen adsorption on neutrophil adhesion, the surface density of the adsorbed kininogen, and the exposure of HK domains 3 and 5 (D(3) and D(5H)), which are responsible for the binding of HK to the neutrophil integrin alpha(m)beta(2) or Mac-1, were examined. On PU, NR(4), and SO(3), kininogen adsorption reached 80% of monolayer coverage when 100 pmol/mL or higher concentration of protein solutions were used. The NR(4) surface adsorbed the most kininogen along with a high exposure of D(3) and D(5H). The availability of D(3) and D(5H) allowed neutrophils to bind to the surface via the Mac-1 receptor; thus, on the NR(4) surface, adsorbed kininogens lost their antiadhesive property, which resulted in a high degree of neutrophil adhesion. Increasing Mac-1 expression by exposure to fMLP increased the neutrophil adhesion on this surface. In contrast, exposure of D(3) and D(5H) on SO(3) was significantly less, because HK binds to anionic surfaces with similar protein sequences used for cell binding. This low binding site exposure preserved the antiadhesive property of HK. GPC was resistant to neutrophil adhesion even in the absence of adsorbed kininogens because of its phosphorylcholine moiety. Thus, both SO(3) coupled with kininogen (or kininogen peptides) and GPC have the potential to markedly reduce neutrophil adhesion to biomaterial devices. PMID- 10515877 TI - A novel approach to arterial thrombolysis. AB - Achieving early, complete, and sustained reperfusion after acute myocardial infarction does not occur in approximately 50% of patients, even with the most potent established thrombolytic therapy. Bleeding is observed with increased concentrations of thrombolytics as well as with adjunctive antithrombotic and antiplatelet agents. A novel approach to enhance thrombolytic therapy is to inhibit the activated form of thrombin-activatable fibrinolysis inhibitor (TAFI), which attenuates fibrinolysis in clots formed from human plasma. Identification of TAFI in rabbit plasma facilitated the development of a rabbit arterial thrombolysis model to compare the thrombolytic efficacy of tissue-plasminogen activator (tPA) alone or with an inhibitor, isolated from the potato tuber (PTI), of activated TAFI (TAFIa). Efficacy was assessed by determining the time to patency, the time the vessel remained patent, the maximal blood flow achieved during therapy, the percentage of the original thrombus, which lysed, the percentage change in clot weight, the net clot accreted, and the release of radioactive fibrin degradation products into the circulation. The results indicate that coadministration of PTI and tPA significantly improved tPA-induced thrombolysis without adversely affecting blood pressure, activated partial thromboplastin time, thrombin clotting time, fibrinogen, or alpha-2-antiplasmin concentrations. The data indicate that inhibitors of TAFIa may comprise novel and very effective adjuncts to tPA and improve thrombolytic therapy to achieve both clot lysis and vessel patency. PMID- 10515876 TI - Circulating activated platelets assist THP-1 monocytoid/endothelial cell interaction under shear stress. AB - Circulating complexes of leukocytes and activated platelets are markers for atherosclerosis, but their interaction with the arterial endothelial lining has not been studied. Therefore, the effect of activated platelets on rolling and adhesion of labeled human THP-1 monocytoid cells to human umbilical vein endothelial cell (HUVEC) monolayers was studied by epifluorescence microscopy in a parallel plate flow chamber. In the absence of activated platelets, THP-1 rolling on resting HUVEC was negligible at shear rates greater than 300 s(-1). Activation of HUVEC with 100 nmol/L phorbol myristate acetate (PMA) increased THP 1 cell adhesion at shear rates less than 400 s(-1). Therefore, a shear rate of 400 s(-1) was identified as a threshold for THP-1 adhesion. THP-1 rolling on activated HUVEC was reduced by 64% after L-selectin inhibition but was not affected by P-selectin inhibition. The addition of 1 to 50 thrombin receptor activating peptide (TRAP)-activated platelets per THP-1 cell enhanced interactions between THP-1 cells and HUVEC, resulting in a steep bell-shaped dose response curve, with a peak of 10 +/- 3 rolling cells/50 seconds at 3 platelets per THP-1 cell (P <.01 v control) with a concomitant 2- to 3-fold increase of firmly adhering cells (P <.01 v control). In reconstituted blood, low numbers of activated platelets had the same effect on THP-1 rolling and adhesion. P-selectin inhibition reduced platelet/THP-1 cell interaction in suspension and deposition of the complexes on the endothelial monolayer. Inhibition of both P- and L selectin reduced THP-1/HUVEC interactions to 14% (P <.01, n = 4). Sialidase digestion and removal of terminal sialic acid residues from HUVEC or THP-1 cells but not from platelets abolished the platelet mediated augmentation of THP-1 cell adhesion. Thus, THP-1 rolling on HUVEC is shear-dependent and largely mediated by L-selectin. P-selectin expressed on activated platelets increases monocytoid cell adhesion to endothelial cells at shear rates found in coronary arteries through interactions with both endothelial and monocytoid cells and may facilitate macrophage accumulation in the vessel wall. PMID- 10515878 TI - Beta(2)-microglobulin identified as an apoptosis-inducing factor and its characterization. AB - Major histocompatibility complex (MHC) molecules play an important role in antigen presentation for induction of tumor as well as cellular and humoral immunities. Recent studies using anti-MHC antibodies demonstrated that antibodies specific for HLA class I molecules induced cellular activation and a type of apoptosis that may be distinct from Fas-dependent or TNFR (tumor necrosis factor alpha receptor)-dependent processes. We purified a previously untested apoptosis inducing factor from HL-60 human leukemic cell-conditioned media to homogeneity and sequenced it. It was identified as beta(2)-microglobulin (beta(2)m), which has been previously known as thymotaxin and is a part of the HLA class I antigen complex. beta(2)m acts on both T-leukemic cells and myeloid leukemic cells to induce apoptosis, which then activates caspase 1 and 3. Cross-linking studies showed that biotinilated beta(2)m recognized an epitope distinct from those recognized by the anti-HLA class I antibody, as reported previously. We demonstrated that beta(2)m plays a previously unrecognized and important role in regulating the elimination of tumor cells, which occurs as a result of the action of beta(2)m as an apoptosis-inducing factor. PMID- 10515879 TI - Fibronectin upregulates gelatinase B (MMP-9) and induces coordinated expression of gelatinase A (MMP-2) and its activator MT1-MMP (MMP-14) by human T lymphocyte cell lines. A process repressed through RAS/MAP kinase signaling pathways. AB - T-lymphocyte migration into tissues requires focal degradation of the basement membrane. In this study, we show that transient adherence to fibronectin induces the production of activated forms of matrix metalloproteinase-2 (MMP-2) and MMP 9, as well as downregulation of tissue inhibitor of metalloproteinase-2 (TIMP-2) by T-cell lines. MMP-2 activation was likely achieved by inducing a coordinated expression of membrane-type matrix metalloproteinase-1 (MMP-14), a major activator of MMP-2. Blocking monoclonal antibodies against alpha4, alpha5, and alphav integrins strongly reduced MMP-2 and MMP-9 production induced by fibronectin. Disrupting actin cytoskeleton organization by cytochalasin D strongly enhanced fibronectin-induced MMP-2 and MMP-9 expression. Inhibiting Src tyrosine kinases with herbimycin A reduced MMP-2 and MMP-9 production with no effect on cell attachment. By contrast, G-protein inhibition by pertussis toxin, or transfection with a dominant negative mutant of Ha-Ras strongly increased fibronectin-induced MMP-2 and MMP-9. Inhibition of PI3 kinase, MAPkinase (MEK1), or p38 MAPkinase by wortmannin, PD 98059, or SB 202190, respectively, strongly promoted fibronectin-induced MMP2 and MMP-9. Cells at high density lost their ability to synthesize MMP-2 and MMP-9 in response to fibronectin and MMP expression was restored by transfection with a dominant-negative mutant of Ha-Ras or by treatment with wortmannin, PD 98059, or SB 202190. Our findings suggest that adhesion to fibronectin transduces both stimulatory (through Src-type tyrosin kinases) and inhibitory signals (through Ras/MAPKinase signaling pathways) for MMP-2 and MMP-9 expression by T lymphocytes and that their relative predominance is regulated by additional stimuli related to cell adhesion, motility, and growth. PMID- 10515881 TI - An inositolphosphate-binding immunophilin, IPBP12. AB - A novel inositolphosphate-binding protein has been identified and shown to be an immunophilin. This protein, which was isolated from human erythrocyte membranes and from K562 (human erythroleukemia) cell membranes, has robust peptidylprolyl cis-trans isomerase activity that is strongly inhibited by nanomolar concentrations of FK506 or rapamycin, indicating a member of the FKBP (FK506 binding protein) class. However, unlike the cytosolic FKBP12, the isomerase activity of this membrane-associated immunophilin is strongly inhibited by nanomolar concentrations of inositol 1,4, 5-trisphosphate (IP(3)), inositol 1,3,4,5-tetrakisphosphate (IP(4)), and phosphatidylinositol 4- and 4,5 phosphates, which are suggested to be physiological ligands. The demonstration of a single 12-kD protein that binds both IP(4) or IP(3) and anti-FKBP12 provides strong support for the inositolphosphate-binding immunophilin having an apparent mass of 12 kD, and it is suggested that the protein might be called IPBP12 for 12 kD inositol phosphate binding protein. When an internal tryptic peptide derived from IPBP12 was sequenced, a sequence also present in human cytokeratin 10 was identified, suggesting a cytoskeletal localization for the immunophilin. While purifying IPBP12, it was found that it is immunoprecipitated with specific proteins that include a protein kinase and a phosphoprotein phosphatase. The latter is indicated to be phosphoprotein phosphatase 2A (PP-2A). It is suggested that immunophilins promote the assembly of multiprotein complexes that often include a protein kinase or a phosphoprotein phosphatase or both. PMID- 10515880 TI - Signaling through CD43 induces natural killer cell activation, chemokine release, and PYK-2 activation. AB - Natural killer (NK) cell activation is the result of a balance between positive and negative signals triggered by specific membrane receptors. We report here the activation of NK cells induced through the transmembrane glycoprotein CD43 (leukosialin, sialophorin). Engagement of CD43 by specific antibodies stimulated the secretion of the chemokines RANTES, macrophage inflammatory protein (MIP) 1alpha, and MIP-1beta, which was prevented by treatment of cells with the specific tyrosine kinase inhibitor genistein. Furthermore, signaling through CD43 increased the cytotoxic activity of NK cells and stimulated an increase in the tyrosine kinase activity in antiphosphotyrosine immune complexes of NK cell lysates. PYK-2 was identified among the tyrosine kinase proteins that become activated. Hence, PYK-2 activation was observed after 20 minutes of CD43 stimulation, reached a maximum after 45 to 60 minutes, and decreased to almost basal levels after 120 minutes of treatment. Together, these results demonstrate the role of CD43 as an activation molecule able to transduce positive activation signals in NK cells, including the regulation of chemokine synthesis, killing activity, and tyrosine kinase activation. PMID- 10515882 TI - Analysis of T cells in paroxysmal nocturnal hemoglobinuria provides direct evidence that thymic T-cell production declines with age. AB - Peripheral blood T cells in patients with paroxysmal nocturnal hemoglobinuria (PNH) comprise a mixture of residual normal and glycosylphosphatidylinositol (GPI)-deficient PNH cells. Using multicolor flow cytometry, we demonstrated significant differences between the proportions of naive and memory cells within these populations. PNH T cells comprise mainly naive cells (CD45RA(+)CD45R0(-)), whereas normal T cells in the same patients were predominantly memory (CD45RA( )CD45R0(+)) cells. Functional analyses showed that GPI-deficient CD45RA(+) T cells can convert to a CD45R0(+) phenotype. We present data from a PNH patient in remission for 20 years who still had significant numbers of GPI-deficient T cells; these showed a normal distribution of naive and memory components. The predominantly naive phenotype of GPI-deficient T cells seen in PNH patients with active disease likely reflects the phenotype of recent normal thymic emigrants. In patients where hematopoiesis was predominantly derived from the PNH stem cell, absolute numbers of both naive PNH CD4(+) cells and CD8(+) cells show an inverse correlation with patient age, implying this age-related decline in T-cell production is secondary to a decrease in thymic activity rather than a stem cell defect. PMID- 10515884 TI - Human immunodeficiency virus nef gene expression affects generation and function of human T cells, but not dendritic cells. AB - Human immunodeficiency virus (HIV)-infected individuals develop an acquired immune deficiency syndrome (AIDS) due to loss in their lymphocyte numbers and cellular defects in T cells and antigen-presenting cells (APC). HIV infection of the thymus results in deficient replenishment of the peripheral naive T-cell pool. The HIV nef gene was shown to be important for progression towards AIDS and cellular depletion of the infected thymus. Here, we demonstrate by retroviral gene transfer that nef expression, in the absence of other HIV genes, impaired human thymic T-cell development. Thymocytes were generated in reduced numbers and downmodulated CD4 and CD8beta cell surface expression. T cells grown from nef expressing thymocytes were hyperproliferative in vitro upon T-cell receptor triggering. Mature dendritic cells (DC) were functional and had normal surface CD4 levels despite nef expression. Thus, nef expression alone may contribute to AIDS development by reduced T-cell generation and T-cell hyperresponsiveness. PMID- 10515883 TI - Monocytoid B cells are distinct from splenic marginal zone cells and commonly derive from unmutated naive B cells and less frequently from postgerminal center B cells by polyclonal transformation. AB - Monocytoid B cells represent a morphologically conspicuous B-cell population that constantly occurs in Toxoplasma gondii-induced Piringer's lymphadenopathy. Although widely believed to be closely related to splenic marginal zone B cells, neither this relationship, nor the B-cell differentiation stage of monocytoid B cells, nor their cellular precursors have been established. We have therefore examined monocytoid B cells for their expression of B-cell differentiation markers and the Ig isotypes at the RNA and protein level as well as for rearranged Ig heavy chain (H) genes and somatic mutations within the variable (V) region. The results obtained were compared with the corresponding features of other B-cell populations. The monocytoid B cells displayed immunophenotypical differences to all other B-cell populations. IgM and IgD expression was absent from most monocytoid B cells at the RNA and protein levels. Unrelated (polyclonal) Ig rearrangements were found in 85 of the 95 cells studied. Seventy four percent of the rearranged VH genes were devoid of somatic mutations, whereas the remaining 26% carried a low number of somatic mutations. The majority of these showed no significant signs of antigen selection. This finding in conjunction with the predominantly unrelated Ig gene rearrangements indicates that most monocytoid B cells arise not by clonal proliferation but by transformation of polyclonal B cells. The B cells undergoing a monocytoid B-cell transformation are in the majority (74%) naive B cells, and only a minority are (26%) non-antigen-selected postgerminal center B cells. Thus, our data show that monocytoid B cells represent a distinct B-cell subpopulation. PMID- 10515885 TI - Early maturation of T-cell progenitors in the absence of glucocorticoids. AB - In the present work, we demonstrated that both fetal liver and thymic T-cell precursors express glucocorticoid receptors (GRs) indirectly suggesting a role for glucocorticoids (GCs) in the earliest events of T-cell differentiation. To evaluate this issue, we analyzed the thymic ontogeny in the progeny of adrenalectomized pregnant rats (Adx fetuses), an in vivo experimental model, which ensures the absence of circulating GCs until the establishment of the fetal hypothalamus-pituitary-adrenal (HPA) axis. In the absence of maternal GCs, T-cell development was significantly accelerated, the process being reversed by in vivo GC replacement. Mature single positive thymocytes (both CD4 and CD8) appeared in 16-day old fetal Adx thymus when in the control fetuses, most thymocytes still remained in the double-negative (DN) CD4(-)CD8(-) cell compartment. In addition, emigration of T-cell receptor (TcR)alphabeta positive cells to the spleen also occurred earlier in Adx fetuses than in control ones. In vitro recolonization of cultured deoxiguanosine-treated mouse fetal thymus lobes with 13-day-old fetal liver cell suspensions from both Adx and control fetuses demonstrated changes in the developmental capabilities of fetal liver T-cell precursors from embryos grown in the absence of GCs. Furthermore, a precocious lymphoid colonization of the thymic primordium from Adx fetuses was evidenced by ultrastructural analysis of both Adx and Sham early thymus. Both findings accounted for the accelerated T cell differentiation observed in Adx fetuses. Together, these results support a role for GCs not only in the thymic cell death, but also in the early steps of T cell differentiation. PMID- 10515886 TI - Resting and cytokine-stimulated human small airway epithelial cells recognize and engulf apoptotic eosinophils. AB - Eosinophils, which are prominent cells in asthmatic inflammation, undergo apoptosis and are recognized and engulfed by phagocytic macrophages in vitro. We have examined the ability of human small airway epithelial cells (SAEC) to recognize and ingest apoptotic human eosinophils. Cultured SAEC ingested apoptotic eosinophils but not freshly isolated eosinophils or opsonized erythrocytes. The ability of SAEC to ingest apoptotic eosinophils was enhanced by interleukin-1alpha (IL-1alpha) or tumor necrosis factor alpha (TNFalpha) in a time- and concentration-dependent fashion. IL-1alpha was found to be more potent than TNFalpha and each was optimal at 10(-10) mol/L, with a significant (P <.05) effect observed at 1 hour postcytokine incubation that was maximal at 5 hours. IL 1alpha stimulation not only increased the number of SAEC engulfing apoptotic eosinophils, but also enhanced their capacity for ingestion. The amino sugars glucosamine, n-acetyl glucosamine, and galactosamine significantly inhibited uptake of apoptotic eosinophils by both resting and IL-1alpha-stimulated SAEC, in contrast to the parent sugars glucose, galactose, mannose, and fucose. Incubation of apoptotic eosinophils with the tetrapeptide RGDS, but not RGES, significantly inhibited their uptake by both resting and IL-1alpha-stimulated SAEC, as did monoclonal antibody against alphavbeta3 and CD36. Thus, SAEC recognize apoptotic eosinophils via lectin- and integrin-dependent mechanisms. These data demonstrate a novel function for human bronchial epithelial cells that might represent an important mechanism in the resolution of eosinophil-induced asthmatic inflammation. PMID- 10515887 TI - In vitro evaluation of fludarabine in combination with cyclophosphamide and/or mitoxantrone in B-cell chronic lymphocytic leukemia. AB - B-chronic lymphocytic leukemia (B-CLL) is characterized by the accumulation of long-lived CD5(+) B lymphocytes. We have analyzed the effect in vitro of the combination of fludarabine with cyclophosphamide and/or mitoxantrone on cells from 20 B-CLL patients. Mafosfamide, the active form of cyclophosphamide in vitro, increased the cytotoxicity of fludarabine in all of the patients studied and produced a significant synergistic effect (P <.01) after 48 hours of incubation. The addition of mitoxantrone to this combination increased the cytotoxic effect in cells from 8 patients, but in the remaining 12 patients no significant increase was observed. The effect of fludarabine and mafosfamide was dose-dependent. Mafosfamide and fludarabine had a synergistic effect in inducing apoptosis of B-CLL cells as determined by DNA staining with propidium iodide and analysis of phosphatidylserine exposure. Mafosfamide significantly increased the apoptosis induced by fludarabine on CD19(+) cells (P =.007), but not on CD3(+) cells (P =. 314). Cell viability was correlated with a decrease in Mcl-1 levels and an increase in p53 levels. These results support that fludarabine in combination with cyclophosphamide and/or mitoxantrone can be highly effective in the treatment of B-CLL. PMID- 10515888 TI - Phenylarsine oxide blocks interleukin-1beta-induced activation of the nuclear transcription factor NF-kappaB, inhibits proliferation, and induces apoptosis of acute myelogenous leukemia cells. AB - Arsenic compounds have recently been shown to induce high rates of complete remission in patients with acute promyelocytic leukemia (APL). One of these compounds, As(2)O(3), induces apoptosis in APL cells via a mechanism independent of the retinoic acid pathway. To test the hypothesis that arsenic compounds may be effective against other forms of acute myelogenous leukemia (AML), we studied the membrane-permeable arsenic compound phenylarsine oxide (PAO). Because interleukin-1beta (IL-1beta) plays a key role in AML cell proliferation, we first tested the effect of PAO on OCIM2 and OCI/AML3 AML cell lines, both of which produce IL-1beta and proliferate in response to it. We found that PAO inhibited the proliferation of both OCIM2 and OCI/AML3 cells in a dose-dependent fashion (0.01 to 0.1 micromol/L) and that IL-1beta partially reversed this inhibitory effect. We then measured IL-1beta levels in these cells by using an enzyme-linked immunosorbent assay and Western immunoblotting and found that PAO almost completely abolished the production of IL-1beta in these AML cells, whereas it did not affect the production of IL-1 receptor antagonist. Because PAO inhibits activation of the transcription factor NF-kappaB and because NF-kappaB modulates an array of signals controlling cellular survival, proliferation, and cytokine production, we also studied the effect of PAO on NF-kappaB activation in AML cells and found that PAO suppressed the IL-1beta-induced activation of NF-kappaB. Because inhibition of NF-kappaB may result in cellular apoptosis, we also tested whether PAO may induce apoptotic cell death in AML cells. We found that PAO induced apoptosis in OCIM2 cells through activation of the cystein protease caspase 3 and subsequent cleavage of its substrate, the DNA repair enzyme poly (ADP-ribose) polymerase. The PAO-induced apoptosis was caspase dependent, because it was completely blocked by the caspase inhibitor Z-DEVD-FMK. Finally, we tested the effect of PAO on fresh AML marrow cells from 7 patients with newly diagnosed AML and found that PAO suppressed AML colony-forming cell proliferation in a dose dependent fashion. Taken together, our data showing that PAO is an effective in vitro inhibitor of AML cells suggest that this compound may have a role in future therapies for AML. PMID- 10515889 TI - A deletion in the gene for transforming growth factor beta type I receptor abolishes growth regulation by transforming growth factor beta in a cutaneous T cell lymphoma. AB - Spontaneous regression of skin lesions is characteristic of lymphomatoid papulosis (LyP), a clonal cutaneous lymphoproliferative disorder. A minority of LyP patients progress to anaplastic large cell lymphoma (ALCL) in which skin lesions no longer regress and extracutaneous dissemination often occurs. In 1 such case, we developed a tumor cell line, JK cells, and show that these cells are resistant to the growth inhibitory effects of transforming growth factor beta (TGF-beta) due to the loss of cell surface expression of the TGF-beta type I receptor (TbetaR-I). Reverse transcriptase-polymerase chain reaction (RT-PCR) and sequencing of JK cell TbetaR-I cDNA clones identified a deletion that spanned the last 178 bp of exon 1, including the initiating methionine. Hybridization of a radiolabeled fragment internal to the deletion was detected in the genomes of TGF beta-responsive cells, but not in JK cells, indicating that they contain no wild type TbetaR-I gene. PCR primers that flanked the deleted TbetaR-I region amplified a single band from JK cell genomic DNA that lacked the last 178 bp of exon 1 and all of the approximately 5 kb of intron 1. This JK cell-specific genomic TbetaR-I PCR product was distinct from products amplified from TGF-beta responsive cells and was also readily detected in tumor biopsies obtained before the establishment of the JK cell line. Our results identify the first inactivating mutation in TbetaR-I gene in a human lymphoma that renders it insensitive to growth inhibition by TGF-beta. PMID- 10515890 TI - Expression of human inducible nitric oxide synthase gene in T-cell lines infected with human T-cell leukemia virus type-I and primary adult T-cell leukemia cells. AB - We examined the expression of messenger RNA (mRNA) of the human inducible nitric oxide synthase (hiNOS) gene in a panel of human T-cell lines. Reverse transcriptase-polymerase chain reaction showed that human T-cell leukemia virus type-I (HTLV-I)-infected T-cell lines (MT-1, SLB-1, and C5/MJ) expressed mRNA for the hiNOS, but TL-Om1 or uninfected Jurkat, H9, and CCRF-CEM did not. The MT-1, SLB-1, and C5/MJ cell lines are infected with HTLV-I and express the viral transactivator Tax, whereas TL-Om1 cells, although derived from adult T-cell leukemia (ATL) leukemic cells, do not express Tax. There was, thus, a correlation between Tax and hiNOS mRNA expression. The transcriptional regulatory region of the hiNOS gene was activated by Tax in Jurkat, in which endogenous hiNOS is induced by Tax. Deletion analysis showed that the region of hiNOS encompassing nucleotides -159 to -111 contained the minimum Tax-responsive elements. Mutations in the NF-kappaB element at position -115 and -106 bp in the hiNOS promoter were still activated by Tax, and a Tax mutant defective for activation of the NF kappaB pathway retained the ability to activate the hiNOS promoter. In addition, overexpression of the dominant-negative mutants of IkappaBalpha and I kappaBbeta failed to reduce Tax-induced activation of hiNOS gene. Furthermore, hiNOS mRNA was detected in leukemic cells from ATL patients. Our results show that the hiNOS promoter contains a minimum Tax-responsive element located between nucleotides 159 and -111, and imply that the expression of the hiNOS gene is involved in the pathogenesis of HTLV-I-associated diseases. PMID- 10515891 TI - Involvement of interleukin-10 (IL-10) and viral IL-6 in the spontaneous growth of Kaposi's sarcoma herpesvirus-associated infected primary effusion lymphoma cells. AB - Primary effusion lymphoma (PEL) is a distinct type of lymphoma associated with Kaposi's sarcoma-associated herpesvirus (KSHV) infection. To determine the factors responsible for the unrestrained proliferation of PEL, we have studied the growth factor requirements of the PEL-derived BCBL-1 and BC-1 cell lines. Both cell lines were found to be autocrine growth factor dependent and to release human interleukin-6 (IL-6), viral IL-6 (vIL-6), and human IL-10 in the culture supernatant. To establish whether these cytokines contribute to autocrine growth, neutralizing antibodies against human IL-6, vIL-6, human IL-10, and soluble IL-10 receptor were used. These experiments showed that human IL-10 and, to a lesser degree, vIL-6 serve as autocrine growth factors for BCBL-1 and BC-1 cells. Thus, human IL-10 and vIL-6 are growth factors released and used by PEL cells for autonomous proliferation and may be critical to the development and progression of PEL. PMID- 10515893 TI - The molecular basis of a case of gamma-glutamylcysteine synthetase deficiency. AB - Gamma-glutamylcysteine synthetase catalyzes the first step in glutathione synthesis. The enzyme consists of 2 subunits, heavy and light, with the heavy subunit serving as the catalytic subunit. A patient with hemolytic anemia and low red blood cell glutathione levels was found to have a deficiency of gamma glutamylcysteine synthetase activity. Examination of cDNA from the patient and her mother showed that she was homozygous and that her mother was heterozygous for a A-->T transversion at nt1109 producing a deduced amino acid change of His370Leu. The partial genomic structure of the catalytic subunit of gamma glutamylcysteine synthetase (GLCLC) was determined, providing some intron/exon boundaries to make it possible to sequence an affected part of the coding region from genomic DNA. The 1109A-->T mutation was not present in the DNA of 38 normal subjects. In the course of these studies we found a diallelic polymorphism in nt +206 of an intron and another polymorphism that consisted of a duplication of a CAGC at cDNA nt1972-1975 in the 3' untranslated region. The 2 polymorphisms were found to be only in partial linkage disequilibrium. PMID- 10515892 TI - Interleukin-10 (IL-10) selectively enhances CIS3/SOCS3 mRNA expression in human neutrophils: evidence for an IL-10-induced pathway that is independent of STAT protein activation. AB - We have recently shown that, in human neutrophils, interleukin-10 (IL-10) fails to induce specific DNA-binding activities to the gamma-interferon response region (GRR), a regulatory element located in the FcgammaRI gene promoter, which is required for transcriptional activation by IL-10 and interferon gamma (IFNgamma) in monocytic cells. In this study, we report that IL-10 is also unable to induce the binding of STAT1 or STAT3 to the serum-inducible element (hSIE/m67), despite the fact that both proteins are expressed in neutrophils. Whereas IFNgamma and granulocyte colony-stimulating factor (G-CSF) are efficient inducers of STAT1 and STAT3 tyrosine phosphorylation in polymorphonuclear neutrophils (PMN), IL-10 fails to trigger STAT1 and STAT3 tyrosine and serine phosphorylation, therefore explaining its inability to induce the FcgammaRI expression in these cells. By contrast, we demonstrate that IL-10 alone represents an efficient stimulus of CIS3/SOCS3 mRNA expression in neutrophils. CIS3/SOCS3 belongs to the recently cloned cytokine-inducible SH2-containing protein (CIS) gene family (which also includes CIS1, CIS2, CIS4, CIS5, and JAB) that is believed to be, at least in part, under the control of STAT transcription factors and whose products are potential modulators of cytokine signaling. Moreover, IL-10 synergizes with lipopolysaccharide (LPS) in upregulating CIS3/SOCS3 mRNA expression in PMN through a mechanism that involves mRNA stabilization. In contrast to CIS3/SOCS3, mRNA transcripts encoding other family members are unaffected by IL-10 in neutrophils. Finally, transfection of CIS3/SOCS3 in murine M1 myeloid cells suppresses LPS-induced growth arrest, macrophage-like differentiation, and nitric oxide synthesis, but not IL-6 mRNA expression. Collectively, our data suggest that, in neutrophils, the activation of STAT1 and STAT3 phosphorylation is neither required for CIS3/SOCS3 induction by IL-10 nor involved in the regulatory effects of IL-10 on cytokine production. PMID- 10515894 TI - ABO blood group antigens on human plasma von Willebrand factor after ABO mismatched bone marrow transplantation. AB - von Willebrand factor (vWF) is synthesized exclusively by endothelial cells and megakaryocytes, and stored in the intracellular granules or constitutively secreted into plasma. ABO blood group antigens are covalently associated with asparagine-linked sugar chains of plasma vWF. The effect of ABO-mismatched bone marrow transplantation (BMT) or blood stem cell transplantation (BSCT) on the expression of ABO blood group antigens on the vWF was examined to obtain information on the origin of these antigens. In ABO-mismatched (HLA-matched) groups, 8 cases of BMT and 4 cases of BSCT were examined. In all cases, the ABO blood groups on red blood cells were gradually converted to the donor's type within 80 to 90 days after the transplantation. The blood group antigens on the vWF were consistent with the recipient's blood group for the period monitored by enzyme-linked immunosorbent assay (ELISA). When vWF was isolated from normal platelets and examined for the blood group antigens using ELISA or immunoblotting, it showed few antigens. However, vWF extracted from veins expressed blood group antigens. These findings indicate that platelet (megakaryocyte)-derived vWF does not contain blood group antigens and that these antigens may be specifically associated with vWF synthesized in endothelial cells and secreted into plasma. Furthermore, it is possible that the persistence of the recipient's blood group antigens on plasma glycoproteins such as vWF, independent of the donor-derived erythrocytes, after ABO-mismatched stem cell transplantation, may influence the immunological system in the production of anti blood group antibodies resulting in the establishment of immunological tolerance in the recipient plasma. PMID- 10515895 TI - Shiga-like toxin-1 receptor on human breast cancer, lymphoma, and myeloma and absence from CD34(+) hematopoietic stem cells: implications for ex vivo tumor purging and autologous stem cell transplantation. AB - The ribosome-inactivating protein, Shiga-like toxin-1 (SLT-1), targets cells that express the glycolipid globotriaosylceramide (CD77) on their surface. CD77 and/or SLT-1 binding was detected by flow cytometry and immunocytochemistry on lymphoma and breast cancer cells recovered from biopsies of primary human cancers as well as on B cells or plasma cells present in blood/bone marrow samples of multiple myeloma patients. Breast cancer cell lines also expressed receptors for the toxin and were sensitive to SLT-1. Treatment of primary B lymphoma, B-cell chronic lymphocytic leukemia, and myeloma B or plasma cells with SLT-1-depleted malignant B cells by 3- to 28-fold, as measured by flow cytometry. Depletion of myeloma plasma cells was confirmed using a cellular limiting dilution assay followed by reverse transcriptase-polymerase chain reaction analysis of clonotypic IgH transcripts, which showed a greater than 3 log reduction in clonotypic myeloma cells after SLT-1 treatment. Receptors for the toxin were not detected on human CD34(+) hematopoietic progenitor cells (HPC). HPC were pretreated with a concentration of SLT-1 known to purge primary malignant B cells and cultured for 6 days. The number of HPC was comparable in toxin-treated and untreated cultures. HPC were functionally intact as well. Colony-forming units (CFU) were present at an identical frequency in untreated and SLT-1 pretreated cultures, confirming that CFU escape SLT-1 toxicity. The results suggest the ex vivo use of SLT-1 in purging SLT-1 receptor-expressing malignant cells from autologous stem cell grafts of breast cancer, lymphoma, and myeloma patients. PMID- 10515896 TI - Correlation between disparity for the minor histocompatibility antigen HA-1 and the development of acute graft-versus-host disease after allogeneic marrow transplantation. AB - Results of a previous study suggested that recipient mismatching for the minor histocompatibility antigen HA-1 is associated with acute graft-versus-host disease (GVHD) after allogeneic marrow transplantation. In that study, most patients received either cyclosporine or methotrexate for GVHD prophylaxis, and a cytotoxic T-cell clone was used to test for HA-1 disparity. To facilitate large scale testing, we developed a method that uses genomic DNA to identify HA-1 alleles. A retrospective study was conducted to correlate HA-1 disparity and the occurrence of acute GVHD in 237 HLA-A2-positive white patients who had received a marrow or peripheral blood stem cell transplant from an HLA-identical sibling. All patients received both methotrexate and cyclosporine for GVHD prophylaxis. The presence of HLA-A*0201 was confirmed in 34 of the 36 HA-1 disparate pairs by sequencing the HLA-A locus. Grades II-IV GVHD occurred in 22 (64.7%) of these 34 patients, compared with 86 (42.8%) of the 201 patients without HA-1 disparity (odds ratio, 2. 45; 95% confidence interval [CI], 1.15 to 5.23; P =.02). Recipient HA-1 disparity showed a trend for association with acute GVHD (odds ratio, 2.1; 95% CI, 0.91 to 4.68; P =.08) when a multivariable logistic regression model was used to include additional risk factors. These data are consistent with results of the previous study, suggesting an association between HA-1 disparity and risk of acute GVHD, but the strength of this association may be lower in patients who received both methotrexate and cyclosporine than in those who received methotrexate or cyclosporine alone. PMID- 10515897 TI - Viral hyperinfection of the central nervous system and high mortality after hematopoietic stem cell transplantation for treatment of Theiler's murine encephalomyelitis virus-induced demyelinating disease. AB - Theiler's murine encephalomyelitis virus (TMEV) establishes a persistent infection in the central nervous system (CNS) leading to an inflammatory demyelinating disease of the CNS in which the histology and clinical course is similar to multiple sclerosis (MS). Disease pathogenesis is primarily due to T cell-mediated destruction of myelin, which has been attributed to cytopathic effects of the virus, but immune-mediated destruction of myelin mediated via both virus-specific and myelin-specific T cells appear to play the major role. To determine if bone marrow transplantation would be an effective therapy for a virus-initiated autoimmune disease and to better separate viral cytopathic effects from immune-mediated demyelination, we ablated the immune system of TMEV infected animals with 1,100 cGy total body irradiation, and then the animal's immunity was reconstituted by transplantation of disease-susceptible SJL/J mice with syngeneic marrow or disease-susceptible DBA/2J with marrow from disease resistant (C57Bl/6 x DBA/2)F1 (B6D2) donors. Hematopoietic transplant performed after onset of disease resulted in 42% mortality in SJL/J syngeneic transplants, 47% mortality in diseased DBA2 recipients restored with marrow from naive B6D2 donors, and 12% in diseased DBA2 recipients receiving marrow from B6D2 donors previously infected with TMEV. Delayed type hypersensitivity (DTH) to both virion and myelin proteins was decreased in surviving mice that underwent transplantation; however, CNS viral titers were significantly elevated compared with nontransplanted controls. We conclude that a functional immune system with appropriate T-cell responses are important in prevention of lethal cytopathic CNS effects from TMEV. Relevant to the clinical use of bone marrow transplantation, attempts to ablate the immune system in viral-mediated immune diseases or virus initiated autoimmune disease may have acute and lethal consequences. Our results raise concern about the attempted use of autologous hematopoietic transplantation in patients with MS, an autoimmune disease with a suspected virus etiology, particularly if the graft is aggressively depleted of lymphocytes. PMID- 10515898 TI - Long-term chimerism and B-cell function after bone marrow transplantation in patients with severe combined immunodeficiency with B cells: A single-center study of 22 patients. AB - We retrospectively analyzed the B-cell function and leukocyte chimerism of 22 patients with severe combined immunodeficiency with B cells (B(+) SCID) who survived more than 2 years after bone marrow transplantation (BMT) to determine the possible consequences of BMT procedures, leukocyte chimerism, and SCID molecular deficit on B-cell function outcome. Circulating T cells were of donor origin in all patients. In recipients of HLA-identical BMT (n = 5), monocytes were of host origin in 5 and B cells were of host origin in 4 and of mixed origin in 1. In recipients of HLA haploidentical T-cell-depleted BMT (n = 17), B cells and monocytes were of host origin in 14 and of donor origin in 3. Engraftment of B cells was found to be associated with normal B-cell function. In contrast, 10 of 18 patients with host B cells still require Ig substitution. Conditioning regimen (ie, 8 mg/kg busulfan and 200 mg/kg cyclophosphamide) was shown neither to promote B-cell and monocyte engraftment nor to affect B-cell function. Eight patients with B cells of host origin had normal B-cell function. Evidence for functional host B cells was further provided in 3 informative cases by Ig allotype determination and by the detection, in 5 studied cases, of host CD27(+) memory B cells as in age-matched controls. These results strongly suggest that, in some transplanted patients, host B cells can cooperate with donor T cells to fully mature in Ig-producing cells. PMID- 10515899 TI - Expression of cell-homologous genes of human herpesvirus-8 in human immunodeficiency virus-negative lymphoproliferative diseases. AB - Human herpesvirus-8 (HHV-8) genome encodes for genes homologous to human cellular genes such as interleukin-6 (IL-6), Cyclin-D, BCL-2, and IL-8 receptor (G-protein coupled receptor [GCR]). We used reverse transcriptase-polymerase chain reaction to study the expression of these viral genes in lymphoproliferative disorders associated with HHV-8 infection. None of these genes was expressed in 1 case of benign, localized Castleman's disease (CD), and only viral IL-6 and viral Cyclin D were transcribed in 2 cases of benign lymphadenopathies with giant germinal center hyperplasia and increased vascularity. In contrast, all 4 genes were transcribed in 1 case of multicentric CD of plasma cell type with aggressive clinical course and in 1 primary effusion lymphoma cell line. Our study provides the evidence that various HHV-8 genes, homologous to cellular genes involved in control of proliferation and apoptosis, may be differently expressed in different lymphoid disorders in vivo. PMID- 10515900 TI - Antifungal agents: mode of action, mechanisms of resistance, and correlation of these mechanisms with bacterial resistance. AB - The increased use of antibacterial and antifungal agents in recent years has resulted in the development of resistance to these drugs. The significant clinical implication of resistance has led to heightened interest in the study of antimicrobial resistance from different angles. Areas addressed include mechanisms underlying this resistance, improved methods to detect resistance when it occurs, alternate options for the treatment of infections caused by resistant organisms, and strategies to prevent and control the emergence and spread of resistance. In this review, the mode of action of antifungals and their mechanisms of resistance are discussed. Additionally, an attempt is made to discuss the correlation between fungal and bacterial resistance. Antifungals can be grouped into three classes based on their site of action: azoles, which inhibit the synthesis of ergosterol (the main fungal sterol); polyenes, which interact with fungal membrane sterols physicochemically; and 5-fluorocytosine, which inhibits macromolecular synthesis. Many different types of mechanisms contribute to the development of resistance to antifungals. These mechanisms include alteration in drug target, alteration in sterol biosynthesis, reduction in the intercellular concentration of target enzyme, and overexpression of the antifungal drug target. Although the comparison between the mechanisms of resistance to antifungals and antibacterials is necessarily limited by several factors defined in the review, a correlation between the two exists. For example, modification of enzymes which serve as targets for antimicrobial action and the involvement of membrane pumps in the extrusion of drugs are well characterized in both the eukaryotic and prokaryotic cells. PMID- 10515902 TI - New insights into human cryptosporidiosis. AB - Cryptosporidium parvum is an important cause of diarrhea worldwide. Cryptosporidium causes a potentially life-threatening disease in people with AIDS and contributes significantly to morbidity among children in developing countries. In immunocompetent adults, Cryptosporidium is often associated with waterborne outbreaks of acute diarrheal illness. Recent studies with human volunteers have indicated that Cryptosporidium is highly infectious. Diagnosis of infection with this parasite has relied on identification of acid-fast oocysts in stool; however, new immunoassays or PCR-based assays may increase the sensitivity of detection. Although the mechanism by which Cryptosporidium causes diarrhea is still poorly understood, the parasite and the immune response to it probably combine to impair absorption and enhance secretion within the intestinal tract. Important genetic studies suggest that humans can be infected by at least two genetically distinct types of Cryptosporidium, which may vary in virulence. This may, in part, explain the clinical variability seen in patients with cryptosporidiosis. PMID- 10515906 TI - Infectious coryza: overview of the disease and new diagnostic options. AB - Infectious coryza is a well-recognized and commonly encountered upper respiratory tract disease of chickens that is caused by the bacterium Haemophilus paragallinarum. The occurrence of recent outbreaks in North America has emphasized that the disease can be significant in meat chickens as well as layer chickens. In developing countries, coryza is commonly complicated by the presence of a range of other infections, resulting in severe disease and significant economic losses. Unusual forms of the disease, involving arthritis and septicemia, again associated with the presence of other pathogens, have been found in South America. Newly recognized bacteria such as Ornithobacterium rhinotracheale and phenotypic variant forms of both H. paragallinarum and close relatives (variant in that they no longer require V-factor for growth in vitro) have increased the difficulty associated with diagnosing the disease. There have been suggestions in both South America and South Africa that new serovars or serovar variants, associated with unusual clinical manifestations and causing vaccine failures, are emerging. Definitive evidence to confirm or deny the role of these "variants" in vaccine failures is currently not available. A new DNA based diagnostic technique, involving PCR, has been recently described and will greatly assist in the diagnosis of infectious coryza. PMID- 10515905 TI - Methods for subtyping and molecular comparison of human viral genomes. AB - The development over the past two decades of molecular methods for manipulation of RNA and DNA has afforded molecular virologists the ability to study viral genomes in detail that has heretofore not been possible. There are many molecular techniques now available for typing and subtyping of viruses. The available methods include restriction fragment length polymorphism analysis, Southern blot analysis, oligonucleotide fingerprint analysis, reverse hybridization, DNA enzyme immunoassay, RNase protection analysis, single-strand conformation polymorphism analysis, heteroduplex mobility assay, nucleotide sequencing, and genome segment length polymorphism analysis. The methods have certain advantages and disadvantages which should be considered in their application to specific viruses or for specific purposes. These techniques are likely to become more widely used in the future for epidemiologic studies and for investigations into the pathophysiology of virus infections. PMID- 10515901 TI - Q fever. AB - Q fever is a zoonosis with a worldwide distribution with the exception of New Zealand. The disease is caused by Coxiella burnetii, a strictly intracellular, gram-negative bacterium. Many species of mammals, birds, and ticks are reservoirs of C. burnetii in nature. C. burnetii infection is most often latent in animals, with persistent shedding of bacteria into the environment. However, in females intermittent high-level shedding occurs at the time of parturition, with millions of bacteria being released per gram of placenta. Humans are usually infected by contaminated aerosols from domestic animals, particularly after contact with parturient females and their birth products. Although often asymptomatic, Q fever may manifest in humans as an acute disease (mainly as a self-limited febrile illness, pneumonia, or hepatitis) or as a chronic disease (mainly endocarditis), especially in patients with previous valvulopathy and to a lesser extent in immunocompromised hosts and in pregnant women. Specific diagnosis of Q fever remains based upon serology. Immunoglobulin M (IgM) and IgG antiphase II antibodies are detected 2 to 3 weeks after infection with C. burnetii, whereas the presence of IgG antiphase I C. burnetii antibodies at titers of >/=1:800 by microimmunofluorescence is indicative of chronic Q fever. The tetracyclines are still considered the mainstay of antibiotic therapy of acute Q fever, whereas antibiotic combinations administered over prolonged periods are necessary to prevent relapses in Q fever endocarditis patients. Although the protective role of Q fever vaccination with whole-cell extracts has been established, the population which should be primarily vaccinated remains to be clearly identified. Vaccination should probably be considered in the population at high risk for Q fever endocarditis. PMID- 10515903 TI - Plant products as antimicrobial agents. AB - The use of and search for drugs and dietary supplements derived from plants have accelerated in recent years. Ethnopharmacologists, botanists, microbiologists, and natural-products chemists are combing the Earth for phytochemicals and "leads" which could be developed for treatment of infectious diseases. While 25 to 50% of current pharmaceuticals are derived from plants, none are used as antimicrobials. Traditional healers have long used plants to prevent or cure infectious conditions; Western medicine is trying to duplicate their successes. Plants are rich in a wide variety of secondary metabolites, such as tannins, terpenoids, alkaloids, and flavonoids, which have been found in vitro to have antimicrobial properties. This review attempts to summarize the current status of botanical screening efforts, as well as in vivo studies of their effectiveness and toxicity. The structure and antimicrobial properties of phytochemicals are also addressed. Since many of these compounds are currently available as unregulated botanical preparations and their use by the public is increasing rapidly, clinicians need to consider the consequences of patients self-medicating with these preparations. PMID- 10515908 TI - Opening the iron box: transcriptional metalloregulation by the Fur protein. PMID- 10515909 TI - Characterization of a new sigma-K-dependent peptidoglycan hydrolase gene that plays a role in Bacillus subtilis mother cell lysis. AB - Bacillus subtilis produces a 30-kDa peptidoglycan hydrolase, CwlH, during the late sporulation phase. Disruption of yqeE led to a complete loss of CwlH formation, indicating the identity of yqeE with cwlH. Northern blot analysis of cwlH revealed a 0.8-kb transcript after 6 to 7.5 h for the wild-type strain but not for the sigma(F), sigma(E), sigma(G), and sigma(K) mutants. Expression of the sigma(K)-dependent cwlH gene depended on gerE. Primer extension analysis also suggested that cwlH is transcribed by Esigma(K) RNA polymerase. CwlH produced in Escherichia coli harboring a cwlH plasmid is an N-acetylmuramoyl-L-alanine amidase (EC 3.5.1.28) and exhibited an optimum pH of 7.0 and high-level binding to the B. subtilis cell wall. A cwlC cwlH double mutation led to a lack of mother cell lysis even after 7 days of incubation in DSM medium, but the single mutations led to mother cell lysis after 24 h. PMID- 10515910 TI - Genetic and biochemical characterization of a high-affinity betaine uptake system (BusA) in Lactococcus lactis reveals a new functional organization within bacterial ABC transporters. AB - The cytoplasmic accumulation of exogenous betaine stimulates the growth of Lactococcus lactis cultivated under hyperosmotic conditions. We report that L. lactis possesses a single betaine transport system that belongs to the ATP binding cassette (ABC) superfamily of transporters. Through transposon mutagenesis, a mutant deficient in betaine transport was isolated. We identified two genes, busAA and busAB, grouped in an operon, busA (betaine uptake system). The transcription of busA is strongly regulated by the external osmolality of the medium. The busAA gene codes for the ATP-binding protein. busAB encodes a 573 residue polypeptide which presents two striking features: (i) a fusion between the regions encoding the transmembrane domain (TMD) and the substrate-binding domain (SBD) and (ii) a swapping of the SBD subdomains when compared to the Bacillus subtilis betaine-binding protein, OpuAC. BusA of L. lactis displays a high affinity towards betaine (K(m) = 1.7 microM) and is an osmosensor whose activity is tightly regulated by external osmolality, leading the betaine uptake capacity of L. lactis to be under dual control at the biochemical and genetic levels. A protein presenting the characteristics predicted for BusAB was detected in the membrane fraction of L. lactis. The fusion between the TMD and the SBD is the first example of a new organization within prokaryotic ABC transporters. PMID- 10515911 TI - A plant-type (beta-class) carbonic anhydrase in the thermophilic methanoarchaeon Methanobacterium thermoautotrophicum. AB - Carbonic anhydrase, a zinc enzyme catalyzing the interconversion of carbon dioxide and bicarbonate, is nearly ubiquitous in the tissues of highly evolved eukaryotes. Here we report on the first known plant-type (beta-class) carbonic anhydrase in the archaea. The Methanobacterium thermoautotrophicum DeltaH cab gene was hyperexpressed in Escherichia coli, and the heterologously produced protein was purified 13-fold to apparent homogeneity. The enzyme, designated Cab, is thermostable at temperatures up to 75 degrees C. No esterase activity was detected with p-phenylacetate as the substrate. The enzyme is an apparent tetramer containing approximately one zinc per subunit, as determined by plasma emission spectroscopy. Cab has a CO(2) hydration activity with a k(cat) of 1.7 x 10(4) s(-1) and K(m) for CO(2) of 2.9 mM at pH 8.5 and 25 degrees C. Western blot analysis indicates that Cab (beta class) is expressed in M. thermoautotrophicum; moreover, a protein cross-reacting to antiserum raised against the gamma carbonic anhydrase from Methanosarcina thermophila was detected. These results show that beta-class carbonic anhydrases extend not only into the Archaea domain but also into the thermophilic prokaryotes. PMID- 10515914 TI - Multiple roles for TnpI recombinase in regulation of Tn5401 transposition in Bacillus thuringiensis. AB - Tn5401 is a class II transposable element derived from the gram-positive bacterium Bacillus thuringiensis. The 4,837-bp transposon encodes a Tn3-like transposase (TnpA) and an integrase-like recombinase (TnpI) and is notable for its unusually long 53-bp terminal inverted repeats (TIRs). The tnpA and tnpI genes are transcribed from a common promoter, designated P(R), that is subject to negative regulation by TnpI. The TIRs of Tn5401 each contain a 38-bp sequence that can be aligned with the 38- to 40-bp TIR sequences of Tn3-like transposons and an adjacent 12-bp sequence that binds TnpI. This unique juxtaposition of TnpA and TnpI binding sites suggests that TnpI may regulate the binding or catalytic activity of TnpA. The results of the present study indicate that TnpI, in addition to functioning as a site-specific recombinase and as a transcriptional repressor, is required for TnpA binding to the TIRs of Tn5401. PMID- 10515913 TI - A second operator is involved in Pseudomonas aeruginosa elastase (lasB) activation. AB - Pseudomonas aeruginosa LasB elastase gene (lasB) transcription is controlled by the two-component quorum-sensing system of LasR, and the autoinducer, 3OC(12)-HSL (N-3-[oxododecanoyl]homoserine lactone). LasR and 3OC(12)-HSL-mediated lasB activation requires a functional operator sequence (OP1) in the lasB promoter region. Optimal activation of lasB, however, requires a second sequence of 70% identity to OP1, named OP2, located 43 bp upstream of OP1. In this study, we used sequence substitutions and insertion mutations in lasBp-lacZ fusion plasmids to explore the role of OP2 in lasB activation. Our results demonstrate that (i) OP1 and OP2 synergistically mediate lasB activation; (ii) OP2, like OP1, responds to LasR and 3OC(12)-HSL; and (iii) the putative autoinducer-binding domain of LasR is not required for synergistic activation from OP1 and OP2. PMID- 10515915 TI - rpoS function is essential for bgl silencing caused by C-terminally truncated H NS in Escherichia coli. AB - From evolutionary and physiological viewpoints, the Escherichia coli bgl operon is intriguing because its expression is silent (Bgl(-) phenotype), at least under several laboratory conditions. H-NS, a nucleoid protein, is known as a DNA binding protein involved in bgl silencing. However, we previously found that bgl expression is still silent in a certain subset of hns mutations, each of which results in a defect in its DNA-binding ability. Based on this fact, we proposed a model in which a postulated DNA-binding protein(s) has an adapter function by interacting with both the cis-acting element of the bgl promoter and the mutated H-NS. To identify such a presumed adapter molecule, we attempted to isolate mutants exhibiting the Bgl(+) phenotype in the background of hns60, encoding the mutant H-NS protein lacking the DNA-binding domain by random insertion mutagenesis with the mini-Tn10cam transposon. These isolated mutations were mapped to five loci on the chromosome. Among these loci, three appeared to be leuO, hns, and bglJ, which were previously characterized, while the other two were novel. Genetic analysis revealed that the two insertions are within the rpoS gene and in front of the lrhA gene, respectively. The former encodes the stationary-phase-specific sigma factor, sigma(S), and the latter encodes a LysR like DNA-binding protein. It was found that sigma(S) is defective in both types of mutant cells. These results showed that the rpoS function is involved in the mechanism underlying bgl silencing, at least in the hns60 background used in this study. We also examined whether the H-NS homolog StpA has such an adapter function, as was previously proposed. Our results did not support the idea that StpA has an adapter function in the genetic background used. PMID- 10515912 TI - Cloning and sequence analysis of two Pseudomonas flavoprotein xenobiotic reductases. AB - The genes encoding flavin mononucleotide-containing oxidoreductases, designated xenobiotic reductases, from Pseudomonas putida II-B and P. fluorescens I-C that removed nitrite from nitroglycerin (NG) by cleavage of the nitroester bond were cloned, sequenced, and characterized. The P. putida gene, xenA, encodes a 39,702 Da monomeric, NAD(P)H-dependent flavoprotein that removes either the terminal or central nitro groups from NG and that reduces 2-cyclohexen-1-one but did not readily reduce 2,4,6-trinitrotoluene (TNT). The P. fluorescens gene, xenB, encodes a 37,441-Da monomeric, NAD(P)H-dependent flavoprotein that exhibits fivefold regioselectivity for removal of the central nitro group from NG and that transforms TNT but did not readily react with 2-cyclohexen-1-one. Heterologous expression of xenA and xenB was demonstrated in Escherichia coli DH5alpha. The transcription initiation sites of both xenA and xenB were identified by primer extension analysis. BLAST analyses conducted with the P. putida xenA and the P. fluorescens xenB sequences demonstrated that these genes are similar to several other bacterial genes that encode broad-specificity flavoprotein reductases. The prokaryotic flavoprotein reductases described herein likely shared a common ancestor with old yellow enzyme of yeast, a broad-specificity enzyme which may serve a detoxification role in antioxidant defense systems. PMID- 10515907 TI - Molecular typing of Borrelia burgdorferi sensu lato: taxonomic, epidemiological, and clinical implications. AB - Borrelia burgdorferi sensu lato, the spirochete that causes human Lyme borreliosis (LB), is a genetically and phenotypically divergent species. In the past several years, various molecular approaches have been developed and used to determine the phenotypic and genetic heterogeneity within the LB-related spirochetes and their potential association with distinct clinical syndromes. These methods include serotyping, multilocus enzyme electrophoresis, DNA-DNA reassociation analysis, rRNA gene restriction analysis (ribotyping), pulsed-field gel electrophoresis, plasmid fingerprinting, randomly amplified polymorphic DNA fingerprinting analysis, species-specific PCR and PCR-based restriction fragment length polymorphism (RFLP) analysis, and sequence analysis of 16S rRNA and other conserved genes. On the basis of DNA-DNA reassociation analysis, 10 different Borrelia species have been described within the B. burgdorferi sensu lato complex: B. burgdorferi sensu stricto, Borrelia garinii, Borrelia afzelii, Borrelia japonica, Borrelia andersonii, Borrelia valaisiana, Borrelia lusitaniae, Borrelia tanukii, Borrelia turdi, and Borrelia bissettii sp. nov. To date, only B. burgdorferi sensu stricto, B. garinii, and B. afzelii are well known to be responsible for causing human disease. Different Borrelia species have been associated with distinct clinical manifestations of LB. In addition, Borrelia species are differentially distributed worldwide and may be maintained through different transmission cycles in nature. In this paper, the molecular methods used for typing of B. burgdorferi sensu lato are reviewed. The current taxonomic status of B. burgdorferi sensu lato and its epidemiological and clinical implications, especiallly correlation between the variable clinical presentations and the infecting Borrelia species, are discussed in detail. PMID- 10515916 TI - Mutation analysis of the 5' untranslated region of the cold shock cspA mRNA of Escherichia coli. AB - The mRNA for CspA, a major cold shock protein in Escherichia coli, contains an unusually long (159 bases) 5' untranslated region (5'-UTR), and its stability has been shown to play a major role in cold shock induction of CspA. The 5'-UTR of the cspA mRNA has a negative effect on its expression at 37 degrees C but has a positive effect upon cold shock. In this report, a series of cspA-lacZ fusions having a 26- to 32-base deletion in the 5'-UTR were constructed to examine the roles of specific regions within the 5'-UTR in cspA expression. It was found that none of the deletion mutations had significant effects on the stability of mRNA at both 37 and 15 degrees C. However, two mutations (Delta56-86 and Delta86-117) caused a substantial increase of beta-galactosidase activity at 37 degrees C, indicating that the deleted regions contain a negative cis element(s) for translation. A mutation (Delta2-27) deleting the highly conserved cold box sequence had little effect on cold shock induction of beta-galactosidase. Interestingly, three mutations (Delta28-55, Delta86-117, and Delta118-143) caused poor cold shock induction of beta-galactosidase. In particular, the Delta118-143 mutation reduced the translation efficiency of the cspA mRNA to less than 10% of that of the wild-type construct. The deleted region contains a 13-base sequence named upstream box (bases 123 to 135), which is highly conserved in cspA, cspB, cspG, and cspI, and is located 11 bases upstream of the Shine-Dalgarno (SD) sequence. The upstream box might be another cis element involved in translation efficiency of the cspA mRNA in addition to the SD sequence and the downstream box sequence. The relationship between the mRNA secondary structure and translation efficiency is discussed. PMID- 10515917 TI - Analysis of protein synthesis rates after initiation of chromosome replication in Escherichia coli. AB - The aim of this study was to investigate whether the synthesis rates of some proteins change after the initiation of replication in Escherichia coli. An intR1 strain, in which chromosome replication is under the control of an R1 replicon integrated into an inactivated oriC, was used to synchronize chromosome replication, and the rates of protein synthesis were analyzed by two-dimensional polyacrylamide gel electrophoresis of pulse-labeled proteins. Computerized image analysis was used to search for proteins whose expression levels changed at least threefold after initiation of a single round of chromosome replication, which revealed 7 out of about 1,000 detected proteins. The various synthesis rates of three of these proteins turned out to be caused by unbalanced growth and the synthesis of one protein was suppressed in the intR1 strain. The rates of synthesis of the remaining three could be correlated only to the synchronous initiation of replication. These three proteins were analyzed by peptide mass mapping and appeared to be the products of the dps, gapA, and pyrI genes. Thus, the expression of the vast majority of proteins is not influenced by the state of chromosome replication, and a possible role of the replication-associated expression changes of the three identified proteins in the cell cycle is not clear. PMID- 10515904 TI - Antifungal activities of antineoplastic agents: Saccharomyces cerevisiae as a model system to study drug action. AB - Recent evolutionary studies reveal that microorganisms including yeasts and fungi are more closely related to mammals than was previously appreciated. Possibly as a consequence, many natural-product toxins that have antimicrobial activity are also toxic to mammalian cells. While this makes it difficult to discover antifungal agents without toxic side effects, it also has enabled detailed studies of drug action in simple genetic model systems. We review here studies on the antifungal actions of antineoplasmic agents. Topics covered include the mechanisms of action of inhibitors of topoisomerases I and II; the immunosuppressants rapamycin, cyclosporin A, and FK506; the phosphatidylinositol 3-kinase inhibitor wortmannin; the angiogenesis inhibitors fumagillin and ovalicin; the HSP90 inhibitor geldanamycin; and agents that inhibit sphingolipid metabolism. In general, these natural products inhibit target proteins conserved from microorganisms to humans. These studies highlight the potential of microorganisms as screening tools to elucidate the mechanisms of action of novel pharmacological agents with unique effects against specific mammalian cell types, including neoplastic cells. In addition, this analysis suggests that antineoplastic agents and derivatives might find novel indications in the treatment of fungal infections, for which few agents are presently available, toxicity remains a serious concern, and drug resistance is emerging. PMID- 10515919 TI - In vitro characterization of peptidoglycan-associated lipoprotein (PAL) peptidoglycan and PAL-TolB interactions. AB - The Tol-peptidoglycan-associated lipoprotein (PAL) system of Escherichia coli is a multiprotein complex of the envelope involved in maintaining outer membrane integrity. PAL and the periplasmic protein TolB, two components of this complex, are interacting with each other, and they have also been reported to interact with OmpA and the major lipoprotein, two proteins interacting with the peptidoglycan. All these interactions suggest a role of the Tol-PAL system in anchoring the outer membrane to the peptidoglycan. Therefore, we were interested in better understanding the interaction between PAL and the peptidoglycan. We designed an in vitro interaction assay based on the property of purified peptidoglycan to be pelleted by ultracentrifugation. Using this assay, we showed that a purified PAL protein interacted in vitro with pure peptidoglycan. A peptide competition experiment further demonstrated that the region from residues 89 to 130 of PAL was sufficient to bind the peptidoglycan. Moreover, the fact that this same region of PAL was also binding to TolB suggested that these two interactions were exclusive. Indeed, the TolB-PAL complex appeared not to be associated with the peptidoglycan. This led us to the conclusion that PAL may exist in two forms in the cell envelope, one bound to TolB and the other bound to the peptidoglycan. PMID- 10515918 TI - Characterization of MexT, the regulator of the MexE-MexF-OprN multidrug efflux system of Pseudomonas aeruginosa. AB - We investigated the regulation of the MexEF-OprN multidrug efflux system of Pseudomonas aeruginosa, which is overexpressed in nfxC-type mutants and confers resistance to quinolones, chloramphenicol and trimethoprim. Sequencing of the DNA region upstream of the mexEF-oprN operon revealed the presence of an open reading frame (ORF) of 304 amino acids encoding a LysR-type transcriptional activator, termed MexT. By using T7-polymerase, a 34-kDa protein was expressed in Escherichia coli from a plasmid carrying the mexT gene. Expression of a mexE::lacZ fusion was 10-fold higher in nfxC-type mutants than in the wild-type strain; however, transcription of mexT as well as the mexT DNA region was unchanged. Located adjacent to mexT but transcribed in opposite direction, the beginning of an ORF termed qrh (quinone oxidoreductase homologue) was identified. Expression of a qrh::lacZ fusion was also found to be activated by MexT. Further, we present evidence for coregulation at the transcriptional and the posttranscriptional level between the MexEF-OprN efflux system and the OprD porin responsible for cross-resistance of nfxC-type mutants to carbapenem antibiotics. PMID- 10515920 TI - Regulation of the transposase of Tn4652 by the transposon-encoded protein TnpC. AB - Transposition is a DNA reorganization reaction potentially deleterious for the host. The frequency of transposition is limited by the amount of transposase. Therefore, strict regulation of a transposase is required to keep control over the destructive multiplication of the mobile element. We have shown previously that the expression of the transposase (tnpA) of the Pseudomonas putida PaW85 transposon Tn4652 is positively affected by integration host factor. Here, we present evidence that the amount of the transposase of Tn4652 in P. putida cells is controlled by the transposon-encoded protein (TnpC). Sequence analysis of the 120-amino-acid-long TnpC, coded just downstream of the tnpA gene, showed that it has remarkable similarity to the putative polypeptide encoded by the mercury resistance transposon Tn5041. As determined by quantitative Western blot analysis, the abundance of TnpA was reduced up to 10-fold in the intact tnpC background. In vivo experiments using transcriptional and translational fusions of the tnpA gene and the reporter gene gusA indicated that TnpC operates in the regulation of the transposase of Tn4652 at the post-transcriptional level. PMID- 10515921 TI - Induction of beta-lactamase influences the course of development in Myxococcus xanthus. AB - Myxococcus xanthus is a gram-negative bacterium that develops in response to starvation on a solid surface. The cells assemble into multicellular aggregates in which they differentiate from rod-shaped cells into spherical, environmentally resistant spores. Previously, we have shown that the induction of beta-lactamase is associated with starvation-independent sporulation in liquid culture (K. A. O'Connor and D. R. Zusman, Mol. Microbiol. 24:839-850, 1997). In this paper, we show that the chromosomally encoded beta-lactamase of M. xanthus is autogenously induced during development. The specific activity of the enzyme begins to increase during aggregation, before spores are detectable. The addition of inducers of beta-lactamase in M. xanthus, such as ampicillin, D-cycloserine, and phosphomycin, accelerates the onset of aggregation and sporulation in developing populations of cells. In addition, the exogenous induction of beta-lactamase allows M. xanthus to fruit on media containing concentrations of nutrients that are normally too high to support development. We propose that the induction of beta-lactamase is an integral step in the development of M. xanthus and that this induction is likely to play a role in aggregation and in the restructuring of peptidoglycan which occurs during the differentiation of spores. In support of this hypothesis, we show that exogenous induction of beta-lactamase can rescue aggregation and sporulation of certain mutants. Fruiting body spores from a rescued mutant are indistinguishable from wild-type fruiting body spores when examined by transmission electron microscopy. These results show that the signal transduction pathway leading to the induction of beta-lactamase plays an important role in aggregation and sporulation in M. xanthus. PMID- 10515922 TI - Hybrid motor with H(+)- and Na(+)-driven components can rotate Vibrio polar flagella by using sodium ions. AB - The bacterial flagellar motor is a molecular machine that converts ion flux across the membrane into flagellar rotation. The coupling ion is either a proton or a sodium ion. The polar flagellar motor of the marine bacterium Vibrio alginolyticus is driven by sodium ions, and the four protein components, PomA, PomB, MotX, and MotY, are essential for motor function. Among them, PomA and PomB are similar to MotA and MotB of the proton-driven motors, respectively. PomA shows greatest similarity to MotA of the photosynthetic bacterium Rhodobacter sphaeroides. MotA is composed of 253 amino acids, the same length as PomA, and 40% of its residues are identical to those of PomA. R. sphaeroides MotB has high similarity only to the transmembrane region of PomB. To examine whether the R. sphaeroides motor genes can function in place of the pomA and pomB genes of V. alginolyticus, we constructed plasmids including both motA and motB or motA alone and transformed them into missense and null pomA-paralyzed mutants of V. alginolyticus. The transformants from both strains showed restored motility, although the swimming speeds were low. On the other hand, pomB mutants were not restored to motility by any plasmid containing motA and/or motB. Next, we tested which ions (proton or sodium) coupled to the hybrid motor function. The motor did not work in sodium-free buffer and was inhibited by phenamil and amiloride, sodium motor-specific inhibitors, but not by a protonophore. Thus, we conclude that the proton motor component, MotA, of R. sphaeroides can generate torque by coupling with the sodium ion flux in place of PomA of V. alginolyticus. PMID- 10515923 TI - Ras signaling is required for serum-induced hyphal differentiation in Candida albicans. AB - Serum induces Candida albicans to make a rapid morphological change from the yeast cell form to hyphae. Contrary to the previous reports, we found that serum albumin does not play a critical role in this morphological change. Instead, a filtrate (molecular mass, <1 kDa) devoid of serum albumin induces hyphae. To study genes controlling this response, we have isolated the RAS1 gene from C. albicans by complementation. The Candida Ras1 protein, like Ras1 and Ras2 of Saccharomyces cerevisiae, has a long C-terminal extension. Although RAS1 appears to be the only RAS gene present in the C. albicans genome, strains homozygous for a deletion of RAS1 (ras1-2/ras1-3) are viable. The Candida ras1-2/ras1-3 mutant fails to form germ tubes and hyphae in response to serum or to a serum filtrate but does form pseudohyphae. Moreover, strains expressing the dominant active RAS1(V13) allele manifest enhanced hyphal growth, whereas those expressing a dominant negative RAS1(A16) allele show reduced hyphal growth. These data show that low-molecular-weight molecules in serum induce hyphal differentiation in C. albicans through a Ras-mediated signal transduction pathway. PMID- 10515924 TI - Functional analysis of glycosyltransferase genes from Lactococcus lactis and other gram-positive cocci: complementation, expression, and diversity. AB - Sixteen exopolysaccharide (EPS)-producing Lactococcus lactis strains were analyzed for the chemical compositions of their EPSs and the locations, sequences, and organization of the eps genes involved in EPS biosynthesis. This allowed the grouping of these strains into three major groups, representatives of which were studied in detail. Previously, we have characterized the eps gene cluster of strain NIZO B40 (group I) and determined the function of three of its glycosyltransferase (GTF) genes. Fragments of the eps gene clusters of strains NIZO B35 (group II) and NIZO B891 (group III) were cloned, and these encoded the NIZO B35 priming galactosyltransferase, the NIZO B891 priming glucosyltransferase, and the NIZO B891 galactosyltransferase involved in the second step of repeating-unit synthesis. The NIZO B40 priming glucosyltransferase gene epsD was replaced with an erythromycin resistance gene, and this resulted in loss of EPS production. This epsD deletion was complemented with priming GTF genes from gram-positive organisms with known function and substrate specificity. Although no EPS production was found with priming galactosyltransferase genes from L. lactis or Streptococcus thermophilus, complementation with priming glucosyltransferase genes involved in L. lactis EPS and Streptococcus pneumoniae capsule biosynthesis could completely restore or even increase EPS production in L. lactis. PMID- 10515925 TI - Unraveling the function of glycosyltransferases in Streptococcus thermophilus Sfi6. AB - Streptococcus thermophilus Sfi6 produces a texturizing exopolysaccharide (EPS) consisting of a -->3)[alpha-D-Galp-(1-->6)]-beta-D-Glcp-(1-->3)-alpha-D-GalpNAc (1--> 3)-beta-D-Galp-(1--> repeating unit. We previously identified and analyzed a 14.5-kb gene cluster from S. thermophilus Sfi6 consisting of 13 genes responsible for its EPS production. Within this gene cluster, we found a central region of genes (epsE, epsF, epsG, and epsI) that showed similarity to glycosyltransferases. In this study, we investigated the sugar specificity of these enzymes. EpsE catalyzes the first step in the biosynthesis of the EPS repeating unit. It exhibits phosphogalactosyltransferase activity and transfers galactose onto the lipophilic carrier. The second step is fulfilled by EpsG, which transfers an alpha-N-acetylgalactosamine onto the first beta-galactoside. The activity of EpsF was determined by characterizing the EPS produced by an S. thermophilus epsF deletion mutant. This EPS consisted of the monosaccharides Gal, Glc, and GalNAc in an approximately equimolar ratio, thus suggesting that epsF codes for the branching galactosyltransferase. epsI probably codes for the beta 1,3-glucosyltransferase, since it is the only glycosyltransferase to which no gene has been assigned and it exhibits similarity to other beta glycosyltransferases. EpsE shows the conserved features of phosphoglycosyltransferases, whereas EpsF and EpsG exhibit the primary structure of alpha-glycosyltransferases, belonging to glycosyltransferase family 4, whose members are conserved in all major phylogenetic lineages, including the Archaea and Eukaryota. PMID- 10515926 TI - Growth phase-dependent variation in protein composition of the Escherichia coli nucleoid. AB - The genome DNA of Escherichia coli is associated with about 10 DNA-binding structural proteins, altogether forming the nucleoid. The nucleoid proteins play some functional roles, besides their structural roles, in the global regulation of such essential DNA functions as replication, recombination, and transcription. Using a quantitative Western blot method, we have performed for the first time a systematic determination of the intracellular concentrations of 12 species of the nucleoid protein in E. coli W3110, including CbpA (curved DNA-binding protein A), CbpB (curved DNA-binding protein B, also known as Rob [right origin binding protein]), DnaA (DNA-binding protein A), Dps (DNA-binding protein from starved cells), Fis (factor for inversion stimulation), Hfq (host factor for phage Q(beta)), H-NS (histone-like nucleoid structuring protein), HU (heat-unstable nucleoid protein), IciA (inhibitor of chromosome initiation A), IHF (integration host factor), Lrp (leucine-responsive regulatory protein), and StpA (suppressor of td mutant phenotype A). Intracellular protein levels reach a maximum at the growing phase for nine proteins, CbpB (Rob), DnaA, Fis, Hfq, H-NS, HU, IciA, Lrp, and StpA, which may play regulatory roles in DNA replication and/or transcription of the growth-related genes. In descending order, the level of accumulation, calculated in monomers, in growing E. coli cells is Fis, Hfq, HU, StpA, H-NS, IHF*, CbpB (Rob), Dps*, Lrp, DnaA, IciA, and CbpA* (stars represent the stationary-phase proteins). The order of abundance, in descending order, in the early stationary phase is Dps*, IHF*, HU, Hfq, H-NS, StpA, CbpB (Rob), DnaA, Lrp, IciA, CbpA, and Fis, while that in the late stationary phase is Dps*, IHF*, Hfq, HU, CbpA*, StpA, H-NS, CbpB (Rob), DnaA, Lrp, IciA, and Fis. Thus, the major protein components of the nucleoid change from Fis and HU in the growing phase to Dps in the stationary phase. The curved DNA-binding protein, CbpA, appears only in the late stationary phase. These changes in the composition of nucleoid associated proteins in the stationary phase are accompanied by compaction of the genome DNA and silencing of the genome functions. PMID- 10515927 TI - Campylobacter jejuni contains two fur homologs: characterization of iron responsive regulation of peroxide stress defense genes by the PerR repressor. AB - Expression of the peroxide stress genes alkyl hydroperoxide reductase (ahpC) and catalase (katA) of the microaerophile Campylobacter jejuni is repressed by iron. Whereas iron repression in gram-negative bacteria is usually carried out by the Fur protein, previous work showed that this is not the case in C. jejuni, as these genes are still iron repressed in a C. jejuni fur mutant. An open reading frame encoding a Fur homolog (designated PerR for "peroxide stress regulator") was identified in the genome sequence of C. jejuni. The perR gene was disrupted by a kanamycin resistance cassette in C. jejuni wild-type and fur mutant strains. Subsequent characterization of the C. jejuni perR mutants showed derepressed expression of both AhpC and KatA at a much higher level than that obtained by iron limitation, suggesting that expression of these genes is controlled by other regulatory factors in addition to the iron level. Other iron-regulated proteins were not affected by the perR mutation. The fur perR double mutant showed derepressed expression of known iron-repressed genes. Further phenotypic analysis of the perR mutant, fur mutant, and the fur perR double mutant showed that the perR mutation made C. jejuni hyperresistant to peroxide stress caused by hydrogen peroxide and cumene hydroperoxide, a finding consistent with the high levels of KatA and AhpC expression, and showed that these enzymes were functional. Quantitative analysis of KatA expression showed that both the perR mutation and the fur mutation had profound effects on catalase activity, suggesting additional non-iron-dependent regulation of KatA and, by inference, AhpC. The PerR protein is a functional but nonhomologous substitution for the OxyR protein, which regulates peroxide stress genes in other gram-negative bacteria. Regulation of peroxide stress genes by a Fur homolog has recently been described for the gram positive bacterium Bacillus subtilis. C. jejuni is the first gram-negative bacterium where non-OxyR regulation of peroxide stress genes has been described and characterized. PMID- 10515928 TI - Arg-52 in the melibiose carrier of Escherichia coli is important for cation coupled sugar transport and participates in an intrahelical salt bridge. AB - Arg-52 of the Escherichia coli melibiose carrier was replaced by Ser (R52S), Gln (R52Q), or Val (R52V). While the level of carrier in the membrane for each mutant remained similar to that for the wild type, analysis of melibiose transport showed an uncoupling of proton cotransport and a drastic reduction in Na(+) coupled transport. Second-site revertants were selected on MacConkey plates containing melibiose, and substitutions were found at nine distinct locations in the carrier. Eight revertant substitutions were isolated from the R52S strain: Asp-19-->Gly, Asp-55-->Asn, Pro-60-->Gln, Trp-116-->Arg, Asn-244-->Ser, Ser-247- >Arg, Asn-248-->Lys, and Ile-352-->Val. Two revertants were also isolated from the R52V strain: Trp-116-->Arg and Thr-338-->Arg revertants. The R52Q strain yielded an Asp-55-->Asn substitution and a first-site revertant, Lys-52 (R52K). The R52K strain had transport properties similar to those of the wild type. Analysis of melibiose accumulation showed that proton-driven accumulation was still defective in the second-site revertant strains, and only the Trp-116-->Arg, Ser-247-->Arg, and Asn-248-->Lys revertants regained significant Na(+)-coupled accumulation. In general, downhill melibiose transport in the presence of Na(+) was better in the revertant strains than in the parental mutants. Three revertant strains, Asp-19-->Gly, Asp-55-->Asn, and Thr-338-->Arg strains, required a high Na(+) concentration (100 mM) for maximal activity. Kinetic measurements showed that the N248K and W116R revertants lowered the K(m) for melibiose, while other revertants restored transport velocity. We suggest that the insertion of positive charges on membrane helices is compensating for the loss of Arg-52 and that helix II is close to helix IV and VII. We also suggest that Arg-52 is salt bridged to Asp-55 (helix II) and Asp-19 (helix I). PMID- 10515929 TI - Ferric enterochelin transport in Yersinia enterocolitica: molecular and evolutionary aspects. AB - Yersinia enterocolitica is well equipped for siderophore piracy, encompassing the utilization of siderophores such as ferrioxamine, ferrichrome, and ferrienterochelin. In this study, we report on the molecular and functional characterization of the Yersinia fep-fes gene cluster orthologous to the Escherichia coli ferrienterochelin transport genes (fepA, fepDGC, and fepB) and the esterase gene fes. In vitro transcription-translation analysis identified polypeptides of 30 and 35 kDa encoded by fepC and fes, respectively. A frameshift mutation within the fepA gene led to expression of a truncated polypeptide of 40 kDa. The fepD, fepG, and fes genes of Y. enterocolitica were shown to complement corresponding E. coli mutants. Insertional mutagenesis of fepD or fes genes abrogates enterochelin-supported growth of Y. enterocolitica on iron-chelated media. In contrast to E. coli, the fep-fes gene cluster in Y. enterocolitica consists solely of genes required for uptake and utilization of enterochelin (fep) and not of enterochelin synthesis genes such as entF. By Southern hybridization, fepDGC and fes sequences could be detected in Y. enterocolitica biotypes IB, IA, and II but not in biotype IV strains, Yersinia pestis, and Yersinia pseudotuberculosis strains. According to sequence alignment data and the coherent structure of the Yersinia fep-fes gene cluster, we suggest early genetic divergence of ferrienterochelin uptake determinants among species of the family Enterobacteriaceae. PMID- 10515931 TI - Initial reactions in anaerobic oxidation of m-xylene by the denitrifying bacterium Azoarcus sp. strain T. AB - The initial enzymatic steps in anaerobic m-xylene oxidation were studied in Azoarcus sp. strain T, a denitrifying bacterium capable of mineralizing m-xylene via 3-methylbenzoate. Permeabilized cells of m-xylene-grown Azoarcus sp. strain T catalyzed the addition of m-xylene to fumarate to form (3-methylbenzyl)succinate. In the presence of succinyl coenzyme A (CoA) and nitrate, (3 methylbenzyl)succinate was oxidized to E-(3-methylphenyl)itaconate (or a closely related isomer) and 3-methylbenzoate. Kinetic studies conducted with permeabilized cells and whole-cell suspensions of m-xylene-grown Azoarcus sp. strain T demonstrated that the specific rate of in vitro (3 methylbenzyl)succinate formation accounts for at least 15% of the specific rate of in vivo m-xylene consumption. Based on these findings, we propose that Azoarcus sp. strain T anaerobically oxidizes m-xylene to 3-methylbenzoate (or its CoA thioester) via (3-methylbenzyl)succinate and E-(3-methylphenyl)itaconate (or its CoA thioester) in a series of reactions that are analogous to those recently proposed for anaerobic toluene oxidation to benzoyl-CoA. A deuterium kinetic isotope effect was observed in the (3-methylbenzyl)succinate synthase reaction (and the benzylsuccinate synthase reaction), suggesting that a rate-determining step in this novel fumarate addition reaction involves breaking a C-H bond. PMID- 10515930 TI - A novel role for Escherichia coli endonuclease VIII in prevention of spontaneous G-->T transversions. AB - In the bacterium Escherichia coli, oxidized pyrimidines are removed by two DNA glycosylases, endonuclease III and endonuclease VIII (endo VIII), encoded by the nth and nei genes, respectively. Double mutants lacking both of these activities exhibit a high spontaneous mutation frequency, and here we show that all of the mutations observed in the double mutants were G:C-->A:T transitions; no thymine mutations were found. These findings are in agreement with the preponderance of C ->T transitions in the oxidative and spontaneous mutational databases. The major oxidized purine lesion in DNA, 7,8-dihydro-8-oxoguanine (8-oxoG), is processed by two DNA glycosylases, formamidopyrimidine DNA glycosylase (Fpg), which removes 8 oxoG opposite C, and MutY DNA glycosylase, which removes misincorporated A opposite 8-oxoG. The high spontaneous mutation frequency previously observed in fpg mutY double mutants was significantly enhanced by the addition of the nei mutation, suggesting an overlap in the substrate specificities between endo VIII and Fpg/MutY. When the mutational specificity was examined, all of the mutations observed were G:C-->T:A transversions, indicating that in the absence of Fpg and MutY, endo VIII serves as a backup activity to remove 8-oxoG. This was confirmed by showing that, indeed, endo VIII can recognize 8-oxoG in vitro. PMID- 10515932 TI - Mechanism of repression of the aroP P2 promoter by the TyrR protein of Escherichia coli. AB - Previously, we have shown that expression of the Escherichia coli aroP P2 promoter is partially repressed by the TyrR protein alone and strongly repressed by the TyrR protein in the presence of the coeffector tyrosine or phenylalanine (P. Wang, J. Yang, and A. J. Pittard, J. Bacteriol. 179:4206-4212, 1997). Here we present in vitro results showing that the TyrR protein and RNA polymerase can bind simultaneously to the aroP P2 promoter. In the presence of tyrosine, the TyrR protein inhibits open complex formation at the P2 promoter, whereas in the absence of any coeffector or in the presence of phenylalanine, the TyrR protein inhibits a step(s) following the formation of open complexes. We also present mutational evidence which implicates the N-terminal domain of the TyrR protein in the repression of P2 expression. The TyrR binding site of aroP, which includes one weak and one strong TyrR box, is located 5 bp downstream of the transcription start site of P2. Results from a mutational analysis show that the strong box (which is located more closely to the P2 promoter), but not the weak box, plays a critical role in P2 repression. PMID- 10515933 TI - MinDE-dependent pole-to-pole oscillation of division inhibitor MinC in Escherichia coli. AB - By inhibiting FtsZ ring formation near the cell ends, the MinC protein plays a critical role in proper positioning of the division apparatus in Escherichia coli. MinC activity requires that of MinD, and the MinE peptide provides topological specificity by suppressing MinC-MinD-mediated division inhibition specifically at the middle of the cell. We recently presented evidence that MinE not only accumulates in an FtsZ-independent ring structure at the cell's middle but also imposes a unique dynamic localization pattern upon MinD in which the latter accumulates alternately in either one of the cell halves in what appears to be a rapidly oscillating membrane association-dissociation cycle. Here we show that functional green fluorescent protein-MinC displays a very similar oscillatory behavior which is dependent on both MinD and MinE and independent of FtsZ. The results support a model in which MinD recruits MinC to its site of action and in which FtsZ ring assembly at each of the cell ends is blocked in an intermittent and alternate fashion. PMID- 10515935 TI - Identification and characterization of major lipid particle proteins of the yeast Saccharomyces cerevisiae. AB - Lipid particles of the yeast Saccharomyces cerevisiae were isolated at high purity, and their proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Major lipid particle proteins were identified by mass spectrometric analysis, and the corresponding open reading frames (ORFs) were deduced. In silicio analysis revealed that all lipid particle proteins contain several hydrophobic domains but none or only few (hypothetical) transmembrane spanning regions. All lipid particle proteins identified by function so far, such as Erg1p, Erg6p, and Erg7p (ergosterol biosynthesis) and Faa1p, Faa4p, and Fat1p (fatty acid metabolism), are involved in lipid metabolism. Based on sequence homology, another group of three lipid particle proteins may be involved in lipid degradation. To examine whether lipid particle proteins of unknown function are also involved in lipid synthesis, mutants with deletions of the respective ORFs were constructed and subjected to systematic lipid analysis. Deletion of YDL193w resulted in a lethal phenotype which could not be suppressed by supplementation with ergosterol or fatty acids. Other deletion mutants were viable under standard conditions. Strains with YBR177c, YMR313c, and YKL140w deleted exhibited phospholipid and/or neutral lipid patterns that were different from the wild-type strain and thus may be further candidate ORFs involved in yeast lipid metabolism. PMID- 10515934 TI - Functional genomics: expression analysis of Escherichia coli growing on minimal and rich media. AB - DNA arrays of the entire set of Escherichia coli genes were used to measure the genomic expression patterns of cells growing in late logarithmic phase on minimal glucose medium and on Luria broth containing glucose. Ratios of the transcript levels for all 4,290 E. coli protein-encoding genes (cds) were obtained, and analysis of the expression ratio data indicated that the physiological state of the cells under the two growth conditions could be ascertained. The cells in the rich medium grew faster, and expression of the majority of the translation apparatus genes was significantly elevated under this growth condition, consistent with known patterns of growth rate-dependent regulation and increased rate of protein synthesis in rapidly growing cells. The cells grown on minimal medium showed significantly elevated expression of many genes involved in biosynthesis of building blocks, most notably the amino acid biosynthetic pathways. Nearly half of the known RpoS-dependent genes were expressed at significantly higher levels in minimal medium than in rich medium, and rpoS expression was similarly elevated. The role of RpoS regulation in these logarithmic phase cells was suggested by the functions of the RpoS dependent genes that were induced. The hallmark features of E. coli cells growing on glucose minimal medium appeared to be the formation and excretion of acetate, metabolism of the acetate, and protection of the cells from acid stress. A hypothesis invoking RpoS and UspA (universal stress protein, also significantly elevated in minimal glucose medium) as playing a role in coordinating these various aspects and consequences of glucose and acetate metabolism was generated. This experiment demonstrates that genomic expression assays can be applied in a meaningful way to the study of whole-bacterial-cell physiology for the generation of hypotheses and as a guide for more detailed studies of particular genes of interest. PMID- 10515936 TI - Pathogenic Yersinia species carry a novel, cold-inducible major cold shock protein tandem gene duplication producing both bicistronic and monocistronic mRNA. AB - Inverse PCR was used to amplify major cold shock protein (MCSP) gene families from a diverse range of bacteria, including the psychrotolerant Yersinia enterocolitica, which was found to have two almost identical MCSP coding regions (cspA1 and cspA2) located approximately 300 bp apart. This tandem gene duplication was also found in Y. pestis, Y. pseudotuberculosis, and Y. ruckeri but not in other bacteria. Analysis of the transcriptional regulation of this MCSP gene in Y. enterocolitica, performed by using both reverse transcriptase-PCR and Northern blot assays, showed there to be two cold-inducible mRNA templates arising from this locus: a monocistronic template of approximately 450 bp (cspA1) and a bicistronic template of approximately 900 bp (cspA1/A2). The former may be due to a secondary structure between cspA1 and cspA2 causing either 3' degradation protection of cspA1 or, more probably, partial termination after cspA1. Primer extension experiments identified a putative transcriptional start site (+1) which is flanked by a cold-box motif and promoter elements (-10 and 35) similar to those found in Escherichia coli cold-inducible MCSP genes. At 30 degrees C, the level of both mRNA molecules was negligible; however, upon a temperature downshift to 10 degrees C, transcription of the bicistronic mRNA was both substantial (300-fold increase) and immediate, with transcription of the monocistronic mRNA being approximately 10-fold less (30-fold increase) and significantly slower. The ratio of bicistronic to monocistronic mRNA changed with time after cold shock and was higher when cells were shocked to a lower temperature. High-resolution, two-dimensional protein gel electrophoresis showed that synthesis of the corresponding proteins, both CspA1 and CspA2, was apparent after only 10 min of cold shock from 30 degrees C to 10 degrees C. The data demonstrate an extraordinary capacity of the psychrotolerant Y. enterocolitica to produce major cold shock proteins upon cold shock. PMID- 10515937 TI - Biochemical and genetic analyses of the role of yeast casein kinase 2 in salt tolerance. AB - Saccharomyces cerevisiae cells lacking the regulatory subunit of casein kinase 2 (CK-2), encoded by the gene CKB1, display a phenotype of hypersensitivity to Na(+) and Li(+) cations. The sensitivity of a strain lacking ckb1 is higher than that of a calcineurin mutant and similar to that of a strain lacking HAL3, the regulatory subunit of the Ppz1 protein phosphatase. Genetic analysis indicated that Ckb1 participates in regulatory pathways different from that of Ppz1 or calcineurin. Deletion of CKB1 increased the salt sensitivity of a strain lacking Ena1 ATPase, the major determinant for sodium efflux, suggesting that the function of the kinase is not mediated by Ena1. Consistently, ckb1 mutants did not show an altered cation efflux. The function of Ckb1 was independent of the TRK system, which is responsible for discrimination of potassium and sodium entry, and in the absence of the kinase regulatory subunit, the influx of sodium was essentially normal. Therefore, the salt sensitivity of a ckb1 mutant cannot be attributed to defects in the fluxes of sodium. In fact, in these cells, both the intracellular content and the cytoplasm/vacuole ratio for sodium were similar to those features of wild-type cells. The possible causes for the salt sensitivity phenotype of casein kinase mutants are discussed in the light of these findings. PMID- 10515938 TI - Identification and characterization of the single-stranded DNA-binding protein of bacteriophage P1. AB - The genome of bacteriophage P1 harbors a gene coding for a 162-amino-acid protein which shows 66% amino acid sequence identity to the Escherichia coli single stranded DNA-binding protein (SSB). The expression of the P1 gene is tightly regulated by P1 immunity proteins. It is completely repressed during lysogenic growth and only weakly expressed during lytic growth, as assayed by an ssb P1/lacZ fusion construct. When cloned on an intermediate-copy-number plasmid, the P1 gene is able to suppress the temperature-sensitive defect of an E. coli ssb mutant, indicating that the two proteins are functionally interchangeable. Many bacteriophages and conjugative plasmids do not rely on the SSB protein provided by their host organism but code for their own SSB proteins. However, the close relationship between SSB-P1 and the SSB protein of the P1 host, E. coli, raises questions about the functional significance of the phage protein. PMID- 10515939 TI - A developmentally regulated gene cluster involved in conidial pigment biosynthesis in Aspergillus fumigatus. AB - Aspergillus fumigatus, a filamentous fungus producing bluish-green conidia, is an important opportunistic pathogen that primarily affects immunocompromised patients. Conidial pigmentation of A. fumigatus significantly influences its virulence in a murine model. In the present study, six genes, forming a gene cluster spanning 19 kb, were identified as involved in conidial pigment biosynthesis in A. fumigatus. Northern blot analyses showed the six genes to be developmentally regulated and expressed during conidiation. The gene products of alb1 (for "albino 1"), arp1 (for "aspergillus reddish-pink 1"), and arp2 have high similarity to polyketide synthases, scytalone dehydratases, and hydroxynaphthalene reductases, respectively, found in the dihydroxynaphthalene (DHN)-melanin pathway of brown and black fungi. The abr1 gene (for "aspergillus brown 1") encodes a putative protein possessing two signatures of multicopper oxidases. The abr2 gene product has homology to the laccase encoded by the yA gene of Aspergillus nidulans. The function of ayg1 (for "aspergillus yellowish green 1") remains unknown. Involvement of the six genes in conidial pigmentation was confirmed by the altered conidial color phenotypes that resulted from disruption of each gene in A. fumigatus. The presence of a DHN-melanin pathway in A. fumigatus was supported by the accumulation of scytalone and flaviolin in the arp1 deletant, whereas only flaviolin was accumulated in the arp2 deletants. Scytalone and flaviolin are well-known signature metabolites of the DHN-melanin pathway. Based on DNA sequence similarity, gene disruption results, and biochemical analyses, we conclude that the 19-kb DNA fragment contains a six-gene cluster which is required for conidial pigment biosynthesis in A. fumigatus. However, the presence of abr1, abr2, and ayg1 in addition to alb1, arp1, and arp2 suggests that conidial pigment biosynthesis in A. fumigatus is more complex than the known DHN-melanin pathway. PMID- 10515941 TI - Horizontal transfer of genetic material among Saccharomyces yeasts. AB - The genus Saccharomyces consists of several species divided into the sensu stricto and the sensu lato groups. The genomes of these species differ in the number and organization of nuclear chromosomes and in the size and organization of mitochondrial DNA (mtDNA). In the present experiments we examined whether these yeasts can exchange DNA and thereby create novel combinations of genetic material. Several putative haploid, heterothallic yeast strains were isolated from different Saccharomyces species. All of these strains secreted an a- or alpha-like pheromone recognized by S. cerevisiae tester strains. When interspecific crosses were performed by mass mating between these strains, hybrid zygotes were often detected. In general, the less related the two parental species were, the fewer hybrids they gave. For some crosses, viable hybrids could be obtained by selection on minimal medium and their nuclear chromosomes and mtDNA were examined. Often the frequency of viable hybrids was very low. Sometimes putative hybrids could not be propagated at all. In the case of sensu stricto yeasts, stable viable hybrids were obtained. These contained both parental sets of chromosomes but mtDNA from only one parent. In the case of sensu lato hybrids, during genetic stabilization one set of the parental chromosomes was partially or completely lost and the stable mtDNA originated from the same parent as the majority of the nuclear chromosomes. Apparently, the interspecific hybrid genome was genetically more or less stable when the genetic material originated from phylogenetically relatively closely related parents; both sets of nuclear genetic material could be transmitted and preserved in the progeny. In the case of more distantly related parents, only one parental set, and perhaps some fragments of the other one, could be found in genetically stabilized hybrid lines. The results obtained indicate that Saccharomyces yeasts have a potential to exchange genetic material. If Saccharomyces isolates could mate freely in nature, horizontal transfer of genetic material could have occurred during the evolution of modern yeast species. PMID- 10515940 TI - Substitution, insertion, deletion, suppression, and altered substrate specificity in functional protocatechuate 3,4-dioxygenases. AB - Protocatechuate 3,4-dioxygenase is a member of a family of bacterial enzymes that cleave the aromatic rings of their substrates between two adjacent hydroxyl groups, a key reaction in microbial metabolism of varied environmental chemicals. In an appropriate genetic background, it is possible to select for Acinetobacter strains containing spontaneous mutations blocking expression of pcaH or -G, genes encoding the alpha and beta subunits of protocatechuate 3, 4-dioxygenase. The crystal structure of the Acinetobacter oxygenase has been determined, and this knowledge affords us the opportunity to understand how mutations alter function in the enzyme. An earlier investigation had shown that a large fraction of spontaneous mutations inactivating Acinetobacter protocatechuate oxygenase are either insertions or large deletions. Therefore, the prior procedure of mutant selection was modified to isolate Acinetobacter strains in which mutations within pcaH or -G cause a heat-sensitive phenotype. These mutations affected residues distributed throughout the linear amino acid sequences of PcaH and PcaG and impaired the dioxygenase to various degrees. Four of 16 mutants had insertions or deletions in the enzyme ranging in size from 1 to 10 amino acid residues, highlighting areas of the protein where large structural changes can be tolerated. To further understand how protein structure influences function, we isolated strains in which the phenotypes of three different deletion mutations in pcaH or -G were suppressed either by a spontaneous mutation or by a PCR-generated random mutation introduced into the Acinetobacter chromosome by natural transformation. The latter procedure was also used to identify a single amino acid substitution in PcaG that conferred activity towards catechol sufficient for growth with benzoate in a strain in which catechol 1,2-dioxygenase was inactivated. PMID- 10515942 TI - Divergent regulation of the evolutionarily closely related promoters of the Saccharomyces cerevisiae STA2 and MUC1 genes. AB - The 5' upstream regions of the Saccharomyces cerevisiae glucoamylase-encoding genes STA1 to -3 and of the MUC1 (or FLO11) gene, which is critical for pseudohyphal development, invasive growth, and flocculation, are almost identical, and the genes are coregulated to a large extent. Besides representing the largest yeast promoters identified to date, these regions are of particular interest from both a functional and an evolutionary point of view. Transcription of the genes indeed seems to be dependent on numerous transcription factors which integrate the information of a complex network of signaling pathways, while the very limited sequence differences between them should allow the study of promoter evolution on a molecular level. To investigate the transcriptional regulation, we compared the transcription levels conferred by the STA2 and MUC1 promoters under various growth conditions. Our data show that transcription of both genes responded similarly to most environmental signals but also indicated significant divergence in some aspects. We identified distinct areas within the promoters that show specific responses to the activating effect of Flo8p, Msn1p (or Mss10p, Fup1p, or Phd2p), and Mss11p as well as to carbon catabolite repression. We also identified the STA10 repressive effect as the absence of Flo8p, a transcriptional activator of flocculation genes in S. cerevisiae. PMID- 10515944 TI - Sulfide-quinone reductase from Rhodobacter capsulatus: requirement for growth, periplasmic localization, and extension of gene sequence analysis. AB - The entire sequence of the 3.5-kb fragment of genomic DNA from Rhodobacter capsulatus which contains the sqr gene and a second complete and two further partial open reading frames has been determined. A correction of the previously published sqr gene sequence (M. Schutz, Y. Shahak, E. Padan, and G. Hauska, J. Biol. Chem. 272:9890-9894, 1997) which in the deduced primary structure of the sulfide-quinone reductase changes four positive into four negative charges and the number of amino acids from 425 to 427 was necessary. The correction has no further bearing on the former sequence analysis. Deletion and interruption strains document that sulfide-quinone reductase is essential for photoautotrophic growth on sulfide. The sulfide-oxidizing enzyme is involved in energy conversion, not in detoxification. Studies with an alkaline phosphatase fusion protein reveal a periplasmic localization of the enzyme. Exonuclease treatment of the fusion construct demonstrated that the C-terminal 38 amino acids of sulfide-quinone reductase were required for translocation. An N-terminal signal peptide for translocation was not found in the primary structure of the enzyme. The possibility that the neighboring open reading frame, which contains a double arginine motif, may be involved in translocation has been excluded by gene deletion (rather, the product of this gene functions in an ATP-binding cassette transporter system, together with the product of one of the other open reading frames). The results lead to the conclusion that the sulfide-quinone reductase of R. capsulatus functions at the periplasmic surface of the cytoplasmic membrane and that this flavoprotein is translocated by a hitherto-unknown mechanism. PMID- 10515943 TI - Sequence and organization of pXO1, the large Bacillus anthracis plasmid harboring the anthrax toxin genes. AB - The Bacillus anthracis Sterne plasmid pXO1 was sequenced by random, "shotgun" cloning. A circular sequence of 181,654 bp was generated. One hundred forty-three open reading frames (ORFs) were predicted using GeneMark and GeneMark.hmm, comprising only 61% (110,817 bp) of the pXO1 DNA sequence. The overall guanine plus-cytosine content of the plasmid is 32.5%. The most recognizable feature of the plasmid is a "pathogenicity island," defined by a 44.8-kb region that is bordered by inverted IS1627 elements at each end. This region contains the three toxin genes (cya, lef, and pagA), regulatory elements controlling the toxin genes, three germination response genes, and 19 additional ORFs. Nearly 70% of the ORFs on pXO1 do not have significant similarity to sequences available in open databases. Absent from the pXO1 sequence are homologs to genes that are typically required to drive theta replication and to maintain stability of large plasmids in Bacillus spp. Among the ORFs with a high degree of similarity to known sequences are a collection of putative transposases, resolvases, and integrases, suggesting an evolution involving lateral movement of DNA among species. Among the remaining ORFs, there are three sequences that may encode enzymes responsible for the synthesis of a polysaccharide capsule usually associated with serotype-specific virulent streptococci. PMID- 10515945 TI - Purification of P(II) and P(II)-UMP and in vitro studies of regulation of glutamine synthetase in Rhodospirillum rubrum. AB - The P(II) protein from Rhodospirillum rubrum was fused with a histidine tag, overexpressed in Escherichia coli, and purified by Ni(2+)-chelating chromatography. The uridylylated form of the P(II) protein could be generated in E. coli. The effects on the regulation of glutamine synthetase by P(II), P(II) UMP, glutamine, and alpha-ketoglutarate were studied in extracts from R. rubrum grown under different conditions. P(II) and glutamine were shown to stimulate the ATP-dependent inactivation (adenylylation) of glutamine synthetase, which could be totally inhibited by alpha-ketoglutarate. Deadenylylation (activation) of glutamine synthetase required phosphate, but none of the effectors studied had any major effect, which is different from their role in the E. coli system. In addition, deadenylylation was found to be much slower than adenylylation under the conditions investigated. PMID- 10515946 TI - Expression of P(II) and glutamine synthetase is regulated by P(II), the ntrBC products, and processing of the glnBA mRNA in Rhodospirillum rubrum. AB - We have studied the transcription of the glnB and glnA genes in Rhodospirillum rubrum with firefly luciferase as a reporter enzyme. Under NH(4)(+) and N(2) conditions, glnBA was cotranscribed from a weak and a strong promoter. In nitrogen-fixing cultures, activity of the latter was highly enhanced by NtrC, but transcription from both promoters occurred under both conditions. There is no promoter controlling transcription of glnA alone, supporting our proposal that the glnA mRNA is produced by processing. PMID- 10515947 TI - Genetic analysis of nif regulatory genes by utilizing the yeast two-hybrid system detected formation of a NifL-NifA complex that is implicated in regulated expression of nif genes. AB - In diazotrophic organisms, nitrogenase synthesis and activity are tightly regulated. Two genes, nifL and nifA, are implicated as playing a major role in this regulation. NifA is a transcriptional activator, and its activity is inhibited by NifL in response to availability of excess fixed nitrogen and high O(2) tension. It was postulated that NifL binds to NifA to inhibit NifA-mediated transcriptional activation of nif genes. Mutational analysis combined with transcriptional activation studies clearly is in agreement with the proposal that NifL interacts with NifA. However, several attempts to identify NifA-NifL interactions by using methods such as coimmunoprecipitations and chemical cross linking experiments failed to detect direct interactions between these proteins. Here we have taken a genetic approach, the use of a yeast two-hybrid protein protein interaction assay system, to investigate NifL interaction with NifA. A DNA fragment corresponding to the kinase-like domain of nifL was PCR amplified and was used to generate translation fusions with the DNA binding domain and the DNA activation domain of the yeast transcriptional activator GAL4 in yeast two hybrid vectors. Similarly, a DNA fragment corresponding to the catalytic domain of nifA was PCR amplified and used to generate translation fusions with the DNA binding domain and the DNA-activation domain of GAL4 in yeast two-hybrid vectors. After introducing appropriate plasmid combinations in yeast cells, the existance of direct interaction between NifA and NifL was analyzed with the MATCHMAKER yeast two-hybrid system by testing for the expression of lacZ and his3 genes. These analyses showed that the kinase-like domain of NifL directly interacts with the catalytic domain of NifA. PMID- 10515948 TI - Localization of periplasmic redox proteins of Alcaligenes faecalis by a modified general method for fractionating gram-negative bacteria. AB - A lysozyme-osmotic shock method is described for fractionation of Alcaligenes faecalis which uses glucose to adjust osmotic strength and multiple osmotic shocks. During phenylethylamine-dependent growth, aromatic amine dehydrogenase, azurin, and a single cytochrome c were localized in the periplasm. Their induction patterns are different from those for the related quinoprotein methylamine dehydrogenase and its associated redox proteins. PMID- 10515949 TI - Porins in the cell wall of Mycobacterium tuberculosis. AB - Lipid bilayer experiments indicated that the cell wall of Mycobacterium tuberculosis contains at least two different porins: (i) a cation-selective, heat sensitive 0.7-nS channel which has a short-lived open state and is probably composed of 15-kDa subunits and (ii) a 3-nS, >60-kDa channel with a long-lived open state, resembling porins from fast-growing mycobacteria. PMID- 10515950 TI - Escherichia coli Lrp (leucine-responsive regulatory protein) does not directly regulate expression of the leu operon promoter. AB - Studies by R. Lin et al. (J. Bacteriol. 174:1948-1955, 1992) suggested that the Escherichia coli leu operon might be a member of the Lrp regulon. Their results were obtained with a leucine auxotroph; in leucine prototrophs grown in a medium lacking leucine, there was little difference in leu operon expression between lrp(+) and lrp strains. Furthermore, when leuP-lacZ transcriptional fusions that lacked the leu attenuator were used, expression from the leu promoter varied less than twofold between lrp(+) and lrp strains, irrespective of whether or not excess leucine was added to the medium. The simplest explanation of the observations of Lin et al. is that the known elevated leucine transport capacity of lrp strains (S. A. Haney et al., J. Bacteriol. 174:108-115, 1992) leads to very high intracellular levels of leucine for strains grown with leucine, resulting in the superattenuation of leu operon expression. PMID- 10515951 TI - Gene disruption studies of penicillin-binding proteins 1a, 1b, and 2a in Streptococcus pneumoniae. AB - The effects of inactivation of the genes encoding penicillin-binding protein 1a (PBP1a), PBP1b, and PBP2a in Streptococcus pneumoniae were examined. Insertional mutants did not exhibit detectable changes in growth rate or morphology, although a pbp1a pbp1b double-disruption mutant grew more slowly than its parent did. Attempts to generate a pbp1a pbp2a double-disruption mutant failed. The pbp2a mutants, but not the other mutants, were more sensitive to moenomycin, a transglycosylase inhibitor. These observations suggest that individually the pbp1a, pbp1b, and pbp2a genes are dispensable but that either pbp1a or pbp2a is required for growth in vitro. These results also suggest that PBP2a is a functional transglycosylase in S. pneumoniae. PMID- 10515952 TI - A novel gene required for rhamnose-glucose polysaccharide synthesis in Streptococcus mutans. AB - Gene rgpG is required for biosynthesis of rhamnose-glucose polysaccharide (RGP) in Streptococcus mutans. Its deduced amino acid sequence had similarity to WecA, which initiates syntheses of enterobacterial common antigen and some O antigens in Escherichia coli. Gene rgpG complemented a wecA mutation of E. coli, suggesting that rgpG may function similarly in RGP synthesis. PMID- 10515953 TI - Occurrence of free D-amino acids and aspartate racemases in hyperthermophilic archaea. AB - The occurrence of free D-amino acids and aspartate racemases in several hyperthermophilic archaea was investigated. Aspartic acid in all the hyperthermophilic archaea was highly racemized. The ratio of D-aspartic acid to total aspartic acid was in the range of 43.0 to 49.1%. The crude extracts of the hyperthermophiles exhibited aspartate racemase activity at 70 degrees C, and aspartate racemase homologous genes in them were identified by PCR. D-Enantiomers of other amino acids (alanine, leucine, phenylalanine, and lysine) in Thermococcus strains were also detected. Some of them might be by-products of aspartate racemase. It is proven that D-amino acids are produced in some hyperthermophilic archaea, although their function is unknown. PMID- 10515955 TI - Efficiency of transcription from promoter sequence variants in Lactobacillus is both strain and context dependent. AB - The introduction of consensus -35 (TTGACA) and -10 (TATAAT) hexamers and a TG motif into the Lactobacillus acidophilus ATCC 4356 wild-type slpA promoter resulted in significant improvements (4.3-, 4.1-, and 10.7-fold, respectively) in transcriptional activity in Lactobacillus fermentum BR11. In contrast, the same changes resulted in decreased transcription in Lactobacillus rhamnosus GG. The TG motif was shown to be important in the context of weak -35 and -10 hexamers (L. fermentum BR11) or a consensus -10 hexamer (L. rhamnosus GG). Thus, both strain- and context-dependent effects are critical factors influencing transcription in Lactobacillus. PMID- 10515954 TI - The modality of enterobacterial common antigen polysaccharide chain lengths is regulated by o349 of the wec gene cluster of Escherichia coli K-12. AB - The assembly of the phosphoglyceride-linked form of enterobacterial common antigen (ECA(PG)) occurs by a mechanism that involves modulation of polysaccharide chain length. However, the genetic determinant of this modulation has not been identified. Site-directed mutagenesis of o349 of the Escherichia coli K-12 wec gene cluster revealed that this locus encodes a Wzz protein that specifically modulates the chain length of ECA(PG) polysaccharides, and we have designated this locus wzz(ECA). The Wzz(ECA)-mediated modulation of ECA(PG) polysaccharide chains is the first demonstrated example of Wzz regulation involving a polysaccharide that is not linked to the core-lipid A structure of lipopolysaccharide. PMID- 10515956 TI - Fast BK-type channel mediates the Ca(2+)-activated K(+) current in crayfish muscle. AB - The role of the Ca(2+)-activated K(+) current (I(K(Ca))) in crayfish opener muscle fibers is functionally important because it regulates the graded electrical activity that is characteristic of these fibers. Using the cell attached and inside-out configurations of the patch-clamp technique, we found three different classes of channels with properties that matched those expected of the three different ionic channels mediating the depolarization-activated macroscopic currents previously described (Ca(2+), K(+), and Ca(2+)-dependent K(+) currents). We investigated the properties of the ionic channels mediating the extremely fast activating and persistent I(K(Ca)). These voltage- and Ca(2+) activated channels had a mean single-channel conductance of approximately 70 pS and showed a very fast activation. Both the single-channel open probability and the speed of activation increased with depolarization. Both parameters also increased in inside-out patches, i.e., in high Ca(2+) concentration. Intracellular loading with the Ca(2+) chelator bis(2-aminophenoxy) ethane-N, N,N',N'-tetraacetic acid gradually reduced and eventually prevented channel openings. The channels opened at very brief delays after the pulse depolarization onset (<5 ms), and the time-dependent open probability was constant during sustained depolarization (< or =560 ms), matching both the extremely fast activation kinetics and the persistent nature of the macroscopic I(K(Ca)). However, the intrinsic properties of these single channels do not account for the partial apparent inactivation of the macroscopic I(K(Ca)), which probably reflects temporal Ca(2+) variations in the whole muscle fiber. We conclude that the channels mediating I(K(Ca)) in crayfish muscle are voltage- and Ca(2+)-gated BK channels with relatively small conductance. The intrinsic properties of these channels allow them to act as precise Ca(2+) sensors that supply the exact feedback current needed to control the graded electrical activity and therefore the contraction of opener muscle fibers. PMID- 10515957 TI - Participation of a chloride conductance in the subthreshold behavior of the rat sympathetic neuron. AB - The presence of a novel voltage-dependent chloride current, active in the subthreshold range of membrane potential, was detected in the mature and intact rat sympathetic neuron in vitro by using the two-microelectrode voltage-clamp technique. Hyperpolarizing voltage steps applied to a neuron held at -40/-50 mV elicited inward currents, whose initial magnitude displayed a linear instantaneous current-voltage (I-V) relationship; afterward, the currents decayed exponentially with a single voltage-dependent time constant (63.5 s at -40 mV; 10.8 s at -130 mV). The cell input conductance decreased during the command step with the same time course as the current. On returning to the holding potential, the ensuing outward currents were accompanied by a slow increase in input conductance toward the initial values; the inward charge movement during the transient ON response (a mean of 76 nC in 8 neurons stepped from -50 to -90 mV) was completely balanced by outward charge displacement during the OFF response. The chloride movements accompanying voltage modifications were studied by estimating the chloride equilibrium potential (E(Cl)) at different holding potentials from the reversal of GABA evoked currents. [Cl(-)](i) was strongly affected by membrane potential, and at steady state it was systematically higher than expected from passive ion distribution. The transient current was blocked by substitution of isethionate for chloride and by Cl(-) channel blockers (9AC and DIDS). It proved insensitive to K(+) channel blockers, external Cd(2+), intracellular Ca(2+) chelators [bis-(o-aminophenoxy)-N,N,N',N'-tetraacetic acid (BAPTA)] and reduction of [Na(+)](e). It is concluded that membrane potential shifts elicit a chloride current that reflects readjustment of [Cl(-)](i). The cell input conductance was measured over the -40/-120-mV voltage range, in control medium, and under conditions in which either the chloride or the potassium current was blocked. A mix of chloride, potassium, and leakage conductances was detected at all potentials. The leakage component was voltage independent and constant at approximately 14 nS. Conversely, gCl decreased with hyperpolarization (80 nS at -40 mV, undetectable below -110 mV), whereas gK displayed a maximum at -80 mV (55.3 nS). Thus the ratio gCl/gK continuously varied with membrane polarization (2.72 at -50 mV; 0.33 at -110 mV). These data were forced in a model of the three current components here described, which accurately simulates the behavior observed in the "resting" neuron during membrane migrations in the subthreshold potential range, thereby confirming that active K and Cl conductances contribute to the genesis of membrane potential and possibly to the control of neuronal excitability. PMID- 10515958 TI - Contrast rectification and distributed encoding By ON-OFF amacrine cells in the retina. AB - The encoding of luminance contrast by ON-OFF amacrine cells was investigated by intracellular recording in the retina of the tiger salamander (Ambystoma tigrinum). Contrast flashes of positive and negative polarity were applied at the center of the receptive field while the entire retina was light adapted to a background field of 20 cd/m(2). Many amacrine cells showed remarkably high contrast gain: Up to 20-35% of the maximum response was evoked by a contrast step of only 1%. In the larger signal domain, C50, the contrast required to evoke a response 50% of the maximum, was often remarkably low: 24 of 25 cells had a C50 value of < or =10% for at least one contrast polarity. Across cells and contrast polarity, the dynamic ranges varied from extremely narrow to broad, thereby blanketing the range of reflectances associated with objects in natural environments. Although some cells resembled "contrast rectifiers," by showing similar responses to contrasts of opposite polarity, many did not. Thus for contrast gain and C50, individual cells could show a strong preference for either negative or positive contrast. In the time domain, the preference was strong and unidirectional: for equal contrast steps, the latency of the response to negative contrast was 20-45 ms shorter than that for positive contrast. The present results, when compared with those for bipolar cells, suggest that, on average, amacrine cells add some amplification, particularly for negative contrast, to the high contrast gain already established by bipolar cells. In the time domain, our data reveal a striking transformation from bipolar to amacrine cells in favor of negative contrast. These and further observations have implications for the input and output of amacrine cell circuits. The present finding of substantial differences between cells reveals a potential substrate for distributed encoding of luminance contrast within the ON-OFF amacrine cell population. PMID- 10515959 TI - Vasopressin and amastatin induce V(1)-receptor-mediated suppression of excitatory transmission in the rat parabrachial nucleus. AB - We examined actions of arginine vasopressin (AVP) and amastatin (an inhibitor of the aminopeptidase that cleaves AVP) on synaptic currents in slices of rat parabrachial nucleus using the nystatin-perforated patch recording technique. AVP reversibly decreased the amplitude of the evoked, glutamate-mediated, excitatory postsynaptic current (EPSC) with an increase in paired-pulse ratio. No apparent changes in postsynaptic membrane properties were revealed by ramp protocols, and the inward current induced by a brief application of alpha-amino-3-hydroxy-5 methylisoxazole-4-propionic acid was unchanged after AVP. The reduction induced by 1 microM AVP could be blocked by a V(1) AVP receptor antagonist, [d(CH(2))(5)(1)-O-Me-Tyr(2)-Arg(8)]-vasopressin (Manning compound, 10 microM). Bath application of an aminopeptidase inhibitor, amastatin (10 microM), reduced the evoked EPSC, and AVP induced further synaptic depression in the presence of amastatin. Amastatin's effects also could be antagonized by the Manning compound. Corticotropin-releasing hormone slightly increased the EPSC at 1 microM, and coapplication with AVP attenuated the AVP response. Pretreatment of slices with 1 microg/ml cholera toxin or 0.5 microg/ml pertussis toxin for 20 h did not significantly affect AVP's synaptic action. The results suggest that AVP has suppressant effects on glutamatergic transmission by acting at V(1) AVP receptors, possibly through a presynaptic mechanism involving a pertussis-toxin- and cholera-toxin-resistant pathway. PMID- 10515960 TI - Regulation of the rebound depolarization and spontaneous firing patterns of deep nuclear neurons in slices of rat cerebellum. AB - Current-clamp recordings were made from the deep cerebellar nuclei (DCN) of 12- to 15-day-old rats to understand the factors that mediate intrinsic spontaneous firing patterns. All of the cells recorded were spontaneously active with spiking patterns ranging continuously from regular spiking to spontaneous bursting with the former predominating. A robust rebound depolarization (RD) leading to a Na(+) spike burst was elicited after the offset of hyperpolarizing current injection. The voltage and time dependence of the RD was consistent with mediation by low threshold voltage-gated Ca(2+) channels. In addition, induction of a RD also may be affected by activation of a hyperpolarization-activated cation current, I(h). A RD could be evoked efficiently after brief high-frequency bursts of inhibitory postsynaptic potentials (IPSPs) induced by stimulation of Purkinje cell axons. IPSP-driven RDs were typically much larger and longer than those elicited by direct hyperpolarizing pulses of approximately matched amplitude and duration. Intracellular perfusion of the Ca(2+) buffer bis-(o-aminophenoxy)-N,N,N',N' tetraacetic acid (BAPTA) dramatically enhanced the RD and its associated spiking, sometimes leading to a plateau potential that lasted several hundred milliseconds. The effects of BAPTA could be mimicked partly by application of apamin, a blocker of small conductance Ca(2+)-gated K(+) channels, but not by paxilline, which blocks large conductance Ca(2+)-gated K(+) channels. Application of both BAPTA and apamin, but not paxilline, caused cells that were regularly spiking to burst spontaneously. Taken together, our data suggest that there is a strong relationship between the ability of DCN cells to elicit a RD and their tendency burst spontaneously. The RD can be triggered by the opening of T-type Ca(2+) channels with an additional contribution of hyperpolarization-activated current I(h). RD duration is regulated by small-conductance Ca(2+)-gated K(+) channels. The RD also is modulated tonically by inhibitory inputs. All of these factors are in turn subject to alteration by extrinsic modulatory neurotransmitters and are, at least in part, responsible for determining the firing modes of DCN neurons. PMID- 10515961 TI - Shift in smooth pursuit initiation and MT and MST neuronal activity under different stimulus conditions. AB - The activity of neurons in extrastriate middle temporal (MT) and medial superior temporal (MST) areas were studied during the initiation of pursuit eye movements in macaque monkeys. The intersecting motion of two stimuli was used to test hypotheses about how these direction- and speed-sensitive neurons contribute to the generation of pursuit. The amplitude and direction of the initial saccade to the target and the initial speed and direction of pursuit were best predicted by a vector-average model of the underlying neuronal activity with relatively short time and spatial separation before a visual pursuit target and a distracter stimulus crossed in the visual field. The resulting eye movements were best described by a winner-take-all model when the time and spatial separation between the two stimuli was increased before the stimuli crossed. Neurons in MT and MST also shifted their activity from that best described by a vector average to a winner-take-all model under the same stimulus conditions. The changes in activity of neurons in both areas were generally similar to each other during these changes in pursuit initiation. Thus a slight alteration in the target motion produced a concurrent shift in both the neuronal processing and the movement execution. We propose that the differences in the oculomotor behavior can be accounted for by shifts in the overlap of active neuronal populations within MT and MST. PMID- 10515962 TI - Modulation of jellyfish potassium channels by external potassium ions. AB - The amplitude of an A-like potassium current (I(Kfast)) in identified cultured motor neurons isolated from the jellyfish Polyorchis penicillatus was found to be strongly modulated by extracellular potassium ([K(+)](out)). When expressed in Xenopus oocytes, two jellyfish Shaker-like genes, jShak1 and jShak2, coding for potassium channels, exhibited similar modulation by [K(+)](out) over a range of concentrations from 0 to 100 mM. jShak2-encoded channels also showed a decreased rate of inactivation and an increased rate of recovery from inactivation at high [K(+)](out). Using site-directed mutagenesis we show that inactivation of jShak2 can be ascribed to an unusual combination of a weak "implicit" N-type inactivation mechanism and a strong, fast, potassium-sensitive C-type mechanism. Interaction between the two forms of inactivation is responsible for the potassium dependence of cumulative inactivation. Inactivation of jShak1 was determined primarily by a strong "ball and chain" mechanism similar to fruit fly Shaker channels. Experiments using fast perfusion of outside-out patches with jShak2 channels were used to establish that the effects of [K(+)](out) on the peak current amplitude and inactivation were due to processes occurring at either different sites located at the external channel mouth with different retention times for potassium ions, or at the same site(s) where retention time is determined by state-dependent conformations of the channel protein. The possible physiological implications of potassium sensitivity of high-threshold potassium A like currents is discussed. PMID- 10515963 TI - Residues in a jellyfish shaker-like channel involved in modulation by external potassium. AB - The jellyfish gene, jShak2, coded for a potassium channel that showed increased conductance and a decreased inactivation rate as [K(+)](out) was increased. The relative modulatory effectiveness of K(+), Rb(+), Cs(+), and Na(+) indicated that a weak-field-strength site is present. Cysteine substituted mutants (L369C and F370C) of an N-terminal truncated construct, (jShak2Delta2-38) which only showed C-type inactivation, were used to establish the position and nature of this site(s). In comparison with jShak2Delta2-38 and F370C, L369C showed a greater relative increase in peak current when [K(+)](out) was increased from 1 to 100 mM because the affinity of this site was reduced at low [K(+)](out). Increasing [K(+)](out) had little effect on the rate of inactivation of L369C; however, the appearance of a second, hyperbolic component to the inactivation curve for F370C indicated that this mutation had increased the affinity of the low-affinity site by bringing the backbone oxygens closer together. Methanethiosulphonate reagents were used to form positively (MTSET), negatively (MTSES), and neutrally (MTSM) charged side groups on the cysteine-substituted residues at the purported K(+) binding site(s) in the channel mouth and conductance and inactivation kinetic measurements made. The reduced affinity of the site produced by the mutation L369C was probably due to the increased hydrophobicity of cysteine, which changed the relative positions of carbonyl oxygens since MTSES modification did not form a high-field-strength site as might be expected if the cysteine residues project into the pore. Addition of the side chain -CH(2)-S-S-CH(3), which is similar to the side chain of methionine, a conserved residue in many potassium channels, resulted in an increased peak current and reduced inactivation rate, hence a higher affinity binding site. Modification of cysteine substituted mutants occurred more readily from the inactivated state confirming that side chains probably rotate into the pore from a buried position when no K ions are in the pore. In conclusion we were able to show that, as for certain potassium channels in higher taxonomic groups, the site(s) responsible for modulation by [K(+)](out) is situated just outside the selectivity filter and is represented by the residues L(369) and F(370) in the jellyfish Shaker channel, jShak2. PMID- 10515964 TI - Increased pyramidal excitability and NMDA conductance can explain posttraumatic epileptogenesis without disinhibition: a model. AB - Partially isolated cortical islands prepared in vivo become epileptogenic within weeks of the injury. In this model of chronic epileptogenesis, recordings from cortical slices cut through the injured area and maintained in vitro often show evoked, long- and variable-latency multiphasic epileptiform field potentials that also can occur spontaneously. These events are initiated in layer V and are synchronous with polyphasic long-duration excitatory and inhibitory potentials (currents) in neurons that may last several hundred milliseconds. Stimuli that are significantly above threshold for triggering these epileptiform events evoke only a single large excitatory postsynaptic potential (EPSP) followed by an inhibitory postsynaptic potential (IPSP). We investigated the physiological basis of these events using simulations of a layer V network consisting of 500 compartmental model neurons, including 400 principal (excitatory) and 100 inhibitory cells. Epileptiform events occurred in response to a stimulus when sufficient N-methyl-D-aspartate (NMDA) conductance was activated by feedback excitatory activity among pyramidal cells. In control simulations, this activity was prevented by the rapid development of IPSPs. One manipulation that could give rise to epileptogenesis was an increase in the threshold of inhibitory interneurons. However, previous experimental data from layer V pyramidal neurons of these chronic epileptogenic lesions indicate: upregulation, rather than downregulation, of inhibition; alterations in the intrinsic properties of pyramidal cells that would tend to make them more excitable; and sprouting of their intracortical axons and increased numbers of presumed synaptic contacts, which would increase recurrent EPSPs from one cell onto another. Consistent with this, we found that increasing the excitability of pyramidal cells and the strength of NMDA conductances, in the face of either unaltered or increased inhibition, resulted in generation of epileptiform activity that had characteristics similar to those of the experimental data. Thus epileptogenesis such as occurs after chronic cortical injury can result from alterations of intrinsic membrane properties of pyramidal neurons together with enhanced NMDA synaptic conductances. PMID- 10515965 TI - Differential modulation of motor neurons that innervate the same muscle but use different excitatory transmitters in aplysia. AB - The medial portion of intrinsic buccal muscle 3 (I3m) is innervated by two excitatory motor neurons, B3 and B9. B3 uses glutamate as its fast transmitter and expresses the neuropeptide FMRFamide, whereas B9 uses acetylcholine as its fast transmitter and expresses the neuropeptide SCP. This preparation was used to study peptidergic modulation of muscles innervated by neurons that use different fast excitatory transmitters. First, we determined the effects of the application of the neuropeptides expressed in these neurons on excitatory junction potentials (EJPs) and contractions. FMRFamide increased the amplitude of EJPs and contractions evoked by B3 while decreasing those evoked by B9. This is the first observation in buccal muscle of a substance that modulates two excitatory neurons innervating the same muscle in opposite directions. SCP increased EJPs contraction amplitude, and the rate of muscle relaxation for both motor neurons. We determined that SCP potently increased cAMP levels in I3m as it does in other buccal muscles. Stimulation of B9 also caused increased cAMP levels in I3m providing independent evidence for SCP release. Finally, stimulation of B9 increased both the contraction amplitude and relaxation rate of B3-evoked I3m contractions in a manner similar to that observed using exogenous SCP. By inhibiting B9's cholinergic transmission with an antagonist, we were able to observe modulatory effects of B9 in the absence of fast excitatory effects. We found that the magnitude of the modulation was dependent on the firing frequency and did occur at frequencies and patterns of firing recorded previously for B9 during ingestive-like motor programs. PMID- 10515966 TI - Dynorphin selectively augments the M-current in hippocampal CA1 neurons by an opiate receptor mechanism. AB - Most electrophysiological studies of opioids on hippocampal principal neurons have found indirect actions, usually through interneurons. However, our laboratory recently found reciprocal alteration of the voltage-dependent K(+) current, known as the M-current (I(M)), by kappa and delta opioid agonists in CA3 pyramidal neurons. Recent ultrastructural studies have revealed postsynaptic delta opiate receptors on dendrites and cell bodies of CA1 and CA3 hippocampal pyramidal neurons (HPNs). Reasoning that previous electrophysiological studies may have overlooked voltage-dependent postsynaptic effects of the opioids in CA1, we reevaluated their role in CA1 HPNs using the rat hippocampal slice preparation for intracellular current- and voltage-clamp recording. None of the delta and mu; receptor-selective opioids tested, including [D-Pen(2,5)]-enkephalin (DPDPE), [D Ala(2)]-deltorphin II (deltorphin), [D-Ala(2), NMe-Phe(4), Gly-ol]-enkephalin (DAMGO), and [D-Ala(2), D-Leu(5)] enkephalin (DADLE), altered membrane properties such as I(M) or Ca(2+)-dependent spikes in CA1 HPNs. The nonopioid, Des-Tyr dynorphin (D-T-dyn), also had no effect. By contrast, dynorphin A (1-17) markedly increased I(M) at low concentrations and caused an outward current at depolarized membrane potentials. The opioid antagonist naloxone and the kappa receptor antagonist nor-binaltorphimine (nBNI) blocked the I(M) effect. However, the kappa selective agonists U69,593 and U50,488h did not significantly alter I(M) amplitudes when averaged over all cells tested, although occasional cells showed an I(M) increase with U50,488h. Our results suggest that dynorphin A postsynaptically modulates the excitability of CA1 HPNs through opiate receptors linked to voltage-dependent K(+) channels. These findings also provide pharmacological evidence for a functional kappa opiate receptor subtype in rat CA1 HPNs but leave unanswered questions on the role of delta receptors in CA1 HPNs. PMID- 10515967 TI - Nociceptin augments K(+) currents in hippocampal CA1 neurons by both ORL-1 and opiate receptor mechanisms. AB - We previously reported (see also the accompanying paper) that dynorphin A significantly enhanced the voltage-dependent K(+) M-current (I(M)) in CA3 and CA1 hippocampal pyramidal neurons (HPNs). Because the opioid-receptor-like-1 (ORL-1) receptor shares a high sequence homology with opioid receptors and is expressed in rat hippocampus, we examined the effects of orphanin FQ or nociceptin, the endogenous ligand for the ORL-1 receptor, using the rat hippocampal slice preparation and intracellular voltage-clamp recording. Current-voltage (I-V) relationships from CA1 HPNs revealed that nociceptin superfusion induced an outward current reversing near the equilibrium potential for K(+) ions. Ba(2+) (2 mM) blocked this effect. The nociceptin-induced current was largest at depolarized membrane potentials, where I(M) is largely activated. Nociceptin concentrations of 0.5-1 microM (but not 0.1 microM) significantly increased I(M) relaxation amplitudes with recovery on washout. Interestingly, both the general opiate antagonist naloxone and the kappa receptor antagonist nor-binaltorphimine (nBNI) inhibited the nociceptin-induced I(M) increases and outward currents in the depolarized range but not the inward current induced at hyperpolarized potentials. The putative ORL-1 receptor antagonist, [Phe(1)Psi(CH(2) NH)Gly(2)]NC(1-13)NH(2) (hereafter ORLAn), blocked most of the nociceptin current near rest but not the I(M) increase. However, ORLAn alone had direct effects similar to those of nociceptin, indicating that ORLAn might be a partial agonist. Our results suggest that nociceptin postsynaptically modulates the excitability of HPNs through ORL-1 and kappa-like opiate receptors linked to different K(+) channels. PMID- 10515968 TI - Synchronized oscillatory discharges of mitral/tufted cells with different molecular receptive ranges in the rabbit olfactory bulb. AB - Individual glomeruli in the mammalian olfactory bulb represent a single or a few type(s) of odorant receptors. Signals from different types of receptors are thus sorted out into different glomeruli. How does the neuronal circuit in the olfactory bulb contribute to the combination and integration of signals received by different glomeruli? Here we examined electrophysiologically whether there were functional interactions between mitral/tufted cells associated with different glomeruli in the rabbit olfactory bulb. First, we made simultaneous recordings of extracellular single-unit spike responses of mitral/tufted cells and oscillatory local field potentials in the dorsomedial fatty acid-responsive region of the olfactory bulb in urethan-anesthetized rabbits. Using periodic artificial inhalation, the olfactory epithelium was stimulated with a homologous series of n-fatty acids or n-aliphatic aldehydes. The odor-evoked spike discharges of mitral/tufted cells tended to phase-lock to the oscillatory local field potential, suggesting that spike discharges of many cells occur synchronously during odor stimulation. We then made simultaneous recordings of spike discharges from pairs of mitral/tufted cells located 300-500 microm apart and performed a cross-correlation analysis of their spike responses to odor stimulation. In approximately 27% of cell pairs examined, two cells with distinct molecular receptive ranges showed synchronized oscillatory discharges when olfactory epithelium was stimulated with one or a mixture of odorant(s) effective in activating both. The results suggest that the neuronal circuit in the olfactory bulb causes synchronized spike discharges of specific pairs of mitral/tufted cells associated with different glomeruli and the synchronization of odor-evoked spike discharges may contribute to the temporal binding of signals derived from different types of odorant receptor. PMID- 10515969 TI - Restrictions on inhibitory circuits contribute to limited recruitment of fast inhibition in rat neocortical pyramidal cells. AB - To further define the operational boundaries on fast inhibition in neocortex, whole cell recordings were made from layer V pyramidal neurons in neocortical slices to evaluate evoked inhibitory postsynaptic currents (IPSCs) and spontaneous miniature IPSCs (mIPSCs). Stimulating electrodes were placed in layers VI and I/II to determine whether simultaneous stimulation of deep and superficial laminae could extend the magnitude of maximal IPSCs evoked by deep layer stimulation alone. The addition of superficial-layer stimulation did not increase maximal IPSC amplitude, confirming the strict limit on fast inhibition. Spontaneous miniature IPSCs were recorded in the presence of tetrodotoxin. The frequency of spontaneous mIPSCs ranged from 10.0 to 33.1 Hz. mIPSC amplitude varied considerably, with a range of 5. 0-128.2 pA and a mean value of 20.7+/-4.1 pA (n = 12 cells). The decay phase of miniature IPSCs was best fit by a single exponential, similar to evoked IPSCs. The mean time constant of decay was 6.4+/ 0.6 ms, with a range of 0.2-20.1 ms. The mean 10-90% rise time was 1.9+/-0.2 ms, ranging from 0.2 to 6.3 ms. Evaluation of mIPSC kinetics revealed no evidence of dendritic filtering. Amplitude histograms of mIPSCs exhibited skewed distributions with several discernable peaks that, when fit with Gaussian curves, appeared to be spaced equidistantly, suggesting that mIPSC amplitudes varied quantally. The mean separation of Gaussian peaks ranged from 6.1 to 7.8 pA. The quantal distributions did not appear to be artifacts of noise. Exposure to saline containing low Ca(2+) and high Mg(2+) concentrations reduced the number of histogram peaks, but did not affect the quantal size. Mean mIPSC amplitude and quantal size varied with cell holding potential in a near-linear manner. Statistical evaluation of amplitude histograms verified the multimodality of mIPSC amplitude distributions and corroborated the equidistant spacing of peaks. Comparison of mIPSC values with published data from single GABA channel recordings suggests that the mean mIPSC conductance corresponds to the activation of 10-20 GABA(A) receptor channels, and that the release of a single inhibitory quantum opens 3-6 channels. Further comparison of mIPSCs with evoked inhibitory events suggests that a single interneuron may form, on average, 4-12 functional synapses with a pyramidal cell, and that 10-12 individual interneurons are engaged during recruitment of maximal population IPSCs. This suggests that inhibitory circuits are much more restricted in both the size of the unit events and effective number of connections when compared with excitatory inputs. PMID- 10515970 TI - Cortical columnar processing in the rat whisker-to-barrel system. AB - Controlled whisker stimulation and single-unit recordings were used to elucidate response transformations that occur during the processing of tactile information from ventral posterior medial thalamus (VPM) through cortical columns in the rat whisker/barrel cortex. Whiskers were either deflected alone, using punctate ramp and-hold stimuli, or in combination with a random noise vibration applied simultaneously to two or more neighboring whiskers. Quantitative data were obtained from five anatomically defined groups of neurons based on their being located in: VPM, layer IV barrels, layer IV septa, supragranular laminae, and infragranular laminae. Neurons in each of these populations displayed characteristic properties related to their response latency and time course, relative magnitudes of responses evoked by stimulus onset versus offset, strength of excitatory responses evoked by the noise stimulus, and/or the degree to which the noise stimulus, when applied to neighboring whiskers, suppressed or facilitated responses evoked by the columnar whisker. Results indicate that within layer IV itself there are at least two anatomically distinct networks, barrel and septum, that independently process afferent information, transforming thalamic input in similar but quantitatively distinguishable ways. Transformed signals are passed on to circuits in supragranular and infragranular laminae. In the case of supragranular neurons, evidence suggests that circuits there function in a qualitatively different fashion from those in layer IV, diminishing response differentials between weak and strong inputs, rather than enhancing them. Compared to layer IV, the greater heterogeneity of receptive field properties in nongranular layers suggests the existence of multiple, operationally distinct local circuits in the output layers of the cortical column. PMID- 10515971 TI - Intrinsic optical signals in rat hippocampal slices during hypoxia-induced spreading depression-like depolarization. AB - In interfaced rat hippocampal slices spreading depression (SD) and hypoxia induced SD-like depolarization are associated with increased light reflectance and decreased light transmittance, indicating increased light scattering. By contrast, mild hypotonicity or electrical stimulation decrease light scattering, which is usually taken to be caused by cell swelling. This difference has been attributed to experimental conditions, but in our laboratory moderate osmotic challenge and SD produced opposite intrinsic optical signals (IOSs) in the same slice under identical conditions. To decide whether the SD-induced IOS is related to cell swelling, we investigated the effects of Cl(-) transport inhibitors and Cl(-) withdrawal on both light reflectance and transmittance, as well as on changes in interstitial volume and tissue electrical resistance. In normal [Cl( )](o), early during hypoxia, there was a slight decrease in light reflectance paired with increase in transmittance. At the onset of hypoxic SD, coincident with the onset of cell swelling (restriction of TMA(+) space), the IOS signals suddenly inverted, indicating sharply increased scattering. The SD-related IOSs started in a single spot and spread out over the entire CA1 region without invading CA3. Application of 2 mM furosemide decreased IOS intensity. When [Cl( )](o) was substituted by methylsulfate or gluconate, the SD-related reflectance increase and transmittance decrease were suppressed and replaced by opposite signals, indicating scattering decrease. Yet Cl(-) withdrawal did not prevent cell swelling measured as shrinkage of TMA(+) space. The SD-related increase of tissue electrical resistance was reduced when bath Cl(-) was replaced by methylsulfate and almost eliminated when replaced by gluconate. The TMA(+) signal is judged to be a more reliable indicator of interstitial space than tissue resistance. Neither application of cyclosporin A nor raising [Mg(2+)](o) depressed the SD-related reflectance increase, suggesting that Cl(-) flux through mitochondrial "megachannels" may not be a major factor in its generation. Fluoroacetate poisoning of glial cells (5 mM) accelerated SD onset and enhanced the SD-induced reflectance increase threefold. This suggests, first, that glial cells normally moderate the SD process and, second, that neurons are the predominant generators of the light-scattering increase. We conclude that light scattering by cerebral tissue can be changed by at least two different physical processes. Cell swelling decreases light scattering, whereas a second process increases scattering. During hypoxic SD the scattering increase masks the swelling-induced scattering decrease, but the latter is revealed when Cl(-) is removed. The scattering increase is Cl(-) dependent, nevertheless it is apparently not related to cell volume changes. Its underlying mechanism is as yet not clear; possible factors are discussed. PMID- 10515972 TI - Rhythmically firing neostriatal neurons in monkey: activity patterns during reaction-time hand movements. AB - While previous studies have identified rhythmically firing neurons (RFNs) in monkey neostriatum and these rhythmic firing patterns have been shown to evolve in neostriatal tonically active neurons (TANs) after dopamine input depletion, the activity patterns of RFNs during motor behavior are still far from completely understood. We examined the single-unit activity patterns of neostriatal neurons, recorded in awake behaving monkeys during a wrist movement task, for evidence of rhythmic activity. Monkeys made ballistic wrist flexion and extension movements in response to vibrotactile cues. Animals held a steady wrist position for 0.5 to 2.0 s while awaiting the onset of the go-cues (hold period). Although the majority of neostriatal neurons (274/306) did not fire rhythmically, approximately 10% of the neurons (32/306) fired rhythmically at 10-50 Hz during the hold period. Most RFNs (28/32) showed significant activity changes during the time between go-cue presentation and movement onset (premovement activity). One half of RFNs exhibited premovement activity that differed as a function of movement direction. Only one RFN may have responded to the delivery of a fruit juice reward. Neuronal firing was analyzed using interspike interval distributions, autocorrelations, and serial correlation techniques. These analyses showed that the activity patterns of most RFNs were consistent with an integrate-and-fire model of neuronal rhythm generation. Changes in RFN activity patterns during the premovement interval and intertrial variations in firing frequency could be explained by changes in the general level of excitatory input. These observations are consistent with the firing properties reported for neostriatal cholinergic interneurons. It has been suggested that tonically active neurons may be cholinergic interneurons and that these neurons show changes in activity related to specific aspects of behavioral paradigms, such as rewards. RFNs may constitute a special class of TANs. The results presented here suggest that RFNs may have a role in movement initiation. We speculate that RFNs may modulate the propagation of cortical oscillations via basal ganglia-thalamic cortical loops. PMID- 10515973 TI - Physiological properties of central medial and central lateral amygdala neurons. AB - Mounting evidence implicates the central (CE) nucleus of the amygdala in the mediation of classically conditioned fear responses. However, little data are available regarding the intrinsic membrane properties of CE amygdala neurons. Here, we characterized the physiological properties of CE medial (CE(M)) and CE lateral (CE(L)) amygdala neurons using whole cell recordings in brain slices maintained in vitro. Several classes of CE neurons were distinguished on the basis of their physiological properties. Most CE(M) cells (95%), here termed "late-firing neurons," displayed a marked voltage- and time-dependent outward rectification in the depolarizing direction. This phenomenon was associated with a conspicuous delay between the onset of depolarizing current pulses and the first action potential. During this delay, the membrane potential (V(m)) depolarized slowly, the steepness of this depolarizing ramp increasing as the prepulse V(m) was hyperpolarized from -60 to -90 mV. Low extracellular concentrations of 4-aminopyridine (30 microM) reversibly abolished the outward rectification and the delay to firing. Late-firing CE(M) neurons displayed a continuum of repetitive firing properties with cells generating single spikes at one pole and high-frequency (> or =90 Hz) spike bursts at the other. In contrast, only 56% of CE(L) cells displayed the late-firing behavior prevalent among CE(M) neurons. Moreover, these CE(L) neurons only generated single spikes in response to membrane depolarization. A second major class of CE(L) cells (38%) lacked the characteristic delay to firing observed in CE(M) cells, generated single spikes in response to membrane depolarization, and displayed various degrees of inward rectification in the hyperpolarizing direction. In both regions of the CE nucleus, two additional cell types were encountered infrequently (< or =6% of our samples). One type of neurons, termed "low-threshold bursting cells" had a behavior reminiscent of thalamocortical neurons. The second type of cells, called "fast-spiking cells," generated brief action potentials at high rates with little spike frequency adaptation in response to depolarizing current pulses. These findings indicate that the CE nucleus contains several types of neurons endowed with distinct physiological properties. Moreover, these various cell types are not distributed uniformly in the medial and lateral sector of the CE nucleus. This heterogeneity parallels anatomic data indicating that these subnuclei are part of different circuits. PMID- 10515974 TI - Increased gamma- and decreased delta-oscillations in a mouse deficient for a potassium channel expressed in fast-spiking interneurons. AB - Kv3.1 is a voltage-gated, fast activating/deactivating potassium (K(+)) channel with a high-threshold of activation and a large unit conductance. Kv3.1 K(+) channels are expressed in fast-spiking, parvalbumin-containing interneurons in cortex, hippocampus, striatum, the thalamic reticular nucleus (TRN), and in several nuclei of the brain stem. A high density of Kv3.1 channels contributes to short-duration action potentials, fast afterhyperpolarizations, and brief refractory periods enhancing the capability in these neurons for high-frequency firing. Kv3.1 K(+) channel expression in the TRN and cortex also suggests a role in thalamocortical and cortical function. Here we show that fast gamma and slow delta oscillations recorded from the somatomotor cortex are altered in the freely behaving Kv3.1 mutant mouse. Electroencephalographic (EEG) recordings from homozygous Kv3.1(-/-) mice show a three- to fourfold increase in both absolute and relative spectral power in the gamma frequency range (20-60 Hz). In contrast, Kv3.1-deficient mice have a 20-50% reduction of power in the slow delta range (2 3 Hz). The increase in gamma power is most prominent during waking in the 40- to 55-Hz range, whereas the decrease in delta power occurs equally across all states of arousal. Our findings suggest that Kv3. 1-expressing neurons are involved in the generation and maintenance of cortical fast gamma and slow delta oscillations. Hence the Kv3. 1-mutant mouse could serve as a model to study the generation and maintenance of fast gamma and slow delta rhythms and their involvement in behavior and cognition. PMID- 10515975 TI - Cutaneous receptive field organization in the ventral posterior nucleus of the thalamus in the common marmoset. AB - The organization of cutaneous receptive fields in the ventroposterior (VP) thalamus of the common marmosets (Callithrix jacchus) was determined from single unit recordings, and these data were correlated with the cytochrome oxidase (CO) histochemistry of the thalamus in the same animals. Under continuously maintained ketamine anesthesia, the receptive fields of a total of 192 single units were recorded from the right VP thalamus using 2 MOhms glass electrodes. After the receptive fields were mapped, the brains were reacted for CO histochemistry on 50 microm coronal frozen sections through the entire VP thalamus. The majority of units were localized to the CO-reactive regions that define the medial and lateral divisions of VP (VPm and VPl). Apart from the expected finding of the face being represented in VPm and the body in VPl, reconstructing the electrode tracks and unit locations in the histological sections revealed a general association between discrete regions of CO reactivity and the representation of specific body regions. Some low-threshold cutaneous units were apparently localized to VPi (the CO weak regions dorsal, ventral, and interdigitating with, the CO regions of VP). These VPi units were clearly part of the same representational map as the VPl and VPm units. We conclude that the low-threshold cutaneous receptive fields of the marmoset are organized in a single continuous representation of the contralateral body surface, and that this representation can most simply be interpreted as being folded or crumpled into the three dimensional space of VP thalamus. The folded nature of the body map in VP may be related to the folded nature of VP as revealed by CO histochemistry. PMID- 10515976 TI - Comparative study of viscerosomatic input onto postsynaptic dorsal column and spinothalamic tract neurons in the primate. AB - The purpose of the present investigation was to examine, in the primate, the role of the postsynaptic dorsal column (PSDC) system and that of the spinothalamic tract (STT) in viscerosensory processing by comparing the responses of neurons in these pathways to colorectal distension (CRD). Experiments were done on four anesthetized male monkeys (Macaca fascicularis). Extracellular recordings were made from a total of 100 neurons randomly located in the L(6)-S(1) segments of the spinal cord. Most of these neurons had cutaneous receptive fields in the perineal area, on the hind limbs or on the rump. Forty-eight percent were PSDC neurons activated antidromically from the upper cervical dorsal column or the nucleus gracilis, 17% were STT neurons activated antidromically from the thalamus, and 35% were unidentified. Twenty-one PSDC neurons, located mostly near the central canal, were excited by CRD and three were inhibited. Twenty-four PSDC neurons, mostly located in the nucleus proprius, did not respond to CRD. Of the 17 STT neurons, 7 neurons were excited by CRD, 4 neurons were inhibited, and 6 neurons did not respond to CRD. Of the unidentified neurons, 23 were excited by CRD, 7 were inhibited, and 5 did not respond. The average responses of STT and PSDC neurons excited by CRD were comparable in magnitude and duration. These results suggest that the major role of the PSDC pathway in viscerosensory processing may be due to a quantitative rather than a qualitative neuronal dominance over the STT. PMID- 10515977 TI - Mossy fiber-granule cell synapses in the normal and epileptic rat dentate gyrus studied with minimal laser photostimulation. AB - Dentate granule cells become synaptically interconnected in the hippocampus of persons with temporal lobe epilepsy, forming a recurrent mossy fiber pathway. This pathway may contribute to the development and propagation of seizures. The physiology of mossy fiber-granule cell synapses is difficult to characterize unambiguously, because electrical stimulation may activate other pathways and because there is a low probability of granule cell interconnection. These problems were addressed by the use of scanning laser photostimulation in slices of the caudal hippocampal formation. Glutamate was released from a caged precursor with highly focused ultraviolet light to evoke action potentials in a small population of granule cells. Excitatory synaptic currents were recorded in the presence of bicuculline. Minimal laser photostimulation evoked an apparently unitary excitatory postsynaptic current (EPSC) in 61% of granule cells from rats that had experienced pilocarpine-induced status epilepticus followed by recurrent mossy fiber growth. An EPSC was also evoked in 13-16% of granule cells from the control groups. EPSCs from status epilepticus and control groups had similar peak amplitudes ( approximately 30 pA), 20-80% rise times (approximately 1.2 ms), decay time constants ( approximately 10 ms), and half-widths (approximately 8 ms). The mean failure rate was high (approximately 70%) in both groups, and in both groups activation of N-methyl-D-aspartate receptors contributed a small component to the EPSC. The strong similarity between responses from the status epilepticus and control groups suggests that they resulted from activation of a similar synaptic population. No EPSC was recorded when the laser beam was focused in the dentate hilus, suggesting that indirect activation of hilar mossy cells contributed little, if at all, to these results. Recurrent mossy fiber growth increases the density of mossy fiber-granule cell synapses in the caudal dentate gyrus by perhaps sixfold, but the new synapses appear to operate very similarly to preexisting mossy fiber-granule cell synapses. PMID- 10515978 TI - Dendritic voltage-gated ion channels regulate the action potential firing mode of hippocampal CA1 pyramidal neurons. AB - The role of dendritic voltage-gated ion channels in the generation of action potential bursting was investigated using whole cell patch-clamp recordings from the soma and dendrites of CA1 pyramidal neurons located in hippocampal slices of adult rats. Under control conditions somatic current injections evoked single action potentials that were associated with an afterhyperpolarization (AHP). After localized application of 4-aminopyridine (4-AP) to the distal apical dendritic arborization, the same current injections resulted in the generation of an afterdepolarization (ADP) and multiple action potentials. This burst firing was not observed after localized application of 4-AP to the soma/proximal dendrites. The dendritic 4-AP application allowed large-amplitude Na(+)-dependent action potentials, which were prolonged in duration, to backpropagate into the distal apical dendrites. No change in action potential backpropagation was seen with proximal 4-AP application. Both the ADP and action potential bursting could be inhibited by the bath application of nonspecific concentrations of divalent Ca(2+) channel blockers (NiCl and CdCl). Ca(2+) channel blockade also reduced the dendritic action potential duration without significantly affecting spike amplitude. Low concentrations of TTX (10-50 nM) also reduced the ability of the CA1 neurons to fire in the busting mode. This effect was found to be the result of an inhibition of backpropagating dendritic action potentials and could be overcome through the coordinated injection of transient, large-amplitude depolarizing current into the dendrite. Dendritic current injections were able to restore the burst firing mode (represented as a large ADP) even in the presence of high concentrations of TTX (300-500 microM). These data suggest the role of dendritic Na(+) channels in bursting is to allow somatic/axonal action potentials to backpropagate into the dendrites where they then activate dendritic Ca(2+) channels. Although it appears that most Ca(2+) channel subtypes are important in burst generation, blockade of T- and R-type Ca(2+) channels by NiCl (75 microM) inhibited action potential bursting to a greater extent than L-channel (10 microM nimodipine) or N-, P/Q-type (1 microM omega-conotoxin MVIIC) Ca(2+) channel blockade. This suggest that the Ni-sensitive voltage-gated Ca(2+) channels have the most important role in action potential burst generation. In summary, these data suggest that the activation of dendritic voltage-gated Ca(2+) channels, by large-amplitude backpropagating spikes, provides a prolonged inward current that is capable of generating an ADP and burst of multiple action potentials in the soma of CA1 pyramidal neurons. Dendritic voltage-gated ion channels profoundly regulate the processing and storage of incoming information in CA1 pyramidal neurons by modulating the action potential firing mode from single spiking to burst firing. PMID- 10515979 TI - Effect of divalent cations on AMPA-evoked extracellular alkaline shifts in rat hippocampal slices. AB - The generation of activity-evoked extracellular alkaline shifts has been linked to the presence of external Ca(2+) or Ba(2+). We further investigated this dependence using pH- and Ca(2+)-selective microelectrodes in the CA1 area of juvenile, rat hippocampal slices. In HEPES-buffered media, alkaline transients evoked by pressure ejection of RS-alpha-amino-3-hydroxy-5-methyl-4 isoxazolepropionic acid (AMPA) averaged approximately 0.07 unit pH and were calculated to arise from the equivalent net addition of approximately 1 mM strong base to the interstitial space. These alkaline responses were correlated with a mean decrease in [Ca(2+)](o) of approximately 300 microM. The alkalinizations were abolished reversibly in zero-Ca(2+) media, becoming indiscernible at a [Ca(2+)](o) of 117+/-29 microM. Addition of as little as 30-50 microM Ba(2+) caused the reappearance of an alkaline response. In approximately one-fourth of slices, a persistent alkaline shift of approximately 0.03 unit pH was observed in zero-Ca(2+) saline containing EGTA. In HEPES media, addition of 300 microM Cd(2+), 100 microM Ni(2+), or 100 microM nimodipine inhibited the alkaline shifts by roughly one-half, one-third, and one-third, respectively, whereas Cd(+) and Ni(2+) in combination fully blocked the response. In bicarbonate media, by contrast, Cd(+) and Ni(2+) blocked only two-thirds of the response. In the presence of bicarbonate, Ni(2+) caused an unexpected enhancement of the alkalinization by approximately 150%. However, when the extracellular carbonic anhydrase was blocked by benzolamide, addition of Ni(2+) reduced the alkaline shift. These results suggested that Ni(2+) partially inhibited the interstitial carbonic anhydrase and thereby increased the alkaline responses. These data indicate that an activity-dependent alkaline shift is largely dependent on the entry of Ca(2+) or Ba(2+) via voltage-gated calcium channels. However, sizable alkaline transients still can be generated with little or no external presence of these ions. Implications for the mechanism of the activity-dependent alkaline shift are discussed. PMID- 10515980 TI - Muscarinic control of dendritic excitability and Ca(2+) signaling in CA1 pyramidal neurons in rat hippocampal slice. AB - The cholinergic system is critically involved in synaptic models of learning and memory by enhancing dendritic [Ca(2+)](i) signals. Diffuse cholinergic innervation suggests subcellular modulation of membrane currents and Ca(2+) signals. Here we use ion-selective microelectrodes to study spread of carbachol (CCh) after focal application into brain slice and subcellular muscarinic modulation of synaptic responses in CA1 pyramidal neurons. Proximal application of CCh rapidly blocked the somatic slow afterhyperpolarization (sAHP) following repetitive stimulation. In contrast, the time course of potentiation of the slow tetanic depolarization (STD) during synaptic input was slower and followed the time course of spread of CCh to the dendritic tree. With distal application, augmentation of the somatic STD and of dendritic Ca(2+) responses followed spread of CCh to the entire apical dendritic tree, whereas the sAHP was blocked only after spread of CCh to the proximal dendritic segment. In dendritic recordings, CCh blocked a small sAHP, augmented the STD, and rather reduced dendritic action potentials. Augmentation of dendritic Ca(2+) signals was highly correlated to augmentation of the STD. The NMDA receptor antagonist DL-2-amino-5 phosphonovaleric acid (APV) blocked approximately 55% of the STD in control and during CCh application. In conclusion, muscarinic suppression of the proximal sAHP can augment firing and thereby Ca(2+) responses. Dendritic augmentation of the STD by blockade of the sAHP and direct enhancement of N-methyl-D-aspartate (NMDA) receptor-mediated currents potentiates Ca(2+) signals even when firing is not affected due to suprathreshold input. In this way, subcellular muscarinic modulation may contribute to parallel information processing and storage by dendritic synapses of CA1 pyramidal neurons. PMID- 10515981 TI - Convergent mechanosensory input structures the firing phase of a steering motor neuron in the blowfly, Calliphora. AB - The first basalar muscle (B1) is 1 of 17 small steering muscles in flies that control changes in wing stroke kinematics during flight. The B1 is often tonically active, firing a single phase-locked action potential in each and every wingbeat cycle. Changes in activation phase alter the biomechanical properties of B1, which in turn cause aerodynamically relevant changes in wing motion. The phase-locked firing of the B1 motor neuron (MNB1), is thought to arise from an interaction of wingbeat-synchronous inputs from the wings and from specialized equilibrium organs called halteres that beat antiphase to the wings and function to detect angular rotation of the body during flight. We investigated how the wing and haltere inputs interact to determine the firing phase of MNB1. Our results indicate that both wing and haltere afferents make strong monosynaptic connections with MNB1, consisting of fast electrical and slow Ca(2+)-sensitive components. Although both the wing and haltere-evoked excitatory postsynaptic potentials (EPSPs) display the two components, their relative contribution is different for the two inputs. Whereas the haltere-evoked EPSP is dominated by the fast electrical component, the wing-evoked EPSP is dominated by a large chemically mediated component and displays an additional prolonged Ca(2+) dependent component that is absent in the haltere-evoked EPSP. Both inputs display an activity-dependent fatigue affecting both electrical and Ca(2+) sensitive components, from which the haltere synapse recovers more rapidly. The net result of these synaptic differences is that the two pathways differ significantly in their relative ability to evoke action potentials in MNB1. Although the haltere pathway displays greater temporal precision, the wing pathway is stronger, judged by its ability to entrain MNB1 within a background of haltere stimulation. We propose a model by which these physiological differences play a functional role in tuning the firing phase of MNB1 during flight. The wing input may serve primarily to set the background firing phase of MNB1, whereas the haltere input serves to transiently advance the firing phase during equilibrium reflexes. PMID- 10515982 TI - Group I mGluR agonist DHPG facilitates the induction of LTP in rat prelimbic cortex in vitro. AB - Long-term potentiation (LTP) of synaptic transmission is a favored neural model for learning and memory. In isolated slices of rat prelimbic cortex, glutamatergic activation of metabotropic receptors (mGluRs) is required for the production of LTP at synapses on layer V neurons. Group I mGluRs are found in neocortex, and in prelimbic cortex they have been located on layer V neurons. We have now investigated whether application of the selective group I mGluR agonist, (S)-3,5-dihydroxyphenylglycine (DHPG) facilitates the induction of LTP. We recorded field potentials in layer V in response to test shocks applied to layer II and measured the population spike peak amplitude and slope. Intracellular recording was used to examine the correspondence between excitatory postsynaptic potentials (EPSPs) and action potentials with components of the field potential, and to further investigate the action of DHPG. Repetitive bursts of stimulation at theta frequencies (TBS) did not consistently alter the magnitude or slope of the population spike (mean response 105+/-4%, mean+/-SE of control at 30 min after TBS ended, n = 9 slices, no significant difference). When DHPG was added to the bathing medium for 10 min during continued test stimulation, the slope and amplitude of the population spike were significantly reduced, but 30 min after wash out of the DHPG, they recovered (mean response 89+/-10% of control, n = 6 slices, no significant difference). However, when TBS was administered in conjunction with bath application of DHPG, LTP of the population spike was induced (mean response 147+/-12% of control at 30 min after TBS ended, P = 0.004, paired t-test, n = 9 slices). We conclude that co-application of DHPG with TBS facilitates the induction of LTP of the population spike, which supports a role for group I mGluRs in the activity-dependent induction of LTP in the prelimbic cortex. PMID- 10515984 TI - Response latency of macaque area MT/V5 neurons and its relationship to stimulus parameters. AB - A total of 310 MT/V5 single cells were tested in anesthetized, paralyzed macaque monkeys with moving random-dot stimuli. At optimum stimulus parameters, latencies ranged from 35 to 325 ms with a mean of 87+/-45 (SD) ms. By examining the relationship between latency and response levels, stimulus parameters, and stimulus selectivities, we attempted to isolate the contributions of these factors to latency and to identify delays representing intervening synapses (circuitry) and signal processing (flow of information through that circuitry). First, the relationship between stimulus parameters and latency was investigated by varying stimulus speed and direction for individual cells. Resulting changes in latencies were explainable in terms of response levels corresponding to how closely the actual stimulus matched the preferred stimulus of the cell. Second, the relationship between stimulus selectivity and latency across the population of cells was examined using the optimum speed and direction of each neuron. A weak tendency for cells tuned for slow speeds to have longer latencies was explainable by lower response rates among slower-tuned neurons. In contrast, sharper direction tuning was significantly associated with short latencies even after taking response rate into account, (P = 0.002, ANCOVA). Accordingly, even the first 10 ms of the population response fully demonstrates direction tuning. A third study, which examined the relationship between antagonistic surrounds and latency, revealed a significant association between the strength of the surround and the latency that was independent of response levels (P < 0.002, ANCOVA). Neurons having strong surrounds exhibited latencies averaging 20 ms longer than those with little or no surround influence, suggesting that neurons with surrounds represent a later stage in processing with one or more intervening synapses. The laminar distribution of latencies closely followed the average surround antagonism in each layer, increasing with distance from input layer IV but precisely mirroring response levels, which were highest near the input layer and gradually decreased with distance from input layer IV. Layer II proved the exception with unexpectedly shorter latencies (P< 0.02, ANOVA) yet showing only modest response levels. The short latency and lack of strong direction tuning in layer II is consistent with input from the superior colliculus. Finally, experiments with static stimuli showed that latency does not vary with response rate for such stimuli, suggesting a fundamentally different mode of processing than that for a moving stimulus. PMID- 10515983 TI - Pain intensity processing within the human brain: a bilateral, distributed mechanism. AB - Functional imaging studies of human subjects have identified a diverse assortment of brain areas that are engaged in the processing of pain. Although many of these brain areas are highly interconnected and are engaged in multiple processing roles, each area has been typically considered in isolation. Accordingly, little attention has been given to the global functional organization of brain mechanisms mediating pain processing. In the present investigation, we have combined positron emission tomography with psychophysical assessment of graded painful stimuli to better characterize the multiregional organization of supraspinal pain processing mechanisms and to identify a brain mechanism subserving the processing of pain intensity. Multiple regression analysis revealed statistically reliable relationships between perceived pain intensity and activation of a functionally diverse group of brain regions, including those important in sensation, motor control, affect, and attention. Pain intensity related activation occurred bilaterally in the cerebellum, putamen, thalamus, insula, anterior cingulate cortex, and secondary somatosensory cortex, contralaterally in the primary somatosensory cortex and supplementary motor area, and ipsilaterally in the ventral premotor area. These results confirm the existence of a highly distributed, bilateral supraspinal mechanism engaged in the processing of pain intensity. The conservation of pain intensity information across multiple, functionally distinct brain areas contrasts sharply with traditional views that sensory-discriminative processing of pain is confined within the somatosensory cortex and can account for the preservation of conscious awareness of pain intensity after extensive cerebral cortical lesions. PMID- 10515985 TI - Robust suppression of afferent-induced excitation in the rat spinal dorsal horn after conditioning low-frequency stimulation. AB - The neuronal plasticity in the spinal dorsal horn induced after conditioning low frequency stimulation of afferent A fibers, and its relationship with spinal inhibitory networks, was investigated with an optical-imaging method that detects neuronal excitation. High-intensity single-pulse stimulation of the dorsal root activating both A and C fibers evoked an optical response in the dorsal horn in transverse slices of 12- to 25-day-old rat spinal cords stained with a voltage sensitive dye, RH-482. The optical response, reflecting the net excitation of neuronal elements along the thickness of each slice, was suppressed after a conditioning low-frequency stimulation (0.2-1 Hz for 10 min) to A fibers in the dorsal root. The degree of suppression was largest in the lamina II of the dorsal horn (48% reduction), where the majority of C fibers terminate, and much less in the deeper dorsal horn (5% reduction in laminae III-IV). The onset of suppression was somewhat slow; after the low-frequency stimulation, the magnitude of excitation gradually decreased, reached the maximum effect 30 min after the conditioning, and remained at the suppressed level for >1 h. Suppression was not observed when the low-frequency stimulation was given during a 20-min perfusion with a solution containing an NMDA-receptor antagonist, DL-2-amino-5 phosphonovaleric acid (30 microM). A brief application of an opioid-receptor antagonist, naloxone (0.5 microM), inhibited the induction, but not the maintenance, of low-frequency stimulus-induced suppression. However, treatments with the GABA(A) receptor antagonist bicuculline (1 microM) and the glycine receptor antagonist strychnine (0.3 microM) did not affect suppression induction and maintenance. In conclusion, conditioning low-frequency stimulation to A fibers interferes with the afferent-induced excitation in the dorsal horn. The low-frequency stimulation-induced suppression is maintained by a reduction of glutamatergic excitatory transmissions in the dorsal horn, not by an enhanced inhibition. Activation of the spinal opioid-mediated system by low-frequency stimulation, but not the inhibitory amino acid-mediated system, is necessary to initiate robust suppression. The long-term depression of afferent synaptic efficacy onto excitatory interneurons likely takes the primary role in the robust suppression of neuronal excitation in the dorsal horn. PMID- 10515986 TI - Epileptiform activity can be initiated in various neocortical layers: an optical imaging study. AB - The initiation site for triggering epileptiform activity was investigated via optical imaging using voltage-sensitive dyes in the neocortical slice perfused with artificial cerebral spinal fluid containing nominally zero magnesium. The neocortical slices (400-microm thick) were harvested from Sprague-Dawley rats (P21-28). Optical imaging was made by using a high speed photodiode array. Spontaneous epileptiform activity emerged 20-40 min after the preparation was perfused with zero-magnesium solution. There was a good correspondence between electrical and optical signals (n = 46), although the details of the two recordings were somewhat different. The initiation sites were measured optically in 11 preparations. Among them, four were found to be located in superficial layers, two were found in middle layers, and five were found in deep layers. Repeated recordings revealed that these initiation sites were relatively stable; shifting of the initiation site was not observed. Therefore spontaneous epileptiform activity could be initiated in various cortical layers, from layer I to layer VI. The activation started from a small area <0.04 mm(3) and spread smoothly from the initiation site to adjacent cortical areas, suggesting that the initiation site is very confined to one of the cortical layers. The initiation sites were distributed randomly in various cortical areas, and no higher probability was found in a special cortical region. Electrical stimulation delivered via a glass microelectrode filled with 2 M NaCl (2-5 MOhms) could reliably trigger epileptiform activity that had the same characteristics as the spontaneous activity. The cortical neurons activated directly by the stimulation were around the electrode's tip and estimated to be within a 50-microm area, suggesting that only a few neurons were needed to form an initiation site. Because the timing for stimulation was arbitrary and the evoked events were initiated independent of discharges of neurons in any other layers, it is likely that the initiation site for epileptiform activity in various cortical layers is independent of the control of layer V pyramidal neurons. Together these finding suggest that the epileptiform focus is confined and can be formed in several (probably all) neocortical layers and in many cortical areas. The initiating neurons may be of different types because neuronal types in various cortical layers are different. PMID- 10515987 TI - Group I metabotropic glutamate receptors mediate an inward current in rat substantia nigra dopamine neurons that is independent from calcium mobilization. AB - Metabotropic glutamate receptors modulate neuronal excitability via a multitude of mechanisms, and they have been implicated in the pathogenesis of neurodegenerative processes. Here we investigated the responses mediated by group I metabotropic glutamate receptors (mGluRs) in dopamine neurons of the rat substantia nigra pars compacta, using whole cell patch-clamp recordings in combination with microfluorometric measurements of [Ca(2+)](i) and [Na(+)](i). The selective group I mGluR agonist (S)-3,5-dihydroxyphenylglycine (3,5-DHPG) was bath-applied (20 microM, 30 s to 2 min) or applied locally by means of short lasting (2-4 s) pressure pulses, delivered through an agonist-containing pipette positioned close to the cell body of the neuron. 3,5-DHPG evoked an inward current characterized by a transient and a sustained component, the latter of which was uncovered only with long-lasting agonist applications. The fast component coincided with a transient elevation of [Ca(2+)](i), whereas the total current was associated with a rise in [Na(+)](i). These responses were not affected either by the superfusion of ionotropic excitatory amino acid antagonists 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and D-2-amino-5-phosphono pentanoic acid (D-APV), nor by the sodium channel blocker tetrodotoxin (TTX). (S) alpha-methyl-4-carboxyphenylglycine (S-MCPG) and the more selective mGluR1 antagonist 7(hydroxyimino)cyclopropa[b]chromen-1a-carboxylate (CPCCOEt) depressed both 3,5-DHPG-induced inward current components and, although less effectively, the associated [Ca(2+)](i) elevations. On repeated agonist applications the inward current and the calcium transients both desensitized. The time constant of recovery from desensitization differed significantly between these two responses, being 67.4+/-4.4 s for the inward current and 28.6+/-2.7 s for the calcium response. Bathing the tissue in a calcium-free/EGTA medium or adding thapsigargin (1 microM) to the extracellular medium prevented the generation of the [Ca(2+)](i) transient, but did not prevent the activation of the inward current. These electrophysiological and fluorometric results show that the 3, 5-DHPG induced inward current and the [Ca(2+)](i) elevations are mediated by independent pathways downstream the activation of mGluR1. PMID- 10515988 TI - Responses of contralateral SI and SII in cat to same-site cutaneous flutter versus vibration. AB - The methods of (14)C-2-deoxyglucose ((14)C-2DG) metabolic mapping and optical intrinsic signal (OIS) imaging were used to evaluate the response evoked in the contralateral primary somatosensory receiving areas (SI and SII) of anesthetized cats by either 25 Hz ("flutter") or 200 Hz ("vibration") sinusoidal vertical skin displacement stimulation of the central pad on the distal forepaw. Unilateral 25 Hz stimulation consistently evoked a localized region of elevated (14)C-2DG uptake in both SI and SII in the contralateral hemisphere. In contrast, 200-Hz stimulation did not evoke elevated (14)C-2DG uptake in the contralateral SI but evoked a prominent, localized region of increased (14)C-2DG uptake in the contralateral SII. Experiments in which the OIS was recorded yielded results that complemented and extended the findings obtained with the 2DG method. First, 25-Hz central-pad stimulation evoked an increase in absorbance in a region in the contralateral SI and SII that corresponded closely to the region in which a similar stimulus evoked increased (14)C-2DG uptake. Second, 200-Hz stimulation of the central pad consistently evoked a substantial increase in absorbance in the contralateral SII but very little or no increase in absorbance in the contralateral SI. And third, 200-Hz central-pad stimulation usually evoked a decrease in absorbance in the same contralateral SI region that underwent an increase in absorbance during same-site 25-Hz stimulation. Experiments in which the OIS responses of both SI and SII were recorded simultaneously demonstrated that continuous (>1 s) 25-Hz central-pad stimulation evokes a prominent increase in absorbance in both SI and SII in the contralateral hemisphere, whereas only SII undergoes a sustained prominent increase in absorbance in response to 200-Hz stimulation to the same central-pad site. SI exhibits an initial, transient increase in absorbance in response to 200-Hz stimulation and at durations of stimulation >1 s, undergoes a decrease in absorbance. It was found that the stimulus-evoked absorbance changes in the contralateral SI and SII are correlated significantly during vibrotactile stimulation of the central pad-positively with 25-Hz stimulation and negatively with 200-Hz stimulation. The findings are interpreted to indicate that 25-Hz central-pad stimulation of the central pad evokes spatially localized and vigorous neuronal activation within both SI and SII in the contralateral hemisphere and that although 200-Hz stimulation evokes vigorous and well maintained neuronal activation within the contralateral SII, the principal effect on the contralateral SI of a 200-Hz stimulus lasting >1 s is inhibitory. PMID- 10515989 TI - Action potential-induced dendritic calcium dynamics correlated with synaptic plasticity in developing hippocampal pyramidal cells. AB - In hippocampal CA1 pyramidal cells, intracellular calcium increases are required for induction of long-term potentiation (LTP), an activity-dependent synaptic plasticity. LTP is known to develop in magnitude during the second and third postnatal weeks in the rats. Little is known, however, about development of intracellular calcium dynamics during the same postnatal weeks. We investigated postnatal development of intracellular calcium dynamics in the proximal apical dendrites of CA1 pyramidal cells by whole cell patch-clamp recordings and calcium imaging with the Ca(2+) indicator fura-2. Dendritic calcium increases induced by intrasomatically evoked action potentials were slight during the first postnatal week but gradually became robust 3 to 6-fold during the second and third postnatal weeks. These calcium increases were blocked by application of 250 microM CdCl(2), a nonspecific blocker for high-threshold voltage-dependent calcium channels (VDCCs). Under the voltage-clamp condition, both calcium currents and dendritic calcium accumulations induced by depolarization were larger at the late developmental stage (P15-18) than the early stage (P4-7), indicating developmental enhancement of calcium influx mediated by high-threshold VDCCs. Moreover, theta-burst stimulation (TBS), a protocol for LTP induction, induced large intracellular calcium increases at the late developmental stage, in synchrony with maturation of TBS-induced LTP. These results suggest that developmental enhancement of intracellular calcium increases induced by action potentials may underlie maturation of calcium-dependent functions such as synaptic plasticity in hippocampal neurons. PMID- 10515990 TI - High-frequency dynamics of regularly discharging canal afferents provide a linear signal for angular vestibuloocular reflexes. AB - Regularly discharging vestibular-nerve afferents innervating the semicircular canals were recorded extracellularly in anesthetized chinchillas undergoing high frequency, high-velocity sinusoidal rotations. In the range from 2 to 20 Hz, with peak velocities of 151 degrees/s at 6 Hz and 52 degrees/s at 20 Hz, 67/70 (96%) maintained modulated discharge throughout the sinusoidal stimulus cycle without inhibitory cutoff or excitatory saturation. These afferents showed little harmonic distortion, no dependence of sensitivity on peak amplitude of stimulation, and no measurable half-cycle asymmetry. A transfer function fitting the data predicts no change in sensitivity (gain) of regularly discharging afferents over the frequencies tested but shows a phase lead with regard to head velocity increasing from 0 degrees at 2 Hz to 30 degrees at 20 Hz. These results indicate that regularly discharging afferents provide a plausible signal to drive the angular vestibuloocular reflex (VOR) even during high-frequency head motion but are not a likely source for nonlinearities present in the VOR. PMID- 10515991 TI - Maturation of lobster stomatogastric ganglion rhythmic activity. AB - The stomatogastric ganglion of the adult lobster, Homarus americanus generates extremely regular pyloric rhythms with a characteristic period of 0.5-1.5 Hz. To study the changes in the pyloric rhythm during embryonic and larval development, we recorded excitatory junctional potentials evoked by lateral pyloric (LP) neuron activity. Early in development the motor discharge of the LP neuron was often irregular, preventing use of conventional analysis methods that rely on extracting burst times to calculate cycle frequency and its variability. Instead, cycle frequency was determined for the LP neuron from the peak of the power spectrum obtained from the occurrence times of excitatory junctional potentials in the p1 muscle. The ratio of the power in the peak to the power from 0 to 3 Hz was used as a relative measure of the regularity of the rhythm. Throughout embryonic and the first larval stage, LP neuron activity is slow, irregular, and only weakly periodic. The regularity of the rhythm increased during midlarval stages, and both the frequency and regularity increased considerably by the postlarval stage LIV. PMID- 10515992 TI - Model for the translational vestibuloocular reflex (VOR). AB - The function of the translational vestibuloocular reflex (tVOR) and the angular vestibuloocular reflex (aVOR) is to stabilize images on the retina during translational and rotational motion, respectively. It has generally been assumed that these two reflexes differ in their central processing because they differ significantly in their primary afferent behavior and characteristics at the motor level. So far, models of the tVOR have focused on the type of processing that the primary afferent signal must undergo before reaching the neural integrator. Here, we propose a model that does not require any prefiltering. It is known that the eye plant requires signals in phase with velocity and position. We propose that the velocity signal is obtained directly from the neural integrator, whereas the position signal is obtained directly from the primary afferents synapsing onto the oculomotor nuclei. This design proved sufficient to simulate eye movements in response to translational motion. PMID- 10515994 TI - GABA(A) and GABA(C) receptors have contrasting effects on excitability in superior colliculus. AB - We have recently found that GABA(C) receptor subunit transcripts are expressed in the superficial layers of rat superior colliculus (SC). In the present study we used immunocytochemistry to demonstrate the presence of GABA(C) receptors in rat SC at protein level. We also investigated in acute rat brain slices the effect of GABA(A) and GABA(C) receptor agonists and antagonists on stimulus-evoked extracellular field potentials in SC. Electrical stimulation of the SC optic layer induced a biphasic, early and late, potential in the adjacent superficial layer. The late component was completely inhibited by 6-cyano-7-nitroquinoxaline 2,3-dione or CoCl(2), indicating that it was generated by postsynaptic activation. Muscimol, a potent GABA(A) and GABA(C) receptor agonist, strongly attenuated this postsynaptic potential at concentrations >10 microM. In contrast, the GABA(C) receptor agonist cis-aminocrotonic acid, as well as muscimol at lower concentrations (0.1-1 microM) increased the postsynaptic potential. This increase was blocked by (1,2,5, 6-tetrahydropyridine-4-yl)methylphosphinic acid, a novel competitive antagonist of GABA(C) receptors. Our findings demonstrate the presence of functional GABA(C) receptors in SC and suggest a disinhibitory role of these receptors in SC neuronal circuitry. PMID- 10515993 TI - Evidence that wakefulness and REM sleep are controlled by a GABAergic pontine mechanism. AB - The pontine microinjection of the inhibitory neurotransmitter GABA and its agonist induced prolonged periods of wakefulness in unanesthetized, chronic cats. Conversely, the application of bicuculline, a GABA(A) antagonist, resulted in the occurrence of episodes of rapid eye movement (REM) sleep of long duration. Furthermore, administration of antisense oligonucleotides against glutamic acid decarboxylase (GAD) mRNA into the same area produced a significant decrease in wakefulness and an increase in REM sleep. Microinjections of glycine, another major inhibitory neurotransmitter in the CNS, and its antagonist, strychnine, did not have any effect on the behavioral states of sleep and wakefulness. These data argue forcibly that 1) GABAergic neurons play a pivotal role in determining the occurrence of both wakefulness and REM sleep and 2) the functional sequelea of inhibitory GABA actions within the pontine reticular formation are excitatory directives and/or behaviors. PMID- 10515995 TI - Priming-induced shift in synaptic plasticity in the rat hippocampus. AB - The activity history of a given neuron has been suggested to influence its future responses to synaptic input in one prominent model of experience-dependent synaptic plasticity proposed by Bienenstock, Cooper, and Munro (BCM theory). Because plasticity of synaptic plasticity (i.e., metaplasticity) is similar in concept to aspects of the BCM proposal, we have tested the possibility that a form of metaplasticity induced by a priming stimulation protocol might exhibit BCM-like characteristics. CA1 field excitatory postsynaptic potentials (EPSPs) obtained from rat hippocampal slices were used to monitor synaptic responses before and after conditioning stimuli (3-100 Hz) of the Schaffer collateral inputs. A substantial rightward shift (>5-fold) in the frequency threshold between long-term depression (LTD) and long-term potentiation (LTP) was observed <1 h after priming. This change in the LTD/P crossover point occurred at both primed and unprimed synaptic pathways. These results provide new support for the existence of a rapid, heterosynaptic, experience-dependent mechanism that is capable of modifying the synaptic plasticity phenomena that are commonly proposed to be important for developmental and learning/memory processes in the brain. PMID- 10515996 TI - Characterization of intracellular reverse transcription complexes of Moloney murine leukemia virus. AB - To examine the early events in the life cycle of Moloney murine leukemia virus (MoMLV), we analyzed the intracellular complexes mediating reverse transcription. Partial purification of the reverse transcription complexes (RTCs) by equilibrium density fractionation and velocity sedimentation indicated that three distinct species of intracellular complexes are formed shortly after cell infection. Only one of these species is able to start and complete reverse transcription in the cell cytoplasm. This RTC is composed of at least the viral genome, capsid, integrase, and reverse transcriptase proteins. The RTC becomes permeable to micrococcal nuclease but not to antibodies. Shortly after initiation of reverse transcription, the viral strong stop DNA within the RTC is protected from nuclease digestion. The sedimentation velocity of the RTC decreases during reverse transcription. After entry into the nucleus, most capsid proteins are lost from the RTC and its sedimentation velocity decreases further. PMID- 10515998 TI - African swine fever virus dUTPase is a highly specific enzyme required for efficient replication in swine macrophages. AB - The African swine fever virus (ASFV) gene E165R, which is homologous to dUTPases, has been characterized. A multiple alignment of dUTPases showed the conservation in ASFV dUTPase of the motifs that define this protein family. A biochemical analysis of the purified recombinant enzyme showed that the virus dUTPase is a trimeric, highly specific enzyme that requires a divalent cation for activity. The enzyme is most probably complexed with Mg(2+), the preferred cation, and has an apparent K(m) for dUTP of 1 microM. Northern and Western blotting, as well as immunofluorescence analyses, indicated that the enzyme is expressed at early and late times of infection and is localized in the cytoplasm of the infected cells. On the other hand, an ASFV dUTPase-deletion mutant (vDeltaE165R) has been obtained. Growth kinetics showed that vDeltaE165R replicates as efficiently as parental virus in Vero cells but only to 10% or less of parental virus in swine macrophages. Our results suggest that the dUTPase activity is dispensable for virus replication in dividing cells but is required for productive infection in nondividing swine macrophages, the natural host cell for the virus. The viral dUTPase may play a role in lowering the dUTP concentration in natural infections to minimize misincorporation of deoxyuridine into the viral DNA and ensure the fidelity of genome replication. PMID- 10515997 TI - An Mpsi-containing heterologous RNA, but not env mRNA, is efficiently packaged into avian retroviral particles. AB - Retroviruses preferentially package full-length genomic RNA over spliced viral messages. For most retroviruses, this preference is likely due to the absence of all or part of the packaging signal on subgenomic RNAs. In avian leukosis-sarcoma virus, however, we have shown that the minimal packaging signal, MPsi, is located upstream of the 5' splice site and therefore is present on both genomic and spliced RNAs. We now show that an MPsi-containing heterologous RNA is packaged only 2.6-fold less efficiently than genomic Rous sarcoma virus RNA. Thus, few additional packaging sequences and/or structures exist outside of MPsi. In contrast, we found that env mRNA is not efficiently packaged. These results indicate that either MPsi is not functional on this RNA or the RNA is somehow segregated from the packaging machinery. Finally, deletion of sequences from the 3' end of MPsi was found to reduce the packaging efficiency of heterologous RNAs. PMID- 10515999 TI - Interleukin-4 diminishes CD8(+) respiratory syncytial virus-specific cytotoxic T lymphocyte activity in vivo. AB - Although interleukin-4 (IL-4) has been implicated in respiratory syncytial virus (RSV)-enhanced disease, the mechanism by which it modulates immune responses to primary RSV infection remains unclear. We have developed a system to investigate the effect of IL-4 on RSV epitope-specific cytotoxic T-lymphocyte (CTL) effector function in vivo, using an H-2K(d)-restricted RSV M2 epitope. BALB/c mice were infected with recombinant vaccinia virus (rVV) constructed to express RSV M2 protein (vvM2) alone or coexpress M2 and IL-4 (vvM2/IL-4). Splenocytes were assessed for M2-specific CTL activity in a direct (51)Cr release assay and intracellular gamma interferon (IFN-gamma) production by fluorescence-activated cell sorting analysis. Mice infected with vvM2/IL-4 had less M2-specific primary CTL activity than those infected with vvM2. M2-specific CTL frequency, as measured by M2 peptide-induced intracellular IFN-gamma production, was diminished in the vvM2/IL-4 group, partially accounting for the reduction of CTL activity. Mice immunized with either construct were challenged intravenously with RSV 4 weeks postimmunization, and direct CTL were measured. These results demonstrate that local expression of IL-4, at the time of antigen presentation, diminishes the cytolytic activity of primary and memory CD8(+) RSV-specific CTL responses in vivo. PMID- 10516000 TI - Herpes simplex virus inhibits apoptosis through the action of two genes, Us5 and Us3. AB - Apoptosis of virus-infected cells occurs either as a direct response to viral infection or upon recognition of infection by the host immune response. Apoptosis reduces production of new virus from these cells, and therefore viruses have evolved inhibitory mechanisms. We previously showed that laboratory strains of herpes simplex virus type 1 (HSV-1) protect infected cells from apoptosis induced by cytotoxic T lymphocytes or ethanol. We have now evaluated the ability of HSV-1 and HSV-2 laboratory and clinical isolates to inhibit apoptosis induced by anti Fas antibody or UV irradiation and explored the genetic basis for this inhibition. HSV-1 isolates inhibited apoptosis induced by UV or anti-Fas antibody. In contrast, HSV-2 clinical isolates failed to inhibit apoptosis induced by either stimulus, although the HSV-2 laboratory strain 333 had a partial inhibitory effect on UV-induced apoptosis. Inhibition of apoptosis by HSV was accompanied by marked reduction of caspase-3 and caspase-8 activity. Deletion of the HSV-1 Us3 gene markedly reduced inhibition of UV-induced apoptosis and partially abrogated inhibition of Fas-mediated apoptosis. Conversely, deletion of the HSV-1 Us5 gene markedly reduced protection from Fas-mediated apoptosis and partially abrogated protection from UV. The Us11 and Us12 genes were not necessary for protection from apoptosis induced by either stimulus. The differences between HSV-1 and HSV-2 in the ability to inhibit apoptosis may be factors in the immunobiology of HSV infections. PMID- 10516001 TI - Nonreplicative RNA recombination in poliovirus. AB - Current models of recombination between viral RNAs are based on replicative template-switch mechanisms. The existence of nonreplicative RNA recombination in poliovirus is demonstrated in the present study by the rescue of viable viruses after cotransfections with different pairs of genomic RNA fragments with suppressed translatable and replicating capacities. Approximately 100 distinct recombinant genomes have been identified. The majority of crossovers occurred between nonhomologous segments of the partners and might have resulted from transesterification reactions, not necessarily involving an enzymatic activity. Some of the crossover loci are clustered. The origin of some of these "hot spots" could be explained by invoking structures similar to known ribozymes. A significant proportion of recombinant RNAs contained the entire 5' partner, if its 3' end was oxidized or phosphorylated prior to being mixed with the 3' partner. All of these observations are consistent with a mechanism that involves intermediary formation of the 2',3'-cyclic phosphate and 5'-hydroxyl termini. It is proposed that nonreplicative RNA recombination may contribute to evolutionarily significant RNA rearrangements. PMID- 10516002 TI - A cell line-based neutralization assay for primary human immunodeficiency virus type 1 isolates that use either the CCR5 or the CXCR4 coreceptor. AB - We describe here a cell line-based assay for the evaluation of human immunodeficiency virus type 1 (HIV-1) neutralization. The assay is based on CEM.NKR cells, transfected to express the HIV-1 coreceptor CCR5 to supplement the endogenous expression of CD4 and the CXCR4 coreceptor. The resulting CEM.NKR-CCR5 cells efficiently replicate primary HIV-1 isolates of both R5 and X4 phenotypes. A comparison of the CEM.NKR-CCR5 cells with mitogen-activated peripheral blood mononuclear cells (PBMC) in neutralization assays with sera from HIV-1-infected individuals or specific anti-HIV-1 monoclonal antibodies shows that the sensitivity of HIV-1 neutralization is similar in the two cell types. The CEM.NKR CCR5 cell assay, however, is more convenient to perform and eliminates the donor to-donor variation in HIV-1 replication efficiency, which is one of the principal drawbacks of the PBMC-based neutralization assay. We suggest that this new assay is suitable for the general measurement of HIV-1 neutralization by antibodies. PMID- 10516003 TI - Palmitoylation of the intracytoplasmic R peptide of the transmembrane envelope protein in Moloney murine leukemia virus. AB - Previously it was reported that the 16-amino-acid (aa) C-terminal cytoplasmic tail of Moloney murine leukemia virus (MoMLV) transmembrane protein Pr15E is cleaved off during virus synthesis, yielding the mature, fusion active transmembrane protein p15E and the 16-aa peptide (R peptide or p2E). It remains to be elucidated how the R peptide impairs fusion activity of the uncleaved Pr15E. The R peptide from MoMLV was analyzed by Tricine-sodium dodecyl sulfate polyacrylamide gel electrophoresis and immunostained with antiserum against the synthetic 16-aa R peptide. The R peptide resolved with an apparent molecular mass of 7 kDa and not the 4 kDa seen with the corresponding synthetic peptide. The 7 kDa R peptide was found to be membrane bound in MoMLV-infected NIH 3T3 cells, showing that cleavage of the 7-kDa R-peptide tail must occur before or during budding of progeny virions, in which only small amounts of the 7-kDa R peptide were found. The 7-kDa R peptide was palmitoylated since it could be labeled with [(3)H]palmitic acid, which explains its membrane association, slower migration on gels, and high sensitivity in immunoblotting. The present results are in contrast to previous findings showing equimolar amounts of R peptide and p15E in virions. The discrepancy, however, can be explained by the presence of nonpalmitoylated R peptide in virions, which were poorly detected by immunoblotting. A mechanistic model is proposed. The uncleaved R peptide can, due to its lipid modification, control the conformation of the ectodomain of the transmembrane protein and thereby govern membrane fusion. PMID- 10516004 TI - A primary determinant of cap-independent translation is located in the 3' proximal region of the tomato bushy stunt virus genome. AB - Tomato bushy stunt virus (TBSV) is a positive-strand RNA virus and is the prototype member of the genus Tombusvirus. The genomes of members of this genus are not polyadenylated, and prevailing evidence supports the absence of a 5' cap structure. Previously, a 167-nucleotide-long segment (region 3.5) located near the 3' terminus of the TBSV genome was implicated as a determinant of translational efficiency (S.K. Oster, B. Wu and K. A. White, J. Virol. 72:5845 5851, 1998). In the present report, we provide evidence that a 3'-proximal segment of the genome, which includes region 3.5, is involved in facilitating cap independent translation. Our results indicate that (i) a 5' cap structure can substitute functionally for the absence of region 3.5 in viral and chimeric reporter mRNAs in vivo; (ii) deletion of region 3.5 from viral and chimeric mRNAs has no appreciable effect on message stability; (iii) region 3.5 represents part of a larger 3' proximal element, designated as the 3' cap-independent translational enhancer (3'CITE), that is required for proficient cap-independent translation; (iv) the 3'CITE also facilitates cap-dependent translation; (v) none of the major viral proteins are required for 3'CITE activity; and (vi) no significant 3'CITE-dependent stimulation of translation was observed when mRNAs were tested in vitro in wheat germ extract under various assay conditions. This latter property distinguishes the 3'CITE from other characterized plant viral 3' proximal cap-independent translational enhancers. Additionally, because the 3'CITE overlaps with cis-acting replication signals, it could potentially participate in regulating the initiation of genome replication. PMID- 10516005 TI - Adeno-associated virus type 2 protein interactions: formation of pre encapsidation complexes. AB - The nonstructural adeno-associated virus type 2 Rep proteins are known to control viral replication and thus provide the single-stranded DNA genomes required for packaging into preformed capsids. In addition, complexes between Rep proteins and capsids have previously been observed in the course of productive infections. Such complexes have been interpreted as genome-linked Rep molecules associated with the capsid upon successful DNA encapsidation. Here we demonstrate via coimmunoprecipitation, cosedimentation, and yeast two-hybrid analyses that the Rep-VP association also occurs in the absence of packageable genomes, suggesting that such complexes could be involved in the preparation of empty capsids for subsequent encapsidation steps. The Rep domain responsible for the observed Rep VP interactions is situated within amino acids 322 to 482. In the presence of all Rep proteins, Rep52 and, to a lesser extent, Rep78 are most abundantly recovered with capsids, whereas Rep68 and Rep40 vary in association depending on their expression levels. Rep78 and Rep52 are bound to capsids to roughly the same extent as the minor capsid protein VP2. Complexes of Rep78 and Rep52 with capsids differ in their respective detergent stabilities, indicating that they result from different types of interactions. Rep-VP interaction studies suggest that Rep proteins become stably associated with the capsid during the assembly process. Rep-capsid complexes can reach even higher complexity through additional Rep-Rep interactions, which are particularly detergent labile. Coimmunoprecipitation and yeast two-hybrid data demonstrate the interaction of Rep78 with Rep68, of Rep68 with Rep52, and weak interactions of Rep40 with Rep52 and Rep78. We propose that the large complexes arising from these interactions represent intermediates in the DNA packaging pathway. PMID- 10516006 TI - SPI-1-dependent host range of rabbitpox virus and complex formation with cathepsin G is associated with serpin motifs. AB - Serpins are a superfamily of serine proteinase inhibitors which function to regulate a number of key biological processes including fibrinolysis, inflammation, and cell migration. Poxviruses are the only viruses known to encode functional serpins. While some poxvirus serpins regulate inflammation (myxoma virus SERP1 and cowpox virus [CPV] crmA/SPI-2) or apoptosis (myxoma virus SERP2 and CPV crmA/SPI-2), the function of other poxvirus serpins remains unknown. The rabbitpox virus (RPV) SPI-1 protein is 47% identical to crmA and shares all of the serpin structural motifs. However, no serpin-like activity has been demonstrated for SPI-1 to date. Earlier we showed that RPV with the SPI-1 gene deleted, unlike wild-type virus, fails to grow on A549 or PK15 cells (A. Ali, P. C. Turner, M. A. Brooks, and R. W. Moyer, Virology 202:306-314, 1994). Here we demonstrate that in the absence of a functional SPI-1 protein, infected nonpermissive cells which exhibit the morphological features of apoptosis fail to activate terminal caspases or cleave the death substrates PARP or lamin A. We show that SPI-1 forms a stable complex in vitro with cathepsin G, a member of the chymotrypsin family of serine proteinases, consistent with serpin activity. SPI-1 reactive-site loop (RSL) mutations of the critical P1 and P14 residues abolish this activity. Viruses containing the SPI-1 RSL P1 or P14 mutations also fail to grow on A549 or PK15 cells. These results suggest that the full virus host range depends on the serpin activity of SPI-1 and that in restrictive cells SPI-1 inhibits a proteinase with chymotrypsin-like activity and may function to inhibit a caspase-independent pathway of apoptosis. PMID- 10516007 TI - Structure-based mutagenesis of the human immunodeficiency virus type 1 DNA attachment site: effects on integration and cDNA synthesis. AB - Sequences at the ends of linear retroviral cDNA important for integration define the viral DNA attachment (att) site. Whereas determinants of human immunodeficiency virus type 1 (HIV-1) integrase important for replication in T lymphocytes have been extensively characterized, regions of the att site important for viral spread have not been thoroughly examined. Previous transposon mediated footprinting of preintegration complexes isolated from infected cells revealed enhanced regions of bacteriophage Mu insertion near the ends of HIV-1 cDNA, in the regions of the att sites. Here, we identified the subterminal cDNA sequences cleaved during in vitro footprinting and used this structure-based information together with results of previous work to construct and characterize 24 att site mutant viruses. We found that although subterminal cDNA sequences contributed to HIV-1 replication, the identities of these bases were not critical for integration. In contrast, the phylogenetically conserved CA dinucleotides located at the ends of HIV-1 contributed significantly to virus replication and integration. Mutants containing one intact CA end displayed delays in peak virus growth compared to the wild type. In contrast, double mutant viruses lacking both CAs were replication defective. The A of the CA appeared to be the most critical determinant of integration, because two different U5 mutant viruses containing the substitution of TG for CA partially reverted by changing the G back to A. We also identified a U5 deletion mutant in which the CA played a crucial role in reverse transcription. PMID- 10516008 TI - Inhibition of replication of reactivated human immunodeficiency virus type 1 (HIV 1) in latently infected U1 cells transduced with an HIV-1 long terminal repeat driven PKR cDNA construct. AB - Treatment of human immunodeficiency virus type 1 (HIV-1)-infected individuals with highly active antiretroviral therapy has effectively decreased viral load to undetectable levels. However, efforts to eliminate HIV-1 from these individuals have been unsuccessful, due to the presence of stable, latent viral reservoirs in resting and active CD4(+) T lymphocytes and macrophages. These latent populations have become critical targets in the effort to eradicate HIV-1 from infected individuals. The mechanisms of HIV-1 latency have been studied by using the HIV-1 infected promonocytic cell line U1. The interferon-inducible double-stranded RNA dependent p68 protein kinase (PKR), a key enzyme in the host-mediated antiviral response, is known to be down-regulated during HIV-1 infection. Therefore, in order to evaluate the role of PKR in the inhibition of replication of reactivated HIV-1 in latently infected U1 cells, we have utilized cDNA constructs containing PKR under the transcriptional control of the HIV-1 long terminal repeat. One PKR transduced clone, U1/106-4:27, inhibited the tumor necrosis factor alpha (TNF alpha)-induced replication of HIV-1 by 99% compared to control U1 cells as measured by syncytium formation and HIV-1 p24 antigen enzyme-linked immunosorbent assay. Western blot analysis showed an increase in PKR expression through 96 h postinduction in the U1/106-4:27 clone, concomitant with maximal increases in phosphorylation of the alpha subunit of eukaryotic initiation factor 2 and NF kappaB activity at 72 h postinduction. These results demonstrate that overexpression of PKR can inhibit the replication of reactivated HIV-1 in latently infected cells and confirm the involvement of PKR in the interferon associated antiviral pathway against HIV-1 infection. Additionally, treatment of the PKR-transduced U1/106-4:27 clone with the protease inhibitor saquinavir (250 nM) completely inhibited TNF-alpha-induced HIV-1 replication. PMID- 10516009 TI - Genetic dissection of cell growth arrest functions mediated by the Epstein-Barr virus lytic gene product, Zta. AB - Expression of the Epstein-Barr virus (EBV) latency-associated genes activates cell cycle progression and drives immortalization of the infected cell. In contrast, progression of the EBV replication program occurs most efficiently in growth-arrested cells. Previous studies showed that the EBV-encoded immediate early transcription factor, Zta, can induce expression of the cyclin-dependent kinase inhibitors, p21 and p27, the tumor suppressor, p53, and cell growth arrest. Moreover, Zta-mediated induction of growth arrest occurs independently of its transcriptional transactivation function. Here we show that substitution of Zta's basic DNA binding domain with the analogous region of the Zta homologue, c Fos, abrogates Zta's ability to induce growth arrest and to induce p21, p27, or p53 expression, suggesting that protein-protein interactions between this region of Zta and key cell cycle control proteins are involved in signaling cell cycle arrest. We also show that despite the crucial role for Zta's basic domain in eliciting cell growth arrest, its amino terminus is required for efficient induction of p27 and it modulates the level of p53 induction. Last, we provide evidence that Zta-mediated inductions of p21, p27, and p53 occur, at least in part, through distinct pathways. Therefore, Zta interacts with multiple growth arrest pathways, a property which may have evolved partly as a means to ensure that lytic replication occurs in a growth-arrested setting in multiple different tissues in various states of differentiation. PMID- 10516010 TI - A strong negative transcriptional regulatory region between the human cytomegalovirus UL127 gene and the major immediate-early enhancer. AB - The region of the human cytomegalovirus (CMV) genome between the UL127 open reading frame and the major immediate-early (MIE) enhancer is referred to as the unique region. DNase I protection analysis with human cell nuclear extracts demonstrated multiple protein binding sites in this region of the viral genome (P. Ghazal, H. Lubon, C. Reynolds-Kohler, L. Hennighausen, and J. A. Nelson, Virology 174:18-25, 1990). However, the function of this region in the context of the viral genome is not known. In wild-type human CMV-infected human fibroblasts, cells permissive for viral replication, there is little to no transcription from UL127. We determined that the unique region prevented transcription from the UL127 promoter but had no effect on the divergent MIE promoter. In transient transfection assays, the basal level of expression from the UL127 promoter increased significantly when the wild-type unique sequences were mutated. In recombinant viruses with similar mutations in the unique region, expression from the UL127 promoter occurred only after de novo viral protein synthesis, typical of an early viral promoter. A 111-bp deletion-substitution of the unique sequence caused approximately a 20-fold increase in the steady-state level of RNA from the UL127 promoter and a 245-fold increase in the expression of a downstream indicator gene. This viral negative regulatory region was also mutated at approximately 50-bp regions proximal and distal to the UL127 promoter. Although some repressive effects were detected in the distal region, mutations of the region proximal to the UL127 promoter had the most significant effects on transcription. Within the proximal and distal regions, there are potential cis sites for known eucaryotic transcriptional repressor proteins. This region may also bind unknown viral proteins. We propose that the unique region upstream of the UL127 promoter and the MIE enhancer negatively regulates the expression from the UL127 promoter in permissive human fibroblast cells. This region may be a regulatory boundary preventing the effects of the very strong MIE enhancer on this promoter. PMID- 10516011 TI - Phosphorylation of simian cytomegalovirus assembly protein precursor (pAPNG.5) and proteinase precursor (pAPNG1): multiple attachment sites identified, including two adjacent serines in a casein kinase II consensus sequence. AB - The assembly protein precursor (pAP) of cytomegalovirus (CMV), and its homologs in other herpesviruses, functions at several key steps during the process of capsid formation. This protein, and the genetically related maturational proteinase, is distinguished from the other capsid proteins by posttranslational modifications, including phosphorylation. The objective of this study was to identify sites at which pAP is phosphorylated so that the functional significance of this modification and the enzyme(s) responsible for it can be determined. In the work reported here, we used peptide mapping, mass spectrometry, and site directed mutagenesis to identify two sets of pAP phosphorylation sites. One is a casein kinase II (CKII) consensus sequence that contains two adjacent serines, both of which are phosphorylated. The other site(s) is in a different domain of the protein, is phosphorylated less frequently than the CKII site, does not require preceding CKII-site phosphorylation, and causes an electrophoretic mobility shift when phosphorylated. Transfection/expression assays for proteolytic activity showed no gross effect of CKII-site phosphorylation on the enzymatic activity of the proteinase or on the substrate behavior of pAP. Evidence is presented that both the CKII sites and the secondary sites are phosphorylated in virus-infected cells and plasmid-transfected cells, indicating that these modifications can be made by a cellular enzyme(s). Apparent compartmental differences in phosphorylation of the CKII-site (cytoplasmic) and secondary-site (nuclear) serines suggest the involvement of more that one enzyme in these modifications. PMID- 10516013 TI - Poliovirus mutants at histidine 195 of VP2 do not cleave VP0 into VP2 and VP4. AB - The final stage of poliovirus assembly is characterized by a cleavage of the capsid precursor protein VP0 into VP2 and VP4. This cleavage is thought to be autocatalytic and dependent on RNA encapsidation. Analysis of the poliovirus empty capsid structure has led to a mechanistic model for VP0 cleavage involving a conserved histidine residue that is present in the surrounding environment of the VP0 cleavage site. Histidine 195 of VP2 (2195H) is hypothesized to activate local water molecules, thus initiating a nucleophilic attack at the scissile bond. To test this hypothesis, 2195H mutants were constructed and their phenotypes were characterized. Consistent with the requirement of VP0 cleavage for poliovirus infectivity, all 2195H mutants were nonviable upon introduction of the mutant genomes into HeLa cells. Replacement of 2195H with threonine or arginine resulted in the assembly of a highly unstable 150S virus particle. Further analyses showed that these particles contain genomic RNA and uncleaved VP0, criteria associated with the provirion assembly intermediate. These data support the involvement of 2195H in mediating VP0 cleavage during the final stages of virus assembly. PMID- 10516012 TI - Nasal immunization of mice with human papillomavirus type 16 (HPV-16) virus-like particles or with the HPV-16 L1 gene elicits specific cytotoxic T lymphocytes in vaginal draining lymph nodes. AB - Human papillomavirus type 16 (HPV-16) infects the genital tract and is closely associated with the development of cervical cancer. HPV-16 initiates infection at the genital mucosal surface; thus, mucosal immune responses are likely to contribute to defense against HPV-16 infection. However, little information is available regarding the induction of immune responses in the genital tract mucosa. In this study, we evaluated the potential of intranasally administered papillomavirus vaccines to elicit both systemic and vaginal immune responses. HPV 16 virus-like particles (VLPs) produced by self-assembly of L1 protein and the HPV-16 L1 gene cloned into a mammalian expression vector were used as vaccines. Intranasally administered VLPs induced serum immunoglobulin G (IgG) and vaginal IgA secretory antibodies. Very weak serum IgG and vaginal IgA responses were found after DNA immunization. Both splenic and vaginal lymphocytes could be activated by intranasal immunization with VLPs and the HPV-16 L1 gene. Activated CD4(+) Th1-like T cells were shown to synthesize gamma interferon, and activated CD8(+) T cells were demonstrated to be cytotoxic. PMID- 10516014 TI - Identification and characterization of the functional elements within the tobacco etch virus 5' leader required for cap-independent translation. AB - Translation in plants is highly cap dependent, and the only plant mRNAs known to naturally lack a cap structure (m(7)GpppN) are viral in origin. The genomic RNA of tobacco etch virus (TEV), a potyvirus that belongs to the picornavirus superfamily, is a polyadenylated mRNA that is naturally uncapped and yet is a highly competitive mRNA during translation. The 143-nucleotide 5' leader is responsible for conferring cap-independent translation even on reporter mRNAs. We have carried out a deletion analysis of the TEV 5' leader to identify the elements responsible for its regulatory function and have identified two centrally located cap-independent regulatory elements (CIREs) that promote cap independent translation. The introduction of a stable stem-loop structure upstream of each element demonstrated that CIRE-1 is less 5' end dependent in function than CIRE-2. In a dicistronic mRNA, the presence of the TEV 5' leader sequence in the intercistronic region increased expression of the second cistron, suggesting that the viral sequence can function in a 5'-distal position. Interestingly, the introduction of a stable stem-loop upstream of the TEV leader sequence or upstream of either CIRE in dicistronic constructs markedly increased their regulatory function. These data suggest that the TEV 5' leader contains two elements that together promote internal initiation but that the function of one element, in particular, is facilitated by proximity to the 5' end. PMID- 10516015 TI - Human immunodeficiency virus type 1-induced hematopoietic inhibition is independent of productive infection of progenitor cells in vivo. AB - Human immunodeficiency virus (HIV)-infected individuals exhibit a variety of hematopoietic dysfunctions. The SCID-hu mouse (severe combined immunodeficient mouse transplanted with human fetal thymus and liver tissues) can be used to model the loss of human hematopoietic precursor cell function following HIV infection and has a distinct advantage in that data can be obtained in the absence of confounding factors often seen in infected humans. In this study, we establish that HIV type 1 (HIV-1) bearing a reporter gene inserted into the viral vpr gene is highly aggressive in depleting human myeloid and erythroid colony forming precursor activity in vivo. Human CD34(+) progenitor cells can be efficiently recovered from infected implants yet do not express the viral reporter gene, despite severe functional defects. Our results indicate that HIV-1 infection alone leads to hematopoietic inhibition in vivo; however, this effect is due to indirect mechanisms rather than to direct infection of CD34(+) cells in vivo. PMID- 10516016 TI - Identification of functional domains in the 14-kilodalton envelope protein (A27L) of vaccinia virus. AB - The mechanism of entry of vaccinia virus (VV) into cells is still a poorly understood process. A 14-kDa protein (encoded by the A27L gene) in the envelope of intracellular mature virus (IMV) has been implicated in virus-cell attachment, virus-cell fusion, and virus release from cells. We have previously described the structural organization of the VV 14-kDa protein, consisting of a triple-stranded coiled-coil region responsible for oligomer formation and a predicted Leu zipper like third alpha helix with an important role in the interaction with a 21-kDa membrane protein (encoded by the A17L gene) thought to anchor the 14-kDa protein to the envelope of IMV (M.-I. Vazquez, G. Rivas, D. Cregut, L. Serrano, and M. Esteban, J. Virol. 72:10126-10137, 1998). To identify the functional domains important for virus entry and release, we have generated VV recombinants containing a copy of the A27L gene regulated by the lacI operator-repressor system of Escherichia coli (VVIndA27L) in the thymidine kinase locus and a mutant form of the A27L gene in the hemagglutinin locus but expressed constitutively under the control of an early-late VV promoter. Cells infected with a VV recombinant that expresses a mutant 14-kDa form lacking the first 29 amino acids at the N terminus failed to form extracellular enveloped virus (EEV). Fusion-from without assays with purified virus confirmed that the fusion process was mediated by the 14-kDa protein and the fusion domain to be contained within amino acids 29 to 43 of the N-terminal region. Competitive inhibition of the infection process with soluble heparin and synthetic peptides and in vitro experiments with purified mutant proteins identified the heparin binding domain within amino acids 21 to 33, suggesting that this domain is involved in virus-cell binding via heparan sulfate. Thus, the N terminus of the 14-kDa protein contains a heparin binding domain, a fusion domain, and a domain responsible for interacting with proteins or lipids in the Golgi stacks for EEV formation and virus spread. PMID- 10516017 TI - Polypyrimidine tract-binding protein binds to the complementary strand of the mouse hepatitis virus 3' untranslated region, thereby altering RNA conformation. AB - Mouse hepatitis virus (MHV) RNA transcription is regulated mainly by the leader and intergenic (IG) sequences. However, a previous study has shown that the 3' untranslated region (3'-UTR) of the viral genome is also required for subgenomic mRNA transcription; deletion of nucleotides (nt) 270 to 305 from the 3'-UTR completely abolished subgenomic mRNA transcription without affecting minus-strand RNA synthesis (Y.-J. Lin, X. Zhang, R.-C. Wu, and M. M. C. Lai, J. Virol. 70:7236 7240, 1996), suggesting that the 3'-UTR affects positive-strand RNA synthesis. In this study, by UV-cross-linking experiments, we found that several cellular proteins bind specifically to the minus-strand 350 nucleotides complementary to the 3'-UTR of the viral genome. The major protein species, p55, was identified as the polypyrimidine tract-binding protein (PTB, also known as heterogeneous nuclear RNP I) by immunoprecipitation of the UV-cross-linked protein and binding of the recombinant PTB. A strong PTB-binding site was mapped to nt 53 to 149, and another weak binding site was mapped to nt 270 to 307 on the complementary strand of the 3'-UTR (c3'-UTR). Partial substitutions of the PTB-binding nucleotides reduced PTB binding in vitro. Furthermore, defective interfering (DI) RNAs harboring these mutations showed a substantially reduced ability to synthesize subgenomic mRNA. By enzymatic and chemical probing, we found that PTB binding to nt 53 to 149 caused a conformational change in the neighboring RNA region. Partial deletions within the PTB-binding sequence completely abolished the PTB induced conformational change in the mutant RNA even when the RNA retained partial PTB-binding activity. Correspondingly, the MHV DI RNAs containing these deletions completely lost their ability to transcribe mRNAs. Thus, the conformational change in the c3'-UTR caused by PTB binding may play a role in mRNA transcription. PMID- 10516018 TI - Site-directed mutagenesis of the virion host shutoff gene (UL41) of herpes simplex virus (HSV): analysis of functional differences between HSV type 1 (HSV 1) and HSV-2 alleles. AB - During lytic herpes simplex virus (HSV) infections, the HSV virion host shutoff protein (UL41) accelerates the turnover of host and viral mRNAs. Although the UL41 polypeptides from HSV type 1 (HSV-1) strain KOS and HSV-2 strain 333 are 87% identical, HSV-2 strains generally shut off the host more rapidly and completely than HSV-1 strains. In a previous study, we identified three regions of the HSV-2 UL41 polypeptide (amino acids 1 to 135, 208 to 243, and 365 to 492) that enhance the activity of KOS when substituted for the corresponding portions of the KOS protein (D. N. Everly, Jr., and G. S. Read, J. Virol. 71:7157-7166, 1997). These results have been extended through the analysis of more than 50 site-directed mutants of UL41 in which selected HSV-2 amino acids were introduced into an HSV-1 background and HSV-1 amino acids were introduced into the HSV-2 allele. The HSV-2 amino acids R22 and E25 were found to contribute dramatically to the greater activity of the HSV-2 allele, as did the HSV-2 amino acids A396 and S423. The substitution of six HSV-2 amino acids between residues 210 and 242 enhanced the HSV-1 activity to a lesser extent. In most cases, individual substitutions or the substitution of combinations of fewer than all six amino acids reduced the UL41 activity to less than that of KOS. The results pinpoint several type-specific amino acids that are largely responsible for the greater activity of the UL41 polypeptide of HSV-2. In addition, several spontaneous mutations that abolish detectable UL41 activity were identified. PMID- 10516019 TI - Construction of a pseudoreceptor that mediates transduction by adenoviruses expressing a ligand in fiber or penton base. AB - Modification of adenovirus to achieve tissue specific targeting for the delivery of therapeutic genes requires both the ablation of its native tropism and the introduction of specific, novel interactions. Inactivation of the native receptor interactions, however, would cripple the virus for growth in production cells. We have developed an alternative receptor, or pseudoreceptor, for the virus which might allow propagation of viruses with modified fiber proteins that no longer bind to the native adenovirus receptor (coxsackievirus/adenovirus receptor [CAR]). We have constructed a membrane-anchored single-chain antibody [m scFv(HA)] which recognizes a linear peptide epitope (hemagglutinin [HA]). Incorporation of HA within the HI loop of the fiber protein enabled the modified virus to transduce pseudoreceptor expressing cells under conditions where fiber CAR interaction was blocked or absent. The pseudoreceptor mediated virus transduction with an efficiency similar to that of CAR. In addition, the HA epitope mediated virus transduction through interaction with the m-scFv(HA) when it was introduced into penton base. These findings indicate that cells expressing the pseudoreceptor should support production of HA-tagged adenoviruses independent of retaining the fiber-CAR interaction. Moreover, they demonstrate that high-affinity targeting ligands may function following insertion into either penton base or fiber. PMID- 10516021 TI - Evaluation of novel human immunodeficiency virus type 1 Gag DNA vaccines for protein expression in mammalian cells and induction of immune responses. AB - Human immunodeficiency virus (HIV)-specific cytotoxic T lymphocytes (CTL) are an important parameter of host defenses that limit viral replication after infection. Induction of effective CTL against conserved viral proteins such as Gag may be essential to the development of a safe and effective HIV type 1 (HIV 1) vaccine. DNA vaccination represents a novel strategy for inducing potent CD8(+) CTL responses in vivo. However, expression of HIV-1 structural proteins by DNA vectors has been hampered by a stringent requirement for coexpression with other viral components, such as Rev and RRE. Furthermore, even with Rev and RRE present, the level of expression of HIV-1 Gag, Pol, or Env is very low in murine cells. These problems have limited our ability to address the key issue of how to generate effective CTL responses to Gag in a mouse model. To overcome this problem, we compared several novel DNA expression vectors for HIV-1 Gag protein expression in primate and mouse cells and for generating immune responses in mice after DNA vaccination. A DNA vector containing wild type HIV-1 gag coding sequences did not induce detectable Gag expression in any of the cells tested. Attempts to increase nuclear export of Gag expression RNA by adding the constitutive transport element yielded only a moderate increase in Gag expression in monkey-derived COS cells and an even lower increase in Gag expression in HeLa cells or several mouse cell lines. In contrast, silent-site mutations in the HIV 1 gag coding sequences significantly increased Gag expression levels in all cells tested. Furthermore, this construct induced both Gag-specific antibody and CTL responses in mice after DNA vaccination. Using this construct, we achieved stable expression of HIV-1 Gag in the mouse cell line p815, which can now be used as a target cell for measuring HIV-1 Gag-specific CTL responses in immunized mice. The DNA vectors described in this study should make it possible to systematically evaluate the approaches for maximizing the induction of CTL responses against HIV 1 Gag in mouse and other animal systems. PMID- 10516020 TI - Replication-defective bovine adenovirus type 3 as an expression vector. AB - Although recombinant human adenovirus (HAV)-based vectors offer several advantages for somatic gene therapy and vaccination over other viral vectors, it would be desirable to develop alternative vectors with prolonged expression and decreased toxicity. Toward this objective, a replication-defective bovine adenovirus type 3 (BAV-3) was developed as an expression vector. Bovine cell lines designated VIDO R2 (HAV-5 E1A/B-transformed fetal bovine retina cell [FBRC] line) and 6.93.9 (Madin-Darby bovine kidney [MDBK] cell line expressing E1 proteins) were developed and found to complement the E1A deletion in BAV-3. Replication-defective BAV-3 with a 1.7-kb deletion removing most of the E1A and E3 regions was constructed. This virus could be grown in VIDO R2 or 6.93.9 cells but not in FBRC or MDBK cells. The results demonstrated that the E1 region of HAV 5 has the capacity to transform bovine retina cells and that the E1A region of HAV-5 can complement that of BAV-3. A replication-defective BAV-3 vector expressing bovine herpesvirus type 1 glycoprotein D from the E1A region was made. A similar replication-defective vector expressing the hemagglutinin-esterase gene of bovine coronavirus from the E3 region was isolated. Although these viruses grew less efficiently than the replication-competent recombinant BAV-3 (E3 deleted), they are suitable for detailed studies with animals to evaluate the safety, duration of foreign gene expression, and ability to induce immune responses. In addition, a replication-competent recombinant BAV-3 expressing green fluorescent protein was constructed and used to evaluate the host range of BAV-3 under cell culture conditions. The development of bovine E1A-complementing cell lines and the generation of replication-defective BAV-3 vectors is a major technical advancement for defining the use of BAV-3 as vector for vaccination against diseases of cattle and somatic gene therapy in humans. PMID- 10516022 TI - Longitudinal phenotypic analysis of human immunodeficiency virus type 1-specific cytotoxic T lymphocytes: correlation with disease progression. AB - Few studies have examined longitudinal changes in human immunodeficiency virus type 1 (HIV)-specific cytotoxic T lymphocytes (CTL). To more closely define the natural history of HIV-specific CTL, we used HLA-peptide tetrameric complexes to study the longitudinal CD8(+) T-cell response evolution in 16 A*0201-positive untreated individuals followed clinically for up to 14 years. As early as 1 to 2 years after seroconversion, we found a significant association between high frequencies of A*0201-restricted p17(Gag/Pol) tetramer-binding cells and slower disease progression (P < 0.01). We observed that responses could remain stable over many months, but any longitudinal changes that occurred were typically accompanied by reciprocal changes in RNA viral load. Phenotypic analysis with markers CD45RO, CD45RA, and CD27 identified distinct subsets of antigen-specific cells and the preferential loss of CD27(+) CD45RO(+) cells during periods of rapid decline in the frequency of tetramer-binding cells. In addition we were unable to confirm previous studies showing a consistent selective loss of HIV specific cells in the context of sustained Epstein-Barr virus-specific cell frequencies. Overall, these data support a role of HIV-specific CTL in the control of disease progression and suggest that the ultimate loss of such CTL may be preferentially from the CD27(+) CD45RO(+) subset. PMID- 10516023 TI - Hierarchal utilization of different T-cell receptor Vbeta gene segments in the CD8(+)-T-cell response to an immunodominant Moloney leukemia virus-encoded epitope in vivo. AB - The CD8(+)-T-cell response to Moloney murine leukemia virus (M-MuLV)-associated antigens in C57BL/6 mice is directed against an immunodominant gag-encoded epitope (CCLCLTVFL) presented in the context of H-2D(b) and is restricted primarily to cytotoxic T lymphocytes (CTL) expressing the Valpha3.2 and Vbeta5.2 gene segments. We decided to examine the M-MuLV response in congenic C57BL/6 Vbeta(a) mice which are unable to express the dominant Valpha3.2(+) Vbeta5.2(+) T cell receptor (TCR) due to a large deletion at the TCR locus that includes the Vbeta5.2 gene segment. Interestingly, M-MuLV-immune C57BL/6 Vbeta(a) mice were still able to reject M-MuLV-infected tumor cells and direct ex vivo analysis of peripheral blood lymphocytes from these immune mice revealed a dramatic increase in CD8(+) cells utilizing the same Valpha3.2 gene segment in association with two different Vbeta segments (Vbeta3 and Vbeta17). Surprisingly, all these CTL recognized the same immunodominant M-MuLV gag epitope. Analysis of the TCR repertoire of individual M-MuLV-immune (C57BL/6 x C57BL/6 Vbeta(a))F(1) mice revealed a clear hierarchy in Vbeta utilization, with a preferential usage of the Vbeta17 gene segment, whereas Vbeta3 and especially Vbeta5.2 were used to much lesser extents. Sequencing of TCRalpha- and -beta-chain junctional regions of CTL clones specific for the M-MuLV gag epitope revealed a diverse repertoire of TCRbeta chains in Vbeta(a) mice and a highly restricted TCRbeta-chain repertoire in Vbeta(b) mice, whereas TCRalpha-chain sequences were highly conserved in both cases. Collectively, our data indicate that the H-2D(b)-restricted M-MuLV gag epitope can be recognized in a hierarchal fashion by different Vbeta domains and that the degree of beta-chain diversity varies according to Vbeta utilization. PMID- 10516024 TI - The nucleocapsid domain is responsible for the ability of spleen necrosis virus (SNV) Gag polyprotein to package both SNV and murine leukemia virus RNA. AB - Murine leukemia virus (MLV)-based vector RNA can be packaged and propagated by the proteins of spleen necrosis virus (SNV). We recently demonstrated that MLV proteins cannot support the replication of an SNV-based vector; RNA analysis revealed that MLV proteins cannot efficiently package SNV-based vector RNA. The domain in Gag responsible for the specificity of RNA packaging was identified using chimeric gag-pol expression constructs. A competitive packaging system was established by generating a cell line that expresses one viral vector RNA containing the MLV packaging signal (Psi) and another viral vector RNA containing the SNV packaging signal (E). The chimeric gag-pol expression constructs were introduced into the cells, and vector titers as well as the efficiency of RNA packaging were examined. Our data confirm that Gag is solely responsible for the selection of viral RNAs. Furthermore, the nucleocapsid (NC) domain in the SNV Gag is responsible for its ability to interact with both SNV E and MLV Psi. Replacement of the SNV NC with the MLV NC generated a chimeric Gag that could not package SNV RNA but retained its ability to package MLV RNA. A construct expressing SNV gag-MLV pol supported the replication of both MLV and SNV vectors, indicating that the gag and pol gene products from two different viruses can functionally cooperate to perform one cycle of retroviral replication. Viral titer data indicated that SNV cis-acting elements are not ideal substrates for MLV pol gene products since infectious viruses were generated at a lower efficiency. These results indicate that the nonreciprocal recognition between SNV and MLV extends beyond the Gag-RNA interaction and also includes interactions between Pol and other cis-acting elements. PMID- 10516026 TI - Genome-wide screening, cloning, chromosomal assignment, and expression of full length human endogenous retrovirus type K. AB - The human genome harbors 25 to 50 proviral copies of the endogenous retrovirus type K (HERV-K), some of which code for the characteristic retroviral proteins Gag, Pol, and Env. For a genome-wide cloning approach of full-length and intact HERV-K proviruses, a human P1 gene library was screened with a gag-specific probe. Both HERV-K type 1 and 2 clones were isolated. Sixteen HERV-K type 2 proviral genomes were characterized by direct coupled in vitro transcription-in vitro translation assays to analyze the coding potential of isolated gag, pol, and env amplicons from individual P1 clones. After determination of long terminal repeat (LTR) sequences and adjacent chromosomal integration sites by inverse PCR techniques, two HERV-K type 2 proviruses displaying long retroviral open reading frames (ORFs) were assigned to chromosomes 7 (C7) and 19 (C19) by using a human rodent monochromosomal cell hybrid mapping panel. HERV-K(C7) shows an altered (YIDD-to-CIDD) motif in the reverse transcriptase domain. HERV-K(C19) is truncated in the 5' LTR and harbors a defective protease gene due to a point mutation. Direct amplification of proviral structures from single chromosomes by using chromosomal flanking primers was performed by long PCR for HERV-K(C7) and HERV-K(C19) and for type 1 proviruses HERV-K10 and HERV-K18 from chromosomes 5 and 1, respectively. HERV-K18, in contrast to HERV-K10, bears no intact gag ORF and shows close homology to HERV-K/IDDMK(1,2)22. In transfection experiments, HERV-K(C7) and HERV-K cDNA-based expression vectors yielded the proteins Gag and cORF whereas HERV-K10 vectors yielded Gag alone. The data suggest that the human genome does not contain an entire, intact proviral copy of HERV-K. PMID- 10516025 TI - Sequence and insertion sites of murine melanoma-associated retrovirus. AB - We previously showed that B16 melanoma cells produce ecotropic melanoma associated retrovirus (MelARV) which encodes a melanoma-associated antigen recognized by MM2-9B6 monoclonal antibody. The biological significance of MelARV in melanoma formation remains unknown. We found that infection of normal melanocytes with MelARV resulted in malignant transformation. It is likely that MelARV emerged from the defective Emv-2 provirus, a single copy of ecotropic provirus existing in the genome of C57BL/6 mice. In the present study, we cloned and sequenced the full-length MelARV genome and its insertion sites and we completed sequencing of the Emv-2 provirus. Our data show that MelARV has a typical full-length retroviral genome with high homology (98.54%) to Emv-2, indicating a close relationship between both viruses. MelARV probably emerged as a result of recombination between Emv-2 and an endogenous nonecotropic provirus. Some observed differences in the gag and pol regions of MelARV might account for the restoration of productivity and infectivity of a novel retrovirus that somatically emerged during melanoma formation. MelARV does not contain any oncogene and therefore might induce transformation by insertional mutagenesis. We sequenced two insertion sites of MelARV. The first insertion site represents the 3' coding region of the c-maf proto-oncogene at 67.0 centimorgans (cM) on chromosome 8. The c-maf proto-oncogene encodes a basic leucine zipper protein homologous to c-fos and c-jun. Insertion of MelARV in BL6 melanoma cells resulted in the up-regulation of c-maf. It is noteworthy that the Emv-2 provirus is also inserted into a noncoding region at 61.0 cM on the same chromosome 8. The second insertion site is the 3' noncoding region of the DNA polymerase gamma (PolG) gene on chromosome 7. The expression of PolG was not affected by the MelARV insertion. Further investigation of the biological significance of MelARV in melanoma formation is being undertaken. PMID- 10516027 TI - Assignment of the multifunctional NS3 protein of bovine viral diarrhea virus during RNA replication: an in vivo and in vitro study. AB - Studies on the replication of the pestivirus bovine viral diarrhea virus (BVDV) were considerably facilitated by the recent discovery of an autonomous subgenomic BVDV RNA replicon (DI9c). DI9c comprises mainly the untranslated regions of the viral genome and the coding region of the nonstructural proteins NS3, NS4A, NS4B, NS5A, and NS5B. To assess the significance of the NS3-associated nucleoside triphosphatase/helicase activity during RNA replication and to explore other functional features of NS3, we generated a repertoire of DI9c derivatives bearing in-frame mutations in different parts of the NS3 coding unit. Most alterations resulted in deficient replicons, several of which encoded an NS3 protein with an inhibited protease function. Three lesions permitted replication, though at a lower level than that of the wild-type RNA, i.e., replacement of the third position of the DEYH helicase motif II by either T or F and an insertion of four amino acid residues in the C-terminal part of NS3. While polyprotein proteolysis was found to be almost unaffected in these latter replicons, in vitro studies with the purified mutant NS3 proteins revealed a significantly impaired helicase activity for the motif II substitutions. NS3 with a DEFH motif, moreover, showed a significantly lower ATPase activity. In contrast, the C-terminal insertion had no negative impact on the ATPase/RNA helicase activity of NS3. All three mutations affected the synthesis of both replication products-negative-strand intermediate and progeny positive-strand RNA-in a symmetric manner. Unexpectedly, various attempts to rescue or enhance the replication capability of nonfunctional or less functional DI9c NS3 derivatives, respectively, by providing intact NS3 in trans failed. Our experimental data thus demonstrate that the diverse enzymatic activities of the NS3 protein-in particular the ATPase/RNA helicase-play a pivotal role even during early steps of the viral replication pathway. They may further indicate the C-terminal part of NS3 to be an important functional determinant of the RNA replication process. PMID- 10516028 TI - Evolution of two types of rhesus lymphocryptovirus similar to type 1 and type 2 Epstein-Barr virus. AB - Rhesus monkeys and other nonhuman Old World primates are naturally infected with lymphocryptoviruses (LCV) that are closely related to Epstein-Barr virus (EBV). A rhesus LCV isolate (208-95) was derived from a B-cell lymphoma in a simian immunodeficiency virus-infected rhesus macaque. The EBNA-2 homologues from 208-95 and a previous rhesus LCV isolate (LCL8664) were polymorphic on immunoblotting, so the EBNA-2 genes from these two rhesus LCV were cloned, sequenced, and compared. The EBNA-2 genes have 40% nucleotide and 41% amino acid identities, and the differences are similar to those between the type 1 and type 2 EBV EBNA-2. Sequence from a portion of the LMP1 gene which is extremely divergent among different LCV was virtually identical between the 208-95 and LCL8664 strains, confirming a common rhesus LCV background. Thus, the EBNA-2 polymorphism defines the presence of two different rhesus LCV types, and both rhesus LCV types were found to be prevalent in the rhesus monkey population at the New England Regional Primate Research Center. The existence of two rhesus LCV types suggests that the selective pressure for the evolution of two LCV types is shared by human and nonhuman primate hosts. PMID- 10516029 TI - Nonrandom distribution of hepatitis C virus quasispecies in plasma and peripheral blood mononuclear cell subsets. AB - The existence of an extrahepatic reservoir of hepatitis C virus (HCV) is suggested by differences in quasispecies composition between the liver, peripheral blood mononuclear cells, and serum. We studied HCV RNA compartmentalization in the plasma of nine patients, in CD19(+), CD8(+), and CD4(+) positively selected cells, and also in the negatively selected cell fraction (NF). HCV RNA was detected in all plasma samples, in seven of nine CD19(+), three of eight CD8(+), and one of nine CD4(+) cell samples, and in seven of eight NF cells. Cloning and sequencing of HVR1 in two patients showed a sequence grouping: quasispecies from a given compartment (all studied compartments for one patient and CD8(+) and NF for the other) were statistically more genetically like each other than like quasispecies from any other compartment. The characteristics of amino acid and nucleotide substitutions suggested the same structural constraints on HVR1, even in very divergent strains from the cellular compartments, and homogeneous selection pressure on the different compartments. These findings demonstrate the compartmental distribution of HCV quasispecies within peripheral blood cell subsets and have important implications for the study of extrahepatic HCV replication and interaction with the immune system. PMID- 10516030 TI - Picornavirus internal ribosome entry site elements target RNA cleavage events induced by the herpes simplex virus virion host shutoff protein. AB - The herpes simplex virus (HSV) virion host shutoff (vhs) protein (UL41 gene product) is a component of the HSV virion tegument that triggers shutoff of host protein synthesis and accelerated mRNA degradation during the early stages of HSV infection. vhs displays weak amino acid sequence similarity to the fen-1 family of nucleases and suffices to induce accelerated RNA turnover through endoribonucleolytic cleavage events when it is expressed as the only HSV protein in a rabbit reticulocyte in vitro translation system. Although vhs selectively targets mRNAs in vivo, the basis for this selectivity remains obscure, since in vitro activity is not influenced by the presence of a 5' cap or 3' poly(A) tail. Here we show that vhs activity is greatly altered by placing an internal ribosome entry site (IRES) from encephalomyocarditis virus or poliovirus in the RNA substrate. Transcripts bearing the IRES were preferentially cleaved by the vhs dependent endoribonuclease at multiple sites clustered in a narrow zone located immediately downstream of the element in a reaction that did not require ribosomes. Targeting was observed when the IRES was located at the 5' end or placed at internal sites in the substrate, indicating that it is independent of position or sequence context. These data indicate that the vhs-dependent nuclease can be selectively targeted by specific cis-acting elements in the RNA substrate, possibly through secondary structure or a component of the translational machinery. PMID- 10516031 TI - Genetic regulation of long-term nonprogression in E-55+ murine leukemia virus infection in mice. AB - Certain inbred mouse strains display progression to lymphoma development after infection with E-55+ murine leukemia virus (E-55+ MuLV), while others demonstrate long-term nonprogression. This difference in disease progression occurs despite the fact that E-55+ MuLV causes persistent infection in both immunocompetent BALB/c-H-2(k) (BALB.K) progressor (P) and C57BL/10-H-2(k) (B10.BR) long-term nonprogressor (LTNP) mice. In contrast to immunocompetent mice, immunosuppressed mice from both P and LTNP strains develop lymphomas about 2 months after infection, indicating that the LTNP phenotype is determined by the immune response of the infected mouse. In this study, we used bone marrow chimeras to demonstrate that the LTNP phenotype is associated with the genotype of donor bone marrow and not the recipient microenvironment. In addition, we have mapped a genetic locus that may be responsible for the LTNP trait. Microsatellite-based linkage analysis demonstrated that a non-major histocompatibility complex gene on chromosome 15 regulates long-term survival and is located in the same region as the Rfv3 gene. Rfv3 is involved in recovery from Friend virus-induced leukemia and has been demonstrated to regulate neutralizing virus antibody titers. In our studies, however, both P and LTNP strains produce similar titers of neutralizing and cytotoxic anti-E-55+ MuLV. Therefore, while it is possible that Rfv3 influences the course of E-55+ MuLV infection, it is more likely that the LTNP phenotype in E-55+ MuLV-infected mice is regulated by a different, closely linked gene. PMID- 10516032 TI - Sendai virus gene start signals are not equivalent in reinitiation capacity: moderation at the fusion protein gene. AB - In paramyxovirus transcription, viral RNA polymerase synthesizes each monocistronic mRNA by recognizing the gene start (S) and end (E) signals flanking each gene. These signal sequences are well conserved in the virus family; nevertheless, they do exhibit some variations even within a virus species. In Sendai virus (SeV) Z strain, the E signals are identical for all six genes but there are four (N, P/M/HN, F, and L) different S signals with one or two nucleotide variations. The significance of these variations for in vitro and in vivo replication has been unknown. We addressed this issue by SeV reverse genetics. The luciferase gene was placed between the N and P gene so that recombinant SeVs expressed luciferase under the control of each of the four different S signals. The S signal for the F gene was found to drive a lower level of transcription than that of the other three, which exhibited comparable reinitiation capacities. The polar attenuation of SeV transcription thus appeared to be not linear but biphasic. Then, a mutant SeV whose F gene S signal was replaced with that used for the P, M, and HN genes was created, and its replication capability was examined. The mutant produced a larger amount of F protein and downstream gene-encoded proteins and replicated faster than wild-type SeV in cultured cells and in embryonated eggs. Compared with the wild type, the mutant virus also replicated faster in mice and was more virulent, requiring a dose 20 times lower to kill 50% of mice. On the other hand, the unique F start sequence as well as the other start sequences are perfectly conserved in all SeV isolates sequenced to date, including highly virulent fresh isolates as well as egg-adapted strains, with a virulence several magnitudes lower than that of the fresh isolates. This moderation of transcription at the F gene may therefore be relevant to viral fitness in nature. PMID- 10516033 TI - trans-Complementation analysis of the flavivirus Kunjin ns5 gene reveals an essential role for translation of its N-terminal half in RNA replication. AB - Recently we described rescue of defective Kunjin virus (KUN) RNAs with small deletions in the methyltransferase and RNA polymerase motifs of the ns5 gene, using BHK cells stably expressing KUN replicon RNA (repBHK cells) as helper (A. A. Khromykh et al., J. Virol. 72:7270-7279, 1998). We have now extended our previous observations and report successful trans-complementation of defective KUN RNAs with most of the ns5 gene deleted or substituted with a heterologous (dengue virus) ns5 sequence. Replication of full-length KUN RNAs with 3'-terminal deletions of 136 (5%), 933 (34%), and 1526 (56%) nucleotides in the ns5 gene was complemented efficiently in transfected repBHK cells. RNA with a larger deletion of 2,042 nucleotides (75%) was complemented less efficiently, and RNA with an even larger deletion of 2,279 nucleotides (84%) was not complemented at all. Chimeric KUN genomic RNA containing 87% of the KUN ns5 gene replaced by the corresponding sequence of the dengue virus type 2 ns5 gene was unable to replicate in normal BHK cells but was complemented in repBHK cells. These results demonstrate for the first time complementation of flavivirus RNAs with large deletions (as much as 75%) in the RNA polymerase gene and establish that translation of most of the N-terminal half of NS5 is essential for complementation in trans. A model of formation of the flavivirus replication complex implicating a possible role in RNA replication of conserved coding sequences in the N-terminal half of NS5 is proposed based on the complementation and earlier results with KUN and on reported data with other flaviviruses. PMID- 10516034 TI - Genetic stability of foamy viruses: long-term study in an African green monkey population. AB - The genetic variability of the envelope surface domain (SU) of simian foamy virus (FV) of African green monkeys was studied. To assess the interindividual diversity of FV, isolates were obtained from 19 animals living together in a monkey house. The monkeys had been imported from Kenya prior to being placed in long-term housing in the research institute. In addition, a simian FV isolate and proviral DNA were obtained from an animal caretaker infected in this setting. DNA of the complete SU (1779 to 1793 bp) was analyzed by PCR and sequencing. The sequences revealed four clusters with high homologies (>95%). Between the clusters, divergencies ranged from 3 to 25%. Obviously, the clusters reflect four different strains or subtypes of simian FV type 3 that were prevalent in the colony. In contrast to lentiviruses, hypervariable regions could not be detected in the FV SU. Furthermore, to analyze the intraindividual diversity of FV, we investigated the virus population within an individual monkey at a given time point and its evolution over 13 years. For this purpose, 22 proviral SU clones generated by PCR from one oral swab and seven isolates obtained from the same animal between 1982 and 1995 were examined. These sequences revealed exceptionally high homology rates (99.5 to 100%), and only a minimal genetic drift was recognized within the series of isolates. In conclusion, the low in vivo divergency of FV SU suggests that genetic variability is not important for the maintenance of FV persistence. PMID- 10516035 TI - Binding of polyomavirus small T antigen to protein phosphatase 2A is required for elimination of p27 and support of S-phase induction in concert with large T antigen. AB - Although polyomavirus large T antigen readily transactivates S-phase-specific enzymes in serum-starved Swiss 3T3 mouse fibroblasts, it is incapable by itself to efficiently drive such cells into S phase. We describe here that this inability correlates with a weak proficiency of the viral protein to induce the synthesis of cyclin A and cyclin E and to stimulate the respective cyclin/cdk activities. Polyomavirus small T antigen, which together with the large T protein supports S-phase induction, strongly contributes to the synthesis of cyclin A. In addition, small T antigen causes a dramatic induction of cyclin A- and, together with large T antigen, of cyclin E-specific protein kinase activity. This latter function of polyomavirus small T antigen correlates with its competence to provoke the elimination of the kinase inhibitor p27(Kip1). An interaction of the small T antigen with the protein phosphatase 2A is essential for this activity. Hence, the ability to drive quiescent Swiss 3T3 cells into S phase results from the capacity of large T antigen to transactivate DNA synthesis enzymes by its interaction with retinoblastoma-type proteins and from the potential of the large and the small T antigens together to stimulate cyclin A synthesis and cyclin A- and cyclin E-dependent protein kinase activity. PMID- 10516036 TI - Human cytomegalovirus 86-kilodalton IE2 protein blocks cell cycle progression in G(1). AB - The 86-kDa IE2 protein of human cytomegalovirus (HCMV) is an important regulator of viral and host cell gene expression. Still, besides its function as a transcription factor, little is known about the biological activities of IE2. Here, we show that IE2 can induce a G(1) arrest in several different cell lines, including HCMV-permissive U-373 cells. The known transcriptional activation domains of IE2 are dispensable for G(1) arrest, favoring a posttranscriptional mechanism mediating this cell cycle effect. We show that like human primary fibroblasts U-373 cells arrest in G(1) upon infection with HCMV. This G(1) arrest occurs within 24 h after infection and in proliferating cells depends on viral gene expression. Our data therefore suggest that IE2 is at least partially responsible for blocking the transition from G(1) to S phase, which is induced when cells are infected with HCMV. PMID- 10516038 TI - Incorporation of wild-type and C-terminally truncated human epidermal growth factor receptor into human immunodeficiency virus-like particles: insight into the processes governing glycoprotein incorporation into retroviral particles. AB - Previous results have indicated that incorporation of surface glycoprotein into retroviral particles is not a specific process and that many heterologous viral and cellular glycoproteins can be incorporated as long as they do not have long cytoplasmic C-terminal regions which were presumed to be sterically inhibitory. In this study, this concept has been directly examined by analyzing the incorporation of the wild-type human epidermal growth factor receptor (Wt-EGFR) and of a C-terminally truncated mutant of Wt-EGFR (Tr-EGFR) into human immunodeficiency virus (HIV)-like particles. Incorporation was directly analyzed at the protein level and by immunogold labelling of enriched HIV-like particles. In agreement with the above concept, Tr-EGFR, with only 7 C-terminal amino acids (aa), was efficiently incorporated into HIV-like particles. Incorporation of the Wt-EGFR species, with 542 C-terminal cytoplasmic aa, was reduced by a factor of about 5 in comparison to that of the Tr-EGFR species. However, the Wt-EGFR species was still very significantly present in the HIV-like particles. A series of control experiments verified that this represents genuine incorporation of Wt EGFR into the membrane of HIV-like particles. These observations allow further speculation as to the processes governing glycoprotein incorporation into retroviral particles and indicate that the internal virus structure of HIV (in particular the matrix layer [MA]) can accommodate much larger heterologous cytoplasmic domains in incorporated glycoproteins than previously assumed. PMID- 10516037 TI - Dual infection of gnotobiotic calves with bovine strains of group A and porcine like group C rotaviruses influences pathogenesis of the group C rotavirus. AB - There is serological evidence that bovine group C rotaviruses exist in the United States, but there are no reports of their isolation. Ninety fecal samples from calves with diarrhea, 81 samples from adult cows with diarrhea (winter dysentery), and 20 fecal samples from healthy adult cows were tested for group C rotaviruses by polyacrylamide gel electrophoresis, immune electron microscopy, and reverse transcription-PCR (RT-PCR). Three samples from adult cow diarrhea cases were positive only by RT-PCR, and a group C rotavirus was isolated from a positive sample in monkey kidney (MA104) cells (WD534tc/C). Genetically and serologically, the WD534tc/C strain was more closely related to the Cowden porcine group C strain than to the Shintoku bovine strain. Because the original cow feces also contained a group A rotavirus (detected after passage in cell culture), we hypothesized that such dual-rotavirus infections might play a role in the pathogenesis and host adaptation of rotaviruses. Thus, we examined the pathogenesis of WD534tc/C alone or combined with virulent (IND/A) or attenuated (NCDV/A) bovine group A rotaviruses in gnotobiotic calves. WD534tc/C alone induced diarrhea without (or with limited) virus shedding in inoculated calves (n = 3). In contrast, all calves coinfected with WD534tc/C and IND/A (n = 2) developed diarrhea and shed both viruses, whereas calves coinfected with WD534tc/C and NCDV/A (n = 3) developed diarrhea but did not shed either virus. Infection with WD534tc/C or NCDV/A alone caused only mild villous atrophy (jejunum and/or ileum), whereas dual infection with both viruses induced lesions throughout the small intestine. Although IND/A alone caused villous atrophy, more widespread small intestinal lesions occurred in calves coinfected with WD534tc/C and IND/A. In conclusion, coinfection of calves with group A rotaviruses enhanced fecal shedding of a bovine group C rotavirus and the extent of histopathological lesions in the small intestines. Thus, our findings suggest a potential novel hypothesis involving dual infections for the adaptation of heterologous rotaviruses to new host species. PMID- 10516039 TI - Generation of adenovirus vectors devoid of all viral genes by recombination between inverted repeats. AB - Direct or inverse repeated sequences are important functional features of prokaryotic and eukaryotic genomes. Considering the unique mechanism, involving single-stranded genomic intermediates, by which adenovirus (Ad) replicates its genome, we investigated whether repetitive homologous sequences inserted into E1 deleted adenoviral vectors would affect replication of viral DNA. In these studies we found that inverted repeats (IRs) inserted into the E1 region could mediate predictable genomic rearrangements, resulting in vector genomes devoid of all viral genes. These genomes (termed DeltaAd.IR) contained only the transgene cassette flanked on both sides by precisely duplicated IRs, Ad packaging signals, and Ad inverted terminal repeat sequences. Generation of DeltaAd.IR genomes could also be achieved by coinfecting two viruses, each providing one inverse homology element. The formation of DeltaAd.IR genomes required Ad DNA replication and appeared to involve recombination between the homologous inverted sequences. The formation of DeltaAd. IR genomes did not depend on the sequence within or adjacent to the inverted repeat elements. The small DeltaAd.IR vector genomes were efficiently packaged into functional Ad particles. All functions for DeltaAd.IR replication and packaging were provided by the full-length genome amplified in the same cell. DeltaAd.IR vectors were produced at a yield of approximately 10(4) particles per cell, which could be separated from virions with full-length genomes based on their lighter buoyant density. DeltaAd.IR vectors infected cultured cells with the same efficiency as first-generation vectors; however, transgene expression was only transient due to the instability of deleted genomes within transduced cells. The finding that IRs present within Ad vector genomes can mediate precise genetic rearrangements has important implications for the development of new vectors for gene therapy approaches. PMID- 10516040 TI - Integrating adenovirus-adeno-associated virus hybrid vectors devoid of all viral genes. AB - Recently, we demonstrated that inverted repeat sequences inserted into first generation adenovirus (Ad) vector genomes mediate precise genomic rearrangements resulting in vector genomes devoid of all viral genes that are efficiently packaged into functional Ad capsids. As a specific application of this finding, we generated adenovirus-adeno-associated virus (AAV) hybrid vectors, first generation Ad vectors containing AAV inverted terminal repeat sequences (ITRs) flanking a reporter gene cassette inserted into the E1 region. We hypothesized that the AAV ITRs present within the hybrid vector genome could mediate the formation of rearranged vector genomes (DeltaAd.AAV) and stimulate transgene integration. We demonstrate here that DeltaAd.AAV vectors are efficiently generated as by-products of first-generation adenovirus-AAV vector amplification. DeltaAd.AAV genomes contain only the transgene flanked by AAV ITRs, Ad packaging signals, and Ad ITRs. DeltaAd.AAV vectors can be produced at a high titer and purity. In vitro transduction properties of these deleted hybrid vectors were evaluated in direct comparison with first-generation Ad and recombinant AAV vectors (rAAVs). The DeltaAd.AAV hybrid vector stably transduced cultured cells with efficiencies comparable to rAAV. Since cells transduced with DeltaAd.AAV did not express cytotoxic viral proteins, hybrid viruses could be applied at very high multiplicities of infection to increase transduction rates. Southern analysis and pulsed-field gel electrophoresis suggested that DeltaAd.AAV integrated randomly as head-to-tail tandems into the host cell genome. The presence of two intact AAV ITRs was crucial for the production of hybrid vectors and for transgene integration. DeltaAd.AAV vectors, which are straightforward in their production, represent a promising tool for stable gene transfer in vitro and in vivo. PMID- 10516041 TI - Rep-mediated nicking of the adeno-associated virus origin requires two biochemical activities, DNA helicase activity and transesterification. AB - The single-stranded adeno-associated virus (AAV) genome is flanked by terminal hairpinned origins of DNA replication (terminal repeats [TRs]) that are nicked at the terminal resolution site (trs) by the AAV Rep protein in an ATP-dependent, site-specific manner. Here we determine the minimal trs sequence necessary for Rep cleavage, 3'-CCGGT/TG-5', and show that this 7-base core sequence is required only on the nicked strand. We also identify a potential stem-loop structure at the trs. Interestingly, Rep nicking on a TR substrate that fixes this trs stem loop in the extruded form no longer requires ATP. This suggests that ATP dependent Rep helicase activity is necessary to unwind the duplex trs and extrude the stem-loop structure, prior to the ATP-independent Rep transesterification reaction. The extrusion of origin stem-loop structures prior to nicking appears to be a general mechanism shared by plant and animal viruses and bacterial plasmids. In the case of AAV, this mechanism of TR nicking would provide a possible regulatory function. PMID- 10516042 TI - The mode and duration of anti-CD28 costimulation determine resistance to infection by macrophage-tropic strains of human immunodeficiency virus type 1 in vitro. AB - We have investigated the ability of anti-CD28 antibody costimulation to induce resistance to macrophage (M)-tropic strains of human immunodeficiency virus type 1 (HIV-1) in vitro. Our results confirm the observations of Levine et al. (15) that stimulation of CD4 T cells with anti-CD3/anti-CD28 antibodies coimmobilized on magnetic beads renders the cells resistant to infection by M-tropic strains of HIV-1. The resistance was strongest when the beads were left in the cultures throughout the experiment. In contrast, stimulation of CD4 T cells with the same antibodies immobilized on the surface of plastic culture dishes failed to induce resistance and resulted in high levels of p24 production. This was true even if the cells were passaged continuously on freshly coated plates. If the beads were removed after initial stimulation, p24 production increased over time and produced a result intermediate to the other forms of stimulation. For beads-in, beads-out, and one-time plate stimulated cultures, resistance to infection correlated with down-regulation of CCR5 expression at the cell surface and with increased production of beta-chemokines. However, cultures of CD4 T cells continuously passaged on anti-CD3/anti-CD28-coated plates produced large amounts of p24 despite decreased levels of CCR5 expression and increasing production of beta-chemokines. Expression of the T-cell activation markers CD25 and CD69 and production of gamma interferon further supported the differences in plate versus bead stimulation. Our results explain the apparent contradiction between the ability of anti-CD28 antibody costimulation to induce resistance to HIV infection when presented on magnetic beads and the increased ability to recover virus from the cells of HIV-positive donors who are on highly active antiretroviral therapy when cells are stimulated by anti-CD3/anti-CD28 immobilized on plastic dishes. PMID- 10516044 TI - Polymorphisms of the cell surface receptor control mouse susceptibilities to xenotropic and polytropic leukemia viruses. AB - The differential susceptibilities of mouse strains to xenotropic and polytropic murine leukemia viruses (X-MLVs and P-MLVs, respectively) are poorly understood but may involve multiple mechanisms. Recent evidence has demonstrated that these viruses use a common cell surface receptor (the X-receptor) for infection of human cells. We describe the properties of X-receptor cDNAs with distinct sequences cloned from five laboratory and wild strains of mice and from hamsters and minks. Expression of these cDNAs in resistant cells conferred susceptibilities to the same viruses that naturally infect the animals from which the cDNAs were derived. Thus, a laboratory mouse (NIH Swiss) X-receptor conferred susceptibility to P-MLVs but not to X-MLVs, whereas those from humans, minks, and several wild mice (Mus dunni, SC-1 cells, and Mus spretus) mediated infections by both X-MLVs and P-MLVs. In contrast, X-receptors from the resistant mouse strain Mus castaneus and from hamsters were inactive as viral receptors. These results suggest that X-receptor polymorphisms are a primary cause of resistances of mice to members of the X-MLV/P-MLV family of retroviruses and are responsible for the xenotropism of X-MLVs in laboratory mice. By site-directed mutagenesis, we substituted sequences between the X-receptors of M. dunni and NIH Swiss mice. The NIH Swiss protein contains two key differences (K500E in presumptive extracellular loop 3 [ECL 3] and a T582 deletion in ECL 4) that are both required to block X-MLV infections. Accordingly, a single inverse mutation in the NIH Swiss protein conferred X-MLV susceptibility. Furthermore, expression of an X-MLV envelope glycoprotein in Chinese hamster ovary cells interfered efficiently with X-MLV and P-MLV infections mediated by X-receptors that contained K500 and/or T582 but had no effect on P-MLV infections mediated by X-receptors that lacked these amino acids. In contrast, moderate expression of a P-MLV (MCF247) envelope glycoprotein did not cause substantial interference, suggesting that X-MLV and P MLV glycoproteins interfere nonreciprocally with X-receptor-mediated infections. We conclude that P-MLVs have become adapted to utilize X-receptors that lack K500 and T582. A penalty for this adaptation is a reduced ability to interfere with superinfection. Because failure of interference is a hallmark of several exceptionally pathogenic retroviruses, we propose that it contributes to P-MLV induced diseases. PMID- 10516043 TI - Transcriptional activation by the product of open reading frame 50 of Kaposi's sarcoma-associated herpesvirus is required for lytic viral reactivation in B cells. AB - Kaposi's sarcoma (KS)-associated herpesvirus (KSHV) is a lymphotropic virus strongly linked to the development of KS, an endothelial cell neoplasm frequent in persons with AIDS. Reactivation from latency in B cells is thought to be an important antecedent to viral spread to endothelial cells during KS pathogenesis. Earlier experiments have posited a role for the transcriptional activator encoded by KSHV open reading frame 50 (ORF50) in such reactivation, since ectopic overexpression of this protein induces reactivation in latently infected B cells. Here we have explored several aspects of the expression, structure, and function of this protein bearing on this role. The ORF50 gene is expressed very early in lytic reactivation, before several other genes implicated as candidate regulatory genes in related viruses, and its expression can upregulate their promoters in transient assays. The protein is extensively phosphorylated in vivo and bears numerous sites for phosphorylation by protein kinase C, activators of which are potent stimulators of lytic induction. The C terminus of the ORF50 protein contains a domain that can strongly activate transcription when targeted to DNA; deletion of this domain generates an allele that expresses a truncated protein which retains the ability to form multimers with full-length ORF50 and functions as a dominant-negative protein. Expression of this allele in latently infected cells ablates spontaneous reactivation from latency and strikingly suppresses viral replication induced by multiple stimuli, including phorbol ester, ionomycin, and sodium butyrate. These results indicate that the ORF50 gene product plays an essential role in KSHV lytic replication and are consistent with its action as a putative molecular switch controlling the induction of virus from latency. PMID- 10516045 TI - Amino acid changes at positions 173 and 187 in the human T-cell leukemia virus type 1 surface glycoprotein induce specific neutralizing antibodies. AB - The nucleotide sequence of human T-cell leukemia virus type 1 (HTLV-1) is highly conserved, most strains sharing at least 95% sequence identity. This sequence conservation is also found in the viral env gene, which codes for the two envelope glycoproteins that play a major role in the induction of a protective immune response against the virus. However, recent reports have indicated that some variations in env sequences may induce incomplete cross-reactivity between HTLV-1 strains. To identify the amino acid changes that might be involved in the antigenicity of neutralizable epitopes, we constructed expression vectors coding for the envelope glycoproteins of two HTLV-1 isolates (2060 and 2072) which induced human antibodies with different neutralization patterns. The amino acid sequences of the envelope glycoproteins differed at four positions. Vectors coding for chimeric or point-mutated envelope proteins were derived from 2060 and 2072 HTLV-1 env genes. Syncytium formation induced by the wild-type or mutated envelope proteins was inhibited by human sera with different neutralizing specificities. We thus identified two amino acid changes, I173-->V and A187-->T, that play an important role in the antigenicity of neutralizable epitopes located in this region of the surface envelope glycoprotein. PMID- 10516046 TI - Induction of syncytia by neuropathogenic murine leukemia viruses depends on receptor density, host cell determinants, and the intrinsic fusion potential of envelope protein. AB - Infection by the neuropathogenic murine leukemia virus (MLV) TR1.3 results in hemorrhagic disease that correlates directly to in vivo syncytium formation of brain capillary endothelial cells (BCEC). This phenotype maps to amino acid 102 in the envelope (Env) protein of TR1.3. Substitution of glycine (G) for tryptophan (W) at this position (W102G Env) in the nonpathogenic MLV FB29 induces both syncytium formation and neurologic disease in vivo. Using an in vitro gene reporter cell fusion assay, we showed that fusion either with murine NIH 3T3 cells or with nonmurine target cells that expressed receptors at or below endogenous murine levels mirrored that seen in BCEC in vivo. In these instances only TR1.3 and W102G Env induced cell fusion. In contrast, when receptor levels on nonmurine cells were raised above endogenous murine levels, FB29 Env was as fusogenic as the neuropathogenic TR1.3 and W102G Env. These results indicate that TR1.3 Env and W102G Env are intrinsically more fusogenic than FB29 Env, that the induction of fusion requires a threshold number of receptors that is greater for FB29 Env than for TR1.3 or W102G Env, and that receptor density on murine NIH 3T3 cells and BCEC is below the threshold for FB29-dependent fusion. Surprisingly, receptor density on NIH 3T3 cells could not be increased by stable expression of exogenous receptors, and FB29-dependent fusion was not observed in NIH 3T3 cells that transiently expressed elevated receptor numbers. These results suggest that an additional undefined host cell factor(s) may limit both receptor expression and fusion potential in murine cells. PMID- 10516047 TI - Detection of bovine spongiform encephalopathy-specific PrP(Sc) by treatment with heat and guanidine thiocyanate. AB - The conversion of a ubiquitous cellular protein (PrP(C)), an isoform of the prion protein (PrP), to the pathology-associated isoform PrP(Sc) is one of the hallmarks of transmissible spongiform encephalopathies such as bovine spongiform encephalopathy (BSE). Accumulation of PrP(Sc) has been used to diagnose BSE. Here we describe a quantitative enzyme-linked immunosorbent assay (ELISA) that involves antibodies against epitopes within the protease-resistant core of the PrP molecule to measure the amount of PrP in brain tissues from animals with BSE and normal controls. In native tissue preparations, little difference was found between the two groups. However, following treatment of the tissue with heat and guanidine thiocyanate (Gh treatment), the ELISA discriminated BSE-specific PrP(Sc) from PrP(C) in bovine brain homogenates. PrP(Sc) was identified by Western blot, centrifugation, and protease digestion experiments. It was thought that folding or complexing of PrP(Sc) is most probably reversed by the Gh treatment, making hidden antigenic sites accessible. The digestion experiments also showed that protease-resistant PrP in BSE is more difficult to detect than that in hamster scrapie. While the concentration of PrP(C) in cattle is similar to that in hamsters, PrP(Sc) sparse in comparison. The detection of PrP(Sc) by a simple physicochemical treatment without the need for protease digestion, as described in this study, could be applied to develop a diagnostic assay to screen large numbers of samples. PMID- 10516048 TI - Sequence and transcriptional analyses of the fish retroviruses walleye epidermal hyperplasia virus types 1 and 2: evidence for a gene duplication. AB - Walleye epidermal hyperplasia virus types 1 and 2 (WEHV1 and WEHV2, respectively) are associated with a hyperproliferative skin lesion on walleyes that appears and regresses seasonally. We have determined the complete nucleotide sequences and transcriptional profiles of these viruses. WEHV1 and WEHV2 are large, complex retroviruses of 12,999 and 13,125 kb in length, respectively, that are closely related to one another and to walleye dermal sarcoma virus (WDSV). These walleye retroviruses contain three open reading frames, orfA, orfB, and orfC, in addition to gag, pol, and env. orfA and orfB are adjacent to one another and located downstream of env. The OrfA proteins were previously identified as cyclin D homologs that may contribute to the induction of cell proliferation leading to epidermal hyperplasia and dermal sarcoma. The sequence analysis of WEHV1 and WEHV2 revealed that the OrfB proteins are distantly related to the OrfA proteins, suggesting that orfB arose by gene duplication. Presuming that the precursor of orfA and orfB was derived from a cellular cyclin, these genes are the first accessory genes of complex retroviruses that can be traced to a cellular origin. WEHV1, WEHV2, and WDSV are the only retroviruses that have an open reading frame, orfC, of considerable size (ca. 130 amino acids) in the leader region preceding gag. While we were unable to predict a function for the OrfC proteins, they are more conserved than OrfA and OrfB, suggesting that they may be biologically important to the viruses. The transcriptional profiles of WEHV1 and WEHV2 were also similar to that of WDSV; Northern blot analyses detected only low levels of the orfA transcripts in developing lesions, whereas abundant levels of genomic, env, orfA, and orfB transcripts were detected in regressing lesions. The splice donors and acceptors of individual transcripts were identified by reverse transcriptase PCR. The similarities of WEHV1, WEHV2, and WDSV suggest that these viruses use similar strategies of viral replication and induce cell proliferation by a similar mechanism. PMID- 10516049 TI - Evolution of envelope sequences of human immunodeficiency virus type 1 in cellular reservoirs in the setting of potent antiviral therapy. AB - In human immunodeficiency virus (HIV)-infected patients treated with potent antiretroviral therapy, the persistence of latently infected cells may reflect the long decay half-life of this cellular reservoir or ongoing viral replication at low levels with continuous replenishment of the population or both. To address these possibilities, sequences encompassing the C2 and V3 domains of HIV-1 env were analyzed from virus present in baseline plasma and from viral isolates obtained after 2 years of suppressive therapy in six patients. The presence of sequence changes consistent with evolution was demonstrated for three subjects and correlated with less complete suppression of viral replication, as indicated by the rapidity of the initial virus load decline or the intermittent reappearance of even low levels of detectable viremia. Together, these results provide evidence for ongoing replication. In the remaining three patients, virus recovered after 2 years of therapy was either genotypically contemporary with or ancestral to virus present in plasma 2 years before, indicating that virus recovery had indeed resulted from activation of latently infected cells. PMID- 10516050 TI - Requirements for RNA replication of a poliovirus replicon by coxsackievirus B3 RNA polymerase. AB - A chimeric poliovirus type 1 (PV1) genome was constructed in which the 3D RNA polymerase (3D(pol)) coding sequences were replaced with those from coxsackievirus B3 (CVB3). No infectious virus was produced from HeLa cells transfected with the chimeric RNA. Processing of the PV1 capsid protein precursor was incomplete, presumably due to inefficient recognition of the P1 protein substrate by the chimeric 3CD proteinase containing CVB3 3D sequences. The ability of the chimeric RNA to replicate in the absence of capsid formation was measured after replacement of the P1 region with a luciferase reporter gene. No RNA synthesis was detected, despite efficient production of enzymatically active 3D(pol) from the 3D portion of the chimeric 3CD. The chimeric 3CD protein was unable to efficiently bind to the cloverleaf-like structure (CL) at the 5' end of PV1 RNA, which has been demonstrated previously to be required for viral RNA synthesis. The CVB3 3CD protein bound the PV1 CL as well as PV1 3CD. An additional chimeric PV1 RNA that contained CVB3 3CD sequences also failed to produce virus after transfection. Since processing of PV1 capsid protein precursors by the CVB3 3CD was again incomplete, a luciferase-containing replicon was also analyzed for RNA replication. The 3CD chimera replicated at 33 degrees C, but not at 37 degrees C. Replacement of the PV1 5'-terminal CL with that of CVB3 did not rescue the temperature-sensitive phenotype. Thus, there is an essential interaction(s) between 3CD and other viral P2 or P3 protein products required for efficient RNA replication which is not fully achieved between proteins from the two different members of the same virus genus. PMID- 10516051 TI - Establishment and characterization of cytopathogenic and noncytopathogenic pestivirus replicons. AB - Defective interfering particles (DIs) of bovine viral diarrhea virus (BVDV) have been identified and shown to be cytopathogenic (cp) in the presence of noncytopathogenic (noncp) helper virus. Moreover, a subgenomic (sg) RNA corresponding in its genome structure to one of those BVDV DIs (DI9) was replication competent in the absence of helper virus. We report here that an sg BVDV replicon which encodes from the viral proteins only the first three amino acids of the autoprotease N(pro) in addition to nonstructural (NS) proteins NS3 to NS5B replicates autonomously and also induces lysis of its host cells. This demonstrates that the presence of a helper virus is not required for the lysis of the host cell. On the basis of two infectious BVDV cDNA clones, namely, BVDV CP7 (cp) and CP7ins- (noncp), bicistronic replicons expressing proteins NS2-3 to NS5B were established. These replicons express, in addition to the viral proteins, the reporter gene encoding beta-glucuronidase; the release of this enzyme from transfected culture cells was used to monitor cell lysis. Applying these tools, we were able to show that the replicon derived from CP7ins- does not induce cell lysis. Accordingly, neither N(pro) nor any of the structural proteins are necessary to maintain the noncp phenotype. Furthermore, these sg RNAs represent the first pair of cp and noncp replicons which mimic complete BVDV CP7 and CP7ins with respect to cytopathogenicity. These replicons will facilitate future studies aimed at the determination of the molecular basis for the cytopathogenicity of BVDV. PMID- 10516052 TI - Charge-to-alanine mutagenesis of the adeno-associated virus type 2 Rep78/68 proteins yields temperature-sensitive and magnesium-dependent variants. AB - The adeno-associated virus type 2 (AAV) replication (Rep) proteins Rep78 and 68 (Rep78/68) exhibit a number of biochemical activities required for AAV replication, including specific binding to a 22-bp region of the terminal repeat, site-specific endonuclease activity, and helicase activity. Individual and clusters of charged amino acids were converted to alanines in an effort to generate a collection of conditionally defective Rep78/68 proteins. Rep78 variants were expressed in human 293 cells and analyzed for their ability to mediate replication of recombinant AAV vectors at various temperatures. The biochemical activities of Rep variants were further characterized in vitro by using Rep68 His-tagged proteins purified from bacteria. The results of these analyses identified a temperature-sensitive (ts) Rep protein (D40,42,44A-78) that exhibited a delayed replication phenotype at 32 degrees C, which exceeded wild type activity by 48 h. Replication activity was reduced by more than threefold at 37 degrees C and was undetectable at 39 degrees C. Stability of the Rep78 protein paralleled replication levels at each temperature, further supporting a ts phenotype. Replication differences resulted in a 3-log-unit difference in virus yields between the permissive and nonpermissive temperatures (2.2 x 10(6) and 3 x 10(3), respectively), demonstrating that this is a relatively tight mutant. In addition to the ts Rep mutant, we identified a nonconditional mutant with a reduced ability to support viral replication in vivo. Additional characterization of this mutant demonstrated an Mg(2+)-dependent phenotype that was specific to Rep endonuclease activity and did not affect helicase activity. The two mutants described here are unique, in that Rep ts mutants have not previously been described and the D412A Rep mutant represents the first mutant in which the helicase and endonuclease functions can be distinguished biochemically. Further understanding of these mutants should facilitate our understanding of AAV replication and integration, as well as provide novel strategies for production of viral vectors. PMID- 10516053 TI - Repeated delivery of adeno-associated virus vectors to the rabbit airway. AB - Efficient local expression from recombinant adeno-associated virus (rAAV)-cystic fibrosis (CF) transmembrane conductance regulator (CFTR) vectors has been observed in the airways of rabbits and monkeys for up to 6 months following a single bronchoscopic delivery. However, it is likely that repeated administrations of rAAV vectors will be necessary for sustained correction of the CF defect in the airways. The current study was designed to test the feasibility of repeated airway delivery of rAAV vectors in the rabbit lung. After two doses of rAAV-CFTR to the airways, rabbits generated high titers of serum anti-AAV neutralizing antibodies. Rabbits then received a third dose of a rAAV vector containing the green fluorescent protein (GFP) reporter gene packaged in either AAV serotype 2 (AAV2) or serotype 3 (AAV3) capsids. Each dose consisted of 1 ml containing 5 x 10(9) DNase-resistant particles of rAAV vector, having no detectable replication-competent AAV or adenovirus. Three weeks later, GFP expression was observed in airway epithelial cells despite high anti-AAV neutralizing titers at the time of delivery. There was no significant difference in the efficiency of DNA transfer or expression between the rAAV3 and rAAV2 groups. No significant inflammatory responses to either repeated airway exposure to rAAV2-CFTR vectors or to GFP expression were observed. These experiments demonstrate that serum anti-AAV neutralizing antibody titers do not predict airway neutralization in vivo and that repeated airway delivery rAAV allows for safe and effective gene transfer. PMID- 10516054 TI - Herpes simplex virus type 1 immediate-early protein Vmw110 inhibits progression of cells through mitosis and from G(1) into S phase of the cell cycle. AB - Herpes simplex virus type 1 (HSV-1) immediate-early protein Vmw110 stimulates the onset of virus infection in a multiplicity-dependent manner and is required for efficient reactivation from latency. Recent work has shown that Vmw110 is able to interact with or modify the stability of several cellular proteins. In this report we analyze the ability of Vmw110 to inhibit the progression of cells through the cell cycle. We show by fluorescence-activated cell sorter and/or confocal microscopy analysis that an enhanced green fluorescent protein-tagged Vmw110 possesses the abilities both to prevent transfected cells moving from G(1) into S phase and to block infected cells at an unusual stage of mitosis defined as pseudo-prometaphase. The latter property correlates with the Vmw110-induced proteasome-dependent degradation of CENP-C, a centromeric protein component of the inner plate of human kinetochores. We also show that whereas Vmw110 is not the only viral product implicated in the block of infected cells at the G(1)/S border, the mitotic block is a specific property of Vmw110 and more particularly of its RING finger domain. These data explain the toxicity of Vmw110 when expressed alone in transfected cells and provide an explanation for the remaining toxicity of replication-defective mutants of HSV-1 expressing Vmw110. In addition to contributing to our understanding of the effects of Vmw110 on the cell, our results demonstrate that Vmw110 expression is incompatible with the proliferation of a dividing cell population. This factor is of obvious importance to the design of gene therapy vectors based on HSV-1. PMID- 10516055 TI - Concatamerization of adeno-associated virus circular genomes occurs through intermolecular recombination. AB - Long-term recombinant AAV (rAAV) transgene expression in muscle has been associated with the molecular conversion of single-stranded rAAV genomes to high molecular-weight head-to-tail circular concatamers. However, the mechanisms by which these large multimeric concatamers form remain to be defined. To this end, we tested whether concatamerization of rAAV circular intermediates occurs through intra- or intermolecular mechanisms of amplification. Coinfection of the tibialis muscle of mice with rAAV alkaline phosphatase (Alkphos)- and green fluorescent protein (GFP)-encoding vectors was used to evaluate the frequency of circular concatamer formation by intermolecular recombination of independent viral genomes. The GFP shuttle vector also encoded ampicillin resistance and contained a bacterial origin of replication to allow for bacterial rescue of circular intermediates from Hirt DNA of infected muscle samples. The results demonstrated a time-dependent increase in the abundance of rescued plasmids encoding both GFP and Alkphos, which reached 33% of the total circular intermediates by 120 days postinfection. Furthermore, these large circular concatamers were capable of expressing both GFP- and Alkphos-encoding transgenes following transient transfection in cell lines. These findings demonstrate that concatamerization of AAV genomes in vivo occurs through intermolecular recombination of independent monomer circular viral genomes and suggest new viable strategies for delivering multiple DNA segments at a single locus. Such developments will expand the utility of rAAV for splicing large gene inserts or large promoter-gene combinations carried by two or more independent rAAV vectors. PMID- 10516056 TI - Mapping of a hypovirus p29 protease symptom determinant domain with sequence similarity to potyvirus HC-Pro protease. AB - Hypovirus infection of the chestnut blight fungus Cryphonectria parasitica results in a spectrum of phenotypic changes that can include alterations in colony morphology and significant reductions in pigmentation, asexual sporulation, and virulence (hypovirulence). Deletion of 88% [Phe(25) to Pro(243)] of the virus-encoded papain-like protease, p29, in the context of an infectious cDNA clone of the prototypic hypovirus CHV1-EP713 (recombinant virus Deltap29) partially relieved virus-mediated suppression of pigmentation and sporulation without altering the level of hypovirulence. We now report mapping of the p29 symptom determinant domain to a region extending from Phe(25) through Gln(73) by a gain-of-function analysis following progressive repair of the Deltap29 deletion mutant. This domain was previously shown to share sequence similarity [including conserved cysteine residues Cys(38), Cys(48), Cys(70), and Cys(72)] with the N terminal portion of the potyvirus-encoded helper component-proteinase (HC-Pro), a multifunctional protein implicated in aphid-mediated transmission, genome amplification, polyprotein processing, long-distance movement, and suppression of posttranscriptional silencing. Substitution of a glycine residue for either Cys(38) or Cys(48) resulted in no qualitative or quantitative changes in virus mediated symptoms. Unexpectedly, mutation of Cys(70) resulted in a very severe phenotype that included significantly reduced mycelial growth and profoundly altered colony morphology. In contrast, substitution for Cys(72) resulted in a less severe symptom phenotype approaching that observed for Deltap29. The finding that p29-mediated symptom expression is influenced by two cysteine residues that are conserved in the potyvirus-encoded HC-Pro raises the possibility that these related viral-papain-like proteases function in their respective fungal and plant hosts by impacting ancestrally related regulatory pathways. PMID- 10516058 TI - cORF and RcRE, the Rev/Rex and RRE/RxRE homologues of the human endogenous retrovirus family HTDV/HERV-K. AB - cORF, a protein encoded by the human endogenous retrovirus family HTDV/HERV-K, contains amino acid motifs which resemble the nuclear import and export signals of the viral regulatory proteins Rev (human immunodeficiency virus) and Rex (human T-cell leukemia virus [HTLV]). In this study, we demonstrated that cORF indeed has a Rev-like function and mapped the cORF-responsive RNA element to a sequence in the 3' long terminal repeat, a localization similar to RxRE, the responsive element in HTLV type 1. Accordingly, we have given the element the designation RcRE. cORF and RcRE stabilize unspliced and incompletely spliced viral transcripts and enhance their nuclear export via the CRM1 export pathway. So far, HTDV/HERV-K is the only endogenous retrovirus family with a complex regulation at the posttranscriptional level. It may be regarded as an intermediate in the evolution from simple to complex retroviruses. PMID- 10516057 TI - Mucosal immunization of cynomolgus macaques with two serotypes of live poliovirus vectors expressing simian immunodeficiency virus antigens: stimulation of humoral, mucosal, and cellular immunity. AB - Poliovirus live virus vectors are a candidate recombinant vaccine system. Previous studies using this system showed that a live poliovirus vector expressing a foreign antigen between the structural and nonstructural proteins generates both antibody and cytotoxic T-lymphocyte responses in mice. Here we describe a novel in vitro method of cloning recombinant polioviruses involving a hybrid-PCR approach. We report the construction of recombinant vectors of two different serotypes of poliovirus-expressing simian immunodeficiency virus (SIV) antigens and the intranasal and intravenous inoculations of four adult cynomolgus macaques with these poliovirus vectors expressing the SIV proteins p17(gag) and gp41(env). All macaques generated a mucosal anti-SIV immunoglobulin A (IgA) response in rectal secretions. Two of the four macaques generated mucosal antibody responses detectable in vaginal lavages. Strong serum IgG responses lasting for at least 1 year were detected in two of the four monkeys. SIV specific T-cell lymphoproliferative responses were detected in three of the four monkeys. SIV-specific cytotoxic T lymphocytes were detected in two of the four monkeys. This is the first report of poliovirus-elicited vaginal IgA or cytotoxic T lymphocytes in any naturally infectable primate, including humans. These findings support the concept that a live poliovirus vector is a potentially useful delivery system that elicits humoral, mucosal, and cellular immune responses against exogenous antigens. PMID- 10516059 TI - Mutations in the DG loop of adenovirus type 5 fiber knob protein abolish high affinity binding to its cellular receptor CAR. AB - The amino acid residues in adenovirus type 5 (Ad5) fiber that interact with its cellular receptor, the coxsackie B virus and Ad receptor (CAR), have not been defined. To investigate this, multiple mutations were constructed in the region between residues 479 and 497 in Ad5 fiber (beta-strands E and F and the adjacent region of the DG loop). The effects of these mutations on binding to CAR were determined by use of cell-binding competition experiments, surface plasmon resonance, and direct binding studies. The mutation effects on the overall folding and secondary structure of the protein were assessed by circular dichroism (CD) spectroscopy. Deletions of two consecutive amino acids between residues 485 and 493 abolished high-affinity binding to CAR; the CD spectra indicated that although there was no disruption of the overall folding and secondary structure of the protein, local conformational changes did occur. Moreover, single site mutations in this region of residues with exposed, surface accessible side chains, such as Thr492, Asn493, and Val495, had no effect on receptor binding, which demonstrates that these residues are not in contact with CAR themselves. This implies the involvement of residues in neighboring loop regions. Replacement of the segment containing the two very short beta-strands E and F and the turn between them (residues 479 to 486) with the corresponding sequence from Ad3 (betaEFAd3-->5 mutation) resulted in the loss of receptor binding. The identical CD spectra for betaEFAd3-->5 and wild-type proteins suggest that these substitutions caused no conformational rearrangement and that the loss of binding may thus be due to the substitution of one or more critical contact residues. These findings have implications for our understanding of the interaction of Ad5 fiber with CAR and for the construction of targeted recombinant Ad5 vectors for gene therapy purposes. PMID- 10516060 TI - Effects of targeting herpes simplex virus type 1 gD to the endoplasmic reticulum and trans-Golgi network. AB - Glycoprotein D (gD) of herpes simplex virus type 1 (HSV-1) was modified to encode targeting signals known to localize proteins to either the endoplasmic reticulum (ER) or the trans-Golgi network. These motifs conferred the predicted targeting properties on gD in transfected cells as judged by immunofluorescence staining, and the exclusion of targeted gD from the cell surface was confirmed by the fact that these molecules exhibited substantially reduced activity in cell-cell fusion assays. Recombinant viruses expressing Golgi-targeted forms of gD grew to wild type levels in noncomplementing cells, exhibited unaltered particle/infectivity ratios, and were found to contain wild-type levels of gD, whereas a recombinant expressing ER-retained gD was helper cell dependent and, when grown on noncomplementing cells, produced virions of low specific infectivity with greatly reduced levels of gD. These data imply that HSV-1 acquires its final membrane from a post-ER compartment and lend support to the view that the virus undergoes de-envelopment and reenvelopment steps during virus egress. PMID- 10516061 TI - Circuit-specific coinfection of neurons in the rat central nervous system with two pseudorabies virus recombinants. AB - Neurotropic alphaherpesviruses have become popular tools for transynaptic analysis of neural circuitry. It has also been demonstrated that coinfection with two viruses expressing unique reporters can be used to define more complicated circuitry. However, the coinfection studies reported to date have employed nonisogenic strains that differ in their invasive properties. In the present investigation we used two antigenically distinct recombinants of the swine pathogen pseudorabies virus (PRV) in single and double infections of the rat central nervous system. Both viruses are derivatives of PRV-Bartha, a strain with reduced virulence that is widely used for circuit analysis. PRV-BaBlu expresses beta-galactosidase, and PRV-D expresses the PRV membrane protein gI, the gene for which is deleted in PRV-BaBlu. Antibodies to beta-galactosidase identify neurons infected with PRV-BaBlu, and antibodies monospecific for PRV gI identify neurons infected with PRV-D. The ability of these strains to establish coinfections in neurons was evaluated in visual and autonomic circuitry in which the parental virus has previously been characterized. The following conclusions can be drawn from these experiments. First, PRV-D is significantly more neuroinvasive than PRV Bartha or PRV-BaBlu in the same circuitry. Second, PRV-D is more virulent than either PRV-Bartha or PRV-BaBlu, and PRV-BaBlu is less virulent than PRV-Bartha. Third, in every model examined, PRV-D and PRV-BaBlu coinfect some neurons, but single infections predominate. Fourth, prior infection with one virus renders neurons less permissive to infection by another virus. Fifth, prior infection by PRV-D is more effective than PRV-BaBlu in reducing invasion and spread of the second virus. Collectively, the data define important variables that must be considered in coinfection experiments and suggest that the most successful application of this approach would be accomplished by using isogenic strains of virus with equivalent virulence. PMID- 10516062 TI - Mutant cells selected during persistent reovirus infection do not express mature cathepsin L and do not support reovirus disassembly. AB - Persistent reovirus infections of murine L929 cells select cellular mutations that inhibit viral disassembly within the endocytic pathway. Mutant cells support reovirus growth when infection is initiated with infectious subvirion particles (ISVPs), which are intermediates in reovirus disassembly formed following proteolysis of viral outer-capsid proteins. However, mutant cells do not support growth of virions, indicating that these cells have a defect in virion-to-ISVP processing. To better understand mechanisms by which viruses use the endocytic pathway to enter cells, we defined steps in reovirus replication blocked in mutant cells selected during persistent infection. Subcellular localization of reovirus after adsorption to parental and mutant cells was assessed using confocal microscopy and virions conjugated to a fluorescent probe. Parental and mutant cells did not differ in the capacity to internalize virions or distribute them to perinuclear compartments. Using pH-sensitive probes, the intravesicular pH was determined and found to be equivalent in parental and mutant cells. In both cell types, virions localized to acidified intracellular organelles. The capacity of parental and mutant cells to support proteolysis of reovirus virions was assessed by monitoring the appearance of disassembly intermediates following adsorption of radiolabeled viral particles. Within 2 h after adsorption to parental cells, proteolysis of viral outer-capsid proteins was observed, consistent with formation of ISVPs. However, in mutant cells, no proteolysis of viral proteins was detected up to 8 h postadsorption. Since treatment of cells with E64, an inhibitor of cysteine-containing proteases, blocks reovirus disassembly, we used immunoblot analysis to assess the expression of cathepsin L, a lysosomal cysteine protease. In contrast to parental cells, mutant cells did not express the mature, proteolytically active form of the enzyme. The defect in cathepsin L maturation was not associated with mutations in procathepsin L mRNA, was not complemented by procathepsin L overexpression, and did not affect the maturation of cathepsin B, another lysosomal cysteine protease. These findings indicate that persistent reovirus infections select cellular mutations that affect the maturation of cathepsin L and suggest that alterations in the expression of lysosomal proteases can modulate viral cytopathicity. PMID- 10516063 TI - Temporal and spatial analysis of Sin Nombre virus quasispecies in naturally infected rodents. AB - Sin Nombre virus (SNV) is thought to establish a persistent infection in its natural reservoir, the deer mouse (Peromyscus maniculatus), despite a strong host immune response. SNV-specific neutralizing antibodies were routinely detected in deer mice which maintained virus RNA in the blood and lungs. To determine whether viral diversity played a role in SNV persistence and immune escape in deer mice, we measured the prevalence of virus quasispecies in infected rodents over time in a natural setting. Mark-recapture studies provided serial blood samples from naturally infected deer mice, which were sequentially analyzed for SNV diversity. Viral RNA was detected over a period of months in these rodents in the presence of circulating antibodies specific for SNV. Nucleotide and amino acid substitutions were observed in viral clones from all time points analyzed, including changes in the immunodominant domain of glycoprotein 1 and the 3' small segment noncoding region of the genome. Viral RNA was also detected in seven different organs of sacrificed deer mice. Analysis of organ-specific viral clones revealed major disparities in the level of viral diversity between organs, specifically between the spleen (high diversity) and the lung and liver (low diversity). These results demonstrate the ability of SNV to mutate and generate quasispecies in vivo, which may have implications for viral persistence and possible escape from the host immune system. PMID- 10516064 TI - Markers for trans-Golgi membranes and the intermediate compartment localize to induced membranes with distinct replication functions in flavivirus-infected cells. AB - Replication of the flavivirus Kunjin virus is associated with virus-induced membrane structures within the cytoplasm of infected cells; these membranes appear as packets of vesicles associated with the sites of viral RNA synthesis and as convoluted membranes (CM) and paracrystalline arrays (PC) containing the components of the virus-specified protease (E. G. Westaway, J. M. Mackenzie, M. T. Kenney, M. K. Jones, and A. A. Khromykh, J. Virol. 71:6650-6661, 1997). To determine the cellular origins of these membrane structures, we compared the immunolabelling patterns of several cell markers in relation to these sites by immunofluorescence and immunoelectron microscopy. A marker for the trans-Golgi membranes and the trans-Golgi network, 1,4-galactosyltransferase (GalT), was redistributed to large foci in the cytoplasm of Kunjin virus-infected cells, partially coincident with immunofluorescent foci associated with the putative sites of viral RNA synthesis. As determined by immunoelectron microscopy, the induced vesicle packets contained GalT, whereas the CM and PC contained a specific protein marker for the intermediate compartment (ERGIC53). A further indicator of the role of cellular organelles in their biogenesis was the observation that the Golgi apparatus-disrupting agent brefeldin A prevented further development of immunofluorescent foci of induced membranes if added before the end of the latent period but that once formed, these membrane foci were resistant to brefeldin A dispersion. Reticulum membranes emanating from the induced CM and PC were also labelled with the rough endoplasmic reticulum marker anti-protein disulfide isomerase and were obviously redistributed during infection. This is the first report identifying trans-Golgi membranes and the intermediate compartment as the apparent sources of the flavivirus-induced membranes involved in events of replication. PMID- 10516065 TI - Observation of measles virus cell-to-cell spread in astrocytoma cells by using a green fluorescent protein-expressing recombinant virus. AB - A recombinant measles virus (MV) which expresses enhanced green fluorescent protein (EGFP) has been rescued. This virus, MVeGFP, expresses the reporter gene from an additional transcription unit which is located prior to the gene encoding the measles virus nucleocapsid protein. The recombinant virus was used to infect human astrocytoma cells (GCCM). Immunocytochemistry (ICC) together with EGFP autofluorescence showed that EGFP is both an early and very sensitive indicator of cell infection. Cells that were EGFP-positive and ICC-negative were frequently observed. Confocal microscopy was used to indirectly visualize MV infection of GCCM cells and to subsequently follow cell-to-cell spread in real time. These astrocytoma cells have extended processes, which in many cases are intimately associated. The processes appear to have an important role in cell-to-cell spread, and MVeGFP was observed to utilize them in the infection of surrounding cells. Heterogeneity was seen in cell-to-cell spread in what was expected to be a homogeneous monolayer. In tissue culture, physical constraints govern the integrity of the syncytia which are formed upon extensive cell fusion. When around 50 cells were fused, the syncytia rapidly disintegrated and many of the infected cells detached. Residual adherent EGFP-positive cells were seen to either continue to be involved in the infection of surrounding cells or to remain EGFP positive but no longer participate in the transmission of MV infection to neighboring cells. PMID- 10516066 TI - Pathogenesis of experimental rhesus cytomegalovirus infection. AB - Human cytomegalovirus (HCMV) establishes and maintains a lifelong persistence following infection in an immunocompetent host. The determinants of a stable virus-host relationship are poorly defined. A nonhuman primate model for HCMV was used to investigate virological and host parameters of infection in a healthy host. Juvenile rhesus macaques (Macaca mulatta) were inoculated with rhesus cytomegalovirus (RhCMV), either orally or intravenously (i.v. ), and longitudinally necropsied. None of the animals displayed clinical signs of disease, although hematologic abnormalities were observed intermittently in i.v. inoculated animals. RhCMV DNA was detected transiently in the plasma of all animals at 1 to 2 weeks postinfection (wpi) and in multiple tissues beginning at 2 to 4 wpi. Splenic tissue was the only organ positive for RhCMV DNA in all animals. The location of splenic cells expressing RhCMV immediate-early protein 1 (IE1) in i.v. inoculated animals changed following inoculation. At 4 to 5 wpi, most IE1-positive cells were perifollicular, and at 25 wpi, the majority were located within the red pulp. All animals developed anti-RhCMV immunoglobulin M (IgM) antibodies within 1 to 2 wpi and IgG antibodies within 2 to 4 wpi against a limited number of viral proteins. Host reactivity to RhCMV proteins increased in titer (total and neutralizing) and avidity with time. These results demonstrate that while antiviral immune responses were able to protect from disease, they were insufficient to eliminate reservoirs of persistent viral gene expression. PMID- 10516067 TI - Scrapie pathogenesis in subclinically infected B-cell-deficient mice. AB - Prion infections can present without clinical manifestations. B-cell deficiency may be a model for subclinical transmissible spongiform encephalopathy, since it protects mice from disease upon intraperitoneal administration of scrapie prions; however, a proportion of B-cell-deficient mice accumulate protease-resistant prion protein in their brains. Here, we have characterized this subclinical disease. In addition, we have studied the possibility that a neurotoxic factor secreted by B cells may contribute to pathogenesis. PMID- 10516068 TI - A lentivirus packaging system based on alternative RNA transport mechanisms to express helper and gene transfer vector RNAs and its use to study the requirement of accessory proteins for particle formation and gene delivery. AB - A lentivirus-based packaging system was designed to reduce the chance of recombination between helper and gene transfer vector sequences by using the constitutive transport element (CTE) derived from Mason-Pfizer monkey virus for expression of the viral proteins and the Rev-Rev response element (RRE) combination for expression of the gene transfer vector. Using this approach, we evaluated a series of human immunodeficiency virus type 1 packaging constructs that express one or more accessory proteins (Vif, Vpr, and Vpu), in addition to the Gag and Pol proteins, for particle formation and virus stock production for gene transfer. Constructs that also express Vpr or both Vpr and Vpu produced more particles, as measured by a p24 assay, than did plasmids that did not contain these sequences. Transactivation experiments showed that the packaging plasmids that encode Vpr or both Vpr and Vpu also expressed a functional single-exon Tat protein. For these constructs, high-titer virus stocks could be prepared in the absence of a cotransfected Tat-expressing plasmid. Amphotropic-envelope pseudotyped virus stocks prepared with all of the packaging constructs, irrespective of whether any of the accessory proteins were coexpressed, were equally efficient in transducing growth-arrested HeLa cells. The combination/mixed packaging system was compared to systems that were based on either the CTE alone or Rev and RRE for expression of both the packaging plasmid as well as the gene transfer vector. The combination/mixed packaging system was comparable to the other systems for production of virus stocks, suggesting that this design may prove to be safer for the eventual deployment of lentivirus vectors for therapeutic purposes. PMID- 10516069 TI - Construction and preliminary characterization of a library of "lethal" preterminal protein mutant adenoviruses. AB - Adenoviruses containing lethal in-frame insertion mutant alleles of the preterminal protein (pTP) gene were constructed with cell lines that express pTP. Thirty in-frame insertion mutant alleles, including 26 alleles previously characterized as lethal and 4 newly constructed mutant alleles, were introduced into the viral chromosome in place of the wild-type pTP gene. The viruses were tested for ability to form plaques at 37 degrees C in HeLa-pTP cells and at 32 degrees C and 39.5 degrees C in HeLa cells. Two of the newly constructed viruses exhibited temperature sensitivity for plaque formation, one virus did not form plaques in the absence of complementation, seven additional mutants exhibited a greater than 10-fold reduction in plaque formation in the absence of complementation, and another eight mutants exhibited stronger phenotypes than did previously characterized in-frame insertion mutants in the plaque assay. These mutant viruses offer promise for analysis of pTP functions. PMID- 10516070 TI - Quasispecies of TT virus (TTV) with sequence divergence in hypervariable regions of the capsid protein in chronic TTV infection. AB - Three hypervariable regions were identified in a central portion of open reading frame 1 of TT virus DNA, which codes for a putative capsid protein of 770 amino acids. TT virus circulates as quasispecies, with many amino acid substitutions in hypervariable regions, to evade immune surveillance of the hosts and to establish a persistent infection. PMID- 10516071 TI - Mucosal but not parenteral immunization with purified human papillomavirus type 16 virus-like particles induces neutralizing titers of antibodies throughout the estrous cycle of mice. AB - We have recently shown that nasal immunization of anesthetized mice with human papillomavirus type 16 (HPV16) virus-like particles (VLPs) is highly effective at inducing both neutralizing immunoglobulin A (IgA) and IgG in genital secretions, while parenteral immunization induced only neutralizing IgG. Our data also demonstrated that both isotypes are similarly neutralizing according to an in vitro pseudotyped neutralization assay. However, it is known that various amounts of IgA and IgG are produced in genital secretions along the estrous cycle. Therefore, we have investigated how this variation influences the amount of HPV16 neutralizing antibodies induced after immunization with VLPs. We have compared parenteral and nasal protocols of vaccination with daily samplings of genital secretions of mice. Enzyme-linked immunosorbent assay analysis showed that total IgA and IgG inversely varied along the estrous cycle, with the largest amounts of IgA in proestrus-estrus and the largest amount of IgG in diestrus. This resulted in HPV16 neutralizing titers of IgG only being achieved during diestrus upon parenteral immunization. In contrast, nasal vaccination induced neutralizing titers of IgA plus IgG throughout the estrous cycle, as confirmed by in vitro pseudotyped neutralization assays. Our data suggest that mucosal immunization might be more efficient than parenteral immunization at inducing continuous protection of the female genital tract. PMID- 10516072 TI - The kissing-loop motif is a preferred site of 5' leader recombination during replication of SL3-3 murine leukemia viruses in mice. AB - A panel of mouse T-cell lymphomas induced by SL3-3 murine leukemia virus (MLV) and three primer binding site mutants thereof (A. H. Lund, J. Schmidt, A. Luz, A. B. Sorensen, M. Duch, and F. S. Pedersen, J. Virol. 73:6117-6122, 1999) were analyzed for the occurrence of recombination between the exogenous input virus and endogenous MLV-like sequences within the 5' leader region. Evidence of recombination within the region studied was found in 14 of 52 tumors analyzed. Sequence analysis of a approximately 330-bp fragment of 44 chimeric proviruses, encompassing the U5, the primer binding site, and the upstream part of the 5' untranslated region, enabled us to map recombination sites, guided by distinct scattered nucleotide differences. In 30 of 44 analyzed sequences, recombination was mapped to a 33-nucleotide similarity window coinciding with the kissing-loop stem-loop motif implicated in dimerization of the diploid genome. Interestingly, the recombination pattern preference found in replication-competent viruses from T-cell tumors is very similar to the pattern previously reported for retroviral vectors in cell culture experiments. The data therefore sustain the hypothesis that the kissing loop, presumably via a role in RNA dimer formation, constitutes a hot spot for reverse transcriptase-mediated recombination in MLV. PMID- 10516073 TI - Persistent zoonotic infection of a human with simian foamy virus in the absence of an intact orf-2 accessory gene. AB - Although foamy viruses (FVs) are endemic among nonhuman primates, FV infection among humans is rare. Recently, simian foamy virus (SFV) infection was reported in 4 of 231 individuals occupationally exposed to primates (1.8%). Secondary transmission to spouses has not been seen, suggesting that while FV is readily zoonotic, humans may represent dead-end hosts. Among different simian species, SFV demonstrates significant sequence diversity within the U3 region of the long terminal repeat (LTR) and 3' accessory open reading frames (ORFs). To examine if persistent human SFV infection and apparent lack of secondary transmission are associated with genetic adaptations in FV regulatory regions, we conducted sequence analysis of the LTR, internal promoter, ORF-1, and ORF-2 on a tissue culture isolate and peripheral blood mononuclear cell samples from a human infected with SFV of African green monkey origin (SFV-3). Compared to the prototype SFV-3 sequence, the LTR, internal promoter, and FV transactivator (ORF 1) showed sequence conservation, suggesting that FV zoonosis is not dependent on host-specific adaptation to these transcriptionally important regions. However, ORF-2 contains a number of deleterious mutations predicted to result in premature termination of protein synthesis. ORF-2 codes in part for the 60-kDa Bet fusion protein, proposed to be involved in the establishment of persistent cellular SFV infections. These results suggest that persistent human infection by SFV and reduced transmissibility may be influenced by the absence of a functional ORF-2. PMID- 10516074 TI - Molecular characterization of a porcine enteric calicivirus genetically related to Sapporo-like human caliciviruses. AB - Porcine enteric calicivirus (PEC) is associated with diarrhea in pigs, and to date it is the only cultivable enteric calicivirus (tissue culture-adapted [TC] PEC/Cowden). Based on sequence analysis of cDNA clones and reverse transcription PCR products, TC PEC/Cowden has an RNA genome of 7,320 bp, excluding its 3' poly(A)(+) tail. The genome is organized in two open reading frames (ORFs), similar to the organizations of the human Sapporo-like viruses (SLVs) and the lagoviruses. ORF1 encodes the polyprotein that is fused to and contiguous with the capsid protein. ORF2 at the 3' end encodes a small basic protein of 164 amino acids. Among caliciviruses, PEC has the highest amino acid sequence identities in the putative RNA polymerase (66%), 2C helicase (49.6%), 3C-like protease (43.7%), and capsid (39%) regions with the SLVs, indicating that PEC is genetically most closely related to the SLVs. The complete RNA genome of wild-type (WT) PEC/Cowden was also sequenced. Sequence comparisons revealed that the WT and TC PEC/Cowden have 100% nucleotide sequence identities in the 5' terminus, 2C helicase, ORF2, and the 3' nontranslated region. TC PEC/Cowden has one silent mutation in its protease, two amino acid changes and a silent mutation in its RNA polymerase, and five nucleotide substitutions in its capsid that result in one distant and three clustered amino acid changes and a silent mutation. These substitutions may be associated with adaptation of TC PEC/Cowden to cell culture. The cultivable PEC should be a useful model for studies of the pathogenesis, replication, and possible rescue of uncultivable human enteric caliciviruses. PMID- 10516075 TI - Association of murine leukemia virus pol with virions, independent of Gag-Pol expression. AB - During the replication cycle of murine leukemia virus (MLV), Pol is normally synthesized as part of a Gag-Pol fusion protein. In this study, the ability of free MLV Pol to be incorporated into virions was examined. When MLV Gag and MLV Pol were coexpressed from separate plasmids in cells, reverse transcriptase (RT) activity associated with Gag core particles at a slightly lower level than did RT activity generated from wild-type Gag-Pol expression. Particles produced in this manner were somewhat less infectious than those produced with wild-type Gag-Pol. A smaller amount of MLV Pol also associated with heterologous human immunodeficiency virus type 1 Gag cores. PMID- 10516076 TI - Minimal incidence of serum antibodies reactive with intact primary isolate virions in human immunodeficiency virus type 1-infected individuals. AB - Immunoglobulin G reactive with primary isolate virions was detected in 36% of serum samples from individuals infected with human immunodeficiency virus type 1. Of these individuals, serum samples from only 7% captured significant quantities of virus. Virion-specific antibody correlated with CD4 counts and, of more significance, primary isolate neutralization. Further dissection of this response should lead to the identification of antibodies and antigenic epitopes for vaccine purposes. PMID- 10516077 TI - Proliferation response to interleukin-2 and Jak/Stat activation of T cells immortalized by human T-cell lymphotropic virus type 1 is independent of open reading frame I expression. AB - Human T-cell lymphotropic virus type 1 (HTLV-1), a complex retrovirus, encodes a hydrophobic 12-kD protein from pX open reading frame (ORF) I that localizes to cellular endomembranes and contains four minimal SH3 binding motifs (PXXP). We have demonstrated the importance of ORF I expression in the establishment of infection and hypothesize that p12(I) has a role in T-cell activation. In this study, we tested interleukin-2 (IL-2) receptor expression, IL-2-mediated proliferation, and Jak/Stat activation in T-cell lines immortalized with either wild-type or ORF I mutant clones of HTLV-1. All cell lines exhibited typical patterns of T-cell markers and maintained mutation fidelity. No significant differences between cell lines were observed in IL-2 receptor chain (alpha, beta, or gamma(c)) expression, in IL-2-mediated proliferation, or in IL-2-induced phosphorylated forms of Stat3, Stat5, Jak1, or Jak3. The expression of ORF I is more likely to play a role in early HTLV-1 infection, such as in the activation of quiescent T cells in vivo. PMID- 10516079 TI - Preferential associations of alleles of three distinct genes argue for the existence of two prototype variants of human herpesvirus 7. AB - We had previously described six distinct alleles of the glycoprotein B (gB) gene of human herpesvirus 7 (HHV-7). The genetic changes corresponding to these alleles did not affect gB gene transcription or translation in in vitro assays. The study of distinct HHV-7-positive human samples showed preferential associations of some gB alleles with some alleles of two other genes, distantly located on the HHV-7 genome, coding for the phosphoprotein p100 (p100) and the major capsid protein (MCP). Two allele combinations, corresponding to 44 and 31% of the samples studied, respectively, were interpreted as the genetic signatures of two major prototype HHV-7 variants. PMID- 10516078 TI - Requirement for CD40 ligand, CD4(+) T cells, and B cells in an infectious mononucleosis-like syndrome. AB - Respiratory challenge with the murine gammaherpesvirus 68 (gammaHV-68) results in productive infection of the lung, the establishment of latency in B lymphocytes and other cell types, transient splenomegaly, and prolonged clonal expansion of activated CD8(+) CD62L(lo) T cells, particularly a Vbeta4(+) CD8(+) population that is found in mice with different major histocompatibility complex (MHC) haplotypes. Aspects of the CD8(+)-T-cell response are substantially modified in mice that lack B cells, CD4(+) T cells, or the CD40 ligand (CD40L). The B-cell deficient mice show no increase in Vbeta4(+) CD8(+) T cells. Similar abrogation of the Vbeta4(+) CD8(+) response is seen following antibody-mediated depletion of the CD4(+) subset, through the numbers of CD8(+) CD62L(lo) cells are still significantly elevated. Virus-specific CD4(+)-T-cell frequencies are minimal in the CD40L(-/-) mice, and the Vbeta4(+) CD8(+) population remains unexpanded. Apparently B-cell-CD4(+)-T-cell interactions play a part in the gammaHV-68 induction of both splenomegaly and non-MHC-restricted Vbeta4(+) CD8(+)-T-cell expansion. PMID- 10516080 TI - The Y-S-L-I tyrosine-based motif in the cytoplasmic domain of the human T-cell leukemia virus type 1 envelope is essential for cell-to-cell transmission. AB - The human T-cell leukemia virus type 1 (HTLV-1) transmembrane glycoprotein has a 24-amino-acid cytoplasmic domain whose function in the viral life cycle is poorly understood. We introduced premature-stop mutations and 18 single-amino-acid substitutions into this domain and studied their effects on cell-to-cell transmission of the virus. The results show that the cytoplasmic domain is absolutely required for cell-to-cell transmission of HTLV-1, through amino acids which cluster in a Y-S-L-I tyrosine-based motif. The transmission defect in two motif mutants did not result from a defect in glycoprotein incorporation or fusion. It appears that the Y-S-L-I tyrosine-based motif of the HTLV-1 glycoprotein cytoplasmic domain has multiple functions, including involvement in virus transmission at a postfusion step. PMID- 10516081 TI - rab5 GTPase regulates adenovirus endocytosis. AB - Adenovirus interaction with alphav integrins is important for virus entry. We have examined the effects of adenovirus attachment on intracellular signaling in HeLa cells, with an emphasis on pathways known to be activated following integrin interaction with other ligands. We found no evidence for [Ca(2+)](c)-mediated signaling or for tyrosine phosphorylation of pp125(FAK), p130(CAS), and paxillin. However, adenovirus attachment is known to activate phosphatidylinositol-3 kinase, which in turn may regulate endocytosis via rab5 GTPase. We found that adenovirus uptake was increased by overexpression of wild-type rab5 and decreased by dominant-negative rab5. These results indicate a role for rab5 in adenovirus entry. PMID- 10516082 TI - Herpes simplex virus type 1 serum neutralizing antibody titers increase during latency in rabbits latently infected with latency-associated transcript (LAT) positive but not LAT-negative viruses. AB - The herpes simplex virus type 1 (HSV-1) latency-associated transcript (LAT) gene is essential for efficient spontaneous reactivation in the rabbit ocular model of HSV-1 latency and reactivation. LAT is also the only viral gene abundantly expressed during latency. Rabbits were ocularly infected with the wild-type HSV-1 strain McKrae or the McKrae-derived LAT null mutant dLAT2903. Serum neutralizing antibody titers were determined at various times during acute and latent infection. The neutralizing antibody titers induced by both viruses increased and were similar throughout the first 45 days after infection (P > 0.05). However, by day 59 postinfection (approximately 31 to 45 days after latency had been established), the neutralizing antibody titers induced by wild-type virus and dLAT2903 diverged significantly (P = 0.0005). The dLAT2903-induced neutralizing antibody titers decreased, while the wild-type virus-induced neutralizing antibody titers continued to increase. A rescuant of dLAT2903, in which spontaneous reactivation was fully restored, induced wild-type neutralizing antibody levels on day 59 postinfection. A second LAT mutant with impaired spontaneous reactivation had neutralizing antibody levels comparable to those of dLAT2903. In contrast to the results obtained in rabbits, in mice, neutralizing antibody titers did not increase over time during latency with any of the viruses. Since LAT is expressed in both rabbits and mice during latency, the difference in neutralizing antibody titers between these animals is unlikely to be due to expression of a LAT protein during latency. In contrast, LAT-positive (LAT(+)), but not LAT-negative (LAT(-)), viruses undergo efficient spontaneous reactivation in rabbits, while neither LAT(+) nor LAT(-) viruses undergo efficient spontaneous reactivation in mice. Thus, the increase in neutralizing antibody titers in rabbits latently infected with LAT(+) viruses may have been due to continued restimulation of the immune system by spontaneously reactivating virus. PMID- 10516083 TI - The trophoblastic epithelial barrier is not infected in full-term placentae of human immunodeficiency virus-seropositive mothers undergoing antiretroviral therapy. AB - To study the mechanism of the placental barrier function, we examined 10 matched samples of term placentae, cord blood, and maternal blood obtained at delivery from human immunodeficiency virus (HIV)-infected mothers with children diagnosed as HIV negative in Sweden. All placentae were histologically normal, and immunochemistry for HIV type 1 p24 and gp120 antigens was negative. Highly purified trophoblasts (93 to 99% purity) were negative for HIV DNA and RNA, indicating that the trophoblasts were uninfected. Although HIV DNA was detected in placenta-derived T lymphocytes and monocytes, microsatellite analysis showed that these cells were a mixture of maternal and fetal cells. Our study indicates that the placental barrier, i.e., the trophoblastic layer, is not HIV infected and, consequently, HIV infection of the fetus is likely to occur through other routes, such as breaks in the placental barrier. PMID- 10516084 TI - Rescue of influenza A virus from recombinant DNA. AB - We have rescued influenza A virus by transfection of 12 plasmids into Vero cells. The eight individual negative-sense genomic viral RNAs were transcribed from plasmids containing human RNA polymerase I promoter and hepatitis delta virus ribozyme sequences. The three influenza virus polymerase proteins and the nucleoprotein were expressed from protein expression plasmids. This plasmid-based reverse genetics technique facilitates the generation of recombinant influenza viruses containing specific mutations in their genes. PMID- 10516086 TI - TT virus--part of the normal human flora? PMID- 10516085 TI - Inhibition of cell-free human T-cell leukemia virus type 1 infection at a postbinding step by the synthetic peptide derived from an ectodomain of the gp21 transmembrane glycoprotein. AB - To investigate the roles of human T-cell leukemia virus type 1 (HTLV-1) envelope (Env) proteins gp46 and gp21 in the early steps of infection, the effects of the 23 synthetic peptides covering the entire Env proteins on transmission of cell free HTLV-1 were examined by PCR and by the plaque assay using a pseudotype of vesicular stomatis virus (VSV) bearing the Env of HTLV-1 [VSV(HTLV-1)]. The synthetic peptide corresponding to amino acids 400 to 429 of the gp21 Env protein (gp21 peptide 400-429, Cys-Arg-Phe-Pro-Asn-Ile-Thr-Asn-Ser-His-Val-Pro-Ile-Leu Gln-Glu-Arg-P ro-Pro-Leu-Glu-Asn-Arg-Val-Leu-Thr-Gly-Trp-Gly-Leu) strongly inhibited infection of cell-free HTLV-1. By using the mutant peptide, Asn407, Ser408, and Leu413, -419, -424, and -429 were confirmed to be important amino acids for neutralizing activity of the gp21 peptide 400-429. Addition of this peptide before or during adsorption of HTLV-1 at 4 degrees C did not affect its entry. However, HTLV-1 infection was inhibited about 60% when the gp21 peptide 400-429 was added even 30 min after adsorption of HTLV-1 to cells, indicating that the amino acid sequence 400 to 429 on the gp21 Env protein plays an important role at the postbinding step of HTLV-1 infection. In contrast, a monoclonal antibody reported to recognize the gp46 191-196 peptide inhibited the infection of HTLV-1 at the binding step. PMID- 10516088 TI - Intestinal absorption of water-soluble vitamins focus on "Molecular mechanism of the intestinal biotin transport process". PMID- 10516089 TI - Molecular mechanism of the intestinal biotin transport process. AB - Previous studies have characterized different aspects of the cellular/membrane mechanism and regulation of the intestinal uptake process of the water-soluble vitamin biotin. Little, however, is known about the molecular mechanisms of the uptake process. In this study, we have identified a cDNA from rat small intestine that appears to be involved in biotin transport. The open reading frame of this cloned cDNA consisted of 1,905 bases and was identical to that identified for the vitamin transporter in placental tissue. Significant heterogeneity, however, was found in the 5' untranslated region of this clone, with three distinct variants (II, III, IV) being identified in the small intestine; the placental variant (variant I), however, was not present in the small gut. Variant II was found to be the predominant form expressed in the rat small and large intestines. Functional identity of the cloned intestinal cDNA was confirmed by stable expression in COS-7 cells, which showed a four- to fivefold increase in biotin uptake in transfected COS-7 cells compared with controls. The induced biotin uptake in transfected COS-7 cells was found to be 1) Na(+) dependent, 2) saturable as a function of concentration with an apparent K(m) of 8. 77 microM and a V(max) of 779.7 pmol. mg protein(-1). 3 min(-1), and 3) inhibited by unlabeled biotin and pantothenic acid and their structural analogs. The distribution of complementary mRNA transcripts of the cloned cDNA along the vertical and longitudinal axes of the intestinal tract was also determined. Results of this study describe the molecular characteristics of the intestinal biotin absorption process and report the identification of a cDNA that encodes a Na(+)-dependent biotin uptake carrier that appears to exist in the form of multiple variants. PMID- 10516090 TI - The astrocytic endothelin system: toward solving a mystery focus on "distinct pharmacological properties of ET-1 and ET-3 on astroglial gap junctions and Ca(2+) signaling". PMID- 10516092 TI - Transforming growth factor-beta1 modulates the expression of vascular endothelial growth factor by osteoblasts. AB - Angiogenesis is essential to both normal and pathological bone physiology. Vascular endothelial growth factor (VEGF) has been implicated in angiogenesis, whereas transforming growth factor-beta1 (TGF-beta1) modulates bone differentiation, matrix formation, and cytokine expression. The purpose of this study was to investigate the relationship between TGF-beta1 and VEGF expression in osteoblasts and osteoblast-like cells. Northern blot analysis revealed an early peak of VEGF mRNA (6-fold at 3 h) in fetal rat calvarial cells and MC3T3-E1 osteoblast-like cells after stimulation with TGF-beta1 (2.5 ng/ml). The stability of VEGF mRNA in MC3T3-E1 cells was not increased after TGF-beta1 treatment. Actinomycin D inhibited the TGF-beta1-induced peak in VEGF mRNA, whereas cycloheximide did not. Blockade of TGF-beta1 signal transduction via a dominant negative receptor II adenovirus significantly decreased TGF-beta1 induction of VEGF mRNA. Additionally, TGF-beta1 induced a dose-dependent increase in VEGF protein expression by MC3T3-E1 cells (P < 0.01). Dexamethasone similarly inhibited VEGF protein expression. Both TGF-beta1 mRNA and VEGF mRNA were concurrently present in rat membranous bone, and both followed similar patterns of expression during rat mandibular fracture healing (mRNA and protein). In summary, TGF-beta1-induced VEGF expression by osteoblasts and osteoblast-like cells is a dose-dependent event that may be intimately related to bone development and fracture healing. PMID- 10516091 TI - Distinct pharmacological properties of ET-1 and ET-3 on astroglial gap junctions and Ca(2+) signaling. AB - Astrocytes represent a major target for endothelins (ETs), a family of peptides that have potent and multiple effects on signal transduction pathways and can be released by several cell types in the brain. In the present study we have investigated the involvement of different ET receptor subtypes on intercellular dye diffusion, intracellular Ca(2+) homeostasis, and intercellular Ca(2+) signaling in cultured rat astrocytes from hippocampus and striatum. Depending on the ET concentration and the receptor involved, ET-1- and ET-3-induced intracellular Ca(2+) increases with different response patterns. Both ET-1 and ET 3 are powerful inhibitors of gap junctional permeability and intercellular Ca(2+) signaling. The nonselective ET receptor agonist sarafotoxin S6b and the ET(B) receptor-selective agonist IRL 1620 mimicked these inhibitions. The ET-3 effects were only marginally affected by an ET(A) receptor antagonist but completely blocked by an ET(B) receptor antagonist. However, the ET-1-induced inhibition of gap junctional dye transfer and intercellular Ca(2+) signaling was only marginally blocked by ET(A) or ET(B) receptor-selective antagonists but fully prevented when these antagonists were applied together. The ET-induced inhibition of gap junction permeability and intercellular Ca(2+) signaling indicates that important changes in the function of astroglial communication might occur when the level of ETs in the brain is increased. PMID- 10516093 TI - Expression of an inwardly rectifying K(+) channel, Kir4.1, in satellite cells of rat cochlear ganglia. AB - Satellite cells are glial cells wrapped around somata of sensory and autonomic ganglion neurons. Neither their functional roles nor electrical properties have been fully clarified so far. Using immunohistochemistry, we found that inwardly rectifying K(+) channel subunit Kir4.1 (also called Kir1.2 or K(AB)-2) was expressed prominently in the satellite cells of cochlear ganglia. The Kir4.1 immunoreactivity was localized specifically at the myelin sheaths of satellite cells wrapping the somata of the ganglion neurons. Developmental expression of Kir4.1 in satellite cells paralleled development of the action potential in the auditory nerve. These results suggest that this channel in satellite cells may be responsible for the regulation of K(+) extruded from the ganglion neurons during excitation. PMID- 10516094 TI - Transport of thiamine in human intestine: mechanism and regulation in intestinal epithelial cell model Caco-2. AB - The present study examined the intestinal uptake of thiamine (vitamin B(1)) using the human-derived intestinal epithelial cells Caco-2 as an in vitro model system. Thiamine uptake was found to be 1) temperature and energy dependent and occurred with minimal metabolic alteration; 2) pH sensitive; 3) Na(+) independent; 4) saturable as a function of concentration with an apparent Michaelis-Menten constant of 3.18 +/- 0.56 microM and maximal velocity of 13.37 +/- 0.94 pmol. mg protein(-1). 3 min(-1); 5) inhibited by the thiamine structural analogs amprolium and oxythiamine, but not by unrelated organic cations tetraethylammonium, N methylnicotinamide, and choline; and 6) inhibited in a competitive manner by amiloride with an inhibition constant of 0.2 mM. The role of specific protein kinase-mediated pathways in the regulation of thiamine uptake by Caco-2 cells was also examined using specific modulators of these pathways. The results showed possible involvement of a Ca(2+)/calmodulin (CaM)-mediated pathway in the regulation of thiamine uptake. No role for protein kinase C- and protein tyrosine kinase-mediated pathways in the regulation of thiamine uptake was evident. These results demonstrate the involvement of a carrier-mediated system for thiamine uptake by Caco-2 intestinal epithelial cells. This system is Na(+) independent and is different from the transport systems of organic cations. Furthermore, a CaM-mediated pathway appears to play a role in regulating thiamine uptake in these cells. PMID- 10516095 TI - MAP kinase cascade is required for p27 downregulation and S phase entry in fibroblasts and epithelial cells. AB - The present report delineates the critical pathway in the G(1) phase involved in downregulation of p27(Kip1), a cyclin-dependent kinase inhibitor, which plays a pivotal role in controlling entry into the S phase of the cell cycle. In resting CCL39 fibroblasts and IEC-6 intestinal epithelial cells, protein levels of p27(Kip1) were elevated but dramatically decreased on serum stimulation, along with hyperphosphorylation of pRb and increased CDK2 activity. In both cell types, expression of ras resulted in an increase of basal and serum-stimulated E2F dependent transcriptional activity and a reduction in p27(Kip1) protein levels as well. The role of the mitogen-activated protein (MAP) kinase cascade in p27(Kip1) reduction and S phase reentry was reinforced by the blockades of serum-induced E2F-dependent transcriptional activity and p27(Kip1) downregulation with the MKK 1/2 inhibitor PD-98059. In both cell lines, downregulation of p27(Kip1) was associated with a repression of its synthesis, an event mediated by the p42/p44 MAP kinase pathway. Using an antisense approach, we demonstrated that p27(Kip1) may control cell cycle exit in both cell types. These data indicate that activation of the MAP kinase cascade is required for S phase entry and p27(Kip1) downregulation in fibroblasts and epithelial cells. PMID- 10516096 TI - Ceramide triggers intracellular calcium release via the IP(3) receptor in Xenopus laevis oocytes. AB - Ceramide, a product of sphingomyelin turnover, is a lipid second messenger that mediates diverse signaling pathways, including those leading to cell cycle arrest and differentiation. The mechanism(s) by which ceramide signals downstream events have not been fully elucidated. Here we show that, in Xenopus laevis oocytes, ceramide-induced maturation is associated with the release of intracellular calcium stores. Ceramide caused a dose-dependent elevation in the second messenger inositol 1,4,5-trisphosphate (IP(3)) via activation of G(q/11)alpha and phospholipase C-betaX. Elevation of IP(3), in turn, activated the IP(3) receptor calcium release channel on the endoplasmic reticulum, resulting in a rise in cytoplasmic calcium. Thus our study demonstrates that cross talk between the ceramide and phosphoinositide signaling pathways modulates intracellular calcium homeostasis. PMID- 10516097 TI - Hypoxia modulates nitric oxide-induced regulation of NMDA receptor currents and neuronal cell death. AB - Nitric oxide (NO) released from a new chemical class of donors enhances N-methyl D-aspartate (NMDA) channel activity. Using whole cell and single-channel patch clamp techniques, we have shown that (Z)-1-[N-(3-ammoniopropyl)-N-(n propyl)amino]-NO (PAPA-NO) and diethylamine NO, commonly termed NONOates, potentiate the glutamate-mediated response of recombinant rat NMDA receptors (NR1/NR2A) expressed in HEK-293 cells. The overall effect is an increase in both peak and steady-state whole cell currents induced by glutamate. Single-channel studies demonstrate a significant increase in open probability but no change in the mean single-channel open time or mean channel conductance. Reduction in oxygen levels increased and prolonged the PAPA-NO-induced change in both peak and steady-state glutamate currents in transfected HEK cells. PAPA-NO also enhanced cell death in primary cultures of rodent cortical neurons deprived of oxygen and glucose. This potentiation of neuronal injury was blocked by MK-801, indicating a critical involvement of NMDA receptor activation. The NO-induced increase in NMDA channel activity as well as NMDA receptor-mediated cell death provide firm evidence that NO modulates the NMDA channel in a manner consistent with both a physiological role under normoxic conditions and a pathophysiological role under hypoxic conditions. PMID- 10516098 TI - Inhibition of Na(+)-K(+)-2Cl(-) cotransport by mercury. AB - Mercury alters the function of proteins by reacting with cysteinyl sulfhydryl (SH(-)) groups. The inorganic form (Hg(2+)) is toxic to epithelial tissues and interacts with various transport proteins including the Na(+) pump and Cl(-) channels. In this study, we determined whether the Na(+)-K(+)-Cl(-) cotransporter type 1 (NKCC1), a major ion pathway in secretory tissues, is also affected by mercurial substrates. To characterize the interaction, we measured the effect of Hg(2+) on ion transport by the secretory shark and human cotransporters expressed in HEK-293 cells. Our studies show that Hg(2+) inhibits Na(+)-K(+)-Cl(-) cotransport, with inhibitor constant (K(i)) values of 25 microM for the shark carrier (sNKCC1) and 43 microM for the human carrier. In further studies, we took advantage of species differences in Hg(2+) affinity to identify residues involved in the interaction. An analysis of human-shark chimeras and of an sNKCC1 mutant (Cys-697-->Leu) reveals that transmembrane domain 11 plays an essential role in Hg(2+) binding. We also show that modification of additional SH(-) groups by thiol-reacting compounds brings about inhibition and that the binding sites are not exposed on the extracellular face of the membrane. PMID- 10516099 TI - Cloning, expression, and characterization of the trout cardiac Na(+)/Ca(2+) exchanger. AB - Isoform 1 of the cardiac Na(+)/Ca(2+) exchanger (NCX1) is an important regulator of cytosolic Ca(2+) concentration in contraction and relaxation. Studies with trout heart sarcolemmal vesicles have shown NCX to have a high level of activity at 7 degrees C, and this unique property is likely due to differences in protein structure. In this study, we describe the cloning of an NCX (NCX-TR1) from a Lambda ZAP II cDNA library constructed from rainbow trout (Oncorhynchus mykiss) heart RNA. The NCX-TR1 cDNA has an open reading frame that codes for a protein of 968 amino acids with a deduced molecular mass of 108 kDa. A hydropathy plot indicates the protein contains 12 hydrophobic segments (of which the first is predicted to be a cleaved leader peptide) and a large cytoplasmic loop. By analogy to NCX1, NCX-TR1 is predicted to have nine transmembrane segments. The sequences demonstrated to be the exchanger inhibitory peptide site and the regulatory Ca(2+) binding site in the cytoplasmic loop of mammalian NCX1 are almost completely conserved in NCX-TR1. NCX-TR1 cRNA was injected into Xenopus oocytes, and after 3-4 days currents were measured by the giant excised patch technique. NCX-TR1 currents measured at approximately 23 degrees C demonstrated Na(+)-dependent inactivation and Ca(2+)-dependent activation in a manner qualitatively similar to that for NCX1 currents. PMID- 10516100 TI - Skeletal muscle contractile activity in vitro stimulates mitogen-activated protein kinase signaling. AB - Physical exercise is a potent stimulator of mitogen-activated protein (MAP) kinase signaling. To determine if this activation is secondary to systemic responses to exercise or due to muscle contractile activity per se, an isolated muscle preparation was developed. Contractile activity in vitro significantly increased p44(MAPK) and p42(MAPK) phosphorylation by 2.9- and 2.4-fold, respectively. Contraction-stimulated MAP kinase phosphorylation was not decreased in the presence of D-tubocurarine or calphostin C, suggesting that neither neurotransmitter release nor diacylglycerol-sensitive protein kinase C mediates the contraction-induced activation of this signaling cascade. However, PD-98059, an inhibitor of MAP kinase kinase (MEK), inhibited the contraction-induced increases in MAP kinase phosphorylation. PD-98059 did not alter contraction induced increases in glucose uptake or glycogen synthase activity, demonstrating that MAP kinase signaling is not necessary for these important metabolic effects of contractile activity in skeletal muscle. These data suggest that contractile activity of the skeletal muscle fibers per se, and not responses to neurotransmitter release, hormones, or other systemic factors, is responsible for the stimulation of MAP kinase signaling with physical exercise. PMID- 10516101 TI - Kinetics of cyclocreatine and Na(+) cotransport in human breast cancer cells: mechanism of activity. AB - The growth-inhibitory effect of cyclocreatine (CCr) and the kinetics of CCr and Na(+) cotransport were investigated in MCF7 human breast cancer cells and its adriamycin-resistant subline with use of (31)P- and (23)Na-NMR spectroscopy. The growth-inhibitory effect in the resistant line occurred at a lower CCr concentration and was more pronounced than in the wild-type line. This correlated with an approximately 10-fold higher affinity of CCr to the transporter in the resistant line. The passive diffusion coefficient of CCr was also higher in the resistant line by three- to fourfold. The transport of CCr was accompanied by a rapid increase in intracellular Na(+). This increase was found to depend on the rate of CCr transport and varied differently with CCr concentration in the two cell lines. It is proposed that the cotransport of CCr and Na(+) followed by increased Na(+) concentration, together with the accumulation of the highly charged phosphocyclocreatine, are responsible for cell swelling and death. PMID- 10516102 TI - Novel role of the Ca(2+)-ATPase in NMDA-induced intracellular acidification. AB - The mechanism involved in N-methyl-D-glucamine (NMDA)-induced Ca(2+)-dependent intracellular acidosis is not clear. In this study, we investigated in detail several possible mechanisms using cultured rat cerebellar granule cells and microfluorometry [fura 2-AM or 2', 7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein AM]. When 100 microM NMDA or 40 mM KCl was added, a marked increase in the intracellular Ca(2+) concentration ([Ca(2+)](i)) and a decrease in the intracellular pH were seen. Acidosis was completely prevented by the use of Ca(2+)-free medium or 1,2-bis(2-aminophenoxy)ethane-N,N,N', N'-tetraacetic acid AM, suggesting that it resulted from an influx of extracellular Ca(2+). The following four mechanisms that could conceivably have been involved were excluded: 1) Ca(2+) displacement of intracellular H(+) from common binding sites; 2) activation of an acid loader or inhibition of acid extruders; 3) overproduction of CO(2) or lactate; and 4) collapse of the mitochondrial membrane potential due to Ca(2+) uptake, resulting in inhibition of cytosolic H(+) uptake. However, NMDA/KCl-induced acidosis was largely prevented by glycolytic inhibitors (iodoacetate or deoxyglucose in glucose-free medium) or by inhibitors of the Ca(2+)-ATPase (i.e., Ca(2+)/H(+) exchanger), including La(3+), orthovanadate, eosin B, or an extracellular pH of 8.5. Our results therefore suggest that Ca(2+) ATPase is involved in NMDA-induced intracellular acidosis in granule cells. We also provide new evidence that NMDA-evoked intracellular acidosis probably serves as a negative feedback signal, probably with the acidification itself inhibiting the NMDA-induced [Ca(2+)](i) increase. PMID- 10516103 TI - Phosphorylation-dependent stimulation of prostanoid synthesis by nigericin in cerebral endothelial cells. AB - Nigericin decreases intracellular pH (pH(i)) and stimulates prostanoid (PG) synthesis in endothelial cells from cerebral microvessels of newborn pigs. Nigericin-induced PG production was abolished by protein tyrosine kinase (PTK) inhibitors and amplified by phorbol 12-myristate 13-acetate (PMA) or protein tyrosine phosphatase (PTP) inhibitors. Nigericin-induced PG production in PMA primed cells was potentiated by PTP inhibitors and abrogated by PTK inhibitors. Phospholipase A(2) (PLA(2)) activity was stimulated by nigericin in a phosphorylation-dependent manner. Nigericin's effects on PG production and PLA(2) activity were reproduced by ionomycin, which activates cytosolic PLA(2) (cPLA(2)). cPLA(2) was immunodetected in endothelial cell lysates. We found no evidence that nigericin's effects are mediated via mitogen-activated protein (MAP) kinase [extracellularly regulated kinase 1 (ERK1) and ERK2] activation: although nigericin stimulated detergent-soluble MAP kinase, its effects were not amplified by PMA or PTP inhibitors. Phosphorylation-dependent stimulation of PG synthesis was also observed when pH(i) was decreased by sodium propionate or a high level of CO(2). Altogether, our data indicate that nigericin and decreased pH(i) stimulate PG synthesis by a protein phosphorylation-dependent mechanism involving cross talk between pathways mediated by PTK and PTP and by protein kinase C; cPLA(2) appears to be a key enzyme affected by nigericin and decreased pH(i). PMID- 10516104 TI - Desensitization to ANG II in guinea pig ileum depends on membrane repolarization: role of maxi-K(+) channel. AB - Desensitization of ANG II tonic contractile response of the guinea pig ileum is related to membrane repolarization determined by Ca(2+)-activated K(+) (maxi K(+)) channel opening. ANG II-stimulated depolarized myocytes presented sustained activation of maxi-K(+) channels, characterized by reduction from 415 to 12 ms of the closed time constant. ANG II desensitization was prevented by 100 nM iberiotoxin, being reversible within 30 min. Depolarization by KCl, higher than 4 mM, impaired desensitization, suggesting that the membrane potential must attain a threshold to counteract the repolarization induced by maxi-K(+) channel opening. Once this value is attained, there is no time dependency because the desensitization process was shut off by addition of KCl along the time course of the tonic response. In contrast, the sustained ACh tonic component was not altered by these maneuvers. We conclude that desensitization of the ANG II tonic component is foremost due to the opening of maxi-K(+) channels, leading to membrane repolarization, thus closing the voltage-dependent Ca(2+) channels responsible for the Ca(2+) influx that sustains the tonic component in this muscle. PMID- 10516105 TI - Endothelins activate Ca(2+)-gated K(+) channels via endothelin B receptors in CD 1 mouse erythrocytes. AB - Cell dehydration mediated by Ca(2+)-activated K(+) channels plays an important role in the pathogenesis of sickle cell disease. CD-1 mouse erythrocytes possess a Ca(2+)-activated K(+) channel (Gardos channel) with maximal velocity (V(max)) of 0.154 +/- 0.02 mmol. l cells(-1). min(-1) and an affinity constant (K(0.5)) for Ca(2+) of 286 +/- 83 nM in the presence of A-23187. Cells pretreated with 500 nM endothelin-1 (ET-1) increased their V(max) by 88 +/- 9% (n = 8) and decreased their K(0.5) for Ca(2+) to 139 +/- 63 nM (P < 0.05; n = 4). Activation of the Gardos channel resulted in an EC(50) of 75 +/- 20 nM for ET-1 and 374 +/- 97 nM for ET-3. Analysis of the affinity of unlabeled ET-1 for its receptor showed two classes of binding sites with apparent dissociation constants of 167 +/- 51 and 785 +/- 143 nM and with capacity of binding sites of 298 +/- 38 and 1,568 +/- 211 sites/cell, respectively. The Gardos channel was activated by the endothelin B (ET(B)) receptor agonist IRL 1620 and inhibited by BQ-788, demonstrating the involvement of ET(B) receptors. Calphostin C inhibited 73% of ET-1-induced Gardos activation and 84% of the ET-1-induced membrane protein kinase C activity. Thus endothelins regulate erythrocyte Gardos channels via ET(B) receptors and a calphostin-sensitive mechanism. PMID- 10516106 TI - Maitotoxin activates a nonselective cation channel and a P2Z/P2X(7)-like cytolytic pore in human skin fibroblasts. AB - Maitotoxin (MTX), a potent cytolytic agent, activates Ca(2+) entry via nonselective cation channels in virtually all types of cells. The identity of the channels involved and the biochemical events leading to cell lysis remain unknown. In the present study, the effect of MTX on plasmalemmal permeability of human skin fibroblasts was examined. MTX produced a time- and concentration dependent increase in cytosolic free Ca(2+) concentration that depended on extracellular Ca(2+) and was relatively insensitive to blockade by extracellular lanthanides. MTX also produced a time- and concentration-dependent increase in plasmalemma permeability to larger molecules as indicated by 1) uptake of ethidium (314 Da), 2) uptake of YO-PRO-1 (375 Da), 3) release of intracellular fura 2 (636 Da), 4) uptake of POPO-3 (715 Da), and, ultimately, 5) release of lactate dehydrogenase (relative molecular weight of 140,000). At the single cell level, uptake of YO-PRO-1 correlated in time with the appearance of large MTX induced membrane currents carried by the organic cation, N-methyl-D-glucamine (167 Da). Thus MTX initially activates Ca(2+)-permeable cation channels and later induces the formation of large pores. These effects of MTX on plasmalemmal permeability are similar to those seen on activation of P2Z/P2X(7) receptors in a variety of cell types, raising the intriguing possibility that MTX and P2Z/P2X(7) receptor stimulation activate a common cytolytic pore. PMID- 10516107 TI - Maitotoxin and P2Z/P2X(7) purinergic receptor stimulation activate a common cytolytic pore. AB - The effects of maitotoxin (MTX) on plasmalemma permeability are similar to those caused by stimulation of P2Z/P2X(7) ionotropic receptors, suggesting that 1) MTX directly activates P2Z/P2X(7) receptors or 2) MTX and P2Z/P2X(7) receptor stimulation activate a common cytolytic pore. To distinguish between these two possibilities, the effect of MTX was examined in 1) THP-1 monocytic cells before and after treatment with lipopolysaccharide and interferon-gamma, a maneuver known to upregulate P2Z/P2X(7) receptor, 2) wild-type HEK cells and HEK cells stably expressing the P2Z/P2X(7) receptor, and 3) BW5147.3 lymphoma cells, a cell line that expresses functional P2Z/P2X(7) channels that are poorly linked to pore formation. In control THP-1 monocytes, addition of MTX produced a biphasic increase in the cytosolic free Ca(2+) concentration ([Ca(2+)](i)); the initial increase reflects MTX-induced Ca(2+) influx, whereas the second phase correlates in time with the appearance of large pores and the uptake of ethidium. MTX produced comparable increases in [Ca(2+)](i) and ethidium uptake in THP-1 monocytes overexpressing the P2Z/P2X(7) receptor. In both wild-type HEK and HEK cells stably expressing the P2Z/P2X(7) receptor, MTX-induced increases in [Ca(2+)](i) and ethidium uptake were virtually identical. The response of BW5147.3 cells to concentrations of MTX that produced large increases in [Ca(2+)](i) had no effect on ethidium uptake. In both THP-1 and HEK cells, MTX- and Bz-ATP-induced pores activate with similar kinetics and exhibit similar size exclusion. Last, MTX-induced pore formation, but not channel activation, is greatly attenuated by reducing the temperature to 22 degrees C, a characteristic shared by the P2Z/P2X(7)-induced pore. Together, the results demonstrate that, although MTX activates channels that are distinct from those activated by P2Z/P2X(7) receptor stimulation, the cytolytic/oncotic pores activated by MTX- and Bz-ATP are indistinguishable. PMID- 10516108 TI - Effects of macromolecular transport and stochastic fluctuations on dynamics of genetic regulatory systems. AB - To predict the dynamics of genetic regulation, it may be necessary to consider macromolecular transport and stochastic fluctuations in macromolecule numbers. Transport can be diffusive or active, and in some cases a time delay might suffice to model active transport. We characterize major differences in the dynamics of model genetic systems when diffusive transport of mRNA and protein was compared with transport modeled as a time delay. Delays allow for history dependent, non-Markovian responses to stimuli (i.e., "molecular memory"). Diffusion suppresses oscillations, whereas delays tend to create oscillations. When simulating essential elements of circadian oscillators, we found the delay between transcription and translation necessary for oscillations. Stochastic fluctuations tend to destabilize and thereby mask steady states with few molecules. This computational approach, combined with experiments, should provide a fruitful conceptual framework for investigating the function and dynamic properties of genetic regulatory systems. PMID- 10516109 TI - Persistence of external chloride and DIDS binding after chemical modification of Glu-681 in human band 3. AB - Although its primary function is monovalent anion exchange, the band 3 protein also cotransports divalent anions together with protons at low pH. The putative proton binding site, Glu-681 in human erythrocyte band 3, is conserved throughout the anion exchanger family (AE family). To determine whether or not the monovalent anion binding site is located near Glu-681, we modified this residue with Woodward's reagent K (N-ethyl-5-phenylisoxazolium-3'-sulfonate; WRK). Measurements of Cl(-) binding by (35)Cl-NMR show that external Cl(-) binds to band 3 even when Cl(-) transport is inhibited approximately 95% by WRK modification of Glu-681. This indicates that the external Cl(-) binding site is not located near Glu-681 and thus presumably is distant from the proton binding site. DIDS inhibits Cl(-) binding even when WRK is bound to Glu-681, indicating that the DIDS binding site is also distant from Glu-681. Our data suggest that the DIDS site and probably also the externally facing Cl(-) transport site are located nearer to the external surface of the membrane than Glu-681. PMID- 10516110 TI - Effects of osmolarity on taste receptor cell size and function. AB - Osmotic effects on salt taste were studied by recording from the rat chorda tympani (CT) nerve and by measuring changes in cell volume of isolated rat fungiform taste receptor cells (TRCs). Mannitol, cellobiose, urea, or DMSO did not induce CT responses. However, the steady-state CT responses to 150 mM NaCl were significantly increased when the stimulus solutions also contained 300 mM mannitol or cellobiose, but not 600 mM urea or DMSO. The enhanced CT responses to NaCl were reversed when the saccharides were removed and were completely blocked by addition of 100 microM amiloride to the stimulus solution. Exposure of TRCs to hyperosmotic solutions of mannitol or cellobiose induced a rapid and sustained decrease in cell volume that was completely reversible, whereas exposure to hypertonic urea or DMSO did not induce sustained reductions in cell volume. These data suggest that the osmolyte-induced increase in the CT response to NaCl involves a sustained decrease in TRC volume and the activation of amiloride sensitive apical Na(+) channels. PMID- 10516111 TI - PLA(2) stimulation of Na(+)/H(+) antiport and proliferation in rat aortic smooth muscle cells. AB - The proliferative properties and the ability to stimulate the Na(+)/H(+) antiport activity of a secretory phospholipase A(2) were studied in rat aortic smooth muscle cells in culture. The requirement of the enzymatic activity of phospholipase A(2) to elicit mitogenesis was assessed by the use of ammodytin L, a Ser(49) phospholipase A(2) from the venom of Vipera ammodytes, devoid of hydrolytic activity. We propose that the proliferative effect is mediated by the same transduction pathway for both proteins. In particular, 1) both secretory phospholipase A(2) and ammodytin L stimulated thymidine incorporation in a dose dependent manner; 2) both proteins affected the cell cycle, as assessed by cell growth and fluorescence-activated cell sorting experiments; 3) both phospholipase A(2) and ammodytin L increased intracellular pH, a permissive factor for cell proliferation, through activation of the Na(+)/H(+) antiport; 4) ammodytin L was able to displace the (125)I-labeled phospholipase A(2) from specific binding sites in a concentration range consistent with that capable of eliciting a cellular response; and 5) the inhibition by heparin was similar for both proteins, taking into account the ratio of heparin to protein. In conclusion, the enzymatic activity of phospholipase A(2) is not required for the stimulation of mitogenesis. The inhibitory effect of heparin combined with its therapeutic potential could help to clarify the role of phospholipase A(2) in the pathogenesis of several preinflammatory situations. PMID- 10516112 TI - Volume-dependent taurine release from cultured astrocytes requires permissive [Ca(2+)](i) and calmodulin. AB - Cell swelling results in regulatory activation of multiple conductive anion pathways permeable toward a broad spectrum of intracellular organic osmolytes. Here, we explore the involvement of extracellular and intracellular Ca(2+) in volume-dependent [(3)H]taurine efflux from primary cultured astrocytes and compare the Ca(2+) sensitivity of this efflux in slow (high K(+) medium induced) and fast (hyposmotic medium induced) cell swelling. Neither Ca(2+)-free medium nor Ca(2+)-channel blockers prevented the volume-dependent [(3)H]taurine release. In contrast, loading cells with the membrane-permeable Ca(2+) chelator 1,2-bis(2 aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA)-AM suppressed [(3)H]taurine efflux by 65-70% and 25-30% under high-K(+) and hyposmotic conditions, respectively. Fura 2 measurements confirmed that BAPTA-AM, but not Ca(2+)-free media, significantly reduced resting intracellular Ca(2+) concentration ([Ca(2+)](i)). The calmodulin antagonists trifluoperazine and fluphenazine reversibly and irreversibly, respectively, inhibited the high-K(+) induced [(3)H]taurine release, consistent with their known actions on calmodulin. In hyposmotic conditions, the effects were less pronounced. These data suggest that volume-dependent taurine release requires minimal basal [Ca(2+)](i) and involves calmodulin-dependent step(s). Quantitative differences in Ca(2+)/calmodulin sensitivity of high-K(+)-induced and hyposmotic medium-induced taurine efflux are due to both the effects of the inhibitors on high-K(+)-induced cell swelling and their effects on transport systems and/or signaling mechanisms determining taurine efflux. PMID- 10516113 TI - Defective function of the cystic fibrosis-causing missense mutation G551D is recovered by genistein. AB - The patch-clamp technique was used to investigate the effects of the isoflavone genistein on disease-causing mutations (G551D and DeltaF508) of the cystic fibrosis transmembrane conductance regulator (CFTR). In HeLa cells recombinantly expressing the trafficking-competent G551D-CFTR, the forskolin-stimulated Cl currents were small, and average open probability of G551D-CFTR was P(o) = 0.047 +/- 0.019. Addition of genistein activated Cl currents approximately 10-fold, and the P(o) of G551D-CFTR increased to 0.49 +/- 0.12, which is a P(o) similar to wild-type CFTR. In cystic fibrosis (CF) epithelial cells homozygous for the trafficking-impaired DeltaF508 mutation, forskolin and genistein activated Cl currents only after 4-phenylbutyrate treatment. These data suggested that genistein activated CFTR mutants that were present in the cell membrane. Therefore, we tested the effects of genistein in CF patients with the G551D mutation in nasal potential difference (PD) measurements in vivo. The perfusion of the nasal mucosa of G551D CF patients with isoproterenol had no effect; however, genistein stimulated Cl-dependent nasal PD by, on average, -2.4 +/- 0.6 mV, which corresponds to 16.9% of the responses (to beta-adrenergic stimulation) found in healthy subjects. PMID- 10516114 TI - Anchoring protein is required for cAMP-dependent stimulation of L-type Ca(2+) channels in rabbit portal vein. AB - Stimulation of cardiac L-type Ca(2+) channels by cAMP-dependent protein kinase (PKA) requires anchoring of PKA to a specific subcellular environment by A-kinase anchoring proteins (AKAP). This study evaluated the possible requirement of AKAP in PKA-dependent regulation of L-type Ca(2+) channels in vascular smooth muscle cells using the conventional whole cell patch-clamp technique. Peak Ba(2+) current in freshly isolated rabbit portal vein myocytes was significantly increased by superfusion with either 0.5 microM isoproterenol (131 +/- 3% of the control value, n = 11) or 10 microM 8-bromoadenosine 3',5'-cyclic monophosphate (8-BrcAMP; 114 +/- 1%, n = 8). The PKA-induced stimulatory effects of both isoproterenol and 8-BrcAMP were completely abolished by a specific PKA inhibitor KT-5720 (0.2 microM) or by dialyzing cells with Ht 31 (100 microM), a peptide that inhibits the binding of PKA to AKAP. In contrast, Ht 31 did not block the excitatory effect of the catalytic subunit of PKA when dialyzed into the cells. These data suggest that stimulation of Ca(2+) channels in vascular myocytes by endogenous PKA requires localization of PKA through binding to AKAP. PMID- 10516115 TI - Pharmacokinetics of insulin-like growth factor I in hypopituitarism: correlation with binding proteins. AB - We investigated the pharmacokinetics of recombinant human insulin-like growth factor I (rhIGF-I) in growth hormone deficiency (GHD). Nine GHD adults [age 25 +/ 3 (SE) yr] received rhIGF-I (60 microgram/kg sc) twice, 10 h apart, and blood was sampled over 24 h. IGF-I and free IGF-I concentrations increased, whereas IGF binding protein 3 (IGFBP-3) and acid labile subunit (ALS) were unchanged during treatment. There was no correlation between absorption or terminal half-life of IGF-I and IGFBP-3 or ALS, but negative correlations with IGF-I clearance (CL/F) and volume of distribution (V/F). Positive correlations between both IGFBP-3 and ALS and IGF-I maximal concentration (C(max)) and time of C(max) (T(max)) were observed. Compared with normal individuals studied similarly (using 80 microgram/kg), GHD subjects showed a normal absorption half-life, a faster elimination half-life, lower C(max), yet normal T(max) and V/F. In conclusion, GHD is associated with normal absorption and distribution of IGF-I yet faster elimination kinetics. Additionally, IGFBP-3 and ALS concentrations modulate the peak concentrations of IGF-I achieved and correlate reciprocally with its V/F and CL/F, underscoring the critical importance of binding proteins in modulating the bioavailability of IGF-I in vivo in humans. PMID- 10516116 TI - Myocardial blood flow, oxygen consumption, and fatty acid uptake in endurance athletes during insulin stimulation. AB - We have previously demonstrated reduced myocardial glucose uptake rates in hearts of endurance athletes, which could be due to increased use of alternative fuels or reduced energy demands. In the present study myocardial blood flow, oxygen consumption, and free fatty acid uptake were measured with [(15)O]H(2)O, [(15)O]O(2), [(18)F]FTHA, and positron emission tomography (PET) in 9 endurance athletes and 11 sedentary men during euglycemic hyperinsulinemia. Compared with sedentary men, athletes had 33% lower myocardial blood flow, 27% lower oxygen consumption, and 20% lower estimated myocardial work per gram of tissue. Myocardial fatty acid uptake rates were not significantly different in endurance athletes (0.83 +/- 0.29) and sedentary men (1.0 +/- 0.31 micromol. 100 g(-1). min(-1), P = 0.232). In conclusion, myocardial blood flow and oxygen consumption per unit mass of myocardium are reduced at rest in endurance athletes. This can be explained by reduced energy requirements per gram of tissue due to anatomic and physiological changes of the athlete's heart. PMID- 10516117 TI - Inhibition of liver protein synthesis during laparoscopic surgery. AB - Previous studies have indicated that laparoscopic surgery is associated with a decline in liver protein synthesis. In this study, the fractional synthesis rate (FSR) of total liver protein and albumin was measured in patients undergoing elective laparoscopic cholecystectomy at different times after commencing the procedure (n = 8 + 8). Liver biopsy specimens were taken after 15 min of surgery in an "early" group and after 49 min of surgery in a "late" group. The liver FSR was higher in the early group (24.1 +/- 4.7%/day) compared with the late group (19.0 +/- 2.8%/day, P < 0.02). The fractional and absolute synthesis rates of albumin were similar in the two groups, 6.4 +/- 1.5 vs. 6.5 +/- 1.0%/day and 97 +/- 19 vs. 96 +/- 18 mg. kg(-1). day(-1) for the early and late groups, respectively. It is concluded that laparoscopic surgery was accompanied by a decrease in total liver protein synthesis rate, which developed rapidly during surgery. In contrast, no change in the synthesis rate of albumin was apparent during the course of surgery. PMID- 10516118 TI - Availability of intestinal microbial lysine for whole body lysine homeostasis in human subjects. AB - We have investigated whether there is a net contribution of lysine synthesized de novo by the gastrointestinal microflora to lysine homeostasis in six adults. On two separate occasions an adequate diet was given for a total of 11 days, and a 24-h (12-h fast, 12-h fed) tracer protocol was performed on the last day, in which lysine turnover, oxidation, and splanchnic uptake were measured on the basis of intravenous and oral administration of L-[1-(13)C]lysine and L-[6,6 (2)H(2)]lysine, respectively. [(15)N(2)]urea or (15)NH(4)Cl was ingested daily over the last 6 days to label microbial protein. In addition, seven ileostomates were studied with (15)NH(4)Cl. [(15)N]lysine enrichment in fecal and ileal microbial protein, as precursor for microbial lysine absorption, and in plasma free lysine was measured by gas chromatography-combustion-isotope ratio mass spectrometry. Differences in plasma [(13)C]- and [(2)H(2)]lysine enrichments during the 12-h fed period were observed between the two (15)N tracer studies, although the reason is unclear, and possibly unrelated to the tracer form per se. In the normal adults, after (15)NH(4)Cl and [(15)N(2)]urea intake, respectively, lysine derived from fecal microbial protein accounted for 5 and 9% of the appearance rate of plasma lysine. With ileal microbial lysine enrichment, the contribution of microbial lysine to plasma lysine appearance was 44%. This amounts to a gross microbial lysine contribution to whole body plasma lysine turnover of between 11 and 130 mg. kg(-1). day(-1), depending on the [(15)N]lysine precursor used. However, insofar as microbial amino acid synthesis is accompanied by microbial breakdown of endogenous amino acids or their oxidation by intestinal tissues, this may not reflect a net increase in lysine absorption. Thus we cannot reliably estimate the quantitative contribution of microbial lysine to host lysine homeostasis with the present paradigm. However, the results confirm the significant presence of lysine of microbial origin in the plasma free lysine pool. PMID- 10516119 TI - Lower recovery of muscle protein lost during starvation in old rats despite a stimulation of protein synthesis. AB - Sarcopenia could result from the inability of an older individual to recover muscle lost during catabolic periods. To test this hypothesis, we compared the capacity of 5-day-refed 12- and 24-mo-old rats to recover muscle mass lost after 10 days without food. We measured gastrocnemius and liver protein synthesis with the flooding-dose method and also measured nitrogen balance, 3-methylhistidine excretion, and the gene expression of components of proteolytic pathways in muscle comparing fed, starved, and refed rats at each age. We show that 24-mo-old rats had an altered capacity to recover muscle proteins. Muscle protein synthesis, inhibited during starvation, returned to control values during refeeding in both age groups. The lower recovery in 24-mo-old rats was related to a lack of inhibition of muscle proteolysis during refeeding. The level of gene expression of components of the proteolytic pathways did not account for the variations in muscle proteolysis at both ages. In conclusion, this study highlights the role of muscle proteolysis in the lower recovery of muscle protein mass lost during catabolic periods. PMID- 10516120 TI - 4-Hydroxyisoleucine: experimental evidence of its insulinotropic and antidiabetic properties. AB - We have recently shown in vitro that 4-hydroxyisoleucine (4-OH-Ile), an amino acid extracted from fenugreek seeds, potentiates insulin secretion in a glucose dependent manner. The present study was designed to investigate whether 4-OH-Ile could exert in vivo insulinotropic and antidiabetic properties. For this purpose, intravenous or oral glucose tolerance tests (IVGTTs and OGTTs, respectively) were performed not only in normal animals but also in a type II diabetes rat model. During IVGTT in normal rats or OGTT in normal dogs, 4-OH-Ile (18 mg/kg) improved glucose tolerance. The lactonic form of 4-OH-Ile was ineffective in normal rats. In non-insulin-dependent diabetic (NIDD) rats, a single intravenous administration of 4-OH-Ile (50 mg/kg) partially restored glucose-induced insulin response without affecting glucose tolerance; a 6-day subchronic administration of 4-OH-Ile (50 mg/kg, daily) reduced basal hyperglycemia, decreased basal insulinemia, and slightly, but significantly, improved glucose tolerance. In vitro, 4-OH-Ile (200 microM) potentiated glucose (16.7 mM)-induced insulin release from NIDD rat-isolated islets. So, the antidiabetic effects of 4-OH-Ile on NIDD rats result, at least in part, from a direct pancreatic B cell stimulation. PMID- 10516121 TI - Glycyrrhetinic acid-induced apoptosis in thymocytes: impact of 11beta hydroxysteroid dehydrogenase inhibition. AB - It has been proposed that glycyrrhetinic acid (GA) enhances endogenous glucocorticoid (GC) action by suppressing the metabolism of the steroid. We show here that marked involution of the thymus occurred within 24 h of a single intraperitoneal administration of GA in mice. Thymocytes from mice treated with GA exhibited DNA cleavage and mitochondrial transmembrane potential disruption, as demonstrated with agarose gel electrophoresis and flow cytometric analysis. Immunocytochemical staining revealed that CD4(+)CD8(+) double positive cells markedly decreased after GA treatment. In contrast to GA in vivo, GA in vitro did not induce apoptosis of cultured thymocytes. These findings suggest that the apoptosis-inducing effect of GA on thymocytes is due to its indirect action. Because GA has been known to inhibit 11beta-hydroxysteroid dehydrogenase (11beta HSD), we measured the enzyme activity in major organs and endogenous corticosterone concentration after GA treatment. The results showed a significant decrease of 11beta-HSD activity (P < 0.0001) and an increase in serum corticosterone concentration (P < 0.005). We concluded that the inhibition of hepatic 11beta-HSD activity by GA has a serious effect on GC metabolism, which results in a significant elevation of systemic GC levels. Apoptosis of thymocytes occurred as a consequence of the elevation in the level of endogenous corticosterone. PMID- 10516122 TI - Potassium control of extrarenal renin secretion in transgenic (mRen-2)27 and normal rats. AB - Plasma active renin and prorenin were followed for 12 h after bilateral, unilateral, and sham nephrectomy (BNx, UNx, and SNx) in anesthetized transgenic (mRen-2)27 rats to compare them with Sprague-Dawley and spontaneously hypertensive rats (SDR and SHR). In Ren-2 rats, active renin and prorenin increased with plasma potassium post-BNx and were augmented by potassium infusion. The increase in prorenin but not active renin was abolished by bilateral adrenalectomy (BADRx). However, this did not reduce prorenin below normal, indicating that the high plasma prorenin Ren-2 phenotype is not only of adrenal origin. SNx and UNx also raised plasma active renin and prorenin in Ren-2 rats, with positive correlations to plasma potassium. In SDR and SHR, active renin fell below prorenin post-BNx, and adrenal ablation and potassium loading (in SDR) modified the decreasing active renin profile consistent with low levels of regulated extrarenal secretion. In Ren-2 rats, adrenal but not extra-adrenal prorenin secretion is potassium sensitive and stress related. The unidentified source of active renin in BNx+BADRx Ren-2 rats is also potassium and stress related. PMID- 10516123 TI - Renin in thymus, gut, hindlimb, and adrenal of (mRen-2)27 and normal rats: secretion and content studies. AB - Thymic ablation and assay of organ renin revealed that one-third of the increasing plasma level of active renin after removal of kidneys and adrenals from Ren-2 rats originates from the thymus. Splanchnic arteriovenous difference and renin content indicate that gut can account for the remainder. Secretion of active renin from these sites correlated significantly with increasing plasma potassium. Prorenin was not secreted from these sites or from hindlimb in amounts sufficient to raise the plasma level, and yet plasma prorenin remained higher than active renin throughout the 12-h protocol. The source of prorenin that accounts for the high plasma prorenin phenotype of the intact conscious Ren-2 rat was not specifically identified. When sensitive assays were used, a low level of active renin secretion from thymus and gut was also apparent 12 h after removal of kidneys and adrenals in normal Sprague-Dawley rats, and plasma prorenin was at this time higher than active renin. A likely source of this extrarenal, extra adrenal renin is the macrophage. PMID- 10516124 TI - Effect of induced metabolic acidosis on human skeletal muscle metabolism during exercise. AB - The roles of pyruvate dehydrogenase (PDH), glycogen phosphorylase (Phos), and their regulators in lactate (Lac(-)) metabolism were examined during incremental exercise after ingestion of 0.3 g/kg of either NH(4)Cl [metabolic acidosis (ACID)] or CaCO(3) [control (CON)]. Subjects were studied at rest, at rest postingestion, and during continuous steady-state cycling at three stages (15 min each): 30, 60, and 75% of maximal oxygen uptake. Radial artery and femoral venous blood samples, leg blood flow, and biopsies of the vastus lateralis were obtained during each power output. ACID resulted in significantly lower intramuscular concentration of [Lac(-)] (ACID 40.8 vs. CON 56.9 mmol/kg dry wt), arterial whole blood [Lac(-)] (ACID 4.7 vs. CON 6.5 mmol/l), and leg Lac(-) efflux (ACID 3.05 vs. CON 6.98 mmol. l(-1). min(-1)). The reduced intramuscular [Lac(-)] resulted from decreases in pyruvate production due to inhibition of glycogenolysis, at the level of Phos a, and phosphofructokinase, together with an increase in the amount of pyruvate oxidized relative to the total produced. The reduction in Phos a activity was mediated through decreases in transformation, decreases in free inorganic phosphate concentration, and decreases in the posttransformational allosteric regulator free AMP. Reduced PDH activity occurred with ACID and may have resulted from alterations in the concentrations of acetyl-CoA, free ADP, pyruvate, NADH, and H(+), leading to greater relative activity of the kinase. The results demonstrate that imposed metabolic acidosis in skeletal muscle results in decreased Lac(-) production due to inhibition of glycogenolysis at the level of Phos and increased pyruvate oxidation at PDH. PMID- 10516125 TI - Reduced beta-cell function contributes to impaired glucose tolerance in dogs made obese by high-fat feeding. AB - The ability to increase beta-cell function in the face of reduced insulin sensitivity is essential for normal glucose tolerance. Because high-fat feeding reduces both insulin sensitivity and glucose tolerance, we hypothesized that it also reduces beta-cell compensation. To test this hypothesis, we used intravenous glucose tolerance testing with minimal model analysis to measure glucose tolerance (K(g)), insulin sensitivity (S(I)), and the acute insulin response to glucose (AIR(g)) in nine dogs fed a chow diet and again after 7 wk of high-fat feeding. Additionally, we measured the effect of consuming each diet on 24-h profiles of insulin and glucose. After high-fat feeding, S(I) decreased by 57% (P = 0.003) but AIR(g) was unchanged. This absence of beta-cell compensation to insulin resistance contributed to a 41% reduction of K(g) (P = 0.003) and abolished the normal hyperbolic relationship between AIR(g) and S(I) observed at baseline. High-fat feeding also elicited a 44% lower 24-h insulin level (P = 0.004) in association with an 8% reduction of glucose (P = 0.0003). We conclude that high-fat feeding causes insulin resistance that is not compensated for by increased insulin secretion and that this contributes to the development of glucose intolerance. These effects of high-fat feeding may be especially deleterious to individuals predisposed to type 2 diabetes mellitus. PMID- 10516126 TI - Sympathetic inhibition, leptin, and uncoupling protein subtype expression in normal fasting rats. AB - To further investigate neural effects on leptin and uncoupling proteins (UCPs), we studied in vivo perturbations intended to block adrenergic input to peripheral tissues. We examined plasma leptin, leptin mRNA, and adipose and muscle UCP subtype mRNA in rats treated with alpha-methyl-p-tyrosine methyl ester (AMPT-ME), which inhibits catecholamine synthesis and 6-hydroxydopamine (6HDA), which is toxic to catecholinergic nerve terminals but, unlike AMPT-ME, does not enter the central nervous system. Intraperitoneal AMPT-ME, 250 mg/kg, was administered at 1800 and 0700 the following day, and rats were killed at 1200-1400. All rats were fasted with free access to water during this time. Intraperitoneal AMPT-ME increased plasma leptin by 15-fold, increased interscapular brown adipose tissue (IBAT) and epididymal fat leptin mRNA by 2- to 2.5-fold, and also increased plasma insulin and glucose concentrations. Intraperitoneal AMPT-ME decreased IBAT UCP-3 mRNA to 40% of control, while it increased epididymal adipose UCP-3 mRNA approximately twofold. Intravenous AMPT-ME, 250 mg/kg, administered to conscious rats for 5 h decreased lumbar sympathetic nerve activity, increased plasma leptin (5.89 +/- 1.43 compared with 2.75 +/- 0.31 ng/ml in vehicle-treated rats, n = 7, P < 0.05), and decreased cardiac rate with no sustained change in blood pressure. Intraperitoneal 6HDA, 100 mg/kg, as a single dose at 1800, increased plasma leptin approximately twofold after 18-20 h, increased IBAT (but not epididymal fat) leptin mRNA by two- to threefold, and decreased IBAT UCP-3 mRNA to 30-40% of control. Neither AMPT-ME nor 6HDA significantly altered mRNA encoding gastrocnemius muscle UCP-3, IBAT UCP-1, or IBAT and epididymal UCP-2. In summary, AMPT-ME and 6HDA increased plasma leptin and upregulated leptin mRNA expression. AMPT-ME also resulted in complex tissue and subtype-specific modulation of adipose UCP mRNA. These data are consistent with interaction between leptin and sympathetic nerve activity (SNA) in regulation of fat cell energy utilization. However, the in vivo modulation of leptin and UCPs appears complex and, beyond a causal effect of SNA per se, may depend on concurrent changes in plasma insulin, glucose, and circulatory hemodynamics. PMID- 10516128 TI - Inverse alterations of BCKA dehydrogenase activity in cardiac and skeletal muscles of diabetic rats. AB - Rat cardiac and skeletal muscles, which have been used as model tissues for studies of regulation of branched-chain alpha-keto acid (BCKA) oxidation, vary greatly in the activity state of their BCKA dehydrogenase. In the present experiment, we have investigated whether they also vary in response of their BCKA dehydrogenase to a metabolic alteration such as diabetes and, if so, to investigate the mechanism that underlies the difference. Diabetes was produced by depriving streptozotocin-treated rats of insulin administration for 96 h. The investigation of BCKA dehydrogenase in the skeletal muscle (gastrocnemius) showed that diabetes 1) increased its activity, 2) increased the protein and gene expressions of all of its subunits (E(1)alpha, E(1)beta, E(2)), 3) increased its activity state, 4) decreased the rate of its inactivation, and 5) decreased the protein expression of its associated kinase (BCKAD kinase) without affecting its gene expression. In sharp contrast, the investigation of BCKA dehydrogenase in the cardiac muscle showed that diabetes 1) decreased its activity, 2) had no effect on either protein or gene expression of any of its subunits, 3) decreased its activity state, 4) increased its rate of inactivation, and 5) increased both the protein and gene expressions of its associated kinase. In conclusion, our data suggest that, in diabetes, the protein expression of BCKAD kinase is downregulated posttranscriptionally in the skeletal muscle, whereas it is upregulated pretranslationally in the cardiac muscle, causing inverse alterations of BCKA dehydrogenase activity in these muscles. PMID- 10516127 TI - The head arterial glucose level is not the reference site for generation of the portal signal in conscious dogs. AB - Experiments were performed on twelve 42-h-fasted, conscious dogs to determine whether the head arterial glucose level is used as a reference standard for comparison with the portal glucose level in bringing about the stimulatory effect of portal glucose delivery on net hepatic glucose uptake (NHGU). Each experiment consisted of an 80-min equilibration, a 40-min control, and two 90-min test periods. After the control period, somatostatin was given along with insulin (7.2 pmol. kg(-1). min(-1); 3.5-fold increase) and glucagon (0.6 ng. kg(-1). min(-1); basal) intraportally. Glucose was infused intraportally (22.2 micromol. kg(-1). min(-1)) and peripherally as needed to double the hepatic glucose load. In one test period, glucose was infused into both vertebral and carotid arteries (HEAD(G); 22.2 +/- 0.8 micromol. kg(-1). min(-1)); in the other test period, saline was infused into the head arteries (HEAD(S)). One-half of the dogs received HEAD(G) first. When all dogs are considered, the blood arterial-portal glucose gradients (-0.52 +/- 0.07 vs. -0.49 +/- 0.03 mM) and the hepatic glucose loads (339 +/- 14 vs. 334 +/- 20 micromol. kg(-1). min(-1)) were similar in HEAD(G) and HEAD(S). NHGU was 24.1 +/- 3.8 and 25.1 +/- 4.6 micromol. kg(-1). min(-1), and nonhepatic glucose uptake was 46.1 +/- 4.2 and 48.8 +/- 7.0 micromol. kg(-1). min(-1) in HEAD(G) and HEAD(S), respectively. The head arterial glucose level is not the reference standard used for comparison with the portal glucose level in the generation of the portal signal. PMID- 10516130 TI - Oligomycin sensitivity of mitochondrial F(1)F(0)-ATPase in diabetes-prone BHE/Cdb rats. AB - BHE/Cdb and Sprague-Dawley rats differ in their mitochondrial DNA sequence for the ATPase 6 ("subunit a") gene. Base substitutions in this sequence result in the substitution of asparagine for aspartate at position 101 and the substitution of serine for leucine at position 129. Differences in sensitivity to oligomycin were observed. When the isolated F(1)F(0)-ATPase complex was studied and ATPase activity was assessed, that which was isolated from the BHE/Cdb rats was less sensitive to oligomycin inhibition than that which was isolated from the Sprague Dawley rats. In contrast, when oxygen consumption was measured [oxygen phosphorylation (OXPHOS)] and a dose-response curve was generated with isolated mitochondria from these two strains, there was a shift to the left for the BHE/Cdb rat mitochondria. These mitochondria were more sensitive to oligomycin inhibition of OXPHOS than were mitochondria isolated from Sprague-Dawley rats. The OXPHOS results are consistent with those from human fibroblasts having either a normal or mutated ATPase 6 gene. PMID- 10516129 TI - Effect of protein intake on plasma and erythrocyte free amino acids and serum IGF I and IGFBP-1 levels in rats. AB - Amino acid (AA) levels in plasma and erythrocytes (RBC) were determined in rats (n = 29) fed diets with 6, 21, and 35% protein, and their association with insulin-like growth factor I (IGF-I), insulin, or IGF-binding protein (IGFBP)-1 levels was studied. Free AA in plasma and RBC were determined by reversed-phase high-pressure liquid chromotography, and IGF-I, IGFBP-1, and insulin plasma levels were determined by RIA. Rats fed the low-protein (6%) diet were growth retarded and had lower serum IGF-I levels and higher serum IGFBP-1 levels than the other two groups (P < 0.0001). In rats fed the low-protein diet, most of the nonessential AA (NEAA) in both plasma and RBC increased, whereas the essential AA (EAA), with the exception of threonine, decreased. When the groups were combined, both RBC and plasma EAA-to-NEAA ratios were positively correlated to IGF-I (r = 0.76 and 0.80, respectively; P < 0.0001) and inversely correlated to IGFBP-1 levels (r = -0.67, P < 0.001 and r = -0.78, P < 0.0001, respectively). A significant inverse correlation was found between RBC glutamate and IGF-I (r = 0.85, P < 0.0001, n = 25) and insulin (r = -0.72, P < 0.001, n = 21), and a positive correlation was found for IGFBP-1 (r = 0.78, P < 0.0001, n = 24). In multiple regression analysis, only IGF-I remained as an independent variable. Threonine was the only EAA with a significant inverse correlation to insulin (r = -0.66, P < 0.001). We hypothesize that AA metabolism is associated to changes in IGF-I, insulin, and IGFBP-1 levels in rats on different protein intakes. PMID- 10516131 TI - Glucocorticoids reverse leptin effects on food intake and body fat in mice without increasing NPY mRNA. AB - Glucocorticoid stimulation of appetite and leptin expression conflicts with leptin inhibition of food intake and suggests that glucocorticoids reduce sensitivity to leptin. To determine if glucocorticoids impair feeding and metabolic responses to leptin, we measured leptin-induced changes in food intake, body weight, hormones, carcass fat, and hypothalamic neuropeptide Y (NPY) mRNA in adrenalectomized mice with and without corticosterone replacement. Leptin infusion (0.5 microgram/h) significantly decreased food intake and body weight in adrenalectomized mice. Corticosterone replacement approximating normal 24-h mean levels restored food intake but did not permit weight gain equivalent to PBS infused controls. Corticosterone levels comparable to stress-induced production completely reversed leptin-induced reductions in weight gain and body fat, despite significant attenuation by leptin of corticosterone-induced increases in plasma insulin levels. Glucocorticoid replacement increased food intake without reversing leptin inhibition of hypothalamic NPY mRNA levels. We conclude that glucocorticoid levels within the physiological range can interfere with leptin action and that glucocorticoid effects are at least partly independent of NPY. PMID- 10516132 TI - Origins of the hydrogen bound to carbon 1 of glucose in fasting: significance in gluconeogenesis quantitation. AB - Healthy subjects ingested (2)H(2)O. (2)H enriched the hydrogen bound to carbon 1 of blood glucose 1.3 to 1.8 times more than the hydrogens bound to carbon 6. Enrichment at carbon 1 was more than at carbon 5 after 14 h, but not after 42 h, of fasting. After overnight fasting, when [2,3-(3)H]succinate was infused, 34 times as much (3)H was bound to carbon 6 as to carbon 1. On [1-(2)H,1-(3)H, 1 (14)C]galactose infusion, the ratios of (2)H to (14)C and of (3)H to (14)C in blood glucose were 30% less than in the galactose. (3)H at carbon 6 was 1% of that at carbon 1 of the glucose. Thus, although the two hydrogens bound to carbon 1 and the two bound to carbon 6 of fructose 6-phosphate (p) during gluconeogenesis are equally enriched in (2)H via pyruvate's equilibration with alanine, one of each is further enriched via hydration of fumarate that is converted to glucose. That hydrogen at carbon 1 of fructose 6-phosphate (P) is also enriched in fructose 6-P's equilibration with mannose 6-P. (2)H from (2)H(2)O at carbon 1 to carbon 2 of blood glucose cannot then quantitate gluconeogenesis because of [1-(2)H]glucose formation during glycogenolysis. Triose-P cycling has a minimal effect on quantitation. (2)H recovery in glucose from [1-(2)H]galactose does not quantitate galactose conversion via UDP-glucose to glycogen. PMID- 10516133 TI - Differential regulation of MAP kinase by contraction and insulin in skeletal muscle: metabolic implications. AB - We have investigated the activation of the extracellular signal-regulated kinases (ERK1 and ERK2) by muscle contraction and insulin in perfused rat skeletal muscle. Both stimuli activated ERK1 and ERK2 by an upstream kinase MAP/ERK kinase (MEK)-dependent mechanism, as the MEK inhibitor PD-98059 inhibited ERK phosphorylation. The presence of the phosphatidylinositol (PI) 3-kinase inhibitors LY-294002 and wortmannin totally eradicated ERK1 and ERK2 phosphorylation in response to insulin but not contraction. Insulin and muscle contraction activated muscle glucose transport, glycogen synthase, and amino acid transport independently of ERK signaling, whereas the PI 3-kinase inhibitors abolished the stimulatory effects of insulin but not those of contraction on these three cellular processes. We conclude that 1) insulin and contraction activate ERK signaling in skeletal muscle; 2) ERK signaling is not necessary for activation of glucose and amino acid transport or glycogen synthase activity by contraction and insulin in skeletal muscle; and 3) insulin-induced activation of MEK, the upstream activator of ERK, is dependent on PI 3-kinase, whereas contraction utilizes a different mechanism. PMID- 10516135 TI - Acute effect of growth hormone to induce peripheral insulin resistance is independent of FFA and insulin levels in rats. AB - To examine whether growth hormone (GH) induces peripheral insulin resistance by altering plasma free fatty acid (FFA) or insulin levels, the effects of GH infusion on insulin-stimulated glucose fluxes were studied in conscious rats under two protocols. In study 1, either saline (n = 7) or human recombinant GH (21 microg. kg(-1). h(-1); n = 8) was infused for 300 min, and insulin-stimulated glucose fluxes were estimated during the final 150-min period of hyperinsulinemic euglycemic clamps. In study 2, hyperinsulinemic euglycemic clamps were first conducted for 150 min (to raise plasma insulin and suppress FFA levels), and saline or GH (n = 7 for each) was subsequently infused for the following 300-min clamp period. In study 1, GH infusion in the basal state did not significantly alter plasma FFA or insulin levels. In contrast, GH infusion decreased insulin stimulated glucose uptake, glycolysis, and glycogen synthesis by 32, 27, and 40%, respectively (P < 0.05). In study 2, GH infusion during hyperinsulinemic euglycemic clamps did not alter plasma FFA or insulin levels (P > 0.05). GH infusion had no effect on insulin-stimulated glucose uptake during the initial 150 min but eventually decreased insulin-stimulated glucose uptake by 37% (P < 0. 05), similar to the results in study 1. These data indicate that GH induces peripheral insulin resistance independent of plasma FFA and insulin levels. The induction of insulin resistance was preceded by suppression of glycogen synthesis, consistent with the hypothesis that metabolic impairment precedes and causes development of peripheral insulin resistance. PMID- 10516134 TI - Exercise and insulin cause GLUT-4 translocation in human skeletal muscle. AB - Studies in rodents have established that GLUT-4 translocation is the major mechanism by which insulin and exercise increase glucose uptake in skeletal muscle. In contrast, much less is known about the translocation phenomenon in human skeletal muscle. In the current study, nine healthy volunteers were studied on two different days. On one day, biopsies of vastus lateralis muscle were taken before and after a 2-h euglycemic-hyperinsulinemic clamp (0.8 mU. kg(-1). min( 1)). On another day, subjects exercised for 60 min at 70% of maximal oxygen consumption (VO(2 max)), a biopsy was obtained, and the same clamp and biopsy procedure was performed as that during the previous experiment. Compared with insulin treatment alone, glucose infusion rates were significantly increased during the postexercise clamp for the periods 0-30 min, 30-60 min, and 60-90 min, but not during the last 30 min of the clamp. Plasma membrane GLUT-4 content was significantly increased in response to physiological hyperinsulinemia (32% above rest), exercise (35%), and the combination of exercise plus insulin (44%). Phosphorylation of Akt, a putative signaling intermediary for GLUT-4 translocation, was increased in response to insulin (640% above rest), exercise (280%), and exercise plus insulin (1,000%). These data demonstrate that two normal physiological conditions, moderate intensity exercise and physiological hyperinsulinemia approximately 56 microU/ml, cause GLUT-4 translocation and Akt phosphorylation in human skeletal muscle. PMID- 10516136 TI - Increased intracellular localization of brain GLUT-1 transporter in response to ethanol during chick embryogenesis. AB - Fetal exposure to ethanol is associated with growth retardation of the developing central nervous system. We have previously described a chick model to study the molecular mechanism of ethanol effects on glucose metabolism in ovo. Total membrane fractions were prepared from day 4, day 5, and day 7 chick embryos exposed in ovo to ethanol or to vehicle. By Western blotting analysis, ethanol exposure caused a mean 7- to 10-fold increase in total GLUT-1 and a 2-fold increase in total GLUT-3. However, glucose uptake by ethanol-treated cells increased by only 10%. Analysis of isolated plasma (PM) and intracellular (IM) membranes from day 5 cranial tissue revealed a mean 25% decrease in GLUT-1 in the PM and a 66% increase in the IM in the ethanol group vs. control. The amount of PM GLUT-3 was unchanged but that of IM GLUT-3 was significantly decreased. The data suggest that GLUT-3 cell surface expression may be resistant to the suppressive effects of ethanol in the developing brain of ethanol-treated embryos. The overall increase in GLUT-1 may reflect a deregulation of the transporter induced by ethanol exposure. The increased IM localization and decreased amount of PM GLUT-1 may be a mechanism used by the ethanol-treated cell to maintain normal glucose uptake despite the overall increased level of the transporter. PMID- 10516137 TI - Downregulation of the human taurine transporter by glucose in cultured retinal pigment epithelial cells. AB - In diabetes, activation of the aldose reductase (AR) pathway and alterations of glucose-sensitive signal transduction pathways have been implicated in depletion of intracellular taurine, an endogenous antioxidant and compatible osmolyte. Cellular taurine accumulation occurs by an osmotically induced, protein kinase C (PKC)-regulated Na(+)-taurine cotransporter (hTT). The effects of ambient glucose on taurine content, hTT activity, and hTT gene expression were therefore evaluated in low and high AR-expressing human retinal pigment epithelial cell lines. In low AR-expressing cells, 20 mM glucose decreased taurine content, hTT transporter activity, and mRNA levels, and these effects were unaffected by AR inhibition (ARI). In these cells, the inhibitory effects of high glucose on hTT appeared to be posttranscriptionally mediated, because 20 mM glucose decreased hTT mRNA stability without affecting hTT transcriptional rate. Inhibition of PKC overcame the decrease in hTT activity in high glucose-exposed cells. In high AR expressing cells, prolonged exposure to 20 mM glucose resulted in intracellular taurine depletion, which paralleled sorbitol accumulation and was prevented by ARI. In these cells exposed to 5 mM glucose, hTT mRNA abundance was decreased and declined further in 20 mM glucose but was corrected by ARI. In 5 mM glucose, hTT transcriptional rate was markedly decreased in high AR-expressing cells, did not decline further in 20 mM glucose, but was increased by ARI to levels above those observed in low AR-expressing cells. Therefore, glucose rapidly and specifically decreases taurine content, hTT activity, and mRNA abundance by AR-unrelated and AR-related posttranscriptional and transcriptional mechanisms. PMID- 10516138 TI - Regulation of fatty acid oxidation of the heart by MCD and ACC during contractile stimulation. AB - We tested the hypothesis that the level of malonyl-CoA, as well as the corresponding rate of total fatty acid oxidation of the heart, is regulated by the opposing actions of acetyl-CoA carboxylase (ACC) and malonyl-CoA decarboxylase (MCD). We used isolated working rat hearts perfused under physiological conditions. MCD in heart homogenates was measured specifically by (14)CO(2) production from [3-(14)C]malonyl-CoA, and ACC was measured specifically based on the portion of total carboxylase that is citrate sensitive. Increased heart work (1 microM epinephrine + 40% increase in afterload) elicited a 40% increase in total beta-oxidation of exogenous plus endogenous lipids, accompanied by a 33% decrease in malonyl-CoA. The basal activity and citrate sensitivity of ACC (reflecting its phosphorylation state) and citrate content were unchanged. AMP levels were also unchanged. MCD activity, when measured at a subsaturating concentration of malonyl-CoA (50 microM), was increased by 55%. We conclude that physiological increments in AMP during the work transition are insufficient to promote ACC phosphorylation by AMP-stimulated protein kinase. Rather, increased fatty acid oxidation results from increased malonyl-CoA degradation by MCD. PMID- 10516139 TI - Lessons from genetically engineered animal models. IV. Nitric oxide synthase gene knockout mice. AB - Nitric oxide is a ubiquitous molecule implicated in a variety of biological processes. The specific action of nitric oxide depends on its enzymatic sources, namely neuronal nitric oxide synthase (nNOS), endothelial NOS (eNOS), and inducible NOS (iNOS), each having distinct tissue localization. Conventional pharmacological antagonists could not distinguish these enzymes or provide models of chronic nitric oxide depletion in whole animals. Several lines of knockout mice have been generated to distinguish the roles of nitric oxide from each enzyme: nitric oxide from nNOS is a major inhibitory neurotransmitter, nitric oxide from eNOS regulates blood flow under physiological conditions, and nitric oxide from iNOS causes hypotension during severe inflammatory conditions. Moreover, the nitric oxides from each isoform have different roles in tissue injury and inflammation. Studies of NOS-deficient animals have also identified redundant and compensatory pathways and revealed the consequences of life-long deficiency of these enzymes. The nNOS-deficient mice develop gastric dilation and stasis, the eNOS-deficient mice develop hypotension and lack vasodilatory responses to injury, and iNOS-deficient mice are more susceptible to inflammatory damage but more resistant to septic shock. PMID- 10516140 TI - Nutrient tasting and signaling mechanisms in the gut. I. Sensing of lipid by the intestinal mucosa. AB - It is well recognized that lipid in the intestine is a potent inhibitor of gastric secretomotor function. Progress has been made in the identification of the "sensor" for lipid in the intestinal wall. Long-chain free fatty acids are the stimulus both for release of CCK and for the production of functional effects. Long-chain triglyceride requires chylomicron formation for absorption, and there is strong evidence that the postabsorptive products of long-chain triglyceride absorption, including chylomicrons and apolipoproteins, are involved in sensory transduction in the intestinal wall. PMID- 10516141 TI - Regulation of small intestinal Na-P(i) type IIb cotransporter by dietary phosphate intake. AB - Dietary restriction of phosphate is a well-known stimulator (acting indirectly via vitamin D(3)) of small intestinal apical Na-P(i) cotransport. In the present study, we document by Western blots and immunohistochemistry that, in mice, a low P(i) diet given for several days leads (in parallel to a stimulation of Na-P(i) cotransport) to an increase of the abundance of the type IIb Na-P(i) cotransporter in the brush-border membrane of mouse enterocytes. Similar results were also obtained by an injection of cholecalciferol. The abundance of the type IIb transcript was investigated by Northern blots. These results indicated that the amount of the type IIb transcript was not changed by either low-P(i) diet or cholecalciferol. It is concluded that stimulation of intestinal Na-P(i) cotransport by low-P(i) diet and vitamin D(3) can be explained by an increased amount of type IIb Na-P(i) cotransporters in the brush-border membrane and that augmentation of type IIb Na-P(i) cotransporters is not related to an increased rate of transcription of the type IIb gene. PMID- 10516142 TI - Altered migration of gut-derived T lymphocytes after activation with concanavalin A. AB - Although activation of lymphocytes is known to be associated with profound changes in homing behavior, it remains unclear how activation alters migration of gut-derived lymphocytes in lymphoid and nonlymphoid organs. The objectives of this study were 1) to compare migration of naive and concanavalin A (ConA) activated T lymphocytes into the gut mucosa, spleen, and liver and 2) to define the role of specific adhesion molecules in this homing process. Fluorescently labeled T lymphocytes collected from rat intestinal lymph were injected into the jugular vein, and the kinetics of appearance of the infused lymphocytes were monitored in ileal Peyer's patches, spleen, and liver. The migration of naive and ConA-activated T lymphocytes into microvessels were compared using an intravital microscope. ConA stimulation significantly increased the rolling velocity of T lymphocytes in postcapillary venules of Peyer's patches, and ConA-stimulated lymphocytes exhibited a loss of the selective adherence properties in Peyer's patches that is normally observed with naive T cells. ConA activation also suppressed the accumulation of T cells in the spleen. On the other hand, the adherence of T cells to hepatic sinusoidal endothelium was significantly increased after ConA activation, especially in the periportal area, and this increase was attenuated by an anti-intercellular adhesion molecule (ICAM)-1 antibody. Flow cytometry analysis revealed a decline in L-selectin expression and an increase in CD11a expression and ICAM-1 on the surface of ConA-treated T cells. In conclusion, activation of gut-derived T lymphocytes with ConA significantly alters their migration path, with a diminished localization to Peyer's patches and spleen and a preferential accumulation in hepatic sinusoids. This altered migration pattern likely results from changes in the expression of leukocyte adhesion molecules such as L-selectin and CD11a. PMID- 10516143 TI - Gastrin stimulates the growth of gastric pit with less-differentiated features. AB - Gastrin stimulates the growth of gastric mucosa by increasing mostly its glandular region but is not known to induce the growth of a pit region where its major constituent cells, gastric surface mucous (GSM) cells, turn over rapidly. To investigate the effect of gastrin on GSM cells, we generated hypergastrinemic mice by expressing a human gastrin transgene. We obtained a hypergastrinemic mouse line whose average serum gastrin level is 671 +/- 252 pg/ml (normal level <150 pg/ml). Gastrin-positive cells were found in the fundic mucosa. The gastric mucosa exhibited hypertrophic growth, which was characterized by an elongated pit with an active proliferative zone, but the glandular region containing parietal cells was normal or reduced in size. The GSM cells contained fewer mucous granules than those of control littermates and lost reactivity to the GSM cell specific cholera toxin beta-subunit lectin. GSM cells along the foveolar region and many mucous neck cells became Alcian blue positive, suggesting the appearance of sialomucin in these cells. We suggest that gastrin stimulates the growth of the proliferative zone of gastric glands, which results in the elongation of the pit region whose GSM cells exhibit less-differentiated features. PMID- 10516144 TI - Temporal changes in TFF3 expression and jejunal morphology during methotrexate induced damage and repair. AB - Trefoil factor TFF3 has been implicated in intestinal protection and repair. This study investigated the spatiotemporal relationship between TFF3 expression and morphological changes during intestinal damage and repair in a rat model of methotrexate-induced small intestinal mucositis. Intestinal tissues from rats with mucositis were collected daily for 10 days. Mucosal damage was characterized by an initial decrease in cell proliferation resulting in crypt loss, villus atrophy, and depletion of goblet cells, followed by hyperproliferation that lead to crypt and villus regeneration and mucous cell repopulation. TFF3 mRNA levels increased marginally during histological damage, and the cell population expressing TFF3 mRNA expanded from the usual goblet cells to include some nongoblet epithelial cells before goblet cell repopulation. TFF3 peptide, however, was depleted during histological damage and normalized during repair, mirroring the disappearance and repopulation of goblet cells. Although there is no temporal relationship between TFF3 levels and crypt hyperproliferation, confirming the nonmitogenic nature of TFF3, the coincidental normalization of TFF3 peptide with repopulation of goblet cells and mucin production after proliferative overshoot suggests that TFF3 may play a role in the remodeling phase of repair. PMID- 10516145 TI - Role of salivary mucin in the protection of rat esophageal mucosa from acid and pepsin-induced injury. AB - The mucosal defensive mechanisms of the esophagus against acid and pepsin remain to be elucidated. In the present study, we investigated the contribution of the salivary mucin in maintaining the integrity of the esophageal mucosa. When an everted esophageal sac, isolated from normal rat, was treated with N-acetyl-L cysteine, a mucolytic agent, the amount of glycoprotein in the gel layer adherent to the epithelium was completely depleted and the susceptibility of the mucosa against acidified pepsin-induced digestion increased. In sialoadenectomized rats, 7 days after extirpation, the amount of glycoprotein adherent to the esophageal epithelium was definitely reduced, and the esophageal mucosa was significantly vulnerable to acidified pepsin-induced digestion compared with the sham-operated rats. Induction of regurgitation of the gastric juices into the esophagus resulted in the development of severe hemorrhagic esophageal lesions only in the sialoadenectomized rats but not in the sham-operated rats. In conclusion, the glycoprotein in the adherent gel layer in rat esophagus, which mainly derives from salivary glands, plays an important role in the preepithelial defense to maintain the integrity of the esophageal mucosa against acid and pepsin. PMID- 10516146 TI - IL-1beta mediates induction of hepatic type 1 plasminogen activator inhibitor in response to local tissue injury. AB - Type 1 plasminogen activator inhibitor (PAI-1), a major physiological inhibitor of plasminogen activation, is an important component of the hepatic acute phase response. We studied the acute phase regulation of murine hepatic PAI-1 in response to systemic toxicity and local tissue injury in both wild-type mice and in mice in which the interleukin (IL)-1beta gene had been inactivated by gene targeting. Endotoxin induced plasma PAI-1 antigen levels and PAI-1 mRNA accumulation in liver to the same extent in both wild-type and IL-1beta-deficient mice. In contrast, turpentine increased plasma PAI-1 and hepatic PAI-1 mRNA accumulation in wild-type mice but not in IL-1beta-deficient mice. Intraperitoneal injection of murine IL-1beta rapidly increased plasma PAI-1 and hepatic PAI-1 mRNA in both wild-type and IL-1beta-deficient mice. These results suggest that IL-1beta is a critical inducer of hepatic PAI-1 gene expression during the acute phase response to local tissue injury. In situ hybridization studies revealed that hepatocytes are the cells primarily responsible for the hepatic expression of the PAI-1 gene induced by lipopolysaccharide and turpentine. PMID- 10516147 TI - Human intestinal epithelial cells express receptors for platelet-activating factor. AB - The intestinal epithelium produces and responds to cytokines and lipid mediators that play a key role in the induction and regulation of mucosal inflammation. The lipid mediator platelet-activating factor (PAF) can be produced and degraded by the human intestinal epithelium and is known to mediate a range of proinflammatory and other biological effects in the intestinal mucosa. In the studies herein, we assessed whether or not human intestinal epithelial cells express cell surface or intracellular PAF receptors (PAF-R), whether expression of these receptors can be regulated, and whether human intestinal epithelial cells respond to PAF. Several human colon epithelial cell lines (HT-29, Caco-2, T84, HCT-8, HCA-7, I407, and LS-174T) were shown by RT-PCR to constitutively express mRNA for PAF-R. In addition, PAF-R expression was demonstrated by immunoblot analysis and PAF-R was shown to be constitutively expressed on the cell surface of several of these cell lines, as assessed by flow cytometry. PAF-R expression by human colon epithelial cells was upregulated by stimulation with retinoic acid but not by stimulation with PAF, proinflammatory agonists (tumor necrosis factor-alpha, interleukin-1, interferon-gamma), or transforming growth factor-alpha. PAF-R on intestinal epithelial cells were functional, as PAF stimulation of the cells increased tyrosine phosphorylation of several cellular proteins, including proteins of 75 and 125 kDa, and this response was blocked by a PAF-R antagonist. Consistent with the findings using cell lines, PAF-R were also constitutively expressed by normal human colon and small intestinal epithelium in vivo, as shown by immunohistology. The constitutive and regulated expression of functional PAF-R by human intestinal epithelium suggests PAF produced by the intestinal epithelial cells or cells underlying the epithelium has autocrine or paracrine effects on intestinal epithelial cells. PMID- 10516148 TI - Role of the vasopressin V(1) receptor in regulating the epithelial functions of the guinea pig distal colon. AB - Vasopressin has a wide spectrum of biological action. In this study, the role of vasopressin in regulating electrolyte transport in the colon was elucidated by measuring the short-circuit current (I(sc)) as well as the Na(+), K(+), and Cl(-) flux in a chamber-mounted mucosal sheet. The cytosolic Ca(2+) concentration ([Ca(2+)](i)) was also measured in fura 2-loaded cells by fluorescence imaging. Serosal vasopressin decreased I(sc) at 10(-9) M and increased I(sc) at 10(-7)-10( 6) M. The decrease in I(sc) was accompanied by two effects: one was a decrease in the amiloride-sensitive Na(+) absorption, whereas the other was an increase in the bumetanide-sensitive K(+) secretion. The increase in I(sc) was accompanied by an increase in the Cl(-) secretion that can be inhibited by serosal bumetanide or mucosal diphenylamine-2-carboxylate. Vasopressin caused an increase in [Ca(2+)](i) in crypt cells. These responses of I(sc) and the [Ca(2+)](i) increase in crypt cells were all more potently inhibited by the vasopressin V(1) receptor antagonist than by the V(2) receptor antagonist. These results suggest that vasopressin inhibits electrogenic Na(+) absorption and stimulates electrogenic K(+) and Cl(-) secretion. In all of these responses, the V(1) receptor is involved, and the [Ca(2+)](i) increase may play an important role. PMID- 10516149 TI - Divergent regulation of human and rat proglucagon gene promoters in vivo. AB - A single mammalian proglucagon gene is expressed in the brain, islets, and intestinal enteroendocrine cells, which gives rise to a unique profile of proglucagon-derived peptides (PGDPs) in each tissue. The biological importance of glucagon, glucagon-like peptide (GLP)-1, and GLP-2 has engendered considerable interest in the factors regulating the synthesis and secretion of the PGDPs in vivo. Although rat proglucagon gene transcription has been extensively studied, the factors important for control of human proglucagon gene expression have not been examined. We now report that, despite conservation of proximal promoter G1 G4 enhancer-like elements, human proglucagon reporter plasmids containing these elements are transcriptionally inactive in islet cell lines. Remarkably, larger human proglucagon promoter fragments, such as the 1604 hGLU-Luc, are expressed in GLUTag enteroendocrine cells but not in islet cell lines. A total of 5775 bases of human proglucagon promoter were required for expression in islet cell lines. Analysis of human proglucagon promoter expression in transgenic mice demonstrated that approximately 1.6 kb of human proglucagon gene sequences directs expression of a human growth hormone reporter gene to the brain and intestinal enteroendocrine cells but not islet cells in vivo. These findings provide the first evidence demonstrating divergence in the mechanisms utilized for tissue specific regulation of the human and rodent proglucagon genes. PMID- 10516150 TI - Changes in growth factor and cytokine mRNA levels after hepatectomy in rat with CCl(4)-induced cirrhosis. AB - Cirrhotic liver is considered to regenerate less actively than normal liver after hepatic resection. However, the mechanisms responsible for this impaired regeneration and the cross talk of implicated factors still remain unclear. In the present study, mRNA levels for cyclins, growth factors, and cytokines were quantitatively assessed by a RT-PCR method at different times after hepatectomy in order to determine the relationships between these factors and the impaired regenerative process observed in cirrhotic liver. In our model of CCl(4)-induced cirrhosis, mRNA levels for cyclins and thymidine kinase provide evidence for the impaired and delayed hepatic regeneration. Moreover, we observed a significant decrease in interleukin (IL)-6 and tumor necrosis factor-alpha mRNA and a significant increase for IL-1beta mRNA. No significant change of hepatocyte growth factor (HGF) mRNA level was detected, contrasting with the decrease both at mRNA and protein levels in the expression of the c-Met/HGF receptor. Therefore, the impaired regeneration of the cirrhotic liver is associated not only with a lowered level of signals that normally promote liver growth but also with a strong decrease in c-Met receptor despite a normal expression of its specific ligand. PMID- 10516151 TI - Phosphodiesterase inhibitors prevent NSAID enteropathy independently of effects on TNF-alpha release. AB - Although the ability of nonsteriodal anti-inflammatory drugs (NSAIDs) to injure the small intestine has been well established in humans and animals, the mechanism involved in this type of injury has yet to be elucidated. The cytokine tumor necrosis factor-alpha (TNF-alpha) has recently been demonstrated to play a critical role in the pathogenesis of NSAID-induced gastric damage. We therefore assessed the possibility that TNF-alpha is similarly involved in the pathogenesis of NSAID-induced small intestinal injury. Administration of multiple doses (n = 4) of diclofenac, but not a single dose, resulted in profound macroscopic damage in the intestine and significantly increased levels of TNF-alpha in intestinal tissue and bile. Pretreatment of rats with a phosphodiesterase inhibitor, pentoxifylline, theophylline, or rolipram, significantly attenuated the macroscopic intestinal ulceration produced by diclofenac administration. However, inhibition of TNF-alpha release with thalidomide or immunoneutralization with a polyclonal antibody directed against TNF-alpha failed to afford any protection. These results suggest that the cytokine TNF-alpha does not play a critical role in NSAID-induced small intestinal injury. Therefore, phosphodiesterase inhibitors mediate their protective effect through a mechanism independent of TNF-alpha synthesis inhibition. PMID- 10516152 TI - Protein meals reduce nausea and gastric slow wave dysrhythmic activity in first trimester pregnancy. AB - First trimester nausea is associated with gastric slow wave dysrhythmias (tachygastria, bradygastria). We tested the roles of meal composition and caloric content on nausea and slow wave rhythm in 14 nauseated pregnant women. Electrogastrography quantified dysrhythmic activity and signal power responses to meals. Symptomatic women reported mild to moderate nausea and exhibited increased dysrhythmias during fasting (P < 0.05). Protein-predominant meals reduced nausea and dysrhythmic activity to greater degrees than equicaloric carbohydrate and fat meals and noncaloric meals (P < 0.05). Meal consistency did not affect symptom responses, although liquid meals decreased dysrhythmias more than solids (P < 0.05). Carbohydrates and fats increased electrogastrographic power to similar degrees as proteins, whereas responses to noncaloric meals were less. In conclusion, protein meals selectively reduce nausea and gastric slow wave dysrhythmias in first trimester pregnancy. Meal consistency is a limited factor in the favorable effects of protein. Electrogastrographic power changes do not explain the symptom response to protein. Thus dietary modulation of gastric myoelectric rhythm with protein supplementation may provide symptomatic benefit in nausea of pregnancy. PMID- 10516153 TI - Measurement of upper esophageal sphincter tone and relaxation during swallowing in premature infants. AB - Upper esophageal sphincter (UES) motor function has not been previously evaluated in premature infants. The motor patterns associated with tonic activity and swallow-related relaxation of the UES were recorded for 1 h after completion of gavage feeding in 11 healthy preterm neonates (postmenstrual age 33-37 wk) with a micromanometric assembly, which included a sleeve sensor specifically adapted for UES recordings. A clearly defined UES high-pressure zone was observed in all premature infants studied. Resting UES pressure ranged from 2.3 to 26.2 mmHg and was higher during periods of irritability and apparent discomfort. During dry swallows, UES pressure relaxed from a resting pressure of 28.2 +/- 4.0 mmHg to a nadir of 1.1 +/- 3.3 mmHg. The mean UES relaxation interval (the time from relaxation onset to relaxation offset) was 0. 31 +/- 0.11 s. We conclude that in premature infants >/=33 wk postmenstrual age the motor mechanisms regulating UES resting pressure and the onset of UES relaxation are well developed. PMID- 10516154 TI - Inhibition of transient LES relaxations and reflux in ferrets by GABA receptor agonists. AB - Transient lower esophageal sphincter (LES) relaxation is the major mechanism of gastroesophageal reflux. This study uses an established ferret model to evaluate GABA(B) receptor agonists' ability to reduce triggering of transient LES relaxations. One hundred sixty manometric/pH studies were performed on 18 conscious ferrets. In untreated animals, intragastric infusion of 25 ml glucose (pH 3.5) led to 2.0 +/- 0.6 reflux episodes over the first 30 min. Twenty-nine of forty-seven reflux episodes occurred during transient LES relaxation, and 18 occurred after downward drifts (<1 mmHg/s) in basal LES pressure. The GABA(B) receptor agonists baclofen (7 micromol/kg ip), CGP-44532, and SKF-97541 (both ED(50) <0.3 micromol/kg) reduced reflux episodes and transient LES relaxations. The putative peripherally selective GABA(B) receptor agonist 3 aminopropylphosphinic acid (80-240 micromol/kg) was ineffective, as was the GABA(A) receptor agonist muscimol (5 micromol/kg). Baclofen's inhibition of transient LES relaxations and reflux was unaffected by low-affinity GABA(B) receptor antagonists CGP-35348 and CGP-36742 at 100 micromol/kg but was reversed by higher-affinity CGP-54626 and CGP-62349 (0.7 micromol/kg) or by CGP-36742 at 200 micromol/kg. Therefore, GABA(B) receptor inhibition of reflux shows complex pharmacology. Our and other data indicate the therapeutic potential for these drugs. PMID- 10516155 TI - Evidence for a feedback inhibition of NO synthesis in enteric synaptosomes via a nitrosothiol intermediate. AB - The exact mechanisms controlling nitric oxide synthase (NOS) activity within enteric neurons are largely unknown. In this study, the effect of exogenous nitric oxide (NO) on NOS activity was investigated in enteric synaptosomes of rat ileum. 3-Morpholinosydnonimine (SIN-1; 10(-4) M) and S-nitroso-N acetylpenicillamine (SNAP; 10(-4) M) significantly inhibited NOS activity by 53% and 48%, respectively. However, superoxide dismutase (SOD; 160 U/ml) as well as the NO scavenger oxyhemoglobin (10(-3) M) did not influence NO donor-induced inhibition. In contrast, the inhibitory effect was antagonized by diethyldithiocarbamate (3 x 10(-4) M), an inhibitor of endogenous Cu/Zn SOD. Inhibition of NOS by exogenous NO was dependent on glutathione (GSH), since the inhibitory effect was augmented in the presence of GSH (5 x 10(-4) M) and antagonized by the GSH-depletor DL-buthionine-SR-sulfoximine (5 x 10(-4) M), suggesting that NO might be protected from extracellular breakdown by reaction with GSH. The reaction product of SIN-1/SNAP and GSH was identified as a nitrosothiol. In the presence of the Cu(+)-chelator neocuproine (10(-5) M), inhibition of NOS by SNAP/SIN-1 was reversed, suggesting that nitrosothiol formation is intermediary. These findings are indicative of a feedback inhibition of enteric NOS, presumably via formation of a nitrosothiol intermediate. PMID- 10516156 TI - Delayed rectifier and Ca(2+)-dependent K(+) currents in human esophagus: roles in regulating muscle contraction. AB - We have examined K(+) channels and their function in human esophageal smooth muscle using perforated patch recording, RT-PCR to identify channel mRNA, and muscle contraction to study the effects of channel blockers. Depolarization revealed at least two types of currents: a 4-aminopyridine (4-AP)-sensitive transient delayed rectifier K(+) (K(V)) and a Ca(2+)-dependent K(+) (K(Ca)) current. K(Ca) current was active at positive potentials and was blocked by tetraethylammonium (TEA), iberiotoxin, and charybdotoxin but was insensitive to 4 AP. The mRNA encoding the gene products of Kv1.2 and Kv1.5 was identified in muscle and dissociated cells, consistent with these channel types contributing to K(V) current. 4-AP increased resting tension of muscle strips, suggesting a role for K(V) in setting the membrane potential. TEA, but not 4-AP, augmented the amplitude and duration of electrically evoked contraction, effects that were abolished by nifedipine. Here we provide the first description of macroscopic K(+) currents in human esophagus. K(V) channels participate in regulation of resting tension, whereas the K(Ca) channel limits depolarization and contraction during excitation. PMID- 10516157 TI - Carrier-mediated uptake of lucifer yellow in skate and rat hepatocytes: a fluid phase marker revisited. AB - Uptake of lucifer yellow (LY), a fluorescent disulfonic acid anionic dye, was studied in isolated skate (Raja erinacea) perfused livers and primary hepatocytes to evaluate its utility as a fluid-phase marker in these cells. However, our findings demonstrated that LY is transported across the plasma membrane of skate hepatocytes largely via carrier-mediated mechanisms. Isolated perfused skate livers cleared 50% of the LY from the recirculating perfusate within 1 h of addition of either 22 or 220 microM LY, with only 4.5 and 9% of the LY remaining in the perfusate after 7 h, respectively. Most of the LY was excreted into bile, resulting in high biliary LY concentrations (1 and 10 mM at the two doses, respectively), indicating concentrative transport into bile canalicular lumen. LY uptake by freshly isolated skate hepatocytes was temperature sensitive, exhibited saturation kinetics, and was inhibited by other organic anions. Uptake was mediated by both sodium-dependent [Michaelis-Menten constant (K(m)), 125 +/- 57 microM; maximal velocity (V(max)), 1.5 +/- 0.2 pmol. min(-1). mg cells(-1)] and sodium-independent (K(m), 207 +/- 55 microM; V(max), 1.7 +/- 0.2 pmol. min(-1). mg cells(-1)) mechanisms. Both of these uptake mechanisms were inhibited by various organic anions and transport inhibitors, including furosemide, bumetanide, sulfobromophthalein, rose bengal, probenecid, N-ethylmaleimide, taurocholate, and p-aminohippuric acid. Fluorescent imaging techniques showed intracellular vesicular compartmentation of LY in skate hepatocyte clusters. Studies in perfused rat livers also indicated that LY is taken up against a concentration gradient and concentrated in bile. LY uptake in isolated rat hepatocytes was saturable, but only at high concentrations, and demonstrated a K(m) of 3.7 +/- 1.0 mM and a V(max) of 1.75 +/- 0.16 nmol. min(-1). mg wet wt( 1). These results indicate that LY is transported into skate and rat hepatocytes and bile largely by carrier-mediated mechanisms, rather than by fluid-phase endocytosis. PMID- 10516158 TI - Kinetics of endothelin-1 binding in the dog liver microcirculation in vivo. AB - Endothelin-1 (ET-1) is a 21-amino acid peptide produced by vascular endothelial cells that acts as a potent constrictor of hepatic sinusoids. Hepatic binding of tracer (125)I-labeled ET-1 was investigated in anesthetized dogs with the multiple-indicator dilution technique with simultaneous measurements of unlabeled immunoreactive ET-1 plasma levels. Despite 80% binding to albumin, tracer (125)I ET-1 was avidly extracted by the liver, with only 15 +/- 6% of the peptide surviving passage through the organ. Exchange of ET-1 between plasma and binding sites, probably located on the surface of liver cells, was quantitatively described by a barrier-limited, space-distributed variable transit time model. Reversible and irreversible parallel binding sites were found. Reversible and irreversible plasma clearances of unbound (125)I-ET-1 were 0.084 +/- 0.033 ml. s( 1). g liver(-1) and 0.17 +/- 0.09 ml. s(-1). g liver(-1), respectively, and the dissociation rate constant for reversible binding was 0.24 +/- 0.12 s(-1). The specific ET(A) receptor antagonist BMS-182874 did not modify binding to either site. The nonspecific ET(A)/ET(B) antagonist LU-224332 dose-dependently reduced irreversible binding only. ET-1 levels in the hepatic vein were significantly lower than in the portal vein but were not different from those in the hepatic artery. The ratio between hepatic vein and portal vein levels (0.64 +/- 0.31) was considerably higher than survival fractions, suggesting a substantial simultaneous release of newly synthesized or stored ET-1 by the liver. These results demonstrate both substantial clearance and production of ET-1 by the intact liver. Hepatic ET-1 clearance is mediated by the ET(B) receptor, with the presence of reversible, nonspecific ET-1 binding at the liver surface PMID- 10516160 TI - Adventitia-dependent influences on vascular function. PMID- 10516162 TI - Mechanism for the effects of extracellular acidification on HERG-channel function. AB - Human ether-a-go-go-related gene (HERG) encodes a K channel similar to the rapid delayed rectifier channel current (I(Kr)) in cardiac myocytes. Modulation of I(Kr) by extracellular acidosis under pathological conditions may impact on cardiac electrical activity. Therefore, we studied the effects of extracellular acidification on I(Kr) function and the underlying mechanism, using HERG expressed in Xenopus oocytes as a model. Acidification [extracellular pH (pH(o)) 8.5-6.5] accelerated HERG deactivation (at -80 mV, the time constant tau of the major component of deactivation was 253 +/- 17, 158 +/- 10, and 65 +/- 5 ms at pH(o) 8.5, 7.5, and 6.5, respectively; n = 7-10 each), with no effects on other gating kinetics except a modest acceleration of recovery from inactivation (at 80 mV, tau of recovery was 4.7 +/- 0.3, 3.8 +/- 0.3, and 1.3 +/- 0.2 ms at pH(o) 8. 5, 7.5, and 6.5, respectively; n = 4-7 each). The following were ruled out as the underlying mechanisms: 1) voltage shift in channel activation, 2) pore blockade by protons, 3) protonation of histidines on the extracellular domain of HERG, 4) acceleration of recovery from C-type inactivation, and 5) interaction between an external H(+) binding site and the cytoplasmic NH(2)-terminal domain (a key determinant of HERG deactivation rate). Extracellular application of diethylpyrocarbonate caused an irreversible acceleration of HERG deactivation and prevented further acceleration by external acidification. Our data suggest that side chains accessible to the extracellular solution mediated the effects of elevating extracellular H(+) concentration on channel deactivation. PMID- 10516161 TI - Role of load in regulating eIF-4F complex formation in adult feline cardiocytes. AB - This study examined whether cardiocyte load increases eIF-4F complex formation. To increase load in vitro, adult feline cardiocytes were electrically stimulated to contract (1 Hz, 5-ms pulses). eIF-4F complex formation, measured by eIF-4G association with eIF-4E, increased 57 +/- 16% after 4 h of contraction compared with controls. eIF-4F complex formation did not increase on electrical stimulation with 2,3-butanedione monoxime (BDM), an inhibitor of active tension. Both insulin and phorbol ester increased eIF-4F complex formation, but these increases were unaffected by BDM. Insulin caused a shift of eIF-4E binding proteins (4E-BPs) into their hyperphosphorylated gamma-isoforms and dissociation of 4E-BPs from eIF-4E. Rapamycin inhibited 4E-BP phosphorylation in response to insulin but had no effect on eIF-4F complex formation. Electrically stimulated contraction caused a partial shift of 4E-BP1 and 4E-BP2 into the gamma-isoforms, but it had no effect on 4E-BP association with eIF-4E. Rapamycin blocked the increase in eIF-4F complex formation in electrically stimulated cardiocytes and depressed contractility. These data indicate that cardiocyte load causes a tension-dependent increase in eIF-4F complex formation that does not require dissociation of 4E-BPs from eIF-4E. PMID- 10516163 TI - Dietary salt increases endothelial nitric oxide synthase and TGF-beta1 in rat aortic endothelium. AB - The amount of NaCl in the diet plays an important role in modulating nitric oxide (NO) synthesis in vivo. In the glomerulus, dietary NaCl also regulates transforming growth factor-beta1 (TGF-beta1) production. We hypothesized that dietary NaCl intake regulated expression of the endothelial isoform of nitric oxide synthase (NOS3) and TGF-beta1 in the aorta. Administration of 8.0% NaCl diet to rats for 7 days did not affect blood pressure but increased steady-state mRNA and protein levels of NOS3 in the arterial wall compared with animals on 0.3% NaCl diet. Northern analysis demonstrated increased steady-state amounts of mRNA of TGF-beta1 in aortas of rats on 8.0% NaCl diet. By ELISA, both total and active TGF-beta1 were increased in these vessel segments. Endothelial denudation of aortic rings reduced active TGF-beta1 secretion to undetectable levels. Addition of a neutralizing antibody to TGF-beta to aortic ring segments attenuated NO production but not to that observed in animals on the 0.3% NaCl diet. The data showed that dietary NaCl intake modulated NOS3 and TGF-beta1 expression in the arterial wall; NOS3 expression was at least partially regulated by endothelial cell production of TGF-beta1. PMID- 10516164 TI - Two types of action potential configuration in single cardiac Purkinje cells of sheep. AB - Membrane potentials and currents of isolated sheep Purkinje and ventricular cells were compared using patch-clamp and microelectrode techniques. In approximately 50% of Purkinje cells, we observed action potentials that showed a prominent phase 1 repolarization and relatively negative plateau (LP cells). Action potential configuration of the remaining Purkinje cells was characterized by little phase 1 repolarization and relatively positive plateau (HP cells). Microelectrode impalement of Purkinje strands also revealed these two types of action potential configuration. In LP cells, the density of L-type Ca(2+) current (I(Ca,L)) was lower, whereas the density of transient outward K(+) current was higher, than in HP cells. Action potentials of HP cells strongly resembled those of ventricular cells. Densities of inward rectifier current and I(Ca,L) were significantly higher in ventricular cells compared with densities in both LP and HP Purkinje cells. Differences in current densities explain the striking differences in action potential configuration and the stimulus frequency dependency thereof that we observed in LP, HP, and ventricular cells. We conclude that LP Purkinje cells, HP Purkinje cells, and ventricular cells of sheep each have a unique action potential configuration. PMID- 10516165 TI - Sympathetic responses to cardiopulmonary vagal afferent stimulation during development. AB - Previous work in our laboratory has demonstrated impairment of cardiopulmonary reflex control of renal sympathetic nerve activity (RSNA) during the newborn period. The present study was designed to test the hypothesis that this delayed maturation is secondary to incomplete central integration of vagal afferent input. Term fetal (135-140 days; n = 6), newborn (3-7 days of age; n = 8), and young adult (6-8 wk old; n = 8) sheep anesthetized with alpha-chloralose underwent vagal afferent nerve stimulation. All animals had undergone prior sinoaortic denervation to eliminate influences from the arterial baroreceptors. After determination of optimal stimulation parameters, RSNA responses to gradual increases in stimulation frequency (1.0-16 Hz) were recorded and compared by one way ANOVA. RSNA decreased progressively with increased frequency of stimulation in all three groups of animals. When comparing the three groups at any given frequency of stimulation, reflex withdrawal of RSNA tended to be more pronounced in newborn lambs (P < 0.05 for 1 and 4 Hz). Heart rate (HR) was also noted to decrease significantly with vagal afferent stimulation in each of the groups, but no significant differences in the reflex decreases in HR were noted among the three groups of animals. These results demonstrate that central integration of vagal afferent input is intact in fetal and newborn sheep. These results suggest that the delayed maturation of cardiopulmonary reflex-mediated changes in RSNA seen early in development appears to depend on intrinsic alterations in baroreceptor function rather than incomplete central integration. PMID- 10516166 TI - NO from smooth muscle cells decreases NOS expression in endothelial cells: role of TNF-alpha. AB - Despite the evidence that cytokines stimulate nitric oxide (NO) production by inducible nitric oxide synthase (iNOS), several reports recently demonstrated that the hypotensive response related to endothelial nitric oxide synthase (eNOS) activity could be inhibited by the same cytokines. The aim of the present work was to analyze whether NO generated by vascular smooth muscle cells (VSMC) could modify eNOS protein expression in endothelial cells. Bovine aortic endothelial cells (BAEC) and bovine VSMC (BVSMC) in coculture were used for the study. Interleukin-1beta (IL-1beta, 10 ng/ml)-treated BVSMC, which expressed iNOS protein, decreased eNOS protein expression in BAEC. The presence of NO antagonists N(omega)-nitro-L-arginine methyl ester (10(-3) mol/l) or N(G) monomethyl-L-arginine (10(-3) mol/l) prevented the decrease in eNOS protein expression induced by IL-1beta-treated BVSMC. Surprisingly, two different NO donors, 3-morpholinosydnonimine (10(-4) mol/l) and S-nitroso-N-acetyl-D,L penicillamine (10(-4) mol/l), failed to modify eNOS expression in BAEC, suggesting the existence of a diffusible mediator released from IL-1beta-treated BVSMC that acts on endothelial cells by reducing eNOS expression. The presence of NO antagonists reduced tumor necrosis factor-alpha (TNF-alpha) production by IL 1beta-stimulated BVSMC. This effect was also produced in the presence of a protein kinase G inhibitor, guanosine-5'-O-(2-thiodiphosphate) trilithium salt. A polyclonal antibody against TNF-alpha prevented eNOS expression in the BAEC-BVSMC coculture. In conclusion, NO by itself failed to modify eNOS protein expression in endothelial cells but increased TNF-alpha generation by IL-1beta-stimulated BVSMC and, in this way, reduced eNOS expression in the endothelium. PMID- 10516167 TI - Hypoxia stimulates proliferation and interleukin-1alpha production in human vascular smooth muscle cells. AB - Several lines of evidence indicate that hypoxia is a stimulus to vascular smooth muscle cell (VSMC) proliferation that occurs in pulmonary hypertension. The present study tested the hypothesis that low O(2) tension directly stimulates human VSMC proliferation by inducing them to produce interleukin (IL)-1, a potent autocrine growth factor for human VSMC. Human VSMC derived from pulmonary artery, aorta, or saphenous vein were incubated in either a normal in vitro O(2) environment (20% O(2)) or in chambers containing low (approximately 1%) or moderate (5%) O(2). Levels of IL-1alpha and IL-1beta mRNA increased in human VSMC after 24-48 h of incubation in low O(2) compared with levels in normoxic cells and then decreased upon subsequent reoxygenation. Levels of cell-associated IL 1alpha also increased progressively after 24-48 h in low O(2); however, detectable IL-1alpha was not released from the cells in the media. IL-1beta was detectable in cell lysates and supernatants; however, the levels were not affected by exposure to low O(2). mRNA encoding for tumor necrosis factor-alpha (TNF-alpha), a related cytokine and VSMC mitogen, was not detectable in human VSMC exposed to either low or 20% O(2). Proliferation of human VSMC was not stimulated during exposure to low O(2), despite the fact that cells remained responsive to the mitogenic effect of exogenous IL-1. Interestingly, however, exposure to 5% O(2) enhanced proliferation of human VSMC but did not induce IL 1alpha production. Inhibition of IL-1 binding to the type I IL-1 receptor by exogenous addition of IL-1-receptor antagonist (10 microgram/ml) did not attenuate the proliferation rates of human VSMC incubated in 20%, 5%, or low O(2) or in human VSMC that were reoxygenated after exposure to low O(2). These results demonstrate two direct and distinct effects of hypoxia on VSMC. Exposure to moderately low O(2) tension induces VSMC proliferation, independent of IL-1, whereas exposure to very low O(2) tension induces production of IL-1alpha. PMID- 10516168 TI - Evidence for an interaction between adducin and Na(+)-K(+)-ATPase: relation to genetic hypertension. AB - Adducin point mutations are associated with genetic hypertension in Milan hypertensive strain (MHS) rats and in humans. In transfected cells, adducin affects actin cytoskeleton organization and increases the Na(+)-K(+)-pump rate. The present study has investigated whether rat and human adducin polymorphisms differently modulate rat renal Na(+)-K(+)-ATPase in vitro. We report the following. 1) Both rat and human adducins stimulate Na(+)-K(+)-ATPase activity, with apparent affinity in tens of nanomolar concentrations. 2) MHS and Milan normotensive strain (MNS) adducins raise the apparent ATP affinity for Na(+)-K(+) ATPase. 3) The mechanism of action of adducin appears to involve a selective acceleration of the rate of the conformational change E(2) (K) --> E(1) (Na) or E(2)(K). ATP --> E(1)Na. ATP. 4) Apparent affinities for mutant rat and human adducins are significantly higher than those for wild types. 5) Recombinant human alpha- and beta-adducins stimulate Na(+)-K(+)-ATPase activity, as do the COOH terminal tails, and the mutant proteins display higher affinities than the wild types. 6) The cytoskeletal protein ankyrin, which is known to bind to Na(+)-K(+) ATPase, also stimulates enzyme activity, whereas BSA is without effect; the effects of adducin and ankyrin when acting together are not additive. 7) Pig kidney medulla microsomes appear to contain endogenous adducin; in contrast with purified pig kidney Na(+)-K(+)-ATPase, which does not contain adducin, added adducin stimulates the Na(+)-K(+)-ATPase activity of microsomes only about one half as much as that of purified Na(+)-K(+)-ATPase. Our findings strongly imply the existence of a direct and specific interaction between adducin and Na(+)-K(+) ATPase in vitro and also suggest the possibility of such an interaction in intact renal membranes. PMID- 10516170 TI - Cardiac autonomic responses to volume overload in normal subjects and in patients with dilated cardiomyopathy. AB - This study evaluated the effects of acute isotonic volume expansion on heart rate variability (HRV) in 10 patients with dilated cardiomyopathy (DCM) and in 10 age- and sex-matched normal volunteers. Echocardiographic left ventricular volumes and HRV measurements by continuous Holter recording were assessed at baseline, at 60 and 120 min during intravenous saline load (0.9% NaCl, 0.25 ml. kg(-1). min(-1)), and 60 min after infusion was terminated. Data analysis was performed by repeated measures ANOVA. After volume expansion, left ventricular ejection fraction increased (F = 9.8; P < 0.001) in normal subjects and decreased (F = 8.7; P < 0.001) in DCM patients. During volume expansion a significant difference was also detectable between the two groups in root-mean-square successive difference (F = 25.2; P < 0.001), percentage of differences between successive normal R-R intervals >50 ms (F = 97.6; P < 0.001), high-frequency power (F = 50.1; P < 0.001), and low-frequency power (F = 41.6; P < 0.001), all of which reflect parasympathetic modulation of heart rate; in fact, these measurements increased in normal subjects and decreased in DCM patients. In normal subjects, the increase in HRV measurements during volume expansion suggests a parasympathetic activation, mediated by stimulation of cardiopulmonary and arterial mechanoreceptors. On the contrary, in DCM patients the parasympathetic withdrawal, already detectable at baseline, increases during volume expansion. PMID- 10516169 TI - A presynaptic mechanism contributes to depression of autonomic signal transmission in NTS. AB - With increasing frequencies of autonomic afferent input to the nucleus tractus solitarii (NTS), postsynaptic responses are depressed. To test the hypothesis that a presynaptic mechanism contributes to this frequency-dependent depression, we used whole cell, voltage-clamp recordings in an NTS slice. First, we determined whether solitary tract stimulation (0.4-24 Hz) resulted in frequency dependent depression of excitatory postsynaptic currents (EPSCs) in second-order neurons. Second, because decreases in presynaptic glutamate release result in a parallel depression of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) and N-methyl-D-aspartic acid (NMDA) receptor-mediated components of EPSCs, we determined whether the magnitude, time course, and recovery from the depression were the same in both EPSC components. Third, to determine whether AMPA receptor desensitization contributed, we examined the depression during cyclothiazide. EPSCs decreased in a frequency-dependent manner by up to 76% in second- and 92% in higher-order neurons. AMPA and NMDA EPSC components were depressed with the same magnitude (by 83% and 83%) and time constant (113 and 103 ms). The time constant for the recovery was also not different (1.2 and 0.8 s). Cyclothiazide did not affect synaptic depression at >/=3 Hz. The data suggest that presynaptic mechanism(s) at the first NTS synapse mediate frequency dependent synaptic depression. PMID- 10516171 TI - EP receptor-mediated inhibition by prostaglandin E(1) of cardiac L-type Ca(2+) current of rabbits. AB - Prostaglandin E(1) (PGE(1)) has cardioprotective effects on the ischemic reperfused heart. To clarify the mechanisms underlying the protective action of PGE(1) on myocardium, we examined the effect of PGE(1) on the L-type Ca(2+) current (I(Ca)) using single atrial cells from rabbits. PGE(1) did not show a significant effect on basal I(Ca) but inhibited the I(Ca) prestimulated by isoproterenol (Iso, 30 nM). This inhibition was concentration dependent (EC(50) = 0.027 microM). Both sulprostone, a specific PGE receptor subtype (EP(1) and EP(3)) agonist, and 11-deoxy-PGE(1), an EP(3) agonist, inhibited the Iso stimulated I(Ca), similar to PGE(1). Pretreatment with pertussis toxin (PTX) abolished the PGE(1) inhibition of I(Ca). Both the application of forskolin plus IBMX and intracellular dialysis with 8-bromoadenosine 3',5'-cyclic monophosphate eliminated the effect of PGE(1). PGE(1) did not show any further inhibition of I(Ca) when the effect of Iso was almost fully antagonized by acetylcholine. Methylene blue (guanylate cyclase inhibitor), KT-5823 (cGMP-dependent protein kinase inhibitor), and erythro-9-(2-hydroxy-3-nonyl)adenine (type II phosphodiesterase inhibitor) did not significantly change the inhibitory effect of PGE(1). These findings suggest that 1) PGE(1) inhibits Iso-stimulated I(Ca) by binding to the EP(3) receptor and 2) the PTX-sensitive and cAMP-dependent pathway is involved in the PGE(1) inhibition of I(Ca), but the nitric oxide-cGMP dependent pathway is not. The PGE(1)-induced antiadrenergic effect shown in this study may contribute to the PGE(1) protection of myocardium against ischemia. PMID- 10516172 TI - Dynamic adaptation of cardiac oxidative phosphorylation is not mediated by simple feedback control. AB - The classic idea about regulation of cardiac oxidative phosphorylation (OxPhos) was that breakdown products of ATP (ADP and P(i)) diffuse freely to the mitochondria to stimulate OxPhos. On the basis of this metabolic feedback control system, the response time of OxPhos (t(mito)) is predicted to be inversely proportional to the mitochondrial aerobic capacity (MAC). We determined t(mito) during steps in heart rate in isolated perfused rabbit hearts (n = 16) before and after reducing MAC with nonsaturating doses of oligomycin. The reduction of MAC was quantified in mitochondria isolated from each perfused heart, dividing oligomycin-sensitive, ADP-stimulated state 3 respiration by oligomycin insensitive uncoupled respiration. The t(mito) to heart rate steps from 60 to 70 and 80 beats/min was 5. 6 +/- 0.6 and 7.2 +/- 0.8 s (means +/- SE) and increased an estimated 34 and 40% for a 50% decrease in MAC (P < 0.05), respectively, which is much less than the 100% predicted by the feedback hypothesis. For steps to 100 or 120 beats/min, t(mito) was 8.3 +/- 0.5 and 11.2 +/- 0.6 s and was not reduced with decreases in MAC (P > 0.05). We conclude that immediate feedback control by quickly diffusing ADP and P(i) cannot explain the dynamic regulation of cardiac OxPhos. Because calcium entry into the mitochondria also cannot explain the first fast phase of OxPhos activation, we propose that delay of the energy-related signal in the cytoplasm dominates the response time of OxPhos. PMID- 10516173 TI - Determinants of mechanical properties in the developing ovine thoracic aorta. AB - We previously reported changes in mechanical properties and collagen cross linking of the ovine thoracic aorta during perinatal development and postnatal maturation, and we now report changes in biochemical composition (elastin, collagen, and DNA contents per mg wet wt) over the same developmental intervals. A comparison of results from the present and previous studies has yielded novel and important observations concerning the relationship between aortic mechanics and composition during maturation. Developmental changes in aortic incremental elastic modulus at low tensile stress (E(low)) closely followed changes in relative elastin content (i.e., per mg wet wt). An 89% increase in E(low) during the perinatal period was associated with a 69% increase in relative elastin content, whereas neither variable changed during postnatal life. Incremental elastic modulus at high tensile stress (E(high)) did not change during the perinatal period but increased 88% during postnatal life. This pattern closely paralleled changes in collagen cross-linking index, which did not change perinatally but almost doubled postnatally. In contrast, relative collagen content (per mg wet wt) increased only slightly from fetal to adult life, a trend that was unrelated to aortic mechanics. Substantial, progressive decreases in measures of wall viscosity (pressure wave attenuation coefficient and viscoelastic phase angle) from fetal to adult life followed the pattern observed for relative DNA (smooth muscle cell) content (per mg wet wt). Our findings suggest that accumulation of elastin per milligram wet weight contributes most to developmental changes in E(low), change in collagen cross-linking is the primary determinant of developmental changes in E(high), and cell accumulation contributes most to developmental changes in wall viscosity. PMID- 10516175 TI - Molecular and functional distributions of chloride conductances in rabbit ventricle. AB - The regulation of cardiac electrical activity is critically dependent on the distribution of ion channels in the heart. For most ion channels, however, the patterns of distribution and what regulates these patterns are not well characterized. The most likely candidates for the genes that encode the cAMP- and swelling-activated chloride conductances in the heart are an alternatively spliced variant of CFTR and ClC-3, respectively. In this study we have 1) measured the density of CFTR and ClC-3 mRNA levels across the left ventricular free wall (LVFW) of the rabbit heart using in situ hybridization and 2) measured the corresponding current density of cAMP- and swelling-activated chloride channels in myocytes isolated from subepicardial, midmyocardial, and subendocardial regions of the LVFW. There was a highly significant gradient in the whole cell slope conductance of cAMP-activated chloride currents; normalized slope conductance at 0 mV was 15.7 +/- 1.8 pS/pF (n = 9) in subepicardial myocytes, 7.8 +/- 1.5 pS/pF (n = 11) in midmyocardial myocytes, and 4.9 +/- 1.1 pS/pF (n = 9) in subendocardial myocytes. The level of CFTR mRNA was closely correlated with the density of cAMP-activated chloride conductances in different regions of the heart, with the level of CFTR mRNA being three times higher in the subepicardium than in the subendocardium. The whole cell slope conductance of swelling-activated chloride channel activity, measured 3-5 min after the commencement of cell swelling, was higher in myocytes isolated from the subepicardium than in myocytes isolated from the midmyocardium or subendocardium. In contrast, there was a uniform expression of ClC-3 mRNA across the LVFW of the rabbit heart. These results suggest that the control of gene expression is an important contributor in regulating the distribution of cAMP-activated chloride channels in the rabbit heart but that it may be less important for the swelling activated chloride channels. PMID- 10516174 TI - Alpha-adrenergic vasoreactivity of canine intrapulmonary bronchial arteries in pacing-induced heart failure. AB - We hypothesized that pacing-induced congestive heart failure alters alpha adrenergic constriction in intrapulmonary bronchial arteries. Cumulative dose responses to norepinephrine (NE), phenylephrine (PE), acetylcholine (ACh) and sodium nitroprusside (SNP) were determined in pressurized vessel segments. ED(50) values for NE and PE were higher for control (-5.34 +/- 0.09 and -4.27 +/- 0.08 M, respectively) vs. paced (-5.73 +/- 0.10 and -5.06 +/- 0.28 M, respectively) groups. Prazosin increased the ED(50) values for NE and PE in both control and paced groups. Yohimbine decreased NE ED(50) in the control group only. Endothelium removal or nitric oxide synthase (NOS) inhibition decreased control but not paced NE ED(50). Maximum vasodilation and sensitivity (i.e., -ED(50) values) were decreased for ACh but were similar for SNP in paced vs. control groups. Secondary segments were more reactive than paired primary segments in both groups, although pacing effects on ED(50) were unrelated to branching order. In conclusion, adrenergic constriction of canine intrapulmonary bronchial arteries is predominantly mediated via alpha(1)-adrenoreceptors and is enhanced after pacing. Endothelium-derived relaxing factor(s) normally opposes alpha adrenergic vasoconstriction but not after pacing in this vasculature. PMID- 10516176 TI - Regulation of respiration in myocardium in the transient and steady state. AB - 1H/(31)P NMR has followed the metabolic response to increased work in the glucose and pyruvate-perfused rat myocardium during a heart cycle and at the steady state. With electrical pacing and dobutamine, the heart O(2) consumption increases by 56%. The phosphocreatine (PCr) level initially declines, but recovers within 15 min to its control level; the oxymyoglobin (MbO(2)) saturation decreases by 15%. Because the MbO(2) signal reflects the intracellular PO(2), the capillary-to-cell O(2) gradient has increased to match the increased O(2) need. However, no transient metabolic fluctuation is observed in either PCr or MbO(2) throughout the entire cardiac cycle in both glucose and pyruvate-/glucose perfused hearts. No systolic-diastolic variation is detectable under either high workload or hypoxic conditions. The results reveal that neither O(2) nor ADP is regulating respiration under increased energy demand in the steady or transient state. PMID- 10516177 TI - Regional alterations in SR Ca(2+)-ATPase, phospholamban, and HSP-70 expression in chronic hibernating myocardium. AB - We sought to identify mechanisms for chronic dysfunction in hibernating myocardium. Pigs were instrumented with a left anterior descending artery stenosis for 3 mo. Angiography demonstrated high-grade stenoses and hibernating myocardium with 1) severe anterior hypokinesis (P < 0.001 vs. shams), 2) reduced subendocardial perfusion [0.73 +/- 0.05 (SE) vs. 1.01 +/- 0.06 ml. min(-1). g(-1) in normal, P < 0.001], and 3) critically reduced adenosine flow (1.0 +/- 0.17 vs. 3.84 +/- 0.26 ml. min(-1). g(-1) in normal, P < 0.001). Histology did not reveal necrosis. Northern blot analysis of hibernating myocardium demonstrated regional downregulation in mRNAs for sarcoplasmic reticulum (SR) proteins phospholamban (0.76 +/- 0.08 vs. 1.07 +/- 0.06, P < 0.02) and SR Ca(2+)-ATPase (0.83 +/- 0.06 vs. 1.02 +/- 0.06, P < 0.05) with no change in calsequestrin (1.08 +/- 0.06 vs. 0.96 +/- 0.05, P = not significant). Heat shock protein (HSP)-70 mRNA was regionally induced in hibernating myocardium (2.4 +/- 0.3 vs. 1.0 +/- 0.11, P < 0.01). Directionally similar changes were confirmed by Western blot analysis of respective proteins. Our results indicate that hibernating myocardium exhibits a molecular phenotype that on a regional basis is similar to end-stage ischemic cardiomyopathy. This supports the hypothesis that SR dysfunction from reversible ischemia may be an early defect in the progression of left ventricular dysfunction. PMID- 10516179 TI - Alteration of gene expression for glycolytic enzymes in aerobic and ischemic myocardium. AB - The purpose of this report was to describe mRNA abundance for the glycolytic enzymes glyceraldehyde-3-phosphate dehydrogenase (GAPDH), pyruvate kinase, and pyruvate dehydrogenase in ischemic and adjacent aerobic myocardium. Mechanical, metabolic, and mRNA data were acquired in a pig model of regulated coronary flow using extracorporeal perfusion. Trials of coronary hypoperfusion included sustained and intermittent exposures of acute ischemia with or without reperfusion. These were compared with a chronic 4-day model of partial coronary stenosis. In ischemic tissues, levels of mRNA, normalized by mRNA for beta-actin, were increased over control values for GAPDH (range 2.7- to 4.6-fold), pyruvate kinase (2.9-fold), and pyruvate dehydrogenase (2.1-fold). It is of interest that increases in mRNA levels over control values were also observed in adjacent aerobic heart muscle from intervention hearts, including 3.6- to 4.5-fold elevations in message for GAPDH and a 2.1-fold increase in signal for pyruvate dehydrogenase. Augmentation in mRNA abundance occurred in as short a time as 40 min of ischemia and was maintained for as long as 4 days in partial coronary stenosis. Whether the former time was of an interval sufficient to affect protein production is problematic, but the latter time was ample to influence enzyme concentration, which may in turn have regulated glycolysis in this condition. PMID- 10516178 TI - Beta-blockade improves adjacent regional sympathetic innervation during postinfarction remodeling. AB - The effect of beta-blockade on left ventricular (LV) remodeling, when added to angiotensin-converting enzyme inhibition (ACEI) after anterior myocardial infarction (MI), is incompletely understood. On day 2 after coronary ligation induced anteroapical infarction, 17 sheep were randomized to ramipril (ACEI, n = 8) or ramipril and metoprolol (ACEI-beta, n = 9). Magnetic resonance imaging was performed before and 8 wk after MI to measure changes in LV end-diastolic, end systolic, and stroke volume indexes, LV mass index, ejection fraction (EF), and regional percent intramyocardial circumferential shortening. (123)I-labeled m iodobenzylguanidine (MIBG) and fluorescent microspheres before and after adenosine were infused before death at 8 wk post-MI for quantitation of sympathetic innervation, blood flow, and blood flow reserve in adjacent and remote noninfarcted regions. Infarct size, regional blood flow, blood flow reserve, and the increase in LV mass and LV end-diastolic and end-systolic volume indexes were similar between groups. However, EF fell less over the 8-wk study period in the ACEI-beta group (-13 +/- 11 vs. -22 +/- 4% in ACEI, P < 0.05). The ratio of adjacent to remote region (123)I-MIBG uptake was greater in ACEI-beta animals than in the ACEI group (0.93 +/- 0.06 vs. 0.86 +/- 0.07, P < 0.04). When added to ACE inhibition after transmural anteroapical MI, beta-blockade improves EF and adjacent regional sympathetic innervation but does not alter LV size. PMID- 10516180 TI - NO mediates postjunctional inhibitory effect of neurogenic ACh in guinea pig small intestinal microcirculation. AB - The present study was designed to evaluate the role of the endothelium as an effector organ of neurally mediated inhibition of vascular tone. Acetylcholine (ACh), either released by stimulation of the submucosal ganglia or applied exogenously, inhibited phenylephrine (PE)-induced constrictions in arterioles of the guinea pig intestinal submucosa. N(G)-monomethyl-L-arginine (L-NMMA), an inhibitor of nitric oxide (NO) synthesis, attenuated the response to superfused ACh by 74% compared with 94% attenuation obtained with N(G)-nitro-L-arginine (L NNA). L-NNA attenuated the response to neurally released ACh by 98% and that to iontophoretically applied ACh by 92%. L-Arginine reversed the effects of both L NMMA and L-NNA. Functional integrity of the endothelium was essential for the neurally mediated inhibition of PE-induced constrictions. However, neurogenic inhibition of neurally evoked constrictions was preserved despite endothelial disruption. It was concluded that at the postjunctional level, the mechanism of action of neurally released ACh was almost exclusively via a NO-dependent pathway, with the source of NO being the vascular endothelium. PMID- 10516181 TI - Enhanced beta-receptor-mediated vasorelaxation in hypoxic porcine coronary artery. AB - To investigate the beta-adrenoceptor-mediated responses in hypoxic coronary arteries, we studied the effect of isoproterenol (Iso) on isolated porcine coronary arteries contracted with endothelin-1 in media aerated with 0, 5, 7.5, and 95% O(2). The concentration-response curve of Iso was significantly shifted to the left by hypoxia (0 and 5% O(2)). In oxygenated and hypoxic arteries, 3 x 10(-8), 10(-6), and 10(-5) M Iso significantly increased the contents of cAMP. However, there was no difference in the increases of cAMP content induced by 3 x 10(-8) M Iso between oxygenated and hypoxic arteries. The content of cAMP induced by high concentrations of Iso (10(-6) and 10(-5) M) was significantly larger in hypoxic than in oxygenated arteries. Furthermore, the potentiation by hypoxia of the Iso-induced vasorelaxation was inhibited by glibenclamide and depolarization by KCl, but not by removal of endothelium and indomethacin. The vasodilatory response to forskolin and dibutyryl cAMP was unaffected by hypoxia. We conclude that activation of the ATP-sensitive K(+) channel may account for the potentiation of the response to Iso in hypoxic coronary arteries. PMID- 10516182 TI - Involvement of ATP-sensitive K(+) channels in spontaneous activity of isolated lymph microvessels in rats. AB - Physiological roles of ATP-sensitive K(+) channels for spontaneous activity in isolated rat mesenteric lymph microvessels (maximum diameter approximately 80-150 microm) were investigated. The lymph microvessels were cannulated with glass micropipettes and pressurized at a perfusion pressure of 6 cmH(2)O. Changes in the diameter and frequency of spontaneous contractions in the lymphatics were measured with videomicroscopy. Pinacidil (K(+)-channel opener) inhibited the spontaneous activity. In the presence of glibenclamide (selective ATP-sensitive K(+)-channel blocker; 10(-7) and 10(-6) M) and tetraethylammonium (TEA; nonselective K(+)-channel blocker; 10(-4) and 10(-3) M), the pinacidil-induced inhibition of the spontaneous contractions in lymph microvessels was significantly reversed. Glibenclamide and TEA themselves, however, did not affect the frequency of spontaneous activity in the lymph microvessels. These results suggest that ATP-sensitive K(+) channels are involved in the regulation of spontaneous activity in the smooth muscles of isolated lymph microvessels of rat mesenteries. PMID- 10516183 TI - Cerebrovascular reactivity to CO(2) and hypotension after mild cortical impact injury. AB - Cerebrovascular reactivity to CO(2) or hypotension was studied in vivo and in vitro [pressurized arteries ( approximately 82 micrometer) and arterioles ( approximately 30 micrometer)] at 1 h after mild controlled cortical impact (CCI) injury in rats. The cortical perfusion response [assessed using laser-Doppler flowmetry (LDF)] to altered CO(2) was diminished (up to 81%) after mild CCI injury. The responses to CO(2) alterations in arteries and arterioles isolated from the injured cortex were similar to responses in vessels isolated from sham injured animals. After mild CCI injury, the autoregulatory response to hypotension (measured using LDF) was maintained or even enhanced, depending on the method used to measure the response. Vessels isolated from the injury site showed a response to changes in pressure similar to that in vessels isolated from sham-injured rats. We conclude that mild CCI injury produces complicated alterations in cerebrovascular control. Whereas the autoregulatory response to hypotension was maintained or even enhanced, the in vivo vascular response to CO(2) was severely compromised. The altered response to CO(2) was not caused by an intrinsic vascular perturbation but rather an altered milieu after mild CCI injury. PMID- 10516184 TI - Ca(2+)-activated Cl(-) current can be triggered by Na(+) current-induced SR Ca(2+) release in rabbit ventricle. AB - The Ca(2+)-activated Cl(-) current [I(Cl(Ca))] contributes to the repolarization of the cardiac action potential under physiological conditions. I(Cl(Ca)) is known to be primarily activated by Ca(2+) release from the sarcoplasmic reticulum (SR). L-type Ca(2+) current [I(Ca(L))] represents the major trigger for Ca(2+) release in the heart. Recent evidence, however, suggests that Ca(2+) entry via reverse-mode Na(+)/Ca(2+) exchange promoted by voltage and/or Na(+) current (I(Na)) may also play a role. The purpose of this study was to test the hypothesis that I(Cl(Ca)) can be induced by I(Na) in the absence of I(Ca(L)). Macroscopic currents and Ca(2+) transients were measured using the whole cell patch-clamp technique in rabbit ventricular myocytes loaded with Indo-1. Nicardipine (10 microM) abolished I(Ca(L)) at a holding potential of -75 mV as tested in Na(+)-free external solution. In the presence of 131 mM external Na(+) and in the absence of I(Ca(L)), a 4-aminopyridine-resistant transient outward current was recorded in 64 of 81 cells accompanying a phasic Ca(2+) transient. The current reversed at -42. 0 +/- 1.3 mV (n = 6) and at +0.3 +/- 1.4 mV (n = 6) with 21 and 141 mM of internal Cl(-), respectively, similar to the predicted reversal potential with low intracellular Cl(-) concentration ([Cl(-)](i)) (-47.8 mV) and high [Cl(-)](i) (-1.2 mV). Niflumic acid (100 microM) inhibited the current without affecting the Ca(2+) signal (n = 8). Both the current and Ca(2+) transient were abolished by 10 mM caffeine (n = 6), 10 microM ryanodine (n = 3), 30 microM tetrodotoxin (n = 9), or removal of extracellular Ca(2+) (n = 6). These properties are consistent with those of I(Cl(Ca)) previously described in mammalian cardiac myocytes. We conclude that 1) I(Cl(Ca)) can be recorded in the absence of I(Ca(L)), and 2) I(Na)-induced SR Ca(2+) release mechanism is also present in the rabbit heart and may play a physiological role in activating the Ca(2+)-sensitive membrane Cl(-) conductance. PMID- 10516185 TI - Computer-controlled heart rate increase by isoproterenol infusion: mathematical modeling of the system. AB - The purpose of this study was mathematical modeling of the heart rate (HR) response to isoproterenol (Iso) infusion. We developed a computerized system for the controlled increase of HR by Iso, based on a modified proportional-integral controller. HR was measured in conscious, freely moving rats. We found that the steady-state HR can be described as a hyperbolic power function of the steady state Iso flow rate. This dependence was coupled with a first-order difference equation to form a pharmacodynamic model that reliably describes the relationship between HR and Iso flow for any arbitrary form of Iso flow function. In simulation studies, we showed that the model continued to follow the HR curve from real-time experiments far beyond the initial "learning interval" from which its parameters were calculated. Our results suggest that the predictive ability and the simplicity of calculating the parameters render this pharmacodynamic model appropriate for use within future advanced, model-based, adaptive control systems and as a part of larger cardiovascular models. PMID- 10516186 TI - Miconazole represses CO(2)-induced pial arteriolar dilation only under selected circumstances. AB - Previous experimental findings have led to the suggestion that guanosine 3',5' cyclic monophosphate (cGMP) plays a permissive role in hypercapnic cerebral vasodilation. However, we recently reported that the technique used to reveal a permissive role for cGMP [cGMP repletion in the presence of nitric oxide synthase (NOS) inhibition] created a situation where CO(2) reactivity was normalized but where different mechanisms (i.e., K(+) channels) participated in the response. In the present study, we examined whether that nascent K(+)-channel dependence is related in any way to an increase in the influence of the miconazole-inhibitable cytochrome P-450 epoxygenase pathway. Using intravital microscopy and a closed cranial window system in adult rats, we measured pial arteriolar diameters during normo- and hypercapnia, first in the absence and then in the presence of a neuronal NOS (nNOS) inhibitor [7-nitroindazole (7-NI)]. This was followed by suffusion of a cGMP analog and then cGMP plus miconazole. Separate groups of rats were used to evaluate whether miconazole either alone or in the presence of 8 bromoguanosine 3', 5'-cyclic monophosphate (8-BrcGMP) or its vehicle (0.1% ethanol) had any effect on CO(2) reactivity and whether miconazole affected K(+) channel opener-induced dilations. Hypercapnic (arterial PCO(2), congruent with65 mmHg) pial arteriolar dilations, as expected, were reduced by 70-80% with 7-NI and restored with cGMP repletion. CO(2) reactivity was again attenuated after miconazole introduction. Miconazole, with and without 8-BrcGMP, and its vehicle had no influence on pial arteriolar CO(2) reactivity in the absence of nNOS inhibition combined with cGMP repletion. Miconazole alone also did not affect vasodilatory responses to K(+)-channel openers. Thus present results suggest that the nascent K(+)-channel dependence of the hypercapnic response found in our earlier study may be related to increased epoxygenase activity. The specific reasons why the pial arteriolar CO(2) reactivity gains a K(+)-channel and epoxygenase dependence only under conditions of nNOS inhibition and cGMP restoration remain to be identified. These findings again call into question the interpretations applied to data collected in studies evaluating potential permissive actions of cGMP or NO. PMID- 10516187 TI - Origins of heart rate variability: relationship of heart rate burst morphology to work duration and load. AB - We are developing a lexicon of specific heart period changes, or lexons, that recur frequently and whose physiological meaning can be read into ambulatory electrocardiogram (ECG). The transient, reversible "burst" of tachycardia induced by exercise initiation can also be seen on ambulatory ECG. We hypothesized that burst morphology depended on the work that preceded it and on baroreceptor activation. Ten subjects with mean age 38 yr (range 17-69 yr) underwent two protocols of semisupine cycling in which load and duration were varied. Burst duration increased with longer cycling times (median values of 18.0, 25.5, and 23.7 s with 1, 3, and 5 s of cycling, respectively; P = 0.033). Burst shape as assessed by heart period exponential decay constant and burst magnitude did not change. To assess the impact of workload, subjects cycled for 5 s at loads of 0, 25, 50, and 75 W. No significant differences were seen in burst duration, burst magnitude, or burst shape. Tachycardia preceded hypotension by 4.6 +/- 2.2 s, which is inconsistent with baroreceptor involvement in the onset of burst tachycardia. Because burst morphology is a nearly quantal response to the initiation of exercise, the presence of a burst on an ambulatory ECG implies the onset of exercise. PMID- 10516188 TI - Depolarization-mediated inhibition of Ca(2+) entry in endothelial cells. AB - The effect of extracellular Cl(-) in regulating ACh-induced Ca(2+) entry into freshly isolated rabbit aortic endothelial cells was studied using Ca(2+) sensitive fluorescence microscopy and patch-clamp electrophysiology. After ACh caused transient Ca(2+) release in Ca(2+)-free medium, readdition of 3 mM Ca(2+) to the bath maintained Ca(2+) entry. Removal of extracellular Cl(-) abolished the plateau phase in Ca(2+) signal and inhibited divalent cation entry. However, in the presence of the K(+) ionophore valinomycin, removal of Cl(-) had no effect on the Ca(2+) plateau. Under current-clamp conditions, substitution of gluconate for Cl(-) induced membrane depolarization. Under voltage clamp, with CsCl in the pipette, ACh activated a slowly developing Cl(-) current, which was blocked by SITS and 5-nitro-2-(3-phenylpropylamino)benzoic acid. Varying the membrane potential by utilizing different extracellular K(+) concentrations in the presence of 5 microM valinomycin demonstrated that depolarization blocked ACh stimulated Mn(2+) entry. These data suggest that ACh-induced Ca(2+) entry in freshly isolated endothelial cells requires the presence of extracellular Cl(-) to maintain a polarized membrane potential. PMID- 10516189 TI - Loss of GSH and thiol enzymes in endothelial cells exposed to sublethal concentrations of hypochlorous acid. AB - We investigated the effect of sublethal concentrations of hypochlorous acid (HOCl) on intracellular thiol groups. Exposure of human umbilical vein endothelial cells to HOCl caused a decrease in cell viability, with concentrations of /=12 h. The blockade of JNK activity by using the negative interfering mutant JNK(K-R) markedly decreased the apoptosis induced by colchicine. Exposure of bovine aortic endothelial cells to laminar shear stress (12 dyn/cm(2)) caused a transient (<2 h) activation of JNK, and there was no induction of apoptosis. The sustained activation of JNK may play a significant role in the apoptosis induced by colchicine. PMID- 10516200 TI - Impaired wound healing and angiogenesis in eNOS-deficient mice. AB - A role for nitric oxide (NO) in wound healing has been proposed; however, the absolute requirement of NO for wound healing in vivo and the contribution of endothelial NO synthase (eNOS) have not been determined. Experiments were carried out using eNOS gene knockout (KO) mice to determine the requirement for eNOS on wound closure and wound strength. Excisional wound closure was significantly delayed in the eNOS KO mice (29.4 +/- 2.2 days) compared with wild-type (WT) controls (20.2 +/- 0.4 days). At 10 days, incisional wound tensile strength demonstrated a 38% reduction in the eNOS KO mice. Because effective wound repair requires growth factor-stimulated angiogenesis, in vitro and in vivo angiogenesis assays were performed in the mice to assess the effects of eNOS deficiency on angiogenesis. Endothelial cell sprouting assays confirmed in vitro that eNOS is required for proper endothelial cell migration, proliferation, and differentiation. Aortic segments harvested from eNOS KO mice cultured with Matrigel demonstrated a significant reduction in endothelial cell sprouting and [(3)H]thymidine incorporation compared with WT mice at 5 days. Capillary ingrowth into subcutaneously implanted Matrigel plugs was significantly reduced in eNOS KO mice (2.67 +/- 0.33 vessels/plug) compared with WT mice (10.17 +/- 0.79 vessels/plug). These results clearly show that eNOS plays a significant role in facilitating wound repair and growth factor-stimulated angiogenesis. PMID- 10516201 TI - ACE inhibition improves cardiac NE uptake and attenuates sympathetic nerve terminal abnormalities in heart failure. AB - Cardiac sympathetic nerve terminal dysfunction plays an important role in the downregulation of myocardial beta-adrenoceptors in heart failure. To determine whether chronic angiotensin-converting enzyme (ACE) inhibition improved cardiac sympathetic nerve terminal function and hence increased myocardial beta adrenergic responsiveness, we administered ACE inhibitors to dogs with chronic right-sided heart failure (RHF) produced by tricuspid avulsion and pulmonary artery constriction. The RHF animals exhibited fluid retention, elevated right heart filling pressures, blunted inotropic response to isoproterenol, and reduced beta-adrenoceptor density. These changes were accompanied by decreases in right ventricular norepinephrine (NE) uptake and neuronal NE histofluorescence and tyrosine hydroxylase immunoreactive profiles. ACE inhibitors had no effect on the production of heart failure but greatly reduced the attenuation of cardiac NE uptake, neuronal NE histofluorescence, and tyrosine hydroxylase immunoreactive profiles. ACE inhibition also improved the inotropic response to isoproterenol and restored myocardial beta-adrenoceptor density. The changes probably are caused by reduction of cardiac NE release by ACE inhibition and may contribute to the beneficial effects of ACE inhibitor therapy in patients with chronic heart failure. PMID- 10516203 TI - Potassium channel-mediated vasorelaxation is impaired in experimental renal failure. AB - Chronic renal failure is associated with increased cardiovascular morbidity and abnormal arterial tone, but the underlying pathophysiological mechanisms are poorly understood. Therefore, we studied the responses of isolated mesenteric arterial rings from Wistar-Kyoto rats in standard organ chambers 6 wk after subtotal (5/6) nephrectomy or sham operation. Subtotal nephrectomy resulted in a 1.7-fold elevation of plasma urea nitrogen, whereas blood pressure was not significantly affected. Endothelium-mediated relaxations of norepinephrine precontracted rings to ACh were impaired in renal failure rats. The nitric oxide (NO) synthase inhibitor N(G)-nitro-L-arginine methyl ester inhibited relaxations to ACh more effectively in the renal failure group, whereas the cyclooxygenase inhibitor diclofenac did not significantly affect the response in either group. Inhibition of Ca(2+)-activated K(+) channels by charybdotoxin and apamin attenuated NO synthase- and cyclooxygenase-resistant relaxations to ACh in control but not renal failure rats and abolished the difference between these groups. Endothelium-independent relaxations to isoproterenol and cromakalim, vasodilators acting via beta-adrenoceptors and ATP-sensitive K(+) channels, respectively, were impaired in the renal failure group, whereas relaxations to the NO donor nitroprusside were similar in both groups. In conclusion, endothelium-mediated relaxation in renal failure rats was impaired in the absence and presence of NO synthase and cyclooxygenase inhibition but not with prevented smooth muscle hyperpolarization. Endothelium-independent relaxations to isoproterenol and cromakalim were also attenuated after 5/6 nephrectomy. These results suggest that impaired vasodilatation in experimental renal failure could be attributed to reduced relaxation via arterial K(+) channels. PMID- 10516202 TI - Further attenuation of endothelium-dependent relaxation imparted by natriuretic peptide receptor antagonism. AB - Nitric oxide (NO) is an important endothelium-derived relaxing factor that functions via activation of soluble guanylyl cyclase and cGMP generation in vascular smooth muscle. Recently, studies have described the synthesis and secretion of C-type natriuretic peptide (CNP) from endothelial cells. This peptide also mediates relaxation via cGMP but through activation of particulate guanylyl cyclase. We tested the hypothesis that endothelium-dependent relaxations to acetylcholine or bradykinin in isolated canine coronary arteries involve both releases of NO and CNP. Rings of canine coronary arteries were incubated with either inhibitors of NO production (N(G)-monomethyl-L-arginine, L-NMMA) or the natriuretic peptide receptor antagonist HS-142-1. CNP caused concentration dependent relaxations of rings with and without endothelium. These relaxations were attenuated by HS-142-1. Relaxations to acetylcholine and bradykinin were attenuated by L-NMMA alone but not attenuated by HS-142-1 alone. Coinhibition with L-NMMA and HS-142-1 significantly inhibited acetylcholine- and bradykinin induced relaxation to a magnitude greater than either inhibitor alone. In summary, a novel interaction between the NO and the natriuretic peptide system is demonstrated by increased attenuation of endothelium-dependent relaxations to acetylcholine and bradykinin when both NO synthase and natriuretic peptide receptors are inhibited. These investigations support the concept of release of multiple endothelium-derived factors in response to acetylcholine- and bradykinin receptor stimulation in endothelial cells, which may include CNP, as well as NO. PMID- 10516204 TI - A (13)C NMR double-labeling method to quantitate local myocardial O(2) consumption using frozen tissue samples. AB - Measurement of local myocardial O(2) consumption (VO(2)) has been problematic but is needed to investigate the heterogeneity of aerobic metabolism. The goal of the present investigation was to develop a method to measure local VO(2) using small frozen myocardial samples, suitable for determining VO(2) profiles. In 26 isolated rabbit hearts, 1.5 mmol/l [2-(13)C]acetate was infused for 4 min, followed by 1.5 min of [1,2-(13)C]acetate. The left ventricular (LV) free wall was then quickly frozen. High-resolution (13)C-NMR spectra were measured from extracts taken from 2- to 3-mm thick transmural layer samples. The multiplet intensities of glutamate were analyzed with a computer model allowing simultaneous estimation of the absolute flux through the tricarboxylic acid cycle and the fractional contribution of acetate to acetyl CoA formation from which local VO(2) was calculated. The (13)C-derived VO(2) in the LV free wall was linearly related to "gold standard" VO(2) from coronary venous O(2) electrode measurements in the same region (r = 0.932, n = 22, P < 0.0001, slope 1.05) for control and lowered metabolic rates. The ratio of subendocardial to subepicardial VO(2) was 1.52 +/- 0.19 (SE, significantly >1, P < 0.025). Local myocardial VO(2) can now be quantitated with this new (13)C method to determine profiles of aerobic energy metabolism. PMID- 10516205 TI - TNF-alpha enhances cardiac myocyte NO production through MAP kinase-mediated NF kappaB activation. AB - We have previously reported that interleukin-1beta (IL-1beta) alone induced nitric oxide (NO) production by neonatal rat cardiac myocytes (CM). The effects of tumor necrosis factor-alpha (TNF-alpha) on inducible NO synthase (iNOS) were not characterized. Unlike IL-1beta, TNF-alpha alone failed to enhance NO production in CM. However, the addition of TNF-alpha to IL-1beta significantly enhanced iNOS mRNA expression, iNOS protein synthesis, and NO production (NO( )(2)). TNF-alpha enhancement of IL-1beta-induced NO(-)(2) production was blocked by PD-98059, a selective mitogen-activated protein (MAP) kinase kinase inhibitor, but not calphostin C (Cal C), a protein kinase C inhibitor. TNF-alpha-enhanced MAP kinase activity was associated with an increase in IL-1beta-stimulated NF kappaB activity. PD-98059, but not Cal C, inhibited both TNF-alpha-enhanced MAP kinase and NF-kappaB activities. Thus TNF-alpha enhancement of IL-1beta-induced NO production is associated with MAP kinase-mediated activation of NF-kappaB. PMID- 10516206 TI - Evidence for peroxynitrite as a signaling molecule in flow-dependent activation of c-Jun NH(2)-terminal kinase. AB - The c-Jun NH(2)-terminal kinase (JNK), also known as stress-activated protein kinase, is a mitogen-activated protein kinase that determines cell survival in response to environmental stress. Activation of JNK involves redox-sensitive mechanisms and physiological stimuli such as shear stress, the dragging force generated by blood flow over the endothelium. Laminar shear stress has antiatherogenic properties and controls structure and function of endothelial cells by mechanisms including production of nitric oxide (NO) and superoxide (O( )(2)). Here we show that both NO and O(-)(2) are required for activation of JNK by shear stress in endothelial cells. The present study also demonstrates that exposure of endothelial cells to shear stress increases tyrosine nitration, a marker of reactive nitrogen species formation. Furthermore, inhibitors or scavengers of NO, O(-)(2), or reactive nitrogen species prevented shear-dependent increase in tyrosine nitration and activation of JNK. Peroxynitrite alone, added to cells as a bolus or generated over 60 min by 3-morpholinosydnonimine, also activates JNK. These results suggest that reactive nitrogen species, in this case most likely peroxynitrite, act as signaling molecules in the mechanoactivation of JNK. PMID- 10516207 TI - Mechanical force-induced signal transduction in lung cells. AB - The lung is a unique organ in that it is exposed to physical forces derived from breathing, blood flow, and surface tension throughout life. Over the past decade, significant progress has been made at the cellular and molecular levels regarding the mechanisms by which physical forces affect lung morphogenesis, function, and metabolism. With the use of newly developed devices, mechanical forces have been applied to a variety of lung cells including fetal lung cells, adult alveolar epithelial cells, fibroblasts, airway epithelial and smooth muscle cells, pulmonary endothelial and smooth muscle cells, and mesothelial cells. These studies have led to new insights into how cells sense mechanical stimulation, transmit signals intra- and intercellularly, and regulate gene expression at the transcriptional and posttranscriptional levels. These advances have significantly increased our understanding of the process of mechanotransduction in lung cells. Further investigation in this exciting research field will facilitate our understanding of pulmonary physiology and pathophysiology at the cellular and molecular levels. PMID- 10516208 TI - Asbestos-induced phosphorylation of epidermal growth factor receptor is linked to c-fos and apoptosis. AB - We examined the mechanisms of interaction of crocidolite asbestos fibers with the epidermal growth factor (EGF) receptor (EGFR) and the role of the EGFR extracellular signal-regulated kinase (ERK) signaling pathway in early-response protooncogene (c-fos/c-jun) expression and apoptosis induced by asbestos in rat pleural mesothelial (RPM) cells. Asbestos fibers, but not the nonfibrous analog riebeckite, abolished binding of EGF to the EGFR. This was not due to a direct interaction of fibers with ligand, inasmuch as binding studies using fibers and EGF in the absence of membranes showed that EGF did not adsorb to the surface of asbestos fibers. Exposure of RPM cells to asbestos caused a greater than twofold increase in steady-state message and protein levels of EGFR (P < 0.05). The tyrphostin AG-1478, which inhibits the tyrosine kinase activity of the EGFR, but not the tyrphostin A-10, which does not affect EGFR activity, significantly ameliorated asbestos-induced increases in mRNA levels of c-fos but not of c-jun. Pretreatment of RPM cells with AG-1478 significantly reduced apoptosis in cells exposed to asbestos. Our findings suggest that asbestos-induced binding to EGFR initiates signaling pathways responsible for increased expression of the protooncogene c-fos and the development of apoptosis. The ability to block asbestos-induced elevations in c-fos mRNA levels and apoptosis by small-molecule inhibitors of EGFR phosphorylation may have therapeutic implications in asbestos related diseases. PMID- 10516209 TI - CFTR involvement in chloride, bicarbonate, and liquid secretion by airway submucosal glands. AB - Previous studies demonstrated that ACh-induced liquid secretion by porcine bronchi is driven by active Cl(-) and HCO(-)(3) secretion. The present study was undertaken to determine whether this process was localized to submucosal glands and mediated by the cystic fibrosis transmembrane conductance regulator (CFTR). When excised, cannulated, and treated with ACh, porcine bronchi secreted 15.6 +/- 0.6 microliter. cm(-2). h(-1). Removal of the surface epithelium did not significantly affect the rate of secretion, indicating that the source of the liquid was the submucosal glands. Pretreatment with diphenylamine-2-carboxylate, a relatively nonselective Cl(-)-channel blocker, significantly reduced liquid secretion by 86%, whereas pretreatment with DIDS, which inhibits a variety of Cl( ) channels but not CFTR, had no effect. When bronchi were pretreated with glibenclamide or 5-nitro-2-(3-phenylpropylamino)benzoic acid (both inhibitors of CFTR), the rate of ACh-induced liquid secretion was significantly reduced by 39 and 91%, respectively, compared with controls. Agents that blocked liquid secretion also caused disproportionate reductions in HCO(-)(3) secretion. Polyclonal antibodies to the CFTR bound preferentially to submucosal gland ducts and the surface epithelium, suggesting that this channel was localized to these sites. These data suggest that ACh-induced gland liquid secretion by porcine bronchi is driven by active secretion of both Cl(-) and HCO(-)(3) and is mediated by the CFTR. PMID- 10516210 TI - PBA increases CFTR expression but at high doses inhibits Cl(-) secretion in Calu 3 airway epithelial cells. AB - Sodium 4-phenylbutyrate (PBA), a short-chain fatty acid, has been approved to treat patients with urea cycle enzyme deficiencies and is being evaluated in the management of sickle cell disease, thalassemia, cancer, and cystic fibrosis (CF). Because relatively little is known about the effects of PBA on the expression and function of the wild-type CF transmembrane conductance regulator (wt CFTR), the goal of this study was to examine the effects of PBA and related compounds on wt CFTR-mediated Cl(-) secretion. To this end, we studied Calu-3 cells, a human airway cell line that expresses endogenous wt CFTR and has a serous cell phenotype. We report that chronic treatment of Calu-3 cells with a high concentration (5 mM) of PBA, sodium butyrate, or sodium valproate but not of sodium acetate reduced basal and 8-(4-chlorophenylthio)-cAMP-stimulated Cl(-) secretion. Paradoxically, PBA enhanced CFTR protein expression 6- to 10-fold and increased the intensity of CFTR staining in the apical plasma membrane. PBA also increased protein expression of Na(+)-K(+)-ATPase. PBA reduced CFTR Cl(-) currents across the apical membrane but had no effect on Na(+)-K(+)-ATPase activity in the basolateral membrane. Thus a high concentration of PBA (5 mM) reduces Cl(-) secretion by inhibiting CFTR Cl(-) currents across the apical membrane. In contrast, lower therapeutic concentrations of PBA (0.05-2 mM) had no effect on cAMP-stimulated Cl(-) secretion across Calu-3 cells. We conclude that PBA concentrations in the therapeutic range are unlikely to have a negative effect on Cl(-) secretion. However, concentrations >5 mM might reduce transepithelial Cl(-) secretion by serous cells in submucosal glands in individuals expressing wt CFTR. PMID- 10516211 TI - Abnormal lung growth and the development of pulmonary hypertension in the Fawn Hooded rat. AB - The Fawn-Hooded rat (FHR) strain develops accelerated and severe pulmonary hypertension when exposed to slight decreases in alveolar PO(2). We recently observed that adult FHR lungs showed a striking pattern of disrupted alveolarization and hypothesized that abnormalities in lung growth in the perinatal period predisposes the FHR to the subsequent development of pulmonary hypertension. We found a reduction in lung weight in the fetus and 1-day- and 1 wk-old FHR compared with a normal rat strain (Sprague-Dawley). Alveolarization was reduced in infant and adult FHR lungs. In situ hybridization showed similar patterns of expression of two epithelial markers, surfactant protein C and 10-kDa Clara cell secretory protein, suggesting that the FHR lung is not characterized by global delays in epithelial maturation. Barium-gelatin angiograms demonstrated reduced background arterial filling and density in adult FHR lungs. Perinatal treatment of FHR with supplemental oxygen increased alveolarization and reduced the subsequent development of right ventricular hypertrophy in adult FHR. We conclude that the FHR strain is characterized by lung hypoplasia with reduced alveolarization and increased risk for developing pulmonary hypertension. We speculate that altered oxygen sensing may cause impaired lung alveolar and vascular growth in the FHR. PMID- 10516212 TI - Modeling the interactions of particulates with epithelial lining fluid antioxidants. AB - Oxidative stress may be a fundamental mode of injury associated with inspired particles. To examine this, we determined the ability of three carbon black particles (CBPs; M120, M880, and R250) and two forms of silicon dioxide, amorphous (Cabosil) and crystalline (DQ12) quartz, to deplete epithelium lining fluid antioxidant defenses. Single and composite antioxidant solutions of uric acid, ascorbic acid (AA), and reduced glutathione (GSH) were examined in the presence of particle concentrations of 150 microgram/ml. Uric acid was not depleted by any particle considered. AA was depleted in a near-linear fashion with time by the three different CBPs; however, AA depletion rates varied markedly with CBP type and decreased in the presence of metal chelators. An initially high GSH depletion rate was noted with all CBPs, and this was always accompanied by the appearance of oxidized glutathione. Exposure to Cabosil or DQ12 did not result in the loss of GSH. Together, these data demonstrate that particle type, size, and surface area are all important factors when considering particle-antioxidant interactions in the airways. PMID- 10516213 TI - cAMP stimulates Na(+) transport in rat fetal pneumocyte: involvement of a PTK- but not a PKA-dependent pathway. AB - To study a cAMP-mediated signaling pathway in the regulation of amiloride sensitive Na(+) transport in rat fetal distal lung epithelial cells, we measured an amiloride-sensitive short-circuit current (Na(+) transport). Forskolin, which increases the cytosolic cAMP concentration, stimulated the Na(+) transport. Forskolin also activated cAMP-dependent protein kinase (PKA). A beta-adrenergic agonist and cAMP mimicked the forskolin action. PKA inhibitors KT-5720, H-8, and myristoylated PKA-inhibitory peptide amide-(14-22) did not influence the forskolin action. These results suggest that forskolin stimulates Na(+) transport through a PKA-independent pathway. Furthermore, forskolin increased tyrosine phosphorylation of approximately 70- to 80-, approximately 97-, and approximately 110- to 120-kDa proteins. Protein tyrosine kinase (PTK) inhibitors (tyrphostin A23 and genistein) abolished the forskolin action. Moreover, 5-nitro-2-(3 phenylpropylamino)benzoate (a Cl(-)-channel blocker) prevented the stimulatory action of forskolin on Na(+) transport via abolishment of the forskolin-induced cell shrinkage and tyrosine phosphorylation. Based on these results, we conclude that forskolin (and cAMP) stimulates Na(+) transport in a PTK-dependent but not a PKA-dependent pathway by causing cell shrinkage, which activates PTK in rat fetal distal lung epithelial cells. PMID- 10516214 TI - Keratinocyte growth factor stimulates bronchial epithelial cell proliferation in vitro and in vivo. AB - Airway epithelial cell (AEC) proliferation is crucial to the maintenance of an intact airway surface and the preservation of host defenses. The factors that regulate AEC proliferation are not known. Keratinocyte growth factor (KGF), also known as FGF-7, is a member of the fibroblast growth factor family and a known epithelial cell mitogen. We studied the influence of KGF on the growth of cultured human bronchial epithelial cells and on bronchial cells of rats treated with KGF in vivo. First, we demonstrated the mRNA for the KGF receptor (KGFR) in both normal human bronchial epithelial (NHBE) cells and BEAS-2B cells (a human bronchial epithelial cell line). KGF caused a dose-dependent increase in DNA synthesis, as assessed by thymidine incorporation, in both cell types, with a maximal twofold increase in NHBE cells after 50 ng/ml KGF (P < 0.001). KGF also induced a doubling in NHBE cell number at 10 ng/ml (P < 0.001). Finally, we determined the effect of intratracheal administration of KGF to rats on proliferation of AEC in vivo. Measuring bromodeoxyuridine (BrdU) incorporation in AEC nuclei, KGF increased BrdU labeling of rat AEC in both large and small airways by approximately threefold compared with PBS-treated controls (P < 0.001). Thus KGF induces proliferation of bronchial epithelial cells both in vitro and in vivo. PMID- 10516215 TI - Critical role of GSH in silica-induced oxidative stress, cytotoxicity, and genotoxicity in alveolar macrophages. AB - The main objective of this study was to evaluate the critical role of glutathione (GSH) in silica-induced oxidative stress, cytotoxicity, and genotoxicity in rat alveolar macrophages (AMs). Silica-induced superoxide radical and hydrogen peroxide formation were determined with lucigenin-dependent chemiluminescence and 2', 7'-dichlorofluorescin diacetate fluorescence test, respectively. The cytotoxicity of silica was estimated by lactate dehydrogenase leakage, and a comet assay was used for examining silica-induced DNA damage in AMs. The intracellular GSH content was modulated by N-acetylcysteine, a GSH precursor, and buthionine sulfoximine, a specific GSH synthesis inhibitor. It was found that silica led to a dose- and time-dependent decrease in GSH content in AMs. N acetylcysteine increased intracellular GSH level and protected against silica induced reactive oxygen species formation, lactate dehydrogenase leakage, and DNA strand breaks in AMs. In contrast, buthionine sulfoximine pretreatment depleted cellular GSH and enhanced the susceptibility of AMs to the cytotoxic and genotoxic effects of silica. It thus appears that GSH plays a critical role in protecting against silica-induced cell injury, most probably through its antioxidant activity. PMID- 10516216 TI - Lung matrix incorporation of plasma fibronectin reduces vascular permeability in postsurgical bacteremia. AB - Plasma fibronectin (pFN) can incorporate into the lung extracellular matrix (ECM) as well as enhance hepatic cell phagocytic removal of bloodborne microparticulate debris that can contribute to lung vascular injury. Treatment of human pFN (hFN) with N-ethylmaleimide (NEM) blocks its ECM incorporation but not its ability to augment phagocytosis. Using hFN purified from fresh human plasma cryoprecipitate, we compared the effect of NEM-treated hFN versus normal hFN on lung transvascular protein clearance (TVPC) in postoperative bacteremic sheep to determine whether the ability of hFN to attenuate the increase in lung endothelial permeability required its ECM incorporation. Sheep with lung lymph fistulas were infused with a sublethal dose of Pseudomonas aeruginosa (5 x 10(8)) 48 h after surgery. In the first study, sheep received either FN-rich human cryoprecipitate, FN-deficient cryoprecipitate, FN purified from cryoprecipitate (hFN), FN-deficient cryoprecipitate reconstituted with purified hFN, or the sterile saline diluent. In the second study, sheep received either 200 mg of purified hFN (group I), 200 mg of NEM-treated hFN (group II), or the saline diluent (group III). In the first study, the increase in TVPC after bacterial challenge was attenuated by FN-rich cryoprecipitate, hFN, or reconstituted FN-deficient cryoprecipitate (P < 0.05) but not by saline and FN-deficient cryoprecipitate. In the second study, TVPC increased by 2 h (P < 0.05) and peaked over 4-8 h (P < 0.05) at 380-420% above baseline in postoperative bacteremic sheep given the diluent (group III). In contrast, intravenous infusion of hFN, but not of NEM-treated hFN, significantly (P < 0.05) attenuated this increase of lung protein clearance. Thus the ability for the intravenously infused purified pFN to attenuate the increase in lung endothelial protein permeability in sheep during postsurgical bacteremia appears to require its ECM incorporation into the interstitial ECM of the lung. PMID- 10516217 TI - alpha-adrenergic blockade restores normal fluid transport capacity of alveolar epithelium after hemorrhagic shock. AB - Activation of beta-adrenergic receptors in the lung is an important mechanism that can prevent alveolar flooding after brief but severe hemorrhagic shock. However, a neutrophil-dependent oxidant injury to the alveolar epithelium prevents the normal upregulation of alveolar fluid clearance by catecholamines after prolonged hemorrhagic shock. Because hemorrhage increases proinflammatory cytokine expression in the lung partly through the activation of alpha-adrenergic receptors, the objective of this study was to determine whether alpha-adrenergic blockade would restore the normal fluid transport capacity of the alveolar epithelium after hemorrhagic shock. Hemorrhagic shock was associated with a significant increase of interleukin-1beta (IL-1beta) concentration in the lung and a failure of the alveolar epithelium to respond to beta-adrenergic agonists, with the upregulation of vectorial fluid transport despite intra-alveolar administration of exogenous catecholamines. In contrast, catecholamine-mediated upregulation of alveolar liquid clearance was restored by pretreatment with phentolamine, an alpha-adrenergic-receptor antagonist. Phentolamine pretreatment also significantly attenuated the shock-mediated increase of IL-1beta concentration in the lung. Additional experiments demonstrated that the inhibition of IL-1beta binding to its receptor by the administration of IL-1 receptor antagonist restored the normal fluid transport capacity of the alveolar epithelium after hemorrhagic shock. In summary, the results of these studies indicate that the activation of alpha-adrenergic receptors after hemorrhagic shock prevents the beta-adrenergic-dependent upregulation of alveolar fluid clearance by modulating the severity of the pulmonary inflammatory response. PMID- 10516218 TI - Inhibition of PARS attenuates endotoxin-induced dysfunction of pulmonary vasorelaxation. AB - Endotoxin (Etx) causes excessive activation of the nuclear repair enzyme poly(ADP ribose) synthase (PARS), which depletes cellular energy stores and leads to vascular dysfunction. We hypothesized that PARS inhibition would attenuate injury to mechanisms of pulmonary vasorelaxation in acute lung injury. The purpose of this study was to determine the effect of in vivo PARS inhibition on Etx-induced dysfunction of pulmonary vasorelaxation. Rats received intraperitoneal saline or Etx (Salmonella typhimurium; 20 mg/kg) and one of the PARS inhibitors, 3 aminobenzamide (3-AB; 10 mg/kg) or nicotinamide (Nic; 200 mg/kg), 90 min later. After 6 h, concentration-response curves were determined in isolated pulmonary arterial rings. Etx impaired endothelium-dependent (response to ACh and calcium ionophore) and -independent (sodium nitroprusside) cGMP-mediated vasorelaxation. 3-AB and Nic attenuated Etx-induced impairment of endothelium-dependent and independent pulmonary vasorelaxation. 3-AB and Nic had no effect on Etx-induced increases in lung myeloperoxidase activity and edema. Lung ATP decreased after Etx but was maintained by 3-AB and Nic. Pulmonary arterial PARS activity increased fivefold after Etx, which 3-AB and Nic prevented. The beneficial effects were not observed with benzoic acid, a structural analog of 3-AB that does not inhibit PARS. Our results suggest that PARS inhibition with 3-AB or Nic improves pulmonary vasorelaxation and preserves lung ATP levels in acute lung injury. PMID- 10516219 TI - Surfactant protein A enhances alveolar macrophage phagocytosis of a live, mucoid strain of P. aeruginosa. AB - In this study, we investigate the interaction between surfactant protein A (SP-A) and a live, mucoid strain of Pseudomonas aeruginosa and identify a mechanism of clearance of this organism by alveolar macrophages. (125)I-labeled SP-A bound live, but not heat-killed, P. aeruginosa organisms in a concentration-dependent manner. Unlabeled SP-A bound live bacteria, protein isolated from whole organisms, and specific proteins of the P. aeruginosa outer membrane. The binding of SP-A to P. aeruginosa and outer membrane components was inhibited by either EDTA or mannose. Phagocytosis assays with fluorescent microscopy demonstrated that the percentage of macrophages with internalized FITC-labeled P. aeruginosa was increased 1.8-fold (19 vs. 35%) by pretreating the live bacteria with SP-A. This finding was confirmed by direct visualization of ingested bacteria by electron microscopy. Adhering macrophages to SP-A-coated surfaces attenuated the increased uptake of P. aeruginosa pretreated with SP-A, suggesting that SP-A acts as an opsonin to stimulate macrophage phagocytosis of this strain of P. aeruginosa. PMID- 10516220 TI - Nitric oxide-induced reduction of lung cell and whole lung thioredoxin expression is regulated by NF-kappaB. AB - We examined whether nitric oxide (NO)-induced inhibition of thioredoxin (Thx) expression is regulated by a mechanism mediated by a transcription factor, i.e., nuclear factor-kappaB (NF-kappaB), in cultured porcine pulmonary artery endothelial cells (PAEC) and in mouse lungs. Western blot analysis revealed that IkappaB-alpha content was reduced by 20 and 60% in PAEC exposed to 8.5 ppm NO for 2 and 24 h, respectively. NO exposure also caused significant reductions of cytosol fraction p65 and p52 content in PAEC. The nuclear fraction p65 and p52 contents were significantly reduced only in PAEC exposed to NO for 24 h. Exposure to NO resulted in a 50% reduction of p52 mRNA but not of the IkappaB-alpha subunit. DNA binding activity of the oligonucleotide encoding the NF-kappaB sequence in the Thx gene was significantly reduced in PAEC exposed to NO for 24 h. Exposure of mice to 10 ppm NO for 24 h resulted in a significant reduction of lung Thx and IkappaB-alpha mRNA and protein expression and in the oligonucleotide encoding Thx and NF-kappaB/DNA binding. These results 1) demonstrate that the effects of NO exposure on Thx expression in PAEC are comparable to those observed in intact lung and 2) suggest that reduced expression of the NF-kappaB subunit, leading to reduced NF-kappaB/DNA binding, is associated with the loss of Thx expression in PAEC and in intact mouse lungs. PMID- 10516221 TI - Increased expression of calreticulin is linked to ANG IV-mediated activation of lung endothelial NOS. AB - This study demonstrates that ANG IV-induced activation of lung endothelial cell nitric oxide synthase (ecNOS) is mediated through mobilization of Ca(2+) concentration and by increased expression and release of the Ca(2+) binding protein calreticulin in pulmonary artery endothelial cells (PAEC). In Ca(2+)-free medium and in the presence of the ANG II AT(1) and AT(2) receptor antagonists losartan and PD-123319 (1 microM each), respectively, ANG IV (5, 50, and 500 nM) significantly increased intracellular Ca(2+) release in PAEC (P < 0.05 for all concentrations). In contrast, ANG IV-mediated activation of ecNOS was abolished by the intracellular Ca(2+) chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N' tetraacetic acid-AM. ANG IV stimulation resulted in significantly increased expression of calreticulin in cells as well as release of calreticulin into the medium of cells as early as 2 h after ANG IV stimulation (P < 0.05). Catalytic activity of purified ecNOS in the absence of calmodulin was increased in a concentration-dependent fashion by calreticulin. Immunocoprecipitation studies revealed that ecNOS and calreticulin were coprecipitated in ANG IV-stimulated PAEC. These results demonstrate that ANG IV-mediated activation of ecNOS is regulated by intracellular Ca(2+) mobilization and by increased expression of calreticulin, which appears to involve interaction of ecNOS and calreticulin proteins in PAEC. PMID- 10516222 TI - Augmentation of eosinophil degranulation and LTC(4) secretion by integrin mediated endothelial cell adhesion. AB - We examined the effect of eosinophil ligation to cultured human umbilical vein endothelial cells (HUVECs) in augmenting the stimulated secretion of leukotriene (LT) C(4) and eosinophil peroxidase (EPO). The effects of adhesion were compared before and after specific blockade with monoclonal antibodies directed against eosinophil surface integrins or endothelial counterligands. Adhesion to HUVECs augmented EPO release caused by formyl-methionyl-leucyl-phenylalanine plus cytochalasin B from 403 +/- 15.3 (BSA control) to 778 +/- 225 ng/10(6) cells for eosinophils exposed to interleukin-1alpha-treated HUVECs (P < 0.05) and also caused a twofold increase in stimulated LTC(4) secretion (P < 0.05). To determine whether augmented secretion resulted directly from adhesive ligation, studies were also performed with paraformaldehyde-treated HUVECs; stimulated secretion of LTC(4) from eosinophils was comparable to that for living HUVECs. Our study is the first demonstration that adhesion to HUVECs through ligation to alpha(4)- or beta(2)-integrin on the eosinophil surface causes augmentation of stimulated secretion of both EPO and LTC(4) and that blockade of adhesion molecules on either eosinophils or HUVECs prevents the priming effect on eosinophil secretion. PMID- 10516223 TI - Inhibition of mucin release from airway goblet cells by polycationic peptides. AB - In the present study, we investigated whether polycationic peptides affect mucin release from cultured airway goblet cells. Confluent primary hamster tracheal surface epithelial cells were metabolically radiolabeled with [(3)H]glucosamine for 24 h and chased for 30 min in the presence of varying concentrations of either poly-L-arginine (PLA) or poly-L-lysine (PLL) to assess the effects on [(3)H]mucin release. Possible cytotoxicity by the polycations was assessed by measuring lactate dehydrogenase release, (51)Cr release, and cell exfoliation. The results were as follows: 1) both PLA and PLL inhibited mucin release in a dose-dependent fashion; 2) there was no significant difference in either lactate dehydrogenase release, (51)Cr release, or the number of floating cells between control and treatment groups; 3) the effects of both PLA and PLL on mucin release were completely blocked by neutralizing the positive charges either by pretreatment with heparin or by N-acetylation of the polycations; and 4) both PLA and PLL completely masked the stimulatory effect of ATP on mucin release. We conclude that these polycationic peptides can inhibit mucin release from airway goblet cells without any apparent cytotoxicity, and the inhibitory effect seems to be attributable to their positive charges. These are the first nonsteroidal agents, to the best of our knowledge, that have been shown to inhibit mucin release from airway goblet cells. PMID- 10516224 TI - Hydrogen peroxide decreases Ca(2+) sensitivity in airway smooth muscle by inhibiting rMLC phosphorylation. AB - The purpose of this study was to determine the mechanism by which hydrogen peroxide (H(2)O(2)), an important inflammatory mediator, relaxes canine tracheal smooth muscle (CTSM). H(2)O(2) caused concentration-dependent relaxations of CTSM strips contracted with ACh or isotonic KCl [EC(50) of 0.24 +/- 0.04 (SE) and 0.23 +/- 0.04 mM, respectively]. Indomethacin (10 microM) decreased the sensitivity of both KCl- and ACh-contracted strips to H(2)O(2). H(2)O(2) increased intracellular cAMP levels, an increase that was abolished by indomethacin. H(2)O(2) did not affect intracellular cGMP levels. In strips treated with indomethacin and contracted with ACh or isotonic KCl, H(2)O(2)-evoked relaxations were accompanied by increases in intracellular Ca(2+) concentration and decreases in regulatory myosin light chain phosphorylation. We conclude that H(2)O(2) decreases Ca(2+) sensitivity in CTSM by decreasing regulatory myosin light chain phosphorylation through inhibition of myosin light chain kinase and/or activation of smooth muscle protein phosphatases. PMID- 10516225 TI - Differential NF-kappaB activation after intratracheal endotoxin. AB - We examined the relationship between nuclear factor (NF)-kappaB DNA binding activity, cytokine gene expression, and neutrophilic alveolitis in rats after intratracheal (IT) instillation of endotoxin [lipopolysaccharide (LPS)]. NF kappaB activation in lung tissue mirrored neutrophilic alveolitis after IT LPS instillation, with NF-kappaB activation and neutrophilic influx beginning 2 h after IT LPS doses of 0.01 mg/kg or greater. In lung lavage fluid cells, however, transient NF-kappaB activation was present in alveolar macrophages by 15 min after IT LPS instillation, followed by a second peak of NF-kappaB activation corresponding to the onset on neutrophilic alveolitis. For cytokines thought to be NF-kappaB dependent, two different patterns of mRNA expression were found. Interleukin (IL)-1alpha, IL-1beta, and tumor necrosis factor-alpha showed increased mRNA by 30 min after IT LPS instillation, but IL-6- and cytokine induced neutrophil chemoattractant mRNAs were not substantially increased until 2 h after IT LPS instillation. Therefore, IT LPS causes differential NF-kappaB activation in air space cells and lung tissue, which likely determines production of key cytokines and directs the evolution of neutrophilic alveolitis. PMID- 10516226 TI - Respiratory syncytial virus upregulates expression of the substance P receptor in rat lungs. AB - Respiratory syncytial virus (RSV) is a major respiratory pathogen in infants. The first goal of this study was to determine whether the infection following endotracheal inoculation of RSV in Fischer 344 rats results in increased inflammatory responses to substance P (SP) either released by capsaicin from sensory nerves or injected into the circulation. Five days after inoculation, the extravasation of Evans blue-labeled albumin after capsaicin or SP was significantly greater in RSV-infected airways than in pathogen-free controls. The peptide-degrading activity of the regulatory enzyme neutral endopeptidase was unaffected by RSV. However, SP(NK(1)) receptor mRNA levels increased fivefold in RSV-infected lungs, and the density of SP binding sites in the bronchial mucosa increased threefold. These data suggest that RSV makes the airways abnormally susceptible to the proinflammatory effects of SP by upregulating SP(NK(1)) receptor gene expression, thereby increasing the density of these receptors on target cells. This effect may contribute to the inflammatory reaction to the virus and could be a target for the therapy of RSV disease and its sequelae. PMID- 10516227 TI - Soluble guanylate cyclase gene expression and localization in rat lung after exposure to hypoxia. AB - The nitric oxide (NO)-cGMP signal transduction pathway plays an important role in the regulation of pulmonary vascular tone and resistance in pulmonary hypertension. A number of studies have demonstrated that endothelial (e) and inducible nitric oxide synthases (NOS) are upregulated in hypoxia-exposed rat lung. These changes in NOS expression have been found to correlate with the process of pulmonary vascular remodeling in hypoxia-induced pulmonary hypertension, and remodeling is increased in the absence of eNOS. In this study, we examined the expression and localization of soluble guanylate cyclase (sGC), the primary receptor for NO, in hypoxia- and normoxia-treated rat lungs. Male Sprague-Dawley rats were exposed to hypoxia (10% O(2), normobaric) or normoxia for 1, 3, 5, and 21 days. The lungs were used for Western analysis of sGC protein, sGC enzyme activity, immunohistochemistry using antiserum against sGC alpha(1)- and beta(1)-subunits, and nonradioactive in situ hybridization (NRISH) using a digoxigenin-labeled sGC alpha(1)-subunit cRNA probe. Western blot analysis revealed a more than twofold increase of sGC protein alpha(1)-subunit in rat lungs exposed to 3, 5, and 21 days of hypoxia, correlating well with sGC enzyme activity. Immunohistochemistry and NRISH demonstrated increased expression of sGC in the smooth muscle cells of the pulmonary arteries and arterioles in the hypoxic rat lungs when compared with normoxic controls. Based on our results, the upregulation of sGC may play an important role in the regulation of smooth muscle tone and pressure in the pulmonary circulation during chronic hypoxia. PMID- 10516228 TI - Developmental shift in the relative percentages of lung fibroblast subsets: role of apoptosis postseptation. AB - We have used the lipophilic, fluorescent dye Nile red and flow cytometry to identify and isolate two rat lung fibroblast subsets, lipid-containing interstitial cells (LICs) and non-LICs (NLICs) and to quantitate developmental changes in the relative percentages of these subsets. A significant decrease was observed in the percentage of LICs (from 79.0 +/- 3.8% on postnatal day 4 to 28.6 +/- 4.2% on day 30; P < 0.0001). To determine whether one or both subsets undergo apoptosis postseptation, fibroblasts from 16- to 18-day rats were treated with BODIPY-conjugated dUTP to label DNA strand breaks, which were then quantitated by flow cytometry. Apoptotic cells were judged to be predominantly LICs based on flow cytometric estimates of cell size and granularity and on light-microscopic colocalization of intracellular lipid and Hoechst-positive apoptotic bodies. Cell proliferation was compared in LICs and NLICs with both an in vitro [(3)H]thymidine incorporation assay and cell cycle analysis of propidium iodide stained cells. Results of both assays indicated that on days 4-5, LICs proliferated more rapidly than NLICs. Tropoelastin and fibronectin mRNA expression, evaluated by RT-PCR, indicated that although tropoelastin mRNA levels did not differ, fibronectin mRNA levels were approximately ninefold greater in LICs. These results demonstrate the feasibility of a flow cytometric assay for the analysis of size, granularity, and intracellular lipid content of neonatal rat lung fibroblast subsets. Subsets differed substantially with respect to proliferative capacity, fibronectin mRNA expression, and incidence of apoptosis postseptation. Together with the observed changes in relative percentages of fibroblast subsets with age, these data suggest that the ratio of LICs to NLICs could be a critical determinant of fibroblast function during lung development. PMID- 10516229 TI - Effects of maturation on adrenergic neurotransmission in ovine cerebral arteries. AB - The present studies examine the hypothesis that multiple adrenergic neuroeffector mechanisms are not fully developed in fetal, compared with adult, ovine middle cerebral arteries. In arteries denuded of endothelium and pretreated with 1 microM atropine to block involvement of muscarinic receptors, 10 microM capsaicin to deplete sensory peptidergic neurons, and 10 microM nitro-L-arginine methyl ester (L-NAME) to block possible influences from nitric oxidergic innervation, transmural stimulation at 16 Hz increased contractile tensions to 9.5 +/- 3.7% (n = 6) of the potassium maximum in adult arteries. Corresponding values in fetal arteries, however, were significantly less and averaged only 1.1 +/- 0.6% (n =10). However, postsynaptic sensitivity to norepinephrine (NE) was similar in the two age groups; NE pD(2) values (-log EC(50)) averaged 6.11 +/- 0.12 (n = 6) and 6.33 +/- 0.09 M (n = 9) in fetal and adult arteries, respectively. Similarly, NE content measured via HPLC was also similar in the two age groups and averaged 32.4 +/- 5.0 (n = 17) and 32.5 +/- 3.9 ng/ng wet wt (n = 13) in fetal and adult middle cerebral arteries, respectively. In contrast, stimulation-induced NE release was greater in fetal than in adult arteries, whether calculated as total mass released [883 +/- 184 (n = 17) vs. 416 +/- 106 pg NE/mg wet wt (n = 13)] or as fractional release [51.1 +/- 5.3 (n = 17) vs. 22.8 +/- 3.8 pg/pg NE content per pulse x 10(-6)]. Measured as an index of synaptic density, neuronal cocaine sensitive NE uptake was similar in fetal and adult arteries [1.55 +/- 0.40 (n = 10) and 1.84 +/- 0.51 pmol/mg wet wt (n = 7), respectively]. Overall, age-related differences in postsynaptic sensitivity to NE, NE release, and NE uptake capacity cannot explain the corresponding age-related differences in response to stimulation. The data thus suggest that total synaptic volume and cleft width, in particular, are probably greater and/or that adrenergic corelease of vasoactive substances other than NE is altered in fetal compared with adult middle cerebral arteries. PMID- 10516230 TI - Pressure and volume overloads are associated with ventricular hypertrophy in male rainbow trout. AB - We investigated whether ventricular hypertrophy in reproductively mature male trout (Oncorhynchus mykiss) is associated with elevated hemodynamic loads. We measured ventral aortic blood pressure, pulse pressure dynamics, and blood volume in cannulated, unanesthetized trout with a wide range of relative ventricle masses (RVM, 0.076-0.199% of body wt). We also investigated in vitro pressure volume dynamics in the bulbus arteriosus taken from trout with a wide range of RVMs. RVM was positively correlated with peak systolic pressure (SBP), mean blood pressure, and pulse pressure. Diastolic pressure and the absolute duration of arterial systole were similar among all animals, but a lower heart rate and a smaller relative duration of arterial systole were correlated with increasing RVM. Blood volume was expanded up to 34% as ventricles enlarged, and clearance of Evans blue dye was greater at higher SBP. Mass, maximal volume, and the pressure volume dynamics of the bulbus were similar among all animals, suggesting that the bulbus did not compensate for ventricular enlargement. This conclusion was supported by the elevated maximal rates of arterial pressure development (+dP/dt) and decay (-dP/dt) observed as RVM increased. We conclude that 1) mature trout are hypertensive and hypervolemic, 2) the dynamics of the bulbus may contribute to increased afterload, and 3) these changes in hemodynamic load may promote ventricular hypertrophy. PMID- 10516231 TI - Effect of glucose supply on ovine uteroplacental glucose metabolism. AB - To test the hypothesis that glucose supply to the uteroplacenta (UP) regulates UP glucose metabolism into oxidative and nonoxidative pathways, we studied eight late-gestation pregnant sheep at low (LG) and high (HG) maternal glucose concentrations (G(M)), using Fick principle and tracer glucose methodology. UP glucose consumption (UPGC) correlated directly with G(M) (r = 0.75, P = 0.0006), and UP glucose decarboxylation (r = 0.80, P = 0.0001), and lactate production (r = 0.90, P = 0.0001) rates were directly correlated with UPGC. The combined fractional production rate for lactate, fructose, and CO(2) from UPGC was the same in LG and HG periods. The fraction of UP oxygen consumption used for glucose oxidation increased by about 50% from LG to HG conditions; however, there was no change in UP oxygen consumption. Nearly half of UPGC was not accounted for by lactate, fructose, and CO(2) production, and about two-thirds of UP oxygen consumption was not accounted for by immediate oxidation of glucose carbon just taken up by the UP. These results indicate that glucose supply directly regulates UP glucose oxidative metabolism and that there is a reciprocal relationship between UP glucose oxidation and the oxidation of other substrates. PMID- 10516232 TI - Leptin inhibits insulin secretion induced by cellular cAMP in a pancreatic B cell line (INS-1 cells). AB - The effect of leptin on insulin secretion is controversial due to conflicting results in the literature. In the present study, we incubated insulin-producing rat insulinoma INS-1 cells for 60 min and examined the effects of recombinant murine leptin (20 nmol/l). We found that leptin (0.1-100 nmol/l) did not affect the insulin response to glucose (1-20 mmol/l). However, when cells were incubated with agents that increase the intracellular content of cAMP, i.e., glucagon-like peptide-1 (100 nmol/l), pituitary adenylate cyclase activating polypeptide (100 nmol/l), forskolin (2.5 micromol/l), dibutyryl-cAMP (1 mmol/l), or 3-isobutyl-1 methylxanthine (100 micromol/l), leptin significantly reduced insulin secretion (by 34-58%, P < 0.05-0.001). In contrast, when insulin secretion was stimulated by the cholinergic agonist carbachol (100 micromol/l) or the phorbol ester 12-O tetradecanoylphorbol 13-acetate (1 micromol/l), both of which activate protein kinase C, leptin was without effect. We conclude that leptin inhibits insulin secretion from INS-1 cells under conditions in which intracellular cAMP is increased. This suggests that the cAMP-protein kinase A signal transduction pathway is a target for leptin to inhibit insulin secretion in insulin-producing cells. PMID- 10516233 TI - Heat stress induces ultrastructural changes in cutaneous capillary wall of heat acclimated rock pigeon. AB - In heat-acclimated rock pigeons, cutaneous water evaporation is the major cooling mechanism when exposed at rest to an extremely hot environment of 50-60 degrees C. This evaporative pathway is also activated in room temperature by a beta adrenergic antagonist (propranolol) or an alpha-adrenergic agonist (clonidine) and inhibited by a beta-adrenergic agonist (isoproterenol). In contrast, neither heat exposure nor drug administration activates cutaneous evaporation in cold acclimated pigeons. To elucidate the mechanisms underlying this phenomenon, we studied the role of the ultrastructure and permeability of the cutaneous vasculature. During both heat stress and the administration of propranolol and clonidine, we observed increased capillary fenestration and endothelial gaps. Similarly, propranolol increased the extravasation of Evans blue-labeled albumin in the skin tissue. We concluded that heat acclimation reinforces a mechanism by which the activation of adrenergic signal transduction pathways alters microvessel permeability during heat stress. Consequently the flux of plasma proteins and water into the interstitial space is accelerated, providing an interstitial source of water for sustained cutaneous evaporative cooling. PMID- 10516234 TI - Sex differences in body weight gains following amygdaloid lesions in rats. AB - Lesions of the most posterodorsal aspects of the amygdala resulted in equal weight gains (mean = 58 g) in male and female rats during a 22-day observation period. However, the absolute weight gains in the first 5 days after lesions were greater in females (+41.4 g) than in males (+18.8 g), as were the longer-term gains relative to their respective control groups. In a second study with female rats, it was found that amygdaloid lesions had little effect on the estrous cycle and that ovariectomy resulted in additional excessive weight gains in both rats with sham lesions and those with amygdaloid lesions. The weight gains produced by amygdaloid lesions and ovariectomy were additive. It is concluded that there is a sex difference in weight gains after amygdaloid lesions, but that the lesion induced obesity is independent of estrogen levels. Similarities to lesions of the ventromedial hypothalamus are noted, and an amygdaloid-ventromedial hypothalamic pathway for the regulation of feeding behavior is proposed. PMID- 10516235 TI - Nitric oxide modulates spontaneous swallowing behavior in near-term ovine fetus. AB - Human and ovine fetuses demonstrate an enhanced rate of swallowing, an activity critical for amniotic fluid regulation. Fetal swallowing may be modulated by both systemic and central factors. Nitric oxide (NO) is a central neuromodulator that has been localized to brain regions regulating thirst and swallowing. We sought to determine if NO contributes to the regulation of spontaneous ovine fetal swallowing. Six time-dated pregnant ewes with singleton fetuses (129 +/- 1 day) were chronically prepared with fetal vascular and lateral ventricle catheters and electrocorticogram (ECoG) and esophageal electromyogram electrodes. After a 2-h control period, fetuses were given lateral ventricle injection of NO synthase inhibitor nitro-L-arginine methyl ester (L-NAME) and monitored for 2 h. NO precursor L-arginine was then injected into the lateral ventricle, and fetuses were monitored for a final 2 h. All fetuses received an additional control study of fetal swallowing before and after lateral ventricle injection of artificial cerebrospinal fluid (aCSF). Data were analyzed with repeated-measures ANOVA and paired t-test (P < 0.05). Suppression of a central NO with central L-NAME significantly reduced mean (+/-SE) spontaneous fetal swallowing (1.2 +/- 0.1-0.6 +/- 0.1 swallows/min low-voltage ECoG; P < 0.01). Restoration of central NO by L arginine significantly increased fetal swallowing to pre-L-NAME levels (1.2 +/- 0.1 swallows/min low voltage). There were no changes in fetal swallowing during the control study of aCSF. Fetal ECoG activity and blood pressure did not change during the study or control aCSF injection. We conclude that NO is an important neuromodulator of fetal swallowing activity. Central NO synthase activity may contribute to the heightened level of spontaneous fetal swallowing and thus amniotic fluid regulation. PMID- 10516236 TI - Cardiovascular and renal responses produced by central orphanin FQ/nociceptin occur independent of renal nerves. AB - The present study investigated the role of the renal nerves in mediating the cardiovascular and renal responses produced by the central administration of the opioid-like peptide orphanin FQ/nociceptin (OFQ/N) in conscious Sprague-Dawley rats. In conscious rats, OFQ/N (10 microgram icv) produced a transient bradycardia and hypotension (nadir 20 min). Although renal sympathetic nerve activity (RSNA) initially remained unchanged, a delayed renal sympathoinhibitory response occurred after recovery (30 min) of blood pressure. By 30 and 70 min postinjection, RSNA decreased to 75 and 66% of control, respectively. Coinciding with the decrease in RSNA, central OFQ/N elicited a diuresis and antinatriuresis that occurred independent of changes in renal hemodynamics. In other studies, intracerebroventricular OFQ/N produced similar cardiovascular and renal excretory responses in bilaterally renal-denervated rats. Finally, in conscious sinoaortic deafferentiated rats, intracerebroventricular OFQ/N produced a rapid decrease in RSNA (55% of control, 10 min; 38% of control, 20 min) that paralleled the onset of the hypotension and bradycardia. These studies demonstrate that in conscious rats, intracerebroventricular OFQ/N produces a centrally mediated inhibition of RSNA which, due to activation of baroreflex mechanisms, is temporally dissociated from the hypotensive and bradycardia responses. As revealed in renal-denervated rats, the cardiovascular and renal excretory responses produced by central OFQ/N occur by a pathway that is independent of intact renal nerves or changes in renal hemodynamics. PMID- 10516237 TI - Hemodynamic effects of platelet-activating factor in nonpregnant and pregnant sheep. AB - The present study was designed to assess the dose-related effects of platelet activating factor (PAF) on systemic, renal, and uterine hemodynamics in nonpregnant sheep and to evaluate how pregnancy might alter these responses. Nonpregnant and pregnant (110 +/- 5 days gestation) ewes were instrumented for conscious measurements of maternal mean arterial pressure (MAP), renal blood flow (RBF), uterine blood flow (UBF), hematocrit, and urinary protein concentration. After recovery, dose-response curves to PAF were generated by systemic infusion at 10, 30, and 100 ng. kg(-1). min(-1) (15 min/dose) into the maternal femoral vein. The above parameters were measured, and renal and uterine vascular resistances (RVR and UVR, respectively) were calculated. In pregnant sheep, PAF increased MAP, RVR, UVR, and urinary protein concentration. We also observed increases in hematocrit, indicative of reduced blood volume secondary to increased systemic microvascular protein permeability. These responses were similar in nonpregnant sheep, with the exception of UVR in nonpregnant ewes being decreased (and thus UBF was increased), whereas in pregnant sheep, UVR was increased, which resulted in decreased UBF. This suggests that pregnancy alters the mechanism of action of PAF within the uterine vasculature in a way that can reduce UBF and thereby potentially compromise placental perfusion. PMID- 10516238 TI - Vesicoanal, urethroanal, and urethrovesical reflexes initiated by lower urinary tract irritation in the rat. AB - Irritation of the urinary bladder causes activation of normally "silent" nociceptive primary afferent fibers. In the present study, it is reported that irritation of the urinary bladder or urethra with infusion of 0.5% acetic acid robustly activates motoneurons that innervate the striated muscle of the external anal sphincter via spinal reflex mechanisms. The activation of anal motoneurons following irritation of the bladder and urethra are termed vesicoanal and urethroanal reflexes, respectively. The reflexes can be mimicked by acute application of capsaicin to the bladder and urethra, and they show desensitization following prolonged topical application of capsaicin or following chronic systemic pretreatment with capsaicin. The reflexes can be demonstrated in chronic spinal cord-transected animals, indicating that the reflex pathways are organized within the spinal cord. The urethroanal reflex is also physiologically activated by urethral distension and/or increases in intraluminal pressure. In addition to activation of anal sphincter activity, slight distension, pressure increases, or instillation of 0.5% acetic acid into the urethra inhibited bladder contractions through activation of an inhibitory urethrovesical reflex. These reflexes are discussed in terms of clinical characteristics of urethritis and prostatitis. Anecdotally, it was discovered that the bladder can buffer acetic acid. PMID- 10516239 TI - Strenuous resistive breathing induces proinflammatory cytokines and stimulates the HPA axis in humans. AB - Interleukin-1beta (IL-1beta) and interleukin-6 (IL-6), powerful stimulants of the hypothalamic-pituitary-adrenal (HPA) axis, increase in response to whole body exercise. Strenuous inspiratory resistive breathing (IRB), a form of clinically relevant "exercise" for the respiratory muscles, produces beta-endorphin through a largely unknown mechanism. We investigated (in 11 healthy humans) whether strenuous IRB produces proinflammatory cytokines and beta-endorphin in parallel with stimulation of the HPA axis, assessed by concurrent measurement of ACTH. Subjects underwent either severe [at 75% of maximal inspiratory pressure (P(m) (max))] or moderate (at 35% of P(m) (max)) IRB. Plasma cytokines, beta-endorphin, and ACTH were measured at rest (point R), at the point at which the resistive load could not be sustained (point F), and at exhaustion [15 min later (point E)]. During severe IRB, IL-1beta increased from 0.83 +/- 0.12 pg/ml at point R to 1.88 +/- 0. 53 and 4.06 +/- 1.27 pg/ml at points F and E, respectively (P < 0. 01). IL-6 increased from 5.30 +/- 1.02 to 10.33 +/- 2.14 and 11.66 +/- 2.29 pg/ml at points F and E, respectively (P = 0.02). ACTH and beta-endorphin fluctuated from 20.87 +/- 5.49 and 25.03 +/- 3.97 pg/ml at point R to 22.97 +/- 4.41 and 26.32 +/- 3.93 pg/ml, respectively, at point F and increased to 46.96 +/- 8.55 and 40.32 +/- 5.94 pg/ml, respectively, at point E (P < 0.01, point E vs. point F). There was a positive correlation between the IL-6 at point F and the ACTH and beta-endorphin at point E (r = 0.88 and 0.94, respectively; P < 0.01) as well as between the increase in IL-6 (between points R and F) and the increases in ACTH and beta-endorphin (between points F and E, r = 0.91 and 0.92, respectively; P < 0.01). Moderate IRB did not produce any change. We conclude that severe IRB produces proinflammatory cytokines and stimulates the HPA axis in humans secondary to the production of cytokines (especially IL-6). PMID- 10516240 TI - Interactive effects of central leptin and peripheral fuel oxidation on estrous cyclicity. AB - A 48-h period of fasting inhibits estrous cycles in Syrian hamsters, and fasting induced anestrus can be prevented by intracerebroventricular treatment with leptin during the fasting period. In the present experiment, the effects of intracerebroventricular leptin were blocked by systemic treatment with inhibitors of metabolic fuel oxidation. Leptin was infused continuously into the lateral ventricles (1 microgram/day) during fasting on days 1 and 2 of the estrous cycle. Intraperitoneal injection of 2-deoxy-D-glucose (2DG) was used to block both central and peripheral glucose oxidation, and intragastric treatment with methyl palmoxirate (MP) was used to inhibit peripheral long-chain fatty acid oxidation during the fasting and leptin-treatment period. 2DG or MP were administered at doses that did not induce anestrus in ad libitum-fed hamsters. Despite elevated central levels of leptin, fasting-induced anestrus occurred in hamsters treated with either 2DG or MP. Thus an elevated intracerebroventricular leptin concentration is not a sufficient condition for normal estrous cycles when fuel oxidation is inhibited. These results raise the possibility that central leptin influences reproduction by indirect effects on peripheral fuel metabolism. PMID- 10516241 TI - Nociceptin modulates renal sympathetic nerve activity through a central action in conscious rats. AB - Nociceptin, an endogenous agonist of the opioid receptor-like(1) receptor, is expressed in the hypothalamus, where it is implicated in autonomic nervous system control. However, the central actions of nociceptin on sympathetic nerve activity have not been studied. We investigated the effect of intracerebroventricularly administered nociceptin (2-10 nmol) on blood pressure, heart rate (HR), and renal sympathetic nerve activity (RSNA) in conscious rats and sinoaortic-denervated (SAD) rats. Intracerebroventricularly administered nociceptin resulted in a dose dependent decrease in mean arterial pressure (MAP) and HR in intact rats. RSNA decreased 31.5 +/- 2.1 and 19.9 +/- 5.0% at a dose of 2 and 5 nmol, respectively. In SAD rats, MAP, HR, and RSNA decreased in a dose-dependent manner, and the maximum responses were larger than those in intact rats. The decrease in HR induced by nociceptin was blocked by propranolol but not by atropine, which indicates that nociceptin is acting by inhibiting cardiac sympathetic outflow. These nociceptin-induced depressor and bradycardic responses were not antagonized by pretreatment with naloxone and nocistatin. These findings suggest that central nociceptin may have a functional role in regulating cardiovascular and sympathetic nervous systems. PMID- 10516242 TI - Effect of adrenocorticotrophic hormone on sodium appetite in mice. AB - A main vector of the effects of stress is secretion of corticotrophin releasing factor (CRF), adrenocorticotrophin (ACTH), and adrenal steroids. Systemic administration of ACTH (2.8 microgram/day sc) for 7 days in BALB/c mice caused a very large increase of voluntary intake of 0.3 M NaCl equivalent to turnover of total body sodium content each day. Intracerebroventricular infusion of ACTH (20 ng/day) had no effect. Intracerebroventricular infusion of ovine CRF (10 ng/h for 7 days) caused an increase of sodium intake. The large sodium appetite stimulating effect of systemic ACTH was not influenced by concurrent systemic infusion of captopril (2 mg/day). Induction of stress by immobilization of mice on a running wheel caused an increase in Na appetite associated with a 50% decrease of thymus weight, indicative of corticosteroid effects. The present data suggest that stress and the hormone cascade initiated by stress evoke a large sodium appetite in mice, which may be an important survival mechanism in environmental conditions causing stress. PMID- 10516243 TI - Effect of an exercise-heat acclimation program on body fluid regulatory responses to dehydration in older men. AB - We examined if an exercise-heat acclimation program improves body fluid regulatory function in older subjects, as has been reported in younger subjects. Nine older (Old; 70 +/- 3 yr) and six younger (Young; 25 +/- 3 yr) male subjects participated in the study. Body fluid regulatory responses to an acute thermal dehydration challenge were examined before and after the 6-day acclimation session. Acute dehydration was produced by intermittent light exercise [4 bouts of 20-min exercise at 40% peak rate of oxygen consumption (VO(2 peak)) separated by 10 min rest] in the heat (36 degrees C; 40% relative humidity) followed by 30 min of recovery without fluid intake at 25 degrees C. During the 2-h rehydration period the subjects drank a carbohydrate-electrolyte solution ad libitum. In the preacclimation test, the Old lost approximately 0.8 kg during dehydration and recovered 31 +/- 4% of that loss during rehydration, whereas the Young lost approximately 1.2 kg and recovered 56 +/- 8% (P < 0.05, Young vs. Old). During the 6-day heat acclimation period all subjects performed the same exercise-heat exposure as in the dehydration period. Exercise-heat acclimation increased plasma volume by approximately 5% (P < 0.05) in Young subjects but not in Old. The body fluid loss during dehydration in the postacclimation test was similar to that in the preacclimation in Young and Old. The fractional recovery of lost fluid volume during rehydration increased in Young (by 80 +/- 9%; P < 0.05) but not in Old (by only 34 +/- 5%; NS). The improved recovery from dehydration in Young was mainly due to increased fluid intake with a small increase in the fluid retention fraction. The greater involuntary dehydration (greater fluid deficit) in Old was accompanied by reduced plasma vasopressin and aldosterone concentrations, renin activity, and subjective thirst rating (P < 0.05, Young vs. Old). Thus older people have reduced ability to facilitate body fluid regulatory function by exercise-heat acclimation, which might be involved in attenuation of the acclimation-induced increase in body fluid volume. PMID- 10516244 TI - Role of ET(B) receptors and nitric oxide in adrenal catecholamine secretion in anesthetized dogs. AB - We examined the effects of sarafotoxin 6c (S6c), an endothelin-B (ET(B)) receptor agonist, on adrenal catecholamine secretion in response to cholinergic stimuli in pentobarbital sodium-anesthetized dogs. Drugs were administered intra-arterially into the adrenal gland through the phrenicoabdominal artery. Infusion of S6c attenuated increases in adrenal catecholamine output induced by splanchnic nerve stimulation. The inhibitory effect of S6c on the catecholamine secretion response was suppressed with a selective ET(B) receptor antagonist N-cis 2, 6 dimethylpiperidinocarbonyl-L-gamma-methylleucyl-D-1-methoxycarbonyl tryptophanyl D-norleucine (BQ-788), a nitric oxide synthase (NOS) inhibitor N(omega)-nitro-L arginine methyl ester, and a neuronal NOS inhibitor 7-nitroindazole monosodium salt (7-NINA). Similar results were obtained with the catecholamine secretion response induced by injection of ACh. 7-NINA alone did not affect these catecholamine secretion responses. These results suggest that ET(B) receptors play an inhibitory role in adrenal catecholamine secretion by activating neuronal NOS, whereas neuronal NOS is unlikely to be involved in regulation of adrenal catecholamine secretion in the absence of simultaneous ET(B) receptor stimulation. PMID- 10516245 TI - Role of cholinergic receptors in adrenal catecholamine secretion in spontaneously hypertensive rats. AB - We investigated the role of nicotinic and muscarinic receptors in secretion of catecholamines induced by transmural electrical stimulation (ES) from isolated perfused adrenal glands of spontaneously hypertensive rats (SHRs) and normotensive Wistar-Kyoto (WKY) rats. ES (1-10 Hz) produced frequency-dependent increases in epinephrine (Epi) and norepinephrine (NE) output as measured in perfusate. The ES-induced increases in NE output, but not Epi output, were significantly greater in adrenal glands of SHRs than in those of WKY rats. Hexamethonium (10-100 microM) markedly inhibited the ES-induced increases in Epi and NE output from adrenal glands of SHRs and WKY rats. Atropine (0.3-3 microM) inhibited the ES-induced increases in Epi and NE output from adrenal glands of SHRs, but not from those of WKY rats. These results suggest that endogenous acetylcholine-induced secretion of adrenal catecholamines is predominantly mediated by nicotinic receptors in SHRs and WKY rats and that the contribution of muscarinic receptors may be different between these two strains. PMID- 10516246 TI - Lesions of the C1 catecholaminergic neurons of the ventrolateral medulla in rats using anti-DbetaH-saporin. AB - Phenylethanolamine-N-methyltransferase (PNMT)-containing neurons in the rostral ventrolateral medulla (RVLM) are believed to play a role in cardiovascular regulation. To determine whether injection of anti-dopamine beta-hydroxylase (DbetaH)-saporin directly into the RVLM in rats could selectively destroy these cells and thereby provide an approach for evaluating their role in cardiovascular regulation, we studied rats 2 wk after unilateral injection of 21 ng anti-DbetaH saporin into the RVLM. There was an approximately 90% reduction in the number of PNMT-positive neurons in the RVLM, although the number of non-C1, spinally projecting barosensitive neurons of this area was not altered. The A5 cell group was the only other population of DbetaH-containing cells that was significantly depleted. The depressor response evoked by injection of tyramine into the RVLM was abolished by prior injection of toxin. The pressor response evoked by injection of glutamate into the RVLM was attenuated ipsilateral to the toxin injection but was potentiated contralateral to the toxin injection. Thus anti DbetaH-saporin can be used to make selective lesions of PNMT-containing cells, allowing for the evaluation of their role in cardiovascular regulation. PMID- 10516248 TI - Contributions of MSNA and stroke volume to orthostatic intolerance following bed rest. AB - We examined whether the altered orthostatic tolerance following 14 days of head down tilt bed rest (HDBR) was related to inadequate sympathetic outflow or to excessive reductions in cardiac output during a 10- to 15-min head-up tilt (HUT) test. Heart rate, blood pressure (BP, Finapres), muscle sympathetic nerve activity (MSNA, microneurography), and stroke volume blood velocity (SVV, Doppler ultrasound) were assessed during supine 30 degrees (5 min) and 60 degrees (5-10 min) HUT positions in 15 individuals who successfully completed the pre-HDBR test without evidence of orthostatic intolerance. Subjects were classified as being orthostatically tolerant (OT, n = 9) or intolerant (OI, n = 6) following the post HDBR test. MSNA, BP, and SVV during supine and HUT postures were not altered in the OT group. Hypotension during 60 degrees HUT in the post-bed rest test for the OI group (P < 0.05) was associated with a blunted increase in MSNA (P < 0.05). SVV was reduced following HDBR in the OI group (main effect of HDBR, P < 0.02). The data support the hypothesis that bed rest-induced orthostatic intolerance is related to an inadequate increase in sympathetic discharge that cannot compensate for a greater postural reduction in stroke volume. PMID- 10516247 TI - Activation of antilipolytic alpha(2)-adrenergic receptors by epinephrine during exercise in human adipose tissue. AB - The involvement of the antilipolytic alpha(2)-adrenergic pathway and the specific role of epinephrine in the control of lipolysis during exercise in adipose tissue (AT) were investigated in healthy male subjects (age: 24.1 +/- 2.2 yr; body mass index: 23.0 +/- 1.6). An in vitro study carried out on isolated adipocytes showed that the weak lipolytic effect of epinephrine was potentiated after blockade of alpha(2)-adrenergic receptor (AR) by an alpha(2)-AR antagonist and reached that of isoproterenol, a beta-AR agonist. The effect of the nonselective alpha(2)-AR antagonist phentolamine on the response of the extracellular glycerol concentration (EGC) in AT during two successive bouts of aerobic exercise (50% maximum O(2) uptake, 60 min duration) was evaluated using the microdialysis method. The metabolic responses measured in perfused probes with Ringer solution were compared with those obtained in perfused probes with Ringer plus 0.1 mmol/l phentolamine. Plasma norepinephrine level was not different during the two exercise bouts, whereas that of epinephrine was 2.5-fold higher during the second exercise. EGC in AT was twofold higher in the second compared with the first exercise, and the same response pattern was found for plasma glycerol. The exercise-induced increase in EGC was higher in the probe perfused with phentolamine compared with the control probe in both bouts of exercise. However, the potentiating effect of phentolamine on EGC was significant during the second exercise bout but did not reach a significant level during the first. These results suggest that epinephrine is involved in the control of lipid mobilization through activation of antilipolytic alpha(2)-AR in human subcutaneous AT during exercise. PMID- 10516249 TI - Seasonal changes of human circadian rhythms in Antarctica. AB - The human circadian rhythms in sleep, activity, plasma melatonin, and rectal temperature were explored under two conflicting time cues in Antarctica: an extreme photoperiod and a strict work schedule. The nine healthy male subjects stayed at the Antarctic zone (latitude 66.5-90 degrees south) for 15 mo including a 13-mo wintering at the Dome station (latitude 77 degrees south). Neither the phases nor the amounts of sleep and daily activity underwent a seasonal change. On the other hand, the peak phase of melatonin rhythm was phase delayed by 4.1 h in winter compared with summer. When the analysis is limited to the Dome data, the seasonal difference was reduced to 1.3 h. Similarly the trough phase of rectal temperature rhythm in two of three subjects was phase delayed by approximately 2 h in winter. From these findings, the sleep or activity rhythm is concluded to be reset predominantly by the work schedule, whereas the circadian rhythm in plasma melatonin and rectal temperature is substantially influenced by the photoperiod. PMID- 10516250 TI - Enantioselectivity of odor perception in squirrel monkeys and humans. AB - With use of a conditioning paradigm, the ability of six squirrel monkeys to distinguish between 10 pairs of enantiomers, i.e., odorants that are identical except for chirality, was investigated. As a group, the animals were only able to discriminate between the optical isomers of alpha-pinene, carvone, limonene, and fenchone, whereas they failed to distinguish between the (+) and (-) forms of beta-citronellol, menthol, rose oxide, 2-butanol, alpha-terpineol, and camphor. With use of a triple forced-choice procedure, 10 human subjects were tested for their ability to discriminate between the same enantiomeric odor pairs in parallel and, with the exception of fenchone, showed a very similar pattern of performance compared with the squirrel monkeys. These findings support the assumption that human and nonhuman primates may share common principles of odor quality perception. Furthermore, the results suggest that, in both species, enantioselective molecular odor receptors may only exist for some, but not all volatile enantiomers and thus that chiral recognition of odorants is not a general phenomenon, but may be restricted to some substances. PMID- 10516251 TI - Effect of brain stem NMDA-receptor blockade by MK-801 on behavioral and fos responses to vagal satiety signals. AB - To test the possible role of N-methyl-D-aspartate (NMDA) glutamate receptors in the transmission of gastrointestinal satiety signals at the level of the nucleus of the solitary tract (NTS), we assessed the effect of fourth ventricular infusion of the noncompetitive NMDA receptor antagonist MK-801 on short-term sucrose intake and on gastric distension-induced Fos expression in the dorsal vagal complex of unanesthetized rats. MK-801, although not affecting initial rate of intake, significantly increased sucrose intake during the later phase of the meal (10-30 min, 8.9 +/- 1.0 vs. 2.9 +/- 0.8 ml, P < 0.01). In the medial subnucleus of the NTS, the area postrema, and the dorsal motor nucleus, MK-801 did not reduce gastric distension-induced Fos expression and itself did not significantly induce Fos expression. In the dorsomedial, commissural, and gelatinosus subnuclei, MK-801 in itself produced significant Fos expression and significantly reduced (-75%, P < 0.05) the ability of gastric distension to induce Fos expression, assuming an additive model with two separate populations of neurons activated by distension and the blocker. Although these results are consistent with NMDA receptor-mediated glutamatergic transmission of vagal satiety signals in general, they lend limited support for such a role in the transmission of specific gastric distension signals. PMID- 10516252 TI - Characterization of cis-elements required for osmotic response of rat Na(+)/H(+) exchanger-2 (NHE-2) gene. AB - The Na(+)/H(+) exchanger (NHE-2) has been implicated in osmoregulation in the kidney, because it transports Na(+) across the cell membrane and efficiently alters intracellular osmolarity. On hyperosmotic stress, NHE-2 mRNA increases in abundance in mouse inner medullary collecting duct (mIMCD-3) cells, suggesting possible transcriptional regulation. To investigate the molecular mechanism of potential transcriptional regulation of NHE-2 by hyperosmolarity, we have functionally characterized the 5'-flanking region of the gene in mIMCD-3 cells. Transient transfection of luciferase reporter gene constructs revealed a novel cis-acting element, which we call OsmoE (osmotic-responsive element, bp -808 to 791, GGGCCAGTTGGCGCTGGG), and a TonE-like element (tonicity-responsive element, bp -1201 to -1189, GCTGGAAAACCGA), which together are shown to be responsible for hyperosmotic induction of the NHE-2 gene. Electrophoretic mobility shift assays suggest that different DNA-protein interactions occur between these two osmotic response elements. However, both DNA sequences were shown to specifically bind nuclear proteins that dramatically increase in abundance under hyperosmotic conditions. Isolation of trans-acting factors and characterization of their specific interaction with these osmotic response elements will further elucidate the transcriptional mechanisms controlling NHE-2 gene expression under hyperosmolar conditions. PMID- 10516253 TI - Differential effects from parapyramidal region and rostral ventrolateral medulla mediated by substance P. AB - Rostral ventrolateral medulla (rVLM) and parapyramidal region (PPr) serve as important medullary control sites for sympathoexcitation. rVLM and PPr have direct projections to the intermediolateral cell column (IML) that are thought to be important in maintaining mean arterial blood pressure (MAP). Substance P (SP) is found in PPr neurons and in and near the subretrofacial area of the rVLM. At least some of these cells project to the IML. We investigated the involvement of SP at the IML in mediating rVLM- and PPr-evoked pressor responses in the chloralose-anesthetized cat. Pressor responses to electrical and chemical PPr and rVLM stimulation were altered after intrathecal injection, at the level of the T1 T3 spinal cord, of either SP antagonist [D-Pro(2), D-Phe(7), D-Trp(9)]-SP, SP antagonist CP 96,345, or SP antiserum. Although MAP and heart rate responses to PPr stimulation were attenuated by intrathecal SP antagonists or antiserum, MAP responses to rVLM stimulation were augmented. Previous studies have revealed differences in transmitters associated with these two areas, even though the general response of both areas is sympathoexcitatory. The present study implies that the identical substance may increase or decrease the MAP response depending on the pathway activated. PMID- 10516254 TI - Developmental changes in renal renin mRNA half-life and responses to stimulation in fetal lambs. AB - In the perinatal period there is increased renin gene expression in the kidney compared with other stages of development. This may be related to changes in responsiveness of the renin gene to stimulation and/or differences in renin mRNA stability as development progresses. To ascertain if either responsiveness or stability changes in fetal life, we studied renin mRNA levels in primary cultures of renal cortical cells obtained from fetal lamb kidneys at two stages (0.7 and 0.9) of gestation after stimulation with isoproterenol, forskolin, or isobutyl methylxanthine and after inhibition of transcription with actinomycin D. Forskolin and isobutyl methylxanthine rapidly increased renin mRNA by at least twofold in the cultured cells from fetuses of both ages, with the sensitivity to stimulation higher in the cells from the mature fetal kidneys. Isoproterenol was effective only in mature fetal cells. In addition, the decay of renin mRNA after cessation of transcription was slower in mature cells compared with immature cells, the half-life being 11.6 +/- 0.8 h in mature cells and 6.6 +/- 0.6 h in immature cells (P < 0.05). The data suggest that increases in both renin mRNA sensitivity to stimulation and in stability can contribute to the enhanced renin expression in the perinatal period. PMID- 10516255 TI - Effect of hepatic glucose infusion on glucose intake and licking microstructure in deprived and nondeprived rats. AB - The effects of hepatic-portal glucose or saline infusions on intake and the temporal distribution of licking (lick microstructure) were evaluated in nondeprived and in 20.5-h food-deprived rats. Rats received portal infusions of isotonic glucose or saline (0.1 ml/min) for 2 h before and then throughout a 90 min period of access to a spout that delivered 12.5% glucose. Overall, a significant treatment-related intake suppression was obtained in nondeprived but not in deprived groups. For both groups, however, there was a significant positive linear relationship between the amount individual rats consumed under the saline (baseline) infusion condition and the extent to which portal glucose infusion suppressed intake. The linear fit for the deprived group was similar in slope, but right shifted, relative to the best fit for the nondeprived group. The individual-subject and group differences in response to portal glucose infusion are discussed in relation to the inconsistent literature on this treatment's short-term intake effects. We focused analysis of the licking pattern on those rats for which a prominent portal glucose infusion effect was obtained (i.e., nondeprived rats with higher than average baseline intakes). Features of the lick pattern associated with taste evaluation (1st min lick rate; lick burst duration) were not significantly affected by portal glucose infusion. Rather, the minute-by minute rate of ingestion under glucose infusion declined more rapidly than under baseline tests, indicating that portal glucose infusion enhanced the inhibitory influence of the accumulating postingestive load. PMID- 10516256 TI - Decreased responsiveness to dietary fat in Otsuka Long-Evans Tokushima fatty rats lacking CCK-A receptors. AB - Adult Otsuka Long-Evans Tokushima fatty (OLETF) rats lack functional cholecystokinin A (CCK-A) receptors, are diabetic, hyperphagic, and obese, and have patterns of ingestion consistent with a satiety deficit secondary to CCK insensitivity. Because dietary fat potently stimulates CCK release, we examined how dietary fat modulates feeding in adult male OLETF rats and their lean [Long Evans Tokushima (LETO)] controls. High-fat feeding produced sustained overconsumption of high-fat diet (30% corn oil in powdered chow) over a 3-wk period in OLETF but not LETO rats. We then assessed the ability of gastric gavage (5 ml, 1-2 kcal/ml x 15 s) or duodenal preloads (1 kcal/ml, 0.44 ml/min x 10 min) of liquid carbohydrate (glucose), protein (peptone), or fat (Intralipid) to suppress subsequent 30-min 12.5% glucose intake in both strains. In OLETF rats, gastric and duodenal fat preloads were significantly less effective in suppressing subsequent intake than were equicaloric peptone or glucose. These results demonstrate that OLETF rats fail to compensate for fat calories and suggest that their hyperphagia and obesity may stem from a reduced ability to process nutrient-elicited gastrointestinal satiety signals. PMID- 10516257 TI - Circadian temperature and melatonin rhythms, sleep, and neurobehavioral function in humans living on a 20-h day. AB - The interaction of homeostatic and circadian processes in the regulation of waking neurobehavioral functions and sleep was studied in six healthy young subjects. Subjects were scheduled to 15-24 repetitions of a 20-h rest/activity cycle, resulting in desynchrony between the sleep-wake cycle and the circadian rhythms of body temperature and melatonin. The circadian components of cognitive throughput, short-term memory, alertness, psychomotor vigilance, and sleep disruption were at peak levels near the temperature maximum, shortly before melatonin secretion onset. These measures exhibited their circadian nadir at or shortly after the temperature minimum, which in turn was shortly after the melatonin maximum. Neurobehavioral measures showed impairment toward the end of the 13-h 20-min scheduled wake episodes. This wake-dependent deterioration of neurobehavioral functions can be offset by the circadian drive for wakefulness, which peaks in the latter half of the habitual waking day during entrainment. The data demonstrate the exquisite sensitivity of many neurobehavioral functions to circadian phase and the accumulation of homeostatic drive for sleep. PMID- 10516258 TI - Allometric scaling of RNA, DNA, and enzyme levels: an intraspecific study. AB - The activities of oxidative and glycolytic enzymes show body size-dependent relationships across a wide variety of taxa; however, the mechanistic basis remains unknown. We sampled white epaxial muscle from rainbow trout (Oncorhynchus mykiss) spanning a 100-fold range in body mass. We measured activities of enzymes from aerobic and anaerobic metabolic pathways, RNA [total RNA and mRNA, pyruvate kinase (PK), citrate synthase (CS), and MyoD mRNA], and total DNA. Total RNA and DNA showed a biphasic relationship with body size, with a break point occurring after fish reached 1 yr of age. In contrast, total RNA/total DNA was constant across the entire size range. Neither CS activity nor CS mRNA levels scaled with body mass. PK activity and PK mRNA levels increased in parallel in yearling fish only (r(2) = 0.91, P < 0.01). This suggests that although PK expression is transcriptionally regulated in yearlings, the molecular mechanisms regulating expression change with growth and age. This was supported by a positive correlation between MyoD and PK mRNA levels (r(2) = 0.17, P < 0.05). PMID- 10516259 TI - Emergence of altered circadian timing in a cholinergically supersensitive rat line. AB - Mammalian circadian rhythms are controlled by the suprachiasmatic nuclei (SCN) in concert with light information. Several neurotransmitters and neural pathways modulate light effects on SCN timing. This study used a line of rat with an upregulated cholinergic system to investigate the role of acetylcholine in rhythmicity. With the use of a selective breeding program based on the thermic response to a cholinergic agonist, we developed a supersensitive (S(ox)) and subsensitive (R(ox)) rat line. The S(ox) rats showed an earlier onset time of melatonin rhythm under a 12:12-h light-dark photoperiod from generation 3 (3 +/- 0.5 h after dark) compared with R(ox) rats (4.5 +/- 0.1 h) and an earlier morning decline in temperature (0.9 +/- 0.3 h before lights on) compared with R(ox) animals (0.1 +/- 0.1 h). Furthermore, the S(ox) animals displayed a significantly shorter free-running period of temperature rhythm than R(ox) rats (23.9 +/- 0.04 and 24.3 +/- 0.1 h, respectively, P < 0.05). The results suggest that the altered circadian timing of the S(ox) rats may be related to the cholinergic supersensitivity, intimating a role for acetylcholine in the circadian timing system. PMID- 10516261 TI - Feeding status and bacterial LPS-induced cytokine and neuropeptide gene expression in hypothalamus. AB - This study determined the effects of feeding status on basal and lipopolysaccharide (LPS)-stimulated cytokine and neuropeptide gene expression in the hypothalamus. With the use of RNase protection assays, we measured mRNA levels of interleukin-1beta (IL-1beta), IL-1 receptor antagonist (IL-1RA), IL-1 receptor type I (IL-1RI), IL-1R accessory proteins (AcP I and II), tumor necrosis factor-alpha (TNF-alpha), transforming growth factor-beta1 (TGF-beta1), glycoprotein 130 (Gp 130), leptin receptor (OB-R), neuropeptide Y (NPY), preprodynorphin, and proopiomelanocortin (POMC). Analyses were done in ad libitum fed, fasted, and fasted and refed rats treated with the intracerebroventricular administration of physiological saline or LPS. The data show that food deprivation increases the basal mRNA expression of IL-1beta, IL-1RA, TNF-alpha, IL-1RI, and IL-1R AcP I, whereas mRNA levels of POMC showed a decrease. Five hours of refeeding returned cytokine levels to those observed in the ad libitum fed group. LPS administration induced a robust upregulation of IL-1beta, TNF alpha, and IL-1RI during all three feeding conditions. Acute food deprivation did not modulate LPS-induced changes in hypothalamic cytokine mRNA profiles. These findings show that 1) cytokine modulation occurs as an adaptive response to the stress of acute fasting and 2) acute fasting does not affect LPS-induced cytokine mRNA modulation in the hypothalamus. The data have implications to gram-negative infections associated with acute anorexia. PMID- 10516260 TI - Analysis of vasoconstrictor responses to histamine in the hindlimb vascular bed of the rabbit. AB - Hemodynamic responses to histamine were investigated in the anesthetized rabbit. Intravenous injections of histamine induced dose-dependent decreases in systemic arterial pressure that were blocked by the H(1)-receptor antagonist pyrilamine but not the H(2) antagonist cimetidine. Injections of histamine and the H(1) agonist 6-[2-(4-imidazolyl)ethylamine]-N-(4-trifuormethylphenyl)-heptan ecardo xamide dimaleate (HTMT) into the hindlimb perfusion circuit increased hindlimb perfusion pressure, whereas the H(2) agonist dimaprit decreased perfusion pressure and the H(3)-receptor agonist R-(-)-alpha-methylhistamine did not alter perfusion pressure. Pyrilamine reduced hindlimb vasoconstrictor responses to histamine and HTMT but did not alter vasodilator responses to dimaprit. Cimetidine reduced the response to dimaprit but did not alter vasoconstrictor responses to histamine or HTMT. The H(3)-receptor antagonist thioperamide was without effect on responses to the histamine agonists. These data suggest the presence of H(1) and H(2) receptors and that histamine for the most part acts by stimulating H(1) receptors to produce vasoconstriction in the hindlimb vascular bed of the rabbit. Responses to histamine, HTMT, and norepinephrine were significantly enhanced by a nitric oxide synthase inhibitor at a time when vasodilator responses to dimaprit were unaltered and responses to acetylcholine were significantly reduced. Responses to histamine and the H(1) and H(2) agonists were not affected by the cyclooxygenase inhibitor meclofenamate or by ATP sensitive K(+) channel, alpha-adrenergic, or angiotensin AT(1) receptor antagonists. The present data suggest that H(1) receptors mediate both systemic vasodepressor and hindlimb vasoconstrictor responses to histamine. PMID- 10516262 TI - Novel TRH analog improves motor and cognitive recovery after traumatic brain injury in rodents. AB - Thyrotropin-releasing hormone (TRH) and certain TRH analogs show substantial neuroprotective effects in experimental brain or spinal cord trauma but also have other physiological actions (autonomic, analeptic, and endocrine) that may be undesirable for the treatment of neurotrauma in humans. We developed a novel TRH analog (2-ARA-53a), with substitutions at the NH(2)-terminus and imidazole ring, that preserves the neuroprotective action of TRH-like compounds while decreasing or eliminating their autonomic, analeptic, and endocrine effects. Rats administered 2-ARA-53a (1.0 mg/kg, n = 17) intravenously 30 min after lateral fluid percussion brain injury showed marked improvement in motor recovery compared with vehicle-treated controls (n = 14). Treatment of mice subjected to moderate controlled cortical impact brain injury, at the same dose and time after trauma (n = 8), improved both motor recovery and cognitive performance in a water maze place learning task compared with vehicle-treated controls (n = 8). In injured rats, no autonomic or analeptic effects were observed with this compound, and endocrine effects were significantly reduced with 2-ARA-53a, in contrast to those found with a typical NH(2)-terminal-substituted TRH analog (YM-14673). These findings demonstrate that the neuroprotective effects of TRH-related compounds can be dissociated from their other major physiological actions and suggest a potential role for dual-substituted TRH analogs in the treatment of clinical neurotrauma. PMID- 10516263 TI - Contractile function is unaltered in diaphragm from mice lacking calcium release channel isoform 3. AB - Skeletal muscle expresses at least two isoforms of the calcium release channel in the sarcoplasmic reticulum (RyR1 and RyR3). Whereas the function of RyR1 is well defined, the physiological significance of RyR3 is unclear. Some authors have suggested that RyR3 participates in excitation-contraction coupling and that RyR3 may specifically confer resistance to fatigue. To test this hypothesis, we measured contractile function of diaphragm strips from adult RyR3-deficient mice (exon 2-targeted mutation) and their heterozygous and wild-type littermates. In unfatigued diaphragm, there were no differences in isometric contractile properties (twitch characteristics, force-frequency relationships, maximal force) among the three groups. Our fatigue protocol (30 Hz, 0.25 duty cycle, 37 degrees C) depressed force to 25% of the initial force; however, lack of RyR3 did not accelerate the decline in force production. The force-frequency relationship was shifted to higher frequencies and was depressed in fatigued diaphragm; lack of RyR3 did not exaggerate these changes. We therefore provide evidence that RyR3 deficiency does not alter contractile function of adult muscle before, during, or after fatigue. PMID- 10516264 TI - Glucose and fatty acid metabolism in the isolated working mouse heart. AB - Although isolated perfused mouse heart models have been developed to study mechanical function, energy substrate metabolism has not been examined despite the expectation that the metabolic rate for a heart from a small mammal should be increased. Consequently, glucose utilization (glycolysis, oxidation) and fatty acid oxidation were measured in isolated working mouse hearts perfused with radiolabeled substrates, 11 mM glucose, and either 0.4 or 1.2 mM palmitate. Heart rate, coronary flow, cardiac output, and cardiac power did not differ significantly between hearts perfused at 0.4 or 1.2 mM palmitate. Although the absolute values obtained for glycolysis and glucose oxidation and fatty acid oxidation are significantly higher than those reported for rat hearts, the pattern of substrate metabolism in mouse hearts is similar to that observed in hearts from larger mammals. The metabolism of mouse hearts can be altered by fatty acid concentration in a manner similar to that observed in larger animals; increasing palmitate concentration altered the balance of substrate metabolism to increase overall energy derived from fatty acids from 64 to 92%. PMID- 10516265 TI - Focal delivery during direct infusion to brain: role of flow rate, catheter diameter, and tissue mechanics. AB - Direct interstitial infusion is a technique capable of delivering agents over both small and large dimensions of brain tissue. However, at a sufficiently high volumetric inflow rate, backflow along the catheter shaft may occur and compromise delivery. A scaling relationship for the finite backflow distance along this catheter in pure gray matter (x(m)) has been determined from a mathematical model based on Stokes flow, Darcy flow in porous media, and elastic deformation of the brain tissue: x(m) = constant Q(o)(3)R(4)r(c)(4)G(-3)mu(-1) 1/5 [corrected] = volumetric inflow rate, R = tissue hydraulic resistance, r(c) = catheter radius, G = shear modulus, and mu = viscosity). This implies that backflow is minimized by the use of small diameter catheters and that a fixed (minimal) backflow distance may be maintained by offsetting an increase in flow rate with a similar decrease in catheter radius. Generally, backflow is avoided in rat gray matter with a 32-gauge catheter operating below 0.5 microliter/min. An extension of the scaling relationship to include brain size in the resistance term leads to the finding that absolute backflow distance obtained with a given catheter and inflow rate is weakly affected by the depth of catheter tip placement and, thus, brain size. Finally, an extension of the model to describe catheter passage through a white matter layer before terminating in the gray has been shown to account for observed percentages of albumin in the corpus callosum after a 4-microliter infusion of the compound to rat striatum over a range of volumetric inflow rates. PMID- 10516266 TI - Regulation of in vitro renin secretion by ANG II feedback manipulation in vivo in the ovine fetus. AB - The renin-angiotensin system is critically important to fetal cardiovascular function and organ development. The feedback regulation of renin secretion by ANG II develops early in gestation yet does not linearly progress from fetal life to adulthood. Renin secretion is elevated in late gestation compared with earlier or postnatal time periods, which suggests that some component of the negative feedback regulation of renin secretion is less sensitive in late gestation. We examined in fetal sheep the age-related consequence of chronic in vivo manipulation of ANG II on renal renin secretion measured in vitro. Immature (101 103 days of gestation) and mature (130-133 days of gestation) fetuses were treated for 72 h with enalaprilat, ANG II or vehicle. Content and basal and isoproterenol-stimulated secretion of prorenin (PR) and active renin (AR) from fetal kidney cortical slices were determined. Enalaprilat pretreatment in vivo increased renal renin content and basal and stimulated secretion of PR and AR in vitro even in immature animals. Immunohistochemical localization showed that enalaprilat treatment caused an age-related recruitment of renin-containing juxtaglomerular cells. Conversely, ANG II pretreatment decreased basal and stimulated PR and AR secretion from immature fetal kidneys, but only inhibited PR secretion from mature kidneys. It also caused an age-related decrease in the percentage of renin-containing juxtaglomerular cells. These results suggest that ANG II feedback modulates not only the synthesis and content of renin, but the sensitivity of the coupling between stimulus and secretion. A critical observation of our study is that the higher renal tissue concentrations of prorenin and active renin in late gestation may be a consequence of reduced sensitivity to ANG II feedback; this is consistent with the increased plasma concentrations of renin found in near-term mammals. PMID- 10516267 TI - Optical imaging of the ventral medullary surface across sleep-wake states. AB - We hypothesized that spontaneous activity declines over widespread areas of the cat ventral medullary surface (VMS) during rapid eye movement (REM) sleep. We assessed neural and hemodynamic activity, measured as changes in reflected 660- and 560-nm wavelength light, from the VMS during sleep and waking states in five adult, unrestrained cats and in two control cats. Relative to quiet sleep, overall activity declined, and variability, assessed by standard deviation, increased by 25% during REM sleep. Variability in activity during waking also increased by 45% over quiet sleep, but mean activity was unchanged. REM sleep onset was preceded by a reduction in the hemodynamic signal from 5 to 60 s before neural activity decline. The activity decline during REM sleep, previously noted in the goat rostral VMS, extends to intermediate VMS areas of the cat and differs from most neural sites, such as the cortex, hippocampus, and thalamus, which increase activity during REM sleep. The activity decline during REM sleep has the potential to modify VMS responsiveness to baroreceptor and chemoreceptor challenges during the REM state. PMID- 10516268 TI - Introduction: glutamate transport, metabolism, and physiological responses. AB - The material covered in this set of articles was originally presented at Experimental Biology '98, in San Francisco, CA, on April 20, 1998. Here, the participants recount important elements of current research on the role of glutamate transporter activity in cellular signaling, metabolism, and organ function. W. A. Fairman and S. G. Amara discuss the five subtypes of human excitatory amino acid transporters, with emphasis on the EAAT4 subtype. M. A. Hediger discusses the expression and action of EAAC1 subtype of the human excitatory amino acid transporter. I. Nissim provides an overview of the significant role of pH in regulating Gln/Glu metabolism in the kidney, liver, and brain. J. D. McGivan and B. Nicholson describe some characteristics of glutamate transport regulation with regard to a specific experimental model of the bovine renal epithelial cell line NBL-1. Finally, T. C. Welbourne and J. C. Matthews introduce the "functional unit" concept of glutamate transport and how this relates to both glutamine metabolism and paracellular permeability. PMID- 10516269 TI - Functional diversity of excitatory amino acid transporters: ion channel and transport modes. AB - Recent studies of glutamate transporters in the central nervous system indicate that in addition to their fundamental role in mediating neurotransmitter uptake, these proteins may contribute to the modulation of a variety of cellular processes. Activation of the excitatory amino acid (EAA) carriers generates an electrogenic current attibutable to ion-coupled cotransport. In addition to this transport-associated current, a substrate-gated thermodynamically uncoupled anion flux has been identified that has been proposed to dampen neuronal excitability. Arachidonic acid has been reported to modulate a variety of membrane proteins involved in cellular signaling. Here we discuss recent findings that indicate arachidonic acid stimulates a previously uncharacterized proton-selective conductance in the Purkinje cell-specific subtype, EAAT4. The unique channel-like porperties of the EAATs, their unexpected localization, and physiological evidence propose a modulatory role for the EAATs in neuronal signaling and suggest a broader role for glutamate transporters than simply the clearance of synaptically released glutamate. Thus, the identification of this arachidonate stimulated proton conductance extends the complexity of mechanisms through which glutamate transporters modulate neuronal excitability. PMID- 10516270 TI - Glutamate transporters in kidney and brain. AB - Glutamate transporters play important roles in the termination of excitatory neurotransmission and in providing cells with glutamate for metabolic purposes. In the kidney, glutamate transporters are involved in reabsorption of filtered acidic amino acids, regulation of ammonia and bicarbonate production, and protection of cells against osmotic stress. PMID- 10516271 TI - Newer aspects of glutamine/glutamate metabolism: the role of acute pH changes. AB - This review focuses on the role of acute pH changes in the regulation of Gln/Glu metabolism in the kidney, liver, and brain. Alterations of proton concentration ([H(+)]) profoundly affect flux through phosphate-dependent glutaminase (PDG) or glutamate dehydrogenase (GDH), the primary enzymes responsible for mitochondrial metabolism of glutamine and glutamate, respectively. In the kidney, acute acidosis stimulates Gln uptake and its metabolism via the PDG pathway. The Glu formed from Gln can be removed via 1) oxidative deamination through the GDH reaction, 2) transamination reactions, and 3) transport of Glu from intracellular to extracellular compartment, thereby diminishing the intramitochondrial pool of glutamate sufficiently to stimulate flux through the PDG pathway. Converse changes may occur with increased pH. In the liver, acidosis diminishes the rate of Gln and Glu metabolism via the PDG and GDH pathways, but stimulates glutamine synthesis (i.e., glutamine recycling). Alkalosis has little effect. Hepatic Gln metabolism via the PDG pathway has a central role in ureagenesis via 1) supplementation of nitrogen for the synthesis of carbamyl phosphate, and 2) providing glutamate for N-acetylglutamate synthesis. In the brain, Gln/Glu metabolism links ammonia detoxification and energy metabolism via 1) detoxification of ammonia and excess glutamate by glutamine synthesis in astrocytes, 2) formation and export of glutamine to neurons where it is metabolized to glutamate and GABA, and 3) production of alpha-ketoglutarate and lactate from Glu and their transport to neurons. Changes in intracellular pH associated with changes in cellular [K(+)] may have a key role in the regulation of these processes of glial-neuronal metabolism of Gln/Glu metabolism. PMID- 10516272 TI - Regulation of high-affinity glutamate transport by amino acid deprivation and hyperosmotic stress. AB - High-affinity glutamate transport activity is induced by stress in NBL-1 cells. Exposure of cells to hyperosmotic medium led to an induction of the EAAC1 glutamate transporter, preceded by a large increase in EAAC1 mRNA levels. Culture of cells in amino acid-free medium also caused a protein synthesis-dependent increase in glutamate transport activity, but this was not accompanied by an increase of either EAAC1 mRNA or protein. Indirect evidence suggests that the increase in EAAC1 activity in the latter case may be due to the synthesis of an activator protein in response to decreased intracellular glutamate concentrations. PMID- 10516273 TI - Glutamate transport and renal function. AB - Brush border gamma-glutamyltransferase-glutaminase activity and the high-affinity glutamate transporter EAAC1 function as a unit in generating and transporting extracellular glutamate into proximal tubules as a signal that modulates intracellular glutamine/glutamate metabolism, paracellular permeability, and urinary acidification. The reported presence of a second glutamate transporter, GLT1, on the antiluminal tubule surface points to specific functional roles for each subtype in physiological and pathophysiological processes. PMID- 10516274 TI - Chloride dependency of renal brush-border membrane phosphate transport. AB - In our present study, we examined the effect of Cl(-) on rabbit renal brush border membrane (BBM) phosphate (P(i)) uptake. It was found that the Na(+) dependent BBM (32)P uptake was significantly inhibited by Cl(-) replacement in the uptake solution with other anions, or by Cl(-) transport inhibitors, including DIDS, SITS, diphenylamine-2-carboxylate (DPC), niflumic acid (NF), and 5-nitro-2-(3-phenylpropylamino)benzoate (NPPB). Intravesicular formate or Cl(-) increased BBM (36)Cl(-) uptake but did not affect BBM (32)P uptake. BBM (22)Na(+) uptake was lowered by Cl(-) replacement in the uptake solution but not by Cl(-) transport inhibitors. Changes in transmembrane electrical potential altered BBM (36)Cl(-) and (32)P uptake in directions consistent with a net inward movement of negative and positive charges, respectively. However, the Cl(-)-dependent BBM P(i) uptake was not affected by changes in transmembrane electrical potential. Finally, a similar Cl(-) dependency of P(i) uptake was also found with BBM derived from rat and mouse kidneys. In summary, our study showed that a component of Na(+)-dependent P(i) uptake was also Cl(-) dependent in rabbit, rat, and mouse renal BBM. The mechanism underlying this Cl(-) dependency remains to be identified. PMID- 10516275 TI - Real-time assessment of alpha-ketoglutarate effect on organic anion secretion in perfused rabbit proximal tubules. AB - To determine the quantitative roles of the basolateral and luminal Na(+) dicarboxylate (Na-DC) cotransporters in establishing and maintaining the alpha ketoglutarate (alphaKG) gradient required for renal tubular secretion of organic anions, we measured net steady-state transepithelial secretion of fluorescein (FL) in real time in isolated, perfused S2 segments of rabbit renal proximal tubules. Net "basal" FL secretion in the absence of exogenous alphaKG had a K(t) of approximately 4 microM and a maximal transepithelial secretion rate (J(max)) of approximately 380 fmol. min(-1). mm(-1) (where K(t) is the FL concentration that produces one-half the J(max)). It could be almost completely inhibited by basolateral p-aminohippurate (PAH). Selective inhibition of the basolateral Na-DC cotransporter indicated that recycling via this transporter of alphaKG that had been exchanged for FL supports approximately 25% of the "basal" FL secretion. Physiological alphaKG concentrations of 10 microM in the bath or 50 microM in the perfusate stimulated net secretion of FL by approximately 30 or approximately 20%, respectively. These data indicate that the basolateral Na-DC cotransporter supports approximately 42% of the net FL secretion. The luminal and basolateral effects of physiological concentrations of alphaKG were additive, indicating that the combined function of the luminal and basolateral Na-DC cotransporters can support approximately 50% of the net FL secretion. This apparently occurs by their establishing and maintaining approximately 50% of the outwardly directed alphaKG gradient that is responsible for driving basolateral FL/alphaKG exchange. The remaining approximately 50% would be maintained by metabolic production of alphaKG in the cells. PMID- 10516276 TI - A role for intracellular calcium in tight junction reassembly after ATP depletion repletion. AB - The integrity of the tight junction (TJ), which is responsible for the permeability barrier of the polarized epithelium, is disrupted during ischemic injury and must be reestablished for recovery. Recently, with the use of an ATP depletion-repletion model for ischemia and reperfusion injury in Madin-Darby canine kidney cells, TJ proteins such as zonula occludens-1 (ZO-1) were shown to reversibly form large complexes and associate with cytoskeletal proteins (T. Tsukamoto and S. K. Nigam, J. Biol. Chem. 272: 16133-16139, 1997). In this study, we examined the role of intracellular calcium in TJ reassembly after ATP depletion-repletion by employing the cell-permeant calcium chelator 1, 2-bis(2 aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-AM (BAPTA-AM). Lowering intracellular calcium during ATP depletion is associated with significant inhibition of the reestablishment of the permeability barrier following ATP repletion as measured by transepithelial electrical resistance and mannitol flux, marked alterations in the subcellular localization of occludin by immunofluorescent analysis, and decreased solubility of ZO-1 and other TJ proteins by Triton X-100 extraction assay, suggesting that lowering intracellular calcium potentiates the interaction of TJ proteins with the cytoskeleton. Coimmunoprecipitation studies indicated that decreased solubility may partly result from the stabilization of large TJ protein-containing complexes with fodrin. Although ionic detergents (SDS and deoxycholate) appeared to cause a dissociation of ZO-1-containing complexes from the cytoskeleton, sucrose gradient analyses of the solubilized proteins suggested that calcium chelation leads to self-association of these complexes. Together, these results raise the possibility that intracellular calcium plays an important facilitatory role in the reassembly of the TJ damaged by ischemic insults. Calcium appears to be necessary for the dissociation of TJ-cytoskeletal complexes, thus permitting functional TJ reassembly and paracellular permeability barrier recovery. PMID- 10516277 TI - Capacitative calcium entry in smooth muscle cells from preglomerular vessels. AB - Calcium entry via voltage-gated L-type channels is responsible for at least half of the increase in cytosolic calcium ([Ca(2+)](i)) in afferent arterioles following agonist stimulation. We sought the presence of capacitative calcium entry in fresh vascular smooth muscle cells (VSMC) derived from rat preglomerular vessels. [Ca(2+)](i) was measured using fura-2 ratiometric fluorescence. Vasopressin V1 receptor agonist (V1R) (10(-7) M) increased [Ca(2+)](i) by approximately 100 nM. A calcium channel blocker (CCB), nifedipine or verapamil (10(-7) M), inhibited the response by approximately 50%. V1R in the presence of CCB increased [Ca(2+)](i) from 106 to 176 nM, confirming that calcium mobilization and/or entry may occur independent of voltage-gated channels. In nominally Ca(2+)-free buffer, V1R increased [Ca(2+)](i) from 94 to 129 nM, denoting mobilization; addition of CaCl(2) (1 mM) further elevated [Ca(2+)](i) to 176 nM, indicating a secondary phase of Ca(2+) entry. Similar responses were obtained when CCB was present in calcium-free buffer or when EGTA was present. In nominally Ca(2+)-free medium, the sarcoplasmic reticulum Ca(2+)-ATPase inhibitors (SRCAI), thapsigargin and cyclopiazonic acid (CPA), increased [Ca(2+)](i) from 97 to 128 and 143 nM, respectively, and to 214 and 220 nM, respectively, when 1 mM extracellular Ca(2+) was added. In the presence of verapamil, the results with CPA acid were nearly identical. In Ca(2+)-free buffer, the stimulatory effect of V1R or SRCAI on the Ca(2+)/fura signal was quenched by the addition of Mn(2+) (1 mM), demonstrating divalent cation entry. These studies provide evidence for capacitative (store- operated) calcium entry in VSMC freshly isolated from rat preglomerular arterioles. PMID- 10516278 TI - NaPO(4) cotransport type III (PiT1) expression in human embryonic kidney cells and regulation by PTH. AB - The aim of the present study was to characterize the type(s) of NaPO(4) cotransporter expressed in the human renal cell line HEK-293 and its regulation by parathyroid hormone (PTH) in wild-type cells and in cells transfected by the PTH/PTH-related protein (PTHrP) receptor. The results showed that human embryonic kidney HEK-293 cells expressed NaPO(4) cotransporter type III (PiT1) mRNA and protein. In contrast, type I (NPT1) or II (NPT2) cotransporter mRNA were not expressed. Na(+)-dependent phosphate uptake followed a Michaelis-Menten model (apparent maximal transport rate and affinity constant: 23.32 +/- 0.69 nmol PO(4). mg protein(-1). 10 min(-1) and 0.147 +/- 0.014 mM KH(2)PO(4), respectively), was stimulated by phosphate deprivation (maximal increase 24.5 +/- 0.8%, P < 0.001, after 15 h of phosphate deprivation), and was inhibited by increasing pH (3.6 +/- 0.2-fold decrease at pH 8.5, P < 0.0001). It was inhibited in a time- and concentration-dependent fashion by PTH in HEK-293 cells stably transfected by PTH/PTHrP receptors but not in parental HEK-293 cells. Maximal inhibition of Na(+)-dependent phosphate transport was observed at 30 min after the addition of 72 nM PTH-(1-34) (31.5 +/- 2.4% inhibition, P < 0.01). PTH inhibition of phosphate transport was maintained in phosphate-deprived cells and reversed by both GF109203X (10(-6) M) or staurosporine (5.5 nM), two protein kinase C inhibitors. Na(+)-dependent phosphate uptake was also significantly inhibited by phorbol 12-myristate 13-acetate (20.9 +/- 3.9% inhibition, P < 0.001) but not by dibutyril-cAMP (10(-4) M) or forskolin (50 microM). The physiological role played by type III NaPO(4) cotransport expression in the overall renal regulation of phosphate homeostasis remains to be established. PMID- 10516279 TI - Nucleotides regulate NaCl transport in mIMCD-K2 cells via P2X and P2Y purinergic receptors. AB - Extracellular nucleotides regulate NaCl transport in some epithelia. However, the effects of nucleotide agonists on NaCl transport in the renal inner medullary collecting duct (IMCD) are not known. The objective of this study was to determine whether ATP and related nucleotides regulate NaCl transport across mouse IMCD cell line (mIMCD-K2) epithelial monolayers and, if so, via what purinergic receptor subtypes. ATP and UTP inhibited Na(+) absorption [measured via Na(+) short-circuit current (I(Na)(sc))] and stimulated Cl(-) secretion [measured via Cl(-) short-circuit current (I(Cl)(sc))]. Using selective P2 agonists, we report that P2X and P2Y purinoceptors regulate I(Na)(sc) and I(Cl)(sc). By RT-PCR, two P2X receptor channels (P2X(3), P2X(4)) and two P2Y G protein-coupled receptors (P2Y(1), P2Y(2)) were identified. Functional localization of P2 purinoceptors suggest that I(Cl)(sc) is stimulated by apical membrane-resident P2Y purinoceptors and P2X receptor channels, whereas I(Na)(sc) is inhibited by apical membrane-resident P2Y purinoceptors and P2X receptor channels. Together, we conclude that nucleotide agonists inhibit I(Na)(sc) across mIMCD-K2 monolayers through interactions with P2X and P2Y purinoceptors expressed on the apical plasma membrane, whereas extracellular nucleotides stimulate I(Cl)(sc) through interactions with P2X and P2Y purinoceptors expressed on the apical plasma membrane. PMID- 10516280 TI - Nitric oxide and renal nerve-mediated proximal tubular reabsorption in normotensive and hypertensive rats. AB - In Inactin-anesthetized Wistar rats with an intact renal innervation, intratubular nitro-L-arginine methyl ester (L-NAME, 10(-4) M) increased proximal fluid uptake (J(va), at 2.47 +/- 0.61 x 10(-4) mm(3). mm(-2). s(-1)) by 17% (P < 0.05), whereas coadministration with sodium nitroprusside (SNP, 10(-4) M) decreased J(va) by 18% (P < 0.01). Similar manipulation of NO generation was without effect in groups of Wistar rats subjected to acute renal denervation. Intratubular aminoguanidine (10(-4) M), a selective inducible nitric oxide synthase (NOS) blocker, had no effect on J(va) in intact kidneys of Wistar rats, but the neuronal NOS (nNOS) blocker, 7-nitroindazole (10(-4) M and 10(-6) M) increased J(va) by 19-23% (both P < 0.001). In stroke-prone spontaneously hypertensive rats (SHRSP), J(va) values in the innervated kidneys were lower (P < 0.05) than in the corresponding Wistar groups and were unchanged by intratubular L-NAME or L-NAME plus SNP. The tonic attenuation of proximal epithelial transport by NO was dependent on the renal sympathetic nerves and appeared to be generated by the nNOS isoform of the enzyme. This role of NO was not evident in the SHRSP. PMID- 10516281 TI - Mechanisms of HCO(-)(3) secretion in the rabbit connecting segment. AB - The connecting tubule (CNT) contains alpha-(H(+)-secreting) and beta-(HCO(-)(3) secreting) intercalated cells and is therefore likely to contribute to acid-base homeostasis. To characterize the mechanisms of HCO(-)(3) transport in the rabbit CNT, in which there is little definitive data presently available, we microdissected the segments from the superficial cortical labyrinth, perfused them in vitro, measured net HCO(-)(3) transport (J(HCO(-)(3))) by microcalorimetry, and examined the effects of several experimental maneuvers. Mean +/- SE basal J(HCO(-)(3)) was -3.4 +/- 0.1 pmol. min(-1). mm(-1) (net HCO( )(3) secretion), and transepithelial voltage was -13 +/- 1 mV (n = 47). Net HCO( )(3) secretion was markedly inhibited by removal of luminal Cl(-) or application of basolateral H(+)-ATPase inhibitors (bafilomycin or concanamycin), maneuvers that inhibit beta-intercalated cell function. Net HCO(-)(3) secretion was not affected by inhibitors of alpha-intercalated cell function (basolateral Cl(-) removal, basolateral DIDS, or luminal H(+)-ATPase inhibitors). Net HCO(-)(3) secretion was stimulated by isoproterenol and inhibited by acetazolamide. These data indicate that 1) CNTs secrete HCO(-)(3) via an apical DIDS-insensitive Cl( )/HCO(-)(3) exchanger, mediated by a basolateral bafilomycin- and concanamycin sensitive H(+)-ATPase; 2) inhibition of cytosolic carbonic anhydrase decreases HCO(-)(3) secretion; and 3) stimulation of beta-adrenergic receptors increases HCO(-)(3) secretion. The failure to influence net HCO(-)(3) transport by inhibiting alpha-intercalated cell apical H(+)-ATPases or basolateral Cl(-)/HCO( )(3) exchange suggests that the CNT has fewer functioning alpha-intercalated cells than the cortical collecting duct. These are the first studies to examine the rate and mechanisms of HCO(-)(3) secretion by the rabbit CNT; this is clearly an important segment in mediating acid-base homeostasis. PMID- 10516282 TI - Vasopressin stimulates sodium transport in A6 cells via a phosphatidylinositide 3 kinase-dependent pathway. AB - The enzyme phosphatidylinositide 3-kinase (PI3K) phosphorylates the D-3 position of the inositol ring of inositol phospholipids and produces 3-phosphorylated inositides. These novel second messengers are thought to mediate diverse cellular signaling functions. The fungal metabolite wortmannin covalently binds to PI3K and selectively inhibits its activity. The role of PI3K in basal and hormone stimulated transepithelial sodium transport was examined using this specific inhibitor. Wortmannin, 50 nM, did not affect basal, aldosterone-stimulated, or insulin-stimulated transport in A6 cells. Wortmannin completely inhibits vasopressin stimulation of transport in these cells. Vasopressin stimulates PI3K activity in A6 cells. Vasopressin stimulation of transport is also blocked by 5 microM LY-294002, a second inhibitor of PI3K. One-hour preincubation with wortmannin blocked vasopressin stimulation of protein kinase A activity in the cells. Sodium transport responses to exogenous cAMP and forskolin, which directly activates adenylate cyclase, were not affected by wortmannin. These results indicate that wortmannin inhibits vasopressin stimulation of Na(+) transport at a site proximal to activation of adenylate cyclase. The results suggest that PI3K may be involved in receptor activation by vasopressin. PMID- 10516283 TI - Regulation of the renal Na-HCO(3) cotransporter. XI. Signal transduction underlying CO(2) stimulation. AB - We have previously shown that CO(2) stimulation of the renal Na-HCO(3) cotransporter (NBC) activity is abrogated by general inhibitors of protein tyrosine kinases. The more selective inhibitor herbimycin also blocked this effect at concentrations known to preferentially inhibit Src family kinases (SFKs). We therefore examined a role for SFKs in CO(2)-stimulated NBC activity. To this end, we engineered OK cells to express the COOH-terminal Src kinase (Csk), a negative regulator of SFKs. CO(2) stimulated NBC activity normally in beta-galactosidase-expressing and untransfected control cells. In contrast, Csk expressing cells had normal baseline NBC activity that was not stimulated by CO(2). CO(2) stimulation increased both total SFK activity and specific tyrosine phosphorylation of Src. The specific MEK1/2 inhibitor PD-98059 completely inhibited the CO(2) stimulation of NBC activity as well as the accompanying phosphorylation and activation of ERK1/2. Our data suggest the involvement of both SFKs, probably Src, and the "classic" MAPK pathway in mediating CO(2) stimulated NBC activity in renal epithelial cells. PMID- 10516284 TI - Ksp-cadherin gene promoter. I. Characterization and renal epithelial cell specific activity. AB - Kidney-specific cadherin (Ksp-cadherin, cadherin 16) is a novel, kidney-specific member of the cadherin superfamily that is expressed exclusively in the basolateral membrane of renal tubular epithelial cells. To characterize the Ksp cadherin gene promoter, a lambda bacteriophage clone containing 3.7 kb of the proximal 5' flanking region of the mouse Ksp-cadherin gene was isolated. The transcription initiation site was mapped by RNase protection assays and 5' rapid amplification of cDNA ends, and a 709-bp intron was identified within the 5' untranslated region. The proximal 5' flanking region was "TATA-less" but contained other consensus promoter elements including an initiator (Inr), GC boxes, and a CAAT box. Potential binding sites were identified for transcription factors that are involved in tissue-specific gene expression including activator protein-2 (AP-2), hepatocyte nuclear factor-3 (HNF-3), basic helix-loop-helix (bHLH) proteins, CCAAT/enhancer-binding protein (C/EBP), and GATA factors. Transfection of luciferase reporter plasmids containing 2.6 kb of the 5' flanking region markedly increased luciferase activity in renal epithelial cells (MDCK and mIMCD-3) but not in mesenchymal cells (NIH 3T3 and MMR1). Deletion analysis identified an 82-bp region from -31 to -113 that was essential for promoter activity in transfected renal epithelial cells. Electrophoretic mobility-shift assays showed that mIMCD-3 cells contain nuclear proteins that bind to this region of the promoter. Mutational analysis showed that sequences within the HNF 3 consensus site and CAAT box were involved in protein binding and promoter activity. We conclude that the proximal 5' flanking region of the mouse Ksp cadherin gene contains an orientation-dependent promoter that is kidney epithelial cell specific. The region of the promoter from -113 to -31 is required for transcriptional activity and contains binding sites for nuclear proteins that are specifically expressed in renal epithelial cells. PMID- 10516285 TI - Ksp-cadherin gene promoter. II. Kidney-specific activity in transgenic mice. AB - Kidney-specific cadherin (Ksp-cadherin, cadherin 16) is a tissue-specific member of the cadherin superfamily that is expressed exclusively in the basolateral membrane of tubular epithelial cells in the kidney. To determine the basis for tissue-specific expression of Ksp-cadherin in vivo, we evaluated the activity of the promoter in transgenic mice. Transgenic mice containing 3.3 kb of the mouse Ksp-cadherin promoter and an Escherichia coli lacZ reporter gene were generated by pronuclear microinjection. Assays of beta-galactosidase enzyme activity showed that the transgene was expressed exclusively in the kidney in both adult and developing mice. Within the kidney, the transgene was expressed in a subset of renal tubular epithelial cells that endogenously expressed Ksp-cadherin and that were identified as collecting ducts by colabeling with Dolichos biflorus agglutinin. In the developing metanephros, expression of the transgene in the branching ureteric bud correlated with the developmental expression of Ksp cadherin. Identical patterns of expression were observed in multiple founder mice, indicating that kidney specificity was independent of transgene integration site. However, heterocellular expression was observed consistent with repeat induced gene silencing. We conclude that the Ksp-cadherin gene promoter directs kidney-specific expression in vivo. Regulatory elements that are sufficient to recapitulate the tissue- and differentiation-specific expression of Ksp-cadherin in the renal collecting duct are located within 3.3 kb upstream to the transcriptional start site. PMID- 10516286 TI - Cation and voltage dependence of rat kidney electrogenic Na(+)-HCO(-)(3) cotransporter, rkNBC, expressed in oocytes. AB - Recently, we reported the cloning and expression of the rat renal electrogenic Na(+)-HCO(-)(3) cotransporter (rkNBC) in Xenopus oocytes [M. F. Romero, P. Fong, U. V. Berger, M. A. Hediger, and W. F. Boron. Am. J. Physiol. 274 (Renal Physiol. 43): F425-F432, 1998]. Thus far, all NBC cDNAs are at least 95% homologous. Additionally, when expressed in oocytes the NBCs are 1) electrogenic, 2) Na(+) dependent, 3) HCO(-)(3) dependent, and 4) inhibited by stilbenes such as DIDS. The apparent HCO(-)(3):Na(+) coupling ratio ranges from 3:1 in kidney to 2:1 in pancreas and brain to 1:1 in the heart. This study investigates the cation and voltage dependence of rkNBC expressed in Xenopus oocytes to better understand NBC's apparent tissue-specific physiology. Using two-electrode voltage clamp, we studied the cation specificity, Na(+) dependence, and the current-voltage (I-V) profile of rkNBC. These experiments indicate that K(+) and choline do not stimulate HCO(-)(3)-sensitive currents via rkNBC, and Li(+) elicits only 3 +/- 2% of the total Na(+) current. The Na(+) dose response studies show that the apparent affinity of rkNBC for extracellular Na(+) ( approximately 30 mM [Na(+)](o)) is voltage and HCO(-)(3) independent, whereas the rkNBC I-V relationship is Na(+) dependent. At [Na(+)](o) v(max) (96 mM), the I-V response is approximately linear; both inward and outward Na(+)-HCO(-)(3) cotransport are observed. In contrast, only outward cotransport occurs at low [Na(+)](o) (<1 mM [Na(+)](o)). All rkNBC currents are inhibited by extracellular application of DIDS, independent of voltage and [Na(+)](o). Using ion-selective microelectrodes, we monitored intracellular pH and Na(+) activity. We then calculated intracellular [HCO(-)(3)] and, with the observed reversal potentials, calculated the stoichiometry of rkNBC over a range of [Na(+)](o) values from 10 to 96 mM at 10 and 33 mM [HCO(-)(3)](o). rkNBC stoichiometry is 2 HCO(-)(3):1 Na(+) over this entire Na(+) range at both HCO(-)(3) concentrations. Our results indicate that rkNBC is highly selective for Na(+), with transport direction and magnitude sensitive to [Na(+)](o) as well as membrane potential. Since the rkNBC protein alone in oocytes exhibits a stoichiometry of less than the 3 HCO(-)(3):1 Na(+) thought necessary for HCO(-)(3) reabsorption by the renal proximal tubule, a control mechanism or signal that alters its in vivo function is hypothesized. PMID- 10516287 TI - Hepatocyte growth factor promotes renal epithelial cell survival by dual mechanisms. AB - Hepatocyte growth factor (HGF) has been shown to protect renal epithelial cells against apoptosis. To define the mechanism by which HGF inhibits apoptosis, we investigated the effect of HGF on the phosphorylation and expression of the Bcl-2 family proteins. Using a human proximal tubular epithelial cell (HKC) line as a model, we demonstrated that constitutive expression of HGF conveyed marked resistance to apoptotic death induced by serum withdrawal. HGF induced rapid phosphorylation of Akt in HKC cells, which was immediately followed by phosphorylation and resultant inactivation of Bad, a pro-apoptotic member of the Bcl-2 family. Pretreatment of the HKC cells with 10 nM wortmannin completely abolished HGF-induced phosphorylation of Akt and Bad, suggesting that this pathway is dependent on phosphoinositide (PI) 3-kinase. Overexpression of Bad increased apoptotic death in wild-type HKC cells but not in HGF-producing H4 cells. Immunoblotting confirmed that the Bad protein over-expressed in H4 cells was fully phosphorylated at both Ser(112) and Ser(136) sites. Prolonged incubation of HKC cells with HGF also dramatically induced expression of Bcl-xL, an anti-apoptotic member of the Bcl-2 family. These results suggest that the anti apoptotic effect of HGF in renal epithelial cells is mediated by dual mechanisms involving two distinct Bcl-2 family proteins. HGF triggers Bad phosphorylation via the PI 3-kinase/Akt pathway, thereby inactivating this pro-apoptotic protein, while simultaneously inducing expression of anti-apoptotic Bcl-xL. PMID- 10516288 TI - JG cell expression and partial regulation of a human renin genomic transgene driven by a minimal renin promoter. AB - In the kidney, renin gene expression is exquisitely localized to the juxtaglomerular (JG) cells lining the afferent arteriole, having the capacity to regulate renin synthesis in response to a variety of physiological cues. We investigated human renin gene expression in transgenic mice containing a genomic construct driven by 149 bp of its proximal promoter to elucidate whether this was sufficient to confer JG-specific expression. Whereas human renin mRNA was permissively expressed in most tissues, the transgene was expressed mainly in JG cells in the kidney. Active human renin and human prorenin were found in the systemic circulation at levels consistent with previous transgenic models. Remarkably, two lines displayed an appropriate upregulation of transgene mRNA in response to angiotensin-converting enzyme inhibition, and two lines exhibited a downregulation of transgene mRNA in response to subpressor and pressor doses of ANG II. Our results suggest that 149 bp of the human renin proximal promoter, in a context of a genomic construct, are sufficient to confer human renin expression in renal JG cells and at least some aspects of appropriate regulation. PMID- 10516289 TI - Defective processing and expression of thiazide-sensitive Na-Cl cotransporter as a cause of Gitelman's syndrome. AB - Gitelman's syndrome is an autosomal recessive disorder of salt wasting and hypokalemia caused by mutations in the thiazide-sensitive Na-Cl cotransporter. To investigate the pathogenesis of Gitelman's syndrome, eight disease mutations were introduced into the mouse thiazide-sensitive Na-Cl cotransporter and studied by functional expression in Xenopus oocytes. Sodium uptake into oocytes that expressed the wild-type clone was more than sevenfold greater than uptake into control oocytes. Uptake into oocytes that expressed the mutated transporters was not different from control. Hydrochlorothiazide reduced Na uptake by oocytes expressing the wild-type gene to control values but had no effect on oocytes expressing the mutant clones. Western blots of oocytes injected with the wild type clone showed bands representing glycosylated (125 kDa) and unglycosylated (110 kDa) forms of the transport protein. Immunoblot of oocytes expressing the mutated clones showed only the unglycosylated protein, indicating that protein processing was disrupted. Immunocytochemistry with an antibody against the transport protein showed intense membrane staining of oocytes expressing the wild type protein. Membrane staining was completely absent from oocytes expressing mNCC(R948X); instead, diffuse cytoplasmic staining was evident. In summary, the results show that several mutations that cause Gitelman's syndrome are nonfunctional because the mutant thiazide-sensitive Na-Cl cotransporter is not processed normally, probably activating the "quality control" mechanism of the endoplasmic reticulum. PMID- 10516290 TI - Evolution of gene expression patterns in a model of branching morphogenesis. AB - Branching morphogenesis of the ureteric bud in response to unknown signals from the metanephric mesenchyme gives rise to the urinary collecting system and, via inductive signals from the ureteric bud, to recruitment of nephrons from undifferentiated mesenchyme. An established cell culture model for this process employs cells of ureteric bud origin (UB) cultured in extracellular matrix and stimulated with conditioned media (BSN-CM) from a metanephric mesenchymal cell line (H. Sakurai, E. J. Barros, T. Tsukamoto, J. Barasch, and S. K. Nigam. Proc. Natl. Acad. Sci. USA 94: 6279-6284, 1997.). In the presence of BSN-CM, the UB cells form branching tubular structures reminiscent of the branching ureteric bud. The pattern of gene regulation in this model of branching morphogenesis of the kidney collecting system was investigated using high-density cDNA arrays. Software and analytical methods were developed for the quantification and clustering of genes. With the use of a computational method termed "vector analysis," genes were clustered according to the direction and magnitude of differential expression in n-dimensional log-space. Changes in gene expression in response to the BSN-CM consisted primarily of differential expression of transcription factors with previously described roles in morphogenesis, downregulation of pro-apoptotic genes accompanied by upregulation of anti apoptotic genes, and upregulation of a small group of secreted products including growth factors, cytokines, and extracellular proteinases. Changes in expression are discussed in the context of a general model for epithelial branching morphogenesis. In addition, the cDNA arrays were used to survey expression of epithelial markers and secreted factors in UB and BSN cells, confirming the largely epithelial character of the former and largely mesenchymal character of the later. Specific morphologies (cellular processes, branching multicellular cords, etc.) were shown to correlate with the expression of different, but overlapping, genomic subsets, suggesting differences in morphogenetic mechanisms at these various steps in the evolution of branching tubules. PMID- 10516291 TI - Time course and permeation of synaptic AMPA receptors in cochlear nuclear neurons correlate with input. AB - AMPA receptors mediate rapid glutamatergic synaptic transmission. In the mammalian cochlear nuclei, neurons receive excitatory input from either auditory nerve fibers, parallel fibers, or both fiber systems. The functional correlates of differences in the source of input were examined by recording AMPA receptor mediated, miniature EPSCs (mEPSCs) in whole-cell voltage-clamp mode from identified neurons. Bushy, octopus, and T-stellate cells of the ventral cochlear nucleus (VCN) and tuberculoventral cells of the dorsal cochlear nucleus (DCN) receive most of their excitatory input from the auditory nerve; fusiform cells receive excitatory inputs from both the auditory nerve and parallel fibers; cartwheel cells receive excitatory input from parallel fibers alone. mEPSCs from bushy, octopus, T-stellate, and tuberculoventral cells had significantly faster decay time constants (0.35-0.40 msec) than did those from fusiform and cartwheel cells (1.32-1.79 msec). Some fusiform cells had two populations of mEPSCs with distinct time courses. mEPSCs in cells with auditory nerve input alone were inhibited by philanthotoxin, a blocker of calcium-permeable AMPA receptors, whereas mEPSCs in cells with parallel fiber input were not. Thus AMPA receptors postsynaptic to the auditory nerve differ from those postsynaptic to parallel fibers both in channel-gating kinetics and in their permeability to calcium. These results confirm the conclusion that synaptic AMPA receptors are specialized according to the source of input (Hunter et al., 1993; Rubio and Wenthold, 1997; Wang et al., 1998). PMID- 10516292 TI - A scorpion alpha-like toxin that is active on insects and mammals reveals an unexpected specificity and distribution of sodium channel subtypes in rat brain neurons. AB - Several scorpion toxins have been shown to exert their neurotoxic effects by a direct interaction with voltage-dependent sodium channels. Both classical scorpion alpha-toxins such as Lqh II from Leiurus quiquestratus hebraeus and alpha-like toxins as toxin III from the same scorpion (Lqh III) competitively interact for binding on receptor site 3 of insect sodium channels. Conversely, Lqh III, which is highly toxic in mammalian brain, reveals no specific binding to sodium channels of rat brain synaptosomes and displaces the binding of Lqh II only at high concentration. The contrast between the low-affinity interaction and the high toxicity of Lqh III indicates that Lqh III binding sites distinct from those present in synaptosomes must exist in the brain. In agreement, electrophysiological experiments performed on acute rat hippocampal slices revealed that Lqh III strongly affects the inactivation of voltage-gated sodium channels recorded either in current or voltage clamp, whereas Lqh II had weak, or no, effects. In contrast, Lqh III had no effect on cultured embryonic chick central neurons and on sodium channels from rat brain IIA and beta1 subunits reconstituted in Xenopus oocytes, whereas sea anemone toxin ATXII and Lqh II were very active. These data indicate that the alpha-like toxin Lqh III displays a surprising subtype specificity, reveals the presence of a new, distinct sodium channel insensitive to Lqh II, and highlights the differences in distribution of channel expression in the CNS. This toxin may constitute a valuable tool for the investigation of mammalian brain function. PMID- 10516293 TI - Developmental expression of an amn(+) transgene rescues the mutant memory defect of amnesiac adults. AB - The Drosophila memory gene amnesiac (amn) has been proposed to encode a neuropeptide protein, which includes regions homologous to vertebrate pituitary adenylyl cyclase-activating peptide (PACAP; Feany and Quinn, 1995). Definitive experiments to link this gene to memory formation, however, have not yet been accomplished (Kandel and Abel, 1995). The experiments described here demonstrate that the putative amn transcript is involved in adult memory formation. With the use of a UAS-amn(+) transgene, we show complete rescue of memory defects in amn(28A), a mutant allele caused by the insertion of a GAL4 enhancer trap transposon (Moore et al., 1998). Study of the amn(28A) reporter reveals widespread expression in the adult brain but also enriched expression in the embryonic and larval nervous systems. To begin addressing the temporal requirement of amn in memory, we asked whether the memory defects could be rescued by restricting transgenic expression to the adult stage. A heat-shock regimen shown previously to rescue fully the amn ethanol sensitivity defect (Moore et al., 1998) failed to rescue the memory defect. These results, coupled with previous genetic and anatomical studies, suggest that adult memory formation and ethanol sensitivity have different temporal and spatial requirements for amn. PMID- 10516294 TI - Role of cell cycle regulatory proteins in cerebellar granule neuron apoptosis. AB - Cerebellar granule neurons (CGNs) undergo apoptosis when deprived of depolarizing concentrations of KCl, but the underlying molecular mechanisms are not yet clear. Although caspases have been postulated to be involved in CGN cell death, inhibitors of caspases failed to prevent apoptosis under our culture conditions, suggesting an involvement of other molecules and pathways. We find that inhibitors of cyclin-dependent kinases--flavopiridol, olomoucine, and roscovitine -protect CGNs from KCl withdrawal-induced apoptosis, suggesting that cell cycle components play a significant role in the death of these neurons. Analysis of the different cell cycle regulatory elements in this model revealed that apoptosis is preceded by an increase in the level of cyclin E protein, with elevated nuclear levels of cyclin D1 and with enhanced activity of the cyclin D1- and E- associated kinases. In addition, there was a significant decrease in the level of the cyclin-dependent kinase (cdk) inhibitor p27. In agreement with these changes, analysis of a major substrate of cyclin-activated cdks, retinoblastoma protein (Rb), showed an increase in the level of phosphorylated forms within 1 hr of KCl withdrawal. Moreover, the overall levels of Rb protein were significantly reduced within 6-12 hr of KCl withdrawal and did so by a caspase-independent mechanism. All of these responses were blocked by cdk inhibitors. These findings indicate that cdks act at an early step in the pathway by which KCl withdrawal induces apoptotic death of cerebellar granule cells and suggest that additional elements of the cell cycle machinery participate in this mechanism. PMID- 10516295 TI - Generation and analysis of GluR5(Q636R) kainate receptor mutant mice. AB - The physiological significance of RNA editing of transcripts that code for kainate-preferring glutamate receptor subunits is unknown, despite the fact that the functional consequences of this molecular modification have been well characterized in cloned receptor subunits. RNA editing of the codon that encodes the glutamine/arginine (Q/R) site in the second membrane domain (MD2) of glutamate receptor 5 (GluR5) and GluR6 kainate receptor subunits produces receptors with reduced calcium permeabilities and single-channel conductances. Approximately 50% of the GluR5 subunit transcripts from adult rat brain are edited at the Q/R site in MD2. To address the role of glutamate receptor mRNA editing in the brain, we have made two strains of mice with mutations at amino acid 636, the Q/R-editing site in GluR5, using embryonic stem cell-mediated transgenesis. GluR5(RloxP/RloxP) mice encode an arginine at the Q/R site of the GluR5 subunit, whereas GluR5(wt(loxP)/wt(loxP)) mice encode a glutamine at this site, similar to wild-type mice. Mutant animals do not exhibit developmental abnormalities, nor do they show deficits in the behavioral paradigms tested in this study. Kainate receptor current densities were reduced by a factor of six in acutely isolated sensory neurons of dorsal root ganglia from GluR5(RloxP/RloxP) mice compared with neurons from wild-type mice. However, the editing mutant mice did not exhibit altered responses to thermal and chemical pain stimuli. Our investigations with the GluR5-editing mutant mice have therefore defined a set of physiological processes in which editing of the GluR5 subunit is unlikely to play an important role. PMID- 10516296 TI - Cannabinoids enhance NMDA-elicited Ca2+ signals in cerebellar granule neurons in culture. AB - A physiological role for cannabinoids in the CNS is indicated by the presence of endogenous cannabinoids and cannabinoid receptors. However, the cellular mechanisms of cannabinoid actions in the CNS have yet to be fully defined. In the current study, we identified a novel action of cannabinoids to enhance intracellular Ca2+ responses in CNS neurons. Acute application of the cannabinoid receptor agonists R(+)-methanandamide, R(+)-WIN, and HU-210 (1-50 nM) dose dependently enhanced the peak amplitude of the Ca2+ response elicited by stimulation of the NMDA subtype of glutamate receptors (NMDARs) in cerebellar granule neurons. The cannabinoid effect was blocked by the cannabinoid receptor antagonist SR141716A and the Gi/Go protein inhibitor pertussis toxin but was not mimicked by the inactive cannabinoid analog S(-)-WIN, indicating the involvement of cannabinoid receptors. In current-clamp studies neither R(+)-WIN nor R(+) methanandamide altered the membrane response to NMDA or passive membrane properties of granule neurons, suggesting that NMDARs are not the primary sites of cannabinoid action. Additional Ca2+ imaging studies showed that cannabinoid enhancement of the Ca2+ signal to NMDA did not involve N-, P-, or L-type Ca2+ channels but was dependent on Ca2+ release from intracellular stores. Moreover, the phospholipase C inhibitor U-73122 and the inositol 1,4,5-trisphosphate (IP3) receptor antagonist xestospongin C blocked the cannabinoid effect, suggesting that the cannabinoid enhancement of NMDA-evoked Ca2+ signals results from enhanced release from IP3-sensitive Ca2+ stores. These data suggest that the CNS cannabinoid system could serve a critical modulatory role in CNS neurons through the regulation of intracellular Ca2+ signaling. PMID- 10516297 TI - Comparing astrocytic cell lines that are inhibitory or permissive for axon growth: the major axon-inhibitory proteoglycan is NG2. AB - Astrocytes, oligodendrocytes, and oligodendrocyte/type 2 astrocyte progenitors (O2A cells) can all produce molecules that inhibit axon regeneration. We have shown previously that inhibition of axon growth by astrocytes involves proteoglycans. To identify inhibitory mechanisms, we created astrocyte cell lines that are permissive or nonpermissive and showed that nonpermissive cells produce inhibitory chondroitin sulfate proteoglycans (CS-PGs). We have now tested these cell lines for the production and inhibitory function of known large CS-PGs. The most inhibitory line, Neu7, produces three CS-PGs in much greater amounts than the other cell lines: NG2, versican, and the CS-56 antigen. The contribution of NG2 to inhibition by the cells was tested using a function-blocking antibody. This allowed increased growth of dorsal root ganglion (DRG) axons over Neu7 cells and matrix and greatly increased the proportion of cortical axons able to cross from permissive A7 cells onto inhibitory Neu7 cells; CS-56 antibody had a similar effect. Inhibitory fractions of conditioned medium contained NG2 coupled to CS glycosaminoglycan chains, whereas noninhibitory fractions contained NG2 without CS chains. Enzyme preparations that facilitated axon growth in Neu7 cultures were shown to either degrade the NG2 core protein or remove CS chains. Versican is present as patches on Neu7 monolayers, but DRG axons do not avoid these patches. Therefore, NG2 appears to be the major axon-inhibitory factor made by Neu7 astrocytes. In the CNS, NG2 is expressed by O2A cells, which react rapidly after injury to produce a dense NG2-rich network, and by some reactive astrocytes. Our results suggest that NG2 may be a major obstacle to axon regeneration. PMID- 10516298 TI - Dendritic calcium spike initiation and repolarization are controlled by distinct potassium channel subtypes in CA1 pyramidal neurons. AB - In CA1 pyramidal neurons of the hippocampus, calcium-dependent spikes occur in vivo during specific behavioral states and may be enhanced during epileptiform activity. However, the mechanisms that control calcium spike initiation and repolarization are poorly understood. Using dendritic and somatic patch-pipette recordings, we show that calcium spikes are initiated in the apical dendrites of CA1 pyramidal neurons and drive bursts of sodium-dependent action potentials at the soma. Initiation of calcium spikes at the soma was suppressed in part by potassium channels activated by sodium-dependent action potentials. Low threshold, putative D-type potassium channels [blocked by 100 microM 4 aminopyridine (4-AP) and 0.5-1 microM alpha-dendrotoxin (alpha-DTX)] played a prominent role in setting a high threshold for somatic calcium spikes, thus restricting initiation to the dendrites. DTX- and 4-AP-sensitive channels were activated during sodium-dependent action potentials and mediated a large component of their afterhyperpolarization. Once initiated, repetitive firing of calcium spikes was limited by activation of putative BK-type calcium-activated potassium channels (blocked by 250 microM tetraethylammonium chloride, 70 nM charybdotoxin, or 100 nM iberiotoxin). Thus, the concerted action of calcium- and voltage-activated potassium channels serves to focus spatially and temporally the membrane depolarization and calcium influx generated by calcium spikes during strong, synchronous network excitation. PMID- 10516299 TI - Modulation of the light response by cAMP in Drosophila photoreceptors. AB - Phototransduction in Drosophila is mediated by a G-protein-coupled phospholipase C transduction cascade in which each absorbed photon generates a discrete electrical event, the quantum bump. In whole-cell voltage-clamp recordings, cAMP, as well as its nonhydrolyzable and membrane-permeant analogs 8-bromo-cAMP (8-Br cAMP) and dibutyryl-cAMP, slowed down the macroscopic light response by increasing quantum bump latency, without changes in bump amplitude or duration. In contrast, cGMP or 8-Br-cGMP had no effect on light response amplitude or kinetics. None of the cyclic nucleotides activated any channels in the plasma membrane. The effects of cAMP were mimicked by application of the non-specific phosphodiesterase inhibitor IBMX and the adenylyl cyclase activator forskolin; zaprinast, a specific cGMP-phosphodiesterase inhibitor, was ineffective. Bump latency was also increased by targeted expression of either an activated G(s) alpha subunit, which increased endogenous adenylyl cyclase activity, or an activated catalytic protein kinase A (PKA) subunit. The action of IBMX was blocked by pretreatment with the PKA inhibitor H-89. The effects of cAMP were abolished in mutants of the ninaC gene, suggesting this nonconventional myosin as a possible target for PKA-mediated phosphorylation. Dopamine (10 microM) and octopamine (100 microM) mimicked the effects of cAMP. These results indicate the existence of a G-protein-coupled adenylyl cyclase pathway in Drosophila photoreceptors, which modulates the phospholipase C-based phototransduction cascade. PMID- 10516300 TI - Presynaptic inhibition of primary olfactory afferents mediated by different mechanisms in lobster and turtle. AB - Presynaptic regulation of transmission at the first olfactory synapse was investigated by selectively imaging axon terminals of receptor neurons in the lobster olfactory lobe and turtle olfactory bulb. In both species, action potential propagation into axon terminals after olfactory nerve stimulation was measured using voltage-sensitive dyes. In addition, in the turtle, calcium influx into terminals was measured by selectively labeling receptor neurons with dextran conjugated calcium indicator dyes. In the lobster, application of the inhibitory transmitters GABA or histamine suppressed action potentials in the terminals. The suppression was blocked by picrotoxin and cimetidine, respective antagonists to lobster GABA and histamine receptors. These results suggest that previously characterized GABA and histaminergic interneurons regulate olfactory input by suppressing action potential propagation into axon terminals of olfactory afferents. In contrast, in the turtle olfactory bulb, neither GABA nor dopamine had any effect on receptor cell action potentials as measured with voltage sensitive dyes. However, calcium influx into axon terminals was reduced by the GABA(B) agonist baclofen and the dopamine D(2) agonist quinpirole, and paired pulse suppression of calcium influx was reduced by the GABA(B) antagonist saclofen. These results indicate that in the turtle, GABA and dopamine mediate presynaptic inhibition not by affecting action potentials directly, as in the lobster, but by reducing calcium influx via GABA(B) and dopamine D(2) receptors. Thus, although mediated by different cellular mechanisms, presynaptic regulation of olfactory input to the CNS, via dual synaptic pathways, is a feature common to vertebrates and invertebrates. This inhibition may be important in the processing of olfactory information. PMID- 10516301 TI - Identification of a cis-acting dendritic targeting element in MAP2 mRNAs. AB - In neurons, a limited number of mRNAs have been identified in dendritic processes, whereas other transcripts are restricted to the cell soma. Here we have investigated the molecular mechanisms underlying extrasomatic localization of mRNAs encoding microtubule-associated protein 2 (MAP2) in primary neuronal cultures. Vectors expressing recombinant mRNAs were introduced into hippocampal and sympathetic neurons using DNA transfection and microinjection protocols, respectively. Chimeric mRNAs containing the entire 3' untranslated region of MAP2 transcripts fused to a nondendritic reporter mRNA are detected in dendrites. In contrast, RNAs containing MAP2 coding and 5' untranslated regions or tubulin sequences are restricted to the cell soma. Moreover, 640 nucleotides from the MAP2 3' untranslated region (UTR) are both sufficient and essential for extrasomatic localization of chimeric mRNAs in hippocampal and sympathetic neurons. Thus, a cis-acting dendritic targeting element that is effective in two distinct neuronal cell types is contained in the 3' UTR of MAP2 transcripts. The observation of RNA granules in dendrites implies that extrasomatic transcripts seem to assemble into multimolecular complexes that may function as transport units. PMID- 10516302 TI - Domains responsible for constitutive and Ca(2+)-dependent interactions between calmodulin and small conductance Ca(2+)-activated potassium channels. AB - Small conductance Ca(2+)-activated potassium channels (SK channels) are coassembled complexes of pore-forming SK alpha subunits and calmodulin. We proposed a model for channel activation in which Ca2+ binding to calmodulin induces conformational rearrangements in calmodulin and the alpha subunits that result in channel gating. We now report fluorescence measurements that indicate conformational changes in the alpha subunit after calmodulin binding and Ca2+ binding to the alpha subunit-calmodulin complex. Two-hybrid experiments showed that the Ca(2+)-independent interaction of calmodulin with the alpha subunits requires only the C-terminal domain of calmodulin and is mediated by two noncontiguous subregions; the ability of the E-F hands to bind Ca2+ is not required. Although SK alpha subunits lack a consensus calmodulin-binding motif, mutagenesis experiments identified two positively charged residues required for Ca(2+)-independent interactions with calmodulin. Electrophysiological recordings of SK2 channels in membrane patches from oocytes coexpressing mutant calmodulins revealed that channel gating is mediated by Ca2+ binding to the first and second E-F hand motifs in the N-terminal domain of calmodulin. Taken together, the results support a calmodulin- and Ca(2+)-calmodulin-dependent conformational change in the channel alpha subunits, in which different domains of calmodulin are responsible for Ca(2+)-dependent and Ca(2+)-independent interactions. In addition, calmodulin is associated with each alpha subunit and must bind at least one Ca2+ ion for channel gating. Based on these results, a state model for Ca2+ gating was developed that simulates alterations in SK channel Ca2+ sensitivity and cooperativity associated with mutations in CaM. PMID- 10516303 TI - The weaver mouse gain-of-function phenotype of dopaminergic midbrain neurons is determined by coactivation of wvGirk2 and K-ATP channels. AB - The phenotype of substantia nigra (SN) neurons in homozygous weaver (wv/wv) mice was studied by combining patch-clamp and single-cell RT-multiplex PCR techniques in midbrain slices of 14-d-old mice. In contrast to GABAergic SN neurons, which were unaffected in homozygous weaver mice (wv/wv), dopaminergic SN neurons possessed a dramatically altered phenotype with a depolarized membrane potential and complete loss of spontaneous pacemaker activity. The gain-of-function phenotype was mediated by a large, nonselective membrane conductance exclusively present in (wv/wv) dopaminergic SN neurons. This constitutively activated conductance displayed a sensitivity to external QX-314 (IC(50) = 10.6 microM) very similar to that of heterologously expressed wvGirk2 channels and was not further activated by G-protein stimulation. Single-cell Girk1-4 expression profiling suggested that homomeric Girk2 channels were present in most dopaminergic SN neurons, whereas Girk2 was always coexpressed with other Girk family members in GABAergic SN neurons. Surprisingly, acute QX-314 inhibition of wvGirk2 channels did not induce wild-type-like pacemaker activity but instead caused membrane hyperpolarization. Additional application of a blocker of ATP sensitive potassium channels (100 microM tolbutamide) induced wild-type-like pacemaker activity. We conclude that the gain-of-function weaver phenotype of dopaminergic substantia nigra neurons is mediated by coactivation of wvGirk2 and SUR1/Kir6. 2-mediated ATP-sensitive K(+) channels. We also show that in contrast to wild-type neurons, all (wv/wv) dopaminergic SN neurons expressed calbindin, a calcium-binding protein that marks dopaminergic SN neurons resistant to neurodegeneration. The identification of two ion channels that in concert determine the weaver phenotype of surviving calbindin-positive dopaminergic SN neurons will help to understand the molecular mechanisms of selective neurodegeneration of dopaminergic SN neurons in the weaver mouse and might be important in Parkinson's disease. PMID- 10516304 TI - Neuroprotective effect of high glucose against NMDA, free radical, and oxygen glucose deprivation through enhanced mitochondrial potentials. AB - Cultured cortical neurons maintained in 25 mM glucose underwent a widespread neuronal death after exposure to NMDA, AMPA, and kainate. Among these, NMDA toxicity was substantially reduced in neurons maintained in 100 mM glucose. NMDA induced increase in [Ca(2+)](i) and reactive oxygen species was attenuated in neurons maintained in high glucose that revealed increased mitochondrial membrane and redox potentials as determined using rhodamine 123 and 3-(4, 5 dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide. p-trifluoromethoxy phenylhydrazone, KCN, and rotenone, the selective inhibitors of mitochondrial potential, abrogated neuroprotective effect of high glucose against NMDA. The neuroprotective action of high glucose was extended against oxygen or combined oxygen-glucose deprivation. The present study provides evidence that prolonged exposure of cortical cells to high glucose attenuates NMDA- and free radical mediated neuronal death via enhanced mitochondrial function. PMID- 10516305 TI - Lack of the p50 subunit of nuclear factor-kappaB increases the vulnerability of hippocampal neurons to excitotoxic injury. AB - Nuclear factor-kappaB (NF-kappaB) is activated in brain cells after various insults, including cerebral ischemia and epileptic seizures. Although cell culture studies have suggested that the activation of NF-kappaB can prevent neuronal apoptosis, the role of this transcription factor in neuronal injury in vivo is unclear, and the specific kappaB subunits involved are unknown. We now report that mice lacking the p50 subunit of NF-kappaB exhibit increased damage to hippocampal pyramidal neurons after administration of the excitotoxin kainate. Gel-shift analyses showed that p50 is required for the majority of kappaB DNA binding activity in hippocampus. Intraventricular administration of kappaB decoy DNA before kainate administration in wild-type mice resulted in an enhancement of damage to hippocampal pyramidal neurons, indicating that reduced NF-kappaB activity was sufficient to account for the enhanced excitotoxic neuronal injury in p50(-/-) mice. Cultured hippocampal neurons from p50(-/-) mice exhibited enhanced elevations of intracellular calcium levels and increased levels of oxidative stress after exposure to glutamate and were more vulnerable to excitotoxicity than were neurons from p50(+/+) and p50(+/-) mice. Collectively, our data demonstrate an important role for the p50 subunit of NF-kappaB in protecting neurons against excitotoxic cell death. PMID- 10516306 TI - Evidence of presynaptic location and function of the prion protein. AB - The prion protein (PrP(C)) is a copper-binding protein of unknown function that plays an important role in the etiology of transmissible spongiform encephalopathies. Using morphological techniques and synaptosomal fractionation methods, we show that PrP(C) is predominantly localized to synaptic membranes. Atomic absorption spectroscopy was used to identify PrP(C)-related changes in the synaptosomal copper concentration in transgenic mouse lines. The synaptic transmission in the presence of H(2)O(2), which is known to be decomposed to highly reactive hydroxyl radicals in the presence of iron or copper and to alter synaptic activity, was studied in these animals. The response of synaptic activity to H(2)O(2) was found to correlate with the amount of PrP(C) expression in the presynaptic neuron in cerebellar slice preparations from wild-type, Prnp(0/0), and PrP gene-reconstituted transgenic mice. Thus, our data gives strong evidence for the predominantly synaptic location of PrP(C), its involvement in the regulation of the presynaptic copper concentration, and synaptic activity in defined conditions. PMID- 10516307 TI - Protofibrillar intermediates of amyloid beta-protein induce acute electrophysiological changes and progressive neurotoxicity in cortical neurons. AB - Alzheimer's disease (AD) is a progressive neurodegenerative disorder that is thought to be caused in part by the age-related accumulation of amyloid beta protein (Abeta). The presence of neuritic plaques containing abundant Abeta derived amyloid fibrils in AD brain tissue supports the concept that fibril accumulation per se underlies neuronal dysfunction in AD. Recent observations have begun to challenge this assumption by suggesting that earlier Abeta assemblies formed during the process of fibrillogenesis may also play a role in AD pathogenesis. Here, we present the novel finding that protofibrils (PF), metastable intermediates in amyloid fibril formation, can alter the electrical activity of neurons and cause neuronal loss. Both low molecular weight Abeta (LMW Abeta) and PF reproducibly induced toxicity in mixed brain cultures in a time- and concentration-dependent manner. No increase in fibril formation during the course of the experiments was observed by either Congo red binding or electron microscopy, suggesting that the neurotoxicity of LMW Abeta and PF cannot be explained by conversion to fibrils. Importantly, protofibrils, but not LMW Abeta, produced a rapid increase in EPSPs, action potentials, and membrane depolarizations. These data suggest that PF have inherent biological activity similar to that of mature fibrils. Our results raise the possibility that the preclinical and early clinical progression of AD is driven in part by the accumulation of specific Abeta assembly intermediates formed during the process of fibrillogenesis. PMID- 10516308 TI - A role for the Eph ligand ephrin-A3 in entorhino-hippocampal axon targeting. AB - Neurons of layers II and III of the entorhinal cortex constitute the major afferent connection of the hippocampus. The molecular mechanisms that target the entorhinal axons to specific layers in the hippocampus are not known. EphA5, a member of the Eph receptor family, which has been shown to play critical roles in axon guidance, is expressed in the entorhinal cortex, the origin of the perforant pathway. In addition, ligands that interact with EphA5 are expressed in distinct hippocampal regions during development of the entorhino-hippocampal projection. Of these ligands, ephrin-A3 mRNA is localized both in the granular cell layer of the dentate gyrus and in the pyramidal cell layer of the cornu ammonis, whereas ephrin-A5 mRNA is only expressed in the pyramidal cell layer of the cornu ammonis. In the dentate gyrus, the ligand protein is not present in the termination zone of the entorhinal efferents (the outer molecular layer of the dentate gyrus) but is concentrated in the inner molecular layer into which entorhinal efferents do not grow. We used outgrowth and stripe assays to test the effects of ephrin-A3 and ephrin-A5 on the outgrowth behavior of entorhinal axons. This functional analysis revealed that entorhinal neurites were repelled by ephrin-A3 but not by ephrin-A5. These observations suggest that ephrin-A3 plays an important role in the layer-specific termination of the perforant pathway and that this ligand may interact with the EphA5 receptor to restrict entorhinal axon terminals in the outer molecular layer of the dentate gyrus. PMID- 10516309 TI - Reorganization and movement of microtubules in axonal growth cones and developing interstitial branches. AB - Local changes in microtubule organization and distribution are required for the axon to grow and navigate appropriately; however, little is known about how microtubules (MTs) reorganize during directed axon outgrowth. We have used time lapse digital imaging of developing cortical neurons microinjected with fluorescently labeled tubulin to follow the movements of individual MTs in two regions of the axon where directed growth occurs: the terminal growth cone and the developing interstitial branch. In both regions, transitions from quiescent to growth states were accompanied by reorganization of MTs from looped or bundled arrays to dispersed arrays and fragmentation of long MTs into short MTs. We also found that long-term redistribution of MTs accompanied the withdrawal of some axonal processes and the growth and stabilization of others. Individual MTs moved independently in both anterograde and retrograde directions to explore developing processes. Their velocities were inversely proportional to their lengths. Our results demonstrate directly that MTs move within axonal growth cones and developing interstitial branches. Our findings also provide the first direct evidence that similar reorganization and movement of individual MTs occur in the two regions of the axon where directed outgrowth occurs. These results suggest a model whereby short exploratory MTs could direct axonal growth cones and interstitial branches toward appropriate locations. PMID- 10516310 TI - Postsynaptic calcium/calmodulin-dependent protein kinase II is required to limit elaboration of presynaptic and postsynaptic neuronal arbors. AB - Neuronal dendritic and axonal arbors grow to a characteristic size and then stabilize their structures. Activity-dependent stop-growing signals may limit neuronal process elaboration. We tested whether endogenous calcium/calmodulin dependent protein kinase II (CaMKII) activity in postsynaptic optic tectal cells is required to restrict the elaboration of neuronal processes in the Xenopus tadpole retinotectal projection. Optic tectal cells were infected with vaccinia viruses that express CaMKII-specific inhibitory peptides. In vivo time-lapse imaging revealed that expression of CaMKII inhibitors blocked the growth restriction that normally occurs during maturation of tectal cell dendritic arbors. Postsynaptic CaMKII inhibition also increased the growth of presynaptic retinotectal axon arbors. The results indicate that endogenous postsynaptic CaMKII activity is required to limit the growth of presynaptic and postsynaptic arbor structures in vivo. PMID- 10516312 TI - Neonatal partial denervation results in nodal but not terminal sprouting and a decrease in efficacy of remaining neuromuscular junctions in rat soleus muscle. AB - Mature motoneurons respond to partial denervation of their target muscle by sprouting to reinnervate denervated fibers, thus maintaining muscle strength in the face of motoneuronal loss caused by injury or disease. Neonatal motoneurons, however, do not expand to innervate more muscle fibers. The present work seeks to understand this developmental change in motoneuron response to partial denervation. It has been suggested that neonatal motor units cannot increase in size because they are already at their maximum size (approximately five times larger than in adulthood). We ruled out this explanation by showing that after partial denervation on postnatal day 14 (P14), when motor units have decreased to their adult size, motoneurons still did not sprout to reinnervate as many fibers as in adulthood. Instead, we found evidence supporting an alternative explanation involving terminal Schwann cells. After partial denervation of neonatal (but not adult) muscles, terminal Schwann cells at denervated endplates undergo apoptosis. We found that terminal (but not nodal) sprouting was absent in partially denervated neonatal muscles. This finding suggests that terminal Schwann cells, previously reported to guide terminal sprouts to denervated endplates in adult muscles, are necessary for the formation and growth of terminal sprouts. Moreover, partial denervation on P14 severely weakened the remaining, uninjured synapses, suggesting that neonatal motoneurons may withdraw terminals after the denervation of nearby fibers. These findings have implications for the interpretation of previous studies on synapse elimination and offer insight into the failure of young motor units to expand after partial denervation. PMID- 10516311 TI - Role of neurotrophin receptor TrkB in the maturation of rod photoreceptors and establishment of synaptic transmission to the inner retina. AB - Brain-derived neurotrophic factor (BDNF) acts through TrkB, a receptor with kinase activity, and mitigates light-induced apoptosis in adult mouse rod photoreceptors. To determine whether TrkB signaling is necessary for rod development and function, we examined the retinas of mice lacking all isoforms of the TrkB receptor. Rod migration and differentiation occur in the mutant retina, but proceed at slower rates than in wild-type mice. In postnatal day 16 (P16) mutants, rod outer segment dimensions and rhodopsin content are comparable with those of photoreceptors in P12 wild type (WT). Quantitative analyses of the photoreceptor component in the electroretinogram (ERG) indicate that the gain and kinetics of the rod phototransduction signal in dark-adapted P16 mutant and P12 WT retinas are similar. In contrast to P12 WT, however, the ERG in mutant mice entirely lacks a b-wave, indicating a failure of signal transmission in the retinal rod pathway. In the inner retina of mutant mice, although cells appear anatomically and immunohistochemically normal, they fail to respond to prolonged stroboscopic illumination with the normal expression of c-fos. Absence of the b wave and failure of c-fos expression, in view of anatomically normal inner retinal cells, suggest that lack of TrkB signaling causes a defect in synaptic signaling between rods and inner retinal cells. Retinal pigment epithelial cells and cells in the inner retina, including Muller, amacrine, and retinal ganglion cells, express the TrkB receptor, but rod photoreceptors do not. Moreover, inner retinal cells respond to exogenous BDNF with c-fos expression and extracellular signal-regulated kinase phosphorylation. Thus, interactions of rods with TrkB expressing cells must be required for normal rod development. PMID- 10516313 TI - A role for HSP27 in sensory neuron survival. AB - Peripheral nerve injury in neonatal rats results in the death of the majority of the axotomized sensory neurons by 7 d after injury. In adult animals, however, all sensory neurons survive for at least 4 months after axotomy. How sensory neurons acquire the capacity to survive axonal injury is not known. Here we describe how the expression of the small heat shock protein 27 (HSP27) is correlated with neuronal survival after axotomy in vivo and after NGF withdrawal in vitro. The number of HSP27-immunoreactive neurons in the L4 DRG is low at birth and does not change significantly for 21 d after postnatal day 0 (P0) sciatic nerve axotomy. In contrast, in the adult all axotomized neurons begin to express HSP27. One week after P0 sciatic nerve section the total number of neurons in the L4 DRG is dramatically reduced, but all surviving axotomized neurons, as identified by c-jun immunoreactivity, are immunoreactive for HSP27. In addition, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling reveals that very few HSP27-expressing neurons are dying 48 hr after neonatal axotomy. In vitro, a similar correlation exists between HSP27 expression and survival; in P0 DRG cultures, neurons that express HSP27 preferentially survive NGF withdrawal. Finally, overexpression of human HSP27 in neonatal rat sensory and sympathetic neurons significantly increases survival after NGF withdrawal, with nearly twice as many neurons surviving at 48 hr. Together these results suggest that HSP27 in sensory neurons plays a role in promoting survival after axotomy or neurotrophin withdrawal. PMID- 10516314 TI - Sonic hedgehog regulates proliferation and inhibits differentiation of CNS precursor cells. AB - Activation of the Sonic hedgehog (Shh) signal transduction pathway is essential for normal pattern formation and cellular differentiation in the developing CNS. However, it is also thought to be etiological in primitive neuroectodermal tumors. We adapted GAL4/UAS methodology to ectopically express full-length Shh in the dorsal neural tube of transgenic mouse embryos commencing at 10 d postcoitum (dpc), beyond the period of primary dorsal-ventral pattern formation and floorplate induction. Expression of Shh was maintained until birth, permitting us to investigate effects of ongoing exposure to Shh on CNS precursors in vivo. Proliferative rates of spinal cord precursors were twice that of wild-type littermates at 12.5 dpc. In contrast, at late fetal stages (18.5 dpc), cells that were Shh-responsive but postmitotic were present in persistent structures reminiscent of the ventricular zone germinal matrix. This tissue remained blocked in an undifferentiated state. These results indicate that cellular competence restricts the proliferative response to Shh in vivo and provide evidence that proliferation and differentiation can be regulated separately in precursor cells of the spinal cord. Thus, Hedgehog signaling may contribute to CNS tumorigenesis by directly enhancing proliferation and preventing neural differentiation in selected precursor cells. PMID- 10516315 TI - Cocaine upregulates the dopamine transporter in fetal rhesus monkey brain. AB - Cocaine is a highly addictive drug that binds to the dopamine transporter (DAT), inhibits the reuptake of dopamine, and initiates multiple actions within midbrain dopaminergic systems. Using the rhesus monkey, we have investigated the consequences of in utero cocaine exposure on the expression of DAT in the fetal brain. By using the selective DAT ligand [125I]RTI-121 and tyrosine hydroxylase (TH) immunocytochemistry, we found that DAT binding sites are highly developed by day 70 of gestation and show a distribution pattern similar to TH. The rank order of specific 3beta-(4-[125I]iodophenyl)tropane-2beta-carboxylic acid isopropyl ester ([125I]RTI-121) binding densities was substantia nigra-ventral tegmental area > putamen > caudate > lateral hypothalamus > accumbens > linear/interfascicular nuclei >/= globus pallidus > prefrontal cortex. Furthermore, we observed that DAT mRNA was differentially expressed within fetal midbrain dopamine neurons with the highest levels detected in the ventral tier of the substantia nigra pars compacta, and the lowest levels in the ventral tegmental area and the linear/interfascicular nuclei. In utero cocaine exposure between days 22 and 70 significantly increased DAT mRNA expression, and the density of [125I]RTI-121 binding sites within midbrain dopamine neurons in the 70 d-old fetus. This increased DAT expression is accompanied by other presynaptic and postsynaptic neuronal changes, which collectively suggest that midbrain dopamine neurons are hypoactive after prolonged cocaine exposure, a state that may be a contributing factor in the development of attention deficit disorders observed in subjects exposed prenatally to cocaine. PMID- 10516316 TI - Bovine CNS myelin contains neurite growth-inhibitory activity associated with chondroitin sulfate proteoglycans. AB - The absence of fiber regrowth in the injured mammalian CNS is influenced by several different factors and mechanisms. Besides the nonconducive properties of the glial scar tissue that forms around the lesion site, individual molecules present in CNS myelin and expressed by oligodendrocytes, such as NI-35/NI-250, bNI-220, and myelin-associated glycoprotein (MAG), have been isolated and shown to inhibit axonal growth. Here, we report an additional neurite growth-inhibitory activity purified from bovine spinal cord myelin that is not related to bNI-220 or MAG. This activity can be ascribed to the presence of two chondroitin sulfate proteoglycans (CSPGs), brevican and the brain-specific versican V2 splice variant. Neurite outgrowth of neonatal cerebellar granule cells and of dorsal root ganglion neurons in vitro was strongly inhibited by this myelin fraction enriched in CSPGs. Immunohistochemical staining revealed that brevican and versican V2 are present on the surfaces of differentiated oligodendrocytes. We provide evidence that treatment of oligodendrocytes with the proteoglycan synthesis inhibitors beta-xylosides can strongly influence the growth permissiveness of oligodendrocytes. beta-Xylosides abolished cell surface presentation of brevican and versican V2 and reversed growth cone collapse in encounters with oligodendrocytes as demonstrated by time-lapse video microscopy. Instead, growth cones were able to grow along or even into the processes of oligodendrocytes. Our results strongly suggest that brevican and versican V2 are additional components of CNS myelin that contribute to its nonpermissive substrate properties for axonal growth. Expression of these CSPGs on oligodendrocytes may indicate that they participate in the restriction of structural plasticity and regeneration in the adult CNS. PMID- 10516317 TI - Dopamine affects parvalbumin expression during cortical development in vitro. AB - This study was undertaken to determine how dopamine influences cortical development. It focused on morphogenesis of GABAergic neurons that contained the calcium-binding protein parvalbumin (PV). Organotypic slices of frontoparietal cortex were taken from neonatal rats, cultured with or without dopamine, harvested daily (4-30 d), and immunostained for parvalbumin. Expression of parvalbumin occurred in the same regional and laminar sequence as in vivo. Expression in cingulate and entorhinal preceded that in lateral frontoparietal cortices. Laminar expression progressed from layer V to VI and finally II-IV. Somal labeling preceded fiber labeling by 2 d. Dopamine accelerated PV expression. In treated slices, a dense band of PV-immunoreactive neurons appeared in layer V at 7 d in vitro (DIV), and in all layers of frontoparietal cortex at 14 DIV, whereas in control slices such labeling did not appear until 14 and 21 DIV, respectively. The laminar distribution and dendritic branching of PV immunoreactive neurons were quantified. More labeled neurons were in the superficial layers, and their dendritic arborizations were significantly increased by dopamine. Treatment with a D1 receptor agonist had little effect, whereas a D2 agonist mimicked dopamine's effects. Likewise, the D2 but not the D1 antagonist blocked dopamine-induced changes, indicating that they were mediated primarily by D2 receptors. Parvalbumin expression was accelerated by dopaminergic reinnervation of cortical slices that were cocultured with mesencephalic slices. Coapplication of the glutamate NMDA receptor antagonist MK801 or AP5 blocked dopamine-induced increases in dendritic branching, suggesting that changes were mediated partly by interaction with glutamate to alter cortical excitability. PMID- 10516318 TI - Astroglial differentiation of cortical precursor cells triggered by activation of the cAMP-dependent signaling pathway. AB - In the developing brain, differentiation of neural precursors into neurons or glial cells occurs in response to neurotrophic factors acting on the cell surface. Intracellular signaling mechanisms that relay information to initiate differentiative responses of neural precursor cells are poorly understood. To investigate whether stimulation of the cAMP-dependent signaling pathway participates in differentiative responses of cells in the developing CNS, we performed experiments using both conditionally immortalized neural precursor cells (RC2.E10 cells) and primary cultures of cells from developing rat cortex. Initially, we determined that RC2.E10 cells retain phenotypic features of neural precursors after inactivation of the immortalizing oncogene, a temperature sensitive mutant of the simian virus 40 large-T antigen (SV40T). We found that, once SV40T is inactivated, RC2.E10 cells cease to divide and die. However, RC2. E10 cells can proliferate in the presence of basic fibroblast growth factor. In addition, they express nestin, a marker of neural precursor cells. Both RC2.E10 cells and primary cortical precursor cells undergo astroglial differentiation in response to cAMP stimulation by treatment with 8-bromo-cAMP. In both cases, cAMP induced astrocyte differentiation is characterized by morphological changes, stimulation of glial fibrillary acidic protein expression, downregulation of nestin expression, and decreased proliferation. No increases in the expression of neuronal or oligodendrocytic markers were observed. Our results support the notion that the developing CNS contains neural precursor cells with the capacity of undergoing astrocyte differentiation in response to increased intracellular cAMP concentrations. PMID- 10516319 TI - Parametric population representation of retinal location: neuronal interaction dynamics in cat primary visual cortex. AB - Neuronal interactions are an intricate part of cortical information processing generating internal representations of the environment beyond simple one-to-one mappings of the input parameter space. Here we examined functional ranges of interaction processes within ensembles of neurons in cat primary visual cortex. Seven "elementary" stimuli consisting of small squares of light were presented at contiguous horizontal positions. The population representation of these stimuli was compared to the representation of "composite" stimuli, consisting of two squares of light at varied separations. Based on receptive field measurements and by application of an Optimal Linear Estimator, the representation of retinal location was constructed as a distribution of population activation (DPA) in visual space. The spatiotemporal pattern of the DPA was investigated by obtaining the activity of each neuron for a sequence of time intervals. We found that the DPA of composite stimuli deviates from the superposition of its components because of distance-dependent (1) early excitation and (2) late inhibition. (3) The shape of the DPA of composite stimuli revealed a distance-dependent repulsion effect. We simulated these findings within the framework of dynamic neural fields. In the model, the feedforward response of neurons is modulated by spatial ranges of excitatory and inhibitory interactions within the population. A single set of model parameters was sufficient to describe the main experimental effects. Combined, our results indicate that the spatiotemporal processing of visual stimuli is characterized by a delicate, mutual interplay between stimulus dependent and interaction-based strategies contributing to the formation of widespread cortical activation patterns. PMID- 10516320 TI - Choosing between small, likely rewards and large, unlikely rewards activates inferior and orbital prefrontal cortex. AB - Patients sustaining lesions of the orbital prefrontal cortex (PFC) exhibit marked impairments in the performance of laboratory-based gambling, or risk-taking, tasks, suggesting that this part of the human PFC contributes to decision-making cognition. However, to date, little is known about the particular regions of the orbital cortex that participate in this function. In the present study, eight healthy volunteers were scanned, using H(2)(15)0 PET technology, while performing a novel computerized risk-taking task. The task involved predicting which of two mutually exclusive outcomes would occur, but critically, the larger reward (and penalty) was associated with choice of the least likely outcome, whereas the smallest reward (and penalty) was associated with choice of the most likely outcome. Resolving these "conflicting" decisions was associated with three distinct foci of regional cerebral blood flow increase within the right inferior and orbital PFC: laterally, in the anterior part of the middle frontal gyrus [Brodmann area 10 (BA 10)], medially, in the orbital gyrus (BA 11), and posteriorly, in the anterior portion of the inferior frontal gyrus (BA 47). By contrast, increases in the degree of conflict inherent in these decisions was associated with only limited changes in activity within orbital PFC and the anterior cingulate cortex. These results suggest that decision making recruits neural activity from multiple regions of the inferior PFC that receive information from a diverse set of cortical and limbic inputs, and that the contribution of the orbitofrontal regions may involve processing changes in reward-related information. PMID- 10516321 TI - Vector averaging occurs downstream from learning in smooth pursuit eye movements of monkeys. AB - How are sensory-motor transformations organized in a cortical motor system? In general, sensory information is transformed through a variety of signal processing operations in the context of distinct coordinate frameworks. We studied the interaction of two distinct operations in pursuit eye movements, learning and vector-averaging, to gain insight into their underlying coordinate frameworks and their sequence in sensory-motor processing. Learning was induced in the initiation of pursuit eye movements by targets that moved initially at one speed for 100 msec and then increased or decreased to a sustained final speed. Vector averaging was studied by comparing the initial eye acceleration evoked by the simultaneous motion of two targets with that evoked by each target singly. Learning caused specific effects on the direction of the vector-averaged responses to two-target stimuli that included one target moving in the direction used to induce learning. Learned increases or decreases in eye acceleration caused the direction of the responses to two-targets to rotate toward or away from the learning direction. Learning also caused nonspecific changes in the responses to two-target stimuli. After any learning protocol, two-target responses usually became smaller, and their directions rotated away from the axis of the target motion used for learning. Quantitative analysis showed that the specific effects of learning were predicted most closely by a model in which vector averaging occurs downstream from the site(s) of learning. We suggest that the pursuit system creates parallel commands for potential movements to each of the targets in two-target stimuli, and that learning occurs in the coordinates of the potential movements. PMID- 10516322 TI - Muscimol inactivation of the dorsal hippocampus impairs contextual retrieval of fear memory. AB - Some models of hippocampal function have suggested a role of the hippocampus in contextual memory retrieval. We have examined this hypothesis by assessing the impact of reversible inactivation of the dorsal hippocampus (DH) on the context specific expression of latent inhibition, a decrement in conditional responding produced by preexposure to a to-be-conditional stimulus. In Experiment 1, rats received tone preexposure either in the context that would later be used for extinction testing (context A) or in a different context (context C); a third group of rats did not receive tone preexposure. All rats then received fear conditioning, which consisted of tone-footshock pairings, in a third distinct context (context B). The following day conditional fear to the tone was assessed in one of the preexposure contexts (context A) by measuring freezing during a tone extinction test. Rats preexposed and tested in the same context exhibited less freezing to the tone than either rats preexposed and tested in different contexts or non-preexposed rats. These results indicate that the expression of latent inhibition is context specific. In Experiment 2, DH inactivation eliminated the context-specific expression of latent inhibition. Compared with saline-infused rats, rats infused with muscimol into the DH exhibited low levels of tone freezing independent of whether they had received tone preexposure in the test context or in a different context. Experiment 3 revealed normal contextual discrimination in rats after DH inactivation. These results suggest the DH is required for contextual memory retrieval in a latent inhibition paradigm. PMID- 10516323 TI - Ventromedial thalamic neurons convey nociceptive signals from the whole body surface to the dorsolateral neocortex. AB - The somatosensory properties of ventromedial (VM) thalamic neurons were investigated in anesthetized rats by examining their responses to calibrated cutaneous stimuli. A population of neurons within the lateral part of the ventromedial thalamus (VMl) showed two peaks of activation after percutaneous electrical stimuli, regardless of which part of the body was stimulated. The early and late peaks were elicited by Adelta- and C-fiber activities with mean conduction velocities of 12.9 +/- 0.9 and 1 +/- 0.2 m/sec, respectively. These responses were strongly depressed or blocked after microinjections within the medullary subnucleus reticularis dorsalis of xylocaine or the NMDA antagonist MK 801. None of the VMl neurons responded to innocuous cutaneous or proprioceptive stimuli. In contrast, all these neurons responded to noxious mechanical and thermal stimulation of the limbs and showed monotonic increases in their discharges to increasingly strong noxious cutaneous stimuli. In addition, some VMl neurons were antidromically activated by stimulation in layer I of the dorsolateral frontal cortex. These findings suggest that the rat VMl conveys and encodes cutaneous nociceptive inputs from any part of the body surface to layer I of the dorsolateral neocortex. This reticulo-thalamo-cortical network may allow any signal of pain to gain access to widespread areas of the neocortex and thus help prime the cortex for attentional reactions and/or the coordination of motor responses. PMID- 10516324 TI - Effects of gravitational load on jaw movements in speech. AB - External loads arising as a result of the orientation of body segments relative to gravity can affect the achievement of movement goals. The degree to which subjects adjust control signals to compensate for these loads is a reflection of the extent to which forces affecting motion are represented neurally. In the present study we assessed whether subjects, when speaking, compensate for loads caused by the orientation of the head relative to gravity. We used a mathematical model of the jaw to predict the effects of control signals that are not adjusted for changes to head orientation. The simulations predicted a systematic change in sagittal plane jaw orientation and horizontal position resulting from changes to the orientation of the head. We conducted an empirical study in which subjects were tested under the same conditions. With one exception, empirical results were consistent with the simulations. In both simulation and empirical studies, the jaw was rotated closer to occlusion and translated in an anterior direction when the head was in the prone orientation. When the head was in the supine orientation, the jaw was rotated away from occlusion. The findings suggest that the nervous system does not completely compensate for changes in head orientation relative to gravity. A second study was conducted to assess possible changes in acoustical patterns attributable to changes in head orientation. The frequencies of the first (F1) and second (F2) formants associated with the steady-state portion of vowels were measured. As in the kinematic study, systematic differences in the values of F1 and F2 were observed with changes in head orientation. Thus the acoustical analysis further supports the conclusion that control signals are not completely adjusted to offset forces arising because of changes in orientation. PMID- 10516325 TI - Chronic morphine treatment alters NMDA receptor-mediated synaptic transmission in the nucleus accumbens. AB - In a study of a possible substrate underlying morphine addiction, we examined NMDA receptor-mediated synaptic transmission of core nucleus accumbens neurons after chronic morphine treatment, using intracellular recording in a slice preparation of rat. We evoked pharmacologically isolated NMDA EPSCs by local stimulation and elicited inward currents by NMDA superfusion. In control slices, Mg(2+) and phorbol 12,13-diacetate (PDAc), a protein kinase C activator, strongly inhibited and increased, respectively, NMDA EPSC amplitudes. The PDAc effects were likely postsynaptic because PDAc enhanced the currents evoked by superfused NMDA to the same extent that it did the NMDA EPSCs. Chronic morphine treatment significantly decreased NMDA EPSC amplitudes and the sensitivity of NMDA EPSCs to Mg(2+) and PDAc, as well as the kinetics of the decay (inactivation rate) of the EPSCs (from 97 +/- 2.5 msec in untreated rats to 78.7 +/- 1.8 msec in slices from treated rats). One week after withdrawal, the Mg(2+) and PDAc effects were still significantly less than those in control slices. Interestingly, 1 week of withdrawal led to an increased NMDA EPSC inactivation rate compared with controls. These data demonstrate that chronic morphine treatment significantly alters NMDA receptor-mediated synaptic transmission in the accumbens, and these effects persist 1 week after withdrawal. These long-term effects may represent an important neuroadaptation in opiate dependence. PMID- 10516326 TI - Presynaptic long-term potentiation in corticothalamic synapses. AB - The thalamus and neocortex are two highly organized and complex brain structures that work in concert with each other. The largest synaptic input to the thalamus arrives from the neocortex via corticothalamic fibers. Using brain slices, we describe long-term potentiation (LTP) in corticothalamic fibers contacting the ventrobasal thalamus. Corticothalamic LTP is input-specific, NMDA receptor independent, and reversible. The induction of corticothalamic LTP is entirely presynaptic and Ca(2+)-dependent. The expression of corticothalamic LTP is associated with a decrease in paired-pulse facilitation (PPF) and blocked by an inhibitor of the cAMP-dependent protein kinase A (PKA). Consistent with an involvement of cAMP and PKA, activation of adenylyl cyclase induced a synaptic enhancement that was associated with a decrease in PPF and occluded LTP. Corticothalamic LTP may serve to enhance the efficacy of cortico-cortical communication via the thalamus and/or to mediate experience-dependent long-term modifications of thalamocortical receptive fields. PMID- 10516327 TI - The spectral main sequence of human saccades. AB - Despite the many models of saccadic eye movements, little attention has been paid to the shape of saccade trajectories. Some investigators have argued that saccades are driven by a rectangular "bang-bang" neural control signal, whereas others have emphasized the similarity to fast arm movement trajectories, such as the "minimum jerk" profile. However, models have not been tested rigorously against empirical trajectories. We examined the Fourier transforms of saccades and compared them with theoretical models. Horizontal saccades were recorded from 10 healthy subjects. The Fourier transform of each saccade was accurately computed using a padded fast Fourier transform (FFT), and the frequencies of the first three minima (M1, M2, M3) in each energy spectrum were measured to a precision of 0.12 Hz. Each subject showed near-linear trends in the relationships among M1, M2, and M3 and the reciprocal of duration (1/T), which we call the "spectral main sequence." Extrapolation of plots did not pass through the origin, indicating a subtle departure from self-similarity. Bivariate confidence regions were established to allow for slope-intercept variability. The nonharmonic relationships seen cannot arise from a rectangular saccadic pulse driving a linear ocular plant. The relationships are also incompatible with minimum acceleration, minimum jerk, or higher-order minimum square derivative trajectories. The best fits were made by trajectories that minimize postmovement variance with signal-dependent noise (). It is concluded that the spectral main sequence is exquisitely sensitive to the saccade trajectory and should be used to test objectively all present and future models of saccades. PMID- 10516328 TI - Two-stage, input-specific synaptic maturation in a nucleus essential for vocal production in the zebra finch. AB - In most songbirds, vocal learning occurs through two experience-dependent phases, culminating in a reduction of behavioral plasticity called song crystallization. At ends of developmentally plastic periods in other systems, synaptic properties change in a fashion appropriate to limit plasticity. Maturation of glutamatergic synapses often involves a reduction in duration of NMDA receptor (NMDAR)-mediated synaptic responses and a coincident reduction in the contribution of NMDARs to synaptic transmission. We hypothesized that similar changes in the zebra finch song system help limit behavioral plasticity during song development. Nucleus robustus archistriatalis (RA) is a key nucleus in the forebrain song motor pathway and receives glutamatergic input from the motor nucleus HVc. RA also receives glutamatergic input, mediated primarily by NMDARs, from the lateral magnocellular nucleus of the anterior neostriatum, which is part of a circuit essential for learning but not song production. We examined whether synaptic maturation occurs in either input to RA by recording synaptic currents in brain slices prepared from zebra finches of different ages. We find the motor input from HVc to RA uses both AMPA receptors (AMPARs) and NMDARs, and synaptic maturation occurs in two phases: an early reduction in duration of NMDAR-mediated synaptic currents in both inputs, and a later reduction in the NMDAR contribution to synaptic responses in the motor pathway. Although NMDAR kinetics change too early to account for crystallization, the reduction of the relative NMDAR contribution to synaptic transmission could contribute to the onset of crystallization. Thus, synaptic maturation events can be temporally distinct and input-specific and may play different roles in behavioral plasticity. PMID- 10516329 TI - Sensory loss by selected whisker removal produces immediate disinhibition in the somatosensory cortex of behaving rats. AB - This study used extracellular unit recordings in behaving animals to evaluate thalamocortical response transformations in the rat whisker-barrel system. Based on previous acute studies using controlled whisker stimulation, we hypothesized that in a cortical barrel adjacent (non-principal) whiskers exert a net inhibitory effect. In contrast, in thalamic barreloid neurons, the effects of neighboring whiskers should be net facilitatory. We evaluated these hypotheses by recording unit activity at 21 sites in 17 animals trained to explore a wire mesh screen with their whiskers. In the middle of the recording session, selected vibrissae were clipped close to the skin surface. The absence of whiskers surrounding the principal whisker was associated with a mean 20% increase in cortical activity and, conversely, a 37% decrease in the thalamic activity. Removal of the principal whisker resulted in a 50% decrease in cortical unit firing. Findings are consistent with the idea that, in the behaving animal, each barrel uses multi-whisker thalamic inputs and local inhibitory circuitry to sharpen the receptive field properties of its constituent neurons. Cortical disinhibition as a consequence of selective whisker removal is likely to be an important factor underlying altered receptive field properties in sensory deprived animals. PMID- 10516330 TI - Behavioral and neural bases of noncoincidence learning in Hermissenda. AB - Neurobiological studies of associative learning and memory have focused nearly exclusively on the analysis of neural plasticity resulting from paired stimuli. A second major category of associative-learning processes, one that has been conspicuously neglected in cellular studies, is that of conditioned inhibition (CI), learning that one stimulus signals the absence of another. The physiological bases of CI are obscure and unexplored. To study the behavioral and neural bases of CI, we exposed the nudibranch mollusc Hermissenda crassicornis to explicitly unpaired (EU) presentations of light and rotation. We report here that Hermissenda exhibited persistent increases in phototactic behavior after EU training. Retardation-of-learning test results provided further evidence that EU animals learned that light signaled the absence of rotation. The increased phototactic behavior of EU animals was paralleled by selective decreases in the magnitude of ocular type B cell photoresponses and the frequency of light elicited action potentials: the first report of a neural correlate of noncoincidence learning. Plasticity arising from explicitly unpaired stimulus presentations raises provocative questions as to how noncoincidence is detected and represented within the nervous system. PMID- 10516331 TI - Mauthner cell-initiated electromotor behavior is mediated via NMDA and metabotropic glutamatergic receptors on medullary pacemaker neurons in a gymnotid fish. AB - Weakly electric fish generate meaningful electromotor behaviors by specific modulations of the discharge of their medullary pacemaker nucleus from which the rhythmic command for each electric organ discharge (EOD) arises. Certain electromotor behaviors seem to involve the activation of specific neurotransmitter receptors on particular target cells within the nucleus, i.e., on pacemaker or on relay cells. This paper deals with the neural basis of the electromotor behavior elicited by activation of Mauthner cells in Gymnotus carapo. This behavior consists of an abrupt and prolonged increase in the rate of the EOD. The effects of specific glutamate agonists and antagonists on basal EOD frequency and on EOD accelerations induced by Mauthner cell activation were assessed. Injections of both ionotropic (AMPA, kainate, and NMDA) and metabotropic (trans-(+/-)-1-amino-1,3-cyclopentanedicarboxylic acid) glutamate agonists induced increases in EOD rate that were maximal when performed close to the soma of pacemaker cells. In contrast, injections in the proximity of relay cells were ineffective. Therefore, pacemaker neurons are probably endowed with diverse glutamate receptor subtypes, whereas relay cells are probably not. The Mauthner cell-evoked electromotor behavior was suppressed by injections of AP-5 and (+/-)-amino-4-carboxy-methyl-phenylacetic acid, NMDA receptor and metabotropic glutamate receptor antagonists, respectively. Thus, this electromotor behavior relies on the activation of the NMDA and metabotropic glutamate receptor subtypes of pacemaker cells. Our study gives evidence for the synergistic effects of NMDA and metabotropic receptor activation and shows how a simple circuit can produce specific electromotor outputs. PMID- 10516333 TI - New features for the journal of neuroscience PMID- 10516332 TI - Methamphetamine-induced neurotoxicity alters locomotor activity, stereotypic behavior, and stimulated dopamine release in the rat. AB - The neurochemical evidence of methamphetamine (MA)-induced toxicity to dopaminergic nerve terminals is well documented; however, the functional consequences are not clearly defined. The present study was designed to investigate whether MA-induced dopamine depletions affect locomotor activity, stereotypic behavior, and/or extracellular dopamine concentrations in the neostriatum. Male rats were treated with a neurotoxic regimen of MA (10 mg/kg, i.p., every 2 hr for four injections) or vehicle and tested for functional effects 1 week later. Animals that had received the neurotoxic regimen of MA showed a reduction in both caudate nucleus and nucleus accumbens dopamine contents of 56 and 30%, respectively. Furthermore, MA-treated rats exhibited a significant attenuation in spontaneous activity, as well as a significant diminution in MA (low dose)-stimulated locomotor activity as compared to vehicle treated rats. However, there were no differences in the MA (low dose)-induced increases in extracellular dopamine concentrations in the caudate nucleus or the nucleus accumbens core of either group. Interestingly, the acute administration of higher doses of MA elicited a significantly augmented stereotypic response and a significantly attenuated increase in the extracellular concentration of dopamine in the caudate nucleus of rats treated with a neurotoxic regimen of MA as compared to vehicle-treated animals. These data indicate that MA-induced neurotoxicity results in abnormal dopamine-mediated behaviors, as well as a brain region-specific impairment in stimulated dopamine release. PMID- 10516334 TI - Functional dichotomy within the vomeronasal system: distinct zones of neuronal activity in the accessory olfactory bulb correlate with sex-specific behaviors. AB - Chemosensory neurons in the vomeronasal organ (VNO) detect pheromones that elicit social and reproductive behaviors in most terrestrial vertebrates. Vomeronasal receptor neurons are chemoarchitecturally divided into two populations based on their position in the VNO, the type of G-protein subunit expressed, the family of putative pheromone receptor expressed, and termination site of their axons in the accessory olfactory bulb (AOB). To investigate the functional implications of these two segregated VNO-AOB pathways, we stimulated mice with pheromonal cues associated with different behavioral contexts and examined cellular activation patterns in the AOB. Exposure of ICR male mice to BALB/c males resulted in aggressive behavior, accompanied by a VNO-dependent increase in c-fos immunoreactivity in a cluster of cells located almost exclusively in the caudal AOB in both strains. This caudal cluster of activated cells did not appear to require the overt display of aggressive behavior because it was present in both the dominant and submissive males and could be evoked when the stimulus animal was anesthetized. In contrast, exposure of an ICR male to an ICR female in diestrus resulted in activation of cells located predominantly in the rostral AOB. Our findings indicate that male-to-male interactions involving interstrain recognition activate a separate population of vomeronasal receptor neurons than chemosensory cues detected in a sexual context. The results suggest that the dichotomy in the peripheral vomeronasal system serves to separate pheromones based on the behaviors they drive. As such, the results provide a bioassay for identifying pheromone molecules. PMID- 10516335 TI - Activity-dependent regulation of potassium currents in an identified neuron of the stomatogastric ganglion of the crab Cancer borealis. AB - Identified neurons of the stomatogastric ganglion of the crab Cancer borealis were voltage-clamped, and the current densities of three K+ currents were measured. The current densities of each of the three K+ currents varied twofold to fivefold in inferior cardiac (IC) neurons from different animals. Conventionally, this degree of variability has been attributed to experimental artifacts. Instead, we suggest that it reflects a natural variability that may be related to an underlying process of plasticity. First, we found that there is no fixed ratio among the three K+ currents. Second, we found that several hours of stimulation with depolarizing current pulses (0.5 sec duration at 1 Hz) altered the current density of the Ca2+-dependent outward current, IK(Ca), and the transient outward current, IA. This stimulation paradigm mimics the normal pattern of activity for these neurons. The effect of stimulation on the IA current density was eliminated when Ca2+ influx was blocked by extracellular Cd2+. In contrast, the K+ current densities of the lateral pyloric (LP) neuron were unaffected by the same pattern of stimulation, and the currents expressed by both the IC and the LP neurons were insensitive to hyperpolarizing pulses at the same frequency. We conclude that the conductance densities expressed by neurons may vary continually depending on the recent history of electrical activity in the preparation, and that intracellular Ca2+ may play a role in the processes by which activity influences the regulation of current densities in neurons. PMID- 10516336 TI - Composition and decomposition of internal models in motor learning under altered kinematic and dynamic environments. AB - The learning process of reaching movements was examined under novel environments whose kinematic and dynamic properties were altered. We used a kinematic transformation (visuomotor rotation), a dynamic transformation (viscous curl field), and a combination of these transformations. When the subjects learned the combined transformation, reaching errors were smaller if the subject first learned the separate kinematic and dynamic transformations. Reaching errors under the kinematic (but not the dynamic) transformation were smaller if subjects first learned the combined transformation. These results suggest that the brain learns multiple internal models to compensate for each transformation and has some ability to combine and decompose these internal models as called for by the occasion. PMID- 10516337 TI - A role for the bed nucleus of the stria terminalis, but not the amygdala, in the effects of corticotropin-releasing factor on stress-induced reinstatement of cocaine seeking. AB - We have shown that intracerebroventricular administration of the corticotropin releasing factor (CRF) receptor antagonist D-Phe CRF(12-41), blocks footshock induced reinstatement of drug seeking in cocaine-trained rats. We now report that D-Phe acts in the bed nucleus of the stria terminalis (BNST), and not in the amygdala, to block footshock-induced reinstatement of cocaine seeking. In addition, CRF injections in the BNST, and not in the amygdala, are sufficient to reinstate cocaine seeking. Rats were trained to self-administer cocaine intravenously on a fixed ratio (FR-1) schedule of reinforcement. After 5 drug free days, animals were returned to the self-administration chambers and given daily extinction and reinstatement test sessions. To test the effects of D-Phe CRF(12-41) on stress-induced reinstatement, rats were pretreated with vehicle or D-Phe in either the BNST (10 or 50 ng per side) or amygdala (50 or 500 ng per side) before being exposed to 15 min of intermittent footshock stress. To test whether injections of CRF itself could induce reinstatement, rats were given vehicle or CRF in either the BNST (100 or 300 ng per side) or amygdala (300 ng per side) 15 min before the session. Injections of D-Phe into the BNST completely blocked footshock-induced reinstatement of cocaine seeking; injections of CRF itself in this structure induced reinstatement. Injections of these compounds into the amygdala were without effect. These findings suggest that activation of CRF receptors in the BNST, but not in the amygdala, is critical for footshock induced reinstatement of cocaine seeking. PMID- 10516338 TI - In search of ideal hemodialysis: is prolonged frequent dialysis the answer? AB - Advances in technology have made it possible to deliver a high Kt/V in a shorter time. The realization that duration of dialysis may be an important predictor of survival independent of dialysis dose has resulted in the popularity of prolonged slow dialysis (PHD). The longer duration and increased frequency of dialysis achieve excellent small- and middle-molecular weight solute clearance and also attenuate the peak concentration of uremic toxins. The slow dialysis process enables the equilibration of tissue and vascular compartments, resulting in better clearance and decreased postdialysis rebound increase in solutes. Gentle, persistent ultrafiltration allows the control of hypertension with minimal antihypertensive use. The intense and more frequent dialysis improves appetite and permits liberalization of diet. This greater dietary protein intake results in a progressive increase in serum albumin level and dry weight. Nocturnal hemodialysis achieves control of hyperphosphatemia without phosphate binders and a significant reduction in serum beta(2)-microglobulin levels. Normalization of extracellular volume, better clearance of uremic toxins, and improved nutrition result in a significant improvement in survival. The flexible time schedule with home hemodialysis and improvement of sleep and neurocognitive function allow better rehabilitation. The available evidence indicates PHD may be closer to the concept of an ideal dialysis, but there is lingering uncertainty about the consequence of prolonged immune stimulation, catabolism, and loss of essential solutes with these therapies. PMID- 10516339 TI - Increased serum and urinary neopterin in nephrotic syndrome indicate cell mediated immune dysfunction. AB - T-cell-mediated immune disturbances are likely but not certain to cause the nephrotic syndrome. Because neopterin (NP) production is closely related to activation of cell-mediated immunity, we addressed the question by measuring serum NP concentrations and urinary NP/creatinine (Cr) ratios, as well as by assessing interstitial lymphocyte and monocyte infiltration in the kidney and activation of the same cell types in peripheral blood. Finally, we observed whether urinary NP/Cr ratios in nephrotic syndrome are changed by steroid therapy. Seventy-four patients with primary glomerulonephritis were divided into 4 groups based on presence or absence of nephrotic syndrome and presence or absence of mesangial proliferation and expansion. Serum and urinary NP concentrations were measured chromatographically. Infiltrating cells in the kidney were identified by immunohistochemistry, and activation of peripheral blood cells was examined by fluorescent surface marker antibodies and flow cytometry. Irrespective of the pathohistology of glomeruli, nephrotic groups showed significantly higher urinary NP/Cr ratios and serum NP concentrations. Nephrotic groups also exhibited more activation of T cells in peripheral blood than did nonnephrotic groups or a healthy control group. Serum NP did not correlate with extent of interstitial renal infiltrates. Steroid therapy decreased urinary NP/Cr ratios in steroid-responsive patients, but not in steroid resistant patients. Increased serum NP concentrations and urinary NP/Cr ratios may reflect disordered cell-mediated immunity in the nephrotic syndrome, irrespective of glomerular histology or interstitial cell infiltration. PMID- 10516341 TI - ACE inhibition induces regression of proteinuria and halts progression of renal damage in a genetic model of progressive nephropathy. AB - Experimental data consistently indicate that renal disease progression is fully prevented in proteinuric glomerulopathies by long-enough angiotensin-converting enzyme (ACE) inhibition therapy. Whether regression of established proteinuria to normal can be achieved is, however, ill defined. The current study was designed with the aim to clarify whether ACE inhibition may induce regression of established proteinuria and renal structural damage in MWF rats, a genetic model of progressive proteinuria and renal injury. Animals treated with the ACE inhibitor lisinopril from 20 weeks of age (time when proteinuria is already important) and age-matched untreated rats were followed for 10 weeks. ACE inhibition normalized systolic blood pressure and progressively reduced proteinuria (from 172 +/- 79 to 81 +/- 23 mg/24 hours). In these animals, a highly significant correlation was obtained between baseline proteinuria and antiproteinuric response. At variance in untreated rats, proteinuria showed a marked increase in the 10-week follow-up period (from 165 +/- 57 to 325 +/- 86 mg/24 hours). Lisinopril prevented the progression of renal damage, as documented by a significantly lower incidence of glomeruli affected by sclerotic lesions (P < 0.01) than in untreated animals after the 10-week study period. Kidney tissue damage was comparable in lisinopril-treated rats and in untreated animals at 20 weeks of age, indicating that structural changes were arrested by the treatment. Thus, in proteinuric MWF rats, late-onset ACE inhibition normalized blood pressure, effectively and progressively restored high protein excretion rate toward normal values, and arrested progression of tissue damage. PMID- 10516340 TI - Can prolonged treatment improve the prognosis in adults with focal segmental glomerulosclerosis? AB - Eighty nephrotic adults with focal segmental glomerulosclerosis (FSGS) and plasma creatinine lower than 3 mg/dL were given corticosteroids (53 patients) or immunosuppressive agents (27 patients) for a median of 16 and 75 weeks, respectively. Forty-two patients responded with complete remission (29 patients, 36%) or partial remission (13 patients, 16%). Twenty-six patients who did not respond were treated again. Two patients obtained complete remission and 13 partial remission. The probability of remission was associated with treatment with corticosteroids (P = 0.0001; RR, 3. 93; 95% CI, 2.00 to 7.72), absence of arterial hypertension (P = 0. 0023; RR, 2.59; 95% CI, 1.41 to 4.79), and a percentage of hyaline glomeruli lower than 5% (P = 0.0152; RR, 2.04; 95% CI, 1.15 to 3.64). The probability of being alive at 110 months without doubling of plasma creatinine was 69%. The risk of renal insufficiency was correlated with mesangial proliferation (P = 0.0025; RR, 5.50; 95% CI, 1.82 to 16.60) and with interstitial fibrosis (P = 0.0231; RR, 4. 44; 95% CI, 1.23 to 16.08) at initial biopsy. Considering partial or complete remission as a time-dependent variable, only the lack of remission (P = 0.0027; RR, 7.23; 95% CI, 1.98 to 26.33) and mesangial proliferation (P = 0.0069; RR, 4.59; 95% CI, 1.52 to 13. 88) were correlated with renal failure. Major side effects were observed in 11 patients (5 infections, 1 peptic ulcer, 2 diabetes, 3 neoplasias). This study shows that 70% of nephrotic adults with FSGS may obtain complete or partial remission and maintain stable renal function for about 10 years when given a prolonged therapy with corticosteroids or immunosuppressive drugs. PMID- 10516342 TI - Growth factors and proinflammatory cytokines in the renal involvement of POEMS syndrome. AB - The POEMS syndrome is a multisystemic syndrome associated with plasma cell dyscrasia, characterized by the combination of polyneuropathy, organomegaly, endocrinopathy, M protein, and skin changes. Renal involvement in POEMS syndrome is rare (26 reported cases). It has been described as membranoproliferative glomerulonephritis-like lesions (MPGN-like), mesangiolytic glomerulonephritis, or thrombotic microangiopathy. Proinflammatory cytokines (TNF-alpha, IL-1, IL-6) have been implicated in the physiopathogenesis of POEMS syndrome, particularly when there is renal involvement. Growth factors (FGF-beta, TGF-beta, PDGF) have been implicated in renal lesions of the same histological type but of different origins. An increase in serum vascular endothelial growth factor (VEGF) has been reported in POEMS syndrome (20 of 22 cases). Circulating levels of these factors were determined in 4 patients with POEMS and renal involvement (3 MPGN-like, 1 MPGN-like, and mesangiolysis) and compared with those obtained in 4 patients with POEMS without clinical renal involvement and in 4 patients with primitive membranoproliferative glomerulonephritis (MPGN). TNF-alpha, IL-1beta, and IL-6 were determined with an immunoradiometric assay, and VEGF, PDGF, FGF-beta, and TGF-beta with an enzyme-linked immunosorbent assay. Among the patients with POEMS syndrome, there was no difference in proinflammatory cytokines and growth factors between those with or without renal involvement. VEGF is the only growth factor that differentiates MPGN in POEMS syndrome from primitive MPGN. PMID- 10516343 TI - Renal asymmetry in children with autosomal dominant polycystic kidney disease. AB - Although for decades autosomal dominant polycystic kidney disease (ADPKD) was considered a disease of adults, our recent longitudinal studies on children from ADPKD families have shown that the disease is evident by ultrasound imaging in approximately 75% of children who are carriers of the ADPKD1 gene, the most common form of ADPKD. Here we report that, in contrast to adults, the disease appears to be unilateral initially in approximately 17% of children. Asymmetric enlargement of the kidneys is also frequently observed. This renal asymmetry can be extreme and lead to diagnostic confusion. We present 2 unusual cases of asymmetric renal involvement that we have observed during the last 10 years. The first is a 14-year-old boy who was scheduled for a nephrectomy to relieve pain and whose family requested a second opinion. The second is a 10-year-old girl who was diagnosed with ADPKD in utero by prenatal ultrasound. After birth, 1 kidney progressively developed cysts and enlarged, whereas the other had only a few tiny cysts and remained normal in size. A review of the literature shows that presentations like these often lead to a nephrectomy or surgical biopsy. A carefully obtained family history and examination of both parents with ultrasound can help to avoid unnecessary invasive procedures. If pain is a prominent symptom, it can be treated by cyst aspiration if there are only a few cysts or a single dominant cyst. The molecular mechanism for extremely asymmetric renal disease remains to be elucidated. PMID- 10516344 TI - Changing trends of histopathology in childhood nephrotic syndrome. AB - This study was conducted to analyze the trend of histopathologic subtypes in idiopathic nephrotic syndrome (INS) in a homogenous racial group in India population. A prospective analysis of 400 consecutive children with INS was performed. Kidney biopsies were performed according to standard indications. Steroids were administered following the Arbeitsgeminschaft fur Padiatrische Nephrologie protocol. Cyclophosphamide was administered to children in the frequent-relapser, steroid-dependent, and steroid-nonresponder categories. Of the various histopathologic subtypes, focal segmental glomerulosclerosis (FSGS) was the most common (87 of 222 subtypes; 39.1%). Children who underwent biopsy between July 1992 and December 1996 (group B, n = 157) were compared with our initial published data of biopsies performed between January 1990 and June 1992 (group A, n = 65), with similar indications for biopsy in both groups. The incidence of FSGS was significantly greater in biopsies performed in the recent period (group B, 47% versus group A, 20%; P = 0.0002). The different clinical and biochemical parameters were also analyzed to differentiate FSGS from the other 2 subtypes. Hypertension (P = 0. 005), renal insufficiency at presentation (P = 0.001), and steroid resistance (P = 0.0006) were significantly greater in children with FSGS. On follow-up (mean, 5.4 years), children with FSGS were at a significantly greater risk for developing renal insufficiency (P = 0. 0001). We conclude there is a shift toward an increasing prevalence of FSGS over the years in the Indian population. This trend has immense therapeutic and prognostic significance. PMID- 10516345 TI - Intraperitoneal erythropoietin in children on peritoneal dialysis: A study of pharmacokinetics and efficacy. AB - The pharmacokinetics and efficacy of intraperitoneal (IP) recombinant human erythropoietin (rHuEPO) were investigated in children undergoing chronic peritoneal dialysis. Eight children were administered a single dose of 100 U/kg of rHuEPO IP with 50 mL of dialysate into a dry peritoneal cavity after nighttime peritoneal dialysis. Serum erythropoietin (EPO) levels were measured at 0, 8, 12, and 24 hours. A mean peak EPO level of 187 mU/mL was obtained at 12 hours. The area under the curve was 5,818 mU/h/mL, and relative bioavailability was similar to that found using subcutaneous (SC) dosing. Nine children completed 11 to 12 weeks of IP rHuEPO therapy. The patients maintained a normal hematocrit (34% +/- 2.3%) with a mean final IP rHuEPO dosage that was not significantly greater than the mean previous SC dosage (IP, 290 +/- 194 U/kg/wk; SC, 279 +/- 126 U/kg/wk; P = not significant). There appeared to be a trend for a slightly increased risk for peritonitis compared with historical controls at our center (relative risk = 3.1; 95% confidence interval, 0.92 to 6.3). IP rHuEPO is effective in children undergoing continuous cycling peritoneal dialysis without requiring increased rHuEPO dosages, but the possibility of an increased risk for peritonitis will need to be further explored. PMID- 10516346 TI - Pharmacokinetics of intraperitoneal epoetin alfa in patients on peritoneal dialysis using an 8-hour "dry dwell" dosing technique. AB - Pharmacokinetic studies of intraperitoneal (IP) epoetin alfa administered to continuous ambulatory peritoneal dialysis (CAPD) patients have shown low bioavailability, primarily attributable to the dilutional effect of coadministered dialysate. However, bioavailability is improved by instilling the dose into a dry peritoneum. The current study was designed to determine whether absorption after administration into a dry peritoneum is improved by extending the dry dosing period from 4 to 8 hours. The pharmacokinetics of a single 100 unit/kg IP epoetin alfa dose were studied in 8 noninfected CAPD patients. The dose was instilled into a dry peritoneum via the peritoneal catheter and allowed to dwell for 8 hours. CAPD was then resumed. Blood samples were collected for 96 hours after the dose. A 14-hour effluent dialysate sample was collected to determine epoetin alfa recovery. Enzyme immunoassay was used for epoetin alfa analysis of serum and effluent. Standard pharmacokinetic methods were employed for analysis of the serum concentration time data. The extent of epoetin alfa absorption was significantly greater than previously reported for a 4-hour dry dwell. The mean (+/-SD) dose-normalized area-under-the-curve (nlAUC(0-infinity)) using the 8-hour dry dwell dosing technique was 6,331 +/- 2,536 mIU. h/mL. This is significantly greater than the value of 2,589 +/- 1,450 mIU. h/mL (two-sided P value = 0.002) from a previous study in which patients received the same 100 unit/kg dose using a 4-hour dry dwell. The absorption of epoetin alfa administered by the intraperitoneal route is improved by extending the time the dose resides in a dry peritoneum. PMID- 10516347 TI - l-2-oxothiazolidine-4-carboxylic acid modulates function of peritoneal mesothelial cells in vitro. AB - The influence of the glutathione precursor, l-2-oxothiazolidine-4-carboxylic acid (OTZ), on the function of human peritoneal mesothelial cells in vitro, in conditions that mimic the in vivo effect of peritoneal dialysis solutions on mesothelium, was studied. Mesothelial monolayers were exposed to dialysis fluids (Dianeal 1.36 or Dianeal 3.86; Baxter Healthcare Corp, Round Lake, IL) that were diluted gradually with pooled-effluent dialysate obtained from patients undergoing continuous ambulatory peritoneal dialysis. In vitro exposure of mesothelium to standard dialysis fluid enhances their susceptibility to injury by hydrogen peroxide. OTZ added to dialysis solution in concentrations of 1 mmol/L prevented the toxic effect of hydrogen peroxide, probably by increasing intracellular glutathione. Mesothelial cells exposed to dialysis fluid become activated, evidenced by increased release of interleukin-6 and hyaluronan. OTZ used in concentrations of 1 mmol/L reduced that effect. Furthermore, the addition of glucose to the culture medium in a concentration of 45 mmol/L inhibits the proliferation of mesothelial cells; the presence of OTZ, 1 mmol/L, partially prevents the inhibitory effect of glucose. The results presented in this report show that by augmenting the intracellular concentration of glutathione in mesothelial cells by the addition of OTZ to the dialysis fluid, we can increase their resistance to the acute toxicity of free radicals and long-term toxicity of glucose. In addition, mesothelial cells with an increased glutathione level are less activated after their exposure to dialysis fluid. PMID- 10516348 TI - Hyperhomocyst(e)inemia and the prevalence of atherosclerotic vascular disease in patients with end-stage renal disease. AB - Hyperhomocyst(e)inemia is now recognized as an independent risk factor for atherosclerotic cardiovascular disease in patients with normal renal function. Hyperhomocyst(e)inemia is common in patients with chronic renal failure. This study is designed to look for an association between hyperhomocyst(e)inemia and atherosclerotic vascular disease in patients with end-stage renal disease (ESRD). Two hundred eighteen patients undergoing hemodialysis were enrolled onto the study and had predialysis bloodwork performed for total homocyst(e)ine, red blood cell folate, and vitamin B(12) levels. A history of clinically significant atherosclerotic vascular disease (ischemic heart disease, cerebrovascular disease, or peripheral vascular disease) was elicited by patient questionnaire and verified by careful inpatient and outpatient chart review. Atherosclerotic vascular disease was present in 45.9% of patients. Mean homocyst(e)ine concentration was 26.7 micromol/L (95% confidence interval [CI], 25.0 to 28.4) overall. Mean homocyst(e)ine concentration was 28.6 micromol/L (95% CI, 25.6 to 31.7) and 25.0 micromol/L (95% CI, 23.2 to 26.8) in patients with and without atherosclerotic disease, respectively (P = 0.036). The adjusted odds ratio for atherosclerotic disease was 2.12 (95% CI, 1.03 to 4.39) for those subjects with a homocyst(e)ine level in the highest quartile compared with the lowest 3 quartiles. In the 126 men, the adjusted odds ratio for atherosclerotic disease was 3.4 (95% CI, 1. 24 to 9.42) for those with homocyst(e)ine levels in the highest quartile compared with the lowest 3 quartiles. No association was found between homocyst(e)ine level and atherosclerotic disease in women. In conclusion, there is an association between hyperhomocyst(e)inemia and atherosclerotic vascular disease in patients undergoing dialysis. Prospective studies need to further examine the relationship between homocyst(e)ine level and atherosclerosis in women with ESRD. PMID- 10516349 TI - Effects of L-carnitine on leukocyte function and viability in hemodialysis patients: A double-blind randomized trial. AB - Excess morbidity and mortality among long-term hemodialysis patients because of infectious complications is partly caused by an impairment of cellular immune defense. We hypothesized this impairment is related to an abnormal carnitine metabolism also present in these patients. In a double-blind, randomized, placebo controlled trial, we investigated the effect of L-carnitine on phagocytic function and viability of blood leukocytes in 17 patients undergoing maintenance hemodialysis. After an observation period of 1 month, the patients received either 10 mg/kg of L-carnitine or placebo intravenously at the end of each hemodialysis session over a period of 4 months. Leukocyte oxidative metabolism was measured by means of luminol-enhanced chemiluminescence and superoxide generation after stimulation with Staphylococcus aureus or phorbol myristate acetate. Killing capacity and phagocytosis of radiolabeled staphylococci were determined. A lactate dehydrogenase (LDH) release test was applied to assess cell viability. We were unable to show an effect of L-carnitine on phagocytic function and viability in vivo. Several clinical parameters were observed during the trial. No statistically significant differences concerning dialysis-related morbidity, anemia, or reduction of blood urea nitrogen and creatinine levels were detected. Additionally, we tested the effect of L-carnitine on phagocytic function after in vitro incubation of blood leukocytes, which also showed no changes. LDH release was decreased, indicating an improved viability of these cells. The latter results were found after in vitro incubation of cells, but could not be confirmed in vivo. In summary, we could not show beneficial effects of L-carnitine administration in hemodialysis patients for the dosage and duration of treatment stated, either on phagocytic function and viability or on the clinical and biochemical parameters observed. PMID- 10516350 TI - Screening plasma aluminum levels in relation to aluminum bone disease among asymptomatic dialysis patients. AB - Aluminum accumulation in plasma and tissues is a well-described complication among persons undergoing peritoneal dialysis or hemodialysis. Excess bone aluminum is associated with low bone formation rates and increased risk for fractures. Current recommendations for care of patients with end-stage renal disease include screening for aluminum toxicity with plasma aluminum levels; patients with levels below 40 microg/L are considered to be at low risk for aluminum bone disease (ABD). We examined data from the Toronto Renal Osteodystrophy Study to evaluate the performance of plasma aluminum levels in screening for ABD. Two hundred fifty-eight unselected patients undergoing peritoneal dialysis (n = 143) or hemodialysis (n = 115) underwent diagnostic bone biopsy and measurement of plasma aluminum level. Sixty-nine patients (26.7%) were identified as having ABD, defined as low or normal bone formation rates with 25% or more bone surface aluminum staining. Plasma aluminum level was strongly associated with the presence of ABD; the odds ratio was 1.4 for each increase of 10 microg/L (95%CI, 1.2, 1.6). However, only 50.1% of patients with a plasma aluminum level of 40 microg/L or greater had ABD, whereas 14.2% of patients with a level below this threshold also had ABD. Using this cutoff level of 40 microg/L, the sensitivity and specificity were 65.2% and 76.7%, respectively. We conclude that although there is a correlation between high aluminum levels and ABD, a patient's plasma aluminum level does not predict well the presence of ABD in spite of a relatively high prevalence of disease. PMID- 10516351 TI - Significance of serum creatinine values in new end-stage renal disease patients. AB - The appropriate use of serum creatinine level as a surrogate for time in the course of renal failure when dialysis commences requires it to be a significant predictor of mortality in incident patients with end-stage renal disease (ESRD). This study evaluated factors that account for variations in creatinine level before the initiation of dialysis and whether incident creatinine level after controlling for these factors was a risk factor for mortality. This is a retrospective cohort study of patients from Maryland and Virginia who initiated dialysis between April 1, 1995, and December 31, 1996, with data ascertained from the Health Care Financing Administration Form 2728. Multivariate models were used to evaluate both the factors that predict incident serum creatinine level and the association between creatinine level and mortality. There were 5, 388 patients followed up for an average of 23.6 +/- 0.2 months. Mean creatinine level was 9.2 +/- 0.1 mg/dL, with case-mix factors most predictive of serum creatinine level and accounting for 9% of its variance. Hematocrit and blood urea nitrogen levels as additional surrogates for progression of renal disease accounted for 7.4% of the variance, whereas the nutritional parameters, body mass index, and albumin level only explained an additional 1% of the total variance in creatinine level. Creatinine level was inversely correlated with mortality risk, and this relationship was sustained both with transformation into an estimated glomerular filtration rate and multivariate adjustment for confounders (relative risk = 0. 96; P < 0.0001). Creatinine values from an incident ESRD population have a weak relationship with the timing of dialysis initiation but represent a strong measure of health status. PMID- 10516352 TI - Changes in venous histology in chronic hemodialysis patients. AB - Because most hemodialysis access fails at the venous side, we studied samples of brachial vein obtained during access creation in 15 patients with end-stage renal disease who gave consent. Veins were examined by computer-assisted histomorphometry, and the results correlated with the patients' clinical data. The mean venous medial width was 239 +/- 31 microm, and mean intimal width was 6.0 +/- 0.9 microm. Mean venous medial width was 358 +/- 74 microm and mean venous intimal width was 9.2 +/- 1.2 microm in the 4 patients who had been undergoing dialysis more than 6 months, compared with 196 +/- 23 microm and 4.9 +/- 0.8 microm, respectively, in the 11 patients undergoing dialysis less than 6 months (P < 0.01). The number of months undergoing hemodialysis correlated well with venous medial width (r = 0.79; P < 0.001). Correlation between number of months undergoing dialysis and intimal width did not reach statistical significance. Medial and intimal widths of the 4 patients with diabetes were not significantly different from those of the patients without diabetes. Serum parathyroid hormone level did not correlate with either medial or intimal venous width. We conclude there may be changes in the veins of hemodialysis patients with time that cause thickening of layers, even in veins not directly used for access. This may affect the creation or survival of subsequent vascular accesses. PMID- 10516353 TI - Racial disparities in renal transplant outcomes. AB - The purpose of our study was to evaluate the association of race and ethnicity with outcomes in the living related donor (LRD) renal transplant population, using multivariable adjustment for potential confounding variables. We prospectively analyzed 14,617 patients from the UNOS Renal Transplant Registry who underwent LRD renal transplantations in the United States between January 1, 1988 and December 31, 1996 using the Cox proportional hazards model. This model adjusts for the effects of potential genetic, social, and demographic confounding variables that may be associated with race or ethnicity long-term graft survival. Blacks were 1.8 times as likely as whites (P < 0.01, RR = 1.77) to suffer graft failure during the 9-year study period, which decreased minimally to 1.7 (P < 0.01, RR = 1.65) after controlling for potential confounding variables. Neither genotypic nor phenotypic HLA matching improved outcomes in blacks. Black renal transplant recipients had lower graft survival even after adjustment for matching and rejection, suggesting that non-HLA or socioeconomic mechanisms may contribute to racial differences in transplantation outcomes. PMID- 10516354 TI - Racial differences in survival in an urban peritoneal dialysis program. AB - We retrospectively evaluated 233 incident patients (61% black, 27% white, and 12% Hispanic/Asian) to our peritoneal dialysis (PD) program from January 1987 to September 1997 to identify any possible racial differences in patient survival. Information collected included clinical features, comorbid conditions, nutritional status, and dialysis dose at initiation of dialysis. The average age was 52 +/- 16 (SD) years, and 49% were men. Diabetes mellitus was present in 41% of patients. Overall follow-up was 31 +/- 24 (median 26) months during which time 21% of patients underwent transplant, 29% of patients transferred to hemodialysis (HD), and 42% of patients died. The Cox proportional hazards analysis, based on intent-to-treat, identified age (RR: 1.03), race (RR: 2.35, white versus black), cardiac disease (RR: 1.97), and serum albumin (RR: 0. 44) to independently predict mortality. Further analysis was performed based on diabetic status, and the analysis identified age (RR: 1.06), race (RR: 2.45, white versus black), and peripheral vascular disease (RR: 2.88) as predictors of mortality in diabetic patients. In nondiabetic patients, age (RR: 1.03), race (RR: 2.24, white versus black), cardiac disease (RR: 2.48), cerebrovascular disease (RR: 3.17), and serum albumin (RR: 0.39) were significant predictors of mortality. The significance of race persisted even after adjusting patients transferring to hemodialysis. The adjusted patient survival at 1, 2, and 5 years was 94%, 87% and 53% for black patients, and 86%, 72%, and 23% for white patients. The adjusted patient survival in diabetics at 1, 2, and 5 years was 92%, 79%, and 37% for black patients, and 82%, 56%, and 9% for white patients. The adjusted patient survival in nondiabetics at 1, 2, and 5 years was 94%, 91%, and 63% for black patients, and 88%, 82%, and 35% for white patients. In conclusion, long-term patient survival is better for black patients than white patients in our peritoneal dialysis program. Peritoneal dialysis should be considered a viable dialytic option for black patients entering an end-stage renal disease program. PMID- 10516355 TI - Racial/ethnic analysis of selected intermediate outcomes for hemodialysis patients: results from the 1997 ESRD Core Indicators Project. AB - Principal goals of the End-Stage Renal Disease (ESRD) Core Indicators Project are to improve the care provided to ESRD patients and to identify categorical variability in intermediate outcomes of dialysis care. The purpose of the current analysis is to extend our observations about the variability of intermediate outcomes of ESRD care among different racial and gender groups to a previously unreported group, Hispanic Americans. This group is a significant and growing minority segment of the ESRD population. A random sample of Medicare-eligible adult, in-center, hemodialysis patients was selected and stratified from an end of-year ESRD patient census for 1996. Of the 6,858 patients in the final sample, 45% were non-Hispanic whites, 36% were non-Hispanic blacks, and 11% were Hispanic. Whites were older than blacks or Hispanics (P < 0.001). Hispanics were more likely to have diabetes mellitus as a primary diagnosis than either blacks or whites (P < 0.001). Even though they received longer hemodialysis times and were treated with high-flux hemodialyzers, blacks had significantly lower hemodialysis doses than white or Hispanic patients (P < 0.001). The intradialytic weight losses were greater for blacks (P < 0.05). The delivered hemodialysis dose was lower for blacks than for whites or Hispanics whether measured as a urea reduction ratio (URR) or as the Kt/V calculated by the second generation formula of Daugirdas (median 1. 32, 1.36, and 1.37, respectively, P < 0.001). Hispanics and whites had modestly higher hematocrits than blacks (33.2, 33.2, and 33.0%, respectively, P < 0.01). There was no significant difference among groups in the weekly prescribed epoetin alfa dose ( approximately 172 units/kg/week). A significantly greater proportion of Hispanic patients had transferrin saturations >/=20% compared with the other two groups (P < 0.001). Logistic regression modeling revealed that whites were significantly more likely to have serum albumin <3. 5(BCG)/3.2(BCP) gm/dL (OR 1.4, p < 0.01); blacks were significantly more likely to have a delivered Kt/V < 1.2 (OR 1.4, P < 0.001) and hematocrit <30%, (OR 1.2; P < 0.05) and both blacks and Hispanics were significantly more likely to have a delivered URR < 65% (OR 1.5, P < 0.001 and 1.2, P < 0.05, respectively). PMID- 10516356 TI - Focal segmental glomerulosclerosis in renal allografts with chronic nephropathy: implications for graft survival. AB - De novo focal segmental glomerulosclerosis (FSGS) in renal allografts most often is diagnosed in association with chronic allograft nephropathy (CN). In this study, we assessed the clinical and pathological variables that correlate with the presence of de novo FSGS and the implications of FSGS for the survival of grafts with CN. The study population included 293 renal allograft recipients (52 living related donor, 241 cadaveric donor) diagnosed with CN by biopsy more than 6 months after transplantation. Patients with recurrent FSGS or FSGS associated with other glomerulopathies were excluded. FSGS was present in 87 patients with CN (30%). FSGS was diagnosed more commonly in the following groups of patients: young (P = 0.04), black (P = 0.02), and those with elevated serum cholesterol levels (P = 0.002) and/or high-grade proteinuria (P < 0. 0001, all univariate analysis). FSGS was diagnosed later after transplantation than CN without FSGS (P < 0.0001), and FSGS correlated with the presence of arteriolar hyalinosis in the biopsy specimen (P = 0.04). Compared with CN without FSGS, FSGS was associated with significantly worse death-censored graft survival (P = 0.008, univariate Cox). In addition, when we analyzed all patients with CN, graft survival correlated by multivariate analysis with the following parameters: serum creatinine level (P < 0.0001) and proteinuria (P = 0.004) at the time of diagnosis, presence of FSGS (P = 0.03), and degree of interstitial fibrosis and tubular atrophy (CN score; P < 0.0001, Cox). Of interest, the use of lipid reducing agents was also associated with improved graft survival in patients with CN (P = 0.0002, univariate Cox), although total lipid levels were not significantly less in patients receiving these drugs. In conclusion, de novo FSGS presents late after transplantation and in association with arteriolar hyalinosis, suggesting these lesions may be related to chronic cyclosporine toxicity. In CN, the presence of FSGS and the severity of interstitial fibrosis are negative independent predictors of graft survival. The possible relationship between lipid-reducing agents and graft survival clearly needs to be examined carefully in future studies. PMID- 10516357 TI - Reversal of sleep apnea hypopnea syndrome in end-stage renal disease after kidney transplantation. AB - Sleep apnea hypopnea syndrome (SAHS) is extremely common in patients with end stage renal disease (ESRD). Although the underlying mechanisms linking these 2 conditions remain to be better defined, it is likely that multiple factors are involved. We report an individual with ESRD with severe SAHS that resolved after kidney transplantation. The improvement in SAHS paralleling the effective treatment of ESRD suggests the pathogenesis involves an unstable breathing pattern, possibly caused by an altered metabolic state, uremia, and changes in volume status. The possibility that elevations in cytokine levels could be involved also is discussed and deserves further attention. PMID- 10516358 TI - Hypocomplementemic urticarial vasculitis or systemic lupus erythematosus? AB - The 2 patients presented here showed the typical signs of hypocomplementemic urticarial vasculitis syndrome (HUVS). During follow-up, there was an inverse correlation between anti-C1q autoantibody titer and C1q antigen concentration in serum in both patients over a period of 2 years. The first patient had nephritis characterized by immune deposits in glomeruli and around the tubules. The histological findings, C1q deposits, and presence of tubuloreticular inclusions in capillary endothelial cells suggested a disease process identical to systemic lupus erythematosus (SLE). The second patient, after a lag phase of 2 years, fulfilled a fourth American College of Rheumatology criteria for SLE when she developed anti-double-stranded DNA antibodies. HUVS and SLE overlap, and the criteria for identifying HUVS as an entity distinct from SLE are lacking. PMID- 10516359 TI - Bilateral renal infarction secondary to paradoxical embolism. AB - Paradoxical embolism is an uncommon but increasingly reported cause of arterial embolic events. Involvement of the kidney is rarely reported. Autopsy studies suggest, however, that embolic renal infarction is underdiagnosed antemortem. We report a case of bilateral, main renal artery occlusion and acute renal failure secondary to paradoxical embolism. Clinical and laboratory data at presentation were not suggestive of renal infarction. Support for the diagnosis of paradoxical embolism, which most commonly occurs across a patent foramen ovale, was made by contrast echocardiography, which provides a sensitive method for detecting right to-left intracardiac shunts. The often subtle presentation of renal infarction suggests patients with peripheral or central arterial embolic events should be carefully observed for occult renal involvement. Contrast echocardiography should be performed when renal infarction occurs without a clear embolic source to evaluate for paradoxical embolism. PMID- 10516360 TI - Kidney transplantation: racial or socioeconomic disparities? PMID- 10516361 TI - When the doctor's master is a bully. PMID- 10516362 TI - Chemokines as therapeutic targets in renal inflammation. PMID- 10516364 TI - A 19-year-old man with hypertension, proteinuria, and renal insufficiency. PMID- 10516365 TI - What is our duty to a "hateful" patient? Differing approaches to a disruptive dialysis patient. PMID- 10516366 TI - Interventional radiologists important to vascular access management. PMID- 10516367 TI - Renal osteodystrophy in Iberoamerica. PMID- 10516368 TI - Optimizing the use of parenteral iron in end-stage renal disease patients: focus on issues of infection and cardiovascular disease. Introduction. PMID- 10516369 TI - National perspective on iron therapy as a clinical performance measure for maintenance hemodialysis patients. AB - The Health Care Financing Administration (HCFA) End-Stage Renal Disease (ESRD) Core Indicators Project collects clinical information on prevalent adult patients receiving in-center hemodialysis care in the United States to assess the quality of care delivered. Although hematocrit values, transferrin saturations (TSATs) and iron prescription practices have improved over the last 5 years, we sought to determine whether there are continued opportunities for improvement of this domain of care. A random sample of 7,292 adult in-center hemodialysis patients was selected for the period October through December 1996. Hematocrit values, TSATs, serum ferritin concentrations, epoetin-alfa dosing, and iron prescriptions were abstracted from 4,991 patient medical records to assess anemia management practices. The mean hematocrit for this cohort was 32.6% +/- 3.5%, and 72% of patients had hematocrit values greater than 30%. Ninety-four percent of patients received epoetin alfa intravenously, with a mean weekly epoetin dose of 202.4 +/- 137.2 U/kg. The mean TSAT was 27.4% +/- 12.6%; 73% of patients had TSATs >/= 20%. The mean serum ferritin concentration was 386 +/- 422 ng/mL; 79% and 12% of patients had serum ferritin concentrations greater than 100 ng/mL and greater than 800 ng/mL, respectively. Nine percent of the sample had TSATs less than 20% and serum ferritin concentrations less than 100 ng/mL. Regardless of the TSAT, approximately three fourths of patients received iron; only about half received IV iron. Of the subset of patients with TSATs less than 20% and serum ferritin concentration less than 800 ng/mL, only 53% were prescribed IV iron. Although substantial improvements have been made in anemia management in hemodialysis patients over the last 5 years, significant opportunities persist to improve iron prescription practices. PMID- 10516370 TI - Iron status as measured by serum ferritin: the marker and its limitations. AB - Assessment of iron status is important, because iron deficiency and overload have pathologic consequences. Serum ferritin, the function of which is unknown, is frequently used to assess labile iron stores for the purpose of ensuring their adequacy for erythropoiesis. However, measurable ferritin levels can be increased when tissue ferritin is released during cellular injury, and erythropoietic blockade can increase the labile iron pool, elevating serum ferritin levels despite suppressed erythropoiesis. Erythropoietin-stimulated red blood cell (RBC) production can quickly decrease the labile cellular iron pool and reduce serum ferritin, unless supplemental iron is supplied. A serum ferritin level less than 12 microg/L indicates that there is no iron in the stores (absolute iron deficiency), and levels above 15 microg/L may still not be sufficient to meet erythropoietic demand. This is particularly true in patients receiving erythropoietin, in which the stimulated erythropoiesis requires extra iron supplies. Because of the limitations of serum ferritin measurements and questions regarding the effects of free iron, clinicians need not be too alarmed by high serum ferritin levels. At the very least, safety concerns must be balanced against the real need for iron supplementation to maintain adequate erythropoiesis. PMID- 10516371 TI - Iron and cardiac disease in the end-stage renal disease setting. AB - Erythropoietin (EPO) therapy and appropriate iron administration are important aspects for managing the anemia of end-stage renal disease (ESRD). Achieving target hemoglobin levels of 11 to 12 g/dL and optimizing iron balance should improve clinical outcomes and increase patient quality of life. However, concerns have been raised about parenteral iron supplementation leading to excessively high iron levels, which may induce increased oxidative stress and risk for cardiovascular disease. Increased oxidative stress is often already present in patients with chronic renal disease and in patients with ESRD undergoing hemodialysis. The "iron hypothesis" proposes that excess iron is associated with increased risk for cardiac disease. While some studies have found an association between high iron levels or increased iron consumption with elevated risk for cardiac disease in subjects without renal disease, others have not found this association. Indeed, several studies suggest that achievement of target hematocrit levels in ESRD patients improves several clinical outcomes and that anemia itself is a risk factor for cardiac disease. Well-designed prospective studies are needed before the relationship between supplemental iron administration, excess iron, and cardiac disease can be firmly established. PMID- 10516372 TI - Pathobiology of the role of iron in infection. AB - Beyond its role in hemoglobin synthesis, iron plays an important part in host defense mechanisms. Sequestration of iron is a mechanism to control bacterial proliferation and virulence. However, uremia results in several immunologic deficits, including impaired lymphocyte mitogenic response and granulocyte function abnormalities such as impaired phagocytosis, respiratory burst, and myeloperoxidase activity. At the other end of the spectrum, iron overload can impair immune function and stimulate bacterial growth and virulence. Overtreatment with iron increases the preexisting risk of infectious complications for uremic patients, as indicated by hyperferritinemia in hemodialysis patients who have impaired polymorphonuclear leukocyte (PMNL) induced bacterial killing. These results suggest that maintaining iron status within normal range is important to control potential increased risk of infection in patients with end-stage renal disease (ESRD). PMID- 10516373 TI - Iron and infection: clinical experience. AB - Bacterial infection is a significant cause of morbidity and mortality in hemodialysis patients, and a number of studies have implicated iron overload as a risk factor for bacterial infection in these patients. While the underlying cause of increased susceptibility to bacterial infection is not completely understood, evidence suggests that iron overload alters the chemotactic and phagocytic properties of neutrophils, thereby reducing their ability to kill invading pathogens. T-cell function also appears to be altered. In addition, high levels of serum iron may promote replication and dissemination of bacterial pathogens that use iron as a growth factor. With the introduction of recombinant human erythropoietin therapy for hemodialysis patients, the need for red blood cell transfusions has been reduced and iron overload occurs much less frequently. Several recent studies have indicated that iron overload, which is suspected in the presence of high serum ferritin levels, may no longer be a significant risk factor for infection in hemodialysis patients receiving erythropoietin therapy. Major risk factors for infection in these patients include history of bacterial infection, immunosuppressive therapy, and vascular access via catheters rather than by arteriovenous fistula. In addition, anemia has recently been linked to an increased incidence of bacterial infection, particularly in patients receiving erythropoietin therapy. Therefore, repletion of iron stores and maintenance of iron balance without iron overload may prove to be important factors in reducing the incidence of bacterial infections in hemodialysis patients. However, the relationships between iron levels, anemia, and susceptibility to bacterial infection require further investigation. PMID- 10516374 TI - The systemic inflammatory response and its impact on iron nutriture in end-stage renal disease. AB - In most chronic disease conditions, the systemic inflammatory response and its mediators play an essential pathogenic role. Protein calorie malnutrition, a prominent feature of end-stage renal disease (ESRD), also develops, largely as a consequence of the systemic inflammatory response. ESRD (uremia), dialysis, systemic metabolic acidosis, and infections activate the systemic inflammatory response. Elevations in C-reactive protein and depressions of serum albumin below 4 g/dL are found in more than 50% of ESRD patients undergoing dialysis. In many patients receiving dialysis, the impact of this acute-phase response on measures of iron metabolism limits the ability to diagnose iron deficiency. Furthermore, there are risks to iron administration, although data linking iron overload to risk of infection in dialysis patients is suggestive, not definitive. It seems reasonable to hypothesize that the greatest risk of iron administration is in patents who are already infected, and the greater risk would be to raise the serum iron level and transferrin saturation precipitously. The total-dose infusion method, which provides all iron required to correct deficiency in 1 dose, is more likely to produce side effects and rapidly raise serum iron levels and transferrin saturation. The use of low-dose intravenous iron supplementation (10 to 20 mg per dialysis treatment or 100 mg every second week) avoids iron overtreatment and should reduce adverse events. In ESRD patients receiving dialysis, the importance of the systemic inflammatory response in the development of protein calorie malnutrition, the impact of the acute-phase response on iron nutriture, and the response to erythropoietin therapy must be considered to achieve an understanding of the altered responses to nutritional therapy in this setting. PMID- 10516375 TI - Beneficial effects of adopting an aggressive intravenous iron policy in a hemodialysis unit. AB - Iron deficiency is the most common cause of a suboptimal response to epoetin therapy, and the treatment of choice is intravenous (IV) iron. It is also increasingly recognized that IV iron can enhance the response to epoetin, even in iron-replete patients. The aim of the present study was to examine the effects of adopting an aggressive IV iron policy in all patients attending a single-center hemodialysis unit. The protocol was simple and practical, and involved administering a weekly IV bolus of 100 mg iron sucrose to all patients with a serum ferritin level of 150 to 1,000 microg/L. Only patients with a serum ferritin level greater than 1,000 microg/L were excluded; patients with a serum ferritin level below 150 microg/L were given a more aggressive IV dosing regimen to get into range for the standard protocol. Among 116 patients included in the study, the mean serum ferritin level increased from 214 microg/L in November 1997 to 564 microg/L in November 1998 (P < 0.0001). Mean hemoglobin increased modestly from 9.6 g/dL to 10.7 g/dL over the same period, but there was a dramatic reduction in mean epoetin dose from 13,277 U/wk to 8,976 U/wk (P < 0.0005), resulting in cost savings of approximately pound 228,000 ($366,000), or pound 152 ($244) per patient per month. No adverse reactions to IV iron were seen among a total of 4,564 injections, and there was no obvious increase in the incidence of infection. This simple, practical IV iron dosing policy resulted in dramatic savings in epoetin dosage and cost with no significant adverse effects. PMID- 10516376 TI - Review of issues relating to iron and infection. AB - Use of erythropoietin (EPO) therapy and iron supplementation has improved the management of anemia in patients with end-stage renal disease (ESRD). As more patients receive supplemental iron, however, concerns are being raised about a potential link between iron and infection. There is biologic plausibility for this link, since iron is a growth factor for bacteria and certain host defense mechanisms are iron-sensitive. Animal models show that injection of iron leads to increased susceptibility to bacterial infection. In some studies, patients with high serum ferritin levels have reduced neutrophil function. However, these studies did not determine whether serum ferritin levels were elevated because of increased iron stores or because of infection. If infection is present, it might cause both the elevated serum ferritin levels and the neutrophil dysfunction. Several clinical studies have found an association between high serum ferritin levels and increased infectious risk. In studies that control for important covariates such as use of catheters and previous infections, the infectious risk associated with iron administration or elevated serum ferritin levels is reduced or eliminated. Collectively, these studies suggest that our current understanding of the relationship between iron and infection is incomplete and further studies are needed. There is no reason to alter current iron treatment strategies based on this literature. PMID- 10516377 TI - Introduction PMID- 10516378 TI - Staging and prognostic factors in soft tissue sarcoma. AB - Significant advances have been made in the understanding of clinicopathologic prognostic factors for soft tissue sarcoma over the past decade. Foremost among these advances is an improved ability to recognize the subset of patients at high risk for recurrent disease based on clinicopathologic data available at the time of initial presentation. Progress has also helped to elucidate specific molecular factors that have independent prognostic significance. This review outlines the updated American Joint Committee on Cancer staging system for soft tissue sarcoma and summarizes the available data on traditional clinicopathologic and molecular prognostic factors. PMID- 10516379 TI - Current aspects of the pathology of soft tissue sarcomas. AB - In soft tissue sarcomas, advances in pathological techniques, including immunohistochemistry, cytogenetics, and molecular genetics, have improved diagnostic accuracy, confirmed or clarified interrelationships between tumor subtypes, and revealed mechanisms of tumorigenesis. Many sarcomas are associated with abnormalities of tumor-suppressor genes, and several types have been found to have specific chromosomal translocations. These data and correlative clinicopathologic studies, although confirming many traditional pathological views, enable refinement or reassessment of terminology and classification of some small round cell, spindle cell, pleomorphic, and lipogenic tumors. New factors are emerging for prediction of tumor behavior, which might ultimately relate to therapy once a wider range of treatment options becomes available. This article reviews these current aspects of sarcoma pathology. PMID- 10516381 TI - Evidence for using adjuvant chemotherapy as standard treatment of soft tissue sarcoma. AB - The role of adjuvant chemotherapy in the treatment of patients with soft-tissue sarcomas remains controversial. The initial observation that extremity tumors might benefit more than tumors of other primary sites prompted careful review of published data. Eight of 9 studies showed improved disease-free and overall survival for the adjuvant chemotherapy group. A meta-analysis of published data showed statistically significant improvement in both disease-free survival and overall survival for adjuvant chemotherapy patients. This analysis prompted a more accurate meta-analysis of individual patient data, which confirmed the results of the meta-analysis of published data. These positive results reflect the efficacy of the chemotherapy of the 1970s and 1980s. The only study of contemporary adjuvant chemotherapy, reported by the Italian Cooperative Group, showed a statistically significant advantage in both disease-free survival and overall survival for patients treated with adjuvant chemotherapy. The studies, taken as a whole, serve as proof of principle that chemotherapy, given early to patients with primary soft-tissue sarcomas of the extremities who are at high risk of developing local recurrence or metastatic disease, can delay and/or prevent relapse and improve cure rate. PMID- 10516380 TI - The local management of soft tissue sarcoma. AB - Soft tissue sarcomas (STS) are rare tumors arising from the connective tissues. STS can arise at any anatomic site, can demonstrate varied behavior and prognosis, and therefore present a formidable challenge in management. The local treatment of STS demands technical complexity in the application of diagnostic tools, including pathology and imaging, as well as treatment approaches, including surgical ablation and reconstruction, radiotherapy, and, in defined cases, chemotherapy. The understanding of the management of these lesions is profoundly dependent on the multidisciplinary setting, where experience has been gained and skills are available to increase the likelihood of a successful result. Several proven options are available for optimal local management, and the choice of approach depends on the prevailing practice and resource profile of the treating center. With modern approaches, the local control rate can be expected to be at least 90% for extremity lesions, which constitute the most common STS. The experience in other anatomic sites is less favorable as a result of a combination of late diagnosis, technically difficult access sites, and possibly less familiarity with these less common presentations. The disappointing results make it all the more important for patients to be referred to a multidisciplinary setting with experience in sarcoma management to maximize the chance of successful local outcome. PMID- 10516382 TI - The reason for confining the use of adjuvant chemotherapy in soft tissue sarcoma to the investigational setting. AB - Adjuvant chemotherapy for soft tissue sarcoma has been the subject of many studies. Most studies have been nonrandomized and therefore by definition inconclusive. In addition, most of the 14 performed randomized studies have had small sample sizes. A meta-analysis on the data of these 14 studies suggested a possible survival improvement, albeit not significant. The outcome of the meta analysis should be interpreted with caution in view of the fact that in 18% of patients, histology data were lacking; ineligibility rates in included studies were high; central pathology was not uniformly performed; and in 6% of patients, sarcomas other than soft tissue sarcomas were included. Because it is generally recommended that meta-analyses of small randomized trials are used only to generate hypotheses for more reliable randomized trials, it is argued that adjuvant chemotherapy in soft tissue sarcomas should remain confined to the investigational setting. PMID- 10516383 TI - Functional outcome in extremity soft tissue sarcoma. AB - The studies reporting functional outcome for patients undergoing limb preservation surgery for extremity soft tissue sarcoma (STS) have evaluated mainly impairments, that is, deficits at an anatomic structure level, such as joint range of motion and strength. Disability, activities of daily living, self care, and mobility have been less frequently evaluated. Review of the literature suggests that approximately 50% of patients treated for STS have significant impairments, whereas the frequency of disability is less. Synthesis of the results is difficult because of the heterogeneity of patient samples, treatment, and the outcomes used to evaluate function. Future studies require the use of standardized definitions and reliable and valid functional outcome measures. Improved patient outcomes can be achieved only by understanding the determinants of these outcomes and by introducing interventions to improve patient functional outcome. PMID- 10516384 TI - The biological significance of failure at the primary site on ultimate survival in soft tissue sarcoma. AB - Local tumor recurrence after complete resection may be due to treatment factors or represent a manifestation of tumor biology. The association of local tumor recurrence, distant metastases, and death in patients undergoing treatment for extremity soft tissue sarcoma (STS) has been described but continues to be enigmatic. After definitive multimodality treatment for extremity STS, local tumor recurrence is associated with development of distant metastasis, and metastases are implicated in subsequent disease-specific death. The relationship is an enigma, and the causality is unclear. Conversely, for patients with retroperitoneal STS, a direct relationship between local tumor recurrence and disease-specific death has been shown. In this article, current concepts are analyzed and reviewed. PMID- 10516385 TI - Approaches to local salvage of soft tissue sarcoma after primary site failure. AB - Improvements in pretreatment evaluation and the wider application of specialized multidisciplinary care have substantially reduced the risk of local recurrence for patients with soft tissue sarcomas arising at any site, and the recurrences that are still seen are often those that are most difficult to manage effectively. The management strategy for an isolated local recurrence of soft tissue sarcoma is usually similar to that used for primary disease. With appropriate pretreatment evaluation and salvage therapy that includes a multidisciplinary approach, most patients with local recurrence can expect local disease control and a good functional outcome. The development of effective management of a local recurrence is often a complex problem. The necessary decisions are influenced by the tumor location, disease extent, and previous local therapy. The need for specialized care is stressed. Patient evaluation, management strategies, and expected outcome for various clinical scenarios are discussed. PMID- 10516386 TI - The treatment of distant metastases in soft tissue sarcoma. AB - This article reviews the current standard approaches to the treatment of metastatic soft tissue sarcoma (STS) and evaluates new chemotherapy agents and novel approaches. A computerized search strategy was used to identify articles examining the role of chemotherapy and surgery in metastatic STS, which were published between January 1992 and December 1998. This search was supplemented by key articles from our files published before 1992. In selecting articles for inclusion in this review, emphasis was placed on randomized data and novel approaches. Only three agents-doxorubicin, ifosfamide, and dacarbazine-have shown significant activity in metastatic STS. Numerous studies have examined the efficacy and toxicity of combining the known active agents in standard doses or in high doses with cytokine support. Promising results, in terms of increased response rates, often have not been reproduced in randomized trials, and there is no convincing evidence of enhanced overall survival. New regimens should be evaluated in randomized trials incorporating quality-of-life endpoints. High-dose chemotherapy with bone marrow/stem cell rescue remains an investigational procedure of uncertain efficacy. Pilot studies have established the feasibility of intraperitoneal chemotherapy, after cytoreductive surgery, in patients with peritoneal sarcomatosis. To date, the efficacy of this approach has not been validated in phase II or III trials. The role of surgery in the treatment of isolated pulmonary metastases is well established. Results of small series raise the possibility that resection of hepatic metastases is beneficial in selected patients. Current chemotherapy options for patients with STS are limited. There is reason to hope that the situation will change with the further development of new agents that have novel and specific mechanisms of action. PMID- 10516388 TI - Structural recovery from sound and aminoglycoside damage in the avian cochlea. AB - Hair cell regeneration in the mature avian cochlea occurs in response to trauma that causes the death of some or all of the existing hair cell population. In general, this trauma has been introduced experimentally by either sound overexposure or treatment of the bird with high doses of aminoglycoside antibiotics. When injured hair cells are ejected from the sensory epithelium, the nonsensory supporting cells respond by re-entering the cell cycle and proliferating or by transdifferentiating directly into hair cells without a mitotic event. The new hair cells mature in a manner similar to that seen during embryonic development. They make connections with the overlying tectorial membrane and the afferent and efferent cochlear nerve processes within the sensory epithelium. This structural regeneration is accompanied by a significant recovery of auditory function and thus allows the animal to regain its hearing ability. This hair cell regeneration is presumably quite beneficial to birds, whose primary means of communication is based on vocalizations and the ability to hear and comprehend them. The prevalence of hearing loss in our society and the isolating impact it has on affected individuals makes the potential for finding ways to induce a similar hair cell regeneration in humans a very tempting goal. Studies of hair cell regeneration over the last 12 years have focused on the mechanisms that regulate the process and how they could be controlled. This review will examine the structural events involved in regenerating hair cells in the avian cochlea after sound damage and aminoglycoside treatment. It will define how hair cells and nerve endings are lost and the tectorial membrane is damaged by the traumatizing stimuli and how the supporting cells and nerve fibers respond by producing new hair cells, a new tectorial membrane and new synaptic connections during recovery. Finally, it will focus on mechanisms that control the proliferation and transdifferentiation of supporting cells and the differentiation of new hair cells. This structural review is accompanied by a companion review that covers the fundamental issues concerning functional recovery in the avian cochlea associated with hair cell regeneration. PMID- 10516389 TI - Functional recovery in the avian ear after hair cell regeneration. AB - Trauma to the inner ear in birds, due to acoustic overstimulation or ototoxic aminoglycosides, can lead to hair cell loss which is followed by regeneration of new hair cells. These processes are paralleled by hearing loss followed by significant functional recovery. After acoustic trauma, functional recovery is rapid and nearly complete. The early and major part of functional recovery after sound trauma occurs before regenerated hair cells become functional. Even very intense sound trauma causes loss of only a proportion of the hair cell population, mainly so-called short hair cells residing on the abneural mobile part of the avian basilar membrane. Uncoupling of the tectorial membrane from the hair cells during sound overexposure may serve as a protection mechanism. The rapid functional recovery after sound trauma appears not to be associated with regeneration of the lost hair cells, but with repair processes involving the surviving hair cells. Small residual functional deficits after recovery are most likely associated with the missing upper fibrous layer of the tectorial membrane which fails to regenerate after sound trauma. After aminoglycoside trauma, functional recovery is slower and parallels the structural regeneration more closely. Aminoglycosides cause damage to both types of hair cells, starting at the basal (high frequency) part of the basilar papilla. However, functional hearing loss and recovery also occur at lower frequencies, associated with areas of the papilla where hair cells survive. Functional recovery in these low frequency areas is complete, whereas functional recovery in high frequency areas with complete hair cell loss is incomplete, despite regeneration of the hair cells. Permanent residual functional deficits remain. This indicates that in low frequency regions functional recovery after aminoglycosides involves repair of nonlethal injury to hair cells and/or hair cell-neural synapses. In the high frequency regions functional recovery involves regenerated hair cells. The permanent functional deficits after the regeneration process in these areas are most likely associated with functional deficits in the regenerated hair cells or shortcomings in the synaptic reconnections of nerve fibers with the regenerated hair cells. In conclusion, the avian inner ear appears to be much more resistant to trauma than the mammalian ear and possesses a considerable capacity for functional recovery based on repair processes along with its capacity to regenerate hair cells. The functional recovery in areas with regenerated hair cells is considerable but incomplete. PMID- 10516390 TI - Pre-attentive discriminability of sound order as a function of tone duration and interstimulus interval: a mismatch negativity study. AB - The present study addressed the pre-attentive processing of sound order. Event related potentials were recorded from reading subjects while they were presented with pairs of two tones differing from each other in frequency (1000 vs. 1500 Hz). The within-pair (silent) interstimulus interval (ISI) was, in separate blocks, varied between 0 and 245 ms to determine the minimum separation in time needed for detecting the reversed order of the two frequencies. In standard tone pairs (p = 0.9), the frequencies were in an ascending order, whereas in the deviant pairs (p = 0.1), their order was reversed. Tone durations of 5 and 20 ms were employed in separate experiments. With the 20-ms stimulus duration, the change-specific mismatch negativity (MMN) component was elicited with all within pair ISIs employed (0, 10, 30, 90 ms). With the 5-ms stimulus duration, however, MMN was elicited only with the 245-ms ISI but not with 95-ms or shorter ISIs. These results show that increased stimulus duration considerably improves perceiving the order of two tones at the pre-attentive level. They also indicate that the accuracy of the processing of temporal information can be probed with MMN. This finding, together with the fact that MMN elicitation does not require the subject's voluntary attention, suggests that MMN might be used in the assessment of temporal processing deficits in clinical disorders in which patients are not motivated or able to give their verbal or motor response. PMID- 10516391 TI - The human olivocochlear system: organization and development. AB - The goals of the present study were to identify olivocochlear neurons in the human brainstem, to establish the time course of their early development and to compare the organization of the human olivocochlear system to that of other mammals. To accomplish these goals, we used immunohistochemistry for choline acetyltransferase (ChAT) and calcitonin gene-related peptide (CGRP) in postmortem brainstems of human subjects ranging in age from 16 fetal weeks to 17 years. By immunostaining, we identified two classes of cells in the superior olivary complex: both classes were seen to be present from the twenty-first fetal week to the seventeenth year. Neurons which are immunostained only for ChAT are located primarily in the dorsomedial, ventral and ventrolateral sectors of the periolivary region. These neurons are predominantly bipolar or multipolar cells, and are probably homologous to medial olivocochlear neurons in other species. A second population of cells is immunoreactive for both ChAT and CGRP. This population includes a cluster of mostly small oval neurons, located on the dorsal edge of the olivary complex, and a variable number of cells found along the margin of the lateral olivary nucleus. These ChAT- and CGRP-immunoreactive cells are likely to be homologous to the lateral olivocochlear system in other mammals. With increasing age, the dorsal cluster of small cells shifts from its original cap-like position over the lateral olivary nucleus to become an extended column of cells lying among the fibers of the olivocochlear bundle. PMID- 10516392 TI - Indomethacin treatment decreases renal blood flow velocity in human neonates. AB - This study was designed to evaluate the effect of indomethacin (ID) on renal perfusion in 13 neonates with symptomatic patent ductus arteriosus (PDA). Serial blood flow velocity in the left renal artery was measured just before and at 10, 30, 45, 60, 75 and 90 min after ID administration. Serum creatinine (Cr), sodium (Na), and osmolarity were measured just before, at 12 and 24 h, and at 3 days after ID administration. Timed urine also was collected for measurement of amount, fractional excretion of Na (FE(Na)), and creatinine clearance (C(Cr)). ID decreased end-diastolic flow velocity of renal artery and increased Pourcelot's index, starting at 10 min and lasting for 75 min (p < 0.05). Serum Cr significantly increased at 12 h, and hourly urine output and C(Cr) decreased for 24 h. Serum Na and osmolarity decreased for a period of at least 3 days (p < 0.05). FE(Na) decreased at 12-24 h (p < 0.05). We conclude that ID treatment can induce significant renal dysfunction due to diminution of renal perfusion in human neonates. PMID- 10516393 TI - Vascular endothelial growth factor in pulmonary lavage fluid from premature infants: effects of age and postnatal dexamethasone. AB - Corticosteroids are used to ameliorate bronchopulmonary dysplasia (BPD). They also affect normal development, including the expression of growth factors such as vascular endothelial growth factor (VEGF). Deep pulmonary lavage specimens were collected on days 1, 3, 7 and 28 of life in 40 infants of <34 weeks of gestation at birth during a randomized controlled trial of two doses of dexamethasone (DEX) at 12 and 24 h of age for BPD prophylaxis. VEGF was measured by enzyme-linked immunosorbent assay. The 18 DEX and 21 control subjects had similar gestations, birth weights and oxygen requirements at study entry. Lavage VEGF tripled between day 1 and 3 in both groups. The day 7 levels were higher in DEX subjects than in controls. DEX and control values were similar on day 28. Higher lavage VEGF levels on days 1 and 3 were also correlated with lower gestational age at birth. Lavage VEGF levels were not associated with the development of BPD. We speculate that these DEX- and age-associated changes in VEGF may affect pulmonary angiogenesis. PMID- 10516394 TI - Associations of IGF-I, IGFBP-1 and IGFBP-3 on intrauterine growth and early catch up growth. AB - Fetal cord blood IGF-I, IGFBP-1 and IGFBP- 3 levels of appropriate-for gestational-age (AGA) and intrauterine growth retardation (IUGR) babies are studied and followed up for 6-9 months, reevaluated for anthropometric measures and the effects of IGF-I, IGFBP-1 and IGFBP-3 on fetal growth and early catch-up growth is investigated. 23 AGA and 21 IUGR babies, totally 44 newborns, were included in the study protocol. IGF-I and IGFBP-3 levels were found to be high in AGAs with respect to IUGR babies and IGFBP-1 is found to be high in IUGR with respect to AGAs. IGF-I was significantly lower in IUGR babies without catch-up growth (group 2b) with respect to AGAs (group 1) and neonates with IUGR and catch up growth (group 2a) and group 2a infants had higher IGF-I values than group 2b infants (p < 0.05). IGFBP-3 levels in group 1 were significantly higher than in the other two groups (p < 0.05), but not significantly different in group 2a with respect to group 2b (p > 0. 05). IGFBP-1 values showed no statistically significant difference with respect to the three different groups (p > 0.05). A good correlation was found between birth weight, postnatal weight and postnatal height and IGF-I and IGFBP-3 levels (p < 0.05) but not with IGFBP-1 levels. Aside from the height of the 3 groups of infants which were similar to each other after the follow-up period, IGF-I was significantly high in IUGR infants with catch-up growth with respect to IUGR infants without catch-up growth, indicating its importance in early catch-up growth of IUGR babies. PMID- 10516395 TI - Early (4-7 days of age) dexamethasone therapy for prevention of chronic lung disease in preterm infants. AB - We conducted a comparative study to evaluate whether early (4-7 days of age) low dose dexamethasone (DEX) therapy in preterm infants with surfactant-pretreated respiratory distress syndrome (RDS) would facilitate extubation and improve the clinical outcome. Twenty-six preterm infants with surfactant-pretreated RDS who were oxygen- and ventilator-dependent at 4 days of postnatal age were enrolled. Twelve infants were in the historical comparison group, and 14 infants were assigned to receive DEX 0.125 mg/kg i.v., every 12 h, for a total of 6 doses. At study entry, the two groups had a comparable clinical status. DEX therapy significantly facilitated weaning from mechanical ventilation (median interval, 6 vs. 24 days, p < 0.005) and shortened duration of oxygen supplementation (9 vs. 28 days, p < 0.05) as compared with the historical comparison group. At 28 days of age, the occurrence of chronic lung disease (CLD) was significantly lower (1/14 vs. 6/12, p < 0.05) and there was a significant decrease in the incidence of ventilator dependence (0/14 vs. 5/12, p < 0.05) in the DEX group. DEX therapy did not influence the incidence of significant complications such as infection, periventricular leukomalacia or retinopathy of prematurity. We conclude that in a selected high-risk group of preterm infants, early low-dose DEX treatment results in improvement in pulmonary outcome without significant side effects. PMID- 10516396 TI - Role of nitric oxide in regulating neonatal porcine pulmonary artery smooth muscle cell proliferation. AB - Nitric oxide (NO), which is known to inhibit systemic vascular smooth muscle cell proliferation, is used in the management of neonatal pulmonary hypertension. Our objectives were to determine: (1) if endogenous NO production by neonatal porcine pulmonary artery smooth muscle cells (PASMCs) varied with oxygen tension in vitro, and (2) the effect of exogenous NO and inducible NO synthase (iNOS) stimulators and inhibitors on PASMC proliferation and apoptosis. PASMCs were exposed to different conditions (varying PO(2), NO donors and scavengers, iNOS stimulators and inhibitors) and proliferation, apoptosis, and cyclic guanosine 5(')-monophosphate (cGMP) assessed. PASMCs proliferated best between 5 and 10% O(2) but cGMP levels were similar at all oxygen levels. NO donors (S-nitroso-N acetyl-penicillamine, NOC-12, NOC-18) inhibited PASMC proliferation in a dose dependent manner with associated cGMP increases, while NO scavengers (carboxy PTIO), iNOS stimulators (interleukin-1beta, lipopolysaccharide), and iNOS inhibitors (aminoethylisothiourea) did not affect proliferation or cGMP. No changes in apoptosis were found at the concentrations of NO donors or iNOS stimulators used. These results suggest that while exogenous NO inhibits PASMC proliferation, endogenous NO may not regulate proliferation during changes in oxygen tension or cytokine levels. Endothelial derived and inhaled NO may attenuate smooth muscle hyperplasia and vascular remodeling. Inducible NOS in porcine PASMCs appears resistant to stimulation with interleukin-1beta or lipopolysaccharide. The mechanisms underlying hypoxia-mediated changes in PASMC proliferation require investigation. PMID- 10516397 TI - Prenatal stress and immune recognition of self and nonself in the primate neonate. AB - The capacity of the neonate to respond to nonself antigens was evaluated in infant monkeys born after normal and disturbed pregnancies. Mixed lymphocyte cultures were used to test the infants' proliferative responses to mitomycin treated stimulator cells, either from a genetically unrelated animal or from a virally transformed monkey cell line. Periods of daily stress for 6 weeks in mid late pregnancy (months 3.0-4.5) resulted in a significant decrease in proliferative responses, whereas the same stressor early in pregnancy (months 1.5 3.0) increased responses by the neonate's cells. Similar to the late stress effect, an inhibition of proliferative responses by neonatal cells was induced by dexamethasone administered for 2 days late in pregnancy at 4.5 months after conception, 1 month before term. These findings demonstrate that certain immune responses at birth are extremely sensitive to prior prenatal events. Further, the bidirectional changes indicate that there may be critical periods in gestation when the same extrinsic events have radically different effects on the fetus. PMID- 10516398 TI - Hypoxia-induced release of peptide growth factors from neonatal porcine pulmonary artery smooth muscle cells. AB - Peptide growth factors are involved in hypoxia-mediated neonatal pulmonary vascular remodeling. The role of hypoxia in the release of selected peptide growth factors from neonatal porcine pulmonary artery smooth muscle cells (PASMC) was examined. PASMC were exposed to different oxygen tensions and the cells were counted electronically and the conditioned media analyzed for basic fibroblast growth factor (bFGF), endothelin-1 (ET-1), platelet-derived growth factor (PDGF), and vascular endothelial growth factor (VEGF). The effect of conditioned media on PASMC proliferation was also measured. Hypoxia (1% oxygen) and hypoxia conditioned media increased PASMC numbers, and this mitogenic effect was abolished by anti-bFGF, but not by anti-PDGF or anti-VEGF. Hpyoxia increased bFGF and VEGF release but not PDGF or ET-1. This suggests that PASMC-derived bFGF and VEGF may participate in hypoxic neonatal pulmonary vascular remodeling. PMID- 10516399 TI - Interleukin-6 receptor is highly expressed in the ganglionic eminence of the human fetal brain. AB - The expression of interleukin-6 receptor (IL-6R) has been investigated immunohistochemically in seven fetal brains with special reference to the ganglionic eminence (GE), a part of the telencephalic proliferative zone. Between the 22nd and 28th gestational week the GE stands out conspicuously due to its very high amount of intensely IL-6R-immunostained cells. Adjacent brain areas exhibit only very weak immunoreactivity. Between the 32nd and 36th week a moderate IL-6R immunolabelling is seen in the remnants of the GE. IL-6 may activate the immature IL-6R-positive cells to secrete a protease which is likely to be involved in GE involution and, perhaps, in the development of hemorrhage frequently occurring in the GE of premature infants. PMID- 10516400 TI - The orthodox view of brain sexual differentiation. AB - The standard view of sexual differentiation of the brain, derived primarily from work with mammals, is that the same steroidal signal which permanently masculinizes the body early in life, androgen, also permanently masculinizes the nervous system. This oversimplified view overlooks the rich diversity of mechanisms produced by natural selection. We review the mechanisms underlying sexual differentiation of what may be the simplest mammalian model, the spinal nucleus of the bulbocavernosus (SNB), a system that is intimately associated with sexual differentiation of the periphery. Indeed, in many instances, early androgen can permanently masculinize the SNB system but, surprisingly, these early influences may depend to some extent on social mediating factors. Furthermore, in adulthood, androgen continues to affect the SNB system in diverse ways, acting on several different loci, indicating a life-long plasticity in even this simple system. Finally, there is evidence that adult androgens interact with social experience in order to affect the SNB system. Thus the SNB system displays a far richer array of interactions than the standard view of sexual differentiation would predict. Examination of other systems and other species, as depicted in the following reports, reveals a far more complicated, and far more interesting perspective on how the brains and behaviors of males and females diverge. PMID- 10516401 TI - Sexual dimorphisms in avian and reptilian courtship: two systems that do not play by mammalian rules. AB - Sexual dimorphisms in the central nervous system exist in numerous vertebrate species, and in many cases these structural differences between males and females parallel differences in the display of reproductive behaviors. Often both the behavioral and anatomical differences are controlled by exposure to gonadal steroid hormones, either during ontogeny or in adulthood. This article reviews some of the evidence supporting the hypothesis that in mammals, testosterone or its metabolites regulate the structure and function of neural and muscle systems involved in the control of masculine sexual behaviors. It then describes data suggesting that the mechanisms regulating sexually dimorphic courtship systems in zebra finches and green anole lizards are not completely parallel to the mammalian systems. Finally, some directions for future study are suggested, with the hope that they will stimulate thought about the nature of comparisons made across vertebrate models when investigators are attempting to determine both which morphological sex differences are important to the control of the reproductive behaviors, and which mechanisms regulating both structure and function are widely employed or are unique. PMID- 10516403 TI - Steroid hormones use non-genomic mechanisms to control brain functions and behaviors: a review of evidence. AB - Progestins, estrogens, androgens, and corticosteroids are capable of modifying brain functions and behaviors by mechanisms that involve the classic genomic model for steroid action. However, experimental evidence indicates that some responses to steroid hormones use non-classical, non-genomic mechanisms. This paper reviews the evidence that steroids can bind to receptors in the plasma membrane, activate cell signaling pathways, and regulate responses on a time scale of seconds or a few minutes. The existence of these alternative regulatory pathways for steroid hormones should make endocrinologists and neurobiologists change how they think about steroid hormones. It is no longer valid to assume that minute-to-minute changes in steroid concentrations are not regulating biologically important, short-term responses, or that the only steroids with biological functions are the ones that bind with high affinity to intracellular steroid receptors. PMID- 10516402 TI - Steroid sensitive sites in the avian brain: does the distribution of the estrogen receptor alpha and beta types provide insight into their function? AB - Studies in avian species have often been useful in elucidating basic concepts relevant to the regulation of reproductive behaviors by sex steroid hormones. Once a link between a steroid hormone and a behavioral response has been established, one can use the localization of steroid hormone receptors in the brain to facilitate the identification of neural circuits that control behavior. The recent identification of a second type of estrogen receptor called estrogen receptor beta or ERbeta has raised new issues about the action of steroid hormones in the brain. A hypothesis has been proposed by Kuiper et al. [1998] based on studies in mammalian species suggesting that ERalpha (the name given to the ER that was previously described) is important for reproduction while ERbeta is more important for non-reproductive functions. In this paper we apply this hypothesis more generally by examining possible functions of ERbeta in avian species. We have initiated studies of the ERbeta in the brain of two avian species, the Japanese quail (Coturnix japonica) and the European starling (Sturnus vulgaris). ERbeta was cloned in both species and the mRNA for this receptor type was localized in the brain employing in situ hybridization histochemistry methods. In both species ERbeta was found to be diffusely present in telencephalic areas consistent with a role for this receptor subtype in cognitive functions. However, ERbeta mRNA was also found in many brain areas that are traditionally thought to be important in the regulation of reproductive functions such as the preoptic region, the bed nucleus of the stria terminalis and the nucleus taeniae. Of the two receptor types, only mRNA for ERalpha was observed in the telencephalic vocal control nucleus HVc of male starlings. Steroid receptors in this nucleus are thought to be an example of an evolutionary specialization that has evolved to coordinate the production of courtship vocalizations with other aspects of reproduction. The lack of ERbeta mRNA expression in HVc is consistent with the hypothesis that ERalpha is preferentially involved in reproductive behaviors while ERbeta is involved in the steroid regulation of other neural functions. However, the widespread occurrence of ERbeta in other nuclei involved in reproductive function suggests that one must be cautious about the general applicability of the above hypothesis until more is known about ERbeta function in these other nuclei. PMID- 10516404 TI - Steroid hormones, dendritic remodeling and neuronal death: insights from insect metamorphosis. AB - Steroid hormones influence neuronal structure and function throughout the animal kingdom, via highly conserved receptor proteins. Insights into steroid effects on neurons and behavior have come from a range of vertebrate species including reptiles, amphibians, fish, birds, rodents and primates. In many instances, steroid hormones regulate the volume of particular regions of the nervous system by affecting both the number of constituent neurons and their size. A major determinant of neuronal number is the process of programmed cell death (PCD), which involves molecular machinery that is conserved across species. This article reviews steroid-mediated PCD and dendritic remodeling during metamorphosis of the hawkmoth, Manduca sexta. Metamorphosis is driven by a class of steroid hormones, the ecdysteroids. During the transformation from larva to pupa to adult moth, accessory planta retractor (APR) motoneurons of Manduca undergo dendritic regression and regrowth, and segment-specific PCD, in response to specific ecdysteroid cues. Experiments utilizing APRs in primary cell culture show that PCD is a direct response to ecdysteroids, regulated by the intrinsic segmental identity of individual APRs. As in other systems, activation of caspases (cysteine proteases) is involved in the execution phase of PCD. Other experiments demonstrate that the ecdysteroid-mediated regression of APRs' dendrites at pupation causes weakening of monosynaptic excitatory inputs from sensory neurons that trigger a larval withdrawal reflex. Thus, the steroid-mediated reduction in dendritic extent is linked to a specific electrophysiological and behavioral change during metamorphosis. The comparative approach, taking advantage of a variety of vertebrate and invertebrate species, holds the most promise for elucidating the full spectrum of steroid effects on neurons and behavior. PMID- 10516405 TI - Sex steroids and communication signals in electric fish: a tale of two species. AB - Weakly electric fish are good model animals to study the evolution of interspecific and sexual differences in communication signals. This is because the neural circuits producing these signals are simple and conserved among related species while the signals are highly species-specific, sexually dimorphic, and under hormonal control. Here we focus on two related species of weakly electric gymnotiform fish that emit a wave-type discharge. These species differ in the direction of the sexual dimorphism of their electric organ discharge (EOD) frequencies and their propensity to produce aggressive communication signals called 'chirps'. Brown ghost (Apteronotus leptorhynchus) males produce high frequency EODs while females produce low frequency EODs. When presented with an EOD mimic, males chirp frequently, while females seldom chirp. By contrast, black ghost (A. albifrons) males discharge at lower EOD frequencies than females, and there is no sex difference in chirping in this species. Accordingly, non-aromatizable androgens raise EOD frequency in brown ghosts, but lower it in black ghosts. Androgens induce chirping in female brown ghosts, but do not increase the propensity to chirp in female black ghosts. Thus, the difference in sexually-dimorphic communication signals between these two species can be explained by differences in their responses to sex steroids. Future studies will elucidate how the neural circuits generating these signals are differentially sensitive to steroids in these species. PMID- 10516406 TI - Clinical course of patients with coronary ectasia. AB - OBJECTIVE: To compare the clinical prognosis between patients with diffuse coronary ectasia and those with localized coronary ectasia. DESIGN: Patients with coronary ectasia were divided into two groups based on the Markis classification (group D: types I-III and group L: type IV), and the clinical manifestations and prognosis were compared between the two groups. RESULTS: Group D patients (52.1 +/- 4 years) were significantly younger than group L patients (62.5 +/- 7 years). During the study, 4 patients in group D died suddenly. Three of the patients had type I coronary ectasia, and 1 had left main coronary ectasia. CONCLUSION: The results of the present study indicate that patients with diffuse coronary ectasia and left main coronary ectasia should be followed carefully. PMID- 10516407 TI - Stroke in paced patients with sick sinus syndrome: influence of left atrial function and size. AB - Patients with sick sinus syndrome have a high prevalence of cerebral ischemia. The present study was designed to establish the prevalence of stroke in patients with sick sinus syndrome and the role of atrial size and function. This prospective study analyzed 100 consecutive patients with sick sinus syndrome without atrial fibrillation who received either dual chamber or ventricular pacemakers. Patients underwent a cranial CT scan at the time of enrollment and again at the end of the study 24 months later. Endpoint of the study was cerebral ischemia. Clinical and echocardiographic features were assessed at the beginning of the study. A multivariate regression analysis was applied to all variables that had at least a marginal univariate predictive value. Cerebral ischemia occurred in 18 patients. Univariate predictors for embolism were age >65 years (p < 0.001), low atrial ejection force (p < 0.01) and a dilated left atrium with spontaneous echo contrast (p < 0.05). These findings identified patients at high risk for the development of peripheral embolism among the group of patients paced for sick sinus syndrome. PMID- 10516408 TI - A quick simple method of determining platelet aggregability following glycoprotein IIb/IIIa receptor inhibitor administration. AB - Platelet aggregation assessment after therapeutic administration of a GP IIb/IIIa inhibitor has been difficult in routine practice due to the inherent nature of platelet preparation and aggregometry. Platelet microaggregation was performed via the utilisation of a Coulter counter. Citrate and heparin were evaluated as the anticoagulants, and ADP and adrenaline were evaluated as the stimulants. Aprotinin was also assessed. ReoPro was utilised in all assays as the GP IIb/IIIa inhibitor. Platelet microaggregation using the described methodology provides a quick, simple, reliable, and clinically valid method of assessing the percentage platelet inhibition due to the presence of a GP IIb/IIIa inhibitor. PMID- 10516409 TI - Long-term site-related differences in the progression and regression of the idiopathic mitral valve prolapse syndrome. AB - The natural history of uncomplicated mitral valve prolapse (MVP) is not clearly understood. To determine the site-related differences in regression and progression of MVP, 112 patients with idiopathic MVP were enrolled in this echocardiographic follow-up study. Cardiovascular complications, including dysarrhythmias (n = 3, 2.7%), overt congestive heart failure (n = 4, 3.6%), progression of mitral regurgitation over one grade (n = 28, 25.0%), newly confirmed chordal rupture (n = 1, 0.9%), and surgical repair (n = 2, 1.8%), were observed in these patients during a follow-up period of 1-13 years (mean, 4.0 +/- 2.8 years). Multivariate analysis and Kaplan-Meier analysis revealed that posterior leaflet prolapse and significant mitral regurgitation (grade >/=2) were considerable risks for cardiovascular complications. Regression of MVP was seen in 17 (18.7%) of the anterior prolapse patients; however, new prolapse was observed in 40 (35.7%) patients, mainly in posterior prolapse patients. These results suggest that site-related differences exist in uncomplicated MVP prognosis and that MVP in the posterior leaflet has a poor outcome compared to that in the anterior leaflet. PMID- 10516410 TI - Central venous catheter mechanical irritation of the right atrial free Wall:A cause for thrombus formation. AB - Thromboembolism is a major complication of long-term central venous catheter, usually associated with catheter or venous occlusion. Intracavitary right atrial thrombosis is currently considered to result from line-tip thrombosis extension. We report three adult patients in whom repeated mechanical trauma to the right atrial wall was probably the main mechanism. Transesophageal echocardiography revealed back and forth movement of the central catheter into a thrombus attached to the right atrial wall, thus suggesting a mechanism of catheter-associated thrombus formation, not previously visualized or suggested. Catheter removal and anticoagulation administration were undertaken with an uneventful clinical course and almost complete disappearance of the thrombi on transesophageal echocardiography follow-up. PMID- 10516411 TI - Gender differences in postinfarction left ventricular remodeling. AB - OBJECTIVE: Previous studies suggest that gender affects the adaptive responses of the heart to some forms of cardiac overload. It is unknown whether gender influences left ventricular (LV) remodeling after myocardial infarction (MI). METHODS: We performed transthoracic echocardiographic-Doppler examinations in age matched male (n = 17) and female (n = 16) rats before, and 1 and 6 weeks after transmural MI or sham surgery. RESULTS: Following large MI (male = 45 +/- 1% LV circumference vs. female = 48 +/- 4%, p = NS), both male and female rats developed progressive LV dilatation. Infarctions caused a similar degree of global and regional LV systolic dysfunction in males and females. Male rats had significant increases in the thickness of the noninfarcted posterior wall by 6 weeks after MI. However, posterior wall thickness did not change in the infarcted female rats. Average myocyte diameter in the noninfarcted region of the heart was also greater in male than female MI rats. The combination of increased cavity size with little change in wall thickness resulted in a greater decline in relative wall thickness in the female rats compared to the males. Male rats with MI showed progressively restricted LV diastolic filling as assessed by transmitral Doppler recordings. Female rats had less of an increase in the ratio of early to late transmitral velocities and less of an increase in the E wave deceleration rate after MI. CONCLUSIONS: Female rats showed a different pattern of LV remodeling than males with less of an increase in thickness of the noninfarcted portions of the left ventricle than males, but comparable LV cavity enlargement and systolic dysfunction. Despite similar infarct size, females developed less pronounced abnormalities of LV diastolic filling. We hypothesize that the gender-related differences in postinfarction LV remodeling may contribute to the different LV filling patterns, and might ultimately relate to differences in clinical outcome. PMID- 10516412 TI - Is the blood flow in the left ventricle during the isovolumic relaxation period a useful parameter of left ventricular systolic and early diastolic performance? AB - Left ventricular (LV) early diastolic performance is determined by LV behavior in the late systole to early diastole and may relate to the physical potential of patients. Isovolumic relaxation flow (IRF) velocity was obtained by continuous Doppler echocardiography in the left ventricle from the apex in 26 patients with atypical chest pain and 63 patients with coronary artery disease (CAD) with or without prior myocardial infarction (MI) who underwent cardiac catheterization. In each patient, a time constant of LV relaxation (tau) was calculated from the LV pressure waves obtained by a catheter-tipped micromanometer. The LV end systolic volume index was measured using contrast left ventriculography. IRF velocity in patients having CAD with prior MI (24.8 +/- 5.4 cm/s) was significantly less than in those with atypical chest pain (41.2 +/- 9.6 cm/s). It was also significantly less than in patients having CAD without prior MI (37.3 +/ 6.8 cm/s). IRF velocity significantly correlated with the time constant tau (r = -0.42, p < 0.001) and LV end-systolic volume index (r = -0.84, p < 0.001). This study indicates that IRF velocity obtained by continuous Doppler echocardiography in the left ventricle provides important information regarding LV systolic performance and early diastolic performance. PMID- 10516413 TI - The cost-effectiveness of losartan versus captopril in patients with symptomatic heart failure. AB - The Losartan Heart Failure ELITE Study recently found that in patients with symptomatic heart failure and a left ventricular ejection fraction of /=65 years with symptomatic heart failure. Data on health care resource utilization were collected as part of the trial. We conducted a cost-effectiveness analysis to estimate the lifetime benefits of treatment and the associated costs. We observed no differences between treatments in the number of hospitalizations, hospital days, and emergency room visits per patient over the trial period. We estimated the total cost of losartan to be USD 54 (95% CI: USD -1,717, USD 1,755) less per patient than captopril over this time frame. We also estimated that over the projected remaining lifetime of the study population, losartan compared to captopril would increase survival by 0.20 years (undiscounted) at an average cost of USD 769 (discounted) more per patient. This cost increase translated into a cost-effectiveness ratio of USD 4,047 per year of life gained for losartan relative to captopril. In patients with symptomatic heart failure, losartan compared to captopril increased survival with better tolerability at a cost well within the range accepted as cost-effective. PMID- 10516414 TI - Antihyperglycemic treatment in diabetics with coronary disease: increased metformin-associated mortality over a 5-year follow-up. AB - Mortality rates are considerably higher in chronic ischemic heart disease (IHD) patients with non-insulin-dependent diabetes mellitus (NIDDM) than in those who are nondiabetics. The relationship between different types of antihyperglycemic pharmacological therapy and mortality rate in this NIDDM population is uncertain. We aimed to examine the survival in NIDDM patients with IHD using various types of oral antidiabetic treatments over a 5-year follow-up period. The study sample comprised 11,440 patients with a previous myocardial infarction and/or stable anginal syndrome, aged 45-74 years, who were screened, but not included in the Bezafibrate Infarction Prevention study. Among them, 9,045 were nondiabetics and 2,395 diabetics. The diabetic patients were divided into four groups on the basis of their therapeutic regimen at screening: diet alone (n = 990), sulfonylureas (n = 1,041), metformin (n = 78) and a combination of a sulfonylurea and metformin (n = 266). All NIDDM groups were similar with regard to age, gender, hypertension, smoking, heart failure, angina and prior myocardial infarction. Crude mortality rate was lower in the nondiabetic group (11.21 vs. 21.8%; p < 0.001). In the diabetic group, mortality was 18.5% for patients on diet alone, 22.5% for those on sulfonylureas, 25.6% for patients on metformin, and 31.6% for the combined sulfonylurea/metformin group (p < 0.01). When analyzing age-adjusted mortality rate and actuarial survival curves, the lowest mortality was found in patients on diet alone and the highest in patients on metformin (alone or in combination with sulfonylureas). After adjustment for variables connected with long-term prognosis, the use of metformin was associated with increased relative risk (RR) for all-cause mortality of 1.42 (95% CI 1.10-1.85), whereas the use of sulfonylureas alone was not [RR 1.11 (95% CI 0.90-1.36)]. NIDDM patients with IHD using metformin, alone or in combination with sulfonylureas, exhibited a significantly increased mortality. Until the results of problem-oriented prospective studies on oral control of NIDDM will be available, alternative therapeutic approaches should be investigated in these patients. PMID- 10516415 TI - Metformin and risk of cardiovascular disease. PMID- 10516416 TI - Doppler velocity and flow through the aortic isthmus in normal term neonates. AB - Our hypothesis was that the relationship between the internal aortic diameter and the Doppler flow velocity across the aortic isthmus could be modeled by applying the principle of conservation of mass flow. The aortic diameter decreased at the isthmus by a mean of 18% (t = 11.02, p < 0.0001), while the flow velocity increased by a mean of 44% (t = 10.09, p < 0.0001). The mean peak mass flow rate was 34. 5 ml/s preisthmus and 32.9 ml/s at the isthmus with excellent correlation (r = 0.830). We conclude that the increase in Doppler velocity observed in the descending aorta can be explained by the normal narrowing observed at the aortic isthmus and application of the continuity equation for conservation of mass flow. PMID- 10516417 TI - Improved one-year survival after acute myocardial infarction in Icelandbetween 1986 and 1996. AB - During the last decade the treatment of patients with acute myocardial infarction (AMI) has changed dramatically. In order to evaluate the overall impact of these changes on mortality and morbidity, we collected data on all patients hospitalized for AMI in Reykjavik, Iceland, during the calendar years of 1986 and 1996. Demographical characteristics of AMI patients did not change significantly between study periods. One-year mortality decreased from 26.3 to 19.7% (p < 0.05). Patients discharged with aspirin or beta-antagonists as well as those who received thrombolytic therapy had decreased 1-year mortality both years. Patients discharged with diuretics, digoxin or antiarrhythmics had increased 1-year mortality. We conclude that the 25% reduction in 1-year mortality is partially due to changes in therapy. PMID- 10516419 TI - Effect of matrix metalloproteinase inhibitors on rat embryo development in vitro. AB - Matrix metalloproteinases appear to play a role in cell migration and tissue remodeling and are postulated to be important in the development of embryos. We hypothesized that inhibition of these proteinases with the synthetic inhibitor batimastat or with endogenous tissue inhibitor of metalloproteinases (TIMP) would alter in vitro development of day 9 rat embryos. Batimastat had a dose-related effect on embryo growth and viability following 48 h in culture, with significant inhibition at 1.0 microM concentration. Dying cells were observed in many organs of living embryos. TIMP-2 at 3.0 microg/ml had a similar effect on embryo growth. TIMP-1 alone or used in combination with TIMP-2 had no obvious effect. No focal malformations were observed. The effective concentrations are comparable to those which inhibit enzyme activity in cell culture. We conclude that matrix metalloproteinases are probably important in mammalian embryo development. PMID- 10516418 TI - Clonality of urogenital organs as determined by analysis of chimeric mice. AB - Though the first mammalian chimera was reported in 1961, suitable markers for different animal strains which are easily detectable in histological sections of all or most organs have not existed. Chimeric mice were produced having an excellent histological marker, the C3H antigen, which is strain-specific and fulfills all the criteria for an ideal strain-specific histological marker. Using male and female C3H-Balb/c chimeric mice we examined epithelial cells of urogenital organs and their morphological or functional units, such as the glomerulus, to determine whether individual organs and their morphological subunits were monoclonal or polyclonal in origin. We found that the epithelial parenchyma of most male and female urogenital organs (the prostate, seminal vesicle, epididymis, ovaries, vagina, kidney, ureter and bladder) and their morphological subdivisions were derived from cells of both input strains, indicating a polyclonal origin for each organ and/or organ component. A notable exception was the uterus in which all individual uterine glands examined (n = 403) were found to be either entirely Balb/c or entirely C3H, indicating a monoclonal origin. The clonality of urogenital structures is discussed in terms of the morphogenesis of the urogenital system. PMID- 10516420 TI - Effects of static electromagnetic fields on chick embryo pineal gland development. AB - The effects of static electromagnetic fields on the development of the chick embryo pineal gland were studied. A total of 144 fertilized White Leghorn eggs were sacrificed after 5, 10 and 15 days of incubation. The stage of development was determined in all embryos using the Hamburger and Hamilton method [J Morphol 49: 88-92, 1951]. The various morphometric parameters (diameter and distance of the pineal gland and its lumen) were measured on serial 7-micron-thick sections. The data were obtained in a morphometer and processed statistically. The intensities of the static electromagnetic fields were 18 and 36 mT. Control and exposed embryos were equally distributed and randomly assigned. After 5 days of incubation, 25% of embryos exposed to a static electromagnetic field of 18 mT had a more advanced stage of development than controls and embryos exposed to 36 mT. On the 10th and 15th day, embryos exposed to either 18 or 36 mT tended to be more developed than controls. In the morphometric study, results were similar for the controls and exposed embryos after 5 and 10 days of incubation. However, the values of the 15-day-old embryos exposed to static magnetic fields were lower than the values of the controls (p > 0.01). These differences were more pronounced in the embryos exposed to 36 mT. These results seem to indicate that static electromagnetic fields affect the development and growth of embryos unequally, and that their action can depend not only on the intensity of the static electromagnetic field, but also on the length of exposure and the organ which is developing. It may be interesting to use these data in ultrastructural and physiological studies. PMID- 10516421 TI - Quantitative analysis of extracellular matrix proteins in hypertrophic layers of the mandibular condyle and temporal bone during human fetal development. AB - A computer image analysis of immunostained extracellular matrix (ECM) proteins (collagen types I, II, III and V, fibronectin and tenascin) in hypertrophic layers in the mandibular condyle and temporal bone of human fetuses, which ranged in gestational age from 12 to 32 weeks, was performed. The percentage of cells positive for proliferating cell nuclear antigen (PCNA) increased in almost the same manner in each region. The level of PCNA was markedly elevated at 16 weeks. The percentage of PCNA-positive cells was low in temporal bone and in the bone forming layer of the mandibular condyle at 24 weeks. Specific concentration patterns of proteins in the ECM were found at each stage of development. The extent of accumulation of fibrillar collagen and of fibronectin differed, while those of other proteins in the ECM, such as tenascin, osteocalcin and osteonectin, were similar at the two sites. PMID- 10516422 TI - Ultrastructural study of the relationship between the morphogenesis of filiform papillae and the keratinization of the lingual epithelium in the mouse. AB - Tongues were removed from fetuses of mice on the 15th day of gestation (E15), from newborns (P0), and from juveniles on the 7th day (p7) and on the 21st day (P21) after birth for examination by light microscopy and transmission electron microscopy. In the fetuses at E15, no rudiments of filiform papillae were visible on the dorsal surface of the tongue. No evidence of keratinization was recognized throughout the entire dorsal lingual epithelium. At P0, rudiments of filiform papillae were compactly distributed over the dorsal surface, as are the filiform papillae in the adult, but their tips were rounder than those of the filiform papillae in the adult. Cell columns in the epithelium, with different degrees of keratinization of the type observed in the matured adult were indistinct. However, a keratinized layer was clearly visible on the tip of each filiform papilla. In juveniles at P7, the filiform papillae on the anterior part of the tongue were long and slender, and the anterior and posterior cell columns of the filiform papillae were identical to those in the adult. These results indicate that, in mice, the morphogenesis of filiform papillae advances in parallel with keratinization of the lingual epithelium from the stage just before birth to a stage a few weeks after birth. PMID- 10516423 TI - Nuclear morphologies of bovine corneal cells as visualized by confocal microscopy. AB - Confocal laser scanning microscopy was used to characterize nuclear morphology of the three cell layers of bovine cornea in vivo. Corneas fixed with formalin were stained with propidium iodide, whereas living cells in nonfixed corneas were stained with PicoGreen. Nuclei in the three corneal cell layers consistently assume strikingly different shapes. Round nuclei were observed throughout the layers of the epithelium of both fixed and living cells. Stromal fibroblasts (keratocytes) showed approximately equal numbers of elliptical and bean-shaped nuclei arrayed in a variety of orientations. Keratocytes near Bowman's layer had almost round nuclei whereas those near Descemet's membrane had more elongated elliptical nuclei. Lobulate nuclei arranged in a regular pattern were observed throughout the endothelium. Some of the lobulate nuclei were large and stained less intensely with the fluorescent dyes. In addition, keratocytes in vitro displayed the same two distinct nuclear morphologies as in vivo. These observations indicate that each cell layer of the cornea contains nuclei with characteristic morphologies and that, in the case of keratocytes, the cells maintain their characteristic nuclear morphologies in vitro. PMID- 10516424 TI - Correlation between surface electromyography and kinematics of the hindlimb of horses at trot on a treadmill. AB - The purpose of this study was to demonstrate the feasibility of surface electromyography in the horse and to correlate electromyographic activity with kinematic data. Surface electromyography of seven hindlimb muscles was recorded in five horses at trot on a treadmill. Simultaneously, kinematic analysis of the hindlimb was performed using a three-dimensional system and a unidirectional accelerometer was attached to the hoof. Electromyographic activities of the gluteus medius, vastus lateralis and two parts of the biceps femoris started in the late part of the swing phase and ended in the late period of the stance phase or the early period of the next swing phase. The semitendinosus showed two bursts of activity. The tensor fasciae latae acted when the previous muscles were inactive. Activity of the extensor digitorum longus was of low level but lasted during most of the step cycle. Correlation between electromyography, kinematics and anatomy helps us to understand the complex role of biarticular muscles: the tensor fasciae latae flexes the hip joint during the swing phase and extends the stifle joint during the stance phase, whereas the semitendinosus extends the hip joint during the stance phase and flexes the stifle joint during the swing phase. Cranial and caudal regions of the biceps femoris were also found to be bifunctional. Surface electromyography correlated with kinematic analysis gives valuable information about the functions and the timing of activity of the hindlimb muscles. PMID- 10516425 TI - Genomic cloning and chromosomal localization of the mouse Mab21l2 locus. AB - A second mouse gene related to the nematode mab-21 gene has been isolated. This gene, Mab21l2, encodes a transcript with an open reading frame discretely organized in a single exon. It shares 93.3% and 55.5% amino acid identity with the human and worm mab-21 respectively. FISH analysis determined that this gene is on chromosome 3 at a position between bands 3E3 and 3F1. This newly identified mouse gene will be useful in future examination of mab-21 gene function in vertebrate models. PMID- 10516426 TI - Assignment of the STOP gene (MAP6) to human chromosome bands 6p12-->p11 by fluorescence in situ hybridization. PMID- 10516427 TI - Assignment of cadherin-4 (R-cadherin, CDH4) to human chromosome band 20q13.3. PMID- 10516428 TI - Monosomy 6 in human cultured fibroblast-like cells permanently stimulated by fibroblast growth factor 1: evidence for selection. AB - The appearance of cells with monosomy 6 (mono6 cells) in cultures of human fibroblast-like cells during long-term stimulation with acidic fibroblast growth factor (FGF1) was confirmed in five of the six lines newly investigated. Aneugenic pretreatment at the start of the cultures accelerated the emergence of mono6 cells, as would be expected if selection, rather than induction, is the main mechanism involved. This could be confirmed by using an incidental rearrangement, der(8)t(6p;8p), that emerged in one of the lines by monitoring the proliferation of the mono6 cells (here monosomic for 6p22.1-->qter) in mixtures with normal cells. During growth in the presence of FGF1, the proportion of mono6 cells increased six fold, whereas in the absence of FGF1, it declined to background levels. Selection rather than induction of the mono6 cells is further supported by their clonal origin, as ascertained on the basis of X-inactivation patterns in three informative cases. In addition, colonies grown in the presence of FGF1 from single cells did not reveal higher proportions of mono6 cells by fluorescence in situ hybridization analysis than those grown without the growth factor. During permanent stimulation with FGF1, the growth of mono6 cells did not become dependent on FGF1, nor did these cells lose their responsiveness to FGF1. Although evidence in favor of selection of preexistent mono6 cells by FGF1 is provided in this study, the contribution of a primary inducing mechanism cannot be entirely excluded. PMID- 10516429 TI - Hybridization-based karyotyping of mouse chromosomes: hybridization-bands. AB - We have developed a method, which we have named hybridization-banding, to identify simultaneously all chromosomes in a mouse metaphase spread. The method uses a combination of hybridization probes labeled with a single fluor to yield a simple, unique, readily identifiable hybridization pattern on each chromosome. The method is superior to Giemsa- or fluorescence-based banding methods for chromosome identification because the hybridization patterns are simpler and easier to identify, and unique patterns can be designed at will for each chromosome. Analysis can be performed with a standard fluorescence microscope, and images can be recorded on film with an ordinary 35-mm camera, making the method useful to many investigators. The method can also be applied to any species for which chromosomes and probes can be prepared. PMID- 10516430 TI - Preferential S-phase pairing of the imprinted region on distal mouse chromosome 7. AB - Fluorescence in situ hybridization (FISH) has been used to visualize the spatial orientation of homologous chromosome regions in interphase nuclei of exponentially growing mouse fibroblasts. Simultaneous labeling of replicating DNA with the halogenated base analog 5-bromodeoxyuridine (BrdU) shows preferential somatic pairing of the imprinted region on distal mouse chromosome 7 during the S phase of the cell cycle. Trans-interactions between oppositely imprinted chromosome regions may be important for the maintenance of imprinting. PMID- 10516431 TI - Nondisjunction in human sperm: comparison of frequencies in acrocentric chromosomes. AB - Acrocentric chromosomes may be particularly predisposed to nondisjunction because of the frequency of trisomy for these chromosomes in human spontaneous abortions and liveborns. Studies of aneuploidy in human sperm have provided data on only a few acrocentric chromosomes, with evidence that chromosome 21 has a significantly increased frequency of disomy. To determine whether other acrocentric chromosomes have a higher frequency of nondisjunction or if chromosome 21 is anomalous, disomy frequencies for chromosomes 13 and 22 were studied by fluorescence in situ hybridization (FISH) analysis of 51,043 sperm nuclei from five normal men for whom the frequency of disomy for chromosomes 15 and 21 was known. The mean frequency of disomy for chromosome 13 (0.19%) did not differ significantly from that for other autosomes; however, the frequency of disomy 22 (1.21%) was significantly elevated (P < 0.001, Mantel-Haenszel chi(2) test). The G-group chromosomes (Nos. 21 and 22) also showed a significantly increased frequency of disomy (0. 75%) compared to acrocentric D-group chromosomes (viz., chromosomes 13 and 15; 0.15%) (P < 0.001, Mantel-Haenszel chi(2) test) and other autosomes (chromosomes 1, 2, 4, 9, 12, 13, 15, 16, 18, and 20; 0. 13%) studied in the same men (P < 0.001, Mantel-Haenszel chi(2) test). PMID- 10516432 TI - Multiple mono- and polymorphic Y-linked copies of the SRY HMG-box in microtidae. AB - Sex determination in mammals is controlled by SRY (sex-determining region of the Y chromosome), a single-copy gene located on the Y-specific region. Several exceptions to this rule have been described: some rodent species present Y specific multiple copies (either mono- or polymorphic) of this gene, and two Ellobius species and one Tokudaia species determine sex without a Y chromosome or the SRY gene. Recently, we have described multiple polymorphic copies of the SRY gene in both males and females of the vole species Microtus cabrerae. The female location and the presence of stop codons in some copies from males and females also suggest that they are nonfunctional copies of this gene (pseudogenes). We have investigated the SRY HMG-box in nine species of the family Microtidae; we report here the presence, in eight of these species, of multiple mono- or polymorphic copies of the SRY gene located on the Y chromosome. PMID- 10516433 TI - Structural unbalanced chromosome rearrangements resolved by comparative genomic hybridization. AB - The identification of unbalanced structural chromosome rearrangements using conventional cytogenetic techniques depends on recognition of the unknown material from its banding pattern. Even with optimally banded chromosomes, when large chromosome segments are involved, cytogeneticists may not always be able to determine the origin of extrachromosomal material and supernumerary chromosomes. We report here on the application of comparative genomic hybridization (CGH), a new molecular-cytogenetic assay capable of detecting chromosomal gains and losses, to six clinical samples suspected of harboring unbalanced structural chromosome abnormalities. CGH provided essential information on the nature of the unbalanced aberration investigated in five of the six samples. This approach has proved its ability to resolve complex karyotypes and to provide information when metaphase chromosomes are not available. In cases where metaphase chromosome spreads were available, confirmation of CGH results was easily obtained by fluorescence in situ hybridization (FISH) using specific probes. Thus the combined use of CGH and FISH provided an efficient method for resolving the origin of aberrant chromosomal material unidentified by conventional cytogenetic analysis. PMID- 10516434 TI - The pig aminoacylase 1 (ACY1) and ribosomal protein L29 (RPL29)/heparin/heparan sulfate interacting protein (HIP) genes are located together at 13q21-->q22, corresponding to human 3p21.1. AB - The pig aminoacylase 1 (ACY1; N-acylamino acid aminohydrolase) gene was isolated from a pig cosmid library and characterized. The gene spans about 4.7 kb and consists of 15 exons. Fluorescence in situ hybridization found ACY1 to be located on pig chromosome 13 in the region q21-->q22. This result and previous reports show that a large part of pig chromosome 13 corresponds to human chromosome 3. BLAST search results reveal that chromosome 13 contains a transcript similar to human ribosomal protein L29 (RPL29)/heparan sulfate/heparin-interacting protein (HIP). The transcript lies near the 3'-flanking region of the pig ACY1 gene; the 2 genes are linked tail-to-tail. The deduced amino acid sequence shows distinct homology to human RPL29/HIP, 96% identical in the N-terminal region. In the corresponding human 3p21.1 region, a deletion closely linked to the ACY1 locus has been observed in carcinoma cells. This suggests that a tumor suppressor gene is located in this region. Comparative mapping suggests also that the human RPL29/HIP gene may be near ACY1. Because many growth factors and cytokines interact with cells via heparin/heparan sulfate-proteoglycan, RPL29/HIP may play an important role on the cell surface by modulating interactions between cells and extracellular molecules. PMID- 10516435 TI - Assignment of the neuronal cochaperone, HSJ1, to human chromosome bands 2q32- >q34 between D2S295 and D2S339 by in situ hybridization and somatic cell and radiation hybrids. PMID- 10516436 TI - Assignment of TLL1 and TLL2, which encode human BMP-1/Tolloid-related metalloproteases, to chromosomes 4q32-->q33 and 10q23-->q24 and assignment of murine Tll2 to chromosome 19. PMID- 10516437 TI - Precise localization of D11S1226 to the human EMK1 gene at chromosome band 11q13 by sequence homology search. PMID- 10516438 TI - Assignment of the human P532 gene (HERC1) to chromosome 15q22 by fluorescence in situ hybridization. PMID- 10516439 TI - Gene structure and chromosome mapping of the human small-conductance calcium activated potassium channel SK1 gene (KCNN1). AB - Small-conductance, calcium-activated potassium channels contribute to the afterhyperpolarization in central neurons and other cell types. Because these channels regulate neuronal excitability, defects in their genes could cause excitability disorders. The human cDNA encoding one such channel, SK1 (KCNN1), was recently cloned. Here we describe the gene structure of KCNN1 and its localization by radiation hybrid mapping to chromosome 19p13.1. PMID- 10516440 TI - Centric fusion differences among Oryx dammah, O. gazella, and O. leucoryx (Artiodactyla, Bovidae) AB - G- and C-banded karyotypes of the genus Oryx were compared using the standard karyotype of Bos taurus. Chromosomal complements were 2n = 56 in O. gazella gazella, 2n = 58 in O. g. beisa and O. g. callotis, 2n = 56-58 in O. dammah, and 2n = 57-58 in O. leucoryx. The number of autosomal arms in all karyotypes was 58. Nearly all variation in diploid number was the result of three independent centric fusions, but one 2n = 57 specimen of O. g. gazella deviated from the normal complement of 2n = 56 due to XXY aneuploidy. A 2;17 centric fusion was fixed in O. g. gazella, whereas O. g. beisa and O. g. callotis lacked this fusion and had indistinguishable karyotypes. Oryx dammah was polymorphic for a 2;15 centric fusion, and O. leucoryx was polymorphic for an 18;19 centric fusion. The five Oryx taxa shared a fixed 1;25 centric fusion; the small acrocentric element involved in the 1;25 fusion was identified by fluorescence in situ hybridization using a cosmid specific to Bos chromosome 25. The X and Y chromosomes were also conserved among the five taxa. Oryx g. gazella differed from the other Oryx species because of the fixed 2;17 centric fusion. This difference reflects an apparently longer period of geographic isolation between O. g. gazella and other populations of Oryx, and it is consistent with the classification of O. gazella and O. beisa as distinct species (see Kingdon, 1997). The lack of monobrachial relationships among the Oryx taxa indicates that sterility barriers between species have not developed. Viability of hybrid offspring constitutes a threat to captive breeding programs designed for endangered species conservation; in the case of Oryx, the 2;15, 2;17, and 18;19 metacentrics could serve as marker chromosomes for assessing hybridization between certain Oryx taxa. PMID- 10516441 TI - Assignment of the murine adenomatous polyposis coli 2 (Apc2) gene to mouse chromosome band 10B5-C2 by in situ hybridisation. PMID- 10516442 TI - M31, a murine homolog of Drosophila HP1, is concentrated in the XY body during spermatogenesis. AB - The formation of the sex vesicle, or XY body, during male meiosis and pairing of the sex chromosomes are thought to be essential for successful spermatogenesis. Despite its cytological discovery a century ago, the mechanism of XY body formation, particularly heterochromatinization of the sex chromosomes, has remained unclear. The HP1 class of chromobox genes are thought to encode proteins involved in the packaging of chromosomal DNA into repressive heterochromatin domains, as seen, for example, in position-effect variegation. Study of the distribution of a murine HP1-like chromodomain protein, M31, during spermatogenesis revealed spreading from the tip of the XY body in mid-stage pachytene spermatocytes to include the whole of the XY body in late-pachytene spermatocytes. We also demonstrate that the formation of the XY body during spermatogenic progression in neonatal mice coincides with the expression of a novel nuclear isoform of M31, M31(p21). These results support the view that a common mechanistic basis exists for heterochromatin-induced repression, homeotic gene silencing, and sex-chromosome inactivation during mammalian spermatogenesis. PMID- 10516443 TI - Factors influencing the course of calcium oxalate stone disease. AB - OBJECTIVE: To assess the influence of previous stone formation, urine and stone composition on the further course of the disease in recurrent calcium stone formers without pharmacological treatment. METHOD: The course of the disease was analysed during a prospective follow-up period by means of Kaplan-Meier estimates. At the start of follow-up the patients were subgrouped with regard to their previous history of stone formation expressed as stone age index (SAI = 100 x number of stones/age), urine composition, stone composition, and sex. In 223 of the patients was it possible to calculate AP(CaOx) index(s), a standardized estimate of the ion-activity product of calcium oxalate. RESULTS: The 446 patients (329 men, 117 women) who were considered representative of an average population of recurrent calcium stone formers, had a 5-year recurrence risk of approximately 50%. Patients with an SAI <2 had a lower recurrence risk than those with an SAI >2 and a corresponding difference was recorded between patients with SAI levels <5 and >5. Furthermore, female patients had a lower risk of new stone formation than male patients. Patients with an AP(CaOx) index(s) of 1.5 or more had a significantly higher recurrence risk than those with a lower index, a difference that was most pronounced in female stone formers. A slightly higher risk of recurrent stone formation during the follow-up period could also be related to the presence of calcium phosphate in the stone, a high AP(CaP) index(s) (a standardized estimate of the ion-activity product of calcium phosphate) and a low concentration of citrate. CONCLUSION: AP(CaOx) index(s) and SAI were the most obvious predictors of the recurrence risk and these two variables, together with information on the sex distribution, might be useful for deriving an expected recurrence risk at a defined point of time in a group of recurrent stone formers. Such an estimate can be valuable for conclusions on the efficacy of different stone-preventive treatments when an appropriate control group is lacking. PMID- 10516444 TI - Clearance of lower-pole stones following shock wave lithotripsy: effect of the infundibulopelvic angle. AB - OBJECTIVE: To assess the effect of anatomic factors, especially the angle of the lower-pole infundibulum, on stone clearance following shock wave lithotripsy (SWL) in order to determine selection criteria for percutaneous nephrolithotomy. METHODS: We retrospectively analyzed 116 patients with single lower-pole stones measuring 11-20 mm treated with SWL. Intravenous urograms were reviewed to measure the infundibulopelvic angle, the angle of the infundibulum to the vertical, and the anatomy of lower-pole calyces. RESULTS: The overall stone-free rate was 52%. Factors most closely associated with a stone-free status were obtuse infundibulopelvic angle, lack of calyceal distortion, and a large infundibular diameter. The infundibulopelvic angle was the only factor to attain significance in predicting stone-free status (p = 0.012). The size of the stone did not predict eventual stone-free status (p = 0.911), but larger stones were more likely to require intervention after SWL. CONCLUSION: For solitary lower pole stones 11-20 mm in size, the angle of the lower-pole infundibulum as it relates to the pelvis plays a role in eventual stone clearance and should be taken into account before choosing a mode of treatment. PMID- 10516445 TI - Comparison of extracorporeal shock wave lithotripsy and ureteroscopy in the treatment of ureteral calculi: a prospective study. AB - 146 patients whose ureteral stones did not pass spontaneously participated in a prospective study on optimal management. Patients were offered two treatment options: extracorporeal shock wave lithotripsy (ESWL) and ureteroscopy (URS). The stone was treated with the technique preferred by the patient. In case of treatment failure after first-line therapy, patients again could decide on how to proceed. Stone analysis could be obtained from 72.6% patients. ESWL was the primary treatment in 66.4% patients. In 2 patients, ESWL was the secondary treatment after failed URS. URS was the first-line therapy in 33.6% patients. In 29 patients URS was done after failed ESWL. For analgesia, sedoanalgesia or spinal anesthesia were used. Analgesia was required in 74.2% ESWL and 100% URS sessions. Following ESWL, 70.1% patients became stone free. In 29.9% ESWL failed. Distal stones had a higher failure rate than proximal or mid-ureteral calculi. Distal stones treated without success were significantly larger than those treated successfully. Failures were switched to URS. Stone analysis could be obtained in 26 patients with failed ESWL: 23/26 consisted of pure whewellite or mixed whewellite stones. Clinically relevant complications were not observed. After URS, 94.9% of the patients became stone free. In distal stones, the stone free rate was 97.5%. There was only 1 relevant complication: a proximal ureteral lesion requiring surgical repair. Our study demonstrates that URS is a safe and highly effective treatment option for ureteral stones. In patients with distal ureteral stones, it should be offered as a first-line treatment. When whewellite is expected as the stone mineral, URS is the treatment of choice. PMID- 10516446 TI - T1 GIII bladder cancer. Management with transurethral resection only. AB - OBJECTIVE: Transurethral resection (TUR) is the elective procedure in the treatment of superficial bladder tumor. The association of intravesical chemotherapy has no influence on survival and cause specific survival. This study was carried out to determine the evolution of T1 GIII bladder carcinoma treated with TUR only. PATIENTS AND METHODS: We retrospectively reviewed the records of 42 consecutive patients with T1 GIII bladder carcinoma. Follow-up was available for 34 patients. No patient received either adjuvant or neoadjuvant therapy. All the patients were treated with TUR only and followed for a median of 40 months. They were followed by cystoscopy, urinary cytology and intravenous urography. RESULTS: Forty-seven percent of patients had a solitary tumor while 53% had multiple tumors. Tumor recurrence occurred in 50% with a disease-free interval until the first relapse of 9.6 months. Progression of the primary tumor was observed in 23.5% of patients. The overall survival rate was 73.6% and the cancer specific survival estimate was 88.2% at mean 36 months of follow-up. CONCLUSIONS: T1 GIII bladder carcinoma may be treated initially with transurethral resection only with acceptable recurrence and progression rates. This would avoid costs and complications of the adjuvant/neoadjuvant therapies. PMID- 10516447 TI - Transurethral resection and surveillance of bladder cancer supported by 5 aminolevulinic acid-induced fluorescence endoscopy. AB - PURPOSE: We determined whether neoplastic disease, which was missed under white light can be found during transurethral resection of bladder cancer by 5 aminolevulinic acid-induced porphyrin fluorescence. MATERIALS AND METHODS: 5 Aminolevulinic acid-induced fluorescence endoscopy was carried out in 328 cases. A 3% 5-aminolevulinic acid solution was instilled intravesically in a mean time of 2.8 h before endoscopy. The fluorescence was excited by a special incoherent light source which provided blue light in addition to white light. RESULTS: In 82 (25%) cases additional neoplastic lesions were found only because of their red porphyrin fluorescence which was induced by 5-aminolevulinic acid. 31% of these neoplastic foci which were found in normal and nonspecific inflamed mucosa had a poorly differentiated histology. CONCLUSIONS: 5-Aminolevulinic acid facilitates detection of neoplastic disease during transurethral resection of bladder cancer and increases the accuracy of diagnosis. PMID- 10516448 TI - Prognostic value of MIB-1 score, p53, EGFr, mitotic index and papillary status in primary superficial (Stage pTa/T1) bladder cancer: a prospective comparative study. The Finnbladder Group. AB - OBJECTIVE: A prospective randomized study was undertaken to determine whether cell proliferation indices (M/V index, MIB1), papillary status, the expression of p53 and epidermal growth factor receptor (EGFr) have prognostic value in superficial (pTa-pT1) bladder cancer (SBC). METHODS: 207 patients with primary SBC were followed up over a period of 4.9 (range 3.7-6.0) years. M/V index and papillary status were assessed by light microscopy, and expression of MIB1, p53 and EGFr was assessed by immunohistochemistry. The results of histopathological analyses were related to the survival data of the patients. RESULTS: Using univariate analysis, stage (p < 0.001), grade (p < 0.001), papillary status (p < 0.001), MIB1 (p < 0.001), M/V index (p < 0.001), EGFr (p < 0.001) and p53 (p = 0.002) were significant predictors of progression. Using multivariate analysis, MIB-1 score and papillary status were independent predictors of progressive disease and cancer-specific survival. Tumor grade was the only independent predictor of recurrence. CONCLUSION: Evaluation of tumor cell proliferation rate by M/V index or by MIB1 immunohistochemistry and assessment of papillary status by light microscopy are useful prognostic tools in tailoring treatment and follow up schedule of patients with SBC. PMID- 10516449 TI - Concomitant radiotherapy and carboplatin in locally advanced bladder cancer. AB - OBJECTIVE: The aim of the study was to assess the efficacy and safety of concomitant radiotherapy (CRT) and carboplatin. PATIENTS AND METHODS: From 1992 until 1997, 67 patients with T3 invasive bladder cancer (IBC) were treated using CRT and carboplatin. X-Ray radiotherapy (10 MeV) was applied using LINAC in a locoregional technique, with a total tumor dose of 65 Gy in 32 fractions. Carboplatin was administered as a bolus infusion once a week, on day 5, up to a total dose of 900 mg. RESULTS: The most frequent toxicity was hematological. Of the 67 treated patients, 92.5% achieved a clinically complete response, and 7.5% developed progressive disease during therapy. The 5-year overall survival was 55% and disease-free survival was 35%. CONCLUSION: CRT and carboplatin appear to be safe and extremely active in the treatment of T3 IBC, but the results should be confirmed in a randomized study. PMID- 10516450 TI - Effect of long-term sitting on serum prostate-specific antigen levels. AB - OBJECTIVE: The aim of this study was to evaluate the effect of long-term sitting on serum prostate-specific antigen (PSA) levels. METHOD: The serum PSA levels of 50 bus drivers under the age of 45 (mean 37.7) years who worked at least 8 h a day for more than 3 weeks were compared with those of 50 healthy surgeons in similar age groups (mean 37.7 years) who spent most of their working time standing. RESULTS: There was no statistically significant difference between the mean PSA level of the study group (1.211 +/- 0.96 ng/ml) and that of the control group (1.214 +/- 0.74 ng/ml; p > 0.05). The PSA levels returned to normal after a 5-day resting period in cases who had higher values than the anticipated 2.5 ng/ml for this age group at the initial determination. CONCLUSIONS: These results suggest that there is no relationship between long-term sitting and serum PSA levels. A second PSA determination after a 5-day resting period may be helpful in cases with higher than normal initial values. PMID- 10516451 TI - Serum levels of free and total prostate-specific antigen in males with liver cirrhosis. AB - OBJECTIVES: We examined the serum concentrations of total and free prostate specific antigen (PSA) from patients with liver cirrhosis to clarify the influence of liver function on serum PSA levels. METHODS: Serum concentrations of total and free PSA, as well as the free/total PSA ratio were measured in 75 male patients with histologically confirmed liver cirrhosis. As a control group, 75 age-matched healthy blood donors were studied. RESULTS: The serum levels of total PSA in liver cirrhosis patients were significantly lower than those in controls (p = 0.0004). However, the serum free/total PSA ratios in liver cirrhosis patients were significantly higher than those of controls (p < 0.0001). CONCLUSIONS: Our results suggest the absence of an obvious clearance disturbance of serum PSA as a result of severe hepatic dysfunction and offer caution against the utility of standard cutoff values of serum PSA or free/total PSA ratios in the detection and staging of prostate cancer in men with severe liver dysfunction. PMID- 10516452 TI - Influence of carbon dioxide on respiratory function during posterior retroperitoneoscopic adrenalectomy in prone position. AB - OBJECTIVE: To evaluate the influences of CO(2) insufflation on changes in blood gas analysis and end tidal CO(2) tension (PetCO(2)) during posterior retroperitoneoscopic adrenalectomy in the prone position. METHODS: Arterial blood gas analysis and measurements of PetCO(2) were carried out during CO(2) insufflation in 16 patients who underwent posterior retroperitoneoscopic adrenalectomy in the prone position (PRA group). The results were compared to 10 patients who underwent open posterior adrenalectomy (OPA group). Ventilation was artificially controlled during the study period in all cases. RESULTS: Arterial pH, PaCO(2), PetCO(2) and PaO(2) were not significantly different between the PRA and OPA groups. However, the PaCO(2)-PetCO(2) gradient in the PRA group was significantly higher than that in the OPA group (p < 0.01). CONCLUSION: Transperitoneal absorption of CO(2) occurs in patients undergoing retroperitoneoscopy in the prone position. The alveolo-arterial CO(2) gradient may be the only parameter which indicates the absorption of CO(2) during PRA. PMID- 10516453 TI - Laparoscopic treatment of parapubic postprostatectomy hernia. AB - We are reporting the case of a parapubic hernia that occurred after radical prostatectomy. This kind of the hernia is caused by the weakening of the attachments of rectus muscles to the pubic bone. Because of its location, it may be misdiagnosed as a far more common direct inguinal hernia. A laparoscopic approach made it possible to precisely diagnose and repair the defect in the abdominal wall. PMID- 10516454 TI - Sacral nerve stimulation and diurnal urine volume. AB - OBJECTIVES: During a screening trial to determine candidacy for sacral nerve stimulation (SNS), several patients were required to repeat the baseline dietary because of discrepancies in the 24-hour urine output before and after successful stimulation. This raised a question regarding the relationship between urine production and neuromodulation. A more complete diary analysis of patients affected by urgency/frequency and/or urge incontinence was therefore carried out to evaluate the possibility of a direct modulatory influence of SNS on urine production. METHODS: Voiding diaries of 40 patients (37 females and 3 males, average age 39.4 years) who underwent SNS were evaluated. Voiding diaries were obtained at baseline, during and after peripheral nerve evaluation (PNE) and after permanent implantation. RESULTS: There was an increase in the average volume/void during PNE in 39 patients. Twenty-four-hour urine volume during PNE was statistically greater than that at baseline. Volume/void and diurnal volume were also significantly greater in follow-up periods after permanent implantation. CONCLUSION: SNS appears to influence not only bladder function but also urine production. Increase in volume/void is paralleled by an increase in 24 hour urine output. The mechanism is unclear, but it is consistent with an altered release of antidiuretic hormone. This observation reflects the direct refractory involvement of the hypothalamus in micturition. PMID- 10516455 TI - Cultural adaptation of a quality-of-life measure for urinary incontinence. AB - OBJECTIVES: To translate and validate a urinary incontinence-specific measure of quality of life (I-QOL) in French, Spanish, Swedish, and German and provide translations only into seven other languages and variants of these languages. METHODS: Quality of life and linguistic experts prepared two forward translations from American English to their native languages and helped to harmonize these translations at a meeting. In the four European countries, the adapted versions of the I-QOL were administered to 259 women with stress, urge, and mixed incontinence. Principal component analyses were used to confirm the proposed measurement model suggested by patient interviews. Psychometric testing was conducted using standardized procedures. RESULTS: Translation procedures resulted in a change in the original instrument's Likert response scale from 4 to 5 points. Principal component analyses confirmed three patient-derived subscales and higher-order factor analysis confirmed a total summary score. In all countries, the internal consistency (alpha) and reproducibility (ICC) were high (alpha ranged between 0.87 and 0.93); (ICC ranged between 0.92 and 0.95). In all countries, I-QOL scores were significantly worse (p < 0.001) as perceived severity of incontinence, use of services, and number of incontinent episodes increased. CONCLUSIONS: The I-QOL has been adapted successfully into eleven languages and six variants of these languages. The cross-sectional psychometric properties of the US version were confirmed in four European countries. The I-QOL fills the need for a valid, international quality-of-life instrument for incorporation in clinical trials covering patients with varying types and severity of urinary incontinence. PMID- 10516456 TI - One-stage hypospadias repair. Experience with 544 cases. AB - PURPOSE: We evaluated one-stage hypospadias repairs in providing a normal looking penis with a normal functioning urethra. Also we looked critically at the effects of the severity of hypospadias, the type of repair and the experience of the surgeon on the outcome. MATERIALS AND METHODS: From 1987 to 1996 we performed 578 primary hypospadias repairs. The type and surgical results as well as the effects of certain variables on outcome were reviewed retrospectively. RESULTS: 544 single-stage hypospadias repairs have been followed up for a mean of 19 months (range 12-49). They included: MAGPI (92), ARAP (78), Mathieu (205), Mustarde (12), Duckett's tubularized preputial flap (142) and Onlay preputial flap (15). Despite an initial overall complication rate of 19%, the final success rate was 96%, after a mean of 1.3 procedures. Complications included fistula in 48 (9%) cases, meatal stenosis or retraction in 28 (5%), residual chordee in 17 (3%), stricture in 14 (2.5%), tubal abnormality in 10 (2%), and flap necrosis in 9 (2%). Complication rates were significantly higher (p < 0.05) when the meatus was proximal, the degree of chordee was moderate or severe and in the early series. Complication rates were also significantly higher with flap procedures and when the urethral plate was resected. Cosmetic defects occurred mainly with meatal advancement procedures. CONCLUSIONS: A repertoire of different types of single stage procedures has allowed the successful treatment of most hypospadias cases presenting to one surgeon. Complication rates increases with the severity of hypospadias or transection of the urethral plate. A cumulative experience allows for better results via a proper selection of the procedure and a perfection of a few techniques. PMID- 10516457 TI - Kock urinary reservoir maturation in children and adolescents: consequences for kidney and upper urinary tract. AB - OBJECTIVE: To study Kock reservoir maturation in children and adolescents and its effects on the kidneys and upper urinary tract. METHODS: Ten boys and 10 girls, aged 10.8-18 years, had Kock reservoir surgery for congenital urinary incontinence. They were followed for 3-10 years, divided into 3 different periods, and assessed with urography and enterocystography, the findings of which were correlated to renal function as measured by (51)Cr EDTA clearance, reservoir endoscopy and patient's history. RESULTS: The reservoir was located in the pelvis and remained in this position throughout the whole follow-up in 75% of patients and in the lower or midabdomen in 25%. Angled efferent nipple seen on enterocystoscopy or enterocystography coincided with nipple dysfunction, reservoir malposition or infrequent reservoir emptying. Upper urinary tract dilatation was detected in 84% of patients 3 months after surgery, 25% at 1 year and 30% at 2-10 years. The dilatation was improved in 56% of patients and unchanged in 25% after 1 year. The situation continued to improve at late follow up. New focal renal scars were radiologically detected in 1 of 19 at early and in another 1 of 17 patients at late follow-up. Progression of old scars was detected in 1 of 19 at early and in 4 of 17 at late follow-up. Eight of 19 cases had deterioration of renal function with a change in the split renal function. Of these 8 patients, 7 reported infrequent reservoir evacuation. CONCLUSIONS: Kock reservoir is a useful form of urinary diversion in children and adolescents with congenital urinary incontinence. Radiological examinations are good methods of follow-up of the maturation of the pouch and its effects on the urinary tract and for detection of complications. Urinary tract dilatation is a frequent finding early after surgery but it subsides in most cases 3-12 months after surgery. Long term efferent nipple dysfunction may be the result of angulation, reservoir stones, malposition and/or overdistension. Permanent renal damage may be due to pyelonephritis, stones, infrequent reservoir emptying or urinary obstruction. A strict regime of reservoir evacuation to avoid overdistension and nipple dysfunction and to decrease the possibility of renal function deterioration is strongly advisable in these patients. It is imperative that their own management of the reservoir is continuously supervised. PMID- 10516458 TI - The use of long-term defunctionalized bladder in renal transplantation: is it safe? AB - OBJECTIVE: Evaluation of the use of defunctionalized bladder in renal transplantation, concerning surgical complications. METHODS: In order to assess the complication rate of ureteral reimplantation in long-term defunctionalized bladder, we compared 20 patients on haemodialysis for more than 15 years (group I) with another 20 patients on haemodialysis for less than 5 years (group II). None of these patients had renal failure due to urological causes or neurogenic bladder. Non-stented extravesical ureteroneocystostomy was done routinely in all patients except 1 in group II who underwent Politano-Leadbetter ureteroneocystostomy and 7 patients in group I who underwent Politano-Leadbetter (3 patients) and pyelo-ureteral anastomosis using the recipient's native ureter (4 patients). The amount of residual urine was insignificant (<100 cm(3)) in both groups. RESULTS: The mean postoperative bladder catheterization period was 7.8 days in group I and 4.2 days in group II. Postoperative urinary tract infections were observed in 9 cases of group I and in 4 cases of group II. No surgical complications occurred in patients of group II, while there were 6 patients with surgical complications in group I: stenosis after a pyelo-ureteral anastomosis (1 case), stenosis after a ureterovesical anastomosis with Politano-Leadbetter technique (1 case), urinary fistulae (3 cases; 1 with Politano-Leadbetter ureteroneocystostomy and 2 cases with pyelo-ureteral anastomosis), and vesico ureteral reflux (1 case with Politano-Leadbetter ureteroneocystostomy). These 6 cases had the lowest bladder capacity (30-150 cm(3)) among our 40 patients. Graft losses were comparable between the two groups and were not due to surgical complications. CONCLUSION: Small defunctionalized bladders can be used in kidney transplantation, but it may represent an increased surgical risk due to difficulty in performing ureteral reimplantation. PMID- 10516459 TI - The role of antibiotics in the treatment of chronic prostatitis: a consensus statement. PMID- 10516461 TI - Human hepatitis viruses -from basic research to treatment. PMID- 10516460 TI - Gleason scores from prostate biopsies obtained with 18-gauge biopsy needles poorly predict Gleason scores of radical prostatectomy specimens. PMID- 10516462 TI - Hepatitis A virus proteins. AB - The RNA genome of hepatitis A virus (HAV) shares common characteristics of the picornavirus family. However, the nucleotide or amino acid sequences are distantly related with other members of the family. Like other picornaviruses, HAV proteins are cleaved from a large polyprotein (PO), but the processing and some products are quite different. The 3C protein is the sole processing enzyme, and the primary cleavage takes place at the 2A/2B site. Several VP1-2A sites are proposed. In some strains, the intermediate VP1-2A polypeptides are assembled in the virion. The VP4 is very small and not detected in the mature virion. Some mutations in 2B, 2C and 3A proteins are identified to enhance viral replication or to induce cytopathogenic effects in the viruses adapted to cell cultures. PMID- 10516463 TI - Hepatitis B virus mutants. AB - During hepatitis B virus (HBV) infection, selection and takeover of mutant viruses are frequent events driven by both humoral and cellular host-immune response and antiviral therapy. Therefore, dynamic studies of the variations of viral mutants over time within the overall viral population in relation with the host-virus interactions are extremely important to better understand the biological and pathogenic role of each mutant. With these premises, we review the more frequent mutations detected in each of the 4 open reading frames of HBV. A detailed analysis of the pathobiologic implications of pre-C region mutations which suppress the expression of the hepatitis B 'e' antigen, will be presented, as these mutations induce a specific change in virus biology and the variations of the ratio between wild-type and mutant correlate with significant events in host-virus interactions. PMID- 10516464 TI - Hepatitis B virus X protein: structure, function and biology. AB - The X gene is conserved among mammalian hepadnaviruses and the X protein, pX, is essential for viral propagation at least in the woodchuck. During the last decade, efforts have centered on elucidating the oncogenic role of pX in hepatitis B virus infection. The accumulating knowledge on pX indicates that it is a multifunctional regulatory protein which modulates many host functions by communicating directly or indirectly with a variety of host targets as is the case for many viral regulatory proteins, such as T antigens, E1A, and human T cell lymphotropic virus tax. pX, which modulates the transcription machinery and/or modulation protein kinase signaling cascades, transactivates many host genes involved in cell proliferation, cytokine networks, acute immune response, and house-keeping functions. Distinct from the transactivation, pX also modulates DNA repair processes by interacting with p53 and/or repair enzymes which may accumulate mutations and sensitize cells to genotoxic stimuli. Several X interaction host proteins remain to be characterized as targets of pX. The biological roles of pX have been addressed in various systems in addition to the role of pX on viral reproduction. pX may affect cell cycle progress, response to apoptotic stimuli, cell transformation, and carcinogenesis in the presence or absence of additional oncogenic factors. These biological roles of pX have not been described in terms of pX functions and targets and remain subjects of future research using improved experimental systems and technologies. Such efforts will identify important function(s) of pX for hepatocarcinogenesis. PMID- 10516465 TI - Hepatitis B virus replication: novel roles for virus-host interactions. AB - Chronic hepatitis B continues to be one of the most widespread and serious viral infections in humans worldwide. Several fundamental aspects of the molecular biology of its causative agent, hepatitis B virus, are meanwhile understood in some detail. However, recent research has emphasized that the dependence of the viral infectious cycle on cellular factors is far greater than previously anticipated. More and more intracellular interactions between viral and cellular components are discovered, and probably each individual step of genome replication will turn out to involve several host factors. Prominent examples are the activation of the viral reverse transcriptase, P protein, by chaperones, and the nucleocytoplasmic trafficking of viral nucleic acids by as yet unidentified components of the host machinery. Some of these new developments will be described here but many more can be expected to follow. Identifying these host factors and characterizing their interactions with the viral components will certainly reveal novel targets for specific antiviral strategies. PMID- 10516466 TI - DNA vaccines. AB - Chronic infections with hepatitis B (HBV) and hepatitis C (HCV) virus are worldwide problems often leading to the development of chronic liver disease and hepatocellular carcinoma. Genetic immunizations with DNA encoding for structural and nonstructural proteins of HCV and HBV in experimental mice generate a broad base CD4+ and CD8+ cellular immune response which may be required for viral clearance from the host. DNA based immunization is a promising antiviral approach for the development of therapeutic and prophylactic vaccine against HBV and HCV. PMID- 10516467 TI - New antiviral agents for the therapy of chronic hepatitis B virus infection. AB - The development of new antiviral strategies for the treatment of chronic hepatitis B remains a major goal since hepatitis virus (HBV) is resistant to interferon treatment as well as to new nucleoside analogs. HBV is a small DNA virus that replicates its genome via a reverse transcription step. The viral polymerase has been the main viral target that was studied to design new antiviral treatments. Active research has led to the discovery of new nucleoside analogs that are potent inhibitors of the viral reverse transcriptase. Among them, lamivudine has also proven antiviral efficacy in clinical trials with a sustained inhibition of viral replication. However, due to the kinetics of viral clearance, long-term antiviral therapy is necessary to eradicate viral infection. These prolonged regimens are associated with the emergence of drug-resistant strains that harbor mutations in the viral polymerase gene within the conserved B and C domains. New approaches using combinations of nucleoside analogs or other strategies, such as immune intervention (DNA vaccine, stimulation of the TH1 response ) or gene therapy (antisense oligonucleotides, dominant negative mutants), should therefore be evaluated in animal models to optimize the current antiviral protocols. PMID- 10516468 TI - Processing and functions of Hepatitis C virus proteins. AB - Hepatitis C virus (HCV) has a positive-stranded RNA genome of about 9.5 kb and a large open reading frame encoding a precursor polyprotein of ca. 3,000 amino acids (aa). This polyprotein is cleaved by host cellular signalase(s) and viral proteases into 10 viral proteins in the order of NH(2)-Core-E1-E2-p7-NS2-NS3-NS4A NS4B-NS5A-NS 5B-COOH. Core and E1/E2 are considered to be a capsid protein and envelope glycoproteins, respectively. NS2-NS5B are putative nonstructural proteins involved in the replication of HCV. NS2/3 is a metalloprotease which cleaves in cis at the NS2/3 junction. NS3 possesses serine protease and RNA helicase activities and is responsible for the cleavage of the remaining nonstructural proteins. NS4A is suggested to be a cofactor for NS3 protease. Although the function of p7, NS4B and NS5A are still unknown, an association of a mutation in NS5A with a susceptibility to interferon (IFN) has been reported. NS5B possesses an RNA-dependent RNA polymerase activity. Most of the current findings in HCV proteins depend on expression studies of HCV cDNA clones because of the lack of an efficient replication system in cell cultures. Therefore, a final assignment of cleavages and functions of HCV proteins has to await the propagation of HCV in cell cultures. PMID- 10516469 TI - Epidemiology of hepatitis C virus in japan. AB - Blood screening for hepatitis C virus (HCV) antibodies at Japanese Red Cross blood centers has indicated an age-dependent prevalence of HCV infection in the general population of Japan: the older the age, the higher the prevalence. The same pattern was corroborated by the sentinel study conducted in an HCV-endemic area. The high prevalence of HCV in the elderly is most likely due to a spread of HCV infection during the turmoil period just after World War II: in particular by illicit intravenous amphetamine abuse. Fortunately, however, since the rate of newly acquired infection is currently too low to influence HCV prevalence in each age group, the total number of HCV carriers (estimated to be 691,852 for the age group 20-64 years) would decrease spontaneously with generation takeover to approximately 50% of the current number during the decade to come. PMID- 10516470 TI - Molecular evolution of hepatitis viruses. AB - Methods and models for analytical and quantitative researches on the evolution of hepatitis viruses using nucleotide sequences and amino acids of hepatitis viruses are described. Methods for the calculation of evolutionary rates and the construction of molecular evolutionary trees, especially by estimating the number of nucleotide substitutions and the number of amino acids, are introduced hereinafter. In addition, the evolution of the hepatitis virus is discussed by applying these methods to hepatitis B virus and hepatitis C virus. PMID- 10516471 TI - Problems in the treatment of hepatitis C with interferon. AB - A marked increase in the rate of eradication of hepatitis C virus (HCV) has been achieved by the combination of interferon-alpha2b and ribavirin when compared with interferon-alpha2b alone. However, even with combination therapy, hepatitis persists in more than half of the patients with chronic herpatitis C and progresses to liver cirrhosis and hepatocellular carcinoma with time. What needs to be kept in mind is that, whether by its natural course or by therapy to suppress liver inflammation or by interferon therapy, the rate of development of hepatocellular carcinoma is reduced if ALT is maintained at low levels. Attention should therefore be focused on the development of drugs which enhance the effect of interferon as well as drugs which suppress liver inflammation. PMID- 10516472 TI - Hepatitis delta virus. AB - The purpose of this article is to give a concise update on our understanding of hepatitis delta virus (HDV). For more extensive background information the reader is directed to published reviews [1-5]. PMID- 10516473 TI - Hepatitis E virus. Advances in HEV biology and HEV vaccine approaches. AB - Hepatitis E, previously known as enterically transmitted, enteric, or epidemic hepatitis, is a worldwide public health problem. The causative agent, the hepatitis E virus, is involved in epidemic, sporadic, and fulminant hepatitis cases worldwide. This review describes the advances in the biology of the hepatitis E virus and the progress made to develop simple and robust serologic assays for the diagnosis of HEV infection. Genomic sequence comparisons with a recently identified US isolate now suggests three genetic groups of HEV viruses. A highly conserved animal isolate found in pigs suggest the coexistence of animal and human isolates of HEV. The use of recombinant technology to develop an effective subunit vaccine capable of providing cross-protection for the most divergent HEV strains has been established and is reviewed. PMID- 10516474 TI - GB virus C/Hepatitis G virus. AB - Hepatitis G virus (HGV) is a positive, single-strand RNA virus that has been classified in the family Flaviviridae. The 5'-untranslated region (UTR) of the HGV genome is lengthy and does not share any significant primary and secondary RNA structures with the 5'-UTR of hepatitis C virus (HCV). The internal ribosome entry site has extraordinarily weak activity. The HGV genome does not seem to encode a nucleocapside protein analogous to HCV. Blood-borne transmission is presumed to be the commonest mode of transmission of the virus. Current infection with HGV is diagnosed by detection of HGV RNA by the polymerase chain reaction (PCR), and past infection with HGV is detectable by testing anti-HGV E2. HGV is distributed worldwide, but its prevalence varies widely from one population to another. Although the prevalence of HGV in association with acute and chronic hepatitis is higher than that in the general population, further prospective studies are needed to demonstrate its relative significance in causing hepatitis and other disease. The major unresolved biological issue at the moment is its hepatotropsim and site of propagation. Recent progress demonstrates that HGV replicates in lymphocytes rather than hepatocytes. HGV may be pathogenic under special conditions, but does not influence carcinogenesis. PMID- 10516475 TI - A novel unenveloped DNA virus (TT virus) associated with acute and chronic non-A to G hepatitis. AB - In 1997, a novel DNA virus was isolated from the serum of a patient with posttransfusion hepatitis of unknown etiology in Japan, and it was named TT virus (TTV) after the initials of the index patient. TTV is a nonenveloped, single stranded and circular DNA virus, and its entire sequence of approximately 3.9 kb has been determined. For being a DNA virus, TTV has a wide range of sequence divergence, allowing the classification into at least 16 genotypes separated by a sequence difference of >30% from one another. The nucleotide sequence of the noncoding region of the TTV genome is conserved, whereas that of the coding region is highly variable. TTV strains with extremely high sequence divergence are common in the same individuals, thereby indicating a mixed infection of TTV strains of different genotypes. An association is found between hepatitis of unknown etiology and the TTV genotypes which are detectable by PCR with primers deduced from the N22 region (genotype 1) in the open reading frame 1 encoding the capsid protein. It would be important to select the primers for specific detection of the TTV genotypes associated with clinical diseases, to further evaluate the capacity of TTV to induce acute and chronic liver disease as well as extrahepatic manifestations. PMID- 10516476 TI - Hepatitis C virus and hepatocarcinogenesis. AB - Although human hepatocellular carcinoma (HCC) is one of the most common types of tumors in the world, the molecular mechanisms underlying hepatitis-C-related human hepatocarcinogenesis are still not clear. HCC is accompanied by virus infections in most cases, and it is suggested that hepatitis B virus and hepatitis C virus (HCV) significantly influence the oncogenic process. The persistence of inflammation following HCV infection is reportedly related to carcinogenesis, and the mechanism of chronic inflammation has been approached by taking viral, immunologic, cytokine and apoptotic responses into consideration. With the progress made in molecular biology, the functional abnormality of oncogenes/tumor suppressor genes has been identified and, apart from the p53 gene, involvement of the IGF-II gene has also been described recently. Furthermore, it has been suggested that uncontrolled proliferation of cancer cells might be based on abnormal regulation of intracellular signal transduction pathways. Here we review the cutting edge of molecular hepatitis C virology in terms of virus-cell interactions, which may contribute to the development of human HCCs. We also discuss the recent progress made in the molecular and cell biology of human hepatocarcinogenesis. PMID- 10516477 TI - Effect of exogenous nitric oxide and inhibitors of nitric oxide synthase on the hypothalamic pituitary adrenal axis responses to neural stimuli. AB - It has been shown that the hypothalamic-pituitary-adrenal (HPA) axis responses to immune-derived stimuli in particular can be modulated by nitric oxide (NO). In the present study we examined the effect of endogenous and exogenous NO on the HPA axis responses to neural stimuli which are not related to immune functions. Intracerebroventricular injection of NOR-3, a donor of NO, had no effect on basal HPA axis activity but significantly attenuated the secretion of median eminence (ME) CRH-41 as well as the serum ACTH and corticosterone (CS) in response to acute photic stimulation in a dose-dependent manner. Intracerebroventricular administration of N-omega-nitro-L-arginine methyl ester (L-NAME), a general NOS inhibitor, significantly enhanced ACTH and CS responses to this stress but did not change the basal levels of these hormones. On the other hand, i.c.v. injection of aminoguanidine, an inhibitor of inducible NO synthase (NOS) but not of neuronal NOS, did not affect the HPA axis responses to photic stimulation. These results suggest that: (1) NO is involved in modulation of the HPA axis responses to neural stimuli which are not dependent on immune factors, (2) the effect of NO is mediated by inhibition of hypothalamic ME CRH-41 secretion, and (3) this effect is probably mediated by neuronal NOS and not by inducible NOS. PMID- 10516478 TI - Decreased type 2 corticotropin-releasing hormone receptor mRNA expression in the ventromedial hypothalamus during repeated immobilization stress. AB - Chronic or repeated stress results in reduction of food intake and body weight in rats. Stress-induced anorexia has been attributed to increased corticotropin releasing hormone (CRH) function in the central nervous system. To explore possible roles of other neuropeptides and peripheral hormones involved in food intake and energy utilization during continuing stress, we examined the impact of repeated immobilization stress on expression of mRNAs coding for CRH, neuropeptide Y (NPY), galanin and pro-opiomelanocortin (POMC) mRNAs in such hypothalamic nuclei as the paraventricular nucleus (PVN), arcuate nucleus (ARC) and dorsomedial hypothalamus (DMH), as well as plasma insulin and leptin concentrations. Changes in type 2 CRH receptor (CRHR-2) mRNA in the ventromedial hypothalamus (VMH), a possible target of anorectic CRH effect, were also examined. Rats were immobilized for 2 h daily for 6 days and sacrificed 24 h after the last immobilization. Immobilized rats had lower food intake and body weight and higher levels of PVN CRH mRNA than controls. Repeated immobiliza tion also lowered plasma insulin and leptin concentrations and VMH CRHR-2 mRNA levels. These results provide additional evidence linking VMH CRHR-2 mRNA levels to plasma leptin concentration. ARC NPY and DMH galanin mRNAs increased following repeated immobilization, while ARC POMC mRNA decreased. DMH NPY mRNA and ARC galanin mRNA were unaltered by immobilization. Since NPY and galanin are considered orexigenic, while the POMC-melanocortin-4 receptor system is apparently anorexigenic, the changes in neuropeptide mRNAs and VMH CRHR-2 mRNA may play counterregulatory roles against anorectic CRH effects. PMID- 10516479 TI - Thyroid hormone coadministration inhibits the estrogen-stimulated elevation of preproenkephalin mRNA in female rat hypothalamic neurons. AB - Expression of the enkephalin gene in ventromedial hypothalamus (VMH) of the female rat has been correlated with the performance of lordosis behavior. By antisense DNA evidence, it has been drawn into a causal role as well. Here, we explored whether, parallel to earlier molecular and behavioral results, thyroid hormone coadministration could disrupt the estrogenic induction of preproenkephalin (PPE) mRNA. As expected, estradiol benzoate treatment to ovariectomized rats led to a large and significant increase in PPE gene expression in the VMH. This increase was inhibited by coadministration of thyroid hormone. The thyroid hormone interference in PPE gene expression was specific to the VMH, as there were no significant effects in the central nucleus of the amygdala or in the caudate/putamen. These in situ hybridization histochemical results form a direct parallel both to previous transcriptional measurements and to reproductive behavior assays in which thyroid hormones were able to oppose estrogenic facilitation. Previous evidence supports the notion of competitive DNA binding and protein/protein interactions providing mechanisms for nuclear thyroid hormone receptors to affect estrogen receptor function, but other, additional mechanisms cannot be ruled out. To date, both oxytocin and PPE gene expression represent potential hypothalamic systems by which thyroid hormones could interfere with estrogen-stimulated female rat reproductive behavior. PMID- 10516480 TI - Central LPS-induced c-fos expression in the PVN and the A1/A2 brainstem noradrenergic cell groups is altered by adrenalectomy. AB - The A1 and A2 brainstem noradrenergic cell groups project to the hypothalamic paraventricular nucleus (PVN), which is involved in integrating the stress response. Bi-directional communication between the brain and immune system is well established, with both neuroendocrine and immune responses being activated by lipopolysaccharide (LPS). The mechanisms underlying such activation and differences between alternative routes of administration remain unclear. We examined activation of the PVN and A1/A2 cell groups, by assessing c-fos mRNA, or counting Fos-positive neurons in either the PVN or in brainstem A1/A2 cell groups 3 h after intracerebroventricular (i.c.v.) LPS, in control and adrenalectomized (ADX) rats. We also measured corticotropin-releasing hormone (CRH) mRNA in the PVN, and plasma corticosterone (CORT) levels. A group of ADX/CORT-replaced animals received i.c.v. LPS, and CRH mRNA and Fos peptide in the PVN were analysed. ADX increased CRH mRNA in the PVN, as did LPS, but no enhancement of this response was seen in LPS/ADX animals. C-fos mRNA also increased in both the PVN and the A2 cell group following LPS, but this response was potentiated by ADX. Fos peptide-containing cells increased in the PVN and A2 following LPS, and this change was amplified by ADX. Only 11.25% of Fos was found in DBH-positive (putative noradrenergic) neurons, suggesting activation of neurons containing other transmitters. ADX/LPS/CORT animals showed numbers of Fos neurons in the brainstem, and CRH mRNA levels in the PVN which were comparable to intact/LPS animals. Central LPS activates the hypothalamo-pituitary-adrenal axis, a process mediated partly by brainstem noradrenergic neurons, suggesting the involvement of afferent/efferent pathways within the brain. Peripheral administration of LPS involves activation of vagal inputs leading to the nucleus tractus solitarius. We suggest that centrally administered LPS activates the A2 cell group by a mechanism independent of the vagus. In the absence of CORT, despite the lack of a CRH mRNA response, an exaggerated c-fos and peptide response to LPS is observed, which is reversed following CORT pretreatment. PMID- 10516481 TI - Sprague-Dawley rats obtained from different vendors exhibit distinct adrenocorticotropin responses to inflammatory stimuli. AB - The purpose of this work was to compare the plasma adrenocorticotropin (ACTH), corticosterone and interleukin-6 (IL-6) responses that rats of the outbred Sprague-Dawley strain obtained from two different vendors: Charles River (CR) and Harlan (HSD). Basal plasma ACTH and IL-6 concentrations were similar in rats from either vendor (HSD or CR), while CR animals exhibited slightly elevated corticosterone levels in late afternoon. Inflammatory stimuli such as lipopolysaccharide (LPS) (1 microgram/kg, i.v.) or turpentine (50 microliter/100 g, i.m.) which induce the production of endogenous cytokines, produced a significantly larger ACTH response in CR, compared to HSD rats, while the overall corticosterone responses were comparable in both rat groups. This could probably not be accounted for by a greater ACTH responsiveness in CR rats per se because CR and HSD rats showed similar peak ACTH responses to electrofootshock. Furthermore, in contrast to when the stimulus was one that induced endogenous cytokine production, the administration of exogenous interleukin-1beta (IL-1beta, 200 ng/kg, i.v.) produced a 2-fold greater rise in plasma ACTH concentrations in HSD rats compared to CR rats. The plasma IL-6 responses to the inflammatory stimuli showed a similar pattern to ACTH, with LPS and turpentine tending to pruduce greater IL-6 responses in CR rats, though these differences were not statistically significant. In contrast HSD rats had a significantly greater IL-6 response to IL-1beta than did CR rats. Collectively, these results show that Sprague-Dawley rats obtained from different commercial sources can differ in immune-neuroendocrine responses to inflammatory stimuli. PMID- 10516482 TI - Effects of chronic 'Binge' cocaine administration on plasma ACTH and corticosterone levels in mice deficient in DARPP-32. AB - The product of the DARPP-32 gene mediates intracellular signals initiated by the binding of dopamine to its receptors. Cocaine administration leads to increased activation of dopamine receptors, and causes activation of the stress-responsive hypothalamic-pituitary-adrenal (HPA) axis. We determined the effects of chronic 'binge' pattern cocaine on HPA activity in mice containing a targeted disruption of the DARPP-32 gene. Mice received three daily injections of cocaine (15 mg/kg/injection) for 14 days, and were sacrificed 30 min after the last injection. We measured the levels of plasma adrenocorticotropin (ACTH) and corticosterone which reflect HPA activity. In wild-type controls, 'binge' cocaine administration significantly increased plasma ACTH and corticosterone levels. In contrast, DARPP-32-deficient mice failed to show a significant elevation of either plasma ACTH or corticosterone levels following 'binge' cocaine. The results indicate that DARPP-32 plays a role in mediating the stimulatory effects of cocaine on the HPA axis. PMID- 10516483 TI - Corticotropin-releasing effect of hexarelin, a peptidyl GH secretagogue, in normal subjects pretreated with metyrapone or RU-486, a glucocorticoid receptor antagonist, and in patients with Addison's disease. AB - GH secretagogues (GHS) are peptidyl and nonpeptidyl molecules which possess strong GH-releasing activity but also stimulatory effect on hypothalamo-pituitary adrenal axis. The ACTH and cortisol responses to Hexarelin (HEX), a peptidyl GHS, are abolished by low-dose dexamethasone pretreatment in normal subjects but are exaggerated and higher than those after hCRH in patients with pituitary ACTH dependent Cushing's disease, in spite of their hypercortisolism. Based on the foregoing, we studied the ACTH, cortisol and GH responses to HEX (2.0 microgram/kg i.v. at 0 min) alone and after metyrapone (2 g p.o. at 23:00 h the night before) or RU-486 (400 mg p.o. at 02:00 h), a glucocorticoid receptor antagonist, in 6 normal women (NS, age 26-34 years). The endocrine responses (mean +/- SEM) to HEX alone were also studied in 8 patients with Addison's disease (AD, 6 males, 2 females, age 30-77 years; last hydrocortisone administration the day before testing). In NS, HEX stimulated basal ACTH (peak, mean +/- SEM: 26.0 +/- 7.8 vs. 10.7 +/- 2.0 pg/ml, p < 0. 05), cortisol (163.2 +/ 18.3 vs. 137.4 +/- 15.4 microgram/l, p < 0.05) and GH (72.6 +/- 23.5 vs. 3.7 +/- 1.3 microgram/l, p < 0.01) levels. Metyrapone markedly increased basal ACTH (294.4 +/- 61.6 pg/ml, p < 0.05), reduced basal cortisol (19.6 +/- 7.2 microgram/l, p < 0.05), while it did not modify GH levels. After metyrapone pretreatment the ACTH response to HEX was clearly increased (DeltaAUC: 2,857.4 +/ 901.9 vs. 367.3 +/- 274.0 pg/ml/h, p < 0.05), while the GH response was not modified. HEX did not stimulate the low cortisol levels after metyrapone pretreatment. RU-486 significantly increased basal ACTH (76.6 +/- 12.5 pg/ml, p < 0.05) and cortisol (312.7 +/- 22.2 microgram/l, p < 0.05), while it did not modify basal GH levels. RU-486 pretreatment did not modify the ACTH, cortisol and GH responses to HEX. In AD, HEX elicited a marked ACTH response (6,619.4 +/- 3,365.8 pg/ml/h; p < 0.01), which was clearly higher (p < 0.01) than that in NS after HEX alone but not significantly different from that after HEX+MET. The GH response to HEX in AD (1,325.6 +/- 284.1 microgram/l/h) was similar to that in NS (1,519.7 +/- 483.8 microgram/l/h). In conclusion, our present data demonstrate that the ACTH-releasing activity of HEX is increased in primary hypoadrenalism as well as in normal subjects after metyrapone but not after RU-486 pretreatment. These findings indicate that in normal subjects as well as in hypocortisolemic patients the ACTH-releasing activity of GHS is enhanced by the lack of negative glucocorticoid feedback. PMID- 10516484 TI - Centrally administered murine leptin stimulates plasma arginine-vasopressin secretion and increases the level of mRNA expression in the supraoptic nucleus of conscious rats. AB - The product of the ob gene protein, leptin, has been suggested to function as an endogenous mediator of the cardiovascular system via sympathetic nerve activity. Moreover, extensive distribution of leptin receptor-like immunoreactivity has been demonstrated in the choroid plexus, cerebral cortex, hippocampus, thalamus and hypothalamus, especially in the paraventricular nucleus (PVN) and supraoptic nucleus (SON). In this study, we have investigated the in vivo effects of leptin on plasma arginine-vasopressin (AVP) secretion and the level of AVP messenger ribonucleotic acid (AVP mRNA) in the SON of conscious rats. Intracerebroventricularly administered leptin increased plasma AVP concentration in a dose-dependent manner (0-400 pmol/rat). The maximal effect was obtained at 15 min after the administration of leptin. Furthermore, in Northern blot analyses, the levels of AVP mRNa in the SON increased approximately 2-fold from the basal level after the administration of leptin. AVP mRNA expression in the PVN was also increased by leptin. However, leptin had no effects on plasma oxytocin (OXT) secretion and OXT gene expression in the SON. In conclusion, leptin is involved in AVP secretion via the central nervous system, however, its physiological role is unknown. PMID- 10516485 TI - Leptin(116-130) stimulates prolactin and luteinizing hormone secretion in fasted adult male rats. AB - Leptin is a hormone secreted by adipocytes with an important role in the control of feeding behavior and neuroendocrine function. Leptin stimulates in vivo LH secretion in fasted female rats and in vitro PRL secretion. Recent data indicate that leptin(116-130), an active fragment of the native molecule, exerts effects similar to those of the native peptide on body weight and food intake. The present study was carried out to determine whether this fragment is also able to stimulate LH and PRL secretion. Adult male rats fasted for 5 days were injected with saline or leptin(116-130) (15 microgram i.c.v.) and LH and PRL concentrations were measured thereafter at 15-min intervals during a 150-min period. Administration of leptin(116-130) increased the frequency of LH pulses (2.0 +/- 0.26 vs. 1.20 +/- 0. 37/150 min; p /=20 mm Hg (A1; B1) and patients with IOP <20 mm Hg (A2; B2). RESULTS: The mean difference of tonometry readings was equal to -0.36 +/- 1. 62 mm Hg in group A, 0.03 +/- 1.77 mm Hg in group A1, -0.61 +/- 1. 45 mm Hg in group A2, 0.23 +/- 1.44 in group B, 0.64 +/- 1.41 mm Hg in group B1, 0.05 +/- 1.42 in group B2. A statistically significant correlation was found in group A, in group A2, B, B1 and B2; a less significant correlation was found in group A1. CONCLUSIONS: The use of the latex caps does not alter the reliability of tonometry readings as long as the cap is applied tightly. Measurement variation in our study is comparable to published data on applanation tonometry. PMID- 10516514 TI - Ultrasound biomicroscopic evaluation of ciliochoroidal effusion after laser treatment. AB - We evaluated ciliochoroidal effusion (CE) by ultrasound biomicroscopy (UBM) following diode endophotocoagulation at the end of the vitreoretinal surgery. The aim of our study was to assess any differences in the CE morphology following diode endophotocoagulation or transpupillary krypton photocoagulation, and to demonstrate the influence of diabetes and intravitreal surgery on CE formation. Sixty-six consecutive patients were divided in to four groups. Twenty-nine patients with proliferative retinopathy underwent transpupillary krypton photocoagulation; 11 diabetic patients underwent vitreoretinal surgery and diode endophotocoagulation; 18 nondiabetics underwent vitreoretinal surgery and diode endophotocoagulation; 8 consecutive nondiabetic patients were the control group and underwent vitreoretinal surgery, without laser treatment. UBM was performed in the four groups before and after laser treatment, if performed. We determined, by UBM, not only the presence, but also the thickness of CE. CE was present in all the patients treated by laser, diabetics and nondiabetics, and its thickness was not correlated with the number of laser spots (p = 0.28). CE was seen ultrasonically in all the patients undergoing transpupillary photocoagulation or endophotocoagulation, regardless of diabetes and surgical trauma. PMID- 10516515 TI - Color doppler imaging of ocular blood flow after topical ketanserin. AB - The authors investigated the effect of ketanserin 0.5% eye drops on intraocular pressure and orbital blood flow in some healthy volunteers. Ketanserin is a selective antagonist of serotonin S(2) receptors; moreover, it produces an alpha(1)-blocking effect. It causes a systemic reduction of blood pressure and capillary vasodilatation. Three hours after topical administration of ketanserin 0.5%, the intraocular pressure was decreased by about 14% from the baseline. At the same time, we observed by color Doppler imaging that the mean percent increase in the peak systolic velocity in the ophthalmic artery (OA) was 5.18% (p < 0.161) and 23.18% (p < 0.001) in the posterior ciliary arteries (PCAs). The mean resistance index of the OA was in contrast reduced by 1.16% (p < 0.669) and by 32.14% (p < 0.003) in the PCAs. These data show that ketanserin 0.5% eye drops may be useful not only to decrease intraocular pressure but also to improve blood flow in the PCAs that supply the optic nerve head. PMID- 10516516 TI - The comparison of intraocular pressure reductions after isometric and isokinetic exercises in normal individuals. AB - The lowering effect of physical exercise on intraocular pressure (IOP) has been reported both in healthy people and those with glaucoma, but a comparison of the lowering effect of isometric and isokinetic exercises on IOP has not been conducted in any study. Our aims were to investigate the relationship between intensity of exercise and IOP, and whether a significant difference in IOP lowering effect existed between isometric and isokinetic exercises. Sixty-seven patients with an age range of 23-40 who had no ocular disease were randomly divided into two groups. While 31 patients in the first group, group A, performed isokinetic exercise with the Cybex 6000 dynamometer, 32 patients in the second group, group B, had isometric exercises with the same machine. IOP was measured in the right eye of patients with Shiotz tonometer just before and 10 min following exercise. Exercise intensity and total energy consumption were determined by the machine for each patient. While IOP values measured before exercise, the degree of exercise applied, and total energy consumption did not differ significantly between groups, both isometric and isotonic exercises lowered IOP significantly. As a result, isometric and isokinetic exercises lowered IOP in ophthalmologically normal subjects with direct relationship to exercise intensity and total energy consumption. Since the pressure lowering effect of isokinetic exercise was more significant, it might prove useful to glaucomatous patients. PMID- 10516517 TI - Refractive errors and ocular motility disorders in preterm babies with and without retinopathy of prematurity. AB - Ocular motility, refraction and visual acuity (VA) were evaluated at the age of 4 years in 136 preterm infants with gestational ages (GAs) at birth of less than 32 weeks. Group 1 (non-retinopathy of prematurity, ROP) included 87 children that had never developed ROP. Group 2 contained 19 children whose ROP had regressed spontaneously. Group 3 (cryo-ROP) was composed of 30 patients who had undergone cryotherapy for severe ROP. Strabismus was found in 13.9% of the total population. chi(2) analysis revealed that strabismus was significantly (p < 0.01) associated with prematurity (i.e. GA <29 weeks), ROP and cryotherapy. Myopia of more than 3 dpt was significantly (p < 0.001) more common in the cryo-ROP infants than in the regressed-ROP and non-ROP groups. The distribution of hypermetropia was similar in all three groups. VA was measured with the E chart. Of the 272 eyes examined, 251 (92.3%) displayed VA of more than 20/25. The majority of these eyes were from the non-ROP group (65.4%), 15.3% had regressed ROP and 21.1% belonged to the cryo-ROP group. Fifteen eyes (8 non-ROP, 3 regressed ROP and 4 cryo-ROP) presented VAs between 20/25 and 20/60. VA of less then 20/60 was found in 6 eyes (2 non-ROP, 1 regressed ROP, 3 cryo-ROP). Cryotherapy did not appear to preclude the development of good VA. PMID- 10516519 TI - Inflammatory response in experimental Staphylococcus and Pseudomonas endophthalmitis. AB - Staphylococcus epidermidis [2.0 x 10(4) colony-forming units (CFU)/0. 1 ml] and Pseudomonas aeruginosa (2.0 x 10(3) CFU/ml) were inoculated in the vitreous humor of rabbits. In S. epidermidis endophthalmitis, the numbers of microorganisms reached a maximum (4. 1 x 10(7) CFU/ml) at day 2 after inoculation and then declined spontaneously. However, clinical scores were observed to be worst at day 5. In P. aeruginosa endophthalmitis, the numbers of microorganisms reached a maximum (9.3 x 10(6) CFU/ml) 36 h after inoculation. However, culture results were persistently positive until day 15. Electroretinogram (ERG) b-wave amplitudes in S. epidermidis endophthalmitis continued to decrease from day 3 (>24%) until day 5, and then recovered to the preinoculative level of amplitudes at day 7. ERG b-wave amplitudes in P. aeruginosa endophthalmitis continued to decrease after 24 h (>24%). ERG b-wave amplitudes from day 7 to day 15 were flat. The inflammatory response continued under the absence of microorganisms in S. epidermidis endophthalmitis. The time in which a maximum in the number of microorganisms was reached was earlier than that in the clinical examination scores in both S. epidermidis and P. aeruginosa endophthalmitis. PMID- 10516518 TI - Secondary glaucoma in patients with uveitis. AB - PURPOSE: To evaluate the prevalence of secondary glaucoma (SG), clinical forms of uveitis more frequently associated with glaucoma, and describe the treatment and complications encountered in a cohort of patients with glaucoma and uveitis during a 10-year period. METHODS: The hospital records of patients with uveitis referred to the Immunology Service of the Massachusetts Eye and Ear Infirmary for a decade were reviewed for cases of SG. RESULTS: One hundred and twenty of the 1,254 patients (9.6%) with uveitis developed SG. SG was more frequent in anterior uveitis (67%) but was also associated with posterior uveitis (13%) and pars planitis (4%). Herpetic keratouveitis (22%), Fuchs' iridocyclitis (19%), juvenile rheumatoid arthritis-associated iridocyclitis (16%), syphilis (14%), and sarcoidosis (12%) were the leading types of uveitis associated with SG. Despite aggressive medical and surgical therapy, SG was associated with progressive visual field loss and optic nerve damage in 39 patients (33%). CONCLUSION: SG is an underappreciated, vision-threatening complication in patients with uveitis. Increased vigilance for emergence of this complicating problem during the care of patients with uveitis is warranted, and medical and surgical treatment for reducing IOP should be especially aggressive in these patients. We hypothesize that earlier, more aggressive treatment of uveitis will reduce the presence of glaucoma as an additional vision-robbing complication of uveitis. PMID- 10516520 TI - The effect of hydroxyurea on rabbit subconjunctival fibroblast culture and use of hydroxyurea in rabbits after glaucoma filtration surgery. AB - In an in vitro study, rabbit subconjunctival fibroblasts were cultured and the effects of an antineoplastic drug, hydroxyurea (HU), on fibroblast proliferation and fibroblast attachment was investigated. The effects of HU were compared with those of mitomycin C (MMC). Different concentrations of HU and MMC were added to culture medium. The HU doses which led to 50% of inhibition (ID(50)) and the dose which led to about 90% of inhibition (subtoxic high dose, STHD) were determined to be 8 and 1,000 microg/ml, respectively. ID(50) of MMC and its STHD which led to about 100% inhibition were found to be 0.01 and 1 microg/ml, respectively. Reversibility studies revealed that inhibition disappeared depending on the dose and incubation period of both HU and MMC. In an in vivo study, glaucoma filtration surgery (GFS) was performed in rabbits which were treated with HU (treatment group) and distilled water (control group). Tissue samples were taken from the subconjunctival area treated at 1 h, 1 day, 5 days and 30 days postoperatively. The biopsy specimens were then placed in tissue culture media. Fibroblast outgrowth rates detected in the HU group were found to be significantly lower than those in the control group in the specimens taken at the end of the first hour. The difference was significant on culture days 9-15 in the biopsy specimens taken on day 1 while it was not significant in those taken on days 5 and 30. PMID- 10516521 TI - Presumed congenital ocular toxoplasmosis in two successive siblings. AB - We present the cases of 2 consecutive siblings with bilateral macular lesions, for which there is strong clinical and laboratory evidence supporting the diagnosis of congenital ocular toxoplasmosis. These cases raise the possibility of maternal parasitemia during Toxoplasma gondii reinfection, leading to transmission to the fetus and congenital ocular toxoplasmosis despite prior immunity and lack of an immune disturbance in the mother. PMID- 10516522 TI - Bilateral granulomatous panuveitis as initial presentation of diffuse systemic T cell lymphoma. AB - A high-grade diffuse T cell lymphoma, initially simulating bilateral panuveitis, was diagnosed by analysis of a vitreous biopsy specimen and a breast tumor in a 57-year-old woman. It responded favorably to aggressive chemotherapy before it relapsed in leukemic transformation. This case emphasizes the misleading initial symptoms of primary intraocular lymphoma and the role of immunophenotyping in the diagnosis and classification of lymphoproliferative ocular disorders. The presentation and management of uveal lymphoid neoplasia are discussed. PMID- 10516523 TI - Das multifokale elektroretinogramm in der diagnostik und verlaufskontrolle lokalisierter Netzhautfunktionsstorungen: fallbericht eines patienten mit chorioretinopathia centralis serosa. AB - The role of multifocal electroretinography (MF-ERG) in the diagnosis and follow up of localized areas of retinal dysfunction is discussed. A 42-year-old male with the preliminary diagnosis of optic neuritis in his left eye was referred for evaluation with the MF-ERG. Simultaneous cone ERGs were obtained from 103 locations within the central 50 degrees of the retina. During an 8-month follow up four MF-ERGs were obtained. Bilaterally reduced paracentral response amplitudes contradicted the preliminary diagnosis. Subsequently central serous chorioretinopathy was diagnosed. Follow-up showed normalization of the MF-ERG responses in the left eye while retinal function in the right eye showed initial worsening. The noninvasive MF-ERG lends itself to follow-up in patients with central serous chorioretinopathy. PMID- 10516524 TI - Hypofluorescent spots on indocyanine green angiography at the recovery stage in multiple evanescent white dot syndrome. AB - PURPOSE: To evaluate the fundus lesions in a young woman. METHODS: Visual function, ophthalmoscopy, electrophysiology, fluorescein angiography, and indocyanine green angiography were performed. RESULTS: A 24-year-old woman had decreased visual acuity (0.2), granularity in the macula, and multiple yellow white patches in the fundus, reduced a wave on electroretinography, hyperfluorescence on fluorescein angiography, and hypofluorescence on indocyanine green angiography in the left eye. When visual acuity improved to 1.0, the white dots disappeared ophthalmoscopically, and fluorescein angiography showed normal findings. Hypofluorescent spots were found, however, on indocyanine green angiography. CONCLUSION: It is possible that signs of multiple evanescent white dot syndrome may remain longer during examination by indocyanine green angiography than by visual function, ophthalmoscopy, or fluorescein angiography. PMID- 10516525 TI - Natural interferon therapy: optic nerve ischemic damage? AB - The purpose of this study was the evaluation of retinal abnormalities during a treatment with natural interferon (IFN-alpha for chronic hepatitis C. Retinal hemorrhages and optic disk edema were found in a 40-year-old woman during IFN alpha therapy. The disk edema and retinopathy resolved after the INF was discontinued. Although retinal abnormalities correlated with IFN therapy have been described recently by some authors, the pathogenesis is still unclear. Anterior ischemic optic neuropathy occurring in a patient treated with IFN is a probable complication of the therapy. PMID- 10516526 TI - Postmortem findings two weeks after oral treatment for metastatic Candida endophthalmitis with fluconazole. AB - The purpose of this histological study was to present postmortem findings in both eyes of a 53-year-old male with liver dysfunction 2 weeks after short-time oral treatment with 200 mg/day fluconazole for metastatic Candida endophthalmitis due to intravenous hyperalimentation for 18 days. Candida had been demonstrated in the venous blood and on the tip of the intravenous catheter. The bilateral fungal endophthalmitis with hypopyon responded well to fungistatic therapy, but the patient suddenly died from heart failure. Both eyes were obtained at autopsy. Candida was demonstrated only in vitreous puff balls but not in the retina or uvea. Fluconazole administered for a short period had little effect in eliminating fungus from vitreous puff balls, which have no blood supply. Prolonged administration of the antifungal drug or vitrectomy should be considered when treating an eye with vitreous puff balls in the presence of fungal endophthalmitis. PMID- 10516527 TI - Sequential treatment: a new way of integrating pharmacotherapy and psychotherapy. PMID- 10516528 TI - Facilitating adjustment to inflammatory bowel disease: a model of psychosocial intervention in non-psychiatric patients. AB - BACKGROUND: There is no consensus about the most appropriate psychosocial interventions for people with inflammatory bowel disease (IBD) or the most appropriate criteria by which to select which patients might benefit from the available interventions. Nonetheless the perception that stress and other subjective factors contribute to suffering in IBD is persistent and professionals are often called upon to offer appropriate support. A model of normal psychosocial adjustment to IBD and the interventions which can improve difficulties with adjustment will facilitate rational therapeutic intervention and needed research in this area. METHODS: A model of normal adjustment to IBD is developed from a synthesis of the empirical literature and clinical experience in a tertiary care medical/surgical IBD centre and is used to identify potential points of psychosocial intervention. RESULTS: Normal adjustment to IBD can be understood as a process involving the interaction of a triad of adaptive challenges: illness uncertainty, loss and change, and suffering. Each of these challenges requires different criteria of psychosocial assessment and may lead to different interventions. CONCLUSIONS: Although the interventions available for improving adjustment to IBD have not been exhaustively investigated, the existing data support the value of further study. The model of psychosocial adjustment presented here provides a synthesis of the existing data and a starting point for further research. PMID- 10516529 TI - The Amsterdam Alexithymia Scale: its psychometric values and correlations with other personality traits. AB - BACKGROUND: This article describes the construction and validation of the Amsterdam Alexithymia Scale (AAS) and explores some of the nomological net of alexithymia. METHODS: Four correlational studies are presented. The internal structure of the AAS is explored by factor analyses on items. Correlations of the AAS with sex and (Guilford) intellectual abilities are established. Mean scores of three different professional groups are compared. Correlations between the AAS and several clinical and personality scales are determined. Students served as subjects in all studies (34738 degrees C) with pulmonary opacity on chest roentgenogram. The American Thoracic Society (ATS) criteria (1993) were used to decide on patient hospitalization. Ninety subjects, of whom 53 (59%) were men, with a mean age (+/ SD) of 38+/-15 years, were randomized: 45 received clarithromycin 500 mg b.i.d. orally for 14 days (CL group), and 45 received clarithromycin plus cefuroxime 500 mg b.i.d. orally for 7 days (CLCE group). Patients were monitored with clinical, radiological, and laboratory controls at 3 and 21 days. There were no significant differences between the two groups with regard to demographic, clinical, physical and laboratory data. RESULTS: The mean time to defervescence was 2.4+/-1.4 and 2.4+/-1.5 days, respectively. Chest roentgenogram clearance was complete in all cases, without statistically significant differences in the time to resolution between both arms. Side effects were mild (no significant differences between groups): 5 patients in the CL group and 3 in the CLCE group showed gastrointestinal symptoms. Two patients (2.2%), both in the CLCE group, needed hospital admission during follow-up, but all 90 patients showed an excellent outcome. A causative agent was determined in 25 cases (28%). Legionella pneumophila, Streptococcus pneumoniae, and Mycoplasma pneumoniae were the most frequent pathogens. CONCLUSION: Empirical treatment of outpatient CAP with clarithromycin can be considered adequate in the Mediterranean area, independently of the microbiological etiology. ATS criteria for admitting patients with CAP are appropriate in this population. PMID- 10516538 TI - Proinflammatory or regulatory cytokines released from BALF macrophages of healthy smokers. AB - BACKGROUND: Chronic smoking influences bronchoalveolar lavage fluid (BALF) cell profiles in healthy subjects, which may alter profiles of inflammatory and regulatory cytokines. OBJECTIVE: We focused on the evaluation of smoking-related changes in the amounts of cytokines released from BALF macrophages. METHODS: We measured the amounts of immunoreactive culture supernatants by using ELISA. RESULTS: The amounts of IL-1 receptor antagonist (IL-1ra) were lower in smokers [n = 10, median 22.1 ng/ml, 25th and 75th percentiles (18.7-39.1)] than in nonsmokers [n = 10, 48.6 (39.2-66.1), p = 0.010]. In smokers, lipopolysaccharide stimulation revealed decreases in the amounts of interleukin-6 (IL-6) [nonsmokers: 2.1 ng/ml (0.68-5.4 vs. smokers: 0.5 (0.03-0.87), p = 0.049] as well as IL-1ra [nonsmokers: 69.2 ng/ml (48.3-83.8) vs. smokers: 27.3 (17.2-56.7), p = 0.028]. A delay in release from intracellular storage was not the cause of the reduced amounts of IL-1ra. In addition, interleukin-1beta (IL-1beta) was positively correlated with IL-6 and granulocyte macrophage colony-stimulating factor in nonsmokers, but not in smokers. Furthermore, the decreases in IL-1ra and interleukin-8 were correlated with the increase in the number of BALF macrophages in smokers, but not in nonsmokers. CONCLUSIONS: Chronic smoking caused changes in the profiles of cytokines released from BALF macrophages in healthy subjects. Decreases in the amounts of regulatory cytokines, but not prominent changes in the amounts of inflammatory ones, were characteristic. PMID- 10516539 TI - Perception of dyspnea during histamine- and methacholine-induced bronchoconstriction. AB - BACKGROUND: Perception of dyspnea is poorly related to bronchoconstriction and may be influenced by distinct psychophysiologic stimuli. OBJECTIVE: This study compared the perceived psychophysiologic changes during histamine- and methacholine-induced bronchoconstriction using verbal as well as nonverbal assessment techniques. METHODS: Perception of dyspnea was studied during induced bronchoconstriction in 40 atopic subjects randomly ascribed to either histamine (n = 20) or methacholine (n = 20) bronchial challenge. A 100% increase in specific airway resistance (sR(aw)) indicated airway hyperresponsiveness (AHR). Dyspnea was verbally assessed by the Borg Scale (BS) and the Asthma Symptom Checklist (ASL). A hand dynamometer (HD) served for nonverbal assessment. Both challenge groups did not differ significantly with respect to age, anthropometric data, smoking and lung function before challenge. RESULTS: AHR did not differ between groups but groups differed significantly with respect to the number of symptoms and to symptom intensity reported after challenge. Subjects who underwent the histamine challenge scored significantly higher on both measures derived from the ASL. BS ratings and HD scores correlated significantly but were not significantly related to the degree of AHR. Accurate and poor perceivers could be discriminated by analysis of the relationship between BS and sR(aw). CONCLUSIONS: These findings suggest that perception of induced dyspnea differs between histamine and methacholine when assessed by a symptom report. PMID- 10516540 TI - Formoterol and salmeterol in partially reversible chronic obstructive pulmonary disease: A crossover, placebo-controlled comparison of onset and duration of action. AB - BACKGROUND: In contrast to the well-known activity profile in asthma, the precise efficacy and optimum dose schedules of long-acting beta(2)-agonists in chronic obstructive pulmonary disease (COPD) are not clear. OBJECTIVE: In this study, we aimed to compare the onset and the duration of action of a single inhalation of formoterol and salmeterol in COPD patients having partially reversible airway obstruction. METHODS: In a double-blind, randomized, crossover and placebo controlled study design, the respiratory functions of 22 patients (mean age 57.3+/-5.4 years) having mild to severe COPD (5 mild, 8 moderate and 9 severe) and partially reversible airway obstruction [mean baseline reversibility of forced expiratory volume in 1 s (FEV(1)) 19.3+/-3.1%] were evaluated after inhalation of 12 microg formoterol and 50 microg salmeterol. RESULTS: Regarding the onset of bronchodilator action, the mean absolute increase of 0.20 liters in FEV(1) 10 min after inhalation of formoterol was significantly higher than baseline and that of placebo (0.04 liters), whereas that of salmeterol (0.11 liters) did not reach statistical significance. At 20 min, both formoterol (0.25 liters) and salmeterol (0.20 liters) produced a significant increase in FEV(1) compared with baseline and with that of placebo (0.04 liters). The peak bronchodilator effects occurring at 60 and 120 min following formoterol (0.39 liters) and salmeterol (0.40 liters) inhalation, respectively, were significantly higher than the corresponding levels of placebo (0.02 and -0.12 liters, respectively). Concerning the duration of action, the 12-hour values of both formoterol (0.25 liters) and salmeterol (0.22 liters) were significantly higher than that of placebo (-0.12 liters). The area under the curve values of FEV(1) of formoterol (3.5+/-1.3 l.h) and salmeterol (3.2+/-1.2 l x h) averaged over 12 h were comparable and higher than placebo values (1.2+/-0.5 l x h). After formoterol inhalation 2 patients experienced tremor and 1 had palpitation; 1 tremor and 1 headache attack were noted after salmeterol. For the pharmacologically predictable side effects, there was no difference between the drugs. CONCLUSIONS: In conclusion, this study revealed that a single dose of 12 microg formoterol and 50 microg salmeterol provided comparable bronchodilation within 12 h and had tolerable side effects in patients with mild to severe COPD having partially reversible airway obstruction. PMID- 10516541 TI - German version of the Epworth Sleepiness Scale. AB - BACKGROUND: The Epworth Sleepiness Scale (ESS) is a questionnaire widely used in English speaking countries for assessment of subjective daytime sleepiness. OBJECTIVE: Our purpose was to translate and validate the ESS for use in German speaking countries. METHODS: A German translation of the ESS was administered to 159 healthy German-speaking Swiss and to 174 patients with various sleep disorders. RESULTS: The mean +/- SD of ESS scores in normals was 5.7+/-3.0, in patients it was 13.0+/-5.1 (p<0.001). Scores were not correlated with age or gender but with the percentage of time spent at an oxygen saturation <90% (R = 0.35, p<0.001), and the respiratory disturbance index (R = 0.26, p<0.001) in primary snorers and sleep apnea patients. Item analysis confirmed internal consistency of the scale (Cronbach alpha = 0.60 in normals, and 0.83 in patients). Follow-up scores in 25 sleep apnea patients on treatment showed a reduction by 7+/-5 points (p<0.05). CONCLUSIONS: Our data validate the ESS for application in German-speaking populations. The simplicity, reliability and the apparent lack of relevant influences of language and cultural background on performance of the ESS makes it a valuable tool for clinical management and research. PMID- 10516542 TI - Effect of nitric oxide synthase inhibition on hemoglobin-oxygen affinity and lipid peroxidation in rabbits during fever. AB - BACKGROUND: Nitric oxide (NO) is one of the most important biologic messengers and takes part in the development of fever. It can influence on the body prooxidant-antioxidant balance by different ways including interaction with hemoglobin (Hb). METHODS: The effects of nitric oxide synthesis inhibition on the febrile response, hemoglobin-oxygen affinity and parameters of lipid peroxidation were studied in rabbits with fever. The fever was induced by intravenous administration of lipopolysaccharide from Salmonella typhi (0.6 microg/kg). Mixed venous blood was taken before the administration and 60, 120 and 180 min after it. The following parameters were measured: half-saturation oxygen pressure (P(50)), concentrations of conjugated dienes, Schiff bases and alpha-tocopherol in plasma and red blood cells, and activity of catalase in red blood cells. RESULTS: The intravenous administration of the nitric oxide synthase inhibitor (N(omega)-nitro-L-arginine; 5x10(-3) M) reduced the lipopolysaccharide-induced rise in body temperature. After 180 min the actual P(50) had decreased from 35.0+/-1.7 to 29.4+/-1.3 mm Hg. An increase in the lipid peroxidation parameters and a decrease of the antioxidant system indices were observed. The administration of L-arginine to prevent nitric oxide synthase inhibition was accompanied by a shift of the oxyhemoglobin dissociation curve rightwards, more marked activation of the free radical processes and a greater elevation of body temperature. The multiple regression analysis showed a close linear correlation between P(50) and conjugated dienes, Schiff bases, alpha-tocopherol and catalase. CONCLUSION: These results suggest that the increased hemoglobin-oxygen affinity found after the inhibition of nitric oxide synthesis lowers the oxygen flow to tissues and its fraction utilized in free radical oxidations, which finally causes a reduction of the fever response to the lipopolysaccharide. PMID- 10516543 TI - Effect of immunomodulators on bleomycin-induced lung injury. AB - BACKGROUND: The role of lymphocytes and their subpopulations in lung fibrosis is as yet unclear. OBJECTIVE: To define the role of immunomodulation in bleomycin induced inflammatory fibrotic lung injury, by testing the effect of two known Th1 inhibitors: linomide and pentoxifylline. METHODS: C57BL/6 mice were treated by a single intratracheal instillation of 0.06 mg bleomycin in 0.01 ml saline or saline alone. Treatment groups included: (1) intratracheal bleomycin and daily treatment with linomide or pentoxifylline; (2) intratracheal bleomycin and daily water; (3) intratracheal saline and daily linomide or pentoxifylline; (4) intratracheal saline and daily water. Linomide and pentoxifylline were available per os in the drinking water from 1 day prior to intratracheal instillation. Animals were studied 14 days after intratracheal instillation. Lung injury was evaluated by total and differential cell count in bronchoalveolar lavage fluid, by a semiquantitative morphological index of lung injury and a quantitative image analysis of cellularity, fibrosis fraction and alveolar wall area fraction, and by biochemical analysis of lung hydroxyproline content. RESULTS: Linomide or pentoxifylline did not cause any lung injury in saline-treated control mice. Overt signs of lung injury were apparent in bleomycin-treated mice. These changes were not affected by daily treatment with linomide or pentoxifylline, which were given in the highest tolerable dose. CONCLUSION: This study does not support the use of linomide or pentoxifylline to prevent or ameliorate lung fibrosis and may suggest that drug-induced differentiation of T lymphocytes into Th1/th2 subpopulations does not affect the evolution of bleomycin-induced lung injury. PMID- 10516544 TI - Scoliosis in Duchenne muscular dystrophy. PMID- 10516545 TI - Mediastinal fibromatosis presenting with superior vena cava syndrome. AB - We encountered a fatal case of mediastinal fibromatosis in a 67-year-old female in whom there was aggressive infiltration into the large vessels, nerves and pericardium. She presented with the superior vena cava syndrome, Horner's syndrome, paralysis of bilateral vocal cords and diaphragm and heart failure. Mediastinoscopical examination revealed an extremely firm tumor adhering to the sternum, trachea and brachiocephalic artery. She died of severe heart failure due to the disturbed dilatation of the heart and ventilatory insufficiency. Although mediastinal fibromatosis is very uncommon and sometimes difficult to diagnose at an early stage, physicians should be aware of this disease for the differential diagnosis of mediastinal tumors. PMID- 10516546 TI - Crohn's disease mimicking sarcoidosis in bronchoalveolar lavage. AB - Granulomatous disorders like sarcoidosis or Crohn's disease are commonly associated with extrapulmonary or extraintestinal manifestations which occasionally may represent the only symptoms. We describe a 28-year-old female patient suffering from atypical erythema nodosum and arthritis. Although the chest x-ray was unremarkable bronchoalveolar lavage revealed lymphocytic alveolitis with an elevated CD4/CD8 ratio of 8 and 11.4 at repeated examinations suggesting a diagnosis of sarcoidosis. Further diagnostic workup included endoscopy of the bowel. The macroscopic aspect and histology of the terminal small bowel and colon ascendens indicated Crohn's disease. The patient recovered on steroids and sulfasalazine. Six months later she developed a perianal abscess for which she needed surgery supporting the diagnosis of Crohn's disease. This is the first case of a significantly (>6) elevated CD4/CD8 ratio in Crohn's disease previously regarded as highly specific for sarcoidosis. PMID- 10516547 TI - Expression of CA125 in thoracic endometriosis in a patient with catamenial pneumothorax. AB - A 40-year-old woman had experienced monthly right thoracic pain and productive cough occurring at the beginning of her menstrual period. X-ray findings indicated a diagnosis of catamenial pneumothorax. The serum CA125 level was very high at 159.6 U/ml. Thoracoscopy showed multiple dark cherry-colored nodules with neovascularization on the diaphragm. Following partial resection of the diaphragm thoracic endometriosis was diagnosed. Immunohistochemical staining of these endometrial cells showed antibodies to CA125. She has been well without recurrence for 15 months, and her serum CA125 level was within the normal range after operation. PMID- 10516549 TI - Chronic lip ulceration in association with an abnormal chest radiograph. PMID- 10516548 TI - Chronic cough due to bronchobiliary fistula. AB - Bronchobiliary fistula is a rare cause of chronic cough. Here we describe a 70 year-old woman complaining of chronic cough and copious dark-yellow watery sputum. The presence of air in the biliary tract in the lower cuts of a computerized tomography scan of the chest and positive bile in the sputum led to the suspicion of bronchobiliary fistula. The diagnosis was confirmed by percutaneous transhepatic cholangiography. Drainage of the intrahepatic biliary tract resulted in complete resolution of her symptoms. PMID- 10516550 TI - Oversight and monitoring of blood safety in the United States. AB - The US blood safety vigilance system is composed of a network of interwoven programs, now organized under a formal structure, with the Assistant Secretary of Health and DHHS Blood Safety Committee bearing overall responsibility. It takes advantage of the breadth of expertise and close collaborative relationship of transfusion medicine and infectious disease scientists within and outside of the government. Core elements include an array of ongoing surveillance programs for monitoring established as well as new and emerging infectious agents that may pose a risk to blood safety, and the existence of historical and contemporary repositories of donor and recipient specimens that enable rapid investigation of putative new risks. This report summarizes the historical events that shaped the US blood safety oversight system, reviews the current organization and decision making processes related to blood safety issues, and highlights key surveillance systems and research programs which monitor the US and global blood supplies for known and potential emerging risks. PMID- 10516551 TI - The French haemovigilance system. AB - Haemovigilance is a national system of surveillance and alarm, from blood collection to the follow-up of the recipients, gathering and analysing all untoward effects of blood transfusion in order to correct their cause and prevent recurrence. In France haemovigilance was created by law and notification of transfusion incidents is a legal obligation. The haemovigilance network associates local correspondents in each hospital and blood centre with regional co-ordinators and is centralised by the Agence Francaise du Sang. After 4 years the incident reporting rate is 2.3 per 1,000 allogeneic blood components transfused, justifying for example increased efforts in the prevention of bacteria-associated transfusion reactions, haemolytic transfusion reactions or vascular overload. However, haemovigilance still has to be strenghtened by improved information management or further progress in standardisation from one region to the other. The most important factor of success is collaboration between blood centres and hospitals. Haemovigilance clearly is the ultimate quality indicator of a transfusion service. PMID- 10516552 TI - Systems contributing to the assurance of transfusion safety in the United Kingdom. AB - In 1996, the United Kingdom launched a voluntary 'haemovigilance' system for confidential reporting of transfusion-related deaths and major adverse events. The Serious Hazards of Transfusion (SHOT) initiative provided the first comprehensive overview of transfusion safety in the UK, with 12 fatalities reported in the first year. The most important finding was that of a total of 169 reports, 47% were 'wrong blood to patient' episodes, of which 16 were ABO incompatible. There were eight transfusion-transmitted infections, three bacterial, four viral and one malarial. A number of other initiatives exist in the UK which also have importance in contributing to transfusion safety. This article reviews these other key contributors, allowing SHOT to be placed in context. PMID- 10516553 TI - Application of a time-resolved fluoroimmunoassay for the analysis of normal prion protein in human blood and its components. AB - BACKGROUND AND OBJECTIVES: To quantify the cellular isoform of prion protein (PrP(c)) in human blood using a new time-resolved dissociation-enhanced fluoroimmunoassay (DELFIA). MATERIALS AND METHODS: The DELFIA was optimised for human blood samples and applied to isolated cell and plasma fractions from blood donations. The physicochemical properties of PrP(c) were analysed. RESULTS: 26. 5% of blood PrP(c) was associated with the platelet fraction, 0.8% with polymorphonuclear leucocytes, 2.4% with mononuclear leucocytes, 1.8% with red cells and 68.5% with plasma (mean values from 4 processed donations). CONCLUSION: The majority of blood PrP(c) is found in the platelet and plasma compartments. PMID- 10516554 TI - Differences in residual white blood cell subset counts in buffy coat-depleted red cell concentrates prepared with bottom and top or quadruple-bag systems. AB - BACKGROUND: For preparation of buffy coat-depleted red cell concentrates (RCCs) in additive solution whole blood is currently collected in The Netherlands both in quadruple-bag and bottom and top bag systems. By using the quadruple-bag system both plasma and buffy coat cells are transferred into integrated satellite bags while the red cells remain in the collection bag. When bottom and top bags are used, the buffy coat remains in the collection bag while both red cells and plasma are transferred into satellite bags. The difference in processing prompted us to perform quantitative analysis of residual WBC subsets in buffy coat depleted RCCs. Differences in removal of specific cells with the buffy coat could improve the outcome of additional filtration procedures aiming at complete removal of specific WBC subsets. STUDY DESIGN AND METHODS: The buffy coat was removed in semiautomated procedures (Optipress I; Compomat G4) from units of whole blood collected in both bag systems. Paired samples were taken before and after removal of the buffy coat for counting and analyzing WBC subsets by flow cytometry using subset-specific monoclonal antibodies. RESULTS: All RCCs met the criteria from the guidelines of the Council of Europe. The percentage of residual total WBCs was lower (p<0.001) in RCCs processed in bottom and top bag systems (26% Compomat and 18% Optipress) as compared to RCCs processed in quadruple-bag systems (43% Compomat). WBC subset analysis in RCCs processed in quadruple-bag systems showed approximately 25% of residual T cells, B cells and monocytes and 60% of residual granulocytes. In contrast, WBC subset analysis in RCCs processed in bottom and top bag systems showed approximately 2% residual T cells, B cells, and monocytes and 35% residual granulocytes; in about 45% of units, lymphocytes and monocytes were even below the detection limit of flow cytometry analysis. CONCLUSION: Buffy coat-depleted RCCs are currently processed in bottom and top bag or quadruple-bag systems, the former being superior to the latter due to selective depletion of lymphocytes and monocytes by 98% (2 logs). The bottom and top procedure is an evident contribution to leukodepletion in blood transfusion, both with and without additional filtration. PMID- 10516555 TI - An international trial demonstrates suitability of a newly developed whole-blood ELISA kit for multicentre platelet HPA-1 phenotyping. AB - BACKGROUND: Neonatal alloimmune thrombocytopenia (NAIT) is in most cases due to pregnant women with HPA-1b platelet phenotype producing antibodies to HPA-1a platelets of the fetus, which may lead to intracranial haemorrhage with subsequent death or life-long morbidity. The availability of sensitive, reliable, straightforward, and inexpensive assays would enable large-scale screening in pregnancy and may help avoid NAIT. METHODS: A recently developed enzyme-linked immunosorbent assay (ELISA) was produced in kit form incorporating modified reagents and enabling distribution to 21 international Platelet Immunology and Blood Centres (see Acknowledgements). The kits were assessed using anticoagulated whole-blood samples stored up to 10 weeks at 4 degrees C. Each centre tested its own blood samples most of which had been previously typed by established assays. RESULTS: Of the 152 samples that were tested, all 31 HPA-1b phenotypes were correctly identified by the kit. The respective mean +/- standard deviation of the specific absorbances of HPA-1b and HPA-1a samples were: 0.52+/-0.15 and 0.12+/-0.06 (p<0.0005). In addition, the modified ELISA showed 100% concordance with PCR-SSP in the phenotyping of 93 donors. Finally, a comparison between freshly prepared and stored kits showed that the kit was stable for at least 2 years at 4 degrees C. CONCLUSIONS: The international trial showed that the modified whole-blood ELISA kit is very well suited for wide-scale screening in pregnancy. Moreover, the ELISA kit could be used for large-scale phenotyping of blood donors, with HPA-1b-typed individuals getting invited to become apheresis platelet donors for patients with NAIT. PMID- 10516556 TI - Prevalence of human parvovirus B19 DNA and IgG/IgM antibodies in Polish haemophilia patients. PMID- 10516557 TI - Determination of IgG subclasses 1-4 in plasmapheresis donors. PMID- 10516558 TI - Threshold concentration of haemoglobin in donor blood. PMID- 10516559 TI - International reference reagents: antihuman globulin. An ISBT/ICSH joint working party report. International Society of Blood Transfusion. International Committee for Standardization in Haematology. AB - An international working party has conducted a study designed to select a suitable reference reagent for antihuman globulin, to replace those first made available in 1987. The chosen preparation contains levels of anti-IgG and anti-C3 (anti-C3c and anti-C3d) potency that are considered suitable to serve for reference when evaluating either polyspecific antihuman globulin reagents or those containing their separate monospecific components. The reference material is available in 2-ml freeze-dried aliquots from seven assigned distribution centres. PMID- 10516560 TI - Mitochondrial DNA variation is an indicator of austronesian influence in Island Melanesia. AB - Past studies have shown a consistent association of a specific set of mitochondrial DNA 9 base pair (bp) deletion haplotypes with Polynesians and their Austronesian-speaking relatives, and the total lack of the deletion in a short series of New Guinea Highlanders. Utilizing plasma and DNA samples from various old laboratory collections, we have extended population screening for the 9-bp deletion into "Island Melanesia," an area notorious for its extreme population variation. While the 9-bp deletion is present in all Austronesian, and many non Austronesian-speaking groups, it is absent in the more remote non-Austronesian populations in Bougainville and New Britain. These results are consistent with the hypothesis that this deletion was first introduced to this region about 3,500 years ago with the arrival of Austronesian-speaking peoples from the west, but has not yet diffused through all populations there. The pattern cannot be reconciled with the competing hypothesis of a primarily indigenous Melanesian origin for the ancestors of the Polynesians. Although selection clearly has operated on some other genetic systems in this region, both migration and random genetic drift primarily account for the remarkable degree of biological diversity in these small Southwest Pacific populations. PMID- 10516561 TI - Distribution of mtDNA haplogroup X among Native North Americans. AB - Mitochondrial DNA (mtDNA) samples of 70 Native Americans, most of whom had been found not to belong to any of the four common Native American haplogroups (A, B, C, and D), were analyzed for the presence of Dde I site losses at np 1715 and np 10394. These two mutations are characteristic of haplogroup X which might be of European origin. The first hypervariable segment (HVSI) of the non-coding control region (CR) of mtDNA of a representative selection of samples exhibiting these mutations was sequenced to confirm their assignment to haplogroup X. Thirty-two of the samples exhibited the restriction site losses characteristic of haplogroup X and, when sequenced, a representative selection (n = 11) of these exhibited the CR mutations commonly associated with haplogroup X, C --> T transitions at np 16278 and 16223, in addition to as many as three other HVSI mutations. The wide distribution of this haplogroup throughout North America, and its prehistoric presence there, are consistent with its being a fifth founding haplogroup exhibited by about 3% of modern Native Americans. Its markedly nonrandom distribution with high frequency in certain regions, as for the other four major mtDNA haplogroups, should facilitate establishing ancestor/descendant relationships between modern and prehistoric groups of Native Americans. The low frequency of haplogroups other than A, B, C, D, and X among the samples studied suggests a paucity of both recent non-Native American maternal admixture in alleged fullblood Native Americans and mutations at the restriction sites that characterize the five haplogroups as well as the absence of additional (undiscovered) founding haplogroups. PMID- 10516563 TI - First rib metamorphosis: its possible utility for human age-at-death estimation. AB - Human first ribs demonstrate predictable, sequential changes in shape, size, and texture with increasing age, and thus, can be used as an indicator of age at death. Metamorphosis of the first rib's head, tubercle, and costal face was documented in a cross-sectional sample of preadult and adult first ribs of known age at death from the Hamann-Todd skeletal collection (Cleveland Museum of Natural History, Cleveland, Ohio). Blind tests of the usefulness of the first rib as an age indicator were conducted, including tabulation of intraobserver and interobserver inaccuracies and biases. First rib age estimates show inaccuracies and biases by decade comparable to those generated by other aging techniques. Indeed, the first rib method is useful as an isolated age indicator. When used in conjunction with other age indicators, the first rib improves the quality of summary age assessments. PMID- 10516562 TI - Looking into the demography of an iron age population in the western Mediterranean. I. Mortality. AB - In this paper, we attempt to reconstruct the mortality pattern of the population buried in S'Illot des Porros (Majorca), an Iron Age necropolis in the western Mediterranean, by means of paleodemographic analysis. The skeletal sample consists of 285 individuals, 93 subadults (under 20 years old) and 192 adults. The aim of this study is twofold: first, to identify and to evaluate the structural anomalies of the skeletal sample, and second, to obtain a possible and realistic description of the biological dynamics of this population, with special reference to its mortality pattern. The study uses current demographic methodology and several demographic models (for comparison). An abridged life table was built to estimate the mortality parameters. To evaluate the likelihood of the estimated data, an indirect analysis, which consisted of a comparison of our results with different population models (Weiss [1973] American Antiquity 38; Coale and Demeny [1996] Regional Model Life Tables and Stable Populations. Princeton: Princeton University Press; Ledermann [1969] Nouvelles tables-types de mortalite. Paris: Presses Universitaires de France), was carried out. An important bias was identified in the case of children, mostly affecting infants but also children between the ages of 1 and 5. This was interpreted as a census error due to taphonomic reasons and to an excluding differential funeral rite. A life expectancy at birth of approximately 28 years was estimated from the observed data. When this bias was removed, the estimated life expectancy at birth dropped to 23 years. The use of the Brass logit system allowed us to sketch a possible mortality profile for this population: low life expectancy, high infant mortality and hard life conditions, which were the cause of the low levels of survivorship in old ages. Am J Phys Anthropol 110:285-301, 1999. PMID- 10516564 TI - The ecological role of the prehensile tail in white-faced capuchins (Cebus capucinus). AB - Prehensile tails appear to have evolved at least twice in platyrrhine evolution. In the atelines, the tail is relatively long and possesses a bare area on the distal part of its ventral surface that is covered with der-matoglyphs and richly innervated with Meissner's corpuscles. In contrast, the prehensile tail of Cebus is relatively short, fully haired, and lacks specialized tactile receptors. Little is currently known regarding tail function in capuchins, and whether their prehensile tail serves a greater role in feeding or traveling. In this paper we examine patterns of positional behavior, substrate preference, and tail use in wild white-faced capuchins (Cebus capucinus) inhabiting a wet tropical forest in northeastern Costa Rica. Observational data were collected over the course of 3 months on adult capuchins using an instantaneous focal animal time sampling technique. Differences in the frequency and context of tail use, and the estimated amount of weight support provided by the tail relative to other appendages during feeding/foraging and traveling were used as measures of the ecological role of this specialized organ in capuchin positional behavior. During travel, quadrupedal walking, leaping, and climbing dominated the capuchin positional repertoire. The capuchin tail provided support in only 13.3% of travel and was principally employed during below branch locomotor activities. In contrast, tail-assisted postures accounted for 40.6% of all feeding and foraging records and occurred primarily in two contexts. The tail was used to suspend the individual below a branch while feeding, as well as to provide leverage and weight support in above-branch postures associated with the extraction of prey from difficult to search substrates. A comparison of tail use in Cebus, with published data on the atelines indicates that both taxa possess a grasping tail that is capable of supporting the animal's full body weight. In capuchins and howling monkeys, the tail appears to be used more frequently and serves a greater weight-bearing role during feeding than during traveling. In Ateles, and possibly Brachyteles, and Lagothrix, however, the prehensile tail serves a dual role in both feeding and forelimb suspensory locomotion. Additional relationships between white-faced capuchin feeding, positional behavior, extractive foraging techniques, and prehensile tail use are discussed. PMID- 10516565 TI - Bilateral asymmetry in skeletal growth and maturation as an indicator of environmental stress. AB - This study examined the efficacy of bilateral asymmetry in epiphyseal union as an indicator of environmental stress affecting the skeleton. We compared the extent of asymmetry in the postcranial skeleton between two cemetery samples excavated from Medieval Kulubnarti, Sudanese Nubia. Past studies have strongly suggested that these ancient Nubians experienced environmental stress-the early Christian period (550-750 AD) population to a greater extent than the late Christian period (750-1450 AD) population. We hypothesized that if bilateral asymmetry is a reflection of stress, then it should be present or greater in the more stressed population, the early Christian period population, while absent or found to a lesser extent in the less stressed population, the late Christian period population. We computed two mean values, representative of right-side and left side epiphyseal union, for each individual in both cemetery samples, and tested for significant differences. Bilateral asymmetry was significant in the combined cemetery sample of 90 individuals (P < 0.019). When cemetery samples were tested separately, bilateral asymmetry was significant for the early Christian period sample (P < 0.001), but not for the late Christian period sample. There were no differences attributable to sex. Finally, we discuss why we conclude that environmental stress was favored over a biomechanic explanation as the cause for asymmetry. To the extent that our results support previous findings that early Christian period individuals were more affected by environmental stress than late Christian period individuals, it is reasonable to consider bilateral asymmetry in skeletal growth and maturation a good indicator of environmental stress. PMID- 10516566 TI - Enamel hypoplasia in deciduous teeth of great apes: do differences in defect prevalence imply differential levels of physiological stress? AB - This paper presents new data on enamel hypoplasia in the deciduous canine teeth of great apes. The enamel defect under consideration is known as localized hypoplasia of primary canines (LHPC), and is characterized by an area of thin or missing enamel on the labial surface of deciduous canine teeth (Skinner [1986a] Am. J. Phys. Anthropol. 69:59-69). Goals of this study are: 1) to determine if significant differences in the frequency of LHPC occur among three genera of great apes, and 2) to evaluate variation in LHPC prevalence among great apes as evidence of differential physiological stress. Infant and juvenile apes with deciduous teeth were examined at the Cleveland Museum of Natural History (n = 100) and at the Smithsonian Institution, National Museum of Natural History (n = 36). Deciduous teeth were observed under oblique incandescent light, with the naked eye and with a 10x hand lens. Enamel hypoplasia was scored using Federation Dentaire International (FDI)-Defects of Dental Enamel (DDE) standards. Hypoplasias were recorded by drawing defect location and size on a dental chart, and by measuring defect size and location with Helios needlepoint dial calipers. The prevalence of LHPC is reported by genus and sex, using two approaches: 1) the frequency of affected individuals-those having one or more deciduous canine teeth scored positive for LHPC; and 2) the number of canine teeth scored positive for LHPC as a percentage of all canine teeth observed. Variation in defect size and location will be described elsewhere. Localized hypoplasia of primary canine teeth was found in 62.5% of 128 individual apes, and in 45.5% of 398 great ape deciduous canines. As in humans, LHPC is the most common form of enamel hypoplasia in deciduous teeth of great apes, while LEH is rare or absent. The distribution and pattern of expression of LHPC in great apes is similar to that described in humans: side differences are not significant, but mandibular canines exhibit the defect two to five times more often than maxillary canine teeth. Differences in LHPC prevalence by sex are small and not significant. Intergeneric differences are large and non-random: chimpanzees (Pan) exhibit a significantly lower frequency of LHPC (22%, n = 50) by individual count, than either the orangutan (Pongo, 88.0%, n = 25) or the gorilla (Gorilla, 88.7%, n = 53). Tooth count prevalences exhibit a similar pattern of variation and are also statistically significant. These findings suggest that large bodied great apes (gorilla and orangutan) may be under greater physiological stress during perinatal and early postnatal development than the chimpanzee. The size, position, and timing of LHPC lesions are currently under analysis and may yield more insight into the etiological origin of this enamel defect. PMID- 10516567 TI - Ontogeny and phylogeny of femoro-tibial characters in humans and hominid fossils: functional influence and genetic determinism. AB - Three different human femoro-tibial characters are selected as functionally relevant and derived hominid characters: femoral bicondylar angle, shape of the femoral distal epiphysis, and the tibial insertion of the lateral meniscus. The timing and mode of formation of these characters are investigated during human ontogeny and are shown to differ considerably. The available hominid fossils (Australopithecus afarensis and early Homo) are interpreted in the light of this ontogenetic analysis with the conclusion that, during hominid evolution, different modes of selection of these features must have occurred. In modern humans, the femoral bicondylar angle proves to be an epigenetic functional feature, which develops during early childhood growth. It is present in all australopithecines and we suggest that it developed following a change in their locomotor behavior and not upon a genomic change: the early practice of bipedal walking, with adducted knee joints, in the locomotor repertoire of infant australopithecines, was sufficient to promote this angle. Later in hominid evolution, the knee joint evolved from having a single insertion of the lateral meniscus on the tibia to a double one. While Australopithecus afarensis exhibits a single insertion, early Homo clearly exhibits a double insertion of the lateral meniscus on the tibia. The double insertion restricts the mobility of the meniscus on the tibial plateau, indicating a habitual practice of full extension movements of the knee joint. Among modern humans, the posterior insertion of the lateral meniscus appears early in fetal life. Consequently in early Homo, this new selected feature developed directly as a result of a genomic change. The derived shape of human distal femoral epiphysis includes a prominence of the lateral lip of the femoral trochlea, an elliptical profile of the lateral condyle, and an anteroposterior lengthening of the epiphysis. Analysis of human fetal and neonatal distal epiphyses shows that the prominence of the lateral lip of the trochlea arises before any use, and thus appears to be genetically determined. However, the postnatal development of this joint shows that this feature is also modified epigenetically by use. It is argued that the hominid femoro-patellar joint would have been reshaped following the process of genetic assimilation (Waddington [1942] Nature 3811:563-565). The prominence of the lateral lip of the femoral trochlea was probably selected following a two-staged process-first epigenetic, then genetic. Far from being a Lamarckian explanation, this concept applies precisely to adaptive characters that are induced by an external stimulus during a single lifetime and are replaced through natural selection by genetically based equivalent characters. The nature of the structures involved in the studied features is shown to be an important parameter determining their mode of development and selection. PMID- 10516569 TI - NMR analysis of peptide structure and bioactivity PMID- 10516568 TI - The anomalous archaic Homo femur from Berg Aukas, Namibia: a biomechanical assessment. AB - The probably Middle Pleistocene human femur from Berg Aukas, Namibia, when oriented anatomically and analyzed biomechanically, presents an unusual combination of morphological features compared to other Pleistocene Homo femora. Its midshaft diaphyseal shape is similar to most other archaic Homo, but its subtrochanteric shape aligns it most closely with earlier equatorial Homo femora. It has an unusually low neck shaft angle. Its relative femoral head size is matched only by Neandertals with stocky hyperarctic body proportions. Its diaphyseal robusticity is modest for a Neandertal, but reasonable compared to equatorial archaic Homo femora. Its gluteal tuberosity is relatively small. Given its derivation from a warm climatic region, it is best interpreted as having had relatively linear body proportions (affecting proximal diaphyseal proportions, shaft robusticity, and gluteal tuberosity size) combined with an elevated level of lower limb loading during development (affecting femoral head size and neck shaft angle). PMID- 10516570 TI - Peptide structural analysis by solid-state NMR spectroscopy. AB - Solid-state nmr spectroscopy provides a robust method for investigating polypeptides that have been prepared by chemical synthesis and that are immobilized by strong interactions with solid surfaces or large macroscopic complexes. Solid-state nmr spectroscopy has been widely used to investigate membrane polypeptides or peptide aggregates such as amyloid fibrils. Whereas magic angle spinning solid-state nmr spectroscopy allows one to measure distances and dihedral angles with high accuracy, static membrane samples that are aligned with respect to the magnetic field direction allow one to determine the secondary structure of bound polypeptides and their orientation with respect to the bilayer normal. Peptide dynamics and the effect of polypeptides on the macroscopic phase preference of phospholipid membranes have been investigated in nonoriented samples. Investigations of the structure and topology of membrane channels, peptide antibiotics, signal sequences as well as model systems that allow one to dissect the interaction contributions in phospholipid membranes will be presented in greater detail. PMID- 10516571 TI - NMR characterization of partially folded and unfolded conformational ensembles of proteins. AB - Studies of unfolded and partially folded proteins provide important insight into the initiation and process of protein folding. This review focuses on the use of nmr in characterization of ensembles of proteins that model the early stages of folding. Analysis of an ensemble includes description of the number of conformations, their structure, relative populations, interconversion rates, and dynamics of subconformations. A chemically synthesized analogue of bovine pancreatic trypsin inhibitor (BPTI), [14-38](Abu), has provided a rare system for characterization of multiple partially folded conformations in slow exchange at near physiological conditions. Multidimensional nmr techniques coupled with selective labeling were used to probe different segments of the polypeptide chain. At each labeled site, there is evidence of slow interconversion between two families of partially folded conformations that in themselves are ensembles of rapidly interconverting conformers. All these conformers display significantly more order in the core relative to the rest of the molecule. For other variants of BPTI that are unfolded at equilibrium, the most ordered structure is also favored in the hydrophobic core residues of the native protein. This is consistent with the hypothesis that the residues that are the first to fold go on to form the most stable, structure-determining part of the protein. PMID- 10516572 TI - Structural characterization of peptide hormone/receptor interactions by NMR spectroscopy. AB - The structural characterization of peptide hormones and their interaction with G protein (guanine nucleotide-binding regulatory protein) coupled receptors by high resolution nmr is described. The general approaches utilized can be categorized into three different classes based on their target: the ligand, the receptor, and the ligand/receptor complex. Examples of these different approaches, aimed at facilitating the rational design of peptides and peptidomimetics with improved pharmacological profiles, based on work carried out in our own laboratory, are given. In the ligand-based approach, the high-resolution structures of bradykinin analogues allowing for the development of a structure-activity relationship for activation of the B1 receptor are described. Studies targeting the receptor are to a large extent theoretical, based on computational molecular modeling. However, experimentally based structural features provided by high-resolution nmr can be used to great advantage, providing insight into the mechanism of receptor function, as illustrated here with results from parathyroid hormone. A similar combination of theoretical methods, supplemented by high-resolution structures from nmr has been utilized to probe the formation and stabilization of the ligand/receptor complex both for parathyroid hormone and cholecystokinin. In each of these three approaches, the importance of well-designed peptide mimetics and accurate structural analysis by high-resolution nmr, will be highlighted. PMID- 10516573 TI - NMR techniques for characterization of ligand binding: utility for lead generation and optimization in drug discovery. AB - Over the last ten years, nmr spectroscopy has evolved into an important discipline in drug discovery. Initially, nmr was most useful as a technique to provide structural information regarding protein drug targets and target-ligand interactions. More recently, it has been shown that nmr may be used as an alternative method for identification of small molecule ligands that bind to protein drug targets. High throughput implementation of these experiments to screen small molecule libraries may lead to identification of potent and novel lead compounds. In this review, we will use examples from our own research to illustrate how nmr experiments to characterize ligand binding may be used to both screen for novel compounds during the process of lead generation, as well as provide structural information useful for lead optimization during the latter stages of a discovery program. PMID- 10516574 TI - Multisubstrate biodegradation kinetics of naphthalene, phenanthrene, and pyrene mixtures. AB - Biodegradation kinetics of naphthalene, phenanthrene and pyrene were studied in sole-substrate systems, and in binary and ternary mixtures to examine substrate interactions. The experiments were conducted in aerobic batch aqueous systems inoculated with a mixed culture that had been isolated from soils contaminated with polycyclic aromatic hydrocarbons (PAHs). Monod kinetic parameters and yield coefficients for the individual compounds were estimated from substrate depletion and CO(2) evolution rate data in sole-substrate experiments. In all three binary mixture experiments, biodegradation kinetics were comparable to the sole substrate kinetics. In the ternary mixture, biodegradation of naphthalene was inhibited and the biodegradation rates of phenanthrene and pyrene were enhanced. A multisubstrate form of the Monod kinetic model was found to adequately predict substrate interactions in the binary and ternary mixtures using only the parameters derived from sole-substrate experiments. Numerical simulations of biomass growth kinetics explain the observed range of behaviors in PAH mixtures. In general, the biodegradation rates of the more degradable and abundant compounds are reduced due to competitive inhibition, but enhanced biodegradation of the more recalcitrant PAHs occurs due to simultaneous biomass growth on multiple substrates. In PAH-contaminated environments, substrate interactions may be very large due to additive effects from the large number of compounds present. PMID- 10516575 TI - Influence of fermentation conditions and microfiltration processes on membrane fouling during recovery of glucuronane polysaccharides from fermentation broths. AB - We have investigated the recovery of exopolysaccharides produced by Sinorhizobium meliloti M5N1 CS bacteria from fermentation broths using different membrane filtration processes: cross-flow filtration with a 7 mm i.d. tubular ceramic membrane of 0.5-microm pores under fixed transmembrane pressure or fixed permeate flux and dynamic filtration with a 0.2 microm nylon membrane using a 16-cm rotating disc filter. With the tubular membrane, the polysaccharide mass flux was mainly limited by polymer transmission that decayed to 10% after 90 min. The mass flux of polymer produced under standard fermentation conditions (70 h at 30 degrees C) stabilized after 70 min to 15 g/h/m(2). This mass flux rises to 36 g/h/m(2) when the mean stirring speed during fermentation is increased and to 123 g/h/m(2) when fermentation is extended to 120 h. In both cases, the mean molecular weight of polysaccharides drops from 4.0 10(5) g/mol under standard conditions to 2.7 10(5) g/mol. A similar reduction in molecular weight was observed when the fermentation temperature was raised to 36 degrees C without benefit to the mass flux. These changes in fermentation conditions have little effect on stabilized permeate flux, but raise significantly the sieving coefficient, due probably to molecular weight reduction and the filamentous aspect of the polymer as observed from SEM photographs. The polymer-mass flux was also increased by reducing transmembrane pressure (TMP) and raising the shear rate by inserting a rod in the membrane lumen. Operation under fixed permeate flux instead of constant TMP inhibited fouling during the first 4 h, resulting in higher sieving coefficients and polymer mass fluxes. The most interesting results were obtained with dynamic filtration because it allows operation at high-shear rates and low TMP. Sieving coefficients remained between 90 and 100%. With a smooth disc, the polysaccharide mass flux remained close to 180 g/h/m(2) at 1500 rpm and cell concentrations from 1 to 3 g/L. When radial rods were glued to the disc to increase wall shear stress and turbulence, the mass flux rose to 275 g/h/m(2) at the same speed and cell concentration. PMID- 10516576 TI - Macroscopic growth of filamentous fungi on solid substrate explained by a microscopic approach. AB - A quantitative model predicting biomass growth on solid media has been developed. The model takes into account steric interactions between hyphae and tips at the microscopic level (competition for substrate and tip-hypha collisions). These interactions effect a slowing down of the hyphal, population-averaged extension rate and are responsible, at the microscopic level, for the distribution of tip orientations observed at the colony border. At the macroscopic level, a limiting value of the colony radial extension rate is attained. A mathematical model that combines hyphal branching, tip diffusion, and biomass growth was proposed to explain such behavior. Experiments using Gibberella fujikuroi were performed to validate the model; good agreement between experiments and simulations was achieved. Most parameters can be measured by simple image analysis on the peripheral growth zone, and they have clear physical meaning; that is, they correspond to properties of single, leading hyphae. The model can be used to describe two-dimensional (2D) solid media fermentation experiments under varying culture conditions; the model can also be extended to consider growth in three dimensional (3D), complex geometry substrates. PMID- 10516577 TI - Conditionally adherent growth of serum-independent CHO cells for automated drug screening and biopharmaceutical production. AB - SSF3 is a CHO cell line adapted for growth in protein-free medium. It grows in suspension unless serum-derived attachment factors such as vitronectin are added to the medium. Serum-independent cell lines, which adhere to the substrate after induction with dexamethasone or constitutively, were created by transfection with a human vitronectin gene under control of the mouse mammary tumor-virus promoter. Substrate attachment and SSF3VN-cell spreading could be prevented with an RGD peptide (arginine-glycine-aspartic acid) confirming that attachment is mediated by an intregrin receptor. Hormone-inducible attachment could be blocked by glucocorticoid antagonist promegestone. All steps in the isolation of stable transfected SSF3VN cell lines could be done in a chemically defined medium avoiding the risk of introduction of serum-derived infectious agents. SSF3VN cells could be grown in protein-free medium in solid-phase large-scale bioreactors. Application in microplates as used in high-throughput screening was demonstrated in an assay of Ca(2+) release from internal stores induced by agonist-binding to recombinant human metabotropic glutamate receptor hmGluR1b. PMID- 10516578 TI - Recombinant glycoprotein product quality in proliferation-controlled BHK-21 cells. AB - We analyzed product quality to determine the applicability of proliferation controlled mammalian cells for recombinant pharmaceutical protein production. Baby hamster kidney (BHK)-21 cells were engineered to express a dicistronic, stabilized, self-selecting growth control system consisting of a beta-estradiol activatable transcription factor IRF-1 fusion protein. IRF-1 activity led to a reduced growth rate, whereas productivity, protein integrity, and glycosylation pattern of the industrially relevant secreted pharmaceutical glycoprotein erythropoietin remained consistent, showing that this technique has the potential for improving the consistency of high-quality pharmaceutical products and thus warrants further development. PMID- 10516579 TI - Dielectric analysis for estimation of oil content in the mycelia of Mortierella alpina. AB - The dielectric behavior of the filamentous fungi Mortierella alpina SAM2104 and 1S-4, which produce polyunsaturated fatty acid enriched oil in the mycelia, was investigated. During the cultivation carried out in a 10-kL fermentor for 12-15 days, the relative permittivity and conductivity of the broth were measured in the frequency range of 100 kHz to 30 MHz. The dielectric parameters, i.e., the amplitude of dielectric relaxation (Deltaepsilon) and the characteristic frequency (f(c)), were obtained by fitting the Cole-Cole equation to the observed dielectric relaxation, and the conductivity of the medium (kappa(a)) was also measured. The value of Deltaepsilon gradually increased from the second day through the end of cultivation, suggesting that volume fraction of the cell increased with oil accumulation. The conductivity of the cytoplasm (kappa(i)) was calculated from the experimental values of f(c) and kappa(a), using a theoretical equation based on an ellipsoidal cell model. As a result, good correlation between the calculated kappa(i) and the oil content was obtained. These findings indicate that dielectric analysis enables us to estimate the oil content in the mycelia of oleaginous fungi and also provides a useful tool for monitoring cell growth and for controlling the cultivation process. PMID- 10516580 TI - Tetracycline-regulated overexpression of glycosyltransferases in Chinese hamster ovary cells. AB - The glycosylation patterns of recombinant therapeutic glycoproteins can be engineered by overexpression of glycosyltransferases in the host cells used for glycoprotein production. Most prior glycosylation engineering experiments have involved constitutive expression of cloned glycosyltransferases. Here we use tetracycline-regulated expression of two glycosyltransferases, N acetylglucosaminlytransferases III and V (GnTIII and GnTV) to manipulate glycoform biosynthesis in Chinese hamster ovary (CHO) cells and to study the effect of glycosyltransferase overexpression on this host. The amount of GnTIII and GnTV in these cells, and the glycosylation patterns of several cellular glycoproteins, could be controlled simply by manipulating the concentration of tetracycline in the culture medium. Using this system, it was found that overexpression of either GnTIII or GnTV to high levels led to growth inhibition and was toxic to the cells, indicating that this may be a general feature of glycosyltransferase overexpression. This phenomenon has not been reported previously, probably due to the widespread use of constitutive promoters, and should be taken into account when designing vectors for glycosylation engineering. The growth inhibition effect sets an upper limit to the level of glycosyltransferase overexpression, and may thereby also limit the maximum extent of in vivo modification of poorly accessible glycosylation sites. Also, such inhibition implies a bound on constitutive glycosyltransferase expression which can be cloned. PMID- 10516581 TI - Visualizing integrated bioprocess designs through "windows of operation". AB - This paper demonstrates a simple graphical approach for the design and analysis of a bioprocess flowsheet in which process interactions are significant. Results are presented showing how the feasible space for operation can be simulated and used both to address key design and operating decisions and to identify suitable trade-offs between operating variables, such as fermentation growth rate and disruption conditions, in order to achieve prespecified levels of process performance. Using verified models to describe the production and isolation of an intracellular protein alcohol dehydrogenase (ADH) in yeast as a test bed, a series of so-called "windows of operation" are developed at growth rates in the range of 0.06-0.28 h(-1) and for a range of overall process specifications. The effects of altering the process design performance specification as defined by the level of cell debris removal and the overall process productivity on the size and position of the feasible space were investigated to demonstrate the sensitivity of the flowsheet to changes in process objectives. Using the approach it has been possible to visualise the processing trade-offs required to increase performance in terms of the level of cell debris removal by 50% and the overall process productivity by 400% from a defined base level. The approach provides a convenient tool when designing integrated bioprocesses by enabling process options to be compared visually and can help in achieving better process designs and accelerating process development for the biological process industry. PMID- 10516582 TI - Mass balance modeling of vanillin production from vanillic acid by cultures of the fungus Pycnoporus cinnabarinus in bioreactors. AB - A systematic two-step procedure for the structural identification of bioprocesses is followed in order to establish a mechanistic model for vanillin production by Pycnoporus cinnabarinus. The first step is devoted to the identification of the underlying reaction structure and the development of a validated mass balance model for the growth of P. cinnabarinus and the biotransformation of vanillic acid into vanillin. The second step is devoted to the kinetic modeling, namely, the estimation of the reaction rates and the calibration of the kinetic parameters. The whole procedure leads to the final set up of a simulation model of the process. The results are supported by the data from five cultures of P. cinnabarinus in bioreactors. PMID- 10516583 TI - Preservation of frozen yeast cells by trehalose. AB - Two different methods commonly used to preserve intact yeast cells-freezing and freeze-drying-were compared. Different yeast cells submitted to these treatments were stored for 28 days and cell viability assessed during this period. Intact yeast cells showed to be less tolerant to freeze-drying than to freezing. The rate of survival for both treatments could be enhanced by exogenous trehalose (10%) added during freezing and freeze-drying treatments or by a combination of two procedures: a pre-exposure of cells to 40 degrees C for 60 min and addition of trehalose. A maximum survival level of 71.5 +/- 6.3% after freezing could be achieved at the end of a storage period of 28 days, whereas only 25.0 +/- 1.4% showed the ability to tolerate freeze-drying treatment, if both low-temperature treatments were preceded by a heat exposure and addition of trehalose to yeast cells. Increased survival ability was also obtained when the pre-exposure treatment of yeast cells was performed at 10 degrees C for 3 h and trehalose was added: these treatments enhanced cell survival following freezing from 20.5 +/- 7. 7% to 60.0 +/- 3.5%. Although both mild cold and heat shock treatments could enhance cell tolerance to low temperature, only the heat treatment was able to increase the accumulation of intracellular trehalose whereas, during cold shock exposure, the intracellular amount of trehalose remained unaltered. Intracellular trehalose levels seemed not to be the only factor contributing to cell tolerance against freezing and freeze-drying treatments; however, the protection that this sugar confers to cells can be exerted only if it is to be found on both sides of the plasma membrane. PMID- 10516584 TI - Photoimmobilization of organophosphorus hydrolase within a PEG-based hydrogel. AB - Organophosphorous hydrolase (OPH) was physically and covalently immobilized within photosensitive polyethylene glycol (PEG)-based hydrogels. The hydroxyl ends of branched polyethylene glycol (b-PEG, four arms, MW = 20,000) were modified with cinnamylidene acetate groups to give water-soluble, photosensitive PEG macromers (b-PEG-CA). The b-PEG-CA macromers underwent photocrosslinking reaction and formed gels upon UV irradiation (>300 nm) in the presence of erythrosin B. Native OPH was pegylated with cinnamylidene-terminated PEG chains (MW = 3400) to be covalently linked with the b-PEG-CA macromers during photogelation. The effect of pegylation on the stability of the enzyme was determined. Furthermore, the effect of enzyme concentration, wavelength of irradiation, and photosensitizer on the stability of the entrapped enzyme was also investigated. The pegylated OPH was more stable than the native enzyme, and the OPH-containing gels exhibited superior stability than the soluble enzyme preparations. PMID- 10516585 TI - Extended serial passaging of mammalian neural stem cells in suspension bioreactors. AB - Neural stem cells (NSCs) are primitive cells that are the "parent" cells of all the cells in the central nervous system (CNS). Their discovery in 1992 opened the door to a multitude of potential therapies and treatments to cure neurodegenerative diseases such as Parkinson's disease, multiple sclerosis, and Huntington's disease, which affect millions of people worldwide and cost billions of dollars in health care each year. This study proposes optimal serial passaging protocols so that mammalian neural stem cells can effectively be grown in suspension culture. We examined stationary culture passaging protocols and developed our own optimal procedure. Also examined was the effect of serially cultivating the neural stem cells in suspension culture for an extended period of time. The cells were grown for over 35 days in suspension with an overall multiplication ratio of over 10(7) with no decrease in growth rate, maximum cell density, or viability. The cells also remained karyotypically normal through 25 doublings and retained their ability to be differentiated into all the major cell types of the CNS-neurons, astrocytes, and oligodendrocytes. For the first time, mammalian neural stem cells were grown on a larger scale in suspension culture and maintained their stem cell characteristics. A semicontinuous scheme for large scale production is also presented. PMID- 10516586 TI - Optimized production of recombinant bluetongue core-like particles produced by the baculovirus expression system. AB - The baculovirus-expression vector system (BEVS) has been widely used for the experimental production of many human and animal single- and multi-unit vaccines, heterologous proteins, and viral insecticides. In this work, the production of recombinant bluetongue virus core-like particles (CLPs), using Sf9 cells in shaker-suspension culture with the SF900 II medium (GIBCO, NY), has been studied. This system involved the simultaneous production of two proteins, VP7 and VP3, and was shown to achieve high volumetric productivities. The key parameters of the time of infection (TOI), and the multiplicity of infection (MOI) were studied. The results show that the peak-volumetric yields and cell-specific yields achieved using low MOIs at low-cell densities were the same as those obtained following infections with a high MOI at high-cell densities. This work establishes the feasibility of using low MOIs in the baculovirus system to produce complex multiprotein particles. PMID- 10516587 TI - Diffusion characteristics of calcium alginate gels. AB - The diffusivity of a protein solute (bovine serum albumin) within calcium alginate gels made from sodium alginate of different guluronic acid content was determined. It was found that protein diffusion within alginate gels, prepared to be isotropic in structure, was greatest for gels prepared from sodium alginate of low guluronic acid content as opposed to those prepared from sodium alginate of high guluronic acid content. This finding was explained in terms of the difference in flexibility of the polymer backbone of the two alginates. The greater the polymer backbone flexibility, the greater the solute diffusivity within the gel. PMID- 10516588 TI - Morphology of single olivocerebellar axons labeled with biotinylated dextran amine in the rat. AB - The morphology of olivocerebellar (OC) axons originating from the inferior olive (IO) was investigated in the rat by reconstructing the entire trajectories of single axons that had been labeled with biotinylated dextran amine. Virtually all of the OC axons entered the cerebellum through the inferior cerebellar peduncle (ICP) contralateral to the IO, with a few exceptions. Although most OC projection was contralateral, a few axons projected bilaterally by crossing the midline within the cerebellum. Collaterals of OC axons could be classified into thick branches and thin collaterals. Thick branches of each OC axon (6.1 +/- 3.7/OC axon, mean +/- SD for n = 16 axons) terminated as climbing fibers (CFs) on single Purkinje cells (PCs) in a one-to-one relationship. Besides terminal arborization around PC thick dendrites, CFs had terminals that surrounded a PC soma, fine branchlets that extended transversely in the molecular layer, and thin retrograde collaterals that re-entered the PC and granular layers. Innervation of a single PC by two CFs originating from the same axon was seen, although infrequently. Concerning thin collaterals, about half of the OC axons had one or only a few collaterals terminating in the white matter of the ICP, most had 1 to 6 collaterals terminating in a single cerebellar nucleus, and all had 3 to 16 collaterals that terminated mainly in the granular layer, but occasionally in the cerebellar white matter and the PC layer. Some swellings of thin collaterals touched somata of presumed Golgi cells and PCs. No OC axons terminated solely in the ICP, cerebellar nucleus or granular layer without giving rise to CFs. PMID- 10516589 TI - Neuronal progenitor-like cells expressing polysialylated neural cell adhesion molecule are present on the ventricular surface of the adult rat brain and spinal cord. AB - In the adult rodent brain, it is now well established that neurons are continuously generated from proliferating neuronal progenitor cells located in the subventricular zone of the lateral ventricle (SVZ) and the dentate gyrus of the hippocampus. Recently, it has been shown that neurons can also be generated in vitro from various regions of the adult brain and spinal cord ventricular neuroaxis. As the highly polysialylated neural cell adhesion molecule (PSA-NCAM) has been shown to be specifically expressed by neuronal progenitor cells of the SVZ and the hippocampus, the present study was designed to determine whether cells expressing this molecule could be detected in the vicinity of the ventricular system of the adult rat brain and spinal cord. After double or triple immunostaining for different neuronal and glial markers, confocal microscopy was used to examine the surface of the ventricular neuroaxis in either 40- to 50 microm-thick transverse vibratome sections cut through different brain regions, or in 200- to 300-microm-thick tissue slices including the intact surface of the brain ventricles or of the spinal cord central canal. In untreated rats, PSA NCAM, microtubule associated protein 2 (MAP2) and class III-beta-tubulin were found to be associated with a number of neuron-like cells located on the surface of the third and fourth ventricles and of the spinal cord central canal. The proliferation of the PSA-NCAM-immunoreactive (IR) neuron-like cells detected on the surface of the third and fourth ventricles was not affected by injection of epidermal growth factor (EGF) or basic fibroblast growth factor (bFGF) into these ventricles, but was stimulated by the combined injection of EGF + bFGF. These data indicate that cells exhibiting features of neuronal progenitors are present on the ependymal surface of the adult rat brain and spinal cord ventricular axis. PMID- 10516590 TI - Spinal and brain circuits to motoneurons of the bulbospongiosus muscle: retrograde transneuronal tracing with rabies virus. AB - Retrograde transneuronal tracing with rabies virus from the left bulbospongiosus muscle (BS) was used to identify the neural circuits underlying its peripheral and central activation. Rats were killed at 2, 3, 4, and 5 days post-inoculation (p.i.). Rabies immunolabelling was combined with immunohistochemical detection of choline acetyltransferase and oxytocin. Virus uptake was restricted to ipsilateral BS motoneurons (2 days p.i.). The onset of transfer (3 days p.i.) visualized interneurons in the dorsal grey commissure (DGC), intermediate zone, and sacral parasympathetic nucleus (SPN), mainly in DGC at L5-S1, and revealed synaptic connections between BS and external urethral sphincter motoneurons. At 4 and 5 days p.i., higher-order interneurons were labelled in other spinal areas and segments. Supraspinal labelling initially involved only Barrington's nucleus, nucleus reticularis magnocellularis, and paragigantocellularis lateralis (4 days p.i.). Later, labelling extended to other populations traditionally associated with control of sexual activity and micturition (periaqueductal grey, paraventricular nucleus, medial preoptic area, prefrontal cortex), but also indicated the intervention of somatic descending motor pathways (vestibulospinal and reticulospinal neurons, "hindlimb" regions of sensorimotor cortex and red nucleus) and cerebellar nuclei in multisynaptic innervation of the labelled motoneurons. Dual color immunofluorescence disclosed multisynaptic links between these motoneurons and thoracolumbar medial sympathetic (choline acetyltransferase immunoreactive) neurons. In contrast, preganglionic neurons in SPN and most oxytocinergic neurons in paraventricular hypothalamic nucleus remained unlabelled, suggesting that parasympathetic and somatic outflow to pelvic organs are probably controlled by separate interneuronal populations and that oxytocinergic spinal projections are more likely to influence sacral autonomic rather than somatic outflow. PMID- 10516591 TI - Expression of the GDNF receptors ret and GFRalpha1 in the developing avian enteric nervous system. AB - The formation of the enteric nervous system (ENS) from neural crest-derived cell precursors requires the growth factor glial cell line-derived neurotrophic factor (GDNF) and the receptors Ret and GDNF family receptor alpha 1 (GFRalpha1). We investigated the location(s), the timing, and the extent to which these GDNF receptors appear in the population of crest-derived precursors that form the avian ENS using immunohistochemistry and in situ hybridization. Sections and whole mounts of embryonic chick gastrointestinal tract were costained with antibodies to the receptors and to HNK-1, a marker for crest-derived cells. Neural crest-derived precursors migrate through the primitive esophagus to colonize the gizzard where an extensive cellular network forms. Ret immunoreactivity (ir) was found in a network of cells in the gizzard at embryonic day (E)3.5. As development proceeded, Ret-immunoreactive cells appeared at progressively more caudal positions and were present in the colon at E7.5. Costaining with Ret and HNK-1 was performed to determine the number of Ret immunoreactive cells in the crest-derived population. Ret appeared in some HNK-1 cells in the esophagus and gizzard at embryonic day (E)3.5. During development, the number of crest cells with Ret increased in the ganglia of the gizzard and small intestine. GFRalpha1-ir was also found in HNK-1 cells in the esophagus at E3.5 but did not appear in the gizzard until E4.5. Surprisingly, the colonizing vanguard of crest-derived cells lacked both Ret- and GFRalpha-ir. Between E4.5 and E6.5, the fraction of HNK-1-positive cells expressing GFRalpha1 increased considerably in the foregut. Ret and GFRalpha1 were coexpressed in many cells at E6.5, and the number of such cells increased as development progressed. In the adult, GFRalpha1 and Ret were found in the neuropil of enteric ganglia. We conclude that the population of cells expressing the receptors increases during development and persists in the adult, findings that support a neurotrophic role for GDNF in the formation and maintenance of the avian ENS. PMID- 10516592 TI - Relative number of cells projecting from contralateral and ipsilateral nucleus isthmi to loci in the optic tectum is dependent on visuotopic location: horseradish peroxidase study in the leopard frog. AB - The leopard frog optic tectum is the principal target of the contralateral retina. The retinal terminals form a topographic map of the visual field. The tectum also receives bilateral topographic input from a midbrain structure called nucleus isthmi. In this study we determined the relative strength of n. isthmi projections to different loci in the tectum. Horseradish peroxidase (HRP) was applied at single superficial tectal locations in a series of leopard frogs. The application sites were distributed across the tectum. Retrogradely filled cells were counted in ipsilateral and contralateral nucleus isthmi. Although all regions of the tectum receive input from both n. isthmi, the relative number of labeled cells in the two n. isthmi is dependent on visuotopic location. Input to the rostromedial tectum representing the visual field ipsilateral to the labeled tectum comes primarily from the contralateral n. isthmi. Input to the caudolateral tectum representing the visual field contralateral to the labeled tectum originates mostly from the ipsilateral n. isthmi. Tectal application sites representing the visual midline had approximately equal numbers of labeled cells in the two n. isthmi. The results are similar at postapplication survival times ranging from 2 to 14 days. Using application of HRP to rostral tectum and application of nuclear yellow to caudal tectum, we show that the anisotropy in isthmi labeling is not due to take up of these labels by isthmotectal fibers passing through the application sites that terminate elsewhere. PMID- 10516593 TI - DLX-1, DLX-2, and DLX-5 expression define distinct stages of basal forebrain differentiation. AB - The homeobox genes in the Dlx family are required for differentiation of basal forebrain neurons and craniofacial morphogenesis. Herein, we studied the expression of Dlx-1, Dlx-2, and Dlx-5 RNA and protein in the mouse forebrain from embryonic day 10.5 (E10.5) to E12.5. We provide evidence that Dlx-2 is expressed before Dlx-1, which is expressed before Dlx-5. We also demonstrate that these genes are expressed in the same cells, which may explain why single mutants of the Dlx genes have mild phenotypes. The DLX proteins are localized primarily to the nucleus, although DLX-5 also can be found in the cytoplasm. During development, the fraction of Dlx-positive cells increases in the ventricular zone. Analysis of the distribution of DLX-1 and DLX-2 in M-phase cells suggests that these proteins are distributed symmetrically to daughter cells during mitosis. We propose that DLX-negative cells in the ventricular zone are specified progressively to become DLX-2-expressing cells during neurogenesis; as these cells differentiate, they go on to express DLX-1, DLX-5, and DLX-6. This process appears to be largely the same in all regions of the forebrain that express the Dlx genes. In the basal telencephalon, these DLX-positive cells differentiate into projection neurons of the striatum and pallidum as well as interneurons, some of which migrate to the cerebral cortex and the olfactory bulb. PMID- 10516594 TI - Aging impairs axonal sprouting response of dentate granule cells following target loss and partial deafferentation. AB - Compared to other brain regions, the hippocampus shows considerable susceptibility to the aging process. Aging may impair the compensatory plastic response of hippocampal neurons following lesions, target loss, and/or deafferentation. We hypothesize that sprouting of dentate granule cell axons (mossy fibers) in response to target loss and partial deafferentation diminishes with age. We quantified mossy fiber sprouting into the dentate supragranular layer (DSGL) following intracerebroventricular kainic acid administration in young adult, middle-aged, and aged rats, using Timm's histochemical method. Mossy fiber ingrowth into the DSGL was assessed in the septal hippocampus at 2- and 4 months postlesion by measuring both the average width and the relative density of sprouted terminals. Kainic acid lesions produced degeneration of CA3 pyramids with sparing of CA1 and dentate granule cells in all age groups. Although young adults demonstrated robust DSGL mossy fiber sprouting, sprouting was significantly reduced in both middle-aged and aged rats. Compared to the case in young adults, the overall sprouting in middle-aged animals was reduced by 52% at 2 months and 50% at 4 months postlesion, whereas in aged rats the sprouting was reduced by 53% at 2 months and 64% at 4 months postlesion. Aged animals also showed an overall reduction of 28% compared to middle-aged animals at 4 months postlesion. Dramatically reduced sprouting in aged animals may represent a deficit in recognition of target loss and partial deafferentation by aged granule cells and/or an impaired up-regulation of factors that stimulate neurite outgrowth in the aged brain. PMID- 10516596 TI - Dendritic morphology of cat retinal ganglion cells projecting to suprachiasmatic nucleus. AB - The morphological properties of cat retinal ganglion cells projecting to the suprachiasmatic nucleus (SCN) of the hypothalamus were studied by using retrograde labeling, in vitro intracellular injection, confocal optical section, and computer three-dimensional reconstruction techniques. A total of 218 stained cells were studied. Neither the dendritic fields nor soma diameters of SCN projecting cells varied with eccentricity. Approximately 50% of cells were concentrated not in the area centralis, but rather in the visual streak. SCN projecting cells showed large and symmetrical dendritic fields (596 +/- 159 microm) and medium to small sized somas (17.2 +/- 3.3 microm). The ramification patterns of SCN-projecting cells were similar. Most cells primarily ramify in either sublamina A or B. Evidence from quantitatively analyzed cells (n = 39) suggests that these cells ramified more frequently in sublamina A (n = 17) than in sublamina B (n = 8). A large number of cells, on the other hand, showed diffuse ramification (n = 14) throughout the inner plexiform layer (IPL). The functional roles of these cells and the corresponding retinal neurocircuitry in circadian entrainment remain to be studied. PMID- 10516595 TI - Decussations of the descending paraventricular pathways to the brainstem and spinal cord autonomic centers. AB - Decussations of descending fibers of the hypothalamic paraventricular nucleus (PVN) were investigated by using Phaseolus vulgaris-leucoagglutinin (PHA-L) in intact and brainstem-operated rats. Fibers descend ipsilaterally along the brainstem and spinal cord and decussate at four levels: 1) Supramamillary decussations (SM). PVN fibers reach this area through the lateral hypothalamus and along the third ventricle in the dorsal hypothalamus. In the posterior hypothalamus some fibers crossover in the SM and terminate in the supramamillary region bilaterally. 2) Pontine tegmentum. PVN fibers run in the lateral part of the tegmentum arching to the basis of the pons. Some fibers crossover under the fourth ventricle. The locus ceruleus and the Barrington's nucleus receive bilateral innervation with ipsilateral dominance. 3) Commissural part of the nucleus of the solitary tract (NTS). The major crossover of PVN fibers is found here. The decussated fibers form a dense network here, and loop rostralward to innervate the entire NTS. A midsagittal knife-cut through the NTS eliminated paraventricular-fibers on the contralateral side. Synaptic contacts between PHA-L labeled boutons and tyrozine hydroxilase-positive neurons were verified in the NTS. The caudal ventrolateral medulla also receives bilateral innervation. 4) Lamina X of the thoracic spinal cord. Paraventricular fibers enter the lateral funiculus ipsilaterally and innervate the intermediolateral cell column (IML). Some fibers cross the midline ventral and dorsal to the central canal running to the contralateral IML, at the level of the decussation. Our results demonstrated that paraventricular projections form a continuous descending pathway on their side of origin, and provide crossover fibers which may terminate segmentally without forming long tracts after crossover. PMID- 10516597 TI - Distribution of FMRFamide-like immunoreactivity in the amphibian brain: comparative analysis. AB - FMRFamide is a small neuropeptide present in particular neurons of the basal forebrain and midbrain of the vertebrate groups studied, especially fishes and mammals. In order to assess interspecies variation, the distribution of FMRFamide like immunoreactivity was studied in the brains of 13 species of amphibian. Although FMRFamide-immunoreactive (IR) terminals occurred throughout much of the brain, IR cell groups were noted in circumscribed regions of the CNS. In the eight anuran species studied, two major populations of labeled perikarya were observed: one in the septopreoptic area and another one in the caudal portion of the diencephalon. The rostrocaudal extent of both and the number of labeled somata in each neuronal group displayed species-specific differences. In urodeles and gymnophiones, labeled perikarya were located in the diencephalon, but there were remarkable species differences in the number of such cells. It is discussed whether sex or season of collection may account for some of the differences observed. The distribution of FMRFamide-IR perikarya, fibers, and pathways in the brain of anurans, urodeles, and gymnophiones was compared. The existence of FMRFamide perikarya in the anterior preoptic neuropil and medial septum appeared to be a feature common to all anurans; labeled neurons in the dorsal thalamus, however, may be present only in the (viviparous) gymnophione Typhlonectes compressicauda. Cerebrospinal fluid contacting FMRFamide neuronal cell bodies and fibers were observed in each of the three taxonomic orders. The data are compared with those previously obtained for other groups of vertebrates. PMID- 10516598 TI - A molecular basis of cell death in olfactory epithelium. AB - When the membrane receptor Fas binds its ligand, Fas ligand (FasL), an apoptotic cascade is initiated in the cell bearing the Fas receptor. The same can be said about the tumor necrosis factor receptor-1 (TNFR1) and its ligand, TNF-alpha. In this study we have shown that the mRNAs of both sets of ligands and receptors, Fas/FasL and TNF-alpha/TNFR1, were present in unperturbed olfactory epithelium. Fas and FasL were shown by immunohistochemistry and by Western blots of bulbectomized animals to be in the neurons and in some non-neuronal (microvillar) cells of unperturbed rat olfactory epithelium. Addition of either FasL or TNF alpha to organotypic cultures of fetal rat olfactory epithelium resulted in a significant increase in the number of apoptotic bodies after 4-6 hours. These data raise the possibility that either or both ligand-receptor pairs participate in cell death in the olfactory epithelium. PMID- 10516599 TI - Structural characterization of marginal (lamina I) spinal cord neurons in the cat: a Golgi study. AB - The neuronal population of the spinal cord lamina I (marginal zone) was structurally characterized, in the cat, by the use of the Golgi method complemented by multivariate analysis of morphometric data. Four cell types were identified, two of them including two subtypes. Fusiform cells accounted for 43% of impregnated cells and presented flame-shaped rostrocaudally elongated perikarya and bipolar, either strictly longitudinal (fusiform A; 37%) or longitudinal and ventral (fusiform B; 6%) dendritic arbors with numerous short pedicled spines. Fusiform cells preferentially occupied the lateral one-third of lamina I. Multipolar cells (22%) had ovoid perikarya with bulging surfaces and numerous primary dendritic trunks. Two subtypes could be distinguished: multipolar A cells (12%) with highly ramified dendrites covered with variably shaped spines and multipolar B cells (10%) with looser and less spiny dendritic arbors expanded for longer distances. Multipolar cells were more commonly found in the medial half of lamina I. Flattened cells (16%) possessed discoid perikarya flattened across the dorsoventral axis and aspiny, scarcely ramified dendritic arbors distributed horizontally within lamina I. They predominated in the intermediate one-third of the lamina. Pyramidal cells had triangular prismatic perikarya partially encased in the white matter overlying lamina I. They represented 19% of the impregnated neurons and were located along the entire lateromedial extent of the lamina. Each neuronal type included a few cells with perikarya and dendritic arbors three times larger than the rest. These so-called giant cells amounted to 6% of the entire lamina I neuronal population. According to the present data, the neuronal population of the spinal cord lamina I of the cat strongly resembles that of the rat (Lima and Coimbra, J. Comp. Neurol. 244:53 71, 1986), which strengthens the functional relevance of this structural classification. PMID- 10516600 TI - Dopaminergic cell group A8 in the monkey: anatomical organization and projections to the striatum. AB - The first part of the study was a quantitative analysis of the distribution of A8 neurons compared with that of A9 and A10 neurons by means of tyrosine hydroxylase and calbindin-D(28K) immunohistochemistry and image analysis in monkeys. Then the striatal projection of A8 neurons was studied using retrograde and anterograde tracing methods. It was compared with that originating in cell groups A9 and A10 by performing injections of the retrograde tracer wheat germ agglutinin conjugated to horseradish peroxidase into different regions of the striatum. Ten percent of all mesencephalic dopaminergic neurons are located in cell group A8. This cell group, along with A10 and the dorsal part of A9, constitutes the dorsal tier, which accounts for 28% of mesencephalic dopaminergic neurons. Double staining experiments showed that the neurons located in the dorsal tier were calbindin positive, whereas those from the ventral tier were not. In terms of anatomical projection, the dorsal tier mainly projects to the ventral part of the associative striatum, with preferential projections of A8 neurons to the ventrocaudal putamen, of A10 neurons to the nucleus accumbens, and of dorsal A9 neurons to both. Conversely, the main targets of the ventral tier of mesencephalic neurons (ventral part of A9) are the sensorimotor putamen and the associative caudate nucleus. In conclusion, each mesencephalic cell group projects primarily to one specific striatal region but also participates, albeit to a lesser extent, in the innervation of all the remaining striatal parts. PMID- 10516601 TI - Medium-sized neurofilament protein related to maturation of a subset of cortical neurons. AB - Neurofilaments are typical structures of the neuronal cytoskeleton and participate in the formation and stabilization of the axonal and dendritic architecture. In this study, we have characterized a murine monoclonal antibody, FNP7, that is directed against the medium-sized neurofilament subunit NF-M. This antibody identifies a subset of neurons in the cerebral cortex of various species including human and in organotypic cultures of rat cortex. In the neocortex of all species examined, the antibody labels pyramidal cells in layers III, V, and VI, with a distinctive laminar distribution between architectonic boundaries. In comparison with other antibodies directed against NF-M, the FNP7 antibody identifies on blots two forms of NF-M that appear relatively late during development, at the time when dynamic growth of processes changes to the stabilization of the formed processes. Dephosphorylation with alkaline phosphatase unmasks the site, making it detectable for the FNP7 antibody. The late appearance suggests that the site is present during early development in phosphorylated form and with increasing maturation becomes dephosphorylated, mainly in dendrites. This event may relate to changes in cytoskeleton stability in a late phase of dendritic maturation. Furthermore, mainly corticofugal projections and only few callosal axons are stained, suggesting a differential phosphorylation in a subset of axons. The antibody provides a useful marker to study subsets of pyramidal cells in vivo, in vitro, and under experimental conditions. PMID- 10516602 TI - Sympathetic-related neurons in the preoptic region of the rat identified by viral transneuronal labeling. AB - The viral transneuronal labeling method was used to localize sympathetic-related neurons in the preoptic region following pseudorabies virus (PRV) injections into either the superior cervical ganglion, stellate ganglion, celiac ganglion, or adrenal gland of rats. A general pattern of infection was detected. First, neuronal labeling was found in the medial preoptic area, medial preoptic nucleus, median preoptic nucleus, and lateral preoptic area, and then it spread to the anteroventral periventricular, anteroventral preoptic, and parastrial nuclei. Finally, the forebrain circumventricular organs: organum vasculosum of the lamina terminalis (OVLT) and subfornical organ (SFO) became infected. Neuropeptide containing preoptic neurons were analyzed following PRV injections in the stellate ganglion. Some thyrotropin-releasing hormone and neurotensin neurons were labeled, but none of the calcitonin gene-related peptide, cholecystokinin, corticotropin-releasing factor, galanin, luteinizing hormone-releasing hormone, enkephalin, substance P, or tyrosine hydroxylase neurons were PRV infected. Two major sympathetic networks appear to be represented in the preoptic region. One is linked to the OVLT, SFO, and anteroventral third ventricular (AV3V) region, sites previously implicated in fluid and electrolyte balance as well as cardiovascular control. The other descending sympathetic pathway appears to target the medial preoptic nucleus as its key nodal point, receiving inputs from infralimbic cortex and limbic regions, such as the lateral septum, medial nucleus of the amygdala, subiculum, and amygdalohippocampal area, and then, projecting caudally to the hypothalamus and brainstem. This second sympathetic network may subserve affiliative, defensive and sexual behaviors. PMID- 10516603 TI - Neurogenesis in the median domain of the embryonic brain of the grasshopper Schistocerca gregaria. AB - Embryonic development in the median domain of the brain of the grasshopper Schistocerca gregaria was investigated with immunohistochemical, histological, and intracellular dye injection techniques. The early head midline is divisible into a dorsal median domain and a ventral median domain based on the orientation of cell somata in each region. At 25% of embryogenesis, a single large midline precursor differentiates in the dorsal median domain and produces a lineage of six neuronal progeny before degenerating. No further precursors arise. In addition, the primary commissure pioneers and a pair of lateral neurons differentiate directly from the ectoderm in this region. Lucifer yellow dye injected into the midline precursor stains only this cell and its progeny. Similarly, there is no dye coupling from the primary commissure pioneers to the midline lineage or to neuroblasts of the brain hemispheres. Neurogenesis in the dorsal median domain therefore proceeds separately within each subset of cells, and is not related to development in the brain hemispheres. Beginning at 42% of embryogenesis, the primary commissure pioneers undergo a morphological transformation and concomittantly express the Term-1 antigen. Expression continues throughout embryogenesis and into the adult, where the midline primary commissure pioneer cells are the only ones labeled by Term-1 in the entire brain. The cellular organization of the dorsal median domain therefore remains remarkably conserved throughout embryogenesis, even as the brain undergoes extensive morphological transformation. PMID- 10516604 TI - Prosomeric map of the lamprey forebrain based on calretinin immunocytochemistry, Nissl stain, and ancillary markers. AB - The structural organization of the lamprey extratelencephalic forebrain is re examined from the perspective of the prosomeric segmental paradigm. The question asked was whether the prosomeric forebrain model used for gnathostomes is of material advantage for interpreting subdivisions in the lamprey forebrain. To this aim, the main longitudinal and transverse landmarks recognized by the prosomeric model in other vertebrates were identified in Nissl-stained lamprey material. Lines of cytoarchitectural discontinuity and contours of migrated neuronal groups were mapped in a two-dimensional sagittal representation and were also classified according to their radial position. Immunocytochemical mapping of calretinin expression in adjacent sections served to define particular structural units better, in particular, the dorsal thalamus. These data were complemented by numerous other chemoarchitectonic observations obtained with ancillary markers, which identified additional specific formations, subdivisions, or boundaries. Emphasis was placed on studying whether such chemically defined neuronal groups showed boundaries aligned with the postulated inter- or intraprosomeric boundaries. The course of diverse axonal tracts was studied also with regard to their prosomeric topography. This analysis showed that the full prosomeric model applies straightforwardly to the lamprey forebrain. This finding implies that a common segmental and longitudinal organization of the neural tube may be primitive for all vertebrates. Interesting novel aspects appear in the interpretation of the lamprey pretectum, the dorsal and ventral thalami, and the hypothalamus. The topologic continuity of the prosomeric forebrain regions with evaginated or non-evaginated portions of the telencephalon was also examined. PMID- 10516605 TI - Biosafety considerations for flow cytometric analysis of human immunodeficiency virus-infected samples. PMID- 10516606 TI - Bivariate analysis of the p53 pathway to evaluate Ad-p53 gene therapy efficacy. AB - BACKGROUND: Gene therapy of human tumors with adenovirus vectors presents a clinical research challenge and a potential opportunity in cancer therapy. One of the research challenges is that endpoints like tumor reduction, time to recurrence, and survival do not provide information about whether a potential therapeutic infects the targeted cells or whether the transferred gene functions or induces a cellular response. Therefore, a flow cytometric approach was developed for a wildtype, p53 encoding adenoviral vector (Ad-p53) that provides (1) the relative level of p53 transferred by p53 immunoreactivity, (2) mdm2 immunoreactivity as an assay of p53 activity, and (3) estimates of the percentage of infected cells by dual parameter analysis (p53 versus mdm2). METHODS: Three prostate cancer cell lines (PC-3, LNCaP, DU 145) that are null, wild-type, and mutant for p53, respectively, and two ovarian cancer cell lines (PA1, MDAH 2774) that are wild-type and mutant for p53, respectively, were tested for immunoreactivity and lack of cross-reactivity with the monoclonal antibodies, DO 7 (anti-p53) and IF2 (anti-mdm2). Optimal dual staining conditions for a flow cytometric assay employing saturating levels of antibody were developed and tested by infection of PC-3, PA1, and MDAH 2774 with Ad-p53 or a control virus, Ad-luc. Dual staining with DO-7 and propidium iodide was used to determine any biological effect of the transferred gene. RESULTS: Neither DO-7 nor IF2 showed appreciable cross-reactions by Western blot analysis of representative prostate or ovarian cell lines. By flow cytometric titration, DO-7 appears to be a high avidity antibody (saturation staining of 10(6) DU 145 cells with 0.5ug) whereas IF2 appears less so (optimum signal to noise ratio at 1ug/10(6) cells). Infection with Ad-p53 was detected at 6 to 48 hours post infection as a uniform relative increase in p53 levels over background p53 levels. Coincident increases in mdm2 immunoreactivity were also detected. DNA content measurements of PA1 and MDAH 2774 cells indicated that G1 arrest and/or apoptosis occurred subsequent to Ad p53 infection. p53 and mdm2 levels and DNA content distributions for Ad-luc infected cells were equivalent to uninfected cells. CONCLUSIONS: A flow cytometric approach to measure the efficacy of an Ad-p53 gene therapy vector was developed that detects not only the gene transferred but also the activity of the transferred gene product. PMID- 10516607 TI - Nuclear morphometry as an intermediate endpoint biomarker in chemoprevention of cervical carcinoma using alpha-difluoromethylornithine. AB - The use of nuclear morphometry as an intermediate endpoint biomarker is described in a Phase I, dose-seeking trial of chemoprevention of cervical cancer, using the agent alpha-difluoromethylornithine (DFMO). Thirty patients with grade III cervical intraepithelial neoplasia (CIN III) were enrolled, and these received daily doses of DFMO at 0.06-1.0 mg/m(2) for a period of 1 month. Fifteen patients were observed to have a complete or partial regressive response to the agent, as assessed by histopathology. No significant differences in cell feature measurements were found between responders and nonresponders in specimens obtained before treatment, indicating that it may be difficult to predict response on the basis of these measurements. In specimens collected after treatment, large differences in morphometric features were observed between responders and nonresponders, indicating a differential effect of DFMO. Significantly modulated features were considered in terms of their correlations with CIN grade, which was determined from an independent set of measurements from archival tissue. Differences between features were consistent with a deletion of cells with high grade nuclei in the responders, and with the persistence of a more heterogeneous population of high grade cells in the nonresponders. Based on an independent set of measurements from archival material, a morphometric index of progression was derived, yielding a quantitative measure of the degree of nuclear atypia in these lesions. When applied to this trial, the morphometric index was seen to be specifically and consistently decreased in responsive lesions, and unchanged in nonresponders. The study indicates that morphometric features fulfill the requirements for an intermediate endpoint biomarker of cervical cancer chemoprevention. PMID- 10516608 TI - Modulation of apoptosis by cytokines in B-cell chronic lymphocytic leukemia. AB - B-cell chronic lymphocytic leukemia (B-CLL) is characterized by the slow and progressive accumulation of monoclonal apparently mature, CD5(+) B lymphocytes. The majority of circulating cells appear to be nondividing, and it has been suggested that a prolonged life span is mainly responsible for the accumulation of the leukemic cells. However, spontaneous programmed cell death by apoptosis occurs when B chronic lymphocytic leukemia cells are cultured in vitro. This may be because of the lack of an unidentified essential cytokine present in vivo. Thus, we investigate interleukin-2 (IL-2), IL-4, IL-6 and IL-10 in vitro effects on apoptosis of B cells from 32 previously untreated patients with B-CLL in initial clinical stages. B cells were isolated from peripheral blood, and apoptosis was measured in these cells immediately after isolation and following incubation in vitro, without and with the different cytokines, for 24 and 48 h. Distribution of cellular DNA content and quantitative analysis of apoptosis were determined by standard propidium iodide staining and flow cytometry. Spontaneous apoptosis occurred in B-CLL cells incubated in vitro in the absence of cytokines. Our results indicate that both IL-2 and IL-4, but not IL-6, inhibit in vitro apoptosis in a large percentage of B-CLL patients. IL-10 increases in vitro apoptotic cell number in stage 0 patients, but not in stage I and II. These data support the hypothesis that IL-2 or IL-4, may be cell survival factors in vivo and that IL-10 might be a candidate for immune therapy of early B-CLL. PMID- 10516609 TI - Enumeration of CD4(+) T-cells in the peripheral blood of HIV-infected patients: an interlaboratory study of the FACSCount system. AB - The aim of the present study was to assess the interlaboratory reproducibility of the FACSCount system for the enumeration of peripheral blood (PB) CD4(+) T-cells. In each of the seven participating centers, both previously stained and unstained PB samples (n = 49) were received and either analyzed or stained and then analyzed. Interlaboratory reproducibility was checked in two different groups of centers (n = 3 and n = 4) where the study was performed in parallel. In addition, both the intralaboratory precision and accuracy of this system were analyzed in comparison with results obtained with conventional flow cytometry. Accordingly, upon comparing both methods, a high degree of correlation was observed in the total number of CD3(+) T-cells (coefficient of correlation of 0.9750 +/- 0.0184, slope of the best linear fit: 0. 9214 +/- 0.0311, y-intercept of 12 +/- 47) as well as in the number of CD3(+)/CD4(+) (coefficient of correlation of 0.9794 +/- 0.1457, slope of the best linear fit: 0.9463 +/- 0.0753, y-intercept of -11 +/- 36) and CD3(+)/CD8(+) (coefficient of correlation of 0.9728 +/- 0.0192, slope of the best linear fit: 0.9682 +/- 0.0735, y-intercept of 7 +/- 95) major subsets. In addition, low coefficients of variation (CV) were obtained for replicates, indicating the method's high degree of accuracy. The present study shows that with respect to the interlaboratory reproducibility reported for most techniques used for the enumeration of PB CD4(+) T-cells, the FACSCount system results in data with much lower coefficients of variance (CVs) (mean CV of less than 10%). Upon measuring the impact on results of different variables associated with either sample preparation or data acquisition and analysis, our study clearly shows that data acquisition and analysis does not influence the results by increasing variability since the coefficients of variation obtained for samples prepared in the same laboratory under the same conditions and read in different laboratories with different instruments were identical to those obtained for the replicates of the same samples read in each individual center. In contrast, interlaboratory variability, although low, significantly increased when sample preparation was carried out in different laboratories, suggesting that pipetting still represents the major source of variability in the FACSCount system. PMID- 10516610 TI - Expression of CD134 (0X-40) on T cells during the first 100 days following allogeneic bone marrow transplantation as a marker for lymphocyte activation and therapy-resistant graft-versus-host disease. AB - CD134 (OX-40) is an activation-associated antigen which functions as a costimulatory receptor for CD4+ T cells. In order to determine the expression of CD134 during immune recovery following allogeneic bone marrow transplantation (BMT), we measured its expression on T cells and T cell subsets during the first 100 days following BMT in 26 patients. CD4+CD134+ T could be seen approximately 14 days following BMT cells in patients who did not develop GvHD which required therapy (n = 20). The percentage of CD4+CD134+ cells continued to increase up to the fourth week following BMT to a maximum of 40-50% of CD4+ T cells (normal = 1 8%). Two patients who developed Grade I-II GvHD and who responded to treatment with pulsed high-dose methylprednisolone (MPD) showed a decline of approximately 40% in CD4+CD134+ T cells was seen within 48 hours of treatment. Four patients who developed GvHD that was not responsive to MPD and who later developed high IV GvHD showed increasing CD4+CD134+ T cells up to 85% of the CD4+ T cells. Additionally, rapid increases in CD134+ T cells following antibody-based T cell therapy were associated with GvHD recurrence. In no cases was the percentage of CD134+ CD4+ T cells predictive of clinical GvHD. In this exploratory study, we have shown that CD134, although not predictive of the initial onset of GvHD, may be a useful tool for monitoring the response to early GvHD therapy and identification of patients at risk for reemergence of GvHD who may benefit from anti-T cell therapy. Cytometry (Comm. Clin. Cytometry) 38: 238-243, 1999. PMID- 10516611 TI - Mitogen-induced T-cell CD69 expression is a less sensitive measure of T-cell function than [(3)H]-thymidine uptake. AB - The most widely used in vitro measure of T-cell function has been the assessment of mitogen induced proliferation by [(3)H]-thymidine incorporation. Mitogens also induce T-cell surface expression of a number of molecules associated with activation, including CD69. Recent reports have suggested that flow cytometric analysis of CD69 expression may be a simpler and faster means of measuring T-cell function. Most studies have been on normal subjects, and the sensitivity of CD69 expression as an in vitro measure of clinical immunodeficiency remains unknown. We address this issue by concurrently measuring mitogen-stimulated T-cell CD69 expression and [(3)H]-thymidine incorporation in a normal population and five immunocompromised patients negative for the human immunodeficiency virus (HIV). All patients had recurrent infections and had known causes of immunodeficiency. Whole blood cultures were setup to measure phytohaemagglutinin A (PHA)- and superantigen staphylococcal enterotoxin B (SEB)-induced CD69 expression at 5, 24, and 72 h, and [(3)H]-thymidine incorporation at 72 h. All immunodeficient patients had lower than normal PHA responses and 3 of 4 had low SEB responses. However in 7 out of 8 of the patient tests, mitogen-induced T-cell CD69 expression was within the normal range. Similar results were found with CD4(+) T cell CD69 expression. This study indicates that measurement of mitogen-induced T cell CD69 expression lacks sensitivity in determining T-cell dysfunction in HIV negative immunodeficient patients. PMID- 10516612 TI - Use of mean platelet component to measure platelet activation on the ADVIA 120 haematology system. AB - Platelet activation results in changes in a number of cell surface molecules including an increase in P-Selectin (CD62P) that may be rapidly and conveniently measured by immunofluorescent flow cytometry. The ADVIA 120 (Bayer) is a new system that facilitates more accurate measurement of platelet volume and in addition provides an approximate measure of the mean refractive index (RI) of the platelets reported as mean platelet component (MPC) concentration. We were interested to determine whether changes in MPC might reflect changes in platelet activation status. To investigate this, the platelet CD62P expression, determined by flow cytometry, and change in MPC, measured on the ADVIA 120 system, was first examined in vitro after stimulation of EDTA anticoagulated whole blood with submaximal concentrations of bovine thrombin in the presence or absence of the thromboxane synthase inhibitor, Ridogrel. Thrombin produced a dose-dependent increase in platelet CD62P expression and a decrease in MPC that could be inhibited by Ridogrel at physiological concentrations. In the second set of experiments, blood from 20 normal controls was collected into both EDTA and sodium citrate (SC) anticoagulants. Within 30 min of venesection and again at 3 h post-venesection after storage at room temperature, the platelet MPC and CD62P expression were determined. Platelets in all samples with both anticoagulants showed very low levels of CD62P expression when first analysed. At 3 h there was a small increase in CD62P expression on platelets in whole blood anticoagulated with SC, but a significant (P < 0.001) increase was observed on platelets anti coagulated with EDTA. A negative correlation was found between the change in MPC of the platelets and the increase in the mean fluorescence intensity (MFI) (r = 0.69, P < 0.001, n = 20) and the percentage (r = -0.72, P < 0.001, n = 20) of CD62P positive platelets at 3 h in blood anticoagulated with EDTA. We conclude that a reduction in MPC as measured by the ADVIA 120 may be used to detect anticoagulant induced, as well as thrombin stimulated, in vitro platelet activation in blood anticoagulated with EDTA. Further, we conclude that platelet activation is negligible for up to 3 h in sodium citrate anticoagulated whole blood. PMID- 10516613 TI - Forum: journal club PMID- 10516614 TI - Effects of ischemia on skeletal muscle energy metabolism in mice lacking creatine kinase monitored by in vivo 31P nuclear magnetic resonance spectroscopy. AB - The aim of this study was to provide in vivo experimental evidence for the proposed biological significance of the creatine kinase (CK)/phosphocreatine (PCr) system in the energy metabolism of skeletal muscle. As a test system we compared hindlimb muscle of knockout mice lacking the cytosolic M-type (M-CK(-)/( )), the mitochondrial ScMit-type (ScCKmit(-)/(-)), or both creatine kinase isoenzymes (CK(-)/(-)), and in vivo 31P-NMR was used to monitor metabolic responses during and after an ischemic period. Although single mutants show some subtle specific abnormalities, in general their metabolic responses appear similar to wild type, in contrast to CK(-)/(-) double mutants. This implies that presence of one CK isoform is both necessary and sufficient for the system to be functional in meeting ischemic stress conditions. The global ATP buffering role of the CK/PCr system became apparent in a 30% decline of ATP in the CK(-)/(-) mice during ischemia. Both M-CK(-)/(-) and CK(-)/(-) showed increased phosphomonoester levels during ischemia, most likely reflecting adaptation to a more efficient utilization of glycogenolysis. While in M-CK(-)/(-) muscle PCr can still be hydrolyzed to provide Pi for this process, in CK(-)/(-) muscle only Pi from ATP breakdown is available and Pi levels increase much more slowly. The experiments also revealed that the system plays a role in maintaining pH levels; the CK(-)/(-) mice showed a faster and more pronounced acidification (pH = 6.6) than muscles of wild type and single knockout mutants (pH = 6.9). PMID- 10516615 TI - Structural information in neuronal tissue as revealed by q-space diffusion NMR spectroscopy of metabolites in bovine optic nerve. AB - 1H NMR diffusion experiments performed on the signal of the metabolites in bovine optic nerve showed that the signal decay due to diffusion is bi-exponential with a slow and a fast diffusing component. Diffusion was measured as a function of the diffusion time, and the data were analyzed as a function of b and q values. Bi-exponential fit was used to analyze the data, and the results were compared with the displacement distribution profiles obtained from the q-space analysis of the data. This q-space analysis showed that the fast diffusing component has a broad displacement distribution and appears not to be restricted. On the other hand, the slow diffusing component appears to be highly restricted to milieu in the order of 1-2 microm. The orientation of the sample with respect to the axis for which diffusion was measured affected mainly the relative sizes of the populations of each component, but had only a small effect on the extracted apparent diffusion coefficients. These results from both the b and the q value analyses suggest that the slow diffusing component is related to restricted diffusion of these metabolites in the axonal fibers, while the fast diffusing component represents diffusion of metabolites in cells and along the long axis of the nerve fibers. It is concluded that q-space analysis of metabolite diffusion enables extraction of structural information about the sample, and that the diffusion of the metabolites in optic nerve is dictated mainly by the cellular medium and microstructure of the tissue. PMID- 10516616 TI - 1H-NMR analysis of microbial-derived organic acids in primary root carious lesions and saliva. AB - In addition to lowered pH values, the molecular profile and concentrations of microbial-derived organic acids in carious dentin are important demineralization parameters involved in the induction, development and progression of dental caries. High-resolution proton ((1)H) NMR spectroscopy was employed to examine the organic acid status of primary root carious lesions. (1)H-NMR analysis of post-neutralized perchloric acid extracts of active carious lesions revealed that at an operating frequency of 600 MHz, the (1)H-NMR-detectable organic acid composition of carious dentin samples (mean molecular percentage content +/- standard error; the mean molecular percentage content is defined here as the mean of the concentration of each (1)H-NMR-visible organic acid/anion expressed as a percentage of total (1)H-NMR-detectable organic acid/anion level in each sample) was acetate 51 +/- 2%, formate 37 +/- 2%, lactate 5 +/- 1%, propionate 3 +/- 0.8%, pyruvate 2.4 +/- 0.3%, n-butyrate 1.2 +/- 0.2%; succinate 0.1 +/- 0.1%; iso butyrate, n- and iso-valerate, and n- and iso-caproate (total) <0.2%. Further components detectable included alanine, glycine, choline, phosphorylcholine, trimethylamine oxide, methanol, glycolate and assorted saccharides. In view of their high dissociation constants (K(a)), our results demonstrate that formic and pyruvic acids (K(a) = 1.77 x 10(-4) and 3.20 x 10(-3) mol/dm(3), respectively) contribute substantially to the decreased pH values associated with active caries lesions (cf. lactate K(a) = 1.40 x 10(-4) mol/dm(3)), and hence the pathogenesis of primary root caries. PMID- 10516618 TI - Myocardial manganese elevation and proton relaxivity enhancement with manganese dipyridoxyl diphosphate. Ex vivo assessments in normally perfused and ischemic guinea pig hearts. AB - Manganese (Mn) dipyridoxyl diphosphate (MnDPDP) is the active component of a contrast medium for liver MRI. By being metabolized, MnDPDP releases Mn(2+), which is taken up and retained in hepatocytes. The study examined whether MnDPDP elevates Mn content and enhances proton relaxivity in normal myocardium, but not in ischemic myocardium with reduced coronary flow and impaired metabolism. Isolated guinea pig hearts were perfused at normal flow or low flow, inducing global subtotal ischemia. Ventricular ATP and Mn contents, T(1) and T(2) were measured. At normal flow tissue Mn content increased from the control level of 4.1 to 70.4 micromol/100g dry wt with MnDPDP (3000 microM), while low-flow perfusion with MnDPDP (3000 microM) resulted in a Mn content of 16.6 micromol/100 g dry wt. Prolonged ischemia (35 and 90 min) reduced tissue Mn down to the control level. T(1) shortening closely paralleled myocardial Mn elevations during both normal and low-flow perfusion. The use of a Mn(2+)-releasing contrast agent like MnDPDP may be a promising principle in MRI assessments of myocardial function and viability in coronary heart disease by revealing a differential pattern of changes in T(1) relative to coronary flow, cell Mn uptake and retention, ion channel function and metabolism. PMID- 10516617 TI - Brain metabolic alterations in Cushing's syndrome as monitored by proton magnetic resonance spectroscopy. AB - Proton magnetic resonance spectroscopy ((1)H MRS) was used to evaluate changes in cerebral metabolites in 13 patients with Cushing's syndrome (including seven with pituitary corticotroph adenomas and six with primary adrenal disease) as compared to 40 normal subjects. Data were recorded in the frontal, thalamic and temporal areas; quantification of the MRS signals demonstrated a statistically significant decrease of the Cho/Cr ratio in the frontal and thalamic areas but not in the temporal area for patients with Cushing's syndrome. The largest decrease in Cho/Cr was measured in the thalamic area of patients with a Cushing's syndrome secondary to an adrenal disease. No statistically significant changes in the NAA/Cr ratio were measured in any of the areas studied. These results suggest that the quantification of choline levels could be helpful for monitoring the cerebral metabolite alterations in patients with hypercortisolism. PMID- 10516619 TI - Quantitative 19F NMR study of trifluorothymidine metabolism in rat brain. AB - Metabolism of trifluorothymidine (TFT) and its transport across the blood-brain barrier (BBB) has been measured quantitatively in rats by fluorine-19 nuclear magnetic resonance spectroscopy ((19)F NMR). It is demonstrated that TFT crosses the BBB in micromolar quantities and is metabolized in brain tissue primarily to its free base trifluoromethyluracil (TFMU) by the enzyme thymidine phosphorylase (TP). It is further proposed that the rate of TFMU production can be used as a measure of cerebral TP. The glycols of both TFMU, and to a lesser degree TFT, are generated via an oxidative route. In contrast, the major pathway for hepatic metabolism of this compound is through reduction of the nitrogen base moiety and generation of 5-6-dihydro species followed by ring degradation. Thus, in addition to TFMU as well as the dihydroxy (glycol)-, and the dihydro-species of both TFT and TFMU, alpha-trifluoromethyl-beta-ureidopropionic acid (F(3)MUPA) and alpha trifluoromethyl-beta-alanine (F(3)MBA) were detected in liver extracts. The total metabolite levels in liver were 2-5 times higher than in the brain. Low levels of fluoride ion were detected in all the extracts from brain and liver, as well as blood and urine. This study characterizes TFT as a potential chemotherapeutic agent for use against brain tumors. PMID- 10516620 TI - Pre-polarized saline: an in vivo feasibility study of a potential contrast agent. AB - The potential for using pre-polarized liquids as contrast agents in vivo is investigated and the feasibility of the method demonstrated. In this study we show the enhancement obtained following intravenous delivery of pre-polarized saline into the antecubital vein of a volunteer. This form of contrast agent provides signal gain on time scales commensurate with its T(1) and allows repeated doses to be administered, thus making alternate acquisitions of data with and without enhancement practicable. PMID- 10516621 TI - The effect of dichloroacetate and alanine on the metabolic recovery of perfused mouse liver after cold ischemia. AB - Pyruvate dehydrogenase has been thought to be involved in the improved recovery of livers, from fasted donors, reperfused with alanine after cold preservation. The aim of this work was to investigate the effect on perfused mouse liver of dichloroacetate, an activator of this enzyme. Livers from fed and fasted animals were perfused with oxygenated Krebs-Henseleit buffer for 30 min, then stored at 4 degrees C in University of Wisconsin solution for 48 h. Then reperfusion at 37 degrees C was performed with Krebs-Henseleit buffer containing 2 mM dichloroacetate for 1 h. (3-(13)C)Alanine (8 mM) was then added and perfusion was continued for a second hour. (31)P-NMR was used to measure nucleoside triphosphate recovery of the livers. At the end of reperfusion, (13)C-NMR spectra of perfusates were recorded. Dichloroacetate (DCA) was found to activate pyruvate dehydrogenase in all cases. However, it decreased the functional recovery of livers from both fed and fasted mice. In order to study the effect of alanine on this DCA deleterious effect, we reperfused the livers according to a modified protocol. The first hour of perfusion without alanine was omitted and the organs were reperfused directly for 1 h in the presence of 2 mM dichloroacetate and 8 mM (3-(13)C)alanine. In this protocol, the deleterious effect of DCA was completely suppressed for livers from fasted mice. These results led to the conclusion that the specific beneficial effect of alanine on livers from fasted livers persists in the presence of DCA and thus cannot be explained solely by the induction of a greater pyruvate dehydrogenase reaction rate. PMID- 10516622 TI - 31P and 1H NMR spectroscopic studies of liver extracts of carbon tetrachloride treated rats. AB - NMR spectroscopy was used to examine hepatic metabolism in cirrhosis with a particular focus on markers of functional cellular hypoxia. (31)P and (1)H NMR spectra were obtained from liver extracts from control rats and from rats with carbon tetrachloride-induced cirrhosis. A decrease of 34% in total phosphorus content was observed in cirrhotic rats, parallelling a reduction of 40% in hepatocyte mass as determined by morphometric analysis. Hypoxia appeared to be present in cirrhotic rats, as evidenced by increased inorganic phosphate levels, decreased ATP levels, decreased ATP:ADP ratios (1.72 +/- 0.40 vs 2.48 +/- 0.50, p < 0.01), and increased inorganic phosphate:ATP ratios (2.77 +/- 0.48 vs 1.62 +/- 0.24, p < 0.00001). When expressed as a percentage of the total phosphorus content, higher levels of phosphoethanolamine and lower levels of NAD and glycerophosphoethanolamine were detected in cirrhotic rats. Cirrhotic rats also had increased phosphomonoester:phosphodiester ratios (5.73 +/- 2.88 vs 2.53 +/- 0.52, p < 0.01). These findings are indicative of extensive changes in cellular metabolism in the cirrhotic liver, with many findings attributable to the presence of intracellular hypoxia. PMID- 10516623 TI - On the role of MRS in drug development. PMID- 10516624 TI - In vivo pharmacokinetics using MRS. PMID- 10516625 TI - Exploring the dynamics of regulation of G protein-coupled receptors using green fluorescent protein. PMID- 10516626 TI - Response-selective C5a agonists: differential effects on neutropenia and hypotension in the rat. AB - Some in vivo activities of two complement C5a agonist analogues have been evaluated by measuring changes in blood pressure and neutropenia in the rat and comparing the results with their receptor affinities in peritoneal macrophages and polymorphonuclear leucocytes (PMNs). In vitro C5a receptor (C5aR) binding experiments showed that YSFKPMPLaR and YSFKD(NMeNle)PlaR had similar affinities for the macrophage C5aR (IC50 0.2, 0.1 microM respectively). In PMNs, the affinity of YSFKPMPLaR (IC50 0.1 microM) was similar to that in macrophages, whereas the affinity of YSFKD(NMeNle)PLaR for the PMN C5aR was >100 microM. Given i.v., YSFKD(NMeNle)PLaR had similar activity to YSFKPMPLaR on blood pressure but did not cause neutropenia. These results demonstrate selectivity of a new C5a agonist in vitro, which is paralleled in vivo. The results suggest the possibility of developing selective agonists of C5a for in vivo use in humans. PMID- 10516627 TI - Autoradiographical and behavioural effects of a chronic infusion of antisense to the alpha2D-adrenoceptor in the rat. AB - 1. The aims of this study were, firstly to use receptor autoradiography to investigate the effect of antisense oligonucleotides to the alpha2D-adrenoceptor on receptor binding and, secondly to measure behavioural and physiological parameters to determine whether the chronic antisense infusion had any effect on alpha2-adrenoceptor function in vivo. 2. A 3 day infusion of antisense to the alpha2D-adrenoceptor significantly reduced specific [3H]-RX821002 binding in the septum (20 - 30%) and anterior hypothalamic area (20 - 30%). beta-Adrenoceptor expression was unaffected in those brain areas examined, indicating the antisense knockdown was specific to the alpha2-adrenoceptors. 3. On the second day of the infusion, the hypothermic response to UK 14,304 was significantly attenuated in the antisense-treated group compared with both vehicle and mismatch controls. The effect was fully reversible and a similar decrease in body temperature was observed in all the treatment groups 4 days after the end of infusion. 4. During the second day of the infusion, the effects of UK 14,304 on behaviour were reduced in the antisense-treated rats, but were not significantly lower than those of the vehicle and mismatch, UK 14, 304 controls. These trends were not observed 4 days after the end of the infusion. 5. In conclusion, antisense has been shown to selectively knockdown alpha2-adrenoceptor expression in specific brain areas. The consequence of this knockdown is a significant attenuation of UK 14,304-induced hypothermia and a reduction in its sedative actions. These changes were fully reversed 4 days after the end of the infusion. PMID- 10516628 TI - Anti-atherosclerotic effects of an angiotensin converting enzyme inhibitor and an angiotensin II antagonist in Cynomolgus monkeys fed a high-cholesterol diet. AB - 1. We investigated the relationship between angiotensin II formation and the development of atherosclerotic lesions in the aorta of monkeys (Macaca fascicularis) fed a high-cholesterol (4% cholesterol and 6% corn oil) diet for 6 months, and studied the effects of an angiotensin converting enzyme (ACE) inhibitor, trandolapril (10 mg kg-1 per day, p.o.), and an angiotensin II type 1 receptor antagonist, 2-butyl-4-(methylthio)-1 [[2'[[[(propylamino)carbonyl]amino]sulfonyl] (1,1'-biphenyl)-4-yl]methyl]-1H imidazole-5-carboxylate (HR 720; 20 mg kg-1 per day, p.o.). 2. The level of low density lipoprotein was significantly increased by the cholesterol diet, whereas that of high-density lipoprotein was significantly decreased. The relative areas of the atherosclerotic lesions in the thoracic aorta in the normal and cholesterol-diet groups were 1.3+/-0.3 and 64+/-10%, respectively. 3. Plasma renin and ACE activities showed no differences between the normal and cholesterol diet groups. ACE activity and the concentration of angiotensin II were significantly increased in the aorta of the cholesterol-fed monkeys. 4. Trandolapril and HR 720 decreased significantly the area of the atherosclerotic lesions in the thoracic aorta of cholesterol-fed monkeys, but not the mean blood pressure and the levels of low-density and high-density lipoproteins. 5. In plasma and aorta, trandolapril, but not HR 720, decreased significantly the ACE activities in the cholesterol-fed monkeys, while both of these drugs decreased significantly the angiotensin II levels. 6. In conclusion, blockade of angiotensin II function in vascular tissues by trandolapril or HR 720 may play an important role in preventing the development of atherosclerotic lesions. PMID- 10516629 TI - Evidence for a role for central 5-HT2B as well as 5-HT2A receptors in cardiovascular regulation in anaesthetized rats. AB - 1. The effects of injections i.c.v. of quipazine, (2 micromol kg-1) and 1-(2,5-di methoxy-4-iodophenyl)-2-aminopropane (DOI; 2 micromol kg-1) on renal sympathetic and phrenic nerve activity, mean arterial blood pressure (MAP) and heart rate were investigated in alpha-chloralose anaesthetized rats pretreated with a peripherally acting 5-HT2 receptor antagonist. 2. Quipazine or DOI caused a rise in MAP which was associated with a tachycardia and renal sympathoinhibition in rats pretreated (i.c.v.) with the antagonist vehicle 10% PEG. These effects of quipazine were completely blocked by pretreatment with cinanserin (a 5-HT2 receptor antagonist) and attenuated by spiperone (a 5-HT2A receptor antagonist). However, pretreatment with SB200646A (a 5-HT2B/2C receptor antagonist) only blocked the sympathoinhibition, while pretreatment with SB204741 (a 5-HT2B receptor antagonist) reversed the sympathoinhibition to excitation as it also did for DOI. Quipazine also caused renal sympathoexcitation in the presence (i.v.) of a vasopressin V1 receptor antagonist. 3. Injection (i.v.) of the V1 receptor antagonist at the peak pressor response evoked by quipazine alone and in the presence of SB204741 caused an immediate fall in MAP. For quipazine alone the renal sympathoinhibition was slowly reversed to an excitation, while the renal sympathoexcitation observed in the presence of SB204741 was potentiated. In both, the quipazine-evoked tachycardia was unaffected. 4. The data indicate that cardiovascular responses caused by i.c.v. quipazine and DOI are primarily due to activation of central 5-HT2A receptors, which causes the release of vasopressin and a tachycardia. This released vasopressin appears to suppress a 5-HT2A receptor-evoked central increase in sympathetic outflow, which involves the activation of central 5-HT2B receptors indirectly by the released vasopressin. PMID- 10516630 TI - Vasodilatation of intrapulmonary arteries to P2-receptor nucleotides in normal and pulmonary hypertensive newborn piglets. AB - 1. The vasodilator responses of isolated intrapulmonary arteries (IPA) to P2 receptor agonists were investigated during adaptation to extrauterine life in the piglet. The effect of pulmonary hypertension on the normal response was determined after exposing newborn animals to chronic hypobaric hypoxia (51 kPa) for 3 days. 2. Adenosine 5'-triphosphate (ATP), 2-methylthioATP (2-meSATP), adenosine 5-O-(2-thiodiphos-phate) (ADPbetaS) and uridine 5'-triphosphate (UTP) induced a relaxation in normal newborn piglet IPA pre-contracted with prostaglandin F2alpha (PGF2alpha). The relaxations were not affected by removal of the endothelium. The responses to ATP and ADPbetaS increased significantly with age. 3. The relaxation responses of IPA to ATP, 2-meSATP and ADPbetaS continued to increase normally after birth in an hypoxic environment. 4. The results of the study show that vasodilatation of porcine intrapulmonary vessels to nucleotides increased during development from foetus to adult; that the vasodilatation to purines was mediated by P2Y-receptors on the vascular smooth muscle rather than on the endothelium; and that the P2Y-receptor mediated relaxation of IPA remained normal in the pulmonary hypertensive neonate. PMID- 10516631 TI - Vasoconstriction of intrapulmonary arteries to P2-receptor nucleotides in normal and pulmonary hypertensive newborn piglets. AB - 1. The vasoconstrictor responses of isolated intrapulmonary arteries (IPA) to P2 receptor agonists was investigated during adaptation to extrauterine life in the normal piglet and the effect of pulmonary hypertension was studied following exposure of newborn animals to chronic hypobaric hypoxia (51 kPa) for 3 days. 2. At resting tone, alpha,beta-methyleneATP (alpha,beta-meATP) (P2X-receptor agonist) contracted intrapulmonary arteries from adult, but not immature pigs, and repeated application desensitized the response. 3. Adenosine 5'-triphosphate (ATP) induced endothelium-independent relaxation at low concentrations at all ages, a variable contractile response to high concentrations developed by 3 days, becoming larger and consistent by 14 days of age. 4. Uridine 5'-triphosphate (UTP) evoked a contractile response in normal intrapulmonary arteries from foetal to adult life, the magnitude of the response increasing with age. Endothelial removal and pre-incubation with Nomega-nitro-L-arginine methyl ester (L-NAME) (100 microM) increased the contractile response of adult vessels. 5. Pre incubation with alpha,beta-meATP (100 microM), increased the contractile response to UTP in both newborn and adult vessels. ATP-induced relaxations were reduced in newborn vessels but there was no effect on the responses of adult vessels. 6. Responses to UTP, ATP and alpha,beta-meATP of intrapulmonary arteries from newborn piglets exposed to chronic hypobaric hypoxia for 3 days were normal. 7. In summary, UTP elicited marked vasoconstriction of porcine IPA at all ages. UTP and ATP responses were consistent with activation of the P2Y4-receptor recently identified in vascular smooth muscle by others. alpha, beta-meATP induced a small vasoconstriction in the adult probably via the P2X1-receptor. Responses remained normal in neonatal pulmonary hypertension. PMID- 10516632 TI - Constitutive activity of the delta-opioid receptor expressed in C6 glioma cells: identification of non-peptide delta-inverse agonists. AB - 1. G-protein coupled receptors can exhibit constitutive activity resulting in the formation of active ternary complexes in the absence of an agonist. In this study we have investigated constitutive activity in C6 glioma cells expressing either the cloned delta-(OP1) receptor (C6delta), or the cloned mu-(OP3) opioid receptor (C6mu). 2. Constitutive activity was measured in the absence of Na+ ions to provide an increased signal. The degree of constitutive activity was defined as the level of [35S]-GTPgammaS binding that could be inhibited by pre-treatment with pertussis toxin (PTX). In C6delta cells the level of basal [35S]-GTPgammaS binding was reduced by 51.9+/-6.1 fmols mg-1 protein, whereas in C6mu; and C6 wild-type cells treatment with PTX reduced basal [35S]-GTPgammaS binding by only 10.0+/-3.5 and 8.6+/-3.1 fmols mg-1 protein respectively. 3. The delta antagonists N, N-diallyl-Tyr-Aib-Aib-Phe-Leu-OH (ICI 174,864), 7 benzylidenenaltrexone (BNTX) and naltriben (NTB), in addition to clocinnamox (C CAM), acted as delta-opioid receptor inverse agonists. Naloxone, buprenorphine, and naltrindole were neutral antagonists. Furthermore, naltrindole blocked the reduction in [35S]-GTPgammaS binding caused by the inverse agonists. The inverse agonists did not inhibit basal [35S]-GTPgammaS binding in C6mu; or C6 wild-type cell membranes. 4. Competition binding assays in C6delta cell membranes revealed a leftward shift in the displacement curve of [3H]-naltrindole by ICI 174,864 and C-CAM in the presence of NaCl and the GTP analogue, GppNHp. There was no change in the displacement curve for BNTX or NTB under these conditions. 5. These data confirm the presence of constitutive activity associated with the delta-opioid receptor and identify three novel, non-peptide, delta-opioid inverse agonists. PMID- 10516633 TI - The role of P-glycoprotein in blood-brain barrier transport of morphine: transcortical microdialysis studies in mdr1a (-/-) and mdr1a (+/+) mice. AB - 1. The aim of this study was to investigate whether blood-brain barrier transport of morphine was affected by the absence of mdr1a-encoded P-glycoprotein (Pgp), by comparing mdr1a (-/-) mice with mdr1a (+/+) mice. 2. Mdr1a (-/-) and (+/+) mice received a constant infusion of morphine for 1, 2 or 4 h (9 nmol/min/mouse). Microdialysis was used to estimate morphine unbound concentrations in brain extracellular fluid during the 4 h infusion. Two methods of estimating in vivo recovery were used: retrodialysis with nalorphine as a calibrator, and the dynamic-no-net-flux method. 3. Retrodialysis loss of morphine and nalorphine was similar in vivo. Unbound brain extracellular fluid concentration ratios of (-/ )/(+/+) were 2.7 for retrodialysis and 3.6 for the dynamic-no-net-flux at 4 h, with corresponding total brain concentration ratios of (-/-)/(+/+) being 2.3 for retrodialysis and 2.6 for the dynamic-no-net-flux. The total concentration ratios of brain/plasma were 1.1 and 0.5 for mdr1a (-/-) and (+/+) mice, respectively. 4. No significant differences in the pharmacokinetics of the metabolite morphine-3 glucoronide were observed between (-/-) and (+/+) mice. 5. In conclusion, comparison between mdr1a (-/-) and (+/+) mice indicates that Pgp participates in regulating the amount of morphine transport across the blood-brain barrier. PMID- 10516634 TI - Activity profiles of dalargin and its analogues in mu-, delta- and kappa-opioid receptor selective bioassays. AB - 1. To elucidate the structural features ensuring action of [D-Ala2, Leu5] enkephalyl-Arg (dalargin), a series of dalargin analogues were tested for their effectiveness in depressing electrically-evoked contractions of the guinea-pig myenteric plexus-longitudinal muscle preparations (mu- and kappa-opioid receptors) and the vasa deferentia of the hamster (delta-opioid receptors), mouse (mu-, delta- and kappa-opioid receptors), rat (similar to mu-opioid receptors) and rabbit (kappa-opioid receptors). The naloxone KB values in the myenteric plexus were also obtained. 2. [L-Ala2]-dalargin was 19 times less potent than dalargin, and its pharmacological activity was peptidase-sensitive. The ratio of delta-activity to mu-activity for [L-Ala2]-dalargin was 6.78, and KB was 7.9 nM. This emphasizes the role that D-configuration of Ala2 plays in determining the active folding of dalargin molecule as well as in conferring resistance to peptidases. 3. [Met5]-dalargin was equipotent to dalargin in the myenteric plexus, but was more potent in the vasa deferentia of hamster and mouse (KB=5.5 nM). Leu5 and the interdependence of Leu5 and D-Ala2 are of importance for the selectivity of dalargin for mu-opioid receptors. 4. Dalarginamide was more potent and selective for mu-opioid receptors than dalargin, whilst dalarginethylamide, though equipotent to dalarginamide in the myenteric plexus, was more potent at delta-opioid receptors (KB=5.0 nM). [D-Phe4]-dalarginamide and N-Me-[D-Phe4] dalarginamide were inactive indicating the contribution of L-configuration of Phe4 to the pharmacological potency of dalargin. 5. N-Me-[L-Phe4]-dalarginamide possessed the highest potency and selectivity for mu-opioid receptors (the ratio of delta-activity to mu-activity was 0.00053; KB=2.6 nM). The CONH2 terminus combined with the N-methylation of L-Phe4 increased the potency and selectivity of dalargin for mu-opioid receptors. PMID- 10516635 TI - YC-1 potentiates nitric oxide-induced relaxation in guinea-pig trachea. AB - 1. The effects of YC-1 (3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole) on tension, levels of cyclic GMP and cyclic AMP were investigated in guinea-pig trachea. We especially studied the combined effect of YC-1 with exogenous or endogenous nitric oxide on these parameters. 2. YC-1 at the concentration 3 or 10 microM, which caused only minor effect by itself, elicited concentration dependent potentiation of sodium nitroprusside (SNP)-induced tracheal relaxation. This relaxation of YC-1 with SNP was reversed by ODQ. 3. Relaxant responses to electric field stimulation (EFS) in the presence of indomethacin, atropine, guanethidine, alpha-chymotrypsin and histamine were also markedly increased by YC 1 (10 microM). In the presence of L-NAME or ODQ, the relaxant effects to EFS were attenuated and the following addition of YC-1 did not further enhance relaxation. 4. YC-1 (10 microM) or SNP (0.3 microM) alone did not induce significant elevation of cyclic GMP levels in the presence of IBMX, whereas simultaneous application of both compounds markedly elevated the cyclic GMP accumulation. In contrast, the cyclic AMP levels were not altered even at the combination of YC-1 and SNP. Additionally, YC-1 also affected cyclic GMP metabolism, since it inhibited the activity of phosphodiesterase type V in human platelets. 5. YC-1 (30 microM) did not scavenge superoxide anion and had no effect on the removal of superoxide anion by superoxide dismutase in a xanthine/xanthine oxidase system. 6. In conclusion, these results indicate that although YC-1 elicits negligible relaxation of guinea-pig trachea by itself, it can potentiate the relaxant responses of exogenous or endogenous NO. This synergistic response of YC-1 is via the elevation of cyclic GMP contents. PMID- 10516636 TI - Reactive oxygen species generation and histamine release by activated mast cells: modulation by nitric oxide synthase inhibition. AB - 1. We have examined the generation of intracellular reactive oxygen species (ROS) and release of histamine by rat peritoneal mast cells (RPMC) in response to stimulation with antigen (ovalbumin), compound 48/80, nerve growth factor (NGF) and substance P (SP). 2. We have also examined the effects of the non-specific nitric oxide synthase inhibitor, L-NAME (100 microM) upon the release of histamine and generation of intracellular ROS in response to the named secretagogues. 3. Ovalbumin (100 - 1000 microg ml-1), compound 48/80 (0.1 - 100 microg ml-1), NGF (0.1 - 100 microg ml-1), and SP (5 - 50 microM), caused a concentration-dependent release of histamine from RPMC. 4. Ovalbumin (1 ng ml-1 - 0.1 microg ml-1), compound 48/80 (1 - 100 microg ml-1), NGF (1 pg ml-1 - 1 microg ml-1), and SP (0.005 - 50 microM) caused a concentration-dependent generation of intracellular ROS by RPMC. 5. Pre-incubation of RPMC with L-NAME (100 microM) caused a significant enhancement of both histamine release and intracellular ROS from RPMC in response to ovalbumin, compound 48/80, NGF and SP. 6. Our data demonstrate that NGF, SP and ovalbumin are capable of causing intracellular ROS generation by RPMC at lower concentrations than those causing significant histamine release and we speculate that this may contribute to the activation of cytokine production. 7. The data also show that NO modulates histamine release, and ROS generation in response to the secretagogues used. This may have significance in pathologies where NO synthesis is decreased, leading to an increased activation of mast cells. PMID- 10516637 TI - Effect of pre-exposure to vasoconstrictors on isoprenaline-induced relaxation in rat aorta: involvement of inducible nitric oxide synthase. AB - 1. The aim of this study was to determine whether a brief (30 min) episode of contractile receptor stimulation could affect the degree of a subsequent vasorelaxation. Therefore, concentration - relaxation curves of the rat aorta to isoprenaline were compared before and after exposure of the tissue to noradrenaline (100 microM) or prostaglandin F2alpha (PGF2alpha, 100 microM). 2. Exposure to noradrenaline enhanced the second maximal relaxant effect of isoprenaline (from 20 - 95% relaxation). This effect was not due to significant differences in precontraction levels and was not modified by the presence of the endothelium. Treatment with PGF2alpha mimicked the actions of noradrenaline on subsequent vasorelaxation to isoprenaline. 3. Before exposure to noradrenaline (100 microM), forskolin (10 microM) did not produce any significant relaxation of the rat aorta. After exposure to noradrenaline, forskolin caused a concentration dependent relaxation with a maximal effect of more than 90% in rings with and without endothelium suggesting that the change in vasorelaxation to isoprenaline occurred downstream from the beta-adrenoceptor. 4. The increase in relaxation due to exposure to noradrenaline was markedly attenuated by treatment with a protein synthase inhibitor (cycloheximide), a nitric oxide (NO) synthase inhibitor (L-NG nitroarginine methyl ester, L-NAME) and an inhibitor of the activation of soluble guanylyl cyclase (methylene blue). 5. Western blot analysis showed an increase of inducible NO synthase (iNOS) in aortic rings exposed to noradrenaline or PGF2alpha. 6. Together, these findings suggest that pretreatment of rat aorta with noradrenaline or PGF2alpha could induce vascular NOS which would in turn result in an increase in isoprenaline-induced vasorelaxation, this increase occurring downstream from receptor activation. Such a mechanism might participate in cardioprotection during preconditioning induced by noradrenaline. PMID- 10516638 TI - Cannabinoid CB1 receptors fail to cause relaxation, but couple via Gi/Go to the inhibition of adenylyl cyclase in carotid artery smooth muscle. AB - 1. The aim of the current study was to characterize which cannabinoid receptors, if any, are present on rat carotid artery smooth muscle. Additionally, the effects of cannabinoids on carotid artery tone, on cyclic AMP accumulation and on forskolin-induced relaxation were examined in the same tissue. 2. Stimulation of carotid arteries with forskolin (10 microM) significantly increased cyclic AMP accumulation, an effect that was inhibited in a concentration-dependent manner by the cannabinoid receptor agonist, methanandamide. 3. Similar inhibition was seen with the CB1 agonist HU-210 but this inhibition was not mimicked by the CB2 agonist, WIN 55,2212-2. 4. The inhibitory effect of methanandamide on cyclic AMP accumulation was prevented by incubation of the arteries with pertussis toxin and was significantly reduced by LY320135, a selective CB1 antagonist, but not by SR 144528, a CB2-selective antagonist. 5. Methanandamide failed to relax carotid arteries pre-contracted with phenylephrine, but inhibited forskolin-induced relaxation of these arteries. This functional inhibition of relaxation by methanandamide was inhibited by CB1-selective (LY320135 and SR 141716A), but not a CB2-selective antagonist (SR 144528). 6. These data demonstrate the presence of functional G protein-linked cannabinoid receptors of the CB1 subtype in the rat carotid artery, but show that these receptors inhibit cyclic AMP accumulation rather than cause relaxation. PMID- 10516639 TI - Inhibition of NOS-2 expression in macrophages through the inactivation of NF kappaB by andalusol. AB - 1. Andalusol, ent-6alpha,8alpha,18-trihydroxy-13(16),14-labdadiene, is a naturally occurring diterpene, isolated from Sideritis foetens (Lamiaceae). This compound exhibited therapeutic activity when evaluated in in vivo models of paw and ear inflammation (Navarro et al., 1997: Z. Naturforsch., 52, 844-849). The pharmacological effects of this diterpene have been analysed on the activation of the macrophage cell line J774 with lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma). 2. Incubation of J774 macrophages with andalusol (0.1 - 100 microM) inhibited the synthesis of nitrite caused by LPS (1 microg ml-1) in concentration and time-dependent manners. The maximal inhibition was observed when andalusol was added 30 min before LPS stimulation and decreased progressively as the interval between andalusol and LPS challenge increased up to 14 h. 3. Incubation of J774 cells with LPS resulted in the expression of NOS-2 protein (130 kDa) as identified by Western blot analysis. The levels of this enzyme decreased significantly in the presence of andalusol (IC50=10.5 microM), suggesting that this diterpene inhibited NOS-2 expression. 4. Andalusol inhibited nuclear factor kappaB activation, a transcription factor necessary for NOS-2 expression in response to LPS and IFN-gamma. This compound also inhibited the degradation of IkappaBalpha favouring the retention of the inactive NF-kappaB complexes in the cytosol. 5. Related compounds to andalusol but lacking the polyol groups were less effective inhibiting NOS-2 expression in LPS-activated macrophages. The present findings provide a mechanism by which the anti-inflammatory properties of this diterpene could be mediated. PMID- 10516640 TI - Receptor density as a factor governing the efficacy of the dopamine D4 receptor ligands, L-745,870 and U-101958 at human recombinant D4.4 receptors expressed in CHO cells. AB - 1. The relationships between the density of dopamine D4.4 receptors and the agonist efficacies of L-745,870 (3-(4-[4-chlorophhenyl]piperazin-1-yl)-methyl-1H pyrrolo [2, 3-b]pyridine) and U-101958 ((1-benzyl-piperidin-4-yl)-(3-isopropoxy pyridin-2-yl)-methyl-a min e) were investigated in Chinese hamster ovary (CHO) cells, after treatment with the gene expression enhancer, sodium butyrate. 2. In CHO cells expressing D4.4 receptors (CHO/D4 cells), dopamine inhibited forskolin stimulated cyclic AMP accumulation (Emax 56+/-1% inhibition, pEC50 7.4+/-0.1, n=10). U-101958 behaved as a partial agonist (39+/-7% the efficacy of dopamine, pEC50 8.1+/-0.3, n=4), whereas L-745,870 had no detectable agonist effect. 3. Receptor density, as estimated by [3H]-spiperone saturation binding was 240+/-30 fmol mg-1 protein (n=8) in CHO/D4 cell homogenates. It reached 560+/-150 (n=6), 1000+/-190 (n=4) and 840+/-120 (n=4) fmol mg-1 protein after treatment with sodium butyrate (5 mM) for 6, 18 and 48 h, respectively. 4. The increase in receptor density was associated with a gradual enhancement of the agonist effects (increased Emax and pEC50 values) of dopamine. The efficacy of U-101958 (relative to dopamine) doubled and L-745,870 was turned into a partial agonist (efficacy 49% relative to dopamine, pEC50 8. 6+/-0.2, n=6, after 48 h treatment with sodium butyrate). These agonist effects of U-101958 and L-745,870 could be antagonized by spiperone (0.1 microM) but not by raclopride (10 microM). 5. The results show that U-101958 and L-745,870 are partial agonists at human dopamine D4.4 receptors expressed in CHO cells. Their efficacy is governed by receptor density. Agonist effects of these two compounds in vivo cannot be excluded under circumstances of increased receptor levels. PMID- 10516641 TI - Novel antimigraineur dotarizine releases Ca2+ from caffeine-sensitive Ca2+ stores of chromaffin cells. AB - 1. The novel antimigraineur, dotarizine (30 microM), increased cytosolic Ca2+ concentration, [Ca2+]c, in fura-2-loaded bovine adrenal chromaffin cells. This increase was transient, reached a peak in about 2 - 5 min (0.53+/-0.07 microM; n=19) and then declined to basal levels over a further 5 min period. 2. This transient rise of [Ca2+]c was mimicked by 1 microM thapsigargin and by 30 microM cyclopiazonic acid (CPA), but not by 30 microM flunarizine. Both thapsigargin and CPA occluded the effects of dotarizine and vice versa. 3. All three compounds suppressed the transient [Ca2+]c rises induced by caffeine (10 mM, 10 s); blockade induced by thapsigargin was irreversible and that induced by CPA and dotarizine was reversible. 4. Of the three compounds, only dotarizine blocked reversibly the [Ca2+]c spikes induced by short pulses of high K+ (70 mM, 5 s), suggesting that dotarizine blocks voltage-dependent Ca2+ channels but CPA and thapsigargin do not. 5. Dotarizine caused a gradual and reversible depletion of endoplasmic reticulum (ER) Ca2+ in chromaffin cells transfected with ER-targeted aequorin. CPA had a similar effect. 6. These data show that dotarizine shares with thapsigargin and CPA the ability to deplete Ca2+ in the ER; this novel action of dotarizine could be relevant to its prophylactic effects in migraine. Unlike thapsigargin and CPA, however, dotarizine additionally and reversibly blocks Ca2+ entry through voltage-dependent Ca2+ channels. PMID- 10516642 TI - Molecular and pharmacological characterization of a functional tachykinin NK3 receptor cloned from the rabbit iris sphincter muscle. AB - 1. A functional tachykinin NK3 receptor was cloned from the rabbit iris sphincter muscle and its distribution investigated in ocular tissues. 2. Standard polymerase chain reaction (PCR) techniques were used to clone a full length rabbit NK3 receptor cDNA consisting of 1404 nucleotides. This cDNA encoded a protein of 467 amino acids with 91 and 87% homology to the human and rat NK3 receptors respectively. 3. In CHO-K1 cells transiently expressing the recombinant rabbit NK3 receptor, the relative order of potency of NKB>>NKA>/=SP to displace [125I]-[MePhe7]-NKB binding and to increase intracellular calcium, together with the high affinity of NK3 selective agonists (e.g. senktide, [MePhe7]-NKB) and antagonists (e.g. SR 142801, SB 223412) in both assays was consistent with NK3 receptor pharmacology. In binding and functional experiments, agonist concentration response curves were shallow (0.7 - 0.8), suggesting the possibility of multiple affinity states of the receptor. 4. Quantitative real time PCR analysis revealed highest expression of rabbit NK3 receptor mRNA in iris sphincter muscle, lower expression in retina and iris dilator muscle, and no expression in lens and cornea. In situ hybridization histochemistry revealed discrete specific localization of NK3 receptor mRNA in the iris muscle and associated ciliary processes. Discrete specific labelling of NK3 receptors with the selective NK3 receptor agonist [125I]-[MePhe7]-NKB was also observed in the ciliary processes using autoradiography. 5. Our study reveals a high molecular similarity between rabbit and human NK3 receptor mRNAs, as predicted from previous pharmacological studies, and provide the first evidence that NK3 receptors are precisely located on ciliary processes in the rabbit eye. In addition, there could be two affinity states of the receptor which may correspond to the typical and 'atypical' NK3 receptor subtypes previously reported. PMID- 10516643 TI - Enhanced phenylephrine-induced rhythmic activity in the atherosclerotic mouse aorta via an increase in opening of KCa channels: relation to Kv channels and nitric oxide. AB - 1. Mice lacking the apolipoprotein E and low density lipoprotein receptor genes (E degrees xLDLR degrees ) develop atherosclerosis. The aim of this study was to investigate changes in endothelium-dependent vasodilation and vasomotion in thoracic aortic rings of E degrees xLDLR degrees mice. 2. K+-induced contractions of the aorta from E degrees xLDLR degrees mice were stronger than those from control mice. The sensitivity of E degrees xLDLR degrees aorta to phenylephrine (PE) was decreased but the maximal contractions were increased. Acetylcholine induced, but not sodium nitroprusside-induced, relaxations of E degrees xLDLR degrees aorta was decreased. 3. PE induced rhythmic activity in both E degrees xLDLR degrees and control aorta but the amplitude was larger in E degrees xLDLR degrees than in control mice. PE-induced rhythmic activity in both E degrees xLDLR degrees and control aorta was augmented by increase in extracellular Ca2+ concentration, but was abolished by removal of the endothelium, the nitric oxide (NO) synthase inhibitor N-nitro-L-arginine methyl ester, the guanylate cyclase inhibitor LY-83583, high K+ solution and ryanodine. 4. 4-Aminopyridine, a voltage dependent potassium (KV) channel blocker, increased basal tension and induced rhythmic activity in E degrees xLDLR degrees aorta but not in control aorta. 5. The Ca2+-activated potassium (KCa) channel blockers tetraethylammonium and charybdotoxin abolished PE-induced rhythmic activity in E degrees xLDLR degrees aorta. 6. In conclusion, opening of Kv channels in E degrees xLDLR degrees mice aorta is reduced and it is susceptible to be depolarized resulting in Ca2+ entry. The vascular smooth muscle is then dependent on compensatory mechanisms to limit Ca2+-entry. Such mechanisms may be decreased sensitivity to vasoconstrictors, or increased opening of KCa channels by NO via a cyclic GMP-dependent mechanism. PMID- 10516644 TI - Novel strategies for the design of receptor-selective vasopressin analogues: Aib substitution and retro-inverso transformation. AB - 1. We determined the pharmacological profile of novel backbone-modified peptides designed as protease-resistant, selective analogues of AVP. Binding affinities of peptides were determined at both V1A and V2 subtypes of vasopressin receptor (VPR). Biological potencies of selected peptides were tested in pressor and antidiuretic bioassays. 2. Substitution of the achiral alpha-aminoisobutyric acid (Aib) at position 4 or 7 of AVP produced peptides that selectively bound the V2 VPR. Both [Aib4]AVP (140 IU mg-1) and [Aib7]AVP (36 IU mg-1) are selective antidiuretic agonists with little or no activity in uterotonic and pressor assays. 3. [Aib4] and [Aib7] derivatives of the linear V1A-selective antagonist [PhaaDTyr(Et)2Arg6Tyr(NH2)9]AVP bound selectively and with high affinity (Kd 0.51 and 4.1 nM respectively) to the V1A VPR. Bioassays confirmed that these peptides were potent antivasopressor agents (pA2 8.10 and 8.36 respectively). 4. A total retro-inverso strategy was used to prepare protease-resistant mimetics of both AVP and linear V1A-selective antagonists. Cyclic retro-inverso mimetics of AVP did not bind either V1A or V2 VPRs. In contrast, rationally designed retro inverso mimetics of linear V1A-selective antagonists selectively bound the V1A VPR. 5. Our findings indicate novel methods to improve the pharmacodynamic and pharmacokinetic parameters of neurohypophysial hormone analogues which could be equally applicable to other peptide-receptor systems. PMID- 10516645 TI - Acetylcholine-induced relaxation of peripheral arteries isolated from mice lacking endothelial nitric oxide synthase. AB - 1. Acetycholine-mediated relaxations in phenylephrine-contracted aortas, femoral and mesenteric resistance arteries were studied in vessels from endothelial nitric oxide synthase knock-out (eNOS -/-) and the corresponding wild-type strain (eNOS +/+) C57BL6/SV19 mice. 2. Aortas from eNOS (+/+) mice relaxed to acetylcholine in an endothelium-dependent NG-nitro-L-arginine (L-NOARG) sensitive manner. Aortas from eNOS (-/-) mice did not relax to acetylcholine but demonstrated enhanced sensitivity to both authentic NO and sodium nitroprusside. 3. Relaxation to acetylcholine in femoral arteries was partially inhibited by L NOARG in vessels from eNOS (+/+) mice, but relaxation in eNOS (-/-) mice was insensitive to a combination of L-NOARG and indomethacin and the guanylyl cyclase inhibitor 1H-[1,2, 4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ). The L NOARG/ODQ/indomethacin-insensitive relaxation to acetylcholine in femoral arteries was inhibited in the presence of elevated (30 mM) extracellular KCl. 4. In mesenteric resistance vessels from eNOS (+/+) mice, the acetylcholine-mediated relaxation response was completely inhibited by a combination of indomethacin and L-NOARG or by 30 mM KCl alone. In contrast, in mesenteric arteries from eNOS (-/ ) mice, the acetylcholine-relaxation response was insensitive to a combination of L-NOARG and indomethacin, but was inhibited in the presence of 30 mM KCl. 5. These data indicate arteries from eNOS (-/-) mice demonstrate a supersensitivity to exogenous NO, and that acetylcholine-induced vasorelaxation of femoral and mesenteric vessels from eNOS (-/-) mice is mediated by an endothelium-derived factor that has properties of an EDHF but is neither NO nor prostacyclin. Furthermore, in mesenteric vessels, there is an upregulation of the role of EDHF in the absence of NO. PMID- 10516646 TI - Modulation of sibutramine-induced increases in extracellular noradrenaline concentration in rat frontal cortex and hypothalamus by alpha2-adrenoceptors. AB - 1. The effects of sibutramine (0.25 - 10 mg kg-1 i.p.) on extracellular noradrenaline concentration in the frontal cortex and hypothalamus of freely moving rats were investigated using microdialysis. The role of presynaptic alpha2 adrenoceptors in modulating the effects of sibutramine in these brain areas was also determined. 2. Sibutramine induced an increase in extracellular noradrenaline concentration, the magnitude of which paralleled dose, in both brain areas. In the cortex, this increase was gradual and sustained, whereas in the hypothalamus it was more rapid and of shorter duration. 3. In both the cortex and hypothalamus, pretreatment of rats with the alpha2-adrenoceptor antagonist RX821002 (3 mg kg-1 i.p.) potentiated increases in the accumulation of extracellular noradrenaline induced by sibutramine (10 mg kg-1 i. p.), by 7 and 10 fold respectively. RX821002 also reduced the latency of sibutramine to reach its maximum effect in the cortex, but not in the hypothalamus. 4. Infusion of RX821002 (1 microM) via the probe increased the accumulation of extracellular noradrenaline induced by sibutramine (10 mg kg-1 i.p.) in both brain areas. In the hypothalamus, the effects of RX821002 on the accumulation of noradrenaline induced by sibutramine were 2 fold greater than those in the cortex. 5. These findings support evidence that sibutramine inhibits the reuptake of noradrenaline in vivo, but that the accumulation of extracellular noradrenaline is limited by noradrenergic activation of presynaptic alpha2-adrenoceptors. Furthermore, the data suggest that terminal alpha2-adrenoceptors in the hypothalamus exert a greater inhibitory effect over the control of extracellular noradrenaline accumulation than do those in the cortex. PMID- 10516647 TI - Block of human aorta Kir6.1 by the vascular KATP channel inhibitor U37883A. AB - 1. A human aorta cDNA library was screened at low stringency with a rat pancreatic Kir6.1 cDNA probe and a homologue of Kir6.1 (hKir6.1) was isolated and sequenced. 2. Metabolic poisoning of Xenopus laevis oocytes with sodium azide and application of the K+ channel opener drug diazoxide induced K+ channel currents in oocytes co-injected with cRNA for hKir6.1 and hamster sulphonylurea receptor (SUR1), but not in oocytes injected with water or cRNA for hKir6.1 or SUR1 alone. 3. K+ channel currents due to hKir6.1+SUR1 or mouse Kir6.2+SUR1 were strongly inhibited by 1 microM glibenclamide. K+-current carried by hKir6.1+SUR1 was inhibited by the putative vascular-selective KATP channel inhibitor U37883A (IC50 32 microM) whereas current carried by Kir6.2+SUR1 or Shaker K+ channels was unaffected. 4. The data support the hypothesis that hKir6.1 is a component of the vascular KATP channel, although the lower sensitivity of hKir6.1+SUR1 to U37883A compared with native vascular tissues suggests the need for another factor or subunit. Furthermore, the data suggest that pharmacology of KATP channels can be determined by the pore-forming subunit as well as the sulphonylurea receptor and point to a molecular basis for the pharmacological distinction between vascular and pancreatic/cardiac KATP channels. PMID- 10516648 TI - Troglitazone and pioglitazone attenuate agonist-dependent Ca2+ mobilization and cell proliferation in vascular smooth muscle cells. AB - 1. The effects of troglitazone and pioglitazone on agonist-induced Ca2+ mobilization and cell proliferation were studied using fluorescent Ca2+ indicator fura-2 AM and incorporation of [3H]-thymidine in rat aortic smooth muscle cells. The patch clamp techniques were also employed. 2. Vasopressin and platelet derived growth factor-BB (PDGF) caused a transient elevation in [Ca2+]i by Ca2+ mobilization from intracellular stores, followed by a sustained rise due to Ca2+ entry. Nicardipine partly inhibited the sustained phase, but La3+ completely abolished it. 3. Troglitazone and pioglitazone did not significantly affect the transient rise elicited by these agonists, but preferentially inhibited the sustained phase of [Ca2+]i. 4. Under voltage clamp conditions, troglitazone and pioglitazone inhibited voltage-dependent L-type Ca2+ current (ICa.L). They also inhibited nonselective cation channels (Icat) elicited by vasopressin in a concentration-dependent manner. The half maximal inhibitory concentrations of troglitazone on ICa.L and Icat were 4.6 and 5.7 microM, respectively. On the other hand, nifedipine and nicardipine did not inhibit Icat. 5. Vasopressin and PDGF increased incorporation of [3H]-thymidine, and nifedipine and nicardipine partly suppressed it. However, the inhibitory effects of La3+ and exclusion of extracellular Ca2+ were more potent than the Ca2+ blocking agents. Troglitazone and pioglitazone also inhibited it concentration-dependently. 6. These results suggest that troglitazone and pioglitazone preferentially inhibited agonist (vasopressin and PDGF)-induced Ca2+ entry and proliferation in rat vascular smooth muscle cells, where the inhibitory effects of thiazolidinediones on ICa.L and Icat might be partly involved. Thus, thiazolidinediones may exert hypotensive and antiatherosclerotic effects. PMID- 10516649 TI - Cannabinoid agonists and antagonists discriminated by receptor binding in rat cerebellum. AB - 1. The effect of allosteric regulators on the binding affinity of a number of cannabinoid receptor ligands of varying efficacy in the rat cerebellum was investigated. 2. Radioligand ([3H]-SR141716A) competition curves were constructed in the presence or absence of sodium ions, magnesium ions and guanine nucleotides. 3. It was found that the presence of these allosteric regulators did not affect the affinity of the two antagonists used but did cause a significant decrease in the affinity of full and partial agonists. 4. This reduction in affinity ranged from a 3.67 fold rightward shift of the displacement curve of a mixed agonist/antagonist (3-(6-cyano-2-hexynyl)-delta-8-tetrahydrocannabinol-O 823) to a 38 fold rightward shift for 3-(1, 1-dimethyl-6-dimethylcarboxamide) delta-8-tetrahydrocannabinol (O-1125), a full agonist. 5. In summary, the results of this study suggest a simple method for the inference of functional data using the classical radioligand binding assay. PMID- 10516650 TI - Thapsigargin-induced endothelium-dependent triphasic regulation of vascular tone in the porcine renal artery. AB - 1. To elucidate the role of thapsigargin-induced Ca2+ entry in endothelial cells in the regulation of vascular tone, changes in Ca2+ and force of smooth muscle were simultaneously monitored in fura-2-loaded strips of porcine renal artery. 2. During phenylephrine-induced sustained contraction, thapsigargin caused an endothelium-dependent triphasic response; an initial relaxation, a subsequent transient contraction, and a sustained relaxation. The initial relaxation and the contraction were associated with a decrease and an increase in [Ca2+]i, respectively. There was no apparent [Ca2+]i decrease during the sustained relaxation. Thapsigargin-induced responses were observed at 10-8 M and higher concentrations, with the maximum response observed at 10-6 M. 3. The transient contraction was inhibited by a cyclo-oxygenase inhibitor (10-5 M indomethacin), a thromboxane A2 (TXA2)/prostaglandin H2 (PGH2) receptor antagonist (10-5 M ONO 3708), and a TXA2 synthase inhibitor (10-5 M OKY-046). 4. During the phenylephrine-induced contraction in the presence of indomethacin, thapsigargin caused an initial, but not a sustained relaxation, in the presence of Nomega nitro-L-arginine methylester (L-NAME). During the contraction induced by phenylephrine plus 40 mM K+-depolarization in the presence of indomethacin, thapsigargin induced both a transient and a sustained relaxation. However, these relaxations were completely abolished in the presence of L-NAME. 5. Thapsigargin caused a large Ca2+ elevation in cultured endothelial cells of the renal artery. The concentration-response relation was thus similar to that for force development in the arterial strips. 6. In conclusion, thapsigargin-induced Ca2+ entry in endothelial cells led to triphasic changes in the tone of the porcine renal artery. The endothelium-dependent contraction was mediated mainly by TXA2. Nitric oxide and hyperpolarizing factor are both involved in the initial relaxation. However, a sustained relaxation was observed which mainly depended on nitric oxide. PMID- 10516651 TI - Evidence that mast cell degranulation, histamine and tumour necrosis factor alpha release occur in LPS-induced plasma leakage in rat skin. AB - 1. In the present study we investigated the role of mast cells during inflammation in rat skin. As the release of several pro-inflammatory mediators, such as histamine and tumour necrosis factor alpha (TNFalpha), occurs following mast cell activation we studied whether mast cell degranulation and the release of both histamine (H) and TNFalpha occurred in a model of lipopolysaccharide (LPS)-induced plasma leakage in rat skin. 2. Plasma leakage in the rat skin was measured over a period of 2 h as the local accumulation of intravenous injection of 125I-human serum albumin (125I-HSA) in response to intradermal injection of LPS. LPS (10 microg site-1) produced an increase of plasma leakage (50.1+/-2.3 microl site-1) as compared to saline (9.0+/-3.2 microl site-1). Histological analysis of rat tissue showed that LPS induced a remarkable mast cell degranulation (59.8+/-2.1%) as compared to saline (13.5+/-2.2%). 3. Ketotifen (10 9 - 10-7 mol site-1), a well-known mast cell-membrane stabilizer, produced a dose related inhibition of LPS-induced plasma leakage by 36+/-3.5%, 47+/-4.0%, 60+/ 3.3% respectively. In addition, ketotifen (10-7 mol site-1) inhibited mast cell degranulation by 59. 2+/-2.7%. 4. Chlorpheniramine maleate (CPM) (10-9 - 10-7 mol site-1), an H1 histamine receptor antagonist only partially inhibited LPS-induced plasma leakage in rat skin (38+/-1.1% at the highest dose). Furthermore, CPM (10 7 mol site-1) did not prevent mast cell degranulation. 5. A polyclonal antibody against TNFalpha (1:500, 1:100, 1:50 v v-1 dilution), locally injected, decreased LPS-induced plasma leakage in the skin by 15+/-2.0%, 24+/-2.1% and 50+/-3.0% respectively. 6. Taken together these results suggest that LPS-induced plasma leakage in rat skin is mediated, at least in part, by mast cell degranulation and by the release of histamine and TNFalpha from these cells. PMID- 10516652 TI - Regulation of induction of nitric oxide synthase and the inhibitory actions of dexamethasone in the human intestinal epithelial cell line, Caco-2: influence of cell differentiation. AB - 1. The inducible isoform of nitric oxide synthase (iNOS) may be involved in the pathogenesis of inflammatory bowel disease. Using the human intestinal epithelial cell line, Caco-2, iNOS expression, regulation and sensitivity to the glucocorticoid, dexamethasone after cytokine exposure and its relationship to the degree of differentiation has been studied. 2. NOS activity, assessed by NO2- and NO3- release, was time-dependently increased after exposure to interferon gamma alone or in combination with interleukin-1beta and tumour necrosis factor alpha. 3. Cytokine-induced iNOS activity was increased with days in culture over 20 days and number of passages, suggesting iNOS up-regulation during enterocyte-like differentiation. This activity was inhibited by the selective iNOS inhibitor 1400 W (0.1 - 100 microM). In addition, iNOS protein induction was confirmed by Western blot. 4. Actinomycin D (5 microg ml(-1) inhibited cytokine-induced iNOS activity, protein expression and mRNA level. Pyrrolidine dithiocarbamate (PDTC: 10 - 200 microM) and 3,4 dichloroisocoumarin (0.1 - 100 microM) reduced cytokine induced iNOS activity and protein expression at both day 10 and 15 after confluence. PDTC also decreased iNOS mRNA levels, suggesting NF-kappaB involvement in its transcription at these times. 5. The tyrphostins A25 and B42 reduced cytokine-induced iNOS activity at both day 10 and 15 after confluence, indicating the JAK-2 kinase is also involved at these times. The tyrphostins also reduced the iNOS protein expression. 6. Dexamethasone (0.1 - 10 microM, for 24 h) reduced cytokine-induced iNOS activity at day 15 and 20 after cell confluence, but not at day 5 or 10. 7. Dexamethasone (5 microM) decreased cytokine-induced iNOS protein expression at day 10 as well as at day 15 after confluence. 8. These findings indicate that iNOS induction and its inhibition by dexamethasone in this human intestinal epithelial cell line is dependent on the degree of differentiation. PMID- 10516653 TI - Tracheal relaxing effects and beta2 adrenoceptor selectivity of S1319, a novel sponge-derived bronchodilator agent, in isolated guinea-pig tissues. AB - 1. S1319 (4-hydroxy-7-[1-(1-hydroxy-2-methylamino)ethyl]-1, 3-benzothiazol-2(3H) one acetate), a novel non-catecholamine beta-adrenoceptor agonist, has been compared with isoprenaline, salbutamol and formoterol for activity in vitro on a range of beta-adrenoceptor containing preparations from guinea-pig. 2. S1319, like isoprenaline, salbutamol and formoterol, relaxed preparations of guinea-pig trachea (contracted by histamine) in a concentration-dependent manner. The relaxing activity of S1319 appeared to be more potent than that of isoprenaline and salbutamol, and similar to that of formoterol (pD2 values of 10.58+/-0.03 vs 7. 60+/-0.01, 7.50+/-0.01 and 10.52+/-0.04, respectively), and was blocked by the beta2-adrenoceptor selective antagonist (ICI 118,551). The intrinsic activity of S1319 was close to 1.0. 3. In the beta1-adrenoceptor containing preparations, guinea-pig right and left atria, a monophasic inotropic response of S1319 was observed. The pD2 value of S1319 for left atrial and right atrial inotropism was 6.70+/-0.15 and 7.81+/-0.01, respectively. 4. The selectivity ratio (trachea/left atrial inotropism) of S1319, formoterol, salbutamol and isoprenaline was 8523, 284, 4.8 and 0.45, respectively. The relative selectivity ratio of S1319 was 18743, 1858 and 30 times greater than that of isoprenaline, salbutamol and formoterol, respectively. 5. Relaxant responses of guinea-pig trachea to S1319 declined rapidly when the agonist was washed from the tissues, with complete recovery within 30 min. The duration of action of S1319 was similar to that of isoprenaline and less than that of salbutamol and formoterol. 6. In summary, S1319, a sponge-derived beta-adrenoceptor agonist, is a potent and selective beta2-adrenoceptor agonist with a short-duration of action in isolated guinea-pig tracheas. PMID- 10516654 TI - Effects of a range of beta2 adrenoceptor agonists on changes in intracellular cyclic AMP and on cyclic AMP driven gene expression in cultured human airway smooth muscle cells. AB - 1. The effects of the selective beta2 adrenoceptor agonists salbutamol, terbutaline and salmeterol and the non-selective beta adrenoceptor agonist isoprenaline on [3H]-cyclic AMP formation and cyclic AMP response element (CRE) driven luciferase expression, assessed using the construct p6CRE/luc, were studied in primary cultures of human airway smooth muscle (HASM) cells. 2. Optimal transfection conditions for transient expression of pGL3 Control were 4 microg DNA/well71 in a 6 well plate and 1.8 microl Transfectam/microg DNA. Expression was maximal at 48 - 72 h. 3. Salbutamol (maximum response 19%, EC50 0.6 microM), terbutaline (maximum response 38%, EC50 2.3 microM) and salmeterol (maximum response 18%, EC50 0.0012 microM) were all partial agonists for cyclic AMP formation compared with isoprenaline (EC50 0.08 microM). However, all of the beta2 adrenoceptor agonists produced increases in CRE-driven luciferase activity, in cultured HASM transfected with the vector p6CRE/luc, which were equivalent or greater (salmeterol) than those seen with isoprenaline. 4. Both salbutamol and salmeterol were more potent at increasing luciferase expression than in elevating cyclic AMP levels in these cells. The potency ratios (EC50 (cyclic AMP)/EC50 (LUC)) for the agents studied were isoprenaline: 0. 2 fold, terbutaline: 3 fold, salbutamol: 24 fold, salmeterol: 38 fold. 5. These data suggest that important quantitative differences exist in the ability of beta2 adrenoceptor agonists to increase whole cell cyclic AMP levels in airway smooth muscle and to drive gene expression via a CRE-driven mechanism. PMID- 10516655 TI - 5-hydroxytryptamine receptors mediating contraction in human small muscular pulmonary arteries: importance of the 5-HT1B receptor. AB - 1. The 5-hydroxytryptamine (5-HT) receptors mediating vasoconstriction in isolated human small muscular pulmonary arteries (SMPAs) were determined using techniques of wire myography and reverse transcription-polymerase chain reaction (RT - PCR). 2. The agonists 5-HT, 5-carboxamidotryptamine (5-CT, unselective for 5-HT1 receptors) and sumatriptan (selective for 5-HT1B/D receptors) all caused contraction and were equipotent (pEC50s: 7.0+/-0.2, 7.1+/-0.3 and 6.7+/-0.1, respectively) suggesting the presence of a 5-HT1 receptor. 3. Ketanserin (5-HT2A selective antagonist, 0.1 microM) inhibited 5-HT-induced contractions only at non physiological/pathological concentrations of 5-HT (>0.1 microM) whilst GR55562 (5 HT1B/1D-selective antagonist, 1 microM) inhibited 5-HT-induced contractions at all concentrations of 5-HT (estimated pKB=7.7+/-0.2). SB-224289 (5-HT1B-selective antagonist, 0.2 microM) inhibited sumatriptan-induced contractions (estimated pKB=8.4+/-0.1) whilst these were unaffected by the 5-HT1D-selective antagonist BRL15572 (0.5 microM) suggesting that the 5-HT1B receptor mediates vasoconstriction in this vessel. 4. RT - PCR confirmed the presence of substantial amounts of mRNA for the 5-HT2A and 5-HT1B receptor subtypes in these arteries whilst only trace amounts of 5-HT1D receptor message were evident. 5. These findings suggest that a heterogeneous population of 5-HT2A and 5-HT1B receptors co-exist in human small muscular pulmonary arteries but that the 5-HT1B receptor mediates 5-HT-induced vasoconstriction at physiological and pathophysiological concentrations of 5-HT. These results have important implications for the treatment of pulmonary hypertension in which the 5-HT1B receptor may provide a novel and potentially important therapeutic target. PMID- 10516656 TI - Structural determinants of the partial agonist-inverse agonist properties of 6' azidohex-2'-yne-delta8-tetrahydrocannabinol at cannabinoid receptors. AB - 1. We have extended previous investigations of four analogues of Delta8 tetrahydrocannabinol (Delta8-THC): 6'-azidohex-2'-yne-Delta8-THC (O-1184), 6' azidohex-cis-2'-ene-Delta8-THC (O-1238) and octyl-2'-yne-Delta8-THC (O-584) and its 1-deoxy-analogue (O-1315). 2. O-1184, O-1238 and O-584 displaced [3H]-CP55940 from specific binding sites on Chinese hamster ovary (CHO) cell membranes expressing CB1 or CB2 cannabinoid receptors, with pKi values of 8.28 to 8.45 (CB1) and 8.03 to 8.13 (CB2). The pKi values of O-1315 were significantly less, 7.63 (CB1) and 7.01 (CB2). 3. All the analogues inhibited forskolin-stimulated cyclic AMP production by CB1-transfected CHO cells (pEC50=9.16 to 9.72). Only O 1238 behaved as a full agonist in this cell line. 4. In mouse vasa deferentia, O 1238 inhibited electrically-evoked contractions (pEC50=10.18 and Emax=70.5%). Corresponding values for O-1184 were 9.08 and 21.1% respectively. At 1 nM, O-1184 produced surmountable antagonism of the cannabinoid receptor agonist, CP55940. However, at 0.1 nM, O-1184 did not attenuate CP55940-induced inhibition of cyclic AMP production by CB1-transfected CHO cells. 5. In CB2-transfected CHO cells, cyclic AMP production was inhibited by CP55940 (pEC50=8.59), enhanced by O-1184 and O-584 (pEC50=8.20 and 6.86 respectively) and not significantly affected by O 1238 or O-1315. 6. At 100 nM, O-1184 and O-1238 produced surmountable antagonism of CP55940 in CB2 cells, decreasing the pEC50 of CP55940 from 8.61 to 7.42 (O 1184) or from 8. 54 to 7.44 (O-1238). 7. These data support the hypothesis that increasing the degree of unsaturation of the aliphatic side-chain of Delta8-THC analogues has little effect on CB1 or CB2 receptor affinity but can reduce CB1 receptor efficacy and reverse the direction of responses elicited at CB2 receptors. PMID- 10516657 TI - Influence of nitrovasodilators and endothelin-1 on rheology of human blood in vitro. AB - 1. The shear stress of flowing blood profoundly influences the release of endothelium-dependent vasodilative and constrictive factors. Conversely, the influence of these mediators such as nitric oxide (NO) or endothelin-1 (ET-1) on blood rheology remains elusive. In the present study the influence of nitrovasodilators and ET-1 on red blood cell (RBC) shape and whole blood viscosity were investigated. 2. Incubation of whole blood with sodium nitroprusside (SNP, 10-5 - 10-2 M), glyceryl trinitrate (GTN, 0.0001 - 0.1 mg mL 1), S-nitroso-N-acetylpenicillamine (SNAP, 10-6 - 10-3 M), and the active metabolite of molsidomine (SIN-1, 10-6 - 10-3 M), but not molsidomine (10-6 - 10 3 M), resulted in significantly increased amounts of methaemoglobin, indicating a relevant interaction with RBCs. Treatment with SNP at 10-2 M induced a marked echinocytosis (morphological index: 2.23+/-0.98 vs -0.17+/-0.10; P<0.001) and increased blood viscosity (haematocrit 45%) at a high shear rate of 94.5 s-1 (6.46+/-0.60 vs 5.07+/-0.35 mPa.s; P<0.01) and a low shear rate of 0.1 s-1 (88.6+/-36.8 vs 42.1+/-11.7 mPa.s; P<0.01). Echinocytosis was probably due to cyanide accumulation. SIN-1 at 10-3 M slightly decreased high shear viscosity (4.88+/-0.28 vs 4. 95+/-0.30 mPa.s; P<0.05). SNAP at 10-3 M slightly increased both high (5.14+/-0.23 vs 5.05+/-0.24 mPa.s; P<0.01) and low shear (53.9+/-7.2 vs 51.2+/-5.9 mPa.s; P<0.05) viscosity. Molsidomine and GTN failed to influence whole blood viscosity. ET-1 (10-9 - 10-6 M) had no effect on RBC shape and viscosity. 3. We conclude that the most important modulators of vascular tone, NO and ET-1, do not affect RBC shape and blood viscosity, which is important from both a physiological and a pharmacological point of view. PMID- 10516658 TI - Evidence that histamine homologues discriminate between H3-receptors in guinea pig cerebral cortex and ileum longitudinal muscle myenteric plexus. AB - 1. The binding of the selective histamine H3-receptor agonist ([3H]-R-alpha methylhistamine) to sites in guinea-pig cerebral cortex and ileum longitudinal muscle myenteric plexus has been characterized and a comparison made of the apparent affinities of a series of H3-receptor ligands. 2. Saturation analysis suggested that [3H]-R-alpha-methylhistamine labelled a homogeneous population of histamine H3-receptors in guinea-pig cerebral cortex (pKD=9.91+/-0. 07; nH=1.07+/ 0.03; n=5) and ileum longitudinal muscle myenteric plexus (pKD=9.75+/-0.21; nH=0.97+/-0.02; n=5). There was no significant difference in the estimated affinity of [3H]-R-alpha-methylhistamine in the two tissues. The cerebral cortex had a significantly higher receptor density (3.91+/-0.37 fmol mg-1 tissue) than the ileum longitudinal muscle myenteric plexus (0. 39+/-0.11 fmol mg-1). 3. Overall, the apparent affinities of compounds, classified as H3-receptor ligands, in cerebral cortex and ileum longitudinal muscle myenteric plexus were well correlated (r=0. 91, P<0.0001) and consistent with the cerebral cortex and ileum longitudinal muscle myenteric plexus expressing histamine H3-receptor population(s) that are pharmacologically indistinguishable by the majority of histamine H3-receptor ligands. However, it was evident that the homologues of histamine within this group of compounds could discriminate between the receptor populations in the two tissues. Thus, the estimated affinity of five imidazole unbranched alkylamines (histamine, homohistamine, VUF4701, VUF4732 and impentamine) were significantly higher in the guinea-pig cerebral cortex than in the ileum longitudinal muscle myenteric plexus assay. PMID- 10516659 TI - Direct block of native and cloned (Kir2.1) inward rectifier K+ channels by chloroethylclonidine. AB - 1. We have investigated the inhibition of inwardly rectifying potassium channels by the alpha-adrenergic agonist/antagonist chloroethylclonidine (CEC). We used two preparations; two-electrode voltage-clamp of rat isolated flexor digitorum brevis muscle and whole-cell patch-clamp of cell lines transfected with Kir2.1 (IRK1). 2. In skeletal muscle and at a membrane potential of -50 mV, chloroethylclonidine (CEC), an agonist at alpha2-adrenergic receptors and an antagonist at alpha1x-receptors, was found to inhibit the inward rectifier current with a Ki of 30 microM. 3. The inhibition of skeletal muscle inward rectifier current by CEC was not mimicked by clonidine, adrenaline or noradrenaline and was not sensitive to high concentrations of alpha1-(prazosin) or alpha2-(rauwolscine) antagonists. 4. The degree of current inhibition by CEC was found to vary with the membrane potential (approximately 70% block at -50 mV c.f. approximately 10% block at -190 mV). The kinetics of this voltage dependence were further investigated using recombinant inward rectifier K+ channels (Kir2.1) expressed in the MEL cell line. Using a two pulse protocol, we calculated the time constant for block to be approximately 8 s at 0 mV, and the rate of unblock was described by the relationship tau=exp((Vm+149)/22) s. 5. This block was effective when CEC was applied to either the inside or the outside of patch clamped cells, but ineffective when a polyamine binding site (aspartate 172) was mutated to asparagine. 6. The data suggest that the clonidine-like imidazoline compound, CEC, inhibits inward rectifier K+ channels independently of alpha receptors by directly blocking the channel pore, possibly at an intracellular polyamine binding site. PMID- 10516660 TI - Differential effects of bepridil on functional properties of troponin C in slow and fast skeletal muscles. AB - 1. Bepridil (BPD) is a pharmacological compound able to bind to the Ca2+ sensor protein troponin C (TnC), which triggers skeletal muscle contraction upon Ca2+ binding. BPD can thereby modulate the Ca2+-affinity of this protein. 2. The Ca2+ sensitizing action of bepridil was investigated on slow and fast isoforms of TnC from skinned slow and fast skeletal muscle fibres, activated by either Ca2+ or Sr2+ ions. 3. Bepridil did not modify the Ca2+ maximal tension of slow and fast fibres, suggesting that binding of the drug to TnC did not induce a change in the number of cross-bridges involved in maximal tension. 4. Sr2+ ions induced lower maximal tension than Ca2+ ions. However, in fast fibres, these lower Sr2+ maximal tensions could be reinforced by bepridil, suggesting an effect of bepridil on the function of site I of fast TnC. 5. Under submaximal tension, bepridil induced an increase in Ca2+ affinity of TnC in both slow and fast fibres. However, slow fibres were more drug reactive than fast fibres, and the increase in tension appeared to be modulated by the Ca2+ concentration. 6. Thus, bepridil exerted a differential effect on slow and fast fibres. Moreover, the results suggest that bepridil was more effective when activation conditions were unfavourable. PMID- 10516661 TI - Catechol-O-methyltransferase activity in CHO cells expressing norepinephrine transporter. AB - 1. We examined the existence of catecholamine metabolizing enzymes (catechol-O methyltransferase, COMT, and monoamine oxidase, MAO) in CHO cells transfected with norepinephrine (NE) transporter (NET) cDNA. 2. NET activity was studied by incubating cells with [3H]-NE (0. 5 microCi ml-1, 20 min) in a Na+ containing medium. Incubation with [3H]-NE lead to [3H] accumulation at 47797+/-4864 d.p.m. per well. Specific inhibitors of NET abolished this uptake. 3. During post-uptake incubation, [3H] leaked rapidly from cells and the extracellular phase comprised 89% of total radioactivity within 40 min. Both [3H] retention and [3H] 'leakage' were largely unaffected by inhibitors for MAO. In contrast, COMT inhibitors, U 0521 and Ro 41-0960, dose-dependently increased intracellular [3H]-NE retention with a maximal increase of 4.5 fold. The EC50 for Ro 41-0960 was 139-times lower than that of U-0521. U-0521 largely inhibited [3H] 'leakage' and doubled the apparent Vmax for [3H]-NE uptake. 4. Addition of U-0521 during uptake incubation increased intracellular NE content by 8 fold. Normetanephrine, the COMT-dependent metabolite of NE, was formed in large quantities during post-uptake incubation. U 0521 significantly inhibited the formation of NMN with an equal preservation of intracellular NE. 5. CHO cells expressing NET possess COMT activity, which is responsible for the metabolism of NE to form lipophilic metabolite normetanephrine. The apparent 'properties' of the NET function expressed in CHO cells changed, after inhibition of COMT, in such a way closer to that described in the native neuronal preparations. PMID- 10516662 TI - On the role of 5-HT1B/1D receptors in modulating transmission in a spinal reflex pathway in the decerebrated rabbit. AB - 1. In decerebrated rabbits, the selective 5-HT1B/1D receptor antagonist GR 127,935 had no significant effects on reflexes evoked in medial gastrocnemius motoneurones by electrical stimulation of the sural nerve, or on arterial blood pressure or heart rate when given by the intrathecal (up to 543 nmol cumulative) or intravenous (up to 1.8 micromol cumulative) routes. 2. In decerebrated, spinalized rabbits, intrathecal GR 127,935 in doses of up to 543 nmol, had no effect on the sural-gastrocnemius reflex. Furthermore, this drug failed to alter enhancement of the sural-gastrocnemius reflex induced by 8-hydroxy-2-(di-n propyl)aminotetralin (8-OH-DPAT), given at 300 nmol kg-1 i.v. 3. In decerebrated, spinalized rabbits, the selective 5-HT1B/1D receptor agonists L-694,247 (cumulative doses of 2 - 243 nmol kg-1 i.v.) and L-741,604 (cumulative doses of 3 - 307 nmol kg-1 i.v.), each caused the sural-gastrocnemius reflex to increase to 140% of pre-drug levels, and arterial blood pressure to rise by about 10 mmHg. Subsequent administration of GR 127,935 at 0.9 - 1.8 micromol kg-1 reversed the pressor effect of the agonists but not the increase in reflexes. The 5-HT1A receptor antagonist WAY-100,635 (185 nmol kg-1 i.v.) also failed to reverse the increase in reflexes, but the 5-HT1B/1D/5-HT2/5-HT7 ligand ritanserin (1.6 micromol kg-1 i.v.) restored reflexes to pre-drug control values after L-741,604 (it was not tested against L-694,247). 4. These data indicate that 5-HT1B/1D receptors do not significantly modulate transmission in the sural-gastrocnemius reflex pathway, and that the enhancement of reflexes by 8-OH-DPAT and L-741,604 is probably mediated by 5-HT7 receptors. PMID- 10516663 TI - Characterization of chemokine CCR3 agonist-mediated eosinophil recruitment in the Brown-Norway rat. AB - 1. The ability of various C-C chemokines to elicit tissue eosinophil infiltration following intradermal injection or peripheral blood eosinophilia following intravenous injection were compared in the Brown-Norway rat. 2. Eotaxin (0.1 - 3 microg site-1) of human and murine origin produced equivalent, dose-dependent increases in eosinophil peroxidase activity in rat dermis 4 h post-injection. 3. Human eotaxin-2 was equipotent with human eotaxin in terms of dermal eosinophil recruitment. Other human CCR3 agonists, such as MCP-3, RANTES and MCP-4 failed to increase dermal eosinophil peroxidase activity at doses up to 1 microg site-1 whereas the latter did produce a small effect at 3 microg site-1. 4. Consistent with observations in vivo, human eotaxin displaced [125I]-eotaxin from rat spleen membranes more potently (IC50=2 nM) than did MCP-4 (IC50=500 nM). RANTES did not compete with the radiolabelled chemokine at concentrations up to 1 microM. 5. Human eotaxin (5 microg) administered intravenously increased circulating eosinophils approximately 3 fold whereas MCP-4 (5 microg i.v.) increased circulating monocytes approximately 3 fold without affecting eosinophil numbers. 6. Dexamethasone pretreatment inhibited eotaxin-induced dermal eosinophil influx only at a steroid dose (0.1 mg kg-1, s.c.) which significantly reduced circulating eosinophil numbers. The steroid also reduced eosinophilia in peripheral blood resulting from systemic eotaxin administration (5 microg, i.v.). 7. These data suggest differences in rat CCR3 relative to other species as surmised from a distinctive rank order of chemokine potency. In addition to its chemotactic effects eotaxin, but not MCP-4, promotes eosinophil recruitment into the circulation. One of the mechanisms by which glucocorticoids, such as dexamethasone, acutely inhibits eotaxin-induced dermal eosinophil influx is to diminish the circulating numbers of these cells available for tissue recruitment. PMID- 10516664 TI - Stimulatory effect of exogenous diadenosine tetraphosphate on insulin and glucagon secretion in the perfused rat pancreas. AB - 1. Diadenosine triphosphate (AP3A) and diadenosine tetraphosphate (AP4A) are released by various cells (e.g. platelets and chromaffin cells), and may act as extracellular messengers. In pancreatic B-cells, AP3A and AP4A are inhibitors of the ATP-regulated K+ channels, and glucose increases intracellular levels of both substances. 2. We have studied the effect of exogenous AP3A and AP4A on insulin and glucagon secretion by the perfused rat pancreas. 3. AP3A did not significantly modify insulin or glucagon release, whereas AP4A induced a prompt, short-lived insulin response ( approximately 4 fold higher than basal value; P<0.05) in pancreases perfused at different glucose concentrations (3.2, 5.5 or 9 mM). AP4A-induced insulin release was abolished by somatostatin and by diazoxide. These two substances share the capacity to activate ATP-dependent K+ channels, suggesting that these channels are a potential target for AP4A in the B-cell. 4. AP4A stimulated glucagon release at both 3.2 and 5.5 mM glucose. This effect was abolished by somatostatin. 5. The results suggest that extracellular AP4A may play a physiological role in the control of insulin and glucagon secretion. PMID- 10516665 TI - Involvement of CYP3A-derived arachidonic acid metabolite(s) in responses to endothelium-derived K+ channel opening substance in monkey lingual artery. AB - 1. In monkey lingual artery strips partially contracted with prostaglandin F2alpha, acetylcholine-induced, concentration-related relaxations were abolished by removal of the endothelium. The response was not significantly influenced by indomethacin but attenuated by NG-nitro-L-arginine (L-NOARG); the effect of the nitric oxide (NO) synthase inhibitor was reversed by L-arginine. 2. The response to acetylcholine resistant to L-NOARG was suppressed in the strips exposed to high K+ media. Charybdotoxin partially inhibited the relaxation, and the remaining relaxation was abolished by additional treatment with apamin, whereas glibenclamide, iberiotoxin or apamin alone was without effect. Relaxations induced by sodium nitroprusside were not influenced by charybdotoxin. 3. The L NOARG-resistant acetylcholine-induced relaxation was inhibited by metyrapone, proadifen and 17-octadecynoic acid, non-selective cytochrome P450 mono-oxygenase (CYP) inhibitors, and progesterone and ketoconazole, inhibitors selective to CYP3A. The inhibitors did not affect the nitroprusside-induced relaxation. Selective inhibitors of other CYP isoforms, such as debrisoquine and lauric acid, did not reduce the response to acetylcholine. 4. Reaction mixture containing human liver microsome rich in CYPs, arachidonic acid and NADPH incubated at 37 degrees C and filtrated relaxed endothelium-denuded monkey lingual artery strips, used as bioassay tissues. This response was abolished in the strips exposed to high K+ media. The response was also suppressed by combined treatment of the assay tissue with charybdotoxin plus apamin, but was not affected by treatment with iberiotoxin. The reaction mixture co-incubated with ketoconazole failed to relax the strips. 5. It is concluded that the monkey lingual arterial relaxation dependent on the endothelium is mediated by NO and also by a charybdotoxin plus apamin-sensitive but iberiotoxin-insensitive Ca2+-activated K+ channel opening substance(s) that may be a CYP3A-derived arachidonic acid metabolite(s). PMID- 10516666 TI - Influence of endothelins and sarafotoxin 6c and L-NAME on renal vasoconstriction in the anaesthetized rat. AB - 1. An investigation was performed in pentobarbitone anaesthetized rats to compare the renal vasoconstrictor actions of endothelin-1 (ET-1), endothelin-3 (ET-3) and sarafotoxin 6c and their dependency on NO production. 2. Intra-renal arterial infusion of ET-1 and ET-3, from 1 - 1000 ng had no effect on blood pressure, but reduced renal blood flow maximally by 82 and 81% with EC50 values of 510+/-18 and 1113+/-17 ng, respectively and correspondingly increased renal vascular resistance and decreased conductance. 3. Direct renal arterial administration of sarafotoxin 6c was without effect on blood pressure but caused a maximum reduction in renal blood flow of 56% at 300 ng and had an EC50 of 86+/-4 ng. 4. Administration of the selective ETA receptor antagonist FR139317 at 0.3 and 1.0 mg kg-1 had no effect on basal levels of blood pressure, renal vascular resistance or renal blood flow. The lower dose of FR139317 had no effect on the ET-1 dose-response curve for renal blood flow while at 1.0 mg kg-1, FR139317 reduced the EC50 to 363+/-32 ng (P<0.05). 5. Infusion of L-NAME, 10 microg kg-1 min-1 increased blood pressure by approximately 15%, increased renal vascular resistance and decreased renal blood flow by some 40%. The EC50 values for renal blood flow were reduced to 358+/-68 ng (P<0.05) for ET-1, 638+/-69 ng (P<0.05) for ET-3 and 55+/-10 ng (P<0.01) for sarafotoxin 6c. The maximal reduction in renal blood flow induced by sarafotoxin 6c was raised (P<0.01) from 56% to approximately 100% and renal vascular resistance increased when NO production was blocked. 6. These results showed that the vasoconstrictor actions of ET-1 and ET 3 on resistance vessels controlling renal blood flow are mediated via ETB rather than ETA receptors. Moreover, both ET-1 and ET-3 dependent vasoconstrictions are slightly attenuated by concomitant NO production. By contrast, sarafotoxin 6c appears much more potent at the renal resistance vasculature and is much more powerfully modulated by NO. PMID- 10516667 TI - Adrenocorticotropin reverses vascular dysfunction and protects against splanchnic artery occlusion shock. AB - 1. Tumour necrosis factor (TNF-alpha) is involved in the pathogenesis of splanchnic artery occlusion (SAO) shock. On the other hand, inhibition of TNF alpha is an important component of the mechanism of action of melanocortins in reversing haemorrhagic shock. We therefore investigated the effects of the melanocortin peptide ACTH-(1 - 24) (adrenocorticotropin fragment 1 - 24) on the vascular failure induced by SAO shock. 2. SAO-shocked rats had a decreased survival rate (0% at 4 h of reperfusion, while sham-shocked rats survived for more than 4 h), enhanced serum TNF-alpha concentrations (755+/-81 U ml-1), decreased mean arterial blood pressure, leukopenia, and increased ileal leukocyte accumulation, as revealed by means of myeloperoxidase activity (MPO=9.4+/-1 U g-1 tissue). Moreover, aortic rings from shocked rats showed a marked hyporeactivity to phenylephrine (PE, 1 nM - 10 microM) (Emax and ED50 in shocked rats=7.16 mN mg 1 tissue and 120 nM, respectively; Emax and ED50 in sham-shocked rats=16.31 mN mg 1 tissue and 100 nM, respectively), reduced responsiveness to acetylcholine (ACh, 10 nM-10 microM) (Emax and ED50 in shocked rats=30% relaxation and 520 nM, respectively; Emax and ED50 in sham-shocked rats=82% relaxation and 510 nM, respectively) and increased staining for intercellular adhesion molecule-1 (ICAM 1). 3. ACTH-(1 - 24) [160 microg kg-1 intravenously (i.v.), 5 min after SAO] increased survival rate [SAO+ACTH-(1 - 24)=80% at 4 h of reperfusion], reversed hypotension, reduced serum TNF-alpha (55+/-13 U ml-1), ameliorated leukopenia, reduced ileal MPO (1.2+/-0.2 U g-1 tissue), restored the reactivity to PE, improved the responsiveness to ACh and blunted the enhanced immunostaining for ICAM-1 in the aorta. 4. Adrenalectomy only in part - but not significantly - reduced the ACTH-induced shock reversal, the survival rate of SAO+ACTH-(1 - 24) adrenalectomized rats being 60% at 4 h of reperfusion; and methylprednisolone (80 mg-1 i.v., 5 min after SAO) had a non-significant effect (10% survival) at 4 h of reperfusion. 5. The present data show that melanocortins are effective also in SAO shock, their effect being, at least in part, mediated by reduced production of TNF-alpha. Furthermore, they demonstrate, for the first time, that this inhibition is responsible for the adrenocorticotropin-induced reversal of vascular failure and leukocyte accumulation. PMID- 10516668 TI - Type I and II metabotropic glutamate receptor agonists and antagonists evoke cardiovascular effects after intrathecal administration in conscious rats. AB - 1. In the present study, the role of metabotropic glutamate receptors (mGluRs) in central cardiovascular regulation in conscious rats was examined. To this end, agonists and antagonists for type I and II mGluRs were administered intrathecally, and the temporal changes in blood pressure and heart rate were recorded. 2. L-glutamate (1 micromol) and the prototypical mGluR agonist (1S,3R) ACPD (0.1 and 0. 3 micromol) both increased mean arterial pressure (MAP) and heart rate (HR), implicating functional mGluRs in the spinal cord. The type I mGluR agonist DHPG (0.01 - 0.1 micromol) evoked increases in MAP (max=25+/-5 mmHg) and HR (max=88+/-23 beats min-1). The duration of action, but not the maximum effects, were dose-related and ranged from approximately 10 min to <90 min and 1 min to >90 min for MAP and HR, respectively. 3. The type I/II mGluR agonist CCG-1 (0.1 and 0. 3 micromol) caused smaller, variable increases in MAP and HR of intermediate duration (5 - 20 min), whereas the type II MGluR agonist APDC (0.1 and 1.0 micromol) caused marked, but transient (3 - 5 min), pressor and tachycardic responses. The highest doses of DHPG and CCG-1, but not APDC, also evoked behavioural responses similar to a spontaneous nociceptive behavioural effect reported previously. 4. The type I and II mGluR antagonists (AIDA and LY307452, respectively) were also given approximately 5 min before the administration of the respective type I and II mGluR agonists (DHPG and APDC). Both compounds caused pressor and tachycardic responses, with the effect of AIDA, but not LY307452, returning to control levels before mGluR agonist administration. AIDA significantly attenuated the overall cardiovascular effects of DHPG, while LY307452 significantly attenuated the overall cardiovascular effects of APDC. 5. These results indicate that functional type I and II mGluRs exist in the spinal cord, and that their activation evokes prolonged cardiovascular effects. PMID- 10516669 TI - Cyclic GMP-associated apamin-sensitive nitrergic slow inhibitory junction potential in the hamster ileum. AB - 1. The mediators of non-adrenergic, non-cholinergic (NANC) inhibitory junction potentials (i.j.ps) in the circular smooth muscle cells of the hamster ileum were studied. 2. Electrical field stimulation (EFS; 0.5 ms duration, 15 V) of the intramural nerves with a train of five pulses at 20 Hz evoked a rapidly developing hyperpolarization (fast i.j.p.) followed by a sustained hyperpolarization (slow i.j.p.). 3. NG-nitro-L-arginine methyl ester (L-NAME; 50 200 microM) and NG-nitro-L-arginine (L-NNA; 50 - 200 microM), NO synthase inhibitors, inhibited or abolished the EFS-induced fast and slow NANC i.j.ps. The effects of these NO synthase inhibitors were reversed by L-arginine (5 mM) but not by D-arginine (5 mM). 4. Exogenously applied nitric oxide (NO; 1 - 100 microM) induced concentration-dependent hyperpolarizations. 5. Oxyhaemoglobin (5 50 microM), NO scavenger, inhibited only the slow i.j.p., and the NO-induced hyperpolarization. 6. 1H-[1,2,4]oxadiazolo[4, 3-a]quinoxaline-1-one (ODQ; 10 microM) and cystamine (10 mM), guanylate cyclase inhibitors, inhibited only the slow i.j.p. Zaprinast (100 microM), a phosphodiesterase type V inhibitor, enhanced the amplitude and duration of the slow i.j.p. 7. Apamin (100 nM), a small conductance Ca2+-activated K+ channel blocker, inhibited only the slow i.j.p., and NO-induced hyperpolarization. A high concentration of 8 bromoguanosine 3':5'-cyclic monophosphate (8-bromo-cGMP; 1 mM)-induced membrane hyperpolarization which was blocked by apamin. 8. These results suggest that NO, or a related compound, may be the inhibitory transmitter underlying the apamin sensitive NANC slow i.j.p. and cyclic GMP mediates the slow i. j.p. in the hamster ileum. It is also likely that NO, without involvement of guanylate cyclase is associated with the fast i.j.p. PMID- 10516671 TI - Psychosocial impact of pediatric BMT on siblings. AB - Although bone marrow transplantation (BMT) has become standard therapy for many life-threatening disorders of childhood, there is little research on the psychosocial impact of BMT on siblings of children undergoing BMT. Such siblings face issues common to any family with a chronic illness. However, the psychological impact on the family is intensified because two family members, usually children, are subjected to intrusive medical procedures. Investigators had earlier noted that sibling donors may be at risk for behavioral problems and anxiety, while nondonors may experience ambivalent feelings of disappointment and relief. It was suggested that psychosocial stages of BMT may parallel the medical transplant process, with high levels of stress experienced pre-BMT, during hospitalization, and post-discharge. Our own group has recently conducted more systematic investigations on the psychosocial effects of BMT on donor vs nondonor siblings of surviving pediatric BMT patients. We found that sibling donors showed significantly more anxiety, lower self-esteem, and more adaptive skills in school than nondonors. Nondonors, on the other hand, showed significantly more school problems. One third of the siblings in each group reported a moderate level of post-traumatic stress. Taken together, our research indicates that BMT affects the life of the child at home and at school and that post-traumatic stress symptomatology is a component of the psychological reaction in siblings. The psychosocial adjustment of siblings is a critical area of investigation in BMT populations. Parents need to know that the BMT process affects every member of the family system, and both parents and professionals need to direct more emotional support and attention to siblings. Studies are needed that focus on interventions designed to reduce levels of sibling psychosocial maladjustment. The psychosocial developmental model of post- traumatic stress disorder is a viable theoretical model that may be used to guide future research. PMID- 10516670 TI - Acetylcholine-induced arteriolar dilation is reduced in streptozotocin-induced diabetic rats with motor nerve dysfunction. AB - 1. Diabetes mellitus produces marked abnormalities in motor nerve conduction, but the mechanism is not clear. In the present study we hypothesized that in the streptozotocin (STZ)-induced diabetic rat impaired vasodilator function is associated with reduced endoneural blood flow (EBF) which may contribute to nerve dysfunction. 2. We examined whether diabetes-induced reductions in sciatic nerve conduction velocity and EBF were associated with impaired endothelium-dependent dilation in adjacent arterioles. We measured motor nerve conduction velocity (MNCV) in the sciatic nerve using a non-invasive procedure, and sciatic nerve nutritive blood flow using microelectrode polarography and hydrogen clearance. In vitro videomicroscopy was used to quantify arteriolar diameter responses to dilator agonists in arterioles overlying the sciatic nerve. 3. MNCV and EBF in 4 week-STZ-induced diabetic rats were decreased by 22% and 49% respectively. Arterioles were constricted with U46619 and dilation to acetylcholine (ACh), aprikalim, or sodium nitroprusside (SNP) examined. All agonists elicited dose dependent dilation in control and diabetic rats, although ACh-induced dilation was significantly reduced in diabetic rats. Treating vessels from normal or diabetic rats with indomethacin (INDO) alone did not significantly affect ACh induced relaxation. However, ACh-induced vasodilation was significantly reduced by treatment with KCl or Nomega-nitro-L-arginine (LNNA) alone. Combining LNNA and KCl further reduced ACh-induced dilation in these vessels. 4. Diabetes causes vasodilator dysfunction in a microvascular bed that provides circulation to the sciatic nerve. These studies imply that ACh-induced dilation in these vessels is mediated by multiple mechanisms that may include the endothelial-dependent production of nitric oxide and endothelial-derived hyperpolarizing factor. This impaired vascular response is associated with neural dysfunction. PMID- 10516672 TI - HLA-identical sibling peripheral blood progenitor cell transplants (PBPCT). AB - Peripheral blood progenitor cells (PBPC) are being investigated as an alternative stem cell source in allogeneic transplantation. The current paper presents a 'state-of-the-art' review of HLA-identical sibling transplants (PBPCT). Medline search and meeting reports were used to identify the latest reports from the various transplant centers. The data are presented systematically with the goal of providing a basis for evidence-based medicine. This approach offered the opportunity to identify threshold CD34+ cell numbers for rapid engraftment, threshold CD3+ cell numbers to prevent acute GVHD, prognostic factors for various outcome parameters and are a strong indication that the infused CD3+ cell dose might influence chronic graft-versus-host disease (GVHD), the most controversial and concerning aspect in allogeneic PBPCT. The value of systematic reviews for future clinical research planning is emphasized. PMID- 10516673 TI - Mobilisation of peripheral blood stem cells with IVE and G-CSF improves CD34+ cell yields and engraftment in patients with non-Hodgkin's lymphomas and Hodgkin's disease. AB - The transplantation of mobilised peripheral blood stem cells is associated with more rapid engraftment than marrow transplantation. We have previously reported that G-IVE (G-CSF, ifosphamide, VP-16, epirubicin) improves the yield of CD34+ cells mobilised in patients with lymphoproliferative disorders compared with cyclophosphamide 3 g/m2 and G-CSF (G/CYCLO). In this study we have extended these observations to a larger series of patients including different lymphoma subtypes. Ninety-seven patients undergoing stem cell mobilisation were studied. Forty-two patients with lymphoproliferative disorders received G-IVE for mobilisation and 55 patients G/CYCLO. The median number of mobilised CD34+cells per leucapheresis was significantly higher for those patients receiving G-IVE: 5.82 x 106/kg (0.19-36) compared with 1.2 x 106/kg (0.04-17), P < 0.001 which resulted in a significantly reduced number of leucapheresis procedures performed in the G-IVE group. When patients were analysed dependent on underlying disease G IVE mobilised significantly more CD34+cells per leucapheresis for all lymphoma types reaching 8.41 x 10(6)/kg (0.2-32) compared to 1.32 x 10(6)/kg (0. 06-17) for patients with high-grade NHL mobilised with G-IVE and C-GCSF respectively (P = 0.012). For patients with low-grade NHL 3. 12 x 10(6)/kg (0.10-24.39) compared to 1.08 x 10(6)/kg (0.04-9.74) were collected (P = 0.04) and for patients with Hodgkin's disease 3.02 x 10(6)/kg (1.48-36) and 1.04 x 10(6)/kg (0.1-12.3) (P = 0.001). Mobilisation with G-IVE resulted in the achievement of clinically significant CD34+ cell thresholds in a significantly higher proportion of patients compared to cyclophosphamide and G-CSF reaching >2.5 x 10(6)/kg CD34+ cells in 88% vs 62% (P = 0.004), >5 x 10(6)/kg in 67% vs18% (P = 0.001) and >10 x 10(6)/kg in 31% vs 14.5% (P = 0.05). Furthermore, an analysis of engraftment demonstrated that there was a significant reduction in the time to achieve platelet counts of >20 and >50 x 10(9)/l in patients receiving each incremental dose of CD34+ cells. We conclude that G-IVE mobilizes significantly more CD34+cells than G/CYCLO in patients with lymphoproliferative disorders. This effect is consistent in patients with high-grade NHL, low-grade NHL and HD and results in fewer failed stem collections and increased CD34+ cells available for transplantation which results in significantly accelerated platelet engraftment post transplant. PMID- 10516674 TI - Administration of recombinant human granulocyte colony-stimulating factor to normal donors: results of the Spanish National Donor Registry. Spanish Group of Allo-PBT. AB - A Spanish National PBPC Donor Registry has recently been established for short- and long-term safety data collection in normal donors receiving rhG-CSF. To date, 466 donors have been included in the Registry. Median (range) dose and duration of rhG-CSF administration was 10 microg/kg/day (4-20) and 5 days (4-8), respectively. Donors underwent a median of two aphereses (range, 1-5). Adverse effects consisted mainly of bone pain (90.2%), headache (16.9%) and fever (6. 1%), but no donor discontinued rhG-CSF prematurely due to toxicity. Side-effects were more frequent in donors receiving >10 microg/kg/day than in those with lower doses (82.8% vs 61.8%; P = 0. 004). A significant decrease between baseline and post-apheresis platelet counts was the most important analytical finding (229 x 10(9)/l vs 140 x 10(9)/l; P < 0.0001), with a progressive reduction in platelet count with each apheresis procedure. One donor developed pneumothorax that required hospitalization due to central venous line placement. The mean CD34+ cell dose collected was 6.9 x 10(6)/kg (range, 1.3-36), with only 14 donors (2.9%) not achieving a minimum target of CD34+ cells of 2 x 10(6)/kg. No definitive information about potential long-term side effects is yet available. However, we hope this National Registry will serve as a useful basis for better monitoring of the efficiency and side-effects of cytokine administration in healthy people. PMID- 10516675 TI - Autologous haematopoietic stem cell transplants for autoimmune disease- feasibility and transplant-related mortality. Autoimmune Disease and Lymphoma Working Parties of the European Group for Blood and Marrow Transplantation, the European League Against Rheumatism and the International Stem Cell Project for Autoimmune Disease. AB - This ongoing multicentre prospective phase I/II trial enrolled 74 consecutive patients from 22 centres worldwide with severe autoimmune disease, 35 with rheumatological disorders, 31 with neurological, five with haematological and three with vasculitides. They were treated with autologous peripheral blood or bone marrow transplants according to predetermined criteria. Two patients died after mobilisation before transplant. Seventy-two patients were given 73 transplants, seven bone marrow, and 66 mobilised peripheral blood stem cell transplants. The graft was manipulated to remove T and/or B cells in 43 cases. All 73 transplants engrafted. Five patients died of transplant-related complications: two from bleeding, three from infections. Two patients died of progressive disease. The transplant-related mortality at 1 year of 9% (1-17%; 95% CI) is comparable to the transplant-related mortality of 6% (3-9%; 95% CI) in patients transplanted during the same period in Europe for non-Hodgkin's lymphoma in sensitive relapse (P = 0.39). Sixty patients are evaluable for response, 40 patients (65%) showed some improvement in their disease. Haematopoietic stem cell transplants are feasible for patients with severe refractory autoimmune disease. Transplant-related mortality is comparable to results in patients with non Hodgkin's lymphoma in responsive relapse. Two-thirds of the patients show at least some response. These preliminary data are promising. Although associated with considerable risk, randomised trials comparing autologous stem cell transplants to conventional therapy are warranted. PMID- 10516676 TI - Bone marrow transplantation for therapy-induced acute myeloid leukemia in children with previous lymphoid malignancies. AB - Twenty-one children who developed therapy-related acute myeloid leukemia after treatment for acute lymphoblastic leukemia received allogeneic bone marrow transplants between January 1990 and June 1997. All had previously received epipodophyllotoxin-containing regimens and 11 had cytogenetic abnormalities involving 11q23. Induction chemotherapy was given to 13 patients and eight patients went directly to BMT. Eleven received marrow from matched siblings, eight from matched unrelated donors and two from haploidentical family members. Conditioning regimens included cyclophosphamide (CY), cytarabine, and total body irradiation. Four patients are alive disease-free between 1118 and 1825 days post BMT resulting in a 3-year DFS of 19%. Ten patients relapsed at a median of 150 days (range 30-664 days) post-BMT and all eventually died of disease. Seven patients died of regimen-related toxicity. The outlook for patients with therapy related AML/MDS remains poor and more effective therapy is needed. PMID- 10516678 TI - Tandem transplant of peripheral blood stem cells for patients with poor-prognosis Hodgkins's disease or non-Hodgkin's lymphoma. AB - To improve the results of high-dose therapy with autologous stem cell transplantation, new conditioning regimens with acceptable toxicity must be developed. The aim of this study was to evaluate the feasibility and toxicity of two myeloablative regimens performed at a 2-month interval. After salvage chemotherapy and collection of peripheral stem cell progenitors (median CD34+ cells/kg: 11 x 106/kg), (n = 15) patients with aggressive non-Hodgkin's lymphoma with poor prognostic factors or refractory Hodgkin's disease (n= 9) received intensified regimens. The first conditioning regimen, consisting of BCNU cyclophosphamide-VP16-mitoxantrone was followed by transplantation of a median number of 4 x 10(6) CD34+ cells/kg; then, after a median interval of 56 days, a second preparative regimen, combining busulfan-aracytine-melphalan or TBI + aracytine-melphalan, was followed by transplantation of a median of 4 x 10(6) CD34+ cells/kg. After regimens 1 and 2, respectively: median time to neutrophil recovery >500/microl was 11 days (both times); median time to platelet counts >50,000/microl was 14 and 36 days, but values > 20,000/microl were reached by days 13 and 16 (P = 0.9); mucositis grade III-IV was observed in 11 and 15 cases. The median number of days with fever >38 degrees C was significantly higher (7.8 days) after the second transplant (P <0.05). Three cases of veno-occlusive disease (VOD) were observed after the second transplant. At a median follow-up of 18 months, 14/24 (58%) patients remained in CR, seven patients had died (two of VOD and five after relapse) and two were alive in relapse. These results indicate that tandem transplants performed at a 2-month interval in poor risk lymphoma can be used with acceptable hematotoxicity. VOD remains the major drawback and hepatotoxic drugs, such as busulfan, should be used with caution. Longer term follow-up of a larger cohort of patients is needed to ascertain the overall efficacy. PMID- 10516677 TI - Syngeneic transplantation in multiple myeloma - a case-matched comparison with autologous and allogeneic transplantation. European Group for Blood and Marrow Transplantation. AB - Twenty-five patients with multiple myeloma received bone marrow grafts (n = 24) or peripheral blood stem cells (n = 1) from twin donors. The outcome was compared in a case-matched analysis to 125 patients who underwent autologous transplantation, and 125 who underwent allogeneic transplantation. Seventeen patients (68%) receiving twin transplants entered complete remission, which was not significantly different from that of autologous (48%) or allogeneic (58%) transplants. The median overall and progression-free survival for the twins was 73 and 72 months, respectively. The overall survival tended to be better (73 vs 44 months) and the progression-free survival was significantly better (72 vs 25 months) than with autologous transplantation and both were significantly better than with allogeneic transplantation. Three of 17 patients who entered complete remission following transplantation had relapsed at follow-up. This relapse rate was significantly lower than following autologous transplantation and similar to the relapse rate with allogeneic transplantation. Only two twins died of transplant-related toxicity. Six further patients died of progressive or relapsing disease. Syngeneic transplantation in multiple myeloma appears to be the treatment of choice if a twin donor is available. A lower relapse risk than in autotransplantation may be due to reinfusion of malignant cells in some patients treated with this modality or to the presence of a graft-versus-myeloma effect in some syngeneic transplants. PMID- 10516679 TI - Autologous bone marrow transplantation in non-Hodgkin's lymphoma patients: effect of a brief course of G-CSF on harvest and recovery. AB - This study compares harvest and hematological recovery data of 100 lymphoma patients who underwent BM harvest either after a short course of G-CSF (16 microg/kg for 3 days) (n = 57) or in steady-state conditions (n = 43). G-CSF allowed the attainment of a significantly higher median number of total nucleated cells x 10(8)/kg (4.4, range 1.4-17, vs 2.1, range 0.6-4.2; P < 0.0001), mononuclear cells x 10(8)/kg (0.55, range 0.20-1.4, vs 0.41, range 0.15-0.76, P < 0.0001) and CFU-GM/ml (310, range 10-5500, vs 80, range 10-3800, P = 0.008), with lower volumes of blood collected (17.5 ml/kg, range 8-31 vs 21.0, range 15-30, P = 0.0001). Hematological recovery was faster in patients who received pre-treated BM (median time to PMN >0.5 x 10(9)/l and to platelets >20 x 10(9)/l was 12, range 10-14, and 13, range 10-18, days, respectively) than in those autotransplanted with steady-state BM (median time to PMN >0.5 x 10(9)/l and to platelets >20 x 10(9)/l 13, range 10-18 and 14, range 10-20 days, respectively, P = 0.004 and P = 0.01). Transfusional requirement was significantly different and patients of the G-CSF group needed shorter hospitalization (17 days, range 12-24, vs 20 days, range 14-32; P = 0.02). These data suggest that treating patients with G-CSF before BM harvest improves the quality of the harvest and accelerates engraftment and hematological recovery. PMID- 10516680 TI - Tacrolimus and minidose methotrexate for prevention of acute graft-versus-host disease after HLA-mismatched marrow or blood stem cell transplantation. AB - Thirty adults with leukemia or lymphoma transplanted with marrow or blood stem cells from 1-antigen mismatched related donors received tacrolimus and minidose methotrexate to prevent acute graft-versus-host disease (GVHD). The group had a median age of 42 years (range 18-56 years). Twenty-seven patients had advanced disease, and 13 were resistant to conventional therapy. Tacrolimus was administered at 0.03 mg/kg/day i.v. by continuous infusion from day -2, converted to oral at four times the i.v. dose following engraftment, and continued to day 180 post-transplant. Methotrexate 5 mg/m2 was given i.v. on days 1, 3, 6 and 11. Mild nephrotoxicity was common before day 100; 69% of patients had a doubling of creatinine, 56% had a peak creatinine greater than 2 mg/dl, and two patients were dialyzed. Other toxicities prior to day 100 thought to be related to tacrolimus included hypertension (45%), hyperkalemia (17%), hyperglycemia (14%), seizures (13%), headache (3%) and hemolytic uremic syndrome (3%). Grades 2-4 GVHD occurred in 59% (95% CI, 38-70%), and grades 3-4 GVHD in 17% (95% CI, 1-32%). Overall survival at 1 year was 29% (95% CI, 12-45%). We conclude that tacrolimus and minidose methotrexate is active post-transplant immunosuppression for patients with 1-antigen mismatched donors. PMID- 10516681 TI - Unreliability of carcinoembryonic antigen (CEA) reverse transcriptase-polymerase chain reaction (RT-PCR) in detecting contaminating breast cancer cells in peripheral blood stem cells due to induction of CEA by growth factors. AB - RT-PCR is increasingly used for the detection of minimal residual disease in solid tumors. Carcinoembryonic antigen (CEA) RT-PCR seemed to be highly specific for detection of tumor cells when tested on PBMC. A very high frequency of RT-PCR amplification product for CEA in PBSC from breast cancer patients mobilized with G-CSF was found. However, this result contrasted with tumor cell detection by immunocytochemistry (ICC) which showed no correlation with RT-PCR results. In addition, CEA mRNA was amplified in most G-CSF-mobilized PBSC samples derived from patients with hematological malignancies and from healthy donors of allogeneic stem cells, although no circulating epithelial cells could be demonstrated by ICC. CEA RT-PCR expression was observed in PBMC from healthy individuals incubated in vitro with G-CSF. These data suggest that CEA transcription can be induced by G-CSF, resulting in a loss of specificity of CEA RT-PCR for tumor cell detection in PBMC. We conclude, CEA RT-PCR may not be recommended to detect tumor cell contamination in peripheral blood from patients treated with G-CSF. This may have implications on tumor cell detection by RT-PCR in tissues where endogenous or exogenous growth factors may induce the transcription of CEA or other genes. PMID- 10516682 TI - Accurate quantitation of residual tumor burden at bone marrow harvest predicts timing of subsequent relapse in patients with common ALL treated by autologous bone marrow transplantation. Nagoya BMT Group. AB - We have investigated whether the extent of residual leukemia at bone marrow harvest can predict subsequent relapse after autologous bone marrow transplantation (BMT). A total of 29 pre- and post-purged marrow samples from 15 patients with high-risk common acute lymphoblastic leukemia were examined. An accurate quantitation of residual disease was achieved by phage library assay using polymerase chain reaction to amplify the third complementarity determining region of the immunoglobulin gene. The estimated rate of disease-free survival at 3 years was significantly higher for the patients with less than 5% residual disease among total B cells than for those with greater than 5% before purging (87.5% vs 0%, P = 0. 0013). Furthermore, among patients with subsequent relapse, there was a linear correlation between the quantitated residual tumor burden of pre-purged marrow and remission duration after BMT (r2 = 0. 888). An accurate quantitative assessment of residual disease in the autograft has a high predictive value for subsequent relapse. A serial assay of residual disease would help us to individualize the treatment for each patient after induction or consolidation therapy. PMID- 10516684 TI - Vascular access devices used during harvest of peripheral blood stem cells: high complication rate in patients with a long-term dialysis central venous catheter. AB - PBSC harvesting requires good quality venous access. The efficacy and complication rate of the venous access devices used during stem cell harvest in 101 consecutive patients were examined. Four different categories of venous access were used: (1) long-term dialysis central venous catheter (dCVC), (2) short-term dCVC, (3) peripheral venous cannulae (PVC), and (4) PVC and conventional central venous catheter. The number of harvest occasions per patient or harvest days per occasion were similar between the various categories of access. Complications during harvest occurred in 13 out of 48 (27%) occasions using a long-term dCVC compared to six out of 97 (6%) in the other three categories pooled together (P < 0.01). Forty-two of the 101 patients received a long-term dCVC to facilitate the harvest. The long-term dCVC was planned to stay in place and also be used as a conventional i.v. line during the following high dose treatment. Twenty-one (50%) of the long-term dCVCs were removed due to complication. Thirteen (31%) of the long-term dCVCs were usable throughout the entire treatment period. In conclusion, we recommend that PBSC harvesting is performed through peripheral venous catheters when practically possible, otherwise via short-term dCVC. PMID- 10516683 TI - Optic disc and retinal microvasculopathy after high-dose chemotherapy and autologous hematopoietic progenitor cell support. AB - The purpose of this study was to prospectively evaluate the retinal and optic nerve changes in patients undergoing high-dose chemotherapy (HDC) followed by autologous hematopoietic progenitor cell support (AHPCS). One hundred and forty patients undergoing HDC and AHPCS underwent extensive pre- and post-transplant ophthalmologic evaluations for development of retinal microvascular complications. One hundred and ten patients received high-dose cyclophosphamide, cisplatin and BCNU; thirty received identical doses of cyclophosphamide and cisplatin, but received paclitaxel instead of BCNU. Thirty-four patients (24%) had retinal findings of either cotton wool spots (CWS) (n = 20) or retinal hemorrhages (n = 18) during follow-up, which ranged from 1 to 12 months. Ten (7%) of these patients, all of whom received BCNU, showed ocular toxicity characterized by CWS 1 to 4 months post transplant (n = 10); optic disc edema (n = 3); and variable vision loss associated with the onset of BCNU-induced pulmonary toxicity. Retinal and optic disc microvascular complications may occur after high-dose chemotherapy followed by AHPCS. The association of ischemic retinal lesions and/or optic disc edema with BCNU-induced pulmonary toxicity and the lack of ocular toxicity in patients that did not receive BCNU may suggest that BCNU is the etiologic agent. PMID- 10516685 TI - Stool smears for diagnosis of intestinal acute graft-versus-host disease. AB - A patient with severe diarrhea was successfully diagnosed as having acute intestinal GVHD on stool smear through detection of detached intestinal epithelial cells with apoptosis. Since a stool smear can be easily obtained non invasively, it is a possible tool for the diagnosis of acute intestinal GVHD. PMID- 10516686 TI - Failure of allogeneic bone marrow transplantation to correct Diamond-Blackfan anaemia despite haemopoietic stem cell engraftment. AB - We report the case of a 10-year-old boy with congenital pure red cell aplasia (Diamond-Blackfan anaemia) who received an allogeneic bone marrow transplant (BMT) from his HLA-identical sister. The transplant was complicated by moderate veno-occlusive disease (VOD). Despite cytogenetic evidence of complete donor haemopoietic stem cell engraftment there was selective failure of red cell engraftment and he remains red cell transfusion-dependent. This is the first case of a stem cell transplant failing to correct the defect in this condition despite engraftment. PMID- 10516687 TI - An unusual case of intrapulmonary granulocytic sarcoma presenting as interstitial pneumonitis following allogeneic bone marrow transplantation for acute myeloid leukaemia and responding to donor lymphocyte infusion. AB - We report a 45-year-old female with AML who underwent a T cell-depleted sibling allograft and relapsed a year later with extramedullary disease involving the lung parenchyma and presenting with the clinical and radiological features of interstitial pneumonitis. The patient was treated with donor lymphocyte infusion (DLI) resulting in complete resolution of the radiological signs. The unusual presentation and the management options are discussed. PMID- 10516688 TI - Vaginal outflow tract obstruction by graft-versus-host reaction. AB - We describe a 25-year-old patient suffering from vaginal outflow obstruction which presented as secondary amenorrhea during hormone replacement therapy. The patient had undergone bone marrow transplantation for acute myelocytic leukemia, which caused ovarian failure. Oral mucositis associated with a chronic GVH reaction also occurred. For 3 years she was treated with HRT and had regular menses which gradually ceased and were associated with dyspareunia and abdominal cramps. Abdominal US examination demonstrated hematocolpus. Sonography guided adhesiolysis of a dense vaginal obstruction allowed free drainage with histologic confirmation of a graft-versus-host reaction. The possibility of vaginal stricture or obstruction should be considered in all patients after BMT who suffer from graft-versus-host disease. PMID- 10516689 TI - Syngeneic bone marrow transplantation for small cell carcinoma of the esophagus. AB - Small cell carcinoma (SCC) of the esophagus is an aggressive tumor which behaves biologically like its pulmonary counterpart. Since standard chemotherapy produces few long-term survivors, high-dose chemotherapy regimens with autologous progenitor cell support have been investigated to improve outcomes. Although these strategies have demonstrated improved response rates, overall survival has not been significantly impacted. Here, we report a unique case of a patient with stage IV SCC of the esophagus who underwent high-dose chemotherapy followed by syngeneic bone marrow transplantation. He remains disease-free 38 months post transplant with minimal long-term sequelae. PMID- 10516690 TI - Methotrexate resistance conferred by transplantation of drug-resistant transgenic marrow cells fractionated bycounterflow elutriation. AB - Introduction of genes conferring drug resistance into hematopoietic cells may allow for improved chemotherapy by protection of normally drug-sensitive cells from the toxic side-effects of antitumor agents. We recently reported that transplantation of murine marrow transgenic for drug-resistant dihydrofolate reductase (DHFR) protected mice from lethal doses of methotrexate (MTX), demonstrating the feasibility of imparting drug resistance to recipients of marrow expressing drug-resistant DHFR activity. In order to optimize this strategy it is necessary to identify the hematopoietic cell populations which mediate drug resistance. For this purpose, we separated committed progenitor populations from primitive hematopoietic cells in DHFR transgenic marrow by counterflow elutriation (CE). As expected, supplementation with a fraction containing committed progenitors afforded protection from MTX-induced aplasia observed early after transplantion in animals administered MTX. In contrast, supplementation with a fraction containing primitive hematopoietic cells depleted of committed progenitors failed to provide immediate protection from early aplasia, but instead contributed to drug resistance 4 to 5 weeks after transplantation. The presence of primitive hematopoietic progenitors in both fractions was evident from Southern hybridization analysis for donor transgenic cells 2 months post-transplant. We conclude that protection from aplasia associated with MTX administration immediately after transplantation requires expression of drug-resistant DHFR activity in more differentiated, committed hematopoietic cells, while primitive DHFR transgenic progenitors contribute to long-term drug resistance. These results help to define the appropriate target cells for improved chemotherapy by protection of hematopoietic cells through the introduction of new genes conferring drug resistance. PMID- 10516691 TI - Effect on cytokine release and graft-versus-host disease of different anti-T cell antibodies during conditioning for unrelated haematopoietic stem cell transplantation. AB - Three different types of anti-T cell antibody were used in patients undergoing haematopoietic stem cell transplantation (HSCT) with an HLA-A, -B and -DR compatible unrelated donor: ATG-Fresenius (ATG-F) (n = 26), Thymoglobuline (TMG) (n = 61) and OKT-3 (n = 45). The groups were comparable regarding diagnosis, stage, age, conditioning and GVHD prophylaxis, Adverse events were less frequent after ATG-F treatment. Levels of IL-2, IL-6, IFN-gamma, TNF-alpha and GM-CSF were increased after OKT-3 infusion. In multivariate analysis OKT-3 treatment (P = 0.01), G-CSF treatment (P = 0.02) and a cell dose >/=2.7 x 108/kg (P = 0.03) gave a faster engraftment. Acute GVHD grades II-IV occurred in 25% of the ATG-F patients, 12% of the TMG-patients and 43% (P < 0.001 vs TMG) of the OKT-3 patients. OKT-3 was associated with acute GVHD in multivariate analysis. TRM was 26% using TMG as compared to 43% in the OKT-3 group (P = 0.03). Patient survival at 4 years was 63%, 50% and 45% in the ATG-F, TMG and OKT-3-treated patients, respectively (NS). Relapses were 8%, 49% and 34%, respectively (ATG-F vs TMG, P = 0.03). Relapse-free survivals were 61%, 40% and 37% (NS). Among CML patients the probability of relapse was 61% in TMG-treated patients, while no patients relapsed in the other two groups. To conclude, the type of anti-T cell antibody affects GVHD and relapse after HSCT using unrelated donors. PMID- 10516692 TI - A prospective randomized trial of Filgrastim (r-metHuG-CSF) given at different times after unrelated bone marrow transplantation. AB - A study was done to compare treatment with Filgrastim (r-metHuG-CSF) given at three different times after unrelated bone marrow transplantation (BMT). Sixty nine patients grafted with HLA-A, -B and -DR-compatible unrelated bone marrow were randomized to Filgrastim (5 microg/kg/day) starting on day 0 (n = 23), day +5 (n = 23) or day +10 (n = 23) after BMT. No significant differences were detected in hematological recovery, days with fever, days on antibiotics, incidence of bacteremia or need for erythrocyte, platelet and granulocyte transfusions between the three groups. Patients given Filgrastim starting on day 0, day +5 or day +10, respectively, reached an absolute neutrophil count (ANC) >0.5 x 109/l on a median of 17, 16 and 16 days after BMT. Starting Filgrastim treatment on day +10, rather than on day 0, reduced the costs of Filgrastim by $1060, with no significant change in the median number of days-to-hospital discharge in the three Filgrastim-treated groups. The incidences of acute and chronic GVHD, transplantation-related mortality, relapse, leukemia-free survival and patient survival (PS) were similar in all groups. PMID- 10516693 TI - Myeloablative chemotherapy with autologous peripheral blood stem cell transplantation for metastatic breast cancer: immunologic consequences affecting clinical outcome. AB - Autologous peripheral blood stem cell transplantation following myeloablative chemotherapy is being increasingly utilized in the treatment of a variety of malignancies. We administered busulfan 16 mg/kg orally, thiotepa 500-700 mg/m2 i.v., and carboplatin 800-1000 mg/m2 i.v. to 56 women with metastatic carcinoma of the breast. Autologous peripheral blood stem cells, which had been collected after a combination of chemotherapy and granulocyte colony-stimulating factor, were infused on day 0. The major toxicities of the conditioning regimen included severe pancytopenia, stomatitis, nausea, emesis, diarrhea, fever, and infection. Transplant-related mortality was 1.8%. The incidence of opportunistic viral infections was 42.9%. Fourteen individuals achieved a complete response. The actuarial survival at 1223 days was 13.7% for the entire group of patients; the actuarial survival at 1009 days was 39.3% among complete responders. The functional status of the immune system was determined following transplantation in a subset of patients. Peripheral blood mononuclear cells were obtained before and after stem cell infusion, and were analyzed phenotypically and functionally. Proliferative and interleukin-2 synthetic ability of these cells was assessed following stimulation with phytohemagglutinin and anti-CD3 antibody. The response to influenza peptides was also ascertained. Proliferative and interleukin-2 synthetic capacity was markedly impaired for over a year. Memory response was virtually absent for up to 2 years following transplantation. The prolonged and marked immunosuppression following this myeloablative regimen was associated with a high incidence of opportunistic viral infections, and may have contributed to disease relapse and progression especially in patients who failed to achieve a complete response following transplantation. PMID- 10516694 TI - High-dose peripheral blood stem cell transplant for multitransfused severe aplastic anaemia patients without antithymocyte globulin in the conditioning regimen. AB - Four multitransfused patients with severe aplastic anaemia (SAA) are described. Two received a BMT after conditioning with cyclophosphamide (Cy) plus antithymocyte globulin (ATG). Both suffered a graft failure (GF) and had a second transplant with PBSC from the original donor. Two other patients received a PBSCT as a first option, with Cy as the only conditioning drug. The four patients received methotrexate (MTX) and cyclosporine (CYA) as post-grafting immunosuppression. The two BMT patients with GF were successfully rescued with a PBSC second transplant. In the two cases where a PBSCT was done as a first option no GF was observed and a successful and complete haematological recovery was achieved. In conclusion, PBSCT rescued two SAA patients with GF after BMT. PBSCT without ATG as a first option produced a quick and complete haematological recovery in two additional patients, suggesting that PBSCT without ATG can be an alternative to BMT plus ATG in SAA as a first transplant option. PMID- 10516696 TI - High treatment-related mortality in cardiac amyloid patients undergoing autologous stem cell transplant. AB - Dose-intensive chemotherapy with PBSC support was recently reported to be feasible in cardiac amyloidosis with some patients achieving post-transplant improvement in performance status. At our center, 11 patients with symptomatic primary systemic amyloidosis and predominant cardiac involvement confirmed by biopsy or increased wall thickness on echocardiogram were evaluated for high-dose therapy. The average time from diagnosis to referral was 11 months (4-26 months). Of the 11 patients, two were not candidates for high-dose therapy, based on poor performance status. The remaining nine patients proceeded to PBSC collection. Three patients died during the mobilization period: two of rapid atrial fibrillation, and the third secondary to progressive heart failure. Six patients proceeded to transplantation. However, one died of sudden cardiac arrest the day of melphalan administration, one following hypotension related to stem cell infusion, and one of hypotensive shock the day following stem cell infusion. Three patients recovered and left the hospital, but one died of a cardiorespiratory event at home within 6 weeks of discharge. Both surviving patients demonstrate objective improvement. A decision to use high-dose therapy and stem cell support in cardiac amyloidosis must balance the substantial morbidity of the procedure with the potential benefits. Transplant regimens should avoid cardiotoxic agents such as cyclophosphamide and DMSO and patients should receive anti-arrythmic therapy. PMID- 10516695 TI - Bone marrow transplantation for patients with Fanconi anemia: reduced doses of cyclophosphamide without irradiation as conditioning. AB - Fanconi anemia (FA), a rare autosomal recessive disease, frequently evolves to bone marrow failure and acute myeloid leukemia, and BMT is the treatment of choice for patients with FA. However, their exquisite hypersensitivity to DNA cross-linking agents is associated with severe complications and several investigators have been looking for the ideal preparatory regimen. We have been involved in a program of progressively decreasing doses of cyclophosphamide (CY) as conditioning therapy, in an attempt to identify the lowest dose of CY capable of maintaining the graft with minimum complications. Here, we describe our experience of allogeneic BMT offered to 16 patients with FA and an HLA-compatible sibling donor, conditioned with 100 mg/kg of CY. The actuarial survival is 88% at approximately 37 months. Mucositis >/= grade II was the most common complication (94%), followed by bacteremias (38%). Veno-occlusive disease and hemorrhagic cystitis did not occur. Sustained engraftment was obtained in 94% of patients, and acute and chronic GVHD was diagnosed in 13% and 7%, respectively. The lowest dose of CY for transplant in FA patients is yet to be determined, but further reductions seem possible. PMID- 10516697 TI - Use of a skin explant model for predicting GVHD in HLA-matched bone marrow transplants - effect of GVHD prophylaxis. AB - Over the last decade we have successfully evaluated the use of a human skin explant assay for predicting acute GVHD in HLA-matched sibling transplants. In the present study, we modified GVHD prophylaxis on an individual patient basis depending on the GVHD outcome predicted by the skin explant model. We have summarised our previous data describing how the skin explant assay results correctly predict GVHD occurrence and severity in 45/56 patients (80%); P< 0.0001, chi2 19.97, df = 1. In a further cohort of 19 patients, all were predicted to develop grade II or above GVHD. These patients were given increased GVHD prophylaxis with the addition of methotrexate and a significant reduction in the expected incidence of GVHD was observed (P = 0.02; chi2 7.7, df = 1; Fisher exact test P = 0.04). The results from these studies suggest that modifying GVHD prophylaxis, based on skin explant assay results, may reduce the expected incidence and severity of GVHD. We suggest that the technique might be used for selective GVHD prophylaxis in T cell non-depleted HLA matched sibling transplants. PMID- 10516698 TI - High-dose therapy supported with immunomagnetic purged autologous bone marrow in high-grade B cell non-Hodgkin's lymphoma. AB - From August 1987 to March 1995, 25 patients with high-grade B cell non-Hodgkin's lymphoma (NHL) were treated with high-dose therapy (HDT) followed by bone marrow purged with immunomagnetic beads. At the time of transplantation, 20 patients were in sensitive relapse and five in first complete or partial remission. Ten patients had secondary high-grade NHL transformed from low-grade NHL. The HDT consisted of TBI followed by high-dose cyclophosphamide. All patients engrafted, except for two patients with early treatment-related death. Eleven patients relapsed, of whom nine died of lymphoma, and two are alive in new CR. The estimated event-free and overall survivals at 5 years were 40% and 48%, respectively, with a median follow-up of 48 months (range 1-123). Eight of the tumours contained the translocation t(14;18) at the major breakpoint region (MBR) of BCL-2. In these patients the presence of tumour cells in the bone marrow graft before and after purging were assessed by PCR. Four of five patients infused with non-detectable minimal residual disease in their autografts are in complete remission, while two of three patients reinfused with t(14;18) positive cells after purging, experienced a fast and aggressive relapse. As found by others, our data suggest that reinfusion of tumour-free autografts obtained by efficient in vivo purging using chemotherapy before harvesting, and/or by in vitropurging of the stem cell products, influence the patients remission status after HDT. PMID- 10516699 TI - Persistent changes in the immune system 4-10 years after ABMT. AB - The aim of the present study was to investigate whether the early changes in the immune system observed after ABMT would persist over years. Eighty-five patients with malignant lymphoma were treated with ABMT in Norway from 1987 until 1993. Of the 46 patients in CR by 1997, 36 were enrolled in our study. Median time from ABMT was 5 years (4-10 years). Immunophenotyping showed an increase in the median number of B cells (0.35 x 109/l in patients vs 0.28 x 109/l in controls), and a decrease in T cells (1.08 vs 1.35 x 109/l). Furthermore, a lower median count of CD4+ T cells (0.54 x 109/l in patients vs0.87 x 109/l in controls) resulted in reduced CD4/CD8 ratios (0.8 in patients vs 1.6 in controls). The subgroup of CD4+ T cells expressing the 'naive' phenotype CD45RA was 19.5% in patients vs 38% in controls. In contrast, the fraction expressing the 'memory' phenotype CD45RO was higher in the ABMT group (76% vs 54%). When stimulated, larger fractions of CD3+CD4+ cells in patients produced IFN-gamma (32% vs 16%) or IL-4 (7% vs 1%) compared to controls; thus a differentiation into the functionally separate subgroups Th1 and Th2, with a dominant Th2 response. Our data further suggest that the decrease in CD4+ T cell counts and the imbalance between CD45RA+ and CD45RO+ subsets persists 4-10 years after ABMT. PMID- 10516700 TI - High-dose corticosteroid therapy for diffuse alveolar hemorrhage in allogeneic bone marrow stem cell transplant recipients. AB - In a series of 74 patients with hematological malignancies undergoing allogeneic bone marrow or peri- pheral blood stem cell transplants from an HLA-identical sibling donor, four developed diffuse alveolar hemorrhage (DAH) between days 0 and 23 post transplant. Diagnosis was made by the radiographic finding of diffuse bilateral lung opacities, and bloody lavage fluid on bronchoscopy. Two patients required mechanical ventilatory support. They were treated with methylprednisolone 0.25-1.5 g/day for at least 4 days with slow tapering thereafter. All patients showed an immediate response and two became long-term survivors with normal respiratory function. Two had a relapse of DAH, developed acute respiratory distress syndrome (ARDS) and died with multi-organ failure. Risk factors for DAH were one or more courses of intensive chemotherapy pretransplant vs no treatment or low-dose chemotherapy (4/4 DAH vs 23/70 no DAH; P = 0.015), and second transplants (2/2 DAH vs 1/70 with no DAH; P = 0.006). These results indicate that DAH is life-threatening but is potentially reversible by prompt treatment with high doses of steroids. PMID- 10516701 TI - Bone mineral density after allogeneic bone marrow transplantation. AB - Bone turnover markers and bone mineral density (BMD) were studied in 25 adult patients (14 females, 11 males) who had undergone allogeneic bone marrow transplantation (BMT). The interval from BMT to the first examination was at least 1 year (mean 3, range 1-10). Mean age of the patients at the time of first evaluation was 42 (range 19-54) years. Blood samples and urine collections for evaluation of biochemical factors reflecting skeletal turnover were performed together with the first BMD measurement. BMD was measured from the lumbar vertebrae (L2 to L4) with computed tomography and results were expressed as Z scores. At the time of the first measurement five patients (20%) had Z-scores < 2.5 s.d. and 12 patients (48%) between -1 and -2.5 s.d. In 12 patients BMD assessments were repeated and it seemed that reduction in BMD had mostly occurred during and shortly after BMT and remained the same during follow-up. The cross linked carboxyterminal telopeptide of type I collagen (ICTP) correlated negatively with BMD (r = -0.45, P = 0.045) as did bone-specific alkaline phosphatase (BAP; r = -0.64, P = 0.002). No correlation between BMD and time interval from diagnosis to BMT, conditioning regimen, corticosteroid use or hospital stay during transplantation was found. In conclusion, bone disease is common after BMT. Our findings demonstrate an increased collagen and bone turnover and a high risk of osteoporosis. BMD measurements must be repeated regularly and collagen markers such as ICTP and BAP can be beneficial in estimating the activity of bone disease. PMID- 10516702 TI - Late onset veno-occlusive disease following high-dose chemotherapy and stem cell transplantation. AB - The original definition of hepatic veno-occlusive disease (VOD), which is still widely accepted, includes onset of the clinical syndrome before day +20 following high-dose chemotherapy (HDC) and stem cell transplantation (SCT). We retrospectively identified four patients following HDC and SCT presenting with late onset VOD occurring at day +24, day +27, day +34 and day +42 post SCT. All patients had moderate VOD, with successful resolution of the VOD before day +100 with optimal supportive therapy. Common risk factors for VOD shared by all four patients included an older age (median age: 60 years), and use of a busulphan containing regimen. Mean and maximum bilirubin levels for all patients during the VOD syndrome were 2.02, 1.76, 5.09, 2.87 mg/dl and 2.5, 2.2, 8.9 and 4.1 mg/dl, respectively, which correlated well with duration of VOD. All patients encountered platelet transfusion-dependent thrombocytopenia during VOD. Ursodeoxycholic acid was used as VOD prophylaxis beginning at a mean of 33 days prior to onset of VOD. As the cellular target of hepatic VOD is as yet unidentified, it is uncertain whether ursodiol or other common characteristics of patients with late onset VOD influence the pathogenesis and natural history of this disease. We believe that the uncommon clinical entity of late onset VOD, a potentially fatal regimen-related toxicity, should not be ignored as a diagnosis of liver disease after 3 or more weeks following HDC and SCT. PMID- 10516703 TI - A prospective randomized trial comparing the toxicity and safety of atovaquone with trimethoprim/sulfamethoxazole as Pneumocystis carinii pneumonia prophylaxis following autologous peripheral blood stem cell transplantation. AB - Pneumonia due to Pneumocystis carinii is an infrequent complication following autologous stem cell transplantation (ASCT) which is associated with a high mortality. Although administration of trimethoprim/sulfa- methoxazole (TMP/SMX) is an effective prophylactic strategy for Pneumocystis carinii pneumonia (PCP), treatment-associated toxicity frequently results in discontinuation of therapy. We have conducted a prospective randomized trial comparing atovaquone, a new anti Pneumocystis agent, with TMP/SMX for PCP prophylaxis following autologous peripheral blood stem cell (PBSC) transplantation. Thirty-nine patients were studied. Twenty patients received atovaquone suspension and 19 patients received TMP/SMX. The median ages were 44 (range 20-68) and 47 (range 32-63) years, respectively. A similar number of patients with solid tumors (14 vs 15) and hematologic malignancies (five vs five) were treated in each group. Either TMP/SMX (160/800 mg) or atovaquone (1500 mg) was administered daily from transplant day -5 until day -1, discontinued from day 0 to engraftment, then resumed 3 days per week until day +100 post-transplant. The median time to engraftment (ANC >0.5 x 109/l) was similar in both groups. Eighty percent of the patients randomized to atovaquone prophylaxis completed the study. Four atovaquone-treated patients were removed from study; two patients (10%) did not receive a transplant and two patients (10%) were removed due to a protocol violation. None of the 16 patients treated with atovaquone experienced treatment associated adverse effects. Of the 19 patients randomized to receive TMP/SMX, 55% completed the study. Nine TMP/SMX-treated patients were removed from the study; one patient (5%) did not receive a transplant and eight patients (40%) were removed due to drug intolerance (P < 0.003). The rate of intolerance to TMP/SMX led to the early discontinuation of this randomized trial. Intolerance of TMP/SMX included elevated transaminase levels (n = 1), nausea or vomiting (n = 3), thrombocytopenia (n = 2) and neutropenia (n = 2). All episodes of TMP/SMP intolerance occurred following transplantation after a median duration of 17.5 (range 2-48) days and a median of 7 (range 1-20) doses. Resolution of adverse side-effects occurred in all eight patients within a median of 7 (range 2-20) days following discontinuation of therapy. Neither PCP nor bacterial infections were identified in any of the patients treated. This prospective randomized study demonstrated that atovaquone is well-tolerated for anti-Pneumocystis prophylaxis in autologous PBSC transplant patients intolerant of TMP/SMX. PMID- 10516704 TI - Assessment of psychosocial adjustment in Chinese unrelated bone marrow donors. AB - This descriptive study assessed the experiences of unrelated bone marrow donation in 27 Chinese donors and compared their mood states with a random sample of 78 Chinese adult non-donors using four scales. The donors demonstrated better mood states in terms of anger- hostility and fatigue compared to non-donors. However, their self-esteem was low (similarly to the non-donors), they did not see themselves as a better person as a result of the donation and they thought they did an act somewhat more generously than usual. This is possibly attributed to the cultural influences of normative obligation. The majority (57.9%) believed that they were not prepared well for the donation, and 40% found the experience emotionally less positive than they expected. Some found the experience physically stressful (10.5%) and some others were worried that their future health may be affected as a result of the donation (15.8%). Although a quarter of the donors was unsure whether they would donate again or whether they would encourage others to donate, the majority would donate again. Such results demonstrate that donors need careful and individualised attention in order to gain a more positive overall experience of the donation. Furthermore, services should be more vigilant in assessing donor needs and intervening when donors experience difficulties due to the donation process. PMID- 10516705 TI - Do patients who are treated with stem cell transplantation have a health-related quality of life comparable to the general population after 1 year? AB - Health-related quality of life (HRQOL) in leukemia and lymphoma patients treated with high-dose chemotherapy followed by allogeneic (SCT) and autologous (ASCT) stem cell transplantation or receiving combination chemotherapy (CT) was prospectively assessed by the EORTC QLQ-C30 and compared with reference data from a general population sample. One year after transplant, the SCT group had functional scores which were close to population values except for lower social (P < 0.0001) and role function (P = 0.0004). More symptoms and problems were reported, especially appetite loss (P = 0. 001) and financial difficulties (P = 0.0001). The ASCT patients reported a less than optimal HRQOL relative to the population 1 year post transplant. Cognitive, physical, role, and social function, dyspnoea, financial difficulties and global quality of life were most impaired (P < 0.001). In the CT group, physical, role and social function, dyspnoea and financial difficulties were impaired 1 year after start of chemotherapy, compared with the general population (P < 0.001). The EORTC QLQ-C30 was supplemented by a high-dose chemotherapy module, the HDC-19, at the 1-year assessment, but no consistent differences were found across groups. Fifteen to 34% of the patients expressed fears of relapse and worries about future health, while 24-30% indicated no participation in sexual activities. PMID- 10516706 TI - Thrombotic microangiopathy associated with reactivation of human herpesvirus-6 following high-dose chemotherapy with autologous bone marrow transplantation in young children. AB - Thrombotic microangiopathy (TMA) is a serious complication of BMT. Several factors are important in the etiology of TMA, such as cyclosporin A, GVHD, irradiation, intensive conditioning chemotherapy and infection, which cause damage to vascular endothelial cells leading to activation of these cells. We describe two young children with TMA following high-dose chemotherapy with autologous BMT. Development of TMA was accompanied by reactivation of HHV-6, which was identified by both an increase in the copy number of HHV-6 DNA in the peripheral blood and a significant increase in antibody titers to HHV-6. Thus, it was suggested that reactivation of HHV-6 together with high-dose chemotherapy played an important role in the pathogenesis of TMA in these patients. Since HHV 6 is known to infect vascular endothelial cells, and CMV which is virologically closely related to HHV-6, has been reported to be a pathogen that causes TMA, infection with HHV-6 of vascular endothelial cells may induce TMA via damage and activation of these cells. PMID- 10516707 TI - Graft-versus-myeloma after withdrawal of immunosuppression following allogeneic peripheral stem cell transplantation. AB - There is growing evidence for a graft-versus-myeloma effect following allogeneic stem cell transplantation. We add to this evidence by reporting complete remission achieved by withdrawal of immunosuppression in a patient with multiple myeloma progressing after HLA-identical sibling peripheral stem cell transplantation. De novo chronic graft-versus-host disease coincided with the anti-myeloma effect and responded to treatment. The patient remains in complete remission 3 years after transplant. PMID- 10516708 TI - Autologous peripheral blood stem cell transplantation for Waldenstrom's macroglobulinemia. AB - We report a 50-year-old male patient with Waldenstrom's macroglobulinemia poorly responsive to conventional chemotherapy, who went on to receive high-dose melphalan as conditioning before autologous PBSC infusion. Autologous hematopoiesis was reconstituted 5 weeks after transplant. Complete remission was confirmed by blood and marrow examination. The patient remains well at 12 months, with no detectable monoclonal IgM in his serum and absence of plasmacytoid lymphocytes in his marrow. Our results suggest that, at least in this case, high dose chemotherapy with autologous PBSC support was an effective therapeutic approach for Waldenstrom's macroglobulinemia. PMID- 10516709 TI - Successful bone marrow transplantation in a case of Griscelli disease which presented in accelerated phase with neurological involvement. AB - Griscelli disease (GD) is a rare disorder characterized by pigment dilution, immunodeficiency and occurrence of accelerated phase consisting of hemophagocytosis, pancytopenia and neurological manifestations. Allogeneic BMT in the early period is an important modality of treatment for GD. We carried out an alloBMT from an HLA-identical sibling donor on a 4-year-old girl who presented in accelerated phase with neurological manifestations including convulsions, strabismus, severe dysarthria, ataxia and clonus. She was treated with etoposide, methylprednisolone and intrathecal methotrexate for 8 weeks and underwent alloBMT after receiving a conditioning regimen including ATG (rabbit, 10 mg/kg x 5 days), Bu/Cy. 8 x 108/kg nucleated bone marrow cells were given. Engraftment occurred early and the post-BMT period was uneventful. Currently, she is at 18 months post BMT with sustained engraftment and with a normal neurological examination except for minimal clonus. Long-term follow-up will determine the prognosis regarding the neurological findings. PMID- 10516710 TI - Hematopoietic stem cell transplantation (HSCT) for Langerhans cell histiocytosis (LCH) in Japan. AB - There exists limited information about the usefulness of hemopoietic stem cell transplantation (HSCT) for the treatment of patients with refractory Langerhans cell histiocytosis (LCH). We report here four Japanese pediatric patients with multisystem LCH disease who underwent HSCT between 1994 and 1997. Two of the four patients are doing well without any relapse. However, neither of them shows improved sequelae 3 to 4 years after allogeneic HSCT, although the graft was rejected in one of the cases. The remaining two patients died of septic shock. A review of the literature of 11 patients revealed four fatalities after the use of HSCT in the treatment of LCH. Three of these were due to active LCH and three deaths occurred within 2 months after HSCT. To establish the usefulness of HSCT for refractory LCH, further studies are required. PMID- 10516711 TI - Next-generation adenoviral vectors: new and improved. PMID- 10516712 TI - "Resistance is futile". PMID- 10516714 TI - In vivo methotrexate selection of murine hemopoietic cells transduced with a retroviral vector for Gaucher disease. AB - The studies described were performed to investigate whether in vivo selection of retrovirus-transduced hemopoietic cells is feasible starting from a low percentage of transduced hemopoietic stem cells (PHSCs). The vector used is an amphotropic bicistronic retroviral vector carrying a cDNA for human lysosomal glucocerebrosidase (hGC) for treatment of Gaucher disease and a methotrexate (MTX) resistant mutant cDNA encoding human dihydrofolate reductase (DHFR). We tested the effect of MTX selection in mice that were either myeloablated or not before infusion of transduced cells. In addition, we determined whether repeated administration of transduced bone marrow cells has an additional effect on the percentage of hGC expressing cells. The results obtained have shown that, in myeloablated mice transplanted once with transduced bone marrow and treated twice weekly with 10 mg/kg of MTX for a total of 6 months, a two- to three-fold increased numbers of hGC expressing cells could be detected in both peripheral blood and bone marrow as compared with non-MTX treated mice. In mice transplanted with transduced bone marrow once every 2 weeks for a total of four times, percentages of hGC expressing cells were not significantly increased as compared with mice transplanted once. In non-ablated mice neither MTX selection nor multiple infusions of transduced bone marrow resulted in detection of hGC expressing cells 6 months after transplantation, indicating that the success of in vivo selection using MTX is highly dependent on the ratio of transduced hemopoietic stem cells transplanted versus residing and untransduced stem cells. PMID- 10516713 TI - Promoter-activated expression of nerve growth factor for treatment of neurodegenerative diseases. AB - Genetic transfer approaches have received recent consideration as potential treatment modalities for human central and peripheral nervous system (CNS and PNS, respectively) neurodegenerative disorders, including Parkinson's disease, Alzheimer's disease, and amyotrophic lateral sclerosis. Transplantation of genetically modified cells into the brain represents a promising strategy for the delivery and expression of specific neurotrophic factors, neurotransmitter synthesizing enzymes, and cellular regulatory proteins for intervention in neurodegenerative diseases. The use of specific regulatable promoters may also provide potential control of gene expression required for dose-specific or time specific therapeutic strategies. In this article, we review the potential use of activated promoters in ex vivo systems for the potential genetic therapy of neurodegenerative disorders, and then describe our own studies using the zinc inducible metallothionein promoter for the regulated expression of nerve growth factor (NGF) in rodent brain transplants. PMID- 10516715 TI - Retrovirus vectors designed for efficient transduction of cytotoxic or cytostatic genes. AB - It is difficult to establish stable packaging cell lines producing retrovirus vectors for the expression of anti-oncogenes with cytotoxic or cytostatic potential, because these genes would also affect the growth of the packaging cell lines. To overcome this problem, we designed a transcriptional unit pBabeLPL for vector RNA production, in which the transcription of the exogenous genes is completely suppressed by the presence of a preceding insertion containing the puromycin resistance gene (puro) and a poly(A) addition signal. This insertion is flanked by a tandem pair of loxP, and is designed to be excised after the introduction of Cre recombinase, when transcription of the exogenous gene will be started from the 5'-LTR. The transcriptional unit car- rying LacZ or p53 as the exogenous gene was introduced into a previously constructed prepackaging cell lines PtG-S2, in which the expression of VSV-G is also designed to be initiated by the introduction of Cre recombinase, while the gag-pol gene is expressed continuously. After the introduction of Cre recombinase by an adenovirus vector, LacZ- or p53-expressing VSV-G-pseudotyped retrovirus vectors with the designed structure were produced at high virus titers. The p53 virus was shown to be able to transduce p53 into the entire population of several human cancer cell lines and to induce their growth arrest at the G1 phase, indicating that this vector producing system will be advantageous for human gene therapy. PMID- 10516716 TI - Delivery of herpes simplex virus amplicon-based vectors to the dentate gyrus does not alter hippocampal synaptic transmission in vivo. AB - Herpes simplex virus type-1 (HSV) amplicon vectors containing neuroprotective genes can alter cell physiology and enhance survival following various insults. However, to date, little is known about effects of viral infection itself (independent of the gene delivered) on neuronal physiology. Electrically-evoked synaptic responses are routinely recorded to measure functional alterations in the nervous system and were used here to assess the potential capability of HSV vectors to disrupt physiology of the hippocampus (a forebrain structure involved in learning that is highly susceptible to necrotic insult, making it a frequent target in gene therapy research). Population excitatory post-synaptic potentials (EPSPs) were recorded in the dentate gyrus (DG) and in area CA3 in vivo 72 h after infusion of an HSV vector expressing a reporter gene (lacZ) or vehicle into the DG. Evoked perforant path (PP-DG) or mossy fiber (MF-CA3) EPSPs slope values measured across input/output (I/O) curves were not altered by infection. Paired pulse facilitation at either recording site was also unaffected. X-gal-positive granule cells surrounded the recording electrode (PP-DG recording) and stimulating electrode tracts (MF-CA3 recording) in animals that received vector, suggesting that we had measured function, at least in part, in infected neurons. Because of the negative electrophysiological result, we sought to deliver a gene with an HSV amplicon which would affect the measured endpoints, as a positive control. Delivery of calbindin D28kpotentiated PP-DG synaptic strength, indicating that our recording system could detect alterations due to vector expression. Thus, the data indicate that HSV vectors are benign, in regard to effects on synaptic function, and support the use of these vectors as a safe method to deliver selected genes to the central nervous system. PMID- 10516717 TI - Intrauterine gene transfer: gestational stage-specific gene delivery in mice. AB - Intrauterine gene transfer in mice by intraplacental microinjection of recombinant adenoviral or retroviral vectors carrying beta-galactosidase (beta gal) reporter gene was analyzed in relation to gestational stage, viral titer and promoters. After injections of viral vectors on days 9.5, 11.5 or 14.5 post coitum (p.c.), embryos, fetuses and adult animals were analyzed for beta-gal expression on days 13.5 p.c., 18.5 p.c. and at 2 months of age, respectively. Injection of adenoviral vectors on day 9.5 or day 11.5 p.c. resulted in high beta gal expression in the heart or liver, respectively. Injection on either day also gave expression in other tissues including vasculature and hindbrain. This temporal pattern of adenoviral transduction correlated with the expression level of integrin beta3 subunit, which is known to be a component involved in adenoviral transduction. Adenovirus-mediated intrauterine gene transfer resulted in persistent beta-gal expression in multiple cell foci in the liver and hearts of 2-month-old adult animals treated in utero, indicating stable integration of the transgene into the host cell genome at a low frequency. Although at low efficiency, injection of retroviral vector on day 9.5 and 11.5 p.c. resulted in beta-gal expression in embryonic liver, while day 9.5 p.c. injection resulted in persistent beta-gal expression in 2-month-old adult heart. This is the first study to show gestational stage-specific gene transfer via intraplacental microinjection of adenoviral vectors. PMID- 10516718 TI - Persistent expression of canine factor IX in hemophilia B canines. AB - We previously demonstrated that direct intramuscular injection of recombinant adeno-associated virus (rAAV) carrying the human FIX (hFIX) cDNA can safely be administered to hemophilic B canines and express human factor IX protein; however, the functional activity of the hFIX protein could not be assessed due to anti-human FIX antibody (inhibitor) formation. To test the therapeutic efficacy of rAAV in hemophilic dogs, rAAV type 2 (rAAV2) carrying canine FIX (cFIX) cDNA was injected into the skeletal muscle of two dogs at doses of 1012-13particles. Circulating cFIX protein levels were maintained for 1 year at levels of 1-2% of normal. Hemostatic correction (WBCT and APTT) paralleled plasma FIX antigen levels. Both dogs still required plasma infusion for spontaneous and traumatic bleeding events. Inhibitors to cFIX protein were not detected in either animal by Bethesda assay. Neutralizing antibodies directed against AAV-2 capsid were pronounced and persistent. Vector DNA and mRNA transcripts were detected only at the injected skeletal muscle tissue. Analysis of both high and low molecular weight DNA identified both replicative episomal and integrated AAV species. These results demonstrate that persistent secretion of the FIX transgene protein, necessary for successful gene therapy of hemophilia B, can be achieved using the parvovirus-based rAAV vector PMID- 10516719 TI - IL-10 expression by CT26 colon carcinoma cells inhibits their malignant phenotype and induces a T cell-mediated tumor rejection in the context of a systemic Th2 response. AB - In spite of the evidence that IL-10 has Th1-immunosuppressive and anti inflammatory effects, it has been shown that IL-10 may reduce the tumorigenic capacity of certain tumor cell types. In order to characterize the responses elicited by IL-10, we explored the effect of transducing murine CT26 colon carcinoma cells with a recombinant retrovirus expressing mIL-10. IL-10 gene transfer of CT26 cells had no effect on tumor cell growth on plastic surface but inhibited the anchorage-independent growth capacity of tumor cells and their metastatic potential as assessed by their invasive and migration ability. Expression of IL-10 also elicited an antitumor immune response involving both CD4+ and CD8+ T cells. Assessment of the immune status of the animals demonstrated that mice injected with CT26-IL10 cells showed prevalence of a systemic and tumor-specific Th2 response. Spleen cells obtained from these mice showed an increased production of IL-4 and no changes in IFNgamma levels, characteristic of a Th2 response. These results demonstrate that IL-10 affects CT26 tumor cell growth by both inhibiting the malignant phenotype and by recruiting and activating a T cell-mediated antitumor response. This T cell response occurs in the context of a shift towards a Th2 response. PMID- 10516720 TI - Intra-articular delivery of a herpes simplex virus IL-1Ra gene vector reduces inflammation in a rabbit model of arthritis. AB - To evaluate the use of HSV-based vectors for arthritis gene therapy we have constructed a first-generation, ICP4 deficient, replication defective herpes simplex virus (HSV) vector (S/0-) and a second-generation HSV vector derivative (T/0-) deficient for the immediate-early genes ICP4, 22 and 27, each carrying a soluble TNF receptor or IL-1 receptor antagonist transgene cassette. A rabbit synovial-fibroblast line in culture, infected by either vector enabled high-level expression of the transgene product. However, following a single intra-articular injection of the vectors into rabbit knee joints, only the second-generation, HSV T/0- vector expressed detectable levels of soluble TNFR in synovial fluid. Synovial lavage fluid from inoculated joints con- tained up to 12 ng/ml of soluble receptor that persisted at detectable, but reduced levels for at least 7 days. When tested in an experimental model of arthritis generated by intra articular overexpression of interleukin-1beta using retrovirus transduced synovial cells, the HSV T/0- vector expressing the interleukin-1 receptor antagonist was found to inhibit leukocytosis and synovitis significantly. The improved levels and duration of intra-articular transgene expression achieved via HSV-mediated gene delivery suggest that an HSV vector system could be used for therapeutic applications in patients with rheumatoid arthritis (RA) and other joint-related inflammatory diseases. PMID- 10516721 TI - An adenoviral vector regulated by hypoxia for the treatment of ischaemic disease and cancer. AB - Recombinant adenoviral vectors have a number of advantages for gene therapy, including transduction of a range of dividing and non-dividing cell types. However, this broad range may be a disadvantage if non-target cells are transduced and are adversely affected by expression of the transferred gene. Here we describe a novel adenoviral vector in which transcription of the transgene is restricted to the patho-physiological condition of low oxygen tension (hypoxia). Hypoxia activates the expression of a number of genes, principally via the stabilisation of members of the bHLH/PAS family of transcription factors that bind to a con- sensus DNA sequence, the hypoxia response element (HRE). We have configured an optimised HRE expression cassette into an adenoviral vector, AdOBHRE. A range of cell types, including primary human skeletal muscle, when transduced with AdOBHRE display a low basal level of transgene expression that is highly induced in hypoxia to levels equivalent to that obtained from the CMV promoter. The AdOBHRE vector could be exploited for transcriptionally targeted gene therapy for the treatment of diseases such as cancer, peripheral arterial disease, arthritis and anaemia where tissue hypoxia is a cardinal feature. PMID- 10516722 TI - Induction of P815 tumor immunity by recombinant Semliki Forest virus expressing the P1A gene. AB - The methylcholantrene-induced P815 mastocytoma tumor is derived from DBA/2 mice and expresses a weak tumor rejection antigen, P815A. The P1A gene, which encodes for the P815A antigen, is silent in most normal tissues with the exception of testis and placenta. These characteristics make P815 an interesting mouse model for the human MAGE-type tumor antigens. Recombinant Semliki Forest virus particles (rSFV) were constructed that expressed variants of the P815 antigen. Such particles, when used for vaccination, express the antigen only transiently since the viral vector is incapable of productive replication. Nevertheless, mice vaccinated with rSFV generated strong CTL responses and were protected against P815 tumor challenge. PMID- 10516723 TI - Nonviral transfer of genes to pig primary keratinocytes. Induction of angiogenesis by composite grafts of modified keratinocytes overexpressing VEGF driven by a keratin promoter. AB - Cultured epithelial grafts have proven to be life-saving in the treatment of large skin losses. It has become apparent that one of the main difficulties of this technology is the overall poor take of the grafts as a consequence of severely damaged dermal beds. Skin substitutes providing both cultured keratinocytes, as an epidermal layer, and a dermal analogous offer a more suitable material for skin repair. Ex vivo transfer of stroma regeneration promoting genes to keratinocytes appears to be an attractive strategy for improving the therapeutic action of these grafts. The use of epidermal-specific promoters as expression drivers of exogenous genes results in both high expression levels and stratum specificity, as shown in transgenic mice studies. Most current gene transfer protocols to primary keratinocytes involve transduction of epidermal cells with retroviral vectors. However, transfer of gene constructs harboring these long DNA fragment promoters cannot be achieved through viral transduction. In this paper, we describe a protocol consisting of lipid-mediated transfection, G418 selection and an enhanced green fluorescence protein (EGFP)-based enrichment step for obtaining high levels of transgene expressing primary keratinocytes. Using this protocol, the cDNA for vascular endothelial growth factor (VEGF), a potent endothelial cell mitogen driven by the 5.2 kb bovine keratin K5 promoter, was stably transfected into pig primary keratinocytes. Genetically modified keratinocytes, expanded on live fibroblast containing fibrin gels and transplanted to nude mice as a composite material, elicited a strong angiogenic response in the host stroma as determined by fresh tissue examination and CD31 immunostaining. Since the formation of a well vascularized wound bed is a crucial step for permanent wound closure, the use of an 'angiogenic' composite material may improve wound bed preparation and coverage with cultured keratinocyte grafts. PMID- 10516724 TI - A photosensitising adenovirus for photodynamic therapy. AB - We have developed a new approach to photodynamic therapy based on adenoviral transduction of the rate-limiting enzyme in heme synthesis. Conventional phototherapy uses porphyrin-based chemical photosensitisers, including delta aminolaevulinic acid (ALA) which is converted to protoporphyrin IX (PpIX) by the enzymes of the heme biosynthetic pathway. The lack of a specific mechanism for targeting chemical photosensitisers and PpIX to tumour cells means that therapeutic irradiation can damage normal tissue and exposure to sunlight following treatment can cause severe burns. The rate limiting enzyme in PpIX synthesis is ALA-synthase (ALA-S). We have developed a new yeast vector system for manipulation of the adeno- virus genome and used it to construct a virus expressing a mutant form of ALA-S lacking the iron response elements which regulate ALA-S translation and the heme regulatory motifs which regulate import of ALA-S into mitochondria. The virus induces a large increase in PpIX expression and confers photosensitivity on cultured cells. Unlike conventional photodynamic therapy, a viral approach makes it possible to restrict photosensitivity by biological rather than purely physical or chemical means. As with HSV thymidine kinase, ALA-S expression is a general mechanism for sensitisation to a therapeutic agent which can easily be adapted to whatever means of gene delivery is most effective. PMID- 10516726 TI - Transduction of axotomized retinal ganglion cells by adenoviral vector administration at the optic nerve stump: an in vivo model system for the inhibition of neuronal apoptotic cell death. AB - Axotomy of the rat optic nerve leads to apoptotic cell death of retinal ganglion cells (RGCs). We have used adenoviral vectors to transduce RGCs from the cut optic nerve stump, a paradigm in which only those neurons are transduced which are directly affected by the axonal lesion. Transgenes encoded by the vectors were p35 and CrmA, which are potent intracellular anti-apoptotic proteins. We found that p35, but not CrmA exerted significant rescue effects on RGCs 14 days after axotomy. Expression of the transgenes was driven by the murine CMV (MCMV) promoter. The respective mRNAs were detectable 7 days but not 14 days after transduction. Since surviving RGCs were present beyond the time-point of detectable transcription of the p35 transgene, we conclude that apoptosis has been efficiently inhibited. In addition, we observed that transduction with two control vectors without a transgene in E1 also resulted in a minor but significant RGC rescue, implicating neuroprotective effects due to adenoviral transduction itself. This system will be useful in dissecting the pathways leading to neuronal cell death after axonal lesions and in the evaluation of the important question whether the cellular suicide program can be reverted to survival by therapeutic gene delivery. PMID- 10516725 TI - Effect of prior exposure to herpes simplex virus 1 on viral vector-mediated tumor therapy in immunocompetent mice. AB - Replication-competent, attenuated mutants of herpes simplex virus type 1 (HSV-1) have been shown to be efficacious for tumor therapy. However, these studies did not address the consequences of prior exposure to HSV, as will be the case with many patients likely to receive this therapy. Two strains of mice, A/J and BALB/c, were infected with wild-type HSV-1 by intraperitoneal injection and the immune response was determined by plaque reduction assay for neutralizing antibody and ELISA for IgG and IgM. Syngeneic tumors, N18 neuroblastoma and CT26 colon carcinoma, were implanted subcutaneously in HSV-1 seropositive and naive A/J and BALB/c mice, respectively. Established tumors were subsequently treated intratumorally with a multi-mutated HSV-1, G207. G207 inhibited tumor growth to a similar extent whether the mice were seropositive or not. We next examined the effect of multiple intratumoral inoculations of a 10-fold lower dose of G207 on tumor growth. In the multiple treatment group (biweekly for 3 weeks), 75% of tumors were cured, whereas no cures were seen in the single treatment group. We conclude that HSV seropositivity should not deleteriously affect the efficacy of G207 tumor therapy, and multiple inoculations of virus should be considered for clinical evaluation. PMID- 10516727 TI - Protective immunization against melanoma by gp100 DNA-HVJ-liposome vaccine. AB - DNA-based vaccine immunization effectively induces both humoral and cell-mediated immunity to antigens and can confer protection against numerous infectious diseases as well as some cancers. Human and mouse melanomas consistently express the tumor-associated antigen interacted with the melanogenesis pathway. Gp100 is immunogenic and has been shown to induce both antibody and cytotoxic T cell (CTL) responses in humans. To explore the potential use of DNA immunization to induce melanoma-specific immune responses, we assessed HVJ-AVE liposome incorporated with plasmid DNA encoding human gp100. The gp100 DNA vaccine was used in a mouse melanoma model to assess immunity against the B16 melanoma of C57BL/6 mice. Intramuscular injection of the DNA-HVJ-AVE liposomes induced both anti-gp100 antibody and CTL responses. Gp100 DNA-HVJ-AVE liposome immunization significantly delayed tumor development in mice challenged with B16 melanoma cells. Mice immunized with gp100 DNA-HVJ-AVE liposomes survived longer compared with control mice immunized with HVJ-AVE liposome alone. These results indicate that immunization with human gp100 DNA by HVJ-AVE liposomes can induce protective immunity against melanoma in this pre-clinical mouse model. This strategy may provide an effective approach for vaccine therapy with tumor-associated antigens against human melanoma. PMID- 10516728 TI - Gene transfer and expression of a non-viral polycation-based vector in CD4+ cells. AB - CD4-selective targeting of an antibody-polycation-DNA complex was investigated. The complex was synthesized with the anti-CD4 monoclonal antibody B-F5, polylysine268 (pLL) and either the pGL3 control vector containing the luciferase reporter gene or the pGeneGrip vector containing the green fluorescent protein (GFP) gene. B-F5-pLL-DNA complexes inhibited the binding of 125I-B-F5 to CD4+Jurkat cells, while complexes synthesised either without B-F5 or using a non specific mouse IgG1 antibody had little or no effect. Expression of the luciferase reporter gene was achieved in Jurkat cells using the B-F5-pLL-pGL3 complex and was enhanced in the presence of PMA. Negligible luciferase activity was detected with the non-specific antibody complex in Jurkat cells or with the B F5-pLL-pGL3 complex in the CD4- K-562 cells. Using complexes synthesised with the pGeneGrip vector, the transfection efficiency in Jurkat and K-562 cells was examined using confocal microscopy. More than 95% of Jurkat cells expressed GFP and the level of this expression was markedly enhanced by PMA. Negligible GFP expression was seen in K-562 cells or when B-F5 was replaced by a non-specific antibody. Using flow cytometry, fluorescein-labelled complex showed specific targeting to CD4+ cells in a mixed cell population from human peripheral blood. These studies demonstrate the selective transfection of CD4+ T-lymphoid cells using a polycation-based gene delivery system. The complex may provide a means of delivering anti-HIV gene therapies to CD4+ cells in vivo. PMID- 10516729 TI - Intratumoral injection of bone-marrow derived dendritic cells engineered to produce interleukin-12 induces complete regression of established murine transplantable colon adenocarcinomas. AB - Stimulation of the antitumor immune response by dendritic cells (DC) is critically dependent on their tightly regulated ability to produce interleukin-12 (IL-12). To enhance this effect artificially, bone marrow (BM)-derived DC were genetically engineered to produce high levels of functional IL-12 by ex vivo infection with a recombinant defective adenovirus (AdCMVIL-12). DC-expressing IL 12 injected into the malignant tissue eradicated 50-100% well established malignant nodules derived from the injection of two murine colon adenocarcinoma cell lines. Successful therapy was dependent on IL-12 transfection and was mediated only by syngeneic, but not allogeneic BM-derived DC, indicating that compatible antigen-presenting molecules were required. The antitumor effect was inhibited by in vivo depletion of CD8+ T cells and completely abrogated by simultaneous depletion with anti-CD4 and anti-CD8 mAbs. Mice which had undergone tumor regression remained immune to a rechallenge with tumor cells, showing the achievement of long-lasting systemic immunity that also was able to reject simultaneously induced concomitant untreated tumors. Tumor regression was associated with a detectable CTL response directed against tumor-specific antigens probably captured by DC artificially released inside tumor nodules. Our results open the possibility of similarly treating the corresponding human malignancies. PMID- 10516730 TI - In vivo gene introduction into keratinocytes using jet injection. AB - Successful keratinocyte gene therapy requires the development of efficient methods of gene transfer to keratinocytes. Jet injection of a solution containing DNA can be used to transfer genes to several tissues in vivo. In this article, we tried to introduce DNA into rat and human keratinocytes using this method. First, we fired a beta-gal expression vector into rat skin at several distances using a jet injector and examined beta-gal activity in the epidermal keratinocytes. The highest activity in keratinocytes was found when the plasmid was fired at 10 cm from the skin surface; the activity lessened as the firing distance became shorter than 10 cm. Next, we transplanted human skin on to a nude rat, fired the vector into the human skin from a distance of 10 cm and examined the beta-gal activity. We also injected the same amount of plasmid with a needle to compare jet with needle injections. The results showed that jet injection of the naked DNA could introduce and express DNA in human keratinocytes in vivo and that jet injection exhibited much higher activity than needle injection. Jet injection of the naked DNA will provide a method for keratinocyte gene therapy in the future. PMID- 10516731 TI - Establishment of an optimised gene transfer protocol for human primary T lymphocytes according to clinical requirements. AB - Current gene therapeutic protocols directed towards the treatment of inherited disorders (eg ADA-SCID) and viral infections (eg AIDS), as well as adoptive immunotherapy approaches are based on the use of genetically modified lymphocytes. Since only insufficient transduction of T cells is obtained using existing techniques, the development of more efficient gene transfer protocols into these cells is of great importance. We present here a protocol for the highly efficient transduction of human primary T cells at high densities (1 x 106/ml) by retroviral infection. Using retroviral vectors encoding a truncated human low-affinity nerve growth factor receptor (DeltaLNGFR) as a gene transfer marker, we obtained transduction frequencies of more than 70% of CD3+ cells after two cycles of infection. Our protocol is based on the use of FBS-free media for both the production of retrovirus-containing supernatant and the cultivation of the primary T cells. Since the protocol presented here works just as efficiently under large-scale conditions, it may be easily adapted to clinical needs and 'good manufacturing practice' (GMP) standards. PMID- 10516732 TI - . . . and losartan was no better than placebo. PMID- 10516733 TI - Toxicity of mercury. AB - A ruling by the European Union heralds the demise of those useful clinical instruments, the mercury thermometer and the mercury sphygmomanometer. The new laws have been passed because of worries about mercury poisoning. Yet you can drink metallic mercury and come to no harm. What does it all mean? There are three forms of mercury from a toxicological point of view: inorganic mercury salts; organic mercury compounds; and metallic mercury. Inorganic mercury salts are water soluble, irritate the gut, and cause severe kidney damage. Organic mercury compounds, which are fat soluble, can cross the blood brain barrier and cause neurological damage. Mercury metal poses two dangers. It can be vaporised: the vapour pressure at room temperature is about 100 times the safe amount, so poisoning can occur if mercury metal is spilled into crevices or cracks in the floorboards. Dentists are occasionally poisoned this way. Mercury easily crosses into the brain, and causes tremor, depression, and behavioural disturbances. A second danger from metallic mercury is that it is biotransformed into organic mercury, by bacteria at the bottom of lakes. This can be passed along the food chain and eventually to man. It was this process that led to the Japanese tragedy at Minimata Bay in the late 1950s when over 800 people were poisoned. It is the need to reduce mercury contamination of the environment which should encourage us to cut the usage of metallic mercury. However, much more metallic mercury is spilled as waste by the chemical industry than is dropped on the floor in the clinic. PMID- 10516734 TI - ABPM comparison of the antihypertensive profiles of the selective angiotensin II receptor antagonists telmisartan and losartan in patients with mild-to-moderate hypertension. AB - The antihypertensive efficacy and tolerability profiles of the selective AT1 receptor antagonists telmisartan and losartan were compared with placebo in a 6 week, multinational, multicentre, randomised, double-blind, double-dummy, parallel-group study of 223 patients with mild-to-moderate hypertension, defined as clinic diastolic blood pressure (DBP) >/=95 and /=140 and /=85 mm Hg. After a 4-week single-blind placebo run-in, eligible patients were randomised to receive telmisartan 40 mg, telmisartan 80 mg, losartan 50 mg, or placebo. Ambulatory blood pressure monitoring (ABPM) after 6 weeks of double-blind therapy showed that all active treatments produced significant (P < 0.01) reductions from baseline in 24-h mean SBP and DBP compared with placebo. During the 18-to-24 h period after dosing, the reductions in SBP/DBP with telmisartan 40 mg (10.7/6.8 mm Hg) and 80 mg (12.2/7. 1 mm Hg) were each significantly (P <0.05) greater than those observed for losartan 50 mg (6.0/3.7 mm Hg), and losartan was no better than placebo. Also for the 24-h mean blood pressure, telmisartan 40 mg and 80 mg were significantly (P< 0.05) better than losartan 50 mg. Compared with losartan, telmisartan 80 mg produced significantly (P < 0.05) greater reductions in both SBP and DBP during all monitored periods of the 24-h period, while telmisartan 40 mg produced significantly greater reductions in SBP and DBP in the night-time period (10.01 pm to 5.59 am) (P < 0.05) and in DBP in the morning period (6.00 am to 11.59 am) (P < 0.05). All treatments were comparably well tolerated. Telmisartan 40 mg and 80 mg once daily were effective and well tolerated in the treatment of mild-to-moderate hypertension, producing sustained 24-h blood pressure control which compared favourably with losartan. PMID- 10516735 TI - QTc dispersion and complex ventricular arrhythmias in untreated newly presenting hypertensive patients. AB - Increased dispersion of ventricular repolarisation (increased QT dispersion) is believed to predispose to arrhythmias associated with sudden death in certain cardiac diseases. Hypertension is also associated with increased risk of sudden death, particularly in those with left ventricular hypertrophy (LVH). Therefore, the first aim of this study is to look into the possible pathogenic role of QT dispersion on the ventricular arrhythmias occurring in a group of never-treated hypertensive patients. The second aim is to look at other possible determinants of QT dispersion (ie, level of blood pressure, hypokalaemia, electrocardiographic LVH and presence or absence of strain pattern) in hypertensive patients, and their relevance to complex ventricular arrhythmias. QTc (corrected QT) was measured in 70 newly presenting (never-treated) hypertensive patients (47 male, 23 female, mean age 51.9 +/- 12.5 years) from a standard 12-lead surface electrocardiogram (ECG). Blood pressure measurements and 24-h ECG holter recordings were performed in all patients. Serum potassium level was measured in 51 of the patients. Ventricular arrhythmias were classified using a modified Lown's scoring system. Maximum QTc, minimum QTc and QTc dispersion for all patients were 442 +/- 30.3 ms, 380 +/- 26.7 ms and 61.5 +/- 21.6 ms respectively. High grade ventricular arrhythmias (Lown's score >/=3) were found in 43% of the patients. The QTc dispersion was strongly correlated with the Lown's classification of arrhythmia and the age of the patients. Patients with more severe ectopy (Lown's score >/=3) were significantly older (57.4 +/- 10.3 years) compared to those with score /=3 Lown's score compared to 39% in the group with LVH but without strain. In the presence of relative hypokalaemia, hypertensive patients with LVH showed more QTc dispersion (85.7 +/- 15.5 ms) and a greater tendency for complex ventricular arrhythmias (100% grade >/=3 Lown's score) compared to those with LVH and normal serum potassium levels (64.1 +/- 22.6 ms and 35%, QTc dispersion and Lown's score >/=3, respectively P = 0. 05). The level of blood pressure had no effect on either the QTc dispersion or the prevalence of complex ventricular arrhythmias. Prevalence of complex ventricular arrhythmias in hypertensive patients is strongly correlated with QTc dispersion and age. When hypertensive patients with LVH have low potassium levels the risk of developing complex ventricular arrhythmias is significantly increased. PMID- 10516736 TI - Hypertension in urban Mexico: the 1992-93 national survey of chronic diseases. AB - The purpose of this work is to estimate the prevalence of hypertension in the urban population of Mexico. We studied a multistage national sample representative of the urban population in 417 cities of over 2500 people. The blood pressure of 14 657 individuals (6053 men and 8604 women) aged 20-69 years was measured after a 5-min rest using a standard mercury sphygmomanometer. The survey personnel had been previously trained and standardised. The main results show a crude prevalence of hypertension, as defined by the JNC VI, of 28.1% in women and 37.5% in men (27.2% and 37.1% age-adjusted). Both genders exhibited a trend of increasing hypertension with age. In individuals under 50 years of age, women had lower rates than men, but the difference disappeared in the older groups. The awareness of hypertension (28%) as well as the success of treatment (22%) were low in our sample. Our results had more similarities than differences with respect to those observed in other national surveys. It is concluded that hypertension in Mexico is an important public health problem similar to that seen in developing and developed nations. Efforts should be aimed at strengthening measures to prevent and control hypertension in Mexico. More information is needed of the sort obtained from longitudinal studies. PMID- 10516737 TI - Relation of ambulatory blood pressure load with left ventricular geometry in untreated patients with mild-to-moderate hypertension. AB - Whether ambulatory blood pressure (ABP) load is associated with left ventricular (LV) geometry was assessed in 335 patients (range 32-72 years) with stage I-II essential hypertension by performing 24-h ABP monitoring and echocardiographic examination. Of these 335 hypertensive subjects, 116 (34.5%) had normal LV geometry, 136 (40.5%) had concentric LV remodelling, 37 (11%) had eccentric LV hypertrophy and 46 (14%) had concentric LV hypertrophy according to the relative wall thickness and left ventricular mass index. Subjects with concentric LV hypertrophy had significantly increased 24-h systolic BP (SBP), diastolic BP (DBP) and mean arterial pressure as well as increased 24-h SBP and DBP load compared to those with normal LV geometry or concentric LV remodelling while there was no difference in the above parameters in comparison with the subjects with eccentric LV hypertrophy. The incidence of patients with normal LV geometry was significantly decreasing and the incidence of patients with LV-CH was significantly increasing as the degree of ABP loads were increasing. Using multiple regression analysis models with each type of LV geometry as a dependent variable and various degree of ABP loads as independent variables, it was revealed that normal LV geometry was significantly related with normal values of 24-h SBP and DBP load (P < 0.05) while there was not any significant relation between concentric LV remodelling and 24-h SBP or DBP load values. Concentric LV hypertrophy was significantly related with increased values of both 24-h SBP and DBP load (P < 0.05) while eccentric LV hypertrophy was significantly related with increased values of 24-h DBP load only (P < 0.05). In conclusion normal LV geometry is associated with normal values of SBP and DBP load while concentric LV hypertrophy is associated with increased values of both SBP and DBP load. PMID- 10516739 TI - Blood pressure and social support observations from Mamre, South Africa, during social and political transition. AB - OBJECTIVE: Social support, by moderating cardiovascular reactivity, has been demonstrated to attenuate the effects of stress on blood pressure in American communities. This is the first report to examine the relationship between social support and blood pressure in a South African context, during a period of infrastructure modernisation and political change. METHODS: A total of 1240 residents (542 men, 698 women) of mixed ethnic origin, older than 14 years and stratified by age and sex, participated in a survey to determine risk factors for hypertension and cardiovascular diseases. Social support was assessed by a questionnaire developed in consultation with the community. It was defined by interactions that may threaten family harmony (score 1) and by networking between relatives, friends, colleagues and neighbours (score 2). RESULTS: Mean blood pressure of the sample was 130/79 mm Hg (s.d. 25/14 mm Hg). Hypertension prevalence was 26.9%. Only 36% of women compared to 57.3% of men (P < 0.0001) were employed. More women (29%) than men (22%) reported threats to family harmony, but social support networks were similarly perceived by both sexes. Systolic and diastolic blood pressure correlated weakly with score 1 (r = 0.096, P < 0.0007) but no association was observed with score 2. Score 1 was not associated with blood pressure by multiple regression analysis, that included confounding by age, sex, BMI, alcohol consumption and smoking status. CONCLUSIONS: Neither threats to family harmony nor networking between relatives, friends or neighbours, significantly influences blood pressure in this community. Measures of social support thought to moderate blood pressure may have limited cross-cultural application. Attitudinal changes during socio-political transition may impact on the generalisability of instruments for measurement. PMID- 10516738 TI - The long-term antihypertensive activity and tolerability of irbesartan with hydrochlorothiazide. AB - The long-term safety, tolerability, and antihypertensive effects of irbesartan/hydrochlorothiazide (HCTZ) were assessed in hypertensive patients (seated diastolic blood pressure [SeDBP] 95-110 mm Hg). Patients (n = 1098) completing two randomised, double-blind trials of irbesartan alone, HCTZ alone, irbesartan/HCTZ combinations, or placebo, took 1 year of open-label therapy starting with irbesartan 75 mg/HCTZ 12.5 mg once daily. If target blood pressure (BP) (<140/<90 mm Hg) was not achieved, the dose was titrated sequentially at 2- to 4-week intervals to irbesartan 150 mg/HCTZ 12. 5 mg, then to irbesartan 300 mg/HCTZ 25 mg. If necessary, adjunctive therapies were added. Mean changes in trough seated systolic BP/SeDBP at months 2, 6, and 12 were -19.1/-14.2 mm Hg (n = 941), -20.7/ -15.7 mm Hg (n = 948), and -20.6/-15.6 mm Hg (n = 898), respectively. From months 2 to 12, normalisation rates (trough SeDBP <90 mm Hg) ranged from 75-85% and total responder rates (normalised or >/=10 mm Hg trough SeDBP reduction) ranged from 81-91%, while target BP was achieved in 65-75% of patients. At all time-points, most patients (>/=87%) were receiving irbesartan/HCTZ alone. Eighty-two patients (7.5%) discontinued the study due to adverse events, with half of these events considered unrelated to study medication. There were no reports of serious adverse events related to study medication. Long-term therapy with irbesartan/HCTZ is safe, well tolerated, and maintains normalised BP in >80% of patients. PMID- 10516740 TI - Differences in blood pressure values by simultaneous auscultation of Korotkoff sounds inside the cuff and in the antecubital fossa. AB - To elucidate whether auscultation of the Korotkoff sounds inside the cuff and in the antecubital fossa leads to different blood pressure (BP) values we measured BP at both sites simultaneously with identical flat stethoscopes in a same-arm test design (part A) in 64 normotensive (N: 32 men, 32 women; mean age: 38.7 +/- 15.1 years) and 67 hypertensive subjects (H: 36 men, 31 women; mean age: 44.6 +/- 12.9 years), and additionally in a same- and opposite-arm test design (part B) in 20 normotensive young women. While in part A systolic BP measured inside the cuff was only slightly higher (N: +1. 6 +/- 3.2 mm Hg; H: +1.0 +/- 1.4 mm Hg), diastolic BP was considerably lower (N: -10.6 +/- 5.6 mm Hg; H: -8.4 +/- 4.9 mm Hg). This result was corroborated by part B with differences in systolic/diastolic BP of +0.8 +/- 1.0/-8.5 +/- 2.2 mm Hg in the same-arm test and +0.4 +/- 4.8/-10.6 +/- 5.2 mm Hg in the opposite-arm test. Subject's age was the main variable determining differences in diastolic BP with significantly higher differences in younger than in older subjects, indicating that the elastic properties of arteries may be responsible for these differences. Our results demonstrate that a modification in the auscultatory technique of BP measurement produces significantly different diastolic BP values, the magnitude of which is important for our conceptions of threshold and target values in diagnosing and treating hypertension. PMID- 10516741 TI - Altered sodium perception in essential hypertensive patients following rapid volume expansion. AB - We investigated sodium and volume-dependent mechanisms in the modulation of adrenal and renal vascular responsiveness to angiotensin II in hypertensive (n = 9) and normal subjects (n = 5) who demonstrated normal responses during steady state salt balance (intact modulation). Adrenal and renal vascular responses to angiotensin II were assessed on four occasions. These studies were performed during steady-state high salt and low salt balance and later during non-steady state conditions, after acute volume expansion with normal saline or dextran infusions. The volume expansion studies were administered while study subjects were in low salt balance. In both the normal and hypertensive patients saline and dextran suppressed plasma renin activity and aldosterone release, induced renal vasodilation and enhanced the renal vascular response to angiotensin II, similar to that observed during high salt balance. Saline also reduced the adrenal response to angiotensin II in normal subjects but the adrenal response to angiotensin II in hypertensives remained enhanced. The slopes of regression lines relating angiotensin II and aldosterone following saline infusion were significantly different (P < 0.05) between normal subjects (21 +/- 6) and hypertensives (110 +/- 33). Volume expansion with dextran did not effect the normal or hypertensive adrenal response to angiotensin II. We have demonstrated that the renal vascular response to angiotensin II is rapidly modulated by volume expansion per se in normal subjects and hypertensive patients with intact steady state renin-angiotensin-aldosterone system responses. However, hypertensives have a delay in resetting the adrenal responsiveness to angiotensin II suggesting that they have altered sodium perception during rapid changes in salt balance. PMID- 10516742 TI - Low dose combinations in the treatment of hypertension: theory and practice. AB - The recent evolution of drug therapy for hypertension has primarily focused on new agents but there has been a renewed interest in the use of low doses of diuretic in combination with other agents. Such low-dose diuretic combinations are rational and their use will almost certainly increase. PMID- 10516743 TI - Early abnormalities in left ventricular diastolic function of sodium-sensitive hypertensive patients. AB - The aim of this study was to evaluate whether salt-sensitivity in essential hypertension produces a significant comparative difference in diastolic function and ventricular mass when compared with sodium-resistance. Recent epidemiological data have demonstrated a positive correlation between sodium intake and arterial pressure. Furthermore, a positive correlation has been detected between sodium intake and left ventricular hypertrophy (LVH) independently of arterial pressure. Thirty-one patients who had never been treated before for uncomplicated hypertension were studied. Each subject received a 30 mmol/per day sodium diet for 14 days, supplemented with a further 190 mmol of sodium in the first study week (220 mmol for the first 7 days and 30 mmol for the second 7 days). Throughout the study compliance was assessed by measuring daily urinary sodium excretion. Sodium sensitivity of blood pressure was defined as the difference (5% or more) between blood pressure at the end of the low and high sodium intake periods. On this basis 16 patients were defined as salt-sensitive (SS) and 15 patients as salt-resistant (SR). The two groups were homogeneous for age, sex and race. Baseline mean arterial pressure (MAP) was comparable between SS (108 +/- 1.8 mm Hg) and SR (107 +/- 2.1 mm Hg, P = NS). Each patient was submitted to M MODE and two-dimensional echocardiogram studies in order to estimate left ventricular mass using the Penn conventional formula and parameters of left ventricular diastolic function. The left ventricular mass measurement showed higher values in the SS group although this did not reach statistical significance (118.4 +/- 4.4 vs 112.0 +/- 4.2 gr/mq, P = NS). Both interventricular septal and posterior wall thickness did not demonstrate significant differences between the two groups. The salt-sensitive group showed impaired left ventricular diastolic function; in particular, the first diastolic peak representing the early maximum of diastolic filling velocity (E) was lower in SS subjects than in SR subjects (71.6 +/- 2.9 vs83.1 +/- 3.3 cm/sec, P < 0.02). No significant difference was detected in the second peak representing the atrial maximum of filling velocity (A) (69.0 +/- 2.3 vs 66.0 +/- 2.0 cm/sec, P = NS). As a consequence the E/A ratio was significantly different (0.73 +/- 0.2 in the SR vs 1.2 +/- 0.04 in the SS group, P < 0.05). Moreover, the peak E integral was lower in SS than in SR subjects (8. 7 +/- 0.6 vs11.2 +/- 0.5 cm, P < 0.005; no difference for the peak A integral (6.0 +/- 0.3 vs 5.7 +/- 0.4 cm, P = NS). The E peak deceleration time was significantly reduced in the SS group (400.3 +/- 13.5 vs 500.9 +/- 12.8 cm/sec, P < 0.001). No significant difference was found for the isovolumetric relaxation time (IVRT) (95.7 +/- 4.3 vs 92.2 +/- 4.0, P = NS). Our data show an impaired diastolic function expressed by a reduced early diastolic filling velocity (peak E) and a significantly abnormal E/A ratio in SS in comparison with SR subjects. Therefore abnormalities in diastolic function are detectable earlier in SS hypertensive subjects than in SR irrespective of actual MAP. PMID- 10516744 TI - Severe hyperkalaemia due to the concomitant use of salt substitutes and ACE inhibitors in hypertension: a potentially life threatening interaction. AB - We report two cases of hyperkalaemia related to the use of the salt substitute 'Lo Salt' in hypertensive patients on treatment with ACE inhibitors. In each case serum potassium returned to the normal range after cessation of the salt substitute. Without vigilance the contribution of the salt substitute to hyperkalaemia would have been overlooked and an ACE inhibitor erroneously withdrawn. PMID- 10516745 TI - Molecular control of cell cycle progression in primary human hematopoietic stem cells: methods to increase levels of retroviral-mediated transduction. AB - Pluripotent hematopoietic stem cells (HSC) are the ideal targets for gene transfer because they can repopulate a sublethally irradiated recipient, giving rise to all lineages of blood cells. Thus, introduction of a corrected gene into HSC (stem cell gene therapy) should ensure persistent transmission of the gene. To date, the most efficient mode of gene delivery is via Moloney murine leukemia virus (MoMuLV)-based retroviral vectors which stably integrate into the genome of the target cell. The quiescent nature of HSC and the fact that MoMuLV-based retroviral vectors can only integrate into dividing cells are major obstacles in gene therapy. While increasing efforts have been directed toward identifying growth factors which facilitate division of primary hematopoietic progenitor and stem cells, little is known about the molecular mechanisms which these cells use to enter cell cycle. In this review, we will discuss the correlation between the hematopoietic inhibitory and growth factors and their impact on the regulation of the cell cycle components. PMID- 10516746 TI - Has the prognosis of adult patients with acute myeloid leukemia improved over years? A single institution experience of 784 consecutive patients over a 16-year period. AB - We reviewed the reports of 784 consecutive patients admitted to our department for newly diagnosed acute myeloid leukemia (AML) over a 16-year period. Median, 5 year and 10-year overall survivals were 9. 5 months, 17.3% and 11.7% respectively. Induction treatment (698 patients) resulted in 50% complete remissions (CR) (from 26.5% in secondary AML to 81.2% in patients <60 years with de novo AML). Period of diagnosis (1980-84/85-89/90-95) demonstrated a major significance for CR achievement and OS in multivariate analysis. In patients >/=60 years (372), CR rate increased (25% to 36.8%, P = 0. 03), and 5-year OS (3.7% to 10.6%, P = 0.022) improved, probably due to an increase in the proportion of patients administered conventional combined chemotherapy (54.5% to 83.8%, P < 0.0001). In younger patients CR rate continuously increased (61.5% to 74.8%, P = 0.028) with an associated improvement of 5-year OS (19.2% to 35.4%). No significant change in DFS and CR durations was observed. This large single center study on a large cohort of unselected AML patients reflects the improvement achieved in the management of AML patients, likely due to improvement of supportive care practices, administration of conventional induction to more elderly patients, and intensification of induction and post-remission treatments in patients <60 years. PMID- 10516747 TI - A phase II trial of induction and consolidation therapy of acute myeloid leukemia with weekly oral idarubicin alone in poor risk elderly patients. AB - We have conducted a phase II outpatient trial testing weekly oral administration of idarubicin (ZAVEDOS-ZVD) alone to determine the rate of objective response and toxicity in poor risk acute myeloid leukemia (AML) patients over 60 years of age. The treatment consisted of three phases: induction, with 20 mg/m2 of ZVD on days 1, 8, 15 and 22; consolidation with 20 mg/m2 of ZVD for 4 weeks; and maintenance with six cycles lasting 3 months and consisting of oral 6 mercapto-purine 2 mg/kg/day, 4 days a week for 2 months; subcutaneous cytarabine 1 mg/kg, once a week for 2 months; and oral ZVD 20 mg/m2 on day 1 and day 8 of the third month. In case of failure after induction course, patients received salvage treatment with 4 weekly oral doses of 40 mg/m2 ZVD. Fifty-one patients with a median age of 76 years were enrolled and could receive induction course. Of these 51 patients, 37 could receive subsequent courses, which consisted either of consolidation, or salvage. Only 11 patients underwent maintenance treatment. Sixty-three percent of patients had to be hospitalized during induction, for a median duration of 14.5 days, and 87% required hospitalization during salvage for a median duration of 17.5 days. Only five patients (38%) required hospitalization during consolidation. There were three toxic deaths (6%), two from hemorrhage and one from pulmonary embolism. The overall response rate was 29%, with 12 patients in complete response (25%) and two in partial response (4%). The median overall survival rate is 4 months for the whole population, and the median DFS is 9.6 months among the 14 responding patients. The results of this trial show that this new weekly schedule of oral ZVD chemotherapy is feasible and effective in poor risk elderly patients with AML. This regimen may be helpful for patients unable to tolerate intensive intravenous regimens, and is a real alternative to palliative treatments. PMID- 10516748 TI - Linkage analysis for ATM in familial B cell chronic lymphocytic leukaemia. AB - B cell chronic lymphocytic leukaemia (CLL) shows evidence of familial aggregation, but the inherited basis is poorly understood. Mutations in the ATM gene have been demonstrated in CLL. This, coupled with a possibly increased risk of leukaemia in relatives of patients with Ataxia Telangiectasia, led us to question whether the ATM gene is involved in familial cases of CLL. To examine this proposition we typed five markers on chromosome 11q in 24 CLL families. No evidence for linkage between CLL and ATM in the 24 families studied and the best estimates of the proportion of sibling pairs that share no, one or both haplotypes at ATM were not different from their null expectations. This would imply that ATM is unlikely to make a significant contribution to the three-fold increase in risk of CLL seen in relatives of patients. PMID- 10516749 TI - CD79b expression in B cell chronic lymphocytic leukemia: its implication for minimal residual disease detection. AB - The surface expression of CD79b, using the monoclonal antibody (Mab) CB3-1, on B lymphocytes from normal individuals and patients with B cell chronic lymphocytic leukemia (CLL) has been analyzed using triple-staining cells for flow cytometry. In addition, the clinical significance of CD79b expression in CLL patients and its possible value for the evaluation of minimal residual disease (MRD) was explored. A total of 15 peripheral blood (PB) samples from healthy blood donors, five bone marrow (BM) samples from normal donors and 40 PB samples from CLL untreated patients were included in the study. In addition we studied the expression of CD79b in B lymphocytes from five CLL patients after fludarabine treatment in order to support our method. The expression of CD79b in B lymphocytes from PB was analyzed by flow cytometry, using simultaneous staining with the Mabs CD22, CD79b, CD19 and CD5, CD79b and CD19. Since normal immature bone marrow B cells are CD79b-/dim+ on their surface, in BM samples we used the combination CD45, CD79b and CD19 selecting mature B lymphocytes according to their bright CD45 intensity. Cell acquisition was performed in two consecutive steps using a live gate drawn on SSC/CD19+ cells. For data analysis, the PAINT-A GATE PRO software (Becton Dickinson) was used. Dilution experiments of CD79b- CLL cells and CD79bdim+ CLL cells with normal PB and BM cells were performed in order to assess the sensitivity level of the technique for detection of CD79b /dim+residual CLL cells. All B lymphocytes from normal samples showed reactivity for the CD79b antigen. In contrast, CD79b was absent in 18/40 CLL patients (42.5%) and 20/40 CLL cases (50%) exhibited a low CD79b expression. Therefore, CD79b- B lymphocytes would be restricted to the CLL population and thus could be considered a 'tumor phenotype' for monitoring MRD in CLL patients. Dilution experiments indicate that the detection limit with this marker almost reaches the levels obtained by molecular biology methods as the PCR technique. All cases studied after fludarabine presented leukemic cells in their PB or BM samples detected by flow cytometry. Upon comparing the clinical and morphological characteristics of CD79b- and CD79b+ cases, all atypical CLL cases included in the present study were CD79b+ and advanced clinical stage (B and C Binet stage) was most frequently observed in CD79b+ cases than in CD79b- cases. PMID- 10516750 TI - Minimally differentiated acute myeloid leukemia in Taiwan: predominantly occurs in children less than 3 years and adults between 51 and 70 years. AB - Acute myeloid leukemia (AML) with minimal differentiation was usually referred to as acute undifferentiated leukemia in the past. With the help of immunophenotyping, this subtype of leukemia was shown to express myeloid antigens on the blasts and was designated AML-M0 by FAB Cooperative Study Group in 1991. Among the 423 consecutive newly diagnosed de novo AML at our institution, 12 (2.8%) were of M0 subtype. The proportion of M0 in AML was higher in children than in adults (8.2% vs 1.7%). Four other M0 patients referred from outside hospitals for immunophenotyping were also included in this study. There were two peaks in age distribution of these 16 patients: less than 3 years and between 51 and 70 years, respectively. Organomegaly was more common in patients with AML-M0 than in those with other subtypes (56.3% vs 29.2%, P = 0.025). The former patients had higher incidences of CD7 and CD34 expression on the leukemic cells than the latter ones (50% vs 16.9%, P = 0.003 and 69.2% vs 37.9%, P = 0.019, respectively). The patients with AML-M0 showed more frequent clonal chromosomal abnormalities in the leukemic cells than other AML patients (83.3% vs 53.9%, P = 0.039); the same is also true for complex cytogenetic aberrations (50% vs 11. 4%, P = 0.004). Adults with AML-M0 showed a lower complete remission (CR) rate and significantly poorer survival than those with non M0-AML. However there was no significant difference in outcome between the two groups of pediatric patients. In conclusion, AML-M0 is a unique subtype of leukemia that has distinct age distribution and shows different clinical and biological characteristics from other AML. Adult patients have poor prognosis. Whether pediatric patients had better outcome than adults needs to be clarified in further studies. PMID- 10516751 TI - The immunophenotype of minimally differentiated acute myeloid leukemia (AML-M0): reduced immunogenicity and high frequency of CD34+/CD38- leukemic progenitors. AB - Minimally differentiated acute myeloid leukemia (AML-M0) is a rare FAB subtype (2 3% of AMLs) of poor prognosis. The aim of our study was to characterize AML-M0 expression and regulation of adhesion/costimulatory molecule involved in immune recognition, to test blast in vitro immunogenicity, and to determine the percentage of leukemia progenitor cells. Here, we demonstrate that alloimmune recognition of AML-M0 in primary mixed lymphocyte reaction, as evaluated by IL-2 secretion of responding T cells, is reduced in comparison with more differentiated subtypes (128 +/- 95 pg/ml vs304 +/- 159 pg/ml, P < 0.05). These data are in line with low blast cell expression of major histocompatibility complex (MHC) class II DR molecules, and of the CD28 ligand B7-2, which plays an important role in AML immune recognition. Adhesion/costimulatory molecules were up-regulated by leukemic cell stimulation via CD40, and, although less efficiently, by gamma-IFN; both stimuli improved blast cell immunogenicity. We also demonstrate that AML-M0 have a very high percentage (40% +/- 30) of CD34+/CD38- leukemic clonogenic precursors in comparison with more differentiated AMLs (2.5% +/- 2) or non-leukemic CD34+hematopoietic precursors (1.8% +/- 0.8). Since the presence of a leukemic cell population at an early differentiation stage has been identified as a poor prognostic factor, we conclude that the high frequency of CD34+/CD38- blasts in AML-M0 may converge with already identified poor prognosis factors such as chemotherapy resistance and cytogenetic abnormalities. The clinical implications of AML-M0 impaired in vitroimmunogenicity and a high percentage of CD34+/CD38- blasts will require comparative analysis of additional patients. The increased immunogenicity of blast cells after CD40 triggering provide interesting clues for AML-M0 immunotherapy, that have to be confirmed with an in vivo leukemia model in mice. PMID- 10516752 TI - Monitoring of minimal residual leukemia in patients with MLL-AF9 positive acute myeloid leukemia by RT-PCR. AB - Twenty-seven patients with AML and MLL gene rearrangement were analyzed by a reverse transcriptase polymerase chain reaction (RT-PCR) for the MLL-AF9 translocation. The MLL-AF9 fusion transcript was detected in six patients. In five patients, the breakpoint of the AF9 gene was located within the recently described site A; in one patient, a novel breakpoint (AF9 site D) mapped to a position 377 bp 3' of site A. Five patients could be serially monitored for a period of 4-23 months. Two patients became two-step PCR negative in bone marrow and peripheral blood. Molecular remission was achieved rapidly after one cycle of induction chemotherapy. Both patients are in continuous complete remission (CR) at 22 and 15 months, respectively. Two patients who had achieved hematological CR did not become PCR negative and MLL-AF9 fusion transcripts were detectable in all samples after induction and consolidation chemotherapy. One patient relapsed 5 months after achieving CR. The other patient received allogeneic bone marrow transplantation from an HLA-identical sibling 2 months after achieving hematological CR and became PCR negative 4 weeks after transplantation. In the fifth patient, hematological CR could not be achieved with two cycles of intensive induction chemotherapy, and MLL-AF9 transcripts were present in all samples tested. Our data indicate that MLL-AF9 RT-PCR is specific for the t(9;11) translocation. PCR negativity can be achieved in responding patients already 1 month after induction chemotherapy. The fast reduction of MLL-AF9 positive blast cells below the detection limit of RT-PCR seems to be a prerequisite for long term CR. The results of RT-PCR may be useful for treatment decisions (eg BMT). PMID- 10516753 TI - The leukemogenic fusion of MLL with ENL creates a novel transcriptional transactivator. AB - Translocations affecting the chromosomal locus 11q23 are hallmarks of infant leukemias. These events disrupt the MLL gene (also ALL-1 or HRX) and fuse the MLL amino terminus in frame with a variety of unrelated proteins. The ENL gene on 19p13.1 is a recurrent fusion partner of MLL. Whereas potential functions have been suggested for isolated domains of either MLL or ENL no experimental data exist for the biological properties of the complete chimeric MLL-ENL protein. We show here that the fusion of MLL with ENL creates a novel molecule that is a potent general transcriptional transactivator in transient reporter gene assays. MLL-ENL strongly transactivated several unrelated promoters including the promoter of Hoxa7 a potential target gene for the unaltered MLL protein. This transactivation capability was cell type specific and it was critically dependent on the contributions of the methyltransferase-homology (MT) region of MLL in combination with the C-terminus of ENL. Squelching experiments and gel retardation studies identified the ENL C-terminus as a binding partner for an unknown factor and the MLL MT region as a unique general DNA binding motif. The potential implications of these findings for the leukemogenesis by MLL-ENL are discussed. PMID- 10516754 TI - Identification of novel chromosomal rearrangements in acute myelogenous leukemia involving loci on chromosome 2p23, 15q22 and 17q21. AB - Chromosomal translocations are frequently linked to multiple hematological malignancies. The study of the resulting abnormal gene products has led to fundamental advances in the understanding of cancer biology. This is the first report of t(2;15)(p23;q22) and t(2;17)(p23;q21) translocations in human malignancy. Patient 1, a 73-year-old male, was diagnosed with myeloblastic (FAB M1 sub-type) AML. Cytogenetic analysis showed a 47,XY,t(2;15)(p23;q22),+13 karyotype. Fluorescent in situ hybridization (FISH) showed that the PML gene was transferred intact to the short arm of chromosome 2 while the ALK gene on chromosome 2p23 was passively transferred to the long arm of chromosome 15. Patient 2 was a 60-year-old male diagnosed with monocytic (FAB M4-type) AML. Cytogenetic analysis showed 46,XY,t(2;17)(p23;q21) karyotype. FISH analysis showed that neither RARalpha nor ALK were disrupted by the translocation. None of the coding region of the three genes studied were translocated in these patients. This raises the possibilities that other neighboring genes could be involved or that noncoding regulatory sequences of the studied genes could be put in contact and deregulate expression of other genes. Alternatively, displacement of ALK, RARalpha and PML to novel positions could lead to loss of their normal regulation PMID- 10516755 TI - Regulation of CD95 expression and CD95-mediated cell death by interferon-gamma in acute lymphoblastic leukemia with chromosomal translocation t(4;11). AB - The regulatory effects of IFNgamma on CD95 expression and CD95-mediated cell death were investigated in three high-risk pro-B acute lymphoblastic leukemia (ALL) lines that carry the chromosomal translocation t(4;11)(q21;q23). These leukemias are characteristically refractory to conventional chemotherapeutic treatments operating through the induction of apoptosis. However, the mechanisms leading to increased cell survival and resistance to cell death in these leukemias are largely unknown. Interferon-gamma (IFNgamma), a potent inhibitor of hematopoiesis, acts in part by upregulating CD95 and sensitizing cells to CD95 induced apoptosis. The t(4;11) lines SEM, RS4;11, and MV4;11 expressed low levels of CD95, but were completely resistant to CD95-mediated death. Addition of IFNgamma markedly upregulated CD95 expression in SEM (8-9-fold), RS4;11 (2-3 fold), and MV4;11 (2-3-fold) lines. However, after treatment with IFNgamma, only an 11% increase in sensitivity to CD95-mediated cell death was observed in SEM cells, whereas RS4;11 and MV4;11 cells remained resistant. Cycloheximide, but not actinomycin D or brefeldin A, increased CD95-specific cell death only in IFNgamma treated RS4;11 cells by approximately 12%. Abundant levels of Bcl-2 and Bcl-XL, known to inhibit CD95-signaling in some cells, were present suggesting a possible role for both molecules in the resistance to CD95-mediated cell death. Resistance of the leukemic blasts to CD95-mediated cell death and the failure of IFNgamma to substantially sensitize the CD95-signaling pathway may contribute to the highly malignant phenotype of pro-B ALL with translocation t(4;11). PMID- 10516757 TI - Apoptosis in patients with myelodysplastic syndromes: differential involvement of marrow cells in 'good' versus 'poor' prognosis patients and correlation with apoptosis-related genes. AB - Apoptosis has been implicated in the pathogenesis of marrow failure in MDS and the coexistence of marrow hypercellularity along with blood cytopenias was seen as evidence of extreme cell death of mainly mature cells in the marrow (ineffective hematopoiesis). We investigated apoptosis in 53 patients with MDS, by using single-step DNA extraction and gel electrophoresis and then by separating fresh marrow mononuclear cells in CD34+ and CD34- populations and in situ single cell evaluation of the process. We also studied the expression of apoptosis-related genes, in total and separated mononuclear marrow cells and correlated the findings with clinical and laboratory characteristics. Patients with apoptosis had increased marrow cellularity, longer overall survival and a longer period for transformation to AML. In 'good' prognosis MDS patients, total mononuclear marrow cells, as well as isolated populations of CD34+ and CD34- cells showed significant degrees of apoptosis; in 'poor' prognosis cases, however, apoptosis was evident only in a large percentage of CD34+ marrow cells and not in total or CD34- cells. Absence of expression of both c-myc and p53 in total marrow cells was associated with significant degrees of apoptosis and in isolated CD34+ and CD34- marrow cells the phenomenon was inversely correlated with the level of bcl-2 expression. In conclusion, marrow apoptosis is detected in both CD34+ and CD34- cells in early MDS and seems to be restricted to CD34+ cells in advanced MDS cases. PMID- 10516756 TI - Quantitative analysis detects ubiquitous expression of apoptotic regulators in B cell non-Hodgkin's lymphomas. AB - To determine whether the expression levels of Bcl-2 family apoptotic regulators are correlated with the histopathological heterogeneity of B cell non-Hodgkin's lymphomas (NHL), we quantified their expression in malignant B cell populations isolated from 33 biopsy samples, including small lymphocytic lymphoma (SLL, n = 9), mantle cell lymphoma (MCL, n = 8), follicular lymphoma (FL, n = 8), and diffuse large cell lymphoma (DLCL, n = 8). Normal B cells purified from reactive lymph nodes and tonsil (n = 3) were used as controls. Cell lysates were analyzed by Western blotting, and signals quantified by densitometry. Expression of Bcl-2 and its homologues, Bcl-xL, Bcl-xS, Bax, Bad, Bak and Bag-1, was detected in all NHL cases, with wide variations between histological subtypes and within each subtype. Statistically significant differences were: (1) a higher level of Bad expression in DLCL compared to FL and MCL; (2) a lower level of Bak expression in FL compared to DLCL, SLL and MCL; and (3) a higher Bag-1 expression level in FL compared to SLL. When compared to NHL cells, normal B cells showed a higher level of Bax expression, and a lower level of Bcl-xL expression. Thus, quantitative analysis shows ubiquitous expression of Bcl-2 family proteins in normal and neoplastic B cells; the variations in expression levels may contribute to both the B-NHL clinicopathological diversity and the different apoptotic sensitivities of normal B cells vs B-NHL cells. PMID- 10516758 TI - Bryostatin 1 enhances paclitaxel-induced mitochondrial dysfunction and apoptosis in human leukemia cells (U937) ectopically expressing Bcl-xL. AB - The effects of the protein kinase C (PKC) activator and down-regulator bryostatin 1 were examined with respect to paclitaxel-induced apoptosis and antiproliferative activity in human myeloid leukemia cells (U937) displaying enforced expression of the anti-apoptotic protein Bcl-xL. Overexpression of Bcl xL blocked various aspects of paclitaxel-mediated apoptosis, including caspase-3 activation, degradation of poly(ADP-ribose) polymerase (PARP), loss of mitochondrial membrane potential (Delta Psim), and release of cytochrome c. However, subsequent (but not prior) exposure of paclitaxel-treated U937/Bcl-xL cells (500 nM; 6 h) to bryostatin 1 (10 nM; 15 h) restored the extent of apoptosis, caspase activation, and mitochondrial damage to levels approximating those in paclitaxel-treated empty-vector control cells (U937/Neo). Potentiation of paclitaxel-induced apoptosis by bryostatin 1 in U937/Bcl-xL cells occurred primarily in the G2M cell population, and was associated with alterations in Bcl xL gel mobility and a reduction in paclitaxel-mediated stimulation of CDK1 activity. Enhancement of paclitaxel-induced apoptosis by bryostatin 1 in Bcl-xL overexpressors was accompanied by a corresponding reduction in clonogenic potential. In contrast to its effects on apoptosis, bryostatin 1 failed to restore paclitaxel-mediated increases in free Bax levels in U937/Bcl-xL cells. Lastly, the actions of bryostatin 1 were mimicked by a pharmacologic inhibitor of the MEK1/MAP kinase pathway (PD98059), but not by SB203580, an inhibitor of p 38 MAP kinase. Moreover, sequential exposure of both U937/Neo or/Bcl-xL cells to paclitaxel followed by bryostatin 1 or PD98059 was associated with a net reduction in MAP kinase activity. Collectively, these findings indicate that protection against paclitaxel-mediated mitochondrial dysfunction and apoptosis in human U937 leukemia cells conferred by Bcl-xL overexpression can be substantially overcome by bryostatin 1 and possibly other agents that interrupt the MAP kinase signal transduction pathway. PMID- 10516759 TI - Bcl-2 family members in childhood acute lymphoblastic leukemia: relationships with features at presentation, in vitro and in vivo drug response and long-term clinical outcome. AB - We have found that, in addition to Bcl-2 and Bax, the expression levels of apoptosis inducers (Bad, Bak) and inhibitors (Bcl-xL, Mcl-1) were highly variable in blasts from 78 children with newly diagnosed acute lymphoblastic leukemia (ALL). The patients were enrolled in the national study ALL-7 of the Dutch Childhood Leukemia Study Group. In contrast to Bcl-2 that inversely correlated with %S-phase cells and WBC, and was lower in T than in B-lineage ALL, the Bcl-2 family members were not found to be associated with features at presentation. These expression levels were also compared with drug resistance in in vitro MTT (methyl-thiazol-tetrazolium) assays for prednisolone, vincristine and asparaginase in 46 children. Protein expression levels of the Bcl-2 family were not found to correlate with in vitroresistance to the individual drugs or the combined drug resistance profile. In addition, neither peripheral blast reduction after 1 week of prednisone monotherapy nor long-term disease-free interval or survival showed a correlation with protein expression. Our results indicate that the anti-proliferative function of Bcl-2 dominates its anti-apoptotic function in ALL, but neither Bcl-2 nor the Bcl-2 family members gained prognostic information in the risk-adapted protocol ALL-7. PMID- 10516760 TI - Perillyl alcohol selectively induces G0/G1 arrest and apoptosis in Bcr/Abl transformed myeloid cell lines. AB - The Bcr/Abl tyrosine kinase that is expressed from the Philadelphia chromosome protects leukemia cells from apoptosis caused by removal of growth factors or by cytotoxic agents and ionizing irradiation. This resistance to apoptosis is associated with a Bcr/Abl-mediated G2/M delay. Therefore, inhibiting Bcr/Abl signaling pathways should block the ability of the Bcr/Abl kinase to protect cells from apoptosis. The monoterpenes, limonene and perillyl alcohol (POH) are new anticancer agents that selectively induce apoptosis in neoplastic cells of a variety of rodent carcinoma models. Since the potential antitumor activities of monoterpenes overlap with signaling pathways affected by the Bcr/Abl kinase, POH and limonene were tested for antileukemia activity. POH, but not limonene selectively induced G0/G1 arrest followed by apoptosis in Bcr/Abl-transformed, but not nontransformed FDC.P1 and 32D myeloid cell lines. In contrast to their greater sensitivity to POH, Bcr/Abl-transformed cells were more resistant than nontransformed cells to several chemotherapy agents and ionizing irradiation. Since in Bcr/Abl-transformed cells, POH induces apoptosis associated with G0/G1 arrest, POH must activate an apoptotic pathway that is not protected by the Bcr/Abl-induced G2/M delay. Monoterpenes may represent novel agents for treating Ph+ leukemias. PMID- 10516761 TI - Cytogenetic analysis of the bipotential murine pre-B cell lymphoma, P388, and its derivative macrophage-like tumor, P388D1, using SKY and CGH. AB - Spectral karyotyping (SKY) and comparative genomic hybridization (CGH) were used to elucidate the divergent cytogenetic make-up of the prototypical bilineage lymphoblastic pre-B lymphoma, P388, and its progenitor macrophage-like tumor, P388D1. P388 was found to be diploid and genomically stable. P388D1 was triploid, highly unstable and characterized by numerous marker chromosomes (Chrs) and composite rearrangements. The karyotype of P388D1 was so complex that its clonal relatedness to P388 would have remained questionable without confirmation by molecular analysis of the clonotypic immunoglobulin heavy-chain and light-chain gene recombinations that coexisted in both tumors. The intrinsic instability of the P388D1 genome was indicated by the observation that only four out of 42 aberrations uncovered by SKY (in a total of 27 metaphases) occurred consistently (100% incidence), whereas 27 changes occurred non-randomly (27 to 96% incidence) and 11 alterations randomly (4 to 11% incidence). Persistent cytogenetic instability was also observed in P388 'macrophages' after phorbol ester- and ionomycin-induced conversion in vitro of P388 lymphoma cells. The 'cytogenetic noise' in these cells was manifested by a multiplicity of sporadic chromosomal aberrations; ie 25 distinct changes were identified by SKY in 40 metaphases. The results in P388D1 and P388 'macrophages' were interpreted to indicate that the myeloid differentiation program in the bipotential pre-B cell lymphoma P388 is invariably characterized by karyotypic instability. The study presented here demonstrates the power of the combined SKY and CGH approach to resolve complicated karyotypes of important and widely used mouse tumors. PMID- 10516762 TI - False human hematopoietic cell lines: cross-contaminations and misinterpretations. AB - The risk of adventitious contamination and subsequent overgrowth of cell lines by unrelated cells is a potential and often recurring problem where cells are grown and studied. This problem of intraspecies and interspecies cross-contamination among human cell lines has been recognized for over 25 years; incidences of cell cross-contamination between 17 and 35% have been reported. The most useful methods to detect human cell cross-contamination are DNA fingerprinting and cytogenetic analysis, each complementing the other. Using this combination, we found that in total 14.8% of the human hematopoietic cell lines received either from the original investigator (n = 117 cell lines) or from secondary sources (n = 72 cell lines) were cross-contaminated with another hematopoietic cell line and were thus false cell cultures. Another problem relates to the fact that not every cell line established from a patient with a hematopoietic malignancy is a malignant cell line; unintended immortalization of non-malignant B cells by 'passenger' Epstein-Barr virus (EBV) leads to the establishment of B lymphoblastoid cell lines (termed EBV+ B-LCLs), an event which is much more frequent than the establishment of a 'true' leukemia-lymphoma-myeloma cell line. These EBV+B-LCLs are most often (albeit not always) unrelated to the malignant clone. The misinterpretation of such EBV+ B-LCLs as true malignant hematopoietic cell lines (particularly in research areas investigating B cell-derived neoplasms such as myeloma) and the indiscriminate use of these cell lines may render some of the results of such studies irrelevant to the pathobiology of the disease concerned. However, a combination of markers commonly allows for an accurate determination of the nature of EBV+ B-LCLs: immunoprofile, cellular morphology, EBV status, and karyotype. In summary, the continuous need for vigilant quality and identity control procedures is emphasized by the high incidences of cross contaminated cell lines. Most laboratories using cells cultured in vitro maintain multiple cell lines. Such cell lines should be monitored regularly for their identity and specific characteristics in order to prevent invalidation of research work due to incidents of cell line cross-contamination or misinterpretation. PMID- 10516763 TI - Genetic manipulation of primitive leukemic and normal hematopoietic cells using a novel method of adenovirus-mediated gene transfer. AB - Gene transfer into early hematopoietic cells has been problematic due to the quiescent nature of primitive cells and the lack of gene transfer vehicles with high efficiency for hematopoietic cell types. Previously, we have shown that adenoviral vectors can be used for the transduction of normal human progenitors with gene transfer efficiencies of approximately 30%. However, this approach is limited by relatively slow uptake kinetics (24-48 h) and a strong dependence on the presence of exogenous cytokines. Thus, we have modified this approach by combining adenoviral vectors with polycations to generate a virus-polycation complex, or VPC. Vehicles of this nature, when composed of conventional adenoviral vectors and polyamidoamine dendrimers, are a highly efficient means of transducing both normal and acute myelogenous leukemia (AML) cells. Moreover, the kinetics of gene transfer are markedly increased using the VPC strategy, with approximately 70% of transduction complete within 2 h. In this study, using viruses that encode green fluorescence protein (GFP), or the T cell costimulatory molecule B7.1 (CD80), we show that VPC-mediated gene transfer is an effective means of transducing normal and AML cells, including those with a highly primitive phenotype. Our data suggest that transient genetic manipulation of primitive hematopoietic cells can readily be achieved and should therefore permit a variety of research and clinical endeavors. PMID- 10516764 TI - Identification of false-positive CBFbeta/MYH11 RT-PCR results. AB - Persistent problems with false positive results were encountered when carrying out a published RT-PCR method to detect the CBFbeta/MYH11 transcripts associated with the inv(16)(p13q22) cytogenetic abnormality in acute myeloid leukaemia. These were shown to be due to amplification of part of the intronic MYH11 sequence, presumably from very small amounts of contaminating DNA or unspliced primary RNA transcripts, amplified because of partial homology of the CBFbeta3 primer to intronic MYH11 sequence. PMID- 10516765 TI - Easy detection of all T cell receptor gamma (TCRG) gene rearrangements by Southern blot analysis: recommendations for optimal results. AB - Southern blot analysis of T cell receptor (TCR) gene rearrangements has proven to be a helpful tool to establish clonality in T cell leukemias and lymphomas. To improve the detection of clonal TCR gamma (TCRG) gene rearrangements by Southern blot analysis, we designed four new Jgamma probes and determined the most optimal restriction enzymes to be used with these probes. Based on detailed analysis of the sequences as well as on hybridization experiments with the TCRGJ21 probe, the Jgamma1.2 and Jgamma2.1 downstream areas were found to be highly homologous, suggesting that during evolution the duplication of the Jgamma region was followed by deletion of the tentative Jgamma2.2 gene segment. Southern blot analysis of 51 T cell acute lymphoblastic leukemias (T-ALL) revealed that all TCRG gene rearrangements can be detected by use of the TCRGJ13 probe in EcoRI digests and the TCRGJ21 probe in PstI digests. Additional probes and digests allow a more precise identification of the exact type of TCRG gene rearrangements in the majority of cases. Almost 90% of the TCRG gene rearrangements in T-ALL involved the Jgamma2 region (16% Jgamma2.1 and 72% Jgamma2.3), whereas Jgamma1 region rearrangements were particularly found in TCRgammadelta+ T-ALL. This information has implications for design of primer sets for PCR analysis at diagnosis and for PCR target choice in detection of minimal residual disease during follow-up of T-ALL patients. PMID- 10516766 TI - Testing for tumor drug resistance in the age of molecular medicine. A contribution to the Debate Round-Table on Phenotypic and Genotypic Analyses of Multidrug Resistance (MDR) in Clinical Hospital Practice. AB - The detection of multidrug resistance (MDR) in clinical hospital practice represents an important strategy to combat clinical tumor drug resistance. Predicting the response of tumors to cytostatic drugs is of prognostic value. The 'Debate Round-Table on Phenotypic and Genotypic Analyses of Multidrug Resistance (MDR) in Clinical Hospital Practice' was launched in 1997 to address specific questions on this topic. The results published thus far are a rich source to learn about the promises and pitfalls of methods, eg surrogate and functional tests and protein or mRNA expression assays as well. In the present paper, some requirements are discussed for applications of drug resistance testing in clinical routine diagnostics. To improve the detection of low-level resistance, established methodologies may be strengthened with respect to: (1) standardization of sample handling, antibodies, PCR primers, and detection reagents; (2) standardization of protocols and far reaching automation in performance and evaluation of results to ensure high quality control criteria. Sophisticated new techniques will feature: (1) high-throughput analyses for the 'horizontal screening' of single drug resistance genes in large numbers of patient samples at economically fair costs; (2) 'vertical screening' of a large number of resistance mechanisms operating upstream or downstream of the actual drug-target interaction sites, in order to detect more complex and multifaceted genotypes and phenotypes of multidrug resistance. PMID- 10516767 TI - Exclusion of Leu1 and Leu2 genes as tumor suppressor genes in 13q14.3-deleted B CLL. AB - The chromosomal region 13q14.3 is frequently deleted in B cell chronic lymphocytic leukemia (B-CLL) and it is supposed that a tumor suppressor gene, involved in this leukemogenesis, is located in this area. The first exons of two genes, Leu1 and Leu2, mapped in a minimally deleted 13q14.3 region, are systematically lost in B-CLL sharing a 13q14.3 deletion. These two genes have been proposed as strong tumor suppressor gene candidates. However, in a study on 15 13q14.3 deleted B-CLL, we found three patients in which this critical region was not involved. Because of these results and that no mutations were detected on the two genes in a previous study, we think that Leu1 and Leu2 can be excluded as tumor suppressor genes. PMID- 10516768 TI - Significantly lower relapse rate for TEL/AML1-positive ALL. PMID- 10516769 TI - Vascular endothelial growth factor at high plasma levels is associated with extranodal involvement in adult T cell leukemia patients. PMID- 10516770 TI - Translisin recognition site sequences flank translocation breakpoints in a Philadelphia chromosome positive chronic myeloid leukemia patient expressing a novel type of chimeric BCR-ABL transcript (E8-INT-A2) PMID- 10516771 TI - Waldenstrom's macroglobulinemia is a biological syndrome which may occur during the evolution of different types of low grade B cell lymphoma. PMID- 10516772 TI - Bimonthly cranial MRI activity following an isolated monosymptomatic demyelinating syndrome: potential outcome measures for future multiple sclerosis 'prevention' trials. AB - Patients with characteristic white matter lesions on MR imaging are at increased risk for the subsequent development of clinically definite MS (CDMS) following an isolated, monosymptomatic demyelinating syndrome (IMDS). These MR positive first onset patients are thus candidates for MS prevention trials. Seven consecutive patients with IMDS and two or more periventricular and/or oval lesions on MR imaging were enrolled into a prospective trial based on serial T2-weighted and enhanced MR imaging at two month intervals for 12 - 15 months. Forty-seven new enhancing lesions were detected with triple dose MR contrast compared to 31 lesions using the standard dose. Four of the seven patients accounted for 98% of all enhancing lesions in this study, while the remaining three patients showed little MRI or clinical disease activity. New T2 lesion counts correlated strongly with enhanced lesion counts (r=0.82 - 0.98). We detected 49 new T2-hyperintense (T2) lesions based on short-interval (2 monthly) follow-up, and 31 new T2-lesions comparing exit and entry examinations. These results suggest several potential MR based strategies for evaluating first onset patients in a phase III MS prevention trial. Since these patients have a small average T2-lesion load, it is quite easy to visually detect new T2 lesions. As a result, at a bimonthly study interval, T2 weighted lesion analysis is an effective measure of cumulative disease activity, provided high-resolution T2-weighted imaging studies are acquired to quantitate new T2-lesions. Enhanced lesion activity remains an important measure of blood - brain - barrier breakdown and may predict short term MRI and clinical disease activity in IMDS patients. PMID- 10516773 TI - Magnetisation transfer of normal appearing white matter in primary progressive multiple sclerosis. AB - Patients with primary progressive multiple sclerosis may develop severe disability despite a paucity of lesions on conventional magnetic resonance imaging, raising the possibility that intrinsic changes in normal appearing white matter (NAWM) contribute to disability. This study has measured magnetisation transfer ratio (MTR), an index of tissue damage, of NAWM in 52 patients with primary progressive multiple sclerosis and 26 healthy controls. Absolute values of MTR were obtained from the genu of the corpus callosum and pons, and mean values were calculated from bilateral regions in the centrum semiovale, frontal white matter, parieto-occipital white matter and posterior limb of the internal capsule. The median MTR was lower in all regions in patients compared to controls. Median values (per cent units) were significantly lower in corpus callosum (39.73 vs 40.63; P=0.01), frontal white matter (39.11 vs 39.59; P=0.01) and centrum semiovale (37.21 vs37.82; P<0.05). This study has demonstrated small but widespread decreases in MTR in NAWM in primary progressive multiple sclerosis supporting the hypothesis that there are intrinsic changes in NAWM which may contribute to disability in this patient group. PMID- 10516775 TI - The effects of anxiety on psychiatric morbidity in patients with multiple sclerosis. AB - Our objective was to assess the point prevalence and effects of clinically significant anxiety in patients with Multiple Sclerosis (MS). One hundred and fifty two consecutive patients with MS attending an outpatient clinic underwent neurological examination and were assessed for psychopathology with the Hospital Anxiety and Depression Scale, the 28 item General Health Questionnaire and a questionnaire probing suicidal thoughts or intent. Clinically significant anxiety, either with or without depression, was endorsed by 25% of patients, three times the rate for depression. Females were significantly more anxious than males. Anxiety co-morbid with depression, rather than anxiety or depression alone, was associated with increased thoughts of self harm, more somatic complaints and greater social dysfunction. Patients with increased psychopathology were not more likely to be taking psychotropic medication. The results provide preliminary evidence that anxiety, which may be often overlooked clinically, is a frequent accompaniment to depression, thereby adding to the morbidity associated with MS. The implications of the findings to MS patients' quality of life are emphasised. PMID- 10516774 TI - Type I interferons and the quality of life of multiple sclerosis patients. Results from a clinical trial on interferon alfa-2a. AB - The objective of the study was to examine whether the beneficial effect of treatment of interferon alfa-2a on multiple sclerosis seen by magnetic resonance imaging is reflected in a corresponding improvement in the quality of life (QoL) and to address the impact of adverse events related to this treatment on the QoL. The study was a randomised double-blinded placebo-controlled treatment trial including 97 relapsing-remitting multiple sclerosis patients. Thirty-two patients received 4.5 MIU recombinant interferon alfa-2a, 32 patients received 9.0 MIU recombinant interferon alfa-2a and 33 patients received placebo treatment for 6 months. All patients were followed up 6 months after end of treatment. QoL was assessed according to the eight scales of the SF-36 Health Survey and measured at baseline, month 3, 6 and 12. The effect found on MRI was not reflected in a corresponding change in the QoL. We found a relationship between the presence of new enhancing lesions and reduced QoL among the placebo patients, whereas this was not found among the patients treated with interferon. The presence of the adverse events fatigue, myalgia, headache and weakness were significantly negatively correlated to several of the QoL dimensions. Conclusively, the treatment with interferon alfa-2a does not seem to improve the patients' QoL after 6 months of treatment, in spite of a marked effect measured by MRI. The treatment is followed by adverse events that negatively affected the QoL. PMID- 10516776 TI - Impaired interleukin-12 production in multiple sclerosis patients. AB - Multiple Sclerosis (MS), a disease of the human central nervous system, is believed to be a T cell mediated autoimmune disorder with genetic and environmental influences. Interleukin-12 (IL-12), a proinflammatory cytokine produced primarily by antigen presenting cells is a potent inducer of interferon gamma (IFN-gamma) and other Th1 cytokines that may play an important role in MS pathogenesis. We have investigated IL-12 production induced by the T cell independent pathway in untreated and IFN-beta treated MS patients, healthy individuals and other neurological disease (OND) patients in response to the human pathogen Staphylococcus aureus. We report that peripheral blood mononuclear cells (PBMC) from untreated MS patients produce normal amounts of the biologically active IL-12 p70 heterodimer but significantly less free IL-12 p40 heavy chain than PBMC from both healthy and disease controls when challenged in vitro with Staphylococcus aureus. Both mRNA expression of the inducible IL-12 p40 chain and protein levels were found to be reduced in untreated MS patients. No decrease in the production of the IL-12 p40 was seen in MS patients on IFN-beta therapy. The decreased production of IL-12 p40 heavy chain is not attributed to increased IL-10 secretion, a defect in the production of cytokines by macrophages or the number of cytokine producing cells. The factor(s) responsible for the decrease in p40 remain to be determined. Since IL-12 p40 antagonizes the biological activity of IL-12 in vitro and in vivo, identification of a defect in the 'natural' antagonist of IL-12, may provide the basis for immune therapy. PMID- 10516777 TI - Increased urinary nitric oxide metabolites in patients with multiple sclerosis correlates with early and relapsing disease. AB - Nitric oxide (NO) has been implicated in the immunopathogenesis of MS as a potential mediator of neuronal loss. To investigate the role of.NO in the development of progressive disease we measured the NO metabolites (nitrate and nitrite) and neopterin, in the urine of 129 patients with demyelinating disease (DD): 23 with clinically isolated syndromes compatible with demyelination and in 46 relapsing remitting (RR) and 60 patients with progressive MS. Eighty-nine of these 129 patients underwent Gd-enhanced MRI. In addition 58 normal control subjects (NC), 19 AIDS and 35 rheumatoid arthritis (RA) patients were studied. Patients with DD, AIDS and RA had significantly elevated urinary nitrate plus nitrite (nit : creat. urine) and neopterin (neopt : creat.urine) to creatinine ratios compared to NC subjects. (Median[25th - 75th%] nit : creat.urine: NC=1183[962 - 1365] vs DD=1245[875 - 2403], AIDS=1686[1231 - 2531], and RA=1950[1214 - 2726] mumol/mol, P<0.001 and median[25th - 75th%] neopt : creat.urine: NC=99[76 - 151] vs DD=163[119 - 266], AIDS=972[653 - 1456], and RA=389[257 - 623] mu mol/mol, P<0.001). Patients with early DD and RR MS had significantly elevated nit : creat.urine compared to patients with progressive MS (nit : creat. urine: 1612[1020 - 2733] vs 1159[790 - 1641] mu mol/mol, P=0.006). The nit : creat.urine and neopt : creat.urine did not correlate with clinical relapse or MRI activity. Excretion of.NO metabolites is increased in patients with early or relapsing-remitting disease.NO appears to be a double-edged sword, mediating tissue damage and modulating complex immunological functions which may be protective in MS. PMID- 10516778 TI - Cytokine analysis in multiple sclerosis by competitive RT - PCR: A decreased expression of IL-10 and an increased expression of TNF-alpha in chronic progression. AB - Multiple sclerosis (MS) is an inflammatory, demyelinating disease that is specific to the central nervous system. Cytokines are thought to be key mediators of the autoimmune attack against central nervous system myelin in MS. To investigate the involvement of cytokines in MS, the mRNA levels of interferon gamma (IFN-gamma), tumor necrosis factor alpha (TNF-alpha), interleukin-4 (IL-4) and interleukin-10 (IL-10) in peripheral blood mononuclear cells without stimulation in vitro were quantified by a competitive reverse transcription polymerase chain reaction technique. The level of IL-10 specific mRNA was significantly decreased in 47 MS patients compared with 42 healthy controls (P<0.0001). TNF-alpha was significantly increased in MS patients compared with healthy controls (P=0.014), especially in the patients with chronic progressive MS (P=0.0003). Thus we conclude that there are significant in vivo alterations in cytokine gene expression in the periphery in MS. PMID- 10516779 TI - Disease steps in multiple sclerosis: a longitudinal study comparing disease steps and EDSS to evaluate disease progression. AB - Clinical assessment of outcome in multiple sclerosis (MS) patients is problematic since the disease can affect different aspects of the central nervous system and follow a variable course. Recently, we developed Disease Steps, a simple approach for evaluating disease progression. Previously, we found that Disease Steps was easy to use, had uniformly distributed scores and low inter-rater variability. Our current objective was to test the long-term use of Disease Steps together with the most widely utilized clinical outcome measure in MS, the Expanded Disability Status Scale (EDSS) in assessing clinical progression. Over 4 years, 804 patients were classified using both EDSS and Disease Steps. Each patient was assessed at least twice. Follow-up results included annual status and time-to event analysis examining median staying times within a level of Disease Steps or EDSS. We found that the two scales behaved similarly and correlated strongly with each other. For both Disease Steps and EDSS, patients with milder levels of disability and relapsing-remitting disease demonstrated a higher likelihood of changing scores over time and shorter median staying times compared to more disabled, chronic progressive patients. These findings have important implications for patient selection in clinical trials and for the design of future measurements of clinical outcome in MS. Furthermore, Disease Steps may serve as a simple, practical tool for the nonspecialty neurologist to follow patients over time and serve as a guide in therapeutic decision making. Our findings further document the general progressive nature of MS when a large cohort is followed in an MS specialty clinic over time. PMID- 10516780 TI - The association of the human herpesvirus-6 and MS. AB - Given the clinical and pathological nature of Multiple Sclerosis (MS), a viral infection has long been hypothesized as part of the etiology. In this study we investigated the possibility that the human herpesvirus-6 (HHV-6) is present in a dormant or active phase in the tissue of MS patients, specifically oligodendrocytes. Using PCR assays of MS and non-MS brain sections with primers prepared against the HHV-6 structural protein 101, the results demonstrated that 36% of MS brains were positive for the virus, while 13.5% of non-MS brains were positive. Antibody to the HHV-6 structural protein was also used in immunohistochemical experiments in brain tissue. 47% (7/15) of MS brains were positive for HHV-6, whereas 0/16 controls were positive. In addition, MS patients demonstrated high immune reactivity to this virus, even when compared to auto immune diseases, which might cause polyclonal activation. Sera obtained from MS and control patients revealed that the IgM response to the HHV-6 virus was significantly elevated in 80% patients compared to 16% non-MS controls, P<.001. The above experiments strongly suggest that a significant number of MS brain samples contain HHV-6 antigens and genomic fragments in a dormant or active phase compared to control specimens and that MS patients mount a brisk, early IgM response. PMID- 10516781 TI - Gait abnormalities in minimally impaired multiple sclerosis patients. AB - Subclinical evidence of gait abnormalities were identified in a group of seven patients with multiple sclerosis, EDSS scored 0 - 2, without functional limitations. A movement analysis technique was used to identify gait parameters indicative of impaired motor function during walking. Abnormalities related primarily to time-distance parameters (reduced speed of progression, shorter strides, prolonged double support phase) and muscular function (premature recruitment of gastrocnemius and late relaxation of tibialis anterior during stance phase) were identified regardless the severity of the clinical score. The gait analysis procedure was able to provide the clinician with evidence of motor abnormalities prior to functional disturbance observable by a trained physician. These minimal dysfunctions may have resulted from reflex mechanisms impaired by delayed transmission through long loop pathways or else as a result of a nonspecific protective gait strategy to improve balance control. The technique described in this study may be useful to identify earlier starting points for follow-up and physiotherapy. PMID- 10516783 TI - Malaria mortality and morbidity in Africa. PMID- 10516782 TI - A profile of multiple sclerosis: the New York State Multiple Sclerosis Consortium. AB - We have obtained a current profile of multiple sclerosis York State through a centralized patient registry and standardized data collection instrument associated with the New York State Multiple Sclerosis Consortium of 12 MS centers located throughout the state. Data from the first 3019 patients with clinically definite MS revealed a clear relationship between MS disease type, duration of disease, and severity of physical disability. Patients with relapsing disease had disease durations approximately half as long as those with progressive forms of the disease (means approximately 6 years versus 11 years). The majority of patients with relapsing disease had Expanded Disability Status Scale (EDSS) scores of 4.0 or less (self-sustained, fully ambulatory), whereas the majority of patients with progressive disease types had EDSS scores of 6.0 or greater (at least unilateral assist for walking). These findings emphasize the importance of early intervention in patients with relapsing disease to slow or prevent the accumulation of physical disability associated with progressive types of disease. Progressive disease was associated with lack of full-time employment and being disabled before the age of 60 years. Patients with relapsing disease were more likely to be employed and have private forms of insurance, whereas patients with progressive types of disease were more likely to have government-supported insurance to cover medical expenses. PMID- 10516784 TI - Health in development. PMID- 10516785 TI - Estimating mortality, morbidity and disability due to malaria among Africa's non pregnant population. AB - The contribution of malaria to morbidity and mortality among people in Africa has been a subject of academic interest, political advocacy, and speculation. National statistics for much of sub-Saharan Africa have proved to be an unreliable source of disease-specific morbidity and mortality data. Credible estimates of disease-specific burdens are required for setting global and national priorities for health in order to rationalize the use of limited resources and lobby for financial support. We have taken an empirical approach to defining the limits of Plasmodium falciparum transmission across the continent and interpolated the distributions of projected populations in 1995. By combining a review of the literature on malaria in Africa and models of acquired functional immunity, we have estimated the age-structured rates of the fatal, morbid and disabling sequelae following exposure to malaria infection under different epidemiological conditions. PMID- 10516786 TI - Mortality and causes of death in Jordan 1995-96: assessment by verbal autopsy. AB - Mortality indicators and causes of death in Jordan were assessed by verbal autopsy. A random sample of 100 clusters of ca. 300 households each were monitored for one year by notification assistants selected from the study area itself. Registered deaths were reported to research assistants who visited the family to complete the verbal autopsy form, which was structured and contained about 100 questions. Causes of death were determined by two physicians according to preset algorithms. A total of 965 deaths were reported among 198,989 persons, giving a crude death rate of 5 per 1000 population per year. The three leading causes of death were diseases of the circulatory system, malignancies and accidents. In the absence of a health information system, verbal autopsy as implemented in Jordan can serve as a reliable substitute. PMID- 10516787 TI - Global burden of Shigella infections: implications for vaccine development and implementation of control strategies. AB - Few studies provide data on the global morbidity and mortality caused by infection with Shigella spp.; such estimates are needed, however, to plan strategies of prevention and treatment. Here we report the results of a review of the literature published between 1966 and 1997 on Shigella infection. The data obtained permit calculation of the number of cases of Shigella infection and the associated mortality occurring worldwide each year, by age, and (as a proxy for disease severity) by clinical category, i.e. mild cases remaining at home, moderate cases requiring outpatient care, and severe cases demanding hospitalization. A sensitivity analysis was performed to estimate the high and low range of morbid and fatal cases in each category. Finally, the frequency distribution of Shigella infection, by serogroup and serotype and by region of the world, was determined. The annual number of Shigella episodes throughout the world was estimated to be 164.7 million, of which 163.2 million were in developing countries (with 1.1 million deaths) and 1.5 million in industrialized countries. A total of 69% of all episodes and 61% of all deaths attributable to shigellosis involved children under 5 years of age. The median percentages of isolates of S. flexneri, S. sonnei, S. boydii, and S. dysenteriae were, respectively, 60%, 15%, 6%, and 6% (30% of S. dysenteriae cases were type 1) in developing countries; and 16%, 77%, 2%, and 1% in industrialized countries. In developing countries, the predominant serotype of S. flexneri is 2a, followed by 1b, 3a, 4a, and 6. In industrialized countries, most isolates are S. flexneri 2a or other unspecified type 2 strains. Shigellosis, which continues to have an important global impact, cannot be adequately controlled with the existing prevention and treatment measures. Innovative strategies, including development of vaccines against the most common serotypes, could provide substantial benefits. PMID- 10516788 TI - Cost-effectiveness of competing diagnostic-therapeutic strategies for visceral leishmaniasis. AB - Reported are the results of a formal decision analysis which facilitated the choice of the most appropriate test-treatment strategy for visceral leishmaniasis in areas where the disease is endemic. The following strategies were compared: treatment of all suspects (strategy A); testing by means of parasitological investigation followed by treatment of positives (strategy B); two-step testing by means of the direct agglutination test (DAT) followed by treatment of patients with high titres as well as those with parasitologically confirmed borderline titres (strategy C); and DAT followed by treatment of positives (strategy D). The results for each strategy were expressed as costs in US$ per death averted. The effectiveness of strategies C and D was close to that of strategy A and far better than that of strategy B. The cost-effectiveness ratio for strategies C and D (US$ 465 per death averted) was not substantially higher than that of testing by means of parasitological investigation followed by treatment of positives (strategy B), which was the most cost-effective strategy at US$448 per death averted. At current prices of antimonial drugs, the cost of test-treatment strategies depends more on the cost of treatment than on that of testing. The use of a sensitive serological test such as the DAT is recommended as the basis of test-treatment strategies for visceral leishmaniasis in areas where the disease is endemic. PMID- 10516789 TI - Pharmaceutical donations by the USA: an assessment of relevance and time-to expiry. AB - This paper assesses the relevance and time-to-expiry of pharmaceutical donations by the USA by means of a convenience sample of two private voluntary organizations. Data were collected on 16,566 donations shipped between 1994 and 1997 for the two organizations to a total of 129 countries. For three field study countries (Armenia, Haiti, and the United Republic of Tanzania), between 37% and 65% of donated unique drug products were on the recipient countries' essential drugs lists, and between 50% and 80% were either on these lists or were permissible therapeutic alternatives. Between 10% and 42% were not listed on either the national essential drugs lists or the WHO Model List of Essential Drugs, nor were they permissible therapeutic alternatives. For the worldwide data set, the median times to expiry when shipment by the organizations took place were 599 and 550 days; about 30% of shipment items had a year or less of shelf life, and about 6% had less than 100 days of shelf-life. Although a majority of the donations fulfilled the criteria of relevance and time-to-expiry, a substantial proportion failed to do so. Actions are proposed with a view to improving the relevance and time-to-expiry of USA pharmaceutical donations. PMID- 10516790 TI - Eradication of poliomyelitis in Cuba: a historical perspective. AB - The eradication of poliomyelitis in Cuba, for which effective vaccines had to be acquired, is reviewed in this article. The strategy for eradication was based on mass immunization campaigns for the annual delivery of two doses of trivalent Sabin oral poliovirus vaccine (OPV). Except during the first campaign in 1962, the ages of the children for immunization were determined through national serological surveys of the entire country, including rural and urban areas. The interruption of wild virus transmission had been suspected since 1967 in Cuba, and since 1970 no studies have detected any wild virus. The important role of political and social organizations in the success of the programme and in the execution of the mass immunization campaigns is underscored. Countries that have successfully interrupted poliovirus circulation should maintain high immunization coverage for as long as there are other countries in the world where poliovirus still exists. PMID- 10516791 TI - Training medical assistants for surgery. AB - A successful programme is reported from Mozambique for training middle-level health workers to perform fairly advanced surgical procedures in remote areas where the services of consultants are virtually unobtainable. Manpower and financial constraints obliged Mozambique to train medical assistants to perform surgical work in rural areas, where three broad priorities were identified: pregnancy-related complications, trauma-related complications, and emergency inflammatory conditions. Since 1984, 20 health workers have emerged from three year courses to become tecnicos de cirurgia (assistant medical officers), and it is expected that there will be 46 by 1999. The training comprises two years of lectures and practical sessions in the Maputo Central Hospital, and a practical internship lasting a year at a provincial hospital. Three workshops organized since 1989 suggest that the upgraded personnel are performing well. More detailed evaluation and follow-up are in progress. Throughout 1995 a follow-up was conducted on 14 assistant medical officers. They performed 10,258 surgical operations, some 70% of which were emergency interventions. Low rates of complication occurred and postoperative mortality amounted to 0.4% and 0.1% in emergency and elective interventions respectively. PMID- 10516792 TI - Diabetes mellitus: a "thrifty" genotype rendered detrimental by "progress"? 1962. PMID- 10516793 TI - Disease burden predicts US government health research funding. PMID- 10516794 TI - Varicella-zoster virus disease. PMID- 10516795 TI - A preamble on parkinsonism. PMID- 10516796 TI - Research on genes: promises and limitations. PMID- 10516798 TI - Can estrogen keep you smart? Evidence from clinical studies. AB - OBJECTIVE: To review and critically analyze the biological plausibility of and the clinical empirical evidence concerning a link between estrogen levels and memory in women. DATA SOURCES: MEDLINE search of the literature published from 1980 to 1998. Studies published between 1952 and 1980 that were known to the author were also included. STUDY SELECTION: Sixteen prospective, placebo controlled studies in humans. DATA SYNTHESIS: Most of the studies that used neuropsychological tests with known reliability and validity found that estrogen maintained aspects of memory in women. CONCLUSIONS: Estrogen specifically maintains verbal memory in women and may prevent or forestall the deterioration in short- and long-term memory that occurs with normal aging. There is also evidence that estrogen decreases the incidence of Alzheimer disease or retards its onset or both. PMID- 10516797 TI - Motor deficits and schizophrenia: the evidence from neuroleptic-naive patients and populations at risk. AB - Patients with schizophrenia and high-risk populations have elevated rates of eye movement abnormalities. However, it is not known whether these abnormalities are specific to eye movements or whether they are also found in more traditional domains of motor control. Most studies examining the motor function of patients with schizophrenia have involved patients treated with medication; abnormalities in motor function could be a result of treatment rather than the disease itself. If motor abnormalities are related to schizophrenia, they should also be found in neuroleptic-naive patients and possibly in high-risk populations in whom eye movement abnormalities are also observed. We reviewed relevant empirical papers published in the last 35 years. Results suggest that approximately one-fifth of neuroleptic-naive patients with schizophrenia have increased rates of parkinsonism and neurological soft signs. In high-risk populations, replicated findings include delayed motor development in preschizophrenia subjects, and poor motor skills in the offspring of patients with schizophrenia. In first-degree relatives, increased rates of neurological soft signs were reported. These findings suggest that motor abnormalities are not limited to eye movements and may constitute markers of vulnerability. The literature has several weaknesses that should be addressed in future studies. PMID- 10516799 TI - Semantic processing deficits in patients with Parkinson's disease: degraded representation or defective retrieval? AB - OBJECTIVE: To determine whether degraded representations (characterized by small differences between word sense frequencies), or defective competitive processes (high levels of word sense lateral inhibition), individually or jointly, can give rise to parkinsonian semantic deficits. DESIGN: Computer model of semantic processing. OUTCOME MEASURES: Correct sense selection, defined by the activation of the word sense unit that first reaches the 0.5 activation threshold. If Parkinson disease (PD)-like errors are observed only at high levels of lateral inhibition, independently of low or high sense frequency deltas (SFDs), this would indicate that a defective competitive process alone could account for the errors. Alternatively, if PD-like errors were observed at any level of lateral inhibition, exclusively with low SFD words, this would indicate that degraded representations alone could account for the errors. RESULTS: Neither degraded representations nor defective competitive processes alone can account for parkinsonian semantic errors. An interaction between the 2, however, correctly reproduces both increased errors and longer latency responses. CONCLUSIONS: Competing explanations for semantic deficits in patients with Parkinson's disease need to be integrated in order to develop effective interventions (e.g., estimating the amount of context required to improve semantic processing performance). PMID- 10516800 TI - Effects of typical antipsychotic drugs and risperidone on the quality of sleep in patients with schizophrenia: a pilot study. AB - OBJECTIVE: To investigate the effects of a newer antipsychotic drug, risperidone (a potent serotonin 5-HT2A/2C and dopamine D2-receptor blocker), on the quantity and quality of sleep in patients with schizophrenia. DESIGN: Prospective pilot study. SETTING: Outpatient treatment at a mental health hospital. PATIENTS: Two groups of age- and sex-matched patients with schizophrenia receiving either risperidone (n = 8) or a typical antipsychotic drug (n = 8), and a group of age- and sex-matched controls (n = 8). OUTCOME MEASURES: Sleep quality, measured by a visual analogue scale, and sleep continuity, measured using a movement index calculated from actigraph data. RESULTS: Patients with schizophrenia had more disturbed sleep than controls. Compared with patients treated with typical antipsychotic drugs, patients treated with risperidone reported significantly better sleep quantity and quality as well as general functioning. CONCLUSION: Improvement by risperidone may be related to 5-HT2A/2C receptor blockade; however, further controlled studies are required to confirm these results. PMID- 10516801 TI - Doppler ultrasonographic examination of the leg veins of patients with Parkinson disease. AB - OBJECTIVE: To determine the incidence of deep venous thrombosis (DVT) in patients with Parkinson disease. DESIGN: Prospective study. SETTING: Outpatient neurology clinic. PATIENTS: Eighty-one patients with Parkinson disease. OUTCOME MEASURES: Duplex ultrasonographic scans consisting of M mode images and compression images, Doppler flow assessment and augmentation of flow assessment. RESULTS: Four patients had leg DVT; in 3 of the patients the thrombi were in calf veins, whereas in 1 patient the thrombosis was in the superficial femoral vein. Of the patients with DVT, 1 (1.23%) had stage 2 Parkinson disease, 1 (1.23%) had stage 2.5, and the other 2 (2.46%) had stage 4. CONCLUSIONS: There was no statistically significant difference in the incidence of DVT among patients who were more severely disabled by Parkinson disease. However, an overall incidence of DVT of 4.9% in a group of asymptomatic patients is clinically meaningful, suggesting that patients with Parkinson disease are at risk for asymptomatic leg DVT. PMID- 10516802 TI - Memory deficits in patients with schizophrenia: preliminary data from the Wechsler Memory Scale-Third Edition support earlier findings. AB - OBJECTIVE: To determine whether memory data presented for a schizophrenia sample in the Technical Manual of the Wechsler Memory Scale-Third Edition support trends identified in a previously published review of studies employing an earlier version of the instrument, the Wechsler Memory Scale-Revised. DESIGN: Archival: reformulation of published data. PATIENTS: Patients with schizophrenia, Alzheimer's disease, Korsakoff's syndrome or traumatic brain injury (TBI) for whom intelligence and memory data were reported in the Technical Manual of the Wechsler Adult Intelligence Scale-Third Edition Wechsler Memory Scale-Third Edition (WAIS-III WMS-III). OUTCOME MEASURES: Mean Full Scale, Verbal, and Performance Intelligence Quotients of the WAIS-III and mean WMS-III Immediate and General Memory Indexes. Single-trial learning and learning slope data were also culled from the WAIS-III WMS-III Technical Manual. RESULTS: Memory indexes for patients with Alzheimer's disease or Korsakoff's syndrome were substantially lower than those for patients with schizophrenia or TBI. In tests of learning processes, patients with schizophrenia had an inferior ability to repeat material presented just once, in comparison with the standardization sample. However, they did relatively better with repeated presentations than patients with Alzheimer's disease or Korsakoff's syndrome. The learning slope for patients with schizophrenia demonstrated an ability to absorb and consolidate increasing amounts of material with repeated exposure that is inconsistent with pronounced memory impairment. CONCLUSIONS: Although patients with schizophrenia exhibit new learning deficiencies, their memory capabilities are not substantially weaker than their general intellectual abilities, and do not approach the memory impairment exhibited by patients with Alzheimer's disease or Korsakoff's syndrome. PMID- 10516803 TI - Pseudo-narcolepsy: case report. AB - This report describes the case of a 44-year-old woman presenting to a Sleep and Alertness clinic with symptoms of narcolepsy. The patient had clinical and polysomnographic features of narcolepsy, which disappeared after disclosure of severe psychological stress. Following a discussion of the differential diagnosis of narcolepsy, alternative diagnoses are considered. The authors suggest that the patient had a hysterical conversion disorder, or "pseudo-narcolepsy." Careful inquiry into psychological factors in unusual cases of narcolepsy may be warranted. PMID- 10516804 TI - The presence of lupus anticoagulant and anticardiolipin antibodies in patients undergoing long-term neuroleptic treatment. PMID- 10516805 TI - Understanding autism: does anyone read new medical journals? PMID- 10516806 TI - [The tooth apex resection of molars]. PMID- 10516807 TI - [Tooth apex resection in the area of the maxillary sinus]. PMID- 10516808 TI - [Drug-induced interstitial pulmonary disease]. PMID- 10516809 TI - [Endothelium at the center of coronary pathologies: new data]. PMID- 10516810 TI - Molecular mimicry between S-antigen and viral peptides. PMID- 10516811 TI - [Epidemiologic and developmental aspects of congenital cardiopathies in the Sfax pediatric service: 123 cases]. PMID- 10516812 TI - [Utility of the Rhame and Sudderth model in nosocomial infection surveillance]. PMID- 10516813 TI - [Association of diabetes mellitus and other organ specific autoimmune diseases: 51 cases]. PMID- 10516814 TI - [Diabetic ketosis and ketoacidosis]. PMID- 10516815 TI - [Role of acupuncture in therapy: personal experience]. PMID- 10516816 TI - [Bourneville tuberous sclerosis: developmental aspects in 7 cases]. PMID- 10516817 TI - [CHARGE association: 4 case reports]. PMID- 10516818 TI - [Clinical and angiographic aspects of angioid streaks]. PMID- 10516820 TI - [Continuing medical education in the private sector]. PMID- 10516819 TI - [Continuing medical education. Experience of the private practice general physician]. PMID- 10516821 TI - [Reflections on continuing medical education in the public sector]. PMID- 10516822 TI - [Continuing medical education of the surgeon. Experience of the Tunisian Association of Surgery]. PMID- 10516823 TI - [Continuing medical education. Quality assurance stage!]. PMID- 10516824 TI - [Continuing medical education in the Internet era]. PMID- 10516825 TI - [Professional practice and continuing medical education. The responsibility of medical faculties. Interfaculty Group of Medical Education]. PMID- 10516826 TI - [Continuing medical education: should it be an ordinal obligation?]. PMID- 10516827 TI - [Current status and problems in continuing medical education in Tunisia]. PMID- 10516828 TI - [Continuing medical education: perspectives, recommendations and conclusions]. PMID- 10516829 TI - [Teaching guide for continuing medical education]. PMID- 10516830 TI - [List of medical specialty societies in Tunisia]. PMID- 10516831 TI - [Medical risk assessment at the turn of the century. International developments and trends--exemplified by some typical changes]. AB - This article analyses the effects that the increasing amount of medical information available today can have on risk assessment, using examples taken from the fields of serological laboratory diagnosis (HIV, Hepatitis C) and sonographic and molecular genetic examination methods. In general, it can be said that new methods of diagnosis help achieve a more exact and refined prognosis, although their results cannot always be implemented consistently, especially in so-called borderline cases. The second section provides a detailed analysis of access to and availability of medical information in proposal forms, drawing on experience gained from the most important markets. PMID- 10516832 TI - [Evaluation of residual working capacity with lung function studies]. AB - Lung diseases are one of the most frequent causes of permanent occupational disability. The assessment of the insured person's remaining ability to work by evaluating the respiratory parameters is by no means trivial. Nevertheless, no clear overview has been published yet. In this paper tables and evaluation schemes are presented for the quick and reliable assessment of the degree of the occupational disability of persons with respiratory diseases. PMID- 10516833 TI - [Hepatitis C. Diagnostic and prognostic aspects from the insurance medicine viewpoint]. AB - Hepatitis C is a chronic infection with potentially serious long-term effects. An acute clinical presentation is the exception, often the disease is only diagnosed through routine screening (e.g. as a blood donor) or work-up for elevated liver enzymes. The silent course of this disease also makes it difficult to interpret epidemiological studies. Potential biases need to be considered which may lead to underestimation or overestimation of prevalence and incidence data. Special attention is needed in evaluating long-term effects as the studies usually deal with small numbers of HCV positives and their selection may not have been randomised. An assessment of the prognosis is difficult, especially in the absence of a series of liver enzyme measurements and if the viral load is unknown. The wide availability of diagnostic tests harbours a potential for anti selection. Caution is therefore required when designing underwriting guidelines; only well documented cases should be accepted. A response to therapy (e.g. with interferon), alone, does not prove a good prognosis, rather, the course over a 2 year follow-up may give relevant prognostic information. PMID- 10516834 TI - [Follow-up and prognosis in chronic polyarthritis]. AB - The spectrum of rheumatoid arthritis (RA) ranges from benign remitting manifestations to rapidly progressive forms with increased mortality. About 10% of the patients show an intractable rapidly progressive course associated with severe extraarticular manifestations. Within the first three years, 70% of the patients develop radiological erosions of the joints and 31% deformities of the hands. Life expectancy is shortened by 3-18 years. Particularly, infections are more frequent causes of death in RA compared with controls. Work disability occurs in about a quarter of the patients within the first three years of RA and in 43-85% after eight to ten years. The ratio of direct to indirect costs is 1:3 in RA. Preliminary data show that regular rheumatological treatment leads to a marked reduction in indirect costs caused by production loss. The most important early indicators of an unfavourable disease course are the large number of swollen joints, early severe functional impairment, highly elevated laboratory markers of inflammation and rheumatoid factor. Knowledge of the current data regarding the course and prognosis of RA are helpful for assessment of the disease for insurance purposes. PMID- 10516835 TI - [Herbal drugs of foreign cultures and medical systems exemplified by Indian incense. Considerations regarding social and insurance medicine expert assessment]. AB - Increase preparations are being used with increasing frequency in Western medicine. In experimental investigations, an anti-inflammatory effect has been shown. Clinical observations relate to increase applications in rheumatoid arthritis, malignant brain tumors, inflammatory bowel diseases and asthma bronchiale. A relevant clinical effect of increase preparations has not been ascertained. In Germany, the registration of increase preparation H 15 as a drug has been withheld by the federal drug registration office (former "Bundesgesundheitsamt"). Health funds may reimburse costs for increase preparations in exceptional cases only. PMID- 10516836 TI - [Immunomodulating mistletoe therapy by lectin standardization: a double-edged sword?]. AB - Taking advantage of an unique, legally generous regulation, proprietary anthroposophic and phytotherapeutic mistletoe preparations are on the market in Germany. One constituent of the extract, the galactoside-specific lectin, is a potent biological response modifier in a very narrow low-dose range. Although the clinical implications of the lectin effects remain to be rigorously defined, this activity already prompted companies to eliminate the common batch-to-batch variations in favor of standardization, keeping the lectin content, which could otherwise vary drastically, purportedly constant. Based on literature data, immunomodulation by the lectin involves enhanced secretion of multifunctional proinflammatory cytokines such as IL-6. The apparently context-dependent ambivalence of their actions includes capacity to serve as autocrine and paracrine tumor growth and survival factors for a wide variety of tumor cell types in vitro and in vivo, as illustrated by the literature presented. The potential for clinical risks is indicated to be non-negligible, e.g. for lymphomas, advanced-stage melanomas and renal cell carcinomas. Moreover, negative effects of immunomodulatory lectin or extract treatment have already been reported. To reliably prove clinical efficacy and exclude lack of undesired side effects for each tumor class and stage, it is mandatory to evaluate the performance of this experimental therapy modality exclusively in relevant preclinical settings and strictly controlled clinical studies to obey the generally accepted rule: primum non nocere. PMID- 10516837 TI - [Comment of J. Fritze: Is therapy of obesity with orlistat medically necessary?]. PMID- 10516838 TI - Violence as a public health emergency. AB - Violence threatens or denies not only the health of those who are directly affected but diminishes the whole human process. Neither the violent acts themselves, nor the repercussions of these tragedies, are limited to one geographic or social setting, and it is not just the frequency of violent actions that threatens the health of the nation. It is the ripple effect that occurs from each of these incidents that affects everyone. This article discusses the problem of national violence in the United States and examines preventive programs. PMID- 10516839 TI - Emergency department evaluation of child abuse. AB - Child abuse has been documented in various forms since the beginning of recorded history. This article reviews the legal aspects of reporting child abuse, the epidemiology of child abuse, various physical manifestations of child abuse, and effective treatment procedures. It is only with the appropriate interventions that physicians can begin to make an impact on the future of abused children. PMID- 10516840 TI - Adolescent violence. AB - Adolescents are the victims, perpetrators, and witnesses of violent acts in the home, at school, and on the streets. Adolescent violence describes a dynamic, destructive, and repercussive process. This article discusses the unique aspects of adolescent violence and the involvement undertaken by the emergency department when adolescent violence results in the need for emergency care. PMID- 10516841 TI - Violence against women. AB - Violence against women has received increasing public attention over the past 20 years. The past two decades have highlighted the problem of intimate partner violence as a major cause of intentional injury to women. Many of these women present themselves to emergency departments where emergency physicians have a unique opportunity to intervene. PMID- 10516843 TI - Management of elder abuse in the emergency department. AB - Emergency physicians are in an ideal position to diagnose and intervene in suspected cases of elder abuse. This article reviews domestic violence against the elderly, highlighting the risk factors for elder abuse, its prevalence, and related medicolegal issues. A special section on mistreatment of the elderly in long-term care facilities is included. In addition to the management of elderly abuse in the emergency department, possible future directions for improved detection of elder abuse or neglect are reviewed. PMID- 10516842 TI - Domestic violence against women in the international community. AB - The social, economic, legal, and linguistic isolation that women in the international community often endure may make it more difficult for them to seek medical, legal, or social assistance in an effort to free themselves from abusive domestic situations. This article describes and defines potentially dangerous domestic situations and discusses the role of the emergency department in lessening the global health threat of violence among women. PMID- 10516844 TI - Violence during pregnancy. AB - This article discusses intimate partner abuse during pregnancy. The population at risk is defined, including risk behaviors, possible identifying factors during presentation to the emergency department, and available outcome data on violence to the fetus and the pregnant mother. Legal and ethical issues are also discussed. Intervention techniques are presented, emphasizing the role of the emergency physician in coordinating referrals to social service agencies and helping victims develop safety plans. PMID- 10516845 TI - Review of psychological issues in victims of domestic violence seen in emergency settings. AB - When a history of domestic violence is discovered during a universal screening in the emergency department, emergency staff typically feel ill equipped to address the underlying psychological, behavioral, and health problems of the victim. This article reviews the known characteristics of ongoing relationships in which one partner exerts coercive control over another, with emphasis on the effects of abuse on the victim's physical and mental health. These effects include actual injury, multiple stress-related physical conditions, substance abuse, and a variety of psychiatric problems, including depression, anxiety and anxiety disorders, dissociation and dissociative disorders, and post-traumatic stress disorder. A patient-centered approach allows emergency response staff to tailor intervention to victims' real and perceived needs, fulfilling their professional obligation to provide meaningful help to female victims of domestic violence seen in emergency settings. PMID- 10516846 TI - Violence in the prehospital setting. AB - The unique position EMS teams hold in their communities (as liaison between the general public and the health care system) allows for a greater understanding of all of the issues effecting patient care and safety. Prehospital care providers are essentially the only medical personnel who are routinely on the streets and in the homes of victims of violence. This article examines the role of EMS systems in regard to interpersonal violence. PMID- 10516847 TI - Sexual assault. AB - According to recent national studies, one in six women and one in 33 men will experience an attempted or completed rape during their lifetime. Although most rapes are never reported, victims that do report them often present to the emergency department for intervention. The emergency physician must be able to treat acute injuries, accurately collect and record evidence, offer STD and pregnancy prophylaxis, offer emotional support (with the social worker and rape crisis advocate), and provide appropriate referrals for follow-up care. The emergency physician can play a crucial role in easing the transition from victim to survivor with initial treatment. This article summarizes current recommendations for evaluation and treatment of sexual assault victims. PMID- 10516848 TI - Firearms and family violence. AB - Firearms contribute significantly to morbidity and mortality in family violence. This article discusses the debate on gun use for protection and guns in the home. Weapons-related risks in the setting of intimate partner violence are closely reviewed. Recommendations for physicians are discussed in the context of firearms and family violence. PMID- 10516849 TI - Violence in the emergency department. AB - Violence in the emergency department is a frequently encountered problem that is often not promptly or adequately addressed. This article outlines the epidemiology of violence in the emergency department, including patients at greatest risk for aggressive behavior. The necessary steps to identify and approach these patients and recommended methods for sedation and restraint are discussed. PMID- 10516850 TI - Factors regulating the growth of metastatic cancer in bone. AB - Metastatic tumor cells can interfere directly with the function of bone cells involved in normal bone remodeling or indirectly by influencing the behavior of hematopoietic, stromal and other cells in bone marrow that interact with bone cells. Recent studies of metastatic cancer have revealed that tumor cells interact closely with vascular endothelial cells, basement membrane and bone marrow stromal cells through cell surface proteins or by releasing factors which affect the function of these cells. Bidirectional interaction between marrow cells and tumor cells can give the latter a selective advantage for growth in bone which can lead to the destruction of or to increased production of bone matrix. Understanding of the mechanisms involved in tumor metastasis and growth in bone has increased in recent years, and in this review we shall describe current knowledge of these mechanisms and of the predilection of certain types of cancers to metastasize to bone, their growth in the bone microenvironment and interactions between them and bone cells. Because metastatic breast cancer has been studied more than any other, we shall focus on it as a representative example, although the general principles apply to other types of cancer and to myeloma. PMID- 10516851 TI - Prospects for the treatment of endocrine-responsive tumours. AB - Much has been achieved over the last 30 years to improve the treatment of hormone dependent cancer of the breast, ovary and prostate. The development of the antioestrogen tamoxifen (Nolvadex) spear-headed a range of drugs that counter the growth-promoting action of the female and male sex steroid hormones. An important additional benefit of endocrine therapies has been their low toxicity compared with conventional cancer chemotherapy thereby providing effective treatment with few serious side-effects and a sustained quality of life. Although some currently available therapies improve patient disease-free survival and overall survival, particularly when given in an adjuvant setting, they are not cures. There is, therefore, a continued need to develop newer therapies that extend the effectiveness of those currently available. This is particularly important when tumours either fail to respond or develop resistance to endocrine therapy. In this review, we examine how our improved understanding of the factors that influence the progression of endocrine-related tumours is leading to the development of novel therapies to treat both hormone-dependent and -independent tumours. PMID- 10516852 TI - Involvement of steroid hormone and growth factor cross-talk in endocrine response in breast cancer. AB - Multiple lines of evidence implicate steroid hormone and growth factor cross-talk as a modulator of endocrine response in breast cancer and that aberrations in growth factor signaling pathways are a common element in the endocrine resistant phenotype. Delineation of these relationships is thus an important diagnostic goal in cancer research, while the targeting of aberrant growth factor signaling holds the promise of improving therapeutic response rates. PMID- 10516853 TI - Prolactin involvement in breast cancer. AB - Normal development and differentiation of the mammary gland are profoundly influenced by prolactin (PRL). In rodent mammary cancer PRL plays a well defined role, but its role, in human breast cancer has not been appreciated until recently. It is now clear that breast tissue, both normal and malignant, is a significant source of extrapituitary PRL. Thus an autocrine/paracrine role of PRL in human breast cancer may be invoked. Both PRL and PRL receptor mRNA are expressed in the vast majority of breast cancer biopsies independent of estrogen and progesterone receptor status. An autocrine/paracrine PRL acting in human breast cancer requires that this hormone's action be blocked at the cellular level, as opposed to suppressing the synthesis and secretion of pituitary PRL. Mutants of PRL or human growth hormone are being explored which act as selective PRL antagonists. In addition, tamoxifen has been shown to act locally at the target tissue by binding directly to the PRL receptor and thus inhibiting PRL's action. These strategies may have clinical relevance in treating PRL-responsive human breast cancer. PMID- 10516854 TI - Tumor suppressor genes in breast cancer. PMID- 10516855 TI - Nesidioblastosis. PMID- 10516856 TI - Combined treatment with the 5 alpha-reductase inhibitor PNU 157706 and the antiandrogen flutamide on the Dunning R3327 prostatic carcinoma in rats. AB - The steroid 5 alpha-reductase enzyme catalyzes the conversion of testosterone to the potent androgen 5 alpha-dihydrotestosterone (DHT). PNU 157706, a novel, potent and selective dual 5 alpha-reductase inhibitor, was reported to be effective in inhibiting the growth of established tumors in the Dunning R3327 rat prostatic carcinoma model. We have studied the efficacy of combined treatment with PNU 157706 and the antiandrogen flutamide in this prostatic tumor in rats. Rats with tumor diameters of about 1 cm were treated orally 6 days a week for 9 weeks with PNU 157706 (10 mg/kg per day) alone or in combination with flutamide (1 and 5 mg/kg per day). Animals were killed 24 h after the last treatment and ventral prostates were removed for testosterone and DHT determination. PNU 157706 reduced the growth of established tumors by 36%; flutamide showed a slight effect at 1 mg/kg per day (24% inhibition), while at the dose of 5 mg/kg per day it reduced tumor growth by 48%. The combination of PNU 157706 with the lower dose of flutamide caused an additive tumor growth inhibition (60%) and the combination with the higher dose of flutamide resulted in a better inhibition of tumor growth (68%) than did either treatment alone. Castration resulted in marked tumor growth inhibition (76%). Ventral prostate weight was more markedly reduced by PNU 157706 treatment than by flutamide; combined treatment was as effective as castration. Prostatic DHT content was markedly reduced by PNU 157706 (93%), whereas prostatic testosterone increased (137%). Concomitant treatment with flutamide partially antagonized the testosterone increase induced by PNU 157706 and did not modify the already considerable suppression of DHT. These data show that the inhibitory effects of PNU 157706 and flutamide on Dunning prostatic tumor growth are additive, thus supporting the rationale of this combination therapy in advanced prostate cancer, in order to achieve adequate androgen blockade with minimal side effects. PMID- 10516857 TI - The road to success: factors affecting the speed of promotion of academic radiologists. AB - RATIONALE AND OBJECTIVES: The authors' purpose was to determine the factors influencing the speed of promotion of academic radiologists. MATERIALS AND METHODS: Three hundred forty-three surveys from faculty members of academic radiology departments with continuous academic careers were analyzed for time in rank at assistant and associate professor levels in relation to publication rate, grant funding rate, and distribution of professional time. Individuals promoted faster than the median time (6 years for assistant professors, 5 years for associate professors) were considered "fast track" and were compared with the remainder of the group. RESULTS: At the assistant professor level, fast track individuals had significantly higher rates of total publications and original articles than did others. At the level of associate professor, fast track individuals had significantly faster rates of publication of original articles, but no significant difference existed in total publication rate. No significant difference was found in the rate of founding of fast track individuals and others. Those with funding were not more likely to be on a fast track than those without funding. Fast track individuals spent significantly more time in administration at the assistant professor level than did other faculty, but no other significant differences were discovered in time distribution at the assistant or associate professor level. CONCLUSIONS: The rate of publishing original articles at the assistant and associate professor levels and the rate of overall publication at the assistant professor level were the most important parameters in predicting speed of promotion. PMID- 10516858 TI - Small-bowel motility disturbances: a comparison of small-bowel series and antroduodenal manometry. AB - RATIONALE AND OBJECTIVES: The authors' purpose was to compare the findings of small-bowel series with those of antroduodenal manometry to determine whether normal findings from a small-bowel series would make it unnecessary to perform antroduodenal manometry. MATERIALS AND METHODS: The findings from 33 small-bowel series performed on patients who had undergone antroduodenal manometry were retrospectively reviewed for abnormalities, including dilatation, transit time, fold thickening, and increased fluid. Antroduodenal manometry findings were classified into the following categories: normal, myopathy, neuropathy, obstructions, or nonspecific conditions. RESULTS: Nine of 12 patients with specific abnormalities at antroduodenal manometry had abnormal results from the small-bowel series. Of seven patients with normal small-bowel series results, three had abnormal antroduodenal manometry results--two had previously undergone vagotomy with neuropathic changes and one had myopathic changes. CONCLUSION: Small-bowel series and antroduodenal manometry are complementary examinations. Only a small number of patients with normal small-bowel series results will have abnormal results at antroduodenal manometry. A large number of patients with motility abnormalities have a combination of nonspecific changes, such as dilatation and increased fluid, at a small-bowel series. PMID- 10516859 TI - How experience and training influence mammography expertise. AB - RATIONALE AND OBJECTIVES: The authors evaluated the influence of perceptual and cognitive skills in mammography detection and interpretation by testing three groups representing different levels of mammography expertise in terms of experience, training, and talent with a mammography screening-diagnostic task. MATERIALS AND METHODS: One hundred fifty mammograms, composed of unilateral cranial-caudal and mediolateral oblique views, were displayed in pairs on a digital workstation to 19 radiology residents, three experienced mammographers, and nine mammography technologists. One-third of the mammograms showed malignant lesions; two-thirds were malignancy-free. Observers interacted with the display to indicate whether each image contained no malignant lesions or suspicious lesions indicating malignancy. Decision time was measured as the lesions were localized, classified, and rated for decision confidence. RESULTS: Compared with performance of experts, alternative free response operating characteristic performance for residents was significantly lower and equivalent to that of technologists. Analysis of overall performance showed that, as level of expertise decreased, false-positive results exerted a greater effect on overall decision accuracy over the time course of image perception. This defines the decision speed-accuracy relationship that characterizes mammography expertise. CONCLUSION: Differences in resident performance resulted primarily from lack of perceptual learning experience during mammography training, which limited object recognition skills and made it difficult to determine differences between malignant lesions, benign lesions, and normal image perturbations. A proposed solution is systematic mentor-guided training that links image perception to feedback about the reasons underlying decision making. PMID- 10516860 TI - Tracheobronchial metal stents: effects of covering a bronchial ostium in pigs. AB - RATIONALE AND OBJECTIVES: The purpose of this study was to examine the effects of placing a metal stent across a bronchial orifice. MATERIALS AND METHODS: Nine pigs were used as test subjects, because the right upper lobe bronchus comes directly off the trachea in these animals. One of three types of metal stents was placed into the trachea of each pig and covered the orifice of the right upper lobe bronchus. Follow-up studies were performed at 1 and 3 months to evaluate the right upper lobe for signs of bronchial obstruction, infection, and atelectasis. The animals were sacrificed at 3 months to study the histopathologic changes of the trachea and lungs. RESULTS: Two upper lobe bronchi remained patent; seven were obstructed by granulation tissue or plugs of mucus and inflammatory cells. Right upper lobe infiltration and atelectasis were seen in eight animals. Interestingly, radiographic opacities were also common in other lung segments. There was a tendency toward fewer and less extensive lung opacities at 3 months compared with that at 1 month. At histopathologic examination, areas of both acute and chronic pneumonia were found in the right upper lobe of all animals. The segment of trachea covered by the stent was lined with a thin layer of granulation tissue containing neutrophils, monocytes, and lymphocytes. The stent luminal surface was covered with columnar, cuboidal, and stratified squamous epithelium. Tracheal stenosis was seen in three animals because of excessive granulation tissue in two and a collapsed stent in one. CONCLUSION: Placement of metal stents in pig trachea covering the orifice of the right upper lobe bronchus resulted in retention of secretions and secondary infection in the right upper lobe and other distant lung segments. PMID- 10516861 TI - US in the emergency department: our experience and proposed resolution of a conflict between emergency medicine and academic radiology. PMID- 10516862 TI - Two-year-old girl with thymic calcifications and liver function abnormalities: a case from the A3CR2 film panel. PMID- 10516863 TI - Apoptosis in cardiac diseases--new opportunities for novel therapeutics for heart diseases. AB - Apoptosis as defined by contemporary science describes a form of cell death that involves discrete genetic and molecular programs, de novo protein expression and unique cellular phenotype. Evidence for the existence of apoptosis in the human heart has been reported in various cardiac diseases, including ischemic and non ischemic heart failure, myocardial infarction and arrhythmias. Among the most potent stimuli that elicit cardiomyocyte apoptosis are: oxygen radicals (including NO), cytokines, (FAS/TNF alpha family of cytokines) and growth factors/energy deprivation. Several complex signal transduction pathways have been implicated in execution of cardiomyocyte apoptosis, including: Fas/TNF alpha receptors signaling, stress or mitogen activated protein kinases (SAPK/MAPK), sphingolipids metabolites (ceramide), G-protein coupled receptor (GPCR) signaling (G alpha i, G alpha q) and NF kappa B activation. Apoptosis of cardiac myocytes may contribute to progressive pump-failure, arrhythmias and cardiac remodeling. The recognition of numerous molecular targets associated with cardiomyocyte apoptosis that are amenable for pharmacologic manipulation, may provide novel therapeutic strategies for diverse cardiac ailments, as recently suggested by pharmacologic studies in experimental animals. PMID- 10516864 TI - Vasomotion of coronary arteries: from nitrates to nitric oxide. AB - The vascular endothelium is a source of substances particularly nitric oxide that act in a paracrine fashion either to contract or relax smooth muscle cells and this function is disturbed in atherosclerotic arteries. The endothelium derived nitric oxide contribute in the basal vasomotor tone of epicardial normal and atheromatous coronary coronary arteries, and of atheromatous coronary stenoses in patients with stable angina. Nitrates, by acting indirectly to increase cGMP in smooth muscle cells, dilate most coronary stenoses and normal coronary segments, decrease left ventricular afterload and diastolic filling pressure and improve the distribution of coronary flow to subendocardial zone. Large and small epicardial coronary vessels responded differently to endothelium-dependent and independent vasodilators. PMID- 10516865 TI - Verapamil in acute myocardial infarction. The rationales of the VAMI and DAVIT III trials. AB - Verapamil is well tolerated in patients with stable and unstable angina pectoris, as well as in patients with threatened infarction. No data exist documenting verapamil to be inferior to beta-blockers in these stages of coronary heart diseases. In the prethrombolytic era i.v. intervention with verapamil did not add any benefit in the early phase of AMI. However, a retrospective analysis suggests the hypothesis that i.v. verapamil given in combination with thrombolysis may improve the prognosis, and an ongoing study (VAMI trial) examines this hypothesis. When given in the late in-hospital phase of AMI to patients without heart failure verapamil significantly reduces mortality and morbidity. When given in the late in-hospital phase to patients with heart failure verapamil did not cause the course or prognosis to deteriorate and might even improve it in patients with residual ischaemia, especially when given in combination with an ACE-inhibitors. A planned study (DAVIT III study) has to confirm these preliminary data. PMID- 10516866 TI - Early treatment with verapamil or diltiazem in patients with acute myocardial infarction: safety and possible beneficial effects. AB - While dihydropyridine calcium antagonists may be harmful in the immediate peri infarction period, the effect of verapamil or diltiazem in these circumstances in uncertain. We utilized the GUSTO-1 formula to calculate the predicted 30-day mortality risk in a cohort of 352 patients with acute myocardial infarction presenting < 6 hours after onset of symptoms, with ECG changes consistent with eligibility for thrombolysis. All patients were treated with an intravenous bolus dose of verapamil followed by oral verapamil (240-360 mg/day) or diltiazem (180 360 mg/day), in the absence of specific contraindications. Predicted 30-day mortality risk was then compared with the actual 30-day mortality rate of the cohort. The actual 30-day mortality of the cohort was 3.7% (95% CI: 2.0,6.3); this was significantly lower than that predicted by the GUSTO-1 formula (7%). A similarly significantly lower mortality (7.5% vs 19.3%) was observed in a "high risk" subset of patients. Of the 13 patients who died, only 4 developed cardiogenic shock. It is concluded that verapamil and diltiazem can be administered safely in a selected patients with evolving acute transmural myocardial infarction. While the current data suggest that this form of treatment may be beneficial, definitive conclusions in this regard should await further randomized studies. PMID- 10516868 TI - Disopyramide decreases the fasting serum glucose level in man. AB - The effect of disopyramide, a class Ia antiarrhythmic drug, on the serum glucose level was evaluated in 6 consecutive in-patients. A 19-hour starvation test was repeated with oral administration of sustained-release disopyramide (150 mg) 0 and 12 hours after starting the test. Serum glucose levels during the starvation test decreased with disopyramide administration from a mean value of 96.5 +/- 1.8 to 85.9 +/- 1.4 mg/dl (24 samples, p < 0.05). The average reduction of the serum glucose level by disopyramide in each patient was 9.7 +/- 2.2 mg/dl. The decrease in the serum glucose level was not related to the serum concentration of disopyramide or serum creatinine levels. The decrease in the serum glucose level was larger in older patients (r = 0.75) and in light patients under 45 kg. These results suggested that disopyramide reduced the fasting serum glucose levels even in normal ranges as a common side effect of the drug, and that not only the occurrence of severe hypoglycemia but also the decrease in glucose levels were influenced by multiple factors including age and body weight. PMID- 10516867 TI - The effect of verapamil on left ventricular remodelling and diastolic function after acute myocardial infarction (the Verapamil Infarction Study on Remodelling and Relaxation--VISOR). AB - The VISOR is a double blind, randomized, placebo-controlled study aimed to assess the effects of early and prolonged administration of verapamil on the left ventricular geometry and diastolic function in patients with anterior acute myocardial infarction treated with thrombolysis. Patients with heart failure or ejection fraction < 45% were excluded. Within 12 hours from starting thrombolysis, 70 patients were given verapamil (5 mg/hour intravenously for the first 24 hours, followed by 120 mg t.i.d. perorally for 6 months) or equivalent placebo. Echocardiograms were performed on admittance, before discharge, after 3 months and 6 months. The following parameters were calculated: left ventricular volumes, ejection fraction, sphericity index, early (E) and late (A) transmitral peak flow velocities and time-velocity integrals with their ratios, deceleration time and half-time of E, isovolumic relaxation time (IVRT), and non-invasive time constant of ventricular relaxation (tau). The basal and the last available parameters were considered for statistical analysis. The effects of the treatment on the left ventricular volumes, ejection fraction, and sphericity index were not statistically relevant. Conversely, a reduction of E/A ratio (P < .05) and increases of A integral (P < .01), deceleration time and half-time of E, IVRT and tau (P < .05) were found in the placebo group and not in the verapamil group. No significant changes in the blood pressure, heart rate, PQ interval, and biochemical parameters were observed in the two groups. In conclusion, in patients with a thrombolysed anterior acute myocardial infarction and preserved systolic function, verapamil can prevent alterations of the diastolic function in absence of effect on ventricular remodelling, and has a good safety profile. PMID- 10516869 TI - Effects of the Ikr-blocker almokalant and predictors of conversion of chronic atrial tachyarrhythmias to sinus rhythm. A prospective study. AB - PURPOSE: To assess the efficacy of the Ikr-blocker almokalant attempting to convert chronic atrial tachyarrhythmias, and to find predictors of conversion, to sinus rhythm. METHODS: The electrophysiological effects of a 6-hour infusion of almokalant, to a total dose of 25 +/- 4 mg, were assessed by ECG and transesophageal atrial electrograms (TAE) in 100 consecutive patients with atrial fibrillation/flutter (n = 95/5) of 8 +/- 12 months' duration (range 1 to 99 months). RESULTS: The conversion rate was 32%. The time to conversion was 3.5 +/- 2.2 hours. During infusion increases in QTtop (292 +/- 35 to 335 +/- 44 ms, p < 0.001, after 30 minutes), QT (387 +/- 40 to 446 +/- 60 ms, p < 0.001), corrected QT (425 +/- 30 to 487 +/- 44 ms, p < 0.001), and QT dispersion (21 +/- 12 to 29 +/- 31 ms, p = 0.02), were paralleled by decreases in T wave amplitude (0.31 +/- 0.19 to 0.23 +/- 0.16 mV, p < 0.001), and atrial rate (425 +/- 78 to 284 +/- 44 beats per minute (bpm) on ECG, and 396 +/- 72 to 309 +/- 44 bpm on TAE), with no differences between converters to sinus rhythm and non-converters. Patients with aberrantly conducted beats, and T wave variation, also increased. Calcium antagonists were more common among converters. A decreasing T wave amplitude predicted conversion. Four patients developed torsades de pointes. CONCLUSIONS: This study demonstrates class III action of almokalant, with a conversion rate of 32% of long-standing, chronic atrial tachyarrhytmias. An early decrease in T wave amplitude was associated with conversion to sinus rhythm. PMID- 10516870 TI - Effect of antihypertensive therapy on aortic distensibility in patients with essential hypertension: comparison with trichlormethiazide, nicardipine and alacepril. AB - To assess the effect of antihypertensive drugs on aortic distensibility, we evaluated the aortic distensibility of 33 hypertensive patients before and after antihypertensive treatment by using cine magnetic resonance imaging. Thirty three hypertensive patients were divided into three groups and treated for 12 weeks with 2-4 mg trichlormethiazide per day (n = 10), 80 mg nicardipine per day (n = 13) and 50 mg alacepril per day (n = 10). There were no significant differences in mean age and mean blood pressure among the three groups. Cine magnetic resonance was performed at ascending and descending aortic levels. Aortic area was measured at the maximum and minimum frames. The effect of antihypertensive therapy on aortic distensibility was evaluated as the percent change from before treatment to after treatment. There were no significant differences in pulse pressure before and after treatment with trichlormethiazide, nicardipine and alacepril. After treatment with these drugs, mean blood pressure in all groups decreased (trichlormethiazide and nicardipine, P < .01; alacepril, P < .05), (the maximum area--the minimum area) and aortic distensibility in all groups increased significantly (each P < .01). Percent changes in aortic distensibility after treatment were significantly higher with nicardipine (ascending, 346.6 +/- 255.9%; descending, 338.8 +/- 246.5%, each P < .05) and alacepril (ascending, 369.7 +/- 238.8%, P < .05; descending, 306.9 +/- 123.3%, P < .01) than with trichlormethiazide (ascending, 146.0 +/- 139.6%; descending, 129.3 +/- 97.5%). In conclusion, nicardipine and alacepril have a beneficial effect on aortic distensibility. PMID- 10516871 TI - The angiotensin II AT1 receptor antagonist candesartan at antihypertensive plasma concentrations reduces damage induced by ischemia-reperfusion. AB - The aim of this study was to investigate if the angiotensin II AT1 receptor antagonist candesartan in antihypertensive plasma concentrations improves myocardial function and limits infarct size in anesthetized pigs. Animals were subjected to 45 min of regional ischemia and 240 min of reperfusion. Starting 60 min before ischemia, two groups of pigs (n = 6 in each) received either candesartan (25 micrograms/kg bolus followed by a continuous infusion at a rate of 14 micrograms/kg/h) or the corresponding volume of vehicle throughout the study period. Left ventricular systolic segment shortening (%SS) was measured by sonomicrometry, and infarct size was determined by triphenyl tetrazolium chloride staining. The plasma concentration of candesartan during the experiment was between 100 and 150 nmol/L, which was considered to be within the therapeutic range. Neither candesartan nor vehicle affected hemodynamics or coronary blood flow prior to ischemia. Compared to vehicle, candesartan improved recovery of %SS in the ischemic area. At 240 min of reperfusion, the %SS was significantly higher in pigs given candesartan than in pigs given vehicle (7.1 +/- 0.87% vs-1 +/- 1.79%; p < 0.01). In both groups the area at risk was approximately 20% of the left ventricle. Infarct size as a percentage of the area at risk was significantly smaller in the candesartan group than in the vehicle group (46 +/- 3.0 vs 73 +/- 3.6%; p < 0.01). The results suggest that angiotensin II AT1 receptor blockade, obtained in antihypertensive plasma concentrations, supports myocardial functional recovery and limits infarct size. PMID- 10516872 TI - Ambulatory norepinephrine infusion in severe idiopathic orthostatic hypotension (Bradbury Eggleston syndrome) PMID- 10516873 TI - Minimal effects of levosimendan on coronary artery smooth muscle tone. PMID- 10516874 TI - Toxic leukoencephalopathy. AB - The white matter of the brain is vulnerable to a wide variety of toxins. Leukoencephalopathy is being increasingly recognized in a number of different patient populations. The detection of early and subtle toxin effects has been facilitated by the advent of magnetic resonance imaging, which offers better resolution of white matter than other neuroimaging methods. Neuropathologic features of leukoencephalopathy are also becoming more completely elucidated. Injury to white matter has been described from cranial irradiation and cancer chemotherapeutic drugs, and from a number of other therapeutic agents, drugs of abuse, and environmental toxins. Many patients have reversible leukoencephalopathy, whereas others experience a progressive and irreversible course. Leukoencephalopathy is associated with neurobehavioral manifestations that may be subtle or devastating, and the syndrome of white matter dementia may result. The pathogenesis of toxic leukoencephalopathy remains largely unknown, and treatment is limited in most cases to prevention by avoidance or minimization of the toxin exposure. However, the prognosis for this syndrome may be relatively favorable because of the frequent sparing of axons even when myelin is affected. Toxic leukoencephalopathy is an emerging clinical disorder that presents the opportunity for improving clinical outcomes in a number of patient groups and for achieving a deeper understanding of the role of white matter in cognitive and emotional function. PMID- 10516875 TI - Midazolam treatment of acute and refractory status epilepticus. AB - Generalized convulsive status epilepticus (GCSE) is a medical emergency requiring prompt resolution. Acute treatment is often delayed by difficulty in obtaining intravenous (i.v.) access. Refractory GCSE is often difficult to treat, and traditional therapy with barbiturates induces hypotension and respiratory depression and prolongs recovery. Midazolam is particularly useful for treating acute GCSE because it has an imidazole ring that is open at low pH, allowing it to be dissolved in aqueous solution for intramuscular injection, but closed at physiologic pH, increasing lipophilicity and rendering good intramuscular absorption, brain penetration, and fast onset of action. When given intramuscularly as a 0.2 mg/kg bolus, it has efficacy at least equal to that of i.v. diazepam, is well tolerated, induces little respiratory compromise, and has a shorter latency to onset of action. Therefore, it should be considered for the treatment of acute GCSE when i.v. access is problematic. For refractory GCSE, continuous i.v. midazolam infusion at 0.1-0.6 mg/kg/hr after a 0.2 mg/kg i.v. bolus is effective and has advantages over traditional therapies because it induces less hypotension and cardiorespiratory depression and can be easily titrated. Further prospective studies are needed to define the role of continuous i.v. midazolam compared to other contemporary therapies. PMID- 10516876 TI - Phenytoin-folic acid interaction: a lesson to be learned. AB - A case of a patient who developed symptomatic phenytoin-induced folic acid deficiency is reported. Folate supplementation of 5 mg/d was followed by a decrease of serum phenytoin concentration to a subtherapeutic level with a breakthrough seizure. Estimation of phenytoin's Km-Vmax Michaelis-Menten pharmacokinetic parameters in this patient demonstrated that folate supplements indeed caused a significant decrease in the Km value. This decrease correlates with a greater affinity of the metabolizing hepatic enzymes for the drug, and hence, with the resultant increase in phenytoin's metabolism and decrease of its serum concentration and anticonvulsive effect. In an era of increasing knowledge of folate's pivotal role in various diseases, we call attention to this drug vitamin interaction, and to the previously suggested recommendation that folate supplementation should be initiated whenever phenytoin therapy commences. Because folic acid dosages as low as 1 mg/d may perturbate phenytoin's metabolism, smaller deficiency preventive doses may be the advisable allowance for phenytointreated patients with normal pretreatment folate levels. This suggestion must be confirmed by a prospective study in a large cohort of patients. PMID- 10516877 TI - Effect of memantine (NMDA antagonist) on Parkinson's disease: a double-blind crossover randomized study. AB - Our aim was to evaluate the effect of Memantine (1-amino 3,5-dimethyl-adamantane hydrochloride) on cardinal symptoms of Parkinson's disease and on the latency, duration, and magnitude of the response to a single dose of L-Dopa and on drug induced dyskinesias. Twelve Hoehn-Yahr III-IV patients with idiopathic Parkinson's disease with motor fluctuations and drug-induced dyskinesias were randomized to the NMDA antagonist memantine or placebo in a cross-over design. A single-dose L-Dopa challenge was performed after each medication arm. A significant drug effect on the Unified Parkinson's Disease Rating Scale motor score was observed in "off" and "on" states (F(1,11) = 13.5; p < 0.003). No significant effect on drug-induced dyskinesias was seen. The results suggest that memantine may improve parkinsonian symptoms independently of dopaminergic drugs and, in contrast to recent findings with amantadine, it has no effect on drug induced dyskinesias. PMID- 10516878 TI - Modification of dopamine D2 receptor activity by pergolide in Parkinson's disease: an in vivo study by PET. AB - It is well known that chronic administration of pergolide and other dopamine agonists may induce a downregulation of dopamine D2 receptors in the rat model of Parkinson's disease (PD). To our knowledge, this effect has not been demonstrated in vivo in patients with PD. At present, the status of striatal dopamine D2 receptors can be studied with use of positron emission tomographic (PET) technology. Five patients with PD chronically treated with levodopa were studied with use of PET and [11C]-raclopride before and after 6 months of pergolide treatment (dose range = 4.5-7.5 mg/d). We found a slight reduction in the specific striatal [11C]-raclopride uptake index (mean reduction 14% in putamen and 9% in caudate) after pergolide treatment. This reduction appears to be related to downregulation of the receptor, although competitive binding of pergolide at the D2 receptor cannot be excluded. PMID- 10516879 TI - Effect of the selective D1 antagonists SCH 23390 and NNC 01-0112 on the delay, duration, and improvement of behavioral responses to dopaminergic agents in MPTP treated monkeys. AB - We assessed the antiparkinsonian response in MPTP-treated monkeys after acute or repeated treatment with oral L-Dopa, subcutaneous administration of L-Dopa methyl ester (LDME) or apomorphine, alone and in combination with D1 antagonists SCH 23390 (SCH) or NNC 01-0112 (NNC). When given alone, the L-Dopa effect occurred within the first hour after treatment. Coadministration of SCH or NNC with L-Dopa significantly delayed the onset of action. The response duration remained unchanged, as did the extent of the antiparkinsonian effect, after SCH, whereas the former became shorter at the higher doses of NNC tested. Bypass of the gastrointestinal tract using parenteral injections of LDME and apomorphine allowed the rapid turning "on" of the animals. Both D1 antagonists administered with LDME delayed the onset and shortened the duration of the therapeutic effect as the dose increased. Pretreatment with SCH failed to block the antiparkinsonian effect induced by apomorphine, but reduced the response duration markedly in a dose-related fashion. Repeated treatment of one monkey with SCH combined with the same dopaminergic drugs produced results similar to those obtained after acute treatment in four animals. The results obtained with parenteral administration of LDME and apomorphine most probably involve pharmacodynamic actions resulting in increased threshold of response. The delay observed with L-Dopa suggests pharmacokinetic interference possibly mediated via dopamine receptors located at the level of the gut. PMID- 10516880 TI - Fulminant CNS perivascular lymphocytic proliferation: association with sargramostim, a hematopoietic growth factor. AB - Sargramostim (GM-CSF) therapy was instituted in a 49-year-old woman with hepatitis C on chronic interferon alpha-2b therapy. Within two weeks, she developed progressive confusion, lethargy, and gait disturbance. At autopsy 4 months later, diffuse perivascular nonmonoclonal lymphoid infiltrates were demonstrated throughout the central nervous system (CNS). As the use of hematopoietic growth factors in clinical practice increases, potential adverse effects, such as the fulminant CNS lymphocytic proliferation in this patient, are more likely to be encountered. PMID- 10516881 TI - "Apraxia of eyelid opening" induced by levodopa therapy and apomorphine in atypical parkinsonism (possible progressive supranuclear palsy): a case report. AB - We report a female patient in whom so-called apraxia of eyelid opening (AEO) developed after the onset of possible progressive supranuclear palsy (National Institute of Neurological Disorders and Stroke criteria) and the introduction of antiparkinsonian medications including levodopa. Although parkinsonian symptoms responded poorly to levodopa, AEO worsened after increasing levodopa dosage and disappeared when levodopa was discontinued. Later, a dose of apomorphine widely accepted for acute tests had no significant effect on limb motor activity but induced AEO. Overall, these observations are grounds for thinking that AEO developing in the course of parkinsonism may be either disease- or drug-related. The possibility of manipulating dopaminergic treatment should always be considered when dealing with AEO associated with parkinsonism. PMID- 10516882 TI - Rhabdomyolysis induced by simvastatin and ketoconazole treatment. AB - Rhabdomyolysis is described as an adverse event of simvastatin therapy either by itself or in combination with other medications. It is unclear whether this phenomenon is specific to simvastatin or to all cholesterol-lowering agents as single-dose therapy or caused by the association of special coadministered medications. We describe two cases in which rhabdomyolysis developed after coadministration of simvastatin (20 mg/d) and the antifungal ketoconazole. The clinical features, blood examination results, and positive outcome were very similar in both cases. We concluded that ketoconazole, an antifungal sterol synthetic inhibitor of the azol group, may induce rhabdomyolysis in patients undergoing treatment with simvastatin, a lipid lowering agent, and increase the possibility of muscle-damaging adverse events of the agents. PMID- 10516883 TI - Trimipramine fails to exert antimanic efficacy: a case of the discrepancy between in vitro rationale and clinical efficacy. AB - Standard mood stabilizers, such as lithium and haloperidol, and anticonvulsants show effectiveness in a maximum of 60%-70% of acutely manic patients. Obviously, there is a clinical need to evaluate other treatment options. Current pathophysiologic concepts suggest that substances with an ameliorating effect on dopaminergic hyperfunction, serotonergic hypofunction, or GABAergic hypofunction might be useful, as may be substances with calcium-antagonistic effects. In vitro, the antidepressant trimipramine exerts dopamine- and calcium-antagonistic properties. Therefore, we conducted an open trial to screen it for antimanic action. We found no clinical benefit in four acutely manic patients receiving up to 400 mg/d of trimipramine. It is concluded that, at least in the case of trimipramine, the pharmacologic profile is not helpful in predicting potential effectiveness in mania. PMID- 10516884 TI - Pramipexole, a nonergot dopamine agonist, is effective against rest tremor in intermediate to advanced Parkinson's disease. AB - We evaluated the efficacy of the nonergot dopamine receptor agonist pramipexole in 16 patients with advanced Parkinson's disease and marked rest tremor during an "on" period. The patients were drawn from a larger placebo-controlled, double blind, randomized trial, which was not originally designed to investigate the effect of pramipexole on tremor. Eleven patients received pramipexole. The first effects were seen with a pramipexole dose of 0.75 mg/d with a reduction of the tremor item A of Unified Parkinson's Disease Rating Scale (UPDRS III, "on" state) by 25% and of rigidity and akinesia by 22%. Under the highest dose, 4.5 mg/d, the tremor score was improved by 61% over baseline (p < 0.0056, Wilcoxon signed rank) and the sum of rigidity and akinesia items by 66% (p < 0.0038, Wilcoxon signed rank). Five patients received placebo and did not improve. Based on this sample of patients, the nonergot dopamine receptor agonist pramipexole appears to have a potent anti-rest tremor action while being effective against akinesia and rigidity. PMID- 10516885 TI - Blood perfusion hyperaemia in response to graded loading of human heels assessed by laser-Doppler imaging. AB - Heel pressure ulcers are important clinical, humanitarian and economic problems arising in part from localized blood flow deficits during loading and inadequate flow recovery. Because there are few data available with regard to the intrinsic physiological responses of heel skin to pressure-induced ischaemia, the present study was undertaken to characterize the main features of the post-loading hyperaemic response. Laser-Doppler perfusion imaging was used to measure hyperaemia in 14 vascularly normal women who were subjected to sequential local heel loading with graded magnitudes (30-140 mmHg) and durations (2.5-20 min). Peak heel perfusion produced by local heating to 44 degrees C for 5 min was used as a comparison standard. All heel loads and durations resulted in hyperaemic responses, with the largest increase in peak response occurring between heel loads of 60 and 120 mmHg. During this transition, peak hyperaemia increased from about 32% to 79% of the local maximal microvascular vasodilatory capacity. Recovery times also increased with both load duration and magnitude, with the longest recovery time being about 7.5 min. Hyperaemic responses and recovery times were analytically dependent on the heel load pressure duration product, with evidence of suppression of the peak response at 1500 mmHg min and a levelling off of recovery time at higher pressure durations. These findings serve to characterize normal physiological perfusion responses to pressure-induced ischaemia at an anatomical site prone to pressure ulceration. The results suggest the possibility of a 'critical' heel loading, above which a near-maximum response is elicited and beyond which vasodilatory recovery potential is blunted. PMID- 10516886 TI - Contributory factors to orthostatic intolerance after simulated weightlessness. AB - Various factors may contribute to orthostatic intolerance (OI) observed after space flights or simulated weightlessness such as bed rest experiments: individual physical and physiological factors (arterial blood pressure (BP), height), physiological changes induced by real or simulated weightlessness (hypovolaemia, increase in venous distensibility), and space flight or simulation conditions (duration and counter-measure application). Our purpose was to test which of these factors were dominant in contributing to the OI. This was assessed in 47 healthy men participating in bed rest experiments of 4, 14, 28, 30 and 42 days, with or without counter-measures (medical stockings, lower-body negative pressure (LBNP), LBNP + muscular exercise). Nineteen subjects did not finish the orthostatic test (60 degrees head-up tilt or stand test) after bed rest. The occurrence of OI was associated with greater height, low resting BP, greater changes in resting lower-limb venous distensibility throughout the bed rest, and absence of counter-measures. PMID- 10516887 TI - Determination of baroreflex gain using auto-regressive moving-average analysis during spontaneous breathing. AB - The heart rate component of the arterial baroreflex gain (BRG) was determined with auto-regressive moving-average (ARMA) analysis during each of spontaneous (SB) and random breathing (RB) protocols. Ten healthy subjects completed each breathing pattern on two different days in each of two different body positions, supine (SUP) and head-up tilt (HUT). The R-R interval, systolic arterial pressure (SAP) and instantaneous lung volume were recorded continuously. BRG was estimated from the ARMA impulse response relationship of R-R interval to SAP and from the spontaneous sequence method. The results indicated that both the ARMA and spontaneous sequence methods were reproducible (r = 0.76 and r = 0.85, respectively). As expected, BRG was significantly less in the HUT compared to SUP position for both ARMA (mean +/- SEM; 3.5 +/- 0.3 versus 11.2 +/- 1.4 ms mmHg-1; P < 0.01) and spontaneous sequence analysis (10.3 +/- 0.8 versus 31.5 +/- 2.3 ms mmHg-1; P < 0.001). However, no significant difference was found between BRG during RB and SB protocols for either ARMA (7.9 +/- 1.4 versus 6.7 +/- 0.8 ms mmHg-1; P = 0.27) or spontaneous sequence methods (21.8 +/- 2.7 versus 20.0 +/- 2.1 ms mmHg-1; P = 0.24). BRG was correlated during RB and SB protocols (r = 0.80; P < 0.0001). ARMA and spontaneous BRG estimates were correlated (r = 0.79; P < 0.0001), with spontaneous sequence values being consistently larger (P < 0.0001). In conclusion, we have shown that ARMA-derived BRG values are reproducible and that they can be determined during SB conditions, making the ARMA method appropriate for use in a wider range of patients. PMID- 10516888 TI - Determinants of post-ischaemic reactive hyperaemia in patients with diabetes mellitus type II. AB - Dysfunction of resistance arteries is thought to be an early reversible stage in the development of atherosclerosis. Dynamics of post-ischaemic reactive hyperaemia are believed to constitute a useful tool for monitoring resistance vessel function. Patient characteristics influencing reactive hyperaemia, however, need to be defined more precisely. Since reactive hyperaemia is a dynamic process, yielding submaximal peak values after 5 min of ischaemia, this period was chosen to investigate the determinants of reactive hyperaemia in 100 type II diabetic patients as well as in 61 control subjects. Reactive hyperaemia was measured by venous-occlusion plethysmography; clinical and laboratory data were acquired by routine methods. Statistical comparison was performed with SYSTAT 5.0 for Apple Macintosh. Overall, no significant differences between diabetic patients and controls were observed by group comparison. In control subjects, only gender showed an influence on peak reactive hyperaemia (females 40.5 +/- 15.3; males 51.8 +/- 17.7 ml min-1 100 ml-1, P < 0.01). In diabetic patients, in addition to gender, actual blood glucose (r = 0.377, P < 0.05) and meal intake (non-fasting 42.8 +/- 19.2; fasting 51.2 +/- 19.5 ml min-1 100 ml-1, P < 0.05) were found to influence reactive hyperaemia. Further investigation revealed a loss of the correlation between peak reactive hyperaemia and actual blood glucose observed in the fasting state (P < 0.001) in non-fasting diabetic patients, indicating an influence of meal intake on resistance vessel reactivity. Our results suggest that, in diabetic subjects, in addition to gender actual blood glucose and the postprandial situation impacts on peak reactive hyperaemia. PMID- 10516889 TI - Atrophy of the quadriceps muscle in children with a painful hip. AB - The objective of this study was to determine the degree of muscle wasting of various components of the quadriceps muscle in children with a painful hip. Between January 1994 and September 1997, 327 consecutive children with a unilateral painful hip and/or limping were evaluated prospectively with ultrasonography. Quadriceps thickness was measured on both sides. Moreover, muscle thickness was measured in 59 control subjects. The patients were divided into eight groups; transient synovitis (n = 134), Perthes' disease (n = 35), slipped capital femoral epiphysis (n = 5), osteomyelitis (n = 4), aspecific synovitis (n = 5), rheumatoid arthritis (n = 3) and miscellaneous (n = 16). In 125 patients, no sonographic and radiological abnormalities were found and during follow-up the symptoms disappeared ('no pathology' group). Ipsilateral muscle wasting was present in all patient groups, whereas the control subjects showed no significant difference in muscle thickness between legs. The degree of muscle wasting was compared between transient synovitis, the 'no pathology' group, Perthes' disease and control subjects. For both quadriceps and vastus intermedius muscles, there was a significant difference between these groups, except between control subjects and the 'no pathology' group. For the rectus femoris muscle, there was a significant difference between these groups, except between transient synovitis and 'no pathology'. Muscle wasting showed a positive correlation with duration of symptoms and pre-existing muscle mass. In conclusion, different diseases show different degrees of muscle wasting, and there are different patterns of muscle wasting of various components of the quadriceps femoris muscle. PMID- 10516890 TI - Delayed respiratory response to exercise of short duration in patients with severe intermittent claudication due to bilateral atherosclerotic vascular disease: normalization after aorto-bifemoral bypass operation. AB - To investigate the respiratory response to exercise in patients with severe intermittent claudication, eight male patients, aged 57 years (range 43-73), with bilateral multi-segment atherosclerotic vascular disease, median maximum walking distance 50 m (range 20-200) and ankle-to-arm pressure index 0.4 (range 0.3-0.6), were studied before and after aorto-bifemoral bypass operation. Ventilation, CO2 output and O2 intake were recorded in the sitting position during 20 min of rest, 1 min of leg exercise on a bicycle ergometer [4.9 kJ (500 kpm)], and 20 min of recovery and rest. Before operation, maximal ventilation and CO2 output per minute were observed 2-4 min after cessation of work, while afterwards peak values were found during the work or the first minute of recovery. Pre operatively, the extra ventilation and CO2 output during the work and recovery period and the recovery times of the ventilation and CO2 output per minute were markedly increased. Afterwards these values were clearly reduced towards normal. It is concluded that patients with severe intermittent claudication show a characteristic delay and prolonged rise in the respiratory response to exercise of short duration, which closely corresponds to the previously described pattern of outflow of hypoxia-generated metabolites from the exercising muscles. The pattern of respiratory response after operation reflects the fact that these patients also suffer from atherosclerotic heart dysfunction. PMID- 10516891 TI - Endothelium-dependent vasodilation and structural and functional changes in the cardiovascular system are dependent on age in healthy subjects. AB - The aim of this study was to evaluate possible associations between endothelium dependent vasodilatation (EDV) and cardiovascular structure and function. EDV could influence peripheral resistance and be affected by atherosclerosis and might thereby influence indices of cardiovascular structure and function. In a group of 31 apparently healthy men and 25 women (age range 20-69 years), EDV was evaluated by infusion of metacholine (4 micrograms min-1), and endothelium independent vasodilatation (EIDV) was assessed by nitroprusside infusion (SNP, 10 micrograms min-1) in the brachial artery. Forearm blood flow (FBF) was measured by venous occlusion plethysmography. Left ventricular (LV) geometry and function and the intima-media thickness in the carotid artery were assessed by ultrasonography. The stroke index to pulse pressure ratio was used to evaluate arterial compliance. Several indices of cardiovascular structure and function were found to be related to an index of endothelial function, the EDV to EIDV ratio. Furthermore, left ventricular mass (LVM), the atrio-ventricular plane displacement, E/A ratio, IVRT, the intima-media thickness of the carotid artery and arterial compliance were all significantly related to both EDV and EIDV in women. However, most indices of cardiovascular structure and function, as well as endothelial function, change with age and only the relation between LV diastolic function and endothelial function in men remained significant (P < 0.05) after including age in multiple regression analysis. Age was related to both cardiovascular structure and function, as well as to endothelial function. Multiple regression analysis showed that ageing generally affects cardiovascular characteristics and endothelial function in parallel in these healthy subjects. PMID- 10516892 TI - A confident decision support system for interpreting electrocardiograms. AB - Computer-aided interpretation of electrocardiograms (ECGs) is widespread but many physicians hesitate to rely on the computer, because the advice is presented without information about the confidence of the advice. The purpose of this work was to develop a method to validate the advice of a computer by estimating the error of an artificial neural network output. A total of 1249 ECGs, recorded with computerized electrocardiographs, on patients who had undergone diagnostic cardiac catheterization were studied. The material consisted of two groups, 414 patients with and 835 without anterior myocardial infarction. The material was randomly divided into three data sets. The first set was used to train an artificial neural network for the diagnosis of anterior infarction. The second data set was used to calculate the error of the network outputs. The last data set was used to test the network performance and to estimate the error of the network outputs. The performance of the neural network, measured as the area under the receiver operating characteristic (ROC) curve, was 0.887 (0.845-0.922). The 25% test ECGs with the lowest error estimates had an area under the ROC curve as high as 0.995 (0.982-1.000), i.e. almost all of these ECGs were correctly classified. Neural networks can therefore be trained to diagnose myocardial infarction and to signal when the advice is given with great confidence or when it should be considered more carefully. This method increases the possibility that artificial neural networks will be accepted as reliable decision support systems in clinical practice. PMID- 10516893 TI - Hip fracture discrimination by imaging ultrasound measurements of the calcaneus. AB - Quantitative ultrasound measurements of the os calcis have recently been upgraded with imaging facilities. This has made measurements of a specific region of interest possible and improved the reproducibility of the method, but the diagnostic ability of imaging ultrasound has not yet been investigated thoroughly. We measured broadband ultrasound attenuation (BUA) and speed of sound (SOS) using imaging ultrasound as well as forearm bone mineral density (BMDarm) using dual-energy X-ray absorptiometry in three age-matched groups of women: (1) 25 women who were admitted to hospital due to a hip fracture; (2) 23 women who were admitted to hospital due to a fall without any fracture; and (3) 26 normal women. Furthermore, BMD of the hip (BMDhip) was measured in a subgroup of the hip fracture patients and those who had fallen. All measurements were performed during the index hospitalization in order to avoid any influence from bone loss due to immobilization after the fracture. We found a -0.48 SD deviation from expected age-matched values in BUA among the hip fracture patients, whereas the patients who had fallen showed a +0.16 SD deviance. For SOS, the figures were 0.70 SD for the hip fracture group and -0.06 SD for the patients who had fallen. For BMD of the arm we found values of -0.65 SD and +0.08 SD, respectively, whereas the figures for BMD of the hip were -0.66 SD and +0.13 SD, respectively. All parameters were significantly lower in the hip fracture group compared with the patients who had fallen. None of the parameters in the patients who had fallen deviated significantly from expected normal age-matched values. Neither BMD of the arm or BMD of the hip separated hip fracture patients and patients who had fallen significantly better than ultrasound (BUA or SOS) did. We conclude that imaging ultrasound (BUA or SOS) separates age-matched groups of hip fracture and non-fracture patients as well as BMD measurements do. PMID- 10516894 TI - Changes in airway dead space in response to methacholine provocation in normal subjects. AB - Measurements of bronchial hyper-responsiveness rely on sensitive techniques for measurement of bronchoconstriction, ideally based on tidal breathing. A potentially useful technique is measurement of airway dead space (VDaw), which reflects the volume of the conducting airways. The aim of this study was to evaluate measurements of VDaw with the single breath test for CO2 (SBT-CO2), compared to spirometric measurements, as a method of measuring bronchial response to methacholine challenge. Nineteen healthy adults were studied. Dosimetric methacholine challenge tests were performed on two study days. Forced expirations or the SBT-CO2 were used to assess the response. There were dose-dependent reductions in the spirometric measurements, with a 10 +/- 10% reduction from the baseline value of forced expiratory volume at the highest dose of methacholine. There was a dose-dependent reduction from the baseline value of VDaw by 19 +/- 9% at the highest dose. There was also a dose-dependent increase in the slope of the alveolar plateau of the SBT-CO2. This study provides support for measurement of VDaw as a means of evaluating bronchial responsiveness after methacholine challenge. In a group of healthy adults, this method shows a greater response but with similar dispersion as measurement of forced expiratory volume after methacholine challenge. PMID- 10516895 TI - Reproducibility of surface EMG during dynamic shoulder forward flexions: a study of clinically healthy subjects. AB - There is a shortage of studies concerning the reproducibility of surface electromyography (EMG) during dynamic contractions. The movement per se introduces factors that could have the potential to affect the characteristics of the electromyogram. The aim of this study was to investigate the during-the-day reproducibility (using intra-class coefficient, ICC) of the peak torque (PT) and the EMG variables (without removing electrodes) of dynamic shoulder forward flexions. Eleven healthy women performed three sets of 10 dynamic maximum right shoulder flexions at 1 h intervals using an isokinetic dynamometer. The PT of each flexion was determined. EMG signals were recorded from four muscles (trapezius, deltoid, infraspinatus and biceps brachii) using surface electrodes, and the mean frequency of the power spectrum (MNF [Hz]) and the signal amplitude (RMS [microV]), were computed. The ability to relax between maximum flexions was calculated as a ratio between the RMS during the passive extension phase and the RMS during the active flexion phase of each contraction cycle. This ratio is the signal amplitude ratio (SAR). The present study showed good reproducibility for PT, MNF and RMS, while the reproducibility of SAR was generally acceptable (fair) according to the criteria used. PMID- 10516896 TI - Different plasma levels of nitric oxide in arterial and venous blood. AB - Experimental investigations suggest that a basal release of nitric oxide (NO) occurs in arterial but not in venous endothelium. We therefore decided to compare plasma levels of NO in the arterial and venous circulation. Parallel blood samples were drawn from the radial artery and brachial vein in 15 healthy drug free women. Nitric oxide levels were assessed by measuring plasma levels of nitrite and nitrate, the two stable oxidation products of NO metabolism. Plasma levels of NO metabolites in arterial blood were significantly higher than in the paired venous blood samples (45.1 +/- 17.7 versus 22.5 +/- 8.5 mumol l-1, respectively, mean +/- SD). The results of this preliminary study strongly suggest that the endothelial release of NO is probably different in arteries and veins in vivo; this is also consistent with previous literature indicating that basal release of NO occurs mainly from the endothelium of arteries but not from that of veins. PMID- 10516897 TI - Pulmonary host defenses. Implications for therapy. AB - Pulmonary host defenses comprise a redundant system of protective mechanisms against invasion of the lungs by pathogenic microbes. The upper and lower airways are uniquely suited to contain and remove organisms that gain access to the respiratory mucosa. If the balance between host and organism is disputed, however, microbial clearance may be ineffective, and infection established. Pulmonary host defense mechanisms, which provide the basis for several current therapeutic strategies, are reviewed. PMID- 10516898 TI - Atypical pathogens in community-acquired pneumonia. AB - The atypical pathogens are an important and significant cause of CAP. The clinical and radiologic manifestations of CAP caused by these pathogens are modulated by the immunologic and physiologic status of the host, and therefore are not pathogen-specific. The range of frequencies found in various studies for the atypical pathogens among the causes of CAP is broad. These frequencies are affected by very important factors that should be recognized. In a significant percentage of patients, an atypical pathogen can be identified together with an additional cause. The significance of multiple causes has not been clarified sufficiently. The principal diagnostic techniques in use today for the causative diagnosis of CAP are serologic tests. Different serologic methods have been used in various studies and diagnostic criteria are not standardized. In the future it is likely that diagnostic testing will be based on the PCR technique on serum samples. The effectiveness and importance of antimicrobial therapy in some patients with atypical pathogen CAP are unclear. The accepted therapy today for atypical pathogen CAP, which is based on erythromycin, will probably be changed in the near future in favor of the new generations of fluoroquinolone or the new macrolide preparations. PMID- 10516899 TI - Drug-resistant pathogens in community- and hospital-acquired pneumonia. AB - Antimicrobial resistance has been a problem since the early days of the antibiotic era, but in recent years, this resistance has increased in the hospital and is being recognized more in the community setting. Respiratory pathogens such as S. pneumoniae and H. influenzae, for example, have developed resistance to traditional antimicrobial therapy, often over a very short period of time. This increase in resistance patterns requires physicians to closely monitor antimicrobial resistance in their community and to appreciate that some antimicrobial resistance mechanisms may result in resistance for a complete class of antibiotics or different classes of antibiotics with similar mechanisms of action. PMID- 10516901 TI - Use of prognostic scoring and outcome assessment tools in the admission decision for community-acquired pneumonia. AB - Prognostic scoring and outcome assessment tools are being developed to provide decision support regarding hospitalization in community-acquired pneumonia. The tools define the risk of mortality and other adverse outcomes using variables available when patients are first seen. Comparison of the specific logic in different tools demonstrates more similarities than differences. Early data suggest that these tools may help physicians safety decrease the rate of admission to the hospital. PMID- 10516900 TI - Fungal pneumonias. The endemic mycoses. AB - Each year, a vast number of individuals are infected with the endemic fungi. An expanding population, along with further land development in endemic areas, will likely continue to place individuals at risk for exposure to these organisms. A high index of suspicion may be required to diagnose histoplasmosis, blastomycosis, or coccidioidomycosis, particularly for patients who do not reside in endemic areas. Although the majority of patients with histoplasmosis, blastomycosis, and coccidioidomycosis experience self-limited infections, treatment is necessary for patients with severe pneumonitis as well as various forms of chronic pulmonary and disseminated infections. The newer azole agents- itraconazole and fluconazole--are useful in the treatment of these infections and have provided alternatives to long-term therapy with amphotericin B for many patients. PMID- 10516902 TI - Diagnostic testing for community-acquired pneumonia. AB - The microbial cause of community-acquired pneumonia can be identified by noninvasive means in the majority of cases, usually within a few days of presentation. The Gram stain and culture of a pretreatment sputum sample are the most useful tests, but have significant limitations. Methods for detecting pneumococcal antigen in respiratory secretions are particularly helpful in patients who have received antibiotics before evaluation. Testing for specific pathogens such as L. pneumophila, M. pneumoniae, or C. pneumoniae should be guided by clinical suspicion in individual circumstances. Invasive procedures are most helpful in patients suspected of having infection with opportunistic or resistant pathogens, and in those whose initial management has been unsuccessful. PMID- 10516903 TI - Radiology of pneumonia. AB - Chest radiography is the imaging technique of choice in evaluating patients with suspected pneumonia because of its low radiation dose, low cost, and wide accessibility. In daily practice, radiographs are used to confirm the clinical diagnosis of pneumonia, characterize the extent and severity of disease, search for complications such as empyema, monitor the response to therapy, and examine for possible alternative or additional diagnoses. Although CT scan has no defined role in the routine assessment of patients with either community-acquired or nosocomial pneumonias, its advantages of superior contrast resolution and cross sectional display can often be helpful in the analysis of complex cases, particularly when radiographic evidence of associated central obstruction, cavitation, lymphadenopathy, or empyema is equivocal. In the immunocompromised patient population, high-resolution CT has been shown to be more sensitive than plain film radiography in the early detection of pulmonary infections. PMID- 10516904 TI - Pneumonia in the elderly. AB - Pneumonia, including community-acquired, long-term care facility-associated, and nosocomial infections, is a major cause of morbidity and mortality in the elderly. The aged with pneumonia often present with atypical features, including confusion, lethargy, and general deterioration of condition (so-called "silent infection"). Further investigations, such as a chest radiograph, are more frequently required for diagnosis, but even these results may be normal early in the course of infection, particularly in dehydrated patients. The elderly are more frequently hospitalized for pneumonia and have a greater need for intravenous therapy, longer hospital stay, more prolonged course, greater morbidity, and, ultimately, a poorer outcome. Yet in many studies it is not chronological age per se that impacts negatively on the manifestations of pneumonia in the elderly but rather the presence of comorbid illness. Antibiotic therapy remains the mainstay of therapy for pneumonia, and both community and hospital-based studies confirm the important positive impact of early appropriate empiric antibiotic therapy on outcome. Attention to nutrition and hydration, the use of pneumococcal and influenza vaccination, and a number of diverse procedures in the hospital setting may help limit the occurrence and impact of such infections. PMID- 10516906 TI - Antibiotic therapy for community-acquired pneumonia. AB - This article takes a broad perspective of community-acquired pneumonia (CAP). The arguments and data that support or refute the current approaches to initial antimicrobial treatment of CAP as outlined in the American Thoracic Society and Infectious Disease Society of America documents are provided. The complex issues involved in the decision of how to properly treat CAP are addressed. PMID- 10516905 TI - Severe community-acquired pneumonia. AB - Severe CAP is a life-threatening condition defined by the presence of respiratory failure or symptoms of severe sepsis or septic shock. It accounts for approximately 10% of hospitalized patients with CAP. The majority of patients with severe pneumonia have underlying comorbid illnesses, with COPD, alcoholism, chronic heart disease, and diabetes mellitus being the most frequent. S. pneumoniae, Legionella spp, GNEB (especially K. pneumoniae), H. influenzae, S. aureus/spp, Mycoplasma pneumoniae, respiratory viruses (especially influenza viruses), and P. aeruginosa represent the most important causative organisms of severe CAP. Rapid initiation of appropriate antimicrobial treatment is crucial for a favorable outcome. Initial antimicrobial treatment should be based on an epidemiological (empiric) approach. Microbial investigation may be helpful in the individual case but is probably more useful to define local antimicrobial policies based on local epidemiologic and susceptibility patterns. Mortality rates range from 21% to 54%. The most important prognostic factors include general health state of the patient, appropriateness of initial antimicrobial treatment, and the existence of bacteremia, as well as factors reflecting severe respiratory failure, severe sepsis, septic hypotension or shock, and the extent of infiltrates in chest radiograph. Initial antimicrobial treatment should consist of a second (or third) generation cephalosporin and erythromycin. Modifications of this basic regimen should be considered in the presence of distinct comorbid conditions and risk factors for distinct pathogens. Promising new approaches of nonantimicrobial treatment, including noninvasive ventilation, treatment of hypoxemia, and immunomodulation, are under investigation. PMID- 10516907 TI - Strategies for early discharge of the hospitalized patient with community acquired pneumonia. AB - The treatment of the hospitalized patient with uncomplicated CAP is changing, to include a brief period of intravenous antibiotics followed by oral therapy. The Classification of Community-Acquired Pneumonia or CoCAP is a stratification tool that categorizes patients as low-risk pneumonia, unstable pneumonia, or complicated pneumonia. Use of validated hospital admission criteria, combined with the CoCAP algorithm and evolving criteria for switching patients from intravenous to oral therapy provides a structure for organizing treatment of patients with CAP for caregivers. Patients who can be discharged early are those from the unstable pneumonia group, which includes patients who have had reversal of their metabolic problems and stabilization of comorbid conditions, and who have not developed any serious pneumonia-related complications. Prolonged courses of intravenous antibiotic therapy are being replaced with 2 to 3 day courses of intravenous hydration and antibiotics; a switch to oral therapy and hospital discharge can be achieved after the patient tolerates one dose of oral therapy. Parameters to watch include vital signs and white blood cell count. Provided these parameters are improving, although they may not have returned to normal, the patient can be switched to oral therapy. Although patient treatment guidelines and critical pathways are becoming widespread in disease management, CAP is one disease in which prospective studies have demonstrated that a reduction in hospital stay is safe. Patients, caregivers, and administrators are happy with the reduction in hospital LOS. Other treatment protocols are being explored, including a single dose of intravenous antibiotic prior to oral switch and all-oral regimens employing the newer fluoroquinolones. PMID- 10516908 TI - Infection of the pleural space. AB - Parapneumonic effusions are common accompaniments of pneumonia that require proper management to prevent progression to empyema. Management decisions require thoughtful individualization of care because of the multiple factors that affect outcome; no one algorithmic approach exists for all patients. Basic principles of care, however, apply to all patients and center on the early detection of infected pleural fluid and the rapid completion of effective pleural drainage and lung re-expansion, when indicated to decrease morbidity and mortality. PMID- 10516909 TI - Nonresolving or slowly resolving pneumonia. AB - Given the variability in rate of radiographic resolution, it remains controversial to decide when to initiate an invasive diagnostic work-up for nonresolving or slowly resolving pulmonary infiltrates. In immunocompetent patients who present with classical features of CAP (i.e., fever, chills, productive cough, new pulmonary infiltrate), clinical response to therapy is the most important determinant for further diagnostic studies. Within the first few days, persistence or even progression of infiltrates on chest radiographs is not unusual. Defervescence, diminished symptoms, and resolution of leukocytosis strongly support a response to antibiotic therapy, even when chest radiographic abnormalities persist. In this context, observation alone is reasonable, and invasive procedures can be deferred. Serial radiographs and clinical examinations dictate subsequent evaluation. In contrast, when clinical improvement has not occurred and chest radiographs are unchanged or worse, a more aggressive approach is warranted. In this setting, we advise fiberoptic bronchoscopy with BAL and appropriate cultures for bacteria, legionella, fungi, and mycobacteria. When endobronchial anatomy is normal and there is no purulence to suggest infection, TBBs should be done to exclude noninfectious causes (discussed earlier) or infections attributable to mycobacteria or fungi. An aggressive approach is also warranted in patients who are clinically stable or improving when the rate of radiographic resolution is delayed. As discussed earlier, what constitutes excessive delay is controversial, and depends upon the acuity of illness, specific pathogen, extent of involvement (i.e., lobar versus multilobar), comorbidities, and diverse host factors. Stable infiltrates even 2 to 4 weeks after institution of antibiotic therapy does not mandate intervention provided patients are improving clinically. Invasive techniques can also be deferred when unequivocal, albeit incomplete, radiographic resolution can be demonstrated. Lack of at least partial radiographic resolution by 6 weeks, even in asymptomatic patients, however, deserves consideration of alternative causes (e.g., endobronchial obstructing lesions, or noninfectious causes). Fiberoptic bronchoscopy with BAL and TBBs has minimal morbidity and is the preferred initial invasive procedure for detecting endobronchial lesions or substantiating noninfectious causes. The yield of bronchoscopy depends on demographics, radiographic features, and pre-test likelihood. In the absence of specific risk factors, the incidence of obstructing lesions (e.g., bronchogenic carcinomas, bronchial adenomas, obstructive foreign body) is low. Bronchogenic carcinoma is rare in nonsmoking, young (< 50 years) patients but is a legitimate consideration in older patients with a history of tobacco abuse. Non-neoplastic causes (e.g., pulmonary vasculitis, hypersensitivity pneumonia, etc.) should be considered when specific features are present (e.g., hematuria, appropriate epidemiologic exposures). Ancillary serologic tests or biopsies of extrapulmonary sites are invaluable in some cases. In rare instances, surgical (open or VATS) biopsy is necessary to diagnose refractory or non-resolving "pneumonias." PMID- 10516911 TI - Diagnostic testing for ventilator-associated pneumonia. AB - This article discusses the interpretation of the diagnostic tests in the management of ventilated patients with suspicion of pneumonia. The specific steps for diagnostic evaluation are identified. An accurate interpretation of the significance of the bacterial burden requires previous evaluation of the sample quality, knowledge of administration of new antibiotics within the prior 48 hours, and evaluation of presence of comorbidities. Finally, the article presents a review of the current debate of impact on outcome. PMID- 10516910 TI - Epidemiology and risk factors for nosocomial pneumonia. Emphasis on prevention. AB - VAP is a complex nosocomial infection, the disease expression and resulting patient outcome of which is dependent on host factors, the causative organism, the timing and adequacy of treatment, and the presence of intrinsic or inducible antibiotic resistance. Significant improvements have been achieved in our ability to reduce the occurrence of VAP in the hospital setting. Clinicians caring for mechanically ventilated patients should strive to develop focused programs for the prevention of VAP, other nosocomial infections, and the occurrence of antibiotic-resistant infections at their institutions. The benefits of such programs are well demonstrated. The components of a PDSA (Plan-Do-STUDY-Act) model that can be simply employed to develop a VAP prevention program are as follows: Stages Plan: 1. Identify potentially modifiable risk factors for VAP at the institutional level. 2. Develop a strategy to modify or prevent the occurrence of these risk factors. [figure: see text] Do: 1. Carry out the planned intervention strategy. 2. Identify problems in the implementation of the designed intervention. 3. Update the intervention with solutions for the identified problems. 4. Collect basic data (e.g., VAP rates, severity of illness). STUDY: 1. Analyze data. 2. Summarize the results. Act: 1. Determine the overall success or failure of the intervention. 2. Identify potential modifications to improve the intervention strategy. 3. Prepare for next PDSA cycle. Inherent in the development and application of such programs is the concept that they are continuous processes striving to improve clinical performance over time (Fig. 3). At any given institution, the most likely approach to the prevention of NP and VAP will be a multifaceted one, employing interventions aimed at reducing the occurrence of aerodigestive tract colonization with pathogenic bacteria and aspiration. To be successful, such quality improvement programs must be embraced at the institutional level. Only in this way can hospitals hope to successfully reduce their rates of VAP and sustain or improve upon those efforts over time. PMID- 10516912 TI - Therapy for ventilator-associated pneumonia. AB - Pneumonia is a serious complication of mechanical ventilation. Pneumonia occurs despite the best efforts at prevention. Multiple methods available to prevent ventilator-associated pneumonia are reviewed, and ventilation-associated pneumonia (VAP) is divided into early versus late onset. The authors discuss the organisms associated with each of these situations, the empiric antibiotic choices, and specific issues related to antibiotic therapy such as resistance, pharmcodynamics, tissue penetration, and types of modifications necessary in empiric choice when the cause of VAP is identified. PMID- 10516913 TI - Radiation processing to improve the quality of fishery products. AB - Extensive investigations, worldwide, in the last 4 decades have shown the benefits of radiation processing for the preservation and microbial quality improvement of seafoods. In the present review, the various factors determining the quality of seafoods are first presented. The basic principles underlying the effects of ionizing radiation and specific effects on food constituents such as proteins, amino acids, lipids, vitamins, and tissue enzymes are discussed. Data on radiation processing of seafoods are reported and discussed with respect to shelf life enhancement under refrigeration by control of bacteria causing spoilage, radiation sensitivity of pathogenic microorganisms and parasites of public health significance and their elimination in fresh and frozen fishery products, control of insect disinfestation in dried fishery products, influence of irradiation on nutritional and sensory quality attributes, detection of irradiation treatment, economics, and international status. PMID- 10516914 TI - Caffeine in coffee: its removal. Why and how? AB - The popularity of coffee as a beverage is ever increasing despite the fact that there are reports antagonized to its consumption. Of the several factors cited, the alkaloid caffeine present in coffee can cause addiction and stimulate the central nervous system. It has an effect on the cardiovascular system with a slight increase in blood pressure and heart output. It undergoes biotransformation in the human body to form methylated derivatives of uric acid. In recent times, much effort has gone into the research on the removal of caffeine in coffee, resulting in a specialty product called decaffeinated coffee. Decaffeination methods mainly employ organic solvents or water or supercritical carbon dioxide. These methods with their attendant advantages and disadvantages are reviewed in this article. PMID- 10516915 TI - Physical polymer surface modification methods and applications in food packaging polymers. AB - Continued innovations in the polymer industry have made polymer surface modification methods a subject of intense research. The importance and necessity of surface modification of plastics are explained, and the advantages of physical surface treatments over the less-sophisticated chemical methods are outlined. Currently available physical surface modification methods for food packaging polymers are reviewed from the food packaging perspective. These physical surface modification methods include flame, corona discharge, UV, gamma-ray, electron beam, ion beam, plasma, and laser treatments. The principle of operation of each method is briefly described, and the advantages and disadvantages of each technique are cited. The extent to which each of these methods can produce the specific modifications desired is discussed. Furthermore, the effects of each treatment on barrier, mechanical, and adhesion properties of food packaging polymers are also examined. Finally, an overview of economic aspects of sophisticated surface modification techniques, including ion beam, plasma, and laser treatments, is presented. PMID- 10516916 TI - Patient assessment and preventive measures for medical emergencies in the dental office. AB - Many practicing dentists have faced few medical emergencies in their practice and often overlook effective preventive measures. The proper and accurate assessment of a patient is paramount in preventing an emergency crisis in the dental office. This article helps the practitioner to evaluate the history and physical examination of the patient, including the patient's medical and surgical history, in order to make informed decisions about treatment options. PMID- 10516917 TI - Management priorities and treatment strategies for medical emergencies in the dental office. AB - The importance of prioritization in the management of the emergency dental patient cannot be overemphasized. Each dentists' ability to recognize and deal with emergencies depends on the dentists' education, the training of the staff, and the available equipment. This article helps the dentist prioritize the management of medical emergencies by examining preventive protocol and treatment options, as well as preparation and training strategies. PMID- 10516918 TI - Chronic pain in the dental patient. AB - This article reviews the pathophysiology and mediators of pain, and addresses the different subtypes of dental pain. An overview of pharmacotherapy is presented. Contributing factors to chronic pain syndromes are reviewed. Alternative methods of pain management are discussed. PMID- 10516919 TI - Infectious disease concerns and possible complications in the dental patient. AB - Several questions and concerns often arise in the management of patients with infectious complications. This article explores the difficult issues surrounding the treatment of patients with infectious diseases: infectivity of the present conditions, medication side effects, and potential complications secondary to the dental treatment are questions to be considered in the evaluation of these patients. Dentists should be aware of the management of oral complications of these conditions and are important members of the health care team involved in the follow-up care of these patients. PMID- 10516920 TI - Bleeding and coagulation problems in the dental patient. Hereditary disease and medication-induced risks. AB - This article addresses the unique problems of hereditary and medically induced coagulation problems in dental patients. Both general practitioners and specialists need to have a full understanding of normal events that lead to clot formation to appreciate how treatments and medications administered can adversely affect the final outcome in this patient group. PMID- 10516921 TI - Dental patients with neurologic and psychiatric concerns. AB - Neurologic emergencies demand that the dentist stop all major procedures and remove all foreign objects from the patient's mouth, secure the ABCs, and do a quick secondary survey to identify possible causes while someone else activates the EMS. Psychiatric emergencies relate mostly to anxiety. They can be anticipated by a thorough medical history, which includes a DAS or DAS-R. Behavioral modifications provide the greatest long-term benefit. Pharmacologic intervention should be considered when other methods fail. It is important to recognize the possible morbidity increase when anxiety is not properly addressed as well as the differential diagnoses to be excluded in cases of sympathetic overexcitation. In preparation for any major dental work, it is important to take a thorough medical history in a nonthreatening environment, including medications, psychiatric history, and drug abuse history. Psychiatric emergencies can be prevented by reducing anxiety. Neurologic problems and drug overdoses can be detected earlier when a high index of suspicion is present, and premedication can be planned. PMID- 10516922 TI - Care of the pregnant patient in the dental office. AB - Attention to a woman's dental health can and should continue throughout pregnancy. The treating dentist must be diligent to the special situations surrounding treating the pregnant patient and the risks involved for both fetus and mother. Undertaking the proper precautions with these patients not only provides the best dental care, but also helps avoid potential complications. Any complication or question that arises with a complicated pregnancy should prompt referral to the patient's obstetrician for further evaluation. PMID- 10516923 TI - Cardiac disease and hypertension. Considerations for office treatment. AB - This article focuses on four classifications of cardiac disease in an effort to provide the information necessary to recognize, process, and react appropriately to either a patient's symptoms or medical history. Each section covers relevant demographic data, discusses pathophysiology, and describes what the practicing dentist must know about the underlying illness to make treatment decisions about patients. Management issues on cardiac patient care are also discussed. PMID- 10516924 TI - Dental care of patients with substance abuse. AB - Patients who abuse alcohol, crack, heroin or prescription drugs, are likely to interact with the dental professional. The dentist should therefore be able to identify problems of abuse and provide informed care and referral. Substance abuse should be a consideration in all patients who present with dental trauma and those who present with frequent vague complaints, multiple pain medication allergies, and regimens with multiple narcotic medications. Polydrug use, either prescription or illicit, is also a possibility, and effective treatment requires prompt recognition. Dentists should be alert to drug-seeking behavior within the context of pain management, and because pain severity is an objective experience, each patient must be treated carefully and sensitively. Unrelieved or unremitting pain can be a relapse trigger and therefore adequate pain control is a necessity in the recovering chemically dependent patient. New modalities, such as coanalgesia with low-dose ketamine in the opioid addicted have been shown to work effectively. In the post-dental surgical patient with chemical dependency, agents with less psychoactive activity than their drugs of abuse, such as extended release morphine (MS Contin) have been tried with variable success. An informed treatment plan includes recognition of substance abuse, appropriate intervention, and referral. This plan may include universal screening, followed by brief interventional therapy for positive patients and in some cases, pharmacological pain control. On discharge from the office, instructions concerning referral to a substance abuse program or, in the case of the patient who may require more immediate treatment, to the emergency department are important. PMID- 10516925 TI - Focused care of pediatric patients in the dental office. AB - No dental text can adequately prepare the practitioner with the necessary expertise to treat all presentations or office complications that may arise in the therapy of children. There are times when consultation with the child's parent and pediatrician may answer necessary treatment-related questions. Most chronic conditions do not prevent needed treatment interventions. Any acute illness or exacerbation of a chronic disease should be cleared by the primary care physician before commencing dental treatment. The mainstay of safe practice requires that the dentist to maintain a basic level of understanding of what constitutes an emergency and that office staff receive basic training and are adequately supplied with emergency equipment. Dentists are cautioned to consult their state board or dental society as well as their insurance carrier as to what constitutes necessary emergency equipment in the office and to what level they are responsible for providing emergency care to their patients. There is a great difference within the dental field just as there is with medical specialties. All practitioners, however, are liable for any acts of consignment, and although the intention is not to dissuade anyone from providing assistance in an emergency, supportive care and an immediate call of 911 to activate the local EMS are important. In addition, maintaining a familiarity with the local hospital and emergency department capabilities as well as travel time and distance is also important. Routine reviews and updates on life-saving interventions and resuscitations are good general practice and will save lives. PMID- 10516926 TI - Anesthesia for office-based oral and maxillofacial surgery. AB - This article is divided into two sections, each encompassing the various aspects of anesthesia used for dental procedures ranging from single tooth extraction to more complex reconstruction and tumor eradication. The various types of anesthesia used for each of these kinds of dental surgery are explored, and the effectiveness and cost efficiency of office-based anesthesia procedures are discussed. PMID- 10516927 TI - Antenatal and neonatal haemoglobinopathy screening in the UK: review and economic analysis. PMID- 10516928 TI - [Study on antibiotics susceptibility and mechanism of carbapenem-resistance in clinical isolates of Pseudomonas aeruginosa]. AB - The susceptibility of 260 strains of Pseudomonas aeruginosa to several antibiotics and the mechanism of resistance to carbapenems were investigated. The number of strains of P. aeruginosa moderately resistant or resistant to ofloxacin, ceftazidime and imipenem (IPM) were 76 (29.2%), 31 (11.9%) and 30 (11.5%), respectively. There was no clear relationship between the drug resistance of P. aeruginosa and serum type. Fourteen strains (46.6%) out of 30 IPM-resistant strains were susceptible to meropenem (MEPM). Twenty seven (90.0%) IPM-resistant strains showed cross resistant to panipenem (PAPM), and 12 strains (44.4%) out of the 27 strains showed high susceptibility to MEPM. P. aeruginosa becomes resistant to IPM and PAPM only by the decrease in the outer membrane permeability of these carbapenems. In contrast, P. aeruginosa becomes equally resistant to MEPM by concurrent occurrence of the increase in the efflux of the antibiotics and the decrease in the outer membrane permeability of the antibiotics. The possibility that both mechanisms are taken place concurrently in P. aeruginosa is considered to be low, and it was also supported by the results of the present study. PMID- 10516929 TI - [Tissue penetration properties of macrolide antibiotics--comparative tissue distribution of erythromycin-stearate, clarithromycin, roxithromycin and azithromycin in rats]. AB - The ability of antibiotics to penetrate the target organs is an important factor for the clinical effects and side effects in the treatment of infection. In the present study, the comparative tissue distribution of 4 kinds of macrolide antibiotics was determined in rats. After oral administration of 20 mg/kg, roxithromycin (RXM) had the highest plasma concentration, and clarithromycin (CAM) has the second highest. The Cmax of RXM and CAM were 2.7 and 1.0 micrograms/ml, respectively. On the other hand, both levels of erythromycin stearate (EM-S) and azithromycin (AZM) were extremely low, with a Cmax of 0.1 microgram/ml. Concentration of the 4 compounds were measurable in the liver, kidney, spleen, lung and heart. The concentration in all tissues for each compound were significantly higher than those in the plasma. AZM had the most sustained and the highest tissue levels. The distribution patterns of RXM and AZM were almost similar to the case of EM-S, in that the highest tissue concentration was observed in the liver, followed in descending order by concentration in the kidney, spleen, lung and heart. On the other hand, CAM had the highest concentration in the lung, and was moderated in the liver. Major clinical indications are infections of the respiratory tracts, and commonly reported side effects are hepatotoxity. Therefore, it is worth noting that the lung levels of CAM were significantly higher than in the liver, as the separation of clinical effects and side-effects. PMID- 10516930 TI - [Clinical studies of faropenem in the field of obstetrics and gynecology]. AB - The clinical effect of faropenem was evaluated in 165 ambulatory patients with various infections in the field of obstetrics and gynecology at 10 institutions in Yamagata Prefecture. The results obtained are summarized below. 1. The rate of efficacy, as determined from the clinical effect following 3- to 7-day repeated administration at a dose of 600 mg/day, was 97.9% (46/47) for intrauterine infections, 92.0% (23/25) for adnexitis, 93.8% (15/16) for external genital infections, 88.9% (8/9) for mastitis, 94.0% (63/67) for cystitis, and 100% (1/1) for cervicitis. The overall efficacy rate was estimated to be 94.5% (156/165). 2. The rate of clinical efficacy, as classified by isolate, was high, 95.1% for Gram positive bacteria, 100% for Gram-negative bacteria, and 100% for anaerobes. As for bacteriological response classified by isolate, the eradication rate was high, 91.4% (74/81) for Gram-positive bacteria, 98.4% (62/63) for Gram-negative bacteria, 89.5% (17/19) for anaerobes, and 93.9% (153/163) in all. 3. No adverse reactions or laboratory abnormalities were observed in any patient. The results presented suggest that faropenem is a highly safe and effective antibiotic for the treatment of obstetric or gynecological infections of various kinds in an ambulatory setting. PMID- 10516931 TI - [Effects of levofloxacin once-a-day therapy on uterine cervicitis]. AB - An investigation was carried out to determine the therapeutic effect of levofloxacin (LVFX) once-a-day oral therapy at the dose of 200 mg/day for 7 days on uterine cervicitis, in comparison with LVFX twice-a-day oral therapy at the dose of 200 mg/day for 7 days. Of the 102 patients enrolled in the study, 90 were subjected to the analysis. The efficacy rate on uterine cervicitis of the once-a day therapy and twice-a-day therapy groups according to the evaluation of the Drug Efficacy Evaluation Committee were 72.0% (36/50) and 82.5% (33/40), respectively. The efficacy rate on uterine chlamydial cervicitis of the once-a day therapy and twice-a-day therapy groups according to the evaluation of the Drug Efficacy Evaluation Committee were 88.0% (22/25) and 85.7% (18/21), respectively. Safety was evaluated as "safe" in 88 of the 90 assessable patients (97.8%). Side effects were seen in two cases, which belong to the once-a-day therapy group; mild candidiasis and mild breast distension sense. As the antimicrobial treatment started, the levels of the inflammatory cytokines, interleukin-6 (IL-6) and interleukin-8 (IL-8) in the cervical mucus, decreased. It is suggested that IL-6 and IL-8 can be useful indicators of the antimicrobial treatment in the uterine cervicitis. These results suggested that the LVFX once-a day therapy can be useful on uterine cervicitis. PMID- 10516932 TI - [Effects of an oxacephem antibiotic on liver function in orthopedic surgery]. AB - The subjects were 531 patients who underwent orthopedic surgery. Flomoxef was administered, and liver function was examined before and after administration. Abnormal liver function after administration of flomoxef was found in 14.3% of patients. In male patients, a high rate of 18.8% was observed. A particularly high rate of 37.0% was obtained among patients who showed GOT values of more than 40 U/L before treatment with flomoxef. The prevalence of abnormal GOT and GPT values after administration of flomoxef was 3.6% and 13.2%, respectively. These values were significantly higher than those obtained with other cephem antibiotics. These rates of occurrence of abnormally high GOT and GPT are obviously higher than those submitted at the time of approval and reported in the drug use investigation. The prevalence of abnormal liver function values was high in patients receiving flomoxef, and particularly high in male patients and patients whose GOT was high before administration of flomoxef. Therefore, sufficient check of liver function appears important when administration of flomoxef to these types of patients is intended. PMID- 10516933 TI - P-glycoprotein and drug therapy in organ transplantation. AB - The role of multidrug resistance and P-glycoprotein (P-gp) in the development of drug-resistant tumor cells has been extensively studied. As more knowledge on the physiological functions of P-gp has accumulated, the effects of P-gp modulation on the pharmacokinetics and the pharmacodynamics of many drugs have become apparent. Solid organ transplant recipients receive numerous medications that are substrates for P-gp. The objective of this review is to discuss the effects of P gp modulation on the pharmacokinetics and the pharmacodynamics of immunosuppressive agents such as cyclosporine, tacrolimus, sirolimus, and corticosteroids. Pharmacokinetic alterations may occur in drug absorption since P gp is in the small bowel, in drug distribution since P-gp functions in the blood brain barrier, in drug metabolism since P-gp and cytochrome P450 3A have linked functions, and in drug elimination since P-gp is in the bile canaliculi and renal tubules. A link between P-gp and organ rejection has been speculated since upregulation of the P-gp pump may restrict immunosuppressant drug entry into immunocompetent cells. A further understanding of P-gp regulation upon chronic exposure to P-gp substrates and inhibitors and the potential administration of selective P-gp inhibitors will enhance our ability to use potent immunosuppressive drugs in organ transplant patients. PMID- 10516934 TI - Assessment of the quality and quantity of drug-drug interaction studies in recent NDA submissions: study design and data analysis issues. AB - This report investigates the quality and quantity of drug-drug interaction studies in recent new drug applications (NDAs). Eighty-nine studies contained in 14 NDAs submitted between December 1995 and November 1996 to the U.S. Food and Drug Administration (FDA) were reviewed. The results indicated that the median number of clinical drug-drug interaction studies per NDA was 6, almost double that of a 1994-1995 survey. In vitro metabolism data were present in 70% of the submissions. More than 50% of the submissions contained interaction studies using a battery of drugs (cimetidine, digoxin, or warfarin) without optimal use of the in vitro metabolism or in vivo mass balance data. Various study designs using a median number of 12 subjects were employed in the evaluation of drug-drug interactions. Some of the important study design factors such as dose size, dosing regimen, dosing duration, and timing of coadministration were considered, although not consistently, by the sponsors in their study design. Seventy-five percent of the studies used normal, healthy male subjects, and 25% used patients for whom the new molecular entities were intended. In 33% of the studies, female subjects were also recruited. Although the majority (80%) of the submissions still used p-values to determine the significance of drug interactions, 30% used a more relevant equivalence approach with 90% confidence intervals for key pharmacokinetic and/or pharmacodynamic parameter ratios to assess the extent of drug interactions. Overall, 82% of the studies concluded no interaction. Although population pharmacokinetic analysis can be a useful tool in studying drug-drug interactions, only 21% of the submissions used this approach. In summary, this assessment reveals that the quantity and quality of drug-drug interaction studies in NDAs have improved over the years. These improvements, as well as others that can be implemented, should result in more informative labeling and better patient care. FDA guidance for industry dealing with the design, analysis, and labeling language of in vivo metabolic drug-drug interactions has been developed to assist sponsors and FDA reviewers with these issues. PMID- 10516935 TI - Disposition of recombinant human interleukin-10 in subjects with various degrees of renal function. AB - The pharmacokinetics of intravenously administered recombinant human interleukin 10 (rHuIL-10) were evaluated in 18 subjects with creatinine clearances (Clcr) between 2.7 and 116.7 mL/min/1.73 m2. Serum samples for rHuIL-10 were obtained over a 48-hour period after a single 25 micrograms/kg i.v. bolus infusion. AUC, total body clearance (Clp), and steady-state volume of distribution (Vdss) were derived by compartmental methods. Analysis of serum concentrations showed statistically significant group differences for log-transformed AUC and original scale Clp (p < 0.01). The AUC and effective half-life increased, while the mean Clp of rHuIL-10 decreased as renal function declined. A linear relationship between AUC and Clcr as well as Clp and Clcr demonstrates that the disposition of rHuIL-10 is altered in subjects with renal insufficiency. No serious adverse events were noted. PMID- 10516936 TI - The pharmacokinetics of extended-release formulations of calcium antagonists and of amlodipine in subjects with different gastrointestinal transit times. AB - The influence of gastrointestinal (GI) transit times on the pharmacokinetics (PK) of three calcium channel blockers (CCBs), recommended for once-daily dosing, was investigated. In a three-way crossover design, the single-dose PK of a controlled delivery formulation of 240 mg diltiazem (DIL), an extended-release formulation of 10 mg felodipine (FEL), and 5 mg amlodipine (AML) were compared in two groups of healthy subjects, with either slow (> 35 h) or rapid (< 15 h) GI transit, as assessed by the metal detector method (EAS II). GI transit significantly affected the PK of DIL. Mean PK parameters in the rapid versus slow transit group were the following: trough levels (C24 h): 22.8 +/- 8.3 versus 49.5 +/- 35.7 ng/ml, p < 0.05; AUC 1134.4 +/- 512.7 versus 1704.7 +/- 1185.6 hng/ml, p < 0.05 (one-sided). Neither AUC nor trough levels of FEL and AML were significantly influenced by transit times, nor was Cmax after any of the three treatments. Variations in PK parameters, as indicated by coefficients of variation, were about twofold higher for both DIL and FEL, compared to AML. Variations in mean residence times were significantly lower for AML compared to DIL and FEL (7% vs. 30% and 17%, p < 0.001 and p < 0.002, respectively). Peak-to-trough ratios (Cmax/C24 h mean) were 1.8 +/- 0.9 for DIL, 7.6 +/- 3.5 for FEL, and 1.7 +/- 0.2 for AML. In conclusion, the predictability of pharmacokinetic behavior both in conditions of rapid or slow GI transit is optimized in drugs with intrinsically slow elimination such as amlodipine. The pharmacokinetics of the CCBs with formulation-based once-a-day characteristics are sensitive to GI transit if these processes are rapid enough to interfere with the formulation-specific release profile. PMID- 10516937 TI - Bioequivalence of 1 and 5 mg tacrolimus capsules using a replicate study design. AB - Tacrolimus (FK506, Prograf) is marketed for the prophylaxis of organ rejection following allogenic liver or kidney transplantation. A previously conducted, randomized, 24-subject, crossover bioavailability study of 1 and 5 mg capsules (one period each) failed to demonstrate bioequivalence. A single-dose, four period, four-sequence, randomized, crossover, replicate study (N = 32) was therefore used to evaluate the bioequivalence of the marketed 1 and 5 mg capsules in healthy volunteers. Tacrolimus blood concentrations were measured serially over 72 hours using a commercially available ELISA assay. Noncompartmental pharmacokinetic parameters were determined. Ninety percent CIs of log-transformed parameter ratios were 90.5-101.9, 87.1-101.7, and 89.7-103.8 for Cmax, AUC0-t, and AUC0-infinity, respectively. Since all values were within 80% to 125%, the capsules are bioequivalent. Based on %CVs, intersubject variability was approximately two to three times greater than intrasubject variability. The safety of single 5 mg oral tacrolimus doses administered to healthy volunteers at 7-day intervals was also ascertained. PMID- 10516938 TI - The effect of pharmacokinetically guided acute intravenous testosterone administration on electrocardiographic and blood pressure variables. AB - Previous studies have demonstrated that intravenous testosterone can dilate coronary arteries and increase exercise treadmill time, but the electrocardiographic and hemodynamic effects are unknown. This trial determined the hemodynamic and electrocardiographic effects of dosing intravenous testosterone to achieve a physiologic and a superphysiologic serum testosterone concentration. Twenty men (70.6 +/- 6.2 years) had individualized testosterone bolus and continuous infusions designed to increase the serum testosterone concentration by two (physiologic) and six times baseline (superphysiologic). The men were studied on three occasions when they were randomly allocated to received a placebo, physiologic testosterone regimen, or superphysiologic testosterone regimen. Blood pressures and 12-lead electrocardiograms (ECGs) were taken preinfusion and 28 minutes after initiating the infusion on each visit. The blood pressure (systolic and diastolic) and ECG variables (PR, QRS, QT, QTc, and RR intervals) preinfusion and during the infusion were compared, and the delta changes in the variables were compared between groups. The physiologic testosterone regimen increased the serum testosterone concentration by 2.39 +/- 0.48 times the preinfusion concentration, while the superphysiologic regimen increased it by 6.22 +/- 0.99 times. No significant changes occurred in the blood pressure or ECG variables in any group versus preinfusion values or between the three groups. Exogenously administered intravenous testosterone does not significantly affect the blood pressure or ECG variables when given to achieve physiologic or superphysiologic concentrations. PMID- 10516939 TI - Pharmacokinetics, safety, and tolerability of single intravenous infusions of an adenosine agonist, AMP 579, in healthy male volunteers. AB - The pharmacokinetics of an adenosine agonist, AMP 579, following intravenous administration were evaluated. Single AMP 579 doses of 20 to 150 micrograms/kg were infused intravenously over 6 hours using a constant-rate or two-step rate of infusion to healthy male volunteers. Plasma and urine samples were collected for measurement of AMP 579 concentration using a validated HPLC assay. An assessment of safety and tolerability was also performed. Based on a noncompartmental method of analysis, the pharmacokinetics of AMP 579 were characterized by rapid systemic clearance (0.77 to 0.85 L/h/kg), a moderate steady-state volume of distribution (0.80 to 0.94 L/kg), and a short terminal elimination half-life (0.84 to 1.13 h). AMP 579 exhibited dose (infusion rate)-proportional pharmacokinetics over the dose range. In addition, little or no unchanged AMP 579 was found in the urine. The primary cardiovascular pharmacodynamic response that was observed was a dose related increase in mean ventricular heart rate. Fifteen minutes prior to the end of the infusion, volunteers administered the highest dose (150 micrograms/kg) exhibited a mean (range) 59% (36%-69%) increase in heart rate as compared to a mean (range) 18% (0%-73%) increase for the placebo group. No clinically relevant changes in the systolic or diastolic blood pressure were observed. The information obtained in this study should allow the design of AMP 579 dosage regimens that would maximize plasma AMP 579 concentrations and minimize the incidence of adverse events. PMID- 10516941 TI - The new topical steroid ciclesonide is effective in the treatment of allergic rhinitis. AB - A randomized, placebo-controlled, double-blind crossover study was performed to investigate the efficacy of ciclesonide nasal spray in allergic rhinitis at the dose level of 200 micrograms per nostril. Twenty-four subjects (13 males, 11 females; median age: 28 years) with a history of allergic rhinitis but free of symptoms at screening entered the study. Ciclesonide and placebo were given for 7 days each with a washout period of at least 14 days in between. In both treatment periods, controlled intranasal allergen provocation with pollen extracts was performed on the 2 days before start of treatment (days -2 and -1) and on all treatment days (days 1 to 7) about 2 hours after administration of the study medication. At 5 and 30 minutes after each allergen provocation, rhinal airflow was measured by anterior rhinomanometry, and the subjective symptoms of obstruction, itching, and rhinorrhea were assessed by means of a standardized visual analog scale. Rhinal airflow improved significantly from day 5, while the subjective symptom of obstruction improved from day 2. Itching and rhinorrhea also improved significantly. The local and systemic tolerability of ciclesonide nasal spray was excellent. The results of this study clearly indicate that the new topical steroid ciclesonide is effective in the treatment of allergic rhinitis without producing local or systemic side effects. PMID- 10516940 TI - Fluoxetine bioequivalence study: quantification of fluoxetine and norfluoxetine by liquid chromatography coupled to mass spectrometry. AB - In this study, the authors assessed the bioequivalence of two fluoxetine tablet formulations in 24 healthy volunteers of both sexes who received a single 20 mg dose of each fluoxetine formulation, and a new sensitive method for the quantification of fluoxetine and norfluoxetine in human plasma was developed. The study was conducted using an open, randomized, two-period crossover design with a 4-week washout interval. Plasma samples were obtained over a 672-hour period. Plasma fluoxetine and norfluoxetine concentrations were analyzed by combined liquid chromatography coupled to mass spectrometry (LC-MS) with positive ion electrospray ionization using selected ion recording (SIR). Kolmogorov-Smirnov's test, histograms, probit plots, and the correlation between norfluoxetine AUC(0 infinity) and fluoxetine AUC(0-infinity) were used to analyze the population distribution. The limit of quantification was 0.15 ng.ml-1 and 0.50 ng.ml-1 for both fluoxetine and norfluoxetine, respectively. Within- and between-run imprecision was less than 13% and 17%, respectively. The pharmacokinetic parameters obtained for fluoxetine and norfluoxetine after the administration of each formulation included AUC(0-672 h), AUC(0-infinity), Cmax, Cmax/AUC(0-672 h), tmax, t1/2, and Ke. The AUC values for fluoxetine were not consistent with a normal distribution, reflecting the existence of two different populations (poor and extensive metabolizers). The mean pharmacokinetic parameters for extensive fluoxetine metabolizers were 27.0 ng ml-1 for Cmax, 2064.0 ng h ml-1 for AUC(0 infinity), and 85.4 h t1/2. The mean pharmacokinetic parameters for norfluoxetine (in extensive metabolizers only) were 2532.0 ng h ml-1 for AUC(0-infinity) and 8.4 ng ml-1 for Cmax. For fluoxetine bioequivalence, the 90% CI of the individual ratio geometric mean for Psiquial/Prozac (including both extensive and poor metabolizers) was 101.6% to 121.1% for AUC(0-672 h) and 86.1% to 102.6% for Cmax. For norfluoxetine, the 90% CI of the individual ratio geometric mean for Psiquial/Prozac (including both extensive and poor metabolizers) was 90.3% to 108.3% for AUC(0-672 h) and 84.5% to 106.3% for Cmax. The new method developed (LC-MS) presented high sensitivity, specificity, and short chromatographic run for the quantification of both fluoxetine and norfluoxetine in human plasma. Since both 90% CI for AUC and Cmax geometric mean ratios were included in the 80% to 125% interval proposed by the U.S. Food and Drug Administration, Psiquial was considered bioequivalent to Prozac according to both the rate and extent of absorption. The finding that there were no significant differences in the bioequivalence assessed by either fluoxetine or norfluoxetine pharmacokinetic parameters indicates that future bioequivalence trials may be performed by quantifying fluoxetine only. PMID- 10516942 TI - No effect of gender or age on binding characteristics of valproic acid to serum proteins in pediatric patients with epilepsy. AB - The gender- and age-related binding characteristics of valproic acid to serum proteins were determined in the pediatric population. Serum samples examined in the study were obtained from 61 pediatric patients (28 males, 33 females) with epilepsy on valproic acid monotherapy. Their ages ranged from 1 to 15 years (mean age with [SD]: 7.8 [3.9] years; < 10 years, n = 41; > or = 10 years, n = 20). The in vivo population binding parameters of valproic acid to serum proteins and theoretical minimal unbound serum fraction (fu) of valproic acid were determined in (1) all, (2) male and female subgroups, and (3) prepubescent (< 10 years) and pubescent (> or = 10 years) subgroups. The association constant (K) was approximately 1.4 times higher in male (0.018 L/mumol) than in female (0.013 L/mumol) patients, while the total concentration of binding sites (n(Pt)) was 1.2 times greater in female (1235 mumol/L) than in male (997 mumol/L) patients. The fu was 0.053 and 0.059 for male and female patients, respectively. The value of K was approximately 1.6 times higher in the pubescent (0.019 L/mumol) than in the prepubescent (0.012 L/mumol) patients, while the n(Pt) was 1.2 times higher in the prepubescent (1244 mumol/L) than in the pubescent (1057 mumol/L) patients. The fu was 0.063 for the prepubescent and 0.047 for the pubescent patients. No significant differences were observed in binding characteristics of valproic acid to serum proteins between male and female or younger and older patients. However, the differences in valproic acid binding to serum proteins appear to be relatively larger in binding affinity than in binding capacity between the two groups. Because no significant differences were observed in serum concentrations of total and unbound valproic acid, albumin, or free fatty acids between any subgroups (male and female, younger and older), the results suggest that gender or age may not be factors for the determination of the binding characteristics of valproic acid to serum proteins in pediatric patients. PMID- 10516943 TI - Trimethoprim/sulfamethoxazole does not affect the steady-state disposition of indinavir. AB - This study evaluates the safety and potential pharmacokinetic interaction between indinavir and trimethoprim/sulfamethoxazole (TMP/SMZ). In a randomized, three period crossover fashion, 12 healthy adults received 1 week of indinavir sulfate 400 mg orally every 6 hours with placebo, TMP 160 mg/SMZ 800 mg orally every 12 hours with placebo, and indinavir sulfate with TMP/SMZ. Plasma indinavir, SMZ, and TMP concentrations were determined after the last dose of each treatment period. Concomitant administration resulted in a 17% decrease in geometric mean trough plasma indinavir concentrations (p = 0.032), an 18% increase in geometric mean AUC0-12 h and Cmax TMP values (p = 0.031 and 0.030, respectively), and a 5% increase in geometric mean AUC0-12 h SMZ values (p = 0.039). None of these effects was considered clinically significant. The combination of indinavir sulfate and TMP/SMZ is generally well tolerated, with no clinically significant pharmacokinetic interaction being noted. PMID- 10516944 TI - Altered pharmacokinetics of indinavir by a novel nonnucleoside reverse transcriptase inhibitor (HBY-097): a pharmacokinetic evaluation in HIV-positive patients. AB - HBY-097 (HBY), an investigational nonnucleoside reverse transcriptase inhibitor (NNRTI), and indinavir (IDV) share a common metabolic pathway, cytochrome P4503A4 (CYP3A4), and may clinically be used together as well as with zidovudine (ZDV). Thus, the potential pharmacokinetic (PK) interaction between these drugs was evaluated. HBY (500 mg Q8H), IDV (800 mg Q8H), and ZDV (200 mg Q8H) were given to 8 HIV-infected subjects. Serial plasma samples were collected at baseline (ZDV and IDV alone) and day 11 (all 3 drugs) to determine PK parameters using noncompartmental analysis. PK parameters for ZDV in the presence and absence of HBY were not appreciably different. However, both the maximum (Cmax) and minimum (Cmin) concentrations of IDV were significantly reduced, from a mean of 7514 +/- 1636 and 146 +/- 81 mcg/L to 4725 +/- 2494 mcg/L and 54 +/- 24 mcg/L (p < .05) after addition of HBY. Furthermore, apparent clearance (CL/F) of IDV before and after 11 days of concomitant HBY administration was significantly higher, from 0.69 +/- 0.14 to 1.94 +/- 0.63 L/h/kg (p < .05) with an associated reduction in area under the curve (AUC0-8) from 16,034 +/- 4903 to 6134 +/- 2701 mg/L/h (p < .05). The increase in IDV CL/F is consistent with the observed metabolic induction effects of other NNRTIs. The results of this trial showed that HBY significantly alters the pharmacokinetic parameters of IDV at the dose studied. PMID- 10516945 TI - What is your diagnosis? Non-ketotic hyperosmolar diabetes mellitus. PMID- 10516946 TI - Current concepts in the diagnosis and treatment of brain tumours in dogs and cats. AB - Intracranial tumours occur relatively frequently in dogs, and less commonly in cats. With the availability of computed tomography (CT) and magnetic resonance imaging, more accurate determination of the location and extent of brain tumours in companion animals has become possible. Following these advances in imaging, precise CT-guided stereotactic techniques for both tumour biopsy and intratumoral drug delivery have been developed for use in cats and dogs. Also, tumour identification methods, such as crush preparation examination, have facilitated rapid tumour identification. The use of improved diagnostic techniques has resulted in an increasing demand for effective therapies for brain tumours. While surgical removal and irradiation remain important treatment considerations in the management of brain tumours of cats and dogs, the development of gene therapy strategies for treatment of intracranial tumours offers much promise, although research in this area is still at an early stage. PMID- 10516947 TI - Laminectomy for 34 dogs with thoracolumbar intervertebral disc disease and loss of deep pain perception. AB - The case details and the results of treatment of 34 dogs with thoracolumbar intervertebral disc disease, without deep pain perception, that had been treated by laminectomy and fenestration, are presented. The association of a number of potential prognostic factors with the neurological outcome is examined. Twenty one dogs (62 per cent) recovered neurological function, seven (21 per cent) failed to recover neurological function and three (9 per cent) developed progressive myelomalacia postoperatively, while three dogs (9 per cent) were euthanized intraoperatively because of diffuse myelomalacia. Twenty of the dogs that recovered neurological function showed a return of deep pain perception within two weeks of decompressive surgery. Statistical analysis showed significant differences in the outcome between dogs that took less than one hour to lose the ability to ambulate and dogs with a longer duration of onset of inability to ambulate. The extent of spinal cord swelling determined by myelography was not found to be a useful prognostic indicator. PMID- 10516948 TI - Transanal endoscopic treatment of benign canine rectal tumours: preliminary results in six cases (1992 to 1996). AB - Transanal endoscopic resection and cautery of benign rectal tumours was performed in six dogs with extensive and/or inaccessible rectal neoplasia. The results were encouraging, with three dogs cured and the quality of life of a further two improved for a significant time. The remaining dog died as a result of rectal perforation. Transanal endoscopic treatment of extensive and/or inaccessible benign canine rectal tumours offers an alternative to more radical techniques such as pull-through surgery. PMID- 10516949 TI - A survey of injuries at five greyhound racing tracks. AB - The number of orthopaedic injuries sustained by racing greyhounds from five greyhound tracks in the state of Wisconsin, USA, was obtained over a two-year period. Calculated injury rates were analysed to predict the probability that a given competitor would have an injury based on track design, temperature, bodyweight, grade of race, race distance, race number, injury location on track and type of trauma. One track had a significantly higher injury rate than the others, and this track was constructed with a decreased initial straightaway, a decreased turning radius in the second turn and an increased turn bank. Increased injury rates were also seen with successively higher grades of race, suggesting a possible correlation with speed. Race distance had a significant effect on racing greyhound injury rates as well. Races measuring 3/16 mile and 7/16 mile resulted in a higher incidence of injury as compared with races with lengths of 5/16 mile and 3/8 mile. Injuries were most likely to occur at the first turn of a race. Temperature, bodyweight, race number and type of trauma had no significant effect on injury rate. Speed, race distance and track design were significant factors that were found to influence the injury rate of the racing greyhound and should be areas to focus on for the prevention of injury. PMID- 10516950 TI - Chronic pneumonia caused by Mycobacterium thermoresistibile in a cat. AB - Mycobacterium thermoresistibile was isolated in pure culture from ultrasound guided pulmonary aspirates taken from a young cat with severe, chronic, pyogranulomatous pneumonia. Thoracic radiography and ultrasonography before therapy demonstrated severe diffuse alveolar disease. Twelve months combination therapy with doxycycline, rifampicin and clarithromycin resolved the infection. Thoracic radiographs taken at the completion of therapy showed multifocal pulmonary mineralisation. M thermoresistibile has been infrequently reported as a human or animal pathogen. This is the first reported pulmonary infection by M thermoresistibile in a cat and documents the successful treatment of the organism in a feline patient. PMID- 10516951 TI - Partial surgical removal of an intramedullary epidermoid cyst from the spinal cord of a dog. AB - An intramedullary space-occupying lesion in the form of an epidermoid cyst was diagnosed in a one-and-a-half-year-old flat-coated retriever. Dorsal laminectomy and durotomy were performed in order to establish the diagnosis followed by excision of one third of the cyst. The remaining cystic tissue that was intimately attached to the spinal cord parenchyma was left in place in order to avoid further damage to the nervous tissue. The dog's neurological status improved dramatically after the surgery, but deteriorated four months later due to recurrence of the cyst. PMID- 10516952 TI - Mesenchymal chondrosarcoma in a young German shepherd dog. AB - An 18-month-old, entire male German shepherd dog was presented with signs indicative of a caudal abdominal space-occupying mass. A needle-core biopsy of this mass failed to establish a definitive diagnosis, but identified a prominent round-cell component. The dog's condition worsened and euthanasia was performed on humane grounds. Histopathology of the mass revealed a mesenchymal chondrosarcoma. PMID- 10516953 TI - Osteomyelitis of the atlanto-occipital region as a sequela to a pharyngeal stick injury. AB - An eight-year-old female Belgian shepherd dog was referred for investigation of chronic neck pain. The dog had sustained a pharyngeal injury 12 weeks previously while catching a stick. Radiographs of the cervical spine revealed signs consistent with a septic arthritis of the atlanto-occipital joint and osteomyelitis of both occipital condyles and the atlas. A foreign body was identified ultrasonographically in the retropharyngeal soft tissues, and a stick was surgically removed from a site ventral to the right side of the atlanto occipital joint. The signs of neck pain started to resolve within a week of surgery. PMID- 10516954 TI - Another busy year for the veterinary poisons information service. PMID- 10516955 TI - Stray dogs--a worldwide problem. PMID- 10516956 TI - Alpha-2 adrenergic modulation of prefrontal cortical neuronal activity related to spatial working memory in monkeys. AB - The effects of systemically administered or iontophoretically applied clonidine (alpha-2 adrenergic agonist) and iontophoretically applied yohimbine (alpha-2 adrenergic antagonist) were examined on prefrontal cortical (PFC) neurons related to spatial working memory (SWM). Systemically administered clonidine (0.04 mg/kg) enhanced SWM-related PFC neuronal activity by 32.5 +/- 14.5%, (mean +/- SD; n = 25 neurons). The facilitatory effect of clonidine was antagonized by iontophoretically applied yohimbine. Iontophoretically applied clonidine enhanced SWM-related PFC neuronal activity by 38.2 +/- 18.6%, (n = 13 neurons), whereas similarly applied yohimbine suppressed it by 34.4 +/- 17.8% (n = 28 neurons). These results indicate that: a) systemically administered clonidine can facilitate SWM-related PFC neuronal activity through actions at alpha-2 adrenoceptors in the PFC; and b) conversely, blockade by yohimbine of alpha-2 adrenoceptors in the PFC suppresses SWM-related neuronal activity. The present study provides neurophysiological evidence that alpha-2 adrenoceptors in the PFC are involved in the cellular mechanisms underlying working memory. PMID- 10516957 TI - Treatment with the noradrenergic alpha-2 agonist clonidine, but not diazepam, improves spatial working memory in normal young rhesus monkeys. AB - Noradrenergic alpha-2 agonists such as clonidine and guanfacine improve working memory performance in aged monkeys. Guanfacine also improves cognition in young monkeys, but there are conflicting reports of the effects of clonidine in young adult human and nonhuman primates. In the present study, high doses of clonidine (0.02-0.1 mg/kg) significantly improved performance of the delayed response task, a test of spatial working memory, in young adult monkeys. Lower doses (0.0001 0.01 mg/kg), similar to those used in human studies (0.001-0.003 mg/kg), had no effect on task performance. In contrast, monkeys experimentally depleted of catecholamines by chronic reserpine treatment have been improved by both dose ranges. These results provide further support for the hypothesis that alpha-2 agonists improve cognition via actions at post-synaptic alpha-2 receptors, and suggest that conflicting results with clonidine in previous studies of prefrontal cortical function may result from insufficient dosage. PMID- 10516959 TI - The brain metabolic patterns of clozapine- and fluphenazine-treated female patients with schizophrenia: evidence of a sex effect. AB - The regional cerebral glucose metabolic rates of clozapine-treated and fluphenazine-treated women with schizophrenia and normal controls were obtained by positron emission tomography (PET) using [18F]-2-fluoro-2-deoxy-D-glucose (FDG) as the tracer. The regional metabolic patterns were compared to each other and to the changes previously observed in men. In women, as in men, both clozapine- and fluphenazine-treatment were associated with lower metabolism in the superior prefrontal cortex and higher metabolism in the medial temporal lobe. In both men and women, clozapine treatment led to a greater lowering of inferior prefrontal cortex activity than fluphenazine, which was statistically significant in the larger male cohort. Fluphenazine led to higher metabolic rates in the lateral temporal lobe than clozapine did, but the differences between the two neuroleptics were not statistically significant in either group. The greatest differences in the female as compared to the male responses to fluphenazine and clozapine were in the cingulate and striatum. As compared to controls, the cingulate metabolic rates of women were reduced by 9.1% and 11.4% on clozapine and fluphenazine, respectively; whereas, men have a statistically nonsignificant reduction of 0.1% with clozapine and a 3.2% increase with fluphenazine. In men, fluphenazine was associated with a much greater elevation in basal ganglia metabolic rates than was clozapine, 23.5% as compared to 3.75%; whereas, in women, basal ganglia metabolic rates are nearly equally increased by fluphenazine (21.6%) and clozapine (15.1%). PMID- 10516958 TI - Comparison of the novel antipsychotic ziprasidone with clozapine and olanzapine: inhibition of dorsal raphe cell firing and the role of 5-HT1A receptor activation. AB - Ziprasidone is a novel antipsychotic agent which binds with high affinity to 5 HT1A receptors (Ki = 3.4 nM), in addition to 5-HT1D, 5-HT2, and D2 sites. While it is an antagonist at these latter receptors, ziprasidone behaves as a 5-HT1A agonist in vitro in adenylate cyclase measurements. The goal of the present study was to examine the 5-HT1A properties of ziprasidone in vivo using as a marker of central 5-HT1A activity the inhibition of firing of serotonin-containing neurons in the dorsal raphe nucleus. In anesthetized rats, ziprasidone dose-dependently slowed raphe unit activity (ED50 = 300 micrograms/kg i.v.) as did the atypical antipsychotics clozapine (ED50 = 250 micrograms/kg i.v.) and olanzapine (ED50 = 1000 micrograms/kg i.v.). Pretreatment with the 5-HT1A antagonist WAY-100,635 (10 micrograms/kg i.v.) prevented the ziprasidone-induced inhibition; the same dose of WAY-100,635 had little effect on the inhibition produced by clozapine and olanzapine. Because all three agents also bind to alpha 1 receptors, antagonists of which inhibit serotonin neuronal firing, this aspect of their pharmacology was assessed with desipramine (DMI), a NE re-uptake blocker previously shown to reverse the effects of alpha 1 antagonists on raphe unit activity. DMI (5 mg/kg i.v.) failed to reverse the inhibitory effect of ziprasidone but produced nearly complete reversal of that of clozapine and olanzapine. These profiles suggest a mechanism of action for each agent, 5-HT1A agonism for ziprasidone and alpha 1 antagonism for clozapine and olanzapine. The 5-HT1A agonist activity reported here clearly distinguishes ziprasidone from currently available antipsychotic agents and suggests that this property may play a significant role in its pharmacologic actions. PMID- 10516960 TI - Developmental regulation of the dopamine D1 receptor in human caudate and putamen. AB - Perturbations in the developmental regulation of the dopaminergic system have been hypothesized to participate in the age-dependent onset of schizophrenia. Although data from studies of non-human primates suggest that dopamine D1-like receptors decrease during adolescence, less information is available concerning changes in human brain. The present study employed quantitative receptor autoradiography to measure D1-like receptor density and affinity in human caudate and putamen. Samples were obtained postmortem from 15 subjects (9 weeks to 49 years), and grouped a priori into three classes: infants, adolescents, and adults. Receptor density and affinity were assessed by saturation binding with [3H]-SCH23390, a D1 receptor antagonist. A decrease in D1 receptor density was observed from infancy to adulthood, with no change in receptor affinity. The temporal pattern of D1-like receptor expression during maturation may play a role in the interaction of dopamine with other neurotransmitter systems, and in the occurrence and pharmacotherapy of neurological and neuropsychiatric disorders. PMID- 10516961 TI - Pertussis toxin lesions of the rat substantia nigra block the inhibitory effects of the gamma-hydroxybutyrate agent, S(-)HA-966 without affecting the basal firing properties of dopamine neurons. AB - S(-)3-amino-1-hydroxypyrrolidone-2 (S(-)HA-966), a potent gamma-hydroxybutyrate like drug, inhibits spontaneous firing and induces a pacemaker-like discharge pattern in nigral dopamine (DA)-containing neurons. Recent evidence has suggested that these effects could be mediated by GABAB receptors and, thus, is likely to involve G protein intermediaries. To test this hypothesis, extracellular single unit recording techniques were used to assess the effects of S(-)HA-966 in animals that had received an intranigral injection of pertussis toxin (PT). Failure to respond to the inhibitory effects of apomorphine was taken as presumptive evidence that PT-sensitive G protein-coupled receptors had been inactivated. No significant differences were observed in the basal firing properties of DA cells recorded in control and PT-lesioned animals. However, in marked contrast to the inhibitory effects observed in uninjected and sham lesioned animals, S(-)HA-966 significantly increased the firing rate of apomorphine-insensitive DA neurons in PT-lesioned rats. The excitatory effects of S(-)HA-966 were accompanied by a significant reduction in bursting activity and an increase in the regularity of firing. These data indicate that the inhibitory effects of S(-)HA-966 are mediated locally within the substantia nigra by a PT sensitive substrate, presumably a G protein-coupled receptor. PMID- 10516962 TI - Gamma-hydroxybutyrate and cocaine administration increases mRNA expression of dopamine D1 and D2 receptors in rat brain. AB - The effects of acute and repeated gamma-hydroxybutyrate (GHB) and cocaine administration on D1 and D2 dopamine receptor mRNA expression were examined using in situ hybridization histochemistry in different rat brain structures rich in GHB receptors. Six hours after a single GHB administration (500 mg/kg i.p.), an increase in D1 and D2 mRNA expression was observed in almost all regions examined; whereas, acute cocaine injection (20 mg/kg i.p.) had no effect. Repeated exposure to GHB (500 mg/kg i.p. twice daily) for 10 days, followed by a 14-h withdrawal period, induced increasing effects on D1 and D2 dopamine receptor mRNA expression, similar to those caused by chronic treatment with cocaine (20 mg/kg i.p. once a day). These effects of GHB and cocaine on dopamine receptor mRNA expression could be a consequence, for both compounds, of the modulation of dopaminergic activity; thus, supporting the benefit of GHB in cocaine substitution therapy. PMID- 10516963 TI - Conditioning to injection procedures and repeated testing increase SCH 23390 induced catalepsy in mice. AB - The cataleptic behavior induced by the dopamine D1 antagonist SCH 23390 (SCH) has proven to be a useful assay for investigating the sensitivity of D1-like dopamine receptor-mediated effects during chronic drug administration. A fundamental flaw in most of these studies may be the involvement of the "repeated measures effect," a behavioral phenomenon well demonstrated for neuroleptic-induced catalepsy but not yet investigated for dopamine D1 antagonists. In this study, mice exposed for various sessions to the bar test presented a strong sensitization to the cataleptic behavior induced by repeated SCH treatment. Conversely, single tested animals exhibited a trend toward decreased catalepsy after repeated SCH treatment, which was in line with the development of a D1-like dopamine receptor supersensitivity suggested by an increase in SKF 38393-induced grooming behavior. Surprisingly, a challenge intraperitoneal saline injection increased the cataleptic behavior of single tested mice after long-term SCH treatment. This "injection-conditioned catalepsy" was also observed after repeated treatment with the dopamine D2 antagonists, haloperidol and metoclopramide. While these findings seem to explain some important contradictory data in the literature, they provide a new and simple animal model of the placebo effect. PMID- 10516965 TI - [Roles of cytokines in wound healing processes]. AB - In recent years, a number of studies have revealed the importance of cytokines and growth factors in the wound healing process. Cytokines, such as epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), transforming growth factor (TGF) alpha 1 and interleukin (IL)-6, have been shown to exhibit the particular ability to stimulate keratinocyte proliferation. In addition, bFGF and TGF beta 1 not only facilitate the migration of monocytes, neutrophils, macrophages, and fibroblasts, but also play a role in the generation of granulation tissue. Cytokine modulation of the repair process in both transitory and chronic wounds remains a very important subject for investigation. In this study, the clinical application of cytokines or cytokine-promoting drugs which were combined with or without collagen matrix is discussed as well as perspectives on wound care in the future. PMID- 10516964 TI - Exercise, antidepressant medications, and enhanced brain derived neurotrophic factor expression. AB - Physical activity and antidepressant treatment have each separately been of significant interest for the management of Alzheimer's disease (AD); particularly the behavioral problems associated with this dementing disorder. We have found that combined antidepressant treatment and physical activity have an additive, potentiating effect on BDNF mRNA expression within several areas of the rat hippocampus. During the 20-day experimental period, animals were treated daily with imipramine (15 mg/kg) or tranylcypromine (7.5 mg/kg) by intraperitoneal injection. Exercising rat groups were given access to running wheels for the duration of the experiment. BDNF mRNA levels were assessed in several cell groups of the hippocampus by in situ hybridization, using a [35S] labelled riboprobe complementary to the full-length BDNF sequence, and computer-assisted densitometry. The combination of physical activity and antidepressant treatment for the 20-day period led to a significant potentiation of full-length BDNF mRNA levels within the dentate gyrus and CA 1, CA 3, and CA 4 cellular fields, above the levels obtained with each intervention alone. These results provide impetus for the study of physical exercise as a potential enhancer of treatment response to antidepressants. PMID- 10516966 TI - [Keloids]. AB - The treatment of keloids has always been a formidable task and no definitive treatment method has yet been established. We have thus studied the histopathological findings to develop a new and effective treatment for keloid scars. In this article, we first describe the morphological and histopathological analysis of keloid lesions and then secondly keloid treatment methods with a mast cell degranulation suppressant that we developed and a modified enucleation method to remove keloid-forming cells and suppress the herniation-like phenomenon. We further summarize other currently available keloid treatment methods. PMID- 10516967 TI - [Artificial dermis]. AB - An artificial dermis, composed of an inner layer of collagen sponge and an outer silicone layer, was developed by modifying the material reported by Yannas et al. When the artificial dermis is placed on full 14-thickness skin defects, the pores of the inner collagen sponge are infiltrated by fibroblasts and capillaries. This cell-infiltrated membrane is gradually converted into a synthesized connective tissue matrix, similar to true dermis, as the original network of collagen sponge is biodegraded. The secondary skin graft takes easily and postoperative contraction is minimal even if a very thin split-thickness skin graft is used. The requirement for two-stage surgery and long hospitalization remain disadvantages of the material. Consequently, application of the material is limited to cases in which the advantages outweigh the disadvantages. Deep skin defects with partial exposure of bone or muscle appear to be the most suitable indications for the material. Shallow but very wide skin defects also seem to benefit from the application of the material. PMID- 10516968 TI - [Tissue engineering]. AB - The new field of tissue engineering, applies the principles of biology and engineering to the development of functional substitutes for the loss or failure of an organ or tissue. This article describes the previous challenges and present status of this interdisciplinary field. PMID- 10516969 TI - [Recent advance in head and neck reconstruction]. AB - The clinical application of microvascular free-flap transfers in reconstructive surgery has expanded tremendously since their introduction. Difficult reconstruction in the head and neck region can now be accomplished in a one-stage procedure using these techniques. Free flaps such as rectus abdominis, forearm, and scapular flap have been used frequently in this region because of their many advantages. They have a long vascular pedicle of a large-caliber vessel with anatomic stability and have ample blood supply. It is easy to harvest, and donor site morbidity in negligible. However, further improvement and refinement of surgical procedures are required to obtain better functional results and increase patients' quality of life. In this article, we describe the most recent concepts and techniques in head and neck reconstruction. PMID- 10516970 TI - [TRAM flap breast reconstruction using a fascia-sparing technique]. AB - Although the TRAM flap has been accepted as one of the most common methods for breast reconstruction utilizing the autologous tissue, its disadvantage is that scarring of the abdominal sheath and muscle may result in postoperative abdominal bulge or hernia. To solve this problem, the authors developed the fascia-sparing technique in TRAM flap breast reconstruction. The technique, in which most of the anterior rectus sheath is preserved, has been applied in 3 patients after radical mastectomy and 7 patients after modified radical mastectomy. With an average follow-up period of 1 year and 2 months, no abdominal bulge or hernia was noted in any patient without the use of prosthetic mesh for the abdominal closure. This fascia sparing technique is particularly effective for TRAM flap reconstruction requiring bilateral rectus abdominis muscle portions and containing only a few minor perforators, for which a DIEP flap is not suitable. PMID- 10516971 TI - [Supramicrosurgical lymphaticovenular anastomosis for the treatment of lymphedema in the extremities]. AB - During the past eight years, we treated obstructive lymphedema of a unilateral upper extremity in 27 females and of a unilateral or bilateral lower extremity in 35 males and females with supramicrosurgical lymphaticovenular anastomoses and/or conservative treatment. The most common cause of upper limb edema was mastectomy with or without subsequent radiation therapy for breast cancer, and that of lower limb edema was hysterectomy with radiation. As an objective assessment of edema, the circumferences of the affected and opposite normal forearms or lower legs were measured 10 cm below the olecranon of the arm or the lower border of the patella. In patients who received conservative treatment (12 arms and 12 legs), the average excess circumferential length of the affected arm and leg was 6.4 and 7.1 cm over that of normal extremities, average duration of edema before treatment was 3.5 and 5.2 years, average period for conservative treatment was 10.6 months and 1.5 years, and average decreased circumferential length was 0.8 and 0.6 cm, respectively. The rate of circumferential decrease over 4 cm was none in arm and 16.7% in leg edema. In patients who underwent surgery (12 arms and 16 legs), the average excess circumferencial length was 8.9 and 9.8 cm, average duration of edema before surgery was 8.2 and 8.9 years, average follow-up after surgery was 2.2 and 3.3 years, and average decrease in excess circumference was 4.1 and 2.7 cm, respectively. The rate of circumferential decrease over 4 cm was 58.3% in arms and 50% in legs. These results indicate that supramicrolymphaticovenular anastomoses have a valuable place in the treatment of obstructive lymphedema. PMID- 10516972 TI - [Application of microvascular surgery in reconstructive digestive tract surgery]. AB - In our institutes, microvascular surgery has been effectively used in reconstructive digestive tract surgery, including esophageal reconstruction and hepatic arterial reconstruction. Free jejunal transfer combined with a gastric pedicle or microvascularly augmented elongated gastric pedicle has been utilized for total esophageal reconstruction. A microvascularly augmented jejunal pedicle or colonic pedicle has been applied in thoracic esophageal reconstructive cases with gastrectomy. Moreover, microvascular surgery has been performed in the reconstruction of the hepatic arterial system in the surgical treatment of pancreatic or bile duct cancer and living related-donor liver transplantation. Some pitfalls in selection of the recipient vessels and handling the intraperitoneal vessels for microvascular anastomosis are also described. Although microvascular surgery has been carried out by plastic and reconstructive surgeons in a team surgical approach, revisions in the medical educational system to create a new-type of surgeon with practical skills and clinical experience in both digestive tract and microvascular surgery will be required in future. PMID- 10516973 TI - Overview of complementary therapies in physical medicine and rehabilitation. AB - This article briefly describes the current usage of alternative and complementary medicine in the United States and among chronically ill users of rehabilitation services. Definitions of alternative and complementary medicine are presented with a few examples from among hundreds of existing therapies, with a focus on therapies currently used in rehabilitation contexts. The role of the National Institutes of Health (NIH), in funding research on these therapies is described. Examples of evidence-based therapies include ginkgo biloba for cerebral insufficiency, and acupuncture for stroke, are presented. Future trends are discussed. PMID- 10516974 TI - Acupuncture in the management of pain of musculoskeletal and neurologic origin. AB - Acupuncture has been used as therapeutic treatment for the health of the Chinese people for more than 3000 years; it is a system for diagnosing and treating disease using fine needles inserted into specific points of the body. Acupuncture can treat a wide variety of conditions. This article discusses the use of acupuncture in the management of pain of musculoskeletal and neurologic origin, with a focus on pain in spinal cord injuries. PMID- 10516975 TI - Auricular acupuncture and auricular medicine. AB - Auricular acupuncture is a treatment system based on normalizing the body's pain and dysfunction through stimulation of points on the ear. Resulting amelioration of pain and illness is believed to be through the reticular formation through the sympathetic and parasympathetic nervous systems. Scalp acupuncture is one of the newest systems of microacupuncture therapy and anesthesia developed in the People's Republic of China; here the needle is inserted and stimulated in the areas directly above the corresponding nerve center. Hand acupuncture is a recently developed type of microreflex system that can be used exclusively or in addition to body acupuncture. Hand acupuncture is indicated to treat all ailments for which acupuncture is traditionally recommended. PMID- 10516976 TI - Scalp acupuncture. PMID- 10516977 TI - Hand acupuncture. PMID- 10516978 TI - Neural therapy. AB - Neural therapy is a treatment system to relieve chronic pain and illness through the injection of local anesthetics into scars, peripheral nerves, autonomic ganglia, trigger points, glands, and other tissues. Treatment is based on normalizing the dysfunctional autonomic nervous system, which initiates or propagates many chronic ailments. Neural therapy has been used widely by European physicians for over 50 years. Theories of action relating to a dysfunctional autonomic nervous system and chronic pain or illness are discussed. PMID- 10516979 TI - Bodywork therapy systems. AB - This article provides an overview of various systems of bodywork including biomechanical and structural systems, movement therapies, and energy-field techniques. Similarities among these systems are described as including the ten principles of movement. Generalizations are also provided regarding the difference between separate systems of bodywork. The review of systems is presented by topical groups within a historic framework. PMID- 10516980 TI - Western movement therapies. AB - Complementary movement therapies have been developed by individuals trained in a variety of disciplines. Movement therapies involve the body, mind, and spirit, and therefore the most informed opinion and recommendations come from personal experience. This article explores nine of the more popular Western movement therapies, which have been chosen for their availability and suitability in the rehabilitation context. PMID- 10516981 TI - Eastern movement therapies. AB - Tai chi, qigong, and yoga represent a class of exercise that differs from the routine strengthening and stretching programs currently employed in physical medicine. These techniques incorporate a "mind-body" approach to the rehabilitation of disorders commonly seen by physical medicine and rehabilitation clinicians. Research into the efficacy of these techniques clearly is in the beginning stages. What little has been conducted thus far is promising. These methods may serve to add valuable contributions to the continuity of care of ambulatory and non-ambulatory patients. PMID- 10516982 TI - Massage techniques in rehabilitation medicine. AB - Massage is an ancient practice that has been integrally incorporated into the management of disease and the maintenance of health across cultures and throughout time. This article discusses the history of massage and the present techniques in practice. The presumed therapeutic effects of massage and the scientific basis to support these ideas are examined. Reported contraindications and complications of massage are reviewed. Scientific research and current trends in the therapeutic use of massage are presented. PMID- 10516983 TI - Prayer and meditation as medical therapies. AB - Prayer and meditation have been used as health-enhancing techniques for centuries. Their use has been investigated more recently in the context of more conventional, allopathic medical approaches. These studies, despite methodological limitations, show some promise for the formal application and integration of these techniques into western medical practice. Some potential benefits from meditation include reduced perceived stress and improvement in mild hypertension. Prayer appears to offer subjective benefit to those who pray; the effects of intercessory prayer on the health status of unknowing individuals requires more investigation. PMID- 10516984 TI - Treatment of musculoskeletal pain with traditional Chinese herbal medicine. AB - Traditional Chinese herbal medicine (TCHM) uses naturally occurring plant, animal, and mineral substances to treat or assist in the treatment of the full spectrum of human disease. With the rise in popularity of alternative or complementary medicine, all physicians will encounter patients using TCHMs. TCHM should be taken under the supervision of a provider whose herbal training and competence is commensurate with the intensity of the herbal regimen and the severity of the clinical condition. TCHM can be valuable in the treatment of all kinds of pain: either as primary or adjunctive therapy depending on the clinical situation. TCHM therapies are prescribed in the framework of a unique diagnostic approach, and are highly specific with regard to type of pain and location of pain. PMID- 10516985 TI - Nutrition and dietary supplements. AB - Quality and number of subjects in blinded controlled clinical trials about the nutrition and dietary supplements discussed here is variable. Glucosamine sulfate and chondroitin sulfate have sufficient controlled trials to warrant their use in osteoarthritis, having less side effects than currently used nonsteroidal anti inflammatory drugs, and are the only treatment shown to prevent progression of the disease. Dietary supplements of ephedrine plus caffeine for weight loss (weight loss being the current first line recommendation of physicians for osteoporosis) show some promise, but are not sufficient in number of study subjects. Phenylpropanolamine is proven successful in weight loss. Both ephedrine and phenylpropanolamine have resulted in deaths and hence are worrisome [table: see text] as an over-the-counter dietary supplement. Other commonly used weight loss supplements like Cola acuminata, dwarf elder, Yohimbine, and Garcinia camborgia are either lacking controlled clinical trials, or in the case of the last two supplements, have clinical trials showing lack of effectiveness (although Garcinia has been successful in trials as part of a mixture with other substances, it is unclear if it was a necessary part of the mixture). Safety of these weight loss supplements is unknown. Chromium as a body building supplement for athletes appears to have no efficacy. Creatine may help more in weight lifting than sprinting, but insufficient study subjects and safety information make more studies necessary. Carbohydrate loading is used commonly before endurance competitions, but may be underused as it may be beneficial for other sport performances. Supplements for muscle injury or cramps have had too few studies to determine efficacy. Although proper rehydration with fluids and electrolytes is necessary, a paucity of actual studies to maximize prophylactic treatment for exercise induced cramping still exists. Nutritional supplements for cardiovascular disorders are generally geared to prevention. The United States Department of Agriculture has good recommendations to prevent atherosclerosis; a stricter version by Ornish was shown to reverse coronary heart disease, and the low meat, high fruit, and vegetable DASH diet has been found to decrease hypertension. The epidemiologic studies of hyperhomocysteinemia are impressive enough to give folic acid (or vitamin B6 or B12) supplements to those with elevated homocysteine levels and test patients who have a history of atherosclerotic disease, but no controlled clinical trials have been completed. Soluble fiber has several positive studies in reduction of cholesterol levels and generally is accepted. The data on vitamin E are the most confusing. This vitamin was not helpful in cerebrovascular prevention in China and not helpful at relatively small doses (50 mg) in the United States or Finland against major coronary events. Levels of 400 mg appeared to decrease cardiovascular disease in the United States in studies based on reports by patients and in one large clinical trial. Vitamin E also was successful in prevention of restenosis after PTCA in one clinical trial. Both of these clinical trials need to be repeated in other developed country populations. Some nutritional and dietary supplements are justifiably useful at this point in time. Several meet the criteria of a late Phase 3 FDA clinical trial (where it would be released for public use), but many dietary supplements have insufficient numbers of studies. Some deaths also have occurred with some supplements. If these supplements were required to undergo clinical trials necessary for a new drug by the FDA, they would not be released yet to the public. Several nontoxic supplements appear promising, though need further study. Because they have essentially no toxicity (such as folic acid with B12, soluble fiber, and vitamin E) and may have efficacy, some of these supplementations may be useful now, without randomized clinical trials. PMID- 10516986 TI - Homeopathy in rehabilitation medicine. AB - Rehabilitationists must be able to assess not only the diagnostic entity, but also the person who is sick. The rehabilitation community's understanding of the holism involved in healing and the limitations of conventional therapies creates an openness to consider the usefulness of unconventional therapies. This article explores the role of homeopathy in rehabilitation medicine. PMID- 10516987 TI - Evolution of magnetic therapy from alternative to traditional medicine. AB - Static or electromagnetic fields have been used for centuries to control pain and other biologic problems, but scientific evidence of their effect had not been gathered until recently. This article explores the value of magnetic therapy in rehabilitation medicine in terms of static magnetic fields and time varying magnetic fields (electromagnetic). A historical review is given and the discussion covers the areas of scientific criteria, modalities of magnetic therapy, mechanisms of the biologic effects of magnetic fields, and perspectives on the future of magnetic therapy. PMID- 10516988 TI - Acupuncture education and integration in the physical medicine and rehabilitation residency. AB - Acupuncture is a growing field of interest to patients, medical students, and physicians. This article outlines the educational efforts and requirements to teach and practice acupuncture in the United States. A rationale for integrating acupuncture into physical medicine and rehabilitation residency programs is included. This article provides a structure for determining the type of education goals and plans most suited to a particular residency. PMID- 10516989 TI - Oral phosphate binders: phosphate binding capacity of iron (III) hydroxide complexes containing saccharides and their effect on the urinary excretion of calcium and phosphate in rats. AB - Phosphate binders that contain aluminum or calcium are frequently prescribed to treat hyperphosphatemia in patients with end-stage renal disease (ESRD), but an accumulation of aluminum can lead to encephalopathy, aluminum-related bone disease (ARBD) such as osteomalacia, anaemia, and resistance to erythropoietin, and calcium accumulation can lead to hypercalcaemia. High phosphate concentrations are reduced in vitro and in vivo by a phosphate adsorption pill, which is synthesized by hydrolyzing ferrous sulfate in the presence of saccharides, to form an iron (III)-saccharide complex that is acid resistant and binds phosphate greater than iron (III) hydroxide alone. Under in vitro conditions, containing 3.26 mg P/dL, the iron (III)-sucrose complex showed the highest phosphate adsorption capacity at pH 2 with artificial gastric juice, 58.9 mg P/g binder. For the 7 day in vivo study, 0% (Group 1), 1% (Group 2), 4% (Group 3), and 8% (Group 4) iron (III)-sucrose complex was admixed into the rodent chow by weight and fed to 15 male Wistar rats. The weight and volume of the feces and urine, and the calcium, iron, and phosphorus excretions in the feces and urine samples were monitored for any signs of irregularity. Total urine outflow was collected during a 24-h period to determine the amount of phosphate recovered, which indicates the ability of the phosphate binder to reduce gastrointestinal phosphate absorption. The fecal iron excretion was significantly effected by the amount of binder ingested throughout the study for Group 2 (p < 0.001), Group 3 (p < 0.01), and Group 4 (p < 0.001). The urinary calcium excretion (mg/rat/24-h) significantly increased by the 7th day for Group 2 (p < 0.05) and Group 4 (p < 0.01) in comparison to the control. Finally, after 7 days, there was a significant drop in the urinary phosphorus levels (mg P/rat/24-h) in a dose dependent manner for Group 2: from 7.82 +/- 1.46 to 1.98 +/- 0.10 mg P/rat/24-h (102 mg P/dL/24-h; p < 0.05); Group 3: from 6.70 +/- 1.14 to 0.16 +/- 0.09 mg P/rat/24-h (6.0 mg P/dL/24-h; p < 0.01); and Group 4: from 8.25 +/- 0.67 to 0.04 +/- 0.01 mg P/rat/24-h (0.9 mg P/dL/24-h; p < 0.01). The results show that this new adsorbent might provide an alternative to conventional aluminum and calcium containing phosphate-binding agents for combating hyperphosphataemia. PMID- 10516990 TI - Role of ticlopidine on adriamycin-induced nephropathy. AB - The effect of ticlopidine on rats with adriamycin nephropathy was observed during 26 weeks. In the ticlopidine-treated nephrotic animals (TNG), proteinuria was less than in the untreated nephrotic animals (NG), but this difference was significant only at week 6 (TNG = 47.27 +/- 16.52 versus NG = 100.08 +/- 13.83 mg/24 h, p < 0.01) and week 26 (TNG = 157.00 +/- 28.73 versus NG = 217.00 +/- 21.73 mg/24 h, p < 0.01) after ADR injection. NG presented severe tubulointerstitial abnormalities with a tubulointerstitial lesion index of 3+. No difference in glomerular lesions was observed among the groups (NG median = 6%, TNG median = 4% and TCG median = 2%). The tubulointerstitial lesion index of TNG was less intense (median = 2+) but not different from those of the control groups (CG median = 1+; TCG median = 0+) nor NG (median = 3+). We concluded that the treatment with ticlopidine produced some partially beneficial effects but did not prevent the development of adriamycin-induced nephropathy. PMID- 10516991 TI - An animal model of septicemia-induced hypercatabolic acute renal failure. AB - A rat model of hypercatabolic acute renal failure (ARF) was developed in order to further investigate the mechanism of this condition. Sprague Dawley rats were separated into three groups: a septicemic group, an ischemic ARF group, and a hypercatabolic ARF group. Septicemia was produced by the i.p. injection of 1 x 10(7) colony-forming units/mL of Escherichia coli. Ischemic ARF was induced by 60 minutes clamping of the left renal artery following a contralateral nephrectomy. Hypercatabolic ARF was produced by combining ischemic ARF with the i.p. injection of 1 x 10(7) colony-forming units/mL of Escherichia coli. The hypercatabolic ARF group exhibited septic clinical features after the surgical procedures. The blood urea nitrogen and the serum creatinine, potassium and carbon dioxide combining power of hypercatabolic ARF were significantly higher than other two groups 24 hours after surgery. In addition, the rats wit hypercatabolic ARF had a greater loss of body weight and a higher mortality rate compared to the other two groups. The features of this form of experimental ARF are similar to the clinical characteristics of hypercatabolic ARF. Consequently, this appears to be a useful model of hypercatabolic ARF. PMID- 10516992 TI - Antierythrocyte antibodies in hemodialysis and kidney transplant patients. AB - Natural antierythrocytic antibodies may be stimulated by bacterial antigens and the immune type may occur as a result of pregnancy or blood transfusions. The prevalence increases with the number of red cell units transfused. Specificity, on the other hand, depends on ethnic backgrounds. The clinical importance of these antibodies is to precipitate hemolytic transfusion reactions and erythroblastosis fetalis. Hemodialysis patients are multitransfused and have a quite variable prevalence of antibodies. Kidney transplant patients with blood group identity do not form antibodies. We studied the presence of both types of antierythrocytic antibodies (natural and immune) in hemodialysis and kidney transplant patients in Brazilian blood transfusion and nephrology services. PMID- 10516993 TI - Treatment of children with steroid refractory idiopathic nephrotic syndrome: the Kuwaiti experience. AB - Data on the treatment and outcome of Kuwaiti children with steroid refractory idiopathic glomerulonephritis (SRIGN), i.e. nephrotic syndrome who failed an eight-week course of prednisone, were collected retrospectively from the records of children attending the two renal centers of Kuwait between January 1, 1990 to December 31, 1996. During those seven years, a total of 34 Kuwaiti children were diagnosed to have SRIGN. Histologically, 22 (65%) of those patients had minimal change, 5 (15%) focal segmental GN, 2 (6%) non-IgA mesangioproliferative GN and one membranous GN. Twenty-two patients had manifested frequent relapses, six were steroid-dependent and six were steroid-resistant. Treatment options were in the following order: (a) small maintenance-dose of corticosteroids (< 0.5 mg/kg/alternate days); (b) cyclophosphamide and or chlorambucil for a single eight week-course or eight then 12 week courses (c) cyclosporin A for three months. The response to therapy was as follows: nine children were cured with low dose corticosteroids; 17 with chlorambucil and/or cyclophosphamide; and five with cyclosporin A. At the end of study, only three children failed such drug therapy, two of who had focal segmental glomerulosclerosis. PMID- 10516994 TI - Impairment of ventilatory response to metabolic acidosis in insulin-dependent diabetic patients with advanced nephropathy. AB - Sudden cardiopulmonary arrest due to a defective respiratory reflex is observed in diabetic patients. Impaired ventilatory response in diabetic patients to acute hypoxia or hypercapnia induced by the inhalation of an artificial gas has been reported. Little is known regarding the respiratory compensatory ability for mild to moderate metabolic acidosis due to renal failure in insulin-dependent diabetic subjects. Arterial blood pH, HCO3-, PaCO2 and PaO2 were measured in 13 insulin dependent diabetic subjects with advanced nephropathy and in 33 non-diabetic subjects with end-stage renal failure. The diabetic group consisted of six predialysis patients and seven on regular hemodialysis (HD) and the non-diabetic group, ten predialysis patients and 23 on HD. Differences between measured partial arterial pressure of carbon dioxide (PaCO2) and predicted PaCO2 determined from HCO3- were examined. PaCO2 was significantly higher in the diabetic than in non-diabetic group (40.0 +/- 7.4 versus 31.1 +/- 5.1 mmHg, p < 0.05 in predialysis, 42.0 +/- 6.4 versus 36.0 +/- 2.6 mmHg, p < 0.05 in HD), though plasma pH was essentially the same for either. Differences in measured PaCO2 and predicted PaCO2 were significantly larger in the diabetic group than in non-diabetic group. Ventilatory response to uremic acidosis may thus be considered impaired in subjects with advanced diabetic nephropathy. PMID- 10516995 TI - Plasma, urinary, and erythrocyte concentrations of guanidino compounds in patients with chronic renal failure. AB - Guanidino compounds are among the most likely candidates for uremic toxins. We determined the plasma, erythrocyte, and urinary concentration of guanidino compounds in 30 hemodialysis patients and 15 patients with chronic renal failure who had not undergone hemodialysis. Guanidino compounds were measured by high performance liquid chromatography. Plasma levels of taurocyamine, guanidinosuccinic acid, alpha-N-acetyl-L-arginine, creatine, guanidinobutyric acid, guanidine, and methylguanidine were significantly increased in patients with chronic renal failure with or without hemodialysis. In contrast, plasma guanidinoacetic acid concentrations were significantly decreased. Erythrocyte concentrations of creatinine, guanidinosuccinic acid, guanidine and methylguanidine were also markedly elevated. No correlation was observed between plasma creatinine concentration and erythrocyte concentration of guanidinosuccinic acid or methylguanidine. However, there was a significant correlation between plasma and erythrocyte methylguanidine, and between plasma and erythrocyte guanidinosuccinic acid. PMID- 10516996 TI - Accelerated hemodialysis: a new safe and simple method of a high filter-low patient blood flow rate hemodialysis. AB - Accelerated hemodialysis (AHD) is a new safe method of hemodialysis that is done using accelerated hemodialysis blood lines (registered). These lines allow partial controlled recirculation of blood through a recirculation segment. AHD allows increase in the filter blood flow without increasing the patient blood flow and thus allows the use of larger filters of higher efficiency, and provides a high blood flow in most parts of the dialysis circuit to permit heparin free dialysis. On the other hand, the patient blood flow can be decreased without increasing the filter blood flow and allows safe sessions for low body weight patients and those with hemodynamic instability or inefficient vascular assess. Twenty-five hemodialysis sessions were conducted with an adolescent girl, 13 years old, 30 kg body weight, on a chronic hemodialysis program. The sessions included: five double-needle hemodialysis sessions (DNHD) with a pediatric filter at a rate of 130 mL/min; five accelerated hemodialysis sessions (AHD) using the same filter at a rate of (65 + 65) mL/min and another five sessions at rate of (130 + 130) mL/min; five (DNHD) were done using an adult hemodialysis filter rate of 130 mL/min; and five (AHD) using the same filter at a rate of (130 + 130) mL/min. In all sessions pre- and post-dialysis levels of BUN and creatinine were measured and post/predialysis ratios were calculated. In AHD the BUN and creatinine level in the patient, and filter segments of the arterial line and in the recirculation segment were measured and the actual patient blood flow was calculated. The efficiency of AHD was decreased by decreasing the actual patient blood flow (p < 0.05) but was not increased by increasing the filter blood flow in both pediatric and adult filters (p > 0.05). There were no significant changes between the intended patient blood flow rate and the actual patient blood flow as calculated from BUN and creatinine levels. The AHD is a simple safe and applicable mode of hemodialysis that needs further studies to verify factors affecting its efficiency and to approve its clinical applications. PMID- 10516997 TI - Acute effects of acetaminophen on renal function and urinary excretion of some proteins and enzymes in patients with kidney disease. AB - The acute effects of acetaminophen, a commonly used as analgesic drug, upon the urinary excretion of some proteins and enzymes as markers of kidney damage, was investigated. Patients with chronic glomerulonephritis (GN) and Balkan endemic nephropathy (BEN), having kidney vulnerable to toxic drugs, were enrolled in the study. Timed urine specimens were collected: before drug administration, and in 3 hour periods for 24 hours after an oral dose of 2 g of acetaminophen. Urinary excretion of albumin before acetaminophen treatment was significantly higher in patients with GN and BEN than in healthy adults, however, beta 2-microglobulin excretion was increased in BEN patients only. Urinary excretion of creatinine markedly increased immediately after acetaminophen ingestion. Urinary excretion of total protein and albumin was not changed after acetaminophen administration. However, acetaminophen treatment produced a significant increase in beta 2 microglobulin excretion in patient with BEN and GN, and in clinically healthy members of nephropathic families. Excretion of aminopeptidase N (APN) activity before acetaminophen treatment was significantly higher in patients with GN, however, NAGA excretion was higher in both GN and BEN patients than in healthy controls. After acetaminophen administration urinary excretion of APN, dipeptidylpeptidase IV (DPP IV), gamma-glutamyltranspeptidase (GGT) and N-acetyl beta-D-glucosaminidase (NAGA) did not increase significantly in any group studied. This study has shown that urinary excretion of APN, DPP IV, NAGA and GGT, as markers of kidney brush border and lysosomal damage, did not change after 2 g of acetaminophen taken orally. beta 2-microglobulin was a marker of acute acetaminophen nephrotoxicity in kidney disease patients and in clinically healthy adults from nephropathic families. PMID- 10516998 TI - Dialysis and pregnancy--a case report and review of the literature. AB - We report on a patient with an eight-year history on maintenance hemodialysis treatment without residual renal function in whom pregnancy was successfully managed through to the 29th week. During this time, under carefully modified dialysis treatment, the nephrologic course, as well as materno-fetal flow relationships were unremarkable. Fetal development was appropriate for gestational age. However, pregnancy was complicated by polyhydramnios, which necessitated i.v. tocolysis. In the 28 + 6th week of gestation, cesarean section was performed because of an antibiotic-resistant fever of unclear origin which ceased within two days of delivery. Although the postnatal course of the adequately developed baby was complicated by the respiratory distress syndrome, normal development continued. We emphasize that the intensive interdisciplinary cooperation of nephrologists and obstetricians is imperative for the successful management of pregnancy under these conditions. In these pregnancies, the main fetal problems consist of premature labor because of polyhydramnios, preterm delivery, intrauterine growth retardation and stillbirth. The mother is threatened by the development of superimposed pre-eclampsia, left ventricular failure because of volume overload and progressive anemia. In order to maintain a well-balanced homeostasis, intensification of dialysis therapy by an increase in frequency and duration is the most important therapeutic approach. Accurate fetal monitoring including frequent examination of the feto-maternal circulation by Doppler sonography as well as attentive surveillance of the mother is required to recognize the above mentioned complications. PMID- 10516999 TI - Acute hyperphosphatemia caused by sodium phosphate enema in a patient with liver dysfunction and chronic renal failure. AB - We report a case of acute hyperphosphatemia secondary to rectal administration of sodium phosphate and sodium biphosphate (Fleet enema). Parathyroid hormone and calcitonin levels were measured along with phosphate clearance and the tubular reabsorption of phosphate. PMID- 10517000 TI - Acute renal failure due to nontraumatic rhabdomyolysis following binge drinking. AB - Nontraumatic rhabdomyolysis is an important but under-recognized cause of acute renal failure. In alcoholics, rhabdomyolysis most frequently develop following muscle necrosis during alcohol-induced coma, but has also been described rarely in those without prolonged coma or seizures. We describe a patient who developed myoglobinuric acute renal failure requiring dialysis following binge drinking in the absence of convulsions or coma. The renal biopsy showed acute tubular necrosis with pigment casts. PMID- 10517001 TI - Well-preserved cholinergic neuronal activity in the brain cortex of the severely uremic rats. PMID- 10517002 TI - Total synthesis and chemical biology of the sarcodictyins. AB - The sarcodictyins A-F and eleutherobin comprise a family of marine-derived diterpenoids with potent cytotoxicities against various tumor cell lines. Investigations have revealed that several of these compounds exert their cytotoxic effects through tubulin binding in a mechanism analogous to that of the clinical anticancer drug Taxol. The biological importance, challenging molecular architecture, and relative scarcity of these natural products have prompted several groups to undertake their total chemical synthesis. In this review, we summarize the current synthetic efforts and examine the preliminary structure activity relationships which have emerged from early combinatorial libraries. PMID- 10517004 TI - Synthesis and in vitro antiprotozoal activity of thiophene ring-containing quinones. AB - A series of quinones (3a-i, 4-9, 11) and aromatic compounds (2a, 2d, 2g) containing the thiophene ring were tested in vitro against the trypomastigote form of Trypanosoma cruzi and the promastigote forms of Leishmania. The quinones 3a-i, 4, 5a, b, 6 and 9 having the thiophene ring fused to a quinone nucleus were the most active members of the series. The electron affinities of the benzo[b]thiophene-4,7-quinones 3, evaluated by their LUMO energies and halfwave potentials, are reported. PMID- 10517003 TI - Isolation and structural determination of new sphingolipids and pharmacological activity of africanene and other metabolites from Sinularia leptoclados. AB - Two new sphingolipids, (2S,3S,4R)-1,3,4-trihydroxy-2-[((R)-2' hydroxytetradecanoyl) amino] tricosane (4) and (2S,3S,4R)-1,3,4-triacetoxy-2 [((R)-2'-acetoxyoctadecanoyl) amino]octadecane (5) along with africanene (1, reasonably good yield), 23-demethylgargosterol (2) and batylalcohol (3) have been isolated from the soft coral Sinularia leptoclados. Preliminary studies for pharmacological activity (blind screening and toxicity studies) of africanene were conducted. Africanene exhibited in vitro and in vivo cytotoxicity, dose dependent hypotensive activity as well as antiinflammatory activity. The pharmacological and toxicity studies on africanene are being reported for the first time and findings strongly encourage further investigation. Compounds 1, 4 and 5 were studied for the antibacterial, antifungal and antiviral activity while compounds 4 and 5 were also studied for the short term in vitro cytotoxic activity. PMID- 10517005 TI - Synthesis and pharmacological evaluation of 4-halo progesterone derivatives as antiandrogen. AB - The pharmacological activity of eight pregnane derivatives 17-alpha acetoxyprogesterone 9, 17-alpha acetoxy-4, 5-epoxypregnan-3, 20-dione 10, 17 alpha acetoxy-4-chloro-4-pregnene-3, 20-dione 11, 17-alpha acetoxy-4-bromo-4 pregnene-3, 20-dione 12, 17-alpha hydroxy-4-bromo-4-pregnene-3, 20-dione 13, 4 chloro-17-alpha hydroxy-4-pregnene-3, 20-dione 14, 17-alpha benzoyloxy-4-bromo-4 pregnene-3, 20-dione 15 and 17-alpha benzoyloxy-4-chloro-4-pregnene-3, 20-dione 16 was determined. These compounds were evaluated as antiandrogens on gonadectomized hamster seminal vesicles. The pharmacological data in this study indicate that compounds 15 and 16 having a C-17 benzoyloxy moiety showed the highest antiandrogenic activity as measured by the reduction of the weight of the seminal vesicles, followed by the steroids 11 and 12 (17-alpha acetoxy group). The free alcohols 13 and 14 exhibited a lower antiandrogenic activity. Apparently, the ester moiety at C-17 is a necessary requirement for the presence of high antiandrogenic activity. Shows the inhibitory effect on the conversion of testosterone (T) to DHT, of the above described steroids as measured by the amount of produced DHT 2 expressed as pmoles of DHT/g of protein/h. Steroids 11, 12 and 16 showed a much higher inhibitory activity on the conversion of testosterone (T) to dihydrotestosterone (DHT) than presently used finasteride 3. PMID- 10517006 TI - Syntheses and evaluation of biantennary oligosaccharide ligands mimicking sialyl Lewis X. AB - Sialyl Lewis X (1) is known to be a ligand of the cell adhesion molecule E selectin. We have synthesized several biantennary glycoside-terminated ligands mimicking sialyl Lewis X (1), and evaluated their binding activity to E-selectin using HL-60 cells expressing sialyl Lewis X epitope and human umbilical vein endothelial cells (HUVECs). These compounds were found to possess moderate binding activities to E-selectin. Among them, di-fucoside analog (8) which has no sialic acid carboxylate group was more active than 2, which had both the sialyl galactose residue and the fucose residue (IC50, 8: 4.7 mM, 2: 11.7 mM). Furthermore, in the rat pleuritic model in vivo induced by carrageenin, 8 was found to reduce neutrophil infiltration at inflammatory lesions. PMID- 10517007 TI - Synthetic studies on glycosphingolipids from protostomia phyla: synthesis of neogala-series glycolipid analogues containing a mannose residue from the earthworm Pheretima hilgendorfi. AB - Two kinds of glycosphingolipid analogues from the earthworm Pheretima hilgendorfi were synthesized as follows: the trisaccharide 2-(tetradecyl)hexadecyl alpha-D mannopyranosyl-(1-->4)-beta-D-galactopyranosyl-(1-->6)-beta-D- galactopyranoside (13) and the tetrasaccharide 2-(tetradecyl) hexadecyl alpha-D-galactopyranosyl-(1 ->6)-[alpha-D-mannopyranosyl-(1-->4)]-beta-D - galactopyranosyl-(1-->6)-beta-D galactopyranoside (20) were synthesized by stepwise condensation of suitably protected monosaccharide units. A 2-(trimethylsilyl)ethyl 2,3,4-tri-O-benzoyl beta-D-galactopyranoside derivative (5) was used as the glycosyl acceptor and thiophenyl derivatives of D-galactose and D-mannose were used as donors respectively. PMID- 10517008 TI - Total synthesis of a natural antioxidant and structure-activity relationships of related compounds. AB - A total synthesis of benzodioxole derivative 1 was achieved via a palladium(0) catalyzed cross-coupling reaction in a 68% overall yield (4 steps). A novel series of benzodioxoles bearing a variety of aromatic and heterocyclic rings was also prepared and the antioxidative activity evaluated using in vitro model systems. Structure-activity studies revealed that i) intramolecular hydrogen bonding in the phenol moiety reduced activity, ii) introduction of disubstituents at the ortho location relative to the phenol increased activity, and iii) the methylenedioxy function contributed to stabilization of the phenoxy radical. Among of these compounds, 5,7-di-(4-methoxyphenyl)-4-methoxy-6-hydroxy-1,3 benzodioxole (7p) was the most favorable agent and more potent than n-propyl gallate. PMID- 10517009 TI - Synthetic studies of an 18-membered antitumor macrolide, tedanolide. 5. Stereoselective synthesis of the C13-C23 part via condensation of two fragments, C13-C17 and C18-C21, by taking advantage of the 3,4-dimethoxybenzyl protecting group. AB - An efficient and stereoselective synthesis of the C13-C23 part (8) was achieved starting from methyl (R)- and (S)-3-hydroxy-2-methylpropionates (9) via coupling of the C13-C17 aldehyde (6), prepared by Evans asymmetric aldol reaction, with the C18-C21 iodoalkene (5b) by taking advantage of the 3,4-dimethoxybenzyl protecting group. PMID- 10517010 TI - Effect of grinding with hydroxypropyl cellulose on the dissolution and particle size of a poorly water-soluble drug. AB - A new benzofuroquinoline derivative, 3,9-bis(N,N-dimethylcarbamoyloxy)-5H benzofuro[3,2-c]quinoli ne-6-one (KCA-098), shows poor oral absorption due to practical insolubility in water. In this study, a co-grinding technique employing a water-soluble polymer was used for improvement of the dissolution rate of KCA 098. Powder X-ray diffraction patterns and IR spectra of KCA-098 showed the conversion of the drug from a crystal state to an amorphous state by grinding with a polymer such as hydroxypropyl cellulose (HPC-SL) or polyvinylpyrrolidone (PVP K30). The particle size of KCA-098 was remarkably reduced to a submicron size by grinding with HPC-SL. The co-ground mixture with HPC-SL showed a rapid dissolution rate and maintained supersaturation for more than 1 h. On the other hand, the co-ground mixture with PVP K30 showed rapid dissolution and supersaturation for a shorter period. These data suggest that the rapid dissolution rate was obtained by the conversion of the drug particles from a crystal to amorphous state by grinding with water-soluble polymers and that a reduction in particle size to the submicron level led to the maintenance of supersaturation due to good dispersion. PMID- 10517011 TI - Synthesis and antiviral and antineoplastic activities of some novel carbocyclic guanosine analogues with a cyclobutane ring. AB - Cyclobutyl nucleoside analogues containing guanine and 8-azaguanine (compounds 5 10) were prepared from (1R,cis)-3-aminomethyl-2,2-dimethylcyclobutylmethanol (1). All were evaluated as antiviral and antitumoral agents in a variety of assay systems. Compounds 6 and 7 showed a noteworthy activity against a respiratory syncytial virus and compound 10 was moderately active against vaccinia virus. Only compound 5 showed some cytostatic activity. PMID- 10517012 TI - Synthesis of fluorine analogs of protoporphyrin potentially useful for diagnosis and therapy of cancer. IV. Synthesis of (trifluorovinyl)vinyl- and (1-chloro-2,2 difluorovinyl)vinyldeuteroporphyrins. AB - Trifluoro or chlorodifluoro analogs of protoporphyrin, the compounds in the title, were synthesized for use in the diagnosis and therapy of cancer. 3- Or 8 acetyldeuteroporphyrin dimethyl esters (2 and 3) were iodinated with iodine in the presence of potassium carbonate to the corresponding iodo compounds (5 and 6). The iodo compounds (5 and 6) were treated with bis(trifluorovinyl)zinc in the presence of tetrakis(triphenylphosphine)-palladium to give trifluorovinyl derivatives (7 and 8) in good yields. Reduction of the acetyl group of 7 and 8 with sodium borohydride afforded the corresponding hydroxyethyl derivatives (9 and 10). Compounds (9 and 10) were dehydrated with methanesulfonyl chloride and triethylamine to give (trifluorovinyl)vinyldeuteroporphyrin dimethyl esters (11 and 12). Treatment of 5 and 6 with bis(1-chloro-2,2-difluorovinyl)zinc in the presence of tetrakis(triphenylphosphine)palladium, followed by similar reactions as above gave (1-chloro-2,2-difluorovinyl)-vinyldeuteroporphyrin dimethyl esters (17 and 18). PMID- 10517013 TI - Synthesis of extremely simplified compounds possessing the key pharmacophore units of taxol, phenylisoserine and oxetane moieties. AB - Straight chain compounds having a phenylisoserine unit and an oxetane ring at the alpha- and omega- position, respectively as extremely simplified analogues of taxol were prepared. None of these compounds showed promising tubulin inhibitory activity. PMID- 10517015 TI - A novel sesterterpenoid, nitiol, as a potent enhancer of IL-2 gene expression in a human T cell line, from the Peruvian folk medicine "Hercumpuri" (Gentianella nitida). AB - A novel sesterterpenoid designated as nitiol (1), possessing enhancement activity of IL-2 gene expression in a human T cell line, was isolated from the Peruvian folk medicine "Hercampuri" (Gentianella nitida). The structure was elucidated by extensive spectroscopic investigation. PMID- 10517014 TI - A collagen network formation effector from leaves of Premna subscandens. AB - As a part of the search for biologically active plant products, M cells, which form a collagen fiber network in vitro after a prolonged culture period, were used. The n-BuOH-soluble fraction of a methanol extract of leaves of Premna subscandens exhibited promotion of collagen network formation by M cells. Extensive isolation work guided by a bioassay afforded a phenylethanoid, acteoside, as an active compound. PMID- 10517016 TI - 12-O-acetylphorbol-13-decanoate potently inhibits cytopathic effects of human immunodeficiency virus type 1 (HIV-1), without activation of protein kinase C. AB - Through bioactivity-guided fractionation, eight phorbol diesters, including five new ones (1-5), were isolated from the seeds of Croton tiglium collected in Egypt. 12-O-Acetylphorbol-13-decanoate (6) and 12-O-decanoylphorbol-13-(2 methylbutyrate) (4) potently inhibited the HIV-1-induced cytopathic effect on MT 4 cells (IC100 values of 7.6 ng/ml and 7.81 micrograms/ml, and CC0 values of 62.5 micrograms/ml and 31.3 micrograms/ml, respectively) without activating protein kinase C. PMID- 10517017 TI - Thiazolidinediones with thyroid hormone receptor agonistic activity. AB - Several thiazolidinedione derivatives (3-7) were designed and synthesized as candidate thyromimetic drugs. Among them, the dihydrogenated compounds, such as 5 2-[[4-(3-tert-butyl-4-hydroxyphenyl)oxy-3,5-diiodophenyl] ethyl]-2,4 thiazolidinedione (6b) and its 3-isopropyl analog (7b), exhibited potent thyroid hormone receptor alpha 1 (TR alpha 1) activation activity. PMID- 10517018 TI - Relationship between acuity for gratings and for tumbling-E letters in peripheral vision. AB - Earlier studies have reported that grating resolution is sampling-limited in peripheral vision but that letter acuity is generally poorer than grating acuity. These results suggest that peripheral resolution of objects with rich Fourier spectra may be limited by some factor other than neural sampling. To examine this suggestion we formulated and tested the hypothesis that letter acuity in the periphery is sampling-limited, just as it is for extended and truncated gratings. We tested this hypothesis with improved methodology to avoid the confounding factors of target similarity, alphabet size, individual variation, peripheral refractive error, and stimulus size. Acuity was measured for an orientation discrimination task (horizontal versus vertical) for a three-bar resolution target and for a block-E letter in which all strokes have the same length. We confirmed previous reports in the literature that acuity for these targets is worse than for extended sinusoidal gratings. To account for these results quantitatively, we used difference-spectrum analysis to identify those frequency components of the targets that might form a basis for performing the visual discrimination task. We find that discrimination performance for the three-bar targets and the block-E letters can be accounted for by a sampling-limited model, provided that the limited number of cycles that are present in the characteristic frequency of the stimulus is taken into account. Quantitative differences in acuity for discriminating other letter pairs (e.g., right versus left letters E or characters with short central strokes) could not be attributed to undersampling of either the characteristic frequency or the frequency of maximum energy in the difference spectrum. These results suggest additional tests of the sampling theory of visual resolution, which are the subject of a companion paper. PMID- 10517019 TI - Sampling limits and critical bandwidth for letter discrimination in peripheral vision. AB - We develop and test two functional hypotheses based on the sampling theory of visual resolution that might account for letter acuity in peripheral vision. First, a letter smaller than the acuity limit provides insufficient veridical energy for performing the task, and, second, the available veridical energy is masked by increased amounts of visible but aliased energy. These two hypotheses make opposite predictions about the effect of low-pass filtering on letter acuity, which we tested experimentally by using filtered letters from the tumbling-E alphabet. Our results reject the masking hypothesis in favor of the energy insufficiency hypothesis. Additional experiments in which high-pass filtered letters were used permitted the isolation of a critical band of spatial frequencies, which is necessary and sufficient for achieving maximum visual acuity. This critical band varied with the particular pair of letters to be discriminated but was in the range 0.9-2.2 cycles per letter. PMID- 10517020 TI - Comparison of cone directionality determined by psychophysical and reflectometric techniques. AB - We measured the directionality of the cones with both a psychophysical (Stiles Crawford I) technique and an optical technique. The two sets of measurements were made in the same subjects, with stimuli as similar as possible used. The two types of measurements gave similar estimates of the location in the pupil toward which the cones were optimally aligned. However, the two measurements gave quite dissimilar estimates of the width of the directional sensitivity. On average, optical measurements were half as broad as psychophysical measurements in the fovea, but there were substantial individual differences. At 2-deg retinal eccentricity the difference between techniques was even more marked. PMID- 10517021 TI - Predicting color from gray: the relationship between achromatic adjustment and asymmetric matching. AB - Achromatic adjustment has been used widely to study color context effects. In the achromatic adjustment procedure, an observer adjusts a test stimulus until it appears black, gray, or white. By its nature, achromatic adjustment directly measures the effect of context only for stimuli that appear gray. We present achromatic loci measured in two contexts and asymmetric color matches measured across the same two contexts. The results indicate that achromatic adjustments, together with a gain-control model, may be used to make accurate predictions of the chromaticity of asymmetric matches. Thus measurements of the effect of context for test stimuli that appear gray may be used to predict the effect of context for stimuli that appear colored. The experiments also indicate that accurate prediction depends on ensuring that observers use similar fixational strategies for the two judgments. PMID- 10517022 TI - Parametric blind deconvolution: a robust method for the simultaneous estimation of image and blur. AB - Blind-deconvolution microscopy, the simultaneous estimation of the specimen function and the point-spread function (PSF) of the microscope, is an underdetermined problem with nonunique solutions that are usually avoided by enforcing constraints on the specimen function and the PSF. We derived a maximum likelihood-based method for blind deconvolution in which we assume a mathematical model for the PSF that depends on a small number of parameters (e.g., less than 20). The algorithm then estimates the unknown parameters together with the specimen function. The mathematical model ensures that all the constraints of the PSF are satisfied, and the maximum-likelihood approach ensures that the specimen is nonnegative. The method successfully estimates the PSF and removes out-of focus blur. The PSF estimation is robust to aberrations in the PSF and to noise in the image. PMID- 10517023 TI - Modified minimum-distance criterion for blended random and nonrandom encoding. AB - Two pixel-oriented methods for designing Fourier transform holograms- pseudorandom encoding and minimum-distance encoding-usually produce higher fidelity reconstructions when combined than those produced by each method individually. In previous studies minimum-distance encoding was defined as the mapping from the desired complex value to the closest value produced by the modulator. This method is compared with a new minimum-distance criterion in which the desired complex value is mapped to the closest value that can be realized by pseudorandom encoding. Simulations and experimental measurements using quantized phase and amplitude modulators show that the modified approach to blended encoding produces more faithful reconstructions than those of the previous method. PMID- 10517024 TI - The role of liquid chromatography-mass spectrometry in the determination of heroin and related opioids in biological fluids. AB - The opioid most commonly sold in the illicit market is heroin. This substance, classified as an analgesic narcotic drug, has an extremely short half-life, and it is rapidly metabolized to 6-monoacetyl-morphine and further to morphine. Morphine is principally metabolized by conjugation to morphine-3 and morphine-6 glucuronides. Morphine itself is a potent analgesic that is frequently used in the pharmacological intervention of cancer pain. The toxicological and clinical evaluation of heroin and morphine have stimulated pharmacokinetic studies in human and animal models. Although a number of methods exist to determine opiates and their metabolites, liquid chromatography (LC) appears to be the technique that can separate without any pretreatment the lipophilic and the hydrophilic analytes of the complete metabolic profile of heroin and/or morphine. Moreover, mass spectrometry (MS) used as a detector for liquid chromatography is unique, because it offers universality and selectivity. Furthermore, efforts have been made to develop LC/MS interfaces that could overcome the previous problem of poor sensitivity. For this reason, in recent years LC combined with MS has been applied to the analysis of opiates--parent drugs and metabolites--in biological fluids. This article reviews the existing literature on the determination, using liquid chromatography coupled to mass spectrometry, of opiate metabolites found in different biological matrices after the administration of the parent compounds. PMID- 10517025 TI - Tomatinase from Fusarium oxysporum f. sp. lycopersici defines a new class of saponinases. AB - Plants produce a variety of secondary metabolites, many of which have antifungal activity. Saponins are plant glycosides that may provide a preformed chemical barrier against phytopathogenic fungi. Fusarium oxysporum f. sp. lycopersici and other tomato pathogens produce extracellular enzymes known as tomatinases, which deglycosylate alpha-tomatine to yield less toxic derivatives. We have cloned and characterized the cDNA and genomic DNA encoding tomatinase from the vascular pathogen of tomato F. oxysporum f. sp. lycopersici. This gene encodes a protein (FoTom1) with no amino acid sequence homology to any previously described saponinase, including tomatinase from Septoria lycopersici. Although FoTom1 is related to family 10 glycosyl hydrolases, which include mainly xylanases, it has no detectable xylanase activity. We have overexpressed and purified the protein with a bacterial heterologous system. The purified enzyme is active and cleaves alpha-tomatine into the less toxic compounds tomatidine and lycotetraose. Tomatinase from F. oxysporum f. sp. lycopersici is encoded by a single gene whose expression is induced by alpha-tomatine. This expression is fully repressed in the presence of glucose, which is consistent with the presence of two putative CREA binding sites in the promoter region of the tomatinase gene. The tomatinase gene is expressed in planta in both roots and stems throughout the entire disease cycle of F. oxysporum f. sp. lycopersici. PMID- 10517026 TI - A novel class of ectomycorrhiza-regulated cell wall polypeptides in Pisolithus tinctorius. AB - Development of the ectomycorrhizal symbiosis leads to the aggregation of fungal hyphae to form the mantle. To identify cell surface proteins involved in this developmental step, changes in the biosynthesis of fungal cell wall proteins were examined in Eucalyptus globulus-Pisolithus tinctorius ectomycorrhizas by two dimensional polyacrylamide gel electrophoresis. Enhanced synthesis of several immunologically related fungal 31- and 32-kDa polypeptides, so-called symbiosis regulated acidic polypeptides (SRAPs), was observed. Peptide sequences of SRAP32d were obtained after trypsin digestion. These peptides were found in the predicted sequence of six closely related fungal cDNAs coding for ectomycorrhiza up regulated transcripts. The PtSRAP32 cDNAs represented about 10% of the differentially expressed cDNAs in ectomycorrhiza and are predicted to encode alanine-rich proteins of 28.2 kDa. There are no sequence homologies between SRAPs and previously identified proteins, but they contain the Arg-Gly-Asp (RGD) motif found in cell-adhesion proteins. SRAPs were observed on the hyphal surface by immunoelectron microscopy. They were also found in the host cell wall when P. tinctorius attached to the root surface. RNA blot analysis showed that the steady state level of PtSRAP32 transcripts exhibited a drastic up-regulation when fungal hyphae form the mantle. These results suggest that SRAPs may form part of a cell cell adhesion system needed for aggregation of hyphae in ectomycorrhizas. PMID- 10517027 TI - Naturally induced secretions of the potato cyst nematode co-stimulate the proliferation of both tobacco leaf protoplasts and human peripheral blood mononuclear cells. AB - Naturally induced secretions from infective juveniles of the potato cyst nematode Globodera rostochiensis co-stimulate the proliferation of tobacco leaf protoplasts in the presence of the synthetic phytohormones alpha naphthaleneacetic acid (NAA) and 6-benzylaminopurine (BAP). With the use of a protoplast-based bioassay, a low-molecular-weight peptide(s) (< 3 kDa) was shown to be responsible for the observed effect. This mitogenic oligopeptide(s) is functionally dissimilar to auxin and cytokinin and, in addition, it does not change the sensitivity of the protoplasts toward these phytohormones. In combination with the mitogen phytohemagglutinin (PHA), cyst nematode secretions also co-stimulated mitogenesis in human peripheral blood mononuclear cells (PBMC). The stimulation of plant cells isolated from nontarget tissue--these nematodes normally invade the roots of potato plants--suggests the activation of a general signal transduction mechanism(s) by an oligopeptide(s) secreted by the nematode. Whether a similar oligopeptide-induced mechanism underlies human PBMC activation remains to be investigated. Reactivation of the cell cycle is a crucial event in feeding cell formation by cyst nematodes. The secretion of a mitogenic low-molecular-weight peptide(s) by infective juveniles of the potato cyst nematode could contribute to the redifferentiation of plant cells into such a feeding cell. PMID- 10517028 TI - The alfalfa (Medicago sativa) TDY1 gene encodes a mitogen-activated protein kinase homolog. AB - Development of root nodules, specifically induction of cortical cell division for nodule initiation, requires expression of specific genes in the host and microsymbiont. A full-length cDNA clone and the corresponding genomic clone encoding a MAP (mitogen-activated protein) kinase homolog were isolated from alfalfa (Medicago sativa). The genomic clone, TDY1, encodes a 68.9-kDa protein with 47.7% identity to MMK4, a previously characterized MAP kinase homolog from alfalfa. TDY1 is unique among the known plant MAP kinases, primarily due to a 230 amino acid C-terminal domain. The putative activation motif, Thr-Asp-Tyr (TDY), also differs from the previously reported Thr-Glu-Tyr (TEY) motif in plant MAP kinases. TDY1 messages were found predominantly in root nodules, roots, and root tips. Transgenic alfalfa and Medicago truncatula containing a chimeric gene consisting of 1.8 kbp of 5' flanking sequence of the TDY1 gene fused to the beta glucuronidase (GUS) coding sequence exhibited GUS expression primarily in the nodule parenchyma, meristem, and vascular bundles, root tips, and root vascular bundles. Stem internodes stained intensely in cortical parenchyma, cambial cells, and primary xylem. GUS activity was observed in leaf mesophyll surrounding areas of mechanical wounding and pathogen invasion. The promoter was also active in root tips and apical meristems of transgenic tobacco. Expression patterns suggest a possible role for TDY1 in initiation and development of nodules and roots, and in localized responses to wounding. PMID- 10517029 TI - Maize streak virus coat protein is karyophyllic and facilitates nuclear transport of viral DNA. AB - Transport of maize streak virus (MSV) DNA into the nucleus of host cells is essential for virus replication and the presence of virus particles in the nuclei of infected cells implies that coat protein (CP) must enter the nucleus. To see if CP is imported into the nucleus in the absence of other viral gene products, the MSV CP gene was expressed in insect cells with a baculovirus vector system, and also in tobacco protoplasts with a cauliflower mosaic virus (CaMV) 35S promoter-driven transient gene expression vector. Immunofluorescent staining showed that the CP accumulated in the nuclei of both insect and tobacco cells. Mutagenesis of a potential nuclear localization signal in the CP resulted in cytoplasmic accumulation of the mutant protein. We have shown previously that the CP binds to single-stranded (ss) and double-stranded (ds) viral DNA. To investigate if CP might also be involved in viral DNA nuclear transport, Escherichia coli-expressed CP, together with TOTO-1-labeled viral ss or ds DNA, was microinjected into maize and tobacco epidermal cells. Both ss and ds DNA moved into the nucleus when co-injected with the CP but not with E. coli proteins alone. These results suggest that, in addition to entering the nucleus where it is required for encapsidation of the viral ss DNA, the MSV CP facilitates the rapid transport of viral (ss or ds) DNA into the nucleus. PMID- 10517030 TI - CFP, the putative cercosporin transporter of Cercospora kikuchii, is required for wild type cercosporin production, resistance, and virulence on soybean. AB - Many species of the fungal genus Cercospora, including the soybean pathogen C. kikuchii, produce the phytotoxic polyketide cercosporin. Cercosporin production is induced by light. Previously, we identified several cDNA clones of mRNA transcripts that exhibited light-enhanced accumulation in C. kikuchii. Targeted disruption of the genomic copy of one of these, now designated CFP (cercosporin facilitator protein), results in a drastic reduction in cercosporin production, greatly reduced virulence of the fungus to soybean, and increased sensitivity to exogenous cercosporin. Sequence analysis of CFP reveals an 1,821-bp open reading frame encoding a 65.4-kDa protein similar to several members of the major facilitator superfamily (MFS) of integral membrane transporter proteins known to confer resistance to various antibiotics and toxins in fungi and bacteria. We propose that CFP encodes a cercosporin transporter that contributes resistance to cercosporin by actively exporting cercosporin, thus maintaining low cellular concentrations of the toxin. PMID- 10517031 TI - Identification of a locus in arabidopsis controlling both the expression of rhizobacteria-mediated induced systemic resistance (ISR) and basal resistance against Pseudomonas syringae pv. tomato. AB - Selected nonpathogenic rhizobacteria with biological disease control activity are able to elicit an induced systemic resistance (ISR) response that is phenotypically similar to pathogen-induced systemic acquired resistance (SAR). Ten ecotypes of Arabidopsis thaliana were screened for their potential to express rhizobacteria-mediated ISR and pathogen-induced SAR against the leaf pathogen Pseudomonas syringae pv. tomato DC3000 (Pst). All ecotypes expressed SAR. However, of the 10 ecotypes tested, ecotypes RLD and Wassilewskija (Ws) did not develop ISR after treatment of the roots with nonpathogenic Pseudomonas fluorescens WCS417r bacteria. This nonresponsive phenotype was associated with relatively high susceptibility to Pst infection. The F1 progeny of crosses between the non-responsive ecotypes RLD and Ws on the one hand, and the responsive ecotypes Columbia (Col) and Landsberg erecta (Ler) on the other hand, were fully capable of expressing ISR and exhibited a relatively high level of basal resistance, similar to that of their WCS417r-responsive parent. This indicates that the potential to express ISR and the relatively high level of basal resistance against Pst are both inherited as dominant traits. Analysis of the F2 and F3 progeny of a Col x RLD cross revealed that inducibility of ISR and relatively high basal resistance against Pst cosegregate in a 3:1 fashion, suggesting that both resistance mechanisms are monogenically determined and genetically linked. Neither the responsiveness to WCS417r nor the relatively high level of basal resistance against Pst were complemented in the F1 progeny of crosses between RLD and Ws, indicating that RLD and Ws are both affected in the same locus, necessary for the expression of ISR and basal resistance against Pst. The corresponding locus, designated ISR1, was mapped between markers B4 and GL1 on chromosome 3. The observed association between ISR and basal resistance against Pst suggests that rhizobacteria-mediated ISR against Pst in Arabidopsis requires the presence of a single dominant gene that functions in the basal resistance response against Pst infection. PMID- 10517032 TI - The Rhizobium etli trpB gene is essential for an effective symbiotic interaction with Phaseolus vulgaris. AB - A mutant strain (CTNUX4) of Rhizobium etli carrying Tn5 unable to grow with ammonium as the sole nitrogen source was isolated and characterized. Sequence analysis showed that Tn5 is inserted into a trpB (tryptophan synthase)-homologous gene. When tested on the roots of Phaseolus vulgaris, strain CTNUX4 was able to induce only small, slightly pink, ineffective (Fix-) nodules. However, under free living conditions, strain CTNUX4 was unable to produce flavonoid-inducible lipo chitin oligosaccharides (Nod factors) unless tryptophan was added to the growth medium. These data and histological observations indicate that the lack of tryptophan biosynthesis affects the symbiotic behavior of R. etli. PMID- 10517033 TI - Molecular characterization of GmFOX2, an evolutionarily highly conserved gene from the mycorrhizal fungus Glomus mosseae, down-regulated during interaction with rhizobacteria. AB - Arbuscular mycorrhizal (AM) fungi form the most wide-spread symbiosis of the plant kingdom. More than 80% of vascular plants are susceptible to colonization by the zygomycetous fungi from the order Glomales, and profit significantly by the nutrient exchange between plant and fungus. However, knowledge of the biology of these fungi still remains elusive because of their obligate biotrophism and, up to now, unculturability. The molecular mechanisms underlying the pre-symbiotic stages and the cell-to-cell communication between AM fungi and other soil microorganisms are, particularly, unknown. Here, we study these aspects by means of a molecular approach to monitor changes in the gene expression of the fungus Glomus mosseae (BEG12) in response to the rhizobacterium Bacillus subtilis NR1. The bacterium was found to induce specific increases in mycelial growth as well as changes in expression of GmFOX2, a highly conserved gene encoding a multifunctional protein of the peroxisomal beta-oxidation. We determined the gene structure and studied its expression in response to rhizobacteria at two time points. The results show that the fungus is able to change its gene expression in response to stimuli other than the plant. PMID- 10517034 TI - Glycopyrrolate: a useful drug in the palliation of mechanical bowel obstruction. PMID- 10517035 TI - Efficacy of lamotrigine on sensory symptoms and pain in peripheral neuropathies. PMID- 10517036 TI - Massage therapy for patients undergoing autologous bone marrow transplantation. AB - The purpose of the current study was to examine the impact of massage therapy on psychological, physical, and psychophysiological measures in patients undergoing autologous bone marrow transplantation (BMT). Patients scheduled to undergo BMT were randomly assigned to receive either (a) massage therapy, consisting of 20 minute sessions of shoulder, neck, head, and facial massage, or (b) standard treatment. Overall effects of massage therapy on anxiety, depression, and mood were assessed pretreatment, midtreatment, and prior to discharge using the State Trait Anxiety Inventory, Beck Depression Inventory, and Brief Profile of Mood States, respectively. The immediate effects of massage were measured via the State Anxiety Inventory, Numerical Scales of Distress, Fatigue, Nausea, and Pain and indices of psychophysiological arousal (heart rate, blood pressure, and respiration rate), collected prior to and following patients' first, fifth, and final massage (on Days--7, midtreatment, and predischarge). Analysis of the data evaluating the immediate effects of massage showed that patients in the massage therapy group demonstrated significantly larger reductions in distress, fatigue, nausea, and State Anxiety than the standard treatment group at Day-7, in State Anxiety at midtreatment, and in fatigue at the predischarge assessment. The overall measures of psychological symptoms measured at pretreatment, midtreatment, and prior to discharge showed no overall group differences, although the massage group scored significantly lower on the State Anxiety Inventory than the standard care group at the midtreatment assessment. The two groups together showed significant declines through time on scores from the Profile of Mood States and State and Trait Anxiety Inventories. PMID- 10517037 TI - Steady-state kinetics and dynamics of morphine in cancer patients: is sedation related to the absorption rate of morphine? AB - Eighteen patients suffering from chronic pain due to cancer completed a balanced, double-blind, double-dummy, two period cross-over trial comparing the pharmacokinetics (PK) and pharmacodynamics (PD) of morphine and its metabolites, morphine-3-glucuronide and morphine-6-glucuronide, after administration of morphine given as controlled-release (CR) tablets (every 12 h) and immediate release (IR) tablets (every 6 h). The same total daily dose of morphine was given in both study periods. Patients received both test formulations for 4 days and on the final day of each period, peripheral venous blood samples for analysis of morphine, morphine-3-glucuronide, and morphine-6-glucuronide were obtained. Pain intensity, sedation, and continuous reaction time (CRT) were assessed. No significant differences could be demonstrated in AUC/dose, Cmin, Cmax or fluctuation index values between the two treatments (IR and CR tablets) for either morphine or its metabolites. Tmax for morphine and its metabolites occurred significantly later after administration of CR tablets than after administration of IR tablets. There were no significant differences between the IR and the CR formulation with respect to analgesia and side effects, and there was no difference in the patients' overall impression of the two treatments. More important, there was no difference between the Tmax and the time to peak sedation after administration of IR tablets (P = 0.63). However, due to the relatively small number of patients and the variability in the data, the statistical power of the test was only 0.074. The risk of a type II error is 0.926. These data demonstrate the PK and PD similarities and differences between CR and IR morphine. They suggest that there may be a relationship between Tmax (determined by absorption rate) and sedation, but further evaluation of this potential relationship is needed. PMID- 10517038 TI - High-dose tramadol in comparison to low-dose morphine for cancer pain relief. AB - Cancer pain treatment following the World Health Organization guidelines is effective and feasible. However, the evidence supporting the use of opioids for mild to moderate pain on the second step of the analgesic ladder is widely discussed. The present evaluation compares the efficacy and safety of high doses of oral tramadol (> or = 300 mg/d) with low doses of oral morphine (< or = 60 mg/d). Patients were included in this nonblinded and nonrandomized study if the combination of a nonopioid analgesic and up to 250 mg/d of oral tramadol was inadequate. 810 patients received oral tramadol for a total of 23,497 days, and 848 patients received oral morphine for a total of 24,695 days. The average dose of tramadol was 428 +/- 101 mg/d (range 300-600 mg/d); the average dose of morphine was 42 +/- 13 mg/d (range 10-60 mg/d). Additional nonopioid analgesics were given on more than 95% of days. Antiemetics, laxatives, neuroleptics, and steroids were prescribed significantly more frequently in the morphine group; the use of other adjuvants was similar in both groups. The mean pain intensity on a 0 100 numerical rating scale (NRS) was 27 +/- 21 (95% CI 26-29) in the tramadol and 26 +/- 20 (95% CI 24-27) in the morphine group (NS). The analgesic efficacy was good in 74% and 78%, satisfactory in 10% and 7%, and inadequate in 16% and 15% of patients receiving tramadol and morphine, respectively (NS). Constipation, neuropsychological symptoms, and pruritus were observed significantly more frequently with low-dose morphine; other symptoms had similar frequencies in both groups. These data suggest that tramadol can be used for the treatment of cancer pain, when nonopioids alone are not effective. High doses of tramadol are effective and safe. PMID- 10517039 TI - Validation of the German version of the Brief Pain Inventory. AB - The Brief Pain Inventory is a comprehensive instrument for pain assessment and has been validated in several languages. A validated German version was not available until now. From March to May 1995 all outpatients of the pain clinic of the Department of Anesthesiology completed a questionnaire with the German versions of the Brief Pain Inventory (BPI) and the SF-36 quality-of-life questionnaire. The BPI was repeated after the consultation. The physician assessed the performance status score of the Eastern Cooperative Oncology Group (ECOG). The questionnaire was completed by 151 patients. Forty-two patients were excluded from evaluation for methodological reasons, so 109 patients were evaluated. As in the original version of the BPI, factor analysis showed a common factor for pain intensity and a second factor for pain-related interference with function. The comparative fit index of 0.86 confirmed this model. Responses before and after consultation correlated closely for the sum scores of the pain intensity items (Perarson correlation r = 0.976) as well as for the interference with function items (r = 0.974). Pain intensity in the BPI correlated with bodily pain in the SF-36 (r = 0.585). Sum scores of the pain interference items were higher in patients with deteriorated ECOG performance status, whereas sum scores of the intensity items were not changed. Validity and reliability of the German BPI were comparable to the original version. The BPI may be advantageous for palliative care patients, as it places only a small burden on the patient and offers easy criteria for evaluation. However, further research is needed to differentiate the impact of pain-related and disease-related interference with function on the BPI, and to find an algorithm for the evaluation of the BPI when values are missing. PMID- 10517040 TI - Methadone response in advanced cancer patients with pain followed at home. AB - Concerns about the safety of therapy with methadone, which may arise because of its pharmacokinetic characteristics and inappropriate dosing, may deter clinicians from using this drug, especially in elderly patients. Experience is accumulating that the drug may be used safely and successfully if low doses are given initially and care is taken in the titration of the dose against the pain. A prospective study was carried out in a consecutive sample of 45 advanced cancer patients followed at home, who had never received other strong opioids for their pain. Patients were treated with an oral liquid preparation of methadone, which was administered 2-3 times daily, according to need. Doses were kept as low as possible and were titrated to achieve acceptable analgesia with minimal adverse effects. The methadone starting dose (MSD) at referral, the maximum dose of methadone (MMD), the days of methadone treatment, the use of other nonopioid analgesics, symptoms associated with methadone therapy, pain intensity, and pain mechanism were recorded. Methadone escalation index percentage (MEI%) and methadone escalation index in mg (MEI mg) were calculated from these parameters. No correlations between age and gender, and MSD, MMD, days on methadone, VAS and symptoms were found. No significant differences were found in pain mechanisms, age, and other parameters, including methadone-related symptoms. Treatment of pain with methadone provides important support to patients with cancer followed at home and the risks are low with individually titrated doses, even in older patients or in the presence of a neuropathic pain mechanism. PMID- 10517041 TI - Pain and loss of function in head and neck cancer survivors. AB - Head and neck cancers are relatively uncommon malignancies and the characteristics of pain and functional impairments in survivors are not well studied. To characterize the incidence, location, severity, types and causes of pain; associated functional impairments, and pain management methods, the medical charts of 40 consecutive outpatients with biopsy-proven head and neck cancers were reviewed. Pain was severe in 52% (N = 21), and was located near sites of tumor origin. Pain was caused by tumor recurrence in 35% (N = 14), treatment sequelae in 30% (N = 12), multiple etiologies in 25% (N = 10), and unrelated causes in 10% (N = 4). Pains were mixed nociceptive and neuropathic pain in 37.5% (N = 15), nociceptive pain in 32.5% (N = 13), myofascial in 13.0% (N = 6), neuropathic in 7.5% (N = 3); and other mixed types in 7.5% (N = 3). Despite the high prevalence of dysphagia (82%), 60% used orally administered opioid-nonopioid analgesics. Physical disfigurement (87.5%; N = 35), dysphagia (62.5%, N = 25), and jaw dysfunction (40.0%; N = 16) were the most frequent physical impairments. Multiple regression analysis showed that the presence of skull base or mandibular bone involvement had significant influence on the severity of pain (P = 0.03, adjusted R2 0.25) We conclude that pain in head and neck cancer can be chronic, severe, and persistent despite completion of oncologic treatment. PMID- 10517042 TI - Clinicians' perceptions of barriers to pain management in AIDS. AB - A growing body of literature has demonstrated the widespread undertreatment of pain in patients with AIDS. While clinician-related barriers to cancer pain management have been studied, to date there has been no systematic attempt to survey clinician-related barriers to the management of pain in patients with AIDS. We surveyed AIDS health care providers' attitudes towards pain management, as well as their perception of the barriers to adequate pain management in patients with HIV disease. Subjects were 492 AIDS care providers attending continuing education symposia on the clinical management of pain in patients with AIDS in 5 major U.S. cities (New York, Philadelphia, San Francisco, Los Angeles, and Miami). Results indicated that the most frequently endorsed barriers to pain management were those regarding lack of knowledge about pain management or access to pain management experts, and concerns regarding potential substance abuse or addiction. Experience in the management of pain in patients with AIDS was inversely correlated with endorsement of barriers related to pain management expertise and concern regarding potential substance abuse. More experienced clinicians were significantly less likely to cite these factors as barriers to pain management. More knowledgeable respondents were significantly more likely to identify barriers to pain management and individuals with more conservative attitudes towards pain management were significantly more likely to cite substance abuse issues or medical concerns as barriers to pain management. PMID- 10517043 TI - Neglect-like symptoms in complex regional pain syndrome: results of a self administered survey. AB - Reflex sympathetic dystrophy (RSD), recently reclassified as a complex regional pain syndrome, type I (CRPS-I), is best known for its disabling sensory symptoms, including pain, allodynia, and abnormal skin temperature. Yet, motor dysfunction is common in CRPS and can result in major disability. In addition to weakness of the involved limb, CRPS patients may develop symptoms akin to a neurological neglect-like syndrome, whereby the limb may feel foreign ("cognitive neglect") and directed mental and visual attention is needed to move the limb ("motor neglect"). Members of the patient support group, the Reflex Sympathetic Dystrophy Syndrome Association (RSDSA), were mailed a questionnaire inserted in their newsletter which inquired about the presence of these neglect-like symptoms; in addition, a separate medical history questionnaire was included to assess adequate documentation for the diagnosis of CRPS. A total of 242 patients returned the questionnaire but only 224 of the questionnaires were analyzed; 15 were excluded due to inadequate documentation of CRPS and 3 were excluded due to non-limb involvement. Eighty-four percent (84%) of these respondents endorsed the presence of at least one neglect symptom and 47% indicated they had both "cognitive" and "motor" neglect symptoms. Of interest, approximately 33% of respondents spontaneously wrote comments regarding the significant disability due to these neglect symptoms and the difficulty explaining these unusual symptoms to their health care providers and family. This patient survey confirms the presence of neglect-like symptoms in a subset of CRPS patients. Neglect-like symptoms need to be addressed and validated by health care providers. PMID- 10517044 TI - Antiemetic prophylaxis with ondansetron and methylprednisolone vs metoclopramide and methylprednisolone in mild and moderately emetogenic chemotherapy. AB - The purpose of the present study was to examine whether its is possible to successfully replace ondansetron (OND) with metoclopramide (MCP) in patients exposed to moderately emetogenic chemotherapy who did not experience severe nausea and vomiting while undergoing OND treatment during their first chemotherapy cycle. After switching to MCP, patients continued with this drug for three cycles, provided that they had adequate control of nausea and vomiting. Otherwise, they were switched back to OND. There were 76 patients, 60 women and 16 men, whose median age was 56 (mean 58) years. Karnofsky performance status score was 100 in 18 patients, 90 in 23, and 80 in 11 patients. No patient had previous chemotherapy. Thirty-four patients had breast cancer and received fluorouracil 500 mg/m2, epirubicin 100 500 mg/m2, and cyclophosphamide 500 mg/m2. Twelve patients had small cell lung cancer and received carboplatin 400 mg/m2 + etoposide 120 mg/m2 x 3 days. Twenty patients with ovarian cancer received carboplatin 350 mg/m2 and cyclophosphamide 500 mg/m2. Ten patients had cancer of unknown primary and received carboplatin 400 mg/m2, epirubicin 60 mg/m2, and etoposide 120 mg/m2 x 3 days. The OND schedule consisted of methylprednisolone 40 mg intravenous bolus followed by OND 8 mg in a 15-min infusion before chemotherapy, followed by OND 4 mg orally x 3 on the same and the next 2 days. Patients who did not experience nausea and vomiting with OND continued with an MCP schedule consisting of methylprednisolone 40 mg bolus followed by MCP 2 mg/kg in a 15-min infusion before chemotherapy, followed by MCP (20 mg x 4 on the day of therapy and the next 2 days after). Patients who failed with MCP or OND continued with OND. Considering our results as a whole, the intensity of nausea does not appear to influence the results of Gralla's scale. The results of Gralla's scale do not appear to be affected by the analysis of the antiemetic results and nausea on the next 2 days following chemotherapy administration. Overall, patients received 145 cycles with OND and 159 cycles with MCP. Of the 76 patients receiving OND-based antiemetic regimen during the first cycle, 13 (21%) experienced severe vomiting (Grade 2, 3) and the remaining 63 (79%) had mild or no vomiting (Grade 0, 1). Patients with Grade 0, 1 vomiting (63, 83%) continued with MCP in the second cycle. The final number of patients who failed on MCP, after 4 cycles of chemotherapy increased to 33 (43%); 43 (57%) were able to complete chemotherapy with MCP. Headache occurred in 15 (10%) cycles with OND and 8 (5%) with MCP. Flushing was noted in 12 (8%), and constipation occurred in 43 (30%) of OND cycles, and extrapyramidal manifestations occurred in 3 (5%) of patients receiving MCP. Diarrhea was noted in 3 (2%) of cycles with OND and in 28 (18%) with MCP. The cost ratio between MCP and OND was 1:14. If we administered OND only in patients who needed it, the overall cost decreased to 44%. Following the strategy applied in the present study, the cost decreased to 47%. PMID- 10517045 TI - Histological findings after long-term infusion of intrathecal ketamine for chronic pain: a case report. AB - Ketamine, a selective, noncompetitive N-methyl-D-aspartate (NMDA)-receptor antagonist, is able to alter pain perception at the spinal level. Little clinical data exist on the intrathecal and epidural use of ketamine in chronic pain. Histopathologic findings after intrathecal injection of ketamine with and without preservatives are rarely reported. This outcome was evaluated in a 72-year-old woman with abdominal pain due to cancer who was treated with the intrathecal application of bupivacaine, clonidine, and morphine. We reached satisfactory pain relief with the addition of ketamine to the mixture for 7 days. On postmortem, focal lymphocytic vasculitis close to the catheter injection site was found. This finding has not been described previously after long-term application of ketamine intrathecally. The intrathecal infusion of ketamine with preservative, or the mixture of ketamine, clonidine, morphine, and bupivacaine resulted in isolated lymphocytic vasculitis of the spinal cord and leptomeninges without any clinical signs of neurological deficit. PMID- 10517046 TI - Epi-arachnoidal drug deposit: a rare complication of intrathecal drug therapy. AB - Intrathecal (i.t.) drug application is accepted as a highly effective treatment option for various neurological conditions. Technical risks and potentially dangerous complications require appreciation. We present the case of a patient treated with i.t. recombinant, human brain-derived neurotrophic factor (rhBDNF) as an experimental therapy for amyotrophic lateral sclerosis (ALS). Five days after starting the i.t. drug infusion, she complained of severe headache and nausea. Radiological studies suggested the catheter was located within the epi arachnoidal space. A deposit of more than 10 ml secluded from the subarachnoidal space was found within this space. I.t. contained a high concentration of the applied drug. Revision of the catheter resulted in complete recovery from symptoms and i.t. infusion could be continued. The epi-arachnoidal positioning of a spinal catheter is a potential cause for treatment failure. If the membrane around the fluid deposit ruptures, the drug could be released into the subarachnoidal space, with the consequence of a potentially life-threatening complication. PMID- 10517047 TI - WISC-III third factor. PMID- 10517048 TI - Violence in the media. PMID- 10517049 TI - Bipolar disorder and ADHD. PMID- 10517050 TI - Nutritional supplements in ADHD. PMID- 10517051 TI - Changes in the practice of child and adolescent psychiatry. PMID- 10517052 TI - Alexithymia in an adolescent with agenesis of the corpus callosum and chronic pain. PMID- 10517053 TI - Child and adolescent abuse and neglect research: a review of the past 10 years. Part I: Physical and emotional abuse and neglect. AB - OBJECTIVE: To review the clinically relevant literature on the physical and emotional abuse and neglect of children and adolescents published during the past 10 years. METHOD: Literature published between 1988 and 1998 was reviewed following a systematic search of Medline, Psychinfo, and the National Clearinghouse on Child Abuse and Neglect. RESULTS: During the last decade there has been substantial progress in understanding the symptomatology associated with maltreatment. However, prevention and intervention research studies are relatively rare and frequently have important methodological limitations. CONCLUSIONS: Child maltreatment research in the next decade needs to focus on understanding factors leading to resilient outcomes and on assessing the effectiveness of psychotherapeutic and psychopharmacological treatment strategies. Increased resources are needed to support child maltreatment research studies and investigators. PMID- 10517054 TI - Cognitive-behavioral group treatments in childhood anxiety disorders: the role of parental involvement. AB - OBJECTIVES: This study examined (1) the effect of a cognitive-behavioral group intervention on anxiety, depression, and coping strategies in school-age children (aged 7-12 years) with Axis I anxiety disorders; and (2) the effect of parental involvement on treatment outcomes. METHOD: Parents and children (N = 62) were randomly assigned to one of three 12-week treatment conditions: parent and child intervention, child-only intervention, and parent-only intervention. Child anxiety, depression, and coping strategies were assessed before and after treatment. RESULTS: All treatment groups reported fewer symptoms of anxiety and depression posttreatment and changes in their use of coping strategies. Children in the parent and child intervention used more active coping strategies posttreatment compared with children in the other 2 treatment conditions. Parents in this treatment condition reported a significantly greater improvement in their children's emotional well-being than parents in the other treatment conditions. CONCLUSIONS: Cognitive-behavioral group interventions reduced symptoms of anxiety and depression in school-age children with anxiety disorders. Concurrent parental involvement enhanced the effect on coping strategies. Further investigation is needed to corroborate the effectiveness of such short-term interventions and the maintenance of treatment effects. PMID- 10517055 TI - Psychometric properties of the Screen for Child Anxiety Related Emotional Disorders (SCARED): a replication study. AB - OBJECTIVE: To replicate and extend work on the psychometric properties of the Screen for Child Anxiety Related Emotional Disorders (SCARED), a child and parent self-report instrument used to screen for children with anxiety disorders. METHOD: The 41-item version of the SCARED was administered to a new sample of 190 outpatient children and adolescents and 166 parents. The internal consistency, discriminant, and convergent validity were assessed. In addition, using discriminant function analysis, a briefer version of the SCARED was developed. RESULTS: Using item analyses and factor analyses on the 41-item version, 5 factors were obtained: panic/somatic, generalized anxiety, separation anxiety, social phobia, and school phobia. In general, the total score and each of the 5 factors for both the child and parent SCARED demonstrated good internal consistency and discriminant validity (both between anxiety and depressive and disruptive disorders and within anxiety disorders). A reduced version of the SCARED yielded 5 items and showed similar psychometrics to the full SCARED. CONCLUSIONS: In a new sample, the authors replicated their initial psychometric findings that the SCARED is a reliable and valid instrument to screen for childhood anxiety disorders in clinical settings. Furthermore, pending future research, the 5-item SCARED appears to be a promising brief screening inventory for anxiety disorders in epidemiological studies. PMID- 10517056 TI - Predicting DSM-III-R disorders from the Youth Self-Report: analysis of data from a field study. AB - OBJECTIVE: To predict DSM-III-R diagnoses from Youth Self-Report (YSR) scores. METHOD: The Diagnostic Interview Schedule for Children Version 2.1c (DISC-2.1c) and YSR were administered to 289 homeless adolescents. Stepwise discriminant analysis identified YSR scales contributing to predictions of DSM-III-R disorders. Paper-and-pencil prediction rules based on YSR "borderline" or "clinical" scores were evaluated. RESULTS: Statistically significant discriminant functions for disruptive disorders, depressive disorders, manic disorders, attention-deficit hyperactivity disorder, schizophrenia, and posttraumatic stress disorder, each based on a unique pair of YSR scales, produced overall hit rates of 0.66 to 0.90. Paper-and-pencil predictions produced comparable results. The weakest overall predictions were for the disruptive behaviors; the best rule ("IF Aggressive OR Delinquent is at least borderline THEN predict oppositional defiant disorder or conduct disorder") produced a 0.72 hit rate. The strongest overall predictions were for schizophrenia; the best prediction rule ("IF [Thought Problems AND Delinquent are at least borderline] AND [at least one is clinical] THEN predict schizophrenia") produced a 0.87 hlt rate. CONCLUSIONS: While the success rates reported here are specific to this sample, it appears that the YSR has good ability to predict DSM-III-R diagnoses as determined by the DISC. Furthermore, it was demonstrated that categorical diagnoses can be treated as locations or cluster sectors in a multidimensional space. PMID- 10517057 TI - Identification of elementary school children at risk for disruptive behavioral disturbance: validation of a combined screening method. AB - OBJECTIVES: To examine the validity of the teacher-rated Inattention/Overactivity With Aggression (IOWA Conners) questionnaire followed by the Conners Abbreviated Symptom Questionnaire (CASQ) as a feasible, combined screening method to identify children at high risk for externalizing behavioral disturbance in a school setting. METHOD: The IOWA Conners and CASQ were administered to the entire population of children, grades K through 6, in a single elementary school. Using a whole-number IOWA Conners threshold score of 18 (1.5 SD), coupled with a CASQ score of 18 (1.0 SD), a high-risk group and an age-, sex-, and classroom-matched low-risk control group were selected for comparison on the Child Behavior Checklist (CBCL), Teacher's Report Form (TRF), Child and Adolescent Functional Assessment Scale (CAFAS), and Diagnostic Interview for Children and Adolescents Revised diagnoses. RESULTS: The data showed significant intergroup differences in CBCL (p < .0002), TRF (p < .0006), and CAFAS (p < .0002) scores and higher numbers of DSM diagnoses (1.7 +/- 0.3 versus 0.1 +/- 0.3, p < .0007) between the high- and low-risk group. The positive predictive value for externalizing diagnoses for the IOWA Conners plus CASQ was 100% and negative predictive value was 70%. CONCLUSIONS: The IOWA Conners combined with the CASQ is a useful initial screening strategy in the school setting for identification of children with disruptive behavioral difficulties. PMID- 10517058 TI - Attention problems versus conduct problems as 6-year predictors of signs of disturbance in a national sample. AB - OBJECTIVE: To test whether attention problems predicted different signs of disturbance than conduct problems over 3 and 6 years. METHOD: Gender-specific criteria for deviance on parents' ratings of attention versus conduct problems were tested as predictors of interview-reported signs of disturbance in a national sample first assessed at ages 4 to 16 years. RESULTS: Males and females deviant on both attention and conduct problems showed higher rates of several signs of disturbance than did those deviant on only one type of problem. Subjects deviant only on conduct problems showed higher rates of several signs than did controls, whereas those deviant only on attention problems exceeded controls mainly on special education services. Unaggressive "delinquent" conduct problems predicted dropping out of school, unwed pregnancy, and total signs for both genders during transitions to adulthood. CONCLUSIONS: Attention problems predict receipt of special education but contribute much less than conduct problems to predicting other signs of disturbance. Differential assessment of aggressive versus unaggressive conduct problems can improve prediction, as can gender specificity in setting criteria for deviance and in testing outcomes. PMID- 10517059 TI - Persistence and desistance of oppositional defiant disorder in a community sample of children with ADHD. AB - OBJECTIVE: To examine the developmental progression of comorbid oppositional defiant disorder (ODD) in a community sample of children with attention-deficit hyperactivity disorder (ADHD) with particular emphasis on persistence and desistance of ODD and the emergence of new cases of conduct disorder (CD). METHOD: A sample of disruptive children was identified from a multiple-gate epidemiological screen and stratified into diagnostic subgroups on the basis of a structured interview. A comparison sample of nondisruptive children was also identified. Group comparisons were performed on demographic, descriptive, family history, and clinical characteristics. Changes in rates of ODD symptoms and diagnostic subgroup membership were assessed after a 4-year longitudinal interval. Predictors of diagnostic group persistence were tested. RESULTS: Few differences distinguished diagnostic subgroups at baseline. Of the 43 children with baseline diagnoses of ADHD + ODD, only 1 (2.3%) was found to have developed CD at follow-up. Over time there was a 57% rate of ODD persistence and a 43% rate of ODD desistance. Negative parenting practices and mothers' psychiatric disorders predicted persistence of ODD. CONCLUSIONS: There was little evidence to show that ODD acted as a precursor to CD. However, when CD was diagnosed at baseline it was always associated with or preceded by ODD (i.e., prodrome). For a subgroup of children with ADHD, comorbid ODD symptoms are relatively unstable and may represent transient developmental perturbations that have little prognostic significance. For a larger subgroup of children with ADHD, ODD symptoms persist into the adolescent years and are associated with adverse parenting practices. PMID- 10517060 TI - Mood, latitude, and seasonality among adolescents. AB - OBJECTIVE: To examine the effect of sex, latitude, and behavior problems on symptoms related to pediatric seasonal affective disorder among seventh and ninth graders. METHOD: A school survey including a modified version of the Seasonal Pattern Assessment Questionnaire was carried out in 2 Finnish cities located in the 60th and 67th northern latitudes. Altogether 1,458 questionnaires were analyzed, representing 89% of the target population. RESULTS: Seasonal changes in mood and behavior were commonly reported among seventh and ninth graders. A high Global Seasonality score (> 95th percentile) was associated with female gender and emotional and hyperactivity symptoms. During February and March, girls living in the 67th latitude reported more seasonal distress than girls living at the 60th latitude. CONCLUSIONS: It is important to recognize pediatric seasonal affective disorder and related problems among adolescents. Seasonal alterations in child and adolescent behavior are not well understood and need to be investigated more thoroughly. PMID- 10517061 TI - The neglected link between eating disturbances and aggressive behavior in girls. AB - OBJECTIVES: Research has linked eating disturbances with behavioral impulsivity. Little is known, however, about whether eating disturbances and aggressive behavior have a tendency to co-occur in the same girls. This article assesses the eating disturbance-aggressive behavior association and then examines the extent to which these factors confer a risk on drug use and attempted suicide. METHOD: Survey data were gathered from 3,630 girls in grades 6 through 12 in the upper Midwest. Girls responded anonymously to questions regarding binge eating and purging, dietary restriction, aggressive behavior, drug use, and attempted suicide. Logistic regression analysis was used to assess the unique contribution of demographic variables, eating disturbances, and aggression on drug use and attempted suicide. RESULTS: Eating disturbances were significantly associated with aggressive behavior. Girls who endorsed binge eating and purging or dietary restriction had odds of aggressive behavior 2 to 4 times higher than girls who did not endorse these items. Logistic regression revealed that eating disturbances and aggressive behavior were significantly associated with both drug use and attempted suicide. CONCLUSIONS: Eating disturbances are significantly associated with aggressive conduct in adolescent girls. The constellation of eating disturbances and aggressive behavior is associated with a greater risk of drug use and attempted suicide. PMID- 10517062 TI - Sleep problems in children with attention-deficit/hyperactivity disorder: impact of subtype, comorbidity, and stimulant medication. AB - OBJECTIVE: To determine the relationship of sleep problems to attention deficit/hyperactivity disorder (ADHD), diagnostic subtype, comorbid disorders, and the effects of stimulant treatment. METHOD: On the basis of clinical diagnostic interviews, children aged 6 to 12 years were assigned to 4 groups: unmedicated ADHD (n = 79), medicated ADHD (n = 22), clinical comparison (n = 35), and healthy nonclinical comparison (n = 36). These groups were compared on 2 sleep questionnaires completed by the parents that assessed current sleep problems and factors associated with sleep difficulties (i.e., sleep routines, sleep practices, child and family sleep history). RESULTS: Factor analysis revealed 3 sleep problem categories: dyssomnias, parasomnias, and sleep-related involuntary movements. Linear regression analyses showed that (1) dyssomnias were related to confounding factors (i.e., comorbid oppositional defiant disorder and stimulant medication) rather than ADHD; (2) parasomnias were similar in clinical and nonclinical children; and (3) the DSM-IV combined subtype of ADHD was associated with sleep-related involuntary movements. However, sleep-related involuntary movements were more highly associated with separation anxiety. CONCLUSIONS: The results suggest that the relationship between sleep problems and ADHD is complex and depends on the type of sleep problem assessed as well as confounding factors such as comorbid clinical disorders and treatment with stimulant medication. PMID- 10517064 TI - Vietnamese parental perceptions of child and adolescent mental illness. AB - OBJECTIVES: To determine the symptoms and behaviors in children which are considered psychopathological by Vietnamese parents, to identify professionals and agencies in the community whom Vietnamese parents would consult if their child had a mental illness, and to determine Vietnamese parents' awareness of existing community mental health services. METHOD: Structured interviews were conducted with a randomized community sample of 283 Vietnamese parents in Perth, Australia. Parents were asked to identify the symptoms and behaviors they considered psychopathological in children, where they would turn for help with a mentally ill child, their knowledge of community mental health services for children, and their understanding of the causes of child psychiatric disorders. RESULTS: Vietnamese parents identified psychotic symptoms, disorientation, and suicidal thoughts and behavior as psychopathological. They preferentially endorsed Western-style treatment approaches but had little awareness of existing community mental health services for children. A biological/chemical imbalance, traumatic experiences, and a metaphysical/spiritual imbalance were identified as the most likely causes of child mental illness. CONCLUSIONS: Despite a different cultural tradition, Vietnamese parents appear open to services provided by Western-trained mental health professionals. Their very limited awareness of child and adolescent mental health services in the community, however, may severely limit their utilization of such services. PMID- 10517063 TI - Premutation female carriers of fragile X syndrome: a pilot study on brain anatomy and metabolism. AB - OBJECTIVE: It was thought that premutation carriers of fragile X syndrome (FraX) have no neurobiological abnormalities, but there have been no quantitative studies of brain morphometry and metabolism. Thus the authors investigated brain structure and metabolism in premutation carriers of FraX. METHOD: Eight normal IQ, healthy female permutation FraX carriers aged 39 +/- 9 years (mean +/- SD) and 32 age-sex-handedness-matched controls (39 +/- 10 years) were studied; in vivo brain morphometry was measured using volumetric magnetic resonances imaging, and regional cerebral metabolic rates for glucose were measured using positron emission tomography and (18F)-2-fluoro-2-deoxy-D-glucose. RESULTS: Compared with controls, FraX premutation carriers had a significant (1) decrease in volume of whole brain, and caudate and thalamic nuclei bilaterally; (2) increase in volume of hippocampus and peripheral CSF bilaterally, and third ventricle; (3) relative hypometabolism of right parietal, temporal, and occipital association areas; (4) bilateral relative hypermetabolism of hippocampus; (5) relative hypermetabolism of left cerebellum; and (6) difference in right-left asymmetry of the Wernicke and Broca language areas. CONCLUSIONS: Premutation carriers of FraX, as defined by analysis of peripheral lymphocytes, have abnormalities in brain anatomy and metabolism. The biological basis for this is unknown, but most likely it includes tissue heterogeneity for mutation status. The findings may be of relevance to people counseling families with FraX and to understanding other neuropsychiatric disorders which are associated with expansion of triplet repeats and genetic anticipation. PMID- 10517065 TI - Psychiatric consultation to the neonatal intensive care unit: liaison matters. PMID- 10517066 TI - Measurement and descriptive statistics. PMID- 10517067 TI - Genetics of childhood disorders: VII. Cloning genes of interest. PMID- 10517068 TI - Childhood-onset systemic lupus erythematosus: a comparative study of African Americans and Latin Americans. AB - This study compared the clinical and serologic features in two different ethnic groups of patients with childhood-onset systemic lupus erythematosus (SLE). One hundred seventy-one SLE patients comprised the study population; 61 (55 girls and 6 boys) were African American with age at onset of 13 +/- 2.9 years, and 110 (97 girls and 13 boys) were Latin American (Colombian) with age at onset of 13 +/- 3.2 years. Clinical, demographic, and laboratory data were obtained by chart review using a standard data collection form. African-American patients more commonly manifested discoid skin lesions, malar rash, pulmonary fibrosis, and pleuritis, and less commonly manifested photosensitivity, livedo reticularis, and vascular thrombosis than did Latin Americans. In addition, there was a higher frequency of anti-dsDNA, anti-Sm, anti-RNP, and anti-Ro positivity among African Americans compared with Latin-American patients. These results suggest the presence of ethnic differences in the clinical expression of SLE. PMID- 10517069 TI - Frequency and factors associated with cardiomyopathy in patients with human immunodeficiency virus infection in an inner-city hospital. AB - The frequency and factors associated with cardiomyopathy were studied among inner city hospital patients with human immunodeficiency virus (HIV) infection. Of 84 patients with HIV infection, 20 (24%) had cardiomyopathy. Fourteen (70%) of the patients with cardiomyopathy did not have clinical evidence of congestive heart failure. There was no significant association between cardiomyopathy and common opportunistic infections or zidovudine treatment. These results indicate that cardiomyopathy is common in patients with HIV infection and often is clinically unsuspected. PMID- 10517071 TI - A survey of the ethnic and racial distribution in orthopedic residency programs in the United States. AB - This study examined the racial and ethnic composition of orthopedic training programs in the United States. A questionnaire was mailed in January 1995 to chairpersons at 159 orthopedic programs in the United States. Eighty-nine (56%) responses were received. The distribution of orthopedic residents and fellows was as follows: white non-Hispanic, 84.2%; Asian, 6.6%; African American, 3.6%; Native American, 2.2%; Puerto Rican, 1.2%; Mexican American, 0.8%; and other Hispanic, 1%. African Americans and Hispanics were under-represented in orthopedic training programs compared with their numbers in the general population. The percentage of residents in these two minority groups also were below goals established by the Council on Graduate Medical Education and the US Government's Healthy People 2000 report. In contrast, Native Americans and Asians were overrepresented. If racial balance is to be achieved in orthopedics, new incentives must be created to encourage more African Americans and Hispanics to enter orthopedic residency training programs. PMID- 10517070 TI - Colorectal cancer surveillance in African-American and white patients at an urban university medical center. AB - Colorectal cancer causes significant morbidity and mortality in the United States. Recommendations for colorectal cancer screening have been developed. This study evaluated the colorectal cancer screening practices of African-American and white patients by internal medicine resident physicians. A retrospective chart review was conducted during 1989-1994. The performance of rectal examination, fecal occult blood testing, and flexible sigmoidoscopy among patients > 50 years was evaluated. The medical records of 200 patients (90 men and 110 women) were reviewed. Ninety-one rectal examinations, 26 fecal occult blood testing, and 30 flexible sigmoidoscopies were performed. There were 129 African-American (54 men and 75 women) and 52 white (26 men and 27 women) patients. Of the African American patients, 57 underwent a rectal examination, 17 had fecal occult blood testing, and 26 underwent flexible sigmoidoscopy. Of the white patients, 24 had a rectal examination, 8 had fecal occult blood testing, and 12 underwent flexible sigmoidoscopy. These results demonstrate that resident physicians adhered poorly to colorectal cancer screening recommendations. There was no statistically significant difference in the screening of African-American and white patients. Increased efforts should be made to improve colorectal cancer screening practices by resident physicians. PMID- 10517072 TI - Effect of prenatal care on infant mortality rates according to birth-death certificate files. AB - Infant mortality has decreased nationwide; however, our national rates still log behind those of other industrialized countries, especially the rates for minority groups. This study evaluates the effect of prenatal care and risk factors on infant mortality rates in Chicago. Using linked infant birth and death certificates of Chicago residents for 1989-1995, a total of 5838 deaths occurring during the first year of life were identified. Birth certificate variables, especially prenatal care, were reviewed. Variables were compared by stratified analysis. Pearson chi 2 analysis and odd ratios (ORs) were computed. Infant mortality rate (IMR) in Chicago decreased from 17 in 1989 to 12.6 in 1995 (P < .0001). Some factors increased IMR several fold: prematurity (OR 17.43), no prenatal care (OR 4.07), inadequate weight gain (OR 2.95), African-American ethnicity (OR 2.55), and inadequate prenatal care (OR 2.03). Compared with no care, prenatal care was associated with lower IMR; however, early care was associated with higher IMR and ORs than later care. These results demonstrate prenatal care is associated with lower IMR; however, compared with late prenatal care, early care does not improve IMR. Further studies should evaluate whether improving the quality of care improves IMRs. PMID- 10517073 TI - Length of pregnancy in African Americans: validation of a new predictive rule. AB - This study evaluated whether a new predictive rule is more accurate for estimating the length of pregnancy in African Americans than Nagele's rule, the accepted standard. After identifying women in early pregnancy, telephone interviews were conducted to obtain information about 16 previously established determinants of gestational length. Based on these data, a linear multivariate regression model was used to predict an estimated delivery date (EDD) for each mother. In addition, the EDD was determined using Nagele's rule. Later, the actual delivery date was compared with the EDD predicted by the new rule and with the EDD predicted by Nagele's rule. Each pregnancy was assigned to its better prediction group, either the new rule's group or the Nagele's rule group. Fifty seven pregnancies were identified prospectively and monitored. The new rule predicted the actual delivery date more accurately in 66% (37/56) of pregnancies, Nagele's rule was a better predictor in 34% (19/56) of pregnancies, and both rules were equally accurate in predicting the delivery date for one pregnancy. The new rule was more precise than Nagele's rule (P = .022) when the binomial distribution was used. When using the linear regression model rule, a more accurate EDD can be determined for African-American women. Moreover, it is possible to predict the risk of preterm delivery (those occurring > 3 weeks earlier than the EDD). PMID- 10517074 TI - A frenzied passion in the head. PMID- 10517075 TI - Contemporary management of rheumatoid arthritis. PMID- 10517076 TI - Update on osteoarthritis. PMID- 10517077 TI - The clinical management of fibromyalgia. AB - Fibromyalgia is characterized by chronic diffuse muscular aches, fatigue and poor sleep; it affects nearly three million individuals in the United States alone, predominantly younger women. The diagnosis of fibromyalgia requires adherence to the American College of Rheumatology criteria and the exclusion of secondary causes and systemic diseases. Treatment with sleep cycle regulators, NSAIDs, and light aerobic exercise is usually helpful. Patients must be reminded that fibromyalgia is often a chronic condition, but can be successfully treated. PMID- 10517078 TI - Treatment of juvenile rheumatoid arthritis. AB - Advances in pediatric rheumatology have closely paralleled those in adult rheumatology, but several unique features of JRA have led to some prominent differences. Methotrexate has clearly become the gold standard for use in moderate to severe disease. A variety of the second-line agents and newer methods of corticosteroid dosing can be used in milder disease or in patients who fail or refuse methotrexate. Biologic agents and newer immunosuppressive agents hold out significant promise for those with the most severe disease. As for adult disease, a cure for arthritis is far away, but the lives of these patients have truly been revolutionized by the wide array of newer therapies. PMID- 10517080 TI - Outpatient disease management: RIQP wants you! PMID- 10517079 TI - Cox-2 inhibitors: a new class of medication? PMID- 10517081 TI - Arthritis and disability in Rhode Island. PMID- 10517082 TI - Reducing the burden of arthritis. PMID- 10517083 TI - [Treatment and prognosis of T 4 renal cell carcinoma]. AB - BACKGROUND: We studied the cases with T 4 renal cell carcinoma (RCC) to characterize the factors associated with prolonged survival and to clarify the indication of extended nephrectomy. MATERIALS AND METHODS: The study population consisted of 53 patients (44 male and 9 female) with pT 4 RCC treated at the Yokohama City University Hospital and its affiliated hospitals from 1965 to 1994. Survival rates were analyzed with respect to clinicopathological factors (patient age, sex, symptom, tumor growing type, tumor size, histological grade, cell type, structural type, lymph node metastasis, vein invasion, distant metastasis and extended nephrectomy). RESULTS: One-year, 2-years, and 3-years survival rates of the cases with T 4 RCC were 30.4%, 16.4%, and 9.4% respectively. In univariate analysis, improved survival were correlated with no extra-urinary symptom (Logrank: p = 0.0048, Wilcoxon: p = 0.0423), no lymphnode metastasis (Logrank: p = 0.1045, Wilcoxon: p = 0.0199), no distant metastases (Logrank: p = 0.0007, Wilcoxon: p = 0.0006), and enforcement of extended nephrectomy (Logrank: p = 0.0018, Wilcoxon: p = 0.0008). In 28 cases with extended nephrectomy, improved survival was correlated with no extra-urinary symptom, no abdominal wall invasion and no distant metastases. In 5 cases with more than 3 year survival after extended nephrectomy, 4 cases were found to have no distant metastases at the time of operation. Non-operative therapy including interferon for 20 cases without extended nephrectomy were almost ineffective. CONCLUSIONS: These results indicate that if curative excision for T 4 RCC cases without distant metastases could be done, some patients might be appropriate candidates for extended nephrectomy. PMID- 10517084 TI - [Establishment of murine model of autoimmune male infertility]. AB - BACKGROUND: Experimental autoimmune orchitis (EAO) has been studied as an animal model for human immunological male infertility. We have already reported the induction of contralateral EAO ("sympathetic orchitis") by unilateral testicular injury in mice. In this paper, we report the induction of autoimmune infertility in such mice as well as EAO. METHODS: Ten to 20 needle punctures were made to the unilateral testis of mice and it was crushed by a needle-holder. Such mice were mated in vivio with female mice. RESULTS: Fifty to 80% mice whose testis was injured unilaterally became infertile in 3 months. Delayed type hypersensitivity (DTH), one of the cell-mediated immunities, to autologous testicular cells (TC) as well as anti-TC antibodies, humoral immunity, were both detected in those mice. CONCLUSION: We have clearly shown that unilateral testicular injury could induce not only the contralateral EAO but also autoimmune infertility in mice. Our present injury model mimics clinical testicular trauma; therefore, this testicular injury model can be very useful in studying the immunological mechanism of EAO and of human immunological male infertility. PMID- 10517085 TI - [Psychogenic lower urinary tract dysfunction in women: patho-physiological investigation for psychogenic frequency-urgency syndrome and psychogenic urinary retention]. AB - PURPOSE: Psychogenic lower urinary tract dysfunction (PLUTD) is composed of two syndromes; psychogenic frequency-urgency syndrome (PFUS) and psychogenic urinary retention (PUR). We evaluated the patho-physiology of PFUS and PUR, and explored the different pathogenesis in these syndromes. MATERIAL AND METHODS: Forty five patients with PLUTD, consisting of 23 patients with PFUS and 22 patients with PUR were investigated by using the psychological tests: CMI (Cornell Medical Index) and TEG (Todai's Egogram), a quantitative perspiration test in 45 females (23 patients with PFUS and 22 patients with PUR), and simultaneous measurements of voiding cysto-urethrography and urodynamic studies using the Life-Tech 6 channel polygraph in 35 patients (17 patients with PFUS and 18 patients with PUR). RESULTS: The prevalence in ages revealed two peaks, 20 years and 50 to 60 years. Over 25% of them had pyuria more than 10/hpf of WBC. Peak flow rate measured by uroflowmetry showed normal range in PFUS group and decreased in PUR group. The functional vesical volume was less than 100 ml in most patients with PFUS. Residual urine in PUR group was significantly greater. Capacity of the PFUS group were able to hold over 400 ml of contrast instilled through the urethral catheter, despite increased desire to void. Over 15% of the study group with PFUS showed uninhibited systolic contraction of detrusor (> 15 cm H2O) during filling phase. The measurement value of urodynamic parameters demonstrated that a periodic follow-up survey of the upper urinary tract should be performed because of the low compliance bladder in the patients with PLUTD. During voiding phase, the women with PFUS had a tendency to be divided into two groups, hypercontractile or acontractile detrusor. The voiding cysto-urethrography (VCUG) showed a tendency of bladder neck opening on patients with PFUS during filling phase. Most of PLUTD cases demonstrated a round to triangle shape on vesical configuration, which led to a spastic condition of detrusor muscle. We attempted to measure the quantitative perspiration using 3 kinds of loading tests; respiratory, arithmetic and psychological load. In the psychological loading test, we asked 98 questions about their daily lives including occupation, living condition, family relationship and sexual activities. Arithmetic loading test consisted of counting in reverse, subtraction and multiplication. The quantitative perspiration rate resulted in a "positive" in many patients with PFUS. Respiration loading test was performed to measure the respiration volume during 3 large inhales. Most patients with PUR tested within the normal range for respiration except for those patients with decreased or no perspiration during the psychiatric loading test. These results may reflect the psychological elements including suppression and subconscious defense mechanism. Neurosis which was diagnosed as having type III to type IV of the Cornell Medical Index was demonstrated in less than under 40% of patients with PFUS and more than 55% patients with PUR. There was no significant trend or difference between PFUS and PUR detected from Todai's Egogram. CONCLUSIONS: Due to the reflection of many psychological responses, it is necessary to investigate from various examinations including psychological, autonomical and classical urological studies for accurate diagnosis of PLUTD. PMID- 10517086 TI - [Histopathological effect and its predictors of neoadjuvant hormonal therapy by combined androgen blockade]. AB - OBJECTIVES: Combined androgen blockade (CAB) uning LH-RH agonist and flutamide has been performed as neoadjuvant therapy for T 2, 3 prostate cancers (CaP). The histological effects of neoadjuvant CAB therapy and influential factors were investigated. METHODS: Materials were 20 CaP cases which were underwent radical prostatectomy (RP) after neoadjuvant CAB therapy. All cases were diagnosed by echo-guided sextant needle biopsies. RP was performed after serum PSA was decreased to undetectable level. Histological effect was evaluated by general rule for clinical and pathological studies on prostate cancer (Japanese Urological Association). All cases were divided 2 groups by histological effects as follows: Group A (poor effect group): G 0 and G 1, Group B (good effect group): G 2 and G 3. Immunostaining of p 53 (mutant type), bcl-2 and Chromogranin A (ChA) were performed for both pretreatment needle biopsy and RP specimen. In addition, pretreatment serum PSA and Gleason grade were also investigated. RESULTS: Down grading were found in 30%. Down staging were found in 35% (7 cases). All 7 cases were negative surgical margins and 5 of 7 were clinical T 3. Negative bcl-2 of biopsy specimen was correlation with down grading (p = 0.008). In the histopathological evaluation, G 0 was 1, G 1 were 10, G 2 were 6 and G 3 were 3 cases. Gleason 4 or 5 elements of biopsy were found in 9/11 cases in Group A but only 3/9 cases in Group B (p = 0.027). The bcl-2 positive cells of biopsy were found in 8/11 cases in Group A but only 1/9 cases in Group B (p = 0.006). The p 53 and/or bcl-2 positive cells of biopsy were found in 10/11 cases in Group A but only 3/9 cases in Group B (p = 0.007). Serum PSA and ChA were not correlation with histological effect of neoadjuvant CAB therapy. But, in 3 cases, ChA positive cell appeared after neoadjuvant therapy. CONCLUSIONS: We could not expect more than 50% cases to show the down grading and down staging. But, in T 3 case, surgical failure could be decrease. We could expect prostate cancer cases without positive bcl-2 cells, p 53 over expression and Gleason 4 x 5 to reveal the good histological effects of neoadjuvant CAB therapy. PMID- 10517087 TI - [Effect of transurethral collagen injection for vesicoureteral reflux in patients with spina bifida]. AB - BACKGROUND: The effect of endoscopic injection of collagen was assessed in spina bifida patients with vesicoureteral reflux (VUR). METHODS: Endoscopic collagen injection was carried out for grade II or worse VUR according to the international classification. Twenty-two ureters were studied in 6 boys and 8 girls (mean: 14.4 years) who were followed up over a period of at least 3 months (mean: 5 months) after surgery they all had a negative preoperative skin test for collagen and were investigated radiologically and urodynamically. Cystograpy was performed 1, 3 and 12 months after surgery and thereafter once a year to detect recurrence of VUR. RESULTS: Anesthesia was not necessary in 4 patients. No adverse reactions occurred to the injection of collagen. VUR disappeared after 1 and 2 collagen injections in 17 (77%) and 2 (9%) ureters, respectively. The therapeutic effect of the single collagen injection showed no relationship to shape of the ureteral orifice, grade of VUR, compliance of the bladder, and presence of detrusor hyperreflexia. CONCLUSIONS: Endoscopic treatment of VUR with collagen injection in spina bifida patients is a simple and less invasive method. We obtained satisfactory short-term results by this method. However, since the risk factor of recurrence remains unclear, sufficient investigation of long-term results is important to determine the role of this method in the treatment of VUR in patients with spina bifida. PMID- 10517089 TI - [The so-called megaureter-megacystis syndrome: a case report]. AB - We encountered a patient with megaureter-megacystis syndrome showing a giant bladder and dilated ureters with marked reflux, which is very rare; to our knowledge, only 2 patients have been reported in Japan. The patient was a 4-year old boy, who showed inborn polyposia and polyuria, and proteinuria at the age of 1 year. He visited the pediatric department in our hospital complaining of cold like symptoms, stomachache and diarrhea. Urinary infection and kidney dysfunction were observed, and the patient was hospitalized for close examination. Bilateral pyelocaliceal hydronephrosis was detected by ultrasonography, and the patient was referred to our department. CT revealed bilateral hydronephrosis (right atrophic kidney), hydroureters and megacystis. Bilateral grade V vesicoureteral reflux, an increase in the bladder volume (> 300 ml), and urination without residual urine were noted by voiding cystourethrography. Uroflowmetry revealed that maximum flow rate was 21.6 ml/sec, voided volume was 110 ml, and residual volume was 24 ml. From these examinations, the patient was diagnosed as having megaureter megacystis syndrome, and underwent antireflux operation of the bilateral ureters using Cohen's procedures. PMID- 10517088 TI - [Development and characterization of a monoclonal antibody which recognizes a new prostate-organ specific antigen]. AB - PURPOSE: Development and characterization of monoclonal antibodies which recognizes a new prostate-organ specific antigen. METHOD: For development of monoclonal antibodies, hybrid cells were prepared by fusion of spleen cells of BALB/c mice immunized with the homogenates of surgically resected prostatic tissue and P 3 x Ag 8 U 1 (P 3 U 1) murine myeloma cells. Supernatants of hybrid clones were primarily screened using an ELISA on human prostatic cancer cell line PC-3 and human bladder cancer cell line T-24. In the secondary screening, they were tested on normal tissues by immunohistochemical staining. To characterize the antigens, biochemical analyses were performed using seminal plasma as an antigen by western blotting and gel filtration, and the reactivity of antibodies were compared with that of antibodies against prostatic acid phosphatase (PAP), prostate-specific antigen (PSA) and gamma-seminoprotein (gamma-Sm). RESULTS: A monoclonal antibody termed KP-9 was obtained and it only reacted with PC-3 and prostate tissues, but did not react with other cell lines and normal tissues. Immunohistochemical staining of prostate tissue revealed that KP-9 stained grandular epithelium and grandular exudate of normal and malignant prostatic tissues, and especially, strongly stained the apical site of grandular epithelium. Western blotting and gel filtration of seminal plasma suggested that the molecular weight of the KP-9 antigen was more than 300,000 and was different from PAP, PSA and gamma-Sm. CONCLUSION: We have developed a monoclonal antibody, KP-9 which specifically reacts with prostatic cancer as well as benign prostatic tissues. The antigen recognized by KP-9 appeared to be a new prostate-organ specific antigen and may be a useful marker for prostatic cancer such as PAP, PSA and gamma-Sm. PMID- 10517090 TI - [A case of submucosal endosalpingiosis in the urinary bladder]. AB - A 47-year-old woman was admitted when a mass in the urinary bladder was pointed out on ultrasound follow-up after hysterectomy for uterine myoma. Cystoscopy, ultrasonography, CT scan and MRI suggested a tumor in the muscle layers of the urinary bladder. Since the possibility of malignancy could not be ruled out, partial cystectomy was performed. The tumor was diagnosed as endosalpingiosis, a subclassification of mullerianosis histologically. The concept of endosalpingiosis has appeared recently and only 3 cases have been reported none of who had severe symptoms. Our case is the forth in the world. One of these cases had been treated with hormonal therapy as endometriosis, with no effect. Therefore, surgery is recommended as the first treatment of choice. PMID- 10517091 TI - [Etiologic, clinical and socio-democratic characteristics of acute diarrhea in Venezuela]. AB - In four cities of Venezuela a study was carried out to evaluate the epidemiological, clinical, and etiological characteristics of acute diarrhea in children under 5 years of age. The study was done between June 1993 and May 1995 and involved children who were seen in a hospital, 2,552 with diarrhea and 793 controls. The Fisher exact test was used for the statistical analysis of the results. Rotaviruses were the most important agents, both in terms of their frequency (30%) and their association with dehydration (58%). Following in importance were Campylobacter spp. (13%) and Escherichia coli classical O serogroups (9%), but their association with diarrhea was only statistically significant among children less than 3 months old, a fact that is particularly important from the standpoint of treatment. The importance of age was confirmed as a determining factor in the prevalence and severity of diarrhea. PMID- 10517092 TI - [Seroprevalence, epidemiology and ocular evaluation of human toxoplasmosis in the rural zone Jauguapita (Parana) Brazil]. AB - Toxoplasmosis is a protozoal zoonosis common among a great variety of species worldwide. The objective of this study was to assess the presence of toxoplasmosis among 345 residents in a rural area in Jagupita municipality, Parana state, Brazil. The frequency of titers in human serum samples was compared with the frequency of titers found in 1,420 samples obtained from various animal species with which local residents came into contact. Titers > or = 16 were considered positive. The highest titer found was 65,536 (1%), and the most frequent titer levels were 256 (29%) and 1,024 (19%). The comparisons between humans and animals revealed a positive and significant correlation between humans and felines (r = 0.78; P = 0.01) and humans and canines (r = 0.64; P = 0.05) in terms of titer distribution. Study participants were also tested with the Amsler grid. Seventy-five of the 345 people (22%) reported some type of ocular degradation. Of these 75, 58 of them (77%) were seropositive for toxoplasmosis. Forty-one of these 58 people underwent an ophthalmologic exam. Of these 41, 9 of them, who were between 34 and 78 years old, presented lesions characteristic of healed chorioretinitis, suggesting ocular toxoplasmosis. None of the 9 had ocular inflammation. Six of the 9 patients (67%) had unilateral lesions; 4 of these 6 presented a titer level of 256. The epidemiological survey showed that the probability of presenting ocular problems was 2.06 times as great for reactive patients as for nonreactive ones. No significant differences were observed in terms of sex, contact with felines or other animals, consumption of raw or rare meat and raw milk, and slaughtering of animals for personal consumption. Our results suggest that toxoplasmosis is common in the region, with a significant incidence of ocular lesions caused by Toxoplasma gondii. Health authorities should increase their monitoring and control activities in order to decrease the risk of toxoplasmic infections, especially among pregnant women. PMID- 10517093 TI - Assessment of nutritional status, cognitive development, and mother-child interaction in Central American refugee children. AB - A cross-sectional study was conducted between July and December 1992 to assess the nutritional status, cognitive development, and mother-child interactions in a group of 153 Nicaraguan refugee children living in Costa Rica. Nutritional status was assessed using anthropometric indices. Cognitive development was assessed with the Bayley Scale of Mental Development. Mother-child interaction was assessed with the Nursing Child Assessment Teaching Scale and Caldwell's Home Observation and Measurement of the Environment Inventory. Correlational analysis was performed to examine the relationship between child cognitive development scores and mother-child interaction measures and also between anthropometric measures and child cognitive development scores. Multiple regression analysis was performed to evaluate the relationship between the mother-child interaction measures and cognitive development scores, after adjusting by anthropometric measures. Thirty-three percent of the children were below the 10th percentile for height-for-age. There was no significant correlation between the total amount of mother-child interaction and child cognitive development. However, certain aspects of the home environment correlated with cognitive development, specifically the manner in which the mother responded emotionally and verbally to her child, and the organization of the child's physical and temporal environment. Multiple regression analysis revealed that the manner in which the mother responded and the child's weight-for-height were important in predicting child cognitive development. The child's weight-for-height and certain aspects of the home environment played an important role in the cognitive development of this refugee population. The findings indicate the importance of assessing nutritional status in this refugee population. PMID- 10517094 TI - [Home hospitalization for patients with acute illnesses]. AB - This study presents the results of 26 months of work, from January 1996 through February 1998, of the Distinct Home Hospitalization Service for Acute Patients. This service managed 20 home beds with two teams, each with a physician and a nurse, with a care approach similar to that for a room in a hospital. Among the items evaluated were the attributes of the admitted population, their illnesses, the form of administering drugs, the satisfaction of the caregivers, the indices of performance, and the costs with this approach. A total of 1,789 patients had home hospitalizations over the period, with a median stay of 4 days. Of the patients, 76.5% were admitted from in-hospital care. The most frequent illnesses were cardiorespiratory ones (45.5%), and the proportion of patients with a terminal illness was 14.2%. Drugs were administered orally with 74% of the patients and parenterally in 26%. The patients' satisfaction level was very high, and the cost of the hospitalization was 70% of that for in-hospital care. Family involvement was key in this approach to care. PMID- 10517095 TI - [Leishmaniasis: knowledge and practice in populations of the Pacific coast of Colombia]. AB - In 1997 a descriptive study with a qualitative emphasis was carried out in order to document, by gender, the knowledge and practices related to cutaneous leishmaniasis among inhabitants 14 years and older in seven communities of Colombia's Pacific coastal department of Choco. Since the control activities carried out by the Choco Sectional Health Services had not had the desired results, the residents of the region were at high risk of contracting leishmaniasis, which they called bejuco (liana) and yatevi. Qualitative data were collected by directly involving each community in discussion workshops and by interviewing knowledgeable informants. Using these materials as a foundation, the researchers prepared a survey with 10 closed-ended questions, which they administered to all persons over 14 years of age in each randomly chosen home visited. The results indicate that 94% of the population knew that leishmaniasis appeared as a skin disease; those not knowing that were more often women than men. With respect to the mode of transmission, 35% of the respondents connected the disease to the bite of an insect, but they did not what the etiologic agent was and thought that the bite was inflicted by a worm that lives in the mountains. In the communities studied, the residents used a great variety of treatments to cure the disease. The treatments were based on plants, chemical substances, burning the lesions with a piece of heated metal, and, to a lesser degree, drugs. Despite being responsible for taking care of sick persons in the household, women were not acquainted with the traditional treatments used in the community. This gender difference in treatment knowledge was statistically significant, the only such statistically significant gender difference found in the research. Of the people surveyed, 45% did not know how to prevent the disease. This was more often true for women; 102 of the 155 respondents saying they did not know how to prevent the disease were women. This research emphasizes the importance of studying the knowledge and practices of local inhabitants before designing and organizing educational programs to control leishmaniasis. PMID- 10517096 TI - [Arterial hypertension and alcoholism among workers in an oil refinery]. AB - The role of alcohol ingestion in the incidence of arterial hypertension has not been completely established. In addition, there are few studies addressing this point in relation to populations of workers. The objective of this study was to evaluate the association between alcoholism and arterial hypertension among workers in an oil refinery in Mataripe, Bahia, Brazil, from 1986 to 1993. We designed a retrospective cohort study with a 7-year follow-up in a stratified systematic sample of 335 workers from the refinery. Arterial hypertension was diagnosed based on blood pressure measurements done during routine medical examinations. At the beginning of follow-up, three groups were defined using the CAGE test of alcohol dependency: nondrinkers (n = 121), CAGE-negative workers (n = 116), and CAGE-positive workers (n = 98). In comparison with the CAGE-negative group, the CAGE-positive group had both greater relative risk and greater attributable risk for developing arterial hypertension (RR = 2.58; AR = 24.95 per 1,000 person-years). The CAGE-positive group also had greater risks compared to nondrinkers (RR = 2.06; AR = 20.97 per 1,000 person-years). The attributable fractions for the same two comparisons of groups were 61% and 51%, respectively. Rate standardization by age or smoking habit did not substantially change the results. Alcoholism is an important risk factor for arterial hypertension. PMID- 10517097 TI - Rapid surveys for program evaluation: design and implementation of an experiment in Ecuador. AB - This paper presents details from the field test of two rapid surveys in Ecuador in 1995. It focuses on how the surveys were designed and implemented, including descriptions of the sampling procedures, the preparation and use of preprogrammed palmtop computers for data entry, the selection criteria for the interviewing team, and how the training was designed. Lessons are drawn that will assist health professionals plan and carry out better rapid data collection in the future. The objective of the study was to evaluate the reliability and validity of data gathered during the rapid surveys as compared with a recent "gold standard" national survey. A two-way factorial design was used to control for differences in sampling (probability versus quasi-probability) and methods of data collection (paper versus palmtop computer). Few differences were detected between the surveys done on palmtops as compared to paper ones, but urban and rural differentials in contraceptive use were less pronounced in the rapid surveys than in the earlier, national survey. This suggests that caution should be exercised in interpreting the disaggregated data in these rapid surveys. In depth interviews revealed two features of the rapid surveys that were especially popular: the palmtops for their speed of data entry, and the short questionnaire for its "low impact" on a respondent's time. The common belief that computers would disturb respondents was not found to be the case. Even with no computer experience, the interviewers rapidly mastered the new technology. PMID- 10517098 TI - Evaluation of institutional cancer registries in Colombia. AB - The four primary objectives of this descriptive study were to: 1) design a quality-measurement instrument for institutional cancer registries (ICRs), 2) evaluate the existing ICRs in Colombia with the designed instrument, 3) categorize the different registries according to their quality and prioritize efforts that will efficiently promote better registries with the limited resources available, and 4) determine the institution with the greatest likelihood of successfully establishing Colombia's second population-based cancer registry. In 1990 the National Cancer Institute of Colombia developed 13 institution-based cancer registries in different Colombian cities in order to promote the collection of data from a large group of cancer diagnostic and treatment centers. During the first half of 1997, this evaluation reviewed 12 registries; one of the original 13 no longer existed. All of the Colombian institutions (hospitals) that maintain institution-based cancer registries were included in the study. At each institution, a brief survey was administered to the hospital director, the registry coordinator, and the registrar (data manager). Researchers investigated the institutions by looking at six domains that are in standard use internationally. Within each domain, questions were developed and selected through the Delphi method. Each domain and each question were assigned weights through a consensus process. In most cases, two interviewers went to each site to collect the information. The university hospitals in Cali, Pereira, and Medellin had substantially higher scores, reflecting a good level of performance. Four of the 12 institutions had almost no cancer registry work going on. Five of the 12 hospital directors considered that the information provided by the cancer registries influenced their administrative decisions. Three of the registries had patient survival data. Four of the institutions allocated specific resources to operate their cancer registries; in the other 8 hospitals there was no clear budget allocation. Seven of the hospital directors could not identify five or more objectives of a cancer registry. Data management was usually poor and resources insufficient at most of the institutions. In summary, the cancer registry system in Colombia varies greatly from institution to institution. A few of the hospitals do a good job while others have neglected the registries. The high, identical total scores for Pereira and Medellin suggest they would be good locations to establish new population-based cancer registries similar to the existing one in Cali. However, the overall characteristics in Pereira may provide a more appropriate environment for the second registry, with Medellin as an alternative. PMID- 10517099 TI - [A program to detect deficient quality of commercialized drugs]. AB - Defective drugs have been found during the postmarketing phase in various countries, regardless of those nations' level of economic development. This paper proposes guidelines for a program to uncover drug defects in the postmarketing surveillance period. Such a program should be run by a center or service organized for that purpose within the framework of a national drug surveillance system. PMID- 10517100 TI - [AIDS and sexually transmitted diseases in the Americas]. AB - This document provides a brief review of the situation of HIV/aids and of sexually transmitted infections (STIs) in the Americas. Among the subjects covered are appropriate care models for HIV/aids, including access to antiretroviral drugs; preventing transmission from mother to child; improved activities for preventing and controlling STIs; and interprogram activities to guarantee the safety of donated blood and blood products. PMID- 10517101 TI - [New dimensions in international health and globalization]. PMID- 10517102 TI - [Introduction: new world of coenzyme science]. PMID- 10517103 TI - [Built-in cofactors: amino acid residue-derived new cofactors]. PMID- 10517104 TI - [Discrimination and specific degradation of mRNAs: a new turn with T4 phage gene silencing]. PMID- 10517105 TI - [Searching for novel ribosomal function]. PMID- 10517106 TI - [Molecular genetics of bipolar disorder]. PMID- 10517107 TI - [Neurotoxicity of Alzheimer's beta-amyloid protein and membrane lipids]. PMID- 10517108 TI - [Current topics of cation transporters in animal, plant and bacteria]. PMID- 10517109 TI - [Genome informatics and structural genomics: progress of the bioinformatics]. PMID- 10517110 TI - [Analysis of electrophoretic pattern with CCD camera]. PMID- 10517111 TI - [DNA sequencing based on single molecule detection]. PMID- 10517112 TI - [Derailed oral anticoagulation with very high INR values and poor response to oral vitamin K--cholestasis as a possible cause]. AB - A 76-year-old man under long term oral anticoagulant treatment showed unclottable prothrombin time (PT) without overt bleeding. After oral administration of vitamin K1, PT remained severely prolonged and the patient was hospitalized. INR was 8.0 and responded to parenteral vitamin K. Cholestasis resulting in poor intestinal vitamin K resorption was assumed to have caused "overanticoagulation". Quick test is a global clotting test for the extrinsic and common pathways of the coagulation system. Increased PT, i.e. decreased Quick percentage, may be due to different conditions and should--if unexplained--be further analyzed by assaying factors II, V, VII, X and fibrinogen. Preanalytical problems, plasma dilution with clotting factor-free volume replacement, decreased vitamin K-dependent clotting factors (oral anticoagulation, intoxication with certain rodenticides, vitamin K deficiency), impaired liver synthetic capacity, disseminated intravascular coagulation, or massive heparin contamination may cause prolonged PT. Newborns physiologically have longer PT and should receive vitamin K prophylaxis. PMID- 10517113 TI - [Recurrent thromboembolisms despite oral anticoagulation in a 76-year-old patient -Trousseau syndrome]. AB - We report the case of a 76-year-old man with recurrent thromboses and low-grade chronic disseminated intravascular coagulation despite therapeutic oral anticoagulation with phenprocoumon. Work-up revealed a bronchial carcinoma (NSCCL) with hilar and mediastinal lymph node metastasis. The clinical condition was consistent with Trousseau's syndrome. Based on reports in the literature, the therapy was changed from phenprocoumon to intravenous unfractionated heparin (UFH), which was effective in controlling the thrombotic coagulopathy. For practical reasons, despite lack of established effectiveness in Trousseau's syndrome, therapy was switched to low-molecular-weight heparin (LMWH, nadroparine) in therapeutic dosage of 100 IU/kg body wt. subcutaneously 12 hourly. The patient remained free from further thromboembolic events during the last 6.5 months of his life. This case suggests that LMWH might be a convenient alternative to the established therapy with UFH in Trousseau's syndrome. PMID- 10517114 TI - [Skin necroses, thrombocytopenia and deep venous thrombosis in subcutaneous thromboembolism prophylaxis with heparin: heparin-induced thrombocytopenia (HIT) type II]. AB - We present a 52-year-old woman who developed a heparin-induced thrombocytopenia type II (HIT II) with deep vein thrombosis, thrombocytopenia and skin necrosis 7 days after initiating subcutaneous prophylaxis with 2 x 5000 U of unfractionated heparin. The platelet count fell from an initial value of 233 x 10(9)/L to 57 x 10(9)/L and normalized within 3 days after stopping heparin. Oral phenprocoumon was started, and her further course was uneventful. The pathogenesis and diagnosis of HIT II is illustrated, and the possible therapeutic options are discussed. To prevent this potentially lethal complication, it is important to begin oral anticoagulation on the first or second day of heparinization, and to stop heparin if the INR-value has been within a therapeutic range for 2 consecutive days. Platelet counts must be checked after 5 to 7 days of heparin therapy. In the case of suspected HIT II, a diagnostic test has to be performed, the heparin must be stopped, and an anticoagulation with either danaparoid or lepirudin is recommended. PMID- 10517116 TI - [Increased thrombin time in a patient with multiple myeloma]. AB - We describe a 56-year-old patient with multiple myeloma and very high paraprotein concentration (IgG kappa). Coagulation studies showed unclottable thrombin and reptilase times caused by impaired fibrin polymerization presumably due to the paraproteinemia. There was no obvious bleeding tendency. The differential diagnosis of thrombin time prolongation includes inhibition of the added thrombin by exogenous heparin, hirudin or seldom by endogenous heparin-like anticoagulants or by acquired (bovine) thrombin antibodies, qualitative fibrinogen disorders (congenital and acquired dysfibrinogenemia), quantitative fibrinogen disorders (severe hypo- and afibrinogenemia) and delayed fibrin polymerization due to fibrin/fibrinogen degradation products, paraproteins and antibodies against fibrin(ogen). In multiple myeloma, thrombin time prolongation may seldom be due to endogenous heparin-like anticoagulants or antibodies to thrombin and more frequently to impaired fibrin polymerization by paraproteins. Simultaneous reptilase time prolongation as present in this case hints to this latter possibility. PMID- 10517115 TI - [Patient with recurrent gastrointestinal hemorrhage in acquired von Willebrand disease]. AB - A 61-year-old female without prior history of bleeding experienced several symptomatic episodes of gastrointestinal bleeding in the last few years. A source of gastrointestinal bleeding could not be found although several tests of occult blood in the stool were positive. Diagnostically, a significant deficiency of von Willebrand factor could be found due to a shortened half-life of the von Willebrand factor. This acquired von Willebrand factor deficiency most likely can be explained by an immunological mechanism (e.g. antibodies against von Willebrand factor). Neither an immunosuppressive therapy with steroids nor cyclophosphamide could normalize the von Willebrand factor and the bleeding continued. Only tranexamic acid (inhibiting fibrinolysis) could at last stop the symptomatic episodes of gastrointestinal bleeding despite the fact of decreasing von Willebrand factor. PMID- 10517117 TI - [2-year-old boy with recurrent mucocutaneous hemorrhage--Glanzmann thrombasthenia. Role of the fibrinogen receptor (glycoprotein IIb-IIIa) in thrombocyte function]. AB - We report the clinical history of a patient suffering from a mucocutaneous bleeding diathesis since his birth. Discussing this case of Glanzmann's thrombasthenia we present essential aspects of clinical approach, diagnostic procedures and management of patients with platelet dysfunctions. Furthermore, we summarize the current understanding of platelet involvement in the haemostatic process and we discuss the role of the fibrinogen receptor glycoprotein IIb-IIIa. PMID- 10517118 TI - [APC resistance--most frequent familial thrombophilia]. AB - Two sisters having suffered from deep vein thrombosis while taking oral contraceptives are presented. Investigation for thrombophilia in 1993 was unrevealing. After the discovery of activated protein C (APC) resistance caused by the factor V R506Q (FV Leiden) mutation in 1993/1994, reinvestigation showed homozygous FV Leiden mutation in both sisters. APC resistance is the most frequent hereditary thrombophilia known so far. Diagnosis, prevalence and clinical significance of this thrombophilic defect are shortly discussed. PMID- 10517119 TI - [Isolated increased aPTT with anamnestic hemorrhagic diathesis--severe FXI deficiency]. AB - A 47-year-old patient with several episodes of bleeding after tonsillectomy, after an excision of a sacral dermoid and after urinary tract surgery presented with a mechanical ileus and was admitted to the department of visceral surgery. Preoperative analysis revealed a prolonged activated partial thromboplastin time (aPTT) of 93 seconds (normal range: 40-60 seconds), whereas the prothrombin time and thrombin time were normal. A mixture of 1 volume patient plasma and 1 volume normal plasma gave a normal aPTT, both before and after incubation of the plasma mixture at 37 degrees C for 1 hour. The patient's history and the prolonged aPTT as the only abnormal clotting test indicated deficiency of either factor VIII, factor IX, factor XI or von Willebrand factor with consecutively diminished factor VIII. Laboratory analysis revealed a factor XI of 4% indicating severe factor XI deficiency. Laparotomy was successfully performed without any hemorrhagic complications under fresh frozen plasma substitution. PMID- 10517120 TI - [A 26-year-old woman with splanchnic vein thrombosis as the initial manifestation of polycythemia vera]. AB - A 26-year-old woman, after cesarean section in the 33rd week of gestation, developed after delivery thrombosis of the popliteal vein, pulmonary embolism and thrombosis of the portal vein. After completion of a six month period of oral anticoagulation, laboratory investigations revealed diminished levels of plasminogen and free protein S antigen as well as APC-resistance due to heterozygous FV R506Q mutation. After six uneventful years, abdominal sonography and magnetic resonance examination, performed because of abdominal pain, showed liver cirrhosis with Budd-Chiari syndrome. Additional hematological investigations led to the diagnosis of polycythemia vera. Association of myeloproliferative disorders, mainly polycythemia vera, with splanchnic venous thrombosis is well known and should always be looked for. PMID- 10517121 TI - [A patient with isolated prolongation of aPTT without hemorrhagic diathesis anamnesis: severe, hereditary factor XII deficiency]. AB - By virtue of a severely prolonged aPTT with a normal thromboplastin time (prothrombin time) and a normal thrombin time, severe FXII deficiency has been diagnosed in a woman without a bleeding diathesis or a history of thromboembolic complications. A deficiency of a factor of the contact activation system (FXII, prekallikrein, high molecular weight kininogen) is usually diagnosed during routine coagulation tests demonstrating a prolonged aPTT. The severe and partial deficiency of FXII, of prekallikrein or high molecular weight kininogen is not associated with a bleeding tendency. In contrast, severely factor XI deficient subjects may suffer from a mild hemorrhagic diathesis, whereas FVIII deficiency (hemophilia A, autoimmune "hemophilia", von Willebrand disease) and FIX deficiency (hemophilia B) are associated with a bleeding tendency of varying severity, depending on the clotting activity of FVIII or FIX, respectively. An isolated prolongation of the aPTT due to a lupus anticoagulant, however, is frequently associated with arterial and/or venous thrombosis. Therefore, in case of a prolongation of the aPTT, its cause has to be determined. PMID- 10517122 TI - [Recurrent hepatitis in oral anticoagulation: coumarin-induced hepatitis]. AB - Coumarin hepatotoxicity resulting in a cholestatic or cytolytic hepatitis is a rare side effect in oral anticoagulant therapy, but has to be considered in any patient receiving coumarin treatment if hepatitis is diagnosed. Substitution with an other coumarin derivative is sometimes feasable, although cross-reactions between coumarin derivatives have been observed. PMID- 10517123 TI - [Life-long hemorrhagic diathesis in a young man with unclottable global coagulation tests--congenital afibrinogenemia]. AB - Congenital afibrinogenemia is a rare autosomal recessive hemostatic disorder leading to unclottable global coagulation tests. Furthermore, it is associated with abnormal platelet aggregation and with severe bleeding episodes if untreated. Surprisingly, thrombotic complications may be observed quite frequently in afibrinogenemic patients following replacement of fibrinogen. A case of congenital afibrinogenemia is described in a patient who suffered from severe bleeding episodes in the absence of replacement therapy but developed a deep vein thrombosis with multiple pulmonary emboli after fibrinogen replacement and surgical treatment of a hip fracture, despite conventional heparin prophylaxis. PMID- 10517124 TI - [Deep venous thrombosis of the leg in acquired thrombophilia- hyperhomocysteinemia as a sequela of undetected celiac disease]. AB - Today hyperhomocysteinemia is a well known and important risk factor for arteriosclerotic vascular and venous thromboembolic disease with a high prevalence in the general population. Elevation of plasma homocysteine levels are caused either by genetic defects in the enzymes involved in homocysteine metabolism or by nutritional deficiencies of vitamin cofactors (folate, vitamin B12, vitamin B6). A number of other factors may influence homocysteine metabolism, such as several disease states and medications. It has been demonstrated, that supplementation of folate, vitamin B12, or vitamin B6 can correct mild and moderate hyperhomocysteinemia. PMID- 10517125 TI - [Decreased Quick percentage, acquired factor X deficiency, hemarthrosis and ecchymosis: amyloidosis]. AB - A 68-year-old woman suffered from spontaneous hemarthrosis of the right elbow joint in april 1994. Cutaneous ekchymoses had been noted since summer 1993. Prothrombin time was prolonged (Quick percentage 40-50%) and was not corrected by prolonged administration of vitamin K. Coagulation studies showed isolated factor X (FX) deficiency without circulating FX inhibitor. This suggested the diagnosis of systemic amyloidosis which was retrospectively confirmed in small bowel biopsy specimens obtained one year before. Bilateral femoral head necrosis and femoral neck fracture due to amyloidosis, necessitated orthopedic surgery. The patient died four weeks postoperatively after several episodes of bleeding complications. Isolated FX deficiency may be hereditary but should--in the clinical context- also evoke the diagnosis of systemic amyloidosis. The possible bleeding tendency associated with amyloidosis is not attributable to acquired FX deficiency alone, but may also be caused by amyloid deposition in the microvasculature leading to acquired vascular hemorrhagic diathesis. PMID- 10517126 TI - [Chronic, hemorrhage-induced iron deficiency anemia in Osler disease]. AB - We report the history of a 77-year-old man with Osler-Weber-Rendu disease (hereditary hemorrhagic telangiectasia) who suffered from recurrent epistaxis and chronic bleeding anemia for many years. Hereditary hemorrhagic telangiectasia is a rare congenital disease, that is characterized by telangiectases of the nasal and oral mucosa, the gastrointestinal tract and the skin. Arteriovenous malformations of the lung and the brain may be present. The genetic and pathologic features, the clinical manifestations and the differential diagnosis are shortly presented. The management is not validated by controlled studies and is mainly based on clinical experience. It involves supportive therapy with iron supplementation and erythrocyte transfusions, if needed, for chronic bleeding anemia, aminocaproic acid, tranexamic acid or oestrogen-progesterone therapy to prevent mucosal bleeding, cauterisation, photocoagulation, transcatheter embolotherapy or surgery to treat the vascular abnormalities. PMID- 10517127 TI - [D-dimer determination in suspected deep venous thrombosis or lung embolism]. AB - Cleavage of crosslinked fibrin by the fibrinolytic enzyme plasmin leads to the formation of fibrin degradation products, among them D-dimers. D-Dimer can easily be measured in plasma or in whole blood by means of monoclonal antibodies directed against epitopes of the D-dimer fragment. Elevated plasma levels of D dimers are characteristic for patients with venous thromboembolism (DVT, PE), but occur also in patients with infectious diseases, malignant neoplasms and heart failure. Given the high sensitivity of ELISA D-dimer assays with respect to venous thromboembolism it is possible to reliably rule out DVT or PE when the plasma concentration of D-dimer is below the cut-off level. Thus, it is possible to rule out DVT in about 30% of outpatients with suspected venous thromboembolism by the measurement of D-dimer-concentration with a validated assay avoiding further diagnostic procedures. PMID- 10517128 TI - [Severe hemorrhage, lymphocytosis and leukoerythroblastic blood picture- disseminated intravascular coagulation in metastatic prostate carcinoma and chronic lymphatic leukemia]. AB - Hemorrhagic complications in CLL are most often related to thrombocytopenia either due to decreased megakaryopoiesis caused by diffuse lymphocytic bone marrow infiltration or to increased peripheral platelet consumption by platelet autoantibodies (immune thrombocytopenia), both being typical manifestations of the disease. Disseminated intravascular coagulation (DIC) does not belong to the spectrum of CLL-related complications. In the case here reported with DIC and newly diagnosed CLL the leukoerythroblastic blood changes hinted to bone marrow infiltration by another neoplastic process in addition to CLL. Bone marrow biopsy revealed infiltration by metastatic carcinoma of the prostate in addition to that by CLL, the former being responsible for triggering severe DIC leading to fatal intracranial bleeding. PMID- 10517129 TI - [32-year-old patient with systemic lupus erythematosus, thrombocytopenia and ischemic cerebrovascular insult and antiphospholipid antibodies]. AB - We report a young woman suffering from systemic lupus erythematosus complicated by severe thrombocytopenia and high-level antiphospholipid antibodies. Thrombocytopenia was refractory to any therapy. In the course of the illness the patient developed an ischemic cerebrovascular stroke. We discuss the clinical features of the antiphospholipid antibody syndrome being primarily characterized by recurrent thromboembolism and address the appropriate prophylactic measures to avoid recurrent events. PMID- 10517130 TI - [46-year-old woman with multiple hematomas and bleeding of the base of the tongue: phenprocoumon poisoning]. AB - A 46-year old nurse complaining of multiple hematomas including bleeding into the tongue was referred for hemostasis evaluation. A very low Quick percentage value, i.e. a severely prolonged prothrombin time with severely depressed vitamin K dependent coagulation factors (FII:C, FVII:C, FX:C) and normal FV:C and fibrinogen level was found. In the absence of cholestasis, malabsorption and broad-spectrum antibiotic therapy, ingestion of vitamin K antagonists was suspected. Three years previously, she had been on oral anticoagulant treatment with phenprocoumon (Marcoumar) for postoperative pulmonary embolism. She denied having voluntarily ingested anticoagulant drugs. A high plasma level of coumarins was found. To exclude accidental ingestion, the patient's son living in the same household was tested as well. Surprisingly, a low level of coumarin was found also in his plasma. We suspect that the patient voluntarily intoxicated herself and gave a low dose of coumarin anticoagulant to her son as well. PMID- 10517132 TI - Health impact of earthquake, Turkey. PMID- 10517131 TI - [An infant with umbilical cord and intracranial hemorrhage--severe factor XIII deficiency]. AB - Based on the description of a severe bleeding disorder in a young child a short overview on the genetics, the epidemiology, the pathophysiology, the clinical manifestations and the laboratory diagnosis of factor XIII deficiency is presented. The impressive clinical signs with bleeding starting in the neonatal period (prolonged bleeding from the umbilical cord), followed by severe, life threatening episodes of intracranial hemorrhages should raise the clinical suspicion of FXIII deficiency. Difficulty of laboratory diagnosis is stressed. The importance of repeating initially negative screening tests and of performing a quantitative FXIII assay in the presence of strong clinical suspicion is strengthened. The diagnosis of factor XIII deficiency is difficult but has important therapeutic consequences: patients with documented severe deficiency should be put on regular substitution with factor XIII concentrates. Appropriately timed periodic infusions of such factor XIII concentrates enable patients to live normal lives, free from catastrophic bleeding episodes. PMID- 10517133 TI - Chagas disease, Venezuela, 1999. PMID- 10517134 TI - Echinococcosis, Uzbekistan. PMID- 10517135 TI - MALDI-TOFMS of chemically modified recombinant hepatitis B surface antigen. PMID- 10517136 TI - Affinity NMR. AB - Because binding ligands are directly detected and identified, the diffusion-based NMR method and NOE pumping approach promise to greatly simplify deconvolution in drug screening. An additional advantage of these techniques is that low-affinity ligands, which might be missed by high-throughput screening, can be detected and could serve as synthetic precursors for higher affinity ligands. The biggest challenge to NMR methodology lies in its sensitivity. Compared with other techniques, such as MS (25), NMR methods for screening mixtures are limited by their relative insensitivity. Because of issues such as solubility, stability, and mass limitation, it is not in general judicious to simply increase the concentration of the mixture. Improvements in hardware and software are necessary to extend the applicability of the affinity NMR method to the screening of larger and more complex mixtures. A boost in sensitivity and screening capacity of NMR technique is possible by the implementation of microcoil (26) and flow probe techniques. An upsurge in the capabilities of mixture analysis could be achieved with a combination of independent and complementary techniques (e.g., HPLC, MS) (27). As a unique, nondestructive, and versatile tool, NMR will continue on its fast track of development in the support of drug discovery. PMID- 10517137 TI - SNP mining. The rush is on. PMID- 10517138 TI - Prediction of substructure and toxicity of pesticides with temperature constrained-cascade correlation network from low-resolution mass spectra. AB - Artificial neural networks are trained to predict the toxicity or active substructures of organophosphorus pesticides and then are applied to screening GC/MS data for environmentally hazardous compounds. Every mass spectral scan in the chromatographic run is classified, and separate chromatograms are obtained for either toxicity or substructure classes. Classification of mass spectra allows the detection of chromatographic peaks from potentially hazardous compounds that may be missing from the reference database. The neural network models predict substructures and toxicity from mass spectra without first determining the complete configurational structure of the pesticides. Temperature constrained-cascade correlation networks (TCCCN) were used because they are self configuring networks that train rapidly and robustly. The toxicity classes are defined by the World Health Organization, and the substructure classes are standard organophosphorus pesticide groupings. The TCCCN models are used to mathematically resolve peaks in the chromatograms by substructure and toxicity. Evaluations yielded classification rates of 97 and 84% for substructure and toxicity, respectively. PMID- 10517139 TI - Multicomponent quantification of diastereomeric hexosamine monosaccharides using ion trap tandem mass spectrometry. AB - A rapid means of stereochemical differentiation and quantification for the hexosamine monosaccharides was achieved using electrospray ionization quadrupole ion trap mass spectrometry. The hexosamine monosaccharides, glucosamine, galactosamine, and mannosamine, were derivatized with [Co(DAP)2Cl2]Cl, and the complex [Co(DAP)2(HexNH2)]Cl was generated. Subjecting this complex to collision induced dissociation provided a unique product ion spectrum for each of the diastereomeric monosaccharide complexes, thus differentiating the stereoisomers. Furthermore, the stereoisomers were quantified. This was achieved by using the relative abundances of product ions from pure standards and using these values to determine the ratio of isomeric products in a mixture. The utility of this quantification method was demonstrated by successfully determining the composition of two- and three-component mixtures of the hexosamines. PMID- 10517140 TI - Reconstitution of acid-denatured holomyoglobin studied by time-resolved electrospray ionization mass spectrometry. AB - Time-resolved electrospray ionization (ESI) mass spectrometry (MS) is a new technique for studying the kinetics of protein folding reactions. It can monitor both changes in the protein conformation and the loss or binding of protein ligands as a function of time. Time-resolved ESI MS was previously used to monitor the acid-induced unfolding of holomyoglobin (hMb). The native form of this protein is characterized by a tightly folded conformation and a heme group that is noncovalently attached to the protein. Acid-induced denaturation induces substantial unfolding of the polypeptide chain and disruption of the heme-protein interactions. In this work, time-resolved ESI MS is used to study the reverse reaction, i.e., reconstitution of acid-denatured hMb. To examine the mechanism and the kinetics of this reaction, a continuous-flow setup with two sequential mixing steps was developed. The data presented in this work show that reconstitution involves the formation of various short-lived intermediates such as tightly folded myoglobin without a heme group and several nativelike forms of the protein that are bound to more than one heme. The occurrence of these transient states is most likely due to the rapid aggregation of free heme in solution. PMID- 10517141 TI - The dominance of arginine-containing peptides in MALDI-derived tryptic mass fingerprints of proteins. AB - Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is a powerful tool for mass finger-printing of peptide mixtures obtained after enzymatic ingel digestion of proteins separated by two-dimensional electrophoresis (2-DE). In the course of a proteome analysis of mycobacteria using mass spectrometric identification, it was found that 94% of the most intense MALDI-MS peaks denote peptides bearing arginine at the C-terminal end. The effect was demonstrated to be equally prominent using an equimolar mixture of the synthetic peptides known to be present in the tryptic digest of the mycobacterial 35 kDa antigen ("synthetic mass map"). In addition, several binary mixtures of synthetic peptides differing exclusively at the C terminus (Arg or Lys) were examined to rationalize the higher sensitivity toward arginine containing peptides. The extent of the effect described depends on the matrix used and may facilitate a more reliable assignment of mass fingerprint data to protein sequences in databases. PMID- 10517142 TI - Fluorescence polarization studies of affinity interactions in capillary electrophoresis. AB - Capillary electrophoresis (CE) combined with molecular recognition for ultrasensitive bioanalytical applications often requires the formation of stable complexes between an analyte and its binding partner. Previous studies of binding interactions using CE involve multiple-step titration experiments and are time consuming. We describe a simple method based on laser-induced fluorescence polarization (LIFP) detection for CE separation, which allows for on-line monitoring of affinity complex formation. Because fluorescence polarization is sensitive to changes in the rotational diffusion arising from molecular association or dissociation, it is capable of providing information on the formation of affinity complexes prior to or during CE separation. Applications of the CE/LIFP method to three binding systems including vancomycin and its antibody, staphylococcal enterotoxin A and its antibody, and trp operator and trp repressor were demonstrated, representing peptide-protein, protein-protein, and DNA-protein interactions. The affinity complexes were readily distinguished from the unbound molecules on the basis of their fluorescence polarization. The relative increase in fluorescence polarization upon complex formation varied with the molecular size of the binding pairs. PMID- 10517143 TI - Development of an ultrasensitive immunoassay for rapid measurement of okadaic acid and its isomers. AB - This report highlights the characteristics of an okadaic acid immunoassay with limits of detection in the subfemtomole range. Two different immunoassay formats were investigated and their characteristics compared in relation to linear ranges, limits of detection, and cross-reactivity with other seafood toxins present in water and/or mussel samples. The developed ELISA system can be manipulated to quantitatively measure total diarrhetic shellfish poisoning (DSP) content or for okadaic acid and dinophysistoxin-1 individual concentrations by variation of the format of the immunoassay. Real mussel samples were validated in percentage recovery test. Calibration curves were established, and aliquots of real samples were tested. Very good recoveries were attained, highlighting the validity of the ELISA system to accurately determine the DSP concentration in mussel samples. PMID- 10517144 TI - Tin speciation in the femtogram range in open ocean seawater by gas chromatography/inductively coupled plasma mass spectrometry using a shield torch at normal plasma conditions. AB - A sensitive method for the determination of ultratrace organotin species in seawater is described. The merits and demerits of derivatization methods using Grignard reagent or sodium tetraethylborate (NaBEt4) were evaluated in terms of derivatization efficiency, applicability to the programmed temperature vaporization (PTV) method, and procedural blanks. The sensitivity of the gas chromatography/inductively coupled plasma mass spectrometry (GC/ICPMS) was improved by more than 100-fold by operating the shield torch at normal plasma conditions, compared with that obtained without using it. The absolute detection limit as tin reached subfemtogram (fg) levels. Furthermore, the detection limit in terms of relative concentration was improved 100-fold by using the PTV method, which enabled the injection of a large sample volume of as much as 100 microL without loss of analyte. When the organotin species in seawater were extracted into hexane with a preconcentration factor of 1000 after ethylation with NaBEt4 and a 100 microL aliquot of the extract was injected into the GC, the instrumental detection limit in relative concentration reached 0.01 pg/L in original seawater. Sources of contamination of organotin species during the sample preparation were examined, and a purification method of NaBEt4 was developed. Finally, the method was successfully applied to open ocean seawater samples containing organotin species at the level of 1-100 pg/L. PMID- 10517145 TI - A minisonicator to rapidly disrupt bacterial spores for DNA analysis. AB - Concerns about the use of anthrax spores as a weapon of mass destruction have motivated the development of portable instruments capable of detecting and monitoring a suspected release of the agent. Optimal detection of bacterial spores by PCR requires that the spores be disrupted to make the endogenous DNA available for amplification. The entire process of spore lysis, PCR, and detection can take several hours using conventional methods and instruments. In this report, a minisonicator and prototype spore lysis cartridge were built to disrupt Bacillus spores in 30 s for rapid, real-time PCR analysis. Utilization of the minisonicator improved PCR analysis by decreasing the limit of detection, reducing the time of detection, and increasing the signal amplitude. Total time of spore disruption and detection using the minisonicator and a microchip PCR instrument was less than 15 min. PMID- 10517146 TI - Automated in-tube solid-phase microextraction coupled with liquid chromatography/electrospray ionization mass spectrometry for the determination of beta-blockers and metabolites in urine and serum samples. AB - The technique of automated in-tube solid-phase microextraction (SPME) coupled with liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) was evaluated for the determination of beta-blockers in urine and serum samples. In-tube SPME is an extraction technique for organic compounds in aqueous samples, in which analytes are extracted from the sample directly into an open tubular capillary by repeated draw/eject cycles of sample solution. LC/MS analyses of beta-blockers were initially performed by liquid injection onto a LC column. Nine beta-blockers tested in this study gave very simple ESI mass spectra, and strong signals corresponding to [M + H]+ were observed for all beta-blockers. The beta blockers were separated with a Hypersil BDS C18 column using acetonitrile/methanol/water/acetic acid (15:15:70:1) as a mobile phase. To optimize the extraction of beta-blockers, several in-tube SPME parameters were examined. The optimum extraction conditions were 15 draw/eject cycles of 30 microL of sample in 100 mM Tris-HCl (pH 8.5) at a flow rate of 100 microL/min using an Omegawax 250 capillary (Supelco, Bellefonte, PA). The beta-blockers extracted by the capillary were easily desorbed by mobile-phase flow, and carryover of beta-blockers was not observed. Using in-tube SPME/LC/ESI-MS with selected ion monitoring, the calibration curves of beta-blockers were linear in the range from 2 to 100 ng/mL with correlation coefficients above 0.9982 (n = 18) and detection limits (S/N = 3) of 0.1-1.2 ng/mL. This method was successfully applied to the analysis of biological samples without interference peaks. The recoveries of beta-blockers spiked into human urine and serum samples were above 84 and 71%, respectively. A serum sample from a patient administrated propranolol was analyzed using this method and both propranolol and its metabolites were detected. PMID- 10517147 TI - Localization of labile posttranslational modifications by electron capture dissociation: the case of gamma-carboxyglutamic acid. AB - Tandem mass spectrometry (MS/MS) of 28 residue peptides harboring gamma carboxylated glutamic acid residues, a posttranslational modification of several proenzymes of the blood coagulation cascade, using either collisions or infrared photons results in complete ejection of the gamma-CO2 moieties (-44 Da) before cleavage of peptide-backbone bonds. However, MS/MS using electron capture dissociation (ECD) in a Fourier transform mass spectrometer cleaves backbone bonds without ejecting CO2, allowing direct localization of this labile modification. Sulfated side chains are also retained in ECD backbone fragmentations of a 21-mer peptide, although CAD causes extensive SO3 loss. ECD thus is a unique complement to conventional methods for MS/MS, causing less undesirable loss of side-chain functionalities as well as more desirable backbone cleavages. PMID- 10517148 TI - A biomimetic phospholipid/alkanethiolate bilayer immobilizing uricase and an electron mediator on an Au electrode for amperometric determination of uric acid. AB - A biomimetic bilayer membrane immobilizing uricase (urate oxidase; EC 1.7.3.3) (UOx) and a redox agent of 1-methoxy-5-methylphenazinium (MMP) was fabricated on an Au electrode substrate with use of the Au substrate coated with a self assembled monolayer of n-octanethiolate (OT/Au) and L-alpha-phosphatidylcholine beta-oleoyl-gamma-palmitoyl (PCOP). The preparation was carried out by successively immersing an Au electrode substrate in an ethanol solution of OT, an MMP aqueous solution, and a suspension of proteoliposome formed by PCOP containing UOx and MMP. The prepared electrode exhibited such fast steady amperometric responses to uric acid as to allow its determination within 20 s after injecting uric acid, indicating that UOx-catalyzed electrochemical oxidation of uric acid was accomplished with assistance of electron mediation by MMP between UOx and the Au substrate. An increase in the response currents with increasing concentration of uric acid was obtained in a concentration range of uric acid found in healthy human blood. Any interference in the current response that is caused by direct anodic oxidation of uric acid or ascorbic acid was not observed at the prepared sensor electrode because the densely packed bilayer effectively blocked the diffusion of these substrates toward the Au surface, making it possible to determine amperometrically uric acid at the electrode with high precision. PMID- 10517149 TI - Dual detection of peptides in a fluorescence binding assay by employing genetically fused GFP and BFP mutants. AB - A competitive fluorescence microplate assay based on a red-shifted green fluorescent protein (rsGFP) and a blue fluorescent protein (BFP) was developed for the detection of two model peptides in the same sample. The assay employed gene fusion to prepare the fluorescently labeled peptide conjugates. Specifically, plasmids were constructed in which the genes encoding for the two small peptides (less than 12 amino acids in length) were fused to either the gene of the rsGFP or the BFP, as desired. The newly constructed plasmids were transformed into E. coli for expression of the fusion proteins. By employing the technique of gene fusion, one-to-one homogeneous populations of peptide-rsGFP or BFP conjugates were produced. These peptide-GFP mutant conjugates exhibited the same excitation and emission spectral characteristics as the unmodified proteins. The naturally fluorescent proteins act as labels to provide sensitive dual detection of the two selected small peptides in a competitive assay format. To our knowledge, this is the first time that mutants of GFP, such as the rsGFP and BFP, have been used as quantitative labels for the development of a dual-analyte fluorescence immunoassay. PMID- 10517150 TI - Multiple-analyte fluoroimmunoassay using an integrated optical waveguide sensor. AB - A silicon oxynitride integrated optical waveguide was used to evanescently excite fluorescence from a multianalyte sensor surface in a rapid, sandwich immunoassay format. Multiple analyte immunoassay (MAIA) results for two sets of three different analytes, one employing polyclonal and the other monoclonal capture antibodies, were compared with results for identical analytes performed in a single-analyte immunoassay (SAIA) format. The MAIA protocol was applied in both phosphate-buffered saline and simulated serum solutions. Point-to-point correlation values between the MAIA and SAIA results varied widely for the polyclonal antibodies (R2 = 0.42-0.98) and were acceptable for the monoclonal antibodies (R2 = 0.93-0.99). Differences in calculated receptor affinities were also evident with polyclonal antibodies, but not so with monoclonal antibodies. Polyclonal antibody capture layers tended to demonstrate departure from ideal receptor-ligand binding while monoclonal antibodies generally displayed monovalent binding. A third set of three antibodies, specific for three cardiac proteins routinely used to categorize myocardial infarction, were also evaluated with the two assay protocols. MAIA responses, over clinically significant ranges for creatin kinase MB, cardiac troponin I, and myoglobin agreed well with responses generated with SAIA protocols (R2 = 0.97-0.99). PMID- 10517151 TI - Enhanced synchronous spectrofluorometric determination of tetracycline in blood serum by chemometric analysis. Comparison of partial least-squares and hybrid linear analysis calibrations. AB - Tetracycline has been determined in human serum samples by a combination of: (1) synchronous fluorescence spectra of whole sera treated with Mg2+, and (2) the multivariate calibration methods of partial least-squares (PLS-1) and a variant of the recently introduced hybrid linear analysis (HLA), which does not require the knowledge of pure-component spectra. The calibration set was designed with 50 sera spiked with concentrations of tetracycline in the range 0.0-4.0 micrograms mL-1'. Studies concerning validation, precision, accuracy and figures of merit (selectivity, sensitivity and limit of determination) were also carried out. A novel wavelength-selection procedure was applied to minimize the effect of nonmodeled interferents present in serum samples containing bilirubin, triglycerides, and salicylate. Overall, the performance of the newly developed HLA approach seems to be better than that of PLS-1. PMID- 10517152 TI - Identification of intact proteins in mixtures by alternated capillary liquid chromatography electrospray ionization and LC ESI infrared multiphoton dissociation Fourier transform ion cyclotron resonance mass spectrometry. AB - Here we propose a novel method for rapidly identifying proteins in complex mixtures. A list of candidate proteins (including provision for posttranslational modifications) is obtained by database searching, within a specified mass range about the accurately measured mass (e.g., +/- 0.1 Da at 10 kDa) of the intact protein, by capillary liquid chromatography electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (LC ESI FT-ICR MS). On alternate scans, LC ESI infrared multiphoton dissociation (IRMPD) FT-ICR MS yields mostly b and y fragment ions for each protein, from which the correct candidate is identified as the one with the highest "hit" score (i.e., most b and y fragments matching the candidate database protein amino acid sequence masses) and sequence "tag" score (based on a series of fragment sequences differing in mass by 1 or 2 amino acids). The method succeeds in uniquely identifying each of a mixture of five proteins treated as unknowns (melittin, ubiquitin, GroES, myoglobin, carbonic anhydrase II), from more than 1000 possible database candidates within a +/- 500 Da mass window. We are also able to identify posttranslational modifications of two of the proteins (mellitin and GroES). The method is simple, rapid, and definitive and is extendable to a mixture of affinity-selected proteins, to identify proteins with a common biological function. PMID- 10517153 TI - Detection of low dose radiation induced DNA damage using temperature differential fluorescence assay. AB - A rapid and sensitive fluorescence assay for radiation-induced DNA damage is reported. Changes in temperature-induced strand separation in both calf thymus DNA and plasmid DNA (puc 19 plasmid from Escherichia coli) were measured after exposure to low doses of radiation. Exposures of between 0.004 and 1 Gy were measured with doses as low as 0.008 Gy yielding significant responses. The double strand, sensitive dye PicoGreen was used as an indicator of DNA denaturation. Calibration plots indicate that fluorescence changes corresponding to amounts as low as 1 ng of double stranded DNA (10(6) copies for plasmid puc 19) are detected by this method. PMID- 10517154 TI - Uniquely folded mini-protein motifs. AB - Mini-proteins containing fewer than 40 amino acids provide simple model systems for studying protein folding and stability as well as serving as scaffolds for the rational design of new functional motifs. This article reviews current progress on the design and characterization of discretely folded mini-protein motifs. PMID- 10517156 TI - De novo design: backbone conformational constraints in nucleating helices and beta-hairpins. AB - A modular approach to synthetic protein design is being developed using conformationally constrained amino acid as stereochemical directors of polypeptide chain folding. An overview of studies aimed at constructing peptide helices using alpha,alpha-dialkyated residues and beta-hairpins using D-Pro as a turn nucleator is presented. The construction of helix-helix motifs and three- and four-stranded structures has been achieved using non-protein amino acids to stabilize specific elements of secondary structures. PMID- 10517155 TI - Extending the concept of template-assembled synthetic proteins. AB - The creation of native-like macromolecules in copying nature's way represents a fascinating challenge in protein chemistry today. In the absence of a detailed knowledge of the complex folding pathway the ultimate goal in protein de novo design, the construction of artificial proteins with predetermined three dimensional structure and tailor-made functions based on a defined, generally valid set of rules, appears to be still out of reach. With progress in synthesis strategies and biostructural characterization methods, topological templates have become a versatile tool for inducing and stabilizing secondary and tertiary structures, such as protein loops, beta-turns, alpha-helices, beta-sheets and a variety of folding motifs. In this article, we extend the concept of template assembled synthetic proteins for the construction of protein-like topologies with multiply bridged, oligocyclic chain architectures termed locked-in tertiary folds that exhibit unique physicochemical and folding properties because of the highly confined conformational space. Furthermore, we show that some fundamental questions in protein assembly can be approached applying the template concept. Using covalent template trapping of self-associated peptide assemblies in aqueous solution the structural and physical forces guiding protein folding, supramolecular assembly and molecular recognition processes can be studied on a molecular level. PMID- 10517157 TI - Perspective: peptides as mimics of transmembrane segments in proteins. AB - Peptide-based approaches to protein structure within membranes have proven enormously valuable. When one focusses on the detailed manner through which membrane proteins actually traverse the cell bilayer, a simple observation emerges: helical peptide segments of 20 amino acids each constitute the only tangible connection between the inside and outside of the cell. Thus, a major step towards understanding the key relationships between biological function and membrane protein structure can be taken through characterization, by composition, sequence, chain length, hydrophobicity and conformation, of hydrophobic peptides designed as mimics of transmembrane segments. PMID- 10517158 TI - The twists and turns of beta-peptides. AB - Recently, it has been discovered that peptides composed of beta-amino acids are capable of adopting novel secondary structures demonstrating that peptides composed of alpha-amino acids are not unique in their ability to fold into well defined structures. Cyclic as well as acyclic peptides composed of beta-amino acid residues adopt turn, helical, and sheet-like conformations. Here, we discuss the synthesis and conformational preferences of individual, substituted beta amino acids as well as the structures that peptides composed of these residues, beta-peptides, may adopt. PMID- 10517159 TI - Characterization of epitope regions of thyrotropin beta-subunit recognized by the monoclonal antibodies mAb279 and mAb299: a chimeric peptide approach. AB - This investigation describes the design, synthesis and evaluation of chimeric peptides related to the bovine thyrotropin beta-subunit, bTSHbeta. The structures of these chimeric peptides were derived from investigations with linear peptides and sequence alignment studies, in association with a homology model of TSHbeta developed from the hCG X-ray crystallographic structure. The structures of these chimeric peptides comprised beta-turn regions of loop L1 [bTSHbeta(14-20)] and loop L3 [bTSHbeta(65-72)] held in close proximity by a bis-beta-alanine linker and the disulfide bond bTSHbeta[Cys16-Cys67]. Linear and cyclic chimeric peptides were evaluated in immunochemical assays for their ability to inhibit the binding of radio-iodinated bTSHbeta [125I-bTSHbeta] to the monoclonal antibodies, mAb279 and mAb299. Previously, mAb279 and mAb299 have been shown to recognize epitopes accessible on the surface of TSHbeta that lie in close proximity to the TSH receptor-binding site. The results indicate that these chimeric peptides can specifically inhibit in a dose-dependent manner the binding of 125I-bTSHbeta to mAb299, while having a lesser effect on the binding with mAb279. Based on these results, it can be concluded that the bTSHbeta-epitope recognized by mAb299 involves contributions from amino residues from the beta-turn regions of the L1 and L3 loops of TSHbeta, and that these loop regions flank part of the receptor binding site of the bTSH beta-subunit. PMID- 10517160 TI - Electrostatic effects on the alpha-helix and beta-strand formation of BPTI(16-36) studied by Monte Carlo simulated annealing. AB - The electrostatic effects on the secondary structure forming tendencies of a peptide fragment with residues 16-36 of bovine pancreatic trypsin inhibitor, BPTI(16-36), are studied using Monte Carlo simulated annealing simulations. We consider three dielectric functions epsilon(r) of distance r: constant dielectric function (epsilon = 2; strong electrostatic interactions) and sigmoidal functions varying from epsilon(0) = 2 to epsilon(infinity) = 47 (intermediate) and to epsilon(infinity) = 78 (weak). Simulations with epsilon = 2 suggest that this peptide exhibits a significant propensity for beta-strand formations in accordance with a beta-sheet structure of the relevant segment in native BPTI. The tendency for alpha-helix formations becomes almost comparable with that of beta-strands in the simulation with epsilon(infinity) = 47, and there appears no appreciable conformational propensity for this case. Finally, the results with epsilon(infinity) = 78 generate low-energy conformations with conspicuous alpha helices. These findings suggest the possibility that the change in electrostatic interactions can be the key factor for the conformational transitions of peptides between alpha-helix and beta-sheet that have recently been observed in experiments. These changes in electrostatic interactions can arise from those in various environmental factors such as conformations of the rest of the protein molecule and solvent conditions. PMID- 10517162 TI - Conformational studies of irreversible HIV-1 protease inhibitors containing cis epoxide as an amide isostere. AB - We have carried out NMR and molecular modeling studies of peptidomimetic HIV-1 protease inhibitors, LB71116: Qc-Asn-Phepsi[(1R,2S)-cis-epoxide]Gly-NH CH(isopropyl)2 where Qc stands for quinaldic acid and LB71148: Qc-(SMe)Pen(O)2 Phepsi[(1R,2S)-cis-epoxide]Gly-NH-CH(isoprop yl)2 where (SMe)Pen(O)2 stands for S methyl-S-dioxo-penicillamine. Through conformational calculations and NMR data analysis, we have obtained preferred conformations of the two inhibitors in solution. To our knowledge, this work is one of the first extensive conformational studies of peptidomimetics containing cis-epoxide amide isostere. The resulting preferred conformations contain extended structures. In these conformations, the psi of Phe(cep) is maintained about 130 degrees and the phi angle of (cep)Gly prefers +/- 150 degrees [where Phe(cep) and (cep)Gly are the residues generated by the replacement of the Phe-Gly peptide bond with cis epoxide]. Two conformations were commonly observed in the preferred conformations of each inhibitor. Through restrained molecular dynamics simulating the hydrogen bond formation between our inhibitor and a water molecule ('flap water'), one of the conformations is assumed as the conformation which can bind to the enzyme without large conformational changes. Recently, we had the opportunity to compare the selected preferred conformation with the binding conformation of LB71116 observed from the X-ray studies of the complex between LB71116 and HIV-1 protease. These two conformations are surprisingly similar to each other. Thus, we can explain high activity and selectivity of our inhibitors to the HIV-1 protease by the similarity between the preferred conformations in solution and the binding conformation. PMID- 10517161 TI - Antibacterial activity of multiple antigen peptides homologous to a loop region in human lactoferrin. AB - An 11-residue peptide (FQWQRNMRKVR) homologous to just over half the loop region of human lactoferricin is thought to be responsible for antimicrobial properties of human lactoferricin. Multiple antigen peptides (MAP) of the 11-residue peptide exerted significant antibacterial effects against a broad spectrum of bacteria including MRSA. More than eight branching was favourable for increasing its antibacterial activity. Our report shows a novel possibility for MAP to increase the activity of antibiotic peptides other than simply to stimulate antibody production, as reported so far. PMID- 10517163 TI - Design of peptides with alpha,beta-dehydro-residues: synthesis, and crystal and molecular structure of a 3(10)-helical tetrapeptide Boc-L-Val-deltaPhe-deltaPhe-L Ile-OCH3. AB - The peptide Boc-L-Val-deltaPhe-deltaPhe-L-Ile-OCH3 was synthesized using the azlactone method in the solution phase, and its crystal and molecular structures were determined by X-ray diffraction. Single crystals were grown by slow evaporation from solution in methanol at 25 degrees C. The crystals belong to an orthorhombic space group P2(1)2(1)2(1) with a = 12.882(7) A, b = 15.430(5) A, c = 18.330(5) A and Z = 4. The structure was determined by direct methods and refined by a least-squares procedure to an R-value of 0.073. The peptide adopts a right handed 3(10)-helical conformation with backbone torsion angles: phi1 = 56.0(6)degrees, psi1 = -38.0(6)degrees, phi2 = -53.8(6)degrees, psi2 = 23.6(6)degrees, phi3 = -82.9(6)degrees, psi3 = -10.6(7)degrees, phi4 = 124.9(5)degrees. All the peptide bonds are trans. The conformation is stabilized by intramolecular 4-->1 hydrogen bonds involving Boc carbonyl oxygen and NH of deltaPhe3 and CO of Val1 and NH of Ile4. It is noteworthy that the two other chemically very similar peptides: Boc-Val-deltaPhe-deltaPhe-Ala-OCH3 (i) and Boc Val-deltaPhe-deltaPhe-Val-OCH3 (ii) with differences only at the fourth position have been found to adopt folded conformations with two overlapping beta-turns of types II and III', respectively, whereas the present peptide adopts two overlapping beta-turns of type III. Thus the introduction of Ile at fourth position in a sequence Val-deltaPhe-deltaPhe-X results in the formation of a 3(10)-helix. The crystal structure is stabilized by intermolecular hydrogen bonds involving NH of Val1 and carbonyl oxygen of a symmetry related (-x, y - 1/2, 1/2 + z) deltaPhe2 and NH of deltaPhe2 with carbonyl oxygen of a symmetry related (x, y1/2, 1/2 + z) Ile4. This gives rise to long columns of helical molecules linked head to tail running along [010] direction. PMID- 10517164 TI - Structure of a paralytic peptide from an insect, Manduca sexta. AB - Paralytic peptide 1 (PP1) from a moth, Manduca sexta, is a 23-residue peptide (Glu-Asn-Phe-Ala-Gly-Gly-Cys-Ala-Thr-Gly-Tyr-Leu-Arg-Thr-Ala-Asp-Gly-Arg -Cys-Lys Pro-Thr-Phe) that was first found to have paralytic activity when injected into M. sexta larvae. Recent studies demonstrated that PP1 also stimulated the spreading and aggregation of a blood cell type called plasmatocytes and inhibited bleeding from wounds. We determined the solution structure of PP1 by two dimensional 1H NMR spectroscopy to begin to understand structural-functional relationships of this peptide. PP1 has an ordered structure, which is composed of a short antiparallel beta-sheet at residues Tyr11-Thr14 and Arg18-Pro21, three beta turns at residues Phe3-Gly6, Ala8-Tyr11 and Thr14-Gly17, and a half turn at the carboxyl-terminus (residues Lys20-Phe23). The well-defined secondary and tertiary structure was stabilized by hydrogen bonding and side-chain hydrophobic interactions. In comparison with two related insect peptides, whose structures have been solved recently, the amino-terminal region of PP1 is substantially more ordered. The short antiparallel beta-sheet of PP1 has a folding pattern similar to the carboxyl-terminal subdomain of epidermal growth factor (EGF). Therefore, PP1 may interact with EGF receptor-like molecules to trigger its different biological activities. PMID- 10517166 TI - Identification of families with hereditary breast and ovarian cancer for clinical and mammographic surveillance: the Modena Study Group proposal. AB - Hereditary factors play a fundamental role in the pathogenesis of breast cancer (BC). Approximately 15-20% of all BCs have been reported to show familial clustering. In spite of the recent demonstration and chromosomal localization of BC predisposing genes, clinical clues and careful inspection of pedigree still remain major instruments in HBC diagnosis. The aim of the present study was to develop minimum operational criteria for the selection of family groups at high risk of developing BC. Following a stepwise procedure, families were stratified into four clusters with increasing probability of genetic involvement. So far, 317 BC-prone families have been identified and distributed in the four groups, and 151 high risk women underwent our clinical and mammographic surveillance program. Among these, after a mean follow-up of 24 months, six BCs and one OC were diagnosed (one BC and one OC occurred in the same woman) and one 'interval' BC was observed. Since the prevalence rate so far detected is dramatically higher than that seen at the first round of Italian population-screening programs, our preliminary data support the usefulness of the proposed procedure in selecting high risk individuals. PMID- 10517165 TI - Treatment of advanced breast cancer with gemcitabine and vinorelbine plus human granulocyte colony-stimulating factor. AB - PURPOSE: A phase II trial was performed to investigate the efficacy and tolerance of gemcitabine, vinorelbine, and recombinant human granulocyte colony-stimulating factor (G-CSF) in advanced breast cancer. PATIENTS AND METHODS: Between April 96 and August 97, 60 patients entered this trial. Forty-five patients were previously untreated and 15 patients had failed previous palliative chemotherapy with (n = 10) or without anthracyclines (n = 5). Therapy consisted of gemcitabine 1000 mg/m2 on days 1 + 15 + 21 and vinorelbine 40 mg/m2 on days 1 + 21, both diluted in 250 ml saline and infused over 30 min. G-CSF was administered at 5 microg/kg/day subcutaneously from days 2-6 and 22-26. Courses were repeated every 5 weeks. Treatment was continued in case of response or stable disease until a total of six courses. RESULTS: The overall response rate was 55.5% for patients who had not received prior palliative chemotherapy (95% confidence interval, 40% 70.3%), including 5 CR (11.1%) and 20 PR (44.4%); 12 patients (27%) had stable disease (SD), and 8 (18%) progressed. Second-line treatment with this regimen resulted in 6/15 (40%) objective remissions, 5 had SD, and 4 PD. The median time to progression was 9.5 months (range, 1.5-28) in previously untreated patients, and 7.0 months (range, 2-23) in those who had failed prior chemotherapy. After a median follow-up time of 15 months, 44 patients (73%) are still alive with metastatic disease. Myelosuppression was commonly observed, though WHO 3 and 4 neutropenia occured in only 9 (15%) and 2 patients (3%), and was never complicated by septicaemia; grade 3 anemia was noted in 2 patients. Severe (WHO grade 3) nonhematologic toxicity was rarely observed, and included nausea/emesis in 3 and constipation in 2 patients. CONCLUSIONS: Our data suggest that gemcitabine and vinorelbine plus G-CSF is an effective and tolerable first- as well as second-line combination regimen for treatment of advanced breast cancer. PMID- 10517167 TI - Mechanism of captopril toxicity to a human mammary ductal carcinoma cell line in the presence of copper. AB - Captopril (D-3-mercapto-2-methylpropanoyl-L-proline) is an angiotensin converting enzyme (ACE) inhibitor, used widely in the treatment of hypertension and congestive heart failure. Captopril also inhibits proliferation of a variety of cell types, including several lacking ACE and renin acitvity. We have previously demonstrated that human mammary ductal carcinoma cells are among the cell types whose mitotic activity is inhibited by captopril. In those cells, captopril also reduces estrogen receptor (ER) and increases progesterone receptor (PR) concentrations. The present study evaluated the mechanism of captopril's antiproliferative action in an ER/PR-negative human mammary ductal carcinoma cell line, Hs578T. Cells grown in a 10% serum medium showed negligible changes in the presence of captopril alone. However, in the presence of subphysiologic concentrations of copper salts or copper-loaded ceruloplasmin, captopril caused a dose-dependent reduction in cell number, thymidine incorporation and mitochondrial dehydrogenase activity. In contrast, iron salts and iron-saturated transferrin had no effect on captopril activity. Catalase and horseradish peroxidase nullified the cytotoxic effects of captopril/Cu++, whereas H2O2 mimicked those effects. These data are consistent with the notion of a copper catalyzed oxidation of captopril, leading to the generation of H2O2 as the cytotoxin to this clinically important cell type. PMID- 10517168 TI - Induction of insulin-like growth factor binding protein expression by ICI 182,780 in a tamoxifen-resistant human breast cancer cell line. AB - Earlier studies in our laboratory demonstrated that the steroidal antiestrogen ICI 182,780 is very effective in abolishing the tamoxifen-resistant proliferation of MCF 7/5-23 cells. In addition, preliminary binding studies showed that ICI 182,780 increased the binding of insulin-like growth factor (IGF)-I to the MCF 7/5-23 cells, although this finding was not the result of an increase in the expression of the insulin-like growth factor-I receptor (IGF-IR). Hence, we reasoned that the inhibition of tamoxifen-resistant cell growth by ICI 182,780 might have been due to increased expression of insulin-like growth factor binding proteins (IGFBPs). We observed the up-regulation of non-insulin-suppressible IGF I binding in both the tamoxifen-sensitive MCF 7/5-21 cell line (1.5-fold) and the tamoxifen-resistant MCF 7/5-23 cell line (2.5-fold) after 5 days of treatment with ICI 182,780 (10(-7) M) in serum-free medium, suggesting a role for cell associated IGFBPs. Affinity cross-linking experiments confirmed the presence of an IGF-I:IGFBP complex of approximately 38-kDa in tamoxifen or ICI 182,780 treated cells. Western ligand blots showed higher levels of a soluble 30-kDa IGFBP in media conditioned by either of the subclones that had been treated with ICI 182,780, an effect consistently opposed by estrogen (E2: 10(-9) M). RT-PCR showed higher levels of IGFBP-5 mRNA than any of the other known IGFBPs, suggesting that this was the major IGFBP subtype. The protein was subsequently identified by Western immunoblotting as IGFBP-5. In conclusion, we postulate that this may be a mechanism contributing to the greater potency of ICI 182,780 in the growth inhibition of the MCF 7/5-23, tamoxifen-resistant cell line. PMID- 10517169 TI - A study on the cost effectiveness of sestamibi scintimammography for screening women with dense breasts for breast cancer. AB - The potential impact of Sestamibi scintimammography (SSMM) on the cost effective management of women with dense breasts is not known. This study addresses this issue quantitatively by examining the impact of SSMM based screening strategies on the approximately 3,000,000 women over 40 with very dense breasts (DY patterns) without palpable masses and who have had one or more prior mammograms, who undergo routine screening each year. Quantitative decision tree sensitivity analysis was used to compare the conventional mammography (MM) strategy (strategy A), which does not subject patients with negative mammograms to any further examination until their next screening, with two decision strategies for screening with SSMM; SSMM after a negative mammogram (strategy B) or SSMM as the only screening test for women already identified as having dense breasts by a previous mammogram (strategy C). Cost effectiveness was measured by calculating the incremental cost effectiveness ratio (ICER) of strategies B and C, which is the cost of achieving an additional year of life in the screening population by choosing a SSMM based decision strategy rather than the conventional strategy. Strategies B and C reduced the number of false negative diagnoses by 62% and 8%, respectively. The ICER was $632,000 and $3.18M per life year for strategy B and C, respectively. To be cost effective, the pre-test probability of cancer in the study population must be greater than 3% for strategy B or the cost of SSMM must be less than $50 for strategy C. These results show the ICER of an SSMM based breast cancer screening strategy in the management of patients with dense breasts is not currently within the range (approximately $50,000 per year life saved) of other commonly performed medical interventions that are considered cost effective. PMID- 10517170 TI - Breast cancer and timing of surgery during menstrual cycle: a 5-year analysis of 248 premenopausal women. AB - In the present report, we retrospectively analyzed the impact of the timing of surgery during menstrual cycle on disease-free and overall survival of 248 premenopausal patients with stage I/II breast cancer who underwent surgery followed by anthracycline-containing adjuvant chemotherapy. With a median follow up of 5 years, no statistically significant differences were observed in disease free or overall survival between women operated upon during the follicular (days 0-14) and the luteal (days 15-32) phase of the menstrual cycle. The impact on disease-free and overall survival of lymph-node status, tumor size and hormone receptor expression, but not of the phase of the menstrual cycle at the time of surgery, was confirmed by univariate and multivariate analysis. However, when combined with hormone receptor status, the phase of the menstrual cycle at the time of surgery proved useful to better define the prognosis of primary breast cancer patients, with significantly longer disease-free and overall survival for patients operated upon during the follicular phase and with positive hormone receptors. PMID- 10517171 TI - Constitutive overexpression of cyclin D1 in human breast epithelial cells does not prevent G1 arrest induced by deprivation of epidermal growth factor. AB - Non-transformed human breast epithelial cell line MCF10A is dependent on exogenous epidermal growth factor (EGF) for continued growth. Complete G1 arrest was rapidly induced following EGF deprivation. The cell cycle arrest was accompanied by increased levels of p27KIP1, a cyclin-dependent kinase inhibitor, and reduced level of cyclin D1. This was associated with strong inhibition of cyclin-dependent kinase 2 and cyclin D1-associated kinase activities. Introduction of exogenous cyclin D1 into MCF10A (MCF10AD1) cells resulted in an accelerated cell growth rate but did not confer colony-forming capacity. Cell cycle arrest was still achieved in MCF10AD1 cells following EGF deprivation. In the great majority of MCF10AD1 clones, accumulation in G1 phase was accompanied by reduced cyclin D1 and increased p27KIP1 protein levels. In two clones where cyclin D remained unchanged during G1 arrest, it was found that more cyclin D1 protein was bound to p27KIP1. The data demonstrate that ectopic expression of cyclin D1 alone could not transform MCF10A cells nor was it sufficient to prevent G1 arrest induced by EGF deprivation. PMID- 10517172 TI - Early age at menopause and breast cancer: are leaner women more protected? A prospective analysis of the Dutch DOM cohort. AB - To investigate the relationship between age at menopause, body mass index, and breast cancer risk, we used data from a prospective cohort study (DOM cohort) in the Netherlands. Participants in this breast cancer-screening project included 10,591 women living in Utrecht, aged 49-66 years at enrolment. During a median follow-up period of 19 years, women attended screening rounds at which anthropometric measurements were taken and questions were asked about menopausal status, age at menopause, medication use and other risk factors for breast cancer. Cox regression analysis was used to investigate the relationship between age at menopause and subsequent breast cancer risk. Breast cancer incidence decreased with an earlier age at menopause. Women with a menopausal age of 44 years or younger had a 34% lower risk of breast cancer, than women with a menopausal age over 54 years (hazard ratio is 0.66 (95% confidence interval 0.43 0.91)). The annual hazard of breast cancer incidence decreased by 2.6% per year reduction in age at menopause. The protective effect of an early age at menopause was stronger for women with a low body mass index (< or = 27 kg/m2; reduction of 44%) than for women with a high body mass index (>27 kg/m2; reduction of 24%), although this difference was not statistically significant (P for interaction = 0.58). This difference was most pronounced in women who had ever smoked. Adjustment for known breast cancer risk factors did not alter the crude risk estimates significantly. In conclusion, this study provides evidence of the protective effect of lower age at menopause on subsequent breast cancer risk. This protective effect may be even stronger in leaner women. PMID- 10517173 TI - Breast conservation therapy over the past 15-20 years. PMID- 10517174 TI - Thy-1 differentiation protein and monocyte-derived cells during regeneration and aging of human placental villi. AB - PROBLEM: The classification of placental villi was reviewed, and regeneration of villous trees in mature human placentae was examined. METHOD OF STUDY: Expression of Thy-1 by placental fibroblasts and pericytes, and markers of endothelial cells and monocyte-derived cells were studied by immunohistochemistry and image analysis. RESULTS: Villous regeneration consists of: (i) dedifferentiation of mature ramuli into young stem villi producing mesenchymal villi; (ii) differentiation of mesenchymal villi into immature intermediate villi; and (iii) differentiation of immature intermediate villi into transitory intermediate villi, branching into the precursors of mature intermediate and terminal villi. These processes are associated with dedifferentiation and redifferentiation of placental monocyte-derived cells. Significant changes of Thy-1 expression by fibroblasts and pericytes accompany aging and degeneration, as well as regeneration of placental villi. CONCLUSIONS: Villous aging and degeneration in normal mature human placenta is compensated by regeneration of villous trees. Lack of villous regeneration may cause chronic fetal distress, due to the increasing demands of the growing fetus on the remaining terminal villi. PMID- 10517175 TI - Hypocomplementemia correlates with intrauterine growth retardation in systemic lupus erythematosus. AB - PROBLEM: The aim of this study was to elucidate fetomaternal risks in systemic lupus erythematosus (SLE)-complicated pregnancy. METHOD OF STUDY: Pregnancy course, complications, and fetal outcome in 82 pregnancies of 55 patients with SLE were investigated. RESULTS: These 82 pregnancies resulted in 14 fetal losses and 66 live births. Without clinical manifestation of SLE-flare, 4 of 8 patients who had low serum complement activity during the pregnancies delivered small-for date neonates. The rate of the intrauterine growth retardation was significantly higher than that observed in pregnancies with normal complement activity. The frequency of premature deliveries (60%) in patients who received more than 15 mg/day of prednisolone was significantly high when compared with pregnancies maintained by 0-15 mg/day (13.1%). CONCLUSIONS: These data demonstrate the preconceptional and perinatal management necessary in SLE and suggest that the pregnancy with hypocomplementemia, the disease activity, and/or a relatively high maintenance dose of corticosteroid should be carefully managed and monitored. PMID- 10517176 TI - Transforming growth factor-beta isoforms and receptors in endometriotic cysts of the human ovary. AB - PROBLEM: The present study examined the presence and cellular distribution of transforming growth factor-beta1, 2, and 3 isoforms and their type I and II receptors in endometriotic cysts of the ovary, relative to their presence in normal endometrial tissue. METHOD OF STUDY: Thirteen control samples of normal endometrium in the proliferative phase and 11 ovarian endometriotic cysts were examined by immunohistochemistry for transforming growth factor-beta1, 2, and 3 isoforms and their type I and II receptors. RESULTS: Immunoreactivity for all ligands and receptors was detected in both normal endometrium and endometriotic cysts. Isoform-specific differences in immunostaining were not detected. Expression of all ligands and receptors was significantly increased in epithelial cells of endometriotic cysts compared with those of normal endometrium. On the other hand, stromal cells in normal endometrium and endometriotic cysts were only faintly immunostained. Inflammatory cells infiltrating among endometriotic stromal cells contained the highest immunostaining intensity for all ligands and receptors. We identified nearly all inflammatory cells as macrophages using a specific antibody. CONCLUSIONS: These findings suggest that macrophages in endometriotic tissue are a major source of transforming growth factor-beta, which may be an important regulator of cell proliferation in endometriotic cysts through paracrine and autocrine actions. PMID- 10517177 TI - RANTES production by cultured primate endometrial epithelial cells. AB - PROBLEM: RANTES (regulated upon activation, normal T cell expressed and secreted), is a chemokine with monocyte, macrophage, T lymphocyte, and eosinophil attractant and activating activities. This mediator has been detected in the peritoneal fluid of patients with endometriosis and in cultures of stromal cells from human endometrial and endometriotic tissue. To determine if endometrial epithelial cells were also a potential source of this mediator, primate endometrial epithelial cells were cultured in vitro and the constitutive and stimulated production of RANTES in these cultures was measured. METHOD OF STUDY: Uterine tissue was obtained from Macaca nemestrina monkeys and the endometrial epithelial cells were isolated and placed in culture for 24-72 hr. RANTES was measured in cell extracts and culture fluids by enzyme-linked immunosorbent assay (ELISA). RESULTS: Constitute release of RANTES was low, ranging from 28-52 ng/mL but addition of interferon gamma (INF-gamma) or the combination of IFN-gamma and tumor necrosis factor alpha (TNF-alpha) produced a marked increase in RANTES production. The greatest release, which was nearly 500-fold greater than the basal level, was observed at 72 hr with the combined addition of TNF-alpha and INF-gamma. Nearly 90% of the stimulated RANTES was released into culture fluids, while cell associated RANTES was minimal constituting only 11.2% of the total. CONCLUSION: These findings indicate that endometrial epithelial cells can produce and release RANTES. This chemokine may be an important attractant and activator of macrophages, T lymphocytes and/or eosinophils in the uterus during the reproductive cycle or implantation. PMID- 10517178 TI - Effect of neonatal treatment with a GnRH antagonist on development of the cell mediated immune response in marmosets. AB - PROBLEM: We examined the effect of neonatal treatment with a gonadotropin releasing hormone (GnRH) antagonist (antide) on the development of cell-mediated immunity in male marmosets. METHOD OF STUDY: Neonatal marmoset twins were treated with either vehicle or antide, and the proliferative response (PR) of lymphoid tissue to mitogens was assessed during infancy, the peripubertal period, and adulthood. RESULTS: Basal proliferation of peripheral blood mononuclear cells (PBMC) from treated peripubertal twins was elevated above control values, but the PR of the cells to T and B cell mitogens was subnormal. Conversely, PBMC from treated infants exhibited an enhanced PR to some of the mitogens employed. In vitro culturing of thymocytes (control or treated) from the three developmental stages with either antide or a GnRH agonist increased basal proliferation, but decreased the PR to mitogens by 60-80%. CONCLUSION: Neonatal treatment with antide alters development of, but does not permanently impair, cell-mediated immunity in the marmoset. GnRH appears to modulate immune responses throughout development in the primate. PMID- 10517179 TI - Characterization of boar spermadhesins by monoclonal and polyclonal antibodies and their role in binding to oocytes. AB - PROBLEM: The role of Ala-Trp-Asn (AWN) and Ala-Gln-Asn (AQN) families of spermadhesive sperm proteins in fertilization. METHOD OF STUDY: The preparation and characterization of polyclonal antibodies against AWN and AQN spermadhesins and one monoclonal antibody (MAb), designated Bo.5, against AWN spermadhesin. The use of biochemical and immunocytochemical methods for characterization of spermadhesins on the sperm membrane of boar spermatozoa and in the cryostat sections of boar reproductive organs. RESULTS: Polyclonal anti-AWN and anti-AQN antibodies specifically reacted with AWN and AQN proteins, respectively. MAb Bo.5 detected the 17-, 16-, and 14-kDa protein members of AWN subfamily. The monoclonal, as well as the polyclonal, AWN antibodies remarkably decreased the sperm binding to the egg surface in an in vitro sperm zona pellucida binding assay. CONCLUSIONS: Presented results demonstrate that polyclonal antibodies and MAb Bo.5 against spermadhesins specifically recognize the membrane-associated antigens and inhibit the binding of sperm to oocytes. Reduced binding of sperm to oocytes, due to the antibodies, indicates the role of these spermadhesins in sperm-egg primary binding. PMID- 10517181 TI - beta-Integrin-collagen interaction reduces chondrocyte apoptosis. AB - We have observed that the spent culture media in suspended chondrocyte cultures is essential for the survival of the cells, since complete change of the spent media induces severe programmed cell death (apoptosis). Moreover, we showed that extracellular matrix (ECM) molecules in the culture media provide vital chondrocyte-matrix interactions; when media are changed, cells are deprived of matrix molecules and undergo apoptosis. In this paper we report that interaction with collagen, a ubiquitous extracellular matrix molecule, is essential for chondrocyte survival. Such an interaction causes chondrocyte aggregation and reduces the level of chondrocyte apoptosis. Hyaluronan, an abundant ECM molecule, can influence the effects of collagen by preventing chondrocyte aggregation. Degradation of hyaluronan with hyaluronidase results in chondrocyte aggregation, and this reduces the level of chondrocyte apoptosis. Experiments with an antibody to integrin beta1 suggest that the collagen-chondrocyte interactions are mediated through integrin beta1, and these interactions may protect chondrocytes from apoptosis. We hypothesize that hyaluronan binds aggrecan and link protein, forming stable ternary complexes, which interact with the chondrocyte surface, perhaps via CD44, and thus maintains a stable chondrocyte-matrix network. PMID- 10517180 TI - Detection of collagenase-induced damage of collagen by 9A4, a monoclonal C terminal neoepitope antibody. AB - To determine whether the collagen network is compromised by collagenase during acute inflammation, a monoclonal antibody (9A4) was developed with specificity for the C-terminal neoepitope sequence generated by collagenase-cleavage of type II collagen (Gly-Pro-Pro-Gly-Pro-Gln-Gly-COOH). 9A4 was shown to detect the collagen collagenase-cleavage neoepitope with a K = 1.7 x 10(-7) M (type II) and K = 2 x 10(-6) M (type I). It does not recognize uncleaved native or denatured collagen. Articular cartilage from control animals is unstained by 9A4. During acute inflammation elicited in hamsters by intra-articular LPS, positive staining for the 9A4 neoepitope indicated the collagen was damaged. Wheel running exercise was used to apply stress to control cartilage and cartilage from animals with damaged collagen. After 6 months of running, the cartilage from normal animals was unaffected. By contrast, in the group with damaged collagen, the cartilage was fibrillated in all animals and in half of those, the cartilage failed and bony eburnation resulted. PMID- 10517182 TI - Ultrastructure of lung elastin and collagen in mouse models of spontaneous emphysema. AB - The tight-skin (Tsk) and beige (bg) mutants of the C57B1/6J strain of mouse spontaneously develop air-space enlargement reminiscent of human emphysema. To determine if this enlargement is accompanied by matrix destruction, as in the human disease, we examined the elastin and collagen matrices of the lungs of both mutants. The ultrastructure of these matrix components was separately visualized by scanning electron microscopy following controlled alkali digestion, which preserves collagen, and formic acid digestion, which enables visualization of elastin. Significant elastin destruction suggestive of an elastolytic process was observed in the lungs of Tsk mice. Thickening of elastin lamellae was observed in the lungs of bg mice, suggesting that congenital matrix remodeling may underlie air-space enlargement in this strain. PMID- 10517183 TI - Evidence of in situ stability of the type IV collagen triple helix in human inflammatory bowel disease using a denaturation specific epitope antibody. AB - A peptide specific antibody (AH1OW1) was raised against an epitope, AH10 (aa 449 463), of the alpha1(IV) chain adjacent to a cleavage site for matrix metalloproteinases (MMP)-2 and -9 within the triple helix of type IV collagen. The antibody only reacted with denatured and reduced preparations of type IV collagen, or with pepsin isolated type IV collagen digested with MMP-2 and MMP-9. The specificity of this antibody for the denatured triple helix was demonstrated by the lack of staining with pre-immune antibody and by pre-incubation of AH1OW1 antibody with excess AH10 peptide epitope. The AH1OWI antibody was used to detect whether proteolysis of type IV collagen occurs in ulcerative colitis, an inflammatory bowel condition often characterised by a large influx of granulocytes and macrophages and an associated tissue destruction. However, no evidence of in situ proteolysis of the basement membrane type IV collagen was observed. Only in the most actively inflamed mucosa was staining with AH1OW1 antibody observed in the mucosal connective tissue. Digestion of frozen sections of bowel with MMP-1, MMP-2, MMP-3 and MMP-9 did not result in the exposure of the AH10 epitope. These data demonstrate the stability of intact type IV collagen and indicate that susceptibility of alpha1(IV) chain to digestion with MMP-2 and MMP 9 may require other proteolytic/denaturing events in the molecule. PMID- 10517184 TI - Collagen breakdown in soft connective tissue explants is associated with the level of active gelatinase A (MMP-2) but not with collagenase. AB - Recent data suggest that gelatinase A (matrix metalloproteinase-2, MMP-2) plays an important role in the degradation of collagen of soft connective tissues. In an attempt to investigate its participation in more detail we assessed the digestion of collagen in cultured rabbit periosteal explants and compared this with the level of active MMP-2 and collagenases. The data demonstrated that both collagen degradation and MMP activity increased with time. Conditioned medium obtained from explants cultured for 72 h showed that the level of active MMP-2 correlated with collagen degradation (r = 0.80, d.f. = 23, P < 0.0001). Such a relationship was not found with collagenase activity (r = -0.08, d.f. = 21, NS). The possible involvement of MMP-2 in collagen degradation was investigated further by incubating explants with selective gelatinase inhibitors (CT1166, CT1399 and CT1746). In the presence of these compounds breakdown of collagen was almost completely abolished (approximately 80%). Finally we assessed whether periosteal fibroblasts had the capacity to degrade collagen type I that conferred resistance to collagenase activity. Breakdown of this collagen did not differ from degradation of normal collagen. Taken together, our data provide support for the view that MMP-2 plays a crucial role in collagen degradation of soft connective tissue. PMID- 10517185 TI - Modulation of keratan sulfate synthesis on lumican by the action of cytokines on human articular chondrocytes. AB - Adult human articular chondrocytes were used to investigate why keratan sulfate/polylactosamine chains are deficient on the lumican residing in the matrix of adult articular cartilage, whereas they are present on the lumican residing in the matrix of juvenile cartilage. Under serum-free conditions with either monolayer cultures, agarose cultures, or micromass cultures, the adult chondrocytes synthesized a form of lumican possessing keratan sulfate/polylactosamine chains. Thus, the adult chondrocytes are capable of producing a proteoglycan form of lumican and this appears to be the default synthesis preference. The micromass culture system proved useful for demonstrating that growth factors/cytokines present in the extracellular milieu are capable of influencing the structure of the keratan sulfate/polylactosamine chains on the secreted lumican. Of particular note was the ability of IL-1beta to promote the secretion of a form of lumican deficient in keratan sulfate/polylactosamine chains, whereas with bFGF, IGF-1 and TGFbeta keratan sulfate/polylactosamine chains were present, though their size or degree of substitution varied. Thus, growth factors/cytokines are able to modulate the molecular form of lumican. Furthermore, additional studies showed that this modulation was not due to the degradation of keratan sulfate/polylactosamine chains following proteoglycan secretion, but represented a direct effect on synthesis. PMID- 10517186 TI - Cyclic compression of articular cartilage explants is associated with progressive consolidation and altered expression pattern of extracellular matrix proteins. AB - In this study, we investigated the biosynthetic response of full thickness, adult bovine articular cartilage explants to 45 h of static and cyclic unconfined compression. The cyclic compression of articular cartilage resulted in a progressive consolidation of the cartilage matrix. The oscillatory loading increased protein synthesis ([35S]methionine incorporation) by as much as 50% above free swelling control values, but had an inhibitory influence on proteoglycan synthesis ([35SO4] incorporation). As expected, static compression was associated with a dose-dependent decrease in biosynthetic activity. ECM oligomeric proteins which were most affected by mechanical loading were fibronectin and cartilage oligomeric matrix protein (COMP). Static compression at all amplitudes caused a significant increase in fibronectin synthesis over free swelling control levels. Cyclic compression of articular cartilage at 0.1 Hz and higher was consistently associated with a dramatic increase in the synthesis of COMP as well as fibronectin. The biosynthetic activity of chondrocytes appears to be sensitive to both the frequency and amplitude of the applied load. The results of this study support the hypothesis that cartilage tissue can remodel its extracellular matrix in response to alterations in functional demand. PMID- 10517187 TI - Collagenase-3 (MMP-13) and its activators in rheumatoid arthritis: localization in the pannus-hard tissue junction and inhibition by alendronate. AB - The hypothesis of the present work was that the pannus tissue overlying the articular hard tissues has an aggressive phenotype and contains the newly discovered collagenase-3 and its endogenous inducers and activators. We therefore analyzed the eventual presence of collagenase-3 and its regulation at the pannus cartilage junction. Collagenase-3 mRNA (in situ hybridization) and enzyme protein (ABC and immunofluorescence staining) were found in the pannocytes in the pannus hard tissue junction. Inflammatory round cells associated with the critical interface contained TNF-alpha and IL-1beta. These cytokines induced collagenase-3 secretion in cultured rheumatoid synovial fibroblasts. Procollagenase-3 activators, stromelysin-1, 72 kDa type IV collagenase/gelatinase and membrane type 1-MMP, were also found in the pannus-hard tissue junction. Active collagenase-3 was inhibited with alendronate (IC50 = 500-750 microM). Collagenase 3, due to its substrate profile and local synthesis in a milieu favoring its activation, might play a major role in the degradation of cartilage type II and bone type I collagens. Alendronate, at concentrations attainable in vivo, is able to inhibit collagenase-3. This might offer an option to control collagenase-3 mediated tissue destruction in rheumatoid arthritis. PMID- 10517188 TI - Is antibiotic therapy of mice and humans useful in Escherichia coli O157:H7 enteritis? PMID- 10517189 TI - Ciprofloxacin and rifampicin versus doxycycline and rifampicin in the treatment of brucellosis. AB - The present study was undertaken to evaluate the efficacy, safety, and patient tolerability of two antibiotic regimens for the treatment of brucellosis: rifampicin 600 mg/day and doxycycline 200 mg/day for 45 days (group 1), versus rifampicin 600 mg/day and ciprofloxacin 1 g/day for 30 days (group 2). Forty patients were diagnosed with brucellosis based on clinical and microbiological findings. The two groups were comparable regarding age and sex distribution. The average number of days without fever and symptoms was lower in group 2 patients than in group 1 patients (mean+/-SD: 3.85+/-1.98 for group 1 vs. 2.78+/-1.03 for group 2, P=0.044). During the 1-year follow-up period, three (15%) patients in group 2 and two (10%) patients in group 1 had clinical relapses; these rates were not significantly different. Ciprofloxacin and rifampicin treatment for brucellosis is as effective as the standard regimen of doxycycline and rifampicin and offers the advantage of a shorter duration of treatment. PMID- 10517190 TI - Prognostic factors influencing mortality in cancer patients with neutropenia and bacteremia. AB - The purpose of this study was to identify risk factors for mortality in neutropenic patients with cancer and bacteremia. A consecutive sample of 438 neutropenic patients (granulocyte count <0.5 x 10(9)/l) with cancer and bacteremia was studied to identify the clinical characteristics associated with mortality at the onset of bacteremia. The mean age of the subjects was 48 years (range, 15-87 years). Most cases of bacteremia (77%) were hospital-acquired and occurred in patients with acute leukemia (48%). Gram-positive organisms caused 233 (53%) episodes of bacteremia, gram-negative organisms caused 151 (34%) episodes, and 48 (11%) episodes were polymicrobial. The overall mortality within 30 days of the onset of bacteremia was 24.4%. The variables found to be independently associated with increased mortality using logistic regression techniques were as follows: shock at the onset of bacteremia (OR, 10; 95% CI, 4.2 23.8), pneumonia (OR,4.4; 95% CI, 1.9-10), uncontrolled cancer (OR,4.3; 95% CI, 1.5-12.7), and absence of prophylaxis with norfloxacin (OR,2.4; 95% CI, 1.3-4.5). The prognostic factors ascertained in this study may help to identify those patients at higher risk of death. Medical intervention addressing some of these factors may improve the outcome of bacteremia in neutropenic patients with cancer. PMID- 10517191 TI - Low prevalence of the immunoglobulin-A-binding beta antigen of the C protein among Streptococcus agalactiae isolates causing neonatal sepsis. AB - Streptococcus agalactiae (Group B streptococcus, GBS) is the most important pathogen causing neonatal sepsis. The role of bacterial proteins contributing to pathogenicity in GBS infections has not yet been clearly determined, but the C protein complex has been suggested to be involved in both virulence and protective immunity. The aim of this study was to assess the prevalence of GBS strains bearing the gene encoding for the beta antigen of the C protein among clinical isolates from 68 neonates with sepsis, 45 newborns colonized without clinical signs of infection, and 50 isolates from pregnant women. The prevalence of the beta antigen gene in all three groups was low (24% vs. 19% vs. 22%) [corrected], and the differences between groups were not statistically significant. Clinical characteristics and cytokine plasma levels did not differ between septic patients with beta antigen-positive and -negative strains. The beta-antigen gene was not found among serotype III isolates, which accounted for roughly half of all the strains isolated. Thus, polymerase chain reaction (PCR) analysis based on the beta antigen gene seems not helpful for distinguishing invasive from colonizing GBS strains. A vaccine based on peptide antigens from the beta antigen of the C protein would most probably not provide protection against the majority of GBS isolates. When analyzing the PCR products of the C protein beta antigen gene by DNA sequencing, a genetic heterogeneity was observed, revealing small repetitive genetic elements within the amplified fragment, an observation that should be studied further. PMID- 10517192 TI - Evaluation of fifteen commercially available serological tests for diagnosis of Lyme borreliosis. AB - The performance of 11 commercially available enzyme immunoassays (EIA) and four Western blot (WB) tests for the detection of IgM and IgG antibodies against Borrelia burgdorferi were compared. A total of 229 serum specimens were used: 26 from patients with early Lyme borreliosis, 13 from patients with late Lyme borreliosis, 62 from healthy controls and 128 from patients with disorders clinically mimicking Lyme borreliosis and/or known to cause cross-reactivity in Lyme borreliosis serological tests (patient control group). In specimens from patients with early Lyme borreliosis, the sensitivity of the individual tests ranged from 35 to 81% for detection of IgM. In late Lyme borreliosis, sensitivity of the tests ranged from 46 to 92%. In healthy controls the specificity of the tests ranged from 89 to 100% and from 82 to 97% for IgM and IgG tests, respectively. In the patient control group, specificity of the tests ranged from 75 to 90% for IgM and from 84 to 100% for IgG tests. The Behring (Germany) and Genzyme Virotech (Germany) IgM EIA tests showed the best performance in detecting early Lyme borreliosis. For the detection of late Lyme borreliosis, the Dako (Denmark) IgG test was the best despite its low sensitivity. The maximum sensitivity of Western blotting for detecting IgM in patients with early Lyme borreliosis and IgG in patients with late Lyme borreliosis was 50 and 46%, respectively. The use of an EIA-WB two-test protocol improved the specificity and positive predictive values of the EIA results but caused a significant loss in sensitivity. Patients with Epstein-Barr virus or cytomegalovirus infection who had a positive reaction in the IgM EIA could not be discriminated from patients with early Lyme borreliosis with the help of Western blotting. Hence, positive and negative predictive values in combination with sensitivity and specificity values indicated that the exclusion of these infections was more relevant than the confirmation of a positive IgM EIA with Western blot. PMID- 10517193 TI - Efficacy of antibiotic therapy for infection with Shiga-like toxin-producing Escherichia coli O157:H7 in mice with protein-calorie malnutrition. AB - Antibiotic therapy for infection with Shiga-toxin-producing Escherichia coli O157:H7 is controversial because of the possibility of its inducing hemolytic uremic syndrome and acute encephalopathy. In a previous study, mice with protein calorie malnutrition were found to be highly susceptible to this pathogen. The efficacy of oral antibiotic therapy in malnourished mice infected with O157 organisms was assessed. Mice fed a low-protein calorie diet were infected intragastrically with 2 x 10(6) colony-forming units of a Shiga-toxin-producing strain of Escherichia coli O157:H7. Infected mice were orally given a therapeutic dose of an antibiotic, including norfloxacin, fosfomycin, kanamycin, ampicillin, clarithromycin or trimethoprim-sulfamethoxazole for 3 days: mice on protocol A received the antibiotic on days 1-3, starting on the day after infection, and mice on protocol B received the antibiotic on days 3-5. The duration of fecal pathogen excretion was shorter and the toxin level in the stool and blood lower in the mice that received protocol A than in untreated mice; all of the mice treated on protocol A survived the lethal infection. All antibiotics except trimethoprim-sulfamethoxazole, administered on protocol B, exhibited the same effect as that exhibited by the respective antibiotic administered on protocol A. Only the mice treated with protocol B of trimethoprim-sulfamethoxazole had a higher toxin level in the blood than untreated controls, resulting in 95% mortality. These results suggest that the antibiotics used in this study, except for trimethoprim-sulfamethoxazole, could reduce the risk of hemolytic uremic syndrome and acute encephalopathy following Escherichia coli O157:H7 infection in humans, and that fosfomycin, in particular, may be relevant for testing in humans. PMID- 10517194 TI - Group A streptococcal meningitis in the antibiotic era. AB - A case of group A streptococcal meningitis is reported and the 51 cases reported in the literature since 1966 reviewed. A total of 24 men and 24 women were included in the study; the mean age (+/-SD) was 20.9+/-25.5 years. Fifty-eight percent of the patients had comorbid conditions, 80% had a distant focus of infection, and 65.8% had blood cultures positive for group A streptococci. Seventy-five per cent of the patients were treated with penicillin. The overall case-fatality rate was 12% (6 patients). Sequelae were more prevalent among children (44%) than among adults (7.7%) (OR=9.43; 95% CI, 1.02-438.95; P=0.03). Group A streptococcus is a rare cause of pyogenic meningitis, affecting mainly children or adults with comorbidity. Although the case-fatality rate is relatively low, neurological sequelae are frequent among survivors, especially children. PMID- 10517195 TI - Mycobacterium malmoense infections in immunocompetent patients. AB - While Mycobacterium malmoense infections were originally restricted to northern Europe, there has been an increasing number of reports of cases of infection in other countries. Two cases of infections due to Mycobacterium malmoense in immunocompetent patients in Germany are presented. In both cases a presumptive diagnosis of tuberculosis was established initially. Mycobacterium malmoense was cultured after a long incubation period (6-8 weeks). The patients were successfully treated with a triple regimen consisting of rifampicin, ethambutol and clarithromycin. The epidemiology and difficulties in diagnosis of Mycobacterium malmoense infection are discussed. PMID- 10517196 TI - Mycobacterium kansasii infection in patients infected with the human immunodeficiency virus. AB - To investigate the clinical and radiographic features and the response to therapy of Mycobacterium kansasii infection in human immunodeficiency virus-infected patients, the clinical charts of 19 cases diagnosed during a 15-year period were reviewed retrospectively. Most patients were male intravenous drug abusers. Mycobacterium kansasii infection occurred late in the course of HIV disease and was associated with advanced immunosuppression. Thirteen patients had pulmonary disease, three extrapulmonary disease (2 with pulmonary involvement), and three pulmonary colonization. Most of them had fever and nonspecific respiratory symptoms; interstitial and alveolar infiltrates were the most common radiographic findings. Fourteen patients were given antituberculous treatment; among these, a clinical response was observed in 85%. Overall mortality was 63%, but only four patients died from active Mycobacterium kansasii disease. HIV infection has become the most important risk factor for Mycobacterium kansasii disease in our setting. Pulmonary infection is the most frequent form of disease and is usually responsive to antituberculous therapy. PMID- 10517197 TI - Disseminated infection due to Nocardia transvalensis coincident with Cryptococcus neoformans variety gattii meningitis. AB - A case of meningitis due to Cryptococcus neoformans var. gattii coincident with disseminated Nocardia transvalensis infection is reported. Nocardia infection initially progressed despite high-dose antimicrobial therapy. Although a specific immunologic defect could not be defined, in vitro lymphocyte proliferation in response to stimulation with the Nocardia isolate was reduced. It is proposed that coinfection with Cryptococcus neoformans may have contributed to the observed impairment of lymphocyte function, leading to disseminated Nocardia disease and a suboptimal treatment response. PMID- 10517198 TI - New lineal immunoenzymatic assay for simultaneous detection of p24 antigen and HIV antibodies. AB - The aim of this study was to evaluate a new lineal immunoenzymatic assay for the simultaneous detection of HIV-1/HIV-2 antibodies and p24 antigen. A total of 320 serum samples were obtained from individuals infected by HIV (HIV 1, n = 183; HIV 2, n = 2), individuals with risk factors for HIV infection (n =49), recipients of multiple transfusions (n =40), and blood donors (n = 46). The Western blot for detection of HIV antibodies and an enzyme immunoassay for detection of p24 antigen, both established techniques, were used for direct comparison. In cases of recent infection, p24 antigen was generally detected at the same time by the lineal test and the established assay. The p24 antigen sensitivity was about 200 pg of HIV antigen per milliliter. The results seem to indicate that the new test could be used with sufficient reliability for screening biological samples (sensitivity, 99.5%; specificity, 94.8%). Use of the lineal immunoenzymatic assay may shorten the amount of time needed to diagnose acute infection with HIV to approximately 2 weeks. PMID- 10517199 TI - Molecular and antibiogram relationships of Acinetobacter isolates from two contrasting hospitals in the United Kingdom and South Africa. AB - The aim of this study was to compare the molecular relationships and antibiograms of nosocomial isolates of Acinetobacter spp. from two acute-care hospitals in Nottingham, UK, and Soweto, South Africa, with different hospital infection control problems and procedures. In contrast to Nottingham, where randomly amplified polymorphic DNA fingerprinting demonstrated that a single multiresistant strain of Acinetobacter baumannii has predominated in the hospital intensive care unit over an 11-year period, the Soweto isolates formed a heterogeneous group of unrelated molecular clusters of different antibiograms, with numerous different strains of Acinetobacter baumannii, Acinetobacter sp. 3 and Acinetobacter sp. 13TU apparently being endemic throughout the Soweto hospital. The contrasting results illustrate the need to maintain exemplary infection control procedures in hospitals where high standards have been achieved and warn of what might result if such measures are diminished. PMID- 10517200 TI - Acute calcular cholecystitis in a patient with brucellosis. PMID- 10517201 TI - Trochanter osteomyelitis and ipsilateral arthritis due to Gemella morbillorum. PMID- 10517202 TI - First isolation of Desulfovibrio species as part of a polymicrobial infection from a brain abscess. PMID- 10517203 TI - Association between submaximal suppression of HIV replication and use of drug combinations without protease inhibitors. PMID- 10517204 TI - Cytomegalovirus polyradiculopathy presenting as bilateral radial nerve palsies in a patient with AIDS. PMID- 10517205 TI - Use of an intraaortic balloon pump in patients with impaired left ventricular function. AB - Prophylactic use of an intraaortic balloon pump (IABP) prior to open-heart surgery in patients with impaired left ventricular function is still under debate. Patients with left ventricular ejection fraction (LVEF) < 40% were therefore compared according to time of IABP placement, viz. preoperative (n = 56), intraoperative (n = 40) or postoperative (n = 17), and also with patients who did not receive mechanical support despite LVEF < 40% (n = 78). The main indication for preoperative IABP insertion was severely impaired left ventricular function (80%), while patients with intraoperative or postoperative IABP placement mainly presented with low cardiac-output syndrome (70%/53%). Preoperative IABP was associated with a low mortality rate (8.9%), whereas patients with intraoperative or postoperative IABP placement had a high mortality risk and an increased catecholamine requirement. Of the patients scheduled for surgery without prophylactic IABP, 19% required intra- or postoperative insertion. Prophylactic placement of IABP thus reduced the mortality rate as well as the postoperative need for mechanical and catecholamine support. Need for intraoperative IABP insertion was associated with high mortality, whereas the outcome after postoperative IABP placement depended on the indication for the measure. PMID- 10517206 TI - Velocity distribution in the ascending aorta in pigs during chronotropic and inotropic stimulation. AB - The influence of heart rate, stroke volume and myocardial contractility on temporal and spatial velocity distribution in the ascending aorta was investigated in 10 pigs. A pulsed Doppler ultrasound technique with intraluminal probe and a single crystal connected to a position-sensitive device was used to measure blood velocity. After baseline registration, the heart rate was increased in two discrete steps of 20 beats/min by right atrial pacing. Isoproterenol infusion was given to increase contractility. Finally, without isoproterenol, the heart rate was again raised to the values found during inotropic stimulation. The first three measuring situations did not differ haemodynamically, apart from increased heart rate and reduced stroke volume. Increased heart rates were not associated with significant change in the parameters for skewness of velocity distribution (peak systolic slope and ratio, maximum skewness slope and ratio). During inotropic stimulation the peak left ventricular dP/dt, aortic systolic pressure, cardiac output and stroke volume were greater than at comparable paced heart rate, and the peak systolic slope of velocity distribution was significantly increased. Velocity distribution in the ascending aorta thus was not altered by increased heart rate alone, whereas skewness of distribution was enhanced by increased inotropic drive of the myocardium and the concomitant central and peripheral vascular changes. PMID- 10517207 TI - Development of velocity profiles and retrograde flow in the porcine abdominal aorta under different haemodynamic conditions. AB - Low and/or oscillating wall shear stresses are related to the development of atherosclerosis and this oscillation is influenced by changes in basic haemodynamics (exercise). The objective of this study was to provide in vivo data on the development of velocity profiles and oscillating blood velocities in the abdominal aorta under varying haemodynamic conditions. Six anaesthetized, 90-kg pigs were used in the study. Abdominal aortic velocity profiles across the anterior-posterior diameter were acquired at different axial positions using 10 MHz pulsed Doppler ultrasound. Measurements were obtained under normal conditions and during cardiac pacing up to 170 beats/min. Velocity profiles were obtained during heart rates ranging between 58 and 169 beats/min, and during flow rates ranging between 0.57 and 2.89 l/min. Main outcome measures included minimum velocities, frequency index, shape of velocity profiles (velocity distribution index), Reynolds' numbers, and Womersley's frequency parameter. Velocity profiles were blunted, with lowest velocities at the distal posterior vessel wall. Multiple regression analysis showed the development of velocity profiles to be inversely correlated with the pulsatility index, Womersley's frequency parameter and the mean Reynolds' number (r = 0.89, p < 0.0005). Minimum velocities were negatively correlated with the PI, Womersley's frequency parameter and positively with the mean Reynolds' number (r = 0.94, p < 10(-8)). Retrograde velocities (and hence oscillating wall shear stresses) were present at mean Reynolds' number < 1000. The oscillation of blood velocities at the wall in the porcine abdominal aorta was highly dependent on general haemodynamics (i.e. flow, heart rate and vessel diameter as expressed in the Reynolds' numbers and Womersley's frequency parameters). The velocity profiles in the abdominal aorta were found to be far from parabolic. These findings have important implications for the understanding and future modelling of the complex haemodynamics in the abdominal aorta and their relation to the development of atherosclerotic disease. PMID- 10517208 TI - Pulmonary blood flow distribution in lobar hypoxia--influence of cardiac output and nitric oxide inhalation. AB - Inhaled NO is reported to be less effective in patients with ARDS if cardiac output is high (> 10 L/min). It has also been demonstrated that increased blood flow and increased shear stress cause an enhancement of endogenous NO production. In one-lung ventilation and regional hypoxia, nitric oxide (NO) delivered to the ventilated lung may decrease blood flow to the nonventilated lung and improve arterial oxygenation. So far, however, results have been divergent. The present study was performed with the hypothesis that inhaled NO would be less effective if cardiac output was increased. In the anaesthetized pig, hypoxia (5% O2) was induced in the left lower lobe. NO was delivered consecutively to the hypoxic lobe and to the other, oxygenated parts, of the lungs during continuous measurement of lobar blood flow and total lung blood flow. Bleeding and infusion of dextran caused variation in cardiac output. It was found that lobar hypoxia per se reduced lobar blood flow from 22.9+/-3.1% to 4.7+/-0.9% of cardiac output. An increase (3.2+/-0.3 L x min(-1)) and a decrease (2.2+/-0.2 L x min(-1)) in cardiac output did not alter the relative perfusion of the hypoxic lobe from baseline cardiac output (2.6+/-0.2 L x min(-1)) values. When NO was delivered to the hypoxic lobe, there was a marked increase in relative lobar perfusion to 19.0+/-2.9% during low cardiac output and 16.5+/-2.7% during high cardiac output without any significant difference between the two NO-induced increases of lobar perfusion. The increase in lobar perfusion tended to depend inversely on total pulmonary blood flow when cardiac output had been reduced by bleeding but without reaching statistical significance (r = -0.42, p > 0.05). The decrease in mean pulmonary artery pressure and PaO2 seen during NO inhalation to the hypoxic lobe did not correlate with the level of cardiac output. When NO was delivered to the oxygenated parts of the lungs, no significant effect on relative lobar perfusion or arterial oxygenation was observed, either at raised or at lowered cardiac output. The findings give no further evidence to show that variations in cardiac output alter the effect of NO inhalation. PMID- 10517209 TI - Surgery for lung cancer in the elderly. AB - In order to assess the appropriateness of lung cancer surgery in the elderly and determine optimal subjects and resection procedure, 75 patients operated on in 1976-1996 at age > or =75 years (including 13 > or =80) were followed up. The operations included limited resection (8), lobectomy (47), bilobectomy (10) and pneumonectomy (10) and were judged to be radical in 59 cases (79%). Perioperative mortality was 9% and morbidity 29%, including 21% major complications. Cumulative 5-year survival was 32%, in stages IA-IIB 27-41%, and cancer-related survival 61 79%. Mortality did not differ significantly between resection types, but morbidity did. Nor did mortality, morbidity or survival differ between the age groups 75-79 and > or =80 years. In stage I cancer there was no significant difference in survival or cancer-related survival after lobectomy vs limited resection. We conclude that age, even >80 years, is not incompatible with curative resection. Lobectomy is the treatment of choice, but a less radical resection may be advisable if there is comorbidity. If more extensive resection is performed, the individual surgical risk must be weighed against the potential long-term benefit. PMID- 10517210 TI - DNA flow cytometry in surgically treated lung cancer--prognostic significance. AB - Although DNA aneuploidy and high proliferative activity (S-phase fraction, SPF) of tumour cells, measured by flow cytometry, have proved to be indicators of poor prognosis in most solid tumours, there have been conflicting results in lung cancer studies. During a four-year period we studied the prognostic significance of DNA ploidy and SPF in 99 surgically treated lung cancer patients. Flow cytometric analysis was done from archival, formalin-fixed, paraffin-embedded tumour specimens. DNA index and SPF were determined, using MultiCycle software with sliced nuclear correction to compensate for debris. There were 61 DNA diploid and 38 DNA aneuploid tumours. The median SPF was 10.2%. Neither ploidy nor SPF was associated with previously known prognostic factors. Survival was poorer in patients with aneuploid tumours than in the other patients, but the difference was not statistically significant. DNA ploidy and SPF thus do not seem to be useful prognostic indicators in surgically treated lung cancer. PMID- 10517211 TI - Left ventricular remodelling in post-myocardial infarction patients with left ventricular ejection fraction 40-50% vs 25-39%. Influence of nisoldipine treatment? An echocardiographic substudy from the DEFIANT II study. AB - OBJECTIVE: Left ventricular (LV) remodelling following acute myocardial infarction has generally been studied in patients with LV ejection fraction (EF) < 40%, and it has been shown that this process can be attenuated by ACE inhibitors. Little is known regarding LV remodelling in patients with LVEF > or = 40% or the effects of treatment in this patient cohort. The DEFIANT II study (Doppler Flow and Echocardiography in Functional cardiac insufficiency) included 542 post-infarction patients with LVEF 25-50% without overt heart failure within 13 days following acute myocardial infarction (AMI). They were then randomized to nisoldipine coat-core (CC) or placebo and followed up for 6 months. DESIGN: Two dimensional echoes were obtained after 8 (5-13) days and 6 months following AMI. SETTING: LV end diastolic (ED) and end systolic (ES) volumes (V) were calculated in 503 patients with technically satisfactory paired echoes using the biplabe method of discs in a core laboratory. SUBJECTS: Group A. 217 patients with baseline EF 40-50%, of whom 112 were randomized to nisoldipine and 104 to placebo (one patient was taken off study medication). Group B. 286 patients with EF 25 39%, of whom 145 were randomized to nisoldipine and 141 to placebo. RESULTS: LVEDV was 175 (+/-45) ml in Group A vs 203 (+/-49) ml in Group B (p = 0.001) at baseline and 184 (+/-48) ml vs 213 (+/-56) ml (p = 0.001), respectively, at 6 months. LVESV at baseline was 97 (+/-42) ml in Group A vs 133 (+/-37) ml in Group B (p = 0.001), and 106 (+/-34) ml vs 134 (+/-45) ml (p = 0.001) at 6 months, respectively. The increase of LVESV was 9 (+/-29) ml in Group A vs 2 (+/-35) ml in Group B (p = 0.007). LVEF decreased by 2 (+/-6)% in Group A vs an increase of 3 (+/-6)% in Group B (p = 0.001). Treatment with nisoldipine had no influence on LV volumes in either of the two groups or in the total study group. CONCLUSION: LV dilatation 6 months following AMI in patients with EF 40-50% was similar in end diastole, but more pronounced in end systole vs patients with EF 25-39%. LV remodelling did not change significantly after nisoldipine treatment. PMID- 10517212 TI - Intramural hydatid cyst of descending aorta complicated by false aneurysm. AB - Hydatid disease is caused by the larval stage of Echinococcus granulosus, and the resultant fluid-filled cysts almost invariably affect the liver. Primary involvement of the aortic wall is very rare. We report a case of hydatid disease presenting as a huge cyst invading the wall of the descending aorta and complicated by a false aneurysm. Diagnostic problems and operative management are reviewed. PMID- 10517213 TI - Subarachnoid haemorrhage with transient myocardial injury and normal coronary arteries. AB - We present a case of cerebral subarachnoid haemorrhage, with T-wave inversions and myocardial akinesia on echocardiography and ventriculography. Acute coronary angiography showed normal arteries. An aneurysm of the right middle cerebral artery was clamped. Echocardiogram was normalized. We discuss coronary spasm as the possible mechanism of myocardial stunning in subarachnoid haemorrhage. PMID- 10517214 TI - Surgical repair of type B aortic dissection complicated by early postoperative lung vein and artery thrombosis. AB - A 24-year old man with Marfan syndrome previously operated for abdominal aortic aneurysm and type A dissection sustained a type B dissection. He underwent graft replacement of the descending and upper abdominal aorta, complicated by infarction of the left upper lobe and lobectomy was carried out. The postoperative course was uneventful. The mechanism for this rare complication is discussed. PMID- 10517215 TI - Severe nodal arrhythmia following direct current cardioversion for atrial flutter. AB - A case of long-lasting nodal arrhythmia and severe hypotension following DC cardioversion for atrial flutter is presented. The patient, treated with the selective serotonin reuptake inhibitor sertraline and with sotalol, thiopental and digoxin, showed no sign of organic disease or drug intoxication. We suggest that drug interaction in combination with the DC shock and an altered sympaticus/parasympaticus balance during anaesthesia provoked the incident. PMID- 10517216 TI - A molecular and clinical study of heterozygous familial hyplercholesterolemia in the Finnish North Karelia. PMID- 10517217 TI - Determination of 4-hydroxyifosfamide concomitantly with ifosfamide and its dechloroethylated metabolites using gas chromatography and a nitrogen phosphorus selective detector. AB - A sensitive gas chromatographic (GC)/nitrogen phosphorus detection (NPD) system was developed for the determination of the antitumor drug ifosfamide (Ifos) and its 2-dechloroethylifosfamide (2-Difos), 3-dechloroethylifosfamide (3-Difos) and 4-hydroxyifosfamide (4-OHIfos) metabolites in human blood. 4-OHIfos was analyzed after coupling with a trapping agent and was used as an indicator of isophosphoramide mustard (IPM). Ifos and its metabolites 2-DIfos, 3-DIfos, 4 OHIfos and the internal standard (trofosfamide) were extracted into chloroform and then resolved by gas chromatography using a Hewlett Packard HP5 capillary column cross-linked with 5% phenyl methyl silicone (30 m; 530 microm I.D.; 2.65 microm film thickness). Precision and accuracy of the assay were determined over a three-day period and a concentration range of 3.25-50 microg/ml for Ifos, 0.8 14 microg/ml for 2D-Ifos, 0.6-10 microg/ml for 3D-Ifos and 0.08-1.40 microg/ml for 4-OHIfos. The limit of quantitation was set at 3.25, 0.80, 0.62 and 0.08 microg/ml, respectively, for Ifos, 2-DIfos, 3-DIfos and 4-OHIfos. The intra- and inter-day coefficients of variation and accuracies were less than 20%, except for a low concentration 4-OHIfos. This assay was then used to provide pharmacokinetic data on antitumor and toxicologic effects following intravenous infusion of Ifos. PMID- 10517218 TI - Precise quantitative determination of phytosterols, stanols, and cholesterol metabolites in human serum by capillary gas-liquid chromatography. AB - Total lipid extraction, solid-phase extraction, saponification, derivatization to trimethylsilyl ether derivatives, then capillary gas-liquid chromatography were used for quantitative analysis of sitosterol, campesterol, stigmasterol, sitostanol, campestanol, lathosterol, desmosterol, and lanosterol in human serum. Details of quality control integral to the accuracy and precision of analyses are included. The method limits of detection and quantitation, respectively, ranged from 0.05 microg/ml and 0.2 microg/ml for sitostanol to 0.4 microg/ml and 1.2 microg/ml for campesterol and campestanol. Analytes were measured at concentrations of 120 ng/ml to 6 microg/ml with standard deviations of 0.02 to 0.12 microg/ml for 55 analyses of a control serum sample conducted over a 2-month period. PMID- 10517219 TI - Simultaneous determination of enantioselective plasma protein binding of aminohydantoins by ultrafiltration and chiral high-performance liquid chromatography. AB - Chiral HPLC methods were developed and utilized for the simultaneous determination of plasma protein binding of enantiomers of two racemic aminohydantoin compounds. Reversed-phase HPLC with the use of a polysaccharide type chiral stationary phase column was employed for the separation and quantitation of the enantiomers of the two compounds with detection limits in the range 5-10 ng/ml in the plasma matrix. The chiral HPLC methods were selective, sensitive and reproducible. The R and S enantiomers of both compounds were baseline-resolved under the chromatographic conditions employed. Ultrafiltration techniques were applied to determining the plasma protein binding for each enantiomer in rat, dog and human plasma. The results clearly show stereoselective binding of the two enantiomers of each compound with higher protein binding of the R enantiomer than the S enantiomer in rat, dog and human plasma. Binding association constants were also determined to be in the range 1.01-14.0 x 10(4) M(-1) at 37 degrees C. Both the protein binding percentage and binding association constant were enantioselective and species-dependent. Such information is important for a clear understanding of the differences in biological activity as well as in pharmacokinetic and pharmacodynamic properties between the two enantiomers of each compound in the drug discovery and development process. PMID- 10517220 TI - Efficient method for preparation of highly purified lipopolysaccharides by hydrophobic interaction chromatography. AB - A method for the efficient preparation of highly purified lipopolysaccharides (LPSs) by hydrophobic interaction chromatography (HIC) has been developed. The procedure can be used for the purification of cell wall bound LPSs after hot phenol-water extraction and for the isolation of extracellular LPSs from the supernatant, respectively. The method described has been tested with artificial mixtures containing LPSs, polysaccharide, protein and RNA and subsequently employed for the preparative purification of two LPSs of different origin, namely the extracellular LPS secreted by Escherichia coli E49 into the culture medium, and the cell wall bound LPS from Pseudomonas aeruginosa VA11465/1. Compared to currently used methods for LPS purification such as enzymatic digestion and ultracentrifugation, the chromatographic separation reported here combines superior purity with minimal loss of LPS, high reproducibility and simple handling. The removal of contaminants such as protein, RNA and polysaccharides and the recovery of LPSs were monitored by appropriate assays. PMID- 10517221 TI - Rapid determination of valaciclovir and acyclovir in human biological fluids by high-performance liquid chromatography using isocratic elution. AB - A rapid high-performance liquid chromatographic assay with isocratic elution is developed for the simultaneous quantification of valaciclovir (VACV) prodrug and its active converted compound, acyclovir (ACV), in biological fluids of treated patients. For serum, the samples are deproteinized with perchloric acid in presence of 1-methylguanosine as the internal standard (IS). For urine and dialysis liquid, the samples are diluted with a mobile phase containing the IS, then filtered. VACV, ACV and the IS are separated on a SymmetryShield RP-8 column with acetonitrile-ammonium phosphate buffer as the mobile phase and detected at 254 nm. The chromatographic time is about 12 min. The relative standard deviations (RSD) of VACV and ACV standards are between 0.5 and 3.5%. Most endogenous nucleosides and their metabolites, psychotropic drugs and drugs of abuse are shown not to interfere with this technique. The method has been applied to study the pharmacokinetics of VACV and ACV in serum, dialysis liquid and urine of renal failure patients on continuous ambulatory peritoneal dialysis (CAPD) under oral treatment of VACV. PMID- 10517222 TI - Simultaneous determination of residual tetracyclines in foods by high-performance liquid chromatography with atmospheric pressure chemical ionization tandem mass spectrometry. AB - We established a method for precisely determining residual tetracycline antibiotics (TCs) in foods by atmospheric pressure chemical ionization liquid chromatography-tandem mass spectrometry (APCI LC-MS-MS) using selected reaction monitoring with an internal standard. By setting the nebulizer probe temperature to 475 degrees C, we were able to use a mobile phase containing oxalic acid without clogging problems at the APCI interface, since oxalic acid decomposes to carbon dioxide and water at high temperature. DMCTC was very effective as an internal standard for determining TCs in various foods. TCs were cleaned up using a Bond Elut ENV cartridge and analysed by APCI LC-MS-MS. The recovery of TCs from various foods including animal tissues, honey, milk, eggs, and fish fortified at levels of 0.05, 0.10, and 0.50 ppm averaged 60.1-88.9%, with an RSD of 1.2-8.7%. The detection limits were 0.001 ppm for OTC and TC, 0.004 ppm for CTC, and 0.002 ppm for DC. The present method was also successfully used to determine TCs in swine kidney samples that were previously found by microbiological assay. PMID- 10517223 TI - Rapid, single-step procedure for the identification of transglutaminase-mediated isopeptide crosslinks in amino acid digests. AB - Tissue transglutaminase (tTG) is a calcium-activated enzyme which can covalently crosslink the epsilon-amino group of a peptide-bound lysine into the gamma carboxamide group of a peptide-bound glutamine, forming a epsilon-(-gamma glutamyl)lysine isopeptide bond. We have developed a sensitive, single-step method for the isolation and detection of tTG-mediated isopeptide bonds from purified proteins and tissue homogenates. This method offers significantly improved resolution over current techniques, and obviates the need for multi column systems or costly fluorescence monitors. By using enzymatic proteolysis, derivatization with phenylisothiocyanate, and a simple elution gradient for HPLC, we were able to determine the frequency of crosslinks in purified fibrin (1.7 mol of isodipeptide per mol of fibrin), crosslinked tau proteins (0.75 mol of isodipeptide per mol of tau), and whole-tissue liver homogenates (0.5 nmol of isodipeptide per mg of total protein). PMID- 10517224 TI - Liquid chromatographic-mass spectrometric determination of the novel, recently identified thioTEPA metabolite, thioTEPA-mercapturate, in urine. AB - An assay for the quantitative determination of the mercapturic acid conjugate of N,N',N"-triethylenethiophosphoramide (thioTEPA-mercapturate) in human urine has been developed. ThioTEPA-mercapturate, a recently identified metabolite of the alkylating anticancer agent thioTEPA, was analyzed using LC-MS and with direct sample injection. Sulphadiazine was used as internal standard. Linearity was accomplished in the therapeutic relevant range of 1-25 microg/ml; recovery was 84% and both accuracy and precision were less than 20% for the lower limit of quantification (1.0 microg/ml) and less than 10% for the other concentration levels. The stability of thioTEPA-mercapturate proved to be satisfactory over a period of 2 months, when kept at -80 degrees C. ThioTEPA-mercapturate urine concentrations of two patients treated with thioTEPA are presented demonstrating the applicability of the assay for clinical samples. PMID- 10517225 TI - Discrimination of granulocyte colony-stimulating factor isoforms by high performance capillary electrophoresis. AB - Granulocyte colony-stimulating factor (G-CSF) is a glycoprotein which acts primarily to stimulate the proliferation, differentiation and activation of committed progenitor cells of the neutrophil-granulocyte lineage into functionally mature neutrophils. The traditional biological assays employed to detect G-CSF are a myeloid bone marrow colony assay, a factor-dependent cell line specific for G-CSF and commercially available immunoassays. However, these methods will not distinguish between glycosylated and non-glycosylated forms of the molecule. In this study high-performance capillary electrophoresis (HPCE) was used to analyse glycosylated and non-glycosylated recombinant human granulocyte colony-stimulating factor (r-met-hG-CSF). Glycosylated G-CSF preparations contained human serum albumin (HSA), added as a protein carrier. Glycosylated and non-glycosylated G-CSFs were prepared in 40 mM Na2HPO4 buffer, pH 2.5, containing hydroxypropylmethylcellulose (HPMC) or 50 mM Na2HPO4 buffer, pH 9.0. Glycosylated G-CSF could be separated into two distinct glycoform populations at the lower pH studied. Differences in migration time and peak shape between glycosylated and non-glycosylated G-CSF were demonstrated. HPCE analysis of G-CSF produced using a baculovirus expression vector system revealed a further distinct G-CSF glycoform and demonstrated the resolving power of the technique. PMID- 10517226 TI - Automated column-switching high-performance liquid chromatography method for the determination of 1-hydroxypyrene in human urine. AB - An on-line sample treatment method to determine 1-hydroxypyrene (1-OHP), a metabolite of polycyclic aromatic hydrocarbons (PAHs), in human urine has been developed. The hydrolysed biological fluid was directly injected into the chromatographic system after only centrifugation. A miniature precolumn loop packed with a preparative phase and coupled on-line to a liquid chromatographic (LC) system was used for analyte enrichment. The analytes were non-selectively desorbed with the LC eluent and cleaned by means of a column-switching procedure comprising two purification columns and an analytical column. Pre-treatment and analysis were performed within 2 and 20 min, respectively. Average 1-OHP recovery reached 99% in the 1-25 microg/l range of urine, and the quantitation limit was 20 ng/l for 100 microl of injected sample. A comparison with a more time consuming off-line method was performed by analysing 120 urine samples of PAH exposed and expected unexposed workers; the statistical treatment indicated that both methods are in agreement. PMID- 10517227 TI - Determination of phenolic derivatives of antipyrine in plasma with solid-phase extraction and high-performance liquid chromatography-atmospheric-pressure chemical ionization mass spectrometry. AB - This manuscript describes a method to determine antipyrine and its phenolic derivatives in plasma by reversed-phase high-performance liquid chromatography mass spectrometry (RP-HPLC-MS). The sample pretreatment consisted of a C18 solid phase extraction (SPE), to remove the salts and proteins. The retention behavior of antipyrine and its phenolic derivatives in the SPE procedure was estimated by the k values determined on a C18 HPLC column at different pH values and with different buffer compositions. Recoveries of antipyrine and its phenolic products were 90% in water and 100% in plasma. Atmospheric pressure chemical ionization (APCI) was used to introduce the components into the mass spectrometer. The mass spectrometer was operated in the single ion monitoring mode (SIM mode) as well as in the selective reaction (SR) mode. The SR mode or tandem MS resulted in the best signal-to-noise ratio, with a detection limit for antipyrine of 6 pg in 20 microl. For the different phenolic antipyrines, different target ions were used and conditions were optimized for each. PMID- 10517228 TI - Solid-phase microextraction gas chromatographic-mass spectrometric method for the determination of inhalation anesthetics in urine. AB - Solid-phase microextraction (SPME) has been applied to the headspace sampling of inhalation anesthetics (i.e. nitrous oxide, isoflurane and halothane) in human urine. Analysis was carried out by gas chromatography-mass spectrometry using a capillary column with a divinylbenzene porous polymeric stationary phase. A SPME divinylbenzene-Carboxen-polydimethylsiloxane coated fiber, 2 cm long, was used, and its performances were compared with those of a Carboxen-PDMS in terms of sensitivity, extraction efficiency, extraction time, fiber coating-urine distribution coefficient. For both fibers, linearity was established over four orders of magnitude, limits of detection were below 100 ng/l for nitrous oxide and below 30 ng/l for halogenated. Precision calculated as %RSD was within 3-13% for all intra- and inter-day determinations. The method was applied to the quantitative analysis of anesthetics in the urine of occupationally exposed people (operating room personnel). PMID- 10517229 TI - Gas chromatographic-mass spectrometric analysis of stable isotopes of cysteine and glutathione in biological samples. AB - A gas chromatographic-mass spectrometric (GC-MS) procedure for the determination of stable isotope labelled glutathione has been applied to animal and human samples. The method, based on preparation of the N,S-ethoxycarbonyl methyl ester derivative of the intact peptide, is rapid and requires little or minor tissue treatment. The same method was applied to cysteine. The method was found to be reliable in terms of within-day and between-day precision, accuracy and linearity. The procedure was applied in humans and animals to determine in vivo the glutathione fractional synthesis rate using labelled cysteine infusion. The glutathione fractional synthesis rate was found to be 22.5%/day in blood from a healthy volunteer and 337+/-29%/day in rat liver. PMID- 10517230 TI - Identification of aminoethylcysteine ketimine decarboxylated dimer in human plasma. AB - Aminoethylcysteine ketimine decarboxylated dimer (AECK-DD) is a natural sulfur containing tricyclic compound detected, until now, in human urine and bovine cerebellum. Recently, the antioxidant properties of this compound, and particularly its protective effect on the in vitro oxidation of low-density lipoproteins, have been demonstrated. In this paper, the identification of AECK DD in human plasma by means of gas chromatography, high-performance liquid chromatography and gas chromatography-mass spectrometry, performed after a simple and fast purification procedure, is reported. PMID- 10517231 TI - Simple method for rapid measurement of trichloroethylene and its major metabolites in biological samples. AB - A simple and rapid, yet sensitive technique was developed for concurrent measurement of trichloroethylene (TCE) and its major metabolites (i.e., trichloroacetic acid, trichloroethanol and dichloroacetic acid) in blood and in solid tissues. The method involves addition of an esterizer (water, sulfuric acid, methanol; 6:5:1; v/v/v) to blood or tissue homogenate in sealed vials, and subsequent gas chromatographic headspace analysis. The procedure should be useful in medical monitoring of TCE exposure as well as in experimental work, notably pharmacokinetic and pharmacodynamic studies pertaining to TCE carcinogenesis. PMID- 10517232 TI - Determination of 11-nor-delta9-tetrahydrocannabinol-9-carboxylic acid in urine using high-performance liquid chromatography and electrospray ionization mass spectrometry. AB - High-performance liquid chromatography with electrospray ionization mass spectrometry was used to determine 11-nor-delta9-tetrahydrocannabinol-9 carboxylic acid (THC-COOH) in urine. After basic hydrolysis of conjugates, the compound was extracted using SPEC-PLUS-3ML-C18 solid-phase extraction columns. A deuterium labelled internal standard (d3-THC-COOH) was added prior to hydrolysis. Separation was performed on a reversed-phase Zorbax Eclipse XDB-C8 analytical column (150x3.0 mm I.D.) using a gradient program from 60 to 80% acetonitrile (4 mM formic acid) at a flow-rate of 0.5 ml/min. The compounds were detected by single ion monitoring of m/z 345 and m/z 348 for the protonated molecules [THC COOH+H]+ and [d3-THC-COOH+H]+, respectively. The precision and accuracy were tested on spiked urine samples in the range 2.5-125 ng/ml. The mean recovery was 95% (n = 58), coefficients of variations were 2.2-4.3% and the limit of detection 2 ng/ml. Diagnostic qualifying ions of THC-COOH (m/z 327 and m/z 299) and d3-THC COOH (m/z 330) were generated using up-front collision-induced dissociation. The relative ion intensities in clinical samples (n = 21) were within +/-20% deviation compared with standards. Using this tolerance and the presence of the ions m/z 327 and m/z 299 at the correct retention times as the acceptance criteria for identification of THC-COOH positive samples, the limit of detection was 15 ng/ml. The LC-MS method complies with the current recommendations on drugs of abuse testing, in which mass spectrometric detection is emphasized. PMID- 10517234 TI - Rapid determination of the anti-cancer drug chlorambucil (Leukeran) and its phenyl acetic acid mustard metabolite in human serum and plasma by automated solid-phase extraction and liquid chromatography-tandem mass spectrometry. AB - A bioanalytical method for the determination of the anticancer drug chlorambucil (Leukeran) and its phenyl acetic acid mustard metabolite in human serum and plasma is described. Automated solid-phase extraction of the analytes is carried out with C18 sorbent packed in a 96 well format microtitre plate using a robotic sample processor. The extracts are analysed by isocratic reversed-phase liquid chromatography using pneumatically and thermally assisted electrospray ionisation (TurboIonspray) with selected reaction monitoring. The method is specific and sensitive, with a range of 4-800 ng/ml in human serum and plasma for both parent drug and metabolite (sample volume 200 microl). The method is accurate and precise with intra-assay and inter-assay precision (C.V.) of <15% and bias <15% for both analytes. The automated extraction procedure is significantly faster than manual sample pre-treatment methods, a batch of 96 samples is extracted in 50 min allowing for faster sample turnaround. The method has been used to provide pharmacokinetic support to biocomparability studies of Leukeran following single doses of oral tablet formulations. PMID- 10517233 TI - Affinity chromatography of plasma proteins (guanidinobenzoatase): use of mimetic matrices and mimetic soluble ligands to prevent the binding of albumin on target affinity matrices. AB - Serum albumin is the most abundant protein in plasma and it has a high capacity to bind many small compounds and macromolecules. In this way, albumin may promote important interferences during affinity chromatography of plasma proteins. Guanidinobenzoatase (GB) is a very relevant plasma protease that seems to be related to tumoral processes. This enzyme may be adsorbed on tailor-made agmatine amide-agarose (CH-A) supports (e.g., the ones having 2 micromol of guanidino groups per ml of agarose attached to the support, through a 6 C aliphatic chain). Such tailor-made supports containing a very low concentration of ionized groups are hardly able to adsorb any protein by anion-exchange. However, they are able to strongly adsorb albumin. In order to solve this problem new mimetic affinity matrices have been designed: (i) by using the same ligand immobilized through a different chemical linkage [guanidino groups attached via secondary amino bonds, (AEA)] or (ii) by using slightly different ligands (e.g., 1,8-octanediamine containing a primary amino group instead of a guanidino one) also attached to the support via amido bonds (CH-DAO). Albumin adsorbs on the target and on the two mimetic matrices while GB is mainly adsorbed on the target one. Moreover, the adsorption of albumin on the affinity matrix (CH-A) is very strongly inhibited by the presence of low concentrations of soluble ligands (e.g., 1,8-octanediamine containing two ionized primary amino groups). On the contrary, the adsorption of GB on CH-A is hardly inhibited by the presence of such mimetic soluble ligand. In this way, the former offering of crude GB samples to AEA plus the use of mimetic inhibitors during adsorption of the extract on CH-A completely prevent the undesirable adsorption of albumin. In a such way, an extremely selective adsorption of GB can be performed. Such an improved chromatography procedure allows a very easy affinity purification and detection of GB. PMID- 10517235 TI - Direct determination of tramadol glucuronides in human urine by high-performance liquid chromatography with fluorescence detection. AB - A sensitive and selective reversed-phase high-performance liquid chromatography method has been developed for the direct determination of three glucuronides of the centrally acting analgesic tramadol (1). Separation of these glucuronides into their diastereomers was achieved by HPLC using ion pair chromatography with nonanesulfonic acid sodium salt and LiChrospher 100 RP 18 as stationary phase. Quantification of O-demethyltramadol glucuronide and N,O-didemethyltramadol glucuronide in human urine was performed by fluorescence detection. The urine samples were purified by a two-step solid-phase extraction. The glucuronides were found to be highly enriched in the 1S,2S-diastereomers. The results of a study with three healthy volunteers are presented. PMID- 10517236 TI - Analyzing mixtures of amino acids and carbohydrates using bi-modal integrated amperometric detection. AB - Described in this work is a new detection methodology - bi-modal integrated amperometric detection - for identifying peaks and as a tool for solving difficult separation problems. Bi-modal integrated amperometry makes it possible to selectively detect amino acids, amino sugars, and carbohydrates following their separation by anion-exchange. Selectivity is gained by two different methods of integrating anodic current on an otherwise identical waveform. As with the single-mode integrated amperometry reported previously, the limits of detection are in the femtomole range and linear calibration plots are possible over three orders of magnitude. This new detection method does not require analyte derivatization. The practical utility of this new technique is demonstrated in the analysis of amino acids and sugars in a recombinant mammalian cell culture medium. PMID- 10517237 TI - Detection and plasma pharmacokinetics of an anti-vascular endothelial growth factor oligonucleotide-aptamer (NX1838) in rhesus monkeys. AB - Aptamers are oligonucleotide ligands selected, in vitro, to bind a specified target protein. The first aptamer to reach human clinical testing is NX1838, a polyethylene glycol conjugated aptamer that inhibits vascular endothelial growth factor. This paper describes the validation of a high-performance liquid chromatographic anion-exchange method for the determination of NX1838 in plasma. Measurements of intact NX1838 had a coefficient of variation of less than 8% and an accuracy between 107% and 115%. The assay was utilized to determine NX1838 plasma pharmacokinetics in rhesus monkeys following a single 1 mg/kg intravenous or subcutaneous dose. Following intravenous administration, the maximum achieved plasma concentration was 25.5 microg/ml with a terminal half-life of 9.3 h and clearance rate of 6.2 ml/h. After subcutaneous administration, the fraction of the dose absorbed into the plasma compartment was 0.78 with a time to peak concentration (4.9 microg/ml) of 8 to 12 h. PMID- 10517238 TI - Determination of hydroxyl free radical formation in human platelets using high performance liquid chromatography with electrochemical detection. AB - The formation of the hydroxyl free radical (HFR) can be quantified indirectly, by measuring two products of the hydroxylation of salicylic acid, 2,3 dihydroxybenzoate (2,3-DHB) and 2,5-dihydroxybenzoate (2,5-DHB). In this study, we used reversed-phase high-performance liquid chromatography with electrochemical (coulometric) detection to measure 2,3-and 2,5-DHB levels in human platelets. The limits of detection of the method were 10 and 5 fmol on column for 2,3-DHB and 2,5-DHB, respectively. We tested the technique by measuring increases in dihydroxybenzoate levels after exposure of platelets to experimentally induced oxidative stress. Then, we measured platelet levels of 2,3 and 2,5-DHB in patients with Parkinson's disease, under therapy with L-DOPA, and in normal subjects. We also measured platelet concentrations of L-DOPA and its major metabolite, 3-O-methyldopa (3-OMD). Parkinsonian patients showed increased levels of both 2,3- and 2,5-DHB. Platelet levels of 2,3-DHB were positively correlated with platelet levels of L-DOPA and 3-OMD. The technique we describe proved simple and extremely sensitive and may represent a useful tool for the study of oxidative stress in humans. PMID- 10517239 TI - Determination of unbound 20(S)-camptothecin in rat bile by on-line microdialysis coupled to microbore liquid chromatography with fluorescence detection. AB - To evaluate the biliary excretion of unbound camptothecin, a flow-through microdialysis probe was constructed for bile sampling. The shunt linear probe was connected from the bile duct, between the liver side to the duodenum to avoid obstruction of the bile duct or bile salt waste. For automatic analysis of microdialysate, an on-line injector was connected to a microbore high-performance liquid chromatographic column with fluorescence detection. Samples were eluted with a mobile phase containing methanol-100 mM monosodium phosphoric acid (35:65, v/v, pH 2.5, adjusted with orthophosphoric acid). The limit of quantification was 1 ng/ml for camptothecin. Following camptothecin administration (5 mg/kg, i.v.), it was found in the bile microdialysate. It was concluded that the in vivo microdialysis technique yields useful data on the biliary excretion of camptothecin. This method is suitable for additional pharmacokinetic studies in rat bile. PMID- 10517240 TI - Simple high-performance liquid chromatographic method for determination of alpha tocopherol in human plasma. AB - A simple high-performance liquid chromatographic method using UV detection was developed for the determination of alpha-tocopherol in human plasma. The method entailed direct injection of the plasma sample after deproteinization using acetonitrile-tetrahydrofuran (3:2). The mobile phase comprised methanol tetrahydrofuran (94:6) and analysis was run at a flow-rate of 1.5 ml/min with the detector operating at 292 nm. A Crestpak C18S (5 microm, 250 mm x 4.6 mm ID) was used for the chromatographic separation. The method had a mean recovery of 93%, while the within-day and between-day coefficients of variation and percentage errors were all less than 7%. The speed, specificity, sensitivity and reproducibility of this method make it particularly suitable for routine determination of alpha-tocopherol in human plasma. Moreover, only a small sample plasma volume (100 microl) is required for the analysis. PMID- 10517241 TI - Rapid and simple micro-determination of carvedilol in rat plasma by high performance liquid chromatography. AB - We studied the use of high-performance liquid chromatography (HPLC) with spectrofluorometric detection, using a solid-phase extraction for a simple, rapid and sensitive determination of plasma carvedilol levels in rats. Extracted aliquots were analyzed by HPLC, using a reversed-phase octadecyl silica column. The analytical mean recovery of carvedilol added to the blank plasma was 94.2%. The detection limit was 3.6 ng/ml in the plasma. The reproducibilities (C.V.) were 2.7-7.5% for the within-day assay, and 2.6-7.4% for the between-day assay, indicating that the method was effective for the determination of carvedilol plasma levels. PMID- 10517242 TI - Determination of famotidine in human plasma and urine by high-performance liquid chromatography. AB - An improved, rapid and specific high-performance liquid chromatographic assay was developed for the determination of famotidine in human plasma and urine. Plasma samples were alkalinized and the analyte and internal standard (cimetidine) extracted with water-saturated ethyl acetate. The extracts were reconstituted in mobile phase, and injected onto a C18 reversed-phase column; UV detection was set at 267 nm. Urine samples were diluted with nine volumes of a mobile phase internal standard mixture prior to injection. The lower limits of quantification in plasma and urine were 75 ng/ml and 1.0 microg/ml, respectively; intra- and inter-day coefficients of variation were < or =10.5%. This method is currently being used to support renal function studies assessing the use of intravenously administered famotidine to characterize cationic tubular secretion in man. PMID- 10517243 TI - Combined method for the determination of gamma-aminobutyric and beta-alanine in cerebrospinal fluid by stable isotope dilution mass spectrometry. AB - A previously described method for the determination of GABA in CSF has been expanded to include both GABA and beta-ALA, using a single GC-MS analysis. A stable isotope labelled internal standard for beta-ALA was synthesised to achieve accurate quantification. This new combined method expands the diagnostic power compared to an isolated GABA measurement. Control values for free and total GABA and free and total beta-ALA are described. Age <2 years: free GABA 0.017-0.067 microM, total GABA 4.2-13.4 microM; free beta-ALA 0.049-0.11 microM, total beta ALA 2.1-4.6 microM. Age >2 years: free GABA 0.032-0.17 microM, total GABA 3.3 12.2 microM; free beta-ALA 0.021-0.058 microM, total beta-ALA 0.91-3.5 microM. PMID- 10517244 TI - Gas chromatographic-mass spectrometric method for quantitative determination of ketotifen in human plasma after enzyme hydrolysis of conjugated ketotifen. AB - A validated method for determination of total amount of ketotifen (unchanged and conjugated) in human plasma has been presented. An enzyme hydrolysis of conjugated ketotifen was conducted with combination of beta-glucuronidase and arylsulfatase. After the enzyme hydrolysis a solid-phase extraction was applied as a cleaning step. The quantitative determination by gas chromatography with mass-spectrometry detection (GC-MS) was performed. Pizotifen has been used as an internal standard. A reliable hydrolysis as well as a satisfactory accuracy, improved precision in the linear region from 0.500 to 10.0 ng/ml plasma, limit of detection of 0.010 ng/ml and prolonged capillary column life have been achieved. PMID- 10517245 TI - Genetic aspects of population policy. AB - Every science begins in folklore and matures as it reacts against dogma and myth. Astronomy developed in the Neolithic, but it did not outgrow astrology until the sixteenth century. Chemistry discarded alchemy at about the same time. On the contrary, the short history of genetics has been concurrent with the pseudo science of eugenics, which, at times, has been widely accepted and incorporated in population policy and directive genetic counselling, with rare opposition by geneticists. Societal pressures are likely to increase with the power of genetic technology, the fear it generates and the perception that population growth threatens human welfare. Without a pertinent ethical code, geneticists are vulnerable to both temptation and opprobrium. The intrusion of eugenics into genetic counselling has been a recent source of concern to societies and congresses of genetics. This review traces the causes of this concern and the manner of its expression in the absence of an international voice for genetics that could address ethical and other common interests. PMID- 10517246 TI - Genetic landmarks through philately--Kabuki theater and Kabuki syndrome. PMID- 10517247 TI - Genetic counselling in multiple sclerosis: risks to sibs and children of affected individuals. AB - Genetic factors are recognized as having important roles in both the overall etiology and the familial aggregation of multiple sclerosis (MS), leading to increased requests for genetic counselling. This paper is designed to provide familial risk data in a practical format for use during genetic counselling for MS. Depending on the amount of genetic sharing among family members, the relative risk of MS compared with that for the general population can range from 1 (adopted sibs and children of the MS proband, with whom they share no genetic material) to 190 (monozygotic co-twins of MS patients, with whom they share 100% of their genetic material). When counselling full sibs of MS patients, risks can be better calculated if information is available on the age of MS onset in the patient and whether or not one parent has MS. PMID- 10517248 TI - Molecular and clinical characterization of a patient with duplication of 1p36.3 and metopic synostosis. AB - Chromosome 1p duplications are rare. There have been only 11 reported cases of isolated 1p duplication, all of which were proximal, interstitial duplications. We present a patient with a terminal duplication of 1p (1p36.3). To our knowledge, this is the first such reported case. Our patient presented with metopic synostosis, rectal stenosis, atrial septal defect, and mildly delayed gross motor development. Molecular characterization using microsatellite marker analysis and fluorescence in situ hybridization (FISH) revealed an area of duplication between p58 and D1S2893, approximately 13 cM in size. We compare our patient's clinical findings with the clinical phenotype found in patients with the corresponding deletion of 1p36.3 and discuss the role of gene dosage in other deletion/duplication syndromes. PMID- 10517249 TI - FISH analysis with locus-specific probes in sperm from two translocation carrier men. AB - Meiotic segregation of normal and derivative chromosomes was analysed in sperm samples from two balanced reciprocal translocation carrier men by use of dual colour fluorescence in situ hybridisation (FISH) technique. The translocations were t(4;8)(p15;p12) and t(15;22)(q(23:q13.2), and the digoxigenin-labelled FISH probes were specific to either the translocated or centric segments of the chromosomes involved in the translocations. A total of 1000 spermatozoa for each probe were analysed and the modes of segregation were described on the basis of signals in each sperm cell. The mean frequency of alternate and/or adjacent-1 (adj-1) segregation types was 69.47%, whereas they were 30.51 and 78.70% for the adjacent-2 (adj-2) and alternate/adj-2 segregation types, respectively. This study illustrated that FISH is a valuable technique for analysing the meiotic segregation products of the heterozygotes in respect to aneuploidy risk. PMID- 10517250 TI - STK11/LKB1 germline mutations are not identified in most Peutz-Jeghers syndrome patients. AB - Germline mutations of the STK11 gene mapped to chromosome 19p13.3 are responsible for Peutz Jeghers syndrome (PJS), a dominant disorder associated with characteristic gastrointestinal hamartomatous polyps and a predisposition to various cancers. We conducted a detailed investigation of germline STK11 alterations by protein truncation test and genomic DNA sequence analysis in ten unrelated PJS families. We identified a novel truncating deletion spanning STK11 exons 2-7 in a single patient and several known polymorphisms. Loss of heterozygosity studies in PJS polyps of four of these patients identified an allelic deletion of D19S886 in another patient. Our results suggest that STK11 mutations account for only a proportion of PJS cases. PMID- 10517251 TI - Possible interaction of genotypes at cystathionine beta-synthase and methylenetetrahydrofolate reductase (MTHFR) in neural tube defects. NTD Collaborative Group. AB - Neural tube defects are a common, complex disorder with genetic and environmental components to risk. We investigated the previously reported interaction between homozygosity for the thermolabile variant at the methylenetetrahydrofolate reductase and heterozygosity for the 844ins68 allele at the cystathionine beta synthase loci in cases with lumbosacral myelomeningocele and their parents. Using control allele frequencies from our sample pooled with those published in the literature, we confirm a marginally significant interaction at these two loci. This finding suggests that additional, larger studies are warranted to investigate this possible interaction in more detail. PMID- 10517252 TI - Short rib-polydactyly syndrome: more evidence of a continuous spectrum. AB - We report a fetus with radiological features of the four established types of short rib-polydactyly syndrome (SRPS). The phenotype of this fetus supports the previously suggested hypothesis that the different subtypes of the short rib and polydactyly syndrome are not single entities, but rather, part of a continuous spectrum with variable expressivity. PMID- 10517253 TI - Aicardi-Goutieres syndrome: monogenic recessive disease, genetically heterogeneous disease, or multifactorial disease? AB - Aicardi-Goutieres syndrome (AGS) is a severe progressive familial encephalopathy, which is usually diagnosed shortly after birth. Using the principle of homozygosity mapping, genome-wide screening of five consanguineous families was performed to search for a homozygous region shared by all affected individuals. A total of 364 markers with an average spacing of 9.9 cM were genotyped, but no homozygous region common to all affected individuals could be found. Regions of homozygosity in affected sibs could only be identified within each family individually. This may reflect genetic heterogeneity, possibly related to clinical heterogeneity, since several syndromes are clinically difficult to distinguish from AGS. Involvement of a small number of genes and/or of an external factor, such as infection, may also explain the absence of a homozygous region common to all affected individuals. PMID- 10517254 TI - Kabuki syndrome: description of dental findings in 8 patients. AB - The cardinal features of Kabuki (Niikawa-Kuroki) syndrome (KS) include characteristic facial dysmorphic features, mild to moderate mental deficiency, skeletal abnormalities, dermatoglyphic abnormalities, and postnatal growth retardation. We identified 8 patients with KS in a genetics clinic over the past 5 years. All were Caucasians, except for 2 who were of mixed Aboriginal and Caucasian descent. All had the facial gestalt, the dermatoglyphic abnormalities characteristic of the syndrome, and developmental delay. Dental abnormalities of permanent teeth were seen in all 8 cases; 6 had missing lower incisors. Five patients had uniquely abnormal upper incisor teeth shape; the upper incisors had a 'flat head' screwdriver-shaped appearance. Other dental abnormalities included missing lower lateral incisors, missing second premolars, and ectopic upper 6 year molars. We believe the presence of the unique dental findings will prove useful in the diagnostic assessment of individuals with KS. PMID- 10517255 TI - Physical activity modulates the effect of a lipoprotein lipase mutation (D9N) on plasma lipids and lipoproteins. AB - We investigated interactions between a mutation (D9N) in the lipoprotein lipase (LPL) gene and physical activity, as well as other lifestyle factors, on lipid traits in a population-based sample of Dutch men and women (n = 379). We used questionnaire information to classify physical activity, alcohol consumption, and smoking habits, while overweight was defined as a body mass index (BMI) > 25 kg/m2. Non-fasting blood samples were used for the determination of lipid traits and the D9N genotype. Fifteen subjects (4%) carried the mutation. They presented with higher levels of total cholesterol, apolipoprotein (apo) B and triglycerides compared to non-carriers. While no interactions with overweight, alcohol consumption, and smoking were found, a strong interaction between the D9N mutation and physical activity became apparent. Physically inactive D9N carriers (n = 5) had considerably higher total cholesterol (+2 mmol/l, p < or = 0.0001) and apo B levels (+63 mg/dl, p < or = 0.0001) compared to non-carriers of this mutation, whereas their high-density lipoprotein (HDL)-cholesterol concentrations were lower (-0.22 mmol/l, p < 0.05). This was not the case for physically active D9N carriers (n = 10). In conclusion, a common variant of the LPL gene (D9N) adversely affects plasma lipid and lipoprotein profiles. However, the unfavorable consequences may be counteracted by physical activity. PMID- 10517256 TI - Serum total IgE levels and CD14 on chromosome 5q31. PMID- 10517257 TI - Association between CAG repeat number in the androgen receptor and male infertility in a Belgian study. PMID- 10517258 TI - High frequency of type 1 GM1 gangliosidosis in southern Brazil. PMID- 10517259 TI - Scalp-ear-nipple (Finlay-Marks) syndrome: a familial case with renal involvement. PMID- 10517260 TI - A Japanese patient homozygous for the H1085R mutation in the CFTR gene presents with a severe form of cystic fibrosis. PMID- 10517261 TI - Symbrachydactyly involving both the hand and foot. PMID- 10517262 TI - High-fat feeding reduced muscle uncoupling protein 3 expression in rats. AB - Uncoupling Protein 3 (UCP3), largely expressed in skeletal muscle, is modulated by cold, thyroid hormones, leptin, fasting-refeeding and exercise training among other factors in a tissue-specific manner. In brown adipose tissue, there is an increase in UCP3 levels after high-fat feeding and beta3-adrenergic agonist treatment. Controversial effects of these agents have been reported in skeletal muscle. The aim of this experimental trial was to evaluate the effect of high-fat feeding and beta3-adrenergic agent treatment on skeletal muscle UCP3 expression levels. Lean rats were fed a cafeteria diet for 30 days and found to have significantly higher fat stores and body weight than control rats at the end of the experimental period. When cafeteria-diet rats were daily i.p. injected with Tertatolol for 30 days; a decrease in total fat mass and body weight was found. Such an effect was not observed in fa/fa rats. Interestingly, gastrocnemius muscle UCP3 mRNA levels were significantly reduced in cafeteria-diet rats when compared to lean animals. Likewise, mitochondrial O2 consumption in gastrocnemius muscle was also significantly decreased (-31%) in cafeteria-diet rats as compared to the control group. It is suggested that the down-regulation of UCP3 gene expression together with the lower O2 consumption observed in high fat fed rats may be linked to lower fatty oxidation, which would promote triglyceride accumulation. PMID- 10517263 TI - Calcium metabolism in bone and teeth of rats during exposure to restriction of motor activity and to swimming exercise. AB - The effects of motor activity restriction for 90 days (hypokinesia, HK) and swimming training (T) on calcium metabolism in rat bones and teeth were evaluated. Male Wistar rats were distributed in four groups: untrained vivarium control rats (UVCR), untrained hypokinetic rats (UHKR), trained hypokinetic rats (THKR) and trained vivarium control rats (TVCR). Hypokinesia was obtained keeping the animals for 90 days in small individual cages which restricted their movements in all directions without hindering food and water intakes. Rats of THKR and TVCR were forced to swim for 15 to 90 minutes everyday. On the 1st, 7th, 15th day of a prehypokinetic period and on the 5th, 10th, 20th, 40th, 60th and 90th day of the hypokinetic period, six rats of each group were decapitated. Radioactive calcium was injected to the animals 70 days before autopsy. Calcium and phosphorus in serum, bones (molars, incisors, upper and lower jaws, parietal, scapular, clavicle, pelvic and tibial bones) and in the respective ash residues were measured. Body and bone weights, and radioactive calcium were also determined. Under prolonged exposure to HK (THKR and UHKR groups), bone weights and bone and ash Ca and P concentrations decreased, whereas serum Ca and P and 45Ca resorption increased, in comparison to the respective values in the UVCR and TVCR groups. Swimming exercise apparently did not modify calcium metabolism in the hypokinetic or control rats. PMID- 10517264 TI - Growth modification of human colon adenocarcinoma cells exposed to a low frequency electromagnetic field. AB - The influence of variable low-intensity, low-frequency electromagnetic fields on culture cells is investigated. Human colon adenocarcinoma cells were exposed to a rectangular and variable magnetic field (1 and 25 Hz; 1.5 mT peak). Cultures were exposed to a dose for 15 and 360 minutes, and after 24 hours incubation, cell viability was measured with neutral red stain. The group treated for 15 minutes showed a statistically significant increase in cell growth with 1 Hz (p < 0.002) and 25 Hz (p < 0.003). In contrast, a significant decrease in cell growth was found in those cultures treated with 1 Hz for 360 minutes (p < 0.02). The effects reported could be influenced by the magnetic field frequency and the exposure time. PMID- 10517265 TI - CCK-mediated response in the activation of 5-HT receptor types in the guinea-pig ileum. AB - The possible interaction between cholecystokinin (CCK) and 5-hydroxytryptamine (5 HT) was evaluated in vitro in the longitudinal muscle-myenteric plexus of the guinea-pig ileum. Devazepide and L-365,260 were used to block CCKA and CCK(B) receptors and ondansetron and tropisetron to block 5-HT3 and 5-HT4 receptors, respectively. The CCK receptor antagonists blocked, in a dose-dependent manner, the response to 5-HT and to the selective agonists at 5-HT3 and 5-HT4 receptors, 2-methyl-5-hydroxytryptamine (2-Me-5-HT) and 5-methoxytryptamine (5-MeOT), respectively. The blockade was almost complete on the first phase of the concentration response curve to 5-HT and for all the concentrations of 5-MeOT tested. In the 2-Me-5-HT-induced contractile response there was a component with the same sensitivity to devazepide and to the selective NK1 receptor antagonist, GR 82334, and another resistant component that was abolished by atropine. However, the blockade of the NK1 receptor did not produce a significant increase in the inhibition obtained when atropine or devazepide were separately tested on the 5-MeOT-induced response. These results suggest that CCK is involved in the 5 HT-induced contractile response, particularly in the response induced by 5-HT4 receptor stimulation. PMID- 10517266 TI - Response of hypophyso-thyroid-axis to surgery under halothane anaesthesia. PMID- 10517267 TI - Providing effective low-cost back lighting for localized surgical procedures. AB - A method of providing back light illumination was developed in the context of injecting rat fetuses. The illuminator had a compact flat design measuring approximately 2.6 cm x 6 cm, with minimal thickness (about 7 mm), and a proportionately large amount of active area. The top working surface was a glass plate through which the illumination was provided. The head provided diffuse light and remained relatively cool during operation. The unit was water proof (suitable for immersion in a warm saline bath) and provided isolation from hazardous voltages. The head was able to be freely positioned, with a small, flexible power lead that exited from a non-active area of the unit. The head was free of sharp or rough edges and was suitable for direct contact with delicate tissues. PMID- 10517268 TI - A macrophage hippocampal slice co-culture system: application to the study of HIV induced brain damage. AB - We have developed an in vitro system that allows the study of the effects of factors released from macrophages on neuronal and glial survival in cultured hippocampal slices. Organotypic hippocampal slice cultures are grown on semi permeable membranes in stationary co-culture with a murine macrophage cell line (RAW 264.7). The two culture systems are separated by a semi-permeable membrane specifically allowing the study of diffusable factors between the two culture systems. The use of the fluorescent exclusion dye propidium iodide as an in vitro marker of cell viability allows the study of progressive toxicity as it evolves in the slice cultures. We demonstrate that the HIV-1 derived nuclear regulatory protein Tat induces toxicity in slice cultures via the production of soluble mediators. The advantages of organotypic cultures over other in vitro systems is discussed as well as the general applicability of this method to the study of other brain pathologies, where macrophage derived factors are thought to play a role in neuronal survival. PMID- 10517269 TI - Non-HPLC separation of water-soluble choline metabolites by two-dimensional high voltage electrophoresis and thin layer chromatography. AB - In cholinergic neurons choline is directed to three main pathways; (1) conversion to phosphorylcholine (PCh) and cytidine diphosphate choline (CDP-choline) for the synthesis of phosphatidylcholine, (2) acylation to the neurotransmitter acetylcholine and (3) oxidation to betaine for the formation of methionine. Thus, the distribution of choline among the different metabolites is important for a better understanding of the regulation of these pathways in neurons. A non-HPLC method for the simultaneous separation of five choline metabolites found in neurons is described. High voltage electrophoresis (HVE) was combined with thin layer chromatography (TLC) to separate choline, PCh, CDP-choline, acetylcholine and betaine. This method is useful in studying the distribution of choline among its different metabolites in radiotracer experiments. Aqueous metabolites from leukemia inhibitory factor treated LA-N-2 cells labeled with [methyl-3H]choline were separated by HVE followed by TLC in the same dimension. Although the separation appeared to be complete, some 'tailing' by PCh significantly elevated the radioactivity measured in CDP-choline. This tailing of PCh was confirmed by subjecting radiolabeled PCh alone to this multiple separation method. Contamination of CDP-choline by PCh was eliminated by subjecting the samples to HVE followed by TLC in the second dimension. This two-dimensional approach was consistently reproducible and achieved excellent resolution of all five metabolites. In addition, this technique also resolved a sixth choline-containing metabolite, glycerophosphorylcholine (GPC), a breakdown product of phosphatidylcholine. PMID- 10517270 TI - Long-term cortical CBF recording by laser-Doppler flowmetry in awake freely moving rats subjected to reversible photothrombotic stroke. AB - This study aimed at developing a laser-Doppler flowmetry (LDF) device suitable for long-term cortical cerebral blood flow (cCBF) measurement in awake, freely moving rats. The device included a flow probe adapter for permanent fixation to the skull bone and a connector that held the flow probe in the adapter in exactly the same position during repeated cCBF recordings. With this LDF recording system, cCBF values were stable and unaltered in awake, freely moving rats up to 4 days after operation compared with initial recordings during anesthesia. Repeated cCBF measurements in rats after transient removal and reattachment of the flow probe revealed a coefficient of variation of 7.0-17.4%. The LDF recording system was applied to rats subjected to a photothrombotic ring stroke lesion. cCBF in the region-at-risk declined to 59-34-26-33% of baseline values (P < 0.01) at 1-2-24 48 h after irradiation with gradually restored cCBF values of 56-87% at 72-96 h post-irradiation (P < 0.01 vs. 24 h). Transcardial carbon black perfusion examination of the brains confirmed the sustained hypoperfusion in the region at risk up to 48 h post-ischemia followed by a consistently occurring late spontaneous reperfusion. In conclusion, a novel laser-Doppler cortical CBF recording system has been set up that allows stable long-term cortical CBF follow up in awake, freely moving rats. PMID- 10517271 TI - Measurement of calcium flux through ionotropic glutamate receptors using Cytostar T scintillating microplates. AB - Human embryonic kidney cells (HEK293), expressing the human GluR4 receptor sub unit of 2-amino-3-hydroxy-methylisoxazol-4-ylpropionic acid (AMPA) type non-NMDA receptors were used, in combination with Cytostar-T scintillating microplates, to develop an assay system for the screening of novel compounds with potential AMPA antagonistic characteristics. Agonist dose responses were measured using the agonists: AMPA; quisqualic acid; L-glutamic acid and kainic acid (KA), and EC50 values of 40, 10, 100 and 100 microM were estimated for each of the agonists, respectively. The AMPA receptor antagonists LY293558 and GYK152466 were tested and shown to inhibit agonist induced [45Ca] influx into the cells. An IC50 value of 600 microM was estimated for the competitive antagonist LY293558 and a value of 100 microM estimated for the non-competitive antagonist GYK152466. The developed assay system is homogeneous, allowing increased assay precision and speed. This allows the potential for automation of the assay and it may be used for screening large numbers of novel compounds. PMID- 10517272 TI - Ultra-miniature headstage with 6-channel drive and vacuum-assisted micro-wire implantation for chronic recording from the neocortex. AB - We describe a head-stage, with precision microtranslators for the chronic placement of micro-wire electrodes in the neocortex, that minimizes compressive damage to the brain. The head-stage has a diameter of 5.8 mm and allows six electrodes, separated by 450 microm on a hexagonal grid, to be individually and continuously positioned throughout a depth of approximately 3 mm. Suction is used to transiently support the dura against a curved array of tubes that guide and stabilize the electrodes as a means to prevent compression of the neocortex as the electrodes breach the dura. With this headstage we recorded extracellular signals in a rat immediately after surgery. Single-unit waveforms at a given electrode position were stable for at least several hours in the freely behaving animal and were obtained throughout the depth of the neocortex for at least 2 months. Electrophysiological records and histological examination showed that the upper layers of the neocortex were intact and minimally damaged after the implantation. PMID- 10517273 TI - A comparison of cross-correlation and surface EMG techniques used to quantify motor unit synchronization in humans. AB - Two methods used to estimate the strength of motor unit (MU) synchronization in a muscle are the direct cross-correlation of MU discharge times, and averaging of the surface electromyogram (SEMG) with respect to discharge of a reference MU. Although indirect, the latter approach has the advantage that a global estimate of MU synchrony can be obtained quickly and easily. The two methods are generally regarded as providing equivalent information on the extent of MU synchronization in a muscle, but this proposition has not previously been tested quantitatively. In the present study, we used both the SEMG technique (189 MUs) and cross correlation of MU discharge (498 MU pairs) to estimate MU synchrony in 28 first dorsal interosseus (FDI) muscles from 16 subjects. Despite considerable overlap in the identity of MUs used to quantify synchrony with each method, linear regression revealed no significant correlation between the estimates of MU synchronization in FDI muscles obtained with the two techniques (r2= 0.04, n = 28). This discrepancy was not due to insufficient sampling of the MU population with the cross-correlation method, although we found evidence for a non-uniform tendency for synchronous discharge in two of 13 motor units providing sufficient data for the analysis. The most likely explanation for the discrepancy between the estimates of MU synchrony is that methodological problems with the SEMG technique limit its accuracy. These problems are difficult to avoid under normal experimental conditions, and we conclude that the SEMG method is not reliable for quantitative comparisons of MU synchrony between muscles and subjects. PMID- 10517274 TI - Examination of the spatial and temporal distribution of sensory cortical activity using a 100-electrode array. AB - This paper introduces improved techniques for multichannel extracellular electrophysiological recordings of neurons distributed across a single layer of topographically mapped cortex. We describe the electrode array, the surgical implant techniques, and the procedures for data collection and analysis. Neural events are acquired through an array of 25 or 100 microelectrodes with a 400 microm inter-electrode spacing. One advantage of the new methodology is that implantation is achieved through transdural penetration, thereby reducing the disruption of the cortical tissue. The overall cortical territory sampled by the 25-electrode array is 1.6 x 1.6 mm (2.56 mm2) and by the 100-electrode array 3.6 x 3.6 mm (12.96 mm2). Using a recording system with 100 channels available, neural activity is simultaneously acquired on all electrodes, amplified, digitized, and stored on computer. In our data, average peak-to-peak signal/noise ratio was 11.5 and off-line waveform analysis typically allowed the separation of at least one well-discriminated single-unit per channel. The reported technique permits analysis of cortical function with high temporal and spatial resolution. We use the technique to create an 'image' of neural activity distributed across the whisker representation of rat somatosensory (barrel) cortex. PMID- 10517275 TI - Ultra-violet light-induced changes in membrane properties in secretory cells. AB - Illumination with ultra-violet is used widely in physiological experiments for the photolysis of caged compounds. In the peptidergic cells of the pituitary gland, as well as cultured PC12 cells, ultra-violet light was found to produce changes in a number of membrane properties. Light of sufficient intensity to produce rapid photolysis of commonly used caged compounds induced changes in K+ and Ca2+ current, as well as changes in membrane capacitance. All responses to light showed a rapid timecourse, activating in a few ms and decaying within 10-50 ms after illumination ended. Experiments with radical scavengers and with inhibitors of cytochrome p450 and phospholipase A2 failed to block the light responses. These rapid responses to light emphasize that experiments employing ultra-violet light in the photorelease of physiological and pharmacological agents require special care for control of light artifacts. PMID- 10517276 TI - A method for increasing the viability of the external portion of lumbar catheters placed in the spinal subarachnoid space of rats. AB - A method for direct catheterization of the lumbar subarachnoid space has recently been developed by Storkson et al. (1996) (J Neurosci Methods 1996;65:167-172) that may potentially improve upon the widely used method of Yaksh and Rudy (1976) (Physiol Behav 1976;17:1031-1036). This 'catheter-through-a-needle' technique inserts the catheter between lumbar vertebrae 5 (L5) and 6 (L6), which has been shown to reduce neurological impairment and post-surgical deaths. However, employing this technique allows the external portion of the chronic indwelling catheters to be easily damaged, resulting in approximately 50% attrition within 4 days after surgery. Therefore, we developed an easy and inexpensive method for protecting the external portion of the catheter that enhances catheter viability beyond 14 days after catheter implantation while also maintaining a low injection volume (8 microl). Moreover, this modification does not significantly alter the implantation methods developed by Storkson et al. (1996) (J Neurosci Methods 1996;65:167-172) and allows for more optimal catheter materials to be incorporated. Chronically implanted catheters (n = 70) with the external portion of the catheter protected, resulted in 4% attrition 7 days after surgery and 11% attrition 14 days after surgery. Approximately 5.5% of animals implanted showed very mild and transient neurological impairment. PMID- 10517277 TI - Nutritional studies on rats and fish (carp Cyprinus carpio) fed diets containing unheated and heated Jatropha curcas meal of a non-toxic provenance. AB - Unheated and heated (121 degrees C, 66% moisture; 15, 30 and 45 min) Jatropha meals of non-toxic provenance from Veracruz state in Mexico were evaluated using rats and fish. With rats, the weight gain was highest for the casein diet followed by heated (30 min; only this treatment was studied using rats) and unheated Jatropha meal containing diets. The protein efficiency ratio (PER) for unheated and heated Jatropha meal containing diets was 37 and 86%, respectively, of the casein diet. On the other hand, the body weight gain, PER and feed conversion ratio of fish were statistically similar for unheated and heated (15, 30 and 45 min) Jatropha meal containing diets fed for a period of 35 days. Although these parameters were statistically similar for the unheated and heated Jatropha meal containing diets, the body weight gain, PER and protein productive value were highest and the feed conversion ratio lowest with 15 min heated Jatropha meal, suggesting that the heat treatment for 15 min is optimal for the meal. Trypsin inhibitor and lectin activities decreased drastically (>83 and 99%, respectively) after 30 and 45 min of heat treatment and after 15 min, the residual lectin activity was negligible and the residual trypsin inhibitor activity was 34%. These results, together with the nutritional parameters investigated, imply that Jatropha trypsin inhibitors and lectins do not have any adverse effects on carp at least up to 35 days of feeding. The nutritional value of Jatropha meal of the non-toxic provenance is high, and potential exists for its incorporation into the diets of monogastrics, fish and possibly humans. PMID- 10517278 TI - Chemical composition of seeds and oil of Xylopia aethiopica grown in Nigeria. AB - The chemical composition and mineral constituents of Xylopia aethiopica, which is valued as a spice in Nigeria, were determined along with the physicochemical characteristics of the seed oil. The seeds had the following chemical compositions moisture (8.43 g/100 g), ash (5.89 g/100 g), crude lipid (9.58 g/100 g), crude protein (12.45 g/100 g) crude fiber (8.66 g/100 g) and carbohydrate (63.65 g/100 g). Calcium and potassium were the major minerals in the seed. The extracted lipid was examined for fatty acid composition. Linoleic (45.1 g/100 g) and oleic (26.5 g/100 g) acids were the predominant unsaturated fatty acids, while palmitic acid (18.0 g/100 g) was the major saturated acid. The iodine value of 97 g/100 g indicates that the seed oil is a non-drying type. PMID- 10517279 TI - Studies on the production of bambara groundnut (Vigna subterranea) tempe. AB - Bambara groundnut, an indigenous African legume, was subjected to fermentation by three strains of Rhizopus. One strain B. arrhizus could not ferment the substrate. Mycelial penetration and binding was good when strains NRRL 2710 (R. oligosporus) and NRRL 1477 (R. stolonifer) were used. Fermentation by both strains resulted in increases of pH, moisture, protein and fat while total carbohydrate decreased by 50%. Sensory evaluation showed that bambara groundnut tempes rated similar (p>0.5) in taste and texture and higher (p<0.05) in color and flavor than soybean tempe. Bambara groundnut would be an acceptable food product in the diet as a good protein supplement. PMID- 10517280 TI - Influence of palm oil (Elaesis guineensis) on health. AB - In recent times there has been a growing research interest in palm oil, one of the major edible plant oils in the tropical countries, because of the link between dietary fats and coronary heart disease. Obtained from a tropical plant, Elaesis guineensis, it has a polyunsaturated fatty acid/saturated fatty acid ratio close to unity and a high amount of antioxidant vitamin A precursors and vitamin E. Palm oil is consumed in the fresh state and/or at various levels of oxidation. Feeding experiments in various animal species and humans have highlighted the beneficial role of fresh palm oil to health. These benefits include reduction in the risk of arterial thrombosis and atherosclerosis, inhibition of cholesterol biosynthesis and platelet aggregation, and reduction in blood pressure. However, a considerable amount of the commonly used palm oil is in the oxidized state which possesses potential dangers to the physiological and biochemical functions of the body. Oxidation is as a result of processing the oil for various culinary purposes. Studies have revealed that relative to fresh palm oil, oxidized palm oil induces an adverse plasma lipid profile, free fatty acids, phospholipids and cerebrosides. Additionally, oxidized palm oil induces reproductive toxicity and organotoxicity particularly of the kidneys, lungs, liver and heart. Available evidence suggests that at least part of the oxidized oil impact on health reflects generation of toxicants due to oxidation. The reduction of the dietary level of oxidized oil and/or the level of oxidation may reduce the health risk associated with consumption of oxidized fats. PMID- 10517281 TI - In vivo degradation and stimulating effect of phaseolin on nitrogen secretion in rats. AB - In short-term feeding experiments, about 78% of the phaseolin administered to rats was degraded regardless of the amounts of phaseolin intubated. In contrast, the total N found in the feces increased rapidly and exceeded the original administered amounts. The bulk of N output was not immunologically related to the glycoprotein. The effects of phaseolin on the stimulation of endogenous N secretion in the small intestine were confirmed from the results of acute experiments. Phaseolin fragments, derived from the breakdown of the native protein, when reapplied intragastrically to rats, were broken down further and to a similar extent as the original glycoprotein and were even more potent related to stimulation of N secretion. It is suggested that this secretagogue biological activity of phaseolin and not its resistance to gut proteolysis, is the main reason for the poor nutritional value of this glycoprotein. PMID- 10517282 TI - Acceptability of supplementary foods based on popped cereals and legumes suitable for rural mothers and children. AB - Eight types of supplementary foods based on popped cereals (wheat, ragi, bajra and sorghum) blended with legumes (soy and bengal gram) and fortified with essential vitamins and minerals were developed on a pilot plant scale. Four of the supplements were prepared with cereals, soy flour (SF) and bengal gram (BG) dhal and the other four were prepared with combinations of cereals and SF. These blends were mixed with jaggery (obtained by boiling juice out of sugarcane) syrup and pressed into compact form. One hundred gram portions of these foods provided 370+/-20 kilocalories and 11+/-1 g protein. Moisture, crude protein, total carbohydrates, total lipids, ash, dietary fiber and energy contents, of all the developed supplements were within the ranges prescribed by the Indian Standards Institute for processed weaning foods and could satisfactorily meet one-third of the Recommended Dietary Allowance (RDA) of these nutrients per day for preschool children. Organoleptic evaluation and feeding trials revealed that the foods were well accepted by rural mothers and children. PMID- 10517283 TI - Reactivity of red palm oil and cyanide ion: implications for the cyanogen content of palm oil-treated gari. AB - Red palm oil was tested for the reactivity of its components with CN-, and alkaline picrate as the color developing reagent. Palm oil components have a low level absorbance at 490 nm which is reduced significantly (p< or =0.01) after reaction with CN-. Hydrolysis of palm oil components, and reaction of the hydrolysis products with CN- significantly increased the absorbance at 490 nm. In contrast, after reaction of palm oil with alkaline picrate, the absorbance at 490 nm is very high; this is reduced significantly by reaction with CN-, hydrolysis with 0.2 M NaOH, and reaction of the hydrolysis products with CN- before treatment with alkaline picrate. The results indicate that palm oil component(s) sequester CN- into a complex which may not be correctly estimated during cyanide quantification, resulting in the absence, or low levels of cyanide in palm oil fried gari as earlier reported. PMID- 10517284 TI - Effect of domestic processing on total and extractable calcium and zinc content of bathua (Chenopodium album) and fenugreek (Trigonella foenum graecum) leaves. AB - Bathua (Chenopodium album) and fenugreek (Trigonellafoenum graecum) stored in polyethylene bags and without packaging for 24 or 48 hours in a refrigerator at 5 or 30 degrees C in polyethylene bags. The fresh leaves were also dried (oven and sun); blanched (5, 10 or 15 min) and cooked in an open pan and a pressure cooker. The processed leaves were analyzed for total and extractable calcium and zinc content. The Ca and Zn content of these leaves varied from 970 to 2230 and 10.50 to 12.30 mg/100 g DM and the percentage HCl-extractability was 80.34 to 83.04 and 82.43 to 83.90, respectively. Non significant effects of drying and storage were observed on total Ca and Zn content and HCl-extractability while blanching and cooking resulted in significant improvement of HCl-extractability of these two minerals. Thus, cooking and blanching are good ways to improve the HCl extractability of Ca and Zn. PMID- 10517285 TI - The bioavailability of magnesium from Wakame (Undaria pinnatifida) and Hijiki (Hijikia fusiforme) and the effect of alginic acid on magnesium utilization of rats. AB - The bioavailability of magnesium from Wakame and Hijiki, and the effects of alginic acid on absorption of dietary magnesium were examined in five groups of rats fed either control, Wakame, Hijiki, AW (containing the same amount of alginate as in the Wakame) and AH (containing the same amount of alginate as in the Hijiki) diets, and animals fed a low magnesium diet (LMg) (twentieth amount of magnesium in the original mineral mixtures as the control). Food intake and body weight gain were decreased by adding sodium alginate to the diets. A large amount of calcium accumulated only in the kidneys of the rats fed the LMg diet. Serum magnesium concentration decreased only in the LMg group. The magnesium content in the defatted left femurs did not differ between the control and Wakame fed animals and also among the animals eating Wakame, Hijiki and AW diets. The breaking force of the right femurs did not differ among all the groups except the LMg group. The ratio of apparent magnesium absorption (%) of the control, LMg, Wakame, Hijiki, AW and AH groups was 82.2, 72.7, 66.9, 50.8, 69.3 and 54.2 in the first experimental period, and was 75.3, 52.1, 57.7, 46.9, 62.6 and 60.5 in the second experimental period, respectively. It was clear that the bioavailability of magnesium in the Wakame fed rats was higher than in those eating the Hijiki. Large amounts of sodium alginate lowered magnesium absorption from the diet. PMID- 10517286 TI - The molecular clock, the biological clock and general physiology and biophysics. PMID- 10517287 TI - Meso-tetraphenylporphyrin in liposomes as a suitable photosenzitizer for photodynamic therapy of tumors. AB - The suitability of a liposomal form of hydrophobic nonsulfonated meso-tetraphenyl porphyrin (TPP) for the photodynamic therapy of tumors was investigated. TPP was solubilized in small unilamellar lipid vesicles prepared by extrusion on a LIPOSOFAST apparatus. These samples were studied by laser-excited time resolved luminescence and triplet-triplet absorption spectroscopy. In this lipid environment TPP was still an efficient singlet oxygen producer, as indicated by the characteristic singlet oxygen phosphorescence at 1270 nm in D2O, when excited with a 28 ns laser pulse at 412 nm. Moreover, unlike with sulfonated TPP (TPPS4), liposomal TPP showed the reduced decay rates of TPP triplet-states with the increasing time of pre-illumination by a Xenon lamp. This was shown in an indirect way, based upon the appearance of a second component of the luminescence decay at 1270 nm in D2O; and by direct TPP triplet state monitoring, detecting triplet-triplet absorption at 440 nm in H2O. The deactivation of higher triplet states was delayed upon pre-illumination. This reflects an irreversible interaction of singlet oxygen with membrane lipids, thus demonstrating the potential of the liposomal form of TPP to efficiently disintegrate tumor cell membranes and to be a suitable preparation for the photodynamic therapy. PMID- 10517288 TI - Characterization of the K+ (Na+)/H+ monovalent cation exchanger in the human red blood cell membrane: effects of transport inhibitors. AB - The (ouabain + bumetanide + EGTA)-insensitive K+ influx (defined as residual K+ influx) in the human erythrocyte was investigated with respect to the characterization of the recently identified K+(Na+)/H+ exchanger (Richter et al. 1997). In particular, the effects of selected ion transport inhibitors on this flux in physiological ionic strength (high ionic strength, HIS) as well as low ionic strength (LIS) solutions were qstudied. The stimulation of the K+ influx observed in LIS medium was further enhanced when DIDS, phloretin, eosin-5 maleimide, furosemide, DIOA, NPPB, or DCDPC was present at a concentration of 0.1 mmol/l. This paradoxical, inhibitor-induced increase of the K+ influx was more pronounced in LIS media where chloride (7.5 mmol/l) was replaced by nitrate. For DNDS, niflumic acid, and MK-196 (0.1 mmol/l) an enhanced K+ transport could only be observed in nitrate-containing LIS solution. Bumetanide and purine riboside, at a concentration of 0.1 mmol/l, did not cause significant changes of the K+ influx in either chloride- or nitrate-containing LIS media. Dipyridamole and ruthenium red (0.1 mmol/l), which are positively charged, significantly reduced the K+ influx in both chloride- and nitrate-containing LIS media. In nitrate containing HIS solution only dipyridamole inhibited the K+ influx. The residual K+ influx in LIS solution was significantly increased by removing internal [Mg2+], and decreased by quinacrine (1 mmol/l). In HIS solution, no effect of altering intracellular Mg2+ occurred but a stimulation of the flux by quinacrine was observed. The results are discussed in terms of a more general surface charge effect of the used inhibitors on the K+(Na+)/H+ exchanger. PMID- 10517289 TI - Different effects of verapamil and low calcium on repetitive contractile activity of frog fatigue-resistant and easily-fatigued muscle fibres. AB - The effects of low calcium and verapamil on contractility of two muscle fibre types (m. iliofibularis, Rana temporaria) upon different stimulation protocols were been compared. Verapamil (0.02 mmol/l) induced temporal excitation contraction coupling failure during single tetanic stimulation and enhanced the decline of tetanic force during 30 s repetitive tetanic stimulation in both fatigue-resistant fibres and easily-fatigued fibres. In contrast to verapamil, low extracellular calcium (0.02 mmol/l) only enhanced the decline of tetanic force in fatigue-resistant during repetitive tetanic stimulation but had no effect on easily-fatigued fibres. The effect of verapamil on the decline of tetanic force in fatigue-resistant fibres was more profound in low calcium conditions. Both verapamil and low calcium eliminated twitch facilitation that appeared after prolonged contractile activity in fatigue-resistant fibres. 4mmol/l Ni+2, used as calcium channel antagonist, had effects similar to low calcium medium. It could be concluded that (i) extracellular Ca2+-requirements for excitation-contraction coupling are different in fatigue-resistant and easily fatigued fibres; (ii) the effects of verapamil on force performance are not entirely dependent upon calcium channel blockade. PMID- 10517290 TI - Passive forces in mammalian skeletal muscle: a freely-jointed and worm-like chain. AB - The passive mechanical properties of whole muscle in active and nonactive states are compared. The experimental results are presented as stress-strain curves, which are analyzed in the framework of the current theoretical background [viz. the freely-jointed chain model (FJCM) and the worm-like chain model (WLCM)] in a semi-quantitative fashion. This analysis shows that both models can explain the mechanical behavior of whole muscle in non-active state. In the active state, the presence of crossbridges alters the mechanical response, leading to a markedly different behavior, as expected. A discussion of the mechanisms involved and the interpretation of the parameters required for the fitting of the stress-strain curves is also presented. PMID- 10517291 TI - Changes in passive electric parameters of human erythrocyte membrane during hyperthermia: role of spectrin phosphorylation. AB - In prefixed by 1 mmol/l OsO4 human erythrocytes, the discocyte shape was preserved upon heating to temperatures which include the denaturation temperature of the main peripheral protein spectrin. Nevertheless, the suspension of fixed cells displayed threshold decrease in its capacitance and resistance at the temperature range where spectrin denaturates. The same changes were established using intact cells and their resealed ghosts. For packed cells (ghosts), the capacitance and resistance decreased about 17% (31%) and 30% (19%). These data indicate a decrease in the beta dispersion of erythrocyte membrane associated, according to a previous study (Ivanov 1997), with the heat denaturation of spectrin at 49.5 degrees C. The amplitude of the 49.5 degrees C decrease in beta dispersion was reversibly reduced in intact erythrocytes and white ghosts following reversible decrease in the phosphorylation of their membrane proteins. It was fully eliminated in ghosts following their resealing with alkaline phosphatase (0.1 mg/ml) which dephosphorylated membrane proteins. These findings are discussed in relation to similar changes found in normal and tumour tissues and cells during hyperthermia. PMID- 10517292 TI - Evaluation of 3-azidiamantane as photoaffinity probe of cytochrome P450. AB - 3-azidiamantane (DIA-N2) has been shown to be a photolabile carbene-generating probe interacting specifically with cytochrome P450 (P450) active centre. To evaluate the modification of P450 by the probe, radiolabelled [9-3H]-3 azidiamantane was prepared by reductive dehalogenation of its precursor, 3-oxo-9 bromodiamantane ethylene ketal. The synthesis was optimized as the proper precursor and reaction conditions were concerned to produce 96% pure product (overall yield 59%). An incorporation efficacy of the probe photoactivated at 366 nm was examined with two different proteins, BSA and rat phenobarbital-inducible P450 2B1, both having hydrophobic binding sites. Under photolysis the photoaffinity probe generated short-lived (> 90%) intermediates binding immediately to the protein. The yield of photoactivated DIA-N2 incorporation was 12% and 11% for BSA and P450, respectively. The presence of reduced glutathione, a scavenger of reactive intermediates, did not affect the probe incorporation markedly. On the other hand, scavengers entering the P450 active centre, methanol and dithiothreitol, reduced the protein labelling by 36% and 42%, respectively. Similarly, at DIA-N2, aminopyrine (substrates), and metyrapone (inhibitor) 50 times molar excess over the probe, prevented its binding by about 40%. In addition, when photoaffinity labelling was carried out with microsomal preparation, the substrate with a high affinity for the P450 2B1, diamantane, (at 20 times molar excess to the probe) caused 47% inhibition of the P450 covalent labelling. These results, suggesting a high specificity of the probe binding, show that it can be applied as a photoaffinity probe for cytochrome P450 2B1 active centre studies. PMID- 10517293 TI - Membrane potential as a modulator of the free intracellular Ca2+ concentration in agonist-activated endothelial cells. AB - We have used combined patch clamp and fura-2 fluorescence to elucidate the role of membrane potential in the regulation of the cytosolic Ca2+ concentration ([Ca2+]i) in a human umbilical vein derived endothelial cell-line, EA.hy926 (EA cells) stimulated with vasoactive agonists, such as ATP, histamine and bradykinin. This stimulation caused hyperpolarization and sustained Ca2+ plateau in nonclamped cells. Clamping agonist-stimulated cells at negative potentials enhanced the amplitude of this plateau, whereas it was smaller at more depolarized potentials, indicating that Ca2+ influx follows its driving force. Depolarization of the membrane by increasing extracellular K+ or by applying charybdotoxin, a blocker of big conductance Ca2+-dependent K+ channels during agonist stimulation diminished the plateau rise in [Ca2+]i. It is concluded that the membrane potential is an efficient regulator of Ca2+ influx during the plateau phase of agonist-mediated Ca2+ signals. In addition, the modulating effects on Ca2+ signals should be interpreted with caution if the membrane potential of the cells is not controlled. PMID- 10517294 TI - Hydrolysis-dependent absorption of disaccharides in the rat small intestine (chronic experiments and mathematical modeling). AB - In order to throw light on the mechanisms responsible for the enzyme-dependent absorption of disaccharides membrane hydrolysis of maltose and trehalose and the absorption of glucose (free and that derived from disaccharides) were studied in isolated loops (20 cm) of the rat small intestine in chronic experiments. The rates of glucose absorption were 0.26-0.81 micromol x min(-1) x cm(-1) when the loop was perfused with a 12.5 to 75.0 mmol/l free glucose solution, which is only insignificantly higher than the rates observed during perfusion with equivalent maltose solutions. The coupling coefficient (the ratio of glucose absorption rate to the rate of disaccharide hydrolysis) decreased from 0.90 to 0.60 with the increasing maltose concentrations in the infusate from 6.25 to 37.5 mmol/l, but remained unchanged (approximately 0.95) within the same range of trehalose concentrations. The permeability of the pre-epithelial barrier was equivalent to that of unstirred water layer of less than 40 microm thickness. Fluid absorption was within the range of 0.73-2.55 microl x min(-1) x cm(-1), and it showed a correlation with the rates of glucose absorption. The results agree with a model developed on the assumption that free glucose and that released from disaccharides share the same membrane transporters. It could be concluded that a close coupling of disaccharide hydrolysis with derived glucose absorption in chronic experiments is achieved mainly due to a high activity of glucose transporters, which are presumably not associated with membrane disaccharidases. The transcellular active transport is a predominant mechanism of disaccharide derived glucose absorption under conditions close to physiological. PMID- 10517295 TI - Hume, bioethics, and philosophy of medicine. PMID- 10517296 TI - Hume on the nonhuman animal. AB - Hume wrote about fundamental similarities and dissimilarities between human and nonhuman animals. His work was centered on the cognitive and emotional lives of animals, rather than their moral or legal standing, but his theories have implications for issues of moral standing. The historical background of these controversies reaches to ancient philosophy and to several prominent figures in early modern philosophy. Hume develops several of the themes in this literature. His underlying method is analogical argument and his conclusions are generally favorable regarding the abilities in animals. Hume does not attribute a moral sense or capacity of judgment to animals, but he does suggest that their actions exhibit moral qualities, such as other-regarding instincts. Hume allows in-kind differences in both demonstrative reason and moral judgment, but in the domains of both causal reason and moral agency he believes there are differences of degree rather than of kind. Hume's most significant philosophical contribution was to move as far as anyone before him to a naturalistic explanation of human and nonhuman minds that invited psychological and epistemological examination of minds by using the identical methods and categories for man and beast. PMID- 10517297 TI - Hume on suicide. AB - Anyone interested in the morality of suicide reads David Hume's essay on the subject even today. There are numerous reasons for this, but the central one is that it sets up the starting point for contemporary debate about the morality of suicide, namely, the debate about whether some condition of life could present one with a morally acceptable reason for autonomously deciding to end one's life. We shall only be able to have this debate if we think that at least some acts of suicide can be moral, and we shall only be able to think this if we give up the blanket condemnation of suicide that theology has put in place. I look at this strategy of argument in the context of the wider eighteenth-century attempt to develop a non-theologically based ethic. The result in Hume's case is a very modern tract on suicide, with voluntariness and autonomy to the fore and with reflection on the condition of one's life and one's desire to carry on living a life in that condition the motivating circumstance. PMID- 10517298 TI - Looking to Hume for justice: on the utility of Hume's view of justice for American health care reform. AB - This essay argues that Hume's theory of justice can be useful in framing a more persuasive case for universal access in health care. Theories of justice derived from a Rawlsian social contract tradition tend to make the conditions for deliberation on justice remote from the lives of most persons, while religiously inspired views require superhuman levels of benevolence. By contrast, Hume's theory derives justice from the prudent reflections of socially-encumbered selves. This provides a more accessible moral theory and a more realistic path to the establishment of universal access. PMID- 10517299 TI - Coming home to Hume: a sociobiological foundation for a concept of 'health' and morality. AB - Assessing the normative status of concepts of health and disease involves one in questions regarding the relationship between fact and value. Some have argued that Christopher Boorse's conception of health and disease lacks such a valuational element because it cannot account for types of harms which, while disvalued, do not have evolutionarily dysfunctional consequences. I take Boorse's account and incorporate some Humean-like sociobiological assumptions in order to respond to this challenge. The possession of moral sentiments, I argue, offers an evolutionary advantage (thus falling within Boorse's definition of normal functional abilities). However, this does not amount to emotivism: on the contrary, these sentiments can be the basis of a value system. This value structure introduces the concept of sympathizing with a fellow being's suffering as the basis of a normative dimension to disease. For example, it holds the disvalue of disease to lie in the fact that disease involves suffering and functional limitations. The naturalistic Humean type of account presented here thus jumps the normative-descriptive divide. When Boorse's account is extended to include social sentiments and behaviors, a conception of health emerges which is broader than Boorse's or Kass's, but narrower than the WHO's. PMID- 10517300 TI - Hume's influence on John Gregory and the history of medical ethics. AB - The concept of medicine as a profession in the English-language literature of medical ethics is of recent vintage, invented by the Scottish physician and medical ethicist, John Gregory (1724-1773). Gregory wrote the first secular, philosophical, clinical, and feminine medical ethics and bioethics in the English language and did so on the basis of Hume's principle of sympathy. This paper provides a brief account of Gregory's invention and the role that Humean sympathy plays in that invention, with reference to key texts in Gregory's work. The paper also considers two interesting and perhaps provocative ways in which Hume can be read through Gregory: first, sympathy as a principle of scientific discovery in Hume's science of man and moral physiology; and sympathy as gendered feminine in Hume's moral philosophy. Hume's principle of sympathy is at the core of Gregory's medical ethics and the histories of Western medical ethics and bioethics pivot on Gregory--and, therefore, on Hume--as it does on few other figures. PMID- 10517301 TI - Help from Hume reconciling professionalism and managed care. AB - Health care systems are widely criticized for limiting doctors' roles as patient advocates. Yet unrestricted advocacy can be unfairly partial, costly, and prejudicial. This essay considers three solutions to the problem of how to reconcile the demands of a just health care system for all patients, with the value of advocacy for some. Two views are considered and rejected, one supporting unlimited advocacy and another defending strict impartiality. A third view suggested by Hume's moral theory seeks to square the moral demands of professional advocacy and just health care systems. A moral basis for limited advocacy exists when it can be justified from a general or moral vantage. Consequently, ethical aspects of professionalism are not necessarily on a collision course with health care systems incorporating managed care. This solution is compatible with goals regarding the importance of humanistic education and professionalism to build patients' trust. PMID- 10517302 TI - Genetics of Type 1 diabetes mellitus. AB - Type 1 diabetes or insulin-dependent diabetes mellitus (IDDM) is the archetypal example of a T cell-mediated autoimmune disease characterised by selective destruction of a single cell type: the insulin-producing beta-cells of the pancreatic islets of Langerhans. The pathogenic equation for IDDM presents a complex interrelation of genetic and environmental factors, most of which have yet to be identified. Based on the observed familial aggregation of IDDM, it is certain that there is a decided heritable genetic susceptibility for developing autoimmune diabetes. The well-known association of IDDM with certain human histocompatibility leukocyte antigen (HLA) alleles of the major histocompatibility complex (MHC) was a major step toward understanding the role of inheritance in IDDM. Landmark molecular biological investigations of diabetes HLA susceptibility genes provided great potential for insights into the molecular basis for the autoimmune nature of the disease, beginning a story that continues to unfold. Although the association of certain HLA alleles with IDDM is very strong, this genetic locus is estimated to account for less than 50% of genetic contributions to disease susceptibility. The search for non-HLA susceptibility genes has received great attention in recent years. Albeit genome wide searches are wrought with controversy, such studies have suggested the association of numerous non-MHC loci with Type 1 diabetes that will require careful follow-up investigation. Cell biological and genetic functional analyses will provide clues that are indispensable for further progress. The necessary studies include research on immunological abnormalities that are present many years before the clinical onset of Type 1 diabetes. PMID- 10517303 TI - Thyroid autoimmunity in children and adolescents with Type 1 diabetes mellitus. AB - Type 1 diabetic children and adolescents often present with autoimmune thyroid disorders. Two hundred and four diabetic patients, less than 20 years old, were studied in order to diagnose these diseases. The prevalence of thyroid autoimmune disorders was 17.6% and, of those, chronic autoimmune thyroiditis was the most frequent. Microsomal autoantibodies correlated more accurately with the presence of chronic autoimmune thyroiditis than thyroglobulin autoantibodies. The thyroid status of most of the patients with positive markers was euthyroidism (77%), but subclinical hypothyroidism (11%), overt hypothyroidism (3%), subclinical hyperthyroidism (3%) and overt hyperthyroidism (6%) were also present. Autoimmune thyroid disorders were the most prevalent immunological processes affecting diabetic patients. No significant associations of thyroid autoimmunity and other autoimmunological disorders, such as celiac disease or presence of other autoimmune antibodies, were found. PMID- 10517304 TI - Maternal effect of Type 2 diabetes mellitus on insulin sensitivity and metabolic profile in healthy young Mexicans. AB - The objective of this study was to identify the maternal effect of Type 2 diabetes mellitus (T2DM) on insulin sensitivity and metabolic profiles of young healthy, Mexican people (n=15). A cross-sectional study was performed in these subjects with a family history of T2DM in both first and second degree relatives on the maternal side and 15 control subjects. The following tests were carried out: insulin tolerance test and metabolic profile. Systolic blood pressure and serum uric acid were significantly higher in probands than controls. Insulin sensitivity was not different between either group studied (4.7+/-0.9 in probands vs 4.5+/-0.8%/min in controls; p=0.52). IN CONCLUSION: family history of T2DM on the maternal side increased both systolic blood pressure and serum uric acid level in probands, without modification in their insulin sensitivity. Triglycerides concentration had a tendency to be higher in probands than controls. PMID- 10517305 TI - Relationship between acute insulin response and vitamin K intake in healthy young male volunteers. AB - To evaluate the effects of vitamin K (VK) on pancreatic function, especially on acute insulin response, 25 healthy young male volunteers were given an oral load of 75 g of glucose, and their mean daily VK intake was estimated by a one-week food check list. After excluding low (<20) and high (> or =25) body mass index (BMI) subjects, the remaining 16 participants were divided into three semi-equal groups according to VK intake. Blood VK status of the low VK intake group tended to be poorer than that of the high intake group (median of 5 samples: prothrombin time; 12.5 vs 12.2s and protein-induced VK absence-factor-II; 23 vs 15 mAU/ml), but fasting plasma glucose status was not markedly different between both groups: [plasma glucose (PG); 87 vs 86 mg/dl, immunoreactive insulin (IRI); 6.7 vs 5.3 microU/ml, HbA1c; 4.8 vs 4.9%]. However, at 30 min after glucose loading, PG of the low VK intake group tended to be higher than those of the high intake group (160 vs 145 mg/dl) and IRI was lower (36.1 vs 52.3 microU/ml). Insulinogenic index (incremental IRI/incremental PG, 0-30 min) of the low VK intake group was significantly lower than that of the high intake group (0.4 vs 0.9). These results suggested that VK may play an important role on the acute insulin response in glucose tolerance. PMID- 10517306 TI - Hyperglycaemia: the bridge between non-enzymatic glycation and oxidative stress in the pathogenesis of diabetic complications. AB - It is generally accepted that high glucose levels for many years are a primary cause of most long-term complications in diabetic patients. Many studies suggest that the central features of diabetic complications are caused by the hyperglycaemia-accelerated formation of non-enzymatic glycated products. Non enzymatic glycation, however, has been recently demonstrated to be linked to glucose auto-oxidative process. At the same time glycated proteins have been shown to be a source of free radicals. These findings raised the hypothesis of a link between oxidative stress and the development of diabetic complications. Some studies have recently demonstrated that antioxidants, such as vitamin C and E, may reduce in vitro and in vivo protein glycation. At the same time some antioxidants act as scavengers of the free radicals produced by non-enzymatic glycation in vitro. Such studies may lead to therapeutic approaches for limiting the damage from glycation and oxidation reactions and for complementing existing therapy for treatment of the complications of diabetes. PMID- 10517307 TI - Fourth Consensus Conference of the European Committee on Hyperbaric Medicine. London, December 4-5, 1998. Hyperbaric oxygen in the management of foot lesions in diabetic patients. PMID- 10517308 TI - Carotid intima media thickness in patients with newly diagnosed T2DM and impaired glucose tolerance. PMID- 10517309 TI - Isolation and partial characterization of an antiviral, RC-183, from the edible mushroom Rozites caperata. AB - A protein of 10,425 Da was purified from the edible mushroom Rozites caperata and shown to inhibit herpes simplex virus types 1 and 2 replication with an IC50 value of < or = 5 microM. The protein designated RC-183 also significantly reduced the severity of HSV-1 induced ocular disease in a murine model of keratitis, indicating in vivo efficacy. HSV mutants lacking ribonucleotide reductase and thymidine kinase were also inhibited, suggesting the mechanism does not involve these viral enzymes. Antiviral activity was also seen against varicella zoster virus, influenza A virus, and respiratory syncytial virus, but not against adenovirus type VI, coxsackie viruses A9 and B5, or human immunodeficiency virus. Characterization of RC-183 by mass spectroscopy, sequencing, and other methods suggests it is composed of a peptide (12 or 13 mer) coupled to ubiquitin via an isopeptide bond between the c-terminal glycine of ubiquitin and the epsilon amino group of a lysine residue in the peptide. The peptide sequence did not match any known sequence. Thus, RC-183 is a novel antiviral that may have clinical utility or serve as a lead compound for further development. Determining the mechanism of action may lead to identification of novel steps in viral replication. PMID- 10517310 TI - Antiviral properties of isoborneol, a potent inhibitor of herpes simplex virus type 1. AB - Isoborneol, a monoterpene and a component of several plant essential oils, showed dual viricidal activity against herpes simplex virus 1 (HSV-1). First, it inactivated HSV-1 by almost 4 log10 values within 30 min of exposure, and second, isoborneol at a concentration of 0.06% completely inhibited viral replication, without affecting viral adsorption. Isoborneol did not exhibit significant cytotoxicity at concentrations ranging between 0.016% and 0.08% when tested against human and monkey cell lines. Isoborneol specifically inhibited glycosylation of viral polypeptides based on the following data: (1) the mature fully glycosylated forms of two viral glycoproteins gB and gD were not detected when the virus was replicated in the presence of isoborneol, (2) no major changes were observed in the glycosylation pattern of cellular polypeptides between untreated and isoborneol treated Vero cells, (3) isoborneol did not affect the glycosylation of gB produced from a copy of the gB gene resident in the cellular genome, and (4) other monoterpenes such as 1,8-cineole and borneol, a stereoisomer of isoborneol, did not inhibit HSV-1 glycosylation. PMID- 10517311 TI - Antiherpetic activity and mode of action of natural carrageenans of diverse structural types. AB - The lambda-carrageenan 1T1, the kappa/iota-carrageenan 1C1 and the mu/nu-type 1C3, isolated from the red seaweed Gigartina skottsbergii, proved to be potent and selective inhibitors of herpes simplex virus (HSV) types 1 and 2. The antiviral IC50 values determined by virus yield inhibition assay in different cell lines ranged from 0.4 to 3.3 microg/ml, and no cytotoxic effects, measured by trypan blue exclusion on stationary or proliferating cells, tetrazolium salt method or cell protein synthesis, were observed. Time of addition and attachment studies suggested that the main target for antiviral action of the three carrageenans was virus adsorption, whereas no effect on virus internalization, or early or late protein synthesis was detected. However, the lambda-carrageenan 1T1 was still significantly inhibitory when added any time after adsorption. The pretreatment of virions with the carrageenans showed that 1C1 and 1C3 lacked direct inactivating effect at concentrations near the antiviral IC50 but 1T1 exerted virucidal action. The cyclization of 1T1 to afford the derivative 1T1T1 maintained the antiviral activity but eliminated the virucidal properties. Thus, the structure of 1T1 seems to be responsible for its differential behavior from 1C1 and 1C3, probably allowing a more stable binding to HSV, leading to virion inactivation. In contrast, 1C1 and 1C3 fail to bind with high affinity to virus alone, but are able to interfere with the interaction between HSV particles and the cell. PMID- 10517312 TI - Antiviral effects of 28-deacetylsendanin on herpes simplex virus-1 replication. AB - The compound purified from the fruit of Melia azedarach exerted an antiviral effect on herpes simplex virus-1 (HSV-1) in Vero cells. It was identified as 28 deacetylsendanin (28-DAS). The 50% inhibitory concentration (IC50) of 28-DAS was 1.46 microg/ml without cytotoxicity at 400 microg/ml on Vero cells. Electron microscopy showed that low electron-dense cores of newly synthesized nucleocapsids remained in swollen nuclei and no extracellular virus particles were observed at 15 h p.i. Consistent with this result, it was confirmed by a plaque assay that few infectious progeny viruses were released from the 28-DAS treated virus-infected cells at 24 h p.i. Intracellular viruses in 28-DAS-treated virus-infected cells were 23% of untreated and infected cells. The synthesis of thymidine kinase (TK) was reduced by 28-DAS at early stage. In conclusion, 28-DAS inhibited the replication of HSV-1, reduced the synthesis of HSV-1 TK, and led to the formation of defective nucleocapsids. PMID- 10517314 TI - Inhibition of HIV-1 infection by zinc group metal compounds. AB - Thirty-seven metal compounds were examined for inhibitory activities against infection with human immunodeficiency virus type 1 (HIV-1). Zinc group metal compounds, namely, zinc acetate, zinc chloride, zinc nitrate, cadmium acetate and mercury chloride, showed anti-HIV-1 activities. Cadmium and mercury compounds at 1-10 microg/ml and zinc compounds at 100 microg/ml strongly inhibited HIV-1 infection, although the cadmium, mercury and zinc compounds had severe cytotoxities at 100, 100 and 1000 microg/ml, respectively. They inhibited transcription of HIV-1 RNA and HIV-1 production at concentrations at which they did not affect the growth of HIV-1-producing cells. They had little effect on syncytium formation resulting from cocultivation of uninfected with HIV-1 producing cells. Nor did they affect HIV-1 DNA synthesis following HIV-1 infection. The metal compounds may owe their anti-HIV-1 effects to inhibition of HIV-1 DNA to RNA transcription, rather than inhibition of the adsorption, penetration or reverse transcription step of HIV-1 infection. PMID- 10517313 TI - Interferon/antioxidant combination therapy for chronic hepatitis C--a controlled pilot trial. AB - The effects of two forms of antioxidative co-therapy were analyzed in 24 interferon-alpha (IFN)-naive patients with chronic hepatitis C who were randomized to either receive IFN monotherapy (3 x 4.5 million units IFN-alpha 2a per week), (group A), or IFN and N-acetylcysteine (N-acetylcysteine (NAC) 1.800 mg/day) plus sodium selenite (400 microg/day) supplementation (group B), or treatment as in group B plus vitamin E (544 IU/day) (group C), over 24 weeks. Changes in histology, normalization of ALT, reduction of viral RNA, serum levels of glutathione, selenium, vitamin E, erythrocyte glutathione peroxidase, trolox equivalent antioxidative capacity (TEAC), thiobarbituric acid reactive substances (TBARS) and protein carbonyl groups were measured. Low baseline TEAC and elevated TBARS indicated increased oxidative stress before therapy, which was not affected by antioxidant supplementation. At the end of treatment complete responses were found in 3/8, 2/8 and 6/8 patients in groups A, B and C, respectively, but liver histology had not significantly improved. Vitamin E treated patients had a 2.4 greater chance (95% CI: 1.05-5.5) of obtaining a complete response and had significantly greater reduction in viral load (P = 0.028) than patients without vitamin E. Relapses, i.e. re-appearance of detectable hepatitis C virus (HCV) RNA and/or re-elevation of ALT-activity occurred in 7 out of the 11 responders within 6 months after termination of therapy (group A: 2/3, group B: 1/2 and group C: 4/6). Thus, no overall beneficial effect of antioxidant/IFN therapy was detected. However, the apparent trend towards a more favorable outcome with vitamin E supplementation warrants to further study this substance as an adjuvant to IFN therapy in chronic hepatitis C. PMID- 10517315 TI - Identification of psi3Tom20, a novel processed pseudogene of the human Tom20 gene, and complete characterization of psi1Tom20 and psi2Tom20. AB - We report the identification and characterization of psi3Tom20, a novel processed pseudogene of the human Tom20 (hTom20) gene, which is 96.2% similarity with the hTom20 cDNA and is 5' and 3' truncated. In addition, we present the complete characterization of psi2Tom20 and psi2Tom20, the two other recently reported members of this pseudogene family. Comparison of the sequences of psi3Tom20 with that of the previously reported psi2Tom20 revealed and corrected an error in the previously determined sequence of psi2Tom20. A detailed analysis of these three pseudogenes, including their flanking regions, is presented. It suggests they probably arose from mRNAs that were polyadenylated at different sites. Possible mechanisms involved in their integration as retroposons are also discussed. PMID- 10517316 TI - Developmental regulation of a cytosolic ascorbate peroxidase gene from tomato plants. AB - This report describes the characterisation of a gene (APX20) from tomato plants that encodes a cytosolic isoform of ascorbate peroxidase which is involved in the detoxification of intracellular H2O2. Expression analysis of promoter-GUS fusions in transgenic plants reveals that the gene is under strict developmental control, becoming transcriptionally active in cells which are apparently undergoing mechanical stimulation generated during the different phases of growth. We show that the APX20 gene contains a large 5' leader intron which is required to confer constitutive gene expression in leaves, but not in other organs of the plant. Based on these observations, we propose that the observed transcriptional regulation of this gene may constitute a basic mechanism for deployment of antioxidative defences in plants, which act to limit the deleterious effects of H2O2 generated during normal plant development. PMID- 10517317 TI - Identification, characterization and chromosomal localization of the cognate human and murine DBF4 genes. AB - The kinase Dbf4p/Cdc7p is required for the G1/S phase transition during the cell cycle and plays a direct role in the activation of individual origins of replication in Saccharomyces cerevisiae. Here, we report the identification and characterization of mouse and human cDNAs whose products are related in sequence to Saccharomyces cerevisiae DBF4 cDNA. Both mammalian Dbf4 proteins contain a putative site for phosphorylation by CDK, PEST protease cleavage sites, nuclear localization signals and a short-looped zinc finger-like domain. Transcription of MmDBF4 is suppressed in mouse NIH3T3 fibroblasts made quiescent by serum starvation. Upon replenishment of the medium, transcript levels increase during progression through G1, peaking as cells enter S phase. MmDbf4p interacts physically with Cdc7p and Mcm2p in vivo. Using fluorescence in situ hybridization (FISH), the human DBF4 gene was localized to chromosome 7 (q21.3), whereas FISH mapped the murine counterpart to band A2 on chromosome 5. The results of chromosome mapping indicate that in both mouse and human the gene is present as a single copy. The structural conservation between Dbf4-related proteins suggests that these proteins play a key role in the regulation of DNA replication during the cell cycle in all eukaryotes. PMID- 10517318 TI - Molecular characterization of mitotic cyclins in rice plants. AB - Cyclins are known to activate cyclin-dependent protein kinases, which are essential for cell cycle progression in eukaryotes. We isolated full-length cDNAs encoding rice mitotic cyclins named CycA1; os; 1 and CycB2;os;1, which are related to A- and B-type cyclins, respectively, from animals. To characterize the function of these mitotic cyclins, as well as that of another B-type cyclin, CycB2;os;2, each cDNA was introduced into yeast cells. When cDNAs encoding CycA1;os;1, CycB2;os; or CycB2;os;2 were overexpressed in the yeast mutant DLI, which is deficient in G1 cyclins, the mutant phenotype was rescued, indicating that these mitotic cyclins are functional in yeast cells. When the cDNA encoding CycB2;os;1 was expressed in the wild-type yeast strain, the cells lost the ability to grow, whereas the expression of either cycA1;os: 1 or cycB2;os;2 did not inhibit growth. In situ hybridization of these mitotic cyclin genes with rice root apices and counterstaining of chromosomes with a DNA-specific dye revealed that cycA1;os;1 is expressed from the G2 phase to the early M phase, while transcripts of cycB2:os;1 and cycB2;os;2 accumulated until the end of mitosis. Our results indicate that these B2-type cyclins may be involved in the control of mitosis, in combination with a G2/M-phase CDK. PMID- 10517319 TI - Four mismatch repair paralogues coexist in Arabidopsis thaliana: AtMSH2, AtMSH3, AtMSH6-1 and AtMSH6-2. AB - By using degenerate oligonucleotides based on the sequence homology between known MutS homologues, three MSH cDNAs belonging to the MSH2, MSH3 and MSH6 families, as defined in eukaryotes, have been isolated from Arabhidopsis thaliana (ecotype Columbia). Genomic sequences for two of these genes (AtMSH2 and AtMSH6-2) were also isolated and determined, whereas the genomic sequence of AtMSH3 was obtained through the Arabidopsis sequencing project, as was the sequence of a second, distinct AtMSH6 homologue (AtMSH6-1). Comparative analysis of the AtMSH2 Landsberg erecta genomic sequence (reported here) and the previously described AtMSH2 Columbia allele revealed several polymorphisms, including the presence of a small, transposon-like element in the 3' untranscribed region of the former allele. Arabidopsis is the first organism to show such divergence of two AtMSH6 genes; the divergence is strongly supported by sequence data and phylogenetic analysis. Southern analysis revealed that the three genes we have isolated exist as single copies, and genetic mapping indicated that AtMSH2 and AtMSH6-2 both reside on chromosome III. Finally, expression of these three genes could only be observed in suspensions of A. thaliana cells. Such a cell suspension divides actively after subculture, and the AtMSH genes are most strongly expressed at this stage. PMID- 10517320 TI - The transcriptional activator CorR is involved in biosynthesis of the phytotoxin coronatine and binds to the cmaABT promoter region in a temperature-dependent manner. AB - A modified two-component regulatory system consisting of the histidine protein kinase CorS and two highly homologous response regulators, CorR and CorP, controls biosynthesis of the polyketide phytotoxin coronatine (COR) by Pseudomonas syringae pv. glycinea PG4180 in a temperature-dependent manner. COR synthesis is maximal at 18 degrees C but does not occur at 28 degrees C. Fusions of CorR and CorP to the maltose-binding protein (MBP) were overproduced in Escherichia coli and P. syringae PG4180, and tested for functionality by complementation of corR and corP mutants of PG4180, respectively. The cmaABT promoter region was defined by deletion mapping, and the DNA-binding capability of CorR and CorP was examined by gel retardation assays. When overproduced in P. syringae at 18 degrees C and purified, MBP-CorR was shown to bind specifically to a 218-bp DNA fragment corresponding to positions -841 to -623 bp upstream of the transcriptional start site of the cmaABT operon. In contrast, MBP-CorP and MBP itself, when overproduced in P. syringae and E. coli at 18 degrees C and 28 degrees C, respectively, did not bind to the 218-bp fragment or to any other DNA fragment analyzed. The CorP protein lacks typical DNA-binding motifs, suggesting that it might modulate the function of CorR. However, addition of purified MBP CorP did not alter the DNA-binding activity of MBP-CorR. On the other hand, this activity was completely abolished when MBPCorR was overproduced at 28 degrees C or in a corS mutant, indicating that the binding of CorR depended on the growth temperature at which it was produced and was controlled by CorS. In addition, overproduction of MBP-CorR in a corP mutant of PG4180 also yielded inactive protein, underlining the importance of CorP for CorR activation. We propose that CorR is activated by CorS at low temperature and that CorP is required for this activation before CorR can bind to DNA. PMID- 10517322 TI - Cis requirements for transposition of Tc1-like transposons in C. elegans. AB - The Caenorhabditis elegans transposons Tc1 and Tc3 are able to transpose in heterologous systems such as human cell lines and zebrafish. Because these transposons might be useful vectors for transgenesis and mutagenesis of diverse species, we determined the minimal cis requirements for transposition. Deletion mapping of the transposon ends shows that fewer than 100 bp are sufficient for transposition of Tc3. Unlike Tc1, Tc3 has a second, internal transposase binding site at each transposon end. We found that these binding sites play no major role in the transposition reaction, since they can be deleted without reduction of the transposition frequency. Site-directed mutagenesis was performed on the conserved terminal base pairs at the Tc3 ends. The four terminal base pairs at the ends of the Tc3 inverted repeats were shown to be required for efficient transposition. Finally, increasing the length of the transposon from 1.9 kb to 12.5 kb reduced the transposition frequency by 20-fold, both in vivo and in vitro. PMID- 10517321 TI - A Nod factor-binding lectin is a member of a distinct class of apyrases that may be unique to the legumes. AB - Recent studies from our laboratory have found that a root lectin from the legume Dolichos hifloris is present on the root surface, binds rhizobial Nod factor and has apyrase activity. To assess the broader significance of this lectin/nucleotide phosphohydrolase (Db-LNP), we have cloned a second related cDNA (Db-apyrase-2) from D. hiflorus, as well as related cDNAs from the legumes Lotus japonicus and Medicago sativa, and from Arabidopsis thaliana, a non-legume. The deduced amino acid sequences of these apyrases were aligned with one another and with the sequences of other apyrases from plants, animals, yeast and protozoa. Phylogenetic analysis shows that Db-LNP has closely related orthologs only in other legumes, while Db-apyrase-2 is more closely related to apyrase sequences from non-leguminous plants. We also show that the orthologs of Db-LNP from M. sativa and Pisum sativum have carbohydrate binding activity. The results suggest that legume LNPs may represent a special class of apyrases that arose by gene duplication and subsequent specialization. PMID- 10517323 TI - Regulated nuclear localisation of the yeast transcription factor Ace2p controls expression of chitinase (CTS1) in Saccharomyces cerevisiae. AB - The yeast transcription factor Ace2p regulates expression of the chitinase gene CTS1 in a cell cycle-dependent manner. Nuclear localisation of Ace2p is restricted to late M and early G phases of the mitotic cell cycle. We show here that this nuclear localisation is directly associated with regulation of CTS1 expression. Using a version of Ace2p tagged with a c-myc epitope, we show that the protein is excluded from the nucleus of cells during most phases of the mitotic cell cycle. A mutant derivative in which one threonine and two serine residues, which are candidate phosphorylation sites, were replaced by alanine (to mimic constitutive dephosphorylation) is localised in the nucleus throughout the cell cycle. The mechanism of localisation of Ace2p therefore involves regulation of its phosphorylation state, and closely resembles that used by the homologous transcription factor Swi5p. The wild-type Ace2 protein associates with Cdc28p in vivo, suggesting this may be the kinase that mediates the phosphorylation event. The stability of the protein is greatly reduced in a mutant that is constitutively localised to the nucleus, but is restored in a deletion derivative which remains in the cytoplasm. Ace2p is therefore controlled throughout the cell cycle at three levels: transcription, nuclear localisation, and proteolysis. PMID- 10517324 TI - Alterations in organization and transcription of the mitochondrial genome of cytoplasmic male sterile sugar beet (Beta vulgaris L.). AB - We have constructed a physical map of the mitochondrial DNA of a cytoplasmic male sterile (CMS) sugar beet line, TK81-MS, and compared it with that published for normal fertile sugar beet (cv. TK81-O) to clarify the differences between the CMS and normal mitochondrial genomes. The TK81-MS genome is present as a single circular molecule of 481.8 kb, or as two molecules of 184.9 and 296.9 kb. The CMS genome was found to be highly rearranged relative to the normal mitochondrial genome, with at least fifteen rearrangement and/or inversion events being required to align the two DNAs. Analysis of transcription patterns of known mitochondrial genes and rearranged regions revealed six genes, coxI, coxII, atpA, atp6, rps3, and orf324, whose expression is altered in the CMS line relative to the normal line. Of these six, only the coxI transcript pattern differs between male-sterile and fertility-restored genotypes, making it likely that the coxI locus is involved in mediating CMS in sugar beet. PMID- 10517325 TI - An artificial enhancer with multiple response elements stimulates prokaryotic transcriptional activation medicated by various regulatory proteins. AB - Regulation of transcription by the form of RNA polymerase that contains sigma(N) involves activation at a distance by activators bound to sites located far upstream of the transcription start site, which contact RNA polymerase bound to the promoter via formation of a DNA loop. At the g/nAp2 promoter, binding sites for the activator NtrC show features characteristic of eukaryotic enhancers. A multiple response element containing binding sites for five sigma(N)-dependent activators from different systems has been cloned in different positions relative to the glnAp2 promoter. These promoter regions indeed allowed activation in vivo by each regulator, thus showing that transcription from an eubacterial promoter may be controlled in a very versatile way by different signals. The activation capability of each activator has been assessed in relation to its concentration, and the presence and relative positions of the corresponding binding sites in the DNA. Results show that most activators can function from any position. However, activation mediated by DctD-L64 was very sensitive to changes in the position of its binding sites. Transcriptional activation by combinations of two regulators was also tested and no significant synergism or interference was detected. Mapping of the 5' ends of the transcripts showed that neither the activator nor the position from which they activate influences selection of the transcription start site. PMID- 10517326 TI - The SOS response is induced by replication fork blockage at a Ter site located on a pUC-derived plasmid: dependence on the distance between ori and Ter sites. AB - A new model system for the study of the SOS response has been developed. In this system the response is induced by blocking the replication fork at a Ter site located in pUC-derived plasmids. Blockage of the fork is dependent on the expression of the Ter binding protein, Tus, encoded on another plasmid, in which the tus gene is under the control of the ara promoter. SOS induction can, therefore, be controlled by arabinose. The extent of the SOS response was monitored by measuring the activity of beta-galactosidase, expressed from a lacZ gene fused to the 5' region of the sfiA gene, a typical SOS-responsive gene. Expression of the fusion gene is completely dependent on recA+ and lexA+ genes. Using this system, we found that the distance between the ori and Ter sites is directly correlated with the strength of SOS induction. The properties of this system are discussed. PMID- 10517327 TI - Heavy metal-mediated activation of the rat Cu/Zn superoxide dismutase gene via a metal-responsive element. AB - The Cu/Zn superoxide dismutase (SODI) catalyzes the dismutation of superoxide radicals produced in the course of biological oxidations. When placed under the control of the rat SOD1 gene promoter and transfected into human HepG2 hepatoma cells, the activity of a chloramphenicol acetyltransferase reporter gene was found to increase three- to four-fold in the presence of heavy metals (cadmium, zinc and copper). Functional analysis of mutant derivatives of the SOD1 gene promoter and the use of a heterologous promoter system confirmed that the induction of the SOD1 gene by metal ions requires a metal-responsive element (MRE) located between positions -273 and -267 (GCGCGCA). It was also shown by gel mobility shift assays that an MRE binding protein is induced by the exposure of the human liver cell line HepG2 to heavy metals. These results suggest that the MRE participates in the induction of the SOD1 gene by heavy metals. PMID- 10517328 TI - Involvement of protein kinase C in the response of Neurospora crassa to blue light. AB - As a first step towards understanding the process of blue light perception, and the signal transduction mechanisms involved, in Neurospora crassa we have used a pharmacological approach to screen a wide range of second messengers and chemical compounds known to interfere with the activity of well-known signal transducing molecules in vivo. We tested the influence of these compounds on the induction of the al-3 gene, a key step in light-induced carotenoid biosynthesis. This approach has implicated protein kinase C (PKC) as a component of the light transduction machinery. The conclusion is based on the effects of specific inhibitors (calphostin C and chelerythrine chloride) and activators of PKC (1,2-dihexanoyl sn-glycerol). During vegetative growth PKC may be responsible for desensitization to light because inhibitors of the enzyme cause an increase in the total amount of mRNA transcribed after illumination. PKC is therefore proposed here to be an important regulator of transduction of the blue light signal, and may act through modification of the protein White Collar-1, which we show to be a substrate for PKC in N. crassa. PMID- 10517329 TI - Interactive regulatory pathways control virulence determinant production and stability in response to environmental conditions in Staphylococcus aureus. AB - The accessory gene regulator (agr) and staphylococcal accessory regulator (sar) loci are important regulators of toxin production in Staphylococcus aureus. In this study we examined how environmental conditions degree of aeration and salt concentration - affect the transcription and translation of mRNAs for alpha haemolysin (Hla) and serine protease (Ssp) via these pathways and influence the stability of these proteins. Using Northern analysis, we have confirmed earlier observations that sarA is involved in the upregulation of RNAIII, the effector molecule encoded by the agr locus. However, this effect was abolished in highly aerated cultures. While sarA does appear to have an up-regulatory effect on hla transcription that is independent of agr, we propose that the PC1839 (sarA) mutant produces less alpha-haemolysin activity mainly as a result of post translational inactivation by proteases. The most obvious phenotypic feature of PC1839 (sarA) is the upregulation of proteases. In this study we show that ssp is repressed by SarA at the transcriptional level. Western analysis using an anti alpha-haemolysin antibody identified a major breakdown product that is only present in the supernatant of strains that are overexpressing serine protease. We have also confirmed that agr exerts a significant regulatory influence on hla at the level of translation, as well as transcription. Finally, the addition of salt upregulates ssp transcription and dramatically downregulates transcription of hla, and is an example of an environmental parameter that affects toxin production independently of agr and sarA. How environmental signals are transduced to control alpha-haemolysin and serine protease production, activity and stability at multiple levels are discussed. PMID- 10517330 TI - The Saccharomyces cerevisiae LEP1/SAC3 gene is associated with leucine transport. AB - Leucine uptake by Saccharomyces cerevisiae is mediated by three transport systems, the general amino acid transport system (GAP), encoded by GAP1, and two group-specific systems (S1 and S2), which also transport isoleucine and valine. A new mutant defective in both group-specific transport activities was isolated by employing a gap1 leu4 strain and selecting for trifluoroleucine-resistant mutants which also showed greatly reduced ability to utilize L-leucine as sole nitrogen source and very low levels of [14C]L-leucine uptake. A multicopy plasmid containing a DNA fragment which complemented the leucine transport defect was isolated by selecting for transformants that grew normally on minimal medium containing leucine as nitrogen source and subsequently assaying [14C]L-leucine uptake. Transformation of one such mutant, lep1, restored sensitivity to trifluoroleucine. The complementing gene, designated LEP1, was subcloned and sequenced. The LEP1 ORF encodes a large protein that lacks characteristics of a transporter or permease (i.e., lacks hydrophobic domains necessary for membrane association). Instead, Lep1p is a very basic protein (pI of 9.2) that contains a putative bipartite signal sequence for targeting to the nucleus, suggesting that it might be a DNA-binding protein. A database search revealed that LEP1 encodes a polypeptide that is identical to Sac3p except for an N-terminal truncation. The original identification of SAC3 was based on the isolation of a mutant allele, sac3-1, that suppresses the temperature-sensitive growth defect of an actin mutant containing the allele act1-1. Sac3p has been previously shown to be localized in the nucleus. When a lep1 mutant was crossed with a sac3 deletion mutant, no complementation was observed, indicating that the two mutations are functionally allelic. PMID- 10517332 TI - Regulation of the expression of the cold shock proteins CspB and CspC in Bacillus subtilis. AB - The small acidic proteins CspB and CspC are the major cold shock-induced proteins of Bacillus subtilis. Analysis of mRNA revealed a transient four-fold increase in the transcription level of both genes during cold shock. The cspB and cspC mRNAs are dramatically stabilised after a temperature downshift from 37 degrees C to 15 degrees C. The data in this study support the idea that the expression of CspB and CspC in B. subtilis during cold shock is regulated mainly at the post transcriptional level, as is also the case with CspA in Escherichia coli. PMID- 10517331 TI - The Escherichia coli heat shock protease HtrA participates in defense against oxidative stress. AB - The serine protease HtrA (DegP), which is indispensable for cell survival at elevated temperatures, is a peripheral membrane protein, localized on the periplasmic side of the inner membrane in Escherichia coli, and the biochemical and genetic evidence indicates that the physiological role of HtrA is to degrade denatured proteins formed in the cellular envelope during heat shock. The aim of this study was to find out if the HtrA protease contributes to protection of the cell against oxidative stress. We compared the influence of various oxidizing agents on htrA mutant cells with their effects on wild-type bacteria, and found that the htrA mutation did not increase sensitivity to hydrogen peroxide or paraquat but made the cell extremely sensitive to ferrous [Fe(II)] ions, which are known to enhance oxidation of proteins. Treatment with ferrous ions caused a larger increase in the level of protein carbonyl groups in the membrane fraction of the cell than in the periplasm and cytoplasm. Iron-induced oxidation of membrane proteins was enhanced in the htrA mutant relative to wild-type cells. Inhibition of the growth of the htrA mutant by iron could be alleviated more efficiently by a nitroxide antioxidant that localizes in the membranes (A-TEMPO) than by a derivative (40H-TEMPO) that acts mainly in the soluble fraction of the cell. Inhibition of the growth of the htrA mutant was more pronounced following treatment with cumene hydroperoxide, which partitions into membranes, than with t butyl hydroperoxide, which forms radical mainly in the cytosol. Both ferrous ions and cumene hydroperoxide, but not hydrogen peroxide, paraquat or t-butyl hydroperoxide, induced synthesis of HtrA. Our results show that HtrA plays a role in defense against oxidative shock and support the hypothesis that HtrA participates in the degradation of oxidatively damaged proteins localized in the cell envelope, especially those associated with the membranes. PMID- 10517333 TI - Integrative recombination of Lactobacillus delbrueckii bacteriophage mv4: functional analysis of the reaction and structure of the attP site. AB - The integrase of the temperate bacteriophage mv4 catalyzes site-specific recombination between the phage attP site and the attB site of the host during lysogenization of Lactobaccillus delbrueckii subsp. bulgaricus. The mv4 integrase also functions in a wide variety of gram-positive bacteria and in Escherichia coli. In this report, in vitro and in vivo recombination assays were developed and the integrase was purified in order to study in greater detail the mv4 attP x attB recombination event. In a cell-free extract of E. coli at 42 degrees C, the mv4 integrase promotes efficient in vitro recombination between a supercoiled attP-containing plasmid and a linear attB fragment. The integrase, which was purified to apparent homogeneity, showed no absolute requirement for accessory factors, unlike the majority of the lambda Int family of recombinases. Deletion derivatives of the attP site were constructed and tested for recombination with the attB site in vitro. A 234-bp DNA fragment containing five scattered putative mv4 Int-binding sites was sufficient for function of the attP site. In contrast to the right arm of attP, most of the left arm could be deleted without drastically reducing the recombination efficiency. In vivo in E. coli, mv4 Int catalyzed recombination in trains between attP and attB sites present on two separate plasmids. This property was used to confirm in vivo the results of the deletion analysis of the attP site performed in vitro. PMID- 10517334 TI - Imbalance in dosage of the genes for the heterochromatin components Sir3p and histone H4 results in changes in the length and sequence organization of yeast telomeres. AB - Telomeric heterochromatin plays an essential role in telomere function, including the regulation of telomere length. We observe that in Saccharomyces cerevisiae an imbalance in the dosage of genes for two protein components of heterochromatin (namely Sir3p and histone H4) causes modifications in telomere length and telomere sequence organization. The effects of Sir3p/H4 imbalance were analyzed in yeast strains in which the wild-type SIR3 gene (normally a single-copy gene) was either absent or present in 20-30 copies, and both histone H4 genes (HHF1 and HHF2) were present or HHF1 was deleted, thus covering a wide range of viable gene dosage combinations. Modifications of telomeres and of subtelomeric regions were identified by analyzing both the overall telomere population and by focusing on two single telomeric regions: the left telomere of chromosome III (LIII) and the right telomere of chromosome XI (RXI). The modifications induced by alteration of the Sir3p/H4 ratio consist of a reduction in the length and an increase in the instability of the terminal block of (C(1-3)A)n repeats and in susceptibility to insertion of Y' elements into this repeat element. Restoration of the wild-type gene ratio (by removal of the extra copies of SIR3 or by complementation with the missing second copy of HHF) restored the original telomere organization, both with respect to the length of the (C(1-3)A)n repeat stretch and the absence of Y' elements. This behavior shows that the stability of the wild-type sequence organization requires maintenance of the normal structure of telomeric heterochromatin. PMID- 10517335 TI - Isolation and characterization of pyrimidine auxotrophs, and molecular cloning of the pyrE gene from the hyperthermophilic archaeon Pyrococcus abyssi. AB - Uracil auxotrophic mutants of the hyperthermophilic archaeon Pyrococcus abyssi were isolated by screening for resistance to 5-fluoro-orotic acid (5-FOA). Wild type strains were unable to grow on medium containing 5-FOA, whereas mutants grew normally. Enzymatic assays of extracts from wild-type P. abyssi and from pyrimidine auxotrophs demonstrated that the mutants are deficient in orotate phosphoribosyltransferase (PyrE) and/or orotidine-5'-monophosphate decarboxylase (PyrF) activity. The pyrE gene of wild-type P. abyssi and one of its mutant derivatives were cloned and sequenced. This pyrE gene could serve as selectable marker for the development of gene manipulation systems in archaeal hyperthermophiles. PMID- 10517336 TI - A novel gene required for cercosporin toxin resistance in the fungus Cercospora nicotianae. AB - Cercosporin, a photosensitizing perylenequinone toxin produced by the plant pathogenic Cercospora fungi, generates the highly toxic singlet oxygen (1O2) upon exposure to light. Cercosporin shows broad toxicity against a wide range of organisms, including bacteria, fungi, plants, and animals; however, Cercospora fungi are resistant to its effects. A novel gene, crg1 (cercosporin-resistance gene) was isolated from a wild-type strain of C. nicotianae by genetic complementation of a C. nicotianae mutant (CS10) which is cercosporin sensitive and down-regulated in cercosporin production. Sequence analysis indicated that crg1 encodes a putative protein of 550 amino acids with four putative transmembrane helical regions, however CRG1 shows no strong similarity to any other protein in sequence databases. Northern analysis identified two transcripts (4.5 and 2.6 kb) that are unaffected by the presence of light or cercosporin. Southern analysis demonstrated that crg1 is present in a single copy in the C. nicotianae genome and can be detected only in Cercospora species. Targeted disruption of crg1 resulted in mutants that, like CS10, are sensitive to cercosporin. However, unlike CS10, crg1 disruption mutants are not down-regulated in toxin production. Both CS10 and the crg1 disruption mutants are unaffected in their response to other 1O2-generating photosensitizers, suggesting that CRG1 functions specifically against cercosporin, rather than against 1O2. PMID- 10517337 TI - Cell type-specific gene expression in the cell cycle of the dimorphic ciliate Eufolliculina uhligi. AB - The life cycle of the unicellular eukaryote Eufolliculina uhligi includes two structurally and physiologically different cell types: a motile swarmer that is arrested in the cell cycle, and a sessile cell (trophont) that feeds and reproduces. These two cell types offer an exceptionally favourable system for the isolation of genes involved in cell cycle regulation and cellular morphogenesis. Differential screening of a trophont cDNA library using a swarmer-subtracted, trophont-specific probe yielded eleven clones that represent trophont-specific transcripts and one clone that represents a swarmer-specific transcript. Sequence analysis showed that seven clones, including the only swarmer-specific one, represent unknown genes, whereas five clones could be identified by sequence comparisons. Two of the clones appear to encode proteins that are involved in the regulation of growth and metabolism. The deduced sequences of three clones resemble potential cell cycle regulators. Data are presented on a putative member of the calcium/calmodulin-dependent protein kinase family and on a TIP120-like sequence, which is the first such sequence to be described since the discovery of the rat TIP120 protein. Furthermore, a unique new sequence is presented, whose features suggest that it represents a protein that is involved in the regulation of cell division. It includes domains characteristic of two different protein families, cyclin-dependent kinases (CDKs) and cyclins, both of which are known to be cell cycle regulators. Based on our results we propose a model for cell cycle regulation in ciliated protozoa. PMID- 10517338 TI - Insulin-like growth factor (IGF)-I rescues breast cancer cells from chemotherapy induced cell death--proliferative and anti-apoptotic effects. AB - Insulin-like growth factor (IGF)-I protects many cell types from apoptosis. As a result, it is possible that IGF-I-responsive cancer cells may be resistant to apoptosis-inducing chemotherapies. Therefore, we examined the effects of IGF-I on paclitaxel and doxorubicin-induced apoptosis in the IGF-I-responsive breast cancer cell line MCF-7. Both drugs caused DNA laddering in a dose-dependent fashion, and IGF-I reduced the formation of ladders. We next examined the effects of IGF-I and estradiol on cell survival following drug treatment in monolayer culture. IGF-I, but not estradiol, increased survival of MCF-7 cells in the presence of either drug. Cell cycle progression and counting of trypan-blue stained cells showed that IGF-I was inducing proliferation in paclitaxel-treated but not doxorubicin-treated cells. However, IGF-I decreased the fraction of apoptotic cells in doxorubicin- but not paclitaxel-treated cells. Recent work has shown that mitogen-activated protein kinase (MAPK) and phosphotidylinositol-3 (PI 3) kinase are activated by IGF-I in these cells. PI-3 kinase activation has been linked to anti-apoptotic functions while MAPK activation is associated with proliferation. We found that IGF-I rescue of doxorubicin-induced apoptosis required PI-3 kinase but not MAPK function, suggesting that IGF-I inhibited apoptosis. In contrast, IGF-I rescue of paclitaxel-induced apoptosis required both PI-3 kinase and MAPK, suggesting that IGF-I-mediated protection was due to enhancement of proliferation. Therefore, IGF-I attenuated the response of breast cancer cells to doxorubicin and paclitaxel by at least two mechanisms: induction of proliferation and inhibition of apoptosis. Thus, inhibition of IGF-I action could be a useful adjuvant to cytotoxic chemotherapy in breast cancer. PMID- 10517339 TI - Suppression of parathyroid hormone-related protein messenger RNA expression by medroxyprogesterone acetate in breast cancer tissues. AB - The level of parathyroid hormone-related protein (PTHrP) expressed in breast cancer tissue is closely related to the incidence of bone metastasis. We examined the PTHrP mRNA expression in breast cancer tissues by coamplification polymerase chain reaction (PCR) in mole ratio to internal standard beta-actin mRNA. The PTHrP expression was higher in premenopausal patients than in postmenopausal patients (P < 0.05). More pronounced difference by menopause found in estrogen receptor (ER) positive groups (P < 0.001) indicated that the PTHrP expression in breast cancer tissue is hormonally regulated and might be altered by endocrine agents. To clarify the changes of PTHrP expression by endocrine therapy of breast cancer, we measured PTHrP expression in the breast cancer tissue incubated for 24 h with 1 x 10(-8) M of estradiol (E2), 1 x 10(-6) M of tamoxifen (TAM) and 1 x 10(-5) M of medroxyprogesterone acetate (MPA). The PTHrP expression was decreased significantly by MPA (P < 0.005), while E2 and TAM did not change the PTHrP expression. Progesterone receptor (PgR) mRNA expression was also examined to confirm that the breast cancer tissue responds to E2 and TAM. The results were well compatible with the better therapeutic effect of MPA reported for the treatment of breast cancer with bone metastases. As a potential candidate for the receptor that mediates the suppressive effect of MPA, androgen receptor (AR) is suggested most probable. Present results also demonstrated that the clinical response of individual tumors is closely associated with the early in vitro changes of gene expression detected in the cancer specimen. PMID- 10517340 TI - Vorozole results in greater oestrogen suppression than formestane in postmenopausal women and when added to goserelin in premenopausal women with advanced breast cancer. AB - The high potency and selectivity of new aromatase inhibitors has translated to greater efficacy and improved tolerability in comparison with established second line hormonal agents for advanced breast cancer in phase III clinical trials. Two pharmacological studies are reported which assess the use of one of these inhibitors, vorozole, in combination or comparison with well-established methods of oestrogen deprivation in pre and postmenopausal patients. When combined with the gonadotrophin-releasing hormone agonist (GnRHa) goserelin in 10 premenopausal patients, vorozole markedly enhanced the suppression of serum levels of oestrone, oestradiol and, oestrone sulphate beyond that achieved by goserelin alone (by a mean 74%, 83%, and 89%, respectively). The combination was well-tolerated and had no significant effects on androgen levels. Vorozole was compared with formestane in 13 postmenopausal women and serum oestrone, oestradiol, and oestrone sulphate levels were suppressed by 47%, 30%, and 70%, respectively, more by vorozole than by the steroidal aromatase inhibitor. Again the tolerability was excellent. The plasma oestrogen levels in the postmenopausal patients on vorozole were lower than in the premenopausal patients on goserelin plus vorozole, indicating that ovarian oestrogen synthesis may be relatively resistant to aromatase inhibition, even during GnRHa treatment. Thus, in both pre and postmenopausal patients substantially greater suppression of oestrogen can be achieved by vorozole compared with alternative approaches. Existing clinical-pharmacological correlates suggest that these increases in pharmacological effectiveness may result in enhanced clinical effectiveness. PMID- 10517341 TI - Hypermethylation of the Wilms' tumor suppressor gene CpG island in human breast carcinomas. AB - CpG island hypermethylation is known to be associated with transcriptional silencing of tumor suppressor genes in neoplasia. We have previously detected aberrantly methylated sites in the first intron of the Wilms' tumor suppressor (WT1) gene in breast cancer. In the present study, we extended the investigation to a CpG island located in the promoter and first exon regions of WT1. Methylation of this CpG island was found to be extensive in MCF-7 and MDA-MB-231 breast cancer cells, as well as in 25% (five of 20 patients) of primary breast tumors. While levels of the known 3.0-kb WT1 mRNAs were decreased or not detected in these cell lines, the expression could be partially restored following treatment with a demethylation agent, 5-aza-2'-deoxycytidine. Surprisingly, a novel 2.5-kb WT1 transcript was expressed at high levels in both untreated and treated MDA-MB-231 cells. This novel transcript was likely a WT1 variant missing the first exon, and therefore escaped the methylation control present in the normal transcript. Our study implicates the future need to investigate the significance of this aberrant transcript as well as the role of WT1 CpG island hypermethylation in breast neoplasia. PMID- 10517342 TI - Quality of life in the first year after breast cancer surgery: rehabilitation needs and patterns of recovery. AB - BACKGROUND: Although mortality rates from breast cancer are declining, many breast cancer survivors will experience physical and psychological sequelae that affect their everyday lives. Few prospective studies have examined the rehabilitation needs of newly diagnosed breast cancer patients, and little is known about the predictors of health-related quality of life (QOL) in this population. METHODS: Between 1987 and 1990, 227 women with early stage breast cancer participated in a prospective longitudinal study in which detailed information was collected through interviews, standardized measures of QOL and psychological distress, and clinical evaluation. Comparisons of physical and treatment-related problems were made according to type of surgical treatment. Multivariate regression analysis was performed to examine the predictors of QOL at one year after surgery. RESULTS: Physical and treatment-related problems were reported frequently one month after breast cancer surgery, and occurred with equal frequency in women receiving modified radical mastectomy or breast conservation treatment. There were no significant differences in problems reported at one year by type of surgery; however, frequently reported problems include 'numbness in the chest wall or axilla,' 'tightness, pulling or stretching in the arm or axilla,' 'less energy or fatigue,' 'difficulty in sleeping,' and 'hot flashes'. There was no relationship between the type of surgery and mood or QOL. Poorer QOL one year after surgery was significantly associated with greater mood disturbance and body image discomfort one month after surgery, as well as positive lymph node involvement. Although the majority of patients experienced substantial disruptions in the physical and psychosocial dimensions of QOL post operatively, most women recovered during the year after surgery, with only a minority (<10%) significantly worsening during that time. CONCLUSIONS: At one year after surgery, most women report high levels of functioning and QOL, with no relationship between the type of surgery and QOL. Women who reported lower levels of QOL at one year after diagnosis had greater mood disturbance and poorer body image one month after surgery, as well as lower income and positive axillary nodes. PMID- 10517343 TI - Rural breast cancer treatment: evidence from the Reaching Communities for Cancer Care (REACH) project. AB - BACKGROUND: Research shows that rural populations are more likely than their urban counterparts to be diagnosed with late-stage cancer, but less is known about appropriateness of cancer treatment in rural locations after diagnosis. The objective of this analysis was to assess the degree to which rural breast cancer treatment was received in concordance with national recommendations. METHODS: Data came from 251 stage I and II breast cancer patients residing in rural North Carolina. State-of-the-art care was defined using the National Cancer Institute's (NCI) physician data query (PDQ) database, and cases were categorized into appropriate primary and/or adjuvant treatment. Chi-square and Fishers' exact tests were used to assess changes in appropriate treatment over time (1991-1996) and between stage. Multiple logistic regression was used to determine whether any patient or disease characteristics were associated with receipt of appropriate treatment. RESULTS: Most (81-90%) of the breast cancer cases received the appropriate primary therapy (mastectomy or lumpectomy followed by radiation therapy); of these, the majority received a mastectomy (66-72%). Fewer women received adjuvant therapy as recommended (27-61%), although significantly more stage II than stage I cases did so (p < or = 0.05). Regression showed that stage and estrogen-receptor (ER) status were associated with appropriate therapy. CONCLUSIONS: The findings suggest that there exist deviations from NCI established treatment recommendations among rural breast cancer patients. More research is needed to develop better methods for dissemination of state-of-the art cancer information to rural physicians and patients, and to understand how treatment decisions are made. PMID- 10517344 TI - Prognostic factors predicting survival from first recurrence in patients with metastatic breast cancer: analysis of 439 patients. AB - We have analyzed retrospectively 439 women with recurrent breast cancer, followed at a single institution, in order to define potential prognostic factors for survival at the time of first recurrence. Median age at the time of first recurrence was 58 and the median disease free interval (DFI) from initial diagnoses to recurrence was 33 months. Thirteen percent of the patients did not receive any adjuvant therapy while 87% received different combinations of chemotherapy, radiotherapy and hormone therapy as adjuvant treatment. With a median follow-up of 44 months from the time of recurrence the median survival (MSR) was 24 months (SE 1.24) and five-year overall survival was 18% (SE 2.02). On the univariate analysis, pathological tumor size (pT) at diagnosis (p < 0.0006), axillary lymph node status at diagnosis (p < 0.00001), negative estrogen receptor (ER) status (p < 0.0001), negative progesterone receptor (PgR) status (p < 0.0001), adjuvant chemotherapy (p < 0.001), disease free interval (p < 0.00001), location of recurrence (p < 0.0002) and number of metastatic sites (> or = 3: p < 0.0003), were significantly associated with shorter survival from first relapse. On the multivariate analysis, only the site of recurrence, axillary lymph node status at diagnosis, ER status and DFI remained independently associated with decreased MSR after first relapse. PMID- 10517345 TI - Schedule-dependent interactions between vinorelbine and paclitaxel in human carcinoma cell lines in vitro. AB - Vinorelbine and paclitaxel are new anticancer agents that bind to distinct sites on tubulin and affect microtubules in opposite ways. Clinical studies of combinations of these agents have been in progress against breast cancer and some solid tumors. To clarify the optimal schedule for this combination, we studied the schedule-dependent cytotoxic effects of vinorelbine and paclitaxel against the human lung carcinoma cell line A549, the breast carcinoma cell line MCF7, the ovarian carcinoma cell line PA1, and the colon carcinoma cell line WiDr in vitro. Tumor cells were incubated with vinorelbine and paclitaxel simultaneously for both 24 h and 5 days. Cells were also incubated with vinorelbine for 24 h, followed by a 24-h exposure to paclitaxel and vice versa. Cell growth inhibition after 5 days was determined by MTT assay. The effects of drug combinations at the concentration producing 80% cell growth inhibition (IC80) were analyzed by the isobologram method (Steel and Peckham). The simultaneous exposures to vinorelbine and paclitaxel for both 24 h and 5 days produced additive effects for all four cell lines. The sequential exposure to vinorelbine followed by paclitaxel produced additive effects for the PA1 and WiDr cells, additive and antagonistic effects for the A549 cells, and antagonistic effects for the MCF7 cells. The sequential exposure to paclitaxel followed by vinorelbine produced additive effects for the A549, and PA1 cells, additive and antagonistic effects for the MCF7 cells, and antagonistic effects for the WiDr cells. Our findings suggest that the simultaneous rather than the sequential administration of vinorelbine and paclitaxel may be the optimal schedule for this combination of these two agents. Applications of this schedule dependency may be beneficial for the treatment of breast cancer and other solid tumors. PMID- 10517346 TI - Association of in vitro invasiveness and gene expression of estrogen receptor, progesterone receptor, pS2 and plasminogen activator inhibitor-1 in human breast cancer cell lines. AB - The invasive potential of tumor cells is usually tested either by in vitro invasion assays which evaluate cell spreading ability in basement membrane-like matrices or by in vivo invasion assays in nude mice. Both methods are laborious and time-consuming. Tumor invasiveness is accompanied by the changes in expression of various genes. The invasive behavior of cells is therefore represented by certain gene expression patterns. The purpose of this study was to investigate whether expression patterns of several genes are characteristic for the invasiveness of cultured cells. We examined the mRNA levels of estrogen receptor (ER), progesterone receptor (PR), estrogen inducible pS2 and plasminogen activator inhibitor-1 (PAI-1) in 23 cell lines derived from benign and malignant breast tissues using a competitive reverse transcription-polymerase chain reaction (cRT-PCR) system. We also evaluated the invasiveness of these cell lines by their ability to penetrate into a collagen-fibroblast matrix. We demonstrate that the gene expression pattern of breast cell lines is clearly associated with their in vitro invasiveness. In general, cells with ER, PR, pS2 but no PAI-1 expression showed a non-invasive phenotype, while cells expressing PAI-1 mRNA but not ER mRNA are invasive. Our study indicates that the invasiveness of breast cancer cell lines is characterized by PAI-1 gene expression and the lack of ER mRNA. This suggests that PAI-1 may participate in the invasive process. PMID- 10517347 TI - Monitoring of olanzapine in serum by liquid chromatography-atmospheric pressure chemical ionization mass spectrometry. AB - A selective assay of olanzapine with liquid chromatography atmospheric pressure chemical ionization (LC-APCI-MS, positive ions) is described. The drug and internal standard (ethyl derivative of olanzapine) were isolated from serum using a solid-phase extraction procedure (C18 cartridges). The separation was performed on ODS column in acetonitrile-50 mM ammonium formate buffer, pH 3.0 (25:75). After analysis of mass spectra taken in full scan mode, a selected-ion monitoring detection (SIM) was applied with the following ions: m/z 313 and 256 for olanzapine and m/z 327 and 270 for the internal standard for quantitation. The limit of quantitation was 1 microg/l, the absolute recovery was above 80% at concentration level of 10 to 100 microg/l. The method tested linear in the range from 1 to 1000 microg/l and was applied for therapeutic monitoring of olanzapine in the serum of patients receiving (Zyprexa) and in one case of olanzapine overdose. Olanzapine in frozen serum samples and in frozen extracts was stable over at least four weeks. The examinations of urine extracts from patients receiving olanzapine revealed peaks of postulated metabolites (glucuronide and N desmethylolanzapine). PMID- 10517348 TI - Use of liquid chromatography-mass spectrometry coupling for monitoring the serralysin-catalyzed hydrolysis of a peptide library. AB - The use of a peptide library of limited size, is considered to be more appropriate for studying a protease with a complex specificity, but very sensitive and efficient analytical techniques must be used. We have designed and synthesized a 49-peptide library of the type Z-AlaXXAla(amide) (X=Ala, Leu, Val, Phe, Ser, Arg, Glu) for studying the Pseudomonas aeruginosa serralysin specificity. All compounds of the peptide library could be identified by a LC-MS procedure. After hydrolysis of the library by pseudomonal serralysin, the LC-MS procedure also allowed the identification of the hydrolysis products and the different cleavage sites of the substrates. PMID- 10517349 TI - Development of a gradient elution high-performance liquid chromatography assay with ultraviolet detection for the determination in plasma of the anticancer peptide [Arg6, D-Trp7,9, mePhe8]-substance P (6-11) (antagonist G), its major metabolites and a C-terminal pyrene-labelled conjugate. AB - [Arg6, D-Trp7,9 mePhe8]-substance P (6-11), code-named antagonist G, is a novel peptide currently undergoing early clinical trials as an anticancer drug. A sensitive, high efficiency high-performance liquid chromatography (HPLC) method is described for the determination in human plasma of antagonist G and its three major metabolites, deamidated-G (M1), G-minus Met11 (M2) and G[Met11(O)] (M3). Gradient elution was employed using 40 mM ammonium acetate in 0.15% trifluoroacetic acid as buffer A and acetonitrile as solvent B, with a linear gradient increasing from 30 to 100% B over 15 min, together with a microbore analytical column (microBondapak C18, 30 cm X 2 mm I.D.). Detection was by UV at 280 nm and the column was maintained at 40 degrees C. Retention times varied by <1% throughout the day and were as follows: G, 13.0 min; M1, 12.2 min; M2, 11.2 min; M3, 10.8 min, and 18.1 min for a pyrene conjugate of G (G-P). The limit of detection on column (LOD) was 2.5 ng for antagonist G, M1-3 and G-P and the limit of quantitation (LOQ) was 20 ng/ml for G and 100 ng/ml for M1-3. Sample clean-up by solid-phase extraction using C2-bonded 40 microm silica particles (Bond Elut, 1 ml reservoirs) resulted in elimination of interference from plasma constituents. Within-day and between-day precision and accuracy over a broad range of concentrations (100 ng/ml-100 microg/ml) normally varied by < 10%, although at the highest concentrations of M1 and M2 studied (50 microg/ml), increased variability and reduced recovery were observed. The new assay will aid in the clinical development of antagonist G. PMID- 10517350 TI - Analysis of cyclophosphamide and five metabolites from human plasma using liquid chromatography-mass spectrometry and gas chromatography-nitrogen-phosphorus detection. AB - An assay method for the quantification of cyclophosphamide (CY) and five metabolites from human plasma is presented. The procedure is adapted to the chemical properties of the compounds of interest: non-polar compounds are extracted into methylene chloride, concentrated and analyzed by GC-NPD after derivatization, and the remaining aqueous fraction is deproteinated with acetonitrile-methanol prior to separation via reversed-phase HPLC and detection using atmospheric pressure ionization (API)-MS. Standard curves are linear over the required range and reproducible over five months. Plasma concentration-time profiles of CY and metabolites from a patient receiving CY by intravenous infusion (60 mg/kg, once a day for two days) are presented. PMID- 10517351 TI - Identification and confirmation of 3-hydroxy metabolite of ketoprofen in camels by gas chromatography-mass spectrometry and nuclear magnetic resonance spectroscopy. AB - The metabolites of ketoprofen were investigated in five camels following intravenous administration of a dose of 2.0 mg/kg body weight. Two metabolites were identified. The first one was purified with thin-layer chromatography. It was identified by gas chromatography-mass spectrometry (GC-MS) in comparison with authenticated reference standard and was found to be hydroxyketoprofen due to reduction of the ketone group of ketoprofen. The second metabolite was purified by high-performance liquid chromatography. It was identified with GC-MS and nuclear magnetic resonance spectroscopy as 3-hydroxybenzolketoprofen resulting from oxidation of the aromatic ring. PMID- 10517352 TI - Reversed-phase high-performance liquid chromatographic investigation of urinary normal and modified nucleosides of cancer patients. AB - Post-transcriptional modifications in RNA give rise to free modified ribonucleosides circulating in the blood stream and excreted in urine. Due to their abnormal levels in conjunction with several tumor diseases, they have been suggested as possible tumor markers. The developed RP-HPLC method has been applied to analyze the urinary nucleosides in 34 urinary samples from 15 kinds of cancer patients. The statistical analyses showed the urinary nucleoside excretion, especially modified nucleoside levels, in cancer patients were significantly higher than those in normal healthy volunteers. Factor analysis was used to classify the patients with cancer and normal healthy humans. It was found that using 15 urinary nucleoside levels or only five modified nucleoside levels as data vectors the factor analysis plot displayed two almost separate clusters representing each group. PMID- 10517353 TI - Analysis of 2-(3-methyl-4-aminophenyl)-benzothiazole (NSC 674495) in plasma by gas chromatography with mass-selective detection. AB - Certain naturally occurring isoflavonoids have been shown to inhibit protein tyrosine kinases, and this has led to investigations of ring-modified structural analogs. Most recently, 2-(3-methyl-4-aminophenyl)-benzothiazole (MAB: NSC 674495) was shown to possess significant activity against certain breast cell cancer lines in vitro and in vivo. Our efforts thus focussed on developing a simple and sensitive method for quantitating MAB in plasma using GC-MS. The GC-MS assay was found to be linear over the range of 0.050 to 5.0 microg/ml, and was applied to monitor the plasma concentration of MAB in a rat dosed with 25 mg/kg as a 1 min intravenous infusion. Plasma was collected at intervals from 3 through 180 min, and concentrations of MAB were determined. Non-linear regression analysis of the plasma concentration-time data revealed that levels declined from a maximum at 3 min of 18 microg/ml to 1 microg/ml at 3 h in a biphasic manner. In another investigation, significant plasma concentrations of a major metabolite was detected and determined to be mono-N-acetylated MAB. PMID- 10517354 TI - Simple and rapid assay for acetaminophen and conjugated metabolites in low-volume serum samples. AB - The use of marker compounds for estimating drug metabolic capacity or pharmacokinetic parameters is common in the biological sciences. Often small laboratory animals are used and thus sample size is a limiting concern. In this report, we describe an assay we developed for measuring the concentration of acetaminophen and its conjugated metabolites in low-volume serum samples. Acetaminophen and metabolites were removed from 10 microl serum samples by a single-step 6% (v/v) perchloric acid deproteination using theophylline as internal standard. Samples were separated in a pH 2.2 sodium sulfate-acetonitrile mobile phase at a flow-rate of 1.5 ml/min on a 15 cm octadecylsilyl column at room temperature. Analytes were detected at a wavelength of 254 nm. The resulting chromatograms showed no interfering peaks from endogenous serum components. The concentration ranges measured were 1.56-200 microg/ml for acetaminophen and acetaminophen sulfate and 3.91-500 microg/ml for acetaminophen glucuronide. The assay was linear in the range of concentrations analyzed. The intra-day and inter day coefficient of variation ranged from 0.4 to 8.2% and 0.2 to 12.3% for acetaminophen, 0.5 to 12.9% and 0.3 to 16.1% for acetaminophen glucuronide, and 0.4 to 8.1% and 0.2 to 14.3% for acetaminophen sulfate, respectively. Results from the experiments show that acetaminophen and its conjugated metabolites can easily and reproducibly be measured in low-volume serum samples and thus may offer an additional method to measure these compounds when the volume of biological samples may be limited. PMID- 10517355 TI - Determination of betulinic acid in mouse blood, tumor and tissue homogenates by liquid chromatography-electrospray mass spectrometry. AB - A rapid and sensitive high-performance liquid chromatography-electrospray MS method has been developed to determine tissue distribution of betulinic acid in mice. The method involved deproteinization of these samples with 2.5 volumes (v/w) of acetonitrile-ethanol (1:1) and then 5 microl aliquots of the supernatant were injected onto a C18 reversed-phase column coupled with an electrospray MS system. The mobile phase employed isocratic elution with 80% acetonitrile for 10 min; the flow-rate was 0.7 ml/min. The column effluent was analyzed by selected ion monitoring for the negative pseudo-molecular ion of betulinic acid [M-H]- at m/z 455. The limit of detection for betulinic acid in biological samples by this method was approximately 1.4 pg and the coefficients of variation of the assay (intra- and inter-day) were generally low (below 9.1%). When athymic mice bearing human melanoma were treated with betulinic acid (500 mg/kg, i.p.), distribution was as follows: tumor, 452.2 +/- 261.2 microg/g; liver, 233.9 +/- 80.3 microg/g; lung, 74.8 +/- 63.7 microg/g; kidney, 95.8 +/- 122.8 microg/g; blood, 1.8 +/- 0.5 microg/ml. No interference was noted due to endogenous substances. These methods of analysis should be of value in future studies related to the development and characterization of betulinic acid. PMID- 10517356 TI - High-performance liquid chromatography analysis, preliminary pharmacokinetics, metabolism and renal excretion of methylprednisolone with its C6 and C20 hydroxy metabolites in multiple sclerosis patients receiving high-dose pulse therapy. AB - A gradient eluent HPLC analysis in human plasma and urine was developed and validated for methylprednisolone (MP), its prodrug methylprednisolone-21 hemisuccinate (MPS) with the metabolites 6beta-hydroxy-6alpha-methylprednisolone (MPA), 20-hydroxymethylprednisolone (MPC), 6beta-hydroxy-20alpha hydroxymethylprednisolone (MPB), 6beta-hydroxy-20beta-hydroxymethylprednisolone (MPE), 20-carboxymethylprednisolone (MPD), methylprednisolone-glucuronide (MPF) and 21-carboxymethylprednisolone (MPX). The column was Cp Spherisorb C8 5 microm, 250 mm x 4.6 mm I.D. (Chrompack, Bergen op Zoom, The Netherlands) with a guard column 75 mm x 2.1 mm, packed with pellicular reversed-phase. The eluent was a mixture of acetonitrile and 0.067 M KH2PO4 buffer, pH 4.5. At t=O, the eluent consisted of 2% acetonitrile and 98% buffer (v/v). Over the following 35 min the eluent changed linearly until it attained a composition of 50% acetonitrile and 50% buffer (v/v). At 37 min (t=37) the eluent was changed over 5 min to the initial composition, followed by equilibration over 3 min. The flow-rate was 1.5 ml/min and UV detection was achieved at 248 nm. Preliminary pharmacokinetic data were obtained from one patient who showed illustrative plasma concentration-time curves and renal excretion-time profiles after a short-lasting infusion (0.5 h) of 1 g of methylprednisolone hemisuccinate. The half-life of prodrug methylprednisolone-21-hemisuccinate (MPS) was 0.3 h, that of metabolite MPX (21 carboxy MP) was 0.4 h and that of the parent drug methylprednisolone (MP) was 1.4 h. The half-lives of the metabolites are almost similar (4 h). The main compounds in the urine are methylprednisolone hemisuccinate (prodrug, 15.0%), methylprednisolone (parent drug, 14.6%), metabolite MPD (20-carboxy, 11.7%), and metabolite MPB (13.2%). The renal clearance values of metabolites MPB, MPC and MPD are approximately 500 ml/min, that of MP is 100 ml/min. PMID- 10517357 TI - Application of the high-performance liquid chromatographic method for separation, purification and characterisation of p-bromophenylacetylurea and its metabolites. AB - This study presents a HPLC method for the separation and purification of p bromophenylacetylurea (BPAU) and its metabolites. The method effectively separated and purified BPAU and its metabolites. Three metabolites of BPAU, M1, M2 and M3 were characterised by mass spectroscopy and nuclear magnetic resonance. They are named as N'-hydroxy-p-bromophenylacetylurea, 4-(4-bromophenyl)-3 oxapyrrolidine-2,5-dione and N'-methyl-p-bromophenylacetylurea, respectively. The major metabolic pathways of BPAU were proposed. The establishment of the HPLC method and characterisation of BPAU metabolites make it possible for further pharmacokinetic studies to explore the mechanism of BPAU-induced delayed neuropathy. PMID- 10517358 TI - Column-switching high-performance liquid chromatographic assay for determination of asiaticoside in rat plasma and bile with ultraviolet absorbance detection. AB - A column-switching high-performance liquid chromatography (HPLC) method is described for the determination of asiaticoside in rat plasma and bile using column-switching and ultraviolet (UV) absorbance detection. Plasma was simply deproteinated with acetonitrile prior to injection and bile was directly injected onto the HPLC system consisting of a clean-up column, a concentrating column, and an analytical column, which were connected with two six-port switching valves. Detection of asiaticoside was accurate and repeatable, with a limit of quantification of 0.125 microg/ml in plasma and 1 microg/ml in bile. The calibration curves were linear in a concentration range of 0.125-2.5 microg/ml and 1-20 microg/ml for asiaticoside in rat plasma and bile, respectively. This method has been successfully applied to determine the level of asiaticoside in rat plasma and bile samples from pharmacokinetics and biliary excretion studies. PMID- 10517359 TI - Optimization of intercalation dye concentration for short tandem repeat allele genotyping using capillary electrophoresis with laser-induced fluorescence detection. AB - DNA analysis using capillary electrophoresis (CE) with laser-induced fluorescence (LIF) detection requires that polymerase chain reaction products either be prepared using primers with fluorescent molecules covalently bonded to them, or stained with a fluorescent intercalation dye following amplification. The intercalation technique has the advantage of allowing fluorescence detection of any double-stranded DNA (dsDNA) product regardless of the amplification primers used. The increased sensitivity of LIF detection is sometimes compromised by the intercalation dye changing the mass to charge ratio of the DNA. The purpose of this study was to evaluate the changes of migration rate, resolution and fluorescent intensity of dye-DNA complexes during electrophoretic separations, and to establish the optimal parameters for short tandem repeats alleles profiling. The alleles of three STR loci THO1, F13A01 and vWFA31 were intercalated with the monomeric dyes TOPRO-1 and YOPRO-1, and their corresponding dimers, TOTO-1 and YOYO-1 (Molecular Probes, Eugene, OR, USA). Alleles intercalated before injection onto the CE column resulted in loss of resolution and sensitivity when compared to the on-column labeling technique. The results of this experimentation were then applied to a STR typing assay using a commercially available polymer and buffer matrix. This assay included development of a unique internal standard used for migration time normalization assignment of alleles. Consequently, the 9 allele and the 9.3 microvariant of the THOI locus were separated and typed correctly with a resolution of 0.49 in less than 20 min, and the only sample preparation necessary after amplification was a dilution step. PMID- 10517360 TI - Importance of high-performance liquid chromatographic analysis of serum N acylneuraminic acids in evaluating surgical treatment in patients with early endometrial cancer. AB - The objectives of this study were the quantification of the two major sialic acid (Sia) forms - N-acetylneuraminic (Neu5Ac) and N-glycolylneuraminic acids (Neu5Gc) - in serum before and after surgical treatment of early endometrial cancer and the relation of their levels with the progress of surgical therapy. The major Sia forms were liberated from sera glycoconjugates by mild acid hydrolysis, separated as per-O-benzoylated derivatives by a highly sensitive reversed-phase HPLC method and detected at 231 nm. Total Sia content in sera of healthy women was not related to age and body weight. Neu5Ac was identified as the major Sia in sera from both cancer patients, healthy individuals as well as in tissue specimens (> or = 94% of total Sia). In patients with endometrial cancer the total Sia level before surgical treatment (709.5 +/- 306.5 mg/l) was significantly higher (p < or = 0.0001) than that of the control group (213.5 +/- 88.7 mg/l). The elevation in Sia level was exclusively due to Neu5Ac. Following surgical therapy, serum Neu5Ac levels (699.4 +/- 305.6 mg/l) were significantly decreased (305.9 +/- 114.5 mg/l). In one case, where Neu5Ac level was increased 15 days and eight months after surgery (1.8 and 2.5 times as compared to control, respectively), a metastasis not detected during surgery was recorded. The obtained results suggest that Neu5Ac level in serum may be used as a tumor marker in evaluating the suitability of surgical treatment in early endometrial cancer. PMID- 10517361 TI - Liquid chromatographic-tandem mass spectrometric method for the determination of the neuraminidase inhibitor zanamivir (GG167) in human serum. AB - An LC-MS-MS method for the analysis of the neuraminidase inhibitor, zanamivir, in human serum is described. Zanamivir was extracted from protein precipitated human serum samples using Isolute SCX solid-phase extraction cartridges and analysed using reversed-phase chromatography with TurboIonSpray atmospheric pressure ionisation followed by mass spectrometric detection. The method uses a stable isotope internal standard, is highly specific and sensitive for a compound of this type and has been used for the analysis of human serum and urine samples from clinical studies. The method was extended to the analysis of serum and plasma samples from pre-clinical studies involving the rat, ferret and cell culture media. The method has been shown to be robust and valid over a concentration range of 10-5000 ng/ml using a 0.2-ml sample volume. The main advantages of this method compared to earlier procedures are primarily specificity, sensitivity, ease of sample preparation, small sample volume and short analysis time (ca. 5 min). PMID- 10517362 TI - Achiral and chiral high-performance liquid chromatography of verapamil and its metabolites in serum samples. AB - Rapid and simple achiral and chiral HPLC assays have been developed for the determination of verapamil and its metabolites in serum samples. Two achiral reversed-phase columns, Hisep C18 (150 x 4.6 mm) and NovaPak C18 (150 x 3.9 mm) were used for the simultaneous separation of all analyzed compounds. An alpha1 AGP column (100 x 4.0 mm) was recommended for successful chiral separations of verapamil and its seven metabolites. All analyses were realised with fluorescence detection at lambda(ex) = 276 nm and lambda(em) = 310 nm. Limits of quantitation were in the range 1.0 to 5 ng/ml for all compounds. Both off-line SPE (SepPak C18 cartridges) and the on-line SPE with a semipermeable surface SDS C8 pre-column, (10 x 4.6 mm) were used for the clean-up and sample preconcentration. Extraction recoveries for all analyzed compounds were 87.7 +/- 5.8 to 92.7 +/- 4.0% for off line SPE and 94.3 +/- 4.2 to 98.2 +/- 5.1% for on-line SPE. The complete assay could be applied for achiral and chiral monitoring verapamil and all its metabolites in serum samples. PMID- 10517363 TI - High-performance liquid chromatographic determination of peroxisomicine A1 (T 514) in genus Karwinskia. AB - A chromatographic method was developed for the T-514 determination in Karwinskia leaves, stems and roots. A C18 analytical column and a mobile phase consisting of methanol and McIlvaine buffer (pH 3) were used. T-514 was detected using a diode array detector and the chromatograms were recorded at 269 and 410 nm. A linear dependence of a peak area on the T-514 concentration (r=0.9991) was obtained in the range of 0.126-12.6 microg/ml. Limits of T-514 quantification (signal-to noise ratio 10) in plant samples were 126 ng/ml at 410 nm and 28 ng/ml at 269 nm. T-514 was extracted from the plant material with ethyl acetate. Optimal extraction conditions were studied: number of extraction steps, volume of extracting agent and extraction time. The extracts were cleaned up using solid phase extraction (SPE). SPE recoveries of 99.9% and 98.4% were achieved for the T 514 concentrations of 1.4 microg/ml and 0.26 microg/ml, respectively. PMID- 10517365 TI - Determination of L-756 423, a novel HIV protease inhibitor, in human plasma and urine using high-performance liquid chromatography with fluorescence detection. AB - A method for the determination of L-756 423, a novel HIV protease inhibitor, in human plasma and urine is described. Plasma and urine samples were extracted using 3M Empore extraction disk cartridges in the C18 and MPC (mixed-phase cation exchange) formats, respectively. The extract was analyzed using HPLC with fluorescence detection (ex 248 nm, em 300 nm), and included a column switching procedure to reduce run-time. The assay was linear in the concentration range 5 to 1000 ng/ml when 1-ml aliquots of plasma and urine were extracted. Recoveries of L-756 423 were greater than 84% over the calibration curve range using the described sample preparation procedures. Intra-day precision and accuracy for this assay was less than 9% RSD and within 7%, respectively. Inter-day variabilities for the plasma (n=17) and urine (n= 10) were less than 5% and 3% for low (15 ng/ml) and high (750 ng/ml) quality control samples. Bovine serum albumin (0.5%) was used as an additive to urine to prevent precipitation of L-756 423 during the storage of clinical samples. The assay was used in support of human clinical trials. PMID- 10517366 TI - Separation and quantification of ropinirole and some impurities using capillary liquid chromatography. AB - Ropinirole, 4-[2-(dipropylamino)ethyl]-1,3-dihydro-2H-indol-2-one, is a potent anti-Parkinson's disease drug developed by SmithKline Beecham Pharmaceuticals. Capillary liquid chromatography (CLC) was used for the separation and quantification of ropinirole and its five related impurities, potentially formed during its synthesis. A simultaneous optimization of three mobile phase parameters, i.e., pH, buffer concentration and acetonitrile content was performed employing an experimental design approach which proved a powerful tool in method development. The retention factors of the investigated substances in different mobile phases were determined. Baseline resolution of the six substances on a C18 reversed stationary phase was attained using a mobile phase with an optimized composition [acetonitrile-8.7 mM 2-(N-morpholino)ethanesulfonic acid adjusted to pH 6.0 (55:45, v/v)]. It was shown that CLC, operated in the isocratic mode under the mobile phase flow-rate of 4 microl/min, can determine the level of these impurities, down to a level of 0.06% of the main component within 25 min. PMID- 10517364 TI - Reversed-phase high-performance liquid chromatographic assay for the adenovirus type 5 proteome. AB - An RP-HPLC assay was developed for a recombinant adenovirus type 5. During chromatography, intact adenovirus dissociated into its structural components (DNA and proteins) and the viral proteome was separated yielding a characteristic fingerprint. The individual components were identified by matrix-assisted laser desorption ionization time-of-flight mass spectroscopy, N-terminal sequencing and amino acid composition. The assay was utilized to measure adenovirus particle concentration through quantification of structural proteins. Each structural protein provided independent measurement of virus concentration allowing verification of accuracy. The assay sensitivity is at or below 2 x 10(8) particles. Contrary to the benchmark spectrophotometric assay, the RP-HPLC assay was shown to be insensitive to contaminants common for partially purified adenovirus preparations. PMID- 10517367 TI - Fast liquid chromatographic-mass spectrometric determination of pharmaceutical compounds. AB - We present fast LC-MS-MS analyses of multicomponent mixtures containing flavones, sulfonamides, benzodiazepines and tricyclic amines. Using a short microbore HPLC column with small particle size, five to eight compounds were partially resolved within 15 to 30 s. TurboIonSpray and atmospheric pressure chemical ionization interfaces were well suited to tolerate the higher eluent flow-rates of 1.2 to 2 ml/min. The methods were applied to biological sample matrices after clean-up using solid-phase or liquid-liquid extraction. Good precision and accuracy (average 8.9 and 97.7%, respectively) were achieved for the determination of tricyclic amines in human plasma. Benzodiazepines were determined in human urine with average precision of 9% and average accuracy of 95% for intra- and inter assay. Detection limits in the low ng/ml range were obtained. An example for 240 injections per hour of demonstrated the feasibility of rapid LC-MS-MS analysis. PMID- 10517368 TI - Assay of 6-mercaptopurine and its metabolites in patient plasma by high performance liquid chromatography with diode-array detection. AB - A reversed-phase high-performance liquid chromatography (HPLC) method was developed to determine 6-mercaptopurine (MP) and seven of its metabolites (6 thioguanine, 6-thioxanthine, 6-mercaptopurine riboside, 6-thioguanosine, 6 thioxanthine riboside, 6-methylmercaptopurine and 6-methylmercaptopurine riboside) simultaneously in human plasma. A volume of 100 microl of plasma was used. Protein was removed from the sample by a simple and easy ultrafiltration step and ultrafiltrate was directly injected onto the HPLC system. Analytes were detected and confirmed with a diode-array detector before quantitation at 295 and 330 nm. The limit of detection for the analytes ranged from 20 to 50 nM. For the majority of patients receiving a 1 g/m2 MP intravenous infusion, MP and all metabolites except 6-thioguanine and 6-methylmercaptopurine riboside were present. This method serves as useful tool to characterize pharmacokinetics and pharmacodynamics of MP in oncology patients, and the small volume of plasma lends itself to pediatric studies. PMID- 10517369 TI - Urine screening of five-day-old newborns: metabolic profiling of neonatal galactosuria. AB - We determined urinary galactose and 4-hydroxyphenyllactic acid (4HPLA) in 4338 of 5-day-old newborns using a newly developed GC-MS screening method. Fifty-two infants were chemically diagnosed as having transient galactosuria based upon elevated urinary galactose levels (4.78-30.53 mg/mg creatinine, control 1.10 +/- 0.89 mg/mg creatinine). These infants did not excrete galactitol or galactonic acid into the urine, which is typical of hereditary galactosemia. Nearly 40% of the transient galactosuria was associated with immature infants (low birth weight or borne before 37 gestational weeks). Immature hepatic function is one explanation for neonatal transient galactosuria, but heterozygotes or the carriers of galactose degradation enzyme deficiencies were also suspected in some of the newborns, judging from the comparisons of urinary galactose and 4HPLA excretion between neonates and patients with galactosemia. PMID- 10517370 TI - Analysis of creatine and creatinine in urine by capillary electrophoresis. AB - Creatine is found in the urine of subjects ingesting creatine monohydrate as an ergogenic aid. Creatinine, the catabolic breakdown product of creatine, is a major constituent of normal urine. It is of interest to follow the excretion of creatine and creatinine in urine as a function of time after creatine ingestion. In this study, creatine and creatinine were analyzed in urine by capillary electrophoresis. The optimization of the method was discussed, with the best results being obtained using a 30 mM phosphate-150 mM sodium dodecyl sulfate buffer at pH 6, with the detector set at 214 nm and an applied voltage of 15 kV across a 45 cm capillary. Verification of the method was provided by HPLC analysis and spiking. The application of the method was demonstrated by analysis of creatine and creatinine in urine samples collected in a 24-h period following creatine ingestion. PMID- 10517371 TI - Simultaneous separation by high-performance liquid chromatography of carbamoyl aspartate, carbamoyl phosphate and dihydroorotic acid. AB - Leflunomide is an immunomodulatory drug which acts by inhibiting dihydroorotic acid dehydrogenase, the fourth enzyme of pyrimidine biosynthesis. We modified our high-performance liquid chromatography method to demonstrate that the principal metabolite in mitogen-stimulated human T-lymphocytes incubated with leflunomide was not dihydroorotic acid, but carbamoyl aspartate. Identification involved preparation of [14C]carbamoyl aspartate from [14C]aspartic acid and mammalian aspartate transcarbamoylase. Accumulation of carbamoyl aspartate indicates that under these conditions the equilibrium constant for dihydroorotase favours the reverse reaction. This HPLC method, enabling simultaneous separation of the first four intermediates in the de novo pyrimidine pathway may be of use in a variety of experimental situations. PMID- 10517372 TI - Purification of fatty acid methyl esters by high-performance liquid chromatography. AB - The determination by gas chromatography (GC) of fatty acid methyl esters (FAMEs) prepared from complex biological samples is subject to interference from cholesterol. During sample injection on the GC system of FAMEs prepared from tissues that contain cholesterol, we observed a major contaminant that co-eluted with docosahexaenoic acid (DHA, 22:6n-3). To address this problem, FAMEs were purified on an amino-phase high-performance liquid chromatography (HPLC) column using a hexane-isopropanol gradient. The HPLC retention times for both the FAME fraction and cholesterol were stable and reproducible when the amino column was used for sample purification. The purified extracts were analyzed by GC without artifacts or impurity peaks after 50 analytical runs. The method described here will be useful for measurement of 22:6n-3 and other fatty acids important for studies of nutrition or pathology. PMID- 10517373 TI - Comparison of plasma sample purification by manual liquid-liquid extraction, automated 96-well liquid-liquid extraction and automated 96-well solid-phase extraction for analysis by high-performance liquid chromatography with tandem mass spectrometry. AB - Three extraction procedures were developed for the quantitative determination of a carboxylic acid containing analyte (I) in human plasma by high-performance liquid chromatography (HPLC) with negative ion electrospray tandem mass spectrometry (MS-MS). The first procedure was based on the manual liquid-liquid extraction (LLE) of the acidified plasma samples with methyl tert.-butyl ether. The second procedure was based on the automation of the manual LLE procedure using 96-well collection plates and a robotic liquid handling system. The third approach was based on automated solid-phase extraction (SPE) using 96-well SPE plates and a robotic liquid handling system. A lower limit of quantitation of 50 pg/ml was achieved using all three extraction procedures. The total time required to prepare calibration curve standards, aliquot the standards and plasma samples, and process a total of 96 standards and samples by manual LLE was three-times longer than the time required for 96-well SPE or 96-well LLE (4 h, 50 min vs. 1 h, 43 min). Even more importantly, the time the bioanalyst physically spent on the 96-well LLE or 96-well SPE procedure was only a small fraction of the time spent on the manual LLE procedure (<10 min vs. 4 h, 10 min). It should be noted that the 96-well SPE procedure incorporated the two steps of evaporation of the eluates to dryness and subsequent reconstitution of the dried extract. The total time required for the 96-well SPE could be reduced by 50% if the eluates were injected directly, eliminating the drying and reconstitution steps, which is achievable when sensitivity is less of an issue. PMID- 10517374 TI - Determination of lorazepam in plasma and urine as trimethylsilyl derivative using gas chromatography-tandem mass spectrometry. AB - A procedure based on gas chromatography-tandem mass spectrometry for identification and quantitation of lorazepam in plasma and urine is presented. The analyte was extracted from biological fluids under alkaline conditions using solid-phase extraction with an Extrelut-1 column in the presence of oxazepam-d5 as the internal standard. Both compounds were then converted to their trimethylsilyl derivatives and the reaction products were identified and quantitated by gas chromatography-tandem mass spectrometry using the product ions of the two compounds (m/z 341, 306 and 267 for lorazepam derivative and m/z 346, 309 and 271 for oxazepam-d5 derivative) formed from the parent ions by collision induced dissociation in the ion trap spectrometer. Limit of quantitation was 0.1 ng/ml. This method was validated for urine and plasma samples of individuals in treatment with the drug. PMID- 10517375 TI - Stroke volume changes during dobutamine-atropine stress echocardiography: the influence of heart rate and ischaemia. AB - BACKGROUND: A decrease in stroke volume during dobutamine-atropine stress echocardiography heralds ischaemia and possible hypotension. Hypotension results from worsening of LV-function (as a result of ischaemia) left ventricular outflow tract obstruction or hypovolemia, while an increase of stroke volume indicates the preservation of myocardial contractile reserve. OBJECTIVE: To assess stroke volume changes during dobutamine stress echocardiography in relation to heart rate and occurrence of ischaemia and to validate a new automated cardiac output measurement device. METHODS: In fifty patients, the stroke volume was assessed using the echocardiographic biplane discs method during a stress echocardiography. These data were reference values for the validation of a new automated cardiac output measurement using the first method as a reference. RESULTS: Stroke volume measured by the biplane discs method and automated cardiac output device decreased from rest to peak stress, respectively, from 54+/-16 to 34+/-9 (63%) ml and 63+/-17 to 38+/-15 (60%) ml (p < 0.001). Stroke volume decreased with increased heart rate and stress-induced ischaemia when assessed by the biplane discs method, but with the automated device it decreased only with increased heart rate. CONCLUSIONS: Both increased heart rate and myocardial ischaemia during dobutamine stress echocardiography cause a reduction of stroke volume. However, the automated device did not detect the effects of stress induced ischaemia on stroke volume. It appears that the biplane discs method is more sensitive for evaluating the effect of ischaemia. PMID- 10517376 TI - The flow-function relationship in patients with chronic coronary artery disease and reduced regional function: a Doppler transesophageal and bidimensional transthoracic echocardiography study. AB - BACKGROUND: Infra-low dose dipyridamole allows one to selectively explore myocardial viability. Transesophageal echocardiography Doppler measurement of left anterior descending coronary artery flow at baseline and following dipyridamole is an efficient tool to assess coronary flow response. Aim of this study was to determine the flow-function relationship during coronary vasodilatory stress in patients with coronary artery disease and baseline dysfunction. METHODS AND RESULTS: Twelve patients with resting dyssynergies and 6 controls underwent assessment of regional function and of left anterior descending blood flow velocity. Flow and function were evaluated at rest and following infra-low dose dipyridamole (0.28 mg/Kg over 4 min). Controls showed a normal function at rest and after dipyridamole. Six patients ('Responders') with resting dyssynergies showed an improvement in segments of left anterior descending artery territory, whereas the other six ones ('Non-responders') showed no functional change. Controls and 'Responders' had similar values of resting peak diastolic left anterior descending artery flow velocity both at rest and after dipyridamole, whereas 'Non-responders' showed a blunted flow response to dipyridamole. CONCLUSION: Myocardial segments with a resting dysfunction and a contractile reserve more often exhibit a residual flow response, whereas segments with fixed pattern show a flat flow response during coronary vasodilator stress. PMID- 10517377 TI - Echocardiographic and Doppler study of patients with heatstroke and heat exhaustion. AB - This is a two-dimensional and Doppler echocardiographic study of the hemodynamic changes in patients with heatstroke and heat exhaustion. It demonstrates that the hemodynamic changes in severe heat exposure reflect a hyperdynamic circulation with tachycardia and high cardiac output states. Relative hypovolemia was more pronounced in patients with heatstroke compared to patients with heat exhaustion. Signs of peripheral vasoconstriction were more often present in patients with heatstroke, while patients with heat exhaustion more often demonstrated peripheral vasodilatation. PMID- 10517378 TI - Assessment of the mechanical properties of coronary arteries using intravascular ultrasound: an in vivo study. AB - The pressure-area relation of coronary arteries provides important information about the mechanical properties of these vessels. In human subjects methodological limitations have precluded measurement of instantaneous compliance and coronary stress in vivo. The purpose of this study was to assess a new method for measuring instantaneous values of coronary artery compliance and wall stress utilizing simultaneously acquired pressure and intravascular ultrasound measurements of vessel area. Ten subjects with coronary artery disease had intravascular ultrasound studies of the proximal left anterior descending or circumflex coronary arteries. Coronary luminal area was measured with a 30-MHz (3F or 3.5F) intravascular ultrasound catheter and simultaneous coronary pressure measured with a 2F micromanometer-tipped catheter. Using this technique the nonlinear pressure-area relation and mean circumferential wall stress were determined over the physiological pressure range. Coronary artery compliance at 100 mmHg ranged from 0.010 to 0.052 mm2/mmHg (mean +/- SD, 0.020+/-0.012 mm2/mmHg). Peak systolic circumferential stress ranged from 0.52 to 2.03 x 10(6) dyn/cm2 (1.09+/-0.42 x 10(6) dyn/cm2). This study describes a new method of determining coronary artery mechanical properties over the physiological pressure range. This technique may be useful in further studies of coronary artery mechanics. PMID- 10517379 TI - Does remodeling occur in the diseased human saphenous vein bypass grafts? An intravascular ultrasound study. AB - BACKGROUND: Coronary artery remodeling is a common phenomenon in human atherosclerotic arteries. Controversies exist concerning the presence of absence of the remodeling process in diseased human coronary saphenous vein bypass grafts. The purpose of the study was to observe the vessel and lumen dimensions in patients who had undergone saphenous vein grafting with intravascular ultrasound to find out whether the remodeling process exists in the diseased human saphenous vein bypass grafts. METHODS: A total of 43 saphenous vein bypass grafts from 43 patients (39 males, 4 females, mean age 63+/-8 years); 1-16 years (mean 9.3+/-4.0 years) after grafting, who had not undergone previous catheter intervention, were studied using intravascular ultrasound. The vessel, lumen and plaque area were measured at the lesion segment as well as in the proximal and distal reference segments. The percent stenosis was calculated. RESULTS: In 43 bypass grafts having severe stenosis before intervention, plaque was eccentric in 69.4% and concentric in 30.6%. No calcification was detected in 75% cases and 25% cases has mild-moderate intimal calcification. The vessel area in the lesion segment was 19.0+/-9.7 mm2, significantly larger than the proximal reference segment 12.8+/-4.0 min2 as well as the distal reference segment 12.9+/-3.6 mm2 (p < 0.001). It was also larger than that of the average area of the proximal and distal reference segments (p < 0.001). The vessel area increased in accordance with plaque area (p < 0.001). A weak relationship existed between vessel area and percent stenosis (r = 0.37, p = 0.04). CONCLUSION: In contrary to previous findings, diseased human saphenous vein bypass grafts undergo focal compensatory enlargement (remodeling) in the presence of plaque formation. The underlying mechanism is probably similar to that in de novo atherosclerosis. PMID- 10517380 TI - Performance of a Fourier-based program for three-dimensional reconstruction of the mitral annulus on application to sparse, noisy data. AB - OBJECTIVES: We investigated the accuracy of mitral annular reconstruction from noisy, sparse data typical of three-dimensional (3D) transthoracic echocardiograms. BACKGROUND: Our Fourier-based method for reconstructing the annulus from dense, accurate 3D transesophageal echo (TEE) data has been validated in vitro with four harmonics in the x, y, and z coordinates (4,4,4). METHODS: Thirteen mitral annuli were reconstructed from 'complete' 3D TEE data using four harmonics (4,4,4) and used to measure area, eccentricity. height, perimeter, and interpeak and intervalley distances; these were the 'true values'. To simulate transthoracic echo data, the TEE data sets were reduced evenly and unevenly (randomly). The complete and reduced data sets were used to reconstruct the annuli using three sets of fitting parameters: (4,4,4), (1,1,3), and (1,1,4). The resulting size and shape measurements were compared with true values. RESULTS: Regardless of the fitting parameters used, area, 2D perimeter, and 3D perimeter measurements were more accurate using reconstructions from evenly reduced than randomly-reduced data sets (p < 0.006), and depended significantly on both data density (p < 0.015 for all) and data distribution (p < 0.02 for all). Perimeter, height, and eccentricity of the reconstructed annuli were more accurately measured using four harmonics (4,4,4). CONCLUSIONS: Mitral annuli can be reconstructed from sparse, noisy data using the (4,4,4) fit if at least 25 points are obtained from evenly distributed imaging planes. These results suggest that detailed analysis of mitral annular size and shape can be made accurately from 3D transthoracic echocardiograms. PMID- 10517381 TI - Regional differences in shape and load in normal and diseased hearts studied by three dimensional tagged magnetic resonance imaging. AB - OBJECTIVES: We aimed to characterize regional geometry in relation to load in two groups of patients with hypertrophic cardiomyopathy (HCM) and right ventricular pressure overload (RVPO) in relation to a group of subjects with normal left ventricular (LV) function. BACKGROUND: Both these diseases are associated with marked changes in LV shape and function, which have not been studied with detailed three dimensional tools. METHODS: Three dimensional (3D) tagged magnetic resonance imaging (MRI) was used to characterize the 3D geometry and regional stresses of the left ventricles in patients with HCM and RVPO. Curvatures, stresses, wall thickness, and endocardial motion were calculated from surface and volume elements. RESULTS: Hearts with RVPO exhibited more circumferential and meridional flattening of the septum than normal and HCM hearts. The stress indices were lowest in the HCM hearts, compared to normal and RVPO hearts, due to the larger thicknesses. There was a more significant difference between lateral wall motion and other regional wall motions in the HCM and RVPO hearts as compared to normal hearts. CONCLUSIONS: It is suggested that curvature and stress mapping by 3D tagged MRI can be used as an important clinical tool for characterizing and distinguishing between healthy and diseased hearts. The results provided here validated previous knowledge on HCM and RVPO known from planary imaging methods. PMID- 10517382 TI - Double outlet right ventricle assessed with magnetic resonance imaging. AB - In this article the value of magnetic resonance (MR) imaging for the evaluation of double outlet right ventricle (DORV) is reviewed from the literature and illustrated with several cases. MR imaging can be used for the determination of cardiac anatomy at initial diagnosis and may provide functional information during the follow-up of patients after surgical correction. PMID- 10517383 TI - Visualization of regional myocardial depolarization by tangential component mapping on magnetocardiogram in children. AB - BACKGROUND: Tangential components to the body surface on magnetocardiography theoretically reflect regional myocardial current sources just below the gradiometer. The usefulness of tangential component mapping on magnetocardiography in determination of regional myocardial abnormalities has not been investigated in children. METHODS: Twenty-six children with ventricular hypertrophy, including a child with a left ventricular diverticulum (aged 7 to 15), and age matched 22 healthy children (aged 7 to 15) were studied. Tangential components on magnetocardiography were measured using a newly-developed super conducting quantum interference device system housed in a magnetically shielded room. Isomagnetic maps and current vector maps were constructed from the data obtained. RESULTS: The peak magnetic fields and current dipoles were demonstrated to be located at the interventricular septum initially, and then were shifted to the anterior and inferior walls of the left ventricle and to the right ventricular outflow tract, successively. In patients with right ventricular hypertrophy whose systolic right ventricular pressure was over 60 mmHg, the peak magnetic fields were located in the right half with rightward directed current vectors throughout ventricular depolarization. In patients with left ventricular hypertrophy, the maximal magnetic fields during depolarization were shifted to the hypertrophic site, showing significantly stronger forces than those in healthy children (35.5+/-11.7 pT vs 26.5+/-11.9 pT, p < 0.01). In a patient with left ventricular diverticulum, two discrete depolarizing current dipoles were visualized. The mean time required in measuring MCGs among all subjects was 10 minutes. CONCLUSION: The time course as well as the location of the regional electrical activities of the myocardium in children can be visualized, in a short time, as a two-dimensional projection to the frontal plane by tangential component mapping on magnetocardiography. PMID- 10517385 TI - New surgical options for the failing heart. AB - Heart failure is common in patients aged over 65 years, and is the fourth leading cause of hospitalization in the United States. Treatment of congestive heart failure involves remodeling the cardiac chamber with ventricular dilatation. New approaches to resolve this problem include medical therapies (digoxin, diuretics, ACE inhibitors, beta-blockers and phosphodiesterase inhibitors) to stabilize patients, followed by chamber remodeling. As yet, surgical intervention in advanced heart failure has been contraindicated, but newly evolving strategies show significant promise. It appears possible that patients can receive surgical therapy to improve cardiac function, followed by state-of-the-art medical therapy for congestive heart failure. This combined medical/surgical approach has led to the evolution of a new subspecialty within cardiology that deals with the management of heart failure. PMID- 10517384 TI - Heart valve replacement: a statistical review of 35 years' results. AB - BACKGROUND AND AIMS OF THE STUDY: Having performed our first heart valve replacement in 1960, we began a prospective lifetime follow up service for all patients, contacting them at least annually to determine survivorship and heart valve complications. METHODS: We reviewed isolated aortic (AVR) and mitral (MVR) valve replacements from 1960 to 1993, with follow up to 1998. In total, 2,942 AVR and 1,579 MVR were performed, with 21,742 and 12,142 patient-years of follow up, respectively. Analysis of the results affords an opportunity to demonstrate the usefulness and necessity of certain statistical methods, including multivariable event-free analyses and cumulative incidence functions. RESULTS: The survival rate was 8% at 30 years for both valve positions. However, an overall survival curve is an artificial composite of patients of increasingly higher risk being served during increasingly safer years of calendar time. One result is that, for AVR, age is not a significant univariate risk factor for operative mortality, but is highly significant after accounting for date of surgery using logistic regression. Long-term mortality is higher for tissue valves than for mechanical valves; but mean age is greater (74 versus 57 years), and after accounting for age using Cox regression, mortality is similar for both valve types. Kaplan-Meier analysis estimates thromboembolism occurrences of 85% for AVR and 95% for MVR at 35 and 34 years, respectively, but the cumulative incidence estimates are only 32% and 41%, respectively. CONCLUSIONS: Prospective follow up for over 35 years has provided an opportunity to illustrate important statistical issues: Multivariate analyses are essential to avoid being misled by excluding important risk factors or including artifactual ones, and the cumulative incidence estimates the percentage of patients who will actually experience a complication. PMID- 10517386 TI - Platelet aggregation and high-intensity transient signals (HITS) in a sheep model of mitral valve replacement. AB - BACKGROUND AND AIMS OF THE STUDY: The composition of microemboli detected as high intensity transient signals (HITS) by Doppler ultrasound in patients with prosthetic heart valves is still debated. Here, platelet aggregation and HITS were investigated in a sheep model. METHODS: Insonation of the carotid artery was performed in 20 sheep with either a mechanical or a biological mitral valve prosthesis in place. The effect of ICI 170809, a 5HT2a antagonist, on the frequency of HITS and on platelet aggregates, counted in arterial blood smears per nine high-power fields, was assessed at three and six months after valve implantation. The mitral transvalvular gradient was measured by transthoracic echocardiography at three and six months. RESULTS: Data are expressed as median and interquartile range. At three months, there were 36 (20-114) HITS/h in the mechanical group, and 0 (0-15) HITS/h in the biological group. At six months, there were 21 (0-82) and 0 (0-2) HITS/h, respectively. The occurrence of HITS was unaffected by either ICI 170809, or by duration of implant in either group. Platelet aggregate counts were higher with the mechanical than with the biological valve at three months, but not at six months. ICI 170809 reduced platelet aggregate counts in both valve types; the reduction was not significant in the bioprosthetic valve group. The pressure gradient across the bioprosthesis increased during the study from 2 (2-3) mmHg to 7.5 (6-10) mmHg, but was unchanged in the mechanical valve. CONCLUSIONS: (i) It was confirmed that the frequency of HITS is higher with the mechanical prosthesis than the bioprosthesis; (ii) circulating platelet aggregates in the bioprosthetic valve group tended to increase as structural valve deterioration occurred; (iii) the frequency of HITS was not influenced by either an increase or a decrease in circulating platelet aggregates; and (iv) HITS detected in patients with prosthetic valves are unlikely to be due to circulating platelet aggregates. PMID- 10517387 TI - High-frequency pressure fluctuations measured in heart valve patients. AB - BACKGROUND AND AIM OF THE STUDY: Due to the risk of thromboembolic complications, mechanical heart valve patients require life-long anticoagulant therapy, in contrast to bioprosthetic valves. The reason for this is still not fully understood. In vitro studies have demonstrated the presence of cavitation bubbles in the vicinity of mechanical heart valves, but not of bioprosthetic valves. When cavitation bubbles collapse, they release a significant amount of energy, which may damage the formed elements of the blood. A correlation between the presence of cavitation bubbles and high-frequency pressure oscillations has been established in vitro. Thus, the aim of this study was to measure and quantify high-frequency pressure oscillations in patients with normal, bioprosthetic or mechanical aortic valves. METHODS: Measurements were performed in six patients with normal aortic valves after coronary bypass surgery, in five patients fitted with a Carpentier-Edwards pericardial bioprosthesis, and in nine patients fitted with a St. Jude Medical or CarboMedics aortic valve. High-frequency pressure fluctuations were measured intraoperatively using a hydrophone placed near the aortic annulus. The root mean square (RMS) value of the high-frequency pressure signals were calculated in the frequency range 35-150 kHz. RESULTS: High frequency pressure fluctuations, with intensities above the noise floor, were registered only in the vicinity of mechanical heart valve prostheses, and not in the vicinity of normal or bioprosthetic valves. The mean value of RMS pressure fluctuations was 0.5 Pa for normal aortic valves, 0.8 Pa for bioprosthetic valves, and 67 Pa for mechanical valves. CONCLUSIONS: This study is the first to show the presence of high-frequency pressure fluctuations in patients with mechanical valves. PMID- 10517388 TI - Prospective evaluation of frequency and nature of transcranial high-intensity Doppler signals in prosthetic valve recipients. AB - BACKGROUND AND AIM OF THE STUDY: In asymptomatic prosthetic valve recipients, high-intensity transient signals (HITS) observed with transcranial Doppler (TCD) are a phenomenon of obscure clinical relevance which nature has not yet been elucidated convincingly. METHODS: Eighty-three patients without carotid disease, history of cerebrovascular accidents, and with negative preoperative TCD undergoing either valve replacement (mitral, n = 11; aortic, n = 56; mitral + aortic, n = 6; 40 mechanical prostheses, 29 biological prostheses, 10 homografts) or mitral repair (n = 10) were evaluated prospectively by means of TCD at discharge, three months and one year after surgery, to analyze the presence, incidence and characteristics of HITS. Furthermore, in 12 patients positive for HITS, TCD was repeated during a 30-min period of 100% O2 inhalation. RESULTS: Twenty-five patients (30%) were positive for HITS at all postoperative controls, although no neurological symptoms were observed. Mechanical prostheses showed a significantly higher incidence of HITS (85%) than biological prostheses (10%, p <0.001), repaired mitral valves (0%, p <0.001) and homografts (0%, p <0.001). At multivariate analysis the presence of a mechanical prosthesis was the only significant predictor of detection of HITS after valve replacement. During O2 inhalation, a significant decrease in the number of HITS per hour (55 +/- 79 versus 22 +/- 31, p = 0.002) occurred, which returned to initial values when room air breathing was resumed. CONCLUSIONS: Prosthetic valve replacement, particularly when mechanical devices are used, is associated with the generation of HITS which persist throughout the follow up period, but remain clinically silent. The decrease of HITS during O2 inhalation strongly supports the hypothesis of the gaseous nature of such signals and confirms the validity of this method in helping to differentiate gaseous microemboli from solid microemboli in prosthetic valve recipients. PMID- 10517389 TI - Aortic valve surgery: where we are and where we shall go. PMID- 10517390 TI - Pulmonary autograft root replacement: mid-term results. AB - BACKGROUND AND AIMS OF THE STUDY: The Ross operation was first performed as a root replacement in 1974, and only limited mid- and long-term results assessing durability and adaptation of the pulmonary root to systemic pressures are available. We reviewed our experience to assess function of the autograft valve and the autograft pulmonary root, and its adaptation to systemic pressures. METHODS: A total of 244 operative survivors (median age 22 years; range: 1 week to 62 years) were reviewed. Clinical follow up (within one year) was available on 98% of cases, and echocardiographic assessment within one year on 93%. Autograft and homograft valve function, aortic annulus diameter, autograft root sinus diameter and ascending aortic diameter were determined on the most recent echocardiogram. RESULTS: Actuarial freedom from autograft valve degeneration (non endocarditis autograft valve reoperation or severe autograft valve insufficiency or valve-related death) was 95 +/- 3% at 5 years and 93 +/-4% at 10 years. Actuarial freedom from all valve-related complications (autograft valve degeneration, autograft valve reoperation, homograft valve reoperation or valve related death) was 90 +/- 4% at 5 years and 83 +/-6% at 10 years. Actuarial freedom from autograft valve replacement was 98 +/- 2% at 5 years and 96 +/- 4% at 10 years. Actuarial survival rate was 98 +/- 2% at 5 years and 86 +/- 9% at 10 years. Aneurysmal dilation of the autograft root occurred in two patients; this was not associated with autograft valve degeneration, and these patients were followed closely. CONCLUSIONS: At 10 years, the Ross root replacement has a low risk of valve degeneration, valve-related complications and autograft valve replacement, and patient survival is excellent. Autograft valve reoperation and homograft valve reoperation have been the only significant late valve-related complications. Techniques to reduce autograft reoperation have been introduced, and hopefully methods to mediate the immunological response to the homograft valve will reduce the incidence of failure. Significant aneurysmal dilation of the pulmonary autograft root is rare. PMID- 10517391 TI - Successful dilatation of the small aortic root for implantation of a larger valve prosthesis. AB - Optimal hemodynamic performance is of paramount importance when implanting a prosthesis in a patient with a small aortic annulus. In order to avoid prosthesis size/patient body surface area mismatch, current techniques advocate aortic annulus patch enlargement or the use of a supra-annular prosthesis to ensure the implantation of an appropriately sized aortic valve. We present the first description of mechanical dilatation of a small aortic annulus as a simple alternative method. PMID- 10517392 TI - Valve orifice area alone is an insufficient index of aortic stenosis severity: effects of the proximal and distal geometry on transaortic energy loss. AB - BACKGROUND AND AIMS OF THE STUDY: Standard measures of hemodynamic severity of aortic valve stenosis vary widely among patients with and without clinical symptoms. Our hypothesis is that valve orifice area alone is not the sole determinant of adverse clinical outcome. Stenotic orifice area ratio is ratio of the cross-sectional stenotic orifice area to the down-stream, ascending aorta cross-sectional area. Determination of workload together with aortic valve orifice area ratio might improve risk stratification among asymptomatic patients with critical aortic stenosis. Accordingly, application of both parameters together might be useful in guiding management decisions in this condition. METHODS: In this study the dependency of transaortic fluid mechanical energy transfer (one component of left ventricular workload) on aortic valve orifice area is shown using modeling and experimental techniques. RESULTS: For a stroke volume of 62 ml at a heart rate of 60 beats/min, the piston work (analogous to left ventricular work) increased by 17% as the stenotic orifice area ratio decreased from 0.60 to 0.25, by 35% as the ratio fell from 0.25 to 0.20, and by 73% as the ratio fell from 0.20 to 0.10. CONCLUSIONS: As predicted by the fundamental fluid mechanical theory, simulated left ventricular work and energy loss in aortic stenosis are influenced not only by the effective stenotic valve orifice area, but also by the geometry of the inflow and outflow conduits, proximal and distal to the valve. These findings might explain clinically observed discrepancies between valve orifice area and the onset of the classical symptoms of severe aortic stenosis that reflect the left ventricular workload. Consideration of the left ventricular work in addition to the effective valve orifice area should enhance clinical evaluation, prognostication and risk stratification among patients with severe aortic stenosis. PMID- 10517393 TI - Exercise hemodynamic performance of the pulmonary autograft following the Ross procedure. AB - BACKGROUND AND AIMS OF THE STUDY: The Ross procedure, in which the aortic valve is replaced with the patient's own pulmonary valve (pulmonary autograft), is considered an excellent alternative for younger patients requiring elective aortic valve replacement. Although resting pulmonary autograft hemodynamics are excellent, exercise hemodynamic data are lacking. The study aim was to measure the hemodynamic performance of the pulmonary autograft with exercise Doppler echocardiography (DE). METHODS: Twenty-four Ross procedure patients (20 males, four females; mean age 46 +/- 11 years) were studied at 25 +/- 14 months after aortic valve replacement with a pulmonary autograft. Patients had baseline supine DE to measure the maximum velocity (Vmax), and the peak and mean pressure gradient across the pulmonary autograft. Effective orifice area was calculated from the continuity equation and indexed to body surface area (EOAi). Patients then underwent symptom-limited upright bicycle exercise with supine DE repeated immediately on stopping exercise. For comparison, 10 normal controls (age 41 +/ 10 years) and five mechanical aortic valve patients (mean age 55 +/- 10 years) were studied. RESULTS: At rest: Ross procedure patients had similar Vmax (1.2 +/- 0.2 m/s), peak gradient (6 +/- 2 mmHg), mean gradient (4 +/- 1 mmHg) and EOAi (1.7 +/- 0.4 cm2/m2) to those of normal controls. Mechanical-valve patients had significantly higher Vmax (2.5 +/- 0.2 m/s, p <0.001), peak gradient (25 +/- 4 mmHg, p <0.001) and mean gradient (14 +/- 3 mmHg, p <0.001) than Ross patients and normal controls. At exercise: Ross procedure patients had similar Vmax (1.8 +/- 0.4 m/s versus 2.1 +/- 0.2, p = NS), peak gradient (14 +/- 6 mmHg versus 17 +/- 4, p = NS) and mean gradient (8 +/- 4 mmHg versus 10 +/- 2, p = NS) to normal controls, with no significant change in EOAi. Mechanical-valve patients had significantly higher Vmax (3.4 +/- 0.3, p <0.001), peak gradient (48 +/- 7 mmHg, p <0.001) and mean gradient (30 +/- 5 mmHg, p <0.001) than Ross patients and normal controls. CONCLUSIONS: Aortic valve replacement using the Ross procedure provides excellent hemodynamic results at rest and on exercise, with DE parameters indistinguishable from those of normal controls. This study provides further support for the use of the Ross procedure as a preferred method of aortic valve replacement in younger patients. PMID- 10517394 TI - Cardiopulmonary exercise testing in patients with 21mm St. Jude Medical aortic prosthesis. AB - BACKGROUND AND AIM OF THE STUDY: Small-sized prostheses may be associated with high transprosthetic gradients, particularly in patients with a body surface area (BSA) >1.70m2, affecting left ventricular mass regression, symptom improvement and long-term survival. However, the influence of such gradients on exercise tolerance has not been clearly defined. The study aim was to verify the utility of cardiopulmonary exercise testing (CPX) in detecting patient-prosthesis mismatch, and to identify the clinical and echocardiographic data that predict exercise tolerance at CPX in patients with a 21mm St. Jude Medical (SJM) aortic prosthesis. METHODS: Twenty patients (one male, 19 females; mean age 66 +/- 9 years) with a 21 mm SJM prosthesis were evaluated by means of 2D echocardiography and CPX at 36 +/- 10 months after operation. Patients were divided into groups on the basis of a BSA of <1.70 m2 (group 1, n = 12) or > or =1.70 m2 (group 2, n = 8). RESULTS: At echocardiography, left ventricular mass reduction was 16 +/- 10% versus 9 +/- 6% in groups 1 and 2, respectively, mean gradient (MG) was 15 +/- 6 versus 17 +/- 4 mmHg (p = NS), effective orifice area index (EOAi) 0.86 +/- 0.10 versus 0.79 +/- 0.09 cm2/m2 (p = 0.05). At CPX, group 2 patients showed a significantly lower exercise duration (p = 0.02), maximum workload (p = 0.02), peak O2 uptake (p = 0.01), anaerobic threshold (AT) (p = 0.03), ventilatory equivalent for CO2 at AT (p = 0.007), and O2 cost of work (p = 0.03). Group 1 patients showed a ventilatory origin for their effort dyspnea, while group 2 patients showed a significant circulatory component. At multivariate analysis, BSA, age, EOAi and MG were independent predictors of CPX results. CONCLUSIONS: In patients with a 21 mm aortic SJM prosthesis and a BSA > or =1.70m2, CPX allows detection of patient-prosthesis mismatch, in terms of impaired exercise tolerance due to circulatory causes. CPX results can be anticipated on the basis of the patient's BSA, age, EOAi and MG. In these patients, technical solutions allowing implantation of a larger prosthesis should be considered whenever an active lifestyle is anticipated after aortic valve replacement. PMID- 10517395 TI - Rest and exercise Doppler hemodynamics of the Medtronic Intact aortic bioprosthesis. AB - BACKGROUND AND AIMS OF THE STUDY: Exercise treadmill testing was used to evaluate the functional rest and stress hemodynamic profile of the Medtronic Intact aortic bioprosthesis. METHODS: A group of 93 patients (mean age at operation 72.9 years; range: 61-79 years) was studied. Mean time to follow up was 20.8 months. The preoperative diagnosis was aortic stenosis (AS; n = 66), aortic regurgitation (AR; n = 19) or AS/AR (n = 8). Left ventricular function was assessed as normal (n = 78), moderate (n = 14) or poor (n = 1). Patients received a range of valve sizes: 21 mm (n = 7); 23 mm (n = 41); 25 mm (n = 32); 27 mm (n = 7); and 29 mm (n = 6). RESULTS: For all valve sizes, Doppler-derived hemodynamics at rest and peak exercise, respectively were: mean aortic valve gradient (AVG) 13.2 +/- 5.2 mmHg and 22.2 +/- 8.9 mmHg; peak aortic valve gradient (AVG) 24.3 +/- 9.6 mmHg and 39.1 +/- 13.5 mmHg; effective orifice area (EOA) 1.39 +/- 0.49 cm2 and 1.38 +/- 0.5 cm2; and effective orifice area index (EOAI) 0.76 +/- 0.26 cm2/m2 and 0.75 +/ 0.26 cm2/m2. Mean and peak AVG decreased as valve sizes increased, while both EOA and EOAI increased as valve sizes increased. CONCLUSIONS: The Medtronic Intact aortic bioprosthesis provides good hemodynamics both at rest and exercise, across the range of implanted valve sizes. PMID- 10517396 TI - Mitral valve reconstruction and replacement for ischemic mitral insufficiency: seven years' follow up. AB - BACKGROUND AND AIMS OF THE STUDY: Patients with ischemic mitral incompetence have a high operative risk whether the valve is repaired or replaced. The advantage of repair over replacement is unclear in this subgroup of patients. METHODS: Between April 1986 and December 1998, 337 patients underwent surgery for ischemic mitral valve insufficiency. Coronary artery bypass grafting (CABG) was carried out concomitantly in 314 cases (93.2%). Valve repair was performed in 140 patients (operative mortality rate 12.1%). The surgical risk in patients with a left ventricular ejection fraction (LVEF) of 10-30% was higher (operative mortality rate 33.3%) than in those with LVEF >30% (operative mortality rate 8.4%). Actuarial survival rates were 75.4%, 66.8% and 61.7% after 2, 5 and 7 years, respectively. Mitral valve replacement was performed in 197 patients (operative mortality rate 14.2%). The surgical risk in patients with a LVEF of 10-30% (operative mortality rate 30.3%) was greater than in those with a LVEF of >30% (operative mortality rate 11.0%).Actuarial survival rates after replacement were 78.6%, 73.4% 67.2% after 2, 5 and 7 years, respectively. RESULTS: Our initial analysis showed that, after mitral valve repair, mortality during follow up was greater in patients with residual mitral valve insufficiency of more than grade I. Subsequent outcome was superior when repair was evaluated perioperatively with transesophageal echocardiography. When mitral insufficiency was persistently more than grade I after repair, mitral replacement was performed. A total of 105 patients was analyzed with no or maximum grade I mitral insufficiency following valve repair; actuarial survival rates were 81%, 78.4% and 77.2% after 2, 5 and 7 years' follow up. CONCLUSIONS: Patients with highly impaired LV function and ischemic mitral insufficiency are at high risk during valve repair or replacement, and cardiac transplantation should be considered for this group. However, patients with ischemic mitral insufficiency and moderately impaired LV function can undergo valve repair or replacement with an acceptable prognosis. The goal of mitral valve repair should be to reduce valvular insufficiency to at least grade I. If this is not the case, then the prognosis of repair is worse than after valve replacement. Thus, the surgeon should replace the valve during the same operation. PMID- 10517397 TI - The long-term effect of successful mitral balloon valvotomy on left atrial size. AB - BACKGROUND AND AIM OF THE STUDY: The study aim was to determine the extent of regression of left atrial (LA) enlargement following mitral balloon valvotomy (MBV) for mitral stenosis. METHODS: Data obtained from 205 patients before, and at a mean of 31.0 +/- 21.1 months (range: 6 to 86.3 months) after successful MBV were analyzed retrospectively. RESULTS: The invasively determined mitral valve area increased from 0.81 +/- 0.27 cm2 at baseline to 1.73 +/- 0.54 cm2 immediately after valvotomy (p <0.0001), and the mean mitral gradient fell from 15.6 +/- 5.3 to 5.4 +/- 2.5 mmHg (p <0.0001). Similar changes were noted in Doppler-determined mitral valve area (0.89 +/- 0.16 to 1.97 +/- 0.29 cm2; p <0.0001) and gradient (12.6 +/- 5.3 to 4.9 +/- 1.7 mmHg; p <0.0001). In comparison with baseline, significant (p <0.0001) reductions were noted at follow up in the echocardiographic anteroposterior (48.7 +/- 6.9 to 42.4 +/- 6.6 mm), superior-inferior (68.5 +/-8.1 to 59.6 +/- 8.2 mm) and medial-lateral LA dimension (51.2 +/- 6.7 to 44.1 +/- 7.7 mm) and calculated LA volume (91.6 +/- 29.1 to 60.7 +/- 23.8 cm3) Patients in atrial fibrillation had larger LA dimensions, but substantially smaller absolute and relative reduction in LA size at follow up than patients in sinus rhythm. Among patients with prevalvotomy LA enlargement, normalization of LA dimension at follow up was seen in 29.2% of patients in sinus rhythm, but in none of the 32 with atrial fibrillation. CONCLUSIONS: Successful MBV results in significant long-term reduction in LA size in most patients, but normalization of LA size is unusual. PMID- 10517398 TI - Papillary muscle misalignment causes multiple mitral regurgitant jets: an ambiguous mechanism for functional mitral regurgitation. AB - BACKGROUND AND AIMS OF THE STUDY: The study aim was to test the hypothesis that asymmetric alignment (misalignment) of the papillary muscles is sufficient to cause incomplete mitral leaflet coaptation and functional mitral regurgitation (MR). METHODS: Different spatial relationships between the papillary muscles and the mitral annulus were investigated in isolated porcine mitral valves in vitro to assess the impact on mitral valve competence. The systolic occlusional leaflet area (OLA) needed to cover the mitral orifice and the anterolateral (ACOM) and posteromedial (PCOM) commissural portion (OLA(ACOM), OLA(PCOM)) were assessed by 2D echocardiography to quantitate incomplete mitral leaflet coaptation. The regurgitant fraction (RF) and MR jet location were assessed by a flow meter and color Doppler ultrasound. RESULTS: Posterolateral dislocation of the posteromedial papillary muscle impaired mitral leaflet coaptation at the corresponding half-portion of the mitral orifice (OLA(PCOM): 351-397 mm2 versus 296 mm2 (normal); p < 0.001) and modified the contralateral part (OLA(ACOM): 354 387 mm2 versus 304 mm2 (normal); p <0.001). The mitral leaflet coaptation line moved in apical and posterior directions, creating a commissural MR orifice at the PCOM side. At the ACOM side, anterior leaflet prolapse and restricted posterior leaflet mobility created an additional commissural regurgitant jet (RF = 0.11-0.13). Symmetrical papillary muscle misalignment restricted mitral leaflet mobility on both sides of the orifice in a synergistic manner (OLA(PCOM): 416-459 mm2 and OLA(ACOM): 427-489 mm2; both p <0.001 versus normal). The central MR jet orifice, which extended towards both commissures, caused more significant MR (RF = 0.15-0.26). CONCLUSIONS: Papillary muscle misalignment caused mitral regurgitant jet ambiguity with an anterior MR jet location following posteromedial papillary muscle displacement. These findings may improve understanding of the relation between myocardial lesion and mitral regurgitant jet location and thereby facilitate rational strategies for valvular interventions. PMID- 10517399 TI - Posterior myocardial infarction complicated by rupture of the posteromedial papillary muscle. AB - A 61-year-old man was admitted with acute posterior myocardial infarction and, on physical examination, was shown to have a mitral regurgitation (MR) murmur. Transthoracic echocardiography (TTE) showed severe hypokinesis of the posterior wall and severe MR by color flow. Right heart catheterization with a balloon tipped catheter revealed a pulmonary artery wedge pressure of 30 mmHg. No 'step up' was seen in blood samples from the right atrium and right ventricle. On angiography, a subtotal occlusion of the mid circumflex artery was found which was angioplastied and stented. As the patient's clinical condition did not improve, he underwent transesophageal echocardiography (TEE) for further evaluation. This showed complete rupture of the posteromedial papillary muscle. The patient underwent urgent surgery with successful mitral valve replacement. The postoperative course was uncomplicated, and clinical improvement seen. This case report underscores the value of TEE in accurate preoperative diagnosis of papillary muscle rupture by providing preoperative anatomic details of the mitral valve apparatus and surrounding structures. PMID- 10517400 TI - Isolated tricuspid valve surgery for severe tricuspid regurgitation following prior left heart valve surgery: analysis of outcome in 34 patients. AB - BACKGROUND AND AIMS OF THE STUDY: Patients with symptoms of right heart failure due to severe tricuspid regurgitation following a prior operation on left heart valves present a difficult problem. The outcome of tricuspid surgery in this setting is not well defined. We describe a single-center experience of isolated tricuspid valve surgery after prior left heart valve surgery, and analyze potential risk factors for a poor outcome. METHODS: Thirty-four patients who underwent isolated tricuspid valve operation for severe tricuspid regurgitation following prior valvular surgery for left-sided valve disease between 1980 and 1997 were identified. Charts were reviewed for clinical, echocardiographic, catheterization and surgical data. Follow up of survivors was conducted by telephone to ascertain functional status. RESULTS: Three patients died in hospital (early mortality rate, 8.8%). At a follow up of 71 +/- 39 months, 13 patients were alive and 21 reached an end-point (three cardiac reoperations, 18 deaths). Event-free actuarial survival at five years was 41.6 +/- 9.2%. Patients who were alive at follow up had a mean NYHA functional class of 2.1 +/- 0.6 compared with 3.4 +/- 0.5 preoperatively; 85% of survivors were symptomatically improved. Predictors of poor outcome were: increased age at the time of tricuspid surgery (p = 0.0007) and higher number of prior cardiac operations (one versus two or three, p-value 0.01, relative risk 3.4). Pulmonary artery systolic pressure, left ventricular ejection fraction, right ventricular function and size, annulus diameter, tricuspid valve pathology, and valve replacement versus repair were not predictive of outcome. CONCLUSIONS: Isolated tricuspid valve surgery for severe tricuspid regurgitation following prior surgery for left-sided heart valve disease can be performed with acceptable early mortality. There remains a high late mortality that is predicted only by age and the number of previous cardiac operations. However, in this selected group of severely symptomatic patients, significant improvement in symptoms are achieved in the survivors. PMID- 10517401 TI - Six-year experience with cryopreserved mitral homografts in the treatment of tricuspid valve endocarditis in HIV-infected drug addicts. AB - BACKGROUND AND AIM OF THE STUDY: In the past, valve homografts have been used in the treatment of aortic endocarditis. This report details our experience in tricuspid valve replacement using cryopreserved mitral homografts in HIV-positive drug addicts with infective endocarditis. METHODS: Five HIV-1-infected drug addicts with active uncontrollable tricuspid valve endocarditis underwent tricuspid valve replacement with a cryopreserved mitral homograft. RESULTS: There was no early mortality, and median follow up was 5 years (range: 1 to 6 years). One late mortality occurred as a result of heroin overdose. Three of the five patients developed six episodes of recurrent bacterial tricuspid endocarditis on the homograft; these were cured successfully with antibiotics. All survivors remain in NYHA functional class I. The latest transthoracic echocardiography examination showed mild, moderate or severe regurgitation in one, two and two patients, respectively. To date, neither homograft calcification nor rupture of the papillary muscle has been detected. CONCLUSIONS: This novel technique is considered to be an adequate approach to these cases of uncontrollable infectious disease. Further episodes of valvular infection can be managed medically. PMID- 10517402 TI - Unalloyed pyrolytic carbon for implanted mechanical heart valves. AB - Materials for implanted heart valves face challenges unmatched for most mechanical applications. The valves must function without interruption for lifetimes in the order of 10(9) cycles in a corrosive, hostile environment. A particular carbon has been widely used for the past three decades. The mechanical behavior of this isotropic pyrolytic carbon (PyC), with and without silicon alloying, was studied in this research program. Several questions were addressed: the fatigue strength at lifetimes of 10(9) cycles, the threshold for crack growth, the validity of fracture mechanics for PyC, and the statistics of ultra high survival under cyclic stress. It was found that cyclic stressing within the scatter band of the static strength did not initiate cracks. Furthermore, artificially induced cracks did not grow under cyclic stressing below a threshold stress intensity factor. This threshold stress intensity factor was valid over a wide range of crack lengths. No failures occurred when batches of 29 specimens were tested above the service stress at 6 x 10(8) cycles. Cyclic stressing did not reduce the static strength. The service stress is a small fraction of the fracture strength; thus, a very high probability of survival can be ensured by a proof test of the assembly. PMID- 10517403 TI - Brucella-mediated prosthetic valve endocarditis with brachial artery mycotic aneurysm. AB - A 39-year-old female with a Hall-Kaster mitral prosthesis developed fever, general malaise and arthralgia 15 years after valve replacement for rheumatic mitral valve disease. Prosthetic valve endocarditis was identified after serial laboratory, clinical and echocardiographic examinations. Penicillin G (40 x 106 units/day, i.v.) + gentamicin (240 mg/day, i.v.) was started as initial therapy. The patient showed no signs of recovery, and penicillin G was replaced with vancomycin (1,000 mg/day, i.v.). There was a gradual reduction in spiking fever, and prominent reductions in erythrocyte sedimentation rate and white cell count. Meanwhile, a tender and pulsatile mass developed in the anterior surface of the left arm; peripheral angiography yielded a diagnosis of brachial artery aneurysm. A successful aneurysmectomy with saphenous vein interposition was performed. Histopathology of the lesion revealed mycotic aneurysm. An initial control SAT for Brucella of 1/80(+) was found to increase. A detailed history showed the patient to have consumed unpasteurized dairy products. Doxycyline (200 mg/day, oral) + co-trimoxazole (2,700 mg/day, oral) + rifampicin (600 mg/day, oral) was administered to treat brucellosis. Later, doxycyline caused intolerable gastrointestinal side effects and was replaced by ciprofloxacin (1,000 mg/day, oral). Subsequently, the patient made an uneventful recovery within one week. Antibiotic treatment was continued for 12 months, with complete resolution of vegetation and paravalvular leakage. During a four-year follow up, the patient showed no signs of relapse. PMID- 10517405 TI - Islands I hardly knew. PMID- 10517404 TI - Fatal bacterial endocarditis following aortic valve replacement in a patient being treated with methotrexate. AB - A 41-year-old man being treated with methotrexate for psoriasis underwent aortic valve replacement. He subsequently developed fulminating bacterial endocarditis. Bacterial endocarditis occurs in 1-2% of cases after prosthetic valve replacement and has a high mortality. The long-term use of methotrexate and similar drugs is increasing in conditions such as psoriasis, rheumatoid arthritis and inflammatory bowel disease. Thus, more patients undergoing heart valve surgery will be taking these preparations for coexisting disease. As methotrexate increases the risk of infection, its perioperative use in these patients requires further evaluation. PMID- 10517406 TI - Women's health on the Web. PMID- 10517407 TI - Experts describe optimal symptom management for hospice patients. PMID- 10517408 TI - Recognizing and treating depression in the elderly. PMID- 10517409 TI - Slowing decline in AIDS deaths prompts concern. PMID- 10517410 TI - Recommendations on stem cell research. PMID- 10517411 TI - From the Food and Drug Administration. PMID- 10517412 TI - From the Centers for Disease Control and Prevention. Availability of hepatitis B vaccine that does not contain thimerosal as a preservative. PMID- 10517413 TI - From the Centers for Disease Control and Prevention. State-specific maternal mortality among black and white women--United States, 1987-1996. PMID- 10517414 TI - From the Centers for Disease Control and Prevention. Satellite broadcast on diagnostic and therapeutic dilemmas for gonococcal and chlamydial infections PMID- 10517415 TI - From the Centers for Disease Control and Prevention. Multiple human exposures to a rabid bear cub at a petting zoo and barnwarming--Iowa, August 1999. PMID- 10517416 TI - Dietary fat and risk of breast cancer. PMID- 10517417 TI - Dietary fat and risk of breast cancer. PMID- 10517418 TI - Left bundle-branch block and the ECG in diagnosis of acute myocardial infarction. PMID- 10517419 TI - Screening mammography for women younger than 50 years. PMID- 10517420 TI - Direct-to-consumer advertising: education or anathema? PMID- 10517421 TI - Direct-to-consumer advertising: education or anathema? PMID- 10517422 TI - Tobacco and alcohol use in children's animated films: Pecos Bill kicks the habit. PMID- 10517423 TI - Tobacco and alcohol use in children's animated films: Pecos Bill kicks the habit. PMID- 10517424 TI - Sexual dysfunction in the United States. PMID- 10517425 TI - Fruit and vegetable intake in relation to risk of ischemic stroke. AB - CONTEXT: Few studies have evaluated the relationship between fruit and vegetable intake and cardiovascular disease. OBJECTIVE: To examine the associations between fruit and vegetable intake and ischemic stroke. DESIGN, SETTING, AND SUBJECTS: Prospective cohort studies, including 75 596 women aged 34 to 59 years in the Nurses' Health Study with 14 years of follow-up (1980-1994), and 38683 men aged 40 to 75 years in the Health Professionals' Follow-up Study with 8 years of follow-up (1986-1994). All individuals were free of cardiovascular disease, cancer, and diabetes at baseline. MAIN OUTCOME MEASURE: Incidence of ischemic stroke by quintile of fruit and vegetable intake. RESULTS: A total of 366 women and 204 men had an ischemic stroke. After controlling for standard cardiovascular risk factors, persons in the highest quintile of fruit and vegetable intake (median of 5.1 servings per day among men and 5.8 servings per day among women) had a relative risk (RR) of 0.69 (95% confidence interval [CI], 0.52-0.92) compared with those in the lowest quintile. An increment of 1 serving per day of fruits or vegetables was associated with a 6% lower risk of ischemic stroke (RR, 0.94; 95 % CI, 0.90-0.99; P =.01, test for trend). Cruciferous vegetables (RR, 0.68 for an increment of 1 serving per day; 95% CI, 0.49-0.94), green leafy vegetables (RR, 0.79; 95% CI, 0.62-0.99), citrus fruit including juice (RR, 0.81; 95% CI, 0.68-0.96), and citrus fruit juice (RR, 0.75; 95% CI, 0.61-0.93) contributed most to the apparent protective effect of total fruits and vegetables. Legumes or potatoes were not associated with lower ischemic stroke risk. The multivariate pooled RR for total stroke was 0.96 (95% CI, 0.93-1.00) for each increment of 2 servings per day. CONCLUSIONS: These data support a protective relationship between consumption of fruit and vegetables-particularly cruciferous and green leafy vegetables and citrus fruit and juice-and ischemic stroke risk. PMID- 10517426 TI - Use of the oral neuraminidase inhibitor oseltamivir in experimental human influenza: randomized controlled trials for prevention and treatment. AB - CONTEXT: Influenza virus neuraminidase is thought to be essential for virus replication in humans; however, to date, available neuraminidase inhibitors are limited to zanamivir, which is topically administered. OBJECTIVE: To determine the safety, tolerability, and antiviral activity of oral neuraminidase inhibitor oseltamivir (GS4104/Ro64-0796) for prevention and the early treatment of influenza in experimentally infected humans. DESIGN: Two randomized, double blind, placebo-controlled trials conducted between June and July 1997. SETTING: Individual hotel rooms; 2 large US university medical schools. PARTICIPANTS: A total of 117 healthy adult volunteers (aged 18-40 years; median age, 21 years) who were susceptible (hemagglutination-inhibition antibody titer < or =1:8). INTERVENTIONS: All subjects were inoculated intranasally with influenza A/Texas/36/91 (H1N1) virus. For the prophylaxis study, oral oseltamivir (100 mg once daily [n = 12], 100 mg twice daily [n = 12], or matching placebo [n = 13], starting 26 hours before virus inoculation) was administered. For the treatment study, the same drug was given (20 mg, 100 mg, or 200 mg twice daily, 200 mg once daily, or matching placebo [n = 16], in each group starting 28 hours after inoculation). All regimens were continued for 5 days. MAIN OUTCOME MEASURES: Comparing placebo groups with pooled treatment groups, for prophylaxis, outcomes included frequency of infection and viral shedding; for treatment, viral shedding in titers. RESULTS: In the prophylaxis study, 8 (67%) of 12 placebo and 8 (38%) of 21 oseltamivir recipients became infected (P = .16; efficacy, 61%); 6 (50%) placebo compared with 0 oseltamivir recipients shed virus (P<.001; efficacy, 100%), and 33% of placebo but no oseltamivir recipient had infection-related respiratory illness (P<.01). Among infected subjects in the treatment study (n = 69), the viral titer area under the curve of the combined oseltamivir groups (n = 56) was lower (median [interquartile range [IQR]], 80 [23-151] vs 273 [79-306] log10 tissue culture-infective doses50 per milliliter x hour; P = .02) than the placebo group (n = 13), and the median (IQR) duration of viral shedding with therapy was reduced from 107 (83-131) to 58 (35-59) hours (P = .003). Oseltamivir treatment also reduced symptom scores (median [IQR] score-hours, 225 [97-349] vs 400 [189-645]; P = .05), and nasal proinflammatory cytokine levels. Transient mild to moderate nausea after dosing was observed in 15 (17%) of 88 oseltamivir and 2 (7%) of 29 placebo recipients (95% confidence interval for difference, -11% to 68%), which was largely prevented by ingestion with food. CONCLUSIONS: In these trials, prophylaxis and early treatment with oral oseltamivir were both associated with significant antiviral and clinical effects in experimental human influenza. PMID- 10517427 TI - Smoking in China: findings of the 1996 National Prevalence Survey. AB - CONTEXT: As the world's largest producer and consumer of tobacco products, China bears a large proportion of the global burden of smoking-related disease and may be experiencing a tobacco epidemic. OBJECTIVE: To develop an evidence-based approach supporting tobacco control initiatives in China. DESIGN AND SETTING: A population-based survey consisting of a 52-item questionnaire that included information on demographics, smoking history, smoking-related knowledge and attitudes, cessation, passive smoke exposure, and health status was administered in 145 disease surveillance points in the 30 provinces of China from March through July 1996. PARTICIPANTS: A nationally representative random sample of 128766 persons aged 15 to 69 years were asked to participate; 120298 (93.8%) provided data and were included in the final analysis. About two thirds of those sampled were from rural areas and one third were from urban areas. MAIN OUTCOME MEASURES: Current smoking patterns and attitudes; changes in smoking patterns and attitudes compared with results of a previous national survey conducted in 1984. RESULTS: A total of 41187 respondents smoked at least 1 cigarette per day, accounting for 34.1% of the total number of respondents, an increase of 3.4 percentage points since 1984. Current smoking continues to be prevalent among more men (63%) than women (3.8%). Age at smoking initiation declined by about 3 years for both men and women (from 28 to 25 years). Only a minority of smokers recognized that lung cancer (36%) and heart disease (4%) can be caused by smoking. Of the nonsmokers, 53.5% were exposed to environmental tobacco smoke at least 15 minutes per day on more than 1 day per week. Respondents were generally supportive of tobacco control measures. CONCLUSION: The high rates of smoking in men found in this study signal an urgent need for smoking prevention and cessation efforts; tobacco control initiatives are needed to maintain or decrease the currently low smoking prevalence in women. PMID- 10517428 TI - The relationship between cyclooxygenase-2 expression and colorectal cancer. AB - CONTEXT: Epidemiological studies have implicated the inducible form of cyclooxygenase (COX-2) in the pathogenesis of colorectal cancer; however, its role is not fully understood. OBJECTIVE: To examine the relationship between the expression of COX-2 in human colorectal cancer and patient survival. DESIGN: Patients diagnosed as having colorectal cancer were evaluated and followed up for up to 9.4 years (median follow-up, 2.7 years). Tumor sections were stained for COX-2 using a rabbit polyclonal antibody raised against human COX-2. The extent of COX-2 staining was graded by 2 observers blinded to outcome. Preabsorption of the anti-COX-2 antibody with a COX-2 peptide abolished the staining, demonstrating the specificity of the assay. SETTING: Gastrointestinal unit of a large general teaching hospital in Dublin, Ireland. PARTICIPANTS: Seventy-six patients (median age, 66.5 years) with colorectal cancer (Dukes tumor stage A, n = 9; Dukes B, n = 30; Dukes C, n = 25; Dukes D, n = 12) whose diagnosis was made between 1988 and 1991. Fourteen normal colon biopsies were stained for COX-2 as controls. MAIN OUTCOME MEASURES: Survival in years following diagnosis compared by extent of COX-2 epithelial staining (grade 1, <1%; grade 2, 1%-19%; grade 3, 20%-49%; grade 4, > or = 50%), Dukes stage, tumor size, and lymph mode metastasis. RESULTS: COX-2 was found in tumor epithelial cells, inflammatory cells, vascular endothelium, and/or fibroblasts. The extent of epithelial staining was heterogeneous, varying markedly among different tumors. Normal tissue adjacent to the tumors also stained weakly for COX-2. No COX-2 was detected in control tissue samples. The Kaplan-Meier survival estimate was 68% in patients who had grade 1 tumor epithelial staining compared with 35% in those with higher grades combined (log-rank chi2 = 5.7; P = .02). Greater expression of COX-2 correlated with more advanced Dukes stage (Kendall tau-b, 0.22; P = .03) and larger tumor size (Kendall tau-b, 0.21; P = .02) and was particularly evident in tumors with lymph node involvement (Kendall tau-b, 0.26; P = .02). CONCLUSIONS: Our data indicate that COX-2 expression in colorectal cancer may be related to survival. These data add to the growing epidemiological and experimental evidence that COX-2 may play a role in colorectal tumorigenesis. PMID- 10517429 TI - Blended payment methods in physician organizations under managed care. AB - CONTEXT: Independent practice associations (IPAs) are developing new methods of physician reimbursement to balance the objectives of encouraging individual productivity and clinical cooperation. The economic literature on payment incentives, derived from nonhealth industries, predicts that methods blending elements of fee-for-service and capitation will outperform exclusive reliance on either form of payment. OBJECTIVE: To identify emerging payment methods within IPA physician groups that contract with managed care organizations. DESIGN AND SETTING: Case studies of 7 large IPAs in the San Francisco, Calif, metropolitan region that served 826000 health maintenance organization (HMO) patients during the summer and fall of 1998. MAIN OUTCOME MEASURE: Payment methods of IPAs for primary care physicians, specialists, and physicians grouped by specialty department within the overall IPA structure. RESULTS: All the IPAs contracted with multiple HMOs for the full range of primary and specialty care physicians' services but paid member physicians using methods that blended elements of fee for-service and subcapitation. For primary care, most IPAs used monthly capitation adjusted for patient age, sex, and selected diagnoses, supplemented with fee-for-service payment for a wide range of visits and procedures, including patient visits in subacute, skilled nursing facility, emergency department, or home settings; for preventive care services; for office procedures requiring expensive supplies; and, most importantly, for borderline primary care procedures that either could be performed directly or referred to specialty care. All the IPAs paid specialty departments on a capitated basis and delegated to the departments responsibility for allocating the budget among individuals. Allocation mechanisms for individual specialists included adjusted fee-for service, referral-based capitation, and blends of both. CONCLUSION: Our results and case studies indicate that IPAs are developing payment methods that blend elements of fee-for-service and capitation in innovative ways for primary care and specialty physicians. PMID- 10517430 TI - Pharmacologic treatment of depression during pregnancy. AB - CONTEXT: Despite the frequency of depression in women of childbearing age, information to guide patients and physicians through a consideration of treatment during pregnancy is limited. OBJECTIVE: To identify risk factors associated with treatment of major depression during pregnancy to help physicians develop treatment plans that optimize clinical care. DATA SOURCES: Reports of prospective controlled trials in English were identified from MEDLINE and Health STAR using the search terms antidepressant during pregnancy and depression during pregnancy, by manually searching bibliographies of review articles, and through discussions with investigators for 1989-1999. STUDY SELECTION: We selected studies in which maternal and infant health outcomes associated with antidepressant exposure were compared with those of non-teratogen-exposed controls. Four studies published since 1993 were identified and included in the analysis. DATA EXTRACTION: We abstracted information about identification of subjects, comparison groups, pregnancy, and birth outcomes. We organized the data along 5 domains of reproductive toxicity: intrauterine fetal death, morphologic teratogenicity, growth impairment, behavioral teratogenicity, and neonatal toxicity. DATA SYNTHESIS: Data were available for tricyclic antidepressants (collectively), fluoxetine, and newer selective serotonin reuptake inhibitors (collectively). Exposure to these agents did not increase risk for intrauterine death or major birth defects. Decreased birth weights of infants exposed to fluoxetine in the third trimester were identified in 1 study. The development of children whose mothers took tricyclics or fluoxetine during gestation did not differ from that of controls. Direct drug effects and withdrawal syndromes occurred in some neonates whose mothers were treated with antidepressants near term. CONCLUSIONS: Although few in number, new information from prospective studies provides a welcome change from decision making based on nonprospective data. Monitoring and interventions for patients with identified risks (eg, poor weight gain) are recommended. PMID- 10517431 TI - The rational clinical examination. Does this patient have breast cancer? The screening clinical breast examination: should it be done? How? AB - CONTEXT: The clinical breast examination (CBE) is widely recommended and practiced as a tool for breast cancer screening; however, its effectiveness is dependent on its precision and accuracy. OBJECTIVE: To collect evidence on the effectiveness of CBE in screening for breast cancer and information on the best technique to use. DATA SOURCES: We searched the English-language literature using the MEDLINE database (1966-1997) and manual review of all reference lists, as well as contacting investigators of several published studies for clarifications and unpublished data. STUDY SELECTION AND DATA EXTRACTION: To study CBE effectiveness, we included all controlled trials and case-control studies in which CBE was at least part of the screening modality; for technique, we included both clinical studies and those that used silicone breast models. All 3 authors reviewed and agreed on the studies selected for inclusion in the pooled analyses. DATA SYNTHESIS: Randomized clinical trials demonstrated reduced breast cancer mortality rates among women screened by both CBE and mammography. Evidence of CBE's independent contribution was less direct; CBE alone detected between 3% and 45% of breast cancers found that screening mammography missed. The precision of CBE was difficult to determine because of the lack of consistent and standardized examination techniques. Studies on CBE precision reported fair agreement (kappa = 0.22-0.59). Pooling trial data, we estimated CBE sensitivity at 54% and specificity at 94%. The likelihood ratio of a positive CBE result is 10.6 (95% confidence interval [CI], 5.8-19.2), while the likelihood ratio of a negative test result is 0.47 (95% CI, 0.40-0.56). Longer duration of CBE and a higher number of specific techniques used were associated with greater accuracy. The preferred technique for CBE includes proper positioning of the patient, thoroughness of search, use of a vertical-strip search pattern, proper position and movement of the fingers, and a CBE duration of at least 3 minutes per breast. The value of inspection is unproved. Professional and lay examiners improved their sensitivity on silicone breast models after being taught this technique. CONCLUSIONS: Indirect evidence supports the effectiveness of CBE in screening for breast cancer. Although the screening clinical examination by itself does not rule out disease, the high specificity of certain abnormal findings greatly increases the probability of breast cancer. PMID- 10517432 TI - Service excellence in health care. PMID- 10517434 TI - Gay men and lesbians in medicine: has discrimination left the room? PMID- 10517433 TI - A future without tobacco: a call for papers. PMID- 10517435 TI - Uneasy partners: the lesbian and gay health care community and the AMA. PMID- 10517436 TI - Domestic partnership benefits at medical universities. PMID- 10517438 TI - JAMA Patient Page: diet. PMID- 10517437 TI - Sexual orientation and youth suicide. PMID- 10517439 TI - Women's health care centers and the women's imaging subspecialty: emerging frontiers in radiology. PMID- 10517440 TI - Unusual breast lesions: radiologic-pathologic correlation. AB - Unusual lesions of the breast can present a diagnostic challenge. These lesions include systemic diseases, benign tumors, and primary and metastatic malignancies. Lymphadenopathy is the most common mammographic finding associated with collagen vascular disease. Wegener granulomatosis may manifest as an irregular, high-density mass simulating breast cancer. Diabetic fibrous mastopathy manifests at mammography as very dense breast tissue and at ultrasonography (US) as an irregular, hypoechoic mass with striking posterior acoustic shadowing simulating malignancy. Fibromatosis simulates malignancy at mammography as an irregularly shaped, uncalcified, high-density mass and at US as an irregular, hypoechoic mass with posterior acoustic shadowing. At US, granular cell tumor may manifest as a solid, poorly marginated mass with marked posterior acoustic shadowing or may appear more benign. At mammography, hamartomas are typically well-circumscribed, round to oval masses with a thin, radiopaque pseudocapsule; at US, they manifest as a sharply defined, heterogeneous oval mass or as normal glandular tissue. Phyllodes tumor manifests at mammography as a large, well-circumscribed oval or lobulated mass; at US, it usually manifests as an inhomogeneous, solid-appearing mass. At mammography, primary breast lymphoma manifests as a relatively circumscribed mass or a solitary, indistinctly marginated, uncalcified mass. Metastatic lesions may manifest mammographically as single or multiple masses or as diffuse skin thickening; at US, they tend to have circumscribed margins with low-level internal echoes. Radiologists should be familiar with the characteristic mammographic appearances of these lesions and should consider benign and systemic causes in the differential diagnosis when malignant-appearing findings are encountered. PMID- 10517441 TI - Radial scar of the breast: radiologic-pathologic correlation in 22 cases. AB - Twenty-two cases were reviewed in which the diagnosis of radial scar (complex sclerosing lesion) of the breast was suspected preoperatively. At mammography, the lesions had a "black star" appearance with long, thin spicules radiating from a radiolucent central area. Excisional rather than core needle biopsy was recommended in all cases. In 13 of 22 cases, including one case of atypical ductal hyperplasia, the lesions proved benign at pathologic analysis. The remaining nine cases were malignant and included one case with a low-nuclear grade cribriform and micropapillary ductal carcinoma in situ adjacent to the lesion. Results of this study confirm the previously reported association of atypical ductal hyperplasia and carcinoma with radial scar. Furthermore, they demonstrate that a mammographic finding suggestive of radial scar may represent a malignancy that mimics the typical imaging findings in these entities. In cases of mammographically suspected radial scar, all members of the management team as well as the patient should be made aware preoperatively of the potential for benign as well as malignant pathologic findings. PMID- 10517442 TI - Imaging spectrum of extracapsular silicone: correlation of US, MR imaging, mammographic, and histopathologic findings. AB - The appearance of free silicone at mammography, ultrasonography (US), and magnetic resonance (MR) imaging is variable. The classic appearance is dense areas of opacity on mammograms, a highly echogenic pattern with or without hypoechoic masses on US images, and foci of low signal intensity on fat suppressed T1-weighted MR images or high signal intensity on water-suppressed T2 weighted MR images. Mammography is a reliable, cost-effective, and readily available means of demonstrating silicone. The major disadvantage of US is that its accuracy depends on the capability of the operator to recognize the abnormality. Although MR imaging outperforms US or mammography in detection of implant rupture, it is not clear that MR imaging is superior in detection of free or residual silicone. The sequelae of noncontained silicone include granuloma formation, fibrosis, and migration. After extrusion from an implant, silicone migrates primarily to local sites, such as the ipsilateral chest wall and axillary nodes. Migration of silicone into the axilla can involve the brachial plexus, resulting in neuropathy. Silicone can also migrate into more distal regions, including the arm and subcutaneous tissues of the abdominal wall. Whatever the source, silicone in breast tissue interferes with the interpretation of mammographic findings. PMID- 10517443 TI - Mammographic findings after breast conservation therapy. AB - Breast conservation therapy for breast cancer involves lumpectomy or segmental mastectomy followed by radiation therapy. Masses, fluid collections, architectural distortion, scarring, edema, skin thickening, and calcifications are posttreatment findings that may mimic or mask local tumor recurrence. Despite the overlap between posttreatment changes and tumor recurrence, the two entities can usually be distinguished by the characteristic mammographic appearances of posttreatment sequelae and by comparing interval findings on successive studies. Postoperative masses and fluid collections slowly diminish in size and usually resolve by 1 year after surgery. Radiation-induced edema gradually resolves; increasing edema may be due to recurrent cancer. Postsurgical scarring usually appears as a poorly marginated soft-tissue mass with interspersed radiolucent areas. Recurrent cancer is usually seen as a mass with no central radiolucent areas. Pleomorphic and granular microcalcifications are important markers for recurrent cancer and can usually be distinguished from the thick, calcified plaques and elongated dystrophic calcifications associated with scarring. PMID- 10517444 TI - Mammography of autologous myocutaneous flaps. AB - Autologous myocutaneous flaps (AMFs) are used increasingly as a method of breast reconstruction after mastectomy for breast cancer. Autogenous breast reconstruction may be performed with a rectus abdominis, latissimus dorsi, or gluteus maximus myocutaneous flap. Mammographic imaging of AMFs is controversial but has been recommended by some authors because mammographic detection of nonpalpable local recurrences in AMFs continues to be reported. At mammography, AMFs have a predominantly fatty appearance with variable density due to the muscle component and postoperative scarring. Normal mammographic findings include the vascular pedicle, surgical clips, and surgical scars, which produce radiopaque lines in predictable locations. Abnormal mammographic findings include fat necrosis appearing as a spiculated mass, noncalcified or calcified lipid cysts, calcifications, lymph nodes, epidermal inclusion cysts, and locally recurrent carcinoma. PMID- 10517445 TI - Breast imaging case of the day. Intramammary and axillary lymph node metastases from infiltrating lobular carcinoma of the breast. PMID- 10517446 TI - Breast imaging case of the day. Fat necrosis of the breast. PMID- 10517447 TI - CT of epithelial ovarian tumors. AB - Ovarian cancer is the second most common gynecologic malignancy in the United States and causes more deaths than any other cancer of the female reproductive system. Approximately two-thirds of patients have tumors that have spread beyond the pelvis at the time of diagnosis. Ovarian tumors arise from the surface epithelium or mesothelium, germ cells, or the gonadal stroma. Epithelial ovarian tumors include serous, mucinous, endometrioid, clear cell, and undifferentiated tumors. In general, the likelihood of malignancy increases with increasing solid tissue elements and thicker septa. Surgery is central to the management of ovarian cancer. At the initial exploratory laparotomy, surgicopathologic staging and debulking of the tumor are undertaken. Patients with advanced cancer frequently undergo second-look surgery after chemotherapy to detect any residual disease. CT can provide staging information for preoperative planning and determination of surgical resectability, demonstrate tumor response to therapy, and allow detection of persistent or recurrent disease. However, a major limitation of CT is the lack of sensitivity for detection of small tumor implants, especially on the small intestine or mesentery. Dedicated CT of the pelvis is best performed with spiral CT. Ovarian carcinoma can spread by means of intraperitoneal implantation, lymphatic invasion, and hematogenous dissemination. PMID- 10517448 TI - Recurrent cervical carcinoma: typical and atypical manifestations. AB - After treatment of cervical carcinoma, recurrent disease may be observed in multiple sites at imaging. Both typical and atypical manifestations of recurrent disease occur. Typical manifestations of recurrent cervical carcinoma involve the pelvis and lymph nodes. Pelvic recurrences may be observed as masses involving the cervix and uterus, vagina or vaginal cuff, parametria, bladder, ureters, rectum, or ovaries and may result in fistula formation or hydronephrosis. Nodal recurrence may be identified as enlarged pelvic and retroperitoneal nodes. Atypical manifestations of recurrent cervical carcinoma are being recognized with greater frequency due to the use of intensive pelvic radiation therapy, the evolution of improved imaging techniques, and the more frequent use of imaging as a means of surveillance. These atypical manifestations may involve the solid organs of the abdomen (focal masses) as well as the peritoneum, mesentery, and omentum (implants); gastrointestinal tract (obstruction, fistula formation, ischemia); chest (metastases to the lung parenchyma, pleura, and pericardium); bones (destructive lesions); and other sites. Familiarity with the imaging features of recurrent cervical carcinoma in these anatomic locations will facilitate prompt, accurate diagnosis and treatment. PMID- 10517449 TI - Sonohysterography: the next step in the evaluation of the abnormal endometrium. AB - Sonohysterography is a simple ultrasound (US) procedure that may be used to evaluate the endometrium. The technique involves placement of a 5-F catheter into the endometrial canal with subsequent instillation of sterile saline solution under US guidance. Fifty patients successfully underwent sonohysterography because of apparent abnormal endometrial thickening at transvaginal US, a nonspecific finding. Patients tolerated this procedure well, and no complications were encountered. In the 39 patients who proved to have endometrial pathologic conditions, sonohysterography demonstrated focal processes (polyps, carcinoma, hamartoma) in 15, diffuse processes (hyperplasia, secretory endometrium) in 21, and both focal and diffuse pathologic conditions in three. If a focal process can be delineated, a visually directed biopsy may be necessary. However, if the process is diffuse, a blind aspiration biopsy may be performed on an outpatient basis. In the majority of patients, the diffuse or focal nature of the disease could not be predicted on the basis of initial transvaginal US. Because sonohysterography allows distinction between diffuse and focal abnormalities, it provides physicians with a cost-effective way to plan the next step in case management. PMID- 10517450 TI - Unusual appearances of uterine leiomyomas: MR imaging findings and their histopathologic backgrounds. AB - Typical appearances of uterine leiomyoma at magnetic resonance (MR) imaging are well established, and diagnosis is usually easy. However, cases that, are extremely difficult to differentiate from other conditions are occasionally encountered. To understand the wide spectrum of MR imaging findings, such unusual appearances can be classified into three categories: degeneration and other histopathologic findings, specific types of unusual leiomyomas, and unusual growth patterns. The common types of degeneration are hyaline (>60% of cases), cystic (approximately 4%), myxoid, and red. Edema is not a phenomenon of degeneration but is a common histopathologic finding (approximately 50% of cases). Hemorrhage, necrosis, and calcification (approximately 4% of cases) may also be observed. Specific types of unusual leiomyomas include lipoleiomyoma and myxoid leiomyoma, which may have MR imaging features characteristic enough to allow differentiation from other gynecologic and nongynecologic diseases. Intravenous leiomyomatosis, metastasizing leiomyoma, diffuse leiomyomatosis, and peritoneal disseminated leiomyomatosis represent unusual growth patterns; other unusual growth patterns are retroperitoneal growth, parasitic growth, and the pattern that may occur in cervical leiomyoma. Because leiomyomas are the most common gynecologic tumors and are exclusively benign, it is important to be familiar with the variety of MR imaging appearances of uterine leiomyomas to distinguish them from other significant diseases. PMID- 10517451 TI - Uterine adenomyosis: endovaginal US and MR imaging features with histopathologic correlation. AB - Uterine adenomyosis is a common gynecologic condition that is characterized by the presence of heterotopic endometrial glands and stroma in the myometrium with adjacent smooth muscle hyperplasia. The histopathologic features of adenomyosis are varied and contribute to its imaging appearance. The accompanying smooth muscle hyperplasia produces the typical gross appearance of adenomyosis and corresponds to areas of decreased echogenicity at endovaginal ultrasonography (US) and areas of decreased signal intensity at magnetic resonance (MR) imaging. Endovaginal US also shows heterogeneity of the myometrial echotexture, which corresponds to small echogenic islands of heterotopic endometrial tissue surrounded by the hypoechoic smooth muscle. On T2-weighted MR images, bright foci are seen in areas of abnormal low signal intensity within the myometrium in approximately 50% of patients. These foci correspond to islands of heterotopic endometrial tissue, cystic dilatation of heterotopic glands, or hemorrhagic foci. With the advent of high-resolution imaging techniques, signs associated with the presence of heterotopic endometrial tissue are being detected with increasing frequency. These signs include myometrial cysts, myometrial nodules, linear striations, pseudowidening of the endometrium, and poor definition of the endomyometrial junction. Pitfalls in diagnosis of uterine adenomyosis include leiomyoma, endometrial carcinoma, myometrial contractions, and muscular hypertrophy. PMID- 10517452 TI - Diffuse and focal adenomyosis: MR imaging findings. AB - Adenomyosis is a common gynecologic disorder that affects women during their menstrual life. Preoperative magnetic resonance (MR) images obtained in 45 patients with pathologically proved adenomyosis who underwent hysterectomy were retrospectively reviewed. Diffuse adenomyosis was seen in 30 cases (66.7%) and focal adenomyosis in 15 cases (33.3%). On T2-weighted MR images, diffuse adenomyosis usually manifested as diffuse thickening of the endometrial myometrial junctional zone (7-37 mm; mean, 16 mm) with homogeneous low signal intensity. T2-weighted MR images were superior to contrast material-enhanced T1 weighted images in the evaluation of junctional zone thickening. High-signal intensity foci were observed on T2-weighted images only in nine cases and on both T1- and T2-weighted images in three cases. Focal adenomyosis manifested on both T2-weighted and contrast-enhanced T1-weighted MR images as a localized, low signal-intensity round or oval mass with a diameter of 2-7 cm (mean, 3.8 cm). All but one of the focal lesions had ill-defined margins. High-signal-intensity foci were noted in all cases of focal adenomyosis, either on T2-weighted images only (four cases) or on both T1- and T2-weighted images (11 cases). MR imaging is useful in diagnosing adenomyosis, differentiating adenomyosis from uterine myoma, and planning appropriate treatment. PMID- 10517453 TI - Endoanal MR imaging of the anal sphincter in fecal incontinence. AB - Fecal incontinence is a major medical and social problem. The most frequent cause is a pathologic condition of the anal sphincter. Endoanal magnetic resonance (MR) imaging allows detailed visualization of the normal anatomy and pathologic conditions of the anal sphincter. The hyperintense internal sphincter appears as a continuation of the smooth muscle of the rectum; the hypointense external sphincter surrounds the lower part of the internal sphincter. A sphincteric defect is seen as a discontinuity of the muscle ring. Scarring appears as a hypointense deformation of the normal pattern of the muscle layer. Two external sphincteric patterns may be misdiagnosed as defects: a posterior discontinuity (often seen in young male patients) and an anterior discontinuity (often seen in female patients). Atrophy of the external sphincter is easily detected on coronal MR images by comparing the thicknesses of all anal muscles. Endoanal MR imaging is superior to endoanal ultrasonography because of the multiplanar capability and higher inherent contrast resolution of the former. Use of endoanal MR imaging may lead to better selection of candidates for surgery and therefore better surgical results. Endoanal MR imaging is the most accurate technique for detection and characterization of sphincteric lesions and planning of optimal therapy. PMID- 10517454 TI - Nongynecologic applications of transvaginal US. AB - Transvaginal ultrasonography (US) is a noninvasive, readily available imaging technique that has greatly enhanced diagnostic sensitivity and accuracy for both gynecologic and nongynecologic disease. High-frequency US probes placed in the vagina allow high-resolution assessment of all the pelvic viscera, including portions of the gut and urinary tract. In addition, they allow visualization of the peritoneum of the pelvic pouch and the pelvic side walls without interference from bowel gas or adipose tissue. Evaluation of these areas requires a modified US technique that includes the use of the highest-frequency probes with angulation of the transducer to allow assessment of the region of interest. In women of childbearing age, the similarity of symptoms in gynecologic and gastrointestinal tract disease in particular underscores the potential utility of transvaginal US, which may, for example, help differentiate appendicitis in a pelvic appendix from pelvic inflammatory disease. Transvaginal US may also help determine the correct course of therapy, thereby improving patient management. Other indications for transvaginal US include assessment for pelvic appendicitis and diverticulitis, rectal and perianal complications of Crohn disease, and ureteric and bladder calculi and tumors as well as evaluation of the anal sphincters in women with fecal incontinence. Transvaginal US is also superior to routine US in the detection and characterization of ascites and peritoneal disease. Transvaginal US examination should include the entire pelvic cavity and contents, especially in women at risk for pelvic sepsis or peritoneal disease. PMID- 10517455 TI - Normal anatomy of the fetus at MR imaging. AB - Owing to recent advances in magnetic resonance (MR) imaging, the role of obstetric MR imaging has increased in cases in which the results of ultrasonography are equivocal. Fast MR imaging sequences, such as T2-weighted fast spin-echo (SE), half-Fourier single-shot fast SE, 0.5-signal-acquired single shot fast SE, and echo-planar imaging, have virtually eliminated the need for fetal premedication, with a concomitant improvement in image resolution and diminished blurring. Artifacts related to maternal respiratory motion and fetal motion no longer limit the anatomic detail that can be demonstrated with MR imaging. With such advances in obstetric MR imaging, knowledge of normal fetal anatomy at MR imaging is essential to detect disease in utero. MR imaging can demonstrate fetal anatomy in detail, especially the brain, thorax, abdomen, pelvis, and vasculature. Major developmental structures of the fetus, particularly the cranial nervous system, naso- and oropharynx, lungs, and major abdominal viscera, can be adequately evaluated with targeted fast MR imaging as early as the beginning of the second trimester. However, MR imaging of the heart remains limited. Fetal MR imaging during the first trimester remains controversial secondary to biosafety issues and is limited due to diminutive fetal size. PMID- 10517456 TI - Single-shot fast spin-echo MR imaging of the fetus: a pictorial essay. AB - Ultrasonography (US) is the modality of choice for prenatal screening, but occasionally additional imaging information is needed. Magnetic resonance (MR) imaging is an attractive alternative but until recently has been limited by motion artifact. Single-shot fast spin-echo MR imaging was used to depict normal and abnormal anatomy in 26 fetuses. Thirteen studies were performed for maternal indications and 13 were performed to evaluate fetal abnormalities identified or suspected at US. Three of the fetal abnormalities involved the central nervous system (CNS) and 10 involved other anatomic sites. Results were correlated with findings at postnatal clinical examination, imaging, and pathologic analysis. MR imaging demonstrated normal fetal anatomy without substantial motion artifact. CNS structures were well visualized as early as 18-20 weeks gestation, as were most other normal anatomic structures except the heart. MR imaging also allowed characterization of a variety of abnormalities of the CNS (Arnold-Chiari malformation, Walker-Warburg syndrome, amniotic band syndrome) as well as of other structures (renal agenesis, multicystic dysplastic kidney, abdominal masses, severe limb-body wall defect, clubfoot with arthrogryposis, diaphragmatic hernia). US findings were confirmed in most cases, and additional information about the precise diagnosis or the severity or location of the anomaly often helped guide clinical management. Single-shot fast spin-echo MR imaging of the fetus is a useful adjunct to US in difficult diagnostic situations. PMID- 10517457 TI - US assessment of the fetal head and neck: a state-of-the-art pictorial review. AB - When attention is paid to the details of normal and abnormal fetal head and neck anatomy, abnormalities that normally would be missed at prenatal ultrasonography can routinely be diagnosed. Five basic views are used to assess the fetal head and neck: a transverse view of the head in the plane of the cavum septum pellucidum and cerebellum, a sagittal and a coronal view of the face to visualize the nose and lips, a sagittal view of the cervical spine, and a transverse view of the orbits to measure the biorbital and interorbital distances. Thickened nuchal fold, a common sign of Down syndrome, can be assessed with transverse images of the head. Transverse views are also useful to demonstrate cystic hygroma, occipital meningocele, and encephalocele, all of which can be associated with other severe anomalies. Micrognathia, cleft lip and palate, and macroglossia, which are best depicted with sagittal and coronal views of the face, are also associated with other fetal abnormalities. Visualization of these entities should prompt further search and amniocentesis. Lymphangioma of the tongue appears similar to macroglossia but is an isolated anomaly. Transverse views through the orbits are helpful for demonstrating orbital teratoma, orbital encephalocele, and hypo- and hypertelorism (the latter two being associated with other abnormalities). Sagittal views of the neck can demonstrate cystic hygroma, teratoma, and an enlarged thyroid. PMID- 10517458 TI - Screening helical CT for evaluation of blunt traumatic injury in the pregnant patient. AB - Pregnant patients who sustain severe blunt trauma are infrequently encountered in most practices. However, detection of internal injuries including those to the gravid uterus is essential since maternal disability or fetal loss are physical and psychologic catastrophes that have long-term effects on the mother and her family. Computed tomography (CT) is commonly used to detect blunt traumatic injuries and can play an important role in the screening of the injured pregnant woman. The normal gravid uterus and physiologic changes of pregnancy can confound CT interpretation. Inhomogeneous enhancement of placental cotyledons, hydronephrosis, and enlarged ovarian veins are normal findings. Avascular regions in the placenta indicate infarction or abruption with impending fetal demise. Although CT can demonstrate uterine rupture and retroperitoneal hemorrhage, direct detection of fetal injuries is rare. Fetal demise is more common when maternal injuries include trauma to the uterus. Although screening ultrasonography can depict fetal distress, use of screening CT allows a concurrent evaluation of multiple areas in the pregnant trauma patient including the uterus. CT is a useful diagnostic tool in the triage of the critically injured pregnant woman. PMID- 10517459 TI - Composite cervicofacial flap for reconstruction of complex cheek defects. AB - The authors present the reconstructive technique for complex cheek defects using the composite cervicofacial flap and study the possibilities, advantages, disadvantages, and results that can be expected. The design follows the classic outline of Mustarde's flap. The skin is undermined for 2 cm anterior to the ear, then after incision of the superficial musculoaponeurotic system (SMAS), undermining is continued below the plane of the SMAS, level with the facial nerve branches. It is continued forward to the facial vessels, which give rise to branches that ensure the blood supply of this composite flap and contribute to its high reliability. In the cervical region, undermining is done beneath the platysma, which is transected transversely in the lower cervical region to allow good upward mobility and satisfactory transposition of the flap. The flap is adapted to the defect and the medial suture line is placed as near as possible to the medial limit of the cheek aesthetic unit. The authors carried out a retrospective study of 7 patients with complex facial reconstruction after excision of malignant lesions. The defects measured from 4x4 cm to 9x7 cm. In 4 patients excision included the periosteum, and in 1 patient excision involved the entire thickness and removed the entire anterior half of the cheek. In 4 patients reconstruction involved the cheek and eyelid. In spite of the advanced age of the patients (88, 69, 91, 67, 70, 82, and 59 years), there was no distal edge necrosis. The only complication was a single case of facial paresis, which resolved spontaneously. The results were considered very good in all 7 patients. The authors conclude that the composite flap increases the possibilities of the cervicofacial flap. It is more mobile, more reliable, thicker, and more adaptable. It can be used in complex cheek defects that involve the periosteum, or even in full-thickness defects. The quality of the results obtained using this flap represents a considerable advance in facial reconstruction. PMID- 10517460 TI - Muscle bow traction method for dynamic facial reanimation. AB - A muscle bow traction method was developed for dynamic facial reanimation utilizing the masseter muscle and a fascial sling. The principle of this method is that the sling around the muscle pulls the oral commissure laterally and backward by the restoring force of the muscle from its relaxed position to its contracted position. The surgical procedure is simple. The sling is passed around the anterior half of the muscle so that the muscle can be bowed anteriorly at its center by the sling. One end of the sling is sutured to the center of the orbicularis oris and the dermis in front of the nasolabial fold, and the other end is sutured to the lower lip and oral commissure. This method was applied to 3 patients with facial palsy and to 1 patient with oral cancer. The restored motion of the oral commissure ranged from 5 to 8 mm when clenching the jaws. The concept of this method differs from those of other muscle transposition methods for facial reanimation in that the force acts at a right angle to the muscle contraction. The advantage of this method is that it is less invasive to the muscle and is a simpler procedure than other conventional muscle transposition methods. PMID- 10517461 TI - Combined radiological and surgical treatment of arteriovenous malformations of the head and neck. AB - Arteriovenous malformations (AVMs) are high-flow lesions. More than 50% of all AVMs are located in the head and neck region. They represent a therapeutic challenge because of their hemodynamic characteristics and their modality of growth. AVMs have a tendency to recur and often require radical resection, making surgical ablation and reconstruction difficult. AVMs require angiography not only for diagnostic purposes but as an initial therapeutic step in the form of embolization. Surgical ablation, which follows a few days after embolization, is facilitated by the reduction in vascularity and shrinkage of the lesion, both of which are afforded by the embolization. These benefits allow for less blood loss at the time of ablation, and less extensive resection. The authors report their experience with 16 patients with extracranial AVMs of the head and neck examined over the last decade. PMID- 10517462 TI - Reversed neurofasciocutaneous flaps based on the superficial branches of the radial nerve. AB - Soft-tissue reconstruction of the hand needs to cover the vital structures with flaps. It is usually difficult to maintain function and form with minimal morbidity. Local tissue is preferable but it is also very valuable. Especially in the distal part of the upper extremity, flap coverage is a challenging problem because of limited reconstructive alternatives. On the dorsum of the hand, flaps can be designed based on the paraneural vascular network of the cutaneous sensory nerves. These paraneural vascular networks send branches to the surrounding tissues. The branches to the skin are known as neurocutaneous perforators. The authors used eight reversed neurofasciocutaneous flaps based on the superficial branches of the radial nerve. Six flaps were based on the branch to the index finger and two flaps were based on the branch to the thumb. All flaps survived completely, and successful flap coverage was achieved in all patients with minimal morbidity. PMID- 10517463 TI - Tuberculosis of the upper extremity. AB - Twelve cases of tuberculosis of the upper extremity over a 5-year period are presented. The average time to diagnosis was 5 months. All patients were treated with a regimen of combination antituberculous chemotherapy for a minimum of 6 months, initial splinting, and intensive physiotherapy. The indications for surgery were limited, with biopsy being the most common procedure. Patients with tenosynovitis underwent tenosynovectomy, and nerve decompression was performed when indicated clinically. Large abscesses were drained. No patient had bony debridement or early arthrodesis to control the infection. The pre- and posttreatment range of motion was recorded, with a mean follow up of 25.4 months. Employing this regimen resulted in resolution of infection and an improved range of motion in 11 patients. PMID- 10517464 TI - Surgical treatment of pseudosyndactyly of the hand in epidermolysis bullosa: histological analysis of an acellular allograft dermal matrix. AB - Recessive dystrophic epidermolysis bullosa is an inherited mechanobullous disorder of skin and mucous membranes. The most striking clinical characteristic of the disease is the formation of blisters following trivial trauma. Repeated cycles of blistering and scarring result in gradual encasement of the hand in an epidermal "cocoon." The authors treated an 11-year-old boy with recessive dystrophic epidermolysis bullosa who presented with hand contractures and interdigital pseudosyndactyly. Treatment included release of contractures and application of a biosynthetic dermal analog. This report is a histological analysis of the dermal matrix 1 year after initial placement of the allograft. Fibroblasts repopulating the dermal allograft had a normal synthetic phenotype and lacked the myofibroblastic features seen in the ungrafted control biopsy. Collagen and elastin in the repopulated dermal allograft had normal dermal orientation and maturity in contrast to the sparse, immature collagen and lack of elastin compared with the dermis of an ungrafted control region. Results of this histological study indicate that treatment of recessive dystrophic epidermolysis bullosa with an acellular human dermal allograft may restore some features of normal dermal architecture. Although the initial results are encouraging, longer follow-up is required before definitive conclusions can be made. PMID- 10517465 TI - Modified Dakin's solution for cutaneous vibrio infections. AB - Vibrio species, specifically Vibrio vulnificus, are known to be endemic to warm saltwater environments. As a human pathogen they are capable of causing severe, progressive, necrotizing infections. The lesions are bullous in nature and often require wide surgical debridement due to the aggressiveness of this organism. The literature supports prophylactic antibiotic therapy for those with preexisting hepatic dysfunction or immunocompromise. The authors routinely implement prophylactic antibiotic coverage with doxycycline 100 mg every 12 hours for vibrio in patients with wounds exposed to or acquired in saltwater. In addition, they institute topical therapy with 0.025% sodium hypochlorite solution (modified Dakin's), based on their in vitro study of vibrio sensitivity to antimicrobials. Over the past 2 years, the authors have treated 10 patients with this protocol for cutaneous vibrio infections confirmed by quantitative cultures. None of these patients experienced progression of infection requiring operative debridement contrary to the aggressive nature of this organism documented in other reports. PMID- 10517466 TI - The fissura antitragohelicina: an anatomic aid to the correction of prominent ears. AB - One of the important features of correction of prominent ears involves the creation of an antihelical fold in the ear cartilage. The precise and symmetrical location of this fold is crucial for the aesthetic result. This study investigated the use of the fissura antitragohelicina, a constant anatomic landmark, as a guide to the correct line for the new antihelix. In the first part of the study, 16 cadaveric ears were dissected. The fissura antitragohelicina was present in each specimen, and measurements of the distance between the fissura antitragohelicina and the helix and the antihelix were recorded. Based on this study, a clinical series of 20 consecutive prominent ear corrections were performed using the fissura antitragohelicina as a guide for the creation of a new, symmetrical antihelical fold. The aesthetic results were satisfactory by subjective assessment in every one of this group of patients. This study showed that the fissura antitragohelicina was a constant, reliable, and simple guide to the creation of the antihelical fold in patients with prominent ears. PMID- 10517467 TI - Modified McIndoe procedure for vaginal agenesis. AB - Between 1982 and 1997, 29 patients with congenital absence of the vagina underwent modified McIndoe vaginoplasty at the Division of Plastic and Reconstructive Surgery, Ege University Medical School, Izmir, Turkey. As a modification, an X incision was utilized instead of a straight-line horizontal or sagittal incision. During the first postoperative week, a perforated Pyrex rigid mold was used. This was replaced with an unperforated mold at the end of the first week. These patients' medical records were reviewed retrospectively. Complications encountered included infection, total lack of skin graft take, stress urinary incontinence, partial graft loss, and vaginal stricture. All complications were treated except the stress urinary incontinence, and the final results were satisfactory. PMID- 10517468 TI - A model for the assessment of sensory recovery of experimental skin grafts. AB - The return of sensation to skin grafts is often suboptimal. Although the reinnervation of skin grafts has been examined by a number of authors during the past century, studies in humans have left a number of unanswered issues, whereas animal studies have been largely confined to histological work and a few electrophysiological studies. Based on knowledge that rats exhibit a reflexive flick of the back skin in response to stimulation, the authors hypothesized that it should be possible to develop a noninvasive model for assessing return of sensation in experimental skin grafts. Full-thickness skin grafts were created, one per animal, on the dorsa of male Sprague-Dawley rats. Sensory testing was performed using a hand-held pinprick device designed to deliver a stimulus of reproducible force. A positive response was observed as a flick of the dorsal skin--a very reliable reflex involving the cutaneus trunci muscle. The stimulus was delivered to each of 25 sectors of the graft on days 9, 13, 16, 20, 40, 60, and 110 postoperatively. Results were analyzed regarding the percentage of grafts responding at each time point as well as the topographical pattern of sensory return. Evidence of sensation was first detected at day 13 at the margins of the skin grafts and then progressed centrally until homogenous reinnervation (94% of the graft surface) was observed at day 40 and was maintained through the end of the study. Growth Associated Protein (GAP)-43 immunostaining was used to document reinnervation of the skin grafts histologically. PMID- 10517469 TI - Quantitative assessment of microhemodynamics in ischemic skin flap tissue by intravital microscopy. AB - Skin flaps are susceptible to ischemia, which may result in tissue necrosis particularly in areas deprived of their original anatomic blood supply. The pathophysiology of skin flap failure has been debated for many years, but due to methodological insufficiencies, every proposed theory has remained hypothetical. The aim of this study was to gain more evidence for the mechanisms involved in flap ischemia by assessing quantitatively hemodynamic parameters such as diameter, flow velocity, and volume flow in the microcirculation of a flap. To this end the authors developed a new island flap on the back of Syrian golden hamsters that allowed intravital microscopic investigation. The flap included an extended portion, which was deprived of its original anatomic blood supply. One hour after flap dissection, blood flow was 42% to 66% lower in all microvessels in the extended area than in the anatomically perfused part of the flap (p<0.05). In the entire microvasculature, a significant gradual decline of blood flow was observed over time. Any blood flow reduction was caused to a major extent by diminished flow velocity. At all times, microvascular diameters were slightly larger in the extended portion of the flap than in the anatomically perfused portion of the flap. The authors conclude that their new model is a unique tool for investigating microhemodynamic mechanisms involved in flap ischemia. This study reveals hypoperfusion of extended flap tissue, which is attributed to diminished arterial perfusion pressure but not to vasoconstriction or arteriovenous shunting. PMID- 10517470 TI - Saphenous neurocutaneous island flap model in the rat: evaluation of vascular supply. AB - Neurocutaneous flaps are utilized routinely in reconstructive surgery and even more so during the past decade. In this study, the vascular supply of the neurocutaneous flap in the rat model is presented and evaluated. Thirty-six flaps (3.5x3 cm2) were dissected on the medial aspect of the leg based on a pedicle of the saphenous nerve, saphenous artery, great saphenous vein, and the surrounding fascial tissues. Animals in the experiment were divided into five groups with different circulatory patterns of pedicle dissections. In group I (N = 12), the pedicle artery, vein, nerve, and fascia were preserved. In group II (neurocutaneous flap model, N = 24), the saphenous artery was transected and the vein, nerve, and fascia were preserved. In group III (intraneural vascular plexus circulation pattern, N = 12), the saphenous artery and the fascia were transected. In group IV (extraneural vascular plexus circulation pattern, N = 12), the saphenous artery and nerve were transected. In group V (N = 12), the entire pedicle was transected completely. Flap survival was evaluated grossly on postoperative day 7. All flaps survived in group I, but in group II 19 of 24 flaps survived completely, 3 of 24 had partial necrosis, and 2 of 24 were completely necrotic. Complete necrosis was observed in all group III flaps. In group IV, 6 of 12 flaps survived completely, 3 of 12 flaps survived partially, and 3 of 12 flaps were necrotic (p<0.05 vs. group I). Only one flap with partial necrosis survived in group V. In group II, the average survival area was not significantly different from group I (p>0.05). In conclusion, the saphenous neurocutaneous flap in the rat is a reliable microsurgical model. The saphenous neurocutaneous flap is commonly supplied by both the intraneural and extraneural vascular plexus, and although the latter is more important, neither provides sufficient vascular supply on its own. PMID- 10517471 TI - Interferon-gamma improves muscle flap microcirculation in double-strand RNA induced inflammation. AB - Endothelial cell (EC) activation and subsequent expression of leukocyte adhesion molecules are initial events in multiple pathological processes. Viral double strand ribonucleic acid (dsRNA) induces EC adhesion protein expression and leukocyte adhesion in vitro. Interferon-gamma (IFN-gamma) has been demonstrated to modulate the expression of certain adhesion proteins. The purpose of this study was to measure the inflammatory response to a viral mimetic--a synthetic dsRNA, polyinosinic-polycytidylic acid (poly-I:C)-on the microcirculation of a muscle flap in a rat model and to determine whether IFN-gamma attenuated the response. Two-stage surgery to create a cremaster muscle end-organ tube flap was performed on 18 male Sprague-Dawley rats in three groups. After intra-arterial injection into the abdominal aorta, the reagents (phosphate-buffered saline bovine serum albumin [PBS-BSA] in groups I and II, and IFN-gamma in group III) were kept for 1 hour in this end-organ system. During the second stage at 16 hours, after injection into the penile vein (PBS-BSA in group I, poly-I:C in groups II and III), the flap was prepared for intravital microscopic measurement. The following parameters were measured: red blood cell velocity; vessel diameter; number of functional capillaries; and number of rolling, sticking, and transmigrating neutrophils and lymphocytes. Wilcoxon's rank sum test was used for statistical comparison. Poly-I:C caused a 70% increase in the main artery diameter and a 7% increase in velocity. But as a consequence of dynamic activation of leukocyte interaction, a 30% drop in functional capillary perfusion was observed. Injury to the entire vascular endothelium was confirmed by a 160% increase in transmigrating leukocytes. Treatment with IFN-gamma inhibited the poly-I:C-induced inflammation, as shown by 88%, 63%, and 85% decreases in rolling, sticking, and transmigrating leukocytes respectively, and by a 28% increase in capillary perfusion. Treating the system with IFN-gamma in advance, inhibited poly-I:C-induced inflammation, shown by marked decreases in rolling, adhering, and transmigrating leukocytes, and a notable increase in perfused capillaries. These observations reflect an inhibitory effect of IFN-gamma on leukocyte adhesion molecule expression in vascular endothelium in response to dsRNA in a muscle flap at the microcirculatory level. PMID- 10517472 TI - Microsurgical replantation of a partial ear, with leech therapy. AB - Ear reconstruction is very difficult to perform and often results in a devastating deformity. The use of microsurgical replantation techniques has allowed very favorable aesthetic results. The authors report a case of partial ear replantation without venous repair with the use of medicinal leeches to decompress the acute venous congestion that occurred during postoperative care. Medicinal leech therapy can be very useful in partial ear replantation in cases with no venous repair. PMID- 10517473 TI - A case of atypical McCune-Albright syndrome requiring optic nerve decompression. AB - McCune-Albright syndrome (MAS) is a disease of noninheritable, genetic origin defined by the triad of cafe-au-lait pigmentation of the skin, precocious puberty, and polyostotic fibrous dysplasia. This syndrome, which affects young girls primarily, has also been reported with other endocrinopathies, and rarely with acromegaly and hyperprolactinemia. The fibrous dysplasia in MAS is of the polyostotic type and, apart from the characteristic sites such as the proximal aspects of the femur and the pelvis, the craniofacial region is frequently involved. A male patient with MAS presented with juvenile gigantism, precocious puberty, pituitary adenoma-secreting growth hormone and prolactin, hypothalamic pituitary gonadal and thyroid dysfunction, and polyostotic fibrous dysplasia causing optic nerve compression. Visual deterioration and its surgical management are presented. PMID- 10517474 TI - Multiple glomus tumor: a case report and review of the literature. AB - Multiple glomus tumors are extremely rare and differ from the more common solitary glomus tumors in their clinical presentation and histological features. The authors report a case of multiple glomus tumors of the right hand in a 65 year-old man, its treatment, and a review of the features of this uncommon tumor. The tumor usually presents as a painful, firm, purplish, solitary nodule of the extremities, especially in the nail bed. Multiple glomus tumors are described as softer, more compressible, bluish nodules and they occur with less frequency than solitary tumors. They are often inherited in an autosomal dominant pattern. The authors present a case of multiple glomus tumors of the right hand, in which many small, painful, red papules were grouped in the right hypothenar region. The patient was treated by wide excision of the lesion and coverage of the defect with an ulnar artery forearm flap. PMID- 10517475 TI - Application of artificial dermis prior to full-thickness skin grafting for resurfacing the nose. AB - Two patients with nasal skin defects resulting from excision of rhinophyma and multiple angiofibromas were treated with artificial dermis followed by full thickness skin grafts taken from the postauricular region. The secondary skin grafts took completely in both patients, and the postoperative results were excellent. Although a two-stage operation is required, application of artificial dermis prior to full-thickness skin grafting is a reliable method for resurfacing the nose. PMID- 10517476 TI - Bartsocas-Papas syndrome with fusion of the lips and posterior fusion defects of the thoracic vertebrae. AB - Bartsocas-Papas syndrome is a rare popliteal pterygial syndrome with multiple anomalies including microcephaly, facial clefts, filiform bands, ankyloblepharon, syndactyly, and other ectodermal anomalies. Affected infants usually die perinatally. The authors present an 8-month-old female infant with manifestations of this syndrome and some additional traits including fusion of the lips, intraoral filiform bands, alopecia totalis, and posterior fusion failure of the vertebrae. The fused lips were opened by incising the fibrotic bands closing her mouth. Details of this patient and a brief review of the literature is presented. PMID- 10517477 TI - Augmentation mammaplasty: the story before the silicone bag prosthesis. AB - Czerny from Heidelberg is generally accepted to have performed the first augmentation mammaplasty in 1895. Since then, a variety of nonsilicone materials have been injected or implanted to augment or to reconstruct the hypoplastic female breast, including autologous tissues, intramammary- or submammary-injected alloplastic materials, and preformed alloplastic materials other than silicone. For various reasons outlined in this review, none was fully acceptable. The introduction of the medical-grade silicone bag prosthesis in the early 1960s improved the results of mammary augmentation dramatically and reduced the incidence of fibrous contracture and implant extrusion. Other methods of breast augmentation became obsolete. PMID- 10517478 TI - Skin-sparing mastectomy with Sun flap closure. AB - Skin-sparing mastectomy lends itself to immediate reconstruction with autologous tissue, allowing a one-stage reconstruction (a nipple-areolar complex is made at the time of the initial surgery). It has been difficult to obtain equally pleasing results when immediate reconstruction with a skin-sparing technique is performed with expanders. The authors describe a modification of previously described techniques that enhance the aesthetic result of expander/implant reconstruction. This modification is called the sun flap closure. PMID- 10517479 TI - Spilling editorial blood. PMID- 10517480 TI - Glans and penile skin amputation as a complication of circumcision. PMID- 10517481 TI - A female patient with frontonasal dysplasia sequence and frontonasal encephalocele. PMID- 10517482 TI - Multiple agminated Spitz nevi of the scalp. PMID- 10517483 TI - Hemorheological alterations in patients with chronic renal failure. Effect of hemodialysis. AB - The rheological profile of 45 patients who had kidney failure and were undergoing periodical hemodialysis was studied to ascertain whether they showed rheological alterations and if these alterations improved after hemodialysis. Our results indicate that the patients studied suffer rheological alterations with respect to the control group, specifically increased plasma fibrinogen (436 +/- 119 vs 271 +/- 43 mg/dl, p < 0.001), increased plasma viscosity (1.30 +/- 0.10 vs 1.22 +/- 0.05 mPa.s, p < 0.001), higher erythrocyte aggregability (8.3 +/- 1.2 vs 7.4 +/- 1.4, p < 0.01) and a lower hematocrit (31.2 +/- 5.9 vs 43.4 +/-3.7, p < 0.001). In contrast, erythrocyte deformability did not seem to be altered with respect to the control group. The analyses carried out immediately after hemodialysis showed that in general the rheological profile worsened during this treatment; there was a significant increase in fibrinogen and in plasma viscosity. The rheological alterations detected may play a role in the development of the atherosclerosis processes often experienced by these patients. PMID- 10517484 TI - Measurement of leukocyte rheology in vascular disease: clinical rationale and methodology. International Society of Clinical Hemorheology. AB - The measurement of leukocyte rheology in vascular disease is a recent development with a wide range of new opportunities. The International Society of Clinical Hemorheology has asked an expert panel to propose guidelines for the investigation of leukocyte rheology in clinical situations. This article first discusses the mechanical, adhesive and related functional properties of leukocytes (especially neutrophils) which influence their circulation, and establishes the rationale for clinically-related measurements of parameters which describe them. It is concluded that quantitation of leukocyte adhesion molecules, and of their endothelial receptors may assist understanding of leukocyte behaviour in vascular disease, along with measurements of flow resistance of leukocytes, free radical production, degranulation and gene expression. For instance, vascular cell adhesion molecule (VCAM-1) is abnormally present on endothelial cells in atherosclerosis, diabetes mellitus and inflammatory conditions. Soluble forms of intercellular adhesion molecule (ICAM-1) or VCAM can be found elevated in the blood of patients with rheumatoid arthritis or infections disease. In the second part of the article, possible technical approaches are presented and possible avenues for leukocyte rheological investigations are discussed. PMID- 10517485 TI - Intramuscular oxygen partial pressure in the healthy during exercise. AB - The oxygen partial pressure (pO2) in the anterior tibial muscle was measured in n=12 (6 physically active and 6 sedentary) apparently healthy subjects. This was the first time a flexible micro catheter with an outer diameter of 0.45 mm was used during skeletal muscular activity in men. A two level tread mill test which is used in the diagnosis of peripheral arterial occlusive disease was chosen to induce physical stress. In the healthy volunteers a pO2 increase was noted at the beginning of exercise. This was followed by a pO2 decrease because of an increased O2 demand in the working muscle. The initial pO2 increase was thought to be due to the recruitment of capillaries and not the subsequently increased heart rate. At rest and during activity pO2 values were higher in physically active subjects than in the sedentary and the exercise induced decrease of pO2 values was slower and in addition to this the compensation to baseline values quicker. PMID- 10517486 TI - Involvement of erythrocyte aggregation and erythrocyte resistance to flow in acute coronary syndromes. AB - The objective of the study was to identify the relative importance of erythrocyte flow resistance and aggregation in acute and chronic coronary syndromes. 117 subjects in five groups were studied: (1) 34 patients shortly after acute myocardial infarction (AMI) before reperfusion therapy; (2) 27 patients with unstable and (3) 21 with stable angina pectoris (AP); (4) 14 age-matched control patients and (5) 21 healthy volunteers. Single erythrocyte transit times were measured using the Cell Transit Analyser. Shear dependent elongation and aggregation was measured by a modified computerized Myrenne aggregometer. Leukocyte count was increased in coronary artery disease (CAD), especially in acute syndromes (mean +/- SD for groups 1-5): 12.2 +/- 4.5; 10.0 +/- 5.4; 8.0 +/- 2.0; 8.0 +/- 3.7; 7.0 +/- 2.0 (pl(-1))). Platelets, hematocrit, fibrinogen, alpha2-macroglobulin did not differ between the groups. Plasma viscosity (mPas) was elevated in AMI and stable AP: 1.34 +/- 0.10; 1.30 +/- 0.09; 1.32 +/- 0.08; 1.27 +/- 0.07; 1.27 +/- 0.05. Erythrocyte filtrability was not different as was the shear dependent deformation. Aggregation parameters such as gammaTmin were elevated in CAD: 180 +/- 70; 159 +/- 60; 166 +/- 59; 115 +/- 43; 113 +/- 51 (s( 1)). Erythrocyte deformability, measured with two independent methods, does not appear to contribute to the pathophysiology of acute coronary syndromes. Erythrocyte aggregation and plasma viscosity were again found increased both in unstable and stable coronary disease. It is unlikely that increased red cell aggregation contributes to emergence of AMI. PMID- 10517487 TI - Elasticity and fracture strain of whole blood clots. AB - The measurement of clot elasticity and fracture strain is carried out using a rotational rheometer with a controlled stress system. The elasticity is quantified by a shear elastic modulus and fracture strain by a critical strain (deformation) when the clot begins to break up. The results indicate a decrease of elasticity and increase of fracture strain of the clot with increasing hematocrit. Moreover, the elastic modulus of the clot is not constant but increasing with deformation. PMID- 10517488 TI - Donor lymphocyte infusions for relapse of chronic myeloid leukemia after allogeneic stem cell transplant: where we now stand. AB - The infusion of lymphocytes from the original marrow donor (donor lymphocyte infusion [DLI]) reinduces complete remission in a high percentage of patients with chronic myeloid leukemia (CML) who relapse after allogeneic stem cell transplant, and thus, is probably the best initial approach to their management. The major predictive factor for response is the disease stage at time of treatment, because patients in molecular or cytogenetic relapse fare better than those in hematologic relapse. Moreover, patients with a short interval between transplant and DLI have a higher probability of response than those with longer intervals. The durability of DLI-induced remissions has not yet been established, but they appear to be prolonged. The observation that DLI can be highly effective for patients in relapse has encouraged the recent development of new strategies designed to minimize the myeloablative regimen and exploit the immunotherapeutic component of the transplant. The principal complication associated with use of DLI is the occurrence of graft-versus-host disease (GVHD). Several approaches have been tested to reduce the incidence or impact of GVHD, based on the ex vivo depletion of alloreactive donor cells or the use of donor T cells transduced with a suicide gene. The incidence of GVHD can also be reduced by starting with low doses of donor cells and "escalating" subsequent doses as required. However, the identification of selective targets for leukemia-reactive immunity is probably the optimal strategy to resolve the problem of GVHD. Although currently minor histocompatibility antigens appear to be the most likely targets for DLI, several groups are focusing on the generation of leukemia-specific immunity. The results obtained by use of tumor-associated antigens presented by dendritic cells are encouraging and may lay the foundations for the use of adoptive immunotherapy in the autologous setting. PMID- 10517489 TI - Ganglioside GD1b supresses immunoglobulin production by human peripheral blood mononuclear cells. AB - Gangliosides are sialic acid-containing glycolipids, that have various immunomodulatory effects. We previously reported that various gangliosides in vitro either inhibited or enhanced spontaneous immunoglobulin (Ig) production by human peripheral blood mononuclear cells (PBMC). GD1b was one of the inhibitory gangliosides. In this study, we further examined the mechanism for the inhibitory effect of GD1b. The inhibitory effect of GD1b was revealed at 0.1 microM, increased dose dependently, and was maximized at 10 microM, which reduced spontaneous IgG, IgM, and IgA production of human PBMC by 50.5%, 52.0%, and 48.3% compared with controls, respectively. GD1b did not affect the proliferation and viability of PBMC. GD1b did not alter Ig production of B cells alone. Interleukin 6 (IL-6) and IL-10 each partially reversed the GD1b-induced inhibition of Ig production by PBMC, and the addition of both cytokines completely reversed the inhibition. When endogenous IL-6 and IL-10 were neutralized by specific antibodies, GD1b did not reveal inhibitory effects on the Ig production. GD1b inhibited IL-6 and IL-10 production of CD4+ T cells, without affecting those of CD8+ T cells, monocytes, or B cells. When CD4+ T cells were preincubated with GD1b and washed and cultured with B cells and monocytes, Ig production was also suppressed. These results suggest that GD1b may indirectly suppress Ig production of B cells in whole PBMC by reducing IL-6 and IL-10 production of CD4+ T cells. GD1b may act as an important inhibitor of human humoral immune responses. PMID- 10517490 TI - Cyclooxygenase-2 is essential for normal recovery from 5-fluorouracil-induced myelotoxicity in mice. AB - Cyclooxygenase (COX) plays a key regulatory role in prostaglandin synthesis. COX 2 is inducible and is the major isoform of inflammatory cells. COX-2-deficient mice were shown to have normal basal hematopoiesis and hematology. We hypothesized that COX-2 induction plays a role in the recovery phase of 5 fluorouracil (5-FU) induced bone marrow injury, because significant macrophage driven phagocytic removal of necrotic debris and stromal cell reorganization of repopulating marrow occur after 5-FU induction of bone marrow necrosis. Hematologic recovery was markedly delayed with moderately severe leukopenia, thrombocytopenia and reticulocytopenia compared to heterozygotes on day 8 or 12 in Cox-2-/- mice. Mild anemia was present in 5-FU-treated Cox-2-/- and Cox-2+/- mice on days 8 and 12, which was more severe in Cox-2-/- mice. Cox-2-/- mice had markedly decreased bone marrow cell counts per femur and reduced numbers of erythroid and myeloid colony-forming cells compared to heterozygote mice on days 8 and 12 post 5-FU. Histologic examination of 5-FU-treated Cox-2-/- mice revealed a failure to repopulate the intact marrow stroma with hematopoietic cells. Accelerated erythropoiesis following phenylhydrazine-induced hemolytic anemia, however, was comparable between Cox-2-/- and Cox+/- mice, as were induced levels of renal erythropoietin mRNA. COX-2 induction is likely a central event in the accelerated hematopoiesis following myelotoxic injury, because recovery from 5-FU induced myeloablation is markedly impaired in Cox-2-/- mice but is normal after phenylhydrazine induction of anemia. PMID- 10517491 TI - Nonmyeloablative conditioning to induce bilateral tolerance after allogeneic bone marrow transplantation in mice. AB - We recently described a new nonmyeloablative method to induce stable and specific transplantation tolerance to allogeneic tissues in adult mice. It included total lymphoid irradiation (TLI) of recipients with six fractions of 200 cGy each, inoculation with donor bone marrow (BM) cells and cyclophosphamide (Cy) for selective elimination or inactivation of residual donor-reactive cells of the host, and infusion with T-cell depleted donor BM cells after Cy. Here, we investigated the possibility to induce stable bilateral graft-vs-host and host-vs graft transplantation tolerance using non-T-cell depleted allogeneic BM. Our results show that the dose of BM required for the induction of transplantation tolerance was inversely correlated with the intensity of the conditioning. Transfer of a low dose (3 x 10(6)) of total donor BM cells to recipients preconditioned with a less intensive regimen (two or three TLI fractions instead of six) diminished graft-vs-host disease (GVHD)-related mortality of recipients to 40% and converted 89% of the survivors into GVHD-free mixed hematopoietic chimeras that maintained donor skin allografts >180 days. A tenfold increase in the number of donor BM cells (3 x 10(7) instead of 3 x 10(6)) reduced the rate of GVHD-related mortality of recipients to 20% and resulted in bilateral transplantation tolerance in 100% of nonirradiated survivors. PMID- 10517492 TI - Human CD34+ hematopoietic cells transduced by retrovirus-mediated interferon alpha gene maintains regeneration capacity and engraftment in NOD/SCID mice. AB - To achieve long-term expression of human interferon alpha-5 (IFNalpha) gene in the bone marrow (BM) hematopoietic microenvironment, replication-deficient retroviral vector LSN-IFNalpha was used to deliver the IFNalpha gene into human BM CD34+ cells. After fibronectin-facilitated transduction, a fraction of CD34+ cells was plated in methylcellulose medium with or without G418 to assess transduction efficiency and the effect of IFNalpha gene transfer on colony formation. Colony-forming assay in the presence of G418 (400 microg/mL) revealed that 41% CFU-GM colonies are G418 resistant after infection with LSN-IFNalpha retrovirus. There was no significant difference in CFU-GM/BFU-E colony formation among IFNalpha gene-transduced CD34+ cells, control vector (LXSN) transduced CD34+ cells and nontransduced CD34+ cells. Another portion of CD34+ cells was grown in liquid medium to measure IFNalpha production. RIA revealed that IFNalpha gene-transduced CD34+ cells produced 72.2 +/- 15.4 U/mL (10(6) cells/24 hours) of IFNalpha compared with 8.3 +/- 2.1 U/mL and 4.3 +/- 1.2 U/mL in LXSN-transduced or nontransduced CD34+ cells, respectively. The remaining portion of transduced CD34+ cells was transplanted into immunodeficient (NOD/SCID) mice to allow analysis of long-term expression of IFNalpha. Transplantation of 1x10(6) CD34+ cells into sublethally irradiated NOD/SCID mice showed that IFNalpha and neo(r) mRNA were detectable in engrafted mouse BM cells for up to 6 months. We conclude that continual local expression of IFNalpha in transduced CD34+ cells does not impair either CD34+ cell growth and differentiation or engraftment and long-term survival in NOD/SCID mice. PMID- 10517494 TI - Measles virus nucleocapsid transcript expression is not restricted to the osteoclast lineage in patients with Paget's disease of bone. AB - Abundant evidence supports a viral etiology for Paget's disease of bone (PD), however, an infectious virus has not been isolated from PD patients. Thus, it is unclear how the virus is maintained for the many years that the disease persists in patients. We considered if a primitive multipotential hematopoietic stem cell (HSC), which is self-renewing, passes the virus to its differentiated progeny and serves as a reservoir for the pathogen. If a primitive stem cell harbored measles virus (MV), then other hematopoietic lineages derived from this stem cell in PD patients should also express MV transcripts. Therefore, because the human hematopoietic stem cell has not been clearly identified or isolated in large numbers, we isolated RNA from highly purified erythroid and multipotential hematopoietic progenitors that are the precursors for erythroid, granulocyte, megakaryocyte and macrophages (CFU-GEMM), and used RT-PCR to determine if MV nucleocapsid transcripts were present. MV transcripts were detected in PD patients in early erythroid (BFU-E) and more primitive multipotential myeloid progenitors (CFU-GEMM). Nonhematopoietic stromal cells from PD patients did not express MV transcripts. The expression of MV transcripts in erythroid progenitors was further confirmed by in situ hybridization using antisense riboprobes to MV nucleocapsid transcripts. Thus, our findings suggest that the pluripotent HSCs may be a potential reservoir for the virus. We propose that when HSCs, which contain MV, divide they produce a second HSC that serves as a reservoir for the virus and also transmit the virus to their more differentiated progeny in the erythroid and myeloid lineages. This mechanism would permit a defective virus to persist in HSCs of PD patients for many years, since HSCs are usually in G0 phase, and then be transmitted to more differentiated cells. This model further suggests that a mature complete virus that affects cell function could only act pathogenetically in the osteoclast lineage, which offers a permissive milieu. PMID- 10517493 TI - Fas ligand is highly expressed in acute leukemia and during the transformation of chronic myeloid leukemia to blast crisis. AB - Fas ligand (FasL) induces apoptosis in susceptible Fas-bearing cells and is critically involved in regulating T-cell immune responses. It is highly expressed in several human malignancies, and a role in the suppression of antitumor immune responses has been suggested. We evaluated FasL expression in leukemia and normal hematopoietic cells. By Western blotting, all acute leukemic cell lines (n = 9) and primary samples of acute leukemic marrow (n = 4) revealed high levels of FasL. In contrast, much weaker signals were observed in samples of normal marrow (n = 5), and either weak or intermediate expression was seen in chronic myeloid leukemia (CML) in chronic phase (n = 7). Additional leukemic samples were examined by immunohistochemistry. Staining for FasL was negative in 7 of 9 cases of chronic-phase CML, whereas all cases of CML in blast crisis (n = 6), acute lymphoblastic leukemia (n = 6), and acute myeloid leukemia (n = 11) stained strongly in 60 to 100% of nucleated cells. FasL+ leukemic cell lines did not trigger Fas-mediated apoptosis in either Jurkat cells or activated human T lymphocytes, possibly related to the intracellular location of the ligand. Western analysis of normal marrow subpopulations revealed that most FasL in marrow mononuclear cells was expressed by CD7+ lymphocytes. FasL also was strongly expressed in CD34+ hematopoietic progenitor cells from both normal and chronic-phase CML marrow, suggesting a correlation with primitive maturation stage. In summary, high levels of FasL expression were associated with aggressive biologic behavior in leukemia, including transformation of CML to blast crisis. This could potentially represent a response to loss of proapoptotic Fas signaling, which is known to occur in acute leukemic blasts. PMID- 10517495 TI - Natural killer cell cytotoxicity of breast cancer targets is enhanced by two distinct mechanisms of antibody-dependent cellular cytotoxicity against LFA-3 and HER2/neu. AB - Treatment of advanced breast cancer with autologous stem cell transplantation is limited by a high probability of disease relapse. In clinical trials, interleukin 2 (IL-2) alone can expand natural killer (NK) cells in vivo and increase their cytotoxic activity against breast cancer cell lines, but this increase is modest. Understanding the mechanisms that mediate NK cell lysis of breast cancer targets may lead to improvements of current immunotherapy strategies. NK cells from normal donors or patients receiving subcutaneous IL-2 were tested in cytotoxicity assays against five breast cancer cell lines. The role of adhesion molecules and antibodies that interact through Fc receptors on NK cells was explored. NK cell lysis of breast cancer targets is variable and is partially dependent on recognition through ICAM-1 and CD18. While blocking CD2 slightly decreased cytotoxicity, contrary to expectations, an antibody against CD58 (the ligand for CD2), failed to block killing and instead mediated an increased cytotoxicity that correlated with target density of CD58. The CD58 antibody-enhanced killing was dependent not only on FcRgammaIII but also on CD2 and ICAM-1/CD18. To further elucidate the mechanism of this CD58 antibody-dependent cellular cytotoxicity (ADCC), another antibody was tested. Trastuzumab (Herceptin), a humanized antibody against HER2/neu, mediated potent ADCC against all the HER2/neu positive breast cancer targets. Unlike CD58 antibody-mediated ADCC, Herceptin ADCC was minimally affected by blocking antibodies to CD2 or ICAM-1/CD18, which suggests a different mechanism of action. This study shows that multiple mechanisms are involved in NK cell lysis of breast cancer targets, that none of the targets are inherently resistant to killing, and that two distinct mechanisms of ADCC can target immunotherapy to breast cancer cells. PMID- 10517496 TI - Thrombopoietin induces an SH2-containing protein, CIS1, which binds to Mpl: involvement of the ubiquitin proteosome pathway. AB - The interaction of thrombopoietin (TPO) with its receptor, Mpl, triggers growth and differentiation of megakaryocytes and their progenitors. The Mpl cytoplasmic domain controls this process through src homology 2 (SH2)-containing target molecules and their receptor docking sites. A novel cytokine inducible SH2 containing protein, CIS1, has been isolated. CIS1 is induced by interleukin-2 (IL 2), IL-3, GM-CSF, and erythropoietin (EPO), but not by IL-6, granulocyte colony stimulating factor (G-CSF), or stem cell factor. To investigate the functional domains of Mpl for induction of CIS1, we examined FDCP-2 cell lines expressing seven carboxyl truncations of the human Mpl cytoplasmic domain. We found that the box1 and box2 regions of Mpl were necessary for induction of CIS1 after TPO stimulation. CIS1 was degraded very quickly and was found to be involved in the ubiquitin-proteosome pathway. A 4-hour depletion of TPO almost completely eliminated CIS1 protein; within 1 hour after TPO stimulation, CIS1 protein reappeared as 37- and 32-kDa proteins in the wild type Mpl-expressing FDCP-2 cells. Further, CIS1 was stably associated with tyrosine-phosphorylated Mpl. The SH2 domains of CIS1, constructed as glutathione S-transferase fusion protein, bound to activated Mpl in vitro. These results suggest that CIS1 may be an important signaling component downstream of Mpl and may regulate the proliferation and differentiation of hematopoietic cells. PMID- 10517497 TI - Ex vivo pharmacological purging of leukapheresis collections with nitrogen mustard: amifostine pretreatment improves both early and late peripheral blood progenitor cell recovery. AB - Ex vivo pharmacological purging of bone marrow has been used to eliminate clonogenic tumor cells contaminating the autograft and potentially responsible of relapse. A considerable improvement of pharmacological purging would be achieved only if normal marrow progenitor cells could be selectively protected by the cytotoxicity of these agents. Amifostine (WR-2721; Ethyol), a phosphorylated aminothiol compound, has been shown to have this property both in vivo and in vitro. We describe here, an experimental model for ex vivo purging of peripheral blood progenitor cell (PBPC) collections based on the combination of 3 mg/ml of amifostine and the alkylating agent nitrogen mustard. Amifostine pretreatment resulted in a statistically significant protection of normal late and early progenitor cells. Under the same experimental conditions, we observed a 4-6 log reduction of contaminating leukemic cells (i.e., K-562 and CEM) and in contrast to the protection of normal peripheral blood progenitor cells, preincubation of contaminating K-562 or CEM with amifostine did not significantly alter the LD95 nitrogen mustard concentration. Moreover, when we tested fresh human leukemia progenitor cells, amifostine pretreatment sensitized the leukemic cells to the cytotoxic effects of NM. PMID- 10517498 TI - Myelopoietin, a chimeric agonist of human interleukin 3 and granulocyte colony stimulating factor receptors, mobilizes CD34+ cells that rapidly engraft lethally x-irradiated nonhuman primates. AB - Myelopoietin (MPO), a multifunctional agonist of interleukin 3 and granulocyte colony-stimulating factor (G-CSF) receptors, was evaluated for its ability to mobilize hematopoietic colony-forming cells (CFC) and CD34+ cells relative to control cytokines in normal nonhuman primates. Additionally, the engraftment potential of MPO-mobilized CD34+ cells was assessed in lethally irradiated rhesus monkeys. Normal rhesus monkeys were administered either MPO (200 microg/kg/day), daniplestim (a high-affinity interleukin 3 receptor agonist) (100 microg/kg/day), G-CSF (100 microg/kg/day), or daniplestim coadministered with G-CSF (100 microg/kg/day each), subcutaneously for 10 consecutive days. The mobilization kinetics were characterized by peripheral blood (PB) complete blood counts, hematopoietic CFC [granulocyte-macrophage CFC (GM-CFC), megakaryocyte CFC (MK CFC)], and the immunophenotype (CD34+ cells) of PB nucleated cells prior to and on day 3 to days 7, 10, 12, and 14, and at intervals up to day 28 following initiation of cytokine administration. A single large-volume leukapheresis was conducted on day 5 in an additional cohort (n = 10) of MPO-mobilized animals. Eight of these animals were transplanted with two doses of CD34+ cells/kg. A maximum 10-fold increase in PB leukocytes (white blood cells) (from baseline 7.8 12.3 x 10(3)/microL to approximately 90 x 10(3)/microL) was observed over day 7 to day 10 in the MPO, G-CSF, or daniplestim+G-CSF cohorts, whereas daniplestim alone stimulated a less than onefold increase. A sustained, maximal rise in PB derived GM-CFC/mL was observed over day 4 to day 10 for the MPO-treated cohort, whereas the daniplestim+G-CSF, G-CSF alone, and daniplestim alone treated cohorts were characterized by a mean peak value on days 7, 6, and 18, respectively. Mean peak values for PB-derived GM-CFC/mL were greater for MPO (5,427/mL) than for daniplestim+G-CSF (3,534/mL), G-CSF alone (3,437/mL), or daniplestim alone (155/mL) treated cohorts. Mean peak values for CD34+ cells/mL were noted within day 4 to day 5 of cytokine administration: MPO (255/microL, day 5), daniplestim+G CSF (47/microL, day 5), G-CSF (182/microL, day 4), and daniplestim (96/microL, day 5). Analysis of the mobilization data as area under the curve indicated that for total CFCs, GM-CFC, MK-CFC, or CD34+ cells, the MPO-treated areas under the curve were greater than those for all other experimental cohorts. A single, large volume (3.0 x blood volume) leukapheresis at day 5 of MPO administration (PB: CD34+ cell/microL = 438 +/- 140, CFC/mL = 5,170 +/- 140) resulted in collection of sufficient CD34+ cells (4.31 x 10(6)/kg +/- 1.08) and/or total CFCs (33.8 x 10(4)/kg +/- 8.34) for autologous transplantation of the lethally irradiated host. The immunoselected CD34+ cells were transfused into autologous recipients (n = 8) at cell doses of 2 x 10(6)/kg (n = 5), and 4 x 10(6)/kg (n = 3) on the day of apheresis. Successful engraftment occurred with each cell dose. The data demonstrated that MPO is an effective and efficient mobilizer of PB progenitor cells and CD34+ cells, such that a single leukapheresis procedure results in collection of sufficient stem cells for transplantation and long term engraftment of lethally irradiated hosts. PMID- 10517499 TI - Cotransplantation of stroma results in enhancement of engraftment and early expression of donor hematopoietic stem cells in utero. AB - Although promising, clinical and experimental efforts at in utero hematopoietic stem cell (HSC) transplantation currently are limited by minimal donor cell engraftment and lack of early donor cell expression after transplantation. We reasoned that cotransplantation of stromal elements (ST) might condition the fetal microenvironment for the engraftment of donor HSC and facilitate precocious bone marrow (BM) hematopoiesis. In this study we cotransplanted sheep ST, derived from adult or fetal BM, with either adult or fetal HSC, into preimmune fetal sheep. We analyzed donor cell chimerism in BM and peripheral blood and compared levels of chimerism achieved with recipients of HSC alone. In all experimental groups, stromal cotransplantation markedly increased the level of peripheral blood donor cell expression at 60 days after transplantation relative to controls. Adult BM-derived stroma cotransplanted with adult HSC provided the highest levels of circulating donor cells, whereas fetal-derived stroma was less effective. In addition, ST cotransplantation resulted in increased donor cell engraftment in the BM and led to significantly increased levels of donor hematopoiesis for over 30 months after transplant. Cotransplantation of stroma may represent a valuable clinical strategy for optimal application of in utero HSC transplantation. PMID- 10517500 TI - Chordoid glioma of the third ventricle: immunohistochemical and molecular genetic characterization of a novel tumor entity. AB - Chordoid glioma of the third ventricle was recently reported as a novel tumor entity of the central nervous system with characteristic clinical and histopathological features (Brat et al., J Neuropathol Exp Neurol 57: 283-290, 1998). Here, we report on a histopathological, immunohistochemical and molecular genetic analysis of five cases of this rare neoplasm. All tumors were immunohistochemically investigated for the expression of various differentiation antigens, the proliferation marker Ki-67, and a panel of selected proto-oncogene and tumor suppressor gene products. These studies revealed a strong expression of GFAP, vimentin, and CD34. In addition, most tumors contained small fractions of neoplastic cells immunoreactive for epithelial membrane antigen, S-100 protein, or cytokeratins. The percentage of Ki-67 positive cells was generally low (<5%). All tumors showed immunoreactivity for the epidermal growth factor receptor and schwannomin/merlin. There was no nuclear accumulation of the p53, p21 (Waf-1) and Mdm2 proteins. To examine genomic alterations associated with the development of chordoid gliomas, we screened 4 tumors by comparative genomic hybridization (CGH) analysis. No chromosomal imbalances were detected. More focussed molecular genetic analyses revealed neither aberrations of the TP53 and CDKN2A tumor suppressor genes nor amplification of the EGFR, CDK4, and MDM2 proto-oncogenes. Our data strongly support the hypothesis that chordoid glioma of the third ventricle constitutes a novel tumor entity characterized by distinct morphological and immunohistochemical features, as well as a lack of chromosomal and genetic alterations commonly found in other types of gliomas or in meningiomas. PMID- 10517501 TI - No complementation between TP53 or RB-1 and v-src in astrocytomas of GFAP-v-src transgenic mice. AB - Human low-grade astrocytomas frequently recur and progress to states of higher malignancy. During tumor progression TP53 alterations are among the first genetic changes, while derangement of the p16/p14ARF/RB-1 system occurs later. To probe the pathogenetic significance of TP53 and RB-1 alterations, we introduced a v-src transgene driven by glial fibrillary acidic protein (GFAP) regulatory elements (which causes preneoplastic astrocytic lesions and stochastically astrocytomas of varying degrees of malignancy) into TP53+/- or RB-1+/- mice. Hemizygosity for TP53 or RB-1 did not increase the incidence or shorten the latency of astrocytic tumors in GFAP-v-src mice over a period of up to 76 weeks. Single strand conformation analysis of exons 5 to 8 of non-ablated TP53 alleles revealed altered migration patterns in only 3/16 tumors analyzed. Wild-type RB-1 alleles were retained in all RB-1+/-GFAP-v-src mice-derived astrocytic tumors analyzed, and pRb immunostaining revealed protein expression in all tumors. Conversely, the GFAP-v-src transgene did not influence the development of extraneural tumors related to TP53 or RB-1 hemizygosity. Therefore, the present study indicates that neither loss of RB-1 nor of TP53 confer a growth advantage in vivo to preneoplastic astrocytes expressing v-src, and suggests that RB-1 and TP53 belong to one single complementation group along with v-src in this transgenic model of astrocytoma development. The stochastic development of astrocytic tumors in GFAP v-src, TP53+/- GFAP-v-src, and RB-1+/- GFAP-v-src transgenic mice indicates that additional hitherto unknown genetic lesions of astrocytes contribute to tumorigenesis, whose elucidation may prove important for our understanding of astrocytoma initiation and progression. PMID- 10517502 TI - Homozygous deletions of the CDKN2C/p18INK4C gene on the short arm of chromosome 1 in anaplastic oligodendrogliomas. AB - Allelic deletions of the short arm of chromosome 1 are common in oligodendrogliomas and have been correlated with chemosensitivity and better prognosis in patients with high-grade oligodendrogliomas. In these tumors, 1p loss is also inversely related to deletions of the CDKN2A gene on 9p, which encodes the key cell cycle regulatory molecule p16INK4A. Because the CDKN2C gene, which encodes the homologous p18INK4C cell cycle regulatory protein, maps to chromosomal band 1p32, CDKN2C is an attractive candidate for the oligodendroglioma suppressor gene on chromosome 1. To evaluate this possibility, we studied 39 high-grade oligodendrogliomas for homozygous deletions and point mutations of the CDKN2C gene, as well as for allelic loss of 1p. Although no mutations were detected in the CDKN2C coding region, two tumors had homozygous deletions that involved CDKN2C. Interestingly, these cases did not have CDKN2A gene deletions. Coupled with the recent report of rare point mutations of CDKN2C in oligodendrogliomas, these findings suggest that CDKN2C inactivation may be oncogenic in a small percentage of human oligodendrogliomas. PMID- 10517503 TI - Clonal origin of recurrent meningiomas. AB - Meningiomas are common intracranial and intraspinal tumors. They are treated primarily by surgical resection. Meningioma recurrence following surgery is frequent despite advances in microneurosurgery. However, it is not clear whether recurrent meningiomas, close or distant to the primary resection site, arise from incomplete resection, dissemination of tumor fragments or from independent tumor growth. In order to address the question of clonality in recurring meningiomas, we examined a series of five patients with a total of 14 tumors for X-chromosome inactivation in the tumor tissues. Four patients with a total of 11 meningiomas were informative for polymorphisms either in the PGK or the AR genes. All recurrent meningiomas were found to be clonal with respect to the primary lesions. This finding suggests a common molecular pathogenesis of primary meningioma and subsequent recurrences (p<0.01). In a sixth patient, we analyzed the NF2 gene for mutations in the primary and 5 recurrent meningiomas. All six lesions carried the identical NF2 mutation, strongly indicating a common origin for these tumors. We conclude that recurrent meningiomas usually arise from dissemination of tumor fragments, most likely at the time of the first surgical resection. Our data should alert to the potential of meningioma cells for seeding during surgical procedures. PMID- 10517504 TI - Axonal pathology in multiple sclerosis. A historical note. PMID- 10517505 TI - Tau and synuclein and their role in neuropathology. PMID- 10517506 TI - Tau-positive glial inclusions in progressive supranuclear palsy, corticobasal degeneration and Pick's disease. AB - The presence of tau-positive glial inclusions has been recently found a consistent feature in the brains of patients with progressive supranuclear palsy (PSP), corticobasal degeneration (CBD) and Pick's disease (PiD). These inclusions are classified based on cellular origin as tau-positive astrocytes, presumably either fibrillary or protoplasmic, coiled bodies and glial threads. Immunohistochemically, their major structural component is abnormal tau proteins, similar to those found in Alzheimer's disease. Nevertheless, their morphology, including ultrastructural profile, has been suggested to be distinctive for each disease. The profile and extent of particular glial inclusions correlate well with disease phenotype. Highly characteristic correlations include tufts of abnormal fibers in PSP, astrocytic plaques and dense glial threads in CBD and ramified astrocytes and small Pick body-like inclusions in PiD. The significance of the inclusions in disease pathogenesis and their biochemical characteristics remain to be clarified. Nevertheless, these distinctive glial lesions most likely reflect fundamental alterations in isoform composition of tau as well as its specific cellular and regional expression in sporadic tauopathies. PMID- 10517507 TI - Comparative biochemistry of tau in progressive supranuclear palsy, corticobasal degeneration, FTDP-17 and Pick's disease. AB - Neurodegenerative disorders referred to as tauopathies have cellular hyperphosphorylated tau protein aggregates in the absence of amyloid deposits. Comparative biochemistry of tau aggregates shows that they differ in both phosphorylation and content of tau isoforms. The six tau isoforms found in human brain contain either three (3R) or four microtubule-binding domains (4R). In Alzheimer's disease, all six tau isoforms are abnormally phosphorylated and aggregate into paired helical filaments. They are detected by immunoblotting as a major tau triplet (tau55, 64 and 69). In corticobasal degeneration and progressive supranuclear palsy, only 4R-tau isoforms aggregate into twisted and straight filaments respectively. They appear as a major tau doublet (tau64 and 69). Finally, in Pick's disease, only 3R-tau isoforms aggregate into random coiled filaments. They are characterized by another major tau doublet (tau55 and 64). These differences in tau isoforms may be related to either the degeneration of particular cell populations in a given disorder or aberrant cell trafficking of particular tau isoforms. Finally, recent findings provide a direct link between a genetic defect in tau and its abnormal aggregation into filaments in fronto-temporal dementia with Parkinsonism linked to chromosome 17, demonstrating that tau aggregation is sufficient for nerve cell degeneration. Thus, tau mutations and polymorphisms may also be instrumental in many neurodegenerative disorders. PMID- 10517508 TI - Fibrillogenesis of tau: insights from tau missense mutations in FTDP-17. AB - Frontotemporal dementia and Parkinsonism linked to chromosome 17 (FTDP-17) is a neurological disorder associated with tau pathology.Tau deposits in FTDP-17 brains consist of polymerized filaments of hyperphosphorylated tau, the morphology of which is determined by the nature of the tau gene mutation observed in each case. A number of mutations associated with FTDP-17 have been identified in the 5' splice site of exon 10 and in exons 9-13 of the tau gene. The exon 10 5' splice site mutations disrupt alternative splicing and thus alter the ratio of 4R and 3R Tau isoforms. The majority of Tau missense mutations decrease its ability to bind tubulin and promote microtubule assembly. The extent of reduction varies depending on the site and nature of the mutation. Some Tau missense mutations also have a direct effect on the rate and the extent of tau filament formation. In the presence of polymerization-inducing agents such as heparin or arachidonic acid, mutant tau forms polymers more efficiently than wild type tau in vitro. Tau mutations affect polymerization at both nucleation and elongation phases. One mutation (R406W) is also known to alter the susceptibility of tau to phosphorylation. Expression of mutant tau in cultured cells changes the cytoskeletal integrity of CHO and COS-7 cells, but none of the tau transfected cells display tau filament inclusions. These findings suggest involvement of at least two mechanisms in the pathogenesis of FTDP-17. PMID- 10517509 TI - Alpha-synuclein in Lewy body disease and Alzheimer's disease. AB - Alzheimer's disease (AD) and Lewy body disease (LBD) are the most common causes of dementia in the elderly population. Previous studies have shown that cognitive alterations in these disorders are associated with synaptic loss. Injury and loss of synapses might be associated with altered function of synaptic proteins. Among them, recent studies have shown that abnormal aggregation and accumulation of synaptic proteins, such as alpha-synuclein, might be associated with plaque formation in AD and Lewy body formation in LBD. Further reinforcing the hypothesis that alpha-synuclein plays a major role in the pathogenesis of these disorders, recent work has shown that mutations that alter the conformation of this molecule are associated with familial forms of Parkinson's disease. The mechanisms by which altered function or aggregation of alpha-synuclein might lead to neurodegeneration are not completely clear; however, new evidence points to a potential role for this molecule in synaptic damage and neurotoxicity via amyloid like fibril formation and mitochondrial dysfunction. In this manuscript we review the data linking alpha-synuclein to the pathogenesis of AD and LBD. PMID- 10517510 TI - Multiple system atrophy: a sporadic synucleinopathy. AB - Multiple system atrophy (MSA) is a sporadic neurodegenerative disease characterized clinically by varying degrees of Parkinsonism, cerebellar ataxia and autonomic dysfunction and pathologically by degeneration in the substantia nigra, putamen, olivary nucleus, pontine nuclei and cerebellum. In addition to selective neuronal loss, iron pigment accumulation and gliosis, myelin pathology is increasingly recognized. In affected white matter, myelin displays signs of degeneration and oligodendroglia contain argyrophilic inclusion bodies, so-called glial cytoplasmic inclusions (GCI). GCI are composed of 10-15-nm diameter coated filaments that are immunoreactive for ubiquitin and alpha-synuclein. Similar inclusions are occasionally found in neuronal cell bodies and cell processes in MSA. Given the presence of inclusion bodies composed of synuclein, it is reasonable to assume that biochemical alterations would be detected in synuclein in MSA and indeed this is the case. In MSA synuclein has biophysical properties that suggest increasing insolubility such as sedimentation in dense fractions in sucrose gradients and ready extraction into detergents and formic acid. Surprisingly, these biochemical modifications in synuclein are more widespread in the brain that the obvious pathology and suggest a fundamental molecular characteristic of the disorder. Similar neuronal, and less frequently glial, inclusions are detected in Lewy body disease, where there is also evidence for biophysical alterations in synuclein. Thus, MSA and LBD are both synucleinopathies, and they may comprise different poles of a disease spectrum that includes sporadic disorders as well as genetically determined disorders such as familial Lewy body Parkinsonism. PMID- 10517511 TI - Transgenic models of tauopathies and synucleinopathies. AB - Rapidly emerging concepts about the pathobiology and defining phenotypes of two major classes of neurodegenerative disease known as tauopathies and synucleinopathies are bringing these diseases into shaper focus. Significantly, recent research has substantially advanced understanding of these neurodegenerative disorders thereby providing fresh opportunities for the development of transgenic (TG) mouse models. Since the availability of such animal models will accelerate efforts to discover more effective therapies, we review the current status of efforts to generate informative TG mouse models for tauopathies and synucleinopathies and other neurodegenerative disorders characterized by prominent filamentous brain lesions. PMID- 10517512 TI - April 1999--44 year old man with a bleeding intracerebral tumor. AB - A 44 year-old man presented with a three month history of increasing headache and evolving left sided hemiparesis that culminated in an haemorrhage into an intracerebral tumour which was partially resected. Histologic, immunohistochemical, electron microscopic and molecular studies are supportive of a diagnosis of primary embryonal rhabdomyosarcoma. While primary rhabdomyosarcoma of the central nervous system is rare, and 72% of previously reported cases are in the paediatric population, there appears to be subset of these tumours occurring supratentorially in the adult. PMID- 10517513 TI - May 1999--16 year old male with an unexpected MRI finding. AB - Four years after resection of a supratentorial pilocytic astrocytoma this 16-year old boy displayed widespread leptomeningeal seeding. Although the primary tumor lacked contrast enhancement, the multiple metastatic nodules were markedly contrast enhancing. Both the initial and disseminated tumor were consistent with a pilocytic astrocytoma. He was treated with vincristin and carboplatinum and the tumor was stable up to Dec. 1998. Dissemination of low-grade intracranial astrocytoma in children occurs in only 4%. It is not a sign of malignancy. The present case is similar to previously published cases. The prognosis of these patients might be quite favorable when treated with radiotherapy and/or chemotherapy. PMID- 10517514 TI - June 1999--22 year old female with intraventricular mass. AB - A 22 year old female presented with a single seizure. CT scan and craniotomy demonstrated an intraventricular papillary tumor with histologic and immunohistochemical features indicative of a choroid plexus carcinoma. Even though the occurrence of this neoplasm is exceptional beyond childhood, pathologists should considered a malignant choroid plexus tumor when postulating the differential diagnosis of intraventricular papillary neoplasms in adults. PMID- 10517515 TI - Mechanism of salmon sperm decondensation by nucleoplasmin. AB - Removal of protamine from DNA-protamine (salmine, protamine from salmon sperm) complexes by nucleoplasmin was examined and compared with that of poly-L-glutamic acid (PLGA) using turbidity and ethidium bromide (EB) treatment methods. When nucleoplasmin or PLGA was added to a DNA-protamine complex solution, turbidity was decreased and the amount of EB intercalated into DNA was increased. These results suggest that nucleoplasmin and PLGA can remove protamine from DNA protamine complexes. The effect of nucleoplasmin was more potent than that of PLGA. Direct interaction of nucleoplasmin with protamine was confirmed by mixing experiments using circular dichroism (CD) and fluorescence spectroscopies. Results suggest that nucleoplasmin is bound to protamine in a 1:1 ratio and that Trp126 is located near a hydrophilic region containing a polyglutamic acid tract of nucleoplasmin which was obviously influenced by its binding with protamine. It would appear that the polyglutamic acid tract in nucleoplasmin plays a critical role for binding with protamine. PMID- 10517517 TI - Conformational transition of native and modified gellan. AB - This paper concerns the characterisation of native gellan by differential scanning calorimetry (DSC) and rheology. The stability of the double helix is characterised by Tm and the enthalpy of conformational change. The role of the external salt concentration is investigated; it is shown that Tm is only slightly modified. At ambient temperature, in 10(-2) M NaCl, native gellan behaves as a loose gel (G' > G''). This behaviour disappears when temperature is larger than 60 degrees C. The comparison with deacylated gellan (commercial sample) shows that the position of conformational transition is much more influenced by the salt concentration; the helical structure is less stable and the conformational transition presents a hysteresis between heating and cooling runs when the external salt concentration increases. The rheological behaviour is that corresponding to a solution (G' < G'') at ambient temperature and in 10(-2) M NaCl. When the salt excess increases, then a stronger gel is formed. The differences between the two types of samples are clearly established as well as the relations between the conformation and the rheology of the systems. PMID- 10517516 TI - Effect of ethanol on the activity and conformation of Penaeus penicillatus acid phosphatase. AB - The effect of ethanol on the activity of Penaeus penicillatus acid phosphatase has been studied. The results show that ethanol significantly inhibits enzyme activity as a non-competitive inhibitor, with Ki 8.75%. The conformational changes of the enzyme molecule induced by ethanol were followed using fluorescence emission, ultraviolet difference and circular dichroism (CD) spectra. Increasing the ethanol concentration caused the fluorescence emission intensity of the enzyme to increase. The ultraviolet difference spectra of the enzyme denatured with ethanol had two negative peaks at 220 and 278 nm, and a positive peak at 240 nm. Increasing the ethanol concentration produced a small shoulder peak at 287 nm in addition to the increases in the negative magnitudes of the 220 and 278 nm peaks. The changes of the fluorescence and ultraviolet difference spectra reflected the changes of the microenvironments of the tryptophan and tyrosine residues of the enzyme. The CD spectrum changes of the enzyme show that the secondary structure of the enzyme also changed. The results suggest that ethanol is a non-competitive inhibitor and the conformational integrity of the enzyme is essential for its activity. PMID- 10517518 TI - Some studies of crosslinking chitosan-glutaraldehyde interaction in a homogeneous system. AB - Chitosan dissolved in acetic acid reacted with glutaraldehyde solution, ranging in concentration from 0.10 to 25.0 x 10(-2) mol dm3. The modified polymers were characterized by means of carbon, hydrogen and nitrogen elemental analysis, scanning electron microscopy, X-ray diffractometry, 13C nuclear magnetic resonance (NMR), infrared and Raman spectroscopies. The uptake of metallic cations in aqueous medium was checked through copper. The obtained data from 13C NMR, infrared and Raman spectroscopies evidenced the formation of an ethylenic double bond in the chitosan glutaraldehyde interaction. These data suggest that free pendant amine groups of chitosan polymer interact with the aldehydic group of the glutaraldehyde to form stable imine bonds, due to the resonance established with adjacent double ethylenic bonds. The crosslinking is formed by the nonuniform length of chains and by terminal unities. The crosslinking formation can involve two chitosan unities belonging, or not, to the same polymeric chain. The sequence of reactions was established for a chitosan:glutaraldehyde molar proportion of 1:20. The degree of crystallinity and particle size decreased as the amount of glutaraldehyde was increased in the polymer. Physical and chemical properties are not just affected for the chitosan glutaraldehyde reaction, but are also affected strongly by the dissolution of the natural chitosan. PMID- 10517519 TI - Thermal conformational changes of bovine fibrinogen by differential scanning calorimetry and circular dichroism. AB - The thermal denaturation of bovine fibrinogen has been investigated using differential scanning calorimetry (DSC) and circular dichroism (CD) spectroscopy. Differential scanning calorimetry measurements were carried out while changing the scan-rate. The transition at 57 degrees C was found to be irreversible and highly scan-rate dependent, suggesting that the denaturation is, at least in part, under kinetic control. The secondary structural changes at various temperatures were monitored by far-ultraviolet CD spectroscopy. These results show that the DSC transition for the thermal denaturation of bovine fibrinogen can be interpreted in terms of a kinetic process, N --> F, where k is a first order kinetic constant that changes with temperature according to the Arrhenius equation. An important transition peak was observed at 78.8 degrees C which is attributed to the C-terminal parts of the Aalpha chains of fibrinogen. PMID- 10517520 TI - Role of reducing terminals in unfractionated and low-molecular-mass heparins in causing free radical generation and loss of structure and activity of trypsin. AB - The role of both the length of saccharide chain and reducing terminals in the heparin molecule in causing oxidative effects on proteins was investigated by employing unfractionated and low-molecular-mass heparins (LMMH), with both intact and reduced reducing terminals on bovine trypsin. The effects of heparin were found to be dependent on both the concentration and time of incubation. Heparins with intact reducing terminals caused significantly higher structural and functional alterations of trypsin compared with heparins with reduced reducing terminals. LMMH was slightly more effective than unfractionated heparin (UNFH) in reducing structural integrity and inhibiting the amidolytic activity of trypsin when used at the same mass, but not molar concentrations. Neither the length of saccharide chains nor the number of intact reducing terminals on the heparin molecule appeared to influence the characteristics of the initial binding of heparin to trypsin, but both these variables crucially affected linkages which in time mediate the inhibition of catalytic activity and the formation of free radicals, ultimately responsible for peptide bond cleavage in trypsin. The results suggest that both a critical number of saccharide units, preferentially lying on shorter chains, and intact reducing terminals in the heparin molecule are involved in setting up the binding which generates radicals and leads to loss of structure and function of the proteinase. PMID- 10517521 TI - Ultrastructural aspects of phytoglycogen from cryo-transmission electron microscopy and quasi-elastic light scattering data. AB - Phytoglycogen particles extracted from the sugary maize mutant su 1 and dispersed in water were studied using transmission electron microscopy (TEM) and light scattering. Dried specimens were either negatively stained with uranyl acetate or shadowed with W/Ta. Frozen-hydrated unstained particles embedded in a thin film of vitreous ice were also observed using cryo-TEM. The particles exhibited a spheroidal shape, with a diameter ranging from 30 to 100 nm. Some of them presented a multilobular morphology and appeared to be formed by smaller subunits, 20-30 nm in diameter, resembling the described beta-particles for animal glycogen. The diameter of stained and ice-embedded particles was measured from electron micrographs. The corresponding size distribution histograms showed that the average weight diameter of ice-embedded particles was higher than that of stained ones. In the latter case, a shrinkage of the particle was believed to occur during the drying process. Light scattering experiments confirmed the diameter of ice-embedded particles and indicated that they could be considered as uniformly dense spheroidal objects. PMID- 10517522 TI - Sulphated polysaccharides of Codium dwarkense Boergs. from the west coast of India: chemical composition and blood anticoagulant activity. AB - Bioassay-guided purification of sulphated polysaccharides from a green marine alga, Codium dwarkense, yielded two products, which contained sulphated arabinan and sulphated arabinogalactan. The product containing arabinan sulphate exhibited stronger blood anticoagulant activity than the one containing sulphated arabinogalactan. PMID- 10517523 TI - A comparative study on viscosity of human, bovine and pig IgG immunoglobulins in aqueous solutions. AB - This paper presents the results of viscosity determinations on aqueous solutions of human, bovine and pig IgG immunoglobulins over a wide range of concentrations and at temperatures ranging from 5 degrees C to 55 degrees C. On the basis of the generalized Arrhenius formula, the viscosity temperature and the viscosity concentration dependence of the solutions are discussed. By applying an asymptotic form of the generalized Arrhenius formula, such rheological quantities as the intrinsic viscosity and Huggins coefficient were calculated. PMID- 10517524 TI - The effect of the chain length of polynucleotides on their binding with platinum complexes. AB - The effect of the length of polynucleotides on their binding with platinum complexes was studied. The highest reaction rate was observed in the reaction with guanosine-containing polynucleotides, whereas cytidine- and adenosine containing polynucleotides were much less efficient. The monoaqua-forms of the platinum complexes exhibited the highest reactivity in the interaction with polynucleotides in solution. The mechanism implies the formation of the monodentate complex at the first stage which is transformed into the corresponding bidentate complex of chelate type at the second stage. Increase in the length of the polynucleotide chain was shown to enhance its interaction with the platinum complexes. PMID- 10517525 TI - Micropurification of beta- and gamma-crystallins from rabbit aqueous humor. AB - Soluble crystallins are normally present in the aqueous humor, originating from the lens, and their concentration may increase in certain conditions such as cataract, possibly contributing to aqueous outflow pathway obstruction, leading to glaucoma. Whether the stability and the tendency of aqueous crystallins to aggregate are different in patients with certain forms of open-angle glaucoma has not so far been established, mainly due to the lack of a suitable purification procedure from this fluid in which crystallins are present at very low concentration together with dozens of other proteins. About 4 microg each of beta and gamma-crystallins were obtained from 20 ml of rabbit aqueous humor by C8 reversed-phase high-performance liquid chromatography (HPLC) and high-performance electrophoresis chromatography (HPEC). The identity of the proteins was confirmed by amino acid analysis following sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and electrophoretic blotting onto polyvinylidene fluoride membranes, with or without previous digestion with Staphylococcus aureus protease V8. PMID- 10517526 TI - Effect of lysine modification on the conformation and indomethacin binding properties of human serum albumin. AB - In order to study the involvement of lysine residues of human serum albumin (HSA) in the binding of indomethacin, HSA was treated with different molar excess of acetic anhydride, succinic anhydride and O-methylisourea which resulted in differently modified preparations: 30%, 62% and 87% acetylated, 20%, 34%, 64% and 78% succinylated and 21%, 43% and 86% guanidinated HSAs. All the preparations were found to be homogeneous with respect to charge as well as size as judged by polyacrylamide gel electrophoresis and gel filtration on a Seralose-6B column. Hydrodynamic and circular dichroic results showed that pronounced conformational changes (both tertiary and secondary structures) were induced in the maximally acetylated (87%) and succinylated (78%) preparations. On the other hand, guanidinated preparations showed no expansion in the hydrodynamic volume. The percent decrease in alpha-helical content was 34% for 87% acetylated, 31% for 78% succinylated and 10% for 86% guanidinated HSAs. A significant increase in the values of Stokes radii and frictional ratios (from 3.43 nm and 1.29 for native HSA to 4.07 nm and 1.52 for 87% acetylated and 4.35 nm and 1.60 for 78% succinylated HSAs, respectively) was also noticed in these highly modified preparations. Fluorescence quench titration results obtained at pH 7.4 and ionic strength 0.15 showed that only 54.1% and 64.7% binding of indomethacin at 4:1 drug/protein molar ratio was retained by 87% acetylated and 78% succinylated HSAs, respectively, as compared to 91% retention in binding in 86% guanidinated preparation. No reversal in the binding of drug to 87% acetylated and 78% succinylated HSA preparations was observed on increasing the ionic strength to 1.0. Therefore, it seems that one or two critical lysine residue(s) that can form salt linkage with the carboxyl group of indomethacin, was (were) probably modified in these preparations. A small decrease in the binding of drug to the guanidinated preparation also confirms the involvement of positive charge, probably contributed by lysine residue(s), in the binding of indomethacin to HSA. PMID- 10517527 TI - Solution properties of the galactomannans extracted from the seeds of Caesalpinia pulcherrima and Cassia javanica: comparison with locust bean gum. AB - The galactomannans from the seeds of Caesalpinia pulcherrima and Cassia javanica were extracted from the milled seeds in water at room temperature. Both products, as well as a commercial sample of locust bean gum (LBG), were purified by precipitation in isopropyl alcohol. The intrinsic viscosity determined for LBG, [eta] = 15.2 dl/g, was slightly higher than those for the other two galactomannans. The dependence of the specific viscosity at zero shear rate on the coil overlap parameter, C[eta], revealed a similar behaviour for the three galactomannans. A master curve was obtained with a critical concentration, C*, at C*[eta] = 3.3. The slope of the curve in the concentrated regime is higher than the values in the range of 3.9-6.6, obtained for the generalized behaviour of several random coil polysaccharides. Dynamic experiments showed that, at the concentrations studied, the behaviour of the galactomannans was typical of systems with predominant entanglement networks in the region between the terminal and plateau zones of frequency response. The correlation between dynamic and steady shear properties (Cox Merz rule) was satisfactory for the three galactomannans. PMID- 10517529 TI - Characterization of a crosslinked high amylose starch excipient. AB - A controlled release excipient based on sodium trimetaphosphate (STMP) crosslinked high amylose starch has been examined by 13C CP/MAS NMR. The dry excipient powder is pressed to a hard tablet whose volume increase in H2O runs parallel to the STMP concentration used. The known polymorph resonances of single helix 'V' starch (hydrated) and double helix 'B' starch (hydrated) both contribute to the spectrum corresponding to the swollen tablet. The wet tablet when loaded with a pharmaceutical agent provides a near zero-order release profile for up to 20 h. The swelling and drug release behaviour is explained in terms of self-assembly of the STMP treated starch nanomolecular particles. These particles are drawn together by "self-assembly" due to formation of amylose double helices as water penetrates the tablet. An optimum level of chemical crosslinking ensures the integrity of the swollen tablet whose sponge-like structure enclosed by a membranous surface is responsible for sustained release. PMID- 10517528 TI - Intracellular depolymerase activity in isolated inclusion bodies containing polyhydroxyalkanoates with long alkyl and functional substituents in the side chain. AB - The in vitro degradation of isolated Pseudomonas oleovorans inclusion bodies containing either poly-3-hydroxynonanoate (PHN), or poly(-3-hydroxy-5 phenylvalerate) (PHPV), or a mixture of these two polymers was investigated. When incubated at 30 degrees C and pH 9, inclusion bodies containing either polyhydroxyoctanoate (PHO), PHN or PHPV exhibited similar degradation rates of approximately 0.94 (+/- 3%) mg/h. The PHN and PHPV components for inclusion bodies containing a mixture of PHN and PHPV showed similar degradation rates; that is the ratios showed little change and remained at approximately 50 wt.% (+/ 3%) for each component. These results contrast markedly with in vivo studies for similar inclusion bodies in whole cells. The results suggest that the synthesis and degradation of these novel polyhydroxyalkanoates by P. oleovorans proceeds by the same enzymatic pathway. In addition, comparisons between the in vivo and in vitro polymer degradation suggest that the activity of the intracellular depolymerase does not control the rate limiting step of PHPV degradation in vivo. Instead, the presence of an aromatic group in the repeating units of this polymer may inhibit the utilization of the monomeric units of PHPV as a reserve carbon source by the cells. PMID- 10517530 TI - Characterization by mass spectrometry of poly(3-hydroxyalkanoates) produced by Rhodospirillum rubrum from 3-hydroxyacids. AB - The sequence distributions of two microbial copolyesters obtained by fermentation of Rhodospirillum rubrum, grown with 3-hydroxyhexanoic or 3-hydroxyheptanoic acids, were determined by analyzing the oligomers prepared by partial pyrolysis or partial methanolysis of these copolyesters using fast atom bombardment mass spectrometry (FAB-MS). Oligomers up to pentamers were identified in the case of partial pyrolysis and up to tetradecamers in the case of partial methanolysis. The comparison between the experimental and calculated peak intensities of FAB mass spectra allows the calculation of compositions and sequence distributions, which in these copolyesters follow Bernoullian statistics, indicating that they are random terpolyesters. PMID- 10517531 TI - Cross-talk of NO, superoxide and molecular oxygen, a majesty of aerobic life. AB - Because nitric oxide (NO) reacts with various molecules, such as hemeproteins, superoxide and thiols including glutathione (GSH) and cysteine residues in proteins, biological effects and metabolic fate of this gaseous radical are affected by these reactants. Although the lifetime of NO is short particularly under air atmospheric conditions (where the oxygen tension is unphysiologically high), it increases significantly under physiologically low oxygen concentrations. Because oxygen tensions in human body differ from one tissue to another and change depending on their metabolism, biological activity of NO in various tissues might be affected by local oxygen tensions. To elucidate the role of NO and related radicals in the regulation of circulation and energy metabolism, their effects on arterial resistance and energy metabolism in mitochondria, mammalian cells and enteric bacteria were studied under different oxygen tensions. Kinetic analysis revealed that NO-dependent generation of cGMP in resistance arteries and their relaxation were strongly enhanced by lowering oxygen tensions in the medium. NO reversibly suppressed the respiration and ATP synthesis of isolated mitochondria and intact cells particularly under low oxygen tensions. Kinetic analysis revealed that cross-talk between NO and superoxide generated in and around endothelial cells regulates arterial resistance particularly under physiologically low oxygen tensions. NO also inhibited the respiration and ATP synthesis of E. coli particularly under low oxygen tensions. Because concentrations of NO and H+ in gastric juice are high, most ingested bacteria are effectively killed in the stomach. However, the inhibitory effects of NO on the respiration and ATP synthesis of H. pylori are extremely small. Kinetic analysis revealed that H. pylori generates the superoxide radical thereby inhibiting the bactericidal action of NO in gastric juice. Based on such observations, critical roles of the cross-talk of NO, superoxide and molecular oxygen in the regulation of energy metabolism and survival of aerobic and microaerophilic organisms are discussed. PMID- 10517532 TI - Antioxidant defence mechanisms: from the beginning to the end (of the beginning). AB - When life first evolved on Earth, there was little oxygen in the atmosphere. Evolution of antioxidant defences must have been closely associated with the evolution of photosynthesis and of O2-dependent electron transport mechanisms. Studies with mice lacking antioxidant defences confirm the important roles of MnSOD and transferrin in maintaining health, but show that glutathione peroxidase (GPX) and CuZnSOD are not essential for everyday life (at least in mice). Superoxide can be cytotoxic by several mechanisms: one is the formation of hydroxyl radicals. There is good evidence that OH* formation occurs in vivo. Other important antioxidants may include thioredoxin, and selenoproteins other than GPX. Nitric oxide may be an important antioxidant in the vascular system. Diet-derived antioxidants are important in maintaining human health, but recent studies employing "biomarkers" of oxidative DNA damage are questioning the "antioxidant" roles of beta-carotene and ascorbate. An important area of future research will be elucidation of the reasons why levels of steady-state oxidative damage to DNA and lipids vary so much between individuals, and their predictive value for the later development of human disease. PMID- 10517533 TI - Glutathione and glutathione-dependent enzymes represent a co-ordinately regulated defence against oxidative stress. AB - Increases in the intracellular levels of reactive oxygen species (ROS), frequently referred to as oxidative stress, represents a potentially toxic insult which if not counteracted will lead to membrane dysfunction, DNA damage and inactivation of proteins. Chronic oxidative stress has numerous pathological consequences including cancer, arthritis and neurodegenerative disease. Glutathione-associated metabolism is a major mechanism for cellular protection against agents which generate oxidative stress. It is becoming increasingly apparent that the glutathione tripeptide is central to a complex multifaceted detoxification system, where there is substantial inter-dependence between separate component members. Glutathione participates in detoxification at several different levels, and may scavenge free radicals, reduce peroxides or be conjugated with electrophilic compounds. Thus, glutathione provides the cell with multiple defences not only against ROS but also against their toxic products. This article discusses how glutathione biosynthesis, glutathione peroxidases, glutathione S-transferases and glutathione S-conjugate efflux pumps function in an integrated fashion to allow cellular adaption to oxidative stress. Co ordination of this response is achieved, at least in part, through the antioxidant responsive element (ARE) which is found in the promoters of many of the genes that are inducible by oxidative and chemical stress. Transcriptional activation through this enhancer appears to be mediated by basic leucine zipper transcription factors such as Nrf and small Maf proteins. The nature of the intracellular sensor(s) for ROS and thiol-active chemicals which induce genes through the ARE is described. Gene activation through the ARE appears to account for the enhanced antioxidant and detoxification capacity of normal cells effected by many cancer chemopreventive agents. In certain instances it may also account for acquired resistance of tumours to cancer chemotherapeutic drugs. It is therefore clear that determining the mechanisms involved in regulation of ARE driven gene expression has enormous medical implications. PMID- 10517534 TI - Down regulation of superoxide dismutases and glutathione peroxidase by reactive oxygen and nitrogen species. AB - The levels of antioxidative enzymes are regulated by gene expressions as well as by post-translational modifications. Although their functions are to scavenge reactive oxygen (ROS) and nitrogen species (RNS), they may also be targets of various oxidants. When ROS and RNS modify the functions of antioxidative enzymes, especially glutathione peroxidase, they may induce apoptotic cell death in susceptible cells. It is conceivable, therefore, that at least a part of the apoptotic pathways mediated by ROS and RNS may be associated with modification of the redox regulation of cellular functions due to elevations of such substances. In this article we review recent findings about the effects of various oxidative conditions associated with alteration of these antioxidative enzymes and the concomitant cellular damage induced. PMID- 10517535 TI - Induction of antioxidant stress proteins in vascular endothelial and smooth muscle cells: protective action of vitamin C against atherogenic lipoproteins. AB - Elevated levels of lipid peroxidation and increased formation of reactive oxygen species within the vascular wall in atherosclerosis can overwhelm cellular antioxidant defence mechanisms. Accumulating evidence implicates oxidatively modified low density lipoproteins (LDL) in vascular dysfunction in atherosclerosis and oxidized LDL have been localized with in atherosclerotic lesions. We here report that human oxidatively modified LDL induce expression of 'antioxidant-like' stress proteins in vascular cells, involving increases in the activity of L-cystine transport, glutathione synthesis, heme oxygenase-1 and the murine stress protein MSP23. Moreover, treatment of human arterial smooth muscle cells with the dietary antioxidant vitamin C markedly attenuates adaptive increases in endogenous antioxidant gene expression and affords protection against smooth muscle cell apoptosis induced by moderately oxidized LDL. As vascular cell death is a key feature of atherosclerotic lesions and may contribute to the plaque 'necrotic' core, cap rupture and thrombosis, our findings suggest that the cytoprotective actions of vitamin C could limit plaque instability in advanced atherosclerosis. PMID- 10517536 TI - Regulatory mechanisms of cellular response to oxidative stress. AB - An antioxidant responsive element (ARE) or electrophile responsive element (EpRE) mediates the transcriptional activation of genes encoding phase II drug metabolizing enzymes. The ARE consensus sequence shows high similarity to an erythroid gene regulatory element, and based on the observation, we have recently found that transcription factor Nrf2 is essential for the coordinate induction of phase II detoxifying enzymes. The expression of anti-oxidative stress enzyme genes is also regulated by Nrf2. Detailed analysis of the regulatory mechanisms of Nrf2 activity has ultimately led us to the identification of a new protein, which we have named Keap1, that suppresses Nrf2 activity by specific binding to its evolutionarily-conserved N-terminal Neh2 regulatory domain. PMID- 10517537 TI - Regulation of gamma-glutamylcysteine synthetase expression in response to oxidative stress. AB - Glutathione (GSH) is synthesized by the activity of two ATP-requiring GSH synthesizing enzymes. Gamma-glutamylcysteine synthetase (gamma-GCS) is the rate limiting enzyme for the GSH synthesis. Gamma-GCS is a heterodimer of heavy, catalytic subunit and light, regulatory subunit and responsive to many stresses, such as heat shock, oxidative stress or cytokines. To know the regulation of the expression of gamma-GCS gene, in the present study, we show evidences that gamma GCS heavy subunit is upregulated by oxidative stress by ionizing radiation and TNF-alpha mediated by nuclear factor-kappaB (NF-kappaB), and impairment of the expression of gamma-GCS by TNF-alpha in diabetic condition. Furthermore we describe the importance of GSH in the regulation of NF-kappaB subunits. PMID- 10517538 TI - Redox regulation and oxidant activation of heme oxygenase-1. AB - The ultraviolet A (UVA, 320-400 nm) component of sunlight has the potential to generate an oxidative stress in cells and tissue so that antioxidants (both endogenous and exogenous) strongly influence the biological effects of UVA. The expression of several genes (including heme oxygenase-1, HO-1; collagenase; the CL100 phosphatase and the nuclear oncogenes, c-fos and c-jun) is induced following physiological doses of UVA to cells and this effect can be strongly enhanced by removing intracellular glutathione or enhancing singlet oxygen lifetime. We have observed that heme is released from microsomal heme-containing proteins by UVA and other oxidants and that activation of HO-1 expression by UVA correlates with levels of heme release. UVA radiation also leads to an increase in labile iron pools (either directly or via HO-1) and eventual increases in ferritin levels. The role of heme oxygenase in protection of skin fibroblasts is probably an emergency inducible defense pathway to remove heme liberated by oxidants. The slower increase in ferritin levels is an adaptive response which serves to keep labile iron pools low and thereby reduce Fenton chemistry and oxidant-induced chain reactions involving lipid peroxidation. In keratinocytes, the primary target of UVA radiation, heme oxygenase levels are constitutively high (because of HO-2 expression). Since there is a corresponding increase in basal levels of ferritin the epidermis appears to be well protected constitutively against the oxidative stress generated by UVA. PMID- 10517539 TI - The role of protein phosphatases in the regulation of mitogen and stress activated protein kinases. AB - It is now established that a family of dual-specificity protein phosphatases are able to interact with mitogen and stress-activated protein kinases in a highly specific manner to differentially regulate these enzymes in mammalian cells. A role for these proteins in negative feedback regulation of MAP kinase activity is also supported by genetic and biochemical studies in yeasts and Drosophila. More recently it has become clear that other classes of protein phosphatase also play key roles in the regulated dephosphorylation of MAP kinases, including tyrosine specific protein phosphatases and serine/threonine protein phosphatases. It is likely that a complex balance between upstream activators and these different classes of MAP kinase specific phosphatase are responsible for determining, at least in part, the magnitude and duration of MAP kinase activation and hence the physiological outcome of signalling. PMID- 10517540 TI - Oxidative stress-inducible proteins in macrophages. AB - Macrophages produce reactive oxygen species such as O2-, H2O2 and *OH that contribute to the pathogenesis of diseases such as inflammation and atherosclerosis. The cells have multiple defense systems against those reactive oxygen species, and we describe here such an oxidative stress-inducible defense system. Upon exposure to reactive oxygen species and electrophilic agents, murine peritoneal macrophages induce stress proteins to protect themselves. Using differential screening, we cloned two novel proteins designated MSP23 and A170 that are induced in the cells by low levels of reactive oxygen species, electrophilic agents and other oxidative stress agents. MSP23 is murine peroxiredoxin I having a thioredoxin peroxidase activity and A170 is known as an ubiquitin- and PKC xi-binding protein. In addition to these two proteins, heme oxygenase-1 (HO-1) and cystine transport activity are also induced in the cells under oxidative stress conditions. Using nrf2-deficient macrophages, we found that transcription factor Nrf2, which is known to interact with antioxidant responsive elements (AREs) in the regulatory sequences of the genes, plays an important role in the oxidative stress-inducible response in the cells. PMID- 10517541 TI - Mouse glutaredoxin - cDNA cloning, high level expression in E. coli and its possible implication in redox regulation of the DNA binding activity in transcription factor PEBP2. AB - We have isolated a cDNA encoding glutaredoxin (GRX) from a mouse splenic cDNA library. This cDNA encoded a protein of 107 amino acids with a calculated molecular weight of 11.9 kDa. The deduced amino acid sequence of glutaredoxin in mouse was highly homologous with that in other mammals (81-89%), containing a putative active sequence of -Cys-Pro-Try-Cys-. Recombinant mouse glutaredoxin expressed in E. coli showed glutathione-disulfide oxidoreductase activity with beta-hydroxyethyl disulfide as its substrate, whereas mutant glutaredoxin (Cys 22, Cys 25 to Ser) showed no activity. In electrophoretic mobility shift assay, we proved that wild type GRX, not mutant one, recovered the DNA-binding activity of a transcription factor, PEBP2, oxidized by diamide. This showed that GRX may be involved in the redox regulation of the DNA-binding activity of PEBP2 as is the case with thioredoxin. PMID- 10517542 TI - Measurement of low breath-alcohol concentrations: laboratory studies and field experience. AB - Recent federal rules and traffic law changes impose breath-alcohol thresholds of 0.02 and 0.04 g/210 L upon some classes of motor vehicle operators, such as juveniles and commercial vehicle operators. In federally regulated alcohol testing in the workplace, removal of covered workers from safety-sensitive duties, and other adverse actions, also occur at breath-alcohol concentrations (BrACs) of 0.02 and 0.04 g/210 L. We therefore studied performance of vapor alcohol and breath-alcohol measurement at low alcohol concentrations in the laboratory and in the field, with current-generation evidential analyzers. We report here chiefly our field experience with evidential breath-alcohol testing of drinking drivers on paired breath samples using 62 Intoxilyzer 5000-D analyzers, for BrACs of 0-0.059 g/210 L. The data from 62 law enforcement breath alcohol testing sites were collected and pooled, with BrACs recorded to three decimal places, and otherwise carried out under the standard Oklahoma evidential breath-alcohol testing protocol. For 2105 pooled simulator control tests at 0.06 0.13 g/210 L the mean +/- SD of the differences between target and result were 0.001 +/- 0.0035 g/210 L and 0.003 +/- 0.0023 g/210 L for signed and absolute differences, respectively (spans -0.016-0.010, 0.000-0.016). For 2078 paired duplicate breath-alcohol measurements with the Intoxilyzer 5000-D, the mean +/- SD difference (BrAC1-BrAC2) were 0.002 +/- 0.0026 (span 0-0.020 g/210 L). Variability of breath-alcohol measurements was related inversely to the alcohol concentration. Ninety-nine percent prediction limits for paired BrAC measurements correspond to a 0.020 g/210 L maximum absolute difference, meeting the NSC/CAOD recommendation that paired breath-alcohol analysis results within 0.02 g/210 L shall be deemed to be in acceptable agreement. We conclude that the field system for breath-alcohol analysis studied by us can and does perform reliably and accurately at low BrACs. PMID- 10517543 TI - Drug testing with alternative matrices II. Mechanisms of cocaine and codeine deposition in hair. AB - A 10-week inpatient study was performed to evaluate cocaine, codeine, and metabolite disposition in biological matrices collected from volunteers. An initial report described drug disposition in plasma, sebum, and stratum corneum collected from five African-American males. This report focuses on drug disposition in hair and sweat collected from the same five subjects. Following a three-week washout period, three doses of cocaine HCl (75 mg/70 kg, subcutaneous) and three doses of codeine SO4 (60 mg/70 kg, oral) were administered on alternating days in week 4 (low-dose week). The same dosing sequence was repeated in week 8 with doubled doses (high-dose week). Hair was collected by shaving the entire scalp once each week. Hair from the anterior vertex was divided into two portions. One portion was washed with isopropanol and phosphate buffer; the other portion was not washed. Hair was enzymatically digested, samples were centrifuged, and the supernatant was collected. Sweat was collected periodically by placing PharmChek sweat patches on the torso. Drugs were extracted from sweat patches with methanol/0.2 M sodium acetate buffer (75:25, v/v). Supernatants from hair digests, hair washes, and sweat patch extracts were processed by solid-phase extraction followed by gas chromatography-mass spectrometry analysis for cocaine, codeine, 6-acetylmorphine, and metabolites. Cocaine and codeine were the primary analytes identified in sweat patches and hair. Drugs were detected in sweat within 8 h after dosing, and drug secretion primarily occurred within 24 h after dosing. No clear relationship was observed between dose and drug concentrations in sweat. Drug incorporation into hair appeared to be dose-dependent. Drugs were detected in hair within 1-3 days after the last drug administration; peak drug concentrations generally occurred in the following 1-2 weeks; thereafter, drug concentrations decreased. Solvent washes removed 50-55% of cocaine and codeine from hair collected 1-3 days after the last drug dose. These data may reflect removal of drug that was deposited by sweat shortly after dosing. Drug removed by washing hair collected 1-3 weeks after the last dose was minimal for cocaine but variable for codeine. Drug in these specimens was likely transferred from blood to germinative hair cells followed by emergence of drug in growing hair. These findings suggest that drug deposition in hair occurs by multiple mechanisms. PMID- 10517544 TI - A retrospective study of buprenorphine and norbuprenorphine in human hair after multiple doses. AB - The analysis of hair has been proposed as a tool for monitoring drug-treatment compliance. This study was performed to determine if buprenorphine (BPR) and norbuprenorphine (NBPR) could be detected in human hair after controlled administration of drug and to determine if segmental analysis of hair was an accurate record of the dosing history. Subjects with dark hair (six males, six females) received 8 mg sublingual BPR for a maximum of 180 days. Single hair collections were made once after BPR treatment and stored at -20 degrees C until analysis. Hair was aligned scalp-end to tip and then segmented in 3-cm sections. For this study, it was assumed that the mean hair growth rate was 1.0 cm/month. Deuterated internal standard was added to hair segments (2-20 mg of hair) and digested overnight at room temperature with 1 N NaOH. Specimens were extracted with a liquid-liquid procedure and analyzed by liquid chromatography-tandem mass spectrometry. The limits of quantitation for BPR and NBPR were 3 pg/mg and 5 pg/mg, respectively, for 20 mg of hair. BPR and NBPR concentrations were highest for all subjects in hair segments estimated to correspond to the subject's period of drug treatment. With one exception, NBPR was present in higher concentrations in hair than was the parent compound. BPR concentrations in hair segments ranged from 3.1 pg/mg to 123.8 pg/mg. NBPR concentrations ranged from 4.8 pg/mg to 1517.8 pg/mg. In one subject, BPR and NBPR were not detected in any hair segment. In some subjects, BPR and NBPR were detected in hair segments that did not correspond to the period of drug treatment, suggesting that drug movement may have occurred by diffusion in sweat and other mechanisms. The data from this study also indicate that there is a high degree of intersubject variability in measured concentration of BPR and NBPR in hair segments, even when subjects receive the same dose for an equivalent number of treatment days. Future prospective studies involving controlled drug administration will be necessary to evaluate whether hair can serve as an accurate historical record of variations in the pattern of drug use. PMID- 10517545 TI - Detection of nandrolone, testosterone, and their esters in rat and human hair samples. AB - Nandrolone and testosterone are anabolic androgenic steroids occasionally abused by athletes. A sensitive, specific, and reproducible gas chromatography-mass spectrometry method for the quantitative determination of nandrolone, testosterone, and their esters in hair has been developed. The limits of quantitation of this method, based on 20 mg of hair, were 50 pg/mg for nandrolone and testosterone, 100 pg/mg for testosterone acetate, and 200 pg/mg for nandrolone-decanoate. Nandrolone-d3 and testosterone-d3 were used as internal standards. This method has been applied to the analysis of these compounds incorporated into rat and human hair. Male Long-Evans rats were given nandrolone decanoate 60 mg/kg intraperitoneally (i.p.) once daily for 10 days over a time period of 14 days. Two of the three rats contained nandrolone in the pigmented hair collected at day 21 at a concentration of 63 and 76 pg/mg, respectively. No drug was found in the corresponding nonpigmented hair. The rat hair samples that tested positive for nandrolone contained also nandrolone decanoate in concentrations of 0.9 and 1.2 ng/mg, respectively. In a separate experiment rats were given testosterone acetate 10 mg/kg i.p. once daily for five days. No testosterone or testosterone acetate was detected in the rat hair samples. Hair specimens were also obtained from four self-reported steroid users. The hair of two subjects were determined to be positive for testosterone in concentrations of 54 and 81 pg/mg. These data demonstrate that it is possible to detect the steroids nandrolone, testosterone, and nandrolone decanoate in hair after systemic administration. PMID- 10517546 TI - Physiological concentrations of DHEA in human hair. AB - In 1974, steroids were added to the list of doping agents banned by the International Olympic Committee because of their effects on the performance of the athletes. Dehydroepiandrosterone (DHEA) is a steroid hormone naturally produced by the adrenal glands and by the ovaries. DHEA can be converted into other hormones, including estrogen and testosterone. In the United States, DHEA is classified as a nutritional supplement. This is not the case in France, where the drug is listed as a doping agent. As athletes can abuse DHEA to benefit from its conversion to testosterone, there is a need to establish the physiological range of DHEA concentrations in human hair. DHEA was investigated in hair obtained from 27 control subjects, including 15 males and 12 females aged 17-42 years. After decontamination with dichloromethane, 100 mg of hair was incubated in 1 M NaOH in presence of 1 ng of testosterone-d3. After neutralization, the extract was purified using solid-phase extraction with Isolute C18 columns and subsequent liquid-liquid extraction with pentane. After silylation, DHEA was analyzed by gas chromatography-mass spectrometry. Results were linear in the range 1-20 pg/mg. Relative extraction recovery was 91.6% with a limit of detection of 0.5 pg/mg. Concentrations were in the range 1.2-6.7 pg/mg (mean value of 4.3 pg/mg) and 0.5 to 10.6 pg/mg (mean value of 5.3 pg/mg) for the males and females, respectively. Extensive chromatographic procedures (two purification steps by solid-phase and liquid-liquid extraction, combined with injection of 4 microL through the column in pulsed mode) were analytical prerequisites for successful identification of DHEA in hair because of the low target concentrations. This new technology may find useful applications in anabolic abuse control. PMID- 10517547 TI - Highly sensitive micro-plate enzyme immunoassay screening and NCI-GC-MS confirmation of flunitrazepam and its major metabolite 7-aminoflunitrazepam in hair. AB - Flunitrazepam (Rohypnol) is a benzodiazepine used in the treatment of insomnia as a sedative hypnotic and as preanesthetic medication in European countries and Mexico. Although it has no medicinal purpose in the United States, the occurrence of its abuse is increasing. Sexual abuse of both men and women while under the influence of so-called "date-rape" drugs has been the focus of many investigations. Reported date-rape drugs include flunitrazepam (FN), clonazepam, diazepam, oxazepam, gamma-hydroxybutyrate, and many others. FN has been banned in the United States because of its alleged use in such situations. Unfortunately, the detection of FN or its metabolites 7-aminoflunitrazepam (7-AFN) and desmethylflunitrazepam in a single specimen such as urine or blood is difficult in criminal situations because of the likelihood of single-dose ingestion and the length of time since the alleged incident. Hair provides a solution to the second of these problems in that drugs tend to incorporate into hair and remain there for longer periods of time than either urine or blood. There are various techniques for the detection of FN in plasma, blood, and urine, but little work has been done with hair. Hair collection is a virtually noninvasive procedure that can supply information on drug use for several months preceding collection. The objective of this paper was to determine if a commercially available micro plate enzyme immunoassay system was sufficiently sensitive for the routine screening of 7-AFN in hair by the development of extraction procedures and optimization of the immunoassay kit. Further, this study used the same solid phase extraction to isolate FN and its major metabolite, 7-AFN, and gas chromatography-mass spectrometry with negative ion chemical ionization for confirmation. Two seven-point standard curves were established ranging from 0.5 pg/mg to 100 pg/mg for 7-AFN and 2.5 pg/mg to 200 pg/mg for FN with respective deuterated internal standards. A replicate analysis of controls was performed to establish inter- and intraday variabilities. Two suicide cases along with one alleged date-rape case and one case of an emergency room patient whose blood screened positive for benzodiazepines were analyzed. All the hair specimens screened positive for benzodiazepines using micro-plate enzyme immunoassay. Two cases, including the date-rape case, were negative for FN and 7-AFN, and two postmortem hair samples were confirmed positive for FN and its metabolite. PMID- 10517548 TI - Immunoassay and GC-MS procedures for the analysis of drugs of abuse in meconium. AB - The analysis of meconium specimens for metabolites of substances of abuse is a relatively accurate method for the detection of fetal exposure to drugs. Most of the methods reported in the literature before the early 1990s relied on radioimmunoassays. The purpose of this study was to develop and validate methods for meconium sample preparation for the screening and gas chromatography-mass spectrometry (GC-MS) confirmation of meconium extracts for cannabinoids, cocaine, opiates, amphetamines, and phencyclidine. EMIT and TDx immunoassays were evaluated as screening methods. The sample preparation method developed for screening included extraction and purification prior to analysis. Cutoff levels were administratively set at 20 ng/g for 11-nor-delta9-THC-9-COOH (THCCOOH) and phencyclidine and at 200 ng/g for benzoylecgonine, morphine, and amphetamines, although lower levels could be detected in meconium using the EMIT-ETS system. Ninety-five meconium specimens were subjected to the screening procedure with GC MS confirmation of presumptive positives. In addition, 30 (40 for cocaine) meconium specimens were subjected to GC-MS analysis for all analytes regardless of the screening results to determine the false-negative rate, if any, of the immunoassay. Although there were no false negatives detected, the GC-MS confirmation rate for the immunoassay-positive specimens was generally low, ranging from 0% for amphetamines to 75% for opiates. The lowest rate of confirmed positives was found with the cannabinoids, suggesting that tetrahydrocannabinol (THC) metabolites other than free 11-nor-9-carboxy-delta9-THC may be major contributors to the immunoassay response in meconium. PMID- 10517549 TI - Identification and analysis of the major metabolites of cocaine in meconium. AB - A gas chromatographic-mass spectrometric (GC-MS) method was used for the simultaneous analysis of cocaine and its metabolites (norcocaine, benzoylecgonine, norbenzoylecgonine, ecgonine methylester, cocaethylene, and the o-, m, and p-hydroxy derivatives of cocaine and benzoylecgonine) in meconium specimens collected from newborns prenatally exposed to the drug in an effort to determine which of these metabolites are more relevant in the confirmation of immunoassay-positive specimens. Significant metabolites included benzoylecgonine and its m- and p-hydroxy derivatives. Reanalysis of immunoassay-positive meconium specimens that previously failed to confirm by GC-MS for benzoylecgonine revealed that the GC-MS confirmation rate could be substantially enhanced by inclusion of m- and p-hydroxybenzoylecgonine in the analysis along with benzoylecgonine. Several specimens were found to be positive for hydroxylated derivative(s) in the absence of benzoylecgonine itself. PMID- 10517550 TI - Correlation of saliva codeine concentrations with plasma concentrations after oral codeine administration. AB - A clinical study was designed to determine if there was a predictable relationship between saliva and plasma codeine concentrations. Drug-free volunteers (n = 17) were administered a 30-mg dose of liquid codeine phosphate. Plasma and saliva specimens were collected at various times for 24 h after administration. Plasma and saliva were analyzed for codeine and morphine by positive-ion chemical ionization gas chromatography-mass spectrometry. The plasma codeine concentrations peaked between 30 min and 2 h after administration and ranged from 19 to 74 ng/mL with a mean of 46 ng/mL. Despite decontamination procedures, elevated saliva codeine concentrations were detected at the early collection times because of contamination of the oral cavity from the liquid codeine. Codeine concentrations in the 15 min specimens ranged from 690 ng/mL to over 15,000 ng/mL. After the initial 2-h period, the mean codeine saliva concentrations declined at a rate similar to that observed in the plasma, but remained 3 to 4 times greater than the plasma concentrations. During the elimination phase, half-life estimates for codeine in plasma and saliva were found to be equivalent, 2.6 and 2.9 h, respectively. However, the area under the curve (AUC) estimate for codeine in saliva was 13 times greater than the plasma AUC. Contamination of the saliva resulted in elevated saliva/plasma (S/P) concentration ratios for the first 1 to 2 h after drug administration. Consequently, S/P ratios in specimens collected in the first 15 to 30 min ranged from 75 to 2580. However, after the absorption phase, a significant correlation between saliva and plasma concentrations was observed (r = 0.809, p < 0.05) and mean S/P ratios remained constant (mean = 3.7). Although small changes in saliva pH were predicted to produce profound changes in the S/P ratios for codeine, this was not observed in the current study. Therefore, saliva codeine concentrations could be used to estimate plasma concentrations through the use of the S/P ratio once the oral contamination has been eliminated. However, these estimates should be made cautiously. One must ensure that oral contamination is not a factor. Also, as with blood-drug concentrations, considerable intersubject variability was observed. PMID- 10517551 TI - A rapid reusable fiber optic biosensor for detecting cocaine metabolites in urine. AB - Analyte 2000, a four-channel fiber optic biosensor (FOB), was used for analysis of cocaine and its metabolites (COC) in human urine using a competitive fluorescence immunoassay. Binding of antibenzoylecgonine monoclonal antibody (mAb) to the casein-benzoylecgonine Ag-coated, tapered optical fibers was inhibited by COC. Bound mAb, which inversely correlated with COC concentration, was quantitated by fluorescence produced by evanescent excitation of bound cyanine dye-tagged antimouse antibody (CY5-Ab). The effective concentration range of benzoylecgonine (BE) for inhibiting the fluorescent signals was 0.75-50 ng/mL, with IC50 of 9.0 ng/mL. This FOB had similar affinities for BE, cocaine, and cocaethylene, but very low affinities for ecgonine and ecgonine methyl ester. A sensitivity of 100% and a specificity of 96% were achieved when 54 human urine specimens were analyzed by FOB (cutoff, 300 ng/mL COC) and GC-MS (cutoff, 150 ng/mL BE). All results were in agreement except for one positive FOB result with a GC-MS BE concentration of 148 ng/mL. In addition, regeneration and reuse of the fiber for multiple analyses were performed successfully with no carryover from specimens containing high COC concentrations to specimens containing low COC concentrations. PMID- 10517552 TI - Determination of morphine, morphine-3-glucuronide, and morphine-6-glucuronide in plasma after intravenous and intrathecal morphine administration using HPLC with electrospray ionization and tandem mass spectrometry. AB - High-performance liquid chromatography (HPLC) coupled to atmospheric pressure ionization (API) mass spectrometry (MS) has become a useful technique in the direct analysis of low concentrations of conjugated opiate metabolites. Previous methods using HPLC with traditional detection methods do not have the sensitivity to detect low concentrations of most conjugated drug metabolites. Methods using gas chromatography-mass spectrometry (GC-MS) require hydrolysis and derivatization of the sample followed by an indirect quantitation of conjugated metabolites. Recently, several reports have described direct analysis of opiates and their glucuronide conjugates by HPLC and API-MS. These methods report lower limits of detection than GC-MS methods and quantitation in the low nanogram-per milliliter range for the glucuronide metabolites of morphine. This report describes an HPLC-electrospray-MS-MS method capable of detecting subnanogram concentrations of morphine (MOR) and its 3- and 6-glucuronide metabolites (M3G and M6G, respectively). The assay has a dynamic range of 250-10,000 pg/mL for M3G and M6G and 500-10,000 pg/mL for MOR. Inter- and intra-assay precision and accuracy varied by less than 8% for all analytes at 750-, 2500-, and 7500-pg/mL concentrations. This assay was used for the determination of MOR, M3G, and M6G in human plasma after intravenous (i.v.) and intrathecal (i.t.) administration of MOR and its effects on the ventilatory response to hypoxia. Peak plasma concentrations of MOR and M6G were measured 1 h after i.v. administration of MOR. Peak concentrations of M3G were measured 2 h after i.v. administration of MOR. After i.t. administration of MOR, peak concentrations of M3G were measured 8 h postdose. MOR was not detected in plasma of patients administered MOR i.t.. Subnanogram concentrations of M6G were measured in the plasma of five of nine patients administered MOR i.t.. PMID- 10517553 TI - Detection of lysergic acid diethylamide (LSD) in urine by gas chromatography-ion trap tandem mass spectrometry. AB - A confirmatory method for the detection and quantitation of lysergic acid diethylamide (LSD) is presented. The method employs gas chromatography-tandem mass spectrometry (GC-MS-MS) using an internal ionization ion trap detector for sensitive MS-MS-in-time measurements of LSD extracted from urine. Following a single-step solid-phase extraction of 5 mL of urine, underivatized LSD can be measured with limits of quantitation and detection of 80 and 20 pg/mL, respectively. Temperature-programmed on-column injections of urine extracts were linear over the concentration range 20-2000 pg/mL (r2 = 0.999). Intraday and interday coefficients of variation were < 6% and < 13%, respectively. This procedure has been applied to quality-control specimens and LSD-positive samples in this laboratory. Comparisons with alternate GC-MS methods and extraction procedures are discussed. PMID- 10517554 TI - Quantitation of alprazolam and alpha-hydroxyalprazolam in human plasma using liquid chromatography electrospray ionization MS-MS. AB - A sensitive and specific electrospray ionization high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS-MS) method has been developed for the quantitative determination of alprazolam (AL) and alpha-hydroxyalprazolam (OH-AL) in plasma. After the addition of deuterium labeled internal standards of AL and OH-AL, plasma samples were buffered to alkaline pH and extracted with toluene/methylene chloride (7:3). Dried extract residues were reconstituted in HPLC mobile phase and injected onto a reversed-phase C18 HPLC column. The analytes were eluted isocratically at a flow rate of 250 microL/min using a solvent composed of methanol and water (60:40) containing 0.1% formic acid. The analyses were performed using selected reaction monitoring. The assay was sensitive to 0.05 ng/mL for both the parent drug and metabolite and linear to 50 ng/mL. The intra-assay percent coefficients of variation (%CV) for AL at 2, 5, and 20 ng/mL were all < or = 5.6. At these concentrations, and all OH-AL intra assay %CVs were < or = 8.4. The interassay variabilities for AL were 11.8%CV, 8.7%CV, and 8.7%CV at 2.0, 5.0, and 20.0 ng/mL, respectively. The OH-AL interassay variabilities were 9.6%CV, 9.2%CV, and 7.8%CV at the same concentrations, respectively. The assay accuracy was less than or equal to +/- 6.6% for both analytes at the three concentrations. The method was used to quantitate AL and OH-AL in plasma samples collected from 10 subjects who were administered a 1-mg oral dose of AL. The mean AL concentration peaked at 11.5 ng/mL 1 h after the dose and AL was detectable for 48 h. The mean OH-AL concentration peaked at 0.18 ng/mL 4 h after the dose and was undetectable by 36 h. Hydrolysis of the plasma samples had little effect on the detected AL concentrations but increased OH-AL concentrations substantially. Plasma/blood ratios for AL and OH-AL exceeded 1 in the study samples. PMID- 10517555 TI - GC-MS determination of flunitrazepam and its major metabolite in whole blood and plasma. AB - A gas chromatography-mass spectrometry method was developed for the analysis of flunitrazepam (FN) and its major metabolite, 7-amino-flunitrazepam (7-amino-FN), in both plasma and whole blood. The method was based on acid hydrolysis of the samples after dilution with HPLC water followed by extraction and derivatization (heptafluorobutyrate) of the resulting benzophenones. Analysis of plasma and whole blood samples from subjects administered 2-mg doses of FN showed that FN was only detected in whole blood (LOD 5 ng/mL) and not in plasma. However, 7 amino-FN was detected in both plasma and whole blood, although the levels were much higher in plasma. 7-Amino-FN was detected for the entire period of specimen collection (12 h), but FN was only detected in whole blood for 4 h after ingestion with peak levels after 1 h. PMID- 10517556 TI - Benzodiazepines in Miami-Dade County, Florida driving under the influence (DUI) cases (1995-1998) with emphasis on Rohypnol: GC-MS confirmation, patterns of use, psychomotor impairment, and results of Florida legislation. AB - Benzodiazepines are central nervous system depressant drugs often detected in biological samples from driving under the influence (DUI) offenders. They are associated with marked psychomotor impairment and represent up to 20% of all Miami-Dade County, Florida DUI urine samples analyzed in our laboratory annually. Flunitrazepam emerged in the mid-1990s as an illegal drug in the U.S. that was predominantly abused recreationally and associated with sexual assaults. Immunoassays for benzodiazepines do not discriminate between different benzodiazepines, and certain metabolites, such as 7-aminoflunitrazepam, react poorly with immunoassay reagents. A simple and sensitive method for the detection and quantitation of major benzodiazepines and metabolites by gas chromatography with mass selective detection is presented. This method was used to confirm benzodiazepines in general and flunitrazepam in particular. Data collected over a three-and-a-half-year period are summarized. Whereas flunitrazepam was present in up to 10% of DUI cases in 1995 and 1996 and had fast become the most frequently encountered benzodiazepine in Miami-Dade County DUI-related urine samples, a dramatic drop in case numbers followed the legal reclassification of the drug as a Schedule I substance in Florida in February 1997. Flunitrazepam was often used alone or in combination with cannabis and cocaine. A recent rise in clonazepam cases coincides with the decrease in flunitrazepam confirmation and may indicate a new trend in the abuse of benzodiazepines in South Florida. PMID- 10517557 TI - Detection of methadone, LAAM, and their metabolites by methadone immunoassays. AB - l-Alpha-acetylmethadol (LAAM) was recently approved as a substitute for methadone. LAAM, methadone, and their common metabolite, methadol, are extensively N-demethylated. The structural similarities of LAAM and its metabolites to methadone suggest that they may cross-react in methadone immunoassays. To test this hypothesis, drug-free urine was fortified with LAAM, norLAAM, dinorLAAM, methadol, normethadol, dinormethadol, methadone, 2-ethylidene 1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP), or 2-ethyl-5-methyl-3,3 diphenylpyrroline (EMDP) at 12 concentrations (0.03 to 100 microg/mL). Samples were analyzed using two enzyme immunoassays (Behring Diagnostics, EIA-b; Diagnostic Reagents, EIA-d); a fluorescent polarization immunoassay (Abbott, FPIA); two enzyme-linked immunosorbant immunoassays (Diagnostix, ELISA-d; STC Technologies, ELISA-s); a kinetic microparticles in solution immunoassay (Roche Diagnostic Systems, KIMS); and a radioimmunoassay (Diagnostic Products, RIA). LAAM had high cross-reactivity with ELISA-d (318.3%), RIA (249.5%), EIA-d (100.8%), KIMS (91.1%), and ELISA-s (75.3%). Methadol also displayed relatively high cross-reactivity as follows: EIA-d (97.8%), KIMS (85.4%), ELISA-d (70.3%), and FPIA (37.7%). Successive N-demethylations of LAAM and methadol were associated with loss of cross-reactivity. The methadone metabolites EDDP and EMDP showed little cross-reactivity. These findings suggest that LAAM use could result in positive immunoassay test results when using many of the commercially available methadone immunoassay kits and that confirmation of LAAM and its metabolites should be considered. PMID- 10517558 TI - Development and evaluation of an improved method for screening of amphetamines. AB - We developed a homogeneous immunoassay method to eliminate false-positive amphetamine results caused by cross-reactive substances, including over-the counter allergy and cold medications. This method uses a neutralizing antibody that binds to amphetamines but does not bind to the labeled amphetamine conjugate used in the assay. The amount of neutralizing antibody is sufficient to reduce the assay signal resulting from authentic amphetamine and methamphetamine, but not the signal resulting from cross-reactants. This concept was implemented using the CEDIA DAU Amphetamines assay on Hitachi 747 and 717 clinical chemistry analyzers. Urine samples were tested using the standard, unmodified reagents in one channel and reagents containing the neutralizing antibody in a second channel. The difference in rate between the two tests was calculated by the analyzer; true-positive samples showed a significantly greater decrease in assay signal in response to neutralizing antibody as compared with false-positive samples. The neutralization method was evaluated in two studies using 448 samples that tested positive in the initial CEDIA DAU Amphetamines screening test. The samples were separated into categories of 154 true-positive samples and 294 false positive samples based upon a secondary screen with the Abbott FPIA Amphetamines assay followed by gas chromatography-mass spectrometry (GC-MS) testing using the HHS (SAMHSA) cutoff criteria. The CEDIA neutralization test successfully identified all 154 of the GC-MS confirmed positive samples. The test successfully identified as false positive 251 out of the 294 (85.4%) samples that failed to confirm by GC-MS. PMID- 10517559 TI - Metabolic production of amphetamine following multidose administration of clobenzorex. AB - The interpretation of urine drug-testing results can have important forensic and legal implications. In particular, drugs that are metabolized to amphetamine or methamphetamine or both pose significant concerns. In this study, clobenzorex, an anorectic drug that is metabolized to d-amphetamine, was administered to five subjects. Each subject took 30 mg daily for seven days, and individual urine samples were collected ad lib for 14 days beginning on the first day the drug was administered. Urine pH, specific gravity, and creatinine values were determined for each sample. Gas chromatography-mass spectrometry (GC-MS) was used to determine the excretion profile of amphetamine and clobenzorex using a standard procedure for amphetamines with additional monitoring of ions at m/z 118, 125, and 364 for the detection of clobenzorex. Peak concentrations of amphetamine were found at 82 to 168 h after the first dose and ranged from approximately 2900 to 4700 ng/mL amphetamine. The use of a regioisomer (3-Cl-benzylamphetamine) as internal standard allowed for accurate quantitation of the parent drug. Peak concentrations of clobenzorex were found at 50 to 120 h after the first dose and ranged from approximately 8 to 47 ng/mL clobenzorex. However, in many samples, clobenzorex was not detected at all. This analysis revealed that the metabolite, (amphetamine) is present in much higher concentrations than the parent compound, clobenzorex. Yet even at peak amphetamine concentrations, the parent was not always detected (limit of detection 1 ng/mL). Thus, in the interpretation of amphetamine-positive drug-testing results, the absence of clobenzorex in the urine sample does not exclude the possibility of its use. PMID- 10517560 TI - Simultaneous determination of amphetamine, methamphetamine, methylenedioxyamphetamine (MDA), methylenedioxymethamphetamine (MDMA), and methylenedioxyethylamphetamine (MDEA) enantiomers by GC-MS. AB - A method is described for the simultaneous determination of the ratio of l- and d enantiomers of amphetamine, methamphetamine, 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxymethamphetamine (MDMA), and 3,4-methylenedioxyethylamphetamine (MDEA) in urine. The assay uses liquid-liquid extraction followed by derivatization with trifluoroacetyl-l-prolyl chloride (l-TPC) and analysis by gas chromatography-mass spectrometry. The assay was developed using prepared samples containing varying concentrations of each of the analytes over a range of percentages of each enantiomer. Results showed the method to provide accurate and reliable results in samples containing > or = 10 ng/mL amphetamine and methamphetamine and > or = 25 ng/mL MDA, MDMA, and MDEA. The assay was used to analyze urine samples from subjects of a controlled MDMA study. Results for each of the eight subjects showed a greater percentage of the l-enantiomer of MDMA initially, and the percentage increased with time postdose. Analysis of the metabolite MDA revealed that the proportion of d-enantiomer was initially greater than the l-enantiomer followed by a gradual increase in the proportion of l enantiomer until it exceeded the amount of the d-enantiomer. In all cases, the l MDA exceeded the d-MDA within the first 36 h postdose. PMID- 10517561 TI - Enantioselective gas chromatography-negative ion chemical ionization mass spectrometry for methylphenidate in human plasma. AB - Therapeutic doses of Ritalin, a racemic mixture of d- and l-threo-methyphenidate, result in low plasma concentrations of methylphenidate. In order to assess the safety and efficacy of methylphenidate, a sensitive analytical method is needed. A gas chromatography-negative ion chemical ionization mass spectrometry (GC-NCI MS) assay capable of measuring both d- and l-enantiomers in human plasma was developed and validated to support clinical studies involving administration of d,l-methylphenidate. d,l-Methylphenidate-d3 is added to 1-mL plasma samples. The plasma samples are made basic, mixed with isopropanol and extracted with hexane. The hexane extracts are then back-extracted into 0.1 N HCl. The acidified aqueous extract is made basic, cooled to ice temperature, and the methylphenidate derivatized with heptafluorobutyryl-l-prolyl chloride. The two diastereomeric derivatives are then extracted into hexane. The hexane extract is evaporated to dryness, reconstituted in ethyl acetate, and analyzed by GC-NCI-MS. This method can accurately (+/- 5% target) and precisely (< 11.1% coefficient of variation) quantitate enantiomers of threo-methylphenidate in human plasma and in the whole blood at concentrations ranging from 0.75 to 100 ng/mL. Plasma samples are stable for up to five freeze-thaw cycles when the duration of each cycle did not exceed 0.5 h. The drug degraded gradually when plasma samples were left at room temperature; a 6% loss at 3 h progressed to 17% at 12 h and to 35% at 24 h. Therefore, it is important that extraction of plasma samples begins within 0.5 h after samples are removed from the freezer. Whole blood stability results show that concentrations of methylphenidate in whole blood, with or without NaF, stored for up to 6 h at room temperature did not deviate from the target concentration by more than 13%. The derivatized methylphenidate in extract is stable at 4 degrees C for up to 10 days. PMID- 10517562 TI - Urinary excretion profiles of 11-nor-9-carboxy-delta9-tetrahydrocannabinol: a delta9-THCCOOH to creatinine ratio study. AB - Monitoring the major cannabinoid metabolite (delta9-THCCOOH) to creatinine ratio (M/C) has been used to predict new drug use. According to Huestis and Cone, the best accuracy (85.4%) for predicting new marijuana use was a ratio > or = 0.5 from two urine specimens collected at least 24 h apart. Manno et al. recommended an M/C ratio of > or = 1.5. Subjects with a history of chronic marijuana use were screened for cannabinoid use by immunoassay (50-ng/mL cutoff), and presumptive positives were confirmed by gas chromatography-mass spectrometry for delta9 THCCOOH (15-ng/mL cutoff). Creatinine was analyzed with a cutoff concentration of 25 mg/dL. The study objective was to apply the criteria from both groups of workers to determine if consecutive urine specimens (collected at least 24 h apart) positive for cannabinoids could be used to differentiate new marijuana use from the excretion of residual cannabinoid metabolite (delta9-THCCOOH) in an uncontrolled setting. Serial urine specimens (826) were collected from 26 individuals. Huestis and Cone and Manno et al. ratios indicated new drug use in 83% and 33% of serial urine specimens collected at least 24 h apart, respectively. Clinically, the Huestis and Cone ratio is recommended because of a lower false-negative rate (7.4%) than the Manno et al. false-negative rate (24%). In legal situations, we recommend using the Manno et al. ratio because of its lower false-positive rate (0.1%) as stated by Huestis and Cone. PMID- 10517563 TI - Long-term stability of abused drugs and antiabuse chemotherapeutical agents stored at -20 degrees C. AB - Stability is an important consideration in the use of specimens for accurate determination of analyte concentrations. To determine the long-term stability for analytes routinely analyzed by mass spectrometry in this laboratory, quality control (QC) results were plotted versus time. The time required for the initial concentration to reach a specified level of deviation (i.e., 15%) was then determined from the slopes. QCs were prepared at 1-3 concentrations in drug-free matrix and stored at approximately -20 degrees C; urines were fortified with 1% sodium fluoride; plasmas (except for cocaines) were prepared with heparin. For cocaine and metabolites, the plasma was either fortified with 2% sodium fluoride or with 2% sodium fluoride and 1 mg% physostigmine after adjustment of the plasma pH to 6.0. In urine, amphetamine, methamphetamine, codeine, morphine, benzoylecgonine (BZE), and 11-nor-9-carboxy-delta9-tetrahydrocannabinol (THCA) slopes did not exceed a 15% deviation before 852 days. Cocaine, however, reached a 15% reduction at 165 days. When cocaine and BZE were prepared in plasma with just 2% sodium fluoride, negative slopes reached 15% deviation within 154 and 111 days, respectively. Further fortification with physostigmide and adjustment of the pH extended this time frame significantly. Delta9-Tetrahydrocannabinol (THC) and THCA in plasma had negative slopes that deviated by 15% just prior to one year. l-Alpha-acetylmethadol (LAAM), methadone, and their N-demethylated metabolites in urine did not have any negative slopes exceeding 15% before 686 days. Several of the compounds had positive slopes. Those for 2-ethylidene-1,5 dimethyl-3,3-diphenylpyrrolidine reached the 15% mark within 98 days. Those for LAAM, norLAAM, and dinorLAAM were concentration dependent. The 25-ng/mL controls reached 15% at 158-216 days. The 700-ng/mL controls reached 15% at 784-1340 days. In plasma, only naltrexone and buprenorphine displayed negative slopes at all three concentrations, reaching the 15% mark as early as 576 and 272 days, respectively. LAAM, norLAAM, dinorLAAM, ibogaine, 6-beta-naltrexone, risperidone, and 9-OH-risperidone did not exceed a 15% deviation before 416 days. To attempt to validate this method, two sets of clinical plasma samples that had been analyzed for buprenorphine were reanalyzed 644 and 869 days after the initial analyses. Those reanalyzed after 644 days were not statistically different from initial analyses, whereas those stored for 869 days were statistically different (p < 0.05). As the average time to reach 15% deviation for the three concentrations of buprenorphine QCs was 782 days, this suggests that extrapolation of QC results gathered over time may provide a reliable method to estimate long-term stability limits for drugs stored under the same conditions as the QC samples. PMID- 10517564 TI - Tissue distribution of mirtazapine (Remeron) in postmortem cases. AB - Mirtazapine (Remeron) is a member of the relatively new class of tetracyclic antidepressants. There are published cases of mirtazapine's detection as an incidental finding in postmortem cases; however, case reports with associated tissue concentrations and interpretations are not available. This report documents the tissue distribution of mirtazapine in eight postmortem cases in which it was identified but did not contribute to the cause or manner of death. The following mean mirtazapine concentrations (milligrams per liter or milligrams per kilogram) were found: heart blood 0.12 (range, < 0.01-0.33, n = 7); peripheral blood 0.09 (< 0.01-0.14, n = 4); urine 0.61 (0.01-3.2, n = 7); liver 0.88 (0.04-3.6, n = 6), kidney 0.21 (0.02-0.48, n = 5); and bile 0.62 (0.11-1.6, n = 6). In each case, the mirtazapine concentration in heart blood was approximately equal to that of peripheral blood, indicating that postmortem redistribution was not a factor in evaluating postmortem blood concentrations in these cases. However, because the liver mirtazapine concentrations were 5-30 times the blood concentrations, the potential for postmortem redistribution cannot be excluded. Additionally, because urine concentrations of the parent compound were consistently greater than the blood concentrations, urine is an adequate screening specimen for mirtazapine. PMID- 10517565 TI - Distribution of mirtazapine (Remeron) in thirteen postmortem cases. AB - Mirtazapine is a new antidepressant agent that entered the United States market in April 1996. To date, the literature provides limited information about therapeutic blood concentrations and virtually no information about postmortem levels. The Los Angeles County Coroner's Toxicology Laboratory has encountered 13 cases where postmortem tissue distributions of mirtazapine were determined. The analysis of mirtazapine from postmortem specimens (2-mL sample size) consisted of an n-butylchloride basic extraction procedure with identification and quantitation on a gas chromatograph-nitrogen-phosphorus detector. Linearity was achieved from 0.025 mg/L to 3.0 mg/L with a limit of quantitation of 0.025 mg/L. Confirmation of mirtazapine was performed on a gas chromatograph-mass spectrometer by comparison with a pure analytical standard. The tissue distribution of mirtazapine are in the following concentration ranges: heart blood 0.03-0.57 mg/L (13 cases), femoral blood 0.04-0.24 mg/L (9 cases), vitreous 0.06-0.10 mg/L (3 cases), liver 0.32-2.1 mg/kg (12 cases), bile 0.40-6.6 mg/L (7 cases), urine 0.12-2.5 mg/L (11 cases), kidney 0.23 mg/kg (1 case), spleen 0.17 mg/kg (1 case), and gastric 0.001-2.7 mg total (9 cases). Mirtazapine was not implicated in the cause of death in any of the 13 cases studied. These cases are being presented to aid the forensic toxicologist in the evaluation of postmortem mirtazapine levels. PMID- 10517566 TI - Experience with a urine opiate screening and confirmation cutoff of 2000 ng/mL. AB - Until recently, most laboratories used an opiate immunoassay screening and confirmation cutoff value of 300 ng/mL for codeine and morphine detection by gas chromatography-mass spectrometry (GC-MS). The cutoff value for opiates was increased to 2000 ng/mL or higher in various laboratories because of concerns that small doses of codeine and foods containing poppy seeds would give a positive opiate-screening result. Workplace drug-testing programs in the U.S. raised the opiate cutoff value to 2000 ng/mL on 30 November 1998. The objective of this study is to describe the results of opiate testing of 8600 urine specimens collected over 24 months with a 2000-ng/mL screening and confirmation (codeine and morphine) cutoff value. Specimens were screened by the EMITdau opiate assay using an in-house 2000-ng/mL morphine calibrator. Presumptive positive findings (N = 621) were analyzed quantitatively by GC-MS for codeine and morphine. One hundred and eighty six urine specimens were positive for codeine and morphine (> 2000 ng/mL), 298 specimens were positive for codeine only (> 2000 ng/mL) and 26 specimens were positive for morphine only (> 2000 ng/mL). All remaining specimens had codeine and morphine values < 2000 ng/mL. The codeine and morphine confirmation rate in this program reduced from 7.1% in 1994-1996 (300 ng/mL cutoff) to 2.1% in 1997-1998 with a 2000-ng/mL cutoff value. The codeine only confirmation rate lowered from 6.6% (300-ng/mL cutoff) to 3.4% (2000-ng/mL cutoff). It was concluded that increasing opiate screening and codeine and morphine confirmation cutoff values led to > 300% reduction in the confirmed positive rate for codeine and morphine and a 47% reduction in codeine-only confirmations in a urine drug-testing program where codeine was the major opiate used. PMID- 10517567 TI - A fatality related to the veterinary anesthetic telazol. AB - A 45-year-old male veterinarian was found dead in bed. Police investigation showed no evidence of trauma or other suspicious circumstances. Autopsy was unremarkable except for cardiomegaly and hepatosplenomegaly. Toxicological analysis revealed the presence of Telazol and ketamine. Telazol is a veterinary anesthetic agent that is composed of equal parts of tiletamine and zolazepam. Tiletamine is a disassociative anesthetic similar to ketamine and phencyclidine, and zolazepam is a diazepine derivative tranquilizer used to minimize the muscle hypertonicity and seizures associated with tiletamine. Quantitation of tiletamine and zolazepam was performed using gas chromatography-mass spectrometry in the selected ion monitoring mode following a solid-phase extraction. Postmortem blood, urine, and liver concentrations of tiletamine were 295 ng/mL, 682 ng/mL, and 196 ng/g, respectively, whereas postmortem concentrations of zolazepam for the same tissues were 1.71 microg/mL, 1.33 microg/mL, and 15.5 microg/g, respectively. Blood and urine ketamine levels were 37 ng/mL and 381 ng/mL, respectively. The cause of death was ruled an acute mixed drug intoxication of tiletamine, zolazepam, and ketamine with the manner of death ruled as unclassified. PMID- 10517569 TI - Acute zolpidem overdose--report of two cases. AB - This report describes two cases of acute zolpidem overdose. The decedent in the first case was a 36-year-old female found dead in bed in her secured home. She had a history of psychiatric illness, including paranoid disorder, depression with panic episodes, and post-traumatic stress disorder. She was treated with risperidone and sertraline. Nine months prior to her death, the decedent was also prescribed zolpidem (Ambien). The postmortem examination revealed white foam within the larynx and upper trachea, which is indicative of pulmonary edema. Toxicological analyses of the urine showed the presence of caffeine, risperidone, and zolpidem. Subsequent quantitation of postmortem iliac serum revealed 5.6 microg/L of 9-hydroxyrisperidone and the following zolpidem concentrations: blood (subclavian), 4.5 mg/L; blood (iliac), 7.7 mg/L; vitreous humor, 1.6 mg/L; bile, 8.9 mg/L; urine, 1.2 mg/L; liver, 22.6 mg/kg; and gastric contents, 42 mg. The second case involved a 58-year old female, also found dead in bed, with white foam around her mouth. The decedent had a 25-year history of hypertension and mental illness--manic depression and schizophrenia. She was medicated with carbamazepine, naproxen, risperidone, and zolpidem. The postmortem examination revealed cardiomegaly, pulmonary edema, hepatomegaly, mild coronary atherosclerosis, and no signs of trauma. Toxicological analyses of the urine showed the presence of zolpidem and carbamazepine and metabolite. Zolpidem concentrations were as follows: blood (iliac), 1.6 mg/L; vitreous humor, 0.52 mg/L; bile, 2.6 mg/L; liver, 12 mg/kg; and gastric contents, 0.9 mg. The zolpidem blood concentrations of these cases are consistent with those of the previously published fatalities. The blood/vitreous humor ratios of zolpidem were 2.81 (subclavian) and 4.81 (iliac) in the first case and 3.08 (iliac) in the second case. These ratios, along with the sampling times of blood and vitreous humor for both cases, are not conclusive to indicate a definitive presence or absence of postmortem drug redistribution of zolpidem. The cause of death for both cases was determined to be acute zolpidem overdose, and manner of death was suicide. PMID- 10517568 TI - Distribution of topiramate in a medical examiner's case. AB - Topimarate (Topamax) is a novel antiepileptic drug. Its mode of action is multifactorial and involves blockage of voltage-dependent sodium channels. The drug was detected in a 15-year-old epileptic who died soon after switching seizure prescriptions. Topimarate was recovered by basic extraction with ethyl acetate and analyzed by gas chromatography-mass spectrometry using selected ion monitoring. Ions monitored were m/z 324 and m/z 110 for topiramate and m/z 98 for the internal standard mepivacane. The drug was quantitated in blood, vitreous humor, bile, stomach content, and liver: the concentrations were 8.9, 12.4, and 10.9 mg/L, 31 mg/total content, and 29 mg/kg, respectively. Topiramate was detected in urine but not quantitated. Other drugs identified in this case were 0.45 mg/L nordiazepam and 0.05 mg/L oxazepam in blood. No alcohol was detected in any of the specimens. The cause of death was seizure disorder with upper respiratory infection. The manner of death was determined as natural. To our knowledge, this is the first report of the presence of topiramate in postmortem specimens. PMID- 10517570 TI - The effect of consumption of Hempen Ale on urine cannabinoid screens. PMID- 10517571 TI - Special issue on the molecular genetics of streptomycetes. PMID- 10517572 TI - Forty years of genetics with Streptomyces: from in vivo through in vitro to in silico. PMID- 10517573 TI - Genetic instability associated with insertion of IS6100 into one end of the Streptomyces lividans chromosome. AB - Analysis of 548 recombinant strains of Streptomyces lividans carrying chromosomal insertions of IS6100 revealed that six mutants contained DNA amplifications. The amplifications differed in size but included IS6100 sequences. Hybridization with representative cosmid clones containing sequences from the unstable regions of the chromosome indicated that, in each mutant, DNA rearrangements affected just one of the chromosome ends. The amplifications were derived either from a region immediately proximal to the terminal inverted repeat (TIR) or further distal, from a previously characterized type I amplifiable unit of DNA. There was no evidence for extensive deletions accompanying the amplifications and chromosome linearity was maintained with, at least in five mutants, clear evidence for no loss of either TIR. The nature of the rearrangements provides evidence that insertions affecting the integrity of a chromosome end can contribute to genetic instability in Streptomyces. PMID- 10517574 TI - Streptomyces genomes: circular genetic maps from the linear chromosomes. AB - Streptomyces chromosomes are linear DNA molecules and yet their genetic maps based on linkage analysis are circular. The only other known examples of this phenomenon are in the bacteriophages T2 and T4, the linear genomic sequences of which are circularly permuted and terminally redundant, and in which replication intermediates include long concatemers. These structural and functional features are not found in Streptomyces. Instead, the circularity of Streptomyces genetic maps appears to be caused by a completely different mechanism postulated by Stahl & Steinberg (1964, Genetics 50, 531-538)--a strong bias toward even numbers of crossovers during recombination creates misleading genetic linkages between markers on the opposite arms of the chromosome. This was demonstrated by physical inspection of the telomeres in recombinant chromosomes after interspecies conjugation promoted by a linear or circular plasmid. The preference for even numbers of crossovers is probably demanded by the merozygosity of the recombining chromosomes, and by the association between the telomeres mediated by interactions of covalently bound terminal proteins. PMID- 10517576 TI - Association of early sporulation genes with suggested developmental decision points in Streptomyces coelicolor A3(2). AB - Cytological analysis of a series of Streptomyces coelicolor A3(2) mutants with disruptions of early sporulation (whi, for white aerial mycelium) genes in an isogenic background has provided new information about the role of whiH, and confirmed and extended previous knowledge about whiG, whiA and whiB. The characteristic straight aerial hyphae of whiG mutants contained normally spaced vegetative-like septa, while mutants in whiA or whiB had abnormally long and coiled aerial hyphae almost devoid of septation. whiG, whiA and whiB were all absolutely required for sporulation septation, and for all visible signs of nucleoid condensation and partitioning and other changes associated with later stages of sporulation. On the other hand, whiH appeared to enhance low basal levels of these processes. Thus, whiH mutant aerial hyphae were divided into loosely coiled fragments of variable sizes by what appeared to be a few sporulation septa. These fragments showed some spore-like characteristics and contained condensed and aberrantly partitioned nucleoids. whiG, whiA and whiB were epistatic to whiH on the criterion that they prevented such fragments from forming in double mutants. These spore-like features and the synthesis of clearly detectable levels of the whiE-directed grey spore pigment were not due to any residual activity of previously studied whiH alleles since they were retained by a constructed whiH null mutant. A model is presented that explains the mutant phenotypes by proposing two early developmental decision points involved in commitment to sporulation septation, one requiring whiG and the other requiring whiA and whiB. PMID- 10517575 TI - Green fluorescent protein as a reporter for spatial and temporal gene expression in Streptomyces coelicolor A3(2). AB - The enhanced green fluorescent protein (EGFP) gene is a modified version of the green fluorescent protein gene of the jellyfish Aequorea victoria with a codon usage that corresponds well to that found in many GC-rich streptomycete genes. Here the use of EGFP as a reporter for the analysis of spatially and temporally regulated gene expression in Streptomyces coelicolor A3(2) is demonstrated. The EGFP gene was inserted into plasmids that can replicate in Escherichia coli, greatly facilitating the construction of EGFP gene fusions. The plasmids can be transferred readily to S. coelicolor by conjugation, whereupon two of them (pIJ8630 and pIJ8660) integrate at the chromosomal attachment site for the temperate phage phiC31. These vectors were used to analyse the spatial and temporal expression of sigF, which encodes a sigma factor required for spore maturation, and of redD, a pathway-specific regulatory gene for the production of undecylprodigiosin, one of the four antibiotics made by S. coelicolor. While transcription of sigF appeared to be confined to developing and mature spore chains, transcription of redD occurred only in ageing substrate mycelium. A further plasmid derivative (pIJ8668) was made that lacks the phiC31 attachment site, allowing the EGFP gene to be fused transcriptionally to genes of interest at their native chromosomal locations. PMID- 10517577 TI - Characterization of the gene for factor C, an extracellular signal protein involved in morphological differentiation of Streptomyces griseus. AB - The gene encoding factor C (facC), an extracellular signal protein involved in cellular differentiation, was cloned from Streptomyces griseus 45H, and the complete nucleotide sequence was determined. The deduced amino acid sequence was confirmed by HPLC/electrospray ionization-mass spectrometry analysis. The full length protein consists of 324 amino acids and has a predicted molecular mass of 34,523 Da. The mature extracellular 286 amino acid protein (31,038 Da) is probably produced by cleaving off a 38 amino acid secretion signal sequence. Southern hybridization detected facC in several other Streptomyces strains, but database searches failed to identify a protein with significant homology to factor C. Expression of facC from a low-copy-number vector in S. griseus 52-1 resulted in a phenotypic effect similar to that given by exogenously added factor C protein. PMID- 10517579 TI - A putative regulatory element for carbon-source-dependent differentiation in Streptomyces griseus. AB - To identify negative regulatory genes for cellular differentiation in Streptomyces griseus, DNA fragments repressing the normal developmental processes were cloned on a high-copy-number plasmid. One of these DNA fragments markedly repressed aerial mycelium and spore formation on solid media containing glucose or galactose, but not on media containing maltose or mannitol. The fragment contained three complete ORFs; precise subcloning revealed that a 249 bp fragment located in the promoter region between ORF1 and ORF3 was sufficient for repression. Quantification of the promoter activities by using a thermostable malate dehydrogenase gene as a reporter showed that the promoter for ORF3 (P(ORF3)) maintained high activity in mycelia grown in the presence of glucose but lost activity rapidly in maltose medium. P(ORF3) activity increased markedly when the promoter sequence was introduced on a high-copy-number plasmid. The results suggested that carbon-source-dependent deactivation of P(ORF3) mediated by a transcriptional repressor may initiate differentiation in S. griseus. PMID- 10517578 TI - Four genes encoding different type I signal peptidases are organized in a cluster in Streptomyces lividans TK21. AB - Four adjacent genes (sipW, sipX, sipY and sipZ) encoding different type I signal peptidases, were isolated on a 7860 bp DNA fragment from Streptomyces lividans TK21. Three of the sip genes constitute an operon and the fourth is the first gene of another operon encompassing three additional, unrelated genes. A DNA fragment containing the four sip genes complemented an Escherichia coli type I signal peptidase mutant when cloned in a multicopy plasmid. Clustering of four different type I signal peptidase genes seems, so far, to be a unique feature of Streptomyces. PMID- 10517580 TI - Involvement of amfC in physiological and morphological development in Streptomyces coelicolor A3(2). AB - amfC plays a regulatory role in aerial mycelium formation in Streptomyces griseus and is distributed widely among Streptomyces species. Disruption of the chromosomal amfC gene in Streptomyces coelicolor A3(2) severely reduced formation of aerial hyphae, indicating that amfC is important in morphological development. In addition, the disruption caused S. coelicolor A3(2) M130 to produce much less actinorhodin, and to produce undecylprodigiosin at a later stage of growth, indicating that amfC also regulates secondary metabolism. S1 nuclease mapping showed that transcription of actII-ORF4, the pathway-specific transcriptional activator in the act gene cluster, was greatly reduced in the amfC disruptants. The defect in secondary metabolite formation was suppressed or overcome by a mutation in sre-1. Consequently, an amfC-disrupted strain derived from S. coelicolor A3(2) M145, an actinorhodin-overproducing strain due to the sre-1 mutation, still produced a large amount of actinorhodin. Extra copies of amfC in strains M130 and M145 did not change spore-chain morphology or secondary metabolite formation. However, the spores in these strains remained white even after prolonged incubation. Since only spore pigmentation was affected, all known whi genes, except whiE, responsible for the polyketide spore pigment formation, were assumed to function normally. S1 nuclease mapping showed that transcription of whiEP1, one of the promoters in the whiE locus, was reduced in S. coelicolor A3(2) containing extra copies of amfC. Introducing amfC into several other Streptomyces species, such as Streptomyces lividans, Streptomyces lavendulae and Streptomyces lipmanii, also abolished spore pigment formation. An increase in the amount of AmfC appeared to disturb the maturation of spores. PMID- 10517581 TI - An AfsK/AfsR system involved in the response of aerial mycelium formation to glucose in Streptomyces griseus. AB - In Streptomyces coelicolor A3(2), a protein serine/threonine kinase (AfsK) and its target protein (AfsR) control secondary metabolism. AfsK and AfsR homologues (AfsK-g and AfsR-g) from Streptomyces griseus showed high end-to-end similarity in amino acid sequence with the respective S. coelicolor A3(2) proteins, as determined by cloning and nucleotide sequencing. AfsK-g and a fusion protein between AfsK-g and thioredoxin (TRX-AfsK-g) produced in high yield as inclusion bodies in Escherichia coli were solubilized with urea, purified by column chromatography and then refolded to an active form by dialysis to gradually remove the urea. AfsR-g was also fused to glutathione S-transferase (GST-AfsR-g); the fusion product in the soluble fraction in E. coli was purified. Incubation of AfsK-g or TRX-AfsK-g in the presence of [gamma-32P]ATP yielded autophosphorylated products containing phosphoserine and phosphothreonine residues. In addition, TRX AfsK-g phosphorylated serine and threonine residues of GST-AfsR-g in the presence of [gamma-32P]ATP. Disruption of chromosomal afsK-g had no effect on A-factor or streptomycin production, irrespective of the culture conditions. The afsK-g disruptants did not form aerial mycelium or spores on media containing glucose at concentrations higher than 1%, but did form spores on mannitol- and glycerol containing media; this suggests that afsK-g is essential for morphogenesis in the presence of glucose. Introduction of afsK-g restored aerial mycelium formation in the disruptants. The phenotype of afsR-g disruptants was similar to that of afsK g disruptants; introduction of afsR-g restored the defect in aerial mycelium formation on glucose-containing medium. Thus the AfsK/AfsR system in S. griseus is conditionally needed for morphological differentiation, whereas in S. coelicolor A3(2) it is conditionally involved in secondary metabolism. PMID- 10517582 TI - Evidence that a single EF-Ts suffices for the recycling of multiple and divergent EF-Tu species in Streptomyces coelicolor A3(2) and Streptomyces ramocissimus. AB - The tsf genes from Streptomyces coelicolor A3(2) and Streptomyces ramocissimus, encoding the guanine-nucleotide exchange factor EF-Ts, were cloned and sequenced. Streptomycetes have multiple and highly divergent EF-Tu species, with EF-Tu1 and EF-Tu3 showing only about 65% amino acid sequence identity, and yet these can apparently interact with a single EF-Ts species. tsf lies in an operon with rpsB, which encodes ribosomal protein S2. The amino acid sequence of S2 from S. coelicolor differs from most other bacterial S2 homologues in having a C-terminal extension of 70 aa residues with a highly repetitive organization, the function of which is unknown. Transcription analysis of the rpsB-tsf operon of S. coelicolor by promoter probing, nuclease S1 mapping and Northern blotting revealed that the genes give rise to a bicistronic transcript from a single promoter upstream of rpsB. An attenuator was identified in the rpsB-tsf intergenic region; it results in an approximately 2:1 ratio of rpsB vs tsf transcripts. Although tuf1, encoding the major EF-Tu, is located in the rpsL ribosomal protein operon, an additional promoter in the fus-tuf1 intergenic region leads to a significant excess of EF-Tu over ribosomes. Most amino acid residues known from the Escherichia coli crystal structure of the EF-Tu-EF-Ts complex to be directly involved in interaction between the two elongation factors are conserved between E. coli and Streptomyces. However, whenever interaction residues in the EF-Tu moiety show divergence among Streptomyces EF-Tu1, EF-Tu2 and EF-Tu3, the single Streptomyces EF-Ts exhibits compensatory substitutions of the corresponding residues. These apparently enable productive interaction to occur with all three EF-Tus. PMID- 10517583 TI - Disruption of sblA in Streptomyces lividans permits expression of a heterologous alpha-amylase gene in the presence of glucose. AB - In a transposition mutant of Streptomyces lividans TK24, the usually glucose repressible expression of a heterologous alpha-amylase gene (aml) became resistant to glucose repression. The transposon had inserted into an ORF called sblA which encodes a 274 aa product sharing significant sequence similarities with various phosphatases that act on small phosphorylated substrates. sblA was transcribed as a monocistronic mRNA and its transcription was enhanced at the transition phase. Because its transcriptional and putative translational start points coincide, sblA is likely to be translated in the absence of a conventional RBS. The sblA-disrupted mutant is characterized by early growth arrest in glucose grown cultures and by partial relief of glucose repression of aml expression. PMID- 10517584 TI - Nitrogen metabolism in Streptomyces coelicolor A3(2): modification of glutamine synthetase I by an adenylyltransferase. AB - An internal adenylyltransferase gene (glnE) fragment from Streptomyces coelicolor was amplified using heterologous PCR primers derived from consensus motifs. The sequence had significant similarity to bacterial glnE genes, and included a motif typical of the C-terminal adenylyltransferase domain of glnE. glnE from S. coelicolor lies on the Asel-C fragment of the chromosome and is localized near glnA (encoding glutamine synthetase I, GSI) and glnII (encoding GSII). To analyse the function of glnE in S. coelicolor, glnE (S. coelicolor E4) and glnA (S. coelicolor HT107) gene replacement mutants were constructed. The GSI activity of the glnE mutant was not down-regulated after an ammonium shock. However, the GSI activity of the wild-type cells decreased to 60% of the original activity. The glnA mutant is not glutamine auxotrophic, but in the gamma-glutamyltransferase assay no GSI activity was detected in unshifted and shifted HT107 cells. By snake venom phosphodiesterase treatment the GSI activity in the wild-type can be reconstituted, whereas no alteration is observed in the E4 mutant. Additionally, the loss of short-term GSI regulation in the E4 mutant was accompanied by an increased glutamine:glutamate ratio. PMID- 10517585 TI - Genes encoding acyl-CoA dehydrogenase (AcdH) homologues from Streptomyces coelicolor and Streptomyces avermitilis provide insights into the metabolism of small branched-chain fatty acids and macrolide antibiotic production. AB - The cloning, using a PCR approach, of genes from both Streptomyces coelicolor and Streptomyces avermitilis encoding an acyl-CoA dehydrogenase (AcdH), putatively involved in the catabolism of branched-chain amino acids, is reported. The deduced amino acid sequences of both genes have a high similarity to prokaryotic and eukaryotic short-chain acyl-CoA dehydrogenases. When the S. coelicolor and S. avermitilis acyl-CoA dehydrogenase genes (acdH) were expressed in Escherichia coli, each of the AcdH flavoproteins was able to oxidize the branched-chain acyl CoA derivatives isobutyryl-CoA, isovaleryl-CoA and cyclohexylcarbonyl-CoA, as well as the short straight-chain acyl-CoAs n-butyryl-CoA and n-valeryl-CoA in vitro. NMR spectral data confirmed that the oxidized product of isobutyryl-CoA is methacrylyl-CoA, which is the expected product at the acyl-CoA dehydrogenase step in the catabolism of valine in streptomycetes. Disruption of the S. avermitilis acdH produced a mutant unable to grow on solid minimal medium containing valine, isoleucine or leucine as sole carbon sources. Feeding studies with 13C triple labelled isobutyrate revealed a significant decrease in the incorporation of label into the methylmalonyl-CoA-derived positions of avermectin in the acdH mutant. In contrast the mutation did not affect incorporation into the malonyl CoA-derived positions of avermectin. These results are consistent with the acdH gene encoding an acyl-CoA dehydrogenase with a broad substrate specificity that has a role in the catabolism of branched-chain amino acids in S. coelicolor and S. avermitilis. PMID- 10517587 TI - Genetic suppression analysis of non-antibiotic-producing mutants of the Streptomyces coelicolor absA locus. AB - The absA locus in Streptomyces coelicolor A3(2) was identified because mutations in it uncoupled sporulation from antibiotic synthesis: absA mutants failed to produce any of the four antibiotics characteristic of S. coelicolor. These mutants are now shown to contain point mutations in the absA1 gene which encodes the histidine kinase sensor-transmitter protein of a two-component signalling system. The absA1 non-antibiotic-producing mutants, which are unpigmented, spontaneously acquire pigmented colony sectors. Genetic analysis established that the pigmented sectors contain second-site suppressive mutations, sab (for suppressor of abs). Phenotypic characterization showed that sab strains produce all four antibiotics; some overproduce antibiotics and are designated Pha, for precocious hyperproduction of antibiotics. A set of sab mutations responsible for suppression was localized by plasmid-mediated and protoplast fusion mapping techniques to the vicinity of the absA locus. DNA cloned from this region was used to construct phage that could transduce sab mutations. Sequence analysis of sab strains defined sab mutations in both the absA1 gene and the absA2 gene; the latter encodes the two-component system's response regulator. PMID- 10517586 TI - A host-vector system for analysis and manipulation of rifamycin polyketide biosynthesis in Amycolatopsis mediterranei. AB - Modular polyketide synthases (PKSs) are a large family of multifunctional enzymes responsible for the biosynthesis of numerous bacterial natural products such as erythromycin and rifamycin. Advanced genetic analysis of these remarkable systems is often seriously hampered by the large size (>40 kb) of PKS gene clusters, and, notwithstanding their considerable fundamental and biotechnological significance, by the lack of suitable methods for engineering non-selectable modifications in chromosomally encoded PKS genes. The development of a facile host-vector strategy for genetic engineering of the rifamycin PKS in the producing organism, Amycolatopsis mediterranei S699, is described here. The genes encoding all 10 modules of the rifamycin PKS were replaced with a hygromycin-resistance marker gene. In a similar construction, only the first six modules of the PKS were replaced. The deletion hosts retained the ability to synthesize the primer unit 3 amino-5-hydroxybenzoic acid (AHBA), as judged by co-synthesis experiments with a mutant strain lacking AHBA synthase activity. Suicide plasmids carrying a short fragment from the 5' flanking end of the engineered deletion, an apramycin resistance marker gene, and suitably engineered PKS genes could be introduced via electroporation into the deletion hosts, resulting in the integration of PKS genes and biosynthesis of a reporter polyketide in quantities comparable to those produced by the wild-type organism. Since this strategy for engineering recombinant PKSs in A. mediterranei requires only a selectable single crossover and eliminates the need for tedious non-selectable double-crossover experiments, it makes rifamycin PKS an attractive target for extensive genetic manipulation. PMID- 10517588 TI - Dispensable ribosomal resistance to spiramycin conferred by srmA in the spiramycin producer Streptomyces ambofaciens. AB - Streptomyces ambofaciens produces the macrolide antibiotic spiramycin, an inhibitor of protein synthesis, and possesses multiple resistance mechanisms to the produced antibiotic. Several resistance determinants have been isolated from S. ambofaciens and studies with one of them, srmA, which hybridized with ermE (the erythromycin-resistance gene from Saccharopolyspora erythraea), are detailed here. The nucleotide sequence of srmA was determined and the mechanism by which its product confers resistance was characterized. The SrmA protein is a methyltransferase which introduces a single methyl group into A-2058 (Escherichia coli numbering scheme) in the large rRNA, thereby conferring an MLS (macrolide lincosamide-streptogramin type B) type I resistance phenotype. A mutant of S. ambofaciens in which srmA was inactivated was viable and still produced spiramycin, indicating that srmA is dispensable, at least in the presence of the other resistance determinants. PMID- 10517589 TI - The Streptomyces coelicolor A3(2) lipAR operon encodes an extracellular lipase and a new type of transcriptional regulator. AB - A region of the Streptomyces coelicolor A3(2) chromosome was identified and cloned by using as a probe the lipase gene from Streptomyces exfoliatus M11. The cloned region consisted of 6286 bp, and carried a complete lipase gene, lipA, as well as a gene encoding a transcriptional activator (lipR). The S. coelicolor A3(2) lipA gene encodes a functional extracellular lipase 82% identical to the S. exfoliatus M11 lipase; the partially purified S. coelicolor enzyme showed a preference for substrates of short to medium chain length. Transcription of lipA was completely dependent on the presence of lipR, and occurred from a single promoter similar to the lipA promoters of S. exfoliatus M11 and Streptomyces albus G. These three Streptomyces lipA promoters have well-conserved -10 and -35 regions, as well as additional conserved sequences upstream of the -35 region, which could function as targets for transcriptional activation by the cognate LipR regulators. The Streptomyces LipR activators are related to other bacterial regulators of a similar size, constituting a previously unidentified family of proteins that includes MalT, AcoK, AlkS, AfsR, five mycobacterial proteins of unknown function and some Streptomyces regulators in antibiotic synthesis clusters. A lipase-deficient strain of S. coelicolor was constructed and found to be slightly affected in production of the polyketide antibiotic actinorhodin. PMID- 10517591 TI - RheA, the repressor of hsp18 in Streptomyces albus G. AB - In Streptomyces albus, Hsp18, a protein belonging to the family of small heat shock proteins, can be detected only at high temperature. Disruption of orfY, located upstream and in the opposite orientation to hsp18, resulted in an elevated level of hsp18 mRNA at low temperature. Genetic and biochemical experiments indicated that the product of orfY, now called RheA (Repressor of hsp eighteen), directly represses hsp18. In Escherichia coli, an hsp18'-bgaB transcriptional fusion was repressed in a strain expressing S. albus RheA. DNA binding experiments with crude extracts of E. coli overproducing RheA indicated that RheA interacts specifically with the hsp18 promoter. Transcription analysis of rheA in the S. albus wild-type and in rheA mutant strains suggested that RheA represses transcription not only of hsp18 but also of rheA itself. PMID- 10517590 TI - End-product control of expression of branched-chain amino acid biosynthesis genes in Streptomyces coelicolor A3(2): paradoxical relationships between DNA sequence and regulatory phenotype. AB - The branched-chain protein amino acids isoleucine, valine and leucine can provide precursors for synthesis of complex polyketide secondary metabolites in streptomycetes; therefore the regulation of their own synthesis is of interest. DNA sequences upstream of ilvBNC, ilvD, leuA, leuB, ilvE and leuCD in Streptomyces coelicolor A3(2) have been obtained in this laboratory or as part of the S. coelicolor genome sequencing project. Upstream of ilvB and leuA, typical features of classical attenuator systems can be discerned, in particular hypothetical short ORFs with runs of Ile/Val/Leu and Leu codons, respectively. No such features are apparent upstream of other genes or gene clusters present. All five upstream regions were fused to xylE (encoding catechol dioxygenase, CO) as a reporter gene in the SCP2*-based low-copy-number vector pIJ2839. All wild-type regions showed strong depression of CO activity in the presence of all three branched-chain amino acids whether or not the attenuation features were present. By site-directed mutagenesis, the Ile/Val/Leu and Leu triplets in the putative attenuator peptides for ilvB and leuA were replaced by ones for other amino acids. In the case of ilvB, this had no effect at all; for leuA, the wild-type regulatory phenotype persisted in at least some experiments. It was concluded that (i) an unknown regulatory mechanism must be operating in the ilv/leu system of S. coelicolor A3(2) in place of classical attenuation; and (ii) it is unsafe to infer the functioning of a regulatory mechanism from sequence homologies alone. PMID- 10517592 TI - Characterization of a vanillic acid non-oxidative decarboxylation gene cluster from Streptomyces sp. D7. AB - The genetics of non-oxidative decarboxylation of aromatic acids are poorly understood in both prokaryotes and eukaryotes. Although such reactions have been observed in numerous micro-organisms acting on a variety of substrates, the genes encoding enzymes responsible for these processes have not, to our knowledge, been reported in the literature. Here, the isolation of a streptomycete from soil (Streptomyces sp. D7) which efficiently converts 4-hydroxy-3-methoxybenzoic acid (vanillic acid) to 2-methoxyphenol (guaiacol) is described. Protein two dimensional gel analysis revealed that several proteins were synthesized in response to vanillic acid. One of these was characterized by partial amino terminal sequencing, leading to the cloning of a gene cluster from a genomic DNA lambda phage library, consisting of three ORFs, vdcB (602 bp), vdcC (1424 bp) and vdcD (239 bp). Protein sequence comparisons suggest that the product of vdcB (201 aa) is similar to phenylacrylate decarboxylase of yeast; the putative products of vdcC (475 aa) and vdcD (80 aa) are similar to hypothetical proteins of unknown function from various micro-organisms, and are found in a similar cluster in Bacillus subtilis. Northern blot analysis revealed the synthesis of a 2.5 kb mRNA transcript in vanillic-acid-induced cells, suggesting that the cluster is under the control of a single inducible promoter. Expression of the entire vdc gene cluster in Streptomyces lividans 1326 as a heterologous host resulted in that strain acquiring the ability to decarboxylate vanillic acid to guaiacol non oxidatively. Both Streptomyces sp. strain D7 and recombinant S. lividans 1326 expressing the vdc gene cluster do not, however, decarboxylate structurally similar aromatic acids, suggesting that the system is specific for vanillic acid. This catabolic system may be useful as a component for pathway engineering research focused towards the production of valuable chemicals from forestry and agricultural by-products. PMID- 10517593 TI - Evidence for a general-purpose genotype in Candida albicans, highly prevalent in multiple geographical regions, patient types and types of infection. AB - Epidemiological studies, using the probe Ca3, have shown that in a given patient population a single cluster of genetically related Candida albicans isolates usually predominates. The authors have investigated whether these local clusters are part of a single group, geographically widespread and highly prevalent as an aetiological agent of various types of candidiasis. An unrooted neighbour-joining tree of 266 infection-causing C. albicans isolates (each from a different individual) from 12 geographical regions in 6 countries was created, based on genetic distances generated by Ca3 fingerprinting. Thirty-seven per cent of all isolates formed a single genetically homogeneous cluster (cluster A). The remainder of isolates were genetically diverse. Using the maximum branch length within cluster A as a cut-off, they could be divided into 37 groups, whose prevalence ranged between 0.3% and 9%. Strains from cluster A were highly prevalent in all but one geographical region, with a mean prevalence across all regions of 41%. When isolates were separated into groups based on patient characteristics or type of infection, strains from cluster A had a prevalence exceeding 27% in each group, and their mean prevalence was 43% across all patient characteristics. These data provide evidence that cluster A constitutes a general purpose genotype, which is geographically widespread and acts as a predominant aetiological agent of all forms of candidiasis in all categories of patients surveyed. PMID- 10517594 TI - A multicopper oxidase gene from Candida albicans: cloning, characterization and disruption. AB - A multicopper oxidase gene from the human pathogenic yeast Candida albicans was isolated and characterized. An open reading frame of 1872 bp, designated CaFET3, was identified, encoding a predicted protein of 624 amino acids and a molecular mass of 70.5 kDa. The identity between the deduced amino acid sequences of CaFET3 and the Saccharomyces cerevisiae FET3 gene is 55%. CaFET3 was localized on chromosome 6. A null mutant (fet3delta/fet3delta) was constructed by sequential gene disruption. Unlike the C. albicans SC5314 wild-type strain the fet3delta mutant was unable to grow in low-iron medium. The lack of growth of a S. cerevisiae fet3delta mutant in iron-limited medium was compensated by transformation with CaFET3. The null mutant strain showed no change in pathogenicity compared with the wild-type strain in the mouse model of systemic candidiasis. PMID- 10517595 TI - A novel copper-binding protein with characteristics of a metallothionein from a clinical isolate of Candida albicans. AB - It is known that clinical isolates of Candida albicans exhibit a high level of resistance to copper salts, although the molecular basis of this resistance is not clear. To investigate this, a novel copper-binding protein was purified from a clinical isolate of C. albicans. The protein was extracted from yeast cells after an induction period of 10 h in a copper-containing suspension medium. It was further purified by size-exclusion chromatography, ultrafiltration and reverse-phase HPLC. All protein fractions were analysed for their protein and copper contents. The copper/protein ratio increased steadily throughout the purification process; the most highly purified fraction showed a 210-fold increase compared to the whole-cell extract, with a recovery of 0.03%. The molecular mass of the protein was 10,000 Da and a reconstitution study using the purified apoprotein suggested that the equivalent extent of Cu(I) binding was approximately 14 mol eq. The amino-terminal segment of the copper-binding protein revealed three Cys-Xaa-Cys motifs, which is typical of a metallothionein (MT), and showed significant homology with mammalian MTs with respect to the positions of the cysteine residues. This is the first report of the isolation of a copper binding protein from C. albicans. PMID- 10517596 TI - Interaction of Salmonella choleraesuis, Salmonella dublin and Salmonella typhimurium with porcine and bovine terminal ileum in vivo. AB - Quantitative experiments on the interaction of Salmonella choleraesuis and Salmonella dublin with porcine and bovine intestinal epithelia yielded no evidence to suggest that host restriction of S. choleraesuis and S. dublin for pigs and calves respectively could be explained in terms of the patterns of intestinal invasion observed in ligated ileal loops in vivo, at 3 h after challenge. No evidence was found to support the idea that Peyer's patches, or specifically M cells, are the major route of entry for these serotypes in vivo. Three hours after loop inoculation, each serotype was recovered in comparable numbers from either absorptive or Peyer's patch mucosae present in the same ileal loop, indicating that both types of tissue are involved in the early stages of the enteropathogenic process induced by both serotypes. More detailed transmission electron microscopic (TEM) analyses of follicle-associated epithelia (FAE) challenged with S. choleraesuis showed that in the same region of FAE, organisms invaded both M cells and enterocytes directly; comparable detailed TEM studies with S. dublin could not be carried out because of the tissue-destructive properties of this serotype. S. dublin was clearly more histotoxic than S. choleraesuis as had previously been found in rabbits: this difference is almost certainly due to a tissue-damaging toxin which is neither host nor gut-tissue specific. The tissue-destructive potential of S. dublin has profound implications for the measurement of and the assignment of significance to the invasiveness of S. dublin. S. dublin was nearly always seen entering gut cells in micro-colonies whereas S. choleraesuis entered mainly as single organisms or small groups of two or three. PMID- 10517597 TI - Identification of O-antigen polymerase transcription and translation start signals and visualization of the protein in Salmonella enterica serovar Typhimurium. AB - The wzy/rfc gene, encoding the O-antigen polymerase, of Salmonella enterica serovar Typhimurium has been previously cloned and sequenced. In the present work, the wzy transcriptional startpoint was initially identified by primer extension. Next, wzy promoter strength in Escherichia coli K-12 was measured, and was found to be greater than that of the induced lac promoter. To define the Wzy translational startpoint, DNA including the wzy promoter and the putative first five residues of the Wzy protein was fused to the N-terminus of glutathione-S transferase, and the fusion protein purified by affinity chromatography. N terminal amino acid sequencing yielded the Wzy translational startpoint. Next, the Wzy protein was C-terminally tagged with the FLAG peptide, and immunoblotting of an S. typhimurium strain expressing a low-copy wzy-FLAG gene (five copies per cell) localized the intact Wzy protein in the cytoplasmic membrane of S. typhimurium cells. The Wzy protein was not well-expressed from a multi-copy wzy FLAG+ plasmid in S. typhimurium, or in E. coli K-12. PMID- 10517598 TI - Disruption of tonB in Bordetella bronchiseptica and Bordetella pertussis prevents utilization of ferric siderophores, haemin and haemoglobin as iron sources. AB - The Bordetella bronchiseptica tonB gene was cloned by detection of a chromosomal restriction fragment hybridizing with each of two degenerate oligonucleotides that corresponded to Pro-Glu and Pro-Lys repeats characteristic of known TonB proteins. The tonB(Bb) gene was situated upstream of exbB and exbD homologues and downstream of a putative Fur-regulated promoter. Hybridization results indicated that the tonB operon and flanking regions were highly conserved between B. bronchiseptica, Bordetella pertussis and Bordetella parapertussis. Disruption of tonB in B. bronchiseptica resulted in inability to grow in iron-limiting media, and inability to utilize alcaligin, enterobactin, ferrichrome, desferroxamine B, haemin and haemoglobin. Although it was not possible to inactivate tonB in a clinical B. pertussis isolate, tonB was disrupted in a laboratory B. pertussis strain previously selected for the ability to grow on Luria-Bertani medium. This B. pertussis tonB mutant shared a similar iron complex utilization deficient phenotype with the B. bronchiseptica tonB mutant. The B. bronchiseptica tonB operon present on a plasmid did not complement an Escherichia coli tonB mutant, but inefficient reconstitution of enterobactin utilization was observed in one fepA mutant harbouring plasmid copies of the B. pertussis fepA homologue and tonB(Bb) operon. PMID- 10517599 TI - A role for the PhoBR regulatory system homologue in the Vibrio cholerae phosphate limitation response and intestinal colonization. AB - To survive and multiply in different environments, Vibrio cholerae has to coordinately regulate the expression of genes involved in adaptive responses. In many pathogens, adaptive responses, including pathogenic responses, are regulated by two-component regulator (TCR) systems. It is likely that members of a TCR family play a role in the regulation of processes involved in intestinal colonization, and therefore pathogenesis, in V. cholerae. We have identified and characterized a TCR system of V. cholerae: this system is a homologue of Escherichia coli PhoBR. The presence of a putative Pho box suggests that the V. cholerae phoBR operon is regulated by inorganic phosphate levels. The phoR and phoB genes are organized the same way as in E. coli. Mutation of the V. cholerae phoB gene affected the expression of the putative Pho regulon, including PhoA, but did not affect the production of cholera toxin. V. cholerae phoB mutants are less able to colonize rabbit intestine than wild-type V. cholerae. The addition of inorganic phosphate at a high concentration to the inoculum only partially restored the ability of the mutants to colonize the intestine, suggesting that the V. cholerae Pho regulon in vivo may not be regulated by inorganic phosphate levels alone. PMID- 10517600 TI - Immunochemical characterization of an Ogawa-Inaba common antigenic determinant of Vibrio cholerae O1. AB - Cholera remains an important public health problem in many parts of the world and the availability of an effective cholera vaccine is important for the prevention of cholera in the countries affected by this disease. Despite the appearance in 1992 of a new serogroup, 0139, of Vibrio cholerae, most of the cholera outbreaks are still caused by V. cholerae O1 biotype El Tor. Vaccine trials in Asia from 1968 to 1971, and studies of the production of serotype-specific antiserum in rabbits and of the protective activity of monoclonal antibodies against diarrhoeal disease in neonatal mice, have led to the conclusion that the Ogawa serotype contains a specific antigenic determinant whereas the Inaba serotype contains a different antigenic determinant that cross-reacts with the Ogawa serotype. By studying the binding of anti-Ogawa monoclonal antibodies to synthetic oligosaccharide fragments mimicking the Ogawa O-specific polysaccharide, it has been shown that the terminal monosaccharide, bearing the 2 O-methyl group in the O-specific polysaccharide, is most probably the serotype specific determinant for the Ogawa strain. However, study of the binding of a monoclonal antibody recognizing both Ogawa and Inaba serotypes suggested partial recognition of the core as well as of the O-specific polysaccharide of the LPS of V. cholerae O1. To further characterize this antigenic determinant that is common to the Ogawa and Inaba serotypes, the core and the O-specific polysaccharide linked to the core of V. cholerae O1 LPS were purified by preparative electrophoresis. The O-specific polysaccharide linked to the core was subjected to periodate oxidation to destroy sugars from the core. Binding studies of these purified saccharide fragments to a monoclonal antibody which is protective in mice and specific to the antigenic determinant common to Ogawa and Inaba serotypes showed that both the core and the O-specific polysaccharide are involved in this common antigenic determinant. This explains how the presence or the absence of the Ogawa-specific antigenic determinant would lead to the expression of two independent antigenic determinants of V. cholerae O1, one specific to the Ogawa serotype and the other common to both Ogawa and Inaba serotypes. PMID- 10517601 TI - Genetic organization of the O7-specific lipopolysaccharide biosynthesis cluster of Escherichia coli VW187 (O7:K1). AB - In previous studies the authors cloned and characterized the DNA sequence of the regions at both ends of the O7-specific lipopolysaccharide (LPS) biosynthesis cluster of Escherichia coli VW187 (O7:K1), and identified the biosynthetic genes for dTDP-rhamnose and GDP-mannose, as well as one of the candidate glycosyltransferases. In this work the complete DNA sequence of a 6.9 kb intervening region is presented. Seven new ORFs were identified. All the functions required for the synthesis and transfer of the O7 LPS were assigned on the basis of complementation experiments of transposon insertion mutants, and amino acid sequence homology to proteins involved in LPS synthesis of other bacteria. Of the seven ORFs, two encoded membrane proteins that were homologous to the O-antigen translocase (Wzx) and polymerase (Wxy), two were involved in the biosynthesis of dTDP-N-acetylviosamine, and the remaining three showed homologies to sugar transferases. The O antigen chain length regulator gene wzz was also identified in the vicinity of the O7 polysaccharide cluster. O7-specific DNA primers were designed and tested for serotyping of O7 E. coli strains. PMID- 10517602 TI - Matrix-binding proteins of Staphylococcus aureus: functional analysis of mutant and hybrid molecules. AB - The fibrinogen-binding protein ClfA and the collagen-binding protein Cna are surface-associated adhesins of Staphylococcus aureus. ClfA has a dipeptide repeat region R composed mainly of serine and aspartate residues, more than 40 of which are required along with the 28-residue region W, the LPXTG motif and region M to display the ligand-binding region A on the cell surface in a functional form. Cna has a 61-residue region W and at least one 187-residue region B linking the collagen-binding region A to peptidoglycan. A cna mutant of S. aureus lacking region B was shown to bind collagen at the same level as wild-type Cna+ cells, indicating that region B is not necessary for ligand binding. Furthermore, altering the number of B repeats did not influence the level of collagen binding. In order to study the ability of C-terminal domains of Cna and ClfA to support functional ligand-binding activity of different adhesins, chimeric proteins were constructed and expressed in S. aureus. Surprisingly, the presence of a single Cna B domain and a nonapeptide linker located between ClfA region A and Cna region WM failed to support fibrinogen binding by S. aureus cells, despite the fact that ClfA region A was detected on the bacterial surface by immunoblotting. In contrast, the ClfA region A-Cna region B hybrid expressed as a recombinant protein in Escherichia coli did bind fibrinogen in Western ligand blots and in an ELISA-type assay. It is concluded that Cna region B cannot support functional display of ClfA region A on the bacterial cell surface. However, the ClfA dipeptide repeat region R and region WM did promote functional surface expression of the Cna collagen-binding domain in a hybrid Cna-ClfA protein. PMID- 10517603 TI - The Mycobacterium tuberculosis katG promoter region contains a novel upstream activator. AB - An Escherichia coli-mycobacterial shuttle vector, pJCluc, containing a luciferase reporter gene, was constructed and used to analyse the Mycobacterium tuberculosis katG promoter. A 1.9 kb region immediately upstream of katG promoted expression of the luciferase gene in E. coli and Mycobacterium smegmatis. A smaller promoter fragment (559 bp) promoted expression with equal efficiency, and was used in all further studies. Two transcription start sites were mapped by primer extension analysis to 47 and 56 bp upstream of the GTG initiation codon. Putative promoters associated with these show similarity to previously identified mycobacterial promoters. Deletions in the promoter fragment, introduced with BAL-31 nuclease and restriction endonucleases, revealed that a region between 559 and 448 bp upstream of the translation initiation codon, designated the upstream activator region (UAR), is essential for promoter activity in E. coli, and is required for optimal activity in M. smegmatis. The katG UAR was also able to increase expression from the Mycobacterium paratuberculosis P(AN) promoter 15-fold in E. coli and 12-fold in M. smegmatis. An alternative promoter is active in deletion constructs in which either the UAR or the katG promoters identified here are absent. Expression from the katG promoter peaks during late exponential phase, and declines during stationary phase. The promoter is induced by ascorbic acid, and is repressed by oxygen limitation and growth at elevated temperatures. The promoter constructs exhibited similar activities in Mycobacterium bovis BCG as they did in M. smegmatis. PMID- 10517604 TI - Molecular characterization of mycobacteria isolated from seals. AB - Tuberculosis (TB) was diagnosed in 10 seals from three species (Arctocephalus australis, Arctocephalus tropicalis and Otaria flavescens) found in South America. The mycobacteria isolated from these cases belonged to the Mycobacterium tuberculosis complex, as determined by RFLP using an IS6110 probe, spoligotyping, analysis of the 16S rRNA gene sequence and by PCR-restriction analysis of hsp65. Polymorphisms in gyrA, katG, oxyR and pncA were investigated in some of the isolates, as well as the presence of the MPB70 antigen. The insertion sequence IS6110 was present in three to seven copies in the genome of the mycobacteria isolated from seals. Using the IS6110 probe, six patterns (designated A, B, C, D, E and F) were identified from 10 different isolates. Patterns A and B were found for the mycobacteria isolated from two and four seals, respectively, indicating an epidemiological relationship between isolates grouped according to their IS6110 RFLP. The mycobacteria isolated from seals shared the majority of their IS6110 DNA-containing restriction fragments, and nine isolates had an identical spoligotype; only one isolate showed a minor difference in its spoligotype. In addition, none of these spoligotypes were found in other M. tuberculosis complex strains. These results suggest that the isolates from seals constitute a unique group of closely related strains. The mycobacteria isolated from seals showed polymorphisms at gyrA codon 95 and katG codon 463, as do group 1 M. tuberculosis, and M. bovis. Group 1 mycobacteria are associated with cluster cases. The spoligotypes found in the mycobacteria isolated from seals lack spacers 39-43, as does M. bovis, but the MPB70 antigen, which is highly expressed in M. bovis and minimally expressed in M. tuberculosis, was not detected in these mycobacteria. The mycobacteria isolated from seals also showed oxyR and pncA polymorphisms specific to M. tuberculosis. In conclusion, the mycobacteria that cause TB in seals in the South-Western Atlantic are a related group, and based on the combination of genetic characteristics, belong to a unique genotypic group within the M. tuberculosis complex. PMID- 10517605 TI - Purification and inhibition by quinolones of DNA gyrases from Mycobacterium avium, Mycobacterium smegmatis and Mycobacterium fortuitum bv. peregrinum. AB - The DNA gyrases from Mycobacterium avium, Mycobacterium smegmatis and Mycobacterium fortuitum bv. peregrinum, which are species naturally resistant, moderately susceptible and susceptible to fluoroquinolones, respectively, were purified by affinity chromatography on novobiocin-Sepharose columns. The DNA gyrase inhibiting activities (IC50 values) of classical quinolones and fluoroquinolones were determined from the purified enzymes and were compared to the corresponding antibacterial activities (MICs). Regarding M. fortuitum bv. peregrinum, which is nearly as susceptible as Escherichia coli, the corresponding MIC and IC50 values of quinolones were significantly lower than those found for M. avium and M. smegmatis (e.g. for ofloxacin, MICs of 0.25 versus 32 and 1 microg ml(-1), and IC50 values of 1 versus 8 and 6 microg ml(-1), respectively). Such a result could be related to the presence of Ser-83 in the quinolone resistance-determining region of the gyrase A subunit of M. fortuitum bv. peregrinum, as found in wild-type E. coli, instead of Ala-83 in M. avium and M. smegmatis, as found in fluoroquinolone-resistant E. coli mutants. The IC50 values of quinolones against the M. avium and M. smegmatis DNA gyrases were similar, while the corresponding MICs were 32-fold higher for M. avium when compared to M. smegmatis, suggesting that an additional mechanism, such as a low cell wall permeability or a drug efflux, could contribute to the low antibacterial potency of quinolones against M. avium. PMID- 10517606 TI - Characterization of haemagglutinin activity of Clostridium botulinum type C and D 16S toxins, and one subcomponent of haemagglutinin (HA1). AB - The 16S toxin and one subcomponent of haemagglutinin (HA), designated HA1, were purified from a type D culture of Clostridium botulinum by a newly established procedure, and their HA activities as well as that of purified type C 16S toxin were characterized. SDS-PAGE analysis indicated that the free HA1 forms a polymer with a molecular mass of approximately 200 kDa. Type C and D 16S toxins agglutinated human erythrocytes in the same manner. Their HA titres were dramatically reduced by employing erythrocytes that had been previously treated with neuraminidase, papain or proteinase K, and were inhibited by the addition of N-acetylneuraminic acid to the reaction mixtures. In a direct-binding test to glycolipids such as SPG (NeuAc alpha2-3Gal beta1-4GlcNAc beta1-3Gal beta1-4Glc beta1-Cer) and GM3 (NeuAc alpha2-3Gal beta1-4Glc beta1-Cer), and glycoproteins such as glycophorin A and/or B prepared from the erythrocytes, both toxins bound to sialylglycolipids and sialoglycoproteins, but bound to neither neutral glycolipids nor asialoglycoproteins. On the basis of these results, it was concluded that type C and D 165 toxins bind to erythrocytes through N acetylneuraminic acid. HA1 showed no haemagglutination activity, although it did bind to sialylglycolipids. We therefore speculate that binding to glycoproteins rather than to glycolipids may be important in causing haemagglutination by type C and D 16S toxins. PMID- 10517607 TI - Thermoprotection by glycine betaine and choline. AB - Glycine betaine is mostly known as an osmoprotectant. It is involved in the osmotic adaptation of eukaryotic and bacterial cells, and accumulates up to 1 M inside cells subjected to an osmotic upshock. Since, like other osmolytes, it can act as a protein stabilizer, its thermoprotectant properties were investigated. In vitro, like protein chaperones such as DnaK, glycine betaine and choline protect citrate synthase against thermodenaturation, and stimulate its renaturation after urea denaturation. In vivo, the internal concentration of glycine betaine is neither increased nor decreased after heat shock (this contrasts with a massive increase after osmotic upshock). However, even in exponential-phase bacteria grown in usual minimal salts media, the internal glycine betaine concentration attains levels (around 50 mM) which can protect proteins against thermodenaturation in vitro. Furthermore, glycine betaine and choline restore the viability of a dnaK deletion mutant at 42 degrees C, suggesting that glycine betaine not only acts as a thermoprotectant in vitro, but also acts as a thermoprotectant for Escherichia coli cells in vivo. PMID- 10517608 TI - Tellurite-mediated thiol oxidation in Escherichia coli. AB - The oxyanion of tellurium, tellurite (TeO3(2-)), is toxic to most micro organisms, particularly gram-negative bacteria. The mechanism of tellurite toxicity is presently unknown. Many heavy metals and oxyanions, including tellurite, interact with reduced thiols (RSH). To determine if tellurite interaction with RSH groups is involved in the toxicity mechanism, the RSH content of Escherichia coli cultures was assayed. After exposure to tellurite, cells were harvested and lysed in the presence of the RSH-specific reagent 5,5' dithiobis(2-nitrobenzoic acid). Upon exposure of tellurite-susceptible cells to TeO3(2-), the RSH content decreased markedly. Resistance to potassium tellurite (Te(r)) in gram-negative bacteria is encoded by plasmids of incompatibility groups IncFI, IncP alpha, IncHI2, IncHI3 and IncHII, as well as the tehAtehB operon from the E. coli chromosome. When cells harbouring a Te(r) determinant were exposed to TeO3(2-), only a small fraction of the RSH content became oxidized. In addition to tellurite-dependent thiol oxidation, the resistance of E. coli mutants affected in proteins involved in disulfide-bond formation (dsb) was investigated. Mutant strains of dsbA and dsbB were found to be hypersensitive to tellurite (MIC 0.008-0.015 microg K2TeO3 ml(-1) compared to wild-type E. coli with MICs of 1-2 microg K2TeO3 ml(-1)). In contrast, dsbC and dsbD mutants showed no hypersensitivity. The results suggest that hypersensitivity to tellurite is reliant on the presence of an isomerase activity and not the thiol oxidase activity of the Dsb proteins. The results establish that the Te(r) determinants play an important role in maintaining homeostasis of the intracellular reducing environment within gram-negative cells through specific reactions with either TeO3(2-) or thiol:tellurium products. PMID- 10517609 TI - Characterization and production of amylovorin L471, a bacteriocin purified from Lactobacillus amylovorus DCE 471 by a novel three-step method. AB - The strongly hydrophobic bacteriocin amylovorin L471 from Lactobacillus amylovorus DCE 471 was isolated and purified to homogeneity from complex culture broth by a novel, rapid and simple three-step protocol including (i) ammonium sulphate precipitation, (ii) chloroform/methanol extraction/precipitation and (iii) reversed-phase HPLC, the only chromatographic step involved. The molecular mass of the peptide was determined to be 4876.9 Da by electrospray mass spectrometric analysis. N-terminal amino acid sequencing identified 35 amino acid residues as being identical to the N-terminal sequence of lactobin A, a bacteriocin from another L. amylovorus strain. These non-identical strains produce bacteriocins that display small differences in molecular mass and inhibitory spectrum. The amino acid sequence of amylovorin L471 shared significant homology with lactacin X, one of the two bactericidal peptides produced by Lactobacillus johnsonii VPI11088. A purified amylovorin L471 preparation permitted confirmation of the inhibitory spectrum previously established with a crude extract. It displayed a bactericidal mode of action on lactobacilli after an extremely rapid adsorption to the target cells. Two Listeria spp. were only weakly sensitive. Amylovorin L471 appears to be produced constitutively. Ethanol not only stimulated specific bacteriocin production but also prevented adsorption of the bacteriocin molecules to the producer cells upon prolonged fermentation. The latter result supports the hypothesis that the apparent inactivation of bacteriocin observed during the stationary phase of batch fermentations is due to adsorption. PMID- 10517610 TI - Oxalic acid production by Aspergillus niger: an oxalate-non-producing mutant produces citric acid at pH 5 and in the presence of manganese. AB - The external pH appeared to be the main factor governing oxalic acid production by Aspergillus niger. A glucose-oxidase-negative mutant produced substantial amounts of oxalic acid as long as the pH of the culture was 3 or higher. When pH was decreased below 2, no oxalic acid was formed. The activity of oxaloacetate acetylhydrolase (OAH), the enzyme believed to be responsible for oxalate formation in A. niger, correlated with oxalate production. OAH was purified from A. niger and characterized. OAH cleaves oxaloacetate to oxalate and acetate, but A. niger never accumulated any acetate in the culture broth. Since an A. niger acuA mutant, which lacks acetyl-CoA synthase, did produce some acetate, wild-type A. niger is apparently able to catabolize acetate sufficiently fast to prevent its production. An A. niger mutant, prtF28, previously isolated in a screen for strains deficient in extracellular protease expression, was shown here to be oxalate non-producing. The prtF28 mutant lacked OAH, implying that OAH is the only enzyme involved in oxalate production in A. niger. In a traditional citric acid fermentation low pH and absence of Mn2+ are prerequisites. Remarkably, a strain lacking both glucose oxidase (goxC) and OAH (prtF) produced citric acid from sugar substrates in a regular synthetic medium at pH 5 and under these conditions production was completely insensitive to Mn2+. PMID- 10517611 TI - Active glycerol uptake is a mechanism underlying halotolerance in yeasts: a study of 42 species. AB - A comparison of 42 yeast species with respect to growth in the presence of high NaCl concentration and characteristics of glycerol uptake is presented. The yeast species were classified into four classes on the basis of their ability to grow in the presence of 1, 2, 3 or 4 M NaCl. Considering that two different types of active-transport systems for glycerol uptake have been described, Na+/glycerol and H+/glycerol symports, glycerol transport was investigated by testing for proton uptake upon glycerol addition in cells incubated in the absence and in the presence of NaCl. Only strains belonging to the two higher classes of salt tolerance showed constitutive active glycerol uptake, and could accumulate glycerol internally against a concentration gradient. Five of these strains exhibited a H+/glycerol symport. All the other strains showed evidence of the activity of a salt-dependent glycerol uptake similar to that described in the literature for Debraryomyces hansenii. The strains within the two lower classes of salt tolerance showed, to varying degrees, glycerol active uptake only when glycerol was used as the carbon and energy source, suggesting that this uptake system is involved in glycerol catabolism. The results within this work suggest that active glycerol uptake provides a basis for high halotolerance, helping to maintain a favourable intracellular concentration of glycerol. The relation between the constitutive expression of such carriers and a higher level of salt stress resistance suggests that this may be an evolutionary advantage for growth under such conditions. PMID- 10517612 TI - Enzyme polymorphism in Pseudomonas aeruginosa strains recovered from cystic fibrosis patients in France. AB - Each of 314 strains of Pseudomonas aeruginosa recovered from 87 French cystic fibrosis (CF) patients was typed by multilocus enzyme electrophoresis to investigate the genetic diversity, the relatedness and the molecular epidemiology of strains isolated from cases of chronic pulmonary colonization. Comparison of allele profiles at 18 enzyme loci identified 17 electrophoretic types (ETs). Of the 314 isolates, 290 (92%) were either ET1 (n = 127) or ET2 (n = 163), which differed only at the shikimate dehydrogenase (SKD) locus. The mean genetic diversity (H) was 0.138. These results suggest that there is cross-colonization between patients and/or that two predominant groups of strains are able to colonize French CF patients. Sequential isolates collected from 18 patients during a period of 12-28 months were analysed to assess genomic variability and its relationship to clinical outcome. Six patients were colonized by a stable strain. For the others, double infections or changes in colonization over time were observed. No relationships were detected between the clinical outcome and the persistence of stable isolates, the emergence of transient superinfecting variants, the presence of multiple ETs or the shift of ET during the monitoring. PMID- 10517613 TI - Contribution of adjuvant to adaptive immune responses in mice against Actinobacillus pleuropneumoniae. AB - The authors have previously demonstrated that adjuvant-mediated differences in early cellular responses to antigens significantly affect subsequent adaptive immune responses. To investigate further the contribution of adjuvant to adaptive immune responses, outer-membrane proteins (OMP) purified from the respiratory pathogen Actinobacillus pleuropneumoniae, given either alone (antigen group) or complexed with SAMA4 (vaccine group), were injected intradermally into groups of mice. Controls were given PBS. Inclusion of adjuvant did not significantly alter the kinetics of antibody responses against OMP in serum or respiratory tract washings (RTW) over 21 weeks. Re-exposure to OMP at 21 weeks also induced identical recall responses in both immunized groups. However, differences between the responses of the vaccine and antigen groups were apparent when sera and RTW were reacted against OMP and OMP-derived polysaccharide antigens (ODPA). Serum and RTW reactivity against protein antigens was stronger in the vaccine group than in the antigen group. Serum and RTW from the vaccine group also reacted against a greater number of proteins than did the antigen group. Although serum reactivity against ODPA was equivalent for both groups, RTW from the vaccine group reacted only faintly against ODPA compared with the antigen group. The results suggested that shifting of antibody reactivity away from polysaccharide antigens toward protein antigens was an adjuvant-mediated effect. The rapid death of controls following intranasal inoculation confirmed that protection was ultimately dependent on the presence of specific antibodies in the serum and respiratory tract. However, since both groups responded equally to intranasal infection with A. pleuropneumoniae, as seen by the rapid clearance of bacteria from the lungs, the biological significance of any differences between the groups was unclear. Knowledge of the effects of adjuvants may provide a rational basis for adjuvant selection and the ability to manipulate immunological outcomes more precisely. PMID- 10517614 TI - Genetic approaches to the identification of the mitis group within the genus Streptococcus. AB - The usefulness and reliability of partial sequence analysis of the manganese dependent superoxide dismutase gene (sodA), autolysin (lytA) gene amplification and species-specific PCR based on the D-alanine:D-alanine ligase (ddl) gene for differentiating each member of the mitis group of the genus Streptococcus was investigated. On the phylogenetic tree based on sodA partial sequences (366 bp) from 96 strains, including all species currently within the mitis group isolated in different geographic areas (mainly Japan and the UK), eight well separated clusters were generated corresponding to recognized species, and all strains fell into those clusters to which they had also been assigned by DNA-DNA hybridization. The Streptococcus pneumoniae sub-cluster was located within the Streptococcus mitis cluster, but the sodA gene of S. pneumoniae was very conserved and therefore could be separated from all other species examined. Furthermore, the lytA gene amplification approach could also be used to differentiate S. pneumoniae from other species. The species-specific amplification product of the ddl gene was successfully detected in Streptococcus sanguinis and Streptococcus gordonii, but failed to be detected in some strains of Streptococcus oralis including the type strain and S. mitis. We conclude that the partial sequence analysis of the sodA gene could be applied globally as a reliable and easy method for the accurate identification of all species currently within the mitis group. PMID- 10517615 TI - Plasmid transfer in the animal intestine and other dynamic bacterial populations: the role of community structure and environment. AB - The transfer of the R1drd19 plasmid between isogenic strains of Escherichia coli BJ4 in batch cultures of laboratory media and intestinal extracts was compared. Using an estimate of plasmid transfer rate that is independent of cell density, of donor:recipient ratios and of mating time, it was found that transfer occurs at a much lower rate in intestinal extracts than in laboratory media. Furthermore, the results suggest that the majority of intestinal plasmid transfer takes place in the viscous mucus layer covering the epithelial cells. Investigation of plasmid transfer in different flow systems harbouring a dynamic, continuously growing population of constant size showed that transfer kinetics were strongly influenced by bacterial biofilm formation. When donor and recipient populations were subjected to continuous mixing, as in a chemostat, transfer continued to occur at a constant rate. When donor and recipient populations retained fixed spatial locations, as in a biofilm, transfer occurred very rapidly in the initial phase, after which no further transfer was detected. From in vivo studies of plasmid transfer in the intestine of streptomycin-treated mice, results were obtained which were similar to those obtained in the biofilm, but differed markedly from those obtained in the chemostat. In spite of peristaltic movements in the gut, and of apparently even distribution of E. coli as single cells in the intestinal mucus, the intestinal environment displays transfer kinetics different from those expected of a mixed, liquid culture, but quite similar to those of a biofilm. PMID- 10517616 TI - Gene estimate rises as US and UK discuss freedom of access. PMID- 10517617 TI - Putting a price on research reading. PMID- 10517618 TI - Japan boosts proteomics and cell biology... PMID- 10517619 TI - While France gives more power to the centre. PMID- 10517620 TI - US and South Africa in pact over AIDS drug prices. PMID- 10517621 TI - Don't leave the biology out of bioinformatics. PMID- 10517622 TI - Courage could win back confidence in science. PMID- 10517623 TI - Sprucing up one's impact factor. PMID- 10517624 TI - Sprucing up one's impact factor. PMID- 10517625 TI - Call a halt to strong-arm tactics over GM crops. PMID- 10517626 TI - Genetics. Engineering a broken heart. PMID- 10517627 TI - Signal transduction. Mating, channels and kidney cysts. PMID- 10517628 TI - Discovery of tetraploidy in a mammal. PMID- 10517629 TI - Psychophysics. Putting plaids in perspective. PMID- 10517630 TI - Effect of vegetables on bone metabolism. PMID- 10517631 TI - Intron size and natural selection. PMID- 10517632 TI - Casein kinase I transduces Wnt signals. AB - The Wnt signalling cascade is essential for the development of both invertebrates and vertebrates, and is altered during tumorigenesis. Although a general framework for Wnt signalling has been elucidated, not all of the components have been identified. Here we describe a serine kinase, casein kinase I (CKI), which was isolated by expression cloning in Xenopus embryos. CKI reproduces several properties of Wnt signals, including generation of complete dorsal axes, stabilization of beta-catenin and induction of genes that are direct targets of Wnt signals. Dominant-negative forms of CKI and a pharmacological blocker of CKI inhibited Wnt signals in Xenopus. Inhibiting CKI in Caenorhabditis elegans generated worms with a mom phenotype, indicative of a loss of Wnt signals. In addition, CKI bound to and increased the phosphorylation of dishevelled, a known component of the Wnt pathway. These data indicate that CKI may be a conserved component of the Wnt pathway. PMID- 10517633 TI - Incorporating rules for responding into evolutionary games. AB - Evolutionary game theory is concerned with the evolutionarily stable outcomes of the process of natural selection. The theory is especially relevant when the fitness of an organism depends on the behaviour of other members of its population. Here we focus on the interaction between two organisms that have a conflict of interest. The standard approach to such two-player games is to assume that each player chooses a single action and that the evolutionarily stable action of each player is the best given the action of its opponent. We argue that, instead, most two-player games should be modelled as involving a series of interactions in which opponents negotiate the final outcome. Thus we should be concerned with evolutionarily stable negotiation rules rather than evolutionarily stable actions. The evolutionarily stable negotiation rule of each player is the best rule given the rule of its opponent. As we show, the action chosen as a result of the negotiation is not the best action given the action of the opponent. This conclusion necessitates a fundamental change in the way that evolutionary games are modelled. PMID- 10517634 TI - The liprin protein SYD-2 regulates the differentiation of presynaptic termini in C. elegans. AB - At synaptic junctions, specialized subcellular structures occur in both pre- and postsynaptic cells. Most presynaptic termini contain electron-dense membrane structures, often referred to as active zones, which function in vesicle docking and release. The components of those active zones and how they are formed are largely unknown. We report here that a mutation in the Caenorhabditis elegans syd 2 (for synapse-defective) gene causes a diffused localization of several presynaptic proteins and of a synaptic-vesicle membrane associated green fluorescent protein (GFP) marker. Ultrastructural analysis revealed that the active zones of syd-2 mutants were significantly lengthened, whereas the total number of vesicles per synapse and the number of vesicles at the prominent active zones were comparable to those in wild-type animals. Synaptic transmission is partially impaired in syd-2 mutants. syd-2 encodes a member of the liprin (for LAR-interacting protein) family of proteins which interact with LAR-type (for leukocyte common antigen related) receptor proteins with tyrosine phosphatase activity (RPTPs). SYD-2 protein is localized at presynaptic termini independently of the presence of vesicles, and functions cell autonomously. We propose that SYD 2 regulates the differentiation of presynaptic termini in particular the formation of the active zone, by acting as an intracellular anchor for RPTP signalling at synaptic junctions. PMID- 10517635 TI - Familial dementia caused by polymerization of mutant neuroserpin. AB - Aberrant protein processing with tissue deposition is associated with many common neurodegenerative disorders; however, the complex interplay of genetic and environmental factors has made it difficult to decipher the sequence of events linking protein aggregation with clinical disease. Substantial progress has been made toward understanding the pathophysiology of prototypical conformational diseases and protein polymerization in the superfamily of serine proteinase inhibitors (serpins). Here we describe a new disease, familial encephalopathy with neuroserpin inclusion bodies, characterized clinically as an autosomal dominantly inherited dementia, histologically by unique neuronal inclusion bodies and biochemically by polymers of the neuron-specific serpin, neuroserpin. We report the cosegregation of point mutations in the neuroserpin gene (PI12) with the disease in two families. The significance of one mutation, S49P, is evident from its homology to a previously described serpin mutations, whereas that of the other, S52R, is predicted by modelling of the serpin template. Our findings provide a molecular mechanism for a familial dementia and imply that inhibitors of protein polymerization may be effective therapies for this disorder and perhaps for other more common neurodegenerative diseases. PMID- 10517636 TI - Congenital heart disease in mice deficient for the DiGeorge syndrome region. AB - The heterozygous chromosome deletion within the band 22q11 (del22q11) is an important cause of congenital cardiovascular defects. It is the genetic basis of DiGeorge syndrome and causes the most common deletion syndrome in humans. Because the deleted region is largely conserved in the mouse, we were able to engineer a chromosome deletion (Df1) spanning a segment of the murine region homologous to the human deleted region. Here we describe heterozygously deleted (Df1/+) mice with cardiovascular abnormalities of the same type as those associated with del22q11; we have traced the embryological origin of these abnormalities to defective development of the fourth pharyngeal arch arteries. Genetic complementation of the deletion using a chromosome duplicated for the Df1 DNA segment corrects the heart defects, indicating that they are caused by reduced dosage of genes located within Df1. The Df1/+ mouse model reveals the pathogenic basis of the most clinically severe aspect of DiGeorge syndrome and uncovers a new mechanism leading to aortic arch abnormalities. These mutants represent a mouse model of a human deletion syndrome generated by chromosome engineering. PMID- 10517637 TI - Polycystin-L is a calcium-regulated cation channel permeable to calcium ions. AB - Polycystic kidney diseases are genetic disorders in which the renal parenchyma is progressively replaced by fluid-filled cysts. Two members of the polycystin family (polycystin-1 and -2) are mutated in autosomal dominant polycystic kidney disease (ADPKD), and polycystin-L is deleted in mice with renal and retinal defects. Polycystins are membrane proteins that share significant sequence homology, especially polycystin-2 and -L (50% identity and 71% similarity). The functions of the polycystins remain unknown. Here we show that polycystin-L is a calcium-modulated nonselective cation channel that is permeable to sodium, potassium and calcium ions. Patch-clamp experiments revealed single-channel activity with a unitary conductance of 137 pS. Channel activity was substantially increased when either the extracellular or intracellular calcium-ion concentration was raised, indicating that polycystin-L may act as a transducer of calcium-mediated signalling in vivo. Its large single-channel conductance and regulation by calcium ions distinguish it from other structurally related cation channels. PMID- 10517638 TI - A polycystic kidney-disease gene homologue required for male mating behaviour in C. elegans. AB - The stereotyped mating behaviour of the Caenorhabditis elegans male is made up of several substeps: response, backing, turning, vulva location, spicule insertion and sperm transfer. The complexity of this behaviour is reflected in the sexually dimorphic anatomy and nervous system. Behavioural functions have been assigned to most of the male-specific sensory neurons by means of cell ablations; for example, the hook sensory neurons HOA and HOB are specifically required for vulva location. We have investigated how sensory perception of the hermaphrodite by the C. elegans male controls mating behaviours. Here we identify a gene, lov-1 (for location of vulva), that is required for two male sensory behaviours: response and vulva location. lov-1 encodes a putative membrane protein with a mucin-like, serine-threonine-rich amino terminus followed by two blocks of homology to human polycystins, products of the autosomal dominant polycystic kidney-disease loci PKD1 and PKD2. LOV-1 is the closest C. elegans homologue of PKD1. lov-1 is expressed in adult males in sensory neurons of the rays, hook and head, which mediate response, vulva location, and potentially chemotaxis to hermaphrodites, respectively. PKD-2, the C. elegans homologue of PKD2, is localized to the same neurons as LOV-1, suggesting that they function in the same pathway. PMID- 10517639 TI - Dystrophin expression in the mdx mouse restored by stem cell transplantation. AB - The development of cell or gene therapies for diseases involving cells that are widely distributed throughout the body has been severely hampered by the inability to achieve the disseminated delivery of cells or genes to the affected tissues or organ. Here we report the results of bone marrow transplantation studies in the mdx mouse, an animal model of Duchenne's muscular dystrophy, which indicate that the intravenous injection of either normal haematopoietic stem cells or a novel population of muscle-derived stem cells into irradiated animals results in the reconstitution of the haematopoietic compartment of the transplanted recipients, the incorporation of donor-derived nuclei into muscle, and the partial restoration of dystrophin expression in the affected muscle. These results suggest that the transplantation of different stem cell populations, using the procedures of bone marrow transplantation, might provide an unanticipated avenue for treating muscular dystrophy as well as other diseases where the systemic delivery of therapeutic cells to sites throughout the body is critical. Our studies also suggest that the inherent developmental potential of stem cells isolated from diverse tissues or organs may be more similar than previously anticipated. PMID- 10517640 TI - Evidence that a free-running oscillator drives G1 events in the budding yeast cell cycle. AB - In yeast and somatic cells, mechanisms ensure cell-cycle events are initiated only when preceding events have been completed. In contrast, interruption of specific cell-cycle processes in early embryonic cells of many organisms does not affect the timing of subsequent events, indicating that cell-cycle events are triggered by a free-running cell-cycle oscillator. Here we present evidence for an independent cell-cycle oscillator in the budding yeast Saccharomyces cerevisiae. We observed periodic activation of events normally restricted to the G1 phase of the cell cycle, in cells lacking mitotic cyclin-dependent kinase activities that are essential for cell-cycle progression. As in embryonic cells, G1 events cycled on schedule, in the absence of S phase or mitosis, with a period similar to the cell-cycle time of wild-type cells. Oscillations of similar periodicity were observed in cells responding to mating pheromone in the absence of G1 cyclin (Cln)- and mitotic cyclin (Clb)-associated kinase activity, indicating that the oscillator may function independently of cyclin-dependent kinase dynamics. We also show that Clb-associated kinase activity is essential for ensuring dependencies by preventing the initiation of new G1 events when cell cycle progression is delayed. PMID- 10517641 TI - Mammalian XRCC2 promotes the repair of DNA double-strand breaks by homologous recombination. AB - The repair of DNA double-strand breaks is essential for cells to maintain their genomic integrity. Two major mechanisms are responsible for repairing these breaks in mammalian cells, non-homologous end-joining (NHEJ) and homologous recombination (HR): the importance of the former in mammalian cells is well established, whereas the role of the latter is just emerging. Homologous recombination is presumably promoted by an evolutionarily conserved group of genes termed the Rad52 epistasis group. An essential component of the HR pathway is the strand-exchange protein, known as RecA in bacteria or Rad51 in yeast. Several mammalian genes have been implicated in repair by homologous recombination on the basis of their sequence homology to yeast Rad51: one of these is human XRCC2. Here we show that XRCC2 is essential for the efficient repair of DNA double-strand breaks by homologous recombination between sister chromatids. We find that hamster cells deficient in XRCC2 show more than a 100 fold decrease in HR induced by double-strand breaks compared with the parental cell line. This defect is corrected to almost wild-type levels by transient transfection with a plasmid expressing XRCC2. The repair defect in XRCC2 mutant cells appears to be restricted to recombinational repair because NHEJ is normal. We conclude that XRCC2 is involved in the repair of DNA double-strand breaks by homologous recombination. PMID- 10517642 TI - Interpreting the folding kinetics of helical proteins. AB - The detailed mechanism of protein folding is one of the major problems in structural biology. Its solution is of practical as well as fundamental interest because of its possible role in utilizing the many sequences becoming available from genomic analysis. Although the Levinthal paradox (namely, that a polypeptide chain can find its unique native state in spite of the astronomical number of configurations in the denatured state) has been resolved, the reasons for the differences in the folding behaviour of individual proteins remain to be elucidated. Here a Calpha-based three-helix-bundle-like protein model with a realistic thermodynamic phase diagram is used to calculate several hundred folding trajectories. By varying a single parameter, the difference between the strength of native and non-native contacts, folding is changed from a diffusion collision mechanism to one that involves simultaneous collapse and partial secondary-structure formation, followed by reorganization to the native structure. Non-obligatory intermediates are important in the former, whereas there is an obligatory on-pathway intermediate in the latter. Our results provide a basis for understanding the range of folding behaviour that is observed in helical proteins. PMID- 10517643 TI - Keeping in touch with research. PMID- 10517645 TI - Occurrence, biosynthesis, and putative role of ecdysteroids in plants. PMID- 10517644 TI - Whither goest the RGS proteins? AB - Studies of the desensitization of G protein-coupled signal transduction have led to the discovery of a family of guanosine triphosphatase-activating proteins (GAPs) for heterotrimeric G protein alpha subunits - the "regulator of G protein signaling" or RGS proteins. In considering both documented and potential functions of several RGS protein family members with demonstrable multidomain compositions (p115RhoGEF, PDZRhoGEF, Axin, Axil/Conductin, D-AKAP2, the G protein coupled receptor kinases [GRKs], the DEP/GGL/RGS subfamily [RGS6, RGS7, RGS9, RGS11], and RGS12), this review explores the shift in our appreciation of the RGS proteins from unidimensional desensitizing agents to multifocal signal transduction regulators. PMID- 10517646 TI - Regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity by side chain oxysterols and their derivatives. PMID- 10517647 TI - Biochemistry and function of nematode steroids. PMID- 10517648 TI - Validation of a noninvasive measure of local myocardial repolarization in a conscious human model: adaptation of repolarization to changes in rate. AB - INTRODUCTION: A commercial pacemaker sensor measure of the unipolar endocardial stimulus to T wave interval may accurately reflect changes in the monophasic action potential duration at 90% repolarization (APD90). This sensor system was used to study the kinetics of adaptation of repolarization duration to changes in heart rate in humans. METHODS AND RESULTS: Patients were studied using an external pacemaker capable of displaying all stimulus to T wave intervals for each paced beat. Right ventricular stimulation was delivered via the pacemaker and compared simultaneously to APD90. Steady-state pacing was simulated by 60 seconds of pacing at cycle lengths (CLs) 350 to 700 msec. Adaptation to a new ventricular rate was analyzed with a sudden 200-msec decrease in CL. The relation between repolarization measure and steady-state CL (n = 16) was linear with a slope of 0.16 and 0.19 for APD90 and stimulus to T wave interval, respectively (P = NS). The adaptation of both repolarization measures to a sudden change in rate were best modeled by a biexponential function. Stimulus to T wave interval exhibited a parallel course to APD90, and an analysis of normalized differences between APD90 and stimulus to T wave interval followed an approximately normal distribution, with 93.5% of the paired differences within 2 SD of the mean. CONCLUSION: A pacemaker sensor measure of stimulus to T wave interval accurately parallels APD90 during both steady-state and sudden changes in rate. Repolarization in human endocardium follows a linear relation to steady-state CL and adapts to a new rate with a biexponential function. This model represents a novel method for studying human cardiac repolarization. PMID- 10517649 TI - Persistent atrial flutter in patients treated for atrial fibrillation with amiodarone and propafenone: electrophysiologic characteristics, radiofrequency catheter ablation, and risk prediction. AB - INTRODUCTION: Antiarrhythmic drugs have been reported to promote the conversion of atrial fibrillation to atrial flutter in patients with paroxysmal atrial fibrillation. However, information about the electrophysiologic mechanism and response to radiofrequency ablation of these drug-induced atrial flutters is limited. Furthermore, the determinants of the development of persistent atrial flutter in patients treated for atrial fibrillation with antiarrhythmic drugs are still unknown. METHODS AND RESULTS: Among the 136 patients treated for atrial fibrillation with amiodarone (n = 96) or propafenone (n = 40), 15 (11%, mean age 65.5 +/- 12.3 years) were identified to have subsequent development of persistent atrial flutter based on surface ECG characteristics during antiarrhythmic drug treatment. The mean interval between the beginning of drug treatment and the onset of atrial flutter was 5.0 +/- 5.5 months. Intracardiac mapping and entrainment studies revealed that 11 patients had counterclockwise typical atrial flutter, and 4 had clockwise typical atrial flutter. All 15 patients underwent successful ablation with creation of complete bidirectional isthmus conduction block. After a mean follow-up of 12.3 +/- 4.2 months, 14 (93%) of 15 patients who underwent successful ablation and continued taking antiarrhythmic drugs have remained in sinus rhythm. Univariate analysis of clinical variables demonstrated that only atrial enlargement was significantly related to the occurrence of persistent atrial flutter. CONCLUSION: In patients with atrial fibrillation, persistent typical atrial flutter might occur during antiarrhythmic drug treatment, and atrial enlargement was a risk factor for the development of such an arrhythmia. Radiofrequency ablation and continuation of pharmacologic therapy offered a safe and effective means of achieving and maintaining sinus rhythm. PMID- 10517650 TI - New approaches for the management of atrial fibrillation: role of ablation of atrial flutter. PMID- 10517651 TI - Differential effects of parasympathetic blockade and parasympathetic withdrawal on heart rate variability. AB - INTRODUCTION: Low heart rate variability (HRV) has been shown to have important prognostic significance in multiple settings. Although this is believed to reflect reduced parasympathetic tone, the physiology of reduced parasympathetic tone has not been elucidated. METHODS AND RESULTS: To evaluate whether parasympathetic withdrawal and partial parasympathetic blockade result in similar changes in HRV, 27 normal volunteers underwent complete beta-adrenergic blockade and then were given (1) graded doses of nitroprusside to achieve baroreflex mediated parasympathetic withdrawal and (2) low-dose atropine (0.01 mg/kg) to achieve partial parasympathetic blockade. Five-minute ECG recordings were obtained for HRV analysis. In 19 subjects, paired 5-minute recordings from each condition were available with mean RR intervals that differed by < 50 msec (low dose atropine: 869 +/- 96 msec and nitroprusside 875 +/- 99 msec). The root mean square of the successive RR interval differences was lower following low-dose atropine than following parasympathetic withdrawal with nitroprusside (16 +/- 11 msec vs 22 +/- 15 msec; P < 0.02). During parasympathetic withdrawal, the low frequency (LF) power was 0.917 +/- 0.602 bpm2 and the high-frequency (HF) power was 0.501 +/- 0.521 bpm2. During partial parasympathetic blockade, the LF and HF powers were significantly lower (0.443 +/- 0.474 bpm2, P < 0.005; and 0.198 +/- 0.207 bpm2, P < 0.02). CONCLUSION: These data confirm that HRV reflects the character of parasympathetic modulation of the heart rate rather than parasympathetic tone per se. Furthermore, this study identifies two distinct physiologic explanations for the finding of low HRV, namely, diminished vagal discharge and resistance of cardiac muscarinic receptors to vagal discharge. Further delineation of the relationships between parasympathetic tone and HRV will allow for better understanding of the pathophysiologic derangements associated with low HRV. PMID- 10517652 TI - Effect of the implantable atrial defibrillator on the natural history of atrial fibrillation. AB - INTRODUCTION: The purpose of our study was to evaluate the effect of repeated cardioversion with an implantable atrial defibrillator on the clinical outcome of patients with atrial fibrillation. METHODS AND RESULTS: The effects of the implantable atrial defibrillator on the total duration of atrial fibrillation, number of atrial fibrillation recurrences, and left atrial size were evaluated prospectively in 16 patients with atrial fibrillation (13 men and 3 women; mean age 58 +/- 11 years). Seven patients had no cardiovascular disease, 5 patients had hypertension, 3 patients had coronary heart disease, and 1 patient had congenital heart disease. Eight patients had paroxysmal atrial fibrillation for a mean duration of 80 +/- 61 months, and eight patients had persistent atrial fibrillation for a mean duration of 68 +/- 119 months. Except for one patient who received digoxin throughout the study, all patients received the same Class I or III antiarrhythmic agent throughout the study. The implantable atrial defibrillator successfully converted 50 (93%) of 54 spontaneous episodes of atrial fibrillation in 12 patients. During the initial 3 months of clinical follow-up, the atrial defibrillator documented 261 +/- 270 hours of atrial fibrillation compared with 126 +/- 172 hours (P = 0.01) during the subsequent 3 months. The left atrial size decreased from 4.4 +/- 0.7 cm at the time of atrial defibrillator implantation to 4.1 +/- 0.6 cm (P = 0.02) 6 months later. The number of atrial fibrillation recurrences did not change. These findings were observed in the absence of changes in drug therapy. No complications were observed. CONCLUSION: Restoration and maintenance of sinus rhythm in patients with atrial fibrillation by repeated cardioversion with an implantable atrial defibrillator was associated with a reduction in the total arrhythmia duration and a reduction in left atrial size. These results suggest that maintenance of sinus rhythm with the atrial defibrillator may reverse the remodeling process associated with atrial fibrillation. PMID- 10517653 TI - The natural history of atrial fibrillation: what is the role of atrial remodeling and what can we learn from the atrial defibrillator? PMID- 10517654 TI - Localization of the sites of conduction abnormalities in a mouse model of myotonic dystrophy. AB - INTRODUCTION: A mouse strain lacking functional myotonic dystrophy protein kinase (DMPK) has recently been developed. DMPK-/- mice exhibit muscular and conduction abnormalities consistent with the disease; however, the site of abnormal cardiac conduction is unknown. METHODS AND RESULTS: Nine homozygous DMPK-/- mice and seven age matched wild-type (WT) controls underwent in vivo electrophysiologic studies using an endocardial 2-French catheter. Baseline intervals as well as Wenckebach and 2:1 cycle lengths were measured to assess AV and ventriculoatrial (VA) conduction. Effective refractory periods (ERP) and functional refractory periods were determined during atrial and ventricular premature stimulation. His bundle recordings were obtained on all the studied animals (16/16). DMPK-/- mice had significantly prolonged PR (48.1 +/- 5.5 vs 40.9 +/- 3.9 msec, P = 0.010) and AH (36.7 +/- 4.0 vs 31.6 +/- 4.8 msec, P = 0.037) intervals compared to WT controls. HV intervals were very significantly prolonged as well (14.7 +/- 2.0 vs 10.3 +/- 0.8 msec; P < 0.0001). Three of 9 DMPK-/- and 1 of 7 WT mice exhibited VA block. Atrial ERP was reached before AV node ERP in 2 (22%) of 9 of the knockout mice and 5 (71%) of 7 of the controls (P = 0.06). Only one mouse (DMPK-/ ) exhibited infra-Hisian block on premature atrial stimulation. CONCLUSION: In this mouse model of myotonic dystrophy, AV conduction abnormalities were localized to the supra-Hisian and infra-Hisian conduction tissues, with a higher predilection to the latter, a finding similar to the human form of the disease. PMID- 10517655 TI - A transgenic model of myotonic dystrophy: will the mouse roar? PMID- 10517656 TI - Effects of mibefradil, a T-type calcium current antagonist, on electrophysiology of Purkinje fibers that survived in the infarcted canine heart. AB - INTRODUCTION: We studied the effects of mibefradil (MIB), a nondihydropyridine T type Ca2+ channel antagonist, on T- and L-type Ca2+ (I(CaT), I(CaL)) currents in Purkinje myocytes dispersed from the subendocardium of the left ventricle of normal (NZPC) and 48-hour infarcted (IZPC) hearts. METHODS AND RESULTS: Currents were recorded with Cs+- and EGTA-rich pipettes and in Na+-K+-free external solutions to eliminate overlapping currents. In all cells, I(Ca) was reduced by MIB (0.1 to 10 microM). No change in the time course of decay of peak I(Ca) was noted. Average peak T/L ratio decreased in NZPCs but not IZPCs with 1 microM MIB. Steady-state availability of I(CaL) was altered with 1 microM MIB in both cell types (mean +/- SEM) (V0.5 = -22 +/- 4 mV for NZPC and -25 +/- 5 mV for IZPC before drug; -63 +/- 9 mV for NZPC and -67 +/- 6 mV for IZPC after drug; P < 0.05). For I(CaT), V0.5 (-50 +/- 3 mV for NZPC and -52 +/- 1 mV for IZPC before drug) shifted to -60 +/- 2 mV (NZPC) and -62 +/- 3 mV (IZPC) (P < 0.05) after drug. We also determined the effects of MIB on spontaneously beating Purkinje normal fibers and on depolarized abnormally automatic fibers from the infarcted heart using standard microelectrode techniques. When NZPC and IZPC fibers were superfused with [K+]o = 2.7 mM, MIB 3 microM and 10 microM had no effect on rate or the maximum diastolic potential, but action potential plateau shifted to more negative values, the slope of repolarization phase 3 decreased, and action potential duration increased. CONCLUSION: MIB blocks L- and T-type Ca2+ currents in Purkinje myocytes but lacks an effect on either normal or abnormal automaticity in Purkinje fibers. PMID- 10517657 TI - Mibefradil, a T-type calcium channel blocker, and abnormal rhythm in subacute myocardial infarction. PMID- 10517658 TI - Canine ventricular myocyte beta2-adrenoceptors are not functionally coupled to L type calcium current. AB - INTRODUCTION: To establish the functional coupling of beta adrenoceptor (betaAR) subtypes beta1AR and beta2AR to L-type calcium current (I(CaL)), we investigated the nonselective agonist isoproterenol (ISO) and the relatively selective beta2AR agonists zinterol (ZIN) and salbutamol (SAL) on I(CaL) in isolated canine ventricular myocytes in the presence and absence of CGP 20712A (CGP) and atenolol (AT), selective beta1AR antagonists, and ICI 118,551 (ICI) a selective beta2AR antagonist. METHODS AND RESULTS: Peak I(CaL) was determined using "patch type" microelectrodes and whole cell voltage clamp. ISO (0.5 microM) increased I(CaL) maximally 3.5 +/- 0.67 fold. ZIN (10.0 microM) and SAL (10.0 microM) increased I(CaL) maximally 1.5 +/- 0.2 fold (n = 5) and 1.4 +/- 0.1 fold (n = 5), respectively. These effects were fully inhibited by CGP (0.3 microM) and AT (1.0 microM), which are inhibitors of beta1AR, but not by ICI (0.1 microM), which is a beta2AR inhibitor. ZIN at relatively lower concentrations (< or = 0.1 microM) did not increase I(CaL). CGP (0.3 microM) but not AT and ICI inhibited I(CaL) in the absence of betaAR agonists. CGP inhibition of I(CaL) was absent in the presence of forskolin (1.0 microM), which increases cAMP levels and I(CaL) by directly stimulating the adenylate cyclase. These data indicate that none of the antagonists affect I(CaL) through an action downstream of betaAR. CONCLUSION: Beta-adrenergic agonists increase I(CaL) via beta1AR but not beta2AR in canine ventricular myocytes. PMID- 10517659 TI - Sequential change in action potential of rabbit epicardium during and following radiofrequency ablation. AB - INTRODUCTION: Although radiofrequency (RF) catheter ablation is used to treat certain cardiac arrhythmias, little is known regarding transient changes in cellular electrophysiology during and following RF delivery. Optical recordings of action potential (OAP) with voltage-sensitive dyes allow immunity from electrical noise during RF delivery. The purpose of this study was to clarify the possible synergistic effects of both the thermal and electrotonic components of RF ablation. METHODS AND RESULTS: In this study, OAPs were recorded on the epicardium of 16 isolated Langendorff-perfused rabbit hearts within or adjacent to lesions made by RF catheters. Hearts were perfused at room temperature with Tyrode's solution containing 2,3-butanedione monoxime and stained by the voltage sensitive dye di-4-ANEPPS. OAPs were recorded before, during, and after RF pulses. Within the lesion, the action potential duration at 80% repolarization (APD80) of OAP decreased rapidly during the RF pulse, without recovery following the pulse. In the border zone surrounding the lesion, the RF energy resulted in a rapid decrease in APD80, which recovered promptly after the pulse (recovery time constant: 82 +/- 37 sec). APD80 was nonlinearly related to temperature during the RF ablation and responded faster to RF ablation than to purely thermal injury. CONCLUSION: The application of RF energy results in significant changes in myocardial cellular electrophysiologic properties. The RF energy has a combination of thermal and electrotonic effects on the myocardial tissue. The results of this in vitro study may illustrate the cellular basis for commonly observed phenomena in clinical practice. PMID- 10517662 TI - Electrophysiology of postinfarction ventricular tachycardia: a paradigm of stable reentry. AB - Sustained monomorphic ventricular tachycardia (VT) is a paradigm of a stable reentrant rhythm. The hallmark of stable reentry is the presence of an excitable gap, which in reentrant VT composes 15% to 45% of the tachycardia cycle length. Resetting allows definition of the extent and pattern of the excitable gap. Site specific resetting responses suggest that the VT circuit has both functionally and anatomically derived characteristics. Entrainment provides information regarding the effects of overdrive pacing on properties of the tissue composing the circuit rather than on properties of the tachycardia itself. These data help us to understand the mechanisms of pharmacologic agents and to direct ablation of reentrant VT. PMID- 10517660 TI - Characterization of a novel missense mutation in the pore of HERG in a patient with long QT syndrome. AB - INTRODUCTION: A new strategy to elucidate the molecular mechanisms underlying the long QT syndrome (LQTS) is now available with genetic mutational analyses and characterization of ion channel mutations. METHODS AND RESULTS: In a 26-year-old woman with LQTS, we identified a novel missense mutation in the pore of HERG by using polymerase chain reaction/single-strand conformation polymorphism (PCR/SSCP) and sequencing of her genomic DNA. The mutation resulted in an amino acid substitution of a positively charged lysine for a highly conserved uncharged asparagine at codon 629 (N629K). Whole cell, patch clamp studies were conducted in COS7 cells by transfecting with wild-type (WT) and/or the mutant N629K HERG. The WT HERG produced an I(Kr)-like, E-4031-sensitive conductance with an inward rectification. In contrast, the cells transfected with the N629K HERG did not display any time-dependent current. Cotransfection of WT and N629K HERG (at a ratio of 1:1) produced a significantly smaller conductance when compared with WT HERG (WT 59.9 +/- 7.3 pA/pF [n = 22] vs WT+N629K 5.5 +/- 2.3 pA/pF [n = 11]; P < 0.01), but did not alter K+ ion selectivity and tail current-voltage dependence. Because aprindine hydrochloride was effective in preventing ventricular tachycardias, we also tested the effect of the drug on WT HERG (I(Kr)) and KvLQT1/KCNE1 (I(Ks)) currents expressed in COS7. CONCLUSION: Functional analyses of a novel missense mutation in the pore of HERG suggest that the mutation causes marked reduction of I(Kr) via a dominant negative effect. PMID- 10517661 TI - Preventing sudden death after repair of tetralogy of Fallot: complex therapy for complex patients. AB - Sudden arrhythmic death in patients with repaired tetralogy of Fallot or its variants has a variety of causes. Consequently, it can serve as a paradigm for management of potentially malignant arrhythmias in all pediatric patients, particularly with regard to the use of nonpharmacologic therapy for management. Five cases are presented as touchpoints for discussion and demonstrate a number of important issues concerning the assessment and reduction of sudden cardiac death risk in these patients. First, there are no clinical parameters that can be used to accurately assess risk. Second, pharmacologic agents alone rarely are adequate therapy. Third, catheter ablation and antitachycardia devices continue to play an ever increasing role in management of these patients, and, finally, additional data are necessary to establish clear management guidelines in patients with congenital heart disease at risk for arrhythmic death. PMID- 10517663 TI - Narrow QRS complex tachycardia initiated by single spontaneous ventricular premature depolarizations: what is the tachycardia mechanism? PMID- 10517664 TI - Transmembrane responses of single guinea pig ventricular cell to uniform electric field stimulus. PMID- 10517665 TI - The M cell. PMID- 10517666 TI - Composite glucocorticoid regulation at a functionally defined negative glucocorticoid response element of the human corticotropin-releasing hormone gene. AB - Glucocorticoid-dependent negative feedback of the hypothalamic-pituitary-adrenal axis is mediated in part through direct inhibition of hypothalamic CRH gene transcription. In the present study, we sought to further localize and characterize glucocorticoid receptor (GR) and AP-1 interactions at a functionally defined negative glucocorticoid response element (nGRE) of the CRH promoter. Transient transfection studies in mouse corticotroph AtT-20 cells demonstrated that internal deletion of the nGRE (-278 to -249 nucleotides) within the context of 1 kb of the intact CRH promoter resulted in decreased 8-BrcAMP stimulation and glucocorticoid-dependent repression of CRH promoter activity. The nGRE conferred transcriptional activation by both cAMP and overexpressed c-jun or c-fos AP-1 nucleoproteins as well as specific glucocorticoid-dependent repression to a heterologous promoter. A similar profile of regulation was observed for the composite GRE derived from mouse proliferin promoter. The CRH nGRE was clearly distinct from the consensus cAMP response element (CRE) at -224 nucleotides, which increased basal activity and cAMP responsiveness of a heterologous promoter but did not confer glucocorticoid-dependent repression. High-affinity binding sites for both GR and AP-1 nucleoproteins were identified at adjacent elements within the nGRE. Mutations that disrupted either GR or AP-1 binding activity were associated with loss of glucocorticoid-dependent repression. These results are consistent with a composite mechanism of glucocorticoid-dependent repression involving direct DNA binding of GR and AP-1 nucleoproteins at discrete adjacent sites within the CRH promoter. PMID- 10517667 TI - RFG (ARA70, ELE1) interacts with the human androgen receptor in a ligand dependent fashion, but functions only weakly as a coactivator in cotransfection assays. AB - Abnormalities of the human androgen receptor (hAR) cause a range of clinical defects in male development. A large proportion of these mutations are single amino acid substitutions in the hormone-binding domain (HBD) that alter AR function by interfering with the capacity of the AR to bind androgen or to form stable hormone-receptor complexes. Prior studies have suggested that the formation of such stable, active hormone-receptor complexes is a crucial step in the modulation of genes by the AR. It is presumed that these hormone-receptor complexes interact with other proteins to participate in the formation of active transcription complexes at the initiation sites of androgen-responsive genes. Using a yeast two-hybrid screening method, we isolated a partial cDNA encoding the carboxy terminus of a protein that interacts with the hAR-HBD (amino acid residues 623-917) in a ligand-dependent fashion in a yeast two-hybrid assay. Sequence analysis of this clone revealed that it encoded a portion of a protein that had been previously characterized as RFG (RET Fused Gene). Using glutathione S-transferase (GST) fusions of the hAR HBD and immunoprecipitation of the in vitro translated proteins, we have demonstrated that this interaction can be reproduced in vitro. To determine the capacity of this protein to modulate the activity of the AR in transfection assays, we expressed full-length RFG in the CV1 and DU145 cell lines, in combination with an AR expression vector and model androgen-responsive genes [mouse mammary tumor virus (MMTV) and PRE2-tk luciferase]. Our results demonstrate that RFG alters the induction of these reporter genes very weakly (no greater than 2-fold compared with transfections without the RFG expression plasmid). Thus, while our findings are in agreement with published reports which indicate that RFG interacts with AR-HBD in a ligand dependent fashion, in our assays RFG does not exert major effects on the activity of the hAR in response to androgen or to other steroid hormones. PMID- 10517668 TI - Effect of overexpression of progesterone receptor A on endogenous progestin sensitive endpoints in breast cancer cells. AB - The human progesterone receptor (PR) is expressed as two isoforms, PRA and PRB, which differ in the N-terminal region and exhibit different activities in vitro, with PRA demonstrating dominant negative inhibitory effects on the activity of PRB and other nuclear receptors. PRA and PRB are expressed in target tissues at comparable levels although cells expressing a predominance of one isoform can be identified. In breast cancers, PRA is expressed at high levels in some tumors, and this may be associated with features of poorer prognosis. To investigate the role of PRA overexpression in PR-positive target cells, the effect of PRA induction on cell proliferation and expression of endogenous progestin-sensitive genes, SOX4 and fatty acid synthetase (FAS), was examined using PR-positive T-47D cell lines, which express a predominance of PRB, in which PRA could be increased 2- to 20-fold over basal levels. No effect of PRA induction was noted on cell proliferation, but marked changes in morphology, consistent with loss of adherent properties, were observed. Increases up to 4-fold in the relative PRA levels augmented progestin induction of SOX4 mRNA expression, and RU486 treatment revealed a progestin agonist effect. There was no consistent effect of PRA induction on progestin-mediated increases in FAS mRNA levels under these conditions. Clones with PRA:PRB ratios greater than 15 were associated with diminished progestin responses on both SOX4 and FAS mRNA expression. These data show that PRA overexpression is associated with alteration in adhesive properties in breast cancer cells and effects on endogenous progestin targets that were dependent on the cellular ratio of PRA:PRB. The results of this study are consistent with the view that PRA expression can fluctuate within a broad range in target cells without influencing the nature of progestin action on downstream targets, but that overexpression of PRA, such as is seen in a proportion of breast cancers, may be associated with inhibition of progestin action and features of poor prognosis. PMID- 10517669 TI - The estrogen receptor enhances AP-1 activity by two distinct mechanisms with different requirements for receptor transactivation functions. AB - Estrogen receptors (ERs alpha and beta) enhance transcription in response to estrogens by binding to estrogen response elements (EREs) within target genes and utilizing transactivation functions (AF-1 and AF-2) to recruit p160 coactivator proteins. The ERs also enhance transcription in response to estrogens and antiestrogens by modulating the activity of the AP-1 protein complex. Here, we examine the role of AF-1 and AF-2 in ER action at AP-1 sites. Estrogen responses at AP-1 sites require the integrity of the ERalpha AF-1 and AF-2 activation surfaces and the complementary surfaces on the p160 coactivator GRIP1 (glucocorticoid receptor interacting protein 1), the NID/AF-1 region, and NR boxes. Thus, estrogen-liganded ERalpha utilizes the same protein-protein contacts to transactivate at EREs and AP-1 sites. In contrast, antiestrogen responses are strongly inhibited by ERalpha AF-1 and weakly inhibited by AF-2. Indeed, ERalpha truncations that lack AF-1 enhance AP-1 activity in the presence of antiestrogens, but not estrogens. This phenotype resembles ERbeta, which naturally lacks constitutive AF-1 activity. We conclude that the ERs enhance AP-1 responsive transcription by distinct mechanisms with different requirements for ER transactivation functions. We suggest that estrogen-liganded ER enhances AP-1 activity via interactions with p160s and speculate that antiestrogen-liganded ER enhances AP-1 activity via interactions with corepressors. PMID- 10517670 TI - 1,25-Dihydroxyvitamin D3 increases nuclear vitamin D3 receptors by blocking ubiquitin/proteasome-mediated degradation in human skin. AB - 1,25-Dihydroxyvitamin D3 (D3) exerts its effects by binding to and activating nuclear vitamin D3 receptors (VDRs) that regulate transcription of target genes. We have investigated regulation of VDR levels in human skin in vivo and in cultured human keratinocytes. Quantitative ligand-binding analysis revealed that human skin expressed approximately 220 VDRs per cell, which bound D3 with high affinity [(dissociation constant (Kd) = 0.22 nM]. In human skin nuclear extracts, VDR exclusively bound to DNA containing vitamin D3 response elements as heterodimers with retinoid X receptors. Topical application of D3 to human skin elevated VDR protein levels 2-fold, as measured by both ligand-binding and DNA binding assays. In contrast, the D3 analog calcipotriene had no effect on VDR levels. Topical D3 had no effect on VDR mRNA, indicating that D3 either stimulated synthesis and/or inhibited degradation of VDRs. To investigate this latter possibility, recombinant VDRs were incubated with skin lysates in the presence or absence of D3. The presence of D3 substantially protected VDRs against degradation by human skin lysates. VDR degradation was inhibited by proteasome inhibitors, but not lysosome or serine protease inhibitors. In cultured keratinocytes, D3 or proteasome inhibitors increased VDR protein without affecting VDR mRNA levels. In cells, VDR was ubiquitinated and this ubiquitination was inhibited by D3. Proteasome inhibitors in combination with D3 enhanced VDR-mediated gene expression, as measured by induction of vitamin D3 24 hydroxylase mRNA in cultured keratinocytes. Taken together, our findings indicate that low VDR levels are maintained, in part, through ubiquitin/proteasome mediated degradation and that low VDR levels limit D3 signaling. D3 exerts dual positive influences on its nuclear receptor, simultaneously stimulating VDR transactivation activity and retarding VDR degradation. PMID- 10517671 TI - p120 acts as a specific coactivator for 9-cis-retinoic acid receptor (RXR) on peroxisome proliferator-activated receptor-gamma/RXR heterodimers. AB - p120 was originally isolated as a novel nuclear co-activator for thyroid hormone receptor. In this study, we characterized its interaction and transactivation of peroxisome proliferator-activated receptor-gamma (PPARgamma) and 9-cis-retinoic acid receptor (RXR) heterodimers. Transient transfection study revealed that p120 enhanced the transcriptional activation of PPARgamma/RXR induced by PPARgamma- or RXR-specific ligands. In the glutathione-S-transferase pull-down assay, while steroid receptor coactivator-1 showed apparent interactions with both RXR and PPARgamma, p120 bound only to RXR in a 9-cis-retinoic acid (RA)-dependent manner and also did not bind to PPARgamma even in the presence of thiazolidinediones. The yeast two-hybrid analysis showed no interaction of p120 with PPARgamma under any conditions, and electophoretic mobility shift assay showed apparent DNA PPARgamma/RXR/p120 complex formation only in the presence of 9-cis-RA. Furthermore, the yeast three-hybrid assay clearly revealed a significant interaction between p120 and PPARgamma via RXR of PPARgamma/RXR heterodimer only in the presence of 9-cis-RA. These findings indicate that p120 acts as a specific co-activator for the RXR of PPARgamma/RXR heterodimer in a 9-cis-RA-dependent manner. PMID- 10517672 TI - Members of the nuclear factor 1 transcription factor family regulate rat 3alpha hydroxysteroid/dihydrodiol dehydrogenase (3alpha-HSD/DD AKR1C9) gene expression: a member of the aldo-keto reductase superfamily. AB - Rat 3alpha-hydroxysteroid/dihydrodiol dehydrogenase (3alpha-HSD/DD; AKR1C9), a member of the aldo-keto reductase (AKR) superfamily, inactivates nearly all steroid hormones by converting 5alpha- and 5beta-dihydrosteroids to their respective 3alpha,5alpha- and 3alpha,5beta-tetrahydrosteroids and protects against circulating steroid hormone excess. It is highly expressed in rat liver comprising 0.5-1.0% of the soluble protein. Previously, we identified a powerful distal enhancer resident at about -4.0 kb to -2.0 kb in the 5'-flanking region of the 3alpha-HSD/DD gene. We now report the functional dissection of this enhancer. Transfection of nested deletions of the 5'-end of the gene promoter linked to chloramphenicol acetyltransferase (CAT) into HepG2 cells located the enhancer activity between (-4673 to -4179 bp). Further internal and 5'-end deletion mutants revealed that a 73-bp fragment (from -4351 to -4279 bp) contained a major enhancer element. This fragment spanned two imperfect direct repeats GTGGAAAAACCCAGGAA and GTGGAAAAAACCCAGGAA and contained three direct repeats of GGAAAAA. This fragment also contained three potential half-nuclear factor 1 (NF1) sites (TGGA-NNNNNGCCA) and a putative CCAAT-enhancer binding protein (C/EBP) binding site. The 73-bp fragment enhanced CAT activity from the basal 3alpha HSD/DD gene promoter. Recombinant C/EBPalpha and C/EBPbeta did not bind to this fragment. Electrophoretic mobility shift assays showed that HepG2 and rat liver nuclear extracts bound to this 73-bp fragment. The 73-bp protein complex was competed out by a NF1 oligonucleotide and was supershifted by an NF1 antibody. When the 73-bp fragment was fused to an alpha1-globin promoter-CAT construct and cotransfected with CCAAT transcription factor 1 (CTF1)/NF1 into Drosophila Schneider SL2 insect cells (which lack NF1-like proteins) trans-activation of CAT activity was observed. These results indicate that members of the NF1 transcription factor family regulate high constitutive expression of the rat 3alpha-HSD/DD gene that is responsible for steroid hormone inactivation. The potential role of NF1 in regulating other AKR genes that have protective roles is discussed. PMID- 10517673 TI - Dynamics of stimulus-expression coupling as revealed by monitoring of prolactin promoter-driven reporter activity in individual, living mammotropes. AB - Single-cell paradigms have greatly expanded our knowledge about stimulus secretion coupling, but the understanding of stimulus-gene expression coupling has lagged behind for lack of a dynamic model sufficiently sensitive to provide single-cell resolution. In the present study, we made continuous indirect measurements within individual, living cells of expression dynamics both before and after treatment with a gene-activating secretagogue. This was accomplished by transfecting (via microinjection) individual, primary mammotropes with a PRL promoter-driven luciferase reporter plasmid, and then quantifying the rate of photonic emissions (reflective of endogenous gene activity). We found that individual cells exhibit spontaneous, random, short-term fluctuations of basal reporter activity and are extremely heterogeneous in terms of responses to a stimulatory agent (TRH). In addition, we found that responses are affected by several factors including the secretory status of the pituitary donor, the manner in which the stimulus is presented, and by the initial level of reporter activity. Moreover, the responsiveness of an individual cell can fluctuate dramatically over time. These results invite speculation that a given cell can "sense" its gene activation state and regulate its response accordingly to satisfy requirements for the corresponding secretory product. PMID- 10517674 TI - Molecular components of large conductance calcium-activated potassium (BK) channels in mouse pituitary corticotropes. AB - Large-conductance calcium- and voltage- activated potassium (BK) channels play a fundamental role in the signaling pathways regulating mouse anterior pituitary corticotrope function. Here we describe the cloning and functional characterization of the components of mouse corticotrope BK channels. RT-PCR cloning and splice variant analysis of mouse AtT20 D16:16 corticotropes revealed robust expression of mslo transcripts encoding pore-forming alpha-subunits containing the mouse homolog of the 59-amino acid STREX-1 exon at splice site 2. RT-PCR and functional analysis, using the triterpenoid glycoside, DHS-1, revealed that native corticotrope BK channels are not functionally coupled to beta subunits in vivo. Functional expression of the STREX-1 containing alpha-subunit in HEK 293 cells resulted in BK channels with calcium sensitivity, single-channel conductance, and inhibition by protein kinase A identical to that of native mouse corticotrope BK channels. This report represents the first corticotrope ion channel to be characterized at the molecular level and demonstrates that mouse corticotrope BK channels are composed of alpha-subunits expressing the mouse STREX-1 exon. PMID- 10517675 TI - Calcitonin receptor-mediated growth suppression of HEK-293 cells is accompanied by induction of p21WAF1/CIP1 and G2/M arrest. AB - We investigated the mechanisms by which calcitonin (CT) suppresses cellular proliferation, using HEK-293 cells stably transfected with either the rat C1a CT receptor (CTR) or the insert-negative form of the human CTR. CT treatment of clonal cell lines expressing either receptor type, but not untransfected HEK-293 cells, strongly suppressed cell growth in a concentration-dependent manner. The reduction in cell growth with CT treatment could not be attributed to cellular necrosis or apoptotic cell death, the latter assessed by both DNA fragmentation analysis and caspase 3 (CPP-32) assay. Growth inhibition was associated with an accumulation of cells in the G2 phase of the cell cycle. CT treatment of the human and rat CTR-expressing cell lines resulted in a rapid and sustained induction of mRNA encoding the cyclin-dependent kinase inhibitor, p21WAF1/CIP1, increased levels of which were maintained at least 48 h after initiation of treatment. Western blot analysis showed a rapid corresponding increase in p21WAF1/CIP1 protein, whereas protein levels of another member of the cyclin dependent kinase inhibitor family, p27kip1, were unchanged. In parallel with the induction of p21, CT treatment reduced levels of p53 mRNA and protein. CT treatment resulted in a specific cell cycle block in G2, which was associated with inhibition of Cdc2/cyclin B kinase activity as measured by histone H1 phosphorylation. There was no evidence for p21 association with this complex despite the inhibition of Cdc2 activity. Evidence that p21 induction was causative of cell growth suppression was obtained from p21 antisense oligonucleotide experiments. Treatment with a p21 antisense oligonucleotide blocked induction of p21 expression and significantly reduced the CT-mediated growth inhibition. These observations suggest that p21 is required for the G2 arrest in response to CT, but argue against a direct role of p21 in the inhibition of Cdc2 activity. These studies suggest a novel regulation of cell cycle progression by CT and will provide a basis for detailed examination of the molecular mechanisms involved. PMID- 10517676 TI - Internalization and recycling pathways of the thyrotropin receptor. AB - Scant information is available to date on the intracellular trafficking of the TSH receptor. In the present study we have used stably transfected L cells that express the TSH receptor, 225I-labeled TSH, and antireceptor antibodies as well as gold-conjugated antireceptor monoclonal antibodies and hormone. The latter allowed us to study, by electron microscopy, the cellular distribution and endocytosis of TSH receptor. The receptor was initially localized on the plasmalemma proper and in clathrin-coated pits but was excluded from smooth vesicles open to the cell surface. It was internalized through clathrin-coated vesicles. Constitutive endocytosis represented 10% of cell surface receptor molecules. Endocytosis was increased 3-fold by incubation with hormone. The majority of internalized receptor molecules (90%) was recycled to the cell surface, whereas the hormone was degraded in lysosomes. This recycling of receptor was inhibited by administration of monensin. Electron microscopic and confocal microscopic studies were repeated in primary cultures of human thyroid cells and showed a distribution, and endocytosis pathways, very similar to those observed in transfected L cells. A previous study has shown the LH receptor to be endocytosed in high proportion and to be degraded in lysosomes. Confocal microscopy and colocalization studies with transferrin receptor confirmed that the highly homologous LH and TSH receptors exhibit, when expressed in the same cells, very different cellular trafficking properties. The use of LH/TSH receptor chimeras showed that transmembrane-intracellular domains contain information orienting the protein toward recycling or degradative pathways. The extracellular domain seems to play a role in the extent of intemalization. These observations should now allow the identification of the molecular signals involved. PMID- 10517677 TI - Dependence of insulin-stimulated glucose transporter 4 translocation on 3 phosphoinositide-dependent protein kinase-1 and its target threonine-410 in the activation loop of protein kinase C-zeta. AB - Previous studies have suggested that 1) atypical protein kinase C (PKC) isoforms are required for insulin stimulation of glucose transport, and 2) 3 phosphoinositide-dependent protein kinase-1 (PDK-1) is required for activation of atypical PKCs. Presently, we evaluated the role of PDK-1, both in the activation of PKC-zeta, and the translocation of epitope-tagged glucose transporter 4 (GLUT4) to the plasma membrane, during insulin action in transiently transfected rat adipocytes. Overexpression of wild-type PDK-1 provoked increases in the activity of cotransfected hemagglutinin (HA)-tagged PKC-zeta and concomitantly enhanced HA-tagged GLUT4 translocation. Expression of both kinase-inactive PDK-1 and an activation-resistant form of PKC-zeta that is mutated at Thr-410, the immediate target of PDK-1 in the activation loop of PKC-zeta, inhibited insulin induced increases in both HA-PKC-zeta activity and HA-GLUT4 translocation to the same extent as kinase-inactive PKC-zeta. Moreover, the inhibitory effects of kinase-inactive PDK-1 were fully reversed by cotransfection of wild-type PDK-1 and partly reversed by wild-type PKC-zeta, but not by wild-type PKB. In contrast to the T410A PKC-zeta mutant, an analogous double mutant of PKB (T308A/S473A) that is resistant to PDK-1 activation had only a small effect on insulin stimulated HA-GLUT4 translocation and did not inhibit HA-GLUT4 translocation induced by overexpression of wild-type PDK-1. Our findings suggest that both PDK 1 and its downstream target, Thr-410 in the activation loop of PKC-zeta, are required for insulin-stimulated glucose transport. PMID- 10517678 TI - Effect of an Asp905Tyr mutation of the glycogen-associated regulatory subunit of protein phosphatase-1 on the regulation of glycogen synthesis by insulin and cyclic adenosine 3',5'-monophosphate agonists. AB - The glycogen-associated regulatory subunit of protein phosphatase-1 (PP-1G) plays a major role in insulin-stimulated glycogen synthesis and thus the regulation of nonoxidative glucose disposal in skeletal muscle. In a general population of Caucasians a polymorphism at codon 905 of PP-1G from an aspartate to tyrosine has been reported to be associated with insulin resistance and hypersecretion. In this report functional studies were performed on rat skeletal muscle L6 cells stably transfected with an Asp905Tyr mutant PP-1G to evaluate the impact of this mutation on cellular responsiveness to insulin and cAMP. Although transfection resulted in a 3-fold increase in mutant PP-1G subunit expression, basal and insulin-stimulated PP-1 catalytic activities were decreased when compared with L6 cells transfected with wild-type PP-1G. The Asp905Tyr mutation resulted in an increase in cellular sensitivity to cAMP agonist, resulting in an inhibition of insulin's stimulatory effect on glycogen synthesis. More importantly, low concentrations of (Bu)2cAMP completely reversed insulin's stimulatory effects on glycogen synthesis when added to insulin-treated cells expressing mutant PP-1G. This was due to a rapid activation of glycogen phosphorylase a and a simultaneous inactivation of glycogen synthase via cAMP-mediated reductions in insulin stimulated PP-1 catalytic activities. We conclude that an Asp905Tyr mutation of PP-1G is accompanied by a relative increase in sensitivity to cAMP agonists as well as a diminished capacity of the mutant PP-1G to effectively mediate the inhibitory effects of insulin on glycogen breakdown via PP-1 activation. PMID- 10517680 TI - The cosmetic uses of Botulinum toxin type A. PMID- 10517681 TI - Waardenburg syndrome. PMID- 10517679 TI - Differential modulation of the tyrosine phosphorylation state of the insulin receptor by IRS (insulin receptor subunit) proteins. AB - In response to insulin, tyrosine kinase activity of the insulin receptor is stimulated, leading to autophosphorylation and tyrosine phosphorylation of proteins including insulin receptor subunit (IRS)-1, IRS-2, and Shc. Phosphorylation of these proteins leads to activation of downstream events that mediate insulin action. Insulin receptor kinase activity is requisite for the biological effects of insulin, and understanding regulation of insulin receptor phosphorylation and kinase activity is essential to understanding insulin action. Receptor tyrosine kinase activity may be altered by direct changes in tyrosine kinase activity, itself, or by dephosphorylation of the insulin receptor by protein-tyrosine phosphatases. After 1 min of insulin stimulation, the insulin receptor was tyrosine phosphorylated 8-fold more and Shc was phosphorylated 50% less in 32D cells containing both IRS-1 and insulin receptors (32D/IR+IRS-1) than in 32D cells containing only insulin receptors (32D/IR), insulin receptors and IRS-2 (32D/IR+IRS-2), or insulin receptors and a form of IRS-1 that cannot be phosphorylated on tyrosine residues (32D/IR+IRS-1F18). Therefore, IRS-1 and IRS-2 appeared to have different effects on insulin receptor phosphorylation and downstream signaling. Preincubation of cells with pervanadate greatly decreased protein-tyrosine phosphatase activity in all four cell lines. After pervanadate treatment, tyrosine phosphorylation of insulin receptors in insulin-treated 32D/IR, 32D/ IR+IRS-2, and 32D/IR+IRS-1F18 cells was markedly increased, but pervanadate had no effect on insulin receptor phosphorylation in 32D/IR+IRS-1 cells. The presence of tyrosine-phosphorylated IRS-1 appears to increase insulin receptor tyrosine phosphorylation and potentially tyrosine kinase activity via inhibition of protein-tyrosine phosphatase(s). This effect of IRS-1 on insulin receptor phosphorylation is unique to IRS-1, as IRS-2 had no effect on insulin receptor tyrosine phosphorylation. Therefore, IRS-1 and IRS-2 appear to function differently in their effects on signaling downstream of the insulin receptor. IRS 1 may play a major role in regulating insulin receptor phosphorylation and enhancing downstream signaling after insulin stimulation. PMID- 10517682 TI - Molluscum contagiosum: its clinical, histopathologic, and immunohistochemical spectrum. PMID- 10517683 TI - Neuropeptides and skin disorders. The new frontiers of neuro-endocrine-cutaneous immunology. PMID- 10517685 TI - Childhood, neonatal, and stillborn pemphigus vulgaris. AB - BACKGROUND: Childhood, neonatal, and stillborn cases of pemphigus vulgaris were reviewed. METHODS: From an examination of the pemphigus vulgaris literature, 46 childhood cases, nine neonatal cases, and three stillborn cases were found and investigated. RESULTS: In the childhood cases, the ratio between the sexes is approximately the same. The mean age of onset is 12 years, with only 11 children being 10 years of age or younger. Some children were treated with adjuvants to corticosteroids, most of them with azathioprine. While only one childhood case has been reported as fatal, the long-term prognosis and the relationship of early treatment and outcome are unknown. Neonatal prognosis, however, is excellent. Both neonatal and stillborn cases are probably the result of transplacental transmission of maternal antibodies. The connection between maternal antibody titer and fetal mortality is unknown. All stillborn cases reviewed died during the eight month of gestation. Immunosuppressive treatment probably affects fetal survival. In women with an active disease, who have had the disease, or who are monozygotic siblings to such patients, the possibility of the fetus developing the disease must be considered. CONCLUSIONS: It is essential that physicians be aware of the existence of childhood pemphigus vulgaris in order to make an early diagnosis and to avoid treatment delay. More childhood case reports are needed to obtain better information on optimal treatment, and authors should be encouraged to report the follow-up of their cases. PMID- 10517684 TI - 'Airborne' contact dermatitis due to Leica immersion oil. AB - BACKGROUND: Contact dermatitis has often been described in healthcare staff, resulting essentially from the use of natural rubber latex gloves, antiseptics, and especially aldehydes. This study reports an unusual cause of contact dermatitis in laboratory technicians. MATERIALS AND METHODS: Four patients working in the bacteriology departments of three different hospitals were seen for airborne contact dermatitis. All were patch tested for specific plastics and glues. RESULTS: For all patients, positive patch reactions were obtained with classic epoxy resins, such as diglycidylether of bisphenol, as well as with new types, such as diglycidylether of bisphenol F and an epoxyacrylate resin. CONCLUSIONS: Although phenols and ether handled by the laboratory technicians and an epoxy mastic applied during floor repair were initially suspected, an immersion oil used in light microscopy proved to be the real cause of the dermatitis. To our knowledge, these are the first reported cases due to this type of contact. PMID- 10517686 TI - The profile of atopic dermatitis in a tertiary dermatology outpatient clinic in Singapore. AB - BACKGROUND: Atopic dermatitis is a common, chronic, relapsing, pruritic, eczematous skin condition occurring in patients with a personal or family history of atopy. The aim of this study is to describe the profile of atopic dermatitis seen at a tertiary referral skin center in a tropical multiracial country. METHODS: A retrospective chart review was conducted of all the patients with atopic dermatitis seen during the first six months of 1994. RESULTS: There were 492 patients, age range from 1 month to 74 years, with an equal sex ratio. The prevalence was 2%. The onset of the disease occurred before the age of 10 years in 61.2% of patients. In 13.6% of patients, the onset was after the age of 21 years. Two hundred and fifty four patients (52%) had "pure" atopic dermatitis without concomitant respiratory allergies; 238 patients (48%) suffered from a "mixed" type, with 23% having allergic rhinitis, 12% having asthma, and 13% having both asthma and allergic rhinitis; 231 patients (47%) had at least one first-degree family member with atopy: atopic dermatitis (17%), asthma (15%), and allergic rhinitis (15%). Most of the patients, 416 (84.5%), had subacute dermatitis at presentation. Ichthyosis vulgaris was present in 38 patients (8%) and pityriasis alba in 13 patients (3%). The most common infective complication was bacterial infection (impetiginized dermatitis, folliculitis, cellulitis) present in 95 patients (19%), followed by viral infections (dermatitis herpeticum, viral warts, and molluscum contagiosum) in 17 patients (3%). Allergies were noted in 43 patients (9%). The most common was drug allergy (penicillin and cotrimoxazole) in 28 patients, followed by food allergy in 11 patients. Common aggravating factors reported included heat, sweating, stress, thick clothing, and grass intolerance. Most patients could be controlled with a fairly simple regimen of moisturizers, topical steroids, and antibiotics for acute flares. Short courses of systemic steroids were used in 78 patients (16%). Three patients were treated with phototherapy: two on combined UVA and UVB (UVAB) and one on oral psoralen photochemotherapy (PUVA). CONCLUSIONS: The pattern of atopic dermatitis in Singapore is similar to that reported in the Western literature, except for a lower prevalence and a significant proportion of adult onset atopic dermatitis. PMID- 10517688 TI - Angiosarcoma of the scalp. AB - An 82-year-old woman was seen at our Dermatology Department for a plaque on the right parietal scalp that had recently increased in size, and bled. The lesion had been present for 3 months. The patient had a previous diagnosis of chronic bronchitis, noninsulin-dependent diabetes mellitus, and hypertension, but no previous history of cancer. Physical examination revealed a 7 x 10 cm plaque, composed of a central necrotic and bleeding surface, surrounded by small purple red satellite nodules. A biopsy showed an ill-defined infiltrative intradermal mass with a pattern of hypercellular sheets of large cells alternating with areas of dilated, irregular, blood-filled channels, dissecting the collagen bundles. The endothelial cells lining these channels were plump and pleomorphic, surrounded by other spindle-shaped cells with pleomorphic and atypical nuclei. The diagnosis of angiosarcoma was made, and the patient was sent to an oncology center for further evaluation and treatment, where a computed tomography head scan was taken revealing no erosion of the skull. The patient refused surgery, so radiotherapy was proposed. One month later, she developed lymph node enlargement of the left anterior cervical nodes. A needle aspiration biopsy was consistent with sarcoma. Two weeks later, she was started on palliative radiotherapy: a programmed dose of 4500 cGy was proposed of which she only received 3000 cGy because of treatment withdrawal and loss to follow-up. During this time, she showed partial initial response, but despite treatment the disease relentlessly progressed, with hemorrhage and severe pain being the most striking features. PMID- 10517687 TI - Congenital self-healing histiocytosis (Hashimoto-Pritzker). AB - BACKGROUND: Congenital self-healing Langerhans cell histiocytosis (CSHLCH) is a rare condition, initially seen at birth or in the neonatal period, with generalized papules, vesicles, or nodules. Affected infants are otherwise well and the skin lesions tend to involute spontaneously within weeks to months. METHODS: Twelve patients with CSHLCH were seen from 1989 to 1998. RESULTS: Eight patients were girls and four were boys and all presented with lesions at birth which disappeared 1-3 months later. The lesions consisted of numerous brownish red papules, papulovesicles, crusts, and nodules distributed on the face, limbs, palms, and soles. Two patients had oral mucosal lesions, and one had ulcerated lesions that evolved leaving hypochromic macules. Light microscopy showed a histiocytic infiltrate in the papillary dermis with epidermotrophism. Two cases were studied by electron microscopy: the Langerhans cells showed Birbeck granules and laminated corpus in their cytoplasm. Immunomarking with S100 protein was performed in all 12 patients and was positive. CD1 was also tested in four cases and was positive. CONCLUSIONS: Because CSHLCH is a rare condition, we emphasize that, although it is usually a benign, self-limited entity, careful evaluation for systemic disease must be performed and long-term follow-up must be carried out to detect evidence of relapse or progression of the disease; this is essential when treating these patients. PMID- 10517690 TI - Vegetative pyoderma gangrenosum: an unusual presentation. PMID- 10517689 TI - Superficial cutaneous sporotrichosis in specific anergic patient. PMID- 10517691 TI - Henoch-Schonlein purpura induced by clarithromycin. PMID- 10517692 TI - Lichen sclerosus et atrophicus in sclerodermatous chronic graft-versus-host disease. PMID- 10517693 TI - Squamometry in seborrheic dermatitis. AB - BACKGROUND: Seborrheic dermatitis is a corticosteroid-responsive condition. Topical corticosteroids exhibit distinct activities according to the type and cellular site of action of the hormone and its vehicle. The latter influences not only drug penetration, but also the structure and function of the horny layer. OBJECTIVE: The present study compared the early response of seborrheic dermatitis to two topical corticosteroids using objective quantifications of the stratum corneum alterations. METHOD: Squamometry was used to supplement the clinical assessment of the therapeutic activity of 0.1% hydrocortisone butyrate in an oil in-water emulsion (Locoid Crelo) and 0.05% betamethasone 17-21 dipropionate lotion (Diprosone lotion). RESULTS: The hydrocortisone butyrate formulation had a faster onset of efficacy as documented by a greater improvement of lesions after a 1-week twice-daily treatment. Clearance or marked improvement was observed in the two treatment groups at completion of the 2-week trial. CONCLUSIONS: As the ranking of the clinical efficacy was not predicted by the intrinsic anti inflammatory potency of the steroids, it is suggested that the vehicle contributed itself to the global in vivo efficacy. The anti-inflammatory action of hydrocortisone butyrate appears to be synergistically implemented by the emollient effect of the specially designed oil-in-water emulsion. PMID- 10517694 TI - The fate of Germany's Jewish dermatologists in the period of National Socialism. PMID- 10517695 TI - Prostaglandin E2 modulates a non-inactivating potassium current in rat neurohypophyseal nerve terminals. AB - A non-inactivating voltage dependent K+ channel current was observed in neuro hypophyseal nerve terminals. This current was sensitive to inhibition by 4 aminopyridine and tetraethyl ammonium chloride, but was not sensitive to inhibition by alpha- or beta-dendrotoxin. Prostaglandin E2 (PGE2) modulated the voltage-dependent K+ channel, through a receptor-mediated process, as indicated by meclofenamate sensitivity, and this involved the activation of G protein(s), as indicated by sensitivity to guanosine-5'-O-(2-thiodiphosphate) (GDPfS). After short periods of incubation (e.g. 5 min), PGE2 increased the non-inactivating current. Following longer incubation periods with PGE2 (e.g. 20 min), the non inactivating current declined. Forskolin and the cyclic adenosine monophosphate (AMP) analogs 8-bromo- and dibutyryl cyclic AMP, and Sp-cyclic AMPs inhibited the current, but did not mimic the increase in current caused by PGE2. Also, the cyclic AMP antagonist Rp-cyclic AMPs did not block the increase in current induced by PGE2. These results indicate that activation of cyclic AMP-dependent protein kinase (PKA) is not involved in mediating the stimulatory actions of PGE2. These observations provide evidence that PGE2 may contribute to the regulation of hormone release from the posterior pituitary by modulating K+ channels. However, the post-receptor mechanisms of subcellular signal transduction underlying this effect remain unknown. PMID- 10517696 TI - Effect of neonatal handling on brain enkephalin-degrading peptidase activities. AB - Neonatal handling decreases neutral endopeptidase 24.11 activity in the amygdala. However, this procedure does not affect aminopeptidase activities in any of the brain areas studied. Neonatal handling has been one of the most commonly used strategies to study the plasticity of the nervous system. The crucial role of the opioids in the control of different aspects of behaviour and development has been well documented. Regarding this subject, the endogenous opioid system might mediate some of the effects induced by neonatal handling. In this work, we have studied the effects of neonatal handling on several enkephalin-degrading peptidases, including soluble and membrane-bound aminopeptidases (puromycin sensitive and -insensitive) and neutral endopeptidase 24.11 in different rat brain areas. Tyrosine aminopeptidase activities were measured fluorimetrically using tyrosine-beta-naphthylamide as substrate and puromycin as selective inhibitor of one of the membrane-enzymes. Dansyl-D-Ala-Gly-Phe(pNO2)-Gly was the fluorogenic substrate for neutral endopeptidase. The reduced neutral endopeptidase 24.11 activity in the amygdala of neonatal handled rats could reduce enkephalin catabolism in this area and it could be responsible for some of the effects induced by neonatal handling. PMID- 10517697 TI - Increased "peripheral-type" benzodiazepine receptor sites and mRNA in thalamus of thiamine-deficient rats. AB - "Peripheral-type" benzodiazepine receptors (PTBRs) are highly expressed on the outer mitochondrial membrane of several types of glial cells. In order to further elucidate the nature of the early glial cell changes in thiamine deficiency, PTBR sites and PTBR mRNA were measured in thalamus, a brain structure which is particularly vulnerable to thiamine deficiency, of thiamine-deficient rats at presymptomatic and symptomatic stages of deficiency. PTBR sites were measured using an in vitro binding technique and the selective radio ligand [3H]-PK11195. PTBR gene expression was measured by RT-PCR using oligonucleotide primers based upon the published sequence of the cloned rat PTBR. Microglial and astrocytic changes in thalamus due to thiamine deficiency were assessed using immunohistochemistry and antibodies to specific microglial (ED-1) and astrocytic (GFAP) proteins respectively. Significant increases of [3H]-PK11195 binding sites and concomitantly increased PTBR mRNA were observed in thalamus at the symptomatic stage of thiamine deficiency, coincident with severe neuronal cell loss and increased GFAP-immunolabelling (indicative of reactive gliosis). Positron Emission Tomography using 11C-PK11195 could provide a novel approach to the diagnosis and assessment of the extent of thalamic damage due to thiamine deficiency in humans with Wernicke's Encephalopathy. PMID- 10517698 TI - Expression of beta-amyloid precursor and Bcl-2 proto-oncogene proteins in rat retinas after intravitreal injection of aminoadipic acid. AB - In order to investigate the role of glia in relation to factors that affect the expression of beta-amyloid precursor protein (betaAPP) and B cell lymphoma oncogene protein (Bcl-2) in the central nervous tissue, the patterns of expression of betaAPP and Bcl-2 in developing and mature rat retinas were studied immunocytochemically after intravitreal injection of alpha-aminoadipic acid (alpha-AAA), a glutamate analogue and gliotoxin that is known to cause injury of retinal Muller glial cells. In normal developing retinas, betaAPP and Bcl-2 were expressed primarily but transiently in a small number of neurons in the ganglion cell layer during the first postnatal week. Immunoreactivity of betaAPP and Bcl-2 appeared in the endfeet and proximal part of the radial processes of Muller glial cells from the second postnatal week onwards. In rats that received intravitreal injection of alpha-AAA at birth, there was a loss of immunoreactivity to vimentin, and a delayed expressed on betaAPP or Bcl-2 in Muller glial cells until 3-5 weeks post-injection. Immunoreactive neurons were also observed in the inner retina especially in the ganglion cell layer from 5 to 35 days after injection. A significant reduction in numerical density of cells with large somata in the ganglion cell layer was observed in the neonatally injected retinas at P56, which was accompanied by an increased immunostaining in radial processes of Muller glial cells. In contrast, no detectable changes in the expression of betaAPP and Bcl-2 were observed in retina that received alpha-AAA as adults. These results indicate that the gliotoxin alpha-AAA has long lasting effects on the expression of betaAPP and Bcl-2 in Muller glial cells as well as neurons in the developing but not mature retinas. The loss of vimentin and delayed expression of betaAPP and Bcl-2 in developing Muller glial cells suggests that the metabolic integrity of Muller cells was temporarily compromised, which may have adverse effects on developing neurons that are vulnerable or dependent on trophic support from the Muller glial cells. PMID- 10517699 TI - Localization of alpha1B-adrenergic receptor in female rat brain regions involved in stress and neuroendocrine function. AB - Activation of alpha1-adrenergic receptors has been linked to the control of blood pressure, neuroendocrine secretion, reproductive behavior and mood. The present study describes the distribution of alpha1B-adrenergic receptor immunoreactivity in female rat brain regions involved in stress and neuroendocrine function. The pattern of immunolabeling seen resembles that obtained in previous in situ hybridization studies. Several hypothalamic areas that control pituitary function showed intense fiber and/or cell immunolabeling, including the paraventricular nucleus of the hypothalamus, the supraoptic nucleus, and the median eminence. Some regions such as the arcuate nucleus, the median eminence, and dorsal hypothalamus exhibit intense labeling of axonal varicosities, while other regions exhibit only perikarya immunolabeling. alpha1B-adrenergic receptor immunoreactivity was also observed in large pyramidal neurons of layer V of the cerebral cortex, the frontal cortex showing a particularly strong immunoreactivity. Virtually all thalamic regions were labeled, especially the lateral and ventral areas. In addition, labeled cells were present in hippocampus, the medial septum, the horizontal and vertical limbs of the diagonal band of Broca, and the caudate putamen. Finally, some midbrain and hindbrain regions important for motor function were immunoreactive. Because ligands specific for alpha1-adrenergic receptor subtypes are not available, the present immunocytochemical study not only addresses the subcellular and regional distribution of alpha1B-adrenergic receptors but may also provide clues about receptor subtype-specific function. PMID- 10517700 TI - HEK-293 cells expressing the human dopamine transporter are susceptible to low concentrations of 1-methyl-4-phenylpyridine (MPP+) via impairment of energy metabolism. AB - Selective dopaminergic neurotoxicity induced by 1-methyl-4-phenylpyridine (MPP+) is believed to be due to the transmembrane uptake by the dopamine transporter and subsequent inhibition of mitochondrial complex I and/or production of free radicals. However, little is known about the molecular sequence of intracellular events leading to cell death induced by low concentrations of MPP+. Here we stably express the human dopamine transporter (hDAT) in human embryonic kidney HEK-293 cells to correlate cytotoxicity and indices of cellular energy metabolism after exposure to low concentrations of MPP+. The permanent ektopic expression of hDAT in HEK-293 cells confers time and dose-dependent cytotoxicity at nanomolar concentrations of MPP+ with an IC50 value of 740 nM after 48 h. MPP+ initially induces a fast increase of cellular NADH content within the first 6 h, followed by a slow reduction of intracellular ATP (IC50 value of 690 nM after 48 h) as well as reduction of intracellular ATP/ADP ratio. These changes of cellular energy metabolism precede reduction of cell viability. The toxic effects of MPP+ are blocked by the hDAT inhibitor GBR12909 with EC50 values of 110 and 60 nM for cytotoxicity and ATP depletion, respectively. Antioxidants such as D-alpha tocopherol and ascorbic acid do not have significant protective effects against MPP+ toxicity. This study shows that HEK-293 cells expressing the hDAT gene are highly sensitive to MPP+ due to (i) transmembrane uptake of MPP+ by the dopamine transporter, (ii) cellular energy depletion, probably caused by inhibition of mitochondrial complex I activity and (iii) that the toxicity is independent from the presence of antioxidants. This cell system may serve as a screening system for endogenous and exogenous compounds with similar effects compared to MPP+ as well as protective agents. PMID- 10517701 TI - Lactate-induced inhibition of glucose catabolism in guinea pig cortical brain slices. AB - Extracellular lactate concentration rises following ischaemic stroke in both the infarcted area and in the surrounding ischaemic penumbra. We investigated the effect of lactate accumulation on glucose metabolism in cortical slices from guinea pigs initially by varying superfusion medium to tissue volumes. Stable intracellular K+ concentrations indicated that a decrease in media/ tissue volume did not impair viability of the tissue, but 13C NMR demonstrated that lactate accumulation in the superfusion medium reduced glucose oxidation with inhibition of glial metabolism via pyruvate carboxylase. The concentration of lactate which had accumulated when significant inhibition was observed was approximately 0.85 mM. In independent experiments we found that superfusion of brain slices with lactate at this concentration (even using a 'high-volume' of superfusion fluid) decreased oxygen consumption by 40 +/- 3%. K(-)-induced depolarisation partially reversed this effect. These results suggest that even low extracellular lactate concentrations may depress metabolic rates in inactive and poorly perfused brain tissue in vivo through inhibition of glial metabolism of glucose. PMID- 10517702 TI - A piece of my mind. Partnership for good dying. PMID- 10517704 TI - Mild cognitive impairment raises Alzheimer disease risk. PMID- 10517703 TI - Researchers urged to tell public how animal studies benefit human health. PMID- 10517705 TI - NEJM editor Jerome P. Kassirer, MD, loses post over "administrative issues". PMID- 10517706 TI - From the Centers for Disease Control and Prevention. Deaths among children aged < or = 5 years from farm machinery runovers--Iowa, Kentucky, and Wisconsin, 1995 1998, and United States, 1990-1995. PMID- 10517707 TI - From the Centers for Disease Control and Prevention. Firearm-associated deaths and hospitalizations--California, 1995-1996. PMID- 10517708 TI - Bartenders' pulmonary function after establishment of a smoke-free workplace. PMID- 10517709 TI - Fixed vs variable costs of hospital care. PMID- 10517710 TI - Fixed vs variable costs of hospital care. PMID- 10517711 TI - Access to essential drugs in poor countries. PMID- 10517712 TI - Access to essential drugs in poor countries. PMID- 10517714 TI - Physicians assessing the Internet: the PAI project. PMID- 10517713 TI - Myocardial infarction associated with ovarian hyperstimulation syndrome. PMID- 10517715 TI - Estimating time to conduct a meta-analysis from number of citations retrieved. PMID- 10517716 TI - Reduction of vertebral fracture risk in postmenopausal women with osteoporosis treated with raloxifene: results from a 3-year randomized clinical trial. Multiple Outcomes of Raloxifene Evaluation (MORE) Investigators. AB - CONTEXT: Raloxifene hydrochloride, a selective estrogen receptor modulator, prevents bone loss in postmenopausal women, but whether it reduces fracture risk in these women is not known. OBJECTIVE: To determine the effect of raloxifene therapy on risk of vertebral and nonvertebral fractures. DESIGN: The Multiple Outcomes of Raloxifene Evaluation (MORE) study, a multicenter, randomized, blinded, placebo-controlled trial. SETTING AND PARTICIPANTS: A total of 7705 women aged 31 to 80 years in 25 countries who had been postmenopausal for at least 2 years and who met World Health Organization criteria for having osteoporosis. The study began in 1994 and had up to 36 months of follow-up for primary efficacy measurements and nonserious adverse events and up to 40 months of follow-up for serious adverse events. INTERVENTIONS: Participants were randomized to 60 mg/d or 120 mg/d of raloxifene or to identically appearing placebo pills; in addition, all women received supplemental calcium and cholecalciferol. MAIN OUTCOME MEASURES: Incident vertebral fracture was determined radiographically at baseline and at scheduled 24- and 36-month visits. Nonvertebral fracture was ascertained by interview at 6-month-interim visits. Bone mineral density was determined annually by dual-energy x-ray absorptiometry. RESULTS: At 36 months of the evaluable radiographs in 6828 women, 503 (7.4%) had at least 1 new vertebral fracture, including 10.1% of women receiving placebo, 6.6% of those receiving 60 mg/d of raloxifene, and 5.4% of those receiving 120 mg/d of raloxifene. Risk of vertebral fracture was reduced in both study groups receiving raloxifene (for 60-mg/d group: relative risk [RR], 0.7; 95% confidence interval [CI], 0.5-0.8; for 120-mg/d group: RR, 0.5; 95% CI, 0.4-0.7). Frequency of vertebral fracture was reduced both in women who did and did not have prevalent fracture. Risk of nonvertebral fracture for raloxifene vs placebo did not differ significantly (RR, 0.9; 95% CI, 0.8-1.1 for both raloxifene groups combined). Compared with placebo, raloxifene increased bone mineral density in the femoral neck by 2.1 % (60 mg) and 2.4% (120 mg) and in the spine by 2.6% (60 mg) and 2.7% (120 mg) P<0.001 for all comparisons). Women receiving raloxifene had increased risk of venous thromboembolus vs placebo (RR, 3.1; 95% CI, 1.5 6.2). Raloxifene did not cause vaginal bleeding or breast pain and was associated with a lower incidence of breast cancer. CONCLUSIONS: In postmenopausal women with osteoporosis, raloxifene increases bone mineral density in the spine and femoral neck and reduces risk of vertebral fracture. PMID- 10517717 TI - Use of a low-literacy patient education tool to enhance pneumococcal vaccination rates. A randomized controlled trial. AB - CONTEXT: Pneumococcal immunization rates for elderly and high-risk patients are only one third to one half the target rate of 60% established by the US Public Health Service. Limited or marginal literacy, which affects nearly 100 million Americans, especially the elderly, may contribute to these low rates of immunization. OBJECTIVE: To determine whether the use of a simple, low-literacy educational tool enhances patient-physician dialogue about pneumococcal vaccination and increases rates of immunization. DESIGN: A randomized controlled trial conducted between May and June of 1998. SETTING: Ambulatory care clinic of a 900-bed public teaching hospital serving a predominantly indigent, low literate, African American, inner-city population. PARTICIPANTS: Of 433 patients who presented for routine primary care, had vaccine indications (age > or =65 years or chronic disease), and had not been previously vaccinated, 221 were randomly assigned to the intervention group and 212 to the control group. Of the total patient population (mean age, 63 years), 280 (64.7%) had less than a high school education, 401 (92.6%) were African American, and 300 (69.3%) were female. INTERVENTION: One-page, low-literacy (below fifth-grade level) educational handout encouraging patients to "ask your doctor about the pneumonia shot" vs a control group (1 -page, low-literacy educational handout conveying information about nutrition). MAIN OUTCOME MEASURES: Vaccination rates (documented by chart audit) of patients who received pneumococcal vaccination and rates of patients who self-reported having discussed vaccination with their physicians. RESULTS: Patients in the intervention group were 4 times more likely to have discussed the pneumococcal vaccine with their physicians than patients in the control group (87/221 [39.4%] vs 21/212 [9.9%]; relative risk [RR], 3.97 [95% confidence interval [CI], 2.71-5.83]), and were more than 5 times as likely to have received the pneumococcal vaccine than the control group (44/221 [19.9%] vs 8/212 [3.8%]; RR, 5.28 [95% CI, 2.80-9.93]). In a multivariate analysis controlling for race, sex, education, insurance status, age, level of physician training, health status, and vaccine indication, only assignment to the intervention group was statistically significantly related to the probability of being immunized or discussing the issue with their physicians (P<.001 for both trends). CONCLUSIONS: A simple, low-literacy educational tool increased pneumococcal vaccination rates and patient-physician discussions about the vaccine in an elderly, low-literate, indigent, minority population. PMID- 10517718 TI - Association between use of unconventional therapies and conventional medical services. AB - CONTEXT: The terms alternative and complementary medicine suggest 2 contradictory possibilities. Whether individuals use unconventional therapies as a substitute for or as an "add on" to conventional medical treatments is uncertain. OBJECTIVE: To determine the association between use of unconventional therapies and conventional medical care in a national sample. DESIGN, SETTING, AND PARTICIPANTS: The 1996 Medical Expenditure Panel Survey was distributed to a probability sample of the noninstitutionalized civilian US population. Of 24676 individuals responding (77.7% response rate), 16068 adults 18 years or older were included in the analysis. MAIN OUTCOME MEASURES: Visits to practitioners for unconventional therapies and conventional medical services, including number of inpatient, outpatient, and emergency department visits and use of 8 types of preventive medical services (blood pressure, cholesterol level, physical examination, influenza vaccination, prostate examination, breast examination, mammography, and Papanicolaou test). RESULTS: During 1996, an estimated 6.5% of the US population had visits for both unconventional therapies and conventional medical care; 1.8% used only unconventional services; 59.5% used only conventional care; and 32.2% used neither. Compared with those with only conventional visits, those who used both types of care had significantly more outpatient physician visits (7.9 vs 5.4; P<.001), and used more of all types of preventive services except mammography. These groups did not differ significantly in inpatient care, prescription drug use, or number of emergency department visits. Individuals in the top quartile of number of physician visits were more than twice as likely as those in the bottom quartile to have used unconventional therapies in the past year (14.5% vs 6.4%; P<.001). The association between unconventional treatments and physician visits remained after adjusting for potential confounders and across different types of unconventional treatment. CONCLUSIONS: In this sample, use of unconventional therapies was substantially lower than has been reported in previous national surveys, but was associated with increased use of physician services. From a health services perspective, practitioner-based unconventional therapies appear to serve more as a complement than an alternative to conventional medicine. PMID- 10517720 TI - Resolving discrepancies between a meta-analysis and a subsequent large controlled trial. AB - CONTEXT: A recent meta-analysis found calcium supplementation to be highly effective in preventing preeclampsia but a large National Institutes of Health trial (Calcium for Preeclampsia Prevention [CPEP]) found no risk reduction due to calcium in healthy nulliparous women. OBJECTIVES: To resolve discrepancies between the results of the meta-analysis and the CPEP trial and to assess the role of effect heterogeneity in the discrepancies. DATA SOURCES: Literature search of English-language articles published prior to July 10, 1997, the date of publication of the CPEP trial, using MEDLINE and by a manual search of bibliographies of published articles. STUDY SELECTION: Trials were included if they reported data on preeclampsia and calcium supplementation. Fourteen trials were systematically evaluated for differences in study design and patient populations. One trial was excluded because its results were reported after publication of the major CPEP results. DATA EXTRACTION: The sample size and number of subjects who developed preeclampsia in the calcium supplementation group vs a control group were recorded and analyzed on an intent-to-treat basis. Each author independently extracted the data. DATA SYNTHESIS: Substantial heterogeneity existed across trials (P = .001). After stratifying studies by the presence of a placebo-controlled group and by high-risk and low-risk populations, the conclusions of the meta-analysis of placebo-controlled trials enrolling a low risk population (relative risk, 0.79; 99% confidence interval, 0.44-1.42; P = .30) were compatible with the conclusions of the CPEP trial that calcium supplementation does not prevent preeclampsia in healthy nulliparous women. In contrast, the data implied a strong beneficial calcium effect (relative risk, 0.19; 99% confidence interval, 0.08-0.46; P = .001) in healthy high-risk subject populations. However, only 225 women were analyzed and because of inconsistent data, these results remain equivocal. CONCLUSIONS: Further studies are needed to establish the efficacy of calcium for preeclampsia prevention in healthy high risk populations. A single summary measure does not adequately describe the findings of a meta-analysis when the observed effects in individual studies differ substantially. In such settings the primary focus should be to identify and incorporate pertinent covariates that reduce heterogeneity and allow for optimum treatment strategies. PMID- 10517719 TI - Behavioral and neuroendocrine characteristics of the night-eating syndrome. AB - CONTEXT: Investigators first described the night-eating syndrome (NES), which consists of morning anorexia, evening hyperphagia, and insomnia, in 1955, but, to our knowledge, this syndrome has never been subjected to careful clinical study. OBJECTIVE: To characterize NES on the basis of behavioral characteristics and neuroendocrine data. DESIGN AND SETTING: A behavioral observational study was conducted between January 1996 and June 1997 in a weight and eating disorders program at the University of Pennsylvania. A neuroendocrine study was conducted from May through August 1997 at the Clinical Research Center of the University Hospital, Tromso, Norway. SUBJECTS: The behavioral study included 10 obese subjects who met criteria for NES and 10 matched control subjects. The neuroendocrine study included 12 night eaters and 21 control subjects. Behavioral study subjects were observed for 1 week on an outpatient basis, and neuroendocrine study subjects were observed during a 24-hour period in the hospital. MAIN OUTCOME MEASURES: The behavioral study measured timing of energy intake, mood level, and sleep disturbances. The neuroendocrine study measured circadian levels of plasma melatonin, leptin, and cortisol. RESULTS: In the behavioral study, compared with control subjects, night eaters had more eating episodes in the 24 hours (mean [SD], 9.3 [0.6] vs 4.2 [0.2]; P<.001) and consumed significantly more of their daily energy intake at night than did control subjects (56% vs 15%; P<.001). They averaged 3.6 (0.9) awakenings per night compared with 0.3 (0.3) by controls (P<.001). In night eaters, 52% of these awakenings were associated with food intake, with a mean intake per ingestion of 1134 (1197) kJ. None of the controls ate during their awakenings. In the neuroendocrine study, compared with control subjects, night eaters had attenuation of the nocturnal rise in plasma melatonin and leptin levels (P<.001 for both) and higher circadian levels of plasma cortisol (P = .001). CONCLUSION: A coherent pattern of behavioral and neuroendocrine characteristics was found in subjects with NES. PMID- 10517721 TI - Glucocorticoid-induced adrenal insufficiency. PMID- 10517722 TI - Consensus statement. Global burden of tuberculosis: estimated incidence, prevalence, and mortality by country. WHO Global Surveillance and Monitoring Project. AB - OBJECTIVE: To estimate the risk and prevalence of Mycobacterium tuberculosis (MTB) infection and tuberculosis (TB) incidence, prevalence, and mortality, including disease attributable to human immunodeficiency virus (HIV), for 212 countries in 1997. PARTICIPANTS: A panel of 86 TB experts and epidemiologists from more than 40 countries was chosen by the World Health Organization (WHO), with final agreement being reached between country experts and WHO staff. EVIDENCE: Incidence of TB and mortality in each country was determined by (1) case notification to the WHO, (2) annual risk of infection data from tuberculin surveys, and (3) data on prevalence of smear-positive pulmonary disease from prevalence surveys. Estimates derived from relatively poor data were strongly influenced by panel member opinion. Objective estimates were derived from high quality data collected recently by approved procedures. CONSENSUS PROCESS: Agreement was reached by (1) participants reviewing methods and data and making provisional estimates in closed workshops held at WHO's 6 regional offices, (2) principal authors refining estimates using standard methods and all available data, and (3) country experts reviewing and adjusting these estimates and reaching final agreement with WHO staff. CONCLUSIONS: In 1997, new cases of TB totaled an estimated 7.96 million (range, 6.3 million-11.1 million), including 3.52 million (2.8 million-4.9 million) cases (44%) of infectious pulmonary disease (smear-positive), and there were 16.2 million (12.1 million-22.5 million) existing cases of disease. An estimated 1.87 million (1.4 million-2.8 million) people died of TB and the global case fatality rate was 23% but exceeded 50% in some African countries with high HIV rates. Global prevalence of MTB infection was 32% (1.86 billion people). Eighty percent of all incident TB cases were found in 22 countries, with more than half the cases occurring in 5 Southeast Asian countries. Nine of 10 countries with the highest incidence rates per capita were in Africa. Prevalence of MTB/HIV coinfection worldwide was 0.18% and 640000 incident TB cases (8%) had HIV infection. The global burden of tuberculosis remains enormous, mainly because of poor control in Southeast Asia, sub-Saharan Africa, and eastern Europe, and because of high rates of M tuberculosis and HIV coinfection in some African countries. PMID- 10517723 TI - Therapy for fracture prevention. PMID- 10517724 TI - Nocturnal eating syndromes: to sleep, perchance to eat. PMID- 10517725 TI - Planning for a kidney transplant: is my doctor listening? PMID- 10517726 TI - JAMA Patient Page: tuberculosis. PMID- 10517727 TI - Pursuing progress in acute coronary syndromes. PMID- 10517728 TI - Warning: the short days of winter may be hazardous to your health. PMID- 10517729 TI - Enoxaparin prevents death and cardiac ischemic events in unstable angina/non-Q wave myocardial infarction. Results of the thrombolysis in myocardial infarction (TIMI) 11B trial. AB - BACKGROUND: Low-molecular-weight heparins are attractive alternatives to unfractionated heparin (UFH) for management of unstable angina/non-Q-wave myocardial infarction (UA/NQMI). METHODS AND RESULTS: Patients (n=3910) with UA/NQMI were randomized to intravenous UFH for >/=3 days followed by subcutaneous placebo injections or uninterrupted antithrombin therapy with enoxaparin during both the acute phase (initial 30 mg intravenous bolus followed by injections of 1.0 mg/kg every 12 hours) and outpatient phase (injections every 12 hours of 40 mg for patients weighing <65 kg and 60 mg for those weighing >/=65 kg). The primary end point (death, myocardial infarction, or urgent revascularization) occurred by 8 days in 14.5% of patients in the UFH group and 12.4% of patients in the enoxaparin group (OR 0.83; 95% CI 0.69 to 1.00; P=0. 048) and by 43 days in 19.7% of the UFH group and 17.3% of the enoxaparin group (OR 0.85; 95% CI 0.72 to 1.00; P=0.048). During the first 72 hours and also throughout the entire initial hospitalization, there was no difference in the rate of major hemorrhage in the treatment groups. During the outpatient phase, major hemorrhage occurred in 1.5% of the group treated with placebo and 2.9% of the group treated with enoxaparin (P=0.021). CONCLUSIONS: Enoxaparin is superior to UFH for reducing a composite of death and serious cardiac ischemic events during the acute management of UA/NQMI patients without causing a significant increase in the rate of major hemorrhage. No further relative decrease in events occurred with outpatient enoxaparin treatment, but there was an increase in the rate of major hemorrhage. PMID- 10517730 TI - Assessment of the treatment effect of enoxaparin for unstable angina/non-Q-wave myocardial infarction. TIMI 11B-ESSENCE meta-analysis. AB - BACKGROUND: Two phase III trials of enoxaparin for unstable angina/non-Q-wave myocardial infarction have shown it to be superior to unfractionated heparin for preventing a composite of death and cardiac ischemic events. A prospectively planned meta-analysis was performed to provide a more precise estimate of the effects of enoxaparin on multiple end points. METHODS AND RESULTS: Event rates for death, the composite end points of death/nonfatal myocardial infarction and death/nonfatal myocardial infarction/urgent revascularization, and major hemorrhage were extracted from the TIMI 11B and ESSENCE databases. Treatment effects at days 2, 8, 14, and 43 were expressed as the OR (and 95% CI) for enoxaparin versus unfractionated heparin. All heterogeneity tests for efficacy end points were negative, which suggests comparability of the findings in TIMI 11B and ESSENCE. Enoxaparin was associated with a 20% reduction in death and serious cardiac ischemic events that appeared within the first few days of treatment, and this benefit was sustained through 43 days. Enoxaparin's treatment benefit was not associated with an increase in major hemorrhage during the acute phase of therapy, but there was an increase in the rate of minor hemorrhage. CONCLUSIONS: The accumulated evidence, coupled with the simplicity of subcutaneous administration and elimination of the need for anticoagulation monitoring, indicates that enoxaparin should be considered as a replacement for unfractionated heparin as the antithrombin for the acute phase of management of patients with high-risk unstable angina/non-Q-wave myocardial infarction. PMID- 10517731 TI - Intracoronary thrombus and platelet glycoprotein IIb/IIIa receptor blockade with tirofiban in unstable angina or non-Q-wave myocardial infarction. Angiographic results from the PRISM-PLUS trial (Platelet receptor inhibition for ischemic syndrome management in patients limited by unstable signs and symptoms). PRISM PLUS Investigators. AB - BACKGROUND: The present study describes the effects of tirofiban, a nonpeptide platelet glycoprotein (GP) IIb/IIIa receptor blocker, on the characteristics of culprit lesions in patients with unstable angina (UA) or non-Q-wave myocardial infarction (NQWMI). METHODS AND RESULTS: Of 1915 patients enrolled in PRISM-PLUS, 1491 had a readable film obtained a median of 65 hours after randomization. A core laboratory examined the culprit lesions for intracoronary thrombus burden (primary end point) and for TIMI flow grade distribution and severity of the obstruction and of underlying coronary artery disease (secondary end points). The combination of tirofiban plus heparin compared with heparin alone significantly reduced the intracoronary thrombus burden of the culprit lesions (OR=0.77, P=0.022), improved the perfusion grade (OR=0.65, P=0.002), and decreased the severity of the obstruction (P=0.037), but it did not influence the severity of the underlying plaque. Persistence of a thrombus in 45% of patients was associated with a 2.4-fold increase in the odds of death at 30 days (P=0.005) and a 2-fold increase in the odds of myocardial infarction (P=0.002). CONCLUSIONS: The addition of tirofiban to heparin reduced the thrombus burden of the culprit lesion and improved distal perfusion in patients with UA or NQWMI, which supports the clinical benefit observed with the combination treatment. PMID- 10517732 TI - Plasma urokinase antigen and plasminogen activator inhibitor-1 antigen levels predict angiographic coronary restenosis. AB - BACKGROUND: The fibrinolytic system is intimately involved in several processes that contribute to restenosis, including clot dissolution, cell migration, and tissue remodeling. However, the role of the individual activators (urokinase [uPA] and tissue plasminogen [tPA] activators) and inhibitors (plasminogen activator inhibitor [PAI-1]) of the fibrinolytic system in maintaining patency after coronary artery angioplasty and stenting is unclear. METHODS AND RESULTS: We prospectively studied 159 patients with stable angina who underwent successful elective angioplasty (n=110) or stenting (n=49) of de novo native coronary artery lesions. Plasma samples were drawn at baseline (before angioplasty) and serially after angioplasty (immediately afterward and 6 hours, 24 hours, 3 days, 7 days, 1 month, 3 months, and 6 months afterward). Antigen and activity assays were performed for uPA, tPA, and PAI-1. Follow-up quantitative coronary angiography was performed in 92% of eligible patients. The overall angiographic restenosis rate (diameter stenosis >50%) was 31% (37% in PTCA patients, 17% in stented patients). At all time periods, including baseline, uPA antigen levels were significantly higher and PAI-1 antigen levels were significantly lower in patients with restenosis. Restenosis rates for patients in the upper tertile of baseline uPA antigen levels were 2-fold higher than for those in the lower 2 tertiles (46% versus 24% and 22%, respectively; P<0.004). In a stepwise regression multivariate analysis, obstruction diameter after the procedure and uPA antigen were significant predictors of follow-up diameter stenosis. CONCLUSIONS: Plasma uPA antigen levels and PAI-1 antigen levels identify patients at increased risk for restenosis after percutaneous coronary revascularization. PMID- 10517733 TI - Preserved endothelium-dependent vasodilation in coronary segments previously treated with balloon angioplasty and intracoronary irradiation. AB - BACKGROUND: Abnormal endothelium-dependent coronary vasomotion has been reported after balloon angioplasty (BA), as well as after intracoronary radiation. However, the long-term effect on coronary vasomotion is not known. The aim of this study was to evaluate the long-term vasomotion of coronary segments treated with BA and brachytherapy. METHODS AND RESULTS: Patients with single de novo lesions treated either with BA followed by intracoronary beta-irradiation (according to the Beta Energy Restenosis Trial-1.5) or with BA alone were eligible. Of these groups, those patients in stable condition who returned for 6 month angiographic follow-up formed the study population (n=19, irradiated group and n=11, control group). Endothelium-dependent coronary vasomotion was assessed by selective infusion of serial doses of acetylcholine (ACh) proximally to the treated area. Mean luminal diameter was calculated by quantitative coronary angiography both in the treated area and in distal segments. Endothelial dysfunction was defined as a vasoconstriction after the maximal dose of ACh (10( 6) mol/L). Seventeen irradiated segments (89.5%) demonstrated normal endothelial function. In contrast, 10 distal nonirradiated segments (53%) and 5 control segments (45%) demonstrated endothelium-dependent vasoconstriction (-19+/-17% and -9.0+/-5%, respectively). Mean percentage of change in mean luminal diameter after ACh was significantly higher in irradiated segments (P=0.01). CONCLUSIONS: Endothelium-dependent vasomotion of coronary segments treated with BA followed by beta-radiation is restored in the majority of stable patients at 6-month follow up. This functional response appeared to be better than those documented both in the distal segments and in segments treated with BA alone. PMID- 10517734 TI - When throughout the year is coronary death most likely to occur? A 12-year population-based analysis of more than 220 000 cases. AB - BACKGROUND: Previous studies have suggested that there is an increase in cardiac events in the morning. Fewer data relate cardiac events to months of the year and season. METHODS AND RESULTS: We analyzed all monthly death certificate data from Los Angeles County, California, for death caused by coronary artery disease from 1985 through 1996 (n=222 265). The mean number of deaths was highest in December at 1808 and January at 1925; the lowest rates were in June, July, August, and September at 1402, 1424, 1418, and 1371, respectively. December and January had significantly higher rates than would be expected from a uniform distribution of monthly deaths (P=0.00001). The percent of yearly coronary deaths was defined by the quadratic U-shaped equation [percent=13.1198-1.5238(month)+0. 0952(month(2)), where January=1, February=2, etc]. When monthly deaths were plotted by year, there was a decrease from 1985 through 1996. Monthly mortality correlated inversely with temperature. During the months with the highest frequency of death (December, January), however, there was an increase in deaths that peaked around the holiday season and then fell, which could not be explained solely on the basis of the daily temperature change. CONCLUSIONS: Even in the mild climate of Los Angeles County, there are seasonal variations in the development of coronary artery death, with approximately 33% more deaths occurring in December and January than in June through September. Although cooler temperatures may play a role, other factors such as overindulgence or the stress of the holidays might also contribute to excess deaths during these peak times. PMID- 10517735 TI - Diuretics shift circadian rhythm of blood pressure from nondipper to dipper in essential hypertension. AB - BACKGROUND: Recently, we found that sodium restriction shifted the circadian rhythm of blood pressure from nondipper to dipper in patients with the sodium sensitive essential hypertension. This study examined whether diuretics can transform the circadian rhythm of blood pressure from nondipper to dipper. METHODS AND RESULTS: We studied 21 patients with essential hypertension during both a baseline period and a period of treatment with hydrochlorothiazide (25 mg daily). The periods lasted 4 weeks each. Twenty-four hour ambulatory blood pressures were measured on the same day of the week at the end of the each period. In nondippers (n=11), but not in dippers (n=10), a significant interaction existed between diuretic therapy and nocturnal fall in systolic and diastolic blood pressure, which indicated that the degree of nocturnal blood pressure fall was affected by diuretic therapy. Nocturnal fall, which was diminished in nondippers, was restored by diuretic therapy with hydrochlorothiazide, indicating that the circadian rhythm of blood pressure shifted from nondipper to dipper patterns. CONCLUSIONS: The present study demonstrated that diuretics can restore nocturnal blood pressure decline in a manner similar to sodium restriction, which suggests that the kidneys and sodium metabolism may play important roles in the genesis of the circadian rhythm of blood pressure. Diuretic-based treatment may have an additional therapeutic advantage of reducing the risk for cardiovascular complications by transforming the circadian rhythm of blood pressure. PMID- 10517737 TI - Angiotensin II stimulates intercellular adhesion molecule-1 (ICAM-1) expression by human vascular endothelial cells and increases soluble ICAM-1 release in vivo. AB - BACKGROUND: We evaluated whether angiotensin II (Ang II) influenced intercellular adhesion molecule (ICAM)-1 expression by human vascular endothelial cells derived from umbilical cord veins (HUVECs) and plasma soluble ICAM-1 levels in vivo. METHODS AND RESULTS: Cultured HUVECs were incubated with Ang II (from 10(-9) to 10(-6) mol/L) with or without candesartan and PD12319 (inhibitors of Ang II AT(1) and AT(2) receptors, respectively) for various times up to 4 hours. Total RNA was then extracted from HUVECs, and Northern blots were probed with a 1.9-kb ICAM-1 cDNA fragment. HUVEC supernatants were used to assess soluble ICAM-1 release by ELISA. Northern blot analysis detected a strong increase of ICAM-1 mRNA after 2 hour incubation with Ang II. The response was inhibited by candesartan. Soluble ICAM-1 release by HUVECs also increased (P<0. 002) after 2-hour Ang II stimulation. In vivo, Ang II (at an initial rate of 1.0 ng. kg(-1). min(-1), to be increased each 30 minutes by 2.0 ng. kg(-1). min(-1) to the final rate of 7.0 ng. kg(-1). min(-1)) was infused in 8 normotensive and 12 essential hypertensive individuals. In the latter, Ang II was reinfused after 4 weeks on either placebo (n=3), losartan (50 mg UID, n=5), or atenolol (50 mg UID, n=4) treatment. Plasma soluble ICAM-1 levels increased after Ang II infusion in hypertensives and normotensives (P<0.0001 after 90 minutes). Losartan reduced baseline soluble ICAM 1 levels (P<0.05) and Ang II-related ICAM-1 increments. CONCLUSIONS: Ang II upregulates ICAM-1 expression by HUVECs and stimulates in vitro and in vivo soluble ICAM-1 release. AT(1) receptor blockade inhibits such endothelial effects of Ang II. PMID- 10517736 TI - Estrogen inhibits vascular smooth muscle cell-dependent adventitial fibroblast migration in vitro. AB - BACKGROUND: Mounting experimental evidence suggests that estrogen treatment protects against neointima formation in response to vascular injury in vivo. Previous studies have suggested that this process includes the activation and migration of adventitial fibroblasts. The present in vitro study was designed to establish a mechanism whereby estrogen attenuates migration of adventitial fibroblasts. METHDS AND RESULTS: Primary cultures of vascular smooth muscle cells (VSMCs) and adventitial fibroblasts were derived from female Sprague-Dawley rats. Reverse transcriptase-polymerase chain reaction and Western blotting were used to determine that expression of the estrogen receptor (ER) was restricted to early passage VSMCs. Migration of transduced (retrovirally mediated) fibroblasts was determined by counting the number of blue lacZ-expressing cells attached to Boyden-type chambers preconditioned under defined experimental conditions. Compared with growth medium alone, chambers treated with medium conditioned by VSMCs demonstrated a 2-fold increase in fibroblast migration, suggesting that VSMCs release soluble factor(s) competent to bind the Transwell membrane and promote fibroblast migration. In contrast, treatment of VSMCs with 17beta estradiol (10(-9) to 10(-7) mol/L) before preconditioning of the chamber induced a dose-dependent inhibition of fibroblast migration. Cotreatment of VSMCs with 17beta-estradiol and the ER antagonist ICI-182780 (10(-7) mol/L) blocked the inhibitory effect of estrogen on fibroblast migration. CONCLUSIONS: These observations suggest a novel mechanism of hormonal vasoprotection by which estrogen directly modulates VSMC expression of factor(s) controlling migration of adventitial fibroblasts via an ER-dependent mechanism. PMID- 10517738 TI - Myocardial uptake of (99m)Tc-N-NOET and (201)Tl during dobutamine infusion. Comparison with adenosine stress. AB - BACKGROUND: The myocardial uptake of (99m)Tc-sestamibi is attenuated by dobutamine stress, resulting in underestimation of ischemia. N-Ethyl-N-ethoxy dithiocarbamato-N-(99m)Tc ((99m)Tc-N-NOET) is a new (99m)Tc-labeled perfusion agent that is highly extracted by the myocardium by a mechanism different from that defined for (99m)Tc-sestamibi. We therefore hypothesized that (99m)Tc-N-NOET uptake would not be attenuated by dobutamine and that (99m)Tc-N-NOET uptake would be comparable to (201)Tl uptake during dobutamine stress. METHODS AND RESULTS: In 28 open-chest dogs, after placement of a stenosis in the left anterior descending coronary artery that reduced flow reserve by >50%, adenosine (300 microgram. kg( 1). min(-1); n=15) or dobutamine (2.5 to 30 microgram. kg(-1). min(-1); n=13) was infused. During adenosine stress, the stenotic-to-normal activity ratio for (99m)Tc-N-NOET was 0.55+/-0.05. The stenotic-to-normal flow ratio was 0.33+/-0.04 at the time of (99m)Tc-N-NOET injection. During dobutamine stress, the stenotic to-normal (99m)Tc-N-NOET activity ratio was 0.63+/-0.04, comparable to the (201)Tl activity ratio of 0.59+/-0.04. The stenotic-to-normal flow ratio was 0.47+/-0.04 at the time of (99m)Tc-N-NOET and (201)Tl injection. The relationship between (99m)Tc-N-NOET uptake and blood flow was comparable for adenosine and dobutamine stress, with no evidence of attenuation of (99m)Tc-N-NOET extraction by dobutamine. Conclusions-In the presence of coronary stenoses that reduced regional flow reserve, the myocardial uptake of (99m)Tc-N-NOET and (201)Tl are closely proportional to blood flow during both adenosine and dobutamine stress, suggesting that the adverse effect of dobutamine on (99m)Tc-sestamibi uptake is a tracer-specific phenomenon rather than a generalized effect. The clinical implication of this finding is that (99m)Tc-N-NOET might be preferable to (99m)Tc sestamibi when used with dobutamine stress for detection of coronary stenoses. PMID- 10517739 TI - Cellular basis for the Brugada syndrome and other mechanisms of arrhythmogenesis associated with ST-segment elevation. AB - BACKGROUND: The Brugada syndrome is characterized by marked ST-segment elevation in the right precordial ECG leads and is associated with a high incidence of sudden and unexpected arrhythmic death. Our study examines the cellular basis for this syndrome. METHODS AND RESULTS: Using arterially perfused wedges of canine right ventricle (RV), we simultaneously recorded transmembrane action potentials from 2 epicardial and 1 endocardial sites, together with unipolar electrograms and a transmural ECG. Loss of the action potential dome in epicardium but not endocardium after exposure to pinacidil (2 to 5 micromol/L), a K(+) channel opener, or the combination of a Na(+) channel blocker (flecainide, 7 micromol/L) and acetylcholine (ACh, 2 to 3 micromol/L) resulted in an abbreviation of epicardial response and a transmural dispersion of repolarization, which caused an ST-segment elevation in the ECG. ACh facilitated loss of the action potential dome, whereas isoproterenol (0.1 to 1 micromol/L) restored the epicardial dome, thus reducing or eliminating the ST-segment elevation. Heterogeneous loss of the dome caused a marked dispersion of repolarization within the epicardium and transmurally, thus giving rise to phase 2 reentrant extrasystole, which precipitated ventricular tachycardia (VT) and ventricular fibrillation (VF). Transient outward current (I(to)) block with 4-aminopyridine (1 to 2 mmol/L) or quinidine (5 micromol/L) restored the dome, normalized the ST segment, and prevented VT/VF. Conclusions-Depression or loss of the action potential dome in RV epicardium creates a transmural voltage gradient that may be responsible for the ST-segment elevation observed in the Brugada syndrome and other syndromes exhibiting similar ECG manifestations. Our results also demonstrate that extrasystolic activity due to phase 2 reentry can arise in the intact wall of the canine RV and serve as the trigger for VT/VF. Our data point to I(to) block (4 aminopyridine, quinidine) as an effective pharmacological treatment. PMID- 10517741 TI - Three-dimensional imaging of aortic aneurysm after balloon angioplasty for coarctation of the aorta. PMID- 10517740 TI - Ticlopidine and clopidogrel. AB - The thienopyridines ticlopidine and clopidogrel are inhibitors of platelet function in vivo. Their mode of action has not been defined, but it appears that they require conversion to as yet unidentified metabolites that are noncompetitive antagonists of the platelet ADP receptor. Inhibition of platelet aggregation with these compounds is delayed until 24 to 48 hours after administration. Maximum inhibition occurs after 3 to 5 days, and recovery is slow after drug withdrawal. Ticlopidine is effective in preventing cardiovascular events in cerebrovascular, cardiovascular, and peripheral vascular disease, with an efficacy that is similar to aspirin. However, its use is associated with significant and sometimes fatal adverse reactions, specifically neutropenia and bone marrow aplasia. Gastrointestinal side effects and skin rashes are common and result in discontinuation of therapy in up to 10% of patients. Clopidogrel is at least as effective as aspirin in preventing cardiovascular events in patients with a history of vascular disease. It appears to be safer than ticlopidine, although its efficacy in acute coronary syndromes or post-coronary-stent insertion has not been reported. Important outstanding issues are whether clopidogrel adds to the benefit of aspirin and whether the combination of these agents is safe. If so, this combination may become the standard for antithrombotic therapy in cardiovascular disease. PMID- 10517742 TI - Four-dimensional cardiac image by helical computed tomography. PMID- 10517743 TI - Mathematical treatment of autonomic oscillations. PMID- 10517744 TI - Mathematical treatment of autonomic oscillations. PMID- 10517745 TI - C-Reactive protein, serum amyloid A protein, and coronary events. PMID- 10517746 TI - C-Reactive protein, serum amyloid A protein, and coronary events. PMID- 10517747 TI - Cardiac neural changes before vasovagal syncope. PMID- 10517748 TI - Relaxation of diaphragm muscle. AB - Relaxation is the process by which, after contraction, the muscle actively returns to its initial conditions of length and load. In rhythmically active muscles such as diaphragm, relaxation is of physiological importance because diaphragm must return to a relatively constant resting position at the end of each contraction-relaxation cycle. Rapid and complete relaxation of the diaphragm is likely to play an important role in adaptation to changes in respiratory load and breathing frequency. Regulation of diaphragm relaxation at the molecular and cellular levels involves Ca(2+) removal from the myofilaments, active Ca(2+) pumping by the sarcoplasmic reticulum (SR), and decrease in the number of working cross bridges. The relative contribution of these mechanisms mainly depends on sarcomere length, muscle tension, and the intrinsic contractile function. Increased capacity of SR to take up Ca(2+) can arise from increased density of active SR pumping sites or in slow-twitch fibers from phosphorylation of phospholamban, whereas impaired coupling between ATP hydrolysis and Ca(2+) transport into the SR or intracellular acidosis reduces SR Ca(2+) pump activity. In experimental conditions of decreased contractile performance, slowed, enhanced, or unchanged relaxation rates have been reported in vitro. In vivo, a slowing in the rate of decline of the respiratory pressure is generally considered an early reliable index of respiratory muscle fatigue. Impaired relaxation rate may, in turn, favor mismatch between blood flow and metabolic demand, especially at high breathing frequencies. PMID- 10517749 TI - Growth hormone restores aged diaphragm myosin composition and performance after chronic undernutrition. AB - The effects of growth hormone (GH) on diaphragm muscle myosin heavy chain (MHC) composition and mechanical performance were investigated in Fischer 344 male rats aged to senescence (24.5 mo of age). Chronic undernutrition (UN), refeeding (RF), and RF+GH were compared with ad libitum feeding by using a model of UN that produced a 50% decrease in body weight over a 12-mo period. The effect of aging was assessed by comparing MHC composition of ad libitum-fed rats at 12 and 24.5 mo of age. At senescence, significant decreases in slow type I (-23%) and fast type IIA (-31%) MHC had occurred with aging. Conversely, UN over this aging period increased types I (32-73%) and IIA (22-23%) MHC and decreased fast types IIB (32-54%) and IIX (30-31%) MHC. RF and RF+GH reversed these shifts back toward control values. At senescence, maximal specific force, maximal velocity, and specific power capacity were not different across treatment groups. During repetitive isotonic contraction trials, the diaphragms of UN rats maintained power production over time (54% of initial power at 60 s), whereas the power production of diaphragms of ad libitum-fed rats fell to 0% (P < 0.05). In comparison with UN rats, the diaphragms of RF and RF+GH rats produced 23 (not significant) and 11% (P < 0.05) of initial power, respectively, suggesting that RF+GH treatment restored performance characteristics after UN. We conclude that RF+GH can reverse alterations in MHC composition and mechanical performance produced by chronic UN in the aged rat diaphragm. PMID- 10517750 TI - Temperature conditioning of nasal air: effects of vasoactive agents and involvement of nitric oxide. AB - Nitric oxide (NO) is released into nasal air, but its function is unknown. We hypothesized that nasal vascular tone and/or flow influences temperature conditioning of nasal air and that NO participates in this process. We measured nasal air temperature (via a thermocouple) and exhaled nasal NO release (by chemiluminescence) in five humans and examined the effects of an aerosolized vasoconstrictor (oxymetazoline), a vasodilator (papaverine), N(G)-nitro-L arginine methyl ester, an inhibitor of NO synthase, or saline (control). Compared with saline (which caused no changes in nasal air temperature or exhaled NO release), oxymetazoline (0.05%) reduced nasal air temperature and NO release (130.8 +/- 15.1 to 81.3 +/- 12.8 nl. min(-1). m(-2); P < 0.01). Papaverine (0.01 M) increased nasal air temperature and NO release (131.8 +/- 13.1 to 157.2 +/- 17.4 nl. min(-1). m(-2); P < 0.03). N(G)-nitro-L-arginine methyl ester reduced nasal air temperature and NO release (123.7 +/- 14.2 to 44.2 +/- 23.7 nl. min( 1). m(-2); P < 0.01). The results suggest that vascular tone and/or flow modulates temperature conditioning and that NO may participate in that function. PMID- 10517751 TI - Targeted deletion of the neutral endopeptidase gene alters ventilatory responses to acute hypoxia in mice. AB - Neutral endopeptidase (NEP) is one of the major endopeptidases responsible for the inactivation of substance P in the carotid body, a neurotransmitter shown to be important in the transduction of hypoxic stimuli. Ventilatory responses to acute hypoxia were measured by indirect plethysmography in unanesthetized, unrestrained wild-type mice and in mice in which the NEP gene was deleted (NEP -/ ). Ventilation was measured while the animals breathed room air: 12% O(2) in N(2) and 8% O(2) in N(2). Deletion of the NEP gene caused marked alterations in both the magnitude and composition of the hypoxic ventilatory response to both 8% O(2) in N(2) and 12% O(2) in N(2), compared with the wild-type mice (C57BL/6J) on the same genetic background as the NEP -/- mice. Treatment of C57BL/6J mice with thiorphan, a NEP inhibitor, resulted in a greater ventilatory response to 8% O(2) because of a significantly greater shortening of expiratory time. The results of these studies demonstrate that NEP plays an important role in modifying the expression of the ventilatory response to acute hypoxia. PMID- 10517752 TI - Ozone-induced hyperresponsiveness and blockade of M2 muscarinic receptors by eosinophil major basic protein. AB - Control of airway smooth muscle is provided by parasympathetic nerves that release acetylcholine onto M(3) muscarinic receptors. Acetylcholine release is limited by inhibitory M(2) muscarinic receptors. In antigen-challenged guinea pigs, hyperresponsiveness is due to blockade of neuronal M(2) receptors by eosinophil major basic protein (MBP). Because exposure of guinea pigs to ozone also causes M(2) dysfunction and airway hyperresponsiveness, the role of eosinophils in ozone-induced hyperresponsiveness was tested. Animals were exposed to filtered air or to 2 parts/million ozone for 4 h. Twenty-four hours later, the muscarinic agonist pilocarpine no longer inhibited vagally induced bronchoconstriction in ozone-exposed animals, indicating M(2) dysfunction. M(2) receptor function in ozone-exposed animals was protected by depletion of eosinophils with antibody to interleukin-5 and by pretreatment with antibody to guinea pig MBP. M(2) function was acutely restored by removal of MBP with heparin. Ozone-induced hyperreactivity was also prevented by antibody to MBP and was reversed by heparin. These data show that loss of neuronal M(2) receptor function after ozone is due to release of eosinophil MBP. PMID- 10517753 TI - Sepsis increases contraction-related generation of reactive oxygen species in the diaphragm. AB - Recent work indicates that free radicals mediate sepsis-induced diaphragmatic dysfunction. These previous experiments have not, however, established the source of the responsible free radical species. In theory, this phenomenon could be explained if one postulates that sepsis elicits an upregulation of contraction related free radical formation in muscle. The purpose of the present study was to test this hypothesis by examination of the effect of sepsis on contraction related free radical generation [i.e. , formation of reactive oxygen species (ROS)] by the diaphragm. Rats were killed 18 h after injection with either saline or endotoxin. In vitro hemidiaphragms were then prepared, and ROS generation during electrically induced contractions (20-Hz trains delivered for 10 min) was assessed by measurement of the conversion of hydroethidine to ethidium. ROS generation was negligible in noncontracting diaphragms from both saline- and endotoxin-treated groups (2.0 +/- 0. 6 and 2.8 +/- 1.0 ng ethidium/mg tissue, respectively), but it was marked in contracting diaphragms from saline-treated animals (19.0 +/- 2.8 ng/mg tissue) and even more pronounced (30.0 +/- 2.8 ng/mg tissue) in diaphragms from septic animals (P < 0.01). This enhanced free radical generation occurred despite the fact that the force-time integral (i.e., the area under the curve of force vs. time) for control diaphragms was higher than that for the septic group. In additional studies, in which we altered the stimulation paradigm in control muscles to achieve a force-time integral similar to that achieved in septic muscles, an even greater difference between control and septic muscle ROS formation was observed. These data indicate that ROS formation during contraction is markedly enhanced in diaphragms from endotoxin-treated septic animals. We speculate that ROS generated in this fashion plays a central role in producing sepsis-related skeletal muscle dysfunction. PMID- 10517754 TI - Theoretical and experimental intravascular gas embolism absorption dynamics. AB - Multifocal cerebrovascular gas embolism occurs frequently during cardiopulmonary bypass and is thought to cause postoperative neurological dysfunction in large numbers of patients. We developed a mathematical model to predict the absorption time of intravascular gas embolism, accounting for the bubble geometry observed in vivo. We modeled bubbles as cylinders with hemispherical end caps and solved the resulting governing gas transport equations numerically. We validated the model using data obtained from video-microscopy measurements of bubbles in the intact cremaster microcirculation of anesthetized male Wistar rats. The theoretical model with the use of in vivo geometry closely predicted actual absorption times for experimental intravascular gas embolisms and was more accurate than a model based on spherical shape. We computed absorption times for cerebrovascular gas embolism assuming a range of bubble geometries, initial volumes, and parameters relevant to brain blood flow. Results of the simulations demonstrated absorption time maxima and minima based on initial geometry, with several configurations taking as much as 50% longer to be absorbed than would a comparable spherical bubble. PMID- 10517755 TI - Acetylcholine chloride and renal hemodynamics during postnatal maturation in conscious lambs. AB - To test the hypothesis that acetylcholine-induced relaxation of the renal artery decreases with postnatal age, we measured parameters of renal hemodynamics before and for 35 s after aortic suprarenal injection of acetylcholine in conscious, chronically instrumented lambs aged approximately 1 wk (n = 5) and approximately 6 wk (n = 5). Acetylcholine was administered in one of five doses ranging from 0 to 10 mg/kg body wt; doses were administered randomly, in the same volume. There were significant age- and dose-dependent changes in renal vascular resistance after acetylcholine administration, such that the response was greater in 1-wk old lambs. After the highest dose tested, renal vascular resistance decreased by 13.6 +/- 7.3 (SD) mmHg. ml(-1). min. g kidney wt in 1-wk-old lambs and by 9.1 +/- 3.2 mmHg. ml(-1). min. g kidney wt in 6-wk-old lambs at 35 s. We also observed a transient renal vasoconstriction before the renal vasodilatation in 6-wk-old lambs but not in 1-wk-old animals. These data provide the first age- and dose dependent effects of exogenous administration of acetylcholine on renal hemodynamics during maturation in conscious animals. PMID- 10517756 TI - Alveolar epithelial fluid transport and the resolution of clinically severe hydrostatic pulmonary edema. AB - To characterize the rate and regulation of alveolar fluid clearance in the uninjured human lung, pulmonary edema fluid and plasma were sampled within the first 4 h after tracheal intubation in 65 mechanically ventilated patients with severe hydrostatic pulmonary edema. Alveolar fluid clearance was calculated from the change in pulmonary edema fluid protein concentration over time. Overall, 75% of patients had intact alveolar fluid clearance (>/=3%/h). Maximal alveolar fluid clearance (>/=14%/h) was present in 38% of patients, with a mean rate of 25 +/- 12%/h. Hemodynamic factors (including pulmonary arterial wedge pressure and left ventricular ejection fraction) and plasma epinephrine levels did not correlate with impaired or intact alveolar fluid clearance. Impaired alveolar fluid clearance was associated with a lower arterial pH and a higher Simplified Acute Physiology Score II. These factors may be markers of systemic hypoperfusion, which has been reported to impair alveolar fluid clearance by oxidant-mediated mechanisms. Finally, intact alveolar fluid clearance was associated with a greater improvement in oxygenation at 24 h along with a trend toward shorter duration of mechanical ventilation and an 18% lower hospital mortality. In summary, alveolar fluid clearance in humans may be rapid in the absence of alveolar epithelial injury. Catecholamine-independent factors are important in the regulation of alveolar fluid clearance in patients with severe hydrostatic pulmonary edema. PMID- 10517757 TI - Human angiotensin I-converting enzyme gene and endurance performance. AB - Human physical performance is strongly influenced by genetic factors. A variation in the structure of the human angiotensin I-converting enzyme (ACE) gene has been reported in which the insertion (I) variant is associated with lower ACE levels than the deletion (D) gene. We have previously reported that the I variant was associated with improved endurance performance in high-altitude mountaineers and British Army recruits. We now examine this genotype distribution in 91 British Olympic-standard runners (79 Caucasians). DNA was extracted from the buccal cells contained in 10 ml of saline mouthwash donated by the subjects, and the I and D variants of the ACE gene were identified by PCR amplification of the polymorphic region. There was an increasing frequency of the I allele with distance run [0.35, 0.53, and 0.62 for /=5,000 m (n = 34), respectively; P = 0.009 for linear trend]. Among 404 Olympic-standard athletes from 19 other mixed sporting disciplines (in which endurance performance was not necessarily a key factor), the I allele did not differ significantly from that found in control subjects: 0.50 vs. 0.49 (P = 0.526). These results support a positive association of the I allele with elite endurance performance. PMID- 10517758 TI - Modeling the influence of body size on V(O2) peak: effects of model choice and body composition. AB - This study examined the bivariate relationship between peak oxygen uptake (V(O2) peak); l/min) and body size in adult men (n = 1,314, age 17-66 yr), using both "simple" and "full" iterative nonlinear allometric models. The simple model was described by V(O2) peak = M(b) (or FFM(b)) exp(c SR-PA) exp(a + d age) epsilon (where M is body mass in kg; FFM is fat-free mass in kg; SR-PA is self-reported physical activity; epsilon is a multiplicative error term; and exp indicates natural antilogarithms). The full model was described by V(O2) peak = M(b) (or FFM(b)) exp(c SR-PA) exp(a + d age) + e (epsilon), where e is a permitted Y intercept term. The M exponent obtained from simple allometry was 0.65 [95% confidence interval (CI), 0.59-0.71], suggestive of a curvilinear relationship constrained to pass through the origin. This "zero Y-intercept" assumption was examined via the full allometric model, which revealed an M exponent of 1.00 (95% CI, 0.7-1.31), together with a positive Y-intercept term (e) of 1.13 (95% CI, 0.54-1.73). The FFM exponents were not significantly different from unity in either the simple or full allometric models. It appears that the curvilinearity of the simple allometric model (using total M) is fictitious and is due to the inappropriate forcing of the regression line through the origin. Utilizing FFM as the body-size variable revealed a linear relationship between body size and V(O2) peak, irrespective of model choice. We conclude that the population mass exponent for V(O2) peak is close to unity. PMID- 10517759 TI - Metabolic response in type I and type II muscle fibers during a 30-s cycle sprint in men and women. AB - The acute metabolic response to sprint exercise was studied in 20 male and 19 female students. We hypothesized that the reduction of muscle glycogen content during sprint exercise would be smaller in women than in men and that a possible gender difference in glycogen reduction would be higher in type II than in type I fibers. The exercise-induced increase in blood lactate concentration was 22% smaller in women than in men. A considerable reduction of ATP (50%), phosphocreatine (83%), and glycogen (35%) was found in type II muscle fibers, and it did not differ between the genders. A smaller reduction of ATP (17%) and phosphocreatine (78%) was found in type I fibers, and it did not differ between the genders. However, the exercise-induced reduction in glycogen content in type I fibers was 50% smaller in women than in men. The hypothesis was indeed partly confirmed: the exercise-induced glycogen reduction was attenuated in women compared with men, but the gender difference was in type I rather than in type II fibers. Fiber-type-specific and gender-related differences in the metabolic response to sprint exercise might have implications for the design of training programs for men and women. PMID- 10517760 TI - The adenosine A(1)-receptor antagonist 8-CPT reverses ethanol-induced inhibition of fetal breathing movements. AB - Administration of either ethanol or adenosine inhibits fetal breathing movements (FBM), eye movements, and low-voltage electrocortical activity (LV ECoG). The concentration of adenosine in ovine fetal cerebral extracellular fluid increases during ethanol-induced inhibition of FBM. The purpose of this study was to determine the effect of a selective adenosine A(1)-receptor antagonist, 8 cyclopentyltheophylline (8-CPT) on the incidence of FBM during ethanol exposure. After a 2-h control period, seven pregnant ewes received a 1-h intravenous infusion of ethanol (1 g/kg maternal body wt), followed 1 h later by a 2-h fetal intravenous infusion of either 8-CPT (3.78 +/- 0.08 microg. kg(-1). min(-1)) or vehicle. Ethanol reduced the incidence of FBM from 44.0 +/- 10.4 to 2.7 +/- 1.3% (P < 0.05) and 51.2 +/- 7.6 to 11.9 +/- 5.0% (P < 0.05) in fetuses destined to receive 8-CPT or vehicle, respectively. In the vehicle group, FBM remained suppressed for 7 h. In contrast, during the first hour of 8-CPT infusion, FBM returned to baseline (31 +/- 11%) and was not different from control throughout the rest of the experiment. Ethanol also decreased the incidence of both low voltage electrocortical activity and eye movements, but there were no differences in the incidences of these behavioral parameters between the 8-CPT and vehicle groups throughout the experiment. These data are consistent with the hypothesis that adenosine, acting via A(1) receptors, may play a role in the mechanism of ethanol-induced inhibition of FBM. PMID- 10517761 TI - Impaired load dependence of diaphragm relaxation during congestive heart failure in the rabbit. AB - The load dependence (LD) of relaxation was studied in the diaphragm of rabbits with congestive heart failure (CHF). CHF (n = 15) was induced by combined chronic volume and pressure overload. Aortic insufficiency was induced by forcing a catheter through the aortic sigmoid valves, followed 3 wk later by abdominal aortic stenosis. Six weeks after the first intervention, animals developed CHF. Sham-operated animals served as controls (C; n = 12). Diaphragm mechanics were studied in vitro on isolated strips, at 22 degrees C, in isotonic and isometric loading conditions. Contractility was lower in the CHF group, as reflected by lower total tension: 1.11 +/- 0.10 in CHF vs. 2.38 +/- 0.15 N/cm(2) in C in twitch (P < 0.001) and 2.46 +/- 0.22 in CHF vs. 4.90 +/- 0.25 N. cm(-2) in C in tetanus (P < 0.001). The index LD was used to quantify the load dependence of relaxation: LD is <1 in load-dependent muscles and tends toward 1 in load independent muscles. LD was significantly higher in CHF than in C rabbits, in both twitch (0.99 +/- 0.01 vs. 0.75 +/- 0.03; P < 0. 001) and tetanus (0.95 +/- 0.02 vs. 0.84 +/- 0.02; P < 0.001). In the CHF rabbits' diaphragm, the fall in total tension was linearly related to the fall in load dependence of relaxation. The decrease in load dependence of relaxation in CHF animals suggests sarcoplasmic reticulum abnormalities. Impairment of the sarcoplasmic reticulum may also partly account for the decrease in contractile performance of diaphragm in CHF animals. PMID- 10517762 TI - Influence of core temperature on autoresuscitation during repeated exposure to hypoxia in normal rat pups. AB - Failure to autoresuscitate by hypoxic gasping during prolonged sleep apnea has been suggested to play a role in sudden infant death. Furthermore, thermal stress brought about by a contribution of infection, overwrapping, or excessive environmental heating has been shown to be associated with an increased risk of sudden infant death, particularly in prone sleeping infants. The present experiments were carried out on newborn rat pups to investigate the influence of "forced" changes in core temperature on their time to last gasp during a single hypoxic exposure and on their ability to autoresuscitate during repeated exposure to hypoxia. On day 5 or 6 postpartum the pups were placed in a temperature controlled chamber regulated to 33, 35, 37, 39, or 41 degrees C and exposed to a single period of hypoxia (97% N(2)-3% CO(2)) and their time to last gasp was determined, or they were exposed repeatedly to hypoxia and their ability to autoresuscitate from primary apnea was determined. Increases in core temperature brought about by changes in ambient temperature from 33 to 41 degrees C decreased the time to last gasp after a single hypoxic exposure and decreased the number of successful autoresuscitations after repeated hypoxic exposures. Thus our data support the hypothesis that forced changes in core temperature brought about by changes in ambient temperature influence protective responses in newborns that may prevent death during hypoxia, as may occur during single or repeated episodes of prolonged sleep apnea. PMID- 10517763 TI - Intravascular macrophage depletion attenuates endotoxin lung injury in anesthetized sheep. AB - We recently showed that we can selectively and safely deplete most (average 85%) of the pulmonary intravascular macrophages in sheep by intravenously infusing liposomes containing dichloromethylene bisphosphonate. After a 1-h stable baseline, we made a 6-h comparison after a 30-min intravenous endotoxin infusion (1 microg/kg) between six anesthetized control lambs and six anesthetized lambs in which the intravascular macrophages had been depleted 24 h previously. Three of the control lambs had been macrophage depleted and allowed to recover their intravascular macrophage population for >/=2 wk. After depletion, both the early and late pulmonary arterial pressure rises were dramatically attenuated. Our main interest, however, was in the acute lung microvascular injury response. The early and late rises in lung lymph flow and the increase in lung lymph protein clearance (lymph flow x lymph-to-plasma protein concentration ratio) were >90% attenuated. We conclude the pulmonary intravascular macrophages are responsible for most of the endotoxin-induced pulmonary hypertension and increased lung microvascular leakiness in sheep, although the unavoidable injury of other intravascular macrophages by the depletion regime may also contribute something. PMID- 10517764 TI - Endurance exercise causes interaction among stress hormones, cytokines, neutrophil dynamics, and muscle damage. AB - We analyzed adaptation mechanisms regulating systemic inflammatory response of the stressed body by using an experimental challenge of repeated exercise bouts and accompanying muscle inflammation. Eight untrained men bicycled at 90 W for 90 min, 3 days in a row. Exercise induced peripheral neutrophilia with a leftward shift of neutrophil nucleus and neutrophil priming for oxidative activity determined by luminol-dependent chemiluminescence. Plasma growth hormone and interleukin-6 rose significantly after exercise and were closely correlated with the neutrophil responses. Serum creatine kinase and myoglobin levels as muscle damage markers rose after exercise in "delayed onset" and were closely correlated with the preceding neutrophil responses. These exercise-induced responses were strongest on day 1, but the magnitude gradually decreased with progressive daily exercise. In contrast, the magnitude of catecholamine responses to exercise sessions gradually rose, possibly suppressing neutrophil oxidative responses. These results indicate that stress-induced systemic release of bioactive substances may determine neutrophil mobilization and functional status, which then may affect local tissue damage of susceptible organs. PMID- 10517765 TI - Dissociation of peak vascular conductance and V(O2) max among highly trained athletes. AB - Previously, a strong relationship has been found between whole body maximal aerobic power (VO(2 max)) and peak vascular conductance in the calf muscle (J. L. Reading, J. M. Goodman, M. J. Plyley, J. S. Floras, P. P. Liu, P. R. McLaughlin, and R. J. Shephard. J. Appl. Physiol. 74: 567-573, 1993; P. G. Snell, W. H. Martin, J. C. Buckley, and C. G. Blomqvist. J. Appl. Physiol. 62: 606-610, 1987), suggesting a matching between maximal exercise capacity and peripheral vasodilatory reserve across a broad range of aerobic power. In contrast, long term training could alter this relationship because of the unique demands for muscle blood flow and cardiac output imposed by different types of training. In particular, the high local blood flows but relatively low cardiac output demand imposed by the type of resistance training used by bodybuilders may cause a relatively greater development in peripheral vascular reserve than in aerobic power. To examine this possibility, we studied the relationship between treadmill VO(2 max) and vascular conductance in the calf by using strain-gauge plethysmography after maximal ischemic plantar flexion exercise in 8 healthy sedentary subjects (HS) and 28 athletes. The athletes were further divided into three groups: 10 elite middle-distance runners (ER), 11 power athletes (PA), and 7 bodybuilders (BB). We found that both BB and ER deviate from the previously demonstrated relationship between VO(2 max) and vascular conductance. Specifically, for a given vascular conductance, BB had a lower VO(2 max), whereas ER had a higher VO(2 max) than did HS and PA. We conclude that the relationship between peak vascular conductance and aerobic power is altered in BB and ER because of training-specific effects on central vs. peripheral cardiovascular adaptation to local skeletal muscle metabolic demand. PMID- 10517766 TI - Effects of salbutamol and Ro-20-1724 on airway and parenchymal mechanics in rats. AB - We investigated the effects of a selective beta(2)-agonist, salbutamol, and of phosphodiesterase type 4 inhibition with 4-(3-butoxy-4-methoxy benzyl)-2 imidazolidinone (Ro-20-1724) on the airway and parenchymal mechanics during steady-state constriction induced by MCh administered as an aerosol or intravenously (iv). The wave-tube technique was used to measure the lung input impedance (ZL) between 0.5 and 20 Hz in 31 anesthetized, paralyzed, open-chest adult Brown Norway rats. To separate the airway and parenchymal responses, a model containing an airway resistance (Raw) and inertance (Iaw), and a parenchymal damping (G) and elastance (H), was fitted to ZL spectra under control conditions, during steady-state constriction, and after either salbutamol or Ro 20-1724 delivery. In the Brown Norway rat, the response to iv MCh infusion was seen in Raw and G, whereas continuous aerosolized MCh challenge produced increases in G and H only. Both salbutamol, administered either as an aerosol or iv, and Ro-20-1724 significantly reversed the increases in Raw and G when MCh was administered iv. During the MCh aerosol challenge, Ro-20-1724 significantly reversed the increases in G and H, whereas salbutamol had no effect. These results suggest that, after MCh-induced changes in lung function, salbutamol increases the airway caliber. Ro-20-1724 is effective in reversing the airway narrowings, and it may also decrease the parenchymal constriction. PMID- 10517767 TI - Effect of supplementation with a cysteine donor on muscular performance. AB - Oxidative stress contributes to muscular fatigue. GSH is the major intracellular antioxidant, the biosynthesis of which is dependent on cysteine availability. We hypothesized that supplementation with a whey-based cysteine donor [Immunocal (HMS90)] designed to augment intracellular GSH would enhance performance. Twenty healthy young adults (10 men, 10 women) were studied presupplementation and 3 mo postsupplementation with either Immunocal (20 g/day) or casein placebo. Muscular performance was assessed by whole leg isokinetic cycle testing, measuring peak power and 30-s work capacity. Lymphocyte GSH was used as a marker of tissue GSH. There were no baseline differences (age, ht, wt, %ideal wt, peak power, 30-s work capacity). Follow-up data on 18 subjects (9 Immunocal, 9 placebo) were analyzed. Both peak power [13 +/- 3.5 (SE) %, P < 0.02] and 30-s work capacity (13 +/- 3.7%, P < 0.03) increased significantly in the Immunocal group, with no change (2 +/- 9.0 and 1 +/- 9.3%) in the placebo group. Lymphocyte GSH also increased significantly in the Immunocal group (35.5 +/- 11.04%, P < 0.02), with no change in the placebo group (-0.9 +/- 9.6%). This is the first study to demonstrate that prolonged supplementation with a product designed to augment antioxidant defenses resulted in improved volitional performance. PMID- 10517768 TI - A computer simulation of free-range exercise in the laboratory. AB - We present a technique for simulating dynamic field (free-range) exercise, using a novel computer-controlled cycle ergometer. This modified cycle ergometer takes into account the effect of friction and aerodynamic drag forces on a 70-kg cyclist in a racing position. It also affords the ability to select different gear ratios. We have used this technique to simulate a known competition cycle route in Cape Town, South Africa. In an attempt to analyze the input stimulus, in this case the generated power output of each cyclist, eight subjects cycled for 40 min at a self-selected, comfortable pace on the first part of the simulated route. Our results indicate that this exercise input excites the musculocardiorespiratory system over a wide range of power outputs, both in terms of amplitude and frequency. This stimulus profile thereby complies with the fundamental requirement for nonlinear (physiological) systems analysis and identification. Through a computer simulation, we have devised a laboratory exercise protocol that not only is physiologically real but also overcomes the artificiality of most traditional laboratory exercise protocols. PMID- 10517769 TI - Constriction to hypoxia-reoxygenation in isolated mouse coronary arteries: role of endothelium and superoxide. AB - The aim of the present study was to determine the role of endothelium and superoxide in the responses of isolated mouse coronary arteries to hypoxia reoxygenation. Isolated mouse coronary artery was cannulated, pressurized at 60 mmHg, and constantly superfused with recirculating Krebs-Ringer bicarbonate solution for continuous measurement of intraluminal diameter (ID) by video microscopy. Under a no-flow condition, hypoxia (0% O(2), 30 min) caused vasoconstriction. Reoxygenation caused a further vasoconstriction (ID change from 111.4 +/- 11.1 to 91 +/- 16.5 microm) that was significantly reduced by removal of endothelium (ID change from 105.4 +/- 27 to 109.9 +/- 23.4 microm). Cu/Zn superoxide dismutase (150 U/ml) did not alter the hypoxic vasoconstriction but abolished the reoxygenation-caused endothelium-dependent vasoconstriction. Hypoxia-reoxygenation markedly enhanced the generation of superoxide that was significantly reduced by either removing the endothelium or treated these endothelium-intact vessels with superoxide dismutase. These results suggest that, in isolated mouse coronary arteries, hypoxia causes vasoconstriction that is independent of endothelium, whereas reoxygenation causes vasoconstriction that is mediated by enhanced generation of superoxide from endothelium. PMID- 10517770 TI - Static respiratory muscle work during immersion with positive and negative respiratory loading. AB - Upright immersion imposes a pressure imbalance across the thorax. This study examined the effects of air-delivery pressure on inspiratory muscle work during upright immersion. Eight subjects performed respiratory pressure-volume relaxation maneuvers while seated in air (control) and during immersion. Hydrostatic, respiratory elastic (lung and chest wall), and resultant static respiratory muscle work components were computed. During immersion, the effects of four air-delivery pressures were evaluated: mouth pressure (uncompensated); the pressure at the lung centroid (PL,c); and at PL,c +/-0.98 kPa. When breathing at pressures less than the PL,c, subjects generally defended an expiratory reserve volume (ERV) greater than the immersed relaxation volume, minus residual volume, resulting in additional inspiratory muscle work. The resultant static inspiratory muscle work, computed over a 1-liter tidal volume above the ERV, increased from 0.23 J. l(-1), when subjects were breathing at PL,c, to 0.83 J. l( 1) at PL,c -0.98 kPa (P < 0.05), and to 1.79 J. l(-1) at mouth pressure (P < 0.05). Under the control state, and during the above experimental conditions, static expiratory work was minimal. When breathing at PL,c +0.98 kPa, subjects adopted an ERV less than the immersed relaxation volume, minus residual volume, resulting in 0.36 J. l(-1) of expiratory muscle work. Thus static inspiratory muscle work varied with respiratory loading, whereas PL,c air supply minimized this work during upright immersion, restoring lung-tissue, chest-wall, and static muscle work to levels obtained in the control state. PMID- 10517771 TI - Presinusoidal vessels predominantly contract in response to norepinephrine, histamine, and KCl in rabbit liver. AB - In rabbit livers, it is not well known which segments of the hepatic vasculature are predominantly contracted by various vasoconstrictors. We determined effects of histamine, norepinephrine, and KCl on hepatic vascular resistance distribution in isolated rabbit livers perfused via the portal vein with 5% albumin-Krebs solution at a constant flow rate. Hepatic capillary pressure was measured by double vascular occlusion pressure (Pdo) and was used to determine portal (Rpv) and hepatic venous (Rhv) resistances. A bolus injection of either histamine or norepinephrine dose-dependently increased portal venous pressure but not Pdo, resulting in a dose-dependent increase in Rpv and no changes in Rhv. KCl (50 mM), when injected in anterogradely perfused livers, contracted the presinusoidal vessels selectively with liver weight loss. Although KCl significantly increased Rhv in retrogradely perfused livers, the increase in Rpv by 400% of baseline predominated over the increase in Rhv by 85% of baseline. In the retrogradely perfused livers, KCl produced an initial liver weight loss followed by a profound weight gain. We conclude that histamine and norepinephrine selectively contract the presinusoidal vessels. The results on KCl effects suggest that this selective presinusoidal constriction might be possibly due to predominant distribution of functionally active vascular smooth muscle in the presinusoidal vessels rather than the hepatic vein in rabbit livers. PMID- 10517772 TI - Effect of training status on fuel selection during submaximal exercise with glucose ingestion. AB - In this study, an oral glucose load was enriched with a [U-(13)C]glucose tracer to determine differences in substrate utilization between endurance-trained (T) and untrained (UT) subjects during submaximal exercise at the same relative and absolute workload when glucose is ingested. Six highly trained cyclists/triathletes [maximal workload (Wmax), 400 +/- 9 W] and seven UT subjects (Wmax, 296 +/- 8 W) were studied during 120 min of cycling exercise at 50% Wmax ( approximately 55% maximal O(2) consumption). The T subjects performed a second trial at the mean workload of the UT group (148 +/- 4 W). Before exercise, 8.0 ml/kg of a (13)C-enriched glucose solution (80 g/l) was ingested. During exercise, boluses of 2.0 ml/kg of the same solution were administered every 15 min. Measurements were made in the 90- to 120-min period when a steady state was present in breath (13)CO(2) and plasma glucose (13)C enrichment. Energy expenditure was higher in T than in UT subjects (58 vs. 47 kJ/min, respectively; P < 0.001) at the same relative intensity. This was completely accounted for by an increased fat oxidation (0.57 vs. 0.40 g/min; P < 0.01). At the same absolute intensity, fat oxidation contributed more to energy expenditure in the T compared with the UT group (44 vs. 33%, respectively; P < 0.01). The reduction in carbohydrate oxidation in the T group was explained by a diminished oxidation rate of muscle glycogen (indirectly assessed by using tracer methodology at 0.72 +/- 0.1 and 1.03 +/- 0.1 g/min, respectively; P < 0.01) and liver-derived glucose (0.15 +/- 0.03 and 0.22 +/- 0.02 g/min, respectively; P < 0.05). Exogenous glucose oxidation rates were similar during all trials (+/-0.70 g/min). PMID- 10517773 TI - Capillary filtration coefficient, vascular resistance, and compliance in isolated mouse lungs. AB - Although many recently produced transgenic mice possess gene alterations affecting pulmonary vascular function, there are few baseline measurements of vascular resistance and permeability. Therefore, we excised the lungs of C57/BL6 mice and perfused them with 5% bovine serum albumin in RPMI-1640 culture medium at a nominal flow of 0.5 ml/min and ventilated them with 20% O(2)-5% CO(2)-75% N(2). The capillary filtration coefficient, a sensitive measurement of hydraulic conductivity, was unchanged over 2 h (0.33 +/- 0.03 ml. min(-1). cmH(2)O(-1). 100 g(-1)) in a control group ventilated with low peak inflation pressures (PIP) but increased 4. 3-fold after high PIP injury. Baseline pulmonary vascular resistance was 6.1 +/- 0.4 cmH(2)O. ml(-1). min. 100 g(-1) and was distributed 34% in large arteries, 18% in small arteries, 14% in small veins, and 34% in large veins on the basis of vascular occlusion pressures. Baseline vascular compliance was 5.4 +/- 0.3 ml. cmH(2)O(-1). 100 g(-1) and decreased significantly with increased vascular pressures. Baseline pulmonary vascular resistance was inversely related to both perfusate flow and microvascular pressure and increased to 202% of baseline after infusion of 10(-4) M phenylephrine due to constriction of large arterial and venous segments. Thus isolated mouse lung vascular permeability, vascular resistance, and the longitudinal distribution of vascular resistance are similar to those in other species and respond in a predictable manner to microvascular injury and a vasoconstrictor agent. PMID- 10517774 TI - Hyperbaric bradycardia and hypoventilation in exercising men: effects of ambient pressure and breathing gas. AB - We sought to determine whether hydrostatic pressure contributed to bradycardia and hypoventilation in hyperbaria. Eight men were studied during exercise at 50, 150, and 250 W while breathing 1) air at 1 bar, 2) helium-oxygen (He-O(2)) at 5.5 bar, 3) sulfur hexafluoride-oxygen (SF(6)-O(2)) at 1.3 bar, and 4) nitrogen oxygen (N(2)-O(2)) at 5.5 bar. Gas densities were pairwise identical in 1) and 2), and 3) and 4), respectively. Increased hydrostatic pressure to 5.5 bar resulted in a modest but significant relative bradycardia on the order of 6 beats/min, in both the absence [1) vs. 2), P = 0. 0015] and presence [3) vs. 4), P = 0.029] of gases that are both denser than normal and mildly narcotic. In contrast, ventilatory responses appeared not to be influenced by hydrostatic pressure. Also, the combined exposure to increased gas density and mild-to moderate inert gas narcosis at a given hydrostatic pressure [1) vs. 3), 2) vs. 4)] caused bradycardia (P = 0.032 and 0.061, respectively) of similar magnitude as 5.5-bar hydrostatic pressure. At the same time there was relative hypoventilation at the two higher workloads. We conclude that heart rate control, but not ventilatory control, is sensitive to relatively small increases in hydrostatic pressure. PMID- 10517775 TI - Mechanical contribution of expiratory muscles to pressure generation during spinal cord stimulation. AB - Lower thoracic spinal cord stimulation (SCS) results in the generation of large positive airway pressures (Paw) and may be a useful method of restoring cough in patients with spinal cord injury. The purpose of the present study was to assess the mechanical contribution of individual respiratory muscles to pressure generation during SCS. In anesthetized dogs, SCS was applied at different spinal cord levels by using a 15-lead multicontact electrode before and after sequential ablation of the external and internal obliques, transversus abdominis (TA), rectus abdominis, and internal intercostal muscles. Paw was monitored after tracheal occlusion. SCS at the T(9) spinal cord level resulted in maximal changes in Paw (60 +/- 3 cmH(2)O). Section of the oblique muscles resulted in a fall in Paw to 29 +/- 2 cmH(2)O. After subsequent section of the rectus abdominis and TA, Paw fell to 25 +/- 2 and 12 +/- 1 cmH(2)O respectively. There was a small remaining Paw (4 +/- 1 cmH(2)O) after section of the internal intercostal nerves. Stimulation with a two-electrode lead system (T(9) + T(13)) resulted in significantly greater pressure generation compared with a single-electrode lead due to increased contributions from the obliques and transversus muscles. In a separate group of animals, Paw generation was monitored after section of the abdominal muscles and again after section of the external intercostal and levator costae muscles. These studies demonstrated that inspiratory intercostal muscle stimulation resulted in only a small opposing inspiratory action (90%, these muscles were subjected to an in situ indirect stimulation protocol, including a series of isolated twitch and tetanic contractions preceding a 3-min fatigue regimen (100-ms trains at 75 Hz applied every 1.5 s). During the first minute of the fatigue regimen, the effects of AChE inhibition were already near maximal, including marked reductions in peak tension and the force-time integral (area), as well as a decrement of compound muscle action potential amplitudes within a stimulus train. Neuromuscular transmission failure was the major contributor of the force decreases in the AChE inhibited muscles. However, despite this neuromuscular transmission failure, muscles of which all AChE molecular forms were nearly completely inhibited were still able to function, although abnormally, during 3 min of intermittent high frequency nerve stimulation. PMID- 10517779 TI - Muscle pump and central command during recovery from exercise in humans. AB - We sought to determine the relative contributions of cessation of skeletal muscle pumping and withdrawal of central command to the rapid decrease in arterial pressure during recovery from exercise. Twelve healthy volunteers underwent three exercise sessions, each consisting of a warm-up, 3 min of cycling at 60% of maximal heart rate, and 5 min of one of the following recovery modes: seated (inactive), loadless pedaling (active), and passive cycling. Mean arterial pressure (MAP), cardiac output, thoracic impedance, and heart rate were measured. When measured 15 s after exercise, MAP decreased less (P < 0.05) during the active (-3 +/- 1 mmHg) and passive (-6 +/- 1 mmHg) recovery modes than during inactive (-18 +/- 2 mmHg) recovery. These differences in MAP persisted for the first 4 min of recovery from exercise. Significant maintenance of central blood volume (thoracic impedance), stroke volume, and cardiac output paralleled the maintenance of MAP during active and passive conditions during 5 min of recovery. These data indicate that engaging the skeletal muscle pump by loadless or passive pedaling helps maintain MAP during recovery from submaximal exercise. The lack of differences between loadless and passive pedaling suggests that cessation of central command is not as important. PMID- 10517780 TI - Portal glucose infusion increases hepatic glycogen deposition in conscious unrestrained rats. AB - It has been demonstrated in the conscious dog that portal glucose infusion creates a signal that increases net hepatic glucose uptake and hepatic glycogen deposition. Experiments leading to an understanding of the mechanism by which this change occurs will be facilitated if this finding can be reproduced in the rat. Rats weighing 275-300 g were implanted with four indwelling catheters (one in the portal vein, one in the left carotid artery, and two in the right jugular vein) that were externalized between the scapulae. The rats were studied in a conscious, unrestrained condition 7 days after surgery, following a 24-h fast. Each experiment consisted of a 30- to 60-min equilibration, a 30-min baseline, and a 120-min test period. In the test period, a pancreatic clamp was performed by using somatostatin, insulin, and glucagon. Glucose was given simultaneously either through the jugular vein to clamp the arterial blood level at 220 mg/dl (Pe low group) or at 250 mg/dl (Pe high group), or via the hepatic portal vein (Po group; 6 mg. kg(-1). min(-1)) and the jugular vein to clamp the arterial blood glucose level to 220 mg/dl. In the test period, the arterial plasma glucagon and insulin levels were not significantly different in the three groups (36 +/- 2, 33 +/- 2, and 30 +/- 2 pg/ml and 1.34 +/- 0.08, 1. 37 +/- 0.18, and 1.66 +/- 0.11 ng/ml in Po, Pe low, and Pe high groups, respectively). The arterial blood glucose levels during the test period were 224 +/- 4 mg/dl for Po, 220 +/- 3 for Pe low, and 255 +/- 2 for Pe high group. The liver glycogen content (micromol glucose/g liver) in the two Pe groups was not statistically different (51 +/- 7 and 65 +/- 8, respectively), whereas the glycogen level in the Po group was significantly greater (93 +/- 9, P < 0.05). Because portal glucose delivery also augments hepatic glycogen deposition in the rat, as it does in the dogs, mechanistic studies relating to its function can now be undertaken in this species. PMID- 10517781 TI - Hindlimb unweighting decreases ecNOS gene expression and endothelium-dependent dilation in rat soleus feed arteries. AB - We tested the hypothesis that hindlimb unweighting (HLU) and the associated reduction in soleus muscle blood flow causes decreased expression of endothelial cell nitric oxide synthase (ecNOS) mRNA and protein and attenuated endothelium dependent vasodilator responses in rat soleus feed arteries (SFA). Male Sprague Dawley rats were exposed to HLU (n = 12) or cage control (Con; n = 12) conditions for 14 days. At the end of this period, SFA were isolated, removed, and cannulated with two glass micropipettes for examination of vasodilator responses or frozen for analysis of ecNOS mRNA and protein expression. RT-PCR of RNA from single SFA was used to measure ecNOS mRNA, and immunoblots on single SFAs were used to measure ecNOS protein content. Results revealed that both ecNOS mRNA and ecNOS protein expression were lower in SFA from HLU rats. Dilation to increased intraluminal flow was attenuated in SFA from HLU rats (Con: 88 +/- 8% vs. HLU: 53 +/- 8%) as was maximal vasodilation to acetylcholine (10(-9)-10(-4) M; Con: 88 +/ 5% vs. HLU: 73 +/- 5%). Sensitivity to the endothelium-independent vasodilator sodium nitroprusside (10(-10)-10(-4) M) was enhanced by HLU (EC(50): Con: 4.46 x 10(-7) M vs. HLU: 5.00 x 10(-8) M). Collectively, these data indicate that the chronic reduction in soleus blood flow associated with the reduced physical activity during HLU results in reduced expression of ecNOS mRNA and protein in SFA and attenuated endothelium-dependent vasodilation. PMID- 10517782 TI - Muscle interstitial glucose and lactate levels during dynamic exercise in humans determined by microdialysis. AB - The purpose of the present study was to use the microdialysis technique to determine skeletal muscle interstitial glucose and lactate concentrations during dynamic incremental exercise in humans. Microdialysis probes were inserted into the vastus lateralis muscle, and subjects performed knee extensor exercise at workloads of 10, 20, 30, 40, and 50 W. The in vivo probe recoveries determined at rest by the internal reference method for glucose and lactate were 28.7 +/- 2.5 and 32.0 +/- 2.7%, respectively. As exercise intensity increased, probe recovery also increased, and at the highest workload probe recovery for glucose (61.0 +/- 3.9%) and lactate (66. 3 +/- 3.6%) had more than doubled. At rest the interstitial glucose concentration (3.5 +/- 0.2 mM) was lower than both the arterial (5.6 +/- 0.2 mM) and venous (5.3 +/- 0.3 mM) plasma water glucose levels. The interstitial glucose levels remained lower (P < 0.05) than the arterial and venous plasma water glucose concentrations during exercise at all intensities and at 10, 20, 30, and 50 W, respectively. At rest the interstitial lactate concentration (2.5 +/- 0.2 mM) was higher (P < 0.05) than both the arterial (0.9 +/- 0. 2 mM) and venous (1.1 +/- 0.2 mM) plasma water lactate levels. This relationship was maintained (P < 0.05) during exercise at workloads of 10, 20, and 30 W. These data suggest that interstitial glucose delivery at rest is flow limited and that during exercise changes in the interstitial concentrations of glucose and lactate mirror the changes observed in the venous plasma water compartments. Furthermore, skeletal muscle contraction results in an increase in the diffusion coefficient of glucose and lactate within the interstitial space as reflected by an elevation in probe recovery during exercise. PMID- 10517783 TI - Lung elastic recoil during breathing at increased lung volume. AB - During dynamic hyperinflation with induced bronchoconstriction, there is a reduction in lung elastic recoil at constant lung volume (R. Pellegrino, O. Wilson, G. Jenouri, and J. R. Rodarte. J. Appl. Physiol. 81: 964-975, 1996). In the present study, lung elastic recoil at control end inspiration was measured in normal subjects in a volume displacement plethysmograph before and after voluntary increases in mean lung volume, which were achieved by one tidal volume increase in functional residual capacity (FRC) with constant tidal volume and by doubling tidal volume with constant FRC. Lung elastic recoil at control end inspiration was significantly decreased by approximately 10% within four breaths of increasing FRC. When tidal volume was doubled, the decrease in computed lung recoil at control end inspiration was not significant. Because voluntary increases of lung volume should not produce airway closure, we conclude that stress relaxation was responsible for the decrease in lung recoil. PMID- 10517784 TI - Differential microvascular response to disuse in rat hindlimb skeletal muscles. AB - The aim of the study was to address discrepant findings in the literature regarding coupling between decreased functional demand during disuse and reduced capillarity. We previously reported [K. Tyml, O. Mathieu-Costello, and E. Noble. Microvasc. Res. 49: 17-32, 1995] that severe disuse of rat extensor digitorum longus (EDL) muscle caused by a 2-wk application of tetrodotoxin (TTX) on the sciatic nerve is not accompanied by capillary loss. Using the same animal model, the present study examined whether this absence of coupling could be explained in terms of 1) too short a duration of disuse and 2) muscle-specific response to disuse. Fischer 344 rats were exposed to either no treatment (control) or to 2- or 8-wk TTX applications. Fiber size, capillary density per fiber cross-sectional area, and capillary-to-fiber (C/F) ratio were determined by morphometry in the EDL muscle (control, 2- and 8-wk groups) and in the superficial portion of medial gastrocnemius (Gas) muscle (control, 2 wk). In both muscles, microvascular blood flow was evaluated by intravital microscopy [red blood cell velocity in capillaries (V(RBC))] and by laser Doppler flowmetry (LDF). Regardless of duration of TTX application or muscle type, TTX-induced disuse resulted in a significant reduction of fiber area (44-71%). However, capillary density increased in EDL muscle (both at 2 and 8 wk) but not in Gas muscle. C/F ratio decreased in EDL muscle at 8 wk (18%) and in Gas muscle (39%). This indicates that the effect on capillarity depended on duration of disuse and on muscle type. V(RBC) and LDF signal were significantly larger in EDL than in Gas muscle. Analysis of change in capillarity vs. V(RBC) suggested that the outcome of disuse may be modulated by blood flow. We conclude that the duration of skeletal muscle disuse per se does not dictate capillary loss, and we hypothesize that discrepant findings of coupling between functional demand and capillarity could be due to the presence/absence of flow-related angiogenesis superimposed on the capillary removal process during disuse. PMID- 10517785 TI - Measurement of cardiac output during exercise by open-circuit acetylene uptake. AB - Noninvasive measurement of cardiac output (QT) is problematic during heavy exercise. We report a new approach that avoids unpleasant rebreathing and resultant changes in alveolar PO(2) or PCO(2) by measuring short-term acetylene (C(2)H(2)) uptake by an open-circuit technique, with application of mass balance for the calculation of QT. The method assumes that alveolar and arterial C(2)H(2) pressures are the same, and we account for C(2)H(2) recirculation by extrapolating end-tidal C(2)H(2) back to breath 1 of the maneuver. We correct for incomplete gas mixing by using He in the inspired mixture. The maneuver involves switching the subject to air containing trace amounts of C(2)H(2) and He; ventilation and pressures of He, C(2)H(2), and CO(2) are measured continuously (the latter by mass spectrometer) for 20-25 breaths. Data from three subjects for whom multiple Fick O(2) measurements of QT were available showed that measurement of QT by the Fick method and by the C(2)H(2) technique was statistically similar from rest to 90% of maximal O(2) consumption (VO(2 max)). Data from 12 active women and 12 elite male athletes at rest and 90% of VO(2 max) fell on a single linear relationship, with O(2) consumption (VO(2)) predicting QT values of 9.13, 15.9, 22.6, and 29.4 l/min at VO(2) of 1, 2, 3, and 4 l/min. Mixed venous PO(2) predicted from C(2)H(2)-determined QT, measured VO(2), and arterial O(2) concentration was approximately 20-25 Torr at 90% of VO(2 max) during air breathing and 10-15 Torr during 13% O(2) breathing. This modification of previous gas uptake methods, to avoid rebreathing, produces reasonable data from rest to heavy exercise in normal subjects. PMID- 10517786 TI - Validity of fan-beam dual-energy X-ray absorptiometry for measuring fat-free mass and leg muscle mass. Health, Aging, and Body Composition Study--Dual-Energy X-ray Absorptiometry and Body Composition Working Group. AB - The aim of the study was to examine the accuracy of fan-beam dual-energy X-ray absorptiometry (DEXA) for measuring total body fat-free mass (FFM) and leg muscle mass (MM) in elderly persons. Participants were 60 men and women aged 70-79 yr and with a body mass index of 17.5-39.8 kg/m(2). FFM and MM at four leg regions were measured by using DEXA (Hologic 4500A, v8.21). A four-compartment body composition model (4C) and multislice computed tomography (CT) of the legs were used as the criterion methods for FFM and MM, respectively. FFM by DEXA was positively associated with FFM by 4C (R(2) = 0.98, SE of estimate = 1.6 kg). FFM by DEXA was higher [53.5 +/- 12.0 (SD) kg] than FFM by 4C (51.6 +/- 11.9 kg; P < 0.001). No association was observed between the difference and the mean of the two methods. MM by DEXA was positively associated with CT at all four leg regions (R(2) = 0.86-0.96). MM by DEXA was higher than by CT in three regions. The results of this study suggest that fan-beam DEXA offers considerable promise for the measurement of total body FFM and leg MM in elderly persons. PMID- 10517787 TI - A model of extravascular bubble evolution: effect of changes in breathing gas composition. AB - Observations of bubble evolution in rats after decompression from air dives (O. Hyldegaard and J. Madsen. Undersea Biomed. Res. 16: 185-193, 1989; O. Hyldegaard and J. Madsen. Undersea Hyperbaric Med. 21: 413-424, 1994; O. Hyldegaard, M. Moller, and J. Madsen. Undersea Biomed. Res. 18: 361-371, 1991) suggest that bubbles may resolve more safely when the breathing gas is a heliox mixture than when it is pure O(2). This is due to a transient period of bubble growth seen during switches to O(2) breathing. In an attempt to understand these experimental results, we have developed a multigas-multipressure mathematical model of bubble evolution, which consists of a bubble in a well-stirred liquid. The liquid exchanges gas with the bubble via diffusion, and the exchange between liquid and blood is described by a single-exponential time constant for each inert gas. The model indicates that bubbles resolve most rapidly in spinal tissue, in adipose tissue, and in aqueous tissues when the breathing gas is switched to O(2) after surfacing. In addition, the model suggests that switching to heliox breathing may prolong the existence of the bubble relative to breathing air for bubbles in spinal and adipose tissues. Some possible explanations for the discrepancy between model and experiment are discussed. PMID- 10517788 TI - Simultaneous measurement of nitric oxide production by conducting and alveolar airways of humans. AB - Human airways produce nitric oxide (NO), and exhaled NO increases as expiratory flow rates fall. We show that mixing during exhalation between the NO produced by the lower, alveolar airways (VL(NO)) and the upper conducting airways (VU(NO)) explains this phenomenon and permits measurement of VL(NO), VU(NO), and the NO diffusing capacity of the conducting airways (DU(NO)). After breath holding for 10-15 s the partial pressure of alveolar NO (PA) becomes constant, and during a subsequent exhalation at a constant expiratory flow rate the alveoli will deliver a stable amount of NO to the conducting airways. The conducting airways secrete NO into the lumen (VU(NO)), which mixes with PA during exhalation, resulting in the observed expiratory concentration of NO (PE). At fast exhalations, PA makes a large contribution to PE, and, at slow exhalations, NO from the conducting airways predominates. Simple equations describing this mixing, combined with measurements of PE at several different expiratory flow rates, permit calculation of PA, VU(NO), and DU(NO). VL(NO) is the product of PA and the alveolar airway diffusion capacity for NO. In seven normal subjects, PA = 1.6 +/- 0.7 x 10(-6) (SD) Torr, VL(NO) = 0.19 +/- 0.07 microl/min, VU(NO) = 0.08 +/- 0.05 microl/min, and DU(NO) = 0.4 +/- 0.4 ml. min(-1). Torr(-1). These quantitative measurements of VL(NO) and VU(NO) are suitable for exploring alterations in NO production at these sites by diseases and physiological stresses. PMID- 10517789 TI - The original presentation of Boyle's law. AB - The original presentation of what we know as Boyle's law has several interesting features. First, the technical difficulties of the experiment were considerable, because Boyle used a glass tube full of mercury that was nearly 2.5 m long, and the large pressures sometimes shattered the glass. Next, Boyle's table of results contains extremely awkward fractions, such 10/13, 2/17, 13/19, and 18/23, which look very strange to us today. This was because he calculated the pressure for a certain volume of gas by using simple multiplication and division, keeping the vulgar fractions. Boyle was not able to express the numbers as decimals because this notation was not in common use at the time. Finally, his contention that pressure and volume were inversely related depended on the reader's comparing two sets of numbers in adjacent columns to see how well they agreed. Today we would plot the data, but again orthogonal graphs were not in general use in 1662. When Boyle's data are plotted by using modern conventional methods, they strongly support his hypothesis that the volume and pressure of a gas are inversely related. PMID- 10517790 TI - Interrupter airway and tissue resistance: errors caused by valve properties and respiratory system compliance. AB - The interrupter technique is used to determine airway and tissue resistance. Their accuracy is influenced by the technical properties of the interrupter device and the compliance of the respiratory system. We investigated the influence of valve characteristics and respiratory system compliance on the accuracy of determining airway and tissue resistance by means of a computer simulation. With decreasing compliance we found increasing errors in both airway and tissue resistance determination of up to 34 and 71%, respectively. On this basis we developed a new occlusion valve, with special emphasis on rapid closing time and tightness in the closed state to improve the accuracy of resistance determination. The newly developed occlusion device greatly improves the accuracy of airway and tissue resistance determination. We conclude that respiratory system compliance is a limiting factor for the accuracy of the interrupter technique. To apply the interrupter technique in patients with extremely low respiratory system compliances, we need sophisticated technical devices. PMID- 10517791 TI - Measurement of limb venous compliance in humans: technical considerations and physiological findings. AB - We conducted a series of studies to develop and test a rapid, noninvasive method to measure limb venous compliance in humans. First, we measured forearm volume (mercury-in-Silastic strain gauges) and antecubital intravenous pressure during inflation of a venous collecting cuff around the upper arm. Intravenous pressure fit the regression line, -0.3 +/- 0.7 + 0.95 +/- 0.02. cuff pressure (r = 0.99 +/ 0.00), indicating cuff pressure is a good index of intravenous pressure. In subsequent studies, we measured forearm and calf venous compliance by inflating the venous collecting cuff to 60 mmHg for 4 min, then decreasing cuff pressure at 1 mmHg/s (over 1 min) to 0 mmHg, using cuff pressure as an estimate of venous pressure. This method produced pressure-volume curves fitting the quadratic regression (Deltalimb volume) = beta(0) + beta(1). (cuff pressure) + beta(2). (cuff pressure)(2), where Delta is change. Curves generated with this method were reproducible from day to day (coefficient of variation: 4.9%). In 11 subjects we measured venous compliance via this method under two conditions: with and without (in random order) superimposed sympathetic activation (ischemic handgrip exercise to fatigue followed by postexercise ischemia). Calf and forearm compliance did not differ between control and sympathetic activation (P > 0.05); however, the data suggest that unstressed volume was reduced by the maneuver. These studies demonstrate that venous pressure-volume curves can be generated both rapidly and noninvasively with this technique. Furthermore, the results suggest that although whole-limb venous compliance is under negligible sympathetic control in humans, unstressed volume can be affected by the sympathetic nervous system. PMID- 10517792 TI - K+ channel blockers and Ca2+ signals in basket cell terminals. PMID- 10517793 TI - A scaffolding for regulation of volume-sensitive Cl- channels. PMID- 10517794 TI - Smooth muscle: PKC-induced Ca2+ sensitisation by myosin phosphatase inhibition. PMID- 10517795 TI - The micro-mechanics of single molecules studied with atomic force microscopy. AB - The atomic force microscope (AFM) in its force-measuring mode is capable of effecting displacements on an angstrom scale (10 A = 1 nm) and measuring forces of a few piconewtons. Recent experiments have applied AFM techniques to study the mechanical properties of single biological polymers. These properties contribute to the function of many proteins exposed to mechanical strain, including components of the extracellular matrix (ECM). The force-bearing proteins of the ECM typically contain multiple tandem repeats of independently folded domains, a common feature of proteins with structural and mechanical roles. Polysaccharide moieties of adhesion glycoproteins such as the selectins are also subject to strain. Force-induced extension of both types of molecules with the AFM results in conformational changes that could contribute to their mechanical function. The force-extension curve for amylose exhibits a transition in elasticity caused by the conversion of its glucopyranose rings from the chair to the boat conformation. Extension of multi-domain proteins causes sequential unraveling of domains, resulting in a force-extension curve displaying a saw tooth pattern of peaks. The engineering of multimeric proteins consisting of repeats of identical domains has allowed detailed analysis of the mechanical properties of single protein domains. Repetitive extension and relaxation has enabled direct measurement of rates of domain unfolding and refolding. The combination of site directed mutagenesis with AFM can be used to elucidate the amino acid sequences that determine mechanical stability. The AFM thus offers a novel way to explore the mechanical functions of proteins and will be a useful tool for studying the micro-mechanics of exocytosis. PMID- 10517796 TI - Sea urchin egg preparations as systems for the study of calcium-triggered exocytosis. AB - This paper reviews recent work in our laboratory on the mechanism of calcium triggered exocytosis. Upon echinoderm egg fertilization, cortical secretory vesicle exocytosis is massive and synchronous. By combining physiological and molecular analyses with a variety of purified membrane isolates containing secretory vesicles that fuse to the plasma membrane or each other, we have characterized the final steps of this calcium-triggered exocytosis. Our kinetic analysis led to a functional definition of the fusion complex whose activation by calcium follows Poisson statistics. The properties of this complex are compared with the properties of the heterotrimeric SNARE protein complex that is present in the cortical vesicle system. Our data do not support the hypothesis that this particular heterotrimeric complex is by itself the biological fusogen. PMID- 10517797 TI - Stimulation of exocytosis without a calcium signal. AB - More than 30 years ago, Douglas (Douglas & Rubin, 1961; Douglas, 1968) proposed that intracellular Ca2+ controls stimulus-secretion coupling in endocrine cells, and Katz & Miledi (1967; Katz, 1969) proposed that intracellular Ca2+ ions control the rapid release of neurotransmitters from synapses. These related hypotheses have been amply confirmed in subsequent years and for students of excitable cells, they dominate our teaching and research. Calcium controls regulated exocytosis. On the other hand, many studies of epithelial and blood cell biology emphasize Ca2+-independent regulation of secretion of mucin, exocytotic delivery of transporters and degranulation. The evidence seems good. Are these contrasting conclusions somehow mistaken, or are the dominant factors controlling exocytosis actually different in different cell types? In this essay, we try to reconcile these ideas and consider classes of questions to ask and hypotheses to test in seeking a more integrated understanding of excitation secretion coupling. Our review is conceptual and narrowly selective of a few examples rather than referring to a broader range of useful studies in the extensive literature. The examples are taken from mammals and are documented principally by citing other reviews and two of our own studies. The evidence shows that protein phosphorylation by kinases potentiates Ca2+-dependent exocytosis and often suffices to induce exocytosis by itself. Apparently, protein phosphorylation is the physiological trigger in a significant number of examples of regulated exocytosis. We conclude that although sharing many common properties, secretory processes in different cells are specialized and distinct from each other. PMID- 10517799 TI - Neurosecretory cells without neurosecretion: evidence of an independently regulated trait of the cell phenotype. AB - Neurosecretion competence is a fundamental property that enables differentiated neurones and professional neurosecretory cells to store neurotransmitters and hormones in specialized organelles, the synaptic-like vesicles and dense granules, and to release them by regulated exocytosis. In our laboratory, the study of rat phaeochromocytoma (PC12) clones that fail to express the above organelles or any other components involved in neurosecretion, whilst maintaining most of the general markers of the parental population, has served to demonstrate that this trait is controlled independently from the rest of the phenotype. The present review focuses on recent advances in elucidating the molecular mechanisms governing neurosecretion competence. Moreover, the opportunities that such neurosecretion-defective PC12 clones offer for the investigation of new aspects of regulated exocytosis and the localization of its components are summarized. PMID- 10517798 TI - Proteins involved in synaptic vesicle trafficking. AB - Neurotransmitter release relies on a series of synaptic vesicle trafficking reactions. We have determined the molecular basis of these reactions by microinjecting, into 'giant' nerve terminals of squid, probes that interfere with presynaptic proteins. These probes affect neurotransmitter release and disrupt nerve terminal structure. From the nature of these lesions, it is possible to deduce the roles of individual proteins in specific vesicle trafficking reactions. This approach has revealed the function of more than a dozen presynaptic proteins and we hypothesize that neurotransmitter release requires the coordinated action of perhaps 50-100 proteins. PMID- 10517800 TI - Identification of an inhibitory Zn2+ binding site on the human glycine receptor alpha1 subunit. AB - 1. Whole-cell glycine-activated currents were recorded from human embryonic kidney (HEK) cells expressing wild-type and mutant recombinant homomeric glycine receptors (GlyRs) to locate the inhibitory binding site for Zn2+ ions on the human alpha1 subunit. 2. Glycine-activated currents were potentiated by low concentrations of Zn2+ (<10 microM) and inhibited by higher concentrations (>100 microM) on wild-type alpha1 subunit GlyRs. 3. Lowering the external pH from 7.4 to 5.4 inhibited the glycine responses in a competitive manner. The inhibition caused by Zn2+ was abolished leaving an overt potentiating effect at 10 microM Zn2+ that was exacerbated at 100 microM Zn2+. 4. The identification of residues involved in the formation of the inhibitory binding site was also assessed using diethylpyrocarbonate (DEPC), which modifies histidines. DEPC (1 mM) abolished Zn2+-induced inhibition and also the potentiation of glycine-activated currents by Zn2+. 5. The reduction in glycine-induced whole-cell currents in the presence of high (100 microM) concentrations of Zn2+ did not increase the rate of glycine receptor desensitisation. 6. Systematic mutation of extracellular histidine residues in the GlyR alpha1 subunit revealed that mutations H107A or H109A completely abolished inhibition of glycine-gated currents by Zn2+. However, mutation of other external histidines, H210, H215 and H419, failed to prevent inhibition by Zn2+ of glycine-gated currents. Thus, H107 and H109 in the extracellular domain of the human GlyR alpha1 subunit are major determinants of the inhibitory Zn2+ binding site. 7. An examination of Zn2+ co-ordination in metalloenzymes revealed that the histidine- hydrophobic residue-histidine motif found to be responsible for binding Zn2+ in the human GlyR alpha1 subunit is also shared by some of these enzymes. Further comparison of the structure and location of this motif with a generic model of the GlyR alpha1 subunit suggests that H107 and H109 participate in the formation of the inhibitory Zn2+ binding site at the apex of a beta sheet in the N-terminal extracellular domain. PMID- 10517801 TI - Modulation by K+ channels of action potential-evoked intracellular Ca2+ concentration rises in rat cerebellar basket cell axons. AB - 1. Action potential-evoked [Ca2+]i rises in basket cell axons of rat cerebellar slices were studied using two-photon laser scanning microscopy and whole-cell recording, to identify the K+ channels controlling the shape of the axonal action potential. 2. Whole-cell recordings of Purkinje cell IPSCs were used to screen K+ channel subtypes which could contribute to axonal repolarization. alpha Dendrotoxin, 4-aminopyridine, charybdotoxin and tetraethylammonium chloride increased IPSC rate and/or amplitude, whereas iberiotoxin and apamin failed to affect the IPSCs. 3. The effects of those K+ channel blockers that enhanced transmitter release on the [Ca2+]i rises elicited in basket cell axons by action potentials fell into three groups: 4-aminopyridine strongly increased action potential-evoked [Ca2+]i; tetraethylammonium and charybdotoxin were ineffective alone but augmented the effects of 4-aminopyridine; alpha-dendrotoxin had no effect. 4. We conclude that cerebellar basket cells contain at least three pharmacologically distinct K+ channels, which regulate transmitter release through different mechanisms. 4-Aminopyridine-sensitive, alpha-dendrotoxin insensitive K+ channels are mainly responsible for repolarization in basket cell presynaptic terminals. K+ channels blocked by charybdotoxin and tetraethylammonium have a minor role in repolarization. alpha-Dendrotoxin sensitive channels are not involved in shaping the axonal action potential waveform. The two last types of channels must therefore exert control of synaptic activity through a pathway unrelated to axonal action potential broadening. PMID- 10517802 TI - Functional characterization of a Na+-phosphate cotransporter (NaPi-II) from zebrafish and identification of related transcripts. AB - 1. We report the molecular identification of a Na+-Pi (inorganic phosphate) cotransport system of the NaPi-II protein family from zebrafish intestine. Following a PCR-related strategy, a DNA fragment from intestine-derived RNA was isolated. Rapid amplification of cDNA ends (3'- and 5'-RACE) resulted in the complete sequence (2607 bp) containing an open reading frame of 1893 bp. 2. The NaPi-II-related protein was expressed in Xenopus laevis oocytes and the resulting transport activity was analysed by electrophysiological means. The apparent Km for Pi was 250 microM (96 mM Na+, -60 mV), and voltage-dependent binding of Na+ exhibited a Km of 67.1 mM (1 mM Pi, -60 mV). 3. Interestingly, the overall transport activity was almost insensitive to changes in the holding potential. The apparent affinity for Na+ decreased under hyperpolarizing conditions, whereas Pi binding showed no voltage dependence. Transport activity was inhibited at low pH, which is characteristic for renal NaPi-II isoforms. 4. The expression of the NaPi-II-related isoform was addressed by reverse-transcription PCR. The mRNA could be detected in intestine, liver, eye and kidney. Unexpectedly, a second NaPi-II-related isoform was identified and found to be expressed in kidney, intestine, liver, brain, eye and prominently in testis. In addition, a shorter amplicon was demonstrated to be an antisense transcript related to the NaPi-II intestinal isoform. PMID- 10517803 TI - Identification of amino acids within the P2X2 receptor C-terminus that regulate desensitization. AB - 1. The ATP-activated P2X2(a) and P2X2(b) receptor splice variants, which differ only in their C-terminal sequences, desensitize at different rates. We used mutational analysis to investigate the involvement of the C-terminal region in receptor desensitization. Rat wild-type and mutant P2X2 receptors were expressed in Xenopus oocytes and currents were measured using the two-electrode voltage clamp technique. 2. Truncating P2X2 at the Lys369 splice site increased the rate of desensitization by >100-fold. Recovery from desensitization was slowed by approximately 5-fold. 3. Addition of Val370 onto the C-terminus of the truncated receptor slowed desensitization by approximately 70-fold. Point mutations that substituted either smaller or larger hydrophobic amino acids for Val370, within the P2X2(a) splice variant, had profound effects on the rate of desensitization. The rate decreased with increasing hydrophobicity but was not dependent upon the precise structure of the side group. 4. A mutant receptor, with only nine amino acids, Val-Asp-Pro-Lys-Gly-Leu-Ala-Gln-Leu, beyond the Lys369 splice site, desensitized at a similar rate to P2X2(a). Injection of the peptide of this sequence into oocytes expressing P2X2(a) increased the rate of desensitization, whereas the eight-residue peptide lacking the valine had no effect. 5. Neutralizing lysines in the vicinity of the splice site increased the rate of receptor desensitization. Substituting glutamine for Lys365 produced the greatest effect ( approximately 30-fold increase), whereas mutating lysines that were further upstream or downstream of this position had progressively less of an effect. 6. We conclude that the C-terminal splice site of the P2X2 receptor is located within a region that is critically involved in regulating the rate of receptor desensitization. The valine at position 370 interacts with an intracellular hydrophobic site to slow the rate of desensitization. Nearby lysines may facilitate this interaction. PMID- 10517804 TI - Muscarinic M1 receptors activate phosphoinositide turnover and Ca2+ mobilisation in rat sympathetic neurones, but this signalling pathway does not mediate M current inhibition. AB - 1. The relationship between muscarinic receptor activation, phosphoinositide turnover, calcium mobilisation and M-current inhibition has been studied in rat superior cervical ganglion (SCG) neurones in primary culture. 2. Phosphoinositide specific phospholipase C (PLC) stimulation was measured by the accumulation of [3H]-cytidine monophosphate phosphatidate (CMP-PA) after incubation with [3H] cytidine in the presence of Li+. The muscarinic agonist oxotremorine methiodide (oxo-M) stimulated PLC in a dose-dependent manner with an EC50 of approximately 3.5 microM. 3. The concentration-response curve for oxo-M was shifted to the right by a factor of about 10 by pirenzepine (100 nM), suggesting a pKB (-log of the apparent dissociation constant) of 7.9 +/- 0.4, while himbacine (1 microM) shifted the curve by a factor of about 13 (pKB approximately 7.1 +/- 0.6). This indicates involvement of the M1 muscarinic receptor in this response. 4. The accumulation of CMP-PA was localised by in situ autoradiography to SCG principal neurones, with no detectable signal in glial cells present in the primary cultures. 5. The ability of oxo-M to release Ca2+ from inositol(1,4, 5)trisphosphate (InsP3)-sensitive stores was determined by fura-2 microfluorimetry of SCG neurones voltage clamped in perforated patch mode. Oxo-M failed to evoke intracellular Ca2+ (Ca2+i) mobilisation in SCG neurones voltage clamped at -60 mV, but produced a significant Ca2+i rise (67 +/- 15 nM, n = 9) in cells voltage clamped at -25 mV. 6. Thapsigargin (0.5-1 microM) caused a 70 % inhibition of the oxo-M-induced Ca2+i increase, indicating its intracellular origin, while oxo-M-induced inhibition of M-current in the same cells was unaffected by thapsigargin. 7. Our results do not support the involvement of InsP3-sensitive calcium mobilisation in M-current inhibition. PMID- 10517805 TI - Caveolin-1 modulates the activity of the volume-regulated chloride channel. AB - 1. Caveolae are small invaginations of the plasma membrane that have recently been implicated in signal transduction. In the present study, we have investigated whether caveolins, the principal protein of caveolae, also modulate volume-regulated anion channels (VRACs). 2. ICl,swell, the cell swelling-induced chloride current through VRACs, was studied in three caveolin-1-deficient cell lines: Caco-2, MCF-7 and T47D. 3. Electrophysiological measurements showed that ICl, swell was very small in these cells and that transient expression of caveolin-1 restored ICl,swell. The caveolin-1 effect was isoform specific: caveolin-1beta but not caveolin-1alpha upregulated VRACs. This correlated with a different subcellular distribution of caveolin-1alpha (perinuclear location) from caveolin-1beta (perinuclear and peripheral). 4. To explain the modulation of ICl, swell by caveolin-1 we propose that caveolin increases the availability of VRACs in the plasma membrane or, alternatively, that it plays a crucial role in the signal transduction cascade of VRACs. PMID- 10517806 TI - Shift from depolarizing to hyperpolarizing glycine action in rat auditory neurones is due to age-dependent Cl- regulation. AB - 1. The inhibitory neurotransmitter glycine can elicit depolarizing responses in immature neurones. We investigated the changes in glycine responses and their ionic mechanism in developing neurones of the rat lateral superior olive (LSO), an auditory brainstem nucleus involved in sound localization. 2. Whole-cell and gramicidin perforated-patch recordings were performed from visually identified LSO neurones in brain slices and glycine was pressure applied for 3-100 ms to the soma. Glycine-evoked currents were reversibly blocked by strychnine. They were mostly monophasic, but biphasic responses occurred in approximately 30 % of P8-11 neurones in perforated-patch recordings. 3. In whole-cell recordings from P2-11 neurones, the reversal potential of glycine-evoked currents (EGly) was determined by the transmembranous Cl- gradient and corresponded closely to the Nernst potential for Cl-, regardless of age. This indicates that Cl- is the principle ion permeating glycine receptors, but is also consistent with a low relative (10 20 %) permeability for HCO3-. The Cl- gradient also determined the polarity and amplitude of glycine-evoked membrane potential changes. 4. Leaving the native intracellular [Cl-] undisturbed with gramicidin perforated-patch recordings, we found a highly significant, age-dependent change of EGly from -46.8 +/- 1.8 mV (P1-4, n = 28) to -67.6 +/- 3.3 mV (P5-8, n = 10) to -82.2 +/- 4.1 mV (P9-11, n = 18). The majority of P1-4 neurones were depolarized by glycine ( approximately 80 %) and spikes were evoked in approximately 30 %. In contrast, P9-11 neurones were hyperpolarized. 5. In perforated-patch recordings, EGly was influenced by the voltage protocol and the glycine application interval; it could be shifted in the positive and negative direction. For a given application interval, these shifts were always larger in P1-4 than in P8-11 neurones, pointing to less effective Cl- regulation mechanisms in younger neurones. 6. Furosemide (frusemide), a blocker of cation-Cl- cotransporters, reversibly shifted EGly in the negative direction in P2-4 neurones, yet in the positive direction in P8-10 neurones, suggesting the blockade of net inward and net outward Cl- transporters, respectively. 7. Taken together, age-dependent changes in active Cl- regulation are likely to cause the developmental shift from depolarizing to hyperpolarizing glycine responses. A high intracellular [Cl-] is generated in neonatal LSO neurones which decreases during maturation. PMID- 10517807 TI - Reconstitution of protein kinase C-induced contractile Ca2+ sensitization in triton X-100-demembranated rabbit arterial smooth muscle. AB - 1. Triton X-100-demembranated smooth muscle loses Ca2+-sensitizing responsiveness to protein kinase C (PKC) activators while intact and alpha-toxin-permeabilized smooth muscles remain responsive. We attempted to reconstitute the contractile Ca2+ sensitization by PKC in the demembranated preparations. 2. Western blot analyses showed that the content of the PKC alpha-isoform (PKCalpha) was markedly reduced and that the smooth muscle-specific protein phosphatase-1 inhibitor protein CPI-17 was not detectable, while the amount of calponin and actin still remained similar to those of intact strips. 3. Unphosphorylated recombinant CPI 17 alone induced a small but significant contraction at constant Ca2+. Isoform selective PKC inhibitors inhibited unphosphorylated but not pre thiophosphorylated CPI-17-induced contraction, suggesting that in situ conventional PKC isoform(s) can phosphorylate CPI-17. 4. Exogenously replenishing PKCalpha alone did not induce potentiation of contraction and only slowly increased myosin light chain (MLC) phosphorylation at submaximal Ca2+. 5. PKC in the presence of CPI-17, but not the [T38A]-CPI mutant, markedly induced potentiation of both contraction and MLC phosphorylation. CPI-17 itself was phosphorylated. 6. In in vitro experiments, CPI-17 was a much better substrate for PKCalpha than calponin, caldesmon, MLC and myosin. 7. Our results indicate that PKC requires CPI-17 phosphorylation at Thr-38 but not calponin for reconstitution of the contractile Ca2+ sensitization in the demembranated arterial smooth muscle. PMID- 10517808 TI - The role of the sarcoplasmic reticulum as a Ca2+ sink in rat uterine smooth muscle cells. AB - 1. The mechanisms responsible for removing calcium ions from the cytoplasm were investigated in single rat uterine myocytes using indo-1. 2. Trains of depolarizing voltage-clamp pulses increased [Ca2+]i. The rate of decay of [Ca2+]i was slowed by inhibition of the sarcoplasmic reticulum (SR) Ca2+-ATPase with cyclopiazonic acid (CPA). However, if the sarcolemmal Na+-Ca2+ exchanger and Ca2+ ATPase were inhibited then recovery of [Ca2+]i was abolished showing that the SR Ca2+-ATPase alone cannot produce decay of [Ca2+]i. 3. In another series of experiments, Ca2+ release from the SR was induced with carbachol in a Ca2+-free solution. Under these conditions responses to repeated applications of carbachol could be obtained. In the presence of CPA, however, only the first application was effective. This suggests that the SR Ca2+-ATPase sequesters a significant amount of Ca2+ into the SR. 4. CPA slowed the rate of decay of [Ca2+]i following carbachol addition by > 50 %. Again, however, after a brief transient fall, decay was abolished when the Na+-Ca2+ exchanger and sarcolemmal Ca2+-ATPase were inhibited. 5. These data show that, although the SR Ca2+-ATPase contributes to the decay of [Ca2+]i, it cannot function effectively in the absence of Ca2+ removal from the cell. These data are discussed in the context of the superficial buffer barrier model in which Ca2+ is taken up into the SR and then released very close to sarcolemmal Ca2+ extrusion sites, i.e. the SR acting in series with the surface membrane extrusion mechanisms. We also suggest that the amount of filling of the SR influences the rate of Ca2+ removal. PMID- 10517809 TI - Direct actions of nitric oxide on rat neurohypophysial K+ channels. AB - 1. Nitric oxide (NO) has been shown to modulate neuropeptide secretion from the posterior pituitary. Here we show that NO activates large-conductance Ca2+ activated K+ (BK) channels in posterior pituitary nerve terminals. 2. NO, generated either by the photolysis of caged-NO or with chemical donors, irreversibly enhanced the component of whole-terminal K+ current due to BK channels and increased the activity of BK channels in excised patches. NO also inhibited the transient A-current. The time courses of these effects on K+ current were very different; activation of BK channels developed slowly over several minutes whereas inhibition of A-current immediately followed NO uncaging. 3. Activation of BK channels by NO occurred in the presence of guanylyl cyclase inhibitors and after removal of ATP or GTP from the pipette solution, suggesting a cGMP-independent signalling pathway. 4. The sulfhydryl alkylating agent N-ethyl maleimide (NEM) increased BK channel activity. Pretreatment with NEM occluded NO activation. 5. NO activation of BK channels occurred independently of voltage and cytoplasmic Ca2+ concentration. In addition, NO removed the strict Ca2+ requirement for channel activation, rendering channels highly active even at nanomolar Ca2+ levels. 6. These results suggest that NO, or a reactive nitrogen byproduct, chemically modifies nerve terminal BK channels or a closely associated protein and thereby produces an increase in channel activity. Such activation is likely to inhibit impulse activity in posterior pituitary nerve terminals and this may explain the inhibitory action of NO on secretion. PMID- 10517810 TI - Dihydropyridine-sensitive ion currents and charge movement in vesicles derived from frog skeletal muscle plasma membranes. AB - 1. Whole-cell voltage clamp experiments were performed in vesicles derived from frog skeletal muscle plasma membranes to characterize the electrophysiological properties of dihydropyridine (DHP) receptors. This preparation allows control of the composition of the internal medium and the recording of currents, without the influence of the sarcoplasmic reticulum (SR). 2. In solutions containing Ba2+, Bay K 8644-sensitive, L-type inward currents were recorded. Peak Ba2+ currents (IBa) averaged 3.0 microA microF-1 and inactivated in a voltage-dependent manner. Half-maximal steady-state inactivation occurred at -40 mV. No major facilitation of tail currents was observed. 3. The time course of activation of L-type Ca2+ channels was voltage dependent and 10 times faster than that in muscle fibres; the current density values were also much lower. 4. Lowering [Mg2+]i from 2 to 0.1 mM shifted the time to peak of IBa versus voltage relation by -13 mV. 5. In solutions that contained mostly impermeant ions, non-linear capacitive currents were recorded. Charge movement with properties resembling charge 1 was observed in polarized vesicles. The charge movement depended on voltage with Boltzmann parameters: Qmax (maximum charge), 45.6 nC microF-1; V (potential at which Q = 0.5 Qmax), -58.4 mV; and k (slope factor), 22. 3 mV. There was no indication of the presence of Qgamma (the 'hump' component of charge movement). 6. In depolarized vesicles, non-linear currents were observed during hyperpolarizing pulses. The currents produced an excessive charge during 'on' transients only. Charge during 'off' transients was linear from -180 to +60 mV. There was no evidence of the presence of charge 2. PMID- 10517811 TI - The role of Ca2+ feedback in shaping InsP3-evoked Ca2+ signals in mouse pancreatic acinar cells. AB - 1. Cytosolic Ca2+ has been proposed to act as both a positive and a negative feedback signal on the inositol trisphosphate (InsP3) receptor. However, it is unclear how this might affect the Ca2+ response in vivo. 2. Mouse pancreatic acinar cells were whole-cell patch clamped to record the Ca2+-dependent chloride (Cl(Ca)) current spikes and imaged to record the cytosolic Ca2+ spikes elicited by the injection of Ins(2,4,5)P3. Increasing concentrations of Ca2+ buffer (up to 200 microM EGTA or BAPTA) were associated with the appearance of steps in the current activation phase and a prevalence of smaller-amplitude Cl(Ca) spikes. Imaging experiments showed that with increased buffer the secretory pole cytosolic Ca2+ signal became fragmented and spatially discrete Ca2+ release events were observed. 3. At higher buffer concentrations (200-500 microM), increasing concentrations of EGTA increased spike frequency and reduced spike amplitude. In contrast, BAPTA decreased spike frequency and maintained large spike amplitudes. 4. We conclude that, during InsP3-evoked spiking, long-range Ca2+ feedback ( approximately 2-4 microm) shapes the rising phase of the Ca2+ signal by acting to co-ordinate discrete Ca2+ release events and short-range ( approximately 40 nm) Ca2+ feedback acts to inhibit further Ca2+ release. PMID- 10517812 TI - Low [ATP] and elevated [Mg2+] reduce depolarization-induced Ca2+ release in rat skinned skeletal muscle fibres. AB - 1. This study examined whether reduced [ATP], raised [Mg2+] and the presence of the metabolites AMP and inosine monophosphate (IMP) affected depolarization induced Ca2+ release from the sarcoplasmic reticulum (SR) in mechanically skinned skeletal muscle fibres of the rat. The amount of Ca2+ released was determined from the extent of SR Ca2+ depletion following a depolarization in the specified conditions with 2 mM free EGTA present to chelate released Ca2+. 2. In the presence of 8 mM total ATP and 1 mM free Mg2+, most of the SR Ca2+ could be released by a single (2-3 s) depolarization. Paired comparisons in the same fibres showed that raising the [Mg2+] from 1 to 3 mM reduced the total amount of Ca2+ released by a single depolarization by approximately 40 %. At 1 mM Mg2+, lowering the [ATP] to 0.5 mM did not cause a detectable change in the total amount of Ca2+ released, but when the release rate was reduced by the presence of 3 mM Mg2+, lowering the [ATP] to 0.5 mM resulted in a further ( approximately 20 %) reduction in the total amount of Ca2+ released. 3. At 1 mM Mg2+ and 0.5 mM ATP, neither the presence of 3 mM AMP alone nor 3 mM AMP plus 3 mM IMP caused a significant change in total Ca2+ release. Furthermore, at 1 mM Mg2+, the combined effect of lowering the [ATP] from 8 to 0.5 mM and simultaneously adding 3 mM AMP and 3 mM IMP did not significantly alter total Ca2+ release. However, when Ca2+ release was already reduced by the presence of 3 mM Mg2+ and 0.5 mM ATP (to approximately 50 %), addition of 3 mM AMP and 3 mM IMP significantly reduced the amount of Ca2+ released a further 2-fold. 4. These results show that depolarization-induced Ca2+ release in mammalian muscle fibres is modulated by the concentration of ATP and its metabolic products, as well as by the free [Mg2+]. Consequently, the (reversible) reduction in Ca2+ release occurring in a muscle fibre after prolonged exercise could result not only from raised [Mg2+] but also from a severe reduction in [ATP] locally near the Ca2+ release channels, with the accompanying build-up of AMP and IMP further exacerbating this effect. PMID- 10517813 TI - Differential effects of caffeine and perchlorate on excitation-contraction coupling in mammalian skeletal muscle. AB - 1. Enzymatically dissociated single muscle fibres of the rat were studied under voltage clamp conditions in a double Vaseline gap experimental chamber. Intramembrane charge movement and changes in intracellular calcium concentration ([Ca2+]i) were measured and the rate of calcium release (Rrel) from the sarcoplasmic reticulum (SR) was calculated. This enabled the determination of SR permeability and thus the estimation of the transfer function between intramembrane charge movement and SR permeability. 2. Perchlorate (3 mM) shifted the membrane potential dependence of intramembrane charge movement to more negative voltages without any effect on the steepness or on the maximal available charge. The drug increased SR permeability at every membrane potential but did not alter the peak-to-steady level ratio. It also increased the slope of the transfer function, indicating a more efficient coupling between the voltage sensors and the ryanodine receptors. 3. Caffeine (1 mM), on the other hand, increased SR permeability without altering the voltage dependence of intramembrane charge movement. It neither prolonged the depolarization-induced increase in [Ca2+]i at short pulse durations nor altered the time to peak of Rrel. The augmentation of SR permeability by the drug was more pronounced during the peak caffeine response than during its steady level. This was manifested in a leftward shift of the transfer function rather than an increase in its slope. 4. These observations indicate that perchlorate and caffeine alter the coupling between the voltage sensors and SR calcium release channels in mammalian skeletal muscle. They do not, however, share a common mechanism for enhancing the depolarization-induced release of calcium from the SR. PMID- 10517814 TI - Fast skeletal muscle troponin T increases the cooperativity of transgenic mouse cardiac muscle contraction. AB - 1. To investigate the functional significance of different troponin T (TnT) isoforms in the Ca2+ activation of muscle contraction, transgenic mice have been constructed with a chicken fast skeletal muscle TnT transgene driven by a cardiac alpha-myosin heavy chain gene promoter. 2. Cardiac muscle-specific expression of the fast skeletal muscle TnT has been obtained with significant myofibril incorporation. Expression of the endogenous cardiac muscle thin filament regulatory proteins, such as troponin I and tropomyosin, was not altered in the transgenic mouse heart, providing an authentic system for the functional characterization of TnT isoforms. 3. Cardiac muscle contractility was analysed for the force vs. Ca2+ relationship in skinned ventricular trabeculae of transgenic mice in comparison with wild-type litter-mates. The results showed unchanged pCa50 values (5.1 +/- 0.04 and 5.1 +/- 0.1, respectively) but significantly steeper slopes (the Hill coefficient was 2.0 +/- 0.2 vs. 1.0 +/- 0.2, P < 0.05). 4. The results demonstrate that the structural and functional variation of different TnT isoforms may contribute to the difference in responsiveness and overall cooperativity of the thin filament-based Ca2+ regulation between cardiac and skeletal muscles. PMID- 10517815 TI - Role of glutamate receptors in transmission of vagal cardiac input to neurones in the nucleus tractus solitarii in dogs. AB - 1. Vagal afferent input from cardiac mechanoreceptors excites neurones in the nucleus tractus solitarii (NTS), but discharge patterns evoked by physiological activation of pressure-sensitive cardiac mechanoreceptors have not been studied in vivo. The role of glutamate receptor subtypes in transmission of afferent activity to the NTS neurones has not been determined. The present study therefore has two aims: first, to characterise the discharge patterns of neurones in the NTS that receive pressure-sensitive vagal cardiac receptor input and second, to determine the roles of ionotropic glutamate receptor subtypes in the transmission of this putative cardiac mechanoreceptor-related activity to NTS neurones. 2. Pulse-synchronous activity of neurones in the NTS evoked by vagal afferent input was recorded extracellularly in an anaesthetised dog model using multibarrel glass electrodes, which allowed picoejection of the glutamate receptor antagonists NBQX or AP5 to block either non-NMDA or NMDA receptors, respectively, during the neuronal recording. Pressure sensitivity of the recorded neurones was examined by monitoring their response to a small increase in arterial blood pressure. Selective pressure activation of carotid sinus baroreceptors in an isolated sinus or selective denervation of aortic baroreceptors were used to test for convergent excitation of the neurones by arterial baroreceptors. 3. Pulse synchronous cardiac-related neuronal activity recorded from neurones in both the right and left NTS was eliminated following section of the left (n = 17) or right (n = 1) vagus nerves. No spontaneous, non-pulsatile activity was observed in these neurones before or after vagotomy. Activity transmitted via left vagal afferents was found to be sensitive to changes in arterial blood pressure. In these neurones, activity was blocked in 13 of 17 neurones by picoejection of NBQX, with the remainder requiring both NBQX and AP5. None of the cardiac-related neurones responded to activation of carotid baroreceptors or denervation of aortic baroreceptors, indicating no convergence of activity from carotid baroreceptors or aortic baroreceptors with pressure thresholds of approximately 130 mmHg or less. 4. The results suggest that vagal pressure-sensitive afferent input from cardiac mechanoreceptors is transmitted primarily by left vagal afferent fibres via non-NMDA receptors to neurones in both the ipsilateral and contralateral NTS. NMDA receptors were also found to have a role in the activation of a small subpopulation of neurones. PMID- 10517816 TI - A novel role for cyclic nucleotide-gated cation channels in lung liquid homeostasis in sheep. AB - 1. Sheep lungs were artificially perfused in situ with warmed whole oxygenated sheep blood. The airspaces of the lungs were filled with liquid containing an impermeant tracer, to allow measurement of the rate of net transepithelial liquid movement under various conditions. 2. Dichlorobenzamil (1.5 x 10-5 M), a blocker of cyclic nucleotide-gated cation channels, inhibited the resting absorption of lung liquid in sheep aged 6 months (n = 5) (from -36.47 +/- 4.62 to -4.36 +/- 5.27 ml h-1, means +/- s.e.m.; P < 0.005, paired t test). Amiloride (10-4 M), a blocker of epithelial sodium channels, had no additive effect to that of dichlorobenzamil. 3. In the lungs of sheep aged 6 months (n = 4), amiloride (10-4 M) partially inhibited the resting absorption of liquid (from -35.21 +/- 8.57 to 11.05 +/- 4.91 ml h-1; P < 0.05, one-tailed paired t test), and dichlorobenzamil (1.5 x 10-5 M) exerted an additive effect to that of amiloride resulting in secretion at +6.29 +/- 3.05 ml h-1 (P < 0. 01, paired t test). 4. In the lungs of sheep aged 6 weeks (n = 3), amiloride (10-4 M) also inhibited the resting absorption of liquid (from -26.36 +/- 14.05 to -5.17 +/- 8.27 ml h-1; P < 0.05, one-tailed paired t test); however, dichlorobenzamil (1.5 x 10-5 M) did not exert an additive effect to that of amiloride. 5. In the lungs of sheep aged 6 months (n = 4), amiloride (10-4 M) partially inhibited the resting absorption of liquid (from -35.70 +/- 8.58 to -6.79 +/- 4.28 ml h-1; P < 0.05, paired t test), and pimozide (1.5 x 10-4 M), another blocker of cyclic nucleotide-gated cation channels, also exerted an additive effect to that of amiloride, resulting in secretion of lung liquid at +15.36 +/- 9.14 ml h-1 (P < 0.05, paired t test). 6. We conclude that cyclic nucleotide-gated cation channels mediate a component of lung liquid absorption in sheep aged 6 months (but not in sheep aged 6 weeks), and that a mechanism for lung liquid secretion (present in fetuses) is retained at 6 months of age. PMID- 10517817 TI - Effect of renal perfusion pressure on renal function, renin release and renin and angiotensinogen gene expression in rats. AB - 1. A study was undertaken to examine the influence of acute renal perfusion pressure (RPP) reduction on renin release, renal renin and angiotensinogen gene expression and the role played by angiotensin II in these responses. 2. In chloralose-urethane anaesthetised rats, reduction of RPP to 60 mmHg for 3 h in vehicle or losartan-treated (5 days at 10 mg kg-1 bis in die (b.i.d.)) rats decreased renal blood flow by 46 and 29 % (both P < 0.001), respectively, glomerular filtration rate by 45 and 57 % (both P < 0.001), respectively, and sodium excretion by 96 and 98 % (both P < 0.01). 3. Chloralose-urethane anaesthesia and surgery caused a rise in plasma renin activity but was associated with a suppression of renal renin (50 %, P < 0.01) and angiotensinogen (40 %, P < 0.05) gene expression. Following reduction of RPP to 60 mmHg for 3 h, plasma renin activity was increased more than 7-fold (P < 0.001) and renal renin gene expression about 2-fold (P < 0.05). 4. Chronic (5 days) blockade of angiotensin II receptors with losartan elevated plasma renin activity some 29-fold (P < 0.001) and caused a marked increase (30-fold, P < 0.05) in renal renin gene expression, compatible with angiotensin II exerting a negative feedback control on renin release and gene expression. Reduction of RPP to 60 mmHg for 3 h in these animals had little effect on renal renin gene expression. 5. From these findings it can be concluded that (a) chloralose-urethane anaesthesia and surgery had a stimulatory effect on renin release but suppressed basal levels of renal renin and angiotensinogen gene expression; (b) acute reduction of RPP for 3 h could stimulate renin gene expression in the renin producing cells; and (c) the negative feedback control of angiotensin II on renin release and synthesis which was evident following chronic losartan treatment was not apparent during short term reduction of RPP. PMID- 10517818 TI - Specificity of synergistic coronary flow enhancement by adenosine and pulsatile perfusion in the dog. AB - 1. Coronary flow elevation from enhanced perfusion pulsatility is synergistically amplified by adenosine. This study determined the specificity of this interaction and its potential mechanisms. 2. Mean and phasic coronary flow responses to increasing pulsatile perfusion were assessed in anaesthetized dogs, with the anterior descending coronary artery servoperfused to regulate real-time physiological flow pulsatility at constant mean pressure. Pulsatility was varied between 40 and 100 mmHg. Hearts ejected into the native aorta whilst maintaining stable loading. 3. Increasing pulsatility elevated mean coronary flow +11.5 +/- 1.7 % under basal conditions. Co-infusion of adenosine sufficient to raise baseline flow 66 % markedly amplified this pulsatile perfusion response (+82. 6 +/- 14.3 % increase in mean flow above adenosine baseline), due to a leftward shift of the adenosine-coronary flow response curve at higher pulsatility. Flow augmentation with pulsatility was not linked to higher regional oxygen consumption, supporting direct rather than metabolically driven mechanisms. 4. Neither bradykinin, acetylcholine nor verapamil reproduced the synergistic amplification of mean flow by adenosine and higher pulsatility, despite being administered at doses matching basal flow change with adenosine. 5. ATP-sensitive potassium (KATP) activation (pinacidil) amplified the pulse-flow response 3-fold, although this remained significantly less than with adenosine. Co-administration of the phospholipase A2 inhibitor quinacrine virtually eliminated adenosine induced vasodilatation, yet synergistic interaction between adenosine and pulse perfusion persisted, albeit at a reduced level. 6. Thus, adenosine and perfusion pulsatility specifically interact to enhance coronary flow. This synergy is partially explained by KATP agonist action and additional non-flow-dependent mechanisms, and may be important for modulating flow reserve during exercise or other high output states where increased flow demand and higher perfusion pulsatility typically co-exist. PMID- 10517819 TI - Modification of activity-dependent increases in cerebellar blood flow by extracellular potassium in anaesthetized rats. AB - 1. The hypothesis that potassium ions mediate activity-dependent increases of cerebral blood flow was examined in rat cerebellar cortex using ion-selective microelectrodes and laser-Doppler flowmetry. Increases of cerebellar blood flow (CeBF) and extracellular potassium concentration ([K+]o) were evoked by stimulation of parallel fibres and climbing fibres, and by microinjection of KCl into the cortex. 2. For parallel fibre stimulation, there was a maximal increase in [K+]o to 6.3 +/- 0.5 mM and in CeBF of 122 +/- 11 %. Climbing fibre stimulation gave a maximal increase in [K+]o to 4.4 +/- 0.2 mM and in CeBF of 157 +/- 20 %. This indicates different maxima for [K+]o and CeBF, dependent on the afferent system activated. 3. [K+]o and CeBF responses evoked by parallel or climbing fibre stimulation increased rapidly at the onset of stimulation, but exhibited different time courses during the remainder of the stimulation period and during return to baseline. 4. Microinjections of KCl into the cortex increased [K+]o to levels comparable to those evoked by parallel fibre stimulation. The corresponding CeBF increases were the same as, or smaller than, for parallel fibre stimulation, and much smaller than for climbing fibre stimulation. This suggests that mediators other than [K+]o are important for activity-dependent cerebral blood flow increases. 5. The present study showed that increased [K+]o is involved in CeBF regulation in the parallel fibre system, but is of limited importance for CeBF regulation in the climbing fibre system. The hypothesis that K+ is a major mediator of activity-dependent blood flow increases is probably not generally applicable to all brain regions and all types of neuronal stimulation. PMID- 10517820 TI - Intercostal expiratory activity in an in vitro brainstem-spinal cord-rib preparation from the neonatal rat. AB - 1. We examined whether expiratory activity can be observed when central chemoreceptors are activated by a decrement in the extracellular pH in an isolated brainstem-spinal cord-rib preparation from 0- to 3-day-old rats. Expiratory activity was defined as the burst activity that occurs in an internal intercostal muscle (IIM) during the silent period between the periodic inspiratory bursts in the C4 ventral root (which contains phrenic motor axons). 2. During perfusion with modified Krebs solution (26 mM HCO3-, 5 % CO2, pH 7.4), there was no consistent activity in IIM, though rhythmic inspiratory motor activity always appeared in the C4 ventral root. 3. When the pH of the perfusate was lowered from about 7.4 to 7.1 by reducing [HCO3-] from 26 to 10 mM, the frequency of the C4 inspiratory rhythm increased, and rhythmic activity appeared in IIM. In most cases, the rhythmic burst in IIM started just after the cessation of the C4 inspiratory burst and coincided with movement of the ribs in a caudal direction. This intercostal expiratory burst was limited to the first half of the expiratory phase. 4. The coordinated reciprocal motor activity between the C4 ventral root and IIM changed to a largely overlapping pattern when strychnine (5 10 microM), a glycine receptor antagonist, was added to the perfusate. 5. These results suggest (i) that the neuronal mechanisms responsible for expiratory motor activity are preserved in this in vitro preparation and (ii) that the glycinergic inhibitory system plays an important role in the coordination between inspiratory and expiratory motor activity during respiration. PMID- 10517821 TI - Transient, reversible apnoea following ablation of the pre-Botzinger complex in rats. AB - 1. In some anaesthetized preparations, eupnoea is eliminated following a blockade or destruction of neurons in a rostral medullary pre-Botzinger complex. 2. Neurons in this region might underlie the neurogenesis of eupnoea, or be the source of an input which is necessary for eupnoea to be expressed. If the latter, any apnoea following ablation of the pre-Botzinger complex might be reversed by an augmentation in 'tonic input.' Contrariwise, this apnoea should be permanent if the neuronal activities of the pre- Botzinger complex are an exclusive generator of the eupnoeic rhythm. 3. Decerebrate, vagotomized, paralysed and ventilated adult rats were studied. Efferent activity of the phrenic nerve was recorded as an index of ventilatory activity. 4. Blockade or destruction of neuronal activities of the pre-Botzinger complex by unilateral and/or bilateral injections of muscimol or kainic acid eliminated eupnoea only transiently. Eupnoea returned following activation of the peripheral chemoreceptors and spontaneously over time. 5. Results do not support the concept that neuronal activities of the pre-Botzinger complex play an exclusive role in the neurogenesis of eupnoea in vivo. Rather, these neuronal activities appear to provide a tonic input to the ponto-medullary circuit which generates eupnoea and/or appear to be one component of this circuit. PMID- 10517822 TI - Proteoid roots. Physiology and development PMID- 10517823 TI - Plant volatiles as a defense against insect herbivores PMID- 10517824 TI - Biosynthesis and immunolocalization of Lewis a-containing N-glycans in the plant cell. AB - We recently demonstrated the presence of a new asparagine-linked complex glycan on plant glycoproteins that harbors the Lewis a (Lea), or Galbeta(1-3)[Fucalpha(1 4)]GlcNAc, epitope, which in mammalian cells plays an important role in cell-to cell recognition. Here we show that the monoclonal antibody JIM 84, which is widely used as a Golgi marker in light and electron microscopy of plant cells, is specific for the Lea antigen. This antigen is present on glycoproteins of a number of flowering and non-flowering plants, but is less apparent in the Cruciferae, the family that includes Arabidopsis. Lea-containing oligosaccharides are found in the Golgi apparatus, and our immunocytochemical experiments suggest that it is synthesized in the trans-most part of the Golgi apparatus. Lea epitopes are abundantly present on extracellular glycoproteins, either soluble or membrane bound, but are never observed on vacuolar glycoproteins. Double-labeling experiments suggest that vacuolar glycoproteins do not bypass the late Golgi compartments where Lea is built, and that the absence of the Lea epitope from vacuolar glycoproteins is probably the result of its degradation by glycosidases en route to or after arrival in the vacuole. PMID- 10517825 TI - A point mutation in the ethylene-inducing xylanase elicitor inhibits the beta-1-4 endoxylanase activity but not the elicitation activity. AB - Ethylene-inducing xylanase (EIX) elicits plant defense responses in certain tobacco (Nicotiana tabacum) and tomato cultivars in addition to its xylan degradation activity. It is not clear, however, whether elicitation occurs by cell wall fragments released by the enzymatic activity or by the xylanase protein interacting directly with the plant cells. We cloned the gene encoding EIX protein and overexpressed it in insect cells. To determine the relationship between the two activities, substitution of amino acids in the xylanase active site was performed. Substitution at glutamic acid-86 or -177 with glutamine (Gln), aspartic acid (Asp), or glycine (Gly) inhibited the beta-1-4-endoxylanase activity. Mutants having Asp-86 or Gln-177 also lost the ability to induce the hypersensitive response and ethylene biosynthesis. However, mutants having Gln 86, Gly-86, Asp-177, or Gly-177 retained ability to induce ethylene biosynthesis and the hypersensitive response. Our data show that the xylanase activity of EIX elicitor can be separated from the elicitation process, as some of the mutants lack the former but retain the latter. PMID- 10517827 TI - Changes in cell wall polysaccharides of green bean pods during development. AB - The changes in cell wall polysaccharides and selected cell wall-modifying enzymes were studied during the development of green bean (Phaseolus vulgaris L.) pods. An overall increase of cell wall material on a dry-weight basis was observed during pod development. Major changes were detected in the pectic polymers. Young, exponentially growing cell walls contained large amounts of neutral, sugar rich pectic polymers (rhamnogalacturonan), which were water insoluble and relatively tightly connected to the cell wall. During elongation, more galactose rich pectic polymers were deposited into the cell wall. In addition, the level of branched rhamnogalacturonan remained constant, while the level of linear homogalacturonan steadily increased. During maturation of the pods, galactose rich pectic polymers were degraded, while the accumulation of soluble homogalacturonan continued. During senescence there was an increase in the amount of ionically complexed pectins, mainly at the expense of freely soluble pectins. The most abundant of the enzymes tested for was pectin methylesterase. Peroxidase, beta-galactosidase, and alpha-arabinosidase were also detected in appreciable amounts. Polygalacturonase was detected only in very small amounts throughout development. The relationship between endogenous enzyme levels and the properties of cell wall polymers is discussed with respect to cell wall synthesis and degradation. PMID- 10517826 TI - Overexpression of a gene that encodes the first enzyme in the biosynthesis of asparagine-linked glycans makes plants resistant to tunicamycin and obviates the tunicamycin-induced unfolded protein response. AB - The cytotoxic drug tunicamycin kills cells because it is a specific inhibitor of UDP-N-acetylglucosamine:dolichol phosphate N-acetylglucosamine-1-P transferase (GPT), an enzyme that catalyzes the initial step of the biosynthesis of dolichol linked oligosaccharides. In the presence of tunicamycin, asparagine-linked glycoproteins made in the endoplasmic reticulum are not glycosylated with N linked glycans, and therefore may not fold correctly. Such proteins may be targeted for breakdown. Cells that are treated with tunicamycin normally experience an unfolded protein response and induce genes that encode endoplasmic reticulum chaperones such as the binding protein (BiP). We isolated a cDNA clone for Arabidopsis GPT and overexpressed it in Arabidopsis. The transgenic plants have a 10-fold higher level of GPT activity and are resistant to 1 microg/mL tunicamycin, a concentration that kills control plants. Transgenic plants grown in the presence of tunicamycin have N-glycosylated proteins and the drug does not induce BiP mRNA levels as it does in control plants. BiP mRNA levels are highly induced in both control and GPT-expressing plants by azetidine-2-carboxylate. These observations suggest that excess GPT activity obviates the normal unfolded protein response that cells experience when exposed to tunicamycin. PMID- 10517828 TI - Cloning and molecular analyses of a gibberellin 20-oxidase gene expressed specifically in developing seeds of watermelon. AB - To understand the biosynthesis and functional role of gibberellins (GAs) in developing seeds, we isolated Cv20ox, a cDNA clone from watermelon (Citrullus lanatus) that shows significant amino acid homology with GA 20-oxidases. The complementary DNA clone was expressed in Escherichia coli as a fusion protein, which oxidized GA(12) at C-20 to the C(19) compound GA(9), a precursor of bioactive GAs. RNA-blot analysis showed that the Cv20ox gene was expressed specifically in developing seeds. The gene was strongly expressed in the integument tissues, and it was also expressed weakly in inner seed tissues. In parthenocarpic fruits induced by 1-(2-chloro-4-pyridyl)-3-phenylurea treatment, the expression pattern of Cv20ox did not change, indicating that the GA 20 oxidase gene is expressed primarily in the maternal cells of developing seeds. The promoter of Cv20ox was isolated and fused to the beta-glucuronidase (GUS) gene. In a transient expression system, beta-glucuronidase staining was detectable only in the integument tissues of developing watermelon seeds. PMID- 10517829 TI - The mur4 mutant of arabidopsis is partially defective in the de novo synthesis of uridine diphospho L-arabinose. AB - To obtain information on the synthesis and function of arabinosylated glycans, the mur4 mutant of Arabidopsis was characterized. This mutation leads to a 50% reduction in the monosaccharide L-arabinose in most organs and affects arabinose containing pectic cell wall polysaccharides and arabinogalactan proteins. Feeding L-arabinose to mur4 plants restores the cell wall composition to wild-type levels, suggesting a partial defect in the de novo synthesis of UDP-L-arabinose, the activated sugar used by arabinosyltransferases. The defect was traced to the conversion of UDP-D-xylose to UDP-L-arabinose in the microsome fraction of leaf material, indicating that mur4 plants are defective in a membrane-bound UDP-D xylose 4-epimerase. PMID- 10517831 TI - The starch-debranching enzymes isoamylase and pullulanase are both involved in amylopectin biosynthesis in rice endosperm AB - The activities of the two types of starch debranching enzymes, isoamylase and pullulanase, were greatly reduced in endosperms of allelic sugary-1 mutants of rice (Oryza sativa), with the decrease more pronounced for isoamylase than for pullulanase. However, the decrease in isoamylase activity was not related to the magnitude of the sugary phenotype (the proportion of the phytoglycogen region of the endosperm), as observed with pullulanase. In the moderately mutated line EM 5, the pullulanase activity was markedly lower in the phytoglycogen region than in the starch region, and isoamylase activity was extremely low or completely lost in the whole endosperm tissue. These results suggest that both debranching enzymes are involved in amylopectin biosynthesis in rice endosperm. We presume that isoamylase plays a predominant role in amylopectin synthesis, but pullulanase is also essential or can compensate for the role of isoamylase in the construction of the amylopectin multiple-cluster structure. It is highly possible that isoamylase was modified in some sugary-1 mutants such as EM-273 and EM-5, since it was present in significant and trace amounts, respectively, in these mutants but was apparently inactive. The results show that the Sugary-1 gene encodes the isoamylase gene of the rice genome. PMID- 10517830 TI - The enzymatic activity of fungal xylanase is not necessary for its elicitor activity. AB - Fungal xylanases from Trichoderma spp. are potent elicitors of defense responses in various plants. To determine whether enzymatic activity is necessary for elicitor activity, we used site-directed mutagenesis to reduce the catalytic activity of xylanase II from Trichoderma reesei. For this, the glutamic acid residue at position 210, which is part of the active center in this family of enzymes, was changed to either aspartic acid (E210D) or serine (E210S). Wild-type and mutated forms of xylanase II were expressed in yeast cells and purified to homogeneity. Compared with the wild-type form of xylanase II, E210D had >100-fold and E210S 1,000-fold lower enzymatic activity. In contrast, these mutated forms showed no comparable drop in elicitor activity. They fully stimulated medium alkalinization and ethylene biosynthesis in suspension-cultured tomato (Lycopersicon esculentum) cells, as well as hypersensitive necrosis in leaves of tomato and tobacco (Nicotiana tabacum) plants. These results provide direct evidence that enzyme activity is not necessary for elicitor activity of fungal xylanase. PMID- 10517832 TI - A splice site mutant of maize activates cryptic splice sites, elicits intron inclusion and exon exclusion, and permits branch point elucidation. AB - DNA sequence analysis of the bt2-7503 mutant allele of the maize brittle-2 gene revealed a point mutation in the 5' terminal sequence of intron 3 changing GT to AT. This lesion completely abolishes use of this splice site, activates two cryptic splice sites, and alters the splicing pattern from extant splice sites. One activated donor site, located nine nt 5' to the normal splice donor site, begins with the dinucleotide GC. While non-consensus, this sequence still permits both trans-esterification reactions of pre-mRNA splicing. A second cryptic site located 23 nt 5' to the normal splice site and beginning with GA, undergoes the first trans-esterification reaction leading to lariat formation, but lacks the ability to participate in the second reaction. Accumulation of this splicing intermediate and use of an innovative reverse transcriptase-polymerase chain reaction technique (J. Vogel, R.H. Wolfgang, T. Borner [1997] Nucleic Acids Res 25: 2030-2031) led to the identification of 3' intron sequences needed for lariat formation. In most splicing reactions, neither cryptic site is recognized. Most mature transcripts include intron 3, while the second most frequent class lacks exon 3. Traditionally, the former class of transcripts is taken as evidence for the intron definition of splicing, while the latter class has given credence to the exon definition of splicing. PMID- 10517833 TI - Expression of a polygalacturonase associated with tomato seed germination. AB - Radicle protrusion from tomato (Lycopersicon esculentum Mill.) seeds to complete germination requires weakening of the endosperm tissue opposite the radicle tip. In common with other cell wall disassembly processes in plants, polygalacturonases (PGs) may be involved. Only calcium-dependent exo-PG activity was detected in tomato seed protein extracts. Chromatographic profiles of a partially acid-hydrolyzed fraction of polygalacturonic acid further digested with seed extract were consistent with the presence of only calcium-dependent exo-PG activity. In addition, a transcript encoding a previously unknown PG was detected prior to the completion of germination. The mRNA, produced from a gene (LeXPG1) estimated by Southern analysis to be represented once in the genome, was also present in flowers (anthers) and in lower amounts in roots and stems. LeXPG1 mRNA abundance was low during seed development, increased during imbibition, and was even greater in seeds that had completed germination. Expression of LeXPG1 during germination predominates in the endosperm cap and radicle tip, and in the radicle appears as a distinct band possibly associated with vascular tissue differentiation. We suggest that PG is involved in cell wall loosening of the endosperm necessary for radicle protrusion from tomato seeds and in subsequent embryo and seedling growth. PMID- 10517834 TI - Arabidopsis alcohol dehydrogenase expression in both shoots and roots is conditioned by root growth environment. AB - It is widely accepted that the Arabidopsis Adh (alcohol dehydrogenase) gene is constitutively expressed at low levels in the roots of young plants grown on agar media, and that the expression level is greatly induced by anoxic or hypoxic stresses. We questioned whether the agar medium itself created an anaerobic environment for the roots upon their growing into the gel. beta-Glucuronidase (GUS) expression driven by the Adh promoter was examined by growing transgenic Arabidopsis plants in different growing systems. Whereas roots grown on horizontal-positioned plates showed high Adh/GUS expression levels, roots from vertical-positioned plates had no Adh/GUS expression. Additional results indicate that growth on vertical plates closely mimics the Adh/GUS expression observed for soil-grown seedlings, and that growth on horizontal plates results in induction of high Adh/GUS expression that is consistent with hypoxic or anoxic conditions within the agar of the root zone. Adh/GUS expression in the shoot apex is also highly induced by root penetration of the agar medium. This induction of Adh/GUS in shoot apex and roots is due, at least in part, to mechanisms involving Ca2+ signal transduction. PMID- 10517835 TI - Genetic analysis of growth-regulator-induced parthenocarpy in Arabidopsis. AB - In Arabidopsis, seedless silique development or parthenocarpy can be induced by the application of various plant growth regulators (PGRs) to unfertilized pistils. Ecotype-specific responses were observed in the Arabidopsis ecotypes Columbia and Landsberg relative to the type of PGR and level applied. The parthenocarpic response was greatest in ecotype Landsberg, and comparisons of fruit growth and morphology were studied primarily in this ecotype. Gibberellic acid application (10 micromol pistil(-1)) caused development similar to that in pollinated pistils, while benzyladenine (1 micromol pistil(-1)) and naphthylacetic acid (10 micromol pistil(-1)) treatment produced shorter siliques. Naphthylacetic acid primarily modified mesocarp cell expansion. Arabidopsis mutants were employed to examine potential dependencies on gibberellin biosynthesis (ga1-3, ga4-1, and ga5-1) and perception (spy-4 and gai) during parthenocarpic silique development. Emasculated spy-4 pistils were neither obviously parthenocarpic nor deficient in PGR perception. By contrast, emasculated gai mutants did not produce parthenocarpic siliques following gibberellic acid application, but silique development occurred following pollination or application of auxin and cytokinin. Pollinated gai siliques had decreased cell numbers and morphologically resembled auxin-induced parthenocarpic siliques. This shows that a number of independent and possibly redundant pathways can direct hormone-induced parthenocarpy, and that endogenous gibberellins play a role in regulating cell expansion and promoting cell division in carpels. PMID- 10517837 TI - Auxin and cytokinin have opposite effects on amyloplast development and the expression of starch synthesis genes in cultured bright yellow-2 tobacco cells. AB - In cultured Bright Yellow-2 (BY-2) tobacco (Nicotiana tabacum) cells, the depletion of auxin (2,4-dichlorophenoxyacetic acid) in the culture medium induces the accumulation of starch. This is accelerated by the addition of cytokinin (benzyladenine). Light and electron microscopic observations revealed that this amyloplast formation involves drastic changes in plastid morphology. The effects of auxin and cytokinin on amyloplast development were investigated by adding auxin or cytokinin to cells grown in a hormone-free culture. Auxin repressed amyloplast development, whereas cytokinin accelerated starch accumulation regardless of the timing of hormone addition. RNA gel-blot analysis revealed that the accumulation of the ADP-glucose pyrophosphorylase small subunit gene (AgpS), granule-bound starch synthase, and starch branching enzyme transcripts were also affected by hormonal conditions. High levels of AgpS, granule-bound starch synthase, and starch branching enzyme transcripts accumulated in amyloplast developing cells grown in auxin-depleted conditions. Furthermore, the addition of auxin to the cells cultured in hormone-free medium reduced the level of AgpS transcripts, whereas the addition of cytokinin increased it, irrespective of the timing of hormone addition. These results suggest that auxin and cytokinin exert opposite effects on amyloplast development by regulating the expression of the genes required for starch biosynthesis. PMID- 10517836 TI - Glucose polyester biosynthesis. Purification and characterization of a glucose acyltransferase. AB - Glandular trichomes of the wild tomato species Lycopersicon pennellii secrete 2,3,4-O-tri-acyl-glucose (-Glc), which contributes to insect resistance. A Glc acyltransferase catalyzes the formation of diacyl-Glc by disproportionating two equivalents of 1-O-acyl-beta-Glc, a high-energy molecule formed by a UDP-Glc dependent reaction. The acyltransferase was purified 4,900-fold from L. pennellii leaves by polyethylene glycol fractionation, diethylaminoethyl chromatography, concanavalin A affinity chromatography, and chromatofocusing. The acyltransferase possesses an isoelectric point of 4.8, a relative molecular mass around 110 kD, and is composed of 34- and 24-kD polypeptides as a heterotetramer. The 34- and 24 kD proteins were partially sequenced. The purified enzyme catalyzes both the disproportionation of 1-O-acyl-beta-Glcs to generate 1,2-di-O-acyl-beta-Glc and anomeric acyl exchange between 1-O-acyl-beta-Glc and Glc. PMID- 10517838 TI - The role of photosynthetic electron transport in the oxidative degradation of chloroplastic glutamine synthetase AB - The stability of chloroplastic glutamine synthetase (GS; EC 6.3.1.2) was investigated under photooxidative stress using wheat (Triticum aestivum L.) leaves, chloroplasts, and chloroplast lysates. Illuminated seedlings sprayed with the superoxide radical (O-(2)) propagator methyl viologen showed rapid GS decline dependent on MV concentration and exposure time. Degradation products of approximately 39 and 31 kD were detected when chloroplast lysates containing both stroma and thylakoids were illuminated in the presence of MV or H(2)O(2). In all cases, GS cleavage was prevented by the addition of the electron transport inhibitor 3-(3, 4-dichlorophenyl)-1,1-dimethylurea. Full protection against degradation could also be obtained by the incorporation of chelators or antioxidant enzymes. Maximal rates of degradation required the presence of transition metals and reducing compounds such as NADPH or dithiothreitol. Similar patterns of GS cleavage were obtained when seedlings were exposed to high doses of irradiation. The results indicate that chloroplastic GS is extremely prone to oxidative cleavage, and that reduced transition metals, presumably resulting from the destruction of iron-sulfur clusters by light-generated O-(2), play a crucial role in the degradation process. The physiological implications of GS lability to oxidative stress are discussed. PMID- 10517839 TI - Cloning, expression, and molecular characterization of a small pea gene family regulated by low levels of ultraviolet B radiation and other stresses. AB - A pea (Pisum sativum) DNA fragment (termed MB3) was isolated by differential display of cDNAs obtained from total leaf RNA of ultraviolet B (UV-B) radiation treated plants. Longer cDNAs were cloned by rapid amplification of cDNA ends in the 3' to 5' direction. Three different, but very similar, cDNAs were cloned, sadA, sadB, and sadC, the major difference between them being a 36-bp deletion in the coding region of sadB. Southern blotting confirmed the occurrence of at least three genes in the pea genome. Database comparisons of the SAD protein sequences revealed high identity (46%) and similarity (77%) with a putative tomato (Lycopersicon esculentum) short-chain alcohol dehydrogenase. Very low levels of UV-B radiation (the biologically effective radiation normalized to 300 nm = 0.08 W m(-2)) was shown to up-regulate expression, a dose considerably lower than that needed to induce expression of the well-known UV-B defensive chalcone synthase and phenylalanine ammonia lyase genes. RNase protection assay revealed that primarily sadA and sadC mRNA accumulation was enhanced by UV-B. In addition to UV B irradiation, ozone fumigation, wounding, aluminum stress, and salt stress induced increased transcript levels of the sad genes in pea. PMID- 10517840 TI - Characterization of stomatal closure caused by ultraviolet-B radiation AB - The effects of ultraviolet-B (UV-B) radiation on stomatal conductance (g(s)) in pea (Pisum sativum L.), commelina (Commelina communis L.), and oilseed rape (Brassica napus L.) plants were investigated. Plants were grown in a greenhouse either with three different high ratios of UV-B to photosynthetically active radiation or with no UV-B radiation. Pea plants grown in the highest UV-B radiation (0.63 W m(-2)) exhibited a substantial decrease of adaxial and abaxial g(s) (approximately 80% and 40%, respectively). With growth in 0.30 W m(-2) of UV B adaxial g(s) was decreased by 23%, with no effect on abaxial g(s), and lower UV B irradiance of 0.21 W m(-2) had no effect on either surface. Although abaxial g(s) increased when leaves were turned over in control plants, it did not in plants grown with the highest UV-B. Adaxial g(s) in commelina and oilseed rape also decreased on exposure to high UV-B (0.63 W m(-2)). For previously unexposed pea plants the time course of the effect of UV-B on g(s) was slow, with a lag of approximately 4 h, and a time constant of approximately 3 h. We conclude that there is a direct effect of UV-B on stomata in addition to that caused by changes in mesophyll photosynthesis. PMID- 10517841 TI - Successive use of non-host plant proteinase inhibitors required for effective inhibition of helicoverpa armigera gut proteinases and larval growth AB - We report on the efficacy of proteinase inhibitors (PIs) from three host plants (chickpea [Cicer arietinum], pigeonpea [Cajanus cajan], and cotton [Gossypium arboreum]) and three non-host (groundnut [Arachis hypogea], winged bean [Psophocarpus tetragonolobus], and potato [Solanum tuberosum]) in retarding the growth of Helicoverpa armigera larvae, a devastating pest of important crop plants. Enzyme assays and electrophoretic analysis of interaction of H. armigera gut proteinases (HGPs) with PIs revealed that non-host PIs inhibited HGP activity efficiently whereas host PIs were ineffective. In the electrophoretic assay, trypsin inhibitor activity bands were detected in all of the host and non-host plants, but HGP inhibitor activity bands were present only in non-host plants (except cotton in the host plant group). H. armigera larvae reared on a diet containing non-host PIs showed growth retardation, a reduction in total and trypsin-like proteinase activity, and the production of inhibitor-insensitive proteinases. Electrophoretic analysis of PI-induced HGP showed differential regulation of proteinase isoforms. Interestingly, HGP activity induced in response to dietary potato PI-II was inhibited by winged bean PIs. The optimized combination of potato PI-II and winged bean PIs identified in the present study and their proposed successive use has potential in developing H. armigera resistant transgenic plants. PMID- 10517842 TI - Phosphatidylinositol 4-kinase associated with spinach plasma membranes. Isolation and characterization of two distinct forms AB - Highly purified plasma membranes from spinach (Spinacia oleracea L.) leaves contained phosphatidylinositol (PtdIns) kinase activity that was firmly associated with the membrane. The enzyme was solubilized by detergent treatment (2% [w/v] Triton X-100) and purified by heparin-Sepharose and Q-Sepharose chromatography. Two enzymically active fractions, QI and QII, both exhibiting PtdIns 4-kinase activity, were resolved and purified 100- to 300-fold over the plasma membrane. QI and QII shared similar high apparent K(m) values for ATP (approximately 0.45 mM) and PtdIns (approximately 0.2 mM) and were insensitive to inhibition by adenosine. While Mg(2+) was the preferred divalent cation, Mn(2+) could partly substitute in the reaction catalyzed by the QII enzyme but not in that catalyzed by QI. Mn(2+) acted synergistically with suboptimal Mg(2+) concentrations to activate not only the QII enzyme, but also to some extent QI. Both enzymes were inhibited by millimolar concentrations of Ca(2+) and micromolar concentrations of wortmannin. The apparent molecular mass for QI was 120 kD, which was determined by SDS-PAGE and western blotting using an antibody against a peptide unique for lipid kinases and the binding of (3)H-wortmannin, and for QII 65 kD as determined by immunodetection and renaturation of PtdIns kinase activity in the 65-kD region of polyacrylamide gels. PMID- 10517843 TI - Rapid and systemic accumulation of chloroplast mRNA-binding protein transcripts after flame stimulus in tomato. AB - It has been shown that tomato (Lycopersicon esculentum) plants respond to flame wounding and electrical stimulation by a rapid (15 min) and systemic up regulation of proteinase inhibitor (pin) genes. To find other genes having a similar expression pattern, we used subtractive cDNA screening between flamed and control plants to select clones up-regulated by flame wounding. We report the characterization of one of them, a chloroplast mRNA-binding protein encoded by a single gene and expressed preferentially in the leaves. Systemic gene expression in response to flaming in the youngest terminal leaf exhibited three distinct phases: a rapid and transient increase (5-15 min) in transcript accumulation, a decline to basal levels (15-45 min), and then a second, more prolonged increase (60-90 min). In contrast, after a mechanical wound the rapid, transient increase (5 min) was followed by a rapid decline to basal levels but no later, prolonged accumulation. In the petiole, the initial flame-wound-evoked transient increase (15 min) was followed by a continuous decline for 3 h. The nature of the wound signal(s) causing such rapid changes in transcript abundance is discussed in relation to electrical signaling, which has recently been implicated in plant responses to wounding. PMID- 10517844 TI - Biochemical and immunocytochemical characterization of two types of myosins in cultured tobacco bright yellow-2 cells AB - We have isolated a myosin (referred to as 170-kD myosin) from lily pollen tubes, which consists of 170-kD heavy chain and calmodulin (CaM) light chain and is responsible for cytoplasmic streaming. A 170-kD polypeptide that has similar antigenicity to the 170-kD myosin heavy chain of lily pollen tubes was also present in cultured tobacco (Nicotiana tabacum) Bright Yellow-2 (BY-2) cells, and possessed the ability to interact with F-actin in an ATP-dependent manner. In addition to this myosin, we identified biochemically another kind of myosin in BY 2 cells. This myosin consisted of a CaM light chain and a 175-kD heavy chain with antigenicity different from the 170-kD myosin heavy chain. In the present study, we referred to this myosin as 175-kD myosin. This myosin was able to translocate rhodamine-phalloidin (RP)-labeled F-actin at an average velocity of about 9 &mgr;m/s in the motility assay in vitro. In contrast, the sliding velocity of RP labeled F-actin translocated by fractions containing the 170-kD myosin was 3 to 4 &mgr;m/s. The velocity of cytoplasmic streaming in living BY-2 cells ranged from 2 to 9 &mgr;m/s. The motile activity of 175-kD myosin in vitro was inhibited by Ca(2+) at concentrations higher than 10(-6) M. Immunoblot analyses using an antiserum against the heavy chain of 170- or 175-kD myosin revealed that in tobacco plants, the 175-kD myosin was expressed in leaf, stem, and root, but not in germinating pollen, while 170-kD myosin was present in all of these plant parts and in germinating pollen. These results suggest that the two types of myosins, 170 and 175 kD, presumably participate in cytoplasmic streaming in BY-2 cells and other somatic cells of tobacco plants. PMID- 10517845 TI - Ascorbate biosynthesis in Arabidopsis cell suspension culture. AB - The biosynthesis of L-ascorbic acid (L-AA) in an Arabidopsis (L.) Heynh. cell suspension culture was studied by quantifying the effects of incubation with a range of potential biosynthetic precursors, analogs, and inhibitors on the intracellular levels of reduced and oxidized forms of L-AA. Our results support the recently published biosynthetic pathway of L-AA from L-galactose (G.L. Wheeler, M.A. Jones, N. Smirnoff [1998] Nature 393: 365-369), but suggest that Arabidopsis cell suspension culture simultaneously contains two other routes leading to L-AA. The possible physiological significance of these alternate routes is discussed. PMID- 10517846 TI - Signaling events leading to crassulacean acid metabolism induction in the common ice plant AB - A rapid, semiquantitative reverse transcriptase-polymerase chain reaction assay was developed to investigate signal transduction events involved in the induction of Crassulacean acid metabolism (CAM) in detached common ice plant (Mesembryanthemum crystallinum) leaves. Transcript abundance of Ppc1, a gene encoding the CAM-specific isoform of phosphoenolpyruvate carboxylase, increased rapidly in response to osmotic stress (dehydration and mannitol), ionic stress (NaCl), and exogenous abscisic acid treatment, but failed to accumulate in response to exogenous cytokinin or methyl jasmonate. Stress-induced accumulation of Ppc1, GapC1, and Mdh1 transcripts was inhibited by pretreating leaves with the calcium chelator ethyleneglycol-bis(aminoethyl ether)-N,N'-tetraacetic acid, suggesting that extracellular calcium participates in signaling events leading to CAM induction. Treatment of unstressed detached leaves with ionomycin, a Ca(2+) ionophore, and thapsigargin, a Ca(2+)-ATPase inhibitor, enhanced Ppc1 transcript accumulation, indicating that elevations in cytosolic [Ca(2+)] are likely to participate in signaling CAM induction. Inhibitors of Ca(2+)- or calmodulin dependent protein kinases (N-[6-aminohexyl]-5-chloro-1-napthalenesulfonamide, Lavendustin C) and protein phosphatase 1 and 2A (okadaic acid) activity suppressed Ppc1 transcript accumulation in response to ionic and osmotic stresses, as well as abscisic acid treatment. These results suggest that both protein phosphorylation and dephosphorylation events participate in signaling during CAM induction. In contrast, pretreatment with cyclosporin A or ascomycin, inhibitors of protein phosphatase 2B activity, stimulated Ppc1 gene expression either directly or indirectly through promoting water loss. PMID- 10517847 TI - A proline-, threonine-, and glycine-rich protein down-regulated by drought is localized in the cell wall of xylem elements. AB - A cDNA clone encoding a proline-, threonine-, and glycine-rich protein (PTGRP) was isolated from a wild tomato species (Lycopersicon chilense) (L.X. Yu, H. Chamberland, J.G. Lafontain, Z. Tabaeizadeh [1996] Genome 39: 1185-1193). Northern-blot analysis and in situ hybridization studies revealed that PTGRP is down-regulated by drought stress. The level of the mRNA in leaves and stems of 8 d drought-stressed plants decreased 5- to 10-fold compared with that in regularly watered plants. The mRNA re-accumulated when drought-stressed plants were rewatered. Antibodies raised against a glutathione S-transferase/PTGRP fusion protein were used to elucidate the subcellular localization of the protein by immunogold labeling. In regularly watered L. chilense plants, PTGRP protein was found to be localized in xylem pit membranes and disintegrated primary walls. Examination of sections from drought-stressed plants revealed a significant decrease in the levels of labeling. In these samples, only a few scattered gold particles were detected in the same areas. In the leaf tissues of plants that had been rewatered for 3 d following an 8-d drought stress, the labeling pattern was similar to that of the regularly watered plants. To our knowledge, PTGRP is the first drought-regulated protein that has been precisely localized in the cell wall. PMID- 10517848 TI - Induced resistance to pathogenic fungi in norway spruce AB - Norway spruce (Picea abies) trees (approximately 16 m high) of a single clone were used to study the effects of fungal infection and wounding on induction of resistance to the bark beetle-associated bluestain fungus Ceratocystis polonica. A dose-response experiment was designed involving three different dosages of fungal (fungus and wound) and sterile agar (wound) pretreatment inoculations (10, 50, or 100 inoculations/m(2) on the stem between 0.8 and 2.0 m high). Three weeks after pretreatment, trees were challenged with a massive C. polonica inoculation (400 inoculations/m(2)). Control trees that received no pretreatment were heavily colonized and killed by the challenge inoculation. The high and medium fungal pretreatments reduced subsequent fungal colonization success by 76% to 97% relative to the control, and fungal pretreatments protected the trees much more efficiently than sterile agar pretreatments. The protection was demonstrated to be local and not systemic in a subsequent experiment, where trees were pretreated with the medium fungal dosage on the lower bole and challenge inoculated further up the stem. Protection was also demonstrated to be pathogen nonspecific, as trees that had been pretreated with a medium dosage of the root rot fungus Heterobasidion annosum showed enhanced resistance to challenge inoculation with C. polonica. PMID- 10517849 TI - Dynamic properties of endogenous phytochrome A in Arabidopsis seedlings. AB - The dynamic behavior of phytochrome A (phyA) in seedlings of the model plant Arabidopsis was examined by in vivo spectroscopy and by western and northern blotting. Rapid accumulation of phyA was observed, reaching a steady state after 3 d. Both red and far-red light initiated a rapid destruction of the far-red light-absorbing form of phytochrome (Pfr); the apparent half-life was only 4-fold longer in far-red than in red light. Furthermore, the Pfr-induced destruction of the red-light-absorbing form of phytochrome (Pr) of phyA occurred in darkness with a rate identical to that of Pfr destruction. A 2-fold decrease in mRNA abundance was observed after irradiation, irrespective of the applied light quality. However, reaccumulation occurred rapidly after far-red but slowly after red irradiation, indicating different modes of regulation of phytochrome expression after light-dark transitions depending on the light quality of the preceding irradiation. The wavelength dependency of the destruction rates was distinct from that of mustard, a close relative of Arabidopsis, and was explained on the basis of Pfr-induced Pr destruction and a simple kinetic two-step model. No dark reversion was detectable in the destruction kinetics after a red pulse. From these data we conclude that Arabidopsis phyA differs significantly in several aspects from other dicot phytochromes. PMID- 10517850 TI - Directed mutation of the Rubisco large subunit of tobacco influences photorespiration and growth. AB - The gene for the large subunit of Rubisco was specifically mutated by transforming the chloroplast genome of tobacco (Nicotiana tabacum). Codon 335 was altered to encode valine instead of leucine. The resulting mutant plants could not grow without atmospheric CO2 enrichment. In 0.3% (v/v) CO2, the mutant and wild-type plants produced similar amounts of Rubisco but the extent of carbamylation was nearly twice as great in the mutants. The mutant enzyme's substrate-saturated CO2-fixing rate and its ability to distinguish between CO2 and O2 as substrates were both reduced to 25% of the wild type's values. Estimates of these parameters obtained from kinetic assays with the purified mutant enzyme were the same as those inferred from measurements of photosynthetic gas exchange with leaves of mutant plants. The Michaelis constants for CO2, O2, and ribulose-1,5-bisphosphate were reduced and the mutation enhanced oxygenase activity at limiting O2 concentrations. Consistent with the reduced CO2 fixation rate at saturating CO2, the mutant plants grew slower than the wild type but they eventually flowered and reproduced apparently normally. The mutation and its associated phenotype were inherited maternally. The chloroplast-transformation strategy surmounts previous obstacles to mutagenesis of higher-plant Rubisco and allows the consequences for leaf photosynthesis to be assessed. PMID- 10517851 TI - Plant succinic semialdehyde dehydrogenase. Cloning, purification, localization in mitochondria, and regulation by adenine nucleotides. AB - Succinic semialdehyde dehydrogenase (SSADH) is one of three enzymes constituting the gamma-aminobutyric acid shunt. We have cloned the cDNA for SSADH from Arabidopsis, which we designated SSADH1. SSADH1 cDNA encodes a protein of 528 amino acids (56 kD) with high similarity to SSADH from Escherichia coli and human (>59% identity). A sequence similar to a mitochondrial protease cleavage site is present 33 amino acids from the N terminus, indicating that the mature mitochondrial protein may contain 495 amino acids (53 kD). The native recombinant enzyme and the plant mitochondrial protein have a tetrameric molecular mass of 197 kD. Fractionation of plant mitochondria revealed its localization in the matrix. The purified recombinant enzyme showed maximal activity at pH 9.0 to 9.5, was specific for succinic semialdehyde (K(0.5) = 15 microM), and exclusively used NAD+ as a cofactor (Km = 130 +/- 77 microM). NADH was a competitive inhibitor with respect to NAD+ (Ki = 122 +/- 86 microM). AMP, ADP, and ATP inhibited the activity of SSADH (Ki = 2.5-8 mM). The mechanism of inhibition was competitive for AMP, noncompetitive for ATP, and mixed competitive for ADP with respect to NAD+. Plant SSADH may be responsive to mitochondrial energy charge and reducing potential in controlling metabolism of gamma-aminobutyric acid. PMID- 10517852 TI - Rapid repression of maize invertases by low oxygen. Invertase/sucrose synthase balance, sugar signaling potential, and seedling survival. AB - We show here that invertase gene expression and the invertase-sucrose (Suc) synthase ratio decrease abruptly in response to low oxygen in maize root tips. In addition to aiding in the conservation of carbon and possibly ATP, this response has the potential to directly affect sugar signaling relative to carbon flux. Experiments were motivated by the potential for a reduced invertase/Suc synthase balance to alter the impact of respiratory and/or membrane carbon flux on sugar signaling. Maize (Zea mays L.) seedlings with 5-cm primary roots were exposed to anoxic (0% [v/v] O2), hypoxic (3% [v/v] O2), and aerobic conditions. Rapid repression of the Ivr1 and Ivr2 maize invertases by low oxygen was evident in root tips within 3 h at both the transcript and activity levels. The speed and extent of this response increased with the degree of oxygen deprivation and differed with genotypes. This decrease in expression also contrasted markedly to that of other genes for respiratory Suc metabolism, such as Suc synthases, which typically increased or remained constant. Although previous work showed that the contrasting effects of sugars on Suc synthase genes were reflected in their regulation by hypoxia and anoxia, the same was not observed for the differentially sugar-responsive invertases. Theoretically advantageous reductions in the invertase/Suc synthase balance thus resulted. However, where this response was extreme (an Oh43 inbred), total sucrolytic capacity dropped below an apparent minimum and root tip viability was reduced. Paradoxically, only Oh43 seedlings showed survival levels >80% (versus <50%) after extreme, long-term stress, suggesting a possible advantage for multiple means of reducing sink activity. Overall, our results demonstrate a rapid change in the regulation and balance of invertases and Suc synthases that could have an immediate impact on limiting the extent of Suc cleavage and reducing the extent of concomitant, hexose-based sugar signaling under low oxygen. PMID- 10517854 TI - Phloem transport of D,L-glufosinate and acetyl-L-glufosinate in glufosinate resistant and -susceptible brassica napus AB - Phloem transport of D,L-[(14)C]glufosinate, D-[(14)C]glufosinate, and acetyl-L [(14)C]glufosinate was examined in the susceptible Brassica napus cv Excel and a glufosinate-resistant genotype (HCN27) derived by transformation of cv Excel with the phosphinothricin-N-acetyltransferase (pat) gene. Considerably more (14)C was exported from an expanded leaf in HCN27 than in cv Excel following application of D,L-[(14)C]glufosinate (25% versus 6.3% of applied, respectively, 72 h after treatment). The inactive isomer, D-glufosinate, was much more phloem mobile in cv Excel than racemic D,L-glufosinate. Foliar or root supplementation with 1 mM glutamine increased D,L-[(14)C]glufosinate translocation in cv Excel but only transiently, suggesting that glutamine depletion is not the major cause of the limited phloem transport. Acetyl-L-[(14)C]glufosinate (applied as such or derived from L-glufosinate in pat transformants) was translocated extensively in the phloem of both genotypes. Acetyl-L-[(14)C]glufosinate was readily transported into the floral buds and flowers, and accumulated in the anthers in both genotypes. These results suggest that phloem transport of D,L-glufosinate is limited by rapid physiological effects of the L-isomer in source leaf tissue. The accumulation of acetyl-L-glufosinate in the anthers indicates that it is sufficiently phloem mobile to act as a foliar-applied chemical inducer of male sterility in plants expressing a deacetylase gene in the tapetum, generating toxic concentrations of L-glufosinate in pollen-producing tissues. PMID- 10517853 TI - The Arabidopsis homolog of yeast TAP42 and mammalian alpha4 binds to the catalytic subunit of protein phosphatase 2A and is induced by chilling. AB - Type 2A serine/threonine protein phosphatases (PP2A) have been implicated as important mediators of a number of plant growth and developmental processes. In an effort to identify plant PP2A substrates and/or regulators, we performed a yeast two-hybrid screen using an Arabidopsis PP2A catalytic subunit cDNA as bait. All true positives identified by this screen were derived from the same gene, which we have named TAP46 (2A phosphatase associated protein of 46 kD). The TAP46 gene appears to be a single-copy gene and is expressed in all Arabidopsis organs. Transcripts derived from this gene are induced by chilling treatment but not by heat or anaerobic stress. Immunoprecipitation assays using antibodies generated to a peptide spanning amino acids 356 to 366 of TAP46 indicate that TAP46 is associated with a type 2A protein phosphatase in vivo. A search of the database identified TAP46 as a homolog of Saccharomyces cerevisiae TAP42 and mammalian alpha4. These two proteins are known to bind to the catalytic subunit of PP2A and to function in the target-of-rapamycin signaling pathway. Our results identify TAP46 as a plant PP2A-associated protein, with a possible function in the chilling response, and suggest that a target-of-rapamycin-like signaling pathway may exist in plants. PMID- 10517855 TI - Modification of distinct aspects of photomorphogenesis via targeted expression of mammalian biliverdin reductase in transgenic Arabidopsis plants. AB - The phenotypic consequences of targeted expression of mammalian biliverdin IXalpha reductase (BVR), an enzyme that metabolically inactivates the linear tetrapyrrole precursors of the phytochrome chromophore, are addressed in this investigation. Through comparative phenotypic analyses of multiple plastid targeted and cytosolic BVR transgenic Arabidopsis plant lines, we show that the subcellular localization of BVR affects distinct subsets of light-mediated and light-independent processes in plant growth and development. Regardless of its cellular localization, BVR suppresses the phytochrome-modulated responses of hypocotyl growth inhibition, sucrose-stimulated anthocyanin accumulation, and inhibition of floral initiation. By contrast, reduced protochlorophyll levels in dark-grown seedlings and fluence-rate-dependent reduction of chlorophyll occur only in transgenic plants in which BVR is targeted to plastids. Together with companion analyses of the phytochrome chromophore-deficient hy1 mutant, our results suggest a regulatory role for linear tetrapyrroles within the plastid compartment distinct from their assembly with apophytochromes in the cytosol. PMID- 10517856 TI - Histidine-41 of the cytochrome b5 domain of the borage delta6 fatty acid desaturase is essential for enzyme activity. AB - Unlike most other plant microsomal desaturases, the Delta6-fatty acid desaturase from borage (Borago officinalis) contains an N-terminal extension that shows homology to the small hemoprotein cytochrome (Cyt) b5. To determine if this domain serves as a functional electron donor for the Delta6-fatty acid desaturase, mutagenesis and functional analysis by expression in transgenic Arabidopsis was carried out. Although expression of the wild-type borage Delta6 fatty acid desaturase resulted in the synthesis and accumulation of Delta6 unsaturated fatty acids, this was not observed in plants transformed with N terminally deleted forms of the desaturase. Site-directed mutagenesis was used to disrupt one of the axial heme-binding residues (histidine-41) of the Cyt b5 domain; expression of this mutant form of the Delta6-desaturase in transgenic plants failed to produce Delta6-unsaturated fatty acids. These data indicate that the Cyt b5 domain of the borage Delta6-fatty acid desaturase is essential for enzymatic activity. PMID- 10517858 TI - Roles of cellulose and xyloglucan in determining the mechanical properties of primary plant cell walls AB - The primary cell walls of growing and fleshy plant tissue mostly share a common set of molecular components, cellulose, xyloglucan (XyG), and pectin, that are required for both inherent strength and the ability to respond to cell expansion during growth. To probe molecular mechanisms underlying material properties, cell walls and analog composites from Acetobacter xylinus have been measured under small deformation and uniaxial extension conditions as a function of molecular composition. Small deformation oscillatory rheology shows a common frequency response for homogenized native cell walls, their sequential extraction residues, and bacterial cellulose alone. This behavior is characteristic of structuring via entanglement of cellulosic rods and is more important than cross-linking with XyG in determining shear moduli. Compared with cellulose alone, composites with XyG have lower stiffness and greater extensibility in uniaxial tension, despite being highly cross-linked at the molecular level. It is proposed that this is due to domains of cross-linked cellulose behaving as mechanical elements, whereas cellulose alone behaves as a mat of individual fibrils. The implication from this work is that XyG/cellulose networks provide a balance of extensibility and strength required by primary cell walls, which is not achievable with cellulose alone. PMID- 10517857 TI - Rates of sugar uptake by guard cell protoplasts of pisum sativum L. Related To the solute requirement for stomatal opening AB - We wished to determine whether the capacity of the sugar uptake mechanisms of guard cells of the Argenteum mutant of pea (Pisum sativum L.) sufficed to support a concurrent stomatal opening movement. Sugar uptake by guard cell protoplasts was determined by silicone-oil-filtering centrifugation. The protoplasts took up [(14)C]glucose, [(14)C]fructose, and [(14)C]sucrose (Suc), apparently in symport with protons. Mannose, galactose, and fructose competed with Glc for transport by a presumed hexose carrier. The uptake of Glc saturated with a K(m) of 0.12 mM and a V(max) of 19 fmol cell(-1) h(-1). At external concentrations <1 mM, the uptake of Suc was slower than that of Glc. It exhibited a saturating component with a K(m) varying between 0.25 and 0.8 mM and a V(max) between 1 and 10 fmol cell(-1) h(-1), and at external concentrations >1 mM, a non-saturating component. At apoplastic sugar concentrations below 4 mM, sugar import was estimated to be mainly in the form of hexoses and too slow to support a simultaneous stomatal opening movement. If, however, during times of high photosynthesis and transpiration, the apoplastic Suc concentration rose and entered the range of non saturating import, absorbed Suc could replace potassium malate as the osmoticum for the maintenance of stomatal opening. PMID- 10517859 TI - Snow-mold-induced apoplastic proteins in winter rye leaves lack antifreeze activity AB - During cold acclimation, winter rye (Secale cereale L.) plants secrete antifreeze proteins that are similar to pathogenesis-related (PR) proteins. In this experiment, the secretion of PR proteins was induced at warm temperatures by infection with pink snow mold (Microdochium nivale), a pathogen of overwintering cereals. A comparison of cold-induced and pathogen-induced proteins showed that PR proteins accumulated in the leaf apoplast to a greater level in response to cold. The PR proteins induced by cold and by snow mold were similar when separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and examined by immunoblotting. Both groups of PR proteins contained glucanase-like, chitinase-like, and thaumatin-like proteins, and both groups exhibited similar levels of glucanase and chitinase activities. However, only the PR proteins induced by cold exhibited antifreeze activity. Our findings suggest that the cold induced PR proteins may be isoforms that function as antifreeze proteins to modify the growth of ice during freezing while also providing resistance to the growth of low-temperature pathogens in advance of infection. Both functions of the cold-induced PR proteins may improve the survival of overwintering cereals. PMID- 10517860 TI - Analysis of the relative increase in photosynthetic O(2) uptake when photosynthesis in grapevine leaves is inhibited following low night temperatures and/or water stress AB - We found similarities between the effects of low night temperatures (5 degrees C 10 degrees C) and slowly imposed water stress on photosynthesis in grapevine (Vitis vinifera L.) leaves. Exposure of plants growing outdoors to successive chilling nights caused light- and CO(2)-saturated photosynthetic O(2) evolution to decline to zero within 5 d. Plants recovered after four warm nights. These photosynthetic responses were confirmed in potted plants, even when roots were heated. The inhibitory effects of chilling were greater after a period of illumination, probably because transpiration induced higher water deficit. Stomatal closure only accounted for part of the inhibition of photosynthesis. Fluorescence measurements showed no evidence of photoinhibition, but nonphotochemical quenching increased in stressed plants. The most characteristic response to both stresses was an increase in the ratio of electron transport to net O(2) evolution, even at high external CO(2) concentrations. Oxygen isotope exchange revealed that this imbalance was due to increased O(2) uptake, which probably has two components: photorespiration and the Mehler reaction. Chilling- and drought-induced water stress enhanced both O(2) uptake processes, and both processes maintained relatively high rates of electron flow as CO(2) exchange approached zero in stressed leaves. Presumably, high electron transport associated with O(2) uptake processes also maintained a high DeltapH, thus affording photoprotection. PMID- 10517861 TI - The Electronic Plant Gene Register. PMID- 10517862 TI - From global expression data to gene networks. AB - Allowing the parallel monitoring of the transcription of thousands of genes, microarrays constitute a powerful technique for functional genomics. In a recent paper, a clustering method and a local alignment software were combined to identify DNA motifs in sets of yeast genes endowed with similar transcription profiles throughout mitosis (1). Identifying various known transcriptional binding sites together with new putative ones, the authors made a significant step towards a systematic characterization of the regulatory structure of genomic networks. BioEssays 1999;21:895-899. PMID- 10517863 TI - Sensory experience and the formation of a computational map of auditory space in the brain. AB - The basic wiring of the brain is first established before birth by using a variety of molecular guidance cues. These connections are then refined by patterns of neural activity, which are initially generated spontaneously and subsequently driven by sensory experience. In the superior colliculus, a midbrain nucleus involved in the control of orienting behaviour, visual, auditory, and tactile inputs converge to form superimposed maps of sensory space. Maps of visual space and of the body surface arise from spatially ordered projections from the retina and skin, respectively. In contrast, the map of auditory space is computed within the brain by tuning the neurons to different localization cues that result from the acoustical properties of the head and ears. Establishing and maintaining the registration of the maps in the face of individual differences in the size and relative positions of different sense organs is an activity dependent process in which the synaptic circuits underlying the auditory representation are modified and calibrated under the influence of both auditory and visual experience. BioEssays 1999;21:900-911. PMID- 10517864 TI - Signaling pathways are focused at specialized regions of the plasma membrane by scaffolding proteins of the MAGUK family. AB - The MAGUKs (membrane-associated guanylate kinase homologs) are a family of proteins that act as molecular scaffolds for signaling pathway components at the plasma membrane of animal cells. They are localized in and required for the formation of several types of cell junctions, including epithelial tight and septate junctions as well as synaptic and neuromuscular junctions. They are also localized at the plasma membrane of other cell types, including erythrocytes, where they contribute to cell shape maintenance. MAGUKs function mainly by binding directly to the cytoplasmic termini of transmembrane proteins as well as to other signal transduction proteins. They appear to hold together elements of individual signaling pathways, thereby contributing to the efficiency and specificity of signaling interactions while simultaneously maintaining the structural specializations of the plasma membrane. BioEssays 1999;21:912-921. PMID- 10517865 TI - At the nexus between pattern formation and cell-type specification: the generation of individual neuroblast fates in the Drosophila embryonic central nervous system. AB - The specification of specific and often unique fates to individual cells as a function of their position within a developing organism is a fundamental process during the development of multicellular organisms. The development of the Drosophila embryonic central nervous system serves as an excellent model system in which to clarify the developmental mechanisms that link pattern formation to cell-type specification. The Drosophila embryonic central nervous system develops from a set of neural stem cells termed neuroblasts. Neuroblasts arise from the ectoderm in an invariant pattern, and each neuroblast acquires a unique fate based on its position within this pattern. Two groups of genes recently have been demonstrated to govern the individual fate specification of neuroblasts. One group, the segment polarity genes, enables neuroblasts that develop in different anteroposterior positions to acquire different fates. The second group, referred to as the columnar genes, ensures that neuroblasts that develop in different dorsoventral domains assume different fates. When integrated, the activities of the segment polarity and columnar genes create a Cartesian coordinate system that bestows unique fates to individual neuroblasts as a function of their position of formation within the ectoderm. BioEssays 1999;21:922-931. PMID- 10517866 TI - The tetratricopeptide repeat: a structural motif mediating protein-protein interactions. AB - The tetratricopeptide repeat (TPR) motif is a protein-protein interaction module found in multiple copies in a number of functionally different proteins that facilitates specific interactions with a partner protein(s). Three-dimensional structural data have shown that a TPR motif contains two antiparallel alpha helices such that tandem arrays of TPR motifs generate a right-handed helical structure with an amphipathic channel that might accommodate the complementary region of a target protein. Most TPR-containing proteins are associated with multiprotein complexes, and there is extensive evidence indicating that TPR motifs are important to the functioning of chaperone, cell-cycle, transcription, and protein transport complexes. The TPR motif may represent an ancient protein protein interaction module that has been recruited by different proteins and adapted for specific functions. BioEssays 1999;21:932-939. PMID- 10517867 TI - Membrane associated matrix metalloproteinases in metastasis. AB - Hematogenous metastasis is postulated to involve tumor cell-initiated degradation of basement membrane barriers and underlying connective tissue matrices. Matrix metalloproteinases (MMP) are zinc-dependent endopeptidases that have been implicated in the proteolytic events of tumor cell invasion. Research has revealed a class of membrane-anchored metalloproteinases (MT-MMPs) and has provided convincing evidence that these enzymes activate latent MMP-2 (72 kDa gelatinase A) on the cell surface. The activation of plasma membrane associated MMP is a potential mechanism for facilitation of cellular metastasis and requires consideration when addressing potential roles of MMPs in tumor progression. This review focuses on potential in vivo regulatory mechanisms of membrane-associated MMP activity in the context of tumor cell interaction with matrix macromolecules. BioEssays 1999;21:940-949. PMID- 10517868 TI - Functions of the retinoblastoma protein. AB - The retinoblastoma protein (pRB) can both positively and negatively regulate transcription. The former correlates with its ability to promote differentiation and the latter with its ability to regulate entry into S-phase. pRB negatively regulates transcription by forming complexes with members of the E2F transcription factor family. These complexes, when bound to E2F sites within certain target genes, actively repress transcription through a variety of mechanisms including physical interaction with adjacent transcriptional activation domains and recruitment of proteins that directly, or indirectly, lead to histone deacetylation. pRB function is, in turn, modulated by phosphorylation mediated by cyclin-dependent kinases. Emerging data suggest that combinatorial control of pRB function may be achieved through the use of different phosphoacceptor sites, different cyclin/cdk docking sites, and different cyclin/cdk complexes. The untimely activation of E2F responsive genes can induce apoptosis. This comes about at least partly through the induction of ARF, which leads to the stabilization and activation of p53. BioEssays 1999;21:950-958. PMID- 10517869 TI - Chaperone-percolator model: a possible molecular mechanism of Anfinsen-cage-type chaperones. AB - Although we have a rather elaborate "working-cycle" for the 60 kDa molecular chaperones, which possess a cavity, and are called Anfinsen-cage-type chaperones to emphasize that they provide a closed, protected environment to help the folding of their substrates, our understanding of the molecular mechanism of how these chaperones help protein folding is still incomplete. The present study adds two novel elements to the mechanism of how Anfinsen-cage-type chaperones (members of the 60 kDa chaperone family) aid protein folding. It is proposed that (1) these chaperones do not generally unfold their targets, but by a multidirectional expansion preferentially loosen the tight, inner structure of the collapsed target protein; and (2) during the expansion water molecules enter the hydrophobic core of the target, this percolation being a key step in chaperone action. This study compares this chaperone-percolator model with existing explanations and suggests further experiments to test it. BioEssays 1999;21:959 965. PMID- 10517871 TI - Evolutionary developmental biology of the cerebral cortex PMID- 10517870 TI - The hybridoma revolution: an offshoot of basic research. AB - In this narrative, I describe how my interest in the nature and origin of antibody diversity led me to tackle the problem by using somatic cell genetic techniques. The first hybridoma (an immortal antibody-secreting cell line derived by fusion of a short-lived lymphocyte and a myeloma cell line) was an offshoot of this approach. Although not intended for such purposes, it soon became obvious that this invention had widespread potential in basic research and industry. Indeed, the technique opened new inroads into the study of complex biological substances and became the method of choice to define new differentiation markers. Hybridomas also allowed us to dissect the immune response to a simple antigen and to demonstrate the critical role of somatic mutations in the generation of high affinity antibodies. Now, monoclonal antibodies can be derived and manipulated in vitro, leading to important new developments in therapeutic applications. BioEssays 1999;21:966-973. PMID- 10517872 TI - Prostate stromal cell-derived hepatocyte growth factor induces invasion of prostate cancer cell line DU145 through tumor-stromal interaction. AB - BACKGROUND: In prostate cancer, several growth factors derived from stromal cells regulate tumor cell growth. Hepatocyte growth factor (HGF) possesses biological activities that promote cancer proliferation and invasion through tumor-stromal interaction. We examined how prostate stromal cell-derived HGF affects invasion of prostate cancer cells through this interaction. METHODS: The effects of HGF, various growth factors (transforming growth factor (TGF)-alpha, TGF-beta1, basic fibroblast growth factor, keratinocyte growth factor, and platelet-derived growth factor), and conditioned medium (CM) from prostate stromal cells (PrSC) on prostate cancer cells (LNCaP, PC-3, and DU145) were determined by collagen gel invasion assay. DU145 cells and PrSC were cocultured for Matrigel invasion chamber assay. Induction activity of CM from cancer cells to stimulate HGF production by PrSC was studied by the ELISA method and Western blotting. RESULTS: LNCaP and PC-3 cells did not respond to any of the factors examined. Invasion of DU145 cells into the collagen gel matrix was induced by HGF and TGF-beta1, but not by any of the other factors tested. When DU145 cells were cultured in CM from PrSC or cocultured with PrSC, the cells acquired invasive potential, and this invasion was inhibited by an antibody against HGF, but not against TGF-beta1. Native-type HGF production in PrSC was enhanced by some unknown inducer(s) produced by cancer cells. CONCLUSIONS: PrSC-derived HGF enhanced invasive activity of the prostate cancer cell line DU145 through tumor-stromal interaction, wherein DU145 cells secreted some HGF-inducer(s) for PrSC. PMID- 10517873 TI - Three-dimensional spheroid cultures of human prostate cancer cell lines. AB - BACKGROUND: Many of the available human prostate cancer (PC) cell lines have lost androgen sensitivity and no longer secrete prostate-specific proteins after serial culturing in cell monolayers. Three-dimensional spheroid cultures have been found to better mimic the in vivo phenotypes of several nonprostatic cell lines. METHODS: We analyzed seven PC cell lines to determine if spheroid culturing results in greater sensitivity to androgens and 1alpha,25(OH)(2) vitamin D(3) (1,25(OH)(2) D(3)) with regards to their growth, differentiation, and apoptotic potential. RESULTS: Only PC-3 cells showed greater sensitivity to the growth-inhibitory effects of 1, 25(OH)(2) D(3), while ALVA-31 showed a diminished response. The regulation of prostate-specific antigen and prostate specific acid phosphatase remained unchanged. However, these studies provided several unique findings not observed in cell monolayers. First, three basic spheroid morphologies were observed with varying degrees of intercellular adhesions. Secondly, the cell lines that formed the tightest spheroids consistently grew at the slowest rates, regardless of their growth rate in monolayers. Lastly, 1,25(OH)(2) D(3) treatment of ALVA-31 and PPC-1 spheroids greatly reduced intercellular adhesions, and rendered ALVA-31 spheroids resistant to apoptotic induction by Fas ligand expressed via a recombinant adenoviral construct. CONCLUSIONS: Our results suggest that spheroid cultures of human PC cells may provide unique insights regarding cell adhesion and apoptotic potential that are diminished or absent in monolayer cultures. PMID- 10517874 TI - Relationship of Ki-67 labeling index to DNA-ploidy, S-phase fraction, and outcome in prostate cancer treated with radiotherapy. AB - BACKGROUND: Our purpose was to evaluate the relationship of Ki-67 labeling index (Ki67-LI) to deoxyribonucleic acid (DNA) ploidy, S phase fraction (SPF), other clinical prognostic factors, and clinical outcome for patients with prostate cancer treated by external beam radiotherapy. METHODS: Tissue was retrieved from 42 patients who underwent transurethral resection of the prostate before treatment with external beam radiotherapy between 1987-1993. DNA histogram profiles were classified as diploid (diploid + near-diploid) and nondiploid (tetraploid + aneuploid). Immunohistochemical staining of Ki-67 by the MIB-1 monoclonal antibody was used to calculate Ki67-LI. Median patient follow-up was 62 months. Treatment failure was defined as two consecutive rises in serum prostate-specific antigen (PSA) or clinical evidence of disease recurrence. RESULTS: The mean and median Ki67-LIs were 3.1 and 2.4, respectively (range, 0 12.4). Mean Ki67-LI values were significantly associated with higher stage, Gleason score, and pretreatment PSA. Nondiploid tumors had significantly higher Ki67-LIs, as did patients who failed radiotherapy over the follow-up period. SPF was not significantly correlated with Ki67-LI. As a categorical variable, the most significant relationships were seen when Ki67-LI was subdivided into thirds around the median (Ki67-LI 1.5-3.5%, and Ki67-LI >3.5%). This trichotomous variable correlated significantly with pretreatment PSA (P = 0.0008), tumor stage (P = 0.016), Gleason score (P = 0.024), and treatment failure (P = 0.0015), but not with DNA-ploidy (P = 0.15). In actuarial univariate analyses, Ki67-LI appeared to be a more significant predictor of patient outcome (P = 0.003) than DNA-ploidy (P = 0.035). CONCLUSIONS: The Ki67-LI correlated with known prognostic factors such as pretreatment PSA, tumor stage, and Gleason score, and was also weakly related to DNA-ploidy. In comparison to DNA-ploidy, Ki67 LI seems to be a better correlate of treatment outcome. PMID- 10517875 TI - Isolation of rat ventral prostate basal and luminal epithelial cells by the STAPUT technique. AB - BACKGROUND: The prostatic epithelium consists principally of basal epithelial cells, luminal epithelial cells, and neuroendocrine cells. Several studies support the concept that among basal cells, a subpopulation of stem cells resides which is capable of giving rise to other stem cells, basal epithelial cells, and also luminal epithelial cells and neuroendocrine cells. Other investigators suggest that luminal epithelial cells can also regenerate prostatic epithelium. Availability of pure populations of basal and luminal epithelial cells will aid in studies on defining the cellular pathways of differentiation during normal and pathological conditions. This study was designed to isolate and characterize pure populations of basal and luminal epithelial cells from adult rat ventral prostates. METHODS: Sequential enzymatic digestion and differential plating permitted the separation of glandular epithelial cells from stromal cells. The glandular epithelial cells were subjected to the STAPUT technique. RESULTS: Two types of cell populations, a large single-cell population and a small single-cell population, were obtained and characterized as basal and luminal epithelial cells by immunostaining for cytokeratin 5 and cytokeratin 8, respectively. CONCLUSIONS: Our results indicate that purified populations of prostatic basal and luminal epithelial cells can be isolated by the STAPUT technique. PMID- 10517876 TI - Effect of prenatal vitamin D (calcitriol) exposure on the growth and development of the prostate. AB - BACKGROUND: We previously found that in the absence of testosterone (T), calcitriol promotes proliferation of normal prostatic stroma, while in the presence of T, it has a differentiating effect on prostatic epithelium. The present study was conducted to determine the effect of calcitriol exposure in utero on the postnatal development of the normal prostate. METHODS: Pregnant rats were injected subcutaneously with either 1.25 microg of calcitriol or vehicle alone on alternate days till delivery. Calcitriol-exposed and control pups were sacrificed at age 25 days (prepuberty), 63 days (postpuberty), or 102 days (adults), and their prostates and seminal vesicles were harvested and weighed. RESULTS: Pups prenatally exposed to calcitriol and sacrificed before puberty (25 days) had a 35% greater mean prostatic weight than controls (0.0314 vs. 0.0422 g, P < 0.007), and calcitriol-exposed adult rats (102 days) had a 68% greater mean prostatic weight than controls (0.1365 vs. 0.2304 g, P < 0.005). No differences were observed in seminal vesicle weights, and in serum calcium and testosterone levels. A disproportionately high mortality rate from sudden death (71%) was observed at puberty in uncastrated male rats prenatally exposed to calcitriol. CONCLUSIONS: These findings suggest that high-dose calcitriol exposure in utero may uniquely influence subsequent prostatic growth. Nonandrogenic steroids such as calcitriol may also be involved in genetic imprinting of the prostate. PMID- 10517877 TI - Loss of heterozygosity and lack of mutations of the XPG/ERCC5 DNA repair gene at 13q33 in prostate cancer. AB - BACKGROUND: Three regions of chromosome 13 were previously identified for having loss of heterozygosity (LOH) in human prostate cancer. One of them, at 13q33, was defined by LOH at markers D13S158 and D13S280. The XPG/ERCC5 gene, a DNA repair gene that when mutated in the germline leads to xeroderma pigmentosum, has been mapped to 13q33, within one megabase of D13S158 and D13S280. This paper describes LOH and mutational analysis of the XPG gene in human prostate cancers, in order to determine whether the XPG gene is involved in the development of prostate cancer. METHODS: LOH of the XPG gene was analyzed in 40 primary prostate cancers and 14 metastases by using the microsatellite assay, and its mutations were examined in 5 cell lines, 14 metastases, and 8 tumors with LOH at 13q33 by using the single-strand conformation polymorphism (SSCP)-direct DNA sequencing analysis. RESULTS: Four of the 29 (14%) informative primary tumors and 4 of 8 (50%) metastases showed LOH for the XPG gene. Analysis of the 8 tumors with LOH at the 13q33 region, 14 metastases, and 5 cell lines of prostate cancer revealed two polymorphisms but no mutation of the gene. The polymorphism in exon 2 did not change the amino-acid sequence of the XPG protein, but the exon 15 polymorphism altered codon 1104 from histidine to aspartic acid. The two polymorphisms also occurred in individuals without prostate cancer. CONCLUSIONS: LOH at XPG in prostate cancer supports the conclusion that the 13q33 region contains a gene important in the development of prostate cancer, while lack of mutations of the gene suggests that XPG is not the target gene involved. PMID- 10517878 TI - Metallothionein isoform 3 expression in the human prostate and cancer-derived cell lines. AB - BACKGROUND: Expression of metallothionein isoform 3 (MT-3) was initially reported to be confined to neural tissues. However, it was recently demonstrated that MT-3 is expressed in epithelial cells of the human kidney. This motivated the current examination of the expression of MT-3 in the human prostate. METHODS: Immunohistochemistry (IHC) was used to localize the expression of MT-3, RT-PCR to determine the expression of MT-3 mRNA, and Western blot analysis to determine the level of MT-3 protein. RESULTS: Selected epithelial and stromal cells of the normal human prostate were shown to have low levels of MT-3 expression. MT-3 was increased in prostatic intraepithelial neoplasia (PIN) lesions and further increased in a highly variable fashion in prostatic adenocarcinoma. In some adenocarcinomas, MT-3 expression exceeded that of nerve. Three cell culture models of prostate cancer were also shown to variably express MT-3. Restriction enzyme analysis confirmed the expression of MT-3 in the cells and tissues. CONCLUSIONS: MT-3 is expressed in the normal human prostate, and expression is enhanced and highly variable in PIN lesions and primary prostate cancer cells. The variable nature of MT-3 expression was also noted in commonly utilized prostate cancer cell lines. PMID- 10517880 TI - Innovative treatment for clinically localized adenocarcinoma of the prostate: the future role of molecular imaging. PMID- 10517879 TI - Androgen-independent expression of hoxb-13 in the mouse prostate. AB - BACKGROUND: Hox genes encode transcriptional regulatory proteins that are largely responsible for establishing the body plan of all metazoan organisms. A subset of Hox genes is expressed during the period of organogenesis and into adulthood. hoxb-13 is a recently-described member of the Hox gene family that is expressed in the spinal cord, hindgut, and urogenital sinus during embryogenesis. METHODS: Northern blot and in situ hybridization analyses of hoxb-13 expression in adult mouse tissues were performed. RESULTS: hoxb-13 mRNA is restricted to the prostate gland and distal colon in adult animals. In situ hybridization of mouse prostate tissue demonstrated that hoxb-13 is expressed in the epithelial cells of the ventral, dorsal, lateral, and anterior prostate lobes. Accumulation of hoxb-13 mRNA is not diminished following castration. CONCLUSIONS: These data demonstrate that hoxb-13 expression is androgen-independent in mouse prostate glands. The identification of hoxb-13 as an androgen-independent gene expressed in adult mouse prostate epithelial cells provides a new potential target for developing therapeutics to treat advanced prostate cancer. PMID- 10517881 TI - Depositional Cyclicity in the Lower Devonian Helderberg Group of New York State. AB - The Helderberg Group of New York State, consisting of a wide range of shallow water carbonate lithologies, contains one of the finest and most complete stratigraphic records of Lower Devonian earth history. High-frequency spatial and temporal rhythmicity of carbonate accumulation in the Helderberg Group has been evaluated by examining thickness frequency distributions of individual sedimentation units in flasered ribbon rock and of individual lithofacies elements. These distributions provide important insights into the sedimentation processes that ultimately controlled stratal thicknesses. Observed thickness frequencies are closely approximated by the exponential end member of the gamma distribution, wherein flaser and lithofacies thickness frequency over each stratigraphic interval is dependent only on net sequence length and the number of stratal elements. Exponentially distributed thickness frequencies would be expected if lithologic change were a Poisson process, that is, if horizons of lithologic change occurred more or less randomly throughout. Such Poisson variation in lithologic thickness suggests a general absence of regularly recurrent high-frequency depositional forcing of stratal thicknesses during accumulation. At the individual flaser-bedded and lithofacies scale, deposition was primarily a stochastic rather than a deterministic process. Thickness distributions of supposedly shallowing-upward lithofacies associations, and magnitudes of sea-level rise inferred from lithologic change across shallowing cycle boundaries, exhibit similar skewed distributions of thickness/magnitude recurrence in Helderberg Group sections. These also are readily interpreted as generally Poisson processes of carbonate accumulation. Skewed frequency distributions of either cycle thickness or water depth change result from the grouping of a small number of elements from an exponential population of randomly distributed sizes. Modal (albeit skewed) cycle thickness distributions are simply an artifact of cycle definition and need not bear any relation to periodic extrabasinal forcing mechanisms. We conclude that thicknesses of flasered bedding units and lithofacies in the Helderberg Group largely reflect the more or less incidental migration of Lower Devonian depositional subenvironments across New York State during sediment accumulation. Stratigraphic extents of shallowing upward lithofacies associations and apparent punctuated deepening across shallowing "cycle" tops are little different from chance groupings of stratal units randomly drawn from an exponential population. If any sea-level control is manifest in these patterns of sediment accumulation, it must have been nearly haphazard with respect to both secular recurrence and magnitude of eustatic change. PMID- 10517882 TI - Tertiary Normal Faulting in the Canyon Range, Eastern Sevier Desert. AB - The contact between pre-Mesozoic and Tertiary rocks in the western Canyon Range, west-central Utah, has been interpreted as a large, low-angle normal fault that marks the breakaway zone of the hypothesized, basin-forming Sevier Desert detachment. Recent fieldwork suggests that the contact may in fact be depositional along much or all of its length. Deformational fabric in the supposed footwall likely traces to the Mesozoic Sevier orogeny rather than to Tertiary detachment faulting. Kinematic indicators at the range front are not generally consistent with low-angle normal-fault motion; instead, well-exposed high-angle faults are the dominant range-bounding structures. The Tertiary conglomerates of the western Canyon Range foothills, previously viewed as an evolving syntectonic deposit related to detachment faulting, are here reinterpreted as three distinct units that reflect different periods and tectonic settings. The pattern in these conglomerates, and in fault-offset gravity-slide deposits that mantle the western foothills, is consistent with block faulting and rotation along several generations of high-angle structures. Local seismic reflection data lend qualitative support to this interpretation, and underscore the need to consider alternative working hypotheses for evolution of the Sevier Desert basin. PMID- 10517883 TI - Lithospheric Mantle Deformation beneath the Indian Cratons. AB - The nature of deformation of the deep continental roots beneath the Archean-Early Proterozoic terrains opens the question whether these ancient terrains have had stable roots since the Precambrian or whether recent plate motions have deformed them. In view of this, we make an attempt to study the thermal structure beneath the cratonic regions of the Indian shield, which vary in lithospheric thickness from 65 km in the Singhbhum craton to 148 km in the Archean Dharwars. The average depth of 104 km to the top of the underlying asthenosphere is consistent with other termination methods and is in fact less than half the 200-400-km depth found in other stable areas of the earth. Similarly, the average reduced heat flow of about 35 mW/m2 and Moho temperature of about 550 degrees C (range: 400 degrees -730 degrees C) for the Indian cratons are also much higher than their counterparts elsewhere. Our study indicates a large-scale deformation of the cratonic mantle lithosphere beneath the Indian shield since the Mesoproterozoic caused by various geodynamic causes, challenging the idea of stability of deep continental roots. PMID- 10517885 TI - Lethal Sandslides from Eolian Dunes. AB - Fossil vertebrates entombed within the Upper Cretaceous Djadokhta Formation of southern Mongolia bear testimony to a heretofore unknown geologic phenomenon: mass wasting of eolian dunes during heavy rainstorms. Evaporation of shallow penetrating rainwater led to progressive calcite accumulation in a thin layer of sand about 0.5 m below the surface of dune lee slopes. During rare heavy rainstorms, a perched water table developed at the top of calcitic zones. Positive pore water pressure led to translational slides and fast-moving sediment gravity flows that overwhelmed animals on the lee slopes of large dunes and in interdune areas. PMID- 10517884 TI - Structural Evolution of the Tuzgolu Basin in Central Anatolia, Turkey. AB - The Central Anatolian segment of the Alpine-Himalayan orogen contains "interior" basins, the largest of which is the Tuzgolu (Salt Lake) basin (>20,000 km2). It is bounded on the east by the Tuzgolu (Salt Lake) fault zone and on the west by the Yeniceoba and Cihanbeyli fault zones. Structural, stratigraphic, and sedimentologic evidence suggests that the Tuzgolu basin started as a fault controlled basin during late Maastrichtian tectonism when the present-day northwest-trending faults that bound the basin were initiated. These faults may have been formed as normal faults suggesting extension or strike-slip faults with a normal component of movement indicating a large transtension at the time of their initiation. The late Maastrichtian faults were reactivated as strike-slip faults in response to late Eocene compression in the region that produced the Central Anatolian thrust belt to the north and the late Eocene south-dipping thrust faults of the Ulukisla basin to the south. This reactivation is suggested by structurally repeated and missing Paleocene-Eocene deposits in some of the basin's wildcat wells. The late Eocene regression in the Tuzgolu basin was caused by the combined effects of Eocene shortening and a large environmental change. Late Eocene evaporites suggest that the basin was dry before the start of the Neotectonic period, while during the Neotectonic itself the Tuzgolu fault zone was reactivated again, predominantly as a normal fault with a right-lateral strike-slip component. This is evidenced by (1) a major unconformity between the post-Eocene Kochisar Formation of the Tuzgolu basin and the underlying Eocene rock units; (2) a well-developed rollover anticline observed on seismic reflection profiles; and (3) a right-step along the Tuzgolu fault zone seen in the field. PMID- 10517886 TI - Lode Gold Deposits and Archean Mantle Plume-Island Arc Interaction, Abitibi Subprovince, Canada. AB - In combination with seismic interpretations and geochronological constraints, the association of juvenile arc-type low-Ti tholeiitic basalts with komatiites in the southeastern Abitibi subprovince, Canada, supports a history of subduction step back following Late Archean mantle plume-island arc interaction. The resulting paired collision zones preserved abundant komatiites and numerous massive sulphide deposits and established the critical metallogenic features to concentrate the majority of Canada's Precambrian gold resources in a small area of the southern Abitibi subprovince. PMID- 10517887 TI - Late Cenozoic Reverse Faulting in the Fall Zone, Southeastern Virginia. AB - A set of en-echelon reverse faults cut Paleozoic metamorphosed igneous rocks of the Piedmont and overlying late Cenozoic sediments at the Old Hickory Heavy Mineral Deposit in the Fall Zone of southeastern Virginia. Diorite of the eastern Slate Belt was faulted over nearshore to shore-face deposits of the Pliocene Yorktown Formation. These NW-SE-striking faults experienced oblique dip-slip movement with a maximum displacement of up to 6 m on individual faults. Faults tip out along strike and are overlain by distinct cobble beds, suggesting that sediment deposition and faulting were contemporaneous. Deformation at Old Hickory may have been formed by reactivation of existing Paleozoic structures under a regionally extensive compressional stress field parallel to the modern one. PMID- 10517889 TI - 1.57-Ga Magmatism in the South Carpathians: Implications for the Pre-Alpine Basement and Evolution of the Mantle under the European Continent: A Reply. PMID- 10517888 TI - Discussion and Reply 1.57-Ga Magmatism in the South Carpathians: Implications for the Pre-Alpine Basement and Evolution of the Mantle under the European Continent: A Discussion. PMID- 10517891 TI - COX-2 in large bowel cancer: a one-sided story. PMID- 10517890 TI - Vaccines against gut pathogens. PMID- 10517892 TI - Are dilating bile ducts a cause for concern? PMID- 10517893 TI - T-cell lymphoma: the real thing. PMID- 10517894 TI - Renal sodium handling in pre-ascitic cirrhosis. PMID- 10517895 TI - Dendritic cell subsets: the ultimate T cell differentiation decision makers? PMID- 10517896 TI - Treatment of fistulas in Crohn's disease with infliximab. PMID- 10517897 TI - Complete and incomplete intestinal metaplasia at the oesophagogastric junction: prevalences and associations with endoscopic erosive oesophagitis and gastritis. AB - BACKGROUND/AIMS: Intestinal metaplasia (IM) is a common finding at the oesophagogastric junction, but the aetiopathogenesis of the different IM subtypes that is, incomplete IM (specialised columnar epithelium, SCE) and complete IM- and their associations with gastro-oesophageal reflux disease and Helicobacter pylori gastritis are unclear. METHODS: 1058 consecutive dyspeptic patients undergoing gastroscopy were enrolled. The gastric, oesophagogastric junctional, and oesophageal biopsy specimens obtained were stained with haematoxylin and eosin, alcian blue (pH 2.5)-periodic acid Schiff, and modified Giemsa. RESULTS: Complete junctional IM was detected in 196 (19%) of the 1058 subjects, and in 134 (13%) was the sole IM subtype. Incomplete junctional IM (SCE) was detected in 101 (10%) subjects, of whom 62 (61%) also had the complete IM subtype. Of patients with normal gastric histology (n = 426), 6% had complete IM and 7% junctional SCE. The prevalence of both types of IM increased with age in patients with either normal gastric histology or chronic gastritis (n = 611). Epithelial dysplasia was not detected in any patients with junctional IM. In multivariate analysis, independent risk factors for incomplete junctional IM were age (odds ratio (OR) 1.3 per decade, 95% confidence interval (CI) 1.2 to 1.6), endoscopic erosive oesophagitis (OR 1.9, 95% CI 1.1 to 3.2), and chronic cardia inflammation (OR 2.9, 95% CI 1.3 to 6.2), but not gastric H pylori infection (OR 1.0, 95% CI 0.6 to 1.7). In univariate analysis, junctional incomplete IM was not associated with cardia H pylori infection. Independent risk factors for "pure" complete junctional IM (n = 134) were age (OR 1.2 per decade, 95% CI 1.0 to 1.4), antral predominant non-atrophic gastritis (OR 2.6, 95% CI 1.3 to 5.2), antral predominant atrophic gastritis (OR 2.1, 95% CI 1.1 to 5.2), and multifocal atrophic gastritis (OR 7.1, 95% CI 2.5 to 19.8). In univariate analysis, junctional complete IM was strongly associated with chronic cardia inflammation and cardia H pylori infection (p<0. 001). CONCLUSIONS: Both complete and incomplete junctional IM are independent acquired lesions that increase in prevalence with age. Although IM subtypes often occur simultaneously, they show remarkable differences in their associations with gastritis and erosive oesophagitis: junctional complete IM is a manifestation of multifocal atrophic gastritis, while the incomplete form (SCE) may result from carditis and gastro oesophageal reflux disease. The frequency of dysplasia in intestinal metaplasia at the oesophagogastric junction appears to be low. PMID- 10517898 TI - Familial hiatal hernia in a large five generation family confirming true autosomal dominant inheritance. AB - BACKGROUND: Familial hiatal hernia has only rarely been documented. AIMS: To describe the pattern of inheritance of familial hiatal hernia within an affected family. SUBJECTS: Thirty eight members of a family pedigree across five generations. METHODS: All family members were interviewed and investigated by barium meal for evidence of a hiatal hernia. RESULTS: Twenty three of 38 family members had radiological evidence of a hiatal hernia. No individual with a hiatal hernia was born to unaffected parents. In one case direct male to male transmission was shown. CONCLUSIONS: Familial inheritance of hiatal hernia does occur. Evidence of direct male to male transmission points to an autosomal dominant mode of inheritance. PMID- 10517900 TI - The clinicopathological features of extensive small intestinal CD4 T cell infiltration. AB - METHODS: Four patients with clinicopathological features suggesting a new distinct entity defining extensive small intestinal CD4 T cell infiltration were observed. RESULTS: All four patients presented with chronic diarrhoea, malabsorption, and weight loss. Biopsy specimens of the small intestine disclosed extensive and diffuse infiltration of the lamina propria by pleomorphic small T lymphocytes, which were positive for CD3, CD4, CD5, and the beta chain of T cell receptor in all three cases studied and negative for CD103 in all three cases studied. It is notable that, in all invaded areas, the infiltrating cells showed no histological change throughout the whole evolution. In three patients, lymphocyte proliferation was monoclonal and there was extraintestinal involvement. In one patient, lymphoproliferation was oligoclonal and confined to the small intestine. In all four patients, there was no evidence of coeliac disease. Although none of the four patients responded to single or multiple drug chemotherapy, median survival was five years. CONCLUSION: Extensive small intestinal CD4 T cell infiltration is a rare entity, distinct from coeliac disease and associated with prolonged survival. PMID- 10517901 TI - Relative power of linkage and transmission disequilibrium test strategies to detect non-HLA linked coeliac disease susceptibility genes. AB - BACKGROUND: Susceptibility to coeliac disease is genetically determined by possession of specific HLA DQ alleles, acting in concert with one or more non-HLA linked genes. The pattern of familial risk is most parsimonious with a multiplicative model for the interaction between these two classes of genes. Haplotype sharing probabilities across the HLA region in affected sibling pairs suggest that genes within the MHC complex contribute no more than 40% of the sibling familial risk of coeliac disease, making the non-HLA linked gene (or genes) the stronger determinant of coeliac disease susceptibility. Attempts to localise these non-HLA linked genes have been carried out using both linkage and association tests. AIMS: To review the evidence for the involvement of non-HLA linked genes in coeliac disease, and to compare the relative merits of linkage and transmission disequilibrium tests (TDT) to detect the non-HLA linked gene (or genes) contributing to the development of coeliac disease. METHODS: Under a range of genetic models the number of affected sibling pairs needed to detect linkage was compared with the number of families required to show a relation between marker and disease, adopting the TDT strategy. RESULTS AND CONCLUSIONS: Power calculations show that, if there is a single major non-HLA linked susceptibility locus, a non-parametric linkage approach may well prove effective. However, if there are a number of non-HLA susceptibility genes, each with small effect, the sample size necessary for linkage studies will be prohibitive and a systematic search for allelic association should be a more effective strategy. PMID- 10517899 TI - Morphological and functional restoration of parietal cells in helicobacter pylori associated enlarged fold gastritis after eradication. AB - BACKGROUND/AIM: Helicobacter pylori infections are associated with hypochlorhydria in patients with pangastritis. It has previously been shown that eradication of H pylori leads to an increase in acid secretion in H pylori associated enlarged fold gastritis, suggesting that H pylori infection affects parietal cell function in the gastric body. The aim of this study was to evaluate the effects of H pylori infection on parietal cell morphology and function in hypochlorhydric patients. PATIENTS/METHODS: The presence of H pylori infection, mucosal length, and inflammatory infiltration were investigated in six patients with enlarged fold gastritis and 12 patients without enlarged folds. Parietal cell morphology was examined by immunohistochemistry using an antibody against the alpha subunit of H(+),K(+)-ATPase and electron microscopy. In addition, gastric acid secretion and fasting serum gastrin concentration were determined before and after the eradication of H pylori. RESULTS: In the H pylori positive patients with enlarged fold gastritis, fold width, foveolar length, and inflammatory infiltration were increased. In addition, the immunostaining pattern of H(+), K(+)-ATPase was less uniform, and the percentage of altered parietal cells showing dilated canaliculi with vacuole-like structures and few short microvilli was greatly increased compared with that in H pylori positive patients without enlarged folds. After eradication, fold width, foveolar length, and inflammatory infiltrates decreased and nearly all parietal cells were restored to normal morphology. On the other hand, altered parietal cells were negligible in H pylori negative patients. In addition, the basal acid output and tetragastrin stimulated maximal acid output increased significantly from 0.5 (0.5) to 4.1 (1.5) mmol/h and from 2.5 (1.2) to 13.8 (0.7) mmol/h (p<0.01), and fasting serum gastrin concentrations decreased significantly from 213.5 (31.6) to 70.2 (7.5) pg/ml (p<0.01) after eradication in patients with enlarged fold gastritis. CONCLUSION: The morphological changes in parietal cells associated with H pylori infection may be functionally associated with the inhibition of acid secretion seen in patients with enlarged fold gastritis. PMID- 10517902 TI - Diabetic intestinal growth adaptation and glucagon-like peptide 2 in the rat: effects of dietary fibre. AB - BACKGROUND/AIMS: Dietary fibre influence growth and function of the upper gastrointestinal tract. This study investigates the importance of dietary fibre in intestinal growth in experimental diabetes, and correlates intestinal growth with plasma levels of the intestinotrophic factor, glucagon-like peptide 2 (GLP 2). METHODS: Male Wistar rats were randomised to the following groups: two streptozotocin-diabetic and two control groups fed either a fibre-containing or a fibre-free diet for three weeks. Intestinal weight, length, and morphometric data (villus height, villus area, crypt depth) were measured. Blood samples were obtained after two weeks for measurement of GLP-2 and enteroglucagon (glicentin, oxyntomodulin). RESULTS: The mean daily consumption of food in the two diabetic groups was 40% higher than in controls. In diabetic rats fed fibre, the increase in intestinal weight from day 0 to 20 was sixfold greater than that of the controls and small intestine weight per cm length was increased by 50%. In the diabetic rats fed a fibre-free diet, intestinal growth was 30% less than in diabetic rats fed fibre, and intestinal weight increased only threefold compared with controls. Morphometric data showed that the intestinal increase in diabetic rats fed fibre was due primarily to growth of the mucosal layer. Villus height and crypt depth increased 60% and 40% respectively, but by only 20% in fibre-free diabetic rats. The plasma levels of GLP-2 parallelled diabetic intestinal growth, whereas plasma levels of enteroglucagon increased regardless of the extent of intestinal growth. CONCLUSIONS: Intestinal growth in experimental diabetes is strongly influenced by the presence of dietary fibre. The effect may be mediated by GLP-2. PMID- 10517903 TI - Fast acting nervous regulation of immunoglobulin A secretion from isolated perfused porcine ileum. AB - BACKGROUND: The intestinal mucosa harbours a large number of nerve fibres and also plasma cells, providing the anatomical basis for studies of neuroimmune interactions. AIMS: To study the effect of different neurotransmitters and electrical stimulation of the extrinsic intestinal nerves on secretion of immunoglobulin A (IgA). METHODS: IgA was measured, using a specific ELISA, in the luminal and venous effluent from isolated vascularly perfused porcine ileal segments with preserved extrinsic nerve supply. RESULTS: Infusion of several neuropeptides stimulated IgA output. Somatostatin (10(-8) M) stimulated IgA secretion in the luminal effluent from 46.6 (14.3) to 79.3 (19.0) microg/5 min and increased the venous output to 148.3 (23.0)% (n=6) of basal output, whereas noradrenaline (10(-6) M) inhibited the secretion (to 49.2 (6.5)% of basal output, n=6). Electrical stimulation of the mixed extrinsic nerves supplying the intestinal segment had no effect by itself. However, electrical stimulation during infusion of alpha adrenergic blockers or coinfusion of both alpha adrenergic and muscarinic blockers resulted in an immediate and significant increase in IgA, an effect that was abolished by nicotinic blockade. CONCLUSION: The extrinsic nerve supply to the intestine could be involved in fast acting regulation of mucosal immune functions. PMID- 10517904 TI - Villous, hypermucinous mucosa in long standing ulcerative colitis shows high frequency of K-ras mutations. AB - BACKGROUND: K-ras mutation is one of the first genetic alterations in classical colorectal carcinogenesis. AIMS: To investigate the role of K-ras mutations in carcinogenesis, in long standing ulcerative colitis. METHODS: A total of 161 microdissected and 100 DNA samples from 13 patients were analysed for K-ras codons 12 and 13 mutations by means of a combination of enriched polymerase chain reaction amplification and temporal temperature gradient electrophoresis. RESULTS: K-ras mutations were found in 21/161 (13%) microdissected samples in 7/13 large bowels (16 and five in codons 12 and 13, respectively), and in 10/100 (10%) mucosal DNA samples (six and four, respectively). One of four patients with six adenocarcinomas had a K-ras mutation in a carcinoma, as well as one of two patients with large dysplasia associated lesion or mass (DALM). Eight of 13 (61%) areas with villous architecture and large, distended goblet cells, had a K-ras mutation, which was significantly more frequent than in low grade dysplasia (one of 23, 4%) but did not reach significance versus high grade dysplasia (four of 14, 28.5%). K-ras mutations were found in one of 20 (5%) flat lesions indefinite for dysplasia, two of 14 (14%) in non-villous, hypermucinous mucosa, and in one of 57 flat areas negative for dysplasia. CONCLUSION: The highest K-ras mutation frequency was found in villous, hypermucinous mucosa. We suggest that this entity should be investigated further as a potential risk lesion for cancer development. It may represent a pathway directly from non-classical dysplasia to cancer, not previously described. PMID- 10517905 TI - Cell specific effects of glucocorticoid treatment on the NF kappaBp65/IkappaBalpha system in patients with Crohn's disease. AB - BACKGROUND/AIMS/PATIENTS: Glucocorticoid treatment is known to reduce nuclear factor-kappa B (NF-kappaB)p65 binding activity and activation in lamina propria cells of patients with Crohn's disease. However, lamina propria cells of glucocorticoid treated patients did not show increased expression of IkappaBalpha, and the hypothesised upregulation of IkappaBalpha by glucocorticoid treatment has not yet been shown in vivo. To investigate whether cells other than lamina propria localised mononuclear cells contribute to increased IkappaBalpha, resection gut specimens from patients matched for Crohn's disease activity index (CDAI) with or without glucocorticoid treatment were studied, and changes in the NF-kappaB/IkappaBalpha system were determined in the lamina propria as well as in underlying submucosal and endothelial cells. METHODS: Changes in the NF kappaB/IkappaBalpha system were determined by immunohistochemistry, electrophoretic mobility shift assay, and western blot analysis in resected gut specimens from patients matched for CDAI and van Hees index with or without long term glucocorticoid treatment. RESULTS: Resection gut specimens from patients with Crohn's disease under glucocorticoid treatment had significantly lower nuclear NF-kappaBp65 levels in mononuclear, epithelial, and endothelial cells than samples from CDAI and van Hees index matched patients not having glucocorticoid treatment. Nuclear NF-kappaBp65 showed a strong positive correlation with both the CDAI (r = 1 for both groups) and the van Hees index (r = 0.605 for untreated and r = 0.866 for glucocorticoid treated specimens). Lower nuclear translocation of NF-kappaBp65 in the glucocorticoid treated group was paralleled by higher IkappaBalpha levels in vascular endothelial cells, but not in infiltrating mononuclear cells. CONCLUSION: A comparison of resection gut specimens from untreated and treated CDAI matched patients with Crohn's disease showed downregulation of NF-kappaB binding activity and NF-kappaBp65 expression and cell specific induction of endothelial IkappaBkappa expression in the glucocorticoid treated group. As the two groups showed similar disease activity (CDAI, van Hees index), the activation of the NF-kappaBp65/IkappaBalpha system must be only part of the inflammatory cascade leading to the clinical appearance of Crohn's disease. PMID- 10517906 TI - Dexamethasone inhibition of leucocyte adhesion to rat mesenteric postcapillary venules: role of intercellular adhesion molecule 1 and KC. AB - BACKGROUND: A previous study showed that the glucocorticoid dexamethasone, at doses of 100 microg/kg and above, inhibited leucocyte adhesion to rat mesenteric postcapillary venules activated with interleukin 1beta (IL-1beta), as assessed by videomicroscopy. AIMS: To identify whether the adhesion molecule, intercellular adhesion molecule 1 (ICAM-1), or the chemokine KC could be targeted by the steroid to mediate its antiadhesive effect. METHODS: Rat mesenteries were treated with IL-1beta (20 ng intraperitoneally) and the extent of leucocyte adhesion measured at two and four hours using intravital microscopy. Rats were treated with dexamethasone, and passively immunised against ICAM-1 or KC. Endogenous expression of these two mediators was validated by immunohistochemistry, ELISA, and the injection of specific radiolabelled antibodies. RESULTS: Dexamethasone greatly reduced IL-1beta induced leucocyte adhesion, endothelial expression of ICAM-1 in the postcapillary venule, and release of the mast cell derived chemokine KC. Injection of specific antibodies to the latter mediators was also extremely effective in downregulating (>80%) IL-1beta induced leucocyte adhesion. CONCLUSIONS: Induction by IL-1beta of endogenous ICAM-1 and KC contributes to leucocyte adhesion to inflamed mesenteric vessels. Without excluding other possible mediators, these data clearly show that dexamethasone interferes with ICAM-1 expression and KC release from mast cells, resulting in suppression of leucocyte accumulation in the bowel wall, which is a prominent feature of several gastrointestinal pathologies. PMID- 10517907 TI - Effect of different prokinetic agents and a novel enterokinetic agent on postoperative ileus in rats. AB - BACKGROUND/AIM: The effects of different prokinetic agents, the motilide erythromycin and the substituted benzamides metoclopramide and cisapride, were investigated in a rat model of postoperative ileus. These effects were compared with that of granisetron, a 5-hydroxytryptamine (5-HT(3)) receptor antagonist, and a novel enterokinetic agent, prucalopride, a 5-HT(4) receptor agonist. METHODS: Different degrees of inhibition of gastrointestinal transit, measured by the migration of Evans blue, were achieved by skin incision, laparotomy, or laparotomy plus mechanical stimulation of the gut. RESULTS: Metoclopramide decreased the transit after laparotomy with or without mechanical stimulation, whereas cisapride increased it after all three operations. Granisetron had no effect on the transit after the three operations when given alone. Prucalopride tended to increase the transit after laparotomy with or without mechanical stimulation when given alone. However, statistical significance was only reached when prucalopride was combined with granisetron. Erythromycin, a motilin receptor agonist, did not improve postoperative ileus in the rat. CONCLUSIONS: Cisapride, but not metoclopramide or erythromycin, is able to improve postoperative ileus in the rat. The results suggest that a combination of 5-HT(3) receptor antagonist and 5-HT(4) receptor agonist properties may be required to obtain a beneficial effect on surgery induced ileus in the rat. Furthermore, they indirectly indicate that stimulation of the excitatory mechanisms is not able to overcome the inhibitory influence of the neural reflex pathways activated during abdominal surgery. PMID- 10517908 TI - Topical diltiazem and bethanechol decrease anal sphincter pressure without side effects. AB - BACKGROUND: Topical nitrates lower anal sphincter pressure and heal anal fissures, but a majority of patients experience headache. The internal anal sphincter has a calcium dependent mechanism to maintain tone, and also receives an inhibitory extrinsic cholinergic innervation. It may therefore be possible to lower anal sphincter pressure using calcium channel blockers and cholinergic agonists without side effects. AIMS: To investigate the effect of oral and topical calcium channel blockade and a topical cholinomimetic on anal sphincter pressure. METHODS: Three studies were conducted, each involving 10 healthy volunteers. In the first study subjects were given oral 60 mg diltiazem or placebo on separate occasions. They were then given diltiazem once or twice daily for four days. In the second and third studies diltiazem and bethanechol gels of increasing concentration were applied topically to lower anal pressure. RESULTS: A single dose of 60 mg diltiazem lowered the maximum resting anal sphincter pressure (MRP) by a mean of 21%. Once daily diltiazem produced a clinically insignificant effect but a twice daily regimen reduced anal pressure by a mean of 17%. Diltiazem and bethanechol gel produced a dose dependent reduction of the anal pressure; 2% diltiazem produced a maximal 28% reduction, and 0.1% bethanechol a maximal 24% reduction, the effect lasting three to five hours. CONCLUSIONS: Topical diltiazem and bethanechol substantially reduce anal sphincter pressure for a prolonged period, and represent potential low side effect alternatives to topical nitrates for the treatment of anal fissures. PMID- 10517910 TI - Differential expression of cyclooxygenase 2 in human colorectal cancer. AB - BACKGROUND: Experimental, clinical, and epidemiological studies have implicated mitogenic metabolites of arachidonic acid such as prostaglandin E(2) (PGE(2)) in colorectal carcinogenesis. Recently, cyclooxygenase 2 (COX-2) which catalyses the conversion of arachidonic acid to PGE(2), has displayed increased levels in human colorectal cancer. AIMS: To evaluate whether there is differential COX-2 expression from different locations (caecum, ascending, transverse, descending, or sigmoid colon, and rectum) in human colorectal cancer. METHODS: Protein levels of COX-2 were determined by western blot analysis in tumours and adjacent normal mucosa of 39 patients with colorectal cancer. RESULTS: There was a notable overexpression of COX-2 protein in tumours located in the rectum (p<0.001) compared with other locations in the colon. Rectal tumours revealed elevated COX 2 protein levels in 18/20 cases compared with 4/19 colonic cases. No association between enhanced COX-2 protein expression in tumour tissue and Dukes's stages was found. CONCLUSIONS: Results suggest that the differential COX-2 expression may be due to differences in gene regulatory factors affecting COX-2 expression and/or reflect secondary changes in tumour progression which may have clinical implications. PMID- 10517909 TI - Decreased and aberrant nuclear lamin expression in gastrointestinal tract neoplasms. AB - BACKGROUND: Altered expression of lamins A/C and B1, constituent proteins of the nuclear lamina, may occur during differentiation and has also been reported in primary lung cancer. AIMS: To examine the expression of these proteins in gastrointestinal neoplasms. PATIENTS: Archival human paraffin wax blocks and frozen tissue from patients undergoing surgical resection or endoscopic biopsy. METHODS: Immunohistochemistry and western blotting using polyclonal antisera against A type lamins and lamin B1. RESULTS: The expression of lamin A/C was reduced and was frequently undetectable by immunohistochemistry in all primary colon carcinomas and adenomas, and in 7/8 primary gastric cancers. Lamin B1 expression was reduced in all colon cancers, 16/18 colonic adenomas, and 6/8 gastric cancers. Aberrant, cytoplasmic labelling with both antibodies occurred in some colonic cancers and around one third of colonic adenomas. Cytoplasmic lamin A/C expression was detected in 3/8 gastric cancers. Lamin expression was reduced in gastric dysplasia, but not intestinal metaplasia, atrophy, or chronic gastritis. Lamin expression was low in carcinomas of oesophagus, prostate, breast, and uterus, but not pancreas. CONCLUSIONS: Reduced expression of nuclear lamins, sometimes together with aberrant, cytoplasmic immunoreactivity is common in gastrointestinal neoplasms. Altered lamin expression may be a biomarker of malignancy in the gastrointestinal tract. PMID- 10517911 TI - Adjuvant treatment of severe acute pancreatitis with C1 esterase inhibitor concentrate after haematopoietic stem cell transplantation. AB - BACKGROUND: With an incidence of 4%, acute pancreatitis is a common complication of bone marrow or peripheral haematopoietic stem cell transplantation, which contributes significantly to morbidity and mortality in these patients. In most cases, the pathogenesis of acute pancreatitis cannot be attributed to a single pathogenetic factor, as treatment toxicity, acute graft versus host disease, infection, and cholestasis may all contribute. Acute pancreatitis is characterised by inflammation and activation of digestive proenzymes leading to autodigestive destruction of the pancreas and systemic activation of protease cascades including the complement system. AIM: To describe the effects of human C1 esterase inhibitor in two children, who developed severe acute pancreatitis with considerable complement activation after allogeneic haematopoietic stem cell transplantation. METHODS: Both children showed clinical features resembling those observed in capillary leakage syndrome. In both patients, treatment with C1 esterase inhibitor concentrate contributed to a rapid clinical stabilisation. CONCLUSIONS: These observations strongly support the proposed pathophysiological concept that early treatment with C1 esterase inhibitor interferes with the activation of the complement system in acute pancreatitis. Inhibition of complement activation prevents its adverse effects on vascular function and permeability, and thus stabilises intravascular fluid status and prevents multiorgan failure in acute pancreatitis. PMID- 10517912 TI - High proportion of mutant K-ras gene in pancreatic juice of patients with pancreatic cystic lesions. AB - BACKGROUND/AIMS: It was recently reported that the quantitative analysis of mutant K-ras gene in pancreatic juice could be useful for the diagnosis of pancreatic cancer. This methodology was applied to patients with pancreatic cystic lesions. METHODS: DNA was extracted from pancreatic juice collected at the time of endoscopy with injection of secretin. The ratio of the K-ras mutant allele to the wild-type allele was measured by two methods to detect and semiquantify mutant K-ras gene: polymerase chain reaction/preferential homoduplex formation assay and enriched polymerase chain reaction/enzyme linked mini sequence assay. RESULTS: A high frequency of K-ras mutation was detected (more than 2% of all K-ras genes) in six of 14 patients (43%) with pancreatic cysts. This frequency was similar to those detected in patients with pancreatic adenocarcinoma and in intraductal papillary neoplasms of the pancreas. In contrast, the frequency of mutation was low (less than 2%) in patients without either pancreatic cysts or pancreatic neoplasms. CONCLUSIONS: K-ras gene mutation may be derived from duct cells in the pancreas with a high potential for tumorigenesis. Therefore careful follow up of patients with a pancreatic cyst is recommended if they are found to have a high level of the mutant gene in the pancreatic juice. PMID- 10517913 TI - The preoperatively normal bile duct does not dilate after cholecystectomy: results of a five year study. AB - BACKGROUND: The common hepatic duct (CHD) is commonly believed to dilate after cholecystectomy but previous studies have either not measured CHD diameter preoperatively or the follow up period is short. AIMS: To measure CHD diameter before and after cholecystectomy. METHODS: Patients undergoing (open) cholecystectomy and operative cholangiography had ultrasonographic measurement of CHD diameter before, and three and six months, and one and five years after cholecystectomy. The normal duct diameter was considered to be 5 mm or less, with an observer error of +/-1 mm. RESULTS: Fifty nine patients with normal diameter ducts were studied. The majority (more than 95%) of patients did not have a dilatation of the CHD beyond 6 mm after cholecystectomy. The CHD appeared to increase as well as decrease with an overall trend towards a minor increase at five years. This was not statistically significant if the margin of error of 1 mm was taken into account. CONCLUSION: A preoperatively normal CHD does not dilate after cholecystectomy and may require further investigation in symptomatic patients. PMID- 10517914 TI - Helical computed tomographic cholangiography versus endosonography for suspected bile duct stones: a prospective blinded study in non-jaundiced patients. AB - BACKGROUND: Helical computed tomography performed after intravenous administration of a cholangiographic contrast material (HCT-cholangiography) may be useful for detecting bile duct stones in non-jaundiced patients. However, this method has never been compared with other non-invasive biliary imaging tests. AIMS: To compare prospectively HCT-cholangiography and endosonography (EUS) in a group of non-jaundiced patients with suspected bile duct stones. METHODS: Fifty two subjects underwent both HCT-cholangiography and EUS. Endoscopic retrograde cholangiography (ERCP), with or without instrumental bile duct exploration, served as a reference method, and was successful in all but two patients. RESULTS: Thirty four patients (68%) were found to have choledocholithiasis at ERCP. The sensitivity for HCT-cholangiography in stone detection was 85%, specificity 88%, and accuracy 86%. For EUS the sensitivity was 91%, specificity 100%, and accuracy 94%. The differences were not significant. No serious complications occurred with either method. CONCLUSIONS: HCT-cholangiography and EUS are safe and comparably accurate methods for detecting bile duct stones in non-jaundiced patients. PMID- 10517915 TI - Renal distal tubular handling of sodium in central fluid volume homoeostasis in preascitic cirrhosis. AB - BACKGROUND/AIMS: Patients with preascitic liver cirrhosis have an increased central plasma volume, and, for any given plasma aldosterone concentration, they excrete less sodium than healthy controls. A detailed study of the distribution of sodium reabsorption along the segments of the renal tubule, especially the distal one, is still lacking in preascitic cirrhosis. METHODS: Twelve patients with Child-Pugh class A cirrhosis and nine control subjects (both groups on a normosodic diet) were submitted to the following investigations: (a) plasma levels of active renin and aldosterone; (b) four hour renal clearance of lithium (an index of fluid delivery to the loop of Henle), creatinine, sodium, and potassium; (c) dopaminergic activity, as measured by incremental aldosterone response to intravenous metoclopramide. RESULTS: Metoclopramide induced higher incremental aldosterone responses, indicating increased dopaminergic activity in patients than controls, which is evidence of an increased central plasma volume (+30 min: 160.2 (68.8) v 83.6 (35.2) pg/ml, p<0.01; +60 min: 140.5 (80.3) v 36. 8 (36.1) pg/ml, p<0.01). Patients had increased distal fractional sodium reabsorption compared with controls (26.9 (6.7)% v 12.5 (3. 4)% of the filtered sodium load, p<0.05). In the patient group there was an inverse correlation between: (a) absolute distal sodium reabsorption and active renin (r -0.59, p<0.05); (b) fractional distal sodium reabsorption and sodium excretion (r -0.66, p<0.03). CONCLUSIONS: These data suggest that in preascitic cirrhosis the distal fractional tubular reabsorption of sodium is increased and critical in regulating both central fluid volume and sodium excretion. PMID- 10517916 TI - Cancer risk in primary biliary cirrhosis: a study in northern England. AB - BACKGROUND: Suggestions that breast cancer may be more common in patients with primary biliary cirrhosis (PBC) have been challenged. It has recently been proposed that total cancer rates may be higher in patients with PBC, as well as liver cancers. AIMS: To investigate these proposals on a strictly defined case series. SUBJECTS: A total of 769 prevalent or incident PBC patients with "definite" or "probable" disease detected in a defined area of the north-east of England during 1987-94. METHODS: Cancer events and deaths were identified by obtaining information from one or more of the following sources: Office for National Statistics (ONS) Central Registers, Regional Cancer Registry, and clinical case records. Standardised cancer incidence (SIR) and mortality ratios (SMR) were calculated using the local region as the standard population. RESULTS: There were 97 cancer events during 1987-96. SIR from cancer registrations for all cancers was 1.7 (95% confidence interval (CI) 1.3 to 2.2), for liver cancer was 74 (95% CI 32 to 146), and for breast cancer was 1.1 (95% CI 0.4 to 2.4). SMR for all cancers was 1. 8 (95% CI 1.4 to 2.4), for liver cancer was 39 (95% CI 20 to 68), and for breast cancer was 0.4 (95% 0.1 to 1.6). The results were similar after excluding the first year of follow up after PBC diagnosis. CONCLUSIONS: There was some evidence of a small increase in overall cancer incidence and mortality in PBC patients. With the exception of liver cancer, it is unlikely that there is a high excess incidence for PBC patients from any cancer at a particular site, and specifically breast cancer. PMID- 10517917 TI - Genomic heterogeneity in synchronous hepatocellular carcinomas. AB - BACKGROUND: Hepatocellular carcinoma (HCC) arising in cirrhosis is frequently multifocal. Whether HCC develops monoclonally or multiclonally is an unresolved question. Of the multiple tumour nodules present in many patients, it has not been established whether the smaller lesions represent intrahepatic metastases or de novo cancers. AIMS: To assess the degree of genomic heterogeneity in synchronous HCCs in cirrhosis. METHODS: The arbitrarily primed polymerase chain reaction technique was utilised to compare the DNA fingerprint of HCCs and regenerative nodules (RNs) removed from cirrhotic explant livers. RESULTS: Polymorphic genomic heterogeneity was noted in 54 HCCs and 31 RNs microdissected. Even satellite nodules in close proximity within the same segment of the liver were found to have distinct genomic patterns. CONCLUSION: Such genomic heterogeneity in synchronous HCCs may explain poor patient survival after surgical resection. If the smaller tumours are de novo lesions rather than metastases (as these data suggest), then current concepts regarding liver resection as a curative treatment modality for HCC may require reassessment. PMID- 10517919 TI - Loss of interstitial cells and a fibromuscular layer on the luminal side of the colonic circular muscle presenting as megacolon in an adult patient. AB - BACKGROUND: Animal studies have shown that the neuromuscular structures on the luminal side of the colonic circular muscle coordinate circular muscle activity. These structures have been identified by electron microscopy in the normal human colon, but have never been thoroughly studied in patients with acquired intestinal hypoganglionosis. AIMS: To perform histological, immunocytochemical, and electron microscopic examinations of the colon of a patient with acquired intestinal hypoganglionosis presenting as megacolon. PATIENT: A 32 year old man with a one year history of constipation and abdominal distention, a massively dilated ascending and transverse colon, and a normal calibre rectum and descending and sigmoid colon. He had a high titre of circulating serum anti neuronal nuclear antibodies. METHODS: Histology, immunocytochemistry (for neurofilaments, neurone specific enolase, synaptophysin, glial fibrillar acidic protein, S100 protein, and smooth muscle alpha-actin), and electron microscopic examinations on the resected colon. RESULTS: The number of ganglion cells and nerve trunks was decreased throughout the colon. Disruption of the neural network and a loss of interstitial cells of Cajal were observed on the luminal side of the circular muscle; in their place, the non-dilated colon contained a hypertrophic fibromuscular layer. CONCLUSIONS: Striking architectural alterations occurred at the site regarded as the source of the coordination of colonic circular muscle activity in an adult patient with acquired intestinal hypoganglionosis presenting as megacolon. PMID- 10517920 TI - Clonal chromosomal abnormalities in congenital bile duct dilatation (Caroli's disease). AB - BACKGROUND: Caroli's disease is a rare congenital disorder characterised by cystic dilatation of the intrahepatic bile ducts and an increased risk of cholangiocellular carcinoma. The cause is unknown, but occasional familial clustering suggests that some cases are inherited, in particular when occurring in association with polycystic kidney disease and germline PKD1 gene mutations. To date, no gene responsible for familial isolated Caroli's disease has been identified, and no genetic investigations of liver tissue from patients with Caroli's disease have been reported. PATIENT/METHOD: A liver biopsy specimen from a patient with isolated Caroli's disease, without any signs of cholangiocellular carcinoma, was short term cultured and cytogenetically investigated after G banding with Wright's stain. RESULT: Cytogenetic analysis disclosed the karyotype 45-47,XX,der(3)t(3;8)(p23;q13), +2mar[cp6]/46,XX[18]. CONCLUSIONS: The finding of an unbalanced translocation between chromosomes 3 and 8 suggests that loss of distal 3p and/or gain of 8q is of pathogenetic importance in Caroli's disease. Alternatively, structural rearrangements of genes located in 3p23 and 8q13 may be of the essence. These chromosomal breakpoints may also pinpoint the location of genes involved in inherited forms of Caroli's disease not associated with polycystic kidney disease. PMID- 10517918 TI - Regulatory peptide receptors in human hepatocellular carcinomas. AB - BACKGROUND: Overexpression of regulatory peptide receptors in selected human tumours is of diagnostic and therapeutic relevance. AIMS: To evaluate the expression of somatostatin, vasoactive intestinal peptide (VIP), substance P, cholecystokinin (CCK) A and B, and neurotensin receptors in hepatocellular carcinoma (HCC). METHODS: In vitro receptor autoradiography for the various peptide receptors using selective iodinated radioligands on tissue sections in 59 cases of HCC. RESULTS: 41% of HCC expressed somatostatin receptors; 47% expressed VIP receptors. VIP receptors were always identified in non-neoplastic liver tissue. Substance P receptors were only identified in 5% of HCC but in the majority of their peritumorous and intratumorous vessels. CCK-A and -B and neurotensin receptors were not detected in HCC. The somatostatin receptors showed high affinity for somatostatin and octreotide. The VIP receptors had high affinity for VIP, pituitary adenylate cyclase activating peptide (PACAP) 27, and a VIP1 selective analogue, suggesting the presence of VIP1/PACAP II type receptors. PACAP I receptors were identified in two cases. Substance P receptors were all of the NK1 subtype. The density of somatostatin receptors in HCC was low compared with the density found in liver metastases of neuroendocrine tumours. The VIP receptor density was always lower in HCC than in adjacent liver tissue. CONCLUSIONS: Somatostatin, VIP, and substance P may have a receptor mediated role in HCC. Substance P receptors may be involved in regulation of tumour associated blood flow; somatostatin receptors and VIP receptors may mediate tumour growth. Diagnostic and therapeutic evaluation of somatostatin and VIP analogues may be of interest in receptor positive HCC. PMID- 10517921 TI - Transplantation of haemochromatosis liver and intestine into a normal recipient. PMID- 10517923 TI - Cerebrospinal fluid testing for the diagnosis of central nervous system infection. AB - The central nervous system (CNS) is susceptible to bacterial, viral, and fungal infections, and prion diseases. Examination of the cerebrospinal fluid (CSF) is crucial in diagnosing these infections. Cerebrospinal tests may directly identify an organism and its nucleic acid and surface constituents by culture, polymerase chain reaction (PCR), or antigen detection. Alternatively, antibody to an organism may be identified in CSF by enzyme-linked immunosorbent assay (ELISA), Western blot, or complement fixation assay. This article discusses how these CSF tests are performed and addresses the sensitivity and specificity of such tests for the diagnosis of selected CNS infections. PMID- 10517924 TI - Overview of acute and chronic meningitis. AB - Meningitis can be subdivided based on time course of onset and duration, cerebrospinal fluid (CSF) profile, and underlying origins into acute aseptic and septic meningitis, recurrent meningitis, and chronic meningitis. These are distinct syndromes that require different management strategies. Most cases of meningitis are caused by infection. The causal agent is generally predictable based on the type of meningitis, host factors, and clues from the history and examination. CSF examination remains the critical diagnostic test. PMID- 10517922 TI - Gastric epithelial dysplasia. PMID- 10517925 TI - Bacterial meningitis. AB - In recent years, investigators have made significant advances in understanding the pathogenesis of bacterial meningitis, particularly with regard to understanding the cascade of biologic events that cause excessive inflammation within the central nervous system (CNS). Nevertheless, the most important event in the field of bacterial meningitis in the past decade is the dramatic decline in the incidence of Haemophilus influenzae meningitis in children as a result of the widespread use of the conjugated H. influenzae type b vaccine. Currently, the most important clinical challenge in this field is the emergence of the drug resistant Streptococcus pneumoniae. This problem has significantly complicated initial management of patients with suspected bacterial meningitis. Preliminary data show promise with new conjugated S. pneumoniae vaccines. PMID- 10517926 TI - Central nervous system tuberculosis. AB - Involvement of the central nervous system (CNS) by Mycobacterium tuberculosis, particularly meningitis, is the most severe form of tuberculous infection. Parenchymal CNS involvement can occur in the form of tuberculoma or, more rarely, abscess. Although surgery was initially advocated as the mainstay of therapy, more recent evidence suggests that parenchymal forms of CNS tuberculosis can be cured with medical treatment alone. Also, damage of the spinal cord, roots, and spine can occur in the form of spinal meningitis, radiculomyelitis, spondylitis, or spinal cord infarction. PMID- 10517927 TI - Fungal infections of the central nervous system. AB - This review discusses a practical approach to the patient with possible fungal infection of the central nervous system (CNS). Difficulties in establishing the diagnosis come from the nonspecific clinical syndromes (subacute meningitis, meningoencephalitis, and brain abscess) and the low isolation rate of fungi from cerebrospinal fluid (CSF). Helpful diagnostic clues often come from knowledge of the patient's geographic travels, risk factors, evidence of systemic organ infection, and fungal serologic tests. Standard and new antifungal agents are evaluated and the initial and suppressive drug management of the common fungal infections is presented. PMID- 10517928 TI - Spirochetal infection of the nervous system. AB - Neurologic infection is a characteristic feature of spirochetes. The neurologic manifestations of spirochetal infection are a source of continuing public concern: Lyme neuroborreliosis in endemic areas, neurosyphilis in HIV infected patients, and neuroborreliosis during outbreaks of relapsing fever. These are reviewed in this article. The techniques for diagnosis and recommendations in the management of these infections are also discussed. PMID- 10517929 TI - Central nervous system involvement in Rickettsial diseases. AB - The Rickettsia are obligate intracellular parasites that are usually spread to humans by insects and typically produce vasculitides. The prototypic rickettsial disorder in the United States is Rocky Mountain spotted fever (RMSF). The differential diagnosis of RMSF and related disorders includes other conditions that produce vasculitis, most importantly meningococcemia. The rickettsial disorders are usually treated effectively with tetracycline derivatives. PMID- 10517930 TI - Encephalitis. AB - Encephalitis is an acute infection of brain parenchyma characterized clinically by fever, headache, and an altered level of consciousness. There may also be focal or multifocal neurologic deficits, and focal or generalized seizure activity. This article discusses the clinical presentation, diagnosis, and treatment of herpes simplex virus (HSV) encephalitis, the arthropodborne viral encephalidities, Rocky Mountain spotted fever, viral encephalitis in the immunocompromised patient, and postinfectious encephalomyelitis. PMID- 10517931 TI - Creutzfeldt-Jakob disease, new variant creutzfeldt-jakob disease, and bovine spongiform encephalopathy. AB - Creutzfeldt-Jakob disease (CJD) is a subacute spongiform encephalopathy (SSE) that is manifested by a variety of neurologic signs that usually include dementia, myoclonus, and an abnormal electroencephalogram (EEG). In 1996, a new variant of CJD (nvCJD) with a somewhat distinctive clinical presentation and neuropathology was reported in adolescents and young adults, a cohort of patients not normally affected with CJD. The appearance of nvCJD coincided temporally and geographically with the emergence of an SSE in cattle known as bovine spongiform encephalopathy (BSE), or mad cow disease. This article discusses the clinical syndrome, pathology, and pathogenesis of classical CJD, nvCJD, and other human SSEs, as well as the link between BSE and nvCJD. PMID- 10517932 TI - AIDS dementia complex. AB - Human immunodeficiency virus (HIV) infection is often complicated by the development of AIDS dementia complex (ADC). This article examines the typical and atypical presentations of ADC, along with aspects of the prevalence and natural history of the disorder. Salient aspects of the neuropathology, neurovirology, neuroimmunology, and pathogenesis are also considered. The intricacies of management of ADC, especially in the era of highly active antiretroviral therapy, are fully evaluated. Finally, this information is synthesized into an approach to the diagnosis of ADC in a particular patient. PMID- 10517933 TI - The penetration of anti-infectives into the central nervous system. AB - The blood-brain barrier, blood-cerebrospinal fluid (CSF) barrier, and meninges are a complex and difficult-to-study system charged with protecting the central nervous system (CNS) from toxins, including drugs. Current estimates of CNS drug exposure are limited to CSF to blood ratios, of which area-under-the curve (AUC) estimates provide the most robust measure of drug exposure. Different classes of drugs and individual drugs within classes have different CNS penetration potential that is dependent upon a variety of biologic and pharmacologic factors. Clinical data (AUC and point ratios) regarding the penetration of several anti infective agents used for the treatment of CNS infections are provided in this article. PMID- 10517934 TI - Disorders that mimic central nervous system infections. AB - Many noninfectious diseases can cause signs, symptoms, and cerebrospinal fluid (CSF) abnormalities simulating central nervous system (CNS) infection. Infection usually can be excluded in these cases by the judicious use of serologic tests and CSF stains and cultures. Then, the correct diagnosis is typically suggested by the history and the concomitant presence of clinical and laboratory evidence of disease in other organ systems. Occasionally, particularly when such evidence is absent, the distinction requires meningeal or brain biopsy. PMID- 10517935 TI - The descriptive epidemiology of brain tumors. AB - Brain tumors have been the subject of controversy both with respect to patterns of occurrence and to potential causes. This article provides a description of current data resources on brain tumors and outlines issues affecting the interpretation of population-based data. A template for estimating regional expected values for brain tumors is provided, and current patterns of incidence, survival, and conditional survival are described. The occurrence of second primary tumors and quality of life studies are also reviewed. PMID- 10517936 TI - The new WHO classification of brain tumors. AB - The current WHO classification greatly advances the ability to predict patient prognosis from pathologic diagnosis by introducing new tumor categories. This improved pathologic stratification is reflected in more accurate interpretations of diagnostic imaging studies. Pathology and oncology have progressed from a gross beginning, through microscopy and special stains, and into the realm of molecular biology and tumor genetics. PMID- 10517937 TI - Pathology of intracranial neoplasms. AB - This article discusses the gross and microscopic pathology of intracranial neoplasms. Primary intracranial neoplasms include tumors of the brain parenchyma; meninges; and rest of mesenchymal, epithelial, and germ cell derivation. The concept of tumor malignancy for primary central nervous system (CNS) neoplasms is somewhat different from that for systemic tumors. Although the capacity to metastasize is a defining feature of systemic malignancies and metastasis to the brain is a relatively common occurrence, primary CNS neoplasms rarely metastasize outside the CNS. PMID- 10517939 TI - Pathology of pediatric brain tumors. AB - In characterizing the pathology of brain tumors, diagnostic and prognostic immunohistochemical and molecular advances are indicated. Brain tumors are described from the perspective of pathologic type consistent with the standard World Health Organization classification as well as by neuroanatomical location with relevant differential diagnosis. Brain tumors associated with genetic syndromes are discussed. PMID- 10517938 TI - Imaging of adult brain tumors. AB - Imaging continues to play an important role in the evaluation and management of patients with primary and metastatic brain tumors. This article provides a generalized framework for evaluating brain tumors in adult patients. The goals of brain tumor imaging, the advantages of CT and MR imaging, and factors which help refine the differential diagnosis are discussed. The imaging appearance of some of the more common tumors is also discussed. PMID- 10517940 TI - Imaging of brain tumors in the pediatric population. AB - Primary brain tumors are the most common solid tumor in children and the second most common cancer after leukemia and lymphoma to affect the pediatric population. The incidence, clinical presentation, imaging features, and treatment outcomes of various pediatric brain tumors are discussed. PMID- 10517941 TI - Imaging of suprasellar and parasellar tumors. AB - A large number of entities can present as a suprasellar or parasellar mass. These include benign and malignant tumors as well as inflammatory and other lesions that can mimic a neoplasm. Imaging features and distinguishing characteristics of juxtasellar masses are described. PMID- 10517942 TI - Contrast issues in brain tumor imaging. AB - The utility of gadolinium-based MR imaging contrast agents and iodinated CT contrast agents is discussed in the context of brain tumor imaging. Although contrast materials are routinely administered for brain tumor imaging to improve lesion conspicuity and characterization, the optimal dose and imaging protocols vary according to lesion type and location, the specific clinical situation, and the imaging equipment that is available. Special problems may be encountered in contrast-enhanced imaging of patients with brain tumors who have undergone treatment with surgery, radiation, or chemotherapy. PMID- 10517943 TI - SPECT and PET imaging of brain tumors. AB - The unique capacity of measuring or visualizing intracellular biochemical processes allows nuclear medicine techniques to determine functional and metabolic activities of various disorders. This article describes the critical role of single-photon emission computed tomography (SPECT) and positron emission tomography (PET) imaging of brain tumors beyond what can be achieved by anatomic methods. PMID- 10517944 TI - Interventional neuroradiology in the treatment of brain tumors. AB - The current role of the interventional neuroradiologist in the treatment of brain tumors is limited to preoperative devascularization of vascular tumors and, occasionally, palliative therapy of metastatic or inoperable disease. Research protocols are in progress examining the role of intraarterial chemotherapy with disruption of the blood-brain barrier and may harbinger an expanded role for endovascular therapy in this group of patients. PMID- 10517945 TI - Neurosurgical management of brain tumors. AB - The combination of surgery and radiation remains the most effective treatment for tumors affecting the central nervous system. This article reviews surgical therapy for brain tumors. Special attention is paid to new approaches to brain tumor therapy and to the interaction between neuroimaging and successful surgery. PMID- 10517946 TI - Imaging of brain tumors after therapy. AB - Imaging is used routinely for the diagnosis and follow-up of patients with brain tumors. Although the latest MR imaging techniques are able to provide an excellent depiction of abnormalities in soft tissue morphology, they are frequently unable to distinguish edema and treatment-induced necrosis from recurrent or residual tumor. For assessment of response to therapy, it is therefore important to use functional imaging modalities such as PET and MR spectroscopy. PMID- 10517947 TI - Temperature effects on the discharge frequency of primary and secondary endings of isolated cat muscle spindles recorded under a ramp-and-hold stretch. AB - The effects of changes in temperature on primary and secondary endings of isolated cat muscle spindles were investigated under ramp-and-hold stretches and different degrees of pre-stretch. Temperature-induced alterations of the discharge frequency were compared over a temperature range of 25-35 degrees C. Both primary and secondary endings responded to warming with increasing discharge frequencies when the spindle was pre-stretched by 5-10% of its in situ length. The following differences between the temperature effects on primary and secondary endings were observed: (1) The temperature coefficients (Q(10)) obtained from the discharge frequencies during the dynamic and static phase of a stretch were similar for endings of the same type, but they were larger in primary endings (range of Q(10): 2.3-3.3; mean: 2.9) than in secondary endings (range of Q(10): 1.6-2.2; mean: 2.0); (2) With primary endings, but not with secondary endings, the temperature sensitivity (imp s(-1) degrees C(-1)) was larger during the dynamic phase than during the static phase of a stretch; (3) In primary endings, the fast and slow adaptive components occurring in the discharge frequency during the static phase of a stretch clearly increased with warming while in secondary endings, the slow decay was less affected, and the fast decay showed no change; (4) In relaxed spindles, the excitatory effect of warming was overlaid by a strong inhibitory effect as soon as the temperature exceeded about 30 degrees C, resulting in an abrupt cessation of the background activity in most secondary endings, but not usually in primary endings. In general, warming induced an enhanced stretch sensitivity in both types of ending, and additionally an inhibitory effect that is obvious only in secondary endings of relaxed spindles. The different effects of temperature on the discharge frequency of primary and secondary afferents are assumed to be caused by different properties of their sensory membranes. PMID- 10517948 TI - Ammonium prepulse: effects on intracellular pH and bioelectric activity of CA3 neurones in guinea pig hippocampal slices. AB - The ammonium prepulse technique was used to study influences of intracellular pH (pH(i)) on bioelectric activity of CA3-neurones in hippocampal slices. 60, 180 or 600 s lasting NH(4)Cl (10 mM) pulses led to a transient intracellular alkalosis (DeltapH(i): up to 0.2 pH-units) in about one-half of the neurones loaded with 2', 7-bis(2-carboxyethyl)-5(6)-carboxyfluorescein-acetoxymethylester (BCECF-AM). No alkalosis was seen in the remainder cells. The amount of alkalosis depended on the actual pH(i) of each neurone and increased when the pH(i) decreased. Washout of NH(4)Cl induced a fall in pH(i) (DeltapH(i): 0.12-0.54 pH-units) which recovered within <20 min. Frequency of spontaneous action potentials remained unchanged during washin of ammonium (60 or 180 s). However, pre-treatment with low concentrations of bicuculline-methiodide (0. 01 microM) or caffeine (0.1 mM), both of which did not change bioelectric activity per se, permitted a burst activity to occur during ammonium-washin in about one-half of the neurones. In all neurones, washout of ammonium inhibited spontaneous and epileptiform activity (elicited by 1 mM caffeine, 20-50 microM bicuculline-methiodide, or 50-75 microM 4-aminopyridine) for 0.98), on most of the analyzers. With the AC assay, only the results obtained with the Hitachi 717 analyzer were correlated with C-HDL values of the PTA method. The linearity and specificity studies were evaluated in the laboratory A on a Kone Specific analyzer. The alpha-CD and PA-D assays were linear for C-HDL values from 0 to 5.56 mmol/l, as observed by increasing amounts of HDL2 + HDL3 or serum without lipoprotein isolated by ultracentrifugation. The specificity of these two methods was evaluated simultaneously, by adding various amounts of lipoproteins isolated by sequential ultracentrifugation. No interference was observed when adding chylomicrons up to 13.4 mmol/l of triglycerides for both methods. Inversely, increased C-HDL values were observed with added VLDL from 6 mmol/l of triglycerides for the PA-D assay and from 8 mmol/l for the alpha-CD assay. No interference was observed with added LDL up to 11.5 mmol/l of C-LDL for the alpha-CD assay and up to 6.7 mmol/l for the PA-D assay. In conclusion, the present multicenter evaluation demonstrates that the new procedures for the direct automation of C-HDL are easy and accurate and most of them correlated well with the classical precipitation method. In addition the study provides arguments for a choice between the different direct C-HDL methods. PMID- 10518059 TI - [Effects of oral contraceptive preparations and smoking on lipoprotein repartition]. AB - We studied the effect of oral contraceptives and smoking on the lipid profile of 251 women and 72 men, 20-29-year-old. In women, taking estroprogestatives, cholesterol, triglycerides, apoproteins AI and B were higher than in controls; HDL-cholesterol was not modified. Lipoprotein analyses in polyacrylamide gradient gel exhibited an increase of the HDL3 fraction at the expense of the HDL2 fraction, with a reduced LDL size. Smoking in addition to estroprogestative absorption accentuated these modifications and led to a decreased HDL-cholesterol (HDL2 fraction essentially), with an increased LDL-cholesterol. In men, smoking resulted in higher levels of total cholesterol, apoprotein B and LDL-cholesterol, without any significant change in LDL size, higher levels of triglycerides and lower level of the HDL2 fraction without any change in HDL-cholesterol. In women, smoking led only to an increase in triglycerides. In summary, analysis of the distribution of HDL subclasses and of LDL size showed an evolution towards a supposed more atherogenic lipid profile in women taking oral contraceptives associated or not with smoking, and in male smokers. PMID- 10518060 TI - [Measurement of antiganglioside autoantibodies by immunodot-blot assay: clinical importance in peripheral neuropathies]. AB - We retrospectively evaluated measurement data and clinical relevance of autoantibodies to gangliosides in peripheral neuropathies (PN). The IgG and IgM antiganglioside autoantibodies were determined by our own immunodot-blot assay on membrane and by enzyme-linked immunosorbent assay (Elisa) in sera of 1,342 patients with peripheral neuropathies. Anti-GM1 and anti-GD1b autoantibodies formed a part of the normal autoantibody repertoire and were common place in 12% of normal subjects and in 14% of disease control groups. Polyclonal IgM antiganglioside autoantibodies were detected in chronic PN, polyclonal IgG antiganglioside autoantibodies were detected in acute PN. Polyclonal IgM anti-GM1 and anti-GD1b autoantibodies were detected in 35 patients out of 48 with treatable multifocal motor neuropathy with persistent conduction blocks. These autoantibodies well discriminated between suspected motor peripheral neuropathies and motor neuron diseases (sensitivity 73%, specificity 83%, positive predictive value 60%, negative predictive value 91%). Monoclonal IgM autoantibodies reacted strongly with gangliosides in 15 patients out of 77 with M-IgM neuropathy (19%). M-IgM autoantibodies differed in their fine specificities with different principal target antigens as demonstrated with cross-reactivity. Such findings provide further evidence for a relationship between neurological syndromes and antiganglioside antibody profiles and also suggest that different gangliosides could be principal target antigens such as GM1, GD1b, GT1b, GD1a or GM2. Polyclonal IgG anti-GM1 and anti-GD1b autoantibodies were detected in 21 patients out of 22 with acute motor axonal Guillain-Barre syndrome with antecedent of infection by Campylobacter jejuni, polyclonal IgG anti-GQ1b autoantibodies in 9 patients out of 10 with Miller-Fisher syndrome. Detection of antiganglioside autoantibodies by immunodot-blot assay which is simple and quick in testing a large panel of gangliosides has become very important in the diagnosis and in the choose of expensive therapeutic strategies in chronic or acute autoimmune neuropathies. PMID- 10518061 TI - [Detection of human papillomavirus DNA by molecular hybridization in tube: interest in cervical neoplasia]. AB - The presence of human papillomavirus (HPV) DNA in 79 cervical specimens obtained from 70 patients was studied by using a molecular hybridization technique performed in tube. The results were compared to those of the cytological and histological studies. The molecular hybridization technique in tube (Hybrid Capture I) detects two groups of HPV types. One group is highly associated with the development of cancer (types 16, 18, 31, 33, 35, 45, 51, 52, 56) whereas the second group (types 6, 11, 42, 43, 44) is not. Among 42 patients with cervical lesions before any treatment, high risk DNA of HPV was found in 50% of those with low grade cytology and 90% with high grade cytology. In total, 32 out of the 42 patients (76%) who presented histological lesions, were actually infected by HPV. Samples were obtained before and after treatment from 9 patients. Seven out of 9 presented high grade cervical intraepithelial neoplasia (CIN) and 2 other patients had low grade CIN. HPV DNA was not detected in any of the patients after treatment. Detection of HPV DNA by molecular hybridization in tube is simple, sensitive, standardized, inexpensive and is well adapted to screening programs. It can be used in complement of the cytological diagnosis, in the surveillance of equivocal cytological abnormalities, and in the follow-up of treated patients. PMID- 10518062 TI - [Assessment of P glycoprotein expression by immunocytochemistry and flow cytometry coupled with functional efflux analysis: application to acute myeloid leukemia]. AB - P glycoprotein (Pgp) expression is associated with failure of anticancer chemotherapy in acute myeloid leukemia (AML). However, a consensus has been difficult to reach, due to the variable results obtained by different methods. Samples of 27 patients with AML were studied here according to international recommendations (Beck, et al. , Cancer Research 1996; 56: 3010-20). Pgp expression was performed by immunocytochemistry (ICC) using the avidin-biotin peroxidase technique with JSB1 and UIC2 monoclonal antibodies. Flow cytometry (FCM) analysis of Pgp was investigated using UIC2 in an indirect immunofluorescent assay. UIC2 staining was measured by the Kolmogorov-Smirnov statistical test and fluorescence intensity ratio. Finally, the rhodamine 123 test (Rh 123) with or without verapamil was performed to detect functional activity. RESULTS: by ICC, results of JSB1 and UIC2 were consistent in 94% of the cases. In 74% of the cases, concordant conclusions were observed by ICC and FCM. Overall, Pgp expression was detected in 67% of the cases (ICC/JSB1+ and ICC/UIC2+ or FCM/UIC2+). Functional activity of Pgp was shown in 59% of the patients. Rh 123 efflux was correlated with Pgp expression in 70% of the 27 studied cases but 3 cases were Pgp-/Rh 123+ and 5 Pgp+/Rh 123-. In conclusion, assessment of Pgp expression by ICC and FCM using two different monoclonal antibodies coupled with functional efflux test is required to identify discordant expression/function cases suggesting a non functional Pgp or another alteration of drug transport. PMID- 10518063 TI - [Blastocystis hominis: epidemiological and clinical remarks from more than 3,500 stool examinations]. PMID- 10518064 TI - [Detection and characterization of "difficult" monoclonal components in serum. Gel-based immunofixation techniques or capillary electrophoresis]. PMID- 10518065 TI - [Evaluation of the Bio-Rad high-performance liquid chromatographic method for plasma total homocysteine assay]. PMID- 10518066 TI - [Equal free and total cortisolemia: a paradox resolved]. PMID- 10518067 TI - [Hygiene and security in laboratory: examples of actions led in quality assurance process]. PMID- 10518068 TI - Analgesia for ventilated neonates: where do we stand? PMID- 10518069 TI - Early recognition of prenatal alcohol effects: A pediatrician's responsibility. PMID- 10518070 TI - Assessing fat absorption. PMID- 10518071 TI - Overweight, central adiposity, and cardiovascular disease risk patterns in children. PMID- 10518072 TI - Treatment of Kawasaki disease: corticosteroids revisited. PMID- 10518073 TI - The tragedy of iron deficiency during infancy and early childhood. PMID- 10518074 TI - The acute chest syndrome of sickle cell disease. PMID- 10518075 TI - Morphine pharmacokinetics and pain assessment in premature newborns. AB - OBJECTIVES: To determine morphine pharmacokinetics in premature neonates varying in postconceptional age (PCA) and evaluate behavioral pain response in relationship to serum morphine concentrations. METHODS: Premature neonates (n = 48), stratified by weeks of PCA (group 1 = 24-27 weeks, group 2 = 28-31 weeks, group 3 = 32-35 weeks, and group 4 = 36-39 weeks) received morphine infusions. Blood samples were drawn at 48, 60, and 72 hours and at discontinuation of morphine, followed by 3 samples obtained during the next 24 hours. Newborns were videotaped during heel lances and restful states, with morphine at steady-state concentrations and without morphine. Pain was assessed by using the Neonatal Facial Coding System (NFCS). Statistical analysis included regression between NFCS score changes from baseline to painful procedure with and without morphine. RESULTS: Morphine clearance for groups 1, 2, 3, and 4 was calculated as 2.27 +/- 1.07, 3.21 +/- 1.57, 4.51 +/- 1.97, and 7.80 +/- 2.67 mL/kg/min, respectively, and correlated with PCA (r = 0.63, P <.001). Pain measured by facial expression was diminished; however, it did not correlate with morphine concentrations. CONCLUSION: Morphine clearance in premature neonates is less than reported, increasing with PCA. Facial activity discloses morphine analgesia; however, it is unrelated to morphine concentrations. PMID- 10518076 TI - Under-recognition of prenatal alcohol effects in infants of known alcohol abusing women. AB - Medical records of 124 women and their infants were analyzed for: (1) documentation of maternal alcohol and other substance abuse and (2) evaluation of exposed infants. These results were compared with the study interview and infant examination. More obstetric nurses documented the presence or absence of alcohol and substance abuse than did pediatricians. More women reported using alcohol in the study interview than documented in the medical records. There was slightly better documentation for cocaine use than for alcohol use in the medical records. One of the 19 infants with documentation of maternal alcohol use was noted to have possible alcohol-related features by the pediatrician, in contrast to 7 infants identified by the study examiner. In addition, 2 of these 19 infants were determined by the study examiner to have fetal alcohol syndrome; neither case was diagnosed by the pediatricians. Continued efforts at education regarding the importance of asking about prenatal alcohol exposure and the spectrum of fetal alcohol effects are needed for early diagnosis. PMID- 10518077 TI - Sudden unexpected deaths in infancy: what are the causes? AB - OBJECTIVES: To determine the incidence of various causes of sudden unexpected death (SUD) within an entire population and to assess the relative importance of an expert autopsy, as well as age of demise, in predicting the likelihood of finding a cause of death. METHODS: We reviewed all cases of SUD in infants aged 1 week to 18 months that occurred in the province of Quebec (Canada) between 1987 and 1996. RESULTS: We identified 623 cases of SUD; in 80% the diagnosis was sudden infant death syndrome (SIDS). Infection was the most common non-SIDS diagnosis (7.1% of all SUDs), followed by cardiovascular anomalies (2.7%), child abuse or negligence (2.6%), and metabolic or genetic disorders (2.1%). The percentage of non-SIDS deaths was much higher for autopsies performed in centers with expertise in pediatric pathology (18% vs 6%, P <.005). The likelihood of a non-SIDS diagnosis was much higher at age ranges atypical, as compared with typical, for SIDS (33% at 7 to 27 days, 19% at 6 to 12 months, and 64% at 12 to 18 months [atypical] vs 15% at 1 to 6 months [typical]; P =.003). CONCLUSION: The study of an entire population provides more reliable data regarding causes of SUDs than does the study of small groups. We recommend that in addition to a thorough investigation of each SUD, autopsies be performed in centers with expertise in pediatric pathology. This recommendation takes on added significance in this era of decreasing SIDS rates. PMID- 10518078 TI - Validation in an animal model of the carbon 13-labeled mixed triglyceride breath test for the detection of intestinal fat malabsorption. AB - OBJECTIVE: To determine, in a rat model of fat malabsorption, the potency of the carbon 13-labeled mixed triglyceride ((13)C-MTG) breath test as a noninvasive, patient-friendly replacement for classic fat balance studies. STUDY DESIGN: Comparison of the percentage of fat absorption, detected by fat balance, with the (13)CO(2) recovery of the (13)C-MTG breath test in rats fed high-fat chow and varying amounts of the lipase inhibitor, orlistat (0, 50, 200, and 800 mg per kilogram of chow), for 5 days. RESULTS: On orlistat administration, total fat absorption decreased from 80.2% +/- 2.2% to 32.8% +/- 3.7% (mean +/- SEM, 0 mg and 800 mg of orlistat per kilogram of chow, respectively; P <.001). Correspondingly, breath (13)CO(2) recovery from (13)C-MTG at 6 hours decreased from 84.5% +/- 7.8% to 42.0% +/- 1.5% of the dose (0 mg and 800 mg of orlistat per kilogram of chow, respectively; P <.001). The 6-hour recovery of breath (13)CO(2) appeared to be highly correlated with the percentage of fat absorption (r = 0.88, P <.001). In rats with fat absorption higher than 70%, however, the coefficient of variation of the (13)C-MTG breath test was 3-fold larger than that of the fat balance. CONCLUSIONS: The (13)C-MTG breath test could potentially replace the fat balance method for comparing fat absorption efficacy between groups. Yet, a considerable interindividual variation of the (13)C-MTG breath test under conditions of relatively mild fat malabsorption does not support its application for diagnostic purposes in individuals. PMID- 10518079 TI - Overweight, fat patterning, and cardiovascular disease risk factors in black and white girls: The National Heart, Lung, and Blood Institute Growth and Health Study. AB - OBJECTIVE: To determine the association of overweight and central adiposity with cardiovascular disease risk factors in black and white 9- and 10-year-old girls. DESIGN: Cross-sectional analysis of baseline data collected from participants in the National Heart, Lung, and Blood Institute Growth and Health Study. Girls were classified as overweight or not with the use of the age- and sex-specific 85th percentiles of the body mass index (kilograms per square meter) distributions from the combined NHANES (I and II) data set. Mean indexes of central adiposity, blood pressure levels, and lipid concentrations and the clustering of risk factors based on published cut points were compared between weight groups by race and by central adiposity group within weight and race groups. RESULTS: Overweight was associated with increased risk factor levels and with increased clustering in both black and white girls. Among overweight girls greater central adiposity was associated with higher risk factor levels and increased clustering. CONCLUSIONS: Given the associations between cardiovascular disease risk factors and both overweight and central adiposity, the secular trends toward increased obesity in American youth portend a worsening of cardiovascular disease risk profiles. PMID- 10518080 TI - Corticosteroids in the treatment of the acute phase of Kawasaki disease. AB - OBJECTIVES: Corticosteroids are considered to be contraindicated during the acute phase of Kawasaki disease (KD) based on unfavorable results in early studies. In our hospital, however, corticosteroids have been used in some cases of KD with satisfactory results. We analyzed outcomes of patients with KD treated with or without corticosteroids. STUDY DESIGN: Medical records of 299 children with KD treated with one of the 4 regimens were reviewed retrospectively. Regimen 1 consisted of aspirin, dipyridamole, and propranolol; regimen 2 was regimen 1 plus prednisolone, 2 mg/kg/d, for 1 week, followed by tapering over 2 weeks; regimen 3 was regimen 1 plus intravenous gamma-globulin (IVGG), 200 or 400 mg/kg/d, for 5 consecutive days; and regimen 4 was regimen 1 plus both prednisolone and IVGG. RESULTS: Although patients treated with regimens 2 and 4 were more ill at presentation than those treated with regimens 1 and 3, respectively, the duration of fever was shorter in the former patient groups (P =.0013). Coronary aneurysms developed least frequently in patients treated with regimen 4 and less frequently with regimen 2 than with regimen 1 (P =.0730). Multiple regression analysis showed significant reductions of fever and coronary aneurysm incidence with prednisolone (P <.0001 and P =.0307, respectively). CONCLUSION: Our data suggest a possible role of corticosteroids in the treatment of the acute phase of KD. PMID- 10518081 TI - The effects of long-term growth hormone treatment on cardiac left ventricular dimensions and blood pressure in girls with Turner's syndrome. Dutch Working Group on Growth Hormone. AB - OBJECTIVE: To assess the effects of long-term growth hormone (GH) treatment for short stature on left ventricular (LV) dimensions and systemic blood pressure (BP) in girls with Turner's syndrome without clinically relevant cardiac abnormalities. STUDY DESIGN: LV dimensions measured by echocardiography and systemic BP were assessed before and during 7 years of GH treatment in 68 girls with Turner's syndrome participating in a randomized dose-response study. These previously untreated girls, age 2 to 11 years, were randomly assigned to 1 of 3 GH dosage groups: group A, 4 IU/m(2)/d; group B, first year 4 IU/m(2)/d, thereafter 6 IU/m(2)/d; group C, first year 4 IU/m(2)/d, second year 6 IU/m(2)/d, thereafter 8 IU/m(2)/d. After the first 4 years, girls >/=12 years of age began receiving 17beta-estradiol, 5 microg/kg body weight per day, for induction of puberty. RESULTS: At baseline the LV dimensions of almost every girl were within the normal range, and the mean SD scores were close to zero. During 7 years of GH treatment, the growth of the left ventricle was comparable to that of healthy girls. No signs of LV hypertrophy were found. Before the start of GH treatment, mean BP was within the normal range but significantly higher than in healthy control subjects. Diastolic BP and systolic BP were above the 90th percentile in 23% and 28% of the girls, respectively. After 7 years of treatment, these percentages were 14% and 36%, respectively (not significantly different from baseline). The SD score of the diastolic BP showed a small decrease after 7 years of treatment. The growth of the left ventricle and the development of BP were not different between the GH dosage groups. CONCLUSIONS: Long-term GH treatment, even at dosages up to 8 IU/m(2)/d, does not result in LV hypertrophy or hypertension in girls with Turner's syndrome. Continued observation into adulthood is recommended to monitor the further development of the relatively high BP and to ensure that GH treatment has no long-term negative effect on the heart. PMID- 10518082 TI - Acquired protein S deficiency caused by estrogen treatment of tall stature. AB - OBJECTIVE: To evaluate the potential thrombogenic changes in the coagulation and fibrinolytic system related to treatment with ethinyl estradiol (200 and 300 microg). SUBJECTS AND METHODS: Twenty-five healthy girls with expected final height exceeding 185 cm, as calculated by the method of Bayley and Pinneau, were treated with 200 microg or 300 microg of ethinyl estradiol. Coagulation and fibrinolytic parameters were determined before and during estrogen treatment and 2 and 4 weeks after estrogen withdrawal. RESULTS: No difference in the effects on hemostasis was found between the 2 treatment groups. All 25 patients developed protein S deficiency during estrogen treatment, which in most girls lasted for 4 weeks after cessation of estrogen administration. During therapy, protein C activity increased, whereas antithrombin did not change. Plasminogen and plasmin alpha(2) antiplasmin complexes significantly increased. Protein S deficiency was accompanied by significantly increased prothrombin fragment 1+2 and fibrinopeptide A. In contrast, thrombin-antithrombin complexes did not change. CONCLUSION: High-dose estrogen treatment to reduce the final height in tall girls is associated with a reversible acquired protein S deficiency with indications of a pre-thrombotic state. Risk of venous thrombo-embolism may be enhanced, especially when additional risk factors for thrombosis are present. PMID- 10518083 TI - Liver disease in Navajo neuropathy. AB - OBJECTIVE: To describe clinical and histologic features of liver disease in infants and children with Navajo neuropathy (NN). METHODS: Physicians at Navajo Area Indian Health Service facilities and neurologists and gastroenterologists at regional referral hospitals were surveyed for identification of patients born between 1980 and 1994 with known or suspected NN. Clinical records and liver histologic findings were reviewed. RESULTS: Liver disease was present in all children with NN. Three clinical phenotypes of NN were observed, based on age at presentation and course: infantile NN presented in 5 infants before 6 months of age with jaundice and failure to thrive and progressed to liver failure before 2 years of age; childhood NN presented in 6 children between 1 and 5 years of age with liver dysfunction, which progressed to liver failure and death within 6 months; and classical NN presented in 9 children with variable onset of liver disease but progressive neurologic deterioration. Liver histologic findings were characterized by multinucleate giant cells, macrovesicular and microvesicular steatosis, pseudo-acini, inflammation, cholestasis, and bridging fibrosis and cirrhosis. Cases of all 3 phenotypes occurred within the same kindred. CONCLUSIONS: Liver disease is an important component of NN and may be the predominant feature in infants and young children. We propose changing the name of this disease to Navajo neurohepatopathy. PMID- 10518084 TI - Orthostatic intolerance and chronic fatigue syndrome associated with Ehlers Danlos syndrome. AB - OBJECTIVE: To report chronic fatigue syndrome (CFS) associated with both Ehlers Danlos syndrome (EDS) and orthostatic intolerance. STUDY DESIGN: Case series of adolescents referred to a tertiary clinic for the evaluation of CFS. All subjects had 2-dimensional echocardiography, tests of orthostatic tolerance, and examinations by both a geneticist and an ophthalmologist. RESULTS: Twelve patients (11 female), median age 15.5 years, met diagnostic criteria for CFS and EDS, and all had either postural tachycardia or neurally mediated hypotension in response to orthostatic stress. Six had classical-type EDS and 6 had hypermobile type EDS. CONCLUSIONS: Among patients with CFS and orthostatic intolerance, a subset also has EDS. We propose that the occurrence of these syndromes together can be attributed to the abnormal connective tissue in dependent blood vessels of those with EDS, which permits veins to distend excessively in response to ordinary hydrostatic pressures. This in turn leads to increased venous pooling and its hemodynamic and symptomatic consequences. These observations suggest that a careful search for hypermobility and connective tissue abnormalities should be part of the evaluation of patients with CFS and orthostatic intolerance syndromes. PMID- 10518085 TI - Diversity in presenting manifestations of systemic lupus erythematosus in children. AB - OBJECTIVE: To describe the diversity in presenting manifestations of systemic lupus erythematosus (SLE) in children. STUDY DESIGN: Initial clinical and laboratory manifestations of 39 children, who fulfilled >/=4 American College of Rheumatology criteria for SLE, were retrospectively analyzed. RESULTS: Median age at onset was 12 years. The male to female ratio was 1:18.5, and racial/ethnic backgrounds were white 41%, black 33%, and Hispanic 26%. Initial manifestations included musculoskeletal 74%, cutaneous 72%, constitutional 67%, neurologic 28%, renal 28%, lymphadenopathy 15%, and Raynaud's phenomenon 10%. Laboratory abnormalities at presentation to our clinic included elevated erythrocyte sedimentation rate 87%, anemia 72%, lymphopenia 59%, leukopenia 31%, proteinuria or cellular casts 44%, low C(3) or C(4) level 77%, antinuclear antibodies 97%, and anti-double-stranded DNA 95%. One third (33%) presented with features not initially suggestive of SLE. Six patients presented with unusual manifestations including parotitis, quadriplegia, chorea, severe abdominal pain, persistent cough, and dizziness. However, 85% of patients with atypical manifestations had abnormal complete blood count or urinalysis results at presentation. CONCLUSION: Presenting manifestations of SLE in children are diverse. A detailed history, thorough review of systems, complete physical examination, complete blood count, urinalysis, and a high index of suspicion help to make the correct diagnosis of SLE in patients with atypical presentations. PMID- 10518086 TI - Cow's milk challenge through human milk evokes immune responses in infants with cow's milk allergy. AB - OBJECTIVES: In order to measure the immune response evoked in breast-fed infants with cow's milk allergy (CMA) by cow's milk challenge through human milk, mothers were given increasing doses of cow's milk after they had been on a cow's milk elimination diet. Another objective was to study the secretion of beta lactoglobulin (BLG) into human milk before and during milk challenge in relation to the appearance of symptoms in infants. STUDY DESIGN: Seventeen asymptomatic mothers who had infants with challenge-proven CMA and 10 asymptomatic mothers who had healthy infants were recruited. Infants ranged in age from 1.8 to 9.4 months. A solid-phase enzyme-linked immunoassay (ELISPOT) was used to assess the total number of immunoglobulin-secreting and specific antibody-secreting cells. Flow cytometry was used to enumerate different lymphocyte subpopulations among peripheral blood lymphocytes primed during provocation by cow's milk antigens. BLG levels were assessed in human milk before the challenge and 1, 2, 3, and 4 hours after the commencement of the challenge. RESULTS: All but one of the infants with CMA showed symptoms of CMA during cow's milk challenge through human milk. There was a significant rise in the total number of immunoglobulin secreting cells in the IgA and IgG classes associated with a positive cow's milk challenge response, but the proportions of peripheral blood B cells bearing CD19, CD23, CD19 and 23, CD5, or CD19 and CD5 were comparable. BLG levels were comparable in both study groups. CONCLUSIONS: Most of the infants with CMA reacted to cow's milk challenge through human milk. Hypersensitivity reactions to food antigens through human milk may be more common than previously thought. PMID- 10518087 TI - Ehlers-Danlos syndrome. PMID- 10518088 TI - Severe iron deficiency anemia in young children. AB - We retrospectively characterized clinical features of 55 patients with severe nutritional iron deficiency anemia. Anemia was commonly discovered in the absence of related complaints. Forty percent of patients were of Southeast Asian ancestry. Most were treated successfully with iron therapy alone; 8 required transfusion. PMID- 10518089 TI - Characterization of esophageal body and lower esophageal sphincter motor function in the very premature neonate. AB - OBJECTIVES: To characterize esophageal body and lower esophageal sphincter (LES) motor function in very premature infants. STUDY DESIGN: Esophageal manometry was performed in 12 very premature infants of 26 to 33 weeks' postmenstrual age (PMA) (body weights of 610-1360 g). Esophageal motor patterns were recorded for 30 minutes with a perfused micromanometric sleeve assembly (outer diameter, 2.0 mm). RESULTS: Esophageal pressure waves triggered by dry swallows were predominantly (84%) peristaltic in propagation sequence. All infants showed tonic LES contraction; the mean resting LES pressure (LESP) for individual infants ranged from 5.0 +/- 4.1 mm Hg to 20.0 +/- 4.8 mm Hg. In all infants the LES relaxed (duration, 5.8 +/- 3.0 seconds; nadir pressure, 1.8 +/- 2.6 mm Hg) in response to pharyngeal swallows. Transient LES relaxations (TLESRs) (duration, 21.7 +/- 8.7 seconds; nadir pressure, 0.1 +/- 1.8 mm Hg) occurred on average 2.6 +/- 1.6 times per study; 86% of these relaxations triggered esophageal body common cavity events known to be associated with gastroesophageal reflux. CONCLUSIONS: Esophageal motor function is well developed in very premature infants. Our data also suggest that TLESR is the predominant mechanism of reflux in these babies. PMID- 10518090 TI - Lower esophageal sphincter position in premature infants cannot be correctly estimated with current formulas. AB - OBJECTIVES: Strobel's formula (Esophageal length = 5 + 0.252 x Height) is frequently used as a guide for determining the distance from the nares to the lower esophageal sphincter (LES) in term infants. The aim of this study was to examine this relationship in premature infants. STUDY DESIGN: The distance from nares to LES was manometrically determined in 156 premature infants (26-40 weeks' postmenstrual age; body weights of 610-3050 g). The ability of body weight, height (body length), head circumference, and postmenstrual age to predict the manometrically determined LES position was evaluated with linear and non-linear regression analyses. RESULTS: Body weight and body length were the most predictive of distance from nares to LES (r(2) = 0.848 and 0.802, respectively). These relationships were non-linear and, in the case of body length, deviated substantially from Strobel's model. CONCLUSIONS: In premature neonates, a different formula is needed for prediction of the distance between nares and LES than that applied to term infants and children. PMID- 10518091 TI - Methylxanthine therapy in premature infants: sound practice, disaster, or fruitless byway? PMID- 10518092 TI - Insights. Pylephlebitis. PMID- 10518093 TI - Etidronate in subcutaneous fat necrosis of the newborn. PMID- 10518094 TI - T lymphocytes and vitamins. PMID- 10518095 TI - The first reported case of Kawasaki disease? PMID- 10518097 TI - Triad Clinical Cancer Control Program: A partnership for data-based community intervention. PMID- 10518096 TI - Overweight, fat patterning, and cardiovascular disease risk factors in black and white boys. AB - PURPOSE: To evaluate the relationships of overweight and fat patterning with cardiovascular disease (CVD) risk factors in black and white boys. DESIGN: Cross sectional analysis of CVD risk factors by weight and central adiposity groups in black and white boys, aged 10 to 15 years. Mean adiposity, lipid, and blood pressure variables were compared between weight and central adiposity groups within race by using linear regression models. Observed clustering of risk factors within weight and adiposity groups was compared with the expected clustering under an assumption of no association. RESULTS: Within each racial group, overweight boys had greater skinfolds, lower high-density lipoprotein cholesterol levels, higher low-density lipoprotein cholesterol and triglyceride levels, and higher systolic and diastolic blood pressure than non-overweight boys. Among overweight boys, greater central adiposity was associated with higher risk factor levels and increased clustering of risk factors. CONCLUSION: Overweight and central adiposity together profoundly affect CVD risk factor levels and risk factor clustering in black and white boys. PMID- 10518098 TI - Reviews in pelvic surgery PMID- 10518099 TI - Peritoneal washing cytology: prognostic value of positive findings in patients with gastric carcinoma undergoing a potentially curative resection. AB - BACKGROUND AND OBJECTIVES: Free cancer cells in the abdominal cavity exfoliated from a tumor are considered to be responsible for peritoneal dissemination, the most frequent pattern of failure in gastric carcinoma patients treated with curative surgery. METHODS: A prospective survival analysis was performed with 91 gastric carcinoma patients treated by potentially curative resection. Cytology was performed for all the patients. The method of Kaplan and Meier was used to construct curves with diagnosis of peritoneal dissemination and cancer death as the end points. Multivariate analysis by Cox's proportional hazards model was performed to identify independent prognostic factors of significance. RESULTS: Patients with a positive cytology result were confirmed to have a greater risk for recurrence in the pattern of peritoneal carcinomatosis and hence a significantly inferior prognosis. Positive cytology was the only significant independent prognostic factor among the curatively resected patients with advanced gastric carcinoma. CONCLUSIONS: Peritoneal lavage cytology should be employed for all advanced cancer undergoing potentially curative resection for added accuracy in the stage classification. The results should also reflect the eligibility of the patients for future clinical trials involving perioperative intraperitoneal chemotherapy. PMID- 10518100 TI - Commentary PMID- 10518101 TI - Intraoperative electron beam radiotherapy in recurrent colorectal carcinoma. AB - BACKGROUND AND OBJECTIVES: The installation of a dedicated linear accelerator in a shielded operating room in 1992 allowed us to start a feasibility study of intraoperative electron beam radiation therapy (IOERT) in colorectal carcinoma. METHODS: From March 1992 to February 1996, 28 patients with recurrent colorectal carcinoma were treated with maximal surgical resection and IOERT to the pelvis (n = 20) or paraortics (n = 8). IOERT dose ranged from 10 to 20 Gy with electron energies of 6-15 MeV. Postoperative external beam radiation therapy (EBRT) of 45 50 Gy was planned for the previously unirradiated patients. RESULTS: IOERT was well tolerated, but 10 (70%) of 13 patients in the previously unirradiated group did not complete the EBRT per protocol. Eight patients (29%) had some morbidity including surgically related fistula distal from IOERT sites. Two patients developed pelvic pain, which can be attributed to IOERT. Three-year local control at sites treated with IOERT was 40% (53% for previously irradiated patients and 27% for previously unirradiated patients). The 3-year actuarial overall survival was 12% (17% for previously irradiated patients and 8% for previously unirradiated patients). CONCLUSIONS: Our initial experience showed that it was feasible to treat poor prognostic colorectal cancer patients with IOERT. The morbidity observed was mainly related to extensive surgery in high-risk patients. Poor local control was obtained in patients treated with low-dose IOERT alone. Hence, previously unirradiated patients are encouraged to complete the planned EBRT or, alternatively, are considered for EBRT preoperatively or are given a higher IOERT dose (up to 20 Gy) if EBRT will not be given. Since IORT doses >20 Gy are associated with nerve toxicity, we currently add limited dose EBRT in the previously irradiated group. Patients with disease located in multiple abdominal sites are no longer considered candidates for IOERT. PMID- 10518102 TI - Ultrasound-guided lumpectomy of nonpalpable breast cancers: A feasibility study looking at the accuracy of obtained margins. AB - BACKGROUND AND OBJECTIVES: Complete excision of a nonpalpable breast cancer after wire localization is a difficult procedure. Often, adequate margins are not obtained, and a second procedure is then required. Prospectively, we studied the feasibility of ultrasound-guided excisions of nonpalpable breast cancers, with particular attention to the accuracy of the procedure in obtaining adequate margins. METHODS: Prospectively, 19 patients with 20 mammographically detected nonpalpable, highly suspect, breast tumors were entered in this feasibility study. In 15 of these, the diagnosis of invasive malignancy was established preoperatively. All patients underwent ultrasound-guided excision with the intent to obtain adequate margins. We also reviewed our own experience with the excision of nonpalpable breast cancers after wire localization. RESULTS: Of the 20 excisions with ultrasound guidance, there were 19 carcinomas and 1 ductal carcinoma in situ. Of the 19 carcinomas, 17 (89%) were excised with adequate margins. Of the 43 carcinomas that were excised after wire localization, only 17 (40%) had been resected with adequate margins. CONCLUSIONS: Ultrasound-guided excision appears to be a reliable procedure for obtaining adequate margins in the resection of nonpalpable breast cancers. Other advantages of this procedure are increased patient comfort and decrease in operating room time. PMID- 10518103 TI - Comparison between a 2- and 3-grade system in predicting metastatic-free survival in extremity soft-tissue sarcoma. AB - BACKGROUND AND OBJECTIVES: The purpose of this study was to determine whether a histologic grading system consisting of 2 or 3 categories had better discrimination for predicting metastasis-free survival in extremity soft-tissue sarcoma. METHODS: One hundred thirty patients with nonmetastatic soft-tissue sarcoma were identified and the histologic grade (3-grade system) for each tumor was determined. For the 2-grade system, grade was determined by collapsing 3 grades into 2. The Kaplan-Meier method was used to estimate disease free survival. RESULTS: By use of a 3-grade system, grade 2 tumors showed a 5.2-fold and grade 3 tumors a 9-fold increased risk of systemic relapse when compared with grade 1 tumors. When grade 2 and 3 tumors were combined, they had a 2.6-fold increased risk of systemic relapse compared with grade 1 tumors. When grade 1 and 2 tumors were combined, grade 3 tumors had an 8.4-fold risk of relapse. After data were controlled for size and depth of tumor, each increase in grade in the 3 grade system showed a successive 2.3-fold increase in risk of systemic relapse. CONCLUSIONS: A 3-grade system may be more appropriate for predicting systemic relapse than 2 grades. A prospective study is required to confirm this. PMID- 10518105 TI - Prognostic factors after extended esophagectomy for squamous cell carcinoma of the thoracic esophagus. AB - BACKGROUNDS AND OBJECTIVES: In Japan, extended esophagectomy with extensive lymphadenectomy has become the standard surgical procedure for carcinoma of the thoracic esophagus. Although mortality and morbidity rates after such extensive esophagectomy have been acceptable, the long-term outcomes are not necessarily satisfactory. METHODS: Among 235 patients with primary squamous cell carcinoma of the thoracic esophagus between June 1981 and March 1998, 143 patients (60.9%) underwent extended esophagectomy with extensive lymphadenectomy. To exclude the effects of surgery-related postoperative complications, 14 patients who died within 90 days after operation were excluded. Thus, clinicopathological characteristics and prognostic factors of 129 patients were retrospectively investigated. RESULTS: Sixty-three patients were alive and free of cancer. Sixty six patients died: 37 of recurrence of the esophageal cancer and 29 of other causes. The 1-, 3-, 5-, and 10-year overall survival rates in the 129 patients were 78.8%, 53.5%, 45.8%, and 30.9%, respectively, and the disease-specific survival rates were 85.7%, 69.1%, 67.9%, and 56.2%, respectively. The factors influencing the disease-specific survival rate were tumor location (upper third vs. non-upper third), Borrmann classification (0, 1 vs. 2, 3), size of tumor (3.0 cm), depth of invasion (T1, 2 vs. T3, 4), number of lymph node metastases (0 or 1 vs. >/=2), time of operation (420 min), amount of blood transfused (/=3 U), lymph vessel invasion (marked vs. not marked), and blood vessel invasion (marked vs. not marked). Among those significant variables, independent prognostic factors for survival determined by multivariate analysis were number of lymph node metastases (P < 0.001), amount of blood transfusions (P = 0.0016), and tumor location (P = 0.0382). CONCLUSIONS: Patients with a single metastatic node after extended esophagectomy should be considered to have excellent prognosis, like patients with pN0 tumors. Patients with multiple involved nodes should receive aggressive postoperative adjuvant treatments. Reduced blood loss during extended esophagectomy and minimal blood transfusions might improve the outcome of curative esophageal resections. PMID- 10518104 TI - Liver fatty acid-binding protein is a new prognostic factor for hepatic resection of colorectal cancer metastases. AB - BACKGROUND AND OBJECTIVES: Liver fatty acid-binding protein (L-FABP) is reported as a biological marker for enterocytic differentiation. We evaluated the prognostic value of L-FABP expression for patients undergoing hepatic resection of colorectal cancer metastases. METHODS: The study group comprised 68 patients who underwent hepatic resection for colorectal cancer metastases between 1982 and 1996 at Niigata University Medical Hospital, Niigata, Japan. L-FABP expression was immunohistochemically studied in metastatic liver tumors and their primary colorectal cancers. The relationship between L-FABP expression and patient prognoses was statistically analyzed. RESULTS: L-FABP was positively stained in 56% (38/68) of liver metastases from colorectal cancers and in 56% (38/68) of their primary tumors. Of 68 cases, 54 (79%) showed similar immunohistochemical findings between primary and metastatic tumors. Patients with L-FABP-positive liver metastases showed better prognosis than patients with L-FABP-negative metastases (P = 0.046). L-FABP expression in primary colorectal cancers more significantly (P = 0.009) affected long-term survival after hepatic surgery. Multivariate analysis revealed that the prognostic effect of L-FABP expression in primary colorectal cancers was exerted independently and that its impact was larger than conventional pathological prognosticators. CONCLUSIONS: L-FABP expression is suitable for use as a new presurgical prognostic factor for patients undergoing hepatic surgery for colorectal cancer metastases. PMID- 10518106 TI - No prognostic significance of p53 expression in esophageal squamous cell carcinoma. AB - BACKGROUND AND OBJECTIVES: It is generally accepted that the overexpression of p53 protein is associated with poor prognosis in breast, colorectal, and other types of cancer. However, the prognostic significance of p53 aberrations in esophageal squamous cell carcinoma has yet to be determined. We attempted to analyze the relationship between p53 expression and the clinicopathologic features of esophageal squamous cell carcinoma by reviewing the medical records of a large patient population. Our study of esophageal squamous cell carcinoma involves the largest patient population to date. METHODS: p53 expression in formalin-fixed, paraffin-embedded samples of 239 patients with primary esophageal squamous cell carcinoma (TNM stage I:79 cases, stage II: 82 cases, stage III: 78 cases), who underwent esophageal resection without additional treatment, were analyzed by immunohistochemical staining using a polyclonal antibody, RSP53. The relationships between p53 immunoreactivity and prognostic factors were determined by the chi(2) test, and the prognostic impact of p53 protein expression was analyzed by univariate and multivariate survival analyses. RESULTS: In 115 (48.1%) of 239 esophageal tumors, nuclear immunoreactivity for the p53 protein was detected. The expression of the p53 protein did not correlate with sex, age, histological grading, lymph node metastasis, vascular invasion, or TNM stage. Similarly, p53 expression did not correlate with prognosis in univariate and multivariate survival analysis. CONCLUSIONS: The expression of the p53 gene product had no impact on the prognosis of esophageal squamous cell carcinoma. PMID- 10518107 TI - Early detection of hepatocellular carcinoma in hepatitis-B-positive renal transplant recipients. AB - Hepatocellular carcinoma (HCC) is a leading cause of malignancy after renal transplantation in Asia, where hepatitis B virus infection is endemic. Early detection and resection are the key to successful treatment because the mortality rate for HCC is high. The value of alpha-fetoprotein monitoring in the early detection of HCC in renal transplant recipients has not been reported before. We describe 2 patients who had successful resection of HCC following early diagnosis by alpha-fetoprotein monitoring. The epidemiology of post-transplant HCC in various parts of the world and its pathogenesis are discussed. PMID- 10518108 TI - Exenterative pelvic surgery. AB - A review of the history, indications, basic technique, end results, and complications of exenterative surgery for pelvic neoplasms is provided. The authors discuss their broad personal experience with the operation. Much of this experience evolved from work at Barnes Hospital and the Ellis Fischel State Cancer Hospital. The techniques are applicable to advanced neoplasms of the cervix uteri, scrotum, urinary bladder, and other, less frequent neoplasms still confined to the pelvis. PMID- 10518109 TI - Regulatory T cells in experimental allergic encephalomyelitis. II. T cells functionally antagonistic to encephalitogenic MBP-specific T cells show persistent expression of fasL. AB - Naive LEW rats and animals that have recovered from active or adoptive EAE contain a subset of T cells that inhibit EAE in secondary recipients and are lytic for MBP-reactive T cell lines in culture. Here we explore various features of these regulatory T cell populations (RTC) with respect to (1) their frequency in animals following immunization with syngeneic MBP-reactive cell lines, (2) their ability to inhibit proliferative responses by MBP-reactive cell lines in culture, (3) their ability to lyse MBP-specific target cells, and especially (4) their prolonged expression of high levels of FasL following activation in culture correlating with their lytic effects on A20, a FasL-sensitive mouse lymphoma cell line. Inhibition studies indicate that mAbs specific for MHC class I and MHC class II molecules inhibit lysis of syngeneic MBP-reactive target T cells, soluble Fas protein shows some inhibition, but none of these agents inhibits the lytic effects of activated RTC on the A20 cell line. PMID- 10518110 TI - Pericytes and periendothelial cells of brain parenchyma vessels co-express aminopeptidase N, aminopeptidase A, and nestin. AB - Within the parenchyma of the CNS, the endothelium of all vessels is surrounded by a layer of cells, pericytes in capillaries and periendothelial or intima smooth muscle cells in other vessels. The origin of these cell types, their relationship, and their role are unclear. However, it has been recently shown that genetically engineered mice that lack pericytes develop microaneurysms at late gestation and die before birth (Lindahl et al. [1997] Science 277:242-245). The goal of this study was to identify in situ molecular markers that would be common to pericytes and periendothelial cells of adult mouse brain. Immunocytochemistry experiments were carried out at the optical and electron microscopic levels on mouse brain sections with antibodies specific for aminopeptidase N, aminopeptidase A, and the intermediate filament nestin. The results of our experiments show that in all brain parenchyma vessels of all sizes, pericytes and periendothelial cells are immunoreactive for aminopeptidase N, essentially at the plasma membrane level, and are also labeled by nestin specific antibodies, which decorate typical intermediate filaments. In addition, brain pericytes and periendothelial cells are also immunoreactive to monoclonal antibodies to aminopeptidase A. In contrast, pericytes and periendothelial cells do not express microglial markers. Taken together these data show that pericytes and periendothelial intima smooth muscle cells share common markers, suggesting a common origin or function, and are distinct from microglia. PMID- 10518111 TI - Spatiotemporal patterns of microglial proliferation in rat brain injured at the postmitotic stage of postnatal development. AB - Changes in the number and distribution of microglial cells proliferating in response to unilateral injury of the cerebral hemisphere were investigated in 30 day-old rats. Twelve hours or 1, 2, 4, or 8 days following the injury the rats were injected with (3)H-thymidine and killed 4 hr later. Brain sections were processed for BSI-B4 isolectin histochemistry followed by autoradiography. During microscopic observations, four morphological types of lectin-positive and autoradiographically labeled cells were distinguished: 1) ramified, 2) hypertrophic, 3) bushy, and 4) macrophages. Subsequently, numbers and locations of the cell types within the injury area were recorded at different stages of the experiment. The earliest signs of the proliferative response were displayed 12 hr after injury by ramified microglia. On the first day after injury, those cells represented about 50% of the whole cell population and were spread at relatively the longest distances from the lesion site. During subsequent stages of the response, a considerable reduction of the area occupied by proliferating microglia corresponded with a dramatic quantitative decrease of their ramified fraction and a simultaneous increase of their more advanced reactive forms and macrophages. PMID- 10518112 TI - Mannose receptor is present in a functional state in rat microglial cells. AB - We studied the expression of the mannose receptor (ManR) in rat microglial cells. Microglial cells are the central nervous system resident macrophages, key participants of the innate immune response. ManR is a differentiation marker and a relevant glycoprotein for the phagocytic and endocytic function of macrophages. Because there is evidence suggesting that ManR could mediate some of the nonenzymatic effects of acetilcholinesterase (AchE) and the enzyme seems to be involved in Alzheimer's disease (AD), we looked for ManR in microglia, evaluating the functionality of the receptor. We isolated microglial cells from the brain of 2-day-old neonatal rats. Microglial cells, identified by their specific staining with the lectin Griffonia simplicifolia, expressed ManR, being detected by immunocytochemistry, Western blot, and immunoprecipitation. Microglial ManR was downregulated by lipopolysaccharide (LPS) and upregulated by dexamethasone, as described for peripheral macrophages. Microglial ManR was functional and able to internalize horseradish peroxidase (HRP), a known ManR ligand, in a mannan inhibitable manner. The presence of a functional ManR in microglia opens the possibility that ManR could participate in multiple physiologic and pathologic conditions in the central nervous system (CNS), including inflammation, ischaemia, and neurodegenerative diseases such as AD. PMID- 10518113 TI - Up-regulation of cyclin D1 occurs in apoptosis of immature but not mature cerebellar granule neurons in culture. AB - Cerebellar granule neurons isolated from 7-day-old rats and cultured in normal medium undergo apoptosis, but remain healthy under depolarizing conditions with elevated K(+) (>==25 mM) or in the presence of brain-derived neurotrophic factor. Northern blot analysis showed that cyclin D1 mRNA was up-regulated in this apoptotic process. Both granule neurons and microglia were immunostained with anti-cyclin D1 antibodies, which is consistent with our previous finding that microglia become activated in response to neuronal cell death under these conditions. Only granule neurons, however, showed an enhanced expression of both mRNA and protein levels of cyclin D1 in the presence of aphidicolin that completely eliminated non-neuronal cells. The entire cell body of granule neurons became immunostained prior to cell shrinkage or nuclear condensation. Moreover, cell cycle blockers and an inhibitor of cyclin-dependent kinases suppressed both increased immunoreactivity and cell death, further substantiating the involvement of an abortive cell cycle in this process. In contrast, both levels of cyclin D1 remained unaltered in mature granule neurons undergoing apoptosis following combined serum withdrawal and low K(+) shift, suggesting developmental stage dependence of granule neuron apoptosis in vitro. This culture system is suitable for further analysis of the role of cyclin D1 in cell death. PMID- 10518115 TI - Protective effects of asiaticoside derivatives against beta-amyloid neurotoxicity. AB - Asiaticoside (AS) derivatives were tested for potential protective effects against Abeta-induced cell death. Of the 28 AS derivatives tested, asiatic acid (AA), asiaticoside 6 (AS6), and SM2 showed strong inhibition of Abeta-induced death of B103 cells at 1 microM. The three AS derivatives were further tested for their effects on free radical injury and apoptosis. All three AS derivatives reduced H(2)O(2)-induced cell death and lowered intracellular free radical concentration, but AA showed the strongest protection. In contrast, SM2 was the most effective blocker of staurosporine-induced apoptosis. These results suggest that the three AS derivatives block Abeta toxicity by acting through different cellular mechanisms. When applied to hippocampal slices, AA, SM2, and AS6 did not alter n-methyl-D-aspartic acid (NMDA) or non-NMDA receptor-mediated synaptic transmission, paired-pulse facilitation or induction of long-term potentiation in the field CA1. These results indicate that the three AS derivatives do not alter physiological properties of the hippocampus at the concentration that blocks Abeta-induced cell death. Therefore AS6, AA, and SM2 can be regarded as reasonable candidates for a therapeutic Alzheimer's disease drug that protects neurons from Abeta toxicity. PMID- 10518114 TI - GABA(A) receptor in growth cones: the outline of GABA(A) receptor-dependent signaling in growth cones is applicable to a variety of alpha-subunit species. AB - The growth cone is responsible for axonal elongation and pathfinding by responding to various modulators for neurite growth, including neurotransmitters. We demonstrated an outline of the gamma aminobutyric acid (GABA)(A)-dependent signaling in growth cones. Here, we examined the effects of the modulators of GABA(A) receptor on the signaling in growth cones. Phenobarbital or propofol, acting on beta-subunit, enhanced the [Cl(-)]infi change and [Ca(2+)](i) elevation by the GABA stimulation to isolated growth cones. Besides, propofol enhanced GABA dependent phosphorylation of growth-associated protein of 43 kDa (GAP-43) by protein kinase C. In contrast, an alpha-subunit acting agent diazepam did not modulate any of the above signals. Next, we examined the effect of the developmental change of alpha-subunit on the outline of the GABA(A)-dependent signaling in growth cones. We also found that the amounts of several different alpha-subunit isoforms developmentally increased or decreased in growth cone membrane (GCM), but that the affinity and density of the [(3)H]diazepam binding sites were similar to those in adult synaptic membrane. Taken together, our results strongly suggest that each step of GABA(A)-dependent signaling in GCM is not modified by diazepam, indicating that the signaling pathway mediated by GABA(A) receptor in growth cones is applicable to any compositional change of alpha-subunit isoforms. PMID- 10518116 TI - Differential effects of staurosporine and retinoic acid on the vulnerability of the SH-SY5Y neuroblastoma cells: involvement of bcl-2 and p53 proteins. AB - Human catecholaminergic neuroblastoma cells (SH-SY5Y) have been widely used in different neurochemical investigations. Quite often these cells are induced to differentiation by various agents, such as staurosporine and retinoic acid. Interestingly, even though both staurosporine and retinoic acid induce similar morphological differentiation in SH-SY5Y cells, we found that these two groups of differentiated cells exhibited opposite vulnerability to harmful chemicals and physical insults. In the present study, cisplatin, 5-fluorouracil (5-FU), N-(2 chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4), 6-hydroxydopamine (6-OHDA), and gamma-radiation were used to assess the tolerance of the differentiated cells. Cell viability was determined by 3-(4,5-dimethylthiazol-2yl)-2, 5 diphenyltetrazolium bromide (MTT) assay. Staurosporine-treated SH-SY5Y cells were more sensitive to these toxic insults than the untreated controls. In contrast, retinoic acid-treated cells became more resistant to the same treatments. The expression of the proteins of the protooncogene Bcl-2 and the tumor suppressor gene p53 following staurosporine or retinoic acid treatment was assessed by Western blot and immunocytochemistry. Retinoic acid increased Bcl-2 and decreased p53 levels, whereas staurosporine decreased Bcl-2 and increased p53 levels. The opposite alteration of Bcl-2 (anti-apoptotic) and p53 (apoptotic) contents in SH SY5Y cells with retinoic acid and staurosporine are attributed to the changes in cell vulnerability. These observations also indicate that caution should be taken when chemically induced differentiated neuroblastoma cells are to be used as an in vitro model for studying neuronal survival. PMID- 10518117 TI - Brain gamma-glutamyl cysteine synthetase (GCS) mRNA expression patterns correlate with regional-specific enzyme activities and glutathione levels. AB - The first and rate-limiting reaction in the formation of glutathione is catalyzed by gamma-glutamylcysteine synthetase (GCS), a dimer composed of a catalytic heavy and a regulatory light subunit. We previously found that heavy subunit GCS mRNA appears to be expressed at high levels in the hippocampus, cerebellum, and cortex of murine brain and at lower levels in the neostriatum (Kang et al. [1997] NeuroReport 8:2053). Here we report that variations in expression of light subunit GCS mRNA in murine brain resembles that of the heavy subunit mRNA with a few minor exceptions. Moreover, levels of GCS activity and glutathione levels in various brain regions appear to correspond to levels of expression of both GCS mRNA subunits. Based on these data, differences in the distribution of expression of the GCS subunits in the brain may therefore have major implications for the susceptibility of various brain regions to oxidative stress and/or mitochondrial damage. PMID- 10518118 TI - Mexiletine and magnesium independently, but not combined, protect against permanent focal cerebral ischemia in Wistar rats. AB - The neuroprotective effect of mexiletine (Mex), a potent Na(+) channel blocker which decreases neuronal energy demands and prevents energy depletion during ischemia, was evaluated in Wistar rats subjected to permanent middle cerebral artery (MCA) occlusion. Postmortem infarct volumes were determined by quantitative image analysis of triphenyltetrazolium (TTC)-stained brain sections. Pretreatment with Mex resulted in a significant infarct volume reduction when administered intraperitoneally, either at the dosage of 50 or 60 mg/kg, 1 hr before MCA occlusion (P < 0.05). Delayed treatment with Mex (50 mg/kg) also had neuroprotective effects when given at 0.5 hr (< 0.05), but not 2-4 hr, after MCA occlusion. Intraarterial administration of MgSO(4) (90 mg/kg), in combination with Mex at 60 mg/kg, showed no additive neuroprotective effect, although each agent independently reduced the MCA occlusion-induced infarction volume (P < 0.05). Our results indicate that a single, acute administration of Mex is neuroprotective against permanent focal cerebral ischemia, but perhaps chronic administration is needed to establish a more effective therapeutic window beyond 0.5 hr. Moreover, the present in vivo data do not favor a combined use of Mg(2+) with Mex for limiting ischemic injury in the brain, since these agents caused cardiopulmonary suppression, which may have led to the loss of the neuroprotective effect of each agent independently. PMID- 10518119 TI - Perinatal supplementation of low doses of ethanol enhances 5-HT restoration in the central nervous system. AB - It has been reported that long-term administration of ethanol has deleterious effects on the central nervous system; the alterations are particularly evident if the exposure occurs during development. Our study shows that rat perinatal administration of 3% and 6% ethanol does not alter development of serotonin (5 HT) pathways in the central nervous system, while their reactive changes triggered by neonatal lesioning are greatly altered. The administration of 5, 7 dihydroxytriptamine (5,7-DHT) within 6 hours from birth causes 5-HT fiber degeneration throughout the central nervous system. The loss of 5-HT is particularly relevant in lumbar spinal cord, occipital cortex, and hippocampus. This early decrease in 5-HT content is followed by a slow and partial recovery. If animals are exposed to 3% ethanol during the perinatal period, there is an enhancement of the 5,7-DHT-induced degeneration that is, however, followed by a faster and greater recovery throughout the central nervous system. Conversely, perinatal exposure to 6% ethanol and 5, 7-DHT administration lead to an irreversible 5-HT loss with no subsequent recovery. The deleterious effects of 6% ethanol are accompanied by a reduced expression of neurotrophin. Thus, our study suggests that chronic exposure to ethanol can influence central nervous system plasticity during development. Low doses may enhance neuronal plasticity and repair perhaps via an increased efficacy of neurotrophic factors, whereas higher doses may negatively affect neural development also by means of the impairment of the expression of neurotrophic factors. PMID- 10518120 TI - Rasagiline, a monoamine oxidase-B inhibitor, protects NGF-differentiated PC12 cells against oxygen-glucose deprivation. AB - In our in vitro model, rasagiline a selective irreversible monoamine oxidase-B (MAO-B) inhibitor, protected nerve growth factor (NGF)-differentiated PC12 cells from cell death under oxygen and glucose deprivation (OGD). The severity of the OGD insult, as expressed by cell death, was time-dependent. Exposure of the cells to OGD for 3 hr followed by 18 hr of reoxygenation caused about 30-40% cell death. Under these conditions, the neuroprotective effect of rasagiline was dose dependent: rasagiline reducing OGD-induced cell death by 68% and 80% at 100 nM and 1 microM, respectively. The neuroprotective effect of rasagiline was also observed when added after the OGD insult (55% reduction in cell death). Under rasagiline treatment, there was a lesser decrease in ATP content in cultures exposed to OGD compared with that in untreated cultures. OGD followed by reoxygenation resulted in a several fold increase in PGE(2) release into the extracellular medium. Rasagiline (100 nM-1 microM) markedly inhibited OGD-induced PGE(2) release. Clorgyline, a monoamine oxidase-A (MAO-A) inhibitor, did not protect NGF-differentiated PC12 cells against OGD-induced cell death. As NGF differentiated PC12 cells contain exclusively MAO type A, these data suggest that the neuroprotective effect of rasagiline under OGD conditions is independent of MAO inhibition. PMID- 10518121 TI - Triggering of soman-induced seizures in rats: multiparametric analysis with special correlation between enzymatic, neurochemical and electrophysiological data. AB - Soman, an anticholinesterasic neurotoxic drug, induces epileptic seizures during severe intoxication. The neuropathological lesions then observed are linked to the appearance of these seizures, but their trigger conditions still remain unknown and a great variability between animals is observed. We have developed a technique allowing, in freely moving rats, the in vivo determination of three sets of neurophysiological data, before and during a soman intoxication. For the same rat, we associated cortical acetylcholinesterase (AChE) activity by microdialysis with both the assay of extracellular acetylcholine (ACh) concentrations and electroencephalographic (EEG) recording and power spectrum analysis (gamma band). Data have been analyzed to define the critical parameters which lead to the epileptic fit. Although we found thresholds for seizure occurrence, AChE inhibition having to be over 65% and ACh over 200-fold the baseline, these two criteria are not sufficient to predict the appearance of seizures. Only animals with no increase of energy in the gamma band early after soman poisoning will then exhibit an epileptic fit. Gamma band energy is modulated by noradrenergic activity and might be related to the sympathetic response to stress. So we can hypothesize, that the variation of energy in gamma band after intoxication, which might be related to stress adaptation strategy, may determine whether or not the animal will exhibit an epileptic fit. PMID- 10518122 TI - Reply PMID- 10518123 TI - Bilirubin rebound. PMID- 10518124 TI - Cytology beyond description and differentials. PMID- 10518126 TI - Percutaneous core needle biopsy vs. fine needle aspiration in diagnosing benign lung lesions. AB - OBJECTIVE: To determine the diagnostic value of percutaneous core needle biopsy (PCNB) in comparison with fine needle aspiration (FNA) in patients with benign pulmonary lesions. STUDY DESIGN: A retrospective review was undertaken of computed tomography-guided PCNBs and FNAs performed between 1988 and 1997. Both FNA and PCNB biopsies were carried out sequentially at the same visit in every patient. RESULTS: A specific benign diagnosis was made in 10/60 cases (16.7%) by FNA and in 49/60 (81.7%) by PCNB. PCNB findings resulted in significant modification of the diagnosis established by FNA. The only significant complication encountered was pneumothorax, at a rate of 11.7%, which is compatible with that reported in the literature for complications induced by FNA alone. CONCLUSION: Radiologically guided PCNB is a safe procedure, can provide sufficient histologic material for a specific diagnosis of peripheral lung disease and can avoid more-invasive surgical procedures in many cases. Our experience demonstrated that the histologic analysis provided by PCNB can greatly increase the diagnostic accuracy in benign pulmonary diseases as compared with the yield of FNA. PMID- 10518125 TI - Amyloid in cytologic specimens. Differential diagnosis and diagnostic pitfalls. AB - OBJECTIVE: To describe and illustrate the characteristic features of amyloid in cytologic preparations and point out its diagnostic pitfalls. STUDY DESIGN: Five fine needle aspirates and one bronchial washing that contained amyloid were retrospectively reviewed. The aspirates were obtained from each of the five following sites: lung, occipital lymph node, thyroid gland, proximal humerus and subcutaneous soft tissue. Smears of all of the aspirates were stained with Papanicolaou stain, and in two cases they were also stained with Diff-Quik. Cell block sections were stained with hematoxylin and eosin. Congo red, CD45 and CD20 were used on selected cases. RESULTS: Amyloid appears as either flocculent material or irregularly shaped fragments with scalloped and pointed edges. The amorphous fragments are acellular and frequently associated with connective tissue cells. They stain eosinophilic to cyanophilic with Papanicolaou stain and deep blue with Diff-Quik. In two cases an exuberant giant cell reaction almost obscured the amyloid. In the thyroid aspirate, the amyloid was misinterpreted as colloid. In bronchial washings and lung aspirates, amyloid has to be distinguished from mucus, alveolar proteinosis, chondroid material and corpora amylacea. When circumferentially surrounded by lymphocytes or plasma cells, flocculent amyloid deposits may simulate adenoid cystic carcinoma. CONCLUSION: Amyloid can be easily overlooked or mistaken for other entities with similar staining qualities. Congo red staining can help to confirm the diagnosis. PMID- 10518127 TI - Diagnosing granulomatous inflammation of the skin. A cytomorphologic approach based on evaluation of cellular reaction patterns. AB - OBJECTIVE: To determine the applicability of recognizing cellular reaction patterns in the cytologic diagnosis of granulomatous inflammation of the skin. STUDY DESIGN: Prospective. Two hundred seventeen cases of clinically suspected granulomatous dermatitis formed the data for this study. May-Grunwald-Giemsa, Ziehl-Neelsen, periodic acid-Schiff, silver impregnation and Gram staining methods were employed. RESULTS: Five cytomorphologic patterns of granulomatous inflammation were identified: epithelioid cell granulomas (77), histiocytic granulomas (20), epithelioid cell granulomas with polymorphous exudate (77), foreign body granulomas (3) and fat necrosis with granulomas (9). CONCLUSION: Correlating clinical presentation with cytomorphologic patterns often yields diagnostic information in the workup of granulomatous inflammation of the skin and frequently obviates the need for biopsy. PMID- 10518129 TI - Fine needle aspiration cytology of reactions in leprosy. AB - OBJECTIVE: To define diagnostic cytomorphologic features of reactions in leprosy. STUDY DESIGN: Part-retrospective, part-prospective, single-blind, controlled study of the applicability of fine needle aspiration cytology in the diagnosis of reactions in leprosy. Cytomorphologic features were compared in 42 clinically diagnosed patients with reactions in leprosy with those in a control group of patients with nonreactional leprosy. The study groups included type 1 and type 2 reactions in 35 and 9 patients, respectively. May-Grunwald-Giemsa and Ziehl Neelsen staining methods were employed. RESULTS: Statistically significant (P < .01) cytomorphologic features of type 1 reaction were the presence of fragments of collagen and elastin; giant cells; giant cells exhibiting elastin phagocytosis; loose, epithelioid cell granulomas; and fibroblasts. Type 2 reaction was characterized in aspirates by the presence of an abundance of neutrophils in a background of lepromatous leprosy (P < .01). CONCLUSION: Criteria that are used in histopathology for the diagnosis of leprosy reactions can be applied satisfactorily to cytologic smears. A good correlation between clinical diagnosis and cytomorphology can be achieved. Multiple-site aspirates from the skin, nerve and lymph nodes are helpful in substantiating the diagnosis. PMID- 10518128 TI - Papillary carcinoma of the breast. Cytologic study of nine cases. AB - OBJECTIVE: To study the cytologic findings of papillary breast carcinoma by fine needle aspiration. STUDY DESIGN: The study group consisted of fine needle aspiration (FNA) specimens of breast tumors from nine patients performed during the period 1988-1997. Eight were female, and one was male. The FNA results were compared with the final histologic diagnosis. RESULTS: The tumor sizes were 4-6.5 cm. The aspirations yielded a good amount of bloody material. The smears revealed high cellularity, papillary clusters, isolated low-to-tall columnar cells, mild to moderate atypia, hemorrhagic background, foam and hemosiderin-laden macrophages, calcification, rare mitoses, palisading row of cells and bipolar cytoplasmic eosinophilic granules. The smears were diagnosed as either suspicious or suggestive of papillary carcinoma. The histologic examination revealed invasive papillary carcinoma. CONCLUSION: Papillary carcinoma of the breast can be diagnosed by using a panel of cytologic findings that includes hypercellularity, papillary clusters, hemorrhagic background, palisading rows of tall columnar cells, cellular atypia and calcification. The interesting finding in this study was the presence of eosinophilic bipolar cytoplasmic granules, which has not been reported before. PMID- 10518130 TI - Ultrastructure of pleural mesothelioma and pulmonary adenocarcinoma in malignant effusions as compared with reactive mesothelial cells. AB - OBJECTIVE: To determine the ultrastructural features of diffuse malignant pleural mesothelioma cells in cytologic specimens from pleural effusions. STUDY DESIGN: We retrospectively studied 35 pleural effusions: 12 diffuse malignant pleural mesotheliomas (8 epithelial type, 4 biphasic type), 12 pulmonary adenocarcinomas and 11 cases of reactive mesothelial cells. RESULTS: In the cytoplasm, reactive and malignant mesothelial cells had more-abundant intermediate filaments (P < .05, P < .01) and fewer free ribosomes (P < .001, P < .001) than adenocarcinoma cells. Reactive mesothelial cells had fewer mitochondria than mesothelioma cells (P < .05). Mesothelioma cells had longer, thinner microvilli on the cell surfaces (P < .001); length/diameter ratios of microvilli were 19.1 +/- 7.0 (mesothelioma) vs. 9.1 +/- 2.2 (adenocarcinoma) and 9.2 +/- 2.4 (mesothelial cells). Giant intercellular junctions (desmosomes or desmosomelike structures > 1 micron in length) were found in eight cases of mesothelioma. Core filaments or rootlets in microvilli were present in two cases of adenocarcinoma. CONCLUSION: Because cytologic specimens from pleural effusions were easy to obtain, we think ultrastructural cytology is useful in distinguishing mesothelioma from adenocarcinoma and benign effusions. PMID- 10518131 TI - Analysis of lifestyle data and cytologic findings in a pilot cervical screening project in rural Vietnam. AB - OBJECTIVE: To evaluate the possibilities of a cervical cytology screening program and the importance of lifestyle parameters in women living in rural Vietnam. STUDY DESIGN: Screening took place in Quangninh province in the north of Vietnam. From 700 women screened for the first time, smears were prepared and read by a cytotechnologist. The number of children and abortions (some women having had 10 abortions) allowed us to study the relationship of these life-style parameters with disturbed vaginal ecology (defined as the presence of Candida, trichomonads or cocci in the smear). RESULTS: The smears from farmers showed the greatest ecologic disturbances in vaginal flora. Water supply turned out to be important for coccoid overgrowth. Scores for both fungal infection and coccoid overgrowth were related to profession. Candidosis proved to be very prominent in cytologically negative smears: the organisms were found (often in abundance) in 95 of 661 negative smears but were absent from cytologically positive smears. CONCLUSION: Since only two cases of CIN 3 were found and no cases of invasive carcinoma, cervical screening for cancer ought not to be the first priority for rural Vietnam. PMID- 10518132 TI - Calculus artifact. A challenge in urinary cytology. AB - OBJECTIVE: To retrospectively review calculus artifact and compare it with instrument artifact and papillary transitional cell carcinoma (TCC). STUDY DESIGN: Voided urine specimens from patients with calculi (65), TCC (low grade, 10, high grade, 34) and history of prior instrumentation (12) were studied. RESULTS: Nineteen specimens of calculus artifact had unremarkable cytology. Forty six specimens had abnormal single cells or papillary clusters and cell balls or a mixture of both. The papillary groups had smooth as well as irregular borders, a cytoplasmic collar and cells with occasional cytoplasmic vacuoles, slightly increased nuclear/cytoplasmic (N/C) ratio and inconspicuous nucleoli. Squamous preponderance and birefringent crystals were seen. In instrumentation artifact, papillary clusters or three-dimensional cell balls had smooth borders, cytoplasmic collars, an occasional cytoplasmic vacuole, normal N/C ratio, regular nuclear membrane and finely granular nuclear chromatin. In TCC, papillary clusters with loss of polarity and irregular borders were present in both grades but were predominant in low grade TCC. No cytoplasmic collar was noted. In high grade TCC, single cells and nuclear alterations were more pronounced, with increased N/C ratio, hyperchromasia, coarse chromatin, irregular nuclear envelopes, prominent nucleoli and rare mitosis. CONCLUSION: Calculus artifact can produce papillary clusters masquerading as papillary TCC. Unlike instrument artifact, there may be significant nuclear atypia, which could be reversible. To avoid diagnostic pitfalls, further investigation is suggested after removal of calculus. PMID- 10518133 TI - False negative cytologic diagnosis of breast carcinoma. AB - OBJECTIVE: To study the reasons for interpretive errors in false negative diagnosis of breast carcinoma on fine needle aspiration cytology material. STUDY DESIGN: We reviewed only those histologically proved malignant cases where the cytologic material was abnormal and to some extent misinterpreted. RESULTS: There were four lobular carcinomas and one each case of in situ, infiltrating duct, medullary and tubular carcinoma. Smears of lobular carcinomas were hypocellular overall, and the cells showed minimal nuclear pleomorphism. In situ, medullary and tubular carcinoma were associated with fibrocystic changes. The presence of bipolar cells and stromal fragments was misleading in cases of infiltrating duct carcinoma. CONCLUSION: The presence of associated fibrocystic disease may be a misleading factor since it may mask a malignancy. Hypocellularity and relatively nuclear monomorphism were the most common reasons for failure to diagnose malignant breast lesions. Careful attention should be paid to extreme nuclear monomorphism and absence of naked bipolar cells. A cytologically atypical or suspicious diagnosis together with radiologic suspicion should suggest a diagnosis of malignancy. PMID- 10518134 TI - Extrauterine malignancies. Role of Pap smears in diagnosis and management. AB - OBJECTIVE: To further elucidate the cytologic manifestations of extrauterine malignancies, to evaluate their possible distinction from primary cervical malignancies and to analyze their clinical significance and role in staging. STUDY DESIGN: Papanicolaou (Pap) smears in 33 cases with abnormal cells originating in histologically proven extrauterine carcinomas were evaluated. These cases came from the files of the Medical College of Pennsylvania and Lankenau Hospitals. RESULTS: Ovary, gastrointestinal tract and breast were the three most frequent primary sites, accounting for 28 of the 33 cases (85%). The histologic types encountered were adenocarcinoma, 29 cases (88%); mucoepidermoid carcinoma, 1 (3%); small cell carcinoma, 1 (3%); cloacogenic carcinoma and large cell lymphoma, 1 (3%). The following diagnoses were rendered at the time of initial evaluation: adenocarcinoma consistent with metastasis, 21 cases; carcinoma, primary versus metastatic, 2; adenocarcinoma, suspicious for endometrial primary, 2; suspicious for carcinoma, 1; and atypical glandular cells, 7. CONCLUSION: The yield for positive Pap smear diagnoses in extrauterine malignancies is best in patients with an established diagnosis of a primary neoplasm. The degree of tumor differentiation and extent of tumor involvement did not appear to correlate with diagnostic yield. There appeared to be no statistically significant association of tumor diathesis with primary versus metastatic carcinoma and presence or absence of documented local involvement of the endometrium, cervix or vagina. Therefore, while Pap smears can serve as a diagnostic tool in the evaluation of extrauterine malignancies, they are best utilized as an adjunct to tumor staging and patient management. PMID- 10518136 TI - Filtration and agar embedding applied to fine needle aspirates for tumor diagnosis by electron microscopy. A combined technique. AB - OBJECTIVE: To develop a novel method for processing of fine needle aspirates subjected to electron microscopic (EM) study. STUDY DESIGN: Included 70 cases of poorly differentiated malignant tumors in which a definitive diagnosis was not possible on light microscopic (LM) examination and that thus required application of an ancillary technique such as FNA/EM, for diagnosis. We have established a novel method of processing, a technique of filtration through nylon mesh filters to eliminate red blood cells (RBCs) and necrotic debris, followed by agar well embedding to avoid loss of diagnostic material during processing without centrifugation at later steps after agar embedding, thus minimizing the time required for processing. It was successfully carried out in 70 cases. RESULTS: The combined technique was extremely effective in eliminating RBCs and necrotic debris. It also avoided further loss of valuable diagnostic material. An accurate diagnosis was rendered in 70 cases; that was not possible by LM alone. The whole procedure saves two to three hours of processing as centrifugation is not required after the agar embedding step. CONCLUSION: This technique was found to be cost- and time-effective, particularly suitable for developing countries, where financial resources are limited. PMID- 10518135 TI - Morphologic analyses of positive peritoneal cytology in endometrial carcinoma. AB - OBJECTIVE: To investigate the relationship between the morphologic features of endometrial adenocarcinoma cells in peritoneal fluids (effusions and washings) and macroscopic intraabdominal adenocarcinoma at laparotomy as well as prognosis. STUDY DESIGN: Seventy-one patients with endometrial adenocarcinoma who showed positive peritoneal cytology at laparotomy were clinically divided into three groups: 25 patients with macroscopic neoplastic seeding in the peritoneal cavity (type 1), 38 patients without macroscopic peritoneal metastasis who survived with no evidence of disease (type 2) and 8 patients without macroscopic peritoneal metastasis who later developed recurrence of adenocarcinoma (type 3). Morphologic features of the adenocarcinoma cells in smears of peritoneal fluids were examined. RESULTS: Most of the smears from type 1 patients showed moderate to high cellularity, scalloped edges of cell clusters and isolated adenocarcinoma cells, whereas these features were seldom observed in type 2 patients. Although not all type 3 patients demonstrated these three features, patients in the series whose specimens exhibited none of the three features did not show any peritoneal lesions or have a recurrence of their disease. CONCLUSION: The finding of endometrial adenocarcinoma cells exhibiting high cellularity, scalloped edge of cell clusters and isolated cells in smears of peritoneal fluid is associated with the presence of intraabdominal macroscopic metastatic lesions and could be regarded as a risk factor for intraabdominal recurrence of carcinoma. PMID- 10518137 TI - Usefulness of estrogen receptor detection using archival Papanicolaou-stained smears. AB - OBJECTIVE: To examine estrogen receptor (ER) detection using cytologic specimens and to compare the results with those obtained by the dextran-coated charcoal (DCC) method and enzyme immunoassay (EIA). STUDY DESIGN: Immunocytochemical staining was conducted on 60 cases of breast cancer resected at our hospital between April 1993 and November 1997 in which ER had been measured by DCC or EIA. Specimens for immunocytochemical staining were prepared by a cell transfer method using archival Papanicolaou-stained imprint smears, and ER staining was performed by the labeled streptavidin method using an anti-ER monoclonal antibody. These results were compared with those obtained by DCC or EIA. RESULTS: In immunocytochemical staining for ER, positive staining was observed in the nuclei of tumor cells. A good correlation was obtained between the immunocytochemical staining results and biochemical results. Five cases were positive in anti-ER staining but negative in biochemical tests, and two cases were negative in anti ER staining and positive in biochemical tests. CONCLUSION: Unlike biochemical assays, the immunocytochemical method does not necessitate use of fresh frozen materials and can be performed even using archival Papanicolaou-stained smears. Immunocytochemical study is a highly useful method for routine ER determination. PMID- 10518138 TI - Analysis of mRNA quality in freshly prepared and archival Papanicolaou samples. AB - OBJECTIVE: To study the feasibility of utilizing mRNA recovered from cytologic Papanicolaou (Pap) specimens as a resource for gene expression studies of normal and diseased cells. STUDY DESIGN: To assess the effects of fixation on mRNA recovery and analysis, fresh Pap samples were processed by three separate methods: (1) routine cytologic fixation (2) 70% ethanol fixation, and (3) air drying without fixation. One-week-old, 1-month-old, 1-year-old and 10-year-old samples were studied to determine the quality of mRNA in archival samples. mRNA quality was analyzed by RT-PCR for the HPRT gene, and by complete transcript amplification. Both heterogeneous (whole slide scrapes) and microdissected cell populations were studied. RESULTS: Reverse transcriptase-polymerase chain reaction (RT-PCR) for the hypoxanthine guanine phosphoribosil transferase gene was positive in all fresh and archival samples and was not affected by fixative, processing methodology or microdissection. Complete transcript amplification followed by gel electrophoresis showed cDNA smears in all fresh samples with a maximum intensity between 1 and 2 kilobases (kb). Amplification of mRNA was not affected by fixation. Smaller cDNA smears were seen in archival specimens with a maximum intensity between 0.5 and 1.5 kb in both one-week-old and one-month-old samples. Smears of approximately 500 base pairs were observed in the 1-year-old and 10-year-old samples. Successful mRNA amplification was possible from microdissected cell populations. CONCLUSION: Messenger RNA recovery and analysis is possible from archival cytologic specimens, suggesting that they can serve as a useful template for RT-PCR analysis of individual genes as well as newly developing high-throughput gene expression methodologies, such as microarrays. Cytologic samples may be particularly useful for study of archival samples as well as diseases from which tissue samples amenable to mRNA-based studies are not available. PMID- 10518139 TI - DNA polymerase chain reaction using fine needle aspiration biopsy smears to evaluate non-Hodgkin's lymphoma. AB - OBJECTIVE: To apply polymerase chain reaction (PCR) analysis to the fine needle aspiration biopsy (FNAB) evaluation of lymphoid proliferations. STUDY DESIGN: We analyzed 37 consecutive archived FNAB malignant lymphoma specimens. Immunophenotypic data from the fine needle aspiration biopsy and excisional biopsy material was available for all specimens. PCR to identify monoclonal rearrangements of the immunoglobulin heavy chain gene, T-cell receptor and translocations involving the bcl-1 and bcl-2 genes was performed. RESULTS: Seventy-eight percent of cases were detected by at least one of these assays. Where DNA analysis was performed on excisional biopsy material, 70% of the cases had identical results; no discordant results for the immunoglobulin heavy chain gene or T-cell receptor were found. In 23% of cases, after review of all available data, a discordant result was thought to be a consequence of a false negative result in DNA analysis of excisional biopsy material. CONCLUSION: These findings indicate that PCR analysis of archived FNAB material, when necessary, provides useful information for diagnosis and staging of malignant non-Hodgkin's lymphomas. PMID- 10518141 TI - Multilobated large B-cell lymphoma diagnosed cytologically. A case report. AB - BACKGROUND: Fine needle aspiration (FNA) biopsy can be used to reliably classify most conditions involving lymph nodes or, at least, significantly reduce the differential diagnosis. CASE: A 70-year-old male presented with an ulcerated mass arising from the left tonsillar fossa and involving the anterior and posterior pillars. A biopsy of the tonsillar mass performed at an outside hospital was interpreted as a large cell undifferentiated carcinoma. Subsequently the patient developed systemic lymphadenopathy. A bone scan showed intense uptake within the medial tibial plateau of the left knee. FNA biopsy of the right axillary mass was interpreted at University of Cincinnati Medical College as a large cell lymphoma, multilobated type. Histologic and immunohistochemical studies of the lymph node confirmed the presence of multilobated B-cell lymphoma. Lymphoma chemotherapy was initially successful but was discontinued due to toxicity. The patient died two months after the initial cytologic diagnosis of lymphoma. CONCLUSION: Multilobated lymphomas are an unusual variant of non-Hodgkin's lymphomas (mostly B-cell type). Cytology and immunocytochemistry are useful diagnostic procedures that can help to diagnose this relatively uncommon type of lymphoma and significantly reduce the possibility of misdiagnosis. PMID- 10518140 TI - Mucinous adenocarcinoma of the parotid gland. Report of a case with fine needle aspiration findings and histologic correlation. AB - BACKGROUND: Mucinous adenocarcinoma rarely arises as a primary tumor within the parotid gland, and only the histologic features of this tumor have been described. CASE: A 4-cm, firm mass arose in the right parotid gland of a 72-year old male over a six-week period. Cystic on computed tomography, the mass, on fine needle aspiration biopsy, yielded monomorphic, moderately atypical cells, both single and clustered, associated with abundant mucoid material and focal necrosis. Tumor cells had eccentric nuclei, prominent nucleoli and occasional cytoplasmic vacuolization. A few binucleated and multinucleated tumor cells were present. Histologic sections of the resected gland showed mucinous adenocarcinoma. A metastatic workup was negative. The differential diagnoses on cytology included other primary tumors of the parotid gland producing mucin or a mucoid matrix and metastatic mucinous adenocarcinomas. CONCLUSION: To our knowledge, this is the first cytologic description of mucinous adenocarcinoma, primary in the parotid gland. PMID- 10518143 TI - Detection of Schistosoma mansoni in bronchoalveolar lavage fluid. A case report. AB - BACKGROUND: Bronchoalveolar lavage (BAL) is a useful tool in the diagnosis of bacterial, viral, fungal and parasitic pulmonary infections. There have been rare reports of parasitic infestations in bronchoalveolar lavage fluid. This is the first case report on detecting a Schistosoma ova in BAL fluid. CASE: A 40-year old, Egyptian male presented with a fever and productive cough. He had a right pleural effusion and segmental collapse of the right lower lobe. BAL fluid showed several ova of Schistosoma mansoni and established the diagnosis of schistosomiasis. Abdominal ultrasound revealed mild hepatic cirrhosis. CONCLUSION: Schistosomiasis should be considered in the differential diagnosis of pulmonary problems in patients with disseminated disease in endemic areas. PMID- 10518142 TI - Diagnosis of microfilaria in gastric brush cytology. A case report. AB - BACKGROUND: Filariasis due to Wuchereria bancrofti is endemic to southern Asia. While the laboratory diagnosis has been conventionally made by demonstrating microfilariae in peripheral blood smears, these have also been occasionally diagnosed on aspiration cytology of various organs. CASE: A 54-year-old male presented with a burning sensation in the epigastrium of five months' duration. Endoscopic brush biopsy revealed numerous sheathed microfilariae of W bancrofti. The patient had had no symptoms suggestive of filarial infection in the past. Cytology revealed numerous microfilariae among lymphocytes and neutrophils. CONCLUSION: This case illustrates that a thorough examination of gastric brushings can at times reveal unexpected findings and may prove to be a useful supplement to endoscopic biopsy. PMID- 10518144 TI - Fine needle aspiration of angiomatoid malignant fibrous histiocytoma. A case report. AB - BACKGROUND: During recent years, reliable cytodiagnostic criteria have been proposed for a number of soft tissue lesions, both benign and malignant. However, cytomorphologic descriptions of angiomatoid malignant fibrous histiocytoma (AMFH) are lacking. We report the cytomorphologic features of this uncommon lesion. CASE: A 10-year-old female presented with a swelling in the thigh that, on fine needle aspiration (FNA), was reported as a soft tissue tumor (vasoformative) of intermediate malignancy. Subsequent excision with histopathologic examination confirmed the diagnosis. CONCLUSION: The differential diagnosis of AMFH based on FNA findings can be difficult and should be made in conjunction with clinical findings. PMID- 10518145 TI - Multicystic autoimmune thyroiditis-like disease associated with HIV infection. A case report. AB - BACKGROUND: Human immunodeficiency virus (HIV) infection and resulting acquired immunodeficiency syndrome (AIDS) may involve virtually every organ system, including the endocrine glands. Thyroid dysfunction most commonly reflects advanced disease and generally resembles euthyroid sick syndrome. Rarely do opportunistic infections, hemorrhage, neoplasms and drugs account for alterations in thyroid tissue. Multiple lymphoepithelial cysts of parotid gland and thymus have been identified, but similar findings in thyroid gland have not been reported. CASE: A 41-year-old, HIV-seropositive woman, asymptomatic for seven years, developed a squamous cell carcinoma of the cervix with local-regional extension. At the same time, bilateral complex thyroid cysts and high titers of antimicrosomal antibodies (1/6,400) were detected. Ultrasound-guided fine needle aspiration biopsy of the thyroid showed a heterogeneous lymphocytic population with a reactive appearance and occasional groups of epithelial cells with an immature squamous pattern, along with cytologic features of autoimmune thyroiditis. Immunocytochemistry was positive for CD20, CD3 and CD5. Immunoglobulin heavy chain gene rearrangement by polymerase chain reaction from cytologic material showed a polyclonal lymphoid population. External radiotherapy resulted in a significant reduction in the pelvic lesion. Four months after diagnosis, abdominal ultrasound displayed multiple hepatic metastasis, the patient's condition rapidly deteriorated, and she died about a month later. CONCLUSION: This case had unique features and probably represented an AIDS related lesion and distinct entity. PMID- 10518146 TI - Fine needle aspiration cytology of plexiform fibrohistiocytic tumor. A case report. AB - BACKGROUND: Plexiform fibrohistiocytic tumors are rare lesions of proposed myofibroblastic origin occurring primarily in infants and children. While the histologic, immunohistochemical and ultrastructural findings have been well described, cytologic description has been limited. CASE: An 8-month-old, male infant presented with a posterior chest wall mass and decreased use of his left arm. Fine needle aspiration biopsy showed a spectrum of plump fibroblastic spindle cells and histiocytelike cells within a finely granular myxoid background. Osteoclastlike giant cells were also noted. CONCLUSION: We report here the cytologic findings of a plexiform fibrohistiocytic tumor from fine needle aspiration biopsy studied using Papanicolaou, Ultrafast Papanicolaou and Diff-Quik stain, with the cytologic differential diagnosis of other spindled and histiocytelike tumors. PMID- 10518147 TI - Fine needle aspiration biopsy of mastitis secondary to empyema necessitatis. A report of two cases. AB - BACKGROUND: Empyema necessitatis is a relatively rare entity. Two instances of mastitis secondary to empyema necessitatis, diagnosed by fine needle aspiration biopsy are reported. CASES: One case was tuberculous in etiology and was initially recognized by cytologic findings of epithelioid and granulomatous cellular reactions and the presence of acid-fast bacilli, which were subsequently cultured and speciated as Mycobacterium tuberculosis. The other case was due to coexisting Actinomyces and Actinobacillus. These organisms were cytologically suggested by "sulfur" granules of filamentous, gram-positive bacilli, admixed gram-negative coccobacilli and Splendore-Hoeppli phenomenon in an exudative cell background and were confirmed by microbiologic culture as Actinomyces israelii and Astinomyces actinomycetemcomitans, respectively. CONCLUSION: The usefulness of fine needle aspiration cytology in the diagnosis of empyema necessitatis, supported by ancillary microbial culture, histochemistry, and radiographic imaging, is well illustrated by these two cases. PMID- 10518148 TI - Horseshoe kidney presenting as a retroperitoneal mass. Report of a case diagnosed by fine needle aspiration cytology. AB - BACKGROUND: Horseshoe kidney is a renal congenital anomaly. It is the result of the fusion of either upper or lower poles of both kidneys, appearing as a horseshoe-shaped structure. This anomaly is very frequent: it can be found in about 1 of every 50-1,000 autopsies). CASE: Computed tomography performed routinely after pancreatitis in a 37-year-old female showed a retroperitoneal mass of uncertain origin. Fine needle aspiration cytology (FNAC) smears evidenced normal renal tissue. Urography confirmed the diagnosis of horseshoe kidney. CONCLUSION: This is the first reported case of horseshoe kidney diagnosed by FNAC. It demonstrates the utility of FNAC for diagnosis of retroperitoneal masses, especially if they are asymptomatic. PMID- 10518149 TI - Cytologic changes mimicking papillary carcinoma of the thyroid after 131I treatment. PMID- 10518150 TI - Fine needle aspiration diagnosis of lymphoepithelial cyst of the parotid gland. PMID- 10518151 TI - Intravascular lymphomatosis presenting as bilateral adrenal enlargement and insufficiency. PMID- 10518152 TI - Fine needle aspiration cytology of a papillary oncocytic neoplasm of the thyroid gland. PMID- 10518153 TI - Fine needle aspiration cytology of hyalinizing trabecular adenoma of the thyroid. PMID- 10518154 TI - A new APA for a new century. PMID- 10518156 TI - Incremental progress in developmental psychopathology: simply complex. PMID- 10518155 TI - Opportunities and challenges for psychiatry. PMID- 10518158 TI - Domestic terrorism with chemical or biological agents: psychiatric aspects. AB - OBJECTIVE: This article highlights the mental health consequences of a domestic terrorist incident involving chemical or biological weapons. METHOD: The author reviews the literature on the neuropsychiatric effects of selected chemical and biological weapon agents, on the psychological sequelae of mass disasters, and on approaches to crisis intervention. RESULTS: Disturbances of behavior, affect, and cognition can result directly from the pharmacological actions of some chemical and biological weapon agents. In addition, an incident involving these agents can have considerable psychological effects on individuals and the community. In either case, some disorders are acute and others are prolonged or delayed in onset. Effective therapeutic intervention involves a broad range of clinical, social, and administrative actions. CONCLUSIONS: Psychiatrists have an important role in the management of a chemical or biological terrorist incident and, along with their other medical colleagues, should train and prepare for it. PMID- 10518159 TI - Molecular mechanisms underlying mood stabilization in manic-depressive illness: the phenotype challenge. AB - OBJECTIVE: The authors critically examine the evidence supporting the hypothesis that lithium's therapeutic effects in bipolar affective disorder are mediated by alterations in the expression of specific genes in critical neuronal circuits. METHOD: Using the heuristic "initiation and adaptation paradigm," the authors appraise the biological effects and underlying molecular and cellular mechanisms of lithium's action. The evidence is critically reviewed, with special attention to the reductive and integrative approaches necessary for identifying lithium's clinically relevant cellular and molecular targets. RESULTS: Lithium's acute effects are mediated through inhibition of specific enzymes involved in two distinct but interacting signaling pathways--the protein kinase C and glycogen synthase kinase 3 beta signaling cascades--that converge at the level of gene transcriptional regulation. The expression of different genes, including transcription factors, is markedly altered by chronic lithium administration. Chronic lithium treatment also robustly increases the expression of the neuroprotective protein Bcl-2, raising the intriguing possibility that some of lithium's effects are mediated through underappreciated neurotrophic/neuroprotective effects. The importance of lithium's effect on circadian rhythms and the related methodological problems in validating the role of specific genes in lithium's therapeutic effects are discussed. CONCLUSIONS: Despite the plethora of lithium effects at the genomic level, direct evidence that the genes identified thus far are responsible for phenotypic changes associated with chronic lithium treatment is still lacking. The combination of sensitive molecular technologies, appropriately designed paradigms, better behavioral analysis, and a chronobiologic approach seems necessary for the future identification of one or more clinically relevant lithium-target genes. PMID- 10518160 TI - Effects of ADHD, conduct disorder, and gender on substance use and abuse in adolescence. AB - OBJECTIVE: The relationships of attention deficit hyperactivity disorder (ADHD), conduct disorder, and gender to substance abuse were studied in a large population-based sample of adolescent twins. METHOD: Structured interviews were administered to 626 pairs of 17-year-old twins (674 girls and 578 boys) and their mothers to generate lifetime psychiatric diagnoses, and computerized measures of current substance use were obtained. Hierarchical logit analyses were performed to assess the independent effects of ADHD, conduct disorder, and gender on current substance use, frequency of substance use, and DSM-III-R diagnoses of substance use disorders. RESULTS: Conduct disorder was found to increase the risk of substance use and abuse in adolescents regardless of gender. In contrast, independent of its association with conduct disorder, an ADHD diagnosis did not significantly increase the risk of substance use problems. CONCLUSIONS: This study found no significant gender differences in the effects of ADHD and conduct disorder on substance use and abuse, although there was some suggestion that girls with ADHD might be at slightly higher risk than boys for substance abuse. In addition, increased risk of substance abuse among adolescents with conduct disorder may be primarily confined to those with persistent conduct disorder. PMID- 10518161 TI - Concurrent and predictive validity of the personality disorder diagnosis in adolescent inpatients. AB - OBJECTIVE: The authors investigated the concurrent and predictive validity of the DSM-III-R diagnosis of personality disorder in adolescents by means of baseline and follow-up assessments of inpatients treated at the Yale Psychiatric Institute. METHOD: One hundred sixty-five hospitalized adolescents were reliably assessed by using a structured interview for personality disorder diagnoses as well as two measures of impairment and distress--the Global Assessment of Functioning Scale and the SCL-90-R. Two years after initial assessment, 101 subjects were independently reassessed with the same measures; their functioning was also assessed at this time. RESULTS: At baseline, adolescents with personality disorders were significantly more impaired than those without personality disorders. At follow-up, adolescents with a personality disorder diagnosis at baseline had used significantly more drugs and had required more inpatient treatment during the follow-up interval. Over time, the scores on the Global Assessment of Functioning Scale and SCL-90-R of adolescents diagnosed with a personality disorder at baseline became more similar to the scores of adolescents without a personality disorder. CONCLUSIONS: The diagnosis of personality disorder in adolescent inpatients has good concurrent validity; however, the predictive validity of the diagnosis is mixed. PMID- 10518163 TI - Children's symptoms in the wake of Challenger: a field study of distant-traumatic effects and an outline of related conditions. AB - OBJECTIVE: The Challenger space shuttle explosion in January 1986 offered an opportunity to determine what, if any, symptoms of posttraumatic stress disorder (PTSD) and bereavement normal latency-age children and adolescents would develop after a distant, horrifying event. METHOD: With a structured interview, the authors assessed the symptoms of 153 randomly selected children from Concord, N.H., and Porterville, Calif. Responses were statistically compared between East Coast children, who saw the event on television and who generally cared more about the teacher aboard Challenger, and West Coast children, who heard about it first; between latency-age children and adolescents; and between children seen 5 7 weeks later and those same children seen 14 months later. RESULTS: More than 60% of the subjects feared at least one stimulus related to Challenger within the first 5-7 weeks of the explosion. The East Coast and latency-age groups appeared significantly more symptomatic than did the West Coast and adolescent groups. Over the 14-month study period, most symptoms dramatically faded. However, adolescents' diminished expectations for the future in general increased, and latency-age children's changed approach to space careers held relatively steady. Three East Coast latency-age children met the DSM-III-R symptom requirements for PTSD in 1986; no children met these in 1987. CONCLUSIONS: Children's symptomatic patterns after Challenger relate to the patterns for PTSD listed in diagnostic manuals and to three symptoms not in the DSM-IV list. To the authors, distant traumas appear to be one of a newly defined spectrum of trauma-related conditions that include relatively evanescent symptoms and a few longer-lasting ones. These symptoms may affect large numbers of normal children. PMID- 10518162 TI - Influence of child and adolescent psychiatric disorders on young adult personality disorder. AB - OBJECTIVE: This study examines associations between childhood psychopathology and young adult personality disorder in a random sample of 551 youths, who were 9 to 16 years old at first assessment. METHOD: Subjects were evaluated for DSM-III-R psychiatric disorders. Information was obtained prospectively from youths and their mothers at three points over 10 years. The predictive effects of prior axis I disorders and adolescent axis II personality disorder clusters A, B, and C on young adult personality disorder were examined in logistic regression analyses. RESULTS: The odds of young adult personality disorder increased given an adolescent personality disorder in the same cluster. Prior disruptive disorders, anxiety disorders, and major depression all significantly increased the odds of young adult personality disorder independent of an adolescent personality disorder. In addition, comorbidity of axis I and axis II disorders heightened the odds of young adult personality disorder relative to the odds of a disorder on a single axis. CONCLUSIONS: Assessment of personality pathology before late adolescence may be warranted. Childhood or adolescent axis I disorders may set in motion a chain of maladaptive behaviors and environmental responses that foster more persistent psychopathology over time. Identification and treatment of childhood disorder may help to reduce that risk. PMID- 10518165 TI - Affective responsiveness in borderline personality disorder: a psychophysiological approach. AB - OBJECTIVE: The aim of the study was to investigate affective responses to emotional stimuli in subjects with borderline personality disorder. METHOD: Twenty-four female patients with borderline personality disorder and 27 normal female comparison subjects were examined. The test stimuli were a set of standardized photographic slides with pleasant, neutral, or unpleasant emotional valence. In addition to self-reports, emotional reactions to the slides were measured by heart rate, skin conductance, and startle response. Psychometric tests for various aspects of impulsiveness were also completed. RESULTS: Neither self-report nor physiological data gave any evidence that the borderline patients showed more intense affective responses than did the normal subjects. The borderline subjects did not produce higher levels of startle amplitude, and while viewing unpleasant slides, they showed a startle potentiation effect that was largely similar to that of the comparison group. In fact, the borderline patients showed low electrodermal responses to all three stimulus categories, which points to physiological underarousal. CONCLUSIONS: The results do not agree with the hypothesis that there is a fundamental, biologically based affective hyperresponsiveness in borderline personality disorder, as is suggested by current theories of affect dysregulation in the disorder. Autonomic underarousal may seriously interfere with a flexible adaptation to environmental stimuli. PMID- 10518164 TI - Panic attacks and suicide attempts in mid-adolescence. AB - OBJECTIVE: The aim of this study was to investigate the association of panic attacks and suicide attempts in a community-based sample of 13-14-year-old adolescents. METHOD: The data are from a survey of 1,580 students in an urban public school system located in the mid-Atlantic region of the United States. Logistic regression methods were used to estimate associations between panic attacks and suicidal ideation and suicide attempts. RESULTS: Controlling for demographic factors, major depression, the use of alcohol, and the use of illicit drugs, the authors found that adolescents with panic attacks were three times more likely to have expressed suicidal ideation and approximately two times more likely to have made suicide attempts than were adolescents without panic attacks. CONCLUSIONS: This new epidemiologic research adds to the evidence of an association between panic attacks and suicide attempts during the middle years of adolescence. PMID- 10518166 TI - Expressed emotion and clinical outcome in borderline personality disorder. AB - OBJECTIVE: This longitudinal follow-up study examined the predictive validity of relatives' expressed emotion in a group of patients diagnosed with borderline personality disorder. METHOD: Thirty-five patients with DSM-III-R-diagnosed borderline personality disorder were followed up 1 year after they were discharged from a psychiatric hospital. Clinical outcome was assessed through interviews with patients and their family members. Expressed emotion in the patients' relatives, assessed at the time of the index admission, was then used to predict patients' subsequent clinical outcomes. RESULTS: Contrary to prediction, relatives' criticism and hostility did not predict how well patients did in the year after discharge. Neither did they predict rates of rehospitalization. Clinical outcome was strongly associated with family levels of emotional overinvolvement, however. Patients whose families scored higher on emotional overinvolvement had better clinical outcomes over the course of the follow-up period. CONCLUSIONS: These findings suggest that the association between expressed emotion and patient outcome may be different for patients with borderline personality disorder than it is for patients with schizophrenia or mood disorders. PMID- 10518167 TI - Effectiveness of partial hospitalization in the treatment of borderline personality disorder: a randomized controlled trial. AB - OBJECTIVE: This study compared the effectiveness of psychoanalytically oriented partial hospitalization with standard psychiatric care for patients with borderline personality disorder. METHOD: Thirty-eight patients with borderline personality disorder, diagnosed according to standardized criteria, were allocated either to a partially hospitalized group or to a standard psychiatric care (control) group in a randomized controlled design. Treatment, which included individual and group psychoanalytic psychotherapy, was for a maximum of 18 months. Outcome measures included the frequency of suicide attempts and acts of self-harm, the number and duration of inpatient admissions, the use of psychotropic medication, and self-report measures of depression, anxiety, general symptom distress, interpersonal function, and social adjustment. Data analysis used repeated measures analysis of covariance and nonparametric tests of trend. RESULTS: Patients who were partially hospitalized showed a statistically significant decrease on all measures in contrast to the control group, which showed limited change or deterioration over the same period. An improvement in depressive symptoms, a decrease in suicidal and self-mutilatory acts, reduced inpatient days, and better social and interpersonal function began at 6 months and continued until the end of treatment at 18 months. CONCLUSIONS: Psychoanalytically oriented partial hospitalization is superior to standard psychiatric care for patients with borderline personality disorder. Replication is needed with larger groups, but these results suggest that partial hospitalization may offer an alternative to inpatient treatment. PMID- 10518168 TI - Differences between clinical and research practices in diagnosing borderline personality disorder. AB - OBJECTIVE: It has been reported that clinicians are less inclined than researchers to use direct questions in ascertaining the presence of personality disorders, and questions have been raised about the validity of research on personality disorders in which diagnoses are based on semistructured diagnostic interviews. This study examined the influence of assessment method on the diagnosis of borderline personality disorder. METHOD: Diagnoses of borderline personality disorder derived from structured and unstructured clinical interviews were compared in two groups of psychiatric outpatients seen in the same practice setting. Five hundred individuals presenting to a general adult psychiatric practice for an intake appointment underwent a routine unstructured clinical interview. After the completion of that study, the method of conducting diagnostic evaluations was changed, and 409 individuals were interviewed with the borderline personality disorder section of the Structured Interview for DSM-IV Personality. RESULTS: Individuals in the structured interview group were significantly more often diagnosed with borderline personality disorder than individuals in the clinical group. When information from the structured interview was presented to the clinicians, borderline personality disorder was much more likely to be diagnosed by them. CONCLUSIONS: The method used to assess borderline personality disorder has a great impact on the frequency with which it is diagnosed. Without the benefit of detailed information from a semistructured diagnostic interview, clinicians rarely diagnose the disorder during a routine intake evaluation. Providing the results of a semistructured interview to clinicians prompts them to diagnose borderline personality disorder much more frequently. This is inconsistent with the notion that personality disorder diagnoses based on semistructured interviews are not viewed as valid by clinicians. PMID- 10518169 TI - Clinical and neurobiological correlates of cytogenetic abnormalities in childhood onset schizophrenia. AB - OBJECTIVE: Cytogenetic abnormalities are increased in schizophrenia, suggesting a possible etiologic contribution. However, their clinical and pathophysiologic roles in the disorder are unknown. To investigate this, a group of children and adolescents participating in a comprehensive study of childhood-onset schizophrenia were screened for chromosomal abnormalities, and their clinical and neurobiological correlates were examined. METHOD: Cytogenetic screening with the use of high-resolution banding, fluorescent in situ hybridization for chromosome 22q11 deletions, and molecular fragile X testing was undertaken in a group of 47 children and adolescents with very early onset of schizophrenia. Clinical, neurobiological (including brain morphometry), and risk factor measures of the subjects with cytogenetic abnormalities were compared with those of the remaining patients without cytogenetic anomalies. RESULTS: Five patients had previously undiagnosed cytogenetic abnormalities. Lower performance IQ and more pronounced premorbid developmental impairments were seen in this subgroup. Rates of obstetric complications, familial schizophrenia spectrum disorders, and familial eye tracking dysfunction were similar for the patients with and without cytogenetic abnormalities. CONCLUSIONS: Cytogenetic abnormalities appear to be increased in childhood-onset schizophrenia, suggesting an association with a very early age at onset. The data from the subgroup of patients with cytogenetic anomalies are consistent with a model in which a childhood onset of schizophrenia is due to a greater impairment of neurodevelopment secondary to the interaction of a number of factors, particularly genetic ones. PMID- 10518170 TI - Lamina-specific alterations in the dopamine innervation of the prefrontal cortex in schizophrenic subjects. AB - OBJECTIVE: Abnormalities in dopamine neurotransmission in the prefrontal cortex have been implicated in the pathophysiology of schizophrenia. However, the integrity of the dopamine projections to the prefrontal cortex in this disorder has not been directly examined. METHOD: The authors employed immunocytochemical methods and antibodies against tyrosine hydroxylase, the rate-limiting enzyme in dopamine biosynthesis, and the dopamine membrane transporter to examine dopamine axons in the dorsomedial prefrontal cortex (area 9) from 16 pairs of schizophrenic and matched control subjects. RESULTS: Compared to the control subjects, the total length of tyrosine hydroxylase-immunoreactive axons was unchanged in the superficial and middle layers of the schizophrenic subjects but was reduced by an average of 33.6% in layer 6. The total length of tyrosine hydroxylase-positive axons in layer 6 was decreased in 13 of the schizophrenic subjects compared to their control subjects. Axons immunoreactive for the dopamine membrane transporter showed a similar pattern of change. In contrast, axons labeled for the serotonin transporter did not differ between schizophrenic and control subjects in any layer examined. In addition, the density of tyrosine hydroxylase-containing axons did not differ between monkeys chronically treated with haloperidol and matched control animals. CONCLUSIONS: These findings reveal that schizophrenia is associated with an altered dopamine innervation of prefrontal cortex area 9 that is lamina- and neurotransmitter-specific and that does not appear to be a consequence of pharmacological treatment. Together, these data provide direct evidence for a disturbance in dopamine neurotransmission in the prefrontal cortex of schizophrenic subjects. PMID- 10518171 TI - Suicidal behavior in patients with schizophrenia and other psychotic disorders. AB - OBJECTIVE: Patients with schizophrenia are known to be at high risk for suicide attempts and dying by suicide. However, little research has been conducted to determine whether the risk for suicidal behavior is elevated among patients with psychosis in general. METHOD: This study evaluated 1-month and lifetime rates of suicidal behavior among 1,048 consecutively admitted psychiatric inpatients (ages 18 to 55 years) with DSM-III-R psychotic disorders. Demographic, clinical, and diagnostic correlates of suicidal behavior were examined. RESULTS: A high rate of suicidal behavior was found in the group: 30.2% reported a lifetime history of suicide attempts, and 7.2% reported a suicide attempt in the month before admission. The highest 1-month and lifetime rates were found in patients with schizoaffective disorder and major depression with psychotic features. Ratings of the medical dangerousness of the most recent suicide attempt on the basis of the extent of physical injury were higher in patients with schizophrenia spectrum psychoses. Agreement was high between emergency room assessments and semistructured interview assessments of suicidal behavior. CONCLUSIONS: Rates of suicidal behavior were high across a broad spectrum of patients with psychotic disorders; patients with a history of a current or past major depressive episode (as a part of major depressive disorder or schizoaffective disorder) were at a greater risk for suicide attempts, but patients with schizophrenia, on average, made more medically dangerous attempts. Risk factors for suicidal behavior in patients with psychosis appear to vary compared to those for the general population. PMID- 10518172 TI - Conjugal loss and syndromal depression in a sample of elders aged 70 years or older. AB - OBJECTIVE: The goal of this study was to describe the association between conjugal loss and both syndromal depression and depressive symptoms in a prospective cohort study of people aged 70 years or older. METHOD: A measure of syndromal depression, the shortform Composite International Diagnostic Interview (CIDI), and a revised version of the Center for Epidemiologic Studies--Depression Scale (CES-D Scale) were administered to a group of 5,449 elders in a longitudinal cohort study. The authors compared the rates of syndromal depression (CIDI diagnosis) and depressive symptoms (six CES-D Scale symptoms) in married participants and those who lost spouses between the first and second waves of assessment. RESULTS: The rate of syndromal depression in the newly bereaved was nearly nine times as high as the rate for married individuals, and the rate of depressive symptoms was nearly four times as high. The percentage of the bereaved respondents who had scores above threshold on the revised CES-D Scale was higher for those interviewed up to 2 years after loss of a spouse than for married respondents. Age, sex, prior psychiatric history, and the expectedness of the death did not differ between depressed and nondepressed newly bereaved subjects. CONCLUSIONS: Recent bereavement is a significant risk factor for syndromal depression in the elderly. Some widows and widowers experienced high levels of depressive symptoms up to 2 years after the loss of their spouses. Neither demographic variables nor variables concerning the nature of the spouse's death predicted bereavement-related depression. PMID- 10518173 TI - White matter hyperintensities and gray matter lesions in physically healthy depressed subjects. AB - OBJECTIVE: Previous studies reported that depressed subjects had more white matter hyperintensities on magnetic resonance imaging scans than control subjects, but the subjects had cerebrovascular disease risk factors. This study used subjects with a history of recurrent major depression and matched comparison subjects, screened to exclude cerebrovascular disease risk factors, to determine whether depressed subjects had more white matter hyperintensities and other lesions. METHOD: A semiautomated volumetric computer program was used to compare numbers and volumes of white matter hyperintensities, basal ganglia lesions, and total lesions in 24 women with a history of recurrent major depression and 24 comparison subjects case-matched on age and education and group-matched on height. In addition, images were measured with the use of a validated categorical scale. All subjects were screened to exclude cerebrovascular disease risk factors. RESULTS: There were no significant differences in the total volumes or total numbers of lesions. However, multiple linear regression showed a significant correlation of age and depression with number of lesions; this was accounted for by a greater number of small lesions (diameter < or = 0.4 cm). CONCLUSIONS: These findings suggest that cerebrovascular disease risk factors most likely mediated the relationship between depression and white matter hyperintensities seen in previous studies. However, the independent effect of depression, as well as an age-by-depression interaction, for small lesions suggests a causal role of depression in certain types of white matter pathology irrespective of other cerebrovascular disease risk factors. The volumetric method used in this study may be more sensitive than other methods in determining lesion characteristics and correlations with clinical variables. PMID- 10518174 TI - Treatment of primary dysthymia with group cognitive therapy and pharmacotherapy: clinical symptoms and functional impairments. AB - OBJECTIVE: This study assessed the efficacy of antidepressant treatment (sertraline) and group cognitive behavior therapy, alone or in combination, in primary dysthymia. The clinical features of dysthymia, as well as the functional impairments associated with the illness (e.g., quality of life, stress perception, coping styles), were evaluated. METHOD: Patients (N = 97) diagnosed with primary dysthymia, but no other current comorbid disorder, received either sertraline or placebo in a double-blind design over 12 weeks. In addition, a subgroup of the patients (N = 49) received a structured, weekly group cognitive behavior therapy intervention. RESULTS: Treatment with sertraline, with or without group cognitive behavior therapy, reduced the functional impairment of depression. The reductions were similar in the drug-cognitive therapy group and in subjects who received the drug alone. Furthermore, while group cognitive behavior therapy alone reduced the depression scores, this effect was not significantly greater than the effect of the placebo. The drug treatment also induced pronounced improvement in the functional measures, and in some respects these effects were augmented by group cognitive behavior therapy. Among patients who responded favorably to cognitive behavior therapy, the improvements in the functional measures were similar to those who responded to drug treatment, whereas such functional changes were not seen among patients who responded to placebo. CONCLUSIONS: Sertraline treatment effectively reduces the clinical symptoms and functional impairments associated with dysthymia. Although the group cognitive behavior therapy intervention was less effective in alleviating clinical symptoms, it augmented the effects of sertraline with respect to some functional changes, and in a subgroup of patients it attenuated the functional impairments characteristic of dysthymia. PMID- 10518175 TI - Cerebral blood flow changes associated with attribution of emotional valence to pleasant, unpleasant, and neutral visual stimuli in a PET study of normal subjects. AB - OBJECTIVE: To assist in the development of a model for the psychopathology of emotions, the present study sought to identify the neural circuits associated with the evaluation of visual stimuli for emotional valence. METHOD: Seventeen healthy individuals were shown three sets of emotionally laden pictures carrying pleasant, unpleasant, and neutral content. While subjects evaluated the picture set for emotional valence, regional cerebral blood flow was measured with the use of [15O] water positron emission tomography. Subjective ratings of the emotional valence of the picture sets were recorded. Data were analyzed by comparing the images acquired during the neutral condition with the unpleasant and pleasant image sets and the unpleasant and pleasant conditions with each other. RESULTS: Processing of pleasant stimuli was associated with increased blood flow in the dorsal-lateral, orbital, and medial frontal cortex relative to the unpleasant condition and in the cingulate, precuneus, and visual cortex relative to the neutral condition. Evaluation of unpleasant stimuli activated the amygdala, visual cortex, and cerebellum relative to the pleasant condition and the nucleus accumbens, precuneus, and visual cortex relative to the neutral condition. CONCLUSIONS: Observing and assigning emotional value to unpleasant stimuli produced activations in subcortical limbic regions, whereas evaluation of pleasant stimuli produced activations in cortical limbic areas. These findings are consistent with the notion of a subcortical and archaic danger recognition system and a system detecting pleasantness in events and situations that is phylogenetically younger, involving primarily the prefrontal cortex. PMID- 10518176 TI - A psychiatrist's reaction to a patient's suicide. PMID- 10518177 TI - Enhancement of CO2-induced anxiety in healthy volunteers with the serotonin antagonist metergoline. AB - OBJECTIVE: The mechanism of action of CO2-induced anxiety is unknown and has been little studied. The authors studied healthy volunteers for the possible influence of serotonin (5-HT) on CO2-induced anxiety. METHOD: Fourteen healthy volunteers received two vital capacity inhalations each of 35% CO2 and of air, preceded once by placebo and once by the 5-HT antagonist metergoline in a double-blind, randomized crossover design. RESULTS: Mean National Institute of Mental Health self-rating anxiety subscale scores increased nonsignificantly after CO2 inhalation; this effect was significantly enhanced by the administration of metergoline. CONCLUSIONS: The authors hypothesize that 5-HT may inhibit CO2 induced anxiety, a function that is lessened by metergoline. PMID- 10518178 TI - Compounds containing cytosolic choline in the basal ganglia: a potential biological marker of true drug response to fluoxetine. AB - OBJECTIVE: Studies have identified two types of antidepressant response: true drug response and placebo pattern response. This study examined the relationship between true drug response and choline-creatine ratios in the basal ganglia of depressed patients treated with fluoxetine. METHOD: The authors evaluated drug free outpatients with major depression before (N = 41) and after (N = 15) 8 weeks of fluoxetine treatment, 20 mg/day, by using proton magnetic resonance spectroscopy. RESULTS: There was a significant difference in the degree of change from baseline to week 8 in choline-creatine ratios between the true drug response group (N = 8) and the placebo pattern response/nonresponse group (N = 7); the true drug response patients had a 20% increase in choline-creatine ratios, and the placebo pattern response/nonresponse patients had a 12% decrease in choline creatine ratios. CONCLUSIONS: These data suggest that true drug response to fluoxetine treatment in depression may be associated with an increase in choline creatine ratios in the basal ganglia. PMID- 10518179 TI - Magnetic resonance assessment of cerebral perfusion in depressed cardiac patients: preliminary findings. AB - OBJECTIVE: The authors investigated the links between depression, cardiac disease, and microcirculatory cerebral blood flow (CBF). METHOD: A magnetic resonance imaging technique based on arterial spin tagging was used to estimate microcirculatory CBF in depressed (N = 5) and comparison (N = 14) elderly subjects with coronary artery disease. Signal intensity ratios corresponding to relative microcirculatory CBF were calculated for four regions on two axial images through the upper and lower halves of the lateral ventricles. RESULTS: On the superior image, estimates of microcirculatory CBF were statistically significantly lower on the left side in the depressed subjects than in the nondepressed group. When the ratios in the superior and inferior images were averaged, the depressed subjects had lower values for both left periventricular regions of interest and the parietal region. CONCLUSIONS: Despite the small study group and indirect estimates of blood flow, these preliminary findings suggest that a relative cerebral hypoperfusion may underlie depression in elderly cardiac patients. PMID- 10518180 TI - Depression and anxiety symptoms in women at high risk for breast cancer: pilot study of a group intervention. AB - OBJECTIVE: The psycho-oncology literature to date contains only one outcome study based on a group model for high-risk relatives of breast cancer patients. The authors set out to study the effects of group intervention in high-risk relatives of breast cancer patients. METHOD: Thirty-three high-risk relatives of breast cancer patients participated in a six-session, 12-hour group intervention model that consisted of educational and psychosocial components. RESULTS: There was a significant reduction of depression symptoms as reported on the Center for Epidemiologic Studies Depression Scale. Similarly, there was a significant reduction of anxiety symptoms as reported on the State-Trait Anxiety Inventory state scale. CONCLUSIONS: In this pilot study, the investigators found the group intervention model effective at reducing symptoms of depression and reactive (not chronic) anxiety. PMID- 10518181 TI - Comparison of ketamine-induced thought disorder in healthy volunteers and thought disorder in schizophrenia. AB - OBJECTIVE: This study sought to determine whether thought disorder induced in healthy volunteers by the N-methyl-D-aspartic acid (NMDA) receptor antagonist ketamine resembles the thought disorder found in patients with schizophrenia. METHOD: The Scale for the Assessment of Thought, Language, and Communication was used to assess thought disorder in healthy volunteers (N = 10) who received subanesthetic doses of ketamine and in a group of clinically stable inpatients with schizophrenia (N = 15) who did not receive ketamine. RESULTS: Mean scores on the Scale for the Assessment of Thought, Language, and Communication for patients with schizophrenia and healthy volunteers receiving ketamine did not differ significantly. Moreover, three of the four highest rated test items in both groups were the same. CONCLUSIONS: These data suggest that ketamine-induced thought disorder in healthy volunteers is not dissimilar to the thought disorder in patients with schizophrenia and provide support for the involvement of the NMDA receptor in a cardinal symptom of schizophrenia. PMID- 10518183 TI - Olanzapine-induced tardive dystonia. PMID- 10518182 TI - Obstetrical complications and childhood-onset schizophrenia. AB - OBJECTIVE: Increased obstetrical complications have been reported in individuals with adult-onset schizophrenia, with several studies finding an association between such complications and an earlier age at onset. Consequently, obstetrical records were examined for individuals with childhood-onset schizophrenia to determine if birth complications were more prevalent. METHOD: The birth records of 36 patients with childhood-onset schizophrenia and 35 sibling comparison subjects were rated for birth complications by two psychiatrists who were unaware of group membership. RESULTS: There were no significant differences between the groups in rates of obstetrical complications. Patients with such complications did not have a relatively earlier age at onset of schizophrenia. CONCLUSIONS: A very early age at onset of schizophrenia is probably not due to birth complications. PMID- 10518184 TI - Pain: cause of agitation in elderly individuals with dementia. PMID- 10518185 TI - Clozapine-induced stuttering: epileptic brain activity? PMID- 10518187 TI - Attention deficit hyperactivity disorder in the prison population. PMID- 10518186 TI - Sildenafil for sexual dysfunction in women taking antidepressants. PMID- 10518188 TI - Neuropsychological functions in patients with schizophrenia. PMID- 10518189 TI - Actual 5-year survival of patients with stage IIIB breast carcinoma: phase II trial of methotrexate, vinblastine, adriamycin, cisplatin, and folinic acid. AB - Patients with stage IIIB breast carcinoma represent only a small proportion of women with breast cancer in western countries but may constitute up to 50% of cases in underdeveloped countries. The prognosis remains poor despite aggressive treatment. Nineteen patients (11 with inflammatory breast carcinoma) received at least three courses of neoadjuvant chemotherapy of methotrexate, vinblastine, adriamycin, cisplatin, and folinic acid (MVAC/FA) followed by mastectomy. Six months of cyclophosphamide, methotrexate, and 5-fluorouracil were given after surgery. Radiation therapy followed chemotherapy. Seventy percent of patients achieved complete and 14% partial response after MVAC/FA chemotherapy alone. Eleven patients (58%) survived 5 years, and 30% survived at least 8 years. The addition of cisplatin in combination chemotherapy used as first-line treatment for stage IIIB breast carcinoma was well tolerated, resulted in higher response rates, and appeared to have an effect on overall survival. PMID- 10518190 TI - Breast cancer metastatic to the uterine cervix: analysis of a rare event. AB - A patient with cervical metastases leading to the diagnosis of a primary breast cancer is presented. To better define the frequency, symptoms, and characteristics of breast cancer metastasizing to the uterine cervix, all cases reported in the literature since 1950 are reviewed. This analysis reveals that in 21% of these cases, metastases were found before or at the same time as the primary tumor. The most common presenting symptom was abnormal vaginal bleeding (57%), but in 32% of the patients, no clinical sign was present. Because 41% of the reported cases were found only at autopsy, the characteristics of this kind of metastatic breast cancer, effectiveness of diagnostic procedures, and treatment options are critically discussed. PMID- 10518191 TI - Selective hepatic arterial chemoembolization for liver metastases in patients with carcinoid tumor or islet cell carcinoma. AB - In many patients with liver metastases from islet cell or carcinoid tumor, vascular occlusion therapy results in prolonged control of symptoms, biochemical response, and also tumor regression. Chemotherapy agents were added to evaluate safety and efficacy. Thirty patients with liver metastases from either carcinoid tumor or islet cell carcinoma underwent sequential vascular occlusion therapy combined with chemotherapeutic agents. In patients with carcinoid tumor, a combination of cisplatin (150 mg) and doxorubicin (50 mg) was used. In patients with islet cell carcinoma, a combination of 5-fluorouracil (350 mg) and streptozotocin (1000-2000 mg) was used. Sixteen patients had carcinoid tumor and 14 had islet cell carcinoma. Biochemical response was observed in 12 of 16 (75%) carcinoid patients and 9 of 10 (90%) islet cell patients. The overall partial response rate was 37% (11/30 patients). Partial response occurred in 4 of 16 (25%) patients with carcinoid tumor and 7 of 14 (50%) with islet cell carcinoma. The median duration of partial responses was 24 months (range, 6-63+ months). The median survival of all patients was 15 months (range, 2-67+ months). No treatment related deaths occurred. Our data suggest that the addition of these chemotherapeutic agents to vascular occlusion, although safe, has no additional benefit. PMID- 10518192 TI - Vinblastine pharmacokinetics in patients with non-small cell lung cancer given cisplatin. AB - The pharmacokinetics of vinblastine were studied in 16 patients with non-small cell lung cancer after a bolus intravenous dose of 3 mg/m2 given before or after cisplatin (100 mg/m2). Venous blood was collected at 0, 10, and 36 hr for analysis by radioimmunoassay. The mean plasma vinblastine concentration at 10 hr was similar when vinblastine was given before (4.8 ng/ml; 95% CI, 3.2-6.3) or after cisplatin (4.9 ng/ml; 95% CI, 2.7-7.1). Plasma vinblastine concentrations in patients given cisplatin were higher than previously reported in patients given vinblastine alone. Patients with plasma vinblastine concentrations less than 2.75 ng/ml at 10 hr experienced less severe neutropenia (37% fall in neutrophil count; 95% CI, 18-55) than those with levels greater than 2.75 ng/ml (69% fall in neutrophil count; 95% CI, 62-77). In conclusion, the pharmacokinetics of vinblastine predict the severity of neutropenia and may be altered when given in conjunction with cisplatin. PMID- 10518193 TI - Enhanced cytotoxic interaction between 5-fluorouracil and 4 hydroperoxycyclophosphamide against L1210 murine leukemic cells: applicability to ex vivo purging. AB - Eradication of contaminated tumor cells in bone marrow is a matter of utmost concern in the setting of autologous bone marrow transplantation. 4 Hydroperoxycyclophosphamide (4-HC) is often used for ex vivo chemical purging of contaminated tumor cells in bone marrow. The marrow from patients pretreated with 5-fluorouracil (5-FU) is enriched with multifactor-responsive high proliferative potential colony-forming cells. To develop an efficient ex vivo chemical purging system, we evaluated interaction between 4-HC and 5-FU. We investigated the antitumor effect of cyclophosphamide, a mother compound of 4-HC, and 5-FU against L1210 ascites tumor in B6D2F1 mice. The median lifespan of the mice treated with 4-HC or 5-FU alone was 8 and 12 days, respectively. The combination of both drugs significantly extended the median lifespan to 18.5 days. The median effect plot analysis indicated a synergistic cytotoxic interaction between 5-FU and 4-HC in 3 (4,5-dimethylthiazol-2-yl)-2,5-diphenyl terazolium bromide (MTT) assay. Clonogenic assay also showed that combination of 4-HC and 5-FU significantly reduced L1210 leukemic colonies to 20% of untreated control. Bone marrow cells from the mice treated with 5-FU at 150 mg/kg body weight was resistant to 4-HC at concentrations as high as 0.2 microgram/ml, which was more than 70% inhibitory concentration for colony formation in L1210 leukemic cells. Findings suggest that sequential treatment with in vivo 5-FU followed by ex vivo 4-HC could selectively enhance antitumor effects of 4-HC in tumor cells remaining in bone marrow. PMID- 10518195 TI - Development of clinical methods of iron deprivation for suppression of neoplastic and infectious diseases. AB - Hosts attempt to withhold growth-essential iron from neoplastic and microbial cells. This review focuses on the development of medical and pharmaceutical methods designed to enhance the iron-withholding defense system. PMID- 10518194 TI - 5-fluorouracil pharmacokinetics: causes for variability and strategies for modulation in cancer chemotherapy. PMID- 10518196 TI - Denial in cancer patients. AB - Denial is a basic mechanism for coping with stressful themes, common in healthy and sick individuals. This article deals with the role and functions of denial in cancer, reviewing empirical studies about the effects of denial on cancer prevention, screening, undergoing tests for early detection, delay in seeking medical attention and getting treatment, complying with medical instructions, and coping with the disease in different stages. Special sections are devoted to the possible role of denial as a risk factor for cancer, the effects of denial on disease course and survival, and the relation of denial to immunocompetence. Major conclusions are that denial may have a positive effect when applied in the first phase of coping, after diagnosis, because it reduces anxiety. This holds also for the terminal stage. The negative effects of denial are that it may interfere with getting treatment (e.g., delay in going to the doctor, not showing up for follow-ups, noncompliance), may disrupt the process of assimilating the stressful event, may affect adversely interpersonal relations, and constitutes a cumulative stressor depressing even immunocompetence. The use of denial varies with the severity of the situation, the patient's personality, and his or her familial and cultural background. A large body of research examined the hypothesis that a tendency toward denial could be one of the risk factors for cancer. Despite evidence supporting the occurrence of denial as a correlate of cancer, a lot of research is necessary to clarify the role of denial in general and of anger specifically as a factor affecting the occurrence of cancer and the course of disease and survival. PMID- 10518197 TI - Effect of managed care on community oncology clinical practice and research. PMID- 10518199 TI - Importance of chemotherapy scheduling: pieces of the puzzle. PMID- 10518200 TI - Single-dose granulocyte colony-stimulating factor concomitant with multifractionated dose chemotherapy: a strategy for maintaining dose intensity. AB - Multifractionated dosing (MFD) schedules for chemotherapy administration such as weekly or twice weekly administration are intended to maximize dose intensity while minimizing toxicity, but the cumulative drug effect may result in neutropenia necessitating interruption of dose fractions and thereby compromising dose intensity. Intermittent granulocyte colony-stimulating factor (G-CSF, Neupogen, Amgen) was administered to patients receiving MFD on three paclitaxel based chemotherapy regimens. Single-dose G-CSF was administered concomitant with the chemotherapy dose fraction when the white blood count was between 2000 and 3500 cells/microliter. A retrospective analysis of the concomitant administration of single-dose G-CSF with chemotherapy in these trials demonstrated that in most patients, G-CSF administration guided by the level or grade of leukopenia permitted maintenance of the chemotherapy dose intensity and completion of the treatment cycle. The common pattern in a six-dose, twice weekly, multifractionated cycle was for G-CSF to be administered with every other chemotherapy dose beginning with the third, fourth, or fifth dose, but some courses required G-CSF administration with each chemotherapy dose fraction. Guidelines for the concomitant use of G-CSF and paclitaxel-based MFD chemotherapy can be used to maintain chemotherapy dose intensity. A prospective study of guidelines for cytokine usage developed on the basis of this retrospective study will be necessary to determine the optimal cytokine dose and schedule for use simultaneously with chemotherapy. PMID- 10518198 TI - Pharmacokinetic modulation of 5-fluorouracil: emulating continuous infusion? PMID- 10518201 TI - Multifractionated dosing for cancer chemotherapy: a novel schedule and a work in progress. PMID- 10518203 TI - Dental students' sexual harassment experiences and attitudes. AB - The objective of this study was to describe the sexual harassment experiences and attitudes of students at the University of Kentucky College of Dentistry. A twenty-six-item questionnaire was developed and administered to 170 dental students in years one through four of the curriculum. Five questions explored students' personal experiences with sexual harassment--whether they had been harassed or had observed harassment; twenty-one questions addressed students' attitudes about sexual harassment. Computations of mean differences in Likert type scale responses for the twenty-one attitude items were completed using independent t-tests for the following variables: gender, whether respondents had been sexually harassed or had observed harassment of others, and years in dental education. Almost 15 percent of the students reported being sexually harassed at least once in dental college. Females were sexually harassed more often than males (p < .01), and second, third, and fourth year students more often than first year students (p < .05). Additionally, 30 percent of the students reported witnessing sexual harassment in the college. Harassers included faculty (88 percent), dental students (8 percent), and others (4 percent). Differences (p < .05) in sexual harassment attitudes were found when responses were analyzed by gender, by whether students had been sexually harassed, by whether they had witnessed harassment, and by years in dental education. The data show that sexual harassment occurs in the college. Dental faculty and students could benefit from programs to educate them about sexual harassment, how to prevent it from occurring, and how to respond if they are sexually harassed. PMID- 10518202 TI - Hematopoietic growth factors for chemotherapy-induced neutropenia. PMID- 10518204 TI - Educational implications for copyright in a digital world. AB - Statutory law and court cases currently leave fair use of copyrighted material poorly defined and fail to provide effective guidance for the use of others' work. Copyright legislation is undergoing significant change, accelerated by the evolution of computing and communication technologies. This paper reviews copyright issues, fair use guidelines, and applicable laws and statutes to help administrators and educators understand and comply with copyright regulations. The paper describes principles of copyright and ownership, the rights of copyright holders, and the conditions under which copyrighted material can be used by others. Recently introduced legislation, such as the 1998 Digital Millennium Copyright Act, may significantly affect how educators can use copyrighted material in the future. The integration of computer and communication technology into education raises a number of intellectual property issues for dental schools. This paper provides some general guidelines regarding copyright issues in academic environments. PMID- 10518205 TI - Employing oral examinations (viva voce) in assessing dental students' clinical reasoning skills. AB - The development and evaluation of a comprehensive oral examination system are described. These examinations are designed to measure, in an authentic manner, graduating students' understanding and use of the clinical reasoning and professional communications skills employed in the diagnosis, treatment, and management of realistic clinical situations. The examination cases were developed from actual clinic cases, enhanced to fit the requirements of the examination. Faculty members were provided with training sessions designed to promote consistency of administration and scoring. A mock oral examination was held to acquaint students with the process and to help reduce their anxiety. The results suggest that the oral examination is a reasonably valid and reliable approach to assessing clinical reasoning skills and that graduating students demonstrated competency in the areas tested. PMID- 10518206 TI - Sources of stress for Australian dental students. AB - The Dental Environment Stress questionnaire was used to identify and quantify sources of stress for 205 Australian Bachelor of Dental Surgery students. A factor analysis revealed negative self-efficacy beliefs accounted for almost one third of the total variance, and despite higher stress levels reported by females, a marked similarity in the dominant patterns emerged for males and females. In testing for differences in residency status, international students expressed significantly more stress from peer pressure, and this is discussed within a socio-cultural context. Irrespective of gender, residency status, and class year, students ranked examinations and grades as the single most stress inducing concern. Overall, stress intensity tended to escalate over time, peaking in the fourth year of training. It is suggested that dental students may be prone to unhealthy perfectionism, placing them at risk for the harmful consequences of chronically elevated stress levels. PMID- 10518207 TI - An intradisciplinary approach to nutrition education of dental and dental hygiene students. PMID- 10518208 TI - Dental education: an excellent investment for the government. PMID- 10518209 TI - Humanities in dental education: a focus on understanding the child. PMID- 10518210 TI - Mouse Lefty2 and zebrafish antivin are feedback inhibitors of nodal signaling during vertebrate gastrulation. AB - Mammalian lefty and zebrafish antivin form a subgroup of the TGF beta superfamily. We report that mouse mutants for lefty2 have an expanded primitive streak and form excess mesoderm, a phenotype opposite to that of mutants for the TGF beta gene nodal. Analogously, overexpression of Antivin or Lefty2 in zebrafish embryos blocks head and trunk mesoderm formation, a phenotype identical to that of mutants caused by loss of Nodal signaling. The lefty2 mutant phenotype is partially suppressed by heterozygosity for nodal. Similarly, the effects of Antivin and Lefty2 can be suppressed by overexpression of the nodal-related genes cyclops and squint or the extracellular domain of ActRIIB. Expression of antivin is dependent on Nodal signaling, revealing a feedback loop wherein Nodal signals induce their antagonists Lefty2 and Antivin to restrict Nodal signaling during gastrulation. PMID- 10518211 TI - Identification of the major intestinal fatty acid transport protein. AB - While intestinal transport systems for metabolites such as carbohydrates have been well characterized, the molecular mechanisms of fatty acid (FA) transport across the apical plasmalemma of enterocytes have remained largely unclear. Here, we show that FATP4, a member of a large family of FA transport proteins (FATPs), is expressed at high levels on the apical side of mature enterocytes in the small intestine. Further, overexpression of FATP4 in 293 cells facilitates uptake of long chain FAs with the same specificity as enterocytes, while reduction of FATP4 expression in primary enterocytes by antisense oligonucleotides inhibits FA uptake by 50%. This suggests that FATP4 is the principal fatty acid transporter in enterocytes and may constitute a novel target for antiobesity therapy. PMID- 10518212 TI - The C. elegans cell death specification gene ces-1 encodes a snail family zinc finger protein. AB - The ces-1 and ces-2 genes of C. elegans control the programmed deaths of specific neurons. Genetic evidence suggests that ces-2 functions to kill these neurons by negatively regulating the protective activity of ces-1, ces-2 encodes a protein closely related to the vertebrate PAR family of bZIP transcription factors, and a ces-2/ces-1-like pathway may play a role in regulating programmed cell death in mammalian lymphocytes. Here we show that ces-1 encodes a Snail family zinc finger protein, most similar in sequence to the Drosophila neuronal differentiation protein Scratch. We define an element important for ces-1 regulation and provide evidence that CES-2 can bind to a site within this element and thus may directly repress ces-1 transcription. Our results suggest that a transcriptional cascade controls the deaths of specific cells in C. elegans. PMID- 10518213 TI - The structural basis for the recognition of diverse receptor sequences by TRAF2. AB - Many members of the tumor necrosis factor receptor (TNFR) superfamily initiate intracellular signaling by recruiting TNFR-associated factors (TRAFs) through their cytoplasmic tails. TRAFs apparently recognize highly diverse receptor sequences. Crystal structures of the TRAF domain of human TRAF2 in complex with peptides from the TNFR family members CD40, CD30, Ox40, 4-1BB, and the EBV oncoprotein LMP1 revealed a conserved binding mode. A major TRAF2-binding consensus sequence, (P/S/A/T)x(Q/E)E, and a minor consensus motif, PxQxxD, can be defined from the structural analysis, which encompass all known TRAF2-binding sequences. The structural information provides a template for the further dissection of receptor binding specificity of TRAF2 and for the understanding of the complexity of TRAF-mediated signal transduction. PMID- 10518214 TI - Calnexin discriminates between protein conformational states and functions as a molecular chaperone in vitro. AB - Although calnexin is thought to function as a molecular chaperone for glycoproteins, a prevalent view is that it cannot distinguish between protein conformational states, binding solely through its lectin site to monoglucosylated oligosaccharides. Using purified components in vitro, calnexin effectively prevented the aggregation not only of glycoproteins bearing monoglucosylated oligosaccharides but also proteins lacking N-glycans, an effect enhanced by ATP. It also suppressed the thermal denaturation of nonglycosylated proteins and enhanced their refolding in conjunction with other cellular components. Calnexin formed stable complexes with unfolded conformers of these proteins but not with the native molecules. Therefore, in addition to being a lectin, calnexin functions as a bona fide molecular chaperone capable of interacting with polypeptide segments of folding glycoproteins. PMID- 10518216 TI - Directed evolution to bypass cyclin requirements for the Cdc28p cyclin-dependent kinase. AB - To identify cyclin-dependent kinase mutants with relaxed cyclin requirements, CDC28 alleles were selected that could rescue a yeast strain expressing as its only CLN G1 cyclin a mutant Cln2p (K129A,E183A) that is defective for Cdc28p binding. Rescue of this strain by mutant CDC28 was dependent upon the mutant cln2 KAEA, but additional mutagenesis and DNA shuffling yielded multiply mutant CDC28 BYC alleles (bypass of CLNs) that could support highly efficient cell cycle initiation in the complete absence of CLN genes. By gel filtration chromatography, one of the mutant Cdc28 proteins exhibited kinase activity associated with cyclin-free monomer. Thus, the mutants' CLN bypass activity might result from constitutive, cyclin-independent activity, suggesting that Cdk targeting by cyclins is not required for cell cycle initiation. PMID- 10518217 TI - A novel cofactor for p300 that regulates the p53 response. AB - The ability of p53 to function as a transcription factor is instrumental in facilitating the response to cellular stress, and p300/CBP proteins, which act as coactivators for diverse transcription factors, participate in regulating p53 activity. We report a novel cofactor for p300 that facilitates the p53 response by augmenting p53-dependent transcription and apoptosis. JMY and p300 associate in physiological conditions, and, during the cellular stress response, the p300/JMY complex is recruited to activated p53. The bax gene is efficiently activated by JMY, and protein isoforms that arise through alternative splicing alter the functional outcome of the p53 response. The results provide compelling evidence that the p300/JMY coactivator complex plays a central role in facilitating the p53 response. PMID- 10518215 TI - SLUG, a ces-1-related zinc finger transcription factor gene with antiapoptotic activity, is a downstream target of the E2A-HLF oncoprotein. AB - The E2A-HLF fusion gene transforms human pro-B lymphocytes by interfering with an early step in apoptotic signaling. In a search for E2A-HLF-responsive genes, we identified a zinc finger transcription factor, SLUG, whose product belongs to the Snail family of developmental regulatory proteins. Importantly, SLUG bears close homology to the CES-1 protein of C. elegans, which acts downstream of CES-2 in a neuron-specific cell death pathway. Consistent with the postulated role of CES-1 as an antiapoptotic transcription factor, SLUG was nearly as active as Bcl-2 or Bcl-xL in promoting the survival of IL-3-dependent murine pro-B cells deprived of the cytokine. We conclude that SLUG is an evolutionarily conserved transcriptional repressor whose activation by E2A-HLF promotes the aberrant survival and eventual malignant transformation of mammalian pro-B cells otherwise slated for apoptotic death. PMID- 10518218 TI - The nature of the nucleosomal barrier to transcription: direct observation of paused intermediates by electron cryomicroscopy. AB - Transcribing SP6 RNA polymerase was arrested at unique positions in the nucleosome core, and the complexes were analyzed using biochemical methods and electron cryomicroscopy. As the polymerase enters the nucleosome, it disrupts DNA histone interactions behind and up to approximately 20 bp ahead of the elongation complex. After the polymerase proceeds 30-40 bp into the nucleosome, two intermediates are observed. In one, only the DNA ahead of the polymerase reassociates with the octamer. In the other, DNA both ahead of and behind the enzyme reassociates. These intermediates present a barrier to elongation. When the polymerase approaches the nucleosome dyad, it displaces the octamer, which is transferred to promoter-proximal DNA. PMID- 10518220 TI - Proteasome active sites allosterically regulate each other, suggesting a cyclical bite-chew mechanism for protein breakdown. AB - In eukaryotes, the 20S proteasome contains two chymotrypsin-like, two trypsin like, and two active sites shown here to have caspase-like specificity. We report that certain sites allosterically regulate each other's activities. Substrates of a chymotrypsin-like site stimulate dramatically the caspase-like activity and also activate the other chymotrypsin-like site. Moreover, substrates of the caspase-like sites inhibit allosterically the chymotrypsin-like activity (the rate-limiting one in protein breakdown) and thus can reduce the degradation of proteins by 26S proteasomes. These allosteric effects suggest an ordered, cyclical mechanism for protein degradation. We propose that the chymotrypsin-like site initially cleaves ("bites") the polypeptide, thereby stimulating the caspase like sites. Their activation accelerates further cleavage ("chewing") of the fragments, while the chymotrypsin-like activity is temporarily inhibited. When further caspase-like cleavages are impossible, the chymotryptic site is reactivated and the cycle repeated. PMID- 10518219 TI - The FHA domain is a modular phosphopeptide recognition motif. AB - FHA domains are conserved sequences of 65-100 amino acid residues found principally within eukaryotic nuclear proteins, but which also exist in certain prokaryotes. The FHA domain is thought to mediate protein-protein interactions, but its mode of action has yet to be elucidated. Here, we show that the two highly divergent FHA domains of Saccharomyces cerevisiae Rad53p, a protein kinase involved in cell cycle checkpoint control, possess phosphopeptide-binding specificity. We also demonstrate that other FHA domains bind peptides in a phospho-dependent manner. These findings indicate that the FHA domain is a phospho-specific protein-protein interaction motif and have important implications for mechanisms of intracellular signaling in both eukaryotes and prokaryotes. PMID- 10518221 TI - Symmetrical mutant phenotypes of the receptor EphB4 and its specific transmembrane ligand ephrin-B2 in cardiovascular development. AB - Ephrin-B2 is a transmembrane ligand that is specifically expressed on arteries but not veins and that is essential for cardiovascular development. However, ephrin-B2 is also expressed in nonvascular tissues and interacts with multiple EphB class receptors expressed in both endothelial and nonendothelial cell types. Thus, the identity of the relevant receptor for ephrin-B2 and the site(s) where these molecules interact to control angiogenesis were not clear. Here we show that EphB4, a specific receptor for ephrin-B2, is exclusively expressed by vascular endothelial cells in embryos and is preferentially expressed on veins. A targeted mutation in EphB4 essentially phenocopies the mutation in ephrin-B2. These data indicate that ephrin-B2-EphB4 interactions are intrinsically required in vascular endothelial cells and are consistent with the idea that they mediate bidirectional signaling essential for angiogenesis. PMID- 10518222 TI - The length of the flexible SNAREpin juxtamembrane region is a critical determinant of SNARE-dependent fusion. AB - The topology of a SNARE complex bridging two docked vesicles could act as a mechanical couple to do work on the lipid bilayer resulting in fusion. To test this, we prepared a series of modified SNARE proteins and determined their effects on SNARE-dependent membrane fusion. When two helix-breaking proline residues are introduced into the juxtamembrane region of VAMP, there is little or no effect on fusion, and the same change in syntaxin 1A only reduced the extent and rate of fusion by half. The insertion of a flexible linker between the transmembrane domain and the conserved coiled-coil domain only moderately affected fusion; however, fusion efficiency systematically decreased with increasing length of the linker. Together, these results rule out a requirement for helical continuity and suggest that distance is a critical factor for membrane fusion. PMID- 10518223 TI - The N-terminal domain of the IP3 receptor gates store-operated hTrp3 channels. AB - In the present work, we studied the interaction and effect of several IP3 receptor (IP3R) constructs on the gating of the store-operated (SOC) hTrp3 channel. Full-length IP3R coupled to silent hTrp3 channels in intact cells but did not activate them until stores were depleted of Ca2+. By contrast, constructs containing the IP3-binding domain activated silent hTrp3 channels in unstimulated cells and restored gating of hTrp3 by IP3 in excised plasma membrane patches. We conclude that the N-terminal domain of the IP3R functions as a gate and is sufficient for activation of SOCs. The sensing and transduction domains of the IP3R are required to maintain SOCs in an inactive state. PMID- 10518224 TI - A Xenopus protein related to hnRNP I has a role in cytoplasmic RNA localization. AB - Cytoplasmic localization of mRNA molecules is a powerful mechanism for generating cell polarity. In vertebrates, one paradigm is localization of Vg1 RNA within the Xenopus oocyte, a process directed by recognition of a localization element within the Vg1 3' UTR. We show that specific base changes within the localization element abolish both localization in vivo and binding in vitro by a single protein, VgRBP60. VgRBP60 is homologous to a human hnRNP protein, hnRNP I, and combined immunolocalization and in situ hybridization demonstrate striking colocalization of hnRNP I and Vg1 RNA within the vegetal cytoplasm of the Xenopus oocyte. These results implicate a novel role in cytoplasmic RNA transport for this family of nuclear RNA-binding proteins. PMID- 10518225 TI - MSH2 and MSH6 are required for removal of adenine misincorporated opposite 8-oxo guanine in S. cerevisiae. AB - Oxidation of G in DNA yields 8-oxo-G (GO), a mutagenic lesion that leads to misincorporation of A opposite GO. In E. coli, GO in GO:C base pairs is removed by MutM, and A in GO:A mispairs is removed by MutY. In S. cerevisiae, mutations in MSH2 or MSH6 caused a synergistic increase in mutation rate in combination with mutations in OGG1, which encodes a MutM homolog, resulting in a 140- to 218 fold increase in the G:C-to-T:A transversion rate. Consistent with this, MSH2 MSH6 complex bound to GO:A mispairs and GO:C base pairs with high affinity and specificity. These data indicate that in S. cerevisiae, MSH2-MSH6-dependent mismatch repair is the major mechanism by which misincorporation of A opposite GO is corrected. PMID- 10518227 TI - TFIIA regulates TBP and TFIID dimers. AB - Dimerization of the TATA-binding protein (TBP) through its DNA-binding domain blocks TBP from accessing DNA and prevents unregulated gene expression. TFIIA plays a central role in loading TBP and its multisubunit counterpart TFIID onto promoter DNA, and it is therefore a candidate for regulating TBP/TFIID dimerization. Here, we show that TFIIA promotes the dissociation of TBP dimers directly and in doing so accelerates the kinetics of DNA binding. TFIID dimer dissociation was found to be slow and rate limiting in DNA binding. TFIIA induced a rapid dissociation of TFIID dimers, allowing TFIID to readily load onto promoter DNA. Together, these results suggest a novel mechanism by which TFIIA assists in regulating gene expression. PMID- 10518226 TI - The centromeric sister chromatid cohesion site directs Mcd1p binding to adjacent sequences. AB - Cohesion between sister chromatids occurs along the length of chromosomes, where it plays essential roles in chromosome segregation. We show here that the centromere, a cis-acting cohesion factor, directs the binding of Mcd1p, a cohesin subunit, to at least 2 kb regions flanking centromeres in a sequence-independent manner. The centromere is essential for the maintenance as well as the establishment of this cohesin domain. The efficiency of Mcd1p binding within the cohesin domain is independent of the primary nucleotide sequence of the centromere-flanking DNA but correlates with high A + T DNA content. Thus, the function of centromeres in the cohesion of centromere-proximal regions may be analogous to that of enhancers, nucleating cohesin complex binding over an extended chromosomal domain of A + T-rich DNA. PMID- 10518228 TI - An open label, randomized study to evaluate the effects of seven monophasic oral contraceptive regimens on hemostatic variables. Outline of the protocol. Oral Contraceptive and Hemostasis Study Group. AB - Complementary to the epidemiological knowledge on the association between oral contraceptive use and the occurrence of venous thromboembolism, a study was designed to obtain more conclusive data regarding the effect of estrogen dose and progestogen type of oral contraceptives on risk markers for the occurrence of venous thromboembolism. The protocol for this multicenter, randomized, open label, parallel group, comparative study is outlined in the present article. A total of 730 healthy, nonsmoking, mulliparous women were treated for six cycles with one of the seven monophasic oral contraceptives tested in this study. The effects of progestogen type (desogestrel, gestodene, levonorgestrel, and norgestimate) and the effects of ethinyl estradiol dose (50, 30, and 20 micrograms) on various hemostatic variables was assessed, including the key components of the anticoagulant and fibrinolytic system, as well as the coagulation system. The primary outcome variables in the study were prothrombin fragment 1 + 2 and D-dimer. PMID- 10518229 TI - Preferred frequency and characteristics of menstrual bleeding in relation to reproductive status, oral contraceptive use, and hormone replacement therapy use. AB - This study addresses attitudes towards changes in menstrual bleeding patterns caused by oral contraceptives (OC) or hormone replacement therapy (HRT) and preferred changes in bleeding pattern with and without use of OC or HRT in relation to reproductive age group. Data were collected by means of telephone interviews with 325 women in each of four age groups (15-19, 25-34, 45-49, and 52 57 years). In total, 80.5% of currently menstruating women preferred one or more changes in bleeding pattern such as less painful, shorter, or less heavy periods, or amenorrhea. The majority of the menstruating women in all age groups preferred to have a bleeding frequency of less than once a month or never, whether the bleeding was spontaneous or induced by OC. In the case of HRT, amenorrhea was most preferred. These findings with respect to preferred bleeding frequency and OC may have important implications for health care providers and for future contraception development. PMID- 10518230 TI - Prevalence of visible disruption of cervical epithelium and cervical ectopy in African women using Depo-Provera. AB - Associations between Depo-Provera (injectable, progesterone-only contraceptive) use and visible disruption of cervical epithelium and cervical ectopy were investigated using data collected as part of a cervical cancer screening study in periurban Cape Town, South Africa. Women were interviewed about their contraceptive use, and underwent a gynecologic examination that included two 35 mm photographs of the cervix after application of 5% acetic acid. Photographs of 723 subjects were reviewed (blind to clinical information and using systematic criteria developed before review) for evidence of atrophy and epithelial disruption, including inflammation and ulceration. The percentage of the cervix covered with columnar epithelium (ectopy) was also estimated from the photographs. A random sample of 85 photographs was reviewed again for reliability. A total of 121 current users of Depo-Provera were no more likely to have evidence of epithelial disruption (38%) than 574 nonusers (39%), odds ratio (OR) = 1.37, 95% CI: 0.89-2.11 adjusting for age and parity. The prevalence of significant ectopy (columnar epithelium covering > 10% of the cervix) was also no different among current Depo-Provera users (OR = 1.22, 95% CI: 0.80-1.86 adjusting for age and parity). Reliability of visual scoring of epithelial disruption and ectopy was excellent (kappa = 0.8). Although the underlying prevalence of visible disruption of cervical epithelium was very high, current use of Depo-Provera was not associated with increased prevalence of visible disruption of the cervical epithelium or with ectopy in this sample of African women. PMID- 10518231 TI - Acceptability of once-a-month injectable contraceptives Cyclofem and Mesigyna: focus group discussion. AB - This article presents the results of focus group discussion sessions conducted with a sample of continuers and discontinuers of the once-a-month injectable contraceptives Cyclofem and Mesigyna, as well as with some of the physicians who participated in the clinical trial. The sessions helped explain certain aspects related to the clients as well as the service delivery setting that affected the acceptability of these contraceptive preparations. Providing counseling and presence of a dedicated clinic staff were very essential to the success of the clinical trial. Social support in the form of both husband knowledge, approval, and support as well as the presence of other close relative or friend users helped in increasing the woman's tolerance of side effects and sometimes helped overcome adverse rumors. Reaction of the women to vaginal bleeding in an Islamic society is discussed and the need for health education in this aspect is strongly recommended. The two preparations are best suited for women who were satisfied with the use of combined hormonal contraceptive pills but had a problem with compliance, and users of other hormonal methods who are free from menstrual irregularities. On the whole, the two preparations had high acceptability among Egyptian women, provided that procedures for the selection of the appropriate candidate are adhered to, and that pre- and postservice counseling and social support are present. PMID- 10518232 TI - A 5-year clinical evaluation of Norplant implants in Senegal. AB - The principal objective of this 5-year clinical study of Norplant implants was to introduce these implants into the family planning program in Senegal and to determine their overall acceptability and safety in Senegalese acceptors. A total of 300 subjects were enrolled into the trial from August 1986 to July 1991. All the women were followed-up for 5 years or until the implants were removed. The pooled cumulative discontinuation rate was 40.8 +/- 2.91 per 100 women resulting in a continuation rate of 59.2 +/- 2.91 per 100 women. Thirteen subjects (4.3%) were lost during the follow-up. Seven pregnancies were reported throughout the 5 years leading to a cumulative pregnancy rate of 3.3 +/- 1.25 per 100 women. Menstrual problems were the reason most often given for early removal during the first 2 years. After year 2, desire for another pregnancy was the main reason for implant removal. The results presented in this study show that the Norplant implant system is a safe, effective, and acceptable method that meets the needs of the Senegalese family planning program. PMID- 10518233 TI - "Pop-out" method of levonorgestrel implant removal. AB - The "pop-out" technique is a method of levonorgestrel implant removal that uses digital pressure to direct implants through a small skin incision. This technique was developed, theoretically, to cause less bruising and patient discomfort by avoiding the use of instruments. The pop-out technique is the primary method used for levonorgestrel implant removal in the Magee-Womens Hospital resident clinic. We performed a retrospective analysis of levonorgestrel implant removals performed between July 1, 1995, and December 31, 1998. Of the 168 removals included in this analysis, 38 were performed by one of two attending physicians, and 130 were performed by the residents with attending supervision. The average time for removal was 12 +/- 5 min (range 2.25-27 min) when the "pop-out" method could be used to remove all six implants, and 14 +/- 7 min (range 2.25-59 min) for all removals. The removal time for residents was inversely proportional to the anticipated level of difficulty of the removal and to the number of previous removals performed. The removal time was significantly faster when residents were supervised by one of the attending physicians as compared with the other attending physician. Only 0.7% (7/1,008) of levonorgestrel implants were fragmented during removal. This review shows that the "pop-out" method is a reasonable alternative to other proposed methods of primary implant removal. The difference in the level of expertise of the attending physician may significantly influence removal time when training clinicians in levonorgestrel implant removal. PMID- 10518234 TI - Methotrexate and misoprostol used alone or in combination for early abortion. AB - The purpose of this study was to compare the outcome and side effects of using the drugs methotrexate and misoprostol, alone or in combination, to induce abortion. A total of 108 subjects who had requested elective termination of pregnancy and medical abortion at 9 weeks gestation or less were randomized into three groups. The first group received 50 mg/m2 intramuscular (i.m.) methotrexate on day 1 and, if the hCG level had risen by > 50% of the initial level on day 4, a second dose was given. They were then followed-up at weekly intervals up to day 21. Group 2 received 800 micrograms vaginal misoprostol on day 1 and, if ultrasound showed a gestational sac on day 4, they received a repeat dose and were re-examined on day 7. Group 3 received 50 mg/m2 methotrexate intramuscularly followed 3 days later by 800 micrograms vaginal misoprostol and were re-examined on day 7. Complete abortion occurred in 25 (69%) of the 36 subjects in group 1, 21 (58%) of the 36 subjects in group 2, and 32 (89%) of the 36 subjects in group 3. The complete abortion rate in group 3 was significantly higher than that of both group 1 and group 2 (p < 0.05). The incomplete abortion rate was significantly higher in group 2 as compared with both of the other groups (p < 0.05). There were significant differences between the mean gestational age of the successful abortions and the failures in group 1 (no abortion occurred at more than 49 days gestation), but not in groups 2 or 3. Vaginal bleeding in subjects who successfully aborted began within 16 +/- 4 days in group 1 after the first dose, and within 24 h in 18 (86%) of the 21 subjects in group 2 and 27 (84%) of the 32 subjects in group 3 after the misoprostol dose. The drugs caused no serious or prolonged side effects. The combination of methotrexate and misoprostol is a more effective abortifacient regimen than when either drug is used alone. PMID- 10518235 TI - Effects of castor bean extract and ricin A-chain on ovulation and implantation in rabbits. AB - The anti-implantation and antiovulation effects of castor bean extract (CBE) and ricin A-chain (RAC) were evaluated in rabbits. Both CBE and RAC, administered intraperitoneally on days 5-9 of pregnancy, exhibited a pronounced decrease in maternal body weight gain and in death of all fetuses. A significant (p < 0.01) decrease of implantation sites resulted after rabbits were treated with RAC on the first 6 consecutive days of pregnancy. When female rabbits were treated with RAC for 10 consecutive days followed by human chorionic gonadotropin (hCG) (50 IU/kg intravenously), there was a 30% reduction in the number of corpora lutae. These data clearly indicate that CBE and RAC possess potent effects on implantation and ovulation in rabbits. The protein contents of castor bean extract, separated by polyacrylamide gel electrophoresis, revealed the presence of several protein bands, ricin toxin being a major constituent of the extract. PMID- 10518236 TI - Changes in daily sperm production rate in rats under the influence of a potent antispermatogenic agent, CDRI 84/35. AB - CDRI 84/35, a potent nonsteroidal antispermatogenic agent, causes total sterility in rats by directly acting on germ cells while having no effect on Sertoli/Leydig cells. This study was conducted to evaluate the effect of the compound on gametogenic activity of testes and to identify stages of spermatogenesis that were affected. Adult male rats administered either compound 84/35 at minimum effective dose or estradiol (5 micrograms) or water only were killed on days 22, 41, and 64 of the treatment period to evaluate the effect on spermatid, spermatocyte, and spermatogonial stages, respectively. Daily sperm production (DSP) was measured employing a homogenization technique. Results showed a decline in testis weight and DSP with a drastic reduction (approximately 95%) in DSP in 84/35-treated rats on day 41 of the treatment period. Estradiol was more potent in reducing the testis weight; however, 84/35 had an edge over estradiol in reducing the DSP. After withdrawal of treatment for 120 days, a phenomenal recovery (> 90%) in DSP per gram parenchyma was noted in 84/35-treated animals. Results indicate a direct effect of estradiol on spermatogonia, whereas 84/35 seems to affect the spermatocyte stage. PMID- 10518237 TI - Preparation of transparent injectable formulation for lipid A analog E5531. AB - We developed a "pH-jump method" to obtain transparent solutions of E5531, a synthetic lipid A analog, at neutral pH for pharmaceutical injection. E5531 has two pKa: pKa1 = 6.0 and pKa2 = 9.3. At pH 11.0, E5531 was dispersed as a dissociated form, and the phase transition temperature Tc of E5531 was determined to be 30 degrees C using differential scanning calorimetry (DSC). Based on these results, the pH-jump method procedure involves dispersing E5531 at pH 11.0 (above pKa2) at 50 degrees C (above Tc) and mixing with a phosphate buffer to neutralize the pH. No degradated products of E5531 were observed at 50 degrees C for 3 hr during dispersing in the alkaline solution. The turbidity of samples prepared with the pH-jump method was similar to that of water and superior to the samples dispersed directly in neutral pH. This method does not need mechanical power and is suitable for large-scale production. PMID- 10518238 TI - Stability and in vitro drug release of flurbiprofen-loaded poly-epsilon caprolactone nanospheres. AB - The effects of temperature and two different initial pH (2.67 and 7.00) on poly epsilon-caprolactone (P epsilon CL) nanospheres loaded with flurbiprofen (FB) (aqueous suspensions) were studied to investigate their influence on the stability and physicochemical characteristics of these drug delivery systems. The drug release behavior was also studied. Release of the associated FB occurred very fast on high dilution in a buffered medium. The stability of the polymeric system depends on the temperature and the initial pH value; it is more degradable with the particles stored at 40 degrees C with an initial pH value of 2.67. PMID- 10518239 TI - Sustained-release interleukin-12 microspheres in the treatment of cancer. AB - Interleukin-12 (IL-12) is a recently discovered cytokine with tremendous antitumor potential. It has been shown to boost the host immune response against experimental cancers in animal models. However, most studies have utilized IL-12 in the solution form, necessitating frequent dosing with higher doses, consequently leading to issues of toxicity. The only attempts at sustaining release have been in the production and use of genetically engineered cells that can secrete IL-12 constantly. These attempts are cost prohibitive and involve extensive labor. This study demonstrates the use of biodegradable albumin microspheres to sustain the release of IL-12. In vitro release of IL-12 from the microspheres was found to fit Higuchi's square-root-of-time model, suggesting diffusion-mediated release. About 46% of the theoretical IL-12 content was released slowly over a period of 24 hr. When administered intraperitoneally to C57BL/6 mice bearing subcutaneous melanomas, the microspheres significantly prolonged the survival when administered at half the weekly dose of the solution formulation. The microsphere dosage form also resulted in generally lower levels of liver and kidney function enzymes, suggesting lower toxicity. PMID- 10518240 TI - Stability of freeze-dried tablets at different relative humidities. AB - The purpose of the study was to evaluate the stability of two different freeze dried tablet formulations at different relative humidities (RHs). The tablets contained 25 mg hydrochlorothiazide (HCT) as a model drug and were prepared by freeze-drying a suspension and an oil-in-water (o/w) emulsion. Formulation A was a rapidly disintegrating tablet and consisted of 80 mg of maltodextrine DE38; 8 mg of polyethyleneglycol (PEG 6000), 8 mg of xanthan gum, and 25 mg of HCT. Formulation B was a lyophilized dry emulsion tablet that consisted of 160 mg of Miglyol 812, 80 mg of maltodextrin DE38, 16 mg of methylcellulose (Methocel) A15LV, and 25 mg of HCT. Tablets were packaged in different packing materials: polyvinylchloride (PVC)/aluminum blister packs, PVC-polyvinylidenechloride (PVDC)/aluminum blister packs, closed containers with a dessicant tablet, and open containers. The tablets were stored at three relative humidities (45%, 60%, and 85% RH) and were characterized on mechanical strength, residual moisture, porosity, content uniformity, and scanning electron microscopy (SEM) during a period of 6 months. After 1 month at 60% and 85% RH, a strong increase in moisture content (from 2.7% to 6.8%) was seen for the tablets packed in the open and closed containers and for the PVC/aluminum blistered tablets. This increase was higher for formulation A compared to formulation B since B contained 160 mg of triglycerides and was more hydrophobic. This increase in water content was correlated with a decrease in mechanical strength. The tablets also showed a change in microstructure and porosity. At a moisture content of 7.2%, formulation A showed a structural "collapse" since water acts as a plasticizer for the amorphous glass, lowering the glass transition temperature Tg. This phenomenon even occurred in PVC/aluminum blister packs at 85% RH. The structural collapse was associated with a complete loss of microstructure as detected by porosimetric analysis and SEM. For the PVC-PVDC/aluminum blistered tablets, the increase in moisture content and decrease in mechanical strength at 85% RH occurred much slower, and the water uptake and strength loss were less intensive. No significant breakdown of HCT could be observed in both formulations with all of the packing materials. Packaging of freeze-dried tablets with PVC/aluminum blister packs, PVC/PVDC/aluminum blister packs, or closed containers did not offer protection against moisture uptake, mechanical strength loss, and structural collapse. PMID- 10518242 TI - The effect of polymorphism on powder compaction and dissolution properties of chemically equivalent oxytetracycline hydrochloride powders. AB - In South Africa, oxytetracycline is identified as an essential drug; many generic products are on the market, and many more are being developed. In this study, six oxytetracycline hydrochloride powders were obtained randomly from manufacturers, and suppliers were compared. It was found that compliance to a pharmacopoeial monograph was insufficient to ensure the optimum dissolution performance of a simple tablet formulation. Comparative physicochemical raw material analysis showed no major differences with regard to differential scanning calorimetry (DSC), infrared (IR) spectroscopy, powder dissolution, and particle size. However, the samples could be divided into two distinct types with respect to X ray powder diffraction (XRD) and thus polymorphism. The two polymorphic forms had different dissolution properties in water or 0.1 N hydrochloride acid. This difference became substantial when the dissolution from tablets was compared. The powders containing form A were less soluble than that containing form B. PMID- 10518241 TI - Optimization of crushing strength and disintegration time of a high-dose plant extract tablet by neural networks. AB - Optimization of crushing strength and disintegration time of a high-dose plant extract tablet was reached after extensive experimentation. Effects of the processing parameters, like compression force and tooling, and also of the excipients were found to be significant. Best results for both disintegration time and crushing strength were obtained with a plant extract that was granulated by roller compaction before compression. To gain more information about the different effects, artificial neural networks (ANNs) and a conventional multivariate method (partial least squares [PLS]) were used for data analysis. The topologies of the neural networks of the feed-forward type were optimized manually and by pruning methods. All methods were tested for contemplated parameters, crushing strength, and disintegration time. In general, ANNs were found to be more successful in characterizing the effects that influence crushing strength and disintegration time than the conventional multivariate methods. PMID- 10518243 TI - Design and evaluation of a new transdermal formulation containing chlorpheniramine maleate. AB - The antihistamine chlorpheniramine maleate (CPM) is used for symptomatic relief of hypersensitive reactions and in pruritic skin disorders. The objective of the present study was to develop a topical formulation that contained CPM to increase patient compliance. Compliance was increased by exploiting foams that, given their application methods, avoid direct contact with the afflicted area. The study also aimed to optimize the permeability of the CPM by discerning an adequate carrier, as well as choosing the correct enhancer. The foams were formulated using aqueous solutions. In vitro studies were carried out using Franz cells with the formulations, as well as with the available pharmaceutical product Polarmin Crema, which contains CPM. These studies showed that the permeability of the CPM in the solutions is increased more then 100 times with respect to the water-in-oil emulsion Polarmin Crema. In particular, the highest permeability was obtained using limonene as an enhancer. PMID- 10518244 TI - Storage conditions for serum deslorelin. AB - The stability of a luteinizing hormone-releasing hormone (LHRH) analog in rat serum was studied under reproduced experimental analysis conditions. Serum samples of deslorelin [D-Trp6, Des-Gly, NH2(10)] LHRH ethylamide were exposed to multiple freeze-thaw cycles to determine the maximum number of cycles the serum sample can be exposed to without producing any quantitative changes in radioimmunoassay (RIA) measurements of deslorelin. A significant cycle effect was observed after completion of the sixth cycle. Serum samples were also stored at standardized -50 degrees C conditions for variable periods of time to determine the effects of acute and chronic storage time on deslorelin stability. Matched pair t-test analysis showed no significant changes in deslorelin values as measured by RIA for a 3-week, 4-month, or 2-year storage period. The conditions in which the rat serum samples were stored prior to analytical analysis were sufficient to prevent detectable degradation of the deslorelin peptide. PMID- 10518245 TI - Kinetic release of theophylline from hydrophilic swellable matrices. AB - The objective of this research was to evaluate the effect of hydroxypropylmethylcellulose (HPMC; Methocel K4M Premium) level and type of excipient on theophylline release and to attempt to predict the drug release from hydrophilic swellable matrices. Formulations containing theophylline anhydrous (10% w/w), Methocel K4M Premium (10%, 30%, and 40% w/w), different diluents (Lactose Fast Flo, Avicel PH-101, and Emcompress), and magnesium stearate (0.75% w/w) were prepared by direct compression at a target weight of 450 mg +/- 5% and target hardness of 7 kp to 10 kp. It was found that, as the percentage of polymer in all formulations increased from 10% to 30% or 40%, the drug release decreased. However, there was no significant difference in drug release between formulations containing 30% polymer and formulations containing 40% polymer. At low levels of polymer, the drug release is controlled by the type of diluent used. Avicel PH 101 formulation gave the highest release, while its corresponding Emcompress formulation gave the lowest release. Formulations containing 30% or 40% polymer gave the same release profiles irrespective of the type of diluent used. In all cases, replacement of a portion of Methocel K4M Premium with any diluent resulted in increase of theophylline release. In addition, this investigation demonstrated that the drug release from hydrophilic swellable matrices can be predicted using only a minimum number of experiments. PMID- 10518246 TI - Evaluation of dosage forms. VI. Studies of commercial oxyphenbutazone tablet dosage forms. AB - Dissolution-dialysis studies of commercial tablets of oxyphenbutazone were carried out to establish the applicability of this technique for the in vitro evaluation of oxyphenbutazone dosage form. While disintegration time and dissolution rate studies did not give a true indication of bioavailability, an excellent correlation was obtained between the dialysis rate constant K and the pharmacokinetic parameters AUC and Cmax. PMID- 10518247 TI - Effect of various salts on the stability of lansoprazole, omeprazole, and pantoprazole as determined by high-performance liquid chromatography. AB - A fast and reproducible reverse-phase high-performance liquid chromatography (HPLC) assay method has been developed for the simultaneous quantitation of omeprazole, lansoprazole, and pantoprazole. The three compounds were monitored at 280 nm using Zorbax Eclipse XDB C8 (5 microns, 150 cm x 4.6 mm i.d.) and a mobile phase consisting of 700:300 phosphate buffer:acetonitrile with the pH adjusted to 7.0 with phosphoric acid. The method was used to study the effect of pH and various salts on the stability of the three compounds. The pH rate profile curve showed that pantoprazole was the most stable compound and lansoprazole the least stable. The stabilities of the compounds in salt solutions were found to be in the following order: phosphate buffer < trisodium citrate < citrate buffer < or = acetate buffer < citric acid < or = monosodium citrate < or = calcium carbonate < sodium bicarbonate < sodium chloride < water. The rate of degradation had a direct relationship with the H+ and salt concentration. PMID- 10518248 TI - Studies of hydroxypropyl methylcellulose donut-shaped tablets. AB - Simple uncoated compressed tablets with a central hole (donut shape) or multihole tablets were prepared. Theophylline and diltiazem hydrochloride were used as model drugs to investigate in vitro drug release from donut-shaped tablets. The effects of hole size, the number of holes, drug solubility, and stirring rate on release kinetics were investigated. As for the donut-shaped tablets, the duration of zero-order drug release could be up to 80-90%. When the hole size was increased, the release rate increased, and the duration of linear drug release was longer. The durations of linear drug release of two-hole and three-hole tablets were longer than that of the single-hole tablets. As the drug solubility increased, the duration of linear drug release was shortened. However, three stirring rates (50 rpm, 100 rpm, 150 rpm) had little effect on the drug release. PMID- 10518249 TI - Nandrolone decanoate is a good alternative for the treatment of anemia in elderly male patients on hemodialysis. AB - OBJECTIVE: To assess the efficiency of nandrolone decanoate (ND) in the control of anemia in elderly male patients on hemodialysis (HD), and to determine its influence on nutritional parameters. DESIGN: A prospective 6-month study with randomization of patients on recombinant human erythropoietin (rHuEPO) therapy into two groups. Group A: rHuEPO was stopped before starting treatment with ND. Group E: rHuEPO was continued. SETTING: Renal unit, tertiary-care center. PATIENTS: 14 male patients in Group A and 19 patients, 12 males and 7 females, in Group E. INTERVENTIONS: In Group A rHuEPO was stopped and ND 200 mg/I.M. weekly was given over six months. RESULTS: The increase in hemoglobin and hematocrit levels was progressive with ND, and significant differences (p < 0.003) were evident after six months (9.6 +/- 1.0 vs 11.0 +/- 1.4 and 28.9 +/- 4.7 vs 33.0 +/ 4.7, respectively), which remained unmodified in Group E. Group A showed a significant increase in serum creatinine, total protein, transferrin and anthropometric parameters. These parameters remained stable and even presented a tendency to decrease in Group E. There was a significant rise in the concentration of triglycerides and a significant decrease in both HDL-cholesterol and apolipoprotein A-1 in Group A. However, lipoprotein (a) decreased significantly. No significant changes were detected in Group E. CONCLUSIONS: The use of ND would allow us an acceptable treatment of anemia as well as a better nutritional condition in elderly male patients on dialysis. PMID- 10518250 TI - Monitoring hypovolemia in healthy elderly subjects by measuring blood pressure response to Valsalva's maneuver. AB - Quantification of hypovolemia by physical examination has limited validity. We explored the use of non-invasive measurement of blood pressure (BP) response to Valsalva's maneuver in monitoring hypovolemia in nine healthy elderly volunteers, recruited from participants of the Nijmegen Annual Four-Days Marches. Heart rate (HR), systolic and diastolic BP, and mean arterial pressure (MAP) response (Finapres) to a Valsalva's maneuver as well as clinical and laboratory assessment of fluid balance were determined 5 minutes before, and 3, 5, and 48 hours after administration of 40 mg furosemide orally. Subjects' (4 males aged 74.2 +/- 3.0 years) weight was 66.1 +/- 9.7 kg, mean BP was 139 +/- 21 over 76 +/- 12 mm Hg. A maximum weight loss of -2.8 +/- 0.9% occurred 5 hours after furosemide administration. Systolic and diastolic BP, HR, clinical assessment scores, and serum creatinine and urea nitrogen did not change during the total study period. Significant changes occurred in Valsalva phase I to phase II systolic BP response (difference +14.2 +/- 11.3 mm Hg, ratio difference -0.09 +/- 0.07 after 5 hours, P < 0.01). Changes after 48 hours did not differ from baseline values. Finapres measurement of Valsalva BP response may be useful in monitoring hypovolemia in the elderly. PMID- 10518251 TI - Renal artery disease in the elderly. AB - This brief review discusses the problem of atherosclerotic renal artery obstruction in the elderly. This disorder is common in the elderly; the overall incidence is estimated to be 10%. The disorder presents with new onset hypertension, a loss of control of BP or a decline in renal function in some patients. In others, the obstruction may be unmasked by the use of angiotensin converting enzyme inhibitors or angiotensin receptor blocker agents. The current approach to the diagnosis of renal artery obstruction is discussed as are the indications for invasive procedures. Careful patient selection for any invasive procedures is particularly important in the elderly since this population has a propensity to higher morbidity. PMID- 10518252 TI - Geriatric urinary incontinence. AB - Urinary incontinence is abnormal at any age. The prevalence of urinary incontinence increases with age due to functional impairments and concurrent medical disease. A detailed history and physical is essential in evaluating these patients. Urinary incontinence is treatable in all age groups when a logical, multifactorial and persistent approach is undertaken. PMID- 10518253 TI - The geriatric patient with obstructive uropathy. PMID- 10518254 TI - Geriatric issues in renal transplantation. AB - There are an increasing amount of data which are compelling us to consider the issue of age in dealing with decisions regarding both renal transplant recipients and donors. These geriatric issues in transplantation can be summarized as follows: (1) The explosion of a geriatric population of patients with ESRD, in association with data showing a survival advantage of transplantation over wait listed dialysis patients, demands an increase in expertise in transplantating patients over 60 years old. (2) The critical shortage in cadaveric organ supply is creating a variety of solutions including expanding the donor pool with older kidneys in which long term survival may be shorter than in kidneys from younger donors. (3) The donor shortage, in association with data demonstrating improved survival of living related and unrelated donor transplants, is generating an increased number of older (> 60 years old) individuals who want to donate to a relative, spouse or friend. Future efforts should be directed toward continued research designed to evaluate the efficacy and safety of these trends. We also need to provide improved training in geriatrics for nephrologists so that we and transplant surgeons can deliver better medical care to an aging population of patients with ESRD. PMID- 10518255 TI - The challenges of geriatric nephrology managed care/disease management. PMID- 10518256 TI - Nephrotic syndrome in the elderly. PMID- 10518257 TI - Reintegrating injury prevention. PMID- 10518258 TI - Injury prevention is growing up! PMID- 10518259 TI - An editor's dilemma--avoiding conflict of interest. PMID- 10518260 TI - Joining with the adults. PMID- 10518261 TI - Emergency department injury surveillance systems: the best use of limited resources? PMID- 10518262 TI - Unintentional childhood injury mortality in Europe 1984-93: a report from the EURORISC Working Group. AB - OBJECTIVE: To examine recent trends in unintentional childhood injury mortality in Europe, and to identify the contribution of specific causes. SETTING: The 15 current member countries of the European Union. METHODS: Analysis of mortality data (1984-93) obtained from the World Health Organisation and national government agencies. RESULTS: Injuries continue to be the leading cause of childhood death in all study countries, with more than 4500 fatalities annually, accounting for over 30% of all child mortality. The major causes of death in all countries were injuries due to motor vehicle traffic accidents, drownings, fire and flames, and falls. Portugal experienced mortality rates double those of most other countries, with the differentials particularly stark early in the study period. Although a decrease in age standardised mortality rates was observed in all countries over the decade, the extent of the decrease varied widely, from 47% in the UK to -11% in Finland. CONCLUSION: The pattern of childhood injury in Europe is similar to that observed elsewhere in the world. None the less, differences in rates of childhood injury mortality persist between countries. Identifying the reasons for these variations between countries may hold the key to the reduction injury rates in Europe as a whole. PMID- 10518263 TI - The "Let's Get Alarmed!" initiative: a smoke alarm giveaway programme. AB - OBJECTIVES: To reduce fires and fire related injuries by increasing the prevalence of functioning smoke alarms in high risk households. SETTING: The programme was delivered in an inner London area with above average material deprivation and below average smoke alarm ownership. The target population included low income and rental households and households with elderly persons or young children. METHODS: Forty wards, averaging 4000 households each, were randomised to intervention or control status. Free smoke alarms and fire safety information were distributed in intervention wards by community groups and workers as part of routine activities and by paid workers who visited target neighbourhoods. Recipients provided data on household age distribution and housing tenure. Programme costs were documented from a societal perspective. Data are being collected on smoke alarm ownership and function, and on fires and related injuries and their costs. RESULTS: Community and paid workers distributed 20,050 smoke alarms, potentially sufficient to increase smoke alarm ownership by 50% in intervention wards. Compared with the total study population, recipients included greater proportions of low income and rental households and households including children under 5 years or adults aged 65 and older. Total programme costs were 145,087 Pounds. CONCLUSIONS: It is possible to implement a large scale smoke alarm giveaway programme targeted to high risk households in a densely populated, multicultural, materially deprived community. The programme's effects on the prevalence of installed and functioning alarms and the incidence of fires and fire related injuries, and its cost effectiveness, are being evaluated as a randomized controlled trial. PMID- 10518264 TI - Urban and rural patterns of bicycle helmet use: factors predicting usage. AB - OBJECTIVES: To document current bicycle helmet use in Winnipeg, Manitoba and nearby rural communities, and to identify target groups for a helmet promotion campaign. METHODS: Cyclist helmet use was observed between 28 May and 20 August 1996 at a sample of urban and rural locations. Age, gender, helmet use, riding companion(s), location type, correct helmet use, and use of headphones were recorded. Univariate and multivariate analyses were performed. Adjusted odds ratios with 95% confidence intervals were calculated from the final models. RESULTS: Altogether 2629 cyclists (70% male, 30% female) were observed: 2316 at 183 urban locations and 313 at 25 rural locations, with nearly equal numbers of children and adults observed. Overall helmet use was 21.3%, with lower use in males (18.9%) than females (26.3%), despite gender only being a significant variable on multivariate analysis for children under 8 years and adults. Urban helmet use was considerably higher (22.9%) than rural use (8.9%). Helmet use increased linearly as mean neighbourhood income increased, with a nearly fourfold difference in use between the highest and lowest income neighbourhoods. Children less than 8 years old and adults had the highest, and teenagers the lowest, use. Significant predictive variables were identified separately by age category to inform targeted programming. CONCLUSIONS: We documented low helmet use in our region, emphasizing the need for a regional helmet promotion campaign as well as future helmet legislation. A marked urban-rural difference in helmet use that has not been previously reported was also identified. Target groups for a future campaign include adolescents, males, rural cyclists, and those in lower income neighbourhoods. PMID- 10518265 TI - Personal and family predictors of children's medically attended injuries that occurred in the home. AB - OBJECTIVE: This study examined the independent contributions of demographic, behavioral, and environmental antecedents of pediatric medically attended injuries that occurred in the home. SETTING: Two household and thirty six American children aged 4-12 in 1988 were drawn from the National Longitudinal Survey of Youth. METHOD: Multiple logistic regression was used to examine whether having a medically attended injury that occurred in the home in 1990 was related to environmental, behavioral, and demographic indicators measured in 1988. To account for individual differences in access to care, results were stratified within samples of children that had, and had not, demonstrated a prior ability to access the medical care system for injury treatment. RESULTS: Among children who did not access the medical care system for injury treatment in 1988, measures of home environmental risk factors did not distinguish those injured at home from those not injured at home in 1990. However, among children who did access the medical care system for injury treatment in 1988, indicators of "dark" (relative risk 4.68, p = 0.019) and "cluttered" (relative risk 4.31, p = 0.038) home environments became significantly and independently associated with home injuries in 1990. CONCLUSION: If not accounted for in data collection or analyses, individual differences in non-financial barriers to medical care may read to an underestimation of the influences of important home environmental risk factors for medically attended injuries. PMID- 10518266 TI - Fit of bicycle safety helmets and risk of head injuries in children. AB - BACKGROUND: Although bicycle helmets are effective in preventing head and brain injury, some helmeted individuals nevertheless sustain head injury. One of the possible reasons may be poor fit of the helmet on the head. This study was undertaken to examine the relationship between helmet fit and risk of injury. METHODS: 1718 individuals who were helmeted riders in a crash were queried on helmet fit and position. A sample of 28 children 2-14 years of age who sustained a head injury while wearing a bicycle helmet and 98 helmeted individuals of the same age treated in the same hospital emergency departments for injuries other than to the head, underwent anthropometric measurements of helmet fit. Measurements were made of the child's head, the helmet, and on a cast made of the child's head. RESULTS: Individuals whose helmets were reported to fit poorly had a 1.96-fold increased risk of head injury compared with those whose helmets fit well. Children with head injuries had helmets which were significantly wider than their heads compared with children without head injuries. Helmet fit was poorer among males and among younger children. CONCLUSIONS: Poor fit of helmets may be associated with an increased risk of head injury in children, especially in males. Helmets may not be designed to provide optimal protection. PMID- 10518267 TI - Descriptive study of sledding injuries in Canadian children. AB - OBJECTIVES: (1) To describe the characteristics of sledding injuries presenting to a pediatric emergency department and (2) To describe the sledding environment that leads to childhood sledding injuries. SETTING: A pediatric hospital emergency department in Ottawa, Canada and identified sledding sites in the region. METHODS: All patients less than 18 years with sled related injuries were included. Questionnaires were completed gathering information on the sled operator, the sled, the sledding site, and the injury. Site visits were made to designated and non-designated sledding hills in the Ottawa region to record data regarding sled operators, sleds, and the sledding environment. RESULTS: Ninety five patients were identified with sledding injuries and 81 (85%) completed the questionnaire. The mean age was 9.9 years (range 8 months to 17 years). The majority were male (63%). Most injuries occurred on non-designated sledding hills in the community (70%). Mild to moderate injuries were most common, however nine patients (11%) were admitted to hospital. Fifty-one per cent had adult supervision at the time of injury compared with 86% observed at the site visits. Common mechanisms of injuries were collisions with objects (33%), falls in icy conditions (28%), and going off jumps (16%). Most serious injuries occurred with contact with motor vehicles. There was no relationship between the type of sled used and the likelihood of injury. CONCLUSIONS: Sledding hills which have obstacles, icy conditions, jumps, or proximity to roads may result in more childhood injuries. Children with no adult supervision are likely at higher risk of injury. PMID- 10518268 TI - The spectrum of prevention: developing a comprehensive approach to injury prevention. AB - OBJECTIVE: The purpose of this paper is to describe the "spectrum of prevention", a framework for developing multifaceted approaches to injury prevention. The value of the tool is that it can help practitioners develop and structure comprehensive initiatives. METHODS: The spectrum is comprised of six inter related action levels: (1) strengthening individual knowledge and skills, (2) promoting community education, (3) educating providers, (4) fostering coalitions and networks, (5) changing organizational practices, and (6) influencing policy and legislation. Activities at each of these levels have the potential to support each other and promote overall community health and safety. CONCLUSIONS: The spectrum of prevention is a tool which can help practitioners and policy leaders move beyond a primarily educational approach to achieve broad community goals through injury prevention strategies that include policy development. This framework has been endorsed and applied in a variety of disciplines, however it has not been formally evaluated, a process that could clarify the scope of its effectiveness. PMID- 10518269 TI - CHIRPP: Canada's principal injury surveillance program. Canadian Hospitals Injury Reporting and Prevention Program. PMID- 10518270 TI - Sensitivity and representativeness of a childhood injury surveillance system. AB - OBJECTIVE: To determine the sensitivity and representativeness of the Canadian Hospitals Injury Reporting and Prevention Program (CHIRPP). SETTING: The study was conducted in the Ottawa-Carleton region of Ontario, Canada (June through August, 1992). METHODS: Surveillance system sensitivity was estimated by dividing the number of injured children attending the Children's Hospital of Eastern Ontario (the only CHIRPP center in Ottawa-Carleton) by the total number of emergency department attended childhood injuries in the region. CHIRPP representativeness was assessed by comparing the injuries missed by the system with those captured on social, demographic, and clinical factors. RESULTS: Sensitivity was 65% (1552/2386). Missed and captured injuries were similar on sex, day, time of presentation, injury intent, and delay before presentation. Children older than 14 years, however, were more likely to be missed by the system; adjusted odds ratio 3.52 (95% confidence interval (CI) 2.87 to 4.32). Conversely, children admitted to hospital were less likely to be missed; adjusted odds ratio 0.43 (95% CI 0.23 to 0.80). CONCLUSION: Given the systematic errors in capture, CHIRPP data should be used cautiously in studies of etiology. PMID- 10518271 TI - House fire injury prevention update. Part II. A review of the effectiveness of preventive interventions. AB - OBJECTIVE: To evaluate and summarize the house fire injury prevention literature. METHODS: MEDLINE (1983 to March 1997) was searched by keyword: fire, burn, etiology, cause, prevention, epidemiology, and smoke detector/alarm. ERIC (1966 to March 1997) and PSYCLIT (1974 to June 1997) were searched by keyword: as above, and safety, skills, education, and training. Other sources included references of retrieved publications, review articles, and books; Injury Prevention hand search; government documents; and internet sources. Sources relevant to residential fire injury prevention were selected, evaluated, and summarized. RESULTS: Forty three publications were selected for review, including seven randomized controlled trials, nine quasiexperiments, two natural experiments, 21 prospective cohort studies, two cross sectional surveys, one case report, and one program evaluation. These studies examined the following types of interventions: school (9), preschool (1), and community based educational programs (5); fire response training programs for children (7), blind adolescents (2), and mentally retarded adults (5) and children (1); office based counseling (4); home inspection programs (3); smoke detector giveaway campaigns (5); and smoke detector legislation (1). CONCLUSIONS: This review of house fire prevention interventions underscores the importance of program evaluation. There is a need for more rigorous evaluation of educational programs, particularly those targeted at schools. An evidence based, coordinated approach to house fire injury prevention is critical, given current financial constraints and the potential for program overload for communities and schools. PMID- 10518272 TI - The Safety City Program: knowledge is power. PMID- 10518273 TI - The changing approach to the epidemiology, prevention, and amelioration of trauma: the transition to approaches etiologically rather than descriptively based. 1968. PMID- 10518274 TI - Inequalities in health. PMID- 10518275 TI - Injury prevention in the Republic of Ireland. PMID- 10518276 TI - Transcobalamin II deficiency with methylmalonic aciduria in three sisters. AB - Transcobalamin II (TC II) is a plasma protein that binds vitamin B12 (cobalamin, Cbl) and facilitates cellular Cbl uptake by receptor-mediated endocytosis. In autosomal recessive TC II deficiency, intracellular Cbl deficiency results in an early onset of megaloblastic anaemia that may be accompanied by neurological abnormalities. Inadequate treatment may lead to neurological abnormalities. We describe three sisters, the daughters of first cousins of Moroccan origin, with TC II deficiency requiring continuous and long-term vitamin B12 treatment. The diagnosis was suspected from the finding of low unsaturated vitamin B12 binding capacity and confirmed by absence of detectable TC II by radioimmunoassay and by inability of cultured fibroblasts to synthesize TC II. PMID- 10518278 TI - Successful pregnancy in severe methylmalonic acidaemia. AB - Methylmalonic acidaemia is an inborn error of metabolism characterized by recurrent episodes of life-threatening ketoacidosis. With improved and intensive treatment, these patients are living into adulthood, but many experience late onset disease complications such as chronic renal failure, chronic pancreatitis and osteopenia. We report the successful delivery of a healthy baby to a 20-year old woman with vitamin B12-unresponsive methylmalonic acidaemia who has these late-onset manifestations of the disease and had plasma methylmalonic acid concentrations of 1900 mumol/L during the first trimester of pregnancy. PMID- 10518277 TI - Towards metabolic sink therapy for mut methylmalonic acidaemia: correction of methylmalonyl-CoA mutase deficiency in T lymphocytes from a mut methylmalonic acidaemia child by retroviral-mediated gene transfer. AB - The pathology associated with mut methylmalonic acidaemia (MMA) is caused by systemic accumulation of methylmalonate. Therefore, removal of methylmalonate from the circulation of affected individuals by an engineered metabolic system is proposed as a potential treatment. The haematopoietic cell is a potential site for such a metabolic system because of its direct contact with the accumulated metabolite and the demonstrated safety and ease in utilizing this cell. In this study, we assessed the feasibility of developing a haematopoietic cell-based methylmalonate sink by analysing propionate/methylmalonate metabolism in a variety of haematopoietic cells. The results show that propionate metabolism and methylmalonyl-CoA mutase (MCM) activity are intact in primary T cells, EBV-B cells, and CD34+ haematopoietic stem cell-derived granulocytes, whereas they are defective in those from a mut MMA child. Moreover, normal T and EBV-B cells clear methylmalonate from the medium at a significant rate. Transduction of MCM deficient T cells with a recombinant retrovirus encoding the human MCM cDNA results in correction of propionate metabolism. These results establish the basis for developing haematopoietic cell-based metabolic sink therapy for mut MMA by T lymphocyte/haematopoietic stem cell-directed gene transfer. PMID- 10518279 TI - Nutritional deficiencies in a patient with glycogen storage disease type Ib. AB - The current mainstay of treatment in glycogen storage disease type I (GSD I) is dietary management that includes providing a frequent source of glucose to prevent hypoglycaemia. To ensure compliance, routine follow-up by a health care team, including a dietitian, experienced in the treatment of GSD is necessary. We describe an adolescent patient with GSD Ib in good metabolic control who was admitted with a 3-month history of weakness, depression, vomiting, decreased appetite and a 11.4-kg weight loss. He had a recent onset of unsteady gait, inability to write, and sore mouth. After an extensive work-up, the patient was found to have vitamin B12, folate, iron and other nutritional deficiencies, which explained his symptoms. The patient improved within 72 h of initiation of total parenteral nutrition and therapeutic doses of deficient micronutrients, with a complete recovery in 2 months. Dietary restrictions, dependence on non-food products (e.g. cornstarch in GSD I), and social and developmental issues place individuals with metabolic disorders at a high risk for developing an array of nutritional deficiencies. This case highlights the importance of both close follow-up of the metabolic control and close monitoring of growth and nutritional intake in individuals with inborn errors of metabolism. This case also illustrates the importance of daily supplementation with appropriate multivitamins, calcium and other minerals needed to meet the Recommended Dietary Allowances (RDAs) in these patients. PMID- 10518280 TI - Adolescent myopathic presentation in two sisters with very long-chain acyl-CoA dehydrogenase deficiency. AB - Two sisters were investigated at the ages of 20 and 13 years owing to persistently increased serum creatine kinase and recurrent episodes of rhabdomyolysis after emotional stress in the older and myalgias in the younger. The finding of increased levels of cis-5-tetradecenoic acid (C14:1) in plasma, severe hypocarnitinaemia and the absence of a pathological dicarboxylic aciduria in both sisters suggested a very long-chain acyl-CoA dehydrogenase (VLCAD) deficiency. Reduced [1-(14)C]palmitate oxidation and deficient mitochondrial VLCAD activity in fibroblasts were found. Mutation analysis revealed compound heterozygosity for Asp365His and Arg410His changes. This late-onset, milder clinical presentation differs from the other two more severe infantile phenotypes described, since there is no hypoglycaemia or cardiac disease. Fatty acid oxidation defects should be investigated in all cases with rhabdomyolysis beginning in adolescence or early adulthood. PMID- 10518281 TI - Long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency: variable expressivity of maternal illness during pregnancy and unusual presentation with infantile cholestasis and hypocalcaemia. AB - Patients with long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD) deficiency present with a Reye-like syndrome, cardiomyopathy, or sudden unexpected death. We describe an unusual presentation in a patient with unsuspected LCHAD deficiency. The proband presented at 2 months of age with an acute infantile hypocalcaemia and vitamin D deficiency associated with occult, unexplained cholestatic liver disease. Sudden, unexpected death occurred at 8 months. Molecular analysis revealed homozygosity for the prevalent LCHAD (1528G > C, E474Q) mutation. The mother had pre-eclampsia during the third trimester of her pregnancy. In a subsequent pregnancy, she developed severe acute fatty liver of pregnancy (AFLP) and intrauterine fetal death at 33 weeks of gestation. In conclusion, infantile hypocalcaemia is an unusual phenotype associated with LCHAD deficiency. The maternal pregnancy history documents that fetal LCHAD deficiency is associated with a spectrum of maternal illnesses ranging from pre-eclampsia to life threatening AFLP. PMID- 10518282 TI - Urinary organic acid screening in children with developmental language delay. AB - The prevalence of 3-methylglutaconic aciduria was evaluated among children with developmental language disorders. A urine specimen was obtained from 40 children referred for developmental language delay to the Tel-Aviv Child Development Center during 12/96-6/97 and from 50 age-matched controls. Urine organic acids were analysed by gas chromatography-mass spectrometry. Urinary 3-methylglutaconic acid was quantified. A mildly increased excretion of 3-methylglutaconic acid was found in 8 children with developmental language delay. The combined excretion of 3-methylglutaconic and 3-methylglutaric acid was increased in 9 patients. There were no differences in the excretion of other organic acids. The patients with elevated 3-methylglutaconic acid did not differ from the other patients with developmental language disorders in any of the parameters evaluated. Mildly elevated urinary levels of 3-methylglutaconic acid may be a marker of a still undefined metabolic disorder presenting with developmental language delay. A further study in large groups of children with different developmental disorders is mandatory. PMID- 10518283 TI - Artefactual pyruvate and 2-oxobutyrate produced by trimethylsilylation of methylmalonic and ethylmalonic acids in the presence of oxygen. AB - Trimethylsilylation of methylmalonic and ethylmalonic acids in the presence of headspace atmospheric oxygen is shown to produce the trimethylsilyl derivatives of pyruvic and 2-oxobutyric acids, along with 2-hydroxy-2-methylmalonic and 2 hydroxy-2-ethylmalonic acids, respectively. This may lead to overestimation of these keto acids, if they were not oximated in the original sample, and the mistaken reporting of the 2-hydroxymalonates. PMID- 10518284 TI - Muscle carnitine acetyltransferase and carnitine deficiency in a case of mitochondrial encephalomyopathy. AB - Profound decrease of the carnitine acetyltransferase activity (0.08 U/g wet weight; 1.67% of control) and carnitine deficiency (total carnitine was 230 nmol/g wet weight in the patient vs 2730 in the controls) was detected in the skeletal muscle of a female paediatric patient. She died of her illness, which included cerebellar symptoms and slight muscle spasticity affecting mainly the lower extremities, at 1 year of age. Histological examination of the autopsy specimens revealed a selective Purkinje cell degeneration in the cerebellum: the cells had abnormal position, were shrunken and decreased in number, and displayed abnormal dendritic trees and fragmented, disorganized axons. Electron microscopy revealed mitochondrial abnormalities in skeletal and cardiac muscle and also in the Purkinje cells. Deletions of the mitochondrial DNA were detected in the muscle in heteroplasmic form (up to 7%). Mainly the ND4-ND4L region was affected, as evidenced by the PCR; however, other regions of the mitochondrial genome also showed deletions of varying size and extent, suggesting multiple deletions of the mitochondrial DNA. PMID- 10518285 TI - Long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency: neonatal manifestation at the first day of life presenting with tachypnoea. PMID- 10518286 TI - Neonatal diagnosis of long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency and implications for newborn screening by tandem mass spectrometry. PMID- 10518287 TI - False negative thiosulphate screening test in a case of molybdenum cofactor deficiency. PMID- 10518288 TI - A new polymorphism in the iduronate-2-sulphatase gene. Implications for the diagnosis of Hunter disease. PMID- 10518289 TI - Eighteen novel mutations in patients with Lesch-Nyhan syndrome or partial hypoxanthine phosphoribosyltransferase deficiency. PMID- 10518290 TI - Normal fluorine-18-labelled 2-fluoro-2-deoxyglucose positron emission tomography and magnetic resonance imaging of the brain in Wolman disease. PMID- 10518291 TI - Bone marrow transplantation in mucopolysaccharidosis type IIIA: a comparison of an early treated patient with his untreated sibling. PMID- 10518292 TI - Mutation analysis of the propionyl-CoA carboxylase alpha-subunit gene in four Japanese patients with propionic acidaemia. PMID- 10518293 TI - Prevalence of comorbid conditions, surgeries, and medication use in a physical therapy outpatient population: a multicentered study. AB - STUDY DESIGN: Prospective, multicenter, observational research study. OBJECTIVES: To investigate the prevalence of comorbidities and medical interventions in physical therapy outpatients and to establish a medical history profile of this population. BACKGROUND: Little research exists that describes the medical history of individuals seeking physical therapy services. The presence of certain comorbid conditions could influence the therapist's examination, evaluation, choice of interventions, treatment outcomes, and choice of outcome measures. METHODS AND MEASURES: Data were obtained from 2433 adults seeking care in 65 rehabilitation clinics located in 20 states. The clinics were located in major US geographic regions, and data were collected during each of the 4 seasons. A self administered questionnaire was used to collect data. The chi 2 test of independence was used for analysis of the association between the presence of disease and sex and geographic region. RESULTS: Forty-two percent of the potential subjects (N = 2433) completed the questionnaire. Skin cancer was reported in 4.5% (N = 110) of the sample. Hypertension (21%, N = 506), depression (15%, N = 354), chronic sinus infection (15%, N = 372), and pneumonia (11%, N = 276) were among the most frequently cited illnesses. The most frequently noted surgeries were orthopaedic procedures (27%, N = 653) and hysterectomy (15%, N = 369), and anti-inflammatories (40%, N = 984) and narcotics (28%, N = 671) were the most commonly prescribed medications. CONCLUSIONS: Many individuals seeking outpatient physical therapy services have extensive medical histories. Understanding the diseases, surgeries, and medications frequently encountered in practice is necessary for developing safe and appropriate interventions and establishing a reasonable prognosis. PMID- 10518294 TI - Physiological measurements of walking and running in people with transtibial amputations with 3 different prostheses. AB - STUDY DESIGN: A 3-factor (foot type, speed, and mode of ambulation) repeated measures experimental design was used. OBJECTIVES: To compare the differences in energy expenditure, gait efficiency, and relative exercise intensity in persons with transtibial amputations with various prostheses. BACKGROUND: There is a need for improved prosthetic designs to accommodate physically active persons with lower-extremity amputations. METHODS AND MEASURES: We used progressive speeds of treadmill walking (53.64, 67.05, 80.46, 93.87, and 107.28 m/min) and running (120.69, 134.1, and 147.51 m/min) with 3 different types of prostheses: the Solid Ankle Cushion Heel (SACH) foot, the Flex-Foot (FF), and the Re-Flex Vertical Shock Pylon (VSP) prosthesis. Five physically active men with unilateral transtibial amputations served as subjects (aged 31.6 +/- 4.28 years). RESULTS: The following statistically significant differences (improvements) between the Re Flex VSP versus the FF and the SACH foot were found. Energy cost: walking (5%), running (11%); gait efficiency: walking (6%), running (9%); relative exercise intensity: walking (4%), running (5%). However, we found no significant differences between the FF and the SACH. CONCLUSIONS: The Re-Flex VSP appears to have a positive effect on energy cost, efficiency, and relative exercise intensity compared with the other prosthetic foot types during walking and running. PMID- 10518295 TI - Diagnosis of mechanical low back pain in a laborer. PMID- 10518296 TI - Association of KT-1000 measurements with clinical tests of knee stability 1 year following anterior cruciate ligament reconstruction. AB - STUDY DESIGN: Prospective, observational study. OBJECTIVES: To determine the association between KT-1000 measurements with an anterior translation force of 89 N and other measures of outcome (the Tegner activity score, the modified Lysholm score, subjective rating of instability, Lachman test, and pivot-shift test) 1 year following anterior cruciate ligament (ACL) reconstruction. BACKGROUND: Health care professionals often use the side-to-side difference measured with the KT-1000 arthrometer to determine ACL integrity during passive motion. It has been postulated that a 5-mm or greater difference between impaired and nonimpaired knees represents a procedural failure. METHODS AND MEASURES: Ninety patients (46 men, 44 women) with a mean age of 30 +/- 8 years were examined 1 year after surgery. Patients were classified in 1 of 3 groups depending on the amount of laxity between the impaired knee and the nonimpaired knee. Seventy percent of the subjects had a side-to-side difference less than or equal to 3 mm (tight), 13% had a difference of between 3 and 5 mm (moderate), and 17% had a difference greater than or equal to 5 mm (loose) on examination using the KT-1000. RESULTS: Mean Lysholm and Tegner scores did not differ significantly among groups. Side-to side differences in KT-1000 measurements at 89 N were not associated with the Lysholm score (r = -0.09) or Tegner score (r = 0.02). Lachman tests were related to involved-knee KT-1000 measurements (r = 0.39) but not to side-to-side differences in KT-1000 measurements (r = 0.15). Similarly, pivot-shift tests were related to involved-knee KT-1000 measurements (r = 0.26) but not to side-to-side differences (r = -0.08). CONCLUSIONS: These results suggest that side-to-side KT 1000 measurements obtained with an anterior translation force of 89 N should not be used in isolation to determine ACL reconstruction success or failure 1 year following surgery. PMID- 10518297 TI - Effects of practice on the ability to perform lumbar stabilization exercises. AB - STUDY DESIGN: Randomized pretest-posttest control group design. OBJECTIVES: To determine the intratester and intertester reliability of a modified isometric stability test and to use this test to evaluate the effects of practice following a 4-week stabilization exercise program with weekly reinstruction. BACKGROUND: Although "stabilization" exercise programs are commonplace in the clinic, the reliability to a tool capable of measuring changes in the ability to perform increasingly difficult stabilization exercises has not, to our knowledge, been reported. In addition, it is not clear if practice improves the ability to perform stabilization exercises. METHODS AND MEASURES: A convenience sample of 44 asymptomatic subjects was pretested using a pressure transducer placed beneath the lumbar spine to detect motion (+/- 4 mm Hg). A series of 7 exercises was attempted, which required increasing levels of muscular control of the lumbar spine for stability. Subjects received a pass or fail for each exercise level based on the pressure gauge readings and the absence of movement compensations. Subjects were assigned randomly to exercise and nonexercise groups, and posttest measurements were taken after 4 weeks. The control group did not receive additional instruction. RESULTS: The weighted kappa coefficient of 0.61 for intratester and 0.62 for intertester represents good agreement. The median level of exercise attainment increased for the exercise group but not for the nonexercise group. CONCLUSION: These results suggest that the modified isometric stability test was reliable and that a 4-week lumbar stabilization exercise program, with weekly intervals of reinstruction and testing, improves the ability to perform progressively difficult lumbar stabilization exercises. PMID- 10518298 TI - Classifying sacroiliac dysfunction. PMID- 10518299 TI - Mixed-function supraoperons that exhibit overall conservation, albeit shuffled gene organization, across wide intergenomic distances within eubacteria. AB - Nearly identical mixed-function supraoperons (defined as nested transcriptional units encoding gene products that function in more than one biochemical pathway) have been found recently in Pseudomonas stutzeri and Pseudomonas aeruginosa. The Pseudomonas serC(pdxF)-aroQp.pheA-hisHb-tyrAc-aroF+ ++-cmk-rpsA supraoperon encodes 3-phosphoserine aminotransferase, a bidomain chorismate mutase/prephenate dehydratase, imidazole acetol-phosphate aminotransferase, cyclohexadienyl dehydrogenase, 5-enolpyruvylshikimate 3-phosphate synthase, cytidylate kinase, and 30S ribosomal protein S1. These enzymes participate in the biosynthesis of serine, pyridoxine, histidine, phenylalanine, tyrosine, tryptophan, and aromatic pathway vitamins and cytidylic acid, in addition to the general role of RpsA in the process of protein synthesis. Features that suggest supraoperon-wide translational coupling are the highly compressed intergenic spacing (including overlapping stop and start codons), as well as possible hairpin structures in mRNA, which could sequester many of the ribosome-binding sites. The hisH-tyrA aroF segment corresponds to the distal genes of the classic Bacillus subtilis supraoperon. Extensive comparative analysis of the member genes of both the Bacillus and Pseudomonas supraoperons from organisms represented in the entire database revealed unmistakable organizational conservation of these genes across wide phylogenetic boundaries, although considerable gene shuffling was apparent. The persistence of aroE-aroB, hisHb-tyrA-aroF, and cmk-rpsA throughout both the gram-negative and gram-positive assemblages of bacteria, but the absence in Archaea, suggests an ancestral gene organization that occurred in bacteria after the separation of the bacterial and archaeal domains. In gram-negative bacteria,the hisHb-tyrAc-aroF grouping may have been expanded (as with the Pseudomonas supraoperon) and then subsequently collapsed (as with the Escherichia serC-aroF supraoperon) via gene shuffling that is herein equated with gene fusion events. PMID- 10518301 TI - Molecular cloning of the dnaK gene region from Bacillus sphaericus in the context of genomic comparisons. AB - The dnaK gene region of Bacillus sphaericus was cloned as a 3.8 kb HindIII fragment and an overlapping 1.7 kb EcoRI fragment by using an internal B. sphaericus specific dnaK gene probe generated by polymerase chain reaction (PCR). Complete DNA sequencing of the two fragments revealed three complete open reading frames (ORFs). These ORFs exhibited a high degree of identity to the grpE dnaK, and dnaJ heat shock genes from other gram-positive bacteria. The order of the genes was found to be grpE-dnaK-dnaJ. Additionally, the 5'-end and 3'-end contained amino acid sequences that were homologous to the C-terminal sequence of the hrcA gene and the N-terminal sequence of ORF35 (yqeT), respectively, from Bacillus subtilis. The entire hrcA gene from B. sphaericus was then isolated by high-fidelity PCR and completely sequenced. A transcription stop site is located between the dnaK and dnaJ genes but not after the dnaJ gene. Consistent with this observation, the dnaJ gene is immediately followed by an ORF that shows a high degree of identity to ORF35 from B. subtilis, Staphylococcus aureus, and Clostridium acetobutylicum. The presence of ORF35 is not indicated in other genera representing the gram-positive bacteria. The amino acid sequence of ORF35 exhibited nearly 30% identity with the methyltransferase for large subunit ribosomal protein L11 from gram-negative Proteobacteria and the related protein from cyanobacteria, other gram-negative bacteria, and Archaea, suggesting the presence of the gene for this protein in the common ancestor of Bacteria and Archaea. The absence of the ORF35 gene in Mycobacterium tuberculosis and other gram-positive bacteria indicates that the loss of this gene must have occurred in an ancestor of other gram-positive bacteria following their divergence from the ancestor of Bacillus/Clostridium/staphylococcus lineage. PMID- 10518300 TI - Mechanisms of spiroplasma genome variation associated with SpV1-like viral DNA inferred from sequence comparisons. AB - Genomes of Spiroplasma citri strains have rearranged frequently during their evolution, partly due to multiple integrated sequences of spiroplasma viruses. To understand better the role of viral sequences in genome evolution, we examined available nucleotide sequences of viruslike elements in the S. citri chromosome. Comparison of integrated and nonintegrated sequences of spiroplasma virus SpV1 C74 DNA suggested that it is an encapsidated form of the circular transposition intermediate belonging to an insertion sequence (IS3) family member. One SpV1-C74 viral DNA fragment was identified as interrupting the remains of a DNA adenine modification methylase gene. A viral DNA insertion of SpV1-R8A2 B DNA had hallmarks of having suffered an internal deletion by a site-specific recombination system. Homologous recombination likely was responsible for several deletions within viral DNA. A homologous recombination event was inferred between part of a viral DNA insertion and a similar chromosomal sequence. Dispersed sequences from SpV1-like C4 open reading frames (ORFs) were identified as involved in a complex deletion-inversion event. Thus, SpV1-like sequences likely have altered spiroplasma genomes by inserting within active genes, destroying their function, by providing targets for site-specific recombination, by mediating deletions of sequences adjacent to their integration sites, and by providing targets for homologous recombination, leading to inversions. PMID- 10518302 TI - Identifying potential conflicts of interest commercially sponsored research: establishing a policy for acceptable standards of disclosure. PMID- 10518303 TI - Genetic aspects of hepatocellular carcinogenesis. AB - Hepatocellular carcinoma (HCC) is linked etiologically to viruses (hepatitis B virus [HBV] and hepatitis C virus [HCV]), chemical carcinogens (i.e., aflatoxins), and other environmental and host factors causing chronic liver injury. Some hepatoblastomas may be linked to inherited gene mutations, but adult hereditary HCC appears to be rare. HCCs display gross genomic alterations, including DNA rearrangements associated with HBV DNA integration, loss of heterozygosity, and, less importantly, chromosomal amplifications and loss of imprinting. Many genes with somatic mutations have now been identified in these tumors. Most frequently involved genes are tumor suppressor genes such as p53, M6P/IGF2R, beta-catenin, p16INK4A, and retinoblastoma genes. Most identified mutations are somatic, but germline mutations of p16INK4A, APC, and BRCA2 have also been reported. Oncogenic activation of several cellular genes such as cyclin D and cyclin A have been described in HCC, but the possible implication of candidate viral oncogenes (i.e., X protein of HBV) is still debated. A comprehensive analysis of all the genetic changes described for HCC demonstrates that at least four different growth regulatory pathways are altered in these tumors. However, each pathway appears to be implicated in a limited fraction of these tumors, suggesting that HCCs are genetically heterogenous neoplasms. This genetic heterogeneity correlates with the heterogeneity of etiologic factors implicated in HCC. PMID- 10518304 TI - Mouse liver tumorigenesis: models, mechanisms, and relevance to human disease. AB - Hepatocytes have a remarkable proliferative capacity, but are quiescent in normal liver. Cell cycle activation in hepatocarcinogenesis can be directly triggered by overexpression of single and combinations of genes or be initiated indirectly by compensatory proliferation in response to liver injury. Work with transgenic and knockout mice indicate that regardless of the initiating cause, constitutive hepatocyte proliferation accompanied by genomic damage are essential factors for liver tumor development. The carcinogenic process is best described as a continuum that involves unregulated hyperplasia, dysplasia, and adenoma formation. The critical steps required for the transition from regulated to constitutive hepatocyte proliferation and the mechanisms of genomic damage in proliferating cells are being investigated. This knowledge should be directly applicable to studies of human liver tumorigenesis. PMID- 10518305 TI - Hepatitis B virus infection and hepatocellular carcinoma: molecular genetics and clinical perspectives. AB - Chronic hepatitis B progresses across a spectrum of asymptomatic carriers, active hepatitis, and liver cirrhosis. With more advanced disease stage, the risk for developing hepatocellular carcinoma (HCC) becomes higher. Recent studies suggest that this progressive risk may reflect an accumulation of multistage genetic mutations in the chromosomes of affected hepatocytes. Mutations of the known candidate genes such as p53 and beta-catenin have been found. Recent genome-wide analysis of HCC chromosomes by comparative genomic hybridization or loss of heterozygosity have identified more new loci implicated in hepatocarcinogenesis. Persistent hepatitis B is essential for inducing these mutations through immune mediated injuries of the hepatocytes and the resulting hyperplasia. Prevention of hepatitis B by active immunization effectively interrupts persistent viral infections in children and subsequently reduces the risk of childhood HCC. Treatment for chronic hepatitis B by interferon or antiviral analogues can control hepatitis B activity, but its effect on controlling HCC remains to be seen. Insights for the hepatocarcinogenesis process should come from a multidisciplinary collaboration to explore important viral and host genes so that new approaches to diagnosis and treatment can be developed. PMID- 10518306 TI - Hepatitis C virus and hepatocellular carcinoma. AB - Hepatitis C virus (HCV) is pathogenetically involved in many cases of hepatocellular carcinoma (HCC) worldwide. HCV-related HCC is on the rise in many developed countries as a consequence of past infections with HCV. The time lag between HCV infection and cancer development is several decades. HCV-related tumors arise in older patients, are almost invariably associated with cirrhosis, and often have a less aggressive course than HCC related to other etiologic factors. In most patients, HCC grows as a single hepatic node for years before generating satellite or distant tumor nodes. However, there are tumors that originate as multifocal disease. Tumor progression and hepatic failure are the leading causes of death in most patients. HCV has been almost invariably detected in tumor tissue of anti-HCV patients with HCV, but it is not clear whether the virus promotes cancer through chronic hepatocellular inflammation, which is per se an important risk factor for HCC, or has a direct role in liver carcinogenesis. No reverse transcriptase activity has been found in infected livers, but there are data suggesting that HCV has oncogenic properties, because its interacts with cellular genes regulating cell growth and differentiation. PMID- 10518307 TI - Epidemiology of primary liver cancer. AB - Liver cancer (LC) ranks fifth in frequency in the world with an estimated number of 437,000 new cases in 1990. In developing countries, incidence rates are two- to three-fold higher than in developed countries. The geographic areas at highest risk are located in Eastern Asia, with age-adjusted incidence rates (AAIRs) ranking from 27.6 to 36.6 per 100,000 in men; Middle Africa, with AAIRs ranking from 20.8 to 38.1 per 100,000 in men; and some countries of Western Africa, with AAIRs ranking from 30 to 48 per 100,000 in men. The geographic areas at lowest LC risk are Northern Europe, Australia, New Zealand, and the Caucasian populations in North and Latin America, with AAIRs below 5.0 per 100,000 in men. Excess of LC incidence among men compared to women is universal, with sex ratios between 1.5 and 3.0. Significant variations in LC incidence among different ethnic groups living in the same geographical area and among migrants of the same ethnic groups living in different areas have been extensively described. The variability of LC incidence rates between countries and within countries, strongly suggests differences in exposure to risk factors. The role of chronic infection with the Hepatitis B and hepatitis C viruses (HBV and HCV) in the etiology of LC is well established. The attributable risk estimates for LC for each of these hepatotropic viruses vary among countries but the combined effects of persistent HBV or HCV infections account for well over 80% of LC cases worldwide. Other documented risk factors such as aflatoxin exposure in diets, cigarette smoking, alcohol consumption, and oral contraceptives may explain the residual variation between and within countries. Interactions between some risk factors have been postulated, and are subject of active research. New laboratory techniques and biological markers such as polymerase chain reaction detection of HBV DNA and HCV RNA, as well as specific mutations related to aflatoxin exposure may help to provide quantitative estimates of the risk related to each these factors. PMID- 10518308 TI - Histopathologic evaluation of hepatocellular carcinoma with special reference to small early stage tumors. AB - Over the past decade extensive studies on small early stage hepatocellular carcinomas (HCCs) have defined their pathomorphologic features. Most early HCCs are well differentiated, with an ill-defined nodular appearance. Proliferation of well-differentiated small HCCs is closely related to tumor dedifferentiation. When a well-differentiated HCC reaches a size of about 1.0-1.5 cm in diameter, less-differentiated cancerous tissues with greater proliferative activity evolve within it. Such a phenomenon is often appreciated grossly and/or histologically as a "nodule-in-nodule" appearance. Subsequently, moderately to poorly differentiated HCC tissues gradually replace the initial surrounding HCC. This replacement of well-differentiated HCC tissue is completed when the tumor reaches a size of about 2-3 cm. Hyperplastic nodular lesions in cirrhotic livers may have a premalignant potency. HCC frequently occurs multicentrically whether synchronously or metachronously, defying complete cure by conventional therapies other than liver transplantation. PMID- 10518309 TI - Imaging diagnosis of hepatocellular carcinoma and premalignant/borderline lesions. AB - With the recent progress in imaging modalities, detection of small, nodular hepatic lesions has become much easier; however, differential diagnosis still remains difficult. Nevertheless, a combination of tomographic imaging techniques and angiography, such as ultrasound (US) angiography, CT arteriography (CTA), and CT during arterial portography (CTAP) has contributed considerably to the differentiation of overt (advanced) hepatocellular carcinoma (HCC) from benign or premalignant/borderline lesions. With such techniques, estimation of the grade of malignancy is also possible. Recently, noninvasive ultrasonographic vascular imaging techniques have been developed, such as color Doppler, power Doppler, and enhanced Doppler imaging. In particular, gray-scale contrast second harmonic imaging may prove useful in the management of HCC and will replace some of the roles of magnetic resonance imaging (MRI) and CT in the near future; ultrasonographic visualization of the vascularity of viable cancer cells is essential for the US-guided interventional therapy of HCC. PMID- 10518310 TI - Liver transplantation for hepatocellular carcinoma. AB - Liver transplantation for hepatocellular carcinoma (HCC) in patients with cirrhosis is a radical treatment of the tumor and associated precancerous state. It is potentially curative in a proportion of patients. The outcomes of early studies of liver transplantation in this indication were initially unfavorable. Selection of transplant candidates at an early stage, in the absence of extrahepatic spread, gives better survival than surgical resection and alternative nonsurgical treatments. Transarterial chemoembolization can be used for preoperative control of the disease. Adjuvant chemotherapy may be indicated in the postoperative period for the prevention of recurrence in patients with histologic features of invasiveness in the surgical specimen. Liver transplantation as the treatment of choice for early HCC in screening programs in cirrhotic patients may become limited by graft availability as the numbers of hepatitis C-related cases increase. Resection may be indicated if the waiting time is likely to be long. PMID- 10518311 TI - Local ablation therapy for hepatocellular carcinoma. AB - Several local ablation methods, most of which can be performed percutaneously under imaging guidance, have recently been developed as minimally invasive therapy for hepatocellular carcinoma (HCC). Among these, percutaneous ethanol injection has been the most widely performed and is now accepted as an attractive alternative to surgery in patients with small HCC. Other local ablation therapeutic options to achieve more effective tumor necrosis with less invasiveness have also been developed. Some of these techniques, such as microwave coagulation therapy and radiofrequency ablation, have already been introduced clinically. However, the real role of these new ablation methods in the treatment of HCC remains to be determined because of the lack of randomization and the small number of patients in the published series. Therefore, further trials, possibly randomized trials to compare the results of different options, are mandatory. Moreover, the prevention of recurrence after local ablation therapy for HCC has become an urgent problem to be resolved. PMID- 10518312 TI - Prognosis of hepatocellular carcinoma: the BCLC staging classification. AB - The classifications of hepatocellular carcinoma (HCC) currently used are based on prognostic factors obtained from studies performed years ago when most tumors were diagnosed at advanced stages and the survival rates were substantially poor. Recent investigations have reviewed the survival of early tumors properly selected to receive radical therapies and the natural outcome of nonsurgical HCC patients. These data enable a new staging system to be proposed, the Barcelona Clinic Liver Cancer (BCLC) staging classification, that comprises four stages that select the best candidates for the best therapies currently available. Early stage (A) includes patients with asymptomatic early tumors suitable for radical therapies--resection, transplantation or percutaneous treatments. Intermediate stage (B) comprises patients with asymptomatic multinodular HCC. Advanced stage (C) includes patients with symptomatic tumors and/or an invasive tumoral pattern (vascular invasion/extrahepatic spread). Stage B and C patients may receive palliative treatments/new agents in the setting of phase II investigations or randomized controlled trials. End-stage disease (D) contain patients with extremely grim prognosis (Okuda stage III or PST 3-4) that should merely receive symptomatic treatment. PMID- 10518313 TI - A 42-year-old woman with liver masses and long-term use of oral contraceptives. AB - A 42-year-old woman with a history of 25-year oral contraceptive use presented with abdominal pain and was found to have two exophytic liver masses. She had no known prior liver diseases, and her serum liver enzyme and AFP levels were normal. One of the masses was a hepatocellular adenoma and the other was a pigmented hepatocellular carcinoma. The exophytic appearance of both lesions was unusual. This case, once more, demonstrated the risk of hepatocellular adenomas to undergo malignant transformation. The reason for the brown pigment deposition in the hepatocellular carcinoma was not clear. The prognosis was expected to be excellent following complete surgical resection. PMID- 10518314 TI - A review on in vitro studies of hemodynamic characteristics in terminal and lateral aneurysm models. AB - Intracranial saccular aneurysms have been a well known cerebrovascular disease for over fourty years in the Taiwan area and over two centuries around the world. Up to now, information pertinent to the genesis, progression, and thrombosis or rupture of saccular aneurysms has been mainly acquired from autopsies and various in vivo studies. The present review provides relevant hemodynamic information gathered from in vitro studies. The parallel results between in vitro and in vivo or between in vitro and autopsy investigations are also addressed. Emphases are placed on the terminal and lateral saccular aneurysms. The effects of the branches's flow-rate ratio, bifurcation angle, aneurysmal size, parent vessel curvature, and Wormersley number on the intra-aneurysmal flow characteristics are examined in detail, and possible risky factors are identified. PMID- 10518315 TI - Hemodynamic and neurohumoral changes after abdominal aortic constriction in rats. AB - Cardiac after-load, neurohumoral reaction and the secondary cardiac hypertrophy were studied in six groups of Sprague-Dawley (SD) rats with abdominal aortic constriction. We found that abdominal aortic constriction above the renal arteries decreased the heart rate and cardiac output, and increased the pulse pressure. These abnormalities would return to normal after constriction ended. Captopril, propranolol and prazosin could reduce the increase of pulse pressure but still had decreased in cardiac output of rats with abdominal constriction. Aortic constriction also increased the aortic impedance and cardiac load but decreased aortic compliance. These changes could also be lessened by captopril, propranolol and prazosin. We have confirmed that aortic constriction can induce secondary cardiac hypertrophy, but the pathogenesis might be due to multiple factors. PMID- 10518316 TI - Biological responses of dog prostate and adjacent structures after meso-tetra-(m hydroxyphenyl) chlorin and aluminum disulfonated phthalocyanine based photodynamic therapy. AB - Further to our work on the feasibility of application of photodynamic therapy (PDT) to the canine prostate, this study evaluates the biological responses of the prostate and adjacent vital structures with meso-tetra-(m-hydroxylphenyl) chlorin (mTHPC) or aluminum disulfonated phthalocyanine (AlS2Pc) based PDT as a preparatory step for clinical trials. Skin photosensitivity was not particularly problematic if light protection could be implemented properly for 2 weeks following sensitization. Prostate PDT was well tolerated by the experimental animals with only minor physical distress. mTHPC was more powerful than AlS2Pc in terms of prostate lesions induced. A large portion of prostate tissue could be destroyed by PDT with 4 punctures. Physical distress was probably caused by severe urethral irritation and aching from acute swelling of the prostate. Although the voiding condition normalized within 10 days, regeneration of urethral epithelium was not complete until 3-4 weeks after PDT. Improper placement of laser fiber caused extensive ecchymosis of the retroperitoneal organs. The biological significance of PDT induced hyperemia in the periprostatic structures remains poorly defined. Neither periprostatic nerve damage nor rectal lesions were seen in dogs receiving either mTHPC or AlS2Pc. Glandular atrophy with papillary cystic regeneration of the prostate was the most prominent finding 90 days after PDT. The glandular architecture was well preserved because the interlobular collagens were less affected than the cellular components of the glands. Our study suggests that PDT with mTHPC and AlS2Pc is safe and promising for necrosing a substantial amount of prostate tissue. The completeness of treatment and long-term therapeutic effectiveness for prostate cancer, however, remains to be determined through further investigation. PMID- 10518317 TI - Minimal residues of porcine reproductive and respiratory syndrome virus in pig carcases and boar semen. AB - Porcine reproductive and respiratory syndrome (PRRS) is a highly contagious disease that affects pigs of all ages worldwide. One of the key features of PRRS pathogenesis is the prolonged viremia in which the virus coexists with antibody. Prolonged viremia raises the concern that porcine products and boar semen may be contaminated by residues of the PRRS virus serving as vehicles for spreading of the virus. To answer this question, we sampled blood, muscles and viscera organs from market pigs slaughtered using an on-the-rail system and utilized a direct reverse transcription-polymerase chain reaction (RT-PCR) to detect residual viral RNA. We found that the overall seropositive rate of the market pigs was 85.4% (205/240), yet only 7.9% (11/140) of the blood samples contained detectable amounts of virus residues, and all tested carcase specimens (n = 472) were negative. The clinical sensitivity of this PCR varied with the tissues tested, with 5 TCID50 per 50 microliters of serum, as determined by means of a spiking experiment. Semen samples (n = 38) of clinically healthy seropositive and seronegative boars were collected from six herds which were routinely subjected to artificial insemination for production purposes. None of the seminal plasma or sperm-rich fractions contained PCR detectable virus residues. However, the possibility of PRRS virus present in semen cannot be totally excluded. We conclude that in naturally infected albeit clinically healthy pigs, the amounts of PRRS virus residues in carcases or semen are minimal. Thus, the risk of these products causing animal health hazards is low. PMID- 10518318 TI - Arginine methylation of a glycine and arginine rich peptide derived from sequences of human FMRP and fibrillarin. AB - N-methylation at the arginine residues in RNA binding proteins with the arginine and glycine rich RGG box has been identified. We show that a synthetic peptide R9 (GGRGRGGGF) with the RGG sequence present in human fibrillarin and fragile X mental retardation protein (FMRP) can be specifically methylated by rat brain extract. A control peptide K9 with all arginines replaced by lysines could not be methylated under the same conditions, indicating that the arginines in the peptide were the methylation sites. A novel missense mutation, which changes an arginine to a histidine in the RGG box region of FMRP in a typical fragile X patient, has been identified. A synthetic peptide with this Arg-->His (GGRGHGGGF) substitution was methylated by our in vitro methylation system to a much less extent. Amino acid analysis of the methylated R9 peptide identified the methylated amino acid as monomethylarginine. The R9 peptide may be useful for further studies on arginine methylation in RGG proteins. PMID- 10518319 TI - Male dispersal in the noctule bat (Nyctalus noctula): where are the limits? AB - Studying the dispersal behaviour of small, volant, and nocturnal animals such as microchiropterans with direct methods (banding--recapture, telemetry) is a very difficult task. The development of easily scorable and highly variable genetic markers nowadays allows us to study some aspects of dispersal indirectly, using population genetics. Here, we applied these indirect methods to characterize male dispersal behaviour in a European bat species. The eight microsatellite loci analysed were highly variable in all the nursing colonies assessed (h = 0.63 0.93). Contrary to what we found in the mtDNA, an AMOVA and F-statistics showed that the overall European population structure of the noctule bat was very weak, indicating a high male dispersal rate. Nevertheless, the population was not totally panmictic (theta = 0.006, p < 0.001), and neither isolation by distance, nor influence of migration could account for this result. Rather, an analysis of pairwise theta-values showed that the population structure might be explained partly by a geographical barrier to gene flow (the Alps), and partly by the fact that there is some limit to the distance the males can disperse. PMID- 10518320 TI - Variable investment, the Continuous Prisoner's Dilemma, and the origin of cooperation. AB - Cooperation is fundamental to many biological systems. A common metaphor for studying the evolution of cooperation is the Prisoner's Dilemma, a game with two strategies: cooperate or defect. However, cooperation is rare all or nothing, and its evolution probably involves the gradual extension of initially modest degrees of assistance. The inability of the Prisoner's Dilemma to capture this basic aspect limits its use for understanding the evolutionary origins of cooperation. Here we consider a framework for cooperation based on the concept of investment: an act which is costly, but which benefits other individuals, where the cost and benefit depend on the level of investment made. In the resulting Continuous Prisoner's Dilemma the essential problem of cooperation remains: in the absence of any additional structure non-zero levels of investment cannot evolve. However, if investments are considered in a spatially structured context, selfish individuals who make arbitrarily low investments can be invaded by higher investing mutants. This results in the mean level of investment evolving to significant levels, where it is maintained indefinitely. This approach provides a natural solution to the fundamental problem of how cooperation gradually increases from a non-cooperative state. PMID- 10518321 TI - The effect of Plasmodium yoelii nigeriensis infection on the feeding persistence of Anopheles stephensi Liston throughout the sporogonic cycle. AB - Vector-borne parasites such as malaria have been shown to modify the feeding behaviour of their invertebrate hosts so as to increase the probability of transmission. However, evolutionary consideration of developmental changes in malaria within Anopheles mosquitoes suggests that the nature of altered feeding by mosquitoes should differ depending on the developmental stage of the parasite. We present laboratory evidence that the feeding persistence of female Anopheles stephensi towards a human host is decreased in the presence of Plasmodium yoelii nigeriensis oocysts (which cannot be transmitted), but increased when the malaria has developed into transmissible sporozoites in the salivary glands. In ten minute trials, 33% of uninfected mosquitoes gave up their feeding attempt before the test period had ended, 53% of those harbouring oocysts had given up, but only 20% of those infected with sporozoites gave up by this time. We conclude that changes in feeding behaviour of mosquitoes mediated by parasite infection are sensitive to the developmental stage of the parasite and that these changes have important implications for malaria epidemiology. PMID- 10518322 TI - Growth, developmental stability and immune response in juvenile Japanese quails (Coturnix coturnix japonica). AB - Stresses are environmental factors which restrict growth or cause a potentially adverse change in an organism. The exposure of developing organisms to environmental stresses may have several physiological consequences including a decrease in immunocompetence. However, mounting an immune response against a foreign antigen may in itself constitute a cost for developing organisms. This cost has potentially long-term consequences for adult function and fitness. This study examines the growth and developmental stability of Japanese quail++ chicks challenged by three non-pathogenic antigens: sheep red blood cells, which assess T-cell-dependent immune responses, and Mycoplasma synoviae and Newcastle disease virus, which assess T-cell-independent responses. Increases in both body mass and wing length were significantly reduced in antigen-challenged birds compared to control birds. Fluctuating asymmetry (FA) in the masses of primary feathers increased from the innermost (1) to the outermost (10) position on the wing. In addition, antigen challenge by M. synoviae and sheep red blood cells was associated with an increase in FA. The cell-mediated response measured by reaction to phytohaemagglutinin was significantly depressed in M. synoviae challenged birds. White blood cell counts, except for monocytes, were elevated in response to all three antigen treatments. Total plasma protein and haematocrit also differed between treatments but exhibited no clear relationship to antigen challenge. Immune responses clearly impose a stress on developing chicks. Additional research will be required to determine the long-term consequences of developmental stress and assess the selective forces that influence the strength of the immune responses of chicks. PMID- 10518323 TI - Predicting variability in biological control of a plant-pathogen system using stochastic models. AB - A stochastic model for the dynamics of a plant-pathogen interaction is developed and fitted to observations of the fungal pathogen Rhizoctonia solani (Kuhn) in radish (Raphanus sativus L.), in both the presence and absence of the antagonistic fungus Trichoderma viride (Pers ex Gray). The model incorporates parameters for primary and secondary infection mechanisms and for characterizing the time-varying susceptibility of the host population. A parameter likelihood is developed and used to fit the model to data from microcosm experiments. It is shown that the stochastic model accounts well for observed variability both within and between treatments. Moreover, it enables us to describe the time evolution of the probability distribution for the variability among replicate epidemics in terms of the underlying epidemiological parameters for primary and secondary infection and decay in susceptibility. Consideration of profile likelihoods for each parameter provides strong evidence that T. viride mainly affects primary infection. By using the stochastic model to study the dependence of the probability distribution of disease levels on the primary infection rate we are therefore able to predict the effectiveness of a widely used biological control agent. PMID- 10518324 TI - Effects of morphine on electrically evoked contractions of the vas deferens in two congeneric rodent species differing in sperm competition intensity. AB - An early prediction of sperm competition theory was that males should adjust the number of sperm they deliver according to the risk of double mating and this has received empirical support in recent years. It has been suggested that adaptive regulation of sperm delivery in mammals may depend on changes in vas deferens contractility. In laboratory mice, the vas deferens is sensitive to opioid agonists and the secretion of endogenous opioid peptides can be affected by social interactions that may be predictive of sperm competition risk. The present experiment was conducted to determine whether morphine, an opioid agonist (at the mu-receptor), has different effects on electrically evoked contractions of the isolated vas deferens in two congeneric rodent species differing in sperm competition intensity. Morphine inhibited contractions of the vas deferens in the non-monogamous deer mouse (Peromyscus maniculatus) but not the monogamous California mouse (Peromyscus californicus). This implies that the vas deferens of P. maniculatus possesses functional mu-receptors and, thus, should be able to respond to changes in the circulating levels of endogenous agonists whose secretion can be affected by social interactions predictive of sperm competition risk. PMID- 10518325 TI - Dietary change and stable isotopes: a model of growth and dormancy in cave bears. AB - In order to discuss dietary change over time by the use of stable isotopes, it is necessary to sort out the underlying processes in isotopic variation. Together with the dietary signal other processes have been investigated, namely metabolic processes, collagen turnover and physical growth. However, growth and collagen turnover time have so far been neglected in dietary reconstruction based on stable isotopes. An earlier study suggested that cave bears (Ursus spelaeus) probably gave birth to cubs during dormancy. We provide an estimate of the effect on stable isotopes of growth and metabolism and discuss collagen turnover in a population of cave bears. Based on a quantitative model, we hypothesized that bear cubs lactated their mothers during their first and second winters, but were fed solid food together with lactation during their first summer. This demonstrates the need to include physical growth, metabolism and collagen turnover in dietary reconstruction. Whereas the effects of diet and metabolism are due to fractionation, growth and collagen turnover are dilution processes. PMID- 10518326 TI - Evidence for on-line visual guidance during saccadic gaze shifts. AB - Rapid orientating movements of the eyes are believed to be controlled ballistically. The mechanism underlying this control is thought to involve a comparison between the desired displacement of the eye and an estimate of its actual position (obtained from the integration of the eye velocity signal). This study shows, however, that under certain circumstances fast gaze movements may be controlled quite differently and may involve mechanisms which use visual information to guide movements prospectively. Subjects were required to make large gaze shifts in yaw towards a target whose location and motion were unknown prior to movement onset. Six of those tested demonstrated remarkable accuracy when making gaze shifts towards a target that appeared during their ongoing movement. In fact their level of accuracy was not significantly different from that shown when they performed a 'remembered' gaze shift to a known stationary target (F3,15 = 0.15, p > 0.05). The lack of a stereotypical relationship between the skew of the gaze velocity profile and movement duration indicates that on line modifications were being made. It is suggested that a fast route from the retina to the superior colliculus could account for this behaviour and that models of oculomotor control need to be updated. PMID- 10518327 TI - Attention without awareness in blindsight. AB - The act of attending has frequently been equated with visual awareness. We examined this relationship in 'blindsight'--a condition in which the latter is absent or diminished as a result of damage to the primary visual cortex. Spatially selective visual attention is demonstrated when information that stimuli are likely to appear at a specific location enhances the speed or accuracy of detection of stimuli subsequently presented at that location. In a blindsight subject, we showed that attention can confer an advantage in processing stimuli presented at an attended location, without those stimuli entering consciousness. Attention could be directed both by symbolic cues in the subject's spared field of vision or cues presented in his blind field. Cues in his blind field were even effective in directing his attention to a second location remote from that at which the cue was presented. These indirect cues were effective whether or not they themselves elicited non-visual awareness. We concluded that the spatial selection of information by an attentional mechanism and its entry into conscious experience cannot be one and the same process. PMID- 10518328 TI - [The 41st congress of the Japanese Society of Clinical Hematology. Akita City, Japan. October 13-15, 1999. Abstracts]. PMID- 10518329 TI - [49th general meeting of the Japanese Society of Allergology. Hiroshima, Japan. October 18-20, 1999. Abstracts]. PMID- 10518330 TI - [The 39th annual meeting of the Japanese Society of Nuclear Medicine. Akita City, Japan. October 5-7, 1999. Abstracts]. PMID- 10518331 TI - [The 29th Western/Eastern regional meetings of the Japanese Society of Nephrology. 1999. Abstracts]. PMID- 10518332 TI - Connecting science and practice. PMID- 10518333 TI - HIV/AIDS and infection control: the debate continues. PMID- 10518334 TI - HIV/AIDS and infection control: the debate continues. PMID- 10518335 TI - HIV/AIDS and infection control: the debate continues. PMID- 10518336 TI - HIV/AIDS and infection control: the debate continues. PMID- 10518338 TI - Codes, codes, codes. PMID- 10518337 TI - HIV/AIDS and infection control: the debate continues. PMID- 10518339 TI - [Guided tissue regeneration in endodontics (2)]. AB - Part I of this article (J Can Dent Assoc 65:394-8) looked at the application of guided tissue regeneration (GTR) in endodontics, the mechanisms of action of GTR and its various presentations. Part II examines the technique for surgically inserting a membrane for GTR, based on a review of the current literature. PMID- 10518340 TI - Photopolymerization of composite resin using the argon laser. AB - Because of the dental profession's increased utilization of light-cured restorative materials, there has been a corresponding increase in research into the light sources used to initiate polymerization. The argon laser is one promising source, as the wavelength of light emitted by this laser is optimal for the initiation of polymerization of composite resins. The literature reflects a strong divergence of opinion about many aspects of the effectiveness of laser curing compared to conventional light curing. Research indicates that the argon laser offers a greater depth and degree of polymerization, less time required and an enhancement of the physical properties of composite resins polymerized. These advantages are offset by reports that the increased polymerization caused by the laser results in increased shrinkage, brittleness and marginal leakage. Dentists interested in the new technology need to monitor ongoing studies. PMID- 10518341 TI - The safety of home bleaching techniques. PMID- 10518342 TI - Statement on the ethical and legal considerations of treating patients with infectious diseases. Canadian Dental Association. PMID- 10518343 TI - HIV/AIDS and infection control: the debate continues. PMID- 10518344 TI - Curing time of dental materials. PMID- 10518345 TI - Licensure requirements and mobility of Canadian dentists. PMID- 10518346 TI - Food hygiene training--another dimension. PMID- 10518347 TI - Teenage pregnancy trends in England and Wales. PMID- 10518348 TI - Cytokines: what are they and what do they do? PMID- 10518349 TI - Antenatal HIV testing: a dilemma. PMID- 10518350 TI - Worldwide wastes. PMID- 10518351 TI - All-Party Parliamentary Group on Skin. PMID- 10518352 TI - Canada goose (Branta canadensis) droppings as a potential source of pathogenic bacteria. AB - Canada goose droppings, collected in parks to which the public had access, were screened for a range of bacteria that could be pathogenic in man. Droppings of Canada geese, and other waterfowl, did contain such bacteria, including some that are well-known causes of illness in man. These bacteria, plus a species of Salmonella that was experimentally inoculated into droppings, were shown to survive and multiply in the droppings for up to one month after their deposition by geese. Canada geese ranged further from water than other waterfowl species and thus distributed their droppings over a larger area of park grassland. This more widespread distribution of their droppings leads Canada geese to pose a greater potential health risk than other waterfowl studied here, but variations in human responses to challenge with bacteria, and variations in human and waterfowl behaviour in public parks, renders quantification of this risk impossible. PMID- 10518353 TI - Work-related stress among nurses: a challenge for health care institutions. AB - The issue of occupational stress amongst health care professionals is currently a major concern in health policy. The main goal of the present study has been to investigate the relationship between levels of stress among nurses, and some of the psychosocial and organisational characteristics of their job. The participants of the cross-sectional survey were female nurses (n = 218) chosen at random from public hospitals in Csongrad Country, Hungary. A self-administered questionnaire was the method used for data collection: this questionnaire contained various items on psychosomatic symptoms, self-perceived health, sociodemographic data, job satisfaction, health risk behaviours, drug consumption, emotional load and social support from peers. The findings suggest that the frequency of common psychosomatic symptoms (e.g. sleeping problems, tension headache, chronic fatigue or palpitations), regular alcohol drinking, heavy smoking, and frequent use of tranquilisers and sleeping pills can be read as an indicator of nurses' work-related stress level. Nurses with only primary education had the highest such levels, while those with baccalaureate-level education had the lowest. Furthermore, nurses aged 51-60 years and those on rotating night shift proved to be vulnerable to stress the most frequently. However, no significant differences were found between nurses working in-theatre and those non-theatre; nor was job satisfaction found to have a significant impact on the levels of stress experienced. The results suggest that supportive relationships with peers may reduce the occurrence of high stress levels among nurses, leading the author to conclude that social support and the psychosocial work climate should be improved in health care institutions. PMID- 10518354 TI - Genital thrush in women: the attitudes and practice patterns of General Practitioners in Teesside and north Yorkshire. AB - This study was performed in order to determine the attitudes and practice patterns of some of the GPs on Teesside and North Yorkshire in the management of their female patients complaining of genital thrush, a term often used to mean vulvo-vaginal candidosis an extremely common condition (Tobin, 1995; Elliott, 1998; Lopez-Martinez et al, 1984). Postal questionnaires were sent to 65 GPs on Teesside and the bordering areas of North Yorkshire. There were 45(69%) completed replies. Nearly all the GP's believed that antibiotic usage was associated with the development of this disorder; less than half (44%) of the patients seen had a confirmatory microbiological test in the form of examination of a high vaginal swab (HVS). The most frequently prescribed antifungal compound was found to be clotrimazole. PMID- 10518355 TI - Cigarette smoking habits among high school boys in a developing country. AB - The aim of this cross-sectional study was to determine the habits, practices, attitudes and knowledge about cigarette smoking among high school boys aged 15-19 years in the United Arab Emirates, and to provide a basis for comparisons with international data. The World Health Organisation questionnaire was used, together with a multi-stage stratified cluster sampling technique; 1,700 subjects aged 15 years and above were randomly selected. A total of 1,486 individuals (87.4%) from among the populations of Al-Ain City, Abu-Dhabi and Dubai Emirates participated in the study. The prevalence of smoking among the studied group was 19%; 28.2% admitted that they had smoked before but had now given up, and the remaining 52.9% denied having ever smoked. Among 18-year-olds (or older) 30.3% smoked. In 70.8% of cases a friend was reported as having been the first source of their cigarette. Fifty-four percent of smokers started between the age of 10 and 15 years. The families of 15.7% of those studied approved of their smoking, while 78.3% did not: 6% did not have an opinion. Nearly two-thirds of the smokers (66.5%) wanted to stop smoking, while the remaining third (33.5%) did not. Differences in parental education (specifically that of the father) were found to have a significant effect on attitudes towards smoking. Contrary to expectations, the highest prevalence of smoking was found among sons of university graduates, and the lowest among sons of illiterate fathers (12.6% and 24.3%, respectively). There was a statistically significant difference in respect to family income and smoking. Among the ex-smokers, religion (40%) and health (26%) were important reasons for giving up smoking. Of the smokers, 33% claimed that stress is the most important factor which makes people smoke. The source of the student's information regarding smoking hazards was lowest from doctors (17-19%), and highest from the media (35%). All student groups were equally aware that smoking is a risk factor for lung cancer, respiratory diseases, and ischaemic heart diseases, but among the smokers only 28.9% were very concerned about the harmful effects of cigarettes. At the time of their graduation (18 years or older), one third of the students were already regular smokers, a figure liable to increase as they start university or work. PMID- 10518356 TI - The relationship between building design and escapes from secure units. AB - The research presented is a survey of all of the identified forensic psychiatry Medium Secure Units (MSUs) within Great Britain. The study examined the numbers and nature of all escapes from these units over a 42-month period. The research focused on the relationship between the building design of the units and the nature of patient escapes. The results showed that overall numbers of patient escapes from the units was low. The average number of escapes was 3.14 per unit over the three-and-a-half-year time-span of the survey. However, the recent increases in bed numbers in medium secure units had been paralleled by a larger percentage rise in escapes. Most escapes from the units occurred from three main areas. These were the perimeter fence, the roof, and internal and external windows. The implications of the methods of escape and their relationship to the building design of MSUs were discussed and recommendations made for future building designs. PMID- 10518357 TI - Nutrition: a necessary adjunct to hospital care? AB - Good nutrition is undoubtedly central to optimum health and to the recovery from illness yet research and clinical surveys repeatedly demonstrate an unacceptably high incidence of malnutrition in hospital patients in whom it delays recovery, increases the incidence of complications and significantly increases the cost of treatment; the human cost is inestimable. PMID- 10518358 TI - Health information systems in developing countries: benefits, problems, and prospects. AB - Health information systems are important support tools in the management of health care services delivery in both developed and less developed countries. An adequate health information system is vital not only for assessing the health needs of populations and groups, but also for planning and implementation of health interventions. It is equally important in the evaluation of programmes from both the perspectives of effectiveness and coverage. This paper examines the practical difficulties of health care provision amidst inadequate statistics to inform decisions. Major obstacles to the introduction of effective health information systems in developing countries are examined, and practical suggestions on measures to overcome them discussed. It is concluded that the establishment of well co-ordinated information collection systems at the various levels of the health care system in developing countries, using appropriate staff, could contribute greatly to improvements in health care delivery. PMID- 10518359 TI - A quality roadmap of a restructured hospital. AB - This paper presents a quality roadmap of a restructured hospital. Specifically, a case study is showcased to reveal how a FOCUS-PDCA model is applied to the Central Portering Services (CPS) within a restructured hospital, to provide better service to other departments within the hospital who can then in turn value add to the patient care delivery chain. The preliminary results suggest that implementers of quality and service improvement programmes using the FOCUS PDCA approach should note some of the key implementation difficulties. These difficulties occur at all levels and include issues relevant to change management, process improvement programmes, accessibility to data, communication and judicious execution of the change process. PMID- 10518360 TI - Lessons to be learned: a case study approach: pseudohyperkalaemia due to thrombocytosis in a case of tubo-ovarian abscess. AB - The case described here is that of a 48-year-old lady who presented with abdominal pain and fever; at a later stage she was found to have hyperkalaemia of uncertain origin. Blood examination revealed there to be marked elevation of the platelets (thrombocytosis) on some occasions. It was then realised that there was correlation between the platelet levels and serum potassium values. During the clotting process the release of potassium from the increased number of platelets caused the serum potassium to be elevated on account of an in vitro effect. The important point is that the raised serum potassium levels were not due to an in vivo phenomenon and, therefore, the patient did not need treatment for this; however, the presence of thrombocytosis was itself a clue to the diagnosis--which was eventually recognised as being due to an infection. At operation a tubo ovarian abscess was discovered to be the cause of the problem. PMID- 10518361 TI - Biochemical markers for assessment of bone metastases in patients with breast cancer. AB - Breast cancer commonly metastasizes to bones, producing both osteolytic and osteoblastic deposits. Different markers for quantitative determination of bone turnover have been developed to evaluate bone metastases of breast cancer. The urinary deoxypyridinoline (Dpd), a crosslink product of collagen molecules found in bone and excreted in urine during bone degradation, and bone specific alkaline phosphatase (B-ALP), an isoenzyme localized in the membrane of osteoblasts and released in circulation during bone formation, were recently described as a group of markers of bone turnover in metastatic cancer. The urinary Dpd/creatinine (Cre) ratios and the serum B-ALP activity were determined in the samples from 148 patients who suffered from breast cancer (BC patients) with or without bone metastases, and 42 healthy women. For comparison, other biochemical markers, e.g. carcinoembryonic antigen (CEA), CA15-3, tissue polypeptide antigen (TPA), tissue polypeptide specific antigen (TPSA), and total alkaline phosphatase (T-ALP) in these samples were also evaluated. The results showed that there was a significant difference in urinary Dpd/Cre ratio between the control group and the patients with breast cancer (BC group) (mean +/- S.D., 5.69 +/- 1.26 vs. 8.19 +/- 3.95 nM/mM, P < 0.05). However, there was no significant difference between their B-ALP activities in the two groups. In addition, the BC patients with bone metastases showed elevated urinary Dpd/Cre ratios and B-ALP activities and ratios of (Dpd/Cre)/B-ALP in compare with BC patients without bone metastases (P < 0.05). Meanwhile, the urinary Dpd/Cre ratios (10.50 +/- 5.04 nmol/mmol) in the advanced stage of BC patients were higher than those in an early stage (7.45 +/- 3.23 nmol/mmol) (P < 0.05), but their serum B-ALP activities increased only in stage IV (P < 0.05). The urinary Dpd/Cre ratios also increased progressively according to the degree of bone metastases (P < 0.05), but their serum B-ALP activities only increased in severe bone metastases (P < 0.05). The results showed that the increase of a bone osteolytic activity took place earlier than that of a bone osteoblastic activity in the metastatic BC patients. In compare with other conventional markers, the best diagnostic efficiency of biochemical markers, analyzed by step wise discriminate analysis, was provided by CEA followed by Dpd/Cre ratio, CA15-3, TPA, TPSA, B-ALP and T-ALP. We conclude that showed the urinary Dpd/Cre ratio was a useful tumor marker to evaluate breast cancer with bone metastases. PMID- 10518362 TI - The experience to use a modified en bloc excision technique in vitrectomy for diabetic traction retinal detachment. AB - Modified en bloc excision is a technique in which all posterior hyaloid is excised except the portions essential for membrane dissection. Then, bimanual dissection techniques allow excision of retained fibrovascular membrane "en bloc" with hyaloid. In a consecutive series of 16 eyes with diabetic traction retinal detachment treated with this technique, visual acuity of 5/200 or better was obtained in 11 (69%) of the cases, and complete macula reattachment was noted in 14 (87%) of 16 eyes. The final visual acuity was improved in 12 (75%) cases. While this surgical technique allows a higher rate of anatomic success and less postoperative morbidity, visual results remain limited by irreversible alteration in retinal function. PMID- 10518363 TI - Evaluation of proliferative activity in middle ear cholesteatoma using proliferating cell nuclear antigen. AB - Middle ear cholesteatoma has a remarkable invasive activity accompanied by destruction of ossicles and temporal bone. Its aggressive growth and high tendency to recur have impact on the postoperative care of the patients. Proliferating cell nuclear antigen (PCNA) is a 36 KDa DNA-delta-polymerase associated protein whose level of synthesis has been found to correlate directly with rates of cellular proliferation. In this present study, we used ABC (avidin biotin complex) technique and monoclonal antibody to PCNA to evaluate the expression of PCNA in 37 cases of cholesteatoma epithelium and 21 cases of normal postauricular skin. The rate of PCNA-positive cells in basal, parabasal, and upper layer of cholesteatoma epithelium tissue is 78% (29 cases), 68% (25 cases), and 41% (15 cases). In each layer of the postauricular skin tissue is 71% (15 cases), 67% (14 cases) and 34% (7 cases). No statistical difference of expression of PCNA-positive cells exists between each layer of cholesteatoma epithelium and normal postauricular skin; however, a tendency of higher PCNA-positive cells in cholesteatoma epithelium was observed. Immunohistochemical method of PCNA has the advantages of spatial architecture preservation, the relative simplicity of the methodology and the rapid acquisition of results. Although the etiology and histopathology of the growth pattern and osteolytic activity of cholesteatoma are unclear, information on cell kinetics may assist in cholesteatoma classification and may help predict the risk of recurrence and bone destruction. The results of this report indicate that cholesteatoma has a similar proliferative activity to the normal postauricular skin, and cholesteatoma itself is not a real tumor, despite its clinical behavior, which is similar to neoplastic cells. It is necessary to further study whether the cell kinetic information we obtained from the PCNA immunohistochemical analysis provides a valuable tool in accessing the prognosis of the cholesteatoma. PMID- 10518364 TI - Detection and identification of Mycobacterium tuberculosis by nested PCR assays in cerebrospinal fluid samples from patients with suspected tuberculous meningitis. AB - We used the nested polymerase chain reaction (PCR) assays developed previously to detect and identify Mycobacterium tuberculosis (M. tuberculosis) in the cerebrospinal fluid (CSF) samples from patients with suspected tuberculous meningitis and non-tuberculous patients. Our nested PCR assays target the multi copy IS6110 insertion element and the single-copy mtp40 genomic DNA of M. tuberculosis. These assays, when used in combination, allowed us to detect a very low number of M. tuberculosis in the CSF samples, which otherwise would be undetectable by the culture method, and to distinguish M. tuberculosis from M. bovis. We applied these nested PCR assays to analyze eleven CSF samples. Among these, five of them were from patients with suspected tuberculous meningitis but all except one were culture negative. Our results of PCR assays show that three of these five are M. tuberculosis positive, one of which is M. bovis positive, and only one is M. tuberculosis negative. The other six CSF samples were from the clinically diagnosed non-tuberculous patients. Surprisingly, two of these so called non-tuberculous patients, a subarachnoid hemorrhage (SAH) and the syndrome of inappropriate antidiuretic hormone secretion (SIADH), were shown M. tuberculosis positive. This finding supports a long-standing argument that tuberculous meningitis is one of the causes of these neurological diseases. These nested PCR assays thus provide the neurologists with an important adjunct, in addition to the patient's clinical presentation and laboratory data, for the diagnosis of tuberculous meningitis. PMID- 10518365 TI - Prognosis of spontaneous intracerebral hemorrhage in hemodialysis patients. AB - During an 8-year period, 32 consecutive patients with chronic renal failure on maintenance hemodialysis were diagnosed to have cerebral hemorrhage. The outcome was determined using the activity of daily life (ADL) at 6 months after hemorrhage. The overall mortality was 64%. Of the 12 surviving patients, no one made a good recovery (back to normality), 5 recovered to ADL grade II, 4 to grade III, 1 to grade IV, and 2 to grade V. Up to 91% of the patients had a history of hypertension. On admission, Glasgow coma scale (GCS) was 15 in 8 cases, 8-14 in 10, and below 8 in 14. The poor prognostic factors showing statistical significance included a poor admission GCS, age above 65 years, and blood sugar level of more than 200 mg/dl. Other factors which apparently were not related to the outcome included sex, history of stroke, acute myocardial infarction, hypertension, and diabetes mellitus, the locations of hemorrhage, the duration of hemodialysis, treatment modality (surgery vs non-surgery), and the laboratory data (blood urea nitrogen, creatinine, platelet count, hemoglobin, prothromin time, and partial thromboplastin time). This study confirmed a poor prognosis for hemodialysis patients with cerebral hemorrhage. More attention should be paid to the control of blood sugar in this group to improve the outcome of cerebral hemorrhage in hemodialysis patients, especially in elderly patients with poor admission GCS. PMID- 10518366 TI - The relation between admission balance and functional outcomes following stroke rehabilitation: a medical center based study. AB - This prospective study evaluated the clinical use of the Fugl-Meyer Balance Scale (FMBS) on stroke patients during hospitalization and assessed the relationship between balance score at admission to the rehabilitation program and functional outcome at discharge. One hundred and sixty-three stroke patients admitted to the in-patient rehabilitation department of a university-based medical center between January 1 and December 31, 1997 were recruited for this investigation. Functional ability was evaluated with the Functional Independence Measure (FIM) instrument, and balance was measured using the 7-item Fugl-Meyer Balance Scale. These measures were assessed both at admission to and discharge from the inpatient rehabilitation program. Pearson correlation and multiple regression analyses were used to determine the relationship between balance and functional ability scores at admission and rehabilitation outcomes at discharge, including length of stay, functional gain, and efficiency. The results demonstrated that the balance score at admission accounted for 6% of the variation in length of stay, once demographic influences were controlled. The FIM efficiency score could possibly be predicted by the balance ability at admission, which accounted for 3% of the variance. However, the balance score could not provide predictive information about the FIM gain beyond that already provided by the FIM score at admission, which accounted for 4% of the variance with demographic factors controlled. Overall, balance ability at admission, assessed by the Fugl-Meyer Balance Scale, had no or at least only little, contribution to account for the variance in rehabilitation outcomes. These findings suggest that the use of Fugl-Meyer Balance Scale at admission to stroke inpatient rehabilitation seemed not to enhance the ability to predict rehabilitation outcomes. PMID- 10518367 TI - Mixed germ cell tumor presenting as intratumoral hemorrhage: report of a case originated from the pineal region. AB - A 17-year-old male patient was brought to our clinic because of sudden onset of headache, vomiting, followed by transient loss of consciousness during a strenuous exercise. Neurologic examinations revealed that the patient had severe sensorimotor and brain stem dysfunction. Examinations of cranial CT and MR imaging showed a huge heterogeneously enhanced tumor originated from the pineal region with tumoral hemorrhage. The tumor markers were found to be high in AFP but not the beta-HCG and CEA. A clinical diagnosis highly suggestive of germ cell tumor was made. Prior to the planned emergency radiation therapy, he received an external ventricular drainage (EVD) and open biopsy of the tumor. Due to a postoperative complication of cerebellar hemorrhage observed 8 hours later, another maneuver was therefore required to extirpate the pineal tumor and cerebellar hematoma. The histological diagnosis proved to be a mixed germ cell tumor with tumoral hemorrhage. Spontaneous intratumoral hemorrhage in germ cell tumor of the pineal region is rare, probably due to compromised venous circulation within the tumor. The bleeding propensity, which may contribute to the formation of cerebellar hematoma, warrants a special attention when a biopsy procedure is to be performed. PMID- 10518369 TI - Peripapillary arterial loop--case report. AB - Peripapillary arterial loops are uncommon congenital retinal arterial anomalies, which are characterized by retinal arterial loops that originate and terminate on the retina beyond the optic disc borders. We report a case of peripapillary arterial loop associated with preretinal and vitreous hemorrhage. The fluorescein angiography demonstrates the arterial loop and leakage from the loop and its capillaries. The origin of hemorrhage of peripapillary arterial loop might result from the loop and its adjacent capillaries. PMID- 10518368 TI - Traumatic ossicular chain discontinuity--report of two cases. AB - In addition to hemotympanum and traumatic eardrum perforation, traumatic ossicular chain discontinuity should also be considered in the differential diagnosis of conductive hearing impairment resulting from head injury. The most common form of these ossicular chain lesions following head injury is incudostapedial joint (I-S joint) separation. We successfully managed two patients with I-S joint separation resulting from head injury through exploratory tympanotomy with ossiculoplasty within the recent 2 years. Both were young females who had sustained head injury resulting from traffic accident with the sequelae of persistent hearing impairment. They both gained significant hearing improvement postoperatively. PMID- 10518370 TI - Expressing estimators of expected quality adjusted survival as functions of Nelson-Aalen estimators. AB - Quality adjusted survival has been increasingly advocated in clinical trials to be assessed as a synthesis of survival and quality of life. We investigate nonparametric estimation of its expectation for a general multistate process with incomplete follow-up data. Upon establishing a representation of expected quality adjusted survival through marginal distributions of a set of defined events, we propose two estimators for expected quality adjusted survival. Expressed as functions of Nelson-Aalen estimators, the two estimators are strongly consistent and asymptotically normal. We derive their asymptotic variances and propose sample-based variance estimates, along with evaluation of asymptotic relative efficiency. Monte Carlo studies show that these estimation procedures perform well for practical sample sizes. We illustrate the methods using data from a national, multicenter AIDS clinical trial. PMID- 10518371 TI - Specification tests for random-effects transition models: an application to a model of the British youth training scheme. AB - We develop diagnostic tests for random-effects multi-spell multi-state models focusing on: independence between the unobserved heterogeneity and observed covariates; mutual independence of heterogeneity terms; and distributional form. They are applied to a transition model of the British youth labor market, revealing significant mis-specifications in our initial model, and allowing us to develop a considerably better-fitting specification that would have been difficult to reach by other means. The improved specification implies reduced estimates of the effectiveness of the youth training scheme (YTS), but we nevertheless retain the conclusion of significant positive effects of YTS on employment prospects. PMID- 10518372 TI - Multi-state models: a review. AB - Multi-state models are models for a process, for example describing a life history of an individual, which at any time occupies one of a few possible states. This can describe several possible events for a single individual, or the dependence between several individuals. The events are the transitions between the states. This class of models allows for an extremely flexible approach that can model almost any kind of longitudinal failure time data. This is particularly relevant for modeling different events, which have an event-related dependence, like occurrence of disease changing the risk of death. It can also model paired data. It is useful for recurrent events, but has limitations. The Markov models stand out as much simpler than other models from a probability point of view, and this simplifies the likelihood evaluation. However, in many cases, the Markov models do not fit satisfactorily, and happily, it is reasonably simple to study non-Markov models, in particular the Markov extension models. This also makes it possible to consider, whether the dependence is of short-term or long-term nature. Applications include the effect of heart transplantation on the mortality and the mortality among Danish twins. PMID- 10518373 TI - Predicting a future lifetime through Box-Cox transformation. AB - In predicting a future lifetime based on a sample of past lifetimes, the Box-Cox transformation method provides a simple and unified procedure that is shown in this article to meet or often outperform the corresponding frequentist solution in terms of coverage probability and average length of prediction intervals. Kullback-Leibler information and second-order asymptotic expansion are used to justify the Box-Cox procedure. Extensive Monte Carlo simulations are also performed to evaluate the small sample behavior of the procedure. Certain popular lifetime distributions, such as Weibull, inverse Gaussian and Birnbaum-Saunders are served as illustrative examples. One important advantage of the Box-Cox procedure lies in its easy extension to linear model predictions where the exact frequentist solutions are often not available. PMID- 10518374 TI - A note on inconsistency of NPMLE of the distribution function from left truncated and case I interval censored data. AB - We show that under reasonable conditions the nonparametric maximum likelihood estimate (NPMLE) of the distribution function from left-truncated and case 1 interval-censored data is inconsistent, in contrast to the consistency properties of the NPMLE from only left-truncated data or only interval-censored data. However, the conditional NPMLE is shown to be consistent. Numerical examples are provided to illustrate their finite sample properties. PMID- 10518375 TI - [Evidence of our history]. PMID- 10518376 TI - Secretory function of adrenal cortex in chronic alcoholis. AB - Three groups of male subjects (healthy subjects, chronic alcoholics with liver cirrhosis and patients with acute viral hepatitis) were included in a 24 hour pattern of excretion of the total and some fractions of 17-ketosteroids (KS), basal concentration of 11-hydroxycorticosteroids (11-OHCS) and dehydroepiandrosterone (DHEA) in plasma, as well as changes of concentration of the same steroids in plasma 15, 30 and 60 minutes after a single i.m. injection of insulin. In regard to healthy subjects and patients with acute viral hepatitis, chronic alcoholics with liver cirrhosis excrete decreased quantities of total and some fractions of 17-KS. In regard to healthy subjects, decreased excretion of the sum androsterone and etiocholanole was established as well as increased DHEA secretion in patients with acute viral hepatitis. In chronic alcoholics with liver cirrhosis basal concentrations of 11-OHCS in plasma and their increase after insulin administration are the same as in healthy subjects, but values of DHEA concentrations in plasma are decreased. It has been pointed to the possibility of damages of the secretory function of adrenal cortex in chronic alcoholics with liver cirrhosis. On the basis of above mentioned results, there is an assumption that adrenal gland primarily provides normal secretion of C21 steroid and thus, satisfying needs for these steroids, increases secretion of DHEA. Follow up of DHEA urinary secretion may provide insight into basal activity of adrenal cortex, whereas the functional state of the liver must be taken into account when interpreting the results. PMID- 10518377 TI - Cytokeratins 7 and 14 in special types of invasive breast carcinomas. AB - Cytokeratins are typical constituents of the cytoskeleton of the epithelial cells and some other cell types. In normal breast tissue the luminal epithelium of the ducts and acini consistently expresses cytokeratin 7 (as well as 8, 18 and 19) while the myoepithelial cells are consistently cytokeratin 14-positive. We studied 20 invasive lobular, 10 medullary and 10 tubular breast carcinomas on expression of cytokeratins 7 and 14 using monoclonal primary antibodies in an automatic immunochemical system. All the carcinomas were cytokeratin 7-positive, but the percentage of positively stained tumor cells varied. The tumor cells of two medullary carcinomas expressed cytokeratin 14, a finding which can be interpreted as a result of squamous differentiation. Mapping of the cytokeratin profile of the special type of breast carcinomas has a practical importance for a more successful typing of adenocarcinomas in primary and metastatic sites. PMID- 10518378 TI - [Physical medicine in the diagnosis and treatment of functional disorders of the spinal column]. AB - INTRODUCTION: Manual (Physical) Medicine is concerned with physiology, pathophysiology, diagnosis and therapy of the musculoskeletal system functional disorders. Functional disorders are caused by direct or indirect injury (muscular spasm), inflammation, sudden movements etc. Functional disorders in regard to decreased joint volume are also called segmental or peripheral joint dysfunction, somatic dysfunction and function blocking. Blocking means: reversible disorder with decreased functional joint movement; muscular spasms caused by neurophysiological changes including decreased movement; pain and functional disorder of internal organs and tissues being part of the joint. Intervertebral joints, intervertebral discs, muscles and their nerve control belong to such control circles which are integrated within an organism. The basic principles of Manual (Physical) Medicine are good knowledge of functional anatomy and three dimensional imaging of biochemical functions of the spine. The most important functions of the spinal column are as follows: it is an organ of axis with a protective and supportive function and it enables normal walk and balance. The basic functional unit in the architecture of the vertebral column is the dynamic spinal segment. Its basic function is movement, posture maintenance and protection of spinal nerve roots. The functional disorder of one part of the dynamic segment cannot be isolated from other parts. Thus functional blocks within the fascial joints, abnormalities within intervertebral discs, changes within the ligament apparatus, and muscular disorders may be both causes and consequences at the same time. The dynamic segment is thus included into a mechanical functional circle. Its mechanical function is tightly connected with the statics of the whole body. Different leg length, severe abnormalities of great leg joints may cause blocks within the fascial joints. The vertebral dynamic segment is also a part of the nervous, functional reflex circle. Clinical symptoms associated with functional disorders within intervertebral joints are as follows: restricted joint mobility and nervous reflex disorders (local segmental and peripheral segmental injuries). MANUAL (PHYSICAL) DIAGNOSIS: In order to make physical diagnosis it is necessary to have good knowledge of anatomy, functional anatomy, kinesiology and neurology. History taking is of great importance for the physician who establishes physical therapy. It is also necessary to note various abnormalities, muscle atrophy and to analyze patient's walk. Palpation at rest reveals changes, skin disorders, subcutaneous and fatty tissues and muscles and it assesses the status of soft tissues. By physical examination of the spine it is possible to diagnose intervertebral joint blocks. Thus examination of joint mobility is performed. It is important to differ physiological, anatomic and pathologic ranges of mobility and to determine the greatest mobility and joint play. Joint play represents the property of the joint to achieve some additional mobility. This examination requires knowledge of a special technique. Palpation of local segmental irritation is performed on all spinal segments after a special technique examining bone-transversal joints as well as sacroiliac joints. Local irritation points, muscle spasms and tendomyosis are being detected. The diagnosis of blocks is also the therapeutic diagnosis at the same time. Physicians using physical medicine must apply x-ray of the spinal column, whereas the aim of physical therapy is to eliminate functional blocks, which occur in the form of reduced reversible mobility in one or more directions. Only physical examination can establish this, while apparatuses cannot objectivize it. Manipulative techniques on the spine should be performed only by specialists. MANUAL (PHYSICAL) THERAPY: Manual (Physical) therapy comprises numerous techniques and is used in various indications. (ABSTRACT TRUNCATED) PMID- 10518379 TI - [Physiologic neonatal body weight loss in a "baby friendly hospital"]. AB - INTRODUCTION: It has been known for centuries that mother's milk is the most optimal nutritive and energetic substance for nutrition of newborn infants. Human milk is naturally adapted to physiological possibilities of nutrition and nutritive requirements of newborn infants. Baby Friendly Hospital Initiative of WHO experts and International Children's Fund are directed to introduction of natural diet and breast feeding as long as possible and they have been accepted in our environment as well. Health workers of these Departments and Hospitals help in education of women giving birth for successful breast-feeding. Successful establishment of lactation should affect physiological body-weight loss reduction in newborn infants. The aim of this paper was to prove that babies' presence induces earlier lactation, and that is how physiological body-weight loss is reduced. MATERIAL AND METHODS: This study included 100 newborn infants of the Baby Friendly Hospital and 100 newborn infants born at the Maternity Ward of the Hospital in Senta in 1997 who were situated in a special ward for newborns. The paper deals with vaginally delivered babies without signs of disease. The study registered as follows: sex of newborn infants, the lowest body-weight during their stay at the Department, date when the body-weight was measured, taking into consideration the parity of their mothers. RESULTS: Statistical data processing revealed that in the group of newborn infants included in the Baby Friendly Program physiological body-weight loss was less (5.32% of the body-weight at birth) than in newborn infants classically managed (5.77% of body-weight at birth). The physiological body-weight loss in the group of the Baby Friendly Program was significantly smaller when expressed in grams. DISCUSSION: According to pediatric literature a 10% physiological loss of body weight in regard to body weight at birth is considered to be normal. Investigations performed at the Maternity Wards in Zrenjanin and Novi Sad (geographically the nearest to our town) have shown results similar to ours. CONCLUSION: Results of our investigation support the Baby Friendly Hospital Initiative of the WHO and International Children Fund and their recommendations for initiating breast feeding of newborn infants as early as possible. We also consider efforts for providing conditions necessary for Baby Friendly Hospital justified because of mother-child close relationship and possibilities for breast-feeding without strict regimens. PMID- 10518380 TI - [Hormonally-active ovarian tumors in 30 years of data (1965-1994)]. AB - INTRODUCTION: Authors have investigated the incidence and distribution of hormonally active ovarian tumors in a 30-year surgical material. Out of 552 ovarian tumors, there were 28 hormonally active tumors (5.07%): 18 granulosa cell tumors, 4 thecomas, 2 arrhenoblastomas, 2 malignant v. Kalden folliculomas. There were laso 2 benign tumors: ectopic adrenal ovarian tumor, and ovarian tube hydatidiform mole. They secrete estrogen and testosterone and their effects are evident: prior to puberty (Praecox pubertas), during the reproductive period, but most often in menopause. In order to make the diagnosis, when uterine hemorrhage occurs, it is necessary to perform explorative curettage, but the tumors must be surgically removed. It is necessary to perform that kind of surgery, which corresponds to the age of the woman. Recurrences may appear even 35 years later. MATERIAL AND METHODS: Hysterectomy with bilateral adnexectomy was performed in 18 (64.28%) postmenopausal women. In 4 (14.29%) women of reproductive age unilateral adnexectomy was performed, while explorative laparotomy was performed also in 4 (14.29%) women. 5-year survival in the first stage of the disease was 75% (20 women). During the first three years 5 (17.86%) women decreased in the IV stage of the disease, as well as one woman (3.57%) in the II b stage, which makes a total of 6 women (21.43%) with fatal outcome. Ovarian tumors also include hormonally active tumors which secrete female and male sex hormones, and whose effects are evident on hormonal receptors. They are divided into benign or malignant, whereas they are all potentially malignant. We differentiate two groups: 1. Feminizing mesenchymomas, which secrete female sex hormones and 2. Virilizing tumors, which secrete male sex hormones. These tumors may occur prior puberty, during the reproductive period and in the postmenopausal period--senium. They can cause minor, long-term or hemorrhages similar to menstruation, as well as hypertrophy of the myometrium. Glandular cystic hyperplasia occurs often, whereas proliferation is rare. Histopathological findings after surgery (for example of myoma) are often surprising, because they reveal hormonally active undetected ovarian tumors or glandular cystic endometrial hyperplasia. All hormonally active tumors are potentially malignant. The aim of this retrospective study was to compare our prior attitudes and interventions with newer attitudes and to make changes on behalf of our patients. RESULTS: Our investigation included 552 adnexal tumors in a 30-year material. Each woman with uterine hemorrhage had undergone explorative curettage and the material was sent for histopathological analysis. During the investigated period 28 hormonally active tumors were found: 2 benign and 26 malignant or potentially malignant tumors (Table 2). There were 18 cases (40.91%) with granulosa cell tumors, 6 cases (13.64%) with cystadenocarcinoma ovarii serosum; 3 cases (6.82%) with borderline tumors; 4 cases (9.09%) with thecoma of the ovarii. Pseudomucious adenocarcinoma was found in 2 cases (4.55%) as well as endometrial adenocarcinoma, malignant mesenchymoma, arrhenoblastoma, malignant folliculoma v. Kalden; whereas malignant teratoma was established in 1 case (2.27%), as well as anaplastic carcinoma, metastatic carcinoma--Krunberg. It is obvious that hormonally active tumors make almost half of the cases (46.43%) and occur mostly at the age of 50-59 years of age (3.4%). Out of 28 patients, 18 underwent hysterectomy and bilateral adnexectomy (64.28%); 4 (14.29%) underwent unilateral adnexectomy that is explorative laparotomy for taking bioptic samples for histological examination (14.29%). Out of 28 patients, 4 were women of reproductive age. All of them underwent adnexectomy and are alive. Hemorrhages usually occur a few years after menopausal period. In one case it occurred 35 years after menopause (arrhenoblastoma), in another 25 years after menopause due to endometrial carcinoma asso PMID- 10518381 TI - [Primary ultrasonic diagnosis of congenital hip dysplasia in neonates at the Senta Hospital]. AB - INTRODUCTION: Congenital hip dislocation is an abnormality which has not been managed in the period of early infantile period. However, clinical and ultrasound screening performed in the first days of life after birth at the Labor and Delivery Departments, provide prompt application of certain preventive and therapeutic measures which are painless, nonaggressive and highly efficient at this age. That is why primary sonoscreening of congenital hip dislocation of newborn infants in the first three days of life has been introduced at the Labor and Delivery Department of the Hospital in Senta. Methodology after Graf using a 5 Mhz linear probe is applied. Examinations are performed in the lateral position, giving a morphological finding, whereas morphodynamic data are processed after Graf classification. In clinically unstable and sonographically critical hips a dynamic hip examination is performed after Harcke. This is a teamwork of a neonatologist, ultrasonographist and child orthopedist and management of pathological hips starts at our department. Depending on indications therapeutic panties. Pavlik harness or Von Rosen band are used. Mothers are instructed how to use these appliances and take care of their newborn infants in these circumstances which helps in accepting the necessary therapeutic procedures. RESULTS: In the period 1991-1998, 14,378 hips were examined at the Labor and Delivery Department of the Hospital in Senta. A normal sonographic finding of Ia type was found in 45.87% of hips, while Ib type was found in 18.58% of hips. There were 31.16% of sonographically immature hips of IIa plus type, whereas a pathological finding was found in 1.39% of hips. In this group of pathological hips there were 0.98% of sonographically critical hips of IIg type. 0.26% of sonographically decentered hips of IID type. 0.09% of decentered hips of IIIa type and 0.06% excentric hips of type IV after Graf. In pathologic hips, check-ups were performed every 3 weeks and every 6 weeks in sonographically immature hips. CONCLUSION: Considering the fact that the first three months are the period of the most intensive maturation, that is the phase of absolute reduction of excentric and decentered hip in newborn infants, check-ups for infants with normal hip clinical and ultrasonographic findings were obligatory after the third month. Results of early nontraumatic and dynamic treatment proved successful on the basis of control clinical, functional, ultrasonographic and radiographic parameters. During the studied period at our territory there were no cases with aseptic necrosis of femoral head, nor necessity for surgical correction of the congenital hip dislocation. PMID- 10518382 TI - [Complications in laparoscopic cholecystectomy]. AB - INTRODUCTION: For more than a century classical cholecystectomy has been a method of choice in surgical management of gallbladder diseases. At the end of eighties and at the beginning of nineties of this century, laparoscopic cholecystectomy has been introduced gradually taking the place of classical. Numerous studies deal with complications associated with this surgical procedure. COMPLICATIONS RELATED TO INSERTION OF VERESS NEEDLE: During insertion of Veress needle, injuries of the frontal abdominal wall may occur (9). Insertion of the needle into the peritoneal cavity may cause: injury of the omentum with consequential hemorrhage; intestinal and mesenteric injuries as well as injuries of urinary bladder and great blood vessels of retroperitoneum. Initial insufflation of CO2 through the needle, if it is inadequately placed, may cause subacute emphysema, intraperitoneal adhesion formation, mediastinal emphysema and pneumothorax (8 10). COMPLICATIONS RELATED TO TROCAR INSERTION--TROCAR INJURIES: Apart from injuries associated with insertion of Veress needle, insertion of trocar may cause injuries of the liver, spleen and falciform ligament (4). Trocar injuries are more severe (8,9). BILE DUCTS INJURIES: Bile ducts injuries are divided into mild (injuries of a smaller bile duct, incomplete ductus cysticus occlusion or partial that is lateral injury of greater ducts) and severe (injuries of duct choledochus or hepatic ducts) (1). Bile ducts injuries are extremely severe complications with high morbidity, long-term hospitalization and may be life threatening (11,12). INTRAOPERATIVE BLEEDING: Intensive and uncontrolled intraoperative bleeding, especially within the operative field, both arterial (from arterial cyst) and venous (gallbladder, vena porte) requires conversion. POSTOPERATIVE HEMORRHAGE: Postoperative hemorrhage usually occurs in cases of cystic artery damage, prolonged hemorrhage from the gallbladder, parenchymal liver injuries and as well as in open cholecystectomy it is an indication for reoperation (8,9). OTHER COMPLICATIONS: Other complications, thermic and mechanic diaphragmatic injuries and infections of incisional injuries and herniations at the place of trocar insertion. OUR EXPERIENCES: In the period June 1995-November 1997, 500 patients underwent laparoscopic cholecystectomy (401 female--80.2% and 99 male--19.8%) aging from 16-78 years of age, average age 46 years. 16 (3.2%) mild complications were recorded (reoperation was necessary in one case): 1) subcutaneous emphysema in 3 patients (0.6%) which gradually disappeared up to 36 hours after surgery; 2) Retzin's space emphysema in 1 patient (0.2%) causing dysuria for 3 days; 3) bilirea--bile excretion through subhepatic drain in 2 patients (0.4%)--from the second to seventh postoperative day (excretion did not exceed 500 ml a day) and it spontaneously stopped; 4) inflammatory hematoma of the falciform ligament (0.02%); 5) in 8 patients (1.6%) postoperative infection of infraxifoid wound was registered; 6) in 1 patient (0.2%) segmental bile duct injury occurred. Out of 500 laparoscopic cholecystectomies conversion was necessary in 19 cases (3.8%). CONCLUSION: In spite of numerous advantages and better comfort for patients, this method may have complications, whereas incidence of bile ducts injuries seems to be higher than in the classical procedure. The more laparoscopic cholecystectomies are performed, the more bile ducts injuries occur. Out of 500 laparoscopic cholecystectomies performed at the Surgery Department of the hospital in Senta, 3.2% of mild complications were registered with 1 case needing reoperation, while in 1 case (0.2%) it was probably type A bile duct injury. PMID- 10518383 TI - [A 3-year study of ultrasound examination of the central nervous system in high risk neonates at the hospital in Senta]. AB - INTRODUCTION: Transfontanellar neurosonography is a noninvasive, precise method for detection and follow-up of morphological and functional changes of the central nervous system in infants and newborn infants, which has been used as sonoscreening since 1996 in General Hospital in Senta in high-risk infant newborns. MATERIAL AND METHODS: In the period 1996-1998 there were 2831 newborn infants at the Labor and Delivery Department. There were 551 (19.46%) newborn infants in the high-risk group. Ultrasonographic examinations of the brain were performed by SIEMENSSONOLINE SL2 apparatus, with a sector probe 3.5 Mhz. Out of the examined high-risk newborn infants 62.25% had a pathological ultrasound finding of the brain including: intracranial hemorrhage and hypoxic ischemic encephalopathies; 117 (21.23%) of newborn infants had intracranial hemorrhage Papile grade I-II; 66 (11.98%) newborn infants had III-IV grade intracranial hemorrhage and 160 (29.04%) had hypoxic ischemic encephalopathies. DISCUSSION: Correlation between pathological CNS findings and Apgar Score (AS) in 143 newborn infants, revealed that all newborn infants with AS: 0-3 had a pathological finding of the brain: HIC or HIE (4 newborn infants). In the group with AS 4-7 (51 newborn infants) pathological sonogram was established in 44 (86.28%), whereas in the group with AS 8-10 (88 newborn infants) pathological ultrasound finding of CNS including HIC or HIE was registered in 40.09% of newborn infants. Changes in regard to HIC of various degree and HIE changes depend on the localization, volume, time of onset (vulnerable period) and spreading time. Compensatory mechanisms as well as great brain plasticity at this age and early rehabilitation can reduce the consequences of brain damages a great deal. That is why it is necessary to inform parents about the changes that have occurred properly in order to ensure correct relationship to their high-risk newborn infant. In this way they cooperate at check-ups and are able to follow the development of their child, noticing possible aggravation going on with multiple stimulation treatment. PMID- 10518384 TI - [Analysis of pathohistological results from the uterine mucosa 1965-1998 at the gynecology department in Senta]. AB - INTRODUCTION: It is a well known fact that there is no invasive uterine carcinoma which was not a "carcinoma in situ" in the initial stage. On the basis of this knowledge it is evident that active systematic detection of malignant uterine neoplasms is of great importance. Incidence of cervical carcinoma had been at the first place for years, but in the last 10 years the number of diagnosed cancer of corpus uteri has significantly increased. Uterine hemorrhage is a problem at any age, but at menopause and senium it is of special importance, because at this age every hemorrhage must be considered carcinomatous, unless something else is established (1). Out of all methods, histopathological analysis of the biopsy material is the most reliable, but only complete, not fractional, uterine abrasion has a full diagnostic value. Explorative curettage was performed in all cases of polypectomies and cervix biopsies (2,3,4). MATERIAL AND METHODS: A retrospective study included all histopathological findings of curettages, polypectomies and biopsies performed due to hemorrhage in the period from 1965 to 1998. During this period 48,606 women were hospitalized at the Gynecology Department of the General Hospital in Senta. 9,475 (19.49%) underwent explorative curettage, partly for therapeutic and partly for diagnostic reasons. The biopsy material was sent for histopathological analysis, disregarding the quantity and macroscopic appearance. RESULTS: The greatest number of explorative curettages were performed in women from 41-50 years of age (41.35%). The number of interventions decreased in the sixth and seventh decade, with sudden increase over 70 years of age. The greatest number of functional bleeding occurred in women from 21 to 50 years of age (33.59%), whereas endometrial hyperplasia was diagnosed in 22.6% of patients 41-50 years of age. Biopsies and polypectomies with explorative curettages were most frequent in patients 61-70 years of age. Uterine carcinoma was diagnosed in 470 patients: out of this number cervical carcinoma was diagnosed in 234 patients, and corpus uteri carcinoma in 236 women. Cervical carcinoma occurred most frequently in the group of patients 41-50 years of age, whereas corpus uteri carcinoma was most frequent in the group 51-70 years of age. It is important to point out the correlation between cervical carcinoma and corpus uteri carcinoma being 1:1.04. CONCLUSION: 1. In order to detect corpus uteri carcinoma, especially in women over 40 years of age, it is necessary to pay special attention to all kinds of uterine bleedings. 2. Explorative curettage of the whole uterine cavity and histopathological analysis of biopsy material are considered to be the most reliable diagnostic methods for detection of carcinoma. 3. The correlation between cervical carcinoma and corpus uteri carcinoma is 1:1.04. It points to a difference in regard to corpus uteri carcinoma established in previous studies. PMID- 10518385 TI - [Effect of the "baby friendly" program on the number of neonatal infections at the maternity ward in Senta]. AB - INTRODUCTION: The aim of this paper was to assess how the Baby Friendly Hospital Initiative affects occurrence, structure and outcome of neonatal infections at our department. MATERIAL AND METHODS: This retrospective study included all newborn infants born in 1995, when all the babies were at the neonatal ward and all the babies born in 1998, who were with their mothers. Newborn infants with low-birth-weight or shortened gestation were excluded. The assumption was that faster onset of lactation and thus breast-feeding decrease incidence of infections, but that there is an increased risk due to hygienic habits of mothers, especially those with no qualifications and difficult living conditions. The paper assesses the percentage of infections occurrence. RESULTS: Occurrence of infection was established clinically, whereas general signs of infection, as well as local signs of infection were confirmed by laboratory and bacteriological findings. Antibiotic therapy was applied. In great number of infections Staphylococcus aureus was isolated. In 1998 a certain increase of low-birth weight and low gestation newborn infants was registered. In mature babies included into the Baby Friendly Program, number of infections has not changed, but the treatment was a little shorter. Infections were much more frequent in low birth-weight and low gestation newborn infants. On the average the treatment in such cases was a little longer, but not only due to infection. DISCUSSION: Baby Friendly Hospital Initiative has not significantly affected the incidence of intrahospital infections in newborn infants. On the average the treatment in mature newborn infants was shorter, probably due to better lactation and transfer of immunoglobulins from mother to child. CONCLUSION: If Baby Friendly Hospital Initiative means adequate epidemiological supervision of mothers, this program does not significantly affect the risk from intrahospital infections. PMID- 10518386 TI - [Cerebellar infarct with complications]. AB - INTRODUCTION: Symptoms of circulatory disorders depend on the volume and localization of the cerebellar infarction. An isolated blood vessel occlusion may have various symptoms due to numerous anastomoses in the cerebellar hemispheres. Dominant symptoms are vertigo, vomiting, nausea and balance disorders. Due to a great number of anastomoses between the three cerebral arteries, if only one of them is occluded, expected symptoms rarely occur, so that vertebral angiography performed in the first 24 hours from the onset of clinical picture, points to a lesion of the cerebellar parenchyma on the basis of radiologic signs of occlusion. Further on, brain computerized tomography solves the diagnostic dilemma, but unfortunately, ischemic cerebellar lesions are often detected only in pathoanatomic findings. Development and course of cerebellar malformation are of great importance for occurrence of symptoms. Cerebellar symptoms often occur in cases of brain-stem lesions. In dorsal hemisection lesions the following symptoms occur: vertigo, vomiting, nystagmus, taste disorders and secretory trophic disorders in the gastrointestinal tract. In cases of spreading to the pyramidal neurons the following symptoms occur: a soft palate and vocal cord paresis, swallowing difficulties and disorders of phonological as well as hemiplegia and sensibility disorders. CASE REPORT: This is a case of a male patient, 60 years of age. He was admitted to the Neuropsychiatric Department because of high blood pressure, severe headache in the back of the head, vertigo and nausea, without vomiting. He complained about weakness in the left side of the body. The somatic status was normal. In regard to neurologic status he had a slow photomotoric reaction of pupils to light, swallowing difficulties, hoarse voice, decreased pharyngeal reflex, symmetrically fast reflexes, walk on wide basis, in Romberg position unstable, whereas other findings were normal. Psychically average. The reaction to therapy was good. Subjectively the patient felt-well, but swallowing difficulties persisted. On the seventh day from admission his status suddenly aggravated, with severe vertigo, vomiting on several occasions, weakness and high temperature, hypotension, tachycardia, left eyelid ptosis with horizontal nystagmus when looking to the left. Lung radiography revealed bilateral pneumonia; computerized tomography of the brain revealed a great ischemic zone in the left cerebellar hemisphere discretely dislocating the brain mass to contralateral side. Resistant hypotension persisted. Surgical findings of the rectum revealed a dark content. Gastroscopy revealed esophageal candidiasis; mucus bleeding was spontaneous and at touch, whereas in the lower third of esophagus there was a mucous lesion. Haematinised blood was found in the stomach and duodenum. The patient's state continually aggravated and after massive stool blood loss a surgery was performed. A great penetrating ulcer was detected at the back duodenal wall 30 mm of size, visualizing the pancreas. A final surgical procedure was performed at the Clinic for Abdominal Surgery in Novi Sad. The postoperative course was good, but on the 7th postoperative day unexpected fatal outcome occurred due to massive lung embolism. Pathoanatomic examination revealed a postencephalomalacic pseudocyst in the left cerebellar hemisphere. CONCLUSION: This is a case report of a patient with lateral medullar syndrome associated with complications which can only partly be explained by the basic disease. Pneumonia and a secretory trophic disorder of the gastrointestinal tract are frequent complications that can be life threatening is spite of intensive management. PMID- 10518387 TI - [Helicobacter heilmanni (Gastrospirillum hominis) as a new cause of gastritis- case report]. AB - INTRODUCTION: During the course of research into Helicobacter pylori, further microorganisms colonizing the gastric mucosa have been detected. The large spiral shaped bacteria deviating from Helicobacter pylori was originally described by Dent. The bacteria was initially named Gastrospirillum hominis, but after sequencing of 16S rRNA it was classified as Helicobacter and named Helicobacter Heilmannii in honour of the pathologist Prof. Dr. Konrad Heilmann. We report two additional cases. CASE REPORTS: In 1997 at the Department of Pathology of General Hospital in Senta 268 gastric biopsies (151 male, 117 female), average age of 50.4 were analyzed. Helicobacter pylori was identified in 176 biopsies (65.7%). Helicobacter Heilmannii was found in two cases (0.74%). The clinical details were as follows: Case 1. A 32-year-old male complained of dyspepsia. Esophagogastroduodenoscopy disclosed hyperemia and scattered erosions in the lower third of stomach. Case 2. a 55-year-old female was admitted at the hospital with lumbar pain, loss of appetite and body weight. Esophagogastroduodenoscopy findings were normal. The patient lived in rural household in contact with various domestic animals. In the biopsy specimens of both cases in the antral gastric mucosa large bacteria resembling a corkscrew were found, 6-10 microns length, consisting of 5-9 regular tight spiral. The bacteria were midly eosinophilic and Gram negative. The bacteria stained strongly with the Warthin Starry silver stain, and with Giemsa stain. The distribution of Helicobacter Heilmannii was patchy in the central zones of the gastric pits, deep or more superficial including the foveolar opening on the mucosal surface. A mild degree of chronic gastritis characterized both cases, with, focally active inflammation and lymphoid aggregates. None of biopsies had atrophy, intestinal metaplasia or epithelial damage. There was a symptomatic improvement after a 4-week course with triple therapy (H2 antagonist & metronidazole & amoxycillin). After treatment rebiopsy specimens displayed mild chronic gastritis and no spiral organisms. CONCLUSION: Our reported cases are the first described cases of gastritis caused by Helicobacter Heilmannii in our region with successful eradication after therapy. Morphologically similar bacteria have been found in the stomachs of domestic animals. It is a general opinion that Helicobacter Heilmannii is characteristic for animals but it is occasionally transmitted to humans. PMID- 10518388 TI - [Anemia in chronic renal insufficiency--case report]. AB - INTRODUCTION: Chronic renal insufficiency is a clinical syndrome including increased urea in the serum and creatinine concentration with or without decrease of urine production. Numerous diseases may induce chronic renal insufficiency. In developed countries these are most often diabetes mellitus, glomerulonephritis and hypertension, whereas in developing countries these are bacterial infections, gall stone, tuberculosis and some parasitic diseases. Chronic renal insufficiency leads to functional disorders of all systems in the body, including hematopoiesis. Anemia is a common complication, while its degree depends on the degree of creatmemia. It is normocytic and normochromic. Anemia is caused by several factors, decreased erythropoietin synthesis being the most important. The stimulus for its secretion is tissue hypoxia. Erythropoietin synthesis is regulated by mechanism of negative retroaction and it stimulates proliferation. It also prevents apoptosis--programmed cell death. Other important factors causing anemia are uremic toxins, which decrease proliferation of erythroid progenitors, damage erythrocytes and shorten their life. Changes in the microvascular system of kidneys also cause anemia mechanically damaging erythrocytes and hemolysis. Patients with chronic renal insufficiency are susceptible to hemorrhages (erosions and duodenal ulcers) being one of the factors causing anemia. Recombinant erythropoietin production and its application in treatment of chronic renal insufficiency patients is of great importance. It is applied subcutaneously and intravenously, whereas the dosage and length of treatment are individual. CASE REPORT: A female patient, (M. R.) 69 years of age, has been treated at the Department of Internal Diseases of the Hospital in Senta 22 times during the last 30 years. Chronic renal insufficiency was caused by chronic pyelonephritis and hypertension. Up to two years ago she had several transfusions and received 7.480 ml of deplasmatic erythrocytes. The last transfusion was in May 1997, due to hemoglobin values lower than 80 g l. In August 1997, the patient acquired recombinant erythropoietin and was hospitalized and treated in our institution by subcutaneous application of erythropoietin (50 mu kg) every other day, with monitoring of hemoglobin, hematocrit, erythrocyte count increase and values of serum iron. The therapeutic response was good. 14 months later she was hospitalized again and the therapy was repeated. The last outpatient follow-up was on January 5, 1999, and hemoglobin was 92 g l. CONCLUSION: Our case report shows that recombinant erythropoietin gives good results in treatment of anemia: correction of anemia, improvement of general status and there is no need for frequent transfusions and thus there are less complications. PMID- 10518389 TI - [Recovery of facial nerve function using sural nerve transplantation after its injury in acoustic nerve surgery]. AB - INTRODUCTION: Statoacoustic n. neurinoma is a benign, slow-growing and usually unilateral tumor. During its growth the tumor exerts pressure on the surrounding anatomic forms within the pontocerebellar angle: cranial nerves, pons, cerebellum. Therefore the first clinical symptoms are ear buzzing and deafness, vision disorders, occipital headache or walking difficulties. The diagnosis of such conditions must be precise, whereas CT (computerized tomography) and MRI (magnetic resonance imaging) are the methods of choice. Surgical tumor removal is the only therapy, but during surgery facial nerve injury occurs. The objective of this paper is a case report of a facial nerve injury and sural nerve transplantation during acoustic neurinoma surgery and recovery of facial nerve function. CASE REPORT: A 23-year-old male patient suffered from ear buzzing in the right ear for a year and a half with gradual development of deafness. Due to frequent headaches and after ophthalmologic examination, the patient was urgently hospitalized at the Neurology Clinic of the Faculty of Medicine in Novi Sad where MRI of the endocranium was performed revealing a tumor of the pontocerebellar right angle, 3 x 3.5 cm in size. The patient has undergone surgery at the Neurosurgery Clinic of the Faculty of Medicine in Pees in Hungary, with suboccipital craniotomy and tumor ablation. During surgery facial nerve injury occurred in the premeatal segment and intraoperative transplantation of sural nerve grafts from the left leg to the distal parts of the facial nerve was performed. The histopathologic finding revealed an acoustic neurinoma (Schwannoma). After surgery a control CT was performed revealing a complete tumor removal. The wound healed per primam intentionem and the patient was released from hospital two weeks later. During the postoperative period physical therapy was performed in the aim of rehabilitation of the facial nerve due to peripheral paralysis. After electrodiagnostic tests using GALVOMED 20 apparatus, massage was performed in the periorbital and perioral regions. Kinesitherapy was also done in front of a mirror several times a day. 6 months after surgery an EMG (electromyography) of m. frontalis dx., m. orbicularis oculi dx. and m. orbicularis oris dx. were performed. The EMG revealed evident reinnervation possibilities. A year after surgery the control MR finding of the endocranium was regular, as well as the control MR two years after surgery (postoperative cyst without signs of recurrence of the removed neurinoma). Control EMG of the m. frontalis dx., m. orbicularis oculi dx., m. orbicularis oris dx. showed signs of reinnervation. DISCUSSION: Tumors of the pontocerebellar angle are usually acoustic nerve neurinomas. 8% of intracranial tumors are Schwannomas. They originate from neurilemmal cells, by rule they grow slowly and are benign tumors. Therefore, for years the only signs pointing to them are ear buzzing and gradual development of deafness. That is why these anamnestic data are important for diagnosis. Headaches, walking difficulties, vision disorders are the usual difficulties due to which patients seek doctor's help. Computerized tomography and magnetic resonance imaging represent the diagnostic methods of choice in establishing the diagnosis. In this case MRI was performed on time. Surgery is the only therapy, but during tumor ablation the facial nerve was injured in the premeatal region. Intraoperative transplantation of grafts taken from sural nerve to proximal and distal parts of the facial nerve provides possibilities for injured nerve regeneration. The process of regeneration of such a nerve is long term and often permanent 40% axon loss occurs. This is the reason to perform a control EMG two years after surgery. During peripheral paralysis rehabilitation is performed in the aim of preventing contractures. It is achieved by passive exercises in front of a mirror a few times a day. (ABSTRACT TRUNCATED) PMID- 10518390 TI - [Methods and sequential approaches in cardiopulmonary and cerebral resuscitation in severe illness at the place of the incident and during transportation]. AB - This paper reviews the most frequent causes of cardiopulmonary arrest and its diagnostics. The order of therapeutic procedures has been defined according to ABC system of cardiopulmonary resuscitation (CPR). Endotracheal intubation has especially been described, as one of the most important methods in the aim of providing air way as well as treatment of electrophysiologic causes of cardiopulmonary arrest. PMID- 10518391 TI - [Military hospitals in Senta (1944-1945)]. AB - Authors have reviewed data on the history of the Soviet Army Military Hospital in Senta during 1944 and activities of the Military Hospital in Senta in the period 1944-1945. The paper also contains a list of the deceased in the Military Hospital with all available data. PMID- 10518392 TI - [Rehabilitation of cardiac patients: yes or no?]. PMID- 10518393 TI - Two-stage cylindrical osseointegrated dental implants. AB - This paper presents an analysis of a 5-year clinical experience with two-stage cylindrical endosseous implants (CEI). A total of 157 implants placed in 93 patients was assessed. Clinical success was analyzed in regard to the following: sex and age distribution of patients, lower or upper jaw into which the implants were placed, prosthetic restorations with CEI or combined with implants of other types or with natural teeth. Certain groups showed a success rate from 88.9% to 100%. CEI proved to be equally good as other domestic and foreign renowned dental implants. This study is a result of a final grant-project IGA MZ CR no. 3014-3. PMID- 10518394 TI - [Comparison of the efficacy of traditional antidepressive agents and the new generation of antidepressives in the treatment of depressive disorders]. AB - INTRODUCTION: Considerable efforts have been made throughout the last 20 years to develop drugs that can replace tricyclic antidepressants as the primary treatment for depression. These efforts are well justified since tricyclic antidepressants, although therapeutically quite efficient, cause several problems, such as side effects and toxicity at therapeutic doses, causing particular problems in the elderly and in patients with congestive heart disease, and giving severe toxicity when taken in overdose. A number of compounds that are pharmacodynamically different from the tricyclics have been developed, and Selective Serotonin Reuptake Inhibitors are such. This group of antidepressives has been developed and widely marked as antidepressant therapy which offers safety and tolerability advantages over tricyclics. Tricyclic antidepressants have been the standard drug treatment for depressive illness, against which the efficacy of new compounds is compared in this investigation. MATERIAL AND METHODS: 30 female inpatients were examined. All patients had to fulfill the inclusion criteria for moderate depressive disorders, according to International Classification of Diseases. The primary efficacy parameters in the study were the changes from baseline Hamilton Rating Scale for Depression with 19 items, total score at endpoint. Patients were evaluated weekly for efficacy and adverse events. Clinicians also rated patients on the Hamilton Rating Scale for Anxiety. In statistical analysis X and T test were used. RESULTS: There was no significant difference between two groups of patients in response to tricyclic antidepressants and selective serotonin reuptake inhibitors. Selective Serotonin Reuptake Inhibitors show less adverse effects compared to tricyclics, so they appeared to be better tolerated. CONCLUSION: The traditional and new antidepressive drugs appeared to be equally effective in the treatment of moderate depressive disorders, but new drugs: Selective Serotonin Reuptake Inhibitors show less adverse effects, compared to the traditional, tricyclic antidepressants. PMID- 10518395 TI - [Modern therapy of cardiac insufficiency]. AB - Heart failure is a lethal, end-stage cardiovascular disease. Recent decrease in mortality rates from cardiovascular diseases has not been accompanied by a reduced mortality from heart failure. Survival, once the heart has used up all its reserves and compensatory mechanisms, is a little better than in cancer. That makes heart failure one of the most important world health problems. MATERIAL AND METHODS: This paper briefly reviews history, present and future of heart failure therapy, as a worldwide problem. Definition and diagnosis of heart failure, mechanism of deterioration of heart function, clinical use of present drugs in heart failure therapy and need for prevention of heart failure are, briefly pointed out. Results of clinical studies are presented, as well as the recommended indication for drug use. Heart failure is not readily identified, defined and evaluated. There is an absence of clear definition. To find a new definition which complies best with clinical practice is an important challenge cardiologists must face. Heart failure is a progressive disease and once the process has started it continues with further deterioration of cardiac function or ends in sudden death. In many patients changes within heart develop long before clinical symptoms occur. The left ventricle goes through a number of adaptations remodeling to compensate increased pressure or volume load or subsequent myocardial infarction. RESULTS AND DISCUSSION: For decades therapy has been focused on relieving symptoms, whereas preventive aspects and prolonging survival received less attention. Conventional therapy with diuretics and cardiotonic glycosides causes regression of symptoms and signs of heart failure, but there is no evidence that these drugs slow down the progression of the disease and reduce mortality. Currently, angiotensin-converting enzyme inhibitors plus diuretics are considered first line therapy for all degrees of heart failure, except for heart failure with atrial fibrillation and a rapid ventricular rate. They are, currently, the only agents with proven ability to decrease mortality. There is evidence about the efficacy of angiotensin converting enzyme inhibitors in asymptomatic left ventricular dysfunction. It is likely that not only patients with significant reduction of systolic function but also other signs of impaired left ventricular dysfunction will benefit from treatment with ACE inhibitors. However, only preventive treatment may decrease the number of patients with new onset of clinical heart failure. Therefore treatment should be introduced early, rather than waiting for heart failure to progress to a more severe stage. Based on these facts ACE inhibitors should be considered as treatment of choice, a first line therapy in most cases of heart failure. A substantial reduction in cardiovascular mortality requires detection and correction of presymptomatic left ventricular dysfunction and risk factors, which predispose to its occurrence. Major contributors to the development of cardiac failure have been delineated and quantified. Identification of high-risk individuals is difficult since signs and symptoms of heart failure are often lacking. A strategy to find these patients must use objective methods to characterize the state of the left ventricle. Despite this, methods for efficient identification presymptomatic candidates for cardiac failure for preventive measures have been developed. High-risk candidates can now be costly-effectively targeted for treatment to delay failure. Early diagnosis and subsequent aggressive medical or surgical treatment are therefore fundamental to improve adverse outcome of heart failure. It is necessary to allow a rational approach to the clinician, indicating the effectiveness of these drugs in patients with evidence of impaired ventricular function. CONCLUSION: Further reduction of morbidity and mortality from heart failure and diseases associated with heart failure is to be expected by early PMID- 10518397 TI - [Current psychological findings in essential hypertension]. AB - Many authors have investigated the role of psychic factors in the pathogenesis of idiopathic hypertension. Experimental studies have confirmed that the emotional stress induces a longterm blood pressure increase. Psychophysiologic investigations have revealed that patients with essential hypertension, when compared to the healthy population, are characterized by prompt reactions at the very initial stage of the disease, accompanied by an elevation of the biochemical and vegetative functional parameters, blood pressure in particular. It is in concordance with findings that hypertonics have hemodynamic reactions at rest quite similar to those found in normotonics under the emotional stress. Certain psychosocial factors cannot be identified as absolute causes of the disease. Their particular role varies from person to person. It depends on particular personality features whether any of these factors will be experienced as a stress. It has therefore been assumed that hypertension is a concomitant vegetative phenomenon, typical for people who have a permanent feeling of anger or for ambitious people who are under a permanent pressure of striving for success. Numerous psychological investigations have led to a conclusion that hypertonics have psychological troubles which cannot generally be considered as specific for hypertension. The question concerning the part played by such behaviour as the primary cause of the disease is still to be answered. Doubtlessly, such behaviour is frequent in hypertonics and it probably contributes to persistence of the disease, since they are more apt to stressogenic situations. These findings have resulted in formulating the therapeutic strategies with psychophysiologic orientation. At the physiological level, they resolve the increased excitement and lower the excessive reactivity of the vegetative nervous system, whereas on the psychological level, applying psychotherapy methods, they achieve both cognitive and behavioural changes. PMID- 10518396 TI - [Reactivation of herpes zoster infection by varicella-zoster virus]. AB - HISTORY: There has been considerable interest in varicella-zoster virus in the middle of the twentieth century. Virus isolation in 1958 had made it possible to find out the complete DNA sequence of the varicella-zoster virus. Molecular identify of the causative agents of varicella and shingles had been proved. ETIOPATHOGENESIS AND HISTOPATHOLOGY: Varicella-zoster virus is a member of the Herpesviridae family. After primary infection which results in varicella, the virus becomes latent in the cerebral or posterior root ganglia. Some of these individuals develop shingles after several decades because of virus reactivation. It is caused by decline of cellular immune response. Circumstances such as old age, hard work, using of steroids or malignancies contribute to the appearance of shingles. Histopathological findings include degenerative changes of epithelial cells such as ballooning, multinucleated giant cells and eosinophilic intranuclear inclusions. EPIDEMIOLOGY: Shingles occur sporadically, mainly among the elderly who have had varicella. There is no seasonal appearance of shingles. Individuals suffering from shingles may be sometimes contagious for susceptible children because of enormous amount of virus particles in vesicle fluid. CLINICAL FEATURES: Clinically, shingles is characterized at first by pain or discomfort in involved dermatome, usually without constitutional symptoms. Local edema and erythema appear before developing of rash. Maculopapular and vesicular rash evolves into crusts. The most commonly involved ganglia are: lumbar, thoracic, sacral posterior root ganglia, then geniculate ganglion of the VIIth cranial nerve and the trigeminal ganglion. The most common complication, postherpetic neuralgia, may last for as long as two or three weeks, sometimes even one year or more. Other complications that may be seen in shingles, but more rarely, are ocular (keratitis, iridocyclitis, secondary glaucoma, loss of sight), neurological (various motor neuropathies, encephalitis, Guillain-Barre syndrome), secondary bacterial infection of vesicles. Immunocompromised patients often develop more severe disease lasting up to two weeks, skin lesions are more numerous and often with hemorrhagic base and there is a high possibility for cutaneous dissemination and visceral involvement including viral pneumonia, encephalitis and hepatitis. Chronic shingles may also be found in immunocompromised hosts, particularly in those with a diagnosis of HIV infection. In patients with HIV infection, shingles is often characterised by radicular pain and itching several days before appearance of skin lesions. Those patients may have two or more dermatomes involved and recurrences of shingles cannot be quite infrequent in those patients. But visceral involvement is rarer than in other immunocompromised patients. Shingles may occur in the second half of pregnancy and usually have a mild course. However, congenital abnormalities has been described in few cases. DIAGNOSIS: The diagnosis of shingles is usually made by history and physical examination. Exceptionally, for example in zoster sine herpete and atypical forms of shingles, virus isolation and serological tests must be used. DIFFERENTIAL DIAGNOSIS: Some other diseases may cause similar skin lesions and rash (varicella, erysipelas, impetigo, enteroviral infections, herpes simplex infections). These diseases are excluded by using detailed history taking and physical examination, laboratory findings, virus isolation and commercially available serological tests. THERAPY: The vast majority of immunocompetent persons with shingles should be treated only by symptomatic therapy. Predominantly it is directed toward reduction of fever and avoiding secondary bacterial skin infection in immunocompetent hosts. Acute neuritis and post herpetic neuralgia require administration of various analgesics, even like amitriptyline hydrochloride and fluphenazine hydrochloride. Acyclovir therapy is limited to ophthal PMID- 10518398 TI - [Anemia in hypothyroidism]. AB - INTRODUCTION: Anemias are diagnosed in 20-60% patients with hypothyroidism. Real values of the degree of anemia are estimated by radioisotopic analysis due to the lower volume of plasma in hypothyroidism causing false high levels of hemoglobin in blood. Anemia is often the first sign of hypothyroidism. Diagnosis of hypothyroidism should be considered in every case of anemia with uncertain etiology because sometimes signs of overt hypothyroidism needn't necessarily be evident. Microcytic, macrocytic and normocytic are regularly described anemias. CLASSIFICATION: Microcytic anemia is usually ascribed to malabsorption of iron and loss of iron by menorrhagia. Macrocytic anemia is caused by malabsorption of vitamin B12, folic acid, pernicious anemia and inadequate nutrition. Pernicious anemia occurs 20 times more frequently in patients with hypothyroidism than generally. Macrocytosis is found in up to 55% patients with hypothyroidism and may result from the insufficiency of the thyroid hormones themselves without nutritive deficit. Normocytic anemia, so-called uncomplicated anemia, arises due to thyroid hormones deficit itself not followed by nutritive deficit. This type of anemia is considered to be an adaptation to a decreased basal metabolism. Thyroid hormones directly or indirectly, through erythropoietin, stimulate growth of erythroid colonies (BFU-E, CFU-E). Normocytic anemia is characterized by reticulopenia, hypoplasia of erythroid lineage, decreased level of erythropoietin, mainly regular erythrocyte survival. Acanthocytosis findings in cytologic blood smear suggest hypothyroidism in about 90% of cases. PMID- 10518400 TI - [Diagnosis of hearing disorders in children with early evoked auditory brainstem potentials]. AB - INTRODUCTION: Early Brainstem Evoked Response Audiometry (BERA) is a modern noninvasive, objective neurophysiological method for evaluation of hearing threshold. This method is applied in the complex otoneurological diagnostic procedure. The aim of the complete functional diagnostic procedure of hearing impairment in children is to determine the hearing level for the purpose of an early, adequate and quality hearing stimulation which affects the morphophysiological development of the complex auditory system. There are programs for early detection of the hearing impairment which give high-risk register. The registry consists of the following factors: family history of childhood hearing impairment, congenital perinatal infection, anatomical malformations involving the head or neck, birth-weight less then 1500 g, hyperbilirubinaemia, asphyxia with acidosis, usage of ototoxic drugs (high dosage of aminoglycosides). Hearing level is determinated by well-formed V wave on the lowest level of the stimulation intensity. MATERIAL AND METHODS: The paper presents an analysis of the results obtained in the period January 1992-December 1995. The diagnostic procedure was applied in 89 children ranging from 9 months to 5 years of age. The children were checked due to suspicion of hearing impairment, because of pathological speech development (undevelop speech, low level of speech development, etc.). Children were hospitalized and a complete diagnostic procedure consisted of: detailed hetero-anamnesis, otolaryngological examination, surdopaedagogical anamnesis and observation; objective audiological diagnostic procedure: BERA, tympanometry and stapedial reflex. BERA was done in a silent, dark, sound and electric-proof room using a Madsen Electronic ERA 2250. In general anaesthesia four silver disc electrodes were put to the vertex, both ear lobes and forehead of the child. Acoustic stimulation was done via earphones by monoaural stimulation by nonfiltrating click (frequency 1-4 kHz). We applied 2000 stimulations, with repetition rate 20/s. We started examination with the highest stimulation intensity--120dB, with decreasing steps of 10dB to the lowest intensity of stimulation with well-formed V wave. Tympanometry was done for evaluation of the middle ear status. The stapedial reflex was used as an indirect sign of hearing level. RESULTS: At the ENT Clinic in Novi Sad during period January 1992-December 1995 89 children ranging from 9 months to 5 years of age, 55 males (61.8%) and 34 females (38.2%) were examined. We found positive family anamnesis (deafness/severe hearing impairment) in 11 cases (12.3%). The other risk factors were found in 25 (28.1%): preterm infants 12 (48%), hypoxia and asphyxia 6 (24%), usage of the ototoxic drugs 3 (12%), hyperbilirubinaemia 2 (8%), exsanguinotransfusion 1 (4%), hydrocephalus 1 (4%). During clinical examination we have found normal otomicroscopical findings in 80 children (89.9%) and in 9 children (10.1%) tubal dysfunction was found. BERA--normal findings were in 22 children (24.7%), mild hearing impairment in 8 (9%), moderate level in 9 (10.1%), severe hearing impairment in 47 (52.8%) and deafness in 3 (3.4%). Tympanometric curve type "A" was found in 76 children (85.4%), type "C" in 9 (10.1%) and type "B" in 4 (4.5%). Stapedial reflex was not registrated in 59 children (66.3%). Habituation procedure (hearing amplification and surdopaedagogical treatment) was applied in 51 children (57.3%) e.g. 86.4% of children with hearing impairment ranging from moderate hearing level to kyphosis. DISCUSSION: Despite early beginning of the development of the complex auditory system damage, there are numerous nondetectable endogenous and exogenous factors. There are isolated hearing impairments or complex hearing impairments with damages of the other system and visible stigmata (syndrome). Well developed auditory function is a necessary precondition for speech development. (ABSTRACT TRUNCATED) PMID- 10518399 TI - [Correlation of ultrasound and radioisotope studies in subacute de Quervain thyroiditis]. AB - INTRODUCTION: The first report on ultrasonic findings in subacute thyroiditis was published in 1975 (1). More detailed studies combining ultrasonographic and clinical findings are still missing. The present paper compares echostructure of the thyroid gland with the level of hormones--triiodthyronine (T3), thyroxine (T4), thyroidstimulating hormone (TSH) and thyroglobulin (tg) and thyroglobulin autoantibodies (tg-ab). MATERIAL AND METHODS: This study included 30 patients with subacute thyroiditis de Quervain. The diagnosis was based on typical clinical symptoms, ultrasonic findings, metabolic test with J-131 uptake with thyroid scintiscan, level of hormones--T3, T4, TSH then tg and tg-ab and fine needle aspiration biopsy of thyroid gland. The patients were followed-up for 6 months, i.e. from the onset of the disease until treatment. Every month the following tests were done: ultrasound of the thyroid gland ("real time" ultrasound with a 5 MHz linear transducer), hormones T3, T4, TSH, then tg and tg at (original radioimunoassay "Vinca" and "Inep" Zemun). RESULTS: During the acute phase of subacute thyroiditis pathological findings were detected by ultrasound in every patient. The gland increased in volume (p 0.05), unclear contours, nonhomogenous echostructure demonstrates in three ways: as hypodense with "pseudocysts", as hypodense, and multiple hypodense area, 6 months later, all patients had normal echostructure of the thyroid gland. In acute phase all patients had low J-131 uptake by the thyroid. Cytological findings of all patients refer to subacute thyroiditis. In the first 6 months the difference between hormone levels and echostructure of the gland is significant (p 0.05). The difference between tg-ab and echostructure of the gland is also significant. Only in the 6th month, there is no difference between these values (p 0.05). In acute phase, after 1 month and 6 months tg levels are not related to echostructure (p 0.05). The difference is significant in the period from the 2th to 6th month. DISCUSSION: Changed ultrasonic findings in subacute thyroiditis have been described in literature. Although sonographic characteristics of the acute phase are well known, reports of long-term ultrasonic findings in subacute thyroiditis are rare (4,5). Ultrasonic findings are not uniform (6,10). They are in correlation with the form and phase of the disease. They may be related to the intensity of inflammation and edema seen in this disorder (11,12). Subacute thyroiditis is followed by hormonal dysbalance, increase of tg level and tg-ab occurrence (3). Recovery is associated with a return to normal functional and ultrasonic characteristics of the thyroid gland (2). By comparing levels of hormones, tg and tg-ab with the ultrasonic findings it can be noted that the thyroid gland has been recovering functionally more than morphologically. Subacute thyroiditis does not cause ultrasonically visible changes in parenchyma of the thyroid gland. CONCLUSION: In serial examinations no correlation was found between sonographic findings and hormone levels (T3, T4, TSH) as in sonographic findings and tg-in acute phase of disease, only in convalescence period. Correlation was found between sonographic findings and tg in acute phase and convalescence period of disease. Ultrasonography provides useful information for follow-up of the course of disease. PMID- 10518401 TI - [Histologic classification and terminology of precancerous lesions of the cervix]. AB - INTRODUCTION: Precancerous changes of the cervix frequently occur in women in their reproductive age and are associated with sexually transmitted diseases. The evolution of these lesions qualifies them as precursors of malignancy, and their origination is associate with various risk factors, human papillomavirus (HPV) being the most important one. A proper clinical approach to and treatment of these changes depend on histologic diagnosis, which must be both terminologically adequate and apprehensive. PREVIOUS CLASSIFICATIONS AND TERMINOLOGY: The continuous change in nomenclature and lack of a uniform terminology has become the source of confusion and misunderstanding between gynecologists and pathologists. The term carcinoma in situ was first introduced in 1930 to denote the lesion, which is a reliable precursor of malignancy. Less intensive epithelial changes of the cervix were classified as dysplasia. Depending on extensive the change was, dysplasias were subclassified into mild, moderate and severe. Carcinoma in situ and various degrees of dysplasias were more precisely defined at the First International Congress of Exfoliative Cytology, which has also enabled the biological differentiation between these entities. The histologic differentiation of these lesions was, however, subjective and quite unreliable. UP-TO-DATE CLASSIFICATION AND TERMINOLOGY: Cellular changes in carcinoma in situ and in severe dysplasias were mutually so similar that pathologists could not make a reproducible difference between these lesions. Therefore, a conclusion was reached that these changes were one and the same process, whereas the differences were merely of quantitative nature. This discovery resulted in a terminological change, i.e. in a unique term--cervical intraepithelial neoplasia (CIN), with its gradation from 1 to 3. The 3rd grade of cervical intraepithelial neoplasia, according to the new terminology, encompassed changes which pathologists could not properly differentiate before. Many other changes with various, mostly descriptive terminology have also been included in the CIN category, thus preventing misunderstanding between pathologists and gynecologists. Besides the CIN classification, which has been most widely used today, there is also a division into only two biologically different categories: low-grade cervical intraepithelial neoplasia (Lo-CIN) and high-grade cervical intraepithelial neoplasia (Hi-CIN). The latter modification is included in the Bethesda system of cytologic diagnoses as low-grade squamous intraepithelia lesion (L-Sil) and high-grade squamous intraepithelial lesions (H-Sil). CONCLUSION: The use of a uniform terminology and classification minimizes the problem of diagnosing precancerous cervical lesions and enables adequate clinical treatment of these patients. PMID- 10518402 TI - [Hyperprolactinemia and disorders of the menstrual cycle]. AB - INTRODUCTION: The authors presented physiological conditions associated with increased prolactin values (sleep, stress, hypoglycemia, nipples stimulation, pregnancy and lactation) as well as the causes of pathological hyperprolactinemia. MATERIAL AND METHODS: The results of radioimmunoassay study have been analyzed in concern to the values of follicle-stimulating hormone (FSH), luteinizing hormone (LH) and prolactin (PRL) obtained from the serum of 100 women with increased PRL levels. The "HOECHT" sets were used for the analysis, whereas the values up to 20.9 microgrammes per litre were rated normal. The patients were distributed into 4 groups in concern to the level of PRL increase: 1st group--twenty (n = 20) patients with PRL values from 21-29.9 micrograms/l, 90 blood samples analyzed; 2nd group-forty (n = 40) patients with PRL values f; 30-49.9 micrograms/l, 183 blood samples analyzed; 3rd group--twenty (n = 20) patients with PRL values from 50-99.9 micrograms/l, 83 blood samples analyzed; 4th group--twenty patients (n = 20) with PRL values more than 100 micrograms/l, 78 blood samples analyzed. The values of FSH and LH recorded in the women with hyperprolactinemia were compared with mean values of the same hormones presented in IU/l from the follicular phase of the cycle in the control group which comprised 50 women of reproductive age having normal ovulatory menstrual cycle. RESULTS: The mean values of FSH and LH in the 1st group have not presented with statistically significant difference in relation to the control group. Prevalence of menstrual disorders was 30%, which was statistically significantly higher than in general population. FSH values in the 2nd group were almost the same as in the control group whereas the values of LH were significantly higher. The rate of polycistic ovary syndrome (PCOS) in this group has been significant, also the increased rate of anovulatory cycles from 30 to 67.5%. A mild increase of menstrual cycle rhythm disorders, from 35 to 40% has been recorded. The values of FSH and LH in the 3rd group were significantly lower than in the control group. The significance level was higher for FSH (p < 0.01) then LH (p < 0.05). There was a sudden increase of the cycle rhythm disorders in this group reaching 90%. The 4th group presented with significantly lower values of FSH and LH in relation to the control group, whereas the cycle rhythm disorders occurred in all patients. DISCUSSION: The obtained results were compared with the literature data and some explanations given. CONCLUSIONS: The values of FSH an LH were statistically significantly lower in the 3rd and 4th group. The 2nd group was characteristic for the sudden increase of the number of anovulatory cycles from 30 to 67.5%, whereas the 3rd group presented with the abrupt increase of menstrual cycle rhythm disorders, from 40 to 90%. PMID- 10518403 TI - [Counseling protocol for early cancer detection at the Women's Health Center in Ruma]. AB - INTRODUCTION: Women's health care office in Ruma Health Center provides health care for women of Ruma and Irig communities. According to census in 1991, Ruma community has 55.063 inhabitants; 28.266 women out of which 13.149 are of reproductive age. Irig community, has 11.696 inhabitants, 6.072 women and 2.526 are of reproductive age. PROTOCOL: According to protocol each woman who consults her gynecologist, is thoroughly examined. The overall examination included the following: anamnesis in regard to risk factors for malignant diseases of female genital organs and breast, speculum examination, cytologic smear, colposcopy, palpatory examination of breast gynecological palpatory examination. On the basis of gathered results, further examinations, if necessary, were carried out: cervical biopsy, endocervical curettage, ultrasound examination, mammography and so on. Two groups of patients were formed on the basis of gathered results--high risk group and "no-risk" group of patients with no risk from malignant disease. The high-risk group of patients has a separate file and is actively controlled; if necessary this group is called for check-ups at intervals from six months to a year periods. The group of "no-risk" patients is controlled once in a three-year period. FORMING THE HIGH-RISK GROUP: High-risk group in regard to a malignant disease of the cervix is formed on the basis of the following findings: 1. Positive family history (mother, aunt, sister, etc.). 2. Positive personal history (positive sexual factor, early sexual relations, early marriage, more sexual partners, more marriages, bad sexual hygiene, human papilloma virus infection (HPV), herpes simplex virus infection, bad living and social conditions, smoking, intra-uterine and oral contraception, immune suppression, and human immune deficiency virus infection (HIV). 3. Positive clinical findings: chronic cervicitis, condylomas. 4. Positive laboratory findings: positive cytological smear, all atypical colposcopic findings, all histopathological findings of precancerous, high oncogenes groups of HPV. 5. Detected cases of malignant disease of the cervix uleri. Cervical biopsy is performed in each woman with positive cytologic and colposcopic findings. Histopathological findings are done according to Bethesda system. HPV classification according to type is done with LSIL changes, and if a high oncogenes HPV type is obtained, women require conisation of the cervix uteri. With HSIL changes, women require cervix conisation. Invasive forms of cervix uteri malignant disease, require treatment in Oncology Institute. High-risk group for endometrial and breast carcinomas is formed on the basis of positive family and personal history, positive clinical and laboratory findings. High-risk women have separate files (marked with "R") and if necessary they are asked for additional check-up. RESULTS: In the framework of the Office for early cancer detection the Cancer Registry (2,3) contains the following: personal data, address, date of disease detection, diagnosis and the disease stage, way of treatment, outcome and a questionnaire on risk factors. Morbidity of precancerous and malignant diseases of female genitalia and breast as well as mortality will be monitored in the forthcoming period. CONCLUSION: We consider the Protocol of work of the Office for early detection of cancer to be an acceptable and compulsory model for protection of women by existing public health services. 1. Each woman who comes to see her gynecologist for examination, should be systematically examined; the risk of malignant disease and necessary further examinations also must be determined. 2. An accurate file on high-risk group must be kept and women should be actively called to come for check-ups, unless they do it in an appointed period--one year at the latest. 3. Women should be kept informed, through local media, about the importance of overall examinations and risk factors for malignant diseases in women. 4. PMID- 10518404 TI - [Persistent dyschromic erythema]. AB - INTRODUCTION: Erythema dyschromicum perstans (ashy dermatosis) is a very rare skin disease included in the group of acquired, idiopathic hypermelanosis, with development of blue-gray macules. This disease appears more frequently in dark coloured persons, especially women in the first and second life decade. CASE REPORT: A male patient, 42 years of age, was admitted to Clinic of Dermatovenereology in Novi Sad due to appearance of slightly pruriginous, brown reddish macules on the trunk, upper and lower extremities, without affecting the skin of the face, scalp, palm soles and visible mucous membranes. Later, the color of the macules changed into blue-gray and new lesions appeared in axilla and flexor side of the big joints, with active, erythematous and thin raised borders. Laboratory findings showed no abnormalities; antinuclear antibodies were negative. Histopathological examination of the skin specimens (which were taken from two different places) showed vacuolar degeneration of the basal cell layer, numerous pigmentophages in papillary dermis and presence of lymphohistiocytic infiltrate in dermis. No history of drug intake or exposure to UV light was established. DISCUSSION: Ashy dermatosis is included in the group of hypermelanosis of unknown origin. As possible etiological factors we can mention ingestion of ammonium nitrate, environmental pollution, hypersensitivity to cobalt chloride and postinflammatory pigmentation. Clinical characteristics: occurrence of blue-gray and gray macules on the trunk, face, neck and extremities (absence on the palms, soles, visible mucous membranes, scalp and nails). In the active phase of the disease, these macules are surrounded by erythematous and thin, raised borders. The lesions are mostly permanent. Due to clinical, histopathological, immunofluorescent and electron microscopy established similarities with lichen planus, it is considered that ashy dermatosis is a variant of lichen planus. Absence of previous drug intake, exposure to UV light, absence of the antinuclear antibodies, clinical picture and histological findings confirm the diagnosis of erythema dyschromicum perstans. CONCLUSION: This case of ashy dermatosis shows that there is a need for differential diagnosis of acquired skin pigmentations, because this dermatosis must also be taken into consideration. PMID- 10518405 TI - [Frontal sinus osteoma as a cause of purulent meningitis]. AB - INTRODUCTION: Osteomas are benign tumours located within bones or developing on them (1). The incidence of osteomas is as follows: frontal, ethmoid and maxillary, while they are extremely rare in the sphenoid sinus (2). Most often they are localized on sutures, and extremely rarely on occipital squama (3). They are often asymptomatic, and can be accidentally detected, by radiographic examination (4). The main clinical symptom is headache of varying intensity and quality, and in most cases not proportional to the size of the osteoma, which ranges from the size of pepper bean to the size of a child's head (5). In addition to headache, there can be sensitivity to pressure in the region of the frontal sinus (6). On exteriorization they give the symptomatology of the organ on which they develop. Depending on the direction of osteoma exteriorization, various complications may occur (6,7,8,9,10). CASE DESCRIPTION: A male patient born 1958, was admitted to the Department of Infectious Diseases on Nov. 29, 1996 with high temperature and strong headaches. The patient had had a traffic accident in 1989. X-ray did not show any injuries of cranial bones, but a frontal sinus osteoma has already been diagnosed. He had been suffering from occasional headaches several years back. Symptoms of the disease started three days prior to admission, with increased body temperature and headaches which persisted in spite of prescribed analgesics. Seven days prior to onset of the disease, the patient had had a cold. When admitted, he was conscious, oriented, with well developed osteomuscular structure. He did not vomit or have photophobia; meningeal signs were negative; febrile. Laboratory blood findings were normal, except for higher sedimentation (SE + 27) and higher blood sugar (BS = 8.1 mmol/l). Due to permanent diffuse headaches a lumbar puncture was performed on Dec. 1, 1996. Laboratory and microscopic examinations showed that the patient had purulent meningitis (Table 1). X-rays of the cranium showed a frontal sinus osteoma, starting from the frontal ethmoid cells and filling the entire right frontal cavity. Electroencephalogram dated Dec. 3 shows no signs of focal or diffuse electro-cortical dysfunction. An otolaryngologist was consulted who recommended surgical extirpation of the osteoma. Axial computerized tomography of the structures of endocranium in 5 and 10 mm sections was done on an outpatient basis on Dec 17, 1996. The findings confirmed existence of frontal sinus osteoma on the right side (without lesions of the frontobasal brain structures) and calcification of fhalx cerebri. After appropriate pre-operative preparation, surgery was performed in general endotracheal anaesthesia on December 6, 1997: Trepanatio sinus frontalis sec. Tato, evacuation osteomatis et obliteratio sinus frontalis lateris dextri. It was found intraoperatively that the right frontal sinus was entirely filled with whitish-yellow osseous tissue. The osseous tumour was completely immobile in relation to the surrounding tissues and filled the entire cavity of the frontal sinus, descending into both front enthmoids towards the right posterior ethmoid and penetrating the upper orbit wall over a radius of about 5 x 10 mm. The osteoma was carefully extirpated from the location with a drill and it was found that its pressure had denuded the dura in the right region of the cranial cavity with the diameter of about 1 cm2. The osteoma was removed in three osseous fragments, total size 4 x 2.5 x 1.5 cm. Upon removal of the osteoma, the sinus walls were explored for possible fracture, due to the head injury from 1989. No signs of fracture were found. Total obliteration of the right frontal sinus was made, with a closure of the nasofrontal channel and osteoplastic reconstruction of the frontal sinus wall. The postoperative course was regular. CONCLUSION: This paper describes an osteoma of the frontal sinus in a 39-year-old patient. (ABSTRACT TRUNCATED) PMID- 10518406 TI - Intensive-care management of a patient with HELLP syndrome--case report. AB - HELLP syndrome belongs to the group of pathological states known as pregnancy induced hypertension or EPH gestosis. The basic criteria for establishing the diagnosis are as follows: H for hemolysis, EL for elevated liver enzymes and LP for low platelets. A pregnant woman, 38 years of age, multipara (V pregnancy, third delivery) has been admitted to the Clinic of Gynecology and Obstetrics in Novi Sad in 36-37 week gestation complaining of nausea, vomiting, epigastric pain, general weakness, exhaustion as well as symptom of previously diagnosed preeclampsia. Due to signs of fetal distress, the patient has undergone urgent cesarean section, giving birth to a female premature newborn infant. Twenty-four hours after delivery all symptoms and signs HELLP syndrome manifested. Being in a critical state, the patient has been transferred to the Institute of Surgery, Clinic of Anesthesiology and Intensive Care with signs of multiple organ failure. With this case report of a patient with HELLP syndrome, we wished to point to importance of continual intensive clinical follow-up, laboratory monitoring and corresponding therapeutic procedures, and at the same time to this relatively rare syndrome. PMID- 10518407 TI - [Sudden cardiac death]. AB - Sudden death occurs unexpectedly, within 24 hours after the onset of subjective symptoms with or without known preexisting conditions. According to Framingham Heart Study, during a 20-year follow-up, 13% of deceased have died of sudden death. Most frequently it occurs in the first 6 months in infants and in the period 45-75 years of age. In more than 80% of cases sudden death is caused by coronary disease, while in 5% of cases the cause is cerebrovascular insult. The mechanism of sudden death is ventricular fibrillation in 65-85%, ventricular tachycardia in 7-10% and electromechanical dissociation in 20-30%. Sometimes sudden death may be caused iatrogenically, by drug intoxication, catheterization and reflex mechanisms--vasovagal reflex and sinus caroticus reflex. Pathoanatomical finding can be positive on myocardium like fibrosis, edema, individual necrosis, cell infiltration or it can be unchanged. In cases of heart failure, resuscitation is performed: airway maintenance, artificial respiration, artificial circulation, drug therapy and electrotherapy. Prevention of sudden death means detection of high-risk patients and application of medical treatment in order to postpone it. Coronary patients with sustained myocardial infarction and ejection fraction lower than 30% and registered tachycardia represent high risk patients. PMID- 10518408 TI - [The family physician within the health care system]. PMID- 10518409 TI - Vitamin K-dependent proteins in the developing and aging nervous system. AB - Although the warfarin embryopathy syndrome, with its neurologic and bone abnormalities, has been known for decades, the role of vitamin K in the brain has not been studied systematically. Recently, it was demonstrated that vitamin K dependent carboxylase expression is temporally regulated in a tissue-specific manner with high expression in the nervous system during the early embryonic stages and with liver expression after birth and in adult animals. This finding, along with the discovery of wide distribution of the novel vitamin K-dependent growth factor, Gas6, in the central nervous system, provides compelling evidence of a biologic role of vitamin K during the development of the nervous system. In animals and bacteria, vitamin K was observed to influence the brain sulfatide concentration and the activity and synthesis of an important enzyme involved in brain sphingolipids biosynthesis. Taken together, previous research results point to a possible role of vitamin K in the nervous system, especially during its development. Hence, the knowledge of the biologic role of vitamin K in the brain may be important for unveiling the mechanisms of normal and pathologic development and aging of the nervous system. The role of the vitamin K-dependent protein Gas6 in activation of signal transduction events in the brain in light of the age-related changes in the nervous system is also discussed. PMID- 10518410 TI - Optimization of nutrition: polyphenols and vascular protection. AB - The role of polyphenols in human nutrition is discussed on the basis of their redox chemistry, which accounts for the observed antioxidant effect and in turn for their protective effect against atherosclerosis. Epidemiologic data, together with experimental pathology and cell biology, support the recommendation that optimal nutrition should contain polyphenols in amounts that may be better described as a "Recommended Optimal Intake" (ROI) than as a "Recommended Dietary Allowance" (RDA). Because a valid procedure to identify polyphenols in plasma is not available, analysis of plasma antioxidant capacity is instead suggested as a suitable index to define the optimal nutritional intake. PMID- 10518411 TI - Vitamin B12 deficiency: a new risk factor for breast cancer? AB - A prospective epidemiologic study found a threshold level for serum vitamin B12, below which an increased risk of breast cancer among postmeno-pausal women was observed. This is the first observation to suggest that B12 status may influence breast carcinogenesis and therefore may be a modifiable risk factor for breast cancer prevention. PMID- 10518412 TI - Mediterranean diet and coronary heart disease: are antioxidants critical? AB - There is substantial evidence that several variants of the Mediterranean diet reduce the incidence of coronary heart disease (CHD) and perhaps other chronic conditions. Recently, the final results of the Lyon Diet Heart Study, a randomized secondary prevention trial, indicated that the Mediterranean diet substantially reduces the rate of recurrence after a first myocardial infarction. Data from this study also suggest that the Mediterranean diet protects against CHD through mechanisms that are independent of traditional CHD risk factors. We postulate that the antioxidant properties of several plant foods in the Mediterranean diet may be critical mediators of the beneficial effects of this diet. PMID- 10518413 TI - Evidence that diet modification reduces in vivo oxidant damage. AB - Because few trials have studied the antioxidant effects of diets rather than vitamin supplements, the results of a recent trial that altered fruit, vegetable, and fat intake in healthy adults are especially valuable. The findings support the hypothesis that changing dietary patterns may decrease the risk of atherosclerosis by favorably altering the balance of oxidant defense and damage. PMID- 10518416 TI - [Progress of the laboratory supporting system of the ordering system]. AB - University hospitals and large public hospitals introduced a first-generation ordering system, which mainly involved an integrated system developed by each institution. This type of system considerably improved the efficiency of hospital jobs, but clinically increased the burden of data-input handling of hospital staffs because the software used was unique to the respective unit. Later, the development of both network technology and package software for the ordering system allowed construction of an easy, low-cost and high-performance ordering system. Most of the recent ordering systems are a type of distributed system which is referred to as a client-server system. In this system the terminal was replaced by personal computer loaded with widely distributed Windows OS, resulting in better performance of multi-tasks. In February 1998, our hospital information system was changed from an intensive host-type to a client-server system, in which the laboratory ordering system was also reconstructed. The laboratory ordering system mainly utilizes EG Main for Windows, package software by Fujitsu Co. Ltd., and has reduced the handling of laboratory ordering jobs with Graphical User Interface and better construction of screen images. In addition to extra-laboratory tests, ordering into this system allowed the database of all the laboratory tests ordered in our hospital to be unified. The previous laboratory ordering system supported laboratory data, especially those of laboratory tests and samples conducted within the last 10 years, and the new system will also provide this function. The new laboratory ordering system is further expected to support reference image-data from physiological tests as well as to allow consultation concerning laboratory test data. These clinical job supporting systems will likely lead to further progress of the total laboratory system. PMID- 10518415 TI - [Platelet activation mediated through membrane glycoproteins: involvement of tyrosine kinases]. AB - Platelet membrane glycoproteins such as GPIb and GPIa/IIa play important roles in platelet functional responses. They are the receptors for specific ligands (GPIb for von Willebrand factors, and GPIa/IIa for collagen), and the ligand-receptor interaction is the first step that elicits downstream intracellular activation signals which finally culminate in platelet aggregation. Although a variety of signal transduction pathways may be involved, tyrosine kinases appear to be most closely related to platelet activation mediated by membrane glycoproteins. Since its discovery in early 1980's, protein tyrosine phosphorylation catalyzed by tyrosine kinases has been recognized to play a role in regulating the cell function of various cells. Platelets have several Src family tyrosine kinases with an SH2 domain and an SH3 domain. Syk with two SH2 domains also appears to play an important role in platelet activation, especially in its early phase. The SH2 domain binds to a phosphorylated tyrosine residue of other proteins, and the SH3 domain recognizes proline-rich domains of target proteins, thus providing the anchoring sites for protein-protein interactions. In this article, some of the recent developments in the signal transduction pathways related with tyrosine kinases are introduced. Several signaling molecules involved in GPIb- or GPIa/IIa mediated platelet activation have been identified. Interestingly, the members participating in these processes are distinct, suggesting a diversity of signal transduction mechanisms. PMID- 10518414 TI - A case of human vitamin A deficiency caused by an inherited defect in retinol binding protein without clinical symptoms except night blindness. AB - Two German siblings were found to suffer from night blindness and mild retinal dystrophy but no other clinical symptoms of vitamin A deficiency. Even though they had no detectable plasma retinal-binding protein (RBP) and their plasma retinol was exceedingly low, they showed normal physiologic functions and growth. Their RBP gene was found to harbor two point mutations. Their post-prandial plasma levels of retinyl esters were normal, and it is likely that they derived their tissue retinol from retinyl esters. PMID- 10518417 TI - [Introduction and some problems of the rapid time series laboratory reporting system]. AB - We introduced an on-line system of biochemical, hematological, serological, urinary, bacteriological, and emergency examinations and associated office work using a client server system NEC PC-LACS based on a system consisting of concentration of outpatient blood collection, concentration of outpatient reception, and outpatient examination by reservation. Using this on-line system, results of 71 items in chemical serological, hematological, and urinary examinations are rapidly reported within 1 hour. Since the ordering system at our hospital has not been completed yet, we constructed a rapid time series reporting system in which time series data obtained on 5 serial occasions are printed on 2 sheets of A4 paper at the time of the final report. In each consultation room of the medical outpatient clinic, at the neuromedical outpatient clinic, and at the kidney center where examinations are frequently performed, terminal equipment and a printer for inquiry were established for real-time output of time series reports. Results are reported by FAX to the other outpatient clinics and wards, and subsequently, time series reports are output at the clinical laboratory department. This system allowed rapid examination, especially preconsultation examination. This system was also useful for reducing office work and effectively utilize examination data. PMID- 10518419 TI - [Acquisition of evidence and facts for a consulting service in the department of clinical laboratory to support effective usage of laboratory tests]. AB - We are developing an integrated laboratory information database to share the evidences and facts with clinicians for effective application of diagnostic laboratory tests for the care of individual patients. It includes high quality evidence acquired from the clinical trials and systematic reviews as well as basic information about the analytical method, related disorders and so on of each laboratory test. We should also have the skills to appropriately obtain information from articles about relevant subjects and obtain the original facts from the laboratory database combined with the database from the department of medical records to review diagnostic data of individual cases. These strategies to acquire and accumulate those kinds of knowledge and information are essential for the management of a consulting service room as part of the clinical laboratory department. We must contribute to the improvement of clinical diagnoses and treatments, and serve as professional consultants with the expertise. PMID- 10518418 TI - [The future guaranteeing the advanced accuracy]. AB - To guarantee the supply of accurate clinical information, the laboratory system uses external quality control, internal quality control, and data checking method. The correlation among these laboratory items and the continuity of the inter-subject variation are applied to self-acting assessment of data. In this text, we introduce other insights, not yet generalized to date, to guarantee the accuracy. First, we emphasize the importance of accurate data communication and redundancy of composition in the information equipment to improve the reliability of information transaction. Secondly, we applied the data checking system which synchronizes with pathophysiological changes in the patients to the diagnosis support system using the hypothesis deduction method. Thirdly, we present the mobility presumption method in the seat of protein electrophoresis crimp as a suitable example to obtain more accurate information processing of the original data. PMID- 10518420 TI - [Utilization of the Internet in cytology]. AB - As no standards for morphologic diagnosis exist, it is difficult to establish quality assurance. We discuss utilization of the internet in cytology, including participation in mailing lists, home pages, internet slide conference and data base use. The newest information concerning cytology can be obtained using the internet. PMID- 10518421 TI - [The future of clinical laboratory database management system]. AB - To assess the present status of the clinical laboratory database management system, the difference between the Clinical Laboratory Information System and Clinical Laboratory System was explained in this study. Although three kinds of database management systems (DBMS) were shown including the relational model, tree model and network model, the relational model was found to be the best DBMS for the clinical laboratory database based on our experience and developments of some clinical laboratory expert systems. As a future clinical laboratory database management system, the IC card system connected to an automatic chemical analyzer was proposed for personal health data management and a microscope/video system was proposed for dynamic data management of leukocytes or bacteria. PMID- 10518422 TI - [Dynamic generation of diagnostic knowledge from a database of well defined case clinical records]. AB - To promote an evidence-based practice of laboratory medicine, we created a database of 1584 typical case records consisting of 30 common diseases. Cases without therapy or major complication were carefully selected. Characteristic clinical symptoms and signs indicating the severity were recorded together with general laboratory test results. A user-friendly computer system for interpretive laboratory diagnosis was also created. It features (1) automatic graphics for visualizing distributions of laboratory values and/or prevalence of clinical symptoms and signs, (2) simultaneous display of distributions of common test values for any combination of diseases, (3) quick subclassification of cases within a disease group by any clinical characteristic, (4) diagnostic guidance by dynamic calculation of a novel similarity index that represents matching of a given set of signs/values with those of typical cases in the database. PMID- 10518423 TI - [Usefulness of a blood gas analyzer using fluorescent method]. AB - Nowadays, blood gas analysis is mainly performed by instruments using electrode methods, which have some disadvantage of instrumentation, troublesome maintenance and measuring procedures. An AVL's new blood gas analyzer works with optodes sensors based on optical fluorescence. We have studied the quality difference among cartridge lots and the effect of hemoglobin concentration on the measurements. We have also studied the relationship between actual measurements and reference values from tonometry. An AVL's new blood gas analyzer has shown good results in the tests. PMID- 10518424 TI - [Pre-beta-migrating lipoprotein A-I concentration in normal full-term maternal blood]. AB - Hyperlipidemia in pregnancy accompanying selectively-raised concentrations of serum apolipoprotein A-I (apoA-I) and high-density lipoprotein-2-cholesterol (HDL2-C) had been considered to be, at least in part, mediated by sex hormones. On the other hand, it is known that oral estrogen-replacement therapies in postmenopausal women raise serum concentrations of lipoprotein A-I without apoA II (LpA-I) originated from the liver. This study was performed to clarify the relations among the concentrations of pre-beta-migrating LpA-I (pre-betaLpA-I) and of serum apoA-I and of HDL-C. And we discussed the origin of pre-beta LpA-I in normal full-term maternal blood. Pre-beta LpA-I concentrations in 12 maternal (mean +/- SD = 33.8 +/- 7.6 mg/dl) and umbilical cord blood (mean +/- SD = 3.1 +/ 1.9 mg/dl) pairs and those in 20 healthy non-pregnant adult female controls (mean +/- SD = 13.5 +/- 6.1 mg/dl) were determined using the crossed immunoelectrophoresis. (1) The higher concentration of maternal pre-beta LpA-I than that of control was contributed to the high concentration of maternal serum apoA-I but not to the high concentration of maternal HDL-C, so it would loosen the correlativity between serum apoA-I concentration and HDL-C concentration because pre-beta LpA-I was cholesterol-poor. (2) It did not correlate with the high concentration of chylomicron, which made us speculate that maternal pre betaLpA-I might be possible to be originated from the liver by the action of endogenous estrogen. And, (3) no correlation between the concentrations of pre beta LpA-I from maternal blood and those from cord blood respectively will support the idea that their lipoprotein metabolisms would be independent with each other. PMID- 10518425 TI - [Examination of soluble cell adhesion molecules in pregnancy induced hypertension]. AB - This study was performed to determine whether or not the soluble-intercellular adhesion molecule-1 (sICAM-1), vascular cell adhesion molecule-1 (sVCAM-1) and endothelial leukocyte adhesion molecule-1 (sELAM-1) are sensitive markers of pregnancy induced hypertension (PIH). sICAM-1 concentrations were significantly higher (p < 0.05) in the mild PIH compared to non-pregnant women and normal pregnant groups. sVCAM-1 concentrations in the mild PIH group and the severe PIH group were significantly higher than the non-pregnant women group (p < 0.0001, p < 0.01, respectively) and the normal pregnant group (p < 0.0001, p < 0.0005, respectively). The concentrations of sELAM-1 in the mild PIH group were also significantly higher compared to normal pregnant group (p < 0.01). Our results suggest that soluble cell adhesion molecules may be useful markers detecting endothelial damage and dysfunction in patients with PIH. PMID- 10518426 TI - [Biological inter- and intra-individual variations of serum immunochemical constituents and their allowable limits of analytical error]. AB - The biological inter- and intra-individual variations of serum immunochemical constituents were estimated by the analysis of variance. Twelve samples taken at weekly intervals were analyzed for 22 healthy individuals. As for immunoglobulin (Ig) G, IgA, IgM, IgE, C-reactive protein, anti-streptolysin O, alpha fetoprotein, complement component 3 and 4, and ferritin, the intra-individual variations (with coefficient of variations (CVs) ranging from 4.8% to 51.7%) were smaller than the inter-individual variations (with CVs ranging from 14.6% to 107.6%). The values for carcinoembryonic antigen (with CVs of inter- and intra individual variation being 31.6% and 39.6%, respectively) and beta 2 microglobulin (with CVs of inter- and intra-individual variation being 14.4% and 13.1%, respectively) were similar. The biological variations of serum immunochemical constituents, examined in this study, were larger than those of serum chemical constituents. Based on our data, we propose allowable limits of analytical error, which is less than one-half of the average intra-individual variation for evaluation of imprecision and is less than one-quarter of the inter plus intra-individual variation for evaluation of inaccuracy. PMID- 10518427 TI - [Reduction in the window period of an acute infection by chemiluminescence enzyme immunoassay of HIV-1 p24 antigen]. AB - To evaluate the detectability of HIV-1 p24 antigen in the window period of an acute infection, 172 HIV-1-infected and 78 HIV-1-uninfected adult serum (plasma) samples were assayed by chemiluminescence enzyme immunoassay (CLEIA, 2 step sandwich method) and EIA. All 78 HIV-uninfected samples (100%) were p24 antigen negative by both methods. Thirty five of the 172 HIV-1-infected samples (32.1%) were p24 antigen negative by EIA, and p24 antigen positive by CLEIA. The HIV-1 RNA levels of the 35 discrepant samples were 8.1 x 10(3)-7.9 x 10(5) copies/ml, TH/I lymphocyte levels of them were less than 200 cells/microliter except 9 cases, and phases of them were AIDSs of CDC 1993 classification except 7 cases. The antigen detection limit of CLEIA was 3.1-6.3 pg/ml, and that of EIA was 12.5 25.0 pg/ml. The coefficient of variations of CLEIA were 3.0-6.3% determined after the assay on 5 samples on 5 different days, 6 times per day. On the basis of these results, it was inferred that the reason for the discrepancy between the two methods was due to the fact that the detection limit of CLEIA was lower than that of EIA. On an antibody-negative RNA-positive (5.0 x 10(4) copies/ml) serum sample in the window period of an acute infection, p24 antigen was detected by CLEIA 7 days earlier than by EIA. CLEIA was judged to be a useful assay for reducing the window period, and a cost-effective (effect/cost > 1.0) method. PMID- 10518428 TI - [Plasma D-dimer levels in patients with deep vein thrombosis]. AB - Hemostatic abnormalities were examined in 40 patients with deep vein thrombosis (DVT), 40 patients treated with warfarin, and 30 healthy volunteers. Plasma D dimer levels were measured by an enzyme linked fluorescent assay (ELFA; Vidas-D dimer) and an enzyme immunosorbent assay (ELISA; D-dimer-ELISA). Plasma levels of D-dimer, thrombin-antithrombin complex (TAT), soluble fibrin monomer (SFM) were significantly higher in patients with DVT than in the other groups. Both the sensitivity and specificity of D-dimer for diagnosis of DVT were the highest among hemostatic molecular markers examined. The most adequate cut off value of Vidas-D-dimer was 0.6 microgram/ml. Plasma levels of Vidas-D-dimer were well correlated with D-dimer ELISA, SFM and TAT. As the time of measurement for Vidas D-dimer is less than one hour, the measurement of D-dimer using a EFLA might be useful for the diagnosis of DVT in bed side. PMID- 10518430 TI - Cost versus care. PMID- 10518429 TI - [Non-hemophilic patient with inhibitor against factor VIII produced after the second delivery]. AB - A 33-year-old woman who had been healthy and had no history of abnormal bleeding developed widespread ecchymoses and intramuscular bleeding 4 months after her second delivery. On admission, laboratory examination data revealed that factor VIII activity was markedly reduced (4%) and APTT was prolonged (119.7s). Factor VIII inhibitor titer was high, at 19 Bethesda units. Her chemical and serological data were normal. No antinuclear antibodies were detected. We concluded that factor VIII inhibitor had been spontaneously produced after the second delivery and was responsible for her bleeding tendency. Prednisolone (60 mg/day) and factor VIII concentrates were administered to stop the bleeding, but factor VIII activity did not increase while factor VIII inhibitor titer increased to 29 Bethesda units. Therefore, treatment with factor VIII concentrate (was stopped while prednisolone was continued resulting in reduction of factor VIII inhibitor titer and improvement of her bleeding tendency. At 8 months after admission, factor VIII inhibitor titer was not detected by the Bethesda method and factor VIII activity and APTT were normal. PMID- 10518432 TI - MedBytes. PMID- 10518431 TI - Respect needs to be earned. PMID- 10518433 TI - Patients' bill of wrongs. PMID- 10518434 TI - The new disease team. PMID- 10518436 TI - Dr Sofie. Frontier surgeon blazed a trial for other women to follow. PMID- 10518435 TI - Women in medicine. PMID- 10518437 TI - Physician report cards. PMID- 10518439 TI - Why people get sick. PMID- 10518438 TI - Heart to heart. Project WATCH enters the fight against cardiovascular disease. PMID- 10518440 TI - Rates of abnormalities and infectious agents in cervical smears from female inmates in Texas: comparison with private and university clinic patients. AB - Inmates are generally considered a high-risk population for gynecologic neoplasia and sexually transmitted diseases. Cervical smears from prisoners of the Texas Department of Corrections (TDC) were expected initially to have higher rates of cellular abnormalities and infectious agents than do smears from the general population. The cytologic findings from 25,522 TDC gynecologic smears were compared with those of 6883 cases from The University of Texas Medical Branch (UTMB) affiliated physician private clinics, and with 56,178 from the UTMB hospital clinics. The period of study was from September 1995 to February 1998. This study revealed a 5.23% higher rate of abnormalities for TDC gynecologic smears as compared with that for the private clinic smears. However, the TDC rate of abnormalities was unexpectedly 1.08% lower than that for the UTMB clinic smears. These unexpected findings were probably the result of a more selected high-risk population referred to the UTMB clinics. The TDC smears showed also the highest incidence of trichomoniasis. PMID- 10518441 TI - Ethnic differences in HIV testing. AB - Testing plays an important role in the early detection of human immunodeficiency virus (HIV) infection and in the formulation of an appropriate management plan for patients who are infected with the virus. Statistical data from the City of Houston Health Department were reviewed for persons who were screened for HIV during 1996 to determine their demographic characteristics, counseling status after testing positive, and availability of medical insurance. Records of 29,085 persons were reviewed in Houston during 1996. Eight hundred eleven cases (3%) tested positive for HIV. Seventy-three percent of the HIV-positive persons received post-test counseling, and 82% of the HIV-positive persons had no health insurance. Of the total number of positive tests, 53% were African American; 28%, white; and 17%, Hispanic. Counseling after a positive test can be an important preventive measure for persons known to be at high risk for the disease. PMID- 10518442 TI - Inhibition of leukotriene formation by diethylcarbamazine modifies the acid-base balance in the rabbits with blast injuries of the lungs. AB - Our previous investigations have shown that leukotrienes are important mediators/modulators in local response of the lungs to the blast injury. The aim of the present study was to investigate the effects of diethylcarbamazine (DEC), an 5-lipoxygenase inhibitor, on the acid-base balance following pulmonary blast injury. The experiments were performed on rabbits (n = 16) subjected to focused blast over-pressure on the middle thoracic region. Immediately prior to blast injury one group was treated with DEC (50 mg/kg, i.v.), and the other with the same volume of saline. Parameters of acid-base balance were measured in arterial and venous blood before and 30 minutes after injury. Obtained results indicated that DEC treatment reduced some disturbances induced by blast injury (prevents edema formation in the lungs, permits respiratory compensation of metabolic acidosis in general circulation, normalization of respirations and slightly improves the oxygen saturation of hemoglobin), in spite of intensified hemodynamic insufficiencies associated with increased hypotension and acidosis in the peripheral circulation. PMID- 10518444 TI - [Analgesic efficacy of Famalgin and Imigran in patients with acute migraine attacks]. AB - Migraine is a syndrome clinically manifested in attacks which dominant symptom is unilateral, rarely generalized, recurrent headache that can last for hours, and occasionally, days. According to different sources, it is considered that about 10-15% of world population suffers from some type of this syndrome. Migraine is most frequently clinically revealed in the age of 30-40, therefore in the most productive period with significant share in treatment costs and a great influence to the working ability of those patients. The aim of this trial was to determine analgesic efficacy of Imigran (sumatriptan) and Famalgin in the treatment of acute attacks of migraine headache. Trial results revealed significant analgesic efficacy of both preparations without significant differences in analgesic effect, with significantly better tolerability of Famalgin. It was concluded that both preparations were efficacious drugs for the treatment of headache as the most prominent clinical symptom of migraine, with significantly rarer and less pronounced adverse effects of Famalgin. PMID- 10518443 TI - [Experimental study of the pathogenesis of frostbite. Part II. Role of the sympathetic-adrenergic system in tissue reperfusion immediately after thawing]. AB - The authors have investigated the role of sympathetic-adrenergic system in reperfusion of muscular tissue of the feet of Wistar rats immediately after thawing using alpha adrenergic blocker phentolamine methanesulfonate (Regitin). Cryoinjury was applied over an experimental model of local, controlled freezing of different intensity for certain experimental groups. Blood flow through microcirculation was measured by scintigraphy, following Tc-99m-pertechnetate clearance that was given in muscular mass of frozen right, as well as unfrozen left feet of rats. Results obtained in some experimental groups were compared mutually, as well as with the control group that was not exposed to cryoinjury. The investigations have revealed that immediately after thawing existed significant reduction, non-dependent of the intensity of freezing, in blood flow in microcirculation of both frozen and unfrozen feet of rats compared to the control group. Significant increase of microcirculatory blood flow in unfrozen feet was caused by blockade of sympathetic-adrenergic system, while the blood flow remained unchanged in the frozen ones. The reduction of blood flow in microcirculation of frozen tissues immediately after thawing was not associated with sympathetic-adrenergic factor. PMID- 10518445 TI - [Myocardial reinfarction]. AB - A total number of 1924 patients were treated for acute myocardial infarction (AMI) at the Clinic for Urgent Medicine at the Military Medical Academy during the period of seven years (1991-1997). These myocardial infarctions were at different locations, the mean age of patients was 63.7 +/- 6.2, in male patients 1192 (61.9%) and 732 (38.1%) in female ones. Out of that number of patients, 406 (21.1%) had recurrent myocard infarction (RMI), 254 (62.6%) males and 152 (37.4%) females, of average age 64.8 +/- 8.3 years. Statistically, no significant differences were observed in those two groups of patients, concerning their age, location of myocardial infarction and administration of fibrinolytic therapy. There were, however, significant differences concerning the complications, primarily cardiac insufficiency, malignant arrhythmias, AV block II0 and III0, applications of temporary or permanent pacemaker and finally mortality. During intrahospital phase of treatment, in the first few months, obtained results revealed that the patients suffering from RMI had multiple and serious complications and that cardiac insufficiency was the main cause of high mortality rate in those patients. PMID- 10518446 TI - [Amputation of the lower extremities in angiopathies and reamputation as a sequelae of postoperative complications]. AB - In the period from January 1985 till January 1995 in the Clinic of Traumatology and Orthopedics of Military Medical Academy were treated 305 patients with 317 primary amputations of the lower extremities that were performed as the sequelae of ischemia and necrosis on the basis of occlusive changes, arteriosclerosis and diabetes. Out of the total number of operated patients, 210 were with diabetes mellitus. The amputations were performed at the level of: foot in 102 (32.17%), lower leg in 145 (45.74%) and upper leg in 70 patients (22.09%). Reamputations were performed in 52 patients (16.4%): at the level of foot in 22 (6.94%), lower leg in 17 (5.36%) and upper leg in 13 patients (4.10%). Tourniquet was used during the surgery of primary amputation in 56 patients. PMID- 10518447 TI - [Arterial hypertension in an industrial population]. AB - The aim of this study was to examine the prevalence and characteristics of arterial hypertension in a group of 1519 industrial workers from the region of Nis. The prevalence of arterial hypertension was 21.3%, and was higher in the group of workers professionally exposed to noise, to lead vapour, in shift and nightly workers than in the workers in workplaces without contact to professional noxiousness. Hypercholesterolaemia and hypertrigliceridaemia were more frequently found in the hypertensive workers than in workers without hypertension. It was observed that just a low number of hypertensive workers behaved in keeping with the prescribed therapy and advices. PMID- 10518448 TI - [Antihistaminics in dermatology]. PMID- 10518449 TI - [Intravenous administration of high doses of immunoglobulins in nephrology]. PMID- 10518450 TI - [The short bowel syndrome]. PMID- 10518451 TI - [Therapeutic use of immunoglobulins]. PMID- 10518452 TI - [Botulinum toxin in the treatment of achalasia]. PMID- 10518453 TI - [Multiple sclerosis and epilepsy]. AB - Epileptic seizures in the patients with multiple sclerosis (MS) were reported more than 100 years ago. The question of mutual physiopathologic association between MS and epilepsy is even nowadays controversial. The question is if epileptic seizures are the symptoms of MS or are just coincidential, i.e., are those two separate neurological diseases? The development of contemporary immunologic, electrophysiologic and neuroradiological diagnostic procedures enabled the diagnosis of MS to be much more certain, and in that way the differentiation of some symptoms and signs, as well as epileptic seizures in those patients became more reliable. In this report are presented three patients in whom the diagnosis of MS was confirmed by clinical, laboratory, immunological, electrophysiological and neuroradiological diagnostic analyses and procedures, and in whom were simultaneously clinically and/or electrophysiologically (EEG) registered epileptic seizures with specific alterations in EEG that were obviously associated with the development of primary disease--MS. PMID- 10518454 TI - [Lateral orbitotomy in the approach to an orbital dermoid cyst]. AB - The choice of surgical approach for the resection of orbital tumor is as a rule determined by the tumor position in the orbital space and the best choice is the most direct approach to orbital tumor. Lateral orbitotomy as the extracranial orbital approach is favorable for radical resection of the tumor, which growth is limited to the anterior, lateral and superolateral orbital segment. PMID- 10518455 TI - [Lymphedema of the leg associated with rheumatoid arthritis]. AB - Only a few cases of the occurrence of chronic lymphoedema as a non-systemic, regional colagenosis associated with a real systemic disease of connective tissue, such as rheumatoid arthritis, have been reported so far. The aim of this article is to review an obridus case of chronic primary lymphoedema in a female patient suffering from rheumatoid arthritis. Reported lymphoedema appeared after many years of remission of rheumatoid arthritis and was not in correlation with its relapse. The doppler ultrasonography, lymphoscintigraphy and histopathological examination confirmed the diagnosis of primary chronic lymphoedema. The operation that consists of omentopexy (with vascularized omental flap) was the treatment of choice for this patient with favorable result. More than a year after the operation, the decreased difference in the circumference of lower limb for more than a 50% is maintained without any occurrence of new episodes of complications of lymphoedema. PMID- 10518456 TI - [Effect of chirality of ribose on nucleic acid structure and function]. AB - Organisms utilize the only D-enantiomer of ribose as a sugar unit of nucleic acids. This homochirality of nucleic acids is considered to be essential for their higher-order structures and functions. Although there had been a few reports on the synthesis of L-deoxynucleosides, effects of the substitution of L ribose for the D-enantiomer on the structure and properties of nucleic acids have hardly been investigated due to difficulties in large-scale synthesis of L deoxynucleosides. We have developed an approach for the efficient, short step synthesis of L-deoxynucleosides, and have investigated the structure and properties of oligonucleotides containing them. The double-helical conformations of an L-hexadeoxynucleotide, L-d(CGCGCG) were clearly shown to be an exact mirror image of those of the corresponding natural one by CD (circular dichroism) and X ray crystallographic analysis. This L-hexadeoxynucleotide was applied to the study of the specific double-stranded DNA recognition mechanism of bleomycin. It was found that the conventional DNA-binding domain of bleomycin binds to the L hexadeoxynucleotide to essentially the same extent as natural one, although bleomycin can not cleave it. This result suggests that the DNA-binding domain is not responsible for the specific DNA recognition. Thus, L-DNA would be useful for distinguishing between enantio-specific and nonspecific interactions in DNA-drug interaction studies. The structures of heterochiral oligonucleotides, which contain an unnatural L-nucleotide residue in the sequence of natural type of DNA chains, were investigated by UV and 1H-NMR experiments. The results demonstrated that the L-nucleotide residue in the heterochiral oligonucleotide forms stable Watson-Crick base-pairing with the complementary natural residue, while the overall duplex stability is slightly decreased. The unusual conformational features of the L-nucleotide residue in the heterochiral DNA may be useful for the design of a novel antisense molecule resistant to nucleases in vivo. PMID- 10518457 TI - [Development and assessment of the pharmaceutical management and guidance services for inpatients considering clinical value and economy]. AB - We developed pharmaceutical management and guidance services for inpatients in a ward of circulatory medicine, considering clinical and economical standpoints. In these services, pharmacists deliver drugs prescribed for inpatients with individual drug information papers, explain to them about their drugs using information papers and give counsel. Since most of the patients were aged people, developing many kinds of diseases and taking many kinds of drugs, they had many problems such as lack of knowledge of the effects of drugs. First, we surveyed views of patients, physicians and nurses on these services. Consequently, all of them advised us that "pharmacists should explain to patients about the prescribed drugs using information papers." The patients preferred pharmacists as expositors of drugs to physicians or nurses. The physicians considered that "pharmacists have to attach importance to clinical information and package-inserts of drugs and explain to patients about drug information using pamphlet in response to the understanding of patients." The nurses wanted to cooperate with pharmacists in "improving medication compliance." On the basis of these views, we improved our services. Next, we made a survey of patients' knowledge about their drugs. We found that in the patients the level of knowledge concerning "ways," "effects" and "reasons" of taking drugs and that of "compliance" and "satisfaction in taking drugs" were improved through these services. The patients reentered in the hospital kept a high level. The ratio of patients taking drugs by themselves increased. Last, we also applied this method to wards of "blood and collagen diseases" and "pediatrics." The demand for these services increased smoothly. We compared these services based on our method with all other services by hospital pharmacists from the viewpoint of economy. We found that only our service method was beneficial. PMID- 10518458 TI - [Pharmacological studies of Reiousan, a drug containing bezoar and ginseng- effects on the blood rheology]. AB - Effects of Reiousan, a crude drug preparation consisting of bezoar and ginseng, on blood rheology were studied. Reiousan improved the deformability of rat erythrocytes exposed to hyperosmorality and treated with phenylhydrazine. The ATP depletion in erythrocytes, the polybrene-induced erythrocyte aggregation and the oxidation of low density lipoprotein were suppressed by Reiousan. Oral administration of Reiousan also improved the erythrocyte deformability in phenylhydrazine-treated rats and delayed the thromboembolic death induced by arachidonic acid in mice. These results suggest that Reiousan has an ameliorative effect on blood rheology related to "Oketsu" syndrome in Kampo diagnostics. PMID- 10518459 TI - [Effect of separation characteristics between salbutamol sulfate (SS) particles and model carrier excipients on dry powder for inhalation]. AB - Most often dry powder for inhalation are formulated as ordered mixtures of a carrier excipient and a micronized drug substance. In the present study, model powder blends were prepared from a mixture of lactose alpha-monohydrate, micro crystalline cellulose pellets or synthesized sugar as carrier particles, and micronized salbutamol sulfate (SS). These ordered mixtures were aerosolized by the multidose JAGO dry powder inhaler (DPI) and their in vitro deposition properties were evaluated by a twin impinger (TI). The separation force between SS particles and carrier particles was investigated by the centrifuge method. In addition, the use of the air jet sieve (AJS) method was investigated to assess the separation behavior of drug particles from carrier excipient. Powder blends were sieved through a 325 mesh wire screen of an air jet sieve at an air pressure of 1500 Pa. The amount of drug deposited at the carrier surface was analysed before and after the sieving to calculate the percentage of the drug retained. A relationship was found between in vitro deposition properties (fine particle fraction, FPF) and the separation characteristics obtained by the centrifuge method and by the AJS method. The AJS method might be a suitable alternative for evaluating separation of a drug particle from carrier particles and hence can be used for the formulation screening of the dry powder inhalation. PMID- 10518460 TI - [Analysis of micronuclei induced under hyperthermic conditions in human lymphocyte culture by fluorescence in situ hybridization (FISH) and spectral karyotyping (SKY) methods]. AB - The spectral karyotyping (SKY) method is a novel molecular cytogenetic technique which simultaneously discerns entire chromosomes. In order to elucidate the origins of micronuclei induced under hyperthermic conditions in human lymphocyte culture, peripheral blood cells were cultured at 40 degrees C or 42 degrees C for 3-24 h, using the cytokinesis-block method with cytochalasin B. The induced micronuclei were identified by the fluorescence in situ hybridization (FISH) and SKY methods. At 42 degrees C for more than 6 h, the frequency of occurrence of micronuclei in binucleated cells rose with increasing incubation time. By the FISH method, 83.3% of micronuclei induced in 24 h culture at 42 degrees C were shown to be positive for the human centromeric probes. By the SKY method, each micronucleus induced under the hyperthermic conditions was identified unequivocally and shown to contain a specific chromosome. These results suggest that the micronuclei induced under the hyperthermic conditions in human lymphocyte culture contain chromosomes which do not migrate to the poles at the anaphase of the cell cycle because of the breakdown of the spindle apparatus. PMID- 10518461 TI - [Evaluation of dissolution behavior for the products containing tegafur and uracil]. AB - Generic drugs are widely used with the object of cost saving in many Japanese therapeutic scenes now. The products containing the same active ingredient(s), even if they are innovative drugs or generic ones, must be designed to possess the equivalent quality. In this report, we observed the dissolution behavior patterns of three generic drugs that contain Tegafur and Uracil, drugs A, B, and C, and compared them with that of an innovative product, UFT. Drugs B and C were similar to UFT in the dissolution rate of Tegafur, but drug A was not. On the dissolution rate of Uracil, all the generic products, drugs A, B and C, did not amount to the level equivalent to that of UFT. Therefore, these generic products did not indicate the same dissolution behavior pattern as UFT. It was suggested that the pharmaceutical technology used in the manufacture was not equivalent even if the products of the same dosage form contain the same kind and content of the active ingredient(s). PMID- 10518462 TI - [Biomimetic oxidation of diphenyl sulfide with electrochemical P-450 model system in CH2Cl2 treated with alkaline solution]. AB - Dichloromethane containing metalloporphyrins [meso tetraphenylporphyrinatomanganese(III) chloride (1) or meso tetraphenylporphyrinatoiron(III) chloride (2)] and Bu4NClO4 was treated with an aqueous solution of NaOH (5%), and subjected to controlled potential electrolysis at -1.0 V (vs. S.C.E. (saturated calomel electrode)) in a divided cell after addition of diphenyl sulfide (3). Diphenyl sulfoxide (4) and diphenyl sulfone (5) were found in an electrolyzed solution as the reaction products. Results obtained from cyclic voltammetry and visible spectrometry suggested that the treatment of dichloromethane containing metalloporphyrins with the aqueous solution of NaOH did not change the fifth ligand of metalloporphyrins from Cl to OH. On the electrode, dissolved dioxygen was reduced to hydrogen peroxide. Compounds 1 and 2 catalyze the oxidation of 3 by hydrogen peroxide without imidazole. Compound 2 showed higher selectivity than compound 1. PMID- 10518463 TI - Retrospective exposure assessment in large occupational cohort studies in China: advantages and concerns. PMID- 10518464 TI - A field comparison of inhalable and thoracic size selective sampling techniques. AB - We measured inhalable, thoracic, and so-called "total" wood dust exposure in British Columbia lumber mill workers. Particle-size selective sampling was conducted using the GSP and Seven hole inhalable samplers, the PEM thoracic sampler and the 37-mm closed-face cassette "total" sampler. All measurements were full-shift personal samples, obtained from randomly selected workers. We obtained intersampler comparison data for the following pairs of instruments: GSP and 37 mm sampler; GSP and seven-hole sampler (SHS); and PEM and 37-mm sampler. The intersampler measurement ratios were estimated as: GSP/37-mm sampler = 4.2; GSP/SHS = 1.7; and PEM/37-mm sampler = 1.6. The GSP/37-mm sampler ratio is consistent with previously reported findings, while PEM/37-mm sampler and GSP/SHS ratios were both larger than expected. We found that in all comparisons, the measurement ratio had significant variability that was greatest at low ambient dust concentrations. Although it was not possible to attribute the source of the variability to specific sampler types, we concluded that the GSP sampler might be susceptible to "projectile" particles not normally aspirated, and may be vulnerable to direct aspiration of dust from accidentally contacted surfaces. The PEM was designed for environmental monitoring, and it is possible that it is unsuited to the higher particulate concentrations found in some occupational settings. Disparities among inhalable sampling techniques such as that between GSP and SHS should be investigated further in light of the proposed adoption of the inhalable method as an industrial standard. PMID- 10518465 TI - Field testing of a personal size-selective bioaerosol sampler. AB - Existing samplers for the collection of bioaerosols have been designed with the aim of maintaining biological stability of the collected material, and in general do not select particles in accordance with international conventions for aerosol sampling. Many have uncharacterised sampling efficiencies and few are designed as personal samplers. If standard personal dust samplers are used for bioaerosols the viability of collected microorganisms may be compromised by dehydration. The objective of this study was to evaluate a novel personal bioaerosol sampler designed to collect the inhalable dust fraction and further subdivide the sample into thoracic and respirable fractions. The new sampler was tested to see whether it enhanced the survival of the collected microorganisms, and was assessed for ease of use in the field and in subsequent laboratory analyses. A number of occupation-related field sites were selected where large concentrations of bioaerosols were to be expected. The prototype sampler was found to be simple to use. Analysis could be carried out with similar efficiency either with all three fractions together for a total count, or separately for size selective data. The sampler performed at least as well as the standard IOM filter method but with the added advantage of size fractionation. The field trials showed that for sampling periods lasting several hours, microorganism survival within the sampler was adequate for culture and identification of the organisms present. This new sampler is now commercially available. In addition to bioaerosol sampling, the principle of size selective sampling using porous foams can be applied to other occupational hygiene problems, and also to indoor air monitoring of PM10 and PM2.5 concentrations. PMID- 10518466 TI - In vitro test of nicotine's permeability through human skin. Risk evaluation and safety aspects. AB - Permeability tests with Franz' diffusion cells and an in vitro test model were made to evaluate the importance of dermal absorption of nicotine as a pathway for intoxication. Studies were carried out to ensure that safety procedures, when spilling nicotine on skin, are sufficient to prevent poisoning. Pure nicotine and nicotine in various concentrations in water or ethanol were applied on human skin or gloves in Franz' cells. Washing was simulated by removing nicotine from skin after 3 or 5 min. Permeation rate (flux) and lag time were calculated and estimated for human skin. Different glove materials were tested for their nicotine breakthrough time. Flux depended on concentration in a non-linear way when nicotine-water solutions were tested. Highest flux was found in 50% w/w nicotine dissolved in water. Solutions with low concentration of nicotine (1% w/w) dissolved in water had a similar permeation rate to 100% nicotine. Flux was found to be low when using ethanol as a vehicle; flux was also pH-dependent. The nicotine-water solution containing acetic acid had the lowest flux. The tests where nicotine was washed away revealed that skin served as a possible nicotine depot, because nicotine concentration in the receptor compartment continued to increase after removing the nicotine from the surface. The length of contact time affected the amount of substance passing the skin, resulting in great difference between 3 and 5 min contact time, 5 min giving higher nicotine concentration and 3 min lower. This emphasizes the importance of washing away nicotine spilled on skin rapidly. Two glove types were tested and they were found to be appropriate in their use with nicotine if changed regularly. PMID- 10518467 TI - Exposure to MDI during the process of insulating buildings with sprayed polyurethane foam. AB - Buildings are often insulated with sprayed-in-place polyurethane foam in spite of the fact that few studies have been carried out on exposure levels to isocyanates during the spraying process. This paper is meant to provide new data on personal exposure to methylene-bis (4-phenylisocyanate) (MDI) while dwellings and office buildings are being insulated with polyurethane foam. An impinger using a 1-(2 methoxyphenyl)piperazine toluene solution as absorbent was used to take personal samples for the sprayer and helper during indoor and outdoor applications. The analytical results show that the levels of exposure were significant, especially for the sprayer, with values of up to 0.077 mg m-3 and 0.400 mg m-3 during outdoor and indoor applications, respectively. The helper's exposure was always lower. PMID- 10518468 TI - Distribution and skewness of occupational exposure sets of measurements in the Norwegian industry. AB - Aggregated occupational sets of exposure measurements from the Norwegian industry registered in the exposure database EXPO at The National Institute of Occupational Health, Oslo were examined with respect to distributions and skewness. Data for lead in blood show a truncated almost normal distribution because of regulations for workers with high lead in blood concentrations. The styrene, dichloromethane and acetone measurements show quasi log-normal distributions possibly because of over-representation of worst-case measurements. The other personal and stationary measurements are relatively good fitted to a log-normal model. The stationary measurements indicate generally lower mean levels than the corresponding personal measurements. The statistical parameter skewness is valuable in connection with an exposure database as a distribution test for raw data and log-transformed data. PMID- 10518469 TI - Substitution of organic solvent cleaning agents in the lithographic printing industry: comment. PMID- 10518470 TI - Association of IgA nephropathy with Clostridium difficile colitis. AB - Immunoglobulin A (IgA) nephropathy, the most common cause of glomerulonephritis worldwide, is usually idiopathic in origin and renal limited. Secondary IgA nephropathy has been associated with systemic disease, including such gastrointestinal tract disturbances as celiac sprue and inflammatory bowel disease. We describe gross hematuria and reversible acute renal failure from IgA nephropathy in a patient with cephalosporin-induced Clostridium difficile colitis. In addition to mesangial IgA and C3 deposition, renal histological examination showed glomerular bleeding, intratubular red blood cell casts, and acute tubular necrosis. To the best of our knowledge, this is the first report of an association between IgA nephropathy and C difficile colitis. PMID- 10518471 TI - Genes to remember. AB - It has been known for several decades that the formation of long-term memory requires gene expression. In recent years, the use of genetic and molecular approaches has led to the identification and characterization of genes and molecules that play a fundamental role in the biological mechanisms underlying learning and memory. From these studies, it appears that molecules and molecular mechanisms essential for the process of memory have been conserved throughout evolution. The cyclic AMP (cAMP)-dependent activation pathway and a cAMP dependent cascade of gene expression have been shown to be essential for memory formation in Aplysia californica, Drosophila melanogaster and rodents. Moreover, members of the transcription factor family cAMP response element binding proteins (CREBs) seem to represent key molecules for transforming incoming information into long-term memory. Here, we review the studies showing that conserved molecules and biological mechanisms are engaged in simple and complex forms of memory. PMID- 10518472 TI - Directional acoustic radiation in the strut display of male sage grouse Centrocercus urophasianus. AB - We present evidence that the acoustic component of the strut display of male sage grouse Centrocercus urophasianus is highly directional and that the nature of this directionality is unique among measured vertebrates. Where vertebrate acoustic signals have been found to be directional, they are most intense anteriorly and are bilaterally symmetrical. Our results show that sage grouse acoustic radiation (beam) patterns are often asymmetric about the birds' anterior posterior axis. The beam pattern of the 'whistle' note is actually strikingly bilobate with a deep null directly in front of the displaying bird. While the sage grouse display serves to attract potential mates, male sage grouse rarely face females head on when they call. The results of this study suggest that males may reach females with a high-intensity signal despite their preference for an oblique display posture relative to those females. We characterized these patterns using a novel technique that allowed us to map acoustic radiation patterns of unrestrained animals calling in the wild. Using an eight-microphone array, our technique integrates acoustic localization with synchronous pressure field measurements while controlling for small-scale environmental variation in sound propagation. PMID- 10518473 TI - Maps of the somata of efferent neurones with axons in the lateral nerves of locust abdominal ganglia. AB - We used the cobalt-backfilling method to map the somata of neurones with axons that project in the two paired lateral nerves of the abdominal neuromeres of the locust Schistocerca gregaria with the objective of expanding and bringing together the incomplete and scattered information on these efferent neurones. We compared somata sizes and positions, and the pathways of primary neurites, with information from previous studies on individual, or groups of, abdominal neurones and we identify many of the somata we mapped. The stained somata belong to paired motor neurones and paired neurosecretory neurones, to unpaired neuromodulatory neurones (dorsal unpaired median, DUM, neurones) and unpaired bilaterally projecting neurones. In different neuromeres, the total number of somata with axons in these lateral nerves ranges from 73 to 106. Within an individual segmental neuromere, approximately 25 % of the somata belong to neurones with axons in nerve 1 (N1) and 35 % to those with axons in nerve 2 (N2) of that segment, while the remaining 40 % belong to neurones with axons in N1 of the next posterior segment. This basic pattern is repeated in all abdominal neuromeres, with differences in the percentages depending on whether the neuromeres are pregenital fused, pregenital unfused or genital. Nerve 1 contains the axons of 26 37 neurones with central somata in different neuromeres, of which 40 % are in the segmental neuromere and 60 % in the next anterior neuromere. In the segmental neuromere, 15 % of somata are ipsilateral to the nerve, 30 % are at the midline and 55 % are contralateral, whereas in the next anterior neuromere, 70 % are ipsilateral, 10 % are at the midline and 20 % are contralateral. Nerve 2 contains the axons of 11-28 neurones in different neuromeres, all of which have somata in the same segmental neuromere from which the nerve projects. Of these, approximately 70 % are ipsilateral, 30 % at the midline and none contralateral, except for the first abdominal and eighth male abdominal neuromeres, where one and two somata, respectively, are contralateral. PMID- 10518474 TI - Evolutionary and acclimation-induced variation in the heat-shock responses of congeneric marine snails (genus Tegula) from different thermal habitats: implications for limits of thermotolerance and biogeography. AB - Heat stress sufficient to cause cellular damage triggers the heat-shock response, the enhanced expression of a group of molecular chaperones called heat-shock proteins (hsps). We compared the heat-shock responses of four species of marine snails of the genus Tegula that occupy thermal niches differing in absolute temperature and range of temperature. We examined the effects of short-term heat stress and thermal acclimation on the synthesis of hsps of size classes 90, 77, 70 and 38 kDa by measuring incorporation of (35)S-labeled methionine and cysteine into newly synthesized proteins in gill tissue. Temperatures at which enhanced synthesis of hsps first occurred (T(on)), temperatures of maximal induction of hsp synthesis (T(peak)) and temperatures at which hsp synthesis was heat inactivated (T(off)) were lowest in two low-intertidal to subtidal species from the temperate zone, T. brunnea and T. montereyi, intermediate in a mid- to low intertidal species of the temperate zone, T. funebralis, and highest in a subtropical intertidal species from the Gulf of California, T. rugosa. Synthesis of hsps and other classes of protein by T. brunnea and T. montereyi was heat inactivated at temperatures commonly encountered by T. funebralis during low tides on warm days. In turn, protein synthesis by T. funebralis was blocked at the upper temperatures of the habitat of T. rugosa. Acclimation of snails to 13 degrees C, 18 degrees C and 23 degrees C shifted T(on) and T(peak) for certain hsps, but did not affect T(off). The heat-shock responses of field-acclimatized snails were generally reduced in comparison with those of laboratory-acclimated snails. Overall, despite the occurrence of acclimatory plasticity in their heat shock responses, genetically fixed differences in T(on), T(peak) and T(off) appear to exist that reflect the separate evolutionary histories of these species and may play important roles in setting their thermal tolerance limits and, thereby, their biogeographic distribution patterns. PMID- 10518475 TI - Sound production by abdominal tymbal organs in two moth species: the green silver line and the scarce silver-line (Noctuoidea: Nolidae: Chloephorinae). AB - Male moths of the chloephorine species Pseudoips prasinana and Bena bicolorana produce clicks (approximately 100 dB peSPL at 10 cm) using ventral tymbal organs located in a cleft in the second abdominal sternite. Large muscles insert on the dorsal part of the tymbal frame and rhythmically flex a thin sheet of cuticle. Normally, each sound-production cycle contains four clicks, the left and right tymbals producing clicks both on active buckling caused by muscle contraction and on the passive elastic return from buckling. Histochemical staining indicated the presence of elastic resilin-like proteins in the tymbals. Obvious differences between the click patterns of the two species reflect differences in their tymbal morphology. P. prasinana has smooth tymbals and produces a single click (300 us, 40 kHz) for each tymbal buckling. In contrast, B. bicolorana has striae on the medial part of the tymbals. Accordingly, it produces many clicks per buckling. The click pattern is a heterogeneous mixture of large clicks at 52 kHz, resembling those of P. prasinana, interspersed with series of broad-band clicks (20-100 kHz) of lower intensity (15-20 dB). Thus, in chloephorine moths, there is a correlation between the structure and function of the smooth and striated tymbals that is strikingly similar to that in arctiid moths, although the two types of tymbals have evolved independently. The hearing of P. prasinana is tuned to its own sounds with lowest threshold (38 dB SPL) at 40-60 kHz. We suggest that sound production in male chloephorines plays a part in sexual acoustic communication. PMID- 10518476 TI - Colour vision of domestic chicks. AB - The colour vision of domestic chicks (Gallus gallus) was investigated by training them to small food containers decorated with tilings of grey and coloured rectangles. Chicks learn to recognise the colour quickly and accurately. Chicks have four types of single-cone photoreceptor sensitive to ultraviolet, short-, medium- or long-wavelength light. To establish how these receptors are used for colour vision, stimuli were designed to be distinguished only by specific combinations of receptors. We infer (1) that all four single cones are used, and (2) that their outputs are encoded by at least three opponency mechanisms: one comparing the outputs of ultraviolet- and short-wavelength-sensitive receptors, one comparing the outputs of medium- and long-wavelength receptors and a third comparing of the outputs of short- and long- and/or medium-wavelength receptors. Thus, the chicks have tetrachromatic colour vision. These experiments do not exclude a role for the fifth cone type, double cones, but other evidence suggests that these cones serve luminance-based tasks, such as motion detection, and not colour recognition. PMID- 10518477 TI - Egg-laying hormone peptides in the aplysiidae family. AB - The neuropeptidergic bag cells of the marine mollusc Aplysia californica are involved in the egg-laying behavior of the animal. These neurosecretory cells synthesize an egg-laying hormone (ELH) precursor protein, yielding multiple bioactive peptides, including ELH, several bag cell peptides (BCP) and acidic peptide (AP). While immunohistochemical studies have involved a number of species, homologous peptides have been biochemically characterized in relatively few Aplysiidae species. In this study, a combination of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MS) and electrospray ionization Fourier transform ion cyclotron resonance MS is used to characterize and compare the ELH peptides from related opisthobranch molluscs including Aplysia vaccaria and Phyllaplysia taylori. The peptide profiles of bag cells from these two Aplysiidae species are similar to that of A. californica bag cells. In an effort to characterize further several of these peptides, peptides from multiple groups of cells of each species were extracted, and microbore liquid chromatography was used to separate and isolate them. Several MS-based sequencing approaches are applied to obtain the primary structures of bag cell peptides and ELH. Our studies reveal that (&agr;)-BCPs are 100 % conserved across all species studied. In addition, the complete sequences of (&egr;)-BCP and ELH of A. vaccaria were determined. They show a high degree of homology to their counterparts in A. californica, with only a few amino acid residue substitutions. PMID- 10518478 TI - Processing of proprioceptive signals by ascending interneurones in the terminal abdominal ganglion of the crayfish. AB - Intersegmental interneurones are crucial for the appropriate coordination of the activity of local circuits located in different body segments. We have analysed the synaptic inputs to ascending intersegmental interneurones from a proprioceptor in the tailfan of the crayfish. Twenty identified interneurones responded during stimulation of the exopodite-endopodite chordotonal organ. Of these 20 interneurones, three were excited phaso-tonically, nine were excited phasically and eight were inhibited. All received convergent exteroceptive inputs from water-motion- or touch-sensitive hairs on the uropods. The effects of simultaneous exteroceptive and proprioceptive stimulation depended upon the identity of an interneurone. For interneurones that were inhibited by proprioceptive stimulation, suprathreshold exteroceptive responses were reduced to a subthreshold level by simultaneous proprioceptive stimulation. In contrast, for interneurones that were excited by proprioceptive stimulation, the simultaneous application of subthreshold proprioceptive and exteroceptive stimulation elicited action potentials. Two of the interneurones that receive proprioceptive input (NE-1 and RC-8) are known to be presynaptic to giant interneurones that mediate and coordinate the tail-flip. Many of the other interneurones that receive proprioceptive inputs in the tailfan are known to excite abdominal extensor motor neurones. Thus, proprioceptive input to these intersegmental interneurones could serve two roles: first, to extend the abdomen during postural movements or prior to escape and, second, to drive the tail-flip escape response. PMID- 10518479 TI - Placental nutrition in the viviparous lizard Niveoscincus metallicus: the influence of placental type. AB - The ion, energy, lipid, nitrogen and fat-soluble vitamin contents of freshly ovulated eggs and neonates of the viviparous lizard Niveoscincus metallicus were measured to quantify uptake of nutrients across the placenta. This species is particularly interesting because it has a chorio-allantoic placenta that is intermediate in complexity compared to viviparous species that have been the focus of other studies. Newly ovulated eggs have a wet mass of 79.6+/-4.6 mg and a dry mass of 41.8+/-2.8 mg, compared to the neonates that have a wet mass of 224.2+/-8.2 mg and dry mass of 37.9+/-1.2 mg. Thus, there is no significant net uptake of dry matter across the placenta. Neonates have significantly less lipid (6.2+/-0.4 mg) than eggs (12.7+/-0.5 mg), but no significant difference in nitrogen (4.1+/-0.3 mg) compared to eggs (4.5+/-0.2 mg). Energy densities reflect the protein and lipid composition and the relative dry masses of the eggs and neonates. There is significantly more energy (1029.1+/-80.0 J) in the egg than in the neonate (858.2+/-38.6 J). The increase in the ash content of the neonates (2.9+/-0.2 mg) compared to fresh eggs (2.1+/-0.3 mg) was not significant, even though there was an approximately threefold increase in the amount of sodium (0.11+/-0.01 mg in neonates, 0.34+/-0.01 mg in eggs) and potassium (0.12+/-0.017 in neonates, 0.40+/-0.01 mg in eggs) in neonates compared to eggs. There was no significant uptake of calcium and magnesium during development. The egg lipids consisted of triacylglycerol (66.7+/-2.3 %), phospholipid (18.9+/-0.7 %), cholesteryl ester (4.9+/-1.6 %) and free cholesterol (5.6+/-1.5 %). The egg phospholipid contained comparatively high proportions of arachidonic and eicosapentanoic acids but low levels of docosahexaenoic acid (DHA), whereas the phospholipid of the neonate was greatly enriched in DHA. In the egg, the predominant vitamin E was (&agr;)-tocopherol (62.6+/-3.4 mg g(-)(1)), although there was some (&ggr;)-tocotrienol (3.5+/-0.3 mg g(-)(1)), and vitamin A was present (1.5+/-0.2 mg g(-)(1)). The ratio of neonate dry mass to egg dry mass of N. metallicus (0.91) lies between that of species with type I (0.78) and type III (1.70) chorio-allantoic placentae, confirming our conclusion that the placenta of N. metallicus is functionally intermediate, as well as intermediate in complexity, between these other two types. PMID- 10518480 TI - The effects of hypercapnia on force and rate of contraction and intracellular pH of perfused ventricles from the land snail Helix lucorum (L.). AB - The effects of hypercapnia, together with low and high levels of extracellular Ca(2+), on heart activity and intracellular pH were examined in isolated perfused hearts from the land snail Helix lucorum. In addition, the intracellular level of Ca(2+) was determined in slices of ventricles superfused with both normal and hypercapnic salines, containing low and high concentrations of Ca(2+), to investigate whether low extracellular pH affects the entry of Ca(2+) into the heart cells. We also examined the effect of a saline that simulated the composition of the haemolymph of snails after estivating for 3 months on the heart activity and intracellular pH. The results showed that hypercapnia causes decreases in the rate and force of heart contraction, and these are more pronounced in the presence of low levels of extracellular Ca(2+). Moreover, the present results indicate that Ca(2+) maintains the contractility of the heart muscle under acidic conditions and seems to act by competing with protons for the Ca(2+ )binding sites on sarcolemma. The negative effect of hypercapnia on heart activity appears to be due to a reduction in extracellular pH rather than to changes in intracellular pH. PMID- 10518481 TI - Effects of zinc on l- AB - Epithelial brush-border membrane vesicles (BBMVs) from the hepatopancreas of the lobster Homarus americanus were prepared using a magnesium precipitation technique and employed in transport experiments designed to demonstrate the effects of external and internal divalent cationic heavy metals on the uptake of l-[(3)H]proline. When BBMVs were exposed to a high external concentration (2.5 mmol l(-)(1)) of Cd(2+), Cu(2+), Fe(2+), Mn(2+) or Zn(2+), l-[(3)H]proline (0.5 mmol l(-)(1)) uptake was significantly (P<0.05) decreased by each metal. However, if a 30 min pre-incubation period with each metal was used before incubation of the vesicles with amino acid and metal, a significant (P<0.05) enhancement of l [(3)H]proline transport occurred. Zinc was the most stimulatory metal of those tested. Proline influxes (1.0 and 2.5 mmol l(-)(1)) were hyperbolic functions of bilateral [Zn(2+)], with a lower apparent zinc half-saturation constant (K(m)) at the higher amino acid concentration. l-[(3)H]proline influx was a hyperbolic function of external [l-proline] (K(m)=2.10+/-0.26 mmol l(-)(1); J(max)=2290+/ 600 pmol mg(-)(1 )protein 10 s(-)(1)) (means +/- s.e.m., N=3), and bilateral exposure to zinc significantly (P<0.05) increased the maximal rate of influx, J(max), of proline (J(max)=4890+/-250 pmol mg(-)(1 )protein 10 s(-)(1)), but had no effect (P>0.05) on apparent l-[(3)H]proline binding to the membranes (K(m)=1.66+/-0.23 mmol l(-)(1)) (means +/- s.e.m., N=3). In the presence of 0.5 mmol l(-)(1 )l-pipecolate, bilateral zinc-stimulated, carrier-mediated, l [(3)H]proline influx was abolished. At low external concentrations of zinc alone (e.g. below 1.0 mmol l(-)(1)), l-[(3)H]proline influx was enhanced by the metal. Enhanced amino acid uptake in the presence of external zinc alone was abolished by l-pipecolate. A model accounting for external and internal zinc enhancements of l-[(3)H]proline influx by the Na(+)-dependent l-pipecolate-sensitive IMINO transport system in these membranes is proposed. PMID- 10518482 TI - The supramolecular organisation of fibrillin-rich microfibrils determines the mechanical properties of bovine zonular filaments. AB - The zonular filaments from the eyes of cows are rich in microfibrils containing fibrillin. Tensile tests, stress-relaxation tests and X-ray diffraction studies were used to study the relationship between the mechanical behaviour of zonular filaments and the molecular packing and structure of the fibrillin-rich microfibrils. Zonular filaments show a non-linear (J-shaped) stress-strain curve and appreciable stress-relaxation. It is proposed that the non-linear properties are due to local variations in waviness in the microfibrils or assemblies of microfibrils, which straighten out and become more regularly aligned with strain. Previous and current X-ray diffraction results consistently show a partial ordering of microfibrils in zonular filaments into staggered aggregates which become more ordered and laterally aligned on stretching. Although the removal and re-addition of Ca(2+) is known to change the molecular structure of fibrillin, no effect was observed on the tensile properties of the zonular filaments. It is hypothesised that strain-induced deformation in the supramolecular aggregate packing may not be Ca(2+)-sensitive but could dominate the mechanical behaviour of microfibrillar arrays in zonular filaments. PMID- 10518483 TI - Inhibiting ventilatory evaporation produces an adaptive increase in cutaneous evaporation in mourning doves Zenaida macroura. AB - We tested the hypothesis that birds can rapidly change the conductance of water vapor at the skin surface in response to a changing need for evaporative heat loss. Mourning doves (Zenaida macroura) were placed in a two-compartment chamber separating the head from the rest of the body. The rate of cutaneous evaporation was measured in response to dry ventilatory inflow at three ambient temperatures and in response to vapor-saturated ventilatory inflow at two ambient temperatures. At 35 degrees C, cutaneous evaporation increased by 72 % when evaporative water loss from the mouth was prevented, but no increase was observed at 45 degrees C. For both dry and vapor-saturated treatments, cutaneous evaporation increased significantly with increased ambient temperature. Changes in skin temperature made only a minor contribution to any observed increase in cutaneous evaporation. This indicates that Z. macroura can effect rapid adjustment of evaporative conductance at the skin in response to acute change in thermoregulatory demand. PMID- 10518484 TI - Sources of magnetic sensory input to identified neurons active during crawling in the marine mollusc Tritonia diomedea. AB - Although the nudibranch mollusc Tritonia diomedea orients to the geomagnetic field, the anatomical site and the mechanism of the geomagnetic transducer are not known. Previous work on semi-intact preparations of Tritonia diomedea in which the brain is intact and nerve connections to the periphery are maintained showed that identifiable pedal ganglion neurons Pd5 fired an increased number of action potentials when the horizontal component of the ambient magnetic field was rotated. This response disappeared when all nerves emerging from the brain were cut, suggesting a peripheral locus for the geomagnetic transducer. In the present work, we recorded intracellularly from Pd5 in preparations in which all peripheral nerves were cut except those containing the axons of neurons Pd5 (pedal nerves 2 and 3). These uncut, mixed, sensory-motor trunks innervate the locomotory epithelium of the foot upon which the animal crawls. In this further reduced preparation, Pd5 again responded to magnetic field rotations with action potentials. To determine the direction of this action potential transmission in response to magnetic field rotations, we analyzed extracellular recordings from nerves containing the Pd5 axons and found that action potentials elicited in Pd5 by magnetic stimuli originate centrally and are transmitted peripherally. In addition, we have explored the behavioral function of Pd5 neurons by simultaneously recording intracellular electrical activity and crawling rate of the semi-intact animal. A significant correlation was found between crawling rate and Pd5 action potential rate. We also found that action potentials in dorsal swim interneurons depolarized both Pd5 and the established locomotion motoneuron Pd21. PMID- 10518485 TI - The hormonal coordination of behavior and physiology at adult ecdysis in Drosophila melanogaster. AB - In insects, ecdysis is thought to be controlled by the interaction between peptide hormones; in particular between ecdysis-triggering hormone (ETH) from the periphery and eclosion hormone (EH) and crustacean cardioactive peptide (CCAP) from the central nervous system. We examined the behavioral and physiological functions of the first two of these peptides in Drosophila melanogaster using wild-type flies and knockout flies that lacked EH neurons. We used ETH from Manduca sexta (MasETH) to induce premature ecdysis and compared the responses of the two types of flies. The final release of EH normally occurs approximately 40 min before ecdysis. It is correlated with cyclic guanosine monophosphate (cGMP) production in selected neurons and tracheae, by an elevation in the heart rate and by the filling of the new tracheae with air. Injection of developing flies with MasETH causes all these events to occur prematurely. In EH cell knockouts, none of these changes occurs in response to MasETH, and these flies show a permanent failure in tracheal filling. This failure can be overcome in the knockouts by injecting them with membrane-permeant analogs of cGMP, the second messenger for EH. The basis for the 40 min delay between EH release and the onset of ecdysis was examined by decapitating flies at various times relative to EH release. In flies that had already released EH, decapitation was always followed within 1 min by the start of ecdysis. Immediate ecdysis was never observed when the EH cell knockout flies were decapitated. We propose that EH activates both ventral central nervous system elements necessary for ecdysis (possibly the CCAP cells) and descending inhibitory neurons from the head. This descending inhibition establishes a delay in the onset of ecdysis that allows the completion of EH-activated physiological processes such as tracheal filling. A waning in the inhibition eventually allows ecdysis to begin 30-40 min later. PMID- 10518486 TI - Clonal analysis of Drosophila embryonic neuroblasts: neural cell types, axon projections and muscle targets. AB - An experimental analysis of neurogenesis requires a detailed understanding of wild-type neural development. Recent DiI cell lineage studies have begun to elucidate the family of neurons and glia produced by each Drosophila embryonic neural precursor (neuroblast). Here we use DiI labeling to extend and clarify previous studies, but our analysis differs from previous studies in four major features: we analyze and compare lineages of every known embryonic neuroblast; we use an in vivo landmark (engrailed-GFP) to increase the accuracy of neuroblast identification; we use confocal fluorescence and Nomarski microscopy to collect three-dimensional data in living embryos simultaneously for each DiI-labeled clone, the engrailed-GFP landmark, and the entire CNS and muscle target field (Nomarski images); and finally, we analyze clones very late in embryonic development, which reveals novel cell types and axon/dendrite complexity. We identify the parental neuroblasts for all the cell types of the embryonic CNS: motoneurons, intersegmental interneurons, local interneurons, glia and neurosecretory cells (whose origins had never been determined). We identify muscle contacts for every thoracic and abdominal motoneuron at stage 17. We define the parental neuroblasts for neurons or glia expressing well-known molecular markers or neurotransmitters. We correlate Drosophila cell lineage data with information derived from other insects. In addition, we make the following novel conclusions: (1) neuroblasts at similar dorsoventral positions, but not anteroposterior positions, often generate similar cell lineages, and (2) neuroblasts at similar dorsoventral positions often produce the same motoneuron subtype: ventral neuroblasts typically generate motoneurons with dorsal muscle targets, while dorsal neuroblasts produce motoneurons with ventral muscle targets. Lineage data and movies can be found at http://www.biologists. com/Development/movies/dev8623.html http://www.neuro.uoregon. edu/doelab/lineages/ PMID- 10518487 TI - The orderly allocation of mesodermal cells to the extraembryonic structures and the anteroposterior axis during gastrulation of the mouse embryo. AB - The prospective fate of cells in the primitive streak was examined at early, mid and late stages of mouse gastrula development to determine the order of allocation of primitive streak cells to the mesoderm of the extraembryonic membranes and to the fetal tissues. At the early-streak stage, primitive streak cells contribute predominantly to tissues of the extraembryonic mesoderm as previously found. However, a surprising observation is that the erythropoietic precursors of the yolk sac emerge earlier than the bulk of the vitelline endothelium, which is formed continuously throughout gastrula development. This may suggest that the erythropoietic and the endothelial cell lineages may arise independently of one another. Furthermore, the extraembryonic mesoderm that is localized to the anterior and chorionic side of the yolk sac is recruited ahead of that destined for the posterior and amnionic side. For the mesodermal derivatives in the embryo, those destined for the rostral structures such as heart and forebrain mesoderm ingress through the primitive streak early during a narrow window of development. They are then followed by those for the rest of the cranial mesoderm and lastly the paraxial and lateral mesoderm of the trunk. Results of this study, which represent snapshots of the types of precursor cells in the primitive streak, have provided a better delineation of the timing of allocation of the various mesodermal lineages to specific compartments in the extraembryonic membranes and different locations in the embryonic anteroposterior axis. PMID- 10518488 TI - Gap junction-mediated transfer of left-right patterning signals in the early chick blastoderm is upstream of Shh asymmetry in the node. AB - Invariant patterning of left-right asymmetry during embryogenesis depends upon a cascade of inductive and repressive interactions between asymmetrically expressed genes. Different cascades of asymmetric genes distinguish the left and right sides of the embryo and are maintained by a midline barrier. As such, the left and right sides of an embryo can be viewed as distinct and autonomous fields. Here we describe a series of experiments that indicate that the initiation of these programs requires communication between the two sides of the blastoderm. When deprived of either the left or the right lateral halves of the blastoderm, embryos are incapable of patterning normal left-right gene expression at Hensen's node. Not only are both flanks required, suggesting that there is no single signaling source for LR pattern, but the blastoderm must be intact. These results are consistent with our previously proposed model in which the orientation of LR asymmetry in the frog, Xenopus laevis, depends on large-scale partitioning of LR determinants through intercellular gap junction channels (M. Levin and M. Mercola (1998) Developmental Biology 203, 90-105). Here we evaluate whether gap junctional communication is required for the LR asymmetry in the chick, where it is possible to order early events relative to the well-characterized left and right hierarchies of gene expression. Treatment of cultured chick embryos with lindane, which diminishes gap junctional communication, frequently unbiased normal LR asymmetry of Shh and Nodal gene expression, causing the normally left sided program to be recapitulated symmetrically on the right side of the embryo. A survey of early expression of connexin mRNAs revealed that Cx43 is present throughout the blastoderm at Hamburger-Hamilton stage 2-3, prior to known asymmetric gene expression. Application of antisense oligodeoxynucleotides or blocking antibody to cultured embryos also resulted in bilateral expression of Shh and Nodal transcripts. Importantly, the node and primitive streak at these stages lack Cx43 mRNA. This result, together with the requirement for an intact blastoderm, suggests that the path of communication through gap junction channels circumvents the node and streak. We propose that left-right information is transferred unidirectionally throughout the epiblast by gap junction channels in order to pattern left-sided Shh expression at Hensen's node. PMID- 10518490 TI - The dominant hemimelia mutation uncouples epithelial-mesenchymal interactions and disrupts anterior mesenchyme formation in mouse hindlimbs. AB - Epithelial-mesenchymal interactions are essential for both limb outgrowth and pattern formation in the limb. Molecules capable of communication between these two tissues are known and include the signaling molecules SHH and FGF4, FGF8 and FGF10. Evidence suggests that the pattern and maintenance of expression of these genes are dependent on a number of factors including regulatory loops between genes expressed in the AER and those in the underlying mesenchyme. We show here that the mouse mutation dominant hemimelia (Dh) alters the pattern of gene expression in the AER such that Fgf4, which is normally expressed in a posterior domain, and Fgf8, which is expressed throughout are expressed in anterior patterns. We show that maintenance of Shh expression in the posterior mesenchyme is not dependent on either expression of Fgf4 or normal levels of Fgf8 in the overlying AER. Conversely, AER expression of Fgf4 is not directly dependent on Shh expression. Also the reciprocal regulatory loop proposed for Fgf8 in the AER and Fgf10 in the underlying mesenchyme is also uncoupled by this mutation. Early during the process of limb initiation, Dh is involved in regulating the width of the limb bud, the mutation resulting in selective loss of anterior mesenchyme. The Dh gene functions in the initial stages of limb development and we suggest that these initial roles are linked to mechanisms that pattern gene expression in the AER. PMID- 10518489 TI - A two-step mechanism generates the spacing pattern of the ciliated cells in the skin of Xenopus embryos. AB - The skin of Xenopus embryos contains a population of specialized ciliated cells that are distributed in an evenly spaced pattern. Here we describe two successive steps that govern the differentiation and the generation of the spacing pattern of these ciliated cells. The first step occurs in the inner or sensorial layer of the non-neural ectoderm where a subset of cells are chosen to differentiate into ciliated-cell precursors. This choice is under the control of lateral inhibition mediated by a Suppressor of Hairless-dependent Notch signaling pathway, in which X-Delta-1 is the putative ligand driving the selection process, and a new Enhancer-of-Split-related gene is an epidermal target of Notch signaling. Because nascent ciliated-cell precursors prevent neighboring cells from taking on the same fate, a scattered pattern of these precursors is generated within the deep layer of the non-neural ectoderm. Ciliated-cell precursors then intercalate into the outer layer of cells in the epidermis. We show that the intercalation event acts as a second step to regulate the spacing of the mature ciliated cells. We propose that the differentiation of the ciliated cells is not only regulated by Notch-mediated lateral inhibition, but is also an example where differentiation is coupled to the movement of cells from one cell layer to another. PMID- 10518491 TI - Patterning the ascidian nervous system: structure, expression and transgenic analysis of the CiHox3 gene. AB - Hox genes play a fundamental role in the establishment of chordate body plan, especially in the anteroposterior patterning of the nervous system. Particularly interesting are the anterior groups of Hox genes (Hox1-Hox4) since their expression is coupled to the control of regional identity in the anterior regions of the nervous system, where the highest structural diversity is observed. Ascidians, among chordates, are considered a good model to investigate evolution of Hox gene, organisation, regulation and function. We report here the cloning and the expression pattern of CiHox3, a Ciona intestinalis anterior Hox gene homologous to the paralogy group 3 genes. In situ hybridization at the larva stage revealed that CiHox3 expression was restricted to the visceral ganglion of the central nervous system. The presence of a sharp posterior boundary and the absence of transcript in mesodermal tissues are distinctive features of CiHox3 expression when compared to the paralogy group 3 in other chordates. We have investigated the regulatory elements underlying CiHox3 neural-specific expression and, using transgenic analysis, we were able to isolate an 80 bp enhancer responsible of CiHox3 activation in the central nervous system (CNS). A comparative study between mouse and Ciona Hox3 promoters demonstrated that divergent mechanisms are involved in the regulation of these genes in vertebrates and ascidians. PMID- 10518492 TI - Regulation of the onset of neural crest migration by coordinated activity of BMP4 and Noggin in the dorsal neural tube. AB - For neural crest cells to engage in migration, it is necessary that epithelial premigratory crest cells convert into mesenchyme. The mechanisms that trigger cell delamination from the dorsal neural tube remain poorly understood. We find that, in 15- to 40-somite-stage avian embryos, BMP4 mRNA is homogeneously distributed along the longitudinal extent of the dorsal neural tube, whereas its specific inhibitor noggin exists in a gradient of expression that decreases caudorostrally. This rostralward reduction in signal intensity coincides with the onset of emigration of neural crest cells. Hence, we hypothesized that an interplay between Noggin and BMP4 in the dorsal tube generates graded concentrations of the latter that in turn triggers the delamination of neural crest progenitors. Consistent with this suggestion, disruption of the gradient by grafting Noggin-producing cells dorsal to the neural tube at levels opposite the segmental plate or newly formed somites, inhibited emigration of HNK-1-positive crest cells, which instead accumulated within the dorsal tube. Similar results were obtained with explanted neural tubes from the same somitic levels exposed to Noggin. Exposure to Follistatin, however, had no effect. The Noggin-dependent inhibition was overcome by concomitant treatment with BMP4, which when added alone, also accelerated cell emigration compared to untreated controls. Furthermore, the observed inhibition of neural crest emigration in vivo was preceded by a partial or total reduction in the expression of cadherin-6B and rhoB but not in the expression of slug mRNA or protein. Altogether, these results suggest that a coordinated activity of Noggin and BMP4 in the dorsal neural tube triggers delamination of specified, slug-expressing neural crest cells. Thus, BMPs play multiple and discernible roles at sequential stages of neural crest ontogeny, from specification through delamination and later differentiation of specific neural crest derivatives. PMID- 10518493 TI - The early-flowering mutant efs is involved in the autonomous promotion pathway of Arabidopsis thaliana. AB - The transition to flowering is a crucial moment in a plant's life cycle of which the mechanism has only been partly revealed. In a screen for early flowering, after mutagenesis of the late-flowering fwa mutant of Arabidopsis thaliana, the early flowering in short days (efs) mutant was identified. Under long-day light conditions, the recessive monogenic efs mutant flowers at the same time as wild type but, under short-day conditions, the mutant flowers much earlier. In addition to its early-flowering phenotype, efs has several pleiotropic effects such as a reduction in plant size, fertility and apical dominance. Double mutant analysis with several late-flowering mutants from the autonomous promotion (fca and fve) and the photoperiod promotion (co, fwa and gi) pathways of flowering showed that efs reduces the flowering time of all these mutants. However, efs is completely epistatic to fca and fve but additive to co, fwa and gi, indicating that EFS is an inhibitor of flowering specifically involved in the autonomous promotion pathway. A vernalisation treatment does not further reduce the flowering time of the efs mutant, suggesting that vernalisation promotes flowering through EFS. By comparing the length of the juvenile and adult phases of vegetative growth for wild-type, efs and the double mutant plants, it is apparent that efs mainly reduces the length of the adult phase. PMID- 10518494 TI - Defining subregions of Hensen's node essential for caudalward movement, midline development and cell survival. AB - Hensen's node, also called the chordoneural hinge in the tail bud, is a group of cells that constitutes the organizer of the avian embryo and that expresses the gene HNF-3(&bgr;). During gastrulation and neurulation, it undergoes a rostral-to caudal movement as the embryo elongates. Labeling of Hensen's node by the quail chick chimera system has shown that, while moving caudally, Hensen's node leaves in its wake not only the notochord but also the floor plate and a longitudinal strand of dorsal endodermal cells. In this work, we demonstrate that the node can be divided into functionally distinct subregions. Caudalward migration of the node depends on the presence of the most posterior region, which is closely apposed to the anterior portion of the primitive streak as defined by expression of the T-box gene Ch-Tbx6L. We call this region the axial-paraxial hinge because it corresponds to the junction of the presumptive midline axial structures (notochord and floor plate) and the paraxial mesoderm. We propose that the axial paraxial hinge is the equivalent of the neuroenteric canal of other vertebrates such as Xenopus. Blocking the caudal movement of Hensen's node at the 5- to 6 somite stage by removing the axial-paraxial hinge deprives the embryo of midline structures caudal to the brachial level, but does not prevent formation of the neural tube and mesoderm located posteriorly. However, the whole embryonic region generated posterior to the level of Hensen's node arrest undergoes widespread apoptosis within the next 24 hours. Hensen's node-derived structures (notochord and floor plate) thus appear to produce maintenance factor(s) that ensures the survival and further development of adjacent tissues. PMID- 10518495 TI - Essential role for the homeoprotein vHNF1/HNF1beta in visceral endoderm differentiation. AB - vHNF1/HNF1beta, a member of the divergent HNF1/vHNF1 homeoprotein family, is expressed in polarized epithelia of several adult organs and may participate in controlling the transcription of specific genes. In addition to this late requirement, vHNF1 may play earlier roles during development, as it is first expressed in the visceral endoderm at the onset of gastrulation. In order to shed light on its function during embryogenesis, we have inactivated the murine gene by homologous recombination. The homozygous mutation results in embryonic lethality by day 7.5 of development and vHNF1(-)(/)(-) embryos display a disorganized visceral endoderm and a significantly reduced size. Studies of ES cell differentiation and aggregation with tetraploid morulae establish that vHNF1 expression is essential for visceral endoderm differentiation, both in vitro and in vivo. Analysis of differentiation markers confirms that vHNF1 is part of a genetic network that directs the expression of HNF4 and downstream endodermal genes. Furthermore, the complementation of the mutant embryos with wild-type visceral endoderm rescues the day 7.5 lethality and reveals an additional phenotype linked to vHNF1 later expression. The examination of chimeric embryos suggests that vHNF1 expression might be cell-autonomously required in the gut for the proper morphogenesis of the embryo. PMID- 10518496 TI - Variant hepatocyte nuclear factor 1 is required for visceral endoderm specification. AB - Genetic and molecular evidence indicates that visceral endoderm, an extraembryonic cell lineage, is required for gastrulation, early anterior neural patterning, cell death and specification of posterior mesodermal cell fates. We show that variant Hepatocyte Nuclear Factor 1 (vHNF1), a homeodomain-containing transcription factor first expressed in the primitive endoderm, is required for the specification of visceral endoderm. vHnf1-deficient mouse embryos develop normally to the blastocyst stage, start implantation, but die soon afterwards, with abnormal or absent extraembryonic region, poorly organised ectoderm and no discernible visceral or parietal endoderm. However, immunostaining analysis of E5.5 nullizygous mutant embryos revealed the presence of parietal endoderm-like cells lying on an abnormal basal membrane. Homozygous mutant blastocyst outgrowths or differentiated embryonic stem cells do not express early or late visceral endoderm markers. In addition, in vHnf1 null embryoid bodies there is no activation of the transcription factors HNF-4alpha1, HNF1alpha and HNF-3gamma. Aggregation of vHnf1-deficient embryonic stem cells with wild-type tetraploid embryos, which contribute exclusively to extraembryonic tissues, rescues periimplantation lethality and allows development to progress to early organogenesis. Our results place vHNF1 in a preeminent position in the regulatory network that specifies the visceral endoderm and highlight the importance of this cell lineage for proper growth and differentiation of primitive ectoderm in pregastrulating embryos. PMID- 10518497 TI - TONDU (TDU), a novel human protein related to the product of vestigial (vg) gene of Drosophila melanogaster interacts with vertebrate TEF factors and substitutes for Vg function in wing formation. AB - The mammalian TEF and the Drosophila scalloped genes belong to a conserved family of transcriptional factors that possesses a TEA/ATTS DNA-binding domain. Transcriptional activation by these proteins likely requires interactions with specific coactivators. In Drosophila, Scalloped (Sd) interacts with Vestigial (Vg) to form a complex, which binds DNA through the Sd TEA/ATTS domain. The Sd-Vg heterodimer is a key regulator of wing development, which directly controls several target genes and is able to induce wing outgrowth when ectopically expressed. Here we show that Vg contains two distinct transcriptional activation domains, suggesting that the function of Vg is to mediate transcriptional activation by Sd. By expressing a chimeric GAL4-Sd protein in Drosophila, we found that the transcriptional activity of the Vg-Sd heterodimer is negatively regulated at the AP and DV boundary of the wing disc. We also identify a novel human protein, TONDU, which contains a short domain homologous to the domain of Vg required for interaction with Sd. We show that TONDU specifically interacts with a domain conserved in all the mammalian TEF factors. Expression of TDU in Drosophila by means of the UAS-GAL4 system shows that this human protein can substitute for Vg in wing formation. We propose that TDU is a specific coactivator for the mammalian TEFs. PMID- 10518498 TI - Transient establishment of anteroposterior polarity in the zebrafish pectoral fin bud in the absence of sonic hedgehog activity. AB - Sonic hedgehog (Shh) is expressed in the posterior vertebrate limb bud mesenchyme and directs anteroposterior patterning and growth during limb development. Here we report an analysis of the pectoral fin phenotype of zebrafish sonic you mutants, which disrupt the shh gene. We show that Shh is required for the establishment of some aspects of anteroposterior polarity, while other aspects of anteroposterior polarity are established independently of Shh, and only later come to depend on Shh for their maintenance. We also demonstrate that Shh is required for the activation of posterior HoxD genes by retinoic acid. Finally, we show that Shh is required for normal development of the apical ectodermal fold, for growth of the fin bud, and for formation of the fin endoskeleton. PMID- 10518499 TI - FGF8 can activate Gbx2 and transform regions of the rostral mouse brain into a hindbrain fate. AB - The mid/hindbrain junction region, which expresses Fgf8, can act as an organizer to transform caudal forebrain or hindbrain tissue into midbrain or cerebellar structures, respectively. FGF8-soaked beads placed in the chick forebrain can similarly induce ectopic expression of mid/hindbrain genes and development of midbrain structures (Crossley, P. H., Martinez, S. and Martin, G. R. (1996) Nature 380, 66-68). In contrast, ectopic expression of Fgf8a in the mouse midbrain and caudal forebrain using a Wnt1 regulatory element produced no apparent patterning defects in the embryos examined (Lee, S. M., Danielian, P. S., Fritzsch, B. and McMahon, A. P. (1997) Development 124, 959-969). We show here that FGF8b-soaked beads can not only induce expression of the mid/hindbrain genes En1, En2 and Pax5 in mouse embryonic day 9.5 (E9.5) caudal forebrain explants, but also can induce the hindbrain gene Gbx2 and alter the expression of Wnt1 in both midbrain and caudal forebrain explants. We also show that FGF8b soaked beads can repress Otx2 in midbrain explants. Furthermore, Wnt1-Fgf8b transgenic embryos in which the same Wnt1 regulatory element is used to express Fgf8b, have ectopic expression of En1, En2, Pax5 and Gbx2 in the dorsal hindbrain and spinal cord at E10.5, as well as exencephaly and abnormal spinal cord morphology. More strikingly, Fgf8b expression in more rostral brain regions appears to transform the midbrain and caudal forebrain into an anterior hindbrain fate through expansion of the Gbx2 domain and repression of Otx2 as early as the 7-somite stage. These findings suggest that normal Fgf8 expression in the anterior hindbrain not only functions to maintain development of the entire mid/hindbrain by regulating genes like En1, En2 and Pax5, but also might function to maintain a metencephalic identity by regulating Gbx2 and Otx2 expression. PMID- 10518500 TI - A novel WD40 protein, CHE-2, acts cell-autonomously in the formation of C. elegans sensory cilia. AB - To elucidate the mechanism of sensory cilium formation, we analyzed mutants in the Caenorhabditis elegans che-2 gene. These mutants have extremely short cilia with an abnormal posterior projection, and show defects in behaviors that are mediated by ciliated sensory neurons. The che-2 gene encodes a new member of the WD40 protein family, suggesting that it acts in protein-protein interaction. Analysis of mutation sites showed that both the amino-terminal WD40 repeats and the carboxyl-terminal non-WD40 domain are necessary for the CHE-2 function. CHE-2 tagged green fluorescent protein is localized at the cilia of almost all the ciliated sensory neurons. Expression of che-2 in a subset of sensory neurons of a che-2 mutant by using a heterologous promoter resulted in restoration of the functions and cilium morphology of only the che-2-expressing neurons. Thus, che-2 acts cell-autonomously. This technique can be used in the future for determining the function of each type of che-2-expressing sensory neuron. Using green fluorescent protein, we found that the extension of cilia in wild-type animals took place at the late embryonic stage, whereas the cilia of che-2 mutant animals remained always short during development. Hence, the abnormal posterior projection is due to the inability of cilia to extend, rather than degeneration of cilia once correctly formed. Expression of che-2 in a che-2 mutant under a heat shock promoter showed that the extension of cilia, surprisingly, can occur even at the adult stage, and that such cilia can function apparently normally in behavior. PMID- 10518501 TI - Regulation of postembryonic G(1) cell cycle progression in Caenorhabditis elegans by a cyclin D/CDK-like complex. AB - In many organisms, initiation and progression through the G(1) phase of the cell cycle requires the activity of G(1)-specific cyclins (cyclin D and cyclin E) and their associated cyclin-dependent kinases (CDK2, CDK4, CDK6). We show here that the Caenorhabditis elegans genes cyd-1 and cdk-4, encoding proteins similar to cyclin D and its cognate cyclin-dependent kinases, respectively, are necessary for proper division of postembryonic blast cells. Animals deficient for cyd-1 and/or cdk-4 activity have behavioral and developmental defects that result from the inability of the postembryonic blast cells to escape G(1) cell cycle arrest. Moreover, ectopic expression of cyd-1 and cdk-4 in transgenic animals is sufficient to activate a S-phase reporter gene. We observe no embryonic defects associated with depletion of either of these two gene products, suggesting that their essential functions are restricted to postembryonic development. We propose that the cyd-1 and cdk-4 gene products are an integral part of the developmental control of larval cell proliferation through the regulation of G(1) progression. PMID- 10518502 TI - nos-1 and nos-2, two genes related to Drosophila nanos, regulate primordial germ cell development and survival in Caenorhabditis elegans. AB - In Drosophila, the posterior determinant nanos is required for embryonic patterning and for primordial germ cell (PGC) development. We have identified three genes in Caenorhabditis elegans that contain a putative zinc-binding domain similar to the one found in nanos, and show that two of these genes function during PGC development. Like Drosophila nanos, C. elegans nos-1 and nos-2 are not generally required for PGC fate specification, but instead regulate specific aspects of PGC development. nos-2 is expressed in PGCs around the time of gastrulation from a maternal RNA associated with P granules, and is required for the efficient incorporation of PGCs into the somatic gonad. nos-1 is expressed in PGCs after gastrulation, and is required redundantly with nos-2 to prevent PGCs from dividing in starved animals and to maintain germ cell viability during larval development. In the absence of nos-1 and nos-2, germ cells cease proliferation at the end of the second larval stage, and die in a manner that is partially dependent on the apoptosis gene ced-4. Our results also indicate that putative RNA-binding proteins related to Drosophila Pumilio are required for the same PGC processes as nos-1 and nos-2. These studies demonstrate that evolutionarily distant organisms utilize conserved factors to regulate early germ cell development and survival, and that these factors include members of the nanos and pumilio gene families. PMID- 10518503 TI - The role of sonic hedgehog in normal and abnormal craniofacial morphogenesis. AB - There is growing evidence that implicates a role for Sonic hedgehog (SHH) in morphogenesis of the craniofacial complex. Mutations in human and murine SHH cause midline patterning defects that are manifested in the head as holoprosencephaly and cyclopia. In addition, teratogens such as jervine, which inhibit the response of tissues to SHH, also produce cyclopia. Thus, the loss of SHH signaling during early stages of neural plate patterning has a profound influence of craniofacial morphogenesis. However, the severity of these defects precludes analyses of SHH function during later stages of craniofacial development. We have used an embryonic chick system to study the role of SHH during these later stages of craniofacial development. Using a combination of surgical and molecular experiments, we show here that SHH is essential for morphogenesis of the frontonasal and maxillary processes (FNP and MXPs), which give rise to the mid- and upper face. Transient loss of SHH signaling in the embryonic face inhibits growth of the primordia and results in defects analogous to hypotelorism and cleft lip/palate, characteristics of the mild forms of holoprosencephaly. In contrast, excess SHH leads to a mediolateral widening of the FNP and a widening between the eyes, a condition known as hypertelorism. In severe cases, this widening is accompanied by facial duplications. Collectively, these experiments demonstrate that SHH has multiple and profound effects on the entire spectrum of craniofacial development, and perturbations in SHH signaling are likely to underlie a number of human craniofacial anomalies. PMID- 10518504 TI - SF/HGF is a mediator between limb patterning and muscle development. AB - Scatter factor/hepatocyte growth factor (SF/HGF) is known to be involved in the detachment of myogenic precursor cells from the lateral dermomyotomes and their subsequent migration into the newly formed limb buds. As yet, however, nothing has been known about the role of the persistent expression of SF/HGF in the limb bud mesenchyme during later stages of limb bud development. To test for a potential role of SF/HGF in early limb muscle patterning, we examined the regulation of SF/HGF expression in the limb bud as well as the influence of SF/HGF on direction control of myogenic precursor cells in limb bud mesenchyme. We demonstrate that SF/HGF expression is controlled by signals involved in limb bud patterning. In the absence of an apical ectodermal ridge (AER), no expression of SF/HGF in the limb bud is observed. However, FGF-2 application can rescue SF/HGF expression. Excision of the zone of polarizing activity (ZPA) results in ectopic and enhanced SF/HGF expression in the posterior limb bud mesenchyme. We could identify BMP-2 as a potential inhibitor of SF/HGF expression in the posterior limb bud mesenchyme. We further demonstrate that ZPA excision results in a shift of Pax-3-positive cells towards the posterior limb bud mesenchyme, indicating a role of the ZPA in positioning of the premuscle masses. Moreover, we present evidence that, in the limb bud mesenchyme, SF/HGF increases the motility of myogenic precursor cells and has a role in maintaining their undifferentiated state during migration. We present a model for a crucial role of SF/HGF during migration and early patterning of muscle precursor cells in the vertebrate limb. PMID- 10518505 TI - A requirement for neuropilin-1 in embryonic vessel formation. AB - Neuropilin-1 is a membrane protein that is expressed in developing neurons and functions as a receptor or a component of the receptor complex for the class 3 semaphorins, which are inhibitory axon guidance signals. Targeted inactivation of the neuropilin-1 gene in mice induced disorganization of the pathway and projection of nerve fibers, suggesting that neuropilin-1 mediates semaphorin elicited signals and regulates nerve fiber guidance in embryogenesis. Neuropilin 1 is also expressed in endothelial cells and shown to bind vascular endothelial growth factor (VEGF), a potent regulator for vasculogenesis and angiogenesis. However, the roles of neuropilin-1 in vascular formation have been unclear. This paper reported that the neuropilin-1 mutant mouse embryos exhibited various types of vascular defects, including impairment in neural vascularization, agenesis and transposition of great vessels, insufficient aorticopulmonary truncus (persistent truncus arteriosus), and disorganized and insufficient development of vascular networks in the yolk sac. The vascular defects induced by neuropilin-1 deficiency in mouse embryos suggest that neuropilin-1 plays roles in embryonic vessel formation, as well as nerve fiber guidance. PMID- 10518506 TI - Mode of action of VegT in mesoderm and endoderm formation. AB - mRNA encoding the T-box transcription factor VegT is located throughout the vegetal pole of the Xenopus egg and is believed to play an important part in endoderm and mesoderm formation. We find that VegT generates endoderm both by cell-autonomous action and by generating TGF-beta family signals, the latter being entirely responsible for its mesoderm-inducing activity. Signalling molecules induced cell-autonomously by VegT include derriere, Xnr4 and activin B. Xnr1 and Xnr2 are also induced, but primarily in a non-autonomous manner. All of these signalling molecules are found in the blastula and gastrula vegetal pole and induce both endoderm and mesoderm in the animal cap assay, and hence are good candidates both for the endogenous zygotic mesoderm-inducing signal and for reinforcing the vegetal expression of endoderm markers. PMID- 10518507 TI - The dorsal-open group gene raw is required for restricted DJNK signaling during closure. AB - During dorsal closure in Drosophila melanogaster, cells of the lateral epidermis migrate over the amnioserosa to encase the embryo. At least three classes of dorsal-open group gene products are necessary for this morphogenetic movement. Class I genes code for structural proteins that effect changes in epidermal cell shape and motility. Class II and III genes code for regulatory components of closure: Class II genes encode Drosophila Jun amino (N)-terminal kinase (DJNK) signaling molecules and Class III genes encode Decapentaplegic-mediated signaling molecules. All characterized dorsal-open group gene products function in the epidermis. Here we report a molecular and genetic characterization of raw, a newly defined member of the Class II dorsal-open group genes. We show that the novel protein encoded by raw is required for restriction of DJNK signaling to leading edge epidermal cells as well as for proper development of the amnioserosa. Taken together, our results demonstrate a role for Raw in restriction of epidermal signaling during closure and suggest that this effect may be mediated via the amnioserosa. PMID- 10518508 TI - Redesign of PNAS: getting ready for the millennium. PMID- 10518509 TI - Chain entropy: spoiler or benefactor in pattern recognition? PMID- 10518510 TI - Double-stranded RNA-activated protein kinase mediates virus-induced apoptosis: a new role for an old actor. PMID- 10518511 TI - An advance in liver-specific gene delivery. PMID- 10518512 TI - Go-ing for the prediction of protein folding mechanisms. PMID- 10518513 TI - Segregation-driven organization in chaotic granular flows. AB - An important industrial problem that provides fascinating puzzles in pattern formation is the tendency for granular mixtures to de-mix or segregate. Small differences in either size or density lead to flow-induced segregation. Similar to fluids, noncohesive granular materials can display chaotic advection; when this happens chaos and segregation compete with each other, giving rise to a wealth of experimental outcomes. Segregated structures, obtained experimentally, display organization in the presence of disorder and are captured by a continuum flow model incorporating collisional diffusion and density-driven segregation. Under certain conditions, structures never settle into a steady shape. This may be the simplest experimental example of a system displaying competition between chaos and order. PMID- 10518514 TI - Simulation of biomimetic recognition between polymers and surfaces. AB - Many biological processes, such as transmembrane signaling and pathogen-host interactions, are initiated by a protein recognizing a specific pattern of binding sites on part of a membrane or cell surface. By recognition, we imply that the polymer quickly finds and then adsorbs strongly on the pattern-matched region and not on others. The development of synthetic systems that can mimic such recognition between polymers and surfaces could have significant impact on advanced applications such as the development of sensors, molecular-scale separation processes, and synthetic viral inhibition agents. Attempting to affect recognition in synthetic systems by copying the detailed chemistries to which nature has been led over millenia of evolution does not seem practical for most applications. This leads us to the following question: Are there any universal strategies that can affect recognition between polymers and surfaces? Such generic strategies may be easier to implement in abiotic applications. We describe results that suggest that biomimetic recognition between synthetic polymers and surfaces is possible by exploiting certain generic strategies, and we elucidate the kinetic mechanisms by which this occurs. Our results suggest convenient model systems for experimental studies of dynamics in free energy landscapes characteristic of frustrated systems. PMID- 10518515 TI - Long-range charge hopping in DNA. AB - The fundamental mechanisms of charge migration in DNA are pertinent for current developments in molecular electronics and electrochemistry-based chip technology. The energetic control of hole (positive ion) multistep hopping transport in DNA proceeds via the guanine, the nucleobase with the lowest oxidation potential. Chemical yield data for the relative reactivity of the guanine cations and of charge trapping by a triple guanine unit in one of the strands quantify the hopping, trapping, and chemical kinetic parameters. The hole-hopping rate for superexchange-mediated interactions via two intervening AT base pairs is estimated to be 10(9) s(-1) at 300 K. We infer that the maximal distance for hole hopping in the duplex with the guanine separated by a single AT base pair is 300 +/- 70 A. Although we encounter constraints for hole transport in DNA emerging from the number of the mediating AT base pairs, electron transport is expected to be nearly sequence independent because of the similarity of the reduction potentials of the thymine and of the cytosine. PMID- 10518516 TI - Crystal structure of the DNA nucleotide excision repair enzyme UvrB from Thermus thermophilus. AB - Nucleotide excision repair (NER) is the most important DNA-repair mechanism in living organisms. In prokaryotes, three enzymes forming the UvrABC system initiate NER of a variety of structurally different DNA lesions. UvrB, the central component of this system, is responsible for the ultimate DNA damage recognition and participates in the incision of the damaged DNA strand. The crystal structure of Thermus thermophilus UvrB reveals a core that is structurally similar to core regions found in helicases, where they constitute molecular motors. Additional domains implicated in binding to DNA and various components of the NER system are attached to this central core. The architecture and distribution of DNA binding sites suggest a possible model for the DNA damage recognition process. PMID- 10518517 TI - Selection of antibodies for intracellular function using a two-hybrid in vivo system. AB - Expression of antibodies inside cells has been used successfully to ablate protein function. This finding suggests that the technology should have an impact on disease treatment and in functional genomics where proteins of unknown function are predicted from genomic sequences. A major hindrance is the paucity of antibodies that function in eukaryotic cells, presumably because the antibodies fold incorrectly in the cytoplasm. To overcome this problem, we have developed an in vivo assay for functional intracellular antibodies using a two hybrid approach. In this assay, antibody, as single-chain Fv (scFv) linked to a transcriptional transactivation domain, can interact with a target antigen, linked to a LexA-DNA binding domain, and thereby activate a reporter gene. We find that several characterized antibodies can bind their target antigen in eukaryotic cells in this two-hybrid format, and we have been able to isolate intracellular binders from among sets of scFv that can bind antigen in vitro. Furthermore, we show a model selection in which a single scFv was isolated from a mixture of half a million clones, indicating that this is a robust procedure that should facilitate capture of antibody specificities from complex mixtures. The approach can provide the basis for de novo selection of intracellular scFv from libraries, such as those made from spleen RNA after immunization with antigen, for intracellular analysis of protein function based only on genomic or cDNA sequences. PMID- 10518518 TI - Peroxynitrite reaction products of 3',5'-di-O-acetyl-8-oxo-7, 8-dihydro-2' deoxyguanosine. AB - Of the DNA bases, peroxynitrite (ONOO-) is most reactive toward 2'-deoxyguanosine (dGuo), but even more reactive with 8-oxo-7, 8-dihydro-2'-deoxyguanosine (8 oxodGuo), requiring a 1,000-fold excess of dGuo to provide 50% protection against the reaction with 8-oxodGuo. Therefore, it seems reasonable that 8-oxodGuo is a potentially important target in DNA and that the structures of the reaction products with ONOO- should be characterized. Using 3', 5'-di-O-Ac-8-oxodGuo as a model compound, the reaction products with ONOO- have been isolated and identified under simulated physiological reaction conditions (phosphate/bicarbonate buffer at pH 7.2). The major reaction product, II, is unstable and undergoes base-mediated hydrolysis to 2,5-diaminoimidazol-4-one, IIa, and 3-(3, 5-di-O-Ac-2-deoxy-beta-D-erythro-pentofuranosyl)-5 iminoimidazolidine -2,4-dione, IIb. The latter compound further hydrolyzes to 3 (3, 5-di-O-Ac-2-deoxy-beta-D-erythro-pentofuranosyl)oxaluric acid, IIc. Other products include 3-(3, 5-di-O-Ac-2-deoxy-beta-D-erythro-pentofuranosyl)-2,4,6 trioxo-[1,3, 5]triazinane-1-carboxamidine, I, which further hydrolyzes to 1-(3, 5 di-O-Ac-2-deoxy-beta-D-erythro-pentofuranosyl)cyanuric acid, Ia. 1-(3,5-di-O-Ac-2 deoxy-beta-D-erythro-pentofuranosyl)parabanic acid, III, is a minor product that also may contribute to formation of IIc. The major products formed in these reactions are biologically uncharacterized but are similar to modified DNA bases that have been shown to be both premutagenic and blocks to DNA polymerization. PMID- 10518519 TI - DNA specificity enhanced by sequential binding of protein monomers. AB - Transcriptional activation often requires the rapid assembly of complexes between dimeric transcription factors and specific DNA sites. Here we show that members of the basic region leucine zipper and basic region helix-loop-helix zipper transcription factor families follow an assembly pathway in which two protein monomers bind DNA sequentially and form their dimerization interface while bound to DNA. Nonspecific protein or DNA competitors have little effect on the rate of assembly along this pathway, but slow a competing pathway in which preformed dimers bind DNA. The sequential monomer-binding pathway allows the protein to search for and locate a specific DNA site more quickly, resulting in greater specificity prior to equilibrium. PMID- 10518520 TI - A multiplasmid approach to preparing large libraries of polyketides. AB - A three-plasmid system for heterologous expression of 6-deoxyerythronolide B synthase (DEBS) has been developed to facilitate combinatorial biosynthesis of polyketides made by type I modular polyketide synthases (PKSs). The eryA PKS genes encoding the three DEBS subunits were individually cloned into three compatible Streptomyces vectors carrying mutually selectable antibiotic resistance markers. A strain of Streptomyces lividans transformed with all three plasmids produced 6-deoxyerythronolide B at a level similar to that of a strain transformed with a single plasmid containing all three genes. The utility of this system in combinatorial biosynthesis was demonstrated through production of a library of modified polyketide macrolactones by using versions of each plasmid constructed to contain defined mutations. Combinations of these vector sets were introduced into S. lividans, resulting in strains producing a wide range of 6 deoxyerythronolide B analogs. This method can be extended to any modular PKS and has the potential to produce thousands of novel natural products, including ones derived from further modification of the PKS products by tailoring enzymes. PMID- 10518521 TI - Salt-induced detour through compact regions of the protein folding landscape. AB - In several cases, inorganic salts have been used to induce partly structured states in protein folding. But what is the nature of these states: Do they represent key stepping stones in the folding process, or are they circumstantial pitfalls in the energy landscape? Here we report that, in the case of the two state protein S6, the salt-induced collapsed state is off the usual folding routes in the sense that it is prematurely collapsed and slows down folding by several orders of magnitude. Although this species is over-compact, it is not a dead-end trap but may fold by alternative channels to the native state. PMID- 10518522 TI - An internal targeting signal directing proteins into the mitochondrial intermembrane space. AB - Import of most nucleus-encoded preproteins into mitochondria is mediated by N terminal presequences and requires a membrane potential and ATP hydrolysis. Little is known about the chemical nature and localization of other mitochondrial targeting signals or of the mechanisms by which they facilitate membrane passage. Mitochondrial heme lyases lack N-terminal targeting information. These proteins are localized in the intermembrane space and are essential for the covalent attachment of heme to c type cytochromes. For import of heme lyases, the translocase of the mitochondrial outer membrane complex is both necessary and sufficient. Here, we report the identification of the targeting signal of mitochondrial heme lyases in the third quarter of these proteins. The targeting sequence is highly conserved among all known heme lyases. Its chemical character is hydrophilic because of a large fraction of both positively and negatively charged amino acid residues. These features clearly distinguish this signal from classical presequences. When inserted into a cytosolic protein, the targeting sequence directs the fusion protein into the intermembrane space, even in the absence of a membrane potential or ATP hydrolysis. The heme lyase targeting sequence represents the first topogenic signal for energy-independent transport into the intermembrane space and harbors two types of information. It assures accurate recognition and translocation by the translocase of the mitochondrial outer membrane complex, and it is responsible for driving the import reaction by undergoing high-affinity interactions with components of the intermembrane space. PMID- 10518523 TI - Cytidine 5'-triphosphate-dependent biosynthesis of isoprenoids: YgbP protein of Escherichia coli catalyzes the formation of 4-diphosphocytidyl-2-C methylerythritol. AB - 2-C-methylerythritol 4-phosphate has been established recently as an intermediate of the deoxyxylulose phosphate pathway used for biosynthesis of terpenoids in plants and in many microorganisms. We show that an enzyme isolated from cell extract of Escherichia coli converts 2-C-methylerythritol 4-phosphate into 4 diphosphocytidyl-2-C-methylerythritol by reaction with CTP. The enzyme is specified by the hitherto unannotated ORF ygbP of E. coli. The cognate protein was obtained in pure form from a recombinant hyperexpression strain of E. coli harboring a plasmid with the ygbP gene under the control of a T5 promoter and lac operator. By using the recombinant enzyme, 4-diphosphocytidyl-[2-(14)C]2-C methylerythritol was prepared from [2-(14)C]2-C-methylerythritol 4-phosphate. The radiolabeled 4-diphosphocytidyl-2-C-methylerythritol was shown to be efficiently incorporated into carotenoids by isolated chromoplasts of Capsicum annuum. The E. coli ygbP gene appears to be part of a small operon also comprising the unannotated ygbB gene. Genes with similarity to ygbP and ygbB are present in the genomes of many microorganisms, and their occurrence appears to be correlated with that of the deoxyxylulose pathway of terpenoid biosynthesis. Moreover, several microorganisms have genes specifying putative fusion proteins with ygbP and ygbB domains, suggesting that both the YgbP protein and the YgbB protein are involved in the deoxyxylulose pathway. A gene from Arabidopsis thaliana with similarity to ygbP carries a putative plastid import sequence, which is well in line with the assumed localization of the deoxyxylulose pathway in the plastid compartment of plants. PMID- 10518524 TI - Correlation of deformability at a tRNA recognition site and aminoacylation specificity. AB - The fidelity of protein synthesis depends on specific tRNA aminoacylation by aminoacyl-tRNA synthetase enzymes, which in turn depends on the recognition of the identity of particular nucleotides and structural features in the substrate tRNA. These features generally reside within the acceptor helix, the anticodon stem-loop, and in some systems the variable pocket of the tRNA. In the alanine system, fidelity is ensured by a G.U wobble base pair located at the third position within the acceptor helix of alanine tRNA. We have investigated the activity of mutant alanine tRNAs to explore the mechanism of enzyme recognition. Here we show that the mismatched pair C-C is an excellent substitute for G.U in alanine-tRNA-knockout cells. A structural investigation by NMR spectroscopy of the C-C RNA acceptor end reveals that the two cytosines are intercalated into the helix, and that C-C exists in multiple conformations. Structural heterogeneity also is present in the wild-type G.U RNA, whereas inactive Watson-Crick helices are structurally rigid. The correlation between functional and structural data suggests that the G.U pair provides a distinctive structure and a point of deformability that allow the tRNA acceptor end to fit into the active site of the alanyl-tRNA synthetase. Fidelity is ensured because noncognate and inactive mutant tRNAs are bound in the active site in an incorrect conformation that reduces enzymatic activity. PMID- 10518525 TI - Two distinct mechanisms drive protein translocation across the mitochondrial outer membrane in the late step of the cytochrome b(2) import pathway. AB - The import of cytochrome b(2) into mitochondria consists of two steps. The translocation of the first part of the presequence across the inner membrane is coupled with the translocation of the tightly folded heme-binding domain across the outer membrane and requires a membrane potential DeltaPsi and the functions of mitochondrial Hsp70 (mHsp70) in the matrix. Once the heme-binding domain has passed the outer membrane, the translocation of the rest of the polypeptide chain across the outer membrane becomes independent of DeltaPsi and mHsp70. Here we analyzed the late DeltaPsi- and mHsp70-independent step in the transport of cytochrome b(2) fusion proteins into the intermembrane space (IMS). The import of the cytochrome b(2) fusion proteins containing two protein domains linked by a spacer segment into mitochondria was arrested at a stage at which one domain folded on each side of the outer membrane, along the pathway that is consistent with the stop-transfer model. The mature-size form of the translocation intermediate could move across the outer membrane in both directions, and the stabilization of the protein domain in the IMS promoted the forward translocation. On the other hand, the intermediate-size form of the translocation intermediate, which retains the anchorage to the inner membrane, was transported into the IMS independently of the stability of the protein domain in the IMS. These results suggest that two distinct mechanisms, the Brownian ratchet and the anchor diffusion mechanisms, can operate for the transmembrane movement of the mature-size form and the intermediate-size form, respectively, of cytochrome b(2) species. PMID- 10518526 TI - Glucocorticoid receptor inhibits transforming growth factor-beta signaling by directly targeting the transcriptional activation function of Smad3. AB - The transforming growth factor-beta (TGF-beta) family of cytokines and glucocorticoids regulate diverse biological processes through modulating the expression of target genes. Here we report that glucocorticoid receptor (GR) represses TGF-beta transcriptional activation of the type-1 plasminogen activator inhibitor (PAI-1) gene in a ligand-dependent manner. Similarly, GR represses TGF beta activation of the TGF-beta responsive sequence containing Smad3/4-binding sites. Using mammalian two-hybrid assays, we demonstrate that GR inhibits transcriptional activation by both Smad3 and Smad4 C-terminal activation domains. Finally, we show that GR interacts with Smad3 both in vitro and in vivo. These results suggest a molecular basis for the cross-regulation between glucocorticoid and TGF-beta signaling pathways. PMID- 10518527 TI - Nonstatistical binding of a protein to clustered carbohydrates. AB - Carbohydrate-derivatized self-assembled monolayers (SAMs) are used as a model system to address issues involving cell-surface carbohydrate-protein interactions. Here we examine the influence of carbohydrate surface density on protein-binding avidity. We show that the binding selectivity of Bauhinia purpurea lectin switches from one carbohydrate ligand to another as the surface density of the carbohydrate ligands increases from values of chi(sugar) approximately 0.1-1.0. Polyvalent binding is possible at all surface densities investigated; hence, the switch in selectivity is not due simply to the achievement of a critical density that permits polyvalent contacts. Instead, secondary interactions at high surface densities promote a switch in carbohydrate binding selectivity. These findings may have implications for how changes in the composition and the density of cell-surface carbohydrates influence biological recognition processes and regulatory pathways. PMID- 10518528 TI - Active-site structure of the soluble quinoprotein glucose dehydrogenase complexed with methylhydrazine: a covalent cofactor-inhibitor complex. AB - Soluble glucose dehydrogenase (s-GDH) from the bacterium Acinetobacter calcoaceticus is a classical quinoprotein. It requires the cofactor pyrroloquinoline quinone (PQQ) to catalyze the oxidation of glucose to gluconolactone. The precise catalytic role of PQQ in s-GDH and several other PQQ dependent enzymes has remained controversial because of the absence of comprehensive structural data. We have determined the crystal structure of a ternary complex of s-GDH with PQQ and methylhydrazine, a competitive inhibitor of the enzyme. This complex, refined at 1.5-A resolution to an R factor of 16.7%, affords a detailed view of a cofactor-binding site of s-GDH. Moreover, it presents the first direct observation of covalent PQQ adduct in the active-site of a PQQ-dependent enzyme, thereby confirming previous evidence that the C5 carbonyl group of the cofactor is the most reactive moiety of PQQ. PMID- 10518529 TI - C-terminal DNA binding stimulates N-terminal phosphorylation of the outer membrane protein regulator OmpR from Escherichia coli. AB - Expression of the porin genes of Escherichia coli is regulated in part by the osmolarity of the growth medium. The process is controlled by the histidine kinase EnvZ and the response regulator OmpR. We have previously shown that phosphorylation of OmpR increases its affinity for the upstream regulatory regions of ompF and ompC. We now report that, in the presence of DNA, there is a dramatic stimulation in the level of phospho-OmpR. This effect is independent of the source of phosphorylation, i.e., stimulation of phosphorylation is observed with a small phosphorylating agent such as acetyl phosphate or with protein catalyzed phosphorylation by the kinase EnvZ. The dephosphorylation rate of phospho-OmpR is affected only slightly by the presence of DNA; thus, the increased level is largely caused by an increased rate of phosphorylation. Stimulation of phosphorylation requires specific binding of DNA by OmpR. Occupancy of the DNA binding domain exposes a trypsin cleavage site in the linker, which connects the phosphorylation domain with the DNA binding domain. Our results indicate that when DNA binds in the C terminus, it enhances phosphorylation in the N terminus, and the linker undergoes a conformational change. A generalized mechanism involving a four-state model for response regulators is proposed. PMID- 10518530 TI - In vitro synthesis of the nucleotide loop of cobalamin by Salmonella typhimurium enzymes. AB - In Salmonella typhimurium, the CobU, CobS, CobT, and CobC proteins have been proposed to catalyze the late steps in adenosylcobalamin biosynthesis, which define the nucleotide loop assembly pathway. This paper reports the in vitro assembly of the nucleotide loop of adenosylcobalamin from its precursors adenosylcobinamide, 5, 6-dimethylbenzimidazole, nicotinate mononucleotide, and GTP. Incubation of these precursors with the CobU, CobS, and CobT proteins resulted in the synthesis of adenosylcobalamin-5'-phosphate. This cobamide was isolated by HPLC, identified by UV-visible spectroscopy and mass spectrometry, and shown to support growth of a cobalamin auxotroph. Adenosylcobalamin-5' phosphate was also isolated from reaction mixtures containing adenosylcobinamide GDP (the product of the CobU reaction) and alpha-ribazole-5'-phosphate (the product of the CobT reaction) as substrates and CobS. These results allowed us to conclude that CobS is the cobalamin(-5'-phosphate) synthase enzyme in S. typhimurium. The CobC enzyme, previously shown to dephosphorylate alpha-ribazole 5'-phosphate to alpha-ribazole, was shown to dephosphorylate adenosylcobalamin-5' phosphate to adenosylcobalamin. Adenosylcobinamide was converted to adenosylcobalamin in reactions where all four enzymes were present in the reaction mixture. This in vitro system offers a unique opportunity for the rapid synthesis and isolation of cobamides with structurally different lower-ligand bases that can be used to investigate the contributions of the lower-ligand base to cobalamin-dependent reactions. PMID- 10518531 TI - Double duplex invasion by peptide nucleic acid: a general principle for sequence specific targeting of double-stranded DNA. AB - Pseudocomplementary PNAs containing diaminopurine.thiouracil base pairs have been prepared and are shown to bind with high specificity and efficiency to complementary targets in double-stranded DNA by a mechanism termed "double duplex invasion" in which the duplex is unwound and both DNA strands are targeted simultaneously, each by one of the two pseudocomplementary peptide nucleic acids (PNAs). On the basis of our results we predict that (for decameric targets) more than 80% of all sequences can be targeted by straightforward Watson-Crick base pairing by using this approach in its present form. Targeting of pseudocomplementary PNAs to the promoter of the T7 phage RNA polymerase effectively inhibits transcription initiation. These results have important implications in the development of gene therapeutic agents as well as for genetic diagnostic and molecular biology applications. PMID- 10518532 TI - A structural snapshot of base-pair opening in DNA. AB - The response of double-helical DNA to torsional stress may be a driving force for many processes acting on DNA. The 1.55-A crystal structure of a duplex DNA oligonucleotide d(CCAGGCCTGG)(2) with an engineered crosslink in the minor groove between the central guanine bases depicts how the duplex can accommodate such torsional stress. We have captured in the same crystal two rather different conformational states. One duplex contains a strained crosslink that is stabilized by calcium ion binding in the major groove, directly opposite the crosslink. For the other duplex, the strain in the crosslink is relieved through partial rupture of a base pair and partial extrusion of a cytosine accompanied by helix bending. The sequence used is the target sequence for the HaeIII methylase, and this partially flipped cytosine is the same nucleotide targeted for extrusion by the enzyme. Molecular dynamics simulations of these structures show an increased mobility for the partially flipped-out cytosine. PMID- 10518533 TI - Interactions of G(h)/transglutaminase with phospholipase Cdelta1 and with GTP. AB - The inositol phosphate hydrolyzing activity of human phospholipase Cdelta1 (PLCdelta1) is markedly inhibited when the enzyme is coexpressed with the human heart G(h)/transglutaminase (TG) in human embryonic kidney cells. Because the cotransfection does not affect the amount of PLCdelta1 in the cells, the depression of phospholipase activity probably is a result of a direct interaction between the two proteins. An ELISA procedure was employed to document the associations of purified TG preparations from a variety of tissues (human red cells, rabbit lens, guinea pig liver) with PLCdelta1. Nucleotides (GTP > GDP > ATP > GMP = ADP, in order of decreasing efficiency) interfered with the formation of the PLCdelta1:TG complex. A conformational change in the TG partner, occurring with nucleotide binding, is thought to be responsible for dissociating the two proteins. The structural rearrangement produces a remarkable shift in the anodic mobility of TG in electrophoresis: TG(slow) + GTP -->/<-- [TG:GTP](fast). Altogether, our findings indicate that GTP controls PLCdelta1 activity by releasing this protein from an inhibitory association with G(h)/transglutaminase. PMID- 10518534 TI - Direct molecular force measurements of multiple adhesive interactions between cadherin ectodomains. AB - Direct-force measurements of the interactions between recombinant C-cadherin from Xenopus demonstrated that the ectodomain of cadherin exhibits multiple adhesive contacts that involve successive domains along the extracellular region of the protein. Contacts between the fully interdigitated antiparallel proteins form the strongest adhesive interaction. A second weaker minimum was measured when the interdigitated proteins were separated by a distance equal to the length of one domain of the extracellular (EC) fragment and corresponding to the antiparallel alignment of domains one through four (EC1 through EC4). The successive rupture of these interactions generates an unbinding force profile that may be optimized to impede the abrupt failure of cadherin-mediated junctions under force. PMID- 10518535 TI - A model of the detection of warmth and cold by cutaneous sensors through effects on voltage-gated membrane channels. AB - Warmth and cold sensations are known to derive from separate warm and cold cutaneous thermoreceptors in the form of differentiated afferent nerves. The firing rate of warm-sensing nerves increases as the temperature increases; the firing rate of cold-sensing nerves increases if the temperature is reduced. I postulate that the primary sensitivity of the warm sensors derives from voltage gated Ca(2+) membrane channels configured such that an increase in temperature opens channels and increases the ion influx while a reduction in temperature increases the ion influx through voltage-gated Na(+) channels in the cold sensory nerve ends. In either case, the initial cation influx causes a small cellular depolarization that further opens Ca(2+) channels, admitting more cations in a positive feedback process that leads to the depolarization of the membrane, thus initiating an action potential pulse. Monte Carlo calculations based on a well defined model of such processes, which include effects of noise, demonstrate quantitative agreement of the model with an extensive body of data. PMID- 10518536 TI - Role of the integrin-associated protein CD9 in binding between sperm ADAM 2 and the egg integrin alpha6beta1: implications for murine fertilization. AB - CD9 is a tetraspan protein that associates with several beta1 integrins, including alpha6beta1. Because alpha6beta1 is present on murine eggs and interacts with the sperm-surface glycoprotein ADAM 2 (fertilin beta), we first asked whether CD9 is present on murine eggs and whether it functions in sperm-egg binding and fusion. CD9 is present on the plasma membrane of oocytes in the ovary as well as on eggs isolated from the oviduct. The anti-CD9 mAb, JF9, potently inhibits sperm-egg binding and fusion in vitro in a dose-dependent manner. JF9 also disrupts binding of fluorescent beads coated with native fertilin or a recombinant fertilin beta disintegrin domain. (Both ligands bind to the egg via alpha6beta1.) Immunohistochemistry showed that CD9 is undetectable in the uterine epithelium, appears basolaterally and as prominent apical patches on the epithelium in the region between the uterus and the oviduct, and then persists apically in the oviduct. The integrin alpha6A subunit is found in similar apical patches in the region between the uterus and oviduct, but is confined to the basal aspect of the epithelium in the uterus and oviduct. Hence, alpha6A and CD9 both are expressed on the apical epithelial surface at the uterine-oviduct junction. These findings correlate with the observation that fertilin beta "knockout" sperm traverse the uterus but do not progress into the oviduct, contributing to the infertility of fertilin beta(-/-) male mice. Our results suggest that high-avidity binding between fertilin beta (ADAM 2) and alpha6beta1 requires cooperation between alpha6beta1 and CD9. Such cooperation may assist sperm passage into the oviduct as well as sperm-egg interactions. PMID- 10518538 TI - Optical recording of signal-mediated protein transport through single nuclear pore complexes. AB - Optical single-transporter recording, a recently established fluorescence microscopic method, was used to study the selective transport of proteins through single nuclear pore complexes (NPCs) of Xenopus oocytes. Recombinant proteins containing either a nuclear localization signal (import protein) or a nuclear export signal (export protein) were generated as transport substrates. To approximate in vivo conditions as closely as possible, a Xenopus egg extract was applied to the cytosolic side and a Xenopus oocyte nuclear extract to the nuclear side of the NPCs. It was found that protein transport through functionally isolated, "patched" NPCs depended on signal sequences, extracts, and metabolic energy, as in vivo. All NPCs were competent for both import and export. The transport direction was strictly determined by the transport signal, and at none of the conditions explored was the import protein exported or the export protein imported, even when the application sides of the extracts were reversed. The mean transport rates of the single NPC were approximately 2 dimers/s for the import protein and approximately 4 dimers/s for the export protein ( approximately 15 microM substrate concentration, 22-24 degrees C), in good agreement with in vivo rates estimated for mammalian cells by microinjection experiments. The study shows that optical single-transporter recording permits the analysis of membrane transport processes not previously accessible to single-transporter recording and thus provides additional possibilities for the elucidation of nucleocytoplasmic transport mechanisms. PMID- 10518537 TI - Regulation of nuclear translocation of forkhead transcription factor AFX by protein kinase B. AB - The regulation of intracellular localization of AFX, a human Forkhead transcription factor, was studied. AFX was recovered as a phosphoprotein from transfected COS-7 cells growing in the presence of FBS, and the phosphorylation was eliminated by wortmannin, a potent inhibitor of phosphatidylinositol (PI) 3 kinase. AFX was phosphorylated in vitro by protein kinase B (PKB), a downstream target of PI 3-kinase, but a mutant protein in which three putative phosphorylation sites of PKB had been replaced by Ala was not recognized by PKB. In Chinese hamster ovary cells (CHO-K1) cultured with serum, the AFX protein fused with green fluorescence protein (AFX-GFP) is localized mainly in the cytoplasm, and wortmannin induced transient nuclear translocation of the fusion protein. The AFX-GFP mutant in which all three phosphorylation sites had been replaced by Ala was detected exclusively in the cell nucleus. AFX-GFP was in the nucleus when the cells were infected with an adenovirus vector encoding a dominant-negative form of either PI 3-kinase or PKB, whereas the fusion protein stayed in the cytoplasm when the cells expressed constitutively active PKB. In CHO-K1 cells expressing AFX-GFP, DNA fragmentation was induced by the stable PI 3 kinase inhibitor LY294002, and the expression of the active form of PKB suppressed this DNA fragmentation. The phosphorylation site mutant of AFX-GFP enhanced DNA fragmentation irrespective of the presence and absence of PI 3 kinase inhibitor. These results indicate that the nuclear translocation of AFX is negatively regulated through its phosphorylation by PKB. PMID- 10518539 TI - The combined absence of the transcription factors Rel and RelA leads to multiple hemopoietic cell defects. AB - Individual Rel/NF-kappaB transcription factors, although dispensable for the development and maturation of most hemopoietic cells, are critical regulators of normal immune function. Redundancy among these proteins prompted us to examine the role of Rel and RelA in hemopoiesis by using mice that lack both subunits. Because of the death of double-mutant fetuses at day 13.5 of gestation (E13.5), E12 fetal liver hemopoietic progenitors were used for in vitro cultures and for repopulating stem cell studies in lethally irradiated normal recipient mice. Most striking, Rel/RelA-deficient hemopoietic precursors failed to promote the survival of myeloablated mice. This phenotype was associated with several defects including a reduction of spleen colony-forming unit progenitors, impaired erythropoiesis, and a deregulated expansion of granulocytes. In vitro progenitor assays also revealed that Rel or RelA serves an antiapoptotic role during monocyte differentiation. Despite the combined loss of Rel and RelA leading to these hemopoietic defects, c-rel(-/-)rela(-/-) stem cells contributed to the development of all lineages in mice engrafted with double-mutant fetal liver cells and normal bone marrow cells, albeit in a reduced fashion compared with controls. Collectively, these data indicate the loss of Rel and RelA does not appear to affect pluripotent stem cells; rather, Rel and RelA serve redundant functions in regulating differentiation and survival of committed progenitors in multiple hemopoietic lineages. PMID- 10518540 TI - beta2-chimaerin is a novel target for diacylglycerol: binding properties and changes in subcellular localization mediated by ligand binding to its C1 domain. AB - The members of the chimaerin family of Rac-GTPase-activating proteins possess a single C1 domain with high homology to those present in protein kinase C (PKC) isozymes. This domain in PKCs is involved in phorbol ester and diacylglycerol (DAG) binding. We previously have demonstrated that one of the chimaerin isoforms, beta2-chimaerin, binds phorbol esters with high affinity. In this study we analyzed the properties of beta2-chimaerin as a DAG receptor by using a series of conformationally constrained cyclic DAG analogues (DAG lactones) as probes. We identified analogs that bind to beta2-chimaerin with more than 100-fold higher affinity than 1-oleoyl-2-acetylglycerol. The potencies of these analogs approach those of the potent phorbol ester tumor promoters. The different DAG lactones show some selectivity for this novel receptor compared with PKCalpha. Cellular studies revealed that these DAG analogs induce translocation of beta2-chimaerin from cytosolic (soluble) to particulate fractions. Using green fluorescent protein-fusion proteins for beta2-chimaerin we determined that this novel receptor translocates to the perinuclear region after treatment with DAG lactones. Binding and translocation were prevented by mutation of the conserved Cys-246 in the C1 domain. The structural homology between the C1 domain of beta2 chimaerin and the C1b domain of PKCdelta also was confirmed by modeling analysis. Our results demonstrate that beta2-chimaerin is a high affinity receptor for DAG through binding to its C1 domain and supports the emerging concept that multiple pathways transduce signaling through DAG and the phorbol esters. PMID- 10518541 TI - Inhibitory role of the host apoptogenic gene PKR in the establishment of persistent infection by encephalomyocarditis virus in U937 cells. AB - Persistent infections by viruses such as HIV-1 and hepatitis B virus can pose long-term health hazards. Because establishment of persistent infections involves close interactions and adjustments in both host and virus, it would be informative to establish a paradigm with which a normally cytolytic viral infection can be easily converted to persistent infection, so that the different stages in developing persistent infection can be examined. Such a model system is described in this paper. Highly cytolytic encephalomyocarditis virus (EMCV) infection was shifted to persistent infection as a result of repressed expression of the double-stranded RNA-dependent protein kinase (PKR) in the promonocytic U937 cells. Because of the apoptogenic potential of PKR, a deficiency of PKR resulted in a delay in virus-induced apoptosis in EMCV-infected U937 cells, allowing the eventual establishment of persistent EMCV infection in these cells (U9K-AV2). That this was a bona fide persistent infection was demonstrated by the ability of infected cells to propagate as long-term virus-shedding cultures; electron microscopy studies showing presence of intracellular EMCV virions and chromatin condensation; detection of virus-induced chromosomal DNA fragmentation and sustained expression of apoptogenic p53 and IL-1beta converting enzyme; and demonstration of active EMCV transcription by reverse transcription-PCR. In addition, a host-virus coevolution was observed in U9K-AV2 cultures over time: U9K-AV2 cells exhibited slower growth rates, resistance to viral super-infection, and cessation of IFN-alpha synthesis, whereas the infectivity of EMCV was drastically attenuated. Finally, data are presented on the suitability of this model to study establishment of persistent infection by other viruses such as Sendai virus and reovirus. PMID- 10518542 TI - BRCA1-associated growth arrest is RB-dependent. AB - BRCA1 is a susceptibility gene for breast and ovarian cancer with growth inhibitory activity for which the mechanism of action remains unclear. When introduced into cells, BRCA1 inhibits growth of some but not all cell lines. In an attempt to uncover the mechanism of growth suppression by BRCA1, we examined a panel of cell lines for their ability to reduce colony outgrowth in response to BRCA1 overexpression. Of all variables tested, only those cells with wild-type pRb were sensitive to BRCA1-induced growth suppression. In cells with an intact rb gene, inactivation of pRb by HPV E7 abrogates the growth arrest imposed by BRCA1. In accordance with these observations, we found that BRCA1 could not suppress BrdUrd uptake in primary fibroblasts from rb-/- mice and exhibited an intermediate ability to inhibit DNA synthesis in rb+/- as compared with rb+/+ cells. We further found that the BRCA1 protein complexes with the hypophosphorylated form of pRb. This binding is localized to amino acids 304-394 of BRCA1 protein and requires the ABC domain of pRb. In-frame deletion of BRCA1 fragment involved in interaction with pRb completely abolished the growth suppressive property of BRCA1. Although it has been reported that BRCA1 interacts with p53, we find the p53 status did not affect the ability of BRCA1 to suppress colony formation. Our data suggest that the growth suppressor function of BRCA1 depends, at least in part, on Rb. PMID- 10518543 TI - Presenilin 2 deficiency causes a mild pulmonary phenotype and no changes in amyloid precursor protein processing but enhances the embryonic lethal phenotype of presenilin 1 deficiency. AB - Mutations in the homologous presenilin 1 (PS1) and presenilin 2 (PS2) genes cause the most common and aggressive form of familial Alzheimer's disease. Although PS1 function and dysfunction have been extensively studied, little is known about the function of PS2 in vivo. To delineate the relationships of PS2 and PS1 activities and whether PS2 mutations involve gain or loss of function, we generated PS2 homozygous deficient (-/-) and PS1/PS2 double homozygous deficient mice. In contrast to PS1(-/-) mice, PS2(-/-) mice are viable and fertile and develop only mild pulmonary fibrosis and hemorrhage with age. Absence of PS2 does not detectably alter processing of amyloid precursor protein and has little or no effect on physiologically important apoptotic processes, indicating that Alzheimer's disease-causing mutations in PS2, as in PS1, result in gain of function. Although PS1(+/-) PS2( -/-) mice survive in relatively good health, complete deletion of both PS2 and PS1 genes causes a phenotype closely resembling full Notch-1 deficiency. These results demonstrate in vivo that PS1 and PS2 have partially overlapping functions and that PS1 is essential and PS2 is redundant for normal Notch signaling during mammalian embryological development. PMID- 10518544 TI - Imino sugars inhibit the formation and secretion of bovine viral diarrhea virus, a pestivirus model of hepatitis C virus: implications for the development of broad spectrum anti-hepatitis virus agents. AB - One function of N-linked glycans is to assist in the folding of glycoproteins by mediating interactions of the lectin-like chaperone proteins calnexin and calreticulin with nascent glycoproteins. These interactions can be prevented by inhibitors of the alpha-glucosidases, such as N-butyl-deoxynojirimycin (NB-DNJ) and N-nonyl-DNJ (NN-DNJ), and this causes some proteins to be misfolded and retained within the endoplasmic reticulum (ER). We have shown previously that the NN-DNJ-induced misfolding of one of the hepatitis B virus (HBV) envelope glycoproteins prevents the formation and secretion of virus in vitro and that this inhibitor alters glycosylation and reduces the viral levels in an animal model of chronic HBV infection. This led us to investigate the effect of glucosidase inhibitors on another ER-budding virus, bovine viral diarrhea virus, a tissue culture surrogate of human hepatitis C virus (HCV). Here we show that in MDBK cells alpha-glucosidase inhibitors prevented the formation and secretion of infectious bovine viral diarrhea virus. Data also are presented showing that NN DNJ, compared with NB-DNJ, exhibits a prolonged retention in liver in vivo. Because viral secretion is selectively hypersensitive to glucosidase inhibition relative to the secretion of cellular proteins, the possibility that glucosidase inhibitors could be used as broad-based antiviral hepatitis agents is discussed. A single drug against HBV, HCV, and, possibly, HDV, which together chronically infect more than 400 million people worldwide, would be of great therapeutic value. PMID- 10518545 TI - Direct visualization of the elt-2 gut-specific GATA factor binding to a target promoter inside the living Caenorhabditis elegans embryo. AB - In analyzing the transcriptional networks that regulate development, one ideally would like to determine whether a particular transcription factor binds directly to a candidate target promoter inside the living embryo. Properties of the Caenorhabditis elegans elt-2 gene, which encodes a gut-specific GATA factor, have allowed us to develop such a method. We previously have shown, by means of ectopic expression studies, that elt-2 regulates its own promoter. To test whether this autoregulation is direct, we fused green fluorescent protein (GFP) close to the C terminus of elt-2 in a construct that contains the full elt-2 promoter and the full elt-2 zinc finger DNA binding domain; the construct is expressed correctly (i.e., only in the gut lineage) and is able to rescue the lethality of an elt-2 null mutant. Multicopy transgenic arrays of this rescuing elt-2::GFP construct were integrated into the genome and transgenic embryos were examined when the developing gut has 4-8 cells; the majority of these embryonic gut nuclei show two discrete intense foci of fluorescence. We interpret these fluorescent foci as the result of ELT-2::GFP binding directly to its own promoter within nuclei of the developing gut lineage. Numerous control experiments, both genetic and biochemical, all support this conclusion and support the specificity of the binding. The approach should be applicable to studying other transcription factors binding target promoters, all within the living C. elegans embryo. PMID- 10518546 TI - Oogenic function of the myogenic factor D-MEF2: negative regulation of the decapentaplegic receptor gene thick veins. AB - The myogenic factor D-MEF2 is required for the proper differentiation of muscle cells during Drosophila embryogenesis and the correct patterning of indirect flight muscles assembled during later metamorphosis. In addition to these essential myogenic functions, mutant D-mef2 adult females are weakly fertile and produce defective eggs. D-MEF2 is expressed in nurse and follicle cells of the wild-type egg chamber. We have analyzed the D-mef2 oogenic phenotype and show that the gene is required for the normal patterning and differentiation of the centripetally migrating follicle cells that are crucial for development of the anterior chorionic structures. D-mef2 alleles exhibit a genetic interaction with a dominant-negative allele of thick veins (tkv), which encodes a type I receptor of the Decapentaplegic-signaling pathway. tkv RNA is overexpressed in D-mef2 mutant egg chambers, and, conversely, forced expression of D-mef2 represses tkv expression. These results indicate a role for D-MEF2 in the regulation of tkv gene expression and Decapentaplegic signal transduction that are essential for proper determination and/or differentiation of the anterior follicle cells. Additionally, they demonstrate a vital function for the D-MEF2 transcription factor in multiple genetic pathways during Drosophila development. PMID- 10518547 TI - Fgfr2 is required for limb outgrowth and lung-branching morphogenesis. AB - The aim of this study was to clarify the role of Fgfr2 during later stages of embryonic development. Of two previously reported gene-targeting experiments, the more extensive Fgfr2 deletion was lethal shortly after implantation, because of trophoblast defects, whereas the less extensive one survived until midgestation with placental insufficiency and defective limb outgrowth [Xu, X., Weinstein, M., Li, C., Naski, M., Cohen, R. I., Ornitz, D. M., Leder, P. & Deng, C. (1998) Development (Cambridge, U.K.) 125, 753-765]. Fgfr2 in the early embryo is expressed in the trophectoderm, and this extra-embryonic localization persists into mid- and late gestation, when Fgfr2 also is expressed in multiple developing organs. To gain insight into the later functions of Fgfr2, fusion chimeras were constructed from homozygous mutant embryonic stem cells and wild-type tetraploid embryos. This allowed survival until term and revealed that Fgfr2 is required for both limb outgrowth and branching lung morphogenesis. The use of fusion chimeras demonstrated that early lethality was indeed because of trophectoderm defects and indicated that in the embryonic cell lineages Fgfr2 activity manifests in limb and lung development. Highly similar lung and limb phenotypes were detected recently in the loss of function mutation of Fgf10, a ligand of Fgfr2. It is likely, therefore, that whereas during early development Fgfr2 interacts with Fgf4, in limb and lung development interactions between Fgf10 and Fgfr2 may be required. Possible epithelial-mesenchymal interactions between the splicing alternatives of Fgfr2 and their specific ligands will be discussed. PMID- 10518548 TI - Reverse homeosis in homeotically reconstructed ribbonworms. AB - Homeosis is the replacement of one body part by another, which may be caused by either developmental or genetic variations. It is particularly obvious in segmented animals, like insects, in which one body segment may be transformed into another. However, homeosis also occurs in animals without overt segmentation that also have detailed positional information specifying their body plan. By grafting, we have artificially generated homeotic ribbonworms of the species Lineus ruber with a duplicated ocellar region replacing the postocellar region anterior to the brain. Such chimeric animals are capable of complete morphogenetic regulation of the anterior-posterior (A-P) pattern. The missing postocellar region is restored by intercalary regeneration, and the anterior duplicated ocellar region is eliminated by a process called transgeneration. Thus, homeosis is reversed, and a completely normal pattern along the A-P axis is restored. This reverse homeosis involves the elimination of the syngeneic eyes and the survival of the grafted allogeneic eye region. LsPax-6, the Lineus sanguineus ortholog of the mammalian Pax-6 gene, which is considered to be a master control gene for eye morphogenesis, is expressed specifically in regenerating, regenerated, and intact eye regions. Our data show that ribbonworm eyes are either maintained or they regress according to their position along the A-P axis, even though there are no obvious segmental boundaries. This system allows us to test the function of LsPax-6 protein not only during eye regeneration but also during maintenance and regression of the eyes. PMID- 10518549 TI - Individuality and adaptation across levels of selection: how shall we name and generalize the unit of Darwinism? AB - Two major clarifications have greatly abetted the understanding and fruitful expansion of the theory of natural selection in recent years: the acknowledgment that interactors, not replicators, constitute the causal unit of selection; and the recognition that interactors are Darwinian individuals, and that such individuals exist with potency at several levels of organization (genes, organisms, demes, and species in particular), thus engendering a rich hierarchical theory of selection in contrast with Darwin's own emphasis on the organismic level. But a piece of the argument has been missing, and individuals at levels distinct from organisms have been denied potency (although granted existence within the undeniable logic of the theory), because they do not achieve individuality with the same devices used by organisms and therefore seem weak by comparison. We show here that different features define Darwinian individuality across scales of size and time. In particular, species-individuals may develop few emergent features as direct adaptations. The interactor approach works with emergent fitnesses, not with emergent features; and species, as a consequence of their different mechanism for achieving individuality (reproductive exclusivity among subparts, that is, among organisms), express many effects from other levels. Organisms, by contrast, suppress upwardly cascading effects, because the organismic style of individuality (by functional integration of subparts) does not permit much competition or differential reproduction of parts from within. Species do not suppress the operation of lower levels; such effects therefore become available as exaptations conferring emergent fitness-a primary source of the different strength that species achieve as effective Darwinian individuals in evolution. PMID- 10518550 TI - Pollinator preference and the evolution of floral traits in monkeyflowers (Mimulus). AB - A paradigm of evolutionary biology is that adaptation and reproductive isolation are caused by a nearly infinite number of mutations of individually small effect. Here, we test this hypothesis by investigating the genetic basis of pollinator discrimination in two closely related species of monkeyflowers that differ in their major pollinators. This system provides a unique opportunity to investigate the genetic architecture of adaptation and speciation because floral traits that confer pollinator specificity also contribute to premating reproductive isolation. We asked: (i) What floral traits cause pollinator discrimination among plant species? and (ii) What is the genetic basis of these traits? We examined these questions by using data obtained from a large-scale field experiment where genetic markers were employed to determine the genetic basis of pollinator visitation. Observations of F2 hybrids produced by crossing bee-pollinated Mimulus lewisii with hummingbird-pollinated Mimulus cardinalis revealed that bees preferred large flowers low in anthocyanin and carotenoid pigments, whereas hummingbirds favored nectar-rich flowers high in anthocyanins. An allele that increases petal carotenoid concentration reduced bee visitation by 80%, whereas an allele that increases nectar production doubled hummingbird visitation. These results suggest that genes of large effect on pollinator preference have contributed to floral evolution and premating reproductive isolation in these monkeyflowers. This work contributes to growing evidence that adaptation and reproductive isolation may often involve major genes. PMID- 10518551 TI - Markov chain Monte Carlo analysis of human Y-chromosome microsatellites provides evidence of biased mutation. AB - We describe a Markov Chain Monte Carlo analysis of five human Y- chromosome microsatellite polymorphisms based on samples from five diverse populations. Our analysis provides strong evidence for mutational bias favoring increase in length at all loci. Estimates of population coalescent times and population size from our two largest samples, one African and one European, suggest that the African population is older but smaller and that the English East Anglian population has undergone significant expansion, being larger but younger. We conclude that Markov Chain Monte Carlo analysis of microsatellite haplotypes can uncover information not apparent when the microsatellites are considered independently. Incorporation of population size as a variable should allow us to estimate the timing and magnitude of major historical population trends. PMID- 10518552 TI - Human and mouse homologs of Escherichia coli DinB (DNA polymerase IV), members of the UmuC/DinB superfamily. AB - To understand the mechanisms underlying mutagenesis in eukaryotes better, we have cloned mouse and human homologs of the Escherichia coli dinB gene. E. coli dinB encodes DNA polymerase IV and greatly increases spontaneous mutations when overexpressed. The mouse and human DinB1 amino acid sequences share significant identity with E. coli DinB, including distinct motifs implicated in catalysis, suggesting conservation of the polymerase function. These proteins are members of a large superfamily of DNA damage-bypass replication proteins, including the E. coli proteins UmuC and DinB and the Saccharomyces cerevisiae proteins Rev1 and Rad30. In a phylogenetic tree, the mouse and human DinB1 proteins specifically group with E. coli DinB, suggesting a mitochondrial origin for these genes. The human DINB1 gene is localized to chromosome 5q13 and is widely expressed. PMID- 10518553 TI - In vivo suppressor mutations correct a murine model of hereditary tyrosinemia type I. AB - Hereditary tyrosinemia type I and alkaptonuria are disorders of tyrosine catabolism caused by deficiency of fumarylacetoacetate hydrolase (FAH) and homogentisic acid dioxygenase (HGD), respectively. Tyrosinemia is a severe childhood disease that affects the liver and kidneys, but alkaptonuria is a more benign adult disorder in comparison. Because HGD is upstream of FAH in the tyrosine pathway, mice doubly mutant in both enzymes were found to be protected from the liver and renal damage of tyrosinemia as hypothesized. Mice mutant at the tyrosinemic locus but heterozygous for alkaptonuria spontaneously developed clonal nodules of functionally normal hepatocytes that were able to rescue the livers of some mice with this genotype. This phenotypic rescue was a result of an inactivating mutation of the wild-type homogentisic acid dioxygenase gene, thus presenting an example of an in vivo suppressor mutation in a mammalian model. PMID- 10518554 TI - The yeast HML I silencer defines a heterochromatin domain boundary by directional establishment of silencing. AB - The eukaryotic genome is divided into functional domains defined in part by local differences in chromatin structure and delimited in many cases by boundary elements. The HML and HMR loci in the yeast Saccharomyces cerevisiae are transcriptionally silent chromosome domains. Each locus is bracketed by two cis acting sequences, designated E and I, that serve to establish and maintain repression of genes within each locus. We show that repression at HML is uniformly high between E and I but decreases sharply beyond I. The region of repression at HML generally correlates with the domain of histone hypoacetylation. Despite the sharp definition of the boundaries of HML, no sequence capable of blocking the spread of heterochromatin resides in the sequences flanking HML. We find, though, that inverting the orientation of I increases silencing outside of HML while weakening silencing within HML. These results indicate that the HML I silencer establishes a boundary between active and inactive chromatin at HML, but does so by organizing inactive chromatin in only one direction. This represents a different mechanism for delimiting the boundaries of a eukaryotic chromosome domain. PMID- 10518555 TI - Drosophila proteins related to vertebrate DNA (5-cytosine) methyltransferases. AB - DNA methylation at CpG residues is closely associated with a number of biological processes during vertebrate development. Unlike the vertebrates, however, several invertebrate species, including the Drosophila, do not have apparent DNA methylation in their genomes. Nor have there been reports on a DNA (5-cytosine) methyltransferase (CpG MTase) found in these invertebrates. We now present evidence for two CpG MTase-like proteins expressed in Drosophila cells. One of these, DmMTR1, is a protein containing peptide epitopes immunologically related to the conserved motifs I and IV in the catalytic domain of the mammalian dnmt1. DmMTR1 has an apparent molecular mass of 220 kDa and, similar to mammalian dnmt1, it also interacts in vivo with the proliferating cell nuclear antigen. During interphase of the syncytial Drosophila embryos, the DmMTR1 molecules are located outside the nuclei, as is dnmt1 in the mouse blastocyst. However, DmMTR1 appears to be rapidly transported into, and then out of the nuclei again, as the embryos undergo mitotic waves. Immunofluorescent data indicate that DmMTR1 molecules "paint" the whole set of condensed Drosophila chromosomes throughout the mitotic phase, suggesting they may play an essential function in the cell-cycle regulated condensation of the Drosophila chromosomes. Through search in the genomic database, we also have identified a Drosophila polypeptide, DmMT2, that exhibits high sequence homology to the mammalian dnmt2 and the yeast CpG MTase homolog pmt1. The expression of DmMT2 appears to be developmentally regulated. We discuss the evolutionary and functional implications of the discovery of these two Drosophila proteins related to mammalian CpG MTases. PMID- 10518556 TI - Disruption of the murine nuclear factor I-A gene (Nfia) results in perinatal lethality, hydrocephalus, and agenesis of the corpus callosum. AB - The phylogenetically conserved nuclear factor I (NFI) family of transcription/replication proteins is essential both for adenoviral DNA replication and for the transcription of many cellular genes. We showed previously that the four murine NFI genes (Nfia, Nfib, Nfic, and Nfix) are expressed in unique but overlapping patterns during mouse development and in adult tissues. Here we show that disruption of the Nfia gene causes perinatal lethality, with >95% of homozygous Nfia(-/-) animals dying within 2 weeks after birth. Newborn Nfia(-/-) animals lack a corpus callosum and show ventricular dilation indicating early hydrocephalus. Rare surviving homozygous Nfia(-/-) mice lack a corpus callosum, show severe communicating hydrocephalus, a full-axial tremor indicative of neurological defects, male-sterility, low female fertility, but near normal life spans. These findings indicate that while the Nfia gene appears nonessential for cell viability and DNA replication in embryonic stem cells and fibroblasts, loss of Nfia function causes severe developmental defects. This finding of an NFI gene required for a developmental process suggests that the four NFI genes may have distinct roles in vertebrate development. PMID- 10518557 TI - Differential effect of an Ig mu transgene on development of pre-B cells in fetal and adult SCID mice. AB - Progression of pro-B lymphocytes to the pre-B stage depends on the expression of a pre-B cell receptor (pre-BCR), consisting of an Ig mu H chain, Ig surrogate light chain, and associated signal transducing chains. Mice that are unable to express a pre-BCR show an arrest of B cell development at the pro-B stage. Such is the case for severe combined immune deficient (SCID) mice in which mu chains are not made because of a defect in V(D)J recombination. When mu chains are made, as in SCID mice bearing a functional mu transgene, then B cell differentiation can proceed to the pre-B stage. However, as reported here, a mu transgene (M54) that promotes development of SCID pre-B cells in adult bone marrow fails to do so in fetal liver. We suggest that a pre-BCR containing the M54 mu chain cannot signal progression of pro-B cells to the pre-B stage in the fetal liver microenvironment. PMID- 10518558 TI - Productive infection of double-negative T cells with HIV in vivo. AB - HIV induces CD4 down-regulation from the surface of infected cells by several independent mechanisms, suggesting an important biological role for this phenomenon. In vitro CD4 down-regulation generates T cells with a double-negative (DN) CD4(-)CD8(-) T cell receptor-alphabeta(+) phenotype. However, evidence that this down-regulation occurs in vivo in HIV-infected subjects is lacking, and viral load or viral production assays invariably focus on CD4(+) T cells. We show here that HIV infection can often be detected in sorted DN cells from peripheral blood and lymph nodes, even when plasma viral load is undetectable. DN T cells infected with HIV represented up to 20% of the cellular viral load in T cells, as determined by DNA PCR. In patients on successful highly active antiretroviral therapy, the viral load decreased in the plasma in CD4(+) and in DN T cells, suggesting that infected DN cells, like CD4(+) cells, contribute to viral production and are sensitive to highly active antiretroviral therapy. Indeed, HIV unspliced and multispliced RNAs were often detectable in DN T cells in spite of the small size of this subset. Infectious virus from DN T cells was transmitted efficiently in coculture experiments with uninfected T cell lymphoblasts, even when viral DNA in the DN cells was barely detectable. We conclude that a discrete population of infected DN T cells exists in HIV-positive subjects, even when the plasma viral load is undetectable. These cells may represent an important source of infectious virus. PMID- 10518559 TI - Persistence of lymphocytic choriomeningitis virus at very low levels in immune mice. AB - Lymphocytic choriomeningitis virus (LCMV), strain WE, is a non-cytopathic RNA virus that is highly adapted to its natural host, the mouse. Acute infection of adult mice leads to generalized virus spread, followed by cytotoxic T lymphocyte mediated virus clearance below the detection levels of conventional assays within 2-3 weeks. Indirect evidence had suggested that virus or viral antigen might persist in the immune mouse. Here we demonstrate LCMV-WE persistence at low levels after infection with 10(2) or 10(6) plaque-forming units, shown as viral genome, viral antigen, and replicative virus using sensitive in vitro and in vivo assays. The finding that LCMV-WE persists in the face of apparently intact immune responses resembles the situation in some viral (hepatitis B and C, HIV) and bacterial (tuberculosis, leprosy) infections in humans; the results are relevant to the understanding not only of other murine and human persistent viral infections but also of protective immunological memory by "infection immunity." PMID- 10518560 TI - Flanking nuclear matrix attachment regions synergize with the T cell receptor delta enhancer to promote V(D)J recombination. AB - Previous studies have identified nuclear matrix attachment regions (MARs) that are closely associated with transcriptional enhancers in the IgH, Igkappa, and T cell receptor (TCR) beta loci, but have yielded conflicting information regarding their functional significance. In this report, a combination of in vitro and in situ mapping approaches was used to localize three MARs associated with the human TCR delta gene. Two of these are located within the Jdelta3-Cdelta intron, flanking the core TCR delta enhancer (Edelta) both 5' and 3' in a fashion reminiscent of the Ig heavy chain intronic enhancer-associated MARs. The third is located about 20 kb upstream, tightly linked to Ddelta1 and Ddelta2. We have previously used a transgenic minilocus V(D)J recombination reporter to establish that Edelta functions as a developmental regulator of V(D)J recombination, and that it does so by modulating substrate accessibility to the V(D)J recombinase. We show here that the Edelta-associated MARs function synergistically with the core Edelta to promote V(D)J recombination in this system, as they are required for enhancer-dependent transgene rearrangement in single-copy transgene integrants. PMID- 10518561 TI - Molecular cloning of a docking protein, BRDG1, that acts downstream of the Tec tyrosine kinase. AB - Tec, Btk, Itk, Bmx, and Txk constitute the Tec family of protein tyrosine kinases (PTKs), a family with the distinct feature of containing a pleckstrin homology (PH) domain. Tec acts in signaling pathways triggered by the B cell antigen receptor (BCR), cytokine receptors, integrins, and receptor-type PTKs. Although upstream regulators of Tec family kinases are relatively well characterized, little is known of the downstream effectors of these enzymes. The yeast two hybrid system has identified several proteins that interact with the kinase domain of Tec, one of which is now revealed to be a previously unknown docking protein termed BRDG1 (BCR downstream signaling 1). BRDG1 contains a proline-rich motif, a PH domain, and multiple tyrosine residues that are potential target sites for Src homology 2 domains. In 293 cells expressing recombinant BRDG1 and various PTKs, Tec and Pyk2, but not Btk, Bmx, Lyn, Syk, or c-Abl, induced marked phosphorylation of BRDG1 on tyrosine residues. BRDG1 was also phosphorylated by Tec directly in vitro. Efficient phosphorylation of BRDG1 by Tec required the PH and SH2 domains as well as the kinase domain of the latter. Furthermore, BRDG1 was shown to participate in a positive feedback loop by increasing the activity of Tec. BRDG1 transcripts are abundant in the human B cell line Ramos, and the endogenous protein underwent tyrosine phosphorylation in response to BCR stimulation. BRDG1 thus appears to function as a docking protein acting downstream of Tec in BCR signaling. PMID- 10518562 TI - Parvalbumin gene transfer corrects diastolic dysfunction in diseased cardiac myocytes. AB - Heart failure frequently involves diastolic dysfunction that is characterized by a prolonged relaxation. This prolonged relaxation is typically the result of a decreased rate of intracellular Ca(2+) sequestration. No effective treatment for this decreased Ca(2+) sequestration rate currently exists. As an approach to possibly correct diastolic dysfunction, we hypothesized that expression of the Ca(2+) binding protein parvalbumin in cardiac myocytes would lead to increased rates of Ca(2+) sequestration and mechanical relaxation. Parvalbumin, which is normally absent in cardiac tissue, is known to act as a soluble relaxing factor in fast skeletal muscle fibers by acting as a delayed Ca(2+) sink. As a test of the hypothesis, gene transfer was used to express parvalbumin in isolated adult cardiac myocytes. We report here that expression of parvalbumin dramatically increases the rate of Ca(2+) sequestration and the relaxation rate in normal cardiac myocytes. Importantly, parvalbumin fully restored the relaxation rate in diseased cardiac myocytes isolated from an animal model of human diastolic dysfunction. These findings indicate that parvalbumin gene transfer offers unique potential as a possible direct treatment for diastolic dysfunction in failing hearts. PMID- 10518563 TI - Spontaneous and antigen-induced production of HIV-inhibitory beta-chemokines are associated with AIDS-free status. AB - The beta-chemokines RANTES, macrophage inflammatory protein (MIP)-1alpha, and MIP 1beta suppress infection by macrophage-tropic strains of HIV and simian immunodeficiency virus (SIV) by binding and down-regulating the viral coreceptor, CCR5. Accordingly, we have examined whether higher levels of CCR5 ligands are associated with a more favorable clinical status in AIDS. A cross-sectional study of 100 subjects enrolled in the Multicenter AIDS Cohort Study at the Baltimore site was conducted to measure chemokine production and lymphocyte proliferation by peripheral blood mononuclear cells (PBMC). Statistical analyses of the data revealed that the production of HIV-suppressive beta-chemokines by HIV antigen stimulated PBMC was significantly higher in HIV-positive subjects without AIDS compared with subjects with clinical AIDS. Increased chemokine production was also correlated with higher proliferative responses to HIV antigens. Both parameters were significantly lower in the AIDS versus non-AIDS group. Notably, significantly higher levels of MIP-1alpha were also observed with unstimulated PBMC from seronegative subjects at risk for HIV infection released as compared with seropositive and non-Multicenter AIDS Cohort Study seronegative subjects. The association of chemokine production with antigen-induced proliferative responses, more favorable clinical status in HIV infection, as well as with an uninfected status in subjects at risk for infection suggests a positive role for these molecules in controlling the natural course of HIV infection. PMID- 10518564 TI - Cholesterol accumulation in tissues of the Niemann-pick type C mouse is determined by the rate of lipoprotein-cholesterol uptake through the coated-pit pathway in each organ. AB - Niemann-Pick type C (NPC) disease is associated with the accumulation of unesterified cholesterol in nearly all tissues and with progressive neurodegeneration. A murine model of this disease, the NPC mouse, was used to determine whether this sequestered cholesterol represented sterol carried in low density lipoprotein (LDL) and chylomicrons (CMs) taken up into the tissues through the coated-pit pathway. By 7 weeks of age, the sterol pool in the NPC mice had increased from 2,165 to 5,669 mg/kg body weight because of the daily sequestration of 67 mg of cholesterol per kg in the various organs. This was 7 fold greater than the rate of accumulation in control mice. The rate of LDL clearance in the NPC mouse was normal (523 ml/day per kg) and accounted for the uptake of 78 mg/day per kg of cholesterol in LDL whereas 8 mg/day per kg was taken up from CMs. Deletion of the LDL receptor in NPC mice altered the concentration of unesterified cholesterol in every organ in a manner consistent with the changes also observed in the rate of LDL cholesterol uptake in those tissues. Similarly, altering the flow of cholesterol to the liver through the CM pathway changed the concentration of unesterified cholesterol in that organ. Together, these observations strongly support the conclusion that, in NPC disease, it is cholesterol carried in LDL and CMs that is sequestered in the tissues and not sterol that is newly synthesized and carried in high density lipoprotein. PMID- 10518565 TI - Impaired growth and fertility of cAMP-specific phosphodiesterase PDE4D-deficient mice. AB - In eukaryotic cells, the inactivation of the cyclic nucleotide signal depends on a complex array of cyclic nucleotide phosphodiesterases (PDEs). Although it has been established that multiple PDE isoenzymes with distinct catalytic properties and regulations coexist in the same cell, the physiological significance of this remarkable complexity is poorly understood. To examine the role of a PDE in cAMP signaling in vivo, we have inactivated the type 4 cAMP-specific PDE (PDE4D) gene, a mammalian homologue of the Drosophila dunce. This isoenzyme is involved in feedback regulation of cAMP levels. Mice deficient in PDE4D exhibit delayed growth as well as reduced viability and female fertility. The decrease in fertility of the null female is caused by impaired ovulation and diminished sensitivity of the granulosa cells to gonadotropins. These pleiotropic phenotypes demonstrate that PDE4D plays a critical role in cAMP signaling and that the activity of this isoenzyme is required for the regulation of growth and fertility. PMID- 10518566 TI - Race-specific HIV-1 disease-modifying effects associated with CCR5 haplotypes. AB - Genetic variation in CC chemokine receptor 5 (CCR5), the major HIV-1 coreceptor, has been shown to influence HIV-1 transmission and disease progression. However, it is generally assumed that the same CCR5 genotype (or haplotype) has similar phenotypic effects in different populations. To test this assumption, we used an evolutionary-based classification of CCR5 haplotypes to determine their associated HIV-1 disease-modifying effects in a large well-characterized racially mixed cohort of HIV-1-seropositive individuals. We demonstrate that the spectrum of CCR5 haplotypes associated with disease acceleration or retardation differs between African Americans and Caucasians. Also, we show that there is a strong interactive effect between CCR5 haplotypes with different evolutionary histories. The striking population-specific phenotypic effects associated with CCR5 haplotypes emphasize the importance of understanding the evolutionary context in which disease susceptibility genes are expressed. PMID- 10518567 TI - Structurally similar oxidized phospholipids differentially regulate endothelial binding of monocytes and neutrophils. AB - We previously have demonstrated that oxidized 1-palmitoyl-2-arachidonoyl-sn glycero-3-phosphorylcholine (OxPAPC), a component of minimally modified low density lipoprotein (MM-LDL), activates endothelial cells to bind monocytes. 1 Palmitoyl-2- (5-oxovaleroyl)-sn-glycero-3-phosphorylcholine (POVPC) and 1- palmitoyl-2-glutaroyl-sn-glycero-3-phosphorylcholine (PGPC), which are present in OxPAPC, MM-LDL, and atherosclerotic lesions, were shown to have a major role in the activation of endothelial cells. We now demonstrate that these two highly similar molecules have dramatically different effects on leukocyte endothelial interactions. POVPC is a potent regulator of monocyte-specific endothelial interactions. Treatment of endothelial cells with POVPC increased monocyte binding by inducing the surface expression of the connecting segment 1 domain of fibronectin; no increase in neutrophil binding was observed. In addition, POVPC strongly inhibited lipopolysaccharide-mediated induction of neutrophil binding and expression of E-selectin protein and mRNA. This inhibition was mediated by a protein kinase A-dependent pathway, resulting in down-regulation of NF-kappaB dependent transcription. In contrast, PGPC induced both monocyte and neutrophil binding and expression of E-selectin and vascular cell adhesion molecule 1. We present evidence to suggest that the two phospholipids act by different novel receptors present in Xenopus laevis oocytes and that POVPC, but not PGPC, stimulates a cAMP-mediated pathway. At concentrations equal to that present in MM LDL, the effect of POVPC dominates and inhibits PGPC-induced neutrophil binding and E-selectin expression in endothelial cells. In summary, our data provide evidence that both POVPC and PGPC are important regulators of leukocyte endothelial interactions and that POVPC may play a dominant role in a number of chronic inflammatory processes where oxidized phospholipids are known to be present. PMID- 10518568 TI - Suppression or induction of apoptosis by opposing pathways downstream from calcium-activated calcineurin. AB - Ca(2+)-mobilizing compounds such as the Ca(2+) ionophore A23187 or the endoplasmic reticulum Ca(2+) ATPase inhibitor thapsigargin can suppress or induce apoptosis in the same cells. The use of different calcineurin inhibitors has shown that both suppression and induction of apoptosis by the Ca(2+)-mobilizing compounds were mediated by calcineurin activation. Ca(2+)-mobilizing compounds activated p38 and p44/42 mitogen-activated protein kinases (MAPKs). Induction of apoptosis by the Ca(2+)-mobilizing compounds was suppressed by an inhibitor of p38 MAPK but not by an inhibitor of p44/42 MAPK. These MAPK inhibitors did not suppress apoptosis induction by wild-type p53 or by withdrawal of IL-6 from IL-6 dependent cells that are mediated by calcineurin-independent pathways. These MAPK inhibitors also did not affect the ability of Ca(2+)-mobilizing compounds to suppress apoptosis. The results indicate that (i) Ca(2+)- mobilizing compounds activate different and opposing pathways that diverge downstream from calcineurin activation that can either suppress or induce apoptosis in the same cells; (ii) p38 MAPK but not p44/42 MAPK is involved in induction of apoptosis but not in its suppression by the Ca(2+)-mobilizing compounds; and (iii) neither p38 nor p44/42 MAPKs mediate induction of apoptosis by some calcineurin-independent pathways. PMID- 10518569 TI - Progesterone receptors in the thymus are required for thymic involution during pregnancy and for normal fertility. AB - Thymocyte development is reported to be inhibited by pregnancy, although the impact of this effect on fertility is unknown. We demonstrate, using progesterone receptor null mutant mice, that the inhibitory effects of pregnancy hormones on T cell development require the presence of functional progesterone receptor (PR). A combination of hysterectomy, thymic immunohistochemistry, and transplant studies reveals that local expression of PR in thymic stromal cells is specifically required for thymic involution to occur. These cells, under the influence of progesterone, block T cell development at the early pre-T cell (CD3(-)CD44(+) CD25(+)) stage of development via a paracrine mechanism. In addition, age-related thymic involution is shown to occur by a separate PR-independent mechanism. Finally, pregnancy studies with thymic transplants from progesterone receptor null mutant mice to wild-type female recipients demonstrate that thymic stromal PR is required for normal fertility. Together, these observations provide evidence for a PR-dependent paracrine mechanism that blocks very early T cell lymphopoiesis during pregnancy and is essential for normal fertility. PMID- 10518570 TI - Underdeveloped uterus and reduced estrogen responsiveness in mice with disruption of the estrogen-responsive finger protein gene, which is a direct target of estrogen receptor alpha. AB - The biological roles of estrogen-responsive finger protein (efp) in vivo were evaluated in mice carrying a loss-of-function mutation in efp by gene-targeted mutagenesis. Although efp homozygous mice were viable and fertile in both sexes, the uterus that expressed abundant estrogen receptor alpha exhibited significant underdevelopment. When the ovariectomized homozygotes were subjected to 17beta estradiol treatment, they showed remarkably attenuated responses to estrogen, as exemplified by decreased interstitial water imbibition and retarded endometrial cell increase, at least, attributable to the lower ratio of G1 to S-phase progression in epithelial cells. These results suggest that efp is essential for the normal estrogen-induced cell proliferation and uterine swelling as one of the direct targets of estrogen receptor alpha. PMID- 10518571 TI - Identification of naturally processed and HLA-presented Epstein-Barr virus peptides recognized by CD4(+) or CD8(+) T lymphocytes from human blood. AB - The broad clinical implementation of cancer vaccines targeting the induction of specific T cell-mediated immunity is hampered because T cell defined tumor associated peptides are currently available for only a restricted range of tumor types. Current epitope identification strategies require a priori the generation of T "indicator" cell lines that specifically recognize the tumor antigenic epitope in in vitro assay systems. An alternative to this strategy is the use of "memory" T cells freshly isolated from the peripheral blood of patients with cancer in concert with sensitive effector cell readout assays (such as the cytokine enzyme-linked immunospot assay) and MS to identify relevant peptide epitopes. In a model system, we have evaluated the capacity of natural Epstein Barr virus (EBV)-transformed B-lymphoblastoid cell line-extracted peptides to activate "memory" viral-specific CD4(+) or CD8(+) T cells freshly isolated from the blood of an EBV-seropositive individual using the IFN-gamma enzyme-linked immunospot assay. After HPLC fractionation and loading onto autologous dendritic cells, multiple naturally processed HLA class I and II-associated lymphoblastoid cell line-derived peptides were isolated that were capable of inducing IFN-gamma spot production by "memory" T lymphocytes. Using MS analysis on a HPLC fraction recognized by CD8(+) T cells, we were able to sequence natural 9-, 10-, and 11 mer peptides naturally processed from the latent EBV antigen LMP-2 (latent membrane protein-2) and presented in the context of HLA-A2. This approach provides a useful methodology for the future identification of MHC-presented viral and tumor epitopes using freshly isolated patient materials. PMID- 10518572 TI - Lentiviral delivery of HIV-1 Vpr protein induces apoptosis in transformed cells. AB - Most current anticancer therapies act by inducing tumor cell stasis followed by apoptosis. HIV-1 Vpr effectively induces apoptosis of T cells after arrest of cells at a G(2)/M checkpoint. Here, we investigated whether this property of Vpr could be exploited for use as a potential anticancer agent. As a potentially safer alternative to transfer of genes encoding Vpr, we developed a method to efficiently introduce Vpr protein directly into cells. Vpr packaged into HIV-1 virions lacking a genome induced efficient cell cycle arrest and apoptosis. Introduction of Vpr into tumor cell lines of various tissue origin, including those bearing predisposing mutations in p53, XPA, and hMLH1, induced cell cycle arrest and apoptosis with high efficiency. Significantly, apoptosis mediated by virion-associated Vpr was more effective on rapidly dividing cells compared with slow-growing cells, thus, in concept, providing a potential differential effect between some types of tumor cells and surrounding normal cells. This model system provides a rationale and proof of concept for the development of potential cancer therapeutic agents based on the growth-arresting and apoptotic properties of Vpr. PMID- 10518573 TI - Visualizing the kinetics of tumor-cell clearance in living animals. AB - Evaluation of potential antineoplastic therapies would be enhanced by noninvasive detection of tumor cells in living animals. Because light is transmitted through mammalian tissues, it was possible to use bioluminescence to monitor (both externally and quantitatively) growth and regression of labeled human cervical carcinoma (HeLa) cells engrafted into immunodeficient mice. The efficacy of both chemotherapy and immunotherapeutic treatment with ex vivo expanded human T cell derived effector cells was evaluated. In the absence of therapy, animals showed progressive increases in signal intensity over time. Animals treated with cisplatin had marked reductions in tumor signal; 5'-fluorouracil was less effective, and cyclophosphamide was ineffective. Immunotherapy dramatically reduced signals at high effector-to-target cell ratios, and significant decreases were observed with lower ratios. This model system allowed sensitive, quantitative, real-time spatiotemporal analyses of the dynamics of neoplastic cell growth and facilitated rapid optimization of effective treatment regimens. PMID- 10518574 TI - Decreased selective uptake of high density lipoprotein cholesteryl esters in apolipoprotein E knock-out mice. AB - Scavenger receptor BI (SR-BI) mediates the selective uptake of high density lipoprotein (HDL) cholesteryl esters (CE) by cells, i.e., the uptake of CE without degradation of HDL protein. Mice with attenuated expression of SR-BI, because of targeted gene mutation (SR-BIatt mice), have increased plasma HDL levels as a result of decreased selective uptake in the liver. To further evaluate the role of SR-BI in lipoprotein metabolism, compound apolipoprotein E knock-out (apoE0)/SR-BIatt mice were bred. Hepatic SR-BI protein was increased (2.3-fold) in apoE0 mice compared with wild type (wt) and was reduced significantly in apoE0/SR-BIatt mice. However, the plasma lipoprotein profile of apoE0 and apoE0/SR-BIatt mice was identical. This was explained by HDL turnover studies that revealed that the selective clearance of HDL CE by the liver and adrenal was already profoundly impaired in apoE0 mice compared with wt (28% of wt in liver). A similar decrease in selective uptake was seen when apoE0 HDL was incubated with isolated apoE0 hepatocytes. The results suggest that apoE plays a major role in the selective clearance of HDL CE by the liver and adrenal gland, possibly by facilitating the presentation of HDL to SR-BI at the cell surface. PMID- 10518575 TI - Arterivirus discontinuous mRNA transcription is guided by base pairing between sense and antisense transcription-regulating sequences. AB - To generate an extensive set of subgenomic (sg) mRNAs, nidoviruses (arteriviruses and coronaviruses) use a mechanism of discontinuous transcription. During this process, mRNAs are generated that represent the genomic 5' sequence, the so called leader RNA, fused at specific positions to different 3' regions of the genome. The fusion of the leader to the mRNA bodies occurs at a short, conserved sequence element, the transcription-regulating sequence (TRS), which precedes every transcription unit in the genome and is also present at the 3' end of the leader sequence. Here, we have used site-directed mutagenesis of the infectious cDNA clone of the arterivirus equine arteritis virus to show that sg mRNA synthesis requires a base-pairing interaction between the leader TRS and the complement of a body TRS in the viral negative strand. Mutagenesis of the body TRS of equine arteritis virus RNA7 reduced sg RNA7 transcription severely or abolished it completely. Mutations in the leader TRS dramatically influenced the synthesis of all sg mRNAs. The construction of double mutants in which a mutant leader TRS was combined with the corresponding mutant RNA7 body TRS resulted in the specific restoration of mRNA7 synthesis. The analysis of the mRNA leader-body junctions of a number of mutants with partial transcriptional activity provided support for a mechanism of discontinuous minus-strand transcription that resembles similarity-assisted, copy-choice RNA recombination. PMID- 10518576 TI - Accumulation of specific RNAs encoding transcriptional factors and stress response proteins against a background of severe depletion of cellular RNAs in cells infected with herpes simplex virus 1. AB - Herpes simplex virus 1 encodes several functions to preclude the shutoff of host response to infection, including degradation of mRNA immediately after infection. To determine whether any cellular mRNAs accumulate in infected cells against a background of severe loss of host RNA, we hybridized cDNAs derived from three different cell lines infected with wild type and a mutant virus to a DNA array containing probes for 588 human genes representing different functional groups. The results were that (i) infected cells accumulated at levels above those of mock-infected cells, a small number of transcripts representing transcriptional factors that could regulate gene expression both positively and negatively, and one stress response protein (GADD45), (ii) the amount and nature of the accumulated transcripts showed limited variability depending on the cell and virus, and (iii) at least some of the proteins encoded by the accumulated transcripts could benefit either the virus or the host. PMID- 10518577 TI - Release of calcium from stores alters the morphology of dendritic spines in cultured hippocampal neurons. AB - The ability to monitor ongoing changes in the shape of dendritic spines has important implications for the understanding of the functional correlates of the great variety of shapes and sizes of dendritic spines in central neurons. We have monitored and three-dimensionally reconstructed dendritic spines in cultured hippocampal neurons over several hours of observation in a confocal laser scanning microscope. In the absence of extrinsic stimulation, the dimensions of dendritic spines of 3-week-old cultured neurons did not change to any significant degree over 3-4 hr in the culture dish, unlike the case with younger cultures. Releasing calcium from stores with pulse application of caffeine causes a transient rise of [Ca(2+)](i) in dendrites and spines, monitored with the calcium dye Oregon-green. Application of caffeine to a dendrite imaged with calcein caused a fast and significant increase in the size of existing dendritic spines and could lead to formation of new ones. This effect is mediated by calcium released from the ryanodine-sensitive stores, as application of caffeine in the presence of ryanodine blocked this effect on the morphology of dendritic spines. Thus, release of calcium from stores is sufficient to produce significant changes in the shape of dendritic spines of cultured hippocampal neurons. PMID- 10518578 TI - Dynamics of spatial summation in primary visual cortex of alert monkeys. AB - One of the fundamental tasks of the visual cortex is to integrate input from different parts of the retina, parsing an image into contours and surfaces, and then assembling these features into coherent representations of objects. To examine the role of the primary visual cortex in the integration of visual information, we measured the response properties of neurons under different stimulus conditions. Surprisingly, we found that even the most conventional measures of receptive field (RF) size were not fixed, but could vary depending on stimulus contrast and foreground-background relationships. On average, the length of the excitatory RF was 4-fold greater for a low-contrast stimulus than for a stimulus at high contrast. Embedding a high-contrast stimulus in a textured background tended to suppress neuronal responses and produced an enlargement in RF size similar to that observed by decreasing the contrast of an isolated stimulus. The results show that RF dimensions are regulated in a dynamic manner that depends both on local stimulus characteristics, such as contrast, and on global relationships between a stimulus and its surroundings. PMID- 10518579 TI - Selective expression of the large neutral amino acid transporter at the blood brain barrier. AB - Amino acid supply in brain is regulated by the activity of the large neutral amino acid transporter (LAT) at the brain capillary endothelial cell, which forms the blood-brain barrier (BBB) in vivo. Bovine BBB poly(A)(+) RNA was isolated from 2.0 kg of fresh bovine brain and size fractionated on a sucrose density gradient, and a size-fractionated bovine BBB cDNA library in the pSPORT vector was prepared. The full-length cDNA encoding the bovine BBB LAT was isolated from this library, and the predicted amino acid sequence was 89-92% identical to the LAT1 isoform. The bovine BBB LAT1 mRNA produced a 10-fold enhancement in tryptophan transport into frog oocytes coinjected with bovine BBB LAT1 mRNA and the mRNA for 4F2hc, which encodes the heavy chain of the heterodimer. Tryptophan transport into the mRNA-injected oocytes was sodium independent and was specifically inhibited by other large neutral amino acids, and the K(m) of tryptophan transport was 31.5 +/- 5.5 microM. Northern blotting with the bovine BBB LAT1 cDNA showed that the LAT1 mRNA is 100-fold higher in isolated bovine brain capillaries compared with C6 rat glioma cells or rat brain, and the LAT1 mRNA was not detected in rat liver, heart, lung, or kidney. These studies show that the LAT1 transcript is selectively expressed at the BBB compared with other tissues, and the abundance of the LAT1 mRNA at the BBB is manyfold higher than that of transcripts such as the 4F2hc antigen, actin, or the Glut1 glucose transporter. PMID- 10518581 TI - Tonic inhibition alternates in paired neurons that set direction of fish escape reaction. AB - Crossed antagonism between activities in neurons subserving alternating movements such as swimming or walking has been described in a number of systems. The role of reciprocal inhibition has been implicated in these activities, but involvement of rhythmic ongoing fluctuations of membrane potential, called synaptic "noise," has not been examined. In the Mauthner (M) cells, which control the direction of escape, this activity is inhibitory. We report that in the zebrafish (Danio rerio), inhibitory synaptic noise exhibits prolonged bursts of rhythmic, inhibitory postsynaptic potentials, which attenuate the M cell's sensibility to excitatory sensory drives. Furthermore, paired intracellular recordings have shown that inhibitory synaptic noise alternates between two distinct states, noisy and quiet, which are out of phase in the two cells. Firing of either M cell resets this pattern by reducing the inhibition in the contralateral one. This suggests that an avoidance reflex in one direction may favor initiation, by the opposite M cell, of a subsequent escape toward a more appropriate location. PMID- 10518580 TI - Evidence relating human verbal memory to hippocampal N-methyl-D-aspartate receptors. AB - Studies in rodents and nonhuman primates have linked the activity of N-methyl-D aspartate (NMDA) receptors within the hippocampus to animals' performance on memory-related tasks. However, whether these receptors are similarly essential for human memory is still an open question. Here we present evidence suggesting that hippocampal NMDA receptors, most likely within the CA1 region, do participate in human verbal memory processes. Words elicit a negative event related potential (ERP) peaking around 400 ms within the anterior mesial temporal lobe (AMTL-N400). Ketamine, an NMDA-receptor antagonist, reduces the amplitude of the AMTL-N400 (in contrast to other hippocampal potentials) on initial presentation, eliminates the typical AMTL-N400 amplitude reduction with repetition, and leads to significant memory impairment. Of the various hippocampal subfields, only the density of CA1 neurons correlates with the word related ERPs that are reduced by ketamine. Altogether, our behavioral, anatomical, and electrophysiological results indicate that hippocampal NMDA receptors are involved in human memory. PMID- 10518582 TI - Resistance to excitotoxin-induced seizures and neuronal death in mice lacking the preprotachykinin A gene. AB - Epileptic seizures are associated with increases in hippocampal excitability, but the mechanisms that render the hippocampus hyperexcitable chronically (in epilepsy) or acutely (in status epilepticus) are poorly understood. Recent evidence suggests that substance P (SP), a peptide that has been implicated in cardiovascular function, inflammatory responses, and nociception, also contributes to hippocampal excitability and status epilepticus, in part by enhancing glutamate release. Here we report that mice with disruption of the preprotachykinin A gene, which encodes SP and neurokinin A, are resistant to kainate excitoxicity. The mice show a reduction in the duration and severity of seizures induced by kainate or pentylenetetrazole, and both necrosis and apoptosis of hippocampal neurons are prevented. Although kainate induced the expression of bax and caspase 3 in the hippocampus of wild-type mice, these critical intracellular mediators of cell death pathways were not altered by kainate injection in the mutant mice. These results indicate that the reduction of seizure activity and the neuroprotection observed in preprotachykinin A null mice are caused by the extinction of a SP/neurokinin A-mediated signaling pathway that is activated by seizures. They suggest that these neurokinins are critical to the control of hippocampal excitability, hippocampal seizures, and hippocampal vulnerability. PMID- 10518583 TI - An infection-based model of neurodevelopmental damage. AB - Perinatal exposure to infectious agents and toxins is linked to the pathogenesis of neuropsychiatric disorders, but the mechanisms by which environmental triggers interact with developing immune and neural elements to create neurodevelopmental disturbances are poorly understood. We describe a model for investigating disorders of central nervous system development based on neonatal rat infection with Borna disease virus, a neurotropic noncytolytic RNA virus. Infection results in abnormal righting reflexes, hyperactivity, inhibition of open-field exploration, and stereotypic behaviors. Architecture is markedly disrupted in hippocampus and cerebellum, with reduction in granule and Purkinje cell numbers. Neurons are lost predominantly by apoptosis, as supported by increased mRNA levels for pro-apoptotic products (Fas, caspase-1), decreased mRNA levels for the anti-apoptotic bcl-x, and in situ labeling of fragmented DNA. Although inflammatory infiltrates are observed transiently in frontal cortex, glial activation (microgliosis > astrocytosis) is prominent throughout the brain and persists for several weeks in concert with increased levels of proinflammatory cytokine mRNAs (interleukins 1alpha, 1beta, and 6 and tumor necrosis factor alpha) and progressive hippocampal and cerebellar damage. The resemblance of these functional and neuropathologic abnormalities to human neurodevelopmental disorders suggests the utility of this model for defining cellular, biochemical, histologic, and functional outcomes of interactions of environmental influences with the developing central nervous system. PMID- 10518584 TI - Cognitive changes and modified processing of amyloid precursor protein in the cortical and hippocampal system after cholinergic synapse loss and muscarinic receptor activation. AB - A number of in vitro studies have shown that activation of muscarinic receptors by cholinergic agonists stimulates the nonamyloidogenic, alpha-secretase processing pathway of amyloid precursor protein (APP). To determine whether increased cholinergic neurotransmission can modify the APP processing in vivo, we administered a muscarinic receptor agonist (RS86) to normal or aged rats and rats with severe basal forebrain cholinergic deficits (induced by 192 IgG-saporin). The levels of the cell-associated APP in neocortex, hippocampus, and striatum, as well as the secreted form of APP (APPs) in cerebrospinal fluid, were examined by Western blots. Additionally, we investigated the association between the altered APP levels and behavioral deficits caused by cholinergic lesions. We found that treatment with muscarinic receptor agonist resulted in decreased APP levels in neocortex and hippocampus and increased levels of APPs in cerebrospinal fluid. Regulation of APP processing by the muscarinic agonist treatment occurred not only in normal rats, but also in aged and cholinergic denervated rats that model this aspect of Alzheimer's disease. Interestingly, we found that elevation of APP in neocortex correlated with the cognitive deficits in water-maze testing of rats with cholinergic dysfunction. These data indicate that increased cholinergic neurotransmission can enhance nonamyloidogenic APP processing in intact and lesioned rats and that APP may be involved in cognitive performance. PMID- 10518585 TI - Differential regulation of mammalian period genes and circadian rhythmicity by cryptochromes 1 and 2. AB - Cryptochromes regulate the circadian clock in animals and plants. Humans and mice have two cryptochrome (Cry) genes. A previous study showed that mice lacking the Cry2 gene had reduced sensitivity to acute light induction of the circadian gene mPer1 in the suprachiasmatic nucleus (SCN) and had an intrinsic period 1 hr longer than normal. In this study, Cry1(-/-) and Cry1(-/-)Cry2(-/-) mice were generated and their circadian clocks were analyzed at behavioral and molecular levels. Behaviorally, the Cry1(-/-) mice had a circadian period 1 hr shorter than wild type and the Cry1(-/-)Cry2(-/-) mice were arrhythmic in constant darkness (DD). Biochemically, acute light induction of mPer1 mRNA in the SCN was blunted in Cry1(-/-) and abolished in Cry1(-/-)Cry2(-/-) mice. In contrast, the acute light induction of mPer2 in the SCN was intact in Cry1(-/-) and Cry1(-/-)Cry2(-/ ) animals. Importantly, in double mutants, mPer1 expression was constitutively elevated and no rhythmicity was detected in either 12-hr light/12-hr dark or DD, whereas mPer2 expression appeared rhythmic in 12-hr light/12-hr dark, but nonrhythmic in DD with intermediate levels. These results demonstrate that Cry1 and Cry2 are required for the normal expression of circadian behavioral rhythms, as well as circadian rhythms of mPer1 and mPer2 in the SCN. The differential regulation of mPer1 and mPer2 by light in Cry double mutants reveals a surprising complexity in the role of cryptochromes in mammals. PMID- 10518586 TI - Long-term doxycycline-controlled expression of human tyrosine hydroxylase after direct adenovirus-mediated gene transfer to a rat model of Parkinson's disease. AB - Developments of technologies for delivery of foreign genes to the central nervous system are opening the field to promising treatments for human neurodegenerative diseases. Gene delivery vectors need to fulfill several criteria of efficacy and safety before being applied to humans. The ability to drive expression of a therapeutic gene in an adequate number of cells, to maintain long-term expression, and to allow exogenous control over the transgene product are essential requirements for clinical application. We describe the use of an adenovirus vector encoding human tyrosine hydroxylase (TH) 1 under the negative control of the tetracycline-sensitive gene regulatory system for direct injection into the dopamine-depleted striatum of a rat model of Parkinson's disease. This vector mediated synthesis of TH in numerous striatal cells and transgene expression was observed in a large proportion of them for at least 17 weeks. Furthermore, doxycyline, a tetracycline analog, allowed efficient and reversible control of transgene expression. Thus, the insertion of a tetracycline-sensitive regulatory cassette into a single adenovirus vector provides a promising system for the development of successful and safe therapies for human neurological diseases. Our results also confirm that future effective gene replacement approaches to Parkinson's disease will have to consider the concomitant transfer of TH and GTP-cyclohydrolase transgenes because the synthesis of the TH cofactor tetrahydrobiopterin may be crucial for restoration of the dopaminergic deficit. PMID- 10518587 TI - Diversity and distribution of nicotinic acetylcholine receptors in the locus ceruleus neurons. AB - The neurons of the locus ceruleus are responsible for most of the noradrenergic innervation in the brain and nicotine potentiates noradrenaline release from their terminals. Here we investigated the diversity and subcellular distribution of nicotinic acetylcholine receptors (nAChRs) in the locus ceruleus both somatically, by combining single-cell reverse transcription-PCR with electrophysiological characterization, and at the level of nerve terminals, by conducting noradrenaline efflux experiments. The proportion of neurons in the locus ceruleus expressing the nicotinic subunit mRNAs varied from 100% (beta2) to 3% (alpha2). Yet, two populations of neurons could be distinguished on the basis of the pattern of expression of nAChR mRNAs and electrophysiological properties. One population (type A) of small cells systematically expressed alpha3 and beta4 mRNAs (and often alpha6, beta3, alpha5, alpha4), and nicotinic agonists elicited large currents with a potency order of cytisine > nicotine. Another population (type B) of cells with large soma did not contain alpha3 and beta4 mRNAs but, systematically, alpha6 and beta3 (and often alpha4) and responded to nicotinic agonists in the order of nicotine > cytisine. The nicotinic modulation of noradrenaline release in the hippocampus displayed an order of potency nicotine > cytisine, suggesting that noradrenergic terminals in the hippocampus originate largely from type B cells of the locus ceruleus. Accordingly, immunocytochemical labeling showed that beta3 is present in hippocampal terminals. The alpha6beta3beta2(alpha4) heterooligomer thus behaves as the main nicotinic regulator of the ceruleo-hippocampal pathway. PMID- 10518588 TI - Impaired cerebellar synapse maturation in waggler, a mutant mouse with a disrupted neuronal calcium channel gamma subunit. AB - The waggler, a neurological mutant mouse with a disrupted putative neuronal Ca(2+) channel gamma subunit, exhibits a cerebellar granule cell-specific brain derived neurotrophic factor deficit, severe ataxia, and impaired eyeblink conditioning. Here, we show that multiple synapses of waggler cerebellar granule cells are arrested at an immature stage during development. Synaptic transmission is reduced at parallel fiber-Purkinje cell synapses. The Golgi cell-granule cell synaptic currents show immature kinetics associated with reduced gamma aminobutyric acid type A receptor alpha6 subunit expression in granule cells. In addition, the mossy fiber-granule cell synapses exhibit N-methyl-D-aspartate (NMDA) receptor-mediated excitatory postsynaptic currents (EPSCs), but not alpha amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor-mediated EPSCs. Our results suggest that voltage-dependent Ca(2+) channels are involved in synapse maturation. This deficient synaptic transmission in the waggler cerebellum may account for their behavioral deficits. PMID- 10518589 TI - Feeding behavior in dopamine-deficient mice. AB - Mice that cannot make dopamine (DA), a condition caused by the selective inactivation of tyrosine hydroxylase in dopaminergic neurons, are born normal but gradually become hypoactive and hypophagic, and die at 3 weeks of age. We characterized the feeding and locomotor responses of these DA-deficient (DA-/-) mice to 3, 4-dihyroxy-L-phenylalanine (L-DOPA) to investigate the relationship between brain DA levels and these complex behaviors. Daily administration of L DOPA to DA-/- mice stimulated locomotor activity that lasted 6 to 9 hr; during that time the mice consumed most of their daily food and water. The minimal dose of L-DOPA that was sufficient to elicit normal feeding behavior in the DA-/- mice also restored their striatal DA to 9.1% of that in the wild-type (WT) mice at 3 hr; then DA content declined to <1% of WT levels by 24 hr. This dose of L-DOPA induced locomotor activity that exceeded that of treated WT mice by 5- to 7-fold, suggesting that DA-/- mice are supersensitive to DA. Unexpectedly, DA-/- mice manifested a second wave of activity 24 to 48 hr after L-DOPA treatment that was equivalent in magnitude to that of WT mice and independent of DA receptor activation. The DA-/- mice approached, sniffed, and chewed food during this second period of activity, but they ate <10% of that required for sustenance. Therefore, DA-/- mice can execute behaviors necessary to seek and ingest food, but they do not eat enough to survive. PMID- 10518590 TI - Endogenous activation of metabotropic glutamate receptors in neocortical development causes neuronal calcium oscillations. AB - Oscillations in intracellular free calcium concentration ([Ca(2+)](i)) occur spontaneously in immature neurons of the developing cerebral cortex. Here, we show that developing murine cortical neurons exhibit calcium oscillations in response to direct activation of the mGluR5 subtype of the group I metabotropic glutamate receptor (mGluR). In contrast, other manipulations that elicit [Ca(2+)](i) increases produce simple, nonoscillatory changes. Furthermore, we find that spontaneous oscillatory [Ca(2+)](i) activity is blocked by antagonists of group I mGluRs, suggesting a specific role for mGluR activation in the promotion of oscillatory [Ca(2+)](i) dynamics in immature cortical neurons. The oscillatory pattern of [Ca(2+)](i) increases produced by mGluR activation might play a role in the regulation of gene expression and the control of developmental events. PMID- 10518591 TI - RNA diversity has profound effects on the translation of neuronal nitric oxide synthase. AB - A comprehensive analysis of the structure of neuronal nitric oxide synthase (nNOS; EC 1.14.13.39) mRNA species revealed NOS1 to be the most structurally diverse human gene described to date in terms of promoter usage. Nine unique exon 1 variants are variously used for transcript initiation in diverse tissues, and each is expressed from a unique 5'-flanking region. The dependence on unique genomic regions to control transcription initiation in a cell-specific fashion burdens the transcripts with complex 5'-mRNA leader sequences. Elaborate splicing patterns that involve alternatively spliced leader exons and exon skipping have been superimposed on this diversity. Highly structured nNOS mRNA 5'-untranslated regions, which have profound effects on translation both in vitro and in cells, contain cis RNA elements that modulate translational efficiency in response to changes in cellular phenotype. PMID- 10518592 TI - Mimicry of erythropoietin by a nonpeptide molecule. AB - Erythropoietin (EPO) controls the proliferation and differentiation of erythroid progenitor cells into red blood cells. EPO induces these effects by dimerization of the EPO receptors (EPOR) present on these cells. To discover nonpeptide molecules capable of mimicking the effects of EPO, we identified a small molecule capable of binding to one chain of EPOR and used it to synthesize molecules capable of inducing dimerization of the EPOR. We first identified compound 1 (N-3 [2-(4-biphenyl)-6-chloro-5-methyl]indolyl-acetyl-L-lysine methyl ester) by screening the in-house chemical collection for inhibitors of EPO binding to human EPOR and then prepared compound 5, which contains eight copies of compound 1 held together by a central core. Although both compounds inhibited EPO binding of EPOR, only compound 5 induced dimerization of soluble EPOR. Binding of EPO to its receptor in cells results in activation of many intracellular signaling molecules, including transcription factors like signal transducer and activator of transcription (STAT) proteins, leading to growth and differentiation of these cells. Consistent with its ability to induce dimerization of EPOR in solution, compound 5 exhibited much of the same biological activities as EPO, such as (i) the activation of a STAT-dependent luciferase reporter gene in BAF3 cells expressing human EPOR, (ii) supporting the proliferation of several tumor cell lines expressing the human or mouse EPOR, and (iii) the in vitro differentiation of human progenitor cells into colonies of erythrocytic lineage. These data demonstrate that a nonpeptide molecule is capable of inducing EPOR dimerization and mimicking the biological activities of EPO. PMID- 10518593 TI - Pharmacological plasticity of cardiac ATP-sensitive potassium channels toward diazoxide revealed by ADP. AB - The pharmacological phenotype of ATP-sensitive potassium (K(ATP)) channels is defined by their tissue-specific regulatory subunit, the sulfonylurea receptor (SUR), which associates with the pore-forming channel core, Kir6.2. The potassium channel opener diazoxide has hyperglycemic and hypotensive properties that stem from its ability to open K(ATP) channels in pancreas and smooth muscle. Diazoxide is believed not to have any significant action on cardiac sarcolemmal K(ATP) channels. Yet, diazoxide can be cardioprotective in ischemia and has been found to bind to the presumed cardiac sarcolemmal K(ATP) channel-regulatory subunit, SUR2A. Here, in excised patches, diazoxide (300 microM) activated pancreatic SUR1/Kir6.2 currents and had little effect on native or recombinant cardiac SUR2A/Kir6.2 currents. However, in the presence of cytoplasmic ADP (100 microM), SUR2A/Kir6.2 channels became as sensitive to diazoxide as SUR1/Kir6. 2 channels. This effect involved specific interactions between MgADP and SUR, as it required Mg(2+), but not ATP, and was abolished by point mutations in the second nucleotide-binding domain of SUR, which impaired channel activation by MgADP. At the whole-cell level, in cardiomyocytes treated with oligomycin to block mitochondrial function, diazoxide could also activate K(ATP) currents only after cytosolic ADP had been raised by a creatine kinase inhibitor. Thus, ADP serves as a cofactor to define the responsiveness of cardiac K(ATP) channels toward diazoxide. The present demonstration of a pharmacological plasticity of K(ATP) channels identifies a mechanism for the control of channel activity in cardiac cells depending on the cellular ADP levels, which are elevated under ischemia. PMID- 10518595 TI - Diet-dependent hypercalciuria in transgenic mice with reduced CLC5 chloride channel expression. AB - Dent's disease is an X-linked inherited disorder characterized by hypercalciuria, nephrocalcinosis, nephrolithiasis, low molecular weight proteinuria, Fanconi's syndrome, and renal failure. It is caused by inactivating mutations in CLC5, a member of the CLC voltage-gated chloride channel family. CLC5 is known to be expressed in the endosomal compartment of the renal proximal tubule, where it may be required for endosomal acidification and trafficking. Although the Fanconi's syndrome and low molecular weight proteinuria in Dent's disease can be explained by disruption of endosomal function in this nephron segment, the pathogenesis of the hypercalciuria in this disease is unknown. We have generated transgenic mice (RZ) with reduced CLC5 expression by introduction of an antisense ribozyme targeted against CLC5. RZ mice are markedly hypercalciuric compared with nontransgenic control mice, at a time when their serum electrolytes and renal function are otherwise normal. This suggests that hypercalciuria in Dent's disease is a direct consequence of CLC5 hypofunction and is not attributable to a gain of function by mutant CLC5, an effect of modifier genes, or a secondary result of nonspecific renal injury. Surprisingly, hypercalciuria in RZ mice is abolished by dietary calcium deprivation, suggesting that the hypercalciuria may be attributable to gastrointestinal hyperabsorption of calcium rather than a renal calcium leak. PMID- 10518594 TI - XK469, a selective topoisomerase IIbeta poison. AB - XK469 (NSC 697887) is a synthetic quinoxaline phenoxypropionic acid derivative that possesses unusual solid tumor selectivity and activity against multidrug resistant cancer cells. We report here that XK469 and its S(-) and R(+)-isomers induce reversible protein-DNA crosslinks in mammalian cells. Under protein denaturing conditions, the protein-DNA crosslinks are rendered irreversible and stable to DNA banding by CsCl gradient ultracentrifugation. Several lines of evidence indicate that the primary target of XK469 is topoisomerase IIbeta. Preferential targeting of topoisomerase IIbeta may explain the solid tumor selectivity of XK469 and its analogs because solid tumors, unlike leukemias, often have large populations of cells in the G(1)/G(0) phases of the cell cycle in which topoisomerase IIbeta is high whereas topoisomerase IIalpha, the primary target of many leukemia selective drugs, is low. PMID- 10518596 TI - Continuous detection of extracellular ATP on living cells by using atomic force microscopy. AB - Atomic force microscopy is a powerful technique used to investigate the surface of living cells under physiological conditions. The resolution of the instrument is mainly limited by the softness of living cells and the interactions with the scanning tip (cantilever). Atomic force microscopy, in combination with myosin functionalized cantilevers, was used in the detection of ATP concentrations in solution and on living cells. Functionally active tips were used to scan the surface of cells in culture and to show that the CFTR+ cell line (S9) had a basal surface ATP concentration that could be detected with atomic force microscopy (n = 10). ATP-dependent signals were not detectable in cells scanned with noncoated or heat-inactivated enzyme-coated tips (n = 9). Enzymatically active tips may serve as a model for future development of atomic force microscopy biosensors that can simultaneously detect topographical and biologically important compounds at the surface microenvironment of living cells. PMID- 10518597 TI - Auxin-induced K+ channel expression represents an essential step in coleoptile growth and gravitropism. AB - Auxin-induced growth of coleoptiles depends on the presence of potassium and is suppressed by K+ channel blockers. To evaluate the role of K+ channels in auxin mediated growth, we isolated and functionally expressed ZMK1 and ZMK2 (Zea mays K+ channel 1 and 2), two potassium channels from maize coleoptiles. In growth experiments, the time course of auxin-induced expression of ZMK1 coincided with the kinetics of coleoptile elongation. Upon gravistimulation of maize seedlings, ZMK1 expression followed the gravitropic-induced auxin redistribution. K+ channel expression increased even before a bending of the coleoptile was observed. The transcript level of ZMK2, expressed in vascular tissue, was not affected by auxin. In patch-clamp studies on coleoptile protoplasts, auxin increased K+ channel density while leaving channel properties unaffected. Thus, we conclude that coleoptile growth depends on the transcriptional up-regulation of ZMK1, an inwardly rectifying K+ channel expressed in the nonvascular tissue of this organ. PMID- 10518598 TI - Abscisic acid signal transduction in guard cells is mediated by phospholipase D activity. AB - In guard cells, the plant hormone abscisic acid (ABA) inhibits stomatal opening and induces stomatal closure through the coordinated regulation of ion transport. Despite this central role of ABA in regulating stomatal function, the signal transduction events leading to altered ion fluxes remain incompletely understood. We report that the activity of the enzyme phospholipase D (PLD) transiently increased in guard cell protoplasts at 2.5 and 25 min after ABA application. Treatment of guard cell protoplasts with phosphatidic acid (PtdOH), one of the products of PLD activity, led to an inhibition of the activity of the inward K+ channel. PtdOH also induced stomatal closure and inhibited stomatal opening when added to epidermal peels. Application of 1-butanol (1-buOH), a selective inhibitor of PtdOH production by PLD, inhibited the increase in PtdOH production elicited by ABA. 1-BuOH treatment also partially prevented ABA-induced stomatal closure and ABA-induced inhibition of stomatal opening. This inhibitory effect of buOH was enhanced by simultaneous application of nicotinamide, an inhibitor of cADP ribose action. These results suggest that in the guard cell, ABA activates the enzyme PLD, which leads to the production of PtdOH. This PtdOH is then involved in triggering subsequent ABA responses of the cell via a pathway operating in parallel to cADP ribose-mediated events. PMID- 10518600 TI - Late maturation of visual spatial integration in humans. AB - Visual development is thought to be completed at an early age. We suggest that the maturation of the visual brain is not homogeneous: functions with greater need for early availability, such as visuomotor control, mature earlier, and the development of other visual functions may extend well into childhood. We found significant improvement in children between 5 and 14 years in visual spatial integration by using a contour-detection task. The data show that long-range spatial interactions-subserving the integration of orientational information across the visual field-span a shorter spatial range in children than in adults. Performance in the task improves in a cue-specific manner with practice, which indicates the participation of fairly low-level perceptual mechanisms. We interpret our findings in terms of a protracted development of ventral visual stream function in humans. PMID- 10518601 TI - DNA structure specificity of Rap endonuclease. AB - The Rap protein of phage lambda is an endonuclease that nicks branched DNA structures. It has been proposed that Rap can nick D-loops formed during phage recombination to generate splice products without the need for the formation of a 4-strand (Holliday) junction. The structure specificity of Rap was investigated using a variety of branched DNA molecules made by annealing partially complementary oligo-nucleotides. On Holliday junctions, Rap endonuclease shows a requirement for magnesium or manganese ions, with Mn(2+)supporting 5-fold more cleavage than Mg(2+). The location of endonuclease incisions was determined on 3' tailed D-loop, bubble, flayed duplex, 5'-flap and Y junction DNA substrates. In all cases, Rap preferentially cleaves at the branch point of these molecules. With a flayed duplex, incisions are made in the duplex adjacent to the single strand arms. Comparison of binding and cleavage specificities revealed that Rap is highly structure-specific and exhibits a clear preference for 4- and 3 stranded DNA over Y and flayed duplex DNA. Almost no binding or cleavage was detected with duplex, partial duplex and single-stranded DNA. Thus Rap endonuclease shows a bias for structures that resemble D-loop and Holliday junction recombination intermediates. PMID- 10518599 TI - Pain modulation by release of the endogenous cannabinoid anandamide. AB - Synthetic cannabinoids produce behavioral analgesia and suppress pain neurotransmission, raising the possibility that endogenous cannabinoids serve naturally to modulate pain. Here, the development of a sensitive method for measuring cannabinoids by atmospheric pressure-chemical ionization mass spectrometry permitted measurement of the release of the endogenous cannabinoid anandamide in the periaqueductal gray (PAG) by in vivo microdialysis in the rat. Electrical stimulation of the dorsal and lateral PAG produced CB1 cannabinoid receptor-mediated analgesia accompanied by a marked increase in the release of anandamide in the PAG, suggesting that endogenous anandamide mediates the behavioral analgesia. Furthermore, pain triggered by subcutaneous injections of the chemical irritant formalin substantially increased the release of anandamide in the PAG. These findings indicate that the endogenous cannabinoid anandamide plays an important role in a cannabinergic pain-suppression system existing within the dorsal and lateral PAG. The existence of a cannabinergic pain modulatory system may have relevance for the treatment of pain, particularly in instances where opiates are ineffective. PMID- 10518602 TI - Rev-mediated nuclear export of RNA is dominant over nuclear retention and is coupled to the Ran-GTPase cycle. AB - The human immunodeficiency virus type-1 Rev protein induces the nuclear export of intron-containing viral mRNAs that harbor its binding site, the Rev response element (RRE). A leucine-rich region of Rev, the activation domain, is essential for function and has been shown to be a nuclear export signal (NES). Although Rev exports viral RNAs that resemble cellular mRNAs, competition studies performed using microinjected Xenopus laevis oocytes have previously indicated that Rev utilizes a non-mRNA export pathway. Here, we show that Rev is able to induce the export of both spliceable and non-spliceable RRE-containing pre-mRNAs and that this activity is not dependent on the location of the RRE within the RNA. Importantly, even RNA molecules of different classes, such as U3 snoRNA and U6 snRNA, which are retained in the nucleus by non-pre-mRNA mechanisms, are exported to the cytoplasm in response to Rev. Consistent with the notion that Rev-mediated export of RRE-containing RNA is mechanistically distinct from the export of processed cellular mRNA, a chimeric Rev protein in which its NES is replaced by the NES of hnRNP A1 does not induce the export of a Rev-responsive mRNA. Finally, we demonstrate that Rev/RRE-activated RNA export is, like other nuclear export pathways, linked to the Ran-GTPase cycle. PMID- 10518603 TI - Improved quantitation of DNA curvature using ligation ladders. AB - It is often desirable to estimate accurately the local shape of DNA molecules. Such measurements are useful in understanding the intrinsic contribution of DNA sequence to curvature, as well as in assessing the effects of chemical modifications. We have been investigating the effects of asymmetric phosphate neutralization on DNA shape using the well-characterized ligation ladder approach developed by Crothers and co-workers [D.M. Crothers and J.Drak (1992) Meth. Enzymol.,212, 46-71]. This technique is remarkably sensitive to differences in DNA shape. We now report a general quantitative assay of DNA curvature that we have validated using a set of phased A(5)tract standards. This approach allows simultaneous estimation of helix axis deflection magnitude and direction when a test sequence is monitored in at least three phasings relative to a reference A(5 6)tract in short DNA duplexes. Analysis using this improved approach confirms our published data on DNA curvature due to electrostatic effects. PMID- 10518604 TI - NMR-derived solution structure of a 17mer hydroxymethyluracil-containing DNA. AB - Incorporation of 5-(hydroxymethyl)-2'-deoxyuridine into DNA in place of thymine by SPO1, a Bacillus subtilis bacteriophage, allows the viral DNA to bind selectively to transcription factor 1. We have synthesized a TF1-binding site: d(5'-ACCHACHCHHHGHAGGT-3')-d(5'-ACCHACAAAGAGHAGGT-3') and studied this molecule using NMR spectroscopy. The chemical shifts of exchangeable and non-exchangeable protons were sequentially assigned. Absence of corresponding NOEs in the imino imino region suggested that the end base pairs did not form Watson-Crick hydrogen bond. Restrained molecular dynamics calculation yielded a family of B-DNA structures whose r.m.s.d. was 0.66 A (all atoms) for the internal 15 bp. The helical twist was 38.5 degrees per step. The base pairs were situated directly on the helix axis (X-displacement = -0.2 A). All sugars exhibited C2'-endo puckering with P = 167.3 degrees and upsilon(max)= 38.2 degrees. The OH groups of all hmU bases resided on the 3' side of the base plane and may affect the base orientation relative to the sugar plane as the average chi value for all hmU was 4 degrees more positive than that of other nucleosides (258 degrees versus 254 degrees ). Positive roll angles (rho) and small flanking twists (omega) at hmU suggested that the two hmU-A base pair steps open toward the minor grooves. PMID- 10518605 TI - Chiral and steric effects in the efficient binding of alpha-anomeric deoxyoligonucleoside N-alkylphosphoramidates to ssDNA and RNA. AB - We report hybridization properties of new phosphate-modified alpha oligonucleoside analogs with non-ionic or cationic internucleotide linkages such as methoxy-ethylphosphoramidate (PNHME), phosphoromorpholi-date (PMOR) and dimethylaminopropylphosphor-amidate (PNHDMAP). First we evaluated the chirality effect of the phosphorus atom on the affinity of alpha- or beta-dodecanucleoside phosphodiesters containing one chirally enriched N -alkylphosphoramidate linkage located in the middle of the sequence d(TCTT-AA*CCCACA). As for P-substituted beta-oligonucleo-tides, a difference in binding behavior between the two diastereoisomers (difference in Delta T (m)) exists in the hybridization properties of alpha-analogs when DNA was the target but this effect was not detrimental to duplex stability. This effect was considerably reduced when RNA was the target. Secondly we studied the effect of steric hindrance around phosphorus on the affinity of fully modified beta- and alpha-oligonucleoside N alkylphosphoramidates for their DNA and RNA targets. This effect was very weak with alpha-analogs whereas it was more pronounced with beta-oligos. PNHME modified alpha-oligonucleosides formed more stable duplexes with DNA (Delta T (m)+9.6 degrees C) and RNA (Delta T (m)+1.4 degrees C) targets than the 'parent' phosphodiester. Finally, base pairing specificity of these alpha-oligonucleo-side N -alkylphosphoramidates for their targets was found to be as high as for natural oligonucleoside phosphodiesters. PMID- 10518606 TI - DNA cleavage by hydroxy-salicylidene-ethylendiamine-iron complexes. AB - Bis(hydroxy)salen.Fe complexes were designed as self-activated chemical nucleases. The presence of a hy-droxyl group on the two salicylidene moieties serve to form a hydroquinone system cooperating with the iron redox system to facilitate spontaneous formation of free radicals. We compared the DNA binding and cleaving properties of the ortho -, meta- and para -(bishydroxy) salen.Fe complexes with that of the corresponding chelate lacking the hydroxyl groups. DNA melting temperature studies indicated that the para complex exhibits the highest affinity for DNA. In addition, this para compound was considerably more potent at cleaving supercoiled plasmid DNA than the regio-isomeric ortho - and meta hydroxy-salen.Fe complexes, even in the absence of a reducing agent, such as dithiothreitol used to activate the metal complex. The DNA cleaving activity of the para isomer is both time and concentration dependent and the complexed iron atom is absolutely essential for the sequence uniform cleavage of DNA. From a mechanistic point of view, electron spin resonance measurements suggest that DNA contributes positively to the activation of the semi-quinone system and the production of ligand radical species responsible for subsequent strand scission in the absence of a reducing agent. The para -hydroxy-salen.Fe complex has been used for detecting sequence-specific drug-DNA interactions. Specific binding of Hoechst 33258 to AT sequences and chromomycin to GC sequences were shown. The para -bis(hydroxy)salen.Fe derivative complements the tool box of footprinting reagents which can be utilised to produce efficient cleavage of DNA. PMID- 10518608 TI - PI-PfuI and PI-PfuII, intein-coded homing endonucleases from Pyrococcus furiosus. II. Characterization Of the binding and cleavage abilities by site-directed mutagenesis. AB - PI- Pfu I and PI- Pfu II from Pyrococcus furiosus are homing endonucleases, as shown in the accompanying paper. These two endonucleases are produced by protein splicing from the precursor protein including ribonucleotide reductase (RNR). We show here that both enzymes specifically interact with their substrate DNA and distort the DNA strands by 73 degrees and 67 degrees, respectively. They have two copies of the amino acid sequence motif LAGLIDADG, which is present in the majority of homing endonucleases and provides some of the catalytic residues necessary for DNA cleavage activity. Site-specific mutagenesis studies showed that two acidic residues in the motifs, Asp149 and Glu250 in PI- Pfu I, and Asp156 and Asp249 in PI- Pfu II, were critical for catalysis. The third residues of the active site triads, as predicted from the structure of PI- Sce I, were Asn225 in PI- Pfu I and Lys224 in PI- Pfu II. Substitution of Asn225 in PI- Pfu I by Ala did not affect catalysis. The cleavage activity of PI- Pfu II was 50-fold decreased by the substitution of Ala for Lys224. The binding affinity of the mutant protein for the substrate DNA also decreased 6-fold. The Lys in PI- Pfu II may play a direct or indirect role in catalysis of the endonuclease activity. PMID- 10518607 TI - PI-PfuI and PI-PfuII, intein-coded homing endonucleases from Pyrococcus furiosus. I. Purification and identification of the homing-type endonuclease activities. AB - We screened for proteins with specific binding activity to Holliday junction DNA from the hyperthermophilic archaeon Pyrococcus furiosus and found a protein that has specific affinity for DNA with a branched structure, like a three-way or four way junction. The protein was identified as one of the two inteins encoded in the gene for ribonucleotide reductase (RNR) by gene cloning. These two inteins were spliced out from the precursor protein as polypeptides with molecular weights of 53.078 and 43.976 kDa, respectively. The amino acid sequences of these inteins have two copies of the LAGLIDADG motif, which is found in the site-specific DNA endonucleases. The purified proteins actually cleaved double-stranded DNA with the sequence of the intein(-)allele, and, therefore, they were designated PI- Pfu I and PI- Pfu II. They generate a 4 bp 3'-OH overhang with a 5'-phosphate, like other known homing endonucleases originating from inteins. The optimal conditions of the DNA cleavage reaction, including temperature, pH, and concentrations of KCl and MgCl(2), have been determined. The high affinity for junction DNA of PI- Pfu I was confirmed using the purified protein. PMID- 10518609 TI - Imidazole-imidazole pair as a minor groove recognition motif for T:G mismatched base pairs. AB - The T:G mismatched base pair is associated with many genetic mutations. Understanding its biological consequences may be aided by studying the structural perturbation of DNA caused by a T:G base pair and by specific probing of the mismatch using small molecular ligands. We have shown previously that AR-1-144, a tri-imidazole (Im-Im-Im) minor groove binder, recognizes the sequence CCGG. NMR structural analysis of the symmetric 2:1 complex of AR-1-144 and GAACCGGTTC revealed that each AR-1-144 binds to four base pairs with the guanine N2 amino group forming a bifurcated hydrogen bond to a side-by-side Im/Im pair. We predicted that the free G-N2 amino group in a T:G wobble base pair can form two individual hydrogen bonds to a side-by-side Im/Im pair. Thus an Im/Im pair may be a good recognition motif for a T:G base pair in DNA. Cooperative and tight binding of an AR-1-144 homodimer to GAACTGGTTC permits a detailed structural analysis by 2D NOE NMR refinement and the refined structure confirms our prediction. Surprisingly, AR-1-144 does not bind to GAATCGGTTC. We further show that both the Im-Im-Im/Im-Py-Im heterodimer and the Im-Im-Im/Im-Im-Im homodimer bind strongly to the CACGGGTC + GACTCGTG duplex. These results together suggest that an Im/Im pair can specifically recognize a single T:G mismatch. Our results may be useful in future design of molecules (e.g. linked dimers) that can recognize a single T:G mismatch with specificity. PMID- 10518610 TI - Murine Stat2 is uncharacteristically divergent. AB - Characterization of the ability of human interferons (IFNs) to rapidly induce genes led to the identification of the first two members of the STAT (signal transducers and activators of transcription) family, Stat1 and Stat2. To study the unique role of this transcription factor in IFN signaling under more physiological conditions, murine Stat2 was isolated and found to be surprisingly divergent. This divergence was most striking in the C-terminal transcriptional activation domain. Studies on murine Stat2 indicate that it functions in IFN signaling. This includes IFN-alpha-dependent activation, nuclear translocation, DNA binding and activation of reporter genes. However, the profound divergence at the C-terminus suggests that murine Stat2 may have evolved to mediate some unique functions as well. To explore this possibility, proteins that interact with the C termini of murine and human Stat2 were examined. These studies indicate that the murine and human C-termini interact with an overlapping, but distinct set of proteins. PMID- 10518612 TI - ConStruct: a tool for thermodynamic controlled prediction of conserved secondary structure. AB - A tool for prediction of conserved secondary structure of a set of homologous single-stranded RNAs is presented. For each RNA of the set the structure distribution is calculated and stored in a base pair probability matrix. Gaps, resulting from a multiple sequence alignment of the RNA set, are introduced into the individual probability matrices. These 'aligned' probability matrices are summed up to give a consensus probability matrix emphasizing the conserved structural elements of the RNA set. Because the multiple sequence alignment is independent of any structural constraints, such an alignment may result in introduction of gaps into the homologous probability matrices that disrupt a common consensus structure. By use of its graphical user interface the presented tool allows the removal of such misalignments, which are easily recognized, from the individual probability matrices by optimizing the sequence alignment with respect to a structural alignment. From the consensus probability matrix a consensus structure is extracted, which is viewable in three different graphical representations. The functionality of the tool is demonstrated using a small set of U7 RNAs, which are involved in 3'-end processing of histone mRNA precursors. Supplementary Material lists further results obtained. Advantages and drawbacks of the tool are discussed in comparison to several other algorithms. PMID- 10518611 TI - Isolation and characterization of the C-terminal nuclease domain from the RecB protein of Escherichia coli. AB - The RecB subunit of the Escherichia coli RecBCD enzyme has been shown in previous work to have two domains: an N-terminal 100 kDa domain with ATP-dependent helicase activity, and a C-terminal 30 kDa domain. The 30 kDa domain had nuclease activity when linked to a heterologous DNA binding protein, but by itself it appeared unable to bind DNA and lacked detectable nuclease activity. We have expressed and isolated this 30 kDa domain, called RecB(N), and show that it does have nuclease activity detectable at high protein concentration in the presence of polyethylene glycol, added as a molecular crowding agent. The activity is undetectable in a mutant RecB(N)protein in which an aspartate residue has been changed to alanine. Structural analysis of the wild-type and mutant RecB(N)proteins by second derivative absorbance and circular dichroism spectroscopy indicates that both are folded proteins with very similar secondary and tertiary structures. The results show that the Asp-->Ala mutation has not caused a significant structural change in the isolated domain and they support the conclusion that the C-terminal domain of RecB has the sole nuclease active site of RecBCD. PMID- 10518613 TI - Whole genome-based phylogenetic analysis of free-living microorganisms. AB - A phylogenetic 'tree of life' has been constructed based on the observed presence and absence of families of protein-encoding genes observed in 11 complete genomes of free-living microorganisms. Past attempts to reconstruct the evolutionary relation-ships of microorganisms have been limited to sets of genes rather than complete genomes. Despite apparent rampant lateral gene transfer among microorganisms, these results indicate a single robust underlying evolutionary history for these organisms. Broadly, the tree produced is very similar to the small subunit rRNA tree although several additional phylogenetic relationships appear to be resolved, including the relationship of Archaeoglobus to the methanogens studied. This result is in contrast to notions that a robust phylogenetic reconstruction of microorganisms is impossible due to their genomes being composed of an incomprehensible amalgam of genes with complicated histories and suggests that this style of genome-wide phylogenetic analysis could become an important method for studying the ancient diversification of life on Earth. Analyses using informational and operational subsets of the genes showed that this 'tree of life' is not dependent on the phylogenetically more consistent informational genes. PMID- 10518614 TI - HIF-1-mediated activation of transferrin receptor gene transcription by iron chelation. AB - Treatment with iron chelators mimics hypoxic induction of the hypoxia inducible factor (HIF-1) which activates transcription by binding to hypoxia responsive elements (HRE). We investigated whether HIF-1 is involved in transcriptional activation of the transferrin receptor (TfR), a membrane protein which mediates cellular iron uptake, in response to iron deprivation. The transcription rate of the TfR gene in isolated nuclei was up-regulated by treatment of Hep3B human hepatoma cells with the iron chelator desferrioxamine (DFO). The role of HIF-1 in the activation of TfR was indicated by the following observations: (i) DFO dependent activation of a luciferase reporter gene in transfected Hep3B cells was mediated by a fragment of the human TfR promoter containing a putative HRE sequence; (ii) mutation of this sequence prevented stimulation of luciferase activity; (iii) binding to this sequence of HIF-1alpha, identified by competition experiments and supershift assays, was induced by DFO. Furthermore, in mouse hepatoma cells unable to assemble functional HIF-1, inducibility of TfR transcription by DFO was lost and TfR mRNA up-regulation was reduced. These results, which show the role of HIF-1 in the control of TfR gene expression in conditions of iron depletion, give insights into the mechanisms of transcriptional regulation which concur with the well-characterized post transcriptional control of TfR expression to expand the extent of response to iron deficiency. PMID- 10518615 TI - Structural organization and regulation of the plasmid-borne type II restriction modification system Kpn2I from Klebsiella pneumoniae RFL2. AB - Kpn 2I enzymes of a type II restriction-modification (R-M) system from the bacterium Klebsiella pneumoniae strain RFL2 recognize the sequence 5'-TCCGGA-3'. The Kpn 2I R-M genes have been cloned and expressed in Escherichia coli. DNA sequence analysis revealed the presence of two convergently transcribed open reading frames (ORFs) coding for a restriction endonuclease (Enase) of 301 amino acids (34. 8 kDa) and methyltransferase (Mtase) of 375 amino acids (42.1 kDa). The 3'-terminal ends of these genes ( kpn2IR and kpn2IM, respectively) overlap by 11 bp. In addition, a small ORF (gene kpn2IC ) capable of coding for a protein of 96 amino acids in length (10.6 kDa) was found upstream of kpn2IM. The direction of kpn2IC transcription is opposite to that of kpn2IM. The predicted amino acid sequence of this ORF includes a probable helix-turn-helix motif. We show that the product of kpn2IC represses expression of the Kpn 2I Mtase but has no influence on expression of the Enase gene. Such a mode of regulation is unique among R-M systems analyzed so far. The Kpn 2I R-M is located on the K.pneumoniae RFL2 plasmid pKp4.3, which is able to replicate in E.coli cells. PMID- 10518616 TI - RPA subunit arrangement near the 3'-end of the primer is modulated by the length of the template strand and cooperative protein interactions. AB - To analyze the interaction of human replication protein A (RPA) and its subunits with the DNA template-primer junction in the DNA replication fork, we designed several template-primer systems differing in the size of the single-stranded template tail (4, 9, 13, 14, 19 and 31 nt). Base substituted photoreactive dNTP analogs-5-[ N -(2-nitro-5-azidobenzoyl)- trans -3-amino-propenyl-1]-2' deoxyuridine-5'-triphosphate (NAB-4-dUTP) and 5-[ N -[ N -(2-nitro-5 azidobenzoyl)glycyl]- trans -3-aminopropenyl-1]-2'-deoxyuridine-5'-triphosphate (NAB-7-dUTP)-were used as substrates for elongation of radiolabeled primer template by DNA polymerases in the presence or absence of RPA. Subsequent UV crosslinking showed that the pattern of p32 and p70 RPA subunit labeling, and consequently their interaction with the template-primer junction, is strongly dependent on the template extension length at a particular RPA concentration, as well as on the ratio of RPA to template concentration. Our results suggest a model of changes in the RPA configuration modulating by the length of the template extension in the course of nascent DNA synthesis. PMID- 10518617 TI - Characterization of a separate small domain derived from the 5' end of 23S rRNA of an alpha-proteobacterium. AB - We demonstrate the presence of a separate processed domain derived from the 5' end of 23S rRNA in ribosomes of Rhodopseudomonas palustris, a member of the alpha ++proteobacteria. Previous sequencing studies predicted intervening sequences (IVS) at homologous positions within the 23S rRNA genes of several alpha proteobacteria, including R.palustris, and we find a processed 23S rRNA 5' domain in unfractionated RNA from several species. 5.8S rRNA from eukaryotic cytoplasmic large subunit ribosomes and the bacterial processed 23S rRNA 5' domain share homology, possess similar structures and are both derived by processing of large precursors. However, the internal transcribed spacer regions or IVSs separating them from the main large subunit rRNAs are evolutionarily unrelated. Consistent with the difference in sequence, we find that the site and mechanism of IVS processing also differs. Rhodopseudomonas palustris IVS-containing RNA precursors are cleaved in vitro by Escherichia coli RNase III or a similar activity present in R.palustris extracts at a processing site distinct from that found in eukaryotic systems and this results in only partial processing of the IVS. Surprisingly, in a reaction unlike characterized cases of eubacterial IVS processing, an RNA segment larger than the corresponding DNA insertion is removed which contains conserved sequences. These sequences, by analogy, serve to link the 23S rRNA 5' rRNA domains or 5.8S rRNAs to the main portion of other prokaryotic 23S rRNAs or to eukaryotic 28S rRNAs, respectively. PMID- 10518618 TI - Exhaustive mining of EST libraries for genes differentially expressed in normal and tumour tissues. AB - A four-step procedure for the efficient and systematic mining of whole EST libraries for differentially expressed genes is presented. After eliminating redundant entries from the EST library under investigation (step 1), contigs of maximal length are built upon each remaining EST using about 4 000 000 public and proprietary ESTs (step 2). These putative genes are compared against a database comprising ESTs from 16 different tissues (both normal and tumour affected) to determine whether or not they are differentially expressed (step 3; electronic northern). Fisher's exact test is used to assess the significance of differential expression. In step 4, an attempt is made to characterise the contigs obtained in the assembly through database comparison. A case study of the CGAP library NCI_CGAP_Br1.1, a library made from three (well, moderately, and poorly differentiated) invasive ductal breast tumours (2126 ESTs in total) was carried out. Of the maximal contigs, 139 were found to be significantly (alpha = 0.05) over-expressed in breast tumour tissue, while 13 appeared to be down-regulated. PMID- 10518619 TI - NMR solution structure of a DNA dodecamer containing a transplatin interstrand GN7-CN3 cross-link. AB - The DNA duplex d(CTCTCG*AGTCTC).d(GAGAC-TC*GAGAG) containing a single trans- diammine-dichloroplatinum(II) interstrand cross-link (where G* and C* represent the platinated bases) has been studied by two-dimensional NMR. All the exchangeable and non-exchangeable proton resonance lines were assigned (except H5'/H5") and the NOE intensities were transformed into distances via the RELAZ program. By combining the NOESY and COSY data (330 constraints) and NMR constrained molecular mechanics using JUMNA, a solution structure of the cross linked duplex has been determined. The duplex is distorted over two base pairs on each side of the interstrand cross-link and exhibits a slight bending of its axis ( approximately 20 degrees ) towards the minor groove. The platinated guanine G* adopts a syn conformation. The rotation results in a Hoogsteen-type pairing between the complementary G(6)* and C(19)* residues which is mediated by the platinum moiety and is stabilized by a hydrogen bond between O6(G(6)*) and N4H(C(19)*). The rise between the cross-linked residues and the adjacent residues is increased owing to the interaction between these adjacent residues and the ammine groups of the platinum moiety. These results are discussed in relation to the slow rate of closure of the monofunctional adducts into interstrand cross links. PMID- 10518620 TI - Identification of an essential proximal sequence element in the promoter of the telomerase RNA gene of Tetrahymena thermophila. AB - Telomerase is a ribonucleoprotein reverse transcriptase that synthesizes and maintains telomeric DNA. Studies of telomeres and telomerase are facilitated by the large number of linear DNA molecules found in ciliated protozoa, such as Tetrahymena thermophila. To examine the expression of telomerase, we investigated the transcription of the RNA polymerase III-directed gene encoding the RNA subunit (TER1) of this enzyme. A chimeric gene containing the Glaucoma chattoni TER1 transcribed region flanked by 5' and 3' Tetrahymena regions was used to identify promoter elements following transformation of Tetrahymena cells. Disruption of a conserved proximal sequence element (PSE) located at -55 in the Tetrahymena TER1 5' flanking region eliminated expression of the chimeric gene. In addition, mutation of an A/T-rich element at -25 decreased expression markedly. A gel mobility shift assay and protein-DNA cross-linking identified a PSE-binding polypeptide of 50-60 kDa in Tetrahymena extracts. Gel filtration analysis revealed a native molecular mass of approximately 160 kDa for this binding activity. Our results point to a similar architecture between ciliate telomerase RNA and metazoan U6 small nuclear RNA promoters. PMID- 10518621 TI - Construction of a recombinant adenovirus for efficient delivery of the I-SceI yeast endonuclease to human cells and its application in the in vivo cleavage of chromosomes to expose new potential telomeres. AB - We have constructed a replication-defective adenovirus vector encoding the yeast I- Sce I endonuclease under the control of the murine cytomegalovirus immediate early gene promoter (AdM Sce I) for efficient delivery of this enzyme to mammalian cells. We present evidence of AdM Sce I-mediated I- Sce I protein expression and cleavage activity in replication-permissive 293 cells, and of cleavage of chromosomes in vivo in both 293 cells and in non-permissive human cells. We have exploited this system for the generation of chromosomes capped by artificial telomeric sequences in cells with integrated plasmids containing telomeric DNA arrays adjacent to an I- Sce I recognition site. The properties of the AdM Sce I virus described here make it a useful tool for studying biological processes involving induction of DNA breaks, recombination and gene targeting in cells grown in culture and in vivo. PMID- 10518622 TI - Gene targeted DNA double-strand break induction by (125)I-labeled triplex-forming oligonucleotides is highly mutagenic following repair in human cells. AB - A parallel binding motif 16mer triplex-forming oligonucleotide (TFO) complementary to a polypurine-polypyrimidine target region near the 3'-end of the SupF gene of plasmid pSP189 was labeled with [5-(125)I]dCMP at position 15. Following triplex formation and decay accumulation, radiation-induced site specific double-strand breaks (DSBs) were produced in the pSP189 SupF gene. Bulk damaged DNA and the isolated site-specific DSB-containing DNA were separately transfected into human WI38VA13 cells and allowed to repair prior to recovery and analysis of mutants. Bulk damaged DNA had a relatively low mutation frequency of 2.7 x 10(-3). In contrast, the isolated linear DNA containing site-specific DSBs had an unusually high mutation frequency of 7.9 x 10(-1). This was nearly 300 fold greater than that observed for the bulk damaged DNA mixture, and >1.5 x 10(4)-fold greater than background. The mutation spectra displayed a high proportion of deletion mutants targeted to the(125)I binding position within the SupF gene for both bulk damaged DNA and isolated linear DNA. Both spectra were characterized by complex mutations with mixtures of changes. However, mutations recovered from the linear site-specific DSB-containing DNA presented a much higher proportion of complex deletion mutations. PMID- 10518623 TI - Codon reading patterns in Drosophila melanogaster mitochondria based on their tRNA sequences: a unique wobble rule in animal mitochondria. AB - Mitochondrial (mt) tRNA(Trp), tRNA(Ile), tRNA(Met), tRNA(Ser)GCU, tRNA(Asn)and tRNA(Lys)were purified from Drosophila melanogaster (fruit fly) and their nucleotide sequences were determined. tRNA(Lys)corresponding to both AAA and AAG lysine codons was found to contain the anticodon CUU, C34 at the wobble position being unmodified. tRNA(Met)corresponding to both AUA and AUG methionine codons was found to contain 5-formylcytidine (f(5)C) at the wobble position, although the extent of modification is partial. These results suggest that both C and f(5)C as the wobble bases at the anticodon first position (position 34) can recognize A at the codon third position (position 3) in the fruit fly mt translation system. tRNA(Ser)GCU corresponding to AGU, AGC and AGA serine codons was found to contain unmodified G at the anticodon wobble position, suggesting the utilization of an unconventional G34-A3 base pair during translation. When these tRNA anticodon sequences are compared with those of other animal counterparts, it is concluded that either unmodified C or G at the wobble position can recognize A at the codon third position and that modification from A to t(6)A at position 37, 3'-adjacent to the anticodon, seems to be important for tRNA possessing C34 to recognize A3 in the mRNA in the fruit fly mt translation system. PMID- 10518624 TI - Design and isolation of ribozyme-substrate pairs using RNase P-based ribozymes containing altered substrate binding sites. AB - Substrate recognition and cleavage by the bacterial RNase P RNA requires two domains, a specificity domain, or S-domain, and a catalytic domain, or C-domain. The S-domain binds the T stem-loop region in a pre-tRNA substrate to confer specificity for tRNA substrates. In this work, the entire S-domain of the Bacillus subtilis RNase P RNA is replaced with an artificial substrate binding module. New RNA substrates are isolated by in vitro selection using two libraries containing random regions of 60 nt. At the end of the selection, the cleavage rates of the substrate library are approximately 0.7 min(-1)in 10 mM MgCl(2)at 37 degrees C, approximately 4-fold better than the cleavage of a pre-tRNA substrate by the wild-type RNase P RNA under the same conditions. The contribution of the S domain replacement to the catalytic efficiency is from 6- to 22 000-fold. Chemical and nuclease mapping of two ribozyme-product complexes shows that this contribution correlates with direct interactions between the S-domain replacement and the selected substrate. These results demonstrate the feasibility of design and isolation of RNase P-based, matching ribozyme-substrate pairs without prior knowledge of the sequence or structure of the interactive modules in the ribozyme or substrate. PMID- 10518625 TI - Detection of homozygous deletions in tumors by hybridization of representational difference analysis (RDA) products to chromosome-specific YAC clone arrays. AB - Representational difference analysis (RDA), a subtractive hybridization method that enriches differences between complex genomes, can be used to isolate fragments deleted in tumor genomes. Usually, most of the clones obtained by this approach result from polymorphic fragments. Therefore, identification of homozygously deleted fragments, which can indicate the presence of tumor suppressor loci, is often tedious. To overcome this limitation, we devised a novel strategy in which labeled RDA products are hybridized in toto against membranes spotted with YAC clones covering a region of interest. In such a way, identified YAC clones provide positional information on homozygous deletions and loss of heterozygosity (LOH) regions. We have tested this approach with a tumor known to have a homozygous deletion within a region of LOH on chromosome 13. RDA was performed using representations generated with restriction enzymes Bgl II, Nco I and Xba I, and the difference products of each experiment were separately hybridized to chromosome 13 YAC filters. When collating the map positions of positive YACs from three different RDA experiments a cluster of hits clearly identified the region on chromosome 13 which comprised the homozygous deletion. This shows that our novel approach can be effective. PMID- 10518627 TI - Editorial PMID- 10518626 TI - CapSelect: a highly sensitive method for 5' CAP-dependent enrichment of full length cDNA in PCR-mediated analysis of mRNAs. AB - Here we present CapSelect as a novel experimental approach for the selective enrichment of full-length cDNAs in PCR-mediated analysis of mRNA sequences. The method combines the 5'-CAP-dependent addition of specifically three to four non templated dCMP residues to the 3'-end of full-length cDNAs by reverse transcriptases in the presence of manganese and the controlled ribonucleotide tailing of cDNA ends by terminal deoxynucleotidyl transferase using rATP. By virtue of the generated terminal sequence motif (5'-dC(3-4)rA(3-4)), full-length cDNAs are selectively anchored to a double-stranded DNA adapter (with a dT(3 4)dG(3)3'-overhang) by T4 DNA ligase. The technique described is highly efficient, discriminates premature termination products and enriches full-length cDNAs. PMID- 10518628 TI - Future challenges for drug delivery. PMID- 10518629 TI - Selected advances in drug delivery and tissue engineering. PMID- 10518630 TI - Mechanistic aspects of iontophoresis in human epidermal membrane. AB - A large number of factors are involved in the movement of ions and molecules across human epidermal membrane (HEM) under the influence of an electric field. These factors and their interplay need to be understood if our knowledge of iontophoretic transport of drugs across HEM is to reach a point where physical models and strategies may be employed for useful quantitative predictions. In a typical in vitro experiment, the fully hydrated HEM is positioned between aqueous compartments of a two-chamber diffusion cell. When a low electric field is applied across the HEM under these conditions, the transport enhancement of ions in the pre-existing pores of the stratum corneum is the result of, (a) the direct interaction of the electric field with the charge of the ion in question, and (b) convective solvent flow (electroosmosis); in the case where the permeant is non ionic under these circumstances, transport enhancement is by convective solvent flow only. At moderate-to-high voltage iontophoresis (> or = around 1.0 V applied across a single HEM), in addition to the direct field effect and convective solvent flow in the pre-existing pores, there can generally be a significant (e.g. 10- to 100-fold enhancement) contribution to transport enhancement arising from new pore induction (electroporation). Much of the recent work in our laboratory has been devoted to defining and quantifying HEM electroporation, and an especially difficult aspect has been that of dealing with the large HEM membrane-to-membrane variabilities with regard to, (a) the extent of new pore induction, and (b) the characteristics of the newly induced pores. Recently we discovered that the extent of relevant (i.e. permeant accessible) pore induction may be correlated to the change in HEM electrical conductance (and quantifiable) if an appropriate matching background electrolyte can be selected having ion sizes comparable to that of the permeant. For example, employing tetraethylammonium (TEA) pivalate (PIV) for which the ion sizes are approximately 3.5 A, but not KCl (ion sizes approximately 1.9 A), as the background electrolyte for TEA (as the permeant) gave very good results; in this example, the sizable contribution of pore induction to iontophoresis was quantitatively factored out from the total iontophoretic enhancement. Experiments with a large number of HEM samples gave good agreement with the Nernst-Planck (N-P) predictions of the direct field effect when TEA-PIV was used as the background electrolyte for TEA transport, but large variations (up to 300%) between N-P predictions and experimental results were observed with KCl as the background electrolyte. Another area of recent effort has been HEM pore size determinations, both at low voltages (i.e. for pre-existing pores) and at voltages where the newly induced pores dominate HEM permeability. The sizes of pre-existing pores of HEM have been determined with the hindered diffusion theory (using experimental fluxes of several probe permeants of different known molecular sizes) to be generally in the range, 10-20 A, by a number of investigators in our laboratory for a large number HEM samples. Deducing pore sizes of electric field induced pores under steady electroporation conditions has been a more challenging task. We succeeded recently in developing a novel method for 'passively' determining pore sizes (i.e. by passive diffusion with hindered diffusion theory) under steady electroporation conditions: by using low frequency (12.5 Hz) a.c. at 2-5 V. We have been able to sustain electroporation at a nearly constant state of electroporation long enough to carry out a set of 'passive' diffusion experiments with relatively good precision to obtain the sizes of the newly induced pores. Studies to date have revealed that the sizes of pores induced with 2-5 V are of the same order of magnitude as those of the pre-existing pores (i.e. 10-20 A). Finally, another research question of interest has been that of pore charge PMID- 10518631 TI - Intestinal MDR transport proteins and P-450 enzymes as barriers to oral drug delivery. AB - Cytochrome P-450 3A4 (CYP3A4), the major phase I drug metabolizing enzyme in humans, and the multidrug efflux pump, MDR or P-glycoprotein (P-gp), are present at high levels in the villus tip enterocytes of the small intestine, the primary site of absorption for orally administered drugs. These proteins are induced or inhibited by many of the same compounds and demonstrate a broad overlap in substrate and inhibitor specificities, suggesting that they act as a concerted barrier to drug absorption. A series of studies from our laboratory of cyclosporine and tacrolimus in humans and a novel cysteine protease inhibitor in rats, dosed concomitantly with inhibitors and inducers of CYP3A4 and P-gp, suggest that gut extraction can be modeled using measures of intestinal metabolism and absorption rate, the latter reflecting changes in P-gp. Results evaluating a preliminary model applied to the CYP3A substrate drugs midazolam, indinavir, saquinavir, and rifabutin suggest that the model may be useful for predicting in vivo intestinal metabolism from in vitro data. PMID- 10518632 TI - Prescribability and switchability of highly variable drugs and drug products. AB - At the AAPS/FDS Workshop (Crystal City, Arlington, VA, March 6-8, 1995), it was agreed that some form of scaling should be permitted for highly variable drugs, although there was no agreement on a method. Currently, much emphasis is focused on developing a practical methodology for individual bioequivalence (IBE) and population bioequivalence (PBE) to replace or complement average bioequivalence (ABE). The latter requires only the mean bioavailabilities of two formulations to be sufficiently similar, whereas PBE also considers their distributions. IBE, on the other hand, is a comparison of the individual responses to the two formulations within subjects and is therefore concerned with switchability (interchangeability) between two multisource formulations. Multisource formulations refer (i) to generic copies of an innovator's formulation or (ii) to different formulations used in stages leading up to the final marked formulation. Evaluation of both PBE and IBE require replicate design studies. The FDA Working Group on IBE has been experimenting with methods in which a one-sided 95% confidence interval is computed based on the Bootstrap technique which ensures that the consumer risk is maintained at 5%. The IBE metric can then be scaled according to the within subject variance of the reference formulation. Thus if the variability of the test formulation (T) is higher than that of the reference (R), the formulation may fail IBE but not ABE. Conversely, if R is more variable than T, then the formulations may be considered to be IBE, even with a difference in means of more than 20%. Experimentation with existing data on our files shows that scaling has a considerable effect on the IBE decision for highly variable drugs. Evidence will also be presented to show that scaling makes the conditions more conservative for potent drugs with steep dose response curves reducing the risk of two generic formulations being BE with the same reference product but not BE with each other. On the other hand, broadening the BE limits for safe, highly variable drugs increases statistical power and reduces the number of subjects required. Even with the introduction of scaling, however, it is clearly difficult to obtain a single IBE criterion suitable to be applied to all drug products/studies. PMID- 10518633 TI - Setting bioequivalence requirements for drug development based on preclinical data: optimizing oral drug delivery systems. AB - The recently proposed Biopharmaceutics Classification System can be used to classify drugs and set standards for scale-up and post-approval changes as well as standards for in vitro/in vivo correlation for immediate and controlled release products. This classification scheme is based on determining the underlying process that is controlling the drug absorption rate and extent, namely, drug solubility and intestinal membrane permeability. Theoretical analysis and experimental results suggest that a permeability/solubility classification scheme can be used to set more rationale drug standards. In particular, high solubility/high permeability, rapidly dissolving drugs may be regulated on the basis of a single point rapid dissolution test while low solubility dissolution rate limited drugs can be regulated based on an in vitro dissolution test that reflects the in vivo dissolution process. This dissolution test may include multiple time points, media change, as well as surfactants in order to reflect the in vivo dissolution process and would be used by the manufacturer for requesting a waiver from a bioequivalence (BE) trial. For controlled release products, the regulation of bioequivalence standards is more complex due to the potential differences in position-dependent permeability/solubility and metabolism of drugs along the gastrointestinal tract. These differences may result in drug absorption rates that are highly transit time dependent. This paper will present the current status of the biopharmaceutic drug classification scheme, the underlying developed data base and its application to optimizing IR and CR products. PMID- 10518635 TI - Harmonization of the timing of non-clinical tests in relation to the conduct of clinical trials. PMID- 10518634 TI - Sterility assurance of microspheres. AB - The International Conference on Harmonization (ICH) provides a forum for constructive dialogues between regulatory authorities and the pharmaceutical industry on the real and perceived differences in the technical requirements for product registration in the EU, US and Japan. Achievement obtained so far is beyond expectation, having strong impacts both favorable and unfavorable on Japanese regulatory authorities and the pharmaceutical industry. The ICH guidelines are very science-oriented and little consideration seems to have been paid to the cultural and legal differences among the three regions. An example of such a difference is the interpretation of the guidelines between the US and Japan. In the US, they are generally recognized as good examples, whereas in Japan, they are usually taken as minimum and rigid requirements. A more flexible approach to guidelines will be necessary in the development of new dosage forms in Japan. In this paper, how to assure sterility of a new dosage form, that is, once-monthly injectable and biodegradable microspheres of Leuprorelin, a super agonist of LHRH, whose pharmaceutical development the author was in charge of at Takeda Chemical Industries, will be introduced. PMID- 10518636 TI - Drug delivery research in India: a challenge and an opportunity. PMID- 10518637 TI - Drug delivery research in Australasia: creating opportunities through collaboration. AB - This paper gives a brief overview of drug delivery research in Australia and New Zealand, known collectively as Australasia, in order to indicate future directions in this area of pharmaceutical research. It describes some of the relevant statistics of the two countries and briefly indicates some of the government incentives to foster the pharmaceutical industries. Examples of Australian drug delivery research such as a controlled release pelleted system for delivery of morphine are presented. The examples for New Zealand relate to veterinary systems such as an intraruminal device for zinc. The paper alludes to the need for collaboration among the various institutions in order to achieve successful outcomes. PMID- 10518638 TI - Future drug delivery research in South Korea. AB - According to the Science Citation Index database, over 50 papers related to drug delivery have been published from South Korea during the last 3 years. For the purpose of this presentation, some of the recently carried out research in our department will be introduced and future research directions presented. Proliposomes are free flowing particles which are composed of drugs, phospholipids and a water soluble porous powder, and immediately form a liposomal dispersion upon hydration. The preparation can be stored sterilized in a dried state and, by controlling the size of the porous powder in proliposomes, relatively narrow range of reconstituted liposome size can be obtained. Because of these properties, proliposomes appear to be a potential alternative to liposomes in design and fabrication of liposomal dosage forms. Thus, we tested the feasibility of this preparation as a sustained transdermal dosage form. Proliposomes containing varying amount of nicotine, a model drug, were prepared using sorbitol and lecithin. Microscopic observation revealed that this preparation is converted to liposomes almost completely within minutes following contact with water. The release pattern of the model drug from this preparation was apparently similar to that of the Exodus((R)) patch, a commercially available transdermal nicotine formulation. Compared with nicotine powder, the nicotine flux across rat skin from proliposomes was initially retarded, and appeared to remain constant. This observation indicated that sustained transdermal delivery of nicotine is feasible using proliposomal formulations under occluded condition. In addition to the investigation of the potential application of proliposomes, we were also interested in the role of the stratum corneum (SC) in the enhanced delivery of drugs from liposomes. Thus, liposome-gel formulation containing hydrocortisone (HC), a model hydrophobic drug, was prepared and used in this study. The study was carried out on both normal and stratum corneum removed skins. Percutaneous absorption of HC across SC removed skin was significantly faster than that across normal skin, suggesting that SC behaves as a penetration barrier for the liposome-bound drugs. Interestingly, the liposome-gel in this study reduced the skin absorption of HC, compared with the conventional ointment formulation. The amount of HC absorbed from the liposome-gel after 8 h into the SC-removed skin was less than one third of that from the conventional ointment. Despite the reduced absorption, a higher and sustained skin concentrations of HC were achieved for the liposome-gel. Drug concentration in both viable and deep skin reached a maximum within 0.5 h. However, drug concentrations in these tissues declined as a function of time for conventional ointment, while those from the liposome-gel were greatly sustained, resulting in a 5-fold higher viable skin drug level was obtained at 8 h after the application. In contrast, plasma concentration of HC at 4 h from the liposome-gel was only one-fourth the value from the conventional ointment in the SC-removed skin. Therefore, the higher and sustained drug concentration in the viable skin appeared not to be due to the enhanced percutaneous absorption but due to retarded diffusion of the drug from the skin. PMID- 10518639 TI - Poly(ethylene glycol)-containing hydrogels in drug delivery. AB - The use of hydrogels as carriers for protein delivery has been a subject of significant recent research. In our recent work, we have shown that diffusion controlled delivery of proteins from hydrogels containing poly(ethylene glycol) (PEG) can be possible and controlled by the three-dimensional structure. In addition, a number of these hydrogel carriers are mucoadhesive and can be used for protein delivery. PEG star polymer gels have also been prepared by gamma irradiation and have been used for protein delivery with and without molecular imprinting. The presence of a large number of functional groups in a small volume makes these polymers important for use in biological and pharmaceutical applications. PEG star polymer hydrogels were synthesized using gamma-irradiation and were characterized using swelling techniques. Equilibrium swelling studies were conducted to investigate the effects of molecular weight, number of star arms, concentration, and radiation dose. PMID- 10518640 TI - Preparation and characterization of polymer micelles from poly(ethylene glycol) poly(D,L-lactide) block copolymers as potential drug carrier. AB - Poly(ethylene glycol)-poly(D,L-lactide) block copolymers (PEG-PLA) with varying composition were prepared through successive ring-opening polymerization of ethylene oxide and D,L-lactide using an anionic initiator, and their property of multimolecular micellization in aqueous milieu was examined in detail from the standpoint of designing carriers for hydrophobic drugs. The heterogeneity of PEG PLA was found to crucially affect the size and distribution of micelles, and narrowly-distributed micelles with sizes of approximately 30 nm in diameter were formed only from PEG-PLA with a substantially narrow molecular weight distribution and an appropriate balance in the length ratio of the PEG and PLA segments in PEG-PLA, indicating the importance of establishing a reliable synthetic route for the block copolymers. PEG-PLA micelles have a considerably low critical association concentration (approximately 1.0 mg/l) which is apparently an advantage in utilizing these micelles as drug carriers in an extremely diluted condition. PMID- 10518641 TI - Mucosal drug delivery using cellulose derivatives as a functional polymer. AB - In order to develop nasal powder preparations with higher bioavailability for peptide delivery, the effect of a combination of hydroxypropyl cellulose (HPC) and microcrystalline cellulose (MCC) used as base materials and microenvironment for the drugs in the preparations was examined. Significant enhanced absorption of leuprolide, calcitonin and FITC-dextran was attained by the addition of a small amount of HPC to MCC. It is suggested that MCC works as an absorption enhancer by causing a locally high concentration of drugs in the vicinity of the mucosa surface. On the other hand, HPC works to increase retention of drugs on the nasal mucosa due to its gel-forming property. In comparison to the powder mixtures of MCC and HPC in which drugs were distributed to the MCC or HPC, selective distribution of drugs to MCC is shown to be a critical factor for enhancing nasal absorption. These results showed that the microenvironment of drugs in the functional base material could critically affect the nasal absorption of drugs. PMID- 10518642 TI - New biodegradable polymers for injectable drug delivery systems. AB - Many biodegradable polymers were used for drug delivery and some are successful for human application. There remains fabrication problems, such as difficult processability and limited organic solvent and irreproducible drug release kinetics. New star-shaped block copolymers, of which the typical molecular architecture is presented, results from their distinct solution properties, thermal properties and morphology. Their unique physical properties are due to the three-dimensional, hyperbranched molecular architecture and influence microsphere fabrication, drug release and degradation profiles. We recently synthesized thermosensitive biodegradable hydrogel consisting of polyethylene oxide and poly(L-lactic acid). Aqueous solution of these copolymers with proper combination of molecular weights exhibit temperature-dependent reversible sol-gel transition. Desired molecular arrangements provide unique behavior that sol (at low temperature) form gel (at body temperature). The use of these two biodegradable polymers have great advantages for sustained injectable drug delivery systems. The formulation is simple, which is totally free of organic solvent. In sol or aqueous solution state of this polymer solubilized hydrophobic drugs prior to form gel matrix. PMID- 10518643 TI - Thermo-responsive drug delivery from polymeric micelles constructed using block copolymers of poly(N-isopropylacrylamide) and poly(butylmethacrylate). AB - To achieve a combination of spatial specificity in a passive manner with a stimuli-responsive targeting mechanism, a temperature-responsive polymeric micelle is prepared using block copolymers of (poly(N-isopropylacrylamide-b butylmethacrylate) (PIPAAm-PBMA)). The micelle inner core formed by self aggregates of PBMA segments successfully loaded with a drug (adriamycin), and the outer shell of PIPAAm chains played a role of stabilization and initiation of micellar thermo-response. Optimum conditions were investigated for the micelle formation and drug loading into the inner cores in a view of micellar stability and function as drug carriers. Outer shell hydrophilicity that prevents inner core interaction with biocomponents and other micelles can be suddenly switched to hydrophobic at a specific site by local temperature increase beyond the LCST (lower critical solution temperature) (32.5 degrees C). These micelles showed reversible structural changes allowing drug release upon heating/cooling thermal fluctuations through the LCST. Polymeric micelles incorporated with adriamycin showed a dramatic thermo-responsive on/off switching behavior for both drug release and in vitro cytotoxicity according to the temperature responsive structural changes of a micellar shell structure. The reversible and sensitive thermo-response of the micelle opens up opportunities to construct a novel drug delivery system in conjunction with localized hyperthermia. PMID- 10518644 TI - Biopharmaceutics of transmucosal peptide and protein drug administration: role of transport mechanisms with a focus on the involvement of PepT1. AB - Non-invasive delivery of peptide and protein drugs will soon become a reality. This is due partly to a better understanding of the endogenous transport mechanisms, including paracellular transport, endocytosis, and carrier-mediated transport of mucosal routes of peptide and protein drug administration. This paper focuses on work related to the elucidation of structure-function, intracellular trafficking, and regulation of the intestinal dipeptide transporter, PepT1. PMID- 10518645 TI - Physiological mechanism for enhancement of paracellular drug transport. AB - We examined the action mechanisms of enhancers that improve paracellular drug transport. For sodium caprate (C10), the increase in the intracellular calcium level was considered to induce the contraction of calmodulin-dependent actin filaments, followed by dilation of the paracellular pathway. Although decanoylcarnitine (DC) also increased the intracellular calcium level, the action was independent of calmodulin and thus, the action mechanism of acylcarnitines was considered to differ from that of C10. Other acylcarnitines, lauroylcarnitine (LC) and palmitoylcarnitine (PC) and organic acids, tartaric acid (TA) and citric acid (CA) decreased the intracellular ATP level and the intracellular pH. From these results, it was considered that one of the action mechanism of acylcarnitines and organic acids is that the intracellular acidosis increases the calcium level through the decrease in ATP levels, followed by opening the tight junction. Membrane dysfunction which was expected from the above mechanism was assessed by the transport function of electrolytes. Membrane conductance, which was increased by C10, LC and PC, returned to the control value during a 3- to 6-h recovery period. On the other hand, Cl(-) ion secretion, which was obtained from short-circuit current (I(sc)), was decreased by these enhancers, but was normalized by C10 but not by LC and PC. Accordingly, C10 can be considered a safer enhancer than acylcarnitines. PMID- 10518646 TI - Recent advances in buccal drug delivery and absorption--in vitro and in vivo studies. AB - In the first part of this study, the aim was to characterize transport of fluorescein isothiocyanate (FITC)-labelled dextrans of different molecular weights as model compounds for peptides and proteins through buccal mucosa. The penetration of these dextrans through porcine buccal mucosa (a nonkeratinized epithelium, comparable to human buccal mucosa) was investigated by measuring transbuccal fluxes and by analyzing the distribution of the fluorescent probe in the epithelium, using confocal laser scanning microscopy for visualizing permeation pathways. The results revealed that passage of hydrophilic compounds such as the FITC-dextrans through porcine buccal epithelium is restricted to permeants with a molecular weight lower than 20 kDa. The permeabilities of buccal mucosa for the 4 and 10 kDa FITC-dextran (of the order of 10(-8) cm/s) were not significantly different from each other or from the much smaller compound FITC. The confocal images of the distribution pattern of FITC-dextrans showed that the paracellular route is the major pathway through buccal epithelium. In the in vivo part of this study, buccal delivery of FITC-labelled dextran 4400 (FD4) and the peptide drug buserelin was investigated in vivo, in pigs. The delivery device consisted of an application chamber with a solution of FD4 or buserelin, and was attached to the buccal mucosa for 4 h using an adhesive patch. A randomized cross over study including intravenous administration and buccal delivery without and with 10 mM sodium glycodeoxycholate (GDC) as an absorption enhancer was performed in pigs. After buccal administration, steady-state plasma levels were rapidly achieved. Co-administration of 10 mM GDC increased the absolute bioavailability from 1.8+/-0.5 to 12.7+/-2.0% for FD4. From the present studies, it is concluded that buccal administration is a suitable route for the delivery for macromolecules and hydrophilic compounds such as peptide drugs. PMID- 10518647 TI - Formula optimization based on artificial neural networks in transdermal drug delivery. AB - The promoting effect of O-ethylmenthol (MET) on the percutaneous absorption of ketoprofen from alcoholic hydrogels was evaluated in rats in vitro and in vivo. Furthermore, a novel simultaneous optimization technique incorporating an artificial neural network (ANN) was applied to a design of a ketoprofen hydrogel containing MET. When a small quantity of MET (0.25-0.5%) was added to the hydrogels, the permeation of ketoprofen increased remarkably, compared with the control. On the other hand, little change in permeation was observed when small amounts of menthol were used (<1%), and at least 2% menthol was required to obtain a promoting efficiency comparable with 0.25% MET. The partitioning of ketoprofen from the hydrogel to the skin was improved by the addition of a small amount of MET, whereas the diffusivity of the drug was enhanced at higher concentration of MET (0.5-1%). For the optimization study, the amount of ethanol and MET were selected as causal factors. A rate of penetration (R(p)) and lag time (t(L)) and total irritation score (TIS) were selected as response variables. A set of causal factors and response variables was used as tutorial data for ANN and fed into a computer. Nonlinear relationships between the causal factors and the response variables were represented well with the response surface predicted by ANN. The optimization of the ketoprofen hydrogel was performed according to the generalized distance function method. The observed results of R(p) and TIS, which had a lot of influence on the effectiveness and safety, coincided well the predictions. PMID- 10518648 TI - Transport of human growth hormone across Caco-2 cells with novel delivery agents: evidence for P-glycoprotein involvement. AB - Emisphere Technologies, Inc. has synthesized a series of small molecules which have been shown to improve protein absorption through mucosal tissue. This enhancement is specific between protein and a particular delivery agent. Despite the specificity of interaction, the mechanism of enhanced tissue penetration is still unclear. The purpose of this work is to understand the enhancement mechanism(s) of these delivery agents by using Caco-2 cells as a model membrane. It was found that the bidirectional transepithelial fluxes of human growth hormone (hGH) in the presence of these delivery agents across human intestinal epithelial Caco-2 cell line showed marked asymmetry. Average permeability coefficient values obtained in the apical (AP) to basolateral (BL) direction were lower than those of the reverse (BL to AP) direction. On the other hand, the fluxes for human growth hormone alone were symmetric. When P-glycoprotein inhibitors were included in the transport medium, the permeability coefficient values of BL to AP direction were significantly decreased while the transport was increased in the reverse direction in the presence of delivery agents. P glycoprotein inhibitors had no effect on the transport of human growth hormone alone. This study shows that human growth hormone alone can be transported across Caco-2 cells in very limited quantities by passive diffusion, but in the presence of delivery agents, human growth hormone can be effluxed in a P-glycoprotein mediated fashion. This also indirectly shows that the human growth hormone has become more lipophilic in the presence of delivery agents. PMID- 10518649 TI - Kinetic and biochemical analysis of carrier-mediated efflux of drugs through the blood-brain and blood-cerebrospinal fluid barriers: importance in the drug delivery to the brain. AB - In this manuscript, our recent studies on the transporters on the blood-brain barrier and blood-cerebrospinal fluid (CSF) barrier responsible for the excretion of ligands from the central nervous system (CNS) to the blood are summarized. By comparing the brain entry of quinidine in normal and mdr 1a knock out mice, the predominant role of P-glycoprotein in the brain distribution of this compound was demonstrated. In addition to P-glycoprotein, the presence of transporters responsible for the efflux of organic anions from the brain has been suggested by a pharmacokinetic analysis of the CNS distribution of cefodizime, a third generation cephalosporin antibiotic. This suggestion was confirmed by demonstrating the presence of a specific mechanism for the elimination of p aminohippuric acid from the brain after microinjection into the cerebral hemisphere. In vitro, the energy-dependent luminal preferential efflux of glutathione-bimane was demonstrated in a monolayer of MBEC4 cells which were derived from mouse brain endothelial cells. Studies with isolated membrane vesicles from MBEC4 cells suggested the presence of a primary active transporter(s) for organic anions, and Western blot analysis indicated the presence of multidrug resistance associated protein (MRP1) and/or its related transporters on MBEC4 cells and freshly isolated rat cerebral endothelial cells. The transcellular transport of 17beta estradiol 17beta-D-glucuronide (E(2)17betaG) across the choroid plexus was also demonstrated by examining the efflux of this compound from CSF after intracerebroventricular administration. The functional significance of organic anion transporting polypeptide (oatp-1) on the brush border membrane of the choroid plexus was demonstrated by comparing the uptake of E(2)17betaG into the isolated choroid plexus and oatp-1 transfected COS 7 cells; in addition, reverse transcription-polymerase chain reaction and Western blot analysis indicated the presence of MRP in the choroid plexus. Together with the direction of transcellular transport, the basolateral localization of MRP on the choroid plexus was suggested. By regulating the activity of these efflux transporters, it is possible to improve the brain entry of certain substrates. PMID- 10518650 TI - Pharmacokinetic considerations in the design of better and safer new antiepileptic drugs. AB - Valproic acid (VPA) is one of the major antiepileptic drugs. However, its anticonvulsant potency is less than the other three major antiepileptic drugs. Furthermore, VPA causes two rare but severe side effects: teratogenicity and hepatotoxicity. We utilized pharmacokinetic considerations in designing various amide derivatives of VPA which are more potent as anticonvulsants than VPA and have the potential to be nonteratogenic and nonhepatotoxic. The following three groups of VPA derivatives were designed and evaluated: (1) Isomers of valpromide (VPD) in order to explore the structural requirements for metabolically stable VPD isomers. Two chiral amides, valnoctamide and propylisopropyl acetamide, have emerged from a stereospecific study as the optimal compounds; (2) Cyclic amide derivatives of VPD. N-Methyl 2,2,3, 3-tetramethylcyclopropane carboxamide (M TMCD) was found to be the optimal compound in this series. M-TMCD is a stable achiral VPD analogue acid which is nonteratogenic. Since M-TMCD contains four methyl substituents it cannot form a metabolite with a terminal double bond, and thus has the potential to be a nonhepatotoxic compound; (3) Conjugation products of VPA and gamma-amino butyric acid (GABA) or glycine. N-valproyl glycinamide (VGD) emerged as the best compound out of this group and is currently undergoing phase II clinical trials. VGD is mainly metabolized to N-valproyl glycine by a nonoxidative hydrolytic metabolic pathway. It did not operate as chemical drug delivery systems of VPA and glycine or GABA, but acted rather as a drug on its own. PMID- 10518651 TI - Analysis of skin disposition of flurbiprofen after topical application in hairless rats. AB - Cutaneous disposition of topically applied flurbiprofen (FP) was evaluated using a new in situ experimental model in hairless rats. A disk-shaped agar gel (3.85 cm in diameter and 0.5 cm in thickness) was subcutaneously implanted in the abdominal region of rats as a drug receptor, and a drug donor cell was subsequently placed above this agar gel. No significant pharmacokinetic modification of FP was observed as a result of this experimental procedure. A bolus injection and a constant intravenous infusion of FP were applied to the rats, followed by an analysis of FP levels in the plasma and agar gels. Using these results, the clearance rate of FP from the systemic circulation to the gel could be calculated. FP (1% gel formulation, 1.0 g/3.14 cm(2)) was then topically applied to the skin of these rats. From these experiments, the amount of FP that migrated from the formulation to the systemic circulation and the amount of FP that migrated directly to the agar gel across the skin, over 10 h, were separately evaluated to be 235.4 and 2.0 microg, respectively. Thus, most of the FP was absorbed into the systemic circulation. The effect of endogeneous vasoactive compounds and penetration enhancers on the FP disposition within skin was also determined. Epinephrine and bradykinin were used as vasoactive compounds that were entrapped in agar gel, and an isopropyl myristate system (IPM system) and a l-menthol-ethanol-glycerin-water system (MEGW system) were used as enhancers in the formulation. Epinephrine enhanced the direct delivery of FP into the agar gel to more than ten times its former level, in spite of the fact that it had no effect on systemic delivery. Bradykinin strengthened systemic delivery slightly, without changing the quantity of FP in the gel. IPM increased only the systemic delivery of FP, as was the case with bradykinin, whereas the MEGW system markedly increased both the blood concentration and the quantity of FP in the gel (13 and 200 times, respectively). This technique has proven to be an effective tool for the quantitative evaluation of cutaneous disposition of a topically applied drug. PMID- 10518652 TI - Simultaneous transport and metabolism of ethyl nicotinate in hairless rat skin after its topical application: the effect of enzyme distribution in skin. AB - An in vitro permeation study of ethyl nicotinate (EN) was carried out using excised hairless rat skin, and simultaneous skin transport and metabolism of the drug were kinetically followed. Fairly good steady-state fluxes of EN and its metabolite nicotinic acid (NA) through the skin were obtained after a short lag time for all the concentrations of EN applied. These steady-state fluxes were not proportional to the initial donor concentration of EN: EN and NA curves were concave and convex, respectively, which suggests that metabolic saturation from EN to NA takes place in the viable skin at higher EN application. Further permeation studies of EN or NA were then carried out on full-thickness skin or stripped skin with an esterase inhibitor to measure their permeation parameters, such as partition coefficient of EN from the donor solution to the stratum corneum and diffusion coefficients of EN and NA in the stratum corneum and the viable epidermis and dermis. Separately, enzymatic parameters (Michaelis constant K(m) and maximum metabolism rate V(max)) were obtained from the production rate of NA from different concentrations of EN in the skin homogenate. The obtained permeation and enzymatic parameters were then introduced to differential equations showing Fick's second law of diffusion in the stratum corneum and the law with Michaelis-Menten metabolism in the viable epidermis and dermis. The calculated steady-state fluxes of EN and NA by the equations were very close to the obtained data. We then measured the esterase distribution in skin microphotographically using fluorescein-5-isothiocyanate diacetate. A higher enzyme concentration was observed in the epidermal cells and near hair follicles than in the dermis. Simulation studies using the even and the partial enzyme distribution models suggested that no significant difference between the models was observed in the skin permeations of EN and NA, whereas concentration-distance profiles of EN and NA were very different. This finding suggests that the total amount of enzyme in skin which converts EN to NA is a determinant of the metabolic rate of EN in skin. The present approach is a useful tool for analyzing simultaneous transport and metabolism of many drugs, especially those showing Michaelis-Menten type-metabolic saturation in skin. PMID- 10518653 TI - Recent advances in retrometabolic design approaches. AB - The retrometabolic drug design approaches simultaneously incorporate structure activity (SAR) and structure metabolism (SMR) relationships in the design process. Two major approaches were developed, the chemical delivery systems (CDS), which allow chemical-enzymatic targeting of drugs via strategic sequential enzymatic activation of the inactive CDSs. On the opposite end of the retrometabolic design loop are the soft drugs (SD), which are designed to have highly improved therapeutic indeces by controlling their metabolism, after they achieve their therapeutic role. One of the most successful SD class is the 'inactive metabolite approach', where the design starts from an inactive metabolite of a drug. Its strategic manipulation yields an isosteric/isoelectronic drug analog, which is enzymatically deactivated to the very inactive metabolite at the desired compartment and with controlled rate. Overall, retrometabolic approaches represent a complex collection of chemical enzymatic means for the design of safer drugs and for their controlled release. Most recent advances involve FDA approval of a soft steroid, as well as the first successful brain targeting of various neuropeptides and their brain-targeted analogs. PMID- 10518654 TI - Improving the pharmacokinetic and pharmacodynamic properties of a drug by chemical conversion to a chimera drug. AB - The purpose of the present study is to investigate a new concept which involves the conjugation of two drugs having different pharmacological activities which is termed a 'chimera drug (mutual prodrug)', in drug delivery optimization using a chemical modification approach. The conjugates of FP, an NSAID, with histamine H(2) antagonists were synthesized, in order to investigate the reduction in gastric damage by NSAID, and their pharmaceutical, pharmacokinetic and pharmacological properties were examined. The limited data obtained herein indicate that the chimera drug composed of FP and PPA was effective in reducing gastric damage by FP, with no changes in its biopharmaceutical properties, compared with the conventional prodrugs such as ester-type prodrugs. PMID- 10518656 TI - Tissue selective drug delivery utilizing carrier-mediated transport systems. AB - This paper describes some successful examples of a tissue selective drug delivery by utilizing specialized transporter(s) expressed in the targeted tissue cells. These are as follows: (1) oral delivery via H(+)/oligopeptide transporter, rat or human Pept1, in the intestine for beta-lactam antibiotics and a newly synthesized dipeptide, L-dopa-L-phenylalanine; (2) tumor cell specific delivery via the newly discovered H(+)/oligopeptide transporter(s) expressed in human fibrosarcoma cell line HT-1080 for model oligopeptides, glycylsarcosine and carnosine; (3) oral and hepatic delivery via an H(+)/monocarboxylate transporter in the intestine and an organic anion transporter in the liver for HMG-CoA reductase inhibitor, pravastatin; and (4) lung selective delivery via some type of transporter and avoidance of transfer into the brain via P-glycoprotein at the blood-brain barrier for a new quinolone antibacterial, HSR-903. PMID- 10518655 TI - Optimizing oral absorption of peptides using prodrug strategies. AB - Prodrug strategies applied to peptides have tended to focus on modification of a single functional group (e.g., N-terminal end). Recently, our laboratory introduced the concept of making cyclic prodrugs of peptides as a way to modify their physicochemical properties sufficiently to allow them to permeate biological barriers (i.e., intestinal mucosa). This cyclization strategy required the development of new 'chemical linkers,' including an acyloxyalkoxy linker, a phenylpropionic acid linker, and a coumarinic acid linker. All three chemical linkers were designed to be susceptible to esterase metabolism (slow step), leading to a cascade of chemical reactions (fast steps) that result in release of the peptide. These cyclic prodrug strategies have been applied to opioid peptides in an attempt to stabilize them to metabolism and/or improve their intestinal mucosal permeation. Specifically, we prepared acyloxyalkoxy-, phenylpropionic acid- and coumarinic acid-based cyclic prodrugs of [Leu(5)]-enkephalin (H-Tyr-Gly Gly-Phe-Leu-OH) and its metabolically stable analog DADLE (H-Tyr-D-Ala-Gly-Phe-D Leu-OH) and determined their metabolic and biopharmaceutical properties. The cyclic prodrugs of these opioid peptides were shown to have: (i) favorable physicochemical properties (e.g., increased lipophilicity) for membrane permeation; (ii) unique solution structures (e.g., beta-turns) that reduce their hydrogen bonding potential; and (iii) metabolic stability to exo- and endopeptidases. The cell membrane permeation characteristics of [Leu(5)] enkephalin, DADLE and the cyclic peptide prodrugs were evaluated using Caco-2 cell monolayers, a cell culture model of the intestinal mucosa. The phenylpropionic acid- and coumarinic acid-based cyclic prodrugs of [Leu(5)] enkephalin and DADLE were shown to have significantly better cell permeation characteristics than the parent opioid peptides. Furthermore, these cyclic prodrugs were shown to be transcellular permeants (in contrast to the opioid peptides, which are paracellular permeants), and were not substrates for polarized efflux systems. Surprisingly, the acyloxyalkoxy-based prodrugs of [Leu(5)]-enkephalin and DADLE were shown to exhibit very low permeation through Caco-2 cell monolayers, which could be attributed to their substrate activity for efflux systems. PMID- 10518657 TI - Liposomes in autoimmune diseases: selected applications in immunotherapy and inflammation detection. AB - In this contribution three examples are discussed of ongoing research where liposomes are used as carrier systems for immunotherapy and inflammation detection in autoimmune diseases. Liposomes can be used as carrier systems of antigenic peptides to peripheral blood mononuclear cells. The second example deals with their use as carrier systems for MHC-peptide complexes for multivalent Ag-presentation to autoreactive T lymphocytes to specifically modulate the activity of these T lymphocytes. The third example relates to our work on long circulating liposomes which are currently being tested in man for their potential to image inflammation sites. PMID- 10518659 TI - Cyclodextrins and carrier systems. AB - This paper describes two new possibilities of using cyclodextrins to increase water solubility and bioavailability of poorly water-soluble drugs intended for targeting delivery by the oral or the parenteral route. They use either amphiphilic cyclodextrin nanoparticles or polymeric nanoparticles containing cyclodextrins. Amphiphilic skirt-shaped cyclodextrins, resulting from the esterification of primary hydroxyl groups by hydrocarbon chains varying from C6 to C14, are capable of forming spontaneously nanoparticles which have been loaded with a series of steroid drugs. The drug in the amphiphilic cyclodextrin nanoparticles is molecularly dispersed and can be released very rapidly. Poly(isobutylcyanoacrylate) nanoparticles can be loaded with natural or hydroxypropyl cyclodextrins. This technique results in a significant increase in the loading capacity of nanoparticles with a series of steroids and in a very rapid release of the drug. Both methods are described as well as their potential interest for water-insoluble drugs. PMID- 10518658 TI - Design of polymeric prodrugs of prostaglandin E(1) having galactose residue for hepatocyte targeting. AB - Based on the relationship between in vivo disposition of macromolecules and their physicochemical and biological characteristics obtained through clearance concept based pharmacokinetic analysis, polymeric prodrugs of prostaglandin E(1)(PGE(1)) were designed stepwise and evaluated on their targeting and therapeutic efficiencies. First poly-L-lysine (PLL) and poly-L-glutamic acid (PLGA) with an ethylenediamine (ED) spacer were modified with 2-imino-2-methoxyethyl 1 thiogalactoside to obtain galactosylated derivatives. After intravenous injection in mice, Gal-ED-PLGA was selectively taken up by the liver parenchymal cells via receptor-mediated endocytosis, while Gal-PLL accumulated in the liver as well as PLL mostly due to electrostatic interaction. Although Gal-ED-PLGA showed good targeting efficacy, its PGE(1) conjugate synthesized with activated PGE(1) by carbonyldiimidazole method failed to show therapeutic effects probably due to inactivation of PGE(1) during conjugation and lack of release in the tissue. In order to overcome these problems, we next conjugated PGE(1) to galactosylated poly-(L-glutamic acid) hydrazide (Gal-HZ-PLGA) in which PGE(1) was easily coupled to Gal-HZ-PLGA via a hydrazone bond in weak acidic solution (pH 5) at room temperature. The PGE(1)-Gal-HZ-PLGA conjugate labeled with [(111)In] or [(3)H]PGE(1) rapidly accumulated in the liver parenchymal cells. In addition, the PGE(1) conjugate effectively inhibited the increase of the GPT level in plasma, while free PGE(1) indicated no therapeutic efficacy even at more than ten times higher doses, in carbon tetrachloride-induced hepatitis mice. These findings suggest potentials of polymeric targeting systems of PGE(1) to hepatocyte utilizing galactose recognition. PMID- 10518660 TI - Gene delivery using liposome technology. AB - Development of more reliable liposomal formulations and preparation methods which can be used for gene therapy instead of commonly used viral vectors is expected. We have already developed the freeze-dried empty (non-drug-containing) liposomes (FDEL) method for mass-production of liposomal products. After these freeze-dried empty liposomes are rehydrated with aqueous drug solutions, many kinds of drugs can be encapsulated highly efficiently, and particle size can be controlled well. This study evaluated the usefulness of this FDEL method for preparation of liposomes containing DNA with a particular attention to the stability of DNA. When the liposomes were prepared by the conventional lipid-film method on a relatively large scale with use of a Potter-homogenizer (a teflon homogenizer), significant degradation and conformational change of DNA was observed during homogenization. Loss of DNA was also significant after extrusion for sizing and sterilization; residual DNA in the final preparation was hardly detected. When the FDEL method was used, on the other hand, no degradation, conformational change or loss of DNA was observed, and particle size was easily controlled. Moreover, there was no significant difference in luciferase activity between the lipid-film method used on a small scale with use of a vortex mixer and the FDEL method after transfection of tumor cells (HRA, HEC-1A and Colo320DM) by the liposomes containing DNA (PGV-C). These findings suggest that the FDEL method is very useful for preparation of liposomes containing DNA. PMID- 10518661 TI - Pulmonary delivery of insulin with nebulized DL-lactide/glycolide copolymer (PLGA) nanospheres to prolong hypoglycemic effect. AB - Insulin loaded PLGA nanospheres having weight mean diameters of 400 nm were prepared by the modified emulsion solvent diffusion method in water. The nanosphere recovery and the drug recovery in the nanospheres were 74.8%+/-4.71 and 46.8%+/-7.01, respectively. Eighty five percent of the drug was released from the nanospheres at the initial burst, followed by prolonged releasing of the remaining drug for a few hours in saline at 37 degrees C. The aqueous dispersions (6 mg/ml) of PLGA nanospheres were nebulized by a sieve type ultrasonic nebulizer to discrete droplets of 5 approximately 7 microm in mean diameters, 75% of which were successfully delivered into the alveolar fraction in a cascade impactor inhaled at 28.3 l/min. The nebulized PLGA nanospheres were administered via a spacer by using a constant volume respirator into the trachea of the fasted guinea pig for 20 min. After the administration of 3.9 I.U./kg insulin with the PLGA nanospheres, the blood glucose level was reduced significantly and the hypoglycemia was prolonged over 48 h, compared to the nebulized aqueous solution of insulin as a reference (6 h). This result could be attributed to the sustained releasing of insulin from the nanospheres deposited widely on to the whole of lung. PMID- 10518662 TI - Mineral precipitation and porosity losses in granular iron columns. AB - As permeable reactive barriers containing zero-valent iron are becoming more widely used to remediate contaminated groundwaters, there remains much uncertainty in predicting their long-term performance. This study focuses on two factors affecting performance and lifetime of the granular iron media: plugging at the treatment zone entrance and precipitation in the bulk iron media. Plugging at the system entrance is due principally to mineral precipitation promoted by dissolved oxygen in the influent groundwater and is an issue in aerobic aquifers or in above-ground canister tests. Designs to minimize plugging in field applications where the groundwater is oxygenated include the use of larger iron particles and admixing sand of comparable size with the iron particles. Beyond the entrance zone, the groundwater in anaerobic and mineral precipitation leads to porosity losses in the bulk iron media, potentially reducing flow through the treatment zone. The nature of the mineral precipitation and the factors that affect extent of mineral precipitation have been examined by a variety of tools, including tracer tests, aqueous inorganic profiles, and surface analytical techniques. At short treatment times, porosity losses as measured by tracer tests are due mainly to Fe(OH)(2) precipitates and possible entrapment of a film of hydrogen gas on the iron surfaces. Over longer treatment times, precipitation of Fe(OH)(2) and FeCO(3) in low carbonate waters and of Fe(OH)(2), FeCO(3) and CaCO(3) in higher carbonate waters begin to dominate porosity losses. The control of pH within the iron media by addition of ferrous sulfide was shown not to reduce significantly calcium and carbonate precipitates, indicating that mineral precipitation is controlled by more than simple carbonate equilibrium considerations. PMID- 10518663 TI - Hydrogeologic modeling for permeable reactive barriers. AB - The permeable reactive barrier technology for in situ treatment of chlorinated solvents and other groundwater contaminants is becoming increasingly popular. Field scale implementation of this and other in situ technologies requires careful design based on the site-specific hydrogeology and contaminant plume characteristics. Groundwater flow modeling is an important tool in understanding the hydraulic behavior of the site and optimizing the reactive barrier design. A combination of groundwater flow modeling and particle tracking techniques was used to illustrate the effect of hydraulic conductivity of the aquifer and reactive media on key permeable barrier design parameters, such as the capture zone width, residence time, flow velocity, and discharge. Similar techniques were used to illustrate the modeling approach for design of different configurations of reactive barriers in homogeneous and heterogeneous settings. PMID- 10518664 TI - Design and construction techniques for permeable reactive barriers. AB - Adequate site characterization, bench-scale column testing, and hydrogeologic modeling formed the basis for the design and construction of permeable reactive barriers for groundwater remediation at various sites, such as Dover Air Force Base, DE and Naval Air Station, Moffett Field, CA. Dissolved chlorinated solvents, such as perchloroethylene (PCE) and trichloroethylene (TCE), have been the focus at many sites because the passive nature of the reactive barrier operation makes such barriers particularly useful for treating groundwater contaminants that can persist in the aquifer for several years. A combination of conventional and innovative site characterization, design, and construction techniques were used at these sites to increase the potential cost effectiveness of field application. PMID- 10518665 TI - Effects of aquifer heterogeneity and reaction mechanism uncertainty on a reactive barrier. AB - This paper addresses impacts of aquifer heterogeneity and reaction mechanism uncertainty on permeable reactive barrier (PRB) performance and describes modeling tools and preliminary guidelines for risk-based design of reactive barriers at heterogeneous sites. A braided stream aquifer was generated stochastically, using a fixed correlation structure and four levels of variability in the hydraulic conductivity field. A vertical, homogeneous barrier was placed in the aquifer. Based on a deterministic design, the size of the PRB for uniform conditions was considered conservative (factor of safety=3.3). Monte Carlo simulation was used to model cis 1,2-DCE reduction by iron metal with uncertainty in the reaction mechanism rate constants. These results were combined with flow and particle tracking results to predict the spatial distribution and flow-averaged concentrations of cis 1,2-DCE and vinyl chloride at the exit face of the PRB. Evaluated on a risk basis, the deterministic design method was found to be unconservative for more heterogeneous aquifers. Uncertainty in the reaction mechanism accentuated the negative effects of aquifer heterogeneity. Several compensating factors that may reduce the vulnerability of reactive barriers to aquifer heterogeneity are discussed. PMID- 10518666 TI - Performance evaluation of a permeable reactive barrier for remediation of dissolved chlorinated solvents in groundwater. AB - A pilot-scale permeable reactive barrier (PRB) consisting of granular iron was installed in May 1995 at an industrial facility in New York to evaluate the use of this technology for remediation of chlorinated volatile organic compounds (VOCs) in groundwater. The performance of the barrier was monitored over a 2-year period. Groundwater velocity through the barrier was determined using water level measurements, tracer tests, and in situ velocity measurements. While uncertainty in the measured groundwater velocity hampered interpretation of results, the VOC concentration data from wells in the PRB indicated that VOC degradation rates were similar to those anticipated from laboratory results. Groundwater and core analyses indicated that formation of carbonate precipitates occurred in the upgradient section of the iron zone, however, these precipitates did not appear to adversely affect system performance. There was no indication of microbial fouling of the system over the monitoring period. Based on the observed performance of the pilot, a full-scale iron PRB was installed at the site in December 1997. PMID- 10518667 TI - Long-term performance monitoring for a permeable reactive barrier at the U.S. Coast Guard Support Center, Elizabeth City, North Carolina. AB - A continuous hanging iron wall was installed in June, 1996, at the U. S. Coast Guard (USCG) Support Center near Elizabeth City, NC, United States, to treat overlapping plumes of chromate and chlorinated solvent compounds. The wall was emplaced using a continuous trenching machine whereby native soil and aquifer sediment was removed and the iron simultaneously emplaced in one continuous excavation and fill operation. To date, there have been seven rounds (November 1996, March 1997, June 1997, September 1997, December 1997, March 1998, and June 1998) of performance monitoring of the wall. At this time, this is the only full scale continuous 'hanging' wall installed as a permeable reactive barrier to remediate both chlorinated solvent compounds and chromate in groundwater. Performance monitoring entails the following: sampling of 10-5 cm PVC compliance wells and 15 multi-level samplers for the following constituents: TCE, cis dichloroethylene (c-DCE), vinyl chloride, ethane, ethene, acetylene, methane, major anions, metals, Cr(VI), Fe(II), total sulfides, dissolved H(2), Eh, pH, dissolved oxygen, specific conductance, alkalinity, and turbidity. Electrical conductivity profiles have been conducted using a Geoprobe to verify emplacement of the continuous wall as designed and to locate upgradient and downgradient wall interfaces for coring purposes. Coring has been conducted in November, 1996, in June and September, 1997, and March, 1998, to evaluate the rate of corrosion on the iron surfaces, precipitate buildup (particularly at the upgradient interface), and permeability changes due to wall emplacement. In addition to several continuous vertical cores, angled cores through the 0.6-m thick wall have been collected to capture upgradient and downgradient wall interfaces along approximate horizontal flow paths for mineralogic analyses. PMID- 10518669 TI - Melt pelletisation of a hygroscopic drug in a high shear mixer. Part 1. Influence of process variables. AB - The applicability of a high shear mixer for melt pelletisation of binary mixtures of sodium valproate and glycerol monostearate was investigated. The effects of binder concentration, impeller speed, jacket temperature and massing time on mean pellet size and size distribution were examined in a 2(4)-factorial design. Binder concentration and impeller speed were found to be the most important variables influencing the mean granule size and size distribution. An increase in each of those accelerated the granule growth. Due to the solubility of the drug in the molten binder very low amounts of binder were necessary for the formation of pellets. The modified high shear mixer was found to be suitable for batch sizes of 1-4 kg; granule growth was delayed with increasing load. A common pellet growth pattern, which can be divided into three phases, was derived and confirmed from all trials. The process was monitored by means of a torque measuring system. The torque-time curve can be used to detect the beginning of the destruction phase. PMID- 10518668 TI - Results of the reactant sand-fracking pilot test and implications for the in situ remediation of chlorinated VOCs and metals in deep and fractured bedrock aquifers. AB - Permeable reactive barriers (PRBs), such as the Waterloo Funnel and Gate System, first implemented at Canadian Forces Borden facility in 1992, are a passive remediation technology capable of controlling the migration of, and treating contaminated groundwater in situ. Most of the PRBs installed to date have been shallow installations created by backfilling sheet-pile shored excavations with iron filing reactive media. More recently continuous trenchers [R. Puls, Installation of permeable reactive barriers using continuous trenching equipment, Proceedings of the RTDF Permeable Barriers Work Group, Virginia Beach, VA, September 1997] and Caissons [J. Vogan, Caisson installation of a pilot scale, permeable reactive barrier in situ treatment zone at the Sommersworth Landfill, NH, Presented to the RTDF Permeable Barriers Work Group, Alexandria, VA, April 1996], and vertical fracturing emplacements [G. Hocking, Vertical hydraulic fracture emplacement of permeable reactive barriers, Progress Report delivered to the Permeable Reactive Barriers Workgroup of the Remedial Technology Development Forum, Beaverton, OR, April 1998] have been used to create reactive barriers in soil. None of the prior methods are capable of adequately addressing groundwater contamination in deep and fractured bedrock aquifers. The purpose of the RSF pilot study was to install reactive media into an impacted bedrock aquifer, and to evaluate the effectiveness of in situ treatment of chlorinated volatile organic compounds (CVOCs) and metals in that type of aquifer. Three discrete fractures were identified and treated and were subjected to testing before and after treatment. Between 300 and 1700 lb. of 1 mm diameter reactive proppants were injected into each zone to facilitate treatment. Monitoring data obtained from adjacent observation wells verified that fracking fluids reached at least 42 ft from the treatment well following hydrofracturing. The concentrations of many of the CVOCs decreased up to 98% based on the results of pre- and post-RSF treatment analyses. Consistent with other research, concentrations of CVOCs were noted to decrease including trichloroethene (TCE), tetrachloroethene (PCE), 1,1,1 trichloroethane (1,1,1-TCA), 1, 1-dichloroethane (1,1-DCA), and 1,1 dichloroethene (1,1-DCE) and increases were noted in concentrations of cis-1,2 dichloroethene (cis-1,2-DCE) and chloroform suggesting that the rate of transformation of the parent compounds to these daughter products is higher than the rate of destruction of the daughter products. The RSF pilot study demonstrated that: (1) zero valent iron foam proppants have the physical and chemical properties necessary to effectively treat CVOCs and metals in groundwater when inserted under high pressures into fractured bedrock. (2) Iron foam reactive media can be placed in bedrock using high pressure hydraulic fracturing equipment and polysaccharide viscosifiers. (3) The extent of the treatment can be monitored in situ using tracers and pressure transducers. (4) Well capacity is increased by improving hydraulic conductivity through hydraulic fracturing and proppant injection. The approximate cost of all of the effort expended in the pilot study was about US$200,000. Full-scale implementations are projected to cost between US$100,000 and US$1,000,000 and would depend on site specific conditions such as the extent and level of impacted groundwater requiring treatment. This technology can potentially be implemented to create treatment zones for the passive treatment of CVOC and metal impacted groundwater in fractured rock aquifers offering a cost-effective alternative to a pump and treat forever scenario. PMID- 10518670 TI - Distribution of salicylic acid in human stratum corneum following topical application in vivo: a comparison of six different formulations. AB - Distribution of salicylic acid in human stratum corneum from treatment of six different formulations was assessed by quantitation of drug content in sequentially tape-stripped stratum corneum after a single 2-h dose was applied unoccluded to skin on the ventral forearm of four female subjects. The profile and total amounts of stratum corneum removed in 20 tape-strips varied among different types of formulations. With or without normalization by the total stratum corneum weights removed, the extent of drug delivery to the stratum corneum decreased in the following order: SA (5%) > > SAC (10%), Duofilm (16.7%) > TSSS (2%) > SAO (10%), Salic (2.5%), the percentage in parentheses indicating the salicylic acid concentration in each formulation. The greatest topical bioavailability was observed for the alcoholic solution containing glycerol (SA). The 10% collodion formulation (SAC) was found to deliver an amount of salicylic acid into the stratum corneum 2-fold greater than 10% ointment formulation (SAO). Use of absorption ointment (TSSS) also increased the uptake of salicylic acid into the stratum corneum in comparison with formulations based on simple ointment (SAO) and oil in water (o/w) cream (Salic). The partitioning of salicylic acid from collodion formulations (SAC and Duofilm) appeared to be concentration independent. The results of this study indicate that topical bioavailability of salicylic acid in the stratum corneum varies substantially among different formulations. PMID- 10518671 TI - Pseudolatex preparation using a novel emulsion-diffusion process involving direct displacement of partially water-miscible solvents by distillation. AB - Pseudolatexes were obtained by a new process based on an emulsification-diffusion technique involving partially water-miscible solvents. The preparation method consisted of emulsifying an organic solution of polymer (saturated with water) in an aqueous solution of a stabilizing agent (saturated with solvent) using conventional stirrers, followed by direct solvent distillation. The technique relies on the rapid displacement of the solvent from the internal into the external phase which thereby provokes polymer aggregation. Nanoparticle formation is believed to occur because rapid solvent diffusion produces regions of local supersaturation near the interface, and nanoparticles are formed due to the ensuing interfacial phase transformations and polymer aggregation that occur in these interfacial domains. Using this method, it was possible to prepare pseudolatexes of biodegradable and non-biodegradable polymers such as poly(D,L lactic acid) and poly(epsilon-caprolactone), Eudragit E, cellulose acetate phthalate, cellulose acetate trimellitate using ethyl acetate or 2-butanone as partially water-miscible solvents and poly(vinyl alcohol) or poloxamer 407 as stabilizing agent. A transition from nano- to microparticles was observed at high polymer concentrations. At concentrations above 30% w/v of Eudragit E in ethyl acetate or cellulose acetate phthalate in 2-butanone only microparticles were obtained. This behaviour was attributed to decreased transport of polymer molecules into the aqueous phase. PMID- 10518672 TI - Stabilization of aerosolized IFN-gamma by liposomes. AB - Aerosolized IFN-gamma is very unstable. We have improved the stability of IFN gamma in the jet nebulizer by adding small liposomes. Aerosolized IFN-gamma was recovered in PBS solution by bubbling and its concentration was determined. After nebulization for 30 min, aerosolized IFN-gamma was detected only 0.4+/-0.2% of the initial amount in the PBS solution and 3.1+/-0.7% in the reservoir. On the other hand, the addition of small liposomes (HSPC/DSPG=10/1 (molar ratio), 45+/ 24 nm) in the nebulizer increased the stability of IFN-gamma, 27.2+/-4.7% of the initial amount in the PBS solution and 25.7+/-12.6% in the reservoir. The present study also examined the effects of composition and concentration of liposomes on the stabilization of aerosolized IFN-gamma. Liposome prepared from distearoyl phosphatidylcholine (DSPC) or hydrogenated soy phosphatidylcholine (HSPC) was very effective for stabilization of aerosolized IFN-gamma (DSPC/DPPG=10/1, HSPC/DSPG=10/1). HSPC/DSPG liposome was efficient at the concentration higher than 12.5 micromols/ml for the stabilization of 5x10(5) JRU/ml of IFN-gamma. In considering the mechanism of this stabilization, the results of gel filtration chromatography suggest that IFN-gamma is inactivated by polymerization or aggregation in nebulization, while the inactivation is suppressed by liposomes due to their adsorption to IFN-gamma. PMID- 10518673 TI - Model development for O(2) and N(2) permeation rates through CZ-resin vials. AB - The headspace of vials containing oxygen-sensitive formulations is filled with a nitrogen blanket. This paper presents the development of a mathematical model to predict the oxygen and nitrogen permeation rates through the walls of plastic CZ resin vials. The model estimates the time required for a nitrogen-filled vial to reach ambient nitrogen and oxygen levels. The permeation of oxygen and nitrogen through the vial is governed by Fick's law and may be described by an exponential equation. Using the values for oxygen and nitrogen permeation through CZ-resin vial, the half-lives for the decrease in nitrogen level and increase in oxygen level was found to be 150 days and 15 days, respectively. This result can be attributed to the greater permeability of CZ-resin vial to oxygen (79.06 cm(3) mm/m(2)-24 h-atm) when compared with nitrogen (12 cm(3)-mm/m(2)-24 h-atm). The ingress of oxygen into CZ-resin vials was determined experimentally and it was found to verify the model. These results indicate that CZ-resin vials may be inappropriate for packaging oxygen-sensitive formulations even in the presence of a nitrogen-filled headspace. PMID- 10518675 TI - Encapsulation characteristics of nystatin in liposomes: effects of cholesterol and polyethylene glycol derivatives. AB - In this study, we characterized the encapsulation of amphipathic nystatin into liposomes with or without cholesterol (CH) and a polyethylene glycol derivative, distearoyl-N-(monomethoxy poly(ethylene glycol)succinyl)phosphatidylethanolamine (DSPE-PEG). The highest encapsulation efficacy of nystatin into liposomes (151 microg nystatin/mg lipid) was obtained with a cholesterol-free lipid composition containing 6 mol% of DSPE-PEG. The encapsulation efficacy was decreased by the incorporation of CH and improved by the incorporation of DSPE-PEG. In liposomes composed of dipalmitoylphosphatidylcholine (DPPC)/CH (2:1, mol/mol), the highest encapsulation efficacy of nystatin liposomes (84 microg/mg lipid) was achieved by the addition of DSPE-PEG and hydration with 9% sucrose solution, as compared with 13 microg/mg lipid without DSPE-PEG. The encapsulated amount increased with increasing amount of DSPE-PEG used and plateaued at 6 mol% of DSPE-PEG. The optimum molecular weight of PEG in DSPE-PEG was 2000 and a larger molecular weight resulted in lower encapsulation. The incorporation of CH affected the self association of nystatin with lipid membranes, which was detected by fluorescence measurement. The molecular interaction between an amino group in nystatin and a phosphate group in DSPE-PEG plays an important role in efficient encapsulation of nystatin. Finally, the encapsulation characteristics of nystatin were compared with those of amphotericin B (AmB). Nystatin more readily associated with CH-free lipid membranes, but, AmB more readily interacted with DSPE-PEG. The results indicated that the differences in the molecular association of AmB or nystatin with lipids or DSPE-PEG are reflected in the encapsulation characteristics in liposomes. PMID- 10518674 TI - Rheological characterization of microcrystalline cellulose and silicified microcrystalline cellulose wet masses using a mixer torque rheometer. AB - The rheological properties of silicified microcrystalline cellulose (Prosolv 50) were compared with those of standard grades of microcrystalline cellulose (Emcocel 50 and Avicel PH 101). Cellulose samples were analyzed using nitrogen adsorption together with particle size, flowability, density and swelling volume studies. The rheological behaviour of the wet powder masses was studied as a function of mixing time using a mixer torque rheometer (MTR). Silicified microcrystalline cellulose exhibited improved flow characteristics and increased specific surface area compared to standard microcrystalline cellulose grades. Although the silicification process affected the swelling properties and, furthermore, the mixing kinetics of microcrystalline cellulose, the source of the microcrystalline cellulose had a stronger influence than silicification on the liquid requirement at peak torque. PMID- 10518676 TI - Surface free energy of ethylcellulose films and the influence of plasticizers. AB - The surface free energy parameters of ethylcellulose (EC) films were determined using the Lifshitz-van der Waals/acid-base approach and the influence of plasticizers on their surface energetics was assessed. Films were prepared by dip coating glass slides in organic solvents containing EC and the advancing angles of drops of pure liquids on the EC films were measured with a contact angle goniometer using the captive drop technique. EC has lower surface free energy than cellulose. The acid-base (AB) term made only a slight contribution to the total surface free energy and the surfaces exhibited predominantly monopolar electron-donicity. The addition of plasticizer (dibutyl sebacate or dibutyl phthalate) resulted in a small decrease in the total surface free energy. The effects of film forming variables, including solvent system, concentration and post-formation treatment (annealing), on the surface free energy parameters of EC films were also investigated. These data were then used to analyze how the surface energetics affect the interaction of the EC films with other surfaces based on interfacial tension, work of adhesion and spreading coefficient calculations. Lifshitz-van der Waals (LW) interactions provided the major contribution to the work of adhesion for EC with all of the solid substrates analyzed. However, the AB interactions contributed significantly to the work of adhesion for EC with 'bipolar' substrates and to the spreading coefficients of EC over substrates. The consideration of work of adhesion and spreading coefficient based on surface free energy parameters may have potential use in evaluating factors affecting film adhesion and, furthermore, in optimizing pharmaceutical film coating processes. PMID- 10518677 TI - Interpolymer complexation. I. Preparation and characterization of a polyvinyl acetate phthalate-polyvinylpyrrolidone (PVAP-PVP) complex. AB - Polyvinyl acetate phthalate (PVAP) and polyvinylpyrrolidone (PVP) readily reacted in ethanol and acidic aqueous solutions to produce an insoluble PVAP-PVP complex. The complex has a pK(a) of 3.8. It is practically insoluble in common organic solvents, but dissolves in dimethylsulfoxide, an alkali or ammonical solution, and a 4:1 (v/v) mixture of methylene chloride and methanol. The powder X-ray diffraction analysis revealed the complex to be an amorphous material. The Fourier-transform infrared spectrum of the complex exhibited characteristics carbonyl stretching vibrations at 1724 and 1657 cm(-1) due to phthalate and acetate moieties in PVAP and cyclic amide groups in PVP, respectively, and at 1632 cm(-1) (appeared as a shoulder) due to cyclic amide groups of PVP bound to PVAP. The proton and carbon-13 (solution and solid-state) nuclear magnetic resonance spectra of the complex showed peak profiles that were linear combinations of those of PVAP and PVP. No new peaks appeared and no change in chemical shifts was observed due to complexation. The spectral data suggest that the interaction between PVAP and PVP probably initially involves the formation of hydrogen bonds between carbonyl groups of PVP and carboxylic groups of PVAP at some point of the polymer chains, causing the hydrophilic parts of the two flexible polymer chains strongly hydrophobic. As a result, the two polymer chains coil up into a compact structure and, consequently, precipitate out from the solution as an insoluble complex. PMID- 10518678 TI - On the stability of ascorbic acid in emulsified systems for topical and cosmetic use. AB - Several O/W microemulsions, O/W and W/O emulsions and a W/O/W multiple emulsion were prepared using non-ionic, non-ethoxylated, skin compatible emulsifiers. Ascorbic acid (vitamin C) was added to the emulsified systems and its stability against oxidation was studied at 45.0 degrees C in aerobic conditions and compared with that in aqueous solutions at different pH values. All emulsified systems provided protection to ascorbic acid, as its degradation rate, which increased with increasing pH, was slower in emulsified systems than in aqueous solutions. The highest protection of ascorbic acid was when it was dissolved in the inner aqueous phase of the W/O/W multiple emulsion, both at 45 and at 20 degrees C for long storage. A pseudo first-order mechanism was hypothesised for ascorbic acid degradation in the experimental conditions for as long as abundant dissolved oxygen was present. PMID- 10518679 TI - Spray-drying of trypsin - surface characterisation and activity preservation. AB - In the present study trypsin mixed with various carbohydrates, i.e. lactose, sucrose, mannitol, alpha-cyclodextrin and dextrin, was spray-dried in order to investigate the effects of spray-drying on this enzyme, with particular emphasis on the effects of interactions between trypsin and the surface formed during spray-drying. The protein was strongly over-represented at the surface of the powder particles, the surface coverage ranging from 10 to 65%, depending on the amount of trypsin in the solids (0.2-5%). This indicates that the protein adsorbs at the air/liquid interface of the spray-droplets, and that this surface is also largely preserved after drying. The surface concentration of protein in the spray dried powders could be controlled by adding a surfactant to the mixture before drying, since the surfactant adsorbs preferentially at the air/liquid interface of the spray droplets, thus expelling protein from the surface. In general, the residual activity of trypsin in these non-optimised formulations was 90% or higher, and in no case less than 82%. It was found that the loss of activity could partly be explained by inactivation of the protein adsorbed at the surface. For mannitol and sucrose, however, the level of inactivation was higher than could be explained by surface inactivation alone, and additional mechanisms must also be considered. PMID- 10518680 TI - A therapeutic dose of primaquine can be delivered across excised human skin from simple transdermal patches. AB - This work investigated the permeation of primaquine across full-thickness excised human skin from two acrylate transdermal adhesives. Primaquine base was formulated with National Starch 387-2516 and 387-2287 to provide aluminium foil backed 1-cm diameter patches, each loaded with 10 mg drug. Other patches were prepared that included Migliol 840 as a potential penetration enhancer. The patches were applied to cadaver skin in Franz-type diffusion cells and the permeation of primaquine determined over a 24-h period. Relatively high fluxes were found, the highest being from those formulations lacking the Migliol 840: 5.68+/-0.30x10(-2) mg cm(-2) h(-1) from 387-2516; 4.94+/-0.20x10(-2) mg cm(-2) h( 1) from 387-2287. It was determined that a simple patch with a diameter of approximately 13 cm(2) could deliver a therapeutic in vivo dose, with possibilities for the treatment and prophylaxis of Plasmodium vivax, P. ovale and P. falciparum forms of malaria. The presence of Migliol 840 failed to produce the anticipated enhancing effect: flux rates that were approximately halved. These results could to a certain extent be rationalised in terms of thermodynamic activity. PMID- 10518681 TI - Influence of the process parameters on the characteristics of starch based hot stage extrudates. AB - The influence of the process parameters on the characteristics of matrix formulations produced by means of hot stage extrusion was investigated using three experimental designs. The first one was designed to evaluate the importance of the screw speed (150-450 rpm) and the feed rate (2-12 kg/h), while the second and the third were designed to study the importance of the temperature profile (60-120 degrees C). The extrudates were produced with a laboratory twin screw extruder equipped with a 3-mm cylindrical die. The formulations consisted of 53% corn starch, 15% sorbitol, 2% glyceryl monostearate and 30% theophylline monohydrate as the model drug. The extrudates were characterized by Karl-Fischer titration, Hg-porosimetry, four-point bending and dissolution testing. From the first design it was concluded that the screw speed and feed rate hardly affected the water content of the extrudates, but that there was a clear influence on the extrudate radius, porosity, mechanical strength and dissolution behaviour. High screw speed-high feed rate processes in comparison with low screw speed-low feed rate processes caused an increase in extrudate radius and porosity and a decrease in mechanical strength and drug release rate from the matrix. It was clear that the contribution of the feed rate was higher than that of the screw speed. Expansion, promoted under certain extrusion conditions, could explain the obtained results. The second and third design revealed that only the maximum barrel temperature and not the whole temperature profile was responsible for the temperature effects on the extrudate characteristics. It was concluded that the maximum barrel temperature was the most critical parameter of the hot stage extrusion process. PMID- 10518682 TI - Measuring water distribution in extrudates using magnetic resonance imaging (MRI). AB - Magnetic resonance imaging (MRI) has been applied to the evaluation of the distribution of water in extrudates produced by extruding pastes. Two model drugs similar in chemical structure were mixed with microcrystalline cellulose (MCC) and with two different amounts of water and extruded at two different extrusion speeds using a ram extruder. Extrudates were collected during the steady-state stage of the extrusion profile and were analysed for the water distribution using MRI. The percolation threshold for each sample was calculated to evaluate the degree of water structure within the sample. The water distribution inside the extrudates was surprisingly uniform. The extrudates produced using the faster extrusion speed had a significant lower percolation threshold, which suggests the existence of a greater water structure in the extrudates. A significant correlation was found between the percolation threshold and the extrusion force, which had been used to provide the extrudates. PMID- 10518683 TI - Preparation and characterization of cationic microspheres for gene delivery. AB - The production and characterization of cationic microparticles based on Eudragit RS and cationic agents (i.e. a cationic acrylic polymer and three different cationic surfactants) for the delivery of nucleic acids is here described. It was found that morphological and dimensional characteristics of microparticles were influenced by the type and concentration of cationic agent employed and by some experimental parameters such as stirring speed, emulsifying agent and type of rotor. The desoxiribonucleotide Defibrotide (DFT) was associated with positively charged microparticles and its in vitro release kinetics from microparticles were determined. A study of the in vitro toxicity of cationic microparticles on cultured human cell line K562 was also performed, demonstrating that DDAB(18) microparticles display very low cytotoxicity. PMID- 10518684 TI - Lysosubtilin modification, Fermosorb, designed for polymeric carrier-mediated intestinal delivery of lytic enzymes: pilot-scale preparation and evaluation of this veterinary medicinal product. AB - Antimicrobial enzymotherapy/enzymoprophylaxis has potential for use as a measure to overcome problems associated with resistance to commonly applied antibiotics. Lysosubtilin, an authorized veterinary medicinal product, when used per os for the treatment and prophylaxis of intestinal infections in newborn calves, is not always efficient due to partial inactivation of lytic enzymes in the gastric region. In this contribution a simple technology for preparation of pH-dependent reversibly dissociating acid stable enzyme-polymer complex (two-component oral delayed-release lysosubtilin formulation, Fermosorb) designed for intestinal delivery of lytic enzymes is described. The technology is based on immobilization of lytic enzymes, using 1% lysosubtilin solution in 10 mM acetate buffer of pH 5.0, onto commercial highly porous carboxylic cation exchanger Biocarb L (v/w ratio 10:1, process duration 1 h) with after-following procedures of vacuum filtration, oven-drying and standardization of the enzyme-polymer complex formed. The technology process of pilot-scale Fermosorb fabrication on the whole revealed itself as simply employed and highly repeatable, totalling in the final lytic enzyme activity yield of 40.2% (the average value obtained from the analysis of the 11 batches running) and approximately 4000 (3938) kg of Fermosorb (200 batches) produced. The proposed technological approach can be successfully applied for fabrication of other enzyme preparations as well and this was shown in the example of Polyferm, a preparation with both lytic and proteolytic enzyme activities. In vitro evaluation of Fermosorb revealed it was more stable when exposed to the acidic environment as well as in storage when compared with the native lysosubtilin. No negative change in the antimicrobial spectrum of action of Fermosorb versus lysosubtilin, influenced by immobilization of lytic enzymes onto Biocarb L, was observed. Moreover, all six lysosubtilin-resistant microbial strains tested have been found to be Fermosorb-susceptible. In vivo evaluation studies performed on 1200 newborn calves revealed 95.2% therapeutic as well as 95.0% prophylactic efficacy of Fermosorb in respect to colibacillosis versus 74.0 and 80.0% for lysosubtilin, respectively, the differences being statistically significant (P<0.01). As a consequence of these studies Fermosorb was authorized for use throughout the former Soviet Union. Data collected during postmarketing surveillance of Fermosorb, which was applied for more than 163,000 newborn calves, confirmed high efficacy (92.3 and 95.5% for treatment and prophylaxis, respectively) and safety of this veterinary medicinal product. PMID- 10518686 TI - Simultaneous determination of chlordiazepoxide and clidinium bromide in pharmaceutical formulations by derivative spectrophotometry. AB - A direct and simple first derivative spectrophotometric method has been developed for the simultaneous determination of clidinium bromide and chlordiazepoxide in pharmaceutical formulations. Acetonitrile was used as solvent for extracting the drugs from the formulations and subsequently the samples were evaluated directly by derivative spectrophotometry. Simultaneous determination of the drugs can be carried out using the zero-crossing method for clidinium bromide at 220.8 nm and the graphical method for chlordiazepoxide at 283.6 nm. The calibration graphs were linear in the ranges from 0.983 to 21.62 mg/l of clidinium bromide and from 0. 740 to 12.0 mg/l of chlordiazepoxide. The ingredients commonly found in commercial pharmaceutical formulations do not interfere. The proposed method was applied to the determination of these drugs in tablets. PMID- 10518685 TI - Apparatus for studying in vitro drug release from medicated chewing gums. AB - An apparatus for in vitro drug release testing of medicated chewing gums has been developed and is described in detail. The effects on the drug release when varying critical instrumental settings such as the chewing stroke frequency, the distance between the chewing surfaces, the twisting movements of these surfaces and the temperature of the test medium have been thoroughly investigated. It has been shown that the drug release can be tuned to obtain suitable drug release profiles for a number of products: Nicorette((R)) and Nicotinell((R)) (active substance nicotine), Travvell((R)) (dimenhydrinate), V6((R)) (xylitol) and an experimental formulation containing meclizine. The main usage of the present apparatus should be within quality control but the present study has also shown that it may be employed within development pharmaceutics since useful in vivo/in vitro relationships may be obtained due to the versatile settings of the critical instrumental parameters. PMID- 10518687 TI - Immunological responses to nasal delivery of free and encapsulated tetanus toxoid: studies on the effect of vehicle volume. AB - In light of growing interest in the intranasal route as a non-invasive mode of immunisation, we have investigated the relationship between the volume of liquid instilled into the nasal passages and the development of subsequent immunological responses. Groups of six mice were intranasally immunised with soluble or microsphere encapsulated tetanus toxoid on days 1, 14 and 28 of the experiment. Microsphere suspensions and tetanus toxoid solutions were nasally instilled in two different volumes of buffer (10 or 50 microl). Nasal instillation of microspheres in 10 microl of buffer generated statistically depressed (P<0.001) tertiary serum anti-toxoid IgG responses in comparison to animals immunised with 10 or 50 microl of soluble vaccine, or 50 microl of microsphere suspension. Relative to other treatments, nasal inoculation of encapsulated toxoid suspended in 50 microl generated statistically (P<0.05) superior levels of specific IgG and IgA antibodies in day 49 lung wash samples. When radiolabelled microspheres were nasally instilled into mouse nares in 50-microl volumes of buffer, a significant portion of the dose (48%) entered the lungs (P<0.001), whereas more particles remained in the nasal passages when a smaller (10 microl) volume of suspension was given (P<0.001). These biodistribution and immunological data indicate that to generate optimal bronchopulmonary and systemic responses in concert following nasal administration, microparticulated vaccines should be administered with a delivery device that targets the formulation to distal regions of the nasal passages and the lower respiratory tract. PMID- 10518688 TI - Comparative mucoretention of sucralfate suspensions in an everted rat esophagus model. AB - A simple, rapid, and reproducible in vitro model was established to quantify the relative esophageal mucoadhesive properties of viscous liquid formulations, and the model was applied to compare marketed sucralfate suspensions (Gastrogel, Antepsin, and Ulcogant) to better understand differences in clinical performance. Rat esophageal mucosal segments were everted onto a glass rod and briefly immersed into a liquid formulation containing 51Cr microspheres. Indirect quantification of the retained formulation provided excellent recovery (98.7 101%) and reasonable precision (1.06-38.3% CV). Mucosal retention profiles of the formulations were determined by rinsing the coated tissue in relevant gastrointestinal fluids using the technique of reciprocating vertical immersion. Dispersions of the mucoadhesive hydrogel Carbopol 934P were employed to initially characterize the performance of the model with respect to composition of the rinse fluids, and type and amount of shear force during rinsing. Retention of Carbopol was sensitive to the mechanics of rinsing and to salivary salts but not mucin in the rinse medium. A sucralfate gel suspension (Gastrogel) showed much greater mucoadhesion and resistance to removal by saliva than two non-gel suspensions (Antepsin, Ulcogant). Results suggest that in situ gelation may be a contributing mechanism for strong esophageal retention. These in vitro results are in general agreement with published human esophageal retention data on similar sucralfate suspensions and lend credence to the everted rat esophagus as a qualitatively predictive in vitro model for development of esophageal mucoadhesive liquids. PMID- 10518689 TI - Rheological characterization of pharmaceutical powders using tap testing, shear cell and mercury porosimeter. AB - Most of the pharmaceutical processes involved in the manufacturing of so lid dosage forms are connected with powder flow properties, at least for some of the intermediate steps. Powder flow characteristics are commonl y investigated by various measurements, such as handling angles, tap tes ting, shear cell measurements, etc. All these approaches allow the calc ulation of indices characterising powder flowability. Unfortunately, th ese methodologies are highly product consuming, which is a limitation in the first steps of a novel drug development, when only a small amount of product is available. The use of mercury porosimetry to evaluate compre ssibility and flow properties of powders could be a new and alternative approach to obtain insight in the rheological properties of granular med ium by the interpretation of the first part of programs (low pressures) . We have developed such an evaluation and compared the results obtaine d with those given by tap testing and shear cell measurements, applied t o four excipients for direct tabletting and three different drugs. Merc ury porosimetry turned out to be a sensitive technique, able to providequantitative information about powder flow properties, complemen ted by an evaluation of particles micro porosity and size distribution, in a single step. These characterisations are obtained with only approx imately 250mg of bulk powder compared to high quantities ( >100g) needed for other methods. PMID- 10518690 TI - Study of Verapamil hydrochloride release from compressed hydrophilic Polyox-Wsr tablets. AB - This study deals with Verapamil hydrochloride release from tablets based on high molecular weight poly(ethylene oxide) (PEO). The drug release proceeds as a controlled diffusion (n=0.44-0.47), which rate is dependent on the molecular weight of PEO. Independent from it, under the conditions of the Half-change test, the drug release practically ceases after 4 h as a result of obtaining low soluble in the water base. The introduction of hydrophilic polymers with pH dependent solubility (Eudragit L, Eudispert hv and Carbopol 934) at concentrations of 10/50% with respect to PEO amount keeping constant the ratio drug: matrix insures relatively complete release both in alkali medium and under the conditions of the Half-change test. Meanwhile drug release kinetics also changes - the release of all models studied runs as a typical abnormal diffusion (a=0.66-0.87), i. e. like a diffusion-relaxation controlled process. The decrease in drug concentration leads not only to retarded release of the drug sample but also to changes in the kinetics of the process. At lower drug concentrations on the matrix from a typical abnormal diffusion it turns into a relaxation controlled diffusion (n(10%)=1). PMID- 10518691 TI - Characterization and evaluation of isomalt performance in direct compression. AB - Isomalt is a sugar substitute with a wide range of potential pharmaceutical applications as a result of its physicochemical properties. Four grades of this material were evaluated for their physical characteristics. Only Palatinit(R) C and F exhibited potential characteristics for direct compression. As expected, the products required lubrification for tabletting. A level of 1% lubricant gave the best performance for Palatinit(R) C, the most compressible grade as shown by compaction profiles generated using a single-punch machine. However, its flow behaviour had to be improved by including 0.5% Aerosil(R) 200 as shown by tablet weight uniformity data. Further evaluation by Heckel analysis showed that isomalt exhibited plastic behaviour and underwent elastic recovery primary in the die. Its dilution potential was examined using powdered paracetamol. Acceptable tablets were produced up to 30% drug dilution, but the tensile strength values were reduced, disintegration time and friability increased as expected. Drug dissolution profiles showed a decreasing dissolution rate with the increase of compression force and drug concentration, but considerable improvement was noted when a disintegrant was included. The physical characteristics of the tablets were relatively stable after half a year storage at different humidities as a result of the low hygroscopicity of isomalt. PMID- 10518692 TI - Slow relaxation of the sub-main transition in multilamellar phosphatidylcholine vesicles. AB - The influence of ionic strength and equilibration time on the appearance of the sub-main transition in fully hydrated multilamellar vesicles composed of phosphatidylcholines has been investigated by means of calorimetry and densitometry. The heat capacity measurements show that the transition enthalpy of the sub-main transition is affected by both salt concentration (KCl) and equilibration time. The small heat capacity peak appearing in vesicles made in pure water is significantly increased upon addition of salt. Furthermore, equilibration of the multilamellar vesicles at low temperatures for several weeks results in a pronounced enhancement of the transition enthalpy of the sub-main transition. Neither salt concentration nor equilibration time affected the transition temperature of the sub-main transition. In the densitometry measurements a small volume change is detectable for high salt concentrations. In order to gain further insight into the physical mechanisms involved in the sub main transition, a Monte Carlo computer simulation study has been carried out using a microscopic model. The combined experimental and simulation results suggest that the sub-main transition involves an acyl chain disordering of phospholipids in lipid bilayer regions that are characterized by a locally decreased lateral pressure most likely caused by a curvature stress. PMID- 10518693 TI - Ion currents of Xenopus laevis oocytes: state of the art. PMID- 10518694 TI - The membrane-bound basic carboxypeptidase from hog intestinal mucosa(1). AB - The carboxypeptidase activity occurring in hog intestinal mucosa is apparently due to two distinct enzymes which may be responsible for the release of basic COOH-terminal amino acids from short peptides. The plasma membrane-bound carboxypeptidase activity which occurs at neutral optimum pH levels was found to be enhanced by CoCl(2) and inhibited by guanidinoethylmercaptosuccinic acid, o phenanthroline, ethylenediamine tetraacetic acid and cadmium acetate; whereas the soluble carboxypeptidase activity which occurs at an optimum pH level of 5.0 was not activated by CoCl(2) and only slightly inhibited by o-phenanthroline, ethylenediamine tetraacetic acid, NiCl(2) and CdCl(2). The latter activity was presumably due to lysosomal cathepsin B, which is known to be present in the soluble fraction of hog intestinal mucosa. Although the membrane-bound enzyme was evenly distributed along the small intestine, it was not anchored in the phospholipidic bilayer via a glycosyl-phosphatidylinositol moiety, as carboxypeptidase M from human placenta is. The enzyme was not solubilized by phosphatidylinositol-specific phospholipase C, but was solubilized to practically the same extent by several detergents. The purified trypsin-solubilized form is a glycoprotein with a molecular mass of 200 kDa, as determined by performing SDS PAGE and gel filtration, which differs considerably from the molecular mass of human placental carboxypeptidase M (62 kDa). It was found to cleave lysyl bonds more rapidly than arginyl bonds, which is not so in the case of carboxypeptidase M, and immunoblotting analysis provided further evidence that hog intestinal and human placental membrane-bound carboxypeptidases do not bear much resemblance to each other. Since the latter enzyme has been called carboxypeptidase M, it is suggested that the former might be carboxypeptidase D, the recently described new member of the carboxypeptide B-type family. PMID- 10518695 TI - Gadolinium induces domain and pore formation of human erythrocyte membrane: an atomic force microscopic study. AB - Lanthanide cations bind to human erythrocyte membranes and enhance cell permeability. It was postulated that this effect is due to their likeness with calcium ions, which have been used to induce perforation of cells. However, the nature and mechanism of the perforation are still not clear. In the present work, the change in surface topography of erythrocyte membranes exposed to various gadolinium species was imaged with an atomic force microscope (AFM) in order to get direct evidence of perforation. The images of the whole cell and regions in nanometer scale showed that the normal surface is featured by closely packed nanometer size particles. The AFM images showed that Gd(3+) binding to erythrocytes led to domain structure at low concentration and pore formation at higher concentration. The domain structures that appeared after incubation with 1.0x10(-6)-1.0x10(-5) mol/l Gd(3+) solution for 30 min are featured by the particles aggregated to form ranges and the separations among them enlarged to gorges. With a higher concentration, 2.5x10(-5) mol/l Gd(3+), the further aggregation developed into crater-shaped 'pores'. By washing with EDTA the 'pores' can be resealed but the domain structure remained. The anionic complex of Gd(3+), [Gd(Cit)(2)](3-) of this concentration, can only induce the domain structure formation. The domain and 'pore' structures mediated by Gd(3+) concentrations might be responsible for both enhanced permeability and perforation. The mechanism of Gd-induced domain formation and perforation is discussed on the basis of aggregation of membrane proteins and the coexistence of different phases of membrane lipids resulting from Gd(3+) binding. PMID- 10518696 TI - Thermodynamics of alcohol-lipid bilayer interactions: application of a binding model. AB - Several recent reports have provided evidence that interactions of small alcohols with lipid bilayer membranes are dominated by adsorption to the membrane-water interface. This mode of interaction is better modeled by binding models than solution theories. In the present study, alcohol-membrane interactions are examined by applying the 'solvent exchange model' [J.A. Schellmann, Biophys. Chem. 37 (1990) 121] to calorimetric measurements. Binding constants (in mole fraction units) for small alcohols to unilamellar liposomes of dimyristoyl phosphatidylcholine were found to be close to unity, and in contrast to partitioning coefficients they decrease through the sequence ethanol, 1-propanol, 1-butanol. Thus, the direct (intrinsic) affinity of the bilayer for these alcohols is lower the longer the acyl chain. A distinction between binding and partitioning is discussed, and it is demonstrated that a high concentration of solute in the bilayer (large partitioning coefficients) can be obtained even in cases of weak binding. Other results from the model suggest that the number of binding sites on the lipid bilayer interface is 1-3 times the number of lipid molecules and that the binding is endothermic with an enthalpy change of 10-15 kJ/mol. Close to the main phase transition of the lipid bilayer the results suggest the presence of two distinct classes of binding sites: 'normal' sites similar to those observed at higher temperatures, and a lower number of high affinity sites with binding constants larger by one or two orders of magnitude. The occurrence of high-affinity sites is discussed with respect to fluctuating gel and fluid domains in bilayer membranes close to the main phase transition. PMID- 10518697 TI - Polyethylene glycol enhances lipoplex-cell association and lipofection. AB - The association between liposome-DNA complexes (lipoplexes) and targeted cell membranes is a limiting step of cationic liposome-mediated transfection. A novel technique was developed where lipoplex-cell membrane association is enhanced by the addition of 2-6% polyethylene glycol (PEG) to the transfection media. Lipoplex-cell association was found to increase up to 100 times in the presence of PEG. Transfection increased correspondingly in the presence of PEG. This increase was found in several cell lines. These results show that lipoplex adsorption to cell membranes is a critical step in liposome-mediated transfection. This step can be facilitated by PEG-induced particle aggregation. PMID- 10518698 TI - Targeting lymph nodes with liposomes bearing anti-HLA-DR Fab' fragments. AB - The ability of liposomes bearing anti-HLA-DR Fab' fragments to target cells expressing the human HLA-DR determinant of the major histocompatibility complex class II (MHC-II) has been evaluated and compared to that of conventional liposomes. Anti-HLA-DR immunoliposomes did not bind to HLA-DR-negative cells. In contrast, a high level of binding was observed following incubation of immunoliposomes with cells bearing important levels of human HLA-DR. The accumulation of conventional and murine anti-HLA-DR immunoliposomes in different tissues has been investigated following a single subcutaneous injection given in the upper back of C3H mice. Anti-HLA-DR immunoliposomes resulted in a much better accumulation in the cervical and brachial lymph nodes when compared to conventional liposomes. The accumulation in the liver was similar for both liposomal preparations, whereas an approximately twofold decrease in accumulation was observed for immunoliposomes in the spleen. Given that HLA-DR surface marker is expressed on monocyte/macrophages and activated CD4+ T lymphocytes, the primary cellular reservoirs of the human immunodeficiency virus (HIV), the use of liposomes bearing surface-attached anti-HLA-DR could constitute a convenient strategy to more efficiently treat this debilitating retroviral disease. Moreover, the reported incorporation of high amounts of host-encoded HLA-DR proteins by HIV particles renders the use of liposomes bearing anti-HLA-DR antibodies even more attractive. PMID- 10518699 TI - The transmembrane protein bacterioopsin affects the polarity of the hydrophobic core of the host lipid bilayer. AB - Influence of the transmembrane protein bacterioopsin (the retinal-free form of bacteriorhodopsin) on the polarity of egg-phosphatidylcholine bilayers was studied by means of a steady-state and time-resolved fluorescence approach exploiting the solvatochromic properties of the 2-anthroyl fluorophore. Introduced in phosphatidylcholine molecules in the form of 8-(2-anthroyl)octanoic acid, this fluorophore probed the hydrocarbon core of the lipid bilayer. As previously shown (E. Perochon et al., Biochemistry 31 (1992) 7672-7682), water molecules were detected in this region of the terminal part of the lipid acyl chains. Their number was considerably reduced upon addition of bacterioopsin to the lipids. This was assessed by a blue shift in the fluorescence emission spectra of the probe and a marked decrease in the fractional population of fluorophores interacting with water, to the benefit of those experiencing a hydrophobic environment. In agreement with current theories, this decrease in the hydration of the bilayer may be linked to an increase in the acyl chain order and a decrease in the lateral diffusion coefficient of lipids near the protein. The data obtained at high protein concentration accounts for a protein/lipid interface which is much less hydrated than the hydrophobic core of a protein-free lipid bilayer. PMID- 10518700 TI - Membrane effects of nitrite-induced oxidation of human red blood cells. AB - The aim of our investigation was to study the red blood cell (RBC) membrane effects of NaNO(2)-induced oxidative stress. Hyperpolarization of erythrocyte membranes and an increase in membrane rigidity have been shown as a result of RBC oxidation by sodium nitrite. These membrane changes preceded reduced glutathione depletion and were observed simultaneously with methemoglobin (metHb) formation. Changes of the glutathione pool (total and reduced glutathione, and mixed protein glutathione disulfides) during nitrite-induced erythrocyte oxidation have been demonstrated. The rates of intracellular oxyhemoglobin and GSH oxidation highly increased as pH decreased in the range of 7.5-6.5. The activation energy of intracellular metHb formation obtained from the temperature dependence of the rate of HbO(2) oxidation in RBC was equal to 16.7+/-1.6 kJ/mol in comparison with 12.8+/-1.5 kJ/mol calculated for metHb formation in hemolysates. It was found that anion exchange protein (band 3 protein) of the erythrocyte membrane does not participate significantly in the transport of nitrite ions into the erythrocytes as band 3 inhibitors (DIDS, SITS) did not decrease the intracellular HbO(2) oxidation by extracellular nitrite. PMID- 10518701 TI - Fluid phase endocytosis and galactosyl receptor-mediated endocytosis employ different early endosomes. AB - Endocytosis may originate both in coated pits and in uncoated regions of the plasma membrane. In hepatocytes it has been shown that fluid phase endocytosis (here defined as 'pinocytosis') is unaffected by treatments that arrest coated pit-mediated endocytosis, indicating that pinocytosis is primarily a clathrin independent process. In this study we have tried to determine possible connections between pinocytosis and clathrin-dependent endocytosis in rat hepatocytes by means of subcellular fractionation, electron microscopy, and by assessing the influence of inhibitors of clathrin-dependent endocytosis on pinocytosis. As marker for clathrin-dependent endocytosis was used asialoorosomucoid (AOM) labelled with [(125)I]tyramine cellobiose ([(125)I]TC). [(125)I]TC-labelled bovine serum albumin ([(125)I]TC-BSA) was found to be a useful marker for pinocytosis. Its uptake in the cells is not saturable, and any remnants of [(125)I]TC-BSA associated with the cell surface could be removed by incubating the cells with 0.3% pronase at 0 degrees C for 60 min. The data obtained by electron microscopy and by subcellular fractionation suggested that early after initiation of uptake (<15 min) [(125)I]TC-BSA and [(125)I]TC-AOM were present in different endocytic vesicles. The two probes probably join prior to their entrance in the lysosomal compartment. The relation between endocytosis via coated pits and pinocytosis was also studied with techniques that induced a selective density shift either in the clathrin-dependent pathway (by AOM-HRP) or in the pinocytic pathway (by allowing uptake of AuBSA). Both treatments indicated that the two probes ([(125)I]TC-AOM and [(125)I]TC-BSA) were early after uptake, at least partly, in separate endocytic compartments. The different distribution of the fluid phase marker and the ligand (internalised via coated pits) was not due to a difference in the rate at which they enter a later compartment, since a lowering of the incubation temperature to 18 degrees C, which should keep the probes in the early endosomes, did not affect their early density distribution. Incubation of cells in a hypertonic medium reduced uptake both of [(125)I]TC-AOM and [(125)I]TC-BSA; the uptake of [(125)I]TC-AOM was, however, reduced much more than that of the fluid phase marker. This finding supports the notion that the two probes enter the cells via different routes. PMID- 10518702 TI - Localization of sterically stabilized liposomes in Klebsiella pneumoniae-infected rat lung tissue: influence of liposome characteristics. AB - Sterically stabilized liposomes are able to localize at sites of infection and could serve as carriers of antimicrobial agents. For a rational optimization of liposome localization, the blood clearance kinetics and biodistribution of liposomes differing in poly(ethylene glycol) (PEG) density, particle size, bilayer fluidity or surface charge were studied in a rat model of a unilateral pneumonia caused by Klebsiella pneumoniae. It is shown that all liposome preparations studied localize preferentially in the infected lung compared to the contralateral non-infected lung. A reduction of the PEG density or rise in particle size resulted in a higher uptake by the mononuclear phagocyte system, lower blood circulation time and lower infected lung localization. Differences in bilayer fluidity did not affect blood clearance kinetics or localization in the infected lung. Increasing the amount of negatively charged phospholipids in the liposome bilayer did not affect blood clearance kinetics, but did reduce localization of this liposome preparation at the site of lung infection. In conclusion, the degree of localization at the infected site is remarkably independent of the physicochemical characteristics of the PEG liposomes. Substantial selective liposome localization can be achieved provided that certain criteria regarding PEG density, size and inclusion of charged phospholipids are met. These properties seem to be a direct consequence of the presence of the polymer coating operating as a repulsive steric barrier opposing interactions with biological components. PMID- 10518703 TI - The regulation of Na(+)-dependent anionic amino acid transport by the rat mammary gland. AB - The regulation of anionic amino acid transport, using radiolabelled D-aspartate as a tracer, by rat mammary tissue explants has been examined. Na(+)-dependent D aspartate uptake by mammary tissue increased between late pregnancy and early lactation and again at peak lactation but thereafter declined during late lactation. In contrast, the Na(+)-independent component of D-aspartate uptake by mammary explants did not change significantly with the physiological state of the donor animals. Premature weaning of rats during peak lactation markedly decreased Na(+)-dependent D-aspartate uptake by mammary tissue. In addition, premature weaning also reduced the effect of reversing the trans-membrane Na(+)-gradient on the fractional loss of D-aspartate from mammary tissue explants. Unilateral weaning of rats during peak lactation revealed that milk accumulation per se reduced the Na(+)-dependent moiety of D-aspartate uptake by mammary tissue suggesting that the transport of anionic amino acids is regulated to match supply with demand. Treating lactating rats with bromocryptine reduced D-aspartate uptake by mammary tissue explants suggesting that the transport of anionic amino acids by the rat mammary gland is regulated by prolactin. PMID- 10518704 TI - Molecular and functional characterization of an amphibian urea transporter. AB - We report the characterization of a frog (Rana esculenta) urea transporter (fUT). The cloned cDNA is 1.4 kb long and contains a putative open reading frame of 1203 bp. In frog urinary bladder, the gene is expressed as two mRNAs of 4.3 and 1.6 kb. The fUT protein is 63.1 and 56.3% identical to rat UT-A2 and UT-B1, respectively. The internal duplication of UT-A2 and UT-B, as well as the double LP box urea transporter signature sequence were found in this amphibian urea transporter. When expressed in Xenopus oocytes, fUT induced a 10-fold increase in urea permeability, which was blocked by both phloretin and mercurial reagents. The fUT protein did not transport thiourea, but the fUT-mediated urea transport was strongly inhibited by this compound. Thus, this amphibian urea transporter displays transport characteristics in between those of UT-A2 and UT-B. PMID- 10518705 TI - Characterization of Mg(2+) efflux from rat erythrocytes non-loaded with Mg(2+). AB - Non-Mg(2+)-loaded rat erythrocytes with a physiological level of Mg(2+)(i) exhibited Mg(2+) efflux when incubated in nominally Mg(2+)-free media. Two types of Mg(2+) efflux were shown: (1) An Na(+)-dependent Mg(2+) efflux in NaCl and Na gluconate medium, which was inhibited by amiloride and quinidine, as was Na(2+)/Mg(2+) antiport in Mg(2+)-loaded rat erythrocytes; and (2) an Na(+) independent Mg(2+) efflux in sucrose medium and choline Cl medium, which may be differentiated into SITS-sensitive Mg(2+) efflux at low Cl(-)(o) (in sucrose) and into SITS-insensitive Mg(2+) efflux at high Cl(-)(o) (in 150 mmol/l choline Cl). PMID- 10518706 TI - Extensive electroporation abolishes experimentally induced shape transformations of erythrocytes: a consequence of phospholipid symmetrization? AB - As shown in earlier work (M.M. Henszen et al., Mol. Membr. Biol. 14 (1997) 195 204), exposure of erythrocytes to single brief electric field pulses (5-7 kV cm( 1)) enhances the transbilayer mobility of phospholipids and produces echinocytes which can subsequently be transformed into stomatocytes in an ATP-dependent process. These shape transformations arise from partly reversible changes of the transbilayer disposition of phospholipids, in agreement with the bilayer couple concept. Extensive membrane modification by repetitive (/=5 microM to monocytes in culture for 24 h, significantly increases low density lipoprotein (LDL) binding and uptake, up-regulates levels of LDL receptors and also induces proinflammatory cytokine (interleukin-1, interleukin-6 and tumour necrosis factor alpha) production and glutathione reductase activity. Because it is known that various cells selectively internalize surface receptors and their ligands through receptor-mediated endocytosis via clathrin-coated pits, we tested whether antibodies raised against the clathrin heavy chain would block the effects of the fibrillar form of C-36 on human monocytes in culture. Addition of excess anti-(clathrin HC) with 10 microM fibrillar C-36 diminished the stimulatory effects of the latter on LDL binding, uptake and LDL receptor levels. In contrast, however, in the presence of anti-(clathrin HC), the potentially cytotoxic effects of fibrils, such as induction of cytokines, free radicals and cytosolic activity of cathepsin D, were much greater than those observed when cells were treated with fibrils alone. These results suggest that endocytosis is the pathway by which C-36 fibrils upregulate LDL receptors, and may be the natural mechanism for fibril clearance. We infer that human monocytes clear C-36 fibrils by a clathrin-dependent pathway, presumably endocytotic, and that loss of this pathway amplifies the cytotoxic effects of the fibrils by increasing their availability to other specific or nonspecific sites through which they exert their cytotoxic effects. PMID- 10518781 TI - Structure and transcriptional regulation of the ovine placental lactogen gene. AB - Ovine placental lactogen (oPL), a member of the growth hormone/prolactin gene family, is produced by chorionic binucleate cells at the maternal-fetal interface, and is thought to modulate metabolic processes and enhance fetal growth. We have determined that the oPL gene contains five exons and four introns, and the transcriptional start site was mapped 91 bp 5' of the initiation codon (AUG). An additional 4.5 kb of 5'-flanking sequence was sequenced and used for transient transfection analysis in human (BeWo) and rat (Rcho-1) choriocarcinoma cell lines to examine trophoblast cell-specific activity. Trophoblast cell-specific transactivation of the reporter gene was conferred by the proximal 1. 1 kb of oPL gene 5'-flanking sequence. Transfection of deletion constructs derived from the 1.1 kb of 5'-flanking sequence resulted in varying profiles of transactivation between the two choriocarcinoma cell lines, but maximal activation in both cell lines resided within the proximal 383 bp of oPL gene 5'-flanking sequence. DNase I protection analysis using ovine chorionic binucleate cell nuclear protein, identified 19 footprints within the 1.1-kb sequence, six of which are located within the 383-bp region. Electrophoretic mobility-shift assays and mutational analysis identified two functional GATA ( 67, -102) sequences as transactivators of the oPL gene. However, a previously undefined element (GAGGAG) residing at -338 and -283 is required for full transactivation, and mutation of either significantly reduces reporter activity. In addition, an AP-2 site (-58) and an E-box (-163) were identified and may coordinate oPL transactivation. Transcriptional regulation of human and rodent PL genes has been previously characterized, and our results indicate that tissue specific regulation of oPL expression may result from cis-acting elements in common with human and rat genes expressed within the placenta. However, our data indicate that regulation of oPL also results from novel cis-acting elements. PMID- 10518782 TI - Cloning, sequencing and heterologous expression of pyrogallol-phloroglucinol transhydroxylase from Pelobacter acidigallici. AB - A genomic lambda-library of Pelobacter acidigallici has been established. Proteolytic digestion of homogeneous pyrogallol-phloroglucinol transhydroxylase from the same microorganism afforded polypeptide fragments whose N-terminal sequences allowed the generation of oligonucleotide primers. Together with primers deduced from the known N-terminal sequences of the two intact subunits these were used in PCR experiments to obtain three amplificates. Screening the lambda-library with the three amplificates led eventually to clones containing the whole gene coding for the transhydroxylase. Sequencing the gene revealed two open reading frames coding for 875 and 275 amino acids which correspond to the alpha- and beta-subunits of THL, respectively. The two subunits are separated by a 48-bp noncoding region. Comparison of the sequence with those of other molybdopterin cofactor (MoCo)-enzymes places THL in the dimethylsulfoxide reductase family. Possible contact sites to the MoCo and to the iron-sulphur clusters were spotted. Using the expression vectors pQE 30 and pT 7-7 three constructs harbouring the THL gene were created. One of them carried a His6-tag at the N-terminus of the alpha-subunit, another at the C-terminus of the beta subunit. Immunoblot analysis showed high expression of THL, but the inclusion bodies could not be refolded to active enzyme. PMID- 10518783 TI - How do acetylcholine receptor ligands reach their binding sites? AB - The access pathway to the binding sites for large competitive antagonists of the nicotinic acetylcholine receptor from Torpedo californica electric tissue was analyzed by binding and photolabeling experiments with alpha-neurotoxins. Binding assays with [125I]alpha-bungarotoxin showed an increase in the number of accessible binding sites upon stepwise solubilization of the receptor-rich membranes. Similarily, ligand binding is facilitated upon fluidization of the membrane by increasing the temperature. The access to the binding sites seems to be sterically 'hindered' in the densely packed membrane state. Using a novel series of large biotinylated photoactivatable derivatives of neurotoxin II, we observed that the accessibility to the alpha/gamma- but not to the alpha/delta binding site was considerably decreased for some derivatives under native conditions. This effect was less apparent at higher temperatures and could be abolished by complete solubilization. These observations support the nonequivalence of the receptor's binding sites. Together, our data suggest (a) that alpha-neurotoxins approach their binding sites from the membrane-facing periphery of the receptor's extramembrane domain rather than through the channel mouth and (b) that different entrance pathways to each binding site exist which vary in their sensitivity to the physical state of the plasma membrane. PMID- 10518784 TI - Corticotropin increases protein tyrosine phosphatase activity by a cAMP-dependent mechanism in rat adrenal gland. AB - Corticotropin signal transduction pathway involves serine/threonine protein phosphorylation. Recent reports suggest that protein tyrosine dephosphorylation may also be an integral component of that pathway. The present study was performed to investigate the role played by protein tyrosine phosphatases (PTPs) on acute response to corticotropin and the hypothetical regulation of PTPs by this hormone. We have used two powerful cell permeant PTP inhibitors, phenylarsine oxide (PAO) and pervanadate (PV), in order to examine the relevance of PTP activity on hormone-stimulated and 8-bromo-adenosine 3',5'-phosphate (8Br cAMP is a permeant analogue of adenosine 3',5'-phosphate)-stimulated steroidogenesis in adrenal zona fasciculata (ZF) cells. In both cases, PAO and PV inhibited the steroid production in a dose-dependent fashion, and had no effect on steroidogenesis supported by a permeant analogue of cholesterol. The effect of hormonal stimulation on PTP activity was analyzed in rat adrenal ZF. In vivo corticotropin treatment reduced phosphotyrosine content in endogenous proteins and produced a transient increase of PTP activity in the cytosolic fraction, reaching a maximum (twofold) after 15 min. Incubation of adrenal ZF with 8Br-cAMP also produced PTP activation, suggesting that it can be mediated by cAMP dependent protein kinase (PKA)-dependent phosphorylation. Detection of PTP activity in an in-gel assay showed three corticotropin-stimulated soluble PTPs with molecular masses of 115, 80 and 50 kDa. In summary, we report for the first time a hormone-dependent PTP activation in a steroidogenic tissue and provide evidence that PTP activity plays an important role in corticotropin signal pathway, acting downstream of PKA activation and upstream of cholesterol transport across the mitochondrial membrane. PMID- 10518785 TI - Studies of the Escherichia coli Trp repressor binding to its five operators and to variant operator sequences. AB - The Escherichia coli Trp repressor binds to promoters of very different sequence and intrinsic activity. Its mode of binding to trp operator DNA has been studied extensively yet remains highly controversial. In order to examine the selectivity of the protein for DNA, we have used electromobility shift assays (EMSAs) to study its binding to synthetic DNA containing the core sequences of each of its five operators and of operator variants. Our results for DNA containing sequences of two of the operators, trpEDCBA and aroH are similar to those of previous studies. Up to three bands of lower mobility than the free DNA are obtained which are assigned to complexes of stoichiometry 1 : 1, 2 : 1 and 3 : 1 Trp repressor dimer to DNA. The mtr and aroL operators have not been studied previously in vitro. For DNA containing these sequences, we observe predominantly one retarded band in EMSA with mobility corresponding to 2 : 1 complexes. We have also obtained retardation of DNA containing the trpR operator sequence, which has only been previously obtained with super-repressor Trp mutants. This gives bands with mobilities corresponding to 1 : 1 and 2 : 1 complexes. In contrast, DNA containing containing a symmetrized trpR operator sequence, trpRs, gives a single retarded band with mobility corresponding solely to a 1 : 1 protein dimer-DNA complex. Using trpR operator variants, we show that a change in a single base pair in the core 20 base pairs can alter the number of retarded DNA bands in EMSA and the length of the DNase I footprint observed. This shows that the binding of the second dimer is sequence selective. We propose that the broad selectivity of Trp repressor coupled to tandem 2 : 1 binding, which we have observed with all five operator sequences, enables the Trp repressor to bind to a limited number of sites with diverse sequences. This allows it to co-ordinately control promoters of different intrinsic strength. This mechanism may be of importance in a number of promoters that bind multiple effector molecules. PMID- 10518786 TI - Mutations of glutamate-84 at the putative potassium-binding site affect camphor binding and oxidation by cytochrome p450cam. AB - Cytochrome P450cam (CYP101) from Pseudomonas putida is unusual among P450 enzymes in that it exhibits co-operative binding between the substrate camphor and a potassium ion. This behaviour has been investigated by mutagenesis of Glu84, a surface residue which forms part of the cation-binding site. Substitutions that neutralize or reverse the charge of this side chain are shown to disrupt the co operativity of potassium and camphor binding by P450cam, and also to influence the catalytic activity. In particular, replacement of Glu84 by positively charged residues such as lysine results in increased high-spin haem fractions and camphor turnover activities in the absence of potassium, along with decreased camphor dissociation constants. However, in the presence of potassium the camphor dissociation constants of these mutants are significantly increased compared with the wild-type, although the camphor turnover activities remain marginally higher. In contrast, substitution by aspartate results in tighter binding of both potassium and camphor, but has little effect on the enzymatic activity. In all cases the reaction remains essentially 100% coupled and gives 5-exo hydroxycamphor as the only product. These results suggest that an anionic side chain at the 84 position is crucial for the co-operativity of camphor and cation binding, and that the physiological role for potassium binding by cytochrome P450cam is to promote camphor binding even at the expense of turnover rate, thus allowing the organism to utilize low environmental concentrations of this substrate for growth. PMID- 10518788 TI - Purification and characterization of thyroid-hormone-binding protein from masu salmon serum. A homolog of higher-vertebrate transthyretin. AB - We purified a thyroid-hormone-binding protein (THBP) from serum of masu salmon at the stage of smoltification when the concentrations of endogenous thyroid hormones in plasma reach the highest levels. All steps of sequential column chromatography suggest that this THBP is responsible for most L-3,5,3' triiodothyronine-binding activity in serum at this stage. The molecular mass of this protein was estimated to be 60 kDa by gel filtration but only 15 kDa by SDS/PAGE, which suggests that it is comprised of four identical subunits. The amino acid sequence of its N-terminal portion was highly similar to those of vertebrate transthyretins. These molecular features indicate that masu salmon THBP is a homolog of transthyretins from tetrapods. However, in contrast with mammalian transthyretins, the affinity of masu salmon transthyretin for L-3,5,3' triiodothyronine was three times greater than for L-thyroxine. This rank order affinity is similar to that of avian and frog transthyretins. Scatchard analysis revealed that masu salmon transthyretin possesses a single class of binding site for L-3,5,3'-triiodothyronine, with a Kd of 13.8 nM at 0 degrees C. Taken together with the data reported by Chang et al. [Eur. J. Biochem. (1999) 259, 534 542], these results suggest that transthyretin has changed from a L-3,5, 3' triiodothyronine-carrier protein to a L-thyroxine-carrier protein during mammalian evolution. PMID- 10518787 TI - Human CDC45 protein binds to minichromosome maintenance 7 protein and the p70 subunit of DNA polymerase alpha. AB - Budding yeast CDC45 encodes Cdc45p, an essential protein required to trigger initiation of DNA replication in late G1 phase. We cloned four and one species of the human Cdc45p homolog cDNA, resulting from different splicing patterns, from HeLa cell and human placenta cDNA libraries, respectively. A comparison of the cDNAs and the genomic sequence showed that the longest encoding a 610-amino acid protein was comprised of 20 exons. One species, which lacks exon 7 and contains the shorter of two exons 18, was identical with the previously reported CDC45L cDNA and constituted 24 out of 28 clones from HeLa cells. Splicing was different in HeLa cells and TIG-1 cells, a human diploid cell line. Human CDC45 protein was found to bind directly in vitro to human minichromosome maintenance 7 protein (hMCM7) and to the p70 subunit of DNA polymerase alpha. The data support a thesis that human CDC45 acts as a molecular tether to mediate loading of the DNA polymerase alpha on to the DNA replication complex through binding to hMCM7. PMID- 10518789 TI - Enzymatic properties of human 25-hydroxyvitamin D3 1alpha-hydroxylase coexpression with adrenodoxin and NADPH-adrenodoxin reductase in Escherichia coli. AB - We have cloned human 25-hydroxyvitamin D3 1alpha-hydroxylase cDNAs from normal subjects and patients with pseudovitamin D-deficient rickets (PDDR), and expressed the cDNAs in Escherichia coli JM109 cells. Kinetic analysis of normal 1alpha-hydroxylase in the reconstituted system revealed that Km values for 25(OH)D3 and (24R), 25(OH)2D3 were 2.7 and 1.1 microM, respectively. The lower Km value and higher Vmax/Km value for (24R),25(OH)2D3 indicated that it is a better substrate than 25(OH)D3 for 1alpha-hydroxylase. These results are quite similar to those of mouse 1alpha-hydroxylase. To establish a highly sensitive in vivo system, 1alpha-hydroxylase, adrenodoxin and NADPH-adrenodoxin reductase were coexpressed in E. coli cells. The recombinant E. coli cells showed remarkably high 1alpha-hydroxylase activity, suggesting that the electrons were efficiently transferred from NADPH-adrenodoxin reductase through adrenodoxin to 1alpha hydroxylase in E. coli cells. Using this system, the activities of four mutants of 1alpha-hydroxylase, R107H, G125E, R335P and P382S, derived from patients with PDDR were examined. Although no significant reduction in expression of these mutants was observed, none showed detectable activity. These results strongly suggest that the mutations found in the patients with PDDR completely abolished 1alpha-hydroxylase activity by replacement of one amino acid residue. PMID- 10518790 TI - N-terminal and C-terminal plasma membrane anchoring modulate differently agonist induced activation of cytosolic phospholipase A2. AB - The 85 kDa cytosolic phospholipase A2 (cPLA2) plays a key role in liberating arachidonic acid from the sn-2 position of membrane phospholipids. When activated by extracellular stimuli, cPLA2 undergoes calcium-dependent translocation from cytosol to membrane sites which are still a matter of debate. In order to evaluate the effect of plasma membrane association on cPLA2 activation, we constructed chimeras of cPLA2 constitutively targeted to the plasma membrane by the N-terminal targeting sequence of the protein tyrosine kinase Lck (Lck-cPLA2) or the C-terminal targeting signal of K-Ras4B (cPLA2-Ras). Constitutive expression of these chimeras in Chinese hamster ovary cells overproducing the alpha2B adrenergic receptor (CHO-2B cells) did not affect the basal release of [3H]arachidonic acid, indicating that constitutive association of cPLA2 with cellular membranes did not ensure the hydrolysis of membrane phospholipids. However, Lck-cPLA2 increased [3H]arachidonic acid release in response to receptor stimulation and to increased intracellular calcium, whereas cPLA2-Ras inhibited it, compared with parental CHO-2B cells and CHO-2B cells producing comparable amounts of recombinant wild-type cPLA2. The lack of stimulation of cPLA2-Ras was not due to a decreased enzymatic activity as measured using an exogenous substrate, or to a decreased phosphorylation of the protein. These results show that the plasma membrane is a suitable site for cPLA2 activation when orientated correctly. PMID- 10518791 TI - Nitration of angiotensin II by .NO2 radicals and peroxynitrite. .NO protects against .NO2 radical reaction. AB - To react with peptides, nitric oxide.NO has to be activated by oxidation, or by coupling with superoxide (O.-2) thereby producing peroxynitrite. In the course of.NO oxidation,.NO2 free radicals and N2O3 may be formed. Using gamma irradiation methods, we characterized the products formed by these nitrogen oxides with angiotensin II. Angiotensin II is specifically nitrated at its tyrosinyl residue by.NO2 or peroxynitrite. Equimolecular amounts of each reagent in K+/Pi solutions at pH 7.4 led to 56% and 5% nitration yields, respectively. Nitrogen oxides produced by autoxidation of.NO, as well as.NO2 under.NO, reacted only with the arginine residue, giving a mixture of peptides containing citrulline, a N-(hydroxylamino-cyanamido-) instead of guanido group, and a conjugated diene derived from an arginine side-chain. However, nitrosation reactions by N2O3 occurred only when the initial concentration of.NO2 was 10 times that able to react with angiotensin II. Thus, in this case.NO appears to protect against.NO2 action. PMID- 10518792 TI - Cloning and characterization of new orphan nuclear receptors and their developmental profiles during Tenebrio metamorphosis. AB - Five PCR fragments corresponding to a part of the DNA-binding domain of different hormone nuclear receptors were isolated from Tenebrio molitor mRNAs. The sequence identity of three of them with known Drosophila nuclear receptors strongly suggests that they are the Tenebrio orthologs of seven-up, DHR3 and beta-FTZ-F1, and thus named Tmsvp, TmHR3 and TmFTZ-F1. The full-length sequences of the other two were established. TmHR78 is either a new receptor of the DHR78 family or the same gene which has evolved rapidly, particularly in the E domain. TmGRF belongs to the GCNF1 family and its in vitro translated product binds to the extended half site TCAAGGTCA with high affinity. The periods of expression of the corresponding transcripts in epidermal cells during Tenebrio metamorphosis were analyzed as a function of 20-hydroxyecdysone titers measured in the hemolymph of the animals taken for RNA extraction. Comparison of the expression profiles of these nuclear receptors with those observed during Drosophila metamorphosis revealed similar temporal correlations as a function of ecdysteroid variations, which further supported the sequence identity data for TmSVP, TmHR3, TmFTZ-F1 and TmHR78. PMID- 10518793 TI - Postsynaptic short-chain neurotoxins from Pseudonaja textilis. cDNA cloning, expression and protein characterization. AB - Two lethal proteins, which specifically bind to the nAChR from Torpedo californica, were isolated from the venom of Pseudonaja textilis, the common brown snake from Australia. The isolated proteins have masses of 6236 and 6345 Da and are structurally related to short-chain neurotoxins from other elapids. Six cDNAs encoding isoforms of related neurotoxins were cloned using the RT-PCR of the venom gland mRNAs. The sequences of the corresponding proteins consist of 57 58 amino acid residues and display several unique features when compared with all known short-chain neurotoxins. Accordingly, they grouped separately in phylogenetic analysis. The six cDNAs were expressed in Escherichia coli and the recombinant proteins were characterized. They have similar masses and display similar toxicities and binding constants to the nAChR as the native toxins isolated from the venom. Thus, a new group of short-chain postsynaptic neurotoxins from the venom of an Australian elapid has been characterized. PMID- 10518795 TI - Pore-forming peptides of Entamoeba dispar. Similarity and divergence to amoebapores in structure, expression and activity. AB - Amoebapore, a 77-residue peptide with pore-forming activity from the human pathogen Entamoeba histolytica, is implicated in the killing of phagocytosed bacteria and in the cytolytic reaction of the amoeba against host cells. Previously, we structurally and functionally characterized three amoebapore isoforms in E. histolytica but recognized only one homolog in the closely related but non-pathogenic species Entamoeba dispar. Here, we identified two novel amoebapore homologs from E. dispar by molecular cloning. Despite strong resemblance of the primary structures of the homologs, molecular modeling predicts a species-specific variance between the peptide structures. Parallel isolation from trophozoite extracts of the two species revealed a lower amount of pore-forming peptides in E. dispar and substantially higher activity of the major isoform from E. histolytica towards natural membranes than that from E. dispar. Differences in abundance and activity of the lytic polypeptides may have an impact on the pathogenicity of amoebae. PMID- 10518794 TI - Biochemical and topological properties of type A MGDG synthase, a spinach chloroplast envelope enzyme catalyzing the synthesis of both prokaryotic and eukaryotic MGDG. AB - MGDG synthase, the enzyme that catalyzes the synthesis of the major chloroplast membrane lipid monogalactosyldiacylglycerol (MGDG), is encoded by a multigenic family. We have analyzed the biochemical properties, subcellular localization and membrane topology of a spinach chloroplast MGDG synthase, a representative member of the type A family from Spinacia oleracea (soMGD A), using a recombinant protein that was functionally overexpressed in Escherichia coli and specific polyclonal antibodies. We demonstrated that soMGD A could catalyze the synthesis of both 'prokaryotic' and 'eukaryotic' MGDG molecular species in vitro, with a selectivity for diacylglycerol similar to that of purified chloroplast envelope MGDG synthase activity. Furthermore, soMGD A was shown to be sensitive to chemical reagents (dithiothreitol, N-ethylmaleimide and o-phenanthroline) known to affect MGDG synthesis by the partially purified enzyme, as well as in isolated chloroplast envelope membranes. In spinach chloroplasts, soMGD A was localized by Western blot analysis in the inner envelope membrane. Topological studies demonstrated that soMGD A is a monotopic enzyme, embedded within one leaflet of the inner envelope membrane from spinach chloroplasts, a structure which may involve amphipathic alpha helices. We further demonstrated that in vitro, soMGD A precursor is imported and processed to its correct mature form in intact chloroplasts. These results show that soMGD A corresponds to a mature polypeptide of approximately 45 kDa. In addition, inactivation kinetics after gamma-ray irradiation strongly suggest that both native chloroplast envelope MGDG synthase and recombinant soMGD A have a functional molecular mass of 95-100 kDa, indicating that they are probably active as homodimers made of two 45-kDa subunits. This study suggests that, in spite of the growing evidence that MGDG synthesis is catalyzed by a multigenic family of enzymes, in spinach leaves both prokaryotic and eukaryotic MGDG syntheses could be attributable to a unique dimeric enzyme, provided that diacylglycerol is transported from the outer membrane to the inner membrane of the chloroplast envelope. PMID- 10518796 TI - Veratryl alcohol-mediated oxidation of isoeugenyl acetate by lignin peroxidase. AB - The mechanism of the veratryl alcohol (VA)-mediated oxidation of isoeugenyl acetate (IEA) by lignin peroxidase, and the subsequent spontaneous Calpha-Cbeta cleavage of IEA to vanillyl acetate were studied. IEA oxidation only occurred in the presence of VA. It probably did not bind to lignin peroxidase as evidenced by an unaffected Km for VA in the presence of IEA, and by the fact that a 10-fold molar excess of the unreactive IEA counterpart, eugenyl acetate, did not affect the IEA oxidation rate. IEA was very efficient in recycling VA. Up to 34 mol of IEA were oxidized per mol VA. Formation of the predominant VA oxidation product, veratraldehyde, was postponed until IEA was almost completely oxidized. Together these findings suggest that IEA was oxidized by VA.+ rather than directly by lignin peroxidase. Thus, VA functioned as a redox mediator during IEA oxidation which is remarkable considering the high calculated ionization potential of 8.81 eV. Regardless of the presence of O2, approximately 2 mol of IEA were consumed per mol H2O2, which indicated that IEA was enzymatically oxidized by one electron to the putative radical cation (IEA.+). After formation of IEA.+, a series of O2 dependent chemical reactions were responsible for Calpha-Cbeta cleavage to the major oxidation product vanillyl acetate, as evidenced by the observation that an N2 atmosphere did not inhibit IEA oxidation, but almost completely inhibited vanillyl acetate formation. GC-MS analyses revealed that under an air atmosphere 1-(4'-acetoxy-3'-methoxyphenyl)-2-propanone, 1-(4'-acetoxy-3'-methoxyphenyl)-1 hydroxy-2-propanone, and 1-(4'-acetoxy-3'-methoxyphenyl)-2-hydroxy-1-propanone were also formed. Formation of the latter two was diminished under an N2 atmosphere. PMID- 10518797 TI - Characterization and molecular cloning of an adenosine kinase from Babesia canis rossi. AB - In the search for immunoprotective antigens of the intraerythrocytic Babesia canis rossi parasite, a new cDNA was cloned and sequenced. Protein sequence database searches suggested that the 41-kDa protein belongs to the phosphofructokinase B type family (PFK-B). However, because of the low level sequence identity (< 20%) of the protein both with adenosine and sugar kinases from this family, its structural and functional features were further investigated using molecular modelling and enzymatic assays. The sequence/structure comparison of the protein with the crystal structure of a member of the PFK-B family, Escherichia coli ribokinase (EcRK), suggested that it might also form a stable and active dimer and revealed conservation of the ATP binding site. However, residues specifically involved in the ribose-binding sites in the EcRK sequence (S and N) were substituted in its sequence (by H and M, respectively), and were suspected of binding adenosine compounds rather than sugar ones. Enzymatic assays using a purified glutathione S-transferase fusion protein revealed that this protein exhibits rapid catalysis of the phosphorylation of adenosine with an apparent Km value of 70 nM, whereas it was inactive on ribose or other carbohydrates. As enzymatic assays confirmed the results of the structure/function analysis indicating a preferential specificity towards adenosine compounds, this new protein of the PFK-B family corresponds to an adenosine kinase from B. canis rossi. It was named BcrAK. PMID- 10518798 TI - Autoantibodies interacting with purified native thyrotropin receptor. AB - Native thyrotropin receptor (TSHR) was purified by immunoaffinity chromatography from membrane extracts of stably transfected L cells. An ELISA test was devised to study anti-TSHR autoantibodies directly. Comparison of native TSHR with bacterially expressed, denatured TSHR showed that the latter was not recognized by the autoantibodies, suggesting that they bind to conformational epitopes only present on the native receptor. The use of deglycosylated TSHR and of purified receptor ectodomain (alpha-subunit) showed that the autoantibodies recognized only the protein backbone moiety of the receptor and that their epitopes were localized entirely in its ectodomain. Autoantibodies were detected in 45 of 48 subjects with untreated Graves' disease and in 26 of 47 healthy volunteers. The affinity for the receptor was similar in the two groups (Kd = 0.25-1 x 10-10 M) and the autoantibodies belonged to the IgG class in all cases. Although the concentration of autoantibodies was higher in Graves' disease patients (3.50 +/- 0.36 mg.L-1) than in control subjects (1.76 +/- 0.21) (mean +/- SEM), there was an overlap between the groups. Receptor-stimulating autoantibodies (TSAb) were studied by measuring cAMP synthesis in stably transfected HEK 293 cells. Their characteristics (recognition of alpha-subunit, of deglycosylated TSHR, nonrecognition of bacterially expressed denatured receptor) were similar to those of the antibodies detected by the ELISA test. TSAb were only found in individuals with Graves' disease. The ELISA test measures total anti-TSHR antibodies, whereas the test using adenylate cyclase stimulation measures antibodies that recognize specific epitopes involved in receptor activation. Our observations thus disprove the hypothesis according to which Graves' disease is related to the appearance of anti-TSHR antibodies not present in normal subjects. Actually, anti-TSHR antibodies exist in many euthyroid subjects, in some cases even at concentrations higher than those found in patients with Graves' disease. What distinguishes the latter from normal subjects is the existence of subpopulation(s) of antibodies directed against specific epitope(s) of the receptor involved in its activation. PMID- 10518799 TI - Impact of C5-cytosine methylation on the solution structure of d(GAAAACGTTTTC)2. An NMR and molecular modelling investigation. AB - The solution structures of d(GAAAACGTTTTC)2 and of its methylated derivative d(GAAAAMe5CGTTTTC)2 have been determined by NMR and molecular modelling in order to examine the impact of cytosine methylation on the central CpG conformation. Detailed 1H NMR and 31P NMR investigation of the two oligomers includes quantitative NOESY, 2D homonuclear Hartmann-Hahn spectroscopy, double-quantum filtered COSY and heteronuclear 1H-31P correlation. Back-calculations of NOESY spectra and simulations of double-quantum-filtered COSY patterns were performed to gain accurate information on interproton distances and sugar phase angles. Molecular models under experimental constraints were generated by energy minimization by means of the molecular mechanics program JUMNA. The MORASS software was used to iteratively refine the structures obtained. After methylation, the oligomer still has a B-DNA conformation. However, there are differences in the structural parameters and the thermal stability as compared to the unmethylated molecule. Careful structural analysis shows that after methylation CpG departs from the usual conformation observed in other ACGT tetramers with different surroundings. Subtle displacements of bases, sugars and backbone imposed by the steric interaction of the two methyl groups inside the major groove are accompanied by severe pinching of the minor groove at the C-G residues. PMID- 10518800 TI - The responses of rat hepatocytes to glucagon and adrenaline. Application of quantified elasticity analysis. AB - The internal control of hepatocyte metabolism has been previously analysed using metabolic control analysis. The aim of this paper is to extend this analysis to include the responses of the cells to hormonal stimulus. Hepatocyte metabolism was divided into nine reaction blocks: glycogen breakdown, glucose release, glycolysis, lactate production, NADH oxidation, pyruvate oxidation, proton leak, mitochondrial phosphorylation and ATP consumption, linked by five intermediates: mitochondrial membrane potential, cytoplasmic NADH/NAD and total cellular ATP, glucose 6-phosphate and pyruvate. The kinetic responses of the reaction blocks to the intermediates were determined previously in the absence of added hormones. In this study, the changes in flux and intermediate levels that occurred upon addition of either glucagon or adrenaline were measured. From comparison of the fractional changes in fluxes and intermediate levels with the known kinetics of the system, it was possible to determine the primary sites of action of the hormones. The results show that the majority of processes in the cell are responsive to the hormones. The notable exception to this is the failure of adrenaline to have a direct effect on glycolysis. The activity change of each metabolic block observed in the presence of either hormone was quantified and compared to the indirect effects on each block caused by changes in metabolite levels. The second stage of the analysis was to use the calculated activity changes and the known control pattern of the system to give a semiquantitative analysis of the regulatory pathways employed by the hormones to achieve the changes in fluxes and metabolite levels. This was instructive in analysing, for example, how glucagon caused a decrease in flux through glycolysis and an increase in oxidative phosphorylation without large changes in metabolite levels (homeostasis). Conversely, it could be seen that the failure of adrenaline to maintain a constant glucose 6-phosphate concentration was due to the stimulation of glycogen breakdown and inhibition of glucose release. PMID- 10518801 TI - Cloning and expression of succinic semialdehyde reductase from human brain. Identity with aflatoxin B1 aldehyde reductase. AB - The neuromodulator gamma-hydroxybutyrate is synthesized in vivo from gamma aminobutyrate by transamination to succinic semialdehyde and subsequent reduction of the aldehyde group. In human brain, succinic semialdehyde reductase is thought to be responsible for the conversion of succinic semialdehyde to gamma hydroxybutyrate. In the present work, we cloned the cDNA coding for succinic semialdehyde reductase and expressed it in Escherichia coli. A data bank search indicated that the enzyme is identical with aflatoxin B1-aldehyde reductase, an enzyme implicated in the detoxification of xenobiotic carbonyl compounds. Structurally, succinic semialdehyde reductase thus belongs to the aldo-keto reductase superfamily. The recombinant protein was indistinguishable from native human brain succinic semialdehyde reductase by SDS/PAGE. In addition to succinic semialdehyde, it readily catalyzed the reduction 9,10-phenanthrene quinone, phenylglyoxal and 4-nitrobenzaldehyde, typical substrates of aflatoxin B1 aldehyde reductase. The results suggest multiple functions of succinic semialdehyde reductase/aflatoxin B1 aldehyde reductase in the biosynthesis of gamma-hydroxybutyrate and the detoxification of xenobiotic carbonyl compounds, respectively. PMID- 10518802 TI - Epitope analysis and molecular modeling reveal the topography of the C-terminal peptide of the beta-subunit of human chorionic gonadotropin. AB - Human chorionic gonadotropin (hCG) belongs to a family of heterodimeric glycoprotein hormones that share a common alpha-subunit and a hormone-specific beta-subunit. Among the gonadotropin beta-subunits, greater than 85% homology exists between lutropin (hLH)beta and hCGbeta in their first 114 amino acid residues. However, unlike hLHbeta, hCGbeta contains a 31-amino acid hydrophilic stretch at its carboxyl end (CTPbeta: C-terminal peptide). Although the crystal structure of deglycosylated hCG has been solved, the topography of CTPbeta remains unknown. In this study, we have attempted to define the topology of CTPbeta using mAb probes. We investigated three epitopes on hCGalpha, which are hidden in the hCGalphabeta dimer. However, these epitopes are not hidden in hLH, which has a similar subunit interface to that of hCG, but lacks CTPbeta. This suggested that these epitopes are not masked at the subunit interface of hLH or hCG. Hence, we hypothesized that, in the case of hCG, these epitopes are masked by the CTPbeta. Consistent with this view, several treatments of hCG that removed CTPbeta unmasked these epitopes and enhanced their reactivity with the corresponding mAbs. In order to localise the position of CTPbeta on the alpha subunit, we used an epitope-mapping strategy [N. Venkatesh & G. S. Murthy (1997) J. Immunol. Methods 202, 173-182] based on differential susceptibility of epitopes to covalent modifications. This enabled us to predict the possible topography of CTPbeta. Further, we were also able to build a model of CTPbeta, completely independently of the epitope-mapping studies, using a homology-based modeling approach [S. Krishnaswamy, I. Lakshminarayanan & S. Bhattacharya (1995) Protein Sci. 4 (Suppl. 2), 86-97]. Results obtained from these two different approaches (epitope analysis and homology modeling) agree with each other and indicate that portions of CTPbeta are in contact with hCGalpha in the native hCG dimer. PMID- 10518803 TI - Structural analysis of the lipopolysaccharide oligosaccharide epitopes expressed by Haemophilus influenzae strain RM.118-26. AB - The structure of the lipopolysaccharide of Haemophilus influenzae mutant strain, RM.118-26, was investigated. Electrospray ionization-mass spectrometry on intact lipopolysaccharide, O-deacylated lipopolysaccharide and core oligosaccharides obtained from lipopolysaccharide after mild acid hydrolysis provided information on the composition and relative abundance of the glycoforms. Oligosaccharide samples were studied in detail using high-field NMR techniques. The structure of the major glycoform containing phosphocholine is identical to the Hex2 glycoform described for H. influenzae RM.118-28 [Risberg, A., Schweda, E.K.H. & Jansson, P. E. (1997) Eur. J. Biochem. 243, 701-707]. A second major glycoform, containing three hexose residues (Hex3), in which a lactose unit, beta-D-Galp-(1-->4)-beta-D Glcp, is attached at the O-2 position of the terminal heptose of the inner core element, L-alpha-D-Hepp-(1-->2)-L-alpha-D-Hepp-(1-->3)-[beta-D-Glcp-( 1-->4)-]- L alpha-D-Hepp-(1-->5)-alpha-Kdo, carries no phosphocholine. Instead this lipopolysaccharide glycoform is partly (40%) substituted by an O-acetyl group linked to the 6-position of the glucose residue in the lactose unit and has the following structure: PMID- 10518804 TI - Ca2+-induced p38/SAPK signalling inhibited by the immunosuppressant cyclosporin A in human peripheral blood mononuclear cells. AB - To understand the effects of the immunosuppressant cyclosporin A (CsA) on Ca2+ mediated intracellular signalling pathways in human peripheral blood mononuclear cells (PBMCs), we investigated its effects on the activity profiles of mitogen activated protein kinase (MAPK) cascades. PBMCs, or subpopulations thereof, were simultaneously stimulated with a phorbol ester and the calcium ionophore ionomycin, in the presence or absence of therapeutic concentrations of CsA. In these primary human cells, CsA significantly inhibited PMA/ionomycin-mediated and ionomycin-mediated activation of the MAPK kinase MKK6, as well as its downstream kinases SAPK2a (p38alpha) and MAPKAP-K2. PMA/ionomycin treatment also mediated activation of SAPK1 (JNKs) which was inhibited by CsA. Treatment with ionomycin alone also resulted in CsA-sensitive activation of SAPK1. With regard to transcription factors targeted by the Ca2+-induced MAPK signalling network, we found CsA to inhibit the ionomycin-mediated phosphorylation of ATF2 at Thr71. We identified the heterodimeric transcription factor ATF2/CREB as constitutively binding to the essential cAMP response element (CRE) site within the Ca2+ regulated DNA polymerase beta promoter and contributing to the activation of this promoter. Our data implicate ATF2 phosphorylation status as a nuclear sensor within PBMCs that monitors converging intracellular Ca2+-signalling pathways. PMID- 10518805 TI - Is beta-poly(L-malate) synthesis catalysed by a combination of beta-L-malyl-AMP ligase and beta-poly(L-malate) polymerase? AB - beta-Poly(L-malate) is supposed to function in the storage and transport of histones, DNA polymerases and other nuclear proteins in the giant syncytical cells (plasmodia) of myxomycetes. Here we report on the biosynthesis of [14C]beta poly(L-malate) from injected L-[14C]malate in the plasmodium of Physarum polycephalum. The effects of KCN, arsenate, adenosine 5'-(alpha, beta methylene)triphosphate, adenosine 5'-(beta, gamma-methylene)triphosphate, guanosine 5'-(beta, gamma-methylene)triphosphate, desulfo coenzyme A and phenylarsinoxid on beta-poly(L-malate) synthesis were studied after their coinjection with L-[14C]malate. The synthesis was not affected by KCN or desulfo coenzyme A, but was blocked by arsenate and adenosine 5'-(alpha,beta methylene)triphosphate. The plasmodium lysate catalysed an L-malate-dependent ATP [32P]pyrophosphate exchange, but was devoid of beta-poly(L-malate) synthetic activity under all experimental conditions tested. The results suggested an extramitochondrial synthesis of beta-poly(L-malate), involving the polymerization of beta-L-malyl-AMP. It is assumed that the lack of synthesis in the lysate is caused by the inactivation of beta-poly(L-malate) polymerase involving a cell injury kinase pathway. Because injected guanosine 5'-(beta, gamma methylene)triphosphate blocks the synthesis, the injury signal is likely to be GTP dependent. PMID- 10518806 TI - The pneumococcal common antigen C-polysaccharide occurs in different forms. Mono substituted or di-substituted with phosphocholine. AB - The structure of the pneumococcal common antigen, C-polysaccharide, from a noncapsulated pneumococcal strain, CSR SCS2, was studied using 1H-NMR, 13C-NMR and 31P-NMR spectroscopy. The dependence of NMR chemical shifts on the variation in pD was also studied. It was established that the C-polysaccharide is composed of a backbone of tetrasaccharide-ribitol repeating units that are linked to each other by a phosphodiester linkage between position 5 of a D-ribitol residue and position 6 of a beta-D-glucopyranosyl residue. The polysaccharide is substituted with one residue of phosphocholine at position 6 of the 4-substituted 2-acetamido 2-deoxy-alpha-D-galactopyranosyl residue. Both galactosamine residues in the polysaccharide are N-acetylated. O)-P-Cho | 6 6)-beta-D-Glcp-(1-->3)-alpha-AATp (1-->4)-alpha-D-GalpNAc-(1-->3)- bet a-D-GalpNAc-(1-->1)-D-ribitol-5-P-(O--> where AAT is 2-acetamido-4-amino-2,4,6-trideoxy-D-galactose and Cho is choline. This structure differs, concerning phosphocholine substituents and N-acetylation, from those reported previously for pneumococcal C-polysaccharide [Jennings, H.J., Lugowski, C. & Young, N.M. (1980) Biochemistry 19, 4712-4719; Fischer, W., Behr, T., Hartmann, R., Peter-Katalinic, J. & Egge, H. (1993) Eur. J. Biochem. 215, 851 857; Kulakowska, M., Brisson, J.-R., Griffith, D.W., Young, N.M. & Jennings, H.J. (1993) Can. J. Chem. 71, 644-648]. The structures of the C-polysaccharides present in three pneumococcal types were also examined. They contain one (in 18B) or two (in 32F and 32A) phosphocholine residues in the repeating unit. The degree of substitution was not determined. The backbone of all examined C polysaccharides was identical and in all cases both galactosamine residues appeared to be N-acetylated. PMID- 10518807 TI - Pyruvate dehydrogenase from Azotobacter vinelandii. Properties of the N terminally truncated enzyme. AB - The pyruvate dehydrogenase multienzyme complex (PDHC) catalyses the oxidative decarboxylation of pyruvate and the subsequent acetylation of coenzyme A to acetyl-CoA. Previously, limited proteolysis experiments indicated that the N terminal region of the homodimeric pyruvate dehydrogenase (E1p) from Azotobacter vinelandii could be involved in the binding of E1p to the core protein (E2p) [Hengeveld, A. F., Westphal, A. H. & de Kok, A. (1997) Eur J. Biochem. 250, 260 268]. To further investigate this hypothesis N-terminal deletion mutants of the E1p component of Azotobacter vinelandii pyruvate dehydrogenase complex were constructed and characterized. Up to nine N-terminal amino acids could be removed from E1p without effecting the properties of the enzyme. Truncation of up to 48 amino acids did not effect the expression or folding abilities of the enzyme, but the truncated enzymes could no longer interact with E2p. The 48 amino acid deletion mutant (E1pdelta48) is catalytically fully functional: it has a Vmax value identical to that of wild-type E1p, it can reductively acetylate the lipoamide group attached to the lipoyl domain of the core enzyme (E2p) and it forms a dimeric molecule. In contrast, the S0.5 for pyruvate is decreased. A heterodimer was constructed containing one subunit of wild-type E1p and one subunit of E1pdelta48. From the observation that the heterodimer was not able to bind to E2p, it is concluded that both N-terminal domains are needed for the binding of E1p to E2p. The interactions are thought to be mainly of an electrostatic nature involving negatively charged residues on the N-terminal domains of E1p and previously identified positively charged residues on the binding and catalytic domain of E2p. PMID- 10518809 TI - Pathophysiology of the inflammatory response. AB - Airway allergic reactions enlist diverse cells and a multitude of chemical mediators that are responsible for the clinical symptoms of allergic rhinitis and asthma. Experiments in vitro and in animal models, as well as increasingly numerous studies in atopic human subjects, are revealing that an orchestrated continuum of cellular activities leading to airway allergic inflammation is set in motion in genetically predisposed individuals at the first exposure to a novel antigen. This sensitization step likely depends on differentiation of and cytokine release by T(H)2 lymphocytes. Among T(H)2-derived cytokines, IL-4 potently enhances B-lymphocyte generation of immunoglobulin E antibodies. The attachment of these antibodies to specific receptors on airway mast cells sets the stage for an acute inflammatory response on subsequent antigen exposure because IgE cross-linking by a bound antigen activates mast cells to release numerous inflammatory mediators. These mast cell-derived mediators collectively produce acute-phase clinical symptoms by enhancing vascular leak, bronchospasm, and activation of nociceptive neurons linked to parasympathetic reflexes. Simultaneously, some mast cell mediators up-regulate expression on endothelial cells of adhesion molecules for leukocytes (eosinophils, but also basophils and lymphocytes), which are key elements in the late-phase allergic response. Chemoattractant molecules released during the acute phase draw these leukocytes to airways during a relatively symptom-free recruitment phase, where they later release a plethora of cytokines and tissue-damaging proteases that herald a second wave of airway inflammatory trauma (late-phase response). The repetition of these processes, with the possible establishment in airway mucosa of memory T lymphocytes and eosinophils that are maintained by paracrine and autocrine cytokine stimulation, may account for airway hypersensitivity and chronic airway symptoms. PMID- 10518810 TI - Mechanisms of intranasal steroids in the management of upper respiratory allergic diseases. AB - Intranasal steroids have proved to be the most effective class of drugs in reducing the symptoms of allergic rhinitis. This clinical response reflects the broad anti-inflammatory activity that has been demonstrated for corticosteroids. Single doses of topical corticosteroids administered before nasal allergen challenge block the late-phase reaction, whereas repeated dosing with intranasal steroids blocks both the early and the late response, as well as the priming phenomenon. Nasal inflammation is accomplished through a number of effector cells and mechanisms, which in turn are produced by director cells through the release of cytokines and chemokines. The anti-inflammatory action of corticosteroids is largely effected through blocking the synthesis and release of these cytokines/chemokines. PMID- 10518811 TI - Intranasal corticosteroids for allergic rhinitis: how do different agents compare? AB - Intranasal steroids have proved to be an effective and safe form of therapy for allergic rhinitis. However, as the number of new glucocorticoid compounds has increased over the past decade, it has become important to consider whether significant differences exist between these agents. Pharmacologically, newer drugs such as mometasone furoate and fluticasone propionate appear to have substantially higher topical potencies and lipid solubilities and lower systemic bioavailabilities than do older compounds. In clinical use, however, all the available drugs appear to be equally effective in controlling symptoms of seasonal and perennial allergic rhinitis. With respect to adverse effects, emerging data suggest that mometasone furoate and fluticasone propionate may have less potential for systemic effects during prolonged use, particularly in children. Newer intranasal steroids appear to have practical advantages over older agents that may favor their use in some groups of patients with allergic rhinitis. PMID- 10518812 TI - A review of the preclinical and clinical data of newer intranasal steroids used in the treatment of allergic rhinitis. AB - The anti-inflammatory activity of corticosteroids has prompted the exploration of their use in the treatment of allergic rhinitis. The development of intranasal steroids has resulted in several agents with quick actions, localized effects, and great efficacy in the treatment of seasonal allergic rhinitis and the prophylactic management of perennial rhinitis. This article presents a concise review of the preclinical and clinical evidence with these new agents and provides data-based guidance for the selection of optimal agents. The survey reveals that mometasone furoate, a new inhaled steroid with topical activity, has the greatest binding affinity for the glucocorticoid receptor, followed by fluticasone propionate, budesonide, triamcinolone acetonide, and dexamethasone. Mometasone furoate also has strong anti-inflammatory activity, with IL-4 and IL-5 inhibition activities equivalent to those of fluticasone propionate. Clinically, both mometasone furoate and fluticasone propionate appear to be well tolerated, to have quick onsets of action, and to be equivalent in efficacy in the treatment of seasonal allergic and perennial rhinitis. Of the intranasal steroids currently available, mometasone furoate has been shown to have the least systemic availability and, consequently, is expected to have the fewest systemic side effects. Some suppression of overnight cortisol levels has been reported with fluticasone propionate (indicative of hypothalamic-pituitary-adrenal axis suppression). PMID- 10518814 TI - Air pollution and asthma. AB - Asthma is a disease syndrome that has captured a great deal of attention for several years. One of the perplexing aspects to asthma is that the prevalence is increasing in most industrialized countries. The reasons for this widespread increase are largely unknown. Another aspect of industrialization is the persistence of air pollution in urban areas. Because much air pollution is due to vehicles, no solution appears in sight. The topic of this article is the association between air pollution and various signs and symptoms of asthma. Air pollution is convincingly associated with many signs of asthma aggravation. These include pulmonary function decrements, increased bronchial hyperresponsiveness, visits to emergency departments, hospital admissions, increased medication use and symptom reporting, inflammatory changes, interactions between air pollution and allergen challenges, and immune system changes. With the exception of exposure to environmental tobacco smoke, common air pollutants have not been shown to cause asthma. It seems prudent for clinicians to counsel their patients about the potential risks of asthma aggravation from common outdoor air pollutants. PMID- 10518815 TI - Chemokines and allergic disease. AB - Understanding the chemokine network has become one of the great challenges for researchers interested in inflammatory mechanisms and inflammation-based diseases. The complexity and diversity of the system provide not only a daunting task for its comprehension but also numerous opportunities for development of new, targeted therapies. It is now certain that chemokines are involved as important mediators of allergic inflammation; the fine details and scope of their roles are now under investigation. Presumably, because of distinct pressures on the immune systems of people living in different geographic regions, genetic variation of ligands, receptors, and regulatory regions in the network have emerged. Establishing the roles of these polymorphisms in determining disease susceptibility or progression among individuals and in distinct ethnic groups will provide a basis for improved understanding and treatment of allergic diseases. PMID- 10518816 TI - Modulation of nitric oxide responses in asthma by extracellular antioxidants. PMID- 10518817 TI - Nitrotyrosine formation in the airways and lung parenchyma of patients with asthma. AB - BACKGROUND: Recent evidence has shown that nitric oxide (NO) levels are increased in asthmatic airways. Although the role of NO in asthma is unknown, reactive metabolites of NO may lead to nitrotyrosine formation and promote airway dysfunction. OBJECTIVE: The aim of this study was to determine whether nitrotyrosine, as a marker of nitrating species, could be found in the airways and lung parenchyma of subjects with asthma who died of status asthmaticus or other nonrespiratory causes. METHODS: Lung tissue specimens were obtained from 5 patients who died of status asthmaticus, 2 asthmatic patients who died of nonrespiratory causes, and 6 nonasthmatic control subjects who died of nonrespiratory causes. Lung sections were stained for immunofluorescence with use of an antinitrotyrosine antibody, followed by a indiocarbocyanine (Cy5, Jackson Immunochemicals, Westgrove, Pa)-conjugated secondary antibody. RESULTS: Nonasthmatic lungs showed little or no nitrotyrosine staining, whereas asthmatic lungs demonstrated significantly more staining of nitrotyrosine residues distributed in both the airways and lung parenchyma. CONCLUSION: This study demonstrates the presence of nitrotyrosine, and hence evidence of formation of nitrating species, in the airways and lung parenchyma of patients with asthma who died of status asthmaticus or other nonrespiratory causes. This finding supports the concept that widespread airway and parenchymal inflammation occurs in asthma, and, more specifically, that NO and its reactive metabolites may play a pathophysiologic role in asthma. PMID- 10518819 TI - Fatal and near-fatal asthma questionnaire: prelude to a national registry. AB - BACKGROUND: Asthma mortality rates continue to be unacceptably high in the United States. As a follow-up to the initiatives proposed by the Asthma Mortality Task Force in 1987, the Committee on Asthma Mortality of the American Academy of Allergy, Asthma, and Immunology developed a questionnaire on fatal and near-fatal asthma. OBJECTIVE: This study assessed completeness of answers from participating physicians and described characteristics of patients with fatal and near-fatal asthma. METHODS: There were 111 survey items intended to characterize patients with fatal or near-fatal exacerbations of asthma. The questionnaire was sent to the members of the American Academy of Allergy, Asthma, and Immunology (approximately 3900), and a total of 143 usable questionnaires were received. RESULTS: Responding physicians had information on most items in the questionnaire; the mean number of responses was 120 of 143 possible, with a median of 128 and a range of 40 to 143. Patient demographics, description of the event, identification of overall risk for the event, use of medications, and characteristics of asthma management had the most complete responses (median response rates were 126-143). Presence of factors contributing to the event had fewer responses (range, 44-125). The physicians frequently had information on some psychologic characteristics (eg, 108 responses for depression and/or hopelessness and 127 responses for social support) but less information on several others (eg, 62 responses for family dysfunction). Statistical analysis of the completed surveys indicated that only 2 characteristics distinguished fatal from near-fatal asthma: progression in minutes (adjusted odds ratio, 4. 89; 95% confidence interval, 2.05-12.90) and absence of a past history of intubation (adjusted odds ratio, 3.55; 95% confidence interval, 1.55-8.97). CONCLUSIONS: There is a need to gather further data on patients with fatal and near-fatal events to design appropriate prospective studies on asthma morbidity and mortality rates. Physicians can contribute important information about these patients. Gathering such data would be enhanced by establishing a national registry on fatal and near-fatal asthma. PMID- 10518818 TI - Influence of interaction of environmental risk factors and sensitization in young asthmatic children. AB - BACKGROUND: The increasing prevalence of asthma and allergy in many countries demands evaluation of potential risk factors to improve the possibility of prevention. OBJECTIVE: We studied the association between exposure to cat and dog allergen and allergic sensitization in young children with asthma and interactions with potential environmental risk factors. METHODS: One hundred eighty-nine young children with asthma were evaluated. IgE antibodies to cat and dog were analyzed. Questionnaires were filled in focusing on exposure to cats and dogs, environmental tobacco smoke (ETS), and signs of home dampness as indicated by window pane condensation (WPC) during the first years of life. House dust was analyzed for content of cat (Fel d 1) and dog (Can f 1) allergen. RESULTS: There was a strong association between the degree of reported exposure to cat and dog and the concentration of the respective allergens in floor dust. A dose-response relationship was found between cat exposure, measured as either reported degree of cat exposure or cat allergen levels in dust, and sensitization both to cat and dog. No such relationship was found between exposure and sensitization to dog. WPC increased the risk for sensitization to cat (odds ratio = 2.6, 95% confidence interval 1.2-5.8), whereas ETS strongly tended to do so both to cat and dog. Interaction was found between exposure to ETS, WPC, and high levels of cat allergen (>8 microg/g dust). The presence of all 3 risk factors revealed a multiplicative interaction with a high risk of sensitization to cat (odds ratio = 42.0, 95% confidence interval 3.7-472.8). CONCLUSIONS: Keeping cats indoors may be a health hazard for infants and young children at risk for development of asthma, particularly when they live in a damp house and their parents smoke. PMID- 10518820 TI - Serum osteocalcin and procollagen as markers for the risk of osteoporotic fracture in corticosteroid-treated asthmatic adults. AB - BACKGROUND: Dual energy x-ray absorptiometry provides the definitive measure of osteoporotic fracture risk. OBJECTIVE: We sought to determine whether metabolic measures of bone formation and/or common features of clinical hypercortisonism provide a useful guide in selecting corticosteroid-treated asthmatic patients for referral for bone densitometry. METHODS: We measured bone density and 8 AM serum osteocalcin, procollagen, and cortisol levels in 52 asthmatic adults aged 60.7 +/ 12.6 years (mean +/- SD). Years of steroid exposure for these patients was 11.8 +/- 10.7 (prednisone) and 11.78 +/- 4.98 (inhaled steroid). Using stepwise logistic regression, we assessed the capacity of the osteocalcin and procollagen levels, with or without the cortisol level, age, clinical features of hypercortisonism, and different lifetime exposures to inhaled and oral steroids for distinguishing between patients with greater or lesser risk of fracture. RESULTS: Osteoporosis, defined as a bone density T score below -2.5, affected 26% of the group at the spine and 63% at the hip. At the spine, greater risk was associated only with lower cortisol levels (P =.003). Diagnostic accuracy was 71%, the false-positive rate was 26%, and the false-negative rate was 31%. At the hip, greater risk was associated with lower cortisol levels (P =.002), longer prednisone exposure, (P =.003), lower current doses of prednisone (P =.01) and inhaled steroid (P =.02), and older age (P =.01). Diagnostic accuracy was 83%, the false-positive rate was 13%, and the false-negative rate was 21%. CONCLUSIONS: Neither osteocalcin nor procollagen nor any of the clinical criteria analyzed proved sufficiently accurate to be reliable as indicators of the risk of fracture in these elderly, corticosteroid-treated asthmatic adults. They are therefore not useful for selecting such patients for diagnostic densitometry. PMID- 10518821 TI - The relationships among environmental allergen sensitization, allergen exposure, pulmonary function, and bronchial hyperresponsiveness in the Childhood Asthma Management Program. AB - BACKGROUND: Sensitivity and exposure to indoor allergens constitutes a risk factor for the development and persistence of asthma in children. OBJECTIVE: Our purpose was to evaluate the relationship between sensitivity and exposure to inhalant allergens and lung function and bronchial responsiveness in a group of children (n = 1041) aged 8.9 +/- 2.1 years with mild to moderate asthma enrolled in the Childhood Asthma Management Program (CAMP). METHODS: With use of the extensive CAMP baseline cross-sectional data on spirometry, bronchial responsiveness, allergen sensitivities, and household allergen levels, the relationship of sensitization and exposure to allergens to lung function and methacholine sensitivity was evaluated. Children who enrolled in CAMP stopped all antiasthma medication except rescue use of albuterol and prednisone for exacerbations during the 5- to 16-week screening period. During the last 2 of these weeks they underwent spirometry and methacholine challenge. Indoor allergen exposures were determined from questionnaires completed by the parent. Household levels of indoor allergens (mite, cat, dog, cockroach, mold) were determined on house dust samples. Allergen sensitivity was determined by percutaneous skin testing with a standard battery of allergens plus locally important pollen and fungal spores. Lung function and bronchial hyperresponsiveness were compared for children sensitive and not sensitive to both indoor and outdoor allergens on skin testing and, if sensitive, for exposed and not exposed to the allergens to which they were positive on skin testing. RESULTS: There was a strong direct correlation between increased sensitivity to inhaled methacholine and skin test sensitivity to tree, weed, Alternaria, cat, dog, and indoor molds. When the relationship was examined by stepwise regression, the skin test sensitivities showing the strongest associations with the concentration of methacholine that caused a 20% fall in FEV(1) were dog (P =.003), Alternaria (P =.01), and cat (P =.05). Children sensitive to any one of the aeroallergens tested were compared for the presence or absence of exposure to that allergen at the time that the methacholine challenge was performed. Those who were sensitive and exposed to weed and cat had greater methacholine sensitivity than those similarly sensitive but not exposed (P =.003 and P =.02, respectively). CONCLUSIONS: Sensitivity to dog or cat dander or Alternaria by skin testing was associated with increased bronchial responsiveness but not decreased lung function in children with mild to moderate asthma. These findings support the important role that sensitization to certain allergens plays in modulating bronchial responsiveness. PMID- 10518822 TI - Elevated plasma eotaxin levels in patients with acute asthma. AB - BACKGROUND: The eosinophil chemotactic and activating effects of eotaxin and the known association of eosinophils with asthma suggest that eotaxin expression is increased during asthma exacerbations. OBJECTIVE: We sought to determine whether plasma eotaxin levels are elevated in patients presenting for emergency treatment of acute asthma and to correlate eotaxin levels with disease activity and responses to treatment. METHODS: A case-control study of plasma eotaxin levels was performed in the 46 patients who presented for emergency asthma treatment and 133 age-, sex-, and ethnicity-matched subjects with stable asthma. RESULTS: Plasma eotaxin levels were significantly higher in 46 patients with acute asthma symptoms and airflow obstruction (520 pg/mL [250, 1100 pg/mL]; geometric mean [-1 SD, +1 SD]) than in 133 subjects with stable asthma (350 pg/mL [190, 620 pg/mL]; P =.0008). Among the patients with emergency asthma flares, those who responded to asthma treatment with an increase in peak expiratory flow rate by an amount equal to at least 20% of their predicted normal value had lower eotaxin levels than those who did not (410 pg/mL [210, 800 pg/mL] and 660 pg/mL [300, 1480 pg/mL], respectively; P =.04). CONCLUSION: These findings imply that eotaxin either is mechanistically involved in acute asthma or serves as a biomarker for activity of the CCR3 receptor ligand system, which is functionally linked to asthma. PMID- 10518824 TI - Identification, isolation, and characterization of Ole e 7, a new allergen of olive tree pollen. AB - BACKGROUND: Olive tree (Olea europaea) pollen is an important cause of pollinosis in countries of the Mediterranean area and California. OBJECTIVE: The aim of this study was to identify and purify a new allergen of olive tree pollen. METHODS: Detection of a pollen allergen was done with individual allergic sera by immunoblotting and ELISA tests. Two allergenic fractions were isolated from olive pollen extract by using gel filtration and reverse-phase HPLC. Molecular characterization was achieved by acid hydrolysis and amino acid analysis, as well as by mass spectrometry. Sequencing of the N-terminal end of the allergen was carried out by Edman degradation of the polypeptide chain. Allergenic characterization was performed with sera from subjects with olive allergy by means of ELISA and immunoblotting after SDS-PAGE. RESULTS: The new allergen Ole e 7 exhibits a high degree of polymorphism. Its molecular mass is in the range of 9875 d to 10,297 d. Twenty-one amino acid residues from the N-terminal end of 2 isoforms of the allergen have been sequenced revealing no homology with proteins contained in database banks. Ole e 7 has an average frequency of about 47% in patients with olive allergy. The strategy of purification of Ole e 7 can be useful on the isolation of new allergens. CONCLUSIONS: A new olive pollen allergen of clinical significance has been purified and characterized, contributing to the study of the complete allergogram of the olive tree pollen. PMID- 10518823 TI - Physician-derived asthma diagnoses made on the basis of questionnaire data are in good agreement with interview-based diagnoses and are not affected by objective tests. AB - BACKGROUND: Defining the phenotype is critical for investigating the genetic etiology of asthma. As part of the Collaborative Study on the Genetics of Asthma (CSGA), the primary objective of which is to identify asthma susceptibility loci, an algorithm was designed to determine diagnoses of definite asthma, probable asthma, less than probable asthma, or no asthma. A respiratory questionnaire was designed to assist in the process of characterizing the asthma phenotype. OBJECTIVE: This study was designed to determine the validity of the CSGA algorithm for the diagnosis of asthma, to determine agreement in assessing an asthma diagnosis between the information obtained by the CSGA questionnaire versus a patient interview by a panel of specialist physicians, and to determine the degree to which objective tests would alter the questionnaire-based certainty of asthma diagnosis. METHODS: An expert panel of asthma clinicians (n = 4) indicated to what degree they were certain that a subject (n = 48) had asthma as determined by using a 6-point Likert scale based on a 20-minute interview (phase I), a review of the CSGA questionnaire (phase II), a review of the questionnaire plus skin test and peripheral blood eosinophilia data (phase III), and a review of phase III information plus pulmonary data (spirometry and methacholine reversibility testing; IV). Intraclass correlation coefficients (ICCs) were calculated between the physicians' interpretation of the likelihood of asthma based on the information they received during each of the phases and between the CSGA algorithm and each of the phases. RESULTS: Interjudge reliability with regard to the degree of certainty with which an asthma diagnosis could be made by interview was excellent (ICC, 98; 95% confidence intervals [95% CIs], 0.87-0.99). We also found that the agreement between the physicians' interview with the patients (phase I) and the CSGA algorithm was good and at least as good with the addition of the CSGA questionnaire data and objective data (ICC, 0. 65-0.75). Good agreement was also observed between the average certainty score from the interview and the CSGA questionnaire (ICC, 92; 95% CI, 0.76-0.93), and ICCs determining the agreement on asthma diagnosis between phase I and phases III and IV, in which objective data were introduced, did not change from the ICCs comparing phase I with phase II (ICC of 0.93 [95% CI, 0.79-0.96] and ICC of 0.91 [95% CI 0.73-0.95], respectively). CONCLUSION: We conclude that the CSGA algorithm is a valid tool for which the diagnosis of asthma can be made at an acceptable level of certainty and that the CSGA questionnaire, interpreted by an asthma specialist, is a useful tool for the diagnosis of asthma in clinical or epidemiologic studies. PMID- 10518825 TI - Increased progenitor cell proliferation in the peripheral blood of patients with bronchial asthma: the role of nitric oxide. AB - BACKGROUND: Asthma exacerbation is associated with increased numbers of circulating CD34(+) progenitor cells, which may migrate to airways and develop into mature cells under the effects of cytokines and hematopoietic factors. Nitric oxide (NO) generation is enhanced in asthma and is known to suppress human hematopoiesis. OBJECTIVES: We studied circulating progenitor cells in the blood of patients with varying severity of asthma and examined the contribution of NO to their proliferation into eosinophil-forming colonies ex vivo. METHODS: With use of multiparameter flow cytometric analyses, the cell numbers and intracellular inducible NO synthase (iNOS) immunoreactivity of circulating CD34(+) cells in peripheral blood was measured. The serum level of GM-CSF or IL-5 was also determined. The colonies grown from progenitor cells were cultured in methylcellulose either in the presence or absence of growth factors, including GM CSF, stem cell factor, and IL-3. RESULTS: A significantly greater number of circulating CD34(+) cells increased together with higher intracellular iNOS immunoreactivity in moderate asthmatics compared with mild intermittent asthmatics and healthy subjects. There was no significant difference in iNOS immunoreactivities or CD34(+) progenitor cell numbers between healthy subjects and those with mild intermittent asthma. Serum levels of GM-CSF or IL-5 were significantly higher in all asthmatics compared with healthy subjects and correlated with circulating CD34(+) cells. A greater number of colonies was grown either in the presence or absence of growth factors with a higher percentage of cells of eosinophil lineage in asthmatics than in health subjects. N(G)-nitro-L arginine methyl ester potentiated and sodium nitroprusside inhibited the colony growth in both asthmatic and healthy subjects without a significant change in the percentage of eosinophil lineage. CONCLUSIONS: The production of NO from progenitor cells or other circulating cells may act in an autocrine or paracrine fashion to regulate progenitor cell growth and colony formation. However, this is not sufficient to control the increased proliferation of progenitor cells observed in asthma. PMID- 10518827 TI - Nuclear factor of activated T cells and YY1 combine to repress IL-5 expression in a human T-cell line. AB - BACKGROUND: IL-5 is an inducible T-cell cytokine with the unique ability to induce eosinophilia without increases in other cell compartments. Regulation of IL-5 expression is controlled primarily at the level of transcription. The role of eosinophilia in allergic disorders indicates IL-5 as a target for therapy. OBJECTIVE: This report aims to increase our understanding of IL-5 gene regulation by identifying distal control elements in the human (h) IL-5 promoter, determining the transcription factors that bind these elements and elucidating their role in control of hIL-5 gene expression. METHODS: Methods used in this study include deoxyribonuclease I footprint analysis, electrophoretic mobility shift assay, and functional analysis by transfection of PER-117 cells with site directed mutants of the hIL-5 promoter. RESULTS: We have identified a protected region in the distal hIL-5 promoter that has sequence homology to the previously identified negative regulatory element within BR3. This protected region has not been previously reported and is shown to contain overlapping binding sites for YY1 and nuclear factor of activated cells. The binding sites exist between positions -447 and -459, and this sequence was named hPRE2-IL5. Substitution mutations that abolish binding of these proteins to hPRE2-IL5 result in a 2- to 3 fold increase in hIL-5 promoter activity in activated human T cells. CONCLUSION: We report the novel combination of YY1 and nuclear factor of activated T cells transcription factors binding to a distal hIL-5 promoter element where both factors are involved in down-regulation of hIL-5 gene expression in human T cell. PMID- 10518826 TI - Frequency and characterization of antigen-specific IL-4- and IL-13- producing basophils and T cells in peripheral blood of healthy and asthmatic subjects. AB - BACKGROUND: Both basophils and T cells are known to secrete IL-4 and IL-13 after activation with either nonspecific stimuli or specific antigen, but the relative contribution of these 2 cell types to overall cytokine production is unclear. OBJECTIVES: To further characterize basophil cytokine production and compare it with that of T cells, we examined the frequency of IL-4- and IL-13-producing basophils and T cells in human PBMCs by means of flow cytometry after activation in allergic asthmatic and normal subjects. METHODS: PBMCs obtained from whole blood after Percoll gradient were activated with specific antigen or ionomycin and fixed. PBMCs were made permeable; stained with antibodies to IgE, CD3, and either IL-4 or IL-13; and analyzed by means of flow cytometry. RESULTS: Preformed cytokines were not detected in unactivated basophils. After ionomycin activation, 60% to 90% of basophils from both control and allergic asthmatic subjects expressed IL-4 and IL-13. Specific antigen induced cytokine expression by 10% to 20% of basophils from the asthmatic group only. After specific antigen activation, basophils accounted for 4 times more IL-4-producing cells than did T cells. IL-4 and IL-13 production at 2 hours was exclusively from basophils. After allergen activation, CD40 ligand was upregulated on a subset of peripheral blood basophils. CONCLUSIONS: These data demonstrate that basophils are the predominant peripheral blood cells that express IL-4 and IL-13 in the first 6 hours after antigen activation and strengthen the putative role of basophils both in IgE production and in the generation of allergic inflammation. PMID- 10518828 TI - Regulation of antigen-induced human T-lymphocyte responses by calcineurin antagonists. AB - BACKGROUND: Cyclosporin A (CS) and tacrolimus (FK506, FK) are calcineurin antagonists used widely as T-cell immunosuppressants; however, their relative efficacy on antigen-stimulated T-cell subsets remains undefined. OBJECTIVE: We have examined the effects of CS and FK on antigen-driven proliferation and cytokine generation from human PBMCs and T-cell clones. METHODS: Proliferation was assessed by tritiated thymidine incorporation. Cytokine generation was assessed by reverse transcription-PCR and ELISA. RESULTS: Ragweed- and tetanus toxoid-driven proliferation of PBMCs was down-regulated equally by CS or FK. Gene expression for proinflammatory cytokines (IL-4, IL-5, IL-13, and IFN-gamma) assessed by reverse transcription-PCR was down-regulated in a concentration dependent manner by either drug. Antigen-induced proliferation of ragweed specific Th0, Th1, or Th2 clones was inhibited by either CS or FK. Cytokine gene expression and protein secretion into culture supernatants (IL-4, IL-5, IL-13, and IFN-gamma) were down-regulated in a concentration-dependent manner by either CS or FK in all relevant T-cell subsets. Interestingly, down-regulation of IL-5 protein generation from Th0 and Th2 clones was consistently less sensitive to either drug than was the effect on either IL-4 or IL-13 protein generation. CONCLUSION: CS and FK promote equivalent down-regulation of Th0, Th1, and Th2 responses; however, IL-5 generation is relatively insensitive to the immunomodulatory effects of calcineurin antagonists. PMID- 10518829 TI - Coprinus comatus (shaggy cap) is a potential source of aeroallergen that may provoke atopic dermatitis. AB - BACKGROUND: Basidiospores are universal components in the air and established as important causes of respiratory allergies. Recent reports indicate that aeroallergens may aggravate eczematous skin lesions in subjects with atopic dermatitis (AD). OBJECTIVE: The aim of the study was to investigate whether spores of Coprinus comatus, a species of basidiomycetes, may elicit delayed-type skin reactions in subjects with an atopic predilection, especially dermatitis. METHODS: Sixty-six study subjects were categorized in groups having AD or respiratory allergy with regard to the skin prick test (SPT) reactivity to C comatus extract. Twenty nonatopic individuals served as control subjects. Atopy patch tests (APTs) were performed with extract of C comatus spore containing tissue at a concentration of 1. 35 mg of protein per gram of petrolatum (Vaseline) and C comatus cap at a concentration of approximately 5 mg of protein per gram of petroleum jelly. APT reactions were evaluated after 48 and 72 hours. RESULTS: Of the subjects with AD completing the study, 12 (32%) of 38 showed a positive APT reaction, with 8 (57%) also having a positive SPT response to C comatus. Only 1 (9%) of 11 subjects with asthma had a positive SPT and APT response to C comatus. No positive test reaction was observed in the nonatopic control subjects or in subjects with respiratory allergy and negative SPT responses to C comatus. CONCLUSION: Our results demonstrate that C comatus can induce delayed-type reactions in atopic individuals, particularly in those with AD. Because spores of Coprinus species are ubiquitous, basidiomycetes have to be considered as possible aeroallergens when investigating causes of eczematous skin lesions in AD. PMID- 10518830 TI - Removal of cockroach allergen from inner-city homes. AB - BACKGROUND: Allergen avoidance has been shown to improve the morbidity and physiology of asthma. Although cockroach allergen has been implicated in chronic asthma, little work has been reported on the feasibility of allergen removal from infested homes. OBJECTIVE: The objective of this study was to examine the effect of professional pest control and home cleaning on cockroach infestation and allergen concentrations in settled dust samples from the kitchens, bedrooms, and television-living rooms of inner-city homes. METHODS: Thirteen homes in inner city Baltimore, Maryland, received a professional cleaning, with vacuuming and a thorough cleaning in the kitchen. Pest control technicians applied abamectin 0.05% gel to the kitchen and, to a limited extent, to the rest of the home and the cleaning was repeated. Technicians visited monthly from month 2 to month 8 to inspect, collect dust samples, and place passive cockroach traps. Bla g 1 concentrations in dust extract were measured by means of ELISA. RESULTS: The number of cockroaches captured in passive traps decreased rapidly in 11 homes, but complete extermination was achieved in only 7 homes. Bla g 1 concentrations were reduced by 93% in kitchens, 77% in television-living rooms, and 74% in bedrooms. The relative reduction in cockroach allergen was not related to successful extermination or to signs of poor house-cleaning. CONCLUSION: We concluded that cockroach extermination is feasible in inner-city homes but that standard house-cleaning procedures are only partially effective in removing residual allergen over 8 months. PMID- 10518831 TI - Specific IgG, but not specific IgE, antibodies to toluene diisocyanate-human serum albumin conjugate are associated with toluene diisocyanate bronchoprovocation test results. AB - BACKGROUND: The role of specific IgG to toluene diisocyanate (TDI) in the pathogenesis of TDI-induced asthma still remains unclear. OBJECTIVE: We sought to evaluate the clinical significance of serum-specific IgG to TDI-human serum albumin (HSA) conjugate in subjects with TDI-induced asthma compared with specific IgE antibody. METHODS: One hundred three subjects were enrolled and divided into 4 groups according to specific bronchoprovocation test (BPT) results: 50 subjects with TDI-induced asthma with positive results on TDI BPT were defined as group 1, 13 symptomatic workers exposed to TDI with negative results on TDI BPT were defined as group 2, 20 unexposed patients with allergic asthma were defined as group 3, and 20 unexposed healthy control subjects were defined as group 4. Serum-specific IgG and IgE antibodies to TDI-HSA conjugate were detected by means of ELISA. RESULTS: The prevalence of specific IgG antibody to TDI-HSA conjugate was significantly higher in group 1 than in group 2 (46% vs 7.7%, P =.01) or group 3 (0%, P <.01). No significant difference was noted between group 2 and group 3 (P >. 05). However, the prevalence of specific IgE antibody to TDI-HSA conjugate was not significantly different between group 1 and group 2 (14% vs 7.7%, P >.05) or group 2 and group 3 (7.7% vs 0%, P >.05). There was no significant difference in prevalence of specific IgE or specific IgG according to the type of asthmatic response during the TDI BPT (P >.05). Overall, statistically significant association was noted between the prevalence of specific IgE and IgG antibodies in 103 subjects (P <.05), but no difference was noted within group 1 subjects only (P >.05). CONCLUSION: These findings demonstrate that the presence of serum-specific IgG is closely related to TDI BPT results, and it may contribute to the development of TDI-induced asthma. PMID- 10518832 TI - Reducing relative humidity to control the house dust mite Dermatophagoides farinae. AB - BACKGROUND: Indoor relative humidity (RH) is the key factor that determines the survival and population development of the house dust mite Dermatophagoides farinae. Maintaining RH below 50% is one recommendation in a comprehensive plan to reduce house dust mites and mite allergen levels in homes. Even when mean daily RH is reduced below 50%, RH may rise above 50% intermittently for brief periods because of activities in the home (eg, cooking, bathing, and ventilation). OBJECTIVE: The purpose of this study was to determine how brief daily periods of moist air alternating with long spells of low ambient RH (0% or 35%) influence population survival and growth of D farinae. METHODS: Population growth was determined for D farinae at daily RH regimens of 2, 4, 6, and 8 hours at 75% or 85% RH alternating with 22, 20, 18, and 16 hours at 0% or 35% RH. RESULTS: D farinae populations declined at daily regimens of 2 hours at 75% or 85% RH alternating with 22 hours at 0% or 35% RH. Daily regimens of 4, 6, and 8 hours at 75% RH alternating with 20, 18, and 16 hours, respectively, at 35% RH provided sufficient moisture for small growths in population size. These growths after 10 weeks were reduced by 98.2%, 98.0%, and 97.3% for daily regimens of 4, 6, and 8 hours, respectively, at 75% RH (with the remainder of the day at 35% RH) compared with the growth of populations continuously exposed to 75% RH. Continuous exposure to 85% RH inhibited population growth, but alternating daily regimens of 16, 18, and 20 hours at 35% RH allowed small populations to develop, although they were reduced by 99.4%, 98.8%, and 99.1% compared with population growth at a continuous 75% RH. CONCLUSION: This study indicates that maintaining mean daily RH below 50%, even when RH rises above 50% for 2 to 8 hours daily, effectively restricts population growth of these mites and thus the production of allergen. To completely prevent population growth of D farinae, RH must be maintained below 35% for at least 22 hours per day when the daily RH is 75% or 85% for the remainder of the day. PMID- 10518833 TI - Allergens of Curvularia lunata during cultivation in different media. AB - BACKGROUND: Despite enormous efforts toward the standardization of fungal extracts, only a few extracts have been characterized that are relevant for the diagnosis and immunotherapy of allergy-associated disorders. OBJECTIVE: The goal of this study was to determine the optimum growth condition of Curvularia lunata, an important fungal allergen for quality raw material, and to analyze the C lunata extract for IgE- and IgG-binding proteins. METHODS: C lunata was grown in synthetic (Czapeck Dox medium), semisynthetic (Sabouraud's broth [SB]), and natural media (potato dextrose [PD]) for different periods of time. The extracts were probed for allergenic and antigenic activity, with pooled patient sera and polyclonal antibodies raised against C lunata by means of ELISAs, immunoblots (in vitro), and intradermal tests (in vivo). RESULTS: The growth of C lunata was better in semisynthetic media (ie, SB) compared with other types of media. Dry weight and protein content was maximum in the 7-day culture of SB. ELISA with pooled sera from C lunata-sensitive patients exhibited that cultures grown in SB for 11 to 13 days and PD plus 1.0% agar for 5 days were the most potent. Intradermal tests with 11- to 13-day SB culture extract showed maximum skin reactivity in allergy patients. Immunoblots with patient sera showed 10 to 14 IgE binding proteins in 5- to 15-day SB cultures. Analysis of 5- to 15-day SB extract with rabbit sera showed 10 to 16 IgG-binding proteins. CONCLUSIONS: The extract from 11- to 13-day SB cultures were most biologically potent (intradermal tests) and showed high antigenic and allergenic reactivity (ELISA and immunoblot). The addition of yeast extract did not affect the dry weight and protein content of the C lunata extract. Furthermore, addition of agar in PD medium increases the dry weight and protein content of the fungal mat. Synthetic media was not suitable for mass cultivation of C lunata. PMID- 10518834 TI - The time-course of milk antigen-induced TNF-alpha secretion differs according to the clinical symptoms in children with cow's milk allergy. AB - BACKGROUND: TNF-alpha secretion by blood mononuclear cells stimulated with cow's milk proteins is significantly higher in infants with active cow's milk allergy (CMA) manifested by digestive symptoms than in children who have recovered from CMA. OBJECTIVE: The current study was undertaken to analyze the kinetics of TNF alpha secretion and to evaluate the usefulness of the measurement of TNF-alpha release in whole blood cultures in the prediction of clinical outcome after milk challenge. METHODS: Blood samples were obtained from 83 children maintained on a cow's milk-free diet and examined just before a cow's milk provocation. Children were divided into 4 groups according to clinical outcome: group I (active CMA with cutaneous symptoms), group II (active CMA with predominantly digestive symptoms), group III (children recovered from CMA), and group IV (control). The kinetics of TNF-alpha secretion was measured in blood cultured for 1 to 5 days at different cow's milk protein concentrations. RESULTS: On day 1 TNF-alpha secretion was significantly higher in group I (485 [453] pg/mL, mean [SD], P <.005) and in group II (269 [102] pg/mL, P <. 005) than that observed in groups III and IV (149 [95] and 87 [71] pg/mL, respectively). Then TNF-alpha was rapidly degraded and a second peak of secretion was observed on day 5 but only in group II (278 [221] pg/mL), whereas in groups I, III, and IV a low secretion was observed (70 [61], 45 [40], and 11 [12] pg/mL, respectively, P <. 02). CONCLUSION: These results show that the pattern of TNF-alpha secretion in response to cow's milk proteins is different in CMA infants with cutaneous or digestive symptoms and suggest that TNF-alpha release might predict clinical relapse on challenge. PMID- 10518835 TI - The influence of initial exposure timing to beta-lactoglobulin on oral tolerance induction. AB - BACKGROUND: Although a number of studies have investigated the induction of oral tolerance to several proteins, relatively little is known about the induction of oral tolerance to beta-lactoglobulin, one of the major antigenic proteins in milk. OBJECTIVE: We investigated the influence of the timing of the initial beta lactoglobulin exposure on oral tolerance induction and examined some characteristics of the tolerogenic immune response. METHODS: BALB/c mice were given beta-lactoglobulin prenatally or from the third or fifth postnatal week, bred for 17 weeks, and compared with unexposed control mice. Specific plasma anti beta-lactoglobulin antibodies (total IgG, IgG subclasses, IgM, and IgE), antigen specific splenocyte responses, frequencies of antibody-producing cells, and cytokine production by splenocytes, intestinal mucosal lymphocytes, and Peyer's patches were analyzed. RESULTS: Differences were observed among the 4 groups of mice in changes of plasma anti-beta-lactoglobulin antibody titers, antigen specific T-cell proliferation, and frequencies of antibody-producing splenocytes, intestinal mucosal lymphocytes, and Peyer's patch cells after the first exposure to beta-lactoglobulin. The onset and duration of the immunologic responses were found to be dependent on the timing of antigen exposure. Prenatal exposure to antigen facilitated the induction of oral tolerance to beta-lactoglobulin, whereas delayed antigen exposure retarded tolerance. The induction of oral tolerance was associated with increased IL-4 and/or IL-10 production and decreased IL-12 production. CONCLUSION: Our results suggest that the timing of initial antigen exposure greatly influences the induction of oral tolerance to beta-lactoglobulin and that altered secretion of regulatory cytokines may be responsible for the differences in antibody production and oral tolerance induction. PMID- 10518836 TI - Anaphylaxis to venom of the Pachycondyla species ant. AB - BACKGROUND: In the southeastern United States, imported fire ants have caused systemic reactions with a high incidence. On the contrary, in Korea Pachycondyla species ants (P chinensis and P solitaria), and the family Formicidae, which are in the genus Pachycondyla and the subfamily Ponerinae, have only occasionally caused systemic reactions. OBJECTIVE: We sought to assess whether commercially available imported fire ant extract would be useful in treating patients with anaphylaxis induced by venom from a Pachycondyla species ant. METHODS: Serum samples were collected from 2 women who had anaphylaxis induced by Pachycondyla species ant venom and from 6 volunteers with no history of having been stung. Specific IgE to Pachycondyla species ant extracts was measured by means of ELISA and possible allergenic components by immunoblot. Cross-reactivity between Pachycondyla chinensis, P solitaria, and imported fire ant extracts was also measured by inhibitory ELISA. RESULTS: Skin prick test responses were strongly positive to the extract of P chinensis (1:20 wt/vol) in the patient. Ten healthy volunteers exhibited negative responses. The 2 patients' sera exhibited high ELISA values, with absorbencies of 0.78 and 0.61 for P chinensis and 0.83 and 0.68 for P solitaria, respectively, and negative ELISA values for the extract of imported fire ants (absorbency <0.01). Imported fire ants showed no inhibition of the IgE binding to P chinensis or P solitaria. Possible allergenic components of Pachycondyla species ant extracts are 29- and 27-kd proteins and, less frequently, 16 kd proteins. CONCLUSION: Our data suggest that patients who have had an anaphylactic reaction to a Pachycondyla species ant might not benefit from immunotherapy with an imported fire ant extract. Immunotherapy with the extract of Pachycondyla species ants is expected to be highly effective. PMID- 10518837 TI - Cross-allergenicity of peanut and lupine: the risk of lupine allergy in patients allergic to peanuts. AB - BACKGROUND: Peanut allergy is common, but cross-allergy between legumes is rare. Proteins from Lupinus albus are increasingly eaten in the form of seeds or additives to wheat flour. The risk of cross-allergenicity is still insufficiently known. OBJECTIVE: We sought to study the risk of cross-allergy to lupine in patients allergic to peanut and to study lupine allergenicity. METHODS: Twenty four patients allergic to peanuts were studied by means of skin prick tests with native lupine flour from Lupinus albus. Double-blind oral challenge tests were performed with lupine flour and peanut in 8 of these patients. Specific IgEs were assayed for peanut, lupine flour, and pollen in 6 sera. RAST inhibition tests for lupine pollen by peanut were performed on 4 of these sera. Peanut and lupine flour immunoblots were carried out for 6 sera, and crossed immunoblot inhibitions for peanut by lupine flour and lupine flour by peanut were carried out for 2 sera. RESULTS: The skin prick test responses with lupine flour were positive in 11 (44%) subjects. The challenge test responses were positive in 7 of 8 subjects at the same doses as with peanut. The major lupine flour allergen (molecular mass, 43 kd) is present in peanuts. The RAST inhibition and immunoblot tests indicated cross-reactivity of peanut with the lupine flour and pollen. CONCLUSIONS: The risk of crossed peanut-lupine allergy is high, contrary to the risk with other legumes. The inclusion of 10% lupine flour in wheat flour without mandatory labeling makes lupine a hidden allergen, presenting a major risk of cross-reaction in subjects already allergic to peanut products. A high sensitizing potential can also be postulated for this legume. PMID- 10518838 TI - Macadamia nut anaphylaxis: demonstration of specific IgE reactivity and partial cross-reactivity with hazelnut. PMID- 10518839 TI - Contact sensitivity to multiple local anesthetics. PMID- 10518840 TI - HLA-DR polymorphism in allergic bronchopulmonary aspergillosis. PMID- 10518841 TI - Meeting needs of infants and young children with asthma: new developments in nebulized corticosteroid therapy. Introduction. PMID- 10518842 TI - Clinical need for a nebulized corticosteroid. AB - Asthma is a leading cause of morbidity in children, leading to frequent emergency room visits and hospitalizations. In recent years, a steady increase in the number of cases of pediatric asthma has been observed. Mortality rates among children also have increased. The diagnosis of asthma in infants and young children relies on symptoms, and most children become symptomatic before the age of 2 years. The prognosis of an individual child is difficult to predict because of a lack of clear markers; however, respiratory symptoms that begin at, or persist through, the ages of 3 to 4 years are significantly associated with future asthma. The inflammatory nature of asthma has prompted recommendations for the routine use of inhaled corticosteroids for all patients with persistent asthma. Studies have indicated that early initiation of corticosteroid therapy can reduce airway inflammation and may prevent more serious asthma or irreversible obstruction in later years. Although treatment guidelines recommend the use of inhaled corticosteroids in infants and children younger than 5 years of age, metered-dose and dry-powder inhalers may be difficult for this population to use because of a lack of coordination or understanding. In addition, there are no commercially available inhaled corticosteroids for use in children younger than 4 years of age in the United States. PMID- 10518843 TI - Present and future treatment of asthma in infants and young children. AB - Asthma is a chronic inflammatory disease of the airways characterized by the local production of inflammatory mediators and an increase in recruitment of inflammatory cells (predominantly eosinophils and mast cells). It has been proposed that the chronic nature of this inflammatory response may be responsible for long-term pulmonary changes including bronchial hyperresponsiveness, airway remodeling, and irreversible airflow obstruction. Much of the information available on the pathogenesis of asthma is based on studies performed in young adults. Because of numerous complications, studies in infants and young children are often difficult to conduct; therefore information on this age group is lacking. Although studies are limited, data suggest that an asthma-like inflammation is present at a very early age, with increases in inflammatory cells and thickening of the lung basement membrane detected in infants and young children. In addition, lung function of children with persistent wheezing was significantly lower by 6 years of age compared with children who had no wheezing episodes during the same period; differences between groups were not apparent at 6 months of age. These data suggest that airway inflammation in young children with asthma is associated with nonreversible impairment of lung function. Recent National Asthma Education and Prevention Guidelines stress the importance of anti inflammatory agents, particularly inhaled corticosteroids, in the treatment of young children with persistent asthma. Given data supporting the presence of an inflammatory response early in the disease course, early intervention with anti inflammatory agents may be indicated at the onset of symptoms to prevent long term, irreversible, impairment of lung function. Although several studies have shown that inhaled corticosteroids may be effective in the treatment of recurrent wheezing in infancy, tools need to be developed to distinguish infants with early onset asthma from those with transient wheezing. PMID- 10518844 TI - Pharmacodynamics and pharmacokinetics of budesonide: a new nebulized corticosteroid. AB - Underutilization of anti-inflammatory agents in the treatment of asthma has received widespread attention. Inhaled glucocorticosteroids are important agents for the management of asthma in children and adults. Budesonide has a high ratio of topical anti-inflammatory to systemic activity and is one of the most extensively used inhaled glucocorticoids. Budesonide inhalation suspension is the first formulation designed to deliver budesonide by way of nebulization for infants and children under 8 years of age with persistent asthma. Budesonide decreases airway hyperresponsiveness and reduces the number of inflammatory cells and mediators present in the airways of patients with asthma. Budesonide appears to be retained within cells, allowing for a once-daily treatment regimen in certain patient groups. After inhalation of nebulized budesonide, absorption is rapid. Data suggest that plasma concentrations of budesonide are similar in adults and children after inhalation of the same nominal dose from a nebulizer. In children 3 to 6 years of age, total systemic availability of budesonide after dosing with a jet nebulizer was approximately 6% of the labeled dose. Budesonide is highly protein bound, undergoes extensive first-pass hepatic metabolism, is metabolized by the liver cytochrome P450 system, and is primarily excreted in the urine as metabolites. In children 3 to 6 years of age, the volume of distribution at steady state of budesonide inhalation suspension is approximately 3 L/kg, with a terminal elimination half-life of 2.3 hours; systemic clearance is approximately 30 mL/kg. The pharmacodynamic and pharmacokinetic properties of budesonide inhalation suspension allow for potent local anti-inflammatory activity with limited systemic exposure. PMID- 10518845 TI - Study designs and challenges in clinical studies conducted in infants and children with asthma. AB - Because of the challenges associated with conducting clinical trials in pediatric patients, most drugs have not been adequately tested in this patient population. Insufficient testing frequently results in product labeling that fails to provide instructions for safe and effective use in pediatric patients. The most prevalent chronic disease in children is asthma; however, very few asthma medications have been evaluated in infants and young children. Budesonide inhalation suspension is an anti-inflammatory corticosteroid administered by nebulization that requires only passive inhalation and is therefore suitable for infants and young children who are unable to use currently available inhalation devices for the administration of asthma medications. The efficacy of budesonide inhalation suspension was evaluated in 3 US clinical trials. All 3 trials were randomized, controlled, double-blind, 12-week studies in children (6 months-8 years of age) with a primary diagnosis of chronic persistent asthma. Symptom assessments conducted in both the morning and evening were the primary efficacy variables. Although the children were young, secondary efficacy evaluations included spirometry and peak expiratory flow collected in both the morning and evening (in a subset of patients capable of performing these maneuvers). Patients who completed or were discontinued from the 12-week double-blind treatment phase were eligible to enter 52-week, open-label extension studies. The design of these 3 studies confirms the feasibility of the performance of placebo-controlled clinical trials of asthma medications in pediatric patients and allows for efficacy evaluations based on symptom assessments together with pulmonary function testing. PMID- 10518846 TI - Efficacy of budesonide inhalation suspension in infants and young children with persistent asthma. Budesonide Inhalation Suspension Study Group. AB - This paper reviews the results from 3 randomized, double-blind, placebo controlled, multicenter studies that assessed the efficacy and safety of once- and twice-daily dosing of budesonide inhalation suspension (BIS) in infants and young children with persistent asthma. In children with mild persistent asthma that was previously treated with bronchodilators or noncorticosteroid anti inflammatory agents (study A), nighttime and daytime asthma symptoms were significantly improved in 0.25-mg once daily, 0.5-mg once daily, and 1.0-mg once daily BIS treatment groups compared with placebo (P Colgate Regular > Crest Regular. In all of the assays the Arm & Hammer Dental Care dentifrice had significantly (p < 0.01-0.001) higher DDR mean values compared to the Colgate Regular and Crest Regular dentifrices. The Colgate Regular dentifrice was significantly (p < 0.01-0.001) superior to the Crest Regular dentifrice in the assays conducted. PMID- 10518864 TI - Preliminary report: laboratory-induced stain removal as assessed by environmental scanning electron microscopy. AB - Environmental scanning electron microscopy (ESEM) was employed to observe stain removal during brushing with Arm & Hammer Dental Care and Crest Regular Toothpaste. ESEM allows serial examinations of the same sample, and does not require a destructive preparative process. Three extracted molars were cleaned, placed into a 96-hour broth culture of Streptococcus mutans, and stain was produced with undiluted chlorhexidine rinse, concentrated coffee and tea for a period of 23 days. After staining, the teeth were examined by ESEM, then brushed using a toothbrushing machine. Imaging was repeated after 5, 10, 15 and 30 seconds of brushing. As seen with ESEM, the Arm & Hammer product had different effects than those from the distilled water control, suggesting something other than that expected from abrasive and mechanical forces alone. There were also differences from the Crest dentifrice removal on this single sample, suggesting a possible difference between the two products. Further studies are needed to confirm and explain these effects. PMID- 10518865 TI - A longitudinal comparison of tooth whitening resulting from dentifrice use. AB - The effect of twice-daily brushing with one of three different dentifrices (Arm & Hammer Dental Care, Arm & Hammer Dental Care Extra Whitening, Crest) on stain removal and tooth whitening was examined in 115 volunteers over a period of 12 weeks. The facial surfaces of 12 anterior teeth were assessed for stain using a published, modified version of a standard stain index. Whiteness was measured on teeth 8 and 9 using a single Vita Lumin-Vaccum Shade Guide for consistency. At baseline, the mean facial stain scores were significantly higher (p < 0.05-0.01) for both Arm & Hammer dentifrices than for Crest. In addition, the tooth shades, as indicated by the stain guide, specifically the b* values representing yellowness, were quantified using a Minolta spectrophotometer. Arm & Hammer Dental Care Extra Whitening formula was found to be significantly better than Crest at removing naturally occurring extrinsic stain. The difference between Arm & Hammer Dental Care Extra Whitening and Crest became significant (p < 0.01) after two weeks of use, and remained intact during the balance of the study, achieving p values of 0.0002 for at least one of the three assessed parameters (total stain, proximal, and facial) at weeks 4 and 12. The study also found that Arm & Hammer Dental Care produced a significant increase in tooth whiteness by week 12, whereas Crest showed no such increase at any time during the study. These results suggest that the two Arm & Hammer Baking Soda products are more effective in reducing stain and increasing whiteness than the standard silica based dentifrice. Their effectiveness is not related to abrasivity since they are less abrasive to tooth enamel than the silica-based product tested. PMID- 10518866 TI - Laboratory assessment of tooth whitening by sodium bicarbonate dentifrices. AB - The increasing emphasis on dental aesthetics has made tooth whitening an important function of a dentifrice. This laboratory study investigated the whitening effect of toothbrushing with sodium bicarbonate-based dentifrices compared with standard dentifrices that do not contain sodium bicarbonate. Six dentifrices and a distilled water control were tested for their ability to whiten teeth with intrinsic stain. The dentifrices contained different abrasive systems: (1) 45% NaHCO3; (2) 65% NaHCO3 (Arm and Hammer Dental Care); (3) 94% NaHCO3; (4) 94% NaHCO3 + 1.5% H2O2; (5) silica (Crest Regular Toothpaste); and (6) dicalcium phosphate (Colgate Regular Toothpaste). After a thorough rubber cup cleaning with a pumice slurry, the intrinsic color of the test teeth with a Vita shade of A3 or darker was measured with a spectrophotometer using the standard L*a*b* color scale. Measurements were taken on a total of 12 teeth per test dentifrice before treatment, and after 30, 60, 90 and 120 minutes of mechanical toothbrushing. No changes in L* (lightness/brightness) or a* (red-green hue) occurred, but significant differences in b* (yellow-blue hue) were observed over time. Compared to baseline tooth color, all four sodium bicarbonate-based dentifrices were significantly effective in removing the yellow intrinsic tooth stain, while the water control, silica dentifrice, and dicalcium phosphate dentifrice demonstrated no significant change. Between-group comparisons showed that the four dentifrices containing sodium bicarbonate were significantly more effective than the water and dicalcium phosphate dentifrice groups. The commercial dentifrice containing 65% sodium bicarbonate was also more effective than the commercial silica dentifrice. Although continued toothbrushing from 30 to 120 minutes resulted in additional stain removal, most of the tooth whitening by the sodium bicarbonate based dentifrices occurred in the first 30 minutes of brushing. In the studies conducted, dentifrices containing high concentrations of sodium bicarbonate were more effective at removing intrinsic tooth stain than dentifrices that do not contain sodium bicarbonate. PMID- 10518867 TI - The effects of dentifrice systems on oral malodor. AB - Chronic oral malodor is a serious concern for about one-fifth of the North American population, and a field of emerging research interest. The present three studies, one involving gas chromatography and two employing odor judge assessment, examined the efficacy of baking soda and other toothpastes in reducing breath odor. The most common cause of oral malodor is elevated levels of volatile sulfur compounds (VSC's), primarily hydrogen sulfide (H2S) and methyl mercaptan (CH3SH), in the breath. Gas chromatography, an accurate means of measuring breath VSC, was employed to evaluate the breath levels of VSC in 11 men after brushing with baking soda-containing dentifrices with or without the addition of Zn++. Dentifrices with either Zn++ or a concentration of baking soda 20% or greater significantly reduced VSC levels. The addition of Zn++ to baking soda dentifrices enhanced the anti-odor effects. In the first organoleptic study, dentifrices containing 20% baking soda and 30% baking soda demonstrated significantly greater ability to reduce breath odor than a standard sodium fluoride/silica dentifrice. The subjects' baseline mouth odor evaluations, initially rated as strong, declined after brushing with the baking soda toothpastes to a barely detectable level at one hour, then rising to a faint level at two hours and moderate levels at three hours. In the second organoleptic study, a dentifrice containing 65% baking soda demonstrated significantly greater ability to reduce breath odor than a standard sodium fluoride/silica tartar control dentifrice, but did not differ significantly from a standard dentifrice containing 0.76% sodium monofluorophosphate in a dicalcium phosphate dihydrate base. The results of these studies indicate that dentifrices containing 20% or more baking soda can confer a significant odor-reducing benefit for time periods up to three hours. PMID- 10518868 TI - Evaluation of the clinical performance and effectiveness of adhesively-bonded metal crowns on damaged canine teeth of working dogs over a two- to 52-month period. AB - In this clinical study, 41 metal full crown restorations of canine teeth were placed in 18 working dogs. Twenty-six canine teeth had severe attrition with no involvement of the pulp cavities; 15 fractured canine teeth were endodontically treated. With the exception of one tooth, at least one-third of the coronal part of each canine tooth was available for a supragingivally performed, minimal tooth crown preparation. A dental resin luting cement technique was used to bond the electrolytically etched crown (made from an alloy of cobalt-chrome-molybdenum) to the tooth. The metal crowns were slightly shorter and with a more rounded tip than the original tooth. Posts or post-and-core techniques were not used. Median follow-up period was 30 months (range 2 to 61 months), at which time 36 crowns were found to be intact and functional. Five crowns were lost; three as a result of subsequent injury and fracture of the tooth below the crown; one as a result of use of less than one-third of the coronal portion of the tooth for retention of the crown; and one as a result of an oblique fracture of the root. PMID- 10518869 TI - Endodontic treatment including apexification in a chow chow with a necrotic immature mandibular canine tooth. AB - A necrotic immature mandibular, canine tooth in a two year-old, male, intact Chow Chow was endodontically treated. This tooth had an open apex, wide root canal, thin dentinal walls, and there was periapical bone resorption. An apexification procedure was used to induce apical closure by calcified tissue formation, with resolution of the periapical inflammation. PMID- 10518870 TI - Dental health status and endodontic treatment of captive brown bears (Ursus arctos ssp.) living in the Bernese bear pit. AB - The teeth of five adult captive brown bears (Ursus arctos ssp.) were examined and radiographed for occlusion, loss of teeth, dental plaque and calculus, and attrition under general anesthesia. Deposits of dental calculus were found in various locations with an overall prevalence of 8% to 15% of all tooth surfaces. In all five animals, severe enamel and dentinal attrition defects were observed in canine teeth with exposed pulp. Cage-chewing behavior is probably the main cause for the dental attrition. The composition of the food and feeding management are most likely responsible for the lack of natural cleaning and the resulting plaque and calculus formation. All affected canine teeth were treated with endodontic procedures using several materials and techniques, and evaluated one- to two-and-a-half years later by clinical examination, radiography, and scanning electron microscopy of silicone casts of the treated teeth. All coronal fillings were tight. The apices were not completely sealed in two teeth. We conclude that the use of adequate and specialized instrumentation and techniques for the treatment of these long, curved, large diameter root canals is more important than the particular endodontic and restorative materials used. The dental health status of zoo animals is an indicator of their general well-being. Preventive measures should be taken in their environment and management to minimize the risk of dental conditions. PMID- 10518871 TI - Feline "odontolysis" in the 1920's: the forgotten histopathological study of feline odontoclastic resorptive lesions (FORL). AB - This article reviews an historic description of feline odontoclastic resorptive lesions (FORL) from the 1920's. Hopewell-Smith describes the complete resorption process of feline permanent premolar teeth in remarkable detail, using 14 excellent photomicrographs. Resorptive lesions in cats were seen in the early part of this century, however the prevalence of this condition in domestic cats appears to have increased concurrently with certain aspects of domestication since the 1960's. PMID- 10518872 TI - Oral malodor and its relevance to periodontal disease in the dog. AB - Oral malodor has been studied extensively in humans but very little work has been done in dogs where it constitutes a significant problem. In this article we review its causes, methods of detection, and strategies for preventing it. Oral malodor arises from microbial metabolism of exogenous and endogenous proteinaceous substrates in the oral cavity and is exacerbated by periodontal disease and poor oral hygiene. Gram negative bacteria found in plaque, in periodontal pockets, and on the dorsum of the tongue are primarily responsible for odor production. The volatile sulfur compounds (VSCs) hydrogen sulfide and methyl mercaptan, which are produced by these bacteria, are not only primarily responsible for the objectionable odor but have been implicated in the pathogenesis of periodontal disease. Assessment of malodor by portable sulfide monitors correlates well with organoleptic measurements. Reduction of microbial load in the oral cavity due to good oral hygiene practices (such as tooth brushing) or by the use of appropriate diets or chews may reduce malodor formation. PMID- 10518873 TI - Studies of oral malodor in the dog. AB - We compared currently used methods for assessing oral malodor and found a significant, positive correlation between the concentration of volatile sulfur compounds (VSCs) in breath and the human perception of malodor, but only within a limited range of VSC production. Daily tooth-brushing from an early age maintains a low and consistent level of oral malodor that does not differ significantly throughout the day. In contrast, dogs with elevated concentrations of VSC production suffer from the highest level of malodor early in the morning, prior to feeding and daily activities. Even in older dogs, periodontal therapy can result in a substantial reduction of oral malodor that remains significantly below pre-treatment values for up to 3 months after treatment. PMID- 10518874 TI - Treatment of asymptomatic internal resorption of a maxillary premolar tooth in a military working dog. AB - An asymptomatic pink discoloration of a maxillary right fourth premolar tooth was discovered during a routine oral examination on a 9 year-old Belgian Malinois dog. A radiolucent lesion was seen in the pulpal chamber on radiographic examination. The lesion had perforated the mesiobuccal root of the tooth. The primary differential diagnosis was idiopathic internal resorption. The tooth was treated by partial resection (removal of the mesiobuccal root and associated crown). A vital pulpotomy and amalgam restoration was performed on the remaining tooth structure. A follow-up 1 year later demonstrated a successful treatment outcome. The animal was asymptomatic and able to perform military duties. Clinical and radiographic signs of healing were evident and the tooth was functional. PMID- 10518875 TI - Use of maxillary and mandibular splints for restoration of normal occlusion following jaw trauma in a cat: a case report. AB - A compression fracture of the maxilla in a young cat was treated by splinting the maxillary canine teeth. Wire-reinforced composite resin was used to keep these teeth from tipping palatally. This technique stabilized the fracture with anatomic reduction and minimal invasion of the tissues. A mandibular symphyseal separation was also stabilized with a cerclage wire and composite resin bonded wire fixation of the mandibular canine teeth. Two years later, occlusion was normal. PMID- 10518876 TI - Enamel decalcification in orthodontics: a survey of Canadian orthodontists. PMID- 10518877 TI - The engaging concept of self-ligation. PMID- 10518878 TI - Endodontic diagnosis. PMID- 10518879 TI - Combined orthodontic, periodontic and endodontic treatment of a fractured maxillary premolar. PMID- 10518880 TI - Giving back to the community and beyond. Dentists making a difference through their volunteer efforts. PMID- 10518881 TI - Entering a new era: periodontics today and tomorrow. PMID- 10518882 TI - The incidence of pulpal complications and loss of vitality subsequent to full crown restorations. PMID- 10518883 TI - Guided tissue regeneration. PMID- 10518884 TI - Subepithelial connective tissue grafts for root coverage. PMID- 10518885 TI - Letters of introduction when selling a practice. PMID- 10518886 TI - Periodontal disease as a risk factor for cardiovascular disease and myocardial infarction. PMID- 10518887 TI - The relationship of palato-gingival grooves and cervical enamel projections to localized periodontal disease. A review of the literature. PMID- 10518888 TI - "Refer" is not a four letter word. PMID- 10518889 TI - Repetitive strain injuries in dentistry. AB - In order to avoid RSI, try to neutralize, modify and energize at work, home and play. These concepts eliminate the risk factors associated with the development of RSI and optimize an individual's physical capabilities to work productively and efficiently. People suffering with an RSI should consult their physician regarding the various treatment options available to them. Ideally, prevention is the key to reducing the costs and problems associated with RSI. However, an individual suffering with RSI should not delay addressing the injury. Treatment is most effective when applied as soon as possible after the onset of injury. PMID- 10518890 TI - Caries diagnosis with the DIAGNOdent laser: a user's product evaluation. AB - In the study, the DIAGNOdent has shown itself to be very accurate in the diagnosis of pit and fissure lesions. In the clinical situation, it showed a 92.1 per cent accuracy in diagnosing lesions, as well as their severity. In addition, when the unit showed a reading of less than 30, it was 100 per cent accurate in the virgin teeth. When the unit indicated that there was no caries in extracted teeth, it was accurate 98.1 per cent of the time. Now, for the first time, dentists can do research as to the speed of progression of caries, as well as what percentage of caries becomes arrested and what percentage deteriorate. PMID- 10518891 TI - An evolution in progress. The integration of social services and public health dental programs. PMID- 10518892 TI - Designing dental programs for those in need. PMID- 10518893 TI - Be prepared--another reason for dental practice appraisals. PMID- 10518894 TI - Keeping smiles on the faces of Toronto's children. PMID- 10518895 TI - Identifying child abuse. PMID- 10518897 TI - Case studies in dental practice computing. PMID- 10518896 TI - A systematic overview of the relationship between infant feeding caries and breast-feeding. PMID- 10518898 TI - Chart count--how many patients do you have? PMID- 10518899 TI - Objective treatment planning. PMID- 10518900 TI - Management considerations for the pregnant or nursing emergency patient. AB - It is important to realize the risk-benefit ratio in all therapeutic modalities in any patient and that these variables vary from patient to patient as do circumstances. The emergency patient who is pregnant or nursing should not be feared or be denied appropriate treatment. It is always best to err on the side of safety and remain conservative. The dental problem can usually be treated with the use of adequate local anaesthesia and supplemental nitrous oxide in the second or third trimester. Nevertheless, situations arise that may warrant consultation with the patient's physician or specialist. This brief overview should refamiliarize the dental practitioner with the medications available to treat the gravid patient. It is important to have an understanding of our common therapeutics as well as the inherent treatment risks and benefits as they relate to the changing maternal-fetal physiology. PMID- 10518901 TI - The top-40 prescription drugs in Canada (1997). PMID- 10518902 TI - Indications for dental anaesthesia. PMID- 10518903 TI - The wand: a user's product review. PMID- 10518904 TI - The prevalence of oral pathoses in a private dental practice: a 30 month survey. AB - The purpose of this study was to determine the prevalence of oral lesions in a private dental practice, which treats a broad spectrum of patients from both urban and rural areas. All oral lesions were documented over a 30 month period in a research programme, which formed part of the normal patient treatment plan. The lesions were analysed according to the prevalence of the lesions, race, age and gender distribution, and the ratio of lesions diagnosed by the practitioner compared with those presented for treatment by the patients. A total of 8418 patients were examined and 151 pathological lesions were diagnosed, representing 46 different conditions. The commonest conditions were fluorosis (11 per cent of lesions) and geographic tongue (11 per cent of lesions seen). Twenty conditions (47.8 per cent) were seen only once. The male:female ratio was 1.17:1. A total of 74 per cent of the lesions were seen in patients between the 2nd and the 5th decades and 25 per cent in the 4th decade. Patients seeking treatment suffered from conditions such as fluorosis causing aesthetic problems. On the other hand premalignant lesions such as leukoplakia and life threatening diseases were usually detected by the dentist, which stresses the importance of a thorough clinical examination of all patients. Fluorosis documented in 17 patients in this study, led to analysis of drinking water in the Dennilton area, which was found to contain toxic levels of fluoride. PMID- 10518905 TI - South African oral hygienists: their profile and perception of their profession and career. AB - The aim of the study was to determine a profile of the oral hygienists in South Africa, their views on the profession, work-place, and the practice of their career, which aspects of the work they enjoy and which are not enjoyable, their opinions on expanding duties for hygienists and which duties should be included, and their perceptions about the status and importance of oral hygienists vis-a vis other health providers. A questionnaire was sent to every third registered oral hygienist and 47 per cent responded. The majority who responded were in the age group 20 to 39 years, had been in practice for less than 15 years, were married, qualified at the Universities of Pretoria and Stellenbosch, and were employed in traditional practice. The larger proportion worked individually and practised from six to eight hours per day. They were happy with the training they received, believed their job was worthwhile, were satisfied with their careers and enjoyed a cordial relationship with dentists. Motivating, educating, assisting patients and communicating with people were the most enjoyable aspects of practice while procedures associated with the treatment of gingivitis and periodontitis and the poor response of patients to treatment were the least enjoyable. The majority preferred expanded duties for hygienists which should include elementary dentistry, local anaesthesia, minor extractions and emergency treatment and they also desired greater independence. Seventy-eight per cent felt that the public does not know what oral hygiene is. The status and importance of the profession were rated comparable to that of physiotherapists, qualified nurses, radiographers and dental therapists but significantly higher than dental assistants. PMID- 10518906 TI - National oral health survey Zimbabwe 1995: quality of restorations. AB - In 1995, a second national oral health survey was carried out, ten years after the first. Application of a multi-stage sampling procedure resulted in 3709 persons being examined. The restorations were assessed using the criteria described by Kroeze et al (1990). Only ditches on the tooth/restoration margins that were wider than 0.4 mm were considered to be carious. The background variables studied were age, gender, type of location, socioeconomic status (SES) and level of education. The prevalence of restorations in all persons examined was 3.4 per cent. Restorations were found much more often among urban (95.5 per cent) than rural people (4.5 per cent) and also among those living in high (75 per cent) compared to low SES suburbs (25 per cent). Amalgam was more often used (89 per cent) than composite resin (10 per cent). The most frequently observed type of restoration was Class I (45 per cent) followed by Class II (39 per cent) and Class III (7 per cent). The prevalence of satisfactory restorations was 83.9 per cent. Failures were due to 'fractured restorations' (6.3 per cent), 'caries at the margin' and 'breakdown of restoration margin', both 4 per cent. Amongst adults, multiple-surface amalgam restorations failed more often than single surface ones. It is concluded that the prevalence of restorations found was very low. There is a need to extend the provision of preventive and restorative oral health care by a more equitable distribution of oral health personnel and by making more finance available to rural and low-SES suburban areas. PMID- 10518907 TI - The role of the community pharmacist as an oral health adviser--an exploratory study of community pharmacists in Johannesburg, South Africa. AB - The aim of this study was to examine to what extent community pharmacists are currently acting as dental advisers, and to determine whether they would be willing to extend this role in the future. The data were collected by means of a random survey of community pharmacists in Johannesburg, and included: incidence and nature of dental enquiries; dental knowledge with respect to preventive measures; willingness to engage in promotion of oral health and prevention of oral disease. The results reveal an anomaly. On one hand community pharmacists seemed to be handling a substantial amount and range of inquiries related to oral health and dental disease, and were willing to fulfil a meaningful role as oral health advisers. On the other hand, however, they have received very little education on these matters and seemed to operate in isolation from dentists and other health professionals, which raises doubts about their ability to play a meaningful role in primary health care. Nevertheless, it is quite feasible for community pharmacists to play the role as envisaged, provided additional training and networking with dentists, oral hygienists, dental therapists and other relevant health professionals takes place. PMID- 10518908 TI - Lingual piercing. PMID- 10518909 TI - Oral health education provided by oral hygienists in private practice. AB - The purpose of this study was to identify the health education services provided by oral hygienists working in private dental practices. A questionnaire was designed, pilot tested and posted to all hygienists registered with the SAMDC. Thirty eight per cent returned the questionnaire. Results indicated that an average of only 6 to 10 minutes was spent on oral hygiene instruction per patient. The reasons given for this were identified as lack of time and resistance of patients to change their habits. Education procedures most often performed were the demonstration of brushing and flossing techniques. The least performed procedures included the use of disclosing agents and periodontal measurements. A statistical significant correlation was found between the number of patients consulted per day and salary paid by the dentist. Hygienists who earn more treat more patients per day, but do not spend less time on providing oral hygiene information. Patient resistance is an obstacle to oral hygiene instruction and more emphasis should be placed on the psychological approach to behavioural change than on techniques of oral hygiene. More time should be spent on oral health education and follow-up appointments should be scheduled to support patients in maintaining good oral health care. PMID- 10518910 TI - Oral management of irradiated head and neck cancer patients. PMID- 10518911 TI - The prevention of endodontic failures. PMID- 10518912 TI - The effect of post-curing on Vickers Hardness of four light-cure resin composite materials. AB - The use of post-curing ovens to post-cure light-cured resin composite restorations leads to a decrease in the negative effects of polymerization shrinkage and an increase in the hardness and wear resistance of the material. The aim of this study was to compare the Vickers Hardness (VH) of four different light-cure resin composite materials. In one group, samples of the four materials, Z100, Tetric Ceram, F2000 and Heliomolar were light-cured only, and in the other group a similar set of samples were light-cured and also post-cured in a DI 500 post-curing oven. VH vests were then carried out on the samples. All samples in the post-cured group had higher VH values (p = 0.000) than the corresponding samples in the light-cured only group. Z100 and F2000 had, in both groups, significantly higher VH values (p = 0.000) than Tetric Ceram and Heliomolar. PMID- 10518913 TI - Reasons why South African qualified dentists are working in the UK. AB - Various reasons are provided why professional people are leaving or had left the country since 1990. A substantial number of these emigrants were dentists who went to practise their profession in the United Kingdom (UK). The purpose of this study was to determine, by way of a questionnaire, why some South African qualified dentists prefer to practise their profession in the UK. The results showed that the high level of crime and violence in South Africa is the most important reason why South African qualified dentists are leaving or have left the country. Economical considerations, either their own financial position or the general economy of the country also appear to be an important reason why South African qualified dentists are moving abroad. PMID- 10518914 TI - A case of oral syphilis. PMID- 10518915 TI - Dental status of schoolchildren from a rural community in Cameroon. AB - This study assessed the dental health status of children in a rural community in Cameroon. The investigation was carried out in 1996 on a total of 403 pupils aged between 5-17 years. The children were divided into 3 age groups, (I-III): I: 5-8 years (n = 157), II: 9-12 years (n = 191), II: 13-17 years (n = 55). The teeth were examined for dmft/DMFT, CPI, opacities and treatment needs. 22.6 per cent of the pupils were caries free, and the following mean dmft/DMFT-values were obtained: Age group I, dmft: 2.26 + 2.84, DMFT: 0.85 + 1.39. Group II, dmft: 0.60 + 1.35, DMFT: 2.76 + 2.74. Group III, dmft: 0.05 + 0.22; DMFT: 5.12 + 3.04. Opacities or other enamel disorders were observed in 16.9 per cent of the children. 68.7 per cent of them needed invasive dental treatment and 61.3 per cent required restorative treatment only. 7.4 per cent needed extractions and 25.6 per cent required prevention/sealant care as sole measure. 6.6 per cent of the children showed CPI-grade 0. The majority scored CPI-grade 1 (37.7 per cent) and grade 2 (49.9 per cent). The reported data suggest that improved dental health mindedness and availability of oral health care services are required in order to reduce oral health diseases. PMID- 10518917 TI - An erupted compound odontoma. PMID- 10518916 TI - Utilisation of oral health services, oral health needs and oral health status in a peri-urban informal settlement. AB - Interviews were conducted with 294 black residents (155 females and 138 males) of a peri-urban informal settlement in Gauteng to ascertain utilisation of oral health services, oral health needs and oral health status. Only 37 per cent of the sample had consulted a dentist or medical practitioner, usually for extractions. Teenagers and employed persons were significantly less likely to utilise dentists than the older age groups and unemployed persons. Forty per cent were currently experiencing oral health problems such as a sore mouth, tooth decay and bleeding/painful gums. Two hundred and twelve (73 per cent) interviewees wanted dental treatment or advice. Residents who rated their oral health status as fair or poor appeared to have the greatest need for oral health services. The use of interviews appears to be a cost-effective method of determining oral morbidity. PMID- 10518918 TI - Permeability of lichen planus lesions and healthy buccal mucosa to water. AB - The resurgence of interest in the oral mucosa as a route for drug delivery requires a thorough understanding of the permeability of this tissue in health and disease. Previous work has indicated that non-keratinized oral mucosa is more permeable than its keratinized counterpart. It has been suggested that pathological hyperkeratotic mucosa, which was previously non-keratinized, would be more permeable than healthy tissue. Equivocal results obtained from animal studies in which chemical or mechanical irritation was used to induce a hyperplastic and hyperkeratotic epithelium, prompted us to conduct a study on the comparison of the permeability to water of lichen planus lesions and healthy buccal mucosa. Buccal mucosa was obtained from six patients with previously confirmed lichen planus and from six clinically healthy patients. Thawed biopsies from each specimen were mounted in flow-through diffusion cells and their permeability to tritiated water determined using a continuous flow-through perfusion system. Specimens were examined histologically before and after permeability experiments. No statistically significant differences between mean steady state flux values (10-20 h) for lichen planus tissue and healthy buccal mucosa were found. These results warrant further studies with other oral conditions associated with hyperkeratosis to establish whether the nature and course of the condition are determinants for the retention or loss of the epithelium's permeability characteristics. PMID- 10518919 TI - The culture of human buccal and vaginal epithelial cells for permeability studies. AB - The objective of the present study was to develop a single improved technique to culture human vaginal and buccal epithelial cells, whereby the cultured cells can be used in drug permeability studies. Cells were obtained from healthy human vaginal and buccal mucosa following vaginal hysterectomies and various oral surgical procedures. Tissue obtained was washed extensively in phosphate buffered saline (PBS, pH 7.3) containing antibiotics and amphotericin-B. Tissue specimens were cut into small pieces and plated out in 24-well plates. After drying, the full medium was added. Cell growth occurred within 4-6 days from primary explants and confluency was reached within 2-3 weeks. Primary explants yielded epithelial cells with minimal fibroblast contamination. After trypsinization, cells were seeded into collagen-coated wells and onto Transwell membranes. Trypsinized cells grew best on collagen-coated surfaces yielding more than one layer. The average steady state flux for the collagen-coated membranes (ccm's) containing either buccal or vaginal epithelial layers towards water was 6-10 times lower than that found for the cell-free ccm's. Fluxes for cultured cells on ccm's were 3x higher than those obtained for intact buccal and vaginal mucosa. Growth and permeability to water of the vaginal and buccal epithelial cells were comparable, confirming the similarity of these two tissues and the suitability of using the former as a model for the latter in permeability to tritiated water. PMID- 10518920 TI - Effect of temperature on permeability of mucosa to water. AB - In previous studies it has been demonstrated that frozen human vaginal mucosa can be used as a model of buccal mucosa for in vitro permeability studies on a variety of chemical compounds, including drugs. However, most of the latter studies have, for the sake of convenience, been conducted at room temperature (+/ 20 degrees C). The objective of the present study was to determine the effects of increased temperature on steady state flux rates of water through vaginal mucosa. Specimens of clinically healthy human vaginal mucosa were obtained from excess tissue removed during a vaginal hysterectomy from a single patient, snap frozen in liquid nitrogen at -85 degrees C and banked for 8 months. After thawing in PBS buffer, seven sections from the vaginal mucosa were mounted in flow through diffusion cells (exposed area 0.039 cm2) and their permeability to tritiated water determined using a continuous flow-through perfusion system at temperatures of 25 degrees, 30 degrees and 37 degrees C. Permeability experiments were performed in triplicate at each temperature setting. Specimens were examined histologically before and after permeability experiments. Mean water flux rates at steady state (16-24 h) were found to be 1760 +/- 22 SEM, 2623 +/- 63 SEM and 4155 +/- 70 SEM cpm. cm-2.min-1, at temperatures of 25 degrees, 30 degrees and 37 degrees C, respectively. A linear regression analysis and plot (r2 = 0.99) displayed a slope of 200 +/- 13 SEM cpm. cm-2.min-1/degree C. The results of this study clearly demonstrated the temperature-dependency of flux rates of water across vaginal mucosa, and this should be taken into account whenever the in vitro vaginal/buccal model is used at room temperature for predicting in vivo buccal drug absorption kinetics. PMID- 10518921 TI - The suitability of five liquid scintillation cocktails for use with a continuous flow-through mucosal perfusion system. AB - Continuous flow-through perfusion systems can be successfully used to study the permeability of small mucosal samples to various permeants. The latter often need to be radiolabelled to enable detection by means of liquid scintillation counting. However, the efficacy of the latter is dependent on the liquid scintillation cocktail used. The present study was conducted to evaluate five liquid scintillation cocktails commercially available in South Africa. Although four out of the five cocktails had similar efficacies in the concentration range 0.05 to 50 nCi, this was not the case in the low range (0.01 to 0.5 nCi). Overall Ready Protein+ was the most efficacious and has the added advantage that this cocktail has protein solubilising properties. The latter is important when we extend our permeability studies to include small proteins. PMID- 10518922 TI - Characterization of purified and unidirectionally reconstituted Pho84 phosphate permease of Saccharomyces cerevisiae. AB - Hydropathy analysis of the amino acid sequence of the Pho84 phosphate permease of Saccharomyces cerevisiae suggests that the protein consists of 12 transmembrane domains connected by hydrophilic loops. The Pho84 protein has been modified by a gene fusion approach, yielding two different N-terminal His-tagged chimeras which can be expressed in Escherichia coli, purified and functionally reconstituted into defined proteoliposomes. The continuous epitopes in the N- and C-terminal sequences of the Pho84 chimeras were shown to be accessible in proteoliposomes containing the purified active Pho84 proteins. Site-specific proteolysis of the immunoreactive N-terminal sequence in the reconstituted protein suggests a unidirectional insertion into liposomes. PMID- 10518923 TI - Structure based development of novel specific inhibitors for cathepsin L and cathepsin S in vitro and in vivo. AB - Specific inhibitors for cathepsin L and cathepsin S have been developed with the help of computer-graphic modeling based on the stereo-structure. The common fragment, N-(L-trans-carbamoyloxyrane-2-carbonyl)-phenylalanine-dimethyla mide, is required for specific inhibition of cathepsin L. Seven novel inhibitors of the cathepsin L inhibitor Katunuma (CLIK) specifically inhibited cathepsin L at a concentration of 10(-7) M in vitro, while almost no inhibition of cathepsins B, C, S and K was observed. Four of the CLIKs are stable, and showed highly selective inhibition for hepatic cathepsin L in vivo. One of the CLIK inhibitors contains an aldehyde group, and specifically inhibits cathepsin S at 10(-7) M in vitro. PMID- 10518924 TI - Functional phage display of leech-derived tryptase inhibitor (LDTI): construction of a library and selection of thrombin inhibitors. AB - The recombinant phage antibody system pCANTAB 5E has been used to display functionally active leech-derived tryptase inhibitor (LDTI) on the tip of the filamentous M13 phage. A limited combinatorial library of 5.2 x 10(4) mutants was created with a synthetic LDTI gene, using a degenerated oligonucleotide and the pCANTAB 5E phagemid. The mutations were restricted to the P1-P4' positions of the reactive site. Fusion phages and appropriate host strains containing the phagemids were selected after binding to thrombin and DNA sequencing. The variants LDTI-2T (K8R, I9V, S10, K11W, P12A), LDTI-5T (K8R, I9V, S10, K11S, P12L) and LDTI-10T (K8R, I9L, S10, K11D, P12I) were produced with a Saccharomyces cerevisiae expression system. The new inhibitors, LDTI-2T and -5T, prolong the blood clotting time, inhibit thrombin (Ki 302 nM and 28 nM) and trypsin (Ki 6.4 nM and 2.1 nM) but not factor Xa, plasma kallikrein or neutrophil elastase. The variant LDTI-10T binds to thrombin but does not inhibit it. The relevant reactive site sequences of the thrombin inhibiting variants showed a strong preference for arginine in position P1 (K8R) and for valine in P1' (I9V). The data indicate further that LDTI-5T might be a model candidate for generation of active-site directed thrombin inhibitors and that LDTI in general may be useful to generate specific inhibitors suitable for a better understanding of enzyme-inhibitor interactions. PMID- 10518925 TI - cDNA cloning, characterization and stable expression of novel human brain carboxylesterase. AB - The DNA sequence encoding a novel human brain carboxylesterase (CES) has been determined. The protein is predicted to have 567 amino acids, including conserved motifs, such as GESAGG, GXXXXEFG, and GDHGD which comprise a catalytic triad, and the endoplasmic reticulum retention motif (HXEL-COOH) observed in CES families. Their gene products exhibited hydrolase activity towards temocapril, p nitrophenyl-acetate and long-chain acyl-CoA. Since the molecular masses of these gene products are similar to those that exist in capillary endothelial cells of the human brain [Yamamda et al. (1994) Brain Res. 658, 163-167], these CES isozymes may function as a blood-brain barrier to protect the central nervous system from ester or amide compounds. PMID- 10518926 TI - Production of a recombinant chitin deacetylase in the culture medium of Escherichia coli cells. AB - With the aid of a signal sequence of a chitinase from Streptomyces lividans, a recombinant chitin deacetylase, whose gene originated from a Deuteromycete, Colletotrichum lindemuthianum, was produced in the culture medium of Escherichia coli cells, existing as a highly active form without the signal peptide. During the production of the recombinant chitin deacetylase, both a slight increase in the value of OD600 nm in the culture medium and a drastic decrease in viable cell number were observed. When penta-N-acetyl-chitopentaose was used as the substrate, the recombinant chitin deacetylase had comparable kinetic parameters to those of the original enzyme from the fungus. The addition of a C-terminal six histidine sequence to the recombinant enzyme caused a slight decrease in the kcat value, and the further addition of a 12 amino acid sequence at its N-terminus caused a further decrease in the value. This production system allowed us to easily produce in the culture media the recombinant chitin deacetylases possessing as good properties as the original enzyme, without any disruption steps of the E. coli cells. PMID- 10518927 TI - A 6 bp Z-DNA hairpin binds two Z alpha domains from the human RNA editing enzyme ADAR1. AB - The Z alpha domain of the human RNA editing enzyme double-stranded RNA deaminase I (ADAR1) binds to left-handed Z-DNA with high affinity. We found by analytical ultracentrifugation and CD spectroscopy that two Z alpha domains bind to one d(CG)3T4(CG)3 hairpin which contains a stem of six base pairs in the Z-DNA conformation. Both wild-type Z alpha and a C125S mutant show a mean dissociation constant of 30 nM as measured by surface plasmon resonance and analytical ultracentrifugation. Our data suggest that short (> or = 6 bp) segments of Z-DNA within a gene are able to recruit two ADAR1 enzymes to that particular site. PMID- 10518928 TI - Topological analysis of an RND family transporter, MexD of Pseudomonas aeruginosa. AB - The membrane topology of a resistance-nodulation-division (RND) family transporter, MexD of Pseudomonas aeruginosa, was determined. Although it had been predicted previously that most RND proteins contain 12 transmembrane helices, three independent computer programs used in the present study predicted that MexD possessed 11, 14 or 17 transmembrane segments. To investigate the topology of MexD more thoroughly, 25 MexD-PhoA (alkaline phosphatase) and 18 MexD-Bla (beta lactamase) fusion plasmids were constructed and analyzed. The resulting topological model had just 12 transmembrane helices and two periplasmic loops of about 300 residues between helices 1 and 2 and helices 7 and 8. It is therefore proposed that the N- and C-termini are located in the cytoplasm and the predicted orientation is consistent with the 'positive-inside rule'. This topological model can be applied to other RND proteins. PMID- 10518929 TI - Molecular cloning of mXCR1, the murine SCM-1/lymphotactin receptor. AB - Single C motif-1 (SCM-1)/lymphotactin is a C-type member of the chemokine superfamily. Previously, we identified its specific receptor XCR1. Here we isolated the murine homologue of XCR1 (mXCR1). To demonstrate its biological activity, we produced recombinant mouse SCM-1 by the baculovirus expression system. B300-19 murine pre-B cells expressing mXCR1 responded to mSCM-1 in chemotactic and calcium-mobilization assays. mXCR1 mRNA was weakly expressed in spleen and lung of normal C57BL/6 mice. In spleen, CD8+ cells and NK1.1+ cells were found to express mXCR1. Identification of mXCR1 will now allow us to study the role of this unique cytokine system in the mouse models of inflammation and immunity. PMID- 10518930 TI - Calcium-induced calcium release mediated by a voltage-activated cation channel in vacuolar vesicles from red beet. AB - Little is known about the mechanisms underlying calcium-induced Ca2+ release (CICR) in plants. The slow-activating vacuolar (SV) channel is both permeable to, and activated by Ca2+, and is therefore a prime candidate for a role in CICR. Cytosol-side-out vacuolar membrane vesicles loaded with 45Ca2+ showed voltage- and Ca(2+)-dependent Ca2+ release, which was sensitive to the SV channel modulators DIDS, protein phosphatase 2B and calmodulin. Significantly, voltage dependent Ca2+ release strongly depended on cytoplasmic Ca2+ concentrations. The results support the notion that CICR occurs in plant cells and that the process can be catalysed by the SV channel on the vacuolar membrane. PMID- 10518931 TI - Role of histone acetylation and DNA methylation in the maintenance of the imprinted expression of the H19 and Igf2 genes. AB - H19 and Igf2 are linked and reciprocally imprinted genes. We demonstrate that the histones associated with the paternally inherited and unexpressed H19 allele are less acetylated than those associated with the maternal expressed allele. Cell growth in the presence of inhibitors of either histone deacetylase or DNA methylation activated the silent Igf2 allele, whereas derepression of the silent H19 allele required combined inhibition of DNA methylation and histone deacetylation. Our results indicate that histone acetylation as well as DNA methylation contribute to the somatic maintenance of H19 and Igf2 imprinting and that silencing of the imprinted alleles of these two genes is maintained via distinct mechanisms. PMID- 10518932 TI - The nascent polypeptide-associated complex (NAC) of yeast functions in the targeting process of ribosomes to the ER membrane. AB - We study here the binding of ribosomes to the endoplasmic reticulum (ER) membrane and its dependence on nascent polypeptide-associated complex (NAC). For this, we use an in vitro translation system in combination with isolated microsomes. Importantly, all components in the system are derived from a single source, Saccharomyces cerevisiae. Ribosome nascent chains (RNCs) of the two naturally occurring invertase species (secreted or cytosolic) were prepared in wild-type, delta alpha NAC or delta alpha beta 1 beta 3 NAC translation lysates and tested for binding to the corresponding microsomal membranes. We provide evidence that NAC prevents binding of RNCs without a signal sequence to yeast membranes. In the absence of NAC, signal-less RNCs are able to bind to ER membranes. However, following puromycin treatment, only very few nascent chains translocate into the lumen, as detected by glycosylation. PMID- 10518933 TI - The delta subunit of rod specific cyclic GMP phosphodiesterase, PDE delta, interacts with the Arf-like protein Arl3 in a GTP specific manner. AB - Recently, we have shown that the delta subunit of the cGMP phosphodiesterase (PDE delta) interacts with the retinitis pigmentosa guanine regulator (RPGR). Here, using the two-hybrid system, we identify a member of the Arf-like protein family of Ras-related GTP-binding proteins, Arl3, that interacts with PDE delta. The interaction was verified by fluorescence spectroscopy and co-immunoprecipitation. Arl3 features an unusually low affinity for guanine nucleotides, with a KD of 24 nM for GDP and 48 microM for GTP. Fluorescence spectroscopy shows that PDE delta binds and specifically stabilizes the GTP-bound form of Arl3 by strongly decreasing the dissociation rate of GTP. Thus, PDE delta is an effector of Arl3 and could provide a novel nucleotide exchange mechanism by which PDE delta stabilizes Arl3 in its active GTP-bound form. PMID- 10518934 TI - Differential interaction patterns in binding assays between recombinant syntaxin 1 and synaptobrevin isoforms. AB - Syntaxin 1 and synaptobrevin play an essential role in synaptic vesicle exocytosis. Two isoforms for each of these proteins, syntaxin 1A and 1B and synaptobrevin 1 and 2, have been found in nerve endings. Previous morphological studies have revealed a characteristic co-localization of syntaxin 1 and synaptobrevin isoforms in nervous and endocrine systems; however, the physiological significance of differential distribution is not known. In the present study an in vitro assay has been used to study a possible isoform specific interaction between syntaxin and synaptobrevin isoforms. The results show that although both syntaxin 1A and 1B may interact with synaptobrevin 1 and 2, this interaction is not uniform, showing different affinity patterns depending on the syntaxin 1/synaptobrevin isoform combination. The addition of SNAP-25 increased the binding capacity of syntaxin and synaptobrevin isoforms without affecting specific interactions. PMID- 10518935 TI - Neurotoxicity of prion peptide 106-126 not confirmed. AB - Prion-related diseases are accompanied by neurodegeneration, astroglial proliferation and formation of proteinase K-resistant aggregates of the scrapie isoform of the prion protein (PrPSc). The synthetic PrP fragment 106-126 was reported to be neurotoxic towards cultured rat hippocampal neurons (Forloni, G., Angeretti, N., Chiesa, R., Monzani, E., Salmona, M., Bugiani, O. and Tagliavini, F. (1993) Nature 362, 543-546) and mouse cortical cells (Brown, D.R., Herms, J. and Kretzschmar, H.A. (1994) Neuroreport 5, 2057-2060). However, we found the viability of these and other neuronal cell types not to be impaired in the presence of PrP106-126 under widely varied sets of conditions. Aged preparations of the peptide as well as synthetic deamidated and isomerized derivatives that correspond to the aging products of the peptide proved also to lack neurotoxicity. Apparently, PrP106-126 cannot serve as a model for the interaction of PrP with neuronal cells. PMID- 10518936 TI - Strong hydrophobic nature of cysteine residues in proteins. AB - The differences between disulfide-bonding cystine (Cys_SS) and free cysteine (Cys_SH) residues were examined by analyzing the statistical distribution of both types of residue in proteins of known structure. Surprisingly, Cys_SH residues display stronger hydrophobicity than Cys_SS residues. A detailed survey of atoms which come into contact with the sulfhydryl group (sulfur atom) of Cys_SH revealed those atoms are essentially the same in number and variety as those of the methyl group of isoleucine, but are quite different to those of the hydroxyl group of serine. Moreover, the relationships among amino acids were also determined using the 3D-profile table of known protein structures. Cys_SH was located in the hydrophobic cluster, along with residues such as Met, Trp and Tyr, and was clearly separated from Ser and Thr in the polar cluster. These results imply that free cysteines behave as strongly hydrophobic, and not hydrophilic, residues in proteins. PMID- 10518937 TI - Kluyveromyces lactis HIS4 transcriptional regulation: similarities and differences to Saccharomyces cerevisiae HIS4 gene. AB - Sequence analysis of the Kluyveromyces lactis HIS4 (KlHIS4) gene promoter reveals relevant differences in comparison to the Saccharomyces cerevisiae HIS4 homologous gene. Among them are the absence of a Rap1 binding site and the presence of only three putative Gcn4 binding consensus sites instead of the five described in the S. cerevisiae promoter. Since these factors are implicated in the general control, we investigated the transcriptional regulation of the KlHIS4 gene under conditions of amino acid starvation and discovered that the mechanisms previously described for S. cerevisiae HIS4 regulation and related to general control are not functional in K. lactis. The expression analysis of the KlHIS4 gene under phosphate starvation or high adenine supply shows that factors, such as Bas1 or Bas2, involved in the basal control may also operate in a different way in K. lactis. Interestingly, and also in contrast to the HIS4 regulation in S. cerevisiae, we found domains for Nit2-like and yeast-Ap1-like binding sequences. Northern analyses showed transcriptional activation under ammonia starvation and oxidative stress. PMID- 10518938 TI - One single mRNA encodes the centrosomal protein CCD41 and the endothelial cell protein C receptor (EPCR). AB - The cDNA encoding the centrosomal protein CCD41 is identical with the cDNA for the endothelial cell protein C receptor. This finding is not due to an artefact, e.g. caused by selection of false positive clones. The segment of the CCD41 cDNA encoding the protein originally termed CCD41 and deletion mutants of it were fused with the nucleotide sequence encoding the enhanced green fluorescent protein (EGFP). Transfection and expression of the full length construct produces a fusion protein mainly located in cell membranes reflecting the receptor-type protein. Deletion mutants, e.g. those where the signal sequence is deleted, result in fusion proteins which are exclusively incorporated into a small perinuclear structure which is the site of the centrosome. This result suggests that post-translational modification, namely deletion of the signal sequence, is decisive for the centrosomal location of the resulting centrosomal protein while the unprocessed protein is incorporated into cell membranes. PMID- 10518939 TI - Molecular architecture of the rod domain of the Dictyostelium gelation factor (ABP120). AB - The Dictyostelium discoideum gelation factor is a two-chain actin-cross-linking protein that, in addition to an N-terminal actin-binding domain, has a rod domain constructed from six tandem repeats of a 100-residue motif that has an immunoglobulin fold. To define the architecture of the rod domain of gelation factor, we have expressed in E. coli a series of constructs corresponding to different numbers of gelation factor rod repeats and have characterised them by chemical crosslinking, ultracentrifugation, column chromatography, matrix assisted laser desorption ionisation (MALDI) mass spectrometry and NMR spectroscopy. Fragments corresponding to repeats 1-6 and 5-6 dimerise, whereas repeats 1-5 and single repeats 3 and 4 are monomeric. Repeat 6 interacts weakly and was present as monomer and dimer when analysed by analytical ultracentrifugation. Proteolytic digestion of rod5-6 resulted in the generation of two polypeptides that roughly corresponded to rod5 and part of rod6. None of these polypeptides formed dimers after chemical crosslinking. Stable dimerisation therefore appears to require repeats 5 and 6. Based on these data a model of gelation factor architecture is presented. We suggest an arrangement of the chains where only the carboxy-terminal repeats interact as was observed for filamin/ABP280, the mammalian homologue of gelation factor. PMID- 10518940 TI - Effect of antibiotics on large ribosomal subunit assembly reveals possible function of 5 S rRNA. AB - Functional large ribosomal subunits of Thermus aquaticus can be reconstituted from ribosomal proteins and either natural or in vitro transcribed 23 S and 5 S rRNA. Omission of 5 S rRNA during subunit reconstitution results in dramatic decrease of the peptidyl transferase activity of the assembled subunits. However, the presence of some ribosome-targeted antibiotics of the macrolide, ketolide or streptogramin B groups during 50 S subunit reconstitution can partly restore the activity of ribosomal subunits assembled without 5 S rRNA. Among tested antibiotics, macrolide RU69874 was the most active: activity of the subunits assembled in the absence of 5 S rRNA was increased more than 30-fold if antibiotic was present during reconstitution procedure. Activity of the subunits assembled with 5 S rRNA was also slightly stimulated by RU69874, but to a much lesser extent, approximately 1.5-fold. Activity of the native T. aquaticus 50 S subunits incubated in the reconstitution conditions in the presence of RU69874 was, in contrast, slightly decreased. The presence of antibiotics was essential during the last incubation step of the in vitro assembly, indicating that drugs affect one of the last assembly steps. The 5 S rRNA was previously shown to form contacts with segments of domains II and V of 23 S rRNA. All the antibiotics which can functionally compensate for the lack of 5 S rRNA during subunit reconstitution interact simultaneously with the central loop in domain V (which is known to be a component of peptidyl transferase center) and a loop of the helix 35 in domain II of 23 S rRNA. It is proposed that simultaneous interaction of 5 S rRNA or of antibiotics with the two domains of 23 S rRNA is essential for the successful assembly of ribosomal peptidyl transferase center. Consequently, one of the functions of 5 S rRNA in the ribosome can be that of assisting the assembly of ribosomal peptidyl transferase by correctly positioning functionally important segments of domains II and V of 23 S rRNA. PMID- 10518941 TI - Energetics of strand-displacement reactions in triple helices: a spectroscopic study. AB - DNA triple helices offer exciting new perspectives toward oligonucleotide directed inhibition of gene expression. Purine and GT triplexes appear to be the most promising motifs for stable binding under physiological conditions compared to the pyrimidine motif, which forms at relatively low pH. There are, however, very little data available for comparison of the relative stabilities of the different classes of triplexes under identical conditions. We, therefore, designed a model system which allowed us to set up a competition between the oligonucleotides of the purine and pyrimidine motifs targeting the same Watson Crick duplex. Several conclusions may be drawn: (i) a weak hypochromism at 260 nm is associated with purine triplex formation; (ii) delta H degree of GA, GT and TC triplex formation (at pH 7.0) was calculated as -0.1, -2.5 and -6.1 kcal/mol per base triplet, respectively. This unexpectedly low delta H degree for the purine triple helix formation implies that its delta G degree is nearly temperature independent and it explains why these triplexes may still be observed at high temperatures. In contrast, the pyrimidine triplex is strongly favoured at lower temperatures; (iii) as a consequence, in a system where two third-strands compete for triplex formation, displacement of the GA or GT strand by a pyrimidine strand may be observed at neutral pH upon lowering the temperature. This original purine to-pyrimidine triplex conversion shows a significant hypochromism at 260 nm and a hyperchromism at 295 nm which is similar to the duplex-to-triplex conversion in the pyrimidine motif. Further evidence for this triplex-to-triplex conversion is provided by mung bean-nuclease foot-printing assay. PMID- 10518942 TI - The solution structure of the domain from MeCP2 that binds to methylated DNA. AB - MeCP2 is an abundant mammalian protein that binds methylated CpG (mCpG) sequences within double-stranded DNA, represses transcription by recruiting histone deacetylases, and is essential for embryonic development. It is one of a family of proteins which mediate the biological consequences of DNA methylation. These proteins each possess a sequence motif of about 70 residues which, in MeCP2, form a domain necessary and sufficient for binding to mCpG. The solution structure of the mCpG-binding domain (MBD) from MeCP2 has been solved and the DNA-binding surface of the domain mapped using NMR spectroscopy. Residues 95-162 of MeCP2 adopt a novel fold forming a wedge-shaped structure. An N-terminal four-stranded antiparallel beta-sheet forms one face of the wedge, while the other face is formed mainly by a C-terminal helical region. The thin end of the wedge is extended by a long loop between beta-strands B and C containing many basic residues. The B-C loop together with residues in strands B, C and D, and at the N terminus of the alpha-helix, appears to form an interface with methylated DNA. Unstructured residues at the NH2 terminus of the domain are also involved in formation of the complex. The presence of numerous arginine and lysine side chains on the DNA-binding surface of MBD is consistent with the requirement for the mCpG site to be flanked by non-specific sequences of base-pairs. The absence of symmetry in the domain implies that recognition does not exploit the symmetry of the binding site. A conserved hydrophobic pocket containing the side-chains of Tyr123 and Ile125 on the positively charged beta-sheet face is a candidate for the region of contact with the methyl-groups of the modified cytosine residues. PMID- 10518943 TI - The binding site for UCH-L3 on ubiquitin: mutagenesis and NMR studies on the complex between ubiquitin and UCH-L3. AB - The ubiquitin fold is a versatile and widely used targeting signal that is added post-translationally to a variety of proteins. Covalent attachment of one or more ubiquitin domains results in localization of the target protein to the proteasome, the nucleus, the cytoskeleton or the endocytotic machinery. Recognition of the ubiquitin domain by a variety of enzymes and receptors is vital to the targeting function of ubiquitin. Several parallel pathways exist and these must be able to distinguish among ubiquitin, several different types of polymeric ubiquitin, and the various ubiquitin-like domains. Here we report the first molecular description of the binding site on ubiquitin for ubiquitin C terminal hydrolase L3 (UCH-L3). The site on ubiquitin was experimentally determined using solution NMR, and site-directed mutagenesis. The site on UCH-L3 was modeled based on X-ray crystallography, multiple sequence alignments, and computer-aided docking. Basic residues located on ubiquitin (K6, K11, R72, and R74) are postulated to contact acidic residues on UCH-L3 (E10, E14, D33, E219). These putative interactions are testable and fully explain the selectivity of ubiquitin domain binding to this enzyme. PMID- 10518944 TI - Accessing the global minimum conformation of stefin A dimer by annealing under partially denaturing conditions. AB - Stefin A folds as a monomer under strongly native conditions. We have observed that under partially denaturing conditions in the temperature range from 74 to 93 degrees C it folds into a dimer, while it is monomeric above the melting temperature of 95 degrees C. Below 74 degrees C the dimer is trapped and it does not dissociate. The dimer is a folded and structured protein as judged by CD and NMR, nevertheless it is no more functional as an inhibitor of cysteine proteases. The monomer-dimer transition proceeds at a slow rate and the activation energy of dimerization at 99 kcal/mol is comparable to the unfolding enthalpy. A large and negative dimerization enthalpy of -111(+/- 8) kcal/mol was calculated from the temperature dependence of the dissociation constant. An irreversible pretransition at 10-15 deg. below the global unfolding temperature has been observed previously by DSC and can now be assigned to the monomer-dimer transition. Backbone resonances of all the dimer residues were assigned using 15N isotopically enriched protein. The dimer is symmetric and the chemical shift differences between the monomer and dimer are localized around the tripartite hydrophobic wedge, which otherwise interacts with cysteine proteases. Hydrogen exchange protection factors of the residues affected by dimer formation are higher in the dimer than in the monomer. The monomer to dimer transition is accompanied by a rapid exchange of all of the amide protons which are protected in the dimer, indicating that the transition state is unfolded to a large extent. Our results demonstrate that the native monomeric state of stefin A is actually metastable but is favored by the kinetics of folding. The substantial energy barrier which separates the monomer from the more stable dimer traps each state under native conditions. PMID- 10518945 TI - Sequence homology and structural analysis of the clostridial neurotoxins. AB - The clostridial neurotoxins (CNTs), comprised of tetanus neurotoxin (TeNT) and the seven serotypes of botulinum neurotoxin (BoNT A-G), specifically bind to neuronal cells and disrupt neurotransmitter release by cleaving proteins involved in synaptic vesicle membrane fusion. In this study, multiple CNT sequences were analyzed within the context of the 1277 residue BoNT/A crystal structure to gain insight into the events of binding, pore formation, translocation, and catalysis that are required for toxicity. A comparison of the TeNT-binding domain structure to that of BoNT/A reveals striking differences in their surface properties. Further, the solvent accessibility of a key tryptophan in the C terminus of the BoNT/A-binding domain refines the location of the ganglioside-binding site. Data collected from a single frozen crystal of BoNT/A are included in this study, revealing slight differences in the binding domain orientation as well as density for a previously unobserved translocation domain loop. This loop and the conservation of charged residues with structural proximity to putative pore forming sequences lend insight into the CNT mechanism of pore formation and translocation. The sequence analysis of the catalytic domain revealed an area near the active-site likely to account for specificity differences between the CNTs. It revealed also a tertiary structure, highly conserved in primary sequence, which seems critical to catalysis but is 30 A from the active-site zinc ion. This observation, along with an analysis of the 54 residue "belt" from the translocation domain are discussed with respect to the mechanism of catalysis. PMID- 10518946 TI - The Cfr10I restriction enzyme is functional as a tetramer. AB - It is thought that most of the type II restriction endonucleases interact with DNA as homodimers. Cfr10I is a typical type II restriction enzyme that recognises the 5'-Pu decreases CCGGPy sequence and cleaves it as indicated by the arrow. Gel filtration and analytical ultracentrifugation data presented here indicate that Cfr10I is a homotetramer in isolation. The only SfiI restriction enzyme that recognises the long interrupted recognition sequence 5'-GGCCNNNNNGGCC has been previously reported to operate as a tetramer however, its structure is unknown. Analysis of Cfr10I crystals revealed that a single molecule in the asymmetric unit is repeated by D2 symmetry to form a tetramer. To determine whether the packing of the Cfr10I in the crystal reflects the quaternary structure of the protein in solution, the tryptophan W220 residue located at the putative dimer dimer interface was mutated to alanine, and the structural and functional consequences of the substitution were analysed. Equilibrium sedimentation experiments revealed that, in contrast to the wild-type Cfr10I, the W220A mutant exists in solution predominantly as a dimer. In addition, the tetramer seems to be a catalytically important form of Cfr10I, since the DNA cleavage activity of the W220A mutant is < 0.1% of that of the wild-type enzyme. Further, analysis of plasmid DNA cleavage suggests that the Cfr10I tetramer is able to interact with two copies of the recognition sequence, located on the same DNA molecule. Indeed, electron microscopy studies demonstrated that two distant recognition sites are brought together through the DNA looping induced by the simultaneous binding of the Cfr10I tetramer to both sites. These data are consistent with the tetramer being a functionally important form of Cfr10I. PMID- 10518947 TI - Intrabody construction and expression. I. The critical role of VL domain stability. AB - We have constructed a panel of hyperstable immunoglobulin VL domains by a rational approach of consensus sequence engineering and combining stabilizing point mutations. These prototype domains unfold fully reversibly, even when the conserved structural disulfide bridge is reduced. This has allowed us to probe the factors that limit the expression yield of soluble immunoglobulin domains in the reducing environment of the cytoplasm (intrabodies). The most important factor is thermodynamic stability, and there is an excellent quantitative correlation between stability and yield. Surprisingly, an unprocessed N-terminal methionine residue can severely compromise VL stability, but this problem can be overcome by changing the amino acid following the initiator methionine residue. Transcription from the strong T7 promoter does not increase the amount of soluble material over that obtained from the tetA promoter, but large amounts of inclusions bodies can be obtained. Elevated temperature shifts protein from a productive folding pathway to aggregation. The structural disulfide bridge does not form in the cytoplasm, but the two consensus cysteine residues can be quantitatively oxidized in vitro. In summary, stability engineering provides a plannable route to the high-yield cytoplasmic expression of functional intrabody domains. PMID- 10518948 TI - Intrabody construction and expression. II. A synthetic catalytic Fv fragment. AB - In general, proteins with structural disulfides cannot be expressed in the reducing environment of the cellular cytoplasm. To overcome this folding problem, we have previously engineered stabilizing mutations, predicted from a consensus sequence analysis, into isolated immunoglobulin VL domains. Here we show that such domains can be used as a framework in the construction of a functional heterodimeric Fv fragment, which was expressed solubly, with high yield in the cytoplasm of Escherichia coli. This designed catalytic intrabody, obtained from grafting the combining site of the esterolytic antibody 17E8, is active in the oxidized and the reduced state. Its construction required no special features on the part of the immunoglobulin, no single-chain linker and introduced no non natural sequence motifs. The potential to design intrabodies with the recognition sequences of arbitrary immunoglobulins opens novel opportunities for gene therapy, cell biology, metabolic engineering and antibody biotechnology. PMID- 10518949 TI - The HU protein from Thermotoga maritima: recombinant expression, purification and physicochemical characterization of an extremely hyperthermophilic DNA-binding protein. AB - The histone-like protein TmHU from the hyperthermophilic eubacterium Thermotoga maritima was cloned, expressed to high levels in Escherichia coli, and purified to homogeneity by heat precipitation and cation exchange chromatography. CD spectroscopical studies with secondary structure analysis as well as comparative modeling demonstrate that the dimeric TmHU has a tertiary structure similar to other homologous HU proteins. The Tm of the protein was determined to be 96 degrees C, and thermal unfolding is nearly completely reversible. Surface plasmon resonance measurements for TmHU show that the protein binds to DNA in a highly cooperative manner, with a KD of 73 nM and a Hill coefficient of 7.6 for a 56 bp DNA fragment. It is demonstrated that TmHU is capable to increase the melting point of a synthetic, double-stranded DNA (poly[d(A-T)]) by 47 degrees C, thus suggesting that DNA stabilization may be a major function of this protein in hyperthermophiles. The significant in vitro protection of double-helical DNA may be useful for biotechnological applications. PMID- 10518950 TI - Stability of a homo-dimeric Ca(2+)-binding member of the beta gamma-crystallin superfamily: DSC measurements on spherulin 3a from Physarum polycephalum. AB - Spherulin 3a (S3a) from Physarum polycephalum represents the only known single domain member of the superfamily of beta gamma eye-lens crystallins. It shares the typical two Greek-key motif and is stabilized by dimerization and Ca(2+) binding. The temperature and denaturant-induced unfolding of S3a in the absence and in the presence of Ca2+ were investigated by differential scanning calorimetry and fluorescence spectroscopy. To accomplish reversibility without chemical modification of the protein during thermal denaturation, the only cysteine residue (Cys4) was substituted by serine; apart from that, the protein was destabilized by adding 0.5-1.8 M guanidinium chloride (GdmCl). The Cys4Ser mutant was found to be indistinguishable from natural S3a. The equilibrium unfolding transitions obey the two-state model according to N2-->2 U, allowing thermodynamic parameters to be determined by linear extrapolation to zero GdmCl concentration. The corresponding transition temperatures TM for the Ca(2+)-free and Ca(2+)-loaded protein were found to be 65 and 85 degrees C, the enthalpy changes delta Hcal, 800 and 1280 kJ/mol(dimer), respectively. The strong dependencies of TM and delta Hcal on the GdmCl concentration allow the molar heat capacity change delta Cp to be determined. As a result, delta Cp = 18 kJ/(K mol(dimer)) was calculated independent of Ca2+. No significant differences were obtained between the free energy delta G degree calculated from delta Hcal and TM, and extrapolated from the stability curves in the presence of different amounts of denaturant. The free energy derived from thermal unfolding was confirmed by the spectral results obtained from GdmCl-induced equilibrium transitions at different temperatures for the Ca(2+)-free or the Ca(2+)-loaded protein, respectively. Within the limits of error, the delta G degree values extrapolated from the transitions of chemical denaturation to zero denaturant concentration are identical with the calorimetric results. PMID- 10518951 TI - New pathways in folding of the Tetrahymena group I RNA enzyme. AB - Previous studies have shown that the earliest detectable step in folding of the Tetrahymena ribozyme is tertiary structure formation of the peripheral element P5abc. This, along with other results, has suggested that P5abc may serve as a scaffold upon which additional tertiary structure is built. Herein we use the onset of oligonucleotide cleavage activity as a readout for native state formation and investigate the effect of P5abc on the rate of folding to the native structure. Despite the early folding of P5abc, its removal to give the E delta P5abc variant decreases the rate of attainment of an active structure less than fivefold (20-100 mM Mg2+, 15-50 degrees C). Furthermore, P5abc added in trans is able to bind the folded E delta P5abc ribozyme and promote oligonucleotide cleavage at least tenfold more rapidly than folding of the wild type ribozyme, indicating that E delta P5abc does not have to first unfold before productively binding P5abc to form the true native state. This suggests that a state with the overall tertiary structure formed but with P5abc unfolded represents a viable on-pathway intermediate for the wild-type ribozyme. These results provide strong evidence for the existence of two pathways to the native state: in one pathway P5abc forms tertiary structure first, and in another it forms late. The pathway in which P5abc forms first is favored because P5abc can fold quickly and because its tertiary structure is stable in the absence of additional structured elements, not because P5abc formation is required for subsequent folding steps. In the course of these experiments, we also found that most of the ribozyme population does not reach the native state directly under standard conditions in vitro, but instead forms an inactive structure that is stable for hours. Finally, the fraction that does fold to the native state folds with a single rate constant of 1 min-1, suggesting that there are no significantly populated "fast-track" pathways that reach the native state directly by avoiding slow folding steps. PMID- 10518952 TI - Functional reconstitution and characterization of AqpZ, the E. coli water channel protein. AB - Understanding the selectivity of aquaporin water channels will require structural and functional studies of wild-type and modified proteins; however, expression systems have not previously yielded aquaporins in the necessary milligram quantities. Here we report expression of a histidine-tagged form of Escherichia coli aquaporin-Z (AqpZ) in its homologous expression system. 10-His-AqpZ is solubilized and purified to near homogeneity in a single step with a final yield of approximately 2.5 mg/l of culture. The histidine tag is removed by trypsin, yielding the native protein with the addition of three N-terminal residues, as confirmed by microsequencing. Sucrose gradient sedimentation analysis showed that the native, solubilized AqpZ protein is a trypsin-resistant tetramer. Unlike other known aquaporins, AqpZ tetramers are not readily dissociated by 1% SDS at neutral pH. Hydrophilic reducing agents have a limited effect on the stability of the tetramer in 1% SDS, whereas incubations for more than 24 hours, pH values below 5.6, or exposure to the hydrophobic reducing agent ethanedithiol cause dissociation into monomers. Cys20, but not Cys9, is necessary for the stability of the AqpZ tetramer in SDS. Upon reconstitution into proteoliposomes, AqpZ displays very high osmotic water permeability (pf > or = 10 x 10(-14) cm3 s-1 subunit-1) and low Arrhenius activation energy (Ea = 3.7 kcal/mol), similar to mammalian aquaporin-1 (AQP1). No permeation by glycerol, urea or sorbitol was detected. Expression of native and modified AqpZ in milligram quantities has permitted biophysical characterization of this remarkably stable aquaporin tetramer, which is being utilized for high-resolution structural studies. PMID- 10518953 TI - Structure of the water channel AqpZ from Escherichia coli revealed by electron crystallography. AB - Molecular water channels (aquaporins) allow living cells to adapt to osmotic variations by rapid and specific diffusion of water molecules. Aquaporins are present in animals, plants, algae, fungi and bacteria. Here we present an electron microscopic analysis of the most ancient water channel described so far: the aquaporin Z (AqpZ) of Escherichia coli. A recombinant AqpZ with a poly(histidine) tag at the N terminus has been constructed, overexpressed and purified to homogeneity. Solubilized with octylglucoside, the purified AqpZ remains associated as a homotetramer, and assembles into highly ordered two dimensional tetragonal crystals with unit cell dimensions a = b = 95 A, gamma = 90 degrees when reconstituted by dialysis in the presence of lipids. Three dimensional reconstruction of negatively stained lattices revealed the p42(1)2 packing arrangement that is also observed with the human erythrocyte water channel (AQP1). The 8 A projection map of the AqpZ tetramer in frozen hydrated samples is similar to that of AQP1, consistent with the high sequence homology between these proteins. PMID- 10518954 TI - An NMR investigation of solution aggregation reactions preceding the misassembly of acid-denatured cold shock protein A into fibrils. AB - At pH 2.0, acid-denatured CspA undergoes a slow self-assembly process, which results in the formation of insoluble fibrils. 1H-15N HSQC, 3D HSQC-NOESY, and 15N T2 NMR experiments have been used to characterize the soluble components of this reaction. The kinetics of self-assembly show a lag phase followed by an exponential increase in polymerization. A single set of 1H-15N HSQC cross-peaks, corresponding to acid-denatured monomers, is observed during the entire course of the reaction. Under lag phase conditions, 15N resonances of residues that constitute the beta-strands of native CspA are selectively broadened with increasing protein concentration. The dependence of 15N T2 values on spin echo period duration demonstrates that line broadening is due to fast NMR exchange between acid-denatured monomers and soluble aggregates. Exchange contributions to T2 relaxation correlate with the squares of the chemical shift differences between native and acid-denatured CspA, and point to a stabilization of native like structure upon aggregation. Time-dependent changes in 15N T2 relaxation accompanying the exponential phase of polymerization suggest that the first three beta-strands may be predominantly responsible for association interfaces that promote aggregate growth. CspA serves as a useful model system for exploring the conformational determinants of denatured protein misassembly. PMID- 10518955 TI - [Cytokines of bone turnover in postmenopause and old age]. AB - BACKGROUND: The aim of the study was the assessment of the influence of cytokines on bone ageing by measuring their level in serum and their secretion in vitro by monocytes from women of different age. METHODS: The levels of cytokines in 34 postmenopausal subjects and 14 old subjects were compared to those measured in 13 cycling subjects who were considered as control group. Subjects suffering from diseases inducing secondary osteoporosis, subjects taking medications that affect bone metabolism and alcohol- or tobacco-consumers were excluded from the study. The levels in serum of (i) the bone stimulating peptide insulin-like growth factor-I (IGF-I), (ii) the inhibitor of bone resorption interferon-gamma (IFN gamma), (iii) the stimulators of bone resorption interleukin-1 beta (IL-1 beta) tumor necrosis factor alpha (TNF alpha) and interleukin 6 (IL-6) were evaluated by immunoassay. IL-1 beta, TNF alpha and IL-6 secreted by monocytes (MO) cultured in vitro from peripheral blood of the same subjects were measured, too. Bacterial endotoxin (LPS) was used as stimulator for cytokine secretion by monocytes. RESULTS: Unlike IFN gamma, which was unaltered, circulating IGF-I level was found significantly diminished in postmenopausal and old subjects compared to control group. Among stimulators of bone resorption, IL-6 was greatly increased in postmenopausal and old subjects, while TNF alpha was reduced in postmenopausal group. In the supernatants of unstimulated monocytes the level of IL-1 beta was consistently decreased in old subjects; TNF alpha was found to be decreased in postmenopausal and old subjects. The stimulation index (SI), calculated as the ratio between the level of cytokines secreted by LPS-stimulated MO and the level of cytokines secreted by unstimulated MO, was found to be significantly increased for IL-1 beta and TNF alpha in postmenopausal subjects vs control group. In the old subjects the SI for IL-6 was enhanced. CONCLUSIONS: The data collected suggest that the measurement of cytokines in serum and supernatants from monocytes may give a picture of the mechanisms regulating bone aging. PMID- 10518956 TI - [Thyroid function in elderly with neoplasms]. AB - BACKGROUND: A variety of severe illnesses can induce changes in thyroid hormone metabolism, leading to findings referred to as "sick euthyroid syndrome" (ESS). These thyroid hormone changes may be mediated in part by cytokines or other inflammatory mediators, acting at the level of the hypothalamus and pituitary gland, the thyroid gland, and the hepatic deiodinase system. The degree of thyroid function disturbance correlates with disease severity and low levels of thyroid hormones predict a poor prognosis in several illnesses. It remains unresolved whether the hormone responses in the ESS represent part of an adaptative response, which lowers tissue energy requirements in the face of systemic illness, or a maladaptive response, which induces damaging tissue hypothyroidism. METHODS: This study examines the incidence of ESS among 220 elderly subjects hospitalized with cancer. In each subjects some individual variables were studied: age, sex, type of cancer, presence of metastasis, rapid shortage of corporeal weight, general clinic condition. The following laboratory parameters were studied: serum glucose, sodium, potassium, calcium, cholesterol, lipids, proteins, leukocytes, serum, lipids haemoglobin, plateletes, VES and the end free triiodothyronine, free tiroxine and thyrotropin. RESULTS: The research points out that ESS is more frequent in the elderly subjects with cancer (incidence 58%). ESS type 1, with FT3 low and FT4 and TSH normal, is the most frequent form. The incidence of ESS is higher in elderly subjects with cancer, with recent and marked lose of corporeal weight (incidence 86%) and in subjects with worst clinical conditions. Finally, a significant direct correlation between FT3/serum cholesterol, FT3/serum proteins, FT3/serum sodium, FT3/FT4 is observed. CONCLUSIONS: The results obtained point out the not yet solved problem of hormone replacement therapy in elderly patients with cancer in bad clinical conditions. PMID- 10518957 TI - [Hepatic and pancreatic disease in patients with acquired immunodeficiency syndrome (AIDS)]. AB - AIDS is frequently expressed through gastrointestinal o abdominal symptoms. In addition, patients with AIDS or ARC frequently have hepatic and biliary symptoms, while pancreatic alterations are found in 4-30% of patients hospitalised for AIDS. Since AIDS patients are immunodepressed, they are subject to opportunistic infection often multifocal and the pathological processes can be present simultaneously. About 2/3 of patients have enlarged liver, steatosis, splenomegaly, lymphoadenopathy, cholecystic and biliary tract abnormalities, alterations of liver function tests, and abdominal discomfort in the upper right quadrant. Jaundice is rare and hepatic failure is not common. Hepatic biopsy is often necessary to establish the diagnosis. The hepatic localisation of an opportunistic pathogenic agent is generally a sign of systemic dissemination which is expressed as granulomatous hepatitis (atypical mycobacteria, frequently mycobacterium avium, or M. tuberculosis representing the reactivation of latent diseases), peliosis hepatis, infection from CMV, HSV, EBV, Pneumocystis carinii, and mycotic infections. Coinfections with the hepatic virus (HBV, HDV, HCV) are also often present. Pharmacological damage may also be present (mainly caused by antibiotic therapies). Neoplasia are rare (hepatic Kaposi's sarcoma associated with cutaneous and gastrointestinal manifestations, or generally metastatic lymphoma). Damage of the biliary tract usually develops after other manifestations of the illness; the most frequent pictures are cholestatic syndromes and cholangitis, while cholecystitis and jaundice are rare. Pancreatic lesions are generally asymptomatic. They are diagnosed during autopsy and are caused principally by opportunistic agents. PMID- 10518958 TI - [Adult-onset Still's disease. Report of 5 cases]. AB - The authors report 5 cases of Still's disease in adults whose symptoms were mainly characterised by high fever, transient exanthema, polyarthralgia and/or polyarthritis, lymphoadenomegaly, splenomegaly and neutrophil leukocytosis. Assays for leukocytosis were positive, as were those for inflammatory markers and serum ferritin was also high in all 3 patients in which it assayed. On the contrary, serum ferritin latex test, Waaler-Rose reaction and all other tests commonly used to diagnose long-term fevers were all negative. All the subjects examined recovered after prolonged steroid therapy. Only one patient reported severe sequelae in the hip joints and subsequently underwent bilateral hip replacement surgery. PMID- 10518959 TI - [Splenic artery aneurysm and portal hypertension. Report of a case]. AB - Visceral arteries aneurysms are an uncommon pathology. Lienal artery (60%), hepatic (20%), superior mesenteric (5.9%) and tripod celiac (4%) are the most involved. Female predominance, portal hypertension, arteriosclerosis, pregnancy, traumatic and infective factors are the most remarkable etiopathogenetic factors. Portal hypertension with splenomegaly might be an important factor in the pathogenesis of intra- and extraparenchimal splenic artery aneurysms. Stress is laid on the importance of arteriography during portal hypertension in pregnancy as a preventive measure because the rupture of aneurysm of the splenic artery, despite laparotomy in emergency is often fatal, and of coeliac angiography prior to liver transplantation; if splenic artery aneurysm is found, ligation of the splenic artery should be performed at the time of transplantation to prevent possible rupture. The different roles of the imaging techniques used are examined. PMID- 10518960 TI - Immunolocalization of MP84 in renal biopsy sections of sickle cell nephropathy patients. PMID- 10518961 TI - [Experience with an "isolation unit" for patients infected with multi-resistant bacteria. Retrospective study of 49 patients]. AB - OBJECTIVES: Perform a retrospective analysis of care in a hospital "isolation unit" for patients infected with multirestant bacteria (MRB), i.e. meticillin resistant staphylococcus aureus (SAMR), broad spectrum beta-lactamase secreting enterobacteria. (BLSE). PATIENTS AND METHODS: Forty-nine patients infected with MRB were cared for in our hospital isolation unit between January 1, 1996 and January 1, 1997. Each patient was in a separate room equipped with a sink and soap distributor, single-use towels, and individual material for patient care (stethoscope, mobile equipment, writing material, etc.). The personnel were given special training in the prevention of nosocomial infections. At admission, and in all patients, bacteriological samples to search for SAMR were acquired from nasal discharge, urine, perineal swabs, wounds and bed sores. Wound, urine and fecal samples were also taken to search for BLSE. Search for other sites of infection depended on the clinical situation. The management protocol in the isolation unit included: isolation, daily antiseptic baths, topical application of antibiotics or antiseptics on all bacteriologically proven sites of SAMR infection, selective decontamination of the digestive tract for patients with BLSE positive stools. Systemic antibiotics were given case by case. RESULTS: Mean duration of stay in the isolation unit was 17 days for SAMR infections and 14 days for BLSE infections. Mean delay to sterilization of the infected sites varied depending on the localization: 2.3 days for blood and 19.4 days for stools. Seven patients died. After leaving the isolation unit, the bacteriological course was followed in 23 patients: there were 7 cases of recurrence at least one site within a mean delay of 34.5 days. CONCLUSION: Use of isolation units provides an interesting solution for health care centers to control spread of multiresistant bacteria. Considering the endemic state of multiresistant bacteria infections in French hospitals, each health care unit should have correctly equipped facilities for isolating infected patients. PMID- 10518962 TI - [Ten-year experience with heart transplantation (1987-1997)]. AB - OBJECTIVES: Analyze ten years experience with heart transplantation at the Dijon University Hospital and determine which parameters control mid and long term outcome. PATIENTS AND METHODS: One hundred thirty six heart transplantations were performed over a 10 year period (1987-1997) in 118 men and 18 women aged 51-87 years. Heart transplantation was indicated on the basis of the following criteria: ejection fraction *20%, pulmonary arteriole resistance < 6 Wood units, peak oxygen uptake < 14 l/kg/min. The Shumway or anatomic technique was used. The triple immunosuppressive protocol combined corticosteroids, azathioprine and cyclosporin. The same team conducted the post-transplantation follow-up with regular programmed consultations in addition to those requested by the general practitioner, the cardiologist or the patient. Follow-up was oriented according to the clinical situation (blood chemistry, cell counts, cyclosporinemia, search for infection, echocardiography, endomyocardial biopsy, coronarography). RESULTS: Five patients (3.6%) died when still on the waiting list. Absolute emergency transplantation was performed for patients (28.1%) including 8 (5.9%) after circulatory assist. Hospital mortality was 11.7% and late mortality was 16.1%. Actuarial survival was 78% at 1 year, 71% at 5 years and 69% at 10 years. Among the survivors, 94% were taking two, three or even four drugs for hypertension. Cyclosporin levels decreased and creatinine levels increased. Episodes of rejection were minimal: 86.57% of the biopsies were * grade 1 and 4.45% * grade 2. Cytomegalovirus infection was documented and treated in 7.55% of the cases. Incidence of graft coronary artery disease was 3.4% at 1 year, 6.5% at 2 years and 7.9% at 3 years. CONCLUSION: Our follow-up structure where the same small team conducts regular examinations together with our approach to heart transplantation appears to be the main factor leading to the quality results obtained in this series. PMID- 10518963 TI - [Action of piribedil in Parkinson disease. Multicenter study]. AB - OBJECTIVES: Several controlled trials have shown that Trivastal (piribedil), a direct dopamine agonist, is active in the treatment of Parkinson's disease. The aim of the present clinical trial was to assess the efficacy of Trivastal 50 mg LP administered as monotherapy in patients naive to treatment with L-dopa. PATIENTS AND METHODS: This 3-month multicenter study was conducted in 113 patients (66 men and 47 women), aged 63.1 +/- 0.6 years, with a 2.1 +/- 0.2 year history of Parkinson's disease and a mean Hoehn and Yahr stage of 1.82. Tremor was the predominant clinical feature in 42 patients; the 71 others presented with the full parkinsonian syndrome. Trivastal 50 mg LP doses were increased stepwise, up to doses of 150-250 mg/day at the end of the 3-month study period. Patients were clinically assessed at 1, 2 and 3 months using the Webster scale and the HARD depression scale. RESULTS: In the 90 patients who completed the study, tremor fell from 1.7 to 1.0 (-41%, p < 0.001), bradykinesia from 1.5 to 0.8 ( 47%, p < 0.001) and rigidity from 1.3 to 0.9 (-31%, p < 0.001). The 32 patients in whom tremor was the predominant feature improved their total score on the Webster scale from 5.8 to 4.7 (-19%, p < 0.05). The 58 patients with the full parkinsonian syndrome improved their total Webster score from 11.8 to 6.9 (-42%, p < 0.001). The depression score fell from 10.2 to 7.3 (p < 0.001), the most marked improvement being in mood and inhibition. CONCLUSION: Monotherapy with Trivastal 50 mg LP at a mean dose of 200 mg/day is effective on the major symptoms of Parkinson's disease. PMID- 10518964 TI - [Latent adrenal gland insufficiency in a HIV-infected patient with phospholipid antibodies]. PMID- 10518965 TI - [Late treatment: main severity factor in flare-up of P. falciparum infection]. PMID- 10518966 TI - [Bacterial translocation in colchicine poisoning]. PMID- 10518967 TI - [Arterial hypertension and left ventricular hypertrophy: a 40-year follow-up in a Framingham cohort study]. PMID- 10518968 TI - [Contribution of the pulmonary angio-scanner to the emergency diagnosis of pulmonary embolism]. PMID- 10518969 TI - [35th Congress of the American Society of Clinical Oncology. Results of an extensive randomized test series]. PMID- 10518970 TI - [African Rickettsia infections]. AB - EPIDEMIOLOGY: African rickettsiasis is transmitted by Rickettsia africae, a cattle tick. Amblyomma spp. is an emerging rickettsiasis in sub-sehalian Afric described in 1992. Seroepidemiology studies conducted in Africa show that it is probably the most widespread rickettsiasis in the sorld. In addition, the development of tourist activities in southern African countries has led to an increase in the number of reported cases in subjects returning from endemic areas. A high serprevalence of anti R. africae anticodies has been recently reported in the guadeloupe (French East Indies) population as well as one documented infection. CLINIC: The clinical expression of African tick rickettsiasis includes fevder, headache, inoculation scar, locoredgional node enlargement, and an inconsistent sometimes vesicular rash. The diagnosis is made on the basis of serological findings and cross absorption of anti R. africae and R. conorii antibodies and/or isolation or gene amplification of R. africae from inoculation scar biopsies. PMID- 10518972 TI - [Pseudohypoparathyroidism and the concept of hormonal resistance. Types Ia and Ic and pseudohypoparathyroidism]. AB - TYPE IA: This familial hereditary condition is characterized by the association of Albright's osteodystrophy, resistance to parathormone (PTH) and a negative PTH test both for urinary phosphorus and cyclic AMP. The condition is caused by an anomalous alpha sub-unit of protein G, impairing its function. The result is defective transmembrane transduction of the PTH mediated signal. Protein G, coupled with all heptahelical membrane-spanning receptors, is ubiquitous, explaining the association of multiple hormone and neurosensory resistances. Resistance of the thyrotrope and gonadotrope axes should be explored to institute appropriate replacement therapy. TYPE IC: This type associates all the clinical and biological features of type Ia pseudohypoparathyroidism but without any protein G defect, suggesting another effector of signal transduction, perhaps adenylate cyclase, is involved. PSEUDOPSEUDOHYPOPARATHYROIDISM: Often in families with type Ia pseudohypoparathyroidism, subjects with Albright's osteodystrophy alone, with no features of PTH resistance, are said to have pseudopseudohypoparathyroidism. Protein G defects are also demonstrated in these subjects, confirming the relationship with type la pseudohypoparathyroidism and explaining the possible multiple hormone resistances observed. The phenotypic variability between type Ia pseudohypoparathyroidism and pseudopseudohypoparathyroidism would be related to genomic imprinting mechanism. PMID- 10518971 TI - [Pseudohypoparathyroidism and the concept of hormonal resistance. Diagnosis, classification and therapy]. AB - CLINICAL AND BIOLOGICAL FEATURES: Pseudohypoparathyroidism is a heterogenous group of conditions with variable clinical and biological features and a common resistance to parathormone (PTH) leading to hypocalcemia associated with high levels of PTH. The classification of these conditions depends on the expression of Albright's osteodystrophy and response to exogenous parathormone: urine phosphorus and cyclic AMP excretion. TYPE IA: The characteristic feature is Albright's osteodystrophy associated with a totally negative response to exogenous PTH. Defective protein G is the cause. Pseudopseudohypoparathyroidism is defined by Albright's osteodystrophy without resistance to PTH. This condition occurs in families with type Ia pseudohypoparathyroidism. It is also related to a defect in protein G. TYPE IB: Type Ib corresponds to PTH resistance alone, without Albright's osteodystrophy. This condition is apparently secondary to anomalous regulation of the gene coding the PTH receptor. TYPE II: Type II is defined by the inconstant presence of Albright's osteodystrophy and a dissociated response to PTH: urinary phosphorus does not respond to PTH (hormone resistance) but urine cyclic AMP increases in response to PTH suggesting anomalous signal transduction downstream from adenyl cyclase. TREATMENT: In all types, treatment is based on combining calcium therapy and vitamin D supplementation under rigorously controlled conditions. PMID- 10518973 TI - [Pseudohypoparathyroidism and the concept of hormonal resistance. Types Ib and II]. AB - TYPE IB PSEUDOHYPOPARATHYROIDISM: Parathormone resistance is the only manifestation of type Ib pseudohyoparathyroidism, the Albright osteodystrophy and multiple hormone resistance described in type Ia are not observed. In type Ib there is a certain preservation of bone sensitivity to parathormone although the main target organ, the kidney, is resistant. Consequently, excessive bone remodeling can be evidenced by X-ray (subperiosteal resorption, fibrocystic osteitis), chemistry (high serum alkaline phosphatase and osteocalcin, and increased urine hydroxyproline), densitometry (lower bone density), and pathology (reduction in trabecular bone volume). The dissociation of the bone and kidney response corresponds to the pseudohypohyperthyroidism described by certain authors. The genetic substratum leading to type Ib pseudohypoparathyroidism remains to be identified. The pathogenic mechanism generally hypothesized concerns a qualitative or quantitative anomaly of the parathormone receptor but seems to be disproved by recent studies. TYPE II PSEUDOHYPOPARATHYROIDISM: Here there is a characteristic lack of a rise in urine phosphorus, signaling parathormone resistance, and stimulated urinary excretion of cyclic AMP, an expression of the integrity of the transmembrane transduction of the parathormone mediated signal and thus protein G and adenylate cyclase. The anomaly could thus involve a transductional effector situated downstream from adenylate cyclase which would explain the symptoms of type II pseudo-hypoparathyroidsm, with both parathormone resistance and Albright osteodystrophy. PMID- 10518974 TI - CT-Based interstitial HDR brachytherapy. AB - PURPOSE: Development, application and evaluation of a CT-guided implantation technique and a fully CT-based treatment planning procedure for brachytherapy. METHODS AND MATERIALS: A brachytherapy procedure based on CT-guided implantation technique and CT-based treatment planning has been developed and clinical evaluated. For this purpose a software system (PROMETHEUS) for the 3D reconstruction of brachytherapy catheters and patient anatomy using only CT scans has been developed. An interface for the Nucletron PLATO BPS treatment planning system for optimization and calculation of dose distribution has been devised. The planning target volume(s) are defined as sets of points using contouring tools and are used for optimization of the 3D dose distribution. Dose-volume histogram based analysis of the dose distribution (COIN analysis) enables a clinically realistic evaluation of the brachytherapy application to be made. The CT-guided implantation of catheters and the CT-based treatment planning procedure has been performed for interstitial brachytherapy and for different tumor sites in 197 patients between 1996 and 1997. RESULTS: The accuracy of the CT reconstruction was tested using first a quality assurance phantom and second, a simulated interstitial implant of 12 needles. These were compared with the results of reconstruction using radiographs. Both methods gave comparable results with regard to accuracy, but the CT based reconstruction was faster. Clinical feasibility was proved in pre-irradiated recurrences of brain tumors, in pretreated recurrences or metastatic disease, and in breast carcinomas. The tumor volumes treated were in the range 5.1 to 2,741 cm3. Analysis of implant quality showed a slightly significant lower COIN value for the bone implants, but no differences with respect to the planning target volume. CONCLUSIONS: The Offenbach system, incorporating the PROMETHEUS software for interstitial HDR brachytherapy has proved to be extremely valuable in routine clinical practice for many tumor sites. Our CT-guided implantation technique together with a fully CT-based planning system has enabled conformal brachytherapy treatment to become routine. PMID- 10518975 TI - [Not a new brachytherapy system]. PMID- 10518976 TI - [Delayed toxicity of brief preoperative irradiation and risk-adjusted postoperative radiotherapy of operative rectal carcinoma. Results of a randomized prospective study]. AB - AIM: Analysis of a randomized study of preoperative radiation therapy for operable carcinoma of the rectum with regard to late sequelae. Results of tumor control and survival, which have already been published in detail are summarized for comparison and for confirmation of the conclusions. PATIENTS AND METHODS: Between January 1988 and October 1993 94 patients with operable carcinoma of the rectum were included in a randomized trial. Fourty-seven patients were treated with 5 x 3.3 Gy (field size 16 x 16 cm, 9 MeV photons) 24 to 48 hours prior to surgery; 46 patients did not receive preoperative irradiation. If risk factors (T4-stage, R1/R2 resection, intraoperative tumor perforation) were present, postoperative irradiation was performed after CT-planning. Total postoperative doses of 41.4 Gy (preoperative irradiation) or 59.8 Gy (surgery only) were applied with doses per fraction of 1.8 to 2.0 Gy. Local control, survival, and pattern of side effects were analyzed at 5 years after conclusion of the trial. RESULTS: The frequency of local recurrence was markedly reduced by preoperative irradiation of R0-resected patients (24% vs 13%, p = 0.08). The time to recurrence was delayed (1.9 vs 3 years). The 5-year actuarial survival rate was significantly higher in the preoperatively irradiated group compared to the not pre-irradiated group (40% vs 28%, p = 0.027). Multivariate analysis revealed UICC grading as the only independent parameter for local control (p = 0.0003), while preoperative irradiation (p = 0.07) and T-stage (p = 0.08) only displayed a trend. For patient survival, age (p = 0.0003). R-status (p = 0.01) and UICC-score (p = 0.001) were significant prognostic factors. Preoperative irradiation had a non-significant effect only (p = 0.078). Radiation-induced side effects with a LENT-SOMA score > 2 were observed neither during frequent follow-up nor at an additional examination of those patients still alive in 1998 (n = 25). Of 4 pre- and postoperatively irradiated patients with risk factors, 3 had side effects grade 1 or 2, predominantly rectal changes, at 5 to 11 years after treatment. CONCLUSIONS: A positive effect on tumor control and survival is achieved with preoperative irradiation with the doses used in this study, with moderate side effects. PMID- 10518977 TI - [Biological prevention and/or therapy possibility of radiation myelopathy. A discussion of recent perspectives]. AB - BACKGROUND: Radiosensitivity of the spinal cord makes both curative first-line treatment of numerous malignancies and re-irradiation of recurrent or second tumors more difficult. This review discusses recent advances in basic research that alter the view on the pathogenesis of radiation myelopathy, possibly offering strategies for prevention and/or therapy. RESULTS: Available data of developmental neurobiology and preclinical studies of demyelinating diseases revealed interesting insights into oligodendrocyte development, intercellular signaling pathways, and myelination processes. Current findings suggest that administration of cytokines could increase proliferation of oligodendrocyte progenitor cells, enhance their differentiation, upregulate synthesis of myelin constituents, and promote myelin regeneration in the adult central nervous system (Table 1). Other compounds might also be able to modulate progression of pathogenic processes that eventually lead to radiation myelopathy. This offers several possible biological prevention and/or treatment strategies, which currently are being investigated in animal studies (Table 2). CONCLUSION: Technical options as well as optimization of fractionation parameters should be given priority in the attempt to reduce iatrogenic neurotoxicity. However, rational biological strategies could offer a new perspective for many patients. PMID- 10518978 TI - Daily amifostine given concomitantly to chemoradiation in head and neck cancer. A pilot study. AB - BACKGROUND: In patients with loco-regionally advanced head and neck cancer conventionally fractionated radiotherapy alone results in poor loco-regional control and survival rates. Treatment intensification by simultaneous administration of cytotoxic drugs produces higher acute morbidity. Therefore chemical radioprotection of normal tissues may be of clinical benefit. PATIENTS AND METHODS: In a pilot study patients with advanced nonresectable head and neck cancer treated with conventionally fractionated radical radiotherapy (60 to 66 Gy total doses) and concomitantly given 5-fluorouracil as protracted venous infusion, 250 mg/sqm/24 h over the entire treatment period were given amifostine 300 mg absolutely before each fraction. Acute treatment related morbidity was scored according to CTC classification and loco-regional control and survival rates were estimated. Comparison was made with a historical control group of identical chemoradiation but without amifostine application. RESULTS: Chemoradiation induced oral mucositis was delayed and showed significant lower degrees at all 10 Gy increments (p < 0.05) except 60 Gy and over (p > 0.05). No significant toxicity was recorded with respect to blood pressure, serum calcium, potassium, hematologic parameters, emesis, nausea or body weight loss. Progression free survival and overall survival probability at 2 years were not statistically different in both cohorts. CONCLUSION: Amifostine given before each fraction of radiotherapy over 6 weeks has no cumulative toxicity, was well tolerated and may reduce treatment induced oral mucositis. No tumor protective effect was observed. PMID- 10518979 TI - [Locally recurrent and metastatic malignant melanoma. Long-term results and prognostic factors after percutaneous radiotherapy]. AB - PURPOSE: Radiotherapy (RT) is used as last resort for patients with advanced cutaneous malignant melanoma (MM). Herein our 20-year clinical experience is presented analyzing different endpoints and prognostic factors in patients with locally advanced, recurrent or metastatic MM. PATIENTS AND METHODS: From 1977 to 1995, 2,917 consecutive patients were entered in the MM registry of our university hospital. RT was indicated in 121 patients (56 females, 65 males) for palliation in locally advanced recurrent and metastatic MM stages UICC IIB to IV. At the time of RT initiation, 11 patients had primary or recurrent lesions which were either not eligible for surgery or had residual disease (R2) after resection of a primary or recurrent MM lesion (UICC IIB); 57 patients had lymph node (n = 33) or in-transit metastases (n = 24) (UICC III), and 53 had distant organ metastases (7 M1a, 46 M1b) (UICC IV). The time from first diagnosis to on-study RT averaged overall 19 months (median: 18; range: 3 to 186 months). In 77 patients conventional RT and in 44 patients hypofractionted RT was applied with 2 to 6 Gy fractions up to a mean total RT dose of 45 (median: 48; range: 20 to 66) Gy. RESULTS: At 3 months follow-up, complete response (CR) was achieved in 7 (64%), overall response (CR + PR) in all (100%) UICC IIB patients, in 25 (44%) and 44 (77%) of 57 UICC III patients, and in 9 (17%) and 26 (49%) of 53 UICC IV patients. Tumor progression during RT occurred in 25 (21%) patients. Patients with CR survived longer (median: 40 months) than those without CR (median 10 months) (p < 0.01). At the time of evaluation and last FU (December 31, 1996), 26 patients were still alive: 6 (55%) stage UICC IIB, 17 (30%) stage UICC III, and 3 (6%) stage UICC IV patients (p < 0.01). Univariate analysis revealed following prognostic factors for CR and long-term survival: UICC stage (p < 0.001), primary location in the head and neck, total RT dose > 40 Gy (all p < 0.05), while age, gender and primary histological subtype had no impact. In multivariate analysis, UICC stage was the only independent favorable prognostic factor for achievement of CR and long-term survival (p < 0.001). CONCLUSION: External RT provides effective palliation in advanced UICC stages. The UICC staging system is a good predictor of initial and long-term tumor response in metastatic MM. Prospective randomized trials using RT with or without adjuvant therapy for advanced MM are justified. PMID- 10518980 TI - 211At-alpha-dose dependence of poly-ADP-ribosylation of human glioblastoma cells in vitro. Suitability in cancer therapy? AB - AIM: It was intended to test the biological response (poly-ADP-ribosylation of cellular proteins) of alpha-particles from extracellular 211At for enhanced damage to human glioblastoma cells in vitro and to discuss its suitability for potential application in therapy of high-grade gliomas. MATERIALS AND METHODS: Confluent cultures of human glioblastoma cells were exposed to different doses of alpha-radiations from homogeneously distributed extracellular 211At. Cellular poly-ADP-ribosylation of all proteins including histones was monitored since it is an indirect but sensitive indicator of chromatin damage and putative repair in both normal and malignant mammalian cells. RESULTS: A significant diminution (average 85.6%) in poly-ADP-ribosylation of total cellular proteins relative to that for non-irradiated glioblastoma cells was observed following 0.025 to 1.0 Gy alpha-radiations. In the dose range of 0.0025 to 0.01 Gy there was an increase with a maximum value of approximately 119.0% at 0.0025 Gy. Below 0.0025 Gy no change in poly-ADP-ribosylation was observed. CONCLUSIONS: Level of cellular poly ADP-ribosylation of proteins at 0.025 to 1.0 Gy of alpha-radiation dose from 211At appears to cause enhanced damage by creating molecular conditions which are not conducive to repair of DNA damages in human glioblastoma cells in vitro. Therefore, it is assumed that clinical application of 211At at least in this dose range might enhance clinical efficacy in radiotherapy of cancer. PMID- 10518981 TI - Three-dimensional treatment planning for postoperative radiotherapy in patients with node-positive cervical cancer. Comparison between a conventional and a conformal technique. AB - PURPOSE: Reduction of irradiated small bowel volume, using a conformal three dimensional treatment planning technique in postoperative radiotherapy of cervical cancer patients. PATIENTS AND METHODS: Large gynecological treatment fields including the para-aortic nodes were analyzed in 15 patients. A conventional treatment plan with anterior and posterior (AP-PA) parallel opposed fields and a 3D 4-field conformal radiotherapy plan with a central blocking of small bowel were compared for each patient. Dose-volume histograms and dose parameters were established. Because of the tolerance constraints of the small bowel, the cumulative dose applied to the target was 48.6 Gy. RESULTS: The mean Tumor Control Probability (TCP) values for both the conventional and the conformal technique were 0.60 and 0.61, respectively, with ranges of 0.56 to 0.67 and 0.57 to 0.66, respectively. The mean volume receiving 95% or more of the prescribed dose (V95) of the small bowel was 47.6% (32.5 to 66.3%) in the AP-PA technique and 14.9% (7.0 to 22.5%) in the conformal technique (p < 0.001), indicating a significant reduction in irradiated volume of small bowel in the higher dose range. The mean Normal Tissue Complication Probability (NTCP) decreased from 0.11 to 0.03 with the conformal plan. In patients who received a pedicled omentoplasty during surgery, the mean V95 for small bowel could be reduced to 8.5% (7.0 to 9.9%). The mean median dose to the kidneys was only slightly elevated in the conformal treatment. Especially the mean dose to the right kidney in conventional vs conformal treatment was 3.3 vs 7.9 Gy. The mean near-minimum dose (D95) to the rectosigmoid decreased from 48.4 to 30.1 Gy in the conformal plan compared to the conventional plan. CONCLUSION: The small bowel dose can be significantly reduced with 3D treatment planning, particularly if a pedicled omentoplasty is performed. This allows dose escalation to the tumor region without unacceptable toxicity for the small bowel. PMID- 10518982 TI - [Postoperative radiochemotherapy of cervix carcinoma]. PMID- 10518983 TI - [Polychemotherapy versus monotherapy in metastatic breast carcinoma]. PMID- 10518984 TI - [Better tumor control by simultaneous radiochemotherapy in advanced head-neck carcinomas]. PMID- 10518985 TI - Introduction: invertebrate axons find their way. AB - The mechanisms that generate the immense complexity of synaptic connections within the developing nervous system have fascinated biologists for decades. Analysis of nervous system development in simple systems, such as insects, has made a major contribution to our understanding of the cellular and molecular mechanisms that control the formation of axon pathways and precise connections. This enterprise has a long, interesting, and somewhat controversial history. This collection of reviews on axon guidance in insects provides a brief update to integrate current molecular and developmental insights in a number of areas from initial axon pathfinding to the recognition of synaptic partners. PMID- 10518986 TI - Axon guidance at the central nervous system midline. AB - A key feature of the central nervous system of most higher organisms is their bilateral symmetry about the midline. The specialised cells that lie at the midline have an essential role in regulating the axon guidance decisions of both neurons that project axons across the midline and those that project on one side. The midline cells produce both attractive and repellent short- and long-range signals to guide axonal growth. The axons themselves express specific receptors that can be dynamically regulated in response to midline-derived signals. In this way, axons extend toward or away from the midline and those that do cross change their behaviour to respond to longitudinal signals on the contralateral side. PMID- 10518987 TI - Glia as mediators of growth cone guidance: studies from insect nervous systems. AB - Growth cones experience many different cues in their journey to their final target. They can respond to a variety of attractive and repulsive cues that can be secreted or cellular. These cues are generated by a wide range of cell types. One subset of cells that play an important role in growth cone guidance are glial cells. Glia secrete guidance cues and express cellular cues on their surface that guide axonal outgrowth. In doing so, glia can act as intermediate targets in growth cone guidance, a process that is conserved between vertebrate and invertebrate nervous systems. Recent work in grasshopper, Drosophila and moth nervous system development has underscored the importance of the instructive role glia play during axonal outgrowth. PMID- 10518988 TI - Postembryonic sensory axon guidance in Drosophila. AB - The peripheral sensory system of the Drosophila adult has been used for the genetic analysis of axon guidance because of its accessibility for experimental manipulation and mutant screens. Wing, leg, antenna, or eye sensory axons are able to pathfind normally under different perturbations, indicating that sensory axon guidance is a highly canalized process. Similarly to other model systems, sensory growth cones seem to use multiple, simultaneous cues for guidance. In addition, sensory axons from peripheral structures seem to be capable of using alternative substrates for pathfinding. Developmental regulation could account for the high stability of axon guidance under experimental and natural perturbation conditions. Despite this flexibility, functional characterization of genes involved in sensory axon guidance is being carried out in situations where there appears to be less system redundancy. PMID- 10518989 TI - 'Identify' and 'lock in': molecular integration during synaptic target recognition. AB - Synaptic target recognition is a complex molecular event. In a differentiating presynaptic terminal, relatively 'rare' molecules first detect the cell identity of the synaptic target. Subsequently, many 'common' molecules continue the process of synaptogenesis. We present a theoretical framework for understanding synaptic target recognition and discuss the features of its molecular components and their integration, drawing on the rapid progress made in recent studies. PMID- 10518990 TI - Intermediate filament assembly: temperature sensitivity and polymorphism. AB - Intermediate filament (IF) proteins are encoded by a large multigene family and form polymers with a uniform diameter of approximately 10 nm. However, although the cytoplasmic representatives all confirm to a unit-type structural principle leading to the formation of extended coiled coils, it is becoming increasingly clear that subunit arrangements and physical properties vary among the different filaments. Thus, the intricate tissue-specific expression pattern of individual IF proteins (especially, their co-expression with other members of the IF protein family or with IF-associated proteins to form obligatory heteropolymers) points to distinct functions acquired during evolution relevant to cellular homeostasis in various tissues. PMID- 10518991 TI - Triplet repeat disorders: discussion of molecular mechanisms. AB - Comparison of the growing number of disorders known to be associated with triplet repeat expansions reveals both common features and a diversity of molecular pathways. Despite significant progress towards the characterization of proteins coded by the mutant genes, the complex nature of these disorders requires identification of all molecular components of the triplet repeat pathways. In this brief review we will discuss recent progress in determining the molecular mechanisms of disorders with unstable trinucleotide mutations. PMID- 10518993 TI - The effect of temperature and protein synthesis on the renaturation of firefly luciferase in intact H9c2 cells. AB - A mild increase in temperature that does not exert an effect on tolerance development or synthesis of heat shock proteins (Hsps) in control cells can stimulate these processes when applied to cells that have previously been heat shocked. To study the underlying mechanism of this effect, H9c2 cells were stably transfected with the gene encoding firefly luciferase (Luc). Heat-shock-induced inactivation of Luc and its subsequent reactivation is frequently used as a model for cellular protein denaturation and renaturation. Luc reactivation was determined following a damaging heat shock (43 or 44 degrees C for 30 min) in cells that were subsequently exposed to either control temperatures (37 degrees C) or various mild hyperthermic conditions (from 38.5 to 41.5 degrees C for 1 h). To prevent changes in Luc activity consequent to new synthesis of Luc, Luc reactivation was monitored in the presence of cycloheximide, an inhibitor of protein synthesis. The results showed that reactivation of Luc was inhibited when heat-treated cells were post-treated under mild hyperthermic conditions. The observed increase in Hsp synthesis under mild hyperthermic post-heat shock conditions therefore appears to be the result of an increase in the period during which denatured proteins are present. In addition, we studied Luc reactivation in the absence of protein synthesis inhibitors. This condition led to much higher Luc activity. By estimating half-life times of Luc, the contribution of new Luc synthesis in this recovery could be determined, and only partially explained the observed increase in Luc reactivation after heat shock. Thus the synthesis of other proteins must be important for the renaturation of heat-damaged proteins. PMID- 10518992 TI - Synaptogenesis in hippocampal cultures. AB - Hippocampal cultures offer unique advantages for the study of neuronal development and synaptogenesis. Studies performed on this model enabled dissection of the temporal sequence of events which lead to the differentiation of pre- and postsynaptic compartments. PMID- 10518994 TI - The possible role of isoforms of cytochrome c oxidase subunit VIa in mammalian thermogenesis. AB - A single cDNA of cytochrome c oxidase subunit VIa was characterised from liver, heart and the thermogenic organ of the partially endotherm tuna fish. The amino acid sequence revealed high identity with subunit VIa from carp and trout, but low identity to subunits VIaL (liver type) and VIaH (heart type) of mammalian cytochrome c oxidase. In reconstituted cytochrome c oxidase from bovine heart, the H+/e- stoichiometry is decreased from 1.0 to 0.5 at high intraliposomal ATP/ADP ratios via exchange of bound ADP by ATP at the matrix domain of the transmembraneous subunit VIaH. Reconstituted cytochrome c oxidase from bovine liver and kidney, containing subunit VIaL, revealed H+/e- ratios below 0.5, independent of the ATP/ADP ratio. The results suggest the evolution of three types of subunit VIa. Subunits VIaH and VIaL are postulated to participate in mammalian thermogenesis. PMID- 10518995 TI - Cholinergic, monoaminergic and glutamatergic changes following perinatal asphyxia in the rat. AB - Perinatal asphyxia (PA) is considered to lead to a variety of brain disorders including spasticity, epilepsy, mental retardation, and minimal brain disorder syndromes and may form the basis for psychiatric and neurodegenerative diseases later in life. We examined markers for neuronal transmission involved in the pathomechanisms of PA and candidates as mediators for long-term sequelae. We tested tyrosine hydroxylase (TH) and the vesicular monoamine transporter (VMAT) representing the monoaminergic system, the vesicular acetylcholine transporter (VAChT), and the excitatory amino acid carrier 1 (EAAC1), a neuronal subtype of the glutamate transporter, using immunohistochemistry on brain sections of rats subjected to graded PA. Three months following the asphyxiant insult immunoreactive (IR)-TH was decreased in striatum, hippocampus, thalamus, frontal cortex, and cerebellum; IR-VMAT was increased, and IR-VAChT was decreased in striatum. IR-EAAC1 glutamate transporter was increased in frontal cortex. The cholinergic, monoaminergic, and glutamatergic changes, still observed 3 months after the asphyxiant insult, may reflect their involvement in the pathomechanisms of PA and indicate mechanisms leading to long-term complications of PA. The variable consequences on the individual markers in several brain regions may be explained by specific susceptibility of cholinergic, monoaminergic, and glutamatergic neurons to the asphyxiant insult. PMID- 10518996 TI - Microtubule organization by the budding yeast spindle pole body. AB - In budding yeast microtubule organizing functions are provided by the spindle pole body (SPB), a multi-layered structure that is embedded in the nuclear envelope throughout the cell cycle. The SPB organizes the nuclear and cytoplasmic microtubules which are spatially and functionally distinct. Microtubule formation in yeast requires the Tub4p-complex, containing the gamma-tubulin Tub4p, and two additional proteins, the SPB components Spc97p and Spc98p. The Tub4p complex assembles in the cytoplasm and is then anchored to the sides of the SPB which organize microtubules. This is achieved by the binding of Spc97p and Spc98p to so called gamma-tubulin complex binding proteins (GTBPs) at the SPB. Spc72p is the yeast GTBP at the cytoplasmic side of the SPB, while Spc110p is the nuclear GTBP. Both GTBPs control the number of Tub4p complexes associated with the SPB and thereby the number of microtubules formed. In addition, the GTBPs may regulate the activity of the Tub4p complex. Homologues of Spc97p and Spc98p have been identified from yeast to mammalian cells and these are also part of gamma-tubulin complexes, suggesting that these related proteins may also interact with GTBPs at the centrosome. Candidates for GTBPs have been identified in mammalian and insect cells. PMID- 10518997 TI - Life cycles of yeast spindle pole bodies: getting microtubules into a closed nucleus. AB - The spindle pole body (SPB) is the principal microtubule organizing center of budding and fission yeast. We have examined SPBs and their associated microtubules from both organisms, using electron microscopy and three-dimensional reconstruction techniques, to identify the structural changes that accompany progression through the cell cycle. In this report, we compare these changes in the two kinds of yeasts and present a model for how microtubules get into a closed nucleus. PMID- 10518998 TI - Centrosomal and non-centrosomal microtubules. AB - While microtubule (MT) arrays in cells are often focused at the centrosome, a variety of cell types contain a substantial number of non-centrosomal MTs. Epithelial cells, neurons, and muscle cells all contain arrays of non-centrosomal MTs that are critical for these cells' specialized functions. There are several routes by which non-centrosomal MTs can arise, including release from the centrosome, cytoplasmic assembly, breakage or severing, and stabilization from non-centrosomal sites. Once formed, MTs that are not tethered to the centrosome must be organized, which can be accomplished by means of self-organization or by capture and nucleation of MTs where they are needed. The presence of free MTs requires stabilization of minus ends, either by MT-associated proteins or by an end-capping complex. Although some of the basic elements of free MT formation and organization are beginning to be understood, a great deal of work is still necessary before we have a complete picture of how non-centrosomal MT arrays are assembled in specific cell types. PMID- 10518999 TI - Microtubule release and capture in epithelial cells. AB - Many differentiated cells including polarised epithelial cells display a non radial, apico-basal microtubule array. In some cells the centrosome disassembles and new nucleating sites are created at more appropriate locations. In others the centrosome remains, but relatively few microtubules radiate from it's immediate environs. Instead, the majority of the microtubule minus-ends are associated with apical cell surface sites. Centrosomal microtubule release and capture is evidently a mechanism exploited by some polarised epithelial cells as a means of producing non-centrosomal, apico-basal microtubule arrays. This involves microtubule nucleation at the centrosome, release and subsequent translocation and capture at the apical sites. Two functionally distinct centrosomal complexes dedicated to the control of microtubule nucleation and anchorage have been suggested to be essential and universal features of all centrosomes. The centrosomal proteins ninein and R2 are potential microtubule anchoring proteins and their discovery has exciting implications for centrosomal organisation and microtubule positioning in cells. PMID- 10519000 TI - Centrosome structure and microtubule nucleation in animal cells. AB - Genetic studies in the budding yeast have led to the molecular characterization of gamma-tubulin associated proteins and to the identification of orthologues in animal cells. While the gamma-tubulin complex is more complex in animal cells than in budding yeast, its function is probably maintained throughout evolution. In this review we discuss some of the possible regulations of the nucleation activity in the light of the centrosome structure. A potential cross-talk between microtubule nucleation and centrosome duplication is suggested by some, still scarce, data. PMID- 10519001 TI - The centrosome cycle in syncytial Drosophila embryos analyzed by energy filtering transmission electron microscopy. AB - We have investigated the centrosome cycle in Drosophila syncytial embryos at the ultrastructural level by using a transmission electron microscope equipped with an electron energy filtering device (Omega filter). This new technique allows the study of uncontrasted thick sections with a high resolution. We have been able to characterize two classes of filamentous structures in the centrosomal apparatus that were not detectable on ultrathin sections. These new filamentous structures are: 1) a very orderly lattice that connects the two centrioles during mitosis; and 2) a fibrogranular connection between the centrosome and the nuclear envelope. The intercentriolar linkage could be involved in the precise timing of separation of the centrioles during late anaphase. The centrosome-nuclear envelope connection probably prevents the loss of centrosomes in this syncytial environment, and ensures the proper migration of the centrosomes along the surface of the nucleus. This connection may also couple the nuclei to the cytoskeleton, thus allowing their migration and their anchorage to the cortex at the blastoderm stage. This thick section analysis has also allowed us to precisely reconstitute the centrosome cycle. From their separation at telophase and throughout most of interphase, centrosomes are composed of a single centriole. We conclude that in the early Drosophila embryo there is an unusual delay between the separation of the parent centrioles and their duplication. This leaves a surprisingly short time to assemble a daughter centriole. PMID- 10519002 TI - Condensation-decondensation of the gamma-tubulin containing material in the absence of a structurally visible organelle during the cell cycle of Physarum plasmodia. AB - Genetic evidence has shown the presence of a common spindle pole organiser in Physarum amoebae and plasmodia. But the typical centrosome and mitosis observed in amoebae are replaced in plasmodia by an intranuclear mitosis devoid of any structurally defined organelle. The fate of gamma-tubulin and of another component (TPH17) of the centrosome of Physarum amoebae was investigated in the nuclei of synchronous plasmodia. These two amoebal centrosomal elements were present in the nuclear compartment during the entire cell cycle and exhibited similar relocalisation from metaphase to telophase. Three preparation methods showed that gamma-tubulin containing material was dispersed in the nucleoplasm during interphase. It constituted an intranuclear thread-like structure. In contrast, the TPH17 epitope exhibited a localisation close to the nucleolus. In late G2-phase, the gamma-tubulin containing elements condensed in a single organelle which further divided. Intranuclear microtubules appeared before the condensation of the gamma-tubulin material and treatment with microtubule poisons suggested that microtubules were required in this process. The TPH17 epitope relocalised in the intranuclear spindle later than the gamma-tubulin containing material suggesting a maturation process of the mitotic poles. The decondensation of the gamma-tubulin material and of the material containing the TPH17 epitope occurred immediately after telophase. Hence in the absence of a structurally defined centrosome homologue, the microtubule nucleating material undergoes a cycle of condensation and decondensation during the cell cycle. PMID- 10519003 TI - Control of centrosome reproduction: the right number at the right time. AB - It is of great importance for the cell to precisely coordinate the doubling of the interphase centrosome with nuclear events during the cell cycle and limit the number of centrosomes it contains at the onset of mitosis to two and only two. The penalties for mistakes are abnormal spindle assembly, inappropriate chromosome distribution, and consequently, genomic instability. We review the functional properties of the mechanisms that control when the centrosome duplicates in the cell cycle and the controls for centrosome copy number. We look to limits that are intrinsic to the centrosome itself and controls imposed by cell cycle linked changes in cytoplasmic conditions. Control of centrosome reproduction is exercised at both levels. PMID- 10519004 TI - Mechanisms of genetic instability revealed by analysis of yeast spindle pole body duplication. AB - Aneuploidy and polyploidy are commonly observed in transformed cells. These states arise from failures during mitotic chromosome segregation, some of which can be traced to defects in the function or duplication of the centrosome. The centrosome is the organizing center for the mitotic spindle, and the equivalent organelle in the budding yeast, Saccharomyces cerevisiae, is the spindle pole body. We review how defects in spindle pole body duplication or function lead to genetic instability in yeast. There are several well documented instances of genetic instability in yeast that can be traced to the spindle pole body, all of which serve as models for genetic instability in transformed cells. PMID- 10519005 TI - Centrosomes and cancer. AB - The centrosome functions as the major microtubule organizing center (MTOC) of the cell and as such it determines the number, polarity, and organization of interphase and mitotic microtubules. Cytoplasmic organization, cell polarity and the equal partition of chromosomes into daughter cells at the time of cell division are all dependent on the normal function of the centrosome and on its orderly duplication, once and only once, in each cell cycle. Malignant tumor cells show characteristic defects in cell and tissue architecture and in chromosome number that can be attributed to inappropriate centrosome behavior during tumor progression. In this review, we will summarize recent observations linking centrosome defects to disruption of normal cell and tissue organization and to chromosomal instability found in malignant tumors. PMID- 10519006 TI - The Xenopus laevis centrosome aurora/Ipl1-related kinase. AB - The cDNA encoding the protein kinase pEg2 was originally cloned through a differential screening performed during the early development of Xenopus laevis. pEg2 orthologues were found in various organisms and were classified in a new family of oncogenic mitotic protein kinases named 'aurora/Ipl1-related kinases' after the Drosophila melanogaster gene aurora and the Saccharomyces cerevisiae gene Ipl1. The catalytic activity of pEg2 is necessary for the mitotic microtubule spindle formation in Xenopus laevis egg extracts. The addition of a dominant negative form of pEg2 to in vitro spindle assembly assays leads to monopolar spindles generated by a defect of centrosome separation. In Xenopus cultured cells, pEg2 was confined around the pericentriolar material once centrosomes were duplicated. The centrosome localization does not depend on the presence of microtubules. However, in vitro, the protein binds to taxol stabilized microtubules independently of its kinase activity. During mitosis the location of the protein changes, in metaphase the kinase localizes on the microtubules at the poles of the mitotic spindle whereas it is not present on astral microtubules. This localization persists until the segregation of the chromosomes is completed. The presence of the kinase on the spindle may reveal another yet unknown function. PMID- 10519007 TI - Cell cycle-dependent localization of monoclonal antibodies raised against isolated Dictyostelium centrosomes. AB - The ultrastructure of the Dictyostelium centrosome is markedly different from that of the well known yeast spindle pole body and vertebrate centriole containing centrosome. It consists of a box-shaped, layered core structure surrounded by a corona with dense nodules embedded in an amorphous matrix. For further structural and biochemical analyses of this type of centrosome we used highly enriched isolated Dictyostelium centrosomes as an antigen to raise 14 new centrosomal monoclonal antibodies. Immunofluorescence microscopy and Western blot analysis revealed that at least 10 of them were directed against different antigens. Immunofluorescence microscopy also showed that the monoclonal antibodies fell into three different groups: A) antibodies localizing to the centrosome during the entire cell cycle; B) antibodies staining the centrosome mainly during mitosis; and C) antibodies labeling centrosome associated structures. All antibodies, except one, exhibited a cell cycle-dependent staining pattern underscoring the highly dynamic properties of the Dictyostelium centrosome. PMID- 10519008 TI - [Role of surgery in treatment of hyperthyroidism]. AB - Hyperthyroidism is treated with antithyroid drugs, radioactive iodine or surgery. The aim of surgery is to obtain long-lasting euthyroidism with a minimal risk of recurrence, secondary hypothyroidism, and complications involving the recurrent nerve or the parathyroids. Depending on the etiology, total or partial resection is indicated in light of these objectives. According to the literature, surgery is not used as first intention treatment for Grave's disease, its role and indications remaining a question of debate. However, surgery is the treatment of choice for toxic nodules, current consensus favoring extracapsular total lobectomy. For multinodular toxic goiter, three are several surgical indications: failure of medical treatment, relapse after drug withdrawal, specific clinical or socioeconomic situations requiring rapid efficacy. Total lobectomy associated with subtotal contralateral hemithyroidectomy appears to be the most appropriate procedure. Surgery may also be indicated for treatment of pregnancy-, diabetes-, or amiodarone-induced hyperthyroidism. PMID- 10519009 TI - [Semiology and etiology of anosmia: apropos of 306 patients]. AB - Chemosensory dysfunction is relatively common. This article describes a series of 306 patients who presented with anosmia. We divided olfactory disorders into those associated with interruption of the transport of stimulus and those associated with damage to either peripheral or central nervous system structures. Nasal and paranasal sinus disease (transport interruption) was found to be causative in 67% of patients presenting with anosmia. These patients are generally 45 years old, the loss of olfaction is progressive and associated with additional nasal symptoms. Upper respiratory infection was found to be causative in 18% of patients. They are generally older, with a mean age of 58 years and are predominantly female (78%). The loss of smell is sudden and anosmia is often accompanied by troublesome parosmias (50%). CT-scan is necessary for the evaluation of a smell dysfunction. PMID- 10519010 TI - [Hypoglossal nerve in its intralingual trajectory: anatomy and clinical implications]. AB - There is little literature on the intralingual trajectory of the hypoglosal nerve. We performed an anatomical dissection on 6 cadavers and completed our study with histological examinations. The 12th cranial nerve enters the lower part of the tongue laterally, reaching the anterior border of the hypoglossal muscle where it follows the ascending lingual artery medially to terminate anteriorly to the lingual V. Its terminal branches spread out horizontally in each half of the tongue. There is a paramedial branch, found in all cases, which projects downwardly, posteriorly and medially at the basilingual portion of the genioglossal muscle. These anatomic findings indicate that basiglossectomy removing the entire base of the tongue can be performed without functional sequelae. A certain degree of somatotopy is also found with specific fibers reaching the protractor and retractor muscles. This nerve distribution supports attempts at selective electrical stimulation of the hypoglossal nerve with the aim of dilating the upper airways in patients with sleep apnea syndrome. PMID- 10519011 TI - [Congenital cholesteatoma of the ear in the child. Clinical, follow-up and therapeutic analysis of a series of 34 cases]. AB - Long a subject of debate, congenital cholesteatomas of the middle ear appear to be a specific clinical entity different from the much more frequent classical acquired cholesteoma. Characteristic features of congenital cholesteatomas are young age at diagnosis, typical peroperative presentation, satisfactory mastoid air cells in almost all cases, and associated congenital malformations, which may involve the otology system or not. Diagnosis is a difficult task due to the long latency period with no clinical manifestations. These congenital cholesteatomas appear to be more aggressive in a mastoid with functioning air cells. Thus open excision does not appear to be appropriate and should be reserved for selected cases. For us, the closed technique with two procedures is more adapted but requires good cooperation with the family. The risk of recurrence is however significant and at least comparable to that of acquired cholesteatomas in children. Follow-up should be persuade as long as possible. Functional results have been encouraging even though ossicular destruction is frequent. The quality of the auditory tube appears to be a determining factor. PMID- 10519012 TI - [Rehabilitation of the paralyzed face by hypoglossal-facial nerve anastomosis. An analysis of 7 cases]. AB - After facial nerve injury, in cerebello-pontine tumors surgery, hypoglossal facial anastomosis is the most common procedure, to rehabilite a paralysed face, if direct facio-facial graft is not possible. This procedure must be done, in a second time, during the next year and followed with a specific reeducation. In seven patients operated between 1985 and 1996, we performed clinical evaluation and electrophysiological examination. The best evaluation is the clinical evaluation using the G. Freyss's facial testing. Best results are seen in early, specific and continued reeducation. All our patients have a good recovery of facial nerve function, but clinical examination and electrophysiological results are not correled with an objective video performance. The management of such patients needs efficient oto-neurosurgical team and specific trained physiotherapists. PMID- 10519013 TI - [Invasive aspergillosis sinusitis in patients infected by the human immunodeficiency virus]. AB - Invasive aspergillus sinusitis invasive is a rare, life threatening infection observed in immunocompromised patients. We report three cases of patients with AIDS. Diagnosis was based on surgical biopsy specimen performed on patients with unilateral sinusitis with facial pain and osteolysis on CT scanning, associated in one patient with neurological disorders. One patient with frontal sinusitis and meningeal involvement died despite therapy. One patient with limited maxillary sinusitis who underwent surgical satisfactory resection and antifungal therapy was successfully treated without relapse 12 months later. One patient with orbital and cavernous sinus extension of ethmoiditis was successfully cured with antifungal therapy by itraconazole. Our results confirm the necessity of early diagnosis when clinical and CTscanning are suggestive and the curability of aspergillus sinusitis. PMID- 10519014 TI - [Common variable immunodeficiency: one or multiple illnesses? 3 clinical cases]. AB - Common variable immunodeficiency (CVID) is a major antibody-deficiency syndrome, associated with increased risk of bacterial infection, as well as autoimmune and granulomatous disease. The clinical and immunological features are heterogeneous. This heterogeneity is expressed by the case reports of three selected patients. These observations will be discussed, with reference to a recent classification of CVID distinguishing four different clinical entities: i) CVID presenting with clinical and immunological features of X-linked agammaglobulinemia; ii) CVID presenting with clinical and immunological features of X-linked hyper-IgM syndrome; iii) CVID associated with systemic granulomatous disease; and iiii) CVID associated with autoimmune manifestations. PMID- 10519015 TI - [Opsoclonus-myoclonus syndrome in the adult. 4 cases]. AB - Four cases of opsoclonus in adults are reported. In all cases, it was associated with myoclonus and cerebellar syndrome. In 2 cases, the clinical and biological picture was suggestive of an infectious origin, and the patients recovered. The last cases are paraneoplastic opsoclonus revelating a pulmonary carcinoma and a breast cancer. The two patients died from metastasis after a transient improvement of ocular disorder. The pathophysiology of opsoclonus is discussed. PMID- 10519016 TI - [Joint involvement in AL amyloidosis]. AB - This review is aimed at defining the frequency, the clinical, biological and radiological presentation and the therapeutic possibilities for AL amyloid arthropathies. The frequency of AL amyloid arthropathy is estimated to be between 2 and 5%. There is usually a bilaterally progressive symmetrical arthritis of multiple joints (predominantly in the upper limb joints) with a chronic evolution. Neurologic or cutaneous symptoms are common. The analysis of synovial fluid sediments is a key test for the diagnosis of amyloid arthropathy. Patients with amyloid arthropathy must be screened for monoclonal gammapathies. The synovectomy is the best symptomatic treatment. A better knowledge of the pathogenesis of amyloid deposits should permit to improve therapeutical strategies. PMID- 10519017 TI - The spectrum of autoimmune thyroid diseases (AITD). PMID- 10519018 TI - Advances in the genetics and immunology of autoimmune polyglandular syndrome II/III and their clinical applications. PMID- 10519019 TI - Type 1 autoimmune polyglandular disease. PMID- 10519020 TI - Lymphocytic hypophysitis. A review of 145 cases. PMID- 10519021 TI - Immunologic approaches in the prevention and treatment of type 1 diabetes and endocrine autoimmune diseases. PMID- 10519023 TI - C reactive protein: a band mistaken as monoclonal gammapathy in acute inflammatory disease. PMID- 10519022 TI - [Methadone and new antiretroviral drugs. Results of therapeutic follow-up]. PMID- 10519024 TI - [Macro-aspartate aminotransferase: a benign biological anomaly to consider]. PMID- 10519025 TI - [Familial Mediterranean fever: discovery of the responsible gene and the implications]. PMID- 10519026 TI - [Imported pediatric malaria in Marseille]. AB - Imported malaria is frequently observed in pediatric practices within geographical areas which have a migrant population. MATERIAL AND METHODS: All the pediatric malaria cases of a university children's hospital (Marseilles, southern France) had been studied retrospectively. The period of the study was from January 1987 to December 1997. Inclusion criteria were based on clinical diagnosis criteria established by WHO. RESULTS: Three hundred and fifteen clinical cases were observed. Ninety-nine percent were confirmed by blood smears. Eighty-six percent of the patients came from the archipelago of the Comoro Islands in the Indian Ocean. Twenty percent were not given chemoprophylaxis, and 77% of the patients with chemoprophylaxis were not compliant. Fever (92%), splenomegaly (61%), vomiting and/or diarrhea (50%) were frequently observed. Neurological signs (23%), especially headaches (15%), were noted. The causative species was Plasmodium falciparum in 76%; coinfections with two species were observed in 9%. Halofantrine was commonly used for therapy (64%), but relapses were noted with this drug. No death was observed during the study. DISCUSSION: Imported pediatric malaria is rare in France. Clinical signs may lead to misdiagnosis when splenomegaly is not obvious, or when vomiting and/or diarrhea, cough or otitis occur. Diagnosis relies on blood smears. Curative medications are chloroquine or halofantrine, with special attention to heart troubles. Mefloquine is rarely used in children. Quinine is reserved for serious attacks. Concerning chimioprophylaxy, medical prescriptions should be adapted to the stay abroad, and patient compliance to medications could be improved. PMID- 10519027 TI - [Recombinant human erythropoietin in premature infants. Evaluation of a one year experience]. AB - Recently, recombinant human erythropoietin (rhEPO) has been claimed to diminish red blood cell transfusions in premature infants. After a year of experience, we investigated whether early rhEPO treatment would reduce the need for transfusion. PATIENTS AND METHODS: Fifty premature infants of gestational age < or = 32 weeks admitted to our NICU in 1997, received rhEPO 750 UI/kg/week from day 3 to 5 for six weeks. They were compared with 50 untreated controls admitted in 1996. RESULTS: The treatment and control groups did not differ for gestational age, weight at birth, CRIB score, and blood losses. We were not able to detect any difference in the number of transfused infants, and in the number of transfusions per infant until discharge. However, treated infants received significantly fewer transfusions per infant between day 16 and day 45 (0.42 +/- 0.67 vs. 0.8 +/- 0.99). Infants with a birth weight between 1,000-1,250 g received fewer transfusions in the EPO group. CONCLUSION: rhEPO treatment can be useful, but in association with other procedures: conservative transfusion criteria, minimization of phlebotomy losses and early iron supplementation. PMID- 10519028 TI - [Effects of puberty on the self-concepts of adolescents]. AB - AIM: This research was aimed at assessing the effects of pubertal maturation on the self-image of adolescents. METHODS AND PATIENTS: Three components of self image have been studied: body image (physical well-being, attractiveness and efficiency), perceived relations with peers of the opposite sex, and self-esteem. A sample of 159 French boys and girls aged from 11 to 16.3 years completed self report scales and a semantic differential. Their pubertal status was measured by school physicians in terms of Tanner stages. RESULTS: Correlational patterns indicated stronger reciprocal links between the different components of self image in girls than in boys. Results from regression analyses confirmed that the impact of pubertal maturation on self-image is qualitatively very different between the two sexes. PMID- 10519029 TI - [Home care of vaso-occlusive crisis in sickle cell disease in Togo]. AB - BACKGROUND: Many people with sickle cell disease manage their pain crises at home. This study aims to describe the home management of sickle cell pain by Togolese patients. PATIENTS AND METHODS: From July 1996 to April 1997, parents of children with sickle cell disease, and some adults with sickle cell disease living in rural and urban regions were interviewed about their home treatment habits during pain crises. RESULTS: A total of 165 patients with sickle cell disease (82 from urban and 83 from rural areas) were selected. The techniques most frequently used for pain management included salicylates (61.8%), paracetamol (37%), non-steroidal anti-inflammatory drugs (15.1%), vasodilators and pentoxifylline (5.4%). Only 4.2% of the patients mentioned adequate hydration. None used other antalgics (weak or strong opium derivatives). No difference was noticed between the treatment habits of rural regions and those of urban regions. CONCLUSION: In order to improve the quality of life of patients with sickle cell disease, information and awareness programs must be organized in order to establish a standard home pain management. Emphasis must be put on the use of salicylates and paracetamol at the correct dosage, the intake of abundant fluids, the easy use of analgesic of the second step of the Word Health Organization, and the systematic treatment of malaria which can induce pain crises. PMID- 10519030 TI - [Favorable outcome of a subdural hematoma diagnosed in utero]. AB - The subdural hematoma detected in utero is unusual but probably under-diagnosed and has a bad prognostic. CASE REPORT: A newborn had a subdural hematoma detected with a prenatal ultrasonography at 31 weeks' gestation, probably in keeping with the regular treatment of AAS. Diagnosis was possible through antenatal MRI at 34 weeks' gestation. The mother chose to follow her pregnancy. The operation at day 3 had no complications and neurological progress at 12 months was very good. CONCLUSION: A subdural hematoma detected in utero may have a good prognostic. Prenatal MRI is useful for the check-up. However, medical and ethical problems remain in these cases. PMID- 10519031 TI - [Long QTc interval complicating halofantrine therapy in 2 children with Plasmodium falciparum malaria]. AB - Halofantrine has been shown to be very effective against multiple drug resistant falciparum malaria. It is usually administered in children at 24 mg/kg at six hour intervals for three doses, and a second therapeutic course one week following the initial treatment is recommended. It is usually well tolerated. However, prolongation of the QT interval has been reported in adults receiving this drug for malaria. CASES REPORTS: Two children experienced a prolongation of the QTc interval while receiving halofantrine. The first child, aged two years, had a prolonged QTc interval (490 ms) six hours after the third administration, at the usual therapeutic dose. The second child, aged six years, had a normal QT interval (360 ms) after the first 24 mg/kg dose and had a prolonged QTc (450 ms) during the second course seven days later, 15 h after the last dose. In both cases, the QTc interval returned to normal values (below 440 ms) rapidly after the end of treatment. CONCLUSION: Cardiotoxic effects are felt to be dose dependant and young children may be particularly at risk due to pharmacological and cardiac immaturity. Therefore, guidelines for drug administration should be followed (administration in a child with an empty stomach, drug not recommended in combination with drugs known to prolong the QTc interval) and monitoring ECG in pediatric patients may be justified. The modalities of the second course in children, which is recommended by the manufacturer to travellers from non-endemic areas, should also be discussed. PMID- 10519032 TI - [Aseptic femur head osteonecrosis during treatment of acute lymphoblastic leukemia in the child]. AB - Avascular femoral head necrosis (AFN) is an uncommon complication of acute lymphoblastic leukemia (ALL) occurring in association with serious functional late effects. One of the many risk factors is high-dose corticosteroid therapy. CASE REPORT: Three children belonging to a series of 266 patients developed AFN. The diagnosis was not made immediately when X-rays were normal. In spite of the fact that treatment was begun as soon as possible, the three children had a difference in the length of their legs, with reduction of their walking perimeter and, in one case, an arthroplasty was necessary. CONCLUSION: If some patients treated for ALL limp or suffer when walking or when practising sports, the diagnosis of AFN is to be evoked. The diagnosis is not only based on simple X rays but also on magnetic resonance imaging, which is more sensitive and reveals lesions earlier. The treatment consists of immobilization of the hip, whether or not associated with surgical procedures. PMID- 10519033 TI - [Subphrenic abscess due to ectopic appendicitis]. AB - BACKGROUND: Idiopathic subphrenic abscesses are uncommon in children. Standard chest X-rays may provide the suspicion of this diagnosis. Initial percutaneous drainage of the collection is usually performed. Surgery is required when the underlying cause remains unknown. CASE REPORT: A 12-year-old boy presented signs of pulmonary abscess. Chest X-rays, ultrasonography and computed tomography established the diagnosis of a right subphrenic abscess, which was percutaneously drained. Surgery disclosed an ectopic appendiceal perforation. CONCLUSION: In children, appendicitis is the main etiology of primary or postoperative subphrenic suppurations. Ectopic appendicitis is an important predisposing factor to this complication. PMID- 10519034 TI - [Anti-rotavirus vaccinations]. AB - Rotavirus is the most common cause of severe acute diarrhea, responsible for 30 to 40 deaths of children each year in France. In order to decrease both mortality and morbidity, vaccines have been designed first from attenuated bovine strains, then from monovalent simian strains, and more recently from reassortant rhesus strains. The live tetravalent human-rhesus reassortant vaccine (RRT-TV) has been shown to be protective in the United States, Finland, and Venezuela despite different environments, in prospective double-blind studies. This vaccine, as the natural infection, decreases by 50% the risk of acute rotavirus diarrhea and by 70 to 100% the risk of severe diarrhea with dehydration. At the present time, its cost limits its use to developed countries. It was put on the market in the United States in October 1998. The challenge is now to make it available in developing countries. PMID- 10519035 TI - [Primary nephrotic syndrome in the black African child]. AB - Idiopathic nephrotic syndrome (INS) in black African children differs from that of children in temperate areas. The main differences are the high rate of corticosteroid non-responders and the low rate of minimal change glomerulopathy in black African children, possibly related to a racial factor. The identification of a high corticosensibility in certain African regions (Togo and Ghana) can lead to the identification of an ethnic factor. Further genetic studies should be carried out in order to provide a better approach to INS in Africa. PMID- 10519037 TI - [Radiology case of the month. A case of cystic tumor of the kidney]. PMID- 10519036 TI - [Prevention of vitamin D deficiency in the child and adolescent. I. Proposal and arguments for use of a decision tree]. AB - A decision-making table, using three questionnaires, is proposed to determine the vitamin D status in children and adolescents. The first questionnaire assesses the vitamin D endogenous synthesis, taking into account the sunlight exposure and the time interval out of the sun. The second questionnaire quantifies the vitamin D dietary intake within three levels: optimal, medium or low. In case of a medium dietary score, a third questionnaire evaluates the daily calcium intake, taking into account the fact that vitamin D metabolism is increased by a low calcium intake (under 400 mg/day). This decision-making table should enable the detection of children and adolescents at risk of vitamin D deficiency and requiring an adapted prophylaxis. Its accuracy will be assessed in prospective surveys. PMID- 10519038 TI - [Varicella vaccine]. AB - Varicella vaccine has been extensively studied in Japan, the United States and Canada since 1974. Its efficacy in healthy children is good, on the order of 94%. It also has a good efficacy in immunocompromised children, such as children treated for acute leukemia or malignant tumors. However, the vaccine has to be administrated during a 2-3 week transient suppression of chemotherapy. In the USA, varicella vaccination is recommended for receptive adults who are particularly exposed to varicella, such as health professionals working with immunocompromised patients. Benefits and cost analysis of a generalized varicella vaccination in children has shown that it would be beneficial due to the cost of parents' work leave during the children's illness. PMID- 10519039 TI - [The pediatrician managing a child with heart disease]. AB - The pediatrician can and must play a pivotal role in the care of children with heart disease. He must provide the preventive care and health supervision as for all children, provide advice to the patient and the family in regards to everyday life, monitor for potential complications or deterioration of the cardiac lesion, and finally play a central role for patients with multiple problems and a chronic illness. The specialist must actively support the generalist with his experience and provide advice for each particular patient. This collaboration should allow the best care to be provided for the patient and maintain the patient and family in an ideal context. PMID- 10519040 TI - [Granuloma annulare in the child]. AB - Granuloma annulare is a benign, common, inflammatory skin lesion of unknown etiology that is seen in both adults and children. The typical lesions are single or multiple small cutaneous papules with an annular distribution. The histology is consistent with an area of fibrinoid degeneration of collagen, surrounded by palisading histiocytes and inflammatory cells. There are four clinically distinct subtypes: localized, generalized, subcutaneous and perforating. Usually a spontaneous resolution is expected. Many medical treatments have been proposed but without evidence of efficacy. The association with insulin dependent diabetes is still being discussed. PMID- 10519041 TI - [The cost of a consultation...]. PMID- 10519042 TI - [Prevalence of growth delay in adolescents 10 to 19 years of age in a rural Togo region (east savanna and littoral area)]. PMID- 10519043 TI - Localization of L-glutamate and glutamate-like receptors at the squid giant synapse. AB - HPLC analysis of the amino acid contents of the second- and third-order giant fibres at the giant synapse in the stellate ganglion of the squid Loligo vulgaris shows that there are significantly higher amounts of L-glutamate and L-aspartate in the second-order (presynaptic) fibre than in the third-order (postsynaptic) fibre. Immunocytochemical staining of sections of the ganglion with an antibody raised against L-glutamate produces specific positive staining in the synaptic region of the second-order fibre. In contrast, staining with antibodies raised against glutamate-receptors (mammalian GluR1 with GluR2/3) produces positive staining in the third-order fibre at the postsynaptic region. These data provide further evidence for the hypothesis that L-glutamate is an excitatory transmitter at the giant synapse. PMID- 10519044 TI - The stabilization of ferrous iron by a toxic beta-amyloid fragment and by an aluminum salt. AB - Aluminum is a recognized neurotoxin in dialysis encephalopathy and may also be implicated in the etiology of neurodegenerative disease, particularly Alzheimer's disease. Alzheimer's disease is suspected to be associated with oxidative stress, possibly due to the pro-oxidant properties of beta-amyloid present in the senile plaques. The underlying mechanism by which this occurs is not well understood although interactions between amyloid and iron have been proposed. The presence of low molecular weight iron compounds can stimulate free radical production in the brain. This study provides a possible explanation whereby both aluminum and beta-amyloid can potentiate free radical formation by stabilizing iron in its more damaging ferrous (Fe2+) form which can promote the Fenton reaction. The velocity, at which Fe2+ is spontaneously oxidized to Fe3+ at 37 degrees C in 20 mM Bis-Tris buffer at pH 5.8, was significantly slowed in the presence of aluminum salts. A parallel effect of prolongation of stability of soluble ferrous ion, was found in the presence of beta-amyloid fragment (25-35). Ascorbic acid, known to potentiate the pro-oxidant properties of iron, was also capable of markedly stabilizing ferrous ions. PMID- 10519045 TI - Noradrenaline and mixed alpha 2-adrenoceptor/imidazoline-receptor ligands: effects on sodium intake. AB - The effect of noradrenaline, and mixed ligands to alpha 2-adrenoceptors (alpha 2 AR) and imidazoline receptors (IR), injected intracerebroventricularly (i.c.v.), on sodium intake of sodium depleted rats, was tested against idazoxan, a mixed antagonist ligand to alpha 2-AR and IR. The inhibition of sodium intake induced by noradrenaline (80 nmol) was completely reversed by idazoxan (160 and 320 nmol) injected i.c.v. The inhibition of sodium intake induced by mixed ligands to alpha 2-AR and IR, UK14,304, guanabenz and moxonidine, was antagonized from 50 to 60% by idazoxan i.c.v. The results demonstrate that noradrenaline, a non-ligand for IR, acts on alpha 2-AR inhibiting sodium intake. The possibility that either alpha 2-AR or IR mediate the effect of mixed agonists on sodium intake remains an open question. PMID- 10519046 TI - Properties of excitatory amino acid transport in the human U373 astrocytoma cell line. AB - In this study, we describe the presence of Na(+)-dependent high-affinity L glutamate transport activity in the human U373 astrocytoma cell line. U373 cells exhibited a robust accumulation of L-glutamate which was predominantly (85%) extracellular Na(+)-dependent. Kinetic analysis of this transport activity revealed that the uptake followed first-order Michaelis-Menten kinetics and was high-affinity in nature. The kinetic parameters estimated by Eadie-Hofstee transformation of the saturable uptake were 37.3 +/- 5.1 microM for K(m) and 0.13 +/- 0.02 nmol min-1 mg-1 protein for Vmax. A total of 14 known inhibitors of high affinity L-glutamate transport were examined for their abilities to inhibit L glutamate uptake by U373 cells. Three compounds, kainate (KA), dihydrokainate (DHK) and alpha-aminoadipic acid produced less than 30% inhibition at 1 mM. The lack of effect of both KA and DHK indicates that the predominant astroglial L glutamate transporter EAAT2 (excitatory amino acid transporter 2) does not contribute to the uptake activity present in these cells. The rank order of inhibitory potency for the remaining 11 compounds tested was L-cysteine sulphinate = L-CCG-III = L-cysteate = L-aspartate = threo-beta-hydroxyaspartate > trans-PDC > D-aspartate = MPDC > beta-glutamate > L-CCG-IV = L-aspartate-beta hydroxamate. Pre-treatment of U373 cells with phorbol ester for 30 min resulted in a 56% decrease in L-glutamate uptake and this effect was blocked in a concentration-dependent manner by the PKC inhibitor bisindolylmaleimide I. Expression of L-glutamate transporters by U373 cells was examined by reverse transcriptase polymerase chain reaction (RT-PCR) and Western analysis. Transcripts for both the EAAT1 and EAAT3 transporter subtypes were detected but not for EAATs 2, 4, and 5. Immunoblot analysis confirmed the presence of EAAT3 protein, however, we were unable to detect EAAT1 protein. In conclusion, the Na(+)-dependent high-affinity L-glutamate transport into human U373 astrocytoma cells appears to be mediated predominantly by the EAAT3 subtype. PMID- 10519047 TI - Cellular dynamics of corneal wound re-epithelialization in the rat. II. DNA synthesis of the ocular surface epithelium following wounding. AB - To determine the fate of ocular surface epithelial cells in response to corneal injury, epithelial cells undergoing DNA synthesis were labeled at discrete time points following abrasion. A 3-mm diameter epithelial defect was made in the center of the rat cornea. One hour prior to sacrifice, animals received an intraperitoneal injection of [3H]-thymidine. DNA synthesis in the regenerating epithelium was of low abundance (< 2%). The undamaged corneal epithelium had a labeling index (LI) that was markedly elevated from unwounded specimens in the first 48 h after abrasion. The LIs of suprabasal cells were increased at 24 h and 36 h; these cells were believed to be displaced basal epithelial cells. Basal and suprabasal cells of the limbus and conjunctiva exhibited increases in LIs from unwounded subjects at singular timepoints (within 24 h of injury). These results, using rigorous statistical analysis, show that DNA synthesis is not impeded--but rather often accelerated--following denuding of the corneal epithelium. Re epithelialization is not dependent on DNA synthesis in the regenerating region, but appears to be related to an increase in DNA synthesis (along with centripetal migration) in the adjacent, undamaged epithelium. The increase in DNA synthesis occurring in the limbus and conjunctiva does not directly supply cells to the regenerating region within the 72 h required for epithelial restitution, but is conjectured to serve in replenishing ocular surface epithelial cells used in the repair process. PMID- 10519048 TI - Sequence of neurodegeneration and accumulation of phosphorylated tau in cultured neurons after okadaic acid treatment. AB - Within neurofibrillary tangles and dystrophic neurites of Alzheimer's disease (AD), the cytoskeletal protein tau is abnormally hyperphosphorylated. In the present study, we examined the effect of okadaic acid (OA), a protein phosphatase inhibitor, in rat cultured neurons. Low concentrations of OA induce degeneration of neurites, rounding of cell bodies, detachment from the substratum, and eventual neuronal death. During OA-induced degeneration, SMI-31 immunoreactivity became punctate in neurites at 6 h after OA treatment, and over time, accumulated in cell bodies and dystrophic neurites. Hyperphosphorylation of tau and marked loss of MAP-2-positive dendrites occurred after 6 h of treatment with OA. Thereafter, AT-8 and PHF-1 immunoreactivity accumulated in cell bodies and subsequently appeared in distal axon-like neurites. These results demonstrate that OA treatment induced hyperphosphorylation of tau and preferential dendritic damage, with subsequent accumulation of phosphorylated tau in cell bodies and dystrophic axon-like neurites. OA-induced neurodegeneration may provide a useful model to study AD. PMID- 10519049 TI - Single unit activity during a Go/NoGo task in the "prefrontal cortex" of pigeons. AB - Single unit activity was recorded during a delayed auditory/visual Go/NoGo task from the neostriatum caudolaterale (NCL) of pigeons, a multimodal associative avian forebrain structure comparable to the prefrontal cortex (PFC). The animals were trained to mandibulate (to open their beak) during the Go period after which they received a drop of water as reward. Neuronal activity changes were observed during the delay period (DELAY) between auditory and visual stimulation, to the onset of the visual stimulus or to the delivery of the reward. In some neurons, responses were related to the behavioral significance of the stimulus such that the neuronal activity was statistically different between Go and NoGo trials. Moreover, some units anticipated the upcoming reward or changed their firing frequency in a correlated manner prior to beak movements. These neuronal activity patterns suggest that the NCL provides a neural network that participates in the integration and processing of external stimuli in order to generate goal directed behavior. PMID- 10519050 TI - Hyperthermia-enhanced serotonin (5-HT) depletion resulting from D-fenfluramine (D Fen) exposure does not evoke a glial-cell response in the central nervous system of rats. AB - D-Fen-induced hyperthermia has been shown to coincide with an enhanced depletion of 5-HT and 5-hydroxyindole acetic acid (5-HIAA). Because these observations have relied on D-Fen exposure at multiple environmental temperatures, some have questioned the validity of the findings. Therefore, this experiment was designed to determine if the correlation between elevated body temperature and 5-HT depletion could be observed when D-Fen exposure occurred in one warm environment (28 degrees C) and to determine if a hyperthermia-enhanced glial-cell response could be evoked by D-Fen exposure. Hyperthermia-enhanced 5-HT and 5-HIAA depletion resulting from D-Fen exposure was dependent on body temperature during drug exposure. In the frontal cortex, 5-HT concentrations ranged from 3 to 45% of control values. Likewise, in the striatum and hippocampus, 5-HT concentrations ranged from 13 to 53% and 6 to 40%, respectively. The 5-HIAA concentrations had a wider range than the 5-HT concentrations for each brain region. In the frontal cortex, striatum and hippocampus, 5-HIAA ranged from 0 to 93%, 15 to 72% and 0 to 83% of control, respectively. In spite of the substantial reductions in 5-HT, there was no detectable glial-cell response. D-Fen-induced hyperthermia does not appear to cause generalized damage to neurons in the frontal cortex, striatum and hippocampus. PMID- 10519051 TI - Posttreatment with the immunosuppressant cyclosporin A in transient focal ischemia. AB - Cyclosporin A (CsA) reduces ischemic brain damage when administered in such a way that its penetration across the blood-brain barrier is enhanced. Since only pretreatment has previously been used in focal ischemia, the objective of the present study was to establish whether posttreatment is efficacious and to assess the window of therapeutic opportunity for CsA. To that end, CsA was given 5 min to 6 h after the start of reperfusion following 2 h transient ischemia, and infarct volume was assessed after 48 h by triphenyltetrazolium chloride staining. Attempts were made to circumvent the BBB to CsA by an intracerebral needle lesion, by an increase in the intravenous CsA dose, or by osmotic opening with intracarotid mannitol. The results were compared to those obtained with FK506. Intravenous CsA in a dose of 10 mg/kg failed to reduce infarct volume, unless preceded by a needle lesion. That procedure, and an increase in CsA dose to 50 mg/kg, reduced infarct volume to about 50% of control, but the higher dose had toxic side effects. The coupled intracarotid infusion of mannitol and CsA (10 mg/kg) was more efficacious, without overt side effects. However, mannitol proved dispensable since CsA alone reduced infarct volume to 30% of control, with a therapeutic window of 3-6 h. When given after 5 min of reflow, CsA reduced infarct volume to 10% of control and was clearly more neuroprotective than FK506. Possibly, this is because CsA blocks the mitochondrial permeability transition pore which is opened under adverse conditions. PMID- 10519052 TI - c-fos gene expression parallels auditory adaptation in the adult rat. AB - This study investigated the pattern of c-fos gene expression corresponding with auditory adaptation to novel sound. Using six groups of adult rats (naive control, 1 h, and 1, 2, 3, and 4 days of continuous stimulation), we quantified c fos expressing cells in the dorsal and ventral cochlear nuclei and found a 54 fold increase in 1 h following novel sound stimuli. The number of reactive cells decreased sharply within 24 h and nearly disappeared by 96 h. Our results reveal that c-fos gene expression in the adult rat is attenuated in parallel with the expected auditory adaptation to novel sounds indicating an association with auditory learning and memory. PMID- 10519053 TI - Dystrophin is required for organizing large acetylcholine receptor aggregates. AB - Dystrophin is a cytoplasmic protein underlying the plasma membrane in normal skeletal muscle. Its absence leads to muscle degeneration as seen in Duchenne muscular dystrophy (DMD) and in mdx mice. One puzzling question in the study of dystrophinopathies is that in mdx muscles the neuromuscular junctions (NMJs) show little, if any, developmental defect, but morphological and functional abnormalities of NMJs are obvious after muscle damage and regeneration begin. This phenomenon leads us to hypothesize that dystrophin may be required for endplate maintenance and/or endplate remodeling in regenerating fibers. Here we show that the absence of dystrophin causes NMJ fragmentation in adult muscle fibers, and greatly reduces both spontaneous and agrin-induced acetylcholine receptor (AChR) clustering activities on cultured myotubes derived from satellite cells. The lower AChR clustering in mdx myotubes originates in the smaller size of each cluster and from a 72% reduction in the occurrence of large (> 10 micron 2) AChR clusters. Our results suggest dystrophin is involved in organizing small AChR clusters into large AChR aggregates during muscle regeneration, although it is not required for initiating the original AChR clustering activity. PMID- 10519054 TI - 7-Nitroindazole, a selective inhibitor of nNOS, increases hippocampal extracellular glutamate concentration in status epilepticus induced by kainic acid in rats. AB - The glutamate extracellular concentration is controlled by metabolic and neuronal pathways via release and uptake mechanisms. Stimulation of glutamate receptors induces neuronal nitric oxide (NO) release, which in turn modulates glutamate transmission. In this study, the influence of neuronally derived NO on hippocampal glutamate extracellular concentration was investigated in conditions of intense metabolic activation, i.e., during status epilepticus induced by systemic kainic acid (KA). Glutamate, arginine and citrulline concentrations were measured by microdialysis coupled to HPLC. Experiments were performed in conscious rats implanted with a microdialysis probe within the hippocampal CA3 area. Three groups were used: (1) rats treated with KA i.p. (12 mg/kg) and vehicle locally, via the microdialysis probe (n = 9); (2) rats given KA i.p. and a selective inhibitor of neuronal NO synthase, 7-nitroindazole (7-NI, 1.25 mM) locally (n = 13); (3) rats treated with saline i.p. and 7-NI locally (n = 7). Infusion of 7-NI or vehicle was performed throughout the second hour of status epilepticus. In groups 1 and 3, no significant modifications of extracellular glutamate, arginine and citrulline concentrations were measured. In group 2, the local application of 7-NI in the hippocampus during status epilepticus significantly increased extracellular glutamate and arginine concentrations, whereas citrulline concentration remained constant. The concomitant increases of extracellular glutamate and arginine concentrations under local 7-NI perfusion in seizure conditions, suggest that glutamate and arginine are linked in a common metabolic pathway and/or that glutamate is involved in the cross-talk between glia and neurons. A cerebrovascular effect of 7-NI which triggers glutamate release may also occur. PMID- 10519056 TI - Insular cortex lesions and taste aversion learning: effects of conditioning method and timing of lesion. AB - The specific role of insular cortex in acquisition and expression of a conditioned taste aversion was assessed using two different conditioning methods, which vary mode of taste delivery. Involvement of insular cortex in the induction of c-Fos-immunoreactivity in the nucleus of the solitary tract, a cellular correlate of the behavioral expression of a conditioned taste aversion, was also assessed. Electrolytic lesions of insular cortex blocked behavioral expression of a conditioned taste aversion and this was evident not only when lesions were placed prior to conditioning, but also when they were made after conditioning but before testing. In contrast to the effects on behavior, lesions did not completely block the c-Fos-immunoreactivity which accompanies re-exposure to the aversive taste. In addition, the blocking of behavioral evidence of aversion conditioning by cortical lesions was seen both in animals trained under an intraoral acquisition procedure and those trained with bottle-conditioning. This contrasts with previous work with amygdala lesions which showed that amygdala was absolutely necessary for taste aversions conditioned with the intraoral method but not for those conditioned using bottle presentation of the taste. Overall, these findings imply that the details of the neural circuitry involved in taste aversion learning, including its anatomical distribution, complexity and degree of redundancy, vary with the type of conditioning method employed. PMID- 10519055 TI - Opioid growth factor and organ development in rat and human embryos. AB - In addition to neurotransmission, the native opioid peptide, [Met5]enkephalin, is a tonically active inhibitory growth molecule that is termed opioid growth factor (OGF). OGF interacts with the zeta (zeta) opioid receptor to influence cell proliferation and tissue organization. We now identify OGF and the zeta receptor in embryonic derivatives including ectoderm, mesoderm, and endoderm of the rat on gestation day 20. Messenger RNA for preproenkephalin (PPE), the precursor of OGF, was detected in the developing cells, suggesting an autocrine production of this peptide. Acute exposure of the pregnant female to OGF resulted in a decrease in DNA synthesis in cells of organs representing all three germ layers, and did so in a receptor-mediated fashion. The influence of OGF was direct, as evidenced in organ culture studies. Blockade of endogenous opioid interaction using naltrexone (NTX) produced an increase in DNA synthesis, indicating the constitutive and functional nature of opioid activity on growth during prenatal life. Human fetal cells contained OGF and the zeta receptor. These data support the hypothesis that endogenous opioid modulation of organ development is a fundamental principle of mammalian embryogenesis, and that OGF has a profound influence on ontogeny. Irregularities in the role of opioids as growth regulators in relationship to the more than 500,000 newborns suffering from birth defects each year in the US needs to be examined. PMID- 10519057 TI - Corticosteroids alter G protein inwardly rectifying potassium channels protein levels in hippocampal subfields. AB - Corticosterone or cortisol, stress hormones in rat and human, respectively, alter neurotransmitter receptor-mediated responses in the brain. Corticosterone could alter these responses by modifying any component of the receptor-effector pathway. Many of these receptors are linked to guanine nucleotide regulatory proteins (G proteins) which, in turn, can activate second messenger systems and/or ion channels, such as G protein inwardly rectifying potassium channels (GIRK). The aim of these experiments was to determine whether corticosterone treatment altered the levels of GIRK proteins in rat hippocampus. Corticosterone treatment selectively altered the levels of GIRK1 and GIRK2 (measured on immunoblots) depending on the subfield of the hippocampus examined. These data lend credence to the hypothesis that corticosterone differentially alters neurotransmitter receptor-mediated responses dependent on the brain area. PMID- 10519058 TI - Peptide transport system-1 (PTS-1) for Tyr-MIF-1 and Met-enkephalin differs from the receptors for either. AB - Tyr-MIF-1 (Tyr-Pro-Leu-Gly-NH2) and Met-enkephalin share a saturable transport system (peptide transport system-1, PTS-1) across the blood-brain barrier but do not readily bind to each other's receptors. This information allows the unique opportunity to differentiate the transport protein(s) from the receptors for either peptide in brain endothelial cells. PTS-1 was studied in vitro by allowing radiolabeled Tyr-MIF-1 (125I-Tyr-MIF-1) to bind to the solubilized proteins of isolated murine brain microvessels in the presence or absence of potential inhibitors. Sephadex chromatography separated bound from free labeled peptide. The binding was saturable as shown by inhibition with increasing concentrations of unlabeled Tyr-MIF-1. 125I-Tyr-MIF-1 binding was not inhibited by an unrelated peptide or iodo-tyrosine. D-Tyr-MIF-1 had no effect, demonstrating the stereospecificity of the system. Met-enkephalin decreased the binding of 125I-Tyr MIF-1 to 84.4% of total, whereas Leu-enkephalin was without effect. Agonists for the mu, delta, and kappa opiate receptors did not change the binding, indicating that the proteins which bound to 125I-Tyr-MIF-1 were not endogenous opiate receptors. The results indicate that, in vitro, Tyr-MIF-1 binds to brain microvessel proteins with characteristics similar to PTS-1. PMID- 10519059 TI - Intraventricular lactate infusion attenuates the transactivational effects of the glucose antimetabolite, 2-deoxy-D-glucose, on hypothalamic vasopressinergic neurons. AB - Glucopenia stimulates neurohypophyseal arginine vasopressin (AVP) secretion and expression of the transcription factor, Fos, by paraventricular (PVN) and supraoptic (SON) magnocellular neurons. Recent studies suggest that central compensatory responses to glucose substrate imbalance are initiated by regulatory signals of periventricular origin. Since the glycolytic endproduct, lactate, is a preferred substrate for central neuronal respiration, we investigated whether intracerebroventricular (i.c.v.) infusion of this monocarboxylate fuel attenuates transactivational effects of glucoprivation on PVN and SON AVP neurons. Continuous intraventricular infusion of sodium lactate (1.0 or 10.0 microM/h) or vehicle was initiated before intraperitoneal (i.p.) injection of the glucose antimetabolite, 2-deoxy-D-glucose (2DG), or saline. Anterior hypothalamic tissue obtained 2 h after systemic injections was processed for colocalization of cytoplasmic AVP- and nuclear Fos-immunoreactivity (Fos-ir). Fos-ir was absent from the PVN and SON of rats treated by i.c.v. infusion of vehicle or either dose of lactate. Intraventricular administration of 10.0 microM lactate/h, but not the lower dose, significantly decreased mean numbers of colabeled AVP neurons in each structure in glucoprivic animals. These data suggest that Fos stimulus transcription cascade is activated in these cells by decreased central availability of this monocarboxylate fuel, and that cellular sources of regulatory signaling of lactate utilization exist within the periventricular CNS. PMID- 10519060 TI - Skeletal muscle injury induced by eccentric muscle action: muscle proteins as markers of muscle fiber injury. AB - Muscular overuse after high force eccentric muscle action is associated with structural damage of the contractile apparatus that can be observed as Z-line steaming and myofibrillar disruption. Mechanical stress is the major contributing factor for inducing muscle injury, which initiates a cascade of processes resulting in skeletal muscle damage. Disturbances in Ca2+ homeostasis with elevated intracellular [Ca2+] activates the nonlysomal cysteine protease, calpain. Calpain is assumed to play an important role in triggering the response of skeletal muscle protein breakdown, of inflammatory changes, and of regeneration processes in response to eccentric muscle action. The inflammatory response is attributed to changes in hormone and cytokine levels in blood and skeletal muscle. To assess the amount of skeletal muscle damage, plasma CK activity and plasma myoglobin levels have been widely used as markers for muscle injury. As the cytosolic proteins do not necessarily reflect the amount of structural damage, structurally bound proteins such as myosin heavy chains and troponin have been investigated. This paper briefly reviews the cascade of events causing muscle cell injury after unaccustomed eccentric muscle action and the potential of muscle proteins as markers of skeletal muscle damage. PMID- 10519061 TI - Free radicals and oxidative stress in exercise--immunological aspects. AB - Reactive oxygen (ROS) and nitrogen species (RNS) are continuously generated in the biological system and play an important role in a variety of physiological and pathological processes. There is evidence that physical exercise augments the generation of ROS/RNS. The present review discusses and compares insights into the generation and function of ROS/RNS such as superoxide, hydrogen peroxide, hypochloric acid, and nitric oxide released by leukocytes in response to exercise. Emphasis is placed on: (a) mechanisms and regulation of ROS/RNS generation in immunocompetent cells with respect to acute exercise and regular training; (b) damaging effects of ROS/RNS in terms of oxidative stress which may be causally involved in features such as exercise-induced damage to muscle tissue and leukocyte DNA; (c) (immuno-) modulating effects of ROS/RNS which include activation of transcription factors; (d) responses of antioxidant stress proteins to acute exercise and regular training; and (e) effects of antioxidants on exercise-induced changes in immune function. Available data suggests that ROS/RNS are involved in the inflammatory response to heavy exercise and therefore exert damaging effects. Several immune functions are influenced by actions of ROS/RNS, and it is hypothesized that adaption to regular training is also modulated in part by free radicals. Furthermore, regular training seems to reduce the capacity of leukocytes for oxidant release and leads to an adaptation of antioxidative mechanisms, which may contribute to a limitation of exercise-induced oxidative stress. PMID- 10519062 TI - Role of heat shock proteins in the exercise response. AB - Heat shock proteins (HSP) are a characteristic set of highly conserved proteins that are synthesized shortly after the organism is exposed to external stress, including physical activity. HSP help to maintain cellular homeostasis and protein conformation, reactivate denatured or malformed proteins, and provide "housekeeping," translocase, and chaperone functions. Some of the conditions known to elicit the cellular stress reaction are similar to those experienced by cells in response to physical exercise. Hyperthermia, ischemia, oxidative, cytokine and muscular stress, glucose deprivation, and alterations in calcium and pH are potent inducers of HSP expression in different types of cells and tissues. This review provides an overview of the cellular heat shock response to exercise. The presently known exercise related HSP are introduced. Their possible roles in response to acute exercise, thermotolerance, adaptation to training, aging, and immunological reactions are discussed. PMID- 10519063 TI - The gastrointestinal system--an essential target organ of the athlete's health and physical performance. AB - An athlete's ability to reach maximum performance is a direct result of physical and muscular performance, muscular and systemic stress tolerance, control and regulation of immune function, and adaptation to physical stress. In this complex sense, the gastrointestinal (GI) tract is also part of the system that controls and regulates adaptation and regeneration of the athlete. A well-balanced GI immune system and an optimized immune competence may protect the athlete from harmful pathogens; it may also protect against dietary as well as inhaled antigens. However, under conditions of mechanical and biochemical stress, the integrity of the GI mucosal block, particularly the epithelial hood, can be damaged, leading to a pathological uptake of toxic or immunogenic substrates. This may occur in endurance athletes, since gut symptomatology, nausea, vomiting, pain, bloating, diarrhea, cramping, and bleeding can be observed in up to half of all participants in endurance events. In addition, composition of stool and fecal microflora in endurance athletes has shown that there may be a specific need for nutritional support for mucosal immunity in highly trained but chronically stressed athletes. Proper diet during training and competition is a significant factor in guarding against GI symptoms and exercise-induced gastrointestinal side effects that may compromise immune competence and physical performance. The present review presents some important suggestions on the possible role of the GI tract in human performance and stress tolerance, and offers new insights about the influence of food quality on the immune system of the gut. PMID- 10519064 TI - Exercise and atherogenesis: where is the missing link? AB - Cardiovascular disease is the principal cause of death in Europe, the United States, and much of Asia. If sedentary people begin exercising on a regular basis, there is epidemiologal evidence of approximately 50% reduction in their risk of developing coronary heart disease. This article is an overview about epidemiology and pathogenesis of atherosclerotic lesions. It is intended to put forth the hypothesis that exercise modulates monocytes and T-lymphocytes and that this modulation is capable of guarding against atherosclerosis or inducing regression of atherosclerotic lesions. The literature does not provide sufficient data for drawing conclusions, but this article introduces a new direction of exercise immunological research. Prevention of atherosclerosis or regression of reversible forms of atherosclerotic lesions such as fatty streaks is the principal goal of preventive efforts. Sports medicine and exercise immunology may contribute significantly to the knowledge base if the cellular and molecular responses of regular exercise for atherogenesis are discovered. PMID- 10519065 TI - [Heterochromatin and single-strand DNA breaks (a hypothesis)]. AB - A hypothesis is put forward which explains the known heterochromatin properties: dense DNA packing, transcriptional inactivity, a tendency for aggregation (adhesiveness) and ectopic contacts. The basic assumption of the hypothesis is that the DNA molecules in heterochromatin are topologically open and contain single-strand breaks in the regions with the same or similar base sequence. PMID- 10519066 TI - [Comparison of the frequency of imi-r resistant Pseudomonas aeruginosa mutants generated by infection of the bacteria with various bacteriophage-transposons]. AB - We compared the frequencies of imipenem-resistant (imi-r) mutants of a Pseudomonas aeruginosa laboratory strain PAO1 infected by the phages-transposons (PT) specific for this pseudomonad species. The frequency of imi-r mutants among the lysogenic bacteria that appeared after infection reflects the frequency of integrative (conserved) PT transposition into ompD2 gene responsible for synthesis of porin, the protein required for the passage of antibiotic through the cell membrane. After infection by either PT the proportion of imi-r mutants among the lysogenic bacteria was higher than that of spontaneous mutants. The imi r mutants induced by PT infection form colonies that differ in morphology when grown on different media. The frequencies of imi-r mutants induced by all PT are similar, except for HW12, PM57, and PM62 assigned to a species of the group B3. The phages of this species induce imi-r mutants at a high frequency. Variations in frequencies and colony morphology of imi-r mutants are discussed. PMID- 10519067 TI - [Frequency of sex chromosome nondisjunction as affected by microtubule destabilization and heat shock in various lines of Drosophila melanogaster]. AB - Vinblastine (Vb), a drug belonging to a group of mitotic poisons, was shown to induce sexual chromosome nondisjunction and loss (SCNL) in oogenesis of different strains of Drosophila melanogaster. Heat shock (37 degrees C, 45 min) decreases the frequency of these events. The highest level of nondisjunction was demonstrated in the temperature-sensitive strain both after heat shock and Vb treatment. No increase of SCNL frequency in oocytes of females treated with griseofulvin (Gf) and Colchicine (Cl), analogues of Vb. PMID- 10519068 TI - [Effect of culture density and degree of expression of the trait "eyeless" and degree of polyteny of giant chromosomes in Drosophila melanogaster]. AB - Effects of culture density on expressivity of character eyeless and polyteny of giant chromosomes were studied in Drosophila melanogaster. Larval crowding was shown to significantly reduce eye facets. The expression of eyeless was sex dependent: in females, this character was expressed more strongly than in males. Larval crowding resulted in a decrease of chromosome polyteny in larval salivary glands. A strong negative correlation was established between the expressivity of eyeless and the degree of giant chromosome polyteny. PMID- 10519069 TI - [Duplications of the wing-hinge structures in the Pth mutants of drosophila melanogaster]. AB - The Pth mutation of Drosophila melanogaster, a unique dominant mutation obtained by us, causes the notum duplications that are followed by the duplications of ventral and dorsal wing-hinge structures. The duplications of the wing hinge have a strictly coordinated structure, can be ranged in a continuous series, and are divided into four distinct types, none of which overlaps with the other. The order of the emergence of the ventral wing-hinge structures was determined in duplication forms, and the shape, size, and location of these structures were compared with normal parameters. The growth of the presumptive wing-hinge region was shown to have a vectorial mode; the directions of the main vectors and their center were determined. A geometrical model is proposed, which adequately explains the strict specificity in the structure of duplications as well as the agreement between the duplication forms to one another in each of the four types. PMID- 10519070 TI - [Modification of the repair processes in chemical mutagenesis in Drosophila melanogaster]. AB - The effect of 1,4-dihydroisonicotinic acid derivative (1,4-DHINA), glutapyrone, on the frequency of sex-linked recessive lethals (SLRL), which were induced by ethyl methanesulfonate (EMS) in spermatozoa, was studied in Drosophila males and females under different exposure conditions. This test was used to analyze repair processes in EMS mutagenesis. Glutapyrone manifested the protective effect after precultivation of males at the stage of larvae and females with glutapyrone. This preparation was shown to have no effect on the frequency of EMS-induced mutations under other conditions (precultivation and subsequent cultivation of mature males with glutapyrone). These data confirmed a key role of indirect mechanisms responsible for the effect of the examined antimutagen. Glutapyrone may be assumed to improve the synthesis or functioning of enzymes involved in the repair of O6-ethylguanine. After adaptive treatment of Drosophila with low doses of the mutagen, glutapyrone increased the fertility of parents but did not suppress chemical mutagenesis. PMID- 10519071 TI - [Supernumerary chromosomes, mosaicism, and dynamics in two populations of Eastern Asian wood mouse (Apodemus peninsulae, Rodentia) from Primorskii Krai at various seasons of the year]. AB - Karyotypes of 47 individuals of the wood mice Apodemus peninsulae from two reserves in Primorskii krai (Kedrovaya Pad' and Ussuriiskii) were studied during spring, summer, and autumn. In each population, variation in the number of B chromosomes (2n = 48 + 0-5B) as well as the intratissue mosaicism determined by variation of the number, size and morphology of supernumerary chromosomes were described. Animals that have one dotlike B-chromosome were first described in both populations as rare variants. Individuals that have bone-marrow cell clones with two or three B-chromosomes were found to be predominant in the populations of Primorskii krai. The number of clones varied from one to three per animal. The frequency of mosaics showed seasonal variation. In the population of the Ussuriiskii reserve, a sharp increase in the frequency of animals with a stable karyotype was detected in autumn, at the phase of increased numbers. The variation for mosaicism was suggested to correlate with the population numbers in mice and to indicate the genetic differences between generations of the population. PMID- 10519072 TI - [Inheritance of pendulum movements in rats]. AB - Inheritance of predisposition to pendulum movements (PMs) in rats was studied by two methods: segregation analysis of binary traits (on pedigrees recorded in the selection archives for cataleptic strain GC) and the classical Mendelian analysis of hybrids between strains PM+ and PM- selected for pronounced PMs and the absence of PMs, respectively. Both methods yields the same result: it was found that predisposition to PMs exhibited a monogenic dominant inheritance with incomplete penetrance. PMID- 10519073 TI - [High allozyme variation in populations of three spiny rat species from Upper Amazonia: association with ecology and evolution]. AB - The allozyme variation in three spiny rat species of the genus Proechimys from Upper Amazonia was studied in relation to their ecology and evolution. The ecological environmental factors and biotopic distribution of species were analyzed. The unusually high allozyme variation was found in P. simonsi and P. sp. (2n = 34) inhabiting native forest biotopes. A relatively low allozyme variation in P. brevicauda was assumed to be associated with eurybiotic properties and the ability of this species to adapt to anthropogenic biotopes. Data on chromosome homeology and reconstruction of chromosome rearrangements in six spiny rat species were correlated with allozyme variation. The results suggested that chromosome rearrangements played the major role in evolution of the spiny rat species, and that the reorganization of the P. brevicauda genome was not random. P. simonsi and P. sp. (2n = 34), which live in native forest biotopes and carry an excessive genomik "informational load", were assumed to be highly susceptible to any novel external factors. These species are potentially able to produce new chromosome forms and are most significantly affected by deforestation. PMID- 10519074 TI - [Genetic demographic characteristics of the rural population of the Tuva Republic: marriage structure and inbreeding]. AB - Marriage structure was studied in three rural populations of the Tuva Republic: the Shinaan population and the populations of Todzhinskii and Bai-Taiginskii raions (districts). The Shinaan and Bai-Taiginskii populations had high levels of endogamy (0.6704 and 0.6050, respectively). In the Todzhinskii population, which was characterized by a mixed ethnic composition, endogamy was 0.3779 for the total population and 0.4626 for Tuvinians; interethnic marriages in this population were rare. The values of marriage assortativeness with respect to birthplace were 19.38, 40.75, 75.87, and 41.87% in the Shinaan, Bai-Taiginskii, all the Todzhinskii populations, and Tuvinian monoethnic marriages in the Todzhinskii raion, respectively. High marriage assortativeness with respect to ethnicity was found. Its values (A') were 91.85 and 93.49% in Tuvinians and Russians, respectively. Tuvinian populations were characterized by high inbreeding. The total (F(it)), random (Fst), and nonrandom inbreeding (Fis) estimated by isonymy were 0.004237, 0.002298, and 0.001944 in the Shinaan population, 0.007292, 0.009448, and -0.002177 in the Bai-Taiginskii population, and 0.003846, 0.004152, and -0.000307 in the Todzhinskii population, respectively (in the latter populations, the F(it), Fst, and Fis values for Tuvinian marriages alone were 0.005000, 0.007222, and -0.002238, respectively). The results obtained indicate that individual territorial groups of Tuvinians retain a high degree of genetic isolation from one another. PMID- 10519075 TI - [Genetic distances and taxonomic analysis of human populations of the Volga-Ural region based on data on DNA polymorphism]. AB - DNA polymorphism was studied in the human diallelic loci MET and D7S23 linked to the cystic fibrosis gene, diallelic locus PAH (the phenylketonuria gene), polyallelic locus ApoB, and hypervariable DNA sequences identified by means of DNA fingerprinting with phage M13 DNA as a probe. The obtained data were used to calculate genetic distances and perform taxonomic analysis of populations of the Volga-Ural region (Turkic and Finno-Ugric ethnic groups). The DNA polymorphic systems studied were demonstrated to be highly informative; their advantages and disadvantages were revealed. According to the data obtained, the genetic distances that were calculated from DNA fingerprints more adequately reflected the genetic relationships between the populations studied than the distances calculated from the allelic frequencies of four DNA loci. It was also found that, in population studies, it would suffice to analyze only the 3.5-6 kb fingerprint fragment that is most suitable for reading, rather than the entire fingerprint obtained. PMID- 10519076 TI - [Analysis of polymorphism of CTG repetitive sequences in the gene of myotonic dystrophy in human populations of the Volga-Ural region]. AB - Distribution of CTG repetitive sequences in the myotonic dystrophy (MD) gene was analyzed in ten populations of the Volga-Ural region, including Tatars, Chuvashes, Maris, Udmurts, Mordovians, Komis, and four ethnogeographical groups of Bashkirs. A total of 25 alleles were found (9 to 14 in individual populations), with each allele containing 5 to 34 trinucleotide repeats. The allele frequency distribution had two peaks corresponding to alleles with 5 and 11-14 CTG repeats. The frequency of the (CTG)5 allele varied from 0.23 to 0.47 in Maris and Mordovians, respectively. Regarding the (CTG)11-14 alleles, those containing 13 and 12 trinucleotides were most frequent in all populations; their frequencies varied from 0.15 in Mordovians to 0.24 in Maris and Bashkirs from the Abzelilovskii raion (district). Alleles with large numbers of repeats (more than 30) were only found in Tatars and Bashkirs from the Abzelilovskii raion, where their frequency was 0.01. The data obtained were compared with those on other human populations from various regions of the world. In general, the populations of the Volga-Ural region took an intermediate position between European and Asian populations (although were somewhat more similar to the latter ones) with respect to the distribution of allelic frequencies of the CTG repetitive sequences. In individual populations, the number of genotypes varied from 13 to 27 in Mordovians and Bashkirs from the Ilishevskii raion, respectively. The observed heterozygosity was the highest (91%) in Udmurts and the lowest (58%) in Mordovians; the average heterozygosity was 81%. Such a high heterozygosity, as well as the revealed differentiation of the populations with respect to the distribution of the allelic frequencies of CTG repetitive sequences in the MD gene, allow this polymorphic DNA locus to be considered a highly informative genetic marker of populations. PMID- 10519077 TI - [Polymorphism of the (CTG)n repeat in the myotonin protein kinase (DM) gene in Belarussian populations: analysis of interethnic heterogeneity]. AB - The highly polymorphic CTG triplet repeat in the 3'-nontranscribed region of the myotonin protein kinase (DM) gene was studied in healthy people from three rural populations (Grodnenskii, Khoinikskii, and Nesvizhskii raions) and the town Bobruisk, Belarus. The allele frequency distribution of this repeat proved to be similar in all the regional groups of Belarussians. PMID- 10519078 TI - [Genetic analysis of anatomical and morphologic traits of the brain, determined by magnetic resonance imaging in families of schizophrenic patients]. AB - Genetic analysis of clinical data and data obtained by magnetic resonance imaging (MRI) on 26 families of schizophrenic patients (26 probands who were patients with schizophrenia, 47 parents, and 15 siblings) revealed an enlargement of the ventricular brain system both in probands and their affected and healthy relatives. Most MRI parameters had high coefficients of inheritance and tended to be linked with positive and negative psychopathological symptoms. Our results confirm the hypothesis of genetic predisposition to the structural changes in the brains of schizophrenic patients and suggest that such MRI characteristics as the width of the anterior horn of the left lateral ventricle in the region of the caudate nucleus, the width of the central region of the left lateral ventricle, the width of the anterior horn of the right lateral ventricle in the region of the caudate nucleus, and the width of the central region of the right lateral ventricle may serve as a marker of predisposition to schizophrenia. PMID- 10519079 TI - [Cloning and characterization of the homologue of the Saccharomyces cerevisiae gene in Drosophila melanogaster]. AB - RAD23 is an evolutionary conserved protein, which is essential for DNA excision repair. It is believed that this protein is present in all eukaryotic organisms from yeast to mammals. In this work, molecular cloning of the Drosophila melanogaster RAD23 gene and an analysis of the encoded protein are reported. PMID- 10519080 TI - Dynamics of capillary electrochromatography. II. Comparison of column efficiency parameters in microscale high-performance liquid chromatography and capillary electrochromatography. AB - In capillary electrochromatography (CEC) the flow of the mobile phase is generated by electrosmotic means in high electric field. This work compares band spreading measured experimentally in several packed capillaries with electrosmotic flow (EOF) and viscous flow under otherwise identical conditions. The data were fitted to the simplified van Deemter equation for the theoretical plate height, H = A + B/u + Cu, in order to evaluate parameters A and C in each mode of flow in the different columns. The ratio of these two parameters obtained with the same column in microscale HPLC (mu-HPLC) and CEC was used to quantify the attenuation of their contribution to band spreading upon changing from viscous flow (in mu-HPLC) to electrosmotic flow (in CEC). The capillary columns used in this study were packed with stationary phases of different pore sizes as well as retentive properties and measurements were carried out under different mobile phase conditions to examine the effects of the retention factor and buffer concentration. In the CEC mode, the value of both column parameters A and C was invariably by a factor of two to four lower than in the mu-HPLC mode. This effect may be attributed to the peculiarities of the EOF flow profile in the interstitial space and to the generation of intraparticle EOF inside the porous particles of the column packing. Thus, band spreading due to flow maldistribution and mass transfer resistances is significantly lower when the mobile phase flow is driven by voltage as in CEC, rather than by pressure as in mu-HPLC. PMID- 10519081 TI - Visualization of sample introduction in liquid chromatography columns. The effect of the frit diameter. AB - A previously developed on-column detection technique using 35 mm SLR cameras [J. Chromatogr. A 826 (1998) 1] was employed to visualize colored sample bands as they elute through frits of differing diameter. Head fittings containing a 4.0 mm frit and a 15.9 mm frit were mounted in a 17 mm I.D. glass column packed with C18 silica with an average particle size of 21 microns. A carbon tetrachloride mobile phase of matching refractive index to that of the silica provides clarity along the column diameter during band migration. The photographs of the migrating sample zones were scanned and analyzed with appropriate imaging software. The smallest diameter frit induced severely parabolic sample distributions at the column inlet compared to the larger frit. Local axial dispersion coefficient values, expressed as local reduced plate height, were calculated as well as local zone velocities at the column inlet. The results demonstrate clearly the need to match the diameter of the inlet frit to the I.D. of the column so as to avoid the initial onset of zone broadening due to the frit. PMID- 10519082 TI - Enzyme amplification as detection tool in continuous-flow systems. I. Development of an enzyme-amplified biochemical detection system coupled on-line to flow injection analysis. AB - The on-line coupling of flow-injection analysis (FIA) to an enzyme-amplified biochemical detection (EA-BCD) system is described. The aim of this study is the development of a detection system able to detect biotin-containing compounds at low concentration levels. The detection system is based on the interaction of biotin with enzyme-labeled affinity proteins. Biotin possesses a high affinity to both streptavidin and anti-biotin Fab fragments, which are both tested. Several biotin derivatives are available with different reactive probes, which can be used to label analytes of interest. Therefore, biotin acts as a universal probe for the enzyme-amplified biochemical detection. Alkaline phosphatase (AP) was used as enzyme label. Several parameters, such as substrate type and concentration, concentration of enzyme-labeled affinity protein, reaction time and reaction temperature were examined. Biotin aminocaproic acid was used as a model compound. In addition to biotin aminocaproic hydrazide, other biotinylation reagents were also examined. With fluorescence detection of the enzyme-generated product, a mass detection limit of 1 fmol was achieved. PMID- 10519083 TI - Enzyme amplification as detection tool in continuous-flow systems. II. On-line coupling of liquid chromatography to enzyme-amplified biochemical detection after pre-column derivatization with biotin. AB - Enzyme-amplified biochemical detection (EA-BCD) was used as a post-column detection technique, coupled on-line with high-performance liquid chromatography (HPLC). The enzyme detection system was developed to detect biotin or biotin containing compounds. Biotinylation is widely used to label analytes of interest ranging from small molecules to proteins and DNA. Naphthalene aldehyde and anthracene aldehyde were used as model compounds. Both compounds were biotinylated off-line with biotin aminocaproic hydrazide (BACH). On-column biotinylation was performed by preconcentration of anthracene aldehyde on copper phthalocyanine. After biotinylation, samples were introduced to the HPLC system. Enzyme-labeled streptavidin, which possesses high affinity to biotin, was added post-column to the HPLC effluent. Excess of enzyme-labeled affinity protein was removed by means of an immobilized biotin column. After separation of free and bound fraction, substrate was added, which was converted to a fluorescent product by the enzyme label. Using alkaline phosphatase as an enzyme label, a mass detection limit after on-column preconcentration and biotinylation of 250 fmol was achieved. PMID- 10519084 TI - Application and comparison of derivatized cellulose and amylose chiral stationary phases for the separation of enantiomers of pharmaceutical compounds by high performance liquid chromatography. AB - The direct high-performance liquid chromatographic separation of three pairs of structurally related enantiomers on derivatized cellulose and amylose chiral stationary phases (Chiralcel OD, Chiralpak AD and Chiralpak AS) was studied using hexane as the mobile phase with 2-propanol or ethanol as modifiers. The separation, retention and elution order of the enantiomers on the different columns using different alcohol modifiers were compared. The effect of structural variation of the solutes on their k' was noted. A reversal of elution order of one enantiomeric pair upon changing the mobile-phase modifier was observed. Chiralcel OD and Chiralpak AD columns provided different elution orders of the enantiomers, including a fourth pair of enantiomers that were not structurally related to the other three pairs. PMID- 10519085 TI - Repeatability and reproducibility of retention data and band profiles on reversed phase liquid chromatography columns. III. Results obtained with Kromasil C18 columns. AB - The reproducibility of the retention data and the band profiles was investigated with Kromasil C18 columns (silica-based monomeric type reversed-phase packing material). High precision data were obtained and statistically compared among five columns from the same batch (column-to-column reproducibility) and six columns, one from each of six different batches (batch-to-batch reproducibility). These data were acquired under five different sets of chromatographic conditions, for a group of 30 neutral, acidic and basic compounds selected as probes following an experimental protocol previously described. Data characterizing the retention time, the retention factor, the separation factor, the column efficiency and the peak asymmetry for the different probe compounds are reported. Factors describing the silica surface interaction with the selected probe compounds, such as the hydrophobic interaction selectivity, the steric selectivity, and the separation factors of basic compounds at different pH values were also determined. The influence of the underlying silica on these data and correlations between the chromatographic and physico-chemical properties of the different batches are discussed. PMID- 10519086 TI - Molecular mechanism of retention in reversed-phase high-performance liquid chromatography and classification of modern stationary phases by using quantitative structure-retention relationships. AB - Quantitative structure-retention relationships (QSRRs) were derived for logarithms of retention factors normalised to a hypothetical zero percent organic modifier eluent, log kw, determined on 18 reversed-phase high-performance liquid chromatography (RP-HPLC) columns for 25 carefully designed, structurally diverse test analytes. The study was aimed at elucidating molecular mechanism of retention and at finding an objective manner of quantitative comparison of retention properties and classification of modern stationary phases for RP-HPLC. Three QSRR approaches were employed: (i) relating log kw to logarithms of octanol water partition coefficient (log P); (ii) describing log kw in terms of linear solvation-energy relationship-based parameters of Abraham; (iii) regressing log kw against simple structural descriptors acquired by calculation chemistry. All the approaches produced statistically significant and physically interpretable QSRRs. By means of QSRRs the stationary phase materials were classified according to the prevailing intermolecular interactions in the separation process. Hydrophobic properties of the columns tested were parametrized. Abilities of individual phases to provide contributions to the overall retention due to non polar London-type intermolecular interactions were quantified. Measures of hydrogen-bond donor activity and dipolarity of stationary phases are proposed along with two other phase polarity parameters. The parameters proposed quantitatively characterize the RP-HPLC stationary phases and provide a rational explanation for the differences in retention patterns of individual columns observed when applying the conventional empirical testing methods. PMID- 10519087 TI - Application of several modified peak purity assays to real complex multicomponent mixtures by high-performance liquid chromatography with diode-array detection. AB - Simple to use interactive self-modelling mixture analysis (SIMPLISMA), orthogonal projection (OPA) and Needle Search (NS) approaches have been applied to the determination of a number of compounds present in a complex multicomponent system. None of these three approaches succeeded completely when they were tested using the whole data matrix. When OPA and NS were applied to three simpler submatrices, obtained by dividing the total data matrix, and where a smaller number of compounds were present, better performance was achieved. PMID- 10519089 TI - Liquid chromatographic-mass spectrometric determination of phenolic compounds using a capillary-scale particle beam interface. AB - A capillary-scale particle beam interface was used to detect 18 phenolic compounds in red wine samples. This technique allows reproducible, library searchable electron ionization spectra at only 1 microliter/min mobile phase flow rate for a sensitive detection of the analytes in complex matrices. The method makes use of a narrow bore, reversed-phase packed capillary column for sample separation. Detection limits were in the low picogram range for most compounds. Sensitivity and response linearity were evaluated for eight phenolic acids, which are often encountered in red wines. The phenolic compound composition was outlined in two red wines obtained using different aging processes. PMID- 10519088 TI - Separation and identification of polar degradation products of benzo[a]pyrene with ozone by atmospheric pressure chemical ionization-mass spectrometry after optimized column chromatographic clean-up. AB - The environmental relevance of oxidized degradation products of polycyclic aromatic hydrocarbons (PAHs) increases due to enhanced combustion of organic matter and fossil fuels. For PAHs consisting of more than three condensed aromatic rings, soot aerosols are the main carrier, on the surface of which they can react with trace gases like ozone. In this study the clean-up procedure and analysis of ozonized benzo[a]pyrene (B[a]P) was optimized. B[a]P and its degradation products were preseparated into three fractions. Different reversed phase materials were evaluated for high-performance liquid chromatographic separation. Among these, a phenyl-modified silica material proved best-suited and the chromatographic separation was optimized on this material. For the detection of separated degradation products, liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry (LC-APCI-MS) was used. With this method, 29 components could be characterized. Besides the three known main degradation products (B[a]P-1,6-dione, B[a]P-3,6-dione, B[a]P-6,12-dione, B[a]P-4,5-dione and 4-oxa-benzo[d,e,f]chrysene-5-one (B[def]C-lactone), were identified for the first time with the help of reference substances. B[def]C-lactone is known as a substance with a mutagenic potential similar to B[a]P. Several other compounds could be tentatively identified. PMID- 10519090 TI - Liquid chromatography with electrospray ionisation mass spectrometric detection of phenolic compounds from Olea europaea. AB - The results demonstrate the potential of electrospray ionisation mass spectrometry for the specific detection of phenolic species in olives. Phenolic compounds were detected with greater sensitivity in the negative ion mode, but results from positive and negative ion modes were complementary with the positive ion mode showing structurally significant fragments. This is demonstrated by the identification of oleuropein and isomers of verbascoside. The structure of the latter were confirmed by retention, mass spectral and nuclear magnetic resonance data. These isomers have not previously been reported in olive. PMID- 10519091 TI - Isocratic separations of closely-related mono- and disaccharides by high performance anion-exchange chromatography with pulsed amperometric detection using dilute alkaline spiked with barium acetate. AB - Mixtures of closely related mono- and disaccharides may be efficiently separated by high-performance anion-exchange chromatography (HPAEC) only when relatively dilute alkaline eluents are employed (i.e., < 20 mM NaOH). The main drawbacks of these eluent solutions are (i) column regeneration between runs, (ii) poor reproducibility of the retention times, and (iii) the need for post-column base addition for enhancing sensitivity. Here, we describe some examples of isocratic separations of carbohydrates by HPAEC coupled with pulsed amperometric detection (PAD) accomplished by carbonate-free alkaline eluents (i.e., 5-20 mM NaOH) obtained upon addition of Ba(OAc)2 (1-2 mM). These separations include aldohexoses (i.e., galactose, glucose, and mannose), aminohexoses (i.e., glucosamine and galactosamine) and their N-acylated derivatives (i.e., N acetylglucosamine and N-acetylgalactosamine) along with some isomeric disaccharides (i.e., lactose, lactulose and epilactose). The separation of closely related isomers of trehalose, alpha,alpha, alpha,beta, and beta,beta, is also presented. It is recommended to add Ba(OAc)2 to NaOH solutions several hours before using the alkaline eluent (i.e., 12-24 h) to ensure complete barium carbonate precipitation in the eluent reservoir. Adopting such a simple strategy can be especially useful for performing carbohydrate separations under isocratic conditions in which no regeneration and or re-equilibration of column between runs is required. Excellent repeatability of retention data throughout a three day working session was observed, with relative standard deviations ranging from 2.0 to 3.7%, and from 0.5 to 2.0%, as day-to-day and within-day values, respectively. In addition, there was no need for postcolumn addition of strong bases to the eluent, and successful applications of the present approach confirmed its validity and practicability with detection limits of simple carbohydrates in the picomole range. PMID- 10519092 TI - Determination of gas phase peroxyacetic acid using pre-column derivatization with organic sulfide reagents and liquid chromatography. AB - The first selective HPLC methods for the determination of peroxyacetic acid (PAA) in gas phase samples have been developed. PAA reacts with 2-([3-{2-[4-amino-2 (methylsulfanyl)phenyl]- 1-diazenyl}phenyl]sulfonyl)-1-ethanol (ADS) to form the corresponding sulfoxide. Sampling may be performed in impingers using aqueous solutions of the reagent or by test tubes with the reagent coated on a solid sorbent. Sulfide and sulfoxide are separated by means of HPLC and detected at a wavelength of 410 nm. The method is highly selective for PAA in the presence of hydrogen peroxide when sampling in impingers. A 10,000-fold excess of hydrogen peroxide leads to the same peak area compared to PAA. Limit of detection is 10( 8) mol PAA, thus corresponding to PAA concentration of 46 ppb when using a sampling time of 10 min with a flow-rate at 500 ml/min. Another sulfide reagent, methyl-p-tolyl sulfide (MTS) has been used in a similar way with impinger sampling. Major advantages of ADS towards MTS are improved UV-Vis spectroscopic properties and reduced volatility. PMID- 10519093 TI - Fast, sensitive and selective liquid chromatographic-tandem mass spectrometric determination of tumor-promoting diterpene esters. AB - A liquid chromatography-tandem mass spectrometry (LC-MS-MS) method was developed to detect tumor-promoting diterpene esters of the tigliane and ingenane types within plant extracts. Fractionation on a C18 high-performance liquid chromatography (HPLC) column was followed by MS-MS-multiple reaction monitoring (MRM) using the precursor-->product ion pairs of m/z 311-->293 and 293-->265 for phorbol esters. The ion pairs m/z 313-->295 and 295-->267 were used for ingenol and deoxyphorbol esters. In a second run, the characteristic ions at m/z 311 and 313 were followed in precursor ion scan mode. These quasi-molecular ions were utilized to obtain full scan spectra of the compounds in product ion scan mode. Due to its selectivity, the present on-line method can be applied for plant cultivar selection and plant product control without time-consuming extraction procedures and complex bioassays. PMID- 10519094 TI - Determination of sulfonylurea degradation products in soil by liquid chromatography-ultraviolet detection followed by confirmatory liquid chromatography-tandem mass spectrometry. AB - A method based on liquid extraction followed by sample enrichment on reversed phase solid-phase extraction was developed for the extraction of five degradation products of four sulfonylurea herbicides (chlorsulfuron, metsulfuron-methyl, thifensulfuron-methyl and tribenuron-methyl) from soil. The compounds have been quantified by LC-UV and identified by tandem LC-MS with electrospray ionization or atmospheric pressure chemical ionization. The limits of detection for the five compounds were between 10 and 50 micrograms kg-1. The method has been applied to the extraction of soil samples after microbial degradation of sulfonylurea herbicides. PMID- 10519095 TI - Gas chromatographic determination of primary alcohols as their formylbenzoic esters by gas-phase luminescence detection. AB - A new and convenient method is described for the derivatization of primary alcohols with p-formylbenzoyl chloride, and the sensitive photometric detection of the resulting formylbenzoic ester derivatives based on their gas-phase luminescence in excited nitrogen. The coupling reaction proceeds rapidly and quantitatively, and the formylbenzoic esters show good GC properties. The minimum detectable amounts of the derivatized alcohols, at a signal three-times the peak to-peak noise, lie between 10 and 100 pg per injection, and their linear ranges cover approximately three-orders of magnitude. PMID- 10519096 TI - Evaluation of a dual-sorbent trap for monitoring organic compounds in air. AB - The performance of a new kind of multi-sorbent trap for use in the simultaneous determination of compounds of different volatility and polarity was investigated. The adsorbents employed for this purpose were Carbograph 2 and Carbograph 5. The performance of this trap was evaluated in terms of thermal desorption and solvent extraction recoveries of substances belonging to the main classes of organic compounds, at different amounts and volumes of air sampled corresponding to concentrations ranging from 0.1 to 1000 mg/m3. The tubes examined allowed the trapping of the compounds used and their complete desorption with the procedure best suited to the analytical problem. PMID- 10519097 TI - Investigation of combwax of honeybees with high-temperature gas chromatography and high-temperature gas chromatography-chemical ionization mass spectrometry. I. High-temperature gas chromatography. AB - The combwaxes of the honeybee species Apis mellifera, Apis cerana, Apis dorsata, Apis laboriosa, Apis florea and Apis andreniformis have been examined by high temperature gas chromatography. Combwax consists of a complex mixture of homologous neutral lipids. These compounds containing up to 64 carbons were chromatographed intact on a 10 m x 0.2 mm high-temperature stable SOP-50-PFD (50% diphenyl/50%-1H,1H,2H,2H-perfluorodecylmethylpolysiloxane)-co ated Duran glass capillary column. The use of this stationary phase results in lower retention values and, at last, in lower thermal stress of the analytes. In order to minimize the discrimination effect due to adsorption and/or degradation, a two step derivatization was performed resulting in the formation of tert. butyldimethylsilyl esters of the long chain fatty acids and trimethylsilyl ethers of complex hydroxyesters, respectively. The derivatization procedure was optimized using a modification of the extended Donike test. In addition this test allows the quantification of the thermal stability of the derivatives performed. The derivatization procedure was applied for combwax analysis. More than 80 compounds were separated and their peak areas semiquantitatively exploited. PMID- 10519098 TI - Preparative thin-layer chromatographic separation and subsequent gas chromatographic-mass spectrometric analysis of monoacylglycerols derived from butter oil by fungal degradation. AB - A semi-micro method has been developed using preparative thin-layer chromatography (TLC) to separate acylglycerols for the subsequent analysis by gas chromatography-mass spectrometry (GC-MS). Monoacylglycerols (MAGs) were formed from butter oil by fungal degradation with Penicillium roquefortii. Total lipids were extracted with hexane-2-propanol (3:2, v/v) and separated on silica gel preparative TLC plates with fluorescence indicator (Merck). The plates were developed in hexane-diethyl ether-formic acid (80:20:2, v/v). Lipid bands were detected under UV light or with iodine vapour, removed and then extracted with hexane-2-propanol (3:2, v/v). The MAG band (RF 0.03) was silylated into trimethylsilyl (TMS) ethers. Structures and composition of MAG-TMS ethers were analysed by GC and GC-MS. Formation of characteristic ions for the identification of sn-1(3)- and sn-2-MAG isomers was discussed. The method is simple, inexpensive and powerful for the separation and analysis of relatively small amounts of MAGs (0.2-5.0 mg) formed from fungal degradation. PMID- 10519099 TI - Comparison of continuous subcritical water extraction and hydrodistillation of marjoram essential oil. AB - Continuous subcritical water extraction (CSWE) and hydrodistillation were compared for the extraction of essential oil from marjoram leaves. Ground marjoram leaves (0.4 g) were subjected to dynamic extraction with water at 50 bar, 150 degrees C and 2 ml/min for 15 min. Hydrodistillation was performed treating 140 g of marjoram leaves with 1000 ml of water for 3 h. When CSWE was used the compounds were removed from the aqueous extract by a single extraction with 5 ml of hexane, detected by gas chromatography-flame ionization detection (GC-FID) and identified by mass spectrometry, electronic impact. The CSWE method is quicker (15 min versus 3 h), provides a more valuable essential oil (with higher amounts of oxygenated compounds and no significant presence of terpenes) and allows substantial savings of costs, in terms both of energy and plant material. The efficiency (in terms of volume of essential oil/1 g of plant) of CSWE is 5.1 times higher than that provided by hydrodistillation. The precision of the overall method (CSWE combined with GC-FID) is good (RSD less than 7.5%) for n = 5. PMID- 10519100 TI - Multiresidue determination of persistent organochlorine and organophosphorus compounds in whale tissues using automated liquid chromatographic clean up and gas chromatographic-mass spectrometric detection. AB - A multiresidue method based on normal-phase LC for the sample clean up of whale tissues extracts prior to GC-MS determination of residues of polychlorinated biphenyls, organochlorine pesticides and derivatives and lipophylic organophosphorus pesticides has been developed. Pesticides were extracted from blubber by fusing and dissolving the fat in n-hexane and from liver and kidney by reflux in n-hexane. Hexanic extracts were directly injected on the silicagel column of the automated LC clean up system, using n-hexane as mobile phase. Diode array detection allowed the on-line monitoring of lipids elution from the LC system. Purified extracts were analysed by GC using mass selective detection. The developed procedure was applied to different tissues from a whale specimen appeared in the Valencian coast, finding high concentrations of OCs (up to 7.3 micrograms g-1 pp'-DDE, and 7.2 micrograms g-1 PCBs). The method was validated by means of recovery tests for all the compounds detected in the whale and also for some other OCs and OPs studied in this paper. The method improves other current methods for the analysis of persistent organochlorines in marine mammals with regard to time of analysis, solvent expend and automation; solvent exchanges are not necessary before GC analysis, and it allows the simultaneous determination of organophosphorus pesticides. PMID- 10519101 TI - Electrospray mass spectrometry and supercritical fluid chromatography of methylated beta-cyclodextrins. AB - Five commercial dimethylated beta-cyclodextrin (DM-beta-CD) samples were analysed by electrospray (or ionspray) mass spectrometry (ESI-MS) and supercritical fluid chromatography (SFC) with evaporative light scattering detection. A silica and a nitro-bonded silica were selected using CO2-methanol-acetonitrile-water and CO2 methanol as mobile phase, respectively. An extensive optimisation scheme was performed for mobile phase selection. Both SFC systems were used for analyses of complex DM-beta-CD samples. Peak identifications were made using off-line ESI-MS. Commercial DM-beta-CDs are impure mixtures of homologues and isomers and analysis reveals that every manufacturer produces a different mixture. PMID- 10519102 TI - Separation of paralytic shellfish poisoning toxins on Chromarods-SIII by thin layer chromatography with the Iatroscan (mark 5) and flame thermionic detection. AB - Thin-layer chromatography (TLC) on Chromarods-SIII with the Iatroscan (Mark-5) and a flame thermionic detector (FTID) was used to develop a rapid method for the detection of paralytic shellfish poisoning (PSP) toxins. The effect of variation in hydrogen (H2) flow, air flow, scan time and detector current on the FTID peak response for both phosphatidylcholine (PC) and PSP were studied in order to define optimum detection conditions. A combination of hydrogen and air flow-rates of 50 ml/min and 1.5-2.0 l/min respectively, along with a scan time of 40 s/rod and detector current of 3.0 A (ampere) or above were found to yield the best results for the detection of PSP compounds. Increasing the detector current level to as high as 3.3 A gave about 130 times more FTID response than did flame ionization detection (FID), for PSP components. Quantities of standards as small as 1 ng neosaxitoxin (NEO), 5 ng saxitoxin (STX), 5 ng B1-toxins (B1), 2 ng gonyautoxin (GTX) 2/3, 6 ng GTX 1/4 and 6 ng C-toxins (C1/C2) could be detected with the FTID. The method detection limits for toxic shellfish tissues using the FTID were 0.4, 2.1, 0.8 and 2.5 micrograms per g tissue for GTX 2/3, STX, NEO and C toxins, respectively. The FTID response increased with increasing detector current and with increasing the scan time. Increasing hydrogen and air flow-rates resulted in decreasing sensitivity within defined limits. Numerous solvent systems were tested, and, solvent consisting of chloroform: methanol-water-acetic acid (30:50:8:2) could separate C toxins from GTX, which eluted ahead of NEO and STX. Accordingly, TLC/FTID with the Iatroscan (Mark-5) seems to be a promising, relatively inexpensive and rapid method of screening plant and animal tissues for PSP toxins. PMID- 10519104 TI - Rapid development of the enantiomeric separation of beta-blockers by capillary electrophoresis using an experimental design approach. AB - A rapid method for determining the separation conditions for chiral resolution of eleven beta-blocking drug substances by capillary electrophoresis is described, using an experimental design approach. An acidic phosphate-triethanolamine buffer and an uncoated fused-silica capillary were used for all experiments. Several modified cyclodextrins were applied as chiral selectors: sulfobutyl ether beta cyclodextrin (SBE-beta CD), dimethyl beta-cyclodextrin (DM-beta CD), carboxymethyl beta-cyclodextrin (CM-beta CD), and hydroxypropyl beta-cyclodextrin (HP-beta CD). Two different fractional factorial experimental designs were applied: (1) a design examining four factors at three levels (3(4-2)) and (2) one examining three factors at two levels (2(3-1)). The factors studied were: type of cyclodextrin, cyclodextrin concentration, pH of the background electrolyte and percentage of organic modifier. Enough resolution for the separation of the enantiomers and even for their quantification was reached. The same scheme is proposed when a fast chiral separation method needs to be developed for other drug families. PMID- 10519103 TI - Widening of the elution window in micellar electrokinetic chromatography with cationic surfactants. II. Cationic additives and modifiers of the electroosmotic flow. AB - In micellar electrokinetic chromatography (MEKC) with cationic surfactants the migration window is significantly narrower than with anionic surfactants. In order to overcome this disadvantage of cationic surfactants, it is investigated whether it is possible to widen the migration window by reducing the velocity of the aqueous phase while the electrophoretic mobility of the micelles is maintained. Short chain alkylammonium compounds, hexamethonium bromide and hydroxypropylmethylcellulose are tested as additives to the separation electrolyte with the potential to improve the migration window via reducing the velocity of the electroosmotic flow. It will be shown that these modifiers can be successfully used in order to widen the migration window in MEKC with cationic surfactant employing an alkyltrimethylammonium bromide as micelle forming agents. Influence of the modifiers selected on retention of neutral and acidic solutes and on efficiency of the separation system is investigated. PMID- 10519105 TI - Analysis of recombinant and modified proteins by capillary zone electrophoresis coupled with electrospray ionization tandem mass spectrometry. AB - A method for rapid characterization of recombinant and modified proteins with known sequences is described. The analytical system consists of a capillary zone electrophoresis (CZE) instrument coupled to an electrospray ionization ion trap tandem mass spectrometer via a sheath-flow interface. Following the procedure consists of proteolytic fragmentation, CZE peptide separation, tandem mass spectrometry (MS-MS) analysis of separated peptides, sequence database search and monitoring of the specific peptides, C 125 S mutated interleukin 2 (S-125-IL2) and bovine beta-casein were characterized as a model of recombinant protein and naturally modified protein, respectively. A tryptic peptide mixture derived from the synthetic salmon calcitonin (s-CT) was also analyzed to test the performance of the system. Although a conventional sheath-flow interface with much higher flow-rate compared to the microspray interface and nanospray interface was used, the proteins were identified at the low picomole level. PMID- 10519106 TI - Separation and purification of isoflavones from Pueraria lobata by high-speed counter-current chromatography. AB - High-speed counter-current chromatography (HSCCC) was applied to the semipreparative separation and purification of puerarin and related isoflavones from a crude extract of Pueraria lobata. Analytical HSCCC was used for the preliminary selection of a suitable solvent system composed of ethyl acetate-n butanol-water (2:1:3, v/v/v). Using the above solvent system the preparative HSCCC was successfully performed yielding six relatively pure isoflavones including puerarin from 80 mg of the crude extract in one-step separation. PMID- 10519107 TI - Supercritical fluid extraction of nylon 6,6 oligomers and their characterization via liquid chromatography coupled with mass spectrometry. AB - This research extends previous studies regarding the application of supercritical fluid extraction (SFE) for the analysis of oligomers from nylon 6,6 fibers. The effects of CO2 pressure, extraction temperature, CO2-modifier percentage, static extraction time and dynamic extraction time on the SFE efficiency of nylon 6,6 oligomers were examined. Results from the SFE methods for oligomer extractions were compared to results from conventional solvent extraction. The extracted oligomers were identified by high-performance liquid chromatography (HPLC) with coupled on-line atmospheric pressure chemical ionization mass spectrometry and HPLC fractionation coupled with off-line liquid secondary ion mass spectrometry. PMID- 10519108 TI - Modified apparatus for voltage gradient gel electrophoresis. AB - We built a modified version of voltage gradient gel electrophoresis system to correct distortions in nucleic acids electrophoretic migration patterns occurring at the edges of the gel when the original voltage gradient apparatus is used. The new device allows correct fractionation of nucleic acids also when electrophoresis is performed at high voltages. PMID- 10519109 TI - [Molecular bases in the search for drugs for treating AIDS. Achievements and possible prospects]. PMID- 10519110 TI - [Bacteriophage T7 RNA polymerase]. PMID- 10519111 TI - [Topoisomerases. Mechanisms of changing DNA topology]. PMID- 10519112 TI - [Telomerase]. PMID- 10519113 TI - [Polymorphism of tumor necrosis factor genes: association with diseases]. PMID- 10519114 TI - [Gene for the human tissue activator plasminogen: regulation of expression by signals, located in the 3'-untranslated region]. PMID- 10519115 TI - [Binding of nucleic acids by a protein, coding the first open reading frame of the gypsy retrotransposon (MDG4)]. PMID- 10519116 TI - [Hybridization of DNA with an oligonucleotide microchip. Correlation and screening for errors]. PMID- 10519117 TI - [Construction of a large-scale physical map of the human genome. Polymorphism of transcribed segments of ribosomal RNA genes]. PMID- 10519118 TI - [Immunochemical detection of the histone-like bacterial protein Hu in covalently bound DNA-protein complexes, obtained in vitro and in vivo]. PMID- 10519119 TI - [Cloning and preliminary characteristics of the 5'-region of the rat estrogen sulfotransferase gene]. PMID- 10519120 TI - [Serratia marcescens extracellular endonuclease. II. Amino acid residues of the active site and hypothetical mechanism of enzyme function]. PMID- 10519121 TI - [Study of bis-netropsin complexes with DNA by Raman spectroscopy]. PMID- 10519122 TI - [Cyclization of structures with shifted loops]. PMID- 10519123 TI - [Stabilizing effect of protein-protein interactions: heat denaturation of barnase and binase complexes with barstar and its C40,82A mutant]. PMID- 10519124 TI - [Conformational transitions beta-structured peptides induced by interaction with DNA]. PMID- 10519125 TI - [Use of results from comparing amino acid sequences and three-dimensional structures of aspartic proteinases in protein engineering of them]. PMID- 10519126 TI - [Crystal packing and structure of B-DNA]. PMID- 10519127 TI - [B1-dID--a new short retroposon from rodents]. PMID- 10519128 TI - [The electron microscopic analysis of synaptonemal complexes in male hybrids]. AB - Cytogenetic studies of sterile male F1 hybrids may be helpful for the understanding of genetic bases of Haldane's Rule. The main purpose of this review is to provide several explanations for various meiotic abnormalities associated with impaired fertility. Results of cytogenetic studies of gametogenesis in vertebrates (mainly mammals) performed using electron microscopy lead to the conclusion that abnormal morphology of synaptonemal complexes is one of the main factors underlying sterility of hybrid males in mammals. Various abnormalities of synaptonemal complexes have been described in male hybrids of primates (lemur), small rodents (hybrids of laboratory mice with wild mice, as well as voles, mole voles, hamsters, rats, and gerbils), and carnivores (silver fox, mustelids), as well as in the shrew, cattle hybrids, buffalo, and fish. PMID- 10519129 TI - [The notochord proteins of true sturgeons carrying the At1 epitope]. AB - Previously we reported the production of monoclonal antibodies against chordin, an acidic glycoprotein from true sturgeon notochord, carrying glycans terminating with 3-sulfoglucuronic acid. In addition, monoclonal antibodies At1, not reacting with chordin, were produced. Here we describe At1 epitope expression in sturgeon tissues and target proteins for At1 antibodies, and test interaction of these proteins with chordin and other molecules carrying glycan with 3-sulfoglucuronic acid. The expression of the antigens carrying At1 epitope during sturgeon development has also been studied. PMID- 10519130 TI - [The "direct mitogen" lead nitrate causes an intensification in lipid peroxidation and Ito cell activation in the rat liver]. AB - We have used biochemical and immunohistochemical methods to study lipid peroxidation and activation of Ito cells in rat liver after a single administration of lead nitrate, a "direct mitogen". Lead nitrate was shown to injure hepatocytes through an increased lipid peroxidation. Response to the injury included increase in the proliferative activity of parenchymal and sinusoidal liver cells. In addition, activation of Ito cells has been noted, which manifested as increased expression of desmin and increased proliferation. However, no transformation of Ito cells into myofibroblasts has been observed. We discuss the possible role of Ito cell activation in creating conditions for the proliferation of liver parenchymal cells after the injury by lead nitrate. PMID- 10519131 TI - [The development of connections of the magnocellular hypothalamic nuclei with the posterior hypophyseal lobe in rat prenatal ontogeny]. AB - The development of projections of the hypothalamic nuclei into the posterior lobe of the pituitary was studied on the fixed brain of rat fetuses from day 15 until day 19 of embryogenesis using retrograde staining with the fluorescent carbocyanine dye DiI. The formation of connections of the supraoptic and retrochiasmatic nuclei of the hypothalamus with the posterior lobe of the pituitary takes place during prenatal development on days 15 and 16-17, respectively, while only an insignificant number of the paraventricular nucleus neurons send their axons to the posterior lobe of the pituitary in rat fetuses. These facts suggest different temporal involvement of the above nuclei in formation of the hypothalamic-hypophysial neurosecretory system in rat fetuses. PMID- 10519133 TI - [Interstrain differences in the maturation of congenital reflexes in 101/HY and CBA mice in early postnatal ontogeny]. AB - We have identified interstrain differences in the rate of formation of certain reflexes and parameters of physical development in CBA/LacSto and 101/HY mice. We have shown that in young 101/HY mice, the maturation of reflexes reflecting the development of the vestibular, neurosensory, and neuromuscular systems, such as surface righting, bar holding, negative geotaxis, and auditory startle response, takes place later during postnatal life. Opening of eyes and auditory canals in mice of all studied groups takes place at the same time. In CBA females, maturation of cliff avoidance reflex occurs later than in 101/HY females; hair also appeared later. Body weight of young 101/HY mice of both sexes is significantly higher during the period of postnatal development from day 2 to day 20 than in the CBA strain. However, the relative brain weight was lower in the 101/HY strain. CBA males had a higher brain weight and also showed a faster rate of formation of inborn reflexes. We discuss possible factors underlying the observed difficulties in the rate of formation of reflexes in 101/HY and CBA young mice in relation to general background information about their genetic and neurobehavioral features. The results provide evidence that these differences are genetically determined; the rate of reflex formation does not depend on the overall physical development of mice and is rather due to a delay and/or abnormalities in nervous system development. PMID- 10519132 TI - [The embryo-protective action of antifein derivatives in chloridin-induced teratogenesis]. AB - We studied the effect of propyl- and ethylnorantifein on chloridine-induced abnormalities of extremities in rat embryos. Chloridine (50 and 25 mg/kg, given through the gastric tube) was administered to rats on day 14 of pregnancy, and its embryotoxic effect was estimated from the state of fetuses and implantation sites on day 20 of prenatal development. Propylnorantifein had fetoprotective properties both after intraperitoneal (10 mg/kg) and after intraamniotic (6 and 0.06 micrograms) administration. Ethylnorantifein under similar conditions does not change the action of chloridine, and it prevents the appearance of developmental abnormalities only at the concentration of 0.06 microgram/embryo. These data are discussed in connection with different effects of antifein derivatives on chromatin proteinkinase, which phosphorylates HMG nonhistone proteins. PMID- 10519134 TI - Characterisation of sulphonylurea and ATP-regulated K+ channels in rat pancreatic A-cells. AB - We have monitored whole-cell and single channel ATP-sensitive K+ (KATP) currents in isolated rat glucagon-secreting pancreatic A-cells. Tolbutamide produced a concentration-dependent decrease in the whole-cell KATP conductance (Ki = 6 microM) and initiated action potential firing. The K+ channel opener diazoxide, but not cromakalim or pinacidil, inhibited electrical activity and increased the whole-cell K+ conductance fourfold. ATP applied to the intracellular face of the membrane inhibited KATP channel activity with a Ki of 17 microM, an effect that could be counteracted by Mg-ADP and Mg-GDP. GTP and UTP did not affect KATP channel activity. Phosphatidylinositol 4,5-bisphosphate activated KATP channels inhibited by ATP after a delay of 90 s. In situ hybridisation demonstrated the expression of the mRNA encoding KATP channel subunits Kir6.2 and SUR1 but not Kir6.1 and SUR2. We conclude that rat pancreatic A-cells express KATP channels with the nucleotide-, sulphonylurea- and K+ channel-opener sensitivities expected for a channel formed by Kir6.2 and SUR1 subunits. PMID- 10519135 TI - Molecular and functional characterization of the small Ca(2+)-regulated K+ channel (rSK4) of colonic crypts. AB - Colonic crypt cells possess basolateral Ca(2+)-regulated K+ channels which support Cl- secretion by providing the necessary driving force. The pharmacological characteristics of these channels were examined in Ussing chamber experiments of rat and rabbit colon mucosa by the use of blockers. The chromanol 293B, a blocker of KVLQT1 channels, and clotrimazole (CTZ), a blocker of small Ca(2+)-activated K+ channels, blocked stimulated Cl- secretion completely. Small conductance Ca(2+)-activated K+ channels (SK) in excised basolateral patches of rat colonic crypts were inhibited concentration dependently by the imidazoles CTZ, NS004 and NS1619 and activated by 1-EBIO. These properties are similar to those of the known human SK channel (hSK4). hSK4-expressing Xenopus laevis oocytes showed ionomycin-activated and CTZ-inhibited K+ currents. When P2Y2 receptors were coexpressed these currents were also activated by ATP. The concentration/response curve was identical to that of rat SK channels. In human colonocytes (T84) exposed to hSK4 antisense probes, but not to sense probes, carbachol-induced K+ currents were attenuated. With RT-PCR an hSK4 could be demonstrated in human colon and in T84 colonocytes. By homology cloning the SK of the rat colon (rSK4) was identified. This protein has a high homology to hSK4 and mouse IK1. These data indicate that the Ca(2+)-activated and imidazole-inhibited basolateral K+ current in the colon is caused by SK4 channels. PMID- 10519136 TI - Endogenous polyamines modulate Ca2+ channel activity in guinea-pig intestinal smooth muscle. AB - The polyamines spermine and spermidine inhibit L-type Ca2+ channels in whole-cell recordings from guinea-pig ileum cells (Gomez and Hellstrand, Pflugers Arch, 430:501-507, 1995 [4]). To study whether they modulate channel activity under physiological conditions, we further investigated their actions on Ca2+ channels and the effects of altered cellular polyamine contents. In inside-out patches, spermine (0.1-1 mM) inhibited channel activity without affecting the amplitude of unitary currents. In cell-attached recordings, addition of spermine to the bath did not influence channel activity in the patch, indicating that its extracellular action is direct and not mediated via passage of the polyamine through the cell membrane. Cellular contents of spermidine and spermine were decreased by about 50% by organ culture of ileum strips for 5 days with the adenosylmethionine decarboxylase inhibitor CGP 48664 (10 microM). This caused enhanced channel activity in cell-attached recordings, suggesting a reduced level of channel block by endogenous polyamines compared with control cells. Whole-cell recordings in the perforated patch mode showed increased current in polyamine depleted cells, while this was not seen when cells were dialysed with the pipette solution. We conclude that polyamines block Ca2+ channels from the inside as well as the outside of the cell membrane, and that endogenous polyamines in smooth muscle modulate Ca2+ channel activity. PMID- 10519137 TI - Skeletal muscle UCP3 and UCP2 gene expression in response to inhibition of free fatty acid flux through mitochondrial beta-oxidation. AB - Studies of starvation and refeeding have implicated the genes coding for uncoupling protein-3 and -2 (UCP3, UCP2) as candidate genes in the regulation of lipids as metabolic fuels in skeletal muscle. To gain insight into the role of free fatty acid (FFA) flux in regulating the expression of these muscle UCP homologues, we recently reported that, in response to the anti-lipolytic agent nicotinic acid, utilized to reduce FFA flux at the input supply (i.e. circulating) level in fed and fasted rats, expression of the UCP3 and UCP2 genes was reduced in the soleus (predominantly slow-oxidative fibres), but not in the gastrocnemius (predominantly fast-glycolytic fibres) or tibials anterior (predominantly fast-oxidative-glycolytic fibres) muscles. In the present study, we examined UCP2 and UCP3 gene expression in these muscles from fed or fasted rats treated with etomoxir, an inhibitor of FFA flux at the output (i.e. mitochondrial oxidation) level. Fasting per se resulted in a threefold increase in serum FFA (P < 0.001) and in marked increases in the messenger ribonucleic acid (mRNA) expression of both UCP2 and UCP3 in all three muscles (P < 0.001). Treatment with etomoxir had no significant effect on serum FFA in the fed rats, but further elevated serum FFA in the fasted rats (P < 0.001). The mRNA levels of both UCP3 and UCP2 in response to etomoxir were significantly reduced in the tibialis anterior muscle in both fed and fasted states (P < 0.01), unaltered in the gastrocnemius muscle in both fed and fasted states and unaltered in the soleus muscle in the fed state, but increased in the fasted state, in parallel with the etomoxir-induced changes in serum FFA levels. Taken together, these results suggest the existence of positive feedback loops between FFA flux and muscle UCPs only in oxidative muscles--with that loop operating at the input FFA supply level for muscles with predominantly slow-oxidative fibres, and at the output FFA oxidation level for muscles with predominantly fast-oxidative glycolytic fibres. PMID- 10519138 TI - Functional and molecular evidence for Na(+)-HCO3- cotransporter in human corneal endothelial cells. AB - Although bicarbonate transport in corneal endothelium has been suggested to be coupled to Na+, the underlying molecular mechanism has not been clarified. In the present study we investigated whether a recently cloned Na(+)-HCO3- cotransporter (NBC-1) is responsible for this process, and, if so, whether the endothelium expresses a separate isoform or one of the other two isoforms that have recently been identified (kNBC-1 from kidney and pNBC-1 from pancreas). Using primers designed for specific and common regions we demonstrated by reverse transcriptase polymerase chain reaction (RT-PCR) that both kNBC-1 and pNBC-1 are expressed in cultured human corneal endothelial cells. In addition functional studies with a pH-sensitive fluorescence probe were performed. In the presence of HCO3-/CO2 a pH regulatory process was demonstrated which depends on the presence of Na+ and membrane potential, but is independent of Cl- and is inhibited by the disulfonic stilbene DIDS. These results support the presence of NBC-1 as the major bicarbonate transport system in corneal endothelium. PMID- 10519139 TI - Mechanisms of the palmitoylcarnitine-induced response in vascular endothelial cells. AB - The mechanisms of Ca2+ mobilization induced by palmitoylcarnitine (Palcar) in rabbit aortic endothelial cells (ETCs) were examined using electrophysiological techniques. The results obtained were compared with those induced by acetylcholine (ACh). When a rabbit aortic muscle preparation with an intact endothelium was treated with 10 microM Palcar, the ACh-induced relaxation was markedly attenuated, whereas endothelium-independent relaxation caused by sodium nitroprusside was not affected. Under perforated-patch whole-cell-clamp conditions, the application of Palcar over the concentration range 0.3 and 10 microM elicited a slowly activating outward current (IPalcar-out), whereas ACh induced a rapidly activating outward current (IACh). A potassium channel blocker, 4-aminopyridine, significantly inhibited both IPalcar-out and IACh. Removal of external Ca2+ almost abolished IPalcar-out. Under the same conditions, however, IACh remained transient. Addition of cation channel blockers SK&F96365 and La3+ inhibited IPalcar-out more effectively than IACh. Application of staurosporine, an inhibitor of protein kinase C, affected neither IACh nor IPalcar-out. In contrast, treatment of ETCs with pertussis toxin (PTX) reduced IACh and almost abolished IPalcar-out. These findings demonstrate that, in ETCs, Palcar induces Ca2+ influx via the activation of PTX-sensitive GTP-binding protein, leading to the activation of Ca(2+)-dependent K+ current and hyperpolarization of the cell. PMID- 10519140 TI - Ca2+ sensitivity of stunned myocardium after skinning using retrograde infusion of detergent. AB - Myofilament Ca2+ desensitization contributes to the contractile dysfunction of ischemic/reperfused ("stunned") myocardium. We examined the presence of reduced Ca2+ sensitivity of isometric force in chemically skinned fibers obtained from stunned myocardium using different modes of applying the detergent Triton X-100. Langendorff-perfused rat hearts underwent 20 min ischemia/20 min reperfusion, which caused a 35 +/- 3% decrease in left ventricular developed pressure, compared to continuously perfused control hearts. Stunned and control hearts were randomly allocated to two different permeabilization protocols: In group A, trabeculae were dissected and immersed in skinning solution containing 1% Triton X-100 for 20 min. Group B hearts remained fixed to the aortic cannula and skinning solution was infused retrogradely for 6 min prior to dissection of trabeculae. Extraction of cytosolic marker proteins was more complete in group-B than in group-A preparations. Group-A preparations from stunned hearts exhibited significant Ca2+ desensitization (pCa50 = 5.07 and 5.15 in stunned and control myocardium, respectively). In group B no such difference was observed, all preparations showing higher Ca2+ sensitivity and maximum force than group-A preparations (pCa50 = 5.32 in stunned versus 5.33 in control hearts). Prolonging group-A skinning to 150 min also abolished the difference in Ca2+ sensitivity between stunned and control myocardium. In conclusion, compared to a conventional protocol, skinning by perfusion results in more complete permeabilization and better preservation of myocardial contractile function. Ischemia/reperfusion at this moderate degree of contractile dysfunction induces Ca2+ desensitization at least partially by factors that can be extracted by thorough skinning. PMID- 10519141 TI - Selectins and beta 2-integrins mediate post-ischaemic venular adhesion of polymorphonuclear leukocytes, but not capillary plugging, in isolated hearts. AB - Leukocytes adhering to venular endothelium and emigrating into the tissue contribute to myocardial reperfusion injury. The aim of the present study was to characterize the contribution of two different families of adhesion molecules, selectins and integrins, to post-ischaemic capillary plugging and venular adhesion of leukocytes in an isolated heart model. Guinea-pig hearts were perfused using the Langendorff technique. After 20 min stabilization global ischaemia was induced for 15 min at 37 degrees C. With the onset of reperfusion 10(7) isolated polymorphonuclear leukocytes (PMN), prelabelled with rhodamine 6G, were infused within 1 min. Perfusion was continued for 2 min to wash out all cells not firmly adhering to the vascular endothelium. Hearts were then arrested, mounted on a microscope stage and perfused with a cardioplegic solution containing 0.01% fluorescein isothiocyanate (FITC)-dextran (MW 150,000). In situ videofluorescence microscopy was used to quantify PMN plugging and adherent PMN. Four groups were studied: control (no treatment or ischaemia, n = 6); ischaemia (no treatment and 15 min ischaemia, n = 5); fucoidin (pretreatment of hearts and PMN with 0.3 mg/ml selectin inhibitor fucoidin and 15 min ischaemia, n = 5) and CD18 (pretreatment of PMN with 0.1 mg monoclonal antibody against CD18 and 15 min ischaemia, n = 5). Capillary plugging by PMN was 25 +/- 5 PMN/mm2 epicardial surface area and increased moderately to 55 +/- 6 PMN/mm2 in reperfused hearts. This increase was not affected by fucoidin or CD18 antibody. In contrast, post ischaemic adhesion of PMN in small venules increased ninefold from 21 +/- 5 to 196 +/- 23 PMN/mm2 endothelial surface area. The increase in PMN adhesion to venular endothelium was blocked completely by pretreatment with fucoidin (19 +/- 5 PMN/mm-2) or CD18 antibody (7 +/- 2 PMN/mm-2). We conclude that selectin interaction alone is not sufficient to account for post-ischaemic PMN adhesion in the small venules of the coronary vasculature, because blocking the integrin subunit CD18 also inhibited PMN adhesion completely. On the other hand, neither integrins nor selectins seem to be involved in post-ischaemic capillary plugging by PMN in our perfused heart model. PMID- 10519142 TI - Activation of an apical Cl- conductance by extracellular ATP in Necturus gallbladder is mediated by cAMP and not by [Ca2+]i. AB - Necturus gallbladder epithelium (NGE) expresses a CFTR-like apical Cl- conductance that can be activated by cAMP. Here, we show that extracellular ATP (100 microM), which is known to elevate intracellular Ca2+ and to hyperpolarize cells by stimulating apical and basolateral K+ conductances, also stimulates an apical Cl- conductance (Ga,Cl), however with a much slower time course. The selectivity sequence of Ga,Cl was SCN- > I- > NO3- > Br- > Cl- >> isethionate (ISE-), but SCN- and I- partially blocked it, which is analogous to observations of CFTR Cl- channels. To disclose a possible role for intracellular Ca2+, gallbladders were incubated with the Ca2+ chelator BAPTA/AM or bathed in solutions containing only submicromolar Ca2+ concentrations. BAPTA partially inhibited the Ca(2+)-mediated hyperpolarization, but did not reduce the ATP dependent activation of Ga,Cl and the latter was also seen in low extracellular Ca2+. On the other hand, the cAMP-antagonist Rp-8-Br-cAMPS strongly inhibited the stimulation of Ga,Cl by ATP (as well as by forskolin), but left the ATP-induced hyperpolarization unchanged. Preincubation with a low concentration of forskolin markedly enhanced the stimulatory effect of ATP, and this effect was not modified by the selective inhibition of protein kinase C. These data suggest the involvement of different signal transduction pathways in the ATP-dependent activation of K+ and Cl- conductances in NGE. The stimulation of the Ga,Cl appears to be mediated by cAMP but not by elevation of intracellular Ca2+. PMID- 10519143 TI - The activation of an apical Cl- conductance by extracellular ATP is potentiated by genistein in Necturus gallbladder epithelium. AB - Necturus gallbladder epithelium (NGE) expresses a CFTR-homologous apical Cl- conductance (Ga,Cl) which can be activated either by elevation of intracellular cAMP or by extracellular ATP. Here we show by microelectrode experiments and impedance analysis that genistein (50 microM), which is known to potentiate the stimulation of Ga,Cl in several cell culture models, also potentiates the stimulation of Ga,Cl by low doses of forskolin in NGE. Moreover, we show that genistein also potentiates the stimulation of Ga,Cl by ATP. In addition genistein renders gallbladders that initially do not respond to ATP sensitive to this stimulant, and it delays the conductance inactivation after ATP removal. Under control conditions Ga,Cl inactivates within < 5 min, but in the presence of genistein a significant Ga,Cl persists even after 60 min. These effects of genistein are not related to inhibition of protein tyrosine kinases, since structurally different inhibitors of the tyrphostin family do not mimic the genistein effects. The data support our conclusion that stimulation of Ga,Cl by ATP is mediated by activation of the cAMP pathway and involves a CFTR-homologous protein. They also favour the view that genistein acts via inhibition of protein phosphatases which dephosphorylate CFTR, but cannot exclude the possibility of a direct interaction with CFTR. PMID- 10519144 TI - The development of post-transplantation hypertension in recipients of an SHR kidney is independent of reinnervation of the graft. AB - The role of sympathetic reinnervation of kidney grafts for the development of renal post-transplantation hypertension was investigated. F1-hybrids (F1H) obtained from crossing spontaneously hypertensive (SHR) and Wistar-Kyoto rats were transplanted with an SHR kidney and bilaterally nephrectomized. Seven (n = 7) and 42 days after transplantation (n = 9), transplanted kidneys were removed and renal norepinephrine concentrations were determined. In addition, mean arterial pressure (MAP) was measured in conscious bilaterally nephrectomized recipients of an SHR (n = 9) or F1H kidney (n = 8) 6 weeks after transplantation. Renal norepinephrine concentrations (ng/g wet kidney weight) decreased dramatically from 348.3 +/- 31.7 ng/g before (n = 7) to 9.9 +/- 2.5 ng/g at 7 days and 6.5 +/- 1.1 ng/g at 42 days after transplantation, indicating that there was no substantial sympathetic reinnervation of the grafts throughout the observation period. Despite the lack of reinnervation of the grafts, recipients of an SHR kidney but not recipients of an F1H kidney developed post transplantation hypertension (MAP 172 +/- 4 mmHg versus 124 +/- 3 mmHg P < 0.001) within 6 weeks after transplantation. We conclude that post-transplantation hypertension in recipients of an SHR kidney does not depend on sympathetic reinnervation of the graft. PMID- 10519145 TI - Permeation of NH3/NH4+ and cell pH in colonic crypts of the rat. AB - Colon cells are subjected to high concentrations of NH3 and NH4+, and a sizeable portion of this buffer is absorbed. The flux of these components into cells causes opposite effects on their pH; this effect is largely used to induce an acid load and to observe the mechanism of acid extrusion from cells. We studied cells of microdissected colon crypts loaded with BCECF and superfused with NH4Cl containing Krebs-Ringer solution. We found a marked transient reduction in pH measured by ratiometric fluorescence microscopy, from a control value of 7.51 +/- 0.041 to 7.15 +/- 0.041 (n = 21), instead of the initial alkalinization found in most cells. This pH was reached at a rate of 0.95 +/- 0.07 pH units/min. Addition of 1 mmol/l furosemide, a blocker of Na+,K+,2Cl- cotransport, to the ammonium solution inverted this acidification toward alkalinization (pH 7.89 +/- 0.041, n = 5), and superfusion with furosemide plus 0.1 mmol/l hexamethylene amiloride, a specific blocker of Na+/H+ exchange, increased this initial alkalinization further to 8.10 +/- 0.117 (n = 7). When Krebs-Ringer with 0 Cl- containing (NH4)2SO4 instead of NH4Cl was superfused, the acid transient was also reverted to alkalinization; however, a higher degree of alkalinization was observed either when 1 mmol/l furosemide was added to the superfusing sulfate solution (when a pH of 7.78 +/- 0.010 was reached), or when ammonium gluconate was used instead of ammonium sulfate. The addition of Ba2+ to the superfusion solution did not alter the initial acidification. These data indicate that in colon crypt cells, basolateral membrane transporters, in particular the Na+,K+,2Cl- cotransporter and the Na+/H+ exchanger (but not Ba(2+)-sensitive K+ channels), mediate the predominant influx of NH4+ ions leading to the initial transient acidification. PMID- 10519146 TI - Maturation and myotonia influence the abundance of cation channels KDR, KIR and CIR differently: a patch-clamp study on mouse interosseus muscle fibres. AB - To detect cation channels, the expression of which is dependent on the physiological state of muscle, single-channel activities of dissociated fibres of the mouse interosseus muscle were recorded using the patch-clamp technique in the cell-attached mode. Fibres were prepared from juvenile and adult wild-type (WT), from chloride channel-deficient myotonic and from denervated adult WT muscles. In all cases delayed-rectifier K+ channels (KDR) with a unitary conductance of 11 pS were recorded in more than 95% of sarcolemmal patches, but with a low, steady state open probability. Inwards-rectifying K+ channels (KIR) with a conductance of 31 pS in 140 mM [K+]o were active in about 50% of the membrane patches from WT and in more than 90% of those from myotonic fibres. A hitherto undescribed, inwards-rectifying, cation channel, provisionally termed CIR, with fast kinetics and a unitary conductance of 36 pS, was active in nearly every membrane patch from juvenile mice, both WT and myotonic. The abundance of CIR decreased during development, but was not changed 7 days after denervation of adult WT muscle. Ca(2+)-dependent K+ channels were seen sporadically. Channels with the characteristics of adenosine 5'-triphosphate (ATP)-sensitive K+ channels were recorded frequently upon excision of membrane patches, but remained inactive in most cell-attached recordings. In conclusion, of the investigated ion channels, only KIR was responsive to the activity pattern of adult muscle, whereas CIR was down-regulated during muscle maturation. PMID- 10519147 TI - Force responses of skinned molluscan catch muscle following photoliberation of ATP. AB - Isometric force responses following flash photolysis of caged-ATP were measured from skinned preparations of the catch muscle anterior byssus retractor of Mytilus (ABRM). When fibres were transferred from Ca(2+)-free to Ca(2+) containing rigor solution (pCa < 4) the force remained low, but flash photolysis produced an extended force increase (half-time, 0.30 +/- 0.07 s, n = 6). When Ca(2+)-activated fibres were transferred to a Ca(2+)-free rigor solution, their force remained at a high level. Flash photolysis produced a rapid force decay (half-time, 0.28 +/- 0.06 s, n = 9) to about 19% of the initial Ca(2+)-activated force. In the presence of 0.5 mM MgADP, the force increase was slowed down by a factor of 3 and the force decay by a factor of 5. These effects of MgADP on crossbridge kinetics are comparable to those observed in vertebrate smooth muscle and are thought to cause "latch", a catch-like state (Fuglsang et al. J Muscle Res Cell Motil 14:666-677, 1993). They are consistent with a model implicating competition between MgADP and MgATP for the nucleotide-binding site on crossbridges. Considering the relatively fast force responses induced by caged ATP photolysis, even in the presence of MgADP, it appears unlikely that the detachment of crossbridges from the rigor state can account for catch-related processes. In view of the low myosin ATPase under maximal activating conditions (0.6 s-1, Butler et al. Biophys J 75:1904-1914, 1998), neither crossbridge attachment nor detachment of rigor crossbridges seems to be the rate-limiting processes of the crossbridge cycle. PMID- 10519148 TI - The actions of altered osmolarity on guinea-pig detrusor smooth muscle contractility and intracellular calcium. AB - The study measured the effects in vitro of changing extracellular osmolarity on the contractility of detrusor smooth muscle strips. The data were interpreted in the context of separate measurements from isolated cells of alterations to the intracellular [Ca2+], [Ca2+]i. Increased osmolarity (300-700 mosmol l-1) reduced phasic contractions but increased resting tension regardless of whether sucrose, LiCl or NaCl were used as osmolytes. [Ca2+]i was decreased slightly only when NaCl increased osmolarity, otherwise it was unchanged. The contractile effects may be explained by tissue shrinkage and reduction of detrusor excitability. Lowered osmolarity (300-64 mosmol l-1) decreased phasic contractions but increased resting tension and [Ca2+]i. The raised resting tension was due solely to low osmolarity and was independent of changes to [Na], [Cl] or ionic strength. The rise of [Ca2+]i was due partly to Ca2+ influx through Na(+)-Ca2+ exchange but a fraction was independent of extracellular Ca, unaffected by Gd3+, and persisted in the presence of caffeine. By contrast, reduction of phasic tension was due mainly to the reduced ionic strength, not osmolarity. The results do not support the presence of functional stretch-activated channels and suggest only a minor role for Na(+)-Ca2+ exchange under these conditions. However, they do suggest an intracellular source of Ca2+, which is independent of the sarcoplasmic reticulum. PMID- 10519149 TI - Tyrosine-kinase-dependent regulation of the nitric oxide synthase gene by endothelin-1 in human endothelial cells. AB - In vascular endothelium, endothelium-derived relaxing factor, predominantly nitric oxide (NO), is synthesized by endothelial NO synthase (eNOS). While regulatory influences on eNOS enzyme activity are widely clarified, little is known about the regulation of the eNOS gene. We investigated the regulatory signaling mechanisms of eNOS mRNA expression and accumulated NO production in human endothelial cells. Northern blot analysis and NO assays demonstrate that the vasoconstrictor peptide endothelin-1 (ET-1) induces the eNOS gene and leads to accumulated NO production. Induction occurs via ETA receptor activation and depends on improved transcript stability. It is maintained for incubation periods of 30-90 min and tapers thereafter. Regulatory signaling mechanisms depend on de novo protein synthesis to control eNOS mRNA fate. Selectively blocking protein tyrosine kinases (PTK) and inhibiting protein kinase C (PKC) inhibit eNOS mRNA expression and accumulated NO secretion. These observations indicate that regulation of eNOS at the genomic level occurs via post-transcriptional mechanisms. Two protein-bound intracellular kinase pathways, PTK and PKC, regulate eNOS mRNA expression and accumulated NO production. PMID- 10519150 TI - Quantification of calcium release from the sarcoplasmic reticulum in rainbow trout atrial myocytes. AB - Using the whole-cell configuration of the patch-clamp technique, we quantified calcium release from the sarcoplasmic reticulum (SR) elicited by short depolarization pulses before and after clearance of the SR Ca2+ content with 10 mM caffeine (CAF). With a loaded SR, the first detectable contraction occurred with a pulse duration of 5 ms. CAF exposure increased this pulse duration to 85 ms and slowed the inactivation of the Ca2+ current (ICa). Repolarization of the cell to -80 mV after a short depolarization elicited a tail current that was attenuated markedly after CAF exposure. The difference between the charge carried by the tail currents obtained before and after CAF exposure was taken as a measure of the Ca2+ released from the SR. SR Ca2+ release increased with increasing SR Ca2+ load over the range of loads examined. In contrast, SR Ca2+ release reached a maximum when the duration of the preceding depolarization exceeded 10 ms. Maximal Ca2+ release was 1.64 amol/pF or 62 microM and elicited a contraction that was 40 +/- 6% of the amplitude of a normal contraction. This release could account for 48 +/- 10% of the total Ca2+ required to activate contraction but only a few percent of the CAF-releasable Ca2+. Thus, contrary to the general view of excitation-contraction coupling in lower vertebrates, SR Ca2+ release in trout atrial myocytes may account for up to 50% of the total Ca2+ transient at room temperature. PMID- 10519151 TI - Osteoblasts derived from load-bearing bones of the rat express both L- and T-like voltage-operated calcium channels and mRNA for alpha 1C, alpha 1D and alpha 1G subunits. AB - Voltage operated calcium channels (VOCCs) are implicated in osteoblastic mechano- and hormonal transduction. Very little, however, is known about the expression of VOCCs in osteoblasts of load-bearing bones. Here we describe two types of whole cell calcium current in rat femoral explant-derived osteoblasts. The first is high-voltage activated and sensitive to nifedipine, Bay K8644 and FPL 64176. The second is low-voltage activated and is sensitive to micromolar concentrations of Ni2+. The properties of these two currents are consistent with those of L-type and T-type calcium currents respectively. T-type currents were detected in most cells on the day of passage, the level of expression being significantly lower on subsequent days. L-type currents were also most common on the day of passage but were detected consistently throughout the 4-day period of study. The reverse transcription polymerase chain reaction with non-specific primers directed against all L-type VOCC alpha 1 subunits and then with specific primers directed against sequences from rat brain alpha 1C (L-type), alpha 1D (L-type) and alpha 1G (T-type) VOCC subunits detected transcripts of appropriate size in all four cases. Products from the three sets of specific primer pairs (alpha 1C, alpha 1D, alpha 1G) were sequenced and were identical to their respective rat brain templates. PMID- 10519152 TI - Capacitance measurements reveal different pathways for the activation of CFTR. AB - We used the Xenopus laevis oocyte expression system to characterize adenosine 3',5'-cyclic monophosphate (cAMP) activation of the cystic fibrosis transmembrane conductance regulator (CFTR). With conventional two-microelectrode voltage-clamp techniques, we recorded transmembrane conductance (Gm) and membrane current (Im). Using five different sine wave frequencies, we also monitored changes of the plasma membrane surface area by recording continuously membrane capacitance (Cm) under voltage-clamp conditions. Impedance spectra recorded in the frequency range 0.1-500 Hz showed that, at least up to 200 Hz, Cm is independent of the frequency. In control oocytes, cAMP (100 microM) treatment did not affect Gm or Im but evoked a small, slowly occurring increase in Cm, probably mediated by cAMP stimulated exocytosis. However, in oocytes expressing CFTR, large simultaneous increases of Gm, Im and Cm occurred after stimulation with cAMP. Oocytes injected with the delta F508 CFTR mutant behaved like control oocytes and cAMP had no additional effects on Gm, Im or Cm. In oocytes injected with wild-type CFTR, adenosine 5'-triphosphate (ATP, 100 microM) did not activate the cAMP-induced augmentation of Im, Gm or Cm further. On the other hand, cAMP-induced increases in Cm were reduced significantly by the specific blockers of protein kinase A (PKA) KT5720 and N-[2-(methylamino-9-ethyl]-5-isoquinolinesulphonamide hydrochloride (H8), whereas the increases in Gm and Im were essentially unaffected by these agents. Reducing intracellular Ca2+ by injection of a Ca2+ chelator 1,2-bis (2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA) prevented PKA-dependent exocytosis while activation of Im and Gm of already inserted CFTR still could be detected. The specific cAMP antagonist adenosine 3',5'-cyclic monophosphothioate Rp diastereomer (RpcAMPS) completely suppressed the effects of cAMP on all parameters. These findings are consistent with the concept of different pathways of CFTR activation by cAMP: already-inserted CFTR Cl- channels are activated directly by cAMP, while traffic of CFTR proteins from an intracellular pool to the plasma membrane and functional insertion into the plasma membrane occurs via cAMP- and Ca(2+)-dependent PKA-mediated exocytosis. PMID- 10519153 TI - Mutation in the leptin receptor (Leprfa) causes fat-storage-independent hyperleptinaemia in neonatal rats. AB - The plasma leptin concentration adjusted for fat mass is affected by mutant gene dosage in older animals segregating for Leprfa. Because the plasma of neonatal rats contains leptin, although their adipocytes contain virtually no triglyceride, we determined whether mutation dose-dependent differences in plasma leptin concentration exist before the postnatal onset of triglyceride storage. Plasma samples were obtained 10 min after birth of each rat pup and leptin concentration determined by radioimmunoassay. Plasma leptin in homozygous wild type (+/+) pups was 1.6 +/- 0.2 ng/ml (n = 20) and 2.4 +/- 0.2 ng/ml in +/fa (n = 32) littermates (least-square means +/- SE, P < 0.05, two-way ANOVA with litter and genotype as factors). The corresponding values for +/fa (n = 21) and fa/fa (n = 15) littermates were 2.4 +/- 0.2 and, 4.0 +/- 0.3 ng/ml respectively (P < 0.001). Leprfa gene dose-dependent elevations in plasma leptin are, therefore, present at birth and constitute the only Leprfa-related phenotypic trait presently known to precede the onset of increased fat storage. PMID- 10519155 TI - Thematic review series. VIII: Phagocyte antimicrobial systems. Introduction. PMID- 10519154 TI - Identification of system y+L as the high-affinity transporter for L-arginine in human platelets: up-regulation of L-arginine influx in uraemia. AB - Kinetic studies of L-arginine transport in human platelets have identified a high affinity, low-capacity transport system [Michaelis-Menten constant (K(m)) about 10 microM] for cationic amino acids that also transports neutral amino acids with high affinity in the presence of Na+ but not K+. These characteristics, together with our kinetic cis-inhibition studies, indicate that saturable L-arginine transport in human platelets is mediated via the system y+L and not the classic cationic transporter system y+. We present here the first evidence that L arginine transport via system y+L is increased twofold in platelets from patients with chronic renal failure. System y+L has been described in human erythrocytes, peripheral blood mononuclear cells and placenta, and up-regulation of system y+L activity in human platelets could explain the paradox of increased nitric oxide (NO) production by uraemic platelets under conditions of decreased plasma L arginine and elevated NG-monomethyl-L-arginine (L-NMMA) concentrations. PMID- 10519157 TI - Myeloperoxidase. AB - Phagocytes respond to stimulation with a burst of oxygen consumption, and much, if not all, of the extra oxygen consumed in the respiratory burst is converted first to the superoxide anion and then to hydrogen peroxide (H2O2). Myeloperoxidase (MPO), which is released from cytoplasmic granules of neutrophils and monocytes by a degranulation process, reacts with the H2O2 formed by the respiratory burst to form a complex that can oxidize a large variety of substances. Among the latter is chloride, which is oxidized initially to hypochlorous acid, with the subsequent formation of chlorine and chloramines. These products of the MPO-H2O2-chloride system are powerful oxidants that can have profound biological effects. The primary function of neutrophils is the phagocytosis and destruction of microorganisms, and the release of MPO and H2O2 into the phagosome containing the ingested microorganism generally leads to a rapid microbicidal effect. Neutrophils from patients with chronic granulomatous disease (CGD) have a microbicidal defect that is associated with the absence of a respiratory burst and, thus, H2O2 production. Neutrophils from patients with a hereditary MPO deficiency, who lack MPO, also have a microbicidal defect, although it is not as severe as that seen in CGD. MPO and H2O2 also can be released to the outside of the cell where a reaction with chloride can induce damage to adjacent tissue and, thus, contribute to the pathogenesis of disease. It has been suggested that pulmonary injury, renal glomerular damage, and the initiation of atherosclerotic lesions may be caused by the MPO system. PMID- 10519156 TI - The NADPH-dependent oxidase of phagocytes. AB - Polymorphonuclear leukocytes (PMNs) represent a prominent cellular element in the innate immune system, serving to ingest exogenous particles and microbes and to kill phagocytosed microorganisms. The microbicidal activity of PMNs depends on the interactions of a broad array of potent systems, including relatively stable degradative proteins as well as labile reactive radicals. These systems can be categorized as oxygen-dependent and nonoxidative mechanisms, although the physiologically relative activity depends on the precisely orchestrated interplay between both systems. The enzyme complex responsible for the activity of the oxygen-dependent system is the respiratory burst oxidase and its important contribution to host defense is best illustrated by the frequent and severe infections seen in individuals whose PMNs lack oxidase activity, namely patients with chronic granulomatous disease (CGD). Multiple elements comprise the oxygen dependent system, and significant advances have been made in the past decade in understanding the protein components of the respiratory burst oxidase, their subcellular distribution in resting PMNs, and their agonist-dependent assembly into a functional system at phagosomal and plasma membranes. In parallel, substantial insights into the molecular bases of CGD have likewise been made. Nonetheless there remain significant gaps in our understanding of the precise functional contributions of particular components of the system, the molecular mechanisms that regulate their coordinated assembly, and the role of related proteins in nonphagocytic cells. PMID- 10519159 TI - Thematic review series. IX: New therapeutic targets for the treatment of heart failure. Introduction. PMID- 10519158 TI - Oxygen-independent microbicidal mechanisms of phagocytes. AB - The principal biological function of phagocytic cells is the destruction of invading microorganisms. Following phagocytosis, microbes are exposed to multiple antimicrobial substances ranging in complexity from simple oxygen radicals to large proteins. These substances disrupt various microbial structures and eventually kill and digest most of the invaders. This review is focused on oxygen independent microbicidal mechanisms in granulocytes and macrophages. PMID- 10519160 TI - Myocardial G protein-coupled receptor kinases: implications for heart failure therapy. AB - The beta-adrenergic signaling cascade is an important regulator of myocardial function. Significant alterations of this pathway are associated with several cardiovascular diseases, including congestive heart failure (CHF). Included in these alterations is increased activity and expression of G protein-coupled receptor kinases (GRKs), such as the beta-adrenergic receptor kinase (beta ARK1), which phosphorylate and desensitize beta-adrenergic receptors (beta ARs). A body of evidence is accumulating that suggests that GRKs, in particular beta ARK1, are critical determinants of cardiac function under normal conditions and in disease states. Transgenic mice with myocardial-targeted alterations of GRK activity have shown profound changes in the in vivo functional performance of the heart. Included in these studies is the compelling finding that inhibition of beta ARK1 activity or expression significantly enhances cardiac function and potentiates beta AR signaling in failing cardiomyocytes. This article summarizes the advances made in the study of beta ARK1 in the heart and addresses its potential as a novel therapeutic target for CHF. PMID- 10519161 TI - The natriuretic peptides in heart failure: diagnostic and therapeutic potentials. AB - The natriuretic peptides are a group of structurally similar but genetically distinct peptides that have diverse actions in cardiovascular, renal, and endocrine homeostasis. Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) are of myocardial cell origin and C-type natriuretic peptide (CNP) is of endothelial origin. ANP and BNP bind to the natriuretic peptide-A receptor (NPR-A), which, via 3',5'-cyclic guanosine monophosphate (cGMP), mediates natriuresis, vasodilation, renin inhibition, antimitogenesis, and lusitropic properties. CNP lacks natriuretic actions but possesses vasodilating and growth inhibiting actions via the guanylyl cyclase-linked natriuretic peptide-B receptor (NPR-B). All three peptides are cleared by the natriuretic peptide-C receptor (NPR-C) and are degraded by the ectoenzyme neutral endopeptidase 24.11 (NEP), both of which are widely expressed in the kidneys, lungs, and the vascular wall. Congestive heart failure (CHF) represents a pathological state in which the activation of the natriuretic peptides exceeds those of all other states. In this brief review, we will attempt to provide an update on important issues regarding natriuretic peptides in CHF, with a focus on their functional importance as a beneficial humoral response in asymptomatic left ventricular dysfunction (LVD), the mechanisms of natriuretic peptide hyporesponsiveness in severe heart failure, the diagnostic and prognostic significance of the natriuretic peptides in CHF, and the therapeutic potential of the natriuretic peptides in this multiorgan syndrome. PMID- 10519162 TI - Role of endothelin-1 in myocardial failure. AB - Endothelin-1 (ET-1) is a potent molecule produced throughout the cardiovascular system; it can exert important effects on both the structure and function of vascular smooth muscle cells and cardiac myocytes. ET-1 appears to play a central role in the physiological regulation of cardiovascular function, particularly in the vasculature. The known actions of ET-1 and the demonstration that plasma ET-1 is elevated in patients with heart failure has raised the possibility that this molecule could play a role in the pathophysiology of heart failure. This thesis has been supported and furthered by studies in animal models of heart failure that demonstrate the salutary, short-term effects of ET-1 receptor antagonists on hemodynamic function, as well as improved ventricular remodeling and survival with long-term administration. Early clinical trials with these ET receptor blockers have demonstrated systemic vasodilation. Long-term trials to determine the effects of ET-1 blockade on symptoms and survival are under way. PMID- 10519163 TI - Cytokines in heart failure: pathogenetic mechanisms and potential treatment. AB - Recent studies have shown that patients with heart failure overexpress a class of biologically active molecules, generically referred to as pro-inflammatory cytokines. This article will review recent clinical and experimental material that suggests that pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), and interleukin-6 (IL-6) may play a role in the pathogenesis of congestive heart failure. In addition, we will review recent studies that suggest that antagonizing cytokines may represent a novel target for heart failure therapy. PMID- 10519164 TI - Voltage-gated T-type Ca2+ channels and heart failure. AB - In the cardiovascular system, two types of voltage-gated Ca2+ channels are present: the L-type and the T-type. Under normal conditions, T-type Ca2+ channels are involved in the maintenance of vascular tone and cardiac automaticity but, since they are not present in contractile myocardial cells, they do not contribute significantly to myocardial contraction. In experimental models of cardiac hypertrophy, myocardial T-type Ca2+ channels are upregulated, which could contribute to the increased incidence of ventricular arrhythmia. In addition, T type Ca2+ channels participate in the regulation of cell proliferation and neurohormonal secretion; through these pathways, T-type Ca2+ channels might participate in myocardial remodeling. The pathophysiological role of T-type Ca2+ channels in heart failure has been investigated using mibefradil, a Ca2+ antagonist that is 10-50 times more potent at blocking T-type than L-type Ca2+ channels. In contrast with classic L-type Ca2+ channel antagonists, miberfradil appears beneficial in many animal models of heart failure; in particular, it does not exert negative inotropic effects nor does it stimulate the neurohormonal system. Furthermore, in the Pfeffer rat model, blockade of T-type Ca2+ channels with mibefradil is associated with an improved survival rate. In humans, however, major metabolic drug interactions independent of T-type Ca2+ channel blockade made it impossible to determine the efficacy of mibefradil in treating heart failure; indeed, these interactions led to the withdrawal of the drug from the market. PMID- 10519165 TI - Heme oxygenase: recent advances in understanding its regulation and role. AB - Heme oxygenase (HO) is responsible for the physiological breakdown of heme into equimolar amounts of biliverdin, carbon monoxide, and iron. Three isoforms (HO-1, HO-2, and HO-3) have been identified. HO-1 is ubiquitous and its mRNA and activity can be increased several-fold by heme, other metalloporphyrins, transition metals, and stimuli that induce cellular stress. HO-1 is recognized as a major heat shock/stress response protein. Recent work from our laboratory has demonstrated several potential consensus regulatory elements in the 5' untranslated region (UTR) of HO-1, including activator protein 1 (AP-1), metal responsive element (MRE), oncogene c-myc/max heterodimer binding site (Myc/Max), antioxidant response element (ARE), and GC box binding (Sp1) sites. Using deletion-reporter gene constructs, we have mapped sites that mediate the arsenite dependent induction of HO-1, and we have shown that components of the extracellular signal-regulated kinase (ERK) and p38 (a homologue of the yeast HOG1 kinase), but not c-jun N-terminal kinase (JNK), mitogen-activated protein (MAP) kinase pathways are involved in arsenite-dependent upregulation. In contrast, HO-2 is present chiefly in the brain and testes and is virtually uninducible. HO-3 has very low activity; its physiological function probably involves heme binding. Products of the HO reaction have important effects: carbon monoxide is a potent vasodilator, which is thought to play a key role in the modulation of vascular tone, especially in the liver under physiological conditions, and in many organs under "stressful" conditions associated with HO-1 induction. Biliverdin and its product bilirubin, formed in most mammals, are potent antioxidants. In contrast, "free" iron increases oxidative stress and regulates the expression of many mRNAs (e.g., DCT-1, ferritin, and transferrin receptor) by affecting the conformation of iron regulatory protein (IRP)-1 and its binding to iron regulatory elements (IREs) in the 5'- or 3'-UTRs of the mRNAs. PMID- 10519166 TI - Effects of cutaneous aspirin on the human stomach and duodenum. AB - Oral aspirin blocks cyclooxygenase in platelets, lowering serum thromboxane concentrations. Oral aspirin also blocks cyclooxygenase in the gastrointestinal mucosa, lowering prostaglandin production and increasing the risk of gastrointestinal ulceration and bleeding. Aspirin placed on the skin also inhibits cyclooxygenase in platelets, but aspirin absorption through skin is slow, which may minimize the gastrointestinal effects. Our objectives in this study were 1) to compare the pharmacokinetic and pharmacodynamic effects of cutaneous and oral aspirin in healthy volunteers and 2) to compare the effects of cutaneous aspirin on gastroduodenal mucosal prostaglandin E2 and F2 alpha content and on mucosal damage, using endoscopy. The bioavailability of cutaneous aspirin was 4%-8% that of oral aspirin. Cutaneous aspirin (750 mg/day for 10 days) significantly lowered serum thromboxane (by 85%) and gastric and duodenal prostaglandins (by 49%-71%); placebo had no effect. Moreover, cutaneous aspirin, but not placebo, resulted in significant gastric mucosal injury. These findings demonstrate that even tiny amounts of aspirin in the blood (2 microM) have inhibitory effects on prostaglandin production in the human stomach and duodenum that result in gastric mucosal damage, even without direct exposure of the stomach to aspirin. PMID- 10519167 TI - Why the physician-scientist? Why the Association of American Physicians? PMID- 10519168 TI - Presentation of the Kober Medal for 1999 to Jean D. Wilson physician-scientist exemplar. PMID- 10519169 TI - Acceptance of the Kober Medal for 1999. PMID- 10519170 TI - [The new variant of Creutzfeldt-Jakob disease: several questions without response]. PMID- 10519171 TI - [Evaluation of biological impregnation of a population exposed to high concentration of arsenic in water supply, Ferrette, 1997]. AB - BACKGROUND: The population of Ferrette has been exposed to well-water with arsenic (As) levels higher than legal threshold. The aim of this study was to assess the relationships between daily tap-water consumption, As quantities thus ingested and biological arsenical impregnation. METHODS: The study was carried out on a sample of 100 people in the town of Ferrette and 100 people in the town of Seppois-le-Bas where the water quality is satisfactory. Ingested water and As were assessed by the mean of a food questionnaire. The quantity of ingested As was related to the body weight and compared to the tolerable daily intake (TDI) of 2 micrograms/kg/d. Biological impregnation was assessed by measuring out As in hair sample. RESULTS: The daily ingested As intake of Ferrette population ranged from 0 to 32 micrograms/kg/d. One half of the population ingested more than 2 micrograms/kg/d. A quarter of the population ingested more than 4.3 micrograms/kg/d. 15% of Ferrette inhabitants yielded an As hair level higher than 0.1 ng/mg [IC95%: 8.7%-23.5%], versus 7% [IC95%: 2.9-13.9%] for the inhabitants of Seppois-le-Bas (p = 0.07). Among those who ingested an amount of As higher than the TDI, 19% were found to have detectable As hair levels, versus 9% for those who ingested less than the daily acceptable amount (p = 0.18). CONCLUSION: One half of the population of Ferrette absorbed an As amount double to the TDI, evidencing the reality of the exposure. We did not find any statistically significant relation between the ingested As amount and biological As impregnation nor between exposure to water containing excessive As level and As biological impregnation. PMID- 10519172 TI - [Capability of health services of the city of Ouagadougou to diagnose sexually transmitted diseases]. AB - BACKGROUND: Quantitative and qualitative shortage of both human and material resources in the diagnosis of sexually transmitted diseases (STD's) remains one of the difficulties in dealing with these diseases in developing countries. The aim of this study was to determine the capability of health and medical laboratories personnel to diagnose STD's prevailing in Ouagadougou. METHOD: The study has been conducted in all the health centres and the medical laboratories of the town. The personnel ensuring out-patient clinics and the director of each medical laboratory have been interviewed. The interview as well as the assessment of the consulting room and the medical laboratories technical equipment were carried out. RESULTS: The range of STD's that the staff was able to point out was limited only to classical diseases. One member of the staff out of five mentioned two probable diagnoses of STD's in a case of a genito-urinary symptom. Errors in the diagnosis of vaginal discharge were significantly more frequent in the paramedical staff than in the medical one (p < 0.01), likewise the number of erroneous diagnoses of urethral discharge was more significant among the health agents in the private sector than those in the public sector (p = 0.04). The number of medical laboratories and the range of medical tests conducted in the town were lacking. Moreover, the results of these tests were not taken into account while prescribing the treatment for STD's. CONCLUSIONS: The above observations indicate a limited capability of the urban health services in establishing diagnoses of STD's on a rational basis. The transformation of current laboratory activities to a centralized specialized laboratory capable of a) identifying microorganisms circulating in the town and determine their magnitude and sensitivity to the main antimicrobials available, b) maintain regular surveillance of microorganism susceptibilities, and c) ensure quality control of laboratory tests conducted at a lower geographical level would contribute to the information and training of health care personnel and better acceptability of diagnosis and treatment strategies by this personnel. PMID- 10519173 TI - [Arterial blood pressure in homozygote patients with drepanocytosis]. AB - BACKGROUND: Relative hypotension has been reported in sickle cell patients. The aim of this study was to compare blood pressure in patients with SS disease and subjects with normal hemoglobin genotype AA and to assess whether the same clinical, biological and socio-demographic variables are associated to the mean arterial pressure in patients with sickle cell disease and normal subjects. METHOD: Blood pressure was measured with a standardized automated oscillometric method in 88 SS patients et 88 AA control subjects seen in the University Hospital of Pointe-a-Pitre (Guadeloupe). A multiple linear regression analysis for mean arterial pressure was done including type of hemoglobin (forced variable), age, sex, body mass index, pulse rate, hemoglobin concentration and interaction terms between type of hemoglobin and other variables. A regression was also fitted separately for each population. A downward stepwise strategy was used to simplify the models. RESULTS: The two groups were similar for age, height and gender ratio and pulse rate. Mean arterial pressure was significantly lower in sickle cell patients (81.6 mmHg in SS patients vs 89.9 mmHg in AA subjects, p < 10(-4)). The final model included type of hemoglobin, age, sex, body mass index, pulse rate and an interaction between type of hemoglobin and age (global F = 22.04, adjusted R2 = 42%). The separate models indicated that sex was associated with mean arterial pressure only in patients with sickle cell disease and that age and hemoglobin concentration was associated with mean arterial pressure only in normal subjects. CONCLUSION: Blood pressure determinants are not similar in the two populations. The effect of age, especially, is not the same in patients with sickle cell disease and in normal subjects. These results confirm that specific patho-physiological models should be defined in sickle cell disease. PMID- 10519174 TI - [Evaluation of mass screening for breast cancer in the Somme district (France) between 1990 and 1996]. AB - BACKGROUND: Randomized clinical trials have demonstrated that screening by mammography can reduce breast cancer mortality by 30% in women aged 50 to 69 years. In France, pilot screening programs were begun in 1989 and particularly in the Somme district. After 6 years and two rounds, the aim of this study was to assess the quality and the effectiveness of this screening program. METHODS: The quality of the campaign was measured concerning effectiveness (recall rate, positive predictive value of screening test or of the screening procedure, intervention rate, breast cancer detection rate) or quality (percentage of in situ cancers, invasive cancers smaller than 10 mm, invasive cancers without lymph node invasion). These indicators were compared with European targets. RESULTS: Attendance rates at 6 years ranged from 33.6% to 37.1%. The recall rate was 14.3% in the first and 7.8% in the second round, 6.4 and 6 cancers were detected per 1,000 women screened during the first and second round, and 38.5% of invasive cancers were of 10 mm or less (27.4% during the first round) and 70.8% had no nodal involvement (68.3% during the first round). CONCLUSION: Results from the screening system are variable yet satisfactory overall, due to the progressive implementation of quality assurance. However attendance rates remain low, whereas actual coverage is 64% in Somme district. PMID- 10519175 TI - [Prognostic factors of secondary complications in pediatric traumatology: a prospective study of 700 patients]. AB - BACKGROUND: The objective of the study was to estimate prevalence of therapeutic complications in a department of orthopedics pediatric surgery, and to determine predictors of minor and serious complications. METHODS: Between November 1996 and 1997, children less than or equal to 17 years of age, admitted for bone fracture (head and face excepted) were selected in a prospective study. In all, 700 children were treated until bone consolidation. Individual, socio-economic, anatomical, environmental and therapeutic factors were progressively registered. All complications were noted. RESULTS: The mean age of the children was 9.2 years; 25 patients could not be traced. Prevalence rate of therapeutic complications was 10.3%. In multivariate analysis using logistic regression, six factors increased the risk of adverse event: kind of fracture (diaphysial), comminuted aspect, compounded fracture and bone displacement, season (spring and autumn), and use of medical transport between accident place and hospital. The same factors, season and compounded fracture excepted, were predictors of serious complications. CONCLUSION: The results show that therapeutic complications depend on anatomical and environmental factors above all, and that medical procedure is not a prognosis factor. So if prevention of fractures is possible before injury, a better knowledge of initial risk can improve prevention of complications. PMID- 10519176 TI - [Guidelines on difficult intubation in anesthesia: evaluation of 2 information diffusion methods]. AB - BACKGROUND: In 1996, experts from the Societe Francaise d'Anesthesie et de Reanimation published guidelines about difficult intubation. We aimed to assess the effectiveness of two diffusion methods of these guidelines, media versus direct mailing plus media diffusion, and the relation between reading of the guidelines and practice behavior and training willingness. METHODS: Data were collected in two different samples of 300 anesthetists from three regions for pre and post-intervention surveys (E1 and E2 samples). Half of the anesthetists from E2, randomly chosen, received a direct mailing of the guidelines (E2a sample). The remaining constituted the E2b sample. Three assessment criteria were used, two concerning practice behavior and one training willingness. Relationship between these criteria and diffusion methods and reading was tested using logistic regression. RESULTS: The response rates were respectively 91%, 80% and 78% in the E1, E2a and E2b samples. The socio-professional features were not statistically different between the three samples. There was no relationship between the criteria and the diffusion methods. The direct mailing did not increase the reading rate (81% and 82% respectively in the E2a and E2b samples). The rate of anesthetists who routinely screened for predictive signs of difficult intubation (one of the practice criteria) was higher in E2a than in E2b (28% and 12% respectively). In the multivariate analysis, the difference only appeared among the sub-group of anesthetists who did not receive the direct mailing. The private practice was associated with a lower rate of routine screening. CONCLUSION: No impact of the diffusion methods on practice behavior and training willingness was found. Reading was inconstantly associated with practice behavior. PMID- 10519177 TI - [Air pollution and health: a synthesis of longitudinal panel studies published from 1987 to 1998]. AB - This article presents a synthetic study of 31 epidemiological panel investigations (published between 1987 and 1998) analyzing how air pollution and health may be related. First, the methodology of these studies is described: choice of population, exposure and health assessment, confounding factors and statistical methods. The main results are presented as follows: for pulmonary performance, respiratory symptoms and medication use per pollutant (PM10 and particles, SO2, O3, NO2). Then the interactions among pollutants or implying other environmental factors are analyzed. Most of the time, an association with particles is found, no matter the health indicator considered. An O3 effect is more often observed with lung function tests than with respiratory symptoms or medication use, the latter being linked first to particles and secondly to SO2. A NO2 effect is more rarely shown. Pulmonary performances are affected by all pollutants. Respiratory symptoms related to air pollution are mostly low respiratory symptoms and cough, particularly with particles and O3. The paper ends by discussing the interest and limits of these studies. It also emphasizes the importance of exposure assessment. Indeed, in order to improve exposure assessment, an essential step in every epidemiological study, new tools need to be developed. PMID- 10519178 TI - [Generalized ROC criteria in the evaluation of several tumor markers]. AB - The main objective of this paper is to present a method of evaluating several tumor markers by using the generalized ROC criterion. This criterion finds the best linear combination of the tumor markers such that the area under the ROC curve is maximized. Confidence intervals for the generalized ROC criteria are also presented. This methodology is applied to 51 patients with advanced colorectal cancer for whom the ACE tumor markers were measured before and during chemotherapy treatment. Two populations were defined according to clinical response to chemotherapy. Each marker taken separately, whether on the raw scale or on the transformed scale, contained 0.5 in the confidence interval and was thus non significant. This was also true for both markers on the raw scale. However, the best linear combination on the logarithms of ACE before and at evaluation gave a significantly better area under the ROC curve. A weighted change in ACE measurements significantly distinguishes between responders and non responders in patients with advanced colorectal cancer. We propose that the methodology presented in this paper be used for the evaluation of several tumor markers. PMID- 10519179 TI - [Obesity in France: contribution of the INSEE-CREDES survey on health and medical care]. PMID- 10519180 TI - [Exclusion of low-risk women from screening programs for cervix uteri cancers: based on mathematical modeling]. PMID- 10519181 TI - [The mode of delivery and the risk of fetomaternal transmission of HIV. A meta analysis of 15 prospective cohort studies]. PMID- 10519182 TI - [Importance of laboratory investigations and trigger factors in chronic urticaria]. AB - Urticaria is one of the most frequent skin disorders. Whereas for the acute form a cause is usually found, the aetiology of chronic urticaria often remains obscure. Infectious or autoimmune origin are presumed aetiologies, whereas trigger factors such as pressure, cold or food additives often induce an urticarial episode. In this study, we investigated the significance of laboratory and supplementary analysis in relation to the aetiology and classification of chronic urticaria. We also looked at a correlation of trigger factors with the aetiology and course of chronic urticaria. Out of 170 patients with chronic urticaria referred to our outpatient allergy clinic within 3 1/4 years, 95 were female (56%) and 75 male (44%). The average age was 37 years. Based on history and clinical signs, laboratory, allergo-immunological, stool and urine samples were performed, as well as allergological skin and physical tests. Of the laboratory parameters, total leukocyte count, C-reactive protein (CRP) and alanine-amino-transferase (ALAT) were the findings most often out of the normal range. In 25% (43/170) chronic urticaria could be attributed to a possible cause (infection [15%], autoimmunity [8%], allergy [1%], urticaria pigmentosa [1%]). Trigger factors were found in 84/170 (49%) patients (physical [29%], pseudoallergic [12%], combination of both [8%]). Follow-up after an average of 22.3 months revealed that 84 patients (63%) no longer suffered from urticarial disorders, while 49 (37%) still complained of hives. In conclusion, laboratory and supplementary investigations were rarely helpful in identifying aetiologic agents, although in 25% chronic urticaria was classified. Trigger factors are not of predictive value either for aetiology or course of chronic urticaria. However, the longer chronic urticaria lasts, the rarer are remissions. In younger patients, chronic urticaria tends to last less long than in elderly persons. PMID- 10519183 TI - [Pediatric renal transplantation: Experience in Lausanne 1971 to 1998]. AB - AIMS OF THE STUDY: Analysis of indications and results of paediatric renal transplantation in a single centre, before and after the introduction of cyclosporine A (CSA). METHODS: Historical retrospective study. RESULTS: 19 transplantations were performed in 14 patients (5 second grafts) between 1971 and 1987 (group I). 13 patients were transplanted between 1988 and 1998 (no second transplant) (group II). In group II, all the patients had immunosuppression with CSA, but none in group I. Group II, with CSA, showed better renal survival than patients without CSA. In group I, obstructive uropathies (posterior urethral valves, pyelo-ureteral junction stenosis, vesico-ureteral reflux) represent a common cause (35%) of terminal chronic renal failure (TCRF), whereas in group II they represent only 15% of the causes and chronic glomerulonephritis is the most common cause (69%) of TCRF. Acute and chronic graft rejections were the cause of 9 and 1 graft losses in group I and II respectively. Living related donors account for 14% of all renal transplantations in group I and 46% in group II. CONCLUSIONS: The incidence of paediatric patients referred to Lausanne for TCRF is stable. We have observed a constant and steady decrease in obstructive uropathies leading to TCRF and renal transplantations, whereas glomerulonephritis are increasingly frequent. Graft survival has much improved since the introduction of cyclosporine A, without an increase in morbidity. In carefully selected cases, intrafamilial renal transplantation provides good results and helps to shorten the time spent on dialysis. PMID- 10519184 TI - [Pica--a qualitative appetite disorder]. AB - Pica is the collective term for any form of qualitative disorder of eating behaviour. If a patient's deviant appetite is fixated on one special object, there are quite a number of corresponding terms available (e.g. geophagia for eating dirt). Pica, although a ubiquitous phenomenon, shows heightened prevalence in certain high-risk populations, i.e. infants, pregnant women, the mentally retarded, special ethnic groups and psychiatric patients. The author briefly summarizes the medical history of the pica concept and the present state of knowledge concerning aetiology and pathogenesis. The numerous possible complications and various therapeutic approaches are pointed out. PMID- 10519185 TI - [A "typical" case of pulmonary embolism]. AB - A 85-year-old woman was admitted to our hospital because of a presumtive diagnosis of pulmonary thromboembolism. The patient presented with a history of progressive dyspnoea and retrosternal pain. 3-4 weeks ago she had noticed a swollen left leg. On examination a 4/6-pansystolic murmur was found. An arterial blood gas analysis showed a reduced oxygen saturation. An electrocardiogram revealed deep S-waves in lead I and pathological Q-waves in lead III. The chest X ray showed cardiomegaly, a pulmonary nodule and an ill-defined opacity inferioposteriorly. Ventilation-perfusion mismatch was demonstrated by lung ventilation-perfusion scanning. Transthoracic echocardiography showed pulmonary hypertension and tricuspid regurgitation. On the 20th hospital day the patient died from multi organ failure. Pulmonary thromboembolism secondary to deep venous thrombosis of the lower extremities was the most likely diagnosis. In view of the patients' history of night sweat, loss of appetite and weight loss a malignant process had to be taken into consideration. A tumor originating from the right ventricle, the right ventricular outflow tract or the pulmonary artery was compatible with the clinical picture of multiple pulmonary emboli. On autopsy a polymorph cellular sarcoma measuring 6 x 3 x 3 cm was found in the right ventricular outflow tract. Section of the lung revealed a single pulmonary metastasis and multiple thromboemboli of various age. Pulmonary artery sarcomas, as described in our case, are extremely rare. The prognosis is poor and often the diagnosis is only made on autopsy. PMID- 10519186 TI - [Lockjaw]. PMID- 10519187 TI - Evidence-based medicine and clinical freedom. PMID- 10519189 TI - [Evidence-based medicine in cardiology with an example of secondary prevention of coronary heart disease]. AB - The fear that application of EBM could lead to a cook-book type medicine has not been substantiated. The challenge is to evaluate and treat the individual patient according to his/her particular situation. The goal of secondary prevention is the reduction of the global risk. As patients with coronary artery diseases have a high event rate (> 20% in 10 years), intensive measures with proven benefit for risk reduction should be undertaken. Guidelines of national or European societies are helpful for the daily routine. However, the application of these guidelines is less widely used than expected which leads to less than optimal treatment. This is a universal phenomenon. The reasons may differ in different countries. Financial problems may be one limiting factor. However, the overal costeffectiveness of these measures for the social system is usually not taken into account. Educational efforts directed to patients, physicians and health care providers will be necessary to overcome this problem. PMID- 10519188 TI - [Epidemiologic evidence in the treatment of cardiovascular risk factors in Germany]. AB - Up to now, approaches neither in primary nor in secondary prevention led to a relevant decline in prevalences of hypertension and hypercholesterolemia. On the contrary, prevalence of hypercholesterolemia increased from 38 to 40% (p < 0.001) while the hypertension prevalence stagnated (at nearly 20%) in the 30 to 69 years old population in West Germany between 1984/85 and 1991/92. The potential of medical advice is not sufficiently used, the compliance of patients regarding medical recommendations to change health behavior is limited. Subsequently the scientific consensus with regard to benefits from normalization of hypertension and hypercholesterolemia does not become effective at the population level. Evidence based medicine needs more efficient evaluation of continued medical education and quality assurance. PMID- 10519190 TI - [Are principles of evidence-based medicine taken into account in the area of pharmacology?]. AB - Scientific literature published in the early 1980s claimed that only 15% of medical interventions are based on scientific evidence (57). Lately, a number of published articles concluded that ca. 80% of the interventions in family medicine (18, 25) and ca. 60% of those in psychiatry fulfil these criteria (24). Did medical practice change so fundamentally? Hard to believe. But to what extent do physicians practice evidence-based medicine as newer publications also constitute a level of science-based interventions across all medical disciplines of only 21% (17)? This article tries to highlight and discuss which observations can be made regarding the compliance to principles of 'evidence based medicine' in one of the most important field of medical interventions--drug therapy. Taking the submarkets of cardiovascular and respiratory drugs as an example, the authors try to identify and discuss effects and obstacles to implement evidence-based pharmacotherapy, and how the latter might be overcome in the future. PMID- 10519191 TI - [Evidence-based medicine from the point of view of health insurance companies--a frame for qualified freedom of therapy and an improved economy?]. AB - The rationality and efficiency of medical care must be improved from the point of view of the statutory health insurance system in Germany. Too many diagnostic and therapeutic procedures are of doubtful usefulness, the high amount of many procedures is unnecessary. 10% of all expenditures in the health care sector or around 25 billion DM could probably be saved if evidence based guidelines are taken as a basis for decisions in medical care. Evidence based medicine not only improves the efficiency, but supports also the therapeutic freedom of the physicians in limiting the vagueness of many medical decisions. EBM is therefore a strategy of the sick fund systems that supports the quality and efficiency of the medical care for the insured people. PMID- 10519192 TI - [Parameter of evidence-based medicine in health care economics]. AB - In the view of scarcity of resources, economic evaluations in health care, in which not only effects but also costs related to a medical intervention are examined and a incremental cost-outcome-ratio is build, are an important supplement to the program of evidence based medicine. Outcomes of a medical intervention can be measured by clinical effectiveness, quality-adjusted life years, and monetary evaluation of benefits. As far as costs are concerned, direct medical costs, direct non-medical costs and indirect costs have to be considered in an economic evaluation. Data can be used from primary studies or secondary analysis; metaanalysis for synthesizing of data may be adequate. For calculation of incremental cost-benefit-ratios, models of decision analysis (decision tree models, Markov-models) often are necessary. Methodological and ethical limits for application of the results of economic evaluation in resource allocation decision in health care have to be regarded: Economic evaluations and the calculation of cost-outcome-rations should only support decision making but cannot replace it. PMID- 10519193 TI - [How do patients evaluate general practice? German results from the European Project on Patient Evaluation of General Practice Care (EUROPEP)]. AB - German results from the European Project on Patient Evaluation of General Practice Care (EUROPEP). As part of the European collaborative study, "EUROPEP" 2224 patients in Germany (response rate 77.2%) filled in a questionnaire to evaluate their general practitioners (GP's) and the care they were delivering over the last 12 month. Overall satisfaction was high: 95.4% reported that they had no reasons to change to another GP. On a five-point scale ranging from 'excellent' to 'poor' the possibility to reach the practice on the phone was evaluated best (73.8% 'excellent') and waiting times were rated worst (31.0% 'excellent'). Also less often rated 'excellent' than other items were 'preparing the patient for what to expect from specialist or hospital care' (53.3%), 'the GP's knowledge on what s/he had done or told the patient during previous contacts' (54.3%) and 'helping the patient to deal with emotional problems related to her/his health status' (54.6%). Relevant differences between practices concerning evaluation of care and sociodemografic background of their patients were detected. Patient evaluation of care can contribute to make practices an their teams more responsive to patients needs. PMID- 10519194 TI - [The Cologne Guidelines Conference: computer-assisted clinical practice guidelines in clinical diagnosis]. AB - Eighteen clinical practice guidelines on interdisciplinary diagnostic issues were developed at the University Hospital of Cologne, Germany. The guideline committee is organized and directed by the quality control program, which also includes the local Cochrane initiative and a wide range of organizational topics. During guideline development, questions of differential diagnosis were addressed to the same extent as the organizational and financial realization. Broad consideration was given to medicolegal implications, but need for interspecialty cooperation was judged to be more critical and even more relevant in this regard. Guidelines were primarily developed as algorithms and translated to text versions secondarily. Critical steps in the decision tree were supported by rated literature and recommendations weighte by criteria as used in evidence-based medicine. For implementation, guidelines were presented to colleagues in a series of short lectures, as print versions containing all literature used in the developing process, and in hypertext format, which is accessible via intranet. Three levels of presentation were chosen in the html-version: algorithm, decision, and information. The former is due to orientation in the guideline, the second displays the binary question, and the latter makes the scientific background available, together with literature and links for more information. Efforts to check effectiveness are currently been made, questions of efficiency will be addressed in future. PMID- 10519195 TI - [The current concepts on the mechanism of energy transformation by molecular motors of differing natures]. PMID- 10519196 TI - [The basic trends in research on the intellectual activities and social behavior of anthropoids]. PMID- 10519197 TI - [The possible role of spiral structures in the functional activity of the regulatory and catalytic subunits of phosphatidylinositol-3-kinases]. PMID- 10519198 TI - [Glycogen, glucose, lactate and pyruvate in the organs of 3 species of adult seals (Pusa sibirica, P. caspica, Phoca vitulina)]. PMID- 10519199 TI - [The interrelationships of the autorhythmic contractile activity of the skeletal musculature and of the smooth musculature of the internal organs]. PMID- 10519200 TI - [The mechanisms of the action of vasoactive intestinal peptide on rat tracheal smooth muscle]. PMID- 10519201 TI - [The cardiotropic activity of the nucleotides found in the composition of the low molecular peptide subfraction obtained from the brain of hibernating susliks Citellus undulatus]. PMID- 10519202 TI - [The optimization of an instrumental-conditioning food-acquisition habit in Macaca mulatta monkeys in a time-deficit situation]. PMID- 10519203 TI - [The synaptic organization of the hypothalamic arcuate nucleus in rats]. PMID- 10519204 TI - [A lactose-specific lectin from the brain of chickens: its isolation and biochemical characteristics]. PMID- 10519205 TI - [The resistance of the muskrat Ondatra zibethicus to hypoxic exposures during diving]. PMID- 10519206 TI - [Biochemical mechanisms of the adaptation in vertebrate to the changing environment: the role of lipids]. PMID- 10519207 TI - [The contribution of evolutionary endocrinology to study of the structure and mechanism of the insulin effect]. PMID- 10519208 TI - [Current problems in biochemistry and physiology of phospholipase D]. PMID- 10519209 TI - [Effect of immunization to biologically active agents from tissues of hibernating animals on the thermoregulation and cardiovascular system in rats and mice]. PMID- 10519211 TI - [Paradoxical sleep and the brain temperature: seasonal interactions of euthermia ("normothermia") and hypometabolism in hibernating susliks Citellus major]. PMID- 10519210 TI - [Effects of gangliosides on the motor activity, learning, and mechanisms of signal transduction regulation in the rat brain]. PMID- 10519212 TI - [Modern theories on endosymbiotic origin of eukaryotic cells]. PMID- 10519213 TI - In vitro replication of Nosema algerae (Microsporidia), a parasite of anopheline mosquitoes, in human cells above 36 degrees C. AB - Microsporidia form a large and ubiquitous group of obligately intracellular parasitic eukaryotes, increasingly recognized as pathogens in humans. Transmission of invertebrate microsporidia to mammals has been considered impossible because temperature seemed to be a limiting factor for development. Nosema algerae, a microsporidian of anopheline mosquitoes, was cultured in human muscle fibroblasts at temperatures of 31 degrees C and 38 degrees C. This is the first record of an invertebrate microsporidian developing in human cells at a temperature above 36 degrees C. The ultrastructure of N. algerae growing in human muscle fibroblasts is similar to that of Brachiola vesicularum, a microsporidian species previously described in the muscle of an AIDS patient. PMID- 10519214 TI - Actin of Histriculus cavicola: characteristics of the highly divergent hypotrich ciliate actins. AB - A macronuclear gene-sized molecule carrying an actin gene from the hypotrich ciliate, Histriculus cavicola, was characterized. Southern blot analysis using a coding region probe suggested that actin in H. cavicola is encoded by a single gene. A comparison of the promoter regions indicated that the H. cavicola actin gene has a TATA box in the 5' flanking region in a position identical to those in other oxytrich ciliates. The coding sequence of this gene is not interrupted by any introns, and codes for a protein of 375 amino acid residues. This protein shares a high degree of similarity with other oxytrichid actins, and a relatively low similarity with actins from other eukaryotes. Comparative analyses of sequences indicated that most of the amino acid substitutions in hypotrich actins are found in surface loops, while the core structures are well-conserved. The sites that interact with DNase I and several regions involved in actin-actin contact have diverged considerably in hypotrich actins, while nucleotide-binding sites are the best-conserved interaction motif. PMID- 10519215 TI - Fish trypanosomes: their position in kinetoplastid phylogeny and variability as determined from 12S rRNA kinetoplast sequences. AB - Fish trypanosomes have traditionally been classified according to the host species from which they were isolated, each isolate being regarded as a distinct species. To test the soundness of this practice, the genetic variabilities of the kinetoplast 12S rRNA-encoding genes of different fish trypanosomes isolates were compared. The DNAs were extracted from trypanosomes cloned from blood samples of 15 donors representing ten different fish species in four orders from waters of three major river systems of Central and Northern Europe. Comparison with other trypanosomatid sequences revealed that the fish trypanosomes form a monophyletic group with Trypanosoma brucei as a sister group. Pairwise comparisons of genetic distances yielded a wide range of continuous variation with no indication of any discontinuities attributable to barriers to gene flow. The genetic distances did not correlate with either the identity of the host species or geography. The host specificity of fish trypanosomes appears to be limited. PMID- 10519216 TI - A novel member of the cyclin-dependent kinase family in Paramecium tetraurelia. AB - Passage through the cell cycle in eukaryotes requires the successive activation of different cyclin-dependent protein kinases. Here, we describe the identification and characterization of a novel class of cyclin-dependent protein kinase, termed Cdk2, in the ciliate Paramecium tetraurelia. It is 301 amino acids long, 7 amino acids shorter than Cdk1, the CDK that is associated with macronuclear DNA synthesis. All the catalytic domains typical of protein kinases can be located within the sequence and putative regulatory phosphorylation sites equivalent to Thr14, Tyr15, and Thr161 in human CDK1 are also conserved. The 'PSTAIRE' region characteristic of most CDKs is perfectly conserved. Cdk2 shares only 48% homology to Cdk1 at the amino acid level, suggesting that the evolutionary separation of Cdk1 and Cdk2 is ancient, and implying that they have different roles in cell cycle regulation. Like Cdk1, Cdk2 does not bind to yeast p13suc1, even though it has better conservation of p13suc1 binding sites than Cdk1 does. The Cdk2 protein level is relatively constant throughout the vegetative cell cycle. Cdk2 exhibits kinase activity towards bovine histone H1 in vitro with the maximal level late in the cell cycle, suggesting it may be involved in the regulation of cytokinesis. Our results further support the view that an analogue of the cyclin-dependent kinase cell cycle regulatory system like that of yeast and higher eukaryotic cells operates in Paramecium and that a family of cyclin-dependent kinases may control different aspects of the Paramecium cell cycle. PMID- 10519217 TI - Ultrastructural characterisation and molecular taxonomic identification of Nosema granulosis n. sp., a transovarially transmitted feminising (TTF) microsporidium. AB - A novel microsporidian parasite is described, which infects the crustacean host Gammarus duebeni. The parasite was transovarially transmitted and feminised host offspring. The life cycle was monomorphic with three stages. Meronts were found in host embryos, juveniles, and in the gonadal tissue of adults. Sporoblasts and spores were restricted to the gonad. Sporogony was disporoblastic giving rise to paired sporoblasts, which then differentiated to form spores. Spores were not found in regular groupings and there was no interfacial envelope. Spores were approximately 3.78 x 1.22 microns and had a thin exospore wall, a short polar filament, and an unusual granular polaroplast. All life cycle stages were diplokaryotic. A region from the parasite small subunit ribosomal RNA gene was amplified and sequenced. Phylogenetic analysis based on these data places the parasite within the genus Nosema. We have named the species Nosema granulosis based on the structure of the polaroplast. PMID- 10519218 TI - Characterization of a Sarcocystis neurona isolate (SN6) from a naturally infected horse from Oregon. AB - An isolate of Sarcocystis neurona (SN6) was obtained from the spinal cord of a horse from Oregon with neurologic signs. The parasite was isolated in cultures of bovine monocytes and equine spleen cells. The parasite divided by endopolygeny and completed at least one asexual cycle in cell cultures in three days. Two gamma interferon knockout mice inoculated with cell culture-derived merozoites became ill 35 d later and S. neurona schizonts and merozoites were found in encephalitic lesions. The parasite in tissue sections of mice reacted with S. neurona-specific antibodies and S. neurona was reisolated from the brain of knockout mice. PMID- 10519219 TI - A novel healing filament in ciliate regeneration. AB - The interphase cells of the hypotrich ciliate Paraurostyla weissei possess a complex fibrillar system surrounding basal bodies in the compound ciliary assemblages, cirri and membranelles. During replacement of the ciliature at cell division, transient filaments precede and accompany the development of ciliary primordia and participate in the formation of the fission furrow. Both fibrillar systems are recognized by monoclonal antibody FXXXIX 12G9. We studied regeneration of cellular fragments after transection employing the mAb 12G9 and found a new cytoskeletal structure involved in healing of the excisional wound. The healing filament is formed at the wound edge, distally and in connection with the bases of cirri closest to the wound. It is visible 5 min after transection. Concomitant with development of new ciliary primordia, the healing filament shrinks and finally disappears together with other transient fibers formed in this process. Ultrastructural analysis of immunolabeled regenerating cells revealed that structures recognized by mAb 12G9 contain fine filaments whose packing and arrangement depends on accompanying cytoplasmic elements and the developmental status of a fragment. Assembly of the healing fiber does not depend on microtubules and microfilaments since it develops in cellular fragments exposed to cold, nocodazole, and Cytochalasin D. On Western blots of whole cell and cytoskeletal extracts of P. weissei the 12G9 antibody identified one protein band whose molecular weight corresponds to 60 kDa. PMID- 10519220 TI - Factors from Trypanosoma cruzi interacting with AP-1 sequences. AB - Interaction between factors from Trypanosoma cruzi extracts and AP-1 sequences was studied by electrophoretic mobility shift assays. Using a double-stranded probe carrying the AP-1 sequence from the SV40 promoter, three specific complexes designated A, B, and C were detected. Complexes A and C were formed when using single-stranded probes. The relative amount of complex B, specific for double stranded DNA, increased as a function of probe length. Complexes were stabilized by cross-linking with UVC irradiation and resolved on denaturing SDS-PAGE. Complex A generated bands of 60- and 39 kDa; complex B produced two bands of 46- and 43 kDa; and complex C generated one band of 43 kDa. The AP-1 binding activity was much higher in purified nuclear preparations than in soluble fractions, and was detected in crude extracts from the three forms of the parasite. The binding signal, however, was much stronger in amastigote and trypomastigote than in the epimastigote forms. Specific binding was increased by oxidative stress. Antibodies raised against peptides corresponding to conserved domains of mammalian c-Jun and c-Fos detected bands of 40- and 60 kDa, respectively, in a nuclear epimastigote preparation. PMID- 10519221 TI - Construction of a normalized cDNA library for the Trypanosoma cruzi genome project. AB - Sequencing of the Trypanosoma cruzi genome is underway. Expressed sequence tags, obtained from cDNA libraries, facilitate mapping and gene discovery. The efficiency of large-scale generation of such tags is increased when using normalized cDNA libraries, where the frequency of individual clones is brought within a narrow range. Repetitive sequencing of abundant clones is therefore minimized. We constructed a normalized cDNA library from epimastigotes of clone CL Brener, and the efficiency of normalization of representative clones was assessed and shown to be adequate. The normalized cDNA library has been distributed to several groups and large-scale sequencing is currently in progress. PMID- 10519222 TI - Characterization of an alpha-tubulin gene of Cryptosporidium parvum. AB - A gene encoding an alpha-tubulin of Cryptosporidium parvum was isolated and characterized. It had no introns, and encoded a 441-amino acid protein whose predicted ORF represented a typical alpha-tubulin protein with a MW of 50.5 kDa. This tubulin had an amino acid sequence similarity with Apicomplexa Plasmodium falciparum and Toxoplasma gondii higher than 88% and shared a number of conserved motifs. PMID- 10519223 TI - Microsporidia at the turn of the millenium: Raleigh 1999. PMID- 10519224 TI - Enterocytozoon bieneusi: a common opportunistic parasite in lungs of HIV-positive patients? PMID- 10519225 TI - Enterocytozoon bieneusi in animals: rabbits and dogs as new hosts. PMID- 10519227 TI - Environmental resistance of Encephalitozoon spores. PMID- 10519226 TI - Isolation of Nosema algerae from the cornea of an immunocompetent patient. PMID- 10519228 TI - Characterization of Nosema algerae isolates after continuous cultivation in mammalian cells at 37 degrees C. PMID- 10519229 TI - Microsporidian molecular phylogeny: the fungal connection. PMID- 10519230 TI - Isolation of a highly divergent gene encoding CDC2-related protein kinase from Encephalitozoon cuniculi (Microspora). PMID- 10519231 TI - Isolating expressed microsporidial genes using a cDNA subtractive hybridization approach. PMID- 10519232 TI - Application of differential display RT-PCR to the analysis of gene expression in a host cell-microsporidian E. cuniculi interaction. PMID- 10519233 TI - Sequencing of several protein-coding genes of the chromosome X from the microsporidian Encephalitozoon cuniculi. PMID- 10519234 TI - Analysis of the major microsporidian polar tube proteins. PMID- 10519235 TI - Polyamine synthesis in Encephalitozoon cuniculi. PMID- 10519236 TI - Production of immunological probes raised against Enterocytozoon bieneusi and Encephalitozoon intestinalis, two microsporidian species causing intestinal infections in man. PMID- 10519237 TI - Susceptibility to apoptosis is reduced in the Microsporidia-infected host cell. PMID- 10519238 TI - Modifications of the cytoskeleton in Encephalitozoon-infected cells. PMID- 10519239 TI - Recent trends in Cryptosporidium research: workshop summary. PMID- 10519240 TI - Emerging and opportunistic intestinal parasites in HIV-infected patients with chronic diarrhea in Rio de Janeiro, Brazil. PMID- 10519241 TI - Which genotypes/species of Cryptosporidium are humans susceptible to? PMID- 10519242 TI - Presence of heterogeneous copies of the small subunit rRNA gene in Cryptosporidium parvum human and marsupial genotypes and Cryptosporidium felis. PMID- 10519244 TI - Cryptosporidium parvum: lectins mediate irreversible inhibition of sporozoite infectivity in vitro. PMID- 10519243 TI - Microsatellite analysis of the human and bovine genotypes of Cryptosporidium parvum. PMID- 10519245 TI - Identification of a putative telomeric repeat DNA binding factor of Cryptosporidium parvum. PMID- 10519246 TI - Preliminary evidence for a mitochondrion in Cryptosporidium parvum: phylogenetic and therapeutic implications. PMID- 10519247 TI - Cryptosporidium parvum: synchronized excystation in vitro and evaluation of sporozoite infectivity with a new lectin-based assay. PMID- 10519248 TI - Characterization of new monoclonal antibodies against Cryptosporidium parvum sporozoites. PMID- 10519249 TI - Effects of Lactobacillus reuteri on Cryptosporidium parvum infection of gnotobiotic TCR-alpha-deficient mice. PMID- 10519250 TI - Improved efficacy of dinitroaniline analogs for use as anti-cryptosporidial drugs. PMID- 10519251 TI - Changes in murine intestinal epithelium following Cryptosporidium parvum infection. PMID- 10519252 TI - Change in the phenotypic profiles of spleen and intestinal lymphocytes in dexamethasone-treated mice infected with Cryptosporidium parvum. PMID- 10519253 TI - Current status of research on Toxoplasma gondii: report from the Sixth International Workshops on Opportunistic Protists. PMID- 10519254 TI - Transmission to guinea pigs of very low doses of oocysts of Toxoplasma gondii in drinking water. PMID- 10519255 TI - Survival of Toxoplasma gondii tachyzoites in goat milk: potential source of human toxoplasmosis. PMID- 10519256 TI - Brain slices organotypic culture, a new model to study Toxoplasma gondii infection. PMID- 10519257 TI - Different PCR systems to detect Toxoplasma gondii tachyzoites or bradyzoites in clinical specimens from patients with and without overt disease. PMID- 10519259 TI - Cytochrome b mutation identified in a decoquinate-resistant mutant of Toxoplasma gondii. PMID- 10519258 TI - Targeting of a nuclear encoded protein to the apicoplast of Toxoplasma gondii. PMID- 10519260 TI - Effect of cytokines and quercetin on Toxoplasma gondii cyst induction in murine astrocytes. PMID- 10519261 TI - Summary of the Pneumocystis research presented at the 6th International Workshop on Opportunistic Protists. PMID- 10519262 TI - Pneumocystis pneumonia, an immunodeficiency-dependent disease (IDD): a critical historical overview. PMID- 10519263 TI - New trend in the epidemiology of Pneumocystis carinii pneumonia (PCP) among AIDS patients. PMID- 10519264 TI - Impaction versus filtration for the detection of Pneumocystis carinii DNA in air. PMID- 10519265 TI - Putative Pneumocystis dormant forms outside the mammalian host, and long-term culture derived from them: initial characterizations. PMID- 10519266 TI - Acquisition and biodiversity of Pneumocystis carinii in a colony of wild rabbits (Oryctolagus cuniculus). PMID- 10519267 TI - Comparison of four methods for extraction of Pneumocystis carinii DNA from pulmonary specimens and serum. PMID- 10519268 TI - Evaluation of a nested PCR for detection of Pneumocystis carinii in serum from immunocompromised patients. PMID- 10519269 TI - Biomolecular approach for Pneumocystis carinii pneumonia (PCP) diagnosis and follow up in AIDS patients. PMID- 10519270 TI - Interactions of two Pneumocystis carinii populations within rat lungs. PMID- 10519271 TI - Pneumocystis carinii trophozoites in the lungs of patients without pneumocystosis. PMID- 10519272 TI - The minimum number of Pneumocystis carinii f. sp. carinii organisms required to establish infections is very low. PMID- 10519273 TI - P.carinii host specificity: attempt of cross infections with human derived strains in rats. PMID- 10519274 TI - Experimental inoculation of immunosuppressed owl monkeys with Pneumocystis carinii f. sp. hominis. PMID- 10519275 TI - Pneumocystis carinii growth kinetics in culture systems and in hosts: involvement of each life cycle parasite stage. PMID- 10519276 TI - Ability of Pneumocystis carinii cysts to seed cultures and infect animals. PMID- 10519277 TI - Mouse derived Pneumocystis carinii in an axenic culture system. PMID- 10519278 TI - Culture of Pneumocystis carinii sp.f. hominis. PMID- 10519279 TI - Distribution and biosynthesis of cis-9,10-epoxystearic acid in Pneumocystis carinii. PMID- 10519280 TI - Type-II major-surface-glycoprotein family of Pneumocystis carinii. PMID- 10519281 TI - Direct correlation of genomic localization and surface expression of the major surface glycoprotein of Pneumocystis carinii. PMID- 10519282 TI - Determination of the maximum frequency of genetic rearrangements associated with Pneumocystis carinii surface antigen variation. PMID- 10519283 TI - Characterization of the Cdc25 phosphatase in Pneumocystis carinii. PMID- 10519284 TI - Cdc2 gene of Pneumocystis carinii hominis and its expression during culture. PMID- 10519285 TI - Assembly of cell wall glucans by Pneumocystis carinii: characterization of the Gsc-1 subunit mediating beta-glucan synthesis. PMID- 10519286 TI - Identification of dihydropteroate (DHPS) gene mutant in Pneumocystis carinii in respiratory samples of HIV+ patients from 1992 to 1997. PMID- 10519287 TI - Contribution of dihydropteroate synthase gene typing for Pneumocystis carinii f.sp. hominis epidemiology. PMID- 10519288 TI - Successful treatment of PCP episodes caused by Pneumocystis carinii with mutant dihydropteroate (DHPS) gene. PMID- 10519289 TI - Recombinant major surface glycoprotein of Pneumocystis carinii elicits a specific immune response but is not protective in immunosuppressed rats. PMID- 10519290 TI - Production of anti-toxin antibodies by immunization with Pichia killer toxin-like antiidiotypic monoclonal antibodies. PMID- 10519291 TI - Glyphosate reduces organism viability and inhibits growth in vitro of Pneumocystis. PMID- 10519292 TI - Effects of sterol inhibitors on the ATP content of Pneumocystis carinii. PMID- 10519293 TI - First steps in the purification and characterization of a Pichia anomala killer toxin. PMID- 10519294 TI - Beta-glucan from Pneumocystis carinii stimulates TNF alpha release from alveolar macrophages. PMID- 10519295 TI - Pneumocystis infection is correlated with a reduction of the total sterol content of human bronchoalveolar lavage fluid. PMID- 10519296 TI - Production of plasminogen activator by alveolar macrophages in experimental Pneumocystis carinii pneumonia. PMID- 10519297 TI - Immunological characterization of surface subtilisin-like protease (SSP) of Pneumocystis carinii. PMID- 10519298 TI - Radiograph quality evaluation for exposure variables--a review. AB - Good quality radiographs are essential for making accurate diagnoses. Many factors influence the quality of radiographs including the x-ray machine specifications and settings, the darkroom environment and processing, and the choice of ancillary x-ray equipment (cassette properties, film/screen selection, use and properties of a grid). In compiling a technique chart, many of these variables are standardized so as to provide dependable guidelines for selecting the appropriate exposure settings (mAs and kVp) for a radiographic study. The systematic evaluation of image blackening, peripheral blackening, and the visibility of the gross image detail and contrast will facilitate the development of a technique chart as well as determining the source of the problem and necessary exposure setting changes for radiographs that are suboptimal. A flow diagram is described that will assist with the systematic evaluation of radiographic quality and provide guidelines for correcting exposure errors. PMID- 10519299 TI - Evaluation of different projections for radiographic detection of tarsal degenerative joint disease in Icelandic horses. AB - Radiographs from 196 tarsi in 98 Icelandic horses were evaluated to compare the accuracy of four different projections in detecting radiographic signs of degenerative joint disease in the distal tarsus. The extent and localization of tarsal degenerative joint disease found in one projection when reading all four projections of the same tarsus together was compared with the combined findings from all four projections. The results of reading individual radiographic projections without knowledge of the other three projections was also evaluated. Degenerative joint disease was detected most frequently in the plantarolateral dorsomedial oblique (P1L-DMO) projection. The location with the highest relative frequency of radiographic findings was the dorsolateral aspect of the centrodistal and tarsometatarsal joints respectively. Radiographic signs of active bone remodelling was detected in 30 (33%) and periarticular osteophytes in 51 (56%) of 91 tarsi with degenerative joint disease. PMID- 10519300 TI - Effects of diet and aging on renal measurements in uninephrectomized geriatric bitches. AB - Under controlled, but varied dietary conditions among geriatric, uninephrectomized Beagle bitches (dogs) observed for 4 years, renal size increase as assessed radiographically and ultrasonographically occurred at variable rates, but on a seemingly continuous basis. The maximum observed mean renal linear parameter increase found was approximately 15%. However, a 10 and 15% increase is a more representative expectation among the 4 parameters (sonographic length, radiographic length, sonographic width, radiographic width) under consideration. The rate of renal size increase was rapid during the first 2 to 3 months following uninephrectomy. Thereafter, the rate of increase was slow, but occurred to varying degrees in both the length and width as assessed radiographically or ultrasonographically. The mechanism creating the size change was hypertrophy, not hyperplasia. Within limits of the 3 diets used in the study, no significant diet effect was found on the rate or degree of long term compensatory hypertrophy. Radiographically and ultrasonographically measured renal length had the greatest correlation with each other as well as with post mortem measurements and are, therefore, the recommended parameter for imaging assessment of compensatory hypertrophy. When the prenephrectomy, radiographic renal lengths and widths were normalized as a ratio of the second lumbar vertebral body length (L2) measured from ventrodorsal radiographs, the diet group means across dogs (approximately three L2 lengths for renal length; two L2 lengths for renal width) were in the middle of the respective previously published normal radiographic ranges for mature dogs (e.g. 2.5 L2 < or = length < or = 3.5 L2; 1.58 L2 < or = width < or = 2.38 L2 lengths). Even after the hypertrophic changes occurred, the radiographic group mean lengths and widths across dogs were still within the specified normal ranges, although toward the upper end of the respective range. This information provides background for clinical interpretation of potential compensatory hypertrophy that may be encountered following uninephrectomy for spontaneous disease in aged dogs. In addition, it appears that available radiographic renal linear ranges for normal mature dogs are applicable to geriatric dogs as well. PMID- 10519301 TI - Effect of reproductive status on feline renal size. AB - Renal length and width dimensions were determined from survey radiography and excretory urography in 28 cats of various sex and reproductive status. Renal dimensions were expressed as a ratio to the length of the second lumbar vertebra. Renal dimensions were not significantly different when males were compared to females. However, significant differences in renal dimensions between intact and neutered cats were identified. Renal length ratios for neutered cats were: left kidney 2.22 +/- 0.14 (mean +/- standard deviation), right kidney 2.29 +/- 0.14. In intact cats, renal length ratios were: left kidney 2.60 +/- 0.19, right kidney 2.65 +/- 0.24. The mean renal length ratios for neutered cats was smaller than previously reported normal values. Thus, reproductive status should be considered when evaluating feline kidneys for alterations in size. Based on this study, normal feline renal length ratios range from 1.9 to 2.6 for neutered cats and 2.1 to 3.2 for intact cats. PMID- 10519302 TI - Magnetic resonance imaging of a choroid plexus carcinoma and meningeal carcinomatosis in a dog. AB - A 6-year-old neutered female Boston Terrier had a slow onset of blindness and behavior changes. Neurologic abnormalities included bilateral visual loss with absent menace responses and visual tracking, mydriatic pupils, slow pupillary light responses and papilledema. On magnetic resonance imaging (MRI) there were multiple cyst-like structures found in the parenchyma of the cerebrum, cerebellum and brainstem. Histopathologically the diagnosis was a choroid plexus carcinoma with meningeal carcinomatosis. The findings differ from previous descriptions of the MRI characteristics of choroid plexus tumors. PMID- 10519303 TI - Use of magnetic resonance angiography for diagnosis of portosystemic shunts in dogs. AB - A prospective study was conducted to determine the sensitivity and specificity of diagnosis of portosystemic shunts (PSS) and the accuracy of anatomically locating single congenital PSS in dogs using magnetic resonance angiography (MRA). MRA was performed on 10 normal dogs and 23 dogs with PSS. Sensitivity and specificity of MRA to diagnose any shunt among all dogs were 80% and 100%, respectively. Among dogs identified with PSS, sensitivity and specificity of MRA for diagnosis of multiple extrahepatic shunts were 63% and 97%, respectively, and for diagnosis of single congenital shunts were 79% and 100%, respectively. Using MRA, radiologists correctly identified shunts as extrahepatic or intrahepatic in 83% of patients and correctly identified the origin and insertion of the shunts in 57% and 97% of patients, respectively. Use of MRA is specific for diagnosis of PSS and is a sensitive indicator of anatomic location of single congenital portosystemic shunts. PMID- 10519304 TI - Congenital atresia of the parotid duct in a horse. AB - Congenital anomalies of the equine salivary glands and their ductal systems are rare. In man, parotid duct atresia is thought to be due to a congenital malformation of the first branchial arch. One horse with unilateral parotid salivary duct atresia is described. Imaging modalities available for accurate diagnosis, and treatment options, are reviewed. PMID- 10519305 TI - Aortoiliac thrombus secondary to a mineralized arteriosclerotic lesion. AB - A 12-year-old, neutered female, Siberian Husky dog presented with a hind limb weakness of one month duration. To facilitate making a diagnosis multiple imaging modalities were performed. These modalities included radiography, ultrasonography, magnetic resonance imaging (MRI), magnetic resonance angiography (MRA) and selective angiography of the abdominal aorta. In this dog, the MRI/MRA studies provided the first documentation of the external iliac thrombi and the collateral circulation via the lumbar arteries. At necropsy, an aortoiliac thrombus caused by a mineralized arteriosclerotic plaque was noted. PMID- 10519306 TI - Use of magnetic resonance imaging for diagnosis of a spinal tumor in a cat. AB - Magnetic resonance imaging was used to identify a spinal mass at the level of the 6th-7th cervical vertebral body in a cat. The MRI images were most consistent with the presence of an intradural, extramedullary meningioma that was confirmed by subsequent histological examination. PMID- 10519308 TI - Computed tomographic evaluation of Finnhorse cadaver forefeet with radiographically problematic findings on the flexor aspect of the navicular bone. AB - Computed tomography (CT) was performed on 12 Finnhorse cadaver forefeet with known radiographic changes in the navicular bone (poor corticomedullary junction, irregular appearance of the flexor central eminence, uneven or unequal thickness of the flexor cortex, and/or irregular outline of the proximal or distal flexor margin). The purpose was to confirm the radiographic findings and to investigate if further information of the flexor aspect of the bone could be gained with CT. In CT, the midsagittal outline as well as the internal structure of the bones varied greatly. Different combinations of trabecular and compact bone were seen within the flexor central eminence. Lucencies within the compact bone were commonly present in the proximal half of the eminence, but in five bones lucencies were also identified in the distal half. Due to partial overlapping of the bone and varying bony composition of the eminence, accurate radiographic evaluation of the shape and internal structure of the flexor central eminence was often found to be difficult. The flexor cortex usually appeared to be thinner in CT than in conventional radiographs. Medullary sclerosis and poor flexor corticomedullary junction were commonly overinterpreted radiographically. New bone formation on the proximal flexor margin of the navicular bone was generally visualized in radiographs, but CT allowed also the evaluation of the internal structure of the bone. In one navicular bone, an avulsion fragment on the distal flexor margin was seen in CT images; radiographically this fragment could not be visualized. It was concluded that the flexor aspect of the navicular bone may be difficult to assess reliably with conventional radiography. PMID- 10519307 TI - Endometriosis and a paraovarian cyst in a rhesus macaque. AB - We describe endometriosis in an aged rhesus macaque. There was a large mass and a related paraovarian cyst, typical of the disease. Endometriosis is a common finding in nonhuman primate. In this report, we also review the pathophysiology of the disease and summarize the historical and more recent relevant literature. Given the frequency of endometriosis in the rhesus monkey and the long-life spans (15-30 years) of nonhuman primates in captivity, endometriosis should be suspected in animals displaying the earliest signs of the disease: anorexia, dysmenorrhea, menorrhagia, irregular menstrual cycles, or infertility. Despite recent advances in the diagnosis and therapeutic strategies for endometriosis in women, the disease remains a significant cause of morbidity and ultimately, a cause of mortality, in the older nonhuman primate. PMID- 10519309 TI - Sonography of the canine stifle. AB - When diagnosing disease of the stifle in dogs ultrasonography is a good addition to clinical and radiological examination. Radiology can evaluate the bony aspects of the joint and their relationship to each other. In contrast, sonography allows visualization of the soft tissue. For most evaluations the 7.5 MHz linear scanner is suited best. Normal stifles of 58 dogs of different breeds were evaluated using a standardized examination procedure. This procedure had been derived from that used in humans. The stifle is divided into several regions which are examined and evaluated. These are the suprapatellar, infrapatellar, lateral, caudal and medial region. One hundred twenty seven patients which had problems associated with the stifle joint were examined sonographically after a clinical and radiographic exam. Osteochondrosis dissecans, ruptured cranial cruciate ligament, meniscal damage, arthritis, tumor, post surgical conditions, injuries of the patella, patellar ligament or tibial tuberosity and luxating patella were examined sonographically and the findings recorded. PMID- 10519310 TI - Ultrasound-guided tissue-core biopsy of liver, spleen and kidney in normal dogs. AB - Six normal dogs were subjected to ultrasound-guided biopsy of the liver, spleen and kidney to examine the accuracy of the technique (i.e. the presence of targeted tissue) and the histologic quality of the biopsies. Five consecutive tissue-core biopsies of each organ were taken on one or more occasions. The accuracy of the technique was 77% for hepatic, 90% for splenic, 53.5% for left kidney and 40% for right kidney biopsies. The histologic quality of the liver and kidney samples was sufficient, although for some samples the diagnostic value was limited by their size and in renal samples either cortical or medullary tissue was sometimes lacking. In contrast, the quality of the splenic sections was not good. The effect of reused and resterilized needles on the quality of the specimens was evaluated by histologic inspection of the samples and by the amount of biopsies lacking tissue. All tissue samples, including those taken with reused or resterilized needles had sharp-cut edges. Twenty-two of the total number of 120 biopsies (18%) contained no tissue. Absence of tissue in the samples was observed in biopsies taken with all needle types. The animals were observed for possible complications of the repeated needle biopsy. Apart from one case of hematuria, no complications were encountered. PMID- 10519311 TI - Ultrasonographic evaluation of biliary cystadenomas in cats. AB - The purpose of this study was to describe the ultrasonographic appearance of biliary cystadenomas in cats and compare the findings to a similar rare form of liver tumor in humans. Biliary cystadenomas are uncommon, benign liver tumors of older cats that may occur as focal or multifocal cystic lesions within the liver. The records of 10 cats which had abdominal ultrasonography and histologic diagnosis of biliary cystadenoma were reviewed. The average age of affected cats was 13.3 years (range 10-16 years). Eight cats were neutered males and two were neutered females. In three cats, the tumors were not seen ultrasonographically due to their small size or from being obscured by near-field reverberation echoes. The remaining seven cats had solitary (4 cats) or multifocal (3 cats) masses corresponding to variable ultrasonographic patterns: multilocular masses containing thin-walled cysts, hyperechoic masses with cystic components, or masses of mixed echogenicity with cystic components. The masses had variable ultrasonographic patterns when multifocal disease was present. Recognizable cysts were evident somewhere within the tumors seen ultrasonographically, although sometimes the cysts appeared very small. The biliary cystadenomas were thought to be clinically silent. Although liver enlargement or a cranial abdominal mass was palpable in 4 cats, no consistent trend of clinical signs, CBC or serum biochemical abnormalities could be directly attributed to biliary cystadenoma. The treatment of choice is surgical resection of the tumor, as continued growth may compress adjacent vital structures within the liver. The differential diagnosis of biliary cystadenomas from other cystic liver lesions such as hepatic cysts, hematomas, abscesses, parasitic cysts, or other liver tumors is discussed. PMID- 10519312 TI - Ultrasonography and fine needle aspirate cytology of the mesenteric lymph node in normal domestic ferrets (Mustela putorius furo). AB - The large mesenteric lymph node of 28 normal ferrets was imaged with ultrasound. The large node, located in the mid-abdomen at the root of the mesentery, was round to ovoid and uniformly hyperechoic. Mean ultrasonographic dimensions of the lymph node were 12.6 +/- 2.6 mm by 7.6 +/- 2.0 mm. Fine needle aspirates of 20 lymph nodes were obtained either using ultrasound guided free-hand techniques or at necropsy. The cytological descriptions were compared to histological descriptions of 13 lymph node core biopsies obtained during laparotomy or necropsy as well as 10 peripheral blood smear differentials. The large mesenteric lymph node of ferrets could be easily imaged and measured by ultrasound and evaluated by fine needle aspirate cytology. Normal lymph node cytology may include an eosinophilic infiltrate. PMID- 10519313 TI - Factors influencing survival after radiotherapy of nasal tumors in 130 dogs. AB - Improvements in survival of dogs with nasal tumors have been slow to develop throughout the past three decades. Despite multiple studies examining various radiation time-dose schema, the advancement of CT-based computerized treatment planning, and the evaluation of detailed staging systems, the optimal treatment regimen, and most important prognostic factors regarding survival remain unclear. In this study, data from four previous studies were combined with data from 44 additional dogs, and this population of 130 dogs was evaluated for factors which influenced survival. Twenty-one dogs were treated with orthovoltage at the University of Pennsylvania. One hundred nine dogs were treated with cobalt photons at North Carolina State University. Sixty-five of these 109 dogs had been previously described. Of the 44 dogs not previously described, 35 were treated with a shrinking field technique. Survival was determined from the medical record, or from information derived by telephone or mail survey. The univariate Cox regression model was used to examine for relationship between various patient, tumor, and treatment variables and survival. Significant relationships identified in the univariate analysis were further analyzed using the multivariate Cox regression model. Median survival of the 130 dogs was 8.9 months (95% C.I., 8-11 months). In the univariate analysis, the following variables were associated with decreased survival: 1) age >10 years old, 2) regional lymph node metastasis, 3) advanced tumor stage, 4) use of megavoltage radiation, 5) overall total dose >55 Gray, and 6) boost technique performed. In a multivariate analysis of 125 dogs with complete data for age, radiation type, and radiation dose, age (p < .001) and radiation type (p = .02) were identified as joint predictors of survival. After adjusting for age, the staging system lost prognostic significance (p = .06). In a subset of dogs that received cobalt radiation, after adjusting for age, dogs treated with a boost technique had decreased survival (p = .001). In general, local control of canine nasal tumors following aggressive radiation therapy is poor. Early diagnosis and selection of appropriate patients is warranted and palliative types of treatment should be considered in dogs with a poor chance of long term survival. PMID- 10519314 TI - A review of portal screen-film technology and five radiologists' evaluations of some existing products. AB - Portal radiographs, radiographs made to document the accuracy of radiotherapy treatment fields, are typically of poor image contrast. Recently, a new portal film and screened-cassette system was marketed, the Kodak EC-L system, with the claim of greatly improved image contrast. This new EC-L system was tested on a canine cadaver exposed to Cobalt-60 teletherapy gamma radiation, and image quality was compared to earlier marketed Kodak portal film products. The EC-L system was found to provide portal images of improved contrast/quality. PMID- 10519315 TI - Selection in a T-dependent primary humoral response: new insights from polypeptide models. AB - Regulatory mechanisms involved in the induction and progression of T-dependent humoral responses have been extensively delineated using a variety of haptens as model antigens. However, several unanswered questions remain with respect to those elicited by structurally more complex molecules. Our own laboratory has been pursuing this latter aspect using designed synthetic peptides as model systems. The cumulative results indeed support that humoral responses to such antigens involve several additional layers of regulation, beyond that identified with haptens. At the first level, the multiplicity of antigenic determinants recognized by the preimmune B-cell pool is soon subject to competitive pressures that restrict, both at the level of repertoire and epitope, fine specificities of early activated clonotypes. Selection at this stage is on the basis of affinity for epitope, which, in turn, is under thermodynamic control. This selected B-cell subset proceeds to populate germinal centers, where further optimization--by way of somatic hypermutation followed by clonal selection--is in favor of increased on-rates of antigen binding. Thus, contrary to findings with hapten antigens, maturation of antibody responses to polypeptides occurs in two discrete, but sequential, stages. The first is for B cells with optimum affinity for the corresponding epitope. This is then followed by further improvement on the basis of increased on-rates of antigen/epitope binding. It is a combination of these two processes which results in the high fidelity of antibodies produced in the secondary response. PMID- 10519316 TI - In situ expression of E-selectin and intercellular adhesion molecule-1 in chronic inflammatory diseases of the gastrointestinal tract. AB - AIM: The study of cell adhesion molecules contributes to our understanding of the inflammatory mechanisms which include the endothelial activation of newly formed or pre-existing vessels, the increase of inflammatory cells' adhesive capability and their migration into perivascular tissues. The aim of the present study was to investigate the local presence and the extent of expression of E-selectin and intercellular adhesion molecule-1 (ICAM-1) in the mucosa of patients with chronic gastritis, chronic inflammatory bowel disease, and controls, as well as to identify possible correlations between in situ expression of the above adhesion molecules and degree of inflammatory activity or therapeutic response. DESIGN: In cryostat tissue sections we examined the immunohistochemical expression and localization of E-selectin and the intercellular adhesion molecule-1 (ICAM-1). Our specimens consisted of 27 cases of chronic gastritis, 42 cases of ulcerative colitis, and 15 cases of Crohn's disease. RESULTS: E-selectin was expressed in capillary endothelia as well as on neutrophils, located either in the lamina propria or in the glandular epithelia or lumina. This marker's expression was associated with the active phase of ulcerative colitis (p<0.0005) and possibly of chronic gastritis (p=0.06). ICAM-1 immunolabelling was localized in endothelia and chronic inflammatory components which had passed through the vascular walls. This marker's immunoreactivity was generally increased in all our specimens compared to normal mucosa and generally tended to correlate with chronic phases of the inflammatory process (p<0.10). CONCLUSIONS: E-selectin regulates the accumulation of neutrophils in the early stages of the inflammatory process and is thus associated at least with the active phase of ulcerative colitis. Whether any post-therapy alteration of E-selectin immunopositivity seems to indicate a good response to drug therapy is well worth investigating in ulcerative colitis patients. ICAM-1 immunoreactivity in lymphoplasmacytic infiltrates might serve as a marker of chronic immune stimulation, which is potentially responsible for the persistence of the inflammatory disorders. PMID- 10519317 TI - Chlamydia pneumoniae may be associated with lung cancer. Preliminary report on a seroepidemiological study. AB - Material from 117 consecutive patients with lung cancer was investigated with respect to serological markers for chronic Chlamydia pneumoniae infection. Specific C. pneumoniae IgA antibodies were found significantly more often in patients with lung cancer than in control groups with coronary heart disease and in healthy controls, even after adjustment for smoking. The results suggest that chronic C. pneumoniae infection is common in patients with lung cancer. PMID- 10519318 TI - Fungus-like hyphochytrids associated with human disease. AB - We report two cases, with liver and brain abscess, respectively, where fungus like organisms belonging to the Hyphochytriomycota were found at the site of inflammation together with Peptococcus in the first and Cysticercus cellulosae in the second case. This is the first time these groups of organisms have been reported in human material. The role of hyphochytrids in human pathology remained uncertain as they were found together with already known human pathogens. PMID- 10519319 TI - Radiation-induced inflammatory malignant fibrous histiocytoma of the ileum. AB - A case of inflammatory malignant fibrous histiocytoma of the ileum seemingly induced by radiation is described. A 50-year-old female with a past history of uterine cervical carcinoma and postoperative radiation therapy presented with abdominal pain, fever and leukocytosis. The subserosa of the distal part of the ileum showed a diffuse dense, neutrophilic and lymphocytic infiltrate with dispersed atypical, short spindle- or plump oval-shaped histiocyte-like cells. Pleomorphic mono- or multinucleated giant cells with bizarre nuclei were also intermingled in the lesion. Immunohistochemically, the tumorous atypical cells were positive for vimentin, alpha-smooth muscle actin, alpha-1 antitrypsin and granulocyte colony-stimulating factor. No EBV genomic sequences were detected by in situ hybridization. Flow cytometry showed an aneuploid DNA content with high S phase fraction. The patient was well with no evidence of tumor at 5 months after surgery. It is important to include this type of tumor in the differential diagnosis of small intestinal lesions accompanied by fever and leukocytosis following radiation. PMID- 10519320 TI - The fluctuating pattern of various genome types of respiratory syncytial virus in Copenhagen and some other locations in Denmark. AB - A semi-nested RT-PCR method based on a region of the F and G glycoprotein genes was established, allowing the simultaneous detection and differentiation of group A and group B isolates of respiratory syncytial virus (RSV). The PCR products were subjected to digestion with restriction endonucleases to further differentiate the isolates. Using, in addition, previously reported studies the prevalence of various genome types in the Copenhagen region over a period of 6 years was established. Furthermore, the prevalence of genome types was determined in a distant region in Denmark during the winters of 1996/97 and 1997/98, and in yet another distant region during the winter of 1997/98. It was shown that the different regions in Denmark to a large extent share the same pool of genome types of RSV. Yet, while the fluctuating patterns of the two groups and various genome types were almost identical at different hospitals in the Copenhagen region, they varied between the different regions. This suggests that epidemics in local communities primarily rely on region-specific herd immunity parameters and emerge from strains endemically circulating in these local communities. Group B strains in Copenhagen showed an overall predominance, being predominant in three of the six epidemic seasons studied, and of almost equal predominance in one season. PMID- 10519321 TI - Multidrug resistance and retroviral transduction potential in human small cell lung cancer cell lines. AB - Multidrug resistance (MDR) remains a major problem in the successful treatment of small cell lung cancer (SCLC). New treatment strategies are needed, such as gene therapy specifically targeting the MDR cells in the tumor. Retroviral LacZ gene containing vectors that were either pseudotyped for the gibbon ape leukemia virus (GALV-1) receptor or had specificity for the amphotropic murine leukemia virus (MLV-A) receptor were used for transduction of five SCLC cell lines differing by a range of MDR mechanisms. Transduction efficiencies in these cell lines were compared by calculating the percentage of blue colonies after X-Gal staining of the cells grown in soft agar. All examined SCLC cell lines were transducible with either vector. Transduction efficiencies varied from 5.7% to 33.5% independent of the presence of MDR. These results indicate that MDR does not severely impair transduction of SCLC cells, and that MLV-A as well as GALV-1 retroviral vectors are suitable for further development of gene therapy in SCLC. PMID- 10519322 TI - The use of CHROMagar Orientation as a primary isolation medium with presumptive identification for the routine screening of urine specimens. AB - The aim of the study was to compare the use of a novel differential culture medium CHROMagar, for both primary isolation and presumptive identificaton, with the method currently used in our laboratory for screening mid-stream-urine samples (MSU). Routine methods (RM) included blotting paper imprinting of all specimens and additional quantitative culture on cysteine lactose electrolyte deficient agar (CLED) for selected samples together with Microbact 12E for further identification. The CHROMagar method (CH) relied on the use of blotting paper imprints, colonial colour and morphology on CHROMagar only. With respect to the 3390 MSU specimens examined, both methods yielded similar results in 3240, including > or = 87% of Escherichia coli, Pseudomonas spp., Staphylococcus spp., Proteus mirabilis/Morganella morganii and Enterobacter/Serratia/Klebsiella/Citrobacter spp. Of the 52 discordant identifications, yeasts were reported as staphylococci on CHROMagar in 10. The overall cost of materials per specimen was US$ 0.30 by RM and $ 0.24 by CH. It took about 3 min to perform each Microbact test. Thus, CHROMagar plus Gram stain and other simple bench tests gave results similar to those using our current method, but had the advantage of saving time and materials. PMID- 10519323 TI - A histomorphometric method for evaluating topographic differences in degree of early atherosclerosis within the human aorta. AB - The aim of the study was to develop an unbiased topographically oriented method of evaluating early atherosclerotic lesions in the aorta and to apply this to a series of human aortas from young adults. A systematic sampling procedure and histomorphometric analysis of intimal thickening is described. Results from a group of 15 young adults (aged 18-40 years) showed a characteristic pattern with increasing intimal thickening when moving distally from the thoracic to the upper and finally to the lower abdominal aorta, but also a shift in the localization of the most pronounced intimal thickening from the posterior to the anterior and back to the posterior aspect. This pattern was found in aortas both with minimal and with more pronounced atherosclerosis, and supports the view that the early intimal thickening precedes the atherosclerotic lesions and marks the sites of predilection for the more advanced disease processes. An increase in intimal thickness with age could be demonstrated in the aortas without overt atherosclerosis. The simple sampling procedure, well-defined sampling sites, and ability to demonstrate and quantitate differences in intimal thickening and plaque morphology make this method well suited for relating morphometric data to other parameters of interest when studying the etiology and dynamics of atherosclerotic disease. PMID- 10519324 TI - Properties and distribution of the putative R3 protein of Streptococcus agalactiae. AB - Strain-variable Streptococcus agalactiae (group B streptococci; GBS) proteins exposed at the bacterial cell surface are important markers in GBS serotyping. These proteins include the c proteins c(alpha) and c(beta) and the R proteins R1 through R4, of which R1 and R4 have been studied most extensively. This study presents the characteristics of a protein which was expressed by a capsular antigen type V GBS strain shown by means of polyclonal and monoclonal antibody testing. Examination of a number of reference and prototype strains by fluorescent antibody testing and Western blotting provided evidence that the serotype V-derived protein was the R3 protein of GBS, previously defined on the basis of immunoprecipitation assays. The putative R3 protein formed ladder-like banding patterns on Western blotting with polypeptides in the 30 kDa to > or = 140 kDa range, was destroyed by pepsin digestion, and partially degraded by trypsin digestion. The protein was expressed by 10 (6.5%) of 153 clinical GBS strains tested, the expression being restricted to isolates of the capsular antigen types II, III, and V. Some isolates expressed both the c(beta) and the R3 protein. Expression in combination with c(alpha) or R4 protein synthesis was not detected. Inclusion of the anti-R3 monoclonal antibody among antibody reagents for GBS serotyping will enhance the discriminatory power of this typing method. PMID- 10519325 TI - Expression of in vitro virulence by Entamoeba histolytica: effect of calmodulin inhibitors. AB - The possible role of calmodulin (CaM) in functions such as cytotoxicity, phagocytosis, and amoebic growth was studied. These are important factors for the in vitro pathogenicity of Entamoeba histolytica HM1 strain. The CaM inhibitors trifluoperazine (TFP), N-(6-aminohexyl)-5-chloro-1-naphthalene-sulfonamide (W-7) and N-(6-aminohexyl)-naphthalenesulfonamide (W-5) were used for these purposes. Cytotoxicity was determined as the ability of amoebae to kill and/or lyse nucleated mouse spleen cells. Phagocytosis by amoebae was assessed by calculating the number of endocytosed human red blood cells. Cytotoxicity by E. histolytica showed a decrease dependent upon the concentration of W-7 and TFP, whereas W-5 did not show a significant inhibiting effect. Phagocytosis was roughly sensitive to W-7; TFP and W-5 did not have any significant effect. Amoebic growth was sensitive to TFP and W-7 CaM inhibitors. These results suggest that CaM participates in the expression of in vitro cytotoxicity of E. histolytica. PMID- 10519326 TI - Demonstration of Chlamydia pneumoniae in tissue by immunohistochemistry. AB - Chronic Chlamydia pneumoniae infection may be difficult to diagnose using routine methods such as culture, antigen detection, serology and polymerase chain reaction (PCR). We describe a method for the immunohistochemical staining of tissue biopsies using a C. pneumoniae-specific monoclonal antibody and a streptavidin/biotin technique. This method provides specific identification of the organism and reveals its actual location in tissues which may or may not be inflamed. Host cells containing C. pneumoniae antigen were easily identified by their bright red color in tissue sections counterstained with hematoxylin. PMID- 10519327 TI - Gender and disease. PMID- 10519328 TI - Do adolescents' own intentions regarding healthy behaviours affect outcome?--a 2 year prospective study. PMID- 10519329 TI - Adverse drug reactions and the package insert. PMID- 10519330 TI - Growth hormone treatment during suppression of early puberty in adopted girls. Swedish Growth Hormone Advisory Group. AB - Girls adopted from developing countries often have early or precocious puberty, requiring treatment with gonadotropin-releasing hormone (GnRH) analogues. During such treatment decreased growth velocity is frequent. The aim of this investigation was to study whether the addition of growth hormone (GH) to GnRH analogue treatment improves height velocity and final height in girls with early or precocious puberty. Forty-six girls with early or precocious puberty adopted from developing countries were randomized for treatment with GnRH analogue or a combination of GH and GnRH analogue. After 2 y of treatment the mean growth in the GH/GnRH analogue group was significantly higher, 14.6 cm, compared to 10.9 cm in the control group. The increase in bone age did not differ, while the difference in predicted adult height increased by 2.7 cm in favour of the combination group. Although data on final height are not yet available, combined GH/GnRH analogue treatment for 2 y resulted in a higher growth velocity and predicted final height compared to GnRH analogue treatment alone. PMID- 10519332 TI - Thyrotoxicosis in children: thirty years' experience. AB - Optimal treatment for thyrotoxicosis remains controversial in adults, but more so in paediatric practice. We have conducted a retrospective review of the records of 76 paediatric patients seen between 1965 and 1995 to determine management practice and outcome of therapeutic interventions. Seventeen are currently on antithyroid drug (ATD) treatment, while four have had their care transferred. Of the remaining 55, 21 (38%) achieved long-term remission with ATD alone following a mean treatment duration of 3.3 y (range 0.5-7 y). Block-replacement (high dose of ATD with thyroxine replacement) was more convenient than the titration regimen (3.4+/-0.3 visits to hospital per year versus 6.1+/-0.4, p<0.001). Surgery (subtotal/total thyroidectomy) was carried out in 27 patients, of whom 24 subsequently became hypothyroid and were treated with thyroxine. I131 was used successfully in six patients, two following surgery. ATD should remain the first line therapy; a block-replacement regimen is more convenient. Surgery in a specialized centre carries a low risk. Caution should still be exercised in the use of I131 in young children. PMID- 10519331 TI - Prevalence of coeliac disease in Turner syndrome. AB - This study was undertaken to investigate the prevalence of coeliac disease in children and adolescents with Turner syndrome. Eighty-seven children and adolescents with Turner syndrome were screened for IgA-antiendomysium antibodies (EMA) and IgA-antigliadin antibodies (AGA), 5% (4/87) being found to be EMA positive, and 15% (13/87) to have AGA levels above normal. Of the 10 patients who were either AGA- or EMA-positive and further investigated with intestinal biopsy, four manifested villous atrophy (i.e. all three of the EMA-positive patients, but only one of the seven AGA-positive patients). The results suggest EMA-positivity to be a good immunological marker for use in screening for coeliac disease, and such screening to be justified in patients with Turner syndrome. PMID- 10519333 TI - Abnormal surfactant composition and activity in severe bronchiolitis. AB - A prospective study of infants under 1 y of age, ventilated for severe viral bronchiolitis, was carried out in four paediatric intensive care units in order to study surfactant activity and composition in this condition. Lung lavage fluid from 24 infants with bronchiolitis, 19 with bronchiolitis and sepsis or cardiac failure and 12 controls were analysed by the "click test" for surfactant activity and for phospholipids. Surfactant activity was present in all controls, but in only 2 of the 24 infants with bronchiolitis alone. The presence of phosphatidylglycerol correlated perfectly with the click test, suggesting that reduced activity is due to changes in surfactant lipid composition. In those with bronchiolitis plus coexisting disease, surfactant activity and phosphatidylglycerol were absent in only half. Surfactant activity and phosphatidylglycerol re-appeared by extubation. Severe viral bronchiolitis is associated with an absence of surfactant activity and PG, which resolves by clinical recovery. Infants with coexisting conditions are not always surfactant deficient. Surfactant administration is likely to be beneficial, but requires a selective approach. PMID- 10519334 TI - Vascular rings: a rare cause of common respiratory symptoms. AB - Upper airway symptoms or dysphagia may be caused by vascular anomalies, forming a ring around the trachea, oesophagus or both. To analyse the clinical presentation, use of various diagnostic techniques, treatment and follow-up we carried out a retrospective study of 38 children who had been diagnosed with a vascular ring between 1981 and 1996. We found 74% of the vascular rings to be symptomatic, with inspiratory stridor and wheezing as the main complaints. The delay between the onset of symptoms and diagnosis of a vascular ring in patients without associated anomalies ranged from 1 to 84 mo. Associated anomalies were found in 53% of cases and 80% of these anomalies consisted of associated cardiovascular malformations. Oesophagography proved to be a valuable diagnostic technique when a vascular ring was suspected. Echocardiography appeared to be of little value for the diagnosis of a vascular ring, but was essential to exclude associated cardiovascular malformations. Although angiography has always been considered to be the gold standard in the determination of the exact anatomy of vascular rings, increasing evidence is available that CT scan or MRI may replace this role. Mortality was related to co-existent tracheal deformities in 5/6 cases. Of the remaining, preoperatively symptomatic patients, relief of symptoms was achieved immediately after surgery in 43% and within 4 y after surgery in 57%. Prolonged and recurrent respiratory complaints or dysphagia in infancy or childhood should alert the paediatrician to the possibility of a vascular ring. PMID- 10519336 TI - Mitochondrial DNA point mutations detected in four cases of sudden infant death syndrome. AB - The aim of this study was to investigate the tRNA(Leu(UUR)) gene and the first part of the ND1 gene in mitochondrial DNA (mtDNA) in cases of sudden infant death syndrome (SIDS). A total of 158 cases of SIDS and 97 controls were included in the study, and the base pairs in the range 3230-3330 were investigated using polymerase chain reaction (PCR) and temporal temperature gradient electrophoresis (TTGE). If a band shift was detected by TTGE, the area investigated and the D loop was sequenced. Three different point mutations (T3290C, T3308C and T3308G) were detected in four of the SIDS cases, while none of the controls were mutated. We also found a high D-loop substitution rate in these four cases. The findings indicate that mtDNA mutations may play a role in some cases of SIDS. PMID- 10519335 TI - A prevalence study of celiac disease in persons with Down syndrome residing in the United States of America. AB - In order to estimate the prevalence of celiac disease in persons with Down syndrome, 105 patients with this chromosomal disorder residing on the East Coast of the United States of America were enrolled in this study. IgA and IgG antigliadin antibodies (AGA) were determined using a fluorescent immunoenzymatic assay, and antiendomysium antibodies (AEA) were measured with immunofluorescence on monkey oesophagus. Of the 105 patients, 5 were positive for AEA, 4 were positive for IgG AGA, and 1 was positive for IgG AGA and AEA. Of the five patients with high titres of AEA, four consented to a jejunal biopsy, which revealed significant villous atrophy. Thus, 4 (possibly 5) patients in this cohort of 105 individuals with Down syndrome have celiac disease. PMID- 10519337 TI - Comparison of lignocaine-prilocaine cream and amethocaine gel for local analgesia before venepuncture in children. AB - The efficacy of lignocaine-prilocaine cream (EMLA) and amethocaine gel (Ametop) in reducing the pain and distress of venepuncture was compared in a single-blind randomized study of 34 children aged 1-14 y. The influences of age, anxiety and past experience were also investigated. Pain was assessed by the researcher using the Observation Scale of Behavioural Distress, and by the parent, doctor and child (if old enough) using a 10-cm, 100-point Visual Analogue Scale (VAS). Doctors also completed a simple rating scale for difficulty of venepuncture. Compared to older children, infants had significantly more anxious parents and were more distressed when being held still. Anticipatory anxiety correlated with higher pain ratings. No differences in the analgesic effect of the two preparations were found. We conclude that whilst EMLA and Ametop are equally effective at reducing the pain of needle puncture, under some circumstances the use of Ametop may be more advantageous. PMID- 10519338 TI - Adverse drug reactions to unlicensed and off-label drugs on paediatric wards: a prospective study. AB - To determine the incidence of adverse drug reactions (ADRs) to unlicensed and off label drugs used in paediatric inpatients, we carried out prospective surveillance on five different paediatric wards in a regional children's hospital for 13 wk. Comparison of the use of each drug with its summary of product characteristics was made to determine whether the drug was used in an unlicensed or off-label manner. The presence of an ADR was determined using previously defined criteria. In total, 4455 courses of drugs were administered to 936 patients in 1046 admissions. In 507 (48%) of the 1046 admissions, patients received one or more unlicensed or off-label drugs. ADRs occurred in 116 (11%) of the 1046 patient admissions. ADRs were associated with 112 (3.9%) of the 2881 licensed drug prescriptions and 95 (6%) of the 1574 unlicensed or off-label drug prescriptions. Use of drugs in an off-label or unlicensed manner to treat children is widespread. ADRs are a significant problem following unlicensed or off-label drug prescriptions. PMID- 10519339 TI - Assessment of stool colour in community management of prolonged jaundice in infancy. AB - Jaundice persisting beyond the first 2 wk of life is often regarded as an indication for investigation to exclude cholestatic liver disease. Most babies with prolonged jaundice have breast milk-related jaundice, which is a benign condition. Cholestatic liver disease is usually accompanied by pale stools and yellow or orange urine. A community programme was established to ascertain the incidence of prolonged jaundice and determine whether abnormal stool and urine colour could be used to assist primary care staff in referral decisions. Data were collected on normal stool and urine colour and used to devise a colour chart and information sheet for parents. Babies with prolonged jaundice were identified and referred for investigation. In all, 3661 babies were recruited into the study, of which 127 were jaundiced at 28 d of age. Of these, 125 were breastfed. The incidence of jaundice in breastfed babies at 28 d was 9.2% (95% CI 7.8% 11.0%) Abnormal liver function tests (LFTs) were common, but no baby had abnormal stool or urine colour and none was found to have liver disease. Jaundiced breastfed babies who are well are unlikely to have serious disease. Elevated LFTs are compatible with a diagnosis of breast milk-related jaundice. Prolonged jaundice in bottle-fed babies, and persistent pallor of stools or yellow/orange urine, are rare and merit immediate referral. Parents and professionals can be advised to report pale stools without generating a large number of unnecessary referrals. Further work is needed to determine whether a colour chart reduces the mean age of referral and treatment of infants with cholestatic liver disease. PMID- 10519340 TI - Speech and language skills in children who required neonatal intensive care: evaluation at 6.5 y of age based on interviews with parents. AB - Speech and language skills at 6.5 y of age were studied in a follow-up of a cohort of children who had required neonatal intensive care (NIC) at Uppsala University Children's Hospital. An interview with the parents indicated that preterm and full-term NIC children were older than control children when they reached certain stages in language development (short sentences, intelligible speech). Absence of babbling was more common in NIC children born at 23-27 wk than in other preterm NIC children, and occurrence of stuttering was more commonly noticed in preterm NIC children born at 23-27 wk than in those born at >32 wk and controls. PMID- 10519341 TI - Do adolescents' own intentions regarding healthy behaviours affect outcome? A two year prospective study. AB - Adolescents' own intentions regarding health behaviours, in addition to their context, are believed to be important for the health habits they chose. This was studied prospectively over a 2-y period. A total of 552 students, 391 aged 13 y and 161 aged 15 y, reported their health and problem behaviours, socioeconomic background and intentions regarding health behaviours through questionnaires in 1991 and in 1993. Outcome dealt with three domains: health habits; acquisition of adult lifestyles; and problem behaviours. The material was analysed for correlations. Significant results were entered into multiple regression stepwise procedures. As expected, already having initiated adult lifestyles or problem behaviours were the most important factors associated with such behaviours 2 y later. Further analyses were then limited to those students who had not started such lifestyles, in order to determine what factors kept them from doing so in a 2-y span. Key predictors for healthy behaviours were adolescents' own decisions not to engage in adult lifestyles or risky behaviours, family processes consistent with support and school satisfaction. Association with peer groups where smoking and drinking were commonplace predicted less optimal behaviours. Gender or socioeconomic factors were not predictive. The results support a comprehensive approach to health promotion during adolescence. PMID- 10519342 TI - Gender differences in children's hospitalization in Catalonia: another inequality? AB - The aim of this study was to describe children's hospitalization and to analyse gender differences by selected diagnostic and procedure groups. Using the Clinical Classification for Health Policy Research Version 2 (CCHPR), 81 888 hospitalizations from the Minimum Basic Data Set of Hospital Discharge (CMBDAH) of 1995 related to children under 15 y of age in Catalonia, Spain, were studied. Hospitalization rates, number of days in hospital, average length of stay (d) in hospital and the standardized hospitalization ratio (SHR) (hospitalization rate in boys/hospitalization rate in girls) were computed. Two independent approaches were taken: (i) gender-specific categories were excluded in order to compare hospitalizations by sex; (ii) a selected group of 17 diagnostic and procedure categories were chosen in order to carry out the specific gender comparison. In both approaches, the selected indicators were compared by age groups, and totals were standardized by age using the direct method. An excess of hospitalization in boys was observed (SHR = 1.18; 95% CI: 1.17-1.19). The number of days spent in hospital and hospitalization rates were higher in infants and were also higher for boys than for girls. The average length of stay in hospital was higher in infants and in girls at all ages. These differences were systematic when stratified by health region, admission and discharge circumstances and hospital category. Except for urinary infection, which was higher in girls (SHR: 0.65; CI 95%: 0.54-0.76), the SHR was higher in boys in all diagnostic and procedure categories analysed (all SHR >1). Specific sex categories, injuries and poisonings cannot on their own explain a higher level of hospitalization among boys. More specific studies are needed to explain the role of the family and the healthcare system in gender inequalities in children's hospitalization. PMID- 10519344 TI - Consequences of pulmonary inflations (sighs) on cerebral haemodynamics in neonates ventilated by high-frequency oscillation. AB - High-frequency oscillation (HFO) is a technique frequently used in neonatal resuscitation, but which has yet to be evaluated. The use of intrathoracic pressures may have an effect on the cerebral circulation of immature neonates. The aim of this study was to examine the variations in cerebral blood velocity and oxygenation during brief pulmonary inflations (sighs), by focusing on alveolar recruitment. In this prospective study performed in 13 intubated and ventilated neonates (alpha = 5%; 1-beta = 80%), mean blood velocity and Doppler Resistance Index were measured, and variations in chromophores concentrations were evaluated by near infrared spectroscopy. Brief inflations at 4 cm H2O above the mean regulated intra-thoracic pressure did not cause any variation in the parameters measured. An explanation for this discordance with animal studies may be the level of pressure chosen, which could be more appropriate for the pulmonary compliance of neonates. PMID- 10519343 TI - Mental health and psychosocial characteristics in adolescent obesity: a population-based case-control study. AB - In this population-based study we compared self-esteem, social background, social and academic competence, behavioural problems and lifestyle in 58 obese adolescents (BMI > or =99.6th percentile or > or =30 kg/m2), aged 14-18 y, with 58 sex- and age-matched controls of normal weight. The instruments used were: I Think I Am, Youth Self Report and a lifestyle questionnaire. The obese group was on average, 40 kg heavier than the controls. The obese individuals rated themselves significantly lower in physical characteristics, but in all other aspects of self-esteem, mental health and social and academic competence there were no differences between the two groups. There were significant socioeconomic differences, with more obese adolescents living with only one parent and with the mothers in the obese group having, in general, lower education than those in the control group. This study confirms previous observations that obesity is associated with special socioeconomic conditions in youth, but that obese adolescents do not differ from their normal-weight peers in other aspects of mental health. PMID- 10519345 TI - Intracardiac fungal masses in high-risk neonates: clinical observations. AB - Four cases of intracardiac fungal masses occurred over 2 y amongst 7 cases of systemic candidiasis in a neonatal referral unit. The gestations and birthweights were 25, 23, 24 and 30 wk and 805, 605, 640 and 1395 g, respectively. The pedunculated, solitary right atrial masses were detected 2-17 d after diagnosing candidemia in 3 cases, whereas it was the presenting feature in the 4th. All had indwelling right atrial catheters and received multiple courses of broad-spectrum antibiotics. The masses were removed successfully in two cases fit for surgery. None survived despite antifungal therapy, including liposomal amphotericin B at 6 mg/kg/d. Early introduction of enteral feeds, minimization of prolonged exposure to broad-spectrum antibiotics and judicious use of central catheters may reduce the incidence of systemic candidiasis in high-risk neonates. Surveillance echocardiography and timely surgical intervention may reduce the mortality and/or morbidity related to intracardiac fungal masses. PMID- 10519346 TI - Cardiac output and ductal reopening during phototherapy in preterm infants. AB - Left ventricular output (LVO), left pulmonary artery blood flow (LPA) and patency of the ductus arteriosus (PDA) were studied with 2D/pulsed Doppler ultrasound before, during and after phototherapy treatment in 27 preterm infants (gestational age < or =32 wk), who were exposed for a minimum of 12 h to phototherapy for non-haemolytic hyperbilirubinemia. In 14 infants (52%) the ductus arteriosus reopened during phototherapy, but ductal patency was not of haemodynamic importance. LVO initially decreased in preterm infants in whom the ductus did not reopen. From 12 h until discontinuation of phototherapy, LVO and LPA were higher than before phototherapy in all infants. After withdrawal of phototherapy, LVO and LPA returned to pre-phototherapy values. PMID- 10519347 TI - Endotracheal tolazoline: pharmacokinetics and pharmacodynamics in dogs. AB - Tolazoline is a potent vasodilator of both arteries and veins and has a powerful effect on the pulmonary vasculature, reducing hypoxic pulmonary vasoconstriction and lowering pulmonary artery pressure. Intravenous tolazoline lowers the mean pulmonary arterial pressure and resistance and increases the cardiac index when given to infants with persistent pulmonary hypertension of the newborn (PPHN). Endotracheally administered tolazoline decreases mean pulmonary arterial pressure and pulmonary vascular resistance, and improves oxygenation without the harmful decline in systemic arterial pressure. The purpose of our study was to examine the pharmacokinetic and pharmacodynamic characteristics of endotracheal tolazoline in order to determine the relationship between endotracheal tolazoline administration, plasma concentration and its effects on the cardiovascular and respiratory systems. Tolazoline was administered endotracheally to 7 newborn dogs, and its serum concentration and the haemodynamic parameters were monitored for 270 min post-delivery. Results are expressed as median and quartiles. It was found that 15 s after dosing, tolazoline plasma concentrations started to increase significantly above baseline levels, reaching a maximum of 2.64 (1.36; 13.16) microg/ml. The extent of tolazoline absorption was 305 (148;453) microg/ min/ml. The volume of distribution was 3.4 (1.6;7.4) 1/kg. The total body clearance was 12.1 (10.9;23.9) ml/min/kg and the elimination half-life was 225 (171;303) min. Endotracheal tolazoline produced an initial short-lived decrease in mean blood pressure in all the dogs, but thereafter the blood pressure increased gradually above baseline levels. Immediately following endotracheal tolazoline significant tachycardia developed, peaking at 90 min. Subsequently, the heart rate gradually decreased and stabilized at values above baseline for 200 min. A single endotracheal dose of tolazoline is effectively absorbed and produces measurable pharmacological effects. Determining the optimal endotracheal dose of tolazoline in the clinical setting requires additional evaluation. PMID- 10519348 TI - Early discharge of preterm infants needing limited special care, followed by domiciliary nursing care. AB - The aim of this study was to evaluate the effect of early discharge, followed by domiciliary nursing care, on infant health and utilization of health services in preterm infants still in need of special care (mainly gavage feeding). In total, 88 infants who were physiologically stable, but in need of further special care such as gavage feeding, were allocated to an early discharge group (EDG = 45 infants) and offered home visits by a nurse backed up by a neonatologist, or to a control group offered standard neonatal care (CG = 43 infants). Infants in the EDG spent 30.6 d (mean) in hospital after birth compared with 46.3 d in the CG (p = 0.003). On average, the domiciliary nurse spent 10.4 h with each family in the EDG, including a median number of 5 home visits, scheduled telephone contact and travelling time. The infants had a mean of 1.7 scheduled visits and 0.4 unscheduled visits to the neonatal ward. The domiciliary nurse received a mean of 0.9 telephone calls from the parents. When the period of domiciliary care in the EDG (post-conceptional age 35.9-38.7 wk) was compared with the corresponding time in hospital in the CG (post-conceptional age 35.6-38.6 wk), no statistical differences were observed in infant health, surgical procedures or medication. However, a reduced incidence of respiratory infections was observed in the EDG (6 versus 16 infants; p = 0.02). Nine infants in the EDG were re-hospitalized. The two groups did not differ in the numbers of rehospitalizations and non-elective contacts with the health services during the first year after discharge. In conclusion, early discharge of preterm infants still requiring special care, followed by domiciliary nursing care, was associated neither with an increased utilization of health services after discharge, nor with infant morbidity after discharge. More information on safety is needed before widespread early discharge can be advocated. PMID- 10519349 TI - Neonatal Group B Streptococcal bacteraemia in India: ten years' experience. AB - Group B Streptococcus (GBS) is an infrequent cause of neonatal septicaemia in many developing countries. In a perinatal centre in India with 60,119 live births between 1988 and 1997, GBS was isolated from blood cultures of 10 babies. Thus the incidence of GBS bacteraemia was 0.17 per 1000 live births. Lethargy, respiratory distress and poor perfusion were the presenting features in eight symptomatic babies. Two babies had meningitis, three required ventilatory support and one died. There were no cases of late onset disease. The low incidence could be due to the low rate of colonisation and high prevalence of protective antibody in the mothers. PMID- 10519350 TI - Prone position in spontaneously breathing infants with pneumonia. AB - This study was designed to evaluate further the effect of prone positioning on oxygen saturation (SpO2) and respiratory mechanics in spontaneously breathing infants with pneumonia. SpO2 and respiratory mechanics were measured in the supine and prone positions in 17 infants. Prone positioning resulted in statistically significant increases in mean (+/- SD) SpO2 (95.52+/-2.87 to 98.00+/-2.40%, p = 0.0002) and respiratory system compliance (5.99+/-2.52 to 7.93+/-4.30 ml/cm H2O, p = 0.02). This suggests that prone positioning is another beneficial supportive measure for spontaneously breathing infants with pneumonia. PMID- 10519351 TI - Non-catheter-related aortic thrombosis and resistance to activated protein C in a premature newborn. PMID- 10519352 TI - Covert biting of the buccal mucosa masquerading as haematemesis or haemoptysis in children. PMID- 10519353 TI - Ring chromosome 17 syndrome with monosomy 17 mosaicism: case report and literature review. PMID- 10519354 TI - Non-invasive oscillometric blood pressure measurement in very-low-birthweight infants: a comparison of two different monitor systems. PMID- 10519355 TI - Recurrent pericarditis in familial Mediterranean fever. PMID- 10519356 TI - Transient hyperphosphatasemia in a renal transplant patient. PMID- 10519357 TI - Behcet's disease, the Silk Road and HLA-B51: historical and geographical perspectives. AB - Behcet's disease (BD), also known as the Silk Road disease, is a blinding inflammatory disorder of young adults found predominantly between the Mediterranean basin and the Orient, and is strongly associated with the major histocompatibility complex (MHC) antigen HLA-B51. In this article we review the history of Behcet's disease since its first description by Hippocrates, the development of the trading routes collectively known as the Silk Road and the effect of population movement on the distribution of HLA-B51. The global distribution of this antigen among healthy control populations bears a striking similarity both to the ancient trading routes and the distribution of Behcet's disease, suggesting a genetic risk that migrated in parallel with population movement between the Mediterranean and Asia. However, certain indigenous Amerindian peoples have a high prevalence of HLA-B51 but no reported cases of BD. Furthermore, a clear genealogical relationship exists between eastern, but not central, Siberian populations with the Amerindians. Since a high level of recombination within the MHC is known to have occurred in these eastern populations before their migration into Beringia, we suggest that disruption of genetic loci in linkage disequilibria with HLA-B51 may be one reason for the absence of disease in these high HLA-B51-bearing populations. However, a contributory influence of environmental factors is not excluded by this data, and the wide variation that exists in relative risk of HLA-B51 even within Europe would support other non-genetic risk factors on the Silk Road which may be absent, or non-contributory to disease, in the Americas. PMID- 10519358 TI - New polymorphic microsatellite markers in the human MHC class I region. AB - The human major histocompatibility complex (MHC) class I region is believed to contain a large number of genes encoding susceptible factors for diseases such as Behcet's disease, Graves disease and psoriasis vulgaris. To identify the causative genes of those diseases, we have conducted large-scale genomic sequencing and determined the 1.8 Mb entire HLA class I region from the MICB gene to the HLA-F gene. During the course of genomic sequencing, a total of 731 microsatellite sequences with dinucleotide to pentanucleotide repeats were found in this region. Previously, we reported that 26 microsatellites between MICB and S on the most centromeric side of the class I region, and between HSR1 and HLA 92/L in the midst of the class I region were highly polymorphic, and served as excellent genetic markers. In this paper, in order to fill the gaps with no known polymorphic microsatellites available in the HLA class I region, 12 new polymorphic microsatellite markers were recruited from the 1.8 Mb region including the remaining class I segments, namely between S and HSR1, and between HLA-92/L and HLA-F The average number of alleles at these new microsatellite loci was 8.2 with a polymorphism content value (PIC) of 0.63. These 38 markers in total almost uniformly interspersed in the HLA class I region will enable us to search precisely for the location of disease susceptible loci within the HLA class I region by association and for linkage analyses. PMID- 10519359 TI - Human platelet alloantigen (HPA)-5a/b mismatch decreases disease-free survival in unrelated bone marrow transplantation. AB - Matching of human platelet alloantigen (HPA) systems 2-6 was retrospectively investigated in 715 unrelated bone marrow transplantations. Of the five HPA systems studied, HPA-5 mismatching was found to have a significant effect on the disease-free survival rate of recipients following transplantation in the HLA-A, B, -C, and -DR allele-matched donor-recipient pairs. The effect of the HPA-5 mismatch was most significant in the recipient group possessing the HLA haplotype A*2402-B*5201, which is a highly frequent haplotype among the Japanese population. However, the probability of development of acute graft-versus-host disease (GVHD) was not increased significantly by the HPA-5 mismatching. These findings suggest that the HPA-5 mismatching decreases the recipient's survival by a mechanism different from that in the case of mismatching of minor antigens found often in transplant recipients developing GVHD. PMID- 10519360 TI - HLA class I expression and chromosomal deletions at 6p and 15q in head and neck squamous cell carcinomas. AB - Loss at the chromosomal region 6p21.3 is a frequent event in head and neck squamous cell carcinomas (HNSCC). Since the human leukocyte antigen (HLA) complex is located at 6p21.3, loss of heterozygosity (LOH) of this region may provide tumour cells with an immune-escape tumour phenotype. In the present study, we have studied the correlation of HLA class I, TAP1 and TAP2 expression and LOH at 6p21.3. HLA class I and TAP1 and TAP2 protein expression was analysed by immunohistochemical procedures. A panel of 41 HNSCC with downregulated HLA class I expression was selected for LOH studies using 5 microsatellite markers located at 6p21.3 (D6S105, D6S265, D6S276, D6S273, D6S291) and 2 markers located at the chromosome 6 centromere (D6S473) and the 6p telomere (D6S277). In addition, LOH of the beta-2-nmicroglobulin (beta2m) gene was studied using 2 microsatellite markers flanking the beta2m gene (D15S126 and D15S153) and was correlated with beta2m and HLA class I expression. In 20/41 (49%) of the HNSCC, allelic loss for at least one locus at 6p21.3 was found. Loss at 15q was found in 4/10 (40%) HNSCC with downregulated beta2m expression and in 12/41 (29%) HNSCC with downregulated HLA class I expression. Our data show that downregulation of HLA class I expression is correlated with loss of chromosomal regions at 6p21.3 in HNSCC. In addition, LOH at 6p21.3 and 15q in 10 paired samples of DNA derived from the primary HNSCC, the lymph node metastases and from peripheral blood lymphocytes (PBLs) was studied. Five (5/10) primary tumours contained the same deletion as the corresponding lymph node metastases. The other cases contained deletions either in the primary tumour (3 cases) or in the lymph node metastases (1 case) or no deletions at all (1 case). PMID- 10519361 TI - DNA typing of HLA class II genes (HLA-DR, -DQ and -DP) in Japanese patients with histiocytic necrotizing lymphadenitis (Kikuchi's disease). AB - The pathogenesis of histiocytic necrotizing lymphadenitis (HNL), which was reported first by Kikuchi et al. and Fujimoto et al. in 1972, is as yet unknown. HNL is frequently reported in Asian countries including Japan, however it is rare in Europe and North America. To elucidate whether the human leukocyte antigen (HLA) alleles and haplotypes are associated with HNL, we performed DNA typing of HLA class II genes (HLA-DR, -DQ, and -DP) in 86 patients with HNL and 525 unrelated healthy Japanese controls with polymerase chain reaction using sequence specific oligonucleotide probes (PCR-SSOP). In this study, we found DPA1*01 and DPB1*0202 allele frequencies in HLA class II genes are significantly higher in HNL patients than in normal controls. It is known that the frequency of DPB1*0202 alleles is extremely low or absent in Caucasians (e.g., French 0.4%, Italian 0.8%) and Negroid (e.g., South African 0%, Hottentot 0%), but relatively frequent in Asians (e.g., Korean 9.9%, Japanese 4.5%). Previous reports have said the incidence of HNL is frequent in Asians but rare in other races. In light of this background, HLA class II genes of HNL and the incidence of HNL in Asian countries, including Japan, might have a positive relationship to DPA1*01 and DPB1*0202 allele. PMID- 10519362 TI - Identification of DRB alleles in rhesus monkeys using polymerase chain reaction sequence-specific primers (PCR-SSP) amplification. AB - Major histocompatibility complex (MHC) class In molecules play a vital role in the regulation of T-cell functions in the mammalian immune system. Two key features characterize the polymorphism of MHC haplotypes in humans and non-human primates: the existence of a large number of alleles, and the high degree of genetic diversity between those alleles. Rhesus monkeys and Chimpanzees have been extensively used as relevant models for human diseases and transplantation We have investigated DRB genes in 19 macaques, members of 3 families, using polymerase chain reaction with sequence-specific primers (PCR-SSP) and denaturing gradient gel electrophoresis (DGGE). After amplification PCR products were purified and subjected direct sequencing. Seven animals (Madison #1) were typed by DDGE also. We report that the DRB haplotypes defined by PCR-SSP exhibit a high degree of concordance with the data obtained by DGGE and direct sequening. Our data show prominent variability in the number of DRB1 alleles ranging from 1-4 per genotype within these families. This analysis demonstrated that most of the amplicons were identical to Mamu-DRB alleles that our PCR primers were to amplify. However, 98-99% similarity was noticed in the case of Mamu-DRB1*0303, Mamu-DRB6*0103 and Mamu-DRB*W201 alleles. The observed mismatches were located in non-polymorphic regions. Thus, family studies in rhesus macaques performed by molecular methods confirmed the multiplicity of Mamu-DRB1 alleles per haplotype and the existence of allelic associations published earlier. In addition, we propose 3 more DRB allele associations (haplotypes): Mamu-DRB1*04-DRB5*03; Mamu DRB1*04-*DRB*W5; Mamu-DRB1*04*W2. The proposed medium-resolution PCR-SSP technique appears to be a highly reproducible and discriminatory typing method for detecting polymorphisms of DRB genes in rhesus monkeys. PMID- 10519363 TI - HLA and tumour necrosis factor (TNF) polymorphisms in ocular Behcet's disease. AB - The role of HLA-B*51 and other major histocompatibility complex (MHC) genes in Behcet's disease (BD) remains unknown. We have performed HLA and tumour necrosis factor (TNF) polymorphism analysis in BD and evaluated their contribution to ocular disease. In this study, 102 patients and 115 controls of Middle Eastern descent were investigated by HLA and B*51 subtyping using novel primers, and by LT alpha NCo 1 and TNF 308 promoter polymorphism analysis. The frequency of the HLA-B*51 family of alleles was raised in patients compared to controls (66% vs. 15%, Pc=2.5x10(-12), OR=10.9). The odds ratio (OR) of this group of alleles for subgroups of patients was as follows: non-ocular patients 7.8, all ocular patients 12.6, blind patients >22. HLA-B*51 subtyping detected B*5101, 07, 08 and 09 alleles, with a similar frequency among patients and controls. HLA-Cw*1602 was associated with B*5108, but was not an independent risk factor for disease. The LT alpha (TNFB*2) allele was associated with HLA-B*51 among patients and the frequency of this allele was significantly higher among completely blind patients compared to both non-ocular patients (P=0.048, OR >3.6) and to healthy controls (P=0.022, OR >4.3). The rare TNF-2 polymorphism at the TNF -308 promoter position was associated with HLA-B*50 (not B*51), and was not associated with BD. Thus, in this population the HLA*B51 family of alleles is a strong risk factor for BD, and in particular the development of ocular disease. HLA-B*51 subtyping did not define new markers for BD. A primary role for TNF gne polymorphisms in BD was not identified, but co-expression of the TNFB*2 allele with HLA-B*51 may contribute to severity of ocular disease. PMID- 10519364 TI - HLA class I and II typing of the patients with Behcet's disease in Saudi Arabia. AB - Thirteen Saudi Arabian patients with Behcet's disease (BD) were typed for HLA-A and -B alleles by the conventional serologic typing and for HLA-DRB1, -DQB1 and DPB1 alleles by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. As a result, the phenotype frequency of the B51 antigen was significantly increased in the patient group as compared to the ethnically matched control group (76.9% in patients vs. 22.20% in controls), but no significant difference was observed in HLA-A, -DRB1, -DQB1 or -DPB1 alleles between the patients and controls, as previously observed in Japanese BD. Further, by HLA-B51 allelic genotyping performed by the polymerase chain reaction sequence specific primers (PCR-SSP) method, all of the B51-positive patients and controls were found to carry one particular allele, B*5101, except one patient with B*5108. PMID- 10519365 TI - Lack of association of MICA transmembrane region polymorphism and Behcet's disease in Spain. AB - We have analyzed the distribution of the major histocompatibility complex (MHC) class I chain-related gene A (MICA) transmembrane alleles among 58 Spanish patients with Behcet's disease (BD) and in 194 ethnically matched healthy controls. The study included the characterization of A4, A5, A5.1, A6 "new" and "old" and A9 MICA-TM alleles using polymerase chain reaction. As previously reported, the serological B51 specificity was increased among this BD patient group (36.25% vs. 19.6% in controls; P=0.009; OR=2.33). The MICA-TM alleles A6 ("new" and "old"), in linkage disequilibrium with HLA-B51 and HLA-B14 respectively, were only slightly increased among patients (70.7% vs. 61.3% in controls; P=NS). We conclude that, in contrast to previous finding reporting a strong association of MICA-TM genes and Behcet disease in Japanese patients, in our population HLA-B51 is more closely associated to Behcet susceptibility than MICA-TM genes. Finally, our data show that in Spain, as occurs in other populations, some MICA-TM alleles exhibit strong linkage disequilibrium with certain alleles of the HLA-B locus. PMID- 10519366 TI - HFE genotypes and haemochromatosis: quantifying the risks of disease. AB - Hereditary haemochromatosis (HH) is an autosomal recessive disease involving mutations in the recently characterised HFE gene linked to HLA-A in the major histocompatibility complex. The known HFE polymorphisms include the wild-type allele, a G-->A substitution at base 845 (845A) and a C-->G substitution at position 187 (187G). Although most cases of HH are accountable by homozygosity of the 845A allele the exact risk of other HFE genotypes, especially those involving the 187G allele has not been determined. We have compiled estimates of disease risk for all known HFE genotypes by re-analyzing published studies. The data show a hierarchical risk calculated as odds ratio (OR) for each genotype 845A/ 845A (OR=2101); 845A/187G (OR=24); 187G/187G (OR=9); 845A/Wt (OR=5); 187G/Wt (OR=2). Interestingly, the disease risk of 187G-genotypes suggests that subtle functional changes in the HFE product can interact with other genetic factors (e.g. trans allele, gender) and environmental factors (e.g. diet) to manifest either as clinical disease, altered iron stores or a normal phenotype. This paradigm is potentially useful in understanding the contribution of HLA alleles to risk of various disorders especially autoimmunity. PMID- 10519367 TI - Platelet glycoprotein (GP) V polymorphisms in Japanese. AB - The glycoprotein (GP) V is a subunit of a platelet receptor for von Willebrand factor, the GPIb/IX/V complex. Recently, polymorphisms in the GPV gene, four involving amino acid changes and five silent, were identified. We developed an allele-specific restriction method to determine the gene frequencies of the non synonymous substitutions in Japanese and Caucasians. The gene frequencies for 341Gly/Arg polymorphism were 0.94 for 341Gly and 0.06 for 341Arg, and 0.95 for 341Gly and 0.05 for 341Arg, in Japanese and Caucasians, respectively. We could not find rare variants for 114Asp/Tyr, 273Met/Ile or 397Leu/Arg. Flow cytometric analysis showed that the 341Arg variant is expressed normally on the platelet membranes, suggesting the possibility of the involvement of the 341Gly/Arg polymorphism as a platelet alloantigen. PMID- 10519368 TI - Polymorphism in the tumor necrosis factor B gene is associated with Palmoplantar pustulosis. AB - We investigated the allele and genotype distribution of a polymorphism of the tumor necrosis factor (TNF) B gene and the frequency of HLA-DR9 in 49 patients with Palmoplantar pustulosis (PPP) and 51 healthy controls. We found that the frequency of TNFB2 in the PPP patients was significantly higher than that in the controls. Furthermore, the DR9-TNFB2 haplotype was significantly more frequent in the PPP patients (P=0.0045). These results suggest that TNFB2 may confer susceptibility to PPP. PMID- 10519369 TI - HLA-DMA and HLA-DMB alleles in German patients with type 1 diabetes mellitus. AB - The HLA-DMA and HLA-DMB genes are located in the HLA-D region between DQ and DP. Four variants of DMA (DMA*0101-0104) and five of the DMB (DMB*0101-0105) have so far been identified. HLA-DM molecules are required in the process of peptide loading to HLA class II antigens, both regulating the dissociation of class II associated invariant chain peptides (CLIP) and the subsequent binding of exogenous peptides to HLA class II molecules. In order to investigate the immunogenetic heterogeneity within the HLA-D susceptibility region, we analysed the distribution of DMA alleles in 125 patients with type 1 diabetes mellitus and 90 healthy controls, and of DMB alleles in 102 patients and 89 healthy controls. Patients and controls were all from central Germany. The polymerase chain reaction (PCR) amplified products were purified and separated on a 10% polyacrylamide gel electrophoresis. Among the four recognized DMA alleles, DMA*0102 was significantly less frequent (12% vs. 28.9%, P<0.01) in patients with type 1 diabetes mellitus. DMB*0101 (70.6% vs. 97.8%, P<5.5x10(-3)) was also reduced in frequency compared to controls. Comparing patients and controls positive for the type 1 diabetes high-risk markers we found a significant association between DMA*0102 and DQA*0501 (9.5% vs. 39.1%, P<0.02), as well as DMB*0101 and DQA*0501 (62.5% vs. 96.2%, P<0.03). In conclusion, DMA*0102 and DMB*0101 contribute to genetic protection to type 1 diabetes mellitus in individuals with high-risk DQA markers in the German population. PMID- 10519370 TI - The relative frequencies of HLA-A*10 alleles in five major United States ethnic populations. AB - The frequency of each A*10 allele was determined in 5 major United States ethnic populations randomly selected from a pool containing 82,979 unrelated individuals. The phenotype frequency of A10 was 10.5% in Caucasians, 14.0% in African-Americans, 21.1% in Asians/Pacific Islanders, 10.6% in Hispanics, and 9.8% in Native Americans. Fifty-nine individuals who had at least one A10 antigen were randomly chosen from each ethnic group for polymerase chain reaction using sequence-specific oligonucleotide probes (PCR-SSOP) typing. Thirteen of sixteen known A10 alleles were identified in this pool. The most common alleles observed were: A*2601 in Caucasians (55%), Hispanics (58%), and Native Americans (45%); A*3402 in African-Americans (34%); and A*3401 in Asians/Pacific Islanders (61%). The African-American and Asian/Pacific Islander populations differ from all other populations in the distribution of A*10 alleles, particularly, A*2601, A*3401, and A*3402. PMID- 10519371 TI - A nucleotide deletion in exon 4 is responsible for an HLA-A null allele (A*0105N). AB - We report herein the identification of a new HLA-A null allele. This allele, A*0105N, was detected during histocompatibility testing of a cord blood donor and the respective mother. Serologic typing results contrasted those obtained with DNA typing that alone showed the presence of HLA-A*01. ThisA*0105N was due to a nucleotide deletion in exon 4 that altered the reading frame, causing a premature termination. PMID- 10519372 TI - HLA-Cw*1701 is associated with two sub-Saharan African-derived HLA haplotypes: HLA-B*4201, DRB1*03 and HLA-B*4202 without DRB1*03. AB - Different extended haplotypes have been described for many ethnic groups, such as African-Americans. The complotype FC(1,90)0 is in linkage disequilibrium with HLA B42, DRB1*0302 in African-Americans and Southern African Xhosa individuals, suggesting a common ancestry. In order to analyze the distribution of Cw*17 alleles (Cw*1701, 1702) in relation to this African-derived extended haplotype, we studied a large panel of samples from African-American individuals and additionally a group of selected samples carrying HLA-B42, DR3 and HLA-B42, non DR3 antigens. HLA alleles were assigned using sequence-specific amplification (SSP) and sequence-specific oligonucleotide probe hybridization (SSOP). We have found that all haplotypes (10 in total) carrying the extended haplotypes [HLA B42, FC(1,90)0, DRB1*0302] were positive for HLA-Cw*1701. Interestingly, HLA B*4201 was found in all samples (17 in total) carrying HLA-B42, DR3, Cw*1701, whereas HLA-B*4202 was found in 10 out of 13 samples from individuals carrying HLA B42, Cw*1701 non-DR3. These findings suggest that HLA-Cw*17 polymorphism is conserved in different ethnic populations and that HLA-B42 alleles seem to separate at least different African-derived haplotypes. The historical context of these findings are important for the study of human evolution and they may be useful for the development of strategies in the search for possible donors in organ transplantation for African-derived populations. PMID- 10519373 TI - Unusual association of the DRB4 null allele, DRB4*0103102N, with HLA DRB1*0402 in a sample of Austrian patients. AB - Tissue typing for HLA class II antigens is routinely performed by serological, and/or DNA-based methods. Accuracy, absence of cross-reactivity and controllable level of resolution are striking advantages of molecular methods. However, a disadvantage of molecular typing, compared to serological methods, is the identification of unexpressed alleles. Whereas serology allows a more or less direct insight into antigen presence, molecular biology is an indirect method and results must also be interpreted by considering the biological pathways of protein expression. We believe that identification of nonexpressed MHC alleles is of importance for transplantation, since nonexpressed MHC allele positive individuals could give rise to antibody formation against the respective expressed MHC product in donor tissue. Usually, the nonexpressed DRB4*0103102N is encountered in association with DRB1*0701-DQB1*03032, which facilitates correct DNA typing. Here we describe the unusual association of this unexpressed DRB4*0103102N, with DRB1*0402-DQB1*0302 in a sample of Austrian patients. PMID- 10519374 TI - Nucleotide sequence of a new HLA-DQB1 allele, DQB1*03033. AB - HLA-genotyping by sequencing of the corresponding polymerase chain reaction (PCR) product allow the identification of a new HLA-DQB1 allele, DQB1*03033. To confirm the finding the entire exon 2 was sequenced. PMID- 10519375 TI - Nomenclature for factors of the dog major histocompatibility system (DLA), 1998. First report of the ISAG DLA Nomenclature Committee. International Society for Animals Genetics. AB - A Nomenclature Committee for factors of the dog major histocompatibility system or dog leukocyte antigen (DLA) has been convened under the auspices of the International Society for Animal Genetics (ISAG) to define a sequence-based nomenclature for the genes of the DLA system. The remit of this committee includes: i) assignment of gene names; ii) rules for naming alleles; iii) assignment of names to published alleles; iv) assignment of names to new alleles; and v) rules for acceptance of new alleles. PMID- 10519376 TI - Nomenclature for factors of the HLA system, update June 1999. WHO Nomenclature Committee for Factors of the HLA System. PMID- 10519377 TI - Prevention of cancer in the next millennium: Report of the Chemoprevention Working Group to the American Association for Cancer Research. PMID- 10519378 TI - Application of complementary DNA microarray technology to carcinogen identification, toxicology, and drug safety evaluation. AB - One major challenge facing today's cancer researchers and toxicologists is the development of new approaches for the identification of carcinogens and other environmental hazards. Here, we describe the potential impact of emerging technologies for measuring gene expression profiles on carcinogen identification and on the general field of toxicology. An example of one of these technologies is the use of cDNA microarray chips. We provide an overview to the key questions that are confronting investigators charged with determining the relative safety of natural or synthetic chemicals to which humans are exposed, followed by a discussion of how cDNA microarray technology may be applied to these questions. Gene chip technology is still a relatively new technology, and only a handful of studies have demonstrated its utility. However, as the technical hurdles to development are passed, the use of this methodology in addressing the questions raised here will be critical to increase the sensitivity of detection of the potential toxic effects of environmental chemicals and to understand their risks to humans. PMID- 10519379 TI - Response of prostate cancer to anti-Her-2/neu antibody in androgen-dependent and independent human xenograft models. AB - Antibody to the Her-2/neu gene product has been shown to inhibit the growth of breast cancer cells overexpressing Her-2/neu and to have clinical utility in treating breast cancer. We studied a recombinant, humanized anti-Her-2/neu antibody (Herceptin) in preclinical models of human prostate cancer. The androgen dependent CWR22 and LNCaP human prostate cancer xenograft models and androgen independent sublines of CWR22 were used. Her-2/neu staining of the parental, androgen-dependent, and androgen-independent CWR22 tumors and LNCaP tumors demonstrated variable Her-2/neu expression. Herceptin was administered i.p. at a dose of 20 mg/kg twice weekly after the xenograft had been established. No effect of Herceptin on tumor growth was observed in any of the androgen-independent tumors; however, significant growth inhibition was observed in both of the androgen-dependent xenograft models, CWR22 (68% growth inhibition at the completion of the experiment; P = 0.03 for trajectories of the average tumor volume of the groups) and LNCaP (89% growth inhibition; P = 0.002). There was a significant increase in prostate-specific antigen (PSA) index (ng PSA/ml serum/mm3 tumor) in Herceptin-treated androgen-dependent groups compared with control (CWR22, 18-fold relative to pretreatment value versus 1.0-fold, P = 0.0001; LNCaP, 2.35-fold relative to pretreatment value versus 0.6-fold, P = 0.001). When paclitaxel (6.25 mg/kg s.c., five times/week) was given to animals with androgen-dependent and -independent tumors, there was growth inhibition in each group. Paclitaxel and Herceptin cotreatment led to greater growth inhibition than was seen for the agents individually. Thus, in these prostate cancer model systems, Herceptin alone has clinical activity only in the androgen-dependent tumor and has at least an additive effect on growth, in combination with paclitaxel, in both androgen-dependent and androgen-independent tumors. Response to Herceptin did not correlate with the PSA levels, because the PSA index markedly increased in the Herceptin-treated group, whereas it remained constant in the control group. These results suggest the utility of Herceptin in the treatment of human prostate cancer. PMID- 10519380 TI - Dominant-negative mutations of the tumor suppressor p53 relating to early onset of glioblastoma multiforme. AB - Previous experiments have suggested that some mutant forms of p53 are able to inactivate the endogenous wild-type p53 protein in a dominant-negative fashion. However, it remains unknown whether tumors with such dominant-negative (transdominant) p53 mutants have a biological significance that is different from that of recessive p53 mutants. In this study, we examined the dominant-negative potential of various p53 mutants using a yeast-based assay in which both wild type and mutant p53 were efficiently expressed. We tested a total of 106 p53 mutants, which were identified in brain tumors, glioblastoma multiforme-derived cell lines, breast cancers, or premalignant lesions and squamous cell carcinomas of oral epithelium or were otherwise created by mutagenesis. In agreement with the previous studies, our results demonstrated that transdominant mutations affected amino acid residues that are essential for the stabilization of the DNA binding surface in the p53 core domain and for the direct interaction of p53 with its DNA-binding sequence. Among 40 patients with sporadic glioblastomas, the average age at diagnosis was significantly younger in the patients with tumors harboring dominant-negative mutations (30.4 +/- 14.7 years, n = 7) than it was in those with recessive mutations (55.2 +/- 18.6 years, n = 9, P < 0.012) and in those without mutations (54.7 +/- 17.1 years, n = 24, P < 0.003). Our data suggest that dominant-negative p53 mutants accelerate development and/or growth of glioblastoma anlagen. PMID- 10519381 TI - Tumor development under angiogenic signaling: a dynamical theory of tumor growth, treatment response, and postvascular dormancy. AB - The effects of the angiogenic inhibitors endostatin, angiostatin, and TNP-470 on tumor growth dynamics are experimentally and theoretically investigated. On the basis of the data, we pose a quantitative theory for tumor growth under angiogenic stimulator/inhibitor control that is both explanatory and clinically implementable. Our analysis offers a ranking of the relative effectiveness of these inhibitors. Additionally, it reveals the existence of an ultimate limitation to tumor size under angiogenic control, where opposing angiogenic stimuli come into dynamic balance, which can be modulated by antiangiogenic therapy. The competitive influences of angiogenically driven growth and inhibition underlying this framework may have ramifications for tissue size regulation in general. PMID- 10519382 TI - Addition of peroxisome proliferator-activated receptor alpha to guinea pig hepatocytes confers increased responsiveness to peroxisome proliferators. AB - The fibrate drugs, such as nafenopin and fenofibrate, show efficacy in hyperlipidemias but cause peroxisome proliferation and liver tumors in rats and mice via nongenotoxic mechanisms. However, humans and guinea pigs appear refractory to these adverse effects. The peroxisome proliferator (PP)-activated receptor alpha (PPAR alpha) mediates the effects of PPs by heterodimerizing with the retinoid X receptor (RXR) to bind to DNA at PP response elements (PPREs) upstream of PP-regulated genes, such as acyl-CoA oxidase. Hepatic expression of PPAR alpha in guinea pigs and humans is low, suggesting that species differences in response to PPs may be due at least in part to quantity of PPAR alpha. To test this hypothesis, we introduced mouse PPAR alpha and its heterodimerization partner, RXR alpha, into guinea pig hepatocytes by transient transfection and determined responsiveness to the PP nafenopin by cyanide-insensitive palmitoyl CoA oxidation (CIPCO). Expression of the mRNA for mouse PPAR alpha in transfected guinea pig hepatocytes was verified using species-specific PCR. In guinea pig hepatocytes transfected with control plasmids and treated with 50 microM nafenopin in the absence or presence of the RXR ligand, 9-cis-retinoic acid (5 microM) gave only a 1.7 +/- 1.5- or 3.3 +/- 1.5-fold induction in CIPCO, respectively. However, addition of ligands to hepatocytes co-transfected with both mPPAR alpha and RXR gave a strong induction of CIPCO (14.8 +/- 8.6-fold). Mouse, human, and guinea pig PPAR alpha showed equivalent function in the CIPCO assays. Thus, quantity of PPAR alpha plays a significant role in the lack of response to PPs in guinea pigs. In humans, however, lack of PPAR alpha may be only one factor dictating lack of response because recent data show that the human acyl-CoA oxidase gene lacks a functional PP response element. PMID- 10519383 TI - Solid tissues removed from ATM homozygous deficient mice do not exhibit a mutator phenotype for second-step autosomal mutations. AB - The presence of increased frequencies of blood-derived and solid tumors in ataxia telangiectasia (A-T) patients, coupled with a role for the ATM (A-T mutation) protein in detecting specific forms of DNA damage, has led to the assumption of a mutator phenotype in A TM-deficient cells. Supporting this assumption are observations of increased rates of chromosomal aberrations and intrachromosomal homologous recombinational events in the cells of A-T patients. We have bred mice with knockout mutations for the selectable Aprt (adenine phosphoribosyltransferase) locus and the Atm locus to examine the frequency of second-step autosomal mutations in Atm-deficient cells. Two solid tissues were examined: (a) the ear, which yields predominately mesenchymal cells; and (b) the kidney, which yields predominately epithelial cells. We report here the lack of a mutator phenotype for inactivating autosomal mutations in solid tissues of the Atm-deficient mice. PMID- 10519384 TI - p53-mutant clones and field effects in Barrett's esophagus. AB - Previous studies have demonstrated multifocal neoplasia in Barrett's esophagus. We evaluated 213 mapped, flow-purified, endoscopic biopsies to determine the distribution of p53-mutant clones in the Barrett's segments of 58 patients who had high-grade dysplasia without cancer. Twenty-nine patients (50%) had p53 mutations in their Barrett's segments, including 3 patients with multiple distinct p53 mutations. p53-mutant clones, including diploid cell populations, underwent expansion from 1 to 9 cm in the Barrett's segment. In 12 of 29 patients (41%) with a p53 mutation, the same mutation was found at every evaluated level of the metaplastic epithelium. This extensive p53-mutant clonal expansion suggests a somatic genetic basis for previous observations of field effects in Barrett's esophagus. PMID- 10519385 TI - Functional activity of ectopically expressed estrogen receptor is not sufficient for estrogen-mediated cyclin D1 expression. AB - Estrogen receptor function can drive cyclin D1 expression and proliferation in human breast cancer cells (MCF-7). Recent studies showing that estrogen receptor positive epithelial cells in the human mammary gland are nonproliferative suggest that the direct mitogenic effect of estrogen on mammary epithelial cells may be acquired during breast cancer development. Because estrogen-dependent cyclin D1 expression has been linked to its mitogenicity, we characterized the ability of estrogen to regulate cyclin D1 expression in estrogen receptor-negative breast cancer cells (MDA-MB-231) and nontransformed human keratinocytes (HaCaT) stably expressing the estrogen receptor. In both cases, estrogen receptor function did not induce cyclin D1 expression. Although MCF-7 cells respond to estrogen by inducing the AP-1 family components c-Fos and c-Jun, HaCaT cells expressing estrogen receptor do not. These results may explain the lack of estrogen dependent cyclin D1 expression and proliferation in cells ectopically expressing the estrogen receptor. Therefore, estrogen receptor function alone is not sufficient for estrogen-dependent cyclin D1 expression and proliferation. Other transcriptional cofactors that allow estrogen receptor to induce expression of AP 1 may be required for estrogen to act as a mitogen. PMID- 10519386 TI - The expression of a truncated HMGI-C gene induces gigantism associated with lipomatosis. AB - Rearrangements of the HMGI-C gene have frequently been detected in human benign tumors of mesenchymal origin, including lipomas. The HMGI-C protein has three AT hook domains and an acidic COOH-terminal tail. The HMGI-C modifications consist in the loss of the C-tail and the fusion with ectopic sequences. Recent results show that the loss of the COOH-terminal region, rather than the acquisition of new sequences, is sufficient to confer to HMGI-C the ability to transform NIH3T3 cells. Therefore, transgenic mice carrying a HMGI-C construct (HMGI-C/T), containing only the three AT-hook domains, were generated. The HMGI-C/T mice showed a giant phenotype, together with a predominantly abdominal/pelvic lipomatosis, suggesting a pivotal role of the HMGI-C truncation in the generation of human lipomas. PMID- 10519387 TI - Loss of Fhit is frequent in stage I non-small cell lung cancer and in the lungs of chronic smokers. AB - Abnormalities of FHIT, a candidate tumor suppressor gene at 3p14.2, have been found frequently in multiple tumor types including non-small cell lung cancer (NSCLC). To investigate whether FHIT inactivation plays a role in early lung tumorigenesis, Fhit levels were determined by immunohistochemistry in tumors from 87 patients with stage I NSCLC and in 372 bronchial biopsy specimens from 86 chronic smokers without evidence of malignancy. We found that 49% of NSCLC specimens demonstrated significantly decreased staining or lack of staining for Fhit. However, Fhit expression status was not significantly associated with disease-free survival or overall survival. Analysis of a subset of 76 specimens on which microsatellite analysis at the FHIT locus was performed did not show a strong association between loss of heterozygosity at FHIT and Fhit expression, suggesting the presence of complex mechanisms of Fhit inactivation. Of 372 bronchial biopsies from chronic smokers, 86 biopsies (23%) exhibited decreased Fhit expression or lack of Fhit expression. In 37 of 86 (43%) subjects, decreased Fhit expression or lack of expression was observed in at least one biopsy site. Loss of Fhit expression was significantly higher in bronchial metaplastic lesions (23 of 49 lesions, 47%) than in histologically normal bronchial epithelium (63 of 323 specimens, 20%; P < 0.001). Smokers with a metaplasia index of > 15% had a higher frequency of loss of Fhit expression than those with a metaplasia index of < or = 15% (P = 0.015). Interestingly, current smokers had a higher rate of loss of Fhit expression than former smokers (P = 0.02). Our data indicate that Fhit expression is significantly reduced in a substantial number of early-stage NSCLC and preneoplastic lesions in chronic smokers. The association between cigarette smoking and Fhit expression suggests a role for FHIT in the initiation of smoking related lung tumorigenesis. PMID- 10519389 TI - Decreased sensitivity to 1-O-octadecyl-2-O-methyl-glycerophosphocholine in MCF-7 cells adapted for serum-free growth correlates with constitutive association of Raf-1 with cellular membranes. AB - We have previously shown that inhibition of MCF-7 cell proliferation by 1-O octadecyl-2-O-methyl-glycerophosphocholine (ET-18-OCH3) is linked to a drug induced decrease in membrane Raf-1 levels and the subsequent inhibition of mitogen-activated protein (MAP) kinase activation in response to growth factor stimulation. We now report that adaptation of MCF-7 cells for growth in a serum free formulation results in decreased sensitivity to growth inhibition by ET-18 OCH3. The decrease in ET-18-OCH3 sensitivity occurred progressively during the adaptation process and correlated with the presence of increasing amounts of inactive Raf-1 that stably associated with MCF-7 cell membranes. ET-18-OCH3 sensitivity could be restored by growing the adapted cells in serum-containing medium, which resulted in the loss of membrane-associated Raf-1. In human normal mammary epithelial cells, which are insensitive to ET-18-OCH3, Raf-1 was also associated with membranes in quiescent cells. In both cell types, incubation with ET-18-OCH3 had no effect on the membrane-Raf-1 levels, suggesting that ET-18-OCH3 induced reduction of Raf-1 levels in growth factor-stimulated MCF-7 cells is due to inhibition of Raf translocation. The activation and termination of the MAP kinase pathway in response to growth factors in the adapted MCF-7 cells and HNME cells occurred without changes to membrane Raf-1 levels. Because membrane translocation is not required to activate Raf in these cells, inhibition of Raf translocation by ET-18-OCH3 subsequent to cell stimulation has no effect on the activation of the membrane-bound Raf and, consequently, the activation of the MAP kinase pathway. The ability of the cells to activate the MAP kinase pathway in the presence of the drugs enables them to resist the growth-inhibitory effects of the drug, leading to the observed ET-18-OCH3 insensitivity of the cells. PMID- 10519388 TI - Extracellular signal-regulated kinase activation is required for up-regulation of vascular endothelial growth factor by serum starvation in human colon carcinoma cells. AB - Vascular endothelial growth factor (VEGF) is a potent angiogenic factor important for colon cancer neovascularization. In previous studies, serum starvation led to induction of VEGF in human colon carcinoma cells. We investigated the possible participation of mitogen-activated protein kinases in serum starvation induction of VEGF in the HT29 human colon carcinoma cell line. The extracellular signal regulated kinases (Erks) 1 and 2 were activated after 3-6 h of serum starvation. Using transient transfection of VEGF promoter-reporter constructs, serum starvation led to an increase in VEGF promoter activity. An inhibitor of phosphorylation of Erk-1/2 blocked the increase of VEGF expression and promoter activity induced by serum starvation. Serum starvation activates several mitogen activated protein kinases, but activation of Erk-1/2 is critical for the up regulation of VEGF mRNA in colon carcinoma cells. PMID- 10519391 TI - High-salt diet induces gastric epithelial hyperplasia and parietal cell loss, and enhances Helicobacter pylori colonization in C57BL/6 mice. AB - A high-salt diet in humans and experimental animals is known to cause gastritis, has been associated with a high risk of atrophic gastritis, and is considered a gastric tumor promoter. In laboratory rodents, salt is known to cause gastritis, and when coadministered, it promotes the carcinogenic effects of known gastric carcinogens. Because Helicobacter pylori has been associated with a progression from gastritis to gastric cancer, we designed a study to determine whether excessive dietary NaCl would have an effect on colonization and gastritis in the mouse model of H. pylori infection. Seventy-two, 8-week-old female C57BL/6 mice were infected with H. pylori strain Sydney, and 36 control mice were dosed with vehicle only. One-half of the infected and control mice were fed a high-salt diet (7.5% versus 0.25%) for 2 weeks prior to dosing and throughout the entire experiment. Twelve infected and 6 control animals from the high-salt and normal diet groups were euthanized at 4, 8, and 16 weeks. At 8 and 16 weeks postinfection (WPI), the colony-forming units per gram of tissue were significantly higher (P < 0.05) in the corpus and antrum of animals in the high salt diet group compared with those on the normal diet. Quantitative urease was significantly higher (P < 0.05) at 4 and 8 WPI in the corpus and antrum of animals on the high-salt diet when compared with controls. At 16 WPI, mice in both the normal and the high-salt diet groups developed moderate to marked atrophic gastritis of the corpus in response to H. pylori infection. However, the gastric pits of the corpus mucosa in mice on the high-salt diet were elongated and colonized by H. pylori more frequently than those in mice on the normal diet. The high-salt diet was also associated with a significant increase in proliferation in the proximal corpus and antrum and a multifocal reduction in parietal cell numbers in the proximal corpus, resulting in the elongation of gastric pits. We conclude that excessive NaCl intake enhances H. pylori colonization in mice and in humans and that chronic salt intake may exacerbate gastritis by increasing H. pylori colonization. Furthermore, elevated salt intake may potentiate H. pylori-associated carcinogenesis by inducing proliferation, pit cell hyperplasia, and glandular atrophy. PMID- 10519390 TI - Additivity of dilantin and vinblastine inhibitory effects on microtubule assembly. AB - Dilantin (phenytoin) is a commonly used antiepileptic agent that is known to decrease conductance of sodium and calcium ions and delay outward potassium currents. Separate from its antiseizure activity, dilantin interferes with microtubule protein polymerization. It induces metaphase arrest and potentiates the effects of the antimitotics vincristine and vinblastine in cell culture. We show here by fluorescence binding studies that dilantin interacts directly with tubulin at a low affinity site [Ka = 3.5 (+/- 2.5) x 10(3) M(-1); Kd = 286 microM]. We quantitatively examined the effect of dilantin on bulk microtubule formation and found that the drug raises the critical concentration for microtubule polymerization in 2 M glycerol identically in the presence or absence of vinblastine. The change in free energy for microtubule polymerization attributable to 400 microM dilantin [deltadelta G = 117 (+/- 28) cal/mol] is additive with vinblastine effects. Under the same conditions, mean microtubule lengths are 7.7 +/- 4.3 microm (n = 558) and 7.4 +/- 4.0 microm (n = 477) in the presence or absence of dilantin, respectively. Dilantin has no effect on vinblastine-induced tubulin spiral formation, as measured by sedimentation velocity. Our data suggest that the mechanism for the antimicrotubule effects of dilantin involves sequestration of tubulin heterodimers in 1:1 drug:tubulin complexes that do not participate in tubulin polymerization. The dilantin binding site is distinct from the Vinca binding site, and these independent binding modes account for the additive effects in vitro. The sequestration of tubulin heterodimers could explain the combined drug synergy in cell cultures if it disrupted interactions with proteins that regulate microtubule dynamics and/or cell cycle events. PMID- 10519392 TI - Tamoxifen-DNA adduct formation in rat liver determined by immunoassay and 32P postlabeling. AB - Tamoxifen (TAM), a nonsteroidal antiestrogen used as a chemotherapeutic and chemopreventive agent for breast cancer, induces liver tumors in rodents and covalent DNA adduct formation in hepatic DNA. Here, we report the development and validation of highly sensitive and specific immunoassays for the determination of TAM-DNA adducts. Rabbits were immunized with calf thymus DNA, chemically modified with alpha-acetoxytamoxifen to 2.4 adducts per 100 nucleotides, and the resulting antisera were characterized by competitive dissociation-enhanced lanthanide fluoroimmunoassay (DELFIA) and chemiluminescence immunoassay (CIA). Compared with DELFIA, the CIA has a much lower background and a 20-fold increase in sensitivity. For the immunogen TAM-DNA, 50% inhibition was at 2.0 +/- 0.11 (mean +/- SE, n = 18) fmol of (E)-alpha-(N2-deoxyguanosinyl)tamoxifen (TAM-dG) adduct in TAM-DNA by DELFIA. For TAM-DNA modified to 4.8 adducts in 10(6) nucleotides, 50% inhibition was at 20.6 +/- 6.6 (mean +/- SE, n = 8) fmol of TAM-dG in TAM-DNA by DELFIA and at 0.92 +/- 0.11 (mean +/- SE, n = 10) fmol of TAM-dG in TAM-DNA by CIA. No inhibition was observed in either assay with up to 20 microg (62.5 nmol of nucleotides) of unmodified DNA. The individual adducts TAM-dG and (Z)-alpha (N2-deoxyguanosinyl)tamoxifen and the individual compounds TAM and 4-OH-TAM gave DELFIA 50% inhibitions at 828, 2229, 5440, and 8250 fmol, respectively. For assay validation, TAM-dG levels were determined by DELFIA, CIA, and 32P-postlabeling in TAM-DNA samples modified in vitro to different levels, and comparable values were obtained in all three assays. Further validation was obtained in vivo in rat liver. DNA adducts of TAM were measurable in rat liver 24 h after a single i.p. dose of 45 mg TAM/kg body weight and after daily p.o. dosing for 7 days with 5.0, 10.0, and 20.0 mg TAM/kg body weight. In addition, TAM-DNA adducts disappeared slowly over 21 days in rats on a control diet that were first given p.o. TAM at 45 mg/kg/day for 4 days. In the rat experiments, TAM-DNA adduct levels determined by CIA compared well with those determined by 32P-postlabeling, although the CIA gave an underestimation at the highest doses. For rat liver samples, the detection limit by CIA was 3 adducts per 10(9) nucleotides (0.2 fmol of adducts per 20 microg of DNA). PMID- 10519393 TI - Loss of responsiveness to transforming growth factor beta induces malignant transformation of nontumorigenic rat prostate epithelial cells. AB - Transforming growth factor (TGF)-betas are multifunctional growth factors, the properties of which include the potent inhibition of epithelial cell growth. Expression patterns of TGF-betas and TGF-beta receptors in the normal prostate indicate that these growth regulators play key roles in prostatic development and proliferative homeostasis. Importantly, TGF-beta receptor levels are frequently diminished in malignant human prostate tissue. To test the hypothesis that loss of TGF-beta responsiveness is causally involved in the tumorigenic process, we have used retroviral transduction to introduce a dominant-negative mutant type II TGF-beta receptor (DNR) into the premalignant rat prostatic epithelial cell line, NRP-152. High-level expression of the DNR abolished the ability of TGF-beta to inhibit cell growth, to promote cell differentiation, and to induce apoptosis, and it partially blocked the induction of extracellular matrix gene expression. When injected into nude mice, NRP-152-DNR cells formed carcinomas at 13 of 34 sites, compared with 0 of 30 sites for parental and control cells (P = 0.0001). We conclude that the type II TGF-beta receptor is an important tumor suppressor in the prostate, and furthermore, that loss of TGF-beta responsiveness can contribute early in the tumorigenic process by causing the malignant transformation of preneoplastic cells. PMID- 10519394 TI - Extracellular matrix and radiation G1 cell cycle arrest in human fibroblasts. AB - It is thought that sublethal doses of radiation cause cells to pause in either G1 or G2 phase, but that then cells with repaired DNA damage reenter the cell cycle. However, it has been observed that gamma-irradiation causes normal human fibroblasts to arrest indefinitely in G1 phase unless the irradiated cells are subcultured. This indicates that cell adhesion plays a role in maintaining the arrest. We now show that the type of extracellular matrix dramatically affects the percentage of cells that arrest in G1 phase. The prolonged radiation G1 arrest in human fibroblasts has been referred to as "senescence-like"; however, we find that smooth muscle alpha-actin is highly expressed in cells that arrest in G1 phase after irradiation. This indicates that the fibroblasts differentiate to myofibroblasts. Together, our results show that the length of radiation G1 arrest in human fibroblasts is affected by the type of extracellular matrix on which the cells are irradiated and that arrest results in myofibroblast differentiation. PMID- 10519395 TI - Apoptosis induced by vitamin D compounds in breast cancer cells is inhibited by Bcl-2 but does not involve known caspases or p53. AB - The hormonally active form of vitamin D3, 1,25-dihydroxyvitamin D3, and its two analogues, EB 1089 and CB 1093, are novel putative anticancer agents with an interesting profile of induction of growth inhibition, differentiation, and apoptosis in tumor cells. To study the signaling pathways mediating these events, we used two human breast cancer cell lines: MCF-7 cells, expressing a wild-type p53 tumor suppressor protein, and T47D cells, lacking a functional p53. Vitamin D compounds induced a growth arrest followed by apoptosis in both cell lines at concentrations ranging from 1 to 100 nM, indicating that p53 is not necessary for growth-inhibitory effects induced by vitamin D compounds. Surprisingly, apoptosis induced by these compounds occurred also independently of known caspases. Inhibition of caspase activation by overexpression of a cowpox-derived caspase inhibitor CrmA or by addition of inhibitory peptides acetyl-Asp-Glu-Val-Asp aldehyde (200 microM), acetyl-Ile-Glu-Thr-Asp-aldehyde (50 microM), and Z-Val-Ala D,L-Asp-fluoromethylketone (1 microM) showed no effect on the induction of growth arrest or apoptosis by vitamin D compounds under assay conditions in which apoptosis induced by TNF or staurosporine was effectively inhibited. Moreover, overexpression of caspase-3 in MCF-7 cells had no sensitizing effect to vitamin D compounds, and neither caspase-3-like protease activity nor cleavage of a caspase substrate poly(ADP)ribose polymerase was detected in lysates from apoptotic cells following the treatment with these compounds. Contrary to CrmA, overexpression of an antiapoptotic protein Bcl-2 in MCF-7 cells conferred a nearly complete protection from apoptosis induced by vitamin D compounds. Taken together, these data indicate that vitamin D compounds induce apoptosis via a novel caspase- and p53-independent pathway that can be inhibited by Bcl-2. This may prove useful in the treatment of tumors that are resistant to therapeutic agents that are dependent on the activation of p53 and/or caspases. PMID- 10519396 TI - Effect of combined treatment with the pure antiestrogen EM-800 and radiotherapy on the growth of human ZR-75-1 breast cancer xenografts in nude mice. AB - Human breast tumors are usually composed of heterogeneous cell populations that exhibit different sensitivities to therapeutic agents. We therefore investigated the effect of treatment with various regimens of the novel pure antiestrogen EM 800, alone or in combination with external beam radiation therapy, on the growth of human ZR-75-1 xenografts in athymic mice. The animals received a maximal dose of EM-800 (300 microg, p.o.) and/or radiotherapy at the dose of 10 Gy. 2.5 Gy fractions were administered over a 9-day period in four sessions of 13.7 min each (250-kilovolt Siemens with 2-mm aluminum filtration at 90 cm from the source origin). EM-800 was administered p.o. once daily, whereas radiotherapy was repeated every 35 days. Tumor size was expressed as a percentage of the initial tumor size, which was assigned a value of 100%. Average tumor size increased by 514% in ovariectomized mice supplemented with estrone alone for 259 days compared with the pretreatment value. Treatment with radiotherapy or EM-800 alone resulted in 11 and 73% decreases in mean tumor size, respectively, whereas combined treatment given simultaneously at the beginning caused a dramatic 98% decrease in tumor size. The start of radiotherapy on day 35 in EM-800-treated mice, or conversely, the start of EM-800 in irradiated mice at the 35-day time interval, resulted in somewhat lower, 88% and 95%, decreases in tumor size, respectively. In animals receiving EM-800 alone, 40% of tumors disappeared, thus indicating a cytotoxic effect caused by the estrogen blockade achieved with the pure antiestrogen. Eighty-six % of the original tumors disappeared under continuous combined treatment. Most importantly, no tumor reappeared under estrogenic stimulation after stopping treatment, thus indicating cure of 86% of the tumors in the group of animals who received the combination therapy. The present data indicate that combined treatment with EM-800 and radiotherapy yields a faster response, a greater decrease in tumor size, and a higher percentage of complete responses or tumor disappearance (cure) than either treatment used alone. The present data also suggest that maximal benefits are achieved when the pure antiestrogen is administered continuously, starting at the same time as radiation therapy and continued without interruption as adjuvant therapy. The present data also clearly show that efficient blockade of estrogens with a potent and pure antiestrogen is not only cytostatic but is cytotoxic and can lead to the disappearance of an important proportion of tumors or cure. PMID- 10519397 TI - Adrenal androgens stimulate the proliferation of breast cancer cells as direct activators of estrogen receptor alpha. AB - Estrogens stimulate the proliferation of many breast tumors and cell lines derived from them. Antiestrogens have therefore become a powerful therapeutic agent to treat breast tumors that express estrogen receptor (ER) alpha. In addition, aromatase inhibitors are now used in postmenopausal women to block the in situ conversion of adrenal androgens to estrogens. This approach can only be successful if ER-alpha in a particular tumor is not directly stimulated by adrenal androgens. We have examined this possibility using several different cell lines as model systems: (a) wild-type MCF7 cells, an ER-alpha-dependent human breast cancer cell line; (b) MCF7SH cells, an estrogen-independent MCF7 variant; (c) Ishikawa cells, an ER-alpha-containing human uterine cell line; (d) ER negative HeLa cells; and (e) budding yeast. Transactivation assays with transfected ER-alpha reporter genes reveal a direct activation of ER-alpha by dehydroepiandrosterone (DHEA), 5alpha-androstene-3beta,17beta-diol, testosterone, and the two nonaromatizable androgens, dihydrotestosterone and 5alpha-androstane 3beta,17beta-diol. The involvement of other steroid receptors could be ruled out with specific antihormones. Moreover, the same set of ligands stimulates the proliferation of the two breast cancer cell lines. At subsaturating and physiologically relevant concentrations of DHEA, DHEA stimulates the proliferation of MCF7SH cells, which correlates with a substantial, albeit submaximal, transcriptional response. Thus, adrenal androgens must also be considered as risk factors in postmenopausal women. PMID- 10519398 TI - Breast cancer risk associated with genotype polymorphism of the estrogen metabolizing genes CYP17, CYP1A1, and COMT: a multigenic study on cancer susceptibility. AB - Estrogen has been proposed to trigger breast cancer development via an initiating mechanism involving its metabolite, catechol estrogen (CE). To examine this hypothesis, we conducted a multigenic case-control study to determine whether polymorphisms of the genes responsible for CE formation via estrogen biosynthesis (CYP17) and hydroxylation (CYP1A1) and CE inactivation (COMT) are associated with an elevated risk for breast cancer in Taiwanese women, and whether the association between genotype and risk may be modified by estrogen exposure. One hundred and fifty breast cancer patients and 150 healthy controls were recruited. PCR-based RFLP assays were used to determine the genotypes of estrogen metabolizing genes. The breast cancer risk associated with individual susceptibility genotypes varied among the three genes and was highest for COMT, followed by CYP1A1 and CYP17. After simultaneous consideration of all three genes and other well-established risk factors of breast cancer, the COMT genotype remained the most significant determinant for breast cancer development and was associated with a 4-fold increase in risk (95% confidence interval, 1.12-19.08). Furthermore, a trend of increasing risk for developing breast cancer was found in women harboring higher numbers of high-risk genotypes (P = 0.006), including the high activity CYP17 (CYP17 A2/A2), high inducibility CYP1A1 (CYP1A1 MspI vt/vt), and low activity COMT (COMT L/L) genotypes. The association of risk with the number of susceptibility genotypes was stronger in women with prolonged estrogen exposure (indicated by a higher number of estrogen exposure years or a higher number of estrogen exposure years between menarche and first full-term pregnancy), women with higher estrogen levels (implied by early menarche), and women with a higher body mass index (> or = 22.5). On the basis of comprehensive profiles of estrogen metabolism, this study supports the possibility that breast cancer can be initiated by estrogen exposure. PMID- 10519399 TI - Antisense cyclin D1 induces apoptosis and tumor shrinkage in human squamous carcinomas. AB - Cyclin D1 plays an essential regulatory role in the G1 phase of the cell cycle. The cyclin D1 gene is amplified in 20-50% of squamous cell carcinomas (SCCs), and the protein is overexpressed in up to 80% of SCCs. Our hypothesis was that gene transduction of antisense (AS) cyclin D1 in human SCCs in vivo would result in tumor reduction. A cyclin D1 cDNA was inserted into an E1/E3-deficient serotype 5 adenovirus (AS cyclin D1) in an AS orientation using homologous recombination. AS cyclin D1 transduction suppressed cyclin D1 protein expression in both cultured cells and tumors. AS cyclin D1 significantly inhibited cell proliferation by both [3H]thymidine incorporation in six SCC cell lines (P = 0.01-0.001) and the conversion of tetrazolium salt to formazan in four SCC cell lines (P = 0.01 0.004). Apoptosis detected in >25% of cells in each cell line 48 h after AS cyclin D1 transduction paralleled the reduction in cyclin D1 protein. Preformed SCCs transduced with AS cyclin D1 were significantly inhibited (P = 0.002-0.005), and apoptosis was prominent in the AS cyclin D1-treated tumors, but not in tumors treated with the control vector. These data extend prior in vitro and ex vivo results and indicate that AS cyclin D1 suppresses SCC growth both in vitro and in vivo through suppression of cyclin D1 protein expression, leading to cellular apoptosis. Our findings suggest that cyclin D1 may have a role in cell survival and that cyclin D1 AS therapy may be useful as an adjunct to standard treatment for SCC. PMID- 10519400 TI - Hypoxia-mediated apoptosis from angiogenesis inhibition underlies tumor control by recombinant interleukin 12. AB - The role of angiogenesis inhibition in the antitumor activity of recombinant murine interleukin 12 (rmIL-12) was studied in K1735 murine melanomas, the growth of which is rapidly and markedly suppressed by rmIL-12 treatment. On the basis of the prediction that tumor ischemia should result from therapeutic angiogenesis inhibition, tumor cell hypoxia was evaluated as a marker of ischemia using the EF5 [2-(2-nitro-1H-imidazol-1-yl)-N-(2,2,3,3,3-pentafluoropropyl)aceta mide] approach. This method measures intracellular binding of the nitroimidazole EF5, which covalently binds to cellular macromolecules selectively under hypoxic conditions. Whereas 1 week of rmIL-12 treatment effectively inhibited K1735 cell induced angiogenesis in Matrigel neovascularization assays, 2 weeks of treatment were needed before severe tumor cell hypoxia was detected in K1735 tumors. The hypoxia that developed was regional and localized to tumor areas distant from blood vessels. The great majority of severely hypoxic tumor cells were apoptotic, and in vitro studies indicated that the degree of hypoxia present within treated tumors was sufficient to trigger K1735 apoptosis. Tumor cell apoptosis was also prevalent in the first week of rmIL-12 treatment when few cells were hypoxic. In vitro studies indicated that this non-hypoxia-related apoptosis was induced directly by IFN-gamma produced in response to rmIL-12 administration. These studies reveal that rmIL-12 controls K1735 tumors initially by IFN-gamma-induced apoptosis and later by hypoxia-induced apoptosis. They also establish hypoxia as an expected result of tumor angiogenesis inhibition and a mediator of its therapeutic effect. PMID- 10519401 TI - Mouse transporter protein, a membrane protein that regulates cellular multidrug resistance, is localized to lysosomes. AB - Mouse transporter protein (MTP), a small, highly conserved mammalian intracellular membrane protein with four putative transmembrane domains, has been implicated in the transport of nucleosides and/or related molecules across intracellular membranes. The production of recombinant MTP in Saccharomyces cerevisiae alters sensitivity of yeast cells to a heterogeneous group of compounds (e.g., antimetabolites, antibiotics, anthracyclines, ionophores, and steroid hormones) by changing the subcellular compartmentalization of these drugs, suggesting that MTP functions similarly in higher organisms. The present study was undertaken to define the intracellular location of MTP in mammalian cells. Native MTP was not detected by indirect immunofluorescence in cell types that expressed MTP mRNA; therefore, a hemagglutinin (HA) epitope-tagged version of MTP was produced in cultured BHK21 cells by transient transfection, and its distribution within cells was determined by confocal microscopy using antibodies directed against the HA epitope and various organellar proteins. Antibodies directed against HA-MTP colocalized with antibodies against late endosomal and lysosomal proteins but not with antibodies against either Golgi or early endosomal proteins. Analysis of subcellular fractions from rat liver by immunoblotting with antibodies directed against MTP demonstrated the presence of a MTP-like protein in Golgi- and lysosome-enriched membranes but not in mitochondria. These results indicate that MTP resides in late endosomes and lysosomes, a finding that is consistent with the proposed role for MTP in the movement of a variety of small molecules across endosomal and lysosomal membranes. MTP shares a number of characteristics with other lysosome-associated proteins. We, therefore, propose that it be redesignated murine lysosome associated protein transmembrane 4. PMID- 10519402 TI - Potent topoisomerase I inhibition by novel silatecans eliminates glioma proliferation in vitro and in vivo. AB - Although topoisomerase inhibitors, such as camptothecin and topotecan, have been widely used in the treatment of nonglial tumors, their application for gliomas has been limited by poor efficacy relative to toxicity that may in part reflect limited bioavailability and blood stability of these agents. However, the potential promise of this class of agents has fostered efforts to develop new, more potent, and less toxic inhibitors that may be clinically relevant. Using a cascade radical annulation route to the camptothecin family, we developed a series of novel camptothecin analogues, 7-silylcamptothecins ("silatecans"), that exhibited potent inhibition of topoisomerase I, dramatically improved blood stability, and sufficient lipophilicity to favor blood-brain barrier transit. We explored the efficacy of a series of these agents against a panel of five high grade glioma cell lines to identify a promising compound for further preclinical testing. One of the most active agents in our systems (DB67) inhibited tumor growth in vitro with an ED50 ranging between 2 and 40 ng/ml, at least 10-fold more potent than the effects observed with topotecan, and at least comparable with those of SN-38, the active metabolite of CPT-11. Because DB67 also exhibited the highest human blood stability of any of the agents examined, this agent was then selected for in vivo studies. A dose-escalation study of this agent in a nude mouse U87 glioma model system demonstrated a concentration-dependent effect, with tumor growth inhibition at day 28 postimplantation (the day control animals began to require sacrifice because of large tumor size) of 61 +/- 7% and 73 +/- 3% after administration of DB67 doses of 3 and 10 mg/kg/day, respectively, for 5 days beginning on postimplantation day 7. Animals that continued treatment with 10 mg/kg/day in three additional 21-day cycles all remained progression free after >90 days of follow-up but later developed enlarging tumors after treatment was stopped. However, a slightly higher dose (30 mg/kg/day) induced complete tumor regression after only two cycles in all study animals and was effective even if treatment was delayed until large, bulky tumors had developed. Application of the 30 mg/kg/day dose to treat established intracranial glioma xenografts led to long-term (>90 day) survival in six of six animals, whereas all controls died of progressive disease (P < 0.00001). No apparent toxicity was encountered in any of the treated animals. In summary, the present studies indicate that silatecans may hold significant promise for the treatment of high grade gliomas and provide a rationale for proceeding with further preclinical evaluation of their efficacy and safety versus commercially available camptothecin derivatives. PMID- 10519403 TI - Application of the Cre recombinase/loxP system further enhances antitumor effects in cell type-specific gene therapy against carcinoembryonic antigen-producing cancer. AB - A considerable number of studies of cancer have shown that the cell type-specific promoter is an effective tool for selective expression of foreign genes in tumor cells. However, few reports have demonstrated significant in vivo antitumor effects using this strategy thus far, possibly because the low activity of such a promoter results in insufficient expression of genes in cancer cells as well as in insignificant antitumor effects, even when the cells are infected by highly efficient gene transfer methods. To overcome this problem, we used the Cre/loxP system for the cell type-specific gene therapy against carcinoembryonic antigen (CEA)-producing cancer. We constructed a pair of recombinant Ads. One expresses the Cre recombinase (Cre) gene under the control of the CEA promoter (Ad.CEA Cre). The other contains the herpes simplex virus thymidine kinase (HSV-TK) gene separated from the strong CAG promoter by insertion of the neomycin resistance (neo) gene (Ad.lox-TK). The HSV-TK gene of the latter Ad is designed to be activated through excisional deletion of the neo gene by Cre enzyme released from the former one only when CEA-producing cells are infected simultaneously with these Ads. Coinfection by these Ads rendered a human CEA-producing cancer cell line 8.4-fold more sensitive to ganciclovir (GCV) compared with infection by Ad.CEA-TK alone, the HSV-TK gene of which is directly regulated by the CEA promoter. On the other hand, coinfection with these Ads did not significantly change the GCV sensitivity of non-CEA-producing cells. Intratumoral injection of Ad.CEA-Cre combined with Ad.lox-TK followed by GCV treatment almost completely eradicated CEA-producing tumors established in the subcutis of athymic mice, whereas intratumoral injection of Ad.CEA-TK with GCV administration at most retarded the growth of inoculated tumors. These results suggest distinct advantages of the Cre/loxP system applied in the conventional cell type-specific gene therapy against cancer. PMID- 10519404 TI - Electromotive versus passive diffusion of mitomycin C into human bladder wall: concentration-depth profiles studies. AB - The objectives of these investigations were: (a) to make a preliminary study to assess concentration-depth profiles of mitomycin C (MMC) in the bladder wall at specified time intervals after passive diffusion (PD); and (b) to conduct a major study to compare concentration-depth profiles after PD and electromotive drug administration (EMDA) of MMC. Full thickness sections of viable human bladder wall were placed in two-chamber cells with urothelium exposed to donor compartments containing 40 mg of MMC in 100 ml of 0.96% NaCl solutions and with serosa-facing receptor compartments containing 0.9% NaCl solutions. In the preliminary study during each of nine experimental sessions, five sections of bladder wall were individually exposed to MMC for either 5, 15, 30, 45, or 60 min. In the major study, an anode and a cathode were sited in the donor and receptor compartments, and 14 paired experiments--current (20 mA)/no current- were conducted over a 30-min period. Bladder wall sections were cut serially into 40-microm slices parallel to the urothelium and analyzed by high-performance liquid chromatography for MMC concentration (microg/g wet tissue weight). Tissue viability and morphology and MMC stability were assessed by trypan-blue exclusion test, histological examination, and mass spectrometry analysis. In the preliminary study (PD only), mean MMC concentrations (microg) at 5, 15, 30, 45, and 60 min were: (a) for urothelium, 15.3, 60.0, 58.2, 60.1, and 57.8, respectively; (b) for lamina propria, 2.2, 18.9, 19.3, 16.1, and 17.3, respectively; and (c) for muscularis, 0.4, 2.0, 1.8, 1.3, and 2.4, respectively. In the comparative study, MMC concentrations and coefficients of variation (CV) were as follows: (a) for urothelium after PD, 46.6 with CV = 69%, and after EMDA, 170.0 with CV = 43% (P < 0.0001); (b) for lamina propria after PD, 16.1, with CV = 60%, and after EMDA, 65.6 with CV = 29% (P < 0.0001); and (c) for muscularis after PD, 1.9 with CV = 82%, and after EMDA, 15.9 with CV = 82% (P < 0.0005). All of the bladder sections remained viable, and the chemical structure of MMC was unchanged. It was concluded that EMDA significantly enhances MMC transport into all of the layers of the bladder wall, and sections of viable human bladder are a reliable tool for assessing different modes of drug delivery. PMID- 10519405 TI - Antitumor efficacy of a novel class of non-thiol-containing peptidomimetic inhibitors of farnesyltransferase and geranylgeranyltransferase I: combination therapy with the cytotoxic agents cisplatin, Taxol, and gemcitabine. AB - Ras malignant transformation requires posttranslational modification by farnesyltransferase (FTase). Here we report on the design and antitumor activity, in monotherapy as well as in combination therapy with cytotoxic agents, of a novel class of non-thiol-containing peptidomimetic inhibitors of FTase and the closely related family member geranylgeranyltransferase I (GGTase I). The non thiol-containing FTI-2148 is highly selective for FTase (IC50, 1.4 nM) over GGTase I (IC50, 1700 nM), whereas GGTI-2154 is highly selective for GGTase I (21 nM) over FTase (IC50, 5600 nM). In whole cells, the corresponding methylester prodrug FTI-2153 is >3000-fold more potent at inhibiting H-Ras (IC50, 10 nM) than Rap1A processing, whereas GGTI-2166 is over 100-fold more selective at inhibiting Rap1A (IC50, 300 nM) over H-Ras processing. Furthermore, FTI-2153 was highly effective at suppressing oncogenic H-Ras constitutive activation of mitogen activated protein kinase and human tumor growth in soft agar. FTI-2148 suppressed the growth of the human lung adenocarcinoma A-549 cells in nude mice by 33, 67, and 91% in a dose-dependent manner. Combination therapy of FTI-2148 with either cisplatin, gemcitabine, or Taxol resulted in a greater antitumor efficacy than monotherapy. GGTI-2154 in similar antitumor efficacy experiments is less potent than FTI-2148 and inhibits tumor growth by 9, 27, and 46%. Combination therapy of GGTI-2154 with cisplatin, gemcitabine, or Taxol is also more effective. Finally, FTI-2148 and GGTI-2154 are 30- and 33-fold more selective and 30- and 16-fold more potent in whole cells than our previously reported thiol-containing FTI-276 and GGTI-297, respectively. Thus, our results demonstrate that this highly potent and selective novel class of non-thiol-containing peptidomimetics inhibits human tumor growth in whole animals and that combination therapy with cytotoxic agents is more beneficial than monotherapy. PMID- 10519406 TI - Transfection of 9-hydroxyellipticine-resistant Chinese hamster fibroblasts with human topoisomerase IIalpha cDNA: selective restoration of the sensitivity to DNA religation inhibitors. AB - In the Chinese hamster lung cell line DC-3F/9-OH-E, selected for resistance to 9 OH-ellipticine and cross-resistant to other topoisomerase II inhibitors, the amount of topoisomerase IIalpha is 4-5-fold lower than in the parental DC-3F cells, whereas topoisomerase IIbeta is undetectable. Cloning and sequencing of topoisomerase IIalpha cDNAs from DC-3F and DC-3F/9-OH-E cells revealed an allele polymorphism, one allele differing from the other by the presence of seven silent mutations and three mutations in the noncoding region. In addition, the mutated allele contains three missense mutations located close to the ATP binding site (Thr371Ser) or to the catalytic site (Ala751Gly; Ile863Thr). To analyze the contribution of these topoisomerase IIalpha alterations to their resistance phenotype, DC-3F/9-OH-E cells were transfected with an eukaryotic expression vector containing the human topoisomerase IIalpha cDNA. In one transfected clone, the amount of topoisomerase IIalpha isoform and the catalytic activity were similar to that in the parental DC-3F cells. These cells, which contain only topoisomerase IIalpha, are then a unique mammalian cell line to analyze the physiological and pharmacological properties of this enzyme. However, the restoration of a nearly normal topoisomerase IIalpha activity in the DC-3F/9-OH-E cells did not have the same effect on their sensitivity to different enzyme inhibitors; a 75% reversion of the resistance, associated with a 2-3-fold increased stabilization of the cleavable complex, was observed with both etoposide and m-AMSA, two drugs that inhibit the DNA religation step in the enzyme catalytic cycle; in contrast, the transfected cells remained fully resistant to ellipticine derivatives that did not induce the stabilization of the cleavable complex. We hypothesized that a trans-acting factor, inhibiting the induction of cleavable complex formation by drugs that are not religation inhibitors, might be present in the resistant cells. However, such a factor was not detected in in vitro experiments, and other hypotheses are discussed. PMID- 10519407 TI - Arabinosylguanine-induced apoptosis of T-lymphoblastic cells: incorporation into DNA is a necessary step. AB - 9-Beta-D-Arabinosylguanine (ara-G) is a recently introduced and effective treatment for T-cell acute lymphoblastic leukemia, but how ara-G and ara-G triphosphate (ara-GTP) kill cells is not known. We hypothesized that, in cycling T-lymphoblastoid cells, ara-G may act directly by incorporation into DNA, which may lead to apoptosis. Hence, blocking the incorporation of ara-G monophosphate (ara-GMP) into DNA may prevent apoptosis. To test this hypothesis, we performed experiments in a T-lymphoblastic leukemia cell line (CCRF-CEM) after synchronization with a double aphidicolin block. Intracellular accumulation of ara-GTP was neither cell cycle dependent nor affected by aphidicolin (53 +/- 5 microM/h without aphidicolin, 50 +/- 5 microM/h with aphidicolin). Cells at the G1-S boundary accumulated 75 +/- 7 microM ara-GTP with minimal incorporation into DNA (5 +/- 2 pmol ara-GMP/mg DNA) and had little biochemical or morphological evidence of apoptosis. In marked contrast, cells in S phase had significantly more ara-G incorporated into DNA (24 +/- 4 pmol ara-GMP/mg DNA), although the cytosolic concentration of ara-GTP (85 +/- 7 microM) was similar to that in the G1-enriched population. In the S-phase cells, there was a corresponding increase in apoptosis (measured as high molecular weight DNA fragmentation and morphological changes), and the incorporation of ara-GTP into DNA resulted in a >95% inhibition of DNA synthesis. There was a direct linear relationship between the number of cells in S phase and both the total number of ara-GMP molecules in DNA and the inhibition of DNA synthesis. Blocking of ara-GTP incorporation into S phase DNA abolished biochemical and morphological features of apoptosis, even in the presence of cytotoxic level of intracellular ara-GTP. Taken together, these data demonstrate that the incorporation of ara-GTP into DNA is the critical event that mediates the induction of apoptosis in CCRF-CEM cells. PMID- 10519408 TI - The polyamine oxidase inhibitor MDL-72,527 selectively induces apoptosis of transformed hematopoietic cells through lysosomotropic effects. AB - Polyamine oxidase functions in the polyamine catabolic pathway, converting N1 acetyl-spermidine and -spermine into putrescine (Put) and spermidine (Spd), respectively, thereby facilitating homeostasis of intracellular polyamine pools. Inhibition of polyamine oxidase in hematopoietic cells by a specific inhibitor, N,N'-bis(2,3-butadienyl)-1,4-butanediamine (MDL-72,527), reduces the levels of Put and Spd and induces the accumulation of N1-acetylated Spd. Although previously thought to be relatively nontoxic, we now report that this inhibitor overrides survival factors to induce cell death of several immortal and malignant murine and human hematopoietic cells, but not of primary myeloid progenitors. Cells treated with MDL-72,527 displayed biochemical changes typical of apoptosis, and cell death was associated with the down-regulation of the antiapoptotic protein Bcl-X(L). However, enforced overexpression of Bcl-X(L), or treatment with the universal caspase inhibitor zVAD-fmk, failed to block MDL-72,527-induced apoptosis in these hematopoietic cells. Despite decreases in Put and Spd pools, MDL-72,527-induced apoptosis was not blocked by cotreatment with exogenous Put or Spd, nor was it influenced by overexpression or inhibition of the polyamine biosynthetic enzyme ornithine decarboxylase. Significantly, MDL-72,527-induced apoptosis was associated with the rapid formation of numerous lysosomally derived vacuoles. Malignant leukemia cells were variably sensitive to the lysosomotropic effects of MDL-72,527, yet pretreatment with the ornithine decarboxylase inhibitor L-alpha-difluoromethylornithine sensitized all of these leukemia cells to the deleterious effects of the inhibitor by stimulating its intracellular accumulation. The lysosomotropic nature of select polyamine analogues may, thus, provide a novel chemotherapeutic strategy to selectively induce apoptosis of malignant hematopoietic cells. PMID- 10519409 TI - Modified vaccinia virus Ankara for delivery of human tyrosinase as melanoma associated antigen: induction of tyrosinase- and melanoma-specific human leukocyte antigen A*0201-restricted cytotoxic T cells in vitro and in vivo. AB - Vaccination with tumor-associated antigens is a promising approach for cancer immunotherapy. Because the majority of these antigens are normal self antigens, they may require suitable delivery systems to promote their immunogenicity. A recombinant vector based on the modified vaccinia virus Ankara (MVA) was used for expression of human tyrosinase, a melanoma-specific differentiation antigen, and evaluated for its efficacy as an antitumor vaccine. Stable recombinant viruses (MVA-hTyr) were constructed that have deleted the selection marker lacZ and efficiently expressed human tyrosinase in primary human cells and cell lines. Tyrosinase-specific human CTLs were activated in vitro by MVA-hTyr-infected, HLA A*0201-positive human dendritic cells. Importantly, an efficient tyrosinase- and melanoma-specific CTL response was induced in vitro using MVA-hTyr-infected autologous dendritic cells as activators for peripheral blood mononuclear cells derived from HLA-A*0201-positive melanoma patients despite prior vaccination against smallpox. Immunization of HLA-A*0201/Kb transgenic mice with MVA-hTyr induced A*0201-restricted CTLs specific for the human tyrosinase-derived peptide epitope 369-377. These in vivo primed CTLs were of sufficiently high avidity to recognize and lyse human melanoma cells, which present the endogenously processed tyrosinase peptide in the context of A*0201. Tyrosinase-specific CTL responses were significantly augmented by repeated vaccination with MVA-hTyr. These findings demonstrate that HLA-restricted CTLs specific for human tumor-associated antigens can be efficiently generated by immunization with recombinant MVA vaccines. The results are an essential basis for MVA-based vaccination trials in cancer patients. PMID- 10519410 TI - Potent activity of soluble B7-IgG fusion proteins in therapy of established tumors and as vaccine adjuvant. AB - Fusion proteins consisting of the extracellular region of murine B7.1 or B7.2 and the Fc portion of murine IgG2a (B7-IgG) were evaluated for their ability to promote antitumor responses. Therapeutic administration of soluble B7-IgG in mice with established tumors induced complete regression of the tumor and increased the survival of mice. In three models, MethA, P815, and MB49, mice with 7-day-old established tumors were cured with two to three treatment cycles of B7-IgG, given twice a week. Even in mice with an established B16/F10 tumor (a poorly immunogenic melanoma), therapeutic treatment with B7-IgG alone slowed tumor growth and increased survival significantly. Still stronger antitumor activity was achieved when B7-IgG was used as a vaccine adjuvant mixed with irradiated tumor cells. In 80% of mice with 7-day-old B16 tumors, tumors regressed completely, and mice survived for at least 80 days. In all tumor models, B7.1-IgG and B7.2-IgG had similar antitumor activity. B7-IgG-mediated tumor rejection was dependent on T cells, specifically CD8 cells, as demonstrated by the failure of B7-IgG to induce tumor regression in severe combined immunodeficient or CD8 depleted mice. In addition, mice that were cured of an established tumor were protected against a rechallenge with the same tumor for at least 4 months, suggesting the generation of memory responses. Surprisingly, the antitumor activity of B7-IgG was independent of IFN-gamma, as demonstrated by tumor rejection in IFN-gamma knockout mice. Our findings demonstrate the potent capacity of B7-IgG to generate or enhance antitumor immune responses and suggest the clinical value of B7-IgG. PMID- 10519411 TI - Expression of OVCA1, a candidate tumor suppressor, is reduced in tumors and inhibits growth of ovarian cancer cells. AB - Loss of all or part of one copy of chromosome 17p is very common in ovarian and breast tumors. OVCA1 is a candidate tumor suppressor gene mapping to a highly conserved region on chromosome 17p13.3 that shows frequent loss of heterozygosity in breast and ovarian carcinomas. Western blot analysis of extracts prepared from breast and ovarian carcinomas revealed reduced expression of OVCA1 compared with extracts from normal epithelial cells from these tissues. Subcellular localization studies indicate that OVCA1 is localized to punctate bodies scattered throughout the cell but is primarily clustered around the nucleus. Attempts to create cell lines that stably expressed OVCA1 from the cytomegalovirus promoter were generally unsuccessful in a variety of different cell lines. This reduction of colony formation was quantified in the ovarian cancer cell line A2780, where it was demonstrated that cells transfected with plasmids expressing OVCA1 had a 50-60% reduction in colony number as compared with appropriate controls, and only a few of these clones expressed OVCA1, albeit at low levels. The clones that expressed exogenous OVCA1 were found to have dramatically reduced rates of proliferation. Reduced growth rates correlated with an increased proportion of the cells in the G1 fraction of the cell cycle compared with the parental cell line and decreased levels of cyclin D1. The low levels of cyclin D1 appeared to be caused by an accelerated rate of cyclin D1 degradation. Overexpression of cyclin D1 was able to override OVCA1's suppression of clonal outgrowth. These results suggest that slight alterations in the level of OVCA1, such as would occur after reduction of chromosome 17p13.13 to hemizygosity, may result in cell cycle deregulation and promote tumorigenesis. PMID- 10519412 TI - Kaposi's sarcoma-associated herpesvirus-encoded v-cyclin triggers apoptosis in cells with high levels of cyclin-dependent kinase 6. AB - Kaposi's sarcoma-associated herpesvirus (KSHV) has a key etiological role in development of Kaposi's sarcoma (KS). v-Cyclin is a KSHV-encoded homologue to D type cyclins that associates with cellular cyclin-dependent kinase 6 (CDK6). v Cyclin promotes S-phase entry of quiescent cells and has been suggested to execute functions of both D- and E-type cyclins. In this study, expression of v cyclin in cells with elevated levels of CDK6 led to apoptotic cell death after the cells entered S phase. The cell death required the kinase activity of CDK6 because cells expressing a kinase-deficient form of CDK6 did not undergo apoptosis upon v-cyclin expression. Studies on the mechanisms involved in this caspase-3-mediated apoptosis indicated that it was independent of cellular p53 or pRb status, and it was not suppressed by Bcl-2. In contrast, the KSHV-encoded v Bcl-2 efficiently suppressed v-cyclin-/CDK6-induced apoptosis, demonstrating a marked difference in the antiapoptotic properties of c-Bcl-2 and v-Bcl-2. In KS lesions, high CDK6 expression was confined to a subset of cells, some of which displayed signs of apoptosis. These results suggest that v-cyclin may exert both growth-promoting and apoptotic functions in KS, depending on factors regulating CDK6 and v-Bcl-2 levels. PMID- 10519413 TI - Identification and characterization of genes associated with human hepatocellular carcinogenesis. AB - Eight cDNAs encoding galectin 4 (Gal-4), UGT2B4 (UDP-glucuronosyltransferase), ribosomal phosphoprotein P0 (rpP0), dek, insulin-like growth factor binding protein (IGFBP) 1, vitronectin, retinoic acid-induced gene E (RIG-E), and CYP3A4 (cytochrome P450 nifedipine oxidase) were identified as differentially expressed genes between human hepatocellular carcinoma (HCC) and matched nontumorous liver tissues. Higher levels of UGT2B4, rpP0, dek, vitronectin, Gal-4, and IGFBP-1 mRNAs combined with a lower level of RIG-E mRNA were observed in at least four of five primary HCCs compared to matched nontumorous liver tissues. Furthermore, a pathological study suggested that the levels of UGT2B4, rpP0, dek, and vitronectin increased and the level of RIG-E decreased with the histological grading. On the other hand, the expression of CYP3A4 mRNA and CYP3A7 (P-450 Fla) mRNA, a transcript found in the fetus and highly homologous to CYP3A4, was higher in all nontumorous liver and some of the carcinoma tissues from five HCC patients, whereas it was significantly lower in normal liver tissues from two non HCC patients. The examination using HCC cell lines HuH-7 and HepG2 under different growth conditions suggested that the expression of dek mRNA was growth associated. In contrast, the expression of Gal-4, UGT2B4, IGFBP-1, and RIG-E mRNAs was regulated in a cell density-dependent manner: the levels of Gal-4, UGT2B4, and IGFBP-1 were undetectably low, whereas the level of RIG-E was high in rapidly proliferating, subconfluent HCC cells in 10% serum; however, the expression levels were reversed in dense, overcrowded cultures. In addition, IGFBP-1 and Gal-4 mRNAs were also induced by reducing the serum concentration to 0.1%. We also demonstrated that sodium butyrate, an inducer of differentiation, up-regulated and down-regulated RIG-E and dek mRNAs, respectively, in a dose dependent manner in HuH-7 cells, supporting, in part, our pathological observation. In summary, therefore, high expression of Gal-4, UGT2B4, rpP0, dek, IGFBP-1, and vitronectin, together with low expression of RIG-E, was correlated with the malignant potential of HCC. CYP3A4 and CYP3A7 could be induced in HCC bearing livers. These transcripts are differentially regulated depending on cell cell contact, serum growth factors, growth and differentiation status, and/or other mechanisms in premalignant and malignant liver cells. PMID- 10519414 TI - Genomic structure, chromosomal mapping, and promoter region analysis of murine uridine phosphorylase gene. AB - Uridine phosphorylase (UPase) plays an important role in the activation of 5 fluorouracil and in the regulation of tissue and plasma concentration of uridine, a potential biochemical modulator of 5-fluorouracil therapy. UPase expression is affected by the c-H-ras oncogene and various cytokines through unknown mechanisms. To understand its expression and regulation, we cloned the murine UPase gene, defined its genomic organization, determined its 5'- and 3'-end flanking sequences, and evaluated the promoter activity. The UPase gene contains nine exons and eight introns, spanning a total of approximately 18.0 kb. Its promoter lacks canonical TATA and CCAAT boxes, although a CAATAAAAA TATA-like box is seen from -41 to -49. Furthermore, IFN regulatory factor 1, c/v-Myb, and p53 binding sites are present in the promoter region, indicating that UPase expression may be directly regulated by cytokines and oncogene products. The 1.2 kb flanking fragment showed promoter activity driving the expression of the luciferase gene in various mammalian cells. A TGGGG repeat sequence is seen in the 3'-end flanking region. This element is considered to be a potential recombination consensus hot spot that may contribute to the encoding of different UPase isoforms present in different tissues, both normal and neoplastic. PMID- 10519415 TI - Carcinoma-associated fibroblasts direct tumor progression of initiated human prostatic epithelium. AB - The present study demonstrates that fibroblasts associated with carcinomas stimulate tumor progression of initiated nontumorigenic epithelial cells both in an in vivo tissue recombination system and in an in vitro coculture system. Human prostatic carcinoma-associated fibroblasts grown with initiated human prostatic epithelial cells dramatically stimulated growth and altered histology of the epithelial population. This effect was not detected when normal prostatic fibroblasts were grown with the initiated epithelial cells under the same experimental conditions. In contrast, carcinoma-associated fibroblasts did not affect growth of normal human prostatic epithelial cells under identical conditions. From these data, we conclude that in this human prostate cancer model, carcinoma-associated fibroblasts stimulate progression of tumorigenesis. Thus, carcinoma-associated fibroblasts can direct tumor progression of an initiated prostate epithelial cell. PMID- 10519416 TI - In vivo prediction of vascular susceptibility to vascular susceptibility endothelial growth factor withdrawal: magnetic resonance imaging of C6 rat glioma in nude mice. AB - One of the hallmarks of tumor neovasculature is the prevalence of immature vessels manifested by the low degree of recruitment of vascular mural cells such as pericytes and smooth muscle cells. This difference in the architecture of the vascular bed provides an important therapeutic window for inflicting tumor selective vascular damage. Here we demonstrate the application of gradient echo magnetic resonance imaging (MRI) for noninvasive in vivo mapping of vascular maturation, manifested by the ability of mature vessels to dilate in response to elevated levels of CO2. Histological alpha-actin staining showed a match between dilating vessels detected by MRI and vessels coated with smooth muscle cells. Switchable, vascular endothelial growth factor (VEGF)-overexpressing tumors (C6 pTET-VEGF rat glioma s.c. tumors in nude mice) displayed high vascular function and significant vascular damage upon VEGF withdrawal. However, damage was restricted to nondilating vessels, whereas mature dilating tumor vessels were resistant to VEGF withdrawal. Thus, MRI provides in vivo visualization of vascular maturity and prognosis of vascular obliteration induced by VEGF withdrawal. PMID- 10519417 TI - Paradoxical inhibition of c-myc-induced carcinogenesis by Bcl-2 in transgenic mice. AB - Here, we investigated changes in apoptosis during tumor progression by analyzing the effect of coexpressing various antiapoptotic genes on the multistage process of c-myc-induced hepatocarcinogenesis in transgenic mice. Whereas continuous c myc gene overexpression in the liver led to cellular hepatocarcinoma, the coexpression of the bcl-2 gene inhibited the emergence of liver tumors, by inhibiting a pretumoral phase characterized by increased proliferation and apoptosis. This antioncogenic effect was specific to Bcl-2 and was not shared by other antiapoptotic genes such as bcl-xL and a dominant negative form of p53. Thus, we have shown that Bcl-2 can have a tumor suppressor effect in vivo on c myc-induced hepatocarcinogenesis during the emergence of neoplastic foci. PMID- 10519418 TI - Overexpression of fibroblast growth factor 1 in MCF-7 breast cancer cells facilitates tumor cell dissemination but does not support the development of macrometastases in the lungs or lymph nodes. AB - Mice bearing primary tumors produced by LacZ-tagged MCF-7 human breast carcinoma cells transfected with fibroblast growth factor (FGF) 1 have frequent micrometastases, but macrometastases are not observed. i.v. injection of FGF-1 transfected tumor cells produced no pulmonary macrometastases, and removal of primary tumors resulted in the disappearance of spontaneous micrometastases. Thus, failure of micrometastases to proliferate was not due to inhibitory factors released from the primary tumor, and the presence of the primary tumor is required for maintenance of the micrometastases. This indicates that the micrometastases result from continued seeding from the primary tumor balanced by clearance from the metastatic site. Tumor emboli trapped in the vessels of lungs and lymph nodes and single tumor cells observed in the pulmonary vein implied that FGF-1-overexpressing MCF-7 cells are deficient in their ability to extravasate. The frequency of tumor cells incorporating bromodeoxyuridine was consistently lower in lung tissues when compared with primary tumors, indicating that disseminated tumor cells were unable to maintain a high rate of proliferation. Increased angiogenesis resulting from FGF-1 production by the transfected cells with a concomitant increased rate of intravasation into developing blood vessels may be the underlying determinant of spontaneous micrometastasis produced by these cells when compared with parental MCF-7 cells. PMID- 10519419 TI - Evidence for clonal outgrowth of androgen-independent prostate cancer cells from androgen-dependent tumors through a two-step process. AB - Prostate cancers require androgen for growth but progress to an androgen independent stage under the selective pressure of androgen ablation therapy. Here we describe a novel human prostate cancer xenograft (LAPC-9) propagated by serial passage in male severe combined immunodeficient mice that expresses prostate specific antigen and wild-type androgen receptor. In response to castration, LAPC 9 cells undergo growth arrest and persist in a dormant, androgen-responsive state for at least 6 months. After prolonged periods of androgen deprivation, spontaneous androgen-independent outgrowths develop. Thus, prostate cancers progress to androgen independence through two distinct stages, initially escaping dependence on androgen for survival and, subsequently, for growth. Through the use of serial dilution and fluctuation analysis, we provide evidence that the latter stage of androgen independence results from clonal expansion of androgen independent cells that are present at a frequency of about 1 per 10(5)-10(6) androgen-dependent cells. We conclude that prostate cancers contain heterogeneous mixtures of cells that vary in their dependence on androgen for growth and survival and that treatment with antiandrogen therapy provides selective pressure and alters the relative frequency of these cells, thereby leading to outgrowths of androgen-independent cancers. PMID- 10519420 TI - Thymidine phosphorylase expression is associated with both increase of intratumoral microvessels and decrease of apoptosis in human colorectal carcinomas. AB - Thymidine phosphorylase (dThdPase)/platelet-derived endothelial cell growth factor is expressed at higher levels in a variety of human carcinomas than it is in adjacent normal tissue. The higher expression is associated with an increase of intratumoral microvessel density (IMVD) and an unfavorable patient prognosis. We examined the role of dThdPase in apoptosis, cell proliferation, IMVD, and p53 expression in human colorectal carcinomas. dThdPase expression was noted in 13 of 36 (36.1%) Dukes' A and B carcinomas and in 13 of 28 (46.3%) Dukes' C and D carcinomas. At least 10 areas consisting of carcinoma cells with diffuse dThdPase expression from the 26 dThdPase-positive tumors (category I) and 10 areas without dThdPase expression from the 38 negative tumors (category II) were selected from each case. For stage A and B tumors, the mean IMVDs were 64.8 +/- 33.7 in category I and 33.2 +/- 12.6 in category II tumors, whereas for stage C and D tumors, the mean IMVDs were 77.6 +/- 27.2 in the category I and 34.7 +/- 14.0 in the category II tumors. The mean IMVD was significantly higher in category I than in category II tumors (P < 0.01). The mean apoptotic indices (AIs; percentage of apoptotic cells) were 2.7 +/- 1.7 in the category I and 5.4 +/- 2.2 in the category II carcinomas of stages A and B and 1.4 +/- 0.5 in category I and 5.3 +/ 2.3 in category II carcinomas of stages C and D, and the value of the mean AI was significantly lower in category I than in category II (P < 0.01), regardless of the Dukes' stage. AI and IMVD showed a significant inverse correlation (P < 0.001). There was no significant difference in the frequency of p53 expression between the two categories. These results indicated that dThdPase expression provides an advantage for tumor growth of human colonic carcinomas not only by increasing the intratumoral microvessels but also by attenuation of apoptosis, which might occur via a p53 gene-independent pathway. PMID- 10519421 TI - A soluble transforming growth factor beta type III receptor suppresses tumorigenicity and metastasis of human breast cancer MDA-MB-231 cells. AB - Transforming growth factor beta (TGF-beta) can promote late stage tumor progression in a number of model systems. In the present study, we have examined whether expression of a truncated soluble extracellular domain of TGF-beta type III receptor (sRIII) in human breast cancer MDA-MB-231 cells can antagonize the tumor-promoting activity of TGF-beta by sequestering active TGF-beta isoforms that are produced by the cancer cells. The secretion of sRIII reduced the amount of active TGF-beta1 and TGF-beta2 in the conditioned medium. This led to a significant reduction of the growth-inhibitory activity of the medium conditioned by sRIII-expressing cells on the growth of mink lung epithelial CCL64 cells in comparison with the medium conditioned by the control cells. The tumor incidence and growth rate of all of the three sRIII-expressing clones studied were significantly lower than those of the control cells in athymic nude mice. Four of five control cell-inoculated mice showed spontaneous metastasis in the lung, whereas none of the sRIII-expressing cell-inoculated mice had any lung metastasis. Thus, our results suggest that the sRIII may be used to antagonize the tumor-promoting activity of TGF-beta. PMID- 10519422 TI - Induction of apoptosis by arachidonic acid in chronic myeloid leukemia cells. AB - The hallmark of chronic myeloid leukemia (CML) is the presence of the bcr-abl oncogene, which is associated with transforming ability and an intrinsic resistance to induction of apoptosis by genotoxic agents. Arachidonic acid (AA), a biologically active fatty acid, plays a crucial role as a mediator of signaling pathways involved in cell proliferation and survival. In this study, we investigated the potential role of AA as a proapoptotic agent in CML. Pretreatment of human CML isolated progenitor cells with AA (100 microM for 18 h) induced 71-75% inhibition of in vitro colony formation of granulocyte-macrophage colony-forming units, multilineage colony-forming units, and erythroid burst forming units. This inhibition was significantly greater than the effect on normal progenitor cells (19-39% growth inhibition of erythroid burst-forming units, multilineage colony-forming units, and granulocyte-macrophage colony forming units). AA also inhibited growth of the bcr-abl-transformed cell line H7.bcr-abl A54. In contrast, a minimal effect of AA on inhibition of cell growth was observed in the parental nontransformed NSF/N1.H7 cell line. The antiproliferative effect of AA was associated with apoptosis. Gamma-linolenic acid, a precursor of AA, also inhibited cell growth, whereas other unsaturated and saturated fatty acids had no effect. Pharmacological inhibition of cyclooxygenase, lipooxygenase, and cytochrome P450 monooxygenase enzymes prior to exposure to AA did not rescue cells from the inhibitory effect of AA. Moreover, 5,8,11,14-eicosatetraynoic acid, a nonmetabolizable arachidonate analogue, also inhibited cell growth, suggesting that the effect of AA did not require further metabolism. Treatment with antioxidants prior to stimulation with AA was also ineffective in preventing its antiproliferative effect. Thus, AA inhibited proliferation of CML cells by inducing apoptotic cell death. The signaling mechanisms of AA-induced inhibition of cell growth appeared to be independent of its conversion into eicosanoids or free radical generation. PMID- 10519423 TI - Could tight garments cause endometriosis? PMID- 10519424 TI - Hormonal prevention of breast cancer: proposal for a change in paradigm. PMID- 10519426 TI - Non-invasive RNA-based determination of fetal Rhesus D type: a prospective study based on 96 pregnancies. AB - OBJECTIVES: To develop a non-invasive method for determining fetal RhD status in order to provide improved care for women most at risk. DESIGN: A prospective study. METHODS: Fetal erythroblasts were enriched from the peripheral circulation of 96 RhD negative women with pregnancies at various stages in gestation using discontinuous density gradients. Amplification of RhD-specific mRNAs was carried out by reverse transcription-polymerase chain reaction assay. RNA, rather than DNA, was selected for amplification because it rarely contaminates samples, thus resulting in fewer false positives; moreover, its presence in multiple copies per cell should enhance the sensitivity of the assay, resulting in fewer false negatives. The study was prospective, relying on postnatal serological confirmation of RhD phenotype. RESULTS: The assay was 75% accurate at predicting fetal RhD status, comparing favourably with standard genomic DNA-based assays. However, we found that accuracy dropped from 85% (29/34) in the third trimester of pregnancy, to 82% (32/39) in the second and 48% (11/23) in the first trimester. Discordant data were due to false negatives in the majority (78%) of cases. CONCLUSIONS: We suggest that reverse transcription may be a useful and perhaps more sensitive alternative to standard genomic polymerase chain reaction in the majority of cases. However, under certain circumstances the absence or reduction of fetal erythroblasts or possibly RhD mRNA in some preparations may compromise the accuracy of the assay. PMID- 10519425 TI - Selective deficit of angiogenic growth factors characterises pregnancies complicated by pre-eclampsia. AB - OBJECTIVE: To compare serum levels of angiogenic growth factors vascular endothelial growth factor (VEGF), placental growth factor (PlGF) and angiogenin in pre-eclamptic women and matched controls. DESIGN: Retrospective analysis of 70 degrees C stored serum of women who developed pre-eclampsia and matched controls. SETTING: Department of Gynaecology and Obstetrics, St Elisabeth Hospital, Curacao, Netherlands Antilles. SAMPLE: Thirty women with pre-eclampsia and 30 normotensive controls matched for age and gestation. RESULTS: VEGF and PIGF serum levels were significantly lower in pre-eclamptic pregnancies, compared with controls (VEGF 0.31 +/- 1.20 vs 18.30 +/- 24.97 pg/mL, P = 0.0004; PlGF 54.19 +/- 32.05 vs 497.95 +/- 340.51 pg/mL, P < 0.0001). Matched couple analysis showed VEGF serum concentrations to be lower in the majority of pre-eclamptic women and PlGF concentrations to be lower in all pre-eclamptic women. Angiogenin serum levels showed no statistical significant difference between pre-eclamptic pregnancies and controls (523.68 +/- 367.55 vs 670.41 +/- 251.54 ng/mL, P = 0.058), with matched couple analysis showing no clear pattern. CONCLUSIONS: Decreased serum levels of VEGF and PIGF characterise, and therefore seem to be of importance during (the development of), pre-eclampsia. This selective deficit of angiogenic growth factors might in part explain the shallow placentation found in this pregnancy complication. PMID- 10519427 TI - Screening for fetal aneuploidies and fetal cardiac abnormalities by nuchal translucency thickness measurement at 10-14 weeks of gestation as part of routine antenatal care in an unselected population. AB - OBJECTIVES: To evaluate first trimester pregnancy screening for fetal aneuploidy and congenital heart defects by maternal age and nuchal translucency measurement and screening for fetal aneuploidies and congenital heart defects by ultrasound in an unselected population. DESIGN: A prospective study. SETTING: Fetal medicine unit, St George's Hospital, London. SAMPLE: 4523 consecutive viable fetuses at 10 14 weeks with a crown-rump length between 38 and 80 mm were scanned transabdominally (93%) or transvaginally (7%). METHODS: Screening was performed by calculating the background risk from maternal age, gestational age and obstetric history, which was then adjusted with the nuchal translucency measurement in relation to crown-rump length (adjusted risk). MAIN OUTCOME MEASURES: Measurements of crown-rump length and nuchal translucency thickness. An adjusted risk of > 1:270 was considered as a positive screening test. Pregnancy outcome was obtained through karyotyping, outcome questionnaires and examination of the newborn infants. RESULTS: Mean maternal age was 29-4 years and mean gestational age 12.2 weeks. Screening was positive in 230/4523 fetuses (5.1%), when the adjusted risk (mean 1:2649) was > 1:270. Fetal karyotype was abnormal in 23 (0.51%) cases, including twelve with trisomy 21, five trisomy 18, one trisomy 13, one trisomy 10, one monosomy X and two triploidies. For a false positive rate of 4.7%, the sensitivity of this test was 78% in detecting any fetal aneuploidy. Only one out of nine major congenital heart defects in this population was found within the 110 euploid fetuses with increased nuchal translucency thickness (> 2.5 mm). CONCLUSION: Screening for fetal aneuploidy by maternal age and nuchal translucency measurement can be effective in an unselected population. However, our results do not support its effectiveness in the detection of cardiac abnormalities. PMID- 10519428 TI - Fetal karyotyping by chorionic villus sampling after the first trimester. AB - OBJECTIVE: To evaluate chorionic villus sampling (CVS) as a technique for karyotyping after the first trimester by examining the incidence of result failure, confined placental mosaicism, and false positive or negative results at different gestational ages. METHODS: During a nine year period between 1989 and 1997, all results of CVS between 8 and 37 weeks of gestation provided by the Regional Cytogenetics Centre were analysed retrospectively by examining indications for CVS, weights of tissue received, gestational age at sampling and karyotype results. RESULTS: There were 2424 chorionic villus samples analysed by the direct method and/or cell culture. In 1548 cases CVS was performed before 14 weeks (Group 1), in 685 between 15 and 20 weeks (Group 2), in 160 between 21 and 28 weeks (Group 3) and in 31 cases after 29 weeks (Group 4). Although there was a trend for an increasing rate of failed direct preparation results from Groups 1 to 4 which were 3.8%, 4.7%, 5.6% and 6.6%, respectively; these results were not significantly different. There were 19 cases of confined placental mosaicism and the incidence was significantly greater in Group 3 compared with Group 1 (P < 0.05), and in Groups 3 and 4 combined compared with Group 1. There were six false positive and one false negative result following direct analysis with no significant differences between gestationar ages. CONCLUSIONS: CVS is a useful test after the first trimester, especially when a fast result is clinically required. However, after 20 weeks, when cordocentesis is available, the higher rate of cytogenetic discordancy between the placenta and the fetus means that cordocentesis may be preferable. PMID- 10519429 TI - Elevated mid-trimester maternal corticotrophin-releasing hormone levels in pregnancies that delivered before 34 weeks. AB - OBJECTIVE: To test whether maternal corticotrophin-releasing hormone levels are elevated in the mid- trimester for those women who subsequently had spontaneous preterm delivery and to assess the clinical utility of the measurement in the prediction of preterm delivery. DESIGN: A prospective observational study. SETTING: Department of Obstetrics and Gynaecology, Prince of Wales Hospital, Hong Kong. POPULATION: 1047 low risk pregnant women recruited at 15-20 weeks of gestation. METHODS: Venous samples were assayed for levels of corticotrophin releasing hormone. The investigators responsible for the laboratory assay were blinded to the obstetric outcome. MAIN OUTCOME MEASURES: Incidence of preterm, term and post-term pregnancies. RESULTS: Those who were delivered spontaneously at a preterm gestational age (before 34 weeks) had significantly higher corticotrophin-releasing hormone levels in the mid-trimester, compared with those who were delivered at term or post-term. There was a trend towards lower corticotrophin-releasing hormone levels with more advanced gestational age at delivery. When the measurement of corticotrophin- releasing hormone was used to predict delivery before 34 weeks, the best cut off was 1.9 MoM, which produced a sensitivity of 72.7% and specificity of 78.4%. This translated to a positive predictive value of 3.6%, negative predictive value of 99.6% and relative risk of 9.4 when the background prevalence of spontaneous preterm delivery before 34 weeks was 1.1%. The likelihood ratio was 3.4. CONCLUSIONS: Mid-trimester maternal corticotrophin-releasing hormone levels are elevated in pregnancies destined to deliver preterm before 34 weeks. When used alone in a low risk population, the measurement has a low predictive power for preterm delivery. However, the likelihood ratio of 3 4 implies that in high risk populations the test may be considerably more valuable. PMID- 10519430 TI - Receptor binding of oxytocin and vasopressin antagonists and inhibitory effects on isolated myometrium from preterm and term pregnant women. AB - OBJECTIVE: To test binding affinities for, and inhibitory effects on, myometrium of some oxytocin and vasopressin antagonists with respect to their therapeutic potential. DESIGN: Receptor binding studies on transfected cell lines. In vitro contractility studies of human myometrium. SETTING: The Research Laboratory of Sanofi Recherche, Centre de Toulouse, France and the Departments of Obstetrics and Gynecology, Lund University Hospital, Sweden and Bialystok University Hospital, Poland. PARTICIPANTS: Nine women delivered by caesarean section preterm and 37 delivered at term for routine obstetric indications. INTERVENTIONS: The binding affinities of oxytocin, arginine vasopressin, atosiban (1-deamino-2-D Tyr(OEt)-4-Thr-8-Om-oxytocin), SR 49059 and SR 121463 for the human oxytocin and different subtypes of vasopressin receptors were determined. Concentration response curves with oxytocin and arginine vasopressin were recorded on myometrium from preterm- and term-delivered women in control experiments and in the presence of 2.5 and 10 nmol/L of SR 49059. Furthermore, using term myometrium, the influence of SR 49059 and SR 121463 in concentrations of 3, 10, 30 and 100 nmol/L on responses to the EC50 concentrations of oxytocin and vasopressin were compared. MAIN OUTCOME MEASURES: Receptor binding affinities. In vitro contractile effects and their inhibitions. RESULTS: Oxytocin had a high affinity for the oxytocin receptor (K(i) in mean = 6.8 nmol/L) and bound, to some extent, to the vasopressin V1a receptor (K(i) = 34.9 nmol/L). Vasopressin displayed higher affinities for vasopressin V1a, V1b and V2 receptors (K(i) = 1.4, 0.8 and 4.2 nmol/L, respectively) than for the oxytocin receptor (K(i) = 48 nmol/L). Atosiban and SR 49059 both had a high affinity for the vasopressin V1a receptor (K(i) = 4.7 and 7.2 nmol/L, respectively, and a moderate one for the oxytocin receptor (K(i) = 397 and 340 nmol/L, respectively). SR 121463 exerted a predominant binding to the V2 receptor (K(i) = 3.0 nmol/L). In the concentration response experiments levels of up to 10 nmol/L of SR 49059 had no influence on the effect of oxytocin on myometrium from women preterm and at term pregnancy. However, a concentration-dependent inhibition of the responses of both these type of tissues to vasopressin was seen. The effects of EC50 concentrations of oxytocin and vasopressin on term pregnant myometrium were markedly inhibited by 10 nmol/L and higher concentrations of SR 49059, the inhibition of the response to vasopressin being more pronounced than that of the oxytocin response. SR 121463 at maximal concentration only caused slight inhibitions of the oxytocin and vasopressin responses. CONCLUSIONS: Atosiban and SR 49059 both have moderate binding affinities for the human oxytocin receptor and high binding affinities for the vasopressin V1a one. We demonstrated that SR 49059 inhibits the response of term myometrium to oxytocin and that of both preterm and term myometrium to vasopressin. These observations suggest a therapeutic potential of SR 49059 in preterm labour. The vasopressin V2 receptor is apparently not involved to any significant degree in the activation of the pregnant human uterus. PMID- 10519431 TI - Leptin concentrations in maternal serum and cord blood: relationship to maternal anthropometry and fetal growth. AB - OBJECTIVE: To determine 1. the relationship between maternal serum leptin concentrations and maternal anthropometry and 2. the relationship between cord serum leptin concentrations at birth and neonatal anthropometry. DESIGN: Prospective cohort study of fetal growth in low-risk pregnancies. SETTING: University teaching hospital. SAMPLE: Thirty-nine women and their babies taking part in a fetal growth study. METHODS: Blood was taken from the women between 10 20 weeks of gestation and from the umbilical cord of their babies at delivery. Serum leptin was measured by radio-immunoassay. Maternal anthropometric measurements were recorded at booking. Neonatal anthropometric measurements were recorded within 48 hours after delivery. Linear regression analysis was used to explore the relationship between serum leptin concentrations and anthropometric measures and multiple regression analysis then applied to determine which variables remained independently associated with leptin. RESULTS: The median (range) leptin concentration in maternal serum was 11.8 ng/mL (1.7-39.7) and in cord blood was 4.2 ng/mL (0.6-21.4). Maternal leptin levels correlated with maternal weight, body mass index, midarm circumference and skinfold thickness, but not with birthweight, placental weight or maternal height. Body mass index and midarm circumference remained significant after multiple regression analysis. Cord leptin levels correlated with birthweight, birthlength, placental weight and skinfold thickness but not with ponderal index. Birthweight and subscapular skinfold thickness remained significant after multiple regression analysis. Cord leptin concentrations did not correlate with maternal leptin concentrations. CONCLUSIONS: We suggest that there are very strong associations between maternal leptin and maternal adiposity in pregnancy, and between cord leptin at delivery and birthweight, as well as other anthropometric markers of fetal growth. PMID- 10519432 TI - Activin A, inhibin A, inhibin B and parturition: changes of maternal and cord serum levels according to the mode of delivery. AB - OBJECTIVE: To evaluate whether activin A, inhibin A, and inhibin B levels in maternal and umbilical artery serum change according to the mode of delivery. DESIGN: Maternal and cord blood specimens were collected at term after spontaneous labour and vaginal delivery, or elective caesarean section. SETTING: Universities of Pisa, Turin, Naples and Udine. POPULATION: Forty-two healthy pregnant women, at 3940 weeks of gestation, divided into two subgroups: group 1 vaginal delivery (n = 21), were delivered of 10 female and 11 male infants; group 2 elective caesarean section (n = 21), were delivered of 11 female and 10 male infants. MAIN OUTCOME MEASURES: Serum activin A, inhibin A, inhibin B concentrations in maternal and umbilical cord blood. RESULTS: At vaginal delivery, maternal serum inhibin A and inhibin B levels were lower and activin A levels higher than at elective caesarean section. Maternal levels of activin A, inhibin A and inhibin B were constantly higher than in umbilical arterial blood, independent of the mode of delivery. No significant difference was observed in umbilical arterial serum levels of the three proteins between the two modes of delivery. Umbilical arterial serum activin A and inhibin A concentrations did not show a significant difference between male and female infants in either vaginal or caesarean section, but male infants showed inhibin B levels significantly higher than female, independent of the mode of delivery. CONCLUSIONS: In the presence of active labour, the human placenta secretes larger amounts of activin A and lesser amounts of inhibin A and inhibin B into the maternal circulation. Inhibin-related proteins in the fetal circulation do not show differences according to the mode of delivery, suggesting that they have a different method of production or metabolic rate compared with maternal activin and inhibins. PMID- 10519433 TI - Misoprostol compared with methylergometrine for the prevention of postpartum haemorrhage: a double-blind randomised trial. AB - OBJECTIVE: To compare the efficacy and side effects of misoprostol, compared with methylergometrine, for the prevention of postpartum haemorrhage. DESIGN: A double blind, randomised clinical trial of 200 women with apparently normal pregnancies. SETTING: University teaching hospital. PARTICIPANTS: Two hundred women with apparently normal pregnancies. METHODS: After the baby had been born, all women received two capsules by mouth and the contents of an ampule by intravenous injection. Each woman only received one active product. The capsules contained either a total of 600 microg misoprostol or placebo, and the ampule 200 microg of methylergometrine or placebo. MAIN OUTCOME MEASURES: Need for further oxytocic drugs, blood pressure, the presence of side effects, mean haemoglobin and haematocrit three days after delivery. RESULTS: Two hundred women completed the study (100 received methylergometrine and 100 misoprostol). Postpartum haemorrhage occurred in 4.3% of the methylergometrine group and 8.3% of the misoprostol group (P = 0.57). The need for further oxytocic drugs was 4.4% and 12.8% after methylergometrine and misoprostol, respectively (P = 0.065). One hour after the birth of the baby there was no difference in the mean systolic blood pressure (117 +/- 12 mmHg versus 115 +/- 11 mmHg) (P = 0.26) or the mean diastolic blood pressure (72 +/- 10 mmHg versus 71 +/- 11 mmHg for the groups receiving methylergometrine or misoprostol, respectively) (P = 0.97). The mean temperature in the misoprostol group rose to 37.4 degrees C, compared with 37 degrees C in the methylergometrine group (P < 0.0001). In the misoprostol group 34% developed fever (> 38 degrees C) compared with 3% in the methylergometrine group (P < 0.0001). Shivering (visual analogue score > or = 8) also occurred more often after misoprostol (42%) than after methylergometrine (8.5%) (P < 0.0001). The haemoglobin level (g/dL) on the third postpartum day was similar for both groups ( 11.0 and 11.2 for methylergometrine and misoprostol, respectively) (P = 0.39). CONCLUSIONS: This study suggests that although protection from postpartum haemorrhage using parenteral methylergometrine and oral misoprostol is nearly equal, misoprostol is associated with more side effects. PMID- 10519434 TI - High resolution imaging of endometriosis and ovarian carcinoma with optical coherence tomography: feasibility for laparoscopic-based imaging. AB - High resolution imaging of gynaecological tissue offers the potential for identifying pathological changes at early stages when interventions are more effective. Optical coherence tomography (OCT) is a high resolution high speed optical imaging technology which is analogous to ultrasound B-mode imaging except reflections of light are detected rather than sound. The OCT technology is capable of being integrated with laparoscopy for real-time subsurface imaging. In this report, the feasibility of OCT for differentiating normal and pathologic laparoscopically-accessible gynaecologic tissue is demonstrated. Differentiation is based on architectural changes of in vitro tissue morphology. OCT has the potential to improve conventional laparoscopy by enabling subsurface imaging near the level of histopathology. PMID- 10519435 TI - Neural networks in the diagnosis of malignant ovarian tumours. AB - OBJECTIVE: To assess the role of neural networks in predicting the likelihood of malignancy in women presenting with ovarian tumours. DESIGN: Retrospective case study. SETTING: University Department of Obstetrics and Gynaecology, St James's Hospital, Leeds. METHODS: Information from 217 cases with histologically proven benign, borderline or malignant tumours was extracted for study. Four variables (age, ultrasound findings with and without colour Doppler imaging and CA125) were entered in the neural network classifier. The neural network results were compared with logistic regression analysis. RESULTS: When used in the neural network the variables of age, CA125 and ultrasound score produced the best result with a sensitivity of 95% and a corresponding specificity of 78% in predicting malignancy. Logistic regression gave a sensitivity or 82% for a specificity of 51%. CONCLUSION: The neural network is a good method of combining diagnostic variables and may be a useful predictor of malignancy in women presenting with ovarian tumours. A comparison of the performance of the neural network with conventional diagnostic methods would be warranted prior to use in clinical practice. PMID- 10519436 TI - Laparoscopic-assisted Doderlein hysterectomy: retrospective analysis of 300 consecutive cases. AB - OBJECTIVE: To assess the clinical outcomes of the Doderlein laparoscopic-assisted hysterectomy. DESIGN: A retrospective study. SETTING: Women's Endoscopic Laser Foundation at South Cleveland Hospital, Middlesbrough and St James's University Hospital, Leeds. POPULATION: Three hundred consecutive women who had a laparoscopic-assisted Doderlein hysterectomy. METHODS: Patients were identified from the laparoscopic hysterectomy theatre log at both sites. Case notes were requested and examined. MAIN OUTCOME MEASURES: Operative time, uterine weight, associated pelvic pathology, blood loss, hospital stay, intra-operative and post operative complications. RESULTS: The operations were performed by eight different surgeons, seven of whom were laparoscopic trainees. The mean operating time was 102 minutes (SD 30). Additional surgery including unilateral or bilateral salpingo-oophorectomy, was carried out in 247 patients (82%). The mean uterine weight was 140 g (SD 74). One hundred and thirty-two women (44%) had a normal pelvis at hysterectomy. The mean drop in haemoglobin and haematocrit was 1.46 g (SD 0.95) and 4.4% (SD 2.8), respectively. The overall complication rate was 18%, of which 6.2% were classed as major. The major complications included four cystotomies, five unscheduled laparotomies, seven post-operative blood transfusions, one pulmonary embolus and two re-operations (within six weeks). The mean hospital stay was three days. CONCLUSIONS: Laparoscopic-assisted Doderlein hysterectomy is an alternative to standard laparoscopic hysterectomy techniques. It has the advantage of being easy to learn and is associated with low complication rates, compared with other laparoscopic and traditional techniques for hysterectomy. PMID- 10519437 TI - Comparison of treatment outcomes for imipramine for female genuine stress incontinence. AB - OBJECTIVE: To assess the efficacy of imipramine as a treatment of genuine stress incontinence and to explore the possible determining factors for treatment success and failure. DESIGN: A prospective study. SETTING: University department of obstetrics and gynaecology. PARTICIPANTS: Forty women with genuine stress incontinence were enrolled. METHODS: Each woman was treated with 25 mg imipramine three times a day for three months. MAIN OUTCOME MEASURES: Each woman had a 20 minute pad test and urodynamic study including uroflowmetry, both filling and voiding cystometry, and stress urethral pressure profile before and after treatment. RESULTS: After treatment, 35% (n = 14) were cured, 25% (n = 10) improved by > or = 50% and in the remaining 40% (n = 16) treatment failed. The efficacy of successful treatment was 60% (95% CI 44.8-75.2). The median age and parity, as well as menopausal status, showed no statistical differences between the two groups. The pre-treatment data including pad weight, functional urethral length, maximal urethral pressure, bladder compliance at urgency, bladder capacity, and average and maximal flow rates showed no statistical differences between the two groups except urethral closure pressure (P = 0.001). Besides, the functional urethral length and urethral closure pressure were significantly improved in the treatment success group. After medication, the median functional urethral length was 3 cm (intraquantile range [IQR] 2.3-3) in the treatment success group vs 2.3 cm (IQR 2-3) in the treatment failure group (P = 0.028). The urethral closure pressure was 77 cmH2O (IQR 61-105) for the treatment success group vs 40 cmH2O (IQR 34-53) in the treatment failure group (P < 0.0001). CONCLUSIONS: The efficacy of imipramine for genuine stress incontinence was 60% (95% CI 44.8-75.2). The pre-treatment high urethral closure pressure may serve as a predictor for treatment success. PMID- 10519438 TI - Postpartum bone mineral density following antenatal dexamethasone therapy. AB - The aim of this study was to determine whether the changes in bone metabolism, which we have demonstrated previously with antenatal dexamethasone therapy, are associated with a lower bone mineral density. We assessed bone mineral density in the proximal femur and lumbar spine using dual photon X-ray absorptiometry after delivery in 15 women who received dexamethasone therapy for fetal lung maturation, and in 30 women who did not have dexamethasone therapy in pregnancy. The absolute bone mineral density, T scores and Z scores at the proximal femur and lumbar spine were similar, and the median values of T and Z scores were positive in both groups. We conclude that antenatal dexamethasone therapy has no long term effect on bone mineral density. PMID- 10519439 TI - Fracture of the sacrum in the absence of osteoporosis of pregnancy: a rare skeletal complication of the postpartum. PMID- 10519440 TI - A bone of contention: an unusual case of secondary infertility. PMID- 10519441 TI - Anal incontinence after vaginal delivery: a prospective study in primiparous women. PMID- 10519442 TI - Primary repair of obstetric anal sphincter rupture using the overlap technique. PMID- 10519443 TI - The performance of screening tests for ovarian cancer: results of a systematic review. PMID- 10519444 TI - Effects of smoking, CYP2D6 genotype, and concomitant drug intake on the steady state plasma concentrations of haloperidol and reduced haloperidol in schizophrenic inpatients. AB - The effects of smoking, CYP2D6 genotype, and concomitant use of enzyme inducers or inhibitors on the steady state plasma concentrations of haloperidol (HAL) and reduced haloperidol (RHAL) were evaluated in 92 schizophrenic inpatients. All but three of these patients received concomitant medication, in many cases with drugs potentially interacting with HAL. Of the 92 patients, 63 were treated orally with HAL in a daily dose of 0.4 to 50 mg; 29 patients were treated intramuscularly with a daily equivalent dose of HAL decanoate (expressed as HAL) of 1.8 to 17.9 mg. A wide interindividual variation in HAL dose and in steady state plasma concentrations of HAL and RHAL was observed. In the patients treated orally, the daily oral dose was about 4 times higher and the dose-normalized HAL (but not RHAL) plasma concentrations were significantly lower in smokers (n = 40) than in nonsmokers (n = 23) (p < 0.01). The dose-normalized RHAL (but not HAL) plasma concentrations and the RHAL/HAL ratio were significantly higher in poor metabolizers (PMs) than in extensive metabolizers (EMs). There was a trend toward an effect of potentially interacting drugs (inducers or inhibitors) on dose, dose normalized HAL and RHAL plasma concentrations, and the RHAL/HAL ratio. In the patients treated intramuscularly, the dose-normalized HAL (but not RHAL) plasma concentrations were significantly lower in smokers than in nonsmokers, but no differences in doses were observed. This naturalistic study of modest sample size in a polymedicated population shows an effect of smoking and CYP2D6 genotype (and to a lesser extent, of interacting drugs) on the kinetics of HAL. PMID- 10519445 TI - Area under the plasma concentration-time curve for total, but not for free, mycophenolic acid increases in the stable phase after renal transplantation: a longitudinal study in pediatric patients. German Study Group on Mycophenolate Mofetil Therapy in Pediatric Renal Transplant Recipients. AB - Mycophenolate mofetil, an ester prodrug of the immunosuppressant mycophenolic acid (MPA), is widely used for maintenance immunosuppressive therapy in pediatric renal transplant recipients. However, little is known about the pharmacokinetics of MPA in this patient population in the stable transplant phase, and dosage guidelines are preliminary. The authors therefore compared the pharmacokinetics of MPA, free MPA, and the renal metabolite MPA glucuronide (MPAG) in the initial (sampling at 1 and 3 weeks) and stable phases (sampling at 3 and 6 months) posttransplant in 17 children (age, 12.0 +/- 0.77 years; range, 5.9 to 15.8 years), receiving the currently recommended dose of 600 mg MMF/m2 body surface area (BSA) twice a day. Plasma concentrations of MPA and MPAG were measured by reverse phase HPLC. Because MPA is extensively bound to serum albumin and only the free drug is presumed to be pharmacologically active, the authors also analyzed the MPA free fraction by HPLC after separation by ultrafiltration. The intraindividual variability of the area under the concentration-time curves (AUC0 12) of MPA throughout the 12-hour dosing interval was high in the immediate posttransplant period, but declined in the stable phase, whereas the interindividual variability remained unchanged. The median MPA-AUC0-12 values increased 2-fold from 32.4 (range, 13.9 to 57.0) mg x h/L at 3 weeks to 65.1 (range, 32.6 to 114) mg x h/L at 3 months after transplantation, whereas the median AUC0-12 values of free MPA did not significantly change over time. This discrepancy can be attributed to a 35% decline of the MPA free fraction from 1.4% in the initial phase posttransplant to 0.9% (p < 0.01) in the stable phase. In conclusion, pediatric renal transplant recipients given a fixed MMF dose exhibit a 2-fold increase of the AUC0-12 of total MPA in the stable phase posttransplant and a 35% decrease of the MPA free fraction, whereas the AUC0-12 of free MPA remains unchanged over time. Because the latter pharmacokinetic variable is theoretically best predictive of the clinical immunosuppressive efficacy of MMF, these findings may have consequences for the dosing recommendations of MMF in renal transplant recipients. PMID- 10519446 TI - Effects of chronic administration of glucocorticoid on midazolam pharmacokinetics in humans. AB - Midazolam (MDZ) is metabolized by CYP3A. Glucocorticoids are potent inducers of CYP3A in humans. The possible interaction between intravenous MDZ and chronically administered glucocorticoids was investigated during surgery in patients. MDZ (0.2 mg/kg) was administered intravenously to 8 patients taking glucocorticoid chronically and 10 patients not taking glucocorticoid. In patients taking glucocorticoid, the AUC0-infinity and CL of MDZ was decreased to 63.9% (16.3 +/- 10.5 vs 25.5 +/- 20.7 microg x min/mL) and increased to 127.5% (16.7 +/- 10.7 vs 13.1 +/- 8.3 mL/min/kg) of that in the control group, respectively. The terminal t1/2 values of MDZ were similar in two groups. In patients taking glucocorticoid, the AUC0-infinity of 1'-hydroxymidazolam (1'-OH MDZ) was 66.7% of that in the control group (7.6 +/- 2.6 vs 11.4 +/- 9.7 microg x min/mL), and the terminal t1/2 of 1'-OH MDZ was significantly (p < 0.01) decreased (1.8 +/- 0.5 vs 3.0 +/- 0.8 hr). Accumulative urinary excretion of 1'-OH MDZ glucuronide was increased to 157.6%. These observations might be results from induction of CYP3A4 and/or UDP glucuronosyltransferase by glucocorticoids. PMID- 10519447 TI - Population pharmacokinetics of gentamicin in preterm neonates: evaluation of a once-daily dosage regimen. AB - Population pharmacokinetic parameters of gentamicin in preterm neonates on a once daily dosage regimen of 3.0 mg/kg given intravenously every 24 hours were established prospectively. In 34 preterm neonates with a mean gestational age of 32 +/- 4 (SD), 182 serum gentamicin levels (91 peak/trough pairs) were determined. Individual adjustments of dose or dosage interval were calculated by computer-aided Bayesian forecasting. The parameters Vd, ke, and CL for each patient were obtained by the nonparametric estimation of maximization method. The predictive power of the model was calculated and the pharmacokinetic estimates were statistically analyzed with SPSS/PC. Cluster analysis showed a division into 2 subpopulations (designated 1 and 2) on the basis of postnatal age. The mean +/- SD postnatal age of subpopulation 1 (n = 29) was 6 +/- 2 days (range 1-7) and of subpopulation 2 (n = 5) 15 +/- 4 days (range 12-24). The mean +/- SD gentamicin relative clearances of subpopulation 1 and subpopulation 2 were 0.0515 +/- 0.0128 and 0.1026 +/- 0.0102 L kg(-1) hr(-1), respectively (p < 0.05). The mean +/- SD values for Vd (Lkg(-1)) in both populations 1 and 2 were 0.6916 +/- 0.1670 and 0.7509 +/- 0.1961, respectively (not significantly different). For ke these data were 0.0744 +/- 0.0200 and 0.1366 +/- 0.0522 (p < 0.05). Statistics showed that the data for Vd and ke of subpopulation 1 were normally distributed (Vd and ke skewness 1.61 and 1.46; kurtosis 3.09 and 3.10 respectively). The model yielded a bias of -0.11 mg/L and a precision of 0.36 mg/L. It is recommended that gentamicin be started in a dosage of 3.5 mg/kg intravenously once-daily under close monitoring. PMID- 10519448 TI - Assessment of acyclovir intraindividual pharmacokinetic variability during continuous hemofiltration, continuous hemodiafiltration, and continuous hemodialysis. AB - The use of intravenous acyclovir can be particularly complicated in pediatric patients with evolving renal impairment, because of intraindividual pharmacokinetic variability linked to the patient's clinical condition. The objective of this study was to use therapeutic drug monitoring data to assess acyclovir intraindividual pharmacokinetic variability during several types of renal replacement therapy. Bayesian adaptive control of acyclovir dosage regimen was performed in a pediatric patient with bone marrow transplant who developed severe renal impairment. Acyclovir pharmacokinetic parameter values corresponding to the different techniques and periods of renal replacement therapy were estimated using USCPACK PC Clinical Programs and therapeutic drug monitoring data. Results showed a wide intraindividual pharmacokinetic variability during CAVH, CAVHDF, and CVVHD, reflecting not only the performance of each dialysis technique but also the difficulty in making use of each one. The acyclovir elimination rate constant was higher during CVVHD compared to CAVH or CAVHDF. Bayesian method appears to be valuable in assessing intraindividual pharmacokinetic variability, as it allows the clinician to deal with sparse routine patient data. PMID- 10519449 TI - High-dose omeprazole: use of a multiple-dose study design to assess bioequivalence and accuracy of CYP2C19 phenotyping. AB - The objectives of this multiple-dose study were to compare the performance of a new formulation of omeprazole (40 mg) with that of an established formulation and to assess the accuracy of CYP2C19 phenotyping during high-dose chronic administration. Twenty-eight healthy subjects were randomized (1:1) to receive 40 mg of either Gasec-40 Gastrocaps (Mepha) or Antra 40 (Astra) daily for 5 days. The pharmacokinetics of omeprazole and the omeprazole/5'-hydroxyomeprazole ratio 3 hours postdose were assessed on day 5. Subjects switched formulations starting on day 6, and all measurements were repeated on day 8. Subjects with metabolic ratios greater than 6 were genotyped for CYP2C19. Gasec-40 was found to be bioequivalent to Antra based on the 90% confidence interval for AUC (102.4-111.7) and Cmax (100.6-120.7). Formulation had no effect on the ratio of omeprazole to 5'-hydroxyomeprazole, which was higher than previously reported with single 20 mg doses of omeprazole. The mean ratio did not differ between day 5 and day 8 but was highly variable: 7 of 28 subjects had more than a 2-fold difference between assessments. In four individuals identified by genotype as extensive metabolizers (EMs), phenotype could not be clearly assigned. The relative bioavailability of omeprazole can be accurately assessed using this multiple-dose study design. Chronic administration of 40 mg doses of omeprazole shifts the metabolic ratio in EMs toward that in poor metabolizers (PMs), apparently because of the nonlinear metabolic clearance of the drug. The assignment of phenotype in patients receiving chronic high-dose omeprazole treatment should be interpreted with caution. PMID- 10519450 TI - Pharmacokinetics of high-dose oral and intravenous dexamethasone. AB - Pharmacokinetics of intravenous and oral pulsed high-dose dexamethasone were studied in four patients with pemphigus vulgaris. Doses for dexamethasone were varied from 100 to 300 mg. Serum concentrations were measured by high-performance liquid chromatographic procedure with diode assay detection. Bioavailability was assessed by comparing the areas under the serum concentration-time curves following oral administration with those of intravenous administration. Mean bioavailability of high-dose oral dexamethasone was 63.4%. Side effects were minor and were limited to temporary facial flushing both after oral and intravenous administration. Oral administration of dexamethasone in pemphigus patients showed to be more convenient and cost effective than administration by the intravenous route. PMID- 10519452 TI - Operator error: a critical determinant of false amikacin and tobramycin concentrations using fluorescence polarization immunoassay kits and TDX analyzer. AB - Amikacin and tobramycin are aminoglycosides used to treat severe Gram-negative infections. Therapeutic monitoring of both drugs is essential to avoid drug toxicity. Fluorescence polarization immunoassay kits for both amikacin and tobramycin are available from Abbott Laboratories, and assays can be run using the TDx analyzer. The test code for amikacin is 3, whereas the test code for tobramycin is 2. Here the authors report that when the operator mistakenly programmed test code 3 (amikacin code) while using the tobramycin kit, the tobramycin values were falsely elevated. The instrument software does not have any mechanism to detect this error. Similarly, when the amikacin kit was used and the operator mistakenly programmed test code 2 (tobramycin code), the amikacin values were falsely lowered. For example, in a serum sample taken from a patient the true amikacin concentration was 20.9 microg/mL. When the amikacin kit was used correctly but test code 2 was programmed, the observed concentration was falsely lowered to 6.1 microg/mL. Similarly, in another specimen the true tobramycin concentration was 3.1 microg/mL. However, when the tobramycin kit was used correctly but test code 3 was used, the observed concentration was falsely elevated to 14.3 microg/mL. The authors also observed a similar effect with controls and spiked specimens containing either amikacin or tobramycin. They conclude that correct programming of the test code is vital to obtaining useful results in amikacin and tobramycin analysis using the TDx analyzer. PMID- 10519451 TI - Drug interaction between mycophenolate mofetil and tacrolimus detectable within therapeutic mycophenolic acid monitoring in renal transplant patients. AB - The possible pharmacokinetic interaction between the new immunosuppressive mycophenolate mofetil (MMF) and tacrolimus (TAC), respectively, was assessed by comparing routinely estimated mycophenolic acid (MPA) plasma trough levels of 15 consecutive renal transplant patients receiving MMF in combination with methylprednisolone (MEP) and cyclosporin A (CSA, 10 patients) or in combination with MEP and tacrolimus (TAC, 5 patients). Coadministration of TAC instead of CSA resulted in a significant increase of mean MPA plasma trough levels [3.4 +/- 1.3 microg/mL (n = 22) versus 1.87 +/- 1.1 microg/mL (n = 57); p < 0.001], despite lower MMF doses [1.5 +/- 0.5 g/d versus 1.7 +/- 0.3 g/d (not statistically significant)]. This elevation in MPA levels is possibly caused by an interaction between MMF and TAC and could lead to a recommendation to monitor MPA plasma levels with appropriate dose adjustment. PMID- 10519453 TI - Acetaminophen (paracetamol) levels in human tears. AB - This study was designed to measure acetaminophen (paracetamol) levels in tears, and to compare it to serum levels. Paracetamol levels were measured in 20 paired tears and serum samples from 10 healthy volunteers, 1 and 2 hours after ingesting 1.5 g paracetamol. Tears were collected using glass microcapillary tubes while stimulating the conjunctiva with a small sponge placed in the lower fornix. Blood samples were taken simultaneously. The samples were analyzed for paracetamol levels using homogeneous enzyme immunoassay. Tears and serum paracetamol levels 1 hour after ingestion were 16.3 microg/mL +/- 7.2 (mean +/- SD), and 21.4 microg/mL +/- 7.7 (mean +/- SD) respectively. Tears and serum levels 2 hours after ingestion were 14.4 microg/mL +/- 7.8 (mean +/- SD), and 17 microg/mL +/- 7.6 (mean +/- SD) respectively. Tears and serum paracetamol levels of all the 20 paired samples (1 h and 2 h after ingestion) were 15.35 microg/mL +/- 7.4, and 19.25 microg/mL +/- 7.8, respectively (mean +/- SD). There was a strong and highly significant correlation between paracetamol levels in serum and in tears 1 and 2 hours after ingestion (r = 0.8, p = 0.005, r = 0.85, p = 0.002 respectively). Mean +/- SD ratio of tears/serum paracetamol levels 1 hour and 2 hours after ingestion were 0.77 +/- 0.21 and 0.81 +/- 0.25 respectively. Delta tears (difference in mean levels at 1 and 2 hours) paracetamol levels is significantly correlated with delta serum levels (r = 0.7, p = 0.025). A reliable, convenient, and feasible noninvasive method is described for measuring paracetamol in tears. There is no information in the literature about tears paracetamol secretion, and little information of tears drugs concentration. PMID- 10519454 TI - Severe side effects and drug plasma concentrations in preterm infants treated with doxapram. AB - A high-performance liquid chromatography method has been developed for simultaneous determination of doxapram and its metabolites including ketodoxapram, the main and only active metabolite. The aim of the study was to evaluate this microtechnique and to report the cases involving severe adverse effects to determine toxic plasma levels in neonates. The method was found to be selective, and showed a good baseline separation of doxapram and metabolites. Recovery, linearity, intraday/interday precision, and limit of detection determined in aqueous solutions and in spiked plasma were satisfactory. The assay is simple, rapid, and plasma-sparing, which represents a true advantage in managing neonates. Case analysis was performed in two consecutive periods: 124 preterm infants in the first period and 173 in the second period. Severe toxic effects were observed in 4 cases in the first period, with doxapram plus keto doxapram levels 9 mg/L. In the second period, only one case was observed. High range plasma concentrations were significantly less frequent in the second period than in the first one. The authors conclude that measuring doxapram plus keto doxapram in plasma may be of interest to avoid severe toxic effects in preterm neonates treated with doxapram. PMID- 10519456 TI - Liquid chromatographic determination of six antiepileptic drugs and two metabolites in microsamples of human plasma. AB - A simple, rapid, sensitive, and reproducible high-performance liquid chromatographic (HPLC) method for simultaneous determination of the antiepileptic drugs (ethosuximide, primidone, lamotrigine, phenobarbital, phenytoin, and carbamazepine) and two metabolites (carbamazepine-diol and carbamazepine-epoxide) in human plasma is described. The procedure involves extraction of the drugs from human plasma (100 microL) with ether using 9-hydroxymethyl-10-carbamyl acridan as an internal standard. The extract was evaporated and reconstituted with mobile phase and then injected onto the chromatograph. The drugs and the internal standard were eluted from a Supelcosil LC-18 stainless steel column at ambient temperature with a mobile phase consisting of a 0.01M phosphate buffer/methanol/acetonitrile (65/18/17, v/v/v) adjusted to a pH of 7.5 with phosphoric acid and a flow rate of 1 mL/min. The effluent was monitored at 220 nm. Quantitation was achieved by using peak area ratio of each drug to the internal standard. The intraassay and interassay coefficients of variation (CV) ranged from 2.43% to 6.25% and from 3.02% to 5.85%, respectively. The absolute (extraction) and relative (analytical) recoveries for the drugs ranged from 70.7% to 104.4% and from 88.3% to 106.1%, respectively. Stability tests showed that the drugs were stable in plasma for at least 4 weeks when stored at -20 degrees C. The method was applied clinically for monitoring the AEDs in epileptic patients. PMID- 10519455 TI - Comparison of the serum barbiturate fluorescence polarization immunoassay by the COBAS INTEGRA to a GC/MS method. AB - The authors evaluated a new Cassette Serum Barbiturates fluorescence polarization immunoassay (FPIA) on the COBAS INTEGRA. The assay was calibrated with secobarbital standards at 0, 0.5, and 4.0 microg/mL. The assay range was 0.030 to 80 microg/mL using an automatic 1/20 postdilution feature. Precision was established for two COBAS INTEGRA instruments for ten days by assaying secobarbital target concentrations ranging from 0.125 to 2.2 microg/mL. The coefficients of variation (CV) for the above target concentrations for the first instrument ranged from 2.7% to 8.3%, and for a second instrument, 3.8 to 8.3%. Seven clinically elevated bilirubin samples were spiked with 0.46 and 1.77 microg/mL secobarbital. Bilirubin interference was less than 10.9 and less than 7.9%, respectively. The average recovery ranged from 85% to 94%. The mean difference in recovery in serum versus plasma was < or = 3%. Fifty-two clinical samples were analyzed for butalbital, pentobarbital, secobarbital, and phenobarbital by GC/MS, and the results were compared to the new Cassette Serum Barbiturates FPIA. The diagnostic sensitivity and specificity were 95% and 100%, respectively. Both FPIA and GC/MS assays are clinically efficacious for monitoring serum barbiturates. PMID- 10519457 TI - Therapeutic monitoring of topiramate: evaluation of the saturable distribution between erythrocytes and plasma of whole blood using an optimized high-pressure liquid chromatography method. AB - Topiramate (TPM) reportedly binds in a saturable manner to erythrocytes but minimally to plasma proteins. Two studies were performed to evaluate this distribution phenomenon. In all studies, TPM was measured with a newly developed, optimized procedure that uses octyldecyl (C-18) solid phase sorbents disks/packed cartridges and a DB-1 methylsilicone capillary gas chromatography (GC) column. Between-run precision coefficients of variation (CVs) (n = 16) ranged from 3.6% 5.6% at concentrations from 3.0 to 15 microg/mL, with low limit of detection of 0.2 to 0.3 microg/mL. For the distribution studies, drug-free whole-blood specimens from five healthy adult volunteers were supplemented with TPM and used to test the influence of TPM concentration and HCT differences on the plasma/blood (P/B) distribution ratio of TPM. In study A, TPM concentration was varied (1-15 microg/mL) and HCT remained constant (40% +/- 5%). In study B, TPM (3 microg/mL) was added to blood specimens comprising a range of HCT values (20% 40%). Study A results were: mean TPM P/B ratios: 0%, 14.2% +/- 5%, 44.2% +/- 4%, 76% +/- 5.5% at 1, 3, 5, 15 microg/mL, respectively. Data between each group were statistically different (p < 0.001). Study B results were: mean TPM P/B ratio: 17.3% +/- 7.3%, 27.5% +/- 10.1%, 39.8% +/- 8% and 56.1% +/- 8.8% at HCT values of 40%, 32%, 26.5%, 20%, respectively. The TPM P/B ratio was significantly inversely correlated to HCT (r = -905, p < 0.001). TPM P/B partitioning was not temperature dependent. Researchers concluded that the saturable binding of TPM to RBC is significant and is correlated to HCT. As a result, TPM in the plasma fraction of whole blood will increase when HCT decreases and as total TPM concentration in whole blood increases. PMID- 10519458 TI - No effect of the new antidepressant reboxetine on CYP2D6 activity in healthy volunteers. AB - The effect of the new antidepressant reboxetine on the activity of the cytochrome P450 (CYP) 2D6 isoenzyme was investigated in 10 healthy volunteers using dextromethorphan as a model CYP2D6 substrate. Each volunteer received a single 30 mg oral dose of dextromethorphan on three different occasions separated by an interval of at least 4 weeks: a) in a control session; b) after 1 week of treatment with reboxetine, 8 mg/day; and c) after 1 week of treatment with paroxetine (an inhibitor of CYP2D6 activity) 20 mg/day. Urine was collected over the next 8 hours for the determination of the dextromethorphan/dextrorphan metabolic ratio. All subjects were classified as extensive metabolizers (EM) with a dextromethorphan/dextrorphan ratio < 0.3. There were no notable changes in the urinary dextromethorphan/dextrorphan ratio in the reboxetine phase as compared to the control session. By contrast, there was a statistically significant increase in the metabolic ratio in the paroxetine phase (p < 0.001), with 4 subjects switching to poor metabolizer (PM) phenotype. These results suggest that reboxetine is unlikely to cause clinically significant interactions with substrates of CYP2D6. PMID- 10519459 TI - Could discontinuing smoking be hazardous for patients administered clozapine medication? A case report. AB - A 35-year-old man with schizophrenia was successfully treated with clozapine at a daily oral dose of 700-725 mg for more than 7 consecutive years. Two weeks after abrupt cessation of chronic heavy cigarette smoking, he suddenly developed tonic clonic seizures followed by stupor and coma. After 2 days of intensive care, the patient recovered completely but could not recall the episode. Clozapine therapy was reinstituted and could be carried out successfully at 425 mg daily, i.e., at an approximately 40% reduction of the daily dose before he stopped smoking. The sudden cessation of smoking most likely caused a rise in plasma concentrations of clozapine and/or clozapine metabolites resulting in the seizure episode. A likely mechanism is that the heavy smoking had induced cytochrome P450-1A2, the main enzyme involved in the metabolism of clozapine. PMID- 10519460 TI - Psychosocial predictors of delay of first sexual intercourse by adolescents. AB - This investigation predicted adolescents' delay of intercourse onset from attitudes, social norms, and self-efficacy about refraining from sexual intercourse. Age, gender, ethnicity, and parental education were also examined as predictors and moderators of the relationships among the 3 psychosocial determinants and onset. The participants (N = 827), part of a cohort initially surveyed in the 9th grade, reported at baseline that they had never engaged in intercourse. The multivariable proportional hazards regression model suggested that adolescents with more positive attitudinal and normative beliefs, as well as those with a parent who graduated from college, were less likely to engage in intercourse in the follow-up period (up to approximately 2 years). Interventions that include an objective to delay onset may benefit from addressing psychosocial determinants, especially attitudes and norms about sexual intercourse. PMID- 10519461 TI - Peer and adolescent substance use among 6th-9th graders: latent growth analyses of influence versus selection mechanisms. AB - This study analyzed peer-influence versus peer-selection mechanisms in adolescent tobacco, alcohol, and marijuana use. Participants were surveyed 3 times, with 1 year intervals, about peers' substance use and their own use; Sample 1 had 1,190 participants (initial mean age = 12.4 years), Sample 2 had 1,277 participants (initial mean age = 11.5 years). Latent growth analyses that were based on composite scores indicated that initial peer use was positively related to rate of change in adolescent use, supporting the influence mechanism; there was little evidence for a selection mechanism. Difficult temperament, poor self-control, and deviance-prone attitudes were related to initial levels for both peer and adolescent use. It is concluded that peer influence is the primary mechanism during middle adolescence. Temperament-related attributes may be predisposing to early experimentation and deviant-peer affiliations. PMID- 10519462 TI - Motivations for condom use: do pregnancy prevention goals undermine disease prevention among heterosexual young adults? AB - Differences in motives for condom use and their implications for understanding frequency of use were investigated in a random, biracial (Black, White) sample of heterosexuals, aged 17 to 25 years (n = 902). Results indicated that sexually active young adults-regardless of race, age, gender, or relationship status-were more likely to use condoms to prevent pregnancy than to prevent disease. Reasons for use mediated the effects of relationship status on condom use per se and moderated the effects of attitudinal and perceptual variables on condom use. Finally, comparisons among condom users motivated by different prevention goals and nonusers (n = 388) revealed that differences among user subgroups were nearly as numerous and, in some cases, more robust than differences between users and nonusers. These findings indicate that condom users comprise distinct subgroups, defined in part by their underlying motives for use, and highlight important conceptual and empirical reasons to distinguish among them. PMID- 10519463 TI - Self-blame attributions in women with newly diagnosed breast cancer: a prospective study of psychological adjustment. AB - Associations between self-blame and anxiety and depression symptoms in a sample of 76 women with breast cancer were investigated. At diagnosis, behavioral self blame was associated with increased distress; at 3 months postdiagnosis, characterological self-blame was positively associated with affective symptoms and behavioral self-blame approached significance (p = .07); and at 6 months, behavioral self-blame was related to increased distress. Prospective analyses revealed that characterological self-blame at diagnosis approached significance in predicting distress at 3 months (p = .055) and was significant in predicting distress at 6 months and at 1 year after diagnosis. These data indicate that behavioral self-blame is a correlate of concurrent affective symptoms, whereas characterological self-blame predicts increased distress over time. Implications for social-cognitive processes in adaptation to breast cancer are discussed. PMID- 10519464 TI - Psychosocial stress and social support are associated with prostate-specific antigen levels in men: results from a community screening program. AB - Perceived stress and satisfaction with social support were assessed in 318 men participating in a prostate-specific antigen (PSA) screening program. We predicted that high stress would be associated with high PSA and high social support with low PSA. We also predicted a Stress x Support interaction (buffering). Hypothesized main effects were confirmed and were not explained by recency of previous rectal examinations or current urinary symptoms. There was no evidence of buffering. Stress and social support were not associated with results of digital rectal examinations. These findings raise the possibility that psychosocial factors promote prostate disease through direct physiologic pathways. However, it is also possible that the data reflect effects of stress on health-related behaviors or that stress scores were affected by participants' anticipation of prostate problems on the basis of prior PSA tests or symptoms. PMID- 10519465 TI - Understanding how people process health information: a comparison of tailored and nontailored weight-loss materials. AB - Health information tailored to meet individuals' unique needs has been shown to be more effective than generic information in promoting risk-reducing behavior changes. To explore mechanisms underlying tailoring's effectiveness, this study randomly assigned 198 overweight adults to receive weight-loss materials that were (a) tailored to the individual, (b) in an American Heart Association (AHA) brochure, or (c) AHA-content formatted to look like tailored materials. Participants who received tailored materials had more positive thoughts about the materials, positive personal connections to the materials, positive self assessment thoughts, and positive thoughts indicating behavioral intention than those who received either of the untailored materials. The tailoring of health information can significantly improve the chances the information will be thoughtfully considered and can stimulate prebehavioral changes such as self assessment and intention. PMID- 10519466 TI - The World Health Organization WHOQOL-100: tests of the universality of Quality of Life in 15 different cultural groups worldwide. AB - The World Health Organization Quality of Life assessment (WHOQOL-100) was developed simultaneously across 15 international field centers and includes 24 facets relating to quality of life, which are grouped into 4 larger domains: physical, psychological, social relationships, and environment. It also includes 1 facet examining overall quality of life and general health perceptions. This article examines the extent to which the WHOQOL-100 assesses quality of life perceptions in different cultures and whether it is structurally comparable in these cultures. Regression analysis showed all 4 domains to be important in assessing quality of life in each of the 15 centers. Structural equation modeling suggested further support for the proposal that there are universal facets and domains that are cross-culturally important in determining quality of life and suggested that the ordering of facets within domains is comparable across cultures. PMID- 10519467 TI - A meta-analysis of psychoeduational programs for coronary heart disease patients. AB - In a meta-analysis of 37 studies, the effects of psychoeducational (health education and stress management) programs for coronary heart disease patients were examined. The results suggest that these programs yielded a 34% reduction in cardiac mortality; a 29% reduction in recurrence of myocardial infarction (MI); and significant (p < .025) positive effects on blood pressure, cholesterol, body weight, smoking behavior, physical exercise, and eating habits. No effects of psychoeducational programs were found in regard to coronary bypass surgery, anxiety, or depression. The results also suggest that cardiac rehabilitation programs that were successful on proximal targets (systolic blood pressure, smoking behavior, physical exercise, emotional distress) were more effective on distal targets (cardiac mortality and MI recurrences) than programs without success on proximal targets. PMID- 10519468 TI - Psychosocial factors and the development of breast cancer: a meta-analysis. AB - A meta-analysis examined the relationship between psychosocial factors and the development of breast cancer. Average effect sizes (Hedges's g) were calculated from 46 studies for 8 major construct categories: anxiety/depression, childhood family environment, conflict-avoidant personality, denial/repression coping, anger expression, extraversion-introversion, stressful life events, and separation/loss. Significant effect sizes were found for denial/repression coping (g = .38), separation/loss experiences (g = .29), and stressful life events (g = .25). Although conflict-avoidant personality style was also significant (g = .19), the effect size was less robust, and a moderate number of future studies with null results would reduce the significance. Results overall support only a modest association between specific psychosocial factors and breast cancer and are contrary to the conventional wisdom that personality and stress influence the development of breast cancer. PMID- 10519469 TI - Specific worry about breast cancer predicts mammography use in women at risk for breast and ovarian cancer. AB - This longitudinal study examined predictors of mammography use among women with a family history of breast cancer participating in a risk assessment and surveillance program (N = 213). Assessed were background variables (age, prior mammography utilization), cognitive variables (perceived vulnerability), and affective variables (cancer worry and general distress). Results of logistic regression analyses predicting adherence 1 year after baseline contact, in which variables of prior utilization, feelings of vulnerability, and general distress were controlled for, indicated that cancer worry and age were significant predictors of mammography adherence. Results suggest that moderate levels of cancer worry facilitate, rather than undermine, adherence. The results have implications for the construction of educational messages that should be designed to acknowledge feelings of cancer-specific worry and to provide guidance in health protective behaviors. PMID- 10519470 TI - Relations of diabetes intrusiveness and personal control to symptoms of depression among adults with diabetes. AB - The generalizability of a model linking illness characteristics to psychosocial well-being was tested in a cross-sectional study of 237 adults with type 2 diabetes. It was hypothesized that diabetic complications increase illness intrusiveness, which in turn increases depressive symptomatology either directly or indirectly by reducing personal control over health outcomes. Illness intrusiveness was defined as the result of disruptions of valued activities and interests due to constraints imposed by the illness. An excellent fit of this model to the data was found using structural equation modeling. The model explained 65% of the variance in depressive symptomatology. Assessment of an alternative model excluding personal control suggested that the extent to which diabetes intrudes in life, rather than diabetic complications per se or personal control, is a key factor in relation to depressive symptomatology in individuals with diabetes. PMID- 10519471 TI - Stage of change as a predictor of success in weight control in adult women. AB - Weight change over 3 years was examined in a large and heterogeneous sample of women as a function of stage of change for weight control. Women were classified into Precontemplation, Contemplation, Preparation, and Action stages on the basis of reports of current and past weight control behaviors and future intentions. Stage of change did not predict success in weight control. Mean weight changes over 3 years were 1.1 kg, 1.0 kg, 2.1 kg, and 2.3 kg for Precontemplation, Contemplation, Preparation, and Action stages, respectively. The findings call into question the generality of the stages-of-change classification system across behavioral domains. PMID- 10519472 TI - Vestibular autorotation test in primary selection of military student pilots using neural networks. AB - BACKGROUND: Selection of student pilots to the Royal Danish Air Force involves a physical examination including a vestibular test. Usually tests for selection proposes are not well documented. HYPOTHESIS: The result of the vestibular autorotation test (VAT) is correlated to the ability to learn to fly in a military context. METHODS: A Multi Layer Perception neural network with three layers configured as a Back Propagation Network was tested using data originating from horizontal VAT of 59 student pilot candidates, given the outcome of the pre jet basic flight check. In the analysis the leave-one-out method was used. RESULTS: Based on horizontal data only the network correctly classified the student pilot candidates as having been passed or rejected within a verification error margin < 0.1. CONCLUSION: The result indicates that VAT performed at the initial physical examination is a powerful tool for the elimination of unfit student pilot candidates when data are analyzed in neural networks. PMID- 10519473 TI - Effects of gender of subjects and experimenter on susceptibility to motion sickness. AB - BACKGROUND: It has been reported that females are more susceptible to motion sickness than males, but these reports have failed to take into account the possible effects of the gender of the experimenter and the subjective nature of reports of symptoms of motion sickness. To deal with the first possible confound, we used male and female experimenters. To deal with the second issue, we recorded gastric myoelectric activity so as to be able to quantify gastric tachyarrhythmia, an objective measure that has been shown previously to correlate highly with severity of symptoms. METHOD: There were 34 male and 34 female participants were assigned to either a male or female experimenter. Symptoms of motion sickness were induced by placing participants in an optokinetic drum for an 8-min baseline period followed by a 16-min rotation period. Electrogastrograms (EGGs) were continuously recorded, and reports of symptoms were obtained from the participants every 3 min during rotation. RESULTS: Comparison of male and female subjects' symptom scores revealed that females had higher symptom scores than males; however, no significant main effects for gender of the subject or experimenter were found. However, on a post-session questionnaire, females reported experiencing significantly more GI symptoms than males. Gender comparisons of the change in gastric tachyarrhythmia power from baseline to rotation yielded no significant differences. CONCLUSIONS: Females report more overall symptoms of motion sickness and significantly more GI symptoms than males, but do not show greater increases in gastric tachyarrhythmia during exposure to a rotating drum. PMID- 10519474 TI - The effects of roll vs. pitch rotation in humans under orthostatic stress. AB - BACKGROUND: It has been known since 1953 that pre-exposure to less than +1 Gz will reduce subsequent +Gz-tolerance. With few exceptions, during operational flying, the transition from hypogravity to hypergravity involves roll as well as pitch rotation. We examined the effect of roll vs. pitch rotation while undergoing transition from hypogravity to +1 Gz on a tilt table. METHODS: Twelve subjects (28-47 yr old) were rotated at 45 degrees x s(-1) from head-up (HU) at 15 degrees relative to gravitational vertical to 135 degrees head-down (HD) and back to the HU position after different HD dwell times. HD dwell times were set at 7, 15, and 30 s. The subject was rotated about the interaural axis (pitch) and about the naso-occipital axis (roll). Both the HD dwell times and axes of rotation were randomized within and across subjects. BP and heart rate were recorded during the HU-HD-HU maneuver. RESULTS: Analysis of variance, repeated measure design revealed that the rate and magnitude of BP decrease induced by the HD to HU maneuver is significantly higher (p < 0.01) in roll than in pitch during all HD dwell times. The decrease of BP at 7s is significantly (p < 0.01) higher than at 15s and 30s. Heart rate increases significantly higher (p < 0.01) in pitch than in roll at 7s-dwell time. CONCLUSION: Our results suggest that the compensatory mechanism to orthostatic stress is more efficient in response to pitch than roll rotation. This is reflected from the findings that the mean magnitude of OH (orthostatic hypotension) and the rate of BP decrease induced by the HD-HU maneuver is significantly greater in roll rotation than pitch rotation. The mean HR increase post HD-HU rotation is significantly higher in the pitch than the roll rotation. The significant rate of BP decrease during HD-HU roll rotation could have important implications for maintaining G-tolerance and spatial orientation during subsequent exposure to hypergravity. PMID- 10519475 TI - Effect of aerobic training on orthostatic tolerance, circulatory response, and heart rate dynamics. AB - BACKGROUND: This study was performed to investigate the effects of aerobic training on orthostatic tolerance and to quantify the post-training changes in cardiovascular response and heart rate variability (HRV). METHODS: Tolerance and circulatory responses to two types of lower body negative pressure (LBNP) were examined and compared in a group of healthy male students before and after 6 mo of aerobic training, and the results were further compared with a group of athletes (runners). Changes in HRV associated with training were analyzed by conventional and time-varying autoregressive spectral analysis, as well as by approximate entropy measurement (ApEn)--a statistic quantifying heart rate "complexity" derived from non-linear dynamics. RESULTS: After aerobic training, there was an initial transient hypotension during the supine -50 mmHg LBNP testing and a significant decrease in tolerance to upright graded LBNP in most of the student-subjects. Moreover, after training, there was a significant decrease in ApEn value of the HRV time series during both supine control and LBNP testing, and the rate of cardiac vagal withdrawal and sympathetic activation during the onset of LBNP was faster than that before training. CONCLUSIONS: The present study has provided further evidence that certain types of aerobic training may affect orthostatic tolerance and may be associated with a loss of complexity of HRV during supine resting and orthostatic stress. PMID- 10519476 TI - Significance of low hematocrit levels in asymptomatic young adults: results of 15 years follow-up. AB - HYPOTHESIS: Periodic complete blood counts are not recommended for disease prevention in low-risk, non-pregnant adults. Consequently, there are few follow up studies of the prevalence of incidentally detected anemia in asymptomatic subjects and its significance for their well-being. The objective of this survey is to determine the frequency of anemia and its predictive value for disease over a 15-yr annual follow-up of a cohort of asymptomatic young males, selected for physical fitness and intelligence. METHODS: One thousand Israeli airmen aged 18 30 yr at entry into this historical-prospective study in 1968 were used as subjects. Hematocrit (Hct) levels were examined annually. On the average each subject had 13.2 tests in the course of the 15 yr follow-up. We arbitrarily defined anemia as a Hct of 40% or less on two or more tests, and compared the prevalence of diagnosed disorders in subjects with and without anemia. RESULTS: During follow-up, anemia was found in 125 (12.5%) of the subjects. On successive annual examinations of the same individual Hct levels varied by 3% or more in 3.5% of those without anemia, and in 10.5% of those with anemia. The frequency of diagnosed disorders, excluding inter-current infections and trauma, was 25.6%, and 10.9% among those with and without anemia, respectively (OR = 2.8, 95% CI 1.8 4.6). Anemia was associated with inflammatory bowel disease (OR = 12.1, 95% CI 2.3 78.6) and malignancy (OR = 3.6, 95% CI 1.1-10.7). It preceded diagnosis only in one case with Waldenstr 246 m's macroglobulinemia, in one case of inflammatory bowel disease and two cases of myocardial infarction. CONCLUSIONS: A finding of anemia doubled the likelihood of chronic disease. However, it had a limited predictive value for subsequent morbidity and did not lead to detection of treatable disorders or to disorders that might otherwise have endangered flight safety. Fluctuations of up to 3% in Hct over time may be viewed as normal in young males. PMID- 10519477 TI - Dantrolene and recovery from heat stroke. AB - Several authors have shown that dantrolene may be effective in the treatment of heat stroke patients. However, the scant data available are still controversial. The aim of this investigation was to establish an animal experimental model for studying the efficacy of this drug both as a prophylactic agent and as a means of hastening the cooling process after heat stroke. Male albino rats were divided into five groups: Sedentary controls (SC), Sedentary+dantrolene (S+D), Exercise controls (EC), and Exercise+dantrolene (E+D, E+D1). The drug (140 mg x kg(-1) body weight) was administered i.v. either prior to subjection to heat stress (40 degrees C) (S+D, E+D) or upon development of heat stroke syndrome (E+D1). In the S+D group, dantrolene administered prior to heat stress (HS) delayed the development of heat stroke by 70%, although colonic temperature (Tc) at the onset of heat stroke was similar to that in group SC (43.0 +/- 0.1 degrees C and 43.2 +/- 0.4 degrees C for S+D and SC, respectively). In E+D animals, dantrolene shortened exercise endurance in the heat by 17.5%, but concomitantly hindered severe Tc rise (40.9 +/- 0.2 degrees C and 40.0 +/- 0.2 degrees C for EC and E+D, respectively). Administration of dantrolene on the development of heat stroke appeared to improve cooling in the exercise group (0.25 degrees C x min(-1) and 0.18 degrees C x min(-1), for the first 1 5 min of cooling, for E+D1 and EC, respectively). The results suggest that dantrolene is effective as a prophylactic agent in sedentary animals only. It also might have application on development of heat stroke. It is hypothesized that the observed rapid cooling is associated with dantrolene's effect on muscle contraction, thus leading to attenuated heat production and peripheral vascular relaxation. PMID- 10519478 TI - Evaluation of cutaneous insensible water loss during hyperbaric exposure in humans. AB - BACKGROUND: Water evaporation diminishes in high pressure environment, however it is unknown whether insensible water dissipation from the human skin falls as a function of the increased environmental pressure. We designed the present study to measure cutaneous insensible water loss at various pressures during exposure to a simulated saturation dive. METHODS: Four healthy male volunteers were exposed to eight different pressures between 1 and 18.4 atmospheres absolute (atm abs). Resting insensible water loss from the skin was measured as change in the body weight and corrected for the weight of the respiratory CO2 - O2 gas exchange and the respiratory water dissipation. RESULTS: We made an equation for the relationship between cutaneous insensible water loss and environmental pressure as: w = 14.5 X p(-0.48), where, w is cutaneous insensible water loss in g x m(-2) x h(-1), and P is the environmental pressure in atm abs. The average cutaneous insensible water loss (15.3 g x m(-2) x h(-1)) at normal atmosphere decreased (p < 0.01) to 4.2 g x m(-2) x h(-1) (reduced by 73%) during a saturation dive to 18.4 atm abs. CONCLUSION: The amount of insensible water loss estimated from the equation was comparable to that of reported observations. PMID- 10519479 TI - Cross validation of USARIEM heat strain prediction models. U.S. ARMY Research Institute of Environmental Medicine. AB - HYPOTHESIS: This study was a cross validation of three heat strain prediction models developed at the U.S. Army Research Institute of Environmental Medicine: the ARIEM, HSDA, and ARIEM-EXP models ability to predict core temperature. METHODS: Seven heat-acclimated subjects completed twelve experimental tests, six in each of two hot climates, at three exercise intensities and two uniform configurations in each climate. RESULTS: Experimental results showed physiological responses as expected with heat strain increasing with work load and level of protective clothing, but with similar heat strain between the two environments matched for wet bulb, globe index. Neither the ARIEM or HSDA model closely predicted core temperatures over the course of the experiment, due mostly to an abrupt initial rise in core temperature in both models. A proportionality constant in the ARIEM-EXP buffered some of this abrupt rise. CONCLUSIONS: Comparisons of the core temperature and tolerance times data with the three models led to the conclusions that for healthy males: 1) the ARIEM and HSDA models provide conservative safety limits as a result of predicting rapid initial increases in core temperature; 2) the ARIEM-EXP most closely represents core temperature responses; 3) the ARIEM-EXP requires modifications with an alternate proportionality coefficient to increase accuracy for low metabolic cost exercise; 4) all of the models require additional input from existing research on tolerance to heat strain to better predict tolerance times; and 5) additional models should be examined to investigate the transient state of the body as it is affected by environment, clothing and exercise. PMID- 10519480 TI - Blood ammonia response during incremental and steady-state exercise in military staff. AB - BACKGROUND: Although in the last few years it has been possible to determine blood ammonia, its application in coaching practice has not yet been fully established. This study was designed to evaluate the blood ammonia response to a laboratory incremental exercise test and three steady-state field tests. METHODS: There were 26 military personnel who performed a submaximal and maximal exercise test on a treadmill, and a field test which included three different constant velocity stages. Gas exchange parameters were monitored throughout the maximal test. Capillary blood samples were obtained from fingertips during the submaximal and field tests for the determination of ammonia and lactate. RESULTS: The ammonia threshold was detected in 23 subjects (88.5%) during the submaximal test. No significant differences were found between the ammonia and lactate thresholds which were shown to be significantly correlated. Blood ammonia levels showed a progressive increase during the last two stages of the field test while lactate levels remained stable at less than 4 mmol x L(-1). CONCLUSIONS: The steady increase in blood ammonia concentration recorded in the field test suggests the possibility of using blood ammonia levels to monitor the duration of exercise although further investigation is required to explore this possibility. Moreover, the assessment of blood ammonia levels during incremental exercise protocols confirms the existence of an ammonia threshold, defined as the intensity of exercise at which ammonia shows a progressive increase. PMID- 10519481 TI - The effects of betaxolol hydrochloride ophthalmic solution on intraocular pressures during transient microgravity. AB - BACKGROUND: Intraocular pressure (IOP) has been found to increase during microgravity. After peaking in the first few hours of orbital flight, IOP slowly decreases to a level that is slightly elevated above baseline IOP's. These modest elevations in IOP do not require treatment. Just as in 1-G, a clinically significant elevation of IOP that occurred during spaceflight would require treatment. We are not aware of previous studies of the efficacy of IOP lowering agents under conditions of microgravity. METHODS: This double-masked, placebo controlled study measured the IOP's of 11 adult subjects (22 eyes) at baseline, preflight, and zero-gravity aboard the NASA KC-135 aircraft, and postflight. One eye of each of the subjects was treated with betaxolol hydrochloride ophthalmic solution 0.5%, while the contralateral eye was treated with normal saline placebo, for 7 d prior to parabolic flight. IOP's were measured by the Tono-Pen 2, a gravity independent tonometer. RESULTS: A modest, but statistically significant reduction of 2.4 mmHg in mean IOP was noted in betaxolol treated eyes at the time of preflight measurement. During zero-G, the mean IOP's of both betaxolol treated eyes and placebo treated eyes increased approximately 20% over preflight levels. Postflight IOP's were similar to preflight IOP's. CONCLUSIONS: The effect of betaxolol on the IOP of eyes treated with for 1 wk prior to exposure to microgravity was statistically significant, but may lack clinical significance in normal eyes. Further research needs to be done to determine the efficacy during microgravity of betaxolol and other agents, in subjects who have upper normal to slightly elevated IOP's at 1 G. PMID- 10519482 TI - Detecting transient cognitive impairment with EEG pattern recognition methods. AB - This paper describes an initial evaluation of a new method for assessing transient states of cognitive impairment associated with intoxication or fatigue: neural network pattern recognition applied to features of the electroencephalogram (EEG) recorded from subjects performing a standardized task. Nine subjects performed a working memory task during an extended testing session occurring over the course of one night, and encompassing an alert baseline period, a state of mild acute intoxication, and a state of fatigue compounded by "hangover" or intoxication after-effects. Relative to the alert baseline, task performance was less accurate in the other test conditions, providing evidence of transient cognitive impairment. These states of impairment were associated with changes in spectral characteristics of the EEG. Neural network-based EEG pattern recognition techniques were used to develop and test detectors of these changes. Brief testing data samples originating from the alert baseline condition could be discriminated from those recorded during the state of acute intoxication with 98% accuracy (p < 0.0001), and from those recorded during the state of fatigue/hangover with 92% accuracy (p < 0.001). Furthermore, networks trained on data from a group of subjects were found to accurately classify data from test subjects who were not part of the training group. These results demonstrate the feasibility of using neurophysiological monitoring methods for detecting transient cognitive impairment. PMID- 10519483 TI - Biochemical characteristics of beta-adrenoceptors in rats after an 18-day spaceflight (LMS-STS78). AB - BACKGROUND: To ascertain whether there was autonomic adaptation with the development of adrenoceptor hypersensitivity under microgravity, the biochemical properties of the beta-adrenoceptors were determined using (125I)iodocyanopindolol (ICYP) binding in rats flown for 18 d onboard the space shuttle. METHODS: This study was performed on heart and kidneys of 3 groups of 12 animals: the flight and 2 ground control (vivarium and AEM) groups. To distinguish the possible role of the corticosteroids, half of each animal group was bilaterally adrenalectomized (ADX rats) with an aldosterone and corticosterone supplementation while the other half was SHAM operated. RESULTS: The Scatchard analysis of the ICYP-binding in both organs revealed no significant alterations in the dissociation constant (Kd) and in the maximal binding capacity (Bmax) between SHAM flight and control groups. The Kd of the beta-adrenoceptors in the cardiac atria of the SHAM flight rats (74 +/- 5 pm) was significantly higher (p < 0.05) than in those of the ADX flight rats (60 +/- 3 pm) while the Bmax was nonsignificantly higher (1925 +/- 370 in SHAM flight rats vs. 1482 +/- 283 fmol x mg(-1) protein in ADX flight rats). No significant change was determined for the Bmax and Kd values in the kidneys of the ADX and SHAM flight rats. CONCLUSIONS: This work performed on animals did not show any obvious effect of microgravity on the beta-adrenergic function in the heart and kidneys. Inflight rodent sacrifice protocols should definitely ensure assessment of the influence of microgravity on the animals. PMID- 10519484 TI - Survey of USAF flight surgeons regarding clinical preventive services, using CHD as an indicator. AB - BACKGROUND: A recent Department of Defense study revealed that nonpharmacologic therapy is not well documented in medical records of individuals identified at risk for coronary heart disease (CHD). Exercise and weight control are often underemphasized relative to dietary and medication interventions, even in medical journal review articles on management of dyslipidemia. METHODS: A literature review of interventions consisting of exercise alone or with diet is presented. A brief survey was developed to assess the knowledge and beliefs of USAF flight surgeons regarding training received for, and delivery of, clinical preventive services. In addition, the respondents were given a scenario patient with high cholesterol to manage. RESULTS AND DISCUSSION: The literature review demonstrates the beneficial effect of exercise alone or with dietary interventions on reducing total cholesterol and increasing high-density lipoprotein (HDL). The survey reveals that USAF flight surgeons believe that more preventive medicine training is needed in the USAF flight surgeon course, graduate medical education, and medical school. Given a scenario patient with hypercholesterolemia, the flight surgeons believe that nonpharmacologic therapy with consults to the base Health and Wellness Center (HAWC), along with follow-up by the physician, is appropriate initial medical treatment rather than pharmacologic therapy. CONCLUSIONS: The primary treatment for prevention of CHD should be an organized exercise and diet program. This treatment is proven effective through clinical trials and is supported by the flight surgeon survey results. Additional benefits of an organized exercise and diet program include decreased obesity with its associated complications, decreased incidence of hypertension, decreased cancer risk, and decreased risk for diabetes mellitus. PMID- 10519485 TI - Record and around the world flights through Singapore by women. AB - Women's efforts to fly as military pilots during World War I were unsuccessful. However, their numbers and achievements increased and, in 1929, they formed the Ninety-Nines and held the first Women's Air Derby, where male pilots escorted them and mechanics were available. In May 1930, England's Amy Johnson became the first woman to fly solo from England to Australia. She was flying for the Air Transport Auxiliary in England when she was killed in an aircraft accident in 1941. New Zealander Jean Batten set new records over the same course in May 1934, from England to New Zealand in Oct. 1936 and from Australia to England in Oct. 1937. Routes were typically via Singapore as were round-the-world efforts by Amelia Earhart (1937), Joan Merriam Smith (1964), Sheila Scott (1966), and Ann Pellegreno (1967). Accounts frequently mention severe weather including monsoon storms. Adequacy of flight planning has been questioned for some. Jerrie Mock, the first woman to fly around the world alone, in 1964, was asked by Saigon Radio if she had a man aboard. Further successes and independence for women in aviation have been slow, but steady. PMID- 10519486 TI - You're the flight surgeon. Carbon monoxide poisoning. PMID- 10519487 TI - The use of virtual reality in spatial disorientation training. PMID- 10519488 TI - Obstetric anesthesia: what have you done for us lately? PMID- 10519489 TI - Anesthetic management of the elderly: measuring function beyond the immediate perioperative horizon. PMID- 10519490 TI - Fast tracking into the new millennium: an evolving paradigm. PMID- 10519491 TI - The importance of professional obligations: nonpatient care obligations of anesthesiologists. PMID- 10519492 TI - Warren Zapol: true-life adventures in science, medicine, and innovation: recipient of the 1999 Excellence in Research Award. PMID- 10519493 TI - Is combined spinal-epidural analgesia associated with more rapid cervical dilation in nulliparous patients when compared with conventional epidural analgesia? AB - BACKGROUND: The combined spinal-epidural technique provides rapid onset of labor analgesia and, anecdotally, is associated with labors of shorter duration. Epidural analgesia, by contrast, has been suggested to prolong labor modestly. It is unclear, however, whether more rapid cervical dilation in patients who receive combined spinal-epidural analgesia is a physiologic effect of the technique or an artifact of patient selection. The authors hypothesized that anesthetic technique may influence the rate of cervical dilation, and we compared the effects of combined spinalepidural with those of epidural analgesia on the rate of cervical dilation. METHODS: One hundred healthy nulliparous parturients in spontaneous labor with singleton, vertex, full-term fetuses were enrolled in a double-blinded manner when their cervical dilation was less than 5 cm. The patients were randomly assigned to receive analgesia via a standardized combined spinal epidural (n = 50) or epidural (n = 50) technique. Data were collected on cervical dilation, pain, sensory level, and motor blockade. RESULTS: When regional analgesia was induced in comparable groups at a mean of 3 cm cervical dilation, the mean initial cervical dilation rates were significantly faster in the combined spinal-epidural group (mean values, 2.1 +/- 2.1 cm/h vs. 1 +/- 1 cm/h; P = 0.0008). Five parturients in the combined spinal-epidural group had a very rapid (> 5 cm/h) rate of mean initial cervical dilation, compared with none of the women in the epidural group. Overall mean cervical dilation rates in patients who achieved full cervical dilation were 2.3 +/- 2.6 cm/h and 1.3 +/- 0.71 cm/h (P = 0.0154) in the combined spinal-epidural and epidural groups, respectively. CONCLUSIONS: In healthy nulliparous parturients in early labor, combined spinal epidural analgesia is associated with more rapid cervical dilation compared with epidural analgesia. Further study is needed to elicit the cause and overall effect of this difference. PMID- 10519494 TI - Randomized trial of hypotensive epidural anesthesia in older adults. AB - BACKGROUND: Data are sparse on the incidence of postoperative cognitive, cardiac, and renal complications after deliberate hypotensive anesthesia in elderly patients. METHODS: This randomized, controlled clinical trial included 235 older adults with comorbid medical illnesses undergoing elective primary total hip replacement with epidural anesthesia. The patients were randomly assigned to one of two levels of intraoperative mean arterial blood pressure management: either to a markedly hypotensive mean arterial blood pressure range of 45-55 mmHg or to a less hypotensive range of 55-70 mmHg. Cognitive outcome was assessed by within patient change on 10 neuropsychologic tests assessing memory, psychomotor, and language skills from before surgery to 1 week and 4 months after surgery. Prospective standardized surveillance was performed for cardiovascular and renal outcomes, delirium, thromboembolism, and blood loss and replacement. RESULTS: The two groups were similar at baseline in terms of age (mean, 72 yr), sex (50% women), comorbid conditions, and cognitive function. After operation, no significant differences in the incidence of early or long-term cognitive dysfunction were observed between the two blood pressure management groups. There were no significant differences in the rates of other adverse consequences, including cardiac, renal, and thromboembolic complications. In addition, no differences occurred in the duration of surgery, intraoperative estimated blood loss, or transfusion rates. CONCLUSIONS: Elderly patients can safely receive controlled hypotensive epidural anesthesia with this protocol. There was no evidence of greater risks, or early benefits, with the use of the more markedly hypotensive range. PMID- 10519495 TI - Risk factors of delayed extubation, prolonged length of stay in the intensive care unit, and mortality in patients undergoing coronary artery bypass graft with fast-track cardiac anesthesia: a new cardiac risk score. AB - BACKGROUND: Risk factors of delayed extubation, prolonged intensive care unit (ICU) length of stay (LOS), and mortality have not been studied for patients administered fast-track cardiac anesthesia (FTCA). The authors' goals were to determine risk factors of outcomes and cardiac risk scores (CRS) for CABG patients undergoing FTCA. METHODS: Consecutive CABG patients undergoing FTCA were prospectively studied. Outcome variables were delayed extubation > 10 h, prolonged ICU LOS > 48 h, and mortality. Univariate analyses were performed followed by multiple logistic regression to derive risk factors of the three outcomes. Simplified integer-based CRS were derived from logistic models. Bootstrap validation was performed to assess and compare the predictive abilities of CRS and logistic models for the three outcomes. RESULTS: The authors studied 885 patients. Twenty-five percent had delayed extubation, 17% had prolonged ICU LOS, and 2.6% died. Risk factors of delayed extubation were increased age, female gender, postoperative use of intraaortic balloon pump, inotropes, bleeding, and atrial arrhythmia. Risk factors of prolonged ICU LOS were those of delayed extubation plus preoperative myocardial infarction and postoperative renal insufficiency. Risk factors of mortality were female gender, emergency surgery, and poor left ventricular function. CRSs were modeled for the three outcomes. The area under the receiver operating characteristic curve for the CRS-logistic models was not significantly different: 0.707/0.702 for delayed extubation, 0.851/0.855 for prolonged ICU LOS, and 0.657/0.699 for mortality. CONCLUSION: In CABG patients undergoing FTCA, the authors derived and validated risk factors of delayed extubation, prolonged ICU LOS, and mortality. Furthermore, they developed a simplified CRS system with similar predictive abilities as the logistic models. PMID- 10519496 TI - Unintended inhalation of nitric oxide by contamination of compressed air: physiologic effects and interference with intended nitric oxide inhalation in acute lung injury. AB - BACKGROUND: Compressed air from a hospital's central gas supply may contain nitric oxide as a result of air pollution. Inhaled nitric oxide may increase arterial oxygen tension and decrease pulmonary vascular resistance in patients with acute lung injury and acute respiratory distress syndrome. Therefore, the authors wanted to determine whether unintentional nitric oxide inhalation by contamination of compressed air influences arterial oxygen tension and pulmonary vascular resistance and interferes with the therapeutic use of nitric oxide. METHODS: Nitric oxide concentrations in the compressed air of a university hospital were measured continuously by chemiluminescence during two periods (4 and 2 weeks). The effects of unintended nitric oxide inhalation on arterial oxygen tension (n = 15) and on pulmonary vascular resistance (n = 9) were measured in patients with acute lung injury and acute respiratory distress syndrome by changing the source of compressed air of the ventilator from the hospital's central gas supply to a nitric oxide-free gas tank containing compressed air. In five of these patients, the effects of an additional inhalation of 5 ppm nitric oxide were evaluated. RESULTS: During working days, compressed air of the hospital's central gas supply contained clinically effective nitric oxide concentrations (> 80 parts per billion) during 40% of the time. Change to gas tank-supplied nitric oxide-free compressed air decreased the arterial oxygen tension by 10% and increased pulmonary vascular resistance by 13%. The addition of 5 ppm nitric oxide had a minimal effect on arterial oxygen tension and pulmonary vascular resistance when added to hospital-supplied compressed air but improved both when added to tank-supplied compressed air. CONCLUSIONS: Unintended inhalation of nitric oxide increases arterial oxygen tension and decreases pulmonary vascular resistance in patients with acute lung injury and acute respiratory distress syndrome. The unintended nitric oxide inhalation interferes with the therapeutic use of nitric oxide. PMID- 10519497 TI - Effect of mivazerol on perioperative cardiac complications during non-cardiac surgery in patients with coronary heart disease: the European Mivazerol Trial (EMIT). AB - BACKGROUND: Mivazerol is a drug with alpha2-agonist properties that reduces post ganglionic noradrenaline availability and spinal efferent sympathetic output. METHODS: A double-blind randomized placebo-controlled trial was conducted in 61 European centers during a 2.5-yr period on 2,854 patients: 1,897 with coronary heart disease and 957 patients without overt coronary heart disease but classified as at high risk for it. The present analysis was restricted to those patients with previous known coronary heart disease of whom 48% had vascular surgery, 32% non-vascular thoracic or abdominal surgery, and 20% orthopedic surgery. Mivazerol or placebo were given intravenously from the induction of anesthesia for up to 72 h. RESULTS: In the 1,897 patients with established coronary heart disease, mivazerol did not reduce the primary endpoint--the combination of myocardial infarction or death--or all-cause deaths significantly. A preplanned subgroup analysis of 904 patients with known coronary heart disease undergoing vascular surgery showed that there were fewer primary endpoints in those receiving mivazerol (risk ratio [RR], 0.67; 95% CL, 0.45-0.98; P = 0.037) and fewer cardiac deaths (6 of 454 vs. 18 of 450: RR, 0.33; 95% confidence limits, 0.13-0.82; P = 0.017). The all-cause death rate was also decreased (RR, 0.41; 95% CL, 0.18-0.91; P = 0.024), although there was no significant reduction in myocardial infarction. CONCLUSION: The alpha2-adrenergic agonist, mivazerol, did not alter the rates of myocardial infarction or cardiac death in patients with known coronary heart disease undergoing noncardiac surgery. However, it may have protected patients undergoing vascular surgery from further coronary events, and a specific study of such patients is now indicated. PMID- 10519498 TI - Effect of epinephrine on lidocaine clearance in vivo: a microdialysis study in humans. AB - BACKGROUND: Local anesthetic nerve block prolonged by epinephrine is thought to result from local vasoconstriction and consequent decreased local anesthetic clearance from the injection site. However, no study has yet confirmed this directly in humans by measuring tissue concentrations of local anesthetic over time. In addition, recent studies have shown that the alpha2-adrenergic receptor agonist, clonidine, also prolongs nerve block without altering local anesthetic clearance. Because epinephrine is also an alpha2-adrenergic receptor agonist, it is possible that epinephrine prolongs local anesthetic block by a pharmacodynamic mechanism and not a pharmacokinetic one. This study was designed to address this issue. METHODS: Microdialysis probes were placed adjacent to the superficial peroneal nerve in both feet of eight volunteers. Plain lidocaine (1%) was injected along one peroneal nerve and lidocaine with epinephrine (2.5 microg/ml) was injected along the other nerve in a double-blinded, randomized manner. The concentration of lidocaine in tissue was measured at 5-min intervals, and sensory block and cutaneous blood flow were assessed by laser Doppler at 10-min intervals for 5 h. The resulting data for lidocaine concentration versus time were fit to a two-compartment model using modeling software. RESULTS: Epinephrine prolonged sensory block by decreasing local blood flow and slowing clearance. There was no evidence of a pharmacodynamic effect of epinephrine. CONCLUSION: Although epinephrine activates alpha2-adrenergic receptors, its mechanism for prolonging the duration of local anesthetic block rests on its ability to decrease local anesthetic clearance and not on a pharmacodynamically mediated potentiation of local anesthetic effect. PMID- 10519499 TI - Randomized trial of informed consent and recruitment for clinical trials in the immediate preoperative period. AB - BACKGROUND: The standard process of obtaining informed consent sometimes prevents physicians or patients from participating in clinical trials, partly because they are concerned about eventual treatment allocation or the physician is concerned the patient might harbor some uncertainty about the best treatment. Alternative randomization methods have been advocated that may address these and other concerns. METHODS: After institutional ethics committee gave its approval, the authors interviewed 770 patients before operation and asked them to consider enrolling in a mock anesthesia trial. Patients were allocated randomly to one of five methods of randomization and consent: one-sided informed consent (the most common approach), prerandomized consent to experimental treatment, prerandomized consent to standard treatment, one-sided physician-modified informed consent, or one-sided patient-modified informed consent. Recruitment rates were compared and sociodemographic and perioperative predictors of recruitment were identified. RESULTS: The randomization method did not result in any significant difference in recruitment rates: one-sided informed consent, 55.6%; prerandomized consent to experimental treatment, 53.3%; prerandomized consent to standard treatment, 53%; one-sided physician-modified informed consent, 60.7%; and one-sided patient modified informed consent, 56.7% (P = 0.66). Multivariate predictors of recruitment were patient age >45 yr (odds ratio, 1.44; 95% confidence interval [CI], 1.08 to 1.93), English-speaking at home (1.49; 1.0 to 2.21), and male researcher-male patient interaction (1.37; 1.20 to 1.57). CONCLUSIONS: No evidence emerged that alternative randomization and consent designs resulted in increased recruitment rates compared with simple one-sided informed consent for a sham anesthesia trial in patients awaiting elective surgery. Older, male patients were more likely to provide consent. PMID- 10519500 TI - Postoperative pain facilitates nonthermoregulatory tremor. AB - BACKGROUND: Spontaneous tremor is relatively common in normothermic patients after operation and has been attributed to many causes. The hypothesis that nonthermoregulatory shivering-like tremor is facilitated by postoperative pain was tested. In addition, the effects of intravenous lidocaine on nonthermoregulatory tremor were evaluated. METHODS: Patients undergoing knee surgery were anesthetized with 2 microg/kg intravenous fentanyl and 0.2 mg/kg etomidate. Anesthesia was maintained with 1.7 +/- 0.8% (mean +/- SD) isoflurane. Intraoperative forced-air heating maintained normothermia The initial 44 patients were randomly allocated to receive an intra-articular injection of 20 ml saline (n = 23) or lidocaine, 1.5% (n = 21). The subsequent 30 patients were randomly allocated to receive an intravenous bolus of 250 microg/kg lidocaine followed by an infusion of 13 microg x kg(-1) x h(-1) lidocaine or an equivalent volume of saline when shivering was observed. Patient-controlled analgesia was provided for all patients: 3.5 mg piritramide, with a lockout interval of 5 min, for an unlimited total dose. Shivering was graded by a blinded investigator using a four point scale. Pain was assessed by a 100-mm visual analog scale (0 = no pain and 100 = worst pain). The arteriovenous shunt status was evaluated with forearm minus-fingertip skin-temperature gradients. RESULTS: Morphometric characteristics and hemodynamic responses were similar in the four groups. Core and mean skin temperature remained constant or increased slightly compared with preoperative values, and postoperative skin-temperature gradients were negative (indicating vasodilation) in nearly all patients. After intra-articular injection of saline, pain scores for the first postoperative hour averaged 46 +/- 32 mm (mean +/- SD), and 10 of the 23 (43%) patients shivered. In contrast, the pain scores of patients who received intra-articular lidocaine were significantly reduced to 5 +/- 9 mm and shivering was absent in this group (P < 0.05). In the second portion of the study, neither intravenous lidocaine nor saline reduced the magnitude or duration of nonthermoregulatory tremor or the patients' pain scores. CONCLUSIONS: Intra-articular, but not intravenous, lidocaine reduced surgical pain and prevented nonthermoregulatory shivering. Therefore, these data indicate that postoperative pain facilitates nonthermoregulatory shivering. PMID- 10519501 TI - Correlation between cerebral oxygen saturation measured by near-infrared spectroscopy and jugular oxygen saturation in patients with severe closed head injury. AB - Near-infrared spectroscopy has been used to monitor cerebral oxygen saturation during cerebral circulatory arrest and carotid clamping. However, its utility has not been demonstrated in more complex situations, such as in patients with head injuries. The authors tested this method during conditions that may alter the arteriovenous partition of cerebral blood in different ways. METHODS: The authors compared changes in measured cerebral oxygen saturation and other hemodynamic parameters, including jugular venous oxygen saturation, in nine patients with severe closed head injury during manipulation of arterial carbon dioxide partial pressure and after mean arterial pressure was altered by vasopressors. RESULTS: The Bland and Altman representation of cerebral oxygen saturation versus jugular oxygen saturation showed a uniform scatter. Values for changing arterial carbon dioxide partial pressure were: bias = 1.1%, 2 SD = +/-21%, absolute value; and those for alterations in mean arterial pressure: bias = 3.7%, 2 SD = +/-24%, absolute value. However, a Bland and Altman plot of changes in cerebral oxygen saturation versus changes in jugular oxygen saturation had a negative slope (alteration in arterial carbon dioxide partial pressure: bias = 2.4%, 2 SD = +/ 17%, absolute value; alteration in mean arterial pressure: bias = -4.9%, 2 SD = +/-31%, absolute value). Regression analysis showed that changes in cerebral oxygen saturation were positively correlated with changes in jugular venous oxygen saturation during the carbon dioxide challenge, whereas correlation was negative during the arterial pressure challenge. CONCLUSIONS: Cerebral oxygen saturation assessed by near-infrared spectroscopy does not adequately reflect changes in jugular venous oxygen saturation in patients with severe head injury. Changes in arteriovenous partitioning, infrared-spectroscopy contamination by extracerebral signal, algorithm errors, and dissimilar tissue sampling may explain these findings. PMID- 10519502 TI - Comparable postoperative pulmonary atelectasis in patients given 30% or 80% oxygen during and 2 hours after colon resection. AB - BACKGROUND: High concentrations of inspired oxygen are associated with pulmonary atelectasis but also provide recognized advantages. Consequently, the appropriate inspired oxygen concentration for general surgical use remains controversial. The authors tested the hypothesis that atelectasis and pulmonary dysfunction on the first postoperative day are comparable in patients given 30% or 80% perioperative oxygen. METHODS: Thirty patients aged 18-65 yr were anesthetized with isoflurane and randomly assigned to 30% or 80% oxygen during and for 2 h after colon resection. Chest radiographs and pulmonary function tests (forced vital capacity and forced expiratory volume) were obtained preoperatively and on the first postoperative day. Arterial blood gas measurements were obtained intraoperatively, after 2 h of recovery, and on the first postoperative day. Computed tomography scans of the chest were also obtained on the first postoperative day. RESULTS: Postoperative pulmonary mechanical function was significantly reduced compared with preoperative values, but there was no difference between the groups at either time. Arterial gas partial pressures and the alveolar-arterial oxygen difference were also comparable in the two groups. All preoperative chest radiographs were normal. Postoperative radiographs showed atelectasis in 36% of the patients in the 30%-oxygen group and in 44% of those in the 80%-oxygen group. Relatively small amounts of pulmonary atelectasis (expressed as a percentage of total lung volume) were observed on the computed tomography scans, and the percentages (mean +/- SD) did not differ significantly in the patients given 30% oxygen (2.5% +/- 3.2%) or 80% oxygen (3.0% +/- 1.8%). These data provided a 99% chance of detecting a 2% difference in atelectasis volume at an alpha level of 0.05. CONCLUSIONS: Lung volumes, the incidence and severity of atelectasis, and alveolar gas exchange were comparable in patients given 30% and 80% perioperative oxygen. The authors conclude that administration of 80% oxygen in the perioperative period does not worsen lung function. Therefore, patients who may benefit from generous oxygen partial pressures should not be denied supplemental perioperative oxygen for fear of causing atelectasis. PMID- 10519503 TI - Pharmacokinetics and pharmacodynamics of vecuronium in rats with systemic inflammatory response syndrome: treatment with NG-monomethyl-L-arginine. AB - BACKGROUND: Insufficient detoxification caused by nitric oxide-related inhibition of cytochrome P450 may be important for metabolism of numerous drugs, including vecuronium. The present study investigated the pharmacodynamics and pharmacokinetics of vecuronium in rats with inflammatory liver dysfunction. METHODS: Male Sprague-Dawley rats (n = 56) were randomly allocated into two groups: In the sepsis group, liver inflammation was established by injection of 56 mg/kg heat-killed Corynebacterium parvum; control rats received the solvent. At day 4, groups were subdivided according to treatment with the nitric oxide synthase inhibitor N(G)-monomethyl-L-arginine (250 mg/kg) or placebo. The aminopyrine breath test was performed to assess cytochrome P450 activity. Rats were anesthetized with propofol and mechanically ventilated. Duration of action of vecuronium (1.2 mg/kg) was measured by evoked mechanomyography (stimulation of the sciatic nerve, contraction of the gastrocnemius muscle). In seven rats of each subgroup a 50% neuromuscular blockade was established by a continuous vecuronium infusion. Vecuronium plasma levels were measured and plasma clearance of vecuronium was calculated. Nitric oxide synthesis was assessed by measuring nitrite/nitrate serum levels. RESULTS: In sepsis/placebo rats, vecuronium-induced neuromuscular blockade was prolonged (144% of contro/placebo), vecuronium plasma levels at 50% neuromuscular blockade were increased (122% of control/placebo), and plasma clearance was decreased (68% of control/placebo). N(G)-monomethyl-L arginine therapy in rats with sepsis improved cytochrome P450 activity and plasma clearance of vecuronium, shortened duration of action of vecuronium, but did not alter the elevated vecuronium plasma levels. CONCLUSIONS: A systemic inflammatory response syndrome with liver dysfunction results in decreased sensitivity to and a decreased elimination of vecuronium. Modulation of nitric oxide synthesis may be a strategy that can be used in the future to improve xenobiotic metabolism in sepsis. PMID- 10519504 TI - Evaluation of interaction between gabapentin and ibuprofen on the formalin test in rats. AB - BACKGROUND: Gabapentin is active in the regulation of facilitated pain states evoked by tissue injury. The mechanism of this action is believed to be through a specific binding site, likely at the spinal level. Nonsteroidal antiinflammatory drugs have a comparable behavioral profile, although their actions are believed to be mediated by cyclooxygenase inhibition at the spinal level. This study was undertaken to determine the nature of the interaction of these two mechanistically distinct antihyperalgesic agents in rats in a model of facilitated processing, the formalin test. METHODS: The effects of intraperitoneal gabapentin and ibuprofen were examined on flinching behavior and cardiovascular response (mean arterial blood pressure [MABP] and heart rate measured in the tail artery) evoked by the injection of formalin (5%; 50 microl). Their interaction was characterized using an isobolographic analysis. RESULTS: Injection of formalin into the hind paw caused a biphasic flinching and parallel increases in MABP. Gabapentin and ibuprofen produced a limited effect on the flinching in phase 1, but both drugs produced dose-dependent suppression of the flinching observed during phase 2 (gabapentin ED50 = 88 mg/kg; ibuprofen ED50 = 19 mg/kg). Gabapentin similarly showed a dose-dependent suppression of the MABP and heart rate response only during phase 2; ibuprofen showed dose-dependent reduction of MABP response in both phases. The isobolographic analysis carried out using equipotent dose ratios in phase 2 revealed an additive interaction between the two drugs. Neither gabapentin nor ibuprofen affected the baseline cardiovascular measures. CONCLUSION: Gabapentin and ibuprofen independently alter the facilitated state as measured by somatomotor and autonomic response. Together these agents interact in an additive fashion if delivered concurrently. This combination may prove useful in managing postinjury pain states in humans. PMID- 10519505 TI - Local anesthetics inhibit muscarinic receptor-mediated activation of extracellular signal-regulated kinases in rat pheochromocytoma PC12 cells. AB - BACKGROUND: Because protein phosphorylation is a key mechanism for controlling cellular functions and extracellular signal-regulated kinase (ERK) plays a role in cellular signal transduction, the authors wanted to determine whether local anesthetics interfere with biochemical signaling molecules. METHODS: Protein tyrosine phosphorylation and ERK activation induced by carbachol, an agonist for muscarinic acetylcholine receptors, were examined in rat pheochromocytoma PC12 cells, a model for investigating signal transduction. Carbachol-induced tyrosine phosphorylated proteins of 44 and 42 kd were determined by Western blot analysis and identified as activated ERK1 and ERK2 using anti-ERK antibody. The ERK activation was blocked by preincubation with atropine or an M3 muscarinic acetylcholine receptor antagonist 4-diphenyacetooxy-1, 1-dimethylpiperidinium, indicating that is was mediated by M3 muscarinic acetylcholine receptor activation. Then, in the presence of local anesthetic, the carbachol-induced tyrosine phosphorylation and ERK activation were evaluated. The effects of three Na+ current-modifying reagents on carbachol-induced ERK activation were also evaluated. RESULTS: Procaine (10(-4) to 10(-3) M) inhibited carbachol-induced tyrosine phosphorylation and ERK activation in a concentration-dependent manner. Although tetracaine, lidocaine, and bupivacaine similarly suppressed carbachol induced tyrosine phosphorylation and ERK activation, neither tetrodotoxin, veratridine, nor ouabain affected the carbachol-induced ERKs activation. Both ERKs were also activated by 4beta-phorbol 12-myristate 13-acetate, an activator of protein kinase C, and fluoroaluminate (AlF4-), respectively, but procaine did not affect ERK activation induced by these two substances. The inhibition of carbachol-induced ERK activation by procaine was not modified by a phosphatase inhibitor, calyculin A. CONCLUSIONS: The current results indicate that local anesthetics inhibit the activity of the signal-transducing molecule(s) leading to M3 muscarinic acetylcholine receptor-mediated ERK activation in PC12 cells. Such action is unlikely to be a result of the drug's action on Na+ channels or on the electrochemical gradients of the neuronal cell membrane. PMID- 10519506 TI - Tooth pulp- and facial hair mechanoreceptor-evoked responses of trigeminal sensory neurons are attenuated during ketamine anesthesia. AB - BACKGROUND: Evidence exists that ketamine, administered systemically using a dose required for inducing a state of anesthesia, may antagonize nociceptive but not innocuous input to lumbar dorsal horn neurons. However, it is unclear whether ketamine exerts this selective action on sensory inputs to trigeminal sensory neurons. The current study was undertaken to compare the responses evoked in trigeminal sensory neurons by electrical stimuli applied to the tooth pulp versus air-puff stimuli applied to facial hair mechanoreceptors (FHMs) during quiet wakefulness versus ketamine anesthesia. METHODS: Accordingly, responses of rostral trigeminal sensory nuclear complex (TSNC) and trigeminothalamic tract neurons evoked by tooth pulp (a source of small-diameter fiber input) and FHMs (a source of larger-diameter fiber input) were recorded extracellularly from chronically instrumented cats before, during, and after recovery from the anesthetic state induced by a single (2.2 mg/kg) intravenous injection of ketamine. RESULTS: Overall, tooth pulp-evoked responses of TSNC neurons were maximally suppressed by 50% within 5 min after the intravenous administration of ketamine. Ketamine also suppressed the FHM-evoked responses of TSNC and trigeminothalamic neurons by 45%. The time course of ketamine's suppressive action was equivalent for tooth pulp- and FHM-evoked responses. However, the recovery of tooth pulp-evoked TSNC neuronal responses at suprathreshold intensities was markedly prolonged compared with neuronal responses driven by threshold stimuli or FHM. CONCLUSIONS: These electrophysiologic results in the chronically instrumented cat preparation indicate that a nonselective suppression of orofacial somatosensory information occurs during ketamine anesthesia. The prolonged recovery of suprathreshold responses of TSNC neurons mediated by small diameter afferent fiber input may partly underlie the analgesic action of ketamine that is clinically relevant at subanesthetic doses. PMID- 10519507 TI - Overexpression of bcl-xL protects astrocytes from glucose deprivation and is associated with higher glutathione, ferritin, and iron levels. AB - BACKGROUND: The possibility of altering outcome from ischemia-like injury by overexpressing the anti-cell death gene bcl-xL was studied. Cells are known to die by different pathways including apoptosis, or programmed cell death, and necrosis. The bcl-xL gene is a member of a family of apoptosis regulating genes and often displays the death-inhibiting properties of the prototype of this family, bcl-2. It is of special interest to study bcl-xL for possible brain protection, because, unlike bcl-2, it is important for normal brain development. METHODS: Overexpression of bcl-xL was achieved in primary astrocyte cultures using a retroviral vector. Cultures of astrocytes overexpressing bcl-xL or a control gene were injured by hydrogen peroxide, glucose deprivation, or combined oxygen and glucose deprivation. Outcome was assessed morphologically and by release of lactate dehydrogenase. We assessed antioxidant effects by measuring glutathione using monochlorobimane, ferritin by immunoblotting, the level of iron spectrophotometrically, and superoxide using iodonitrotetrazolium violet and dihydroethidium. RESULTS: Protection by bcl-xL was found against glucose deprivation and hydrogen peroxide exposure but not combined oxygen and glucose deprivation. Higher levels of superoxide were found, without increased levels of lipid peroxidation. Overexpression of bcl-xL was associated with elevated glutathione levels, elevated ferritin levels, and increased amounts of iron. The increased glutathione contributed to the protection from glucose deprivation. CONCLUSIONS: Overexpression of bcl-xL protects astrocytes from oxidative injury with the same spectrum of protection seen previously for bcl-2. The increased antioxidant defense observed should be beneficial against both apoptotic and necrotic cell death. The effects on levels of ferritin and iron are novel and identify a new area of interest for this gene family. Whether this relates to the effects of these genes on mitochondrial function remains to be elucidated. PMID- 10519508 TI - Xenon does not trigger malignant hyperthermia in susceptible swine. AB - BACKGROUND: Xenon is a noble gas with anesthetic properties currently under investigation for use in humans. This study was performed to evaluate whether xenon may trigger malignant hyperthermia in susceptible swine. METHODS: Nine malignant hyperthermia-sensitive swine (Pietrain) were initially anesthetized with pentobarbital and then ventilated with 70% xenon in oxygen for 2 h. Heart rate, mean arterial pressure, cardiac output, body temperature, arterial and mixed-venous blood gases, and plasma catecholamine and lactate levels were measured every 10 min both during xenon-oxygen ventilation and after a 30-min xenon washout phase followed by subsequent administration of halothane (1% inspired) and succinylcholine (3 mg/kg intravenous). During the investigation, no malignant hyperthermia-specific therapy was instituted. RESULTS: Xenon exposure did not induce any changes in metabolic and hemodynamic parameters nor elevations of the plasma catecholamine levels indicative for an episode of malignant hyperthermia. By contrast, in all animals, within 20 min after the administration of halothane and succinylcholine, fulminant and fatal malignant hyperthermia episodes were initiated. CONCLUSIONS: The authors conclude that xenon does not trigger malignant hyperthermia in susceptible swine. PMID- 10519509 TI - Direct inhibition of the N-methyl-D-aspartate receptor channel by high concentrations of opioids. AB - BACKGROUND: Electrophysiologic and receptor binding studies showed that some opioids have noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist properties. METHODS: The effects and mechanisms of action of various opioid compounds were examined on four kinds of heteromeric NMDA receptor channels, namely the epsilon1/zeta1, epsilon2/zeta1, epsilon3/zeta1, and epsilon4/zeta1 channels, expressed in Xenopus oocytes. Furthermore, the action sites of opioids on NMDA receptor channels were investigated by site-directed mutagenesis. RESULTS: Meperidine inhibited four kinds of channels to a similar extent with inhibitor concentrations for half-control response (IC50s) of 210-270 microM. Morphine, fentanyl, codeine, and naloxone also inhibited NMDA receptor channels with affinities comparable to meperidine. Opioid inhibition exhibited voltage dependence and was quite effective at negative potentials. Opioids also shifted the inhibition curve of Mg2+ to the right. Furthermore, replacement of the conserved asparagine residue with glutamine in the channel-lining segment M2 of the zeta1 subunit, which constitutes the block sites of Mg2+ and ketamine, reduced the sensitivity to opioids, whereas that of the epsilon2 subunit barely affected the opioid sensitivity. CONCLUSIONS: These results, together with previous findings, suggest that the low-affinity NMDA receptor antagonist activity is a common characteristic of various opioid compounds, and that the inhibition is a result of channel-block mechanisms at the site, which partially overlaps with those of Mg2+ and ketamine. This antagonist property of opioids may be clinically significant in the spinal cord following epidural or intrathecal administration, after which the cerebrospinal fluid concentrations of some opioids reach the high micromolar level. PMID- 10519510 TI - Quantitative and qualitative effects of isoflurane on movement occurring after noxious stimulation. AB - BACKGROUND: Anesthetic potency is assessed by determination of the anesthetic concentration that prevents gross, purposeful movement in response to noxious stimulation. It is unclear whether anesthetics cause a progressive decrease in the number and force of limb movements evoked by noxious stimulation, or a step decrease (consistent with an all-or-none effect at the site of action). The authors hypothesized that isoflurane and halothane would progressively depress the movement response. METHODS: Isoflurane minimum alveolar concentration (MAC) was determined in rats (N = 14) using a clamp applied to a hind paw. Lateral head movements and flexions of the forelimbs and hindlimbs were measured with force transducers. Isoflurane was adjusted to 0.6, 0.9, 1.1, and 1.4 MAC, the noxious stimulus applied, and the force and number of limb and head movements determined. Force and movement determinations were made in seven additional halothane anesthetized rats. RESULTS: Isoflurane MAC was 1.3 +/- 0.1%. In general, if movement occurred after application of the noxious clamp, the head and all limbs were involved. At 0.6 MAC, the median number of extremity and head movements was 3.5 (10th-90th percentile, 2.0-11.4) with force generated per movement (force/movement) = 6.4 (2.0-13.2) N-s. Movement number decreased to 2.1 (0.25 4.2) at 0.9 MAC (P < 0.05), but force/movement was unchanged at 4.5 (0.4-15.1) N s (Newton-second). At 1.1 MAC, movement number and force/movement decreased to 0.2 (0.0-1.5) and 0.1 (0.0-3.2) N-s, respectively (P < 0.005). No significant movement occurred at 1.4 MAC. The halothane-anesthetized rats had similar findings, although at 0.6 MAC they generated more movements (10.5 [5.2-19.8]) than the rats receiving isoflurane (P < 0.05). CONCLUSIONS: The results indicate that increasing anesthetic concentration from 0.6 to 0.9 MAC had little effect on the motor system controlling the force of limb movements, and the neural system generating repeated limb movements was depressed, consistent with a differential anesthetic effect at separate sites. PMID- 10519511 TI - Intrathecal adenosine interacts with a spinal noradrenergic system to produce antinociception in nerve-injured rats. AB - BACKGROUND: Adenosine analogs produce antinociception in animal models of acute pain, reduce hypersensitivity in models of inflammatory and nerve-injury pain, and stimulate neurotransmitter release in the brain. Adenosine itself is entering clinical trials for analgesia, and the current study examined the effect, mechanisms of action, and interaction with noradrenergic systems of intrathecal adenosine in a rat model of neuropathic pain. METHODS: The left L5 and L6 spinal nerve roots were ligated and, 1 week later, an intrathecal catheter was inserted in male rats. Withdrawal threshold to mechanical stimulation of the left hind paw was determined before and after surgery, confirming mechanical hypersensitivity. The effects of intrathecal adenosine, clonidine, and their combination on withdrawal threshold were determined, and reversal of the effects of adenosine by adenosine and alpha2-adrenergic antagonists and by destruction of noradrenergic nerve terminals was tested. Finally, spinal cord slices were perfused in vitro with the adenosine agonist 5'-N-ethylcarboxamide adenosine, and norepinephrine release was measured. RESULTS: Intrathecal adenosine and clonidine reduced hypersensitivity and interacted in an additive manner. The effects of adenosine were blocked by intrathecal injection of A1 but not A2 adenosine receptor antagonists, by an alpha2-adrenergic antagonist, and by destruction of spinal noradrenergic nerve terminals. Perfusion of spinal cord slices with 5'-N ethylcarboxamide adenosine resulted in a concentration-dependent increase in norepinephrine release. CONCLUSION: These data support clinical examination of intrathecal adenosine alone and with clonidine in the treatment of chronic pain states that include a component of mechanical hypersensitivity and suggest that, after nerve injury, adenosine acts to reduce hypersensitivity through spinal norepinephrine release. PMID- 10519512 TI - Inhibitory effects of propofol on acetylcholine-induced, endothelium-dependent relaxation and prostacyclin synthesis in rabbit mesenteric resistance arteries. AB - BACKGROUND: Propofol (2,6-diisopropylphenol) modulates endothelium-dependent relaxation in some arterial preparations. The effect of propofol on endothelium dependent, prostacyclin-mediated responses in mesenteric resistance arteries has not yet been clarified. METHODS: The effect of propofol was examined on acetylcholine-induced membrane potential changes in the presence of N(G)-nitro-L arginine (L-NOARG) in endothelium-intact rabbit mesenteric resistance arteries in vitro. The effects of propofol were also examined on the endothelium-dependent relaxation and prostacyclin synthesis that was induced by acetylcholine in the presence of L-NOARG and nicardipine. The effect of propofol on the relaxation induced by a prostacyclin analogue was examined in strips treated with L-NOARG and diclofenac. RESULTS: Acetylcholine produced an initial and a slow membrane hyperpolarization. Propofol, 10 microM, and diclofenac each inhibited the acetylcholine-induced slow hyperpolarization, but not the initial hyperpolarization. Acetylcholine produced an endothelium-dependent relaxation that was significantly inhibited by propofol, 10 microM, and diclofenac. Propofol, 10 microM, greatly inhibited the acetylcholine-induced synthesis of prostacyclin, as did diclofenac. Propofol, 10 microM, had no effect on the relaxation induced by a prostacyclin analog. CONCLUSIONS: In rabbit mesenteric resistance arteries, propofol inhibits the synthesis of prostacyclin and thus attenuates acetylcholine-induced, endothelium-dependent responses. Our results may help to explain why some actions seen with propofol in some preparations (e.g., vasoconstriction) are not seen after the endothelium is removed. PMID- 10519513 TI - Inhaled nitric oxide: basic biology and clinical applications. PMID- 10519514 TI - Anticoagulation monitoring during cardiac surgery: a review of current and emerging techniques. AB - The literature does not consistently support the importance of anticoagulation monitoring techniques during CPB. This is best reflected by studies that have evaluated the impact of the ACT method on blood loss and transfusion outcomes. Inconsistent findings from studies that evaluated the impact of ACT monitoring may be related to either suboptimal study design (i.e., retrospective, unblinded, nonrandomized) or possibly the diagnostic inprecision of the ACT method used in these studies. There are a small number of well-controlled studies, some of which suggest that bleeding and transfusion outcomes can be improved by refining heparin monitoring techniques, either by sustaining better anticoagulation during CPB or by optimizing protamine doses (i.e., when empiric protocols result in excessive protamine doses). More well-controlled studies are needed to better define the importance of anticoagulation management during CPB. PMID- 10519515 TI - Nonpatient care obligations of anesthesiologists. PMID- 10519516 TI - Management of anesthesia for the pregnant surgical patient. PMID- 10519517 TI - Cardiogenic failure after isolated neurologic injury: a report of two cases. PMID- 10519518 TI - Use of the laryngeal mask for exchange of orotracheal tubes. PMID- 10519519 TI - Delayed-onset epinephrine-induced pulmonary edema. PMID- 10519520 TI - Vagotonia and cardiac arrest during spinal anesthesia. PMID- 10519521 TI - Photosensitivity and perioperative polyneuropathy complicating orthotopic liver transplantation in a patient with erythropoietic protoporphyria. PMID- 10519522 TI - Questions use of nasal cannula for oxygen supplementation during cataract surgery. PMID- 10519523 TI - Cost-efficient end-tidal carbon dioxide monitoring via Hudson-style mask. PMID- 10519524 TI - Carbon dioxide monitoring during deep conscious sedation using nasopharyngeal airways. PMID- 10519525 TI - Inflation of the endotracheal tube cuff in the pharynx for ventilation of paralyzed patients with unanticipated difficult airway. PMID- 10519526 TI - Life-threatening bilateral pneumothorax caused by misconnection of the laser lens cooling system during gynecologic laparoscopy. PMID- 10519527 TI - Web site reviews. Hewlett Packard Anesthesiology Village PMID- 10519528 TI - Wanted: a new order in protein nomenclature. PMID- 10519529 TI - Europe strengthens its hand in bioscience website talks... PMID- 10519530 TI - And India protects its past online. PMID- 10519531 TI - Lasker awards honour three pioneers in ion channels. PMID- 10519532 TI - cDNA sequence databank will safeguard research access. PMID- 10519533 TI - Online archive must serve authors as well as publishers. PMID- 10519534 TI - A view from Kansas on that evolution debate. PMID- 10519535 TI - Wellcome for education on science in society. PMID- 10519537 TI - Medicine and biology are more than biomedicine. PMID- 10519536 TI - Confidentiality is vital to bioweapons control. PMID- 10519538 TI - How many billions to go? PMID- 10519539 TI - Myosin steps backwards. PMID- 10519540 TI - Apoptosis. Cutting red-cell production. PMID- 10519541 TI - Homogeneous catalysis. Controlled green oxidation. PMID- 10519542 TI - The haemoglobin enzyme. PMID- 10519543 TI - A cell for all reasons. PMID- 10519544 TI - Ernst Ludwig Wynder 1922-99. PMID- 10519545 TI - Evolution of an antifreeze glycoprotein. PMID- 10519546 TI - Aquatic sex pheromone from a male tree frog. PMID- 10519547 TI - An enzymatic globin from a marine worm. PMID- 10519548 TI - The origins of insect metamorphosis. AB - Insect metamorphosis is a fascinating and highly successful biological adaptation, but there is much uncertainty as to how it evolved. Ancestral insect species did not undergo metamorphosis and there are still some existing species that lack metamorphosis or undergo only partial metamorphosis. Based on endocrine studies and morphological comparisons of the development of insect species with and without metamorphosis, a novel hypothesis for the evolution of metamorphosis is proposed. Changes in the endocrinology of development are central to this hypothesis. The three stages of the ancestral insect species-pronymph, nymph and adult-are proposed to be equivalent to the larva, pupa and adult stages of insects with complete metamorphosis. This proposal has general implications for insect developmental biology. PMID- 10519549 TI - Dorsoventral lineage restriction in wing imaginal discs requires Notch. AB - The formation of boundaries that prevent the intermixing of cells is an important developmental patterning mechanism. The compartmental lineage restrictions that appear in the developing imaginal discs of Drosophila are striking examples of such boundaries. However, little is known about the cellular mechanism underlying compartmental lineage restrictions. The dorsoventral (D/V) lineage restriction that arises late in the developing wing imaginal disc requires the dorsal expression of the transcription factor Apterous and it has been hypothesized that apterous (ap) maintains compartmentalization by directly regulating the expression of molecules that modify cell adhesion or affinity. However, ap expression also regulates signalling between dorsal and ventral compartments, resulting in high levels of Notch signalling at the D/V boundary. Here we show that the formation of Notch-dependent boundary cells is required for the D/V lineage restriction. PMID- 10519550 TI - Fringe-dependent separation of dorsal and ventral cells in the Drosophila wing. AB - The separation of cells into populations that do not intermix, termed compartments, is a fundamental organizing principle during development. Dorsal ventral compartmentalization of the Drosophila wing is regulated downstream of the apterous (ap) gene, which encodes a transcription factor that specifies dorsal wing fate. fringe (fng) is normally expressed by dorsal cells downstream of ap; here we show that fng plays a key role in dorsal-ventral compartmentalization. Loss of fng function causes dorsal cells to violate the compartment boundary, and ectopic expression of the Fng protein causes ventral cells to violate thecompartment boundary. Fng modulates signalling through the Notch receptor. Notch and its ligands are essential for formation of the dorsal ventral compartment border, and repositioning the stripe of Notch activation that is normally established there appears to reposition the compartment border. However, activation of Notch does not itself confer either dorsal or ventral cell location, and fng can influence compartmentalization even within regions of ubiquitous Notch activation. Our results indicate that the primary mechanism by which fng establishes a compartment border is by positioning a stripe of Notch activation, but also that fng may exert additional influences on compartmentalization. PMID- 10519551 TI - Silencing of TGF-beta signalling by the pseudoreceptor BAMBI. AB - Members of the transforming growth factor-beta (TGF-beta) superfamily, including TGF-beta, bone morphogenetic proteins (BMPs), activins and nodals, are vital for regulating growth and differentiation. These growth factors transduce their signals through pairs of transmembrane type I and type II receptor kinases. Here, we have cloned a transmembrane protein, BAMBI, which is related to TGF-beta family type I receptors but lacks an intracellular kinase domain. We show that BAMBI is co-expressed with the ventralizing morphogen BMP4 (refs 5, 6) during Xenopus embryogenesis and that it requires BMP signalling for its expression. The protein stably associates with TGF-beta-family receptors and inhibits BMP and activin as well as TGF-beta signalling. Finally, we provide evidence that BAMBI's inhibitory effects are mediated by its intracellular domain, which resembles the homodimerization interface of a type I receptor and prevents the formation of receptor complexes. The results indicate that BAMBI negatively regulates TGF-beta family signalling by a regulatory mechanism involving the interaction of signalling receptors with a pseudoreceptor. PMID- 10519552 TI - Tom22 is a multifunctional organizer of the mitochondrial preprotein translocase. AB - Mitochondrial preproteins are imported by a multisubunit translocase of the outer membrane (TOM), including receptor proteins and a general import pore. The central receptor Tom22 binds preproteins through both its cytosolic domain and its intermembrane space domain and is stably associated with the channel protein Tom40 (refs 11-13). Here we report the unexpected observation that a yeast strain can survive without Tom22, although it is strongly reduced in growth and the import of mitochondrial proteins. Tom22 is a multifunctional protein that is required for the higher-level organization of the TOM machinery. In the absence of Tom22, the translocase dissociates into core complexes, representing the basic import units, but lacks a tight control of channel gating. The single membrane anchor of Tom22 is required for a stable interaction between the core complexes, whereas its cytosolic domain serves as docking point for the peripheral receptors Tom20 and Tom70. Thus a preprotein translocase can combine receptor functions with distinct organizing roles in a multidomain protein. PMID- 10519553 TI - Negative regulation of erythropoiesis by caspase-mediated cleavage of GATA-1. AB - The production of red blood cells follows the sequential formation of proerythroblasts and basophilic, polychromatophilic and orthochromatic erythroblasts, and is promoted by the hormone erythropoietin (Epo) in response to tissue hypoxia. However, little is known about the negative regulation of this process. Death receptors are a family of surface molecules that trigger caspase activation and apoptosis in a variety of cell types. Here we show that immature erythroid cells express several death receptors whose ligands are produced by mature erythroblasts. Exposure of erythroid progenitors to mature erythroblasts or death-receptor ligands resulted in caspase-mediated degradation of the transcription factor GATA-1, which is associated with impaired erythroblast development. Expression of a caspase-resistant GATA-1 mutant, but not of the wild type gene, completely restored erythroid expansion and differentiation following the triggering of death receptors, indicating that there is regulatory feedback between mature and immature erythroblasts through caspase-mediated cleavage of GATA-1. Similarly, erythropoiesis blockade following Epo deprivation was largely prevented by the expression of caspase-inhibitory proteins or caspase-resistant GATA-1 in erythroid progenitors. Caspase-mediated cleavage of GATA-1 may therefore represent an important negative control mechanism in erythropoiesis. PMID- 10519554 TI - Cytochrome P450 2C is an EDHF synthase in coronary arteries. AB - In most arterial beds a significant endothelium-dependent dilation to various stimuli persists even after inhibition of nitric oxide synthase and cyclo oxygenase. This dilator response is preceded by an endothelium-dependent hyperpolarization of vascular smooth muscle cells, which is sensitive to a combination of the calcium-dependent potassium-channel inhibitors charybdotoxin and apamin, and is assumed to be mediated by an unidentified endothelium-derived hyperpolarizing factor (EDHF). Here we show that the induction of cytochrome P450 (CYP) 2C8/34 in native porcine coronary artery endothelial cells by beta naphthoflavone enhances the formation of 11,12-epoxyeicosatrienoic acid, as well as EDHF-mediated hyperpolarization and relaxation. Transfection of coronary arteries with CYP 2C8/34 antisense oligonucleotides results in decreased levels of CYP 2C and attenuates EDHF-mediated vascular responses. Thus, a CYP epoxygenase product is an essential component of EDHF-mediated relaxation in the porcine coronary artery, and CYP 2C8/34 fulfils the criteria for the coronary EDHF synthase. PMID- 10519555 TI - Ascaris haemoglobin is a nitric oxide-activated 'deoxygenase'. AB - The parasitic nematode Ascaris lumbricoides infects one billion people worldwide. Its perienteric fluid contains an octameric haemoglobin that binds oxygen nearly 25,000 times more tightly than does human haemoglobin. Despite numerous investigations, the biological function of this molecule has remained elusive. The distal haem pocket contains a metal, oxygen and thiol, all of which are known to be reactive with nitric oxide. Here we show that Ascaris haemoglobin enzymatically consumes oxygen in a reaction driven by nitric oxide, thus keeping the perienteric fluid hypoxic. The mechanism of this reaction involves unprecedented chemistry of a haem group, a thiol and nitric oxide. We propose that Ascaris haemoglobin functions as a 'deoxygenase', using nitric oxide to detoxify oxygen. The structural and functional adaptations of Ascaris haemoglobin suggest that the molecular evolution of haemoglobin can be rationalized by its nitric oxide related functions. PMID- 10519556 TI - A chain initiation factor common to both modular and aromatic polyketide synthases. AB - Antibiotic-producing polyketide synthases (PKSs) are enzymes responsible for the biosynthesis in Streptomyces and related filamentous bacteria of a remarkably broad range of bioactive metabolites, including antitumour aromatic compounds such as mithramycin and macrolide antibiotics such as erythromycin. The molecular basis for the selection of the starter unit on aromatic PKSs is unknown. Here we show that a component of aromatic PKS, previously named 'chain-length factor', is a factor required for polyketide chain initiation and that this factor has decarboxylase activity towards malonyl-ACP (acyl carrier protein). We have re examined the mechanism of initiation on modular PKSs and have identified as a specific initiation factor a domain of previously unknown function named KSQ, which operates like chain-length factor. Both KSQ and chain-length factor are similar to the ketosynthase domains that catalyse polyketide chain extension in modular multifunctional PKSs and in aromatic PKSs, respectively, except that the ketosynthase domain active-site cysteine residue is replaced by a highly conserved glutamine in KSQ and in chain-length factor. The glutamine residue is important both for decarboxylase activity and for polyketide synthesis. PMID- 10519558 TI - Vertebral artery flow and cervical manipulation: an experimental study. AB - BACKGROUND: Spinal manipulation therapy is used by millions of patients each year to relieve symptoms caused by biomechanical dysfunction of the spine. Cerebrovascular accidents in the posterior cerebral circulation are a feared complication, but little research has been done on vertebral artery hemodynamics during cervical manipulation. OBJECTIVE: The purpose of this study was to develop an experimental model for investigations of volume blood flow changes in the vertebral arteries during premanipulative testing of these vessels and during spinal manipulation therapy of the cervical spine. DESIGN AND SETTING: An experimental study in a university biomedical laboratory. MATERIAL AND METHODS: The vertebral arteries were exposed in 8 adult pigs after extensive mediastinal dissection. Volume blood flow was measured on both sides simultaneously by advanced transit-time flowmetry. RESULTS: After cervical manipulation, the vertebral artery volume blood flow increased significantly for 40 seconds before returning to baseline values in less than 3 minutes. We found no significant changes in volume flow during premanipulative testing of the vertebral arteries (DeKleyn's test). CONCLUSION: We present an experimental model for investigations of vertebral artery hemodynamics during biomechanical interventions. We found a modest and transient effect of cervical manipulation on vertebral artery volume flow. The model may have further applications in future biomechanical research, for example, to determine whether any of several spinal manipulative techniques imposes less strain on the vertebral artery, thereby reducing possible future cerebrovascular accidents after such treatment. PMID- 10519557 TI - Myosin VI is an actin-based motor that moves backwards. AB - Myosins and kinesins are molecular motors that hydrolyse ATP to track along actin filaments and microtubules, respectively. Although the kinesin family includes motors that move towards either the plus or minus ends of microtubules, all characterized myosin motors move towards the barbed (+) end of actin filaments. Crystal structures of myosin II (refs 3-6) have shown that small movements within the myosin motor core are transmitted through the 'converter domain' to a 'lever arm' consisting of a light-chain-binding helix and associated light chains. The lever arm further amplifies the motions of the converter domain into large directed movements. Here we report that myosin VI, an unconventional myosin, moves towards the pointed (-) end of actin. We visualized the myosin VI construct bound to actin using cryo-electron microscopy and image analysis, and found that an ADP-mediated conformational change in the domain distal to the motor, a structure likely to be the effective lever arm, is in the opposite direction to that observed for other myosins. Thus, it appears that myosin VI achieves reverse direction movement by rotating its lever arm in the opposite direction to conventional myosin lever arm movement. PMID- 10519559 TI - Preliminary study of the effects of a placebo chiropractic treatment with sham adjustments. AB - OBJECTIVE: To identify aspects of the delivery of placebo chiropractic treatments by using sham adjustments that may cause a treatment effect and that may affect the success of blinding. DESIGN AND SETTING: Two-period crossover design in a chiropractic college research clinic. SUBJECTS: Eighteen volunteer staff, students, and faculty of the chiropractic college who reported low-back pain within the last 6 months. INTERVENTIONS: Flexion-distraction technique was used to perform chiropractic adjustments, and a hand-held instrument (Activator adjusting instrument) with the pressure gauge set on the 0 was used to perform sham adjustments. The treatment period was 2 weeks, with a total of 4 visits. MAIN OUTCOME MEASURES: The Visual Analog Scale (VAS) for pain and Global Well Being Scale (GWBS). RESULTS: Although VAS and GWBS scores improved with both treatments, a somewhat greater improvement occurred in most cases with the active treatment. Eight of 14 patients interviewed believed that the placebo had a treatment effect. CONCLUSION: This study provided preliminary information that was useful in planning the protocol for a placebo chiropractic treatment in the randomized clinical trial for which it was designed. PMID- 10519560 TI - The importance of normalization in the interpretation of surface electromyography: a proof of principle. AB - OBJECTIVE: To demonstrate the errors in surface electromyogram (EMG) interpretation that can be made when the EMG signal is not normalized. DESIGN: A case study as a proof of principle. MAIN OUTCOME MEASURES: The EMG amplitude between the upper and lower portions of the rectus abdominis in one subject during a trunk curl when the EMG signal was normalized (expressed as a percentage of a maximum voluntary contraction) and the amplitude when the signal was expressed in raw, unnormalized arbitrary units or raw millivolts directly read from the instrumentation. RESULTS: Interpretation of the unnormalized EMG signal suggests that a large difference in neural activation of the upper and lower sections of the rectus abdominis is occurring. In this condition the average activity in the lower rectus is 60.9% of that in the upper portion. This interpretation is incorrect. When the EMG signal is normalized, the differences between the upper and lower segments are reduced. When normalized, the lower segment activity is equal to that of the upper segment. CONCLUSIONS: Because of the inherent EMG signal variability, clinical interpretation of surface EMG signals requires normalization of the signal for physiologic interpretation and for comparison between bilateral muscles and between the same muscle on different days and between different subjects. PMID- 10519561 TI - Biological activity of Melaleuca alternifola (Tea Tree) oil component, terpinen-4 ol, in human myelocytic cell line HL-60. AB - BACKGROUND: Tea tree oil is an aboriginal Australian traditional medicine for bruises, insect bites, and skin infections. It was rediscovered in the 1920s as a topical antiseptic that is more effective than Phenol. Previous studies have demonstrated its antiseptic qualities, but its effects on human white blood cells have never been investigated. OBJECTIVE: To test the hypothesis that tea tree oil exerts its antiseptic action through white blood cell activation. METHODS: Crude oil and the purified "active" component were studied by using a model system that responds to bioactive components by induction of differentiation in white blood cells. Methods used included white blood cell oxidative burst assay (nitroblue tetrazolium [NBT] dye reduction); cell proliferation assay (tritiated thymidine incorporation); cell surface differentiation marker assay (flow cytometric quantitation of phycoerythrin-anti-CD 11b binding); cell viability assay (trypan blue exclusion); and cellular differentiation enzyme assay (white cell esterase staining). RESULTS: Collectively, five assays that measure differentiation in white blood cells indicated monocytic differentiation after treatment with either crude oil or the purified active component. Both the crude oil and the purified active component, (+:-) terpinene-4-ol, caused a similar type and amount of differentiation. The culture of cells in medium containing serum caused more activation than in medium containing no serum. CONCLUSION: The antiseptic activity of tea tree oil appears to be due, in part, to white blood cell activation. PMID- 10519562 TI - A psychological profile of fibromyalgia patients: a chiropractic case study. AB - BACKGROUND: Fibromyalgia is a chronic condition characterized by widespread musculoskeletal pain and tenderness. Reversible modulation of the pain threshold is believed to contribute to the pathogenesis of this condition, and psychosocial stress is known to alter the pain threshold. OBJECTIVE: To describe and compare the psychological profile of fibromyalgia patients attending chiropractic clinics in Australia. SETTING: Chiropractic clinics located in 5 Australian states and the Australian capital territory with practices in inner city, suburban, coastal, and rural areas were included. SUBJECTS: Chiropractic patients with acute and chronic biomechanical conditions, fibromyalgia, and who were undergoing maintenance care were included in the study. METHOD: A case study to explore the psychological profile of fibromyalgia patients was undertaken. The Distress and Risk Assessment Method (DRAM) and Sense of Coherence (SOC) questionnaires were used to ascertain and compare the distress, sense of coherence, and manageability levels of patients with fibromyalgia with patients having maintenance chiropractic care. Purposive sampling of practitioners and convenience sampling of patients fulfilling the study's inclusion criteria were undertaken. Patients were asked to complete two questionnaires and chiropractors to complete one questionnaire and an interview. RESULTS: While more than half of the patients in the fibromyalgia group were distressed, fewer than 1 in 7 maintenance patients were found to be distressed according to the DRAM questionnaire. With several individual exceptions, fibromyalgia patients also tended to have lower SOC and manageability scores than the maintenance group. CONCLUSION: This study supports the view that fibromyalgia may represent a problem of coping with various environmental stresses, including psychosocial stresses. However, owing to individual variation, a diagnosis of fibromyalgia at the clinical level does not accurately predict whether a particular patient is distressed or has a low SOC score. Screening of fibromyalgia patients may help determine whether intensive counseling and stress management by the chiropractor or another health professional should be contemplated. PMID- 10519563 TI - Spinal pain syndromes: nociceptive, neuropathic, and psychologic mechanisms. AB - BACKGROUND: Pain continues to be the main symptom reported by patients. Frequently, clinicians incorrectly diagnose patients and resulting treatments are ineffective, which may promote the development of chronic pain. This situation may arise as a result of a lack of clarity in the literature regarding pain syndromes. OBJECTIVE: To discuss the differences between nociceptive, neuropathic, and psychologic induction of pain and provide important clinical correlates to aid in diagnosis and treatment. DATA SOURCES: The data were accumulated over a period of years by reviewing contemporary articles and books and subsequently retrieving relevant papers. Articles also were selected from MEDLINE searches and from manual library searches. DATA SYNTHESIS: Nociceptive pain syndromes are responsible for the majority of pain complaints in clinical practice. Care must be taken to avoid the common mistake of the diagnosis of neuropathic pain, which can lead to inappropriate treatments. CONCLUSION: Although the treatment of neuropathic pain is difficult, sufficient evidence in the literature demonstrates that the treatment of nociceptive pain should be multimodal and involve spinal manipulation, muscle lengthening/stretching, trigger point therapy, rehabilitation exercises, electrical modalities, a variety of nutritional factors, and mental/emotional support. PMID- 10519564 TI - Chiropractic care and ochronotic arthropathy. AB - OBJECTIVE: To discuss the case of a patient with ochronotic arthropathy whose symptoms were treated with chiropractic care. An emphasis is placed on this condition's radiographic features. CLINICAL FEATURES: A 59-year-old woman with pain in her low back, right knee, and left ankle sought chiropractic evaluation. Black pigmentation was found in the sclera of both eyes, and homogentisic acid was present in the urine. Orthopedic evaluation revealed uncomplicated, nonspecific joint pain, and radiographs demonstrated characteristic spinal changes. INTERVENTION AND OUTCOME: The patient was treated with chiropractic manipulation, physiotherapy modalities, bracing, and exercises. This type of therapy was successful in reducing the symptoms and helped decrease the severity and frequency of acute exacerbations. CONCLUSION: Ochronotic arthropathY is a rare metabolic disorder that can be diagnosed from spinal radiographs. Chiropractic care is an appropriate tool for reducing its symptomatology. PMID- 10519565 TI - The empty sella. AB - OBJECTIVE: To discuss the diagnostic imaging findings of an empty sella in a chiropractic patient with emphasis on magnetic resonance imaging (MRI) of normal and abnormal pituitary appearances. CLINICAL FEATURES: A 44-year-old woman started having headache, dizziness, nausea, vomiting, and diarrhea after an argument with her boyfriend. She had been treated for acute torticollis for three weeks when the new symptoms began. Consultation with an internist led to an MRI examination of the cerebellopontine angles to exclude an acoustic neuroma. The MRI demonstrated an enlarged empty sella. There was no history of pituitary tumor or other sellar disease. INTERVENTION AND OUTCOME: There was complete remission of the symptoms after 1 additional dizzy spell that occurred 3 days after the initial symptom. No intervention was performed, but the stress levels in her life had been reduced. CONCLUSION: An enlarged empty sella can be present without symptoms and can represent an incidental finding on radiography and MRI. However, an enlarged sella seen on lateral cervical spine radiographs should prompt further evaluation to rule out pituitary disease. The normal pituitary has a varied appearance and signal intensity on MRI depending on the patient's age and pregnancy status. PMID- 10519566 TI - Research design in chiropractic. PMID- 10519567 TI - Structural rehabilitation of the spine and posture: rationale for treatment beyond the resolution of symptoms. PMID- 10519568 TI - Dysafferentation: a novel term to describe the neuropathological effects of joint complex dysfunction--a look at likely mechanisms of symptom generation. PMID- 10519569 TI - Effects of aqueous instillation. PMID- 10519570 TI - Ciprofloxacin monotherapy and bacterial keratitis. PMID- 10519571 TI - Vogt-Koyanagi-Harada syndrome after cutaneous injury. PMID- 10519572 TI - Neural retinal cell transplantation. PMID- 10519573 TI - Neural retinal cell transplantation. PMID- 10519574 TI - Small clear-zone radial keratotomy. PMID- 10519575 TI - IOP measurements after PRK. PMID- 10519576 TI - IOP measurements after PRK. PMID- 10519577 TI - The 50th anniversary of the intraocular lens and a quiet revolution. PMID- 10519578 TI - Clinical and electrophysiologic results after intracameral lidocaine 1% anesthesia: a prospective randomized study. AB - OBJECTIVE: To evaluate the efficacy and safety of intracameral lidocaine in cataract surgery compared to peribulbar anesthesia. DESIGN: A prospective, randomized, controlled study. PARTICIPANTS: A total of 200 consecutive cataract patients (200 eyes) participated. INTERVENTION: Eyes were randomly assigned to two groups: one group received 0.15 ml intracameral 1% unpreserved lidocaine combined with topical anesthesia (oxybuprocaine); the other group received 6 ml prilocaine peribulbar before phacoemulsification with sclerocorneal tunnel incision. MAIN OUTCOME MEASURES: Duration of surgery was measured; implicit time and amplitudes of the b-waves of the photopic electroretinogram (ERG) potentials (single-flash ERG and the 30-Hz flicker ERG) were recorded; frequencies of intraoperative problems, complications, intraoperative, and postoperative pain were evaluated. RESULTS: After lidocaine anesthesia combined with topical anesthesia, similar complications were found, longer operation time (P < 0.001), and significantly better visual acuity immediately after surgery (P < 0.001). The ERG amplitudes were not significantly reduced after 0.15-ml intracameral lidocaine half an hour after surgery (P > 0.05). CONCLUSION: Intracameral lidocaine 1% combined with topical anesthesia can be recommended as an alternative procedure to peribulbar anesthesia in cataract surgery with corneoscleral tunnel incision. PMID- 10519579 TI - Endophthalmitis in cataract surgery: results of a German survey. AB - OBJECTIVE: To document perioperative prophylactic treatment and to evaluate the risk factors for endophthalmitis after cataract surgery. DESIGN: Cross-sectional study via anonymous survey. PARTICIPANTS: Four hundred sixty-nine centers in Germany were queried. RESULTS: A total of 311 (67%) questionnaires were received, with each center reporting an average of 900 cataract surgeries per year (total, 340,633 surgeries in 1996). Respondents reported a total of 267 cases of endophthalmitis, which resulted in a mean responder-specific endophthalmitis rate of 0.148% versus a median rate of 0%. Statistical analysis via Poisson regression suggested that sclerocorneal incisions were associated with a reduced incidence of endophthalmitis (odds ratio, 0.35; 95% confidence interval, 0.24-0.51). Antibiotics used intraocularly (odds ratio, 0.65; 95% confidence interval, 0.43 0.98) and the preoperative application of diluted povidone-iodine on the conjunctiva (odds ratio, 0.59; 95% confidence interval, 0.36-0.99) were associated with a reduced risk of postoperative infection. Immune deficiencies (66%), diabetes mellitus (62%), occlusion of the lacrimal system (40%), and skin diseases (33%) were regarded as risk factors for endophthalmitis by the respondents. When cataract surgery is performed solely under inpatient conditions, the use of systemic antibiotics as well as the periocular injection of antibiotics at the end of the operation were associated (although not significantly) with a trend toward reducing the incidence of postoperative infection. Conversely, flushing the lacrimal drainage system, using eye shields, and cutting the eyelashes had no demonstrable effect in preventing endophthalmitis. The use of preoperative topical antibiotics (odds ratio, 2.38; 95% confidence interval, 1.21-4.68) and the performance of more than 20% of the surgeries in an outpatient center (odds ratio, 2.0; 95% confidence interval, 1.24 3.21) were associated with a detrimental effect on the development of endophthalmitis. CONCLUSIONS: Although the appropriate antibiotic agent and dosage are not yet established, the administration of intracameral antibiotics and the application of povidone-iodine on the conjunctiva significantly reduced the relative risk of postoperative endophthalmitis in this survey. Because the study was not individual based but rather on aggregate questionnaire, the results have to be interpreted with care. PMID- 10519580 TI - Objective and subjective evaluation of photic phenomena after monofocal and multifocal intraocular lens implantation. AB - OBJECTIVE: To objectively measure and compare halo, flicker, and glare disability in pseudophakic eyes with monofocal (MONO) and multifocal (MULTI) intraocular lenses (IOLs) with respect to the influence of corneal surface quality, astigmatism, and age. DESIGN: Prospective case series. PARTICIPANTS: This clinical trial involved 28 eyes of 28 patients after small-incision cataract surgery with a MONO silicone IOL and 28 eyes of 28 patients with zonal progressive silicone IOL. INTERVENTION: A computer program objectively determined halo, glare, and flicker. Corneal surface quality and astigmatism were measured using computerized videokeratography. Ray-tracing analysis was performed based on the videokeratography data to calculate retinal peak distance and distortion index. A questionnaire was sent to all patients to evaluate the incidence of subjective photic phenomena. RESULTS: Mean halo size (square degrees +/- standard deviation) valued 6.1 (+/- 1.3) in the MONO group and 7.2 (+/- 2.3) in the MULTI group with no statistically significant difference between MONO and MULTI. Flicker (in % contrast to add) was -0.7 (+/- 2.9) in the MONO group and -1.0 (+/- 4.2) in the MULTI group with no statistical differences. Glare (in % contrast to add) was 5.5 (+/- 16.5) in the MONO group and 6.5 (+/- 18.0) in the MULTI group with no statistical differences. Patients in the MONO group older than 70 years of age had significantly more glare than those younger than 70 years (P = 0.017). In the MULTI group, patients with corneal shape irregularities (peak distance > 6.0 microm) or astigmatism (> 1 diopter) had statistically significant greater halos than did patients with regular corneal shape (peak distance < or = 6.0 microm) or astigmatism (< or = 1 diopter) (P < 0.005). Three of 27 MONO patients and 9 of 28 MULTI patients noticed light sensations (mainly halos) after surgery that were not present before surgery, with the majority not being bothered by these at all. CONCLUSION: In monofocal as well as in multifocal eyes, halo and glare disability occurred. Patient age, corneal surface quality, and IOL design played an important role in these photic phenomena. Because these photic phenomena may be more prevalent in night driving conditions, the authors' study suggests that night driving ability, especially in the elderly patient with pseudophakia, should be examined carefully. PMID- 10519581 TI - Low-cost high-volume extracapsular cataract extraction with posterior chamber intraocular lens implantation in Nepal. AB - OBJECTIVE: To improve current clinical practices and ways of thinking about the problem of curable Third-World blindness resulting from cataract. DESIGN: A two site prospective, nonrandomized, comparative clinical trial. PARTICIPANTS: Patients from 2 distinct surgical venues underwent cataract surgery following the same carefully outlined protocol: 62 consecutive cases from the Tilganga Eye Centre in Katmandu, Nepal, and 207 cases from a remote eye camp in rural Chaughada, Nepal. INTERVENTION: Extracapsular cataract extraction with posterior chamber intraocular lens (IOL) implantation surgery using a technique developed by Dr. Sanduk Ruit of the Tilganga Eye Centre in conjunction with the Medical Directorate of the Fred Hollows Foundation of Australia. Also presented is the teaching method used to help make local doctors proficient in this technique. MAIN OUTCOME MEASURES: Visual acuity recorded at 2 months after surgery as well as surgical complications. RESULTS: Preoperative visual acuities for the 62 patients from the Tilganga Eye Centre ranged from 20/60 to light perception only (4 patients were untested). At 2 months after surgery, 87.1% had a best-corrected visual acuity of 20/60 or better. There were zero major surgical complications reported from the Tilganga group. Of the 207 patients at the Chaughada eye camp, preoperative visual acuities (recorded for 177 [85.5%]) ranged from 20/200 or greater to light perception only. One hundred eighty-nine (91.3%) of the patients returned for an examination at 2 months after surgery, at which time 54.5% had an uncorrected visual acuity of 20/60 or greater, improving to 74.1% with correction. There were six (2.9%) surgical complications documented at Chaughada. CONCLUSIONS: Because the average operative time using the technique presented here is less than 10 minutes per case and the cost per surgery is less than $20, the surgical results are significant in addressing the massive problem of cataract blindness in the Third World. With some changes in preoperative care, a simplified surgical technique, the development of local lens factories, and the implementation of teaching programs, effective lens implantation cataract surgery can be done in high volume at affordable costs in remote areas of underserved nations. PMID- 10519582 TI - Diabetic retinopathy in a black population: the Barbados Eye Study. AB - OBJECTIVE: The distribution of diabetic retinopathy in black populations is largely unknown. The authors present retinopathy data from the predominately black participants of the Barbados Eye Study (BES). DESIGN AND PARTICIPANTS: Prevalence study of 4631 participants based on a random sample of the Barbados population 40 to 84 years of age (84% participation). MAIN OUTCOME MEASURES: Diabetes was defined as self-reported history of physician-diagnosed diabetes or glycosylated hemoglobin greater than 10% (>2 standard deviations above the population mean of persons without a diabetes history). Retinopathy was assessed by independent gradings of 30 degrees color stereo fundus photographs of the disc and macula. RESULTS: Diabetes was present in 19.4% of black (n = 4314), 15.2% of mixed (black and white; n = 184), and 7.5% of white/other (n = 133) self-reported racial groups. In the black/mixed population, regardless of diabetes status, the prevalence of retinopathy was 5.9%. In the 636 black and mixed participants with diabetes, the prevalence of retinopathy was 28.5%: 19.8% had minimum changes, 7.7% had moderate changes, and 0.9% had severe retinopathy. Clinically significant macular edema (CSME) was found in 8.6% of those with diabetes. CONCLUSIONS: In the population of African origin, approximately 1 in 17 persons had retinopathy. Among those with diabetes, 28.5% had retinopathy and 8.6% had CSME. These results highlight the clinical and public health relevance of diabetic retinopathy in the black population. PMID- 10519583 TI - Management of submacular hemorrhage with intravitreous tissue plasminogen activator injection and pneumatic displacement. AB - OBJECTIVE: To investigate the efficacy and safety of treating thick submacular hemorrhages with intravitreous tissue plasminogen activator (tPA) and pneumatic displacement. DESIGN: Retrospective, noncomparative case series. PARTICIPANTS: From 5 participating centers, 15 eligible patients had acute (<3 weeks) thick subretinal hemorrhage involving the center of the macula in eyes with pre existing good visual acuity. Hemorrhages were secondary to age-related macular degeneration in 13 eyes and macroaneurysm and trauma in 1 eye each. METHODS: The authors reviewed the medical records of 15 consecutive patients who received intravitreous injection of commercial tPA solution (25-100 microg in 0.1-0.2 ml) and expansile gas (0.3-0.4 ml of perfluoropropane or sulfur hexafluoride) for thrombolysis and displacement of submacular hemorrhage. After surgery, patients maintained prone positioning for 1 to 5 days (typically, 24 hours). MAIN OUTCOME MEASURES: Degree of blood displacement from under the fovea, best postoperative visual acuity, final postoperative visual acuity, and surgical complications. RESULTS: In 15 (100%) of 15 eyes, the procedure resulted in complete displacement of thick submacular hemorrhage out of the foveal area. Best postprocedure visual acuity improved by 2 lines or greater in 14 (93%) of 15 eyes. After a mean follow up of 10.5 months (range, 4-19 months), final visual acuity improved by 2 lines or greater in 10 (67%) of 15 eyes and measured 20/80 or better in 6 (40%) of 15 eyes. Complications included breakthrough vitreous hemorrhage in three eyes and endophthalmitis in one eye. Four eyes developed recurrent hemorrhage 1 to 3 months after treatment, three of which were retreated with the same procedure. CONCLUSIONS: Intravitreous injection of tPA and gas followed by brief prone positioning is effective in displacing thick submacular blood and facilitating visual improvement in most patients. The rate of serious complications appears low. Final visual outcomes are limited by progression of the underlying macular disease in many patients. PMID- 10519584 TI - Transpupillary thermotherapy of occult subfoveal choroidal neovascularization in patients with age-related macular degeneration. AB - OBJECTIVE: To evaluate the efficacy of transpupillary thermotherapy for the treatment of occult subfoveal choroidal neovascularization (CNV) in patients with age-related macular degeneration. DESIGN: A retrospective, noncomparative case series. PARTICIPANTS: Sixteen eyes of 15 consecutive patients who presented with occult subfoveal choroidal neovascularization secondary to age-related macular degeneration. INTERVENTION: After informed consent was obtained, 16 eyes of 15 patients were treated with transpupillary thermotherapy. All patients underwent pretreatment fluorescein angiography and were deemed untreatable by the Macular Photocoagulation Study standard. Transpupillary thermotherapy was delivered using a diode laser at 810 nm. A variable spot size of 1.2 mm, 2.0 mm, or 3.0 mm was used depending on the size of CNV. The diode laser was delivered through a contact lens, and treatment was initiated in one spot for 60 seconds' duration at a power range between 360 and 1000 mW. The end point was an area of no visible color change to a light-gray appearance. MAIN OUTCOME MEASURES: In all eyes, outcome was assessed by Snellen chart visual acuity and clinical examination. In 10 of 16 eyes, preoperative and postoperative fluorescein angiography and optical coherence tomography were available. In the remaining 6 of 16 eyes, exudation was measured by postoperative clinical examination alone. RESULTS: Three eyes (19%) showed a two-or-more-line improvement in visual acuity over a period of 6 to 25 months. Mean follow-up was 13 months. Visual acuity remained stable (no change or one-line improvement) in nine treated eyes (56%). The remaining four eyes (25%) showed a decline (equal to one-line worsening or greater) in visual acuity. Fifteen eyes (94%) demonstrated decreased exudation on fluorescein angiography, optical coherence tomography, and/or clinical examination. CONCLUSIONS: Transpupillary thermotherapy shows no deleterious side effects in treating occult subfoveal choroidal neovascularization. A randomized, prospective study is necessary to evaluate treatment efficacy. PMID- 10519585 TI - Verteporfin photodynamic therapy retreatment of normal retina and choroid in the cynomolgus monkey. AB - OBJECTIVE: This study evaluated the effect of repeated photodynamic therapy (PDT) applications on normal primate retina and choroid using an intravenous infusion of liposomal benzoporphyrin derivative (verteporfin). DESIGN: This was an experimental study in a primate model. ANIMALS/CONTROLS: Six cynomolgus monkeys were used as experimental subjects and one monkey was used as a control subject. INTERVENTION: Three consecutive PDT treatments at 2-week intervals were applied over the center of the fovea or the optic nerve of each eye. Verteporfin was delivered by intravenous infusion at a dose of 6 mg/m2, 12 mg/m2, or 18 mg/m2. Laser irradiation was then applied using a diode laser (689 nm) with light doses and spot sizes kept constant. MAIN OUTCOME MEASURES: Findings were documented by fundus photography, fluorescein angiography, and light and electron microscopy. RESULTS: A cumulative dose response was seen angiographically and histologically with more severe damage to the retina and choroid noted at higher dye doses. Photodynamic therapy applied to the macula using the 6-mg/m2 verteporfin dose showed recovery of choriocapillaris, with mild retinal pigment epithelium and outer photoreceptor damage at 6 weeks. At this dose, the optic nerve showed few focal sites of axon atrophy and capillary loss. Treatments over the macula using the 12-mg/m2 and 18-mg/m2 doses led to chronic absence of choriocapillaris and photoreceptors at 6 weeks. One of two optic nerves became atrophic after PDT applications using dye doses of 12 mg/m2, and both optic nerves became atrophic in the 18-mg/m2 dye dose group. CONCLUSION: Limited damage to the retina, choroid, and optic nerve was present in primates treated with multiple PDT sessions using 6 mg/m2 verteporfin with light doses and the timing of irradiation kept constant. However, PDT using higher dye doses of 12 mg/m2 and 18 mg/m2 led to significant chronic damage to the normal retina, choroid, and optic nerve. PMID- 10519586 TI - High-altitude retinopathy and altitude illness. AB - OBJECTIVE: To determine the relationship between high-altitude retinopathy (HAR) and other altitude-related illnesses and establish a classification system for HAR. DESIGN: Observational case series. PARTICIPANTS: All 40 climbers among 3 Himalayan expeditions who ascended to altitudes between 16,000 and 29,028 feet above sea level (summit of Mt. Everest) were examined for signs of HAR and altitude illness (AI). METHODS: All subjects had dilated fundus examinations before the ascent, intermittent fundus, and medical examinations during the climb and a dilated fundus and medical examination within 2 days after attaining their highest altitude. MAIN OUTCOME MEASURES: Careful fundus drawings or fundus photography or both were obtained for all participants. All subjects gave a subjective assessment of their symptoms of acute mountain sickness (AMS) and were assessed clinically for signs of high-altitude cerebral edema (HACE). RESULTS: Nineteen of 21 climbers who ascended above 25,000 feet developed HAR. Fourteen of 19 climbers who attained altitudes between 16,000 and 25,000 feet were found to have retinopathy. A grading system for HAR describing the severity of the retinopathy was developed. Correlation of the retinopathy with other AI showed that AMS was endemic and that a statistically significant correlation exists between HAR and HACE (P = 0.0240). CONCLUSION: Recognizing advancing grades of HAR may allow physicians to recommend initiating empiric treatment with oxygen, steroids, diuretics and immediate descent to prevent HAR progression, macular involvement, or potentially fatal HACE. High-altitude retinopathy is both a significant component of and a predictor of progressive AI. PMID- 10519587 TI - Indocyanine green angiography in birdshot chorioretinopathy. AB - OBJECTIVE: Birdshot chorioretinopathy (BC) is an ocular inflammatory disease involving both the retina and the choroid. The study goal was to evaluate indocyanine green angiographic features in BC to assess choroidal involvement. DESIGN: Retrospective, observational case series. PARTICIPANTS: Fifty-two patients with BC documented with at least 1 concomitant fluorescein and indocyanine green angiogram. INTERVENTION: Indocyanine green angiography (ICGA) was performed according to a standard protocol used for inflammatory disorders. MAIN OUTCOME MEASURE: Indocyanine green angiographic signs were correlated with fundus photographs, fluorescein angiography, degree of inflammatory activity, and stage of disease. RESULTS: In active disease, three main features were observed. The principal finding, found in 100% of patients, was the presence of hypofluorescent dark dots during the intermediate phase of angiography; their evolutionary pattern was twofold, becoming either isofluorescent or remaining hypofluorescent at the late phase of angiography. The other two signs were fuzzy, indistinct choroidal vessels and late-diffuse choroidal hyperfluorescence. In chronic longlasting disease, the characteristic finding was the presence of hypofluorescent dark dots that persisted in the late phase of disease and is theorized to correspond either to chorioretinal atrophy (irregular geographic pattern) or to persistent choroidal granulomas (round oval form). CONCLUSIONS: Consistent ICGA findings in 52 patients allowed the authors to establish a fairly precise ICGA semiology for BC. This procedure enabled the authors to assess choroidal involvement, and, in selected cases, it also was found to be of diagnostic help and useful to monitor therapeutic intervention. PMID- 10519588 TI - Computer-vision-enabled augmented reality fundus biomicroscopy. AB - PURPOSE: To guide treatment for macular diseases and to facilitate real-time image measurement and comparison, investigations were initiated to permit overlay of previously stored photographic and angiographic images directly onto the real time slit-lamp biomicroscopic fundus image. DESIGN: Experimental study in model eyes, and preliminary observations in human subjects. METHODS: A modified, binocular video slit lamp interfaced to a personal computer and framegrabber allows for image acquisition and rendering of stored images overlaid onto the real-time slit-lamp biomicroscopic fundus image. Development proceeds with rendering on a computer monitor, while construction is completed on a miniature display interfaced directly with one of the slit-lamp oculars. Registration and tracking are performed with in-house-developed software. MAIN OUTCOME MEASURES: Tracking speed and accuracy, ergonomic acceptability. RESULTS: Computer-vision algorithms permit robust montaging, tracking, registration, and rendering of previously stored photographic and angiographic images onto the real-time slit lamp fundus biomicroscopic image. In model eyes and in preliminary studies in a human eye, optimized registration permits near-video-rate image overlay with updates at 3 to 10 Hz and misregistration errors on the order of 1 to 5 pixels. CONCLUSIONS: A prototype for ophthalmic augmented reality (image overlay) is presented. The current hardware/software implementation allows for robust performance. PMID- 10519589 TI - Rhegmatogenous retinal detachment after block excision of epithelial implantation cysts and tumors of the anterior uvea. AB - OBJECTIVE: To report on frequency, time of occurrence, treatment, and final outcome of rhegmatogenous retinal detachment after block excision, combined with corneoscleral tectonic grafts, of intraocular epithelial implantation cysts and tumors of the anterior uvea involving the anterior chamber angle. DESIGN: Noncomparative case series. PARTICIPANTS: The study included 144 patients who had consecutively undergone scleral full-thickness block excision of tumors of the anterior uvea (n = 87) or intraocular epithelial implantation cysts (n = 57). Diameter of the block excision ranged between 5.5 and 20 mm. In 39 patients, the tumor extended posterior to the ora serrata. INTERVENTIONS: Retinal detachment surgery. MAIN OUTCOME MEASURES: Retinal detachment rate, visual acuity, risk factors. RESULTS: Retinal detachment occurred in 10 (6.9%) of 144 patients (2 of 57, or 3.5% of patients with cysts; 8 of 87 or 9.2% of patients with tumors) 3 to 12 months after block excision. Six patients underwent primary pars plana vitrectomy with temporary endotamponade by silicone oil, which was removed 3 to 9 months later. In four patients, a scleral buckling procedure only was performed. After a mean follow-up of 31 months after silicone oil removal or the buckling procedure (median, 30 months; range, 13-42 months), the retina has remained attached in all patients operated on. Visual acuity increased from 2/20 (median; range, light perception-6/20) to 8/20 (median; range, 2/20-12/20). In the eyes with retinal detachment compared to the eyes without retinal detachment, the block excision was significantly larger (13.75+/-4.86 mm vs. 9.41+/-3.02; P = 0.01) and was located significantly more posteriorly (limbus distance of posterior excision margin: 6.75+/-2.87 vs. 4.35+/-3.24 mm; P = 0.01). CONCLUSIONS: Scleral buckling procedures and primary pars plana vitrectomy with temporary ocular endotamponade can give acceptable results in eyes with rhegmatogenous retinal detachment occurring after block excision of epithelial implantation cysts or tumors of the anterior uvea. Despite intraoperative vitreous prolapse or tumor extension posterior to the ciliary body, rhegmatogenous retinal detachment occurs in fewer than 10% of patients undergoing block excision of cysts or tumors of the anterior uvea. Size and posterior location of the block excision are the main risk factors for rhegmatogenous retinal detachment, which becomes unlikely later than 12 months after surgery. PMID- 10519590 TI - A phase I/II study of subconjunctival carboplatin for intraocular retinoblastoma. AB - PURPOSE: To evaluate the efficacy and toxicity of subconjunctival carboplatin for intraocular retinoblastoma. DESIGN: A phase I/II clinical trial (noncomparative case series). PARTICIPANTS: Eleven children with bilateral retinoblastoma who had 13 eyes containing active tumor. METHODS: Subconjunctival carboplatin (1.4-2.0 ml of a 10-mg/ml solution) was administered. Ophthalmologic examinations with the patient under anesthesia were performed before each injection, and response and toxicity were assessed. MAIN OUTCOME MEASURES: Tumor response and toxicities. RESULTS: A median of three injections per eye was administered (range, 1-7 injections per eye) at a median interval of 21 days between injections (range, 14 35 days). The median dose was 20 mg/injection (range, 14-20 mg). Toxicities included transient periorbital edema in four eyes and optic atrophy in one eye that also received laser photocoagulation and cryotherapy. No nonocular toxicities were noted. Three of five eyes with vitreous disease that could be evaluated had major responses. Two of five eyes with retinal tumors had a response, but neither eye with subretinal disease responded. CONCLUSIONS: Subconjunctival carboplatin as used in this protocol is effective for intraocular retinoblastoma. It appears to be safe, but additional study and longer follow-up are required, particularly to determine the risk of optic atrophy. PMID- 10519591 TI - Prevalence of systemic and ocular disease in infantile exotropia: comparison with infantile esotropia. AB - OBJECTIVE: Exotropia in infancy is believed to be associated with an increased prevalence of neurologic, ocular, and craniofacial abnormalities; however, the prevalence of coexisting ocular and systemic disease in these patients is unknown. In this study, the prevalence of ocular disease and systemic illness was determined in patients diagnosed with exotropia in infancy. DESIGN: Observational comparative case series. PARTICIPANTS: Medical records of 70 patients diagnosed with exotropia in the first year of life were reviewed and compared with records of 136 patients diagnosed with esotropia before 1 year of age. INTERVENTION: Patients with no disorders (other than latent nystagmus, dissociated vertical deviation, or oblique muscle overaction) were grouped as "simple" strabismus. Patients with systemic disorders (including prematurity, neurologic disease, and genetic disease) and patients with ocular disorders (including congenital nystagmus, other strabismus, ptosis, and any condition associated with loss of vision [except amblyopia]) were grouped as "complex" strabismus. MAIN OUTCOME MEASURES: Prevalence of coexisting systemic and ocular disorders. The demographics, strabismus measurements, and types of coexisting disease in the simple and complex groups were compared. RESULTS: A high percentage of both exotropia (67%) and esotropia (49%) patients had a coexisting ocular or systemic abnormality. Exotropia patients with a constant strabismus were more likely to have coexisting ocular or systemic disease than those with an intermittent strabismus. Smaller angles of exotropia or esotropia were associated with a higher likelihood of coexisting ocular or systemic diseases. Systemic disorders were found more frequently than ocular disorders in both the exotropia and esotropia groups. In 25% of all patients referred for evaluation of strabismus, an additional ocular or systemic abnormality was discovered by the ophthalmologist. CONCLUSION: Patients presenting to a university hospital-based practice in the first year of life with exotropia were more likely than those presenting with esotropia to have coexisting ocular and systemic disease. Both groups had a notably high prevalence of associated disorders. The percentages measured in this population may not be applicable to other practices because of referral bias. However, clinicians should consider that children presenting with infantile exotropia and esotropia appear to be at risk for coexisting ocular or systemic disease. PMID- 10519592 TI - Isolated sulfite oxidase deficiency: review of two cases in one family. AB - OBJECTIVE: The authors describe two cases of isolated sulfite oxidase deficiency found in one family. This is a rare autosomal-recessive disorder presenting at birth with seizures, severe neurologic disease, and ectopia lentis. It can be easily missed with metabolic screening; however, the finding of lens subluxation stresses the importance of ophthalmic assessment in making the diagnosis. DESIGN: Two observational case reports. INTERVENTION/METHODS: Ophthalmic assessment, biochemical assay for specific urinary and plasma metabolites, magnetic resonance imaging, and gene sequencing were used to make the diagnosis of the disease in the proband. The diagnosis was subsequently recognized in a previously affected sibling after the postmortem neuropathology was reviewed. Mutation analysis was performed on cultured fibroblasts from the proband to identify and categorize the specific mutation responsible for the disease in the family. From this, future prenatal detection of sulfite oxidase deficiency is possible. MAIN OUTCOME MEASURES: The diagnosis of sulfite oxidase deficiency was established in this family, enabling appropriate genetic counseling and recurrence risk estimation. RESULTS: Point mutations were found in both alleles of the sulfite oxidase gene in the proband. The first is a 623C-->A mutation, which predicts an A208D substitution, and the second is a 1109C-->A, which predicts an S370Y substitution. Both residues A208D and S370Y are critical for sulfite oxidase activity. CONCLUSIONS: Isolated sulfite oxidase deficiency is a rare heritable disease for which mutation analysis can allow accurate prenatal screening. It often is difficult to diagnose by clinical presentation alone, but the critical finding of lens subluxation accompanying seizures and diffuse neurologic disease in an infant should alert the physician to the diagnosis. PMID- 10519593 TI - Initial endothelial cell density and chronic endothelial cell loss rate in corneal transplants with late endothelial failure. AB - OBJECTIVE: To determine when corneal transplants develop the decreased endothelial cell density that predisposes to late endothelial failure (LEF). DESIGN: Noted cohort study within a prospective case series. PARTICIPANTS: The authors compared 21 grafts that developed LEF with the remaining transplants (controls) in a longitudinal cohort study of 500 consecutive penetrating keratoplasties by 1 surgeon. Eyes regrafted during the study, fellow eyes of bilateral cases, and patients who did not want their data used for research purposes were excluded, leaving 389 grafts in 389 patients for analysis. INTERVENTION: Penetrating keratoplasty. MAIN OUTCOME MEASURES: Endothelial cell density before surgery and at 2 months and 1, 3, 5, and 10 years after surgery. RESULTS: Grafts with LEF had lower median endothelial cell densities than other grafts, both before surgery (2710 cells/mm2 vs. 2991 cells/mm2; P = 0.02) and 2 months after surgery (1895 cells/mm2 vs. 2501 cells/mm2; P < 0.001), a difference that was maintained through 5 postoperative years. Despite having lower preoperative cell densities, the grafts with LEF had larger median endothelial cell losses 2 months after keratoplasty (31% vs. 16%, P = 0.002). The endothelial cell loss subsequent to the 2-month examination, however, was not increased in the grafts with LEF. Risk factors for developing LEF included a low endothelial cell density before surgery (P = 0.007) and 2 months after surgery (P = 0.002), aphakia or pseudophakia (P = 0.04), older recipient age (P = 0.002) and older donor age (P = 0.03), and occurrence of an endothelial rejection episode (P = 0.03). CONCLUSIONS: Corneal grafts with LEF, the major cause of graft failure after 5 postoperative years, fail from low initial endothelial cell density rather than an increased rate of chronic postoperative cell loss. Improved techniques of corneal preservation should decrease the rate of LEF. In addition to low preoperative and 2-month postoperative endothelial cell density, a higher risk of LEF is also seen in patients with aphakia or pseudophakia, older recipient age, older donor age, and occurrence of an endothelial rejection episode. PMID- 10519594 TI - Laser in situ keratomileusis for correction of myopia and astigmatism after penetrating keratoplasty. AB - PURPOSE: To determine the safety and effectiveness of laser in situ keratomileusis (LASIK) for visual rehabilitation of residual myopia and astigmatism after penetrating keratoplasty. DESIGN: Prospective, noncomparative case series. PARTICIPANTS: LASIK was performed on 23 eyes of 22 patients unable to wear glasses or contact lenses after penetrating keratoplasty due to anisometropia, high astigmatism, and/or contact lens-intolerance. METHODS: All patients underwent LASIK for visual rehabilitation after penetrating keratoplasty. Uncorrected visual acuity and best spectacle-corrected visual acuity, degree of anisometropia, and corneal transplant integrity were recorded before surgery, as well as at 1 month, 3 months, 6 months, and 12 months after LASIK surgery. RESULTS: The mean spherical equivalent before surgery was -7.58+/ 4.42 diopters (D), which was reduced to -1.09+/-2.01 D, -0.79+/-1.84 D, -0.77+/ 1.25 D, and -1.57+/-1.20 D, respectively, at 1, 3, 6, and 12 months after LASIK. The mean cylinder before surgery was 3.64+/-1.72 D, which was reduced to 1.98+/ 1.15 D, 1.64+/-1.14 D, 1.48+/-0.92 D, and 1.29+/-1.04 D, respectively, at 1, 3, 6, and 12 months after LASIK. Spherical equivalent anisometropia was reduced from a mean of 6.88+/-4.4 D to 1.42+/-1.05 D at the final examination. Best-corrected visual acuity remained the same or improved in 21 of 23 eyes and decreased by 1 and 3 lines in 2 patients. There were no surgical flap or corneal transplant complications. CONCLUSIONS: LASIK is a viable treatment alternative for myopia and astigmatism after penetrating keratoplasty in patients who are contact lens intolerant. LASIK is more effective in treating myopia than astigmatism after penetrating keratoplasty. PMID- 10519595 TI - Photorefractive keratectomy for hyperopia: long-term nonlinear and vector analysis of refractive outcome. AB - PURPOSE: To characterize the refractive changes after excimer laser photorefractive keratectomy for the correction of hyperopia over a follow-up up to 3 years and to assess refractive stability and changes in astigmatism. DESIGN: Noncomparative, nonrandomized, retrospective, interventional case series. PARTICIPANTS: Thirty-eight hyperopic eyes of 28 patients (age range, 33-62 years) with refraction in the range +1.00 to +8.00 diopters (D). Mean attempted correction was +3.33+/-0.98 D (range, +1.00 to +4.00 D). Data were compared to those from 216 eyes treated for myopia in the range -1.00 to -12.70 D. INTERVENTION: The hyperopic correction was made using an erodible mask inserted in the laser optical pathway to produce a circular ablation measuring 6.5 mm in diameter. An axicon was then used to create a blend transition zone from 6.5 mm up to 9.4 mm in diameter. Eyes were evaluated 3 to 11 times (5.5+/-2.4) over a 3- to 34-month follow-up (16.8+/-8.4 months). MAIN OUTCOME MEASURES: Vector analysis of refractive error, applying a nonlinear statistical model fitting the spherical equivalent, and the sphere component data. The fit parameters were the long-term error at stabilization (epsilon(infinity)), the amount of regression (epsilon0), being the difference of refractive errors immediately after surgery and at stabilization, and the time constant (T1/2) giving the temporal scale length by which the overcorrection halves (regression half-life). Cylinder was analyzed by a linear regression. RESULTS: The initial overcorrection was much larger after hyperopic treatments than myopic ones (epsilon0 = -3.26+/-0.35 D vs. +1.43+/-0.35 D), and it takes typically four times longer to regress (T1/2 = 3.30+/-0.91 months). Refractive stabilization is reached after more than 1 year, with a satisfactory refractive result. The hyperopic treatment induces a mean astigmatism of 1.00 D, which remains constant throughout the follow-up, and tends to be aligned along the with-the-rule meridian. CONCLUSIONS: The advantages of a reasonably well-designed algorithm to correct hyperopia (epsilon(infinity) = +0.20+/-0.23 D) are counterbalanced by the long time to refractive stabilization and by the induced astigmatism. PMID- 10519596 TI - Treatment of persistent corneal epithelial defect by autologous serum application. AB - OBJECTIVE: To evaluate the efficacy of autologous serum application for the treatment of persistent epithelial defect. DESIGN: Prospective, clinical, noncomparative case series. PARTICIPANTS: A total of 16 eyes were studied. INTERVENTION: Autologous serum was prepared from the patients and diluted to 20% by saline. The patients were instructed to use the autologous serum six to ten times a day. The concentration of vitamin A, epidermal growth factor (EGF), and transforming growth factor-beta (TGF-beta) was measured at 1 week and 1 month stored in the refrigerator and 1 month and 3 months in the freezer. MAIN OUTCOME MEASURES: Time to closure of epithelial defect. RESULTS: Vitamin A, EGF, and TGF beta were stable during the 1 month in the refrigerator and 3 months in the freezer. Among 16 persistent epithelial defects, 7 (43.8%) healed within 2 weeks, 3 (18.8%) healed within 1 month, and the remaining 6 (37.5%) did not respond within 1 month. No apparent side effect of autologous serum application was observed. CONCLUSIONS: Autologous serum application healed 43.8% of persistent defect within 2 weeks and 62.5% within 1 month. PMID- 10519597 TI - Presumed periorbital lupus vulgaris with ocular extension. AB - OBJECTIVE: To report an unusual case of lupus vulgaris that spread to the left anterior ocular surface. DESIGN: Case report. PARTICIPANT: An 18-year-old woman presented with an 8-month history of an infiltrative skin lesion affecting the left lower eyelid and cheek area, left globe, right lacrimal sac area, together with a cystic lesion in the foot area. TESTING/INTERVENTION: The authors describe the clinical findings, radiologic study, and histopathologic study of the conjunctiva, skin, liver, and ankle lesion. The patient was treated with antituberculous medications for 3 months. MAIN OUTCOME MEASURES: Healing of the skin, conjunctival, and bone lesions. RESULTS: The lesion of the face healed, leaving scar tissue. The left eye showed symblepharon with loss of its anterior surface features. The right eye showed no symblepharon, the bones of the foot healed with no deformity, and the liver function test results normalized after 3 months of antituberculous medications. CONCLUSION: Lupus vulgaris can be associated with multiple system involvement. Its clinical presentation and behavior depend on the patient's immunity and duration of the disease. Early diagnosis and appropriate management may cure the disease with no life threatening sequelae. PMID- 10519598 TI - Symptomatic compression of the optic nerve by the carotid artery: clinical profile of 18 patients with 24 affected eyes identified by magnetic resonance imaging. AB - OBJECTIVE: To characterize the clinical features and course of patients with magnetic resonance imaging (MRI)-defined optic nerve compression by the supraclinoid carotid artery. DESIGN: Retrospective, observational case series. PARTICIPANTS: Eighteen patients with 24 affected eyes were identified by reviewing case records from the author's referral-based neuro-ophthalmology practice. Predetermined inclusion and exclusion criteria were applied to potential participants. MAIN OUTCOME MEASURES: The following variables were abstracted from the medical record: age, gender, presenting symptoms, past medical problems, visual acuity, color vision, visual field, pupillary reactions, optic disc appearance, other neurologic signs, and previously documented and follow-up examinations. RESULTS: There were eight women and ten men ranging in age from 28 to 86 years (median age, 72 years) at the time of diagnosis. Ten (56%) of 18 patients had hypertension. Twelve patients had unilateral optic neuropathy, whereas 6 patients had bilateral optic neuropathy. One patient presented with subacute superior orbital fissure syndrome due to mass effect of a dolichoectatic carotid artery. Another patient had oculomotor nerve palsy with signs of aberrant regeneration due to intracavernous mass effect of a dolichoectatic carotid artery. One patient had a bitemporal hemianopia associated with bilateral compression of the immediate prechiasmatic optic nerves by dolichoectatic carotid arteries. The predominant pattern of visual field loss in most patients reflected nerve fiber bundle injury. A central scotoma or absolute central visual field loss was noted in only 6 (25%) of 24 affected eyes. Most patients demonstrated saucerlike excavation of the optic disc. Progression of visual acuity loss occurred at a relatively slow rate. CONCLUSIONS: Although uncommon, intracranial compression of the optic nerve by the carotid artery should be considered in a patient with unexplained or progressive unilateral or bilateral optic neuropathy. This entity can be diagnosed using clinical skills to exclude more common causes of optic nerve injury and coronal-oriented MRI to confirm anatomic compression of the symptomatic optic nerve. Although many affected patients have excavation of the optic disc and nerve fiber bundle visual field defects, most have additional signs atypical for glaucoma, minimizing the potential for diagnostic confusion between the two disorders. PMID- 10519599 TI - Expansion of the human microphthalmic orbit. AB - OBJECTIVE: To determine the effects of long-term, incremental enlargement of an orbital tissue expander on bone and eyelid growth in microphthalmia. DESIGN: A prospective, noncomparative case series. PARTICIPANTS: Five consecutive patients with microphthalmos treated with orbital expansion were evaluated. INTERVENTION: A tissue expander was placed into the orbits of five children (age, 10 months-6 years) with unilateral microphthalmos and gradually enlarged by saline injections. MAIN OUTCOME MEASURE: The midorbital width of each patient was determined from axial computed tomographic scans before insertion of the device. The length of the normal and abnormal eyelid fissures was measured at surgery. The postexpansion dimensions of both the normal and microphthalmic orbits and the eyelids were remeasured when the expanders were removed. The residual deficits between the normal and the microphthalmic sides were expressed in percentages. RESULTS: Gradual inflation of the expander to a diameter of 22 mm reduced the average preoperative orbital dimension deficit of the group from 14.6% (range, 8% 25%) to 3.8% after surgery (range, 0.5%-6.3%). The average pre-expansion eyelid length deficit for the group was 17.5% (range, 12%-26%) compared to 2.3% (range, 0.0%-5.3%) after expansion. The average expansion period was 56.8 weeks (range, 20-100 weeks). Two outpatient surgical procedures were required in each patient. CONCLUSION: Incremental inflation of a tissue expander placed within the microphthalmic orbit induced sufficient osseous and eyelid growth to ameliorate the major stigmata of this syndrome in all patients treated. PMID- 10519600 TI - The relationship between glaucoma and myopia: the Blue Mountains Eye Study. AB - OBJECTIVE: To quantify the relationship between myopia and open-angle glaucoma, ocular hypertension (OH), and intraocular pressure (IOP) in a representative older population. DESIGN: Cross-sectional population-based study of 3654 Australians 49 to 97 years of age. METHODS: Subjects with any myopia (> or =-1.0 diopter [D]) were identified by a standardized subjective refraction and categorized into low myopia (> or =-1.0 D to <-3.0 D) or moderate-to-high myopia (> or =-3.0 D). Glaucoma was diagnosed from characteristic visual field loss, combined with optic disc cupping and rim thinning, without reference to IOP. Ocular hypertension was diagnosed when applanation IOP was greater than 21 mmHg in either eye in the absence of glaucomatous visual field and optic disc changes. MAIN OUTCOME MEASURE: General estimating equation models were used to assess associations between eyes with myopia and either glaucoma or OH. RESULTS: Glaucoma was present in 4.2% of eyes with low myopia and 4.4% of eyes with moderate-to-high myopia compared to 1.5% of eyes without myopia. The relationship between glaucoma and myopia was maintained after adjusting for known glaucoma risk factors, odds ratio (OR) of 2.3, and 95% confidence intervals (CI) of 1.3 to 4.1 for low myopia. It was stronger for eyes with moderate-to-high myopia (OR, 3.3; CI, 1.7-6.4). Only a borderline relationship was found with OH, OR of 1.8 (CI, 1.2-2.9) for low myopia, and OR of 0.9 (CI, 0.4-2.0) for moderate-to-high myopia. Mean IOP was approximately 0.5 mmHg higher in myopic eyes compared to nonmyopic eyes. CONCLUSIONS: This study has confirmed a strong relationship between myopia and glaucoma. Myopic subjects had a twofold to threefold increased risk of glaucoma compared with that of nonmyopic subjects. The risk was independent of other glaucoma risk factors and IOP. PMID- 10519601 TI - Age-related changes in intraocular pressure in a large Japanese population: a cross-sectional and longitudinal study. AB - OBJECTIVE: To assess the influence of aging, blood pressure, and body mass index (BMI) on intraocular pressure (IOP) in a large Japanese population. DESIGN: Cross sectional and longitudinal study. PARTICIPANTS: The participants in this study were 69,643 Japanese men and women 20 to 79 years of age. They were office workers and their family members who had received annual health examinations from 1989 to 1997. The records of the participants who received health examinations were reviewed retrospectively. Each participant was examined according to a standard protocol, including tonometry with a noncontact tonometer, anthropometric measurements, and blood pressure measurements. The data from the subjects' most recent visit were analyzed cross-sectionally. The data from the 68,998 men and women of the total participants who were born between the 1920s and 1960s were used in longitudinal analysis. TESTING: Tonometric and anthropometric measurements. MAIN OUTCOME MEASURES: Mean values of IOP, systolic blood pressure, diastolic blood pressure, and BMI were determined. The relationship among IOP, age, systolic blood pressure, diastolic blood pressure, and BMI was studied using the multiple linear regression model with cross sectional analysis. In longitudinal analysis, regression coefficients of IOP against age, systolic blood pressure, diastolic blood pressure, and BMI were calculated using the mixed effect model. RESULTS: The mean (+/-standard deviation) IOP values for men and women were 11.9+/-2.5 and 11.5+/-2.4 mmHg, respectively. In cross-sectional analysis, IOP decreased significantly with age (P < 0.001). However, longitudinal analysis showed that IOP increased significantly with age in both men and women (P < 0.001). Systolic blood pressure and BMI were positively correlated to IOP in both the cross-sectional and longitudinal studies. CONCLUSION: The authors found an inconsistency in the change in IOP against age between cross-sectional and longitudinal analysis. It is suspected that birth cohort differences in ocular characteristics influence IOP in the Japanese population. PMID- 10519602 TI - Neodymium: YAG transconjunctival laser revision of late-failing filtering blebs. AB - OBJECTIVE: To study the effectiveness of neodymium:YAG (Nd:YAG) laser transconjunctival revision of late-failing filtering blebs. DESIGN: Retrospective, noncomparative case series. PARTICIPANTS: Thirty consecutive patients from a glaucoma referral practice at the Ohio State University Department of Ophthalmology. INTERVENTION: Transconjunctival Nd:YAG laser revision of blebs diagnosed as failing secondary to episcleral fibrosis. MAIN OUTCOME MEASURE: Reduction in intraocular pressure (IOP) and survival of bleb. RESULTS: Reduction in IOP was attained in 24 (80%) of 30 patients with a mean prelaser IOP of 21.21 mmHg and a mean 1-week postlaser IOP of 13.97 mmHg (P < 0.0001). These results were maintained at the 1-, 3-, and 12-month follow-up visits with mean IOPs of 16.31 mmHg (P = 0.0008), 14.81 mmHg (P < 0.0001), and 15.25 mmHg (P = 0.0003), respectively. Two-year data were available on a small number of patients (4) with a mean of 15.25 mmHg (P = 0.0085). Mean time interval between trabeculectomy and laser revision was 31 months. CONCLUSION: Late-failing filtering blebs are often amenable to Nd:YAG transconjunctival revision. PMID- 10519603 TI - Heidelberg retina tomography and optical coherence tomography in normal, ocular hypertensive, and glaucomatous eyes. AB - PURPOSE: To evaluate optic disc and retinal nerve fiber layer (RNFL) appearance in normal, ocular-hypertensive, and glaucomatous eyes undergoing confocal scanning laser ophthalmoscopy and optical coherence tomography (OCT). DESIGN: Prospective, cross-sectional study. PARTICIPANTS: Seventy-eight eyes of 78 consecutive normal (n = 17), ocular-hypertensive (n = 23), and glaucomatous subjects (n = 38) were enrolled. METHODS: Each patient underwent complete ophthalmic examination, achromatic automated perimetry, confocal scanning laser ophthalmoscopy (Heidelberg Retinal Tomography [HRT]), and OCT. Topographic HRT parameters (disc area, cup-disc ratio, rim area, rim volume, cup shape measure, mean RNFL thickness, and cross-sectional area) and mean OCT-generated RNFL thickness were evaluated in each group. MAIN OUTCOME MEASURES: OCT and HRT assessment of optic disc and RNFL anatomy. RESULTS: OCT RNFL thickness showed no difference between normal and ocular-hypertensive eyes (P = 0.15) but was significantly less in glaucomatous eyes (P < 0.001). HRT measurements of rim area, cup-disc ratio, cup shape measure, RNFL thickness, and RNFL cross-sectional area were significantly less in glaucomatous eyes (all P < 0.005) and were correlated with mean OCT RNFL thickness (all P < 0.02). RNFL thickness using OCT or HRT was highly correlated with visual field mean defect during achromatic perimetry (P < 0.0001). CONCLUSION: Both HRT and OCT can differentiate glaucomatous from nonglaucomatous eyes. RNFL thickness measurements using OCT correspond to disc topographic parameters using HRT. PMID- 10519604 TI - Control of intraocular pressure elevations after argon laser trabeculoplasty: comparison of brimonidine 0.2% to apraclonidine 1.0%. AB - OBJECTIVE: To determine whether brimonidine 0.2% can control intraocular pressure (IOP) spikes as well as apraclonidine 1.0% can in those patients undergoing argon laser trabeculoplasty (ALT). DESIGN: Prospective, randomized, double-masked, clinical trial. PARTICIPANTS: A total of 56 eyes of 41 patients with open-angle glaucoma or ocular hypertension were entered in the study; 46 eyes of 41 patients were eventually used for the final analysis. INTERVENTION: Patients were randomized to receive either brimonidine 0.2% or apraclonidine 1.0% before and after 360 degrees ALT. Both patient and physician were masked as to which agent each patient received. MAIN OUTCOME MEASURES: Intraocular pressure measurements were recorded before surgery and at 1, 2, and 4 hours after surgery. The difference between the preoperative IOP (baseline) and the highest recorded postoperative IOP was recorded as the maximum IOP change. The mean of the maximum IOP change for each group was analyzed using a two-sample, one-tailed t test. RESULTS: The mean of the maximum IOP change in the brimonidine 0.2% group was 2.6+/-3.6 mmHg, and the mean for the apraclonidine 1.0% group was -2.3+/-3.7 mmHg (P = 0.8). No patient had a pressure spike greater than 10 mmHg. CONCLUSIONS: Brimonidine 0.2% appears to be as effective as apraclonidine 1.0% in preventing IOP spikes after argon laser trabeculoplasty. PMID- 10519605 TI - Micronized progesterone: clinical indications and comparison with current treatments. AB - OBJECTIVE: To integrate and evaluate the pharmacokinetic, endocrine, and clinical effects of micronized progesterone formulations. DESIGN: Published articles concerning the pharmacokinetics of orally administered progesterone and the potential clinical uses of oral micronized progesterone were reviewed. Results concerning their use for secondary amenorrhea, premenopausal bleeding disorders, luteal phase dysfunction, termination of premature labor, hormone replacement therapy, and premenopausal syndrome are summarized. Critical issues to be resolved through ongoing preclinical and clinical research are highlighted. RESULT(S): Because of the enhanced bioavailability of oral micronized progesterone, the compound may be useful for a variety of therapeutic indications. Oral micronized progesterone is available in France, and a formulation recently has been approved in the United States for the treatment of secondary amenorrhea and postmenopausal hormone replacement therapy. A large body of evidence, including the Postmenopausal Estrogen/Progestin Interventions study, suggests that the use of a combination of estrogen and oral micronized progesterone is optimal for long-term hormone replacement therapy. There also are data indicating that oral micronized progesterone could be of potential use for the treatment of premenopausal bleeding disorders, luteal phase disorders, and premature labor. CONCLUSION(S): Oral micronized progesterone has widespread clinical potential, particularly for the treatment of secondary amenorrhea and dysfunctional premenopausal bleeding, and as a component of postmenopausal hormone replacement therapy. PMID- 10519606 TI - Statistically valid infertility research. PMID- 10519607 TI - High estradiol levels and high oocyte yield are not detrimental to in vitro fertilization outcome. AB - OBJECTIVE: To evaluate the impact of elevated peak E2 levels and a high number of retrieved oocytes on implantation in patients undergoing assisted reproductive techniques. DESIGN: Retrospective study. SETTING: University-based IVF program. PATIENT(S): One hundred six patients undergoing 106 IVF cycles. High responders were defined as those who had peak E2 levels of >3,000 pg/mL on the day of hCG administration (n = 38) or >15 retrieved oocytes (n = 48). Their IVF outcomes were compared with those of patients whose peak E2 levels were < or =3,000 pg/mL (n = 68) or who had < or =15 retrieved oocytes (n = 58). INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Implantation and pregnancy rates. RESULT(S): There were no statistically significant differences in age, basal FSH level, basal E2 level, number of ampules of gonadotropins required, fertilization rate, number of ETs, implantation rate, or pregnancy rate between normal and high responders or between women who did and did not become pregnant. In addition, no differences were detected when outcome was analyzed according to the stimulation regimen used. CONCLUSION(S): Elevated peak E2 levels and high oocyte yield are not detrimental to IVF outcome. More studies are needed to characterize the threshold E2 levels above which implantation rates are reduced. PMID- 10519608 TI - Cessation of gonadotropin-releasing hormone analogue (GnRH-a) upon down regulation versus conventional long GnRH-a protocol in poor responders undergoing in vitro fertilization. AB - OBJECTIVE: To determine whether a controlled ovarian hyperstimulation (COH) regimen that involves GnRH agonist (GnRH-a) discontinuation before administration of gonadotropins would benefit poor responders. DESIGN: A prospective, randomized controlled trial. SETTING: Hospital-based IVF Unit. PATIENT(S): Sixty-three patients with previous poor response to COH and/or high basal FSH level (> or =9 mIU/mL) undergoing 78 IVF-ET cycles. INTERVENTION(S): In both groups, administration of GnRH-a was started in the midluteal phase. Whereas in the study group (40 cycles), it ended before administration of gonadotropins, in controls (38 cycles) GnRH-a treatment was continued throughout the follicular phase. MAIN OUTCOME MEASURE(S): Ovarian stimulation patterns and IVF outcome. RESULT(S): A significantly higher cancellation rate was noted in the study group than in the controls (22.5% versus 5%, respectively). The new and control regimens resulted in similar stimulation characteristics and clinical pregnancy rates (11% versus 10.3%, respectively). In 13 patients with a basal FSH level that was not persistently high, the new regimen resulted in a significantly higher number of retrieved oocytes compared with the standard protocol (7.6+/-1.03 versus 4.0+/ 0.68, respectively). CONCLUSION(S): Whereas for most low responders, the new COH regimen offers no further advantage, future prospective studies may demonstrate whether it can confer a benefit for a subset of patients with a basal FSH level that is not persistently high. PMID- 10519609 TI - Prediction of the rates of fertilization, cleavage, and pregnancy success by cumulus-coronal morphology in an in vitro fertilization program. AB - OBJECTIVE: To examine the relation between the grading of cumulus-coronal morphology at oocyte retrieval and the rates of fertilization, cleavage, and pregnancy success in IVF-ET cycles. DESIGN: Retrospective study. SETTING: University-affiliated medical center. PATIENT(S): Infertile women who underwent IVF-ET treatment. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Fertilization and cleavage of the oocytes and the pregnancy outcome. RESULT(S): Mature grade 3 cumulus-oocyte complexes (COCs) constituted the highest percentage among all grades and had a higher fertilization rate than COCs of other grades (77% versus 65%, 43%, and 28% for grades 2, 1, and 4, respectively). The cleavage and polyspermy rates did not correlate with cumulus-coronal morphology grading. The pregnancy rate was higher in cycles with >50% grade 3 COCs than in cycles with < or =50% grade 3 COCs (32% versus 16%). In cycles with >80% grade 3 COCs, the pregnancy rate was 57%. The correlation between the percentage of grade 3 COCs and the pregnancy rate was independent of patient age and the number of COCs retrieved. CONCLUSION(S): The cumulus-coronal morphology grade correlates with the fertilization rate but not with the cleavage or polyspermy rate. In vitro fertilization cycles that have a greater percentage of grade 3 COCs have an increased chance of resulting in pregnancy. The cumulus-coronal morphology grade predicts pregnancy success in IVF-ET cycles. PMID- 10519610 TI - Development and integration of an extended embryo culture program. AB - OBJECTIVE: To study and evaluate a sequential, extended embryo culture system. DESIGN: Prospective study. SETTING: University-affiliated IVF clinic. PATIENT(S): All couples who were treated between October 1997 and July 1998. INTERVENTION(S): A standard human tubal fluid plus 10% serum substitute supplement (SSS) culture medium was used. The embryos were transferred to extended culture medium (S2 or G2) on day 3. MAIN OUTCOME MEASURE(S): Blastocyst formation and implantation and pregnancy rates. RESULT(S): Forty percent of the 20 donated cryopreserved embryos progressed to the blastocyst stage by day 6. Clinically, 7 (5.6%) of the 125 cycles did not result in a transfer. Blastocyst formation rates ranged from 33% 63% in the five study groups. Implantation rates ranged from 15%-52% and pregnancy rates ranged from 37%-75%. CONCLUSION(S): Extended culture to day 5 or 6 results in acceptable blastocyst formation rates, implantation rates, and pregnancy rates. PMID- 10519611 TI - Multifetal pregnancy reduction in cases of threatened abortion of triplets. AB - OBJECTIVE: To investigate the course of pregnancy and fetal outcome after first trimester multifetal pregnancy reduction (MFPR) in patients with triplet pregnancies and uterine bleeding. DESIGN: Case series of patients with threatened triplet pregnancies considered for MFPR. SETTING: Department of Obstetrics and Gynecology, Rabin Medical Center, Petah-Tiqva, Israel. PATIENT(S): Forty-two patients with triplet pregnancies and first-trimester uterine bleeding. INTERVENTION(S): At 10-15 weeks' gestation, MFPR with intracardiac injection of potassium chloride was performed. The procedures were performed 7-10 days after cessation of bleeding (9-13 weeks) or in the presence of minimal uterine bleeding (14-15 weeks). In patients with heavy uterine bleeding, MFPR was postponed. MAIN OUTCOME MEASURE(S): Early- and late-pregnancy complications related to the procedure, pregnancy outcome, and fetal survival. RESULT(S): Performance of MFPR at 14-15 weeks was associated with a higher abortion rate (38.5%), lower mean gestational age at delivery (30.6 weeks), and lower mean twin birth weight (1,376+/-218 g and 1,014+/-202 g) than was performance of MFPR at 10-13 weeks (18.8%, 33.2 weeks, and 1,720+/-245 g and 1,596+/-170 g, respectively). Abortion occurred in four of the five patients with moderate to heavy uterine bleeding who did not undergo MFPR; the fifth patient gave birth prematurely at 28 weeks, and two of the newborns died. CONCLUSION(S): Pregnancy outcome and fetal mortality and morbidity in triplet pregnancy after MFPR are directly correlated with duration and amount of first-trimester bleeding. PMID- 10519612 TI - Lack of association between serum antibodies to Chlamydia trachomatis and a history of recurrent pregnancy loss. AB - OBJECTIVE: To study the relation between recurrent pregnancy loss (RPL) and infection with Chlamydia trachomatis, and to compare the prevalence of antibodies to C. trachomatis in women with primary and secondary RPL. DESIGN: Prospective comparative study. SETTING: University hospital and university student health center. PATIENT(S): Seventy patients with RPL were selected from women attending an RPL outpatient clinic; 40 normal parous women and 94 asymptomatic sexually active women served as controls. INTERVENTION(S): Blood samples were collected during the clinical examinations for RPL. MAIN OUTCOME MEASURE(S): Serum immunoglobulin (Ig) G and IgA antibodies were detected by two independent methods, a recombinant ELISA specific to the genus Chlamydia and microimmunofluorescence testing specific to the species C. trachomatis. RESULT(S): There was no statistically significant difference in the frequencies of IgG or IgA between the women with RPL and the controls. The antibody frequencies were similar in the women with primary and secondary RPL. CONCLUSION(S): The presence of serum antibodies to C. trachomatis is not associated with RPL. Women with primary and secondary RPL do not differ with respect to the prevalence of antichlamydial antibodies. Thus, women with RPL do not benefit from screening for chlamydial IgG or IgA antibodies. PMID- 10519613 TI - The World Health Organization multinational study of breast-feeding and lactational amenorrhea. III. Pregnancy during breast-feeding. World Health Organization Task Force on Methods for the Natural Regulation of Fertility. AB - OBJECTIVE: To determine the risk of pregnancy during lactational amenorrhea relative to infant feeding status. DESIGN: Prospective longitudinal study. SETTING: Five developing and two developed countries. PATIENT(S): Four thousand one hundred eighteen breast-feeding mother-infant pairs. INTERVENTION(S): Infant feeding practices, menstrual status, and pregnancy were measured. MAIN OUTCOME MEASURE(S): Life-table rates of pregnancy. RESULT(S): In the first 6 months after childbirth, cumulative pregnancy rates during amenorrhea, depending on how the end of amenorrhea was defined, ranged from 0.9% (95% confidence interval [CI] = 0%-2%) to 1.2% (95% CI = 0%-2.4%) during full breast-feeding, and from 0.7% (95% CI = 0.1%-1.3%) to 0.8% (95% CI = 0.2%-1.4%) up to the end of partial breast feeding. At 12 months, the rates ranged from 6.6% (95% CI = 1.9%-11.2%) to 7.4% (95% CI = 2.5%-12.3%) during full breast-feeding, and from 3.7% (95% CI = 1.9% 5.5%) to 5.2% (95% CI = 3.1%-7.4%) up to the end of partial breast-feeding. CONCLUSION(S): These results support the Bellagio Consensus on the use of lactational amenorrhea for family planning, and confirm that the lactational amenorrhea method is a viable approach to postpartum contraception. PMID- 10519614 TI - The World Health Organization multinational study of breast-feeding and lactational amenorrhea. IV. Postpartum bleeding and lochia in breast-feeding women. World Health Organization Task Force on Methods for the Natural Regulation of Fertility. AB - OBJECTIVE: To describe and compare the duration of lochia in seven groups of women; to investigate the occurrence of a possible "end-of-puerperium" bleeding episode; and to determine the frequency of bleeding episodes before postpartum day 56, which applies to the practice of the lactational amenorrhea method of contraception. DESIGN: Prospective longitudinal study with fortnightly follow-up, beginning within 7 days of delivery. SETTING: Five developing and two developed countries. PATIENT(S): Four thousand one hundred eighteen breast-feeding women. INTERVENTION(S): Postpartum lochia and all days of bleeding per vaginam were recorded. MAIN OUTCOME MEASURE(S): Duration of lochia, frequency of an end-of puerperium bleeding episode, and frequency of postlochia bleeding episodes within 56 days of delivery. RESULT(S): The median duration of lochia was 27 days; it varied significantly among the centers (range, 22-34 days). In 11% of the women, lochia lasted >40 days. An end-of-puerperium bleeding episode around the 40th day postpartum was reported by 20.3% of the women. Bleeding within 56 days of delivery (separated from lochia by at least 14 days) occurred in 11.3% of the women and usually was followed by a confirmatory bleeding episode 21-70 days later. CONCLUSION(S): The duration of lochia varied significantly among the study populations, and long durations were not unusual. The significance of the end-of puerperium bleeding episode is unknown. Most users of the lactational amenorrhea method will not experience a postlochia bleeding episode before postpartum day 56. PMID- 10519615 TI - Treatment with flutamide improves hyperinsulinemia in women with idiopathic hirsutism. AB - OBJECTIVE: To investigate insulin metabolism and its modifications induced by the administration of flutamide, a specific antiandrogen compound, in women with idiopathic hirsutism (IH) and in nonobese women with polycystic ovary syndrome (PCOS). DESIGN: Prospective, randomized trial. SETTING: Endocrinological Centre of the Department of Obstetrics and Gynecology, University of Caligari, Caligari, Italy. PATIENT(S): Thirty-two women with normal body mass index participated in the study: 11 with clinical and hormonal features of PCOS and 21 age- and weight matched normally cycling women with IH (n = 11) and without IH (n = 10, controls). INTERVENTION(S): Each subject with PCOS or IH was assigned randomly to receive either flutamide tablets (250 mg twice a day) or placebo for > or =5 months. Twelve subjects (6 with PCOS, 6 with IH) received flutamide and 10 (5 with PCOS, 5 with IH) received placebo. All subjects ingested 75 g of glucose and then underwent an oral glucose tolerance test (OGTT), 3-7 days after spontaneous or medroxyprogesterone acetate (5 mg daily for 5 days)-induced menses. In women with PCOS or IH, the OGTT was repeated at the fourth month of treatment. MAIN OUTCOME MEASURE(S): Fasting and OGTT-stimulated levels of glucose, insulin, and C peptide. RESULT(S): Both fasting and OGTT-stimulated levels of insulin and C peptide were significantly higher in women with PCOS and in those with IH than in controls. Placebo did not modify parameters of glucose metabolism. Flutamide was capable of significantly blunting fasting and OGTT-stimulated secretion of insulin only in women with IH. CONCLUSION(S): Hyperinsulinemia exists in women with IH as well as in nonobese women with PCOS. Treatment with flutamide can completely reverse hyperinsulinemia only in women with IH, which suggests that the efficacy of the drug is dependent on peripheral androgen hyperactivity. PMID- 10519616 TI - Lack of insulin-like growth factor binding protein-3 variation after follicle stimulating hormone stimulation in women with polycystic ovary syndrome undergoing in vitro fertilization. AB - OBJECTIVE: To investigate serum and follicular fluid (FF) insulin-like growth factor-I (IGF-I) and insulin-like growth factor binding protein-3 (IGFBP-3) behavior in superstimulated cycles in patients with polycystic ovary syndrome (PCOS). DESIGN: Controlled clinical study. SETTING: Department of Obstetrics and Gynecology, University of Naples. PATIENT(S): Thirty-two patients with regular menses and tubal and/or male factor infertility and 21 patients with PCOS undergoing IVF. INTERVENTION(S): The IVF program used leuprolide acetate suppression followed by sequential hMG in the subsequent cycle. After follicular development, hCG administration was followed 34-36 hours later by oocyte retrieval. MAIN OUTCOME MEASURE(S): E2, GH, IGF-I, and IGFBP-3 assayed by RIA and immunoradiometric assay. RESULT(S): The controls and patients with PCOS showed similar increases in E2 and GH titers in response to FSH stimulation. Serum IGF-I did not change in either group and was equivalent in the FF. Patients with PCOS had a higher FF IGFBP-3 titer and did not show the decrease in serum IGFBP-3 levels of the control group after FSH stimulation. CONCLUSION(S): The apparent failure of IGFBP-3 reduction in patients with PCOS alters IGF-I bioavailability. Increased sequestration of IGF-I affects ovarian steroidogenesis and may explain the poor response to gonadotropin stimulation. PMID- 10519617 TI - Insulin action during variable hyperglycemic-hyperinsulinemic infusions in hyperandrogenic anovulatory patients and healthy women. AB - OBJECTIVE: To determine whether 3-month GnRH analogue (GnRH-a) administration to hyperandrogenic anovulatory patients and healthy women affects glucose utilization or endogenous glucose production (EGP) in the postabsorptive state and during variable hyperglycemic-hyperinsulinemic infusions. DESIGN: Prospective, nonrandomized study. SETTING: Academic research environment. PATIENT(S): Twelve hyperandrogenic anovulatory patients and 11 healthy women matched by body mass index and waist to hip circumference ratio. INTERVENTION(S): Variable hyperglycemic-hyperinsulinemic infusions replicated physiological increases in circulating glucose and insulin levels before and after 3-month GnRH a administration. MAIN OUTCOME MEASURE(S): Glucose utilization and EGP. RESULT(S): In the postabsorptive state, plasma glucose and insulin levels, glucose utilization, and EGP were similar in hyperandrogenic patients and healthy women. During variable hyperglycemic-hyperinsulinemic infusions, glucose use increased and EGP decreased to similar degrees in both groups of women. Three month GnRH-a administration to hyperandrogenic patients and healthy women did not affect plasma glucose and insulin levels, glucose utilization and EGP in the postabsorptive state, or glucose utilization and EGP during variable hyperglycemic-hyperinsulinemic infusions. CONCLUSION(S): Glucose use and EGP in the postabsorptive state and during variable hyperglycemic-hyperinsulinemic infusions are similar in hyperandrogenic anovulatory patients and healthy women of similar body fat distribution and are unaffected by 3-month GnRH-a administration. PMID- 10519618 TI - Identification of seminiferous tubule aberrations and a low incidence of testicular microliths associated with the development of azoospermia. AB - OBJECTIVE: To evaluate the use of percutaneous testicular sperm aspiration in the assessment of azoospermia and its association with seminiferous tubule microliths. DESIGN: Case report. SETTING: Tertiary care fertility center in a university hospital. PATIENT(S): Male undergoing infertility evaluation. INTERVENTION(S): Testicular biopsy and percutaneous testicular aspiration. MAIN OUTCOME MEASURE(S): Serum hormone analysis, sperm concentration in semen, spermatogenesis in samples from testicular biopsies and aspirations, and microlith composition. RESULT(S): A patient presented for infertility evaluation with a history of severe oligospermia that progressed to azoospermia. The serum testosterone concentration (357 ng/dL) and LH concentration (9.2 mIU/mL) were normal and the serum FSH concentration (18.3 mIU/mL) was elevated. Testicular biopsy results indicated spermatogenic hypoplasia with limited spermatozoa. Seminiferous tubules obtained by percutaneous testicular aspiration were structurally aberrant, with multiple diverticula. Microliths averaging 120 microm in diameter were observed within and blocking the seminiferous tubules. The microliths were composed of calcium phosphate (hydroxyapatite) in both the core and peripheral regions. Electron microscopy revealed a high degree of collagen like material within the peripheral zone. CONCLUSION(S): The presence of seminiferous tubule microliths is associated with the development of azoospermia. In patients with a low incidence of seminiferous tubule microliths and aberrant seminiferous tubule architecture, percutaneous testicular aspiration may provide a diagnostic advantage over testicular biopsy. PMID- 10519619 TI - Aneuploidy frequencies in semen fractions from ten oligoasthenoteratozoospermic patients donating sperm for intracytoplasmic sperm injection. AB - OBJECTIVE: To determine aneuploidy frequencies in pellet and swim-up semen fractions from 10 infertile men with severe oligoasthenoteratozoospermia (OAT) who were donating sperm for intracytoplasmic sperm injection and to determine whether the swim-up isolation method would successfully separate aneuploid from haploid sperm. DESIGN: Prospective study. SETTING: Infertility clinic and molecular genetics laboratory. PATIENT(S): Ten patients with severe OAT. INTERVENTION(S): Cytogenetic analyses by fluorescence in situ hybridization to determine aneuploidy frequencies for chromosomes 1, 13, 18, 21, X, and Y in sperm from swim-up and pellet fractions. MAIN OUTCOME MEASURE(S): Gametic aneuploidy was scored in sperm fractions separated by the swim-up technique and clinical results after intracytoplasmic sperm injection were tabulated. RESULT(S): In all cases, chromosome aneuploidy levels in patients were significantly greater than in controls. The type and percentage of aneuploid sperm for all patients with OAT found in both swim-up and pellet fractions were not different, with the exception of diploid sperm, which remained in the pellet fraction. After ET, 2 (20%) of 10 couples achieved successful pregnancies. CONCLUSION(S): The data show significantly higher rates of diploidy, autosomal disomy and nullisomy, sex chromosome disomy and nullisomy, and total aneuploidy in sperm from all separated fractions obtained from all patients with OAT versus controls. This patient population with OAT may be at increased risk of producing aneuploid offspring. PMID- 10519620 TI - Advanced surgical sperm recovery is a viable option for intracytoplasmic sperm injection in patients with obstructive or nonobstructive azoospermia. AB - OBJECTIVE: To determine whether advanced sperm retrieval is appropriate in cases of obstructive and nonobstructive azoospermia. DESIGN: Prospective controlled study. SETTING: Tertiary care center. PATIENT(S): Men with obstructive and nonobstructive azoospermia, and their partners. INTERVENTION(S): Surgical sperm retrieval followed by intracytoplasmic sperm injection (ICSI) after 4 or 48 hours. MAIN OUTCOME MEASURE(S): Fertilization and pregnancy rates. RESULT(S): Advanced and fresh surgical sperm recoveries for ICSI were performed in 54 and 230 cycles, respectively. Patient demographics and cycle parameters were comparable. Two hundred forty-one (56.3%) of 428 injected eggs in the advanced retrieval group were fertilized, compared with 955 (56.6%) of 1,686 eggs in the fresh retrieved group (P=.94). There was no statistically significant difference in the pregnancy rates per ET between groups: 38.2% (18 of 47) in the advanced retrieval group and 39.9% (73 of 183) in the fresh sperm recovery group (P=.97). CONCLUSION(S): Testicular and epididymal sperm recovery can be safely performed 48 hours before ICSI. This facilitates planning, and, in cases of failure to retrieve sperm, hCG administration and ovum pick-up can be canceled, thereby reducing costs and eliminating the risk of ovarian hyperstimulation. PMID- 10519621 TI - Antioxidant supplementation in vitro does not improve human sperm motility. AB - OBJECTIVE: To determine the effects of supplementation of preparation media with ascorbate and alpha-tocopherol on subsequent sperm motility and reactive oxygen species production. DESIGN: Prospective study to analyze postpreparation human sperm motility parameters and reactive oxygen species production following antioxidant supplementation. SETTING: Andrology Laboratory, Royal Maternity Hospital, Belfast, Northern Ireland. PATIENT(S): Sixty patients attending the Andrology Laboratory for semen analysis. INTERVENTION(S): Normozoospermic and asthenozoospermic semen samples (n = 10 for each control and antioxidant group) were prepared by Percoll density centrifugation (95%-47.5%) in media supplemented with ascorbate or alpha-tocopherol to different concentrations within physiologic levels. Controls were included that were not exposed to antioxidant. MAIN OUTCOME MEASURE(S): Sperm motility parameters were assessed using computer-assisted semen analysis. The generation of reactive oxygen species was determined using luminol dependent chemiluminescence. RESULT(S): The production of reactive oxygen species by sperm was reduced by supplementation in vitro with ascorbate and alpha tocopherol. However, progressive motility, average path velocity, curvilinear velocity, straight-line velocity, and linearity were decreased significantly, with the greatest inhibition observed with the highest concentrations of antioxidants. CONCLUSION(S): Supplementation of preparation media with ascorbate and alpha-tocopherol, either singly or in combination, is not beneficial to sperm motility. PMID- 10519622 TI - Potential adverse effect of semen processing on human sperm deoxyribonucleic acid integrity. AB - OBJECTIVE: To examine the effect of standard Percoll density-gradient centrifugation on human sperm DNA denaturation. DESIGN: Prospective, observational study. SETTING: University-based infertility clinic. PATIENT(S): Twenty-five nonazoospermic men. INTERVENTION(S): Semen samples (n = 25) were obtained from consecutively seen nonazoospermic men presenting for infertility evaluation. Samples were processed by two-layer and four-layer Percoll density gradients. Sperm motility and sperm chromatin structure (evaluated by flow cytometry analysis of acridine orange-treated spermatozoa) were monitored before and after semen processing. Sperm chromatin integrity was expressed as the percentage of spermatozoa that demonstrated denatured DNA. MAIN OUTCOME MEASURE(S): Sperm motility and DNA integrity. RESULT(S): Mean sperm motility improved significantly after processing with two-layer and four-layer Percoll gradients compared with whole semen (54% and 57% motility versus 44% motility, respectively). In contrast, the percentage of sperm with denatured DNA increased after processing with two-layer and four-layer Percoll gradients compared with whole semen (34% and 32% versus 18%, respectively). CONCLUSION(S): Our data demonstrate that the improvement seen in sperm motility after Percoll processing is not associated with a similar improvement in sperm DNA integrity. These data suggest that we reexamine current sperm processing techniques to minimize sperm DNA damage and the potential transmission of genetic mutations in assisted reproductive cycles. PMID- 10519623 TI - Influence of age, diagnosis, and cycle number on pregnancy rates with gonadotropin-induced controlled ovarian hyperstimulation and intrauterine insemination. AB - OBJECTIVE: To determine whether age, diagnosis, and cycle number influence cycle fecundity associated with gonadotropin-induced controlled ovarian hyperstimulation/IUI. DESIGN: Retrospective analysis. SETTING: The Center for Reproductive Medicine at the Brigham and Women's Hospital, a tertiary care academic medical center. PATIENT(S): Two hundred seventy-four women who underwent controlled ovarian hyperstimulation with gonadotropins and IUI. INTERVENTION(S): Infertility treatment with gonadotropins and IUI. MAIN OUTCOME MEASURE(S): Pregnancy rates according to patient age, infertility diagnosis, and number of treatment cycles. RESULT(S): Pregnancy rates decreased with increasing patient age. The cumulative pregnancy rates varied greatly by diagnosis from 13% for patients with male factor infertility to 84% for patients with ovulatory factor infertility. Average cycle fecundity was considerably less varied by diagnosis. All pregnancies among patients with male factor infertility and tubal factor infertility were achieved during the first two cycles. CONCLUSION(S): There is a clear age-related decline in fecundity associated with gonadotropin-induced controlled ovarian hyperstimulation/IUI. Patients <40 years of age and those with male factor infertility or tubal factor infertility have a particularly poor prognosis. PMID- 10519624 TI - A levonorgestrel-releasing intrauterine system for the treatment of dysmenorrhea associated with endometriosis: a pilot study. AB - OBJECTIVE: To evaluate the efficacy and safety of an intrauterine system releasing 20 microg of levonorgestrel per 24 hours in the long-term treatment of recurrent dysmenorrhea in women already operated on conservatively for endometriosis. DESIGN: A prospective noncomparative pilot study. SETTING: A tertiary care and referral academic center for patients with endometriosis. PATIENT(S): Twenty parous women with recurrent moderate or severe dysmenorrhea after conservative surgery for endometriosis who did not want further children. INTERVENTION(S): A levonorgestrel-releasing intrauterine system was inserted in each woman within 7 days of the start of a menstrual cycle. MAIN OUTCOME MEASURE(S): Variations in severity of dysmenorrhea during treatment according to a 100-mm visual analogue scale and a 0-3-point verbal rating scale, modification of a pictorial blood-loss assessment chart devised to evaluate the amount of menstrual flow, and degree of satisfaction after 12 months of therapy. RESULT(S): One woman was lost to follow-up after achieving amenorrhea and expressing satisfaction, and 1 requested system removal because of weight gain and abdominal bloating. In another subject, the levonorgestrel intrauterine system was expelled 3 months after insertion. The menstrual patterns in the remaining 17 women were characterized by amenorrhea in 4 cases, hypomenorrhea or spotting in 8, and normal flow in 5. Baseline and 12-month follow-up mean +/- SD blood loss scores were 111+/-36 and 27+/-26, respectively. At the same time, mean +/- SD visual analogue and verbal rating scale scores dropped, respectively, from 76+/-12 to 34+/-23 points and from 2.5+/-0.5 to 1.2+/-0.5 points. Four women were very satisfied with treatment, 11 were satisfied, 2 were uncertain, and 3 were dissatisfied at 12-month follow-up. CONCLUSION(S): Because of the amenorrhea or hypomenorrhea induced in most women, a levonorgestrel intrauterine system greatly reduced menstrual pain associated with endometriosis and achieved a high degree of patient satisfaction. PMID- 10519625 TI - Electrical activation and in vitro development of human oocytes that fail to fertilize after intracytoplasmic sperm injection. AB - OBJECTIVE: To determine whether electrically stimulated Ca2+ influx can "rescue" fertilization and early embryogenesis in human oocytes that fail to fertilize after intracytoplasmic sperm injection (ICSI). DESIGN: Prospective, randomized trial of a laboratory procedure. SETTING: A research laboratory at a university medical center. PATIENT(S): Discarded oocytes from ICSI-IVF cycles. INTERVENTION(S): Oocytes (n = 104) that showed no evidence of fertilization 16-24 hours after ICSI were assigned to three treatment groups: group 1 (one direct current electrical pulse at 1.36-1.50 kV/cm for 40-60 micros), group 2 (three pulses every 15-20 minutes), or group 3 (treated the same as group 2 but with no electrical stimulation). MAIN OUTCOME MEASURE(S): After stimulation, the oocytes were cultured in vitro for 3-5 days. Oocytes that displayed two pronuclei and a second polar body within 16 hours were considered to have fertilized normally. Fertilization and embryo cleavage rates were compared between groups. RESULT(S): Fertilization occurred in 26 (70%) of 37 and 38 (78%) of 49 group 1 and 2 oocytes, respectively, but in only 5 (27%) of 18 group 3 oocytes. Within 3 days, group 2 embryos routinely developed beyond the two-cell to four-cell stage (61% versus 13% in group 1); 11% of these oocytes developed to the morula or early blastocyst stage. Sex chromosome analyses indicated 10 male and 8 female embryos. CONCLUSION(S): Oocytes that fail to fertilize by 24 hours after ICSI can resume apparently normal fertilization and early embryonic development in response to electrical stimulation. Moreover, the degree of cytoplasmic activation as determined by the number of pulses applied affects fertilization efficiency and early embryonic development. PMID- 10519626 TI - Heterotopic ovarian transplantation without vascular pedicle in syngeneic Lewis rats: six-month control of estradiol and follicle-stimulating hormone concentrations after intraperitoneal and subcutaneous implants. AB - OBJECTIVE: To assess the recovery, maintenance, and quality of ovarian function by comparing the success of autotransplantation in intraperitoneal (IP) and SC locations over a 6-month period in syngeneic Lewis rats. DESIGN: Experimental animal study. SETTING: Unit of Experimental Research at the Barcelona University School of Medicine. ANIMAL(S): Female syngeneic Lewis rats. INTERVENTION(S): The animals were randomized to one of three groups: group A, control group with ovariectomy (n = 15); group B, ovariectomy and IP autologous heterotopic transplant of ovarian tissue without vascular pedicle (n = 14); and group C, ovariectomy and SC autologous heterotopic transplant (n = 15). MAIN OUTCOME MEASURE(S): Serum levels of FSH and 17beta-E2 and vaginal cytology. RESULT(S): In groups B and C, E2 serum concentrations from day 7 and day 10 onward remained comparable to basal levels, and significantly higher than in group A, throughout the 6-month period. In group B from day 7 after surgery onward, and in group C from day 10 after surgery onward, FSH concentrations remained low (comparable to basal levels) throughout the follow-up period. Vaginal cytology of groups B and C showed trophic maturation between days 4 and 10 after ovariectomy and insertion of the ovarian tissue implant, whereas the control group remained atrophic. There were no statistically significant differences between IP and SC implants. CONCLUSION(S): A heterotopic autotransplant of ovarian tissue without vascular pedicle in syngeneic Lewis rats is successful for > or =6 months. PMID- 10519627 TI - Effects of misoprostol on lipoprotein (a) levels of ovariectomized rats. AB - OBJECTIVE: To determine the effects of misoprostol on plasma lipoprotein (a) concentrations of ovariectomized rats. DESIGN: Controlled prospective study. SETTING: Animal research laboratory. ANIMAL(S): Four-month-old female Sprague Dawley rats. INTERVENTION(S): Blood samples were obtained before and 60 days after ovariectomy, and the rats were divided into three groups. Group I (five rats) was treated with vehicle (water); groups II and III (nine and eight rats, respectively) were treated with oral misoprostol at 100 and 200 microg/kg/d, respectively, for 60 days, after which blood was drawn again. MAIN OUTCOME MEASURE(S): Serum lipoprotein (a) levels. RESULT(S): The median lipoprotein (a) level before ovariectomy was 10.8 mg/dL (range, 10.6-46.5 mg/dL). Sixty days after ovariectomy, the level increased significantly to 15.9 mg/dL (range, 10.6 36.9 mg/dL). After treatment, there was no change in lipoprotein (a) levels in the vehicle-treated group (range, 16.3-21.1 mg/dL); however, the lipoprotein (a) levels decreased significantly in the group treated with 100 microg/kg/d of misoprostol, from 15.4 mg/dL to 10.8 mg/dL, and in the group treated with 200 microg/kg/d of misoprostol, from 17.1 mg/dL to 10.6 mg/dL. CONCLUSION(S): Misoprostol caused a significant decrease in lipoprotein (a) levels. PMID- 10519628 TI - Expression of alpha6 and beta4 integrin subunits throughout the menstrual cycle: no correlation with uterine receptivity. AB - OBJECTIVE: To compare the expression of alpha6 and beta4 integrin subunit levels throughout the menstrual cycle in the endometrium of healthy women and infertile women. DESIGN: Prospective study. SETTING: An academic teaching hospital. PATIENT(S): Endometrium was collected from 58 women, including healthy subjects (n = 28) and patients undergoing diagnostic testing for infertility (n = 30). INTERVENTION(S): Endometrial biopsies were performed throughout the menstrual cycle in healthy women and infertile women. MAIN OUTCOME MEASURE(S): Immunohistochemical staining intensity of alpha6 and beta4 using the semiquantitative immunohistochemical score, compared using regression analysis and analysis of variance with Scheffe's correction. RESULT(S): There was no correlation between menstrual cycle phase and endometrial integrin subunit alpha6 or beta4 on glandular or luminal epithelium. Even women with identified luteal phase defects had indistinguishable patterns of expression for these integrin subunits during the window of implantation. CONCLUSION(S): The integrin subunits alpha6 and beta4 are expressed uniformly throughout the menstrual cycle on glandular and luminal epithelium. These integrins may have a housekeeping role in anchoring endometrial epithelium and do not appear to be useful markers of uterine receptivity. PMID- 10519629 TI - Survival of cryopreservation and thawing with all blastomeres intact identifies multicell embryos with superior frozen embryo transfer outcome. AB - OBJECTIVE: To assess the impact of survival of cryopreservation and thawing with all blastomeres intact on the outcome of multicell frozen ET. DESIGN: Retrospective study. SETTING: Academic assisted reproductive technology program. PATIENT(S): One hundred sixteen exclusively multicell frozen ETs in 78 patients. INTERVENTION(S): Frozen ET. MAIN OUTCOME MEASURE(S): Relation of embryonic blastomere survival to the outcome of frozen ET (i.e., pregnancy). RESULT(S): When at least one embryo survived with all blastomeres intact, the total pregnancy rate (biochemical, clinical, or delivered) was 37.7%, the clinical pregnancy rate was 24.6%, and the delivered pregnancy rate was 18.8%. When no embryo survived with all blastomeres intact, the corresponding rates were 10.6%, 8.5%, and 6.4%. The differences in the total pregnancy rate and the clinical pregnancy rate were statistically significant. The delivered pregnancy rates approached statistical significance. CONCLUSION(S): Multicell embryonic survival of cryopreservation and thawing with all blastomeres intact identifies embryos with superior developmental potential. PMID- 10519630 TI - Effects of peritoneal macrophages from women with endometriosis on endometrial cellular proliferation in an in vitro coculture model. AB - OBJECTIVE: To study the effects of peritoneal macrophages on endometrial cellular proliferation in an in vitro coculture model and to compare the magnitude of these effects between macrophages from women with endometriosis and normal women. DESIGN: Controlled study of peritoneal macrophage function. SETTING: University hospital. PATIENT(S): Patients with a normal peritoneal cavity (n = 15) and with pelvic endometriosis (n = 20) undergoing laparoscopy. INTERVENTION(S): Peritoneal macrophages were cocultured with endometrial epithelial and stromal cells; endometrial cell cultures without macrophage coculture acted as controls. MAIN OUTCOME MEASURE(S): Endometrial cellular proliferation measured by 3H-thymidine incorporation. RESULT(S): Endometrial epithelial cells cocultured with peritoneal macrophages from women with endometriosis showed significantly increased proliferation compared with cocultures using macrophages from normal women when assessed at 24 hours (1.56 versus 1.03 times, respectively, over control) and at 72 hours (1.55 versus 1.10 times over control). Endometrial stromal cells cocultured with peritoneal macrophages from women with endometriosis similarly exhibited increased proliferation compared with cocultures using macrophages from normal women when assessed at 24 hours (1.65 versus 1.17 times over control) and at 72 hours (1.65 versus 1.21 times over control). CONCLUSION(S): Peritoneal macrophages of patients with endometriosis stimulate cellular proliferation of endometrial epithelial and stromal cells in vitro. PMID- 10519631 TI - Birth of a healthy infant after conception with round spermatids isolated from cryopreserved testicular tissue. AB - OBJECTIVE: To report a case of nonobstructive azoospermia in which round spermatids recovered from thawed testicular tissue were used for injection. DESIGN: Case report. SETTING: Reproductive Medicine Unit, S.I.S.ME.R. PATIENT(S): A 33-year-old azoospermic man. INTERVENTION(S): Intracytoplasmic sperm injection with frozen-thawed spermatids. MAIN OUTCOME MEASURE(S): Fertilization, embryo cleavage, pregnancy, and delivery. RESULT(S): Birth of a healthy, chromosomally normal girl. CONCLUSION(S): Frozen-thawed testicular round spermatids from a patient with a history of incomplete spermatogenesis can maintain their viability and their capacity to fertilize and to lead to full-term pregnancy. PMID- 10519632 TI - Value of estradiol response after human chorionic gonadotropin administration in predicting in vitro fertilization success. AB - OBJECTIVE: To determine whether the serum E2 response after the administration of exogenous hCG is predictive of outcome during IVF. DESIGN: Prospective, noncomparative cohort. SETTING: Two academic centers and one private-practice IVF program. PATIENT(S): Two hundred twenty-two couples undergoing IVF for infertility arising from ovarian dysfunction, asthenoteratospermia, endometriosis, tubal disease, or unexplained infertility. MAIN OUTCOME MEASURE(S): Implantation, pregnancy, and miscarriage rates were compared in cycles that demonstrated an increase, decrease, or plateau in the serum E2 level on the day after hCG administration. The effects of age, cause of infertility, and maximum E2 value on outcome were evaluated. RESULT(S): Ninety-two cycles resulted in a clinical pregnancy and 130 cycles failed. Of 115 cycles in which the E2 level rose, 42 (37%) resulted in an ongoing pregnancy; among cycles with plateauing E2 responses, 20 of 69 (29%) resulted in a pregnancy. Fifteen of 38 (39%) of cycles exhibiting a drop in serum E2 resulted in an ongoing pregnancy. No statistically significant differences in ongoing pregnancy rates were noted in the increasing, plateauing, or decreasing E2 response groups. CONCLUSION(S): E2 values obtained on the day after hCG administration are not predictive of outcome in women undergoing IVF. PMID- 10519633 TI - Relaxin-like factor expression in a human ovarian Sertoli-Leydig cell tumor. AB - Relaxin-like factor (RLF), a new member of the insulin-like growth factor family, is a reliable marker for normal Leydig cells in the human postpubertal testis (1). Expression of the RLF gene appears to be developmentally regulated, given that only during puberty is RLF expression up-regulated. We recently demonstrated down-regulation of the human RLF gene in testicular Leydig cell tumors indicating dedifferentiation of the Leydig cells within the tumor (2). Ovarian Sertoli Leydig cell tumors (SLCTs), histologically typed as androblastomas, are rare, potentially malignant sex-cord stromal tumors exhibiting testicular-like structure and differentiation of various degrees. In the present study, we investigated the expression of RLF, 17alpha-hydroxylase, 3beta-hydroxysteroid dehydrogenase (3beta-HSD), Ki-67, and cytokeratin 18 in a human ovarian SLCT of low differentiation. PMID- 10519634 TI - Outpatient laparoscopic tubal anastomosis and subsequent fertility. AB - OBJECTIVE: To determine the reproductive outcome of women who undergo laparoscopic tubal anastomosis. DESIGN: Observational prospective study. SETTING: University-affiliated infertility medical center. PATIENT(S): One hundred two patients seeking reversal of tubal sterilization. INTERVENTION(S): Laparoscopic tubal anastomosis was performed with a one-suture technique. MAIN OUTCOME MEASURE(S): Pregnancy rate. RESULT(S): There were 69 isthmic-isthmic, 16 isthmic ampullary, 12 cornual-isthmic, and 5 ampullary-ampullary anastomoses. The mean operative time was 71.35 minutes. Eight patients had bilateral tubal obstruction on postoperative hysterosalpingography. Sixty-nine patients (70%) conceived. Sixty-four (65.3%) had ongoing intrauterine pregnancies, 15 (21.7%) had spontaneous abortions, and 5 (7.2%) had ectopic pregnancies. CONCLUSION(S): This study demonstrates that laparoscopic tubal anastomosis can be done safely and successfully on an outpatient basis, reducing costs and postoperative morbidity while accelerating the patient's return to normal activities. PMID- 10519635 TI - Resectoscopic treatment of uterus didelphys with unilateral imperforate vagina complicated by hematocolpos and hematometra: case report. AB - OBJECTIVE: To describe a technique for treating hematocolpos and hematometra in patients with uterus didelphys and unilateral imperforate vagina involving the use of resectoscopy under ultrasonographic control. DESIGN: Case report. SETTING: University hospital. PATIENT(S): A 13-year-old girl with uterus didelphys with unilateral hematometra, hematocolpos, and ipsilateral renal agenesis. The girl complained of severe abdominal pain, which appeared with each of her menses. INTERVENTION(S): The intervention was performed by a vaginoscopic approach to preserve the integrity of the hymen. The first incision on the vaginal wall was performed in correspondence with the hematocolpos under continuous ultrasonographic guidance with the use of a straight resectoscopic loop. Resection of the vaginal septum was continued with the use of an angled resectoscopic loop until almost complete excision of the septum was achieved. MAIN OUTCOME MEASURE(S): Clinical, echographic, and vaginoscopic findings before the operation and 2 and 6 months after the operation. RESULT(S): The surgical procedure was easy to perform. Almost complete excision of the septum was achieved with just a few passages of the resectoscope. Complete drainage of both the hematocolpos and the hematometra was confirmed by ultrasonography. The postoperative period was completely uneventful. Clinical and vaginoscopic evaluations 6 months after the operation confirmed the integrity of the hymen, the complete resolution of clinical symptoms, and the persistence of a large communication between the two vaginas. CONCLUSION(S): Resectoscopic excision under ultrasonographic guidance of the vaginal septum in a girl with uterus didelphys with unilateral hematometra and hematocolpos was effective and easy to perform, and it fully respected the integrity of the reproductive system. PMID- 10519636 TI - Increased recovery of swim-up spermatozoa by application of "antigravitational" centrifugation. AB - OBJECTIVE: To compare the quantity and quality of sperm recovered following the 10-minute application of an antigravitational force during the swim-up procedure with the standard 60-minute swim-up (SSU) procedure. DESIGN: Prospectively controlled in vitro study. SETTING: Private andrology laboratory and hospital based infertility practice. INTERVENTION(S): Equal aliquots of semen were evaluated following various intervals of antigravitational centrifugation swim-up (ACSU). ACSU and SSU were then compared. PATIENT(S): Thirty-eight men undergoing therapeutic testing. MAIN OUTCOME MEASURES: Sperm concentration, sperm motility, and progressive sperm motility. RESULTS: The number of sperm recovered from the ACSU procedure was significantly higher than from the SSU procedure. No significant differences in percent motile sperm and progressive motile sperm recovery between the two procedures were observed. CONCLUSION: The ACSU procedure yields a higher number of motile spermatozoa in a much shorter time. PMID- 10519637 TI - Delay in the application of the cover glass is a potential source of error with the Makler Counting Chamber. AB - OBJECTIVE: To determine the effect of delaying the application of the cover glass of a Makler Counting Chamber (Sefi Medical Instruments, Haifa, Israel) on the apparent concentration of sperm or Dynabeads (Dynal Pty Ltd, Carlton, Victoria, Australia). DESIGN: Systematic alteration of the time taken to apply the cover glass. SETTING: Comprehensive infertility service. PATIENT(S): Fresh semen from a patient or washed sperm from a donor. INTERVENTION(S): For the Makler Counting Chamber, a 5-microL drop of the sperm or bead suspension was placed in the center of the lower platform and the cover glass was applied either immediately (0 seconds) or after a delay of 5, 10, or 30 seconds. MAIN OUTCOME MEASURE(S): The concentration of beads or sperm counted using an Improved Neubauer Hemocytometer or a Makler Counting Chamber. RESULT(S): There was a progressive increase in sperm concentration with a longer delay in applying the cover glass when sperm was suspended in either seminal plasma or culture medium. Repeating the experiments with Dynabeads resuspended in phosphate-buffered saline, serum, or seminal plasma showed the same trend, although the suspension of beads in serum seemed to settle the least over the first 10 seconds. CONCLUSION(S): Delay in the application of the cover glass is a potential source of error with the Makler Counting Chamber and should be avoided. PMID- 10519638 TI - Adjusting for "treatment-independent pregnancies". PMID- 10519639 TI - "Female power"--and a straight look into the female past. PMID- 10519640 TI - "Female power"--and a straight look into the female past. PMID- 10519641 TI - Needle insertion for pneumoperitoneum! PMID- 10519642 TI - Residual effect of clomiphene citrate? PMID- 10519643 TI - The role of hydrosalpinx in IVF: are we getting closer to an answer? PMID- 10519644 TI - Is antivenom the most successful therapy in scorpion victims? PMID- 10519645 TI - Purification and characterization of a platelet aggregation inhibitor acidic phospholipase A2 from Indian saw-scaled viper (Echis carinatus) venom. AB - An acidic phospholipase A2 (EC-I-PLA2) has been purified from the Indian saw scaled viper (Echis carinatus) venom through a combination of column chromatography and electrophoresis. EC-I-PLA2 has a molecular weight of 16000 by SDS-PAGE. It was focussed between pH 4.2 and 4.8 by isoelectro focussing. EC-I PLA2 was non-lethal to mice and devoid of neurotoxicity, myotoxicity, anticoagulant activity and cytotoxicity. It induced mild oedema in the foot pads of mice. The purified PLA2 inhibited ADP, collagen and epinephrine induced human platelet aggregation and the inhibition was both dose and time dependent. PMID- 10519646 TI - Primary culture of venom gland cells from the South American rattlesnake (Crotalus durissus terrificus). AB - Primary cultures of venom gland cells from the South American rattlesnake (Crotalus durissus terrificus) were attempted. At first, six different cell types were obtained including potentially secreting epithelial-like cells. Nonepithelial cell cultures were later invaded by fibroblast-like cells. Cultures of epithelial-like gland cells were successfully maintained, after testing different culture conditions by varying the media, incubation temperature, use of dissociating agents and adhesion substrates. The best results were achieved using plates precoated with rattlesnake skin collagen and incubation in CMRL 1415 modified for snake gland cells plus 10% fetal calf serum at 30 degrees C. The presence of venom could be demonstrated in the supernatant of five out of six epithelial-like gland cell cultures tested by ELISA, in the very first passages. After the third passage, however, venom amounts dropped to undetectable values. A total of 23 venom gland cell lines were obtained and are kept frozen in the laboratory; among them, five epithelial-like gland cell lines with up to 12 passages, that were continuously cultured for more than 30 weeks. The methodology described here was successfully applied to C. d. terrificus kidney cells culturing, developed to be used as negative control. PMID- 10519647 TI - Aromatic substitutions in alpha-conotoxin ImI. Synthesis of iodinated photoactivatable derivative. AB - Conotoxin ImI is a specific marker of alpha7 nicotinic acetylcholine receptors. To study the role of aromatic indole group of tryptophan 10 in biological activity of ImI, the analogue containing tyrosine at this position was synthesized by solid-phase peptide synthesis. The analogue obtained, as well as its iodinated derivatives, were shown to be active against rat brain alpha7 acetylcholine receptor expressed in Xenopus oocytes. Attachment of bulky aromatic p-benzoylbenzoyl group to N-terminal alpha-amino group of iodinated [Tyr10]ImI only slightly affected the biological activity of the analogue. The data obtained suggest that indole ring of tryptophan 10 is not absolutely necessary for biological activity of conotoxin ImI, and that the N-terminus can accommodate a large aromatic group without loss of biological activity. PMID- 10519649 TI - Toxicity of French strains of the dinoflagellate Prorocentrum minimum experimental and natural contaminations of mussels. AB - Mediterranean strains of Prorocentrum minimum do not appear to have the same toxic component as Japanese strains since they showed no cytotoxicity for hepatocytes in culture. However, their toxic components, which appear to block calcium channels, were detectable by the immobilisation test on Diptera larvae. A bio-accumulation experiment in the laboratory showed that the toxins could accumulate in nearly equivalent amounts in the hepatopancreas and meat of cultured mussels. The same toxicity was found in natural samples collected in a period of bloom of P. minimum. These results suggest that P. minimum could be responsible for shellfish toxicity in the natural environment and thus present a risk for human health. PMID- 10519648 TI - Indian red scorpion (Buthus tamulus) venom-induced augmentation of cardiac reflexes is mediated through the involvement of peripheral 5-HT3 and central 5 HT1A receptor subtypes. AB - The present study was undertaken to identify 5-HT receptor subtypes involved in Buthus tamulus (BT) venom-induced augmentation of cardiac reflexes elicited by phenyldiguanide (PDG). Intravenous injection of PDG (10 microg/kg) produced parallel decrease in mean arterial pressure (MAP) and heart rate (HR) in urethane anaesthetized rats (r=0.82; p < 0.001). Injection of PDG (1-40 microg/kg, i.v.) produced concentration-dependent decrease in time-response area of the HR. After BT venom (20 microg/kg) the concentration-response curve was shifted to the left. Further, fall of MAP and HR in response to submaximal concentration of PDG (10 microg/kg) were augmented significantly. Pretreatment with 5-HT3 receptor antagonist (ondansetron; 10 microg/kg) intravenously, blocked the BT venom induced augmentation of PDG reflex but spiperone (100 microg/kg; 5-HT1A/5-HT2 antagonist) or ketanserin (300 microg/kg; 5-HT2 antagonist) failed to do so. Afferent discharges elicited by PDG (10 microg/kg) in vagus nerve were doubled after exposure to BT venom. Ondansetron (100 microg/kg, i.v.) totally abolished the discharges after exposure to BT venom but not by spiperone or ketanserin. Intracerebroventricular injection of spiperone (100 microg/kg) but not ketanserin or ondansetron, blocked the BT venom-induced augmentation of PDG reflex. Results show that the BT venom-induced augmentation of reflex elicited by PDG is mediated through the involvement of 5-HT3 receptors peripherally and 5-HT1A type of receptors centrally. PMID- 10519650 TI - A pharmacological examination of venom from the Papuan taipan (Oxyuranus scutellatus canni). AB - The Papuan taipan (Oxyuranus scutellatus canni) is the third most venomous terrestrial snake in the world, however, little is know about the pharmacology of the venom. In the chick biventer cervicis muscle, venom (10 microg/ml) abolished nerve-mediated twitches (time to 90% inhibition (t90) 44+/-5 min, n = 9). This inhibition was unaffected by prior incubation of the venom with the phospholipase A inhibitor 4-bromophenacyl bromide (4-BPB; 0.72 mM) (t90 48+/-7 min, n = 8). The mouse phrenic nerve diaphragm preparation displayed greater sensitivity to venom (10 microg/ml) (t90 25+/-1 min, n = 6). In the chick biventer muscle, venom (10 microg/ml) significantly inhibited responses to acetylcholine (1 mM) and carbachol (20 microM), but not KCI (40 mM), indicating activity at post-synaptic nicotinic receptors. Venom (10 microg/ ml) did not affect direct muscle stimulation. Venom (3-30 microg/ml) produced dose-dependant contractions of the guinea-pig ileum. Contractile responses were significantly inhibited by indomethacin (1 microM) or prior incubation of the venom with 4-BPB (0.72 mM) indicating involvement of a PLA component. In rat phenylephrine (0.3 microM) precontracted aortae, venom (3-100 microg/ml) produced endothelium-independent relaxation which was unaffected by prior incubation of venom (30 microg/ml) with 4-BPB (0.72 mM). In anaesthetised rats, 10 microg/kg (i.v.) venom produced rapid respiratory and cardiovascular collapse while 5 microg/kg (i.v.) venom produced only a small transient decrease in mean arterial blood pressure. Prior administration of 5 microg/kg (i.v.) venom enabled subsequent administration of 10 and 100 microg/kg (i.v.) venom without respiratory or cardiovascular collapse. Further work is required to identify specific toxins with the above pharmacological activity. PMID- 10519651 TI - Characterization of a basic phospholipase A2-homologeu myotoxin isolated from the venom of the snake Bothrops neuwiedii (yarara chica) from Argentina. AB - A basic protein was isolated by CM-Sephadex C-25 chromatography from the venom of Bothrops neuwiedii from Argentina, and named B. neuwiedii myotoxin I. This protein exerted local myotoxic and edema-forming effects in mice, with potencies comparable to other myotoxins isolated from Bothrops spp. venoms. When injected by i.v. route at doses up to 4.7 mg/kg of body weight, the toxin was not lethal. In vitro, the toxin had no detectable phospholipase A2 activity on egg yolk phospholipids. B. neuwiedii myotoxin I appeared as a homodimer in sodium dodecylsulphate-polyacrylamide gel electrophoresis, with a subunit molecular weight of 15 kD. Gel immunodiffusion revealed a pattern of partial antigenic identity between the newly isolated myotoxin and myotoxin II from Bothrops asper venom. The sequence of B. neuwiedii myotoxin I was determined for the first 40 amino acid residues, showing high homology to several class II phospholipase A2 myotoxins of the Lys-49 family from crotalids. Altogether, results suggest that this toxin is a new member of the Lys-49 phospholipase A2-homologues with myotoxic, cytolytic, and edema-inducing activities. PMID- 10519652 TI - Studies on the specificity of CNF, a phospholipase A2 inhibitor isolated from the blood plasma of the South American rattlesnake (Crotalus durissus terrificus). I. Interaction with PLA2 from Lachesis muta muta snake venom. AB - A phospholipase A2 inhibitor has been previously purified and cloned from the blood plasma of the South American rattlesnake, Crotalus durissus terrificus. This inhibitor, named CNF for Crotalus neutralizing factor, interacts with crotoxin, the main neurotoxin from C. d. terrificus venom, abolishing its phospholipase A2 activity. Crotoxin is a heterodimer of an acidic subunit (CA) and a basic phospholipase A2 (CB). CNF acts by forming a stable non-toxic complex with CB, replacing CA in the toxic CA-CB of crotoxin. In the present investigation, we have shown that CNF has a broader specificity. It is able to inhibit the PLA2 activity of the whole venom from the bushmaster snake (Lachesis muta muta), a species evolutionary related to Crotalus. Inhibition experiments have been carried out with four PLA2 active components isolated from L. m. muta venom, one basic and three acidic ones. CNF inhibition is not restricted to the basic PLA2, but extended to the three acidic forms as well. PMID- 10519653 TI - Blomhotin: a novel peptide with smooth muscle contractile activity identified in the venom of Agkistrodon halys blomhoffii. AB - A novel peptide has been isolated from the venom of Agkistrodon halys blomhoffii using a bioassay that monitors the stimulant effect on rat stomach fundus. The 11 amino acid peptide, named blomhotin, was purified to homogeneity by gel filtration column chromatography and reverse-phase HPLC. The amino acid sequence of blomhotin was determined to be pGlu-Gly-Arg-Pro-Pro-Gly-Pro-Pro-Ile-Pro-Arg, which is similar to that of bradykinin-potentiating peptides which themselves cause no contraction of smooth muscle. The contraction induced by blomhotin showed homologous desensitization, implicating the involvement of a blomhotin specific site in the response. PMID- 10519654 TI - The effects of antivenom on the in vitro neurotoxicity of venoms from the taipans Oxyuranus scutellatus, Oxyuranus microlepidotus and Oxyuranus scutellatus canni. AB - The venoms of the inland (Oxyuranus microlepidotus), coastal (O. scutellatus) and Papuan (O. s. canni) taipans are among the most potent in the world. The present study compared the in vitro neurotoxic effects of these venoms and the protective effects of taipan antivenom. Venom (10 microg/ml) from all three snakes abolished nerve-mediated twitches of the chick biventer cervicis muscle preparation with the following rank order of potency (based on the time taken to inhibit 90% of the twitch response; t90): O. microlepidotus (27+/-3 min) > O. scutellatus (42+/ 3 min) = O. S. canni (48+/-5 min). This inhibitory effect of all three venoms was primarily postsynaptic in origin as evidenced by the inhibition of responses to exogenous acetylcholine (ACh; 1 mM) and carbachol (CCh; 20 microM), but not potassium chloride (40 mM). In contrast, the presynaptic neurotoxins taipoxin (3 microg/ml) and paradoxin (3 microg/ml) abolished nerve-mediated twitches without producing a significant effect on contractile responses to exogenous agonists. Prior incubation of the tissue with taipan antivenom (1 unit/ml for 10 min) markedly attenuated the inhibitory effects of taipoxin (3 microg/ml) and paradoxin (3 microg/ml), as well as O. scutellatus (10 microg/ml) and O. s. canni (10 microg/ml) venom. However, in the presence of antivenom, O. microlepidotus venom (10 microg/ml) still abolished nerve-mediated twitches and responses to ACh and CCh. The results of the current study indicate that taipan antivenom, raised against O. scutellatus venom, is effective, in vitro, against the neurotoxic effects of venom from the Papuan and coastal taipans, as well as the presynaptic effects of venom from the inland taipan. However, the antivenom appears less effective against the postsynaptic effects of the latter. It is possible that inland taipan venom contains a component not neutralised by the antivenom which may contribute to the extreme potency of this venom. PMID- 10519655 TI - Purification and partial characterization of the phospholipase A2 and co-lytic factor from sea anemone (Aiptasia pallida) nematocyst venom. AB - Functional nematocysts of one specific morphological class, the penetrant microbasic mastigophores, were isolated from the sea anemone, Aiptasia pallida. These nematocysts contain a multicomponent venom composed of several proteins, including those with neurotoxic, hemolytic, and lethal activities. Hemolytic activity is produced by at least three synergistic venom proteins. One of these proteins is identified as a phospholipase A2 (EC 3.1.1.4) which exists in two isozymic forms, alpha and beta, with molecular weights of 45,000 and 43,000, respectively. The beta isozyme has been purified to homogeneity. It is a single chained glycoprotein with an isoelectric point (pI) of 8.8 and represents 70% of the phospholipase activity of the venom. The activity of the beta isozyme is relatively labile and is inactivated by 3.5 M urea or by heating at 45 degrees C. It is most stable at pH 4.0 and loses 50% of its activity at pH values below 3.5 and above 8.0. A second venom protein has also been purified. It is essential for the hemolytic activity of the venom and is termed co-lytic factor (CLF). It is a monomeric glycoprotein having a pI of 4.5. CLF has a molecular weight of approximately 98,000, a sedimentation coefficient of 4.8 S, and is prolate in shape, having a frictional ratio of about 1.6. CLF constitutes about 1.25% of the total venom protein and is assayed by reversing fatty acid inhibition of the venom hemolysis activity. PMID- 10519656 TI - Hypertension and abdominal pain: uncommon presentation after exposure to a pine caterpillar. AB - The pine caterpillar Thaumatopoea wilkinsoni is found in pine woods all over Israel. Contact with its hair usually results in local reactions. Systemic reactions after contact with caterpillar hairs are known in other caterpillar species, but have been described only once after contact with T. wilkinsoni. We describe a group of adolescents who were exposed to T. wilkinsoni while camping in a pine wood. Three of them were referred to an emergency department. They had severe pruritus, pain and edema at the contact sites, with papular and urticarial rashes. Two of the patients had abdominal pain and one patient had hypertension for several hours. The hypertension resolved spontaneously. CONCLUSION: Skin eruptions are the most common manifestations of T. wilkinsoni contact, however, although systemic manifestations are rare, abdominal pain and hypertension may occur. PMID- 10519657 TI - A neurophysiological method of rapid detection and analysis of marine algal toxins. AB - We have examined the effectiveness of the in vitro rat hippocampal slice preparation as a means of rapidly and specifically detecting the marine algal toxins saxitoxin, brevetoxin, and domoic acid and have identified toxin-specific electrophysiological signatures for each. Brevetoxin (PbTX3, 50-200 nM) produced a significant reduction in orthodromic population spike amplitude which was quick to reverse during a 50 min wash-out, while antidromic population spikes and field EPSPs exhibited only slight reductions, and fibre spiof orthodrokes showed no change at all. Domoic acid (100 nM) produced a robust, reversible increase in amplitude mic spikes, and the appearance of multiple spikes (i.e., epileptiform activity) within minutes of toxin wash-in. Other notable features of the domoic acid signature included a significant decrease in amplitude of the field EPSPs, and a complete absence of effect on either antidromic or fibre spikes. Fifty nanomolar saxitoxin (PSP) abolished all responses in all slices. Only antidromic spikes showed any recovery during wash-out. Field EPSP and fiber spike analysis further demonstrated that the preparation is capable of reliably detecting saxitoxin in a linearly responsive fashion at toxin concentrations of 25-200 nM, and tests of naturally contaminated shellfish confirmed the utility of this assay as a screening method for PSP. Our findings suggest that the in vitro hippocampal slice preparation has potential in the detection and analysis of three marine algal toxins important to the shellfish industry. PMID- 10519658 TI - Ciguatoxin reduces larval survivability in finfish. AB - Ciguatoxins are lipophilic polyether toxins which concentrate in the viscera and flesh of coral reef associated finfish (Hessel et al., 1960). In this study, we quantify the adverse effects of ciguatoxin on fish embryos by microinjection into the egg yolk of medaka (Oryzias latipis) embryos. Embryos microinjected with 0.1 0.9 pg/egg (ppb) of ciguatoxin exhibit cardiovascular, muscular, and skeletal abnormalities and those injected with higher levels (1.0-9.0 pg/egg) exhibit significantly reduced hatching success. The sensitivity of embryonic fish to direct oocyte exposure indicates that maternal transfer of low levels of ciguatoxin may represent an unrecognized threat to the reproductive success of reef fish and a previously undetected ecological consequence of proliferation of ciguatoxin-producing algae in reef systems increasingly impacted by human perturbations. PMID- 10519659 TI - Subcellular distribution of tetrodotoxin in puffer fish liver. AB - The liver homogenate of puffer fish was fractionated into blood cell, nuclear, mitochondrial, microsomal and cytosol fractions by the differential centrifugation method. All the five fractions were toxic to mice, although the toxin amount was significantly high in the cytosol fraction. Analyses by HPLC and LC-FABMS demonstrated that tetrodotoxin is the major toxic principle in each fraction. These results reveal that tetrodotoxin is widely distributed in organelles in liver cells, though predominantly in the cytosol fraction. PMID- 10519660 TI - Latrotoxins and secretory systems. Meeting held in Gaeta, Italy 18-21 October 1998. PMID- 10519661 TI - Bibliography of toxinology. PMID- 10519662 TI - Visceral and renal perfusion during thoracoabdominal aneurysm repair. PMID- 10519664 TI - Surgical venous thrombectomy. AB - Rehabilitation of the technique of venous thrombectomy is justified, but, in order for this technique to be effective, it must only be performed in selected cases. In the authors' view it is of the utmost value in young patients when the venous thrombosis occurs accidentally, after traumatism or surgery and when a diagnostic is made without delay. PMID- 10519663 TI - Thoracoabdominal aneurysm repair. AB - Aneurysms that extend from the descending thoracic aorta into the abdomen and also those that involve the visceral segments of the upper abdominal aorta are traditionally classified as thoracoabdominal. Besides the surgical exposure difficulties associated with repair of these aneurysms, the temporary interruption of renal, splanchnic, and perhaps even spinal cord flow introduces a number of potential complications that makes surgical repair of these aneurysms a daunting task. The exact incidence of thoracoabdominal aneurysms is unknown, but population studies suggest a prevalence at least a log fold less than the more common infrarenal abdominal aortic aneurysm. The poor prognosis of nonsurgically treated aneurysmal disease of the descending thoracic and abdominal aortas has been reasonably well established. Few patients with thoracoabdominal aneurysms survive beyond five years as a result of not only aneurysm rupture but also death from advanced co-morbid medical disease. Since the first successful report of thoracoabdominal aneurysm repair over 40 years ago, a number of remarkable contributions have been made in the field. These advances have led to a significant decline in operative mortality as well as procedure related morbidity. Spinal cord ischemia remains a perplexing and devastating complication following thoracoabdominal aneurysm repair. A number of surgical adjuncts have been developed over the years to reduce the incidence of cord ischemic complications, yet a great deal of controversy still exists with regards to the optimal protective strategy. A description of the incidence, natural history, and classification of thoracoabdominal aneurysms, along with associated repair techniques centered on reducing spinal cord ischemic complications will form the basis for this review. PMID- 10519665 TI - Thoracoabdominal aneurysm--how I do it. PMID- 10519666 TI - Thoracoabdominal aortic aneurysm. How we do it. AB - A systematic approach to paraplegia risk in the surgical treatment of thoracoabdominal aortic aneurysms based on effective strategies identified from the experimental literature is discussed. With this approach, collateral blood flow, rather than direct intercostal reimplantation, moderate hypothermia and endorphin receptor, is emphasized blockade. The result has been a 10-fold reduction in paraplegia risk in elective patients and a 5-fold reduction in acute patients. This reduction in paralysis risk has resulted in improved short- and long-term survival. PMID- 10519667 TI - Thoracoabdominal aortic aneurysm repair: how I do it. AB - There remains no consensus on the operative management of Thoracoabdominal aortic aneurysm (TAA). Our approach emphasizes operative expediency and simplicity (without circulatory assist techniques), avoiding anticoagulation and systemic hypothermia. The technique involves a fundamental clamp/sew method with specific adjuncts directed against the principle complications: epidural cooling (introduced in 1993) for spinal cord protection, regional renal hypothermia, and in-line mesenteric shunting to minimize visceral ischemia. In a cohort of over 200 TAA patients (50% Types I & II) treated during the past decade perioperative mortality has been 8% and paraparesis/paraplegia occured in 7%. These figures are halved for patients treated in elective circumstances. PMID- 10519668 TI - How I do it: thoracoabdominal aortic aneurysm graft replacement. AB - A technique of aortic graft replacement in thoracoabdominal aortic aneurysm repair is described. The author's experience with the surgical adjuncts of cerebrospinal fluid drainage and distal aortic perfusion, and the evolution and rationale of thoracoabdominal aortic aneurysm classification are discussed. Interpretation of the large amount of data that the author has accumulated would have been impossible without such a system. PMID- 10519669 TI - Reoperative aortic surgery. AB - The presentation of long-term complications after conventional aortic surgery and the treatment of patients that have had reoperative aortic operations are reviewed. Ninety-seven consecutive patients that had 102 subsequent aortic operations at a tertiary referral center were studied. Presenting symptoms, demographics, risk factors, indications for initial and second procedures, operative techniques and outcomes were recorded in a computerized database. There were 70 men and 27 women studied, with an average age of 64 years. First operations were performed primarily for aneurysm (56%) and occlusive disease (44%). The interval between procedures ranged up to 23 years, with a mean of 6 years. Indications for reoperation were subsequent aneurysm (65), graft occlusions (25) and/or infections (24). Seventy-three percent of the subsequent aneurysms were true metachronous aneurysms; the others were associated with the graft or an anastomosis. Para-anastomotic aneurysms may be more common with a primary end-to-side graft configuration. One-third of subsequent aneurysms were not palpable and asymptomatic. Graft occlusion can be treated safely with elective repeat bypass (mortality 0%). Graft infections that require total graft removal remain a challenging problem (mortality 17%). Although surgical approach for reoperations utilized more extensive exposure and proximal clamping, 59 elective aneurysm cases had a 5.1% mortality rate; eight emergent procedures for ruptured aneurysms resulted in 88% mortality. Reoperation for graft occlusion or infection showed a similar high mortality rate with emergent cases. In this referral practice, graft occlusion and infection are relatively less frequent, whereas metachronous aneurysm formation is now the most common indication for reoperation. These aneurysms often remain undetected until symptoms occur; frank rupture is usually lethal. As elective repair with modern reoperative techniques can be safely performed, routine computed tomographic examination is advisable at least every 5 years after aortic operations. PMID- 10519670 TI - The New England Society for Vascular Surgery: on the first quarter century. PMID- 10519671 TI - Takayasu's arteritis with renovascular hypertension: results of surgical treatment. AB - Renal artery stenosis caused by Takayasu's arteritis is an important cause of hypertension in young patients in the Far East. The role of surgery in Takayasu's arteritis is not as well-defined as in atherosclerosis or fibromuscular dysplasia. In this retrospective review, the author reports the results of 19 renal artery reconstructions in 12 young patients (median age 23.5 years, range 10-46 years) presenting with renovascular hypertension and Takayasu's arteritis, and discusses the different surgical options. The procedures performed included aortorenal bypass using vein (five), aortorenal bypass using polytetrafluoroethylene (PTFE) (eight), iliorenal bypass using vein (four), reimplantation of renal artery (one) and aortic replacement graft-renal bypass (one). Postoperatively, all 12 patients had a successful outcome with improved hypertension. There was no perioperative mortality, and complications included two early graft thrombosis and one late graft occlusion. These results support the view that surgical treatment for renovascular hypertension in Takaysu's disease is safe and effective. PMID- 10519672 TI - Retinoic acid suppresses intimal hyperplasia and prevents vessel remodeling following arterial injury. AB - Vitamin A and its derivatives (retinoids) are capable of inhibiting vascular smooth muscle cell proliferation in vitro. The present study examines the effect of two retinoids, all-trans retinoic acid and 13-cis retinoic acid, on intimal hyperplasia following arterial injury. After receiving varying doses of all-trans retinoic acid or 13-cis retinoic acid, 78 male Sprague-Dawley rats underwent standard balloon catheter denudation of the left common carotid artery. Morphometric analysis and immunohistochemistry for proliferating cell nuclear antigen was performed at early and late time points. Intimal/medial ratios were reduced in a dose-dependent fashion for animals treated with all-trans retinoic acid (P = 0.001) and 13-cis retinoic acid (P = 0.004). Proliferating cell nuclear antigen labeling indices were reduced after treatment with all-trans retinoic acid and 13-cis retinoic acid at early time points post-injury. At a dose of 10 mg/kg, both all-trans retinoic acid and 13-cis retinoic acid inhibited vessel remodeling as measured by increases in luminal diameter (P < 0.05) and external elastic lamina (P < 0.05). Retinoids are an attractive clinical option for the treatment of restenosis following angioplasty and arterial surgery. PMID- 10519673 TI - Cardiac assessment with thallium scanning prior to aortic aneurysm repair. AB - Coronary artery disease occurs commonly in patients with aortic aneurysms and is a major cause of morbidity and mortality. The role of screening and intervention for cardiac disease prior to aneurysm repair is controversial. The outcome after cardiac screening with thallium scanning and/or angiography in 102 consecutive patients undergoing aortic aneurysm repair was documented. Significant coronary artery disease was found in 34 (33%) patients and two patients had either coronary artery bypass or angioplasty prior to aneurysm repair. There was no cardiac mortality after aneurysm repair and the overall mortality on an intention to-treat basis was 2%. There was good correlation between prior history of cardiac events, electrocardiography (ECG) and the results of screening with thallium scanning and angiography. There was no correlation between cardiac history, ECG and the incidence of cardiac events in the postoperative period. Significant coronary artery disease was found in 33% of patients without a cardiac history or abnormal ECG. Cardiac screening with thallium scanning confirmed a high incidence of significant coronary disease in patients with aortic aneurysm. In this study, cardiac intervention followed by expedient aneurysm repair in 20 patients was associated with zero mortality. The short-term benefit of such a policy is difficult to prove and its main advantage may be better long-term survival. PMID- 10519674 TI - Clinical, biochemical and histochemical assessment of pretreatment with glucose insulin-potassium for patients undergoing mitral valve replacement in the third and fourth functional groups of the New York Heart Association. AB - In this study, the potentially beneficial effects of preoperative treatment with glucose, insulin and potassium in a randomized series of 30 consecutive patients undergoing mitral valve replacement, who were in the third and fourth functional groups of the New York Heart Association scale, were investigated. Fifteen patients received glucose, insulin and potassium, and 15 patients received the same volume of normal saline. The characteristics of the groups did not differ. Papillary muscle-biopsy samples were obtained at the time of surgery and analysed for glycogen, both biochemically and histochemically. The clinical course of all patients was monitored closely during the first 24 hours after surgery. The patients receiving glucose, insulin and potassium had higher glycogen levels (43 +/- 13.54 micromol/g) (P < 0.001). In addition, they required less inotropic pharmacological support (scored by the Gradinac method), had fewer ventricular arrhythmias and exhibited improved haemodynamic indices: cardiac output increased (P < 0.025 to P < 0.005), while systemic vascular resistance decreased (P < 0.001). Pretreatment with glucose, insulin and potassium did not, however, affect the patients' postoperative wedge pressure and mortality. The results of this study suggest that glucose, insulin and potassium pretreatment may be beneficial in unfit patients undergoing mitral valve replacement. PMID- 10519675 TI - Management of late complications after classic Fontan procedure by conversion to total cavopulmonary connection. AB - Classic Fontan procedures, such as atriopulmonary connection or atrioventricular connection, are associated with right atrial dilatation and several late complications. Seven patients with univentricular heart who presented with exercise intolerance (n = 7), severely dilated right atrium (n = 7) with pulmonary vein compression (n = 3), atrial arrhythmias resistant to medical treatment (n = 4), cyanosis (n = 4), diffuse effusions and oedema (n = 1), and protein-losing enteropathy (n = 1), underwent conversion to total cavopulmonary connection 5.8-14.4 years after a previous atriopulmonary connection (n = 6) or atrioventricular connection (n = 1). A 14-year-old boy who, preoperatively, was in ventricular failure and a very poor state died early after conversion because of low cardiac output. All survivors had either marked or partial clinical improvement with regression of cardiomegaly, absence of pulmonary vein compression or cyanosis, and recovery of sinus rhythm. Conversion to total cavopulmonary connection appears to be effective in the treatment of late complications after classic Fontan procedures. It should be considered early in symptomatic patients, before significant ventricular dysfunction and clinical deterioration ensue. PMID- 10519676 TI - A right atrial approach in redo postinfarction ventricular septal defect. AB - Despite improvements in surgical techniques, post-infarction ventricular septal defect remains a surgical challenge that is associated with significant early and late mortality. Furthermore, the recurrence of the defect after primary correction occurs in approximately 10-25% of patients, and the operative risk increases because of a difficult dissection that is often complicated by previous patent grafts. The repair of recurrent ventricular septal defect has generally been performed by ventriculotomy in the infarcted zone. The authors propose an alternative approach that, when the rupture is posterior, allows its complete visualization, and avoids any further ventriculotomy in an already impaired ventricle. PMID- 10519677 TI - Unusual cause of a stroke in a patient with seronegative rheumatoid arthritis. AB - The case of a patient who suffered a severe cerebrovascular accident caused by an aortic rheumatoid granuloma is presented. PMID- 10519678 TI - Successful one-stage resection of intravenous leiomyomatosis of the uterus with extension into the heart. AB - Intravenous leiomyomatosis of the uterus is a rare neoplasm characterized by nodular masses of benign smooth muscles with intraluminal growth to the inferior vena cava and, in some cases, to the heart. It may cause abdominal and cardiovascular symptoms and is a serious risk of death when it reaches the tricuspid valve. Surgery is the best treatment and must be applied as soon as possible using cardiopulmonary bypass. The authors report a new case that had cardiac involvement and was successfully resected. The symptoms, imaging diagnosis, pathological and histopathological findings, tumors that mismatched the intravenous leiomyomatosis (IVL), and the use of circulatory arrest and deep hypothermia are discussed. A review of the literature is included. PMID- 10519679 TI - Discharge planning and transitional care: issues in Thai nursing. AB - Discharge planning is part of the care process that places nurses in a pivotal position in facilitating continuity of care for clients. This ethnographic study explored the current discharge practices of Thai nurses and examined how transitions from hospital to home were incorporated into Thai nursing practice. The study was conducted with registered nurses in an acute hospital in central Thailand. Results indicated that the discharge process in this community was highly informal with several factors affecting the effectiveness of nurses' discharge functions. Consequently, strategies to improve this care process were proposed for further implementation. PMID- 10519680 TI - Hermeneutic analysis: a qualitative decision trail. AB - Qualitative analysis is problematic from two perspectives, which exist in the science and art debate [Tesch, R., 1990. Qualitative Research. Analysis types and software tools. The Falmer Press, London; Robson, C., 1993. Real World Research. Blackwell, Oxford]. The science view claims that the absence of clear and agreed analysis processes, which can be found in the quantitative domain, attracts labels to qualitative analysis of intuitive artistry and personal journeys which are considered 'unscientific'. This thinking remains dominant despite the growth of systematic qualitative analysis supported through computer analysis systems [Tesch, 1990]. However, in the art domain there is real resistance to the development of set methods of analysis which does view qualitative analysis, "as more of an art than a science" [Robson, 1993, p. 370]. This paper offers a contribution to the qualitative analysis tension through the promotion and illustration of a decision making trail. This option supports the principles of academic rigour in qualitative research [Guba, E.G., Lincoln, Y.S., 1981. Effective Evaluation. Jossey Bass, San Francisco] as a decision trail permits the research community to make their own judgements concerning the process of analysis, the overall trustworthiness of the research and therefore its presented interpretations. PMID- 10519681 TI - Working across cultures: a model for practice in developing countries. AB - The aim of this study was to identify the influence that cultural values have on the practice of nurses working in developing countries in primary health care. This was achieved through the exploration of their common experiences of practice using an ethnographic approach with ethnosemantic analysis as the research tool. Western expatriate nurses represent a dominant Western culture and so carry with them the values and beliefs of the West into their practice, thus potentially creating greater dependence rather than supporting the primary health care principles exhorted through Alma Ata of self reliance and community participation. The study showed that the effect is determined by the values that influence practice. These values are either modified by the expatriate nurses to create an enabling environment or there is a failure to modify their values so promoting dependence through a 'helping' or 'aid giving' approach. PMID- 10519682 TI - The art of nursing. AB - It is a common assumption that nursing is an art. Art in this sense is sometimes taken to mean a fine art, and sometimes a skill or craft. This paper reviews some of the arguments for nursing as an art and concludes that nursing is not a fine art, but that it is an art in the more general sense of a craft or skill performed by people. It is further argued that, in most instances, to say that nursing is an art tells us little or nothing that is useful and the word 'art' is redundant in this context. PMID- 10519683 TI - Caring attributes, professional self concept and technological influences in a sample of Registered Nurses in eleven countries. AB - Caring, the theoretical foundation of nursing, is identified as one of the core values by Hospital Authorities world-wide to be actualised in clinical practice. Exactly how caring attributes relate to nurses' professional self image and quality nursing service in the context of a contemporary technological environment have yet to be operationalised. In total, 1957 Registered Nurses from 11 different countries were involved in the study which aimed to: develop an understanding of and compare the responses to items relating to caring, professional self concept and technological influences. To collect data an instrument containing 104 Likert items was administered to RNs working in general hospitals. The instrument contained sections which examined professional self concept, technological influences and caring attributes. Descriptive and inferential statistics revealed that many of the Asian nurses in the sample were younger, had less experience yet were more qualified than their 'western' colleagues. The mean scores for the caring attributes for nurses from the Philippines, Sweden and South Africa were significantly higher than those from China (Beijing), Korea, China (Hong Kong) and Scotland. The Korean sample demonstrated the lowest mean score for professional self concept while the New Zealand sample the highest. The Australian, Canadian, NZ, Scotland and Swedish samples were strongly of the opinion that the increase in technology has not brought about the any more spare time in nursing and generally had a more negative opinion about the influence of technology when compared to those from China (Beijing), Philippines, China (Hong Kong) and Singapore. The results are discussed in the light of the cultural differences in the sample and recommendations for future research are considered. PMID- 10519684 TI - The science of meaning in chronic illness. AB - A generation of 'insider research' into the subjective experience of being ill and the social context in which illness experience is played out provides us with a developing theoretical knowledge of what has come to be understood as the 'meaning' of illness. In this paper, a critical analysis of the tradition of meaning research forms the basis for an interpretive synthesis of what we now know and where future research might most appropriately be directed. Problems arising from some of the traditional approaches to knowledge development are articulated and potential solutions arising from more recent methodological advances are proposed. The relevance of this analysis for nursing practice knowledge and for the strategies for nursing's scientific development are considered. PMID- 10519685 TI - Is physical activity as effective in reducing risk of cardiovascular disease as estrogen replacement therapy in postmenopausal women? AB - Coronary heart disease (CHD) is known as a disease of postmenopausal women. While hormone replacement therapy is recommended increasingly to postmenopausal women for the prevention of CHD, the potential impact of non-pharmacologic interventions on cardiovascular disease reduction has been largely unexplored. The purpose of this article is to develop the understanding of the research literature on three risk factors by which estrogen and physical activity, a non pharmacologic intervention, may confer cardiovascular protection in postmenopausal women. Three risk factors are lipoproteins, systolic pressure, and changes in the vascular endothelial functions. Effects of estrogen and physical activity on these biological factors are compared. PMID- 10519686 TI - Problem drinkers in acute care settings: validation of an assessment instrument. AB - This article reports a validation study of a screening procedure for detecting those patients whose patterns of alcohol consumption places them at risk of developing alcohol-related health problems. The sample comprised 998 patients who were admitted to the wards of a general hospital for treatment of conditions which were not primarily alcohol-related. Twenty-six per cent of patients reported drinking more than published guidelines for low-risk drinking. The use of a diary was found to be a valid means of assessing patients' levels of alcohol consumption, and its reliability and validity was confirmed as a method of distinguishing potential problem drinkers from those whose level of alcohol consumption does not place them at risk. As such, the diary is recommended as an assessment tool which can readily be incorporated into standard assessment procedures. PMID- 10519687 TI - A study of minimal interventions for problem drinkers in acute care settings. AB - This article reports an investigation of three minimal interventions for potential problem drinkers in general hospital wards. The interventions were: (a) brief advice; (b) the provision of health education literature; (c) a combination of both the advice and literature. One year after recruitment the mean levels of alcohol consumption and the number of alcohol-related problems reported by the cohort was significantly reduced. These reductions were supported by reductions in the mean levels of GGT and AST, but not in mean MCV. No statistically significant treatment effects were found. The results are presented and implications for nursing are discussed. PMID- 10519688 TI - Hyperthermia and liposomes. AB - Hyperthermia and liposomal drug delivery are treatment modalities that have been used to treat cancer over the last two decades. More recently, the two therapies have been used together in an attempt to exploit their mutual interactions against cancer. The goal of this review is to explore the literature related to combined hyperthermia and liposomal drug delivery for cancer therapy. The motivation behind combining hyperthermia and liposomal drug delivery is discussed from a physical chemical and physiological standpoint. Two types of therapeutic ratios were calculated for in vivo studies from across the literature. These ratios compared the results obtained from hyperthermia and liposomes to hyperthermia and free drug as well as to liposomes without hyperthermia. These two therapeutic ratios were applied to both tumour drug uptake and tumour growth delay studies. In all studies reviewed, hyperthermia in combination with liposomal drug showed an enhanced therapeutic effect compared to either treatment modality alone or hyperthermia and free drug. Future work needs to be focused on optimizing thermosensitive liposomes and understanding the effect of thermal dose on liposomal drug delivery. Though not currently used in the clinic, this combination therapy seems to hold great promise towards improving current cancer therapeutic regimens. PMID- 10519689 TI - Radiation therapy, cisplatin and hyperthermia in combination in management of patients with recurrent carcinomas of the head and neck with metastatic cervical lymph nodes. AB - Twenty-one patients with recurrent carcinomas of the head and neck with metastatic cervical lymph nodes were treated with radiation therapy, cisplatin and hyperthermia in combination, in an attempt to investigate any potential contribution in terms of safety, response, duration of palliation and quality of life. Patients not initially treated with radiation therapy were treated with a median dose of 70 Gy and patients initially treated with radiation therapy with a median dose of 30Gy. The median number of weekly cisplatin courses was five and the median number of twice weekly local external ultrasound hyperthermia sessions was five. Average T90, Average T50 and Average T10 were 39.9 +/- 1.2 degrees C, 42.4 +/- 1.3 degrees C and 44.5 +/- 0.8 degrees C, respectively, and Average CEM 43 degrees C T90, Average CEM 43 degrees C T50 and Average CEM 43 degrees C T10 were 7.8+/-9.6min, 22.6 +/- 18.8min and 39.3 +/- 25.1min, respectively. Mean follow-up was 1 year. Nodal complete response was achieved in eight patients and palliation of presenting symptoms in 19. Overall survival was 39% at 1 year. Grade 3 acute skin toxicity was observed in one patient and Grade 3 acute haematological toxicity in one. Radiation therapy, cisplatin and hyperthermia in combination appear to be safe and might improve response, prolong duration of palliation and reinstate quality of life in patients with recurrent carcinomas of the head and neck with metastatic cervical lymph nodes. PMID- 10519690 TI - Use of whole body hyperthermia as a method to heat inaccessible tumours uniformly: a phase III trial in canine brain masses. AB - In this study, whole body hyperthermia (WBH) was assessed as a means of heating intracranial tumours uniformly. Twenty-five dogs received radiation therapy and 20 the combination of radiation and WBH. Total radiation dose was randomly assigned and was either 44, 48, 52, 56 or 60 Gy. Because of WBH toxicity, intercurrent disease or tumour progression, seven of the 45 dogs received less than the prescribed radiation dose. For WBH, the target rectal temperature was 42 degrees C for 2h and three treatments were planned. In five of the 20 dogs randomized to receive WBH, only one WBH treatment was given because of toxicity. WBH toxicity was severe in six dogs, and resulted in death or interruption in treatment. Most tumours did not undergo a complete response, making it impossible to differentiate tumour recurrence from brain necrosis as a cause of progressive neuropathy. Therefore, survival was the major study endpoint. There was no survival difference between groups. One-year survival probability (95% CI) for dogs receiving radiation therapy alone was 0.44 (0.25, 0.63) versus 0.40 (0.19, 0.63) for dogs receiving radiation and WBH. There was no difference in the incidence of brain necrosis in the two treatment groups. Results suggest that use of WBH alone to increase the temperature of intracranial tumours as a means to improve radiation therapy outcome is not a successful strategy. PMID- 10519691 TI - Thermometric analysis of intra-cavitary hyperthermia for esophageal cancer. AB - Thermometric analysis was carried out in 51 patients with esophageal cancer treated with intra-cavitary hyperthermia combined with radio chemotherapy, to test whether temperature index (T20, T50) and T90) could be used as an indicator for tumour control. Hyperthermia was administered by intra-cavitary microwave applicator. The T20, T50 and T90 were deducted from the temperature sensors T0 and T3 situated at the center of the tumour surface and 3cm from it. Eighteen patients with local control > or =36 months were named long term control patients (LC), 24 patients with local recurrence within 24 months (LR) (there were no events occurring between 24 and 36 months) and nine patients died of metastasis without local recurrence (DM). The overall survival rates were 80.4 +/- 5.6% at 1 year, 38.3 +/- 6.9% at 3 years and 31 +/- 6.7% at 5 years, respectively. Chi square test showed no influence of the number of hyperthermia sessions on the local control (p > 0.25). The 5-year local control rate was 18.8% for the patients with T90 < 43 degrees C and 45% for those with T90 > or = 43 degrees C (p < 0.01). The average T90 was 43.76 +/- 0.74 degrees C for the LC patients and 43.17 +/- 0.57 degrees C for those LR (p = 0.024). The mean T90 was higher than 43 degrees C in 94.4% of LC, whereas in 58.8% of LR. The study suggested that T90 was a good parameter for thermal dose in the intracavitary hyperthermia for the treatment of esophageal cancer. PMID- 10519692 TI - Behaviour of heat-treated bone marrow cells in long-term bone marrow cultures. AB - The effect of prior heat treatment on the ability of bone marrow cells to form long-term bone marrow culture has been studied. Bone marrow cells were heated for various times in the temperature range of 39-43 degrees C and then cultured in the modified Dexter type suspension culture. At weekly intervals, the behaviour of the cultures in terms of stroma formation and confluency, cellular viability, and myelopoiesis were evaluated. The results show that there was a dose-dependent decrease in the number of viable cells in the non-adherent fraction of the cultures. Cytological analysis of these cells showed a strong shift towards macrophage population in the successive weeks of the cultures and also as a function of heat dose delivered to these cultures. The stroma formation was delayed or inhibited as a function of the heat dose. The number of granulocyte macrophage colony forming cells (CFU-GM) in both adherent and non-adherent fraction of the cultures were decreased substantially after hyperthermia treatment. At 41 degrees C and higher temperatures, the CFU-GM were severely diminished in both fractions. The dose response experiments showed that the decrease in the number of CFU-GM was dependent on the heat dose. The results suggest that CFU-GM is an extremely sensitive target in the hyperthermia treatment of bone marrow cells and heat-treated bone marrow cells lose their ability to maintain long-term cultures. PMID- 10519693 TI - A multi-user networked database for analysis of clinical and temperature data from patients treated with simultaneous radiation and ultrasound hyperthermia. AB - A database was developed using commercially available development software that allows the entry of clinical data and automatically analyses temperature and power data from a commercial ultrasound hyperthermia system. The database can be accessed via network connections by more than one authorized user, thus facilitating the entry, management, and analysis of clinical data. The software automatically estimates ultrasound induced temperature artifacts and calculates thermal dose parameters such as T90s, equivalent minutes at 43 degrees, and time at or above index temperatures using the corrected temperatures. These parameters also become part of the database. Digital photographs of treatment setup, probe placement, and tumour or normal tissue response can be included in the database for documentation and reference. Ultrasound diagnostic images that document the depth and reproducibility of probe placement can be scanned into the PC and included in the database as well. This short communication documents experiences developing this tool that may be useful to other investigators. PMID- 10519694 TI - Localized current field hyperthermia in carcinoma of the cervix: 3-D computer simulation of SAR distribution. AB - Software was developed for the 3D simulation of SAR distribution for a 500 kHz localized current field hyperthermia system to be used in patients with carcinoma of the cervix. This hyperthermia system was specifically designed for use with a modified Fletcher-Suit intracavitary applicator. It consists of software modules for data input, tetrahedral grid generation and a numerical calculation of SAR distribution using an adaptive, multilevel finite element code. The AVS (Advanced Visual System, Inc.) system was used for the visual presentation of the results. A quasi-static approach was employed for the determination of SAR distribution. Results of the performed numerical tests were presented and they showed an important, clinically relevant ability to obtain a selective power deposition. This selective power deposition depended on the applicator geometry, i.e. the distance between the components of a Fletcher-Suit applicator and their relative position and the use of different modes of excitation. PMID- 10519695 TI - An electric field measurement system, using two-dimensional array of diodes. AB - In hyperthermia induced by electromagnetic applicators, one way to obtain information about the energy absorption is by measurement of electric field strength (E). This paper describes a system which can measure E-distributions, using a two-dimensional array of diodes. It was designed to be used on patient skin, during hyperthermia treatments of superficial tumours, providing additional data for applicator power control. The first prototype consists of a sheet with Schottky diode sensors with a spacing of 2.5cm, connected to high-resistance leads, printed with carbon ink. The rectified diode voltages are passed through a multiplexer unit to an AD-DA card in a PC. The sensitivity of the sensors is linearly proportional to the electric field and to the length of the extended diode connection pins. Relative E-field distributions obtained with 64 sensors are updated within 1s. Phantom measurements, performed with the sensor matrix under a Lucite Cone Applicator (LCA) and under a 2 x 2 array of Current Sheet Applicators (CSAs) were compared with infrared measurements of the temperature rise after a short power pulse. A fair agreement was found between the square of the diode voltages and the infrared distributions. Movement of a single CSA over the sensor matrix can be visualized clearly by the system. The diode matrix E field measurement system is sufficiently fast and accurate to give valuable feedback for power steering for an array of LCAs and CSAs. The system has the potential of being a helpful tool in other fields of quality assurance as well. PMID- 10519696 TI - Otogenic lateral sinus thrombosis in children. AB - INTRODUCTION: The clinical picture of lateral sinus thrombosis (LST) has changed with the advent of antibiotics, as have the utility of various diagnostic tests. LST may appear in children as a complication of acute otitis media, but nowadays it is more frequently encountered in adults with long-standing chronic ear disease. METHOD: A retrospective study of all the pediatric patients with LST between 1982 and 1997. RESULTS: Thirteen cases of LST were diagnosed and treated by our department. In six cases, LST was due to acute otitis media and in the remaining cases it was due to chronic otitis media. Headache, fever, aural discharge and mastoid tenderness were the most frequent findings in these patients and four patients were initially diagnosed with meningitis. In the majority of the patients, LST was accompanied with other intracranial complications, such as perisinus abscess, brain abscess and meningitis. One patient with multiple brain abscesses, unresponsive to several drainage procedures, died. The other patients recovered and have since been followed-up as out-patients. CONCLUSION: LST may be difficult to diagnose due to previous antibiotic treatment and to the overlap of clinical findings with other entities such as meningitis. Despite the value of modern imaging techniques in the investigation of the disease, a high index of suspicion based on the clinical picture is warranted. Our results are consistent with those of other recent studies, who found that mortality of LST has dropped below 10%. PMID- 10519697 TI - Von Willebrand disease as cause of unanticipated bleeding following adeno tonsillectomy. AB - Von Willebrand disease (vWD) is a frequent autosomal bleeding disorder. We report two unsuspected patients over 6 years of age with this disease operated by tonsillectomy and adenoidectomy (T&A) at our Department. Preoperative hematologic work-up (with activated partial thromboplastic time, prothrombin time, fibrinogen, and platelet count) was normal. Both patients had undergone previous adenoidectomy without complications. In both cases, profuse bleeding was noted in the immediate postoperative period. The therapeutic measures in these situations are discussed and a review of the current literature concerning preoperative hematologic evaluation of T&A is included. PMID- 10519698 TI - Complicated acute sinusitis and the computed tomography anatomy of the ostiomeatal unit in childhood. AB - The ostiomeatal unit is postulated to be a critical area in the pathogenesis of sinus disease and accurate assessment of this anatomical area has made possible by the coronal computed tomography (CT) scan. Data from the CT scans of 24 patients with complications of acute sinusitis were retrospectively reviewed and compared with a set of normative data of the infundibular length, and width and the uncinate angle in 196 scans of healthy children. In the patients with complicated sinusitis the infundibular length was found to be less and the infundibular width greater than the normative data. No difference in the mean uncinate angle of the two groups was shown. One, therefore, has to assume that the pathological process at the ostiomeatal unit is more likely to be mucosal than bony. PMID- 10519699 TI - Magnetic resonance imaging procedures to study the concurrent anatomic development of vocal tract structures: preliminary results. AB - The vocal tract structures undergo drastic anatomic restructuring during the course of development from infancy to adulthood. This study demonstrates the feasibility of using MRI to examine the growth processes of the vocal tract. This method affords precise and detailed visualization of the soft tissues in the oro pharyngeal region, while also providing images of related bony and cartilaginous structures. Information on anatomic restructuring contributes to the understanding of how speech emerges and develops, and it also establishes normative information that can be used in the assessment of developmental anomalies. This paper describes the method used to measure and examine the concurrent anatomic development of the various vocal tract structures during early childhood. Preliminary results from two pediatric subjects indicate that there is synchrony of growth in the different structures-both soft and hard tissues-, and that such synchronous growth appears to persist during periods of growth spurts. PMID- 10519700 TI - Diagnostic value of tympanometry in infants in clinical practice. AB - One hundred and twenty-one visits of 58 infants (2-11 months of age) were evaluated in the Finnish Otitis Media Vaccine Trial. Infants were examined with tympanometry (Grason-Stadler GSI 38 Autotymp) and pneumatic otoscopy by one study doctor. Diagnosis of otitis media was verified by myringotomy in 74% of cases. Tympanometry was technically successful in 94% of ears. The success rate was statistically significantly higher (P < 0.05) among infants less than 7 months of age than those above 7 months. The sensitivity of tympanometry (type B) to detect ears with middle ear fluid was 0.70 and the specificity 0.98 with a positive predictive value of 0.93 and negative predictive value of 0.94. The sensitivity was somewhat lower in the younger age group (0.61); specificity and positive and negative predictive values were good in both age groups. The high success rate and high negative and positive predictive values of tympanometry make it a useful aid for assuring the correct diagnosis of otitis media in infants in routine clinical practice. PMID- 10519701 TI - Vestibular compensation in infants and children with congenital and acquired vestibular loss in both ears. AB - In children with semicircular canal anomalies, vestibular compensation during their development and growth was studied. The damped rotation test elicited 'absence or poor per-rotatory nystagmus and absence of post-rotatory nystagmus in all cases. Development of gross motor and balance function was seriously delayed in each case during the first 2 or 3 years of life. Thereafter, during the pre school age, all children could achieve most landmarks of motor development, such as head control, independent walking and running. However, balance functions at the age of entrance of the elementary school (6 years old) were variously impaired in each case. The better case could swim under water but the poor case could not maintain static balance with eyes closed. These motor skills due to vestibular compensation presumably depend on integration of the compensatory input from visual, somatosensory and proprioceptive senses, and the maturation of motor control systems in the cerebellum, basal ganglia and motor cortex. PMID- 10519702 TI - Laryngeal mucosal histology in laryngomalacia: the evidence for gastro oesophageal reflux laryngitis. AB - OBJECTIVE: To describe the histopathological changes of the mucosa in laryngomalacia; look for any relationship with gastro-oesophageal reflux and to describe the histological changes of reflux laryngitis in laryngomalacia. METHODS: We examined serial histological sections from nine cases of laryngomalacia, who had aryepiglottoplasty and compared the histopathological features with five cases of postintubation inflammatory laryngitis and five age matched autopsy specimens of normal larynx. RESULTS: Five of the cases of laryngomalacia had mild inflammation in the form of basal cell hyperplasia and chronic inflammation close to the basement membrane. Deeper subepithelium was oedematous. Two cases had moderate and two cases severe inflammation. The latter showed ulceration and a dense band of chronic inflammation in the immediate subepithelium with underlying oedema. Three of the cases had gastro-oesophageal reflux proven by barium swallow. Two of these showed intraepithelial eosinophils. CONCLUSIONS: A band of inflammation of variable intensity just beneath the epithelium with oedema deep to it is the most important histological feature of laryngomalacia. The presence of intraepithelial eosinophils appears to be a histological indication for reflux aetiology of the inflammation. PMID- 10519703 TI - Encephalocraniocutaneous lipomatosis with otolaryngologic manifestations: a rare neurocutaneous syndrome. AB - Encephalocraniocutaneous lipomatosis (ECCL) is a rare congenital disorder and was first described in 1970. The main clinical features of the syndrome include convulsions beginning in infancy, mental retardation, and unilateral cutaneous and ophthalmologic lesions with ipsilateral cerebral manifestations. A 14-year old caucasian boy with ECCL associated with otolaryngologic manifestations is reported. To our knowledge, this is the first case of ECCL with otolaryngologic manifestation in the English literature. PMID- 10519704 TI - Choanal and ileal atresia: a new syndrome or association? AB - Choanal atresia is a relatively common congenital malformation which is often associated with other anomalies. On the other hand, ileal atresia is very rare, mostly nonsyndromic and occasionally associated with other anomalies. The association of choanal and ileal atresia is unknown. Here we report the first instance of bilateral choanal atresia and ileal atresia in a full term male infant and describe the subsequent surgical treatment of both conditions. The association is unique and may represent a syndrome. PMID- 10519705 TI - Ectopic thymus presenting as a solid submandibular neck mass in an infant: case report and review of literature. AB - Solid ectopic cervical thymus is an extremely uncommon etiology of a neck mass in an infant. It occurs in the line of descent of the thymus from the angle of the mandible to the superior mediastinum. Nine cases of ectopic cervical thymus in infants have been reported in the literature. Only two of nine cases were solid, the remaining seven were thymic cysts. A preoperative diagnosis is seldom considered and is often misdiagnosed as a possible malignancy or a lymph node. We present a case of a 2-month-old infant with an asymptomatic enlarging right neck mass. Patient underwent complete excision of the mass. PMID- 10519706 TI - NHS direct: here and now. PMID- 10519707 TI - Growth hormone insensitivity: a widening diagnosis. PMID- 10519708 TI - Confirmation of deafness in infancy. AB - AIM: To assess delay in confirming hearing impairment in infants identified by universal neonatal screening and to investigate the causes. PATIENTS: Infants identified from 25 199 babies screened from January 1992 to December 1997. METHODS: A two stage transient evoked oto-acoustic emission test (TEOAE), with a threshold auditory brainstem response (ABR) recording undertaken on those who failed. The screen identified infants with a permanent congenital hearing impairment (PCHI) averaging 40 dBnHL or worse in the best ear. Those with less impairment were also ascertained. The positive predictive value (PPV) of the ABR test and measures of delay between identification and eventual diagnosis were analysed. RESULTS: A targeted PCHI was found in 1.18/1000 neonates. The PPV of the ABR for confirming a targeted PCHI was 100% when the ABR threshold was >/= 80 dBnHL. Nine of 11 infants with this threshold had severe or profound permanent deafness. The delay from ABR to audiological certainty was about 1 month diagnosis was confirmed around 3 months. There was uncertainty when the ABR was 40-80 dBnHL. The PPV was 60% and 8% when the ABR thresholds were 70 dBnHL and 50 dBnHL, respectively. 85 of 111 infants with ABR thresholds in this range had a temporary conductive impairment. Their early diagnosis depended upon the type and degree of hearing impairment and diagnosis was delayed to about 8 months in these infants. CONCLUSIONS: Hearing impairments identified by universal screening are delayed in all but those with severe or profound bilateral PCHI. This delay can be reduced by applying in early infancy a battery of audiological tests and requires further exploration. PMID- 10519709 TI - Cause of death in cerebral palsy: a descriptive study. AB - BACKGROUND: Cause specific research on death certification in chronic disease has rarely involved cerebral palsy. AIMS: To evaluate cause of death information in people known to have cerebral palsy by: describing the cause of death distribution; determining case ascertainment using death certification as the data source; and analysing the choice of wording and its arrangement in the "cause of death statement". STUDY CASES AND SETTING: People with early or late impairment cerebral palsy who died by 30 June 1998, on the population based Mersey Cerebral Palsy Register born 1966-91 to mothers resident locally. STUDY DESIGN: Descriptive study of the multiply coded cause of death statements from National Health Service Central Register flagging. RESULTS: Death certificate copies were acquired for all 282 (13.4%) of the 2102 registered cases who died. Cerebral palsy was the most common "underlying cause of death" (95 of 282; 33.7%) and was mentioned in a further 61 cases. The underlying cause of death was more likely to be cerebral palsy with increasingly severe disability and was derived from Part II in 16 of 95 cases. CONCLUSIONS: The potential of death certification for case ascertainment of cerebral palsy is important, but limited, even with multiple cause coding. Mortality data need careful interpretation as a proxy source for examining trends and patterns in cerebral palsy. PMID- 10519710 TI - Growth patterns of breast fed and formula fed infants in the first 12 months of life: an Italian study. AB - AIM: To compare the growth patterns of breast fed and formula fed Italian infants in the first 12 months of life using World Health Organisation (WHO) reference data. METHODS: The growth patterns of 73 breast fed infants (36 male, 37 female) and 65 formula fed infants (35 male, 30 female) were compared. Solid foods were introduced with the same weaning schedules from the 5th month in both groups. The weight for age (WA), length for age (LA), and weight for length (WL) z scores (National Center for Health Statistics-WHO data) were calculated at birth, 1, 2, 3, 4, 6, 9, and 12 months. RESULTS: Breast fed infants had the highest z scores (WA, WL) at birth. Breast fed groups had significantly higher growth indices at 1 month (WA, LA), 2 months (WA) and 3 months (WA, LA) of age. Compared to breast fed groups, formula fed infants showed significantly higher WA z score changes in the 1-2, 2-3, 3-4, and 4-6 month intervals. LA z score changes were higher for breast fed infants at 0-1 month and for the formula fed infants at 4-6 months. In the 6-12 month interval growth indices progressively increased for the formula fed infants and declined for infants breast fed for longer (12 months). The 0-12 month changes in WA, LA, and WL z scores were positive for formula fed infants and negative for the 12 month breast fed group. Nevertheless, the 12 month breast fed group showed an absolute WA z score just below 0 (mean (SEM) -0.04 (0.26)) at 12 months. CONCLUSION: The growth pattern of breast fed and formula fed Italian infants differs in the first 12 months of life. This questions the validity of current reference values for monitoring the growth of breast fed infants. Growth indices in breast fed groups, high at birth and closer than expected to the reference at 12 months, may reflect differences in genetic factors, intrauterine conditions, or both. PMID- 10519712 TI - Chronic pain from the appendix PMID- 10519711 TI - Smoke alarm installation and function in inner London council housing. AB - AIM: To determine the prevalence of and predictors for installed, functioning smoke alarms in council (public) housing in a low income, multi-ethnic urban area. DESIGN: Cross sectional study. SETTING: 40 materially deprived electoral wards in two inner London boroughs. PARTICIPANTS: Occupants of 315 addresses randomly selected from council housing lists, with 75% response rate. MAIN OUTCOME MEASURES: Installation and function of smoke alarms based on inspection and testing. RESULTS: 39% (95% confidence interval (CI) 33% to 46%) of council tenants owned a smoke alarm, 31% (95% CI 25% to 38%) had an installed alarm (of which 54% were correctly installed), and 16% (95% CI 12% to 22%) had at least one installed, functioning alarm. Alarms most commonly failed because they lacked batteries (72%). In multivariate modelling, having an installed, functioning alarm was most strongly associated with living in a house versus a flat (apartment) (odds ratio (OR) 3.2, 95% CI 1.1 to 10.0), having two resident adults versus one (OR 2.8, 95% CI 1.2 to 6.5), and recognising stills from a Home Office television smoke alarm campaign (OR 2.4, 95% CI 1.1 to 5.5). CONCLUSIONS: Fires are a leading cause of child injury and death, particularly among those younger than 5 years of age and those in social classes IV and V. Smoke alarms are associated with a significantly reduced risk of death in residential fires, and are more protective in households with young children. Few council properties in a multi-ethnic, materially deprived urban area had any installed, functioning smoke alarms, despite a high risk of residential fires and fire related injuries in such areas. Effective methods to increase the prevalence of installed and functioning alarms must be identified. PMID- 10519713 TI - Respiratory function in childhood following repair of oesophageal atresia and tracheoesophageal fistula. AB - AIM: To determine the relation between respiratory function in infancy and at school age in children who have undergone oesophageal atresia and tracheoesophageal fistula repair, and assess the value of infant respiratory function testing; and to examine the effect of bronchodilators. METHOD: Fourteen children (6 girls, and 8 boys) who had undergone respiratory function testing in infancy were retested at school age (7-12 years). Measurements included lung volume, airways resistance, peak flow, and spirometry. Clinical problems were investigated by questionnaire. Twelve children had repeat measurements after taking salbutamol. RESULTS: Predominant complaints were non-productive cough and dysphagia, but even those children with major problems in infancy reported few restrictions at school or in sport or social activities. Respiratory function and clinical findings at school age appeared unrelated to status in infancy, such that even the patients with severe tracheomalacia requiring aortopexy did not have lung function testing suggestive of malacia at school age. Most patients showed a restrictive pattern of lung volume which would appear to result from reduced lung growth after surgery rather than being a concomitant feature of the primary congenital abnormality. Although six children reported wheeze and four had a diagnosis of asthma, only one responded to salbutamol. This suggests that a tendency to attribute all lower respiratory symptoms to asthma may have led to an overdiagnosis of this condition in this patient group. CONCLUSION: Respiratory function testing in infancy is of limited value in medium term prognosis, but may aid management of contemporary clinical signs. In children respiratory function testing is valuable in assessing suspected asthma and effects of bronchodilators. PMID- 10519714 TI - Accuracy of clinical assessment of heart murmurs by office based (general practice) paediatricians. AB - AIM: To determine the diagnostic accuracy of physical examination by office based (general practice) paediatricians in the evaluation of heart murmurs. DESIGN: Each of 30 office based paediatricians blindly examined a random sample of children with murmurs (43% of which were pathological). Sensitivity and specificity were calculated and were related to paediatricians' characteristics. RESULTS: Mean (SD) sensitivity was 82 (24)% with a mean specificity of 72 (24)% in differentiating pathological from innocent murmurs, with further investigations requested for 54% of assessments. The addition of a referral strategy would have increased mean sensitivity to 87 (20)% and specificity to 98 (8)%. Diagnostic accuracy was not significantly related to the paediatricians' age, education or practice characteristics, but was related to referral practices and confidence in assessment. CONCLUSIONS: Diagnostic accuracy of clinical assessment of heart murmurs by office based paediatricians is suboptimal, and educational strategies are needed to improve accuracy and reduce unnecessary referrals and misdiagnosis. PMID- 10519715 TI - The natural history of ventricular septal defects. AB - AIMS: To correlate the size and position of isolated ventricular septal defects with closure rate in a cohort of children with mean follow up of more than six years. DESIGN: A birth cohort was identified using the northern region cardiac database. The following were noted from case notes: defect size, position, means of closure, and age at closure. RESULTS: 68 children were identified. 49 defects were small, 14 were moderate, and 5 were large. 13 cases required surgical closure, including 12 perimembranous defects. 35 defects closed spontaneously. Nine of the small muscular defects remained open and five of the small perimembranous defects remained open. The spontaneous closure rate for muscular defects was significantly greater than for perimembranous defects. Mean age of follow up for patients who still have defects is 76 months. CONCLUSIONS: The position of a ventricular septal defect is extremely relevant to its natural history. Perimembranous defects accounted for most of the moderate and large defects that required surgical intervention. After more than six years almost a third of all perimembranous and just over two thirds of all muscular defects closed spontaneously. PMID- 10519716 TI - Diagnostic markers of infection: comparison of procalcitonin with C reactive protein and leucocyte count. AB - BACKGROUND: Procalcitonin has been advocated as a marker of bacterial infection. OBJECTIVE: To evaluate diagnostic markers of infection in critically ill children, comparing procalcitonin with C reactive protein and leucocyte count in a paediatric intensive care unit (PICU). METHODS: Procalcitonin, C reactive protein, and leucocyte count were measured in 175 children, median age 16 months, on admission to the PICU. Patients were classified as: non-infected controls (43); viral infection (14); localised bacterial infection without shock (25); bacterial meningitis/encephalitis (10); or septic shock (77). Six children with "presumed septic shock" (without sufficient evidence of infection) were analysed separately. Optimum sensitivity, specificity, predictive values, and area under the receiver operating characteristic (ROC) curve were evaluated. RESULTS: Admission procalcitonin was significantly higher in children with septic shock (median 94.6; range 3.3-759.8 ng/ml), compared with localised bacterial infection (2.9; 0-24.3 ng/ml), viral infection (0.8; 0-4.4 ng/ml), and non-infected controls (0; 0-4.9 ng/ml). Children with bacterial meningitis had a median procalcitonin of 25.5 (7.2-118.4 ng/ml). Area under the ROC curve was 0.96 for procalcitonin, 0.83 for C reactive protein, and 0.51 for leucocyte count. Cut off concentrations for optimum prediction of septic shock were: procalcitonin > 20 ng/ml and C reactive protein > 50 mg/litre. A procalcitonin concentration > 2 ng/ml identified all patients with bacterial meningitis or septic shock. CONCLUSION: In critically ill children the admission procalcitonin concentration is a better diagnostic marker of infection than C reactive protein or leucocyte count. A procalcitonin concentration of 2 ng/ml might be useful in differentiating severe bacterial disease in infants and children. PMID- 10519717 TI - Opportunistic immunisation in hospital. AB - AIM: To assess the potential for administering catch up and scheduled immunisations during hospital admission. METHODS: Immunisation status according to the child's principal carer was checked against official records for 1000 consecutively admitted preschool age children. Junior doctors were instructed to offer appropriate vaccination before discharge, and consultants were asked to reinforce this proactive policy on ward rounds. RESULTS: Excluding those children who were not fully immunised against pertussis through parental choice, 142 children (14.2%) had missed an age appropriate immunisation and 41 were due a scheduled immunisation. None had a valid contraindication. Only 43 children were offered vaccination on the ward but uptake was 65% in this group. CONCLUSIONS: Admission to hospital provides opportunities for catch up and routine immunisations and can contribute to the health care of an often disadvantaged group of children. These opportunities are frequently missed. Junior doctors must be encouraged to see opportunistic immunisation as an important part of their routine work. PMID- 10519719 TI - Can body size predict infant energy requirements? AB - Traditionally, infant energy requirements have been predicted from body size or age, whereas in older children and adults, physical activity is also taken into account. However, the extent to which body size determines energy use in individual infants has not been considered. Data on 232 measurements of total energy expenditure obtained in 124 infants aged 1.5 to 12 months were used to assess the relation between body size and energy use in individuals. Age, weight, and fat free mass consistently predicted total energy expenditure with an error of 21-23%. This contrasts greatly with the error of 10% with which infant basal metabolism can be predicted from anthropometry. Body size is a poor index of the total energy requirements of individual infants, and predictive equations generated from data on healthy infants will be inappropriate for disease states where physical activity or growth is altered. PMID- 10519718 TI - Effect of an individualised training programme during weight reduction on body composition: a randomised trial. AB - OBJECTIVE: To study the effect of a standardised training programme focusing on maintenance of fat free mass during weight reduction by energy reduction in obese children. DESIGN: Randomised trial of physical training programme and dietary advice (group A) versus dietary advice alone (group B). SUBJECTS: Thirty obese children and adolescents (14 group A, 16 group B) participated in the 12 week long programme; 20 children (10 group A, 10 group B) were also reassessed after one year. MEASUREMENTS: Fat free mass was estimated from the resistance index, obtained by bioelectrical impedance analysis at baseline, after four, eight, and 12 weeks in all subjects, and after one year in 20 subjects. RESULTS: The mean (SD) change in fat free mass was significantly different between the two groups after 12 weeks (group A, 2.68 (3.74) kg; group B, 0.43 (1.65) kg). The change in body weight after one year was inversely correlated with the change in fat free mass after 12 weeks (r = -0. 44), as assessed in the 20 subjects. CONCLUSIONS: A standardised training programme as used in this study can prevent reduction in fat free mass during weight loss in obese children. Reduction in fat free mass during weight reduction might be a risk factor for regain of weight. PMID- 10519720 TI - Epidemiology of pyridoxine dependent and pyridoxine responsive seizures in the UK. AB - OBJECTIVE: To study the epidemiology of pyridoxine dependent seizures and other forms of pyridoxine responsive seizures. DESIGN: Monthly notifications to the British Paediatric Surveillance Unit over two years. Questionnaire follow up. SETTING: UK and the Republic of Ireland. PATIENTS: Children aged 15 years or younger whose seizures respond to pyridoxine. INTERVENTIONS: None. MAIN OUTCOME MEASURES: Numbers of children with definite, probable, and possible pyridoxine dependent seizures or other seizures responsive to pyridoxine. RESULTS: Point prevalence and birth incidence: 1/687 000 and 1/783 000, respectively (definite and probable cases); 1/317 000 and 1/157 000, respectively (all types of pyridoxine responsiveness). NOTIFICATIONS: Pyridoxine dependency: 14 definite, 9 probable, and 10 possible cases; neonatal seizures not meeting case definitions: 7; infantile spasms: 5. Eight of 18 families of definite/probable cases had 2 affected siblings. Just over a third had atypical presentations and just under a third had features and/or initial diagnoses of birth asphyxia and neonatal hypoxic ischaemic encephalopathy. CONCLUSIONS: Pyridoxine dependency is rare. Atypical presentations are relatively frequent. A trial of pyridoxine is justified in all cases of early onset intractable seizures or status epilepticus, whatever the suspected cause. PMID- 10519721 TI - Preliminary report of a toxicity study of hydroxyurea in sickle cell disease. French Study Group on Sickle Cell Disease. AB - AIM: To evaluate the tolerance of hydroxyurea in children affected with sickle cell disease. DESIGN: Questionnaire study of French physicians likely to treat patients with sickle cell disease. Data were collected on 101 children with sickle cell disease, treated for a median of 22 months, 36 of whom were treated for more than three years. 13 children were younger than 5 years of age at inclusion. RESULTS: Hydroxyurea was stopped for medical reasons in 11 patients: 6 failures, 1 pregnancy, 1 cutaneous rash, 1 leg ulcer, 1 lupus. Acute lymphoblastic leukaemia occurred in a girl treated for 1.5 months with hydroxyurea, this short interval arguing against a causative association. One 17 year old boy had paraparesis after 8 years of treatment. CONCLUSIONS: No major short or medium term toxicity was related to hydroxyurea in this cohort of 101 children. However, the number of children treated for more than 3 years is too few to make firm conclusions on the long term tolerance of this drug. PMID- 10519722 TI - Parent perceptions of symptom severity in Tourette's syndrome. AB - The families of 66 consecutive children with Tourette's syndrome were surveyed for their perception of symptom significance using a questionnaire. Families considered attention deficit and learning difficulties to be most significant, while motor and vocal tics were least important. When present, episodic rage was the most impairing symptom. Physicians must be aware of the significance of these comorbid symptoms for patients with Tourette's syndrome. PMID- 10519723 TI - Vincristine treatment revealing asymptomatic hereditary motor sensory neuropathy type 1A. AB - A 5 year old boy developed severe weakness after receiving vincristine for treatment of acute lymphoblastic leukaemia. Although weakness improved after the discontinuation of vincristine, other symptoms suggestive of a neuropathy persisted. Neurophysiological and genetic analysis at age 8 years indicated that vincristine had induced symptoms of a hereditary sensory motor neuropathy type 1A, which had previously been asymptomatic; his genetically affected mother was also asymptomatic. PMID- 10519724 TI - B cell non-Hodgkin's lymphoma in a girl with the DiGeorge anomaly. AB - The DiGeorge anomaly (DGA) is occasionally associated with cellular immunodeficiency. We report a female infant diagnosed with complete DGA, who developed fatal, high grade, non-Hodgkin's lymphoma that expressed Epstein-Barr virus (EBV). Non-Hodgkin's lymphoma should be considered in children with DGA. PMID- 10519725 TI - Diagnosis and management of late complications after liver transplantation. PMID- 10519726 TI - The role of radiology in the evaluation of stridor. PMID- 10519727 TI - Clinical and laboratory findings in referrals for mitochondrial DNA analysis. PMID- 10519728 TI - Pertussis is increasing in unimmunised infants: is a change of policy needed? PMID- 10519730 TI - Paediatrics and child health PMID- 10519729 TI - How to manage warts. PMID- 10519731 TI - Q&A about this issue PMID- 10519732 TI - Novel delivery systems for biotechnology drugs: recent progress toward improved therapies. PMID- 10519733 TI - Guidelines for the clinical use of benzodiazepines: pharmacokinetics, dependency, rebound and withdrawal. Canadian Society for Clinical Pharmacology. AB - Principles of benzodiazepine selection are outlined for various psychiatric indications and diverse populations (the elderly, and drug and alcohol abusers). Benzodiazepines are still among the most commonly used classes of medications, and they differ in their pharmacodynamic properties. They have varied uses as monotherapy or as adjunctive medication because of their efficacy in the treatment of conditions involving a dysfunction of the GABAergic system or where neuronal inhibition is required. In multiple therapy, benzodiazepines augment the efficacy of other drugs such as lithium in mania, antipsychotics in psychotic agitation and selective serotonin reuptake inhibitors in panic disorder. Benzodiazepines can produce dependence and tolerance in most patients; predisposed individuals are at greater risk. Short- and intermediate-beta half life compounds carry a greater risk of rebound and withdrawal reactions, and drug dependence than long acting agents. Adverse effects include sedation, psychomotor and cognitive impairment, memory loss, potentiation of other central nervous system depressants and treatment-emergent depression. Drug potency and beta elimination half-life are reviewed and compared as pharmacokinetic variables. PMID- 10519734 TI - Comparative effects of simvastatin and cholestyramine on plasma lipoproteins and CETP in humans. AB - Cholesteryl ester transfer protein (CETP) mediates neutral lipid transport in plasma, resulting in a net transfer of cholesteryl ester from high density lipoprotein to very low density lipoprotein. CETP gene expression is regulated by cholesterol, and plasma CETP level increases in patients with hyperlipidemia and with cholesterol feeding. Simvastatin, unlike cholestyramine, reduces hydroxymethylglutaryl coenzyme A reductase activity and may decrease a cellular pool of cholesterol, which is regulatory for CETP gene expression. The effects of simvastatin and cholestyramine on plasma lipids and CETP in 24 male and 19 female patients with primary hypercholesterolemia were compared. Following a four-week placebo period, patients were randomly assigned to receive either simvastatin or cholestyramine. Medication was increased in a stepwise fashion (from 10 to 40 mg for simvastatin and from 8 to 24 g for cholestyramine) as required at six-week intervals to maintain a low density lipoprotein cholesterol (LDL-C) level below 3.4 mmol/L. At the end of the 18-week study, the mean dose of simvastatin was 28.6 mg/day and of cholestyramine 19.3 g/day. Simvastatin was more effective than cholestyramine in lowering LDL-C (-36.8% versus -27. 2%; P=0.031) and triglycerides (-8.5% versus +12.5%; P=0.045). Plasma CETP level decreased by 14.8% following treatment with simvastatin (P=0.003) but did not change following cholestyramine treatment. This study demonstrates that, compared with cholestyramine, simvastatin results in more favourable improvements in the plasma lipoprotein profile and also lowers plasma levels of CETP. PMID- 10519735 TI - Prescription and nonprescription drug use among at-risk community-dwelling seniors in Ottawa-Carleton. AB - BACKGROUND: The Council on Aging Ottawa-Carleton conducted an evaluation study aimed at reducing the inappropriate use of medication by community-dwelling seniors. During a 17-month period from 1990 to 1992, 278 of 415 seniors consented to participate. All were new referrals to home care, all were 65 years of age or older and all were taking a minimum of six prescription medications. OBJECTIVE: To describe the patterns of prescription and nonprescription drug use among high risk elderly people living in the Ottawa-Carleton region. ANALYSIS: Descriptive statistics and logistic regression were used to explore relationships between the numbers of drugs (prescription and nonprescription), drug classes and drug subclasses, and sex, age and number of prescribing physicians. RESULTS: The top 10 prescription or nonprescription drugs taken by study participants were consistent among diseases and health problems most commonly found among the elderly. Comparisons by rank for subclasses of drugs indicated that eye, ear, nose and throat (EENT), and respiratory drugs were considerably more prevalent among males than females, while women took more hormones and anxiolytics, sedatives and hypnotics. Age also appeared to have an effect; the proportions of respiratory drugs and hormones decreased with age. In contrast, vitamins and EENT drugs were most prevalent among the oldest seniors. These findings were supported by the results of multivariate analyses. Having three or more prescribing doctors was positively associated with the use of laxatives. Findings from multivariate analyses indicated that for prescription drugs, the youngest age group was about four times more likely than the oldest age group to be taking at least eight drugs. For nonprescription drugs, the only statistically significant finding was that those with three or more doctors were twice as likely as those with one or two doctors to take at least one nonprescription drug. PMID- 10519736 TI - Identifying appropriate subjects for abuse liability studies using prestudy pharmacological testing. AB - Drug abuse liability testing usually involves a subject population of individuals who are current or former drug abusers. To determine whether choosing subjects from a wider and more diverse population of nondrug abusers results in subjective response patterns that are pharmacologically associated with drug abuse liability, a series of studies were conducted using the prescription opiate hydromorphone and the barbiturate secobarbital to prescreen subjects before their acceptance into large multidrug, multidose abuse liability studies. The results of these prescreening studies show that there were some subjects who were not able to report consistently and reliably relevant drug effects, even though they received pharmacologically active doses as measured by objective indexes such as pupil diameter for the opiate study and psychomotor impairment for the barbiturate study. To test the consistency of these results, several of the subjects from the hydromorphone screening were retested with hydrocodone. The results demonstrate that subjects who were unable to report positive subjective effects of hydromorphone were also unable to respond appropriately to hydrocodone, and subjects who responded appropriately to hydromorphone in the prescreening study consistently responded to another opiate. Using this prescreening methodology has allowed sensitive and reliable data on the abuse liability of benzodiazepine agonists and partial agonists, barbiturates, carbamate compounds, amphetamines, selective serotonin reuptake inhibitors and prescription opiate compounds to be generated using a nondrug abusing population. In conclusion, due to the relative heterogeneity of a population of nondrug abusers compared with a drug abusing population, it is necessary to prescreen subjects for their ability to detect and report subjective drug effects, and to distinguish the effects of an active drug from those of placebo. Selecting subjects from this population has several advantages over selecting subjects from the drug abusing population that outweigh any inconvenience of this simple prescreening methodology. PMID- 10519737 TI - Pilot study to improve health outcomes for medication-induced headache sufferers. AB - The objectives of this prospective cohort study were to identify and encourage individuals with symptoms consistent with medication-induced headache (MIH) to seek medical help, and to determine the health outcomes of these patients. All community pharmacists (approximately 150) on Newfoundland's Avalon Peninsula were provided with continuing education material on MIH and were asked to display a poster inviting individuals taking analgesics more than two days a week for headache to speak to their pharmacist. Pharmacists were asked to provide a patient information pamphlet to individuals with medication use and symptoms consistent with MIH, to request them to contact a study nurse and to log the number of potential MIH patients encountered. The study nurse supervised the completion of headache and quality of life questionnaires, and advised the subjects to seek help from their family physicians. Forty-six pharmacists returned logs identifying 142 potential MIH subjects: 21 contacted the study nurse, 17 completed the questionnaires and 11 were classified as MIH sufferers. MIH sufferers reported a reduction in headache frequency (P<0.02) and analgesic consumption (P<0.05) when re-interviewed after three months. A health care team approach can improve health outcomes for some MIH sufferers. Further work to evaluate strategies to encourage chronic headache sufferers to seek help is required. PMID- 10519740 TI - Routine dental diagnostic imaging in hematopoietic cell transplantation PMID- 10519739 TI - Routine dental diagnostic imaging in hematopoietic cell transplantation. PMID- 10519741 TI - Temporomandibular disorders: a review of current understanding. AB - OBJECTIVE: The purpose of this article is to conduct a narrative review of current evidence regarding the understanding, evaluation, management, and treatment of temporomandibular disorders to provide a broad perspective and updated introduction to an important and controversial subject with rapidly changing developments and limited well-designed research. DATA SOURCES: Studies were identified through a search of MEDLINE for 3 topics (temporomandibular disorder, temporomandibular joint, and chronic pain) over a 10-year period (January 1988 to August 1998) and of bibliographies of identified studies and review articles. STUDY SELECTION: More than 5000 articles were produced. In-depth review of all of this literature was beyond the scope of the present article, which is intended to provide an overview. The amount and diversity of the literature and the limitations of covering such a broad topic being recognized, the papers selected were those that reviewed limited topics or studied focused areas. This report is not a systematic (qualitative) or meta-analysis (quantitative) review. An acknowledged limitation of this narrative review method lies in the potential for bias in selection. The referenced works do not include all papers reviewed; only pertinent literature and reviews with comprehensive references were selectively included. CONCLUSIONS: Advances in basic and clinical science have resulted in important changes in the understanding and management of temporomandibular disorders. Many treatments are not supported by research, and the role of dentistry is changing to a more diagnostic and management-based model from the hands-on treatment procedures of the past. The present science-based understand-ing of a biopsychosocial disorder is important in properly and responsibly dealing with patients with temporomandibular disorders. PMID- 10519742 TI - Sarcoidosis: medical and dental implications. PMID- 10519743 TI - Is wire osteosynthesis obsolete for fixation of Le Fort I osteotomies? PMID- 10519744 TI - Medical legal considerations in temporomandibular disorders. AB - Dentists commonly see patients with temporomandibular disorders who are involved in litigation, and dentists are occasionally accused of temporomandibular disorder-related negligence. Rapidly changing scientific knowledge must be reflected in dental practice and in the courtroom, where dentists act as experts. In this article, concepts pertaining to assessments of patients with temporomandibular disorders for impairment and disability and concepts pertaining to legal causation are discussed, especially in relation to motor vehicle accidents. Recommendations for avoiding malpractice suits are presented. PMID- 10519746 TI - Preradiation dental extraction decisions in patients with head and neck cancer. AB - OBJECTIVE: The first objectives of this international survey were to study how dental-related and radiotherapy-related risk factors influence clinicians' preradiation dental decision making in patients with head and neck cancer and to evaluate clinicians' degree of certainty in making such decisions. A further objective was to examine the correlation of clinicians' policies with a policy based on a model for dental decision support that was presented in an earlier article. STUDY DESIGN: A consensus questionnaire was mailed to 54 oral maxillofacial surgeons and hospital-based dentists at a number of international locations. The responses were aggregated and anonymously analyzed through use of a multiple regression procedure. RESULTS: Forty-four clinicians returned the questionnaire (response rate, 81%). Nine clinicians (20%) were using printed clinical guidelines for preradiation dental screening. Eighty-eight percent of clinicians' preradiation decisions and 49% of their certainty could be explained by the studied risk factors. Not all risk factors were significant at P <.001. Clinicians' policies showed high correlation (.85) with the policy based on the model for dental decision support. CONCLUSIONS: The findings support our previous assumption that policies in this field seem to be based primarily on clinical experience and opinions rather than on evidence-based clinical guidelines. We conclude that the clinical usefulness and validity of the model for dental decision support should now be tested and that it could also serve as a training tool. PMID- 10519745 TI - The effect of midazolam sedation on indicators for myocardial ischemia. AB - OBJECTIVE: The purpose of this investigation was to examine hemodynamic and electrocardiographic responses to local anesthesia. STUDY DESIGN: Seventy-five patients with heart disease were treated in 2 groups with lignocaine 2%, adrenaline 1:50,000, and vasopressin 0.25 IU, either alone or with midazolam. Heart rate, blood pressure, and electrocardiogram values were recorded, and the rate-pressure product and pressure-rate quotient were calculated as indicators for myocardial ischemia. RESULTS: Sedation and anesthesia induced significant changes to the means for heart rate and systolic and mean blood pressure. For the anesthetic group, the maximum value for the rate-pressure product was 12,168 (95% CI = 1368), the minimum value for the pressure-rate quotient was 1.39 (95% CI = 0.04), and one patient exhibited minor changes to the S-T segment. For the sedated group, the rate-pressure product maximum was 9,882 (95% CI = 1,226) and the pressure-rate quotient fell to 1.13 (95% CI = 0.06), but no patient experienced ischemia. CONCLUSION: Values for these indicators suggested that treatment did not generate significant ischemic risk. PMID- 10519747 TI - Specific expression of interleukin-1 beta in temporomandibular joints with internal derangement: correlation with clinical findings. AB - OBJECTIVE: Interleukin-1 beta appears to play an important role in the pathophysiology of joint diseases. The aim of this study was to analyze the expression of interleukin-1 beta in temporomandibular joint internal derangement. STUDY DESIGN: Using an immunohistochemical technique with specific antibodies, we examined 20 human temporomandibular joint samples from patients with internal derangement of the temporomandibular joint: 5 extirpated disks and 15 biopsy specimens from the synovitic area of the temporomandibular joint upper compartment. We also examined 2 control specimens. The evaluation of interleukin 1 beta expression compared with clinical findings. RESULTS: Interleukin-1 beta was predominantly localized in the synovial lining cells and endothelial cells of blood vessels. Statistically significant correlation was found between interleukin-1 beta expression and some clinical findings. CONCLUSIONS: The results suggest that interleukin-1 beta may be involved in the pathogenesis of temporomandibular joint internal derangement and that the intensity of interleukin-1 beta expression may correlate with clinical findings, especially pain. PMID- 10519748 TI - Evaluation of electronic dental anesthesia in children. AB - OBJECTIVE: The purpose of this study was to determine the effectiveness and acceptance of electronic dental anesthesia in comparison with local anesthesia in restorative procedures in children. STUDY DESIGN: Twenty-eight children, aged 6 to 12 years, participated in the study. Each patient had symmetric teeth requiring class I cavity preparations. One tooth was treated with electronic dental anesthesia; the antimere tooth was treated with local anesthesia. The tooth and the method were selected at random, and the two restorations were performed at the same appointment. Pain was assessed by means of two pain scales, the color scale and the sound, eye, and motor scale. Behavior was assessed through use of the North Carolina Behavior Rating Scale. The ratings of pain and behavior were made at 4 separate intervals (after rubber dam clamp placement, with the handpiece operating adjacent to the tooth, during penetration of the dentin-enamel junction of the tooth, and 5 minutes postoperatively). The recorded values for the steps of the restorations completed with electronic dental anesthesia and local anesthesia were analyzed by means of chi(2) analysis to determine any statistically significant difference between the techniques at a level of P <.05. RESULTS: Although the success rate of electronic dental anesthesia was less than that of local anesthesia, there was no significant difference between the two methods. On the other hand, 53.6% of the patients preferred electronic dental anesthesia, whereas 35.7% preferred local anesthesia. CONCLUSIONS: In restorative dental care in children, electronic dental anesthesia appears to be beneficial in reducing discomfort, as judged from behavioral observations and self-reports. PMID- 10519750 TI - The evaluation of cutaneous, genital, scalp, nail, esophageal, and ocular involvement in patients with oral lichen planus. AB - OBJECTIVE: Lichen planus, in its classical presentation, involves the oral cavity and skin. This study evaluated patients with oral lichen planus for extraoral manifestations of the disease. STUDY DESIGN: A total of 584 patients with oral lichen planus were evaluated for cutaneous, genital, scalp, nail, esophageal, and ocular lichen planus. RESULTS: Extraoral manifestations included cutaneous lichen planus in 93 patients, genital lichen planus in 19% of 399 examined women and 4.6% of 174 examined men, nail involvement in 11 patients, lichen planopilaris in 6 patients, esophageal lichen planus in 6 patients, and conjunctival lichen planus in 1 patient. Thirty-three patients developed lichen planus in 3 or more sites. CONCLUSIONS: Because a relatively high percentage of patients with oral lichen planus develop extraoral manifestations, a thorough evaluation should routinely be performed. A complete history and physical examination by a multidisciplinary group of health care providers uncovers common and uncommon extraoral features of the disease. PMID- 10519749 TI - Contemporary issues in the diagnosis of oral pemphigoid: a selective review of the literature. AB - Pemphigoid is a group of bullous diseases that have a diversified morphologic presentation and affect the skin, oral mucosa, and other mucosal membranes, alone or in combination. In the literature, the condition has been subclassified into bullous pemphigoid and cicatricial pemphigoid (mucous membrane pemphigoid) on the basis of the primary organ of involvement. In addition to the clinical presentation and a subepithelial vesicle or bullae on routine histologic analysis, the diagnosis is based on direct and indirect immunofluorescence studies. Recent investigations indicate that different clinical groups of patients with pemphigoid produce autoantibodies to different molecules within the basement membrane zone. Based on these recent observations and a review of the literature, a viewpoint is presented that not all patients with cicatricial pemphigoid should be grouped together. Rather, they should be classified into subgroups-ocular, oral, etc-on the basis of the clinical phenotype and long-term follow-up. Such a division will facilitate the provision of appropriate and relevant treatment plans; if the clinical course changes, the diagnosis can be adjusted. This strategy will prevent patients with disease limited to the oral cavity from receiving systemic drugs or agents that may be more harmful than beneficial. PMID- 10519751 TI - Oral health in hospitalized and nonhospitalized community-dwelling elderly patients. AB - OBJECTIVE: The purpose of this study was to compare hospitalized and nonhospitalized home-dwelling elderly patients in a single community with respect to oral health and general health and to study risk factors for edentulousness in these patients. The study hypothesis was that hospitalized elderly patients would have poorer oral health than nonhospitalized elderly patients. STUDY DESIGN: Oral health status was examined according to the World Health Organization's guidelines for 181 hospitalized patients (mean age, 81.9 +/- 5.8 years) in a geriatric ward and for 254 home-living patients (mean age, 76.9 +/- 5.6 years). Data regarding the patients diseases and medications came from hospital records and doctors prescriptions and were categorized on the basis of the International Classification of Diseases. Differences between the hospitalized and nonhospitalized patients, between genders, between age groups, and between the various disease and medication groups were analyzed. Logistic regression was used to analyze the effects of study variables on edentulousness. RESULTS: The mean number of teeth was 10. 3 +/- 7.6 in the hospitalized patients and 16.3 +/- 7.4 in the nonhospitalized patients (P <.001). The mean number of decayed teeth was 1.3 +/- 2.2 in the hospitalized patients and 0.6 +/- 0.9 in the nonhospitalized patients (P <.01). All dentate patients had poor periodontal health. Community Periodontal Index scores were between 2 and 4 in 94.8% of the hospitalized patients and 98.6% of the nonhospitalized patients. Edentulousness was observed in 66.3% of the hospitalized patients and 42.1% of the nonhospitalized patients (P <.001). In both groups, female gender (odds ratio, 2.0; CI, 1.3-3. 1) and age between 80 and 89 years (odds ratio, 2.5; CI, 1.5-4.4) were the strongest risk factors for edentulousness. The number of drugs used daily also correlated significantly with the loss of teeth (P <.05). In the nonhospitalized patients, edentulousness correlated significantly with cardiovascular diseases and drugs taken daily (P <.01), whereas in the hospitalized patients such an association was not found. CONCLUSIONS: The results of this study confirmed our hypothesis that hospitalized elderly patients who had many concomitant diseases and used many drugs daily had worse dental health than nonhospitalized home-dwelling elderly patients. The nature of a patient's illness was not a significant factor in this respect. Female gender and age between 80 and 89 years were the strongest factors for edentulousness in both patient groups. PMID- 10519752 TI - Ultraviolet B irradiation: a new therapeutic concept for the management of oral manifestations of graft-versus-host disease. AB - Ultraviolet irradiation inhibits the proliferative responses of lymphoid cells to mitogens and alloantigens by inactivation of T lymphocytes and antigen-presenting cells. Its immunosuppressive capacity led to the introduction of UV irradiation into clinical practice for the treatment of dermatologic manifestations of chronic graft-versus-host disease. The cumulative experience with psoralen-UV-A rays in the treatment of cutaneous and oral graft-versus-host disease was the incentive for the application of oral UV-B rays in 2 patients with oral graft versus-host disease signs and symptoms after allogeneic marrow transplantation. Intraoral UV-B irradiation (0.02 mJ/cm(2)) was administered 2 or 3 times per week on an ambulatory basis; the dose was increased by 0. 02 mJ/cm(2) every fourth session. Both patients responded early and satisfactorily, displaying only minimal side effects at a relatively low cumulative dose. Intraoral UV-B proved a valuable modality in the treatment of resistant chronic oral graft-versus-host disease. PMID- 10519753 TI - Gingival fibrous nodule. AB - Five patients with small, exophytic, asymptomatic fibrous nodules, single or multiple, of the mandibular mucogingival zone of the anterior labial gingiva are presented. Such nodules are benign and resemble small fibromas. The clinical and histologic appearance distinguishes them from entities such as retrocuspid papilla, reticular mandibular gingival ridges, oral hamartomas associated with Cowden's syndrome, tuberous sclerosis oral lesions, and oral nodules secondary to epidermolysis bullosa. Gingival fibrous nodule, or gingival nodule, is the proposed name for this lesion, which is essentially a variation of normal oral structures that may be mistaken for disease. PMID- 10519754 TI - Focal parotid necrosis in systemic lupus erythematosus: case report and review of the literature. AB - Systemic lupus erythematosus (SLE) is a disease that may affect a number of organ systems, particularly the joints, skin, kidneys, heart, lungs, and immune system. Salivary gland involvement is usually associated with Sjogren's syndrome, in which lymphocytic acinar infiltrates predominate histologically. We present the case of a 29-year-old woman with SLE who developed bilateral parotid enlargement with a unilateral focus of parotid necrosis that was consistent with a cystic mass on computerized tomography. A biopsy of this lesion was histologically similar to a cervical lymph node biopsy in the same patient, with both specimens showing loss of architecture and foci of necrosis consisting of nuclear dust, histiocytes, and scattered plasma cells without formation of granulomata or presence of multinucleated giant cells; these findings are classic for SLE lymphadenopathy. We believe this to be the first reported case of focal necrosis in the parotid gland directly associated with SLE. PMID- 10519756 TI - Cell cycle-associated proteins in melanotic neuroectodermal tumor of infancy. AB - OBJECTIVE: The purpose of the present study was to compare the immunohistochemical expression of cell cycle-associated proteins in neuroblastic and melanocytic cell populations of melanotic neuroectodermal tumor of infancy. STUDY DESIGN: Three cases of melanotic neuroectodermal tumor of infancy were selected. The immunohistochemical expression of MDM-2, p53, proliferating cell nuclear antigen, cyclin D1, and cyclin A was assessed through use of the streptavidin-biotin-peroxidase complex technique. RESULTS: Positive immunostaining for MDM-2, proliferating cell nuclear antigen, cyclin D1, and cyclin A was occasionally observed in the large melanin-containing epithelioid cells. CONCLUSIONS: These data suggest that MDM-2 expression may be important for the development of melanotic neuroectodermal tumor of infancy and that the melanocytic cell population, not the neuroblastic one, is the proliferative component of the tumor. PMID- 10519755 TI - Solitary fibrous tumor of the buccal mucosa: report of a case with immunohistochemical studies. AB - We describe a case of a solitary fibrous tumor of the buccal mucosa and report the results of immunohistochemical studies of the lesion. Solitary fibrous tumors are extremely rare in the intraoral region. These tumors are generally difficult to diagnose because of their broad range of morphologic characteristics. We regard the expression of CD34 within the appropriate clinical and morphologic setting, in the absence of reactivity for other specific markers of differentiation, as evidence supporting the diagnosis of solitary fibrous tumor. PMID- 10519757 TI - Superficial mucocele: report of 4 cases. AB - Four cases of the lesion first described as superficial mucocele by Eveson in 1988 are reported. All of the lesions developed in adult women; two of the women had concurrent oral lichen planus. The mucoceles were found on the soft palate, the buccal mucosa, and the upper and lower labial mucosa. The etiologic factors and pathogenesis of this lesion are discussed. PMID- 10519758 TI - Tumor-doubling time and onset of pulmonary metastasis from adenoid cystic carcinoma of the salivary gland. AB - Adenoid cystic carcinoma (ACC), an uncommon malignancy in the head and neck region, invades diffusely and often metastasizes to the lung, although the growth rate is very slow. A retrospective study was conducted in 30 patients with ACC to ascertain the frequency of pulmonary metastasis, the doubling time of metastatic tumor deposits, and the time of onset for pulmonary metastasis. The following results were obtained: (1) Of 30 patients with ACC, 21 had pulmonary metastases (4 initially and 17 during observation), 7 were free of metastases but have not been observed for 5 years, and 2 were free of metastases for more than 5 years but less than 10 years after the initial treatment. The cumulative metastasis rate at 5 and 10 years for this group of patients was 70% and 100%, respectively. (2) Patients with T1 or T2 tumors that have a tubular or cribriform histopathologic pattern showed pulmonary metastases about 20 months later than those with T3 or T4 tumors and a solid pattern. However, the final metastasis rate did not differ between the 2 groups after a long period. (3) The tumor doubling time of the metastatic deposits of ACC was 86 to 1064 days with an average of 393 days, which was much longer than that of most other malignant neoplasms reported previously. (4) The time of onset of pulmonary metastasis was calculated to be much earlier (average of 227 months) before the first visit. These findings suggest that the treatment method for ACC should be chosen with the consideration that many of the patients may have occult pulmonary metastases at the time of their initial evaluation. PMID- 10519759 TI - Examination of the roots of paramolar tubercles with computed tomography: report of 3 cases. AB - Computed tomography was used to examine root anatomy in 3 cases of paramolar tubercles. The images clearly showed the structure of the paramolar tubercles, including their root canal morphology. The root of the paramolar tubercle was united with the distobuccal root in each case. Canals were observed in all tubercles and were connected with the canals in distobuccal roots at various levels. In one case, the imaging information was helpful for endodontic treatment. The clinical importance of knowledge of the root and canal anatomy of these paramolar tubercles is discussed. PMID- 10519760 TI - Application of titanium-alloy endodontic implants in conjunction with periradicular surgery. AB - OBJECTIVE: To evaluate the outcome of placement of titanium-alloy endodontic implants in conjunction with periradicular surgery. STUDY DESIGN: Twenty-four teeth were treated with endodontic implants with corresponding periradicular surgery by using SuperEBA cement (Harry J Bosworth Co, Skokie, Ill) as a sealer. The results of the endodontic implant surgery were evaluated clinically and radiographically from 2 to 4 years after treatment. RESULTS: Twenty-two teeth were treated successfully, whereas 2 teeth were treated unsuccessfully, for a success rate of 92%. CONCLUSIONS: Titanium-alloy endodontic implants in conjunction with periradicular surgery may provide good short-term results. Sealing between the endodontic implant and the dentin is likely an important factor for success. PMID- 10519761 TI - Localization and changes in superoxide dismutase immunoreactivity in rat pulp after tooth preparation. AB - OBJECTIVE: To examine the distribution of superoxide in the uninflamed and inflamed dental pulp by characterizing the immunoreactivity of the detoxifying antioxidant enzymes, manganese and copper-zinc superoxide dismutases (MnSOD and CuZnSOD, respectively). STUDY DESIGN: In 12 rats, mesial cavity preparations were made on the maxillary right first molar; left molars were unoperated controls. After 5 days, the rats were killed, and histologic sections were processed by using MnSOD and CuZnSOD immunoreactivity, and the extent of inflammation was evaluated on alternate sections stained with hematoxylin and eosin. RESULTS: In the hematoxylin and eosin-stained sections, inflammation was consistent with round-cells: macrophages, lymphocytes, and plasma cells that coalesced into a distinct leukocytic "lesion", which obliterated portions of the underlying pulp. Both MnSOD and CuZnSOD immunoreactivity increased dramatically in inflammatory cells within the leukocytic lesion and in the tissue surrounding the lesion. CONCLUSIONS: The findings suggest that the protective role of SOD increases within pulp cells that are undergoing inflammatory stimulation. SOD immunoreactivity may be an early indicator of stress in pulp. PMID- 10519762 TI - Value of contrast-enhanced magnetic resonance imaging in differentiation of hemangiomas from lymphangiomas in the oral and maxillofacial region. AB - OBJECTIVE: The purpose of this study was to investigate the ability of contrast enhanced magnetic resonance imaging to differentiate hemangioma from lymphangioma in the oral and maxillofacial region. STUDY DESIGN: Contrast-enhanced magnetic resonance imaging was performed in 20 patients (21 masses: 17 hemangiomas and 4 lymphangiomas) through use of either a 0.2-T permanent system or a 0. 5-T superconductive system and spin-echo pulse sequences. RESULTS: After intravenous administration of contrast medium, enhancement was observed in all hemangiomas in areas corresponding to those with high signal on T(2)-weighted images. Unequivocally increased signal was observed in 10 masses, and slightly increased signal was observed in 7 masses. On the other hand, none of the lymphangiomas showed an enhancing mass on contrast-enhanced T(1)-weighted images. CONCLUSIONS: Although contrast-enhanced T(1)-weighted imaging may not improve delineation of masses in all cases, it can be used to differentiate between deep hemangiomas and lymphangiomas. PMID- 10519763 TI - Multiple idiopathic resorption in the primary dentition: review of the literature and case report. AB - Resorption of primary teeth is a normal physiologic process, except when it occurs prematurely. Resorption of permanent teeth is considered abnormal, and multiple etiologic factors have been implicated. A significant number of cases are represented by idiopathic resorption. Multiple idiopathic root resorption stands as a separate physiologic entity that has been described as affecting the entire permanent dentition. Multiple idiopathic resorption of primary teeth has not been previously reported. A case is described and a differential diagnosis is provided. The specific radiographic diagnostic criteria for this condition affecting the primary dentition are outlined. PMID- 10519764 TI - Hyperaeration of the sphenoid sinus: cause for concern? AB - A case of exuberant pneumatization of the left sphenoid sinus into the pterygoid process and floor of the middle cranial fossa is presented. The fact that pneumatization of the sphenoid sinus is frequently atypical is of clinical import because there is an intimate relationship between the contents of a hyperaerated sinus and adjacent vital facial and cranial structures. It is imperative that clinicians determine the location and extent of the walls of the sphenoid sinus and its relationship to adjacent vital structures whenever endoscopic sinus surgery is contemplated to avoid morbid consequences during surgery. PMID- 10519765 TI - The use of panoramic radiographs to localize displaced maxillary canines. AB - OBJECTIVE: The purpose of this investigation was to develop a reliable method of diagnosing the position of a displaced maxillary canine on the basis of a single panoramic radiograph. STUDY DESIGN: A total of 115 panoramic radiographs depicting 164 displaced maxillary canines were evaluated. The ratio of the width of the displaced canine to the width of the homolateral central incisor (the canine-incisor index) and the ratio of the width of the displaced canine to the width of the contralateral canine (the canine-canine index) were calculated. The height of the crown of each displaced canine was classified in the vertical plane, relative to the adjacent incisor, as apical, middle, or coronal. RESULTS: There was an overlap in the canine-incisor index ranges of the buccal (0.94-1. 45) and palatal (1.15-1.29) canines in the apical zone. In the middle and coronal zones, a clear difference could be seen between the canine-incisor indices of labially (0.78-1.11) and palatally (1.15-1.7) located canines. A cut-off point of 1.15 was determined. CONCLUSIONS: Provided that vertical restriction and the canine-incisor index are used, the panoramic radiograph can serve as a useful indicator for determining the position of an unerupted maxillary canine. PMID- 10519766 TI - Assessments of the physical performance of 2 generations of 2 direct digital intraoral sensors. AB - OBJECTIVE: To evaluate the performance of 4 intraoral direct digital sensors regarding their fundamental physical characteristics. The sensors are made by Schick Technologies Inc (Long Island City, NY) and Gendex Dental Systems (Milan, Italy). STUDY DESIGN: The sensors were exposed by using a Prostyle dental x-ray machine (Planmeca Oy, Helsinki, Finland) operating at 50 kV, 8 mA and various exposure times. Three test phantoms were used: a homogeneous 10-mm thick aluminum block, an aluminum block with a pattern of holes of varying sizes, and a resolving power target. Digital images were transferred as 8 bit TIFF files and analyzed by using a personal computer. RESULTS AND CONCLUSIONS: Some improvements could be observed in the physical performance of the new generation of direct digital radiographic sensors when compared with the earlier generation. Smaller pixels and higher quantum efficiency have improved sensor performance. PMID- 10519767 TI - Challenges for occupational health from work in the information society. PMID- 10519768 TI - Impact of a changing U.S. workforce on the occupational injury and illness experience. PMID- 10519769 TI - Form of employment in a project-intensive economy. PMID- 10519771 TI - How organizational change influences gender segregation in the workplace. PMID- 10519770 TI - Economic aspects of the work life in transition. PMID- 10519772 TI - Consequences of the flexible labor market on working conditions from a gender perspective. PMID- 10519773 TI - Promotion of work ability during ageing. PMID- 10519774 TI - Occupational injury fatalities among older workers in the United States, 1980 1994. PMID- 10519775 TI - Age and gender differences in exposure patterns and low back pain in the MUSIC Norrtalje study. PMID- 10519776 TI - Risks of fatal injuries to farm workers 55-years of age and older. PMID- 10519777 TI - Young workers at risk when working in agricultural production. PMID- 10519778 TI - Where African-American women work and the nonfatal work-related injuries they experienced in the U.S. in 1996, compared to women of other races. PMID- 10519779 TI - A minority with few occupational accidents: the case of Swedish-speaking Finns. PMID- 10519780 TI - Cold-related non-fatal injuries in Alaska. PMID- 10519781 TI - Workplace health promotion in Sweden: A collaborating network with 15 EU member states. PMID- 10519782 TI - Good occupational health service practice. PMID- 10519783 TI - Management of a sophisticated hearing conservation program. PMID- 10519784 TI - Real time indoor air monitoring system and analysis method. PMID- 10519785 TI - Enhanced particle filtration in a non-problem office environment: preliminary results from a double-blind crossover intervention study. PMID- 10519786 TI - Predicting system interactions in the design process. PMID- 10519787 TI - Intra-organization work for change: A model and two tools. PMID- 10519788 TI - The relationship of organizational factors to employee health and overall effectiveness. PMID- 10519789 TI - Surface haulage truck research. PMID- 10519790 TI - A workplace safety device for operators of remote-controlled continuous mining machines. PMID- 10519791 TI - Beryllium contamination inside vehicles of machine shop workers. PMID- 10519792 TI - Use of ambient aerosol for testing agricultural cabs for protection against pesticide aerosol. PMID- 10519793 TI - Evaluating engineering controls during asphalt paving using a portable tracer gas method. PMID- 10519794 TI - Workplace violence in Finland: high-risk groups and preventive strategies. PMID- 10519795 TI - A Swedish industrial research program 'Co-operative for Optimization of Industrial Production Systems Regarding Productivity and Ergonomics' (COPE). PMID- 10519796 TI - ELMERI observation method for predicting the accident rate and the absence due to sick leaves. PMID- 10519797 TI - Strategies to promote well-being in small enterprises. PMID- 10519798 TI - NIOSH control technology and intervention efforts for small businesses. PMID- 10519799 TI - Reducing injuries and illnesses among construction workers. PMID- 10519800 TI - Swedish contribution to European research network activities on detection and prevention of injuries due to occupational vibration exposures. PMID- 10519801 TI - Development of new technique for risk assessment using physiologically based toxicokinetic models. PMID- 10519803 TI - Community Partners for Healthy Farming: involving communities in intervention planning, implementation, and evaluation. PMID- 10519802 TI - Pathophysiological mechanisms behind work-related muscle pain syndromes. PMID- 10519804 TI - Danger of drilling into sealed and filled plow frames. PMID- 10519806 TI - Computerized accident reconstruction and training for metal/non-metal mines. PMID- 10519805 TI - Farm work planning simulation in multi-media: A comparative evaluation. PMID- 10519807 TI - Safer mine hoisting with conveyance position and load monitoring. PMID- 10519808 TI - Safety climate dimensions associated with occupational exposure to blood-borne pathogens in nurses. PMID- 10519809 TI - Effects of a preventive message in the organizational context: occupational latex allergy in hospitals. PMID- 10519810 TI - Evaluation of the Permea-Tec pads as new technology for the detection of chemical breakthrough in PPC. PMID- 10519811 TI - Incidence of occupational asthma: A comparison by reporting systems. PMID- 10519812 TI - Common surfactants form contact allergens at normal handling and storage. PMID- 10519813 TI - Development of a combined irritancy/phenotypic analysis assay for the identification and differentiation of chemicals with the potential to elicit irritation, IgE-mediated, or T cell mediated hypersensitivity responses. PMID- 10519814 TI - Role of sensitization routes in the development of type I hypersensitivity to natural rubber latex in mice. PMID- 10519815 TI - Health and exposure surveillance of Siberian asbestos miners: A joint Finnish American-Russian project. PMID- 10519816 TI - Hazardous occupational exposure and lung disease among nylon flock workers. PMID- 10519817 TI - Asphalt fumes: exposure to PAH and amines. PMID- 10519818 TI - Exposure to monoterpenes in Finnish sawmills. PMID- 10519819 TI - Determination of nicotine as an indicator of environmental tobacco smoke in restaurants. PMID- 10519820 TI - Rapid method for measurement of micro-organisms in waste water cleaning plants. PMID- 10519821 TI - In vitro toxicity of silica substitutes used for abrasive blasting. PMID- 10519825 TI - Effects of an Intergenerational Choir for Community-Based Seniors and College Students on Age-Related Attitudes. AB - The purpose of this study was to examine attitudes of college students and senior citizens towards each other by incorporating successful components of a senior citizens' music program into the "Adopt-A-Choir" program established in a university music education/therapy program. Data collection was accomplished using the Age Group Evaluation and Description Inventory (AGED), providing insight into attitudes classified in 4 domains/evaluative scales: Goodness, Positiveness, Vitality, and Maturity. Subjects ware members of the Senior Singers (n = 15, 15) and music education/therapy students enrolled in the Woman's Glee Club at a local university (n = 15, 12). Results of the pre/post AGED survey warn compared using the Wilcoxon Matched Pairs test, with means increasing significantly for the four domains (p <.02). Gains were greatest for the seniors, suggesting a stronger move from negative to positive attitudes. Though the gains were smaller with the university students, all changes were positive, with one exception: in the "vitality" domain, university student attitude ratings decreased on the continuum for "timid-assertive." Informal predictors further suggested the partnership between seniors and university students was enjoyed by both sets of participants. PMID- 10519826 TI - Effects of Information on Elementary Band Students' Attitudes Toward Individuals with Special Needs. AB - The purpose of this study was to assess elementary music students' attitudes toward individuals with disabilities and compare potential methods of altering these attitudes. An adapted Disability Factor Scale was initially administered to four elementary school bands who would be attending the same junior high where an individual with a hearing impairment was to be mainstreamed. Prior to the second administration, each school participated in a different preparation: (a) normal rehearsal, (b) videotape of individuals with disabilities anticipating in music activities, (c) videotape with disability label provided, and (d) videotape, label, and attribution of successful music participation. Examination showed a 1.00 point difference between highest and lowest rated disabilities on a 6-point scale. Females demonstrated a more positive attitude than males for 6 of the disabilities. Rank ordering indicated similarities between genders with visual scars most accepted and visual impairments, amputation, and epilepsy least accepted. Results of the second administration showed no differences among schools. Of the 10 disabilities addressed on the questionnaire, four were portrayed on the videotape. Scores for students who viewed the videotape showed no difference between means for statements referring to disabilities that were viewed versus those not viewed. PMID- 10519827 TI - A Descriptive Analysis of Music Therapists' Perceptions of Delivering Services in Inclusive Settings: A Challenge to the Field. AB - The purpose of this study was to examine the perceptions of music therapists toward inclusion (providing services within general education settings) and to determine their willingness to provide their services in these settings. A questionnaire was sent to 560 music therapists of which 373 responded (67%). A descriptive analysis indicated that although the vast majority of music therapists are providing their services in a segregated setting, they (a) overwhelmingly know about inclusion, (b) perceive benefits to clients with and without disabilities, and (c) are willing to provide their services within an inclusive setting. Why then do therapists so overwhelmingly provide their services in noninclusive settings? Possible answers to this question as well as the challenge this creates to the field of music therapy are discussed. PMID- 10519828 TI - Effects of Active Versus Passive Group Music Therapy on Preadolescents with Emotional, Learning, and Behavioral Disorders. AB - This study attempted to compare the behavioral effects of active, rhythm-based group music therapy vs. those of passive, listening-based group music therapy on preadolescents with emotional, learning, and behavioral disorders. It was hypothesized that preadolescents who participated in active music therapy would more significantly improve target behaviors than those involved in passive music therapy. Achenbach's Teacher Report Form (TRF) was used to confirm changes among subjects in attention, motivation, and hostility as rated by homeroom teachers. Twelve music therapy sessions were conducted over a 4-month period with three different groups of subjects (n = 16), with two groups participating in active music therapy and the other receiving passive music therapy. Results indicate that subjects improved significantly after receiving both music therapy interventions. The most significant change in subjects was found on the aggression/hostility scale. These results suggest that group music therapy can facilitate the process of serf-expression in emotionally disturbed/learning disabled adolescents and provide a channel for transforming frustration, anger, and aggression into the experience of creativity and self-mastery. Discussion of results also includes recommendations for chousing one music therapy approach over another based on personality types and/or clinical diagnoses of subjects. PMID- 10519829 TI - Presentation of Aural Stimuli to Newborns and Premature Infants: An Audiological Perspective. AB - The purpose of this study was twofold: (a) to examine extant research in the field of music with premature and full term infants in order to identify protocols being used in the presentation of musical stimuli to neonates and (b) to use knowledge gleaned from audiology as a basis for suggesting a standardized protocol for use of musical stimuli with infants. Articles considered appropriate for inclusion in the analysis met the following criteria: (a) presented data for the effects of music on a dependent measure, (b) had subjects who were identified as either premature or term newborns receiving treatment after birth and prior to discharge from the hospital, and (c) used music for some or all of the aural stimuli. Articles (N = 20) were categorized by demographic information, types of aural stimuli, independent variables, dependent measures, and protocol used to present the musical stimuli. Of primary importance to this study was the protocol used in each study to present musical stimuli. Data regarding total duration of stimuli per day, longest duration of stimuli per day, method of stimuli presentation, placement of speakers, decibel level of stimuli, and where;he decibel level was measured reveal that there is no standard protocol being followed with regard to the presentation of aural stimuli. Recommendations include future research on (a) determining a minimum gestational age where music therapy may be appropriate, (b) determining the frequency spectrum perceived by a premature infant, (c) determining the decibel levels reaching the ear drum and assessing appropriate levels for minimum stimulation with maximum results, and (d) carefully considering the method of stimulus presentation as it will have an impact on the decibel level reaching the ear drum of these infants. PMID- 10519830 TI - Effects of Social versus Musical Antecedents on Communication Responsiveness in Five Children with Developmental Disabilities. AB - The present study involved a comparison of social versus musical antecedents on communication responsiveness in five children with developmental disabilities. During the social antecedent condition, the teacher presented opportunities for the children to greet, name objects, and request materials. In the musical antecedent condition, these same opportunities were embedded within a music/singing activity. A reversal design was used to compare the percentage of opportunities with appropriate communication responses across the two conditions. For three of the five children, the musical antecedent condition was associated with higher percentages of appropriate communication responses. For the other two children, the two conditions were associated with approximately equal rates of appropriate communication. Across both conditions, appropriate responses were more likely during opportunities for greeting and requesting than during opportunities to name objects. The results suggest that embedding communication opportunities within a musical activity may lead to increased appropriate communication responses for some children with developmental disabilities. PMID- 10519831 TI - The Contributions of Wayne Ruppenthal to the Field of Music Therapy. AB - This paper examines the career of Wayne Ruppenthal, considered one of the early pioneers in the field of music therapy. He began his practice in the late 1940s and his clinical accomplishments at Topeka State Hospital, spanned nearly two decades during the height of Freudian psychoanalysis and "milieu" therapy prescribed by The Menninger Foundation. Ruppenthal received his education at The University of Kansas in Lawrence, Kansas, and was the first graduate of the Master's of Music Education in Functional Music program in 1948. His contributions to the profession were significant and enduring and included establishing formal clinical practice and training standards, assisting with development of the National Association for Music Therapy (NAMT), and promoting the credibility of music therapy through published research. He retired from Topeka State Hospital in 1968. This paper is dedicated to the memory of Wayne Ruppenthal, who died on August 31, 1997. PMID- 10519832 TI - The Effect of Therapist and Activity Characteristics on the Purposeful Responses of Probable Alzheimer's Disease Participants. AB - The purpose of this study was to examine the efficacy of individual therapist sessions on the purposeful responses of probable Alzheimer's disease (AD) participants (hereby designated as "dementia participants". Three music therapists and one occupational therapist individually presented a combined total of 29 sing-along and exercise sessions to a group of seven participants, six having a diagnosis of probable Alzheimer's disease (moderate to severe cognitive decline), and one nondementia "control" participant, at an adult day care center specializing in dementia. Informed consent procedures were given to all study participants and their families. Sixteen sing-along sessions of 25-45 minutes duration consisted of live guitar playing and singing by individual music therapists. Thirteen exercises sessions of 20 minutes in length consisted of individual therapist (music or occupational) leading of exercises to a music tape ("Everyone Can Move" by Farnan & Johnson, 1988). Independent variables and their response definitions included: activity type (Sing-along, exercise); session content (music intervals, nonmusic intervals); therapist type (music, occupational); and therapist styles (singing, guitar playing). Participant dependent measures included singing/humming to songs, and movement/exercise to a therapist model (purposeful responses). All sessions were videotaped. Results indicated that the six dementia participants as a group purposefully responded significantly more during exercise than during sing-along sessions, but one of the six participants had extremely low exercise participation scores. That one participant's scores in sing-along sessions were in the range of the majority (4 of 5) of the other dementia participants' singing scores. Further analysis to help explain the discrepancy in this participant's purposeful responding indicated no significant difference in responding to individual music or occupational therapists during exercise activities, but a significant difference in individual responding to 3 different music therapists during the sing-along activities. Addi6onal analyses of the sing-along activities revealed significant differences in the total percentage of singing and guitar playing by the individual music therapists. Implications of the results for music therapy programming with dementia clients are discussed. PMID- 10519833 TI - Creativity in QualitativeMusic Therapy Research. PMID- 10519834 TI - Standards of Integrity for Qualitative Music Therapy Research. PMID- 10519835 TI - Embracing Complexity: The Creation of a Comprehensive Research Culture in Music Therapy. PMID- 10519836 TI - Science as Metacritique. PMID- 10519837 TI - Rhythmic Auditory Stimulation in Gait Training for Patients with Traumatic Brain Injury. AB - Rhythmic auditory stimulation (RAS) was studied in a frequency entrainment design and as a therapeutic stimulus to facilitate gait patterns in 8 traumatically brain injured individuals (5 male/3 female; mean age 30 +/- 5 years) with persisting gait disorder, 4-24 months postinjury. During entrainment, with RAS frequency matched to baseline cadence, velocity and stride symmetry both increased by an average of 18%. Increases contributing to the velocity improvement were seen in both stride length (7%) and cadence (8%). With RAS accelerated 5% over the fast walking step rate of the patients, 5 patients could entrain to a higher step frequency. The 2 patients with the slowest baseline gait velocity could not entrain to faster RAS frequencies. After 5 weeks of daily RAS training, 5 patients' mean velocity increased significantly (p /=80) and only one was severely disabled. Of the 48 with shunts only 20 were achievers (OR 11.2, 95% confidence interval (95% CI) 1.3-96.8). Lack of achievement in these 48 was associated with revisions of the shunt, particularly when revisions were performed after the age of 2. Sixteen patients had never required a revision and 11 (69%) were achievers; 10 had had revisions only during infancy and five (50%) were achievers; 22 had had revisions after their second birthday and only four (18%) were achievers (p<0.001). Elective revisions were not performed in this cohort and in 75% of patients revisions had been preceded by clear symptoms of raised intracranial pressure. CONCLUSION: Revisions of the shunt, particularly after the age of 2, are associated with poor long term achievement in adults with spina bifida. PMID- 10519864 TI - Comparative study between chemiluminescence assay and two different sensitive polymerase chain reactions on the diagnosis of serial herpes simplex virus encephalitis. AB - OBJECTIVE: A prospective study was undertaken on the diagnosis of herpes simplex encephalitis (HSVE) by comparing chemiluminescence assay (CL) and two different sensitive polymerase chain reactions (PCRs). METHODS: The materials comprised 53 serial CSF samples from 31 patients with acute encephalitis with suspected HSVE. Each CSF was distributed to three independent laboratories to perform quantitative measurements by CL, the low sensitive (single) PCR, and high sensitive (nested) PCR. The CL provided a method of detecting HSV itself and the small fragment with HSV antigenicity which was composed of viral component proteins. The serial CSFs were found retrospectively to comprise 24 samples from 11 patients with HSVE due to HSV1 and 29 samples from 20 patients with non-HSVE. RESULTS: the CL showed 50 to 48 000 pfu/ml in all samples of HSVE (except one) taken from the 3rd to the 25th day. The low sensitive PCR demonstrated 50 to 47 000 pfu/ml in only six samples of HSVE. The high sensitive PCR disclosed less than 100 to 120 000 copies/ml in 11 samples of HSVE. At the acute stage from the 1st to 7th day, the sensitivities of CL and the high sensitive PCR were 100%, but that of the low sensitive PCR was 75%. The sensitivity of CL was significantly higher than those of both PCRs after the acute stage on the 15th to 32nd day. The specificities and positive predictive values of the three methods were 100%. However, the negative predictive value of CL was significantly higher than that of the low sensitive PCR. CONCLUSIONS: The sensitivity of CL is equivalent to that of the high sensitive PCR during the acute stage and significantly higher than that of the high sensitive PCR after the acute stage. A clear difference in sensitivity exists between the different PCRs. A combination of the PCR, chemiluminescence assay, and serological antibody diagnosis is currently considered the most effective approach for the clinical diagnosis of HSVE. PMID- 10519865 TI - Surgery for suspected neurogenic thoracic outlet syndromes: a follow up study. AB - OBJECTIVES: To assess the outcome of surgical treatment for thoracic outlet syndrome (TOS), and to compare the outcome in patients with and without an underlying cervical rib. METHODS: a heterogeneous group of 40 patients (33 women, seven men; aged 22-62 years) were evaluated 3 months to 20 years after surgery for suspected neurogenic TOS. Forty nine operations had been performed: cervical ribs were removed in 23 patients, together with fibrous band excision in nine. In the 17 without a cervical rib the thoracic outlet was decompressed by resection of the first thoracic rib in nine, and by other operations in eight. RESULTS: After surgery patients reported improved pain (33/36), sensory disturbance (30/35), hand muscle strength (14/27), and hand function (23/34). Postoperatively TOS recurred in two, and symptoms continued to progress in three patients in whom other diagnoses eventually emerged. Surgical complications were recorded in 10 patients, but were transient and did not result in permanent symptomatic sequelae. CONCLUSIONS: Surgical treatment of suspected neurogenic TOS relieves pain and sensory disturbance (90%), but is less effective for muscle weakness (50%). Surprisingly, surgery relieved sensory and motor abnormalities to a similar degree in patients both with and without a cervical rib. Ideally, patients require early operation to forestall permanent hand muscle denervation, but, our retrospective analysis fails to identify any single preoperative diagnostic criterion for TOS, particularly in patients lacking a radiographic cervical rib. PMID- 10519866 TI - Progressive polyradiculoneuropathy in diabetes: correlation of variables and clinical outcome after immunotherapy. AB - OBJECTIVE: To quantify the progression of diabetic polyradiculoneuropathy-a condition in which immune factors have been implicated-after immunotherapy. METHODS: The study evaluated 15 consecutive patients with this condition. All patients were older than 40. Four had type I diabetes and six were women. The duration of pre-existing diabetes varied from 2 to 20 years. The clinical presentation was dominated by painful progressive motor weakness, with or without exacerbation of sensory symptoms. The weakness involved all limbs, but was often asymmetric. RESULTS: Electrophysiological testing showed a predominantly axonal polyneuropathy, with more recent denervating polyradiculopathy. Analysis of CSF showed increased protein in 14 and oligoclonal bands in five. Quantitative autonomic tests showed abnormalities in all patients. Sural nerve biopsy was performed in 14 patients; all showed fibre loss and segmental demyelination, four had occasional onion bulbs, and 10 showed various inflammatory infiltrates. After immunomodulating therapy, there was no further deterioration and clinical improvement occurred in all patients. Sweat responses, cardiovascular reflexes, and sural nerve fibre density correlated best with functional outcome. There was no significant difference between plasmapheresis and intravenous gammaglobulin. CONCLUSION: Immunotherapy may improve this condition, but only certain variables correlate with rapid therapeutic response. PMID- 10519867 TI - Risk factors for spread of primary adult onset blepharospasm: a multicentre investigation of the Italian movement disorders study group. AB - OBJECTIVES: Little is known about factors influencing the spread of blepharospasm to other body parts. An investigation was carried out to deterrmine whether demographic features (sex, age at blepharospasm onset), putative risk, or protective factors for blepharospasm (family history of dystonia or tremor, previous head or face trauma with loss of consciousness, ocular diseases, and cigarette smoking), age related diseases (diabetes, hypertension), edentulousness, and neck or trunk trauma preceding the onset of blepharospasm could distinguish patients with blepharospasm who had spread of dystonia from those who did not. METHODS: 159 outpatients presenting initially with blepharospasm were selected in 16 Italian Institutions. There were 104 patients with focal blepharospasm (mean duration of disease 5.3 (SD 1.9) years) and 55 patients in whom segmental or multifocal dystonia developed (mainly in the cranial cervical area) 1.5 (1.2) years after the onset of blepharospasm. Information was obtained from a standardised questionnaire administered by medical interviewers. A Cox regression model was used to examine the relation between the investigated variables and spread. RESULTS: Previous head or face trauma with loss of consciousness, age at the onset of blepharospasm, and female sex were independently associated with an increased risk of spread. A significant association was not found between spread of dystonia and previous ocular diseases, hypertension, diabetes, neck or trunk trauma, edentulousness, cigarette smoking, and family history of dystonia or tremor. An unsatisfactory study power negatively influenced the validity and accuracy of the negative findings relative to diabetes, neck or trunk trauma, and cigarette smoking. CONCLUSIONS: The results of this exploratory study confirm that patients presenting initially with blepharospasm are most likely to experience some spread of dystonia within a few years of the onset of blepharospasm and suggest that head or face trauma with loss of consciousness preceding the onset, age at onset, and female sex may be relevant to spread. The suggested association between edentulousness and cranial cervical dystonia may be apparent because of the confounding effect of both age at onset and head or face trauma with loss of consciousness. The lack of influence of family history of dystonia on spread is consistent with previous findings indicating that the inheritance pattern is the same for focal and segmental blepharospasm. PMID- 10519868 TI - Time course of symptomatic orthostatic hypotension and urinary incontinence in patients with postmortem confirmed parkinsonian syndromes: a clinicopathological study. AB - OBJECTIVE: Although both orthostatic hypotension and urinary incontinence have been reported in a number of parkinsonian syndromes, such as Parkinson's disease (PD), multiple system atrophy (MSA), dementia with Lewy bodies (DLB), corticobasal degeneration (CBD), and progressive supranuclear palsy (PSP), differences in the evolution of these features have not been studied systematically in pathologically confirmed cases. METHODS: 77 cases with pathologically confirmed parkinsonian syndromes (PD, n=11; MSA, n=15; DLB, n=14; CBD, n=13; PSP, n=24), collected up to 1994, formed the basis for a multicentre clinicopathological study organised by the NINDS to improve the differential diagnosis of parkinsonian disorders. The present study determined the time course that is, latency to onset and duration from onset to death, of symptomatic orthostatic hypotension, and urinary incontinence in the NINDS series. Furthermore, the diagnostic validity of a predefined latency to onset within 1 year of disease onset of symptomatic orthostatic hypotension or urinary incontinence was analysed. RESULTS: Significant group differences for latency, but not duration, of symptomatic orthostatic hypotension and urinary incontinence were found. Latencies to onset of either feature were short in patients with MSA, intermediate in patients with DLB, CBD, and PSP, and long in those with PD. Symptomatic orthostatic hypotension occurring within the first year after disease onset predicted MSA in 75% of cases; early urinary incontinence was less predictive for MSA (56%). CONCLUSION: Latency to onset, but not duration, of symptomatic orthostatic hypotension or urinary incontinence differentiates PD from other parkinsonian syndromes, particularly MSA. PMID- 10519869 TI - Bradykinesia akinesia inco-ordination test (BRAIN TEST): an objective computerised assessment of upper limb motor function. AB - OBJECTIVES: A simple and rapid computerised keyboard test, based on the alternating finger tapping test, has been developed to quantify upper limb motor function. The test generates several variables: (1) kinesia score: the number of keystrokes in 60 seconds; (2) akinesia time: cumulative time that keys are depressed; (3) dysmetria score: a weighted index calculated using the number of incorrectly hit keys corrected for speed; (4) incoordination score: a measure of rhythmicity which corresponds to the variance of the time interval between keystrokes. METHODS: The BRAIN TEST(Copyright ) was assessed on 35 patients with idiopathic Parkinson's disease, 12 patients with cerebellar dysfunction, and 27 normal control subjects. RESULTS: The mean kinesia scores of patients with Parkinson's disease or cerebellar dysfunction were significantly slower than normal controls (Parkinson's disease=107 (SD 28) keys/min v cerebellar dysfunction=86+/- (SD 28) v normal controls=182 (SD 26), p<0.001) and correlated with the UPDRS (r =-0.69, p<0.001). The akinesia time is very insensitive and was only abnormal in patients with severe parkinsonism. The median dysmetria (cerebellar dysfunction=13.8 v Parkinson's disease=6.1 v normal controls=4.2, p=0.002) and inco-ordination scores (cerebellar dysfunction=5.12 v Parkinson's disease=0.84 v normal controls=0.15, p=0.002) were significantly higher in patients with cerebellar dysfunction, in whom the dysmetria score correlated with a cerebellar disease rating scale (r=0.64, p=0.02). CONCLUSION: The BRAIN TEST(Copyright ) provides a simple, rapid, and objective assessment of upper limb motor function. It assesses speed, accuracy, and rhythmicity of upper limb movements regardless of their physiological basis. The results of the test correlate well with clinical rating scales in Parkinson's disease and cerebellar dysfunction. The BRAIN test will be useful in clinical studies. It can be downloaded from the Internet (). PMID- 10519870 TI - Blink reflex R2 changes and localisation of lesions in the lower brainstem (Wallenberg's syndrome): an electrophysiological and MRI study. AB - OBJECTIVES: Pathways of late blink reflexes are detected by high resolution MRI. Electronically matched stroke lesions superimposed to an anatomical atlas show the suspected course. METHODS: Fifteen patients with infarction of the lower brainstem, MRI lesions and electrically elicited blink reflexes were examined. The involved structures in patients with R2 and R2c blink reflex changes were identified by biplane high resolution MRI with individual slices matched to an anatomical atlas at 10 different levels using digital postprocessing methods. RESULTS: The blink reflexes were normal in five of 15 patients (33%) and showed loss or delay of R2 and R2c to stimulation ipsilaterally to lesion (R2-i and R2c i) in eight (53%). Loss or delay of R2-i/R2c-i was seen in lesions covering the entire trigeminal spinal tract and nucleus (TSTN) at at least one level. These infarctions were located more dorsally within the medulla. Patients with normal blink reflexes showed lesions sparing or involving the TSTN only partially. They more often had incomplete Wallenberg's syndromes and MRI lesions were located more ventrally. CONCLUSIONS: Using digital postprocessing MRI methods it was possible to identify central pathways of late blink reflex in patients with Wallenberg's syndrome. This method is suggested as a new approach to identify incompletely understood functional structures of the brainstem. PMID- 10519872 TI - Re-emergent tremor of Parkinson's disease. AB - OBJECTIVE: To characterise postural tremors in patients with Parkinson's disease. Rest tremor is a well recognised cardinal symptom of Parkinson's disease, but postural tremors associated with the disease may cause more disability than the more typical rest tremor. Postural tremor of Parkinson's disease has been attributed to enhanced physiological tremor, clonus, or coexistent essential tremor. It is postulated that one type of postural tremor in Parkinson's disease represents a rest tremor that re-emerges after a variable delay while maintaining posture, hence "re-emergent tremor". METHODS: Accelerometry, peak frequency, peak frequency amplitude, root mean square (RMS) amplitude, and latency were determined in 18 patients (mean age: 63.2 (SD 9.8) years) with Parkinson's disease who had clinically evident postural tremor, 20 (mean age: 66.9 (SD 5. 8) years) with typical essential tremor, and seven (mean age: 68.7 (SD 15.3) years) with the combination of pre-existing essential tremor and subsequent Parkinson's disease (essential tremor/Parkinson's disease). Latency, the time interval starting with the assumption of an outstretched posture and ending with the onset of postural tremor, was measured by marking the start time by a pulse produced from interrupting a beam to a photocell when the arm reached a horizontal position. RESULTS: The latency for the re-emergent tremor (9.37 (SD 10.66) s), present in 12 of 18 patients with Parkinson's disease, was significantly (p<0.0005) longer than the latency for postural tremor of essential tremor (1.29 s in one patient, absent in 19 others); five of seven essential tremor/Parkinson's disease patients had an observed latency (6.57 (SD 8.23 s) which was also significantly (p<0.005) longer than that for essential tremor. There was no difference in the mean tremor frequency ( approximately 5.5 Hz) between the re-emergent tremor and the more typical Parkinson's disease rest tremor. The amplitudes were generally higher for the postural tremor associated with Parkinson's disease compared with those of essential tremor. CONCLUSION: These studies suggest that the re-emergent tremor of Parkinson's disease can be differentiated from the postural tremor of essential tremor and that it may share pathophysiological mechanisms with the more typical rest tremor. PMID- 10519871 TI - Unilateral focal lesions in the rostrolateral medulla influence chemosensitivity and breathing measured during wakefulness, sleep, and exercise. AB - OBJECTIVES: The rostrolateral medulla (RLM) has been identified in animals as an important site of chemosensitivity; in humans such site(s) have not been defined. The aim of this study was to investigate the physiological implications of unilateral lesions in the lower brainstem on the control of breathing. METHODS: In 15 patients breathing was measured awake at rest, asleep, during exercise, and during CO(2) stimulation. The lesions were located clinically and by MRI; in nine patients they involved the RLM (RLM group), in six they were in the pons, cerebellum, or medial medulla (Non-RLM group). All RLM group patients, and three non-RLM group patients had ipsilateral Horner's syndrome. RESULTS: Six of the RLM group had a ventilatory sensitivity to inhaled CO(2) (V/P(ET) CO(2)) below normal (group A: V/P(ET) CO(2), mean, 0.87; range 0.3-1.4 l. min(-1)/mm Hg). It was normal in all of the non-RLM group (group B: V/P(ET) CO(2), mean, 3.0; range, 2.6 3.9 min(-1)/mmHg). There was no significant difference in breathing between groups during relaxed wakefulness (V, group A: 7.44 (SD 2.5) l.min(-1); group B: 6.02 (SD 1.3) l.min(-1); P(ET) CO(2), group A: 41.0 (SD 4.2) mm g; group B: 38.3 (SD2.0) mm Hg) or during exercise (V/VO(2): group A: 21 (SD 6. 0) l.min( 1)/l.min(-1); group B: 24 (SD 7.3) l.min(-1)/l.min(-1)). During sleep, all group A had fragmented sleep compared with only one patient in group B (group A: arousals, range 13 to > 60 events/hour); moreover, in group A there was a high incidence of obstructive sleep apnoea associated with hypoxaemia. CONCLUSION: Patients with unilateral RLM lesions require monitoring during sleep to diagnose any sleep apnoea. The finding that unilateral RLM lesions reduce ventilatory sensitivity to inhaled CO(2) is consistent with animal studies. The reduced chemosensitivity had a minimal effect on breathing awake at rest or during exercise. PMID- 10519873 TI - Can stroke physicians and neuroradiologists identify signs of early cerebral infarction on CT? AB - Doctors managing acute stroke are expected to recognise signs of early infarction on CT before choosing thrombolytic treatment, according to recent trials and guidelines. The ability of 13 physicians and two neuroradiologists to recognise early infarct signs and decide whether patients should be randomised in a hypothetical stroke treatment trial was tested. Only 65% of the CT scans from 14 stroke patients were correctly identified as normal or abnormal (95% CI 60-69%). Neither observer experience nor knowledge of symptoms significantly improved recognition of abnormality, although experience did significantly improve the observers' ability to reproduce their results. Parenchymal hypodensity was the least well recognised sign. Only 45% (95% CI 40%-50%) of patients were identified correctly for the hypothetical acute stroke treatment trial. Early infarction on CT is not well recognised even by experienced doctors. Part of the problem may be in understanding the definitions of the extent of infarction. These difficulties should be considered in the design of acute stroke treatment trials and in the introduction of any new acute stroke treatments. PMID- 10519874 TI - Inverse relation between Braak stage and cerebrovascular pathology in Alzheimer predominant dementia. AB - The most common neuropathological substrates of dementia are Alzheimer's disease, cerebrovascular disease, and dementia with Lewy bodies. A preliminary, retrospective postmortem analysis was performed of the relative burden of each pathology in 25 patients with predominantly Alzheimer's disease-type dementia. Log linear modelling was used to assess the relations between ApoE genotype, Alzheimer's disease, and cerebrovascular disease pathology scores. Sixteen of 18 cases (89%) with a Braak neuritic pathology score 24 hours), preeclampsia, and the use of tocolytic drugs, antenatal exposure to betamethasone was associated with a lower risk of cystic periventricular leukomalacia than was either the absence of glucocorticoid therapy (adjusted odds ratio, 0.5; 95 percent confidence interval, 0.2 to 0.9) or exposure to dexamethasone (adjusted odds ratio, 0.3; 95 percent confidence interval, 0.1 to 0.7). The adjusted odds ratio for the group of infants whose mothers had received dexamethasone as compared with the group of infants whose mothers had not received a glucocorticoid was 1.5 (95 percent confidence interval, 0.8 to 2.9). CONCLUSIONS: Antenatal exposure to betamethasone but not dexamethasone is associated with a decreased risk of cystic periventricular leukomalacia among very premature infants. PMID- 10519897 TI - Images in clinical medicine. Tuberculous meningitis. PMID- 10519899 TI - Growth hormone therapy in adults and children. PMID- 10519898 TI - Racial differences in the treatment of early-stage lung cancer. AB - BACKGROUND: If discovered at an early stage, non-small-cell lung cancer is potentially curable by surgical resection. However, two disparities have been noted between black patients and white patients with this disease. Blacks are less likely to receive surgical treatment than whites, and they are likely to die sooner than whites. We undertook a population-based study to estimate the disparity in the rates of surgical treatment and to evaluate the extent to which this disparity is associated with differences in overall survival. METHODS: We studied all black patients and white patients 65 years of age or older who were given a diagnosis of resectable non-small-cell lung cancer (stage I or II) between 1985 and 1993 and who resided in 1 of the 10 study areas of the Surveillance, Epidemiology, and End Results (SEER) program (10,984 patients). Data on the diagnosis, stage of disease, treatment, and demographic characteristics of the patients were obtained from the SEER data base. Information on coexisting illnesses, type of Medicare coverage, and survival was obtained from linked Medicare inpatient-discharge records. RESULTS: The rate of surgery was 12.7 percentage points lower for black patients than for white patients (64.0 percent vs. 76.7 percent, P<0.001), and the five-year survival rate was also lower for blacks (26.4 percent vs. 34.1 percent, P<0.001). However, among the patients undergoing surgery, survival was similar for the two racial groups, as it was among those who did not undergo surgery. Furthermore, analyses in which adjustments were made for factors that are predictive of either candidacy for surgery or survival did not alter the influence of race on these outcomes. CONCLUSIONS: Our analyses suggest that the lower survival rate among black patients with early-stage, non-small-cell lung cancer, as compared with white patients, is largely explained by the lower rate of surgical treatment among blacks. Efforts to increase the rate of surgical treatment for black patients appear to be a promising way of improving survival in this group. PMID- 10519900 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 31-1999. A 33-year-old man with wide-complex tachycardia and a left ventricular mass. PMID- 10519901 TI - Exogenous reinfection in tuberculosis. PMID- 10519902 TI - Immune reconstitution. PMID- 10519903 TI - Periventricular leukomalacia--prospects for prevention. PMID- 10519904 TI - Racial disparity in rates of surgery for lung cancer. PMID- 10519905 TI - Beta-peptides: twisting and turning. AB - Oligomers of beta-amino acids (beta-peptides), which are readily available by standard meth ods either in solution or on solid support, adopt a large variety of different secondary structures in solution and in the solid state. beta Peptides 4, 5 and 10 fold into a helix with 3 residues per turn and 14-membered H bonded rings (314 helix) that is left-handed for 5 and 10 and right-handed for 2 (due to the reversal of the chirality of the building blocks), as was clearly demonstrated by two-dimensional NMR-spectroscopy. This helix thermally is very stable in methanol solution upon heating. As shown by NMR- and CD-spectroscopy, it is partially populated even at 100 C (Figure 3). Another helix was dis covered for mixed beta-peptide 8 in methanol solution: it is characterized by 12- and 10- membered turns (Figure 4, left) and its central 10-membered turn has been found in the solid state of a geminally disubtituted beta-peptide (Figure 4, right). This central 10-membered turn was used as a scaffold to attach beta-amino acid residues that prefer a linear (non-helical) conformation (beta-peptide 21): a hairpin (pleated sheet-turn-pleated sheet) structure was determined in solution by NMR-spectroscopy (Figure 5). In contrast to this antiparallel pleated-sheet, a parallel pleated sheet was found for a beta-tripeptide in the solid state. For the first time it was possible to observe reversible peptide folding in MD simulations by studying beta-peptides (Figure 6) and to determine folding pathways and intermediates. beta-Peptides are a new class of promising peptidomimetics. They are resistant against the degradation by proteolytic enzymes such as pepsin, elastase, carboxypeptidase A, pro nase or proteasom 20S. A variety of beta-amino acids (27-34) was shown to be non- mutagenic by Ames tests and beta-peptides 47 and 48 reveal large elimination half-lives of 3 h (for 47) and 10 h (for 48) in the serum of rodents (Figure 7). Conjugates of alpha- and beta- peptides are efficient ligands for the HLA*B27 MHC Class I protein, a five fold increase of binding (2.0 microM for 55) compared to a natural peptidic ligand 51 was observed. Furthermore, beta-peptides are able to mimic natural a peptidic hormones such as somatostatin. The cyclo-beta-tetrapeptide 57 binds to the five human somatostatin receptors in the micromolar range. In addition, several other non-natural oligomers such as beta-peptide nucleic acids (built from 58 and 59), beta-peptoids (60), oligomers of anthranilic acids and beta sulfonamido peptides are discussed. PMID- 10519906 TI - Recent advances in the medicinal chemistry of taxoids with novel beta-amino acid side chains. AB - Beta-Amino acids have been recognized as an important class of compounds in the design and synthesis of potential pharmaceutical drugs and also for the study of enzymatic reaction mechanisms. Among the beta-amino acid family, isoserines (alpha-hydroxy-beta-amino acids) are probably the most important members because many of them are potent enzyme inhibitors and they also serve as essential building blocks for biologically and medicinally important molecules such as Taxol. Taxol (paclitaxel) and Taxotere (docetaxel) are currently considered to be the most important drugs in cancer chemotherapy. This review describes recent advances in the chemistry of isoserines and taxoid anticancer agents at the biomedical interface including (i) the development of highly efficient method for the synthesis of isoserine side chains of taxoids and (ii) the synthesis and structure-activity relationship (SAR) study of taxoids featuring discovery and development of the "second generation" taxoid anticancer agents that possess exceptional activities against drug-resistant cancer cells. PMID- 10519907 TI - Chemical process synthesis of beta-amino acids and esters. AB - The importance of beta-amino acids and esters for the synthesis of potential therapeutic agents and biologically active compounds is well known and the subject of this special issue. This review outlines some of the recent approaches reported for the synthesis of both racemic and enantiomeric beta-amino acids and esters with emphasis on those used for large scale production. This compilation is written from a chemical process perspective focusing on the practical application of these processes for large scale synthesis. A survey of procedures described in patent publications as well as current advances from chemical process research groups and results from our laboratory are included with emphasis on asymmetric Michael-type additions, addition of metal enolates to imines/sulfinimines, classical and enzymatic resolutions, and reduction of enantiomeric enamines. PMID- 10519908 TI - Structure-activity relationships of beta-amino acid-containing integrin antagonists. AB - Interest in the development of specific antagonists of the beta3 family of integrins (platelet alphaIIbbeta3 and the vitronectin receptor alphavbeta3) has been principally driven by efforts to design more potent antithrombotic agents than either aspirin or the thienopyridine-type ADP receptor modulators. The platelet fibrinogen receptor (aIIbb3) and the vitronectin receptor (alphavbeta3) bind the RGD tripeptide sequence found within adhesive ligands. Because of this, many approaches to antagonists of beta3 receptors have utilized an RGD mimetic to identify antagonists. Integrin antagonists of many structurally diverse classes have been discovered. One of the larger beta3 integrin antagonist classes employs beta-amino acids to mimic the aspartate residue of the RGD mimetic. Structure activity investigations have revealed the potent activity of agents which have substituents appended to both the alpha and beta position of the beta-amino acid units of these antagonists. Several clinical candidates targeting platelet aIIbb3 contain these beta-amino acid units and are currently being evaluated clinically. PMID- 10519909 TI - Recent advances in the enantioselective synthesis of beta-amino acids. AB - The introductory section of this review presents some of the currently most compelling beta-amino acid targets, according to their structural types: alpha- and beta-aryl substituted, olefinic and alkynyl, alpha, alpha- and alpha,beta disubstituted, cyclic and conformationally restricted, fluorine-containing, and phosphonic analogous beta-amino acids. The main section highlights some of the very new (1996-1998), promising methodology for the enantioselective synthesis of beta-amino acids, with especial emphasis on catalytic and enzymatic processes, as well as methods based on "chiral pool", self-regeneration of stereogenic centers , diastereoselective nucleophilic additions to prochiral double bonds, and enantioselective reactions in the presence of chiral additives. PMID- 10519910 TI - Glucagon-like peptide-1, a gastrointestinal hormone with a pharmaceutical potential. AB - Glucagon-like peptide-1 (GLP-1) is an insulinotropic hormone secreted from endocrine cells in the gut mucosa in response to meal ingestion. It is an important incretin hormone; mice with a null mutation in the GLP-1 receptor gene develop glucose intolerance. In addition, it inhibits gastrointestinal secretion and motility and is thought to be part of the "ileal brake" mechanism. Perhaps because of the latter actions it inhibits food intake, but intracerebral injection of GLP-1 also inhibits food intake. The insulinotropic effect is preserved in patients with type 2 diabetes mellitus, in whom also glucagon secretion is inhibited. Thus upon i.v. GLP-1 infusion blood glucose may be completely normalised. Because its actions are glucose-dependent hypoglycaemia does not develop. However, GLP-1 is metabolised extremely rapidly in vivo, initially by a mechanism that involves the enzyme dipeptidyl peptidase-IV. It is currently being investigated how GLP-1 or analogues thereof can be employed in practical diabetes therapy. Promising solutions include the development of stable analogues and inhibitors of the degrading enzyme. PMID- 10519911 TI - Pharmaceutical VIP: prospects and problems. AB - Recently, multiple receptors for vasoactive intestinal peptide (VIP) have been molecularly cloned and our understanding of VIP chemistry and mechanisms of action has been broadened. The following review outlines the physiological effects of the hormone from growth regulation, reproduction, bronchodilation, vasodilation and immune interactions to neurotrophism. VIP-based drug design and non-invasive innovative delivery modes are discussed with emphasis on tumor diagnosis and treatment, impotence treatment and neuroprotection. PMID- 10519912 TI - Actions of CRF and its analogs. AB - Corticotropin-releasing factor (CRF), urocortin, sauvagine and urotensin I form the CRF family. These peptides bind with different affinities to two subtypes of CRF receptor (CRFR), CRFR1 and CRFR2. The latter exists as two splice variants, the neuronal CRFR2a and the peripheral CRFR2b. CRFR is a G protein-dependent receptor which acts mainly through Gs enhancing cAMP production. However, CRFR1 expressed in neutrophils of the spleen in response to immunologic stimulation and psychological stress does not seem to function through Gs, as indicated by the inability of CRF to stimulate the cAMP production of CRFR1+ neutrophils. Besides the two receptors, a 37 kD CRF binding protein (CRF-BP) binds several CRF peptides with high affinity. CRFR and CRF-BP do not share a common amino acid sequence representing the ligand binding site. In view of the unusually slow offrate of CRF-BP, it is proposed that CRF-BP provides an efficient uptake of free extracellular CRF. Thus, the time of exposure of CRFR to CRF or urocortin can be limited. At this time, the fate of the ligand CRF-BP complex is unclear. CRFR1 is not only involved in the hypophyseal stimulation of corticotropin release, but hippocampal CRFR1 mediates enhancement of stress-induced learning. CRFR1 may also be involved in basic anxiety. In contrast, at least in the mouse, CRFR2 of the lateral intermediate septum mediates tonic impairment of learning. In response to stressful stimuli or after local injection of high CRF doses, CRFR2 mediates anxiety. Effects requiring CRFR2 can be blocked specifically by the recently developed peptidic antagonist antisauvagine-30. PMID- 10519913 TI - Molecular characterization of the ligand-receptor interaction of neuropeptide Y. AB - Neuropeptide Y (NPY) consists of 36 amino acids and is one of the most abundant peptides in the peripheral and central nervous system. Several subtypes of NPY receptors have been described (Y1- y6) using segments and analogues of NPY. The Y1-, Y2- and the Y5-receptor, which have been cloned, belong to the G-protein coupled hormone receptor family and will be specially addressed, because they are the endogenous binding sites of neuropeptide Y in human. In contrast, Y4 receptors recognize endogenous PP, Y3 receptors are discussed controversially and the y6-receptor is truncated in human. In this review, we summarize the data of neuropeptide Y with respect to ligand binding, selectivity, receptor structures and ligand-receptor complexes by using ligand analogues, site directed mutagenesis and photoaffinity labeling. PMID- 10519914 TI - Calcitonin. AB - The peptide calcitonin (CT) was initially discovered in 1962 as a novel hypocalcemic hormone. This hypocalcemic response was principally due to a potent inhibitory action of CT on osteoclast mediated bone resorption and it is this action which underlies its widespread clinical use for the treatment of bone disorders, including Paget's disease, osteoporosis and hypercalcemia of malignancy. In this article we review the basic physiology of CT action, structure-function studies on CT peptides, cloning of CT receptors and the identification of isoforms of the receptor derived from alterative splicing of the receptor mRNA. We also review the state of understanding on CT receptor mediated signaling and receptor regulation, along with developing concepts of how CT peptides interact with the receptor, including how the receptors may interact with receptor activity modifying proteins to produce novel phenotypes. Finally, current therapeutic use is reviewed, and the potential for expanded use that may come with advances in delivery of peptides or CT mimetics. PMID- 10519915 TI - Design and applications of parathyroid hormone analogues. AB - The endocrine parathyroid hormone (PTH) is the major regulator of serum calcium levels. In contrast, the autocrine/paracrine parathyroid hormone-related peptide (PTHrP) has been associated with organism development. Both are secreted as much larger molecules but have their major functions associated with their N-terminal 34 residues. They share a common receptor expressed in organs critical to PTH function - bone, kidney, and intestine. PTH and PTHrP receptor activation stimulates adenylyl cyclase (AC), phospholipase C (PLC), and phospholipase D (PLD) in target cells. It has been possible to separate the AC-stimulation from that of PLC. AC-stimulation requires at least the N-terminal 28 residues of PTH and PLC-stimulation requires a minimum of residues 29-32-NH2. Intermittent administration of PTH stimulates bone growth and requires AC-stimulation. The shortest linear sequence of hPTH with essentially full anabolic activity for bone growth-stimulation is hPTH(1-31)NH2. Two applications are postulated for PTH and PTHrP-based pharmaceuticals - treatment of bone loss due to osteoporosis and reversal of the hypercalcemic effect of malignancy. PTHrP analogues which strongly inhibit PTHrP AC-stimulation showed promise for the treatment of malignancy-associated hypercalcemia in animal trials but failed in human ones. However, both animal and human trials of hPTH have shown significant bone growth stimulating effects. New deletion, substitution and cyclized analogues of PTH show great promise both for greater in vitro activity and possibly for improved delivery and greater specificity as agents for restoration of bone loss in osteoporosis. PMID- 10519916 TI - Chemistry and structure activity relationships of bafilomycin A1, a potent and selective inhibitor of the vacuolar H+-ATPase. AB - Bafilomycin A1, a macrolide antibiotic isolated from the fermentation of Streptomyces spp., is a potent and selective inhibitor of vacuolar-type proton translocating ATP-ases (V-ATPases) and was used to study the physiological role of this class of enzymes. An extensive chemical effort on the unusual structure of this macrolide led to the synthesis of significantly different bafilomycin derivatives. None of the new analogues was more potent than the parent compound but provided a significant amount of information about the structural requirements for the inhibitory activity of bafilomycin A1 in particular on chicken osteoclast (cOc) ATPase. The vinylic methoxy group adjacent to a carbonyl function, the dienic system and the hydroxy group at position 7 were recognized to be essential features for bafilomycin V-ATPase-inhibitory activity. This information was utilized to design simplified novel derivatives as inhibitors of bone resorption. PMID- 10519917 TI - Steroidal antiandrogens and 5alpha-reductase inhibitors. AB - The purpose of this work is to synthesize a pregnane derivative with a high antiandrogenic effect or a high inhibitory activity for the enzyme 5 alpha reductase type 2. Benign prostatic hyperplasia and prostate cancer are androgen dependent diseases which afflict a large percentage of the male population. Dihydrotestosterone 3, a 5 alpha-reductase metabolite of testosterone 2 has been implicated as a causative factor in the progression of these diseases, largely through the clinical evaluation of males who are genetically deficient of steroid 5 alpha-reductase enzyme. As a result of this study, the inhibition of this enzyme has become a pharmacological strategy for the design and synthesis of new drugs. The advent of finasteride 22 "figure 5" a 5 alpha-reductase inhibitor, has greatly alleviated the symptoms associated with benign prostatic hyperplasia. On the other hand, the discovery of cyproterone acetate 4 "figure 2" alone or in combination with the antiandrogens flutamide 14 "figure 3" or bicalutamide 21 has greatly reduced the misery of prostate cancer. Prostate cancer kills about 40,000 men in the USA and approximately 400,000 prostatectomies are performed each year. In our laboratory we have recently synthesized ten new progesterone derivatives 17 alpha-acyloyloxy-6-halo (chloro, bromo) 16 beta-methyl-4, 6-pregnadiene-3, 20 diones (54a-54e and 55a-55e), "figure 10". These steroids were evaluated as antiandrogens and exhibited a much higher activity than the commercially available cyproterone acetate 4. The same compounds were also evaluated as 5 alpha-reductase inhibitors and showed a slightly higher inhibitory activity than that of finasteride 22, the drug of choice today for the treatment of benign prostatic hyperplasia In another study we synthesized several new 4-halo (bromo and chloro) 17 alpha-benzoyloxy and also 4-halo-17 alpha-acetoxy progesterone derivatives (58-63) "figure 13". These compounds were prepared from the commercially available 17 alpha-acetoxy progesterone 56. The pharmacological evaluation of these steroids "figure 14" indicated that the 17 alpha-benzoyloxy derivatives (4-chloro and bromo) 62 and 63 were very potent antiandrogens. On the other hand, the 4-halo (bromo and chloro) 17 alpha-acetoxy (58, 59) and the 17 alpha-benzoyloxy-4-chloro analog 63 showed a very high inhibitory activity for the enzyme 5 alpha-reductase type 2 "figure 15". PMID- 10519919 TI - Systemic antifungal agents against AIDS-related opportunistic infections: current status and emerging drugs in development. AB - No effective drug was available for the treatment of systemic fungal infections until the discovery of Amphotericin B in 1953. Since then flucytosine, azoles and later the triazoles, have now become available. The current interest in the development of new antifungal agents can partially be explained by the dramatic rise in the number of AIDS cases and the subsequent suppression of the immune system in patients with the disease. For example, over 90% of those diagnosed to be HIV-positive contract a fungal infection during the course of their illness. Other conditions that have spurred the development of new systemic antifungal agents include the increase in the frequency of bone marrow and organ transplants, the use of antineoplastic agents, long-term use of corticosteroids and even the indiscriminate use of antibiotics. The emergence of fungi resistant to currently available agents, especially the azoles, has made the need for new and effective antifungal agents more urgent. This review article focuses on agents targeted against opportunistic fungal infections, i.e., fungal infections which, in contrast to immunocompetent individuals, may cause serious life threatening illness in immunocompromised individuals. Agents currently on the market or undergoing clinical development, as well as potential new agents that have been discovered, are discussed. PMID- 10519918 TI - Bioactivities of chalcones. AB - This review outlines the different bioactivities of a variety of chalcones. The cytotoxic, anticancer, chemopreventative and mutagenic properties of a number of chalcones are described followed by an account of various of these unsaturated ketones as antimicrobial agents. The antiviral, antiprotozoal and insecticidal activities of a variety of chalcones are reviewed as well as the enzyme inhibitory properties and miscellaneous activities of some of these molecules. PMID- 10519923 TI - Morphological priming in the German mental lexicon. AB - We present results from cross-modal priming experiments on German participles and noun plurals. The experiments produced parallel results for both inflectional systems. Regular inflection exhibits full priming whereas irregularly inflected word forms show only partial priming: after hearing regularly inflected words (-t participles and -s plurals), lexical decision times on morphologically related word forms (presented visually) were similar to reaction times for a base-line condition in which prime and target were identical, but significantly shorter than in a control condition where prime and target were unrelated. In contrast, prior presentation of irregular words (-n participles and -er plurals) led to significantly longer response times on morphologically related word forms than the prior presentation of the target itself. Hence, there are clear priming differences between regularly and irregularly inflected German words. We compare the findings on German with experimental results on regular and irregular inflection in English and Italian, and discuss theoretical implications for single versus dual-mechanism models of inflection. PMID- 10519924 TI - Goal attribution without agency cues: the perception of 'pure reason' in infancy. AB - The proper domain of naive psychological reasoning is human action and human mental states but such reasoning is frequently applied to non-human phenomena as well. The studies reported in this paper test the validity of the currently widespread belief that this tendency is rooted in the fact that naive psychological reasoning is initially restricted to, and triggered by, the perception of self-initiated movement of agents. We report three habituation experiments which examine the necessary conditions under which infants invoke a psychological principle, namely the principle of rational action, to interpret behaviour as goal directed action. Experiment 1 revealed that the principle of rational action already operates at 9 (but not yet at 6) months of age. Experiment 2 demonstrated that perceptual cues indicating agency, such as self propulsion, are not necessary prerequisites for interpreting behaviour in terms of the principle of rational action. Experiment 3 confirmed that this effect cannot be attributed to generalisation of agentive properties from one object to another. These results suggest that the domain of naive psychology is initially defined only by the applicability of its core principles and its ontology is not restricted to (featurally identified) object kinds such as persons, animates, or agents. We argue that in its initial state naive psychological reasoning is not a cue-based but a principle-based theory. PMID- 10519925 TI - When and why do people avoid unknown probabilities in decisions under uncertainty? Testing some predictions from optimal foraging theory. AB - When given a choice between two otherwise equivalent options - one in which the probability information is stated and another in which it is missing - most people avoid the option with missing probability information (Camerer & Weber, 1992). This robust, frequently replicated tendency is known as the ambiguity effect. It is unclear, however, why the ambiguity effect occurs. Experiments 1 and 2, which separated effects of the comparison process from those related to missing probability information, demonstrate that the ambiguity effect is elicited by missing probabilities rather than by comparison of options. Experiments 3 and 4 test predictions drawn from the literature on behavioral ecology. It is suggested that choices between two options should reflect three parameters: (1) the need of the organism, (2) the mean expected outcome of each option; and (3) the variance associated with each option's outcome. It is hypothesized that unknown probabilities are avoided because they co-occur with high outcome variability. In Experiment 3 it was found that subjects systematically avoid options with high outcome variability regardless of whether probabilities are explicitly stated or not. In Experiment 4, we reversed the ambiguity effect: when participants' need was greater than the known option's expected mean outcome, subjects preferred the ambiguous (high variance) option. From these experiments we conclude that people do not generally avoid ambiguous options. Instead, they take into account expected outcome, outcome variability, and their need in order to arrive at a decision that is most likely to satisfy this need. PMID- 10519926 TI - DNA pulsed macrophage-mediated cDNA expression library immunization in vaccine development. AB - cDNA expression library immunization (cDELI), based on the use of a large number of pathogen's cDNA clones, has been previously used in our laboratory in an experimental model of murine Taenia crassiceps cysticercosis. In this study we show that ex vivo immunization of mice with macrophages pulsed in vitro with plasmid DNA containing cDNA expression library (2x10(4) clones) leads to significant reduction of parasite load. Additionally, pathogen-specific proliferation of splenocytes from immunized mice is demonstrated indicating the induction of cellular protective immune response and the efficacy of the proposed strategy to identify vaccine candidate antigens. Our method may represent an attractive tool in vaccine discovery applicable to any host-pathogen system and may be useful especially for the pathogens with large genomes and complicated life cycles. PMID- 10519927 TI - Priming and boosting immunity to respiratory syncytial virus by recombinant replication-defective vaccinia virus MVA. AB - Intranasal and intramuscular immunizations of mice with the highly attenuated MVA strain of vaccinia virus expressing the respiratory syncytial virus (RSV) F or G glycoprotein induced higher RSV antibody titers than those achieved by infection with RSV and greatly restricted the replication of RS challenge virus in both the upper and lower respiratory tracts. In addition, a recombinant MVA expressing both RSV F and G was stable and was as immunogenic as a combination of two single recombinant viruses. The levels of antibodies to RSV F and G, induced by previous intranasal infection with attenuated RSV, were boosted by intramuscular immunization with recombinant MVA. These data support further development of recombinant MVA as a RSV vaccine. PMID- 10519928 TI - Investigation of mucosal immunisation in pulmonary clearance of Moraxella (Branhamella) catarrhalis. AB - Moraxella (Branhamella) catarrhalis is a common cause of otitis media in children and respiratory infection in adults with chronic obstructive pulmonary disease. To identify immune responses that may facilitate the development of a mucosal vaccine, a mouse model to study pulmonary responses was established. Regimes involving intra-Peyer's patch, intratracheal and intranasal routes of immunisation with killed M. catarrhalis were investigated. A mucosal immunisation regime of a primary intra-Peyer's patch immunisation with an intratracheal boost resulted in significantly enhanced pulmonary clearance of bacteria compared to controls following an intratracheal challenge with live bacteria. Additional intratracheal boosts did not induce further enhancement of clearance. Intra Peyer's patch immunisation alone, intratracheal and intranasal immunisations did not induce enhanced clearance. The levels of specific IgG and IgA in serum and bronchoalveolar lavage fluid correlated with pulmonary clearance. The present study showed that mucosal immunisation induced enhanced pulmonary clearance of M. catarrhalis following live bacterial challenge. This mucosal immunisation model has demonstrated that a mucosal vaccine, particularly an oral vaccine, would be feasible. PMID- 10519929 TI - The cost-effectiveness of varicella vaccine programs for Australia. AB - OBJECTIVE: to examine the cost-effectiveness of three different varicella vaccination programs compared with no vaccination program. DESIGN: cost effectiveness study. Simulations of the costs and consequences of chickenpox and the vaccination programs over a 30-year period. Direct (health-care) costs only were used in the simulations. SETTING: Australia.Participants/subjects: annual birth cohorts of infants (12-months old) and adolescents (12 years old). INTERVENTIONS: strategy I (no vaccination) was compared with three different varicella vaccination programs: strategy II - all infants; strategy III - adolescents without a history of varicella; and strategy IV ('catch-up')- all infants plus, for the first 11 years, adolescents without a history. OUTCOME MEASURES: fatalities and hospitalisations for varicella and its complications (encephalitis, pneumonitis, long-term disability). RESULTS: the average cost per case of chickenpox averted was $64, $530 and $418 in the infant, adolescent and catch-up programs, respectively. The infant program was the most cost-effective of the three. This program could avert 4. 4 million cases, 13,500 hospitalisations and 30 fatalities for chickenpox over a 30-year period. RESULTS were sensitive to the price of the vaccine and the discount rate, but relatively insensitive to changes in vaccine efficacy, coverage rates or vaccine complication rates. Improved accuracy of a negative varicella history in adolescents would substantially reduce the costs of the adolescent and catch-up programs making these programs feasible. CONCLUSIONS: the infant vaccine program is the preferred program, but the direct costs of any of the vaccination programs considered here are greater than the direct costs of no vaccination program. PMID- 10519930 TI - The antigenic and immunogenic properties of synthetic peptide immunocontraceptive vaccine candidates based on gamete antigens. AB - In this study we have investigated the immunogenicity and antigenicity of synthetic immunocontraceptive vaccine candidates containing T- and B-cell epitopes arranged in different geometries. Two epitopes were selected, a B-cell epitope from fox sperm lactate dehydrogenase C(4) and a B-cell epitope from murine zona pellucida protein 3. Both were immunogenic in BALB/c mice when coupled to defined T helper cell determinants, eliciting antibodies which bound to the corresponding B-cell epitope and also to the recombinant protein. Because each of these proteins represent important components in the fertilisation process the results encourage further investigation of synthetic immunocontraceptive vaccines. PMID- 10519931 TI - Embryo vaccination of turkeys against Newcastle disease infection with recombinant fowlpox virus constructs containing interferons as adjuvants. AB - Recombinant fowlpox viruses (rFPV) expressing the fusion and hemagglutinin neuraminidase glycoproteins of Newcastle disease virus (NDV) as well as chicken type I interferon (IFN) or type II IFN were used to vaccinate specific pathogen free (SPF) turkeys in ovo. No significant changes in the hatchability, survival rate, performance and weight gain were observed after vaccination with the rFPV vaccines in comparison to diluent-inoculated embryos. The rFPV-NDV-IFN-II construct induced the onset of anti-NDV antibody production in SPF birds at one week post hatch, one week earlier than other vaccine constructs. Three to five weeks post hatch, the turkeys were challenged with the neurotropic velogenic NDV strain Texas GB (NDV-GB-Tx). The rFPV-NDV-IFN-II construct was the most protective vaccine against NDV. rFPV vaccines significantly (p<0.05) suppressed the mitogenic response of peripheral blood leukocytes in vaccinated turkeys in comparison to placebo inoculated controls at 25 days post vaccination. Birds vaccinated with rFPV-NDV-IFN-I construct did not have an inhibition in the mitogenic response. PMID- 10519933 TI - Bacterial cell envelopes (ghosts) but not S-layers activate human endothelial cells (HUVECs) through sCD14 and LBP mechanism. AB - Bacterial cell-envelopes (called ghosts) and surface layers (S-layers) are discussed to be used as vaccines and/or adjuvants, consequently it is necessary to find out which immunomodulatory mediators are induced in human cells. The present work focuses on the effects of ghosts (Escherichia coli O26:B6), S-layers (Bacillus stearothermophilus) in comparison with LPS and antibiotic-inactivated whole bacteria (E. coli O26:B6) on human umbilical vein endothelial cells (HUVEC) with regard to the release of interleukin 6 (IL-6) and the expression of surface E-selectin and the role of lipopolysaccharide binding protein (LBP), soluble CD14 (sCD14) and serum for this activation. Endothelial cells responded to ghosts, whole bacteria and LPS with IL-6 release up to 15000 pg/ml and surface E-selectin expression, while in contrast the response to S-layers with IL-6 release up to 500 pg/ml was very weak. Compared to LPS, 10-100-fold higher concentrations of bacterial ghosts and whole bacteria were required to induce the cytokine synthesis and E-selectin expression. IL-6 release and E-selectin expression of HUVECs were reduced in the absence of serum and equivalent to unstimulated samples. We have also studied the role of CD14 and LBP for the activation of endothelial cells using antiCD14 and antiLBP antibodies (Ab). AntiCD14 and antiLBP Ab both inhibited IL-6 release and E-selectin expression in a dose dependent manner after stimulation with ghosts, whole bacteria and LPS but had no effect on S-layers stimulated cells. AntiCD14 Ab inhibited more effectively than antiLBP Ab. These findings suggest that bacterial ghosts but not S-layers activate HUVECs through sCD14 and LBP dependent mechanisms. PMID- 10519932 TI - Distribution of adjuvant MF59 and antigen gD2 after intramuscular injection in mice. AB - MF59, which is an adjuvant approved for human use, typically elicits higher antibody titers than alum when used in combination with a variety of recombinant and natural subunit antigens. The mechanisms responsible for the adjuvant action of MF59 are not fully understood. In particular, little is known about the in vivo distribution of MF59 and of antigen after intramuscular (i.m.) injection. The goal of the present study was to determine the distribution of MF59 injected with soluble antigen gD2 from type 2 herpes simplex virus (HSV) and to compare the distribution of gD2 injected with or without MF59. At 4 h, 36% of the injected dose of labeled MF59 was in the quadriceps muscle and about 50% was in the inguinal fat surrounding the muscle. Half of the initial amount of labeled MF59 in muscle was detected 42 h after injection. The amount of labeled MF59 in the draining lymph nodes was maximal 2 d after injection, which represented 0.1 0.3% of the injected dose. At 4 h, 12% of the injected dose of labeled gD2 was found in the muscle. The presence of MF59 did not significantly modify the distribution of gD2. The results indicate that MF59 and gD2 distribute and are cleared independently after i.m. injection. Importantly, MF59 is unlikely to have a repository effect, whereby it slowly releases the antigen. PMID- 10519934 TI - Safety and immunogenicity of phoP/phoQ-deleted Salmonella typhi expressing Helicobacter pylori urease in adult volunteers. AB - Salmonella typhi Ty800, deleted for the Salmonella phoP/phoQ virulence regulon has been shown to be a safe and immunogenic single dose oral typhoid fever vaccine in volunteers. This promising vaccine strain was modified to constitutively express a heterologous protein of Gram negative bacterial origin, Helicobacter pylori urease subunits A and B, yielding S. typhi strain Ty1033. Seven volunteers received single oral doses of > or = 10(10) colony forming units of Ty1033; an eighth volunteer received two doses 3 months apart. Side effects were similar to those observed previously in volunteers who received the unmodified vector Ty800. All volunteers had strong mucosal immune responses to vaccination as measured by increases in IgA-secreting cells in peripheral blood directed against S. typhi antigens. Seven of eight volunteers had convincing seroconversion as measured by increases in serum IgG directed against S. typhi flagella and lipopolysaccharide antigens by ELISA. No volunteer had detectable mucosal or humoral immune responses to the urease antigen after immunization with single doses of Ty1033. A subset of three volunteers received an oral booster vaccination consisting of recombinant purified H. pylori urease A/B and E. coli heat labile toxin adjuvant 15 days after immunization with Ty1033. None of three had detectable humoral or mucosal immune responses to urease. Expression of a stable immunogenic protein in an appropriately attenuated S. typhi vector did not engender detectable mucosal or systemic antibody responses; additional work will be needed to define variables important for immunogenicity of heterologous antigens carried by live S. typhi vectors in humans. PMID- 10519935 TI - Humoral and cellular immune responses to HIV-1 nef in mice DNA-immunised with non replicating or self-replicating expression vectors. AB - OBJECTIVE: HIV accessory protein Nef is expressed early in the infectious cycle of the virus and has been shown to be an effective immunogen in humoral and cellular immune responses. We have used two different self-replicating pBN vectors and one non-replicating pCGal2 derived (pCG) vector expressing HIV-1 Nef in DNA immunisation of mice in order to determine their efficiency in raising humoral and cellular immune responses. DESIGN AND METHODS: The expression of Nef by the three plasmids was tested by transfections into COS-1 cells. Balb/c mice were immunised with the pBN-NEF and pCGE2-NEF constructs using gold particle bombardment. Immunoblotting and immunocytochemistry were used to detect in vitro expression of Nef. 51Cr release assay, ELISA and immunoblotting were used to detect cellular and humoral immune responses in immunised mice. RESULTS: Efficient in vitro expression of Nef was detected in pBN and pCGE2-NEF transfected cells, in pBN-NEF transfected cells the expression lasting up to three weeks. Anti-Nef antibodies in sera of 13 of 16 pBN-NEF immunised mice were detected within four weeks after the last immunisation, whereas only 2 of 12 pCGE2-NEF immunised mice had very weak anti-Nef antibodies. Twelve of the pBN-NEF immunised mice (75%) and 6 the pCGE2-NEF immunised mice (50%) showed Nef-specific cytotoxic T lymphocyte (CTL) responses within four weeks. CONCLUSIONS: We conclude that the three eukaryotic expression vectors tested are capable of inducing a cell mediated immune response towards HIV-1 Nef and should be considered as part of a genetic HIV vaccine. PMID- 10519937 TI - Immunization of dogs with a DNA vaccine induces protection against rabies virus. AB - Rabies is a fatal encephalomyelitis which is transmitted to man, mostly by dogs in developing countries. This zoonosis can be prevented by vaccination of humans before or after exposure. However, a more radical approach is possible, involving the elimination of the principal vector/reservoir by vaccinating dogs. The vaccine must be effective, safe and inexpensive. Mass production of plasmids is possible and DNA-based immunization with a plasmid encoding the antigen responsible for inducing protection seems to be more cost-effective than classical techniques involving cell culture. Beagles were immunized by intramuscular (i.m.) injection with a plasmid encoding the rabies virus (PV strain) glycoprotein. Neutralizing antibodies against both wild-type rabies virus and European Bat Lyssaviruses (EBL1 and EBL2) were detected after a single injection and a boost, but levels of neutralizing antibodies against EBL1 were low. Moreover, all vaccinated dogs were protected against a lethal challenge with a wild-type dog rabies strain. This is one of the first studies to demonstrate that dogs can be protected by DNA vaccines, and opens important perspectives for rabies control. PMID- 10519936 TI - Studies on a Hib-tetanus toxoid conjugate vaccine: effects of co-administered tetanus toxoid vaccine, of administration route and of combined administration with an inactivated polio vaccine. AB - A freeze-dried tetanus toxoid (T) conjugated Haemophilus influenzae type b (Hib) vaccine, was reconstituted in either diphtheria toxoid (D), DT or a combined DT and inactivated polio vaccine (IPV), and administered in an open randomized trial either intramuscularly (i.m. ) or subcutaneously (s.c.) to 287 Swedish infants at three, five and 12 months of age. When not included in the mixture, IPV was administered s.c. at a separate site. The geometric mean concentrations of Hib antibodies after primary and booster vaccinations were 1.0 and 11.6 microg/ml, respectively, with no differences related to co-administration of the carrier protein T. Antibodies against T were induced by the T conjugated Hib vaccine (Hib T) alone in 69/95 infants aged six months, and in 87/93 children aged 13 months, although infants receiving both Hib-T and T had significantly higher concentrations. Antibody responses to Hib, D, T or polio were not negatively influenced by administration route or by mixing with IPV, except that the mixed vaccine DT-IPV induced lower anti-polio GM titers after primary vaccination than did separate IPV. More local reactions were induced by the s.c. than by the i.m. route (P-values from 0.001 to 0.01). Slight increases in rates of local reactions and febrile events (>/=38 degrees C) occurred by order of dose. The low Hib antibody concentrations after the first two doses in this and other Swedish studies are unlikely to be of clinical relevance. The tetanus antibody response from T as a carrier protein alone was not sufficient for basic tetanus immunization, but should be considered in future use of additional T conjugated vaccines. PMID- 10519939 TI - At-birth immunisation against hepatitis B using a novel pre-filled immunisation device stored outside the cold chain. AB - We evaluated the immunogenicity of hepatitis B (HB) vaccine in UniJect, a pre filled, non-reusable injection device, stored at tropical temperatures for up to one month and used to give the first dose of HB vaccine to newborns. Infants in Tabanan district, Bali, Indonesia, were given their first dose of HB vaccine with UniJect stored out of the cold chain, UniJect stored in the cold chain; or standard syringe, needle and multidose vial stored in the cold chain. Subsequent doses were given by usual means and blood samples drawn 4-6 weeks after the third dose. No significant differences were found in seroconversion rates or geometric mean titres of HB surface antibody between the three groups. PMID- 10519940 TI - Generation of a parainfluenza virus type 1 vaccine candidate by replacing the HN and F glycoproteins of the live-attenuated PIV3 cp45 vaccine virus with their PIV1 counterparts. AB - Parainfluenza virus type 1 (PIV1) is a major cause of croup in infants and young children, and a vaccine is needed to prevent the serious disease caused by this virus. In the present study, a live attenuated PIV1 vaccine candidate was generated by modification of the extensively-studied PIV3 cold-passaged (cp) cp45 vaccine candidate using the techniques of reverse genetics. The HN and F glycoproteins of the PIV3 cp45 candidate vaccine virus were replaced with those of PIV1. This created a live attenuated PIV1 vaccine candidate, termed rPIV3-1 cp45, which contained the attenuated background of the PIV3 cp45 vaccine virus together with the HN and F protective antigens of PIV1. Three of the 15 mutations of cp45 lie within the HN and F genes, and those in the F gene are attenuating. Thus, some attenuation might be lost by the HN and F glycoprotein replacement. To address this issue we also constructed a derivative of PIV3 cp45, designated rPIV3 cp45 (F(wt)HN(wt)), that possessed wild type PIV3 HN and F glycoproteins but retained the 12 other cp45 mutations. rPIV3 cp45 (F(wt)HN(wt)) replicated in the respiratory tract of hamsters to a level three- to four-fold higher than rPIV3 cp45, indicating that loss of the two attenuating mutations in the cp45 F gene effected a slight reduction in the overall attenuation of cp45 for hamsters. However, the chimeric rPIV3-1 cp45 virus was about 5-fold more restricted in replication in hamsters than rPIV3 cp45 and about 15- to 20-fold more restricted than rPIV3 cp45 (F(wt)HN(wt)). This suggests that two components contribute to the attenuation of the new chimeric rPIV3-1 cp45 PIV1 vaccine candidate: one being the 12 cp45 mutations, which provide most of the observed attenuation, and the other resulting from the introduction of the heterologous PIV1 HN and F proteins into PIV3 (i.e., a chimerization effect). rPIV3-1 cp45 was observed to be immunogenic and protective against challenge with wild type PIV1 in hamsters. This virus shows sufficient promise that it should be evaluated further as a candidate live attenuated vaccine strain for preventing severe lower respiratory tract PIV1 disease in infants and young children. PMID- 10519938 TI - Foreign gene expression in Corynebacterium pseudotuberculosis: development of a live vaccine vector. AB - A defined phospholipase D mutant of Corynebacterium pseudotuberculosis, designated Toxminus, was used as a live vector to express and deliver a range of candidate vaccine antigens to sheep. Expression levels of the foreign genes in Toxminus were evaluated when directed from a number of different promoters, both constitutively expressed and inducible, as fusions with expressed genes including a signal sequence, and from chromosomal and episomal loci. In general expression levels were low and it appeared that some of the recombinant proteins were tolerated by C. pseudotuberculosis Toxminus better than others. Gene expression was however sufficiently high for three of the genes to elicit antibody responses specific to the recombinant protein following a single dose of the live Toxminus vector vaccine. This work suggests that C. pseudotuberculosis Toxminus has potential for development as a live veterinary vaccine vector. PMID- 10519941 TI - Protective effects against simian immunodeficiency virus agm (SIVagm) infection in cynomolgus monkeys immunized with a recombinant vaccinia virus expressing the SIVagm envelope gene. AB - Protective immunity induced by a recombinant vaccinia virus expressing the envelope (Env) protein of a simian immunodeficiency virus strain, SIVagm, (SEN RVV), was evaluated in cynomolgus monkeys (Macaca fascicularis). Three monkeys were immunized twice with SEN-RVV and boostered with the purified SIVagm Env protein. These monkeys developed high titers of anti-SIVagm Env antibody, especially after boostering. After challenge with polyclonal SIVagm, no virus was recovered from two of the monkeys and no provirus DNA was detected in one of these two. After autopsy, however, proviral DNA was detected in the spleen of this monkey. These results suggest that this immunization regimen could not completely protect the monkeys from SIV infection but that it did reduce the replicability of the challenged virus. PMID- 10519942 TI - Needle-free, non-adjuvanted skin immunization by electroporation-enhanced transdermal delivery of diphtheria toxoid and a candidate peptide vaccine against hepatitis B virus. AB - To explore the feasibility of non-adjuvanted, needle-free skin immunization, we examined the immune responses of mice immunized by three routes with two antigens (Ag) differing in molecular size and lipophilicity. Diphtheria toxoid (DT) or a myristylated peptide (MYR) administered either by transdermal electroporation (EP) or intradermally (i.d.) with a needle elicited markedly different responses. The EP route elicited higher responses to MYR than i.d. immunization, but lower responses to DT. Intraperitoneal (i.p.) immunization evoked responses against MYR that were higher than those evoked by EP. It was inferred that EP is a promising technique for non-adjuvanted skin immunization, specially with low molecular weight, weakly immunogenic Ag. PMID- 10519943 TI - Antibodies against pneumococcal polysaccharides after vaccination in HIV-infected individuals: 5-year follow-up of antibody concentrations. AB - We studied the production of IgG antibodies against eight pneumococcal polysaccharide serotypes (PPS) 1, 4, 6B, 9V, 14, 18C, 19F and 23F after vaccination of 50 HIV-infected adults with 23-valent Pneumovax((R))23 and the course of the antibodies against four PPS during the following years. Mean antibody concentrations against PPS 18C, 19F and 23F were sigificantly lower in the patients with CD4(+)-lymphocyte counts <200x10(6)/l than in healthy controls; mean antibody concentrations against PPS 1, 4, 9V, 6B and 14 were similar in HIV infected individuals and controls. Although it has been assumed that polysaccharides induce a T-cell-independent immune response, our results indicate that some PPS are T-cell-independent type 2 antigens. The rates of decline of mean antibody concentrations in HIV-infected individuals and in healthy controls were similar during 5 y after vaccination. However, as a consequence of the low postvaccination antibody concentrations against several PPS, within 3 y after vaccination most HIV-infected individuals had antibody concentrations below the level which is assumed to be required for protection. PMID- 10519944 TI - Phase I trial of two recombinant vaccines containing the 19kd carboxy terminal fragment of Plasmodium falciparum merozoite surface protein 1 (msp-1(19)) and T helper epitopes of tetanus toxoid. AB - The safety and immunogenicity of 2 yeast-derived, blood-stage malaria vaccines were evaluated in a phase l trial. Healthy adults were given 2 or 3 doses of alum adsorbed vaccine containing the 19 kDa carboxy-terminal fragment of the merozoite surface protein-1 (MSP-1(19)) derived from the 3D7 or the FVO strain of Plasmodium falciparum fused to tetanus toxoid T-helper epitopes P30 and P2. The first 2 doses of MSP-1(19) were well tolerated. Hypersensitivity reactions occurred in 3 subjects after the third dose of MSP-1(19), including bilateral injection site reactions in 2 (one with generalized skin rash), and probable histamine-associated hypotension in 1. Serum antibody responses to MSP-1(19) occurred in 5/16, 9/16 and 0/8 subjects given 20 microg of MSP-1(19), 200 microg of MSP-1(19), and control vaccines (hepatitis B or Td), respectively. Both MSP 1(19) vaccines were immunogenic in humans, but changes in formulation will be necessary to improve safety and immunogenicity profiles. PMID- 10519945 TI - Vaccination against lyme disease with recombinant Borrelia burgdorferi outer surface protein A (rOspA) in horses. AB - Eight 1-year-old ponies were vaccinated with recombinant OspA (ospA gene derived from B. burgdorferi B31) with adjuvant (aluminium hydroxide). Four ponies were used as non-vaccinated controls with adjuvant. One hundred and twelve days after the first vaccination, the vaccinated and non-vaccinated ponies were challenged by exposure to B. burgdorferi-infected adults tick (Ixodes scapularis) collected from Westchester County, New York (tick infection rate >/=60%). Protection from infection was evaluated by culture for B. burgdorferi from three monthly skin biopsies taken near the site of tick bites. B. burgdorferi was not isolated from any of the vaccinated ponies. In contrast, three of four control ponies challenged by tick exposure were skin culture positive. At the time of tick exposure, vaccinated ponies had antibody to B. burgdorferi demonstrable by KELA (kinetic-ELISA), western blot and a serum growth inhibition assay. Antibodies in the challenge control ponies were only detectable by two to three months after tick exposure and remained at intermediate levels until termination of the study. By western blot analysis, antibodies to OspA first appeared in the sera of vaccinated ponies three weeks after the first vaccination. The absence of additional bands, known to develop when the animal is infected, suggests that infection was blocked after tick exposure of vaccinated ponies. Results from this study show that vaccination with recombinant OspA protected ponies against infection after experimental challenge with B. burgdorferi-infected ticks. PMID- 10519946 TI - Peptide vaccination using nonionic block copolymers induces protective anti-viral CTL responses. AB - High molecular weight nonionic block copolymers have been developed as vaccine adjuvants. We employed these adjuvants in water-in-oil emulsion and multiple emulsion formulations with a synthetic peptide-based antigen vaccine to test their ability to prime anti-viral CD8(+) T cell responses. Vaccines were made using the H-2(d)-restricted immunodominant peptide from lymphocytic choriomeningitis virus (LCMV), NP118-126, and administered to BALB/c ByJ (H-2(d)) mice. Peptide-containing emulsions were able to induce NP118-126 specific CTL and IFN-gamma secreting CD8(+) T cells in the vaccinated mice and these responses were maintained for at least 90 days post immunization. At all times, the responses induced by the copolymer formulations were equal to, or better than, formulations based on incomplete Freund's adjuvant (IFA). In addition, the responses induced by prophylactic vaccination using the multiple emulsion formulation resulted in accelerated viral clearance following infection with a strain of LCMV (clone 13) that causes a persistent infection in naive adult mice. These results indicate that peptide vaccination using a formulation based on high molecular weight nonionic block copolymer in a simple water-in-oil or a multiple emulsion format can induce virus-specific CD8(+) T cell responses and confer protection sufficient enough to prevent the establishment of a persistent infection. PMID- 10519947 TI - Intranasal IFN-gamma gene transfer protects BALB/c mice against respiratory syncytial virus infection. AB - Respiratory syncytial virus (RSV) is a major respiratory pathogen in infants, young children and the elderly and causes severe bronchiolitis and asthma. In an effort to develop a preventive IFN-gamma therapy against RSV infection, an intranasal gene transfer strategy was utilized. Intranasal administration of a plasmid expressing the IFN-gamma cDNA (pIFN-gamma) resulted in the expression of IFN-gamma in murine lungs and decreased RSV replication. The mice administered with pIFN-gamma and then infected with RSV exhibited a significant decrease in broncho-alveolar lavage lymphocyte and neutrophil counts. A significant reduction in epithelial cell damage, infiltration of mononuclear cells in the peribronchiolar and perivascular regions, and thickening of the septa was observed in the lungs of mice treated with pIFN-gamma when compared to controls. These results suggest that intranasal IFN-gamma gene transfer results in decreased RSV replication and pulmonary inflammation and may be useful against RSV infection. PMID- 10519948 TI - Long-term effects of PZP immunization on reproduction in white-tailed deer. AB - A 6-year study was conducted to determine the long-term effects of porcine zona pellucida (PZP) vaccine on the immune and hormonal responses, and reproduction of the white-tailed deer. The first 2 years of active immunization resulted in an 89% reduction in fawning. Vaccination with PZP produced reversible infertility lasting 1-4 years. Infertility was directly related to immune titers to PZP. Doe fertility was restored when the antibody titer dropped to minimal levels, but following re-immunization, infertility was reestablished. Reduction in fawning throughout the 6-year study was 76%. It was also observed that immune responses among deer were variable, especially in the first year of treatment. Variability was also observed among deer for the duration of infertility following the initial vaccination. PMID- 10519949 TI - Research funding by the CAG reaches its highest level ever! PMID- 10519950 TI - Prophylaxis for spontaneous bacterial peritonitis. PMID- 10519951 TI - Antibacterial activity of Helicobacter pylori. PMID- 10519952 TI - Canadian Helicobacter Study Group Consensus Conference on the Approach to Helicobacter pylori Infection in Children and Adolescents. AB - Gastric infection with Helicobacter pylori is common in both children and adults, but children are considerably less susceptible to peptic ulcers and other pathological sequelae. As a result, the risk to benefit ratio of diagnostic studies and therapeutic regimens for H pylori in adults are likely different from those in pediatric populations. These guidelines for the management of pediatric H pylori infection, developed by the Canadian Helicobacter Study Group, are designed to identify when the diagnosis and treatment of H pylori may improve patient care. Given the low prevalence of this infection in Canada, it is important to recognize that indiscriminate testing and treatment programs in children are not recommended, and indeed may threaten the optimal care of children. Diagnostic tests should be employed judiciously and be reserved for children who are most likely to derive measurable benefit, such as those likely to have peptic ulcer disease. At this time a test and treat strategy in children cannot be considered prudent, evidence based or cost effective. It is appropriate to limit diagnosis and treatment to children and adolescents in whom H pylori has been identified during endoscopic investigation. PMID- 10519953 TI - Clinical practice guidelines and Helicobacter pylori infection in children. AB - The objective of this paper is to review the principles, methods and issues behind the development of clinical practice guidelines. Practice guidelines have been defined as "systematically developed statements to assist practitioner and patient decisions about appropriate health care for specific clinical circumstances". The ultimate goal of guidelines is to improve patient outcomes; however, they may also be used as tools to decrease health care costs, improve medical education and enhance quality assurance. Evidence-based guidelines use explicit methods to link recommendations to the quality of the underlying research. Following development of the guideline, implementation and evaluation are key steps. The ultimate aim of guideline development is to influence physician knowledge, attitudes and behaviour. PMID- 10519954 TI - Current therapy for Helicobacter pylori infection in children and adolescents. AB - Helicobacter pylori infects approximately 50% of the world's population and is a definitive cause of gastroduodenal disease (ie, gastritis, duodenal and gastric ulcers) in children and adults. Four consensus conferences held around the globe have brought together clinicians, scientists, epidemiologists and health care economists to discuss the role of the gastric pathogen H pylori in human gastroduodenal disease. At each of these conferences, the overriding objective was to reach a consensus on the development of practical guidelines for the diagnosis and treatment of H pylori-infected individuals. However, it was not until the Canadian H pylori Consensus Conference, held in November 1997, that the issues of H pylori infection in children were addressed. Therapies for H pylori infection in children, presented in part at the First Canadian Pediatric H pylori Consensus Conference, held in Victoria, British Columbia, November 1998, are reviewed in this paper. PMID- 10519955 TI - Helicobacter pylori: novel therapies. AB - The ideal therapy for Helicobacter pylori would cure the infection without resulting in the development of antibiotic resistance. Current therapies have variable cure rates; the reasons for treatment failure include bacterial resistance and poor compliance. Some antibiotics, such as furazolidone, may be affordable agents to treat this infection worldwide. New proton pump inhibitors, such as rabeprazole, can potentiate antibiotics. Nutriceuticals and probiotics demonstrate interesting in vitro activity against H pylori. Children rarely have symptoms to this infection and, therefore, are a suitable group in which to assess different nonaggressive therapies. PMID- 10519956 TI - Novel diagnostic tests to detect Helicobacter pylori infection: A pediatric perspective. AB - Because of the widespread problem of Helicobacter pylori infections, there is an increased need for rapid, reliable and inexpensive diagnostic tests. Five recently developed tests that offer potential advantages because they are less invasive or permit easier acquisition of samples than available tests are assessed. The tests assessed are whole blood, saliva and urine assays that measure systemic antibody response to H pylori, stool tests that measure H pylori antigens and string tests that recover H pylori organisms. PMID- 10519957 TI - Local and systemic antibody responses in humans with Helicobacter pylori infection. AB - Immunization can prevent or cure an otherwise chronic helicobacter infection in several animal models despite the chronic nature of natural helicobacter infections. Differences in the antigenic specificity of the antibodies may contribute to the protection observed in these experimental animals. The goal of the present study was to compare the local and systemic antibody responses of humans with chronic Helicobacter pylori infection with those of an individual with spontaneous resolution of infection to find an immunological correlate of protection. Spontaneous resolution of infection was accompanied by a change in immunoblot profiles. Whereas a broad range of H pylori antigens was recognized in chronically infected patients (including the patient who ultimately cleared the infection spontaneously), resolution of infection in the absence of therapeutic agents resulted in the recognition of only several immunodominant antigens. The most dominant antigen was approximately 66 kDa in molecular mass. Immunoblot analysis demonstrated that these antibodies were specific for the structural subunits of the urease enzyme. These studies suggest that the success of antihelicobacter immunization may be due to the ability of vaccination to induce an immune response against antigens that are normally not immunodominant during the course of infection. PMID- 10519958 TI - Transmission of Helicobacter pylori infection. AB - Helicobacter pylori infection is one of the most common bacterial infections worldwide. It is accepted as the major cause of chronic gastritis, peptic ulcer, carcinoma of the distal part of the stomach and gastric lymphoma. However, how and when the infection is acquired remain largely unknown. Identification of mode of transmission is vital for developing preventive measures to interrupt its spread, but studies focused on this issue are difficult to implement. From epidemiological studies, it is known that there are great differences in the prevalence of infection in different populations and in ethnic groups originating from high prevalence regions. This is likely related to inferior hygienic conditions and sanitation. In developing countries, infection occurs at a much earlier age. In developed countries, the prevalence of infection is related to poor socioeconomic conditions, particularly density of living. Humans seem to be the only reservoir of H pylori, which spread from person to person by oral-oral, fecal-oral or gastro-oral routes. Most infections are acquired in childhood, possibly from parents or other children living as close contacts. Infection from the environment or from animals cannot be entirely excluded. PMID- 10519959 TI - Pathobiology of Helicobacter pylori infection in children. AB - In the pediatric population, the associations of Helicobacter pylori with gastritis, gastric ulcer, duodenitis and duodenal ulcer, and with duodenal gastric surface metaplasia and disorders of the D cell- G cell axis resulting in hypergastrinemia, are well established and in many ways resemble their counterparts in adults. Eradication of H pylori invariably results in the reversal of these diseases with time. There are also suggestions that gastric surface metaplasia is more extensive in children with H pylori, and may be the site of duodenal H pylori infection and associated duodenal erosions or ulcers. There is no consensus as to whether H pylori in children is more or less severe than in adults. In one pediatric cohort, H pylori was associated with increased intensity of inflammation, while other studies suggest that acute inflammation may be less intense in children overall but that chronic inflammation may be increased in intensity, including lymphoid hyperplasia, which in turn may correlate with endoscopic nodularity. Lymphoid hyperplasia and nodular gastritis appear to be more frequent in children than in adults and usually regress following H pylori eradication. However, in children, other diseases or morphological abnormalities, including some loss of glands (atrophy), occasionally intestinal metaplasia, lymphoproliferative diseases including low grade mucosal-associated lymphoid tissue lymphoma, lymphocytic gastritis and hypertrophic gastritis/Menetrier's disease, are much less frequently associated with H pylori than in adults. Other associations are rarely seen in children, primarily because the time required for these to develop takes the individual to adulthood; for example, while intestinal metaplasia occurs in the pediatric population, the complications of adenoma/dysplasia and carcinoma are rare. In adults, inflammatory and hyperplastic polyps, atrophic gastritis and pernicious anemia, and in some patients granulomas (granulomatous gastritis), may also be associated with H pylori infection. Greater awareness of the spectrum of diseases associated with H pylori may well lead to their increased recognition in the pediatric population. Some diseases, particularly Crohn's disease, but also human immunodeficiency virus infection, have a negative association with H pylori that appears not to be simply a result of the excess antibiotic therapy that these patients receive. These variations in association and reactions to H pylori, some of which are age-related, may allow the different host responses to H pylori that occur in humans to be examined. PMID- 10519960 TI - Helicobacter pylori infection and childhood recurrent abdominal pain: lack of evidence for a cause and effect relationship. AB - BACKGROUND: Recurrent abdominal pain is a common complaint among children and adolescents. Apley's criteria - at least three discrete episodes of abdominal pain of sufficient severity to interrupt normal activities that occur over a period of three or more months - are often used to define this chronic pain syndrome. OBJECTIVE: To summarize the extent and quality of the published evidence for a cause-and-effect relationship between Helicobacter pylori infection and childhood recurrent abdominal pain. MATERIALS AND METHODS: The MEDLINE bibliographic database (January 1983 to July 1998) was searched to identify pertinent (English language) studies. The search was restricted to prospective, controlled studies reporting empirical data on children up to 18 years of age. RESULTS: Six studies were identified. Five of the six were case control studies, while the remaining study assessed the effectiveness of antibiotic therapy. Only one study was community based, with the remaining studies conducted in the tertiary hospital setting. The evidence for a causal relationship was inconsistent; of the five case-control studies reviewed, the odds ratios ranged from 0.32 to 1.80. Two studies demonstrated statistically significant results; however, the findings were conflicting. The only treatment trial was limited because of methodological flaws. CONCLUSIONS: Current evidence suggests no association between H pylori infection and recurrent abdominal pain in children. The evidence to date indicates that routine investigation for H pylori infection in children who present with the classical symptoms of recurrent abdominal pain based on Apley's criteria is not warranted. PMID- 10519962 TI - How to reap benefits from other people's money. PMID- 10519961 TI - Determinants of disease outcome following Helicobacter pylori infection in children. AB - Helicobacter pylori infection is primarily acquired during childhood, causes chronic, active gastritis and peptic ulcer disease, and is associated with the development of gastric malignancies. However, only a small number of infected individuals ever develop the more severe sequelae of peptic ulcer disease and gastric cancers. Therefore, the identification of bacterial and host factors that play a role in determining the outcomes and pathophysiology of infection is a major focus of current research. Recent advances in the understanding of disease pathogenesis are critically considered, with particular reference to the pediatric population. PMID- 10519963 TI - [Multiple actions of EGCG, the main component of green tea]. AB - Green tea, the most popular beverage in Japan and China, contains epicatechin derived compounds which have been characterized in some epidemiological studies as having protective effects against cancer. Epigallo catechin-O-gallate (EGCG), the most active epicatechin in green tea was previously found to block the in vitro growth of many cancer cell lines and in vivo to strongly reduce tumor growth in cancer-bearing animals. Today, green tea consumption by animals is shown to markedly reduce VEGF-induced angiogenesis, a neo-vascularization process occurring in several physio-pathological conditions. EGCG is also able to inhibit endothelial cell growth in vitro and angiogenesis process in vivo. Elsewhere, EGCG has been described as a potent inducer of apoptosis and an inhibitor of telomerase activity. Because EGCG is acting on different processes, it could trigger various molecular mechanisms of action. Its anti-oxidant properties could explain its antagonistic action in some inflammatory processes. In summary, although no direct molecular target has been so far elucidated for EGCG, the multi-potentialities of this molecule, along with its broad bioavailability, render it very attractive as a putative curative drug for various diseases such as dermatosis, gout, atherosclerosis and cancer. PMID- 10519964 TI - [Beta-catenin mutations in a common skin cancer: pilomatricoma]. AB - The normal hair development requires WNT signalling pathways. A constitutive activation of beta-catenin, one of the components of this cascade, induces an abnormal proliferation of hair matrix cells. This activation is observed in a human skin tumours called pilomatricoma. Mutations of the beta-catenin gene are detected in 75% of the tumours analysed. PMID- 10519965 TI - [LY231514. A multitargeted antifolate (MTA)]. PMID- 10519967 TI - [Adult's anaplastic CD30+ large cell lymphomas]. AB - Anaplastic large-cell lymphomas were recognized by Stein in 1985. Less than fifteen years were necessary to confirm this entity, as well as her phenotype and to characterize the t(2;5) (p23;q35) chromosomal abnormality. This rare subgroup of non-Hodgkin's lymphomas (15% of peripheral T cell lymphomas and 8% of all diffuse aggressive lymphomas) is individualized in the Real classification. This disease, which had a bimodal age distribution, is clinically characterized by a diffuse nodal involvement and the frequency of extranodal involvement, especially skin and lungs. Primitive cutaneous anaplastic large cell lymphomas belong to the cutaneous CD30+ lymphoproliferative diseases spectrum. Among peripheral T cell and diffuse aggressive lymphomas, they have the better prognosis. We present in this paper a review of the recent advances in the knowledge, treatment and prognosis of this peculiar entity. PMID- 10519966 TI - [Bisphosphonates and bone metastases]. AB - Bisphosphonates, potent inhibitors of bone resorption have been emerging as the standard treatment of tumor-induced hypercalcemia during the 90's. All uncontrolled phase II studies up to 1992 had demonstrated efficacy in reducing morbidity in terms of bone pain, fracture and hypercalcemia. Other studies on intravenous bisphosphonates, with no other anti-tumor treatment, even demonstrated sclerosis of osteolytic breast cancer bone metastases. Randomised phase III studies only began after 1992. In multiple myeloma, one study with oral clodronate has reported a decrease in bone events and two other studies, one with intravenous pamidronate and the other with oral clodronate have both reported a decrease in skeletal events and bone pain. In breast cancer patients with bone metastases, five large studies have been reported: three with intravenous pamidronate, one with oral pamidronate and one with oral clodronate. All these studies have demonstrated the superiority of bisphosphonates over placebo on both bone pain and bone events, but have failed to show an increase in duration of survival. Bisphosphonates should therefore be considered as an important part of the palliative treatment in breast cancer patients with bone metastases. On the other hand, no definite conclusion can be drawn on the role of bisphosphonates in the treatment of prostatic carcinoma bone metastases yet. However, bisphosphonates should be considered as part of the standard therapy in managing painful lesions in patients with multiple myeloma, breast cancer and prostatic cancer. Nevertheless, further studies are needed with bisphosphonates in the adjuvant setting before bone metastases appear. Could new and more potent bisphosphonates such as zoledronate further reduce bone metastases morbidity? PMID- 10519968 TI - [Initial medical treatment for breast cancer]. AB - Rationale for primary medical treatment of breast cancer relies on experimental data showing that the incidence of metastatic disease is dependent on the primary tumour mass and tumoral angiogenesis. Although this concept may be applied to both chemotherapy and hormonotherapy, only the first was extensively explored for patients with locally advanced breast cancer, and more recently in smaller tumours. Despite high clinical +/- radiological response rates, only pathologic information, carefully assessed in both the primary and axillae lymph nodes, stands out as the major source of prognostic information on patients' outcome. Recent developments in chemotherapy (dose-intensity, new drugs) do not seem to influence these results, indicating the possible limitations of conventional chemotherapy. Of 6 published randomized trials comparing neo versus adjuvant strategy, none showed any significant impact of primary chemotherapy on survival, with in some a trend towards delayed and/or distant recurrences if neoadjuvant treatment given. That some recent reports suggest that local relapse rate might be increased after conservative treatment following induction chemotherapy in subgroups analyses should cause oncologists to revise the role for post neoadjuvant treatment conservative surgery without calling into question the global strategy. Through sequential samplings, neoadjuvant medical treatment provides indeed the opportunity (a) to identify molecular mechanisms associated with pathologic response and (b) to study the possibility to guide the choices for induction treatment and patients' populations submitted to primary medical treatment. PMID- 10519969 TI - [Decision committees in oncology: from standard to actual cases]. AB - Medical decision making, especially in oncology, is more and more assisted by pre established therapeutic protocols. But applying these protocols for particular cases is sometimes difficult; then, local expertise intervenes. For solving these difficult cases, Alexis-Vautrin like other centers, organizes committees for therapeutic decisions. This paper aims to describe, from a field study, the tools for medical decision making and the difficulties of using them. These result of the analysis of decision committee for breast cancers in a center using of protocols, which the first were established twenty years ago. This analysis was carried out by a team of oncologists and ergonomists; it stresses on collective decision making as a mean for adapting rules and exchanging knowledge. Furthermore, these findings lead to discuss using of decision committee for advancement and particularization of protocols themselves. PMID- 10519970 TI - [Endothelin receptors in benign human tumours of uterine muscle]. AB - The endothelins (ET1, ET2, ET3) are a family of peptides that exert vasoactive and mitogenic effects. ETs bind to at least two subtypes of receptors: the ETA subtype is ET1 selective whereas the ETB subtype binds ET1, ET2 and ET3. By RT PCR, we detected ETA receptor mRNA and ETB receptor mRNA in leiomyoma and in homologous myometrium distal from the tumor. Despite the presence of four spliced variants of ETA receptors, we identified a single class of ETA-binding sites. The level of ETB receptor mRNA was found to be higher in myometrium versus leiomyomas. Using complementary pharmacologic approach, we demonstrated the predominance of ETA receptors in normal myometrium (75% of total receptors). Both ETA and ETB transcripts coexist in leiomyomas, but we have reported only ETA binding sites. Because of growth properties of ET1, we suggest a role for this peptide in the tumoral development of human uterine smooth muscle. PMID- 10519971 TI - Multiple signalling pathways exist in the stress-triggered regulation of gene expression for catecholamine biosynthetic enzymes and several neuropeptides in the rat adrenal medulla. AB - A critical component of the response to stress is the coincident activation of the hypothalamic-pituitary-adrenal axis and the sympathoadrenal system - comprised of sympathetic ganglia and the adrenal medullae. The sympathoadrenal system produces the catecholamines - noradrenaline and adrenaline, and several neuropeptides, involved in the homeostatic mechanisms that govern the adaptation to stress. This brief survey aims to provide a general overview of the present knowledge about the impact of stress on neurotransmitter gene expression in the adrenal medulla, with particular attention paid to the apparent heterogeneity in stress-evoked signals and regulatory pathways. PMID- 10519972 TI - Conducted vasomotor responses in arterioles: characteristics, mechanisms and physiological significance. AB - Micropipette application of certain vasoconstrictor or -dilator substances onto the surface of arterioles induces both a local vasomotor response and a response which is propagated up- and downstream along the vessel, a so-called conducted vasomotor response. In some vascular beds conducted vasoconstrictor and dilator responses are detectable more than a millimetre from the site of agonist delivery. While agonists such as acetylcholine, noradrenaline, and KCl almost invariably give rise to conducted vasomotor responses others, such as sodium nitroprusside or vasopressin, do not. Conducted vasomotor responses in arterioles appear to rely on passive electrotonic spread of the change in membrane potential induced by the agonist at the tip of the pipette. Presumably the current spreads up- and downstream along the arteriolar wall through endothelial or smooth muscle cell gap junctions. Whether the electrical signal is conducted primarily through the endothelial or the smooth muscle cell layer or both is currently not known, but it may depend on the agonist used. Experiments have suggested that conducted vasodilation in skeletal muscle feed arterioles plays an important role in the development of functional hyperaemia at the onset of exercise. In the kidney, conducted vasoconstriction is believed to be responsible for the upstream contraction of the afferent arteriole and interlobular artery known to occur in response to activation of the macula densa. Therefore conducted vasoconstriction could be important for the tubuloglomerular feedback mechanism. Finally, experimental studies have shown that conduction of vasomotor responses in arterioles may be altered in pathological conditions associated with microvascular dysfunction such as arterial hypertension and sepsis. PMID- 10519973 TI - Post-ischaemic dysfunction does not correlate with release of cardiac troponin T in isolated rat hearts. AB - Cardiac troponin T (cTnT) is a highly sensitive and specific serum marker of irreversible cardiomyocyte injury. It is not clear whether cTnT also is a suitable marker of subtle, reversible injury. In the present investigation the relationship between cTnT release and function during the first 30 min of reperfusion after 30 min of global ischaemia, was investigated in isolated, retrogradely perfused rat hearts. Left ventricular systolic (LVSP), end-diastolic (LVEDP) and developed (LVDP) pressures, heart rate (HR), and coronary flow (CF) were measured. In one series of experiments (n=7) the kinetics of cTnT release during 30 min of reperfusion was investigated. An early, short-lasting peak of cTnT release appeared after 30 s of reperfusion. Then cTnT release gradually increased with a maximum after 20 min (from 0.08 +/- 0.03 before ischaemia to 2.16 +/- 0.40 ng min-1) (mean +/- SEM). In a second series of experiments (n=52) the relationship between cTnT release and cardiac function was investigated after 20 min of reperfusion. At this time point LVEDP increased (0 to 62 +/- 3 mmHg) and LVDP decreased (84 +/- 2 to 33 +/- 3 mmHg), but without any correlation with cTnT release. cTnT release was positively correlated to LVSP (P < 0.04, r=0.29), and negatively correlated to HR (P < 0.03, r=-0.31). cTnT concentration in the coronary effluent increased in parallel to increasing CF (P < 0.03, r=0.31). In conclusion, during the early reperfusion period there was no consistent correlation between cTnT release and dysfunction after global ischaemia in the isolated rat heart. Release of cTnT and post-ischaemic function appear to provide supplementary information in this particular model. PMID- 10519974 TI - Heart rate variability in healthy volunteers during normobaric and hyperbaric hyperoxia. AB - Inhaled supranormal partial pressure of oxygen induces bradycardia and peripheral vasoconstriction. The exact mechanism of the decreasing heart rate is not clear, but the autonomic nervous system is partly involved. In the present study the role of the autonomic nervous system in hyperoxic bradycardia was evaluated by using the power spectral analysis of heart rate variability. Ten healthy volunteers participated in four experiments: (i) hyperbaric oxygen treatment (100% oxygen at 2.5 ATA), (ii) hyperbaric air treatment (O2 21% at 2.5 ATA), (iii) oxygen treatment at normal pressure (100% O2, 1 ATA) and (iv) air breathing at normal pressure (21% O2, 1 ATA). During the experiments, ECG was registered and subjected to power spectral analysis. The volunteers rated their perception of temperature, ear discomfort, sweating and excitement on a visual analogue scale. Statistical comparison of the results of the four trials was conducted with a two-way ANOVA for repeated measurements. Heart rate decreased during all interventions, but there were no statistically significant differences between the sessions. High frequency variability of heart rate variability and Hayano's index of HF power increased and LF/HF ratio decreased with increasing partial pressure of oxygen. Our results suggest, that normobaric and hyperbaric hyperoxia increase parasympathetic influence in the regulation of the heart. PMID- 10519975 TI - No role of interstitial adenosine in insulin-mediated vasodilation. AB - The mechanisms behind the vasodilatory effect of insulin are not fully understood, but nitric oxide plays an important role. We have investigated the possibility that insulin mediates vasodilatation in the human skeletal muscle via an increase in extracellular adenosine concentrations. In eight healthy subjects (H) and in four subjects with a complete, high (C5-C6/7) spinal cord injury (SCI) a hyperinsulinaemic (480 mU min-1 kg-1), isoglycaemic clamp was performed. SCI subjects were included as it has been proposed that adenosine and adenine nucleotides may be released from nerve endings in the skeletal muscle. Adenosine concentrations in the extracellular fluid (ECF) of skeletal muscle in the thigh were measured by means of the microdialysis technique. Leg blood flow (LBF) was measured by termodilution. In response to insulin infusion, LBF always increased (P < 0.05) (from 228 +/- 25 and 318 +/- 18 mL min-1 to 451 +/- 41 and 530 +/- 29 mL min-1, SCI and H, respectively [mean +/- SEM]). Concentrations of adenosine in the muscle ECF did not change with infusion of insulin and did not differ between groups (before: 147 +/- 55 [SCI] and 207 +/- 108 [H] nmol L-1; during: 160 +/- 36 [SCI] and 165 +/- 74 [H] nmol L-1). No significant correlation between concentrations of adenosine and corresponding LBF rates was achieved (LBF=[ 0.0936. Adenosine] + 475. R=-0.092, P=0.22, number of samples=181, number of subjects=12). CONCLUSION: the mechanism by which insulin mediates an increase in skeletal muscle blood flow is not associated with adenosine in the ECF. PMID- 10519976 TI - Pituitary adenylate cyclase-activating polypeptide inhibits the cyclooxygenase pathway of rat cerebral microvessels. AB - The presence of nerve endings containing pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal polypeptide (VIP) around cerebral microvessels suggests that these peptides have regulatory roles in the cerebral microcirculation. Prostanoids synthesized by the cerebrovascular endothelium have a determining role in the regulation of the brain circulation. In the present study, the effects of PACAP and VIP on the cyclooxygenase pathway of cerebral microvessels were investigated. The isolated microvessels were incubated with 1 14C-arachidonic acid and different concentrations of the peptides. The prostanoids formed were separated by means of overpressure thin-layer chromatography, and were quantitatively determined by liquid scintillation. Higher concentrations (10-7 and 10-6 mol L-1) of PACAP significantly inhibited the activity of the cyclooxygenase pathway, whereas VIP had no significant effect on it. As regards the cyclooxygenase metabolites, the syntheses of thromboxane A2 and prostaglandin D2 were inhibited significantly. PACAP and VIP are known to increase the intracellular cAMP level in the cerebral microvessels and in the present experiments the protein kinase A inhibitor H-89 attenuated the effect of PACAP on prostanoid synthesis. It is concluded that the cyclooxygenase pathway of rat cerebral microvessels is more sensitive to PACAP than to VIP. The inhibitory effect of PACAP on prostanoid synthesis is mediated via a cAMP-dependent pathway. By inhibiting the formation of vasoactive prostanoids, PACAP can decrease the vasoreactivity of the microvessels. PMID- 10519977 TI - Change point in VCO2 during incremental exercise test: a new method for assessment of human exercise tolerance. AB - The main purpose of this study was to present a new method to determine the level of power output (PO) at which VCO2 during incremental exercise test (IT) begins to rise non-linearly in relation to power output (PO) - the change point in VCO2 (CP-VCO2). Twenty-two healthy non-smoking men (mean +/- SD: age 22.0 +/- 0.9 years; body mass 74.5 +/- 7.5 kg; height 181 +/- 7 cm; VO2max 3.753 +/- 0.335 l min-1) performed an IT on a cycloergometer. The IT started at a PO of 30 W, followed by gradual increases of 30 W every 3 min. Antecubital venous blood samples were taken at the end of each step and analysed for plasma lactate concentration [La]pl, blood PO2, PCO2 [HCO3-]b and [H+]b. In the detection of the change-point VCO2 (CP-VCO2), a two-phase model was assumed for the 'third-minute data' of each step of the test. In the first phase, a linear relationship between VCO2 and PO was assumed, whereas in the second, an additional increase in VCO2 was allowed, above the values expected from the linear model. The PO at which the first phase ends is called the change point in VCO2. The identification of the model consists of two steps: testing for the existence of the change point, and estimating its location. Both procedures are based on suitably normalized recursive residuals (see Zoladz et al. 1998a. Eur J Appl Physiol 78, 369-377). In the case of each of our subjects it was possible to detect the CP-VCO2 and the CP VO2 as described in our model. The PO at the CP-VCO2 amounted to 134 +/- 42 W. The CP- VO2 was detected at 136 +/- 32 W, whereas the PO at the LT amounted to 128 +/- 30 W and corresponded to 49 +/- 11, 49 +/- 8 and 47 +/- 8.6% VO2max, respectively, for the CP-VCO2, CP-VO2 and the LT. The [La]pl at the CP-VCO2 (2.65 +/- 0.76 mmol L-1), at the CP-VO2 (2.53 +/- 0. 56 mmol L-1) and at the LT (2.25 +/- 0.49 mmol L-1) were already significantly higher (P < 0.01, Students t-test) than the value reached at rest (1.86 +/- 0.43 mmol L-1). Our study illustrates that the CP-VCO2 and the CP-VO2 occur at a very similar power output as the LT. We therefore postulate that the CP-VCO2 and the CP-VO2 be applied as an additional criterion to assess human exercise tolerance. PMID- 10519978 TI - Maximal strength and power characteristics in isometric and dynamic actions of the upper and lower extremities in middle-aged and older men. AB - Muscle cross-sectional area of the quadriceps femoris (CSAQF), maximal isometric strength (handgrip test and unilateral knee extension/flexion), the shape of isometric force-time curves, and power-load curves during concentric and stretch shortening cycle (SSC) actions with loads ranging from 15 to 70% of one repetition maximum half-squat (1RMHS) and bench-press (1RMBP) were examined in 26 middle-aged men in the 40-year-old (M40) (mean age 42, range 35-46) and 21 elderly men in the 65-year-old age group (M65) (mean age 65, range 60-74). Maximal bilateral concentric (1RMHS and 1RMBP), unilateral knee extension (isometric; MIFKE and concentric; 1RMKE) strength and muscle CSA in M65 were lower (P < 0.001) than in M40. The individual values of the CSAQF correlated with the individual values of maximal concentric 1RMHS, 1RMKE and MIFKE in M65, while the corresponding correlations were lower in M40. The maximal MIFKE value per CSA of 4.54 +/- 0.7 N m cm-2 in M40 was greater (P < 0. 05-0.01) than that of 4.02 +/ 0.7 N m cm-2 recorded in M65. The maximal rate of force development of the knee extensors and flexors in M65 was lower (P < 0.01-0.001) and the heights in squat and counter-movement jumps as much as 27-29% lower (P < 0.001) than those recorded in M40. M65 showed lower (P < 0.001) concentric power values for both upper and lower extremity performances than those recorded for M40. Maximal power output was maximized at the 30-45% loads for the upper extremity and at the 60 70% loads for the lower extremity extensors in both age groups. Muscle activation of the antagonists was significantly higher (P < 0.01-0.001) during the isometric and dynamic knee extension actions in M65 than in M40. The present results support a general concept that parallel declines in muscle mass and maximal strength take place with increasing age, although loss of strength may vary in both lower and upper extremity muscles in relation to the type of action and that ageing may also lead to a decrease in voluntary neural drive to the muscles. Explosive strength and power seem to decrease with increasing age even more than maximal isometric strength in both actions but power was maximized at the 30-45% loads for the upper and at the 60-70% loads for the lower extremity action in both age groups. High antagonist muscle activity may limit the full movement efficiency depending on the type of muscle action, testing conditions and the velocity and/or the time duration of the action, especially in the elderly. PMID- 10519979 TI - Calphostin C is an inhibitor of contraction, but not insulin-stimulated glucose transport, in skeletal muscle. AB - Wortmannin selectively impairs insulin-stimulated glucose transport in skeletal muscle. To search for an inhibitor specific for contraction-stimulated glucose transport, we screened a number of calmodulin and PKC inhibitors for their ability to impair contraction- and insulin-stimulated 2-deoxyglucose uptake in incubated rat soleus muscles. In concentrations that did not reduce contraction induced force output, among calmodulin inhibitors W-7 inhibited both contraction- and insulin-stimulated glucose transport by up to 50% (P < 0.05), while Calmidazolium impaired only insulin-stimulated glucose transport (P < 0.05), and Trifluoperazine and Phenoxybenzamine did not influence glucose transport. In concentrations that did not reduce force generation, among PKC inhibitors Calphostin C specifically inhibited contraction-stimulated glucose transport (P < 0.05), whereas insulin-stimulated transport was impaired by Rottlerin and Bisindolylmaleimide I (P < 0.05), and both contraction- and insulin-stimulated glucose transport were inhibited by RO-31-8220 (P < 0.05). Calphostin C did not reduce contraction-induced increase in AMP-activated protein kinase (AMPK) activity. In conclusion, we have identified specific inhibitors of both contraction- and insulin-stimulated glucose transport. Both calmodulin and different isoenzymes of the PKC family may be involved in contraction- and insulin-stimulated glucose transport. Calphostin C does not influence glucose transport during contractions via stimulation of AMPK. Calphostin C may be used to unravel signal transduction in contraction-stimulated glucose transport. PMID- 10519980 TI - Attenuated insulin action on glucose uptake and transport in muscle following resistance exercise in rats. AB - A previous study reported elevations of insulin-mediated muscle protein synthesis following four days of resistance exercise in rats (Fluckey et al. 1996. Am J Physiol 270, E313-E319). The purpose of this study was to determine if insulin stimulated muscle glucose uptake (a-v diff.) and 2-deoxyglucose (2-DG) transport were altered under similar conditions. The protocol consisted of squat-like exercises during four sessions with progressively increased weight (70-190 g). Each session consisted of 50 repetitions and sessions were separated by 48 h. Sixteen hours after the last exercise session, basal glucose uptake in perfused hindlimbs was not different (P > 0.05) between exercised (n=6) and non-exercised controls (n=6). However, there was a significant (P < 0.05) attenuation of insulin-stimulated (20 000 microU mL-1) glucose uptake in exercised vs. non exercised rats (491 +/- 31 vs. 664 +/- 58 micromol glucose-1 g-1 [15-min insulin period]-1, respectively). Following resistance exercise, insulin-stimulated 2-DG transport, measured during the last 10 min of the perfusion period, was significantly reduced (P < 0.05) in the soleus, white gastrocnemius and extensor digitorum longus muscles. Additionally, GLUT-4 glucose transporter protein content was significantly reduced (P < 0.05) in white gastrocnemius and extensor digitorum longus muscles. These results demonstrate that insulin-stimulated glucose uptake and transport are reduced after resistance exercise. Furthermore, the applied resistance exercise protocol causes directionally opposite changes of insulin action in two major metabolic pathways, i.e. glucose transport and protein synthesis. PMID- 10519981 TI - Mechanisms associated with hypoxia- and contraction-mediated glucose transport in muscle are fibre-dependent. AB - The purpose of this study was to examine the effects of hypoxia and muscle contractions on rates of 2-deoxyglucose (2-DG) transport in red and white portions of the gastrocnemius muscle of the rat. 2-DG transport was measured during the last 10 min of a 60-min hindlimb perfusion in male Wistar rats ( approximately 300 g), with or without muscle contractions of one limb. The medium was gassed with either 95% oxygen and 5% carbon dioxide or 95% nitrogen and 5% carbon dioxide to achieve normal or hypoxic conditions, respectively. Muscle contractions began after 30 min of perfusion and consisted of isometric muscle actions (200-ms trains, 100 Hz; one train per second) for two sets of 5 min, with 1-min rest between sets. 2-DG transport in white gastrocnemius was higher (P < 0.05) than basal during hypoxia (4.8-fold) and following contractions using oxygenated or hypoxic medium (4.6-fold and 5.4-fold, respectively; n=6 for each group). 2-DG transport was not different (P > 0.05) between these stimulated conditions. Similarly, 2-DG transport in red gastrocnemius was 5.1- and 4.8-fold higher (P < 0.05) than basal during hypoxia and following contractions in oxygenated medium, respectively. However, 2-DG transport following contractions during hypoxic conditions in red gastrocnemius was, unlike white gastrocnemius, higher (8.9-fold over basal; P < 0.05) than in all other conditions. These results suggest that mechanisms associated with hypoxia- and muscle contraction mediated glucose transport are fibre type-dependent, with additive effects of the two stimuli in fast-twitch, oxidative fibres. PMID- 10519982 TI - 5q--syndrome. PMID- 10519983 TI - The impact of new-variant Creutzfeldt-Jakob disease on blood transfusion practice. PMID- 10519984 TI - Management of relapsed acute myeloid leukaemia. PMID- 10519985 TI - Cardioprotection. PMID- 10519986 TI - Origin of mutation in sporadic cases of haemophilia A. AB - The aim of this study was to define the origin of mutation in sporadic cases of severe haemophilia A. The series was composed of 31 families with sporadic severe haemophilia A in the geographical catchment area of the Malmo haemophilia centre. The mutation was characterized in 29/31 families: inversion type 1 (n = 11), inversion type 2 (n = 3), other inversion (n = 1), small or partial deletion (n = 6), insertion (n = 2), non-sense mutation (n = 4) and mis-sense mutation (n = 2). Of 29 probands, eight carried a de novo mutation, whereas the proband's mother was found to carry the mutation in 21/29 families. Of the 21 carrier mothers, 16 had de novo mutations (i.e. the proband's maternal grandfather and grandmother were non-carriers). Owing to the lack of samples from the grandparents, origin could not be determined in the remaining five families. Polymorphisms of the FVIII gene were used to determine whether the de novo mutation of the carrier mother was of paternal or maternal origin. In 15/16 cases the mutation was of paternal origin and in 1/16 cases of maternal origin. In the series as a whole, mutation frequency was 6-fold higher in males than in females, but no differences in the ratio of sex-specific mutations rates was found among different types of mutation. PMID- 10519987 TI - Pathological events in platelets of Wiskott-Aldrich syndrome patients. AB - The Wiskott-Aldrich syndrome (WAS) is a severe X-linked platelet/immunological disorder arising from mutations of the gene WASP. At the clinical level, the major platelet abnormalities are small size and low number, both partially correctable by splenectomy. To identify underlying pathological events, we examined WAS platelets at various stages of their lifetime. In spleen sections from WAS patients, fluorescence microscopy showed dramatic co-localization of markers of platelets (CD41) and macrophages (CD68) compared to non thrombocytopenic controls, suggesting that WAS splenic macrophages are involved in platelet removal. Study of isolated WAS blood platelets by flow cytometry showed substantial enhancement of surface exposure of phosphatidylserine (PS), a signal for engulfment by macrophages. Isolated resting WAS platelets were also aberrantly susceptible to microparticle release, and plasma samples of WAS patients contained > 5 times normal numbers of platelet-derived microparticles which may explain the small size of circulating platelets. Measurements with the Ca2+ sensitive dye fluo-3 revealed significantly increased Ca2+ levels, 310 +/- 13 nmol/l for WAS platelets versus 106 +/- 12 nmol/l for normal platelets, and also prolongation of agonist-induced Ca2+ flux. Cumulatively, these studies identify abnormal events occurring in WAS platelets: increased Ca2+ levels and enhancement of two Ca2+ dependent processes, PS exposure and microparticle release; these abnormal events may contribute to the in vivo decrease of platelet number and reduction of platelet size in this disease. PMID- 10519988 TI - Age-related changes in thrombopoietin in children: reference interval for serum thrombopoietin levels. AB - We studied thrombopoietin (TPO, Mpl ligand) values using a sensitive ELISA in 254 serum samples obtained from disease-free children and adult volunteers. TPO was detected in all samples, and its values ranged widely from 0.25 to 9.18 fmol/ml. When analysed by dividing the subjects into 11 age groups, the mean TPO levels from birth to 1 month of age were increased (3.73-5.92 fmol/ml). The highest values were found 2 d after birth; TPO levels then gradually decreased to adult levels (0.83 fmol/ml). The relationship between TPO values and platelet counts was not significant in all subjects (r = 0.27) or in children alone (r = 0.12). In children > 1 month of age a 95% reference interval for serum TPO values was determined from 0.58 to 3.27 fmol/ml. A significant correlation was found between TPO values in serum and plasma; serum TPO values = -0.257 + 4.039 x plasma TPO values (r = 0.951, P < 0.001, n = 22). This study is the first to report age dependent changes in blood TPO levels throughout child development. Serum TPO values were significantly high up to 1 month of age and were correlated with plasma TPO levels. PMID- 10519989 TI - Influence of three potential genetic risk factors for thrombosis in 43 families carrying the factor V Arg 506 to Gln mutation. AB - The factor V (FV) Arg 506 to Gln mutation is the most common abnormality observed in familial thrombophilia. Many studies have shown that its clinical expression differs among families and among carriers. Some thrombotic patients carry an additional genetic risk factor such as protein C, protein S or antithrombin deficiency. We sought to identify other genetic risk factors potentially favouring expression of the thrombotic phenotype in 370 members of 43 families with the FV Arg 506 to Gln mutation. We analysed three candidate polymorphisms in genes involved in the PC anticoagulant pathway, consisting of two polymorphic sites in the 5' non-transcribed region of the PC gene, -1654 C/T and -1641 A/G, with three known combinations (TA, CA and CG) that influence the protein C plasma level; one polymorphic site (4070 A/G) in exon 13 of the FV gene, which influences the plasma factor V concentration, and one polymorphic site (677 C/T) in the methylenetetrahydrofolate reductase gene, which is often associated with moderate hyperhomocysteinaemia. The distribution of these different polymorphisms was similar in patients with a history of thrombosis and those who remained asymptomatic, ruling out the possibility that each of these polymorphisms alone can play a role in the onset of thrombosis in carriers of the FV Arg 506 to Gln mutation. PMID- 10519990 TI - Standardization of the PFA-100(R) platelet function test in 105 mmol/l buffered citrate: effect of gender, smoking, and oral contraceptives. AB - The PFA-100(R) (PFA) diagnostic system for the detection of platelet dysfunction was evaluated to determine reference ranges in a normal population. The PFA determines the primary haemostasis capacity (PHC) of anticoagulated whole blood, expressed by the system's closure time (CT). In this study the CT reference ranges were determined for blood samples collected in 105 mmol/l (3.2%) buffered citrate and the effect of gender, smoking, and use of oral contraceptives on reference ranges was assessed. Each of the 309 healthy blood donors from five blood centres was confirmed to have normal platelet function before inclusion in the study. Blood samples were tested in duplicate with both the collagen/epinephrine (Col/Epi) and collagen/ADP (Col/ADP) test cartridges. PFA reference ranges (90% central intervals of measured closure times) for both cartridge types were similar for all groups. Subgroup analysis showed that neither gender nor oral contraceptive usage had any effect on PHC. The 95% cut off value for the Col/Epi CT was slightly higher for smokers than for non smokers, an effect more pronounced in female than in male donors. However, the small difference did not justify establishment of specific reference ranges for smokers. Data from all included subjects were pooled to calculate the CT reference ranges for blood samples collected in 105 mmol/l buffered citrate (Col/Epi 82-150 s; Col/ADP 62-100 s). Normal levels of fibrinogen, as well as normal platelet counts and normal haematocrit levels, appeared not to influence the PHC. Because slight but significant differences of the reference ranges were observed between some of the participating sites, in-house confirmation of these reference range guidelines is recommended. PMID- 10519991 TI - Selective secretion of chemoattractants for haemopoietic progenitor cells by bone marrow endothelial cells: a possible role in homing of haemopoietic progenitor cells to bone marrow. AB - To elucidate the mechanisms by which haemopoietic progenitor cells lodge in the bone marrow, we examined the secretion of chemoattractants for haemopoietic progenitor cells by bone marrow and lung endothelial cells. The bone marrow endothelial cells, but not lung endothelial cells, secreted chemoattractants for the haemopoietic progenitor cell line, FDCP-2, and normal haemopoietic progenitor cells. Checkerboard analysis demonstrated that the conditioned medium of the bone marrow endothelial cells had chemotactic activity and random motility-stimulating activity. The bone marrow endothelial cells expressed stromal-cell-derived factor 1 (SDF-1) mRNA and produced SDF-1 protein, whereas the lung endothelial cells did not. Adhesion of FDCP-2 cells to the bone marrow endothelial cells was partially inhibited by anti-SDF-1 antibody. These findings suggest that the chemoattractants for haemopoietic progenitor cells including SDF-1 and random motility-stimulating factor(s) selectively secreted by the bone marrow endothelial cells may contribute to the homing of haemopoietic progenitor cells to bone marrow. PMID- 10519992 TI - Purification, amplification and characterization of a population of human erythroid progenitors. AB - In humans, studies of the erythroid cell lineage are hampered by difficulties in obtaining sufficient numbers of erythroid progenitors. In fact, these progenitors in bone marrow or peripheral blood are scarce and no specific antibodies are available. We describe a new method which allows proliferation in liquid culture of large numbers of pure normal human erythroid progenitors. CD34+ cells were cultured for 7 d in serum-free conditions with the cytokine mixture interleukin (IL)-3/IL-6/stem cell factor (SCF). This resulted in cell expansion and the appearance of a high proportion of CD36+ cells which were purified on day 7. Methylcellulose clones from these cells were composed of 96.6% late BFU-E and 3.4% CFU-GM. These CD36+ cells could be recultured with the same cytokine mixture plus or minus erythropoietin (Epo) for a further 2-7 d. In both conditions further amplification of CD36+ cells was observed, but Epo induced a more dramatic cell expansion. Glycophorin-positive mature cells appeared only in the presence of Epo, and terminal red cell differentiation was observed after 7 d of secondary culture. Cells obtained from adult CD34+ progenitors mostly contained adult haemoglobin, whereas cord blood-derived cells contained equal proportions of adult and fetal haemoglobin. Activation of STAT5 and tyrosine phosphorylation of the Epo receptor and JAK2 were observed after Epo stimulation of these cells. This new method represents a straightforward alternative to the procedures previously described for the purification of normal erythroid progenitors and is useful in the study of erythropoietic regulation. PMID- 10519993 TI - Role of FcgammaRI (CD64) in erythrocyte elimination and its up-regulation in thalassaemia. AB - To examine any role of the high affinity Fcgamma class I receptor (FcgammaRI) (CD64) in erythrocyte elimination by mononuclear phagocytes (MP) in thalassaemia (thal), we investigated the in vitro interaction of beta-thalassaemic erythrocytes with monocytes (Mo) whose FcgammaR expression had been modulated by cytokines. Treatment of Mo with interferon (IFN)-gamma or interleukin (IL)-10 which up-regulate FcgammaRI, caused a dose-dependent increase in binding of beta thalassaemic erythrocytes, whereas stimulation with IL-4 which down-regulates the receptor, reduced this interaction, in a dose-dependent manner, to that of normal erythrocytes. Binding of thalassaemic erythrocytes by IFN-gamma or IL-10-treated Mo was inhibited by FcgammaRI-specific reagents. In addition, Mo expression of FcgammaRI and HLA class II DR was determined by flow cytometry in Thai patients with HbH disease (alpha1/alpha2 or alpha1/Hb Constant Spring) (n = 15) or beta degrees -thal/HbE (n = 16). In both groups of patients FcgammaRI expression was increased as compared to normal controls (n = 14): mean fluorescence intensity (+/-SD) 124.79 +/- 38. 77 in HbH disease and 121.86 +/- 18.23 in beta degrees thal/HbE versus 91.94 +/- 17.36 in normal controls (P < 0.01 and P < 0.001, respectively). In contrast, HLA class II DR expression was similar in patients and controls. The results suggest that, in thalassaemia, up-regulated FcgammaRI on mononuclear phagocytes plays a role in their interaction with erythrocytes and that this process can be modified by cytokines. PMID- 10519994 TI - A novel mutation of delta-aminolaevulinate dehydratase in a healthy child with 12% erythrocyte enzyme activity. AB - Cloning, expression and phenotype studies of the defective gene for delta aminolaevulinate dehydratase (ALAD) in a family with an asymptomatic girl who had ALAD deficiency were carried out. The proband was identified by neonatal ALAD screening, and had erythrocyte ALAD activity at 12% of the normal control. She was heterozygous for ALAD deficiency, which was inherited from her father. Nucleotide sequence analysis of the cloned ALAD cDNA revealed C36 to G and T168 to C mutations on the same allele. The former mutation resulted in F12L substitution, whereas the latter was a silent mutation. All family members who had decreased ALAD activity had the same mutation. Expression of the mutant ALAD cDNA in Chinese hamster ovary cells produced an ALAD protein without significant enzyme activity. Additionally, the mutant ALAD cDNA which encodes F12L substitution produced an aberrant migration pattern in polyacrylamide gel electrophoresis under denaturing conditions. This finding probably reflects an abnormal folding of the F12L protein, since the mutation occurred in the alpha1 helix of the N-terminal arm of the enzyme, which is involved in the extensive quaternary interactions among the subunits. This is also the first report of ALAD gene mutation in an asymptomatic subject. PMID- 10519995 TI - An in vitro system for expression analysis of mutations of the beta-globin gene: validation and application to two mutations in the 5' UTR. AB - We describe the setting up of an in vitro expression system for the analysis of mutations of the beta-globin gene. The system is based on the stable transfection of a normal or mutated beta-globin gene into mouse erythroleukaemia (MEL) cells. The expression construct contains an Agamma gene as an internal control and both globin genes are under the control of the HS2 element of the beta LCR. The system enables analysis of transcription, RNA processing and transport, as well as mRNA stability. With non-mutant genes, high-level expression of both beta and Agamma genes is seen and both mRNAs are stable. The system was validated by comparing the expression of the beta654 thalassaemia splicing mutation in MEL cells with its well-characterized expression in vivo. The level of the initial transcript, the proportion of abnormally spliced mRNA and its instability during erythroid cell maturation were all faithfully reproduced. The system was used to examine the mechanism by which two mutations in the beta-globin 5' untranslated region (5' UTR) result in beta thalassaemia. Surprisingly, the mechanism appeared to differ in the two cases, with the C-G substitution at position +33 affecting transcription, whereas the -T deletion at position +10 resulted in a translational defect. The stably transfected MEL cells, with an internal control and an endogenous enhancer, appear to be a valid and realistic experimental model, superior to transient expression studies. This system should find wide application in the analysis of the effects and mechanisms of gene inactivation in mutations affecting the beta-globin as well as other genes. PMID- 10519996 TI - Suppression of CDA II expression in a homozygote. AB - The CDAN2 gene, responsible for congenital dyserythropoietic anaemia, type II (CDA II), was recently mapped to 20q11.2. We report data on an additional member of a previously studied CDA II family. This member had always been regarded as haematologically normal. Unexpectedly, she had the same microsatellite assortments around the CDAN2 alleles as her three sisters with CDA II. In particular, she was a homozygote for microsatellites D20S863 and D20S841. This prompted an analysis of all facets of her phenotype. The Ham test was negative. The bone marrow smears contained a normal proportion of binucleate erythroblasts. Electron microscopy revealed the absence of extensive stretches of cisternae beneath and parallel to the inner surface of the erythroblast plasma membrane. Proteins of the endoplasmic reticulum, which contaminate the reticulocyte plasma membrane in CDA II patients, were missing. Only the shape of the band 3 peak appeared slightly altered. This case exemplifies that homozygosity (or compound heterozygosity) for a deleterious gene may be silenced, or almost completely silenced. In recessively inherited diseases, suppressed phenotypes tend to be overlooked in siblings where both patients and unaffected individuals are expected. PMID- 10519997 TI - Persistent leucocyte abnormalities in children years after previous long-term low dose radiation exposure. AB - Two hundred and eighty-nine children who had been educated in a kindergarten during 1983-92 and received continuous low-dose whole body gamma-irradiation from 60Co-contaminated steel window frames in their classrooms were investigated for residual effects on their haematological tissues. Another 751 children, sex and age-matched, received similar but much lower exposure in an elementary school with classrooms built with contaminated steel rebars. The peripheral leucocytes of these children were examined 5-7 years after they had stopped using these irradiated classrooms. Children who received higher exposure in the kindergarten were shown to have a significant decrease in total leucocytes and neutrophils and an increase in eosinophils. Moreover, they were shown to be at significantly higher risk of developing relative leucopenia and neutropenia, but not lymphocytopenia, than those who received a lower exposure at the elementary school. Children from the kindergarten who had much higher exposure were shown to have a significant lowering of total leucocytes and neutrophils, and an increase in eosinophils years after exposure. Residual adverse haematological effects on the exposed children are strongly suspected. PMID- 10519998 TI - A long-term study of patients with chronic natural killer cell lymphocytosis. AB - Chronic natural killer cell lymphocytosis is a persistent state of natural killer (NK) cell (CD3-CD16/CD56+) excess in the peripheral blood that is not associated with clinical lymphoma. In 16 consecutive patients (median age 60.5 years, range 7-77), males were overrepresented (M:F 7:1) and the median absolute NK cell count was 4.09 x 10(9)/l (range 1.2-16.6). Bone marrow examination was performed in 14 patients and showed atypical granulomata in two; chromosome studies in seven patients were normal. Clonal T-cell receptor gene rearrangement was not found in any of 12 patients evaluated. At presentation, seven patients (44%) had no clinical symptoms or signs and the others had vasculitic skin lesions (three patients), non-neutropenic fever (three patients), recurrent neutropenic infection (two patients), musculoskeletal symptoms (two patients), peripheral neuropathy (two patients), aphthous ulcers (one patient), and splenomegaly (one patient). Five patients had anaemia, five had neutropenia, and two had thrombocytopenia. After a median follow-up of 5.1 years (range 0-10.2) from immunophenotypic diagnosis or 5.7 years (range 0.1-14.1) from documentation of absolute lymphocytosis, vasculitic glomerulonephritis developed in one patient, accelerated splenomegaly developed in a patient receiving myeloid growth factor treatment, and severe aplastic anaemia developed in one patient. Treatment with nonsteroidal anti-inflammatory drugs or immunosuppressive agents was variably successful. PMID- 10519999 TI - Results at a single centre of immunosuppression with cyclosporine A in 66 children with aplastic anaemia. AB - A retrospective analysis of cyclosporine A (CsA) monotherapy administered to 66 children with aplastic anaemia (AA) at a single centre was carried out. The study was conducted on 66 children (F34/M32) with a median age of 10.5 years. 30 children (45%) achieved complete or partial remission within a median of 8 weeks. The response rate was 3/19 (16%) in cases of very severe aplastic anaemia (vSAA), 16/34 (47%) in severe aplastic anaemia (SAA) and 11/13 (85%) in nonSAA. 10 remitters (33%) relapsed after CsA cessation or dose tapering and six of those responded again on CsA alone. The only variable predictive for response was granulocyte count before treatment. The actuarial probability of survival was 51% at 4 years (85% in moderate AA, 50% in SAA and 32% in vSAA). The optimal dosage of CsA has yet to be established. PMID- 10520000 TI - TGF-beta is not the principal immunosuppressive component in coagulation factor concentrates. AB - Coagulation factor concentrates are known to inhibit a variety of immune reactions when assessed in vitro. This study assessed the immunomodulatory activity of a wide range of coagulation factor concentrates by measuring their inhibition of PHA-stimulated lymphocyte proliferation and reduction in IL-2 secretion. The hypothesis that TGF-beta is responsible for most of these effects was tested by measuring biologically active TGF-beta and immunoreactive TGF-beta1 in the concentrates and comparing the levels recorded with immunosuppressive activity. In addition, the coagulation factors were compared directly with a standard preparation of TGF-beta in a TGF-beta-specific bioassay and in lymphocyte proliferation assays. Although there was a broad correlation between levels of total or active TGF-beta and immunosuppressive activity across all of the coagulation factors tested, individual data sets showed clear discrepancies. Implying that TGF-beta probably serves as a surrogate marker for other immunomodulatory contaminants and that neither TGF-beta nor any other single substance could account for all of the immunosuppressive activity observed. Furthermore, there was a difference of more than 100-fold in the relative potencies of coagulation factors and pure TGF-beta, when compared in immunosuppression assays, indicating that the different assays did not measure the same substance. Whereas anti-TGF-beta antibody almost completely blocked the activity of coagulation factor concentrates (TGF-beta-specific bioassay) and abrogated the effect of authentic TGF-beta (immunosuppression assays) at high concentrations it achieved <50% reversal of the immunosuppressive effects of coagulation factors in immunosuppression assays. These findings indicated that TGF-beta accounted for only a minor proportion of the immunosuppressive activity in most coagulation factor concentrates. PMID- 10520001 TI - Abnormalities of the p53 gene in juvenile myelomonocytic leukaemia. AB - Juvenile chronic myelomonocytic leukaemia (JMML) is a rare myeloproliferative disorder of childhood. Fewer than 30% of cases of JMML terminate in a blast crisis; however, its molecular mechanism is unknown. Since mutation and/or deletion of the p53 gene has been reported to be associated with disease progression in a wide variety of human cancers, including adult-type chronic myelogenous leukaemia, we studied the p53 gene in 20 patients with JMML (16 samples in chronic phase and seven at blast crisis). Exons 4-8 of the p53 gene, which cover all the hot spots of point mutations, were amplified by the polymerase chain reaction (PCR) method and subjected to mutation screening by single-strand conformation polymorphism analysis. No mobility shift of single strand DNA of PCR products in polyacrylamide gel electrophoresis, indicating point mutations, was found in 19/20 patients. DNA of the remaining patient in the chronic phase failed to be amplified by PCR and Southern blot analysis with XbaI digested genomic DNA revealed a gross rearrangement (presumed deletion) of the p53 gene. These data indicate that abnormalities of the p53 gene are rare in JMML and not responsible for acute transformation, but could be involved in the pathogenesis of some cases of JMML. PMID- 10520002 TI - Increased serum levels of beta2m-free HLA class I heavy chain in multiple myeloma. AB - Serum levels of beta2-microglobulin (beta2m)-free HLA class I heavy chain (FHC) in 94 patients with multiple myeloma (MM) were higher than in 29 patients with monoclonal gammopathy of undetermined significance (MGUS) (P = 0.023) and in 97 sex- and age-matched healthy controls (P < 0.0001). Spearman correlation analysis indicated that in MM, FHC correlated with beta2m (r = 0.31, P = 0. 003) and the percentage of bone marrow plasma cells (BMPC%) (r = 0. 36, P = 0.002), whereas beta2m, in addition to BMPC% (r = 0.43, P = 0.0003), also correlated with creatinine levels (r = 0.63, P < 0.0001), haemoglobin levels (r = -0.35, P = 0.0007) and patient age (r = 0.34, P < 0.0011). Furthermore, MM patients with poor prognosis (beta2m >/= 6 mg/l) displayed higher FHC levels than those with a better prognosis (beta2m < 6mg/l) (P < 0.021). At variance from beta2m, these levels were not influenced by renal failure, as indicated by the lack of Spearman correlation of FHC with creatinine concentration and of statistical significance between the median FHC concentration of MM patients with creatinine < 176.6 micromol/l and those with creatinine >/= 176.6 micromol/l (P = 0.3). Stratification of patients according to disease activity and stage showed that FHC levels were only statistically different (P = 0.04) for disease activity, whereas beta2m and C-reactive protein were not. Taken together, our data indicate that serum FHC may be a useful disease marker in MM. PMID- 10520003 TI - Bryostatin and CD40-ligand enhance apoptosis resistance and induce expression of cell survival genes in B-cell chronic lymphocytic leukaemia. AB - Modulating signal transduction pathways represents a promising approach for altering the biological behaviour of haemopoietic malignancies. B-cell chronic lymphocytic leukaemia (B-CLL) cells were treated in vitro with CD40-ligand (CD40L) (CD154) or the protein kinase C modulator Bryostatin-1, exploring the effects on: (a) sensitivity to apoptosis induction by chemotherapeutic drugs (fludarabine, dexamethasone) or anti-Fas antibody; (b) expression of apoptosis regulatory proteins (Bcl-2, Bcl-X, Mcl-1, Bax, Bak, BAG-1, Flip, XIAP); (c) expression of cell surface co-stimulatory antigens (CD80 [B7.1]; CD54 [ICAM-1]; CD70); and (d) expression of immune modulatory receptors (CD27, CD40, CD95 [Fas]). CD40L and Bryostatin decreased both spontaneous and drug-induced apoptosis in most B-CLL specimens tested. Apoptosis resistance was associated with CD40L- and Bryostatin-induced elevations in the anti-apoptotic Bcl-2 family protein Mcl-1. CD40L also induced striking increases in the levels of the anti apoptotic protein Bcl-XL in B-CLLs. CD40L stimulated increases in the surface expression of CD40, CD54, CD69, CD70, CD80 and CD95, whereas Bryostatin induced expression of CD40, CD54, CD69 and CD95 but not the co-stimulatory molecules CD70 and CD80. Despite elevations in the expression of CD95 (Fas), anti-Fas antibodies failed to induce apoptosis of CD40L- and Bryostatin-treated B-CLL cells. This Fas resistance was associated with increased expression of the Fas-antagonist Flip in CD40L-treated, and with elevations in the caspase inhibitor XIAP in Bryostatin treated B-CLLs. The potential anti-apoptotic properties of CD40L and Bryostatin should be taken into consideration when employing these agents in clinical trials involving patients with B-CLL. PMID- 10520004 TI - Prognostic evaluation in multiple myeloma: an analysis of the impact of new prognostic factors. AB - We have analysed the prognostic information for survival of presenting features in an unselected series of 394 myeloma patients. 15 variables with significant prognostic information were identified, among these were some not previously or only recently reported: serum levels of hepatocyte growth factor (HGF), interleukin-6 (IL-6), C-terminal cross-linked telopeptide of collagen I (ICTP) and soluble interleukin-6 receptor (sIL-6R). In a multivariate Cox analysis six variables were significantly and independently associated with poor survival: high age, low W.H.O.-performance status (PS), high serum levels of calcium, beta 2-microglobulin (beta-2M), IL-6 and sIL-6R. A risk score formed to predict survival for each percentile of the patient population allowed an efficient separation of prognostic groups. The discriminating power of the model compared favourably with three other previously published staging systems applied to the study population. Exclusion of IL-6 and sIL-6R from the model only marginally decreased the efficacy of the separation. The predictive value of some variables (sIL-6R, beta-2M and W.H.O.-PS) decreased significantly over time. We conclude that formation of a risk score based on independent variables is an efficient way to separate prognostic groups, that the contribution of new and not easily available parameters should be thoroughly evaluated before inclusion in prognostic models for clinical use and that the predictive value of parameters may decrease over time. PMID- 10520005 TI - Efficacy and toxicity of IFN-alpha2b combined with cytarabine in chronic myelogenous leukaemia. AB - Newly diagnosed chronic myelogenous leukaemia (CML) patients (n = 65) were treated with interferon (IFN)-alpha2b (5 x 106 IU/d s.c.) combined with monthly courses of cytarabine (20 mg/d s.c. for 14 d). Median age of patients enrolled was 45 years. The endpoints of the study were clinical efficacy and toxicity. The survival rates at 3 years and 5 years were 77% and 56%, respectively. The rate of complete haematological response was 60%. Evaluation of cytogenetic response was available in 29/65 patients. A complete cytogenetic response was seen in 3/29 patients (10%). W.H.O. toxicity grade 3-4 occurred in only 22/523 evaluable treatment cycles. Since the study protocol required intermittent or definitive discontinuation of cytarabine in case of moderate leucopenia (white blood cells (WBC) <5 x 109/l), combined cytopenia (WBC < 5 x 109/l, platelets <100 x 109/l), and isolated moderate thrombocytopenia (<100 x 109/l), the drug had to be discontinued temporarily or definitively in 200 cycles and the dose of cytarabine had to be reduced in 35 cycles. Thus, only 25% of the planned dose of cytarabine could be administered. At this dosage it would appear that cytarabine had no effect on survival and did not improve remission rates. We conclude that a clinical benefit for the addition of cytarabine to the treatment of CML with IFN might only be achieved by the administration of a higher cumulative dose of cytarabine, suggesting that lower leucocyte counts of 2-4 x 109/l have to be tolerated. PMID- 10520006 TI - Secondary leukaemia and myelodysplasia after autografting for lymphoma: results from the EBMT. EBMT Lymphoma and Late Effects Working Parties. European Group for Blood and Marrow Transplantation. AB - Between 1978 and 1996 more than 7500 lymphoma transplants have been reported to the European Bone Marrow Transplantation (EBMT) Lymphoma Registry. This has been examined to establish the incidence of secondary leukaemia and myelodysplasia and to relate this to possible prognostic factors. 131 centres representing 4998 patients responded to a questionnaire. This identified 66 patients with post transplant myelodysplastic syndrome (MDS)/acute myeloid leukaemia (AML). The actuarial risk for MDS/AML at 5 years post-transplant (+/-95% CI) was 4.6% (3.1 6.8) for Hodgkin's disease and 3.0% (2.0-4. 3) for non-Hodgkin's lymphoma. Multivariate analysis for all patients demonstrated an effect of age at transplant, radiotherapy at conditioning, number of transplants and interval between diagnosis and transplant as risk factors. For patients with NHL, grade of histology was important (low grade > intermediate or high-grade); for Hodgkin's disease, female sex was identified as a risk factor. These findings suggest that the incidence of MDS/AML may not be greater following an autograft than after conventional chemotherapy. PMID- 10520007 TI - A population-based study of childhood myelodysplastic syndrome in British Columbia, Canada. AB - Myelodysplastic syndrome (MDS) is considered to be very rare in children. However, the only two published population-based studies reported widely divergent incidence figures. To further explore the epidemiology of childhood MDS and to evaluate the accuracy of cancer registry and treatment trial data, we conducted a population-based study of children aged 0-14 years in British Columbia (BC), Canada, between 1982 and 1996. MDS was diagnosed in 31 cases corresponding to an annual incidence of 3.2 per million children or 6% of all leukaemias, compared with an incidence of 6.0/million for acute myeloid leukaemia (AML), and of 0.5/million for chronic myeloid leukaemia. There was a non significant (P = 0.19) trend toward an increase in MDS incidence with time, the increase was partly explained by an increasing number of patients with Down syndrome. Associated abnormalities were found in 48% of the MDS cases with Down syndrome as the most common (seven cases). Only one third of the MDS cases were correctly registered in the Cancer Registry and less than half of the eligible MDS patients were enrolled on a cooperative group study. Data on MDS from treatment-based studies and cancer registries were inaccurate and seemed to significantly underestimate the incidence of MDS in children. PMID- 10520008 TI - Kaposi's sarcoma-associated herpesvirus (KSHV/HHV-8) DNA sequences are absent in leukapheresis products and ex vivo expanded CD34+ cells from multiple myeloma patients. AB - Recently it was reported that Kaposi's sarcoma-associated herpesvirus (KSHV/HHV 8) infects bone marrow (BM) dendritic cells (DC) in multiple myeloma (MM) patients and therefore might play a role in MM development. Because of the use of myeloid growth factors like GM-CSF and G-CSF for the mobilization of peripheral blood progenitor cells (PBPC), the subsequent increase of DC precursors might imply a risk for KSHV contamination in PBPC grafts. Therefore, in this study leukapheresis products and ex vivo cultured CD34+ cell suspensions were analysed. KSHV DNA could not be amplified in any of them. PMID- 10520009 TI - Acute lymphoblastic leukaemia occurring as second malignancy: report of the GIMEMA archive of adult acute leukaemia. Gruppo Italiano Malattie Ematologiche Maligne dell'Adulto. AB - Between July 1992 and June 1996, 901 new cases of adult acute lymphoblastic leukaemia were recorded in the GIMEMA Archive of Adult Acute Leukaemia; 21 of them (2.3%) had a previous primary malignancy (PM). We found that secondary acute lymphoblastic leukaemia cases (sALL) presented with older age, a high incidence of pre-pre-B immunophenotype and a significantly higher prevalence of cancer among relatives compared to de novo ALL. The leukaemogenic activity of the cytotoxic drugs employed for the treatment of PM may have played a potential role in only a proportion of patients, opening the possibility that some sALL patients may have developed two or more malignancies due to individual predisposing factors. PMID- 10520010 TI - A novel BCR-ABL transcript e2a2 in a chronic myelogenous leukaemia patient with a duplicated Ph-chromosome and monosomy 7. AB - A novel BCR-ABL transcript was detected by multiplex RT-PCR in a patient with Philadelphia chromosome (Ph) positive chronic myelogenous leukaemia (CML) in accelerated phase. Sequencing of the aberrant transcript revealed an in-frame e2a2 fusion that included a 9 basepairs insertion. Cytogenetic analysis showed t(9;22), an additional Ph chromosome and monosomy 7. The clinical course was dismal: therapy was poorly tolerated, and the patient died in blast crisis 10 months after diagnosis. These data support the association of additional Ph and monosomy 7 with poor prognosis and suggest that the novel e2a2 BCR-ABL transcript may be related to an aggressive clinical course. PMID- 10520011 TI - Revision of the diagnosis of T-zone lymphoma in the father of a patient with autoimmune lymphoproliferative syndrome type II. AB - Autoimmune lymphoproliferative syndrome (ALPS) is a disease of childhood characterized by typical clinical and laboratory findings. Here we describe an adult patient presenting with lymph node enlargement and splenomegaly. Pathological examination of an adenopathy supported the diagnosis of malignant T zone lymphoma. The patient was treated accordingly. 3 years later his child was diagnosed with ALPS. Therefore the diagnosis of the father's disease was reconsidered. Review of the slides and functional tests led to the diagnosis of ALPS in both father and son. ALPS should be considered as a possible differential diagnosis in adult patients presenting with rare types of T-cell lymphomas. PMID- 10520012 TI - The effect of p53 dysfunction on purine analogue cytotoxicity in chronic lymphocytic leukaemia. AB - To clarify the role of p53 in the killing of chronic lymphocytic leukaemia (CLL) cells by purine analogues, we examined the cytotoxic effects of chlorodeoxyadenosine and fludarabine on CLL cells that had been characterized according to their p53 functional status. Cases of CLL with p53 dysfunction (n = 7) displayed slight, but significant, resistance to nucleoside-induced cell killing when compared with cases with functionally intact p53 (n = 12). The small difference between the two groups indicated that p53 plays a minor role in such killing. These findings suggest that the poor therapeutic response to purine analogues observed in patients with p53 defects is likely to be caused by the emergence, on a background of genomic instability, of CLL-cell clones that are resistant to nucleoside-induced killing for reasons unrelated to p53. PMID- 10520013 TI - Improved outcome in haemophagocytic lymphohistiocytosis after bone marrow transplantation from related and unrelated donors: a single-centre experience of 12 patients. AB - Haemophagocytic lymphohistiocytosis (HLH) is an autosomal recessive disease with histiocytic and lymphocytic infiltrations in multiple organs. Cure seems possible only by allogeneic bone marrow transplantation (BMT), but matched sibling donors (MSD) are restricted and high mortality rates are associated with BMT from unrelated donors (URD). We report on 12 consecutive HLH patients with an improved outcome following URD transplants. Eight patients received BMT from URD, four from MSD. Five patients had signs of active HLH at the time of BMT. The conditioning regimen consisted of 20 mg/kg busulphan, 60 mg/kg VP-16 and 120 mg/kg cyclophosphamide and, in case of URD, 90 mg/kg antithymocyte globulin. The doses of busulphan and VP-16 were reduced during the programme to 16 mg/kg and 30 mg/kg, respectively. Using a fivefold graft-versus-host disease (GVHD) prophylaxis, GVHD was absent or mild in 10, and moderate or severe in two patients undergoing unrelated transplants. One patient with URD experienced graft failure and was retransplanted on day 37. Major toxicities were hepatic veno occlusive disease in five, capillary leak syndrome in two, pneumonia in three, sepsis in one, severe mucositis in one and seizures in two patients. All patients are alive without HLH after a median follow-up of 24.5 months. One patient has chronic GVHD, another patient has severe retardation. Three patients show slight to moderate development delay. These results indicate that in HLH, BMT from matched unrelated donors should be performed. Incomplete resolution of disease activity need not impede a successful outcome. PMID- 10520014 TI - Absolute CD4+ T-lymphocyte and CD34+ stem cell counts by single-platform flow cytometry: the way forward. AB - To determine the potential advantage of single-platform technology in the enumeration of CD4+ T lymphocyte and CD34+ stem cells, data has been analysed from the UK NEQAS for Leucocyte Immunophenotyping schemes. The inter-laboratory CVs for CD4+ T lymphocyte counts were consistently lower for single-platform (mean 13.7%, range 10-18.3%) compared to dual-platform methodology (mean 23.4%, range 14.5-43.7%). Subgroup analysis of single-platform users demonstrated mean overall inter-laboratory CVs of 17.2%, 13% and 7.1% for the FlowCount, TruCount and volumetric approach respectively. The lowest inter-laboratory CVs obtained for a single sample by each single platform approach were 4% (TruCount), 4.4% (volumetric), 4.6% (FACSCount) and 12.7% (FlowCount). Similarly, the mean inter laboratory CV for CD34+ stem cell enumeration using non-standardized single platform approaches was 18.6% (range 3.1-36.9%) compared to 28.6% (range 19 44.2%) for the dual-platform technology. Our results suggest absolute cell subset enumeration should be performed by single-platform technology and that such an approach should improve the quality control of multi-centre clinical trial data for CD4+ T lymphocyte and CD34+ stem cells. PMID- 10520015 TI - Resolution of immune haemolytic anaemia with allogeneic bone marrow transplantation after an unsuccessful autograft. AB - Autologous transplantation of lymphocyte-depleted peripheral blood stem cells (PBSC) has been proposed for treatment of patients with severe autoimmune disease. However, several patients have been reported to achieve only transient remissions. We report on a child with thalassaemia intermedia and immune-mediated haemolytic anaemia, given an autologous lymphocyte-depleted PBSC transplant, who relapsed 7 weeks after transplant. A complete remission, lasting 18 months to date, was obtained with allogeneic bone marrow transplantation (BMT) from an HLA matched unrelated donor. This experience indicates that, in selected cases, allogeneic BMT may be the treatment of choice for life-threatening autoimmune disease. A graft-versus-autoimmunity effect may favour the eradication of the recipient autoaggressive lymphocytes. PMID- 10520016 TI - Laparoscopic splenectomy: single-centre experience of a district general hospital. AB - We report the results of 20 consecutive laparoscopic splenectomies performed on haematology patients for a number of indications. Our series includes patients up to 77 years of age at the time of surgery and removal of spleens weighing up to 3530 g. The most significant benefit is the early rate of discharge post operatively (median 2 d); however, there is a risk of conversion to open laparotomy (in this series 3/20, 15%). We show that laparoscopic splenectomy can be offered as a therapeutic option to patients unfit for conventional laparotomy and that even large and bulky spleens can be removed safely using this approach. PMID- 10520017 TI - Increased macrophages, high serum M-CSF and low serum cholesterol in myelodysplasia and Kawasaki disease. PMID- 10520018 TI - Translocation (11;14)(q13;q32) in CD5-positive B-cell lymphoma associated with haemophagocytic syndrome. PMID- 10520019 TI - Images in haematology. Hepatic haemopoiesis in beta-thalassaemia trait. PMID- 10520020 TI - Management of sickle cell disease: recent advances and controversies. PMID- 10520021 TI - Is it dominantly inherited beta thalassaemia or just a beta-chain variant that is highly unstable? PMID- 10520022 TI - Coagulation history, Oxford 1951-53. PMID- 10520023 TI - Haemopoietic progenitor cell lines generated by the myeloproliferative leukaemia virus: a model system to analyse murine and human lineage-affiliated genes. AB - Multipotential progenitor and stem cells occur with a low frequency in haemopoietic tissue. As a result, it is often difficult to obtain sufficient numbers of cells to undertake many of the assays that would be informative about the molecular events involved in the regulation of lineage-affiliated genes within these multipotent cells. To circumvent this problem, we have used the myeloproliferative leukaemia virus (MPLV) to generate a phenotypically diverse array of haemopoietic progenitors from adult mouse bone marrow and embryonic blood. These cells could be expanded to perform a variety of analyses that would not previously have been possible using analogous primary cells. The validity of these assays was supported by the observation that the phenotype of several MPLV infected lines was very similar to previously described primary haemopoietic progenitor cells. By using mice transgenic for the human alpha and beta globin gene clusters, we have shown that human genes may also be investigated. In addition, this strategy has a wide potential applicability including the rescue of haemopoietic progenitors from mouse embryos lacking genes critical for their survival as well as the study of any haemopoietic gene for which an appropriate transgenic mouse is available. PMID- 10520024 TI - Pregnancy in bone marrow failure syndromes: Diamond-Blackfan anaemia and Shwachman-Diamond syndrome. AB - Pregnancy in bone marrow failure syndromes has risk to mother and fetus. There are fewer than 30 reports of cases with Diamond-Blackfan anaemia (DBA), and none with Shwachman-Diamond syndrome (SD). We report two DBA and one SD cases. One DBA mother received transfusions intra-partum, and the other only post-partum. Both required caesarean sections (C-sections) for failure of labour to progress and severe pre-eclampsia respectively. Both subsequently resumed pre-pregnancy steroid-induced control of anaemia. approximately 40% of DBA pregnancies required maternal transfusions; 25% delivered by C-section. The SD patient also had Ehlers Danlos (ED) syndrome and urticaria pigmentosa (UP). Her blood counts were adequate until week 38, when the platelet count dropped and a C-section was performed. Pregnancy management in marrow failure disorders requires obstetricians with expertise in high-risk pregnancies, and haematologists with experience with marrow failure syndromes. PMID- 10520025 TI - Long-term decrease of CD4+CD45RA+ T cells and impaired primary immune response after post-traumatic splenectomy. AB - Congenital or acquired absence of the spleen and functional hyposplenism are associated with abnormalities of host defence such as an increased susceptibility to infection with encapsulated bacteria. The effects of the lack of the spleen on cell-mediated immunity are largely unknown. In the present study we have investigated peripheral blood lymphocyte subpopulations in healthy adults who had undergone splenectomy because of severe abdominal trauma > 4 years before the study. The results show a significant reduction in the percentage of CD4+ T cells due to a selective and long-term decrease in the percentage of CD4+CD45RA+ lymphocytes, the CD4+ T-cell subset mainly involved in primary immune responses to newly encountered antigens. Levels of the reciprocal CD45RO+CD4+ T-cell subset were comparable between splenectomized and control individuals, as were lymphoproliferative responses and IFN-gamma production to recall antigens. Decreased levels of CD4+CD45RA+ cells were accompanied by an impairment in primary immune responsiveness, as assessed by investigating T-cell proliferation to stimulation with keyhole limpet haemocyanin and by measuring antibody responses following primary immunization with a clinically relevant T-dependent antigen, hepatitis A vaccine, in vivo. These findings suggest a possible role of the spleen in the generation, maintenance and/or differentiation of naive, unprimed T cells or their precursors, which might have a possible functional relevance for primary immune responses following splenectomy. PMID- 10520026 TI - A simple, robust, validated and highly predictive index for the determination of risk-directed therapy in acute myeloid leukaemia derived from the MRC AML 10 trial. United Kingdom Medical Research Council's Adult and Childhood Leukaemia Working Parties. AB - Data on 1711 patients, aged up to 55 years, in the MRC AML 10 trial were used to create a prognostic index for use in risk-directed therapy decision making for younger patients with acute myeloid leukaemia (AML). Two parameters, response after course 1 and cytogenetics, were strongly predictive of outcome. For patients with complete remission, partial remission and resistant disease, 5-year survival from the start of course 2 was 53%, 44% and 22% and relapse rates were 46%, 48% and 69% respectively, and for patients with favourable, intermediate and adverse karyotypic abnormalities, survival was 72%, 43% and 17% and relapse rates were 34%, 51% and 75% respectively (all P < 0.0001). Patients with FAB type M3 but no cytogenetic t(15;17) also had a low relapse rate (29%). These three factors were combined to give three risk groups: good (favourable karyotype or M3, irrespective of response status or presence of additional abnormalities), standard (neither good nor poor), poor (adverse karyotype or resistant disease, and no good-risk features). Survival for these three groups was 70%, 48% and 15% respectively and relapse rates were 33%. 50% and 78% (both P < 0.0001). The index is simple (based on just three parameters), robust (derived from 1711 patients), highly discriminatory (55% survival difference between good and poor risk) and validated, so can be applied in the clinical setting to assist with therapeutic decisions as in the current AML 12 trial. PMID- 10520027 TI - Real-time RT-PCR for the detection and quantification of AML1/MTG8 fusion transcripts in t(8;21)-positive AML patients. AB - AML1/MTG8 was quantified relative to the expression of the GAPDH housekeeping gene by real-time RT-PCR in 22 patients with t(8;21)-positive acute myeloblastic leukaemia (AML) at initial diagnosis and in seven of these patients also during/after chemotherapy and allogeneic bone marrow transplantation. Real-time PCR was able to specifically detect and quantify AML1/MTG8 over a 5 log range. The detection limit for t(8;21)-positive cells was a dilution of 1:105. The AML1/MTG8 expression varied considerably among the 22 AML patients at intial diagnosis with a ratio AML1/MTG8:GAPDH of 0.5135+/-0.536 (range 0.1-2.14, median 0.318). In six patients with t(8;21)-positive AML a marked decline of AML1/MTG8 could be induced by chemotherapy. These patients are in ongoing complete haematological remission (CR) with a constant low-level AML1/MTG8 expression. In another patient a rapid rise of AML1/MTG8 transcripts could be detected in CR after allogeneic bone marrow transplantation and the patient relapsed 10 weeks later. In conclusion, real-time RT-PCR is a suitable approach for the quantification of AML1/MTG8 transcripts in the monitoring of AML patients with t(8;21) during/after chemotherapy and can provide data of prognostic relevance. PMID- 10520028 TI - All-trans retinoic acid regulates adhesion mechanism and transmigration of the acute promyelocytic leukaemia cell line NB-4 under physiologic flow. AB - The success of all-trans retinoic acid (ATRA) in the therapy of acute promyelocytic leukaemia (APL) has received increased attention. Unfortunately, life-threatening multiorgan failure commonly occurs, i.e. retinoic acid syndrome, and is thought to be the result of organ infiltration by leukaemic cells. We hypothesized that ATRA-induced differentiation of APL cells leads to adhesion receptor alterations responsible for leucocyte extravasation from the blood into tissue. Changes in adhesive properties of the APL cell line NB-4 in response to ATRA were investigated using a parallel plate flow chamber under conditions that recapitulate physiologic flow conditions. Untreated NB-4 cells initially tether and roll on activated human umbilical vein endothelial cell monolayers using a combination of E-selectin, P-selectin and alpha4 integrin. After ATRA treatment, > 80% of initial NB-4 cell attachment to endothelial cells was E-selectin dependent. Stable arrest (firm adherence) of NB-4 cells on activated endothelium was also altered by ATRA treatment. Untreated NB-4 cells used alpha4 integrin to arrest on endothelium, but beta2 integrin dependent arrest was induced by ATRA. With the acquisition of beta2 integrin function, ATRA-treated cells acquired the ability to transmigrate through activated endothelium. Thus, ATRA dramatically altered the adhesion phenotype on NB-4 cells: ATRA induced rolling largely attributable to E-selectin, abrogated alpha4 integrin dependent rolling, and promoted acquisition of beta2 integrin dependent firm adherence and transmigration. These findings represent novel cellular and differentiation effects of ATRA, and, to our knowledge, are the first demonstration that a therapeutic agent differentially regulates alpha4 and beta2 integrin on the same leucocyte. PMID- 10520029 TI - fas-mediated lysis of chronic lymphocytic leukaemia cells: role of type I versus type II cytokines and autologous fasL-expressing T cells. AB - Given the known role of the fas cytolytic pathway in B-cell regulation, we evaluated whether fas-fasL interactions might induce chronic lymphocytic leukaemia (CLL) cell death. De novo CLL cells expressed a low level of surface fas, and were not lysed by fasL-bearing cells. CLL cells cultured in media containing the type I cytokines interleukin (IL)-12 or interferon (IFN)-alpha had increased fas expression, and were readily lysed by fasL-bearing cells. In contrast, the type II cytokine IL-4 did not increase CLL cell fas, and abrogated type I cytokine-induced fas up-regulation. With prolonged culture, IL-4 exposed CLL cells expressed an intermediate level of fas; however, such CLL cells were resistant to fas-mediated lysis. These results indicate that IL-4 inhibits fas mediated killing of CLL cells at the level of both fas receptor expression and post-receptor events. Additionally, we have defined in vitro culture conditions which generate fasL-bearing T cells from CLL patients; such T cells efficiently mediated fas-based lysis of autologous fas-positive CLL cells. We therefore conclude that type I and type II cytokines differentially regulate the fas pathway in CLL cells, and that a combination of type I cytokines and fasL expressing T cells may represent a new approach to the immunotherapy of CLL. PMID- 10520030 TI - Molecular features of primary mediastinal B-cell lymphoma: involvement of p16INK4A, p53 and c-myc. AB - Primary mediastinal B-cell lymphoma (PMBL) shows chromosome 9p anomalies in 50% of cases. Based on reports that p16INK4A gene, located on this chromosomal arm, is frequently altered in aggressive lymphomas, we analysed for alterations of this gene in 27 cases of PMBL, which were part of a series of 32 PMBL cases that have been characterized for alterations in c-myc, p53, N-ras, bcl-1, bcl-2, bcl-6 and for Epstein-Barr virus (EBV) infection. Four cases showed p16INK4A gene anomalies, including three with promoter methylation and one homozygous deletion. Eight PMBLs showed c-myc rearrangements. Three additional cases showed sequence variations in the c-myc P2 promoter, two of which consisted of the same germline variation involving a novel polymorphic XhoI site. Four tumours contained p53 gene mutations and three had clonal EBV infection. One case had a bcl-6 rearrangement. In conclusion, our study shows that p16INK4, c-myc and p53 alterations occur in 15%, 25% and 13% of PMBLs, respectively. EBV monoclonality was found in 9% of cases, whereas no abnormality was detected in bcl-1, bcl-2 and N-ras. Thus, none of the common genetic aberrations seen in other types of non Hodgkin's lymphomas appears to be stringently involved in the pathogenesis of this unique lymphoma type. PMID- 10520031 TI - VH gene sequences from a novel tropical splenic lymphoma reveal a naive B cell as the cell of origin. AB - The prevalence of malaria and other infections in tropical Africa provides a setting for the emergence of B-cell tumours distinct from that in Western countries. Attempts to draw comparisons with Western lymphomas have led to difficulties, with so-called African chronic lymphocytic leukaemia (CLL) having a different pattern of incidence from Western CLL. Splenomegaly is common in African CLL, and this has posed diagnostic problems in differentiating the tumour from malaria-associated hyper-reactive malarial splenomegaly (HMS). One feature of the splenomegalic form of African CLL is that the tumour cells often possess short but fine cytoplasmic projections reminiscent of those observed in Western splenic lymphoma with villous lymphocytes (SLVL). Analysis of Ig VH genes both facilitates discrimination between clonal B-cell tumours and HMS, and reveals the differentiation status of the cell of origin. This study indicated that VH genes of nine cases of clonal splenic B-cell tumours with villous lymphocytes from Ghana were relatively unmutated, consistent with an origin from a naive B cell. These features differ from SLVL which arises from a post-follicular antigen selected B cell. One possibility is that these splenic B-cell tumours derive from a splenic T-independent B cell, with malaria infection as a potential influence. PMID- 10520032 TI - High-sensitive immunophenotyping and DNA ploidy studies for the investigation of minimal residual disease in multiple myeloma. AB - Sensitive techniques for monitoring minimal residual disease (MRD) in multiple myeloma (MM) are needed to evaluate the effectiveness of new intensive treatment strategies. The aim of the present study was to explore the applicability and sensitivity of flow cytometry immunophenotyping and DNA ploidy studies for the investigation of residual myelomatous plasma cells (PC) in MM patients. Bone marrow (BM) samples from 61 untreated MM patients were immunophenotypically analysed with a panel of 21 monoclonal antibodies, using a high-sensitive method based on a two-step acquisition procedure through a SSC/CD38 -CD138+ 'live-gate'. Overall, in 87% of MM cases, PC displayed an aberrant phenotype at diagnosis. The most important aberrant criteria were: antigen over-expression of CD56 (62%), CD28 (16%) and CD33 (6%) and asynchronous expression of CD117 (28%), sIg (21%) and CD20 (10%). DNA aneuploidy was found in 62% of cases. The simultaneous use of these two techniques allowed the detection of aberrant/aneuploid PC in 95% of the cases. Based on dilutional experiments, the detection limit of both techniques ranged from 10(-4) to 10(-5). In 29 stem cells harvests and 19 BM samples obtained 3 months after autologous transplantation, we have investigated the presence of residual myelomatous PC; they were detected in 44% of the stem cell collections and in 61% of the BM samples obtained after transplant. The percentage of pathological PC did not significantly change during the days of harvest. In summary, the present study shows that the combined use of immunophenotyping and DNA ploidy studies is a suitable approach for MRD investigation in MM patients based on their applicability (95% of cases) and sensitivity (up to 10(-5)). PMID- 10520033 TI - Serum interleukin-6 has no discriminatory role in paraproteinaemia nor a prognostic role in multiple myeloma. AB - We determined interleukin-6 (IL-6) levels in the serum of 212 well-defined patients with newly diagnosed paraproteinaemia and evaluated its discriminatory value and prognostic role in multiple myeloma (MM). Results were compared with serum neural cell adhesion molecule and beta-2-microglobulin, both established prognostic MM markers. Paraproteinaemia-related diagnoses were: MM (60), other haematological diseases (46), solid tumours (35), autoimmune diseases (17) and monoclonal gammopathy of unknown significance (MGUS) (54). The range of IL-6 levels in all diagnostic groups overlapped widely and did not serve as a discriminatory marker in newly diagnosed paraproteinaemia even when patients with infection or fever (42) were excluded. In MM high IL-6 levels (>/= 50 pg/ml) were not associated with a shorter survival (P = 0.24). We compared our results with 20 published studies on serum IL-6 in paraproteinaemia and/or MM. IL-6 data have to be related to the assay used (bio- or immunoassay) and to the status of MM (newly diagnosed, during therapy, progressive disease). We conclude that serum IL 6 is not specific for paraproteinaemia-related diseases and will not serve as a reliable discriminatory or prognostic marker in paraproteinaemia and MM. PMID- 10520034 TI - Platelet c-mpl expression is dysregulated in patients with essential thrombocythaemia but this is not of diagnostic value. AB - Essential thrombocythaemia (ET) can be difficult to discriminate from an occult case of reactive thrombocytosis (RT). Since thrombopoietin (TPO) is the primary regulator of thrombopoiesis, we have investigated whether levels of TPO and/or its receptor, c-mpl, are of value in the differential diagnosis of ET. Plasma TPO levels in patients with ET, RT and other myeloproliferative disorders (MPDs) did not differ significantly from normal controls. However, surface c-mpl expression was significantly reduced in platelets from 18 ET patients, 0-65.5% of controls (P < 0.001). Immunoblots on five of these and five additional patients were consistent with absent or reduced c-mpl protein levels. The surface c-mpl expression results were significantly different from those in eight RT patients (21. 3-95.5%, P = 0.0015), but there was considerable overlap between the two groups and a reduced level was not restricted to ET. Furthermore, c-mpl expression in ET patients was not different from eight patients with other MPDs (0-87.6%, P = 0.06), nor could it differentiate between ET patients with monoclonal and polyclonal haemopoiesis. Although a low or absent c-mpl level is suggestive of a primary rather than a secondary thrombocytosis, it is insufficiently discriminatory to be used as a diagnostic marker for ET. PMID- 10520035 TI - The PNH phenotype cells that emerge in most patients after CAMPATH-1H therapy are present prior to treatment. AB - Paroxysmal nocturnal haemoglobinuria (PNH) cells are deficient in glycosylphosphatidylinositol (GPI) linked antigens due to a somatic mutation of the PIG-A gene in a haemopoietic stem cell. It appears that a PNH clone reaches detectable proportions only when there is selection in its favour. GPI-deficient T lymphocytes have been identified in patients treated with CAMPATH-1H, a monoclonal antibody against the GPI-linked CD52 molecule. CAMPATH-1H selects for cells that are deficient in CD52 (such as PNH-like cells) promoting the development of a PNH-like clone (analogous to PNH). We report that 10/15 patients with chronic lymphocytic leukaemia developed PNH-like lymphocytes after therapy with CAMPATH-1H. The remaining five patients developed no PNH-like cells at any stage, including one patient who received 12 weeks of therapy. The inactivating PIG-A mutation has been identified in one patient. This mutation was detectable by an extremely sensitive mutation-specific PCR-based analysis in the patient's mononuclear cells prior to CAMPATH-1H therapy. The frequency and phenotype of GPI deficient lymphocytes after CAMPATH-1H and the detection of a PIG-A mutation in the lymphocytes prior to CAMPATH-1H therapy indicated that such mutations were present in a very small proportion of cells prior to selection in their favour by CAMPATH-1H. This suggests that a large proportion of individuals have cells with PIG-A mutations that are not detectable by flow cytometry and thus may have the potential to develop PNH. PMID- 10520036 TI - Allogeneic bone marrow transplantation in aggressive non-Hodgkin's lymphoma (excluding Burkitt and lymphoblastic lymphoma): a series of 73 patients from the SFGM database. Societ Francaise de Greffe de Moelle. AB - The place of allogeneic bone marrow transplantation (BMT) in the treatment of aggressive non-Hodgkin's lymphoma (NHL) remains controversial. We conducted a retrospective study of French experience in allografting NHL between 1984 and 1994. To improve the homogeneity of the study population, cases of low-grade, Burkitt and lymphoblastic NHL were excluded. 73 patients were included in the analysis. Median age at transplantation was 35 years (range 9-61 years); 64 patients were in stage IV and 45 had bone marrow involvement at diagnosis. At the time of transplantation, 46 patients had sensitive disease (25 in complete remission; CR). The overall survival (OS) and progression-free survival (PFS) rates were 41% and 40% respectively at 5 years (median follow-up of survivors 90 months). The probability of disease progression was 30% at 5 years, and only one relapse occurred after 15 months. 32 patients died of transplantation-related complications. In multivariate analysis, pretransplant complete remission was the main factor associated with longer survival (OS at 60 months of 76% among the 25 patients in CR at transplant and of 23% among the 48 patients not in CR at transplant). Neither acute nor chronic graft-versus-host disease (GvHD) influenced the relapse rate. In conclusion, in this high-risk population the overall results of allogeneic BMT were encouraging, despite a high transplant related mortality rate. We believe this procedure should be studied further in prospective controlled trials. PMID- 10520037 TI - Competitive repopulation of retrovirally transduced haemopoietic stem cells. AB - Gene transfer into haemopoietic stem cells (HSC) may be useful in gene therapy for a variety of inherited and acquired human diseases. Cell division is required for retroviral transduction, and cytokine stimulation is often used to increase mitosis of quiescent HSC. Exposure to cytokines has been shown to have an unfavourable effect on the engraftment of these cells when competed with unmanipulated HSC. We now show that a similar engraftment defect is present when HSC are manipulated and transduced with the human multiple drug resistance (MDR) gene. The extent of the unfavourable competition depended on the relative numbers of cytokine-treated and fresh cells when the two populations of cells were administered simultaneously into marrow-ablated isogenic mice. When the manipulated transduced cells were given 2 or 4 d before the unmanipulated cells there was a much greater engraftment of the manipulated cells. The data suggested that the manipulated cells were at a relative disadvantage for marrow engraftment as compared to fresh cells, presumably due to the more efficient homing and engraftment properties of these latter unmanipulated cells. However, the manipulated cells had no intrinsic inability to engraft when they were the predominant donor cell population. In all cases the percent of MDR transduced cells in the engrafting manipulated cells remained relatively constant at about 25-30%. These results have implications for the use of manipulated transduced stem cells in gene therapy, suggesting that administering them before adding fresh cells can overcome their engraftment defect. PMID- 10520038 TI - Selective depletion of major and minor histocompatibility antigen reactive T cells: towards prevention of acute graft-versus-host disease. AB - Development of acute graft-versus-host disease (aGVHD) following HLA-identical sibling bone marrow transplantation (BMT) remains a serious complication. A selective depletion of T cells has proved to be effective in preventing aGVHD but is associated with relapse and increased incidence of infection. As aGVHD is directed mainly against epithelial tissues we examined whether it would be feasible to selectively deplete T cells reactive with epithelial cells whilst preserving other specificities. Donor T cells which express HLA-DR, CD25, CD69 and CD71 activation markers after cocultivation with patient keratinocytes were depleted using magnetic cell separation techniques. Depletion of major as well as minor histocompatibility antigen activated T cells revealed a significant (P = 0.004 and P = 0.031, respectively) 10-fold decrease in the frequency of donor T lymphocyte precursors reactive with patient keratinocytes. The frequency reactive with third-party and patient peripheral blood mononuclear cells, including leukaemia cells, remained unchanged, supporting the notion that aGVHD and graft versus-leukaemia (GVL) may be separable. This alloantigen-specific depletion may be used in matched unrelated as well as HLA-identical sibling BMT for reducing aGVHD whilst conserving GVL. PMID- 10520039 TI - Distinct patterns of apoptosis in association with modulation of CD44 induced by thrombopoietin and granulocyte-colony stimulating factor during ex vivo expansion of human cord blood CD34+ cells. AB - The insufficient number of haemopoietic stem cells (HSCs) in cord blood (CB) is the major potential limitation to widespread use of CB for marrow replacement. Cytokine-mediated ex vivo expansion has been proposed as a means of increasing the number of CB HSCs for transplantation. However, the biology of CB HSCs during cytokine-mediated ex vivo expansion, such as apoptosis or expression of adhesion molecules, has not yet been elucidated. We have investigated the patterns of apoptosis and CD44 expression on human CB CD34+ cells during ex vivo expansion. CD34+ cells isolated from human CB were cultured in a stroma-free liquid culture system with thrombopoietin (TPO), flt3-ligand (FL), stem cell factor (SCF), and/or granulocyte-colony stimulating factor (G-CSF). During the culture, for up to 5 weeks, apoptosis was measured by staining with 7-amino-actinomycin D (7-AAD) along with concurrent immunophenotyping of CD34 and CD44 with three-colour flow cytometry. In the cultures with TPO, an apoptotic fraction with down-regulated CD44 appeared from the fourth day up to the second week. G-CSF also induced apoptosis but in a different manner; the apoptotic fraction without down regulation of CD44 appeared unremittingly for up to 5 weeks. FL did not induce apoptosis or down-regulation of CD44. These findings show that apoptosis is indeed involved in the regulation of CB CD34+ cells in ex vivo expansion and the patterns of apoptosis are dependent on the type of cytokines used. The distinct patterns of apoptosis suggest different mechanisms of TPO and G-CSF in inducing apoptosis, which still remains to be elucidated. PMID- 10520040 TI - Evidence that anti-HBc but not HBV DNA testing may prevent some HBV transmission by transfusion. AB - Blood donor screening for antibody to hepatitis B core antigen (anti-HBc) implemented in some countries as a surrogate marker for non-A, non-B hepatitis has been superseded by anti-HCV screening. To assess the value of anti-HBc screening for the detection of hepatitis B surface antigen-negative blood donations that might contain infectious HBV, HBV genomic detection and recipient testing were used. Blood donations were screened and confirmed by multiple anti HBc assays. Donations containing isolated anti-HBc and those with anti-hepatitis B surface antigen (anti-HBs) level < 0.1 IU/ml were tested for the presence of HBV DNA. Recipients of previous donations from the corresponding donors during the previous 5 years were traced and tested for markers of HBV infection. Of 103 869 donations screened, 586 (0.56%) were anti-HBc positive, two of which contained HBsAg, and 413 (0.4%) had protective (>/= 0.1 IU/ml) levels of anti HBs. Anti-HBs < 0.1 IU/ml was found in 102 of these donations (0.1%) and isolated anti-HBc in 69 (0.07%). No donations with isolated anti-HBc were HBV DNA confirmed positive. Of 278 recipients of previous donations from 171 donors at risk of HBV carriage, 12 had markers of HBV infection. Six recipients had other identified risk factors. An association with blood transfusion was considered probable in two and possible in four recipients. None of the six corresponding donors had detectable HBV DNA 6-40 months after the implicated donation. The frequency of HBV transmission by chronic carriers negative for hepatitis B surface antigen was estimated in this study to be 1 in 52,000 donations (CI 0.3 7.8/100,000) from HBsAg-negative donors. Such HBV infectious donations may not be detected by DNA amplification. PMID- 10520041 TI - Identification of a factor VIII peptide, residues 2315-2330, which neutralizes human factor VIII C2 inhibitor alloantibodies: requirement of Cys2326 and Glu2327 for maximum effect. AB - Factor VIII (FVIII) inhibitor alloantibodies react with combinations of the A2, C2 and A3-C1 domains of the FVIII molecule. Some inhibitors block binding of FVIII to both von Willebrand factor (VWF) and phospholipid, and recognize a C2 domain epitope which overlaps both binding sites. In order to determine the essential binding regions for alloantibodies inhibitory for FVIII activity, we have performed inhibitor neutralization assays and competitive inhibition assays using 10 overlapping synthetic peptides spanning the carboxy-terminal region of the C2 domain (residues 2288-2332). We found one peptide (2315-2330, L9) which neutralized the anti-FVIII activity of four out of five different C2 alloantibodies by 50%, 39%, 47% and 57%, respectively. Neutralization of these alloantibodies by recombinant C2 domain (residues 2173-2332) was 68%, 50%, 59%, 86% and >95%, respectively. The inhibitor which was not neutralized by L9 peptide and reacted by immunoblotting with peptide 2218-2307, did not prevent binding of FVIII to VWF and only partially inhibited binding of FVIII to phosphatidylserine. Mutants of the L9 peptide were prepared in which each residue from 2315-2330 was sequentially substituted by glycine. Inhibitor neutralization experiments using these peptides demonstrated that Arg2320 and Cys2326 or Glu2327 are important for the effect of L9 peptide, since their substitution by glycine reduced its neutralizing effect by 60% to >90%, suggesting that they are crucial for formation of the one of the C2 inhibitor epitopes. PMID- 10520042 TI - Bleeding and thrombosis in 55 patients with inherited afibrinogenaemia. AB - Knowledge of the spectrum of symptoms in patients with inherited afibrinogenaemia is limited by the rarity of this coagulation defect. We compared a large series of 55 afibrinogenaemic patients from Iran with 100 patients with severe factor VIII deficiency. In afibrinogenaemia there was a higher frequency of mucosal-type bleeding symptoms but joint and muscle bleeding was less frequent and severe than in haemophilia. Umbilical cord bleeding was relatively frequent only in afibrinogenaemic patients. Two young patients developed spontaneous thrombotic episodes and three women had recurrent abortions. Overall, in afibrinogenaemia bleeding symptoms are qualitatively different and less severe than in haemophilia. Afibrinogenaemia can also be accompanied by thrombotic manifestations. PMID- 10520043 TI - Racial background is a determinant of average warfarin dose required to maintain the INR between 2.0 and 3.0. AB - Warfarin is a commonly used prophylactic agent for the prevention of thromboembolic disease. We hypothesized that racial background influenced warfarin dosage, and tested this by recording the international normalized ratio (INR) in 867 patients aged 40-90 routinely passing through our Anticoagulation Service whose target INR was 2-3. Mean (95% confidence interval) dose was 4.1 (4.0-4.2) mg/d in 737 Caucasians, 5.5 (4.9-6.1) mg/d in 72 Asians, and 6.7 (5. 8 7.6) mg/d in 58 Afro-Caribbeans (P < 0.05) between each group). In a subgroup of 302 (41 Asians, 22 Afro-Caribbeans, 239 Caucasians), body mass index did not influence warfarin use. Despite small numbers, we conclude that racial background, but not body mass index, is a determinant of warfarin dosage. The reasons for this could be genetic, cultural (diet related), or both. PMID- 10520044 TI - Factor V Leiden and the common haemochromatosis mutation HFE C282Y: is there an association in familial venous thromboembolic disease? AB - The involvement in venous thrombosis of the two most common mutations of the hereditary haemochromatosis gene (HFE C282Y and HFE H63D) was investigated in 239 patients with objectively proven venous thrombosis. Neither mutation showed an increased prevalence in the cohort (HFE C282Y: 13.0% (95% CI 9.3-17.8) patients, 16.2% (95% CI 14.3-18.2) controls; HFE H63D: 28.3% (95% CI 22.9-34.3) patients, 28.1% (95% CI 25.8-30.6) controls. Neither mutation was increased in patients with factor V Leiden (FVL) compared to those without. However, HFE C282Y was increased among patients who had both FVL and a family history of thrombosis (7/20), compared with those with FVL and no family history (1/22) (relative risk 7.97, 95% CI 1.5-43.1, P = 0.016). PMID- 10520045 TI - Born to clot: the European burden. PMID- 10520046 TI - Unusual presentation of primary autoimmune neutropenia. PMID- 10520047 TI - Airway vascularity in asthma. PMID- 10520048 TI - Interleukin-12 and allergic tissue response. PMID- 10520049 TI - Occupational asthma and interleukin-8. PMID- 10520050 TI - Environmental and lifestyle factors may act in concert to increase the prevalence of respiratory allergy including asthma. PMID- 10520051 TI - Neuro-immune interactions in the lung. PMID- 10520052 TI - Expression of interleukin (IL)-12 (p40) and IL-12 (beta 2) receptors in allergic rhinitis and chronic sinusitis. AB - BACKGROUND: Interleukin (IL)-12 is a relatively new and structurally distinct TH1 associated cytokine produced by B cells and macrophages, which may play a suppressive role in the development of allergic sinonasal mucosal responses. OBJECTIVE: We investigated the expression of IL-12 (inducible p40 subunit) and its receptor (IL-12R beta2 subunit) in tissue biopsies of naturally exposed patients with allergy-associated (ACS) and nonallergy-associated chronic sinusitis (NCS) and compared it with controls. We also examined IL-12 and IL-12R expression in biopsies from a ragweed allergen challenge model. In the allergen challenge model, the effect of pretreatment with topical corticosteroids on IL-12 and IL-12R expression was assessed. METHODS: To detect IL-12 and IL-12R mRNA, we employed the technique of in situ hybridization using digoxigenin-labelled riboprobes. RESULTS: In both ACS and NCS subjects there was decreased expression of IL-12 as compared with control (P < 0.05). IL-12R (beta2) expression was decreased in ACS subjects as compared with control (P < 0.05), however, there was no significant difference found between NCS subjects and control. In the allergen challenge subjects, there was a significant decrease in IL-12 expression following challenge (P < 0.05). This effect was abrogated by pretreatment of the subjects with topical corticosteroids. However, IL-12R (beta2) expression showed no change following allergen challenge while pretreatment with topical corticosteroids resulted in increased expression of the (beta2) receptor after allergen challenge (P < 0.05). CONCLUSION: Our data suggest that IL-12 plays a role in the in vivo suppression of the allergic inflammatory response and that the control of this suppression may be exerted largely via the IL-12 (beta2) receptor. PMID- 10520053 TI - Short-term preseasonal birch pollen allergoid immunotherapy influences symptoms, specific nasal provocation and cytokine levels in nasal secretions, but not peripheral T-cell responses, in patients with allergic rhinitis. AB - BACKGROUND: [corrected] Birch pollen allergic rhinitis can be sufficiently treated with specific subcutaneous allergoid immunotherapy (IT). However, little is known about the clinical and immunological effects of short-term therapy protocols. OBJECTIVE: To investigate the clinical efficacy of a birch pollen allergoid IT using seven preseasonal injections and to evaluate immunological parameters that might explain clinical findings. METHODS: Thirty-seven patients were included into the study and randomized to either a symptomatic treatment or allergoid IT plus symptomatic treatment. Patients were examined during the pre-IT season, at two extraseasonal visits both before and after IT and during the post IT season. At each visit, nasal secretion samples were taken and analysed for levels of IL-4, IL-5 and IFNgamma. In addition, short-term birch-specific T-cell lines (TCLs) were cultured from peripheral blood mononuclear cells of 10 patients of the IT group, both before and after IT, and the ratios of lymphocyte subpopulations were determined. Cytokine production by TCLs (IL-4, IL-5, IFNgamma, IL-10) and proliferation of TCLs in response to stimulation with birch pollen allergen were measured. RESULTS: It was possible to evaluate 27 patients in accordance with the study protocol. Clinical symptoms and medication intake were reduced as a result of the IT as were nasal secretion levels of IL-5 (P = 0.007). IFNgamma was increased in nasal secretions (P = 0.01), while IL-4 was not measurable in most samples. No effect was found on proliferation of birch pollen reactive TCLs, cytokine production by TCLs and the frequency and ratio of CD4+ and CD8bright or CD45RA+ and CD45RO+ cells in peripheral blood (all P > 0.05). Conclusion Preseasonal IT with a birch pollen allergoid is clinically effective in allergic rhinitis and influences cytokine production in the nose, but does not modulate the measured responses of peripheral blood T cells. PMID- 10520054 TI - Effects of house dust mite avoidance measures on Der p 1 concentrations and clinical condition of mild adult house dust mite-allergic asthmatic patients, using no inhaled steroids. AB - BACKGROUND: Exposure to house dust mite (HDM) allergens often results in worsening of asthma. Therefore, avoidance of exposure to HDM allergens is often proposed. Unfortunately, the most effective and feasible avoidance strategy is still not completely assessed. Consequently, we investigated the effects of a combined HDM avoidance strategy on HDM allergen concentrations and clinical condition of allergic, mild asthmatic, patients using no inhaled steroids. METHODS: Asthmatic patients, allergic to HDM, using no inhaled corticosteroids, were randomly allocated to an active (n = 76) or a placebo allergen-avoidance group (n = 81). Avoidance measures consisted of applying Acarosan(R) (placebo: water) to the living room and bedroom floors, and the use of HDM-impermeable covers for mattresses and bedding (placebo: cotton covers for mattresses only). Effects on allergen concentrations (Der p 1), FEV1, bronchial hyperresponsiveness, peak flow parameters and asthma symptom scores were studied during 20 weeks and controlled for the allergic status of the patients. RESULTS: The active covers reduced Der p 1 concentrations to 9.4% (P = 0.0001), and were always significant lower than in the placebo group (P = 0.0002). Acarosan(R) resulted in slight but significant decreases (twofold, P = 0.0001), both on living room and bedroom floors, but concentrations were never significantly lower than the placebo group. Although the combined avoidance strategy resulted in a considerable reduction in allergen load in the active group, no differences were seen between the two groups in any of the clinical parameters during the follow up period in this group of allergic asthmatics, using no inhaled corticosteroids. Corrections for the allergic status did not alter these results. CONCLUSIONS: The combined avoidance strategy was effective in reducing HDM allergen concentration. This was especially achieved by the allergen-impermeable covers, while the effects of Acarosan(R) were only marginal. However, this allergen reduction was not reflected in a convincing improvement in clinical condition in this group of mild allergic asthmatics, using no inhaled steroids. Perhaps, a longer follow-up period would have resulted in more pronounced effects. PMID- 10520055 TI - Effect of topical corticosteroids on seasonal increases in epithelial eosinophils and mast cells in allergic rhinitis: a comparison of nasal brush and biopsy methods. AB - BACKGROUND: Nasal brushing and nasal biopsy are well-tolerated sampling techniques. Seasonal grass pollen-induced rhinitis is characterized by epithelial mast cell infiltration and seasonal increases in both epithelial and sub-mucosal eosinophils. OBJECTIVE: To compare the ability of the nasal brush and nasal biopsy techniques to detect natural seasonal increases in eosinophils and mast cells, and to assess the influence of topical corticosteroid. METHODS: Nasal brush samples and nasal biopsies were collected from 46 grass pollen-sensitive seasonal rhinitis patients before the grass pollen season and at the peak of the pollen season following 6 weeks' treatment with either fluticasone propionate aqueous nasal spray (200 microg, twice daily) or placebo nasal spray. RESULTS: Placebo patients showed seasonal increases in epithelial eosinophils both with nasal brushing (P < 0.0001) and biopsy (P < 0.001). Epithelial mast cell numbers also increased during the pollen season as detectable by brushing (P < 0.0001) and biopsy (P < 0.03). Changes in cell numbers measured by nasal brushing correlated with those observed with nasal biopsy, both for eosinophils and mast cells (P < 0.05). Sub-mucosal eosinophils but not mast cells also increased during the pollen season (P < 0.002). Nasal brushing and biopsy revealed that fluticasone treatment inhibited seasonal increases in epithelial eosinophils (P < 0.00001) and epithelial infiltration by mast cells (nasal brushing P < 0.00001 and nasal biopsy P < 0.01). Fluticasone also inhibited seasonal increases in sub mucosal eosinophils (P < 0.001) and significantly reduced nasal symptoms (P < 0.001). CONCLUSION: Nasal brushing harvests sufficient inflammatory cells from the surface of the nasal mucosa to be used in lieu of nasal biopsies in observation of the effect of drugs on the nasal epithelium. PMID- 10520057 TI - Comparison of the effects of fluticasone propionate, aqueous nasal spray and levocabastine on inflammatory cells in nasal lavage and clinical activity during the pollen season in seasonal rhinitics. AB - BACKGROUND: Treatment options for allergic rhinitis include antihistamines, decongestants, anticholinergics, cromolyn sodium and corticosteroids. As the nose is a small organ, comprising less than 1% of total body mass and surface area, it seems logical to confine treatment of rhinitis to the diseased organ. OBJECTIVE: To evaluate the effects of therapy with intranasal fluticasone propionate (FP), both on subjective symptoms and pathophysiological mechanisms, in rhinitis patients during pollen season when the patients were symptomatic. METHODS: We used a double-blind, placebo (PLA)-controlled, randomized, double dummy, parallel group study of the effect of 6 weeks treatment. The double-blind comparison was made between the following three treatments: FP aqueous nasal spray, 200 microg taken once daily, levocabastine (LEV) nasal spray, 200 microg taken twice daily and PLA nasal spray. Clinical evaluation and the levels of cells and mediators in nasal washing were performed before and after treatments. Twenty-four patients (11 men and 13 women, aged 17-50 years, mean age 30.1 +/- 8.5) with strictly seasonal allergic rhinitis to Parietaria entered the study. Clinical evaluation and the levels of inflammatory cells (eosinophils and activated eosinophils, i.e. EG2+) and their mediators (tryptase, eosinophil cationic protein, eosinophil protein X and neutrophil myeloperoxidase) in nasal-lavage were performed before and after treatments. RESULTS: Treatment with FP significantly increased, with respect to placebo, the percentage of days without sneezing (P < 0. 001), nasal blockage (P < 0.001), rhinorrhea (P < 0.001), nasal itching (P < 0.001). Furthermore, treatment with FP showed additional benefits with respect to LEV. The percentage of days without nasal blockage was significantly higher in the FP group that in the placebo group (P = 0.018). The same applied to rhinorrhea (P = 0.009). The percentages of days without sneezing and itching were instead not significantly different between the two groups. As expected, no significant differences were observed in baseline medians of the rhinitis symptom scores as well as in mean values of all mediators and eosinophils in nasal lavages of the various groups under study. After treatment the mean of subjective symptoms as well as all values in nasal lavage level fell significantly only in the FP group, whereas no significant changes were observed either in LEV or PLA groups. Accordingly, significant differences were observed at the end of the treatments between the values of fluticasone group vs LEV and PLA group values. Significant correlations between these values and symptom scores were found, according with literature data suggesting a pathogenetic role for these mediators and eosinophils in rhinitis. CONCLUSION: FP (200 microg once daily) affords a significant degree of improvement in rhinitis control during pollen season, as measured by subjective and objective parameters, compared with LEV (200 microg twice daily) and PLA. The therapeutic benefits of intranasal FP are reflected in, and may be caused by, the decrease in nasal inflammatory cells. PMID- 10520056 TI - Long-term effects of corticosteroid nasal spray on nasal inflammatory cells in patients with perennial allergic rhinitis. AB - BACKGROUND: The effect of long-term topical nasal corticosteroid therapy on nasal inflammatory cells is unclear. OBJECTIVES: To investigate the long-term effect of fluticasone propionate aqueous nasal spray (FPANS) on nasal mucosal inflammatory cells and efficacy in a 1-year study in patients with perennial allergic rhinitis. METHODS: In a 1-year, double-blind, placebo-controlled study of duration we investigated the influence of a topical corticosteroid (FPANS), on Langerhans' cells (CD1a+ cells), T cells, mast cells, eosinophils and macrophages in nasal mucosa in 42 patients with perennial allergic rhinitis. Efficacy was evaluated by nasal symptom score. RESULTS: The FPANS group experienced significantly less sneezing and nasal itching compared with the placebo group. The total symptom score in the FPANS group declined significantly in comparison with baseline (P = 0.007) and placebo group (P = 0.009). After 1 year of active treatment, a significant decrease was seen in the epithelium in numbers of Langerhans' cells, CD3+, CD4+, CD8+ cells, mast cells and eosinophils. In the lamina propria, there was a significant decrease in eosinophils. CONCLUSION: These findings show that FPANS treatment results in a decrease of nasal inflammatory cells. Furthermore, the efficacy of FPANS improves after prolonged treatment. PMID- 10520058 TI - A retrospective study of the relationship between childhood asthma and respiratory infection during gestation. AB - BACKGROUND: Wheeze in children has been found to be associated with prior antepartum haemorrhage and raised levels of IgE in cord blood, and acute wheezing episodes are intimately linked with respiratory viral infections. OBJECTIVE: To assess the relationship between maternal presentation with respiratory tract infections in pregnancy and childhood asthma, taking into account factors which could affect presentation. METHODS: This was a case-control study of 200 asthmatic children, 5-16-year-old, age-matched with one control, having no recorded history of wheeze. Data on respiratory tract infections, maternal wheeze, atopy and smoking was collected from primary care records. Deprivation score was assessed according to small residential areas and subjects were equally distributed between four general practices in Plymouth, UK. RESULTS: Presentation with respiratory tract infections during pregnancy was significantly associated with childhood asthma (OR 1.69, 95% confidence interval 1.05-2.77, P = 0.03). The association was marginally stronger for infections in the first trimester (OR 2.30, 95% CI 1.05-5.41, P = 0.04) and for those with cough during pregnancy (OR 2.24, 95% CI 1.23-4.22, P = 0.007). The associations remained significant after allowing for the effect of the independent variables (gender, maternal smoking, maternal wheeze, allergic rhinitis, eczema, asthma treatment in pregnancy and deprivation [Townsend] score), using multiple logistic regression analysis (ORs and 95% CIs 1.91, 1.14-3.22; 2.32, 1.01-5.34 and 2.29, 1.17-4.48, respectively). There was also an association between numbers of presentations with respiratory infections and childhood asthma (test for trend, P = 0.02). CONCLUSIONS: This study has shown an association between presentation with respiratory infection during gestation and childhood asthma. The results were not affected by the other independent variable factors studied and therefore provide some evidence to support the theory that respiratory viruses may be implicated in the aetiology of asthma. PMID- 10520059 TI - Serum eosinophilic cationic protein and blood eosinophil counts for the prediction of the presence of airways inflammation in children with wheezing. AB - BACKGROUND: Serum eosinophilic cationic protein (ECP) concentrations may be useful noninvasive markers of airways inflammation in atopic asthma. However, the usefulness of serum ECP measurement for the prediction of airways inflammation in children with a history of wheezing is unknown. OBJECTIVE: To determine the test accuracy of serum ECP and blood eosinophil percentage as noninvasive markers of eosinophilic airways inflammation. METHODS: Bronchoalveolar lavage (BAL) fluid and peripheral blood samples for eosinophil percentages and serum ECP were obtained from children undergoing elective surgery and who gave a history of wheezing in the previous year. Sensitivity, specificity and likelihood ratios (LH) and the area under the curve (AUC) for the receiver operator characteristic (ROC) curve were calculated for each blood marker for the prediction of airways inflammation defined by a BAL eosinophil percentage > 0.86. Data were analysed on the basis of how recently symptoms had occurred. RESULTS: Seventy-seven children (median age 6.75 years) were studied. An AUC of 0.75 (log serum ECP concentration) and 0.76 (log blood eosinophil percentage) was obtained for predicting airways inflammation. A serum ECP > 13 microg/L yielded a LH of 4.4, whereas using a cutoff blood eosinophils > 4% yielded a LH of 1.9, for the prediction of elevated eosinophils in BAL. Serum ECP and eosinophil percentages in BAL and blood were lowest (not statistically significant) when last symptoms had occurred more than 12 weeks previously. CONCLUSIONS: Serum ECP and blood eosinophil percentages are useful markers for predicting eosinophilic airways inflammation in wheezing children. PMID- 10520060 TI - VEGF levels in asthmatic airways do not correlate with plasma extravasation. AB - BACKGROUND: Vascular permeability/vascular endothelial growth factor (VEGF) is a multifunctional cytokine which plays a role in chronic inflammation and angiogenesis. Its expression in bronchoalveolar lavage (BAL) has not been determined although VEGF may be relevant to the pathophysiology of asthma in which oedema is an important feature. METHODS: We studied VEGF, albumin and IgA immunoreactive levels in the BAL fluids obtained from 27 chronic stable asthmatics, nine untreated chronic bronchitis patients and 15 control subjects. RESULTS: BAL fluid levels of VEGF and VEGF normalized to IgA were not significantly different in any patient group. Both asthmatic steroid- and non steroid-treated groups had significantly lower albumin levels in their BAL fluids explaining most of the 179% increased VEGF normalized to albumin ratios in non steroid treated asthmatics. Moreover, VEGF and albumin markers correlated in control subjects (r = 0.73, P = 0.006) and in chronic bronchitics (r = 0.75, P = 0.03, Spearman test), but not in asthmatics. VEGF was inversely correlated with asthma severity (GINA/NHLBI scores) in non-steroid treated asthmatics (tau = - 0.52, P = 0.009, Kendall test). CONCLUSIONS: Thus, the potential role of VEGF in asthma requires further studies on bronchial biopsies and induced sputum. PMID- 10520061 TI - Neutrophil activation following TDI bronchial challenges to the airway secretion from subjects with TDI-induced asthma. AB - BACKGROUND: The immunopathological mechanism for occupational asthma induced by toluene diisocyanate (TDI) remains to be further clarified. There have been few reports suggesting involvement of neutrophils in inducing bronchoconstriction after TDI inhalation. OBJECTIVES: To further understand the role of neutrophils in the pathogenesis of TDI-induced asthma. MATERIALS AND METHODS: Eight TDI induced asthmatic subjects were classified as group I, and five exposed workers who had complained of work-related symptoms and worked in the same workplace, but showed negative bronchial challenges were enrolled as controls (group II). Serum neutrophil chemotactic activity during TDI bronchial challenge test was measured by the Boyden chamber method. Induced sputum was collected before and after the TDI bronchial challenge test. The myeloperoxidase (MPO) and interleukin (IL) -8 levels in the sputum were measured using RIA and ELISA. RESULT: Serum neutrophil chemotactic activity significantly increased at 10 min (P = 0.01), then decreased at 60 min (P = 0.02) and remained unchanged for up to 420 min (P = 0.07) in group I subjects, while no significant changes were found in group II subjects (P > 0.05). MPO and IL-8 were abundantly present in the sputum of all the TDI-induced asthmatic subjects and they increased significantly at 420 min after the bronchial challenges (P = 0.02, P = 0.03, respectively), while no significant changes were noted in group II subjects (P > 0.05). CONCLUSION: These findings support the view that activated neutrophils may contribute to bronchoconstriction induced by TDI which may be associated with IL-8 release. PMID- 10520062 TI - Means of increasing sensitivity of an in vitro diagnostic test for aspirin intolerance. AB - BACKGROUND: Pseudo-allergic reactions caused by aspirin (acetyl salicylic acid; ASA) often resemble immediate-type hypersensitivity reactions consisting of urticaria and angioedema or rhinoconjunctivitis, asthma and nasal polyps. In the last few years, a new in vitro assay based on determination of sulfidoleucotrienes from isolated leucocytes (cellular allergen stimulation test CAST) has been introduced for type I allergies and pseudoallergic reactions. In ASA intolerance, there is only limited experience using this assay with - in some studies - only moderate sensitivity. Furthermore, the necessity to use freshly isolated leucocytes from untreated patients is inconvenient for routine settings. OBJECTIVE: The purpose of our study was to search for possibilities of increasing the sensitivity of the test and to use stored blood samples which would permit shipping, two requirements for the clinical suitability of this test. PATIENTS AND METHODS: Leucotriene release in response to ASA and other non-steroidal anti inflammatory drugs (NSAIDs) was analysed in 38 ASA-intolerant patients (predominantly airway-related symptoms n = 22; predominantly cutaneous symptoms n = 16) and 50 controls. The diagnosis of ASA intolerance was established by history and placebo-controlled oral challenge tests. RESULTS: Using 24 h-stored leucocytes obtained from 10 ASA-intolerant patients and 10 healthy controls there were no significant differences of leucotriene release by resting, ionomycin-, and anti FcepsilonRIalpha-stimulated leucocytes compared with freshly isolated leucocytes. Analysis of ASA + C5a-mediated leucotriene release by stored blood samples in combination with indomethacin- and diclofenac-mediated leucotriene release in ASA-intolerant patients (n = 38) resulted in an increased sensitivity (from 50 to 72.7% in ASA-intolerant patients with predominantly airway-related symptoms and from 81 to 100% in ASA-intolerant patients with predominantly skin symptoms) compared with assays in which only ASA + C5a-mediated leucotriene release has been determined. Moreover, the specificity of the assay remained high (96.7% when analysing different NSAIDs compared with > 99% when analysing only ASA + C5a-mediated leucotriene release). CONCLUSION: In vitro stimulation with ASA + C5a leucocyte stimulation with other NSAIDs should be performed to achieve a higher sensitivity. This finding can be explained by the clinical observation of a high ratio of cross-reactivities between the mentioned NSAIDs. PMID- 10520063 TI - Amalgam allergy associated with exacerbation of aspirin-intolerant asthma. AB - BACKGROUND: Aspirin-intolerant asthma can be induced not only by acidic analgesics (including acetylsalicylic acid), which effectively inhibit cyclo oxygenase, but also by cross-reactivity with paraben, and other chemical additives. OBJECTIVE: We examined whether amalgam allergy is involved in the pathogenesis of a aspirin-intolerant asthma. METHODS: We present the first case of aspirin-intolerant asthma that improved after the removal of dental amalgam. In addition, we performed both the methacholine provocation testing and sulpyrine provocation testing before and after the removal of dental amalgam. RESULTS: In addition, the methacholine concentration causing a 20% fall in FEV1 in provocation tests rose significantly, though hypersensitivity to analgesics evaluated with sulpyrine provocation testing did not decrease. These results suggest that amalgam sensitization is involved in bronchial hyperresponsiveness in aspirin-intolerant asthma. CONCLUSION: Sensitivity to amalgam may cause exacerbation of aspirin-intolerant asthma in some patients. To the best of our knowledge, this is the first case report of amalgam allergy associated with aspirin-intolerant asthma. PMID- 10520064 TI - The contribution of 'holiday deaths' to seasonal variations in asthma mortality in England and Wales. AB - BACKGROUND: In younger age groups there is a summer peak in asthma deaths, but whether this is due to social or environmental factors is not known. One suggested social factor is a summer holiday away from home during which there may be a lack of compliance with medication or greater difficulty in getting medical help. In older age groups there is a winter peak in asthma deaths. OBJECTIVE: To study the contribution of 'holiday deaths' to seasonal variations in asthma mortality in England and Wales. METHODS: Routinely collected mortality statistics were used (all asthma deaths of persons dying in England and Wales, 1991-93 and 1995). Deaths occurring 40 miles or more from home were estimated using the District Health Authority in which the person was usually resident; the registration district of death; and the 'transferability code' (derived by the Office for National Statistics). RESULTS: There were 484 asthma deaths in people aged 0-34 years, and 6337 asthma deaths in people aged 35 years or more. Deaths estimated to occur 40 miles or more from home contributed little (16%) to the summer peak in asthma deaths age 0-34 years, and nothing to the winter peak in asthma deaths age 35 years or more. CONCLUSIONS: Holidays away from home do not play an important part in explaining the seasonal variation of either young or old asthma deaths. Other social or environmental factors are more important. PMID- 10520065 TI - Allergy to the pine processionary caterpillar (Thaumetopoea pityocampa). AB - BACKGROUND: The contact with the pine processionary caterpillar (Thaumetopoea pityocampa) induces dermatitis and ocular lesions by a mechanic and toxic mechanism. However, IgE-mediated hypersensitivity to this caterpillar has been demonstrated in two recent studies. OBJECTIVE: To find if an IgE-mediated mechanism was operative among patients with suspected previous reactions to processionary caterpillars, particularly exposed workers. METHODS: Fifty-five patients were studied by skin prick test (SPT), and specific IgE detection by immunoblotting. RESULTS: A total of 58.18% patients had a positive SPT for caterpillar extract. Positive SPT patients had more generalized cutaneous reactions (47%) and oedema (50%) as well as a shorter latency period (mean, 36 min) and duration of cutaneous lesions (mean, 26 h) than the patients with negative SPT. A total of 60% of the positive SPT patients had occupational exposure to the processionary caterpillar. The occupationally exposed workers showed significant symptoms from October to December. The anaphylactic reactions only appeared in allergic patients with occupational exposure and were also more frequent from October to December. These patients with anaphylactic reactions had a major size of SPT and the exercise was found in them to be a variable that increased the symptoms. The IgE immunoblot detected in the caterpillar extract several reactive bands with apparent molecular weights from to 35-4 kDa in 72% of the cases with positive SPT. CONCLUSIONS: Allergic reactions to T. pityocampa urticating hairs have different clinical characteristics than those induced by a toxic-irritative mechanism and are more frequent than suspected. Allergic reactions to this caterpillar among occasional visitors to pine-wood areas, and particularly in pine-forest workers, should be taken into consideration by allergists. PMID- 10520066 TI - Interleukin-10 is localized to and released by human lung mast cells. AB - BACKGROUND: Mast cells control the local inflammation by producing many kinds of cytokines. Interleukin (IL)-10 is one of the important cytokine that upregulate or downregulate inflammation. OBJECTIVE: The aim of this study is to ascertain whether IL-10 is produced from human lung mast cells by cross-linkage of high affinity Fcepsilon receptors (FcepsilonRI). METHODS: Mast cells were purified using affinity magnetic selection with mAb YB5.B8 (> 93% pure). Mast cells were precultured with human myeloma IgE (3 microg/mL) for 16 h and then washed, and stimulated with anti-IgE in the presence or absence of recombinant human stem cell factor (rhSCF). We have studied the production of IL-10 by using reverse transcription-PCR, enzyme-linked immunosorbent assay and immunocytochemistry. RESULTS: We found that human lung mast cells were immunocytochemically stained with anti-IL-10 mAb after IgE-dependent stimulation. The activation of mast cells via FcepsilonRI enhanced the intensity of the IL-10 mRNA signal. Anti-IgE (1 microg/mL) induced a median IL-10 release of 301.7 (7.8-1532.4) pg/106 mast cells/24 h. In contrast, mast cells released only a small amount of IL-10 in the absence of anti-IgE. This difference was statistically significant (P = 0.02, n = 11). CONCLUSION: Our findings indicate that human lung mast cells are capable of producing IL-10 in response to IgE-dependent stimulation. PMID- 10520067 TI - Prevalence of atopy and urban air pollution: dirty business. PMID- 10520068 TI - Indoor fungal exposure--does it matter, and what can be done about it? PMID- 10520069 TI - Allergen immunotherapy and mast cells. PMID- 10520070 TI - Anti-inflammatory mechanisms of leukotriene modulators. PMID- 10520071 TI - Eosinophil markers in childhood asthma. PMID- 10520072 TI - Airway wall remodelling: does it occur and what does it mean? PMID- 10520073 TI - A connectionist theory of asthma. PMID- 10520074 TI - Gaseous air pollution and atopy. AB - BACKGROUND: Photochemical air pollutants are commonly thought to be implicated in the gradual increase in the prevalence of atopy. However, no epidemiological data are available. METHODS: To clarify this issue, we performed a cross-sectional epidemiological survey in 2604 primary school children, 10 and 11 years old, living in seven communities among which some have the highest photochemical exposure in France. The mean levels of the main gaseous air pollutants (SO2, NO2 and O3) were measured during a 2-month period in 1993. The protocol included a standardized questionnaire, skin prick tests to common aeroallergens and in the atopic children, collection of a sample of mattress dust to measure group 1 mite allergens. Atopy was only defined on the basis of the skin prick tests. RESULTS: Percentage of positive skin tests and the number of positive skin tests were similar in the different communities looked at. The distribution of dust samples with a group 1 allergen level greater than 2 microg/g dust, was also similar. Logistic regression analysis including potential confounding factors, as well as the mean level of air pollutants, did not demonstrate any association between atopy and mean SO2, NO2 and O3 levels. CONCLUSION: The increase in photochemical air pollutants is unlikely to be a major determinant for the recent increase in the prevalence of atopy. PMID- 10520075 TI - Prevalence and residential determinants of fungi within homes in Melbourne, Australia. AB - BACKGROUND: Recent epidemiological studies suggest that the adverse respiratory health effects caused by the inhalation of fungal propagules are substantial. Knowledge of the prevalence and environmental determinants of indoor fungal levels is essential in designing effective avoidance measures. AIM: To investigate the prevalence of fungi and the influence of residential characteristics on levels of fungi within homes in Melbourne, Australia. METHODS: Floor dust and air samples were collected from bedrooms in 485 houses over 1 year. The dust was analysed for ergosterol, a marker of cumulative fungal biomass exposure. Total and genera-specific fungal propagules were identified in air samples. Details of the relevant residential characteristics were documented using a questionnaire. Independent predictors (P < 0.05) of ergosterol and total fungal propagules were identified by multiple linear regression. RESULTS: Fifty five percent of the houses had viable fungal propagules exceeding 500 CFU/m3. Cladosporium and Penicillium were identified as the most prevalent and abundant fungal genera in indoor air. The median ergosterol level in bedroom floor was 3.8 microg/g of dust. Multivariate analysis showed that total fungal propagules in indoor air were lower in bedrooms with a ceiling fan, without visible mould, and those that were more frequently vacuumed, had a solid fuel fire, had windows closed at the time of the sampling or lacked pets. The presence of more than one cat had the greatest effect on total fungal propagules. Ergosterol levels were significantly lower in homes without old fitted carpets, visible mould or pets and those with frequent airing and regular use of an extractor fan in the kitchen. Old wall-to-wall carpets had the greatest effect on ergosterol. CONCLUSIONS: High indoor fungal exposures were associated with infrequent ventilation or vacuuming, presence of pets, visible mould and old carpets. PMID- 10520076 TI - Grass pollen immunotherapy decreases the number of mast cells in the skin. AB - BACKGROUND: Allergen injection immunotherapy is effective for summer hay fever and reduces cutaneous sensitivity to grass pollen. OBJECTIVE: We have addressed whether this effect of immunotherapy may be due to a decrease in mast cell numbers in the skin. METHODS: Total mast cells and mast cell subtypes in the dermis were measured by dual immunocytochemistry in 40 adult patients who had received either 'active' grass pollen immunotherapy or placebo injections for 9 months in a double-blind clinical trial. RESULTS: Clinical improvement in hay fever was accompanied by a greater than 10-fold reduction in the immediate cutaneous response to grass pollen (P = 0. 0002) and a sevenfold decrease in mast cell numbers in the skin (P = 0.0001). The number of mast cells after immunotherapy correlated with the clinical response in terms of seasonal symptoms (r = 0.61, P = 0.001) and rescue medication use (r = 0.75, P = 0.0001). Specific double immunostaining showed that the majority of mast cells (greater than 60%) were tryptase/chymase-positive (MCTC) and the remainder tryptase-only (MCT) cells. Following immunotherapy both subtypes were equally reduced. CONCLUSION: One mechanism by which immunotherapy may act is to reduce mast cell numbers with a consequent reduction in immediate allergic sensitivity. PMID- 10520077 TI - Circadian variation of urinary eosinophil protein X in asthmatic and healthy children. AB - BACKGROUND: It is suggested that urinary eosinophil protein X (EPX) is a noninvasive tool to monitor bronchial inflammation in asthmatic children. However, circadian variation of the number and activation of eosinophils might possibly influence urinary EPX excretion. OBJECTIVE: Measurements of urinary EPX (radioimmunoassay) were used to investigate circadian variation of eosinophilic activation and to monitor bronchial inflammation in children with asthma before and after treatment with corticosteroids. METHODS: Urinary EPX excretion (microg/mmol creatinine) was measured in the morning and afternoon in 22 stable asthmatics and in 16 nonatopic, nonasthmatic controls to investigate circadian variation. Additionally, EPX excretion in the afternoon was analysed in 21 children with chronic asthma before and after 6 weeks of treatment with inhaled corticosteroids, and in seven children within 24 h of admission due to an asthma exacerbation and again 3 months after discharge. RESULTS: EPX excretion in the first morning urine sample of the day compared with the afternoon urine sample was significantly higher both in children with asthma (n = 22; mean +/- standard deviation: 179.7 +/- 97.3 vs 60.9 +/- 40.7 microg/mmol creatinine, P = 0.0001) and in nonatopic nonasthmatic controls (n = 16; 114.5 +/- 57.1 vs 53.4 +/- 29.0 microg/mmol creatinine, P = 0.0001). EPX excretion decreased significantly after 6 weeks of anti-inflammatory treatment in the group of children with chronic asthma (n = 21; 124.7 +/- 84.6 vs 87. 5 +/- 61.9 microg/mmol creatinine, P = 0.02) and in the group of children with an acute asthma exacerbation 3 months after discharge (n = 7; 233.2 +/- 174.5 vs 75.8 +/- 59.5 microg/mmol creatinine, P = 0.02). CONCLUSION: This study suggests a circadian variation of EPX excretion in children with asthma and in nonatopic, nonasthmatic controls. Measurement of EPX excretion is helpful monitoring therapy in asthmatic children if circadian variation is considered. PMID- 10520078 TI - Eosinophil protein X and eosinophil cationic protein as indicators of intestinal inflammation in infants with atopic eczema and food allergy. AB - BACKGROUND: The aim of this study was to evaluate the presence of allergic intestinal inflammation in infants with food allergy and atopic eczema before and after elimination diet, and to evaluate the use of eosinophil protein X (EPX) and eosinophil cationic protein (ECP) in the monitoring of inflammatory activity. METHODS: The study material comprised 25 infants with atopic dermatitis and food allergy. Thirteen healthy infants served as controls. Faecal and serum samples were collected before an elimination diet (on the first visit to the hospital) and approximately 3 months later for the determination of EPX and ECP. RESULTS: Before the elimination diet, infants with atopic dermatitis demonstrated markedly higher faecal concentrations of EPX and ECP than healthy controls (P = 0. 0003, P < 0.0001, respectively). The faecal concentrations of EPX and ECP showed a distinct decrease as a result of an adequate elimination diet in patients with favourable clinical response (P = 0.0027, P = 0.004, respectively). CONCLUSIONS: The results indicate the presence of marked intestinal inflammation in patients with atopic dermatitis and food allergy. The determination of faecal ECP and especially of faecal EPX provides a promising noninvasive tool in monitoring intestinal inflammation and disease activity in infants with atopic eczema and food allergy. PMID- 10520079 TI - Intracellular expression and serum levels of eosinophil peroxidase (EPO) and eosinophil cationic protein in asthmatic children. AB - BACKGROUND: Eosinophils are involved in the chronic inflammatory response in asthma and their basic proteins are thought to play a major pathophysiological role in this process. While serum levels of basic proteins have been used to monitor the ongoing allergic disease, little is known about the intracellular expression of these proteins in clinical situations. OBJECTIVE: The aim of the study was to determine the intracellular expression of eosinophil cationic protein (ECP) and eosinophil peroxidase (EPO) in asthmatic children and control subjects and relate it to serum levels of both proteins, lung function tests and immunoglobulin (Ig)E levels. METHODS: Serum ECP and EPO concentrations were determined by immunoassays in 13 asthmatic children (mean age: 9 +/- 1 years, mean FEV1: 92 +/- 10% predicted, geometric mean PC20 histamine 0.5 mg/mL) and 10 age-matched, healthy control subjects. A flow cytometric single cell assay was employed to detect intracellular ECP and EPO in peripheral blood eosinophils. RESULTS: While serum concentrations of both ECP (asthma: median 15.0 microg/L [range 3.6-57.7] vs control: 5.9 microg/L [2.7-9.1]; P = 0.02) and EPO (22.9 microg/L [5.2-82.5] vs 7. 2 microg/L [2.5-12.7]; P = 0.008) were significantly elevated in asthmatics, the intracellular expression of ECP and EPO (measured as mean fluorescence intensity) was decreased (EG1: 55.3 [17.7-120.8] vs 100.3 [46.5 264.4]; P = 0.01; EG2: 80.2 [24.1-135.3] vs 133.7 [32. 1-244.9]; P = 0.04 and EPO: 49.7 [23.1-155.8] vs 94.9 [28.8-115.2]; P = 0.03). In asthmatics there was a significant correlation of FEV1 with intracellular ECP and of bronchial hyperresponsiveness with serum EPO and ECP. Furthermore, total IgE levels were positively correlated with serum EPO only. CONCLUSION: We conclude that in asthmatics the intracellular content of ECP and EPO in peripheral eosinophils is reduced possibly due to degranulation. Epitope masking in activated eosinophils or a shift to early bone marrow-derived progenitors with less granule proteins are further possible explanations. PMID- 10520080 TI - Characterization of eosinophils and detection of eotaxin in skin chamber fluid after challenge with relevant allergen in patients with mild asthma. AB - BACKGROUND: A selective recruitment of eosinophils to sites of allergic inflammation is suggested to be controlled by regulation of cytokines, chemokines and adhesion molecules. OBJECTIVE: The aim of this study was to examine whether allergen challenge in skin chambers, applied on patients with allergic rhinitis and mild asthma, results in a selective influx of activated eosinophils and detectable levels of cytokines/chemokines related to eosinophil recruitment, such as interleukin (IL)-5 and eotaxin. METHODS: A skin blister was induced on the volar aspect of each forearm; one contained PBS-heparin buffer (control) and the other was challenged with relevant allergen. Peripheral blood was drawn before the allergen was applied to the skin chamber, and the expression of CD9, CD11b and EG2-epitope on intracellular eosinophil cationic protein (ECP) was analysed in eosinophils. Chamber fluid was collected 8 h after allergen application and analysed for differential cell counts, expression of eosinophil activity markers, the presence of ECP, eotaxin, and IL-5. RESULTS: The number of recruited leucocytes was equal in the allergen-challenged chambers and in controls. However, the number of eosinophils was significantly increased in the allergen challenged chambers, and elevated levels of released ECP were measured. Moreover, the eosinophils recruited were activated, as shown by increased expression of EG2 and CD11b, and decreased expression of CD9, in comparison with blood eosinophils. In the skin chamber fluids, higher levels of eotaxin were detected in the allergen-challenged chambers than in controls, but there were no detectable levels of IL-5. CONCLUSION: We have demonstrated a selective recruitment of eosinophils, and higher levels of released ECP and eotaxin, in skin chambers stimulated with allergen, as compared with control chambers. Allergen challenge in skin chambers is a useful tool for studies of eosinophil recruitment, their state of activation, and their involvement in the allergic inflammatory response. PMID- 10520081 TI - Sputum cellular and cytokine responses to inhaled endothelin-1 in asthma. AB - BACKGROUND: Endothelin (ET)-1 is a 21-amino acid peptide which has potent bronchoconstrictor activity. Animal studies show elevation of ET-1 during experimental airway inflammation, and inhibition of inflammation by endothelin antagonists, suggesting pro-inflammatory activity for ET-1. OBJECTIVE: We wanted to assess any acute influence that bronchoconstrictor doses of inhaled ET-1 might have on cells, tumour necrosis factor (TNF)-alpha, interleukin (IL)-1beta, nitrite (NO2) and albumin in induced sputum in asthma. METHODS: Bronchial challenge was performed using nebulized ET-1 (nebulized dose range 0.96-15.36 nmol) and placebo in 10 adult asthmatic subjects in a randomized double-blind placebo-controlled cross-over study. Sputum induction was performed 30 min and 4 h after placebo or ET-1 bronchial challenge. RESULTS: All subjects experienced dose-dependent bronchoconstriction to inhaled ET-1 with a mean (range) PC15 forced expiratory volume in 1 s (FEV1) to ET-1 of 9.45 (1.2-21.7) nmol. Comparing ET-1 with placebo inhalation, there was no change in sputum differential cell counts, TNFalpha, IL-1beta, NO2 or albumin at 30 min or 4 h after inhalation, nor was there a difference in these parameters at 4 h compared with 30 min after ET-1 inhalation. There was no fall in FEV1 at 4 h after ET-1 inhalation, suggesting that ET-1 inhalation is not associated with a late bronchoconstrictor response. CONCLUSIONS: We conclude that inhaled ET-1 does not appear to stimulate an acute inflammatory response in asthma as assessed by differential cell count, TNFalpha, IL-1beta, NO2 and albumin concentrations in induced sputum. PMID- 10520082 TI - Effects of ONO-1078 (pranlukast) on cytokine production in peripheral blood mononuclear cells of patients with bronchial asthma. AB - BACKGROUND: ONO-1078 (pranlukast) is a leukotriene receptor antagonist developed in Japan. This drug has been shown to be useful in oral treatment of bronchial asthma. The present study was undertaken to assess the effects of this drug on the production of cytokines in the peripheral blood mononuclear cells of patients with asthma under stimulation with specific antigens. METHODS: Peripheral blood mononuclear cells from mite antigen-positive asthmatic patients (immunoglobulin E RAST score > 3) were incubated for 72 h in the presence of mite antigen (10 microg/mL). The supernatant of the culture was subjected to enzyme-linked immunosorbent assay (ELISA) to quantify interleukin (IL) -4, IL-3, IL-5, and granulocyte macrophage-colony stimulating factor (GM-CSF). Other peripheral blood mononuclear cells from the same patients were incubated for 72 h in the presence of both mite antigen (10 microg/mL) and ONO-1078 (0.5, 1, or 10 microg/mL), followed by ELISA of the supernatant to quantify the cytokines. RESULTS: Production of IL-4, IL-5, and GM-CSF by mononuclear cells under stimulation with mite antigen was markedly suppressed when they were exposed to ONO-1078 at a concentration of 10 microg/mL. CONCLUSION: The results suggest that ONO-1078 acts directly on peripheral blood mononuclear cells and that blockade of leukotriene receptors on blood mononuclear cells by the cysteinyl-leukotriene receptor antagonist (LTRA) pranlukast (ONO-1078) can dose-dependently inhibit release of immunoreactive TH2-type cytokines (IL-3, IL-4, GM-CSF, and possibly IL-5), but not of the TH1-type cytokine IL-2, when stimulated by mite allergen in vitro. The data may provide clues to the mechanism by which a number of LTRA including zafirulukast and montelukast can reduce airway, sputum and blood eosinophil counts in clinical asthma. It supports animal studies showing that anti-IL-5 antibodies partially block cys-LT-induced airway eosinophilia, suggesting that cys-LTs may cause secondary release of IL-5 from an unknown cell-type. These findings indicate that ONO-1078 suppresses the production of IL-4 (a cytokine that affects IgG antibody production), IL-5, and GM-CSF (cytokines that affect eosinophil activation) by peripheral blood mononuclear cells under stimulation with specific antigens in patients with bronchial asthma. Because of its anti inflammatory effects, ONO-1078 should be useful in the treatment of bronchial asthma. PMID- 10520083 TI - Increased carbon monoxide in exhaled air of patients with seasonal allergic rhinitis. AB - BACKGROUND: Carbon monoxide (CO) can be detected in exhaled air and is increased in asthmatic patients. However, it is uncertain whether exhaled CO is increased in patients with allergic rhinitis. OBJECTIVE AND METHODS: To study whether exhaled CO is increased in patients with allergic rhinitis, exhaled CO concentrations were measured on a CO monitor by vital capacity manoeuvre in 86 patients with seasonal allergic rhinitis during and out of the cedar pollen season. RESULTS: During the season, exhaled CO concentrations were 3. 6 +/- 0.3 p.p.m. and decreased to 1.2 +/- 0.1 p.p.m. out of the season. The values of exhaled CO out of the season were similar to those in age-matched non-smoking healthy control subjects (1.2 +/- 0. 1 p.p.m.). Exhaled CO concentrations were significantly higher in patients with symptoms than in those without symptoms (P < 0.01). Exhaled CO concentrations in patients did not differ significantly among oral and nasal exhalation, and oral exhalation with an expiratory resistance (P > 0.20). CONCLUSION: These findings suggest that allergic rhinitis increases the concentration of CO in exhaled air and increases in exhaled CO may be derived from lower airways. PMID- 10520085 TI - T-cell reaction to local anaesthetics: relationship to angioedema and urticaria after subcutaneous application--patch testing and LTT in patients with adverse reaction to local anaesthetics. AB - BACKGROUND: Local anaesthetics are known to elicit T-cell reactions after epicutaneous application, namely contact dermatitis. In addition, adverse reactions like urticaria and angioedema are rather common after submucosal or subcutaneous injection. The pathogenesis of these side-effects, which appear frequently hours after application, is unknown, but thought to be not immunoglobulin E-mediated, since immediate skin tests are mostly negative. OBJECTIVES: We investigated whether patients who developed urticaria and angioedema after subcutaneous application have a T-cell sensitization to local anaesthetics, which might be responsible for the symptoms. METHODS: Twenty patients with generalized and/or local cutaneous reactions after LA were examined with intradermal testing using a standard panel of six LAs and patch testing using between seven and nine LAs in vaseline and four LAs in PBS. In 10 patients, a lymphocyte transformation test (LTT) was performed. RESULTS: Only 2/20 patients had an immediate skin reaction (positive intradermal test), whereas 6/20 patients had a positive delayed skin reaction (positive patch test). In 6/10 subjects the LTT was positive. CONCLUSIONS: Delayed appearance of urticaria and angioedema after subcutaneous application of local anaesthetics may be related to a T cell- mediated sensitization, which might be detected by patch testing or LTT. PMID- 10520084 TI - Submucous turbinectomy decreases not only nasal stiffness but also sneezing and rhinorrhea in patients with perennial allergic rhinitis. AB - BACKGROUND: Turbinate surgery, in which mucous epithelium is resected, has often been used for patients with perennial allergic rhinitis, since the mucous epithelium is the principal site for immunoglobulin (Ig)E-mediated allergic reactions and chronic inflammation. OBJECTIVE: To assess the effect of submucous turbinectomy on allergic rhinitis. METHODS: Sixty patients with severe perennial allergic rhinitis underwent submucous turbinectomy and were followed-up for 1 year. Nasal symptoms were evaluated with a standard symptom score. Rhinometry was used to evaluate nasal congestion, and nasal provocation tests in vivo were performed to evaluate allergic reactions. In 16 cases, biopsies from the nose were also available for immunohistochemical analysis. These examinations were performed before and after submucous turbinectomy. RESULTS: The mean total nasal symptom score (7.2 +/- 1.7, mean +/- SD before surgery) was significantly reduced after surgery (1.2 +/- 1.4, P < 0.0001), and the effect of the surgery on nasal symptoms continued for at least 12 months (1.9 +/- 1.8, P < 0.0001). Submucous turbinectomy reduced both nasal discharge and sneezing, as well as nasal stiffness. Histopathological examination following surgery revealed that the lamina propria was occupied by fibrous tissues, and that the number of vessels, nasal glands, eosinophils and infiltrating IgE+ cells decreased in the turbinate. There were no significant differences in the levels of either house dust mite specific or non-specific IgE in the serum between before and after surgery. CONCLUSION: Submucous turbinectomy preserving the ciliary epithelium is a powerful strategy for improving nasal symptoms induced by allergic reaction via the reduction in the number of allergy-related cells in the nose. PMID- 10520086 TI - Association of FcepsilonR1-beta polymorphisms with asthma and associated traits in Australian asthmatic families. AB - BACKGROUND: Asthma is a genetically complex disease, and is characterized by elevated serum immunoglobulin E (IgE) levels, elevated blood eosinophil counts and increased airway responsiveness. Polymorphisms in the beta subunit of the high affinity receptor for IgE (FcepsilonR1-beta) have been previously associated with these phenotypes and with an increased risk of asthma. OBJECTIVE: To investigate the association of all known bi-allelic polymorphisms in FcepsilonR1 beta to asthma and quantitative traits associated with asthma in a selected sample of Australian asthmatic children and their nuclear families. METHODS: Australian Caucasian nuclear families (n = 134 subjects) were recruited on the basis of a child proband with current, severe, symptomatic asthma. The quantitative traits assessed included serum levels of total IgE and specific IgE to house dust mite and mixed grass, blood eosinophil counts and the dose-response slope of the forced expiratory volume in 1 s to histamine provocation. RESULTS: Neither the Leu181 nor the E237G mutations were detected in this population. Allele B of RsaI intron 2 (RsaI_in2*B) was significantly associated with physician-diagnosed asthma (ever) (P = 0.002). Alleles of both the RsaI_in2 and RsaI exon 7 (RsaI_ex7) polymorphisms were significantly associated with loge total serum IgE levels and the combined RAST index. RsaI_ex7 was also associated with loge blood eosinophil counts. These associations were independent of age, sex and familial correlations. CONCLUSION: This study supports a role for the FcepsilonR1-beta gene or a nearby gene in the pathogenesis of asthma. PMID- 10520087 TI - The use of phage-peptide libraries to define the epitope specificity of a mouse monoclonal anti-Der p 1 antibody representative of a major component of the human immunoglobulin E anti-Der p 1 response. AB - BACKGROUND: More than 80% of individuals who are sensitive to the dust mite Dermatophagoides pteronyssinus produce immunoglobulin (Ig) E antibodies to Der p 1, the most significant domestic allergen. There is therefore considerable interest in elucidating the interaction between human IgE and Der p 1, as a basis for developing strategies for therapeutic intervention. OBJECTIVES: We have therefore sought to determine the Der p 1 epitope recognized by a mouse monoclonal anti-Der p 1 antibody (mAb 2C7) representative of a major component of the human IgE anti-Der p 1 response. METHODS: M13 15mer and T7 9mer bacteriophage peptide display libraries were screened with mAb 2C7. Mimotope sequences were defined and compared with the native Der p 1 sequence and with those of three homologous molecules, namely chymopapain, papain and actinidin. The sequence of a candidate epitope was then located in the three-dimensional model of Der p 1 and the corresponding sequences in the homologous molecules were studied for accessibility in the three-dimensional structure. RESULTS: We have demonstrated that it is possible to isolate phage clones with peptide inserts specific for mAb 2C7. Examination of the sequences obtained and the location of the corresponding epitope within the three-dimensional model of Der p 1 has shown that mAb 2C7 recognizes a conformational epitope comprising the sequence Leu147-Gln160. The relevance of the identified epitope was established by showing that native Der p 1 can block the binding of specific phage clones to mAb 2C7. Similar sequences were identified within the three-dimensional structures of chymopapain, papain and actinidin, thereby providing a structure-based explanation for immunological cross-reactivity. CONCLUSION: The identification of the Der p 1 sequence Leu147 Gln160 as a potential epitope recognized by a major component of the human IgE anti-Der p 1 response may provide therapeutic opportunities for disrupting the interaction between IgE and this important allergen. PMID- 10520088 TI - The prevalence of immunoglobulin E antibodies to the proteins of rubber (Hevea brasiliensis) latex and grass (Phleum pratense) pollen in sera of British blood donors. AB - BACKGROUND: Although there have been many studies of the prevalence of latex allergy in populations deemed to be at risk, little is known of the potential allergic susceptibility to latex products prevailing in the general population. OBJECTIVE: To assess the possible prevalence of allergy to latex goods in a population of blood donors by measurement of specific antilatex immunoglobulin (Ig) E in blood, to relate this to prevalence of antigrass IgE in the blood donations, and to assess the prevalence of antibodies to grass, house dust mite and cat allergens in those donors having antilatex IgE antibodies. METHODS: Sera from two groups of donations obtained in the English West Midlands were assayed. A group of 2000 donations obtained in midwinter was assayed for antilatex and antigrass pollen IgE. A group of 5000 midsummer donations was assayed for total IgE, and antilatex IgE and the sera giving a positive reaction, assayed for antigrass pollen, antihouse dust mite and anticat IgE. The nature of the principal latex and grass pollen polypeptides reacting with IgE in the sera was assessed by immunoblotting. RESULTS: Anti-latex IgE was detected in approximately 4% of the winter and 7% of the summer donations. The prevalence of antigrass IgE in the winter donations was approximately 20% and amongst the latex-positive sera approximately 84% contained antigrass IgE. Of the summer donations of latex positive sera, 96% contained antigrass, 48.6% antimite IgE and 34% anticat IgE. The prevalence of both antilatex and antigrass IgE was age and sex related. Inhibition studies indicated cross-reactivity of IgE with latex and grass pollen proteins. CONCLUSIONS: Whilst 4-7% of the population may have serum IgE reacting with latex, the levels are low compared with those reacting with the aeroallergens studied. The apparent cross-reactivity of some serum IgE with both latex and grass pollen taken with other evidence suggests that, in some individuals, allergy to latex may arise from an initial sensitization by grass pollen. PMID- 10520089 TI - Plagiarists publish and perish. PMID- 10520090 TI - Pre-registration diploma students: a quantitative study of entry characteristics and course outcomes. AB - Nurse education has been transformed over the last decade and continuing change is likely. Nurse educators are responsible for meeting the quality assurance standards of local stakeholders and student retention and progress are important aspects of this process. As part of a monitoring exercise, an enquiry was set up to review pre-registration selection and recruitment strategies and to establish if there were any significant relationships between the characteristics of pre registration diploma entrants and their academic achievement or completion rates. A multi-factorial tree-based technique was used for this purpose. This is one of the first British studies to consider both academic performance and completion rates for pre-registration diploma students. Four cohorts (N = 355) were studied. There was marked heterogeneity in student characteristics with a wide age distribution, a significant proportion of mature entrants with previous care experience, and considerable diversity in terms of education. Education and age were significant predictors of academic achievement: entrants with a minimum of two A levels and mature women with recent study experience did particularly well. Younger recruits with modest educational qualifications on entry performed less well in their assessments of theoretical knowledge. Younger students tended to leave more regularly, and well-qualified entrants showed a greater tendency to complete, although these relationships were not statistically significant. Multi factorial analysis demonstrated that organisational and course characteristics have a conjoint influence on course outcomes. Although the study is concerned with Project 2000 in the United Kingdom, there are lessons to be drawn concerning the selection and support of non-traditional recruits into nursing. PMID- 10520091 TI - Assessing educational effectiveness: the impact of a specialist course on the delivery of care. AB - An exploratory study was undertaken in South Wales to assess the changes in clinical practice brought about by a specialized pharmacology module designed for Community Mental Health Nurses (CMHNs), in our institution. The respondents were the seven CMHNs who completed the course in 1997 and returned to clinical practice, and seven CMHN comparators, matched on the basis of work experience and location. In order to assess the impact of the module, the practice, attitudes and knowledge of the respondents were investigated before and after the course and 6 months later, using semistructured interviews, nonparticipant observation and questionnaires. The three data sets were analysed using the constant comparative method, and relevant themes were identified and refined. While we obtained some objective measures of positive educational impact, these should be considered in relation to contextual and confounding variables. Both reported and observed behaviours indicated that the main benefit from the course was increased awareness and monitoring of the side-effects of medication. Respondents had not implemented change uniformly; several factors determined how they modified working practices, including service pressures and the support of colleagues. PMID- 10520092 TI - Taking care as a relationship: a phenomenological view. AB - The purpose of this study was to understand and analyse the meanings given to caring of patients on the isolation ward by nursing students, focusing particularly on the aspects of communication and interpersonal relationships. The data were collected from individual interviews with 18 nursing students who were performing nursing practice on the isolation ward. The results, analysed and interpreted according to existential phenomenology, describe the structure of the phenomenon 'taking care on the isolation ward' from a relational perspective. The students described their difficulties and anxieties, as well as their willingness to take care of isolated patients, resulting in the overcoming of obstacles and in contacting and becoming involved when taking care of these patients. PMID- 10520093 TI - The 'greying' of the United Kingdom nursing workforce: implications for employment policy and practice. AB - One in five nurses on the United Kingdom (UK) professional register is aged 50 years or older. Over the next few years, the profession will lose, through retirement, many of its most experienced practitioners. The significance for policy makers and for employers of this age-shift is two-fold. Firstly it is clear that greater numbers of nurses and midwives are reaching, or soon will reach, potential retirement age. Secondly many more nurses are now reaching their middle years and they are likely to have different requirements and attitudes to nursing work. This paper examines the employment policy and practice of the ageing of the UK nursing population. The paper examines data from official sources, and information from attitudinal surveys and case studies with employing organizations to assess the major effects of the ageing of the nursing workforce. Key findings are that the age profile of those nurses working in the National Health Service appears to be 'younger' than that of the total population, with the age profile of nurses working in nursing homes and as practice nurses being older than that of the NHS nursing workforce. However, the overall age profile of NHS nurses masks considerable variation between specialties and trusts, and the 'pool' of potential nurse returners from which the NHS and other employers attempts to recruit, is declining in numbers, as it too ages. Other major issues requiring policy attention are the provision of appropriate flexible hours to older nurses who have caring responsibilities, improving access to continuing professional development, and reducing pension provision inflexibility. PMID- 10520094 TI - Health professionals' perspectives on service delivery in two Northern Ireland communities. AB - This research builds on the findings of an ethnographic study of health inequalities in two small, rural communities in Northern Ireland. Through further analysis of existing data, this second study aimed to explore health professionals' perspectives on issues of service delivery relevant to government policy on primary care. Anthropological fieldwork was conducted for two consecutive 4-month periods during 1995 and 1996 in one predominantly Catholic and one predominantly Protestant town. To preserve confidentiality, the locations have been given the pseudonyms, respectively, of Ballymacross and Hunterstown. Research tools included fieldwork journals and a fieldwork diary, meetings with key informants, tape-recorded interviews, group discussions, participant observation and use of secondary material such as census data, local newspapers and community health profiles. Interviews with 15 health workers revealed that there was not a coherent approach to achieving health gain, little collaborative enterprise and minimal interaction between the different professional groups. The National Health Service (NHS)-employed primary care professionals, more than local community workers, appeared to be demoralized, exhausted and suspicious of the business-orientated health service. In this respect, the primary care-led NHS appeared not to be working. It is concluded that a shared health agenda should be developed by NHS-employed primary care professionals and local community workers to create a health-inducing environment at community level. This needs to be complemented by the establishment of formal mechanisms for inter-agency working at local, professional and government levels. PMID- 10520095 TI - Methodological issues in the development and use of instruments to assess patient nutritional status or the level of risk of nutritional compromise. AB - This paper presents the results of an appraisal of the evidence for the effectiveness of methods of nutritional assessment currently in use by nurses and from this develops concepts which may be applied to many areas of nursing practice. The paper first shows how the approach to nutritional assessment by nurses has changed over time, producing the present search for methods which are relatively quick and easy to use. It then describes the limited nature of the evidence of effectiveness of these methods and considers the reasons for this situation. It suggests that the difficulty of validation is an important factor and describes the three most common approaches to validation, showing how problems arise from lack of clarity in the definition of terms and the assumption of a simple relationship between the level of risk of nutritional compromise and actual nutritional status. In conclusion, it is suggested that these difficulties illustrate principles applicable to many areas of nursing care, and a definition of 'segments' of care processes, each with well-defined purposes and outcome measures, is proposed. Such an approach will help to demonstrate the complexity and value of nursing activity. PMID- 10520096 TI - Physical touch in nursing studies: a literature review. AB - In nursing contexts a distinction is made between two types of touch: physical touch and therapeutic touch. Physical touch may be experienced as therapeutic, but that is not its explicit purpose in the same way as with therapeutic touch. Most of the touch studies reviewed in this article are from the United States of America, Canada and the United Kingdom and thus represent the culture of modern western society. The area covered by these studies is far from coherent, and even the results are to some extent contradictory. It follows that it is difficult to draw any firm conclusions from this review of the concepts, methods and main results of touch studies. PMID- 10520097 TI - Blood gas analysis: a study of blood loss in intensive care. AB - This paper describes a quantitative study conducted on an intensive care unit in the north of England. It involved the collection of data from the existing records of 65 patients consecutively sampled from a predetermined date provided that they stayed more than 24 hours and had an arterial line in situ. As patient records were used, ethical approval was not necessary. The objectives of the study were to quantify the mean number of blood gas samples taken per patient and estimate the mean blood loss resulting from this, including discard volume. Limitations include reliance on records and lack of an economic evaluation. The results show that blood loss in this study was greater than that reported elsewhere. Patients who were ventilated for 24 hours or more had a statistically significant greater blood loss when compared to those who were not (P < 0.001). A subgroup of patients undergoing renal replacement therapy had the greatest blood loss (mean 55.18 ml per day). This loss was statistically significant when compared to patients not in acute renal failure (P=0.007). When patients undergoing multiple therapies normally associated with increased sampling were compared to patients not receiving such therapies, there was no statistically significant difference in blood loss. The need to change current nursing practice to reduce iatrogenic anaemia is emphasized. PMID- 10520098 TI - Vital signs and nurses' choices of titrated dosages of intravenous morphine for relieving pain following cardiac surgery. AB - Postoperative pain, if unrelieved, will impede patients' recovery. A theoretical model of factors which influence nurses' choices of titrated dosages of intravenous (IV) morphine was constructed for this study. This study aimed to examine whether or not a patient's vital signs would influence nurses' choices of titrated dosages of IV morphine for relieving pain following cardiac surgery. A survey design with a vignette and questionnaire method was used to collect data. The vignette developed by McCaffery & Ferrell was modified and adapted for this study. It described the pain reports and vital signs of two patients on postoperative day 1 following cardiac surgery. Convenience sampling was used to seek voluntary participation from 29 registered nurses working in the cardio thoracic intensive care unit of a private hospital in Sydney, Australia. A protocol of the unit allowed nurses to titrate IV morphine against the pain of patients following cardiac surgery. The results showed that the pain assessment of the two patients in the vignette documented by the nurses were not consistent. The titrated dosages (a bolus dosage and a maintenance dosage) chosen by the nurses for the patient with slightly elevated vital signs differed significantly from the titrated dosages chosen by the same group of nurses for another patient with vital signs at the lower end of the stable range (t=3. 33, d.f.=25, P < 0.01 for a bolus dosage and t=3.73, d.f.=25, P < 0. 01 for a maintenance dosage). Different risk factors were stated by the nurses in titrating the bolus and maintenance dosages of IV morphine. The importance of accepting patients' verbal reports of pain as well as the provision of optimal dosages of IV morphine for pain relief is highlighted. A disadvantage of using a vignette and questions method is that the patients' clinical status is somewhat unreal. Further studies, however, were also recommended. PMID- 10520099 TI - Patients' evaluation of pain and nurses' management of analgesics after surgery. The effect of a study day on the subject of pain for nurses working at the thorax surgery department. AB - The effect of a study day on the subject of pain for nurses working at the thorax surgery department The aims of this investigation were: to describe patients' evaluation of pain and the treatment of pain after thorax surgery via sternotomy; to repeat the evaluation with another group of patients following a study day for nurses, featuring pain and pain treatment; and to examine whether the study day influenced the nurses in their treatment of pain. The investigation included daily evaluation of pain using a visual analogue scale (VAS), and an interview with the patients before discharge, where they were asked to review their experience of pain and its treatment. The nurses on the thorax surgery ward and on the intensive care unit (ICU) completed a questionnaire before and after the study day. Finally, a retrospective study of the case notes of the patients taking part was carried out. The results of the investigation showed a low assessment of pain by most patients during the daily evaluation. Asked to recall their pain when interviewed, the rating was higher. A small group of patients had more evident pain than others. When administering opiates the ICU nurses often chose a lower dose than the standing order prescribed. After the study day the nurses gave larger doses of intravenous opioids and the patients experienced less pain. PMID- 10520100 TI - Improving patients' postoperative sleep: a randomized control study comparing subcutaneous with intravenous patient-controlled analgesia. AB - One hundred female patients undergoing major reconstructive plastic or gynaecological surgery were randomized to either receive subcutaneous patient controlled analgesia (PCA) (bolus dose 2.5 mg diamorphine in 1 ml with a 20 minute lockout) or intravenous PCA (bolus dose 0.5 mg diamorphine in 1 ml with a 5-minute lockout). Data were collected by questionnaire and interview to evaluate the intervention on pain scores, quality of sleep on the first postoperative night, postoperative nausea and vomiting (PONV) and overall patient acceptability. The subcutaneous PCA group experienced less 'worse pain' (P < 0.01) and less sleep disturbance due to pain (P < 0.001). Subcutaneous PCA would appear to offer patients a safe and effective means of analgesia and may offer significant advantages over the intravenous route of administration. PMID- 10520101 TI - An embedded decisional model of stress and coping: implications for exploring treatment decision making by women with breast cancer. AB - Treatment decision making by women with breast cancer has been recognized to be an inherently stressful process. However, past decisional theory and research has failed to fully elucidate the personal, transactional and relational nature of choice behaviour. The purpose of this paper is to explore an embedded decisional model of stress and coping that locates key assumptions of Janis and Mann's conflict-theory model (CTM) of decision making within Lazarus and Folkman's transactional framework. Through combining decisional and stress and coping theories, a model is developed that addresses the theoretical limitations of the CTM and provides greater specificity within decision-making research. The paper examines the complexity of treatment decision making within the context of the constructs of causal antecedents, primary appraisal, secondary appraisal, coping and adaptational outcomes. Examples specific to women with breast cancer are provided to illustrate the potential application of the embedded model. The implications of this inclusive and comprehensive decisional theory for future knowledge development and research in the area of treatment decision making are also discussed. PMID- 10520102 TI - Psychiatric nursing in prison: the state of the art? AB - Psychiatric nursing in prisons has received criticism from within and outside the profession in recent years. In England and Wales this amongst other issues has prompted a review of forensic health care by the United Kingdom Central Council for Nursing, Midwifery & Health Visiting (UKCC). The status of forensic psychiatric nursing as a specialty has also been disputed in the literature and the role of nurses working in this field is seen by some to be more about social control than caring. These arguments are set in the present situation of increasing numbers of mentally ill individuals in the prison system and a crisis in the availability of beds in secure units, a situation that is paralleled throughout the western world. The standard expected of health care services in prisons is equivalence with the service the public receives from the National Health Service (NHS). The number of prisoners needing transfer to hospital has increased during the last decade, resulting in competition for a limited number of suitable beds. The effect on health care centres (HCCs) in English prisons is that they now must provide long-term care for seriously mentally ill prisoners. This paper outlines the development of British psychiatric services in prisons. It then describes the HCC in Her Majesty's prison (HMP) Belmarsh and reports on recent radical changes in the management structure of this service. The aim of these changes is to produce a clinical environment in which psychiatric nurses can deliver high quality care in an area beset with difficulties for clinicians and managers and to further progress towards the goal of equivalence. These advances have been achieved through a shift of emphasis in management structure that increases the number of clinical posts and minimizes administrative and security-based responsibilities held by clinical grades. We conclude that although external contracts are necessary, much can be achieved through internal review and changes in policy. PMID- 10520103 TI - A national survey of practice nurse involvement in mental health interventions. AB - BACKGROUND: The move in the United Kingdom (UK) from institutional to community care has led to an inevitable increase in the involvement of practice nurses (PNs) in mental health care. Around 20 000 PNs are currently working in the United Kingdom (UK). However, the extent and nature of PN involvement in delivering mental health interventions has not been adequately explored. AIM: This study aimed to quantify practice nurses' involvement in delivering mental health interventions in primary care settings. METHOD: A questionnaire was sent to a random sample of 1500 practice nurses registered with the Practice Nurse Forum at the Royal College of Nursing. Sixty per cent of questionnaires were returned; however, once non-eligible respondents were removed an adjusted response rate of 54% was achieved. RESULTS: Practice nurses play a significant role in the assessment and treatment of mental health problems, most frequently via the administration of depot antipsychotics and the screening for depression. However, antipsychotic side-effects were infrequently monitored and PNs' understanding of treatment issues in depression is poor. These findings may be associated with the reported lack of mental health training that PNs have received. CONCLUSIONS: The findings of this study have important implications for the training of practice nurses in mental health, specifically in the areas of medication management and the detection of mental disorders. PMID- 10520104 TI - Training for transformation: reorientating primary health care nurses for the provision of mental health care in South Africa. AB - Using programme research, this paper reports on the evaluation of a programme designed to orientate primary health care nurses towards the provision of a comprehensive approach to care. In addition to training in psychiatric care, this was deemed necessary in order to facilitate comprehensive integrated primary mental health care in South Africa. Nurse-patient consultations were evaluated on indicators of comprehensive care before and after the programme. Interviews were also conducted with the participants individually and in a group. The results indicate that there are several factors which mediate the provision of comprehensive care by primary health care nurses. These include individual factors as well as contextual factors, inter alia, the structure and organization of the health care system, which historically has been organized to promote biomedical care. Furthermore, biomedicine has dominated training models in South Africa, instilling in nurses a biomedical approach to patient care. PMID- 10520105 TI - Women's perceptions of phenomena they label premenstrual tension: normal experiences reflecting ordinary behaviour. AB - Remarkably little empirical knowledge exists about premenstrual tension (PMT), a construct that has been studied intensively during the last 30 years. Practically nothing is known about the perceptions women have about PMT. The purpose of this study is to describe the perceptions healthy women have about PMT. Seventeen women, who had 4 years earlier participated in a concurrent diary study assessing the prevalence of PMT, were interviewed. The interviews were analysed by means of a qualitative content analysis. Four main categories describing the women's perceptions were formed: (1) individual experiences of phenomena referred to as PMT; (2) phenomena referred to as PMT reflect ordinary experiences in healthy women; (3) biopsychosocial explaining of phenomena referred to as PMT; (4) internal and external resources used to manage the variability of phenomena referred to as PMT. The content of these categories reflects the participants' perceptions of PMT as the common, normal and very variable experiences which women handle by the use of management strategies commonly used in contemporary society to deal with life in general. Experiences of women differ with respect to what they experience and the timing and persistence of those experiences. This the participants explain from a biopsychosocial perspective. It is suggested that women have adapted a medical term, PMT, to describe what they consider normal female phenomena. PMID- 10520106 TI - A discussion of the legal aspects of female genital mutilation. AB - The purpose of this paper is to examine the position of the nurse/midwife in the United Kingdom when involved with the care of a woman or female child who has suffered genital mutilation, which is an illegal practice in this country and most other areas of the world. The types of circumcision commonly practised are introduced, the prevalent reasons for the continuation of the practice among certain ethnic groups are presented, and the range of issues to be considered by the nurse is examined. These include international and national legal aspects which do not exist in isolation and are considered in context with cultural, medical, human rights and gender issues. Nursing legal issues include child protection, consent, advocacy and confidentiality, which invoke the Code Of Professional Conduct of the United Kingdom Central Council for Nursing Midwifery & Health Visiting. Midwives and nurses working in the field of gynaecology have raised questions regarding possible courses of action to take when presented with this issue. Increased knowledge can help to inform those decisions. Therefore, implications for future practice are addressed, together with recommendations to assist nurses with decision making when faced with this scenario in the future. PMID- 10520107 TI - Violence in mental health care: the experiences of mental health nurses and psychiatrists. AB - Violence against mental health service personnel is a serious workplace problem and one that appears to be increasing. This study aimed to ascertain the extent and nature of violence against mental health nurses and psychiatrists, and to identify what support, if any, they received following exposure to violence. Mental health staff working within five West Midlands Trusts in the United Kingdom were surveyed using a postal questionnaire to investigate the extent and nature of violence they encountered in their daily work. There was an overall response rate of 47%, which included a response rate for psychiatrists of 60% (n=74) and for mental health nurses of 45% (n=301). Though both groups experienced violence at work, nurses were found: to have been exposed to violence significantly more during their career; to have been a victim of violence within the previous 12 months of the survey; and to have suffered a violent incident involving physical contact. Whilst a higher proportion of nurses than psychiatrists received some support following a violent incident, a large proportion of both groups did not receive any, although most felt in need of it. The implications of this study for training and management are discussed. PMID- 10520108 TI - Trends in ophthalmology services, nursing skill-mix and education: 2nd national survey. AB - This paper reports on a replication study of the first national survey commissioned by the RCN Ophthalmic Nursing Forum to assess the changing skill-mix of staff caring for ophthalmic patients and the nature of ophthalmic services. After minor modifications to the original questionnaire, it was circulated to the total population of ophthalmic units and hospitals in the United Kingdom (n=181). Following quantitative data analysis, it was found that the numbers of nurses holding ophthalmic nursing qualifications has risen to almost 50%, although this did not reach statistical significance. This increase has occurred alongside a significant rise in numbers of institutions offering ophthalmic nurse education and the expansion of day case surgery. Regional variations exist, however, to the extent that the increase in ophthalmic nurses and provision of ophthalmic nurse education has not been reflected nearly as much in Scotland as the rest of the UK. Overall, it was also found that the proportion of C, G and I grade nurses has fallen significantly, whilst the numbers of nurses graded D and F has risen. The implications of the results are discussed and recommendations for future research made. PMID- 10520109 TI - The nurse's role in drug handling within municipal health and medical care. AB - In Sweden, elderly care has been transferred from the county councils to the municipalities, resulting in changes in the work tasks of nurses. Today, nurses within municipal health and medical care carry sole responsibility for large groups of care receivers, especially during evenings and weekends. A consequence of this heavy work load is that the nurses have to delegate a number of tasks to subordinate staff. Drug administration is a task which is often delegated. Cases of malpractice involving drug administration, reported from the municipal health and medical service to the regional supervisory units of the National Board of Health and Welfare during a 3-year period, have been analysed in order to gain a deeper understanding of the underlying causes of malpractice. In addition, interviews were conducted with eight nurses, the objective being to find out how they rate their knowledge within this area. The analysis showed that the majority of errors occurred in nursing homes and that the most common errors were administration of the wrong drug, and levels of drugs administered exceeding the prescribed ones. These errors were also common among subordinate staff who had been delegated responsibility for drug administration. Of the nurses interviewed, six out of eight had been close to making errors or had noticed a colleague making an error in conjunction with drug handling. Not all nurses reported errors made to the physician in charge immediately on discovery. It is concluded that through the transformation of elderly care, nurses working within municipal health and medical care have been given a new professional role. This places new demands on their competence, requiring them to make independent judgements and to take their own initiatives. PMID- 10520110 TI - An evaluative study of clinical supervision based on Proctor's three function interactive model. AB - This paper reports on a study of the benefits reported from participation in clinical supervision by registered nurses (n=201) working in a large English community and mental health NHS Trust. It summarizes the emergent practical and theoretical 'key ingredients' of clinical supervision in the United Kingdom and argues that these provide a basis for generalizable research practice. The study is based on Proctor's three function interactive model previously commended as a guide for supervisory practice and evaluation. Within this study, the three functions underpin instrument design but are also a primary focus of evaluation. The development of this instrument through semi- structured interviews with supervisors and supervisees is described. The study aims to assess and compare reported benefits in each of the three functions of accountability, skill development and support in order to examine the effects of contract use, length of experience of clinical supervision and length of service as a registered nurse on reported benefits by using non-parametric statistical analysis. The results indicate that reported benefits are experienced in almost equal proportion across each of these three functions. Statistical analysis indicates a significant positive correlation between experience of clinical supervision and its reported benefits. An inverse correlation is reported between length of service and overall benefits; however, no similar reduction over time against normative benefits was found. There was no relationship between contract use and reported benefits. Limitations of the study are discussed with reference to bias, interview transcription and overlap between the three functions. The paper concludes that nurses report clear benefits from clinical supervision in each of the three functions. This validates the three function interactive model and demonstrates that clinical supervision is used to critically examine and change nursing practice. The content and usage of contracts is identified as an aspect that merits further study. PMID- 10520111 TI - Implementing new health visiting services through action research: an analysis of process. AB - An action research project based in a single fundholding practice on the South Coast of England aimed to identify needs relevant to families with resident children, then to use the contracting system to redirect health visiting practice to meet those needs. The naivety of this plan was well recognised, so the processes that occurred during implementation of the proposed changes were recorded throughout. An analysis of these participant observation data revealed various organisational constraints and facilitators that arose during contract negotiations. Two new full time health visiting posts were established, each with an innovative and somewhat controversial focus. One health visitor was employed to establish a community development project in an underserved area of the town, while the other was to provide a home visiting service for the families of school aged children. The processes involved in structuring decisions about 'health needs' and how these are best met are analysed for each of the two new posts. The analysis reveals powerful influences that affect the implementation of new health visiting services. PMID- 10520112 TI - Practice development: ambiguity in research and practice. AB - Practice development activity occupies an ambiguous position in relation to both clinical practice and research. In practice, it is seen at times as an added extra to normal work despite arguably being an inherent part of professional practice. In research, it fails to demonstrate the rigour of being generalisable because of its explicit location in a specific care environment. The study reported in this paper explored the implications of this ambiguity for practitioners who seek to develop health care practice. Ten focus groups were held with health care researchers and practitioners concerning the processes of developing practice in the North East of England. The results demonstrate how people manage themselves and the uncertainty that surrounds the use of research in clinical practice. The paper argues for an appreciation of reflexive forms of research, such as action and practitioner research, which do not disassociate research and practice and in which practitioners have a role in knowledge creation as well as knowledge implementation. PMID- 10520113 TI - A hermeneutic textual analysis of suffering and caring in the peri-operative context. AB - Hermeneutic text interpretation is discussed in this paper as a possible way of releasing nurses' knowledge, in order to develop a deeper understanding of professional caring. The text is a nurse's story, which has been gathered by means of the critical incident technique. The intention is to elicit the knowledge of professional caring in the stories and the language of caring science, which is concealed in a nurse's reality. Hermeneutic text interpretation through reading a text and having a dialogue with the nurse's reality will give a voice and a language to silent knowledge. PMID- 10520114 TI - Representing nursing judgements in the electronic health record. AB - The naming of nursing phenomena and representing the phenomena in a standardized manner suitable for encoding in computer-based systems is a challenge for the nursing profession at the national and the international level. Considerable progress has been made in the development of classification systems for nursing practice. The focus of this article is on language systems developed to represent nursing judgements in computer-based systems, in particular the electronic health record. A review of two current systems and their proposed revisions (North American Nursing Diagnosis Association, NANDA, Taxonomies I and II, and the International Classification for Nursing Practice, ICNP, Alpha and Beta versions), according to the features suggested by the Computer-based Patient Record Institute (CPRI) for classification systems appropriate for implementation in computer-based systems, suggests that the evolving versions extend the current versions in terms of sufficient granularity (depth and level of detail) and atomic and compositional character. However, it is not clear from the literature available to date whether the characteristics that are most closely related to definition of a formal terminology (i.e. clear and non-redundant representation of concepts, syntax and grammar for logical constructions of compositional terms, synonyms and language independence) will be part of the evolving vocabularies. Formal terminology models and related tools have the potential to complement, extend, and refine existing nursing classification systems. PMID- 10520115 TI - Nursing research conference organised by the Royal College of Nursing of the United Kingdom Research Society, held at Keele University, Keele, England, 23-25 April 1999. PMID- 10520117 TI - Media reviews PMID- 10520118 TI - Media reviews PMID- 10520116 TI - Media reviews PMID- 10520120 TI - Media reviews PMID- 10520119 TI - Media reviews PMID- 10520121 TI - Media reviews PMID- 10520122 TI - Forthcoming contents of volume 30, number 5, november 1999 PMID- 10520123 TI - Oestrogen receptors in the brainstem of the female sheep: relationship to noradrenergic cells and cells projecting to the medial preoptic area. AB - Oestrogen regulates the secretion of gonadotropin releasing hormone (GnRH) and this could be mediated by noradrenergic systems originating in the brainstem. Whilst it is known that noradrenergic cells possess oestrogen receptors (ER), it is not known whether ER-immunoreactive (-ir) cells in the brainstem project to the regions of the hypothalamus in which GnRH neurons are found. We have used dual-label immunocytochemistry to determine the extent to which ER-alpha is found in noradrenergic cells in the brainstem of the ovariectomized (OVX) ewe. Noradrenergic/adrenergic cells were identified by immunostaining for dopamine beta-hydroxylase (DBH). Cells that stained for both DBH and ER were found in both the A1 and A2 cell groups, with the highest levels found in the most caudal regions. In the A1 group, at the most caudal extent, 73% of ER-ir cells were DBH positive and 19% of DBH-ir cells were ER-positive. The degree of co-localization decreased in a linear manner towards the rostral brainstem. In the caudal half of A2, 9-14% of ER-ir cells were DBH-positive and 20-25% of DBH cells were ER positive. Less than 2% of DBH-ir cells in the A5 group were dual-labelled and none of the cells in the A6 and A7 groups were ER-positive. The retrograde tracer FluoroGold was injected into the preoptic area of nine OVX ewes and labelled cells were examined in the brainstem to determine the extent of co-localization of ER. Only injections in the rostroventral part of the medial preoptic area near to the organum vasculosum of the lamina terminalis resulted in the labelling of cells in the brainstem. One ewe with very strong labelling of the brainstem was selected for detailed mapping. In the ventrolateral medulla, half the ER-ir cells in the most caudal regions were retrogradely labelled. Almost all the ER-ir cells in the mid-region of the ventrolateral medulla were retrogradely labelled but no co-localization of retrograde tracer and ER was observed rostral to obex. There were many ER-ir cells and retrogradely-labelled cells in the nucleus of the solitary tract but only a few double-labelled cells. Similarly, numerous ER-ir cells and retrogradely labelled cells were observed around the lateral edges of the caudal fourth ventricle and across to the lateral parabrachial nucleus but there were few double-labelled cells. These results suggest differential regulation of noradrenergic cells by oestrogen, with a direct action of the hormone confined to the cells in the most caudal region of the A1 and A2 cell groups. The cells of the caudal ventrolateral medulla which contain ER-ir cells that project to the preoptic area may be important in the mediation by noradrenaline of the actions of oestrogen on GnRH secretion in the ewe. PMID- 10520124 TI - Altered pituitary sensitivity to corticotropin-releasing factor and arginine vasopressin participates in the stress hyporesponsiveness of lactation in the rat. AB - The regulation of the activity of the hypothalamic-pituitary-adrenal (HPA) axis is modified during lactation, wherein a blunted stress-induced adrenocorticotropic hormone (ACTH) and glucocorticoid secretion is coupled with elevated basal secretion of these hormones. The involvement of pituitary modifications in lactation-induced stress hyporesponsiveness has yet to be established. In this study we tested the hypothesis that the pituitary sensitivity to corticotropin-releasing factor (CRF) and arginine vasopressin (AVP) is altered in lactation in the rat. We examined the effect of exogenous CRF (0.1-5 microg/rat), AVP (0.01-0.5 microg/rat), and AVP (0.01-0.5 microg/rat)+CRF (0.1 microg/rat) on the ACTH response of virgin, mid-lactating (lactation day 10 12) females, as well as nursing females separated from their pups for 48 h. Additionally, to determine if changes in CRF- or AVP-receptor densities might mediate alterations in pituitary sensitivity, we compared pituitary CRF- and AVP receptor binding by autoradiography in pregnant, mid-lactating, and virgin female rats. While both virgin and lactating female rats exhibited significant ACTH responses to CRF, the responses to the highest doses of CRF (2.0 and 5.0 microg/rat) were greater in virgin than in lactating females. Separation of the litter for 48 h partially restored pituitary responsiveness to 2.0 microg of CRF. Conversely, whereas lactating females displayed robust ACTH secretion following a high dose of AVP or following a combination of AVP+CRF, the response of virgin females was much smaller. These modifications in pituitary responsiveness were not accompanied by significant differences in pituitary CRF-and AVP-receptors levels between female groups. Our results demonstrate that a reduction in pituitary sensitivity to CRF, but not to AVP occur during lactation in the rat which mediates, at least in part, the stress hyporesponsiveness of lactation. PMID- 10520125 TI - Prior exposure To oxytocin mimics the effects Of social contact and facilitates sexual behaviour In females. AB - The purpose of this study was to determine whether pretreatment with oxytocin could mimic the effects of social contact and enhance sexual receptivity in female prairie voles. Female prairie voles require prolonged exposure to males to become sexually active and oxytocin has been shown to play a major role in the establishment of social bonds between males and females. Therefore, we hypothesized that prior exposure to exogenous oxytocin, in the absence of males, would enhance sexual activity in females. Two experiments were conducted to test this hypothesis. Experiment 1 examined the capacity of oxytocin to enhance sexual behaviour in females undergoing natural oestrus. Sexually naive female prairie voles received a daily subcutaneous injection of 20 microg oxytocin or isotonic saline for 5 days before being placed with a sexually experienced male for 48 h. Females treated with oxytocin were significantly more likely to mate during this period than saline-treated females. In experiment 2 the ability of oxytocin to increase subsequent sensitivity of sexually naive females to oestradiol was tested. Females that received oxytocin pretreatment, as in experiment 1, followed by oestradiol displayed a significant increase in sexual receptivity when compared to females treated with saline and oestradiol or oestradiol only. The results supported the hypothesis that prior exposure to oxytocin can mimic the effects of social contact, and can facilitate sexual receptivity by increasing the sensitivity of females to very low doses of oestradiol. PMID- 10520126 TI - Effects of sexual interactions with a male on fos-like immunoreactivity in the female quail brain. AB - Sexual interactions can cause changes in plasma hormone levels and activate immediate early genes within the mammalian brain. There are marked anatomical differences between the regions activated that relate directly to the sexual specific behaviour and neuroendocrinology of each sex. The aim of this study was to determine if such a sexual dimorphism exists in birds by examining the brain regions stimulated in adult virgin female Japanese quail (Coturnix japonica) during sexual behaviour and comparing this to previously reported data concerning males. Female quail were allowed to freely interact with adult males and both female and male sexual behaviour was recorded. Contrary to previous findings in male quail, no significant induction of Fos-like immunoreactive (FLI) cells was observed following sexual interactions in the preoptic area of females; this area is fundamentally involved in the control of male-type copulatory behaviour. Sexual interactions significantly induced FLI cells in the hyperstriatum ventrale, the part of the archistriatum just lateral to the anterior commissure, and the nucleus intercollicularis. Moreover, prominent activation was detected throughout most of the ventromedial nucleus of the hypothalamus, a region reported to be rich in oestrogen receptors. FLI induction was not a consequence of sexual behaviour induced changes in luteinizing hormone (LH) as plasma LH levels were unaltered. Instead, brain activation must be a consequence of copulation-associated somatosensory inputs or direct stimuli originating from the male. Male quail, like the majority of other birds, lack an intromittant organ (penis) so that the somatosensory inputs to the female are rather different from those in mammals; the precise nature of these inputs is yet to be determined. PMID- 10520128 TI - The effect of GHRP-6 on the intracellular Na+ concentration of rat pituitary cells in primary culture. AB - The objective of the present study was to further investigate the ionic mechanism of the action of GHRP-6 on male rat pituitary cells in culture. A synthetic hexapeptide, GHRP-6 stimulates the secretion of growth hormone both in vivo and in vitro. It is generally accepted that Ca2+ and protein kinase C but not cAMP are involved in the signal transduction pathway of the action of GHRP-6. Ca2+ influx through voltage-gated Ca2+ channels and mobilization of internal stored Ca2+ are thought to be responsible for an increase in cytosolic Ca2+ concentration. For activation of the voltage-gated Ca2+ channels, however, it is not determined whether the membrane Na+ permeability plays a role. To answer this question, we measured intracellular Na+ concentration of the pituitary cells with ion imaging technique. We found that GHRP-6 increased [Na+]i; the Na+ response depended on the presence of extracellular Na+ and was blocked by Gd3+, known as a blocker of nonselective cation channels but not by tetrodotoxin, a blocker of the voltage-gated Na+ channel; thapsigargin, an inhibitor of endoplasmic reticulum Ca2+ ATPase, had no effect on the response; Ca2+ chelating agent, BAPTA had no inhibitory effect on the response; ouabain, an inhibitor of Na+-K+ ATPase, did not block the rise in [Na+]i induced by GHRP-6; somatostatin, which hyperpolarizes the cells by activating K+ channels, suppressed the response. These data clearly showed that GHRP-6 increased [Na+]i in the rat pituitary cells including somatotrophs. The rise in [Na+]i is likely to be due to an increase in the membrane Na+ permeability which should depolarize the cells, thereby activating the voltage-gated Ca2+ channels. This process leads to an influx of Ca2+ and subsequent increase in [Ca2+]i which results in an exocytotic release of GH. PMID- 10520127 TI - Immunoreactive neurotensin in gonadotrophs and thyrotrophs is regulated by sex steroid hormones in the female rat. AB - In addition to regulating anterior pituitary function by being released from the median eminence, mammalian neurotensin (NT) may also exert an autocrine or a paracrine action within the anterior pituitary. In this study, using double immunostaining with elution restaining, we identified the specific anterior pituitary cells which express NT immunoreactivity (NT-IR) during the rat oestrous cycle. In the normal cycling rat, NT-IR was present in both gonadotrophs and thyrotrophs and displayed plastic changes along the oestrous cycle. Both the number of TSH-NT positive cells and the intensity of immunological reaction were elevated during dioestrus, and decreased through pro-oestrus and early oestrus. NT-IR was also high in both follicle stimulating hormone (FSH)- or luteinizing hormone (LH)-positive cells during early pro-oestrus, and decreased during late pro-oestrus. Treatment of intact rats with either the anti-oestrogens Tamoxifen or LY117018, or the anti-progestagen RU486 prevented the normal expression of NT IR in thyroid-stimulating hormone (TSH)-, FSH-, and LH-positive cells during pro oestrus. Bilateral ovariectomy induced a dramatic reduction in the number of NT IR cells. This effect was completely prevented by treatment of ovariectomized rats with oestradiol and progesterone, and was unaffected by the concurrent administration of a GnRH antagonist. Furthermore, administration of an anti oestrogen together with an anti-progestagen to ovariectomized-oestrogen, progesterone-treated rats, blocked the stimulatory effect of ovarian hormones on NT-IR in anterior pituitary cells. These findings demonstrate that, in female rats, NT is specifically localized in gonadotrophs or thyrotrophs. In addition, they strongly suggest that changes in circulating concentrations of ovarian steroids may control both NT synthesis in, and release from, these cells. PMID- 10520129 TI - Interactive effects of tamoxifen and oestrogen upon the nigrostriatal dopaminergic system: long-term treatments. AB - In the present report adult female rats were ovariectomized (OVX) and assigned to one of four treatment conditions. Treatments consisted of administering pellets containing 17beta-oestradiol (E), tamoxifen (TMX), a combination of TMX and E or no further treatment (OVX). Animals received these treatments immediately following OVX and were maintained in these conditions for a 40-day period. Subsequently, the corpus striatum (CS) was dissected from each animal and prepared for determinations of basal and amphetamine stimulated DA output using in-vitro superfusion. No statistically significant differences among the four treatment groups were obtained for basal dopamine output. The highest levels of amphetamine-stimulated dopamine responses were obtained from E treated rats. These values were significantly greater than that obtained from OVX rats and rats treated with a combination of TMX+E. The significance of these findings is that they indicate both a non-traditional central nervous system site and mechanism of action through which tamoxifen-oestrogen interactions can function. Such data may have important implications for administration of tamoxifen to premenopausal women as this anti-oestrogen may compromise nigrostriatal dopaminergic function under conditions where oestrogenic modulation is present. PMID- 10520130 TI - Evidence for expression of galanin receptor Gal-R1 mRNA in certain gonadotropin releasing hormone neurones of the rostral preoptic area. AB - Previous studies have shown that galanin plays an important role in the regulation of gonadotropin releasing hormone (GnRH) release. At present, it is not known if this role is exerted by direct or indirect interactions between galanin producing neurones and GnRH neurones. The objective of this study was to determine whether or not GnRH neurones could express galanin receptor Gal-R1 mRNA. Dual in-situ hybridization experiments were carried out with digoxigenin labelled cRNA probes encoding GnRH in combination with 35S-labelled riboprobe encoding the galanin receptor Gal-R1. In order to detect possible variations in the expression of the Gal-R1 mRNA under different physiological conditions, male rats, intact female rats throughout the phases of the oestrous cycle, ovariectomized (OVX) and steroid-treated rats were analysed. The results show that many cells expressing Gal-R1 mRNA were present throughout the preoptic area. Gal-R1 mRNA-expressing cells were observed in very close proximity with GnRH neurones. In the female rat, some GnRH neurones located in the rostral preoptic area/vascular organ of the lamina terminalis expressed Gal-R1 mRNA. These double labelled cells were observed at all times of the oestrous cycle, except during diestrus at 08.00 h and pro-oestrus at 18.00 h. Conspicuously, at oestrus 1800 h, we found that 21.6% of rostral GnRH neurones expressed the Gal-R1 mRNA. In addition, dual-labelled GnRH neurones were seen in OVX animals but not in oestrogen plus progesterone-treated ones. In the male rat, colocalization of GnRH mRNA and Gal-R1 receptor mRNA was not observed. In the medial preoptic area, no double-labelled GnRH neurones were detected regardless of the endocrine conditions. These results suggest that, in addition to a possible indirect action of galanin on GnRH cells via neurones located at close proximity, the effects of galanin on GnRH can be mediated by direct activation of galanin receptors in rostral GnRH neurones. This study also shows that expression of Gal-R1 mRNA in GnRH cells is influenced by the levels of circulating gonadal steroids. PMID- 10520131 TI - A study of the gonadotropin releasing hormone neuronal network in the median eminence of the rhesus monkey ( Macaca mulatta) using a post-embedding immunolabelling procedure. AB - The purpose of this study was to describe the ultrastructural features of gonadotropin releasing hormone (GnRH) axonal processes in the median eminence of the monkey, using a post-embedding immunogold labelling procedure. Evidence was also sought to evaluate the view that release of this peptide may be governed by direct inputs to GnRH axons in the median eminence. Plastic embedding was used to preserve ultrastructure, and a polyclonal rabbit anti-GnRH was used as primary antibody. Immunogold labelling with 15-nm particles was almost exclusively found overlying dense core vesicles (dcvs) and preabsorption of the primary antibody with synthetic GnRH eliminated this labelling. Morphometric analysis was performed on tissue from two monkeys. Four types of profiles containing GnRH immunoactive dcvs were observed. Type I profiles were morphologically unremarkable with a cross sectional area of approximately 0.6 microm2 and probably represent intervaricose axon segments. Type II profiles, which were nominally larger than Type I structures, were characterized by a high density of round microvesicles, which were frequently concentrated along the neuronal membrane to form 'synaptoid' contacts with adjacent glia. Two additional and large GnRH profiles (>5 microm2) were observed. One (Type III) contained a high density of dcvs and mitochondria, and was considered analogous to an axonal swelling or Herring body in the magnocellular hypothalamo-neurohypophysial system. The Type IV structure, which was considered not to be a Herring body because of the relative low density of mitochondria was innervated by a classical symmetrical synapse. The functional significance of these observations is discussed. PMID- 10520132 TI - Kappa-opioid receptor activation inhibits post-spike depolarizing after potentials in rat supraoptic nucleus neurones in vitro. AB - Endogenous agonists acting at kappa-opioid receptors modulate the discharge activity of hypothalamic supraoptic nucleus vasopressin cells in vivo. Phasic activity in vasopressin cells is known to depend critically on intrinsic mechanisms involving post-spike depolarizing after-potentials and we hypothesized that inhibition of phasic bursting by an endogenous kappa-agonist may result from reducing the magnitude of depolarizing after-potentials. To investigate this possibility, intracellular sharp electrode recordings were obtained from supraoptic nucleus cells impaled in superfused explants of rat hypothalamus. Bath application of the selective kappa-agonist, U50,488H (0.1-1 microM), decreased the spontaneous firing rate of magnocellular neurosecretory cells (by 94. 0+/ 4.5% at 1 microM, mean+/-SEM; P = 0.02, n = 4). U50,488H did not alter membrane potential (0.9+/-0.8 mV hyperpolarization at 1 microM, P = 0.17, n = 8) or input resistance (11.0+/-4.5% increase at 1 microM, P = 0.09, n = 5). U50,488H (0.1 and 1 microM, both n = 5) reduced depolarizing after-potential amplitude (by 29.9+/ 9.3 and 78.0+/-10. 6%, respectively, P<0.001) in eight cells in which the baseline membrane potential was kept constant by dc-current injection and in which a depolarizing after-potential was evoked every 25-40 s by a brief (40-80 ms) train of 3-6 action potentials (the number of spikes in the trains was kept constant for each cell). Thus, kappa-opioid receptor activation reduces depolarizing after-potential amplitude in supraoptic nucleus cells and this may underlie the reduction in burst duration of vasopressin cells caused by an endogenous kappa-agonist in vivo. PMID- 10520133 TI - Progestins' rapid facilitation of lordosis when applied to the ventral tegmentum corresponds to efficacy at enhancing GABA(A)receptor activity. AB - Progestins may have actions in the midbrain though gamma-aminobutyric acid (GABA)A/benzodiazepine receptor complexes (GBRs) that are relevant for sexual receptivity. The efficacy and time course of various progestins to enhance lordosis when applied to the ventral tegmental area (VTA), following progesterone to the ventral medial hypothalamus (VMH) was investigated. Ovariectomized, oestrogen-primed rats and hamsters with contralateral VMH/VTA cannulae were tested for lordosis before and after implants of P to the VMH and progestins to the VTA. The progestins were P, 5alpha-pregnan-3,20-dione (DHP), 5alpha-pregnan 3alpha-ol-20-one(3alpha,5alpha-TH P), 5alpha-pregnan-3alpha,21-diol-20-one (THDOC), 5beta-pregnan-3alpha-ol-20-one(3alpha,5beta-THP) , 17alpha-ol-6-methyl 4,6-pregnadiene-3,20-dione-17-acetate (megestrol acetate, MA), and 6-chloro-17-ol 4,6-pregnadiene-3, 20-dione-17-acetate (chlormadinone acetate, CA). Progestins' effects on GABA-mediated chloride influx and SR 95531 binding in cortical and midbrain tissue, respectively, were examined in rats and hamsters. 3alpha,5alpha THP and THDOC implants to the VTA were the most effective at immediately facilitating lordosis of rats and hamsters. Two hours later all other progestins, except MA and CA, increased lordosis in rats; only P, 3alpha,5alpha-THP, and THDOC were effective in hamsters. The progestins' effectiveness at facilitating lordosis were similar to their effects on GABA-stimulated chloride influx and SR 95531 receptor binding (3alpha,5alpha-THP and THDOC>P>DHP>3alpha, 5beta-THP>MA and CA). These findings suggest that progesterone lordosis enhancing effects in the rodent VTA may be via GBRs. PMID- 10520134 TI - Mating stimuli influence endogenous variations in the neurosteroids 3alpha,5alpha THP and 3alpha-Diol. AB - Progesterone facilitates and 5alpha-dihydrotestosterone (DHT) inhibits female sexual behaviour in rodents; their metabolites, 5alpha-pregnan-3alpha-ol-20-one (3alpha,5alpha-THP) and 5alpha-androstane-3alpha-17beta-Diol (3alpha-Diol), may influence the onset and termination of lordosis. Changes in these and related steroids in hormonal states associated with differences in receptivity were investigated. Rats were assigned to oestrus, metoestrus, dioestrus, pro-oestrus, mated, gestational days 5-7, 12-14, 18-20, or post-partum conditions; rats 9+ months of age were considered older. Pro-oestrus rats were exposed to the mating arena, sight and smell of a male with no tactile contact, artificial vaginocervical stimulation, standard mating, or no mating. Progesterone, 5alpha pregnane-3,20-dione, 3alpha,5alpha-THP, oestradiol, testosterone, DHT, 3alpha Diol, dehydroepiandrosterone, and corticosterone were measured in plasma and whole brain, midbrain, hypothalamus, cortex, amygdala, hippocampus. 3alpha,5alpha THP and 3alpha-Diol changed with reproductive state and mating stimuli. Plasma and whole brain 3alpha,5alpha-THP and 3alpha-Diol were significantly increased in pro-oestrus versus dioestrus rats; plasma 3alpha,5alpha-THP was decreased and 3alpha-Diol increased in mated versus pro-oestrus rats. The midbrain and hypothalamus had the most evident changes in 3alpha,5alpha-THP and 3alpha-Diol between dioestrus versus pro-oestrus and pro-oestrus versus mated rats. 3alpha,5alpha-THP and 3alpha-Diol were altered differently in response to mating stimuli. 3alpha,5alpha-THP was greater in the midbrain of mated versus pro oestrus rats; other mating-relevant stimuli decreased 3alpha,5alpha-THP. Midbrain 3alpha-Diol was increased with exposure to a male /=5 days. Most patients with DH did not perceive the condition as severe and did not seek treatment (75.1%). Only 23.3% used a desensitizing dentifrice. The results indicated that self-reporting of DH is lower than reported in a dental hospital population and was not perceived as a major dental problem by most patients in a general dental practice population. PMID- 10520146 TI - Adjustment of dental occlusion in treatment of chronic cervicobrachial pain and headache. AB - The hypothesis that the response to conventional physical therapy of patients suffering from chronic cervicobrachial pain and/or headache can be improved by adjusting dental occlusion, was tested. Forty patients seeking treatment were interviewed and examined prior to treatment, and 6 weeks, 12 months and 60 months after treatment. All patients underwent routine physical therapies. They were pairwise matched for age, gender and type of dental occlusion, and randomly allocated to a true occlusal adjustment group or to a mock adjustment group. The patients and the examiners were unaware of the type of dental treatment given. The outcome variables included subjective pain and discomfort, cervical spine mobility and pain on movement, and comparison of relative EMG activities. The short-term response to therapy was good in both groups. In the long-term, however, the response was significantly better in the patients who had undergone occlusal adjustment than in the mock-adjusted controls. PMID- 10520147 TI - In vitro permeability and scanning electron microscopy study of acid-etched and ground enamel surfaces protected with dental adhesive coating. AB - Clinical procedures, such as acid etching and reshaping of the teeth supporting removable partial dentures by grinding off some enamel surface, increase the permeability of dental enamel. Teeth take several months in vivo to partially recover from such damage. In the meantime, the tooth is more susceptible to carious decay. To prevent this the ground or etched enamel should be effectively protected. Using electron paramagnetic resonance (EPR) and a two-chamber diffusion cell the authors studied the influence of adhesive resin applied to the ground and acid-etched enamel surfaces on the diffusion of spin label TMAPO (2,2,6-6 tetramethyl-4-acetamido-piperidine-1-oxyl) molecules through the enamel. The enamel permeability was measured in samples exposed to 1-min etching with 37% phosphoric acid, in samples etched for 5 min, and in samples ground with a diamond bur. Next, all the treated enamel surfaces were coated with Scotchbond Multi-Purpose Plus(R) dental adhesive system and the permeability measurements repeated. Scanning electron microscopy (SEM) was used to study the porosity of enamel surfaces. The adhesive resin film covering the etched or ground enamel surfaces was found to decrease significantly the diffusion through dental enamel. This finding confirms the clinical value of dental adhesives used to protect ground or accidentally acid-etched enamel surfaces. SEM analysis showed that adhesive resin covers the porous surface of the acid-etched and ground enamel tightly. PMID- 10520148 TI - Quantitative analysis of masseter and temporalis EMGs: a comparison of anterior guided versus balanced occlusal concepts in patients wearing complete dentures. AB - The lack of easily measurable, objective physiological activity parameters of the masseter and temporalis muscle during jaw movements in humans has led to the consideration to revise data of surface electromyographies (EMGs) by applying a computerized quantification method. The aim of this follow-up analysis was to get quantitative data out of EMG-records of an earlier study. These records were obtained with two different splints, splint 1 providing an anterior front-canine guidance and splint 2 providing bilateral balanced occlusion. Utilizing a computer aided integration method led to numeric results which statistically proves the prediction of the previous investigation. Applying the integration method, the EMG raw signal was transformed into area-values which enabled a statistical work up of the data. Wilcoxon test statistics shows a significant (P<0.05) lower muscle activity in patients wearing dentures providing anterior front-canine guidance compared to those with balanced occlusion. It is concluded that the neuromuscular activity of the elevator muscles is highly reproducible and that the neuromuscular function is similar in edentulous subjects to that found in people with natural teeth. Furthermore, the study statistically proves earlier visual data that all those subjects, whose muscle activities were observed with anterior guidance (splint 1) compared to bilateral balanced occlusion (splint 2) showed significantly lower values with regard to subjects wearing splint 2. PMID- 10520149 TI - Histological features of artificial secondary caries adjacent to amalgam restorations. AB - Artificially induced carious lesions on either side of in vitro un/restored amalgam cavities were examined to establish the degree of randomness of caries development. Class I cavities were cut in 132 extracted premolars: twelve teeth were not restored, and 120 teeth were restored with one of 20 different restoration combinations of silver amalgam, base and varnish. After ageing for periods of 3 months and 1 year, the 12 unrestored teeth and 80 of the restored teeth were subjected to an in vitro bacterial challenge for 36 days, while the other 40 specimens were challenged in acidified broth (pH 4.0). Sections were then prepared for polarized light microscopy. Carious lesion configuration on either side of the cavity was noted, and outer, wall and dentine lesions measured. Data underwent Fisher's exact test, a chi-squared goodness of fit test and a Student's paired t-test with P<0.05. Except for dentine lesions, acid broth and unrestored specimens showed carious lesions having similar size and occurrence on either side of the cavity. Two unrestored cavities showed caries resistance. Restored, bacterially challenged specimens were significantly different regarding total and wall lesion distribution and wall lesion width and area on either side of the cut cavity. Acid broth challenge will promote regular caries development at the tooth-restoration interface. The random caries pattern which developed in restored bacterially challenged specimens indicates that the tooth-restoration interface forms a diverse environment providing sites of varying susceptibility to caries. PMID- 10520150 TI - The effect of accelerated ageing on the mechanical properties of soft denture lining materials. AB - The purpose of this study was to determine the effect of simulated ageing on the physical properties of some soft denture liners. The hardness, tensile strength and percentage elongation values were determined before and after ageing treatment. At the end of the experimental ageing process softening was observed only in Simpa and Ufigel L materials. It was established that ageing had insignificant effect on the hardness of other materials. It can be presumed that there will not be significant time-related changes in the hardness of these materials. Molloplast B preserved its highest value after the ageing process, whilst that of Simpa decreased to become comparable with Ufigel P. PMID- 10520151 TI - The long-term effect of oromaxillofacial trauma on the function of the temporomandibular joint. AB - Thirty patients with temporomandibular disorders (TMD) after trauma to the orofacial region were followed for 6 months. They were assessed at 10 days, 1 month, and 6 months. It was found that patients with combined fractures of the mandible and condyle showed more effects on function of the temporomandibular joint after 1 month than patients with condylar fracture only. This difference was not so apparent after 6 months. Patients with trauma but no fracture showed similar effects to those with non-condylar fractures. There was a tendency for arthrogenous patients to develop myogenous problems with time. A group of patients with TMD but no trauma, showed more favourable long-term response to conservative treatment. Long-term follow-up of these patients for TMD problems is recommended. PMID- 10520152 TI - Dental asymmetry in temporomandibular disorders. AB - In our previous study, it was reported that facial asymmetry due to mandibular lateral displacement (MLD) was significant in patients with temporomandibular disorders (TMD). In this study, dental asymmetry in TMD patients was investigated by means of PA cephalogram and study model. Lateral deviation of the midline of the mandibular occlusal table (PA-mid) and right-left difference of the molar relationship (Molar-Diff.) were examined, and their relationship with MLD was studied. PA-mid and Molar-Diff. were significantly correlated with MLD. In most cases, displaced side of the midline of the lower occlusal table was coincident with that of the mandibular skeletal midline. These results suggest that in TMD patients asymmetries in occlusal relationship of the midline of the mandibular teeth and molars were mainly due to a mandibular skeletal asymmetry and not merely due to a dental malposition on the alveolar basal bone. Many cases had a more distal occlusal relationship of the first molar on mandibular displaced side compared with that on the opposite side. A high incidence of Class II relationship was found (61.8% as a whole) and more remarkable on the mandibular displaced side. Midline discrepancy and right-left difference of the molar relationship seem to be important occlusal characteristics in patients with TMD. PMID- 10520153 TI - Temporomandibular disorders in saudi females seeking orthodontic treatment. AB - The aim of this study was to describe the prevalence of signs and symptoms of temporomandibular disorders (TMD) in a group of patients seeking orthodontic treatment. One hundred and ninety one consecutive prospective orthodontic female patients, divided into three age groups of 8, 14 and 18 years, were examined for TMD signs and symptoms and orthodontic treatment need (IOTN). The percentages of signs and symptoms found were 41 and 30%, respectively. No significant association was found between IOTN and TMD signs and symptoms. The youngest age group reported significantly less headache and temporomandibular joint (TMJ) noise. Headache was significantly associated with all TMD symptoms and with tenderness to palpation. In conclusion, the results indicate that malocclusion could not be considered as a primary aetiologic factor for TMD within the age range studied. PMID- 10520154 TI - Malassezia pachydermatis: a review. AB - Malassezia pachydermatis is of importance in both veterinary and human medicine. Its taxonomic status and physiological characteristics are now better understood. Skin disease associated with this lipophilic yeast is now commonly recognized, especially in dogs. However, further studies are required to elucidate the mechanisms which allow this yeast to proliferate and induce disease. Skin colonization is common in pet carnivores which consequently constitute a source of M. pachydermatis for susceptible humans. In the future, the development of efficient typing systems should allow better understanding of the transmission mechanisms. PMID- 10520155 TI - Oxidative stress sensitivity and superoxide dismutase of a wild-type parent strain and a respiratory mutant of Candida albicans. AB - It is important to know responses of the pathogenic fungi to reactive oxygen species by which hosts protect themselves against fungal infection. In the present study, sensitivities to the superoxide radical (O2-) and superoxide dismutase (SOD) were compared between a wild-type parent strain and a respiration deficient mutant of Candida albicans. When their survival was examined on an agar medium containing an intracellular O2- generator, paraquat (PQ), the parent strain was selectively killed by increasing the PQ concentration. In contrast, when cells of both strains were illuminated in a riboflavin solution, they exhibited similar sensitivity to O2- generated extracellularly by photo-reduced riboflavin. There were no large differences in sensitivity to hydrogen peroxide in the two strains. Thus, the high tolerance of the mutant to PQ was suggested to result from low intracellular O2- generation by PQ due to the respiratory deficiency. It is generally accepted that fungal cells contain manganese (Mn)-SOD in the mitochondria and copper and zinc (CuZn)-SOD in the cytoplasm. Cyanide insensitive SOD activity (attributable to Mn-SOD) was dominant in the parent strain throughout growth phases, whereas cyanide-sensitive activity (attributable to CuZn-SOD) occurred in the mutant. The activity bands of Mn- and CuZn-SODs were clearly separated by electrophoresis of the cell extracts of both strains on non denaturing polyacrylamide gels. The electrophoretic profiles obtained were consistent with the results of the activity assay. These results showed that the respiratory deficiency affected oxidative stress sensitivity and SOD in C. albicans. PMID- 10520156 TI - Molecular taxonomy of Trichophyton mentagrophytes and T. tonsurans. AB - Most members of the anamorph genus Trichophyton are anthropophilic and have evolved with the human host. Classical parameters for the identification of dermatophytes include clinical features, cultural characteristics, conidial morphology and physiological test results. Phenotypic variability and pleomorphism due to culturing on artificial media is common among this group of organisms and has led to the description of numerous species. The validity of taxa around T. mentagrophytes and T. tonsurans was verified. Morphological and physiological features were compared to results of three different molecular techniques (sequencing of the internal transcribed spacer (ITS) region of the ribosomal operon, PCR fingerprinting and amplified fragment length polymorphism (AFLP) analysis). Twenty-four species or varieties investigated could be reduced to five taxa and were reclassified or synonymized as Trichophyton tonsurans, T. interdigitale, T. mentagrophytes, T. simii and T. erinacei. PMID- 10520157 TI - Asymptomatic carriage of Cryptococcus neoformans in the nasal cavity of the koala (Phascolarctos cinereus). AB - Over a 22-month period, sequential nasal and skin swabs were obtained from 52 healthy captive koalas (Phascolarctos cinereus) from the Sydney region. Cryptococcus neoformans was isolated in 17 koalas from 64 of 262 (24%) nasal swabs and from nine of 262 (3%) skin swabs. Prevalence of nasal colonization varied seasonally from 12% (3/25) to 38% (10/26). Cryptococcus neoformans var. gattii alone was cultured from 37, var. neoformans alone from 22 and both varieties from five nasal swabs. Of 33 koalas sampled on three or more occasions, organisms were isolated persistently from six, occasionally from eight and never from 19. Two koalas were persistently and heavily (>/=100 colonies/plate) colonized by C. neoformans var. gattii and two with var. neoformans. Isolation of C. neoformans var. gattii from the skin was low grade and sporadic. No koalas from which C. neoformans was persistently isolated showed clinical signs of cryptococcosis and all except one had a negative latex cryptococcal antigen test, therefore the nasal cavity was presumed to be colonized by, rather than infected with, C. neoformans. Preliminary observations of koalas from Coffs Harbour indicated a much higher prevalence of colonization by C. neoformans, suggesting that environmental factors influenced the extent of carriage by C. neoformans. PMID- 10520158 TI - Antifungal activity of cerebrospinal fluid against Cryptococcus neoformans and Candida species. AB - The effect of human cerebrospinal fluid (CSF) on the growth of Cryptococcus neoformans and Candida species was tested in RPMI-1640. CSF alone was highly fungistatic for both yeasts and inhibited growth in a concentration-dependent manner. Unlike human serum, CSF did not collaborate with fluconazole for killing C. neoformans. Molecular sieve fractionation of CSF on a G-200 Sephadex column yielded a highly antifungal fraction with a molecular weight around 66 kDa. On SDS-PAGE this fraction migrated as a major and a minor band corresponding to the mobility of bovine serum albumin. These novel findings suggest that CSF contains a factor(s) that provides resistance to the growth of C. neoformans or Candida species. PMID- 10520159 TI - Effect of dental treatment and/or local application of amphotericin B to carious teeth on oral colonization by Candida. AB - Microbiological analyses of saliva and swabs were obtained from carious lesions of 54 children and adolescents with carious teeth, and of 49 boys and girls with healthy teeth. Candida species were isolated from the saliva of 36 (66.7%) subjects with active caries, but from the saliva of only one (2%) of the 49 caries-free subjects. Candida was detected in material removed from the carious lesion in 44 (81.5%) of the children with caries. Thirty patients with carious teeth and colonized by Candida were randomly divided into three groups of 10 individuals each, and either treated by complete dental restoration, by local application of amphotericin B or by a combination of dental treatment plus amphotericin B. The final microbiological control showed that thorough dental treatment alone eliminated fungi from the oral cavity in 90% of cases, whereas the local application of amphotericin B alone had a minimal effect on the candidal colonization of carious lesions. When, in addition to dental treatment, amphotericin B was applied, fungi were completely eliminated from the oral cavity of all subjects. PMID- 10520160 TI - Inhibition of sterol 14 alpha-demethylation of Candida albicans with NND-502, a novel optically active imidazole antimycotic agent. AB - To investigate the mode of action of the newly synthesized optically active imidazole compound, NND-502, (-)-(E)-[4-(2, 4-dichlorophenyl)-1,3-dithiolan-2 ylidene]-1-imidazolylacetonit rile, its effect on ergosterol biosynthesis in cell free extracts of Candida albicans was examined and compared with that of the (S) enantiomer of NND-502 in addition to lanoconazole and bifonazole, both of which are clinically used for the treatment of dermatomycoses. NND-502 was found to interfere with ergosterol biosynthesis by inhibition of sterol 14alpha demethylase, while no interference due to the (S)-enantiomer of NND-502 was found, indicating that the stereochemical orientation of the 2, 4-dichlorophenyl group plays an important role in the interaction with the enzyme. In terms of drug concentration exerting 50% inhibition of ergosterol biosynthesis, NND-502 was 2.5 and 28 times more effective than that of lanoconazole and bifonazole, respectively. PMID- 10520161 TI - Identification and cloning of GCA1, a gene that encodes a cell surface glucoamylase from Candida albicans. AB - Adherence of yeast cells of Candida albicans to human oesophageal cells is greater when cells are grown in 500 mM D-galactose in comparison to D-glucose at the same concentration. Moreover, a 190 kDa mannoprotein (MP190) from a yeast cell wall preparation is highly expressed when cells are grown in the presence of galactose but less so in glucose. We now report on the identification of the MP190 and the isolation of its encoding gene. MP190 was purified, and three internal peptides were isolated and sequenced. Each of the three peptides showed significant homology (65-85%) with a glucoamylase (GAM1) from the yeast, Schwanniomyces occidentalis. In order to isolate the C. albicans homologue of GAM1 (GCA1), we probed a genomic library with a 0.9-kb internal fragment of the S. occidentalis GAM1 and isolated a 2.3-kb clone that corresponded to the 5' region of the gene. Polymerase chain reaction (PCR) amplification was used to isolate the remainder of the open reading frame. GCA1 encodes a 946 amino acid protein containing three putative hydrophobic, membrane-spanning domains and 15 potential N-glycosylation sites. Both Gca1p and GAM1 are novel to the family of glycosyl hydrolases. Northern analysis indicated that GCA1 is transcribed to a greater extent in galactose than in sucrose or glucose. Also, using reverse transcriptase (RT)-PCR, we observed expression of GCA1 in a rat model of oral candidiasis, indicating that Gca1p is expressed during disease development. PMID- 10520162 TI - Feral pigeons as carriers of Cryptococcus laurentii, Cryptococcus uniguttulatus and Debaryomyces hansenii. AB - We collected fresh droppings and cloaca samples from feral pigeons Columba livia in the southern Swedish city of Malmo, and isolated the following fungi: Debaryomyces hansenii var. hansenii, Cryptococcus laurentii and Cryptococcus uniguttulatus. The first two species are known to be pathogenic to humans. No strains of Cryptococcus neoformans var. neoformans were found. Our results indicate that feral pigeons can be carriers of medically significant fungi other than Cryptococcus neoformans var. neoformans. PMID- 10520163 TI - Comments on airborne Aspergillus and incidence of invasive aspergillosis. PMID- 10520164 TI - Development and plasticity of interstitial cells of Cajal. AB - Interstitial cells of Cajal (ICC) are the pacemakers in gastrointestinal (GI) muscles, and these cells also mediate or transduce inputs from the enteric nervous system. Different classes of ICC are involved in pacemaking and neurotransmission. ICC express specific ionic conductances that make them unique in their ability to generate and propagate slow waves in GI muscles or transduce neural inputs. Much of what we know about the function of ICC comes from developmental studies that were made possible by the discoveries that ICC express c-kit and proper development of ICC depends upon signalling via the Kit receptor pathway. Manipulating Kit signalling with reagents to block the receptor or downstream signalling pathways or by using mutant mice in which Kit or its ligand, stem cell factor, are defective has allowed novel studies into the specific functions of the different classes of ICC in several regions of the GI tract. Kit is also a surface antigen that can be used to conveniently label ICC in GI muscles. Immunohistochemical studies using Kit antibodies have expanded our knowledge about the ICC phenotype, the structure of ICC networks, the interactions of ICC with other cells in the gut wall, and the loss of ICC in some clinical disorders. Preparations made devoid of ICC have also allowed analysis of the consequences of losing specific classes of ICC on GI motility. This review describes recent advances in our knowledge about the development and plasticity of ICC and how developmental studies have contributed to our understanding of the functions of ICC. We have reviewed the clinical literature and discussed how loss or defects in ICC affect GI motor function. PMID- 10520165 TI - Temporal and spatial rhythmicity of jejunal wall motion in rats. AB - Isolated segments of jejunum of fasted rats exhibit regular rhythmic contractions at the same frequency as slow-waves. The aim of the present study was to search for a possible spatial rhythmicity of this activity. Using a video imaging technique, jejunal segments of 50 rats were studied. Only experiments (n=76) with no propagated contractions at visual inspection were included in the study. After the measurement, a spectral analysis of the diameter variations was performed. The bands were characterized by four parameters: level, main frequency, amplitude and phase. At each level, the phase varied, suggesting that the same rhythmic phenomenon occurred, but with a delay as a function of the spatial position. In 58 measurements, the rhythmic activity had a frequency near 0.50 Hz and in 18, near 0.25 Hz. Phase difference was found in 32 segments (42%). The variation with distance was linear as a function of time and its length was greater for the low frequency group than for the high-frequency group (25.6 +/- 9.4 vs. 33.3 +/- 5.2 mm, P=0.015). By contrast, the speed of propagation was not significantly different. The wavelength lambda of the spatial rhythmicity was 27.7 +/- 23.2 and 9.8 +/- 4.2 mm (P=NS) in the high- and low-frequency groups, respectively. This corresponds to a speed of propagation of v=lambda*f, where f is the frequency of the wall motion (7.0 +/- 5.2 vs. 5.2 +/- 2.2 mm sec-1, P=NS). PMID- 10520166 TI - Progressive alterations in circular smooth muscle contractility in TNBS-induced colitis in rats. AB - The present study was designed to investigate inflammation-induced changes in smooth muscle responses to acetylcholine and the tachykinins that may contribute to the abnormal motility associated with inflammatory bowel disease. Colitis was induced in male Sprague-Dawley rats by intrarectal administration of trinitrobenzenesulphonic acid in ethanol. After either 4 h (acute) or 7 days (chronic) the distal colon was taken for in vitro measurement of smooth muscle tension and histological assessment. Acute colitis featured injury and neutrophilic infiltration confined to the mucosa while chronic inflammation showed marked injury, lymphocytic infiltration and muscle thickening. Acute inflammation increased responses to substance P and acetylcholine but decreased responses to neurokinin A. The enhanced response to substance P was dependent on nerves, while the decreased response to neurokinin A reflected a reduction in activity at the level of the smooth muscle. In the saline group, there was evidence of cholinergic interaction with substance P, but not neurokinin A. Substance P modulation of cholinergic nerves was absent in acute inflammation. Responses to all neurotransmitters were decreased in the chronic stage. These data demonstrate progressive changes in the smooth muscle function during acute and chronic colitis that may contribute to the abnormal motility associated with inflammatory bowel disease. PMID- 10520167 TI - Effect of truncal vagotomy on gallbladder bile kinetics in conscious dogs. AB - Previous studies on the effects of vagotomy on gallbladder (GB) motility have yielded conflicting results. The aim of this study was to evaluate the effects of vagotomy on GB motility and bile kinetics using a new method. Twelve dogs were divided into two groups of six (control and pyloroplasty) and, 4 weeks later, underwent truncal vagotomy. A catheter secured in the GB fundus was used to monitor GB volume. After injecting polyethylene glycol (PEG) into the GB, combined measurements of GB volume and PEG concentration enabled GB emptying and bile kinetics to be estimated. Seven and five of the 12 vagotomized dogs were classified as having large and normal fasting GB volumes, respectively. Postprandial GB emptying was impaired when the fasting GB volume was enlarged. In the fasting state, bile kinetics of vagotomized dogs were significantly smaller than the control values. The emptying ability of the GB of vagotomized dogs with large fasting GB volumes was reduced considerably both in the postprandial and the fasting states. Such retention of bile in the GB after vagotomy may facilitate cholesterol crystal nucleation and stone growth. PMID- 10520168 TI - Motility in the Roux-Y limb after distal gastrectomy: relation to the length of the limb and the afferent duodenojejunal segment--an experimental study. AB - Following gastrectomy, the longer is a Roux-Y limb constructed to restore digestive continuity the higher the frequency of postoperative symptoms. The aim of this experimental study was to test how the level of the jejunal transection and the length of the Roux limb affect the motility of the constructed limb and in particular the onset and the propagation of activity fronts (AFs). Three months after a distal Roux-en-Y gastrectomy, electromyographic tracings were recorded in six groups of rats grouped according to the level of the transection (20 or 40 cm from the pylorus) and the length of the limb (10, 20 or 30 cm). Animals in which a simple laparotomy or laparotomy + jejunal transection was performed, served as controls. During the interdigestive period, all animals had AFs in the limb which were independent from those recorded in the duodenum. In the limb, the mean time interval between two AFs was shorter (P < 0.01) and more irregular than in controls. An increase in limb length was associated with a lower incidence of completely propagated AFs (P < 0.05) and a higher incidence of irregularly propagated AFs (P < 0.01). When propagation of the AFs was analysed both in the limb and in the jejunum distal to the anastomosis, propagation abnormalities were more frequent. Below the gastrojejunal anastomosis, for an intestinal length of either 20 or 30 cm, the frequency of abnormal AFs was not different when this length was either only a limb or a limb with the 10 cm of distal jejunum below the jejuno-jejunal anastomosis. Interruption of AFs by a meal was irregular in the limb and more rarely observed in the 30-cm than in 10 cm limbs (P < 0.05). Interruption of AFs was shorter than in controls (P < 0.01). In the duodenum and the jejunum proximal to the limb, the interval between AFs was higher than in controls and in the Roux-Y limbs (P < 0.001). Intraluminal concentrations of bacterial strains were not different in the different types of limb while lactobacillus concentrations and pooled concentrations of bacteria were higher than in controls (P < 0.05). No relationship was found between the incidence of myoelectric abnormalities and intraluminal bacterial concentrations. Increasing the length of a Roux-Y limb resulted in more frequent disturbances in AFs in the limb but had no significant consequence on the overall rate of abnormal AFs in the jejunum distal to the transection. Motor response to food intake was also reduced. Motor changes were not related to intraluminal bacterial concentrations. PMID- 10520169 TI - Control systems of gastrointestinal motility are immature at birth in dogs. AB - Networks of interstitial cells of Cajal (ICC) in the myenteric plexus (Myp) or circular muscle (CM) function as pacemakers for gastrointestinal slow waves. ICC in contact with muscle and closely associated with nerves in the CM may mediate inhibitory neurotransmission. We wondered if ICC in Myp and CM and their connections are immature at birth and mature first in the proximal gut in association with nerves. Tissues from lower esophageal sphincter (LES), pylorus (PYL), small intestine (SI) and colon (CO) of 18 term fetal dogs taken from six females were fixed and prepared for ultrastructural examination and studied. Ganglia were present where expected in the Myp and submucous plexus (SMP). ICC cells were present in the Myp of PYL, SI and CO and appeared to have normal relationships to the outer border of CM as in adults. ICC in CM were found associated with nerves in the LES and in PYL, but not in SI or CO. However, axons in CM were everywhere usually free of glial covering, indicating ongoing migration or development. No organized deep muscular plexus (DMP) in SI or submuscular plexus (SP) in colon was present. Visible gap junctions were absent everywhere except for very rare ones between circular muscle cells. We conclude that at birth the neural and ICC networks of CM are more immature in intestine and colon than in oesophagus and stomach. Development of nerve and ICC of CM in oesophagus and stomach apparently precedes that in the remaining gut. However networks in these regions have not achieved adult organization and ICC and smooth muscle cells are anatomically poorly coupled. These findings suggest the reasons that gut motility at birth will not be adult in pattern are because ICC, nerve and muscle control systems are not fully differentiated. Further developmental delays in ICC and nerve maturation could have serious consequences for feeding of infant animals. PMID- 10520170 TI - Multisite optical recording of excitability in the enteric nervous system. AB - A multisite optical recording technique consisting of an array of 464 photodiodes was used to measure dynamic changes in transmembrane potentials (Vm) of guinea pig and mouse enteric neurones stained with the voltage-sensitive dye Di-8 ANEPPS. Optical recordings of Vm changes in enteric neurones which were evoked by depolarizing current pulses or synaptic activation mirrored the Vm changes measured intracellularly in the same neurone. Action potentials had fractional change in fluorescence of -0.09 +/- 0.06% and their peak to peak noise level was 20 +/- 14% of the action potential amplitude. Optical recordings after electrical stimulation of interganglionic nerve strands revealed slow EPSPs, nicotinergic supra- and subthreshold fast EPSPs as well as propagation of action potentials along interganglionic strands. Local application of acetylcholine onto a single ganglion induced reproducibly and dose dependently action potential discharge demonstrating the feasibility of neuropharmacological studies. The optical mapping made it possible to record action potentials simultaneously in a large number of neurones with high spatiotemporal resolution that is unattainable by conventional techniques. This technique presents a powerful tool to study excitability spread within enteric circuits and to assess differential activation of enteric populations in response to a number of stimuli which modulate neuronal activity directly or through synaptic mechanisms. PMID- 10520171 TI - Possible role of nitric oxide in postoperative ileus: a comparative study. AB - In the present study, the possible involvement of nitric oxide (NO) in the pathogenesis of postoperative ileus was investigated indirectly by measuring nitrate, a stabile metabolite of NO. Plasma levels and 24-h urinary excretion of nitrate and nitrite were determined in the peri-operative period in three different groups of patients undergoing surgery: group 1 (LT, n=11) underwent a laparotomy, group 2 (LS, n=12) underwent a laparoscopic procedure, whereas group 3 underwent an extra-abdominal procedure (EA, n=9). Duration of postoperative ileus was assessed clinically using first occurrence of flatus and defaecation as the end of the period of ileus. Postoperative ileus lasted significantly longer in the LT group (first flatus after 3.0 [3.0-4.0] days) compared with the LS (1.0 [1.0-2.0] days) and EA (1.0 [1.0-3.0] days) groups. Urinary nitrate excretion increased significantly in the LT and EA groups during the first 24 h after surgery (from 797.0 [214.0-810.0] and 551.5 [438.3-1215.8] to 2079.0 [889.0 4644.0] and 1102.5 [315.3-1238. 0] micromol/24 h, median [IQR]), but normalized before the end of postoperative ileus. Plasma levels of nitrate were unchanged after surgery, whereas CRP levels were significantly increased in all groups (LT > LS=EA). In the first 24 h following surgery, urinary nitrate excretion is increased, suggesting increased endogenous synthesis of NO postoperatively. As no correlation was found between urinary nitrate excretion and duration of postoperative ileus, we conclude that assessment of nitrate has no value in predicting clinical outcome after surgery. PMID- 10520172 TI - Receptors for innate pathogen defence in insects are normal activation receptors for specific immune responses in mammals. AB - The Toll genes code for activation receptors for innate microbial defence in insects and have recently been found to code for the Lps receptor in murine B lymphocytes. In this study, I will present evidence to show that this family of receptors are responsible for induction of specific thymus-independent immune responses in mice. T cells do not possess these pattern-recognizing receptors, but they possess structures capable of delivering activation signals to these receptors on B cells. PMID- 10520173 TI - Is there a maternally induced immunological imprinting phase a la Konrad Lorenz? AB - In mammals, IgG antibodies are transferred from mothers to the offspring. Since these maternal antibodies result mainly from thymus-dependent immune responses which have undergone immune maturation through somatic hypermutations, they represent the highest quality of the collective maternal immunological experience. Maternal antibodies not only confer passive immunity as long as the newborn's immune system has not fully developed, but also exert an active stimulation as indicated by their regulatory influence on isotype expression, long-term idiotypic alterations, determination of the adult B and T cell repertoire, induction of antigen reactive IgM as well as an affinity enhancement of a proportion of early primary antibodies. The fact that several of these features can only be induced during limited sensitive periods shortly after birth is reminiscent of the behavioural imprinting as defined by Konrad Lorenz. We therefore propose that during early ontogeny there is an immunological imprinting phase with characteristics analogous to behavioural imprinting: (i) the internal imprinting effect is induced by external signals, (ii) in contrast to normal learning, immunological imprinting is also only possible during certain development phases and (iii) it is characterised by an (almost) irreversible result. Hence, if particular immunological experiences are only possible during such sensitive phases, maternal immunoglobulins and consequently the mother's immunological experience is of prime importance for the start of the ontogenetic development of the immune system. PMID- 10520174 TI - A single-chain fusion molecule consisting of peptide, major histocompatibility gene complex class I heavy chain and beta2-microglobulin can fold partially correctly, but binds peptide inefficiently. AB - The function of major histocompatibility complex class I (MHC-I) molecules is to sample peptides from the intracellular environment and present these peptides to CD8+ cytotoxic T lymphocytes (CTL). We have attempted to develop a general approach to produce large amounts of pure and active recombinant MHC-I molecules. A convenient source of MHC-I molecules would be a valuable tool in structural and biochemical analysis of MHC-I, and in experiments using MHC-I molecules to enable specific manipulations of experimental and physiological CTL responses. Here we describe the generation of a recombinant murine MHC-I molecule, which could be produced in large amounts in bacteria. The recombinant MHC-I protein was expressed as a single molecule (PepSc) consisting of the antigenic peptide linked to the MHC-I heavy chain and further linked to human beta2-microglobulin (hbeta2m). The PepSc molecule was denatured, extracted, purified and folded using a recently developed in vitro reiterative refolding strategy. This led to the formation of soluble, recombinant MHC-I molecules, which migrated as monomers of the expected size when submitted to non-reducing sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). Serological analysis revealed the presence of some, but not all, MHC-I-specific epitopes. Biochemically, PepSc could bind peptide, however, rather ineffectively. We suggest that a partially correctly refolded MHC-I has been obtained. PMID- 10520175 TI - Staphylococcus aureus Cowan strain 1 activation of B-chronic lymphocytic leukaemia cells augments the response to CD40 stimulation. AB - The signals involved in regulating the proliferation, differentiation and survival of B-chronic lymphocytic leukemia (B-CLL) cells are fully understood. B CLL cells have been found to respond poorly to various activation signals and only after successful Epstein-Barr virus (EBV) transformation has it been possible to maintain such cells in long-term cultures. In this work we describe a new method to activate and induce proliferation in B-CLL cells and to maintain such cells in long-term culture for longer than 1 month. We used a combination of protocols in an attempt to mimic some of the signals of a thymus-dependent immune response. The B-CLL cells were first activated with Staphylococcus aureus Cowan strain 1 (SAC) particles plus thioredoxin (Trx), followed by stimulation with interleukin (IL)-2 + Trx. This treatment primed the cells for further stimulation with anti-CD40 monoclonal antibody (MoAb) presented on irradiated CD32L cells (the CD40-system) or soluble CD40 Ligand, and a combination of Trx and cytokines (IL-4 + IL-10), which allowed the cells to be maintained for up to 1 month with preserved viability and a variable rate of proliferation. However, induced proliferation of the B-CLL cells was limited to approximately 1 month, suggesting that additional signals are required to facilitate further proliferation. PMID- 10520176 TI - Quillaja saponin formulations that stimulate proinflammatory cytokines elicit a potent acquired cell-mediated immunity. AB - We examined the ability of various Quillaja saponins in iscom-matrix formulations to induce proinflammatory cytokines, such as interleukin (IL)-1alpha and IL-6, and to stimulate acquired immune responses to influenza virus envelope proteins. The A-fraction of Quillaja saponins (QH-A) was shown to stimulate antigen presenting cells (APC) to produce proinflammatory cytokines, and elicited a high primary antigen-specific antibody response and potent cell-mediated responses, as measured by T-cell proliferation, production of cytokines and cytotoxic T lymphocyte (CTL) activity. The C-fraction of Quillaja saponins (QH-C) was shown to have a low capacity to stimulate proinflammatory cytokines and elicited low primary antibody and T-cell responses. However, the QH-C iscom-matrix mediated a potent booster effect, resulting in a high secondary antibody response. The ability of APC to discriminate and to respond to QH-A formulations more efficiently than to QH-C with release of proinflammatory cytokines, which precedes a potent acquired immune response, identifies an important mechanism through which some adjuvants may exert their immunoenhancing activities. PMID- 10520177 TI - Regulation of effector cell functions through the ligation of lymphocyte function associated antigen-1 and intracellular adhesion molecule-1 leads to spontaneous regression of a rat histiocytoma. AB - Cell adhesion molecules mediate cell-to-cell interactions, thereby regulating effective immune response against target cells. We have investigated the intercellular cross-talk between lymphocyte function-associated antigen-1 (LFA-1) and its counter-receptor, intercellular adhesion molecule-1 (ICAM-1). This interaction regulates the postadhesion events leading to the regression of a rat histiocytoma, AK-5, which is mediated by natural killer (NK) cells through antibody-dependent cell cytotoxicity (ADCC). Repeated systemic administration of anti-cell adhesion molecule (anti-CAM) monoclonal antibodies (MoAbs), i.e. anti LFA-1 and anti-intracellular adhesion molecule-1 (anti-ICAM-1), in AK-5 tumour bearing animals enhanced tumour growth and caused delayed regression. The immune suppression achieved after the administration of these antibodies could be attributed to the diminished cytotoxic, as well as apoptotic, activity of NK cells. The MoAbs selectively reduced the adhesion of NK cells to the tumour cells in vitro. Flow cytometric analysis showed down-regulation of Fas-ligand expression on NK and T cells following MoAb administration in vivo. Marked reduction in the lymphokine-activated killer (LAK) activity of NK cells was also observed after in vitro blocking of LFA-1 and ICAM-1 molecules. In addition, continuous MoAb administration induced suppression of the cytokine response. These results underline the importance of ligand binding between LFA-1 and ICAM 1, which could provide co-stimulation for the effector-cell functions via signalling events to induce cytotoxicity, apoptosis and cytokine production, resulting in the efficient regression of AK-5 tumour in syngeneic hosts. PMID- 10520178 TI - Participation of CD1 molecules in the presentation of bacterial protein antigens in humans. AB - Human CD1 molecules, expressed on the surface of professional antigen-presenting cells (including dendritic cells, Langerhans' cells, B cells and activated monocytes) are structurally homologous to major histocompatibility complex (MHC) class I and class II molecules. CD1b and CD1c have been shown to present nonpeptide bacterial antigens to T cells. We hypothesized that CD1 molecules may also be involved in the presentation of bacterial protein antigens. Human peripheral blood mononuclear cells (PBMC) were exposed to two medically important proteins, tetanus toxoid (TT) and purified protein derivative (PPD), with and without murine monoclonal antibodies (MoAbs) specific for CD1a, CD1b and CD1c. All the MoAbs substantially inhibited the proliferative responses of PBMC to TT and PPD. Simultaneous interaction of CD1 and MHC class II molecules was even more inhibitory to these antigen-specific proliferative responses. In contrast, neither mixed lymphocyte reaction nor superantigen and mitogenic responses were affected by CD1-specific antibodies, indicating a certain restriction pattern in antigen presentation. Our findings suggest that, besides MHC class I and II molecules, there is a family of nonpolymorphic cell surface molecules that is able to present certain bacterial protein antigens to T cells. PMID- 10520179 TI - Tolerance or rejection: A delicate balance as judged by exposure of heart transplanted rats to the immunomodulator Linomide. AB - When applied in rodent transplant models most immunosuppressive drugs yield adequate graft protection for as long as the drug is given, and permanent graft survival is often induced. The immunomodulator, Linomide, previously shown to stimulate T cells and prevent apoptosis, usually reduces or abolishes both tolerance induction and the graft-protective effect of the immunosuppressive drug. By chance, we observed that Linomide alone exerted a modest but unequivocal graft-protective effect in the BN to WF strain combination. This finding was analysed by simple genetic mapping of rat strains. Untreated WF recipients kept BN grafts for a median of 8 days, whereas Linomide treatment prolonged graft survival to 12. 5 days (P = 0.0001). In control groups (DA to LEW, BN to LEW, DA to WF and WF to BN), median graft survival was 5.5-7 days irrespective of whether Linomide was given. However, the BN to F1 (LEW x WF) combination also manifested slightly longer graft survival in the presence of Linomide. F1 (BN x WF) to WF grafts survived a median of 15 days without Linomide and 46 days with Linomide treatment. Both in the presence and absence of Linomide, two of the control graft combinations [F1 (BN x DA) to WF and F1 (BN x WF) to BN] manifested 6-7-day graft survival. Taken together, our results suggest a delicate balance between unresponsiveness and rejection, while a single agent (Linomide) may either cause on its own long-term survival of allografts in one setting or rejection despite optimal immunosuppression in another setting. PMID- 10520180 TI - Inhibitory effect of haptoglobin on granulocyte chemotaxis, phagocytosis and bactericidal activity. AB - Human haptoglobin (Hp) is synthesized at hepatic and extrahepatic sites as an acute-phase reactant protein (APP). We investigated the effects of Hp on granulocyte function. The chemotaxis of freshly isolated human granulocytes and differentiated HL-60 cells in response to the bacterial tripeptide, f-met-leu phe, was inhibited in the presence of a physiological concentration of Hp, but chemotaxis in the presence of the proinflammatory cytokine interleukin-8 (IL-8) was not inhibited. Phagocytosis of viable Escherichia coli, as well as fluoresceinated nonviable E. coli, was inhibited. Hp also reduced granulocyte intracellular bactericidal activity against E. coli. The observed inhibitory effects of Hp on granulocyte function are similar to those reported for C reactive protein and suggest that APPs dampen the acute inflammatory response. PMID- 10520181 TI - Minute defects in the expression of MHC E molecules lead to impaired protection from autoimmunity in NOD mice. AB - The E complex of the major histocompatibility complex (MHC) can prevent the spontaneous development of diabetes in nonobese diabetic (NOD) mice transgenic for the Ea gene. None of three promoter-mutated Ea constructs with Ea expression directed to different subsets of immunocompetent cells exerts full protection in NOD mice. The promoter-mutated constructs are all capable of mediating intrathymic elimination of I-E-restricted T cells. Thus, thymic negative selection is not responsible for the protective effect but a more complex effect is likely. Here we show that combinations of two or three different mutated Ea constructs do not protect against intra-islet insulitis either. We also show that spleen cells from protected animals are sufficient to protect NOD mice in adoptive transfer experiments. The only detectable expression defects in splenic cells or cells influencing the repertoire of splenic cells are in the B-cell compartment. Furthermore, in three construct combinations, the differences to wild-type expression are extremely small. Thus, we conclude that even minute disturbances of the E expression pattern might reduce the protection of NOD mice from insulitis. PMID- 10520182 TI - Antigenic definition of plasma membrane proteins of Bacillus Calmette-Guerin: predominant activation of human T cells by low-molecular-mass integral proteins. AB - Mycobacterial plasma membrane proteins, in particular the detergent-soluble or 'integral' ones, comprise a class of mostly unexplored antigens capable of inducing potent activation of human T cells. Plasma membrane isolated from culture-grown Bacillus Calmette-Guerin (BCG; Indian vaccine; Danish strain) was subjected to a Triton X-114-based biphasic extraction procedure for isolation of peripheral (water-soluble) and integral proteins (PMP and IMP). A distinction between the two protein pools was evident from results of SDS-PAGE and immunoblotting using antisera raised in rabbits. An enzyme-linked immunosorbant assay with a panel of WHO-IMMYC monoclonal antibodies against various mycobacterial antigens revealed that three well-known antigens, 19 kDa, 33/36 kDa (proline rich) and 38 kDa (PstS homologue), were part of the IMP pool; and another such antigen, 14/16 kDa alpha-crystallin homologue, partly constituted the PMP pool. Apparently, antigenically distinct species of the immunomodulatory moiety lipoarabinomannan partitioned in aqueous and detergent phases. Human T cell proliferation assays in donors comprising tuberculoid leprosy and pulmonary tuberculosis patients and healthy BCG vaccinees showed significantly greater potency of IMP over PMP and this immunodominance appeared to be directed towards CD4+ cells. IMP of < 56 kDa were resolved by 'continuous elution SDS-PAGE' into 15 fractions which, after extraction of SDS, were used in T-cell proliferation assays for the identification of immunodominant constituents. Proteins falling within three low-molecular-mass zones (all < 35 kDa) performed better than the rest, particularly a approximately 22 kDa fraction, which strongly stimulated T cells from all five donors. Partial overlap between IMP and secreted proteins, as noticed in this study, could provide clues to immunodominance of the latter. The apparent uniqueness and a high T-cell activating potency make mycobacterial IMP attractive candidates for designing future vaccines or immunotherapeutic agents. PMID- 10520183 TI - High peptide affinity for MHC class I does not correlate with immunodominance. AB - Cytotoxic T (Tc)-cell responses against influenza virus infection in BALB/c (H 2d) mice are dominated by Tc clones reactive to the viral nucleoprotein (NP). Here, we report investigations using recombinant vaccinia viruses (VV) encoding major histocompatibility complex (MHC) class I H-2Kd molecules differing by a single amino acid from glutamine (wild-type, Kdw) to histidine (mutant, Kdm) at position 114 located in the floor of the peptide-binding groove. Influenza infected target cells expressing Kdw were strongly lysed by Kd-restricted Tc cells against A/WSN influenza virus or the immunodominant peptide of viral NP (NPP147-155), whereas infected Kdm-expressing targets gave little or no lysis, respectively, thus showing the immunodominance of NPP147-155. Kdm-expressing target cells saturated with synthetic NPP147-155 (10-5 M) were lysed similarly to Kdw-expressing targets by NPP147-155-specific Tc cells. Thus the defect in influenza-infected Kdm-expressing targets was quantitative; insufficient Kdm peptide complexes were expressed. Tc-cell responses against four other viruses or alloantigens showed no effect of Kdm. When peptide transport-defective cells were infected with VV-Kdw or VV-Kdm and co-infected with a recombinant VV encoding an endoplasmic reticulum-targeted viral peptide, two influenza haemaglutinin peptides caused higher expression of Kdw than NPP147-155 indicating their higher affinity for Kdw. These results are inconsistent with the hypothesis that immunodominance in the anti-influenza response reflects high affinity of the immunodominant peptide, but are consistent with skewing of the Tc-cell receptor repertoire. PMID- 10520184 TI - Lymphocyte stimulation by CD3-CD28 enables detection of low T cell interferon gamma and interleukin-4 production in rheumatoid arthritis. AB - The analysis of cytokine production is increasingly important in defining the course of an immune response and in evaluating specific therapies of immune diseases. In rheumatoid arthritis (RA), a dysregulation in T1/T2 cell balance, as defined by the production of their specific cytokines, IFN-gamma and IL-4, respectively, is suggested. A predominance of T1-cell mediated macrophage activity in the joint plays a key role in the destruction of articular cartilage and subchondral bone, whereas local T2 cell activity, mitigating disease, fails. However, analysis of the cytokines defining both T cell subsets is difficult and spontaneous production is often below detection limits. Several stimuli are therefore used to increase cytokine production. In the present study we examined whether stimulation of peripheral blood T cells in the context of mononuclear cells (PB MNC) by CD3-CD28 is a reliable method for assessing IFN-gamma and IL-4 production and is representative for the spontaneous production of these cytokines. The production of IFN-gamma and IL-4 following CD3-CD28 stimulation of RA PB MNC correlated significantly in a ratio 1 : 1 with production following ionomycin-PMA stimulation. In samples with detectable spontaneous production of IFNgamma and IL-4, production following CD3-CD28 stimulation was significantly higher than in stimulated samples with undetectable spontaneous production. Moreover, in the case of spontaneous production there was a significant positive linear correlation between the CD3-CD28 stimulated and spontaneous IFNgamma and IL-4 production, although production of both cytokines was not equally enhanced. Serial sampling did not show significant daily or weekly variation in stimulated cytokine production. The results demonstrate that a pecific T-cell stimulation by CD3-CD28 is a reliable way to enhance IFN-gamma and IL-4 production above the detection limit and so measure the T1/T2 cell balance in RA. PMID- 10520185 TI - Enhanced expression of integrins and CD66b on peripheral blood neutrophils and eosinophils in patients with rheumatoid arthritis, and the effect of glucocorticoids. AB - The aim of this study was to elucidate signs of granulocyte activation by studying adhesion and phagocytosis receptors on peripheral blood granulocytes from patients with rheumatoid arthritis (RA), and to observe the effect of glucocorticoids. Analyses by flow cytometry showed elevation of the neutrophil and eosinophil expression of the alpha- and beta-chains of the beta2-integrin Mac 1 (CD11b/CD18) and of the CEA-gene family member 6 (CGM6, CD66b). Expression of the adhesion receptor antigens CD11a, CD29, CD49d, CD49f and CD44, and the Fcgamma receptors II and III, was unaffected. Treatment with low-dose prednisolone reduced the expression of CD11b on neutrophils and of CD11b, CD18 and CD66b on eosinophils to the same level as that found in healthy controls. Metyrapone treatment increased the surface expression of CD35 and CD49f on eosinophils, but did not affect surface expression on neutrophils. Activation of blood granulocytes may be important for the increased recruitment of neutrophils and eosinophils to the synovial cavity in RA. Treatment with low doses of glucocorticoids in RA normalizes the enhanced expression of the studied adhesion molecules in eosinophils but has minor impact on neutrophil activation. Endogenous glucocorticoid production seems to have minimal or no effect on the expression of adhesion and phagocytosis receptors on circulating granulocytes. PMID- 10520187 TI - Going international in doctoral education in nursing: the creation of INDEN. International Network of Doctoral Education in Nursing. PMID- 10520186 TI - Serum Gm allotype development during childhood. AB - Gm allotypes are genetic variants of the immunoglobulin heavy G chains (IGHG) of IgG molecules, coded from chromosome 14q32, characterized by differences in amino acid epitopes of the constant heavy G chains and inherited in the Mendelian manner. Gm allotypes have influence on IgG subclass levels, and serum Gm allotype levels have been given for different Gm genotypes in adults. Four hundred and thirty healthy children, aged 1-15 years, were examined for serum Gm allotypes and IgG subclasses from the six most common Gm genotypes and different age groups were measured using competitive enzyme-linked immunosorbant assay and radial immunodiffusion methods. Quantities (in g/l) of G1m(a) and G1m(f) of IgG1, G2m(n) and G2m(-n) of IgG2 and G3m(g), and G3m(b) of IgG3 are given. Different maturation rates of the alternative Gm allotypes within IgG1, IgG2 and IgG3 were shown. G2m(n) development was strikingly retarded compared with G2m(-n) from the gamma2 locus. This was found comparing IgG2 levels from homozygous G2m(-n-n) and G2m(nn) individuals, but was also seen in heterozygous G2m(n-n) genotypes. From the gamma1 locus G1m(f) levels dominated significantly, but inconstantly, over G1m(a) levels in heterozygous G1m(af) individuals. In homozygous G1m genotypes, G1m(aa) compared with G1m(ff) of the same age, one or the other dominated, sometimes significantly. Serum levels of G3m(b) from the gamma3 locus of homozygous G3m(bb) individuals were increased significantly compared with G3m(g) levels of homozygous G3m(gg) individuals, in ages over 3 years. However, in heterozygous G3m(gb) individuals G3m(b) dominance was not evident. There is a relatively rapid development of G1m(f) molecules and a retarded development of G2m(n) in the Gm(f;n;b) haplotype. In comparison, G1m(a) is retarded and G2m(-n) is enhanced in the Gm(a;-n;g) haplotype. The retarded serum G2m(n) development is comparable with serum IgA development during childhood. Different maturation rates of Gm allotypes within the same IgG subclass provide further explanation for the variation of the antibody response during childhood. Quantitative Gm allotype determinations give information of the activity from IGHG genes. The genetic variation constitutes an additional basis for evaluation of IgG antibodies in different diseases in childhood. PMID- 10520188 TI - Maternal perceptions of family-provider relationships and well-being in families of children with Down syndrome. AB - The purpose of this study was twofold: (a) to describe parental perceptions of family-provider relationships, and (b) to explore links between parental perceptions of family-provider relationships and well-being in families with children who have Down syndrome. Mailed questionnaires were used to collect data from 94 families that include a child with Down syndrome. Data from 89 mothers are the focus of this report. The results indicate that when mothers of children with Down syndrome believe that their family's relationship with health care providers is positive and family-centered, they feel more satisfied with the care that their child is receiving and they are more likely to seek help from health care providers. In addition, when a discrepancy exists between what mothers want the family-provider relationship to be and what they believe the relationship is, mothers feel less satisfied with the care that their child is receiving. Finally, higher levels of individual and family well-being are reported by mothers who (a) want, and believe they have, positive family-centered relationships with providers, and (b) feel more satisfied with care received. Results of this study contribute to a better understanding of the role that health care providers play in individual and family adaptation to chronic conditions. PMID- 10520189 TI - Initiation of physical restraint in nursing home residents following restraint reduction efforts. AB - In this pilot study a one group pretest posttest design was employed to identify resident characteristics and environmental factors associated with initiation of physical restraint. Predictors of restraint initiation for older adults were examined using secondary analysis of an existing data set of nursing home residents who were subjected to a federal mandate and significant restraint reduction efforts. Lower cognitive status (OR = 1.5 [for every 7-point decrease in Mini-Mental State Examination], 95% CI = 1.0, 2.1) and a higher ratio of licensed nursing personnel (OR = 3.7, 95% CI = 1.2, 11.9) were predictive of restraint initiation. Key findings suggest that restraint initiation occurs, despite significant restraint reduction efforts, when a nursing home resident is cognitively impaired or when more licensed nursing personnel (predominantly licensed practical nurses) are available for resident care. Achievement of restraint-free care in nursing homes requires specific and individualized approaches for residents who are cognitively impaired, as well as greater attention to staff mix of registered nurses, licensed practical nurses, and nursing aides. PMID- 10520190 TI - Inhibitors of prostaglandin synthesis do not improve food intake or body weight of tumor-bearing rats. AB - Interleukin-1 (IL-1) and tumor necrosis factor (TNF) are immunoregulatory cytokines that mediate many aspects of the acute phase response to infection and injury. It has been hypothesized that these cytokines mediate the onset of the cachexia-anorexia syndrome with tumor growth. The anorexigenic effects of IL-1 are mediated in part by prostaglandins (PG). Therefore, the purpose of the present study was to determine if administration of ibuprofen (ibu) or indomethacin (indo), which inhibit PG synthesis, would affect the food intake and body weight of tumor-bearing rats. Rats were implanted with the Morris 7777 hepatoma, a tumor known to induce anorexia and weight loss in rats, and weight loss and leukocyte synthesis of IL-1 and TNF in mice. Treatment with indo or ibu did not improve food intake or body weight in the tumor-bearing rats. However, administration of ibu coincident with tumor implantation did result in smaller tumor mass compared to placebo-treated controls. The results of the present study suggest that PG synthesis is not a major factor in the onset of anorexia in this animal model of tumor-induced anorexia. However, further studies of the effects of inhibitors of PG synthesis on the kinetics of tumor growth are clearly indicated. PMID- 10520191 TI - The relationship of authority to decision-making behavior: implications for redesign. AB - Redesigning health care environments has occurred in response to cost and quality pressures. Efforts to redesign the nursing practice environment have focused on the structure and process of nursing care delivery. When redesign efforts address the structure of nursing practice systems to facilitate one important process, nurses' participation in decision making, better patient and organizational outcomes are expected. The purpose of this study was to determine if two dimensions of structure: administrative (decentralization) and professional authority (expertise) influence the process of participation in decision making for two kinds of decisions (caregiving and condition-of-work) that nurses make. The stratified sample consisted of 300 registered nurses working on medical surgical units. Administrative and professional authority accounted for a small but significant amount of variation in participation in decision making. Because the extent of explained variation was small, the findings may challenge the prevailing assumption that greater authority for decision making results in the exercise of that authority. Redesign of the practice environment therefore must incorporate multiple factors in achieving greater participation in decision making. PMID- 10520192 TI - Development and psychometric testing of the Breastfeeding Self-Efficacy Scale. AB - Many new mothers discontinue breastfeeding prematurely. Researchers have shown that maternal confidence is an important factor in the continuation of breastfeeding. The purpose of this methodological study was to develop and conduct preliminary psychometric assessment of an instrument to measure confidence in new breastfeeding mothers. Using self-efficacy theory as a conceptual framework, the Breastfeeding Self-Efficacy Scale (BSES) was developed and content validity was judged by a panel of experts and through interviews with experienced breastfeeding mothers. Following a pilot test, the revised BSES was assessed with 130 in-hospital breastfeeding mothers for reliability and validity, including internal consistency, principal components factor analysis, comparison of contrasted groups, and correlations with measures of similar constructs. Support for predictive validity was demonstrated with positive correlations between BSES scores and infant feeding patterns at 6 weeks postpartum. Following further testing, this instrument may be used to identify new mothers with low breastfeeding confidence who are at high risk to prematurely discontinue breastfeeding. PMID- 10520193 TI - A feminist critique of research on menopausal experience of Korean women. AB - Despite the increasing number of studies on the menopausal experience of Asian women, the focus of the studies has been on simple comparisons of their symptoms with Western women's and other disease-oriented research topics. To propose directions for future research on menopause, we analyzed and critiqued 158 studies on the menopausal experience of a group of Asian women-Korean women. The studies were retrieved through a search of computerized databases in the United States and South Korea, and they were reviewed, analyzed, and critiqued with a feminist perspective. Many of the studies have problems with (a) conceptualization, including ethnocentric and androcentric views of menopause, biomedical perspectives, and language difficulties; (b) research methods, such as inadequate instruments, passive relationships between researchers and research participants, culturally inappropriate communication styles, inadequate study designs, and homogeneous research participants; and (c) interpretation and communication of study findings. These issues undermine the conclusions drawn about the nature of the menopausal experience of Korean women. PMID- 10520194 TI - A comparison of traditional approaches to hierarchical linear modeling when analyzing longitudinal data. AB - Longitudinal designs typically involve repeated time-ordered observations for each individual (or unit). Such designs are uniquely suited to studying changes over time within individuals, and relating these to individual characteristics to identify processes and causes of intra- individual changes and interindividual differences in physiologic and psychological development. The purpose of this paper is to compare and contrast univariate and multivariate ANOVA with repeated measures to hierarchical linear modeling as approaches to analyzing such longitudinal data. This will enable researchers to choose the approach that best meets their research needs, and it will enable them to compare research results that are reported using one analytical approach with results that are reported using the other approach. PMID- 10520195 TI - Simultaneous three wavelength imaging with a scanning laser ophthalmoscope. AB - BACKGROUND: Various imaging properties of scanning laser ophthalmoscopes (SLO) such as contrast or depth discrimination, are superior to those of the traditional photographic fundus camera. However, most SLO are monochromatic whereas photographic systems produce colour images, which inherently contain information over a broad wavelength range. METHODS: An SLO system has been modified to allow simultaneous three channel imaging. Laser light sources in the visible and infrared spectrum were concurrently launched into the system. Using different wavelength triads, digital fundus images were acquired at high frame rates. RESULTS: Favourable wavelengths combinations were established and high contrast, true (red, green, blue) or false (red, green, infrared) colour images of the retina were recorded. The monochromatic frames which form the colour image exhibit improved distinctness of different retinal structures such as the nerve fibre layer, the blood vessels, and the choroid. CONCLUSIONS: A multi-channel SLO combines the advantageous imaging properties of a tunable, monochrome SLO with the benefits and convenience of colour ophthalmoscopy. The options to modify parameters such as wavelength, intensity, gain, beam profile, aperture sizes, independently for every channel assign a high degree of versatility to the system. PMID- 10520196 TI - Key adhesion molecules are present on long podia extended by hematopoietic cells. AB - BACKGROUND: We recently reported that CD34(+) hematopoietic cells and the KG1a cell line extend long, thin podia. These podia can dynamically extend and retract, often adhere to the substrate, and appear to connect cells up to 300 microm apart. The surface receptors found on these podia have not been described. METHODS: By using time-lapse fluorescent microscoscopy and immunostaining techniques, we describe a method for detecting surface receptors on these podia. This includes an in situ antibody staining procedure without fixing cells. RESULTS: We demonstrate, using CD34 selected mobilized peripheral blood cells and KG1a cells, that adhesion molecules known to play important roles in blood-cell migration and adhesion are present on these podia. These include: CD11a, CD18, CD29, CD34, CD45, CD49d, CD49e, and CD62L. Additionally, CD54 and CD44 were present on the podia extended by KG1a cells, but were not detectable on the primary CD34(+) cells. The integrin CD49d localized at the base of these podia in a time-dependent manner in KG1a cells. The frequency and morphology of these long podia on three myeloid leukemia-cell lines (KG1a, MV4-11, and AML-193) and a CD34 negative T-cell line (CEM) are also compared. KG1a and CEM cell lines extend long, dynamic podia that are similar to the podia on primary CD34(+) cells in morphology and adhesion molecule expression. The AML-193 and MV4-11 cell lines, however, did not extend these long podia. CONCLUSIONS: We describe a technique that provides a method of detecting surface receptors on thin cell membrane projections. These results support the likely role of these podia in cell migration and cell-cell communication. PMID- 10520197 TI - Human basophils express CD22 without expression of CD19. AB - BACKGROUND: Even modern automatic cell counters cannot count basophils precisely. Therefore, we need a rapid, accurate, precise, and easy method for counting basophils. METHODS: Using flow cytometry, basophils (CD22+/CD19-) and B cells (CD22+/CD19+) were counted. Within a large lymphocyte light scatter gate, % basophils (G%baso) and % B cells (G%B) were determined from the total count. Another method of analysis was to make two regions (R1 for basophils and R2 for B cells) and to determine in those the % basophils (R1%baso) and % B cells (R2%B) without gating. The flow cytometric basophil counts of the blood of 21 normal controls and 43 chronic myelogenous leukemia (CML) patients were compared with manual basophil count (Ma%baso) and basophil count by Coulter electronic cell counter (Hialeah, FL) (Auto%baso). CD22+/CD19- cells were sorted by a FACSCalibur (Becton Dickinson, San Jose, CA). RESULTS: The G%baso of all samples was 4.66 +/- 5.35%, and R1%baso was 4.23 +/- 4.88%, and they were well-correlated (r = 0.996, P < 0.001). The G%B of all samples was 1.55 +/- 1.68%, and R2%B was 1.59 +/- 1.67%, and they were also well-correlated (r = 0.993, P < 0.001). Their correlation was better in normal controls than in CML. G%baso was well-correlated to Ma%baso (r = 0.827) and Auto%baso (r = 0.806), and R1%baso was well-correlated to Ma%baso (r = 0.831) but showed poor correlation to Auto%baso (r = 0.734). Auto%baso revealed the poorest correlation to Ma%baso (r = 0.692). The sorted CD22+/CD19- cells were all basophils (99.48 +/- 0.30%), and they revealed CD13, CD33, and dim CD45 expression, whereas CD3, CD14, CD16, and HLA-DR were not detected on them. CONCLUSIONS: We discovered a specific marker combination to identify basophils (CD22+/CD19-), and we suggest that flow cytometric analysis using these markers is an easy, reliable, and accurate method of basophil counting. PMID- 10520198 TI - Anomalous changes in forward scatter of lymphocytes with loosely packed membranes. AB - BACKGROUND: Forward scatter (FSC) is generally associated with cell size and has been suggested as a way to differentiate apoptotic from viable cells. Among spleen cells cultured for 48 h, a population of cells (population B) was found to have decreased forward and increased side scatter relative to freshly purified cells (population A). Interestingly, population B was not present early in analysis; this report explores the change in FSC of population B. METHODS: Using a Coulter (Hialeah, FL) Epics Elite ESP flow cytometer, changes in forward scatter and lipid packing of spleen cells were measured. RESULTS: Over time, the FSC of unfixed cells in population B increased from that of the debris field, to reach a stable value by 30 sec (population A's FSC remained constant). When fixed, populations A and B exhibited constant FSC. Population B cells displayed altered lipid packing as reported by MC540, and the FSC changes were mimicked by Nonidet P-40 treatment of freshly purified spleen cells. CONCLUSIONS: Data emphasize the importance of delaying measurements on unfixed cells until FSC readings have stabilized, and suggest that flow cytometry may be a useful tool in studying lipid packing. PMID- 10520200 TI - Annexin V used for measuring apoptosis in the early events of cellular cytotoxicity. AB - BACKGROUND: Current cytotoxic assays, including Cr release and fluorescent assays, do not directly measure the proportion of target cells which are killed by apoptosis. Cell-mediated cytotoxicity induced by CTLs and NK cells is mainly regulated by the perforin-granzyme, the Fas ligand (Fas L), and the Tumor Necrosis Factor (TNF)-alpha pathways. Perforin generates pores in the membrane of target cells, allowing granzyme B to enter and initiate apoptosis. The other effectors, Fas L and TNF-alpha act by an apoptosis mechanism, leading to DNA fragmentation. A three color flow cytometric method to measure cell-mediated cytotoxicity induced by CTLs or NK cells is described. METHODS: The fluorochromes used are: PKH-26, a stable membrane dye for the labeling of the effector cells, annexin V-FITC which allows the direct evaluation of early apoptotic cells and propidium iodide which distinguishes membrane permeabilized and late apoptotic cells. RESULTS: By eliminating through gating PKH-26 positive effector cells, we obtain a direct estimation of the percentage of target cells in the early stages of apoptosis as well as the percentage of target cells dying after late apoptosis and membrane permeabilization. The cytotoxic activity of IL-2 stimulated PBL against K562, Jurkat and KYM-1 was evaluated. CONCLUSIONS: This rapid and novel assay permits the discrimination of the cell death mechanisms occurring during a cytotoxic response and to precisely evaluate the contribution of apoptosis in the early phases of cell-mediated cytotoxicity. PMID- 10520199 TI - Comparative analysis of apoptosis in HL60 detected by annexin-V and fluorescein diacetate. AB - BACKGROUND: Our aim was to compare and evaluate apoptosis formation as detected by propidium-iodide (PI)/annexin-V or PI/fluorescein-diacetate (FDA) as dose response parameters in a human promyelocytic leukemia cell line, HL60. METHODS: In exponentially growing HL60 cells, apoptosis was induced by ionizing radiation, hyperthermia, topotecan, and cytosine beta-D-arabinofuranoside. At 4 consecutive days following induction, apoptosis was detected by double-labelling, either with PI/annexin-V or PI/FDA. Forward and side scatter, red (PI), and green (FDA or annexin-V) fluorescence were measured by flow cytometry. RESULTS: While light scatter discriminated between morphologically damaged and undamaged cells, fluorescence differentiated vital, apoptotic, and dead cells. Equal proportions of these three subpopulations were detected by both staining techniques. Occasionally, early and mature apoptoses were identified as distinct clusters. During the 4-day observation period, no pronounced maxima of the apoptotic fractions were obtained with either treatment modality. The gradual increases usually showed a delay of 1-2 days. CONCLUSIONS: FDA and annexin-V are equally suitable for detecting apoptosis. Separation improves with time after induction, indicating that, with respect to test specificity, mature apoptoses are superior to early stages. However, the sensitivity towards low rates of apoptosis after weak induction appears limited with both staining procedures. PMID- 10520201 TI - Sensitive analysis of recombination activity using integrated cell surface reporter substrates. AB - BACKGROUND: Recombination processes play a crucial role in the functioning of the immune system and are also involved in mutation events that result in various malignancies. So far the study of recombination activity has frequently relied on the use of reporter substrates that are limited by low sensitivity as well as tedious and distorting readout procedures. METHODS: Immunoglobulin class switch recombination substrates were generated which, upon recombination, resulted in the surface expression of human CD4 or murine MHC class I H-2K(k) and thus allowed for cytometric evaluation. RESULTS: Recombining cells harboring integrated reporter substrates were analyzed by immunofluorescence and flow cytometry and could easily be isolated by high-gradient magnetic cell sorting (MACS). The analysis was not influenced by cloning efficiencies, as would be the case after drug selection, or prokaryotic recombination that might occur after analysis of recovered substrates in bacteria. In addition, cytometric readout is much faster, as it can be performed immediately after recombination. The substrate exhibited properties compatible with the detection of immunoglobulin class switch recombination and permitted the detection of recombination events down to 10(-5) per cell and generation. CONCLUSIONS: The high sensitivity of this system allows precise detection of very rare recombination events and thus permits the study of cell types with extremely low recombination activities. PMID- 10520202 TI - Particle classification from light scattering with the scanning flow cytometer. AB - BACKGROUND: The differential light-scattering pattern, an indicatrix, provides the most complete characterization of the optical properties of a particle. Particle classification can be performed on the basis of particle parameters retrieved from the indicatrices. This classification extends the ability of flow cytometry in particle recognition. METHODS: The scanning flow cytometer (SFC) permits an acquisition of traces of light scattering signals, i.e., native SFC traces, from single particles. The acquired native SFC traces are transformed into indicatrices. The performance of the SFC in measurements of indicatrices has been demonstrated for the following particles: lymphocytes, erythrocytes, polystyrene particles, and milk-fat particles. RESULTS: The structure and profile of the indicatrix for each particle type have been found to be unique. Classification of polystyrene particles has been performed on the basis of the map formed by particle refractive index and size. The polystyrene particles were classified using this map into different size categories ranging from 1.4-7 microm, with a size deviation of 0.07 microm. CONCLUSIONS: The method based on analysis of native SFC traces shows better performance in particle classification than the method based on the particle refractive index and size map. The classification performance of the SFC will be useful, for example, for particle sorting and particle identification, and with additional fluorescent measurements may have applications in multiparameter particle-based immunoassay. PMID- 10520203 TI - Fluorescence enhancement of DNA-bound TO-PRO-3 by incorporation of bromodeoxyuridine to monitor cell cycle kinetics. AB - BACKGROUND: The detection of DNA-incorporated bromodeoxyuridine (BrdUrd) in mammalian cells is a well-known and important technique to study cell cycle. The use of TO-PRO-3 for detection of BrdUrd substitution of DNA by dual-laser flow cytometry has been investigated. METHODS: Fluorescence enhancement of TO-PRO-3 in BrdUrd-labeled cells is registered in combination with the fluorescence emission of the intercalating dye propidium iodide (PI) as a total DNA stain to give bivariate DNA/BrdUrd histograms. By the low concentration of only 0.3 mircoM TO PRO-3, BrdUrd detection is optimized, and undisturbed total DNA content by PI can be detected as well. TO-PRO-3 is excited by a red HeNe laser and PI by an argon ion laser. RESULTS: In order to understand the binding of TO-PRO-3, energy transfer from PI to TO-PRO-3 has been measured as well as the influence of an external DNA binding dye such as Hoechst 33258 with Adenine-Thymine (AT) binding specificity. Cell cycle studies of human SCL-2 keratinocytes and mouse 3T3 cells prove the method to be as generally applicable as the classical BrdUrd/Hoechst quenching technique, but without need for expensive ultraviolet laser excitation. No BrdUrd sensitivity could be found for the similar dyes TO-PRO-1 and YO-PRO-3, whereas TO-PRO-5 and YOYO-3 showed only very little sensitivity to BrdUrd labeling as compared with TO-PRO-3. CONCLUSIONS: Cell cycle studies of mammalian cells can be done by dual-laser flow cytometry without the need for ultraviolet lasers by using the BrdUrd-dependent fluorescence enhancement of TO-PRO-3. Total DNA content can be measured simultaneously using PI. PMID- 10520204 TI - Orientation of the chromophore dipoles in the TOTO-DNA system. AB - BACKGROUND: Flow cytometry has been applied successfully to the sizing of medium to large-sized DNA molecules, thanks to the excellent staining properties of cyanine chromophores such as TOTO (homodimer of thiazole orange) (Petty et al.: Anal Chem 67:1755-1761, 1995). The hydrodynamic flow, used to focus the sample molecules in a small laser-illuminated volume, is also responsible for their alignment, thereby allowing the determination of the TOTO-dipole orientation with respect to the DNA axis (Agronskaia et al.: Appl Opt 38:714-719, 1999). METHODS: We present model calculations of the fluorescence yield of TOTO-stained DNA measured in a flow-cytometric setup with high numerical aperture. The models consider different orientations of the chromophore dipoles. RESULTS: Comparison of measurement and calculation suggests that the absorption dipoles of the TOTO molecule make a mean angle of 61 degrees with the helix axis of the DNA molecule. This mean angle can be the consequence of two binding modes. CONCLUSIONS: Our results indicate that any model with a significant contribution of perpendicularly-oriented chromophores fails to reproduce the experimental results. PMID- 10520205 TI - Detection of baculovirus-infected insect cells by flow cytometric side-scatter analyses. AB - BACKGROUND: The baculovirus expression vector system (BEVS), utilizing the Autographa californica nuclear polyhedrosis virus (AcNPV), has turned out to be an attractive alternative for high-level expression (<600 mg/l) of recombinant proteins. However, there is a shortage of reliable methods for monitoring the infection process in situations where marker proteins cannot be used. METHODS: Three recombinant baculoviruses, FastBac1-wtGFP, VTBac-GFP, and VL1392-hIL 2Ralpha, all having the genes inserted under the transcriptional control of the polyhedrin gene promoter of the Autographa californica nuclear polyhedrosis virus (AcNPV), were used to infect Spodoptera frugiperda (Sf9) and Mamestra brassicae (IZD-MB-0503) insect cells. The infection process of the recombinant baculoviruses was monitored by flow cytometric side-scatter and fluorescence intensity analyses over a period of 6-96 h. RESULTS: A clear correlation between the side-scatter (SSC) signal and the relative fluorescence was observed for both of the infected cell lines, compared to noninfected cells. Comparison of SSC histograms from noninfected insect cells with cells infected with the nonfluorescent recombinant baculovirus VL1392-hIL-2Ralpha showed a clear increase of SSC for the infected cells. CONCLUSIONS: The SSC parameter can therefore be utilized for flow cytometric monitoring of a baculovirus infection process in situations where suitable markers are not available. PMID- 10520207 TI - Reply to the letter by dr. vinogradov PMID- 10520206 TI - Chromatin signal in genome size measurements. PMID- 10520208 TI - Fieller's theorem and linkage disequilibrium mapping. AB - Linkage disequilibrium mapping exploits the fact that at genetic markers close enough to a disease locus on a particular chromosome, we expect to find an association between the disease and marker alleles. Furthermore, the magnitude of the association is expected to follow a unimodal curve when plotted against location, with the peak at the disease location. In practice, for real data, we usually see deviations from such a curve due to other influences such as evolutionary variability, mutation, and selection. Here we propose fitting a quadratic curve to data of this nature, estimating the location of the disease locus by the point at which the curve is maximum. A key feature of our method is the use of transformations of both location and disequilibrium, so that departures from a unimodal curve are incorporated by fitting the curve not to the original location and disequilibrium values but to the transformed values. In addition, we estimate the covariances between the disequilibrium values at linked loci using either a multinomial approximation or a bootstrap procedure. The location estimate from our method is the ratio of two quantities that, in large samples, are normally distributed, and so we use Fieller's theorem to obtain a confidence interval for the disease gene location. We successfully apply our method to data from several published studies in which the true disease gene location is known. PMID- 10520209 TI - Lung cancer risk in families of nonsmoking probands: heterogeneity by age at diagnosis. AB - In an earlier investigation, we did not detect a major genetic component to lung cancer in families of nonsmoking lung cancer probands. However, heterogeneity with respect to familial aggregation, based on probands' age at diagnosis, was evident. We reanalyzed our previously collected data of 257 families, stratified by age at diagnosis of the probands, using complex segregation analysis. We specifically tested the effects of a Mendelian diallelic gene, history of tobacco use, and history of selected chronic lung diseases in families with a proband diagnosed at the age of 60 years or older and in families with a younger proband (i.e. , under 60 years of age). Cases were identified from the Metropolitan Detroit Cancer Surveillance System. Information on lung cancer occurrence, smoking history, and chronic respiratory diseases in first-degree relatives was obtained for 210 older probands and for 47 younger probands. In older probands' families, no evidence of a major genetic effect was detected. A history of emphysema and tobacco-smoke exposure were found to be significant risk factors. In younger probands' families, a Mendelian codominant model with significant modifying effects of smoking and chronic bronchitis best explained the observed data. Our results suggest the presence of a high-risk gene contributing to early onset lung cancer in a population where the probands are nonsmokers. PMID- 10520210 TI - Familial aggregation of breast cancer with early onset lung cancer. AB - Site-specific familial aggregation and evidence supporting Mendelian codominant inheritance have been shown in lung cancer. In characterizing lung cancer families, a number of other cancers have been observed. The current study evaluates whether first-degree relatives of early onset lung cancer cases are at increased risk of breast cancer. Families were identified through population based lung cancer cases and controls under 40 years of age. Cases were ascertained through the Metropolitan Detroit SEER registry; controls through random-digit dialing. Data were available for 384 female relatives of 118 cases and 465 female relatives of 161 controls. Breast cancer in relatives was evaluated after adjusting for age, race, sex, and smoking status of each family member and the sex and age of the probands. A positive family history of early onset lung cancer increased breast cancer risk among first-degree relatives 5. 1 fold (95% CI, 1.7-15.1). Relatives of cases with adenocarcinoma of the lung were at highest risk (RR = 6.3, 95% CI 2.0-20). Mean age of breast cancer diagnosis among relatives of cases was 52.2 years and not statistically different from relatives of controls. Three case families also reported early ovarian cancers (mean age of diagnosis of 35 years). These findings suggest that shared susceptibility genes may act to increase risk of early onset lung and breast cancer in families. PMID- 10520211 TI - Apolipoprotein A-IV-2 allele: association of its worldwide distribution with adult persistence of lactase and speculation on its function and origin. AB - Apolipoprotein A-IV (apo A-IV) is a 46-Kd plasma glycoprotein that may play a major role in intestinal lipid absorption. A genetic polymorphism in the apo A-IV gene, apo A-IV-2, encodes a His-->Gln substitution at codon 360 that alters the biological function of this apolipoprotein. As the worldwide distribution of the apo A-IV-2 allele appeared similar to the frequency of a genetic polymorphism that determines the persistence of lactase into adulthood, we examined the relationship between the apo A-IV-2 and lactase persistence polymorphisms by compiling the prevalence of adult lactase persistence in all populations in which the frequency of the apo A-IV-2 allele has been determined. Across 29 groups, there was an extremely strong correlation (4 = 0.937, P < 0.000001) between apo A IV-2 allele frequency and the prevalence of adult lactase persistence. Apo A-IV-2 allele frequency was highest in Iceland, an ancient Viking colony, and decreased across Europe in a north-to-south and west-to-east gradient, generally following hypothetical isoclines for the lactase persistence gene. There were no correlations between the population frequencies of the apo E2, E3, or E4 alleles and either the prevalence of lactase persistence or the frequency of the apo A-IV 2 allele. In light of the effects of the apo A-IV-2 polymorphism on lipid metabolism, we speculate that the apo A-IV-2 allele may have originated in ancient Scandinavia, spread by conferring a nutritional advantage in the setting of a lifelong high milkfat intake, and was later carried southwards by the Viking incursions into Europe. PMID- 10520212 TI - Methionine synthase D919G polymorphism is a significant but modest determinant of circulating homocysteine concentrations. AB - Elevation in plasma homocysteine concentration has been associated with vascular disease and neural tube defects. Methionine synthase is a vitamin B(12)-dependent enzyme that catalyses the remethylation of homocysteine to methionine. Therefore, defects in this enzyme may result in elevated homocysteine levels. One relatively common polymorphism in the methionine synthase gene (D919G) is an A to G transition at bp 2,756, which converts an aspartic acid residue believed to be part of a helix involved in co-factor binding to a glycine. We have investigated the effect of this polymorphism on plasma homocysteine levels in a working male population (n = 607) in which we previously described the relationship of the C677T "thermolabile" methylenetetrahydrofolate reductase (MTHFR) polymorphism with homocysteine levels. We found that the methionine synthase D919G polymorphism is significantly (P = 0.03) associated with homocysteine concentration, and the DD genotype contributes to a moderate increase in homocysteine levels across the homocysteine distribution (OR = 1.58, DD genotype in the upper half of the homocysteine distribution, P = 0.006). Unlike thermolabile MTHFR, the homocysteine-elevating effects of the methionine synthase polymorphism are independent of folate and B(12) levels; however, the DD genotype has a larger homocysteine-elevating effect in individuals with low B(6) levels. This polymorphism may, therefore, make a moderate, but significant, contribution to clinical conditions that are associated with elevated homocysteine. PMID- 10520213 TI - Density-dependent resistance to apoptosis in retinal cells. AB - PURPOSE: To study effects of cell density on retinal cell survival. METHODS: Apoptotic cell death was induced in cultured retinal cells seeded at higher or lower density by various stimuli including simulated ischemia, excitotoxicity and antibody against heat shock protein 27 (hsp27). Quantitative analysis of apoptotic cells was performed using terminal deoxynucleotidyl transferase mediated dUTP nick end labeling technique and flow cytometry. Cytoskeleton was examined using immunocytochemistry and specific staining of actin by phalloidin and DNase I. In addition, alterations in the cytoskeletal proteins, bcl-2 family of proteins and hsp27 were studied using western blotting. RESULTS: Incubation of the cells under apoptotic stimuli caused higher rates of apoptosis in lower density cultures as determined by TUNEL technique and flow cytometric analysis. Both morphologic examination of cytoskeleton and western blotting revealed that after incubation with various stressors, degradation of actin and tubulin was more prominent in lower density cultures compared to higher density cultures. The expression of bcl-2 and bcl-xL was higher and the expression of bax was lower in lower density cultures compared to higher density cultures at basal condition. After incubation with stressors, bcl-2 and bcl-xL expressions decreased and bax expression increased in both lower and higher density cultures. However, we observed that the expression of hsp27 was higher in higher density cultures than in lower density cultures in the presence or absence of apoptotic stimuli. CONCLUSIONS: These findings demonstrate that retinal cells are more resistant to apoptosis in higher density cultures, independent of the inducer. This might be partly due to protective activity of endogenous hsp27 in the cells at higher density, which contributes to cytoskeletal integrity in response to apoptotic stimuli. PMID- 10520214 TI - Expression of sarco/endoplasmic reticular Ca(2+)-ATPase in human lens epithelial cells and cultured human lens epithelial B-3 cells. AB - PURPOSE: Sarco/Endoplasmic Reticulum Ca(2+ )-ATPase (SER-CAs) isoforms, SERCA2b (100Kd) and SERCA3 (97Kd), are co-expressed in some non-muscle tissues. Both types of SERCAs play important roles in Ca(2+) regulation in non-muscle tissues such as brain and platelets. We tested whether these two SERCAs are present in human lens epithelial cells, human lens epithelial B-3 cells (HLE B-3 cells) and rat lens epithelial cells providing a basis for further studies the role of SERCAs in cataract formation. METHODS: Lens epithelial cells from rat lens, human lens and cultured human lens epithelial B-3 cell line (HLE B-3) were used to extract mRNA and membrane proteins. Reverse transcription polymerase chain reaction (RT-PCR) was employed to detect SERCA3 and SERCA2b mRNA using SERCA2b and SERCA3 primers. Western blotting was used to detect SERCA3 and SERCA2b proteins using anti-SERCA2b and 3 antibodies. RESULTS: A 270 bp fragment was amplified with SERCA2b primers and a 186 bp fragment amplified with SERCA3 primer in human lens epithelial cells, human lens epithelial B-3 cells and rat lens epithelial cells. Both fragments are the expected SERCA2b and SERCA3 products. The sequences of these two fragments in human lens epithelial cells are 100% and 98% homologous to the published partial SERCA2b and SERCA3 mRNA sequence, respectively. The amino acid sequences translated from both PCR products are 100% homologous to the published SERCA2b and SERCA3 sequences in GeneBank. Western blotting confirmed that the expected 100 KDa and a 97 KDa proteins are recognized by anti-SERCA2b and anti-SERCA3 antibodies. CONCLUSION: Sarco/Endoplasmic Reticulum Ca(2+)-ATPase (SERCAs) isoforms 2b and 3 mRNAs are expressed in human lens epithelial cells, the cultured human lens epithelial B-3 cell line (HLE B-3) and rat lens epithelial cells. SERCA2b and SERCA3 may play an important role in the regulation of calcium homeostasis in lens epithelial cells. PMID- 10520215 TI - Expression of integrin receptors in the human trabecular meshwork. AB - PURPOSE: To examine the expression of integrin receptors in the human trabecular meshwork. METHODS: The expression of integrins in human tissues was visualized by immunohistochemical staining using integrin-specific antibodies. Immunoprecipitation was performed after biotin labeling of cell surface proteins. Reverse transcriptase-polymerase chain reaction was used to detect the presence of mRNA species for integrin subunits. RESULTS: Human trabecular meshwork tissues obtained from donors 2 to 65 years old stained positively for integrins alpha1, alpha3, alpha4, alpha5, alpha6, alphav, beta1, beta3, beta4 and beta5. The staining was observed mostly around the edges of trabecular beams in association with trabecular meshwork cells. The staining intensity did not appear to vary with donor age. In addition, immunoprecipitation of tissue extracts revealed the presence of integrin alpha2 and confirmed the absence of beta2. Results from polymerase chain reactions were consistent with these findings. CONCLUSIONS: A spectrum of integrin receptors that may have important roles in the cell-matrix interactions are demonstrated in the human trabecular meshwork. The repertoire identified in tissues is similar to that found in cultured cells except that the beta4 expression in tissues is lost in cultures. PMID- 10520216 TI - Quantitative analysis of leukocyte dynamics in retinal microcirculation of rats with short-term ischemia-reperfusion injury. AB - PURPOSE: To characterize the time course of complete recovery of leukocyte velocities in the retinal microcirculation of rats from short-term (5-minute) retinal ischemia. METHODS: After 5 minutes of retinal ischemia produced by clamping the optic nerve, resulting in the occlusion of both central retinal artery and the central retinal vein, we used acridine orange (AO) and a scanning laser ophthalmoscope (SLO) to observe the velocities of leukocytes in precapillary arteriole ( v(a)), capillary (v(c)) and postcapillary venule (v(v)). Measurements were taken at reperfusion time points of 5, 10, 15, 20, 30, 40, 50, 60 and 80 minutes for the ischemic eyes and at 12, 22, 42 and 62 minutes for the control eyes, respectively. RESULTS: Each control velocity was arteriole 25.1 +/- 4.3 mm/ s; venule 16.9 +/- 3.2 mm/s; and capillary 1.54 +/- 0.31 mm/s, respectively. The leukocyte velocities after 5 minutes of ischemia in arteriole and venule recovered completely within 80 minutes of reperfusion; however, the recovery patterns were different. The recovery pattern showed a biphasic increase in arterioles and a monophasic increase in venules. The velocity in capillaries half recovered rapidly, within 5 minutes of reperfusion, but the subsequent recovery was slower and was not complete even at 80 minutes of reperfusion. CONCLUSIONS: In this study, leukocyte velocities in arterioles, venules and capillaries exhibited different recovery patterns following retinal ischemia and subsequent reperfusion. Capillaries, at least within 80 minutes of reperfusion, may have difficulty recovering completely from even short-term (5-minute) ischemia. PMID- 10520217 TI - Preferential adenovirus-mediated transduction of cells at the sites of laser photocoagulation in the rat eye. AB - PURPOSE: This study aimed to investigate the feasibility of recombinant adenovirus-mediated gene transfer into cells implicated in the development of choroidal neovascularization (CNV). METHODS: A rat model of CNV which used laser photocoagulation was developed. Gene delivery into the laser spots was investigated following subretinal injection of recombinant adenoviruses, AdRSVlacZ, AdCMVlacZ or AdCMVgfp. Immunohistochemical analysis was performed using a proliferating cell nuclear antigen antibody and a cytokeratin-specific antibody to identify the cell types transduced by the recombinant adenoviruses. RESULTS: At 7 days post-injection, lacZ expression was detected in 51.6 +/- 13.2% and 71.2 +/- 19.3% of laser spots in AdRSVlacZ- and AdCMVlacZ-injected eyes, respectively. By 28 days post-injection, lacZ expression was only present in AdCMVlacZ-injected eyes. In vivo fundus fluorescent photography of AdCMVgfp injected eyes detected gfp expression in 79.9 +/- 12.7% and 35.6% +/- 19.7% of laser spots at 4 and 7 days post-injection, respectively. Although fundus fluorescent photography did not detect the gfp signal at 10 days post-injection, fluorescent microscopy revealed a gfp signal in 81.3 +/- 6.0% of laser spots. Immunohistochemical analysis detected retinal pigment epithelial (RPE) cells as the most predominant proliferating cell type in the laser spots, although several other proliferating cell types were also identified. X-gal staining showed that the majority of transduced cells were those present in the laser spots. CONCLUSIONS: It is proposed that following laser photocoagulation, proliferating RPE cells are susceptible to adenovirus-mediated gene delivery and that they may be suitable targets for the delivery of antiangiogenic factors by recombinant adenoviruses in order to inhibit developing CNV membranes. PMID- 10520218 TI - Naphthalene-induced cataract model in rats: A comparative study between slit and retroillumination images, biochemical changes and naphthalene dose and duration. AB - PURPOSE: The purpose of the study was to compare different methods of photographic evaluation of cataract formation in rats in response to different regimes of naphthalene treatment. Furthermore, we intended to study the relationship between cataract extension and biochemical parameters. METHODS: Brown Norway rats were treated with 0.10-1.5 g naphthalene/kg body weight, twice a week for ten weeks to induce cataract or placebo. Slit illumination and retroillumination (SI and RI) photographs were produced by an EAS-1000 instrument to document cataract formation as light-scattering intensity. The degree of the cataractous changes was quantified in SI photographs by the peak height and the integrated peak area, and in RI photographs by threshold setting. Finally, the lens concentration of Na(+) and K( +) and the protein composition were analyzed and correlated to the photographic analysis. RESULTS: The degree of the cataractous changes was most linearly related to dose and duration when the integrated peak area was estimated. However, protein fractions were non-linearly related to the cataractous changes estimated. Alterations in concentration of Na(+) and K(+) were small or insignificant, which indicate that naphthalene induced cataract is not caused by osmotic changes. The lowest possible naphthalene dose to induce cataractous changes was between 0.10 and 0.50 g/kg twice a week for ten weeks. CONCLUSIONS: 0.50 and 1.0 g naphthalene/kg twice a week appeared to be optimal, because the rats in these groups were healthy and the cataractous changes were consistent between animals. Thus, the combination of the animal model with the cataract quantification system has the potential to be useful and reliable in studies of cataract-preventive compounds. PMID- 10520219 TI - Depletion of NK cell activity results in growth of hepatic micrometastases in a murine ocular melanoma model. AB - PURPOSE: To study the role of natural killer (NK) cells in growth of spontaneous hepatic metastasis in a murine intraocular melanoma model. METHODS: Tissue culture B16-LS9 melanoma cells were analyzed by flow cytometry for MHC class I expression of all haplotypes and inoculated into the posterior compartment (PC) of one eye of C57BL6 mice. The eyes were enucleated at 12 days post-inoculation and histologically examined for tumor growth. One group of mice (n = 10) were given intraperitoneal injections of anti-asialo GM1 for NK cell depletion post enucleation and a second group of mice (n = 9) served as controls. The mice were sacrificed at 24 days post-inoculation and necropsies were performed to determine the number and size of metastasis. RESULTS: The B16-LS9 cells failed to express MHC class I antigen. Tumor grew in the PC of all eyes and metastasized to the lungs and livers of all mice, with the average number of hepatic micrometastases greater in the NK depleted group versus the control group (p =.009). There was no significant difference in the average number of pulmonary metastases in the treated versus the control group (p =.072). Hepatic metastases grew to an average diameter of 600 microm in diameter in two NK depleted mice. CONCLUSIONS: NK depletion in this model of metastatic ocular melanoma results in increased number and growth of hepatic micrometastases. PMID- 10520220 TI - Biphasic intraocular pressure response to calcitonin gene-related peptide. AB - PURPOSE: To examine the effects of calcitonin gene-related peptide (CGRP) on intraocular pressure (IOP). METHODS: IOP was periodically measured in rabbits treated with intravitreal injection (20 ael) of either: 1) CGRP (10(-4) approximately 10(-7) M) into one eye; 2) CGRP (10(-4) or 10(-6) M) into both eyes 30 min after intravitreal administration of CGRP-(8-37) (10( -3) M), a CGRP1 receptor antagonist, into one eye; 3) CGRP (10(-4 ) or 10(-6) M) into one eye 30 min after intravenous administration (200 mg/kg) of either Nw-nitro-L-arginine methyl ester (L-NAME), a nonselective inhibitor of nitric oxide synthase (NOS), or aminoguanidine (AG), a selective inhibitor of inducible NOS; or 4) CGRP (10( 4) or 10(-6) M) into one eye along with intraperitoneal indomethacin (50 mg/ kg at -1 and 4 h). RESULTS: CGRP (10(-4) and 10(-5) M) produced a biphasic IOP response, which consisted of an early ocular hypertensive phase and a subsequent sustained hypotensive phase. While CGRP (10(-6) M) yielded only a profound IOP reduction. CGRP-(8-37) significantly inhibited the IOP elevation induced by CGRP (10(-4) M) and completely abolished the IOP reduction induced by CGRP (10(-6) M). The IOP increase induced by CGRP (10(-4) M) was completely abolished by L-NAME, but not affected by AG. The IOP reduction induced by CGRP (10(-6) M) was not affected by L-NAME. Indomethacin did not significantly affect the IOP responses to CGRP. CONCLUSIONS: CGRP dose-dependently produces a biphasic IOP response, which is mediated by CGRP1 receptors. It is suggested that constitutive NOS is involved in the early ocular hypertensive response. PMID- 10520221 TI - Intermediate filament, laminin and integrin expression in lacrimal gland acinar cells: comparison of an immortalized cell line to primary cells, and their response to retinoic acid. AB - PURPOSE: The goal of this study was to characterize intermediate filament, integrin and laminin expression by rabbit lacrimal gland acinar cells in culture, to determine whether retinoic acid (RA) alters expression of these proteins and to compare primary cells to an immortalized rabbit lacrimal gland acinar cell line using flow cytometric analysis. METHODS: Primary cells, maintained in serum free medium, were exposed to 10(-6) M retinoic acid for 24 hours. Immortalized cells were grown in defined medium with Nu-Serum and exposed to retinoic acid. Cells were labeled with monoclonal antibodies to cytokeratins (AE1, AE2, AE3, AE5, CK10/13, CK18), integrins (alpha(3), alpha(6), alpha(V), beta(1), beta(2), beta(3) and beta(4)), laminin, or vimentin and with FITC-conjugated secondary antibodies. Cells were analyzed for antigen expression by flow cytometry and immunocytochemistry. RESULTS: Primary and immortalized cells expressed type I and type II epithelial cytokeratins (AE1 and AE3), cytokeratin 18, and cytokeratin 3 (AE5) Both cell types were negative to AE2 and CK10/13. Primary and immortalized cells expressed vimentin in culture, with immortalized cells expressing this protein at higher levels. Lacrimal acinar cells appear to synthesize laminin which was detected intracellularly in both cells types. Integrins alpha(6) (CD49f) and alpha(V) (CD51) were expressed by primary and immortalized cells. Expression of integrin alpha(6) was 10-fold higher in immortalized cells compared to primary cells. Retinoic acid increased integrin alpha( V) expression by primary and immortalized cells 1.3-fold and 3-fold, respectively, and caused a slight increase in integrin alpha(6) expression by primary cells. Both cell types also expressed integrins beta( 1), beta(2) and beta(3), but beta(4) was detected only in immortalized cells. Lacrimal acinar cells do not express integrin alpha(3). CONCLUSIONS: Expression of cytokeratins, laminin and integrins by primary and immortalized cells was similar, suggesting that the immortalized cell line is a good model for the study of lacrimal structure and function. Since retinoic acid up-regulated only integrin alpha(V), but not cytokeratins, these cells appear to be highly differentiated. Flow cytometry is a useful method for analysis of protein expression by lacrimal acinar cells. PMID- 10520222 TI - Increased cleavage of the c-terminal serine from alpha-A crystallin present in the high molecular weight aggregate fraction from human and bovine lenses. AB - PURPOSE: To determine the relative amount of serine-173 cleavage from the C terminus of alpha-A crystallin from the high molecular weight aggregate fraction and lower molecular weight alpha crystallin fraction from water soluble proteins of human and bovine lenses. METHODS: Using gel permeation chromatography, the high molecular weight aggregate fraction and lower molecular weight alpha crystallin fractions were purified from soluble proteins of human lenses and mature bovine lenses. Alpha-A crystallin present in these fractions was digested with trypsin, followed by the use of reverse phase chromatography and synthetic peptide standards to identify and quantitate the relative amount of in vivo cleavage of C-terminal serine -173. RESULTS: For all human and bovine lenses studied, alpha-A crystallin in the high molecular weight aggregate fraction had undergone more truncation at the C-terminus, than the alpha-A crystallin present in the lower molecular weight alpha crystallin fraction from the same lens. CONCLUSIONS: Relative to the alpha-A crystallin present in the lower molecular weight alpha fraction, the alpha-A crystallin present in the high molecular weight aggregate fraction from the same human and bovine lenses has undergone increased cleavage of the C-terminal serine residue, suggesting a common mechanism that may contribute towards formation of protein aggregation in vivo. PMID- 10520223 TI - Effect of topical interleukin-1 receptor antagonist (IL-1ra) on corneal allograft survival in presensitized hosts. AB - PURPOSE: Topical interleukin-1 receptor antagonist (IL-1ra) promotes corneal allograft survival in naive hosts by suppressing local antigen-presenting cell function and allosensitization. The purpose of these experiments was to test whether IL-1ra could likewise promote transplant survival in pre-sensitized hosts. METHODS: Orthotopic corneal grafts were performed into immunized (N = 65) and naive (N = 40) BALB/c recipients from fully disparate (C57BL/6) or minor H only disparate (B10.D2) mice. Allograft recipients were randomized to receive treatment with IL-1ra or vehicle only. RESULTS: Immunization to both MHC and minor H alloantigens was associated with accelerated graft rejection. IL-1ra treatment had no significant impact on overall graft longevity or the early incidence of graft rejection in pre-immunized recipients of fully-disparate corneal allografts (P = 0.25). Similarly, although rejection of corneal grafts in hosts immunized to minor H alloantigens occurred more slowly in mice treated with IL-1ra than in controls (P = 0.04), overall graft survival at 8 weeks was comparable (P = 0.07) among the two groups. CONCLUSIONS: The accelerated rejection of corneal allografts in presensitized hosts is not materially affected by IL-1ra, particularly if the host has been immunized to donor MHC alloantigens. These data confirm that the role of IL-1ra in corneal transplantation is primarily limited to downmodulating the afferent, not expression, arm of alloimmunity. PMID- 10520224 TI - Differential rapid adhesion of bovine ocular surface epithelial cells to laminin isoforms. AB - PURPOSE: To examine the ability of bovine corneal and conjunctival epithelial cells to adhere to different types of exogenous laminin preparations. METHODS: The ability of bovine corneal and conjunctival epithelial cells in primary culture to attach to laminin isolated from human placenta or from mouse EHS tumor was measured using a short-term colorimetric adhesion assay. Focal adhesion formation in response to interaction with laminins was determined by immunofluorescence microscopy using antibodies to vinculin and morphometric analysis. The influence of laminin on the secretion of adhesion complex proteins by bovine corneal epithelial cells in culture was analyzed using immunofluorescence microscopy. RESULTS: In short-term assays, primary bovine corneal epithelial cells demonstrate rapid and efficient adhesion to placental laminin, and significantly more cells contain focal adhesions, compared to those incubated on EHS laminin. In contrast, primary bovine conjunctival epithelial cells adhere equally well to placental and EHS laminin over a range of substrate concentrations. Additionally, the percentage of cells containing focal adhesions is not significantly different. In primary bovine corneal epithelium, the deposition of collagen type IV and collagen type VII into extracellular network like structures is inhibited in cells cultured on placental laminin compared to cells cultured on EHS laminin. CONCLUSIONS: In vitro, bovine corneal epithelial cells attach more rapidly and efficiently to exogenous placental laminin compared to EHS laminin. However, this isoform inhibits the ready formation of adhesion complex-like structures in culture. The laminin isoform found in human placental preparations may therefore modulate corneal epithelial cell motility as opposed to permanent adhesion. PMID- 10520225 TI - The effect of selective cyclooxygenase-2 inhibitor on corneal angiogenesis in the rat. AB - PURPOSE: Eicosanoids that are present in inflamed tissues are thought to play a significant role in angiogenesis. Cyclooxygenase, a key enzyme in eicosanoid synthesis, has recently been shown to exist in two isoforms: the constitutive COX 1 and the inducible COX-2. This study was undertaken to determine the role of COX 2 in the corneal angiogenic response. METHODS: Angiogenesis in the rat cornea was provoked by chemical cautery. Either NS-398, a selective COX-2 inhibitor, or indomethacin, a non-selective COX inhibitor, was applied topically 3 times daily for 4 days. Neovascularization was quantitated by digital image analysis in corneal flat preparations. To test their inhibitory effects on eicosanoid synthesis, normal or cauterized corneas were incubated in the culture medium with the inhibitor. Prostaglandin E2 in the medium was assayed using an enzyme-linked immunosorbent assay. RESULTS: Both NS-398 and indomethacin significantly inhibited corneal neovascularization with the % inhibition of 36.4 +/- 9.6%, and 38.5 +/- 9.0%, respectively, when applied topically at a concentration of 0.1% (p <.001). Neither reduced the angiogenic response at a concentration of 0.01% or below. PGE(2) production in the cauterized cornea was 2.0 times higher than that in the controls. In normal corneas, indomethacin inhibited PGE(2) synthesis by 80%, whereas NS-398 inhibited it by no more than 20%. In contrast, in injured corneas, both indomethacin and NS-398 inhibited PGE(2) synthesis in a similar fashion, with a maximal inhibition rate of 75 to 80%. CONCLUSIONS: Our results suggest that COX-2 induction in cauterized corneas increases the level of eicosanoids, which result in corneal angiogenesis. PMID- 10520226 TI - Effect of diabetes and fructose/non-fructose diet on the optical quality (cataracts) of the rat lens. AB - PURPOSE: This study measures the effect of moderate and severe levels of diabetes on the optical performance of the rat lens, and evaluates the effect of dietary fructose on diabetic lens damage. METHODS: Moderate and severe diabetes were induced in 250 g rats (Harlan Sprague Dawley) with streptozotocin (35 and 55 mg/kg body weight iv). Animals were fed either a control (glucose/cornstarch) or a 40% (wt:wt) fructose and cornstarch diet and then sacrificed after 7 or 12.5 weeks. These two diets were also fed to two groups in which diabetes was not induced and these rats were also sacrificed at 7 or 12.5 weeks. Lenses removed from animals were analyzed in terms of average focal length (mm), focal length variability (spherical aberration, mm) and relative transmission of light (scatter or number of pixels excited by a refracted laser beam) using an automated laser scanning system. RESULTS: Diabetes disrupted rat lens optical function, both at 7 and 12.5 weeks, as indicated by an increase in focal length variability (FLV). This was true for control and fructose diets. For example, after 12.5 weeks on the diets average FLV values of 0.10 +/- 0.00 mm [n = 9], 0.11 +/- 0.01 mm [n = 9],(control and fructose diets, no diabetes) and 0. 48 +/- 0.04 mm [n = 10], 0.95 +/- 0.11 mm [n = 9], (control and fructose diets, severe diabetes), were measured. The difference between control and fructose diets was significant at 12.5 weeks in the severe diabetes group (p 4)-beta-D-Glcp-(1-->4)-[alpha Neu5Ac-(2--> 3)]-beta-D- Galp-(1-->4)]n. A tetrasaccharide corresponding to this repeat unit has been synthesized by us [E. Eichler, H.J. Jennings, D.M. Whitfield, J. Carbohydr. Chem., 16 (1997) 385-411]. Since the protective epitopes are believed to involve several repeat units, methods to extend this tetrasaccharide were examined. This objective requires a glycosylation of the unreactive OH-4 of the beta-L-Rhap, which was accomplished by coupling a D-Galp glycosyl trichloroacetimidate donor with a beta-L-Rhap-(1-->4)-D-Glcp acceptor. Subsequent coupling of this trisaccharide as a donor to an alpha-Neu5Ac-(2-->3)-D Galp disaccharide acceptor gave a pentasaccharide. The pentasaccharide was deprotected and enzymatically sialylated using an alpha-(2-->3)-sialyltransferase from Campylobacter jejuni to give the title hexasaccahride alpha-Neu5Ac-(2-->3)- beta-D-Galp-(1-->4)-beta-L-Rhap-(1-->4)-beta-D-Glcp-(1-->4)-[alpha -Neu5Ac- (2- >3)]-beta-D-Galp-(1-->O)-(CH2)3N3. PMID- 10520253 TI - UDP-6-deoxy-6-fluoro-alpha-D-galactose binds to two different galactosyltransferases, but neither can effectively catalyze transfer of the modified galactose to the appropriate acceptor. AB - The effect of substitution of the HO-6 of D-galactose with fluorine on the ability of alpha-(1-->3)-galactosyltransferase (EC 2.4.1.151) and beta-(1-->4) galactosyltransferase (EC 2.4.1.22) to catalyze its transfer from UDP to an appropriate acceptor was determined. HPLC analyses indicated that each transferase properly catalyzed formation of the expected product [beta-D-Gal-(1- >4)-D-GlcNAc] for the beta-(1-->4)-galactosyltransferase and alpha-D-Gal-(1-->3) beta-D-Gal-(1-->4)-D-GlcNAc for the alpha-(1-->3)-D-galactosyltransferase] when UDP-alpha-D-Gal was the substrate. When UDP-6-deoxy-6-fluoro-alpha-D-galactose (6) was used in conjunction with each transferase, no product indicative of transfer of 6-deoxy-6-fluoro-D-galactose to its respective acceptor sugar was identified. 6-Deoxy-6-fluoro-D-galactose (3) was obtained by hydrolysis of methyl 6-deoxy-6-fluoro-alpha-D-galactopyranoside, synthesized by the selective fluorination of methyl alpha-D-galactopyranoside with diethylaminosulfur trifluoride (DAST), with aqueous trifluoroacetic acid. Acetylation of 3 gave crystalline 1,2,3,4-tetra-O-acetyl-6-deoxy-6-fluoro-beta-D-galactopyranose, which was converted to the corresponding 1-alpha-phosphate and used for the synthesis of 6. PMID- 10520254 TI - Transglycosylation of cellobiose by partially purified Trichoderma viride cellulase. AB - A commercial cellulase from Trichoderma viride was fractionated into three fractions, F1, F2, and F3, in order to investigate transglycosylation activities. Among these fractions, F3, which demonstrated highly hydrolytic activity toward p nitrophenyl beta-D-glucopyranoside and Avicel, most effectively catalyzed the transglycosylation of cellobiose and converted cellobiose into beta-Glc-(1-->6) beta-glc-(1-->4)-Glc and beta-Glc-(1-->6)-beta-Glc-(1-->6)-beta-Glc(1-->4)-Glc. The F3 fraction contained the enzyme to catalyze beta-glucosyl transfer toward only the C-6 position of the sugar acceptor, and thus it is expected to be of use for syntheses of functional oligosaccharides. PMID- 10520255 TI - Introduction of a carboxyl group through an acetal as a new route to carboxylic acid derivatives of sugars. AB - A new class of carboxylic acid derivatives of sugars is described. Acetalation of mono- and disaccharides with a functionalized vinylic ether or a diethoxybutanoate afforded mono- and diacetals bearing an ester group. Their saponification led to the corresponding carboxylic acid acetals in which the length of the acetal side chain can be modulated. PMID- 10520256 TI - Enzymatic glycosylation of reducing oligosaccharides linked to a solid phase or a lipid via a cleavable squarate linker. AB - Reducing oligosaccharides were converted into their corresponding glycosylamines, and these were reacted with 3,4-diethoxy-3-cyclobuten-1,2-dione (squaric acid diethyl ester). The resulting derivatives could be linked to amino-functionalized lipids, solids, or proteins. Treatment of the obtained lipid or solid conjugates with aqueous bromine or, alternatively, with ammonia-ammonium borate cleaved the linkage and regenerated the oligosaccharide glycosylamines, which were in turn rapidly hydrolyzed to the reducing oligosaccharides. To demonstrate the usefulness of this linkage in enzymatic oligosaccharide synthesis, lactose was linked to a lipid or a solid phase, the obtained conjugates were then subjected to two enzymatic glycosylations (either consecutively or 'one-pot'). The resulting materials were then cleaved to give, in both cases, the expected reducing tetrasaccharide (lacto-N-neotetraose) in good yield. PMID- 10520257 TI - The structure of the core part of Proteus mirabilis O27 lipopolysaccharide with a new type of glycosidic linkage. AB - The structural assignment of the intact lipopolysaccharide core from Proteus mirabilis O27 has been completed based on a combination of chemical degradation studies, NMR spectroscopy and ES-MS spectroscopy. The overall core structure is as follows: [formula: see text] where all sugars are in the pyranose form except the N-acetylglycosamine residue, Hep refers to L-glycero-alpha-D-manno heptopyranose and alpha-DDHep to D-glycero-alpha-D-manno-heptopyranose. Bold italics indicate non-stoichiometric substituents. A new type of glycosidic linkage has been discovered wherein a GalNAc residue is linked as an open form acetal to the 4- and 6-positions of a 2-amino-2-deoxygalactopyranose residue. This structural element is abbreviated GaloNAc-4,6-, where the 'o' indicates the open form of the sugar residue. PMID- 10520259 TI - A novel trisaccharide glycolipid biosurfactant containing trehalose bears ester linked hexanoate, succinate, and acyloxyacyl moieties: NMR and MS characterization of the underivatized structure. AB - A Gram-positive actinomycete growing on n-hexadecane secreted a family of anionic glycolipid surfactant homologs. The major homolog, with a molecular weight of 1210.6347, had the formula C58H98O26. Following mild alkaline saponification, 1H and 13C NMR spectroscopy were used to characterize the non-reducing trisaccharide backbone: beta-Glcp-(1-->3)-alpha-Glcp-(1<-->1)-alpha-Glcp ('laminaratrehalose'). Hexanoate, succinate, 3-hydroxyoctanoate, and 3-hydroxydecanoate were found in 3:1:1:1 molar ratio using GC-EIMS analysis of fatty acid methyl esters (FAME) prepared by transesterification. We found that the beta-hydroxy acids bore secondary hexanoate chains in 3-O-ester linkage, giving acyloxyacyl anions of appropriate m/z in FABMS and FABMS/MS spectra. COSY, HETCOR, HMBC, and HMQC NMR experiments established the acylation pattern: succinate at C-2 of the terminal alpha-glucopyranose ring; hexanoate at C-3" of the beta-glucopyranose ring; 3 hexanoyloxyoctanoate and 3-hexanoyloxydecanoate at the 2'- and 4-positions. In FABMS spectra, the homologs flanked the molecular ion by +/- 14 and +/- 28 amu, suggesting heterogeneity in acyl chain length. PMID- 10520260 TI - Chemical structure of a polysaccharide from Campylobacter jejuni 176.83 (serotype O:41) containing only furanose sugars. AB - A neutral polysaccharide was obtained by hot phenol-water extraction of biomass from Campylobacter jejuni 176.83 and subsequently separated from acid-liberated core oligosaccharide of lipopolysaccharide by sequential GPC on Bio-Gel P6 and TSK-40 columns. All sugar components of the trisaccharide repeating unit of the polysaccharide were found to be of the furanose ring form. The major trisaccharide contained beta-L-arabinose, 6-deoxy-beta-D-altro-heptose (beta-D-6d altHep) and 6-deoxy-beta-L-altrose (beta-L-6d-Alt), whereas in the minor trisaccharide the beta-L-6d-Alt is replaced by its C-5 epimer alpha-D-Fuc. On the basis of 1H and 13C NMR spectroscopic studies, including 2D ROESY, HMQC and HMQC TOCSY experiments, the following structures of the repeating units were established: [formula: see text] PMID- 10520258 TI - Hydrolysis of wheat bran and straw by an endoxylanase: production and structural characterization of cinnamoyl-oligosaccharides. AB - Hydrolysis of wheat bran and wheat straw by a 20.7 kDa thermostable endoxylanase released 35 and 18% of the cell-wall xylan content, respectively. Separation of the cinnamoyl-oligosaccharides (accounting for 6%) from the bulk of total oligosaccharides was achieved by specific anion-exchange chromatography. The cinnamoyl-oligosaccharides were further purified by preparative paper chromatography (PPC) and their molecular weight was determined by MALDI-TOF mass spectrometry. The partially purified hydrolysis end-products contained from 4 to 16 and from 4 to 12 pentose residues for wheat bran and straw, respectively, and only one cinnamic acid per molecule. The primary structure of the new feruloyl arabinoxylopentasaccharide from wheat bran hydrolysis, which has been determined using 2D NMR spectroscopy, is O-beta-D-xylopyranosyl-(1-->4)-O-[5-O- (feruloyl) alpha-L-arabinofuranosyl-(1-->3)]-O-beta-D-xylopyranosy l-(1-->4) -O-beta-D xylopyranosyl-(1-->4)-D-xylopyranose. PMID- 10520261 TI - Structural studies on the acidic exopolysaccharide from Haloferax denitrificans ATCC 35960. AB - The structure of a linear, acidic exopolysaccharide isolated from the Archaeon Haloferax denitrificans ATCC 35960 has been determined using NMR spectroscopy. The sugar residues in the repeating unit of the polysaccharide were identified as Gal and GlcA2,3NAc after the assignment of the 1H and 13C resonances using COSY, HOHAHA, HMQC and HMQC-TOCSY experiments. The sequence of the residues in the polysaccharide was established from the inter-residue connectivities observed in the HMQC-NOESY plot. The only sugar released on acid hydrolysis was shown to be D Gal by GLC analysis, while the absolute configuration of the acidic sugars was shown to be D by comparison of the carbon chemical shifts with those of model compounds. Partial acid hydrolysis yielded a tetrasaccharide, terminated by D-Gal at the reducing end, whose structure confirmed that of the repeating unit of the polysaccharide as-->4)-beta-D-GlcpA2,3NAc-(1-->4)-beta-D-GlcpA2, 3NAc-(1-->4) alpha-D-GlcpA2,3NAc-(1-->3)-alpha-D-Galp- (1-->, where D-GlcpA2,3NAc is 2,3 diacetamido-2,3-dideoxy-D-glucopyranosiduronic acid. PMID- 10520262 TI - Ozonolytic depolymerization of polysaccharides in aqueous solution. AB - The selective oxidation of beta-D-glycosidic linkages of polysaccharides by ozone has great utility as a general method for depolymerization of polysaccharides. Here we describe a 'one-step' method whereby polysaccharides dissolved in water or basic solutions are depolymerized by ozonolysis. The oxidation of glycosidic linkages of unprotected carbohydrates by ozone is complicated by several side reactions. We describe here optimized conditions for carrying out ozonolysis degradation. We also characterize the major pathways for unwanted degradation by various side reactions. In the preferred oxidation pathway, the aldosidic linkage is oxidized to an aldonic ester function that hydrolyzes under the basic conditions employed to give a free aldonate, with cleavage of the polysaccharide chain. Nonselective degradation pathways include oxidative degradation by radical species that oxidize glycosyl residues to formic, acetic, and oxalic acids. The nonselective degradation caused by acids is minimized by basic buffers. The products of polysaccharide depolymerization form a size distribution around a nominal molecular weight, and the average molecular weight of the products can be controlled by the rate or amount of ozone passed through the reaction mixture. The ozonolysis method described herein provides a convenient, inexpensive, and controllable means for generating small polysaccharides or large oligosaccharide fragments. PMID- 10520263 TI - The O-chain polysaccharide of the lipopolysaccharide of Xanthomonas campestris pv. begoniae GSPB 525 is a partially L-xylosylated L-rhamnan. AB - The O-chain polysaccharide (OPS) of the lipopolysaccharide of Xanthomonas campestris pv. begoniae GSPB 525 was found to contain L-rhamnose and L-xylose in the ratio 1:0.6. The OPS lacked strict regularity because of nonstoichiometric xylosylation of the main rhamnan chain. Based on methylation analysis, Smith degradation, and 1H and 13C NMR spectroscopy, including COSY, TOCSY, NOESY, and H detected 1H,13C heteronuclear multiple-quantum coherence (HMQC) experiments, the following structure of the OPS was established: [formula: see text]. PMID- 10520264 TI - Further data on the structure of brown seaweed fucans: relationships with anticoagulant activity. AB - The composition, molecular weight (MW), anticoagulant activity and nuclear magnetic resonance spectra of various low-molecular-weight fucans (LMWFs) obtained by partial hydrolysis or radical depolymerization of a crude fucoidan extracted from the brown seaweed Ascophyllum nodosum are compared. Fucose units were found mainly sulfated at O-2, to a lesser extent at O-3, and only slightly at O-4, contrary to previously published results for fucoidans from other brown seaweeds, and fucose 2, 3-O-disulfate residues were observed for the first time. As the sulfation pattern excluded an alpha-(1-->2)-linked fucose backbone and a high proportion of alpha-(1-->4) linkages was found, it would appear that the concept of fucoidan structure needs to be revised. Anticoagulant activity is apparently related not only to MW and sulfation content, as previously determined, but also (and more precisely) to 2-O-sulfation and 2,3-O-disulfation levels. PMID- 10520265 TI - Chemoenzymatic synthesis of the Salmonella group E1 core trisaccharide using a recombinant beta-(1-->4)-mannosyltransferase. AB - The chemical synthesis of the bacterial O-antigen from Salmonella serogroup E1, 3 O-(4-O-beta-D-mannopyranosyl-alpha-L-rhamnopyranosyl)-alpha-D-galactos e, presents a particular challenge because it contains a beta-(1-->4) mannosidic linkage to L-rhamnose. We report a chemoenzymatic synthesis of this crucial antigenic material which culminates in the enzymatic formation of the critical beta-mannosyl connection catalyzed by Salmonella GDP-alpha-D-Man:alpha Rha1-->3 alpha Gal-PP-Und beta-(1-->4)-mannosyltransferase (ManT beta 4). In comparison with previous synthetic routes, this method is advantageous since it utilizes intermediates, available in significant yield, which can be readily derivatized from the reducing end to present flexibility for analog construction, while the enzymatic construction of the Man1-->4Rha glycosidic bond is both rapid and occurs in high yield. Furthermore, the reported spectroscopic and enzymatic structural characterization of the trisaccharide product furnishes the first indisputable functional link between wbaO and ManT beta 4 and clearly sets the stage for the future mechanistic study and exploitation of this fascinating glycocatalyst. PMID- 10520266 TI - Structure of an acidic O-specific polysaccharide of Proteus mirabilis O5. AB - The following structure of the O-specific polysaccharide of Proteus mirabilis O5 was established by 1H and 13C NMR spectroscopy at 500 MHz, including two dimensional COSY, TOCSY, NOESY, and H-detected 1H, 13C heteronuclear multiple quantum coherence (HMQC) experiments: [formula: see text] where O-acetylation of alpha-D-GlcNAc at both positions is nonstoichiometric. PMID- 10520267 TI - Structure of the O-specific polysaccharide for Acinetobacter baumannii serogroup O1. AB - A polymeric fraction containing D-galactose, N-acetyl-D-galactosamine, and N acetyl-D-glucosamine was isolated from the lipopolysaccharide produced by the reference strain for Acinetobacter baumannii serogroup O1. By means of NMR spectroscopy, methylation analysis, and chemical degradation, the repeating unit of the polymer was identified as a branched trisaccharide of the following structure. [formula: see text]. PMID- 10520268 TI - Studies of pectic enzymes produced by Talaromyces flavus in submerged and solid substrate cultures. AB - Tons of peel and rag are generated each year by industries of citrus fruit juices. These by-products are used either for the elaboration of pectin or as substrate for enzyme production. Talaromyces flavus produces extracellular pectinesterase and polygalacturonase after 24 h in submerged culture supplemented with 0.5-0.8% citrus pectin preceded by preculture for 24 h in 2% (w/v) sucrose or in solid substrate culture on passion fruit peel, lemon or orange pulp pellets after 3-6 days of incubation. Chromatographic profiles in a CM-Sepharose column of liquid and solid cultures were very similar, consisting of one endopoligalacturonase (endo-PG I) and one pectinolytic complex constituted by an endopoligalacturonase (endo-PG II) and pectinesterase. Pectin and pectate lyases were undetectable in both media. In Talaromyces flavus the synthesis of pectinases was repressed by glucose and finally controlled by the concentration of products from pectic enzymes degradation. PMID- 10520269 TI - Transcription level of operon ftsYEX and activity of promoter P1 of rpoH during the cell cycle in Escherichia coli. AB - In Escherichia coli, the genes of the main heat-shock proteins are under the control of the product of gene rpoH, protein sigma 32. The distal promoter P1 of rpoH is located within the terminator of the division operon ftsYEX which could imply some coupling between ftsYEX transcription termination, P1 transcription and cell division. To study the possibility of this coupling, the level of transcription of ftsYEX and the activity of promoter P1 of rpoH were examined in synchronous cultures. Results indicate that transcription of ftsYEX and of rpoH from P1 is continuous, suggesting that ftsYEX transcription termination and P1 activity are not coupled to the cell cycle. PMID- 10520271 TI - Insoluble glucans synthesized by cariogenic streptococci: a structural study. AB - Of the three cariogenic streptococci grown in four various culture media, the strain Streptococcus mutans 20,381 was found to produce large amounts of extracellular glucosyltransferase and water-insoluble, adhesive exopolysaccharide when grown in batch culture on brain-heart infusion broth. Methylation and nuclear magnetic resonance analyses revealed that the insoluble polymers synthesized by the crude glucosyltransferase preparations were mixed-linkage (1- >3), (1-->6)-alpha-D-glucans (so-called mutans) with a greater proportion of 1,3 to 1,6 linkages and major branch points of 3,6-linked glucose. The percentage content of different types of linkages in glucans varied widely and depended on the strain of cariogenic bacteria used to produce glucosyltransferase, and on the kind of medium utilized to cultivate mutans streptococci. The potential application of insoluble glucan produced by mutans streptococci is discussed. PMID- 10520270 TI - Production and shelf-life studies of low cost beverage with soymilk, buffalo cheese whey and cow milk fermented by mixed cultures of Lactobacillus casei ssp. shirota and Bifidobacterium adolescentis. AB - A study was performed to develop a fermented milk beverage with the aim to increase the potential application of buffalo cheese whey and soymilk. A mixed substrate was prepared by selective combination, which contained buffalo cheese whey 35%, soymilk 30% and cow milk 35%. The substrate mixture was fermented by a mixed culture of Lactobacillus casei shirota and Bifidobacterium adolescentis at 37 degrees C for 8 h keeping a 1:1.5 proportion between the lactic and bifidobacteria within a 5% (v/v) inoculum size. The fermented beverage was lightly extra-flavoured with vanilla essence and subjected to chemical, microbiological and sensory evaluations during storage for 28 days at 4 degrees C. Except a slight variation in the acidity, no other properties changed even after 28 days. There were no contaminating organisms (Salmonella and coliforms), which indicated the sanitary and hygienic conditions of the processing and the viable cells of the bacterial strains was well within recommended limits (6.8 x 10(8) cells for L. casei and 2.3 x 10(7) cells for Bifidobacterium). No negative changes were found in the sensory characteristics of the beverage allowing its good acceptability in all during the storage period. PMID- 10520272 TI - [Heat and vestibular stimulation]. AB - In 1905 Barany analyzed the nystagmus that appears during the irrigation of the ear with cold and hot water, justifying the phenomenon due to convectives currents in the horizontal semicircular canal. This hypothesis has been widely accepted until doubt was casted on it by the experiences carried out in the Skylab. Of the various mechanisms that could be responsible for the vestibular stimulation in the caloric tests, those which entail convection currents or endolymph expansion would need--we think--thermal gradients very superior to those occurring during these tests, that according to our calculations--always pro such hypothesis--seem insufficient. PMID- 10520273 TI - [Electronystagmography of sudden head impulses (head-only impulsive rotational testing, head thrust test, head impulse test)]. AB - The basis of the head-thrust test is when a head thrust is done in the horizontal plane, the ipsilateral semicircular canal is the only which gets the gaze stabilization, being the other HSC completely inhibited. In this paper are investigated the ENG records according the test and analyzed the main parameters obtained. As conclusion we admit that the main parameters are the compensating ocular movements amplitude and their latencies. PMID- 10520274 TI - [Measure of the slow phase angular speed by the trigonometric method]. AB - We compare trigonometric and traditional systems for nystagmus slow phase speed measures. Results are detailed in Table I. Mean difference between both procedures is 0.88 +/- 0.51 degree/sec. so we conclude that trigonometric system is a reliable method for these measures. PMID- 10520276 TI - [Caloric surface and assessment of the caloric bithermal test. Toward an intelligent system of vestibular diagnosis]. AB - The theoretical basis for bithermal caloric test assessment by means of caloric surfaces is described. This system supports a computer program for the automatic appraisal of the test, as a first step in the development of our intelligent system for vestibular diagnosis. PMID- 10520275 TI - [Polymorphic reticulosis of the rhinopharynx]. AB - "Malignant centrofacial granulomatosis" is an unusual but devastating spectrum of lymphoproliferative disorders that is now thought to include several entitites, such as polymorphic reticulosis or peripheral T-cell lymphoma. Characteristically there are destructive centrofacial lesions which often produce difficulties for the diagnostic to the clinician and pathologist. The case of a 71-year-old man diagnosed in our Department as polymorphic reticulosis sitting on the rhinopharynx is reported. PMID- 10520277 TI - [Head-thrust test: its validity as a diagnostic clinical test]. AB - The head-thrust test is presented as an ideal test to investigate the horizontal vestibulo-ocular reflex. We review its theoretical basis, and assess it as a diagnostic test with a 22 patient series. Its specificity accounts for 100% and the sensibility for 54%. Would this test substitute in the future the caloric test? PMID- 10520278 TI - [Epithelial-myoepithelial carcinoma of smaller salivary gland]. AB - The epithelial-myoepithelial carcinoma of salivary glands is rarely presented and it's considered as an intermediate grade tumor. Usually are located in the parotid gland, however sometimes occur on minor salivary glands. An epithelial myoepithelial carcinoma sitting on a palatine minor salivary gland of long course is reported. PMID- 10520279 TI - [Epithelioid amelanotic melanoma of the nose and paranasal sinuses]. AB - Mucous melanomas are rare, representing 1-3% of melanomata and 3% of malignant nasosinusal tumors. The more usual location is the nasal cavity and paranasal sinuses. In early stages occur with a poor and unespecific features, which delays the diagnosis and worsen the prognosis. An amelanotic melanoma case is reported and bibliographic review of these tumors are made. PMID- 10520280 TI - [The blink reflex in the electrophysiologic exploration of Bell's palsy and its prognostic value]. AB - Conventional electrophysiology of the facial nerve gives only information about the functional condition of its extracranial segment. The blink-reflex triggered for the electric stimulation of supraorbital nerve allows the study of the whole nerve path, even the intraosteal, where the great majority of its pathology settles. We have realized this tests in 43 Bell's palsy cases (at the beginning of the paralysis and three weeks later) and correlated the results with the clinic course, in order to set up the pronostic value of the procedure. The cases with shorter tendency of blink reflex along the three first week have a better outcome. PMID- 10520281 TI - [Anaplastic carcinoma of the thyroid in a young man. A report of a case]. AB - Anaplastic thyroid carcinoma is a tumor that has a very ominous prognosis, and is characteristic of people older than 60 years. Generally occurs as a neck lump of rapid growth rate, arousing compressive symptoms on nearby structures. We report the case of a 29-year-old patient who presented an anaplastic thyroid carcinoma. He was treated with surgery, chemotherapy and radiotherapy, but died 10 months after being diagnosed. PMID- 10520282 TI - [The enhanced lethality of cells in suspension during simultaneous exposure to pulsed electrical and shock-wave acoustic fields]. AB - We present the results of comparative experimental studies of the damaging effect of intense electrical field with a microsecond duration range, shock acoustic wave, and their combination on the ovary carcinoma cells. We found that upon a combined effect of pulse electrical and acoustic shock-wave fields of a moderate amplitude, practically all cells died. A qualitative physical model is proposed, which accounts for enhanced damage upon combined effect, on the one hand, by a decreased threshold of electric intensity of the cytoplasmic membrane breakdown at a temporal phase of pressure compression of the shock-wave pulse and, on the other hand, by a growth of cavitational embryos at a temporal phase of the action of stretching tensions of the shock-wave pulse, whose role is played by hydrophobic type pores appeared as a result of the membrane electrical breakdown. Possibilities of practical application of the proposed combined effect are discussed: in oncology for suppression of tumor growth and in biotechnology for cell disintegration and ecologically pure sterilization of liquid media. PMID- 10520283 TI - [The structural transformations of X-oligomers]. AB - Mechanism of self-assembly and the three-dimensional organization of the intermediate soluble forms of X-oligomers in the presence of non-denaturing urea concentration have been studied by dynamic light scattering, analytical ultracentrifugation, and viscometric methods. Swedberg and Kuhn-Mark-Hauwink analysis of the obtained hydrodynamic data accounting the concentration effect on translational friction demonstrated formation of equilibrium single-stranded rod like protofibrils with end-to-end arrangement of the peripheral domains of X monomers. Upon elongation the single-stranded protofibrils form double-stranded structures through lateral aggregation. We infer that alpha C domains are involved in neither stabilization of the single-stranded structures nor their dimerization. PMID- 10520286 TI - [In Process Citation] AB - A complex cytophysiological study of the resistance of Aegilops speltoides K-389 and wheat-aegilops seedlings against Erysiphe graminis tritici was carried out. Ae. speltoides proved to be very resistant at the stage of pathogen penetration and endurant to the powdery mildew after infection. The wheat-aegilops lines had varying features of resistance against the powdery mildew but did not retain a sufficiently high resistance of Ae. speltoides at the stage of seedlings. Since the studied lines showed a high resistance in the field, their resistance appears to be determined by the genetic system of protection, predominantly at the adult stage. This should be taken into account when using Ae. Speltoides as a source of resistance by controlling the character of its expression not only at the adult state, but also at the stage of seedlings. PMID- 10520285 TI - [The effect of mutagenic environmental factors on recombination in Drosophila melanogaster]. AB - We studied recombination in Drosophila melanogaster males exposed in a thermoelectric power station in Moscow for six years and, for comparison, in another thermoelectric power station during 1994. The frequency of recombination in the experimental males was two to three times that in the control. The annual differences in the frequency of recombination in the experimental males were not statistically significant. The differences between the data obtained for two different thermoelectric power stations were not statistically significant as well. A marked increase in the frequency of recombination in the D. melanogaster males exposed at thermoelectric power stations may be considered as an adequate reaction to the presence of a sufficient amount of effective mutagens in discharges of thermoelectric power stations. PMID- 10520284 TI - [The biosynthesis of 2H-labelled inosine by Bacillus subtilis bacteria in a heavy hydrogen medium]. AB - We studied biosynthesis of the purine ribonucleoside inosine by the strain of Bacillus subtilis, which was adapted to deuterium through plating to individual colonies on 2% agar with 2H2O and subsequent selection. For growing the strain, a special heavy hydrogen medium with a high level of deuteration (89-90 at. % 2H) based on 2% hydrolyzate of the biomass of the methylotrophic Brevibacterium methylicum as a source of 2H-labeled growth substrates was obtained on 98 vol % 2H2O and 2 vol % [U-2H] methanol. We provide experimental data on the strain growth and accumulation of inosine in the culture medium. A study of the level of inosine deuteration using mass-spectroscopy of fast atoms has shown a polymorphism of deuterium incorporation in the molecule (isotope composition of inosine: 4 at. 2H, 20%; 5 at. 2H, 38%; 6 at. 2H, 28%; 7 at. 2H, 14%), deuterium being incorporated in the ribose and hypoxanthine fragments of the molecules. PMID- 10520287 TI - [Disorders of energy metabolism in the rat brain under extreme exposures: a new method for assessing the energy state of the brain]. AB - We propose a new method to evaluate energy state of the brain based on vital assay for phosphate metabolites and intracellular pH in the brain. A proposed Z index describes a set of correlations between phosphates involved in brain energy metabolism assayed in vivo by magnetic resonance spectroscopy on 31P nuclei. We tried to use Z-index as a quantitative test for disturbed energy metabolism. Potentialities of Z-index for evaluating rat brain energy state after extreme influences (gamma [correction of X-] irradiation and postoperative shock of the brain cortex) have been demonstrated. PMID- 10520288 TI - [The indices of free-radical oxidation in the umbilical cord blood and placenta of inhabitants of the Altai Territory]. AB - Free-radical processes were studied in the umbilical blood and placenta of women from the regions of the Altai Territory, which were affected to different extents by nuclear tests on the Semipalatinsk grounds in 1949-1965. The data was obtained, which suggest changes of free-radical processes, from studied materials from women in labor in the regions most affected by the consequences of tests. The activity of erythrocytic superoxide dismutase was decreased, thus suggesting the formation of structural-functional defects of the erythrocytes. The data corresponds to the results obtained earlier when studying free-radical processes in the venous blood samples from female residents of the Altai Territory. PMID- 10520289 TI - [The precision parameters of visual perception]. AB - Here we demonstrate that retina resolution is a system parameter, depending not only on the size of receptive fields (RF) of the ganglion cells (GC), but as also on their number and the picture complexity. The curve of visual perception accuracy as a function of these three variables is nonmonotone with pronounced extremum corresponding to the RF size. The system approach allows us to reconsider the accuracy of picture approximation by various retina zones, while its application to the on- and off-subsystems of GC reveals a new function of the lateral geniculate bodies (LGB) which is to processes these afferents. We have deciphered "abnormal" alternation of ipsi- and contralateral projections in the six LGB layers. Neural network simulation demonstrated that LGBs improve the reception accuracy; being transparent for accurate reception they screen out indistinct reception and block its access to the visual cortex. Resumption and refinement of the screened information is related to eye movements and to superior colliculi (SC) assuming that SC inverses LGB function, allocating the map of indistinct pixels (image contours) by processing the on- and off-afferens. PMID- 10520290 TI - [The factors influencing the completeness of skin regeneration in mammals]. AB - In the majority of experimental studies on skin regeneration, which were conducted on the backs of rats and mice, wound healing resulted in the formation of epithelialized scars of the connective tissue. Using various methods to influence the process of skin regeneration in this and other models, we were able to obtain in some cases the organotypic regeneration accompanied by the formation of skin derivatives. We conclude that we have detected factors affecting the extent of posttraumatic skin regeneration in mammals. PMID- 10520291 TI - [A comparative study of the hemolytic activity of triterpene polyols of the dammarane series]. AB - We studied membranotropic effect of dammarane-type triterpenes (isolated from birch leaves) with a variable side chain structure, as well as the number and configuration of OH groups as a function of temperature and pH. Polyols with an acyclic side chain and four OH groups proved to have a higher hemolytic activity as compared to polyols with three or five hydroxyl groups. The triterpenes, with tetrahydrofurane cycle as the side chain, exert a hemolytic effect at a more acidic pH (5.0) and higher temperature (45 degrees C), as compared to the polyols with acyclic side chain. Irrespective of the side chain structure, the compounds with a 24(S) configuration lysed the erythrocytes at lower rate, as compared to the compounds with a 24(R) configuration. PMID- 10520293 TI - [Body-related locus of control in healthy and chronically ill adolescents]. AB - The present contribution analyses body-related locus of control beliefs in healthy and chronically ill adolescents. Moreover, we asked which bearing locus of control beliefs have on psychosocial adaptation. We examined a sample of asthmatic and diabetic adolescents and a sample of healthy adolescents aged 12 to 16 using the KLC-questionnaire. The questionnaire measures the locus of control concerning health, appearance and physical ability. Regarding the structure of body-related locus of control beliefs, the results show no differences between healthy and chronically ill adolescents; but the ill adolescents show less external locus of control. In the sample of chronically ill adolescents the locus of control is independent of severity and duration of their illness. Significant correlations between external locus of control and psychosocial adaptation could only be found in healthy adolescents. The results are discussed based on illness specific experiences and general characteristics of adolescent body image. PMID- 10520294 TI - [Sexual violence in the reality of life in mentally handicapped young girls and women--results of a research project]. AB - Subject of the study is the situation of 12- to 25-year-old girls and women from Berlin with a mental handicap who have experienced sexualised violence. 367 residential institutions of the Behindertenhilfe Berlin (aid for the handicapped) as well as 15 selected advice and consultancy facilities with amongst others a range of offers on problems of sexual violence. The return from the residencies was 68.7%. The gained individual data for the first time show the belonging to a sex for the residents. The group that was relevant for the study--altogether 147 girls and young women--was cared for in 31 institutions. The heads of the institutions provided information about more than 116 of these inhabitants. Every third to fourth resident (32 out of 116) was concerned by sexualised violence. This high extend of experience of sexualised violence in mentally handicapped female residents is directly opposed to an utterly insufficient psychosocial care situation. On one side the access to the questioned advice and consultancy centres is considerably limited for this clientele. Furthermore, heads of institutions of the Behindertenhilfe rather fall back on experts with medical psychiatric knowledge, but not necessarily with knowledge of the handling of problems of sexual violence. Suggestions for the necessary improvement of the care situation of concerned girls and women with a mental handicap are being made. PMID- 10520292 TI - [The situation of young people in Germany--results of a representative population survey]. AB - Youth and adolescence is one of great periods of upheaval during a human lifetime. Separation from parents and forming a stable individual identity are major themes. The present study outlines results relevant to this issue taken from three representative surveys of German population. The average results for various psycho-social parameters applying to young people in the country as a whole reflect a well-integrated, relatively contented and stable generation. Differentiating these results according to place of residence (East vs. West Germany) demonstrates, firstly, that psycho-social gap between East and West is closing faster among the younger generation than among the older sections of the population and, secondly, that East German youth is particularly burdened by special problems. The diminishing role of family as a stabilizing "counterpole" in the face of increasing objective difficulties and the possible consequences of this trend for young people is discussed in connection with the findings. PMID- 10520295 TI - [The water impermeability of discontinuous animal integuments]. AB - Discontinuous integuments formed by macroscopic discrete elements are characters mainly of land animals, both vertebrates and invertebrates. Waterproof properties of these integuments manifest themselves in capture and preservation of some air in their layers during submergence. The functions of integument air layer are various in different groups of animals, viz. participation in respiration (invertebrates with integuments in the form of plastron), insulation (especially birds and mammals), regulation of pelage state in the water (mammals). The principal condition of watertightness is high density of integument elements forming a system of capillaries with variable cross-sections. The general physical principle underlying the watertightness of integuments consist in a effect of surface tension arresting the water penetration deep into the coats soon as meniscus of a liquid has attend the widening section of intraintegumental capillary. For the watertightness of plastron formed by a single hair layer considerable stiffness of the whole system and high values of capillary pressure are important. Multilayered pliable mammalian pelage is characterized by a rather low capillary pressure because its effect is supplemented by the pressure that arise in the pelage air layer as a result of its compression during submergence. In birds with their structurally complicated integuments and diverse locomotory adaptations a wide spectrum of variability of plumage watertightness is observed. PMID- 10520296 TI - [The ecology of communities of related animal species (research approaches and methods exemplified by terrestrial vertebrates)]. AB - The review is dedicated to some old and modern problems in studying of the assemblages of phylogenetically related species among terrestrial vertebrates. The classical approach by Robert Mac-Arthur, discussed elsewhere summarized briefly. Although an explanative potential of competition theory seems exhausted there is no new paradigm that can simply explain species diversity in natural communities. From the beginning of 90's community ecology became very complex and controversial discipline. At the same time it is still a pioneering science. We have not enough comparative data on geo2 graphically distinct communities studied under the unified methods of data collection and analysis. Some recent results of intercontinental comparisons signify to the high degree of individualism in structure of geographically isolated communities. This makes doubtful any extrapolation from one region to another. Examination and explanation of processes is still the most contradictory field in community ecology. Confusion of causes and effects is continuing to be serious methodological problem. Much of attention was paid recently to the macroecological approach. A desire for reconsideration of some "old rules" of biodiversity is explained by advances in paleozoology, biogeography and systematic, as well as by some unsatisfactory explanations within the framework of competition theory. PMID- 10520297 TI - [The determination of the species classification of Baikal planarian cocoons found in the stomach of the black grayling (Thymallus arcticus baicalensis) by a comparative analysis of the nucleotide sequences of the ribosomal RNA gene]. AB - Comparative analysis of nucleotide sequences of gene 18S of ribosome RNA was carried out. The results show that the genetic sequences of the given locus could be used as a molecular marker to identify the species of planaria irrespective of ontogenetic stage. The articles deals with problem of specific determination of cocoons of Baikal planaria from the stomach of Baikal black grayling using comparative analysis of nucleotide sequences of ribosome RNA fragments with known sequences determined earlier for Baikal planaria. The cocoons belong to two species of Rimacephalus. The authors discuss also the importance of feeding relationships of planaria and benthophage fish to investigate the biotic factors that influence the evolution of Baikal planaria. PMID- 10520298 TI - Synthesis and antimicrobial activity in vitro of new amino acids and peptides containing thiazole and oxazole moieties. AB - 2-(Pyrrolidinyl)thiazole-4-carboxylic acid 5d, 2-(1-aminoalkyl)thiazole-4 carboxamides and hydrazides 8, 10 have been synthesized using alanine, valine, and proline as educts. In addition oxazole amino acids derived from leucine 20a and alanine 20b and some peptides 13, 14, 16 containing the 5-ring heterocyclic backbone modifications have been prepared. The thiazole and oxazole containing amino acids and peptides showed moderate antibacterial activity in vitro against various Gram-positive (Staphylococcus aureus, Bacillus cereus, etc.) and Gram negative (Escherichia coli, Protens vulgaris, etc.) bacteria, fungi (Candida albicans), and yeast (Saccharomyces cerevisae, etc.). PMID- 10520299 TI - Synthesis and inhibitory effect on platelet aggregation and antihypertensive activity of 1-hydroxy-2,5-dihydro-1H-imidazole-2-carboxylic acid 3-oxides, 1,3 dihydroxyimidazolidine-2-carboxylic acids, and 1,4-dihydroxy-2,3 piperazinediones. AB - The reaction of 1,2-hydroxylamino-oximes 1 with pyruvic or glyoxylic acid and of 1,2-bishydroxylamines 4 with glyoxylic acid resulted in 2,5-dihydro-1H-imidazole 2-carboxylic acid 3-oxides 2 and 2-imidazolidinecarboxylic acids 5, respectively. It was found that hydroxylamino acids 2 and 1,4-dihydroxy-2,3-piperazinediones 7 showed marked inhibitory effect on platelet aggregation. 1-Hydroxy-2-methyl 1,5,6,7,8,8a-hexahydro-2H,4H-cycloheptimidazole -2-carboxylic acid 3-oxide 2e exhibited antihypertensive activity. PMID- 10520300 TI - Pyrrolidine-2,4-diones with affinity to the N-methyl-D-aspartate (glycine site) receptor, Part II. 5-Arylidene-pyrrolidine-2,3,4-trione 3-oximes as NMDA receptor antagonists. AB - A series of oximes deriving from 5-arylidene-pyrrolidine-2,3,4-triones and pyridine-2,3,4-triones has been prepared. The presence of the tautomeric nitrosoenol was proven in solutions of alpha-ketooxime 7a. The binding affinity of the new oximes toward the N-methyl-D-aspartate (glycine site) receptor has been measured as a basis for more detailed structure-activity relationship studies. Oxime 13b showed the highest binding potency acting as glycine antagonist in nanomolar concentration. PMID- 10520301 TI - Synthesis and antiproliferative activity of novel 3-(indazol-3-yl)-quinazolin 4(3H)-one and 3-(indazol-3-yl)-benzotriazin-4(3H)-one derivatives. AB - Several new 3-(indazol-3-yl)-quinazolin-4(3H)-one and 3-(indazol-3-yl) benzotriazin-4(3H)-one derivatives 5 and 6 were synthesized and tested for their in vitro antiproliferative activity against Raji, K562, and K562-R cell lines. The pharmacological screening showed that some 2, 6, or 7-substituted quinazolinones 5 posses a significant antiproliferative activity, with a percentage growth inhibition ranging from 44.8% to 100% at 50 microM, which was higher than that showed by the unsubstituted derivative 5a previously synthesized. For the most active compounds 5d, 5f, and 5g the IC50 were recorded. PMID- 10520302 TI - Antiulcer effect of the N- and O-beta-D-glucopyranosides of 5-aminosalicylic acid. AB - Starting from methyl 5-nitrosalicylate (20) the N- and O-beta-glucopyranosyl derivatives (24, 28) of 5-aminosalicylic acid were prepared. The LD50 values of these compounds were determined on mice, and the inhibitory effect of 24 (0.83 mmol/kg) and 28 (1.2 mmol/kg) on gastric ulcer on rats, induced by indomethacin was investigated. PMID- 10520303 TI - Synthesis and anti-hepatitis B virus activity of some 2,3-dihydroxyprop-1-yl unnatural hetaryls. AB - The sodium salts of some hetaryls of the quinoxalin-2-ones 2-4, phthalazine-1,4 dione 5, phthalazin-1-one 6, and pyridazin-6-ones 7 and 8 were alkylated with (+/ ) 2,3-O-isopropylidene-1-O-(4-toluenesulfonyl)glycerol (1) to give the respective tetraseco-nucleosides 9-15. Their deisopropylidenation with 70% acetic acid in water gave the corresponding 2,3-dihydroxyprop-1-yl hetaryls 16-22. Compounds 16 22 showed varying inhibition activity against Hepatitis B virus (HBV) with low to moderate cytotoxicity, where 18 and 21 showed the highest replication inhibition and low cytotoxicity. PMID- 10520304 TI - [New diabetes criteria: impact on prevalence of a high risk cardiovascular population in Germany]. PMID- 10520305 TI - [Prevalence of newly diagnosed type 2 diabetes, impaired glucose tolerance and abnormal fasting glucose in a high risk population. Data from the RIAD study using new diagnostic criteria for diabetes]. AB - BACKGROUND AND OBJECTIVE: In 1997 the American Diabetes Association (ADA) introduced new criteria for the diagnosis of type 2 diabetes mellitus, reducing the upper limit of normal fasting blood sugar from 140 to 126 mg/dl. A level of between 110 and 126 < mg/dl (6.1-7.0 mmol/l) was added as a new category, "impaired fasting glucose" (IFG) as an at-risk factor. It was the aim of this study to determine what effect these new criteria will have on the prevalence of type 2 diabetes and other glucose tolerance stages in the German population. PATIENTS AND METHODS: The analysis was based on data collected in the "Risk factors in IGT for atherosclerosis and diabetes" (RIAD) study. 1139 persons of an at-risk population group (aged 40-70 years) were investigated. Most of them were relatives of diabetics and/or themselves had obesity and/or dyslipidaemia. All known diabetics were excluded. All subjects underwent an oral glucose test with 75 g glucose, and plasma glucose values were used for classifying them into different glucose tolerance stages. RESULTS: According to the new ADA criteria, confirmed by WHO in 1998, the prevalence of type 2 diabetes mellitus was shifted from 11% to 15.1%, that of IGT from 28.8% to 26.2%. An impaired fasting glucose was found in 27.1% of the population cohort, 9.2% of whom had 2-hour plasma glucose levels corresponding to the diabetes criteria. Men in the first age decade (40-49 years) had a 14% prevalence of previously undetected type 2 diabetes, double that in women. Hyperlipidaemia, hypertension and obesity were significantly more common in diabetics and persons with IGT than in normoglycaemic persons. CONCLUSIONS: The prevalence of previously unknown type 2 diabetes is very high in this at-risk part of the population. Using the criteria for impaired fasting glucose, 9.2% diabetics would remain undetected. We, therefore, support the recommendation that an oral glucose tolerance test be performed in those of the population aged over 45 years and in younger at-risk persons so that any indicated treatment can be initiated at the earliest. Measurement of fasting glucose is adequate in the population aged under 45 years not at special risk. PMID- 10520306 TI - [The percutaneous endoscopic gastrostomy catheter as a therapeutic possibility in recurrent anus praeter prolapse]. AB - HISTORY AND ADMISSION FINDINGS: A 67-year-old man with schizophrenia, well controlled by drugs, had undergone a left hemicolectomy with a terminal transverse colostomy and rectal stump closure (Hartmann's operation) for perforation of the sigmoid colon with diffuse peritonitis. 3 months later a large invagination of the transverse colon necessitated relaparotomy with further extensive resection of the colon and a new colostomy. Subsequent mild mucosal erosions were treated conservatively. He was referred to our hospital after another irreducible colon invagination through the colostomy. On admission there was an obvious, 15 cm long, prolapsing invagination of the colon with dark-blue swollen mucosa as a sign of venous obstruction. TREATMENT AND COURSE: After complicated manual reduction of the prolapsed invagination the transverse colon was fixed under coloscopy in the region of the right flexure by percutaneous endoscopic gastrostomy (PEG). Subsequently the repositioned colon portion was fixed near the colostomy with three more PEGs. These were brought out percutaneously from the intestinal lumen entirely by palpation. The four PEG tubes were removed 4 weeks later and examination after a further 4 weeks showed a good result. CONCLUSION: After endoscopic repositioning of a gastric volvulus, sigmoid volvulus or upside-down stomach, the affected organ can in certain circumstances be anatomically fixed by PEG. This minimally invasive method was successfully used by us for the first time in the repair of a prolapsed colostomy. PMID- 10520307 TI - [Abdominal actinomycosis after stomach surgery in a patient with long-term rheumatoid arthritis treated with methotrexate]. AB - HISTORY AND ADMISSION FINDINGS: A 78-year-old woman had a 30-year history of rheumatoid arthritis, of late treated with prednisolone and methotrexate. A week before admission she had first noticed a mass about 3 cm in diameter, at the lower end of a scar from a Billroth II gastric resection for gastric ulcer, performed 4 months before. She reported to have lost 6.5 kg in weight. On admission a moderately mobile, hard mass was palpated on the abdomen. INVESTIGATIONS: Ultrasound and computed tomography revealed a superficial, inhomogenous space-occupying lesion with poorly circumscribed margins. TREATMENT AND COURSE: After two days the skin over the mass became reddened and a laparotomy was performed because an incarcerated herniation was suspected. An abscess and inflammatory adhesions were found in the area of the transverse colon, histologically shown to be a chronic purulent abscess with granular clusters of pathogens indicating actinomycosis. After 3 weeks' treatment with imipenem i.v. the patient became free of symptoms, oral doxycyclin was continued for a further 6 months. CONCLUSION: Actinomycosis should be considered in the differential diagnosis of a tumour of undetermined benignity in the region of the head, chest or abdomen in immunosuppressed patients. This bacterial infection should be thought of especially if the gastrointestinal mucosa has been penetrated by invasive procedures. PMID- 10520308 TI - [Management of Amanita phalloides poisoning]. PMID- 10520309 TI - [Sodium-iodine symporter of the thyroid gland. Discovery, characterization, clinical relevance and prospects]. PMID- 10520310 TI - [Glucose infusions--a possible effective therapeutic option in ifosfamide-induced encephalopathy]. PMID- 10520311 TI - [Diagnosis of lung embolism]. PMID- 10520312 TI - [Virologic evolution in HIV-infected subjects with an undetectable viral load in the first trimester of 1997]. AB - OBJECTIVES: Assess the evolution of viral load in HIV-infected patients whose viral load was < 500 copies/ml during the first three months of 1997. PATIENTS AND METHODS: The viral load during the first three months of 1998 was recorded for comparison. Data were collected from the DM12 database implanted in the CISIH and concerned 2113 patients. RESULTS: Virological failure was observed in one third of the patients in the study cohort whose viral load was undetectable in 1997 (< 500 copies/ml) as their load was > 500 copies/ml one year later in 1998. The percentage of patients in a situation of virological failure in 1998 who were taking tritherapy in 1997 (38%) was lower than that in those who were not (47%). The clinical and immunological characteristics were however similar. The difference in the therapeutic regimen would probably explain the difference. CONCLUSION: These findings demonstrate that virological failure is an important problem. Other analyses should be conducted in other cohorts to determine the influence of non compliance or resistance to antiretroviral treatments. Future therapeutic strategies should take into account the results of such studies. PMID- 10520313 TI - [Behcet's disease and factitious manic-depressive psychosis: a case of Munchausen syndrome]. AB - BACKGROUND: Munchausen syndrome is frequently observed in men unlike other factitious conditions. The patient presents a characteristic triad: apparently acute but factitious disorders, migration from hospital to hospital resulting in unnecessary explorations and treatments, and fabulated medical history. CASE REPORT: A 57-year-old man was hospitalized in the psychiatric unit for alleged insomnia, psychomotor excitation and multiple hallucinatory phenomena. The factitious nature of the patientis condition was rapidly suspected in light of the large number of previous unconfirmed medical conditions and a rather unbelievable history. DISCUSSION: Unlike the classical description of Munchausen's syndrome, this patient had no history of surgery. This unusual aspect should not exclude the clinical diagnosis as for some patients, the invasive nature of certain explorations may be a valid substitute for surgery. PMID- 10520314 TI - [Intravascular leiomyomatosis of uterine origin. a case of pseudo-metastatic cavo cardial thrombus]. AB - BACKGROUND: Leiomyomatosis is a benign smooth muscle tumor which can provoke serious complications in case of intracaval or intracardiac extension. CASE REPORT: A 61-year-old woman had undergone hysterectomy at the age of 45 years for a hemorrhagic fibroma. She underwent surgery for infiltrative breast cancer 3 months before hospitalization and was taking tamoxifen 30 mg/day. In the cancer context, the diagnosis of cavo-cardiac metastatic thrombus was proposed but not confirmed at pathology. The diagnosis of uterine tissue intravascular leiomyomatosis was established on the basis of pathology findings and immunohistochemistry results. DISCUSSION: Five other cases of leiomyomatosis after hysterectomy have been reported in the literature. PMID- 10520315 TI - [Gingival metastasis revealing a renal adenocarcinoma]. PMID- 10520316 TI - [Isolated unilateral adrenal metastasis and simultaneous rectal adenocarcinoma]. PMID- 10520317 TI - [Hairy cell leukemia during treatment of psoriatic polyarthritis with methotrexate]. PMID- 10520318 TI - [What are the consequences of a delay in decision making after breast cancer detection?]. PMID- 10520319 TI - [40th meeting of the National French Society of Internal Medicine Strasbourg, 10, 11 and 12 June 1999. Concerns of the internist about the thyroid]. PMID- 10520320 TI - [A novel idea in continuing medical education: the English Inter-University of Medicine Diploma (DIVAM). A superfluous concern or an imperative necessity?]. PMID- 10520321 TI - [The key to the calcium regulation and calcium metabolism: G protein coupled membrane calcium receptor]. AB - CALCIUM RECEPTOR: The calcium receptor can iperceivei small vanations in extracellular calcium concentrations and adapt cell response. The icalciostati was hypothesized for several years before its discovery on parathyroid and other cells, particularly renal cells. It can couple with protein G and has 7 transmembrane domains. FUNCTION: The calcium receptor is stimulated by increased extracellular calcium level which, via a GDP/GTP exchange by the heterotrimeric protein G and a transduction network, triggers release of intracellular calcium ion stores and influx of extracellular calcium ions through the cationic channels. A FUNDAMENTAL ROLE: The calcium receptor controls not only parathormone and thyrocalcitonin secretion, but also plays an important role in calcium transfer from the mother to fetus, in reabsorption of calcium by the large ascending cortial portion of the Henle loop, and in reabsorption of water by the collecting tubes. DISEASE STATES: Mutations affecting the calcium receptor sequence cause hereditary or familial parathyroid diseases. In addition, decreased expression of the calcium receptor protein and signal has been demonstrated in primary hyperparathyroidism as well as in secondary hyperparathyroidism. The calcium receptor is also expressed in a large variety of tissues and could be implication in other disease states. THERAPEUTICS: Medical treatment of primary hyperparathyroidism using specific calcium receptor agonists derived from phenylalkylamines is now possible. The efficacy of these drugs should be demonstrated in the treatment of secondary hyperparathyroidism and chronic renal failure. Other therapeutic implications are possible. PMID- 10520322 TI - [Update on hepatitis G virus]. AB - HEPATITIS G VIRUS: The hepatitis G virus is an RNA virus with a genomic organization and variability similar to the hepatitis C virus but with a much different pathogenic power which remains to be elucidated. HEPATITIS G: No cases of chronic hepatitis have been observed. Coinfections with HCV and HIV do occur but do not aggravate the prognosis. Anti E2 antibodies are found in the serum and correspond to elimination and neutralization of the HVG genome. There is a high prevalence in transplant recipients (31%), but no modification in liver or kidney grafts has been reported. PMID- 10520324 TI - [Endoscopic view of Kaposi sarcoma of the stomach]. PMID- 10520323 TI - [Bone marrow transplantation in the treatment of autoimmune diseases. ISAMAIR Group]. AB - EXPERIMENTAL BASIS AND CLINICAL OBSERVATIONS: Remission of an autoimmune disease has been observed in certain patients after bone marrow allograft from a healthy donor. Autoimmune disease in the donor can also be transmitted to an unaffected recipient. These phenomena would be hematopoietic-dependent. BONE MARROW ALLOGRAFTS: Indications for the treatment of refractory autoimmune diseases are exceptional due to the related mortality even in patients without malignant hematologic disease. A NEW THERAPEUTIC CONCEPT: Therapeutic intensification, followed with autologous peripheral stem cell grafts, a procedure with a mortality below 3% in 1997, constitutes a therapeutic alternative in these difficult indication concerning severe refractory autoimmune diseases including: sclerodermia, vasculitis, lupus, inflammatory myositis, autoimmune cyopenia. PMID- 10520326 TI - [Medical research in Germany--present and future aspects. Roentgen Address at the 80th German Roentgen Congress in Wiesbaden 13 May 1999]. PMID- 10520325 TI - [Radiological horizons. Diagnostic radiology: image, expectations and reality]. PMID- 10520327 TI - [MR defecography at 1.5 Tesla with radial real-time imaging and reduced image field]. AB - PURPOSE: To evaluate a new technique for MR defecography with real-time imaging using radial k-space profiles. MATERIALS AND METHODS: A catheter-mounted condom was inserted into the rectum of 16 patients and filled in situ by a mixture of Nestargel and Gadolinium. After multiplanar imaging of the pelvis by high resolution T2-weighted turbo-spin echo sequences, defecation was imaged by a gradient echo sequence with radial k-space filling using a reduced field of view (rFOV) in real-time. The documentation was performed on an S-VHS recorder. RESULTS: At a constant background signal, radial k-space filling yields a real time impression. An interactive software allowed the operator to modify the slice thickness, slice plane, flip angle and slice angulation during scanning, resulting in an optimum imaging quality of the defecation. CONCLUSIONS: This new imaging technique allows real-time MR defecography in a high-field scanner and provides all anatomical und functional information of the defecation. PMID- 10520328 TI - [Malignant lymphoma: magnetic resonance tomography findings in residual supradiaphragmatic space-occupying lesions]. AB - PURPOSE: Evaluation of MR imaging in patients with Hodgkin's lymphoma and high grade non-Hodgkin's-lymphoma and mediastinal residual mass after first line chemotherapy. MATERIALS AND METHODS: MR imaging (1.5 T) was performed in 36 patients (Hodgkin's lymphoma n = 26, NHL n = 10) after first line chemotherapy). Twenty patients had inactive residual mass, 16 patients had residual lymphoproliferative lesions. T1- and T2-weighted spin echo images were visually analysed by a score index (range 1-5) as well as quantification of enhancement by signal-intensity-ratios SImax/SIplain). RESULTS: For the differentiation between residual lymphoproliferative activity and inactive residual mass, the highest accuracy was obtained for the signal intensity of residual mass on T2-w-SE compared to pectoralis muscle (94% sensitivity, 80% specificity, likelihood ratios: 4.0 [LR+]; 0.3 [LR-]). The cut-off value of the SI ratio was calculated retrospectively at 1.96 (p > 0.05). CONCLUSIONS: Differentiation between inactive (fibrotic) and lymphoproliferative (active) residual mediastinal mass is possible by MR imaging using as parameter the size reduction after therapy and the signal intensity on T2-w-SE in comparison to pectoralis muscle. Thus study suggests an additional value using the SI ratio for the differentiation. PMID- 10520329 TI - [Performance of different CT angiography imaging modalities in detecting renal artery stenoses]. AB - PURPOSE: Purpose of the study was to compare the sensitivity and specificity of various display modalities in the detection of renal artery stenosis. In particular, the difference between hard-copy reading and interactive analysis at the workstation was assessed. MATERIALS AND METHODS: Selected patients (n = 31) with expected suboptimal conditions for CT angiography due to long-standing hypertension and compromised renal function were included. Six radiologists evaluated independently a total of 77 renal arteries with 49 renal artery stenoses proven by angiography. Image analysis included: mode A: interactive display, analysis of multiplanar reformats and axial sections at the workstation, mode B: visualization of MIPs in the coronal and axial planes as hard copies only, mode C: visualization of MIPs and axial sections as hard copies only. RESULTS: The following sensitivities and specificities were found for internal readers and (readers from outside institutions): mode A: 94.8%, 87.9%, mode B: 97.7% (95.1%), 80.3% (75.4%); mode C: 97.0% (95.3%), 78.8% (76.6%). The differences were statistically not significant (p > 0.05). CONCLUSIONS: Standardized hard copies of MIPs plus axial CT provide sufficient accuracy to detect renal artery stenosis compared to interactive imaging even in this highly selected group of patients. PMID- 10520330 TI - [Angiography detection of closed palmar arcs]. AB - PURPOSE: Angiographic evaluation of the prevalence of an anastomosis between the superficial and deep palmer arcs in our patients. MATERIALS AND METHODS: Between January 1st 1991 and December 31st 1997, all selectively performed angiographies of the upper extremities were evaluated retrospectively with special focus on an anastomosis between the superficial palmer arc (SPA) and the deep palmer arc (DPA). The indications for performing a selective angiography of the upper extremity were presurgical planning of a skin/muscle transplantation (n = 42), creation of a hemodialysis fistula (n = 4) and evaluation of vascular diseases such as vasculitis (n = 9), peripheral occlusive disease (n = 3) and M. Winiwarter-Buerger (n = 8). RESULTS: 66 selective angiographies of the palm of 60 patients (39 male, 21 female, mean age 52 years old) could be used for evaluation. Angiographies of both hands were performed in 6 patients. Closed superficial palmar arcs and closed deep palmar arcs were detected in 31.8% and 84.8% of the patients, respectively. CONCLUSIONS: The rates of anastomosed superficial and deep palmar arcs in our study are comparable with those of former angiographic investigations, but they are considerably lower than the rates based on anatomical or sonographical studies. Reasons for the discrepancy are discussed. PMID- 10520331 TI - [Diagnostic value of signal-enhanced color Doppler ultrasound in space-occupying lesions of the skin and skin appendages]. AB - PURPOSE: Prospective evaluation of the diagnostic outcome of plain and signal enhanced color Doppler sonography for the differentiation between benign and malignant cutis-involving lesions. MATERIALS AND METHODS: 39 patients with 40 suspected malignant skin lesions received an preoperative ultrasound examination before and after application of 4 g Levovist i.v. Spectral Doppler, conventional color and power Doppler were performed. The images were analyzed semiquantitatively using a specially developed software. The diagnoses were confirmed by histological analysis (37 cases) or by follow up controls (3 cases). RESULTS: Whereas the B-mode criteria such as echogenoity, borderline and homogenecity, and the spectral Doppler analysis were not useful for differentiation between malignant and benign lesions, the analysis of the intratumorously visible number of vessels and of the ratio of the vascularized area using a special software, especially after injection of the signal enhancing agent, provided valuable information. In 10 of 18 benign but only in 1 of 22 malignant lesions, no intratumorous vessels were visible after application of Levovist (plain: 10/18 benign and 6/22 malignant lesions). The number of visible vessels increased strongly after signal enhancement. Using a "ratio of the vascularized area (percentage vessel area) > 5%" as a criterion of malignancy, 2 false positive and 2 false negative results were observed after application of Levovist (without/with Levovist sensitivity 73%/91%, specificity 89%/89%). CONCLUSIONS: The signal-enhanced color Doppler sonography provides useful additional information for the differentiation between benign and malignant cutis involving lesions, especially the semiquantitative vascularization analysis. The latter is superior to B-mode ultrasound and to spectral Doppler analysis. The signal enhancer increases the sensitivity, whereas the specificity remains unchanged. PMID- 10520332 TI - [Slice sensitivity profile and image pixel noise of multi-slice spiral CT in comparison with single slice spiral CT]. AB - PURPOSE: Presentation and evaluation of slice sensitivity profile and pixel noise of multi-slice CT in comparison to single-slice CT. METHODS: Slice sensitivity profiles and pixel noise of a multi-slice CT equipped with a 2D matrix detector array and of a single-slice CT were evaluated in phantom studies. RESULTS: For the single-slice CT the width of the slice sensitivity profiles increased with increasing pitch. In spite of a much higher table speed the slice sensitivity profiles of multi-slice CT were narrower and did not increase with higher pitch. Noise in single-slice CT was independent of pitch. For multi-slice CT noise increased with higher pitch and for the higher pitch decreased slightly with higher detector row collimation. CONCLUSIONS: Multi-slice CT provides superior z resolution and higher volume coverage speed. These qualities fulfill one of the prerequisites for improvement of 3D postprocessing. PMID- 10520333 TI - [Cohen's kappa or McNemar's test? A comparison of binary repeated measurements]. AB - This article intends to illustrate the combination of McNemar's significance test and Cohen's kappa coefficient in the comparison of repeated binary measurements. Both methods are standard statistical tools of major relevance for the evaluation and comparison of clinical imaging methods and thus have an impact on the corresponding publications. The interpretation of results obtainable with these methods will be illustrated to facilitate their use based on recent statistical software. Examples will further outline limitations and possible pitfalls in their application to clinical data. PMID- 10520334 TI - [Statistical analysis of German radiologic periodicals: developmental trends in the last 10 years]. AB - PURPOSE: To identify which statistical tests are applied in German radiological publications, to what extent their use has changed during the last decade, and which factors might be responsible for this development. MATERIALS AND METHODS: The major articles published in "ROFO" and "DER RADIOLOGE" during 1988, 1993 and 1998 were reviewed for statistical content. The contributions were classified by principal focus and radiological subspecialty. The methods used were assigned to descriptive, basal and advanced statistics. Sample size, significance level and power were established. The use of experts' assistance was monitored. Finally, we calculated the so-called cumulative accessibility of the publications. RESULTS: 525 contributions were found to be eligible. In 1988, 87% used descriptive statistics only, 12.5% basal, and 0.5% advanced statistics. The corresponding figures in 1993 and 1998 are 62 and 49%, 32 and 41%, and 6 and 10%, respectively. Statistical techniques were most likely to be used in research on musculoskeletal imaging and articles dedicated to MRI. Six basic categories of statistical methods account for the complete statistical analysis appearing in 90% of the articles. ROC analysis is the single most common advanced technique. Authors make increasingly use of statistical experts' opinion and programs. CONCLUSIONS: During the last decade, the use of statistical methods in German radiological journals has fundamentally improved, both quantitatively and qualitatively. Presently, advanced techniques account for 20% of the pertinent statistical tests. This development seems to be promoted by the increasing availability of statistical analysis software. PMID- 10520335 TI - [MR angiography of peripheral vessels with automatic tracking table technique at 1.0 in comparison with intra-arterial digital subtraction angiography]. AB - PURPOSE: Contrast-enhanced (CE) 3D-MR angiography of peripheral arteries was performed in 8 patients with peripheral arterial occlusive disease by applying a new tracking technique on a 1.0 T system (Magnetom Harmony, Siemens). The studies were compared with intra-arterial digital angiography as gold standard. MATERIALS AND METHODS: Imaging of the distal aorta, pelvis, upper and lower limb arteries was accomplished with a Flash-3D-sequence (TR/TE = 6.2/3.2 ms) within 26 s acquisition time of each region after a single bolus of 30 ml contrast agent. Individual circulation time was determined by a test bolus before each examination. RESULTS: 112 vessel segments were evaluated. MR angiography achieved a sensitivity of 89% and a specificity of 100% for detecting high grade stenoses and vessel occlusions. CONCLUSIONS: Tracking CE 3D-MR angiography with a 1.0 T MR imager proved to be a promising method in evaluating hemodynamically significant stenoses and occlusions of peripheral arteries. However, its definite role in the diagnostic work-up of peripheral arterial occlusive disease has to be evaluated in larger prospective studies. PMID- 10520336 TI - [MR-track pointer. A reusable device for localization during interventions]. AB - PURPOSE: Development of an helpful instrument for a better planning and orientation during MR guided interventions. MATERIAL AND METHODS: We developed a reusable, sterilizable instrument that can be filled both with a solution of Gd DTPA (1%) and with a sodium chloride solution (0.9%). The pointer has a diameter of 3 and 5 mm resp. and its length is 100-150 mm. The instrument can be combined with an interactive stereotactic tracking device. RESULTS: The MR-Track-Pointer can be fixed to the two handpieces of the integrated interactive tracking system without problems. The pointer can be seen both on T1w and T2w sequences inside and outside of the tissue. This new instrument can be interactively used for reliable planning the biopsy trajectory, planning craniotomies and identifying structures which are only visible on MR images. CONCLUSIONS: The MR-Track-Pointer is an ideal supplement for the integrated virtual tracking system. It permits a minimal invasive orientation and facilitates the exact localisation of suspect lesions in sensible regions during MR guided interventions, e.g. diagnostic biopsies and tumour resections. PMID- 10520337 TI - [A smooth pneumatic motion device for dynamic MRI imaging of the cervical spine]. AB - PURPOSE: To develop and evaluate a new, pneumatically operated motion apparatus for the dynamic functional MR examination of the cervical spine. MATERIALS AND METHODS: A metal-free, pneumatically operated motion apparatus for functional dynamic MR imaging of the cervical spine was developed with respect to the geometry of a short-bore, superconducting high-field MR system. The examination protocol included a T2-weighted multi-slice turbospin echo sequence (T2-TSE, TR/TE = 2610/115, matrix 254/512) and a dynamic T2-weighted single-shot sequence (T2-TSE, TR/TE = 1110/110, matrix 255/256, acquisition time 1 s) in a sagittal plane. In order to evaluate the new motion apparatus and the examination protocol, 10 healthy subjects and 10 patients were examined. RESULTS: The new motion apparatus allowed us to perform a passive stepless inclination and reclination motion of the patient's cervical spine within the MR scanner without leaving the magnet bore. The maximum motion degrees of the cervical spine were within -35 degrees (reclination) and 30 degrees (inclination). Due to the T2 contrast and the high spatial resolution of the dynamic sequence, the myelon, the surrounding cerebrospinal fluid and the discoligamental structures were imaged with a good contrast and allowed a sufficient evaluation of the cervical spinal channel in all functional positions. CONCLUSIONS: This new motion apparatus allows a standardized and stepless dynamic functional MR examination in a short bore high-field MR scanner. By the use of fast T2-weighted spin echo sequences it has been shown to be of high value for the evaluation of the cervical myelon, the anterior and posterior cerebrospinal fluid and the discoligamental structures. PMID- 10520338 TI - [Comparison of conventional and high resolution 2D RARE MRCP in diagnosis of pancreaticobiliary diseases]. AB - PURPOSE: Evaluation of high-resolution 2D-RARE MRCP in the diagnosis of pancreaticobiliary diseases. MATERIALS AND METHODS: 23 patients with various pancreaticobiliary diseases were evaluated using conventional and high-resolution 2D-RARE sequences. RESULTS: High-resolution RARE MRCP allowed for improved delineation of biliary and pancreatic ducts at the expense of reduced signal-to noise ratio. Pathological findings were significantly (p < 0.001) better demonstrated with high resolution RARE MRCP (score 1.65 +/- 0.54) as compared to the conventional RARE sequence (score 2.58 +/- 0.62). CONCLUSIONS: High resolution 2D-RARE-MRCP improves the delineation of biliary and pancreatic ducts. This sequence should replace the conventional 2D-RARE sequence. PMID- 10520339 TI - ["Trap door" fractures of the orbital floor]. PMID- 10520340 TI - [Rare manifestations of Wegener disease in the parotid gland]. PMID- 10520341 TI - [Mycetoma simulating, cystic fatty tissue necrosis after breast augmentation with autologous fatty tissue injection]. PMID- 10520342 TI - [Pneumatosis cystoides coli]. PMID- 10520343 TI - [Primary non-Hodgkin lymphoma of the knee joint involving the synovial membrane]. PMID- 10520344 TI - Current and future concerns for malaria in Africa. PMID- 10520345 TI - Assessment of drug resistance to the malaria parasite in residents of Kampala, Uganda. AB - OBJECTIVE: To assess drug-resistance of the malaria parasite in elite residents of Kampala city, Uganda. DESIGN: Recruited into the study were patients with complaints of fever, backache and headache or general malaise and body joint pains, could recall their previous treatment for the current complaints and could show laboratory reports indicating presence of malaria parasites. Blood was taken from those patients and examined for malaria parasites. SETTING: Kampala Diagnostic and Imaging Consultants Clinic, Kampala, Uganda from 1994 to 1997. RESULTS: Out of 268 patients, 27%, 26%, 12%, 11%, 6%, 6% and 5% strains of malaria parasites were respectively resistant to chloroquine, quinine, metakelfin, fansidar, halfan, artenam and camoquine. Double drug resistance was also observed in the patients who had taken chloroquine and quinine (21%), chloroquine and fansidar (16%) and quinine and fansidar (10%) out of 86. Some strains exhibited resistance to chloroquine, quinine and fansidar (12.6%) out of 71. R III was observed in 17 strains of malaria parasites, eight of them were for chloroquine, four for fansidar and three for quinine. Twenty-six patients had frequent recurrence of malaria lasting for over one year. CONCLUSION: One third of Plasmodium falciparum strains by 1997 had acquired resistance to chloroquine and quinine and some were gradually acquiring multi-drug resistance, leading to frequent recurrence of malaria and use of many different types of antimalarials. PMID- 10520346 TI - Knowledge, attitudes and practices of mothers and knowledge of health workers regarding care of the newborn umbilical cord. AB - OBJECTIVE: To determine the knowledge, attitudes and practices (KAP) of mothers and the knowledge of health workers regarding care of the newborn umbilical cord. DESIGN: Cross-sectional survey. SUBJECTS: Mothers with infants less than three months of age attending well child clinics and health workers (HW) in the clinics, maternity and newborn units of public health, facilities serving an urban slum area in Nairobi, Kenya. RESULTS: Of the 307 mothers interviewed, 91% and 28% of mothers knew of the need for hygiene whilst cutting and tying the cord, respectively. Regarding postnatal cord care, 40% had good knowledge and 66% good practice. Fifty-one percent of mothers knew and 54% practised postnatal cord care for the appropriate duration of time. Seventy-nine percent of mothers were afraid of handling an unhealed cord. After multivariate analysis, the following variables showed significant independent association with good maternal KAP; increased level of education (OR 2.3, p < 0.001), living in middle class areas rather than slums (OR 1.5, p < 0.03), increased maternal age (OR 1.8, p < 0.001), acquisition of knowledge from a HW rather than from other sources (OR 1.5, p < 0.001), and living in stone/brick houses rather than mud houses (p = 0.01). Fifty per cent of HW had correct knowledge on type of postnatal cord care, and 79% had correct knowledge on duration required for the same. The knowledge of 50% on type of care was incorrect by international standards, but was in keeping with Nursing Council of Kenya teaching. CONCLUSION: Mothers had good knowledge on the need for hygiene when cutting the cord, had poor knowledge and practice in other aspects of cord care, and were afraid of handling the cord. Poor KAP was associated with young, poor mothers of low education, who had acquired their knowledge from sources other than HW. The knowledge of a large proportion of HW was incorrect and outdated. We recommend that health education on cord care be given at all levels of contact with mothers and that knowledge of all primary HW on cord care be updated. PMID- 10520348 TI - How safe is motherhood in Nigeria?: the trend of maternal mortality in a tertiary health institution. AB - OBJECTIVE: To determine the magnitude and trend of maternal mortality in Jos University Teaching Hospital, Jos, Nigeria. DESIGN: Retrospective study. SETTING: Jos University Teaching Hospital, Jos, Nigeria. SUBJECT: All women dying in pregnancy, labour and puerperium. MAIN OUTCOME MEASURES: Maternal mortality ratio, trend of maternal mortality, age, antenatal booking status, educational status, main causes of maternal death, factors contributing to maternal deaths. RESULTS: The maternal mortality ratio was 739/100,000 total deliveries and trend rose from 450/100,000 in 1990 to 1,060/100,000 total deliveries in 1994. About 33% of all maternal deaths occurred among teenagers. The risk factors for maternal deaths included adolescence, grand multiparity, illiteracy and non utilisation of antenatal services. The main causes of maternal mortality were haemorrhage (28.1%), sepsis (21.3%) and eclampsia (15.7%). The contributions of complicated induced abortion and anaesthetic deaths in this study are worthy of mention. CONCLUSION: The maternal mortality ratio is unacceptably high in Jos University Teaching Hospital more particularly because of the rising trend. Socio cultural and economic factors contributed immensely to the high maternal mortality in Jos. The objective of the World Health Organisation (WHO) to reduce maternal mortality by 50% by the year 2000 will not be achieved in this part of Nigeria. Nonetheless, improvement of the nation's economy coupled with a stable policy and provision of intrastructural facilities will assist to significantly reduce maternal mortality. PMID- 10520347 TI - Growth and development of abandoned babies in institutional care in Nairobi. AB - OBJECTIVE: To determine the pattern of growth and development of institutionalised infants and to compare the outcome with that of infants living with their biological mothers. DESIGN: A cross-sectional survey. SETTING: Seven children's homes; Kenyatta National Hospital's New Born Unit and Well Baby Clinics in Nairobi, Kenya. PARTICIPANTS: Eighty-two abandoned babies who fulfilled the selection criteria were recruited and for each abandoned baby two mothered babies matched for age and sex were selected from the well baby clinics. MAIN OUTCOME MEASURES: Anthropometric measures of weight, length, head circumference and left mid arm circumference (LUMAC) were taken and the mean values and Z scores determined to demonstrate growth pattern and nutritional status of the babies. The Revised Denver Development Screening Test (RDDST) was used to assess the development pattern of infants. RESULTS: Seventy per cent of infants were below six months old and 73% were abandoned within the first week of life. Abandoned babies were significantly thinner with the mean LUMAC of 10.8 cm versus 12.3 cm (p = 0.02) Institutionalised babies were significantly wasted (p = 0.00001) and stunted (p = 0.00001). Abandoned babies were significantly delayed in development (p < 0.0001). In all the four sectors tested for, institutionalised babies showed significant delay, p < 0.0001 in each sector. CONCLUSION: This study demonstrates that infants under institutional care have poorer growth and development compared to mothered infants. PMID- 10520349 TI - Rapid epidemiological mapping of onchocerciasis in areas of Uganda where Simulium neavei sl is the vector. AB - OBJECTIVE: To test whether Rapid Epidemiological Mapping of Onchocerciasis (REMO) was suitable for mapping of onchocerciasis in foci where S. neavei sl is the primary vector. DESIGN: Topographical maps of scale 1:250,000 were used in demarcating regions into ecotopographic divisions and zones in order to identify potential onchocerciasis endemic areas. SETTING: The study was conducted in Kabarole and Nebbi districts. High-risk communities were selected 30 km from each other, and closest to rivers where vector breeding appeared likely. Secondary and additional communities were selected 10 km and 20 km away from high-risk communities, respectively. SUBJECTS OR PARTICIPANTS: Communities were mobilised for nodule palpation. A sample of thirty males aged at least 20 years, from each community that had lived in the area for at least ten years, were randomly selected and examined. INTERVENTIONS: Individuals positive for at least one nodule were expressed in terms of Nodule Prevalence Rates (NPR) which were used to map the distribution of onchocerciasis. MAIN OUTCOME MEASURES: Coefficient of variation (CV) of Nodule Prevalence Rates between high risk secondary communities. RESULTS: In Kabarole district, the results indicated a low coefficient of variation (CV) of 75 in NPR between high risk and secondary communities while in Nebbi district, higher CV of 187.4 was attained. The less varied NPR implies that communities in Kabarole were almost equally exposed to onchocerciasis while highly varied situation in Nebbi indicated decreasing NPR with increasing distance from high-risk communities. CONCLUSION: REMO is applicable in areas where S. neavei sl is the primary vector, for identification and mapping communities requiring mass treatment with ivermectin. PMID- 10520350 TI - Familial tendency and dietary association of goitre in Gamo-Gofa, Ethiopia. AB - OBJECTIVE: To assess the familial tendency and dietary association of goitre. DESIGN: Cross-sectional study with descriptive and analytical components. SETTING: Goma-Gofa, south Ethiopia. SUBJECTS: Five hundred and ninety seven elementary school children aged 6-18 years and their biological parents. RESULTS: Prevalence of goitre was found to be 51.7% of which 21.7% was visible goitre. The mean urinary iodine extraction levels indicated adequate dietary intake of iodine by the study group. A significant association (p < 0.001) was established for familial tendency of goitre between parents and their children. Consumption of halleko (Moringa stenopetala), a leafy vegetable common in the study area, of more than two times per day was significantly (p < 0.005) associated with causation of goitre. CONCLUSION: These results strongly suggest that goitre prevalence in Gamo-Gofa, Ethiopia is due to familial tendency as well as dietary factors. PMID- 10520351 TI - HIV sero-prevalence among tuberculosis patients in Kenya. AB - OBJECTIVE: To determine HIV seroprevalence among tuberculosis patients and the burden of HIV attributable tuberculosis among notified patients in Kenya. DESIGN: A cross-sectional anonymous unlinked HIV seroprevalence survey. SETTING: Tuberculosis diagnostic clinics of the National Leprosy Tuberculosis Programme in 19 districts. SUBJECTS: One thousand nine hundred and fifty-two newly notified tuberculosis patients. INTERVENTIONS: Selection and registration of eligible subjects followed by obtaining 5 ml of full blood for haemoglobin testing and separation of serum for HIV testing by ELISA. MAIN OUTCOME MEASURES: HIV seroprevalence per district and burden of HIV attributable tuberculosis among tuberculosis patients. RESULTS: A total of 1,952 eligible patients were enrolled. The weighted seroprevalence in the sample was 40.7% (range 11.8-79.6% per district). The seroprevalence was significantly higher among females and patients with sputum-smear negative tuberculosis. Chronic diarrhoea, female sex, oral thrush and a negative sputum were independent risk factors for HIV infection. The Odds ratio for HIV infection in female tuberculosis patients aged 15-44 years, was 5.6 (95% CI 4.5-6.9) compared with ante-natal clinic attenders. The population attributable risk was 0.22 in 1994. CONCLUSION: The HIV epidemic has had a profound impact on the tuberculosis epidemic in Kenya and explains about 41% of the 94.5% increase of registered patients in the period 1990-1994 and 20% of all registered patients in 1994. Repetition of the survey with inclusion of a more representative control group from the general population may provide a more accurate estimation of the burden of HIV attributable tuberculosis. PMID- 10520352 TI - Re: Laparo-endoscopic surgery: moving towards the new millennium. PMID- 10520353 TI - Vitamin D deficiency rickets: socio-demographic and clinical risk factors in children seen at a referral hospital in Addis Ababa. AB - OBJECTIVE: To test the association between vitamin D deficiency rickets and protein-energy malnutrition in Ethiopian children. SETTING: Ethio-Swedish Children's Hospital, a tertiary health facility catering for children coming from Addis Ababa and the surrounding districts. DESIGN: A case-control study. SUBJECTS: One hundred and fifty seven children under three years of age who had vitamin D deficiency rickets constituted the cases. Controls were the same number of children matched with cases for sex and age within one month and were seen within a week of case recruitment. RESULTS: Cases and controls were similar (p < 0.5) in terms of maternal education, paternal age, paternal education and family size. Factors strongly and independently associated with rickets were underweight, nutritional status [OR = 12.7 (95% CI 4.47-11.08)], marasmus [OR = 6.3 (95% CI 2.81-19.66)], lack of exposure to sunshine [OR = 3.5 (95% CI 1.33 5.84)] and non-married maternal marital status [OR = 5.1 (95% CI 2.90-10.62)]. CONCLUSION: Protein-energy-malnutrition is strongly associated with vitamin D deficiency rickets. Intervention strategies targeting vitamin D deficiency rickets should give emphasis to children with protein energy malnutrition. Further work will be required to define the causal links between rickets and the risk factors identified in the present study. PMID- 10520354 TI - Training of surgeons in Kenya at the University of Nairobi teaching hospital. AB - OBJECTIVE: To determine the number of surgeons trained by the Department of Surgery, Faculty of Medicine, College of Health Sciences, University of Nairobi, since its inception in 1967. DESIGN: This was a retrospective (1975-1987) and prospective study (1988-1999). SETTING: Kenyatta National Hospital, a National Referral Hospital and University of Nairobi Teaching Hospital. SUBJECTS: All surgeons trained by the Department of Surgery of the University of Nairobi for postgraduate MMed in general surgery, anaesthesia, ENT surgery and ophthalmology from 1975 to March 1999. RESULTS: Two hundred and eighty-five surgeons with Master of Medicine degree were trained by the Department of Surgery of the University of Nairobi between 1975 and March 1999. They included 181 (63.51%) general surgeons; 46 (16.14%) anaesthetists; 35 (12.28%) ophthalmologists and 23 (8.07% ear, nose and throat (ENT) surgeons. One hundred and seventy-six, (61.75%) were from retrospective studies; 94 (32.98%) were from prospective studies while 15 (5.26%) were from both retrospective and prospective studies. Two hundred and thirty-two (81.40%) surgeons were Kenyans while 53 (18.60%) were foreigners. The majority, 42 (79.24%) of the foreigners were from other African countries. Thirty one (58.50%) were from neighbouring Uganda, Sudan, Tanzania, Ethiopia and Zambia. There was also one PhD in anaesthesia and one MD in urology during the same period. CONCLUSION: The University of Nairobi, Department of Surgery based at Kenyatta National Hospital has played a very significant and leading role in the training of surgeons for Kenya and even other African and foreign countries since its inception. Of the forty surgeons who constitute the academic staff in the Departments of General Surgery, Orthopaedic Surgery and Ophthalmology of the University of Nairobi, thirty-five surgeons (87.50%) have been trained by the Faculty of Medicine at KNH. PMID- 10520355 TI - Role of cortisol in the deterioration of glucose tolerance in Sudanese pregnant women. AB - OBJECTIVE: To study the state of insulin resistance which occurs in pregnancy and the possible role of cortisol in augmentation of this state. DESIGN: Case-control study. SETTING: Khartoum teaching hospital, Khartoum north hospital, Soba hospital, Ibrahim Malek hospital, maternity hospital and Fath-Elbasheer referral centre from January to August 1996. SUBJECTS: Thirty pregnant women with gestational diabetes mellitus(GDM) and thirty pregnant women with impaired glucose tolerance(IGT) were compared with thirty control pregnant women with normal glucose tolerance. RESULTS: The GDM and the IGT groups were found to have significantly higher levels of serum cortisol than that of the control group [937.2 +/- 79.4 nmol/l (mean +/- S.E.) and 794.2 +/- 60.5 vs 597.3 +/- 30.9 respectively, p < 0.0006]. CONCLUSION: Cortisol has a great role to play in the deterioration of glucose tolerance in pregnancy. PMID- 10520356 TI - Maternal age, parity and gestational age on the size of the newborn in Addis Ababa. AB - OBJECTIVE: To assess birthweight, length and head circumference of live births, and to examine the effect of maternal age, parity and gestational age on birth size of the live births. DESIGN: A prospective study. SETTING: Tikur Anbessa, Zewditu, Gandhi memorial and St. Pauls hospitals, in Addis Ababa, Ethiopia. SUBJECTS: Four thousand two hundred and six consecutive live births from July 1996 to January 1997. MAIN OUTCOME MEASURES: Weight, length and head circumference of the newborns. RESULTS: Among the 4206 consecutive live births, 4047 were singleton and 159 were twins. Two thousand one hundred and sixty (51.7%) were males and 2046 (48.6%) were females. The mean birthweight of singleton live births was 3065 +/- 465 g, with modal group of 3000-4000 g. The incidence of low birthweight was 9.1%. The mean length for all singleton live births was 48.6 +/- 3.3 cm and modal group of 46-52 cm. The mean head circumference was 34.4 + 2.9 cm; modal group 31-37 cm. The incidence of low birthweight of the newborns was significantly higher for females, younger maternal age, primiparas and pre-term babies. On the multivariate analysis, gestational age and sex of the newborn respectively had significant effects on birthweight, length and head circumference of the neonates controlling for the other variables. On the other hand parity and maternal age had significant effects only on the birthweight of the neonates. CONCLUSION: This study has provided information on the effects of some of maternal characteristics on the size, particularly length and head circumference of newborns which were not given emphasis on previous birthweight studies. We suggest proper recording and analysis of birthweight, length and head circumferences be given importance for monitoring and evaluating maternal and child health programmes. PMID- 10520357 TI - Cervico-facial necrotising fasciitis occurring with facial paralysis: case report. AB - Necrotising fasciitis is a soft tissue infection, usually polymicrobial, characterised by necrosis of fascia and subcutaneous tissue. It frequently involves the groin, abdomen and extremities, but rarely involves the cervico facial region. A case is presented of a 70-year old man who, following a futile attempt to extract a lower left first molar, developed a cervico-facial necrotising faciitis with facial nerve paralysis. Bacteriological investigations revealed the presence of Klebsiella spp and viridans streptococci. It is emphasized that early detection of this disease followed by aggressive surgical debridement and antibiotic therapy are most important. PMID- 10520358 TI - Re: A Sokoto modification of Bohler-Braun splint. PMID- 10520359 TI - African oral histoplasmosis mimicking lip carcinoma: case report. AB - A case of localised African histoplasmosis with an unusual presentation in a 56 year old Nigerian farmer is reported. The lesion presented as an ulcer clinically mimicking squamous cell carcinoma of the lower lip. An incisional biopsy and culture studies confirmed African histoplasmosis and the ulcer healed spontaneously without treatment. This case is reported to highlight the unusual location and clinical course of African histoplasmosis. PMID- 10520360 TI - Typhoid fever and the Widal test. PMID- 10520361 TI - Prevalence and clinical features of tuberculosis in Ethiopian diabetic patients. AB - OBJECTIVE: To determine the prevalence and clinical characteristics of tuberculosis (TB) in diabetic patients. DESIGN: This study was a cross-sectional survey based on the retrospective analysis of data on tuberculosis in diabetic patients. SETTINGS: The study was conducted at Endocrinology and Metabolism Unit of the Department of Internal Medicine, Faculty of Medicine, Addis Ababa, Ethiopia. SUBJECTS: Seventy-eight cases of tuberculosis among 1352 diabetic patients were included from September 1989 to 1996. MAIN OUTCOME MEASURES: Clinical evaluation, chest x-ray, acid fast bacilli (AFB) in sputum or measures tissue and histopathologic characteristic of biopsy specimens. RESULTS: Seventy eight cases of TB were identified among 1352 diabetic patients giving a prevalence of 5.8%. Among 1352 diabetic patients, 713 (52.7%) were males, 639 (43.3%) were females, 619 (45.8%) were IDDM and 733 (54.2%) were NIDDM. The mean age of the 71 TB patients whose records could be retrieved was 34.0 +/- 11.9 years, 42(59%) were males, 29 (41%) were females. Fifty-four (76.1%) were IDDM and 17(23.9%) were NIDDM, 17 of the IDDM had clinical characteristics of malnutrition-related diabetes mellitus (MRDM), 36 (56%) of 71 patients were admitted for management. The three most common symptoms of tuberculosis were fever (80.5%), sweating (80.4%) and cough (70.5%). Twenty six (36.6%) of 71 cases were positive for AFB and six (8.5%) were seropositive for HIV. Fifty-six (78.9%) had pulmonary,eight (11.2%) had extrapulmonary and seven(9.8%) had disseminated TB. Forty-eight of 53 abnormal chest x-ray showed unilateral involvement. Thirty eight of 41 (53.6%) had completed TB treatment, relapse occurred in seven (9.8%), eight (11.2%) are currently on treatment, 13 (18.3%) were lost to follow up, four (5.6%) defaulted and three (4.2%) died. The relative risk of developing TB in IDDM and NIDDM patients was being 26 times and seven times than the general population respectively. CONCLUSION: The prevalence of tuberculosis in the diabetic population is high and this warrants a prospective study to determine association between pulmonary tuberculosis and diabetes. PMID- 10520362 TI - In-vitro disk diffusion sensitivity of meropenem against bacterial pathogens in Harare. AB - OBJECTIVE: To compare the in-vitro sensitivity of meropenem with imipenem and other antibiotics against clinically significant bacteria. DESIGN: A longitudinal survey. SETTING: Department of Medical Microbiology, in a tertiary care university hospital. SUBJECTS: Specimens obtained from patients attending various clinics at tertiary care and teaching hospital in Harare. Those submitted to the Public Health Bacteriology Laboratory were analysed. MAIN OUTCOME MEASURES: Rates of resistance or susceptibility of the various bacteria to the antibiotics employed in the study. RESULTS: There was excellent in-vitro bacterial activity of meropenem against virtually all clinically significant Gram positive and Gram negative isolates when compared with other antibiotics such as imipenem, ciprofloxacin, gentamicin, penicillin, ampicillin, fusidic acid, tetracyclines, erythromycin and clindamycin (p < 0.5). All isolates of Streptococcus pyogenes, Pseudomonas aeruginosa, Enterobacteriaceae, Neisseria meningitidis were susceptible to meropenem. Meropenem showed 99% overall in-vitro sensitivity against Gram positive and Gram negative bacteria. About 80% of staphylococci were resistant to penicillin whereas at least 20-25% of S. aureus, coagulase negative staphylococci, S. pyogenes showed resistance to ampicillin, erythromycin, gentamicin, tetracycline and clindamycin. CONCLUSION: Meropenem is not included in the list of routinely tested antibiotics in our laboratory, a major tertiary laboratory in the country. As a result of the ultra-broad spectrum of activity, we recommend its inclusion in our routine antibiotic sensitivity testing and observe that there is a great potential for meropenem in the treatment of infections caused by several genera of bacteria in our environment. PMID- 10520363 TI - Immunoglobulin profile in HIV-1 infected children in Dar es Salaam. AB - OBJECTIVE: To determine immunoglobulin levels in HIV-1 seronegative and HIV-1 seropositive children at different clinical stages of HIV infection. DESIGN: Cross-sectional study. SETTING: Lugalo and Mwanayamala mother and child clinics in Dar es Salaam. SUBJECTS: Two hundred and ninety-nine children aged 18 months to five years. MAIN OUTCOME MEASURES: Estimation of immunoglobulin classes (IgG, IgA, IgM, IgD and IgE). RESULTS: Mean serum levels of all immunoglobulin classes were significantly higher (p < 0.0001) among the HIV-1 seropositive children (IgG = 22.9 g/l, IgA = 2.2 g/l, IgM 2.7 g/l, IgD 101.5 IU/ml and IgE 887.7 IU/ml) than among the HIV-1 seronegative children (IgG = 11.3 g/l, IgA = 1.0 g/l, IgM = 1.5 g/l, IgD = 27.8 UI/ml and IgE 341.3 UI/ml). The percentages of HIV-infected children with immunoglobulin concentrations above mean level were 83% for IgG, 77% for IgA, 78% for IgM, 73% for IgD and 78% for IgE. CONCLUSION: HIV seropositive children have higher levels of immunoglobulin than seronegative children. There was no correlation between the levels of immunoglobulin classes and CDC clinical staging. PMID- 10520364 TI - Prevalence of malnutrition in Kenya. AB - BACKGROUND: Prevalence of malnutrition among pre-school children can be used to determine the need for nutrition surveillance, nutritional care, or appropriate nutritional intervention programmes. Such data also indicate the target groups and where interventions are required. OBJECTIVE: To determine the at risk groups, extent and magnitude, and regional distribution of malnutrition. DESIGN: A cross sectional study. SETTING: The survey was conducted in 14 districts representative of the eight provinces of Kenya. SUBJECTS: Six thousand, four hundred and nineteen children (3294 males and 3125 females) aged six to 72 months selected using the cluster sampling technique from eight provinces were studied. MAIN OUTCOME MEASURES: Anthropometric measures of height/length and weight were used to do the assessment. RESULTS: The prevalence of stunting, wasting and underweight were 37%, 6% and 27% respectively. Stunting was highest among the 12 23 months age group (44.8%). A statistically significant difference (p = < 0.005) was found between boys and girls with regard to stunting. This difference was more remarkable when the two were stratified by age group where 29% of the boys were stunted compared to 20% of the girls. Geographically, it was found that there exists great regional disparities with a low (22.6%) in Kiambu and a high (56.5%) in Kwale districts. CONCLUSION: These results show that malnutrition is still a serious public health problem in Kenya and requires urgent attention. The problem since the first survey in 1977 shows an upward trend, suggesting deterioration over the years. Well thought out and targeted intervention programmes are long overdue. The results of this survey and others emphasize the importance of having a well established surveillance system which would ensure necessary and timely action. PMID- 10520365 TI - Histological evidence of hypertrophy and ischaemia in sigmoid volvulus among Africans. AB - OBJECTIVE: To document histological evidence of hypertrophy and ischaemia in sigmoid volvulus among Africans. DESIGN: Retrospective case series study of the histology of sigmoid volvulus over seven years with cadaveric controls. SETTING: King Edward VIII Teaching Hospital, University of Natal, Durban, South Africa. SUBJECTS: Fifty African patients with sigmoid volvulus and nine cadavers with normal sigmoid colon. RESULTS: There was hypertrophy of the submucosa, muscularis propria and nerve plexuses with features of ischaemia in the patients' specimens. Their veins were thrombosed and recanalized while mesentery and submucosa had fibrosis and vascular hyalinization. There was also hypertrophy and hyperplasia of Meissner's nerve plexus. In the autopsy study, normal African sigmoid specimen showed similar ischaemic features but specimens from the four Indian patients in the study did not have such abnormalities. CONCLUSION: We postulate that chronic ischaemia may account for postoperative anastomosis dehiscence in some cases where the resection margins involved the hypertrophic segment. The benefit of an extended resection with anastomosis being effected on the bowel with apparent normal thickness to avoid this possibility should be investigated. PMID- 10520366 TI - Knowledge on lactational amenorrhoea and contraception in Kocaeli, Turkey. AB - OBJECTIVE: To determine the knowledge of women about lactational amenorrhoea and contraceptive properties of breastfeeding. DESIGN: A prospective, randomised descriptive study. SETTING: Kocaeli University School of Medicine, Department of Obstetrics and Gynecology. SUBJECTS OR PARTICIPANTS: Nine hundred and twenty-two women in their reproductive ages. INTERVENTION: A questionnaire was filled by doctors or nurses during face to face interview. MAIN OUTCOME MEASURES: There was significantly less knowledge for the importance of frequency and duration of suckling (p < 0.0001). The education increases the knowledge of lactational amenorrhoea as a interruptus contraceptive method. RESULTS: More than fifty-three per cent of women were using one of the modern contraceptive methods, 23.86% were using natural methods and 22.78% not using any family planning method. Intrauterine devices (30.15%), coitus interuptus (21.69%) and condom (16.48%) were the most common contraceptive methods. Nearly fifty-two per cent of women were not aware of the contraceptive property of breastfeeding, 25.68% of women knew lactation had a protective effect from pregnancy, 48.16%, did not know the importance of frequency and duration of suckling on fertility reducing effect of lactation. CONCLUSION: The level of knowledge on lactational amenorrhoea and frequency of suckling was significantly low in our study, especially in the illiterate group. Since efficacy of natural family planning depends on the compliance of women, education of women about lactation is very important. Family planning programmes should be focussed on breastfeeding and type of breastfeeding practices used, especially where there are no contraceptive alternatives. PMID- 10520367 TI - Re: "Bonsai-like appearance": a unique feature of multicentric hepatoma. PMID- 10520368 TI - Experience with gynaecological laparoscopies in Wad Medani Hospital, Sudan. AB - OBJECTIVE: To assess the value and safety of laparoscopy in gynaecological practice in a tertiary care centre in Sudan to determine the magnitude of tubal disease as an aetiological factor in female infertility. DESIGN: A prospective case series study. SETTING: Department of Obstetrics and Gynaecology in a tertiary care Teaching Hospital in Sudan. SUBJECTS: Seven hundred and three women selected for laparoscopy for various reasons. MAIN OUTCOME MEASURES: Indications for laparoscopy findings and complications. RESULTS: Infertility was the main indication (94.32%). Tubal disease was diagnosed in 46.6% of all infertile women studied. The overall complication rate was 22.76 per 1000; two major complications and no death. CONCLUSION: Laparoscopy is a valuable and safe procedure and and is useful in solving patients' problems, especially infertility. Tubal disease is a major aetiological factor in female infertility. PMID- 10520369 TI - Long-term complications of inguinal hernia repair. AB - OBJECTIVE: To assess the pattern of long term complications of inguinal hernia repair. DESIGN: A prospective, descriptive study. PATIENTS: Eighty six consecutive patients who presented with symptoms and signs of long-term complications of inguinal hernia repair. SETTING: Out patient clinic of a rural hospital at Mityana in Uganda. RESULTS: A number of long term complications of inguinal hernia repair were discovered. The most frequent was recurrence of hernia (42%) followed by stitch absesses/sinuses (24.2%). Others included intestinal obstruction, faecal/urine fistulae, painful scars/neuromas, unilateral/bilateral testicular atrophy, impotence, hydrocele, multiple incision scars on same side and hypertrophic scars. Complications were more prevalent between 50-70 years. In children the majority of repairs were done below 10 years. CONCLUSION: Majority of these complications could be avoided by first investigating for the aetiology of the hernia in elective cases, use of better surgical techniques and expertise. Good follow up is essential to avert distressing complications like testicular atrophy, faecal and urine fistulae. PMID- 10520370 TI - Nerve regeneration through biodegradable gelatin conduits in mice. AB - OBJECTIVE: To test if fabricated gelatin conduits can be used to bridge nerve inter-stump gaps and support regeneration. DESIGN: Experimental laboratory study. SETTING: Department of Anatomy and Neurobiology, Graduate School of Medicine, Kyoto University, Japan. SUBJECTS: Twenty-four adult mice. INTERVENTION: Mouse's sciatic nerve was resected and both proximal and distal nerve stumps sutured into each end of a gelatin conduit, to bridge a 7-mm gap. MAIN OUTCOME MEASURES: Nerve regeneration. RESULTS: At one week post-implantation, a scaffolding fibrin matrix containing few mononuclear cells formed inside the conduit. At three weeks, a well regenerated nerve composed of myelinated and unmyelinated axons, associated Schwann cells and surrounding perineurial sheath bridged the gap. CONCLUSION: Biodegradable gelatin conduits direct and support nerve regeneration and are therefore promising tools for use in entubulization repair of nerve defects. PMID- 10520371 TI - Shift work and sleep disorder among textile mill workers in Bahir Dar, northwest Ethiopia. AB - OBJECTIVE: To assess the length and quality of sleep among shift workers at Bahir Dar textile mill. DESIGN: A cross sectional study using structured questionnaire that contained sociodemographic variables, duration of work, work schedule, number of sleeping hours, sleep disorders, and associated reasons for such disorders. SETTING: A textile mill in Bahir Dar, northwest Ethiopia. SUBJECTS: Three-hundred ninety four random sample of production workers of the mill. OUTCOME MEASURES: Sleep disorders, and the impact of external and home environment on sleep. RESULTS: The mean duration of work in the factory was 25.4 +/- 7.1 years. Ninety-seven per cent of the study population work in a rotating eight hourly shift system. The mean number of hours a worker sleeps after a worked shift was 5.1 +/- 2.3. Two hundred thirty (58.4%) claimed to experience a sleep disorder. Sleep disturbance was significantly associated with rotating shift work, external environmental noise, and working in the spinning department. CONCLUSION: The majority of the workers in Bahir Dar textile mill experienced sleep disturbances as detailed in the study methodology. PMID- 10520372 TI - The arrow-head which went through the brain. AB - An 18-year old man was admitted into hospital being fully conscious, with a thirteen centimetre long metal arrow-head entirely lodged intracranially, having entered through the right orbit. Pre- and post-operative neurological condition, treatment and investigations are described. The arrow-head was removed through a partial occipital craniectomy without any major haemorrhage. The patient not only survived the operation, but was also discharged in an astonishing improved neurological condition. PMID- 10520373 TI - Epidural Bilharzioma mansoni compressing the spinal cord: case report. AB - A case of an epidural Bilharzioma mansoni (epidural granuloma due to Schistosoma mansoni) compressing the spinal cord at T11-T12 is presented. The patient, a 20 year old African man, started complaining of recurrent back pain since 1993 and became paraparetic in 1996. The myelography showed a complete block at T12 and the CT scan showed a mass at T11-T12 compressing the spinal cord. Through a bilateral laminectomy of T 10, T11 and T12, the bilharzioma was completely removed. The histopathology and the laboratory tests confirmed the diagnosis of granuloma due to Schistosoma mansoni. The patient recovered completely and was seen last time more than one year after surgery. Not a similar case has been found in the literature and the authors presume that this is the first case ever successfully treated by surgery and chemotherapy and reported in the world literature. PMID- 10520374 TI - Post malaria cerebellar ataxia and ocular flutter: report of two cases. AB - We report two cases of post malarial cerebellar ataxia presenting with severe ocular flutter, in a female of 72 years and a male of 20 years. Both patients had falciparum malaria infection. CT scans of both patients were within normal limits for the age. Cerebellar signs as well as ocular flutter responded very well to moderate doses of prednisolone therapy. PMID- 10520375 TI - [Fundamentals and results of cytostatic chemotherapy in some of the most common solid tumors]. AB - In this review we discuss established cytostatic chemotherapies as well as developments and perspectives for the three most common tumour entities. Adjuvant and neoadjuvant therapy models for primary breast cancer are discussed, established and newer concepts in palliative care presented and open questions about high dose chemotherapy raised, which need to be settled before their routine use. Developments in the field of lung cancer therapy such as adjuvant and neoadjuvant approaches are pointed out and the importance of multimodal and interdisciplinary treatment modalities is underlined. Newer cytostatic drugs are compared with established agents. In the field of colorectal cancer several new thymidilatsynthase-inhibitors as well as drugs with different mode of action are available. With these new agents, [table: see text] cytostatic therapy of advanced stages and most probably also in the adjuvant setting could markedly be improved. PMID- 10520376 TI - Quantitative tissue carcinoembryonic antigen (T CEA) assay as a screening test for severe dysplasia in colorectal adenomas. AB - Relationship between the serum (S CEA) and the tissue (T CEA) carcinoembryonic antigen concentrations with regard to the degree of dysplasia in colorectal adenomas was investigated. Our study included 56 single or multiple colorectal adenomas in 46 patients. The measurements of T CEA concentrations were performed using the quantitative CEA-EIA method (Abbott) modified for wet tissue, obtained from heads of the adenomas. As a control point the mucosa near adenoma and the rectal mucosa were used. Our results suggest that the T CEA concentrations from the head of the adenoma demonstrate a highly significant difference between mild and severe dysplasia (P < 0.001), between mild dysplasia and invasive adenocarcinoma (P < 0.001) and a significant difference between mild and moderate dysplasia (P < 0.05). On the other hand, the S CEA concentrations corresponding to these cases showed no such differences. In conclusion, we suggest the quantitative measurement of T CEA concentrations as a screening test for severe dysplasia in colorectal adenomas. PMID- 10520377 TI - [In vitro evaluation of potentially effective gemcitabine combination therapy for exocrine pancreatic carcinoma]. AB - The aim of the present study was to define potentially synergistic gemcitabine drug combinations for the treatment of pancreatic adenocarcinoma by using a tumour-specific in vitro screening system. The anticancer drug screening system used for these experiments consisted of four different established human pancreatic adenocarcinoma cell lines (BxPc-3, Panc-1, ASPC-1, Ca-pan-1) and the Microculture Tetrazolium (MTT) Assay for quantification of cytotoxic drug effects. To define single agent activities, dose-response curves, IC 50 values, and in order to validate the test system, in a first step gemcitabine and several conventional anticancer drugs including 5-FU, cisplatin, epirubicin, and mitomycin C were tested at 10 different concentrations ranging from 0.001 to 100 micrograms/ml. The effectiveness of various gemcitabine combinations was subsequently determined by using clinically relevant in vitro drug concentrations, and was rated as synergistic, additive or subadditive according to the criteria of Aapro. Overall, a heterogeneous chemosensitivity pattern was noted within the four tested cell lines. In agreement with the known chemotherapeutic refractoriness of pancreatic cancer, major cytotoxic effects were only seen with use of rather high drug concentrations. Investigation of various drug exposure times revealed a superior antiproliferative activity of gemcitabine and the other compounds in case of prolonged incubation. During subsequent drug combination experiments, gemcitabine + cisplatin and gemcitabine + epirubicin resulted in synergistic activity in 2/4 cell lines each. As opposed to the poor activity of single agents, a > 50% growth inhibition (in vitro response) was noted in 3 and 2 cell lines, respectively. Experimental data obtained with this pancreatic cancer specific in vitro screening system suggest that dose escalation or prolonged administration of gemcitabine, as well as the combination of this drug with cisplatin or epirubicin might result in improved therapeutic results. Encouraging preliminary results obtained in phase II studies seem to support the potential clinical relevance of the described disease oriented screening system. PMID- 10520379 TI - Visualization of non-Hodgkin's lymphoma by high dosed somatostatin receptor specific scintigraphy and extended single photon emission tomography. AB - The study aimed to increase the sensitivity of somatostatin receptor (SR) specific scintigraphy for the detection of non-Hodgkin's lymphoma (NHL). Ten selected patients presenting with histologically proven NHL and with 50% to 100% bone marrow involvement were injected with 20 micrograms octreotide labeled with a mean of 254 MBq 111In. The results were recorded with a double head gamma camera by long-time SPET (60 sec per frame, 3 interval) of neck/thorax and abdomen/pelvis. To show bone marrow displacement by lymphoma cells, SPET of the same regions (15 sec per frame, 3 interval) was recorded 3 to 7 days later after i.v. administration of 0.5-1 mg monoclonal anti-granulocyte antibody (Mab 250/183) labeled with a mean of 454 MBq 99mTc. This modality showed a person related sensitivity of 70%, a lesion related sensitivity of 48% (29/60), 60% (22/37) above and 30% (7/23) below the diaphragm. The sensitivity in detecting bone marrow involvement was 10%. Only 80% bone marrow infiltration with lymphoma cells in nodular configuration was shown by In-111-octreotide scintigraphy correlating with cold lesions in the anti-granulocyte scan. There was no false positive result; the smallest lesion correctly identified by SR scintigraphy had with a diameter of 1 cm, the largest lesion missed measured 3.5 cm. In conclusion, doubling the doses of octreotide and radiolabel and extended SPET recording improved to some extent the patient related sensitivity and visualized nodular bone marrow involvement in 10% of patients. The lesion related sensitivity improved mainly above but not below the diaphragm. PMID- 10520378 TI - [Inhibition of 18F-FDG uptake in glioblastoma cells by FDG and glucose]. AB - It was the aim of the study to compare the inhibition of 18F-2-Fluor-D-deoxy glucose uptake (18F-FDG) in tumor cells by various concentrations of FDG carrier or D-glucose in an experimental model using tissue culture and positron emission tomography (PET). Glioblastoma cells in culture were incubated with 18F-FDG with and without added carrier or in presence of glucose concentrations in the range from 0-5 mmol/L. Cellular uptake of 18F-FDG was measured after 20 min. of incubation in PBS-buffer containing different sugar concentrations. The uptake was determined with a PET camera. The similarity of the kinetics of the FDG and glucose uptake are backing the hypothesis that both substrates use the same carrier system. The more intense inhibition of the 18F-uptake by FDG can be explained by the different intracellular metabolism of both substrates. The results explain the clinical experience that there is an optimal 18F-FDG uptake in the patient's tumor when the blood glucose level is as low as possible and the specific activity of 18F-FDG is very high. PMID- 10520380 TI - [Use of electrocardiographic placement control of central venous catheters in Austria]. AB - After placement of a central venous catheter the correct position of the catheter tip has to be verified. The use of intravascular ECG tracing via a guide-wire or via the saline-filled lumen of the catheter enables immediate and safe control of the position. Only if complications (e.g. pneumothorax) are suspected, further clinical and radiological diagnostics are necessary. Up to now, no data on the routine clinical use of this method are available. In April 1998, a semi structured questionnaire was sent to the 518 heads of anaesthesiological, surgical and medical departments in Austria (33% of the questionnaires were returned). The subclavian (56%) and internal jugular veins (35%) are most frequently used for catheter insertion in Austria. Verification of the catheter tip placement by ECG-guidance is used in only 8% of cases, while radiographs are performed in most cases. Uncertainty with respect to forensic consequences of using the ECG-guidance for control of the catheter tip placement and the possible necessity of an additional radiograph are the main problems seen by the heads of the departments. After placement of a central venous line, measures for the verification of the catheter tip and measures for the control of possible complications have to be considered separately. Intravascular ECG tracing is unable to detect complications. Concerning the verification of the catheter tip position many studies confirm the easy handling, relevance of results and cost savings for this method. Its use for the control and documentation of the tip location is considered a standard. In Austria the consequent use of the method would offer the chance for significant reductions of treatment costs. PMID- 10520381 TI - Dry flowmeters. PMID- 10520382 TI - Pulsed-field gel electrophoresis is a useful tool in the monitoring of methicillin-resistant Staphylococcus aureus epidemic outbreaks in the intensive care unit. AB - We wished to determine how pulsed-field gel electrophoresis may be of use in monitoring methicillin-resistant Staphylococcus aureus (MRSA) outbreaks in the intensive care unit (ICU). A retrospective epidemiological analysis was conducted. All 27 ICU patients and 11 patients from other hospital wards from whom MRSA was isolated over a one year period were included in the study. Seventeen of the 27 ICU MRSA isolates were analysed by pulsed-field gel electrophoresis for clonality and compared with the 11 other hospital isolates genotypes over the same period. During three MRSA outbreaks, five MRSA genotypes were identified in ICU whilst the same five genotypes and three additional were found in the rest of the hospital. Pulsed-field gel electrophoresis analysis was useful in identifying clonality of ICU MRSA infections and establishing that they were imported from hospital wards, rather than arising de novo in ICU. We were further able to identify clonal clusters within the unit linked by temporal and geographical proximity, suggestive of cross-infection. Pulsed-field gel electrophoresis typing might be additionally useful in tracing the source of human and/or environmental factors if a genotype were persistently identified. PMID- 10520383 TI - Estimation of alveolar deadspace fraction using arterial and end-tidal CO2: a factor analysis using a physiological simulation. AB - The alveolar deadspace as a fraction of alveolar ventilation (VDalv/VTalv), while technically difficult to measure, is an objective monitor of pulmonary disease progression and a predictor of successful weaning from mechanical ventilation. The aim of the study was to examine the relationship between the arterial to end tidal PCO2 gradient (Pa-E'CO2) and VDalv/VTalv and between (Pa-E'CO2)/PaCO2 and VDalv/VTalv using the Nottingham Physiology Simulator, an original, validated physiology simulation. The relationships were observed while pulmonary shunt, anatomical deadspace, ventilatory minute volume and metabolic rate were varied. The relationship between Pa-E'CO2 and VDalv/VTalv was non-linear and was affected significantly by all the factors except anatomical deadspace. The relationship between (Pa-E'CO2)/PaCO2 and VDalv/VTalv (best fit: VDalv VTalv = 1.135 x (Pa E'CO2)/PaCO2-0.005) during normal physiological conditions was approximately linear and less influenced by physiological variation. Shunt and anatomical deadspace caused some inaccuracy, although they are unlikely to prevent the clinical usefulness of this formula. PMID- 10520384 TI - Anaesthetic management of patients undergoing lung volume reduction surgery for treatment of severe emphysema. AB - Lung volume reduction surgery can improve lung function in patients with emphysema. We report our anaesthetic experience, problems and the physiological data of eight patients. Our aims were prevention of air trapping and air leaks, good analgesia and early recovery and mobilization. We were able to achieve these aims using pressure limited ventilation, lumbar epidural diamorphine, propofol infusions and intensive physiotherapy. Hypoxia during one-lung ventilation was the main intraoperative problem. Air leaks, infection and pulmonary hypertension were the main postoperative problems. PMID- 10520385 TI - Patient-controlled analgesia in postoperative cardiac surgery. AB - The purpose of this study was to assess, in the early postoperative period of cardiac surgery, the efficacy of patient-controlled analgesia (PCA) versus nurse administered intravenous morphine followed by oral acetaminophen with or without codeine. Patients undergoing coronary bypass and/or valvular surgery were recruited. All were under 75 years of age and were in stable angina with no ischaemic attacks within the last three months. Visual analog scores (VAS) were used for pain assessment. Pulmonary function tests were done preoperatively and measured every six hours after surgery until discharge from the intensive care unit. Patients allocated to the PCA group received morphine intravenously by a PCA Plus Micro Delivery Device for at least 48 hours. Patients entered into the nurse-administered intravenous morphine group received intravenous morphine followed by oral acetaminophen with or without codeine in 24 to 36 hours according to the clinical assessment of the critical care nurse. The data showed that the quality of pain control and pulmonary function were comparable in both groups. The equipotent morphine dosage requirements were also not statistically different. It was concluded that there was no significant advantage in using PCA routinely in the early postoperative period after cardiac surgery. Furthermore, repetition of PCA instructions was often required during the study period. PMID- 10520386 TI - Prevention of postoperative nausea and vomiting in gynaecological laparotomies: a comparison of tropisetron and ondansetron. AB - In a randomized, double-blind study, the antiemetic efficacy of a single bolus of tropisetron 5 mg (group T, 37 patients), ondansetron 4 mg (group O, 39 patients) or saline (group C, 45 patients) given at induction was compared in a homogeneous group of 121 patients undergoing gynaecological laparotomy and receiving postoperative patient-controlled intravenous morphine for 24 to 48 hours. Fewer group T and group O patients developed severe nausea compared to group C (P < 0.01, log rank test in Kaplan-Meier analysis). Group T patients also had lower nausea scores than group O at 8 to 16h (P < 0.05). The overall incidences of severe nausea in groups T, O, and C were 5.4%, 17.9%, and 44.4% respectively (P < 0.001, group T vs group C; P < 0.05 group O vs group C). In conclusion, the 5 hydroxytryptamine 3 receptor antagonists tropisetron and ondansetron were superior to placebo in preventing PONV. PMID- 10520387 TI - The haemodynamic effects of propofol in combination with ephedrine in elderly patients (ASA groups 3 and 4). AB - The marked vasodilator and negative inotropic effects of propofol are disadvantages in frail elderly patients. We investigated the safety and efficacy of adding different doses of ephedrine to propofol in order to obtund the hypotensive response. The haemodynamic effects of adding 15, 20 or 25 mg of ephedrine to 200 mg of propofol were compared to control in 40 ASA 3/4 patients over 60 years presenting for genito-urinary surgery. The addition of ephedrine to propofol appears to be an effective method of obtunding the hypotensive response to propofol at all doses used in this study. However, marked tachycardia associated with the use of ephedrine in combination with propofol occurred in the majority of patients, occasionally reaching high levels in individual patients. Due to the risk of this tachycardia inducing myocardial ischemia, we would not recommend the use in elderly patients of any of the ephedrine/propofol/mixtures studied. PMID- 10520388 TI - Reduction of preoperative investigations with the introduction of an anaesthetist led preoperative assessment clinic. AB - Preoperative investigations, when used to screen for disease not clinically evident, have been shown to be unnecessary. The aim of this study was to rationalize the ordering of preoperative investigations by introducing guidelines and screening all investigations ordered at a new Day of Surgery Admissions clinic. Two hundred and one elective general and ear, nose and throat (ENT) patients attending this clinic at Sir Charles Gairdner Hospital from July to September 1997 were induced in a prospective study group. These were compared to a retrospective control group of 168 elective general and ENT surgical patients who had been admitted for surgery during May to July 1996. Patient demographics were similar for both groups. There were also similar proportions of each surgical subtype and degrees of surgical complexity in each group. There were significant reductions in most types of investigations (electrocardiogram, chest X-ray, liver function tests, urea and electrolytes, full blood examination, coagulation profile) ordered with the Day of Surgery Admissions clinic intervention. This resulted in an estimated reduction of preoperative investigation costs by 38%. It was concluded that the clinic intervention was associated with a reduction in indiscriminate preoperative investigation ordering patterns. PMID- 10520389 TI - The effect of single dose intravenous dexamethasone in tonsillectomy in children. AB - A prospective, randomized, double-blinded, placebo-controlled clinical trial was conducted in 41 patients evaluating the effect of a single preoperative dose of intravenous dexamethasone on postoperative vomiting and pain in children undergoing elective tonsillectomy. Dexamethasone was found to significantly reduce the incidence of vomiting in the first 24 hours postoperatively (P = 0.02), the time to first intake of solids (P = 0.001), the need to administer a rescue antiemetic (P = 0.005) and intravenous fluid therapy requirements (P = 0.006) in the postoperative period. No significant difference was found between the dexamethasone and placebo groups in the time to first intake of fluids, pain scores or analgesic requirement postoperatively. These results indicate that dexamethasone substantially reduces morbidity after tonsillectomy in children. PMID- 10520390 TI - Bacterial contamination of propofol in the operating theatre. AB - There have been several reports of propofol becoming extrinsically contaminated with bacteria. These reports have usually related to infusions or delays in administration after the ampoule has been opened. This observational study was performed to examine bacterial contamination of propofol during usual practice in the operating theatres of a single large hospital group. One hundred samples of propofol were collected and cultured. Samples were taken immediately after administration in cases where the delay between opening the ampoule and administration was at least 15 minutes. The samples were classified according to whether the propofol was kept in the ampoule or a syringe after opening the ampoule and whether the intended use was for a single patient or multiple patients. The time between opening the ampoule and administration was recorded. There were three positive bacterial cultures. These samples all came from ampoules used for more than one patient, without the later dose (does) being drawn into a syringe at the time the ampoule was opened. This common clinical practice, especially in paediatric anaesthesia, does not comply with the manufacturer's recommendations. The clinical significance of the bacterial contamination detected is not clear. It is recommended that propofol should be handled in an aseptic fashion and measures taken to minimize the risk of bacterial contamination. PMID- 10520391 TI - Ruptured abdominal aortic aneurysm--outcome in a community teaching hospital intensive care unit. AB - Ruptured abdominal aortic aneurysm (RAAA) is a surgical emergency associated with a high mortality often requiring postoperative intensive care. Our objectives were to assess the outcome of RAAA management in a nontertiary community hospital intensive care unit (ICU) and to compare this with historical data from tertiary hospitals. We also sought to identify variables related to outcome and evaluate the potential of an organ failure score to identify patients at increased risk of death. The study was a retrospective chart review of patients with RAAA over 11 years (1986-1996 inclusive) at Manly District Hospital, a 210 bed community teaching hospital with eight intensive care beds. Forty patients were identified in the study period as having been admitted to ICU after RAAA surgery. There was an overall hospital mortality rate of 47.5% and intensive care mortality rate of 42.5% for successfully operated RAAA. Five variables were significantly different between survivors and non-survivors. These were age, total amount of blood products required, duration of operation, development of hypotension (systolic blood pressure < 90 mmHg) in ICU postoperatively, and APACHE II score at Day 1 ICU. A trend was also found between mortality rate and the number of failed systems after 48 hours intensive care stay. Mortality for a patient with zero failed systems was 38%, one failed system 42%, two 58% and three 67%. Based on these results, management of RAAA in a non-tertiary setting appears appropriate with postoperative care occurring in an ICU where there is adequate equipment and medical and nursing staff experienced in the care of complex critical illness. PMID- 10520392 TI - Anaesthesia and postoperative pain management for bilateral lung volume reduction surgery. AB - Bilateral lung volume reduction surgery was introduced into Australia in 1995 for treatment of selected patients with emphysema. We present our experience of the anaesthetic management of our first 55 cases and describe factors associated with outcome. There were four postoperative deaths (7%). Mean (SD) total operation time was 231 (72) minutes. Median intensive care unit (ICU) stay was 26 hours. There was a significant improvement in postoperative lung function (FEV1, VC, 6 minute walk test, all P < 0.001). Eight patients (15%) required reintubation for respiratory failure; three of these patients subsequently died. With multivariate analysis, total operation time was the only significant predictor of length of ICU stay R2 = 0.25, P = 0.001), which itself was the only significant predictor of hospital stay duration (R2 = 0.36, P < 0.001). PMID- 10520393 TI - Supplementary oxygen and the laryngeal mask airway--evaluation of a heat-and moisture exchanger. AB - Heat-and-moisture exchangers (HMEs) are routinely used in anaesthesia for the humidification and warming of inspired gases. The use of the Laryngeal Mask Airway (LMA) is widespread, and many anaesthetists choose to leave it in situ until the patient regains consciousness. In this study, we have investigated the effectiveness of an HME as a means of administering supplemental oxygen via LMA in the immediate postoperative period. Oxygen was administered at varying flow rates via the gas sampling port of the HME and the oxygen delivery recorded, using end-tidal oxygraphy as a measure of alveolar PO2. At an oxygen flow rate of 4 l.min-1, the HME provided a mean end-tidal oxygen concentration of 36.2% (+/- 6.2), which compares favourably to other previously described devices. The HME thus represents a convenient, effective and economical means of oxygen supplementation via the LMA. PMID- 10520395 TI - Rupert Walter Hornabrook--Australia's first full-time anaesthetist. AB - Rupert Walter Hornabrook, born in 1871, was the first physician in Australia to devote his medical practice solely to anaesthesia. He was appointed to the Royal Melbourne Hospital in 1909, after an adventurous earlier career which ranged from service in plague hospitals in India to the Boer War in South Africa. Hornabrook was a colourful character whose work in anaesthesia included popularizing the use of the ethyl chloride-ether sequence in the early 1900s, when chloroform and closed inhalers were the norm. He was also a vigorous campaigner for the recognition of anaesthesia as a branch of medicine in its own right. PMID- 10520394 TI - The effect of devices used to reduce the risk of blood spillage or needlestick injury on the flow of intravenous infusion systems. AB - We evaluated the effect of two devices used to reduce needlestick injury and blood spillage on the flow of saline, polygeline and blood through intravenous infusion equipment and their effects on methods of increasing flow. The devices studied all reduced flow compared with control. The reductions were less for the reflux valve (< or = 9%) and greater for the anaesthesia extension set (< or = 59%), with little further reduction in flow when both were used in series (< or = 60%). Reductions in flow increased with increasing viscosity of the fluid infused, being greatest with blood. The flow reduction produced by the reflux valve was more than compensated by increasing pressure or increasing to the next larger cannula size. The flow reduction produced by the anaesthesia extension was compensated by increasing pressure but not by increasing cannula size. PMID- 10520396 TI - Combined spinal-epidural analgesia in the management of labouring parturients with mitral stenosis. AB - We report the use of combined spinal-epidural analgesia during labour in three parturients with moderately severe mitral stenosis. In each case, rapid analgesia was achieved using intrathecal fentanyl 25 micrograms without major haemodynamic changes. Maintenance analgesia was then established gradually using a dilute epidural infusion of bupivacaine 0.1% and fentanyl 0.0002%, with the avoidance of large or rapid boluses of local anaesthetic. Supplementary analgesia in the latter stages of labour was provided using slow epidural boluses of fentanyl, with or without a low concentration of bupivacaine, which was sufficient to allow controlled instrumental deliveries. We conclude that combined spinal-epidural analgesia is a useful technique for providing analgesia and maintaining haemodynamic stability in parturients with mitral stenosis. PMID- 10520397 TI - Remifentanil in emergency caesarean section in pre-eclampsia complicated by thrombocytopenia and abnormal liver function. AB - We describe the use of remifentanil in a woman with severe pre-eclampsia who presented for emergency caesarean section. Remifentanil was effective in obtunding the hypertensive response to laryngoscopy and intubation. Previous studies have found no significant adverse effects of remifentanil on the neonate. With its short duration of action, the use of this new opioid has several potential advantages in the above setting. Further studies are required to explore the use of remifentanil as an adjunct to obstetric general anaesthesia. PMID- 10520398 TI - Massive transfusion and hyperkalaemic cardiac arrest in craniofacial surgery in a child. AB - Hyperkalaemia is a recognised complication of massive blood transfusion. We present a case of hyperkalaemic cardiac arrest in a male infant of 12 months, who was undergoing craniofacial surgery for sagittal craniosynostosis. At the time of arrest the patient had received a massive transfusion of predominantly irradiated packed red cells over a two-hour period, and had a measured plasma potassium concentration of 10.1 mmol/l. Cardiopulmonary resuscitation was successful after 15 minutes. On the basis of our laboratory data and a review of the available literature, we recommend the use of fresh, non-irradiated packed red cells whenever possible in paediatric surgery. PMID- 10520399 TI - Caesarean section in a patient with paramyotonia congenita. AB - This case report details spinal anaesthesia for an elective caesarean section in a patient with the rare condition of paramyotonia congenita. There are few case reports of anaesthesia in this condition and none in the Australian anaesthetic literature. This case highlights the need for the avoidance of hypothermia and depolarizing muscle relaxants, the safety of spinal anaesthesia and a conservative approach to the management of plasma potassium concentration. The subsequent review outlines the current literature and discusses other issues involved in the anaesthetic management of this disorder. PMID- 10520400 TI - Anaphylactic reaction to aprotinin. PMID- 10520401 TI - Effects of midazolam--lest we forget: amnesia! PMID- 10520402 TI - Propofol abuse. PMID- 10520403 TI - Laerdal mask leak. PMID- 10520404 TI - [The effectiveness of oral cyproterone acetate in combination with ethinylestradiol in acne tarda of the facial type]. AB - BACKGROUND: Cyproterone acetate is a potent antiandrogen with strong progestational activity. In combination with ethinyloestradiol, it has been shown to be of clinical benefit to women displaying signs of androgenization, such as acne, seborrhoea and hirsutism. In addition, this combined oestrogen/progestogen preparation provides effective contraceptive protection. OBJECTIVE: The aim of this open-label, multicentre study was to determine the efficacy and tolerability of a preparation containing cyproterone acetate and ethinyloestradiol in women with various grades of facial acne. PATIENTS: A total of 890 women aged from 15 to 50 years with grade I-IV facial acne according to the classification of Plewig and Kligman, participated in the study. METHODS: Patients received six cycles of a preparation containing 2 mg cyproterone acetate and 0.035 mg ethinyloestradiol (EE/CPA). Changes from baseline counts of individual acne lesions (open and closed comedones, papules, pustules, nodes and cysts) were monitored together with the simultaneous presence of seborrhoea and hirsutic signs. Reductions in the number of all and individual lesions were classified as: 75-100%, "very good"; 50-75%, "good"; 25-50%, "moderate"; < or = 25%, "absence of therapeutic effect". Body weight and blood pressure were recorded throughout the study. Adverse events and reasons for premature withdrawal from treatment were documented. RESULTS: A "good" or "very good" therapeutic response (i.e. reduction in the total lesion count of > 50%) was seen in 82.8% (95% CI 80.1-85.5) of patients after six cycles of EE/CPA. A similar response was achieved in the different grades of acne with a significant decrease in the number of all lesions throughout the study (p < 0.05). A greater than 50% reduction in open and closed comedones, papules, pustules, nodes and cysts was observed in 75.6%, 80.0%, 88.4% and 85.1% of patients, respectively. By the end of the study, 64.3% of women displayed a lower grade of acne and only 4.9% experienced exacerbation of the condition. EE/CPA was well tolerated, with only 3.4% of patients discontinuing treatment because of adverse events. The majority of events were described as mild, and their incidence declined as the study progressed. No clinically significant changes in blood pressure and body weight were recorded. CONCLUSION: EE/CPA is an effective treatment for acne of all grades and all types of lesion. The preparation is well tolerated and would appear to be an appropriate treatment for women with these symptoms of androgenization. PMID- 10520406 TI - [Genes, dyslipoproteinemias and atherosclerosis: from the experimental to the therapeutic]. PMID- 10520405 TI - [Metabolic and trophic role of catecholamines in the development of white adipose tissue]. AB - The fat cell is of key significance to the physiologist investigating the mechanisms controlling lipid storage, mobilization and utilization as well as other functions of the adipose tissue. Insulin and catecholamines are the major hormonal regulators of lipolysis. Four adrenoceptor subtypes are involved in the adrenergic regulation of fat cell lipolysis. The control of adenylyl cyclase activity involves stimulatory beta 1-, beta 2- and beta 3-adrenergic receptors and inhibitory alpha 2-adrenergic receptors. Their control of lipolysis is subjected to variations according to the anatomical localization of adipose tissue deposits. In humans, lipolysis differs in visceral and subcutaneous deposits. Changes in beta- and alpha 2-adrenoceptor ratios and function have been proposed to explain the lipolytic disturbances. Human and rodent white adipocytes differ dramatically with respect to the balance between alpha 2 and b-adrenergic receptors. Human adipocytes express mainly alpha 2 and few b3-adrenergic receptors while the reverse is true for rodent adipocytes. Preadipocyte alpha 2 adrenergic receptor stimulation initiates proliferation mediated by MAPkinase activation and cytoskeleton re-arrangements. We have generated transgenic mice on a b3-adrenergic receptor gene knock-out background which express human alpha 2 adrenergic receptors selectively in white and brown fat cells by using an adipocyte-specific promoter. No phenotype was noticed in the mice fed with a standard diet, by contrast a large increase in body weight was observed when the animals are fed with a high fat diet. The weight gain concerns fat deposits and is mainly characterized by a large increase in fat cell number. This phenotype is due to an interaction between two genes and the diet since the unique combination of a high fat diet, absence of b3-adrenergic receptors and presence of alpha 2 adrenergic receptors promotes hyperplastic development of fat deposits and increased weight gain. PMID- 10520407 TI - [Insulin resistance. A public health problem]. PMID- 10520408 TI - [How to assess insulin action in man in research and clinical practice]. AB - Various methods have been proposed to assess insulin action in vivo, from the most complex to the simplest. All methods are based on the comparison of plasma concentrations of glucose and insulin, but can be differentiated by some important characteristics: evaluation in the basal state, after administration of exogenous insulin or after stimulation of insulin secretion; measurement in conditions of normo, hyper- or hypoglycaemia; and assessment using or not a modeling approach. For research purpose, the most informative techniques, such as the "euglycaemic hyperinsulinaemic clamp" or the intravenous glucose tolerance test combined with the minimal model approach, should be preferred. Easier tests may be used as alternative approaches, such as the fixed insulin-glucose infusion or the continuous infusion of glucose with model assessment (CIGMA). In daily practice, the clinician can often use simpler indices, such as fasting insulin concentrations, eventually analysed in comparison with corresponding glucose levels using the HOMA method. The only easy to perform dynamic maneuver is the short insulin tolerance test, but it is subject to several criticisms. As every approach for measuring insulin action has its own advantages and disadvantages, the selection essentially depends on studied populations (diabetic or not), primary objectives and, most importantly, available means. PMID- 10520409 TI - [Insulin resistance: therapeutic approaches]. AB - NIDDM is characterized by a decrease in insulin sensitivity of the liver, the muscles and adipocytes. Diet, exercise and control of excess body weight are the first step of the treatment; they are even able to prevent NIDDM. In this paper the drugs that may improve insulin sensitivity are described with their different specific action on liver, muscles, or adipocytes. Drugs from the thiazolidinedione class act by enhancing the sensitivity to insulin of adipose tissue; they are high-affinity ligands for peroxisome proliferator-activated receptor gamma 2 (PPAR gamma 2 being the predominant form expressed in adipocytes) Hepatotoxicity and weight gain are sides effects of thiazolidinedione. Acipimox (a nicotinic acid analogue) is a NEFA lowering drug that suppress lipolysis, but after a few days of utilisation there is a compensatory free fatty acid rise. Recent data on Metformin action on hepatic insulin sensitivity are discussed and combination with Troglitazone is presented. Vanadyl sulfate may also improve insulin sensitivity but there is no long term human studies. PMID- 10520410 TI - [Current data on iron metabolism]. AB - Iron is required for cellular life. However, abnormalities of its metabolism may lead to iron deficiency or iron overload, both conditions which are deleterious. Therefore, stock and distribution of iron in the body must be very stable. Classically, four major proteins are involved in iron metabolism: (a) transferrin which is implicated in its plasmatic transport, (b) transferrin receptor which regulates iron-transferrin uptake, (c) ferritin, the major iron storage protein, and (d) IRP (Iron Regulatory Protein) which regulates both the entry and storage of iron by linking to the IRE (Iron Responsive Element), a nucleotidic sequence found on transferrin receptor and ferritin mRNA. Thus, IRP adapts gene expression to the iron cellular status. Recent data give informations about new proteins involved in iron metabolism: HFE whose gene is mutated in genetic hemochromatosis, ceruloplasmin which permits cellular iron egress and frataxin which is implicated in the exit of iron from mitochondria. PMID- 10520411 TI - [Molecular genetics of hemochromatosis]. AB - Hemochromatosis is a recessive disorder of iron metabolism characterized by progressive iron loading of parenchymal organs, which accounts for clinical complications such as cirrhosis, diabetes mellitus, cardiopathy, endocrine dysfunctions and arthropathy. Clinical complications, which usually develop after the third or fourth decade of life, can be fatal but may be prevented by phlebotomy if iron excess is detected at a very early stage. The hemochromatosis gene (HFE), located 4.5 megabases telomeric to the HLA-A locus, encodes an HLA class I like protein and two missense mutations, C282Y and H63D in complete disequilibrium have been identified within this gene. Due to its high frequency in the general population, the involvement of H63D in the pathogenesis of the disease remains controversial, and it might correspond to a minor mutation. Conversely, the C282Y mutation is tightly linked to the disease, as it accounts for 80 to 100% of the hemochromatosis cases in Northern Europe. The lower frequency observed, in the patients, in Italy and South of France led to imagine either the implication of other mutations or of other genes. The C282Y mutation is absent in Asia and Africa and is present in the general population with a decreasing gradient of frequency from Northern to Southern Europe. The prevalence of the disease was usually estimated to be 3% but the observed frequency of the C282Y homozygotes is 5% in our breton population raising the question of the penetrance of the disease, and consequently the use of the genotypic test for its systematic screening. As HFE encodes a membrane protein similar to HLA class I protein, its contribution to iron overload is not obvious. The normal protein is predicted to to be expressed at the cell surface in association with beta 2 microglobulin, a localization for which C282Y is critical as it disrupts this association. This protein has also been shown to form a stable complex with the transferrin receptor leading to a decreased affinity for transferrin. A better knowledge of its function will help to decipher iron and different metal-ions metabolism. Although the exact role of the HFE protein is unknown, the genotypic test allows the clinicians to ascertain their diagnosis and genetic counselling. PMID- 10520412 TI - [The diagnosis of hemochromatosis in the era of the gene]. AB - The discovery of the hemochromatosis gene has deeply changed and simplified the diagnosis of the disease. In a given individual, establishing the diagnosis relies, from now on, on a simple blood sample showing the couple: elevated transferrin saturation and homozygous C282Y mutation (= C282Y +/+). Liver biopsy should only be performed when iron overload is massive in order to detect cirrhosis (or bridging fibrosis), i.e. in a prognostic view. Practically, liver biopsy is confined to the following two situations: when the C282Y +/+ patient exhibits hepatomegaly and/or an increase in serum transaminases and/or a serum ferritin level above 1,000 micrograms/L; whenever, despite a strong bio-clinical suspicion of iron overload, genetic testing does not show the expected homozygosity for C282Y. At the family level, evaluating the risk for hemochromatosis is now "instantaneous" thanks to genetic testing. One must, however, keep in mind in interpreting the data of the family members that: clinical expression of the homozygous status is not constant; heterozygosity for C282Y does not per se lead to significant iron overload, but may constitute a co factor exacerbating (or increasing the risk of) other hepatic or non hepatic diseases. Heterozygosity exposes also to the risk of homozygosity among the offspring; this knowledge of C282Y status must be balanced by the negative impact from the standpoint of possible societal genetic discrimination. PMID- 10520413 TI - [Transcription factors of the anterior pituitary and combined hypopituitarism]. AB - In the recent years, spontaneous and experimental models of hypopituitarism have underlined the involvement of a number of homeodomain transcription factors in different forms of congenital anterior pituitary hormone deficiencies. Indeed, abnormalities of the transcription factor Pit-1 are responsible for combined deficiencies affecting thyrotroph, somatotroph and lactotroph cell lineages both in dwarf mouse strains (Snell, Jackson) and in human patients. More recently, alterations of the Prop-1 gene have been shown to induce a similar phenotype in the Ames mice, and alterations of the human homolog gene have been evidenced in patients with anterior pituitary deficiencies involving the same three lineages together with the gonadotrophs. Gene knock-out experiments have demonstrated the importance of other transcription factors such as Lhx3 and Lhx4 in the development of the normal pituitary gland. These findings illustrate the potential involvement of anomalies of these and many other factors in the various forms of multiple hypopituitarism. PMID- 10520414 TI - Idiopathic growth hormone deficiency: presentation, diagnostic and treatment during childhood. AB - The clinical and biological presentation of idiopathic growth hormone (GH) deficiency (GHD) varies greatly, demonstrating the variety of its pathogenic features and explaining why it is difficult to diagnose. We examined 48 patients (26 males) with certain idiopathic GHD diagnosed at 4.8 +/- 0.7 yr. The symptoms that led to the diagnosis of GHD were low growth rate (33 cases), hypoglycemia (12 cases), microphallus (1 case) and in 2 cases the GHD was diagnosed from magnetic resonance imaging (MRI) performed for delayed mental development (1 case), or congenital blindness (1 case). The 2 other cases were diagnosed from routine GH evaluation performed at birth because of idiopathic GHD in siblings. Thirteen had congenital malformation. Twenty three cases (48%) had features suggesting that the GHD was of antenatal origin. Six of them were born by breech delivery. Twenty one cases (44%) had features suggesting a hypothalamic origin. The decrease in growth rate occurred before 0.5 year in 21 (55%), before 1 year in 27 (71%) and before 2 years in 30 (79%): 8 patients (21%) maintained a normal growth rate after this age. Among these 8 patients, 5 had signs suggesting an antenatal origin and 4 had severe episodes of hypoglycemia from birth. The mean GH peak after the pharmacological stimulation test was 3.6 +/- 0.5 micrograms/l. The mean plasma insulin-like growth factor 1 (IGFI) was 0.1 +/- 0.02 U/ml. The GH deficiency was associated with deficiencies of thyrotropin in 26 (54%) and of adrenocorticotrophic hormone in 17 (35%) patients. Among the 15 patients of pubertal age, 9 (60%) had gonadotrophin deficiency. No patient had diabetes insipidus. The MRI showed pituitary stalk interruption syndrome in 39 patients and normal pituitary anatomy in 6 patients. GH treatment reduced the difference between target and actual heights from 3.5 SD (before) to I SD (after 3 years) in the 39 more recently seen patients given 0.5-0.6 U/kg/w GH in 6 or 7 weekly injections. Height gain during the first year and cumulative height gain over 3 years (SD) was correlated negatively with height (SD) at the start of treatment (p < 0.01). We conclude that most of the patients with GHD have features suggesting an antenatal origin. Despite this early origin, the decreased growth rate may occur after 2 years. PMID- 10520415 TI - [Hormone replacement therapy in menopause and the risk of cerebrovascular accident]. AB - BACKGROUND AND PURPOSE: Hormone replacement therapy relieves climacteric symptom and prevents postmenopausal osteoporosis. A protective effect of estrogen against coronary heart disease remains debatable and inconclusive results have been reported with respect to the risk of stroke. We have therefore performed an updated quantitative assessment of the risk for stroke associated with hormone replacement therapy. METHODS: MEDLINE database was used. Studies analyzing postmenopausal women and considering any subtypes of stroke--i.e. fatal or non fatal, ischaemic or haemorrhagic--as the outcome of interest were selected. An overall estimate was calculated as a weighted average of the odds ratios or relative risks, with the weights being the reciprocal of their variance. Random effects models were used to take into account the heterogeneity of data. RESULTS: Six case-control studies, seventeen cohort studies and one randomized trial were selected between 1978 and 1998. Seven studies assessed the risk of ischaemic stroke associated with hormone replacement therapy and the pooled estimate of the risk was 1.12 (95% confidence interval, 1.01 to 1.25). The random effects model showed an increased risk of 18% (relative risk 1.18, 95% confidence interval, 0.98 to 1.43). Regarding haemorrhagic stroke, the analysis based on two case control studies and one cohort study showed a significantly reduced risk of 35%. Lastly, based on five studies, no significant change in the risk of subarachnoidal hemorrhage was found. CONCLUSION: This updated analysis suggests an increased risk for ischaemic stroke among postmenopausal women who use oral estrogen replacement therapy. No data regarding transdermal estrogen are available. PMID- 10520416 TI - [Results of international clinical trials with raloxifene]. AB - A new drug class called Selective Estrogen Receptor Modulators (SERM) could combine ideal properties for a product designed for menopausal women. The most widely studied member of this class is raloxifene which is currently marketed in several countries for the prevention of osteoporosis in menopaused women. This product is a nonsteroidal derivative of benzothiophene which, like estrogens, has a preventive effect against bone loss involving the spine and peripheral skeleton and a cholesterol lowering effect, both in the ovariectomized rat and in menopausal women. Unlike estrogens, raloxifene does not stimulate breast or uterine tissue. These interesting properties make raloxifene a possible preventive treatment for osteoporosis and other menopause-related risks for menopausal women of all ages. Multicenter studies have been conducted in recently menopausal women who received either raloxifene at the doses of 30, 60, or 150 mg/day or a placebo in a randomized protocol. All subjects were also given calcium supplementation. Bone density was measured twice a year for 36 months by dual X-rays absorptiometry and showed a significant decrease at all sites in the placebo group while there was a significant increase in the spine, the hip and the overall skeleton for all three raloxifen groups. After 24 months of treatment, mean increase over placebo was 2.4% for 60 mg raloxifene measured on the spine and total hip and 2% for the overall skeleton. Markers of bone formation (serum osteocalcin and bone alkaline phasphatase) and resorption (urinary CrossLaps) decreased significantly reaching, after 3 to 6 months of treatment, the levels observed in non menopausal women. In addition, total serum cholesterol as well as LDL-cholesterol decreased significantly in a dose dependent fashion in all groups treated with raloxifene. Serum HDL-cholesterol and triglycerides did not very significantly during treatment. Hot flashes were the most frequently observed undesirable effect, at a frequency slightly higher in the raloxifene group (25%) than in the placebo group (18%). This undesirable effect was of low intensity and generally occurred during the first months of treatment. It did not cause a higher drop out rate (raloxifen 1.5%; placebo 2.1%). The preliminary data at two years follow-up suggest that raloxifene is not associated with an increased risk of breast cancer. In conclusion, raloxifene is a particularly interesting drug for menopausal women showing very promising efficacy and clinical tolerance. PMID- 10520417 TI - Steroidogenic factor-1 (SF-1), a specific transcriptional factor of differentiation and function of steroidogenic cells. PMID- 10520418 TI - Transcription factors in developing endocrine pancreas. PMID- 10520419 TI - [Disorders of the vitamin D nuclear receptor: from children to transgenic mice]. PMID- 10520420 TI - [10 years' experience of using cultured human skin cells for the treatment of thermal burns]. AB - The authors have developed a unique method of treating extensive burns by transplantation of the cultivated cells on the affected area. The main elements of the transplant are not keratinocytes but fibroblasts, and this is a principal difference with other methods. The transplanted fibroblasts actively synthesize DNA, collagen, fibronectin, glycosaminoglycanes which comprise the extracellular matrix formed by cells. The method is simple, cost-effective, provides high percentage of fibroblast transplant survival, essential acceleration of the burn area epithelialization as compared to other methods. This method was applied in the treatment of 517 patients in various hospitals of the country. PMID- 10520421 TI - [A morphological and electron radioautographic study of the wound in the diabetic foot syndrome]. AB - Two groups of patients with a grave form of diabetes mellitus type I and II with diabetic foot syndrome have been examined. Group 1 (21 cases) consisted of patients whose foot was amputated. The cause of the necrotic process was ischemia produced by alteration of the arterial vessels aggravated by infection. Patients of group 2 (18 cases) were treated with resultant wound healing and foot preservation. The material was subdivided into two subgroups: 1) tissue from the "infectious" foot with great amount of bacteria and inflammation reaction; 2) tissue from the "ischemic" foot with considerable alterations of microvessels structure. In both groups well developed granulation tissue was observed after the treatment; almost all the cells of this tissue included 3H-uridin and labelling with 3H-thymidine was increased in fibroblasts, endotheliocytes and pericytes. PMID- 10520422 TI - [The survivability time of central nervous system cells]. AB - Our investigations show that vital functions of some neurons and glial cells may survive for 4-8 hours after death and disappear in parallel with destructive changes in cells. Up-to-date morphological techniques demonstrate that neurons and other cells of the CNS preserve biological activity not for several minutes as believed so far but up to 4-8 hours. It is too early to judge about functional importance of this phenomenon and further investigations into this problem are necessary. Temporal parameters of RNA synthesis in the brain cells give additional knowledge about CNS cells survival and this may be of interest for reanimatology and transplantology. PMID- 10520423 TI - [The identification of pericyte-like cells in the subendothelium of human blood vessels]. AB - We identified pericyte-like cells in various regions of human vascular bed using 3G5 monoclonal antibodies against brain capillary pericytes' surface antigen. Pericyte-like cells were revealed in the subendothelial layer of the arterioles, venules, brain microvascular bed and in vasa vasorum adventitia. In arteries and veins of small, medium and large caliber pericyte-like cells were found in intimal subendothelial space. Pericyte-like cells form subendothelial network and may play an important role in physiological processes of vascular wall function. PMID- 10520425 TI - [A systems pathologicoanatomical analysis of the causes of maternal mortality]. AB - Principal nosological forms of the maternal mortality are presented using terminology of the International Classification of Diseases (ICD-10), and a number of morphological markers are described facilitating establishment of primary causes of the maternal mortality. A new approach is suggested to the analysis of autopsies of women in childbirth in the context of the functional system "mother-her target organs-uteroplacental area-placenta-fetus, newborn". The primary cause of death is formulated by summation of macro- and microscopic changes in the above organs and areas. PMID- 10520426 TI - [Morphological and pathogenetic differences in the respiratory disorders of newborn infants in the first days of life based on data from the composition of tracheobronchial smears]. AB - Six variants of pulmonary pathology are described basing on the differences in composition of tracheobronchial wash-offs. This composition was studied by electron microscopy and biochemical assay of phospholipids in 76 neonates during the first day of life as well as by structural investigation of the lungs in 16 children who had died within two days after birth. The composition of the tracheobronchial wash-offs was compared with lung alterations associated with intrauterine hypoxia, infection, delay of respiratory parenchyma development, brain trauma. PMID- 10520424 TI - [Multiple primary breast cancer]. AB - Primary multiplicity (PM) of mammary carcinoma is found in 669 (39%) cases of 1690 invasive tumors. PM is more typical for invasive ductal carcinoma with predominance of the intraductal component (67%), Paget cancer (61%), invasive lobular carcinoma (55.5%) and tubular carcinoma (52%). Invasive PM develops in the presence of the corresponding background with areas of grave dysplasia and intraepithelial carcinoma. True PM does not influence the disease prognosis. PMID- 10520427 TI - [The status of the autonomic centers in hydrocephalus of different origins in fetuses and newborn infants]. AB - Developmental defects (partial atrophy, hyperplasia, hypotrophy, hypertrophy, deformation, dysplasia) in the higher vegetative centers in cases of hydrocephaly in fetuses and newborns are described. PMID- 10520428 TI - [Mucosal morphology in the latent form of terminal ileitis in children based on biopsy data]. AB - Morphological study of serial section biopsies from 22 children was made to develop new diagnostic criteria of a latent form of terminal ileitis. The following changes were observed: fibroblastic reaction around the lymphoid follicles, degenerative changes of enterocytes close to the lymphoid follicles; disruption of basal membrane over the lymphoid follicles; penetration of lymphoid cells into the intestinal lumen and their adhesion to the apical membrane of enterocytes; neuroma-like structures as an issue of productive vasculitis. The frequency of the symptoms above 62% does not depend on the sex, age and treatment method. PMID- 10520429 TI - [A "sugar" tumor of the lungs]. AB - Histological structure of a rare benign tumor ("sugar" tumor of the lung) is described. PMID- 10520430 TI - [A statistical analysis of liver diseases (based on the autopsy data from the clinics of the I. M. Sechenov Moscow Medical Academy for 1978-1987)]. AB - 6170 autopsy protocols have been analysed and the conclusion is made that alcoholic liver cirrhosis is predominating in the structure of liver diseases. Viral liver cirrhosis is less important by its incidence than alcoholic one. Liver tumors are of importance against the background of alcoholic and viral cirrhosis. Primary liver tumors, autoimmune and metabolic diseases occur rarely. The material confirms the fact that alcohol is a principal etiological factor in development of liver diseases. PMID- 10520432 TI - [The place and tasks of a pathologicoanatomical service in St. Petersburg under the conditions of public health reform]. PMID- 10520431 TI - [A morphological analysis of the results of using the capron drainage-tampon with a sorbent for the treatment of acute nonspecific metroendometritis]. AB - The surface of carbon-mineral sorbent (CMS-1) granules and that of the capron cartridge were investigated using scanning electron microscopy in the course of acute nonspecific metroendometritis treatment after late abortion. The use of this sorbent for drainage of the uterine cavity decreases a toxic and antigenic effect on the organism. The use of the capron cartridge excludes the loose of the sorbent granules and their contact with the tissues, does not hinder the drainage and, at the same time, capron serves as a filter preserving active granule surface from a rapid disactivation by large fragments of tissue detritus. PMID- 10520433 TI - [Congenital ichthyosiform erythrodermas: aspects of their etiology and pathogenesis]. AB - The term "inherited ichthyosiform erythrodermia" (IIE) designates a heterogeneous group of inherited disturbances of keratinization. Lamellar ichthyosis (LI), inherited nonbullous ichthyosiform erythrodermia, bullous inherited ichthyosiform erythrodermia (BIIE) and some other conditions. Processes of terminal cell differentiation and epidermal keratinization are affected in all the above conditions. A decrease of transglutaminase activity is observed in L1, this being connected with mutations in chromosomes 14q, 2g33-35. In BIIE which is characterized by dense groups of tonofilaments in the suprabasal layer, mutations of keratin I and X coded by chromosomes 2 and 17 are identified. When sporadic forms of the disease occur, the main question is establishment of the genotype phenotype correlation which depends on detailed characteristics of each patient and accurate study of the ultrastructural disturbances. PMID- 10520434 TI - Sexual assault prevention programs: current issues, future directions, and the potential efficacy of interventions with women. AB - Current problems facing the primary prevention of sexual assault are reviewed. Effective sexual assault prevention programs for both males and females have been slow to develop due to the fact that the etiologies of sexual assault have not been identified. Although dissemination of prevention programs has become increasingly popular in recent years, few programs have evaluated the extent to which the constructs identified in the interventions are effective at decreasing rates of sexual assault. This article discusses previous studies in sexual assault prevention programs, methodological and conceptual problems that currently exist in the field, pragmatic difficulties regarding program implementation and evaluation, and recommendations for future research with an emphasis on interventions with female participants. PMID- 10520435 TI - Evaluating recovery from anorexia nervosa and bulimia nervosa: integrating lessons learned from research and clinical practice. AB - The aims of this article are to critically examine the conceptualization and measurement of recovery, within the eating disorder outcome literature, and suggest possible ways of developing the clinical utility of outcome research. First, definitions and measures of recovery operationalized in outcome studies are critically reviewed, highlighting the variety of definitions and measures used in outcome research. Two important caveats in the outcome literature are identified: absence of clients' views on their recovery from outcome evaluations and dissociation of outcome research from negotiated interpersonal and organizational meanings of recovery. These caveats form the focus of the second section of the article. A need for greater integration between research and clinical perspectives on recovery is identified, and the final section of the article suggests several proposals for enhancing current research on recovery from eating disorders. These proposals particularly advocate development of methods and measures that can accommodate diversity of clients' experiences of recovery, while remaining informative to both researchers and clinicians. PMID- 10520436 TI - A bidirectional associative memory explanation of posttraumatic stress disorder. AB - Network theory is a relatively recent explanatory development to improve our understanding of emotions and emotional disorders. Connectionist neural networks are introduced as mathematically well-defined network theories that have been suggested as PTSD models, partly because of their ability to form and retrieve memories. Explanatory requirements that a comprehensive PTSD theory must have are reviewed. A connectionist neural network system called the bidirectional associative memory (BAM) is extended to encode emotion and cognition and is then shown to satisfy all PTSD explanatory requirements and consequently constitutes a comprehensive PTSD theory. Empirical work pertaining to a fundamental assumption that learning alters brain structure in ways that can be detected by neuroimaging is reviewed. Novel predictions are made, and relevance to neuroscience is discussed. PMID- 10520437 TI - Factors associated with caregiver burden in mental illness: a critical review of the research literature. AB - This article reviews caregiver burden studies that evaluated burden of care for a mentally ill relative using measurement instruments with established validity and reliability. The review identifies aspects of caregiving that are most burdensome to caregivers. It describes the nature of the relationships between variables and different dimensions of caregiver burden, and identifies mixed findings that are theoretically relevant to caregiver burden. The review discusses research findings in light of the methodological issues and research designs characterizing the literature, and briefly summarizes the effects of burden on the caregiver's life. Finally, it identifies advances made in this line of research in recent years and highlights areas that need further attention in future research work. Summary tables are included. PMID- 10520438 TI - Mental disorder and cross-cultural psychology: a constructivist perspective. AB - The predominant Western approach to understanding mental disorder, as indicated in the Diagnostic and Statistical Manual of Mental Disorders (DSM), is based on a biomedical perspective which sees mental disorders as "natural kinds" or discrete entities which manifest as dysfunction within individuals. Following from this is the view that the DSM's primary syndromes are universal, based on the assumption that this dysfunction is similar across diverse human populations. The cross cultural literature, however, reveals significant differences in the manifestation of these syndromes across ethnic groups, thereby challenging the universalist position. In response to this shortcoming of the predominant contemporary conceptualization of mental disorder, a constructivist approach is offered which, it is argued, has a number of important advantages over the traditional view. Finally, the implications of a constructivist definition are discussed, demonstrating the important connection between theory and practice. PMID- 10520439 TI - Cognitive-behavioral treatment for rapists: can we do better? AB - A review of treatment studies with rapists suggests that the currently used cognitive-behavioral treatment strategies remain limited in their success. The current article proposes that some reasons for the limited success may be that current treatment approaches do not adequately address the heterogeneity of the population, emphasize changing patterns of physiological arousal and cognitive distortions rather than psychological acceptance, and neglect to address differences in the function of sexually aggressive behavior among individuals. With the hope of decreasing rates of victimization and preventing recidivism by rapists, this article offers several treatment suggestions that should be tested empirically to determine if treatment efficacy can be increased with this population. PMID- 10520440 TI - Cognitive impairment in elderly who are not yet demented. AB - Patients with neuropathological changes of Alzheimer disease may not be demented during initial evaluation of memory disturbance. Understanding current issues regarding the patient with incipient degenerative dementia should help identify those at greatest risk for progression and may help delay onset of symptoms. PMID- 10520441 TI - Migraine: clinical features and diagnosis. AB - Migraine is a syndrome characterized by recurrent headaches with or without aura. Triggers include foods, hormonal changes, and stressors. Migraine must be differentiated from other unilateral headache disorders and from headaches due to other neurologic and systemic diseases. PMID- 10520442 TI - Treatment options for coronary stent restenosis. AB - Treatment of in-stent restenosis with balloon angioplasty alone is adequate for focal lesions but is associated with a 50% recurrence rate for diffuse lesions. For diffuse in-stent restenosis, debulking with atherectomy or laser can significantly reduce the recurrence rate. PMID- 10520443 TI - Orthopedic management of pyogenic arthritis. AB - Septic arthritis results from bacterial invasion of the joint cavity. Joint infection is curable if timely, appropriate treatment is rendered. Antimicrobial drugs and removal of harmful substances produced by host defenses interacting with the organism are essential for restoring joint function. PMID- 10520444 TI - Recent advances in pharmacological treatment of type 2 diabetes mellitus. AB - Several new pharmacological agents attempt to correct abnormalities in the pathogenesis of type 2 diabetes mellitus. The availability of agents with different mechanisms of action and side-effect profiles permits the design of individualized regimens that address the various pathophysiologic abnormalities. PMID- 10520445 TI - Basic therapy for rheumatoid arthritis: nonsteroidal anti-inflammatory drugs. AB - Nonsteroidal anti-inflammatory drugs (NSAIDs) are often the initial treatment for rheumatoid arthritis. NSAID-induced inhibition of cyclooxygenase-2 (COX2) and cyclooxygenase-1 appears to correlate with clinical efficacy and toxicity, respectively. Newer NSAIDs with greater COX2 selectivity offer the promise of less toxic therapy. PMID- 10520446 TI - Role of restenosis in determining adverse outcome after coronary angioplasty vs bypass surgery. AB - A group of patients who did not have restenosis after percutaneous transluminal coronary angioplasty (PTCA) was compared with a group receiving coronary artery bypass grafting (CABG). Restenosis only partially accounted for the greater morbidity after PTCA compared with CABG. PMID- 10520447 TI - Carotid artery stenting: rationale, indications, and results. AB - Carotid stenting is a percutaneous, minimally invasive treatment for carotid stenosis. It does not carry the surgical risks of carotid endarterectomy and therefore can be applied to the elderly and to patients with comorbidities. Complications and late events appear low. PMID- 10520448 TI - Bovine ornithine decarboxylase gene: cloning, structure and polymorphisms. AB - Bovine ornithine decarboxylase (ODC) genomic clones were isolated from a bacteriophage lambda DASH genomic library. A total of 9452 bp sequence was determined which covers the entire sequence of the bovine ODC gene. Sequence analysis showed that the bovine ODC gene consisted of 12 exons which encode a protein identical to that inferred from a bovine ODC cDNA. Comparison of the structure and nucleotide sequence of the bovine, human and mouse ODC genes revealed that the gene was highly conserved. Primer extension analysis demonstrated that the transcription start point of bovine ODC mRNA was located 378 bp upstream from the A residue in the translation initiation codon. The 5' untranslated region (UTR) of ODC mRNA was highly G + C rich, particularly in its 5'-most portion, and computer predictions suggested a very stable secondary structure for this region, with an overall free energy of formation of -134.4 kcal/mol. Conserved sequences and potential promoter elements including a TATA box, a possible CCAAT element, SP1 ranscription factor binding sites (GC boxes) and cAMP response elements (CRE) were identified in the 5'-flanking region of the gene. Two polymorphic restriction sites, a TaqI and a MspI, were mapped to the ODC gene and PCR-based methods for detection of the 2 polymorphisms were developed. PMID- 10520449 TI - Sequence analysis of the rDNA internal transcribed spacer 2 of five species of South American human malaria mosquitoes. AB - The rDNA internal transcribed spacer 2 (ITS2) was sequenced for 5 species of mosquitoes that may be important vectors of human malaria in certain regions of South America and are difficult to distinguish by morphology: Anopheles evansae, An. nuneztovari, An. rangeli, An. strodei and An. trinkae. ITS2 sequences from samples collected in Ecuador, Bolivia, Venezuela and Brazil were aligned and compared in order to determine the usefulness of this spacer for the elaboration of species specific primers and DNA probes. The ITS2 was found to be different in size (ranging from 333 to 397 bp) and sequence between all pairs of species. Highly variable regions were found primarily at the 3' end of the spacer and were interspersed with relatively conserved sites. Instraspecific sequence variation was limited to a single transversion between specimens of An. rangeli from distant geographic locations suggesting concerted evolution and homogenization of the ITS2. PMID- 10520450 TI - Type 2 rice metallothionein-like gene has two introns. AB - A type 2 rice metallothionein-like gene was isolated from root by PCR and sequenced. The PCR fragment was designated as pcr1460, which overlaps with OsMT 2, a cDNA sequence previously characterized, with the presence of two additional segments, 583 and 613 bp in length. These segments are recognized as introns which divide the coding sequence into three exons, 65, 78 and 106 base pairs in length. The sequences flanking the introns conform with the GT/AG rule for splice junctions, and one exonic open reading frame can be identified in each of the introns. The observation that MT-like gene has two introns is the first of such a finding obtained from monocotyledonous plants. PMID- 10520451 TI - Nucleotide sequence of exon 2 to 4 of the R-ras gene in the hermaphroditic fish Rivulus marmoratus. AB - We have cloned the ras homologue from the mangrove rivulus (Rivulus marmoratus) after low-stringency plaque hybridization of the mangrove rivulus genomic DNA library. This mangrove rivulus ras homologue showed a 76% amino acid homology to the human R(related)-ras gene with identical exon/intron boundary and was named the mangrove rivulus R-ras gene. The mangrove rivulus R-ras gene spanned 1.8 kb and consisted of at least 5 exons. The exon/intron boundaries coincided with the rule of GT/AG of consensus splice acceptor and donor sequences. To our knowledge this is the first report that fish also have the R-ras gene with identical exon/intron boundaries and extensive homology with human. PMID- 10520452 TI - Nucleotide sequence of oat (Avena sativa L.) cDNA encoding an auxin-binding protein (ABP1). AB - We isolated and determined a nucleotide sequence of a cDNA clone encoding a protein homologous to maize major auxin-binding protein (ABP1) from a cDNA library of oat coleoptiles. The deduced amino acid sequence of this clone contained an N-linked glycosylation signal and an ER-retention signal. Furthermore, two domains that were important to interact with auxins, were conserved in this clone at amino acid level. PMID- 10520453 TI - An oligo-screening strategy to fill gaps found during shotgun sequencing projects. AB - During the course of projects to sequence human and nematode cosmids we encountered great difficulties generating contiguous sequence in regions with repetitive DNA (Alu repeats in humans and tandem or inverted repeats in the nematode). We have developed a simple and efficient strategy to fill gaps. By screening M13, plasmid or phagemid libraries with oligonucleotides flanking the gap, clones are identified that contiguate the cosmid sequence. Our method has been integrated into the GAP4 sequence assembly program. The strategy reduces both time and costs in large scale sequencing projects. PMID- 10520454 TI - Two phosphoglycerate kinase cDNAs from Arabidopsis thaliana. AB - The deduced amino acid sequences of two Arabidopsis thaliana cDNAs share 94% sequence identity among each other. They are 90 to 93% identical to chloroplast phosphoglycerate kinases (chl-PGK) from other plant species. PMID- 10520455 TI - An Arabidopsis thaliana cDNA homologous to cellulase. AB - A cDNA containing an ORF encoding a protein homologous to cellulase has been isolated from an Arabidopsis cDNA library. PMID- 10520457 TI - Sequence of a cDNA coding for a 1-aminocyclopropane-1-carboxylate synthase homolog from Phalaenopsis. AB - A cDNA coding for 1-aminocyclopropane carboxylate (ACC) synthase from Phalaenopsis was cloned and sequenced. Comparison to ACC synthases from Doritaenopsis showed only 67-68% homology at the protein level implicating there are at least two kinds of ACC synthase in orchids. PMID- 10520456 TI - The complete cDNA sequence encoding dog gastric lipase. AB - Oligodeoxyribonucleotide ligation to single-stranded cDNA (SLIC) and polymerase chain reaction (PCR) techniques were used to clone an entire dog gastric lipase (DGL) cDNA. The size of the cDNA is confirmed by Northern blot analysis. The DGL is synthesized as a 379-amino acid mature polypeptide with a molecular mass of 43176 Da which is preceded by a 19-amino acid signal sequence located at the NH2 terminus. Comparison of the signal sequences reveals a high degree of similitude between the DGL, the human gastric lipase (HGL), the rabbit gastric lipase (RGL) and the rat lingual lipase (RLL). PMID- 10520458 TI - The cloning and sequencing of ribosomal protein S18 of parasitic protozoa, Entamoeba histolytica. AB - A cDNA clone encoding ribosomal protein S18 has been isolated from parasitic protozoa, Entamoeba histolytica. The cDNA insert was 495 nucleotides long and an open reading frame (468 nucleotides including the stop codon) coded for 156 amino acid protein which was highly homologous with human, plant and yeast counterparts. PROSITE analysis of the ribosomal protein S18 in eukaryote revealed homology with prokaryotic ribosomal protein S13 signature, which is known to be involved in the initiation of translation. PMID- 10520459 TI - Multiple potential regulatory elements in the 5' flanking region of the human alpha 1a-adrenergic receptor. AB - In spite of their critical importance in myocardial hypertrophy and benign prostatic hyperplasia, nothing is known about mechanisms underlying transcriptional regulation of alpha 1a-adrenergic receptors (alpha 1aARs). Therefore we cloned 6.2 kb of novel sequence upstream of the initiator ATG in the human alpha 1aAR gene. Sequence analysis reveals a TATA-less promoter, the presence of several initiator (Inr) consensus sequences, multiple GC rich regions consistent with Sp-1 binding, and consensus sequences for AP-1 and AP-2 as well as putative cis transcriptional regulatory elements for binding of CREB (cyclic AMP response element binding protein), glucocorticoids, estrogen, and insulin. Compared to the alpha 1bAR, the alpha 1aAR has several more cis regulatory elements, suggesting more complex regulation. The importance of alpha 1aARs in human disease makes it imperative to determine mechanisms underlying transcription and ultimately expression of this receptor. These studies can now be undertaken with the availability of human alpha 1aAR 5'-flanking and 5' untranslated sequence. PMID- 10520460 TI - [Cytoskeletal factors determine the maximum length of cultured fibroblasts]. PMID- 10520461 TI - [Characteristics of cytotoxic proteins, expression of some in nonlymphoid cells is activated upon contact by cells from different organisms]. PMID- 10520462 TI - [Effects of stimulating various segments of amygdala's central nucleus on appearance of the vago-vagal reflex]. PMID- 10520463 TI - [Transposition of the ribosomal RNA locus in Drosophila melanogaster]. PMID- 10520464 TI - [Demand of each of 64 codons in genetic overlapping areas]. PMID- 10520465 TI - [Cloning the human tag7 gene and study of its genomic organization]. PMID- 10520466 TI - [Structural changes in DNA-Ca2+-dipalmitoylphosphatidylcholine complexes during changes in the molar ratio of nucleotide/lipid. Microcalorimetric study]. PMID- 10520467 TI - [Distribution of HindIII-repeats in genomes of Caucasian lizards of the Lacerta species reflect their phylogenetic affiliation]. PMID- 10520468 TI - [Influenza A virus M1 matrix protein is similar to protease inhibitors]. PMID- 10520469 TI - Vaccines. Overview. PMID- 10520470 TI - Francis field trial of inactivated poliomyelitis vaccine: background and lessons for today. PMID- 10520471 TI - Public health considerations for the introduction of new rotavirus vaccines for infants: a case study of tetravalent rhesus rotavirus-based reassortant vaccine. PMID- 10520472 TI - Evaluation of vaccines for the prevention of pneumonia in children in developing countries. PMID- 10520473 TI - Use of the case-control approach in vaccine evaluation: efficacy and adverse effects. PMID- 10520474 TI - Design and interpretation of vaccine field studies. AB - There are many different effects to consider when evaluating vaccines in the field. In this review, we have covered some of the various measures and issues related to study design and interpretation of the different measures. We emphasize that in designing and understanding vaccine studies, it is necessary to be specific about what the effect of interest is and about the assumptions underlying the interpretation of the results. Halloran et al. (81) present design, analysis, and interpretation of vaccine studies in more detail. PMID- 10520475 TI - Combination vaccines: issues in evaluation of effectiveness and safety. PMID- 10520476 TI - Improving immunization coverage rates: an evidence-based review of the literature. PMID- 10520477 TI - The theory of planned behaviour: self-identity, social identity and group norms. AB - The aim of the present study was to examine further the role that self-identity plays in the theory of planned behaviour and, more specifically, to: (1) examine the combined effects of self-identity and social identity constructs on intention and behaviour, and (2) examine the effects of self-identity as a function of past experience of performing the behaviour. The study was concerned with the prediction of intention to engage in household recycling and reported recycling behaviour. A sample of 143 community residents participated in the study. It was prospective in design: measures of the predictors and intention were obtained at the first wave of data collection, whereas behaviour was assessed two weeks later. Self-identity significantly predicted behavioural intention, a relationship that was not dependent on the extent to which the behaviour had been performed in the past. As expected, there was also evidence that the perceived norm of a behaviourally relevant reference group was related to behavioural intention, but only for participants who identified strongly with the group, whereas the relationship between perceived behavioural control (a personal factor) and intention was strongest for low identifiers. PMID- 10520478 TI - Community care and mental disorder: an analysis of discursive resources. AB - The number of people with mental disorders living in the community has recently increased with further increases likely. This study provides a post-attitudinal examination of the discursive resources on which ordinary New Zealanders draw when talking about 'community care'. Four common resources were identified: dual community, rights, disorder and patronization. Each of these resources is examined by using a range of analytic concepts which illustrate the rhetorical achievements and social practices found in the data. We argue that the dual community resource works to position the disordered as being outside the community which is contrary to the broad aim of community care. The analysis of talk of rights was cast as an ideological dilemma for participants who endorsed both universality and conditionality of rights for the disordered. The disorder resource was notable for its flexible rhetorical deployment, while patronization contributed to the positioning of the disordered as subordinate. The implications of these resources are discussed in terms of existing notions of stigma and possibilities for change centred around affiliative resources. PMID- 10520479 TI - Change in adult self-esteem: a longitudinal assessment. AB - This is a longitudinal investigation of self-esteem change in an adult population. The analysis addresses two limitations in earlier studies: the use of convenience samples of children and adolescents, and cross-sectional or short duration longitudinal studies of self-esteem change. Participants are 97 randomly selected married couples interviewed at two points in time separated by 13 years. Two components of the self were measured: self-esteem and reflected appraisals (perception of others' evaluation). Contrary to previous research on self-esteem change, a significant decline was found in all components of the self for both husbands and wives. The decline in self-esteem was not a function of age, education or income. The decline was more likely to occur for high, rather than low, self-esteem participants. This finding is attributed to the demands on higher self-esteem participants to maintain or enhance self-esteem and the caution of low self-esteem participants to engage in behaviours that would threaten the self. PMID- 10520480 TI - Born to abuse? Negotiating identity within an interpretative repertoire of impairment. AB - The focus here is on the constraints and possibilities afforded by the mediation of experience though semi-deterministic notions of the intergenerational transmission of abuse (the 'cycle of abuse' theory). A discourse analytic approach is adopted to accounts in which mothers identify themselves as having been abused in childhood. This identification is seen as inviting negative assumptions and inferences, among them the assumption that childhood abuse is associated with later abusive behaviour. The identity work occasioned by this, and the interpretative repertoires which ground participants' constructions of the general mechanisms through which abuse may be transmitted, are examined. The analysis explores some of the consequences of adopting particular explanatory frameworks for the credentialling of a fully autonomous adult identity. It is suggested that paying attention to this kind of identity work can increase our understanding of the constitutive power of discursive frameworks and the limits of individual resistance within them. PMID- 10520481 TI - A bench-scale investigation of land treatment of soil contaminated with diesel fuel. AB - A bench-scale investigation (soil pan testing) was conducted with the objective of studying degradation rates of diesel contaminated soil (2500 and 10,000 ppm by weight of total petroleum hydrocarbons (TPH) to dry weight of soil) under different treatment conditions over a 17 week testing period. The greatest degradation of the diesel contaminated soil was obtained with the addition of nutrients (Co = 10,000 ppm of TPH; k = 0.19 week-1). 'k' for soil not amended with nutrients was 0.07 week-1. The control cell (C0 = 2500 ppm TPH), with sodium azide (to suppress degradation) was compared with an experimental cell of 2500 ppm initial concentration of TPH without nutrient amendment. The control cell exhibited a relatively low uniform degradation (k = 0.08 week-1) of TPH over the duration of the experiment with reasonable first-order kinetic regression statistics. PMID- 10520482 TI - Cyclodextrin inclusion: catalytic effects on the degradation of organophosphorus pesticides in neutral aqueous solution. AB - alpha-, beta and gamma-Cyclodextrins were found to have promotive inclusion catalytic effects on the degradation of organophosphorus pesticides solubilized in neutral aqueous media. Pesticide degradations were especially accelerated in such systems as alpha-cyclodextrin plus diazinon and beta-cyclodextrin plus chloropyrifos. The above findings are in contrast with the inhibitive catalytic effects of cyclodextrins on pesticide degradations in alkaline media. Cyclodextrin catalytic effects found here were considerably affected by differences in the substitutional and hetero-cyclic properties of the aromatic rings of pesticides and also by the sizes of cyclodextrin cavities. PMID- 10520483 TI - Studies on nitric oxide (NO) formation by the green alga Scenedesmus obliquus and the diazotrophic cyanobacterium Anabaena doliolum. AB - This study provides preliminary evidence that NO production could be a general attribute of algae. Anabaena doliolum was found to be a better NO producer than Scenedesmus and Synechoccocus. Experiments conducted with inhibitors of photosynthesis (DCMU), ATP synthesis (DCCD), and the uncoupler (2,4-DNP) and its analog arsenate clearly revealed that inhibition of nitrite assimilation through the blockage of nitrite reductase (NiR) is primarily responsible for NO emission. A linear relationship between nitrite concentration in the culture medium and NO in the exhaust gas supports the view that accumulation of nitrite is responsible for NO formation. A failure of Scenedesmus, grown in the medium substituted with W for Mo, to produce either NO/NO-2 in light or a 'light-off' peak, and a resumption of these activities upon the addition of Mo proved beyond doubt that a functional nitrate reductase (NR) is necessary for the production of nitrite and NO by algae grown on nitrate as the nitrogen source. Moreover, the appearance of a NO peak immediately after nitrite supplementation under dark conditions in W substituted cultures with or without glucose ruled out an enzymatic role of NR in NO emission. PMID- 10520484 TI - Degradation pathways of PCBs upon UV irradiation in hexane. AB - The photodegradations of eight individual PCB congeners (5, 31, 52, 77, 87, 126, 138, 169) in hexane have been investigated employing a mercury lamp. All degradation reactions of the above mentioned PCB congeners are of the pseudo first order. The principal products of PCB decomposition are the less chlorinated biphenyls, and no PCB-solvent adducts are found. Symmetrical and coplanar PCB congeners show lower photoreactivities. The reactivities of the chlorine atoms at various positions of PCB rings are generally in the order: ortho > meta > para. Photodechlorinations occur mainly on the more substituted rings, when the numbers of chlorine atoms on the two phenyl rings are unequal. During photodegradation, some coplanar PCB congeners are formed, which make the TEQ of solutions to decrease slowly or even to increase. PMID- 10520485 TI - Advanced oxidation of a pulp mill bleaching wastewater. AB - The degradation, by several advanced oxidation reactions, of a pulp mill ECF bleaching effluent, was studied. The initial biodegradability of the organic matter present in the effluent, estimated as the BOD5/COD, was low (0.3). When the effluent was submitted to ozonation and to five different advanced oxidation systems (O3/UV, O3/UV/ZnO, O3/UV/TiO2, O2/UV/ZnO, O2/UV/TiO2), the biodegradability increase significantly. After five minutes of reaction, the O3/UV system appears as the most efficient in to transform the organic matter to more biodegradable forms. A similar effect was observed when the effluent was submitted to an activated sludge treatment. The COD, TOC and toxicity reduction correlated well with the biodegradability enhancement after AOPs treatments. PMID- 10520486 TI - Antimony in municipal waste. AB - Antimony content in municipal waste was studied. Sampled municipal waste was dried, crushed and analyzed. Antimony determinations were performed by Kjeldahl decomposition--batch hydride generation spectrometry and neutron activation analysis. Overall content of antimony in waste was 40-50 g/t raw waste. It was estimated than 20% of the annual production of antimony was discarded as municipal waste in Japan. Leaching of antimony from antimony-added materials may occur, because "small tips" involved considerable amounts of antimony. PMID- 10520487 TI - Pesticide residues in air from coastal environment, south India. AB - Chlorinated pollutants are transported through atmosphere. India is one of the point source countries for these pollutants [1]. In this study the concentration of DDT and HCH were evaluated in air from a tropical coastal environmental (at Parangipettai--southeast coast of India). DDT and HCH ranged in concentrations from 0.16 to 5.93 ng m-3 and 1.45 to 35.6 ng m-3 respectively. The ban on DDT in agriculture is reflected from the low residue levels recorded, predominantly by metabolites other than the parent compounds. PMID- 10520488 TI - PCDDs, PCDFs, PCBs and HCB in marine and estuarine sediments from Queensland, Australia. AB - Concentrations of 2,3,7,8-chlorine substituted PCDDs, PCDFs, selected PCB congeners and HCB were determined in sediment samples collected from sites along the east coast of Queensland in northern Australia. PCDDs were detectable in all sediment samples while PCDFs, PCBs and HCB were mainly found in sediment samples collected from sites in the Brisbane metropolitan area. The results provide evidence that an unidentified source for higher chlorinated PCDDs exists along the Queensland coast. PMID- 10520489 TI - Levels of synthetic musks; bromocyclene and PCBs in eel (Anguilla anguilla) and PCBs in sediment samples from some waters of Berlin/Germany. AB - The purpose of this study was to identify firstly the extent of contamination both of the aquatic biota and of sediments and secondly any regional differences in the concentrations of these compounds. In 1996, polychlorinated biphenyls (PCBs, including their coplanar congeners) were measured in 58 eel and 50 sediment samples. Furthermore, the veterinary pharmacetical bromocyclene and the nitro musks were determined in 84 (1995) and 122 (1996) eel samples. Polycyclic musk fragrances were additionally determined in analyses carried out in 1996. The mean values obtained during the two measurement periods (1995 and 1996) were 24 and 12 micrograms/kg fw. for musk xylene, 41 and 39 micrograms/kg fw. for musk ketone, 14 and 7 micrograms/kg fw. for bromocyclene and, in 1996, 592 micrograms/kg fw. for HHCB (maximum: 4131 micrograms/kg fw.). Decreasing contamination levels are seen for musk xylene and bromocylene but not for musk ketone. This tendency, and the high amounts of polycyclic musk fragrances in the edible parts of eel show that these musks are widely being used in place of nitro musks and that they reach the aquatic system via waste waters, especially those of sewage treatment plants. The mean levels of PCBs in the biota samples of a highly polluted area were 1227 micrograms/kg fw. or 119 pgTEQ/g (sediment: 203 micrograms/kg dw.) and of a slightly polluted area 340 micrograms/kg fw. or 49 TEQ/g (sediment: 47 micrograms/kg dw.). PMID- 10520490 TI - Environmental fate of synthetic pyrethroids during spray drift and field runoff treatments in aquatic microcosms. AB - The aquatic fate and persistence of synthetic pyrethroids under spray drift and field runoff treatment regimens were determined in outdoor pond microcosms. In this paper, the experimental design and construction of outdoor microcosms is presented, as well as the aquatic fate of tralomethrin and deltamethrin. Tralomethrin is rapidly degraded to deltamethrin, with a half-life of 12.7 hours under spray drift conditions. Degradation profiles of tralomethrin in water indicated rapid conversion of deltamethrin and to less active isomers and then to decamethrinic acid (BR2CA). After 24 hours, the percent radioactivity of tralomethrin was 25% of the test material in the water column. In sediment, tralomethrin was immediately converted to deltamethrin. Deltamethrin is rapidly degraded with a half-life of 8 to 48 hours, depending on mechanisms of introduction into water. Degradation profiles of deltamethrin in water indicated rapid conversion of deltamethrin to decamethrinic acid (BR2CA), comprising approximately 90% of the radioactivity in the aqueous phase at 168 hours. Extraction and analysis of fathead minnows (Pimephales promelas) after 96 hours revealed that tissue residues contained parent compounds and metabolites alpha-R deltamethrin, trans-deltamethrin and Br2CA. Fish residues are directly related to aqueous concentrations, thus bioavailability under field runoff regimes were an order of magnitude lower than tissue residues under spray drift conditions. Plant tissue was found to significantly accumulate pyrethroids. PMID- 10520491 TI - A population model applied to HIV transmission considering protection and treatment. AB - An epidemiological population model is proposed to assess the impact of protection and/or treatment strategies applied to HIV infection. Sex-education campaigns are the available protection strategy, and drug (or association of drugs) administration is the treatment strategy considered. In this model we assumed recruitment and differential mortality rates for the homosexual population. In addition to the classical threshold contact rate related to the establishment of the disease, we obtained a threshold input rate. PMID- 10520492 TI - A mathematical model of the ecology of Lyme disease. AB - A mathematical model of enzootic Lyme-disease transmission in a natural focus is presented. This model is based on the life history of the vector tick Ixodes scapularis Say and the primary reservoir host Peromyscus leucopus. Using this model, the threshold condition for the disease to be able to invade a nonenzootic region is determined as a function of the various possible transmission chains operating throughout the year. These expressions show that the transmission chain in which ticks acquire the disease from mice in the fall and transmit it back to mice as nymphs in the spring is the most important chain (contributing approximately 87% of the elasticity of the threshold for the parameter choices examined). Equilibrium disease levels were examined under the assumption of a constant tick population; these levels were determined as a function of tick and mouse density, the vertical transmission rate, the infectivity of mice, and the survivorship parameters of the ticks and of the tick-host contact rates. Vertical transmission has a disproportionately large effect, since unfed infected larval ticks have two opportunities to feed on mice, rather than only one opportunity (as for a newly infected unfed nymph). Finally, a global sensitivity analysis based on Latin hypercube sampling is performed, in which is shown the importance of quantifying the natural history of infection in mice, and of elucidating the contribution of other hosts for I. scapularis than mice. PMID- 10520493 TI - [Long-term oncological results of hepatectomy performed after selective portal embolization]. AB - BACKGROUND: Preoperative selective portal vein embolization (PSPVE), usually of the right portal branch, allows some patients to undergo an hepatectomy which was initially impossible as it would have left an insufficient amount of liver parenchyma. PSPVE induces relative atrophy of the embolized part of the liver, inducing compensatory hypertrophy of the nonembolized part (future remaining liver). Its technical aspects, its tolerance, its immediate results and indications are currently well-known, but long-term results of PSPVE followed by hepatectomy are unknown. The objective of this retrospective study was to assess the long-term survival of this unusual therapeutic approach. PATIENTS AND METHODS: Forty-one patients with initially unresectable (for volumetric reasons) malignant lesions of the liver, underwent PSPVE followed by hepatectomy between September 1987 and September 1998. In two-thirds of cases, the primary tumor was a colorectal adenocarcinoma. The mean number of resected lesions was 4.6 per patient, the mean size of the free margin was 4.3 mm, and 26.8% of the patients presented a (resectable) extrahepatic tumor. RESULTS: Overall 5-year survival (including the two postoperative deaths) was 31.3%, and 5-year survival without recurrence was 24%. For the 27 patients with colorectal metastases (the only homogenous subgroup of the series), overall 5-year survival was 28.6%. Although this result was lower than those obtained with classical hepatectomy (34.4%), it can be considered to be satisfactory, due to the number and size of the lesions. CONCLUSION: In conclusion, it is justified to use PSPVE to make initially unresectable very large liver tumors resectable, in view of the good survival results. PMID- 10520494 TI - [Selective segmental colectomy in diverticular sigmoiditis. The surgical risk is not increased after 70 years of age]. AB - Segmental colectomy for complicated diverticulitis is often indicated, between attacks, in elderly patients. We report a retrospective study of operated patients over the age of 70. The aim of this retrospective study was to evaluate the age risk of consecutive patients operated between diverticular attacks by segmental colectomy, and whether such a surgical procedure is justified, as the life expectancy of these patients was 74 years for men and 81.9 years for women in 1996. PATIENTS AND METHOD: From January 1990 to December 1996, 117 patients were operated, between attacks, for complicated sigmoid diverticulitis; 31% (n = 37) were over the age of 70 years. They were 16 men and 21 women with an overall mean age of 77 (range: 70-85 years). Indications for surgery were repeated attacks (n = 17), large bowel stenosis (n = 14) and colovesical fistula (n = 6). The operation was performed an average 8 weeks after the last infectious episode (3-10 weeks). RESULTS: Among patients over the age of 70 years, the postoperative morbidity rate was 40% and there was no mortality. One patient was reoperated for an anastomotic abscess and was managed by protective colostomy and pelvic drainage with conservation of the anastomosis. CONCLUSION: This retrospective study shows that in our experience, one third of patients operated for a complication of diverticulitis were over the age of 70 years. In this subgroup, left colectomy, when performed between attacks carries an acceptable specific morbidity and no mortality. A further study could compare the percentage and outcome of operated patients with those who were denied surgery. PMID- 10520495 TI - [Long-term results of the treatment of eventrations by intraperitoneal non absorbable prosthesis (149 patients)]. AB - The authors report a series of 149 cases of incisional hernia, operated between 1983 and 1993, by insertion of a non-absorbable prosthetic mesh within the intraperitoneal cavity. This series consisted of 93 women and 56 men, with a mean age 57 years. One third of repairs were performed because of primary treatment failure. One or more operative risk factors were present in 127 patients. A non absorbable intraperitoneal prosthetic mesh was inserted with tension to allow good musculo-aponeurotic repair. Postoperative mortality was 0.6%. All but 13 of the patients, were reviewed with a mean follow-up of 83 months. Twenty eight patients (20%) developed recurrence. In 8 cases, the cause of recurrence was failure of prosthetic mesh insertion because of excessive tension. Three patients (1.7%) developed a fistula in contact with the prosthetic mesh, that had to be removed. A small bowel fistula was observed in 2 cases after an intraoperative wound in 1 case, and a colonic fistula in 1 case. The results of incisional hernia repair with nonabsorbable intraperitoneal prosthetic mesh can be compared with these of other techniques using prosthetic materials. This technique does not require dissection of the intermediate planes and avoids undermining which causes substantial bleeding. The risk of sepsis is also decreased by deep placement of the prosthesis. The exceptional cases of fistula or the possibility of migration of the prosthesis are not exclusively observed with this technique, but must clearly encourage a very strict aseptic technique, with placement of omentum between the prosthetic mesh and the viscera. PMID- 10520496 TI - [Outcome in cervical recurrences of papillary or follicular thyroid cancer]. AB - The aim of this study was to evaluate the treatment and outcome of patients with local recurrence (LR) of differentiated thyroid carcinoma. This retrospective study concerned patients treated between 1974 and 1990 for papillary or follicular thyroid cancer. Our patients had at least one LR. LR diagnosed within 6 months after thyroidectomy and patients with increased serum thyroglobulin levels were excluded. Thirty one patients (80% female) aged 15 to 84 years had at least one LR. LR was diagnosed 7 to 200 months after thyroidectomy (mean 63.7). There were 25 papillary and 6 follicular cancers. There were 1.5 LR per patient (range 1-6). LR were treated by radioiodine in 21 cases and by surgery in 22 cases. Among the 22 surgically treated patients, 7 had nodal recurrences, 7 had nodes and tumor, 3 had only tumor, 1 had recurrence in the remnant thyroid. After a mean follow-up of 75.8 months, 11 patients had distant metastases, 11 had died from their thyroid carcinoma (7 after metastases). Three of the 7 patients with nodal recurrence died. In one third of cases, LR announced distant metastases. Node recurrence had a poor prognosis. PMID- 10520497 TI - [Indications and results of extemporaneous examination of pelvic lymph nodes in the surgical strategy of stage Ib or II cancers of the cervix uteri]. AB - The aim of this study was to evaluate the accuracy of frozen section examination of lymph nodes and its place in the surgical management of early stage cervical cancer. This study was based on 80 patients with stage Ib (n = 76) or IIa (n = 4) invasive cervical carcinoma, with tumor size < 3 cm, treated by radical hysterectomy with pelvic lymphadenectomy with frozen section examination of pelvic lymph nodes. A total of 718 nodes were submitted to frozen section examination. Only one case of false-negative result was found (micrometastases). The sensitivity of frozen section examination in detecting metastatic nodes was 92.3%, its specificity was 100% and its negative predictive value was 97%. Frozen section diagnosis of pelvic nodes is a reliable procedure and should be carried out on obturator, external iliac and common iliac nodes. It should, therefore be performed in patients with early cervical carcinoma to avoid routine para-aortic lymphadenectomy. PMID- 10520499 TI - [Intraductal papillary mucinous tumors of the pancreas: diagnosis, treatment and prognosis]. AB - Intraductal papillary and mucinous tumors of the pancreas (IPMT) have been recently well defined histologically. Recent reports have described their diagnostic and therapeutic modalities. The malignant potential of these lesions warrants their surgical resection. The main difficulty of their management is the pre-operative assessment of ductal extension and grade of dysplasia. Despite the use of various imaging modalities (ultrasound, endoscopic ultrasonography, CT scan, retrograde pancreatography, pancreatic MRI), examination of the pancreatic margin on frozen section remains mandatory to ensure complete resection of epithelial lesions. This usually requires partial pancreatic resection. The long term outcome is favourable for lesions with no invasive component. Recurrence in case of invasive malignant transformation appear to be more frequent than previously reported. PMID- 10520498 TI - [Study of sustained forces and the working space of endoscopic surgery instruments]. AB - The instruments currently used in endoscopic surgery are limited by several factors, in particular their reduced working space. In order to develop instruments with manifold degrees of freedom (DOF), the elementary actions performed by the existing instruments must be defined. MATERIAL AND METHODS: We have broken down into elementary movements the actions performed by the currently used instruments and analyzed them by measuring the strain on the instruments and on their working volume. The elementary actions were performed in vitro (Pelvitrainer) on an animal model (pig) and also in the course of clinical practice. A total of 6,750 measurements were carried out for the following actions: grasping, cutting, dissecting, suturing and knotting. RESULTS: The largest working volumes were measured for the knotting and suturing actions (198 degrees in axial rotation and 69 degrees in lateral translation). The range of the working space was between 8 degrees and 52 degrees. Forces exerted on the instrument were between 0.5 and 12 Newton (N). Forces exerted on the headpoint were between 0.4 and 10.5 N whereas the friction forces were between 0.5 and 1.5 N. COMMENTS: By analyzing the elementary actions of endoscopic surgery, we were able to quantify the directions and dimensions of forces sustained by the instruments and were also able to measure the working volume involved when performing the main actions in endoscopic surgery. The results obtained provide a basis for the development of more sophisticated instruments. PMID- 10520500 TI - [Value of preoperative drainage of the bile ducts in obstructive jaundice]. AB - Hepato-biliary surgery for obstructive jaundice is associated with high morbidity and mortality rates. Experimental and clinical studies on obstructive jaundice revealed endotoxaemia, coagulation disorders and depressed immune function. Many studies have been carried out to identify the operative risk factors. The serum bilirubin level seemed to be a significant factor. Biliary decompression via a percutaneous or endoscopic retrograde approach was therefore proposed to improve the surgical outcome. The first retrospective studies have suggested a reduction of morbidity and mortality. Subsequent randomized studies have not confirm the benefit of preoperative biliary drainage because of procedure-related complications. The article reviews the literature on preoperative biliary drainage and proposes the indications, choice of method and optimal duration of biliary drainage. PMID- 10520501 TI - [Prevention of pancreatic fistula after cephalic duodenopancreatectomy]. AB - A pancreatic fistula occurs in about 10% of cases after Whipple's procedure. This complication is associated with a mortality rate ranging from 7% to 30%. The main predisposing factor of pancreatic fistula is a soft pancreatic parenchyma. Several procedures have been proposed to decrease the rate of this complication. Occlusion of the residual stump is infrequently used and does not clearly reduce the rate of this complication. Pancreaticojejunostomy is the technique most frequently used. No alternative technique is clearly superior to pancreaticojejunostomy. Pancreaticogastrostomy and pancreaticojejunostomy have equivalent early results. Superiority of transient intubation of the Wirsung duct and mucosa-to-mucosa anastomosis is not demonstrated. Among the 7 controlled randomized studies which tested somatostatin or its analogs, many have methodological insufficiencies which prevent definite conclusions. Meanwhile, most studies suggest that these drugs decrease the rate of pancreatic fistula after pancreaticoduodenectomy. Further evaluation in high-risk patients (soft pancreatic parenchyma) is advisable. PMID- 10520502 TI - [Origin of animal experimentation legislation in the 19th century]. AB - The first legislation in the world, designed to protect animals used in research, was passed in England in 1876, and is still in force today. It is one of the strictest in Europe. At the same period, France had no such law, and was the country conducting the greatest amount of animal experimentation. Comparing, these two countries, in the middle of the 19th century, can account for this difference. The most important difference seems to be related to the theological question: are animals endowed with a soul? Saint Augustine, claimed, in the 4th century, perhaps because of an experiment with the centipede, that animals do not have a soul. In the 17th century, Rene Descartes, using a different philosophical system, reached a similar conclusion, in France. On the other hand, under the influence of Charles Darwin, England rejected the Roman Catholic conclusion, about the soul of animals. The industrial revolution, occurring earlier in England than in France, also changed the society, developing urban areas, where people were cut off from rural life and changing human relationships with animals. The industrial revolution enabled the development of the press, giving impetus to public opinion. These facts, combined with a caution of science, which was more developed in England than in France, brought about the first important "anti-doctor" campaign. PMID- 10520503 TI - [Vascular control during left splenopancreatectomy in cancer]. AB - Distal pancreatectomy remains the gold standard for resection of left-sided pancreatic carcinoma. For oncologic and surgical reasons, the control of splenic vessels is an important phase of this operation. Based on anatomical considerations, the two resection techniques are presented in this paper: by first dividing the pancreatic neck or by first removing the spleen. PMID- 10520504 TI - [Late cutaneous fistula after laparoscopic cholecystectomy]. PMID- 10520505 TI - [Severe hemobilia after percutaneous transhepatic drainage: radiological and surgical management]. PMID- 10520506 TI - [Huperzine a: an acetylcholinesterase inhibitor with high pharmacological potential]. AB - Huperzine A is an alkaloid isolated from a chinese club-moss. The molecule is a potent and selective inhibitor of acetylcholinesterase. Several pharmacological and clinical studies showed that huperzine A improves the mnesic capacity and cognitive functions. Huperzine A was also found to be a neuroprotective agent. This molecule which possesses a high pharmacological potential is under clinical evaluation for the palliative treatment of Alzheimer's disease. In addition, its use in the pretreatment of poisoning by organophosphorous nerve agents could be another indication. PMID- 10520507 TI - [Effect of presynaptic dopaminergic agonism on the immobility of mice in the forced swimming test]. AB - Administration of 0.05 mg/kg (i.p.) of apomorphine (APO) or of the two dopamine (DA) lipoamides, DA steatamide (S-DA) or DA palmitamide (P-DA), at doses of 10 or 30 mg/kg (i.p.), induced an anti-immobility effect, in the forced swimming test in mice, of the same order as those of imipramime (30 mg/kg, i.p.). At the aforementioned doses APO, S-DA or P-DA could be acting on the dopaminergic autoreceptors and cause a decrease of the turnover and/or the release of the brain DA in the striatum. In contrast, DA linoleamide (L-DA), which has no action on the brain DA, did not exhibit anti-immobility effect. It is suggested that the decrease of turnover and/or release of the brain DA, induced by APO, S-DA or P DA, could constitute a clue of the extent of the extrasynaptic action developed by these drugs whose antiglutamatergic incidence could cause the anti-immobility effect, observed with these derivatives, as well as their possible antidepressive action. PMID- 10520508 TI - [Prenatal diagnosis of genetic diseases in France]. AB - Prenatal diagnosis has been introduced in medicine in the seventies with aminocenteses and amniotic cells cultures. It was applied to the diagnosis, during the second trimester of pregnancy for chromosomal abnormalities (mainly Down syndrome in women 38 years of age) and inborn errors of metabolism with very severe handicaps). Since 1970, obstetrical techniques have improved giving access to several fetal biological samples, knowledge in genetics has identified more diseases and biological analyses have become more accurate. During the same time legislation has been instituted: in France the law of 1994 July 29th established rules for prenatal diagnosis. Among theses rules are defined the objective of prenatal diagnosis, the requirement for medical genetic counselling and official authorizations for cytogenetic, infectious diagnosis, biological diagnosis of genetic diseases (biochemical, molecular genetic, hematology, immunology) and maternal plasma markers of chromosomal abnormalities (Down syndrome). Recently (1997 may 28th) have been established "pluridisciplinary prenatal diagnosis centers", including complementary, clinical and biological services to insure the safety of Prenatal Diagnosis. Preimplantation Diagnosis (PID) inherited diseases has become recently possible with the techniques of in vitro fertilization, blastomere biopsy of the early embryo and DNA analysis of the single blastomere. Only a very few centres worldwide offer PID. In France PID is not yet allowed but the National Ethical Committee examined the question and legislation is about to be published. PMID- 10520509 TI - [Substituted thiazolidinediones and thioxothiazolidinones: synthesis and structure]. AB - Synthesis and physico-chemical properties of 3-(4-bromobenzyl)-, 3-(4 chlorobenzyl)-5-arylidene-thiazolidine-2,4-diones and 3-(4-chlorobenzyl)-4-thioxo 5-arylidene-thiazolidin-2- ones are described. Twelve new products were synthesized by the aldolisation-crotonisation reaction from aromatic aldehydes and N-alkylated thiazolidinediones or thioxothiazolidinones. Seven compounds were preliminary tested for their bacteriostatic activity. PMID- 10520510 TI - [Comparison of the thermostability of natural (sulfoprolamine and sodium usnate) and synthetic (climbazole and piroctone olamine) antidandruff agents]. AB - We compared thermostability of various natural (sulfoprolamine and sodium usnate) or synthetic (climbazol and piroctone olamine) antidandruff agents in aqueous diluted solution at pH around 7. Thermodegradation of these solutions was studied by an isothermal method in thermostatically controlled ovens, at three temperatures (50, 70 and 90 degrees C). For each molecule, we determined at 20 degrees C t90% (time necessary to obtain a decrease of 10% of the initial concentration, value which shows the stability of the product). The present study shows that piroctone olamine is the most stable antidandruff agent among those studied. PMID- 10520511 TI - [Reduction of renal uranium uptake by acetazolamide: the importance of urinary elimination of bicarbonate]. AB - Acetazolamide was compared with bicarbonate for the treatment of contamination with uranium. Uranium was injected peritoneally in rats, and its distribution was investigated. Acetazolamide was three times more efficient than bicarbonate in reducing the renal content of uranium. On the other hand, it had no effect on hepatic or skeletal content. In this study, renal physiology provides the basis for understanding the mode of action of acetazolamide and bicarbonate. In this context, it is of interest to determine the alkalinity of the urine, with the aim of knowing whether bicarbonate is present to mobilize uranium. PMID- 10520512 TI - [Standardization of the aerial parts of Alchemilla]. AB - Dried aerial parts of Alchemilla xanthochlora Rothm. (16 batches), A. glabra Neygenf. (1 batch), A. coriacea Buser (2 batches) and A. filicaulis Buser (3 batches) present a similar flavonoid and tanin pattern. In the case of A. xanthochlora, the mean levels of the principal compounds were: total flavonoids 2.22%, glucuronyl-3 quercetol 1.18%, tanins 16.0% and ellagic acid 0.36%. The flavonoid levels were higher before flowering and the tanin levels higher during flowering. Four commercial batches were examined for a comparative study. Pharmacopoeial specifications were proposed for a revision of the monograph "Alchemillae herba". PMID- 10520513 TI - [Standardization of hazel leaf]. AB - Dried leaves of 9 harvested batches and 5 batches of commercial origin from Corylus avellana L. were examined. The levels of principal polyphenolic compounds averaged respectively: total flavonoids 2.58 and 2.58%, myricitrin 1.09 and 1.35%, quercitrin 0.30 and 0.40%, tannins 5.2 and 6.5%. Specifications were discussed for a French Pharmacopoeial monography. PMID- 10520514 TI - [Standardization of the sour orange flower and leaf]. AB - Dried flowers (1 batch) and leaves (6 batches) of sour orange Citrus aurantium L. had a similar flavonoid pattern. But the flavonoid levels of flowers were higher than those of leaves. The mean levels of the principal flavonoid compounds were respectively: total flavonoids 12.35 and 1.06%, neohesperidin 5.44 and 0.08%, naringin 1.93 and 0.06%, eriocitrin 0.38 and 0.25%. 18 batches of commercial origine were also examined for a comparative study. Specifications were proposed for a revision of the monographs "Sour orange flower" and "Sour orange leaf" of the French Pharmacopoeia. PMID- 10520515 TI - [New derivatives of 1-methyl-phthalazine with antimicrobial and fungistatic action]. AB - In this paper we present the antimicrobial and antifungical action of some new salts of 1-methyl-phtalaziniums and their corresponding ylids. We also present conclusions concerning the relation between structure and biological activity. The test was performed using the diffusimetric method with rustless steel cylinders based on the diffusion on the tested substances on gelose surface. The salts of 1-methyl-phtalaziniums are always more active comparatively than their corresponding ylids. All the compounds but not all derivatives are more active on the Candida albicans. PMID- 10520516 TI - [The pneumonitis mortality trend in Italy in the 1975-94 period]. PMID- 10520517 TI - [Streptococcus pneumoniae meningitis in Italy--1994-98]. PMID- 10520518 TI - [The antibiotic resistance and serotypes of invasive Streptococcus pneumoniae strains isolated in Italy]. PMID- 10520520 TI - [Community-acquired pneumonitis (CAP). The viewpoint of the general practice physician]. PMID- 10520521 TI - [The national health plan: its applications and outlook]. PMID- 10520519 TI - [Nosocomial infections of the lower airways: the risk factors in COBP]. PMID- 10520522 TI - [The prevention of infectious diseases in adults and the elderly]. PMID- 10520523 TI - [The clinical efficacy of antipneumococcal vaccination. A brief report]. PMID- 10520524 TI - [Prevention in the elderly: the vaccine center, the hospital and the family physician]. PMID- 10520525 TI - [An antipneumococcal vaccination program for the elderly of Bologna]. PMID- 10520526 TI - [Anti-influenza and antipneumococcal vaccinations in the Sicily area]. PMID- 10520527 TI - [A proposal for antipneumococcal vaccination in the Lombardy region]. PMID- 10520529 TI - [Development and validation of a graphic method for spatial interpretation and evaluation of biplanar coronary angiograms]. AB - Visual evaluation and measurement of the lesion geometry is impaired by the foreshortening of the radiographically displayed target coronary segment. For this reason, we developed and validated a simple graphic procedure that facilitates the spatial interpretation and allows an objectifiable assessment of the view of the target coronary segment. The method computes a spatial axis of the coronary segment imaged in two planes and determines its inclination in the radiation path of each projection view. A triangle made up of the axis of the imaged segment, the unforeshortened axis of the imaged segment and the foreshortening height is displayed in each projection plane. The shape of the triangle indicates the degree of foreshortening while its position in the angiogram indicates the orientation of the spatial axis relative to the observer. The method was validated by comparing calculated and true foreshortening in the radiographic views of a centimetre grid obtained in the usual angiographic projections. The method has an accuracy and precision of 0.05 +/- 0.62%. It is applied clinically to evaluate biplane segmental visualization during coronary interventions and to select valid segmental views for measurements during quantitative biplane coronary angiography. This application may considerably facilitate the interpretation and assessment of biplane angiograms. PMID- 10520528 TI - [The Working Group on the Prevention of S. pneumoniae Infections in Italy. The final report]. PMID- 10520530 TI - [Calculation of internal from external electrical field strength and its effect on cardiac pacemaker patients]. AB - What electric field strength is developed inside a person entering the electric field produced by, e.g., a high-voltage transmission line? This question may be of vital importance for a pacemaker patient. With the aim of finding an answer, known phenomena are combined in such a way that the mean current density within the thorax at the level of the heart can be determined. In this way it can be demonstrated that the internal electric field of a grounded person is twice that of an ungrounded person, that the internal field is 0.72 mV/m per 1 kV/m external field, and that the most sensitive pacemaker can be influenced by 2 kV/m. It is also clear that the field strength limit of 5 kV/m proposed by ICNIRP for the general public cannot be justified by the basic limit of 2 mA/m2. PMID- 10520531 TI - Two-dimensional Fourier representation used in the bioelectric forward problem. AB - The objective of this paper is the application of two-dimensional discrete Fourier transformation for solving the integral equation of the bioelectric forward problem. Therefore, the potential, the source term, and the integral equation kernel are assumed to be sampled at evenly spaced intervals. Thus the continuous functions of the problem domain can be expressed by their two dimensional discrete Fourier transform in the spatial frequency domain. The method is applied to compute the surface potential generated by an eccentric dipole in a homogeneous spherical conducting medium. The integral equation for the potential is solved in the spatial frequency domain and the value of the potential at the sampling points is obtained from inverse Fourier transformation. The solution of the presented method is compared to both, an analytic solution and a solution gained from applying the boundary element method. Isoparametric quadrilateral boundary elements are used for modeling the spherical volume conductor in the boundary element solution, while in the two-dimensional Fourier transformation method the volume conductor is represented by a parametric boundary surface approximation. PMID- 10520532 TI - [Possibilities of spinal endoscopy within the scope of minimal invasive intervention--an experimental study]. AB - The purpose of the present study was to evaluate, in vitro, a newly designed spinaloscope with a diameter of 1.8 mm, with integrated portals for instruments and irrigation. The 0 degree optical system has a resolution of 6,000 pixels. The instrument portals can be used for surgical lasers, biopsy forceps or burrs. We carried out our evaluations on fresh (unfixed) human lumbar spine specimens. The position of the endoscope was documented by CT scans. The endoscope was introduced into the spinal canal via the hiatus sacralis using a blunt trocar. The various structures and tissues were clearly identifiable and included the dura, the lig. flavum, the lig. long. posterior, spinal nerves, small pieces of disc material and various fibrous bands. The usefulness of the biopsy forceps was also shown. PMID- 10520533 TI - [An optical method for determining deformation and form changes in polyethylene surfaces of explanted acetabular cups of hip endoprostheses]. AB - AIM: Most methods used for the determination of volumetric wear of polyethylene cups are based on the assumption that the head of the prosthesis penetrates the cup in "cylindrical" fashion. The new accurate optical method is independent of this disputable assumption. METHOD: The articulating surface of the cup is scanned with light and a data set of 60,000 pixels obtained in this way is stored in a computer. Data obtained from used cups were compared with those obtained from unused cups. The volumetric wear was calculated directly by threefold integration. To assess the changes in surface shape, the data are fitted by an ellipsoid whose long axis defines the mean direction of load. A total of 18 retrieved and 3 unused cups of different types were studied. RESULTS: The unused acetabular cups deviated only slightly from ideal hemispheres. The surfaces showed rotational symmetry, and an undulation having an amplitude of 0.1 mm between dome and equator. For all explanted cups, the assumption of cylindrical penetration of the head into the polyethylene was shown not to represent the true situation. The cup expands in all directions, and the volumetric wear is underestimated by 50% with the traditional methods. The data suggest that long term survival may be jeopardized when the main direction of loading is centered on the dome of the cup. Ceramic heads were associated with smaller rates of volumetric wear. CONCLUSION: The new optical method is characterised by short measuring times, precision and simple application. Analysis of the wear patterns of polyethylene components using this technique may contribute to a further understanding of the complex mechanisms of aseptic loosening. PMID- 10520534 TI - Investors in people. PMID- 10520535 TI - Mike Grace interviews QDA winner John Barnet-Lamb. Quality Development Award. PMID- 10520536 TI - The benefits of the Benevolent Fund. PMID- 10520538 TI - Replantation of teeth an option. PMID- 10520537 TI - Industry in crisis over bleaching issue. PMID- 10520539 TI - Evidence of poor oral health in ethnic minorities. PMID- 10520540 TI - Nurses needed in fight against pain. PMID- 10520541 TI - Dental problems for nuclear workers. PMID- 10520542 TI - The psychology of dental patient care. 9. Communicating effectively: some practical suggestions. PMID- 10520543 TI - Dental implants. 3. Assessment of the dentition and treatment options for the replacement of missing teeth. PMID- 10520544 TI - Multiple sclerosis, dental caries and fillings: a case-control study. AB - OBJECTIVES: To investigate the association between multiple sclerosis, dental caries, amalgam fillings, body mercury and lead. DESIGN: Matched case-control study. SETTING: Leicestershire in the years 1989-1990. SUBJECTS: Thirty-nine females with multiple sclerosis (of recent onset) were matched with 62 controls for age, sex and general practitioner. METHODS: Home interview of cases and controls within which there was an assessment of the DMFT index and blood and urine mercury and lead levels. RESULTS: The odds of being a MS case increased multiplicatively by 1.09 (95% CI 1.00, 1.18) for every additional unit of DMFT index of dental caries. This represents an odds ratio of 1.213 or a 21% increase in risk of MS in relation to dental caries in this population. There was no difference between cases and controls in the number of amalgam fillings or in body mercury or lead levels. There was a significant correlation between body mercury levels and the number of teeth filled with amalgam (controls: r = +0.430, P = 0.006, cases: r = +0.596, P = 0.001). CONCLUSION: There was evidence of excess dental caries among MS cases compared with the controls. This finding supports the strong geographical correlation between the two diseases. A further study of this association is recommended. PMID- 10520545 TI - Fluoride release, weight loss and erosive wear of modern aesthetic restoratives. AB - OBJECTIVE: In this investigation, the in vitro sustained fluoride release, weight loss and erosive wear of three conventional glass ionomer cements (Fuji IX, ChemFil Superior, Ketac-Silver), three resin-modified glass ionomer cements (Fuji II LC, Vitremer, Photac-Fil), a polyacid-modified resin composite (Dyract), and a resin composite control material (Z100) were compared. METHODS: The amounts of fluoride released and weight changes were measured for 12 weeks using a fluoride electrode with TISAB III buffer. After 12 weeks, the specimens were recharged with fluoride using 2 mL of 1.23% APF gel. The recharged specimens were assessed for the amounts of fluoride released and weight changes over another 12 weeks. At the end of the experiment, the specimens were examined with SEM and surface profilometry. RESULTS: All materials, with the exception of Z100, showed the highest initial fluoride release rates during the first 2 days, dropping quickly over 2 weeks and becoming largely stabilised after 5 weeks, in an exponential mode. The recharging of the specimens with APF gel caused a large increase in the amounts of fluoride released during the first 2 days only. Analyses for all cements showed strong correlations between mean weight loss and cumulative fluoride release over a 5-week period following the application of the APF gel. SEM and surface profilometry found that roughness increased from the polyacid modified resin composite to the conventional glass ionomer cements. CONCLUSIONS: APF gel caused erosive wear of the glass ionomer cements especially, and the wear correlated well with the weight losses. To minimise surface erosion, APF gel should not be used on these cements, especially as the recharging effects are transitory. PMID- 10520546 TI - A closer look at General Dental Service orthodontics in England and Wales. II: What determines appliance selection? AB - AIM: To elucidate factors that influence choice of appliance type in General Dental Service (GDS) orthodontics in England and Wales. METHOD: Records were obtained for 1527 cases, representing a systematic 2 per cent sample of GDS cases completed during 1990-91. Evaluation involved Discriminant Analysis to find the most influential factors in appliance choice. Factors considered included patient and practitioner characteristics, and features of the malocclusion as assessed by Occlusal Indices. RESULTS: Full data were available for 1217 cases. 24 per cent of treatments included use of dual- and 26 per cent single-arch fixed appliances. Appliance choice was predictable in 55 per cent of cases. Older patients, orthodontically qualified practitioners, high Peer Assessment Rating score at start, permanent dentition, lower grades of the Dental Health Component of the Index of Orthodontic Treatment Need at start, and practitioners with high gross earnings from orthodontics, all tended to be associated with more frequent use of fixed appliances. CONCLUSIONS: Possession of a diploma or membership in orthodontics was associated with more frequent use of both dual- and single-arch fixed appliances. Better appliance selection, and thus more effective treatments in the GDS, may result from a greater availability of practitioners with formal postgraduate training in orthodontics. PMID- 10520563 TI - Default nominal inflection in Hebrew: evidence for mental variables. AB - According to the 'word/rule' account, regular inflection is computed by a default, symbolic process, whereas irregular inflection is achieved by associative memory. Conversely, pattern-associator accounts attribute both regular and irregular inflection to an associative process. The acquisition of the default is ascribed to the asymmetry in the distribution of regular and irregular tokens. Irregular tokens tend to form tight, well-defined phonological clusters (e.g. sing-sang, ring-rang), whereas regular forms are diffusely distributed throughout the phonological space. This distributional asymmetry is necessary and sufficient for the acquisition of a regular default. Hebrew nominal inflection challenges this account. We demonstrate that Hebrew speakers use the regular masculine inflection as a default despite the overlap in the distribution of regular and irregular Hebrew masculine nouns. Specifically, Experiment 1 demonstrates that regular inflection is productively applied to novel nouns regardless of their similarity to existing regular nouns. In contrast, the inflection of irregular sounding nouns is strongly sensitive to their similarity to stored irregular tokens. Experiment 2 establishes the generality of the regular default for novel words that are phonologically idiosyncratic. Experiment 3 demonstrates that Hebrew speakers assign the default regular inflection to borrowings and names that are identical to existing irregular nouns. The existence of default inflection in Hebrew is incompatible with the distributional asymmetry hypothesis. Our findings also lend no support for a type-frequency account. The convergence of the circumstances triggering default inflection in Hebrew, German and English suggests that the capacity for default inflection may be general. PMID- 10520564 TI - Children's understanding that utterances emanate from minds: using speaker belief to aid interpretation. AB - Children interpreted an utterance made by a protagonist with a false belief, such as, 'I would like the car in the garage.' Calculating the speaker's belief in conjunction with the literal meaning of the utterance would lead to the correct interpretation that the intended referent is the car on the track, given that the car in the garage swapped places with the one on the track. In Experiments 1 and 2, many children aged around 4 and 5 years wrongly indicated the car in the garage. In contrast, many correctly indicated the car on the track when it was unnecessary to consider the speaker's belief because the utterance was, 'the car I put in the garage'. Six-year-olds found both kinds of utterance equally easy in Experiment 1, while 3-year-olds had equal difficulty with both. In Experiments 2 and 3, the speaker gave an ambiguous utterance and many children aged between 3 and 6 years successfully used information about the speaker's belief to identify which of several candidate referents was intended. We discuss the results in relation to characteristics of utterance comprehension and consider implications for developments in understanding the mind by children beyond 4 years. PMID- 10520565 TI - Language acquisition in the absence of explicit negative evidence: how important is starting small? AB - It is commonly assumed that innate linguistic constraints are necessary to learn a natural language, based on the apparent lack of explicit negative evidence provided to children and on Gold's proof that, under assumptions of virtually arbitrary positive presentation, most interesting classes of languages are not learnable. However, Gold's results do not apply under the rather common assumption that language presentation may be modeled as a stochastic process. Indeed, Elman (Elman, J.L., 1993. Learning and development in neural networks: the importance of starting small. Cognition 48, 71-99) demonstrated that a simple recurrent connectionist network could learn an artificial grammar with some of the complexities of English, including embedded clauses, based on performing a word prediction task within a stochastic environment. However, the network was successful only when either embedded sentences were initially withheld and only later introduced gradually, or when the network itself was given initially limited memory which only gradually improved. This finding has been taken as support for Newport's 'less is more' proposal, that child language acquisition may be aided rather than hindered by limited cognitive resources. The current article reports on connectionist simulations which indicate, to the contrary, that starting with simplified inputs or limited memory is not necessary in training recurrent networks to learn pseudonatural languages; in fact, such restrictions hinder acquisition as the languages are made more English-like by the introduction of semantic as well as syntactic constraints. We suggest that, under a statistical model of the language environment, Gold's theorem and the possible lack of explicit negative evidence do not implicate innate, linguistic specific mechanisms. Furthermore, our simulations indicate that special teaching methods or maturational constraints may be unnecessary in learning the structure of natural language. PMID- 10520566 TI - First outbreak of influenza in the United Kingdom this season hits Orkney. PMID- 10520567 TI - Post licensing evaluation of meningococcal C conjugate vaccine efficacy. PMID- 10520568 TI - Fertility control in wild populations of animals. PMID- 10520569 TI - Bidding on the future? The limits of paying for gametes. PMID- 10520570 TI - The diagnostic and prognostic value of traditional semen parameters. PMID- 10520571 TI - Brown-colored semen in men with spinal cord injury. AB - The interesting condition of brown-colored semen has often been observed during assisted ejaculation of men with spinal cord injury (SCI). This condition has not been reported in the literature, and its cause is unknown. To investigate this condition, the present study examined the incidence and quality of brown semen and its relationship to level of SCI, time since SCI, number of successive ejaculations, ejaculation frequency, and ejaculation method in a total of 664 semen specimens from 162 SCI men. In addition, a microscopic evaluation was performed on brown semen specimens from SCI men, not-brown specimens from SCI men, and normally colored specimens from normal men. The results showed that 27% of SCI subjects had brown semen on at least one ejaculation. There was no difference between men producing and men not producing brown semen in age, level of injury, or years postinjury. The duration of an ejaculation, number of successive ejaculations, and frequency of ejaculation were not associated with occurrence of brown semen. Sperm concentration and sperm motility were not significantly different in brown and not-brown specimens. Specimens from subjects who produced brown semen had similar pH but lower volume than specimens from subjects who did not produce brown semen. Brown semen had a thin consistency more often than not-brown semen. Brown specimens contained intact red blood cells (RBCs) and/or heme pigment more often than not-brown specimens; however, one half and one third of the specimens, respectively, contained neither RBCs nor heme pigment. The cause of brown semen is unknown but may relate to seminal-vesicle dysfunction. PMID- 10520572 TI - Estradiol and high-dose dihydrotestosterone treatment causes changes in cynomolgus monkey prostate volume and histology identical to those caused by testosterone alone. AB - To evaluate the effects of testosterone (T), dihydrotestosterone (DHT), and estradiol (E2) on the regulation of prostate growth and tissue composition, the following study was conducted in a nonhuman primate model. Fifteen adult, long term castrated cynomolgus monkeys were randomly assigned to receive implants filled with T (0.19 +/- 0.01 g), DHT alone (0.21 +/- 0.01 g), or (99%) DHT + (1%) E2 (0.21 +/- 0.01 g). Prior to and at 4-week intervals during the treatment phase of 252 days, prostate volumes (PV), body weight, ejaculate weight, hormone levels (of T, DHT, and E2), and red blood cell count were measured. Five adult, intact monkeys served as controls for prostate volume and histology. At the end of the study, histological analysis of an ultrasound-guided prostate biopsy was performed. T levels increased significantly in the T group compared with baseline (P < 0.01) and with the DHT and DHT + E2 groups (P < 0.05). Both groups receiving DHT showed higher DHT levels than did animals in the T group (P < 0.001). E2 levels in all groups increased over time (P < 0.05), although significant differences (P < 0.01) could only be detected between the DHT + E2 and the DHT group. Prostate volume in all groups increased (at baseline: T = 1.03 +/- 0.12 ml, DHT = 1.08 +/- 0.15 ml, DHT + E2 = 1.13 +/- 0.09; at day 252: T = 5.83 +/- 1.00, DHT = 4.72 +/- 0.9, DHT + E2 = 5.05 +/- 0.62) over time (P < 0.001), whereas no differences could be detected between the groups. Prostate biopsy could be performed successfully in 15 out of 20 monkeys. Prostate tissue evaluation between the treatment groups and the evaluated intact monkeys revealed no differences in the status of secretory epithelia, nuclear chromatin, excretory vacuoles, interstitial stroma, smooth muscles, and total functional status, whereas the prostate of a long-term castrated monkey showed severe atrophy. Thus, both androgens fully restored prostate volume and ejaculatory function. Highly supraphysiological DHT serum levels are not associated with abnormal volumetric or histological changes of the prostate. Comparing the DHT group with the DHT + E2 group, an additional stimulatory effect of normal or slightly elevated estrogens on the prostate cannot be found in the presence of highly supraphysiological DHT levels. PMID- 10520573 TI - Serum dihydrotestosterone and testosterone concentrations in human immunodeficiency virus-infected men with and without weight loss. AB - Weight loss is an important determinant of disease outcome in human immunodeficiency virus (HIV)-infected men. Others have suggested that a defect in dihydrotestosterone (DHT) generation contributes to weight loss in HIV-infected men. To determine whether DHT levels correlate with weight loss independently of changes in testosterone levels, we prospectively measured serum total- and free testosterone and DHT levels in 148 consecutive HIV-infected men and 42 healthy men. Thirty-one percent of HIV-infected men had serum testosterone levels less than 275 ng/dL, the lower limit of the normal male range; of these, 81% had normal or low LH and FSH levels (hypogonadotropic), and 19% had elevated LH and FSH levels (hypergonadotropic). Overall, serum testosterone, free-testosterone, and DHT levels were lower in HIV-infected men than in healthy men, but serum DHT to-testosterone ratios were not significantly different between the two groups. Serum total- and free-testosterone levels were lower in HIV-infected men who had lost 5 lb or more of weight in the preceding 12 months than in those who had not lost any weight. Serum DHT levels and DHT-to-testosterone ratios did not differ between those who had lost weight and those who had not. Serum testosterone and free-testosterone levels, but not DHT levels, correlated with weight change and with Karnofsky performance status. We also performed a retrospective analysis of data from a previous study in which HIV-infected men with serum testosterone levels less than 400 ng/dL had been treated with placebo or testosterone patches designed to nominally release 5 mg testosterone over 24 hours. Serum testosterone to-DHT ratios did not change after testosterone treatment. Changes in fat-free mass were correlated with changes in both serum testosterone (r = 0.42, P = 0.018) and DHT (r = 0.35, P = 0.049) levels. Serum total- testosterone and DHT levels were highly correlated with one another, and when the change in serum testosterone was taken into account, serum DHT levels no longer showed a significant correlation with change in fat-free mass. We conclude that DHT levels are lower in HIV-infected men than in healthy men but that neither DHT levels nor DHT-to-testosterone ratios correlate with weight loss. During testosterone treatment, serum DHT levels increase proportionately, but the increments in serum testosterone correlate with the change in fat-free mass. Our data do not support the hypothesis that a defect in DHT generation contributes to weight loss in HIV infected men independently of changes in testosterone levels; it is possible that such a defect might exist in HIV-infected men with more severe weight loss. PMID- 10520574 TI - Effect of N-acetylsphingosine (C2) and the ceramidase inhibitor (1S,2R)-D-erythro 2-(N-myristoylamino)-1-phenyl-1-propanol on the regulation of Sertoli cell function. AB - In the present study, a possible role of ceramide in the regulation of Sertoli cell function was investigated. Intracellular ceramide levels were increased by adding N-acetylsphingosine (C2) or by inhibiting ceramidase with (1 S,2R)-D erythro-2-(N-myristoylamino)-1-phenyl-1-propanol (MAPP). Cultured Sertoli cells were stimulated for 3 days with different doses of C2, MAPP, and their corresponding inactive analogs. The effect of these drugs was evaluated along four well-known Sertoli cell parameters: lactate and transferrin secretion, gamma glutamyl transpeptidase (gamma-GTP) activity, and estradiol production. C2 and MAPP increased lactate production and decreased transferrin secretion. The inactive analogs did not produce any effect. In FSH (follicle-stimulating hormone)-stimulated cultures, C2 and MAPP produced a further increment in lactate production and decreased FSH-stimulated transferrin secretion. No effect was observed under basal or FSH-stimulated gamma-GTP activity, and both treatments decreased estradiol production in response to FSH. Results obtained in dbcAMP (dibutyryladenosine 3':5'-cyclic monophosphate)-stimulated cultures suggest that the observed effects of ceramide on transferrin secretion are secondary to a decrease in cAMP production, whereas the effects of ceramide on lactate and estradiol productions are post-cAMP formation regulatory events. In summary, our results show that ceramide can regulate Sertoli cell function. Similar to what has been observed for other signaling molecules, ceramide can interact with the FSH-dependent pathway, but the precise steps involved in this interaction are still unknown. PMID- 10520575 TI - The development of male reproductive organ abnormalities after neonatal exposure to tamoxifen is genetically determined. AB - Responses of the male reproductive organs to neonatal exposures of tamoxifen (Tx) were examined in seven different strains of mice (A/J, AKR/J, BALB/cAnN, C3H/HeJ, C57BL/6J, DBA/ 2J, and FVB/N). Male mice were given daily subcutaneous injections of 2 microg Tx from postnatal day 1 to 5, and the testes, epididymides, ductus deferens, and seminal vesicles were examined at 3 months of age. At necropsy, the testes and seminal vesicles of Tx-treated groups were significantly smaller in size than those of the control groups in all strains. Histologically, the testes of AKR/J, BALB/cAnN, and FVB/N mice showed no abnormality after neonatal treatment with Tx. In contrast, the testes of A/J, C3H/HeJ, C57BL/6J, and DBA/ 2J mice were often necrotic and highly disorganized, with severe inflammation. In the connective tissue surrounding these testes, relatively large arteries were involved in the inflammation, and the vascular lumen was occluded by the thickened tunica interna, suggesting that these testicular changes were due to infarction. Similarly, the epididymides and ductus deferens of Tx-treated A/J, C3H/HeJ, C57BL/6J, DBA/2J, and FVB/N mice showed chronic pyogranulomatous inflammation. The epithelium of the seminal vesicles of Tx-treated A/J, C3H/HeJ, C57BU6J, DBA/2J, and FVB/N mice also exhibited moderate hyperplasia, with squamous metaplasia. These results indicate that neonatal exposure to Tx causes various abnormalities of the male reproductive organs in postpubertal mice, depending on the strains, and suggest that a genetic component plays a major role in determining the phenotypic variation observed. PMID- 10520576 TI - Construction and preliminary characterization of a series of mouse and rat testis cDNA libraries. AB - We have constructed a series of 23 cDNA libraries from mouse and rat testicular cells. These include libraries made from whole, intact adult testes; seminiferous tubule cells from adult testes; combined populations of primary spermatocytes from 18-day-old mouse testes; and isolated populations of primitive type A spermatogonia, type A spermatogonia, type B spermatogonia, preleptotene spermatocytes, leptotene plus zygotene spermatocytes, juvenile pachytene spermatocytes, adult pachytene spermatocytes, round spermatids, Sertoli cells from 6-, 8-, 17-, and 18-20-day-old mice, and peritubular cells from 18-20 day old mice, all recovered from outbred white Swiss (CD-1) mice. We also constructed libraries from whole adult testes from five other lines of mice: C57 Bl6/J, C3 HEB, BDF-1, Balb/c, and 129 Sv. Finally, there are two libraries made from populations of Sertoli cells and peritubular cells isolated from testes of 20-day old Sprague-Dawley rats. Enzymatic dissociation, followed by gradient separation or plating/lysing techniques, was used to prepare populations of specific cell types in purities of 85-98%. cDNAs were synthesized from poly A+ mRNA primed with oligo dT and unidirectionally cloned into the lambda Uni-Zap XR expression vector from Stratagene. Primary titers ranged from 2.1 x 10(5) to 2.9 x 10(8) plaque forming units, and insert sizes averaged 1.0-1.2 kb. These libraries have been amplified once and submitted to the American Type Culture Collection (ATCC) for distribution to interested investigators. ATCC accession numbers are provided. PMID- 10520578 TI - In vitro sperm-binding assay to distinguish differences in populations of human sperm or damage to sperm resulting from cryopreservation. AB - Annually, >1.3 million men are members of couples seeking help because of infertility. Semen from many of these men contains reasonable numbers of motile and normal sperm, but for a subset of individuals, many sperm are deficient in ability to bind to the zona pellucida during in vitro fertilization. Diagnosis of this defect has been hampered by lack of a low-cost test. Molecular similarity exists between the perivitelline membrane of a hen's egg and the mammalian zona pellucida. These facts and some preliminary data led to evaluation of binding of human sperm during incubation for 60 minutes at 37 degrees C to an extract of chicken perivitelline membrane coated in microwell assay plates. The sperm binding assay had inter- and intraassay plate variations of 21 and 12%, respectively, using washed fresh sperm. All seminal samples were normal, except a few that had 36 to 50% motile sperm with a low rate of sperm movement (if there is a low rate of movement, World Health Organization [WHO] criterion for normalcy is >50% motile). Nevertheless, this sperm-binding assay detected differences among individuals in percentage of sperm bound. Based on data for two to four ejaculates from each of eight occasional sperm donors, the coefficient of variation for ejaculates within donor averaged 31%, and means for the donors differed (P < 0.02). Percentage of sperm bound ranged from <1 to 38% for fresh semen from 57 men and from <1 to 13% for frozen-thawed semen from 34 men. Percentage of motile sperm accounted for <30% of the variation in percentage of sperm bound. In a direct comparison based on 17 ejaculates, aliquots evaluated fresh averaged 13% sperm bound, versus 2% for frozen-thawed aliquots. We concluded that the egg membrane substrate used in these microwell assay plates might serve as the basis for a diagnostic assay. However, it remains to be established whether samples of human semen with a low percentage of sperm binding indeed have relatively low fertilizing potential. PMID- 10520577 TI - Presence and tyrosine phosphorylation of c-met receptor in human sperm. AB - The c-met receptor is a p190MET tyrosine kinase proto-oncoprotein that through its binding to its ligand, designated hepatocyte growth factor (HGF), induces mitogenic, motogenic, and morphogenic activities in a variety of cell types. The present study was conducted to examine whether or not the c-met receptor is expressed and tyrosine phosphorylated in the human sperm cell. The Western blot analysis, using a monoclonal antibody (MAb2) directed against the extracellular domain of the c-met receptor, showed a specific band of 195 kDa corresponding to the intact c-met receptor in the detergent-solubilized human sperm preparation (HSP). This protein band was not recognized by the control myeloma lg (immunoglobin). In the immunoprecipitation procedure, a similar specific band of 195 kDa and a 145-kDa band corresponding to the beta-subunit of c-met receptor were seen. In the indirect immunofluorescence technique, the c-met receptor was localized predominantly in the acrosomal region of the sperm cell. The c-met receptor was tyrosine phosphorylated/autophosphorylated during capacitation and in the cell-free in vitro kinase assay. Incubation of human sperm with hepatocyte growth factor (HGF) or MAb2 to c-met receptor enhanced the degree of tyrosine phosphorylation/autophosphorylation of the c-met receptor up to 5.1-fold. These findings indicate that the c-met receptor is present in the acrosomal region of human sperm cell and is tyrosine phosphorylated, which is enhanced by HGF and the receptor antibody. The c-met system may have an important role in sperm function. PMID- 10520580 TI - Assessment of bacterial community structure in soil by polymerase chain reaction and denaturing gradient gel electrophoresis. AB - Bacterial community structure was studied in a Flevo silt loam (FSL) soil microplot, as well as in 15 other soils, by using DNA extraction followed by molecular fingerprinting. Total community DNA was extracted and purified by a direct method, which yielded amplifiable DNA of high molecular weight for all soils. A variable region of the 16S rRNA gene was then amplified by PCR with bacterial primers, resulting in a mixture of amplicons separable via denaturing gradient gel electrophoresis (DGGE). The DGGE profiles of FSL soil were indicative of dominant soil bacterial types, as evidenced by assessing the amplification of Enterobacter cloacae and Arthrobacter sp. targets in a soil DNA background. These targets produced barely detectable bands when present in soil DNA at roughly 5 x 10(6) genome equivalents per g dry soil, and strong bands at 27-fold higher levels. The PCR-DGGE analysis of the FSL soil was highly reproducible. Furthermore, different single versus composite topsoil samples yielded similar DGGE profiles with respect to major bands. In addition, samples taken along vertical soil cores (0-45 cm depth) revealed relative stability of the DGGE profiles. The profiles produced with DNA obtained from different aggregate size fractions of this soil were also similar with respect to the main bands. Moreover, FSL topsoil samples taken over a 1-year period (fallow soil) yielded stable profiles. These data suggested that the soil bacterial communities thus determined were dominated by a limited number of stable and ubiquitous types. The 16 soils, representing varying types and geographical locations, were assessed for differences in their bacterial DGGE profiles. There were striking differences between the profiles obtained for these soils. Evidence was found for the hypothesis that similar soil types tend to contain similar structures of the dominating bacterial types as revealed by the DGGE profiles. PMID- 10520579 TI - Increased in vitro binding of fresh and frozen-thawed human sperm exposed to a synthetic peptide. AB - Prosaposin is a well-characterized, approximately 68-kDa protein found in many tissues and as a normal component of human semen. A fragment of prosaposin apparently is involved in primary sperm-egg binding. We hypothesized that binding of sperm from some men to egg investments would be increased by in vitro exposure of their sperm to a synthetic fragment of human prosaposin (FertPlus peptide). Hence, we evaluated samples of washed fresh or frozen-thawed human sperm after a 10-minute exposure to synthetic FertPlus peptide at 0 (control), 80, 160, 320, 640, or 1280 pM, followed by 1:50 dilution for evaluation of binding. The criterion of response was mean percentage of sperm bound to a substrate prepared from chicken egg membranes after sperm were incubated for 60 minutes at 37 degrees C in substrate-coated wells of a sperm-binding assay plate. For each seminal sample, data were normalized against the percentage of sperm bound for control aliquots, providing values for relative binding. With fresh sperm, relative binding was increased (P < 0.01) by exposure of sperm to peptide, and the effect was especially obvious at 1280 pM. Higher doses were not tested. Collectively at three study sites, exposure of fresh sperm to 1280 pM peptide substantially increased (above 99% confidence interval; on the basis of duplicate control samples) percentage of sperm bound for 25 of 74 (34%) samples. For frozen thawed sperm, exposure to 1280 pM peptide increased binding for 29 of 65 (45%) samples. We concluded that for >30% of men, exposure of their sperm to this synthetic fragment of prosaposin at 1280 pM increased binding of sperm to an egg membrane substrate similar to that offered by the zona pellucida. PMID- 10520581 TI - Quantitation of bacterial adherence by image analysis. AB - In studies of the adherence of pathogenic bacteria to host eukaryotic cells in vitro, the counting of the bacteria is often challenging, especially if many experiments are involved. We developed a method to use digital imaging and computer-aided recognition for the quantitation of bacteria attached to cultured cells. We employed an immunocytochemical method to stain the bacteria and leave the hosts cells relatively unstained. We describe this method for use with five species of bacteria, Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis, Staphylococcus aureus, and Chlamydia pneumoniae. To demonstrate an application of this method, we studied the attachment of H. influenzae and S. pneumoniae to target epithelial cell lines derived from the respiratory tract. PMID- 10520582 TI - Novel method for screening bacterial colonies for phosphatase activity. AB - Current methods for screening large numbers of bacterial colonies for phosphatase activity, rely heavily on the use of colorimetric assays. While such methods have been applied extensively in the laboratory, they are not without their drawbacks. We here describe a precipitating fluorescent probe that can be used to screen phosphatase activity in bacterial colonies. This probe can be incorporated directly into agar plates used to culture the organisms of interest. The approach offers several advantages over current methodologies including the ability to monitor the development of phosphatase activity with colony development, and the ability to distinguish between activity arising from cell-bound and cell-free enzyme. This enzyme probe was successfully used to detect and isolate phosphatase producing bacteria from activated sludge. PMID- 10520583 TI - Automatic enumeration of adherent streptococci or actinomyces on dental alloy by fluorescence image analysis. AB - The aim of the present study was to develop an automated image analysis method to quantify adherence of Streptococcus sanguinis or Actinomyces viscosus on surfaces of a currently used dental alloy. Counting such bacterial strains was difficult because of their arrangement, thus S. sanguinis being a coccus arranged in chains or pairs, and A. viscosus a long complexly arranged polymorph rod. Direct counting of fluorescently stained adherent bacteria was done visually and with image analysis methods. To differentiate these two morphotypes, two programs were developed: (i) for streptococci, thresholding and selection of the object maxima, and (ii) for actinomyces, two step thresholding and processing of the characteristic points of the object skeletons. The triplicate enumerations for each bacterial strain were not significantly different (p > 0.005) and correlations between visual counting and automated counting were significant (r = 0.91 for S. sanguinis and r = 0.99 for A. viscosus, p <00.0001). These rapid and reproducible methods, allowed us to count either cocci or rods, adherent on an inert substratum, in high density conditions. PMID- 10520584 TI - Combination of biomolecular and stable isotope techniques to determine the origin of organic matter used by bacterial communities: application to sediment. AB - Natural isotopic composition is a good tool to trace organic matter in ecosystems. Recent studies used a combination of molecular and stable isotope techniques to determine the origin of the organic carbon used by bacteria in the water column. In our study, we show that this procedure can be used for analysis of sediment bacterial communities with few modifications. In the water column, bacterial recovery is done before DNA extraction. In the sediment, we tested qualitatively and quantitatively a direct and indirect extraction of DNA. The direct extraction was the most efficient. It recovered between 3.1 and 15.8 microg DNA g(-1) dry sediment and the contamination of field samples by eucaryotic DNA was less than 13%. In this preliminary study of the salt marsh ecosystem, the delta(13)C values of DNA (-26 to - 24%) recovered from the sediment were close to the delta(13)C values of halophytic plants (-26.4 and - 25.3%) showing a relationship between plants and microorganisms. Thus, this procedure can be used to trace the flow of carbon through the sediment microbial biomass and to understand the variation of bacterial activity according to the inputs of allocthonous and autochtonous organic matter. PMID- 10520585 TI - Amplification of fluorescently labelled DNA within gram-positive and acid-fast bacteria. AB - Representative organisms from a variety of Gram-positive genera were subjected to varying regimes in order to optimise the intracellular amplification of DNA. The bacteria were subjected to treatments with paraformaldehyde, muramidases and mild acid hydrolysis to discover which regime made each organism permeable to the amplification reagents yet allowed retention of the fluorescein-labelled amplified products within the cell. Scanning electron micrographs were used to corroborate the effectiveness of the treatments, as seen by fluorescent photomicrographs, with the damage caused to the bacterial walls. A combination of mutanolysin and lysozyme was found most effective for Bacillus cereus, whereas permeabilisation of Streptomyces coelicolor, Lactococcus lactis and Clostridium sporogenes was most effective when exposed to lysozyme only. Surprisingly, direct amplification with no pre-treatment gave the brightest fluorescence in Mycobacterium phlei. Comparing the techniques of whole cell PCR, primed in situ labelling (PRINS), and cycle PRINS showed that under the conditions used the strongest intensity of fluorescence was obtained with in situ PCR; only L. lactis and M. phlei produced signals with cycle PRINS, fluorescence was not seen for any of the organisms with PRINS. PMID- 10520586 TI - Convenient and versatile DNA extraction using agarose plugs for ribotyping of problematic bacterial species. AB - We describe a convenient, versatile and safe method for preparing bacterial DNA for ribotyping analysis. In this method, extraction of bacterial DNA from Salmnonella typhi and Burkholderia pseudomallei. and subsequent restriction endonuclease digestion, was performed in agarose blocks/plugs thus minimizing shearing and loss of DNA, problems commonly associated with liquid phase phenol extraction. Digested DNA in the plugs was then electrophoresed directly, transferred to nylon membranes and hybridized with labeled rDNA probes in the usual manner to provide reproducible restriction patterns. This method is particularly useful for bacterial species where standard DNA extraction in the liquid phase using phenol has been problematic (e.g. B. pseudomallei) but can be used for any bacterial species. The DNA extracted within the agarose plugs can be stored for long periods and can be used in other, widely-used typing methods such as pulsed-field gel electrophoresis (PFGE) and PCR-based techniques. Embedding live cells directly in agarose plugs also minimizes the risk of exposure to these virulent human pathogens among laboratory workers. PMID- 10520587 TI - An immunochemical in situ approach to detect adaptation processes in the photosynthetic apparatus of diatoms of the Wadden Sea sediment surface layers. AB - Intertidal Wadden Sea sediment surface layers located near the North Sea shore at Dangast (Germany) were subjected to quantitative chlorophyll and protein extractions followed by SDS-PAGE and Western immunoblotting. During the study, benthic diatoms were almost exclusively the only group of microphytobenthos in this area performing oxygenic photosynthesis. Three successive extractions with 90% acetone yielded more than 98% of the extractable pigments. The absorption spectra of the extracts of sediment samples were nearly identical to those obtained from the diatom Phaeodactylum tricornutum. Ten repetitive extractions with SDS containing loading buffer used for SDS-PAGE ensured that more than 98% of the extractable protein was recovered. Subsequent Western immunoblotting with an antiserum directed against the subunits of the main light harvesting complex of the diatom Cyclotella cryptica showed that the antiserum immunodecorated selectively subunits of diatomaceous light harvesting complexes. This finding demonstrated that a taxon specific class of polypeptides could be visualized and quantified directly in sediment samples. In shading experiments, shaded sedimient areas generally revealed higher amounts of light harvesting subunits which could be immunodecorized. The improved mnethodological approach and the results are discussed in the context of the current development of direct molecular methods for the investigation of activities and adaptation processes of specific groups of microorganisms in their natural habitats. PMID- 10520588 TI - Nucleic acid sequence-based amplification (NASBA) detection of medically important Candida species. AB - Nucleic Acid Sequence Based Amplification (iNASBA), an isothermal amplification technique for nucleic acids, was evaluated for the identification of medically important Candida species using primers selected from 18S rRNA sequences conserved in fungi. An RNA fragment of 257 nucleotides was amplified for Candida albicans. Nineteen different fungi were tested for rRNA amplification with the NASBA. All were positive when analyzed on agarose gel, whereas human RNA was negative. For the identification of Candida species, NASBA amplification products were analyzed in an enzyme bead-based detection format, using species-specific biotinylated probes and a generic Candida HRPO probe or a membrane-based system using biotinylated probes and avidin-HPRO. Discrimination of the major human pathogenic Candida spp. was based on a panel of biotinylated probes for C. krusei, C. tropicalis, C. albicans, C. glabrata, and C. lusitaniae. Using rRNA dilutions obtained from pure cultures of C. albicans, the combination of NASBA and the enzymatic bead-based detection yielded a sensitivity equivalent to 0.01 CFU. In a model system using 1 ml of artificially contaminated blood as few as 1 10 CFU of C. albicans could be detected. Testing of 68 clinical blood samples from patients suspected of candidemia showed that eight samples were positive for C. albicans and one for C. glabrata. Testing of 13 clinical plasma samples from patients suspected of fungemia identified the presence of C. albicans in two specimens. The whole procedure of sample preparation, amplification and identification by hybridization can be performed in 1 day. This speed and the observed sensitivity of the assay make the NASBA a good alternative to PCR for the detection of candidemia. PMID- 10520589 TI - Effects of storage on measurements of potential microbial activities in stream fine benthic organic matter. AB - Sample storage can significantly influence measured microbial activities in stream fine benthic organic matter (FBOM), possibly confounding effects of sample variability and short-term changes in activity. Denitrification potential, acetylene reduction and respiration rates, mineralizable N and extractable ammonium concentrations, and beta-glucosidase and phosphatase enzyme activities of FBOM from first-order mountain streams in the western Oregon Cascade Mountains were assayed at various times after collection to determine potential storage effects. Denitrification potential, phosphatase activity, and extractable ammonium remained stable over a minimum of 11 h of storage at 5 degrees C. Mineralizable N concentrations, respiration rates, and beta-glucosidase activity all decreased within 12 h of collection. Results varied for acetylene reduction. Once assay conditions were established, denitrification potential and respiration rates were linear with incubation time. Based on paired t-tests, measures of acetylene reduction, denitrification potential, respiration rate, beta glucosidase activity, and phosphatase activity were generally similar at a 1-wk interval within the same stream reaches. PMID- 10520590 TI - Evaluation of two automated enzyme-immunoassays for detection of thermophilic campylobacters in faecal samples from cattle and swine. AB - We evaluated the performance of two enzyme-immunoassays (EIA) for the detection of naturally occurring, thermophilic Campylobacter spp. found in faecal samples from cattle (n = 21 and n = 26) and swine (n = 43) relative to the standard culture method, and also assuming that none of the tests was the definitive standard. The primary isolation both for the culture and the EIA methods was carried out by overnight selective enrichment in Preston broth. The results showed good sensitivities for both EIA methods in cattle (95% and 84%) and swine (88% and 69%) samples. However, when testing cattle samples, EIA-2 method resulted in a rather low specificity (32%). This seemed to be partially due to the isolation of nonthermophilic species. In conclusion, EIA-1 method may provide a simple and fast tool with good accuracy in cattle and swine samples for automated screening of large number of samples. PMID- 10520591 TI - Development of an agar lift-DNA/DNA hybridization technique for use in visualization of the spatial distribution of Eubacteria on soil surfaces. AB - While microbial growth is well-understood in pure culture systems, less is known about growth in intact soil systems. The objective of this work was to develop a technique to allow visualization of the two-dimensional spatial distribution of bacterial growth on a homogenous soil surface. This technique is a two-step process wherein an agar lift is taken and analyzed using a universal gene probe. An agar lift is comprised of a thin layer of soil that is removed from a soil surface using an agar slab. The agar is incubated to allow for microbial growth, after which, colonies are transferred to a membrane for conventional bacterial colony DNA/DNA hybridization analysis. In this study, a eubacterial specific probe was used to demonstrate that growing bacterial populations on soil surfaces could be visualized. Results show that microbial growth and distribution was nonuniform across the soil surface. Spot supplementation of the soil with benzoate or glucose resulted in a localized microbial growth response. Since only growing colonies are detected, this technique should facilitate a greater understanding of the microbial distribution and its response to substrate addition in more heterogenous soil systems. PMID- 10520592 TI - PCR-based quantitation of Cryptosporidium parvum in municipal water samples. AB - A PCR method for the quantitation of Cryptosporidium parvum oocysts in municipal drinking water samples was investigated. Quantitative PCR uses an internal standard (IS) template with unknown target numbers to compare to standards of known concentrations in a standard curve. The IS template was amplified using the same primers used to amplify a portion of a 358 bp gene fragment that encodes a repetitive oocyst wall protein in C. parvum. Municipal water samples spiked with known numbers of C. parvum oocysts were tested by quantitative PCR using the IS and the Digene SHARP Signal System Assay for PCR product detection. The absorbance readings for target DNA and IS templates versus the number of molecules of the target DNA were plotted to generate standard curves for estimating oocyst numbers. The method allowed the quantitation of oocysts from log 3 to log 5 spiked into municipal water samples. PMID- 10520593 TI - Flow cytometric application of the Sabin and Feldman dye test in the diagnosis of toxoplasmosis. AB - Although time-consuming and requiring live parasites, the Sabin and Feldman dye test (DT) is still considered the 'gold standard' among the serological tests for toxoplasmosis diagnosis. The present study was initiated to compare detection of dead parasites using optical microscopy with flow cytometry and a fluorescent nonvital dye, propidium iodide. After incubation with sera (N = 150) and a complement source, tachyzoites were washed, then stained using a fluorescein conjugated Toxoplasma-specific antiserum. Dead tachyzoites were detected by flow cytometry after addition of propidium iodide. Intra- and inter-assay reproducibilities of percentages of dead parasites varied between 7 and 14%, and 8 and 21%, respectively. When comparing flow cytometry with the classical DT, no discrepancy was noted for positive (N = 118) and negative sera (N = 32). Correlation was good (r = 0.85) for positive sera. In conclusion, when easily available, flow cytometry is a very sensitive, specific and time-sparing method to detect specific antibodies to Toxoplasma gondii. PMID- 10520594 TI - Estimation of temperature dependent growth rate and lag time of Listeria monocytogenes by optical density measurements. AB - An automated turbidimetric system, Bioscreen C, was used to monitor growth of ten strains of Listeria monocytogenes at different temperatures. Several methods for estimation of maximum specific growth rate (mu(max)) and lag time (lag) from turbidimetric data were compared to values estimated from viable count data. By using a calibration factor, reliable estimations of mu(max) could be obtained from turbidimetric measurements. On the other hand, accurate estimations of lag required some viable count data. PMID- 10520595 TI - Assessment of a magnetic bead-EIA based kit for rapid diagnosis of fish pasteurellosis. AB - The accuracy of the magnetic beads-EIA based BIONOR AQUAEIA-Pp kit for the rapid diagnosis of pasteurellosis was evaluated. The kit reacted with all the Photobacterium damselae subsp. piscicida strains included in this study, with a detection limit of 10(4) bacteria/ml. However, non-specific reactions were observed with isolates of Ph. damselae subsp. damselae or Ph. histaminum when the bacterial concentration was high (10(9)-10(10) bacteria/ml). Similar findings in specificity and sensitivity were observed when the kit was applied to experimentally infected fish tissues. However, since those bacterial species are not usually found in the fish species susceptible to pasteurellosis, the AQUAEIA kit appears applicable for a rapid screening of the disease. In addition, when the kit was utilized to analyze cultured populations of seabream, it allowed the detection of the pathogen, not only in individuals affected by the disease, but also in asymptomatic carrier fish. Furthermore, the positive detection of Ph. damselae subsp. piscicida in broodstock gonads, seminal, and ovaric fluids, and also in eggs indicated the possibility of vertical transmission of pasteurellosis. PMID- 10520596 TI - Standardisation of restriction fragment length polymorphism analysis for Mycobacterium avium subspecies paratuberculosis. AB - DNA from 1008 strains of Mycobacterium avium subspecies paratuberculosis, digested by restriction endonucleases PstI and BstEII, was hybridised with a standard IS900 probe prepared by PCR and labelled non-radioactively by ECL. DNA fingerprints were scanned by CCD camera and analysed using the software Gel Compar (Applied Maths, Kortrijk, Belgium). Thirteen restriction fragment length polymorphism (RFLP) (PstI) types were detected, which where designated as A, B, C, D, E, F, G, H, I, J, K, L and M in accordance with the study of Pavlik et al. (1995) [Pavlik, I., Bejckova, L., Pavlas, M., Rozsypalova, V., Koskova, S., 1995. Characterization by restriction endonuclease analysis and DNA hybridization using IS900 of bovine, ovine, caprine and human dependent strains of Mycobacterium paratuberculosis isolated in various localities. Vet. Microbiol. 45, 311-318]. Twenty RFLP (BstEII) types were detected and designated as C1-3, C5, C7-20, S1 and I1 in accordance with the study by Collins et al. 1990 [Collins, D.M., Gabric, D.M., de Lisle, G.W., 1990. Identification of two groups of Mycobacterium paratuberculosis strains by restriction endonuclease analysis and DNA hybridization. J. Clin. Microbiol. 28, 1591-1596]. A combination of both RFLP (PstI) and RFLP (BstEII) results revealed a total of 28 different RFLP types. All the RFLP types and detailed protocols are available at Intemet web site WWW...: http:/ /www.vri.cz/wwwrflptext.htm. PMID- 10520597 TI - Optimization of the growth conditions of the extremely thermophilic microorganisms Thermococcus celer and Pyrococcus woesei. AB - Growth medium components and cultivation conditions for the extremely thermophilic Archaea Thermococcus celer and Pyrococcus woesei were optimized. A culture media based in marine water was formulated. Both Archaea demonstrated to be strictly anaerobic with optimal growth temperature of 85 degrees and 95 degrees C, respectively. Sodium sulfide, but not cysteine, was used as a sulfur and reductive capacity source. It was observed that hydrogen sulfide could be replaced by 30 microM titanium (III) nitrile acetate. The addition of elemental S(o) enhanced growth of both microorganisms, with T. celer far more sensitive than P. woesei to the absence of S(o). P. woesei utilized maltose as a carbon source, while T. celer was able to use only peptides from yeast extract, peptone and tryptone as its carbon source. Optimum carbon source concentrations were 1.25 g/L for T. celer and 5 g/L for P. woesei. Although both Archaea required peptides as a nitrogen source, the addition of ammonia chloride to a nitrogen-limited media did not stimulate growth, which suggests that neither Archaea appear to metabolize ammonia. The growth of P. woesei, but not T. celer, was stimulated considerably in the presence of iron. Co, Ni, Zn, Mo. Mn and Mg were essential trace elements needed for optimal growth of both bacteria. PMID- 10520598 TI - Regulation of eukaryotic DNA replication and nuclear structure. AB - In eukaryote, nuclear structure is a key component for the functions of eukaryotic cells. More and more evidences show that the nuclear structure plays important role in regulating DNA replication. The nuclear structure provides a physical barrier for the replication licensing, participates in the decision where DNA replication initiates, and organizes replication proteins as replication factory for DNA replication. Through these works, new concepts on the regulation of DNA replication have emerged, which will be discussed in this minireview. PMID- 10520599 TI - The alphaMbeta2 integrin and its role in neutrophil function. AB - Neutrophils are the first cell type to arrive at the injury sites and play a critical role in host defense, by virtue of its ability to adhere and transmigrate through endothelium, to phagocytose foreign pathogens, and to produce free oxygen radicals and proteolytic enzymes. Yet, inappropriate neutrophil activation causes tissue damage and various inflammatory diseases. These physiological and pathological functions of neutrophils depend on the engagement of certain surface receptors, especially alphaMbeta2, the major beta2 integrin receptor present on neutrophil surface. Understanding of the molecular mechanisms underlying ligand binding by alphaMbeta2, as well as the roles of alphaMbeta2-ligand interactions in neutrophil functions will enable us to regulate more precisely neutrophil activities: that is, to promote their host defense functions, and at the same time to minimize their deleterious effects on normal cells. PMID- 10520600 TI - The Notch-Hes pathway in mammalian neural development. AB - A wide variety of neurons and glial cells differentiate from common precursor cells in the developing nervous system. During this process, Notch-mediated cell cell interaction is essential for maintenance of dividing cells and subsequent generation of cell type diversity. Activation of Notch inhibits cellular differentiation, and abnormality of the Notch pathway leads to premature neuronal differentiation, the lack of some cell types, and severe defects of tissue morphogenesis. Recent data demonstrate that Notch fails to inhibit cellular differentiation in the absence of the bHLH genes Hes1 and Hes5, which functionally antagonize the neuronal bHLH genes such as Mash1. These results indicate that the two Hes genes are essential effectors for the Notch pathway and that neuronal differentiation is controlled by the pathway "Notch-Hes1/Hes5 |Mash1". PMID- 10520601 TI - Expression of a soluble form of CTLA4 on macrophage and its biological activity. AB - Interaction between cytotoxic T lymphocyte-associated antigen-4 (CTLA4, CD152) and B7 molecules (B7-1 and B7-2) is of importance in the cellular events of lymphocyte, including antigen-specific T-cell activation and induction of autoreactive T-cell. We describe here the first introduction of a murine soluble CTLA4 gene, CTLA4Ig, to Mm1 cells, a macrophagic cell line. CTLA4Ig was successfully expressed on Mml cells and the expressed CTLA4Ig was found to be functionally active in their binding to B7 molecules by flow cytometry and immunofluorescence studies. The biological activity of CTLA4Ig from the transfected Mm1 cells was studied and showed inhibitory activity on mixed lymphocyte culture. A high CTLA4Ig producing macrophagic cell line was obtained. As Mm1 cells were regarded as difficult for gene transfection and there had so far been no report on expression of CTLA4Ig gene on Mm1 cells, these results suggested that the CTLA4Ig expressing Mm1 cells could be useful for analysis of CTLA4 and B7 molecule interaction in both macrophage and T-cell. PMID- 10520602 TI - The developmental fate of green fluorescent mouse embryonic germ cells in chimeric embryos. AB - Primordial germ cells (PGCs), as precursors of mammalian germ lineage, have been gaining more attention as a new resource of pluripotent stem cells, which bring a great possibility to study developmental events of germ cell in vitro and at animal level. EG4 cells derived from 10.5 days post coitum (dpc) PGCs of 129/svJ strain mouse were established and maintained in an undifferentiated state. With an attempt to study the differentiation capability of EG4 cells with a reporter protein: green fluorescence protein, and the possible application of EG4 cells in the research of germ cell development, we have generated several EG4-GFP cell lines expressing enhanced green fluorescence protein (EGFP) and still maintaining typical characteristics of pluripotent stem cells. Then, the differentiation of EG4-GFP cells in vitro as well as their developmental fate in chimeric embryos which were produced by aggregating EG4-GFP cells to 8-cell stage embryos were studied. The results showed that EG4 cells carrying green fluorescence have a potential use in the research of germ cell development and other related studies. PMID- 10520603 TI - Cloning and analysing of 5' flanking region of Xenopus organizer gene noggin. AB - Xenopus organizer specific gene Noggin possesses nearly all the characteristic properties of the action of organizer to specify the embryonic body axis. To analyze how the maternal inherited factors control its expression pattern, we cloned the 5' regulatory region of noggin gene. The 1.5 kb upstream sequence could direct reporter gene to express in vivo and data from deletion analysis indicated that a 229 base pair fragment is essential for activating noggin expression. We further demonstrated that the response elements within this regulatory region were indeed under the control of growth factor activin and Wnt signaling pathway components. PMID- 10520604 TI - Random amplified polymorphic DNA analysis of eel genome. AB - Eel family is a huge one, in which many kinds of eels especially some migratory eels, bear strong resemblance to each other, and are therefore difficult to be identified. In this study 29 random primers were used to make RAPD analysis for Japanese eel (Anguilla japonica), European eel (Anguilla anguilla) and Pike eel (Muraenesox cinereus). And totally 299 fragments were counted. Shared or specific fragments were counted and genetic similarity or genetic distance were calculated. The genetic similarity between Japanese eel and Pike eel is 0.68 and the genetic distance between them is 0.32; those between European eel and Pike eel are 0.72 and 0.28 respectively, and between Japanese eel and European eel are 0.74 and 0.25 respectively. The method has been shown to be suitable to molecular identification of eels. It provides an alternative approach to determine the relationship between species. PMID- 10520605 TI - Retrovirus-mediated herpes simplex virus thymidine kinase gene therapy approach for hepatocellular carcinoma. AB - The therapeutic effect of herpes simplex virus thymidine kinase/ganciclovir (HSV tk/GCV) system on hepatocellular carcinoma was studied in this experiment. The tk containing retroviral recombinants were used to infect hepatoma cells (BEL-7402) and the cells were treated with ganciclovir (0-1000 microg/ml). The results showed that HSV-tk gene could be efficiently transferred in vitro into hepatoma cells and stably expressed. The growth potential of the tk-containing cells was significantly inhibited by GCV (P<0.01) as compared to the non-tk-containing cells. The antitumor effect of HSV-tk/GCV system was also produced ex vivo in tk containing tumor of nude mice as characterized by a marked decrease in tumor growth after GCV treatment contrary to a progressive enlargement of non-tk containing tumors. Although the histological examination demonstrated that the efficiency of the gene transfer was less than 30%, the killing effect of HSV tk/GCV system on hepatocellular carcinoma was still significantly generated. The proper mechanism of HSV-tk gene therapy on hepatic tumor referred as "bystander effect" in therapeutic approach has not been found in this study and required to be explored further. PMID- 10520606 TI - Involvement of gene expressions in apoptosis of vascular endothelial cells induced by rattlesnake venom. AB - Formation of apoptotic bodies is a typical character of apoptotic cell death, but how the processes are controlled is not known. In this study, we compared two apoptosis inducing systems in vascular endothelial cells (VEC). We found that the formation of apoptotic bodies during apoptosis induced by rattlesnake venom, which is an unique and specific apoptosis inducer to vascular endothelial cells, was much faster than that induced by deprivation of survival factors (aFGF and serum). When we blocked the synthesis of mRNAs in cells treated with rattlesnake venom by DRB (5, 6-dichloro-1-beta-D-ribofuranosylbenzimidazole), an inhibitor of transcription, the formation of apoptotic bodies was dramatically inhibited. We examined the expression of p53 gene and found that its expression was much higher in apoptosis induced by rattlesnake venom than that in apoptosis induced by deprivation of aFGF and serum. Our results suggest that gene expression is important and p53 gene may play a major role in inducing the formation of apoptotic bodies in VEC. PMID- 10520607 TI - The obligation of researchers to precept. PMID- 10520608 TI - Review of hazards associated with children placed in adult beds. AB - OBJECTIVES: To identify and assess dangers associated with placing children younger than 2 years to sleep in adult beds. This article focuses on overlying, wedging, and strangulation hazards and the relationship of these hazards to children's sleeping environments. DESIGN: A retrospective review and analysis of data collected by the US Consumer Product Safety Commission on deaths of children younger than 2 years in standard adult beds, daybeds, and waterbeds. The review included incident data from January 1990 through December 1997. RESULTS: The 8 year records showed a total of 515 deaths of children younger than 2 years who were placed to sleep on adult beds. Of these deaths, 121 were reported to be due to overlying of the child by a parent, other adult, or sibling sleeping in bed with the child and 394 were due to entrapment in the bed structure. Most of these deaths seem to have resulted from suffocation or strangulation caused by entrapment of the child's head in various structures of the bed. CONCLUSIONS: Placing children younger than 2 years to sleep in adult beds exposes them to potentially fatal hazards that are generally not recognized by the parent or caregiver. These hazards include overlying by a parent, sibling, or other adult sharing the bed; entrapment or wedging of the child between the mattress and another object; head entrapment in bed railings; and suffocation on waterbeds. Parents and caregivers should be alerted to these avoidable hazards. PMID- 10520609 TI - The bedtime pass: an approach to bedtime crying and leaving the room. AB - OBJECTIVE: To evaluate a novel intervention for bedtime problems. DESIGN: We used an ABAB withdrawal-type experimental design. SETTING: The intervention was prescribed in an outpatient primary health care context and evaluated in the home setting. PARTICIPANTS: Two normally developing boys aged 3 and 10 years were the primary participants. Twenty parents and 23 practicing pediatricians rated the acceptability of the intervention. INTERVENTION: A bedtime pass, exchangeable for 1 excused departure from the bedroom after bedtime. MAIN OUTCOME MEASURES: For both primary participants, instances of crying and/or coming out from the bedroom after bedtime; for the 20 parents and 23 pediatricians, comparative ratings of acceptability for the pass and 2 other commonly used approaches to bedtime problems (ignoring crying and letting children sleep with their parents). RESULTS: Crying and coming out from the bedroom reduced to zero rates in both children. Pediatricians rated using the pass as significantly more acceptable than letting children sleep with parents and equivalent to ignoring. Parents rated the pass as more acceptable than either alternative. CONCLUSION: The bedtime pass provides pediatricians with a readily usable, potentially effective, and highly acceptable novel intervention for bedtime problems, one of the most common complaints in outpatient pediatrics. PMID- 10520610 TI - Culture and the care of children with chronic conditions: their physicians' views. AB - BACKGROUND: Little is known about physicians' perceptions of the influence of culture on the health care of children with chronic and disabling conditions. OBJECTIVE: To identify physicians' perceptions of the impact of the family's ethnocultural background on the health care of school-aged children with chronic conditions and recommendations for improving care. DESIGN: Qualitative study in 2 midwestern metropolitan areas. SETTING: General community. PARTICIPANTS: Convenience sample of 52 physicians nominated by 60 African American, Hispanic, and European American families of school-aged children with chronic conditions. METHODS: In-depth interviews were conducted with the physicians. Content analytic techniques were used to analyze the data. RESULTS: In 44% of the responses, the physicians reported that ethnocultural background did not influence the care the child received, noting that comparable care was provided to all of their patients. In 14% of the responses, the effect was unknown. The overall effect was negative in 26% of the responses and positive in 16%. Physicians' recommendations focused on 4 topics: improving the training and education of health care professionals and families; ensuring good communication between the child, family, and health care professionals; supporting families; and improving the access and provision of services for children from diverse cultural backgrounds. CONCLUSION: Although the majority of participants reported that ethnocultural background did not affect the care the child received from the health care system, physicians' recommendations reflected awareness of the influence of culture on the care of children with chronic conditions and the need for further training on this issue. PMID- 10520611 TI - Diagnosis of attention-deficit/hyperactivity disorder and use of psychotropic medication in very young children. AB - CONTEXT: Increases in diagnosis and treatment of attention-deficit/hyperactivity disorder (ADHD) have elicited public and professional concern. Research suggests that this trend warrants the inclusion of previously underdiagnosed children and adults. It is not clear whether this trend includes young children. OBJECTIVE: To identify patterns of diagnosis and treatment of ADHD in very young children over time. DESIGN: Descriptive study of Michigan Medicaid claims data. PATIENTS: Inclusion criteria included recorded ADHD diagnosis, continuous Medicaid eligibility during a 15-month period, and age 3 years or younger at the first date of service. MAIN OUTCOME MEASURES: Diagnoses of ADHD, conditions commonly comorbid with ADHD, other chronic health conditions, and injuries; treatments such as psychological services and psychotropic medication; and the number of ambulatory visits. RESULTS: We identified 223 children aged 3 years or younger diagnosed with ADHD. Many had conditions commonly comorbid with ADHD (44%), other chronic health conditions (41%), and injuries (40%). More than half received psychotropic medication (57%); fewer received psychological services (27%). Twenty-two different psychotropic medications were used. Patterns included more than 1 psychotropic medication (46%) in 30 combinations of simultaneous use and 44 combinations of sequential use. The mean number of ambulatory visits was 18. CONCLUSIONS: Children aged 3 years or younger had ADHD diagnosed and received markedly variable psychotropic medication regimens. Little information is available to guide these practices. The presence of comorbid conditions and injuries attests to these children's vulnerability. Resources must be identified that will enable physicians to better respond to the compelling needs of these children and their families. PMID- 10520612 TI - Beliefs about Papanicolaou smears and compliance with Papanicolaou smear follow up in adolescents. AB - OBJECTIVE: To explore qualitatively adolescent girls' understanding of Papanicolaou smears and barriers to compliance with Papanicolaou smear follow-up appointments. DESIGN: Qualitative analysis, using 3 focus groups and 15 in-depth, semistructured individual interviews. SETTING: Adolescent Clinic and Young Parents' Program at Children's Hospital, Boston, Mass. MAIN OUTCOME MEASURES: Beliefs and attitudes about Papanicolaou smears and barriers to compliance with Papanicolaou smear follow-up. RESULTS: The mean (+/- SD) age of the 15 interview participants was 18.7 (+/- 1.9) years. Knowledge about Papanicolaou smears and pelvic examinations was poor. Most participants believed that their peers receive Papanicolaou smear screening and perceived teenagers to be susceptible to cervical cancer. Perceived benefits to getting Papanicolaou smears were prevention and early detection or diagnosis, and reported barriers included pain or discomfort, embarrassment, fear of finding a problem, fear of the unknown, denial, poor communication or rapport with the provider, not wanting to look for trouble, lack of knowledge, and peers' advice. Participant-generated strategies for how providers could overcome barriers to Papanicolaou smear screening included education and the development of trusting, consistent relationships with providers. Participant-generated strategies for how providers could enhance appointment-keeping among adolescents included telephone and written reminders. CONCLUSIONS: These data support a behavioral theory-based model of adolescent compliance with Papanicolaou smear follow-up, which may help to develop strategies to enhance compliance with Papanicolaou smear follow-up appointments. These strategies include providing in-depth education about Papanicolaou smears, addressing barriers to Papanicolaou smear follow-up, focusing on appropriate provider behaviors, and instituting an appointment reminder system. PMID- 10520613 TI - Parental influence on adolescent sexual behavior in high-poverty settings. AB - BACKGROUND: African American adolescents living in high-poverty urban settings are at increased risk for early sexual initiation and sexually transmitted diseases. OBJECTIVE: To determine whether parental strategies to monitor their children's social behavior and to communicate with them about sexual risks help to reduce the initiation of risky sexual behavior and prevent the resulting adverse health outcomes. METHODS: To assess the viability of these strategies, we surveyed a stratified cross-section of African American children aged 9 to 17 years (N = 355) living in urban public housing. Talking computers were used to increase the confidentiality and comparability of the interviews across the wide age range. RESULTS: Children who reported high levels of parental monitoring were less likely to report initiating sex in pre-adolescence (aged < or = 10 years) and reported lower rates of sexual initiation as they aged. Children who reported receiving both greater monitoring and communication concerning sexual risks were also less likely to have engaged in anal sex. Communication was also positively related to the initiation of condom use and consistent condom use. The protective correlates of these parenting strategies were independent of the type of guardian (mother vs other family member). CONCLUSION: Interventions with parents and other guardians to increase monitoring and communication about sexual risks seem to be promising health-promotion strategies for adolescents in high-risk settings. PMID- 10520614 TI - The type 2 family: a setting for development and treatment of adolescent type 2 diabetes mellitus. AB - OBJECTIVE: To identify physical, behavioral, and environmental features of adolescents (aged 11-17 years) with type 2 diabetes mellitus and their families to define the involvement of known risk factors and to define a profile of at risk individuals. DESIGN AND METHODS: A total of 42 subjects from 11 families with an adolescent in whom type 2 diabetes was previously diagnosed participated. All subjects underwent anthropometric measurement and completed food frequency and eating disorder questionnaires, and were classified according to the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition. In addition, laboratory tests to determine levels of hemoglobin A1c, fasting glucose, C peptide, insulin, and proinsulin were performed. RESULTS: Type 2 diabetes had been diagnosed in 5 of 11 mothers and 4 of 11 fathers before the study. Type 2 diabetes was diagnosed in 3 of the remaining 7 fathers during the study. In 3 families, both parents were affected with type 2 diabetes. As a group, participants were obese, with a body mass index higher than the 95th percentile for probands and fathers, and higher than the 85th percentile for mothers and siblings. The sum of skin fold measurements was above the 95th percentile for the probands, their siblings, and the parents. All groups had high fat intake and low fiber intake. None of the subjects participated in a structured or routine exercise program, and most reported no regular physical activity. Three of the probands met the criteria for binge-eating disorder, and 6 additional patients had notable characteristics of the disorder. Mothers affected with type 2 diabetes had markedly abnormal hemoglobin A1c levels, indicating poor control. There were no group differences in fasting concentrations of insulin, proinsulin, or C peptide. However, a third of the mothers with type 2 diabetes, and all but 1 of the siblings, had evidence of insulin resistance. CONCLUSIONS: Adolescents in whom type 2 diabetes has been diagnosed, as well as their first degree family members, are obese. In addition, the incidence of diagnosed and undiagnosed type 2 diabetes or of insulin resistance in the families of adolescents with type 2 diabetes is striking. Probands and other family members have lifestyles characterized by high fat intake, minimal physical activity, and a high incidence of binge eating. These findings indicate that the families of adolescents with type 2 diabetes share many anthropometric and lifestyle risk factors. The design of treatment programs for adolescents with type 2 diabetes will need to address the lifestyle and health habits of the entire family. PMID- 10520615 TI - Depot medroxyprogesterone acetate use in inner-city, minority adolescents: continuation rates and characteristics of long-term users. AB - OBJECTIVE: To identify continuation rates of depot medroxyprogesterone acetate (Depo-Provera) and characteristics of long-term users in a population of inner city, minority adolescents with high pregnancy rates. DESIGN: Retrospective medical record review. SETTING: An inner-city adolescent clinic and an adolescent pregnancy program. METHODS: A review of the medical records of 250 females aged 13 to 20 years (mean +/- SD, 16.8 +/- 1.1 years), 62.9% Hispanic and 34.2% African American, receiving a first depot medroxyprogesterone acetate injection for contraception between August 1993 and June 1996 was conducted using a standardized form. The mean +/- SD age at menarche of the subjects was 11.6 +/- 1.4 years, and the mean +/- SD age at first intercourse was 14.1 +/- 1.3 years; the mean number of lifetime sex partners was 2.4. Of the subjects, 73.6% had used condoms, 32.0% used oral contraceptives, and none used implants. Of the 201 subjects for whom there were data in the medical records regarding prior fertility, 172 (85.6%) had been pregnant, and 145 (72.1%) had a child. Life table analysis was used to measure depot medroxyprogesterone acetate continuation rates and to compare subgroups of adolescents. RESULTS: Depot medroxyprogesterone acetate continuation rates were found to be 70.3% at 6 months, 48.3% at 9 months, 31.5% at 12 months, and 12.8% at 24 months. The most common reason for depot medroxyprogesterone acetate discontinuation was missed appointments (41.7%). Subjects were followed up for a mean +/- SD of 1.3 +/- 0.7 years after discontinuation of depot medroxyprogesterone acetate use; 46.7% became pregnant. Among those 156 adolescents who discontinued depotmedroxyprogesterone acetate use, 40.0% restarted the method at some later time. Continuation of depot medroxyprogesterone acetate use was more likely if age at first intercourse was younger than 13 years (P = .04). Continuation rates were not related to age, ethnicity, age at menarche, number of sex partners, use of other contraceptives, prior pregnancy, or having a child. CONCLUSIONS: In this study, just less than one third of the adolescents continued depot medroxyprogesterone acetate use for 1 year or longer. This suggests that depot medroxyprogesterone acetate does not function as a long-term method for most inner-city adolescents. The only characteristic that was associated with successful continuation of depot medroxyprogesterone acetate use was young age at first intercourse, implying that experience may be the main determinant of continuation. PMID- 10520616 TI - The spontaneous passage of esophageal coins in children. AB - OBJECTIVES: To determine the likelihood of spontaneous passage of esophageal coins to the stomach in children and to determine the effect of initial coin location on spontaneous passage. DESIGN: Retrospective review of medical records and radiographs. SUBJECTS: Consecutive patients 18 years or younger presenting during a 24-month period (October 1995 to September 1997) whose evaluation revealed an esophageal coin. SETTING: The emergency department of a large, urban academic children's hospital. MAIN OUTCOME MEASURES: Independent measures were time between ingestion and radiographs, initial location of the coin, and categorization of case as "simple" (patients without a history of esophageal disease or surgery, with a single esophageal coin lodged less than 24 hours, and with no respiratory compromise on presentation) or "complex." Dependent measures were spontaneous passage of the coin to the stomach and the time to passage. RESULTS: A total of 116 cases were included in the analysis, of which 84 were simple and 32 complex. Among the 84 simple cases, the coin was initially located in the proximal third of the esophagus in 54 (64%), the middle third in 7 (8%), and the distal third in 22 (26%). For the 32 complex cases, the initial location of the coin was the proximal third of the esophagus in 27 (84%) and the middle third in 5 (16%). Subsequent radiographs were obtained in the emergency department in 58 (69%) of the simple cases. Among these cases, spontaneous passage of the coin to the stomach occurred in 16 (28% [95% confidence interval, 21%-41%]). By initial coin location, spontaneous passage in this group occurred in 22% (7/32) of proximal, 33% (2/6) of middle, and 37% (7/19) of distal esophageal coins (P >.05). Subsequent radiographs were obtained in 14 (44%) of the complex cases; no coin had passed spontaneously to the stomach in these patients (0% [95% confidence interval, 0%-20%]). CONCLUSIONS: Children with a single esophageal coin seen within 24 hours of ingestion, who have no history of esophageal disease and no respiratory compromise on presentation, have a 28% chance of spontaneous passage of the coin to the stomach. Coins in the upper as well as the lower esophagus pass spontaneously. Observing these children for 12 to 24 hours prior to invasive procedures will reduce complications and costs. PMID- 10520617 TI - Etiology of alcohol use among Hispanic adolescents: sex-specific effects of social influences to drink and problem behaviors. AB - BACKGROUND: Hispanic adolescents seem to be at greater risk for alcohol use; a greater understanding of the factors that predict alcohol use among Hispanic youth is needed. Social influences to drink and other problem behaviors often predict adolescent alcohol use. However, most past research has concentrated on samples of predominantly white adolescents residing in suburban areas. OBJECTIVES: To determine which demographic factors, social influences, and problem behaviors are associated with alcohol use among Hispanic adolescents and to eludicate the difference in the origins of alcohol use depending on sex. DESIGN: Cross-sectional study. SETTING: Middle schools in New York City. PARTICIPANTS: This study focuses on 1410 adolescents in grade 7 from inner-city schools who identified themselves as Hispanic at the baseline assessment of an investigation of alcohol and other drug use. MAIN OUTCOME MEASURES: Alcohol initiation, alcohol consumption, and future drinking. RESULTS: The findings showed that social influences to drink and reported problem behaviors were associated with alcohol use across and within sex groups. In particular, friends' drinking was related to alcohol initiation, consumption, and plans to drink in the future across sexes and within both sex groups. Other predictors (mother's drinking, siblings' drinking, ease of obtaining alcohol, deviance, cigarette smoking, and marijuana use) exhibited sex-specific effects. CONCLUSION: These findings lend support to teaching social resistance skills to improve Hispanic adolescents' ability to resist social influences to drink and use other drugs. PMID- 10520618 TI - Conscious sedation with remifentanil and midazolam during brief painful procedures in children. AB - OBJECTIVE: To test the hypothesis that remifentanil, because of its favorable pharmacokinetic properties, would be advantageous to use in combination with midazolam to provide analgesia and sedation during brief painful procedures. DESIGN: Prospective observation and data collection. SETTING: University hospital. PATIENTS: Seventeen children aged 2 to 12 years, who underwent 20 brief, painful procedures. INTERVENTIONS: Administration of intravenous midazolam hydrochloride, 0.05 mg/kg, and remifentanil hydrochloride, 1 microg/kg, followed by a remifentanil infusion at 0.1 microg x kg(-1) x min(-1). The dose was titrated at 5-minute intervals to levels of sedation and analgesia. MAIN OUTCOME MEASURES: Successful remifentanil doses, times to discharge readiness, side effects, complications, and requirement for additional medications. RESULTS: The technique was successful in 17 of 20 procedures. The mean +/- SD successful dose was 0.4 +/- 0.2 microg x kg(-1) x min(-1). Four children developed hypoxemia that abated with mild stimulation; 1 child became unresponsive and required positive pressure ventilation. The mean +/- SD time to reach discharge criteria was 9.5 +/ 4.3 minutes. Hypoxemia was avoided in 10 of 13 patients by continuous stimulation throughout the procedure. CONCLUSION: The use of remifentanil and midazolam during brief, painful procedures results in rapid times to discharge but is complicated by a high incidence of life-threatening respiratory depression at subtherapeutic levels. PMID- 10520619 TI - Are the federal and state governments complying with the Synar Amendment? AB - BACKGROUND: In 1992, Congress enacted the Synar Amendment, requiring states and territories to enact a law prohibiting the sale of tobacco to minors and to enforce that law in a manner that could reasonably be expected to decrease the availability of tobacco to minors. The Department of Health and Human Services (DHHS) was mandated to withhold block grant funding from noncompliant states. OBJECTIVE: To determine if DHHS and applicant states and territories are complying with the Synar Amendment. DATA SOURCES: Fiscal year 1997 substance abuse block grant applications from 59 states and territories. MAIN OUTCOME MEASURES: Whether applicants had enacted a tobacco sales law, conducted enforcement inspections, penalized violators, and conducted a statewide survey, and whether DHHS regulations and actions were consistent with the statutory requirements of the Synar Amendment. RESULTS: Two applicants failed to enact appropriate laws, 15 failed to conduct enforcement inspections, 18 failed to provide a single example of a violator being penalized, and 1 failed to conduct a survey. Nineteen applicants failed to meet the statutory requirements, but none were sanctioned by DHHS as required by the Synar Amendment. The DHHS regulations, as implemented, do not require states to enforce their laws or to achieve illegal tobacco sales rates low enough to reduce the availability of tobacco to minors. CONCLUSIONS: The states and DHHS are violating the statutory requirements of the Synar Amendment, rendering it ineffective. Few states have implemented effective enforcement programs, and national surveys confirm that there has been no measurable reduction in the availability of tobacco to youths. PMID- 10520620 TI - Consequences of incomplete carbohydrate absorption from fruit juice consumption in infants. AB - BACKGROUND: Most infants consume fruit juices by 6 months of age. However, fruit juices containing sorbitol may be associated with carbohydrate malabsorption without clinical symptoms. We hypothesized that increased physical activity and metabolic rate may be associated with carbohydrate malabsorption. METHODS: Physical activity and metabolic rate were determined in 14 healthy infants ([mean +/- SD] age, 5.1 +/- 0.8 months; weight, 7.8 +/- 1.1 kg; length, 67 +/- 4.2 cm; and body fat, 26% +/- 5%) for 3 hours in a respiratory chamber. Seven were fed pear juice, and the other 7 were fed white grape juice (120 mL) after a 2-hour fast. Pear juice contains sorbitol and a high fructose-glucose ratio, whereas white grape juice is sorbitol free and has a low fructose-glucose ratio. Carbohydrate absorption was determined by breath hydrogen gas analysis. The study was double-blinded. RESULTS: When compared with the infants without carbohydrate malabsorption (peak breath hydrogen level < 20 ppm above baseline), 5 of the 7 infants fed pear juice and 2 of the 7 infants fed white grape juice exhibited carbohydrate malabsorption (peak breath hydrogen level > or = 20 ppm above baseline; P < .01). These infants also exhibited both increased physical activity (P < .001) and metabolic rate (P < .05) after juice consumption in comparison with infants with normal carbohydrate absorption. When grouped according to the type of juice consumed, only infants fed pear juice exhibited increases in physical activity (P < .01). CONCLUSIONS: Carbohydrate malabsorption is associated with increased physical activity and metabolic rate in infants. Most of the infants who had carbohydrate malabsorption consumed pear juice. Therefore, fruit juices containing sorbitol and high levels of fructose may not be optimal for young infants. PMID- 10520621 TI - Radiological case of the month. Intrauterine stab wound to the head of a 29-week fetus. PMID- 10520622 TI - Picture of the month. Zellweger (cerebro-hepato-renal) syndrome. PMID- 10520623 TI - Pathologic case of the month. Extraskeletal myxoid chrondosarcoma. PMID- 10520624 TI - Exotic diseases close to home. PMID- 10520625 TI - Nipah-virus encephalitis--investigation of a new infection. PMID- 10520626 TI - Prevention of atherosclerosis in childhood. PMID- 10520627 TI - Should all pregnant women be screened for hypothyroidism? PMID- 10520628 TI - A stimulating new target for cancer immunotherapy. PMID- 10520629 TI - Diagnosing dementia with Lewy bodies. PMID- 10520630 TI - Bed rest: a potentially harmful treatment needing more careful evaluation. AB - BACKGROUND: Bed rest is not only used in the management of patients who are not able to mobilise, but is also prescribed as a treatment for a large number of medical conditions, a procedure that has been challenged. We searched the literature for evidence of benefit or harm of bed rest for any condition. METHODS: We systematically searched MEDLINE and the Cochrane library, and retrieved reports on randomised controlled trials of bed rest versus early mobilisation for any medical condition, including medical procedures. FINDINGS: 39 trials of bed rest for 15 different conditions (total patients 5777) were found. In 24 trials investigating bed rest following a medical procedure, no outcomes improved significantly and eight worsened significantly in some procedures (lumbar puncture, spinal anaesthesia, radiculography, and cardiac catheterisation). In 15 trials investigating bed rest as a primary treatment, no outcomes improved significantly and nine worsened significantly for some conditions (acute low back pain, labour, proteinuric hypertension during pregnancy, myocardial infarction, and acute infectious hepatitis). INTERPRETATION: We should not assume any efficacy for bed rest. Further studies need to be done to establish evidence for the benefit or harm of bed rest as a treatment. PMID- 10520631 TI - Influence of maternal hypercholesterolaemia during pregnancy on progression of early atherosclerotic lesions in childhood: Fate of Early Lesions in Children (FELIC) study. AB - BACKGROUND: Children generally have low cholesterol and no clinical manifestations of atherosclerosis, but fatty-streak formation begins in fetuses and is greatly increased by maternal hypercholesterolaemia during pregnancy. In the FELIC study we assessed the evolution of such lesions during childhood. METHODS: Computer-assisted imaging was used to measure the area of the largest individual lesion and the cumulative lesion area per section in serial cross sections through the entire aortic arch and abdominal aorta of 156 normocholesterolaemic children aged 1-13 years, who died of trauma and other causes. Children were classified by whether their mother had been normocholesterolaemic (n=97) or hypercholesterolaemic (n=59) during pregnancy. Atherosclerosis was correlated with 13 established or potential risk factors. Findings The largest fatty streaks in the aortic arch of children younger than 3 years of hypercholesterolaemic mothers were 64% smaller than those previously found in corresponding fetuses (p<0.0001), which suggests that fetal fatty streaks may regress after birth. In the two groups, lesion size in the aortic arch and abdominal aorta increased linearly with age (r=0.87-0.98). However, lesions progressed strikingly faster in children of hypercholesterolaemic mothers than in those of normocholesterolaemic mothers (p<0.0001). Conventional risk factors for atherosclerosis in children or mothers correlated with lesion size, but did not account for the faster progression of atherogenesis in normocholesterolaemic children of hypercholesterolaemic mothers. INTERPRETATION: Our results suggest that maternal hypercholesterolaemia during pregnancy induces changes in the fetal aorta that determine the long-term susceptibility of children to fatty-streak formation and subsequent atherosclerosis. If so, cholesterol-lowering interventions in hypercholesterolaemic mothers during pregnancy may decrease atherogenesis in children. PMID- 10520632 TI - Current and potential impact of fetal diagnosis on prevalence and spectrum of serious congenital heart disease at term in the UK. British Paediatric Cardiac Association. AB - BACKGROUND: Assessment of the effect of fetal diagnosis on the prevalence of congenital heart disease at term requires national ascertainment because referral patterns are not rigorously structured. METHODS: Between 1993 and 1995, all 17 paediatric cardiac centres in the UK submitted to a database lists of all fetuses diagnosed, and all infants needing surgery or interventional catheterisation or dying in the first year of life because of structural heart disease; details included the postal area of residence. FINDINGS: There were 4799 affected pregnancies, 4165 babies born alive, 1124 fetal diagnoses, and 567 terminations of pregnancy because the fetus had structural heart disease. Thus, a fetal diagnosis was made in 23.4% of affected pregnancies (11.7% of all affected livebirths) with geographical variability in diagnostic rates. INTERPRETATION: Fetal cardiac screening has an effect on the prevalence and types of congenital heart disease seen at term because many affected pregnancies are terminated. If detection rates of affected fetuses rose nationally to those seen in the 15 postal areas where detection rates were significantly higher than the national average in 1993-95, we would expect about 218 fewer affected individuals to be born annually. PMID- 10520633 TI - The epidemiology of chronic pain in the community. AB - BACKGROUND: Chronic pain is recognised as an important problem in the community but our understanding of the epidemiology of chronic pain remains limited. We undertook a study designed to quantify and describe the prevalence and distribution of chronic pain in the community. METHODS: A random sample of 5036 patients, aged 25 and over, was drawn from 29 general practices in the Grampian region of the UK and surveyed by a postal self-completion questionnaire. The questionnaire included case-screening questions, a question on the cause of the pain, the chronic pain grade questionnaire, the level of expressed needs questionnaire, and sociodemographic questions. FINDINGS: 3605 questionnaires were returned completed. 1817 (50.4%) of patients self reported chronic pain, equivalent to 46.5% of the general population. 576 reported back pain and 570 reported arthritis; these were the most common complaints and accounted for a third of all complaints. Backward stepwise logistic-regression modelling identified age, sex, housing tenure, and employment status as significant predictors of the presence of chronic pain in the community. 703 (48.7%) individuals with chronic pain had the least severe grade of pain, and 228 (15.8%) the most severe grade. Of those who reported chronic pain, 312 (17.2%) reported no expressed need, and 509 (28.0%) reported the highest expressed need. INTERPRETATION: Chronic pain is a major problem in the community and certain groups within the population are more likely to have chronic pain. A detailed understanding of the epidemiology of chronic pain is essential for efficient management of chronic pain in primary care. PMID- 10520634 TI - Outbreak of Nipah-virus infection among abattoir workers in Singapore. AB - BACKGROUND: In March 1999, an outbreak of encephalitis and pneumonia occurred in workers at an abattoir in Singapore. We describe the clinical presentation and the results of investigations in these patients. METHODS: Clinical and laboratory data were collected by systemic review of the case records. Serum and cerebrospinal fluid (CSF) samples were tested for IgM antibodies to Nipah virus with an IgM capture ELISA. Reverse-transcriptase PCR was done on the CSF and tissue samples from one patient who died. FINDINGS: Eleven patients were confirmed to have acute Nipah-virus infection based on raised IgM in serum. Nipah virus was identified by reverse transcriptase PCR in the CSF and tissue of the patient who died. The patients were all men, with a median age of 44 years. The commonest presenting symptoms were fever, headache, and drowsiness. Eight patients presented with signs of encephalitis (decreased level of consciousness or focal neurological signs). Three patients presented with atypical pneumonia, but one later developed hallucinations and had evidence of encephalitis on CSF examination. Abnormal laboratory findings included a low lymphocyte count (nine patients), low platelet count, low serum sodium, and high aspartate aminostransferase concentration (each observed in five patients). The CSF protein was high in eight patients and white-blood-cell count was high in seven. Chest radiography showed mild interstitial shadowing in eight patients. Magnetic resonance imaging (MRI) showed focal areas of increased signal intensity in the cortical white marker in all eight patients who were scanned. The nine patients with encephalitis received empirical treatment with intravenous aciclovir and eight survived. INTERPRETATION: Infection with Nipah virus caused an encephalitis illness with characteristic focal areas of increased intensity seen on MRI. Lung involvement was also common, and the disease may present as an atypical pneumonia. PMID- 10520635 TI - Fatal encephalitis due to Nipah virus among pig-farmers in Malaysia. AB - BACKGROUND: Between February and April, 1999, an outbreak of viral encephalitis occurred among pig-farmers in Malaysia. We report findings for the first three patients who died. METHODS: Samples of tissue were taken at necropsy. Blood and cerebrospinal-fluid (CSF) samples taken before death were cultured for viruses, and tested for antibodies to viruses. FINDINGS: The three pig-farmers presented with fever, headache, and altered level of consciousness. Myoclonus was present in two patients. There were signs of brainstem dysfunction with hypertension and tachycardia. Rapid deterioration led to irreversible hypotension and death. A virus causing syncytial formation of vero cells was cultured from the CSF of two patients after 5 days; the virus stained positively with antibodies against Hendra virus by indirect immunofluorescence. IgM capture ELISA showed that all three patients had IgM antibodies in CSF against Hendra viral antigens. Necropsy showed widespread microinfarction in the central nervous system and other organs resulting from vasculitis-induced thrombosis. There was no clinical evidence of pulmonary involvement. Inclusion bodies likely to be of viral origin were noted in neurons near vasculitic blood vessels. INTERPRETATION: The causative agent was a previously undescribed paramyxovirus related to the Hendra virus. Close contact with infected pigs may be the source of the viral transmission. Clinically and epidemiologically the infection is distinct from infection by the Hendra virus. We propose that this Hendra-like virus was the cause of the outbreak of encephalitis in Malaysia. PMID- 10520636 TI - "Something moving in my head". PMID- 10520637 TI - Identification of a Kunjin/West Nile-like flavivirus in brains of patients with New York encephalitis. AB - Molecular analysis of brains from patients of the recent New York City encephalitis outbreak reveals the presence of a flavivirus not previously described in the Americas. PMID- 10520638 TI - Retinoic acid for treatment of multicentric Castleman's disease. AB - Multicentric Castleman's syndrome has an aggressive course with poor prognosis, and its treatment remains uncertain. We report a woman with multicentric Castleman's disease that was successfully treated with prednisone and retinoic acid. PMID- 10520639 TI - Truth telling in the case of a pessimistic diagnosis in Japan. AB - We investigated how physicians in Japan convey a poor prognosis of advanced cancer. Physicians tended to give patients optimistic accounts of their prognosis, while they were inclined to give the families pessimistic accounts. PMID- 10520640 TI - Down's syndrome screening in the UK in 1998. AB - The extent of antenatal screening for Down's syndrome with serum or ultrasound markers has increased over the past decade. We here present a survey of screening in the UK in 1998 and compare the results with similar surveys from 1991 and 1994. PMID- 10520641 TI - Association of CCR5 delta32 with reduced risk of asthma. AB - We report that individuals carrying the CCR5 delta32 mutation, a naturally occurring variant of the C-C chemokine receptor 5 (CCR5), are at reduced risk of developing asthma. These data suggest a possible explanation for the high prevalence of this mutation in the general population. PMID- 10520642 TI - Fatal uncertainty: death-rate from use of ecstasy or heroin. AB - We provide a 25-fold range for the ecstacy-related death rate per 10,000 15-24 year-old users in the UK: from 0.2 to 5.3, compared with the death rate of 1.0 from road traffic accidents in the same age-group. The heroin-related death rate in 15-24-year-old heroin users was much higher, but also imprecisely estimated: from 9.1 to 81.5 deaths per 10,000 15-24-year-old users. Data deficiencies which inhibit the calculation of drug-specific rates in this population should be remedied. PMID- 10520643 TI - Olestra increases faecal excretion of 2,3,7,8-tetrachlorodibenzo-p-dioxin. AB - Two patients with chloracne had concentrations of 2,3,7,8-tetrachlorodibenzo-p dioxin (TCDD) of 144,000 and 26,000 pg/g blood lipids. Olestra, a non-digestible, lipophilic dietary fat substitute accelerated the patients' intestinal excretion of TCDD by eight to ten fold. This is sufficient to reduce the normally observed elimination half life of TCDD from about 7 years to 1-2 years. PMID- 10520644 TI - Beware of dogs licking ears. AB - A patient with right-sided chronic purulent otorrhoea developed meningitis due to Pasteurella multocida transmitted by a dog that frequently licked his ear. We suggest that patients with a perforated tympanic membrane should avoid being licked on their ears by animals. PMID- 10520645 TI - Inroads made into transplantation problems. PMID- 10520646 TI - Japanese government offers guidelines for stressed workers. PMID- 10520647 TI - New paradigms, amid scepticism colour evidence-based medicine meeting. PMID- 10520648 TI - Influenza. AB - Influenza is the most frequent cause of acute respiratory illness requiring medical intervention because it affects all age groups and because it can recur in any individual. During the past three decades, efforts to prevent and control influenza have focused primarily on the use of inactivated influenza vaccines in elderly people and in individuals with chronic medical conditions that put them at risk for complications. However, the continuing impact of influenza in these and other population groups has motivated the development of novel approaches for prevention and control of influenza. Several important advances in the field of influenza have occurred in the last few years. An experimental live, attenuated, intranasally administered trivalent influenza vaccine was shown to be highly effective in protecting young children against influenza A H3N2 and influenza B. New antiviral drugs based on the structure of the neuraminidase molecule were assessed in clinical trials and found to be effective against influenza A and B viruses. The expected use of these new antiviral agents has accelerated the development of rapid point-of-care diagnostic tests. The availability of new diagnostic tests, new antiviral drugs, and new vaccines will undoubtedly alter our approaches to influenza control and have an impact on clinical practice. PMID- 10520649 TI - Carbon dioxide and the critically ill--too little of a good thing? AB - Permissive hypercapnia (acceptance of raised concentrations of carbon dioxide in mechanically ventilated patients) may be associated with increased survival as a result of less ventilator-associated lung injury. Conversely, hypocapnia is associated with many acute illnesses (eg, asthma, systemic inflammatory response syndrome, pulmonary oedema), and is thought to reflect underlying hyperventilation. Accumulating clinical and basic scientific evidence points to an active role for carbon dioxide in organ injury, in which raised concentrations of carbon dioxide are protective, and low concentrations are injurious. We hypothesise that therapeutic hypercapnia might be tested in severely ill patients to see whether supplemental carbon dioxide could reduce the adverse effects of hypocapnia and promote the beneficial effects of hypercapnia. Such an approach could also expand our understanding of the pathogenesis of disorders in which hypocapnia is a constitutive element. PMID- 10520650 TI - Continuum of palliative care: lessons from caring for children infected with HIV 1. PMID- 10520651 TI - Decline in child health in rural Papua New Guinea. PMID- 10520652 TI - Laparoscopic hernia surgery. PMID- 10520653 TI - Laparoscopic hernia surgery. PMID- 10520654 TI - Laparoscopic hernia surgery. PMID- 10520655 TI - Laparoscopic hernia surgery. PMID- 10520656 TI - Laparoscopic hernia surgery. PMID- 10520657 TI - Non-alcoholic steatohepatitis. PMID- 10520658 TI - Non-alcoholic steatohepatitis. PMID- 10520659 TI - Non-alcoholic steatohepatitis. PMID- 10520660 TI - Non-alcoholic steatohepatitis. PMID- 10520661 TI - Antioxidants and Friedreich's ataxia. PMID- 10520662 TI - Survival benefit of trandolapril. PMID- 10520663 TI - Mutations of cationic trypsinogen in chronic pancreatitis. PMID- 10520665 TI - Heart rate variability and hormone replacement therapy. PMID- 10520664 TI - Risks of hormone replacement therapy. PMID- 10520666 TI - Lonomia obliqua and haemorrhagic syndrome. PMID- 10520667 TI - Risk of transmission of BSE via drugs of bovine origin. PMID- 10520668 TI - Are European-specific guidelines needed for prevention of opportunistic infections in HIV-1? PMID- 10520669 TI - Are disability weights universal? WHO/NIH Joint Project CAR Study Group. PMID- 10520670 TI - Hypoglycaemia and driving. PMID- 10520671 TI - The Nobel chronicles. 1975: Renato Dulbecco (b 1914), David Baltimore (b 1938), and Howard Martin Temin (1934-94). PMID- 10520672 TI - Inconsistencies in the "societal perspective" on costs of the Panel on Cost Effectiveness in Health and Medicine. AB - A key recommendation of the recent Panel on Cost-Effectiveness in Health and Medicine was that cost-effectiveness analyses be carried out from a societal perspective. The authors show that two of the Panel's recommendations concerning costs are not consistent with a societal perspective, and how to correct those inconsistencies. In its recommendations concerning costs resulting from morbidity, the Panel advises excluding lost income from costs in the belief that individuals take income changes into account when they respond to the quality-of life questions that are used to calculate quality-adjusted life years (QALYs). It is shown that even if individuals do consider income changes in responding to these quality-of-life questions, this recommendation would seriously underestimate production losses due to morbidity, since individuals do not bear a major part of lost production. In its recommendations concerning costs resulting from mortality, the Panel does not require that health care costs for "unrelated" illness and non-health care consumption and production during added life years be included in the Reference Case. It is shown that omitting these costs will seriously distort comparisons of programs at different ages and favor programs that extend life over those that improve quality of life. This can be corrected by including total consumption minus production in added life-years among costs. PMID- 10520673 TI - Realistic rigor in cost-effectiveness methods. PMID- 10520674 TI - Improving the Panel's recommendations. PMID- 10520675 TI - Theoretically correct cost-effectiveness analysis. PMID- 10520676 TI - Life expectancy estimation with breast cancer: bias of the declining exponential function and an alternative to its use. AB - BACKGROUND: Life expectancy gain (LEG) is an outcome measure commonly estimated with a declining exponential function in a Markov model. The accuracy of such estimates has not been objectively evaluated. PURPOSE: To compare LEGs from declining exponential function estimates with those calculated from population data, using published screening mammography studies as examples. METHOD: SEER based population data are used to compare LEG calculation with declining exponential function estimation and empiric population data in a new model, the "nested" Markov. RESULTS: Analyses of the LEG of mammographic screening based on the declining exponential function significantly overestimate LEGs for younger women and underestimate them for older women. Because of offsetting errors, all age analyses paradoxically appear accurate. CONCLUSION: Declining exponential function estimates of LEGs for chronic diseases with low mortality rates and long time horizons are liable to significant bias, especially with limited age cohorts. PMID- 10520677 TI - How patients' preferences for risk information influence treatment choice in a case of high risk and high therapeutic uncertainty: asymptomatic localized prostate cancer. AB - To assess how patients' preferences for non-numerical risk information are related to their tendency to choose early surgical treatment for asymptomatic gland-confined prostate cancer (a choice with high risk and high therapeutic uncertainty), the authors conducted a cross-sectional study of 228 patients receiving continuing care in a general medicine clinic. After being provided with three data disclosures related to the treatment decision, subjects were given a choice between surgery-now and watchful waiting. Data about surgical complications were presented in numerical format. The subjects were also asked whether they preferred communication with their physician about the chance (probability) of adverse outcomes-related to management strategies-in terms of words (such as possible or probable) or numbers (such as percentages). Of the 226 patients who chose either surgery-now or watchful waiting, 71.2% preferred risk information in terms of words only or numbers only: 44% words only, and 56% numbers only. Younger patients (OR = 1.06 per year; CI = 1.02-1.10, p = 0.0008) and those wanting risk communication in terms of words only (OR = 2.41; CI = 1.24 4.70, p = 0.01) tended to prefer surgery-now over watchful waiting as the management strategy for asymptomatic gland-confined prostate carcinoma. The authors conclude that there is a significant association between patients' preferences for risk communication with their physicians in terms of words only and a tendency to prefer early surgical intervention for prostate cancer when surgical risk data are provided numerically. PMID- 10520678 TI - Development of a pediatric multiple organ dysfunction score: use of two strategies. AB - BACKGROUND: An organ dysfunction (OD) scoring system for critically ill children is not yet available, and the method for developing such a system is not well defined. The aim of this study was to compare two developmental methods for assessing OD in critically ill children. METHODS: Consecutive admissions between January and May 1997 in three French and Canadian pediatric intensive care units (PICUs) were studied prospectively. Physiologic data were selected using a Delphi method; the most abnormal values during PICU stay were recorded. The outcome measure was the vital status at PICU discharge. Six organ systems were studied: hepatic, cardiovascular, renal, hematologic, respiratory, and neurologic. For each of the six organ systems, the PEdiatric Multiple OD (PEMOD) system included one variable and the PEdiatric Logistic OD (PELOD) system included several variables. Severity levels and relative weights of ODs were determined according to the mortality rate (PEMOD) or by logistic regression (PELOD). RESULTS: There were 594 admissions, including 51 deaths (9%). Severity levels and relative weights of ODs were: four levels graded from 1 to 4 for the PEMOD system and three levels with scores of 1, 10, and 20 for PELOD system. For both systems, calibrations were good (p = 0.23 and p = 0.44 respectively). The PELOD system was more discriminant than the PEMOD system (areas under the ROC curves 0.98 and 0.92, respectively, p < 10(-5)). Moreover, with the PEMOD system, four ODs did not contribute significantly to the prediction of PICU outcome. CONCLUSIONS: The PELOD system was more discriminant and had the advantage of taking into account both the relative severities among ODs and the degree of severity of each OD. PMID- 10520679 TI - Medical decision making in the choice of a thrombolytic agent for acute myocardial infarction. Quebec Acute Coronary Care Working Group. AB - Little is known about how physicians make decisions when the evidence is incomplete or controversial. While thrombolysis improves survival following acute myocardial infarction (AMI), conflicting evidence exists as to any specific agent's superiority, particularly if cost-effectiveness is considered. Using a Bayesian hierarchical model, the authors examined the patient, physician, and hospital characteristics that are related to the decision-making process concerning the choice of thrombolytic agent in a prospective registry of 1,165 AMI patients receiving thrombolysis. Tissue plasminogen activator (t-PA) was administered to 432 patients (31.8%) and streptokinase (SK) to the remainder. The presence of an anterior infarction, a previous myocardial infarction, low blood pressure, a cardiologist decision maker, younger age, and receiving treatment within six hours after the start of symptoms were independent predictors of receiving t-PA. The levels of importance that physicians accorded to these patient characteristics differed according to their practicing institutions. Generally, they followed evidence-based medicine and reasonably targeted high risk patients to receive the more expensive t-PA. However, they also preferentially treated younger patients, where only a small absolute advantage appears to exist. PMID- 10520680 TI - Evaluation of a decision-support system for preoperative staging of prostate cancer. AB - The usefulness and effectiveness of a decision-support system for preoperative staging of prostate cancers (PCES) were evaluated. The study population consisted of 43 consecutive patients with the preoperative diagnosis of prostate cancer who underwent surgical operation. Results obtained using the PCES were compared with staging by four urology attending physicians and five urology residents. The effect of PCES consultation on the physicians' staging of prostate cancer was also evaluated. To confirm the usefulness of the clinical findings of prostate specific antigen, prostate-specific antigen density, prostate volume, and abnormal Gleason score in the PCES, their receiver operating characteristic (ROC) curves for diagnosis of advanced prostate cancer were plotted. The values of the areas under the curves were 0.772, 0.800, 0.531, and 0.752. The stage of prostate cancer was correctly determined by the PCES for 38 of the 43 patients, yielding 88.4% preoperative diagnostic accuracy. The PCES was significantly more accurate than two of the attending physicians and all residents. PCES consultation improved the residents' staging accuracy to approximately that of the attending physicians. The effect of PCES consultation on the residents' staging was significantly (p < 0.001) greater than the effect on the physicians' staging. The PCES may be useful in the preoperative staging of prostate cancers, especially during residency. The system's accuracy in determining the stage of advanced prostate cancer may make it possible to avoid unneccesary surgical operations. PMID- 10520682 TI - The discrepancy between risky and riskless utilities: a matter of framing? AB - Utilities differ according to whether they are derived from risky (gamble) and riskless (visual analog scale, time-tradeoff) assessment methods. The discrepancies are usually explained by assuming that the utilities elicited by risky methods incorporate attitudes towards risk, whereas riskless utilities do not. In (cumulative) prospect theory, risk attitude is conceived as consisting of two components: a decision-weight function (attentiveness to changes in, or sensitivity towards, chance) and a utility function (sensitivity towards outcomes). The authors' data suggest that a framing effect is a hitherto unrecognized and important factor in causing discrepancies between risky and riskless utilities. They collected risky evaluations with the gamble method, and riskless evaluations with difference measurement. Risky utilities were derived using expected-utility theory and prospect theory. With the latter approach, sensitivity towards outcomes and sensitivity towards chance are modeled separately. When the hypothesis that risky utilities from prospect theory coincide with riskless utilities was tested, it was rejected (n = 8, F(1,7) = 132, p = 0.000), suggesting that a correction for sensitivity towards chance is not sufficient to resolve the difference between risky and riskless utilities. Next, it was assumed that different gain/loss frames are induced by risky and riskless elicitation methods. Indeed, identical utility functions were obtained when the gain/loss frames were made identical across methods (n = 7), suggesting that framing was operative. The results suggest that risky and riskless utilities are identical after corrections for sensitivity towards chance and framing. PMID- 10520681 TI - How should effectiveness of risk communication to aid patients' decisions be judged? A review of the literature. AB - Risk-communication interventions are associated with benefits at both the individual and the public health level. However, the types of outcomes used to assess the effectiveness of risk-communication interventions vary greatly. This makes synthesis of the research in systematic review difficult, and limits both the implementation of advances in clinical practice and further research. This article reviews the outcomes used in risk-communication publications, particularly those addressing individual decisions about treatment. From the traditional cognitive and behavioral research outcomes of patient knowledge, risk perception, and compliance, the emphasis has shifted towards more affective outcomes, including satisfaction, assessment of the information provided and the decision-making process, and certainty about whether the best option has been chosen. These affective outcomes may be more specific and sensitive measures for risk-communication research. Further development and validation of measurement scales to address these issues is needed. PMID- 10520683 TI - Motivating mammography adherence in elderly Latinas: a test of three mathematical models of decision making. AB - Many elderly Latinas do not have mammography every one to two years as recommended by cancer organizations. To elucidate the causal factors underlying this behavior, 52 Latinas, aged 65 and over, were asked to judge the likelihood of having yearly mammography in 79 different scenarios constructed from factor levels of cost, perceived risk, and the source of a recommendation (none, a recognized cancer organization, a doctor), assuming a convenient mammography facility. A configural-weight-averaging model, with different parameter values for the 30 adherers (women who reported having had mammography at least twice in the preceding four years) and the 22 non-adherers, gave a good fit to the data and did well in predicting reported mammography adherence (r = 0.85). According to this model, offering free mammography would not induce non-adherers to adhere; they would require a recommendation, and value a doctor's as highly as that of a recognized cancer organization, but reported never having received one from either source. All 52 women reported never receiving risk information from any source. These results have direct educational and dissemination implications for cancer organizations. PMID- 10520684 TI - Do people value their own future health differently from others' future health? AB - The purpose of this study was to investigate whether time preferences for own health are the same as time preferences for others' health. A random sample of the general public was sent a postal questionnaire containing six choices between ill health in the near future and ill health in the further future. They were asked to indicate the maximum duration of more distant ill health they would be willing to accept in return for a specified delay in the onset of the period of ill health. For half of the sample the questions were set in the context of their own health and for the other half in terms of others' health. The median implied discount rates were not statistically different, 0.061 for own health and 0.062 for others' health. A multilevel analysis of the determinants of these implied discount rates provided additional evidence of the similarity of time preferences for own health and others' health. PMID- 10520685 TI - Effects on preferences of violations of procedural invariance. AB - BACKGROUND: In studies of health preferences, utilities for hypothetical health states cannot always be successfully measured. One marker for unsuccessful measurement is violation of "procedural invariance": when the ranking of two health states varies across assessment procedures. Using preference values based on unsuccessful measurement may result in misinterpretation of patients' attitudes about health. OBJECTIVE: The authors sought to determine whether people who violated procedural invariance had different preferences than people who satisfied it. METHODS: They performed secondary analyses of three completed studies that used the same two assessment procedures, identifying participants who violated procedural invariance and comparing the mean standard gamble (SG) and visual analog scale (VAS) scores of violators and satisfiers. PARTICIPANTS: Experiment 1, 30 healthy volunteers and 30 patients with cardiac arrhythmias; experiment 2, 139 patients with depressive illness; experiment 3, 98 family members of patients with schizophrenia. RESULTS: Rates of violation of procedural invariance ranged from 16% to 32%. Violation of procedural invariance was not associated with age, education level, race, or gender. Subjects with violations of procedural invariance had, in general, less ability to discriminate among states and less reliable VAS and SG measurements, and sometimes had different mean SG and VAS values. CONCLUSIONS: Violation of procedural invariance of preferences across scaling methods may be a signal for failure of the measurement process. Researchers should test for procedural invariance and consider reporting data separately for satisfiers and violators. PMID- 10520686 TI - Whose preferences count? AB - An important consideration when choosing how to allocate health care resources is the improvements in patients' health-related quality of life (HRQoL) that alternative allocations generate. There is considerable debate about whose preferences should be used when measuring and valuing HRQoL. This debate has usually been in terms of whether the values of patients or the general public are the most appropriate. It is argued in this paper that this is a false dichotomy that does not facilitate understanding of empirical evidence. Nor, more importantly, does it address one of the most important issues in the debate about whose preferences count, that is, whether the fact that many people adapt to poor health states should be taken into account when ascribing values to those states. A conceptual framework is developed to facilitate a more fruitful discussion of the issues relating to the question of whose preferences should count. PMID- 10520687 TI - Assessing convergent validity of health-state utilities obtained using different scaling methods. AB - A number of empirical studies have attempted to assess the convergent validity of health-state utilities obtained using two or more scaling methods (standard gamble, time tradeoff, rating scale, magnitude estimation, equivalence technique, and willingness-to-pay). The data from these studies can be mapped onto an N x K matrix, where N and K are the numbers of respondents and health states, respectively, and each matrix cell consists of a pair of health-state utilities, one obtained using scaling method X and the other obtained using scaling method Y. The Pearson's rassessing convergent validity can then be computed as 1) the unraveled correlation over all N x K data pairs, 2) the mean within-respondent correlation, 3) the mean within-health-state correlation, or 4) the correlation of the across-respondents means. These four different ways of computing the correlation do not necessarily yield the same results. The appropriateness of each method of computing the correlation is considered. PMID- 10520688 TI - Sensitivity analysis on a chance node with more than two branches. AB - Sensitivity analysis is an essential part of decision analysis. The literature on medical decision analysis suggests the use of two-branch chance nodes in decision trees to avoid logical inconsistencies during sensitivity analysis. The authors show that the two-branch decomposition is not appropriate for sensitivity analysis when multiple outcomes from a single state cannot be disentangled into a sensible sequence of events. They recommend retaining the natural structure of the tree and propose two sensitivity-analysis methods for use on chance nodes with three or more branches. PMID- 10520689 TI - Does DEALE-ing stack the deck? PMID- 10520690 TI - A call to standardize measures for judging the efficacy of interventions to aid patients' decision making. PMID- 10520691 TI - On the role of framing effects in assessment of health-related utilities. PMID- 10520692 TI - Nuclear medicine pioneers: Henry N. Wagner, Jr., MD. PMID- 10520693 TI - Public affairs update. PMID- 10520694 TI - Fused image tomography: where do we go from here? PMID- 10520695 TI - SPECT image analysis using statistical parametric mapping in patients with Parkinson's disease. AB - This study investigated alterations in regional cerebral blood flow (rCBF) in patients with Parkinson's disease using statistical parametric mapping (SPM). METHODS: Noninvasive rCBF measurements using 99mTc-ethyl cysteinate dimer (ECD) SPECT were performed on 28 patients with Parkinson's disease and 48 age-matched healthy volunteers. The Parkinson's disease patients were divided into two groups, 16 patients with Hoehn and Yahr stage I or II and 12 patients with Hoehn and Yahr stage III or IV. We used the raw data (absolute rCBF parametric maps) and the adjusted rCBF images in relative flow distribution (normalization of global CBF for each subject to 50 mL/100 g/min with proportional scaling) to compare these groups with SPM. RESULTS: In patients with stage I or II Parkinson's disease, we found a diffuse decrease in absolute rCBF in the whole brain with sparing of the central gray matter, hippocampus and right lower temporal lobe compared with healthy volunteers. Adjusted rCBF increased in both putamina and the right hippocampus. In patients with stage III or IV disease, rCBF decreased throughout the whole brain. Adjusted rCBF increased bilaterally in the putamina, globi pallidi, hippocampi and cerebellar hemispheres (dentate nuclei) and in the left ventrolateral thalamus, right insula and right inferior temporal gyrus. CONCLUSION: SPM analysis showed that significant rCBF changes in Parkinson's disease accompanied disease progression and related to disease pathophysiology in the functional architecture of thalamocortex-basal ganglia circuits and related systems. PMID- 10520696 TI - Evaluation of phosphoinositide turnover on ischemic human brain with 1-[1-11C] butyryl-2-palmitoyl-rac-glycerol using PET. AB - It is important to evaluate cerebral function from neural signal transduction in ischemic brain in judging morbid state and prognosis. We synthesized 1-[1-(11)C] butyryl-2-palmitoyl-rac-glycerol (DAG) for the purpose of imaging the second messenger on PET and applied it to clinical cases of cerebral infarction. METHODS: Five patients, who had ischemic stroke, were examined with PET. [15O] CO2 and [15O]-O2 inhalation methods were applied to cerebral blood flow (CBF), oxygen extraction fraction (OEF) and cerebral metabolic rate of oxygen (CMRO2). For the measurement of phosphoinositide turnover after intravenous injection of DAG, dynamic PET data were collected continuously for 46 min. Arterial blood samples were taken to evaluate changes in the serum concentration of DAG. To quantify the metabolic activity of inositol phospholipid, the incorporation constant k*(DAG) was calculated on the basis of the kinetics of DAG. RESULTS: The plasma concentration of DAG increased rapidly and peaked 30 s after injection of DAG solution. In the normal cortex, DAG concentration increased gradually and reached a plateau between 15 and 20 min after injection. In the ischemic core (infarction), DAG concentration increased slowly, and its peak concentration was lower than that in normal tissue. In comparison with blood flow and metabolic parameters, k*(DAG) showed the best correlation with CMRO2, suggesting a reflection of neuronal activity. Locally, CBF and CMRO2 gradually decreased from the normal area toward the ischemic center (infarction), whereas k*(DAG) and OEF significantly decreased only in the ischemic center. CONCLUSION: The k*(DAG) of ischemic brain, including that caused by infarction, significantly correlated with CMRO2, suggesting that metabolic activity of inositol phospholipid reflects neural viability. Maintained metabolic activity of inositol phospholipid in the region around the ischemic core indicated preservation of the signal transduction system through the metabotropic receptor. PMID- 10520697 TI - Differential diagnosis of thymic tumors using a combination of 11C-methionine PET and FDG PET. AB - We assessed the usefulness of PET studies in making a differential diagnosis of thymic tumors by using 11C-methionine (MET) and 18F-fluorodeoxyglucose (FDG). METHODS: We examined 31 patients with thymic tumors, including 14 patients with thymic cancer, 9 with invasive thymoma, 5 with noninvasive thymoma and 3 with thymic cysts. The histological diagnosis was confirmed by either surgery or biopsy. MET PET and FDG PET were performed in 28 and 29 patients, respectively. Both the MET and FDG uptakes were evaluated by the standardized uptake value (SUV). RESULTS: MET uptake was not substantially different among thymic cancer (4.8 +/- 1.4), invasive thymoma (4.3 +/-1.1) and noninvasive thymoma (4.5 +/- 1.2), bat MET uptake in thymic cysts (0.9 +/- 0.1) was lower than that in the other three tumors (P < 0.01). The FDG uptake in thymic cancer (7.2 +/- 2.9) was higher than that in invasive thymoma (3.8 -/+ 1.3), noninvasive thymoma (3.0 +/- 1.0) and thymic cysts (0.9) (P < 0.01). MET uptake in thymic tumors correlated with the FDG uptake (r = 0.65), whereas MET uptake in thymic cancer was lower than FDG uptake (FDG/MET ratio = 1.52 +/- 0.52) but was higher than FDG uptake in both invasive and noninvasive thymoma (FDG/ MET ratio = 0.86 +/- 0.33). To differentiate thymic cancer from thymoma, a receiver operating characteristic (ROC) analysis was performed. The area under the curve of FDG PET was 0.90, whereas the FDG/MET ratio was 0.87. CONCLUSION: The MET PET, FDG PET and the FDG/MET ratios were unable to differentiate benign thymic tumors from malignant ones, although FDG PET was considered to be useful in the differential diagnosis between thymic cancer and thymoma. Although the difference in the uptake ratio between FDG and MET suggests a different origin of the tumors, the FDG/MET ratio is not considered to be useful as a complementary method for the differential diagnosis of thymic tumors. PMID- 10520698 TI - Detection of bone metastases in patients with endocrine gastroenteropancreatic tumors: bone scintigraphy compared with somatostatin receptor scintigraphy. AB - Scintigraphy with somatostatin analogs is a sensitive method for the staging and therapeutic management of patients with endocrine gastroenteropancreatic (GEP) tumors. The aim of this study was to compare prospectively somatostatin receptor scintigraphy (SRS) using 111n-pentetreotide with bone scintigraphy using 99mTc hydroxymethylene diphosphonate for the detection of bone metastases. METHODS: One hundred-forty-five patients with proven endocrine GEP tumors were investigated. Patients were classified according to the presence of bone metastases as indicated by CT, MRI or histologic data. Group I included 19 patients with confirmed bone metastases, and group II included 126 patients without bone metastases. RESULTS: In group I, SRS was positive in all 19 patients with bone metastases, and bone scintigraphy was positive in 17 patients. Bone metastases were found to occur predominantly in patients with liver metastases. In group 11, 5 patients had recent bone surgery for fracture or arthritis. SRS showed bone uptake in 4 of these patients, and bone scanning showed abnormal uptake in 5. In 7 of the remaining 121 group II patients, SRS was negative and bone scanning showed abnormal bone uptake suggesting bone metastases. The detection of bone metastases was of major prognostic value, because 42% of group 1 patients died during a 2-y follow-up. CONCLUSION: In patients with GEP tumors, the accuracy of SRS appears to be similar to that of bone scintigraphy for the detection of bone metastases. PMID- 10520699 TI - Prediction of hematologic toxicity after radioimmunotherapy with (131)I-labeled anticarcinoembryonic antigen monoclonal antibodies. AB - This study was undertaken to determine the factors affecting myelotoxicity after radioimmunotherapy (RAIT) with 131I-labeled anticarcinoembryonic antigen (anti CEA) monoclonal antibodies (MAbs). METHODS: Ninety-nine patients who received 131I-labeled MN-14 or NP-4 anti-CEA MAbs for the treatment of CEA-producing cancers were assessed for platelet and white blood cell (WBC) toxicity based on the common Radiation Therapy Oncology Group (RTOG) criteria. Univariate and multivariate regression analyses were used to identify the statistically significant factors affecting toxicity among the following variables: red marrow dose, baseline platelet and WBC counts, bone or marrow (or both) metastases, prior chemo- or radiotherapy, timing of prior chemo- or radiotherapy in relation to RAIT, type and number of prior chemotherapeutic regimens, age, sex, antibody form and cancer type. RESULTS: Red marrow dose, baseline platelet or WBC counts and multiple bone or marrow (or both) metastases were the only significant factors affecting hematologic toxicity according to both univariate and multivariate analyses, whereas chemotherapy, 3-6 mo before RAIT, was significant according to multivariate analysis. In this retrospective study, the multivariate regression equations using these four variables provided an exact fit for postRAIT platelet toxicity grade (PltGr) and WBC toxicity grade (WBCGr) in 40% and 46%, respectively, of the 99 patients included in the analysis. Moreover, severe (grade 3 or 4) PltGr and WBCGr could be classified accurately in all cases, whereas nonsevere (grade 0, 1, or 2) PltGr and WBCGr could be classified accurately in all but 6 of 13 cases of grade 2 toxicity, in which a severe toxicity grade was estimated using the regression equations. CONCLUSION: Red marrow dose, baseline blood counts, multiple bone or marrow (or both) metastases and recent chemotherapy are the most important factors related to hematologic toxicity after RAIT. This study provides a simple model for predicting myelotoxicity with reasonable accuracy in most patients. In addition, the identification of bone or marrow (or both) metastases and recent chemotherapy as significant factors for myelotoxicity may be important in the future design of clinical trials. PMID- 10520700 TI - Comparison of dual-head coincidence PET versus ring PET in tumor patients. AB - This study compared the multiring detector (Ring-PET) and the dual-head coincidence imaging system (DH-PET) for staging/ restaging neoplastic patients before or after surgery or radiochemotherapy. METHODS: Seventy patients with suspected tumor recurrence or metastatic dissemination received an intravenous dose of 18F-fluorodeoxyglucose (FDG) under overnight fasting and were studied in sequence with a dedicated positron emission tomograph with Ring-PET and a DH-PET. Ring-PET studies were performed 45-75 min postinjection and were followed by a DH PET scan approximately 3 h postinjection. Number and location of the hypermetabolic lesions detected on DH-PET and Ring-PET reconstructed images were blindly assessed by three independent observers. RESULTS: DH-PET identified all 14 head lesions detected by Ring-PET, 53 of 63 thoracic lesions and 36 of 45 abdominal lesions. Of the 19 lesions not identified by DH-PET, 6 were smaller than 10 mm, 8 were between 10 and 15 mm and 1 was 18 mm; dimensions of 4 bone lesions were not available. A concordant restaging, based on location and number of lesions detected, was found in all 14 patients with head tumors, in 28 of 30 patients with thoracic tumors and in 24 of 26 patients with abdominal tumors. CONCLUSION: We found a good agreement between Ring-PET and DH-PET assessment of oncologic patients in detecting hypermetabolic lesions > or = 10-15 mm. PMID- 10520701 TI - Sensitivity in detecting osseous lesions depends on anatomic localization: planar bone scintigraphy versus 18F PET. AB - Radionuclide bone scanning (RNB) is considered to be the most practical screening technique for assessing the entire skeleton for skeletal metastases. However, RNB has been shown to be of lower sensitivity than MRI and CT in detecting osteolytic metastases. A prospective study was designed to evaluate the accuracy of planar RNB versus tomographic bone imaging with 18F-labeled NaF and PET (18F PET) in detecting osteolytic and osteoblastic metastases and its dependency on their anatomic localization. METHODS: Forty-four patients with known prostate, lung or thyroid carcinoma were examined with both planar RNB and 18F PET. A panel of reference methods including MRI of the spine, 1311 scintigraphy, conventional radiography and spiral CT was used as the gold standard. RNB and 18F PET were compared by a lesion-by-lesion analysis using a five-point score for receiver operating characteristic (ROC) curve analysis. RESULTS: 18F PET showed 96 metastases (67 of prostate carcinoma and 29 of lung or thyroid cancer), whereas RNB revealed 46 metastases (33 of prostate carcinoma and 13 of lung or thyroid cancer). All lesions found with RNB were also detected with 18F PET. Compared with 18F PET and the reference methods, RNB had a sensitivity of 82.8% in detecting malignant and benign osseous lesions in the skull, thorax and extremities and a sensitivity of 40% in the spine and pelvis. The area under the ROC curve was 0.99 for 18F PET and 0.64 for RNB. CONCLUSION: 18F PET is more sensitive than RNB in detecting osseous lesions. With RNB, sensitivity in detecting osseous metastases is highly dependent on anatomic localization of these lesions, whereas detection rates of osteoblastic and osteolytic metastases are similar. Higher detection rates and more accurate differentiation between benign and malignant lesions with 18F PET suggest the use of 18F PET when possible. PMID- 10520702 TI - Scintigraphic imaging and absorption of a 5-aminosalicylic acid enema in patients with ileorectal anastomosis. AB - Ileorectal anastomosis (IRA) is a possible surgical treatment for hyperacute and drug-unresponsive forms of ulcerative colitis (UC). UC relapses in the rectal remnant usually are prevented by chronic administration of 5-aminosalicylic acid (5-ASA) in topical formulations. The relationships between intestinal absorption and pattern of luminal spread of 5-ASA enemas are still unknown in patients with IRA. We correlated the absorption of a 5-ASA enema with its spread in the distal bowel of patients with IRA as assessed by 99mTc radioenema imaging. METHODS: Eight patients with UC in remission and previous IRA received a therapeutic 50-mL 5-ASA enema labeled with 99mTc-sulfer colloid. Absorbed 5-ASA and its major metabolite, acetyl 5-ASA, were measured in plasma, and dynamic images of radiolabeled enema were obtained for 6 h. The retrograde ileal spread (RIS) was determined and expressed as percentage of total enema radioactivity. Plasma levels of 5-ASA and acetyl 5-ASA were measured in six healthy volunteers after administration of the same enema volume with no radiolabeling. RESULTS: The mean 5-ASA plasma level was 0.70 microg/mL (range 0.37-0.95 microg/mL) in patients and 0.96 microg/mL (range 0.78-1.16 microg/mL) in healthy volunteers (P = not significant), and the mean acetyl 5-ASA plasma levels were 0.89 microg/mL (range 0.44-1.19 microg/mL) and 0.84 microg/mL (range 0.51-1.02 microg/mL), respectively (P = not significant). Radioenema imaging allows RIS assessment of patients with IRA. The mean value was 8.5% (range 2%-19.3%) of administered radioactivity, which correlated significantly with the total absorption of 5-ASA in the IRA group (P = 0.033, linear correlation test). Rectal wall contractions recognized by dynamic radioenema imaging were defined as a common cause of RIS episodes. CONCLUSION: In IRA patients, 5-ASA plasma levels were similar to those in healthy volunteers after administration in enema. Only part of a 50-mL 5-ASA enema reaches the ileum, and radiolabeled imaging shows the degree and number of these RIS episodes. The absorption of 5-ASA can increase in patients compared with healthy volunteers, in the presence of either occasional but significant ileal spread associated with postural factors and abdominal wall contraction or multiple moderate episodes of radioenema backdiffusion related to rectal wall motility. PMID- 10520703 TI - Evaluation of neoadjuvant therapy response of osteogenic sarcoma using FDG PET. AB - According to the current treatment protocol of the Cooperative Osteosarcoma Study (COSS), monitoring preoperative chemotherapy response and estimating grade of tumor regression in patients with osteosarcoma is mandatory before surgical removal of the tumor, particularly if a limb salvage procedure is intended. In addition, response to neoadjuvant chemotherapy is considered as an important prognostic indicator. The aim of this prospective study was to assess the usefulness of 2-(18F) fluoro-2-deoxy-D-glucose (FDG) PET in the noninvasive evaluation of neoadjuvant chemotherapy response in osteosarcoma. METHODS: In 27 patients with osteosarcoma, we determined tumor-to-background ratios (TBRs) of FDG uptake with PET, before and after neoadjuvant chemotherapy according to COSS 86c or COSS 96 protocols, respectively. We compared changes in glucose metabolism of osteosarcomas with the histologic grade of regression in the resected specimen, according to Salzer-Kuntschik, discriminating responders (grades I-III; n = 17) and nonresponders (grades IV-VI; n = 10). RESULTS: The decrease of FDG uptake in osteosarcomas expressed as a ratio of posttherapeutic and pretherapeutic TBRs showed a close correlation to the amount of tumor necrosis induced by polychemotherapy (P < 0.001; Spearman). With a TBR ratio cutoff level of 0.6, all responders and 8 of 10 nonresponders could be identified by PET. In addition, lung metastases of osteosarcoma were detected with FDG PET in 4 patients. CONCLUSION: FDG PET provides a promising tool for noninvasive evaluation of neoadjuvant chemotherapy response in osteosarcoma. This could imply consequences for the choice of surgical strategy, because a limb salvage procedure cannot be recommended in patients nonresponsive to preoperative chemotherapy unless wide surgical margins can safely be achieved. PMID- 10520704 TI - Preoperative assessment of residual hepatic functional reserve using 99mTc-DTPA galactosyl-human serum albumin dynamic SPECT. AB - Preoperative assessment of residual hepatic functional reserve offers important strategic information for hepatic resection. To predict the postoperative residual liver function, we assessed the value of hepatic 99mTc diethylenetriamine pentaacetic acid-galactosyl-human serum albumin (99mTc-GSA) clearance estimated by dynamic SPECT analysis. METHODS: We investigated 114 consecutive patients with liver disease, including 55 hepatectomy cases. One minute after injection of 185 MBq 99mTc-GSA, 15 serial dynamic SPECT images were obtained every minute. The initial five sets of SPECT images were analyzed by Patlak plot to estimate the sequential initial hepatic 99mTc-GSA clearance (mL/min) as an index of hepatic function. The sum of hepatic 99mTc-GSA clearance of the segments immune from resection was categorized as predicted residual 99mTC GSA clearance. In the hepatectomy cases, scintigraphy was performed before and 37 +/- 10 d after the operation. RESULTS: Good correlation was observed between the total hepatic 99mTc-GSA clearance and conventional hepatic function tests: plasma retention rate of iodocyanine green (ICG) at 15 min (ICG R15), r = -0.600, P < 0.0001, n = 94; plasma disappearance rate of ICG (K ICG), r = 0.670, P < 0.0001, n = 83; cholinesterase, r = 0.539, P < 0.0001, n = 121; serum albumin, r = 0.421, P = 0.0001, n = 123; and hepaplastin test, r = 0.456, P < 0.0001, n = 120. There was good correlation between the predicted residual 99mTc-GSA clearance and the postoperative total hepatic 99mTc-GSA clearance in patients who underwent segmentectomy or lobectomy (r = 0.84, P < 0.0001, n = 28) and between the pre- and postoperative total hepatic 99mTc-GSA clearance in patients who underwent subsegmentectomy (r = 0.91, P < 0.0001, n = 25). Five patients who had postoperative complications due to hepatic insufficiency (2 patients died of postoperative hepatic failure within 2 mo after operation) showed significantly lower predicted residual 99mTc-GSA clearance compared with the patients without complications (90.3 +/- 37.2 versus 320.9 +/- 158.8 mL/min; P < 0.005). CONCLUSION: The total hepatic 99mTC-GSA clearance reflected hepatic function. In addition, preoperative predicted residual hepatic 99mTc-GSA clearance was a good indicator of postoperative hepatic function and early prognosis. 99mTc-GSA dynamic SPECT is assumed to be a useful method for determining the surgical strategy in patients with hepatic tumor and especially in patients with hepatic dysfunction. PMID- 10520705 TI - Clinical usefulness of scintigraphy with 99mTc-galactosyl-human serum albumin for prognosis of cirrhosis of the liver. AB - Scintigraphy with 99mTc-diethylenetriamine pentaacetic acid-galactosyl-human serum albumin (99mTc-GSA) is useful for evaluating hepatic functional reserve. We assessed the clinical usefulness of this technique, including its value in establishing a prognosis, in patients with cirrhosis of the liver. METHODS: Scintigraphy with 99mTc-GSA was performed in 10 healthy subjects, 42 patients with chronic hepatitis and 158 patients with cirrhosis. Computer acquisition of gamma camera data were started just before the injection of 99mTc-GSA. Time activity curves for the heart and liver were generated from regions of interest (ROIs) for the heart and the entire liver. A receptor index was calculated by dividing the radioactivity of the liver ROI by that of the liver-plus-heart ROI 15 min after the injection. An index of blood clearance was calculated by dividing the radioactivity of the heart ROI at 15 min by that of the heart ROI at 3 min. RESULTS: The median receptor index was lower in patients with cirrhosis than in patients with chronic hepatitis or in healthy subjects, and the median index of blood clearance was higher. The receptor index was significantly lower when a complication (varices, ascites) was present. The index of blood clearance was significantly higher when a complication (varices and ascites) was present. Correlation of the two indices with classic indicators for functional reserve was significant. On the basis of the receptor index, the patients with cirrhosis were divided into two groups of roughly equal size: group A, receptor index over 0.85, and group B, receptor index 0.85 or less. On the basis of the index of blood clearance, the patients with cirrhosis were divided into two groups of roughly equal size: group A, index of blood clearance < 0.70, and group B, index of blood clearance > or = 0.70. The cumulative survival rates were lower in group B than in group A. CONCLUSION: Scintigraphy with 99mTc-GSA is clinically useful, especially in establishing the prognosis of patients with cirrhosis of the liver. PMID- 10520706 TI - Quality assurance in PET: evaluation of the clinical relevance of detector defects. AB - Defective detector blocks in PET may cause serious image artifacts. To estimate the influence of malfunctioning detectors on image quality, a method is described for transferring the actual detector defect onto previously acquired scans. METHODS: Consequences of detector defects of varying types and extensions were simulated in phantom studies as well as in clinical 18F-fluorodeoxyglucose investigations. First, a condition frame was obtained by dividing the sinogram of a blank measurement, obtained with rod sources on the defective PET camera, by the sinogram of a reference blank acquired before the appearance of the defect. Second, the sinogram of a previously acquired typical patient study was multiplied by the condition frame and reconstructed. Thereafter, images from corrupted sinograms were compared visually with their originals. For repairing defective sinograms, linear interpolation and the constrained Fourier space method were tested. RESULTS: The effects of detector defects can be simulated accurately in patient studies. The correction methods applied are especially helpful in cases of (a) several neighboring defective detectors and small study objects, (b) small hot artifacts and (c) several nonadjacent defective detectors. Linear interpolation is faster than the constrained Fourier space method; it is more widely applicable and provides similar results. CONCLUSION: The proposed approach allows specific evaluation of clinical consequences of detector defects. This technique simplifies the decision as to whether a planned patient study can be performed or must be postponed. Even in cases of serious detector problems, sinogram repair may help eliminate image artifacts and minimize the loss of image quality. PMID- 10520707 TI - Regions of interest in the venous sinuses as input functions for quantitative PET. AB - As clinical PET becomes increasingly available, quantitative methods that are feasible in busy clinical settings are becoming necessary. We investigated the use of intracranial blood pools as sources of an input function for quantitative PET. METHODS: We studied 25 patients after the intravenous injection of [18F]6 fluoro-L-m-tyrosine and compared sampled blood time-activity curves with those obtained in small regions of interest (ROIs) defined in the blood pools visible in the PET images. Because of the comparatively large dimensions of the blood pool at the confluence of the superior sagittal, straight and transverse sinuses, a venous ROI input function was chosen for further analysis. We applied simple corrections to the ROI-derived time-activity curves, deriving expressions for partial volume, spillover and partition of tracer between plasma and red blood cells. The results of graphic and compartmental analysis using both sampled [Cs(t)] and ROI [Cr(t)] venous input functions for each patient were compared. We also used an analytic approach to examine possible differences between venous and arterial input functions in the cerebral circulation. RESULTS: Cr(t) peaked significantly earlier and higher than Cs(t) in this patient population, although the total integral under the curves did not differ significantly. We report some apparent differences in the results of modeling using the two input functions; however, neither the graphically determined influx constant, Ki, nor the model parameter that reflects presynaptic dopaminergic metabolism, k3, differed significantly between the two methods. The analytic results suggest that the venous ROI input function may be closer to the arterial supply of radiotracer to the brain than arterialized venous blood, at least in some patient populations. CONCLUSION: We present a simple method of obtaining an input function for PET that is applicable to a wide range of tracers and quantitative methods and is feasible for diagnostic PET imaging. PMID- 10520708 TI - Noise removal using factor analysis of dynamic structures: application to cardiac gated studies. AB - Factor analysis of dynamic structures (FADS) facilitates the extraction of relevant data, usually with physiologic meaning, from a dynamic set of images. The result of this process is a set of factor images and curves plus some residual activity. The set of factor images and curves can be used to retrieve the original data with reduced noise using an inverse factor analysis process (iFADS). This improvement in image quality is expected because the inverse process does not use the residual activity, assumed to be made of noise. The goal of this work is to quantitate and assess the efficiency of this method on gated cardiac images. METHODS: A computer simulation of a planar cardiac gated study was performed. The simulated images were added with noise and processed by the FADS-iFADS program. The signal-to-noise ratios (SNRs) were compared between original and processed data. Planar gated cardiac studies from 10 patients were tested. The data processed by FADS-iFADS were subtracted to the original data. The result of the substraction was studied to evaluate its noisy nature. RESULTS: The SNR is about five times greater after the FADS-iFADS process. The difference between original and processed data is noise only, i.e., processed data equals original data minus some white noise. CONCLUSION: The FADS-iFADS process is successful in the removal of an important part of the noise and therefore is a tool to improve the image quality of cardiac images. This tool does not decrease the spatial resolution (compared with smoothing filters) and does not lose details (compared with frequential filters). Once the number of factors is chosen, this method is not operator dependent. PMID- 10520709 TI - Prediction of recovery of left ventricular dysfunction after acute myocardial infarction: comparison between 99mTc-sestamibi cardiac tomography and low-dose dobutamine echocardiography. AB - The aim of this study was to evaluate the role of 99mTc-sestamibi cardiac imaging and dobutamine echocardiography in detecting myocardial viability early after acute myocardial infarction. METHODS: Forty-nine patients (mean age 52 +/- 10 y) underwent coronary angiography, low-dose dobutamine echocardiography, radionuclide angiography and rest 99mTc-sestamibi imaging within 10 d after myocardial infarction. Of these patients, 19 were revascularized and 30 were treated medically. Resting echocardiogram and radionuclide angiography were repeated 8 mo later to evaluate segmental functional recovery and changes in left ventricular (LV) ejection fraction, respectively. RESULTS: In revascularized patients, 61 of 108 akinetic or dyskinetic segments showed functional recovery. In these patients, sensitivity in predicting segmental functional recovery was 87% for sestamibi imaging and 66% for dobutamine echocardiography (P < 0.001), whereas specificity and accuracy were comparable. Sestamibi activity (> or =55% of peak) was the strongest predictor of segmental functional recovery (P < 0.001) and of LV ejection fraction improvement > or =5% (P < 0.01) after revascularization. In medically treated patients, 60 of 149 akinetic or dyskinetic segments showed functional recovery. In these patients, the majority (94%) of segments with contractile reserve on dobutamine were viable on sestamibi imaging and 86% of them improved function at follow-up. Functional recovery was poor in segments without contractile reserve either with (38%) or without (62%) preserved sestamibi uptake. Inotropic response was the best predictor of segmental (P < 0.001) and global (P < 0.01) LV functional improvement in medically treated patients. CONCLUSION: Dobutamine echocardiography predicts spontaneous functional recovery after acute myocardial infarction. However, sestamibi imaging is useful to identify patients with dysfunctional myocardium without contractile reserve who may benefit from coronary revascularization. PMID- 10520711 TI - Clues to prognosis in congestive heart failure. PMID- 10520710 TI - Left ventricular volumes and ejection fraction calculated from quantitative electrocardiographic-gated 99mTc-tetrofosmin myocardial SPECT. AB - We compared the left ventricular (LV) end-diastolic volume (EDV), end-systolic volume (ESV) and ejection fraction (LVEF) as calculated by Cedars automated quantitative gated SPECT (QGS) to those determined by first-pass radionuclide angiography (FPRNA) and contrast left ventriculography (LVG) in a group of 21 patients (mean age 61.4 +/- 9.2 y). METHODS: A total of 740 MBq 99mTc-tetrofosmin was administered rapidly into the right cubital vein at rest, and FPRNA was performed using a multicrystal gamma camera. One hour after injection, QGS was performed with a temporal resolution of 10 frames per R-R interval. LVG was performed within 2 wk. RESULTS: The EDV, ESV and LVEF calculated by QGS were highly reproducible (intraobserver, r = 0.99, r = 0.99 and r = 0.99, respectively; interobserver, r = 0.99, r = 0.99 and r = 0.99, respectively; P < 0.01) and were more consistent than those determined by FPRNA (intraobserver, r = 0.97, r = 0.95 and r = 0.93, respectively; interobserver, r = 0.86, r = 0.96 and r = 0.91, respectively; P < 0.01). There was a good correlation between EDV, ESV and LVEF by FPRNA and those by LVG (r = 0.61, r = 0.72 and r = 0.91, respectively; P < 0.01), and there was an excellent correlation between QGS and LVG (r = 0.73, r = 0.83 and r = 0.87, respectively; P < 0.01). The mean EDV by QGS (100 +/- 11.3 mL) was significantly lower than by FPRNA (132 +/- 16.8 mL) or LVG (130 +/- 8.1 mL), and the mean ESV by QGS (53.8 +/- 9.3 mL) was lower than by FPRNA (73.0 +/- 13.3 mL). Ejection fraction values were highest by LVG (57.1% +/- 3.2%), then QGS (51.8% +/- 3.0%) and FPRNA (48.9% +/- 2.4%). CONCLUSION: QGS gave more reproducible results than FPRNA. LV volumes and LVEF calculated by QGS correlated well to those by LVG. PMID- 10520712 TI - Radiolabeled thymidine: a sensitive tracer for early tumor response and recurrence after irradiation. AB - This study evaluated the sensitivity of a radiolabeled thymidine tracer for assessment of early tumor response and recurrence after irradiation. METHODS: SW707 human colon carcinoma implanted into nude mice was irradiated with 6 or 20 Gy. Tumor volume was determined for an interval of 14 d. At 4, 8 and 24 h and at 2, 3, 7, 10 and 14 d after irradiation, [14C]thymidine uptake into the tumor was determined with a liquid scintillation counter and the intratumoral distribution of [14C]thymidine was visualized and evaluated semiquantitatively by autoradiography using a phosphor imager. RESULTS: In both groups, tumor volume decreased until day 7 after irradiation; afterward, regrowth occurred in only the group that had received 6 Gy. A decrease in thymidine uptake was found as early as 8 h after irradiation. On day 3 after irradiation, thymidine uptake increased again in the 6-Gy group, before the increase in tumor volume, but remained unchanged in the 20-Gy group. Also on day 3, multiple foci of thymidine uptake suggesting proliferation preceding tumor recurrence were seen on autoradiographs from the 6-Gy group but not from the 20-Gy group. Histological findings correlated with the results of autoradiography. CONCLUSION: The results show that radiolabeled thymidine is a sensitive tracer for assessment of early tumor response and recurrence after irradiation. The rapid decrease in uptake, however, does not allow any prediction about tumor recurrence. PMID- 10520713 TI - Oncological applications of FDG PET imaging. PMID- 10520714 TI - Radiation dose to the testes after 131I therapy for ablation of postsurgical thyroid remnants in patients with differentiated thyroid cancer. AB - Radioiodine-131 is used in differentiated thyroid cancer (DTC) for ablation of postsurgical thyroid remnants and destruction of metastases. The question may be raised of whether 131I treatment of DTC in male patients may give an irradiation dose to the testes that could impair fertility. Few data in the literature concern the dose absorbed by the testes after 1311 therapy for DTC. Because 131I kinetics may be altered by the hypothyroid condition commonly present at the time of treatment and by the radioiodinated iodoproteins released by the damaged thyroid tissue, the dose values reported in the International Commission on Radiological Protection (ICRP) tables for euthyroid men may not be appropriate. To clarify this problem, three male subjects undergoing 131I therapy for ablation of thyroid remnants shortly after thyroidectomy for DTC were studied. METHODS: The mean administered activity was 1256 MBq, and the duration of the study was 2 wk. The gamma dose was measured by thermoluminescent dosimeters (TLDs) applied to the lower poles of the testes. Correction factors were calculated for the distance of the TLD from the center of the testes and for attenuation by the testes of the gamma rays reaching the TLD. After correction, the gamma dose to the testes ranged from 21 to 29 mGy. The gamma dose calculated by the Medical Internal Radiation Dose (MIRD) method from blood and urine samples was similar (18-20 mGy) to that measured by TLDs. The beta dose was estimated by the MIRD method from blood activity and testicular volume and ranged between 14 and 31 mGy. RESULTS: The total (beta and gamma) doses to testes were 30, 33 and 43 microGy/MBq in the three subjects. CONCLUSION: These values are close to those derived from the ICRP tables (26-37 microGy/MBq 131I) for euthyroid subjects. The present data indicate that significant irradiation is delivered to the testes after the administration of the 131I ablative dose to thyroidectomized patients. The relevance of the radiation absorbed by testes on fertility remains to be established. PMID- 10520716 TI - Biodistribution, dosimetry and metabolism of 11beta-methoxy-(17alpha,20E/Z) [123I]iodovinylestradiol in healthy women and breast cancer patients. AB - The biodistribution and dosimetry of the 20E and 20Z stereoisomers of 11 beta methoxy-(17alpha,20)-[123I]iodovinylestradiol (MIVE) were evaluated in six healthy women. Tumor uptake and metabolism of the 20Z isomer were evaluated in 13 women referred after abnormal mammography or after discovery of a suspect mass at physical examination. METHODS: The radiopharmaceuticals were prepared from their corresponding stannyl intermediates and administrated intravenously. Blood samples were drawn at different time intervals and urine was collected for up to 24 h. Metabolites were detected by radiochromatography. Tissue distribution was followed for up to 24 h by scintigraphic imaging. The dosimetry was computed according to the Medical Internal Radiation Dose scheme. RESULTS: The 20E and 20Z isomers exhibit similar biodistribution and dosimetry patterns. Chromatographic analysis of plasma samples of healthy volunteers and cancer patients, as well as in vitro plasma incubations, confirmed the in vivo stability of (20Z)-[123I]MIVE. Radioactivity was rapidly cleared from the blood by the liver and excreted through the gut, which received the highest radiation dose (0.211 mGy/MBq). The effective doses for the adult female and male phantom were 0.054 and 0.046 mSv/MBq, respectively. Among the 13 patients imaged with (20Z)-[123I]MIVE, 3 had fibrocystic disease with no focal uptake, 8 had good agreement with in vitro estrogen receptor determination and 2 were false-positive. CONCLUSION: The radiation dose after intravenous administration of 20E- or (20Z)-[123I]MIVE at imaging dose levels is within acceptable limits. There was a good correlation between uptake of (20Z)-[123I]MIVE and the presence of estrogen receptors in breast cancer patients. PMID- 10520715 TI - Preparation of alpha-emitting 213Bi-labeled antibody constructs for clinical use. AB - Preclinical evaluation of alpha particle-emitting 213Bi-labeled antibody constructs have demonstrated the specificity and potency of these agents in a variety of cancer systems. The transition of a 213Bi-radiolabeled antibody from a preclinical construct to a clinical drug represented a difficult task that involved development of reliable and validated methods to provide multiple MBq quantities of a pure, immunoreactive agent that met pharmaceutical standards to treat patients. METHODS: The methods used for the preparation of (213Bi)CHX-A diethylenetriamine pentaacetic acid (DTPA)-HuM195, an alpha particle-emitting anti-CD33 antibody construct for therapy of myeloid leukemias, is used as a specific example. This article describes methods for reagent purification, drug labeling, radioprotection and chromatographic purification. Quality of the drug is evaluated using radiochemical incorporation and purity assays with instant thin-layer chromatography (ITLC) and high-performance liquid chromatography (HPLC), determination of cell-based antibody total immunereactivity, small animal safety, pyrogen level, sterility and radionuclidic purity. RESULTS: Sixty-seven doses were prepared. Individual doses ranged from 148 to 814 MBq. Specific activities ranged from 329 to 766 MBq/mg. The radiolabeling efficiency (median +/ SD) of CHX-A-DTPA-HuM195 with 213Bi was 81% +/- 9% (n = 67) after 9 min. The construct was purified by size-exclusion chromatography and was found to be 99% +/- 2% pure (n = 67) by either ITLC or HPLC methods. The immunoreactivity of (213Bi)CHX-A-DTPA-HuM195 was 89% +/- 9% (n = 44) and was independent of the specific activity. The formulated pharmaceutical was found to contain < or =4 +/- 1 EU/mL pyrogens (n = 66); all samples examined were sterile. An 225Ac radionuclidic impurity was present at a level of 0.04 +/- 0.03 x 10(-6)/mL (n = 10) in a product volume of 7.4 +/- 0.5 mL (n = 67). Each of the 67 doses was injected intravenously into patients without complication as part of a phase I clinical trial. CONCLUSION: These data show that 213Bi-labeled antibody constructs can be prepared and administered safely to humans at a wide range of therapeutic levels. PMID- 10520717 TI - Noninvasive quantification of cerebral blood flow using 99mTc-ECD and SPECT. AB - The aim of this study was to develop a simple, noninvasive method for quantifying regional cerebral blood flow (rCBF) using 99mTc-ethyl cysteinate dimer (ECD) by a single SPECT scan and single venous sampling. METHODS: Using a three-compartment model, we introduced the regional brain fractionation index (BFI), Cb(Ts)/integral of 0-Ts Ca(tau)dtau [Ca(t), arterial input; Cb(t), brain activity]. Regional BFI obtained at the optimum time Ts (min) was converted to rCBF using an exponential function, which was obtained by analyzing the relationship between regional BFI and rCBF (= F) obtained by the standard 133Xe inhalation SPECT method. The integral of the concentration of 99mTc-ECD in arterial blood corrected for physical decay [Ca(t)] in BFI was estimated from a single venous blood sample obtained at the optimum time Tv using the regression line obtained by analyzing the relationship between the integral of Ca(t) and venous sample data. The data come from three groups of patients. The first group of patients (n = 16) underwent a complete 99mTc-ECD BFI study with measurement of Ca(t) and dynamic SPECT scanning, as well as a 133Xe inhalation study to measure rCBF The results were used to analyze the relationship between regional BFI and rCBF (obtained with 133Xe) and to determine the optimum time Ts for obtaining BFI. Data from the second group of patients (n = 15) were used to analyze the relationship between the integral of Ca(t) and venous sample data and to determine the optimum time Tv for one-point venous blood sampling. Finally, the third group of patients (8 patients, 10 studies) was used to validate the current method by comparing the results with 133Xe inhalation SPECT. RESULTS: Regional BFI obtained at time Ts = 20 min showed good agreement (r = 0.907; a = 0.552, b = 0.962) with rCBF. The venous sample data obtained at time Tv = 6 min showed a good correlation (r = 0.988) with BFI. In comparing rCBF values thus obtained and those obtained by the 133Xe method, we found a good correlation (r = 0.917, slope = 1.01). CONCLUSION: The proposed method has three advantages: (a) accurate quantification of rCBF without underestimation in the high flow range, (b) simplicity and noninvasiveness and (c) the ability to use any type of SPECT camera for the study. PMID- 10520718 TI - A novel, simple method of functional spleen volume calculation by liver-spleen scan. AB - Spleen enlargement is commonly associated with portal hypertension from cirrhosis and may cause thrombocytopenia. Thus, accurate assessment of spleen size may be helpful in the clinical evaluation. Spleen length is not a precise estimate of spleen size because of the variation in spleen configuration, and spleen volumes measured by edging techniques can be tedious. We present a new method of measuring the functional spleen volume by liver-spleen scan (LSSs), validation experiments and some clinical data. METHODS: The method involves measurement of the total spleen counts by SPECT and dividing by a representative voxel concentration on a single frame to obtain the organ volume. Validation included phantom studies and clinical evaluation in 443 consecutive patients, including 216 with histologic assessments of chronic liver disease (CLD) and 11 healthy volunteers. RESULTS: A calibration factor determined from phantoms was used to convert the calculated volume (CV) to the "true" volume (V): V = CV (0.956) - 66.5 (r = 0.9991; P < 0.001). The volume calculations were validated in a second group of phantoms (r= 0.981; P < 0.0001). Spleen volumes were expressed as volume (cm3) and as volume per pound ideal body weight (IBW) (cm3/lb) (the conversion factor to convert cm3/lb IBW to cm3/kg IBW is 2.2). Clinical studies of reproducibility included demonstration of a significant (P < 0.0001) linear correlation between volumes calculated from repeat LSSs within 9 mo of the initial LSS in 11 healthy volunteers and 32 patients with CLD: y = 1.02x - 25; r = 0.968. The correlation with spleen volumes from autopsy or splenectomy was significant: y = 0.766x + 57; r = 0.845; P < 0.001. The normal spleen volume in 11 patients was 201 +/- 77 cm3 and 1.43 +/- 0.68 cm3/lb IBW (upper limits of normal: 335 cm3 or 2.5 cm3/lb IBW). In 443 consecutive LSSs over 15 mo, half of the patients had spleen volumes above the upper limits of healthy volunteers, and CLD was present in 90.9% of these patients. In 216 patients with histologically proven liver disease, a progressive increase in the percentage of spleen volumes above the upper limits of normal was noted from no fibrosis (10%) to mild to moderate fibrosis (36.7%) to early cirrhosis (52%) to advanced liver disease (75%). The correlation of spleen volume with platelet count was excellent (r = 0.7635; P < 0.005). CONCLUSION: This novel spleen volume measurement detects serious liver disease and correlates with splenic hyperfunction. PMID- 10520719 TI - A novel potential application for 99mTc-HMPAO: endothelial cell labeling for in vitro investigation of cell-biomaterial interactions. AB - Good adherence of endothelial cells (ECs) seeded on vascular prostheses and cell retention under flow conditions are important factors to consider in the use of functionalized prostheses in vascular surgery. Because 111In-oxine radiolabeling presents disadvantages, we wondered whether, because of its well-known physical properties, 99mTc-hexamethyl propyleneamine oxime (HMPAO or exametazime) could be used. METHODS: The cytotoxicity of unlabeled HMPAO and 99mTc-HMPAO at increasing concentrations and activities was tested on monolayers of the EC line EA-hy-926. The influence of temperature and time on tracer incorporation into cells was also tested. The optimal labeling conditions were applied to evaluate the retention of ECs seeded on polyester grafts under flow conditions by gamma camera detection. RESULTS: The activity of 10 MBq/10(6) cells corresponding to 4.5 microg/10(6) cells of unlabeled HMPAO, applied for 3 h at 37 degrees C (cellular uptake = 18%), was the best compromise between the maintenance of cell viability and metabolic activity and efficient detection by the gamma camera. Spontaneous leakage was observed and analyzed by high-performance liquid chromatography. A cell loss of 13% after 180-min exposure to shear stress was obtained. CONCLUSION: Our data thus indicate the feasibility of using such a radiolabeling technique to investigate EC-biomaterial interactions. PMID- 10520720 TI - 131I radioimmunotherapy and fractionated external beam radiotherapy: comparative effectiveness in a human tumor xenograft. AB - This article compares the effectiveness of radiation delivered by a radiolabeled monoclonal antibody, 131I-labeled A33, that targets colorectal carcinoma, with that of 10 fractions of conventional 320 kVp x-rays. METHODS: Human colorectal cancer xenografts (SW1222) ranging between 0.14 and 0.84 g were grown in nude mice. These were treated either with escalating activities (3.7-18.5 MBq) of 131I labeled A33 or 10 fractions of 320 kVp x-rays (fraction sizes from 1.5 to 5 Gy). Tumor dosimetry was determined from a similar group of tumor-bearing animals by serial kill, tumor resection and counting of radioactivity in a gamma counter. The relative effectiveness of the two radiation therapy treatment approaches was compared in terms of tumor regrowth delay and probability of tumor cure. RESULTS: The absorbed dose to tumor per MBq administered was estimated as 3.7 Gy (+/-1 Gy; 95% confidence interval). We observed a close to linear increase in tumor regrowth delay with escalating administered activity. Equitumor response of 1311 monoclonal antibody A33 was observed at average radiation doses to the tumor three times greater than when delivered by fractionated external beam radiotherapy. The relationship between the likelihood of tumor cure and administered activity was less predictable than that for regrowth delay. CONCLUSION: The relative effectiveness per unit dose of radiation therapy delivered by 131I-labeled A33 monoclonal antibodies was approximately one third of that produced by fractionated external beam radiotherapy, when measured by tumor regrowth delay. PMID- 10520721 TI - 99mTc-MIBI in differentiated thyroid carcinoma. PMID- 10520722 TI - Attribution of use of perchlorate in parathyroid scintigraphy. PMID- 10520723 TI - Beneficial effects of MET-88 on left ventricular dysfunction and hypertrophy with volume overload in rats. AB - We examined the effects of MET-88 on haemodynamics and cardiac hypertrophy in rats with an aortocaval shunt (A-V shunt). On the day of surgery, an A-V shunt was produced by using an 18-gauge needle in Wistar rats as described by Garcia and Diebold. MET-88 and captopril were orally administered to rats 1 week after surgery, and the administration was continued for 3 weeks. Four weeks after the surgery, A-V shunt-operated rats had biventricular hypertrophy and higher right atrial pressure (RAP) and left ventricular end-diastolic pressure (LVEDP) than sham-operated rats. Compared with untreated A-V shunt rats, those treated with MET-88 showed significant attenuation of the development of left ventricular (LV) hypertrophy and of the increased LVEDP. Captopril-treated A-V shunt rats also failed to show increases in LV weight and LVEDP. In in vitro studies, MET-88 had no effect on renin and angiotensin-converting enzyme (ACE) activities in the plasma of normal rats. These results suggest that MET-88 improved LV hypertrophy and LV dysfunction in rats with an A-V shunt. Furthermore, the data indicate that the beneficial effects of MET-88 may be attributed to some pathway, not involving the renin-angiotensin system, such as myocardial energy metabolism, venous return, etc. We conclude that MET-88 may be a novel agent for the therapy of chronic heart failure. PMID- 10520724 TI - Differential effects of pinacidil and levcromakalim on the contractions elicited electrically or by noradrenaline in the portal vein of the rabbit. AB - The present study was undertaken to examine the antivasoconstrictor effects of pinacidil and levcromakalim, two potassium channel openers (PCOs), on the isolated rabbit portal vein and to define the role for different subtypes of pre- and/or post-synaptic K+ channels in the antivasoconstrictor action of the PCOs. The vein strips were contracted by electrical field stimulation (EFS) or by exogenous noradrenaline (NA). The results of this study showed that pinacidil produced a more potent inhibition of the neurogenic contractions (pD2 = 6.04 +/- 0.05) than of contractions induced by exogenous NA (pD2 = 4.90 +/- 0.10). Glibenclamide (1 microM), a selective blocker of adenosine triphosphate (ATP) sensitive K+ channels (K(ATP)), did not affect the pinacidil-induced inhibition of contractions evoked by exogenous NA. In contrast, glibenclamide (0.1-10 microM) significantly antagonized the effect of pinacidil on EFS evoked contractions in a noncompetitive manner. There was no difference between the inhibitory effects of levcromakalim on neurogenic contractions (pD2 = 7.58 +/- 0.05) and contractions evoked by exogenous NA (pD2 = 7.64 +/- 0.08). Glibenclamide (1 microM) antagonized in the same manner the levcromakalim-induced inhibition of neurogenic contractions and contractions evoked by exogenous NA. Moreover, glibenclamide competitively antagonized the effect of levcromakalim on EFS induced contractions of the rabbit portal vein (pA2 = 6.40 +/- 0.10). Charybdotoxin (0.4 microM) and apamin (0.1 microM) did not influence the inhibitory effects of pinacidil and levcromakalim, both on contractions evoked by EFS and contractions evoked by exogenous NA. These results suggest that the antivasoconstrictor effect of levcromakalim might be postsynaptic and associated with opening of the smooth muscle K(ATP) channels. In contrast, it is hypothesized that the effect of pinacidil on neurogenic contractions is due to an interference with K(ATP) channels in the neuromuscular synapse. It seems that the action of pinacidil on the NA contractions is mediated by another still undefined mechanisms of pinacidil. PMID- 10520726 TI - Antagonism by pimozide of olanzapine-induced hypothermia. AB - The atypical antipsychotic olanzapine (2.5-20 mg/kg) produced hypothermia in rats. The decrease in rectal temperature caused by olanzapine (2.5-20 mg/kg) was blocked by the selective dopamine D2 receptor antagonist pimozide (0.5 and 1 mg/kg) but not by the dopamine D1 receptor antagonist SCH 23390 (0.5 and 1 mg/kg). The dopamine D1/D2 receptor agonist apomorphine (3 mg/kg) and the selective dopamine D2 receptor agonist talipexole (0.5 mg/kg) produced hypothermia in rats. Olanzapine (10 and 20 mg/kg) significantly blocked hypothermia produced by both apomorphine and talipexole while the lower doses (2.5 and 5 mg/kg) of olanzapine failed to block it. The present results demonstrate that olanzapine behaves as a partial agonist at brain DA D2 receptor populations involved in thermoregulation in the rat. PMID- 10520725 TI - Antioxidant properties of aminoguanidine. AB - It is well known that aminoguanidine (AG) can diminish advanced glycosylation of proteins, which might be beneficial in preventing chronic diabetic complications. Recent reports suggested an inter-relationship between glycosylation of protein and free radical damage. In the present study, we examined the free radical scavenging properties of AG. Electron paramagnetic resonance using the spin-trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) was performed to determine the superoxide and hydroxyl radical scavenging abilities of AG. These experiments revealed that AG was an effective hydroxyl radical scavenger even though it expressed a direct inhibitory effect on the xanthine oxidase activity at high concentrations (AG > or = 5 mM). In the second part of the study, allophycocyanin was used as an indicator of free radical mediated protein damage. In the assay, 2,2'-azobis(2 amidinopropane) hydrochloride (AAPH) was used as a peroxyl radical generator, and the loss of allophycocyanin fluorescence was monitored. The antioxidant effect of AG was expressed in oxygen-radical absorbing capacity (ORAC), where one ORAC unit equals the net protection produced by 1 microM Trolox (a water soluble analogue of vitamin E) as a control standard. AG exhibited a significant dose-dependent effect against free radical damage. These radical scavenging properties of AG may contribute to protective effects during glycation and explain the prevention of diabetic complications. PMID- 10520727 TI - Fusobacterium necrophorum haemolysin stimulates motility of ileal longitudinal smooth muscle of the guinea-pig. AB - Fusobacterium necrophorum haemolysin (0.5-3.1 mg protein/mL) dose-dependently induced contractions of the isolated ileal longitudinal smooth muscle of the guinea-pig. The haemolysin (3.1 mg protein/mL) -induced maximum contraction of 75% of the response to 60 mM K+ declined within 17 min and the muscles then demonstrated rhythmic contractions. Tetrodotoxin (3.1 x 10(-6) M) had no effect on the contraction due to the haemolysin. After incubation in Ca(2+)-free medium, the ileal response to the haemolysin was lost. Verapamil, a Ca2+ channel blocker, dose-dependently inhibited the contraction to the haemolysin. The rabbit anti serum against F. necrophorum haemolysin inhibited the haemolysin-induced contraction of ileal muscle. The bacterial haemagglutinin and the lipopolysaccharide had no effect on the response of ileal muscle. These findings suggest that the haemolysin-induced direct stimulation of ileal motility dependant on Ca2+ influx will increase the probability of contact of F. necrophorum and ileal mucosa and could increase the chances of colonization for F. necrophorum. PMID- 10520728 TI - Analysis of Helicobacter pylori binding site on HEp-2 cells and three cell lines from human gastric carcinoma. AB - Helicobacter pylori (H. pylori) is a pathogen responsible for chronic gastritis and peptic ulcer diseases. It colonises the gastric mucus layer and adheres to the gastric epithelial cell surface. As this adherence is the first step of infection, it is important to study the adherence mechanism. The aim of this study was to analyse the specific binding assay of H. pylori to HEp-2 cells and three gastric phenotype cell lines, AGS, MKN-45 and AZ-521. H. pylori NCTC 11637 grown on agar plates was harvested and used in experiments. H. pylori was inoculated to pre-cultured cell monolayers. Adhered bacteria were labelled with an anti-H. pylori antibody and an FITC-conjugated secondary antibody and quantified by using a fluorescent plate reader. Microbial adherence to HEp-2 cells increased with incubation time and incubated concentration of H. pylori. No further increase was obtained with four or more hours of incubation or with a concentration of 4 x 10(7) bacteria/well or more. Scatchard analysis revealed a linear plot and the Bmax value was 88.3. Similar adherence patterns were obtained when AGS, AZ-521 and MKN-45 cells were used for adherence assays, but they had a lower binding affinity than HEp-2 cells and AZ-521. MKN-45 cells had less receptors than HEp-2 and AGS cells. In conclusion, H. pylori adhered to the cell surface could be quantified by this assay method. H. pylori adhesion to cell surfaces has a single population of binding site and one type of binding site on HEp-2, AGS, AZ-521 and MKN-45 cells. PMID- 10520729 TI - Pharmacological characterization of muscarinic receptors implicated in rabbit detrusor muscle contraction and activation of inositol phospholipid hydrolysis in rabbit detrusor and parotid gland. AB - In the present study, we evaluated the pharmacological characteristics of the functional muscarinic receptors implicated in rabbit detrusor contraction and coupled to inositol phospholipid turnover in rabbit detrusor and parotid gland. The selectivity of several muscarinic antagonists for detrusor vs. salivary gland muscarinic receptors was also examined. The affinities for the muscarinic m1-, m2 and m3-receptor subtypes were determined using membranes from human cloned receptors expressed in CHO-K1 cells using [3H]-N-methyl scopolamine as a radioligand. Anti-muscarinic activity was determined in isolated rabbit detrusor by measuring the displacement of the contractile response to carbachol, and in rabbit detrusor and rabbit parotid by measuring the displacement of inositol phospholipid hydrolysis (total inositol phosphate accumulation) to carbachol. A significant correlation was found between the potencies to antagonize carbachol induced rabbit detrusor contraction (pK(B)) and the affinities (pKi) for the m3 receptor subtype (r = 0.93, P = 5 x 10(-6)). Lower, but significant, correlations [0.88 (P = 6.3 x 10(-5)), 0.72 (P = 4.6 x 10(-3))] were obtained with m1- or m2 receptor subtypes, respectively. Each muscarinic antagonist tested displayed similar potency to antagonize carbachol-stimulated inositol phospholipid hydrolysis in rabbit detrusor and parotid (r = 0.96, P = 8 x 10(-3)). A significant correlation was found between the potencies to antagonize carbachol stimulated inositol phospholipid hydrolysis (pK(B)), determined in rabbit detrusor and rabbit parotid, and the affinities (pK(i)) for the m3-receptor subtype [r = 0.96 (P = 0.01), 0.99 (P = 5 x 10(-5)), respectively] and for the m1 receptor subtype [r = 0.98 (P = 3.5 x 10(-3)), 0.94 (P = 0.02), respectively] but not for the m2-receptor subtype [r = 0.33, 0.57, ns, respectively]. In each in vitro assay, methoctramine (preferential M2 selective antagonist) and pirenzepine (preferential M1 selective antagonist) were slightly potent. We suggest that the muscarinic receptor implicated in the response to carbachol in rabbit detrusor and parotid gland corresponds to the M3-subtype. None of the muscarinic antagonists studied in rabbit tissues displayed preferential affinity for the detrusor. PMID- 10520730 TI - Effects of oxazepam and acetaminophen on cicletanine metabolism in rat hepatocytes and liver microsomes. AB - Cicletanine, a racemic furopyridine derivative synthesized as racemate, is used as an antihypertensive agent. Its two enantiomers are involved in the pharmacological effects of the drug. Cicletanine is metabolized by conjugation enzyme systems (phase II) into sulfoconjugated or glucuroconjugated enantiomers. As oxazepam and acetaminophen are widely prescribed, especially to elderly patients, these two drugs may be co-administered with cicletanine. The metabolic profile and the kinetics of biotransformation were studied by using rat hepatocytes and liver microsomes. Cicletanine was extensively metabolized by rat hepatocytes. More than 80% of the drug was biotransformed after a 3 h incubation. The formation of glucuroconjugated metabolites was characterized by the following kinetic parameters, i.e. Vmax = 2.05 +/- 0.21 nmol/min/mg protein and Km = 287 +/ 6.7 microM for (-)-cicletanine, and Vmax = 1.44 +/- 0.12 nmol/min/mg protein and K(m) = 171 +/- 4.1 microM for (+)-cicletanine. Oxazepam inhibited the glucuronidation of cicletanine in both rat hepatocytes and liver microsomes with a competitive-type inhibition, i.e. K(i) = 129 +/- 7.5 and 152 +/- 19.7 microM for (-)-cicletanine and (+)-cicletanine, respectively. The co-incubation of acetaminophen with cicletanine showed that only sulfoconjugation was inhibited in rat hepatocytes. Glucuronidation was not modified by acetaminophen. As natriuric activity is due to sulfoconjugated (+)-cicletanine, acetaminophen could potentially modulate in vivo the pharmacological effect of cicletanine. The data of the in vitro study reported here suggested an interaction between cicletanine and oxazepam or cicletanine and acetaminophen. However, the clinical impact of such a drug interaction needs further evaluation. PMID- 10520731 TI - Interspecies variability and drug interactions of clozapine metabolism by microsomes. AB - Cytochrome P450 expression in liver is influenced by several factors, including species, sex and strain. We compared metabolism formation of clozapine in different species (rat, mouse, guinea-pig, dog, monkey and man) so as to choose between species to further validate interaction studies. Liver microsomes of male and female Sprague-Dawley rats, hairless rats, OF1 mice, Balb C mice and Dunkin Hartley albino guinea-pigs, male beagle dogs, male cynomolgus monkeys and man were used to investigate in vitro metabolism of clozapine. This process was dependent on the presence of NADPH and on the presence of microsome protein. In addition, we observed the formation of desmethyl- and N-oxide metabolites, with the rate of formation of each of these compounds varying with species, sex and strain of microsomes incubated. The desmethyl- and N-oxide metabolites formed were statistically greater in male than in female rats, mice in the two strains studied, as well as for the guinea-pigs. Levels of desmethyl clozapine formed were high for the rats and no significant difference in clozapine biotransformation was observed between Sprague-Dawley and hairless rats. For man, the formation of metabolites of clozapine was comparable with guinea-pig, dog and monkey. In addition, we screened the effect of 52 molecules, representative of 11 different therapeutic classes, on the metabolism of clozapine by rat liver microsomes. We found that most of the calcium channel blockers (diltiazem, felodipine, isradipine, lacidipine, nicardipine and nitrendipine), antifungals (ketoconazole, miconazole) and two anticancer drugs (paclitaxel, teniposide) caused more than 50% inhibition of clozapine metabolism in vitro. The extent of inhibition was increased in a concentration-dependant manner. Complementary clinical and pharmacokinetic studies should be performed to confirm these results. PMID- 10520732 TI - Effects of 50mg amisulpride on EEG, psychomotor and cognitive functions in healthy sleep-deprived subjects. AB - Amisulpride, a substituted benzamide, binds selectively to the dopamine D2- and D3-receptors. It has higher affinity for limbic compared to striatal dopamine receptors in vivo. At low doses, amisulpride facilitates dopamine transmission via a selective blockade of presynaptic D2- and D3-receptors. Amisulpride is an active antipsychotic compound effective at low doses for negative symptoms and at high doses for positive symptoms of schizophrenia. The CNS profile of multiple doses of a low dosage regimen of amisulpride (50 mg once daily for 4 days) was assessed in a randomised, double-blind, 3-way crossover, placebo-controlled study carried out in 12 young sleep-deprived (for 36 h) subjects, using EEG and various measures of psychomotor and cognitive functions. Caffeine slow release (600 mg) was used as a positive reference. Multiple doses of 50 mg amisulpride once daily were devoid of any detrimental effects on EEG and psychomotor performance and cognitive function after total sleep deprivation. In addition, 50mg amisulpride partially antagonized the deleterious effects of sleep deprivation on EEG and subjective sedation as shown by trends, and a significant increase in EEG relative beta power and a decrease in subjective sedation. These effects were more pronounced at the end of sleep deprivation, suggesting possible alerting effects of amisulpride at this dose level. Caffeine significantly antagonized the detrimental effects of sleep deprivation on vigilance (increase in EEG beta waves, speed of reaction, sustained attention and reduction in subjective sedation). In conclusion, the present results demonstrate that 50 mg amisulpride is devoid of detrimental effects on EEG, psychomotor and cognitive performance after sleep deprivation, a situation well-known to amplify such effects if they exist. Moreover, some data suggest possible alerting effects of this low dosage regimen of amisulpride. PMID- 10520733 TI - Prediction of methotrexate elimination after high dose infusion in children with acute lymphoblastic leukaemia using a population pharmacokinetic approach. AB - High-dose methotrexate (HD-MTX) with leucovorin rescue is a component of therapy in children with acute lymphoblastic leukaemia. Since MTX toxicity is related to drug exposure, a monitoring of serum MTX concentrations at H24, H48, H72 and until the concentration is less than 0.2 micromol/L is commonly performed. However, a number of patients may reach concentrations of less than 0.2 micromol/L long before the next sampling is scheduled. The aim of our study was to develop a Bayesian method predicting the time at which MTX concentration reaches 0.2 micromol/L in order to decrease the number of samples drawn and to allow for a more rapid patient discharge. Methotrexate population parameters were estimated from a retrospective analysis of 60 infusions in 23 children and MTX concentrations were predicted from an independent set of 20 courses in 14 children with a Bayesian approach using either one (H48) or two (H24 and H48) samples. The following population parameters were obtained using a two compartment model: CL = 3.51 L/h (inter-individual variability: 66%), Vd = 8.67 L (58%), k12 = 0.0044 h(-1)(105%), k21 = 0.039 h(-1)(25%). Clearance and Vd were found to increase with weight and age respectively. Both sampling schedules tested for the Bayesian estimation enabled accurate prediction of concentrations and provided satisfactory precision despite a small bias. When considering the ability to predict the time at which the threshold was reached, the one-sample (H48) schedule gave the best results. We conclude that a sampling schedule involving only one sample and Bayesian parameter estimation may be able to predict the delay necessary to reach 0.2 micromol/L in each individual. PMID- 10520734 TI - Report from the second European congress of pharmacology. PMID- 10520735 TI - A new signal peptidase gene from Streptomyces lividans TK21. AB - Using synthetic oligonucleotides derived from known signal peptidase genes and a multicopy plasmid as a vector, a signal peptidase gene (sipZ) from Streptomyces lividansTK21 has been cloned. The primary structure of the gene has been determined and the amino acid composition of the SipZ protein inferred. SipZ is 258 aa long and showed homology to other type I signal peptidases, containing like them an N-terminal transmembrane anchor. Alignment of SipZ with other known SPases allowed the identification of a conserved sequence of amino acids specific for Gram-positive bacteria. PMID- 10520736 TI - The Sip(Sli) gene of Streptomyces lividans TK24 specifies an unusual signal peptidase with a putative C-terminal transmembrane anchor. AB - Type I signal peptidases (SPases) are a widespread family of enzymes which remove signal peptides from proteins translocated across cellular membranes. Here, we report the first isolation of a gene coding for type I signal peptidase of Streptomyces, denoted Sip(Sli). The sip(sli) gene specifies a protein of 291 amino acids. Thus Sip(Sli) is much larger (approximately 100 amino acids) than other known SPases of Gram-positive bacteria and resembles SPases of Gram negative bacteria, showing the highest degree of similarity to an SPase of the cyanobacterium Phormidium laminosum. Sip(Sli) contains conserved serine and lysine residues, which are believed to be required for the catalytic activity. Similar to other known SPases from Gram-positive bacteria, Sip(Sli) seems to have only one N-terminal transmembrane anchor. In addition, Sip(Sli) seems to contain a second transmembrane anchor at the C-terminus, which is an unusual feature for type I signal peptidases. PMID- 10520737 TI - Sequencing of 42kb of the APO E-C2 gene cluster reveals a new gene: PEREC1. AB - Through the sequencing of a 42kb cosmid clone we describe a new gene, designated PEREC1, located approximately 1.5kb centromeric of the human apolipoprotein (APO) E-C2 cluster. The combination of dotplot analysis, predicted coding potential and interrogation of the Expressed Sequence Tag (EST) database determined the genomic organisation of PEREC1. Sequence alignment with multiple overlapping ESTs confirmed the predicted splice sites. The predicted cDNA and amino acid sequences of PEREC1 have extensive similarity to the Caenorhabditis elegans protein, C18E9.6. Conserved structural and functional motifs have been defined by combining nucleotide and amino acid analyses to identify third base degeneracy and therefore selection at the protein level. The Poliovirus Receptor Related Protein2 gene (PRR2), previously mapped to chromosome 19q13.2 by Fluorescent In Situ Hybridisation, has also been located approximately 17kb centromeric of APO E. PMID- 10520738 TI - Molecular cloning of a leucine zipper motif-containing novel cDNA specifically expressed in adult mouse testis. AB - A novel cDNA clone was isolated from a mouse testis cDNA library. This cDNA is 2,020 nucleotides in size and predicted to encode 527 amino acid residues with a molecular weight of 59.9 kDa. Protein sequence motif analysis revealed that the predicted protein contained one leucine zipper motif in its N-terminal portion and two basic domains in its C-terminal portion. Northern blot analysis of adult ICR mouse organs using the cDNA as probe demonstrated that an approximately 2.0 kb transcript was specifically expressed in testis. We therefore termed this novel cDNA tsec-2, testis-specifically expressed cDNAs-2. Results of Southern blot analysis suggested that the tsec-2 gene may exist as a single copy in the mouse genome. PMID- 10520739 TI - Isolation and identification by sequence homology of a second putative C5-DNA methyltransferase gene from Ascobolus immersus. AB - I report the cloning of a new Ascobolus gene (masc 2) that potentially encodes a C5-DNA-methyltransferase. The putative protein exhibits the two domains characteristic of eukaryotic maintenance DNA-methyltransferase: a large N terminal domain and a C-terminal domain containing all ten catalytic motifs arranged in the canonical order. A new type of eukaryotic DNA-methylase gene (masc 1) has been recently found in Ascobolus. Masc1 is essential for the de novo methylation and dispensable for methylation maintenance. The masc2 gene could encode the methylase involved in this maintenance. PMID- 10520740 TI - Cloning of a cDNA for xDOR2, a novel TR2-related nuclear orphan receptor, expressed during neurulation in Xenopus laevis embryos. AB - We isolated from neurulating Xenopus laevis (X. laevis) embryos a cDNA encoding a novel nuclear orphan receptor, Developmental Orphan Receptor 2 (xDOR2), closely related to Ambystoma mexicanum DORI (aDOR1) and to Testicular Receptor-2 (TR2) orphan receptor family members. The xDOR2 cDNA sequence which is truncated both at its 5' and 3' ends predicts a protein sequence of 542 amino acids. While the DNA-binding domain of xDOR2 shares 91%, and 92%, identity with those of aDOR1 and the TR2s, respectively, considerable divergence is observed at both extremities of the peptides. At the N-terminus, xDOR2 is 66% identical to aDOR1 and 64% to the TR2s. At the C-terminus, xDOR2 which is longer by 126 amino acids compared to aDOR1, shares 62%, identity with aDOR1 and appears to contain a complete ligand binding domain. The DOR receptors appear to form a subtype distinct from the TR2s, and may play a role in the development and differentiation of neural tissues. PMID- 10520741 TI - Cloning of a cDNA encoding ovine keratinocyte growth factor. AB - A cDNA encoding the ovine keratinocyte growth factor (KGF) has been cloned. A 622 bp cDNA, containing the entire protein coding region, was amplified from an ovine adenocarcinoma fibroblast cell line. The cDNA was found to share 95% nucleotide identity with dog KGF, 94% identity with human KGF, 90% identity with mouse KGF and 88% identity with rat KGF. The predicted ovine amino-acid sequence shares 98.5, 98, 96 and 94% identity, respectively, with the corresponding dog, human, mouse and rat proteins. The KGF gene is present as a single copy in the ovine genome. PMID- 10520742 TI - Diversity of the limited-host range iaaH gene of Agrobacterium vitis strain Ag162. AB - The indole-3-acetamide hydrolase gene (iaaH) of the limited-host range strain AG162, a biotype III strain of Agrobacterium tumefaciens has been the subject of several studies and reviews, but its primary structure has not been previously reported. In the course of our own work we found that this gene hybridizes only weakly to a nucleic acid probe corresponding to the iaaH gene from a biotype I strain of A. tumefaciens. Analysis of the primary structure of the Ag162 iaaH gene revealed that it is diverse from biotype I iaaH genes and, surprisingly, also from the iaaH genes of previously characterized biotype III Agrobacterium strains. PMID- 10520743 TI - Molecular cloning and characterization of a mouse gene with homology to the Duffy antigen receptor for chemokines. AB - The Duffy antigen receptor for chemokines (DARC) is a receptor for both CXC and CC chemokines. We have cloned a mouse gene with a predicted amino acid sequence homology of approximately 63% to human DARC and localized this gene to mouse chromosome 1 between the Xmv41 and D1Mit166 loci. We further demonstrated that, like the human gene, the mouse gene exhibits a single intron of 462 bp which interrupts the open reading frame between the codons for the seventh and eighth amino acid residues. Northern blot analyses revealed that putative mDARC mRNA is highly expressed in adult mouse spleen and skeletal muscle and in whole embryos between embryonic days 8.5 to 12. Northern blot analysis of hemangiosarcomas which develop spontaneously in the spleen of the Eker rat reveal expression of mRNAs which hybridize with both DARC and CXCR2 probes, suggesting a potential role of these receptors in the angiogenesis associated with tumor formation. PMID- 10520744 TI - Assembly of a complete zebrafish mitochondrial 16S rRNA gene from overlapping expressed sequence tags. AB - A complete zebrafish mitochondrial 16S ribosomal RNA gene was assembled from our existing zebrafish EST clones by aligning them with the carp mitochondria 16S rRNA sequence. The overall homology between our assembled zebrafish mt 16S rRNA and the carp mt 16S rRNA is 83.5%. As the number of zebrafish ESTs grows, the assembly of more full-length cDNA sequences from overlapping EST data could be expected. PMID- 10520745 TI - The division and cell wall gene cluster of Enterococcus hirae S185. AB - A chromosomal 10355-bp segment of Enterococcus hirae S185 contains nine orfs which occur in the same order as the MraW-, FtsL-, PBP3-, MraY-, MurD-, MurG-, FtsQ-, FtsA- and FtsZ-encoding genes of the division and cell wall clusters of Escherichia coli and Bacillus subtilis. The E. hirae DNA segment lacks the genes which in E. coli encode the ligases Ddl, MurC, MurE and MurF and the integral membrane protein FtsW. The encoded E. hirae and E. coli proteins share 25% to 50% identity except FtsL and FtsQ (approximately = 14% identity). PMID- 10520746 TI - Computer sequence analysis of human highly conserved zinc finger modules. AB - We defined a sub-family of zinc finger proteins by computer analyses and comparisons of five new finger domains against protein databases. This subclass of the cysteine-cysteine/histidine-histidine motif shows additional well conserved amino acid patterns and belongs to the human kox and gli-Kruppel gene family, sharing also the same stretches of regulatory zinc finger-containing proteins of mouse and Xenopus. We particularly describe ZF6 cDNA which contains the most interesting sequence, encoding a putative multi-domain regulatory protein. PMID- 10520748 TI - Nucleotide sequence of the GLABROUS1 gene of Arabidopsis thaliana ecotype Columbia. AB - The GLABROUS1 (GL1) gene from Arabidopsis thaliana ecotype Columbia was isolated and sequenced. Comparison of the nucleotide sequence with that of the gl1-2allele revealed that two sites were changed in gl1-2. One was a 14-bp deletion in exon 3, which seemed most likely to be the cause of the mutated phenotype. The other was a change from TC to CT in the 5' untranslated region. Since the 5' upstream sequence required for the GL1 gene expression has not been clearly specified, one cannot rule out the possibility that this change may contribute to the phenotype of gl1-2. A comparison was also made with the GL1 gene of ecotype Wassilewskija and revealed some lack of sequence conservation. The changes observed may have little influence on the function or the expression of the GL1 gene product. PMID- 10520747 TI - Isolation of two human cDNAs, HLP3 and HLP4, homologous to the neuron-specific calcium-binding protein genes. AB - We have cloned and sequenced two types of human cDNA, HLP3 and HLP4, encoding a protein of 191 amino acid residues with four EF-hand calcium-binding motifs. The HLP3 and HLP4 proteins are homologous to the neuron-specific calcium-binding proteins and are likely human counterparts of the neural visinin-like protein(NVP) 1 and NVP2 identified in the rat brain (Kajimoto et al. 1993), displaying 98% and 99% amino acid identities with these sequences, respectively. The human HLP3 and 4 mRNAs are detected only in the brain and found at high amount in the cerebral cortex and cerebellum by Northern blot analysis. PMID- 10520749 TI - The tdcE gene in Escherichia coli strain W3110 is separated from the rest of the tdc operon by insertion of IS5 elements. AB - Unlike other Escherichia coli K-12 strains, W3110 contains multiple copies of the insertion sequence IS5. Some of these IS5 elements have been involved in tandem duplication of a portion of the chromosome which includes, amonst others, the tdcABC-DEFG operon genes. The nucleotide sequence and insertion site of one of these elements, IS5P, was determined. It was shown that IS5P has inserted within the coding sequence of the tdcA gene and is flanked, not by the remaining portion of the tdcA gene, but by the extreme 3' end of the tdcD gene. In other E. coli K 12 strains the tdcD gene and three other genes, tdcE, tdcF and tdcG, all form part of the tdc operon. Our results demonstrate that during the duplication event the tdcABCgenes have been amplified and separated from the remaining genes tdcE, tdcF and tdcG of the operon, which are each present in single copy. PMID- 10520750 TI - Analysis of a 69-kb contiguous genomic sequence at a putative tumor suppressor gene locus on human chromosome 6q27. AB - Multiple neoplasias including B-cell non-Hodgkin's lymphoma, breast carcinoma, and ovarian carcinoma, have been associated with frequent deletions of the distal region on the long arm of human chromosome 6, suggesting the presence of one or more tumor suppressor gene(s) at this locus. Loss of heterozygosity analysis of breast and ovarian tumors has further restricted the minimal region of loss within 6q27. To further characterize this genomic region for gene content including putative tumor suppressor genes as well as other elements that may contribute to tumorigenesis, a 68940-bp contiguous sequence, encompassing markers D6S193 and D6S297, was generated by random shotgun sequencing of a cosmid, P1, and PAC contig. In addition, exon trapping was performed utilizing a subset of these clones. Sixteen trapped exons, ranging in size from 44 to 399 bp, span this approximately 69-kb region. Many other putative exons have been identified computationally. Further analysis has identified 13 potential promoters and 13 putative polyadenylation sites in the region. Northern analysis identified a transcript mapping within this interval that is expressed in ovarian, breast, and lymphoid-derived tumor cell lines. Consideration of these data, together with the demonstration of several regions of high CpG content, suggests the possibility of several genes at this locus. PMID- 10520751 TI - Cloning and characterization of a family of cDNAs from human histiocyte macrophage cells encoding an arginine-rich basic protein related to the 70 kD U1 snRNP splicing factor. AB - This paper describes the cloning and characterization of five cDNA members of a novel family of mRNAs, termed hm-1, isolated from human U937 macrophage cells. Two family members (clones 46 and 11) show complete mRNA features [including ribosome binding sites (RBS), polyadenylation signals, and poly(A) tails], and encode the same protein (designated HM-1), but differ substantially in their 5' untranslated regions. The three other cDNAs (clones 20, 60, and 38) appear to represent partial cDNAs. The protein sequences deduced from the five hm-1 cDNAs are identical (some truncated), except for one Trp --> Cys substitution. Full length HM-1 is 246 amino acids long, has a predicted MW of 29431, is rich in arginine residues, has a pI of 10.25, and a mean hydrophobicity index of -1.23. HM-1 contains no obvious hydrophobic N-terminal cleavable signal sequence, and no potential N-glycosylation sites, but does contain three highly conserved motifs present in U1-70K splicing factors, and contains numerous C-terminal Arg/Asp and Arg/Glu dipeptides characteristic of "RD" family members that function as regulators of mRNA splicing. Northern hybridizations indicate that hm-1 is a family of mRNAs differentially expressed in a variety of human tissues. PMID- 10520752 TI - Isolation and structural and genetic analysis of the mouse enkephalin gene and its d(AC/TG)n repeats. AB - Enkephalins, the endogenous opioids, mediate a wide variety of intercellular communications through ontogeny and their involvement has been suggested in drug addiction and alcohol abuse as well as in various neuropsychiatric disorders. In order to generate a genetic model, we have isolated the mouse enkephalin (mENK) gene, analyzed its regulatory region and compared its structure to the well characterized rat ENK (rENK) gene. We analyzed 2600 bp and found 3 highly homologous regions: The highest level (98%) of positional and sequence homology between mice and rats was in the TATA/proximal regulatory region. This region contains all the inducible regulatory elements (enkCRE1, NF1, AP-2, NFkappaB, etc.) and also an octamer-like element at -543 bp. This high homology is interrupted in both mice and rats by the typically polymorphic d(AC/TG)n and d(TC/GA)n dinucleotide repeats positioned between nucleotides -670 and -950. The position and orientation of these repetitive elements differ substantially in the two species. Genomic PCR analysis of the d(AC/TG)n repeat in various mouse strains, including aberrant behavioral or neurological phenotypes, showed lack of polymorphism at this repeat. The positional and sequence homologies between the rat and the mouse ENK genes decrease in more upstream regions due to the presence of nonhomologues repetititve DNA sequences. PMID- 10520753 TI - Structure of the gene coding for calcineurin B (PPP3R1) and mapping to D2S358 D2S1778 (chromosomal region 2p15). AB - Calcineurin is a protein phosphatase with an important role in signal transduction; its calcium-binding regulatory subunit, calcineurin B, is widely present in the brain and is coded by the PPP3R1 gene which was mapped recently to human chromosome 2. Calcineurin has long been considered a candidate for psychiatric and/or monogenic brain disorders. The present study reports the intron-exon structure of the PPP3R1 gene with the proximal intronic sequences, its genetic mapping to D25358-D251778 on chromosome 2p15, and its exclusion in a genetic disorder mapped proximal to this locus. PMID- 10520754 TI - Identification of an elicitin gene cluster in Phytophthora cinnamomi. AB - Elicitins are a group of highly conserved proteins secreted by species of Phytophthora and a species of the related genus Pythium, Pythium vexans. Some of these proteins act as inducers of the necrotic hypersensitive-like response and the associated systemic acquired resistance phenomenon, in some species. We cloned and characterised the cinnamomin-beta and -alpha genes and two related elicitin genes from Phytophthora cinnamomi. These four open reading frames (ORFs) are clustered in tandem pairs. Two out of these four genes present homologies with the basic and acidic elicitin groups; but the two others encode, if expressed, elicitin isoforms exhibiting homologies with the class II of highly acidic elicitins. PMID- 10520756 TI - Structure-based drug design approaches for predicting binding affinities of HIV1 protease inhibitors. AB - Computational assessment of the binding affinity of enzyme inhibitors prior to synthesis is an important component of computer-assisted drug design (CADD) paradigms. The free energy perturbation (FEP) methodology is the most accurate means of estimating relative binding affinities between two inhibitors. However, due to its complexity and computation-intensive nature, practical applications are restricted to analysis of structurally-related inhibitors. Accordingly, there is a need for methods that enable rapid assessment of large number of structurally-unrelated molecules in a suitably accurate manner. In this review, the FEP method is compared with regression-based methods that employ multivariate models to assess the advantages of each in the estimation of relative binding affinities of inhibitors to an enzyme. Semiquantitative predictions of relative binding free energies of human immunodeficiency virus 1 (HIV1) protease inhibitors are also presented and compared with the corresponding FEP results. The results indicate that the regression-based methods and the FEP method are useful in the semi-quantitative and quantitative assessment of relative binding affinities of enzyme inhibitors, respectively, prior to synthesis. PMID- 10520755 TI - Nucleotide sequence of a marsupial interleukin-10 cDNA from the Australian brushtail possum (Trichosurus vulpecula). AB - A cDNA encoding a marsupial interleukin-10 (IL-10), was isolated from Australian brushtail possum alveolar macrophages. The cDNA of 1604 bp had an open reading frame of 522 bp coding for a protein of 174 amino acids. Its deduced amino acid sequence had an identity of 60% with cat, 58% with pig, 56% with human and cow, 52% with mouse and 53% with rat IL-10. The expression of IL-10 was up-regulated in both LPS-stimulated and Mycobacterium bovis-infected possum alveolar macrophages. PMID- 10520757 TI - Coagulation factor Xa: the prothrombinase complex as an emerging therapeutic target for small molecule inhibitors. PMID- 10520758 TI - Analysis of the kinetics of reversible enzyme inhibition by a general algebraic method. Application to multisite inhibition of the phosphoglycerate kinase from Trypanosoma brucei. AB - The action of an inhibitor on a stationary enzyme reaction is described by a simple equation, which reflects how the progressive binding of inhibitor molecules influences the existence and the productivity of the enzyme forms. This allows deduction of the structure of the enzyme system from the experimental results, using new type of plots (1/[I], 1/[I](a)v) where a = 0,1,2,... in complement to the usual graphs. A reaction scheme is thereby logically built. This method may be used without any theoretical calculation. It is valid whatever the inhibitor, when the association reactions of the substrates and the inhibitor to the enzyme are in rapid equilibrium, and with dead end inhibitors, more generally for steady state enzyme reactions. This method may be adapted to enzyme activation. An original inhibition mode is described with particular bifunctional molecules: cooperative binding of the inhibitor to the enzyme, outside the active site, by direct mutual interaction of two inhibitor molecules, and locking of the conformational changes that normally precede the release of the products. PMID- 10520759 TI - The antifungal activity of 2,2'-diamino-4,4'-dithiazole derivatives is due to the possible inhibition of lanosterol-14-alpha-demethylase. AB - Aryl/alkyl sulfonylamido-, arylsulfenylamido-, arylcarboxamido- and ureido/thioureido/guanidino derivatives of 2,2'-diamino-4,4'-dithiazole were prepared by reaction of the title compound with sulfonyl/sulfenyl halides, sulfonic acid anhydrides, acyl chlorides, tosyl isocyanate, aryl/allyl isocyanates or isothiocyanates. Mono- as well as bis-derivatized compounds have been obtained. Several of the newly synthesized compounds act as effective antifungal agents against Aspergillus and Candida spp., some of them showed activities comparable to ketoconazole (with minimum inhibitory concentrations in the range of 0.2-1.8 microg/mL) but possessed lower activity as compared to itraconazole. Greatest activity was detected against A. niger, and least activity against C. albicans. The mechanism of action of these compounds probably involves inhibition of ergosterol biosynthesis, and interaction with lanosterol-14-alpha demethylase (CYP51A1), since reduced amounts of ergosterol were found by means of HPLC in cultures of the sensitive strain A. niger treated with some of these inhibitors. Thus, the compounds reported here and the azole antifungal derivatives might possess a similar mechanism of action at molecular level. PMID- 10520760 TI - Macrostatin, a novel macromolecular inhibitor of topoisomerases produced by Streptomyces avermitilis no. C-127. AB - A novel inhibitor of topoisomerases designated as Macrostatin has been isolated from the culture filtrate of Streptomyces avermitilis strain No. C-127 and purified by successive chromatography on Dowex, activated charcoal, gel filtration and cellulose. It is an acidic macromolecule having 45 kD molecular weight as determined by gel filtration. Macrostatin inhibited topoisomerase I and II in a noncompetitive manner with Ki = 3.7 and 1.3 nM respectively. Macrostatin differed from well-known inhibitors of topoisomerase such as camptothecin, etoposide and doxorubicin which induce topoisomerase-mediated DNA cleavage by stabilizing the cleavable complex or intercalation into DNA strands. Macrostatin had neither ability to stabilize the cleavable complex nor ability to intercalate into DNA strands. It was suggested that Macrostatin inhibits topoisomerase by a direct action on the enzyme. PMID- 10520762 TI - Interventional Strategies in the Treatment of Atrial Fibrillation. Proceedings of an international symposium. Dresden, Germany, March 19-20, 1999. PMID- 10520761 TI - Isolation of antioxidant principle from Azadirachta seed kernels: determination of its role on plant lipoxygenases. AB - An antioxidant principle was isolated from Azadirachta indica seed using high pressure liquid chromatography with a hydrophobic reverse-phase chromatography column. The eluted molecule had lambdamax at 224 and 272 nm and was a potent inhibitor of plant lipoxygenases. In in vivo studies of horsegram during germination, low levels of lipoxygenase activity and lipid peroxides were found upon treatment with the Azadirachta extract. The antioxidant property of Azadirachta indica has not been previously reported. PMID- 10520763 TI - Pharmacological therapy of atrial fibrillation. AB - Pharmacological treatment of atrial fibrillation aims at reducing symptoms and possible complications of this frequently encountered arrhythmia. Prevention of embolic events by means of oral vitamin K antagonists is therefore mandatory in most patients presenting with atrial fibrillation. Whereas in paroxysmal atrial fibrillation reduction in frequency and duration of episodes is the treatment goal, symptomatic relief of persistent atrial fibrillation can be achieved by two treatment strategies: 1) restoration of sinus rhythm, referred to as rhythm control, and 2) limitation of the average and maximal ventricular heart rate, referred to as rate control. This review provides data from relevant clinical studies addressing the effectiveness and limitations of currently available drug therapy according to the two treatment approaches. PMID- 10520764 TI - Pacing therapy for atrial fibrillation. PMID- 10520765 TI - Intra-atrial defibrillation in humans. AB - The frequency of atrial fibrillation (AF) and its economic impact on the health care system ensure that AF will be a focus for future research. At present, the chronic use of an atrioverter (IAD) is a major challenge in the nonpharmacological treatment of AF. Studies in animals as well as in humans have demonstrated the feasibility and safety of internal atrial defibrillation. The major issues that have to be addressed are the pain perception and the potential risk of inducing life-threatening ventricular arrhythmias during delivery of low energy atrial shocks. A first step to this novel approach is a physician activated device. At the very beginning, the IAD should be restricted to highly selected patients with drug refractory, poorly tolerated, recurrent AF episodes. The extenuation of this therapy to wider subsets of patients should be dependent on the initial results with regard to clinical efficacy and safety as well as patient tolerance. Finally, cost-effectiveness as well as quality of life studies are needed to demonstrate the benefit of this specific therapy among other therapeutic strategies available for the management of AF. A new arrhythmia management system that combines both detection and treatment in the atrium as well as in the ventricle may represent an important milestone and a significant improvement in the management of patients with both supraventricular and ventricular tachyarrhythmias. PMID- 10520766 TI - An anatomic approach to prevention of atrial fibrillation: pulmonary vein isolation with through-the-balloon ultrasound ablation (TTB-USA). AB - Our current understanding is that atrial fibrillation (AF) is initiated most often by a focal trigger from the orifice of or within one of the pulmonary veins. Though mapping and ablation of these triggers appears to be curative in most patients with paroxysmal AF, there are a number of limitations to ablating focal triggers via mapping and ablating the earliest site of activation with a "point" radiofrequency lesion. One way to circumvent thesen limitations is an anatomically-guided ablative approach. By electrically isolating one or more pulmonary veins from the left atrium with a circumferential lesion, firing from within those veins would be unable to reach the body of the atrium, and thus could not trigger atrial fibrillation. We have developed a novel over-the-wire catheter design which integrates a cylindrical ultrasound transducer within a saline filled balloon, termed TTB-USA (through-the-balloon ultrasound ablation) in order to produce narrow circumferential zones of hyperthermic tissue death at the pulmonary vein ostia. Animal studies show great promise, and clinical trials will begin soon. PMID- 10520767 TI - Catheter ablation of pulmonary vein foci for atrial fibrillation: PV foci ablation for atrial fibrillation. AB - While experimental and human mapping studies have documented multiple wavelet reentry as the electrophysiological mechanism maintaining atrial fibrillation, recent evidence shows that nearly all paroxysms of atrial fibrillation are initiated by trains of rapid discharges from the pulmonary veins. Radiofrequency catheter ablation targeting these initiating triggers has resulted in an overall 69% freedom from atrial fibrillation at a follow-up of 8 +/- 4 months in a population of 110 patients with paroxysmal atrial fibrillation. Six of the targeted pulmonary veins (4%) developed pulmonary vein stenosis; none requiring specific treatment. Catheter ablation of pulmonary vein foci initiating atrial fibrillation is therefore an effective curative modality for paroxysmal atrial fibrillation. PMID- 10520768 TI - Catheter ablation of atrial flutter. AB - Typical atrial flutter in humans is the consequence of a stable macro-reentrant circuit produced by the unique right atrial architecture providing anatomic barriers and functional blocks to conduction. Mapping studies have indicated that the so-called isthmus between the inferior aspect of the tricuspid annulus and the ostium of the inferior caval vein is a critical zone for maintenance of atrial flutter. An anatomically guided approach with placement of a transmural and contiguous lesion line throughout the isthmus has established as curative treatment of typical atrial flutter. Electrophysical criteria indicating complete bidirectional isthmus conduction block after ablation proved to be superior with respect to redurrences of atrial flutter compared with the noninducibility criterion. The gold standard for prove of complete conduction block is the recording of double potentials along the entire isthmus ablation line. Recently, it proved possible to reduce the period of fluoroscopy during isthmus ablation by using electro-anatomical mapping. PMID- 10520769 TI - Evolution of the maze III procedure: are modifications necessary? AB - The maze III procedure has been the culmination of multiple surgical approaches for supraventricular arrhythmias. Its success in curing atrial fibrillation has generated multiple modifications which constitute attempts to simplify the operation, particularly with associated mitral or multiple valvular pathology. Our preference in these patients, however, has been to employ the original maze III procedure without modification. This review tracks the development of the maze III procedure and its modifications and compares the early outcomes in patients requiring the maze procedure combined with mitral and additional valvular procedures. PMID- 10520770 TI - Antiarrhythmic surgery for treatment of atrial fibrillation--new concepts. AB - Curative treatment of atrial fibrillation is one of the main challenges of todays electrophysiology. The ideal treatment strategy should be effective, safe, and easy to apply to allow a widespread use. In addition, curative treatment should not only aim on the restoration of sinus rhythm but also restore mechanical atrial function to improve hemodynamics thereby avoiding anticoagulation. With respect to percutaneous catheter ablation no treatment concept with proven efficiency to cure chronic permanent atrial fibrillation is currently available. Surgical techniques such as the Corridor operation and the left atrial isolation procedure have been shown to effectively store sinus rhythm but these procedures do not restore biatrial transport function. Cox's Maze procedure is highly effective, however, it is an extensive and very time consuming technique which precludes the widespread application of this operation. Thus, new intraoperative treatment concepts are currently under intense clinical investigation. Most new concepts aim on the application of contiguous radiofrequency-induced lesion lines in the atria. Some of the new treatment strategies are based on the replacement of the surgical incisions of the Maze procedure using inrtraoperative radiofrequency coagulation thereby preventing functional determined reentrant circuits. Other new concepts aim on the induction of contiguous atrial lesion lines to eliminate anatomical determined atrial reentrant circuits. The main advantage of these new concepts when compared to the Maze procedure is a significantly shorter treatment time of approximately 20 minutes. In addition, some treatment strategies can also be applied in conjunction with minimally invasive cardiac surgery. The initial results reported with application of new treatment concepts indicate that approximately 60-80% of patients operated on can be cured from atrial fibrillation. Randomised studies with these new treatment strategies are necessary to validate the results and to outline which treatment concept may prove superior to others. Based on the progress made, it can be expected that intraoperative ablation of chronic permanent atrial fibrillation will become an important curative treatment strategy. PMID- 10520771 TI - Electrosurgical treatment of atrial fibrillation with a new intraoperative radiofrequency ablation catheter. AB - BACKGROUND: To assess the efficacy of a new catheter for intraoperative radiofrequency ablation. METHODS: We operated 35 mitral patients with atrial fibrillation of which 27 had chronic atrial fibrillation with a mean duration of 8 +/- 6 years. Most patients were in functional class III or IV. All patients were operated under cardiopulmonary bypass using sternotomy in 29 patients, right thoracotomy in 5 and left thoracotomy in 1. RESULTS: Bilateral pulmonary vein isolation with radiofrequency catheter ablation was achieved in 7 +/- 4 minutes. There was no mortality or morbidity. Out of the 27 patients with one to three months follow-up 60% were out of atrial fibrillation and 48% had both atria contracting (scores 3 and 4). A longer time is required to assess the end result of these techniques, because the complete healing of the ablation lesions takes 3 to 6 months. CONCLUSIONS: We conclude that with appropriate tools and settings the use of intraoperative radiofrequency catheter ablation is fast, safe and effective. Its indications can be extended to other types of atrial fibrillation patients. PMID- 10520772 TI - The potential role of the cooled tip radiofrequency ablation catheter in the Cox Maze III procedure. PMID- 10520773 TI - Treatment of atrial fibrillation in open heart surgery--the potential role of microwave energy. AB - Treatment of atrial fibrillation by creating linear lesions in patients undergoing heart surgery (mitral valve replacement or coronary bypass grafting) receives increasing interest. Conventional rhythm surgery require tremendous operative effort and increases the risks of perioperative complications. For that reason various alternative methods for creating linear lesion are under investigation. This paper discusses several techniques for creation of transmural lesions in open heart surgery, e.g. ultra sound ablation, high current DC shocks, laser ablation, cryoablation, radio frequency ablation, cooled radio frequency ablation, and microwave ablation. PMID- 10520774 TI - Intraoperative microwave ablation for curative treatment of atrial fibrillation in open heart surgery--the MICRO-STAF and MICRO-PASS pilot trial. MICROwave Application in Surgical treatment of Atrial Fibrillation. MICROwave Application for the Treatment of Atrial Fibrillation in Bypass-Surgery. AB - Concomitant microwave ablation for curative treatment of atrial fibrillation (AF) was performed in 18 patients with history of chronic atrial fibrillation and indication for open heart surgery, 11 patients with mitral valve replacement and 7 patients with coronary artery bypass grafting. There were no perioperative complications. During the postoperative period most of the patients had intermittent AF, they received low dose Sotalol therapy and electric cardioversions. Up to now seven patients have reached follow-up day 90. One patient has persistent AF. Two patients had typical atrial flutter that was electrically converted to sinus rhythm (SR), isthmus ablation is planned. The other four patients have SR, one patient without cardioversions. These four patients show recovered atrial function with observed A-wave for transmitral flow. Under visual guidance the continuous atrial lesion lines could be induced effectively and safely by the intraoperative device Lynx. PMID- 10520775 TI - The role of laser in cardiac surgery. AB - With lasers tissue can be removed or coagulated. Laser coagulation has the same effect as radiofrequency or microwave applications. So far lasers are barely used in atrial fibrillation but it has been shown that a total block of the AV-node or a change of the PQ-time are possible with a percutaneous laser-catheter. Also the myocard can be coagulated if the laser radiation is delivered through the mitral valve without any damage to it. Short pulsed lasers produce shockwaves and thus mechanical injury to tissue. This mechanical damage to the myocard along with a controlled coagulation may lead to an interpretation of the effectiveness of TMLR. PMID- 10520776 TI - 50th anniversary historical article. A century of cardiac arrhythmias: in search of Jason's golden fleece. PMID- 10520777 TI - Recombinant hirudin (lepirudin) for the improvement of thrombolysis with streptokinase in patients with acute myocardial infarction: results of the HIT-4 trial. AB - OBJECTIVES: The purpose of this study was to compare recombinant hirudin and heparin as adjuncts to streptokinase thrombolysis in patients with acute myocardial infarction (AMI). BACKGROUND: Experimental studies and previous small clinical trials suggest that specific thrombin inhibition improves early patency rates and clinical outcome in patients treated with streptokinase. METHODS: In a randomized double-blind, multicenter trial, 1,208 patients with AMI < or =6 h were treated with aspirin and streptokinase and randomized to receive recombinant hirudin (lepirudin, i.v. bolus of 0.2 mg/kg, followed by subcutaneous (s.c.) injections of 0.5 mg/kg b.i.d. for 5 to 7 days) or heparin (i.v. placebo bolus, followed by s.c. injections of 12,500 IU b.i.d. for 5 to 7 days). A total of 447 patients were included in the angiographic substudy in which the primary end point, 90-min Thrombolysis in Myocardial Infarction (TIMI) flow grade 3 of the infarct-related artery, was evaluated, while the other two-thirds served as "safety group" in which only clinical end points were evaluated. As an additional efficacy parameter the ST-segment resolution at 90 and 180 min was measured in all patients. RESULTS: TIMI flow grade 3 was observed in 40.7% in the lepirudin and in 33.5% in the heparin group (p = 0.16), respectively. In the entire study population the proportion of patients with complete ST resolution at 90 min (28% vs. 22%, p = 0.05) and at 180 min (52% vs. 48%, p = 0.18) after start of therapy tended to be higher in the lepirudin group. There was no significant difference in the incidence of hemorrhagic stroke (0.2% vs. 0.3%) or total stroke (1.2% vs. 1.5%), reinfarction rate (4.6% vs. 5.1%) and total mortality rate (6.8% vs. 6.4%) at 30 days, as well as the combined end point of death, nonfatal stroke, nonfatal reinfarction, rescue-percutaneous transluminal coronary angioplasty and refractory angina (22.7 vs. 24.3%) were not statistically different between the two groups. CONCLUSIONS: Lepirudin as adjunct to thrombolysis with streptokinase did not significantly improve restoration of blood flow in the infarct vessel as assessed by angiography, but was associated with an accelerated ST resolution. There was no increase in the risk of major bleedings with lepirudin compared to heparin. PMID- 10520778 TI - Impaired coronary blood flow in nonculprit arteries in the setting of acute myocardial infarction. The TIMI Study Group. Thrombolysis in myocardial infarction. AB - OBJECTIVES AND BACKGROUND: While attention has focused on coronary blood flow in the culprit artery in acute myocardia infarction (MI), flow in the nonculprit artery has not been studied widely, in part because it has been assumed to be normal. We hypothesized that slower flow in culprit arteries, larger territories infarcted and hemodynamic perturbations may be associated with slow flow in nonculprit arteries. METHODS: The number of frames for dye to first reach distal landmarks (corrected TIMI [Thrombolysis in Acute Myocardial Infarction] frame count [CTFC]) were counted in 1,817 nonculprit arteries from the TIMI 4, 10A, 10B and 14 thrombolytic trials. RESULTS: Nonculprit artery flow was slowed to 30.9 +/ 15.0 frames at 90 min after thrombolytic administration, which is 45% slower than normal flow in the absence of acute MI (21 +/- 3.1, p < 0.0001). Patients with TIMI grade 3 flow in the culprit artery had faster nonculprit artery CTFCs than those patients with TIMI grades 0, 1 or 2 flow (29.1 +/- 13.7, n = 1,050 vs. 33.3 +/- 16.1, n = 752, p < 0.0001). The nonculprit artery CTFC improved between 60 and 90 min (3.3 +/- 17.9 frames, n = 432, p = 0.0001), and improvements were related to improved culprit artery flow (p = 0.0005). Correlates of slower nonculprit artery flow included a pulsatile flow pattern (i.e., systolic flow reversal) in the nonculprit artery (p < 0.0001) and in the culprit artery (p = 0.01), a left anterior descending artery culprit artery location (p < 0.0001), a decreased systolic blood pressure (p = 0.01), a decreased ventriculographic cardiac output (p = 0.02), a decreased double product (p = 0.0002), a greater percent diameter stenosis of the nonculprit artery (p = 0.01) and a greater percent of the culprit artery bed lying distal to the stenosis (p = 0.04). Adjunctive percutaneous transluminal coronary angioplasty (PTCA) of the culprit artery restored a culprit artery CTFC (30.4 +/- 22.2) that was similar to that in the nonculprit artery at 90 min (30.2 +/- 13.5), but both were slower than normal CTFCs (21 +/- 3.1, p < 0.0005 for both). If flow in the nonculprit artery was abnormal (CTFC > or = 28 frames) then the CTFC after PTCA in the culprit artery was 17% slower (p = 0.01). Patients who died had slower global CTFCs (mean CTFC for the three arteries) than patients who survived (46.8 +/- 21.3, n = 47 vs. 39.4 +/- 16.7, n = 1,055, p = 0.02). CONCLUSIONS: Acute MI slows flow globally, and slower global flow is associated with adverse outcomes. Relief of the culprit artery stenosis by PTCA restored culprit artery flow to that in the nonculprit artery, but both were 45% slower than normal flow. PMID- 10520779 TI - Angiotensin-converting enzyme inhibition reduces monocyte chemoattractant protein 1 and tissue factor levels in patients with myocardial infarction. AB - OBJECTIVES: We investigated the effects of enalapril therapy on plasma tissue factor (TF), tissue factor pathway inhibitor (TFPI) and monocyte chemoattractant protein-1 (MCP-1) levels in patients with acute myocardial infarction. BACKGROUND: Macrophages express TF in human coronary atherosclerotic plaques. Both TF and TFPI are major regulators of coagulation and thrombosis. Monocyte chemoattractant protein-1 is a monocyte and macrophage chemotactic and activating factor. METHODS: In a randomized, double-blind, placebo-controlled study beginning about two weeks after myocardial infarction, 16 patients received four weeks of placebo (placebo group) and another 16 patients received four weeks of enalapril 5 mg daily therapy (enalapril group). We performed blood sampling after administration of the doses. RESULTS: There were no significant differences in the serum angiotensin-converting enzyme (ACE) activity, plasma TF, free TFPI or MCP-1 levels before administration between the enalapril and placebo groups. In the enalapril group, ACE activity (IU/liter) (14.0 before, 5.2 on day 3, 5.8 on day 7, 6.3 on day 28), TF levels (pg/ml) (223, 203, 182, 178) and MCP-1 levels (pg/ml) (919, 789, 790, 803) significantly decreased by day 28. However, the free TFPI levels (ng/ml) (28.2, 26.5, 26.8, 28.4) did not change. These four variables were unchanged during the study period in the placebo group. CONCLUSIONS: This study demonstrated that administration of enalapril reduces the increased procoagulant activity in patients with myocardial infarction associated with inhibition of the activation and accumulation of macrophages and monocytes. PMID- 10520780 TI - Low-level exercise echocardiography detects contractile reserve and predicts reversible dysfunction after acute myocardial infarction: comparison with low dose dobutamine echocardiography. AB - OBJECTIVES: The aim of this study was to evaluate low-level exercise echocardiography (LLEE) in detecting contractile reserve and predicting functional improvement of akinetic myocardium early after acute myocardial infarction (AMI). BACKGROUND: Experimental and clinical studies have shown that low-dose dobutamine enhances contractile function of dyssynergic but viable myocardium in patients with recent AMI. We hypothesized that endogenous catecholamines produced during a LLEE test could serve as a myocardial stressor to elicit contractile reserve. METHODS: Fifty-two consecutive patients with first AMI and > or =2 akinetic segments in the infarct-related territory underwent 5 +/ 2 days after AMI low-dose dobutamine echocardiography (LDDE) (5, 10 and 15 microg/kg/min) and LLEE (25 W during 3 min on a supine bicycle, with continuous echocardiographic recording). Both tests were performed on the same day, in random order. Follow-up echocardiography was obtained one month later. Regional wall thickening was semi-quantitatively assessed using a 16-segment, 5-grade scale model. Contractile reserve was defined as improvement in wall thickening of > or =1 grade. RESULTS: Mean increase in heart rate during stress tests was 15 +/ 7 beats/min with LLEE and 13 +/- 6 beats/min with LDDE (p = NS). Contractile reserve was detected in 119 (55%) of 217 akinetic segments at LLEE and in 137 (63%) segments at LDDE. At follow-up study, functional improvement was identified in 139 (64%) segments. Sensitivity, specificity and positive and negative predictive values for predicting functional recovery were 81%, 92%, 95% and 73%, respectively, for LLEE, and 91%, 86%, 92% and 84%, respectively, for LDDE. Moreover, there was a good correlation between systolic wall thickening measured in the center of the dyssynergic area during stress tests and at follow-up study: r = 0.77, p < 0.001 with exercise testing and r = 0.73, p < 0.001 with dobutamine testing. CONCLUSIONS: Low-level exercise echocardiography provides a promising alternative to LDDE for identifying myocardial viability and predicting reversible dysfunction early after AMI. PMID- 10520781 TI - Mechanisms and clinical significance of transient atrioventricular block during dobutamine stress echocardiography. AB - OBJECTIVES: The purpose of this study was to investigate the possible mechanism and the clinical significance of transient atrioventricular block (AVB) during dobutamine stress echocardiography (DSE). BACKGROUND: Transient AVB occurs rarely during DSE; however, the mechanisms responsible for blocks are unclear. METHODS: A retrospective analysis of clinical, echocardiographic, catheterization, revascularization and head-up tilting test data was conducted in patients who developed transient AVB during DSE. RESULTS: A total of 302 patients with known or suspected coronary artery disease (CAD) underwent DSE before coronary angiography between November 1997 and August 1998. Transient AVB developed in 12 patients during the test. Mobitz I block was noted in six patients and Mobitz II block in the other six patients. Nine of these 12 patients were subsequently shown to have CAD and three had no significant coronary artery stenosis. Mobitz II block was observed only in patients with CAD, while Mobitz I block occurred in three patients with and three patients without CAD (p < 0.05). Eight of the nine patients with CAD underwent a successful coronary angioplasty with or without stenting and a repeat DSE revealed no recurrence of heart block except in one patient. Head-up tilting test in the 12 patients revealed a positive response in three of the nine patients with and all three patients without CAD. A negative head-up tilting test was likely to be observed in patients with, as compared with those without, CAD in this study population (p < 0.05). CONCLUSIONS: Transient AVB is not an infrequent manifestation during DSE. Both myocardial ischemia and neurally mediated vagal reflex may be responsible for this phenomenon. The development of Mobitz II block during DSE is indicative of the presence of CAD. A successful revascularization in patients with CAD who develop transient AVB may abolish this phenomenon. PMID- 10520782 TI - Improvement of myocardial blood flow to ischemic regions by angiotensin converting enzyme inhibition with quinaprilat IV: a study using [15O] water dobutamine stress positron emission tomography. AB - OBJECTIVES: This study was designed to analyze the effects of acute angiotensin converting enzyme (ACE) inhibition on myocardial blood flow (MBF) in control and ischemic regions. BACKGROUND: Although animal studies indicate an improvement of MBF to ischemic regions after ACE inhibition, this effect has not been conclusively demonstrated in patients with coronary artery disease. METHODS: Myocardial blood flow was analyzed in ischemic and nonischemic regions of 10 symptomatic patients with coronary artery disease using repetitive [15O] water positron emission tomography at rest and during maximal dobutamine stress before and after ACE inhibition with quinaprilat 10 mg i.v. To exclude the possibility that repetitive ischemia may cause an increase in MBF, eight patients underwent the same protocol without quinaprilat (placebo patients). RESULTS: Rate pressure product in control and quinaprilat patients was comparable. In placebo patients, repetitive dobutamine stress did not change MBF to ischemic regions (1.41 +/- 0.17 during the first stress vs. 1.39 +/- 0.19 ml/min/g during the second stress, p = 0.93). In contrast, MBF in ischemic regions increased significantly after acute ACE inhibition with quinaprilat during repetitive dobutamine stress (1.10 +/- 0.13 vs. 1.69 +/- 0.17 ml/min/g, p < 0.015). Dobutamine coronary reserve in ischemic regions remained unchanged in placebo patients (1.07 +/- 0.11 vs. 1.10 +/- 0.16, p = 0.92), but increased significantly after quinaprilat (0.97 +/- 0.10 vs. 1.44 +/- 0.14, p < 0.002). Total coronary resistance decreased after ACE inhibition (123 +/- 19 vs. 71 +/- 10 mm Hg x min x g/ml, p < 0.02). CONCLUSIONS: Angiotensin-converting enzyme inhibition by quinaprilat significantly improves MBF to ischemic regions in patients with coronary artery disease. PMID- 10520783 TI - Prediction of functional recovery of viable myocardium after delayed revascularization in postinfarction patients: accuracy of dobutamine stress echocardiography and influence of long-term vessel patency. AB - OBJECTIVES: We sought to evaluate dobutamine stress echocardiography (DSE) for predicting recovery of viable myocardium after revascularization with cineangiography as a gold standard for left ventricular (LV) function. We studied the influence of late vessel reocclusion on regional LV function. BACKGROUND: Dobutamine stress echocardiography is a well established evaluation method for myocardial viability assessment. In previous studies the reference method for assessing LV recovery was echocardiography, long-term vessel patency has not been systematically addressed. METHODS: Sixty-eight patients with a first acute myocardial infarction (AMI) and residual stenosis of the infarct related artery (IRA) underwent DSE (mean +/- standard deviation) 21 +/- 12 days after AMI to evaluate myocardial viability. Revascularization of the IRA was performed in 54 patients by angioplasty (n = 43) or bypass grafting (n = 11). Coronary angiography and LV cineangiography were repeated at four months to assess LV function and IRA patency. RESULTS: Sensitivity and specificity of DSE for predicting myocardial recovery after revascularization were 83% and 82%. In the case of late IRA patency, specificity increased to 95%, whereas sensitivity remained unchanged. In the 16 patients with myocardial viability and late IRA patency, echocardiographic wall motion score index decreased after revascularization from 1.83 +/- 0.15 to 1.36 +/- 0.17 (p = 0.0001), and left ventricular ejection fraction (LVEF) increased from 0.52 +/- 0.06 to 0.57 +/- 0.06 (p = 0.0004), whereas in five patients, reocclusion of the IRA prevented improvement of segmental or global LV function despite initially viable myocardium. CONCLUSIONS: Dobutamine stress echocardiography is reliable to predict recovery of viable myocardium after revascularization in postinfarction patients. Late reocclusion of the IRA may prevent LV recovery and influence the accuracy of DSE. PMID- 10520784 TI - Thrombin generation after the abrupt cessation of intravenous unfractionated heparin among patients with acute coronary syndromes: potential mechanisms for heightened prothrombotic potential. AB - OBJECTIVES: The purpose of this study was to determine the mechanistic basis for thrombin generation and increased prothrombotic potential after the abrupt cessation of intravenous (i.v.) unfractionated heparin among patients with acute coronary syndromes. BACKGROUND: A "rebound" increase in prothrombotic potential has been observed biochemically and clinically after the abrupt cessation of unfractionated heparin (UFH) among patients with acute coronary syndromes. Although the mechanism is unknown, tissue factor and the extrinsic coagulation cascade, both operative in atherosclerotic vascular disease and arterial thrombosis, are thought to be centrally involved. METHODS: In a single-center, pilot study, 30 patients with either unstable angina or non-ST segment elevation myocardial infarction who had received a continuous i.v. infusion of UFH for 48 h were randomly assigned to: 1) abrupt cessation, 2) i.v. weaning over 12 h or 3) subcutaneous weaning over 12 h. RESULTS: Thrombin generation (prothrombin fragment 1.2) was evident within 1 h of UFH cessation, increased progressively (by nearly two-fold) at 24 h (p = 0.002) and correlated inversely with tissue factor pathway inhibitor concentration (r = -0.61). Thrombin generation was greatest among patients randomized to abrupt cessation (1.6-fold increase at 24 h) and least in those with i.v. weaning. CONCLUSIONS: Thrombin generation after the abrupt cessation of UFH may represent a drug-induced impairment of physiologic vascular thromboresistance in response to locally generated tissue factor. A dosing strategy of abbreviated i.v. weaning attenuates but does not prevent heparin rebound among patients with acute coronary syndromes. PMID- 10520785 TI - Effects of adjunctive balloon angioplasty after intravascular ultrasound-guided optimal directional coronary atherectomy: the result of Adjunctive Balloon Angioplasty After Coronary Atherectomy Study (ABACAS). AB - OBJECTIVES: This study was conducted to evaluate: 1) the effect of adjunctive percutaneous transluminal coronary angioplasty (PTCA) after directional coronary atherectomy (DCA) compared with stand-alone DCA, and 2) the outcome of intravascular ultrasound (IVUS)-guided aggressive DCA. BACKGROUND: It has been shown that optimal angiographic results after coronary interventions are associated with a lower incidence ofrestenosis. Adjunctive PTCA after DCA improves the acute angiographic outcome; however, long-term benefits of adjunctive PTCA have not been established. METHODS: Out of 225 patients who underwent IVUS-guided DCA, angiographically optimal debulking was achieved in 214 patients, then theywere randomized to either no further treatment or to added PTCA. RESULTS: Postprocedural quantitative angiographic analysis demonstrated an improved minimum luminal diameter (2.88 +/- 0.48 vs. 2.6 +/- 0.51 mm; p = 0.006) and a less residual stenosis (10.8% vs.15%; p = 0.009) in the adjunctive PTCA group. Quantitative ultrasound analysis showed a larger minimum luminal diameter (3.26 +/- 0.48 vs. 3.04 +/- 0.5 mm; p < 0.001) and lower residual plaque mass in the adjunctive PTCA group (42.6% vs. 45.6%; p < 0.001). Despite the improved acute findings in the adjunctive PTCA group, six-month angiographic and clinical results were not different. The restenosis rate (adjunctive PTCA 23.6%, DCA alone 19.6%; p = ns) and target lesion revascularization rate (20.6% vs. 15.2%; p = ns) did not differ between the groups. CONCLUSIONS: With IVUS guidance, aggressive DCA can safely achieve optimal angiographic results with low residual plaque mass, and this was associated with a low restenosis rate. Although adjunctive PTCA after optimal DCA improved the acute quantitative coronary angiography and quantitative coronary ultrasonography outcomes, its benefit was not maintained at six months. PMID- 10520787 TI - Nuclear cardiology after angioplasty and stent placement: beyond sensitivity and specificity. PMID- 10520786 TI - Early recovery of coronary flow reserve after stent implantation as assessed by positron emission tomography. AB - OBJECTIVES: The aim of this study was to quantitatively evaluate myocardial flow reserve in patients early after coronary stent implantation using positron emission tomography. BACKGROUND: Delayed restoration of coronary flow reserve after percutaneous transluminal coronary angioplasty (PTCA) has been observed using a variety of techniques. Altered distal vasoregulation as well as residual stenosis have been considered possible explanations for this phenomenon. Although the implantation of stents may influence some of these mechanisms, little data are available characterizing coronary flow reserve early after stent placement. METHODS: In 14 patients 1.6 +/- 0.6 days after stenting, N-13-ammonia positron emission tomographic studies were performed at rest and during adenosine-induced vasodilation. Myocardial blood flow was quantified using a three-compartment model. Rest and stress flow data, as well as coronary flow reserve of stented vascular territories, were compared with that of remote areas. RESULTS: The stenosis decreased from 72.1 +/- 7.3% to 3.7 +/- 6.7% after stent implantation. Coronary flow in the stented areas did not differ significantly from that in remote areas either at rest (76.1 +/- 18.5 and 75.7 +/- 17.7 ml/min/100 g, respectively), or during maximal vasodilation (205.5 +/- 59.9 and 179.4 +/- 47.4 ml/min/100 g, respectively). In addition, there was no significant difference in the calculated values of coronary reserve of these two regions (2.74 +/- 0.64 and 2.43 +/- 0.55, respectively). CONCLUSIONS: The mechanical support of dilated arteries by a stent not only restores the macroscopic integrity of epicardial arteries, but also results, in contrast to conventional PTCA procedures, in early recovery of flow reserve. PMID- 10520788 TI - Tissue characteristics of restenosis after percutaneous transluminal coronary angioplasty in diabetic patients. AB - OBJECTIVES: The purposes of this study were to analyze coronary specimens from patients with diabetes mellitus (DM) and to compare them with specimens from patients without DM. BACKGROUND: Diabetes mellitus is associated with an increased incidence of restenosis after percutaneous transluminal coronary angioplasty (PTCA). Increased hypercellular smooth muscle cell proliferation with exaggerated intimal hyperplasia formation may be responsible for this predisposition. METHODS: Eighteen coronary atherectomy specimens with restenosis after PTCA from patients with DM were compared with 18 coronary atherectomy specimens with restenosis after PTCA from patients without DM. Total and segmental areas were quantified on trichrome-stained tissue of hypercellular tissue, collagen-rich sclerotic tissue, atheroma and thrombus. Demographic and angiographic data were similar in both groups. RESULTS: The percentage of total plaque area composed of hypercellular tissue was lower in restenotic specimens from patients with DM than in restenotic specimens from patients without DM (19 +/- 6% vs. 44 +/- 5%; p = 0.003). The percentage of collagen-rich sclerotic tissue area was larger in restenotic specimens from patients with DM than in restenotic specimens from patients without DM (77 +/- 9% vs. 53 +/- 4%; p = 0.004). The percentages of atheroma and thrombus were similar in both groups. CONCLUSIONS: Intimal hypercellular tissue content is reduced in restenotic tissue from patients with DM. Collagen-rich sclerotic content is increased in restenotic lesions from patients with DM. These results suggest an accelerated fibrotic rather than a proliferative response in diabetic lesions from patients with restenosis after PTCA. PMID- 10520789 TI - Final results of the STent versus directional coronary Atherectomy Randomized Trial (START) AB - OBJECTIVES: This study was designed to compare primary stenting with optimal directional coronary atherectomy (DCA). BACKGROUND: No previous prospective randomized trial comparing stenting and DCA has been performed. METHODS: One hundred and twenty-two lesions suitable for both Palmaz-Schatz stenting and DCA were randomly assigned to stent (62 lesions) or DCA (60 lesions) arm. Single or multiple stents were implanted with high-pressure dilation in the stent arm. Aggressive debulking using intravascular ultrasound (IVUS) was performed in the DCA arm. Serial quantitative angiography and IVUS were performed preprocedure, postprocedure and at six months. The primary end point was restenosis, defined as > or =50% diameter stenosis at six months. Clinical event rates at one year were also assessed. RESULTS: Baseline characteristics were similar. Procedural success was achieved in all lesions. Although the postprocedural lumen diameter was similar (2.79 vs. 2.90 mm, stent vs. DCA), the follow-up lumen diameter was significantly smaller (1.89 vs. 2.18 mm; p = 0.023) in the stent arm. The IVUS revealed that intimal proliferation was significantly larger in the stent arm than in the DCA arm (3.1 vs. 1.1 mm ; p < 0.0001), which accounted for the significantly smaller follow-up lumen area of the stent arm (5.3 vs. 7.0 mm2; p = 0.030). Restenosis was significantly lower (32.8% vs. 15.8%; p = 0.032), and target vessel failure at one year tended to be lower in the DCA arm (33.9% vs. 18.3%; p = 0.056). CONCLUSIONS: These results suggest that aggressive DCA may provide superior angiographic and clinical outcomes to primary stenting. PMID- 10520790 TI - When should we start randomized trials for new devices? PMID- 10520791 TI - Economic impact of GPIIB/IIIA blockade after high-risk angioplasty: results from the RESTORE trial. Randomized Efficacy Study of Tirofiban for Outcomes and Restenosis. AB - OBJECTIVE: This study was conducted to assess the impact of GPIIb/IIIa blockade with tirofiban on costs during the initial hospitalization and at 30 days among patients undergoing high-risk coronary angioplasty. BACKGROUND: GPIIb/IIIa blockers are a new class of compounds that have been shown in clinical studies to prevent complications after high-risk angioplasty. METHODS: The RESTORE trial was a multinational, blinded placebo-controlled study of 2,197 patients randomized to tirofiban or placebo following coronary angioplasty. This economic assessment was a prospective substudy of the RESTORE trial, and included 1,920 patients enrolled in the U.S. Costs were estimated for the U.S. cohort based on their utilization of healthcare resources and on costs measured directly in 820 U.S. patients at 30 sites. RESULTS: There was a 36% difference in the rate of the composite event of death, myocardial infarction (MI) and revascularization at two days between tirofiban and placebo (8% vs. 12%, p = 0.002). This difference was attributed to a reduction in nonfatal MI, repeat angioplasty, coronary surgery and stent placement. These clinical benefits followed a similar trend at 30 days, with a 16% reduction in the composite event (p = 0.10). In-hospital cost, including professional and study drug costs, was $12,145 +/- 5,882 with placebo versus $12,230 +/- 5,527 with tirofiban (p = 0.75). The 30-day cost was $12,402 +/- 6,147 with placebo versus $12,446 +/- 5,814 with tirofiban (p = 0.87). CONCLUSIONS: Tirofiban has been shown to decrease in-hospital and possibly 30-day events after high-risk angioplasty. The beneficial clinical effects of tirofiban in high-risk patients can be achieved at no increased cost. PMID- 10520792 TI - Periprocedural quantitative coronary angiography after Palmaz-Schatz stent implantation predicts the restenosis rate at six months: results of a meta analysis of the BElgian NEtherlands Stent study (BENESTENT) I, BENESTENT II Pilot, BENESTENT II and MUSIC trials. Multicenter Ultrasound Stent In Coronaries. AB - OBJECTIVES: We aimed to identify periprocedural quantitative coronary angiographic (QCA) variables that have predictive value on long-term angiographic results and to construct multivariate models using these variables for postprocedural prognosis. BACKGROUND: Coronary stent implantation has reduced the restenosis rate significantly as compared with balloon angioplasty in short de novo lesions in coronary arteries >3 mm in size. Although the postprocedural minimal luminal diameter (MLD) is known to have significant bearing on long-term angiographic results, no practically useful model exists for prediction of angiographic outcome based on the periprocedural QCA variables. METHODS: The QCA data from patients who underwent Palmaz-Schatz stent implantation for short (<15 mm) de novo lesions in coronary arteries >3 mm and completed six months of angiographic follow-up in the four prospective clinical trials (BENESTENT I, BENESTENT II pilot, BENESTENT II and MUSIC) were pooled. Multiple models were constructed using multivariate analysis. The Hosmer-Lemeshow goodness-of-fit test was used to identify the model of best fit, and this model was used to construct a reference chart for prediction of angiographic outcome on the basis of periprocedural QCA variables. RESULTS: Univariate analysis performed using QCA variables revealed that vessel size, MLD before and after the procedure, reference area before and after the procedure, minimal luminal cross-sectional area before and after the procedure, diameter stenosis after the procedure, area of plaque after the procedure and area stenosis after the procedure were significant predictors of angiographic outcome. Using multivariate analysis, the Hosmer-Lemeshow goodness-of-fit test showed that the model containing percent diameter stenosis after the procedure and vessel size best fit the data. A reference chart was then developed to calculate the expected restenosis rate. CONCLUSIONS: Restenosis rate after stent implantation for short lesions can be predicted using the variables percent diameter stenosis after the procedure and vessel size. This meta-analysis indicates that the concept of "the bigger the better" holds true for coronary stent implantation. Applicability of the model beyond short lesions should be tested. PMID- 10520793 TI - Coronary artery distensibility in diabetic patients with simultaneous measurements of luminal area and intracoronary pressure: evidence of impaired reactivity to nitroglycerin. AB - OBJECTIVES: This study investigated whether noninsulin dependent diabetes mellitus (NIDDM) adversely affects the elastic properties of the coronary arteries in patients with coronary artery disease (CAD) and NIDDM. BACKGROUND: Attenuated vascular smooth muscle dilation to exogenous donors of nitric oxide, such as nitroglycerin, has been observed with forearm blood flow studies in patients with NIDDM. METHODS: Twenty patients with CAD and NIDDM (diabetics), and 20 patients with only CAD (nondiabetics) were evaluated. Intracoronary ultrasound (ICUS) imaging with simultaneous intracoronary pressure (P2) recordings were performed at the imaging site with 0.014 in fiber-optic high fidelity pressure monitoring wire. The same wire was used as guide wire for the ICUS catheter. Sites with less than 50% luminal stenosis by ICUS were studied. Recordings were done before and after 300 microg of intracoronary nitroglycerin (IC-NTG). Electrocardiographic tracings recorded simultaneously with ICUS images were used for timing. Systolic and diastolic cross-sectional lumen area (CSLA) and coronary artery distensibility (C-DIST) were measured, C-DIST = [(systolic CSLA-diastolic CSLA)/[(intracoronary pulse pressure) x (diastolic CSLA)]] x 1,000. RESULTS: Diabetics had smaller CSLA (diabetics = 8.6 +/- 0.6 mm2, nondiabetics = 11.5 +/- 0.5 mm2, p < 0.01). Although C-DIST was similar before IC-NTG in the two groups, it became significantly lower in diabetics after IC-NTG (diabetics C-DIST = 3.02 +/- 0.14 mm Hg(-1), nondiabetics C-DIST = 4.21 +/- 0.15 mm Hg(-1), p < 0.01). Degrees of circumference involved, total plaque burden and composition were similar in both groups. CONCLUSIONS: Noninsulin dependent diabetes mellitus reduces C-DIST after IC-NTG administration. PMID- 10520794 TI - Long-term outcome of patients with unexplained syncope treated with an electrophysiologic-guided approach in the implantable cardioverter-defibrillator era. AB - OBJECTIVES: We evaluated the long-term outcome of patients with coronary artery disease and unexplained syncope who were treated with an electrophysiologic (EP) guided approach. BACKGROUND: Electrophysiologic studies are frequently performed to evaluate unexplained syncope in patients with coronary artery disease. Patients with this profile who have inducible ventricular tachycardia are considered at high risk for sudden death and increased overall mortality, and therefore are often treated with an implantable cardioverter-defibrillator (ICD). The impact of this EP-guided strategy is unknown because there are no data comparing the long-term outcome of ICD recipients with that of noninducible patients. METHODS: We evaluated 67 consecutive patients with coronary artery disease and unexplained syncope. All patients were treated with an EP-guided approach that included ICD implantation in patients with inducible ventricular tachycardia. RESULTS: Electrophysiologic testing suggested a plausible diagnosis in 32 (48%) of these patients. Inducible monomorphic ventricular tachycardia was the most common abnormality. Despite frequent appropriate therapy with ICDs, the total mortality for patients with inducible monomorphic ventricular tachycardia was significantly higher than for noninducible patients. The respective one- and two-year survival rates were 94% and 84% in noninducible patients and 77% and 45% in inducible patients (p = 0.02). CONCLUSIONS: Electrophysiologic testing suggests an etiology for unexplained syncope in approximately 50% of patients and risk stratifies these patients with regard to long-term outcome. Patients who receive an ICD for the management of inducible ventricular tachycardia have a high incidence of spontaneous ventricular arrhythmias requiring ICD therapy. However, despite ICD implantation and frequent appropriate delivery of ICD therapies, patients with inducible ventricular tachycardia have a significantly worse prognosis than do those who are noninducible. PMID- 10520795 TI - Relative effectiveness of the implantable cardioverter-defibrillator and antiarrhythmic drugs in patients with varying degrees of left ventricular dysfunction who have survived malignant ventricular arrhythmias. AVID Investigators. Antiarrhythmics Versus Implantable Defibrillators. AB - OBJECTIVES: We sought to assess the effect of baseline ejection fraction on survival difference between patients with life-threatening ventricular arrhythmias who were treated with an antiarrhythmic drug (AAD) or implantable cardioverter-defibrillator (ICD). BACKGROUND: The Antiarrhythmics Versus Implantable Defibrillators (AVID) study demonstrated improved survival in patients with ventricular fibrillation or ventricular tachycardia with a left ventricular ejection fraction (LVEF) < or =0.40 or hemodynamic compromise. METHODS: Survival differences between AAD-treated and ICD-treated patients entered into the AVID study (patients presenting with sustained ventricular arrhythmia associated with an LVEF < or =0.40 or hemodynamic compromise) were compared at different levels of ejection fraction. RESULTS: In patients with an LVEF > or =0.35, there was no difference in survival between AAD-treated and ICD treated patients. A test for interaction was not significant, but had low power to detect an interaction. For patients with an LVEF 0.20 to 0.34, there was a significantly improved survival with ICD as compared with AAD therapy. In the smaller subgroup with an LVEF <0.20, the same magnitude of survival difference was seen as that in the 0.20 to 0.34 LVEF subgroup, but the difference did not reach statistical significance. CONCLUSIONS: These data suggest that patients with relatively well-preserved LVEF (> or =0.35) may not have better survival when treated with the ICD as compared with AADs. At a lower LVEF, the ICD appears to offer improved survival as compared with AADs. Prospective studies with larger patient numbers are needed to assess the effect of relatively well-preserved ejection fraction (> or =0.35) on the relative treatment effect of AADs and the ICDs. PMID- 10520796 TI - Learning more about indications for implantable cardioverter-defibrillators. PMID- 10520797 TI - Detection of myocardial injury during radiofrequency catheter ablation by measuring serum cardiac troponin I levels: procedural correlates. AB - OBJECTIVES: In the present prospective controlled study, we measured blood levels of cardiac troponin I (cTnI) in patients undergoing radiofrequency (RF) catheter ablation (RFA), and we sought to investigate the degree of myocardial injury incurred by the application of RF energy and determine its procedural correlates. BACKGROUND: Measurement of serum creatine kinase (CK) levels after RFA may underestimate the degree of myocardial injury due to its thermal inactivation by RFA. Cardiac troponin I is a newer, more specific marker of myocardial injury, which may circumvent this limitation; its use in this setting has rarely been studied. METHODS: In 118 consecutive patients, 67 men and 51 women aged 38 +/- 19 years undergoing RFA for a variety of arrhythmias, cTnI and creatine kinase isoenzyme (CK-MB) levels were measured before, immediately after and 4 to 24 h after RFA. Cardiac troponin I was also measured in 39 patients (control group) having only electrophysiologic studies (EPS) without RFA. RESULTS: All RFA procedures were uncomplicated, lasted 3.2 +/- 2.0 h and included delivery of 16 +/- 22 (median: 9) RF current applications. Baseline cTnI levels averaged 0.17 +/ 0.18 ng/ml, rose to 0.88 +/- 1.12 at the end of RFA and to 2.19 +/- 2.46 at 4-24 h later. Creatine kinase isoenzyme was found to be elevated (>6 microg/l) in 32 patients (27%), while cTnI levels were increased (> or =1 ng/ml) in 80 patients (68%) (p = 0.0001). Cardiac troponin I levels correlated with the number of RF lesions applied (r = 0.53, p < 0.0001), the site of RFA, being higher with ventricular > atrial > annular lesions (p = 0.012) and the approach to the mitral annulus (transaortic > transseptal, p = 0.004). In a control group of 39 patients undergoing EPS, all but one patient had normal cTnI or CK-MB. CONCLUSIONS: The degree of myocardial injury incurred by RFA is far more accurately assessed by cTnI levels rather than by CK-MB measurements. Cardiac troponin I levels correlate with the number of RF lesions applied, the site of RFA and the approach to the mitral annulus. PMID- 10520798 TI - Large T wave inversion and QT prolongation associated with pulmonary edema: a report of nine cases. AB - OBJECTIVES: The purpose of this study is to describe a new clinical electrocardiographic phenomenon characterized by diffuse symmetrical T wave inversion and QT prolongation after recovery from an episode of cardiogenic but nonischemic pulmonary edema. BACKGROUND: A variety of clinical conditions, but not acute pulmonary edema, have been previously associated with giant negative T waves and QT prolongation in the postevent electrocardiogram. METHODS: In nine patients not suspected of having ischemic heart disease, new large or global T wave inversion with QT prolongation was observed after resolution of acute cardiogenic pulmonary edema. Each patient underwent detailed clinical evaluation including testing for myocardial injury and a coronary ischemic etiology. RESULTS: There were seven women and two men with ages ranging from 32 to 79 years. The etiology of pulmonary edema was diverse, but acute myocardial infarction and significant coronary artery disease were ruled out in each case. During the index event, most patients had elevated blood pressure, sinus tachycardia, minimal nonspecific ST and T wave changes and normal QT intervals. Large inverted T waves with marked prolongation of the QT intervals evolved within 24 h after clinical stabilization. The electrocardiographic changes gradually resolved in one week. There was no in-hospital mortality. CONCLUSIONS: Acute cardiogenic but nonischemic pulmonary edema may cause deep T wave inversion and QT prolongation after resolution of the symptoms. The repolarization abnormalities may last for several days. These electrocardiographic changes do not adversely effect short-term prognosis. PMID- 10520799 TI - In-hospital versus out-of-hospital presentation of life-threatening ventricular arrhythmias predicts survival: results from the AVID Registry. Antiarrhythmics Versus Implantable Defibrillators. AB - OBJECTIVES: This study describes the outcomes of patients from the Antiarrhythmics Versus Implantable Defibrillators (AVID) Study Registry to determine how the location of ventricular arrhythmia presentation influences survival. BACKGROUND: Most studies of cardiac arrest report outcome following out of-hospital resuscitation. In contrast, there are minimal data on long-term outcome following in-hospital cardiac arrest. METHODS: The AVID Study was a multicenter, randomized comparison of drug and defibrillator strategies to treat life-threatening ventricular arrhythmias. A Registry was maintained of all patients with sustained ventricular arrhythmias at each study site. The present study includes patients who had AVID-eligible arrhythmias, both randomized and not randomized. Patients with in-hospital and out-of-hospital presentations are compared. Data on long-term mortality were obtained through the National Death Index. RESULTS: The unadjusted mortality rates at one- and two-year follow-ups were 23% and 31.1% for patients with in-hospital presentations, and 10.5% and 16.8% for those with out-of-hospital presentations (p < 0.001), respectively. The adjusted mortality rates at one- and two-year follow-ups were 14.8% and 20.9% for patients with in-hospital presentations, and 8.4% and 14.1% for those with out-of hospital presentations (p < 0.001), respectively. The adjusted long-term relative risk for in-hospital versus out-of-hospital presentation was 1.6 (95% confidence interval [CI] 1.3-1.9). CONCLUSIONS: Compared with patients with out-of-hospital presentations of life-threatening ventricular arrhythmias not due to a reversible cause, patients with in-hospital presentations have a worse long-term prognosis. Because location of ventricular arrhythmia presentation is an independent predictor of long-term outcome, it should be considered as an element of risk stratification and when planning clinical trials. PMID- 10520800 TI - Electromechanical left ventricular behavior after nonsurgical septal reduction in patients with hypertrophic obstructive cardiomyopathy. AB - OBJECTIVES: To investigate the electromechanical consequences of nonsurgical septal reduction in a group of patients with hypertrophic obstructive cardiomyopathy (HOCM). BACKGROUND: Patients with HOCM may benefit symptomatically from nonsurgical septal reduction as an alternative to dual chamber pacing and sensing (DDD) pacing and surgical myectomy. METHODS: We studied 20 symptomatic patients with HOCM (12 men), mean age 52 +/- 17 years, before and after septal reduction using echocardiography and electrocardiogram (ECG). RESULTS: Septal reduction with a significant rise in cardiac enzymes was successfully achieved in all patients resulting in a 50% reduction in resting left ventricular (LV) outflow tract gradient within 24 h of procedure and an 80% reduction after six months. Left ventricular outflow tract diameter increased at 24 h with a further increase six months later. QRS duration increased by 35 ms at 24 h after procedure associated with right bundle branch block (RBBB) and significant rightward axis rotation in 16 patients. R-wave amplitude in V1 fell by 7 +/- 4 mm in 15/20 patients, 13 of whom developed reduction of septal long axis excursion. Left-axis deviation appeared in three patients and septal q-wave was suppressed in 12 long-axis excursion; peak shortening and lengthening rates all fell at the septal site by 20% at 24 h. Only septal excursion returned back to baseline values at six months. Wall motion also became incoordinate so that postejection septal shortening increased by three times control values at 24 h and by four times six months later. CONCLUSIONS: Nonsurgical septal reduction is associated with a drop in LV outflow tract obstruction and the creation of a localized myocardial infarction (MI) increasing LV outflow tract diameter. The technique also results in a consistent alteration of septal activation and secondary incoordination. The latter could play a significant role in gradient reduction and symptomatic improvement in a manner similar to that seen with DDD pacing. PMID- 10520801 TI - Changes in left ventricular filling and left atrial function six months after nonsurgical septal reduction therapy for hypertrophic obstructive cardiomyopathy. AB - OBJECTIVES: The purpose of this study was to evaluate changes in left ventricular (LV) filling, left atrial (LA) volumes and function six months after nonsurgical septal reduction therapy (NSRT) for hypertrophic obstructive cardiomyopathy (HOCM). BACKGROUND: Patients with HOCM frequently have enlarged left atria, which predisposes them to atrial fibrillation. Nonsurgical septal reduction therapy results in significant reduction in left ventricular outflow tract (LVOT) obstruction and symptomatic improvement. However, its effect on LV passive filling volume, LA volumes and function is not yet known. METHODS: Thirty patients with HOCM underwent treadmill exercise testing as well as 2-dimensional and Doppler echocardiography before and six months after NSRT. Data included clinical status, exercise duration, LVOT gradient, mitral regurgitant (MR) volume, LV pre-A pressure and LA volumes. Left atrial ejection force and kinetic energy (KE) were computed noninvasively and were compared with 12 age-matched, normal subjects. RESULTS: New York Heart Association (NYHA) class was lower and exercise duration was longer (p < 0.05) six months after NSRT. The LVOT gradient, MR volume and LV pre-A pressure were all significantly reduced. HOCM patients had larger atria, which had a higher ejection force and KE, compared with normal subjects (p < 0.01). After NSRT, LV passive filling volume increased (p < 0.01), whereas LA volumes, ejection force and KE decreased (p < 0.01). Reduction in LA maximal volume was positively related to changes in LV pre-A pressure (r = 0.8, p < 0.05) and MR volume (0.4, p < 0.05). Changes in LA ejection force were positively related to changes in LA pre-A volume (r = 0.7, p < 0.01) and KE (r = 0.81, p < 0.01). The increase in exercise duration paralleled the increase in LV passive filling volume (r = 0.85, p < 0.05). CONCLUSIONS: Nonsurgical septal reduction therapy results in an increase in LV passive filling volume and a reduction in LA size, ejection force and KE. PMID- 10520802 TI - Functional anatomy of mitral regurgitation: accuracy and outcome implications of transesophageal echocardiography. AB - OBJECTIVES: This study was performed to determine the accuracy and outcome implications of mitral regurgitant lesions assessed by echocardiography. BACKGROUND: In patients with mitral regurgitation (MR), valve repair is a major incentive to early surgery and is decided on the basis of the anatomic mitral lesions. These lesions can be observed easily with transesophageal echocardiography (TEE), but the accuracy and implications for outcome and clinical decision-making of these observations are unknown. METHODS: In 248 consecutive patients operated on for MR, the anatomic lesions diagnosed with TEE were compared with those observed by the surgeon and those seen on 216 transthoracic echocardiographic (TTE) studies, and their relationship to postoperative outcome was determined. RESULTS: Compared with surgical diagnosis, the accuracy of TEE was high: 99% for cause and mechanism, presence of vegetations and prolapsed or flail segment, and 88% for ruptured chordae. Diagnostic accuracy was higher for TEE than TTE for all end points (p < 0.001), but the difference was of low magnitude (<10%) except for mediocre TTE imaging or flail leaflets (both p < 0.001). The type of mitral lesions identified by TEE (floppy valve, restricted motion, functional lesion) were determinants of valve repairability and postoperative outcome (operative mortality and long-term survival; all p < 0.001) independent of age, gender, ejection fraction and presence of coronary artery disease. CONCLUSIONS: Transesophageal echocardiography provides a highly accurate anatomic assessment of all types of MR lesions and has incremental diagnostic value if TTE is inconclusive. The functional anatomy of MR defined by TEE is strongly and independently predictive of valve repairability and postoperative outcome. Therefore, the mitral lesions assessed by echocardiography represent essential information for clinical decision making, particularly for the indication of early surgery for MR. PMID- 10520803 TI - Progression of mitral regurgitation: a prospective Doppler echocardiographic study. AB - OBJECTIVES: This study was performed to define the rates and determinants of progression of organic mitral regurgitation (MR). BACKGROUND: Severe MR has major clinical consequences, but the rates and determinants of progression of the degree of regurgitation are unknown. Quantitative Doppler echocardiographic methods allow the quantitation of regurgitant volume (RVol), regurgitant fraction (RF) and effective regurgitant orifice (ERO) to define progression of MR. METHODS: In a prospective study of MR progression, 74 patients had two quantitative Doppler echocardiographic examinations of MR (with at least two methods) 561 +/- 423 days apart without an intervening event. RESULTS: Progression of MR was observed, with increase in RVol (77 +/- 46 ml vs. 65 +/- 40 ml, p < 0.0001), RF (47 +/- 16% vs. 43% +/- 15%, p < 0.0001), and ERO (50 +/- 35 mm2 vs. 41 +/- 28 mm2, p < 0.0001). Annual rates (95% confidence interval) were, respectively, 7.4 ml/year (5.1, 9.7), 2.9%/year (1.9, 3.9) and 5.9 mm2/year (3.9, 7.8). However, wide individual variation was observed, and regression and progression of RVol >8 ml was found in 11% and 51%, respectively. In multivariate analysis, independent predictors of progression of RVol were progression of the lesions, particularly a new flail leaflet (p = 0.0003), and progression of mitral annulus diameter (p = 0.0001). Regression of MR was associated with marked changes in afterload, particularly decreased blood pressure (p = 0.008). No significant effect of treatment was detected. CONCLUSIONS: Organic MR tends to progress over time with increase in volume overload (RVol) due to increase in ERO. Progression of MR is variable and determined by progression of lesions or mitral annulus size. These data should help plan follow up of patients with organic MR and future intervention trials. PMID- 10520804 TI - Mitral balloon valvotomy for patients with mitral stenosis in atrial fibrillation: immediate and long-term results. AB - OBJECTIVES: The purpose of this study was to examine the effect of atrial fibrillation (AF) on the immediate and long-term outcome of patients undergoing percutaneous mitral balloon valvuloplasty (PMV). BACKGROUND: There is controversy as to whether the presence of AF has a direct negative effect on the outcome after PMV. METHODS: The immediate procedural and the long-term clinical outcome after PMV of 355 patients with AF were prospectively collected and compared with those of 379 patients in normal sinus rhythm (NSR). RESULTS: Patients with AF were older (62 +/- 12 vs. 48 +/- 14 years; p < 0.0001) and presented more frequently with New York Heart Association (NYHA) class IV (18.3% vs. 7.9%; p < 0.0001), echocardiographic score >8 (40.1% vs. 25.1%; p < 0.0001), calcified valves under fluoroscopy (32.4% vs. 18.8%, p < 0.0001) and with history of previous surgical commissurotomy (21.7% vs. 16.4%; p = 0.0002). In patients with AF, PMV resulted in inferior immediate and long-term outcomes, as reflected in a smaller post-PMV mitral valve area (1.7 +/- 0.7 vs. 2 +/- 0.7 cm2; p < 0.0001) and a lower event free survival (freedom of death, redo-PMV and mitral valve surgery) at a mean follow-up time of 60 months (32% vs. 61%; p < 0.0001). In the group of patients in AF, severe post-PMV mitral regurgitation (> or =3+) (p = 0.0001), echocardiographic score >8 (p = 0.004) and pre-PMV NYHA class IV (p = 0.046) were identified as independent predictors of combined events at follow-up. CONCLUSIONS: Patients with AF have a worse immediate and long-term outcomes after PMV. However, the presence of AF by itself does not unfavorably influence the outcome, but is a marker for clinical and morphologic features associated with inferior results after PMV. PMID- 10520805 TI - Low prevalence of valvular heart disease in 226 phentermine-fenfluramine protocol subjects prospectively followed for up to 30 months. AB - OBJECTIVES: This investigation sought to determine the effect of phentermine fenfluramine (phen-fen) on the prevalence of valvular heart disease in 226 obese subjects enrolled in a prospective, strict weight loss, research protocol. BACKGROUND: Early reports have suggested that the use of phen-fen for weight loss may be associated with increased valvular heart disease. Such reports were based on small numbers of patients, limited data on dose and duration of phen-fen therapy, and no correlation with matched controls. METHODS: All subjects underwent transthoracic echocardiography for significant valvular lesions within a mean of 97 days from the manufacturer's announcement of the voluntary withdrawal of fenfluramine and dexfenfluramine. All echocardiograms were interpreted by two independent readers. RESULTS: The study population included 183 women and 43 men with a mean age of 46.9 +/- 8.9 years and mean starting body mass index of 39.8 +/- 7.7 kg/m2. Using the Food and Drug Administration criteria, significant aortic regurgitation was detected in 15 subjects (6.6%) and mitral regurgitation in 3 subjects (1.3%). Only one patient had significant regurgitation of both aortic and mitral valves. No valves had severe regurgitation. Significant valvular disease did not correlate with the dose or duration of phen-fen therapy. Furthermore, the prevalence of valvular regurgitation is comparable to the normal offspring in the Framingham Heart Study, who are similar in age, gender, and geographical location. CONCLUSIONS: Phen-fen therapy is associated with a low prevalence of significant valvular regurgitation. Valvular regurgitation in our subjects may reflect age-related degenerative changes. PMID- 10520806 TI - Fen/phen and valvular heart disease: if it sounds too bad to be true, perhaps it isn't. PMID- 10520807 TI - The influence of abciximab use on clinical outcome after aortocoronary vein graft interventions. AB - OBJECTIVES: The purpose of this study was to evaluate the effect of abciximab use on clinical outcome in aortocoronary vein graft interventions. BACKGROUND: Although large randomized trials have demonstrated a significant benefit of abciximab use in the setting of percutaneous coronary interventions, there is relatively little data with respect to the use of this agent in percutaneous vein graft interventions. METHODS: Three hundred and forty-three patients were identified; 210 undergoing vein graft intervention without abciximab and 133 patients with abciximab. RESULTS: There were differences in baseline clinical and angiographic characteristics between the two groups; advanced age, unstable angina, older vein grafts and thrombus containing lesions were relatively common in both groups. Angiographic and procedural success rates were similar with or without the use of abciximab (89% vs. 92%, p = 0.15, and 85% vs. 91%, p = 0.12, respectively). The in-hospital composite end point of death/Q-wave myocardial infarction (QWMI)/repeat revascularization was similar between the two groups. Utilizing statistical modeling to adjust for baseline differences between the groups, abciximab use did not influence the cumulative long-term composite end point of death/MI/repeat revascularization. CONCLUSIONS: This study demonstrates that in this relatively high-risk population undergoing aortocoronary vein graft interventions, the administration of abciximab periprocedurally does not appear to reduce major adverse clinical events. PMID- 10520808 TI - Neuroendocrine activation in heart failure is modified by endurance exercise training. AB - OBJECTIVES: The purpose of this study was to determine whether endurance exercise training could buffer neuroendocrine activity in chronic heart failure patients. BACKGROUND: Neuroendocrine activation is associated with poor long-term prognosis in heart failure. There is growing consensus that exercise may be beneficial by altering the clinical course of heart failure, but the mechanisms responsible for exercise-induced benefits are unclear. METHODS: Nineteen heart failure patients (ischemic disease; New York Heart Association [NYHA] class II or III) were randomly assigned to either a training group or to a control group. Exercise training consisted of supervised walking three times a week for 16 weeks at 40% to 70% of peak oxygen uptake. Medications were unchanged. Neurohormones were measured at study entry and after 16 weeks. RESULTS: The training group (n = 10; age = 61 +/- 6 years; EF = 30 +/- 6%) and control group (n = 9; age = 62 +/- 7 years; EF = 29 +/- 7%) did not differ in clinical findings at study entry. Resting levels of angiotensin II, aldosterone, vasopressin and atrial natriuretic peptide in the training and control groups did not differ at study entry (5.6 +/- 1.3 pg/ml; 158 +/- 38 pg/ml; 6.1 +/- 2.0 pg/ml; 37 +/- 8 pg/ml training group vs. 4.8 +/- 1.2; 146 +/- 23; 4.9 +/- 1.1; 35 +/- 10 control group). Peak exercise levels of angiotensin II, aldosterone, vasopressin and atrial natriuretic peptide in the exercise and control groups did not differ at study entry. After 16 weeks, rest and peak exercise hormone levels were unchanged in control patients. Peak exercise neurohormone levels were unchanged in the training group, but resting levels were significantly (p < 0.001) reduced (angiotensin -26%; aldosterone 32%; vasopressin -30%; atrial natriuretic peptide -27%). CONCLUSIONS: Our data indicate that 16 weeks of endurance exercise training modified resting neuroendocrine hyperactivity in heart failure patients. Reduction in circulating neurohormones may have a beneficial impact on long-term prognosis. PMID- 10520809 TI - Angiotensin I-converting enzyme and plasminogen activator inhibitor-1 gene variants: risk of mortality and fatal cardiovascular disease in an elderly population-based cohort. AB - OBJECTIVES: We studied the contribution of putative risk genotypes at the angiotensin I-converting enzyme inhibitor (ACE D/D) and plasminogen activator inhibitor-1 (PAI-1 4G/4G) loci to all-cause and cardiovascular mortality in a population-based cohort. BACKGROUND: The ACE D/D and PAI-1 4G/4G genotypes have been consistently associated with elevated plasma activities of the gene products. Their role in cardiovascular disease, although explored intensively, is still equivocal. METHODS: The ACE and PAI-1 genotypes were determined in 648 subjects > or =85 years old. In a cross-sectional analysis, the genotype distributions in a subset of 356 elderly subjects who were born in Leiden, The Netherlands, were compared with those in 250 young subjects whose families originated from the same geographic region. In addition, the complete cohort of elderly subjects was followed over 10 years for all-cause and cardiovascular mortality and was stratified according to genotype. RESULTS: In the cross sectional analysis, the ACE and PAI-1 genotype distributions were similar in elderly and young subjects. In the prospective follow-up study, however, the age adjusted risk of fatal ischemic heart disease was increased threefold (95% confidence interval [CI] 1.2 to 7.6) in elderly men carrying the PAI-1 4G/4G genotype. The risk of all-cause mortality was not increased among elderly subjects carrying the PAI-1 4G/4G (relative risk [RR] 0.9, 95% CI 0.7 to 1.1) or the ACE D/D genotype (RR 0.9, 95% CI 0.7 to 1.1), nor did we observe elevated risks of death from all cardiovascular diseases combined. There was no interaction between the genotypes. CONCLUSIONS: The PAI 4G/4G genotype may be a risk factor for fatal ischemic heart disease in elderly men. The impact of moderately increased ACE and PAI-1 activities associated with the ACE D/D and PAI 1 4G/4G genotypes is too small to affect mortality in the general population. PMID- 10520810 TI - The effects of chronic prostacyclin therapy on cardiac output and symptoms in primary pulmonary hypertension. AB - OBJECTIVES: This study evaluated the response to prostacyclin dose reduction in patients with primary pulmonary hypertension (PPH) who developed high cardiac outputs. BACKGROUND: Patients on prostacyclin require chronic upward dose titration to overcome tolerance to the medication. No upper limit of effective dose has been described. METHODS: We studied 12 patients with PPH treated with chronic prostacyclin therapy who presented in high cardiac output states. Each patient underwent prostacyclin dose reduction under hemodynamic guidance targeted to reduce the cardiac index to < or =4 liter/min/M2, unless rebound pulmonary hypertension occurred. Following dose reduction, patients were observed for changes in the effectiveness of the prostacyclin. RESULTS: Patients were treated for 39 +/- 20 months, resulting in a 71% reduction in pulmonary vascular resistance compared to baseline. At the time of their most recent evaluation their cardiac outputs were increased to 10.1 +/- 2.3 liter/min. The patients underwent a 39% dose reduction (range 12% to 78%) resulting in a change of mean PAP from 45 to 46 mm Hg (p = NS), cardiac index from 7.4 +/- 1.4 to 4 +/- 0.74 liter/min/M2 (p = 0.01), and pulmonary vascular resistance from 3.7 +/- 1.7 to 4.7 +/- 1.5 units (p < 0.001). In no instance did rebound pulmonary hypertension occur. However, the patients all retained their clinical benefit without a return of tolerance. CONCLUSIONS: Excessive prostacyclin in PPH can lead to a high cardiac output state, suggesting it has important positive inotropic effects. In this circumstance, reducing the dose can allow the cardiac output to return to normal without worsening the clinical state. PMID- 10520811 TI - Effect of orally active prostacyclin analogue on survival of outpatients with primary pulmonary hypertension. AB - OBJECTIVES: This study sought to investigate the effect of beraprost sodium (BPS), an orally active prostacyclin analogue, on the survival of outpatients with primary pulmonary hypertension (PPH). BACKGROUND: Continuous intravenous administration of epoprostenol (prostacyclin) has been shown to improve survival in PPH. However, the effect of oral BPS on survival in PPH remains unknown. METHODS: Fifty-eight consecutive patients with PPH who could be discharged after the first diagnostic catheterization for PPH were retrospectively divided into two groups: patients treated with BPS (BPS group, n = 24) and those without BPS (conventional group, n = 34). The baseline demographic and hemodynamic data did not significantly differ between the two. RESULTS: Twenty-seven patients died of cardiopulmonary causes in the conventional group during a mean follow-up period of 44 +/- 45 months. In contrast, only 4 patients died of cardiopulmonary causes in the BPS group during a mean follow-up period of 30 +/- 20 months. In a subsample (n = 15) of patients in the BPS group, mean pulmonary arterial pressure and total pulmonary resistance significantly decreased, respectively, by 13% and 25% during a mean follow-up period of 53 days. Among the variables previously known to be associated with the mortality in PPH, the absence of BPS therapy and the reduced cardiac output were independently related to the mortality by a multivariate Cox proportional hazards regression analysis (both p < 0.05). The Kaplan-Meier survival curves demonstrated that the one-, two- and three-year survival rates for the BPS group were 96%, 86% and 76%, respectively, as compared with 77%, 47% and 44%, respectively, in the conventional group (log-rank test, p < 0.05). CONCLUSIONS: The oral administration of BPS may have beneficial effects on the survival of outpatients with PPH as compared with conventional therapy alone. PMID- 10520812 TI - Validation of a new noninvasive method (contrast-enhanced transthoracic second harmonic echo Doppler) for the evaluation of coronary flow reserve: comparison with intracoronary Doppler flow wire. AB - OBJECTIVES: We tested the hypothesis that coronary flow reserve (CFR) in the left anterior descending coronary artery (LAD) as assessed by a new noninvasive method (contrast-enhanced transthoracic second harmonic echo Doppler) is in agreement with CFR measurements assessed by intracoronary Doppler flow wire. BACKGROUND: Contrast-enhanced transthoracic second harmonic echo Doppler is a novel noninvasive method to detect blood flow velocity and reserve in the LAD. However, it has not yet been validated versus a gold-standard method. METHODS: Twenty-five patients undergoing CFR assessment in the LAD by Doppler flow wire were also evaluated by contrast-enhanced transthoracic Doppler to record blood flow in the distal LAD at rest and during hyperemia obtained by adenosine i.v. infusion. In five patients CFR was evaluated twice (before and after angioplasty). RESULTS: As a result of the combined use of i.v. contrast and second harmonic Doppler technology, feasibility in assessing coronary flow reserve equaled 100%. The agreement between the two methods was high. In fact, in all but five patients the maximum difference between the two CFR measurements was 0.38. Overall, the prediction (95%) interval of individual differences was -0.69 to +0.72. Reproducibility of CFR measurements was also high. The limits of the agreement (95%) between the two measurements were -0.32 to +0.32. CONCLUSIONS: Coronary flow reserve in the LAD as assessed by contrast-enhanced transthoracic echo Doppler along with harmonic mode concurs very closely with Doppler flow wire CFR measurements. This new noninvasive method allows feasible, reliable and reproducible assessment of CFR in the LAD. PMID- 10520813 TI - Gene transfer of nitric oxide synthase: effects on endothelial biology. AB - OBJECTIVES: The purpose of the study was to investigate the role of nitric oxide (NO) in monocyte-endothelial interaction by augmenting NO release via transfection of human endothelial cells (ECs) with EC NO synthase (eNOS) DNA. BACKGROUND: Enhancement of NO synthesis by L-arginine or shear stress reduces endothelial adhesiveness for monocytes and inhibits atherogenesis. To elucidate further the underlying mechanism, we augmented NO synthase expression by transfection of human EC. METHODS: Liposome-mediated transfection of EC was performed with a plasmid construct containing the gene encoding eNOS. Expression of eNOS was confirmed by reverse transcription-polymerase chain reaction (RT PCR). Endothelial cells were exposed to human monocytoid cells, and adherent cells were quantitated using a computer-assisted program. Nitric oxide was measured by chemiluminescence. RESULTS: The NO levels were not different in EC that were either not transfected, transfected with beta-gal or liposomes only. The nitric oxide synthase (NOS) transfection increased NO release by +60% (n = 6), which increased further when EC were stimulated by shear stress (24 h) by +137% (n = 5) as compared with untransfected, unstimulated EC (both p < 0.05). The RT-PCR revealed diminished monocyte chemotactic protein-1 (MCP-1) expression in eNOS transfected EC. There was an inverse relation between NO levels and monocyte binding (r = -0.5669, p < 0.002). Stimulation of EC with tumor necrosis factor-alpha (TNF-alpha; 250 U/ml) led to a decrease in NO synthesis, and an increase in monocyte binding. Cells transfected with NOS were resistant to both effects of TNF-alpha. CONCLUSIONS: Endothelial cells transfected with eNOS synthesize an increased amount of NO; this is associated with diminished MCP-1 expression and monocyte-endothelial binding. The reduction in monocyte endothelial binding persists even after cytokine stimulation. PMID- 10520814 TI - Differential effects of alpha- and gamma-tocopherol on low-density lipoprotein oxidation, superoxide activity, platelet aggregation and arterial thrombogenesis. AB - OBJECTIVES: This study was designed to examine the differential effects of alpha- and gamma-tocopherol on parameters of oxidation-antioxidation and thrombogenesis. BACKGROUND: Experimental studies have shown that antioxidants, such as vitamin E (alpha-tocopherol), improve atherosclerotic plaque stability and vasomotor function, and decrease platelet aggregation and tendency to thrombus formation. METHODS: Sprague Dawley rats were fed chow mixed with alpha- or gamma-tocopherol (100 mg/kg/day) for 10 days. A filter soaked in 29% FeCl3 was applied around the abdominal aorta to study the patterns of arterial thrombosis. The aortic blood flow was observed and continuously recorded using an ultrasonic Doppler flow probe. ADP-induced platelet aggregation, low-density lipoprotein oxidation induced by phorbol 12-myristate 13-acetate (PMA)-stimulated leukocytes, superoxide anion generation and superoxide dismutase (SOD) activity were also measured. RESULTS: Both alpha- and gamma-tocopherol decreased platelet aggregation and delayed time to occlusive thrombus (all p < 0.05 vs. control). Both alpha- and gamma-tocopherol decreased arterial superoxide anion generation, lipid peroxidation and LDL oxidation (all p < 0.05 vs. control), and increased endogenous SOD activity (p < 0.05). The effects of gamma-tocopherol were more potent than those of alpha-tocopherol (p < 0.05). CONCLUSIONS: This study indicates that both alpha- and gamma-tocopherol decrease platelet aggregation and delay intraarterial thrombus formation, perhaps by an increase in endogenous antioxidant activity. Gamma-tocopherol is significantly more potent than alpha tocopherol in these effects. PMID- 10520815 TI - Antioxidant vitamins: sorting out the good and the not so good. PMID- 10520816 TI - The influence of low afterload on the nature of the stress-velocity relationship. AB - OBJECTIVES: Correct assessment of contractility by conventional methods during manipulation of afterload is often disappointing. To this purpose, the stress velocity relationship offers assessment of contractility at different levels of afterload. We decided to study the influence of afterload on the nature of the stress-velocity relation. BACKGROUND: Although linear at baseline conditions in a population older than two years, data in newborns or after administration of low dose dobutamine suggest a different nature of this relationship at low afterload. METHODS: Ten healthy piglets (five to six weeks; 11 to 13 kg) were studied. End systolic meridional wall stress (ESWS) and rate-corrected velocity of circumferential fiber shortening (VcFc) were measured in these piglets at baseline, after balloon occlusion of the descending aorta, and at nitroprusside infusion rates of 1, 2 and 5 microg/kg/min. To eliminate inotropic influences mediated by reflex tachycardia, we subsequently studied five piglets and six adult pigs after bilateral cervical vagotomy. RESULTS: The ESWS changed from a baseline mean of 50 g/cm2 to 137 g/cm2 after balloon occlusion and to 19 g/cm2 at 5 microg/kg/min of nitroprusside. The VcFc changed from 1.19 c/s (circumference/second) to values of 0.9 c/s and 1.73 c/s, respectively. The ensuing stress-velocity regression line proved to be curvilinear instead of linear. The steeper slope at low afterload could suggest enhanced contractility compared to expected values had the relationship been linear. CONCLUSIONS: Data from young piglets and adult pigs suggest a curvilinear relationship of the stress-velocity relationship. This could probably explain some of the "hypercontractile states" encountered in conditions with low afterload. PMID- 10520817 TI - The language of lysis: a proposal to standardize the nomenclature. PMID- 10520818 TI - Going, going . . . gone. PMID- 10520820 TI - AHA/ACC scientific statement: Assessment of cardiovascular risk by use of multiple-risk-factor assessment equations: a statement for healthcare professionals from the American Heart Association and the American College of Cardiology. PMID- 10520819 TI - ACC/AHA Guidelines for Coronary Artery Bypass Graft Surgery: A Report of the American College of Cardiology/American Heart Association Task Force on Practice Guidelines (Committee to Revise the 1991 Guidelines for Coronary Artery Bypass Graft Surgery). American College of Cardiology/American Heart Association. PMID- 10520821 TI - Hypertension detection, treatment and control: a call to action for cardiovascular specialists. PMID- 10520822 TI - A radical view of Helicobacter pylori. PMID- 10520823 TI - Does repeated paracentesis prevent spontaneous bacterial peritonitis? PMID- 10520824 TI - AICF and DIC in liver cirrhosis: expressions of a hypercoagulable state. PMID- 10520825 TI - The Rome criteria process: diagnosis and legitimization of irritable bowel syndrome. PMID- 10520826 TI - Reflux laryngitis: pathophysiology, diagnosis, and management. AB - Gastroesophageal reflux disease is felt to be associated with a variety of laryngeal conditions and symptoms of which "reflux laryngitis" is perhaps the most common. The most likely mechanism for laryngeal injury and symptoms is secondary to direct acid and pepsin contact, although studies concerning the cause and effect between gastroesophageal reflux disease and laryngeal disorders are conflicting. Likewise, the most effective method to diagnose such patients is unclear. Empiric treatment of patients with reflux laryngitis has been shown to be effective though none of the studies are controlled. PMID- 10520827 TI - Esophageal submucosal glands: structure and function. AB - A three-tiered defense system exists in the esophagus, which serves a dual purpose of both limiting the degree of gastroesophageal reflux and minimizing the risk of acid-induced mucosal injury. The antireflux barrier, composed of both the lower esophageal sphincter and the diaphragmatic pinchcock, is the first line of defense and serves to limit the frequency and volume of refluxed gastric contents. When the antireflux barrier fails, the second line of defense, esophageal clearance, comes into play and serves to limit the duration of contact between gastric contents and the esophageal epithelium. Mechanisms involved in esophageal clearance include gravity and esophageal peristalsis, which remove volume, and secretions from swallowed saliva and esophageal submucosal glands, which neutralize acid. The third line of defense, tissue resistance, is necessary when acid contact time is prolonged such as when esophageal clearance is either ineffective or not operative (e.g., during sleep). Most studies that have examined esophageal clearance mechanisms have focused on the roles of esophageal peristalsis and salivary secretion, but the role of submucosal gland secretions is less well understood. This article reviews the structure and function of esophageal submucosal glands and discusses the potential role of their secretory products in esophageal clearance and tissue resistance. PMID- 10520828 TI - Gastroesophageal reflux and Barrett's esophagus in adults born with esophageal atresia. AB - OBJECTIVE: Postoperative morbidity after correction of esophageal atresia is partly determined by gastroesophageal reflux disease, which has been proven to affect from one-half to two-thirds of patients during childhood. We conducted a follow-up study to test our hypothesis that, if former patients still show gastroesophageal reflux at adult age, they are at high risk for developing Barrett's esophagus, which is considered to be premalignant. METHODS: Of 69 patients born between 1971 and 1978, all having undergone a primary anastomosis, 24 had died, five of them because of aspiration. Of the 45 survivors, 39 could be traced; they all completed a questionnaire inquiring after symptoms related to the esophagus. Of these patients, 34 underwent an additional esophagogastrocopy. RESULTS: Only nine of the 39 patients had no symptoms at all; 30 had mild to severe dysphagia symptoms, and 13 had mild to severe reflux symptoms. Esophagogastrocopy in 34 patients revealed that the anastomosis was still recognizable in all cases, but stenoses were not found. Six patients showed a small hiatal hernia, and one a large one. The incidences of reflux symptoms (13/39, p < 0.01), reflux esophagitis (9/34, p < 0.01) and Barrett's esophagus (2/34, p < 0.001) were significantly higher than in the normal population. CONCLUSIONS: This group seems to be at risk for developing Barrett's esophagus. As this is the first follow-up study of a consecutive group of adult esophageal atresia patients, we think it is advisable to perform an esophagogastroscopy in all patients at adulthood until more long term follow-up data are available. PMID- 10520829 TI - Biomarker studies in reversed Barrett's esophagus. AB - OBJECTIVE: The purpose of this study was to use biomarkers to assess for cancer risk in Barrett's esophagus patients with either squamous islands or complete reversal of their intestinal metaplasia to squamous epithelium. METHODS: The biomarkers included proliferation characteristic using Ki-67, p53 abnormalities using immunohistochemical methods with two antibodies, DO-1 and DO-7, and ornithine decarboxylase (ODC) activity. RESULTS: Eleven patients had complete reversal produced by a combination of acid suppression and thermal injury (multipolar electrocoagulation). Ki-67 staining was indistinguishable from that of normal squamous esophageal epithelium, i.e., basal layer staining only. All 11 cases were negative for p53. ODC activity was low and in the range for normal squamous epithelium. Fourteen patients had squamous islands (partial reversal) after prolonged proton pump inhibitor therapy. Multilayer Ki-67 staining occurred in nine cases (64%), and six (43%) had areas of positive p53 staining. CONCLUSIONS: Initial biomarker studies suggest that completely reversed squamous epithelium is biologically similar to normal squamous epithelium and of low cancer risk. In contrast, partial reversal, manifest as squamous islands, is accompanied by biomarker abnormalities. PMID- 10520830 TI - Detection of Epstein-Barr virus in esophageal squamous cell carcinoma in Taiwan. AB - OBJECTIVE: Recently, an association between viral infection and the development of esophageal carcinoma has been reported, particularly the human papilloma virus (HPV) and Esptein-Bar virus (EBV). However, geographic variation in carcinogenesis is realized. In this study, we investigate the viral carcinogenesis and the biologic effect of viral infection on esophageal squamous cell carcinoma (ESCC) in Taiwan. METHODS: To determine the association of viral infection (EBV and HPV) with ESCC, we applied polymerase chain reaction (PCR), immunohistochemistry, and in situ hybridization (ISH) to examine 119 surgical specimens from different sites of esophagus in 31 ESCC patients. Additionally, an immunoperoxidase method was used to detect EBV latent membrane protein-1 (LMP-1), p53, CD45RO (UCHL-1), Fas ligand (Fas L), and RNA ISH with oligonucleotide sequences was used to detected interleukin-6 (IL-6) mRNA. RESULTS: By PCR, EBV DNA was detected in 11 cases (35.5%). Expression of EBERs in ESCC was further confirmed with ISH. Nonetheless, no LMP-1 expression was detected. On the other hand, human papillomavirus (HPV) was identified in only one case (3.2%) of ESCC. Furthermore, HPV was located by ISH in the distant normal region rather than in tumor cells. In EBV-positive cases, accumulation of p53 protein was detected in 10 lesions (91%); CD45RO+ lymphocytes together with expressions of FasL and IL-6 were respectively identified in 100%, 63.6%, and 54.5% of 11 EBV-positive lesions. Interestingly, in the EBV-negative cases (n = 20), p53 protein was detected in 40% of lesions; CD45RO 30%; FasL 50%, and IL-6 10%. CONCLUSIONS: In this study, no correlation was found between the presence of EBV in ESCC and the patients' age, sex, as well as survival. Although our results indicate that EBV could be associated with ESCC, the clinical role of EBV in ESCC remains to be determined. PMID- 10520831 TI - Association of obesity with hiatal hernia and esophagitis. AB - OBJECTIVE: Although weight loss is commonly recommended for symptoms of gastroesophageal reflux, a relationship between excessive body weight and esophageal reflux has not been established. The aim of this study was to determine whether obesity is associated with the presence of a hiatal hernia (HH) and/or an endoscopic diagnosis of esophagitis. METHODS: Retrospective case control studies were done using 1389 patients who underwent gastric analysis and upper GI endoscopy between 1974 and 1995. After excluding patients with Zollinger Ellison syndrome, 189 cases of esophagitis with 1024 controls were identified. In a separate analysis of the database, 151 cases of HH with 1053 controls were also identified. Patients were classified by body mass index (BMI) as: thin (BMI <20 kg/m2), normal (BMI 20-25), mildly obese (BMI 25-30), and obese (BMI >30). RESULTS: Excessive body weight was significantly associated with the presence of HH, the probability of HH increasing with each level of BMI (p < 0.01), as well as with esophagitis (OR 1.8; 95% CI 1.4-2.1). HH was independently associated with esophagitis (OR 4.2 95% CI 2.9-6.1); when controlled for the effect of HH, the association between BMI and esophagitis diminished but remained significant. In the population as a whole, for the presence of esophagitis multiple logistic regression indicates BMI and hiatal hernia were significant factors but gender and race did not appear to be. CONCLUSIONS: Excessive body weight is a significant independent risk factor for hiatal hernia and is significantly associated with esophagitis, largely through an increased incidence of hiatal hernia. Whites are more likely to have the combination of esophagitis and hiatal hernia than are blacks. PMID- 10520832 TI - Prevalence and impact of upper gastrointestinal symptoms in the Canadian population: findings from the DIGEST study. Domestic/International Gastroenterology Surveillance Study. AB - OBJECTIVE: The prevalence and impact of upper gastrointestinal (GI) symptoms in the general population are poorly defined. Most data are obtained from selected samples derived from patients presenting to health care providers. As part of a larger international effort (The DIGEST study), we examined the prevalence of upper GI symptoms among the general Canadian population, as well as their psychosocial and economic impact. METHODS: A sample of 1036 adults was studied, its demographic characteristics closely matching those of the general Canadian population. A validated detailed questionnaire measured the prevalence, severity, and frequency of 15 digestive symptoms, as well as demographic information, use of medication and medical resources, other illnesses, and dietary habits. The Psychological General Well-Being Index, a self-administered questionnaire, assessed the individual's subjective sense of well-being. RESULTS: Of the sample population, 28.6% reported substantial symptoms in the preceding 3 months, the majority (111/153 subjects) for >1 yr; 34.1% reported having never experienced significant GI symptoms. The most bothersome symptoms were primarily related to dysmotility-like symptoms in 54.9% of those with chronic symptoms, ulcer-like symptoms in 12.4%, and related to heartburn in 42.5%. Chronic upper GI symptoms were associated with a highly significant (p < 0.001) decrease in all facets of the Psychological General Well Being Index. CONCLUSIONS: Upper GI symptoms are very prevalent in the general Canadian population and substantially affect the quality-of-life and psychological well-being of those affected. Dysmotility-like symptoms, rather than heartburn, are the most common chronic upper gastrointestinal symptoms in the general population. PMID- 10520833 TI - Cyclic vomiting syndrome in adults: clinical features and response to tricyclic antidepressants. AB - OBJECTIVE: Cyclic vomiting syndrome (CVS) has been described infrequently in adults, and treatment in both children and adults remains unsatisfactory. We report clinical features of a group of adults with CVS and anecdotal outcome from open-label treatment with tricyclic antidepressants, medications that have some efficacy in other unexplained gastrointestinal disorders. METHODS: Clinical data were examined from 17 adult patients with CVS seen over a 10-yr period, each having been treated with a tricyclic antidepressant. Outpatient records were reviewed, clinical outcome was extracted using a priori criteria, and findings were compared with 37 patients having usual functional nausea and vomiting who also received tricyclic antidepressant therapy. RESULTS: Symptoms in CVS began at age 35 yr (range 14-73 yr); the average episode length was 6 days (range 1-21 days) and the symptom-free interval averaged 3.1 months (range 0.5-6 months). Vomiting cycles typically began without warning, and fewer than one-third of the subjects reported a prodrome or potential trigger event, such as menstrual periods, pregnancy, or large meals. Sleep was seemingly beneficial in 23.5%. Tricyclic antidepressant therapy was associated with complete remission in 17.6% and partial response in 58.8%, but was less effective than for functional nausea and vomiting (p = 0.02). CONCLUSIONS: CVS is a rare diagnosis with distinctive features in adults. Duration of episodes and cycles varies considerably across subjects. In open-label, uncontrolled use, tricyclic antidepressants appear beneficial for some subjects but are less effective in CVS than in chronic, persistent functional nausea and vomiting. PMID- 10520834 TI - Simultaneous measurement of gastric emptying of a solid meal by ultrasound and by scintigraphy. AB - OBJECTIVE: Although ultrasonic imaging may represent a valid alternative to scintigraphy for measurement of gastric emptying, most studies comparing the two methods have been carried out with liquid meals. The aim of this study was to compare scintigraphic and ultrasonographic measurements of gastric emptying of a solid meal in healthy subjects and in patients with possible delay in emptying. METHODS: Nineteen subjects were studied: five controls, six patients with gastroesophageal reflux, and eight patients with dysmotility-like dyspepsia. Gastric emptying was measured by both scintigraphy and ultrasonography after ingestion of an 800-calorie solid, realistic meal containing 99mTc-labeled chicken liver. Scintigraphic measurements were made every 15 min for 6 h, and ultrasonic imaging of antral sections was undertaken every 15 min for the first 1 h and every 30 min thereafter. Total emptying times were calculated independently using the two methods, and the emptying patterns recorded by the two methods were compared. RESULTS: Maximal antral dilation occurred 30 min (range 0-90 min) after the end of the meal and persisted until 96 +/- 42 min, by which time gastric radioactivity had decreased from its maximum by 43% +/- 23%. From this time on, the antral cross-sectional area returned toward the basal value, declining faster than the gastric counts recorded by scintigraphy. Total emptying times measured by ultrasound and by scintigraphy were in good agreement in all subjects, with a mean difference of only 4.5 min (limits of agreement, -17.1 to 21.6 min). CONCLUSIONS: Ultrasonographic measurement of antral cross-sectional area provides a valid alternative to scintigraphy for the measurement of total gastric emptying of a solid meal. It is less reliable if other parameters of gastric emptying such as T(1/2) are required. PMID- 10520835 TI - Evaluation of gastric emptying and motility in diabetic gastroparesis with magnetic resonance imaging: effects of cisapride. AB - OBJECTIVE: The motor mechanisms that underlie both slow gastric emptying in diabetic gastroparesis and its acceleration by cisapride are poorly understood. We have recently shown that magnetic resonance imaging (MRI) allows concurrent evaluation of both gastric emptying and regional gastric motility. METHODS: Emptying and motility were measured in eight diabetic patients with previously demonstrated delayed gastric emptying using a rapid MRI technique during oral administration of cisapride and placebo. Studies were performed in a double blind fashion and each patient acted as his own control. Subjects were studied supine for 120 min in a 1.5 Tesla MRI scanner after ingestion of 500 ml of 10% Intralipid. Gastric emptying corrected for the volume of secretions was determined every 15 min using transaxial scans. Each transaxial scan was followed by 120 coronal scans at 1 s intervals. Coronal scans were angled to provide simultaneous imaging of the proximal and distal stomach. MRI studies were also performed in seven diabetic patients with normal emptying who served as disease controls. RESULTS: Emptying was slower in the gastroparetic patients (t(1/2): 124 +/- 10 min) compared to patients with normal emptying (81 +/- 9 min, p < 0.05). Cisapride accelerated gastric emptying (74 +/- 5 vs 124 +/- 10 min) in patients with gastroparesis. The contraction amplitudes in the proximal stomach of gastroparetic patients were increased during cisapride treatment (17.2% +/- 1.8% vs 13.2% +/- 0.6%; p < 0.02), whereas antral contraction frequency, amplitude, and velocity were unchanged. CONCLUSIONS: We conclude that cisapride-induced acceleration of liquid gastric emptying in diabetic gastroparesis does not appear to result from changes in antral contractility, but may be related to changes in proximal gastric tone or gastric outlet resistance. PMID- 10520836 TI - Inhibition of pentagastrin-induced gastric acid secretion by intravenous pantoprazole: a dose-response study. AB - OBJECTIVE: The purpose of this study was to compare the gastric acid inhibitory ability of increasing doses of intravenous (i.i.) pantoprazole with that of i.v. famotidine and placebo. Pentagastrin was infused continuously in healthy subjects as a model for patients with Zollinger-Ellison syndrome. METHODS: Pentagastrin (1 microg/kg/h) was infused to stimulate maximum acid output in 39 subjects over a 25-h period. After 60 min of pentagastrin infusion, subjects received a single dose of i.v. pantoprazole (20, 40, 80, or 120 mg), i.v. famotidine (20 mg), or saline placebo. The variables measured were onset of response (time until acid output fell to < 10 mEq/h), duration of response (time acid output remained < 10 mEq/h), and cumulative acid output over 24 h. RESULTS: All doses of i.v. pantoprazole produced a dose-dependent suppression of acid output to < 10 mEq/h. Single i.v. doses of pantoprazole, 80 and 120 mg, suppressed acid output by > 90% in all subjects for < or = 21 h and had an onset of action of < 1 h. CONCLUSIONS: Intravenous pantoprazole has a rapid onset and a clear dose-related effect, with a significantly longer duration of action than that of i.v. famotidine. PMID- 10520837 TI - Isolation of Helicobacter pylori from vomitus in children and its implication in gastro-oral transmission. AB - OBJECTIVE: The route of transmission of Helicobacter pylori (H. pylori) is unclear. Gastro-oral transmission via contaminated vomitus has been proposed as an important mode of transmitting H. pylori, especially in children. This pilot study attempted to isolate H. pylori from the vomitus of children. METHODS: Children presenting for evaluation with gastroenteritis-associated vomiting were studied. Fresh vomitus samples were collected for detection of H. pylori by bacteriological culture and polymerase chain reaction, (PCR). A rapid, whole blood test was used to determine the H. pylori status of patients. RESULTS: A total of 18 children with mean age of 6 yr were studied; four had a positive serology test. Among these four children, H. pylori was isolated from vomitus by culture in one child and by PCR in two. An 18-month-old girl with negative serology had H. pylori detected in vomitus by PCR. Six months later, she had seroconversion confirmed, suggesting that she had an acute H. pylori infection on initial presentation. CONCLUSIONS: This is the first study reporting successful isolation of H. pylori from naturally produced vomitus. The result implies that transmission of H. pylori infection by vomitus, especially in children, is possible. PMID- 10520838 TI - Eradication of Helicobacter pylori normalizes elevated mucosal levels of epidermal growth factor and its receptor. AB - OBJECTIVE: Helicobacter pylori (H. pylori) infection has been linked to gastric cancer. The factors that promote carcinogenesis remain unknown. Epidermal growth factor (EGF) has been shown to be a potent epithelial mitogen and oncoprotein when sustained over expression occurs. Our aim was to compare gastric mucosal levels of EGF and its receptor (EGFR) among controls, H. pylori infected subjects, and subjects following H. pylori eradication using quantitative flow cytometric analysis. METHODS: Patients referred for evaluation of dyspepsia underwent EGD and six antral biopsies were performed (two each for rapid urease testing (RUT), histopathology, and flow cytometry). Controls were those found to be H. pylori negative while subjects had confirmed infection. The study patients were treated, then had repeat EGD with biopsies. RESULTS: There were 17 controls and 28 cases. Mean EGF and EGFR values were 2.69 and 2.46 for controls and 4.67 and 4.64 for subjects. Subjects' mean EGF was 73% higher (p = .035) and EGFR was 88% higher (p = 0.029) than controls. After treatment, the subjects' mean values declined 55% (p = 0.0001) for EGF and 40% (p = 0.002) for EGFR. Three subjects had persistent infection and showed no change in their EGF/EGFR levels. No difference was found among factor levels with respect to endoscopic findings. CONCLUSIONS: Both EGF and EGFR from gastric antral biopsies are increased nearly 2-fold in infection with H. pylori. Infection eradication reduces levels of both factors to those of controls. One major pathogenic mechanism for gastric mucosal hyperproliferation and possibly carcinogenesis related to H. pylori may be the over expression of EGF and increased receptor density of EGFR on gastric mucosal cells. PMID- 10520839 TI - Evaluation of upper gastrointestinal tract symptoms in patients infected with HIV. AB - OBJECTIVE: Upper gastrointestinal tract (UGI) symptoms are frequent in patients infected with the human immunodeficiency virus (HIV), but little published information exists about their characteristics or methods of evaluation. We evaluated the prevalence of nonesophageal UGI symptoms in a referral population, the utility of esophagogastroduodenoscopy (EGD) for diagnosis, and clinical predictors of abnormal endoscopic findings in patients infected with HIV. METHODS: All HIV-infected patients referred to.the outpatient gastroenterology clinics were prospectively evaluated using recognized symptom questionnaires. EGD indications, results, and the patients' self-reported symptom scores were compared. HIV-infected patients undergoing EGD were compared with HIV-infected patients not receiving an EGD and with symptomatic non-HIV-infected patients undergoing EGD. RESULTS: A total of 201 patients completed 280 questionnaires. Among 93 patients who underwent endoscopy, severe symptoms occurring at least several times per week included: anorexia (70%), upper abdominal pain (34%), vomiting (32%), or a recent weight loss of approximately 15 lb (31%). Patients undergoing EGD had more frequent/severe symptoms, but did not have differences in overall well-being or mean GI symptom score. The frequency of substantial and treatable endoscopic findings among patients infected with HIV was comparable to that found in the non-HIV-infected control group. There were no independent symptoms predicting substantial or treatable disease on EGD. CONCLUSIONS: We conclude that: 1) upper gastrointestinal symptoms are common in HIV-infected patients referred for GI consultation; 2) symptomatic HIV patients have a high prevalence of both treatable and untreatable upper GI pathologies; 3) and physicians use symptom frequency and severity to select patients for EGD, but these factors correlate poorly with abnormalities on EGD. Given this discrepancy, longitudinal study is needed to determine whether treating endoscopic abnormalities improves UGI symptoms. PMID- 10520840 TI - Efficacy of endoscopic clipping for bleeding gastroduodenal ulcer: comparison with topical ethanol injection. AB - OBJECTIVE: Although endoscopic clipping is used widely for the treatment of bleeding gastroduodenal ulcers, clinical trials on its efficacy are scarce. The aim of this study is to assess the efficacy and safety of endoscopic clipping for hemostasis from bleeding gastroduodenal ulcers. METHODS: The present study was designed as a retrospective study using historical controls. One hundred consecutive patients with bleeding gastroduodenal ulcers were treated by endoscopic clipping. The preceding 91 consecutive patients treated by endoscopic pure ethanol injection were regarded as controls. Forty-nine of the clipping group and 41 of the ethanol group had lesions at sites difficult to perform endoscopic manipulation. Hemostatic rates, rebleeding rates, amounts of blood transfusion, and durations of hospital stay were analyzed. RESULTS: The hemostatic rate was 96% in both clipping and ethanol groups, whereas the rebleeding rate was lower (15% vs 29%, p = 0.023) in the former than the latter. In technically difficult cases, the hemostatic rate was comparable (96 vs 90%). CONCLUSION: In patients with bleeding gastroduodenal ulcers, endoscopic clipping may be a choice of therapy because of a low rebleeding rate compared with pure ethanol injection. PMID- 10520841 TI - Transnasal endoscopy for enteral feeding tube placement in critically ill patients. AB - OBJECTIVE: Early enteral feedings may improve outcomes in critically ill patients. Recently, transnasal endoscopy with an ultrathin transnasal endoscope has been shown to be of value for diagnostic endoscopy without conscious sedation. We developed a technique for the placement of postpyloric feeding tubes in critically ill patients using transnasal endoscopy. We describe our initial experience in a consecutive series of patients. METHODS: We collected data on consecutive intensive care unit patients undergoing bedside transnasal endoscopy for nasoenteric feeding tube placement using a standardized technique. Tube position was confirmed in all patients with a plain abdominal radiograph. Tube placement was deemed successful if the feeding tube traversed the pylorus. RESULTS: Transnasal endoscopy was completed in all fourteen patients, as was placement of a feeding tube. Feeding tubes were successfully placed in the jejunum or duodenum in 13 of the 14 patients (93%). Tubes remained in place from 3 to 45 days (mean 16 days). Two patients required conscious sedation during tube placement, and two ultimately required percutaneous gastrostomy. CONCLUSIONS: Transnasal endoscopy allows simple and successful postpyloric feeding tube placement at the bedside of critically ill patients. This method can facilitate early enteral feeding in intensive care units. PMID- 10520842 TI - Hyoscyamine as a pharmacological adjunct in colonoscopy: a randomized, double blinded, placebo-controlled trial. AB - OBJECTIVE: Investigators have assessed the utility of antispasmodic agents in colonoscopy, with conflicting results. The aim of this study is to determine the effects of premedication with hyoscyamine, an anticholinergic antispasmodic, on outcomes in colonoscopy. METHODS: A total of 165 patients undergoing elective colonoscopy were randomized in a double blinded fashion to one of three arms: intravenous hyoscyamine (0.25 mg), oral hyoscyamine (0.25 mg), or placebo, administered 20-40 min before colonoscopy. Primary outcome measures included insertion time to cecum, patient's assessment of pain, and physician assessment of spasm. Secondary outcome measures included amount of analgesic medications used, total procedure time, amount and type of pathology visualized, and physician assessment of patient's pain. RESULTS: Bivariate analysis showed no difference between the three groups in insertion time (13.8 min, 14.8 min, and 13.8 min for placebo, intravenous hyoscyamine, and oral hyocyamine, respectively), analgesic medication necessary, or any other primary or secondary outcome variable. Multivariate analysis controlling for potential confounders also failed to demonstrate any differences between the groups. Women had higher procedure duration and analgesic requirement, and reported more pain than did men. CONCLUSIONS: This randomized, double blinded, placebo-controlled trial did not demonstrate efficacy of either intravenous or oral hyoscyamine as a premedication for colonoscopy. PMID- 10520843 TI - Treatment of chronic bleeding from gastric antral vascular ectasia (GAVE) with estrogen-progesterone in cirrhotic patients: an open pilot study. AB - OBJECTIVE: Gastric antral vascular ectasia (GAVE) is a rare cause of chronic bleeding in cirrhotic patients. Treatment of GAVE with surgical or nonsurgical portal decompression, beta-blockers, or endoscopic therapy provides disappointing results. In the present study, we evaluated the efficacy of estrogen-progesterone therapy, which has been reported to control chronic bleeding in gastrointestinal vascular malformations, such as Osler-Weber Rendu disease or angiodysplasia, in GAVE-related chronic bleeding. METHODS: Six cirrhotic patients who bled chronically from GAVE were included. Three had alcoholic cirrhosis, two cryptogenic cirrhosis, and one primary biliary cirrhosis. Grade 1 esophageal varices were noted in four patients. Bleeding could not be controlled by beta blockers, and endoscopic therapy was not considered given the extension of the antral vascular lesions. RESULTS: Before the start of therapy, transfusion requirements averaged 3.5 units/month over a 1.5-11 month period of observation. Patients were then treated with a combination of ethynil estradiol 30 microg and noretisterone 1.5 mg daily. During follow-up (range 3-12 months), bleeding did not recur in four patients; in one patient, treatment with estrogen progesterone decreased the need for transfusions from 4 units/month to 1.4 unit/month; this patient stopped the treatment inadvertently after 6 months and severe anemia recurred with a need for 4 units of blood in the following month; reintroduction of the treatment resulted in an increase of hemoglobin levels without the need for blood transfusions during the following 4 months. In the last patient, a 5 month treatment did not improve chronic bleeding. CONCLUSIONS: The present study suggests that estrogen-progesterone therapy is useful in the treatment of chronic bleeding related to GAVE; however, these findings require confirmation by a controlled trial. PMID- 10520844 TI - Predictive value of the Rome criteria for diagnosing the irritable bowel syndrome. AB - OBJECTIVE: Our aim was to examine the predictive value of the Rome criteria and absence of so-called "red flags" of clinical practice for diagnosing irritable bowel syndrome. Red flags were relevant abnormalities on physical examination, documented weight loss, nocturnal symptoms, blood in stools, history of antibiotic use, and family history of colon cancer. METHODS: In retrospective studies, 98 patients who had one or more Rome criteria and lacked red flags were identified by chart review of a 1-yr period. In prospective studies, 95 patients were identified who met the Rome criteria and lacked red flags. Sensitivity, specificity, predictive value of Rome criteria, and absence of red flags were determined. Consultant's final diagnosis was the gold standard. Investigations before and after referral were recorded and reason for referral was determined in prospective studies. RESULTS: In the retrospective series, the Rome criteria and absence of red flags had a sensitivity of 65%, specificity of 100%, and positive predictive value of 100%. None of these patients required revision of their diagnosis during a 2-yr follow-up. In the prospective study, the positive predictive value was 98%. More than 50% of the patients in this group had been referred because of diagnostic uncertainty and 24% had had an abdominal ultrasound; 66% of those <45 yr old underwent at least partial colonic evaluation. CONCLUSIONS: These findings suggest that the Rome criteria combined with a lack of red flags have a very high predictive value for diagnosing irritable bowel syndrome. Application of these diagnostic criteria has the potential to alter utilization of health care resources. PMID- 10520845 TI - The effects of migration on ulcerative colitis: a three-year prospective study among Europeans and first- and second- generation South Asians in Leicester (1991 1994). AB - OBJECTIVE: Our aim was to measure prospectively the incidence of ulcerative colitis in Leicester City and to compare this with a previous retrospective study in the same area. We also sought to compare the incidence and disease extent in the European community with that of the South Asian community and to compare the disease extent between first- and second-generation South Asian migrants. METHODS: A 3-yr prospective study of ulcerative colitis in the city of Leicester took place from October 1, 1991 to September 30, 1994 and included all cases resident in Leicester City and diagnosed as having ulcerative colitis, regardless of the extent and severity of the disease. RESULTS: Extensive colitis was commoner in second-generation migrants than in the first generation (chi2 = 4.3, p = 0.04) and was comparable to the European community. The annual average incidence of ulcerative colitis was 9.1/10(5) population/yr (95% confidence interval [CI] 7.1-11.3), which is similar to the previous retrospective study. However, the annual average incidence of ulcerative colitis in the European population was 7.0/10(5) population/yr (95% CI 5-9.5), whereas that of the South Asian population was 17.2/10(5) population/yr (95% CI 11.8-24.3), confirming that the risk of ulcerative colitis in this particular community is exceptionally high. CONCLUSIONS: These early results suggest that the disease pattern follows that of the indigenous population after only one generation and requires monitoring over the next decade. The incidence of ulcerative colitis in the South Asian population is high and continuing to rise. PMID- 10520847 TI - Symptom differences in moderate to severe IBS patients based on predominant bowel habit. AB - OBJECTIVE: We sought to determine if irritable bowel syndrome (IBS) patients with different bowel habit predominance differ in self-reported viscerosensory symptoms related to the upper and lower gastrointestinal (GI) tract, somatosensory symptoms, and constitutional functions. METHODS: Six hundred and twenty-five Rome criteria-positive IBS patients completed a bowel symptom questionnaire (BSQ), psychological symptom checklist (SCL-90), and health status (SF-36). Bowel habit predominance for IBS patients was determined using the Rome criteria for functional constipation (IBS-C; n = 140) and functional diarrhea (IBS-D; n = 216). The BSQ included questions about viscerosensory symptoms of the upper (chest pressure, bloating, fullness, early satiety, nausea) and lower GI tract (bloating, pain, incomplete evacuation), somatosensory symptoms related to the musculoskeletal system (pain in neck/shoulders, lower back/hip, muscles/joints), and constitutional functions (sleep, appetite, libido). Analysis was further conducted between the IBS-C and IBS-D patients, controlling for gender and quality of sleep, and using the Bonferroni correction to control for multiple comparisons. RESULTS: Female gender was more prevalent among IBS-C than IBS-D (77% vs 56.1%, p < 0.01), whereas age did not differ (40.2 +/- 1.2 yr vs 39.5 +/- 1.0 yr). Symptoms referred to the upper GI were more prevalent in IBS-C than IBS-D: early satiety (56.7% vs 33.9%, p < 0.004), fullness (63.2% vs 38.5%, p < 0.05), and a trend for upper bloating (80.3 vs 62.6%). IBS-C patients reported higher severity ratings for lower GI bloating (p < 0.001). IBS-C more commonly reported musculoskeletal symptoms (92.2% vs 75.4%, p < 0.001), as well as impairment in sleep (31.3 vs 17.5%, p < 0.009), appetite (35.0% vs 18.4%, p < 0.015) and sexual function (45.2% vs 33.1%, p < 0.0021). There were no differences in SCL-90 and SF-36 scores. CONCLUSIONS: Compared with the IBS-D group, the IBS-C patients show greater prevalence of a wide range of symptoms referred to the upper and lower abdomen, musculoskeletal, and constitutional functions. These findings may be related to differences in autonomic or perceptual responses to visceral and somatic stimuli, and are likely to have implications for treatment responses in the two subgroups. PMID- 10520846 TI - Rectal dialysate and fecal concentrations of neutrophil gelatinase-associated lipocalin, interleukin-8, and tumor necrosis factor-alpha in ulcerative colitis. AB - OBJECTIVE: Neutrophil gelatinase-associated lipocalin (NGAL) is a newly described neutrophil lipocalin that may bind the proinflammatory bacterial tripeptide N formylmethionyl-leucyl-phenylalanine. In situ hybridization and immunohistochemical studies have shown a strong NGAL expression in colonocytes and neutrophils in ulcerative colitis (UC). Because NGAL is highly protease resistant, it should be ideal for in vivo fecal and dialysate studies. Our aim was to investigate the potential of NGAL as a disease activity marker in UC and to compare it with IL-8 and TNF-alpha. METHODS: Twenty-three patients with UC, 14 with Crohn's disease (CD), 19 patients with acute infectious enterocolitis, and 20 healthy controls were included. The disease activity of UC and CD was scored semiquantitatively. Concentrations of NGAL, IL-8, and TNF-alpha were determined in rectal dialysis fluid, feces, and serum using sandwich enzyme-linked immunosorbent assays. The total protein concentration in feces and dialysate fluid was measured, and the amount of markers was expressed as ng/mg protein. RESULTS: In healthy controls and non-IBD (irritable bowel disease) colitis, the median values for NGAL in feces were 183 ng/mg protein and 546 ng/mg protein (p < 0.01), respectively. When separating UC into clinical activity groups (remission, mild/moderate, and severe disease activity) the corresponding values of NGAL were 442 ng/mg (p > 0.05), 605 ng/mg (p < 0.02), and 3646 ng/mg (p < 0.001, compared with controls), respectively, and in quiescent colonic CD 368 ng/mg (p > 0.05) and in active stages 751 ng/mg (p < 0.01). NGAL levels in dialysis fluid listed in the same order were: 11 ng/mg for controls, 71 ng/mg (p > 0.05) for non-IBD colitis, 100 ng/mg (p < 0.02), 179 ng/mg (p < 0.01), and 2053 ng/mg (p < 0.001) for UC, and 14 ng/mg (p > 0.05) and 121 ng/mg (p < 0.02) for CD, respectively. Serum NGAL concentrations did not differ between UC and CD in quiescent versus active stages. A significant increase of NGAL in both feces and dialysate with increasing disease activity of UC was found (p = 0.02 and p = 0.003, respectively). CONCLUSIONS: The NGAL content in rectal dialysate and particularly in feces seems to be a reliable marker for severe disease activity in UC, whereas serum NGAL concentrations do not reflect disease activity. PMID- 10520848 TI - Lactase enzyme, detected immunohistochemically, is lost in active celiac disease, but unaffected by oats challenge. AB - OBJECTIVES: The loss of lactase activity that occurs in active celiac disease resolves on adherence to a gluten-free diet that excludes the cereals wheat, barley, rye, and oats. Recently, an immunohistochemical technique has been described to evaluate lactase expression in primary hypolactasia. We have adapted this method to study lactase activity in adult celiac patients and to assess its value as a diagnostic tool. In addition, given the results of two recent studies suggesting the safety of reintroducing oats cereal into the celiac diet, we have also evaluated the response of lactase expression to oats exposure. METHODS: Duodenal biopsies from 26 patients were stained for lactase expression using an indirect immunoperoxidase method. Eleven disease control patients had normal architecture and nine had features of active celiac disease. Ten patients, who had celiac disease in clinical and histological remission, underwent oats challenge for 12 wk. RESULTS: Confluent expression of lactase was observed in the 11 control patients with normal histology, whereas staining was absent in the nine patients with active celiac disease. All 10 patients with treated celiac disease had normal lactase expression after exposure to oats. CONCLUSIONS: The immunohistochemical technique used in this study provides an easy, reliable method of assessing lactase enzyme and confirms the value of this index as a marker of celiac disease activity. The results of two recent studies demonstrating the lack of oats toxicity in adult celiac patients have been further corroborated by our findings, which show the preservation of lactase enzyme after oats challenge. PMID- 10520849 TI - Mesenteric blood flow is related to disease activity and risk of relapse in Crohn's disease: a prospective follow-up study. AB - OBJECTIVE: The diagnostic significance of increased splanchnic blood flow in Crohn's disease is unclear. This prospective study was therefore undertaken to define the role of Doppler sonography in the assessment of disease activity and in the prediction of early relapse. METHODS: Splanchnic flowmetry was performed in 59 patients with Crohn's disease and 20 healthy volunteers during fasting and 30 min after ingestion of a standardized meal. Twenty-one patients measured during the active state and in clinical remission were followed-up for 6 months. Hemodynamic parameters of the superior and inferior mesenteric arteries and the portal vein were related to clinical (Crohn's disease activity index [CDAI]), laboratory (C-reactive protein), and endoscopic (Crohn's Disease Endoscopic Index of Severity) parameters of disease activity. RESULTS: The postprandial mean velocity of the superior mesenteric artery correlated closest with clinical activity (CDAI, p < 0.005) and C-reactive protein (p < 0.01), but was unrelated to endoscopic activity. All patients in remission after 6 months (9/9) showed an increase in postprandial pulsatility index of the superior mesenteric artery, compared with an initial measurement during active disease (+28%). In contrast, the majority of patients with later relapse or surgery (11/12) had decreased pulsatility index during initial remission (-20%). The positive predictive value of this index for maintenance of remission was 0.82. CONCLUSIONS: Postprandial flow measurements in the superior mesenteric artery are closely related to clinical but not endoscopic disease activity in patients with Crohn's disease. The repeated measurement of the postprandial pulsatility index allows estimation of the risk of recurrence. PMID- 10520850 TI - Assessment of body composition by bioelectrical impedance in adolescent patients with celiac disease. AB - OBJECTIVE: Assessment of body composition is of primary importance in the management of celiac adolescents. We aimed to evaluate body composition by dual energy x-ray absorptiometry and bioelectrical impedance in celiac adolescents on a gluten-free diet to investigate whether impedance may provide an alternative method to assess nutritional status in these patients. METHODS: We studied body composition in 43 adolescents affected by celiac disease on a gluten-free diet for > or = 1 yr and 30 healthy subjects. Fat, fat-free, and bone masses were assessed by dual-energy x-ray absorptiometry. Fat and fat-free masses were also assessed by bioelectrical impedance. All anthropometric measurements were performed according to standard procedures. RESULTS: All patients had a significantly lower body weight, height, fat-free mass, bone mineral density (p < 0.001), and body mass index (p < 0.01) compared with controls. In contrast, parameters predicting fat compartment (sum of skinfolds and fat mass) did not differ from those of controls. No significant difference was found between patients strictly adherent to a gluten-free diet and patients partially compliant. Compared with dual-energy x-ray absorptiometry measurements, bioelectrical impedance showed a high accuracy to estimate fat-free mass (R2 = 0.97) and limited accuracy for fat mass (R2 = 0.75). Furthermore, impedance was more reliable for estimating hydration of soft tissue underlying the fat-free mass changes. CONCLUSIONS: In adolescents with celiac disease, after a mean of 1 yr of gluten-free diet all the parameters assessing body compartments, except fat mass, were affected, compared with healthy controls. Bioelectrical impedance holds promise for routine assessment of body composition changes in celiac adolescents on a gluten-free diet. PMID- 10520851 TI - Intestinal permeability test as a predictor of clinical course in Crohn's disease. AB - OBJECTIVE: The clinical course of Crohn's disease is often unpredictable. The aim of this study was to select the most useful parameters able to predict clinical relapses. METHODS: One hundred-thirty Crohn's disease patients in clinical remission were followed every 4 months for 2 yr or until clinical relapse. Demographic and clinical data were recorded and intestinal permeability (lactulose/mannitol [L/M] test) and biochemical tests (white blood cell count, erythrocyte sedimentation rate, C-reactive protein, alpha1 acid glycoprotein, and serum iron) were performed at study entry. A subgroup of 54 patients had clinical follow-up and repeated tests every 4 months. RESULTS: Fifty-two patients (40%) relapsed during the 2-yr follow-up. A significant correlation was found between relapse and gender (p = 0.030) but not between relapse and age, extent and type of disease, previous surgery, or therapy. Increased L/M test (p = 0.0001) and decreased serum iron level (p = 0.0057) were associated with clinical relapse. Time-dependent analysis, performed on patients receiving serial evaluation, showed that L/M test alteration was the only variable that could predict a relapse (RR 8.84, 95% confidence interval [CI] 1.41-53.37; p < 0.05). CONCLUSIONS: The L/M test identifies Crohn's disease patients in apparent remission, but with a high risk of clinical relapse, better than clinical and biochemical indices. Different treatment strategies might be suggested for this subgroup of patients. PMID- 10520852 TI - Epidemiology of Crohn's disease in Israel: a survey of Israeli kibbutz settlements. AB - OBJECTIVE: The prevalence of Crohn's disease ranges from 10 to 70 cases per 100,000 population, and is 3-8 times more common among Jews. However, this excess risk is not evident in the Jewish population of Israel. Recently we have described a significant increase in the prevalence and incidence of Crohn's disease in the south of Israel. The aim of this study was to confirm this trend in a stable population found in communal (kibbutz) settlements. METHODS: We repeated a community-based survey in 124,400 kibbutz residents, 10 yr after our first study. This population represents 5% of the Jewish population of Israel. All Crohn's disease patients were located by contacting the kibbutz clinics of the 269 kibbutz settlements (100% compliance). Data was updated to December 31st, 1997, which was designated the point prevalence date, and included information on gender, age, origin, education, profession, extent of the inflammatory process, clinical spectrum of the disease, therapy, complications of the disease, and treatment. The average annual incidence for the 10 yr was calculated from the prevalence data. Only cases with a definite diagnosis of Crohn's disease made in a recognized gastroenterology unit were accepted into the study. RESULTS: There were 81 confirmed cases of Crohn's disease and the prevalence rate rose from 25.5/100,000 in 1987 to 65.1/100,000 in 1997 (p < 0.001). The mean annual incidence rate for this period (1987-1997) is 5.0/100,000/yr. Prevalence rates were higher in women than men, and in those born in Israel or Europe/America than in Asia/Africa. The mean age at presentation of the disease was lower in 1997 than in 1987, 37.4 +/- 17.0 and 45.0 +/- 17.0 yr, respectively (p = 0.041). Prevalence was highest in men with > 16 yr of education, and in women with 11-12 yr education, 119.7 and 100.3/100,000, respectively. CONCLUSIONS: During the decade 1987-1997, the prevalence of Crohn's disease has increased in Israel and is approaching the rates in Europe and America. PMID- 10520853 TI - The longitudinal high-pressure zone profile in patients with fecal incontinence. AB - OBJECTIVE: The aim of this study was to evaluate differences in the longitudinal high-pressure zone (HPZ) profile between incontinent patients and healthy controls. METHODS: One hundred and fifty-six patients with fecal incontinence (mean age 63 yr; 139 women, 17 men) and 25 healthy controls (mean age 54 yr; 20 women, five men) underwent anorectal manometry with a perfused catheter using a station pull-through technique. RESULTS: Maximum resting pressure (MRP) and maximum squeeze pressure (MSP) were lower in patients (p < 0.001) and the HPZ was shorter in patients (p < 0.05). The ratio MRP/sum of resting pressures within HPZ did not differ between the groups. The severity of incontinence measured as an incontinence score was correlated to MSP (p < 0.05) and sum of squeeze pressure within HPZ (p < 0.05), but not to any other variable. The relative pressure amount during rest in the distal half of HPZ was higher in controls (63% vs 56%, p < 0.05), but this was not seen during squeeze. CONCLUSIONS: These results suggest that the main difference between incontinent patients and healthy controls is a greater magnitude of the pressure profile in the latter group. Furthermore, the relative pressure accumulation during rest tended to be more distally located within the HPZ. PMID- 10520854 TI - Unsuspected infection is infrequent in asymptomatic outpatients with refractory ascites undergoing therapeutic paracentesis. AB - OBJECTIVE: Large-volume paracentesis is a safe and effective means of treating patients with refractory ascites. However, there is limited information regarding the need for ascitic fluid studies in asymptomatic outpatients presenting for therapeutic paracentesis. The aim of this prospective study was to define the incidence and natural history of peritoneal fluid infection in asymptomatic outpatients undergoing therapeutic paracentesis. METHODS: Over a 13-month period, 118 therapeutic paracenteses were performed in 29 outpatients with decompensated cirrhosis (Child-Pugh class B = 38%, C = 62%). After a brief medical history and physical examination, ascitic fluid cell count with differential and culture were obtained from all participating subjects. Seven (24%) of the subjects were receiving norfloxacin prophylaxis, accounting for antibiotic coverage during 40% of the procedures performed. The clinical course and outcome of study subjects during a mean follow-up of 137 days was reviewed. RESULTS: All 118 (100%) of the ascitic fluid samples demonstrated absolute neutrophil counts of <250/mm3 (mean = 6.5 +/- 22.5 pmn/mm3). Asymptomatic bacterascites was identified from three of the 118 (2.5%) fluid samples, but all of these subjects spontaneously recovered without treatment or sequelae. During follow-up, six episodes of symptomatic or hospital-associated peritoneal fluid infection were identified in study participants, emphasizing the importance of fluid studies in other clinical settings. CONCLUSIONS: Although further studies are needed, the routine culture of ascitic fluid in asymptomatic outpatients with refractory ascites requiring therapeutic paracentesis may not be necessary when there is a low index of suspicion for occult infection. In circumstances of clinical uncertainty, however, obtaining ascitic fluid cell counts with differential is recommended to insure patient safety. PMID- 10520855 TI - Disseminated intravascular coagulation in liver cirrhosis: fact or fiction? AB - OBJECTIVE: Cirrhosis is commonly associated with haemostatic dysfunction. The similarities of laboratory tests of disseminated intravascular coagulation (DIC) to those found in cirrhosis has led to the belief that DIC is a feature of the haemostatic failure of cirrhosis. METHODS: The aim of this study was to determine whether DIC is part of the coagulopathy of cirrhosis by applying quantitative tests for prothrombin fragment 1 + 2, antithrombin III, thrombin-antithrombin complex, and specific fribrinogen degradation products levels (XDP), as well as the thrombelastograph for detecting the Clot Lysis Index. RESULTS: Fifty-two stable cirrhotic patients (33 men, 19 women; mean age, 58.8 yr; range, 24-72 yr) with differing etiologies were studied. On tests of thrombin generation: thrombin antithrombin complexes, fibrin(ogen) degradation products, and prothrombin fragments 1 + 2 were not found to be significantly different from an age- and gender-matched control group (p = 0.18, 0.3, and 0.67, respectively), whereas albumin, Factor V, fibrinogen, antithrombin III, and alpha2-antiplasmin were all significantly low (p = 0.0004, 0.002, 0.06, 0.004, and 0.004, respectively), reflecting reduced synthetic function and correlation in ascitic and non-ascitic patients. There was no correlation between impaired synthesis (antithrombin III and alpha2-antiplasmin) and indices of DIC (prothrombin fragment 1 + 2, thrombin antithrombin complexes, and XDP) (p = not significant). The percentage of patients with high prothrombin fragments 1 + 2 and thrombin antithrombin levels in each Child grade group was similar. Thrombin time was significantly elevated in the cirrhotic group (a manifestation of low fibrinogen levels). The Clot Lysis Index as measured by thrombelastography was significantly abnormal, indicating mild hyperfibrinolysis. CONCLUSION: We conclude that DIC is not part of the coagulopathy in stable liver cirrhosis without recent complications. PMID- 10520856 TI - Anticardiolipin antibodies in patients with liver disease. AB - OBJECTIVE: Our aim was to test the hypothesis that anticardiolipin antibodies (aCL) may cause an antiphospholipid syndrome and thrombotic events in patients with liver disease. METHODS: aCL were measured in 116 healthy controls and 372 patients with liver disease of different stage and etiology: 136 cases secondary to hepatitis C virus (HCV) infection, 139 due to hepatitis B virus (HBV) infection, 69 with alcoholic liver damage, and 28 cryptogenic in origin. Prior thrombotic events were recorded. The results were related to age, gender, stage, severity, and etiology of the liver disease, as well as to the occurrence of organ- and nonorgan-specific autoantibodies. RESULTS: aCL were positive in 4.4% of controls and in 18.8% of patients (p < 0.0002). Patients with aCL were more frequently men with an advanced cirrhosis and simultaneous occurrence of anti smooth-muscle antibodies (ASMA) in serum (p < 0.0006); their liver damage was often secondary to HBV (37.3%) or alcohol abuse (18.5%). At conditional logistic regression analysis, only the presence of ASMA (odds ratio [OR] = 3.02, 95% confidence interval [CI] 1.7-5.5, p = 0.0003), HBV (OR = 3.4, 95% CI 1.6-7.2, p = 0.0013), or alcoholic liver disease (OR = 5.3, 95% CI 2.3-12.2, p = 0.0001) were independently associated with aCL. Thrombosis was encountered in 24 patients (6.4%). At conditional logistic regression analysis, thrombosis was significantly associated with advanced age (OR = 1.07, 95% CI 1.0-1.1, p = 0.0094), development of hepatocellular carcinoma (OR = 17.8, 95% CI 1.6-196.0, p = 0.01), HBV etiology (OR = 6.3, 95% CI, 1.6-24.6, p = 0.0076), or cryptogenic liver disease (OR = 54.8, 95% CI 5-599.9, p = 0.001). Of the five patients with newly documented portal thrombosis during the follow-up, only one tested positive for aCL. CONCLUSIONS: In patients with nonautoimmune liver disease, aCL production is an epiphenomenon of the liver damage and is not associated with thrombotic complications. These data do not support the hypothesis that HCV is a cause of the antiphospholipid syndrome. PMID- 10520857 TI - Screening for hepatocellular carcinoma in patients with advanced cirrhosis. AB - OBJECTIVE: Most available data on screening for hepatocellular carcinoma (HCC) in patients with cirrhosis originate from Asia and Europe. These data may not be applicable to patients from the United States because of geographic variation in the underlying etiology and other factors. Our aim was to assess the risk of HCC in U.S. patients with cirrhosis undergoing standardized screening. METHODS: All cirrhotic patients evaluated for liver transplantation at our institution from January 1, 1994-December 31, 1997 were included in this study. The screening strategy included initial screening, which was offered to all patients and consisted of alpha-fetoprotein (AFP), abdominal ultrasound, and computed tomography (CT) scan, and extended screening, which was performed only on transplant-eligible patients and consisted of semiannual AFP and ultrasound. RESULTS: During the study period, 285 patients with cirrhosis were evaluated for transplantation and underwent initial screening. Of these, 166 were eligible for transplantation and underwent extended screening during a median follow-up of 15 months (range 6-42 months). Twenty-seven HCC were found, 22 during initial screening and five during extended screening. The cancer-free proportions of the cohort who underwent extended screening at 1, 2, and 3.5 yr were 98.6% +/- 1.4%, 96.4 +/- 1.8%, and 77.1% +/- 1.7%, respectively (mean +/- SE). Hepatitis C, either alone or in part, was the etiology in 63% of patients with HCC. The sensitivity of CT scan (88%) was significantly higher than AFP >20 ng/ml (62%) and ultrasound (59%) for detecting HCC (p < 0.001). CONCLUSIONS: In patients with established cirrhosis, the risk of detecting HCC is maximal at the baseline screening (7%). Hepatitis C was the most common etiology for cirrhosis in study. In U.S. patients with established cirrhosis, CT scan exhibited higher sensitivity for detecting HCC than ultrasound or AFP. PMID- 10520858 TI - Prevalence of hypoxemia in 102 Japanese patients with alcoholic and nonalcoholic cirrhosis. AB - OBJECTIVE: Liver cirrhosis is often accompanied by arterial hypoxemia in the absence of cardiopulmonary disease. The aim of this study was to investigate the relationship between various clinicopathological conditions and the hypoxemia seen in Japanese patients with liver cirrhosis. METHODS: In 102 consecutive patients with alcoholic (N = 45) and nonalcoholic (N = 57) cirrhosis not associated with cardiopulmonary disease, we performed lung perfusion scintigraphy, contrast echocardiography, and arterial blood gas analysis and measured oxygen consumption. RESULTS: No abnormality was seen in pulmonary blood flow in cirrhotic patients, but 38 (38%) of them had a decreased partial pressure of oxygen (PaO2). The hypoxemic patients did not show any pulmonary signs or symptoms. The hypoxemia was not associated with the Child-Pugh grade, and was observed in 32 (71%) of the 45 alcoholic patients but in only six (11%) of the 57 nonalcoholic patients (p < 0.001). Oxygen consumption was significantly higher in the alcoholic group than in the nonalcoholic group (p < 0.0001), and a high incidence of oxygen consumption was seen in all 45 (100%) of the alcoholic patients and in 34 (60%) of the nonalcoholic subjects, the difference being significant (p < 0.01). The relationship between oxygen consumption and PaO2 in the 102 cirrhotic patients showed an inverse correlation (r = -0.85, p < 0.0001). Among the alcoholic patients, the incidence of hypoxemia did not differ between the 33 smokers and the 12 nonsmokers. After 1 wk of abstinence from alcohol a significant increase (p < 0.0001) in the PaO2 was seen in 14 of 19 patients with alcoholic cirrhosis. CONCLUSIONS: We conclude that the hypoxemia in Japanese patients with liver cirrhosis occurs mainly in drinking alcoholic patients, presumably due to an increased oxygen consumption by alcohol. PMID- 10520859 TI - A pilot study on the hemodynamic effect of short-term ursodeoxycholic acid therapy in patients with stable liver cirrhosis. AB - OBJECTIVE: Total serum bile acid concentrations are elevated in individuals with liver disease. Ursodeoxycholic acid (UDCA) therapy in such patients results in a further significant rise in plasma levels to the extent that it becomes the major circulating bile acid. In laboratory animals, bile acids, such as taurocholic acid, have also been shown to possess a diuretic-like action, as they can promote diuresis, natriuresis, and kaliuresis by inhibiting tubular sodium reabsorption. The aim of the present study was to assess the effect of 1 month's UDCA therapy on cardiovascular function in cirrhotic patients. METHODS: Two groups of patients with cirrhosis were studied, six with primary biliary cirrhosis (PBC) and six with postnecrotic liver cirrhosis (PNC). Cardiovascular function was assessed by determination of blood pressure, heart rate, and by two-dimensional and pulsed Doppler echocardiography. RESULTS: In PBC patients, 1 month's treatment with UDCA significantly reduced diastolic volume without changing systolic, diastolic, and mean blood pressures, heart rate, systolic and stroke volumes, ejection fraction, cardiac output, and systemic vascular resistance. In PNC patients, UDCA significantly reduced cardiac output, with a tendency to reduce left ventricular volumes, without any changes in systolic, diastolic, and mean blood pressures. CONCLUSIONS: UDCA caused reductions in diastolic volume in the PBC patients and cardiac output in the PNC patients. Such reductions are not unlike that seen in individuals treated with diuretics. This diuretic-like action deserves further study, particularly in cirrhotic patients who are also being treated with diuretics or show evidence of cardiac myopathy. PMID- 10520860 TI - Gallbladder hyporesponsiveness to an exogenous nitric oxide donor, glyceryl trinitrate, in patients with advanced liver cirrhosis. AB - OBJECTIVE: An increased production of nitric oxide (NO) in liver cirrhosis has been documented. NO could intervene in regulating gallbladder contraction, as suggested by clinical and experimental studies. Our aim was to investigate the influence of an NO donor on gallbladder motility in cirrhotic patients in relation to the severity of liver cirrhosis. METHODS: The subjects were six controls and 18 patients with liver cirrhosis (six in each Child class). Gallbladder emptying was monitored by ultrasound for 90 min after a mixed meal (14 g fat, 425 kcal). Fasting gallbladder volume, minimal residual volume, ejection fraction, area under emptying curve, and half contraction time of the gallbladder were assessed at 15-min intervals. The patients were evaluated on two consecutive days, with or without perlingual administration of 0.5 mg glyceryl trinitrate (GTN). Statistical analysis was performed by the two-tailed Student's t test and Pearson's correlation coefficient. RESULTS: GTN significantly reduced gallbladder motility in controls and compensated cirrhotics (p < 0.02), but had no effect upon gallbladder emptying in Child class B and C cirrhotics. CONCLUSIONS: Gallbladder hypocontractility in liver cirrhosis is related to the severity of the disease. This study is the first to show that GTN has no effect upon gallbladder motility in advanced liver cirrhosis when administered in doses that induce relaxation in controls and compensated cirrhosis. PMID- 10520861 TI - Prospective evaluation of unexplained chronic liver transaminase abnormalities in asymptomatic and symptomatic patients. AB - OBJECTIVES: It is currently recommend to perform a liver biopsy for patients with chronically elevated liver function tests (LFT) of unknown etiology (marker negative). The necessity and benefits of these recommendations are unknown. The aims of this study were to determine the prevalence of marker-negative LFT in patients referred for evaluation of chronically elevated LFT; to determine the prevalence of diseases that may be associated with marker-negative abnormal LFT; and to assess whether a liver biopsy alters the management of such patients. METHODS: We conducted a prospective observational study of 1124 adults referred for evaluation of chronically elevated LFT. Patients who consented to a liver biopsy were eligible. Marker-negative abnormal LFT was defined as the absence of accepted serum markers for infectious, metabolic, autoimmune, or hereditary liver disease, the absence of a history of alcohol or hepatotoxic drug use, and the absence of signs of chronic liver disease. RESULTS: Eighty-one of 1124 eligible patients were marker-negative. Liver biopsies in the 81 marker-negative patients revealed: normal histology (eight), steatosis (41), steatohepatitis (26), fibrosis (four), and cirrhosis (two). All 73 abnormal liver biopsies had some degree of steatosis. There were no significant associations between histological findings and the presence of obesity (p = 0.13), hyperlipidemia (p = 0.4), or diabetes (p = 0.9). There were no significant associations when classifying patients by gender or by symptoms. CONCLUSION: In the setting of marker-negative elevated LFT, the most likely histological diagnosis is fatty metamorphosis of the liver with occasional associated fibrosis. PMID- 10520862 TI - Molecular epidemiology of hepatitis C infection in U.S. veteran liver transplant recipients: evidence for decreasing relative prevalence of genotype 1B. AB - OBJECTIVE: The U.S. Veteran population represents a unique patient group to study different HCV genotypes because of geographically diverse exposures. The aim of this study was to characterize the distribution of HCV genotypes in U.S. veterans undergoing liver transplantation (OLT), trace genotypes to modes of acquisition (risk behavior and location), and evaluate the relative prevalence of HCV genotypes according to the time of acquisition. METHODS: Between 10/88 and 12/95, 110 primary OLTs were performed in U.S. Veterans at our center. Forty-nine (45%) patients had detectable HCV-RNA by PCR at the time of OLT. Determination of HCV genotypes was performed by restriction fragment length polymorphism of the 5' noncoding region and classified according to Simmonds et al. RESULTS: Twenty three of 49 (47%) veterans had 1a, 17 (35%) 1b, two (4%) 2a, three (6%) 2b, two (4%) 3a, two (4%) mixed (1a/2a, 1b/2a). This distribution of HCV genotypes was comparable to the genotypic distribution of a contemporary cohort of nonveteran OLT recipients at the University of Washington. There was a statistically significant association between illicit injection drug use (IDU) and 1a, with 63% of 1a patients having IDU whereas only 14% of 1b patients admitted to IDU (p = 0.03). All patients in whom the mode of acquisition was unknown had genotype 1b (p = 0.04). Intranasal cocaine use was strongly correlated with IDU (p = 0.002). Patients who had tattoos but no history of blood transfusion (BT) or recreational drug use had genotype 1 (2 had 1a, 2 had 1b; p = NS). Twenty-two (45%) patients had serological evidence of prior hepatitis B (HBV) infection. Patients who had genotype 2a, 2b, 3a, or mixed were much more likely to have had HBV (seven of nine, 78%) than patients with genotype 1a or 1b (15 of 40, 37.5%) (p = 0.03). There was no significant correlation between BT, dates, or military branch of service, high risk behavior in Southeast Asia, level of education, ethnicity, and particular genotype(s). Whereas the proportion of 1b accounting for HCV infection in patients with a first exposure before 1968 was 50%, all patients with a first exposure post-1975 were non-1b (p = 0.04), suggesting a change in the epidemiology of HCV in our cohort. CONCLUSIONS: In U.S. Veterans undergoing OLT: 1) 45% had PCR-confirmed HCV infection, 2) 1a was the predominant genotype and was associated with IDU, and 3) a significant decrease in the prevalence of genotype 1b from the pre-Vietnam era to post-1975 suggests a changing epidemiology of HCV genotypes. PMID- 10520863 TI - Clinical significance of TT virus infection in patients with chronic hepatitis C. AB - OBJECTIVE: The TT virus (TTV) is a novel DNA virus that has recently been identified. The clinical significance of TTV infection in patients with chronic hepatitis C has not been determined. The aim of this study was to determine the prevalence and possible role of TTV in a well characterized population with chronic hepatitis C infection. METHODS: Ninety patients with chronic HCV and known time of HCV acquisition were selected from approximately 250 patients followed at our institution. Characteristics including age, sex, histology, and length of disease were recorded. Direct sequencing of the NS5 region was used for HCV genotyping. TTV DNA detection was based on PCR. RESULTS: TTV infection was present in 24 of 90 (27%) HCV patients. Patients were divided into four groups based on stage of disease; chronic hepatitis (CH, 29 patients), compensated cirrhosis (CC, 17 patients), decompensated cirrhosis (DC, 28 patients), and hepatocellular carcinoma (HCC, 16 patients). TTV was present in 2/29 (7%), 2/17 (12%), 11/28 (39%), and 9/16 (56%) in those with CAH, CC, DC, and HCC respectively. TTV was significantly more prevalent among those with advanced disease (DC and HCC) compared to those with stable disease (CH and CC; p = 0.001). Mean age, sex, and the time from exposure to HCV to development of complications were similar in TTV-positive and -negative patients. TTV infection was more common in patients infected with HCV genotype 1b. Univariate analysis showed that length of HCV infection, HCV genotype 1b, and TTV infection were important in predicting the stage of liver disease in HCV patients. However, after adjusting for length of HCV infection, TTV but not HCV genotype was important in predicting the stage of liver disease. CONCLUSIONS: We conclude that 1) TTV infection is common in patients with chronic HCV; 2) TTV infection is more prevalent among patients with advanced HCV-associated liver disease (DC and HCC) than in those with stable disease (CH and CC); and 3) TTV infection is more common in patients with HCV genotype 1b but is independent from genotype in predicting the stage of HCV-associated liver disease. PMID- 10520864 TI - Usefulness of measurement of Lens culinaris agglutinin-reactive fraction of alpha fetoprotein as a marker of prognosis and recurrence of small hepatocellular carcinoma. AB - OBJECTIVE: Lens culinaris agglutinin-reactive fraction of alpha-fetoprotein (AFP L3%) is a recently described marker of hepatocellular carcinoma (HCC), and its usefulness has been demonstrated in many studies. We evaluated the usefulness of serial measurement of AFP-L3% as a marker of prognosis and recurrence after treatment of small HCC. METHODS: AFP-L3% was measured before and after initial treatment in 60 patients with small HCC (maximum diameter < or = 2 cm). AFP-L3% was taken as the ratio of AFP-L3 to total AFP and multiplied by 100%, and levels > or = 10% were considered positive. Outcomes and recurrence were compared between patients AFP-L3%-negative after initial treatment (Group A, n = 43) and patients who were AFP-L3%-positive after initial treatment (Group B, n = 17). RESULTS: Before treatment, AFP-L3% was positive in 14 (23.3%) of the 60 patients. The cumulative survival rate of Group A was significantly longer (p = 0.0091) than that of Group B. The recurrence rate was significantly higher in Group B (p = 0.0104) than in Group A. When recurrence was limited to intrahepatic metastasis, the recurrence rate was significantly higher in Group B (p = 0.0064). However, the recurrence rate of multicentric occurrence did not differ significantly between Groups A and B. CONCLUSIONS: Measurement of AFP-L3% after treatment may be useful for understanding prognosis and recurrence of HCC. PMID- 10520865 TI - Marked differences in the frequency of microsatellite instability in gastric cancer from different countries. AB - OBJECTIVE: Previous studies have reported variable rates of microsatellite instability (MSI) in gastric cancer. We investigated the frequency of MSI in invasive gastric carcinoma of patients from three geographic regions. METHODS: Genomic DNA from gastric cancer and nontumor tissue from 22 Korean, 20 Colombian, and 26 U.S. patients was amplified with five microsatellite markers. RESULTS: MSI was more frequently seen in gastric cancer from Korea, affecting 50% of patients, in contrast with gastric cancers from the U.S. (7%) and Colombia (15%) (p = 0.003 and p = 0.03, respectively). MSI at one locus was significantly more frequent in gastric cancer from individuals >65 yr (p = 0.01). MSI was similarly associated with both diffuse and intestinal types of gastric cancer. CONCLUSIONS: MSI affects the two major histological types of gastric cancer, and was more frequent in gastric cancer from Korea than in the other countries, suggesting that the relative importance of different pathways of gastric carcinogenesis may vary in diverse regions of the world. PMID- 10520866 TI - How does colorectal cancer present? Symptoms, duration, and clues to location. AB - OBJECTIVE: Most colorectal cancers still present with symptoms because screening, although effective, is not yet widely practiced. A careful history and physical examination are still the usual methods for suspecting colorectal cancer and ordering appropriate investigation. Therefore, we studied the symptoms, duration, and clues to location of colorectal cancer. METHODS: We reviewed both hospital and office records for 204 consecutive patients with colorectal cancer, first diagnosed after symptoms, at one regional referral center from 1983-87. We abstracted data on demographic characteristics, presence and duration of 15 symptoms, and characteristics of the tumors. RESULTS: The 194 patients included in the study were similar to those with colorectal cancer described elsewhere in terms of age, gender, and tumor location (58% distal to the splenic flexure), and stage (56% stage A or B). The most common symptoms were rectal bleeding (58%), abdominal pain (52%), and change in bowel habits (51%); the majority had anemia (57%) and occult bleeding (77%). The median duration of symptoms (from onset to diagnosis) was 14 wk (interquartile range 5-43). We found no association between overall duration of symptoms and the stage of the tumor. Patient age, gender, and proximal cancer location were also not associated with a longer duration of symptoms before diagnosis. We developed a rule for predicting a distal location of cancer using multiple logistic regression. Independent predictors were (odds ratio [95% CI]): Hb (1.34 for each g/dl [1.16-1.54]); rectal bleeding (3.45 [1.71 6.95]); constipation (3.16 [1.38-7.24]); and proximal symptoms (at least one of anorexia, nausea, vomiting, abdominal pain, or fatigue) (0.48 [0.20-1.02]). The rule had sensitivity of 93% and a specificity of 47%, with an area under the ROC curve of 0.79. CONCLUSIONS: Until prevention of colorectal cancer is more common, we must continue to rely on clinical findings for detecting this cancer. Our results will remind physicians to keep colorectal cancer on the differential diagnosis of "chronic" gastrointestinal symptoms, and our decision rule may prompt earlier investigation with colonoscopy. PMID- 10520867 TI - A look back. The microscope. PMID- 10520868 TI - Heterotopic gastric mucosa in the upper esophagus ("inlet patch"): a rare cause of esophageal perforation. AB - We report the case of a 21-yr-old woman who presented with a perforation of an upper esophageal ulcer on a patch of gastric-type mucosa. Despite surgical closure of the perforation and reinforcement with a pleuro-muscular flap the patient developed an esophageal leakage and died in the postoperative period. Heterotopic gastric mucosa in the upper esophagus is usually an asymptomatic abnormality, discovered incidentally during endoscopic studies carried out for some other reason; however, complications secondary to the inlet patch acid secreting capacity can arise, and this has to be kept in mind to elude life threatening conditions. PMID- 10520869 TI - Hemobilia, a rare cause of acute pancreatitis after percutaneous liver biopsy: diagnosis and treatment by endoscopic retrograde cholangiopancreatography. AB - We here report the case history of a 75-yr-old woman who developed pancreatitis and recurrent symptomatic, cholestasis-induced hemobilia after percutaneous liver biopsy. An endoscopic sphincterotomy with clot extraction led to relief of symptoms. The risk of hemobilia after percutaneous liver biopsy is less than one per 1000 procedures, and only two cases of acute pancreatitis after percutaneous liver biopsy have previously been reported. To our knowledge, this is the first case in which endoscopic retrograde cholangiopancreatography was used to both diagnostic and therapeutic ends. PMID- 10520870 TI - Carcinosarcoma of the ampulla of Vater: a case report with immunohistochemical and ultrastructural studies. AB - Carcinosarcoma of the duodenum has not been reported previously, although this type of tumor has been detected in other organs. We present here a case of carcinosarcoma of the duodenum, including immunohistochemical and electron microscopical findings. An ulcerating tumor, located in the duodenal ampullary region, contained two divergent components: ordinary differentiated tubular adenocarcinoma, and sarcomatoid tissue composed of spindle tumor cells. Immunohistochemically, the adenocarcinoma cells were stained with antibodies against epithelial markers including keratin and CA19-9. In contrast, the sarcomatoid tissue was strongly positive for vimentin and was focally positive for myoglobin, keratin, and CA19-9. We speculate that the sarcomatoid element of the carcinosarcoma arose from part of the ordinary adenocarcinoma tissue. PMID- 10520871 TI - Is "negative" positive? PMID- 10520872 TI - A lot of heartburn, a little cancer. PMID- 10520873 TI - Molecular detection of micrometastases: science on stage. PMID- 10520874 TI - Glucocorticosteroids are not effective in alcoholic hepatitis. PMID- 10520875 TI - Corticosteroids are effective in patients with severe alcoholic hepatitis. PMID- 10520876 TI - Re: Richter et al.: possibly protective properties of Helicobacter pylori in connection with GERD. PMID- 10520877 TI - Acute abdomen due to perforation of colon by ingested chicken bone: diagnosis and endoscopic treatment. PMID- 10520878 TI - Adrenal gland sampling during transjugular liver biopsy leads to misdiagnosis of hepatocellular carcinoma. PMID- 10520879 TI - Iron deficiency and dysphagia. PMID- 10520880 TI - Low efficacy of ranitidine bismuth citrate plus clarithromycin combination on Helicobacter pylori in a Turkish population. PMID- 10520882 TI - How long for the routine Helicobacter pylori antimicrobial susceptibility testing? The usefulness of the string test to obtain Helicobacter for culture. PMID- 10520881 TI - Soluble interleukin-6 receptor levels in liver cirrhosis. PMID- 10520883 TI - Misdiagnosed anorectal varices resulting in a fatal event. PMID- 10520884 TI - Re: Compliance curves during peritoneal dialysate infusion. PMID- 10520885 TI - Incidence, risk factors, and clinical significance of neurological symptoms in adult patients with acute gastroenteritis. PMID- 10520886 TI - IgA-tissue transglutaminase (tTG)-antibodies are highly sensitive serum markers for celiac disease. PMID- 10520887 TI - Manometry for chest pain. PMID- 10520888 TI - Vaginal variceal hemorrhage in a patient with primary biliary cirrhosis: a case successfully treated by balloon-occluded retrograde transvenous obliteration. PMID- 10520889 TI - Is Helicobacter pylori-negative duodenal ulcer masked by the high prevalence of H. pylori infection in the general population? PMID- 10520890 TI - Iron-deficiency anemia in premenopausal women: why not consider atrophic body gastritis and Helicobacter pylori role? PMID- 10520891 TI - Ashkenazi Jews and colon cancer. PMID- 10520892 TI - Re: van den Boogert et al.: current endoscopic modalities for ablating Barrett's esophagus with high grade dysplasia. PMID- 10520893 TI - Multiple anomalies of pancreaticobiliary ductal system: report of a case. PMID- 10520894 TI - Interferon plus ribavirin: a cautionary note. PMID- 10520895 TI - Cirrhosis and diabetes: C, the difference. PMID- 10520896 TI - A rheumatoid factor specific mimotope identified by a peptide display library. AB - We screened a 10 amino acid peptide display library in filamentous phage with B'20, a monoclonal high affinity IgM rheumatoid factor (RF) expressing the VkIIIa dependent 4C9 idiotype. Using direct and indirect selection techniques, 12 B'20 reactive peptides were identified, 9 of which belonged to one of two motifs. Binding of B'20 to phage-bearing peptides was inhibited by both IgG and 4C9 antiidiotype. Synthetic peptides corresponding to the two motifs inhibited the Fc binding of a low avidity IgA B'20 construct. Purified IgM from 6/8 RF-positive RA patients and 8/11 monoclonal RFs with VkIII-encoded light chains bound to the phage, whereas none of the four monoclonal RFs with VkI or VkII encoded light chains bound. Phage binding appeared to be RF specific. Three 4C9 positive/RF negative cell lines from RA patients did not bind to phage nor did three B'20 mutants that had lost RF specificity, whereas two mutants that retained RF specificity also retained phage binding. We propose that there is a common epitope(s) recognized by VkIII encoded RFs that is mimicked by the structure of these peptides. Such mimotopes might be exploited to design novel agents that interfere with autoantibody binding. PMID- 10520897 TI - Lumbosacral acute demyelinating polyneuropathy following hepatitis B vaccination. AB - We report a patient who presented with an acute inflammatory demyelinating polyneuropathy, that followed the second injection of a hepatitis B vaccination, and characterized by motor and sensory deficit restricted to lower limbs and perineum, and persistent bladder dysfunction. The relationship between the preceding event and neurological disease is discussed. PMID- 10520898 TI - Autoantibody to the liver arginase present in sera of patients with autoimmune hepatitis and chronic hepatitis. AB - We have reported previously that immunization of rat liver arginase in rabbits induced autoantibody that is reactive with their own liver arginase and has cytotoxic activity to their hepatocytes. This promoted us to investigate whether or not such an autoantibody is present in sera of patients with certain hepatic disorders, since the liver arginase is dominant in the liver and highly homologous in structure among ureotelic animals. By Western blot analysis, sera from patients with chronic hepatitis and autoimmune hepatitis were shown to have an autoantibody reacting with purified human liver arginase. Since the autoantibody was also reactive with liver arginase of rat origin to almost the same extent as that of human origin, ELISA with rat liver arginase as a coating antigen was developed and used for the quantification of the autoantibody. Prominent increase of the anti-liver arginase autoantibody was found in autoimmune hepatitis, moderate increase in chronic liver diseases, and no increase in acute hepatitis or normal controls by the ELISA. These results suggest that the increased anti-liver arginase autoantibody might involve in some parts in the pathophysiology of the hepatitic disorders. Assay of the autoantibody can also be utilized as a marker for the differentiation of certain hepatitis. PMID- 10520899 TI - Analysis of autoantibody epitopes on human thyroid peroxidase. AB - A number of studies have indicated that the major autoantibody epitopes on human thyroid peroxidase (TPO) are conformational and are formed by two overlapping immunodominant regions on the TPO molecule. In order to investigate further autoantibody reactivity with TPO, we have studied the TPO binding characteristics of sera from patients with autoimmune thyroid disease (n = 20), autoimmune adrenal disease (Addison's disease; n = 8) and apparently healthy blood donors (n = 9) using recombinant TPO expressed with a series of truncations and internal deletions. This material was obtained using an in vitro transcription/translation system in the presence of 35S-methionine and the reactivity of TPO autoantibodies tested in an immunoprecipitation assay. In addition, we have studied the effects of denaturing purified recombinant TPO by reduction and/or sodium dodecyl sulphate on its reactivity with TPO autoantibodies by Western blotting analysis. These studies show that TPO autoantibodies can recognise TPO in Western blotting analysis when large amounts of purified TPO are run on the gels and the blotted proteins renatured prior to addition of antibody. Under these conditions TPO autoantibodies in all 20 Graves' or Hashimoto's sera tested reacted strongly with blots of non-reduced TPO but reduction of TPO had a marked effect on the ability of autoantibodies to recognise it in Western blotting analysis. Analysis of TPO autoantibody binding to 35S-labelled TPO proteins containing N-terminal, central or C-terminal deletions indicated that all modifications studied caused a statistically significant lowering of binding. In the case of some modifications, there were differences in the reactivity of TPO autoantibodies in sera from patients with Addison's disease compared to TPO autoantibodies in autoimmune thyroid disease and/or healthy blood donor sera. Overall, our results of analysis of T PO autoantibody binding in Western blotting and with modified TPO proteins in immunoprecipitation assays suggest that the main autoantibody binding sites on the TPO molecule involve extensive amino acid sequences. Our studies also suggest that TPO autoantibodies from patients with autoimmune thyroid disease, Addison's disease and apparently healthy blood donors show some differences in epitope recognition on TPO and this approach may allow differentiation between disease related and unrelated TPO autoantibodies. PMID- 10520900 TI - Cytokines and apoptotic molecules in experimental melanin-protein induced uveitis (EMIU) and experimental autoimmune uveoretinitis (EAU). AB - The cytokine profile and occurrence of apoptosis during experimental melanin protein induced uveitis (EMIU) were investigated and compared with that of experimental autoimmune uveoretinitis (EAU). EMIU or EAU was induced in Lewis rats. Eyes were collected at different time points after immunization. Cytokine mRNA expression was identified in the inflammatory cells in the uvea of EMIU rats; IL-2, IFN-gamma and IL-12 increased at the peak of the inflammation, and then tapered off as inflammation subsided. IL-4 and IL-10 increased at the peak of ocular inflammation, and persisted with inflammation resolved. Fas and FasL were expressed consistently in ocular resident cells of EMIU, but were elevated in EAU. In EAU, Bcl-2 expression showed a sharp peak in inflammatory cells but not in the resident cells. In EMIU, high levels of Bcl-2 were present and persisted in both ocular resident and inflammatory cells. Expression of Bax was relatively stable in both EAU and EMIU. Cellular DNA fragmentation was detected in the retinal glial cells of EAU and some inflammatory cells of EMIU. In EMIU, the dynamics of Th1 cytokines were consistent with the ocular inflammation, whereas persistent expression of Th2 cytokines was consistent with their known regulatory role. The continuous high expression of Bcl-2 and the high ratio of Bcl-2 to Bax in the eyes of EMIU may possibly contribute to prevention of ocular tissue damage, and of inflammatory cells from undergoing apoptosis, thus resulting in chronic recurrent inflammation. PMID- 10520901 TI - HLA class II restriction of autoreactive T cell responses in pemphigus vulgaris: review of the literature and potential applications for the development of a specific immunotherapy. AB - Pemphigus vulgaris (PV) is a life-threatening autoimmune bullous disease of the skin and mucous membranes which requires immunosuppressive therapy, most commonly a combination of glucocorticoids and additional immunosuppressive agents. Since the side effects of long-term immunosuppressive therapy contribute to the poor prognosis of this disorder, there is considerable interest in a more specific treatment of this severe skin disease. PV may serve as a model disease for the development of a specific immunotherapy, because its pathogenesis as well as involved immunogenetic factors are well-characterized. This review focuses on the characterization of autoreactive T cell responses to desmoglein 3 (Dsg3), the autoantigen of PV, that presumably regulate the production of autoantibodies by providing help to the autoreactive B cells. Current knowledge on T cell epitopes of Dsg3 and the HLA class II alleles that restrict Dsg3-specific autoreactive T cell responses, as well as potential applications for a specific immunotherapy of PV, are described. PMID- 10520902 TI - Presidential address: toward physician competency. PMID- 10520903 TI - Prospective controlled study of polytetrafluoroethylene versus saphenous vein in claudicant patients with bilateral above knee femoropopliteal bypasses. AB - BACKGROUND: Although several studies have compared the patency rates of polytetrafluoroethylene (PTFE) and saphenous vein grafts (SVG) for the above knee location, none have compared the 2 grafts when implanted in the same patient with claudication who needs bilateral above knee femoropopliteal bypasses. METHODS: Forty-three patients (86 limbs) with bilateral disabling claudication who had superficial femoral artery occlusion and above knee reconstitution with 2- to 3 vessel runoff were analyzed. Patients were treated on one side with PTFE and on the other side with SVG. They were sequentially assigned to PTFE-SVG alternating with SVG-PTFE. All patients were followed using duplex ultrasound and ankle/brachial indexes at 1 month and every 6 months thereafer. RESULTS: The perioperative complication rates were 5% for PTFE and 12% for SVG. There was no operative death or perioperative amputation for either procedure. The Kaplan Meier estimate of primary, assisted primary, and secondary patency rates at 72 months were 68%, 68%, and 77% for PTFE and 76%, 83%, and 85% for SVG. There were no statistically significant differences between primary and secondary patency rates for both grafts; however; the assisted primary patency rates were higher for SVG (P < .05). The crude limb salvage rate at 72 months was 98% for PTFE and 98% for SVG. There were no risk factors identified that had an impact on graft patency. CONCLUSIONS: PTFE and SVG for above knee bypasses have comparable patency and limb salvage rates in claudicant patients with bilateral superficial femoral artery occlusion and 2- to 3-vessel runoff This may justify the use of PTFE for above knee locations in these selected patients. PMID- 10520904 TI - Initial experience with laparoscopic live donor nephrectomy. AB - BACKGROUND: Advances in laparoscopic instruments and video technology have made laparoscopic donor nephrectomy (LDN) feasible. We report our initial experience with this technique. METHODS: A retrospective review of 30 open donor nephrectomies and our first 30 LDNs was performed to assess donor and recipient outcome and resource usage. RESULTS: LDN was successfully completed in 26 donors (87%). The increased operative time and costs were balanced by less postoperative pain, earlier discharge, earlier return to normal activity and work, fewer incision problems, and less personal financial loss. Recipient outcome was not affected. CONCLUSION: LDN is technically feasible and safe, and recipient graft outcomes are equivalent. Convalescence is shortened, and there is less personal financial loss. LDN offers significant benefit to the donor and may result in increased organ donation. PMID- 10520905 TI - Zone I retroperitoneal hematoma identified by computed tomography scan as an indicator of significant abdominal injury. AB - OBJECTIVE: All zone I retroperitoneal hematomas (Z1RPHs) identified at laparotomy for blunt trauma traditionally require exploration. The purpose of this study was to correlate patient outcome after blunt abdominal trauma with the presence of Z1RPH diagnosed on admission computed tomography (CT) scan. METHODS: This is a retrospective review of patients with blunt trauma who were admitted to a Level 1 trauma center and who underwent CT scan during a 40-month period. All scans with a traumatic injury were reviewed to identify and grade Z1RPH as mild, moderate, or severe. Patients requiring operative treatment were compared with those who were observed. Statistical analysis was performed with Student's t test and chi square test, with P < .05 considered significant. RESULTS: Eighty-five (15.5%) of the CT scans were positive for Z1RPH. None of the 50 patients with a mild Z1RPH had their treatment altered. Of the 29 patients with a moderate or severe Z1RPH, 8 required celiotomy. The patients requiring celiotomy had significant elevations of solid viscus score (SVS) (4.9 +/- 1.6 versus 1.8 +/- 0.3), abdominal Abbreviated Injury Scale (3.8 +/- 0.3 versus 2.6 +/- 0.3), and transfusion requirements (13 +/- 4 versus 2 +/- 1). All patients (N = 4) with an SVS >4 required operative treatment. Seventy-two percent of patients with more than 1 intra-abdominal injury required abdominal exploration. CONCLUSIONS: The presence of a moderate or severe Z1RPH and more than 1 intra-abdominal injury or an SVS >4 on admission CT scan is an important radiographic finding. This injury pattern should be considered a contraindication for nonoperative treatment of the associated solid organ injury. PMID- 10520906 TI - Endoscopic drainage of the pancreatic pseudocyst. AB - BACKGROUND: Pancreatic pseudocyst is a common complication of chronic pancreatitis occurring in 20% to 40% of cases. Pseudocysts can be treated by endoscopic cystenterostomy or transpapillary drainage, percutaneously with computed tomography guidance or operatively. METHODS: A total of 36 endoscopic pancreatic pseudocyst drainage procedures were performed in 29 patients with 34 pseudocysts. Eighty percent presented with chronic pain, 25% had recurrent pancreatitis, and approximately one half of the patients had either gastric outlet obstruction or a palpable abdominal mass. RESULTS: Thirty-six endoscopic drainage procedures were performed, 27 cystenterostomies and 9 transpapillary drainages. Endoscopic treatment achieved complete resolution of the pseudocyst in 24 of 29 patients (83%), and the other 5 (17%) eventually required surgery. Two patients required distal pancreatectomy because of their pancreatic pathology, 2 cystgastrostomies for persistence of the pseudocyst, and 1 external drainage of an infected pancreatic cyst. The mean follow-up after the initial drainage was 16 months. There were no deaths attributed to the procedures and no complication that required surgery. Only 1 nonadherent pseudocyst (cystadenoma) required immediate operation after attempted endoscopic drainage. CONCLUSIONS: The conclude that endoscopic drainage of pancreatic pseudocysts can be both safe and effective, and definitive treatment. It should be considered as an alternative option before standard surgical drainage in selected patients. PMID- 10520907 TI - Differential expression of prostaglandin E2 and interleukin-6 in occlusive and aneurysmal aortic disease. AB - BACKGROUND: Both aortoiliac occlusive disease (AIOD) and abdominal aortic aneurysm disease (AAA) are traditionally considered degenerative conditions that are caused by atherosclerosis. Although it is becoming apparent that the pathophysiology of each condition has its own determinants, inflammation is thought to play a role in each. The purpose of this study was to analyze the inflammatory cytokines interleukin-6 (IL-6) and prostaglandin E2 (PGE2) in aortic disease and compare AAA with AIOD, as well as to compare both with normal aorta. METHODS: Aortic tissue was harvested at the time of aortic reconstructive surgery for AAA (n = 13) and AIOD (n = 14) or at time of organ harvest for normal (n = 16) aortic specimens. Whole organ cultures were immediately established, and the culture medium was collected after 72 hours. An enzyme-linked immunosorbent assay was used to assay for PGE2 and a lymphoproliferative assay was used to quantitate IL-6. Statistical analysis was performed using paired 2-tail t tests. RESULTS: Normal aorta expressed much less PGE2 (384 +/- 67 ng/mL) than either AAA (11,093 +/- 7,411 ng/mL) (P < .001) or AIOD (13, 719 +/- 3,355 ng/mL) (P < .002). However there was no statistically significant difference in PGE2 expression between the AAA and AIOD groups (P = . 44). The IL-6 assay also showed that normal aorta had very little expression (1,861 +/- 334 U/mL) compared with either AAA (14,329 +/- 4,159 U/mL) (P = . 02) or AIOD (39,805 +/- 8,426) (P < .001). Comparison between AAA and AIOD revealed significantly higher expression of IL-6 by the AIOD cultures (P = .03). CONCLUSIONS: AAA and AIOD are associated with increased expression of the proinflammatory cytokines PGE2 and IL-6. However, AIOD is associated with a much higher level of IL-6 expression than is AAA, although the level of PGE2 expression is the same. This differential expression of IL-6 may help explain the pathogenesis of these 2 distinct aortic diseases. PMID- 10520908 TI - Ultrasound-guided needle biopsy of the breast. AB - BACKGROUND: To determine the role of office-based ultrasound in the early clinical evaluation of breast masses, a consecutive series of diagnostic and interventional breast ultrasounds performed in the surgeon's office were prospectively studied. METHODS: A series of 1028 diagnostic ultrasounds were performed in 662 patients over 2 years. The clinical-pathologic data from those patients undergoing ultrasound-guided fine-needle aspiration biopsy (FNAB; n = 267 patients) and/or core needle biopsy (CNB; n = 210 patients) were reviewed. RESULTS: Of the 267 patients undergoing initial FNAB, 179 cysts were identified; 25 patients underwent no additional intervention, and 63 patients with apparently solid lesions underwent subsequent CNB. Core needle biopsy was the initial interventional approach in 147 cases. Of the 210 total patients in whom a CNB was performed, needle biopsy pathologic findings included:fibroadenoma, 57 patients; fibrocystic breast change, 82 patients; carcinoma, 53 patients; abscess/cyst, 12 patients; and other, 6 patients. Operative excision was performed in 106 of these 210 patients. There was a significantly higher false-negative rate among those patients who underwent an initial FNAB (20%; 2/10 patients) as compared with those patients undergoing CNB (3.6%; 2/55 patients; P < . 05). No cancers have been identified in those patients undergoing a benign CNB and followed for 6 to 30 months (median, 18 months). CONCLUSION: Office-based diagnostic ultrasound and interventional ultrasound that uses core needle biopsy is an effective adjunct to the early clinical evaluation of breast masses. PMID- 10520909 TI - Diagnostic and therapeutic video-assisted thoracic surgery resection of pulmonary metastases. AB - BACKGROUND: Appropriateness of video-assisted thoracic surgery (VATS) pulmonary metastasectomy for curative intent has been a controversial topic. We reviewed our experience with VATS wedge resection for peripheral lung metastases to determine the efficacy and potential adverse consequences of this approach for pulmonary metastasectomy. METHODS: One hundred seventy-seven patients underwent VATS resection of pulmonary metastases. Diagnostic resection (VATS-dx) was performed for 78 patients when percutaneous biopsy was unsuccessful or not feasible. Potentially curative resections (VATS-rx) were performed for 99 patients. The histologic findings in this group included colorectal (68), renal (7), sarcoma (6), breast (4), melanoma (3), head/neck (3), lymphoma (2), uterine (1), and "other" (5). The average number of lesions resected was 1.4 (range, 1 7). RESULTS: VATS resection was successfully performed for all VATS-dx and VATS rx patients. There were no perioperative deaths. Longitudinal follow-up demonstrated a mean survival of 18 months in the VATS-dx group and 28 months in the VATS-rx group. In the VATS-rx group, 37 (37%) of 99 were free of disease, at a mean follow-up interval of 37 months. Of the 57 recurrences, 5% were local, 26% were regional, and 69% were distant. CONCLUSIONS: Results with VATS resection of peripheral pulmonary metastases for diagnostic and potentially curative intentions appear comparable with historical results by "open" thoracotomy. Careful patient selection based on high-resolution helical CT scanning is important to avoid compromise of therapeutic intent. Conversion to thoracotomy is indicated when lesions identified preoperatively are not found or when technical problems encountered may compromise surgical margins when resecting lung metastases for potential cure. PMID- 10520910 TI - Use of omeprazole in the management of giant duodenal ulcer: results of a prospective study. AB - BACKGROUND: Giant duodenal ulcer (GDU) is generally thought to require surgical intervention. Proton pump inhibitors have beneficial effects in peptic ulcer disease, but their role in GDU disease is unknown. We examined the use of omeprazole in GDU management. METHODS: Twenty-eight patients were diagnosed with GDU. One patient required immediate operative intervention. The remaining 27 were placed on omeprazole (40 mg daily). When ulcer healing was documented by endoscopy, the patients were placed on oral histamine-2 receptor antagonist therapy. RESULTS: Of the 28 study patients, 20 (71.4%) did not require operative intervention, and 8 (28.6%) required operation for ulcer complications. Of the 15 patients with adherent clot or a visible vessel at initial endoscopy, 7 (46.7%) required operative intervention, as compared with 1 (7.7%) of the 13 patients without a visible vessel or adherent clot. This difference was statistically significant (P < .05). Twenty-three patients underwent antral biopsy and/or enzyme-linked immunosorbent assay for Helicobacter pylori, and 9 (39.1%) had a positive result. CONCLUSIONS: Omeprazole is effective in the treatment of GDU disease. An adherent clot or a visible vessel at endoscopy indicates a higher likelihood of complications requiring operation. The relatively low H pylori infection rate, as compared with other peptic ulcer disease, may indicate a different pathophysiology in GDU. PMID- 10520912 TI - Classification and management of perforations complicating endoscopic sphincterotomy. AB - BACKGROUND: The management of perforations after endoscopic sphincterotomy (ES) is controversial. The purpose of this study was to analyze the treatments and outcome of patients with ES perforations. METHODS: Between January 1994 and July 1998, in a series of 6040 endoscopic retrograde cholangiopancreatographies, 2874 (48%) ESs were performed: 40 patients (0.6%) with perforation were identified and retrospectively reviewed. RESULTS: All patients (n = 14) with guidewire perforation (group I) were recognized early, managed medically, and discharged after a mean hospital stay of 3.5 days. Twenty of 22 patients with periampullary perforation (group II) were identified early; 18 patients (90%) had aggressive endoscopic drainage, and none required operation. Of the 2 patients identified late, 1 patient required operation and subsequently died. Mean hospital stay for this group was 8.5 days. Only 1 of 4 patients with duodenal perforations (group III) was identified early; all required operation; 1 patient died, and the mean hospital stay was 19.5 days. CONCLUSIONS: ES perforation has 3 distinct types: guidewire, periampullary, and duodenal. Guidewire perforations are recognized early and resolve with medical treatment. Periampullary perforations diagnosed early respond to aggressive endoscopic drainage and medical treatment. Postsphincterotomy perforations diagnosed late (particularly duodenal) require surgical drainage, which carries a high morbidity and mortality rate. PMID- 10520911 TI - Continuous paravertebral extrapleural infusion for post-thoracotomy pain management. AB - BACKGROUND: Continuous thoracic epidural analgesia is considered by many the gold standard for post-thoracotomy pain control but is associated with its own complications. In this study we compare continuous paravertebral extrapleural to epidural infusion for post-thoracotomy pain control. METHODS: In a prospective fashion, 50 patients were randomized to receive either paravertebral or epidural infusion for post-thoracotomy pain control. The anesthesia department placed epidurals, and the operative surgeon placed unilateral paravertebral catheters. Patients were evaluated for analgesic efficacy and postoperative complications. RESULTS: We found that both methods of analgesia provide adequate postoperative pain control. Epidural infusion demonstrated an improved efficacy early in the postoperative course but provided statistically similar analgesia to paravertebral by postoperative day 2. Neither group demonstrated a greater number of pain-related complications. Narcotic-induced complications such as pruritus, nausea/vomiting, and postural hypotension/mental status changes/respiratory depression were seen with statistically similar frequency in both epidural and paravertebral arms. Urinary retention, however, was noted to be significantly more frequent in patients with epidural catheters. Drug toxicity was not observed with either epidural or paravertebral infusion. CONCLUSIONS: We recommend continuous paravertebral infusion as an improved method of post-thoracotomy analgesia that can be placed and managed by the surgeon. PMID- 10520913 TI - Results of gastric interposition for reconstruction of the pharyngoesophagus. AB - BACKGROUND: Free jejunal transfer has become the standard technique for reconstruction of the proximal pharynx and hypopharynx. Gastric tube interposition is an effective alternative when resection extends below the thoracic inlet. This study was done to determine current indications, review morbidity and mortality rates, and to define clinical and pathologic determinants of survival associated with this procedure. METHODS: We reviewed the records of 32 patients who underwent gastric tube interposition for reconstruction of the pharyngoesophagus from 1987 to 1997. RESULTS: The overall complication rate was 50%. Complications were more frequent in the reoperative group (22% vs 66%, P < .05). The overall fistula rate was 31%. The overall mortality rate was 12%. Ultimately, 71% of patients resumed oral feedings. The 5-year actuarial survival rate was 22%. Unfavorable prognostic factors associated with significantly reduced survival (P < . 05) included margin positive resection, positive lymph node involvement, and operations done for recurrent tumor CONCLUSIONS: Reconstruction of the pharyngoesophagus with gastric tube interposition is indicated for primary tumors of the hypopharynx and cervical esophagus with inferior extension below the thoracic inlet and recurrent tumors or benign strictures in which free jejunal transfer is not feasible or has failed. It can be done with acceptable morbidity and mortality and provides reasonable expectations for long-term survival and resumption of oral intake. PMID- 10520914 TI - Preservation of the recurrent laryngeal nerves in thyroid and parathyroid reoperations. AB - BACKGROUND: The recurrent laryngeal nerve (RLN) is vulnerable to injury in thyroid and parathyroid reoperations because of the presence of scar tissue and displacement of the nerve from its normal position. METHODS: Since 1993, we have performed 132 reoperations for recurrence of thyroid or parathyroid carcinoma (102 cases), persistent hyperparathyroidism (21 cases), and recurrent goiter (9 cases). One or both RLNs were identified in all cases (208 nerves). Exposure of the nerve was accomplished by a lateral approach (159 nerves), a low anterior approach (41 nerves), or the identification of the nerve between the larynx and the upper pole of the thyroid, in parathyroid reoperations (8 nerves). Dissection was then done while the nerve was kept in view at all times. RESULTS: Preoperatively, unilateral vocal cord paralysis was noted in 6 patients. Resection of a functioning RLN encased with a tumor was intentionally carried out in 5 patients. The RLNs were identified and preserved in all other cases. Among these 121 patients, transient hoarseness lasting up to a month occurred in 12 patients. CONCLUSIONS: Careful identification and exposure of the RLN through a previously undissected area can be done safely in thyroid and parathyroid reoperations and resulted in no permanent recurrent nerve injuries in our experience. PMID- 10520915 TI - Gangrenous cholecystitis: analysis of risk factors and experience with laparoscopic cholecystectomy. AB - BACKGROUND: Gangrenous cholecystitis occurs in up to 30% of patients admitted with acute cholecystitis. Factors predicting gangrenous disease in patients with acute cholecystitis remain poorly defined, making preoperative diagnosis difficult. Identification of these factors and early diagnosis of gangrenous cholecystitis will indicate more aggressive treatment, earlier operation, and a lower threshold for conversion of laparoscopic to open cholecystectomy. METHODS: We reviewed our experience with acute cholecystitis during the 2-year period of 1995 to 1996. Admitting history, physical examination, operative report, laboratory and radiology data, and pathology report were analyzed for each patient. Acute cholecystitis and its gangrenous complication were diagnosed by both gross and microscopic examination. RESULTS: One hundred fifty-four patients were admitted to the hospital with acute cholecystitis and underwent cholecystectomy; gallbladder gangrene was found in 27 (18%) of these patients. Four patients with gallbladder gangrene underwent open cholecystectomy and 23 patients underwent laparoscopic cholecystectomy, of which 15 (65%) were completed laparoscopically and 8 (35%) had open conversion as a result of severe inflammation. Risk factors for gallbladder gangrene included male gender, age older than 50 years, history of cardiovascular disease, and leukocytosis greater than 17,000 white blood cells/mL. CONCLUSIONS: Older male patients (age older than 50 years) with history of cardiovascular disease, leukocytosis greater than 17,000 white blood cells/mL, and acute cholecystitis have increased risk of gallbladder gangrene and conversion of laparoscopic cholecystectomy to open cholecystectomy. Urgent laparoscopic cholecystectomy with low threshold for conversion to open cholecystectomy should be considered in these patients at high risk for gallbladder gangrene. PMID- 10520916 TI - Higher prevalence of abdominal aortic aneurysms in patients with carotid stenosis but without diabetes. AB - BACKGROUND: We compared abdominal aortic aneurysm (AAA) prevalence in 3 groups of patients at the Hines Veterans Affairs Medical Center: (1) patients with 50% or more carotid stenosis, (2) patients with less than 50% stenosis, and (3) patients screened for the Aneurysm Detection and Management (ADAM) study. METHODS: Of all the patients referred to the vascular laboratory for carotid duplex examination during a 12-month period, patients with 50% or more carotid stenosis underwent ultrasonography of the abdominal aorta unless they had a previous scan or previous aortic surgery (group 1, n = 374). Patients with less than 50% carotid stenosis who had been screened for ADAM comprised group 2 (n = 139). They were compared with all patients screened for ADAM at our center during the same time period (group 3, n = 2477). RESULTS: AAA of 3.0 cm or more were present in 18.2%, 12.2%, and 7.2% of groups 1, 2, and 3, respectively; AAA of 4.0 cm or more were present in 8.3%, 5.8%, and 2.1% of groups 1, 2, and 3, respectively. Among patients with carotid stenosis, those patients without diabetes accounted for the observed increase in prevalence (21.9 % > or = 3.0 cm and 10.2% > or = 4.0 cm vs 9.2% and 2.8% in patients with diabetes). CONCLUSIONS: The relative risk of AAA is 2 to 3 times greater in patients with carotid stenosis compared with patients undergoing routine screening. However, only patients without diabetes account for the increased prevalence. Selective AAA screening of patients who are not diabetic with carotid stenosis is recommended. PMID- 10520917 TI - Civilian rectal trauma: a changing perspective. AB - BACKGROUND: Recently the Organ Injury Scaling Committee of the American Association for the Surgery of Trauma developed a Rectal Injury Scaling System (RISS). Little data exist regarding its clinical utility. METHODS: We retrospectively reviewed 45 patients with rectal injuries to assess the impact of the RISS on patient management and outcome. We compared RISS grade I patients (group I, partial-thickness injury) with patients with grades 2, 3, and 4 injuries (group II, full-thickness injury). RESULTS: Group II underwent distal rectal washout and repair of the injury twice as often and had a significantly higher rate of diversion of the fecal stream. This was associated with a 3-fold increase in complications. The only complications in group I were in patients managed with diversion of the fecal stream and distal rectal washout. CONCLUSIONS: Our data suggest that aggressive surgical management for RISS grade I injury may not be necessary. Implementation of therapy based on the RISS may improve outcomes of civilian rectal trauma. PMID- 10520918 TI - Medical ethics curriculum for surgical residents: results of a pilot project. AB - INTRODUCTION: This study sought to develop and evaluate a medical ethics curriculum designed specifically for surgical residents. METHODS: The learning needs of surgical residents relevant to ethics were determined by using a structured literature review and synthesis strategy. We identified 5 primary areas of importance for ethics education for surgical residents: withdrawing and withholding treatment, advance directives, do-not-resuscitate orders, informed consent, and communicating bad news. Learning objectives were developed, and teaching plans were designed for four 90-minute interactive teaching episodes on the basis of adult learning principles. We surveyed residents using a published survey instrument modified for surgery to identify residents' beliefs about the usefulness of ethics training, confidence in addressing ethical issues, and factual knowledge of ethics questions. RESULTS: Twenty surgical residents at a single institution completed the pretest and posttest close-ended surveys. Results showed that although 88% had formal ethics exposure in medical school, 93% considered ethics education at the resident level to be a "very important" or "important" topic. Residents' confidence in addressing ethical issues showed statistically significant improvement between pretest and posttest surveys for 13 of 23 items. There were no statistically significant linear relationships between postgraduate year of residency and the pretest confidence items or the number of correct responses on the pretest multiple-choice items. CONCLUSIONS: Despite the prevalence of ethics education during medical school, surgical residents welcome formal instruction on numerous ethical issues pertinent to surgical practice. A focused curriculum can be developed that has a measurable impact on residents' confidence in addressing ethical issues. PMID- 10520919 TI - Surgical portosystemic shunts for treatment of portal hypertensive bleeding: outcome and effect on liver function. AB - BACKGROUND: Since the advent of liver transplantation and transjugular intrahepatic portosystemic shunts (TIPS), the role of surgical portosystemic shunts in the treatment of portal hypertension has changed. However, we have continued to use portosystemic shunts in patients with noncirrhotic portal hypertension and in patients with Child's A cirrhosis. METHODS: We performed 48 surgical portosystemic shunt procedures between 1988 and 1998. The outcomes of these patients were evaluated to assess the efficacy of this treatment. Data from 39 of 48 patients were available for analysis. The average follow-up was 42 months. RESULTS: Liver function generally remained stable for the patients; only 2 patients had progressive liver failure and required transplant procedures. Gastrointestinal bleeding (3 patients), encephalopathy (3 patients), and shunt thrombosis (3 patients) were rare. Patient survival was 81% at 4 years, similar to survival with liver transplantation (P = .22). CONCLUSIONS: Surgical shunts remain the treatment of choice for prevention of recurrent variceal bleeding in patients with good liver function and portal hypertension. PMID- 10520920 TI - Learning sentinel node biopsy: results of a prospective randomized trial of two techniques. AB - BACKGROUND: Evidence indicates that sentinel node (SN) biopsy can accurately predict axillary nodal status. Debate exists as to the optimal method of SN identification. METHODS: Patients with clinical T1 or T2 tumors and negative axillae were randomized to SN localization with blue dye (B) alone (n = 50) or blue dye plus radioactivity (B+R) (n = 42). Patients undergoing needle localization (n = 47) were assigned to blue dye. RESULTS: The SN was identified in 110 patients (79%) and contained metastases in 28. The SN predicted the axillary nodal status in 96% of cases. The SN identification rate did not differ between B (88%) or B+R (86%) but was significantly lower in patients requiring localization (64%). The time to SN identification also did not differ between B and B+R. The number of cases done by an individual surgeon was a significant predictor of SN identification. A stepwise logistic regression analysis of factors influencing the success of SN identification identified tumor location, needle localization, number of operations, and body mass index as significant predictors. CONCLUSIONS: Our study does not identify any advantage for the use of the more expensive and complex method of SN identification using B+R compared with B alone, even for surgeons learning the techniques. PMID- 10520921 TI - Reoperative laparoscopic antireflux surgery. AB - BACKGROUND: Laparoscopic antireflux surgery (LAP) is becoming increasingly used for the surgical treatment of medical recalcitrant gastroesophageal reflux disease (GERD). We sought to determine the utility of remedial LAP approaches to antireflux surgery. METHODS: From March 1996 to December 1998, 15 patients underwent remedial LAP to manage medically recalcitrant recurrent GERD after LAP (n = 8) or open antireflux procedure (n = 1) and/or troublesome postfundoplication complications (dysphagia 6, gas bloat 4). The remedial LAP surgery consisted of conversion from Nissen to Toupet fundoplication to manage dysphagia or gas bloat symptoms (n = 7), revision of IAP Nissen fundoplication (n = 7) and LAP revision of a failed open Nissen fundoplication (n = 1) for recurrent reflux. RESULTS: The remedial LAP repair was accomplished in all patients. Findings at operation included disrupted fundoplication (n = 6), incomplete or inappropriately positioned fundoplication (n = 2), paraesophageal hernia (n = 3), or a normal total fundoplication among patients with primary dysphagia (n = 4). Follow-up symptom scoring beyond 3 months of remedial surgery demonstrated a change from the preoperative mean dysphagia, heartburn, gas bloat, and regurgitation score (P < .05). Follow-up GERD testing (manometry, upper gastrointestinal tract, pH testing) was normal in 13 of the 15 patients. CONCLUSIONS: Reoperative antireflux surgery can be accomplished using LAP approaches without compromise of therapeutic intent or increased surgical morbidity. Surgeons sufficiently experienced with these LAP repairs may consider repeat LAP instead of open surgery for patients with recurrent GERD or postfundoplication problems. PMID- 10520922 TI - The treatment of malignant endobronchial obstruction with laser ablation. AB - BACKGROUND: We compared a new endoscopic treatment for malignant endobronchial obstruction known as photodynamic therapy (PDT) with the more established therapy of neodymium: yttrium-aluminum garnet laser (Nd:YAG) therapy. METHODS: A retrospective review was conducted of the medical records at our institution from 1988 to 1999 of patients treated for bronchial obstruction by thermal laser vaporization (Nd:YAG) or by PDT using the tunable dye laser in combination with a light-sensitive dye (PDT). The Nd:YAG procedure vaporized the obstructing neoplasm, whereas the PDT procedure photoablated the obstruction. Thirty-day mortality and morbidity rates were analyzed for both treatment groups using chi square analysis. RESULTS: Of the 102 patients who were suitable for review, 83 received treatment with the Nd:YAG laser and 19 patients received treatment with PDT. Morbidity rates were comparable in both groups (22% for Nd:YAG vs 31% for PDT; P > .05). Equally common complications in both groups were respiratory failure and hypoxemia. Five Nd:YAG patients (6%) died within 30 days after treatment (3 of respiratory failure, 2 of massive hemoptysis), whereas 2 patients (10%) in the PDT group (1 of massive hemoptysis, 1 of acute myocardial infarction) died (P > . 05). CONCLUSIONS: PDT and Nd:YAG have similar mortality and morbidity rates. In our experience, PDT is a better choice for the treatment of malignant bronchial obstruction because it is technically easier, potentially safer, and does not require general anesthesia. PMID- 10520923 TI - Management of diagnostic dilemmas of the pancreas by ultrasonographically guided laparoscopic biopsy. AB - INTRODUCTION: Pancreatic lesions may be difficult to diagnose because of small size or inaccessibility. Such lesions are being seen with increasing frequency because of advances in pancreatic imaging techniques. In the past 18 months we have evaluated 14 patients whose pancreatic lesions could not be diagnosed by traditional means, including percutaneous biopsy. METHODS: With the patient under general anesthesia, the anterior surface of the pancreas was exposed by a three trocar laparoscopic technique. The lesion was located by laparoscopic ultrasonography. A core biopsy needle was inserted into the lesion under simultaneous visual and ultrasonographic guidance using picture-in-picture techniques. RESULTS: The main diagnostic dilemma encountered was the differentiation of pancreatic cancer from pancreatitis. Other conditions were lymphoma and renal cell carcinoma. Excellent tissue samples were obtained, allowing diagnosis and planning of treatment in all cases. Operative time ranged from 1 to 4 hours, and length of stay ranged from 1 to 3 days. Blood transfusions were not required, and there were no complications. Alcohol nerve block was performed laparoscopically in one patient in this group after the diagnosis was made by frozen section. CONCLUSIONS: Direct ultrasonographically guided laparoscopic biopsy provides rapid, safe diagnosis of pancreatic lesions. PMID- 10520924 TI - The expression of genes in the ubiquitin-proteasome proteolytic pathway is increased in skeletal muscle from patients with cancer. AB - BACKGROUND: The intracellular mechanisms of muscle cachexia in patients with cancer are not known. To assess the role of the ubiquitin-proteasome proteolytic pathway in cancer-induced muscle breakdown, we determined messenger RNA levels for ubiquitin and several 20S proteasome subunits in muscle from patients undergoing surgery for cancer METHODS: A biopsy specimen was obtained from the rectus abdominis muscle in patients undergoing laparotomy for cancer (n = 6) or noncancer disease (n = 6). Tissue levels of mRNA for ubiquitin and the 20S proteasome subunits HC3, HC5, HC7, and HC9 were determined by dot blot analysis. RESULTS: The mRNA levels for ubiquitin and the 20S proteasome subunits were 2 to 4 times higher in muscle from patients with cancer than in muscle from control patients. CONCLUSION: This is the first report of increased expression of genes in the ubiquitin-proteasome proteolytic pathway in muscle tissue from patients with cancer. Cancer-induced muscle catabolism may at least in part reflect ubiquitin-proteasome-dependent protein breakdown. PMID- 10520925 TI - Clinical pathway implementation improves outcomes for complex biliary surgery. AB - BACKGROUND: Complex biliary surgery is associated with significant morbidity, prolonged hospital stay, and high cost. Clinical pathway implementation has the potential to standardize treatment and improve outcomes. Therefore the aim of this analysis was to determine whether clinical pathway implementation and/or feedback of outcome data would alter hospital stay, charges, and mortality rates for complex biliary surgery at an academic medical center METHODS: Pre- and postoperative length of stay, hospital charges, and mortality rates were monitored for 36 months before (period 1) and for 2 18-month periods (periods 2 and 3) after implementation of a clinical pathway for hepaticojejunostomy. Outcome data were provided to the surgeons 18 months after pathway implementation to determine whether further clinical practice improvement was possible. RESULTS: From 1991 to 1997, 339 patients underwent hepaticojejunostomy at The Johns Hopkins Hospital for malignant and benign biliary obstruction. Total length of stay was 13.3 +/- 0.9 days for period 1 compared with 12.5 +/- 0.8 days for period 2 (not significant) and 10.1 +/- 0.3 days for period 3 (P < .01 vs period 1; P < .03 vs period 2). Hospital charges averaged $24,446 during period 1 compared with $23,338 during period 2 and $20,240 during period 3 (P < .01 vs periods 1 and 2). Hospital mortality rate was 4.5% during period 1 compared with 0.7% during periods 2 and 3 (P < .05). CONCLUSIONS: These data suggest that implementation of a clinical pathway for hepaticojejunostomy reduces hospital mortality rates and that feedback of outcome data to surgeons results in further clinical practice improvement. Thus clinical pathway implementation and feedback are effective methods to control costs at an academic medical center. PMID- 10520926 TI - Femoral-infrapopliteal bypass with prosthetic grafts. AB - BACKGROUND: Patients who require prosthetic infrapopliteal of the lower extremity have historically had dismal long-term results. This study examined the outcome of patients undergoing femoral-distal arterial bypass with expanded polytetrafluoroethylene (ePTFE) grafts. METHODS: Femoral to infrapopliteal artery bypasses with ePTFE performed between 1990 and 1997 were reviewed. Graft patency, limb salvage, and survival rates were calculated by actuarial analysis. Different anastomotic adjuncts (direct end-to-side anastomosis, vein patch anastomosis, and arteriovenous fistula [AVF] anastomosis) were compared with the log-rank test. RESULTS: Seventy-four femoral-infrapopliteal bypasses with ePTFE were performed in 67 patients for limb salvage. At 24 months the primary patency, assisted primary patency, and secondary patency rates were 40% +/- 10% (SEMAI), 48% +/- 11%, and 52% +/- 11%, respectively. Limb salvage was successful in 62 % +/- 10% of the bypasses at 24 months. Forty-six percent of the bypasses were performed with an AVF; 35% were performed with direct end-to-side anastomosis, and 19% were performed with a vein patch anastomosis. Apparent trends in favor of the AVF group did not attain statistical significance with 24-month patency rates of 65% +/- 19%, 44% +/- 16%, and 35% +/- 20%, respectively, in the 3 subgroups. Limb salvage rates were similar (64% +/- 17%, 56% +/- 15%, and 76% +/- 22%, respectively) at 24 months. CONCLUSIONS: Patency and limb salvage rates are sufficient to justify the use of ePTFE grafts in infrapopliteal bypass when adequate autogenous material is unavailable. PMID- 10520927 TI - Cryotherapy extends the indications for treatment of colorectal liver metastases. AB - BACKGROUND: Hepatic resection for colorectal metastases has been established as the best option for patients with 4 or less lesions meeting specified criteria. Recently, the use of intraoperative ultrasound has increased the detection of previously occult liver lesions, and cryotherapy has allowed the treatment of liver lesions in inaccessible areas with less destruction of normal liver in the case of multiple lesions. 14e prospectively performed hepatic resection or cryotherapy to test the hypothesis that more than 4 liver metastases could be safely and successfully treated with improved long-term survival. METHODS: From August 1993 to January 1999, 137 patients with liver metastases from colorectal cancer were treated with hepatic resection or cryotherapy at the Medical College of Wisconsin. Preoperative and postoperative computed tomography scans, intraoperative assessments of lesion number and curability, number of blood transfusions administered, length of stay, complications experienced, and overall survival rates were reviewed. RESULTS: One hundred thirty-seven patients were explored. Treatment consisted of resection alone in 34, cryotherapy alone in 20, both treatments in 52, and no treatment was possible in 31 patients. "Curability" was defined as complete resection or cryotherapy of all identifiable tumor at the conclusion of the operation. A Cox proportional hazards model demonstrated that survival was determined by the destruction of all identifiable metastases (P < . 001) and was not statistically influenced by age, gender type of therapy, or the number of metastases treated. CONCLUSIONS: Surgical treatment of colorectal liver metastases remains the best option for patients with this disease. A key factor in overall survival is the destruction or resection of all identifiable disease and not the number of tumors per se. Using cryotherapy as an addition to the surgical arsenal, patients previously deemed unresectable because of the number of lesions have a chance for long-term survival. This study demonstrates improved long-term survival for "cured" patients with more than 4 metastatic lesions, thereby extending the indications for resection/ablation. PMID- 10520928 TI - An analysis of male and female breast cancer treatment and survival among demographically identical pairs of patients. AB - BACKGROUND: Male breast cancer is rare, and there are no large comparative studies to guide treatment. We used National Cancer Data Base data on 4755 men and 624,174 women who had breast cancer (1985-1994) to identify equivalent groups of male and female breast cancer patients. METHODS: For each man with breast cancer, the next woman treated at the same hospital was sought who matched the man's age (within 5 years), ethnicity, income category, and stage. We identified 3627 closely matched pairs of male and female patients with breast cancer. RESULTS: Men were more likely to be treated with mastectomy (modified radical, 65% of men versus 55.1% of women; radical, 2.5% of men versus 0.9% of women; simple, 7.6% of men versus 3.4% of women; P <.001), and more likely to receive radiation therapy after mastectomy (men, 29%; women, 11%; P <.001). Men treated with lumpectomy were less likely to receive radiation therapy (men, 54%; women, 68%; P <. 001). Men were also less likely to receive chemotherapy (26.7% of men versus 40.6% of women; P <. 001) after any surgical treatment. CONCLUSIONS: This large comparative study is the first to detail stage-specific differences in contemporary treatment strategies for highly comparable groups of men and women treated for breast cancer. Further studies of male breast cancer should focus on identifying prognostic factors and defining optimal therapy. PMID- 10520929 TI - Comparison of symptomatic and quality of life outcomes of laparoscopic versus open antireflux surgery. AB - BACKGROUND: Even though laparoscopic antireflux procedures have become the surgical treatment of choice for gastroesophageal reflux disease (GERD), little quantitative data exist comparing symptomatic and quality of life outcomes between laparoscopic and standard open procedures. This study was done to compare short-term outcomes. METHODS: All patients referred for surgical treatment of GERD are prospectively followed with a disease-specific reflux symptom score (the GERD-HRQL, best score 0, worst score 50) and a generic quality of life questionnaire (the SF-36, best score 100, worst score 0). Patients are evaluated preoperatively and at least 6 weeks postoperatively. Patients were treated with either laparoscopic or open Nissen (360-degree wrap) or Toupet (270-degree wrap) fundoplications. RESULTS: Sixty patients underwent laparoscopic surgery (LS) and 20 open surgery (OS). LS and OS had significant improvement in the median GERD HRQL scores, 27 to 3 and 27 to 1, respectively, both P < .000001. LS had statistically significant improvements in the SF-36 domains of mental health (62 to 71.5, P = .05) and general health (57 to 67, P = .004). There was no worsening in any of the other 6 domains. OS produced a worsening score in the domain of physical functioning (75 to 67.5, P = .02). LS had better postoperative scores compared with OS in the domains of physical functioning (80 vs 67.5, P = .05) and trended to better scores in bodily pain (64 vs 51.5, P = .09). CONCLUSIONS: LS produces equivalent improvement in reflux symptoms compared with OS, with improved general quality of life outcomes. PMID- 10520930 TI - Surgical decompression of ductal obstruction in patients with chronic pancreatitis. AB - BACKGROUND: Recurrent acute pancreatitis often leads to chronic obstructive ductal disease requiring operative decompression. METHODS: From 1983 through 1998, 124 patients with ductal obstruction underwent lateral pancreaticojejunostomy (78 patients), distal pancreatectomy with end-to-side pancreaticojejunostomy (27 patients), distal pancreatectomy with placement of a pancreas with a filleted duct within a jejunal limb (15 patients), or pancreaticoduodenectomy (4 patients). Preoperative symptoms included abdominal and back pain (99%), nausea with vomiting (99%), and diarrhea with weight loss (11%). Associated conditions included hypertension (20%) and diabetes mellitus (12%). Endoscopy in 106 patients demonstrated distal stricture (37%), proximal stricture (36%), pseudocyst (30%), chain of lakes (15%), calcification and debris (19%), and bile duct stricture (8%). RESULTS: Two patients died, one of an unrecognized esophageal perforation during intubation and the other of leakage of a 1-layer pancreaticojejunostomy. Thirty-six patients developed 53 complications including intra-abdominal abscess (7 patients) and bleeding requiring reoperation in 1 patient. Pain relief was complete in 61 patients, substantial in 39 patients, moderate in 11 patients, minimal in 8 patients, and nonexistent in 3 patients with multiple stones and narrow duct. Ten patients died, with 6 deaths as a result of pancreatic cancer Two other patients may have died of pancreatic cancer. CONCLUSIONS: Lateral pancreaticojejunostomy is the procedure of choice in most patients. Recurrent pancreatitis usually follows alcoholic binges. Long-term follow-up must assess for pancreatic cancer. PMID- 10520931 TI - Follicular or Hurthle cell neoplasm of the thyroid: can clinical factors be used to predict carcinoma and determine extent of thyroidectomy? AB - BACKGROUND: Fine-needle aspiration biopsy (FNAB) and frozen section exam are of limited or no value in distinguishing benign and malignant follicular or Hurthle cell neoplasms of the thyroid gland. METHODS: Patients who underwent thyroidectomy for treatment of a follicular or Hurthle cell neoplasm between 1990 and 1998 were identified and evaluated for age, gender, head and neck irradiation, nodule size, and cytologic atypia to determine whether clinical factors were predictive of carcinoma. RESULTS: Of the 352 patients evaluated for nodular thyroid disease, 75 (21%) underwent thyroidectomy after an indeterminate FNAB finding, 66 with follicular and 9 with a Hurthle cell neoplasm. Seventeen (23%) of the patients had carcinoma-follicular variant of papillary (10), follicular (6), and Hurthle cell (1). Carcinoma was diagnosed in 15 of 64 women and 2 of 11 men (P > .05). The mean age was 43 +/- 21 years and 50 +/- 16 years, respectively, in patients with and without carcinoma (P > . 05). Three patients had previous neck irradiation and none had carcinoma. Mean nodule size was 4.2 +/ 2.7 cm and 4.3 +/- 3.5 cm, respectively, in patients with and without carcinoma (P > . 05). Cytologic atypia was present in 8 of 17 patients with carcinoma and 20 of 58 patients without carcinoma (P > .05). CONCLUSIONS: Clinical factors were not helpful in predicting carcinoma in patients with an indeterminate FNAB finding and thus cannot be used to reliably select patients for more extensive thyroidectomy. PMID- 10520933 TI - Hereditary motor and sensory neuropathy: the plot thickens. PMID- 10520934 TI - Ventricular volume and transmural pressure gradient in normal pressure hydrocephalus. PMID- 10520932 TI - Optimizing the management of blunt splenic injury in adults and children. AB - BACKGROUND: The treatment for splenic injury is evolving to an increased use of nonoperative management. We studied patients with blunt injury to the spleen to determine the overall success with splenic salvage and the reason that adults and children have different outcomes. METHODS: Patient records were reviewed retrospectively for information and parameters that may influence outcome. Patients were categorized by age and type of management. RESULTS: Two hundred sixty-seven patients (222 adults; 45 children < 16 years old) with blunt splenic trauma were treated over a 7.5-year period. Adults had a significantly higher injury severity score (ISS; 27.2 +/- 0.9 vs 19.9 +/- 2.0; P < .05), splenic injury score (SIS; 2.8 +/- 0.1 vs 2.3 +/- 0.1; P < .01), and mortality rate (11.7% vs 2.2%; P < .05) compared with children. Eighty-six adults and 3 children had emergent operation; 23 patients had splenorrhaphy. Nonoperative management was selected initially in 178 patients; 83% (105 adults and 42 children) were treated successfully. The ISS and SIS of patients in whom nonoperative management failed were different from those patients in whom treatment was successful (ISS, 27.5 +/- 2.1 vs 20.6 +/- 1.0; SIS, 3.6 +/- 0.2 vs 2.1 +/- 0.1; P < .05) but were similar to those patients who needed initial emergent operation. Adults and children who had successful nonoperative management had similar ISSs (21.4 +/- 1.1 vs 18.4 +/- 2.0) and SISs (2.0 +/- 0.1 vs 2.3 +/- 0.1). Overall splenic salvage was achieved in 64% of patients (57% of adults and 96 % of children). Salvage increased from 50% to 85% during the study period. CONCLUSIONS: Splenic preservation is possible in most adults and children with blunt injury with the appropriate use of both operative salvage and nonoperative treatment. The higher salvage rate and decreased need for operation in children is due to their lower severity of overall injury and splenic injury. Operative salvage has become less common in adults because more patients are selected for nonoperative management. PMID- 10520935 TI - Molecular pathogenesis of Friedreich ataxia. AB - Friedreich ataxia, the most common type of inherited ataxia, is itself caused in most cases by a large expansion of an intronic GAA repeat, resulting in decreased expression of the target frataxin gene. The autosomal recessive inheritance of the disease gives this triplet repeat mutation some unique features of natural history and evolution. Frataxin is a mitochondrial protein that has homologues in yeast and even in gram-negative bacteria. Yeast organisms deficient in the frataxin homologue accumulate iron in mitochondria and show increased sensitivity to oxidative stress. This suggests that Friedreich ataxia is caused by mitochondrial dysfunction and free radical toxicity. PMID- 10520936 TI - The molecular pathogenesis of Pelizaeus-Merzbacher disease. AB - In 1885, Pelizaeus described 5 boys in a single family with nystagmus, spastic quadriparesis, ataxia, and delay in cognitive development. In 1910, Merzbacher reexamined this family, which then included 14 affected individuals, including 2 girls, and found that all affected family members shared a common female ancestor. Also, he noted that the disease was passed exclusively through the female line without male-to-male transmission. Pathological analysis of brain tissue from one affected individual showed that most of the central white matter lacked histochemical staining for myelin, although there were occasional small regions of preserved myelin, giving the sections a "tigroid" appearance. The description of this family provides the clinical, genetic, and pathological basis for Pelizaeus-Merzbacher disease (PMD): an X-linked disorder of myelination classically characterized by nystagmus, spastic quadriparesis, ataxia, and cognitive delay in early childhood. PMID- 10520937 TI - Polymerase chain reaction in the diagnosis and management of central nervous system infections. AB - Polymerase chain reaction (PCR) is a broadly applied laboratory test for the diagnosis of a wide variety of central nervous system (CNS) diseases, including genetic and autoimmune diseases, malignant neoplasms, and infections. With its ability to detect minute amounts of DNA or RNA contained in tissues or fluids, PCR has improved the rapidity and accuracy of diagnosis, enhanced understanding of pathogenesis, and helped identify infectious causes for diseases previously considered idiopathic. In addition, PCR can be performed on a variety of tissues preserved in different ways--even archival specimens can be used to provide important epidemiological information. By making quick and precise diagnoses, appropriate treatments can be instituted, and unnecessary or invasive investigations can be avoided. PMID- 10520938 TI - Beneficial effect of siphoning in treatment of adult hydrocephalus. AB - OBJECTIVE: To increase awareness about the treatment of adult patients with shunt nonresponsive hydrocephalus--a state characterized by marked ventriculomegaly, low intracranial pressure, and a patent cerebrospinal fluid diversionary shunt. DESIGN: Retrospective analysis of hospital and outpatient records. PATIENTS: Four patients with symptomatic ventriculomegaly and patent ventriculoperitoneal shunts treated with a protocol of progressive ventricular hypotension induced by external cerebrospinal fluid drainage. RESULTS: Severe clinical manifestations exhibited by the patients, including parkinsonian features, Parinaud syndrome, and extensor posturing, completely reversed once a normalization of ventricular size was achieved. External ventricular drainage pressures as low as -30 cm H2O were required to reduce ventricular size. All patients finally received a shunt incorporating a standard medium differential pressure valve with no antisiphon device. CONCLUSIONS: Shunt siphoning may be an essential mechanism by which cerebrospinal fluid shunting is effective in many patients with adult hydrocephalus. Cerebrospinal fluid shunts that contain an antisiphon device are ineffective in these patients, despite the attainment of "physiologic" intracranial pressures. Based on reported experimental and clinical evidence, it seems that the cause of this condition may be related to abnormally high intracranial compliance. PMID- 10520940 TI - Glutamine synthetase in cerebrospinal fluid, serum, and brain: a diagnostic marker for Alzheimer disease? AB - OBJECTIVES: To determine whether the glutamine synthetase (GS) level in cerebrospinal fluid (CSF) is a useful biochemical marker in the diagnosis of Alzheimer disease (AD), and to assess the source of GS (brain vs. blood derived) in CSF. METHODS: Sandwich enzyme immunoassay and immunoblotting were applied to detect GS in CSF and in serum from neurologically healthy control subjects and patients with neurodegenerative diseases, including AD. The origin of GS was estimated by the concentration gradients of CSF to serum and ventricular to lumbar CSF. In addition, postmortem brain tissue from controls and patients with AD was analyzed using immunohistochemistry for expression of GS. RESULTS: Levels of GS were significantly increased in lumbar CSF from patients with AD (20+/-12 pg/mL; P = .01) and to a lesser extent in patients with vascular dementia and amyotrophic lateral sclerosis. In CSF of controls, GS levels were 4+/-3 pg/mL. The GS concentration gradients were less than 1:10 for CSF to serum and 2:1 for ventricular to lumbar CSF. Immunoreactivity of GS was most prominent in astrocytes from temporal neocortex of patients with AD, suggesting a relationship between astrocyte reactions and increased GS levels in CSF. CONCLUSIONS: Level of GS in lumbar CSF of patients with AD is increased significantly but nonspecifically, probably related to the strong astrogliosis in brain. Glutamine synthetase in lumbar CSF is mainly brain derived. PMID- 10520939 TI - Clinical and pathological evidence for a frontal variant of Alzheimer disease. AB - OBJECTIVE: To evaluate the clinical and pathological features of a subgroup of patients with Alzheimer disease (AD) who exhibited early and disproportionately severe impairments on tests of frontal lobe functioning. We hypothesized that these patients would exhibit a greater degree of either neurofibrillary tangle (NFT) or senile plaque pathology in the frontal lobes than would patients with typical AD. DESIGN AND OUTCOME MEASURES: We examined the neuropsychological profiles and senile plaque and NFT accumulation in the frontal, entorhinal, temporal, and parietal cortices in 3 patients with AD who exhibited disproportionate frontal impairments during early stages of dementia (frontal AD) and 3 matched patients with typical AD (typical AD). RESULTS: Compared with the typical AD group, the frontal AD group performed significantly worse on 2 tests of frontal lobe functioning and on the Wechsler Adult Intelligence Scale-Revised Block Design test. No significant group differences were found on other tests. Analysis of brain tissue samples demonstrated that, despite comparable entorhinal, temporal, and parietal NFT loads, the frontal AD group showed a significantly higher NFT load in the frontal cortex than the typical AD group. Senile plaque pathology in the frontal and entorhinal cortices did not differentiate the 2 groups. CONCLUSIONS: We identified a subgroup of patients with pathologically confirmed AD who presented in the early stages of dementia with disproportionate impairments on tests of frontal lobe functioning and had a greater-than-expected degree of NFT pathology in the frontal lobes, suggesting the existence of a frontal variant of AD that has distinctive clinical and pathological features. PMID- 10520941 TI - Sympathetic skin response and heart rate variability in patients with Huntington disease. AB - OBJECTIVE: To examine the autonomic nervous system functions in patients with Huntington disease. BACKGROUND: Although patients with Huntington disease frequently experience vegetative symptoms, it is not clear if there is dysfunction of the autonomic nervous system. METHODS: Sympathetic skin response (SSR) latency and amplitude from both palms and soles and R-R (heart rate) interval variation (RRIV) at rest and during the Valsalva maneuver were examined in 22 patients and 21 age-matched controls. Unified Huntington's Disease Rating Scale scores were determined in all the patients. RESULTS: Our data are reported as means +/- SEMs. The SSR latencies in patients (mean palm latency, 1835.8+/ 110.7 milliseconds; mean sole latency, 2625.3+/-226.9 milliseconds) were prolonged compared with controls (mean palm latency, 1359.5+/-28 milliseconds [P<.01]); mean sole latency, 2038.1+/-44.9 milliseconds [P<.01]) and amplitudes in patients (mean amplitude, 1063.1+/-237.7 microV) were smaller compared with controls (mean amplitude, 1846.3+/-251.2 microV [P<.05]). The RRIV in patients both at rest (mean RRIV in patients, 3.7%+/-0.4% vs. controls, 9.7%+/-0.6% [P<.01]) and during the Valsalva maneuver (mean RRIV in patients, 6.3%+/-1.6% vs. controls, 14.5%+/-1.2% [P<.01]) was lower compared with controls. Furthermore, the prolonged SSR latencies, smaller amplitudes, and lower RRIV in patients compared with controls closely correlated with the various components of the Unified Huntington's Disease Rating Scale scores (total behavior score and SSR latency, R = 0.6 [P<.01]; total behavior score and SSR amplitude, R = -0.5 [P<.05]; total behavior score and RRIV, R = -0.4 [P<.05]; verbal fluency and SSR latency, R = -0.5 [P<.05]; verbal fluency and SSR amplitude, R = 0.5 [P<.05], verbal fluency and RRIV, R = 0.5 [P<.05]; total functional capacity and SSR latency, R = -0.6 [P<.01]; total functional capacity and SSR amplitude, R = 0.5 [P<.05]). CONCLUSION: These results suggest that there is autonomic nervous system dysfunction in patients with Huntington disease. PMID- 10520942 TI - Progression of clinical deterioration and pathological changes in patients with Alzheimer disease evaluated at biopsy and autopsy. AB - OBJECTIVES: To quantify the progression of senile plaques, neurofibrillary tangles, cerebral amyloid angiopathy, and microglial activation in the cortex and white matter of patients with Alzheimer disease evaluated at both biopsy and subsequent autopsy and correlate these changes with the progression of neurologic impairment. SETTING: Academic referral center for patient with Alzheimer disease. PATIENTS: Four patients meeting the clinical criteria for Alzheimer disease, enrolled in a pilot study for the evaluation of response to intracerebroventricular administration of bethanechol chloride. The patients were followed up until death occurred and autopsy was performed. RESULTS: All 4 patients had progressive deterioration from the time of biopsy to autopsy (9-11 years). Pathological investigations showed a striking increase in the density of senile plaques and neurofibrillary tangles in 2 of 4 patients from biopsy to autopsy, and a significant increase in microglial activation in 1 of 4 cases. Severity of cerebral amyloid angiopathy varied significantly among patients, 1 of whom displayed striking amyloid deposition with associated subcortical white matter atrophy. CONCLUSIONS: These unique data demonstrate that the progressive neurologic impairment in Alzheimer disease is accompanied by a significant increase in senile plaque and neurofibrillary tangle counts in the frontal cortex and, possibly in some patients, by increased microglial cell activation. Cerebral amyloid angiopathy was associated with significant white matter disease. PMID- 10520943 TI - Course of depression during the initiation of interferon beta-1a treatment for multiple sclerosis. AB - OBJECTIVE: To examine the hypothesis that increases in depression after initiation of treatment with interferon beta-1a for multiple sclerosis can be explained as representing a return to pretreatment levels of depression. DESIGN: Level of depression in patients with multiple sclerosis was assessed at 3 time points: 2 weeks before initiation of interferon beta-la treatment, at initiation of treatment, and at 2-month follow-up. SETTING: A health maintenance organization. PATIENTS: Fifty-six patients with confirmed relapsing forms of multiple sclerosis. MAIN OUTCOME MEASURE: The depression-dejection scale of the Profile of Mood States. RESULTS: Patients who scored high on the depression measure 2 weeks before the initiation of interferon beta-1a treatment showed significant reduction in depression at the initiation of treatment. However, depression returned nearly to initial levels within 2 months. CONCLUSIONS: These findings suggest that increases in depression after initiation of interferon beta 1a treatment are related to level of depression 2 weeks before initiation of treatment. Physicians should assess history of depression for all patients in whom interferon beta-1a treatment is initiated. Patients with a recent history of depression are at risk for increased depression within 2 months after starting interferon beta-1a treatment, even though they may not be depressed at the time of treatment initiation. PMID- 10520944 TI - Psychiatric medication and abnormal behavior as predictors of progression in probable Alzheimer disease. AB - OBJECTIVE: To examine whether the use of psychiatric medication and the presence of abnormal behaviors affects the progression of Alzheimer disease. DESIGN: Cross sectional with longitudinal follow-up and the likelihood of arriving at 4 end points: (1) Mini-Mental State Examination score of 9 or lower; (2) Blessed Dementia Rating Scale score of 15 or higher for activities of daily living; (3) nursing home admission; and (4) death, evaluated using a proportional hazard model with 9 variables: psychosis, insomnia, wandering, aggression, psychomotor agitation, depression, and use of antidepressants, antipsychotic agents, or sedatives/hypnotics. SETTING: Multidisciplinary dementia research clinic. PATIENTS: We examined baseline and follow-up behavioral symptoms and the use of psychiatric medication in 179 mildly to moderately impaired patients with probable Alzheimer disease participating in a longitudinal study of dementia. Patients were observed from 2.4 to 172 months (mean duration +/- SD, 49.5+/-27.4 months). RESULTS: Nine patients (5%) were taking sedatives/ hypnotics; 16 (9%), antipsychotic agents; and 22 (12%), antidepressants at study entry. Patients taking antipsychotic agents had lower Mini-Mental State Examination scores and higher Blessed Dementia Rating Scale scores for activities of daily living than patients not taking any medication. Using proportional hazard analysis with time dependent covariates for individual psychiatric symptoms and medications, we found that the development of psychosis was associated with functional decline (time to Blessed Dementia Rating Scale score of > or =15), institutionalization, aggression, and agitation with functional decline after adjusting for age at study entry, education, Mini-Mental State Examination scores, and Blessed Dementia Rating Scale scores. Use of antipsychotic medication was associated with functional decline, and sedatives/hypnotics with death. Neither the presence of psychiatric symptoms nor use of medication was associated with rate of cognitive decline (time to Mini-Mental State Examination score of < or =9). CONCLUSIONS: These findings indicate that the use of antipsychotic agents and sedatives can affect the natural course of Alzheimer disease. Psychosis, agitation, and aggression are important predictors of outcome, even when the effects of medication to treat them is taken into account. PMID- 10520945 TI - Change in cognitive function in older persons from a community population: relation to age and Alzheimer disease. AB - OBJECTIVE: To examine change in cognitive function in older persons sampled from a community population, and its relation to age and Alzheimer disease. DESIGN: Prospective cohort study with an average of 3.5 years of follow-up. SETTING: East Boston, Mass--a geographically defined, urban, working-class community. PARTICIPANTS: A stratified, random sample of persons 65 years and older underwent uniform, structured clinical evaluation for Alzheimer disease. The 388 persons (89.2% of those eligible) who completed at least 1 annual follow-up evaluation were studied: 97 had Alzheimer disease at baseline; 95 developed Alzheimer disease during the study; and 196 were unaffected. OUTCOME MEASURES: Eight cognitive performance tests were administered, then converted to population weighted z scores and averaged to create a composite summary measure of cognitive function. Initial level of and change in this score were the outcome measures. RESULTS: In the population as a whole, many persons experienced a decline in cognitive performance, and age was related to both initial level and rate of decline. Analyses were conducted in 3 subgroups: persons with Alzheimer disease at baseline, those who developed Alzheimer disease during the study, and those who remained unaffected. In both Alzheimer disease subgroups, substantial cognitive decline was observed, but neither initial level nor rate of decline was related to age. In unaffected persons, little cognitive decline was evident, and there was a small, inverse association of age with initial level of cognitive function. CONCLUSION: In a general population sample, there was little evidence of cognitive decline during a 3.5-year period among persons who remained free of Alzheimer disease. PMID- 10520947 TI - A case of sporadic Pick disease with onset at 27 years. AB - BACKGROUND: Pick disease is an uncommon cause of dementia in middle age, and young-onset cases have rarely been reported. SETTING: A specialist hospital. PATIENT: Patient with onset of cognitive impairment at the age of 27 years whose cerebral biopsy specimen demonstrated Pick cells and tau-positive Pick bodies. CONCLUSION: Pick disease should be considered in the differential diagnosis of dementia in young adults. PMID- 10520946 TI - A novel type of hereditary motor and sensory neuropathy characterized by a mild phenotype. AB - BACKGROUND: Three loci for autosomal dominant hereditary motor and sensory neuropathy type I (HMSN I) or Charcot-Marie-Tooth disease type 1 (CMT1) have been identified on chromosomes 17p11.2 (CMT1A), 1q21-q23 (CMT1B), and 10q21.1-q22.1 (designated here as CMT1D). The genes involved are peripheral myelin protein 22 (PMP22), myelin protein zero (MPZ), and the early growth response element 2 (EGR2), respectively. Probably a fourth locus (CMT1C) exists since some autosomal dominant HMSN I families have been excluded for linkage with the CMT1A and CMT1B loci. Four loci for autosomal dominant hereditary motor and sensory neuropathy type II (HMSN II) or Charcot-Marie-Tooth disease type 2 (CMT2) have been localized on chromosomes 1p35-p36 (CMT2A), 3q13-q22 (CMT2B), 7p14 (CMT2D), and 3p (HMSN-P). OBJECTIVE: To describe the clinical, electrophysiologic, and neuropathological features of a novel type of Charcot-Marie-Tooth disease. PATIENTS AND METHODS: We performed linkage studies with anonymous DNA markers flanking the known CMT1 and CMT2 loci. Patients and their relatives underwent clinical neurologic examination and electrophysiologic testing. In the proband, a sural nerve biopsy specimen was examined. RESULTS: Linkage studies excluded all known CMT1 and CMT2 loci. The clinical phenotype is mild and almost all affected individuals remain asymptomatic. Electrophysiologic and histopathological studies showed signs of a demyelinating neuropathy, but the phenotype is unusual for either autosomal dominant HMSN I or HMSN II. CONCLUSION: Our findings indicate that the HMSN in this family represents a novel clinical and genetic entity. PMID- 10520948 TI - The pathology of shingles: Head and Campbell's 1900 monograph. AB - Shingles (herpes zoster) and postherpetic neuralgia, a chronic neuropathic pain syndrome that can persist after the shingles lesions heal, were studied by eminent neurologists of the 19th century. Autopsy studies were used to establish sensory neural pathways in the peripheral and central nervous systems. More recently, zoster and postherpetic neuralgia have served as models for the study of the pathogenesis and treatment of neuropathic pain. Postherpetic neuralgia has the cardinal clinical features of all neuropathic pain syndromes, including sensory abnormalities, ongoing pain, and allodynia (touch-induced pain). Unlike most other neuropathic pain syndromes, such as trigeminal neuralgia or nerve root compressions, shingles has a well-defined pathogenesis and onset, as well as visible lesions, and is therefore uniquely suitable for study. PMID- 10520949 TI - The mere presence of low levels of carboxyhemoglobin is not causal proof for altered neuropsychological performance. PMID- 10520950 TI - Therapeutic serum levels for new antiepileptic drugs. PMID- 10520951 TI - The role of intermolecular interactions: studies on model systems for bacterial biofilms. AB - The mechanical stability of biofilms and other microbial aggregates is of great importance for both the maintenance of biofilm processes and the removal of undesired biofilms. The binding forces are weak interactions such as London dispersion forces, electrostatic interactions and hydrogen bonds. In a first attempt to rank their contribution, the viscosity of solutions of extracellular polymeric substances (EPS) from a mucoid strain of Pseudomonas aeruginosa is measured. In order to distinguish the binding forces, substances are chosen which individually address the different types of bonds. Polyacrylic acid is identified as a suitable model system for EPS when molecular interactions are studied. Electrostatic interactions and hydrogen bonds are found to be the dominating forces among macromolecules within the biofilm. PMID- 10520952 TI - Dehydrophenylalanine analogues: conformational characterization. AB - Few dehydrophenylalanine (deltaPhe) analogues (X-deltaPhe-Phe-Gly-X1, X = Ac-; Boc-; Z-; X1 = OMe; OH; ONH2) of virus replication inhibiting peptide (Z-D-Phe Phe-Gly) were synthesized, and their solution conformations were investigated by 1H NMR, UV and circular dichroism (CD) spectroscopy. Homogeneity of these analogues was also assessed by reverse phase-high performance liquid chromatography (RP-HPLC) using water-acetonitrile gradients. PMID- 10520953 TI - Thermal features of the bovine serum albumin unfolding by polyethylene glycols. AB - Albumin showed very poor affinity for polyethylene glycol molecular weight (Mw) 1000 (30 M(-1)) and Mw 8000 (400 M(-1)) (PEG 1000 and PEG 8000). Polyethylene glycol of low Mw favours the ionization of the tyrosine (TYR) residues of albumin. Such variation might be a consequence of the change in dielectric constant at the domain of the protein by PEG binding. PEGs of high Mws stabilize the native compact state of human albumin showing negative preferential interaction with the protein. Interaction between PEGs and albumin is thermodynamically unfavourable, and becomes even more unfavourable for denatured proteins whose surface areas are larger than those of native ones leading to a stabilization of the unfolded state, which is manifested as a lowering of the thermal transition temperature. PEG 8000 perturbs the structure of the protein surface, partially modifying the layer of water and the microenvironment of the superficial aromatic residues (tryptophan, TRP and TYR) which is in agreement with the modifications of the UV spectrum of albumin by PEG 8000 and circular dichroism (CD) spectrum at high temperatures. PMID- 10520954 TI - Structural investigation of beta-lactoglobulin gelation in ethanol/water solutions. AB - The aggregation and gelation properties of beta-lactoglobulin (BLG), a globular protein from milk, was studied in hydro-ethanolic solutions (50/50% (v/v)) at room temperature. The phase state diagrams as a function of pH and ethanol concentration showed that a gel structure appeared after a period ranging from 1 min to 1 week depending on the physico-chemical conditions. The aggregation kinetics, studied by infrared spectroscopy and dynamical rheological measurements, highly depended upon the pH; the process being the fastest at pH 7. Alcohol-induced aggregation of BLG was characterized by the formation of intermolecular hydrogen bonded beta-sheet structures. Small angle neutron scattering indicated that the aggregates structures in the final gels were similar at pH 7, 8 and 9. Through the data obtained at the molecular and macroscopic levels, it can be concluded that the kinetics of gelation were pH dependent while the spatial arrangements of the aggregates were similar in the final structures. The heterogeneous structures formed in hydro-ethanolic gels could be analysed in terms of a phase separation, the syneresis being the final visible state. PMID- 10520955 TI - Linkage of proton binding to the thermal unfolding of Sso7d from the hyperthermophilic archaebacterium Sulfolobus solfataricus. AB - In this study the pH dependence of the thermal stability of Sso7d from Sulfolobus solfataricus is analyzed. This small globular protein of 63 residues shows a very marked dependence of thermal stability on pH: the denaturation temperature passes from 65.2 degrees C at pH 2.5 to 97.9 degrees C at pH 4.5. Analysis of the data points out that the binding of at least two protons is coupled to the thermal unfolding. By linking the proton binding to the conformational unfolding equilibrium, a thermodynamic model, which is able to describe the dependence upon the solution pH of both the excess heat capacity function and the denaturation Gibbs energy change for Sso7d, is developed. The decreased stability in very acid conditions is due to the binding of two protons on identical and noninteracting sites of the unfolded state. Actually, such sites are two carboxyl groups possessing very low pKa values in the native structure, probably involved in salt bridges on the protein surface. PMID- 10520956 TI - Comparison between the physicochemical behaviour of two microbial polysaccharides: RMDP17 and rhamsan. AB - This paper concerns the properties of two ionic polysaccharides with very close chemical structures. pH metric and conductimetric measurements showed that they behave similarly from a polyelectrolytic point of view. From optical rotation and differential scanning calorimetry (DSC) measurements, the two polymers probably adopt a double helical conformation which is destabilised by deacetylation. The main differences concern the stability of this ordered conformation and the ability of these double helices to associate to form gels. The results support a higher thermal stability of the ordered conformation for deacetylated RMDP17 (about 8 degrees C), whereas deacetylated rhamsan has a better ability to form gels. PMID- 10520957 TI - Antimicrobial films produced from chitosan. AB - Antimicrobial films were prepared by dissolving chitosan into hydrochloric, formic, acetic, lactic and citric acid solutions. Below 40 degrees C, the counter ions could be classified into two groups based on their effect on zero-shear-rate viscosity in 2% solutions of organic acids. Chloride and citrate produced solutions with much lower viscosities than formate, acetate and lactate. At higher temperatures, these differences vanished, and the activation energies of viscous flow were all similar between 40 and 60 degrees C. Films prepared from these solutions were evaluated in tension for Young's modulus, stress and elongation at yield and break points. Films made from hydrochloric, formic and acetic acids were hard and brittle, whereas those from lactic and citric acids were soft and could be stretched. Good correlation was found between Young's modulus and volume of the counter ion. Film properties are essentially governed by the volume of the counter ion and not by the interactions between this counter ion and the macromolecule. Results suggest that acetate has the maximum molecular volume above which the film strength decreases very rapidly. PMID- 10520958 TI - Gel--sol transition in kappa-carrageenan systems: microviscosity of hydrophobic microdomains, dynamic rheology and molecular conformation. AB - The effect of gel-sol transition in kappa-carrageenan systems on the microviscosity of hydrophobic microdomains, as well as its relation to macroscopic rheology and molecular conformation, was studied in kappa-carrageenan systems. The microdomains were probed by 1,3-di(-1-pyrenyl)propane (P3P) for which the excimer intensity (Ie) provides relative measures of the microviscosity in the immediate probe surroundings. In particular the applicability of P3P to monitor the gel--sol transition was proved, the results showing a dramatic decrease in microviscosity in the vicinity of the transition point. The corresponding changes in rheological properties and carrageenan conformation were investigated by dynamic viscometry (DV) and optical rotation (OR), respectively. The temperature of onset of the transition as indicated by the microviscosity data (T0) was found to correlate well with the OR and DV-results. The application of microviscosity and OR-measurements allowed an estimation of the helical content at T0 to be determined. P3P-data indicate a microenvironment viscosity for the probe sites in the kappa-carrageenan system comparable to that found in SDS micelles. PMID- 10520959 TI - The effect of gamma-irradiation on the temperature dependence of D.C. electrical conductivity of dry bone. AB - The effect of gamma-irradiation with doses from 10 to 500 kGy on the electrical conductivity (g) of dry bone was studied. Temperature measurement of electrical conductivity were made from 393 to 533 K. The dependence obtained indicates the increase in g with temperature. An increase in irradiation dose resulted in a decreased g value for each dose up to temperature 462 K. Temperature 462 K was interpreted as the temperature of collagen melting point in dry bone. Above 462 K, g values were dose independent. A dose of 500 kGy shifted the melting point to lower temperature. In addition, the activation energy for the charge conduction process was calculated. Obtained values for electrical conductivity and activation energy were typical for dielectrics and indicated degradation of the organic component of bone. PMID- 10520960 TI - Isolation and analysis of a novel acidic polysaccharide from the case of squid pen. AB - A new non-sulphated acidic polysaccharide with an average molecular mass of 55 kDa was isolated from squid pen case after papain digestion and beta-elimination. This polysaccharide contains mainly L-iduronic acid, D-glucuronic acid, D galactosamine, D-glucosamine and significant amounts of neutral sugars as glucose, galactose and fucose. The polysaccharide was not degraded to the relative disaccharides by chondroitinases ABC, AC and B, hyaluronidase and keratanase or by treatment with heparinases, suggesting a structure different from those of known glycosaminoglycans. The polysaccharide cannot form self aggregates. PMID- 10520961 TI - Failure to attract and retain clinician/scientist faculty puts our profession at risk. PMID- 10520962 TI - Neil Goldsworthy and his legacy: Sydney's Institute of Dental Research. AB - Dr. Neil Goldsworthy had a profound effect on the development of dental research in the biological sciences in Australia, including measures for the prevention of dental caries. He played a major role in the establishment of the Institute of Dental Research in Sydney in 1946 and was Director until his sudden death in 1960. Reviewing the Institute's activities provides the opportunity for his achievements to receive due recognition and to show how they influenced its subsequent development. As one who has been associated with the Institute for most of the last 50 years, it is an honor for me to undertake this task. PMID- 10520963 TI - Localization and changes in NADPH-diaphorase reactivity and nitric oxide synthase immunoreactivity in rat pulp following tooth preparation. AB - Inflammatory changes in the dental pulp are accompanied by release of a wide variety of chemical mediators. Nitric oxide, an oxidative free radical produced by the enzyme nitric oxide synthase (NOS), has been implicated in multiple inflammatory processes, which makes it a suitable marker for changes which likely occur following tooth pulp insult. Since limited information on nitric oxide in the pulp is available, it is necessary first to examine relative distributions of NOS in uninflamed and inflamed rat pulp. We accomplished this by characterizing regions of nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) activity and the distribution of both macrophage NOS (macNOS) and neuronal NOS (nNOS) immunoreactivity in normal and inflamed rat molar pulp at multiple time points. The results showed that: (1) deep cavity preparation on the mesial surface of the molar produced a time-dependent inflammation, with acute inflammation early progressing to chronic, granulomatous inflammation with necrosis later that spread preferentially down the mesial root; (2) control (non prepared) teeth showed a relatively faint and homogeneous distribution of NADPH-d and macNOS reactivity but no discernible nNOS reactivity; (3) inflamed teeth displayed localized increased intensity of NADPH-d and macNOS reactivity surrounding the inflamed area of pulp, but no increased nNOS activity; (4) pulp vessels supplying the inflamed area showed increased NADPH-d reactivity, but no increased macNOS or nNOS reactivity; and (5) neither NADPH-d, macNOS, nor nNOS reactivity was observed in pulpal nerves. Therefore, nitric oxide may mediate the pulpal inflammatory response through its effects on the paralesional pulp tissue and surrounding endothelial/vascular structures. PMID- 10520964 TI - Co-increase of nerve fibers and HLA-DR- and/or factor-XIIIa-expressing dendritic cells in dentinal caries-affected regions of the human dental pulp: an immunohistochemical study. AB - Neuro-immune interaction has been suggested to play some modulatory role in the immunodefense of the dentin/pulp complex. In this study, we performed a simultaneous immunohistochemical observation of neural elements and pulpal dendritic cells (PDCs) on human carious teeth, to obtain morphological evidence for neuro-immune interaction in response to dentinal tubule-derived carious stimuli. Human third molars bearing a pulp-exposure-free caries lesion were studied. Immunoperoxidase staining was performed with anti-HLA-DR, anti coagulation factor XIIIa, and anti-CD14 as PDC markers, and anti-low-affinity nerve growth factor receptor (NGFR), anti-protein gene products 9.5, and anti calcitonin gene-related peptide as nerve markers. The carious teeth usually exhibited localized accumulation of both PDCs and nerve fibers immunoreactive to each marker, in the para-odontoblastic region corresponding to the pulpal end of carious dentinal tubules. Semi-quantitative digital densitometry revealed that pixel numbers corresponding to factor-XIIIa- and NGFR-immunoreactivity were significantly higher in the carious regions than those in the non-carious regions of the same teeth as well as those in the corresponding regions of intact teeth. Classification of specimens with respect to caries depth showed that the co increase was most apparent in teeth with superficial caries. The increase of PDCs was less pronounced in carious teeth with reparative dentin. These findings suggest that both pulpal nerves and PDCs respond promptly and actively to dentinal tubule-derived carious stimuli. The synchronized accumulation of the two structures suggests an increased opportunity for neuro-immune interaction that may be of significance in the modulation of pathological processes in the dental pulp. PMID- 10520965 TI - Capsaicin induces cystatin S-like substances in submandibular saliva of the rat. AB - Irritating dietary substances such as tannin and papain have been reported to alter the morphology of salivary glands and their secretions. Such alterations can be one line of protection from toxic or irritating substances in food. We investigated the effects of dietary capsaicin (a pungent ingredient of hot red pepper) on the rat submandibular gland and its secretions. Several groups of animals were offered either control diets or diets containing capsaicin (from 0.0001 to 0.1%) for seven days. Higher concentrations suppressed food consumption for two days, after which only the highest concentration continued to reduce intake. The relative weight of the salivary glands in capsaicin-diet groups increased in a dose-dependent fashion, and new proteins appeared in the submandibular saliva. Chromatographic and electrophoretic properties of these proteins were identical or similar to those of isoproterenol-induced proteins. After affinity chromatography of the new protein fraction on a Cm-papain Sepharose 4B column, SDS-electrophoresis of the eluate revealed three major bands (15,500, 16,500, and 28,000 kDa). Hydrolysis of N-benzoyl-D,L-arginine-p nitroanilide by papain (a cysteine protease) decreased in the presence of the new protein fraction, suggesting that these proteins have cystatin-like activity (inhibition of cysteine protease). Denervation of the glossopharyngeal nerve suppressed induction of these proteins. The results suggest that dietary capsaicin induces cystatin S-like substances in submandibular saliva by stimulating the reflex arc involving the glossopharyngeal nerve. These proteins likely facilitate ingestion of diets containing the irritating substance. PMID- 10520966 TI - Autocrine regulation of osteoclast formation and bone resorption by IL-1 alpha and TNF alpha. AB - Bone resorption is regulated by the cytokines within marrow cells that mediate osteoclast formation and activation. IL-1 and TNF induce bone resorption by stimulating the production of osteoclast-like multinucleated cells and by increasing the bone-resorbing activity of formed osteoclasts. This study was designed to detect IL-1 and TNF in osteoclasts in vitro and to determine whether these cytokines up-regulate osteoclast differentiation and bone resorption. The production of IL-1 alpha, -beta, and TNF alpha, beta in osteoclasts was examined immunohistochemically and by in situ hybridization. In the co-culture of C57BL/6N mouse bone marrow and MC3T3-G2/PA6 cells, a colony of osteoclasts formed, and IL 1 alpha and TNF alpha were detected. However, IL-1 beta and TNF beta were not detected. To investigate the role of IL-1 alpha and TNF alpha from osteoclasts, we enumerated TRAP-positive cells and measured the resorption pit areas in the presence of antibodies against IL-1 alpha and TNF alpha. The addition of antibodies against IL-1 alpha and TNF alpha to the co-culture system decreased the number of TRAP-positive colonies at seven days after incubation (anti-IL-1 alpha, 25.0 +/- 2.3%; anti-TNF alpha, 41.7 +/- 3.7%; anti-IL-1 alpha + anti-TNF alpha, 40.5 +/- 1.3%; and control, 100%), and the ratio of mononuclear to multinuclear cells had changed (anti-IL-1 alpha, 90:10; anti-TNF alpha, 75:25; anti-IL-1 alpha+ anti-TNF alpha, 88:12; and control, 60:40). The total pit areas per dentin slice also decreased with the addition of antibodies (anti-IL-1 alpha, 28,828; anti-TNF alpha, 49,249; anti-IL-1 alpha + anti-TNF alpha, 30,685; and control, 303,139 mm2). These results suggest that local production of IL-1 alpha and TNF alpha by osteoclasts is an important mechanism for regulating the osteoclast differentiation and bone resorptive process. PMID- 10520968 TI - Establishment of oral anaerobes during the first year of life. AB - Anaerobic species constitute a significant part of the bacterial community of the mouth. Although the time and species involved in the primary colonization of infants are of great importance by forming the basis for further colonization, the development of the oral anaerobic microflora with age is still inadequately understood. In the present study, time and succession of colonization of oral anaerobes were longitudinally examined in 44 healthy Caucasian infants at 2, 6, and 12 months of age. Unstimulated saliva samples were quantitatively cultured on non-selective Brucella blood agar and several selective media for the isolation of anaerobic micro-organisms. The most frequent anaerobic finding in two-month old infants was Veillonella spp. The Prevotella melaninogenica group also represented early colonizing species, and the frequency increased remarkably during the first year of life, whereas the Prevotella intermedia group organisms seemed to be late colonizers. Fusobacterium nucleatum, non-pigmented Prevotella spp., and Porphyromonas catoniae were occasional findings in subjects at 2 months but frequent findings in those at one year of age. F. nucleatum was the most frequent strictly anaerobic species in one-year-old infants; other fusobacteria were also occasionally found. The frequency of facultative/micro-aerophilic corroding rods and Capnocytophaga spp. started to increase toward the end of the first year. Except for the common presence of facultative/micro-aerophilic Actinomyces spp., other anaerobic gram-positive species were only occasionally present in these infants. Once established, early-colonizing species tended to persist in the mouth. Our longitudinal study demonstrated the establishment of several anaerobic species with steadily increasing frequencies during the first year of life. PMID- 10520969 TI - Saliva, salivary micro-organisms, and oral health in the home-dwelling old elderly--a five-year longitudinal study. AB - High scores of chair-side salivary microbial tests have been found to be related to an increased prevalence and incidence of coronal and root caries. Many elderly face an increased risk of the growth of oral microbes, and previous studies have reported high salivary microbial counts in elderly populations. The aim of this follow-up study was to compare, at five-year intervals, stimulated salivary flow rates with the numbers of selected salivary micro-organisms (mutans streptococci, lactobacilli, and yeasts) in a group of home-dwelling elderly in Helsinki. A further aim was to study the influence of baseline microbial counts on five-year root caries increments and rates of tooth loss. The baseline study population was comprised of 270 subjects who were all participants in the population-based Helsinki Aging Study. Salivary flow rates and microbial conditions were determined as part of their dental examination at the Institute of Dentistry, University of Helsinki, in 1990-1991. Of these subjects, 110 underwent a follow up examination in 1995-1996. Commercially available kits (Dentocult SM strip mutans for mutans streptococci, Dentocult LB for lactobacilli, and Oricult N for yeasts) were used for the enumeration of micro-organisms, after the collection of paraffin-wax-stimulated whole saliva. The stimulated whole saliva flow rates of the subjects were significantly lower at the follow-up than at baseline (paired t test, difference -0.16 mL/min; p < 0.05), whereas buffer capacity was higher (paired t test, difference 0.19 on a three-unit scale; p < 0.05). Apart from lower salivary lactobacilli counts at follow-up (paired t test, difference -0.44 CFUs/mL of saliva; p < 0.001), no changes were found in salivary microbial levels. Salivary microbial counts were clearly associated with the subjects' dentition types: More denture-wearers had high microbial counts than persons with natural dentitions. None of the salivary factors correlated with the root caries incidence or the number of teeth lost during the five-year follow-up. PMID- 10520967 TI - Recombinant human bone morphogenetic protein-2 stimulates osteoblastic differentiation in cells isolated from human periodontal ligament. AB - Periodontal ligament cells may play an important role in the successful regeneration of the periodontium. We investigated the effects of recombinant human bone morphogenetic protein-2 (rhBMP-2), one of the most potent growth factors that stimulates osteoblast differentiation and bone formation, on cell growth and osteoblastic differentiation in human periodontal ligament cells (HPLC) isolated from four adult patients. rhBMP-2 induced no significant changes in cell growth in any of the HPLCs. rhBMP-2 at concentrations over 50 ng/mL significantly stimulated alkaline phosphatase (ALPase) activity and parathyroid hormone (PTH)-dependent 3', 5'-cyclic adenosine monophosphate accumulation, which are early markers of osteoblast differentiation, in the HPLCs. rhBMP-2 (500 ng/mL) also slightly enhanced the level of PTH/PTH-related peptide receptor mRNA expression in these cells. While interleukin-1 beta enhanced ALPase activity stimulated with rhBMP-2, tumor necrosis factor-alpha inhibited the rhBMP-2 stimulated activity. Interleukin-6 induced no significant changes in ALPase activity stimulated with rhBMP-2. Although HPLCs, whether treated with rhBMP-2 or not, could not produce measurable amounts of osteocalcin, which is a marker of more mature osteoblasts, 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] induced osteocalcin mRNA expression and protein synthesis in these cells. rhBMP-2 inhibited 1,25(OH)2D3-induced osteocalcin synthesis in HPLCs at both the mRNA and protein levels. These results suggest that rhBMP-2 provides an anabolic effect on periodontal regeneration by stimulation of osteoblastic differentiation in human periodontal ligament cells, and that this stimulatory effect is differentially modulated by inflammatory cytokines during the course of periodontal regeneration. PMID- 10520970 TI - Fluoride release from restorative materials and its effects on dentin demineralization. AB - As the use of adhesive restorative materials has increased during the last several years, interest in adhesive materials that release fluoride has also grown. The purpose of this study was to measure fluoride release from several adhesive restorative materials and to evaluate its effect on dentin resistance to demineralization and on bacterial metabolism in a modified in vitro system. Standardized cavities (1.8 mm in diameter) were prepared in bovine teeth that had been ground to dentin. One cavity in each tooth was restored with one of the following restorative systems: (a) Single Bond and Z100; (b) Single Bond and Tetric Ceram; (c) Fuji Bond LC and Z100; (d) Fuji Bond LC and Tetric Ceram; (e) Fuji II LC; or (f) Fuji IX GP. The other cavity in each tooth was "restored" with wax as a control. For each restorative system, 12 specimens were evaluated for fluoride release during the first 24 hrs after restoration placement. Dentin adjacent to the restored sites was subjected to lactic acid challenge (pH 4.3) for 3 hrs, after which calcium release was measured. Another 12 specimens in each group were stored for 24 hrs in de-ionized water, and were exposed to an S. mutans suspension (1:1 THB/de-ionized water and 50 mM glucose, A660 = 0.2) for 6 hrs, followed by calcium release and pH measurement. Bulk specimens of each material were also made and stored in water. Fluoride released from Fuji Bond LC, Fuji IX GP, and Fuji II LC in bulk was significantly greater than from the other materials. In the restored dentin specimens, increased resistance to demineralization from a lactic acid challenge was directly related to fluoride release. The same effects were seen as a result of the S. mutans challenge. While fluoride release from restorative materials increased the resistance of dentin to demineralization in this system, the clinical relevance of the findings is not known. PMID- 10520972 TI - Smoothness of human jaw movement during chewing. AB - Human limb movements are successfully modeled based on the assumption that the central nervous system controls the movements by maximizing movement smoothness. Movement smoothness is quantified by means of a time integral of squared jerk (jerk-cost), where jerk is defined as the rate of change in acceleration. This study was performed to investigate whether the control of human masticatory vertical jaw movements can also be explained by a minimum-jerk (maximum smoothness) model. Based on the assumption that minimum-jerk models account for vertical jaw-opening and -closing movements during chewing, the actual time profile of the movement trajectory was simulated by the model. The simulated jerk costs and peak velocities were compared with those obtained by actual measurements of jaw movements during chewing. Jerk-costs and peak velocities of the jaw movements during chewing were significantly correlated with those predicted by minimum-jerk models (P < 0.0001, r between 0.596 and 0.799). The minimum-jerk models predicted closing movement trajectories more accurately than opening movement trajectories (jaw opening, root-mean-square error = 1.19 mm; jaw closing, 0.52 mm, t = 4.375, P < 0.0001). The results indicated that the vertical jaw movement control during chewing was represented by the minimum-jerk control model and that the vertical jaw-closing movement is smoother than the opening movement during gum-chewing. PMID- 10520973 TI - Molecular neurobiology for practicing psychiatrists, Part 1: overview of gene activation by neurotransmitters. PMID- 10520971 TI - Variations in dental anxiety among middle-aged and elderly women in Sweden: a longitudinal study between 1968 and 1996. AB - Cross-sectional studies have shown that older individuals are significantly less dentally anxious than younger ones. However, research has not been able to show if this is a cohort effect or an effect of fear declining with age. If it is a cohort effect, dental anxiety among the elderly may pose a greater-than-expected problem for the providers of dental services. With the exception of longitudinal studies in children and a three-year follow-up on adults, no truly longitudinal epidemiological studies concerning dental anxiety have been performed. The aim of this project was to investigate how dental anxiety changes with aging. In a longitudinal population study of women in Goteborg, Sweden, starting in 1968, 1462 women aged 38 to 54 participated. A representative subsample of 778 women took part in a psychiatric examination where an investigation of dental anxiety was included. The same questions were also included when these women were re examined in 1974, 1992, and 1996. Three hundred seventy-five women were still eligible for investigation in 1996. In 1968-69, 48 (12.8%) of the participating women assessed themselves as "very afraid" or "terrified" when visiting the dentist, and in 1996 the frequency was 21 (5.6%) among the same women. In 1968 69, 180 women (48%) reported no dental anxiety when visiting the dentist, and 28 years later the frequency was 230 (61%). In the three youngest age groups, dental anxiety decreased significantly (p < 0.001) over the 28-year period. Older compared with younger women reported significantly less dental anxiety, and this was an age effect rather than a cohort effect. Thus, this longitudinal study supported the hypothesis that dental fear, like many other general and specific phobias, declines with age. PMID- 10520974 TI - Antidepressant medication change in a clinical treatment setting: a comparison of the effectiveness of selective serotonin reuptake inhibitors. AB - BACKGROUND: This investigation focuses on the 3 most frequently used selective serotonin reuptake inhibitors (SSRIs) (paroxetine, fluoxetine, sertraline) and examines the rate of medication switches as a measure of effectiveness. We answer 2 questions: (1) What is the likelihood that a patient starting treatment with an SSRI will complete treatment with the same agent? and (2) Depending on the initial SSRI agent used, do patients switch at different frequencies? METHOD: A retrospective chart review was performed on 2779 patients treated in a university outpatient clinic from March 1995 to January 1997. Of these, 263 patients given antidepressants were randomly selected: 214 were prescribed SSRIs; 24, novel antidepressants; and 25, tricyclic antidepressants. RESULTS: There was no significant difference in rate of switching between the different classes of antidepressant (p = .1) nor between drugs within the SSRI class (p = .513). When medication change was the independent factor, significant differences between the groups were total time in treatment and number of visits (p < .001 and p = .011, respectively). Age, education, and Clinical Global Impressions-Severity of Illness scale scores (admission, discharge, and change) were not significantly different between the groups. CONCLUSION: Approximately 25% of patients started with an SSRI will switch to another antidepressant in the course of their treatment. The SSRIs appear to be equivalent in effectiveness. They are not interchangeable, because patients who discontinue one SSRI for lack of tolerability or response can generally be treated effectively with another. PMID- 10520975 TI - Efficacy of fluvoxamine in the treatment of major depression with comorbid anxiety disorders. AB - BACKGROUND: Major depression with comorbid anxiety disorder is associated with poor antidepressant outcome compared with major depression without comorbid anxiety disorder. The purpose of our study was to assess changes in depressive symptoms and anxiety levels in outpatients with major depression with comorbid anxiety disorder following 12 weeks of open treatment with fluvoxamine. METHOD: We enrolled 30 outpatients (mean +/- SD age = 39.4 +/- 11.3 years; 16 women and 14 men) with DSM-IV major depressive disorder accompanied by one or more current comorbid DSM-IV anxiety disorders in our study. Patients were treated openly with fluvoxamine initiated at 50 mg/day, with an upward titration to a maximum of 200 mg/day (mean +/- SD dose = 143 +/- 45 mg/day). Efficacy assessments included the 17-item Hamilton Rating Scale for Depression (HAM-D-17) and Clinical Global Impressions-Severity of Illness (CGI-S) and Improvement (CGI-I) scales for both depression and anxiety. Intent-to-treat analysis was used to assess outcome. RESULTS: The mean +/- SD number of comorbid anxiety disorders per patient was 2.1 +/- 1.1. Following fluvoxamine treatment, the mean +/- SD HAM-D-17 score dropped from 20.2 +/- 3.3 to 1 1.0 +/- 7.0 (p < .0001). The mean +/- SD depression CGI-S score dropped from 4.0 +/- 0.6 to 2.4 +/- 1.1 (p < .0001), and the mean +/- SD anxiety CGI-S score decreased from 4.1 +/- 0.8 to 2.5 +/- 1.2 (p < .0001). Eighteen (60%) of the 30 patients had CGI-I scores < or = 2 for both anxiety and depression at endpoint, with 53% showing a > or = 50% reduction in HAM-D-17 scores at endpoint. CONCLUSION: Although preliminary, our findings suggest that fluvoxamine is effective in treating outpatients with major depression with comorbid anxiety disorder, having a significant effect on both depression and anxiety symptoms. Further double-blind, placebo-controlled trials are needed, in a larger sample, to confirm our findings. PMID- 10520977 TI - Hoarding in obsessive-compulsive disorder: a report of 20 cases. AB - BACKGROUND: We describe the demographic characteristics, hoarding phenomenology, comorbid disorders, family histories, and treatment response of 20 adult obsessive-compulsive disorder (OCD) patients exhibiting hoarding behavior. METHOD: We utilized the Structured Clinical Interview for DSM-III-R, the Yale Brown Obsessive Compulsive Scale, and a semistructured interview to gather data. RESULTS: We studied 9 women and 11 men. Their hoarding began from age 5 years to age 46 years (mean +/- SD age at onset = 20 +/- 11 years); hoarding was evident before the onset of other OCD symptoms in 9 patients. The most commonly hoarded items were newspapers and magazines, junk mail, old clothes, notes or lists, and old receipts. Hoarded material occupied from one room plus most or all closets to more than one room plus all closets, the garage, and yard. Seven patients rented additional storage space for hoarded items. Eighty-four percent of patients reported a family history of hoarding, and 80% grew up in a household where someone else hoarded. The most frequent primary motives for hoarding were fears of discarding something useful and discarding something that would be needed in the future. Lifetime prevalence of major depression and of impulse-control disorders, especially compulsive shopping, were high; only 3 patients met DSM-IV criteria for obsessive-compulsive personality disorder. Response of hoarding to selective serotonin reuptake inhibitors was less robust than is expected for obsessive-compulsive disorder. CONCLUSION: Whether hoarding behaviors mark a subset of obsessive-compulsive disorder patients with a different pathophysiology or functional anatomy deserves investigation. PMID- 10520976 TI - Cognitive-behavioral therapy as an adjunct to serotonin reuptake inhibitors in obsessive-compulsive disorder: an open trial. AB - BACKGROUND: We report the results of an open trial of cognitive-behavioral therapy (CBT) using exposure and ritual prevention as an adjunct to serotonin reuptake inhibitors (SRIs) in obsessive-compulsive disorder (OCD). We hypothesized that exposure and ritual prevention would significantly reduce OCD symptoms in patients who remained symptomatic despite an adequate trial of an SRI and enable patients to discontinue their medication. METHOD: OCD patients taking an adequate dose of an SRI > or = 12 weeks who remained symptomatic (i.e., a Yale Brown Obsessive Compulsive Scale [Y-BOCS] score > or = 16) were eligible. While taking a stable dose of an SRI, patients received 17 sessions of exposure and ritual prevention. For the intent-to-treat group, the paired t test was used to compare scores on the Y-BOCS, the National Institute of Mental Health (NIMH) Global OCD scale, the Clinical Global Impressions scale, and the Hamilton Rating Scale for Depression before and after exposure and ritual prevention. RESULTS: Six of 7 eligible patients entered the study, and 5 completed it. All 6 improved on all OCD measures. The mean +/- SD Y-BOCS score was 23.8 +/- 2.6 prior to exposure and ritual prevention and 12.2 +/- 4.3 after it (p < .001). The mean percentage decrease on the Y-BOCS was 49% (range, 26%-61%). Patients were rated by the therapist and rated themselves as much (N = 4) or very much (N = 2) improved. Blood drug levels did not change in most patients during exposure and ritual prevention; thus, the improvement was attributed to this type of therapy. No patients discontinued their medication. CONCLUSION: This open trial suggests that CBT using exposure and ritual prevention can lead to a significant reduction in OCD symptoms in patients who remain symptomatic despite an adequate trial of an SRI. PMID- 10520978 TI - Naltrexone in the treatment of dissociative symptoms in patients with borderline personality disorder: an open-label trial. AB - BACKGROUND: Dissociative phenomena, including flashbacks, are common in patients with borderline personality disorder and posttraumatic stress disorder (PTSD). Although dissociative symptoms can be severe and may interfere with psychotherapy, there is no established pharmacotherapy for these symptoms. Evidence suggests that alterations of the endogenous opiate system contribute to dissociative symptoms in patients with borderline personality disorder and PTSD. METHOD: We treated 2 groups of female borderline personality disorder patients (N = 13, with an overlap of 5 patients between the 2 groups; all met the diagnostic criteria of DSM-IV and the revised Diagnostic Interview for Borderline Patients) who experienced prominent dissociative phenomena including flashbacks with the nonselective opiate receptor antagonist naltrexone, 25 to 100 mg q.i.d., for at least 2 weeks. A self-rated questionnaire measuring dissociation, analgesia, tonic immobility, and tension (DAISS) was applied to 9 patients, who completed it for 7 consecutive days before and during treatment with naltrexone. In addition, 9 patients (with an overlap of 5 patients from the other group) completed a flashback protocol. RESULTS: DAISS scores reflected a highly significant reduction of the duration and the intensity of dissociative phenomena and tonic immobility as well as a marked reduction in analgesia during treatment with naltrexone. Six of 9 patients reported a decrease in the mean number of flashbacks per day. CONCLUSION: These observations support the hypothesis that an increased activity of the opioid system contributes to dissociative symptoms, including flashbacks, in borderline personality disorder and suggest that these symptoms may respond to treatment with opiate antagonists. In view of these results, a placebo-controlled, double-blind study to assess the potential benefit of naltrexone in a more rigorous way appears justified. PMID- 10520979 TI - Efficacy, safety, and gradual discontinuation of clonazepam in panic disorder: a placebo-controlled, multicenter study using optimized dosages. AB - BACKGROUND: The purpose of this multicenter, double-blind, placebo-controlled study was to evaluate the efficacy and safety of optimized dosages of clonazepam for the treatment of panic disorder and assess the tolerability of a schedule for gradual discontinuation. METHOD: Adult patients with panic disorder with or without agoraphobia (DSM-III-R criteria) were randomly assigned to receive either placebo or clonazepam in individually adjusted doses over 3 weeks to approximate an optimal dosage, which was then maintained for an additional 3 weeks, amounting to a 6-week therapeutic phase. The daily dose range was 0.25 to 4.0 mg administered in 2 divided doses. In the following 7-week discontinuance phase, the doses were tapered gradually to cessation. RESULTS: At the therapeutic endpoint, clonazepam (N = 222) proved clinically and statistically superior to placebo (N = 216) in change in the number of panic attacks and in Clinical Global Impressions-Severity of Illness (CGI-S) and CGI-Change scores, Patient's Global Impression of Change scores, amount of fear and avoidance associated with phobic symptoms, and duration of anticipatory anxiety. The gradual tapering of clonazepam was not associated with symptoms suggestive of withdrawal syndrome. Although patients taking clonazepam experienced some clinical worsening compared with the status achieved at endpoint, particularly in terms of number of panic attacks, no deterioration was observed using their condition at baseline as point of reference. No overall evidence of rebound was found. All regimens were generally well tolerated. Somnolence was the main adverse event associated with clonazepam therapy. The percentage of patients who reported adverse events was higher in the clonazepam group than in the placebo group, as was the mean number of adverse events per patient. CONCLUSION: In this placebo-controlled trial, clonazepam was an efficacious and safe shortterm treatment of the symptoms of panic disorder. Discontinuance during and after slow tapering was well tolerated. PMID- 10520980 TI - Do depressed subjects who have failed both fluoxetine and a tricyclic antidepressant respond to the combination? AB - BACKGROUND: Recent evidence suggests that the combination of fluoxetine and desipramine may provide a rapid and effective treatment for depression. METHOD: The current study evaluated 13 subjects with DSM-III-R nonpsychotic major depression who had previously failed either desipramine or imipramine and who were currently unsuccessfully treated with fluoxetine. Desipramine or imipramine was added to fluoxetine and Hamilton Rating Scale for Depression (HAM-D) scores, Beck Depression Inventory (BDI) scores, and plasma tricyclic levels were monitored for 3 weeks. RESULTS: Of the 13 subjects, 7 (54%) had a greater than 40% decline in HAM-D scores and 4 of these (31%) had 50% or greater decline in HAM-D. At week 3, responders (767 +/- 282 nmol/L) had a significantly higher mean tricyclic level as compared with nonresponders (515 +/- 95 nmol/L, F = 25.1, p < .0001), and change in BDI scores was significantly correlated with tricyclic level (r = -0.60, p < .05). CONCLUSION: These findings suggest that in some subjects the positive clinical effect of combining fluoxetine and a tricyclic antidepressant may be related to the plasma levels of the tricyclic compound. PMID- 10520981 TI - A case report of reduction in alcohol craving and protection against alcohol withdrawal by gabapentin. PMID- 10520982 TI - The nosology of compulsive skin picking. PMID- 10520983 TI - Transient syncope and ECG changes associated with the concurrent administration of clozapine and diazepam. PMID- 10520984 TI - Treatment of inhalant abuse with risperidone. PMID- 10520985 TI - The role of norepinephrine in the treatment of depression. PMID- 10520987 TI - The ties that bind: localization of the sister-chromatid cohesin complex on yeast chromosomes. PMID- 10520986 TI - Anger attacks: correlates and significance of an underrecognized symptom. AB - BACKGROUND: Anger attacks over provocations described as trivial by the individual are an underrecognized symptom associated with aggressive acts. They are usually followed by guilt and regret. Anger attacks among mothers are an important problem because they are often directed at the woman's spouse and/or children. This study examines the prevalence and correlates of anger attacks in a psychiatric clinic for women who are either pregnant or up to 18 months postpartum. METHOD: Fifty consecutive consenting patients were assessed at initial presentation with the Structured Clinical Interview for DSM-IV Axis I Disorders, a modified Anger Attacks Questionnaire, self-reports of psychiatric symptoms and psychosocial variables, and clinician ratings. RESULTS: Thirty (60%) of 50 patients reported anger attacks. Of those with anger attacks, 76.7% worried about them, and 73.3% had tried to prevent them. Compared with women without anger attacks, those with anger attacks were significantly more likely to report higher state and trait anger (p < .001), have a diagnosis of unipolar depression (p < .01), report more aggression directed at immediate family, and avoid their children. Both groups displayed little angry affect in the interview, thus appearing similar at assessment. CONCLUSION: Anger attacks in response to children and spouse were common in this group of women and were associated with subjective distress. Because those with and without anger attacks appear similar at interview, inquiring about the presence of anger attacks is important to ensure that they become a focus of treatment. PMID- 10520988 TI - "Kissin' cousins": intimate plasma membrane-ER interactions underlie capacitative calcium entry. PMID- 10520989 TI - Last but not least: regulated poly(A) tail formation. PMID- 10520990 TI - A novel role for 3-O-sulfated heparan sulfate in herpes simplex virus 1 entry. AB - Herpes simplex virus type 1 (HSV-1) binds to cells through interactions of viral glycoproteins gB and gC with heparan sulfate chains on cell surface proteoglycans. This binding is not sufficient for viral entry, which requires fusion between the viral envelope and cell membrane. Here, we show that heparan sulfate modified by a subset of the multiple D-glucosaminyl 3-O-sulfotransferase isoforms provides sites for the binding of a third viral glycoprotein, gD, and for initiation of HSV-1 entry. We conclude that susceptibility of cells to HSV-1 entry depends on (1) presence of heparan sulfate chains to which virus can bind and (2) 3-O-sulfation of specific glucosamine residues in heparan sulfate to generate gD-binding sites or the expression of other previously identified gD binding receptors. PMID- 10520991 TI - CCR7 coordinates the primary immune response by establishing functional microenvironments in secondary lymphoid organs. AB - The proper function of immune surveillance requires well-coordinated mechanisms in order to guide the patrolling immune cells through peripheral tissues and into secondary lymphoid organs. Analyzing gene-targeted mice, we identified the chemokine receptor CCR7 as an important organizer of the primary immune response. CCR7-deficient mice show severely delayed kinetics regarding the antibody response and lack contact sensitivity and delayed type hypersensitivity reactions. Due to the impaired migration of lymphocytes, these animals reveal profound morphological alterations in all secondary lymphoid organs. Upon activation, mature skin dendritic cells fail to migrate into the draining lymph nodes. Thus, in order to bring together lymphocytes and dendritic cells to form the characteristic microarchitecture of secondary lymphoid organs, CCR7 is required to rapidly initiate an adoptive immune response. PMID- 10520992 TI - Cosuppression of nonhomologous transgenes in Drosophila involves mutually related endogenous sequences. AB - Cosuppression refers to the phenomenon in which silencing among dispersed homologous genes occurs. Here we demonstrate that two nonhomologous reciprocal fusion genes, white-Alcohol dehydrogenase (w-Adh) and Adh-w, exhibit cosuppression using the endogenous Adh sequence as an intermediary. Deletion of the endogenous Adh gene eliminates the interaction, while reintroduction of an 8.6 kb Adh fragment restores the silencing. Using truncated Adh constructs, a nontranscribed segment in the Adh regulatory region was found to be one of the sequences required for homology recognition. The silencing interaction is initiated during early development. The silenced transgenes are associated with the Polycomb group complex of chromatin proteins. PMID- 10520993 TI - Targeted mutagenesis of Tsix leads to nonrandom X inactivation. AB - During X inactivation, mammalian female cells make the selection of one active and one inactive X chromosome. X chromosome choice occurs randomly and results in Xist upregulation on the inactive X. We have hypothesized that the antisense gene, Tsix, controls Xist expression. Here, we create a targeted deletion of Tsix in female and male mouse cells. Despite a deficiency of Tsix RNA, X chromosome counting remains intact: female cells still inactivate one X, while male cells block X inactivation. However, heterozygous female cells show skewed Xist expression and primary nonrandom inactivation of the mutant X. The ability of the mutant X to block Xist accumulation is compromised. We conclude that Tsix regulates Xist in cis and determines X chromosome choice without affecting silencing. Therefore, counting, choice, and silencing are genetically separable. Contrasting effects in XX and XY cells argue that negative and positive factors are involved in choosing active and inactive Xs. PMID- 10520994 TI - Plexin-neuropilin-1 complexes form functional semaphorin-3A receptors. AB - Class 1 and 3 semaphorins repulse axons but bind to different cell surface proteins. We find that the two known semaphorin-binding proteins, plexin 1 (Plex 1) and neuropilin-1 (NP-1), form a stable complex. Plex 1 alone does not bind semaphorin-3A (Sema3A), but the NP-1/Plex 1 complex has a higher affinity for Sema3A than does NP-1 alone. While Sema3A binding to NP-1 does not alter nonneuronal cell morphology, Sema3A interaction with NP-1/Plex 1 complexes induces adherent cells to round up. Expression of a dominant-negative Plex 1 in sensory neurons blocks Sema3A-induced growth cone collapse. Sema3A treatment leads to the redistribution of growth cone NP-1 and plexin into clusters. Thus, physiologic Sema3A receptors consist of NP-1/plexin complexes. PMID- 10520995 TI - Plexins are a large family of receptors for transmembrane, secreted, and GPI anchored semaphorins in vertebrates. AB - In Drosophila, plexin A is a functional receptor for semaphorin-1a. Here we show that the human plexin gene family comprises at least nine members in four subfamilies. Plexin-B1 is a receptor for the transmembrane semaphorin Sema4D (CD100), and plexin-C1 is a receptor for the GPI-anchored semaphorin Sema7A (Sema K1). Secreted (class 3) semaphorins do not bind directly to plexins, but rather plexins associate with neuropilins, coreceptors for these semaphorins. Plexins are widely expressed: in neurons, the expression of a truncated plexin-A1 protein blocks axon repulsion by Sema3A. The cytoplasmic domain of plexins associates with a tyrosine kinase activity. Plexins may also act as ligands mediating repulsion in epithelial cells in vitro. We conclude that plexins are receptors for multiple (and perhaps all) classes of semaphorins, either alone or in combination with neuropilins, and trigger a novel signal transduction pathway controlling cell repulsion. PMID- 10520996 TI - MT1-MMP-deficient mice develop dwarfism, osteopenia, arthritis, and connective tissue disease due to inadequate collagen turnover. AB - MT1-MMP is a membrane-bound matrix metalloproteinase (MT-MMP) capable of mediating pericellular proteolysis of extracellular matrix components. MT1-MMP is therefore thought to be an important molecular tool for cellular remodeling of the surrounding matrix. To establish the biological role of this membrane proteinase we generated MT1-MMP-deficient mice by gene targeting. MT1-MMP deficiency causes craniofacial dysmorphism, arthritis, osteopenia, dwarfism, and fibrosis of soft tissues due to ablation of a collagenolytic activity that is essential for modeling of skeletal and extraskeletal connective tissues. Our findings demonstrate the pivotal function of MT1-MMP in connective tissue metabolism, and illustrate that modeling of the soft connective tissue matrix by resident cells is essential for the development and maintenance of the hard tissues of the skeleton. PMID- 10520997 TI - The structure of the ligand-binding domain of neurexin Ibeta: regulation of LNS domain function by alternative splicing. AB - Neurexins are expressed in hundreds of isoforms on the neuronal cell surface, where they may function as cell recognition molecules. Neurexins contain LNS domains, folding units found in many proteins like the G domain of laminin A, agrin, and slit. The crystal structure of neurexin Ibeta, a single LNS domain, reveals two seven-stranded beta sheets forming a jelly roll fold with unexpected structural similarity to lectins. The LNS domains of neurexin and agrin undergo alternative splicing that modulates their affinity for protein ligands in a neuron-specific manner. These splice sites are localized within loops at one edge of the jelly roll, suggesting a distinct protein interaction surface in LNS domains that is regulated by alternative splicing. PMID- 10520999 TI - Physical activity intervention studies with health-related outcomes: some issues. PMID- 10520998 TI - Inhibiting HIV-1 entry: discovery of D-peptide inhibitors that target the gp41 coiled-coil pocket. AB - The HIV-1 gp41 protein promotes viral entry by mediating the fusion of viral and cellular membranes. A prominent pocket on the surface of a central trimeric coiled coil within gp41 was previously identified as a potential target for drugs that inhibit HIV-1 entry. We designed a peptide, IQN17, which properly presents this pocket. Utilizing IQN17 and mirror-image phage display, we identified cyclic, D-peptide inhibitors of HIV-1 infection that share a sequence motif. A 1.5 A cocrystal structure of IQN17 in complex with a D-peptide, and NMR studies, show that conserved residues of these inhibitors make intimate contact with the gp41 pocket. Our studies validate the pocket per se as a target for drug development. IQN17 and these D-peptide inhibitors are likely to be useful for development and identification of a new class of orally bioavailable anti-HIV drugs. PMID- 10521000 TI - Upper extremity kinematics and body roll during preferred-side breathing and breath-holding front crawl swimming. AB - Front crawl swimmers often restrict the number of breaths they take during a race because of the possible adverse effects of the breathing action on resistance or stroke mechanics. The aim of this study was to determine whether differences exist in the kinematics of the trunk and upper extremity used during preferred side breathing and breath-holding front crawl swimming. Six male swimmers performed trials at their 200 m race pace under breathing and breath-holding conditions. The underwater arm stroke was filmed from the front and side using video cameras suspended over periscope systems. Video recordings were digitized at 50 Hz and the three-dimensional coordinates of the upper extremity obtained using a direct linear transformation algorithm. Body roll angles were obtained by digitizing video recordings of a balsa wood fin attached to the swimmers' backs. The swimmers performed the breathing action without any decrement in stroke length (mean +/- s: breathing 2.24 +/- 0.27 m; breath-holding 2.15 +/- 0.22 m). Stroke widths were similar in the breathing (0.28 +/- 0.07 m) and breath-holding (0.27 +/- 0.07 m) trials, despite swimmers rolling further when taking a breath (66 +/- 5 degrees) than when not (57 +/- 4 degrees). The timing of the four underwater phases of the stroke was also unaffected by the breathing action, with swimmers rolling back towards the neutral position during the insweep phase. In conclusion, the results suggest that front crawl swimmers can perform the breathing action without it interfering with their basic stroke parameters. The insweep phase of the stroke assists body roll and not vice versa as suggested in previous studies. PMID- 10521001 TI - Test of performances strategies: development and preliminary validation of a comprehensive measure of athletes' psychological skills. AB - We report the initial stages of validation of the 64-item Test of Performance Strategies, a self-report instrument designed to measure the psychological skills and strategies used by athletes in competition and during practice. Data were obtained from a sample of 472 athletes competing across a range of performance standards in a wide variety of sports. Exploratory factor analyses of their responses produced eight competition strategy subscales and eight practice strategy subscales, each consisting of four items. Internal consistencies of the subscales ranged from 0.66 to 0.81 (x = 0.75). Correlations among strategies were examined within and between performance contexts. Subgroups defined by age, sex and current standard of performance in sport differed significantly in their psychological skills and strategies. PMID- 10521002 TI - Development and initial validation of an instrument to assess the motivational qualities of music in exercise and sport: the Brunel Music Rating Inventory. AB - Equivocal results of the psychophysical effects of music have been explained in part by the insensitivity of researchers to important personal and situational variables when selecting music. The aim of the present study was to operationalize a conceptual framework for the prediction of psychophysical responses to music into a music rating inventory to assess the motivational qualities of music in exercise and sport environments. An initial item pool was developed and administered to 334 aerobics instructors. Exploratory factor analysis produced a 13-item, four-factor structure (association, musicality, cultural impact and rhythm response), which accounted for 59.2% of the variance. This model demonstrated acceptable fit indices when tested using confirmatory factor analysis on 314 exercise-to-music participants, and was better than an alternative two-factor model. When cross-validated using multisample confirmatory factor analysis, the model also showed an acceptable fit overall, although some invariance in the rhythm response factor was evident that can be attributed to the exclusive use of synchronous music by aerobics instructors. The Brunel Music Rating Inventory appears to be a valid and reliable tool for both researchers and practitioners to assess the motivational qualities of music in exercise and sport environments. PMID- 10521003 TI - Stability of questionnaire items in sport and exercise psychology: bootstrap limits of agreement. AB - We describe an alternative method for assessing the stability of individual questionnaire items in sport and exercise psychology. To date, the Pearson product-moment correlation coefficient has been widely used in psychometrics. We propose an alternative non-parametric method based on proportion of agreement. Ninety-two male university students completed the revised 9-item Social Physique Anxiety Scale on two occasions, separated by a 2-week interval. Point estimates of the proportion of direct within-individual agreement between the two occasions were calculated separately for each item of the Social Physique Anxiety Scale. Estimates of uncertainty of the agreement were calculated using a bootstrapping resampling technique. For each item, 2000 bootstrap samples (each n = 92 pairs) were redrawn from the original sample. The sample statistic was calculated for each bootstrap sample to provide a bootstrap sampling distribution. The 95% confidence intervals (CI) were then calculated using the percentile method. The three most problematic items were items 7, 8 and 10 (as labelled in the original 12-item scale). These items demonstrated an agreement of 0.46 (95% CI= 0.36 0.56), 0.42 (95% CI = 0.33-0.52) and 0.41 (95% CI = 0.32-0.51) respectively. Our proposed method measures absolute agreement between test-retest responses, is free of normal assumptions, does not depend on high between-individuals variance, and can be applied successfully to individual items in the development of psychological tests. PMID- 10521004 TI - Athletic injury, psychosocial factors and perceptual changes during stress. AB - In this study, we measured changes in state anxiety, visual perception and reaction time during stress among 196 collegiate athletes participating in 10 sports. The athletes also completed measures of life events and social support at the beginning of their seasons. Measures of life events stress, social support, perceptual changes and changes in reaction time during stress were used as predictors of the number of injuries. For the entire sample, the only significant predictor of injury was negative life events stress (R = 0.45, P < 0.001). Following the suggestions of Smith et al., simple correlations were performed for those with least social support (bottom 33%, n = 65). Among this group, those individuals with more negative life events and greater peripheral narrowing during stress incurred more injuries than those with the opposite profile. Our findings are in line with the model of Andersen and Williams, in that those individuals who were low in a variable that buffers stress responsivity (i.e. social support), their negative life events and peripheral narrowing under stress (large and medium effect sizes, respectively) were substantially related to their number of injuries. PMID- 10521005 TI - The quantification of joint laxity in dancers and gymnasts. AB - The aim of this study was to determine the range of movement in gymnastic and dance populations. Sixty-five participants (41 females, 24 males; mean age 21.4 years) were assessed. The sample included dancers and gymnasts ranging from novice and club standard to international and professional status. Non specialized physical education students acted as controls. Range of movement was measured at the shoulders, hips, lumbar spine and ankles using a Loebl hydrogoniometer, and inherent joint laxity was assessed using Beighton and coworkers' adaptation of the Carter and Wilkinson 9-point scale. The right and left sides of the body were assessed and measures of active and passive motion were recorded. A graded increase in laxity was observed from controls, through novice gymnasts, to dancers and finally international gymnasts. The greater laxity of females than males was also confirmed. Dancers and gymnasts had a greater passive range of movement in all joints, which was partly inherited and partly acquired. There was a large difference between their active and passive ranges, which appeared to render the joints unstable. PMID- 10521006 TI - Dennis Patrick Cantwell, M.D.: who taught us how to fish. PMID- 10521007 TI - Psychotropic medication treatment patterns among school-aged children in foster care. AB - This study describes the level of psychotropic medication use and patterns of such treatment among school-aged children in foster care. Structured survey interviews were conducted in the foster homes of 302 randomly selected children, aged 6-12 years, who were living in foster care for 6 months or more and placed from three county service areas. Follow-up mental health assessments using the existing system of care format were completed on 255 children. Sixteen percent of these school-aged children in foster care were found to have taken psychotropic medication during their lifetime. The most common class of medication used in the past year was stimulants (62%). Children who were older, from Caucasian and biracial backgrounds, and who lived in a group home more likely to have taken psychotropic medication in the past year. Among those children who received a clinician diagnosis of a severe psychiatric disorder for which medication is an accepted component of care, boys were more likely to receive medication treatment than girls. Sociodemographic characteristics and placement history variables may be influential in the level of psychotropic medication use among this population. Further research to examine the appropriateness and level of benefit of medication treatment in this population is needed. PMID- 10521008 TI - Use of psychotropic medications in special education students with serious emotional disturbance. AB - The three-year usage of psychotropic medication was investigated for the first time in elementary school students classified by the special education category serious emotional disturbance. Almost 40% of the original 89 students were on a medication at baseline, primarily stimulants (26%), and multiple medications were not common (17%). Over the three time points of followup, 52% of the constant 54 students used a medicine at least once, principally stimulants followed by, in descending order, antipsychotics, antidepressants, and Clonidine. The order of distribution was constant at each point of follow-up. Only 24% received a medication at all three time points (rarely the same specific medicine), and only 9% received therapy at each time. Overall, 41% of the ongoing students received no medication or therapy over the course of the study. The medication needs, as well as the therapy needs, of this highly dysfunctional group of students appear unmet. PMID- 10521009 TI - Unravelling sleep problems in treated and untreated children with ADHD. AB - This study investigated parental perception of sleep problems in stimulant treated and untreated children with Attention Deficit Hyperactivity Disorder (ADHD). Parents of 135 psychiatric clinic referred children and 83 pediatric outpatients completed a sleep questionnaire and the Child Behavior Checklist. Moderate to severe "sleep problems" reportedly occurred at least once a week in 19.3% of children with ADHD, 13.3% of the psychiatric controls, and 6.2% of the pediatric controls. Children with ADHD treated with stimulants were reported to display a higher prevalence of nightly "severe" sleep problems than did untreated children with ADHD. Almost a third (29%) of stimulant treated ADHD children were reported to display increased sleep latency or insomnia every night versus 10% of untreated children with ADHD. Despite the high prevalence of sleep related problems in ADHD, the significance of the association between delayed sleep onset and ADHD with regard to etiology and management of ADHD is still poorly understood. PMID- 10521010 TI - Childhood inattention-overactivity, aggression, and stimulant medication history as predictors of young adult outcomes. AB - This study examined the contributions of childhood symptom dimensions and aspects of methylphenidate (MPH) treatment to the prediction of young adult outcomes in boys who were referred to a child psychiatry outpatient clinic. They were diagnosed with hyperkinetic reaction of childhood/minimal brain dysfunction, and given MPH for an average of 30 months. Including significant effects and statistical trends, childhood Inattention-Over-activity was uniquely associated with fewer than 10% of adult outcomes such as schizotypic features, impairment on the Global Assessment Scale (GAS), and unemployment. Childhood aggression was uniquely associated with 38% of adult outcomes such as lifetime diagnoses of major depression, drug abuse disorder, and antisocial personality disorder; MMPI PD, PA, and SC scores; and six additional measures of adult impairment and life circumstances-extending external validation of the two-factor model to young adulthood. For 20 young adult outcomes (63%), aspects of childhood treatment with MPH had no lasting effects. For one adult outcome (3%), a lasting negative effect of childhood drug treatment was found; better initial response to medication was associated with not graduating from high school. For 11 young adult outcomes (34%), however, aspects of childhood MPH treatment had positive effects that lasted long after treatment was discontinued. Higher dosage was associated with fewer diagnoses of alcoholism or suicide attempts. Better response to medication was associated with lower MMPI D scores and better social functioning. Longer medication duration was associated with fewer schizotypic features, lower MMPI MA scores, higher WAIS Performance and Full Scale IQs, and better WRAT Reading and Arithmetic performance. PMID- 10521011 TI - Do stimulants improve self-esteem in children with ADHD and peer problems? AB - The self-esteem of children with Attention Deficit Hyperactivity Disorder (ADHD) has been shown to be low. The effects of stimulant medication upon their self esteem have not been systematically studied. The present study employed a reliable self-report instrument to measure the self-esteem of children with ADHD medicated with stimulants vs. those who were unmedicated. Results showed that stimulants were associated with significantly higher self-esteem. Children with ADHD prescribed stimulants reported feeling more intelligent and more popular than unmedicated children with ADHD. Children with ADHD and Oppositional Defiant Disorder (ODD) prescribed stimulants reported feeling better behaved. Significant correlations indicated that higher doses were associated with higher self-esteem. The present results suggest a need for a well-controlled study to determine if stimulants were responsible for the observed differences in self-esteem. PMID- 10521012 TI - No effect of adjunctive fluoxetine on eating behavior or weight phobia during the inpatient treatment of anorexia nervosa: an historical case-control study. AB - A six-week open label clinical trial investigated the response to fluoxetine in adolescents hospitalized for treatment of anorexia nervosa. Patients were drawn from consecutive admissions to a specialty treatment service and received fluxoetine as an add-on to their multidisciplinary treatment regimen beginning three weeks to one month postintake. Global clinical severity ratings of eating behavior and weight phobia were obtained at baseline and then at biweekly intervals. These ratings were compared to those obtained from matched historical case-controls who received an identical course of inpatient therapy but without adjunctive pharmacotherapy. Analyses failed to show any beneficial or detrimental effect of fluoxetine on these clinical measures. We conclude that fluoxetine has no robust additive or synergistic therapeutic benefit when measured against the effects of intensive, multidisciplinary inpatient therapy. PMID- 10521014 TI - Medication treatment in adolescents vs. adults with psychotic mania. AB - An epidemiologic sample of first admission psychotic patients is used to examine naturalistic treatment of psychotic mania. Specifically, we examined if 15-20 year olds receive different medications, or respond differently from patients over age 30 who are also early in their course of illness. The major difference in the two groups, besides their age, is the presence of comorbid externalizing disorder and substance abuse in the younger group. This report further examines the impact of this comorbidity on the presence of further episodes and overall outcome. The findings indicate that acute treatment is very similar in the two groups, and that over a 4 year follow-up, about 40% of patients discontinue medication. In spite of that, 32% of the YOUTH subjects and 48% of the ADULT group did not have a second episode. Of those externalizing YOUTH with a single episode, 64% took no medication during follow-up. However, the major difference between admission, 24 month follow-up and 48 month follow-up was the discontinuation of substance/alcohol use and the improvement in functioning. Medication did not invariably make that difference. This community sample reveals that at 48 months, a significant minority of young and adult subjects hospitalized with psychotic mania will have a single episode, and that short term course in younger patients is considerably worse than longer term course. PMID- 10521013 TI - Psychopharmacology of pediatric posttraumatic stress disorder. AB - OBJECTIVE: To review the current knowledge of pharmacotherapy in the treatment of Post-traumatic Stress Disorder (PTSD) as it applies to children and adolescents and to provide a rational approach to medication use in Pediatric PTSD. METHOD: The literature on the psychopharmacology of Pediatric PTSD is reviewed. Additionally, literature is reviewed on the neurobiological systems presumptively involved in trauma as well as studies in the pharmacology of adult PTSD, as they pertain to the treatment of Pediatric PTSD. RESULTS: There are too few studies in the current Pediatric PTSD literature to confirm treatment recommendations. Downward extrapolation from the adult literature combined with an understanding of the neurobiology of PTSD and its comorbid conditions may serve as the basis for a rational pharmacotherapy of PTSD in childhood. The effectiveness of targeting pharmacological agents at PTSD symptom clusters and associated comorbid conditions remains to be verified in controlled clinical trials. CONCLUSIONS: The state of psychopharmacology for Pediatric PTSD is in its earliest stages. While there are insufficient numbers of controlled pharmacological trials to make firm recommendations, the field requires a starting point for a rational psychopharmacological approach. Pharmacotherapy may provide symptom relief of both the debilitating primary symptoms and the comorbid conditions in children suffering from PTSD. PMID- 10521015 TI - Technical note: development of a transcervical oocyte recovery procedure for sheep. AB - An oocyte recovery procedure was developed and evaluated to determine whether a transcervical embryo recovery procedure is feasible with our method, which includes estradiol-17beta (E2) and oxytocin (OT) treatments, for dilating the cervix in ewes. On d 6 of an estrous cycle, oocytes were recovered either transcervically or with a laparotomy procedure. In the laparotomy group, ovulation rate was determined during the procedure and was used to calculate the percentage ofoocytes recovered. The laparotomy procedure was a standard uterine flush, and 12 mL of PBS was used to flush each uterine horn. In the transcervical group, the ovaries in each ewe were evaluated ultrasonically to determine ovulation rate. For transcervical recovery, 100 microg of E2 were injected i.v. on d 5 to increase cervical OT receptors, and 100 USP units of OT were injected i.v. 10 to 12 h later to dilate the cervix. Approximately 25 min after OT, ewes were placed in dorsal recumbency in a Commodore cradle, and a modified Foley catheter was passed through the cervix and into the uterus for injection (80 to 210 mL) and aspiration of PBS. The PBS was aspirated with a vacuum pump. The percentage of PBS recovered was greater (P<.01) at laparotomy than with the transcervical procedure (85.8 vs. 36.2%). Despite that difference, oocyte recovery did not differ significantly between the two groups (67% for laparotomy vs. 50% for transcervical; [oocytes recovered/number of corpora lutea] x 100), and there was no evidence that the transcervical procedure damaged the oocytes; the zona pellucida remained intact around all of the oocytes. In conclusion, a procedure that includes E2-OT-induced cervical dilation, passage of a modified Foley catheter into the uterus, and incremental infusion and aspiration of media through the catheter can be used to recover oocytes transcervically from ewes. This procedure may make transcervical embryo recovery feasible for sheep. PMID- 10521017 TI - Prediction of clean mohair, fiber diameter, vegetable matter, and medullated fiber with near-infrared spectroscopy. AB - Four experiments were conducted in three separate years to test the utility of near-infrared spectroscopy (NIRS) to predict the clean mohair content of Angora goat fleece. Mohair fleece samples were obtained each year from yearling billies at the conclusion of the Angora Goat Performance Test conducted at the Texas A&M University Research Station, Sonora. In Exp. 1 (n = 293) and Exp. 2 (n = 256), fleeces were scanned with a Pacific Scientific (Silver Spring, MD) near-infrared spectrometer fitted with a fiber-optic probe, and calibrations were developed for clean mohair content. In Exp. 3, 59 mohair fleeces collected at the Texas A&M Research Station in San Angelo were sampled four times each. Each sample was scanned with the same spectrometer in reflectance mode fitted with a transport mechanism. This mechanism allowed the instrument to scan a 15-cm2 segment of the fleece sample. Conventional procedures to determine reference values for mohair yield, vegetable matter content, fiber diameter, and percentage of medullated and kemp fibers were conducted. Prediction equations were developed that related NIR spectra to reference values for yield and diameter parameters and were used to predict mohair characteristics for each fleece sample. The predicted and reference values were subjected to a simple analysis of variance to determine variation within and across samples. In Exp. 1, mohair base was related to NIR spectra with R2 = .46 and standard error of calibration (SEC) = 2.84%. In Exp. 2, similar repeatability errors for mohair base could be obtained for both reference and NIRS-derived values. Fiber diameter and medullated fibers were poorly related to NIR spectra. When samples were scanned using the transport mechanism (Exp. 3), R2 and SEC were .82 and 1.19% for mohair base and .93 and .98 microm for fiber diameter, respectively. The CV for mohair base and diameter were 1.0 and 1.4%, whereas those for predicted mohair base and diameter were 1.4 and 3.4%, respectively. The increased variation within samples for predicted values represents sampling error and lack of fit between NIRS and the laboratory determined values. When the samples from Exp. 1 and 2 were rescanned with the NIRS transport (Exp. 4), R2 and SEC were .79 and 2.03% for mohair base and .52 and 3.49 microm for fiber diameter. The fiber optic probe would facilitate real time analysis on the shearing floor, but our data indicate that the spectral limitations so far are too severe. A large sample device such as the transport gave excellent results for predicting mohair base and fiber diameter. PMID- 10521016 TI - Estradiol-17 beta-oxytocin-induced cervical dilation in sheep: application to transcervical embryo transfer. AB - Experiments were conducted to determine whether exogenous estradiol-17beta (E2) and oxytocin (OT) can be used to improve transcervical (TC) embryo transfer (ET) procedures for sheep. Our concerns that the E2-OT treatment may alter luteal function prompted Exp. 1, in which 32 ewes were assigned to treatments in a 2x2 factorial array. On d 7 after onset of estrus, ewes received i.v. either 100 microg of E2 or diluent; 12 h later, ewes received i.v. either 400 USP units of OT or saline. To monitor luteal function, progesterone was measured in jugular blood collected from d 7 to 18. The treatments did not affect progesterone concentrations. Two trials were conducted in Exp. 2. In Trial 1, ewes were assigned to one of three treatments: TC transfer with E2-OT treatment to dilate the cervix, laparoscopic ET with E2-OT treatment, or laparoscopic ET with an equivalent diluent that did not dilate the cervix. In Trial 2, ewes were assigned to treatments in a 2x2 factorial array: TC or laparoscopic ET on d 6; E2-OT treatment for cervical dilation or diluents on d 6. Transferred embryos were recovered on d 12 in Trial 1 and d 14 in Trial 2, evaluated morphologically for development, and scored. Treatments did not affect the percentage of transferred embryos recovered. However, mode of transfer decreased (P<.01) the mean embryo development score. The E2-OT treatment increased (P<.01) the development score of embryos transferred transcervically, indicating that cervical dilation may improve the chances of embryos surviving after TC transfer. In conclusion, E2-OT treatment did not affect luteal function, and the E2-OT treatment can be used to enhance the success of TC embryo transfer in sheep. PMID- 10521018 TI - Influence of free fatty acid content on the feeding value of yellow grease in finishing diets for feedlot cattle. AB - Holstein steers (n = 96; 375 kg) were used in a 144-d growth-performance trial to evaluate influence of level (42, 28.5, and 15%) of FFA content on feeding value of yellow grease. Two sources of yellow grease were compared: conventional yellow grease (CYG), containing 15% FFA, and griddle grease (GG), containing 42% FFA. Dietary treatments consisted of an 88% concentrate finishing diet supplemented with either 1) 0% fat, 2) 5% GG, 3) 2.5% GG and 2.5% CYG, or 4) 5% CYG. Fat supplementation increased ADG (11%; P<.05), feed efficiency (9%; P<.05), diet NE (6.4%; P<.05), carcass weight (4%; P<.10), dressing percentage (1%; P<.10), and kidney, pelvic, and heart fat (20%, P<.05). Increasing the FFA in supplemental fat increased (linear effect, P<.10) DM intake, ADG, and feed efficiency and decreased (linear effect, P<.10) retail yield. These improvements in performance were primarily due to increased DM intake. The NEm and NEg values of supplemental fats were not affected by FFA content, averaging 4.98 and 3.85 Mcal/kg, respectively. Treatment effects on characteristics of ruminal and total tract digestion were evaluated using four Holstein steers (180 kg) with cannulas in the rumen and proximal duodenum. Supplemental fat did not influence (P>.10) ruminal or total tract digestion of OM, ADF, starch or N. Postruminal fatty acid digestion was less (P<.10) for fat-supplemented diets than for unsupplemented diets (73.0 vs. 78.6%). The decrease in postruminal fatty acid digestibility with fat supplementation was mainly due to a decreased (16.7%; P<.05) digestibility of C18:0. Postruminal digestibility of the supplemental fat was 68%. There were no treatment effects (P>.10) on ruminal pH. Ruminal biohydrogenation of fatty acids was directly proportional to estimates of methane production. We conclude that the feeding value of conventional yellow grease and griddle grease is similar and that differences in the FFA content of yellow grease will not negatively affect diet acceptability and growth performance of feedlot cattle. PMID- 10521019 TI - Influence of body condition score on live and carcass value of cull beef cows. AB - Mature beef cows (n = 88) were slaughtered to determine the influence of body condition score (BCS) on carcass and live animal value. Cows were weighed and assigned a BCS (9-point scale), 24 h before slaughter. Hide and by-products weights were recorded during harvest. After a 48-h chill period, the right side of each carcass was fabricated into boneless subprimal cuts, minor cuts, lean trim, fat, and bone. Weights were recorded at all stages of fabrication. Carcass values (U.S.$/100 kg of hot carcass weight) were calculated for U.S. Utility and U.S. Cutter grades, as well as for the Utility/Cutter mix for each BCS. Gross value included the carcass value and the value of the hide and byproducts, whereas net value was calculated after harvest and fabrication costs and by product value were considered. Live value (U.S.$/100 kg of live weight) was computed by dividing the net value by the animal's live weight 24 h before harvest. The value of the hide and by-products for BCS-2 cows was greater (P<.05) than for cows assigned a BCS of 3 through 8. Even though U.S. Utility carcasses from BCS-8 cows produced the least (P<.05) valuable subprimal cuts from the chuck, loin, and round, the gross and net values of BCS-8 cows were greater (P<.05) than those of BCS-3, 4, 5, and 6. Within the grade of U.S. Cutter, carcasses from BCS-6 cows had the highest (P<.05), and BCS-2 cows had the lowest (P<.05), gross and net values. Across the U.S. Utility/Cutter mix, cows designated with a BCS of 7 and 8 had greater (P<.05) gross and net values than cows assigned a BCS of 6, or lower. Live value increased linearly (P = .0002) from a low of $76.10/100 kg for BCS-2 cows to a high of $90.84/100 kg for BCS-7 cows. Carcasses from BCS-6 cows were relatively lean (8.4 mm of fat opposite of the longissimus muscle), and approximately 73% of the carcasses achieved a quality grade of U.S. Utility. Moreover, carcasses from BCS-6 cows had the highest total carcass values and live values comparable (P>.05) to BCS-7 cows. Information from this study can be used by the non-fed beef industry to establish a value-based marketing system. Data from this study would indicate that marketing cull beef cows at a BCS of 6 could optimize economic returns to both cow-calf producers and non-fed beef packers. PMID- 10521020 TI - Additive genetic relationships between heifer pregnancy and scrotal circumference in Hereford cattle. AB - The objective of this study was to determine an appropriate method for using yearling scrotal circumference observations and heifer pregnancy observations to produce EPD for heifer pregnancy. We determined the additive genetic effects of and relationship between scrotal circumference and heifer pregnancy for a herd of Hereford cattle in Solano, New Mexico. The binary trait of heifer pregnancy was defined as the probability of a heifer conceiving and remaining pregnant to 120 d, given that she was exposed at breeding. Estimates of heritability for heifer pregnancy and scrotal circumference were .138+/-.08 and .714+/-.132, respectively. Estimates of fixed effects for age of dam and age were significant for heifer pregnancy and bull scrotal circumference. The estimate of the additive genetic correlation between yearling heifer pregnancy and yearling bull scrotal circumference was .002+/-.45. Additional analyses included models with additive genetic groups for scrotal circumference EPD for heifer pregnancy or heifer pregnancy EPD for scrotal circumference to account for a potential nonlinear relationship between scrotal circumference and heifer pregnancy. Results support the development of a heifer pregnancy EPD because of a higher estimated heritability than previously reported. The development of a heifer pregnancy EPD would be an additional method for improving genetic merit for heifer fertility. PMID- 10521021 TI - Technical note: use of PCR-single-strand conformation polymorphism analysis for detection of bovine beta-casein variants A1, A2, A3, and B. AB - We have optimized the polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP) technique to screen the most frequent variants (A1, A2, A3, and B) of the bovine beta-casein gene. Five partly overlapping PCR products (233, 234, 265, 466, and 498 bp) of Exon VII of the beta-casein gene that encompass the target point mutations were heat-denatured, separated on nondenaturing polyacrylamide gels, and silver-stained. Simultaneous detection of all variants in reference samples of known genotypes (A1A2, A2A2, A1A3, A1B, and A2B) was best achieved on 17% polyacrylamide (100:1 acrylamide:bis-acrylamide ratio) gels with the PCR product of 234 bp. These results were confirmed by sequencing the allele-specific SSCP bands directly excised from polyacrylamide gels. A population of 65 anonymous samples belonging to various breeds was then analyzed twice, without discrepancies in a blind trial. Routine beta-casein genotyping using PCR-SSCP is proposed as a cost-effective, fast, and sensitive technique. PMID- 10521022 TI - Economic values for meat quality traits in pigs. AB - The aim for this study was to derive economic values for meat quality traits. Meat quality experts from 10 slaughter and retail companies in Switzerland were asked to give an interview in which they indicated their willingness to pay different prices for carcasses from different quality classes. Economic values for the seven meat quality traits color (L*), drip loss, intramuscular fat content, iodine value, pH1 (45 min after slaughtering), pH2 (24 to 30 h after slaughtering), and proportion of premium cuts were derived from the answers. For each trait, a weighted mean of these economic values was calculated using the number of pigs slaughtered by a company per year as a weighting factor. Population mean and standard deviation of the investigated traits were taken from station test results of Swiss Large White pigs (intramuscular fat content, iodine value, pH1, pH2, and proportion of premium cuts) and from the literature (color and drip loss). Weighted means (range among companies) of economic values per slaughter pig on the basis of one phenotypic standard deviation of the trait in Swiss Francs (SFr.) were -22.92 (0 to -62.46) for color, -10.27 (0 to -43.64) for drip loss, 10.84 (0 to 29.43) for intramuscular fat content, -29.90 (-15.54 to 43.58) for iodine value, 11.50 (.67 to 19.79) for pH1, 0 (0) for pH2, and 10.92 ( 2.30 to 33.67) for proportion of premium cuts. PMID- 10521023 TI - Variance components and breeding values for growth traits from different statistical models. AB - Estimates of genetic parameters resulting from various analytical models for birth weight (BWT, n = 4,155), 205-d weight (WWT, n = 3,884), and 365-d weight (YWT, n = 3,476) were compared. Data consisted of records for Line 1 Hereford cattle selected for postweaning growth from 1934 to 1989 at ARS-USDA, Miles City, MT. Twelve models were compared. Model 1 included fixed effects of year, sex, age of dam; covariates for birth day and inbreeding coefficients of animal and of dam; and random animal genetic and residual effects. Model 2 was the same as Model 1 but ignored inbreeding coefficients. Model 3 was the same as Model 1 and included random maternal genetic effects with covariance between direct and maternal genetic effects, and maternal permanent environmental effects. Model 4 was the same as Model 3 but ignored inbreeding. Model 5 was the same as Model 1 but with a random sire effect instead of animal genetic effect. Model 6 was the same as Model 5 but ignored inbreeding. Model 7 was a sire model that considered relationships among males. Model 8 was a sire model, assuming sires to be unrelated, but with dam effects as uncorrelated random effects to account for maternal effects. Model 9 was a sire and dam model but with relationships to account for direct and maternal genetic effects; dams also were included as uncorrelated random effects to account for maternal permanent environmental effects. Model 10 was a sire model with maternal grandsire and dam effects all as uncorrelated random effects. Model 11 was a sire and maternal grandsire model, with dams as uncorrelated random effects but with sires and maternal grandsires assumed to be related using male relationships. Model 12 was the same as Model 11 but with all pedigree relationships from the full animal model for sires and maternal grandsires. Rankings on predictions of breeding values were the same regardless of whether inbreeding coefficients for animal and dam were included in the models. Heritability estimates were similar regardless of whether inbreeding effects were in the model. Models 3 and 9 best fit the data for estimation of variances and covariances for direct, maternal genetic, and permanent environmental effects. Other models resulted in changes in ranking for predicted breeding values and for estimates of direct and maternal heritability. Heritability estimates of direct effects were smallest with sire and sire maternal grandsire models. PMID- 10521024 TI - Germplasm evaluation in beef cattle--Cycle IV: postweaning growth and puberty of heifers. AB - Postweaning growth, puberty, and pregnancy traits were evaluated for 783 F1 heifers sired by Angus, Hereford, Charolais, Shorthorn, Galloway, Longhorn, Nellore, Piedmontese, and Salers bulls and out of Angus and Hereford dams in Cycle IV of the Germplasm Evaluation (GPE) Program at the U.S. Meat Animal Research Center. The Hereford and Angus sires included a sample of bulls born from 1982 to 1985 (1980s HA) as well as reference sires born from 1963 to 1970 (REF HA) used in previous cycles of the GPE program. Breed group of sire had a significant (P<.01) effect on age and weight at puberty, on 200-, 400-, and 550-d weights, on ADG from 200 to 400 and from 400 to 550 d, and 550-d hip height, but it did not influence (P<.05) pregnancy rate. Mean age and weight at puberty were predicted from the cumulative distribution because of censoring of data in each tail of the distribution. Sire breed group rankings (and predicted means in days) for age at puberty were as follows: Piedmontese (332), Shorthorn (338), Charolais (348), REF HA (348), Galloway (351), 1980s HA (352), Salers (355), Longhorn (357), and Nellore (405). Sire breed group rankings (and predicted means in kilograms) for weight at puberty were Longhorn (283), Piedmontese (298), Galloway (305), REF HA (309), Shorthorn (329), 1980s HA (330), Salers (338), Nellore (341), and Charolais (345). Sire breed group rankings (and least squares means in kilograms) for 200-d weight were Charolais (229), Salers (225), Nellore (221), Shorthorn (220), Piedmontese (215), 1980s HA (215), Galloway (209), REF HA (206), and Longhorn (197), with differences >8.3 kg significant. Rankings for 400-d weight (kilograms) were Charolais (390), Shorthorn (384), Salers (380), 1980s HA (374), Nellore (364), REF HA (356), Piedmontese (353), Galloway (348), and Longhorn (321), with differences >11.5 kg significant. Rankings for 550-d weight (kilograms) were Charolais (445), Salers (430), Shorthorn (429), 80's HA (422), Nellore (420), Piedmontese (401), REF HA (398), Galloway (389), and Longhorn (371), with differences >11.7 kg significant. Rankings for 550-d hip height (centimeters) were Nellore (132.2), Charolais (131.9), Salers (129.9), Shorthorn (129.5), Piedmontese (126.7), 1980s HA (126.1), Longhorn (125.3), Galloway (121.7), and REF HA (121.5), with differences >1.35 cm significant. Breed of sire had significant effects on growth and puberty traits of heifers. PMID- 10521025 TI - Influence of body condition score on carcass characteristics and subprimal yield from cull beef cows. AB - Mature beef cows (n = 83) were slaughtered to measure the influence of body condition score (BCS) on carcass characteristics and subprimal yields. All cows were weighed and assigned BCS, based on a 9-point scale, 24 h before slaughter. Cows were slaughtered, and, after a 48-h chilling period, quality and yield grade data were collected on the left side of each carcass. The right side was quartered, fabricated into primal cuts, and weighed. Each primal cut was further processed into boneless subprimal cuts, minor cuts, lean trim, fat, and bone. Cuts were progressively trimmed to 6.4 and 0 mm of external and visible seam fat. Weights were recorded at all stages of fabrication, and subprimal yields were calculated as a percentage of the chilled carcass weight. Live weight, carcass weight, dressing percentage, fat thickness, longissimus muscle area, muscle:bone ratio, and numerical yield grade increased linearly (P = .0001) and predicted cutability and actual muscle-to-fat ratio decreased linearly (P = .0001) as BCS increased from 2 to 8. Carcasses from BCS-8 cows had the most (P<.05) marbling. The percentage of carcasses grading U.S. Utility, or higher, was 16.7, 20.0, 63.6, 43.3, 73.3, 100.0, and 100.0% for cows assigned a BCS of 2, 3, 4, 5, 6, 7, and 8, respectively. At 6.4 mm of fat trim, carcasses from BCS-5 cows had higher (P<.05) shoulder clod yields than carcasses from cows having a BCS of 6, 7, and 8. Carcasses of BCS-2 cows had lower (P<.05) strip loin yields than carcasses from BCS-3, 4, 5, 6, and 7 cows. Top sirloin butt yields were higher (P<.05) for carcasses of BCS-2, 3, 4, and 5 cows than those of BCS-6, 7, or 8 cows. Carcasses from BCS-7 and 8 cows had lower (P<.05) tenderloin and inside round yields than carcasses of BCS-5, or less, cows. At both fat-trim levels, carcasses from BCS-5 cows had higher (P<.05) eye of round yields than cows assigned BCS of 2, 7, or 8. When subprimal cuts were trimmed to 6.4 mm of visible fat, carcasses from BCS-5 cows had higher (P<.05) total lean product yields than cows assigned a BCS of 2, 4, 7, and 8. Regardless of fat trim, total fat yields increased (P = .0001) and total bone yields decreased (P = .0001) linearly as BCS increased from 2 to 8. Although carcasses from BCS-5 and 6 cows had the highest yields of lean product, cattle producers and packers may benefit most by marketing and(or) purchasing BCS 6 cows because a higher percentage of their carcasses had quality characteristics deemed desirable for fabrication into boneless subprimal cuts. PMID- 10521026 TI - Influence of body condition score on by-product yield and value from cull beef cows. AB - Mature beef cows (n = 122) representing British and Continental phenotypes were slaughtered to measure the influence of body condition score (BCS) on by-product yield and value. All cows were weighed and assigned BCS, based on a 9-point scale, 24 h before slaughter. By-product weights were obtained during the slaughter process and included blood, feet (with hooves attached), oxlips, tongue, gullet, trachea, cheek meat, head meat, skull, tripe, honeycomb tripe, large and small intestines, spleen, mesenteric fat, weasand meat, kidneys, heart, lungs, and oxtail. By-product yields were calculated as a percentage of the animal's live weight taken 24 h before slaughter. By-product values were computed by multiplying the weight of each piece removed during the slaughter process by the 1997 average price. Live weight increased linearly (P<.001) as BCS increased from 2 to 8, whereas Continental cows were approximately 86 kg heavier (P<.05) at slaughter than British cows. Cows assigned a BCS of 2 or 3 had greater (P<.05) skull, feet, tongue, tripe, honeycomb tripe, trachea, and lung yields than cows assigned a BCS of 4 or higher. On the other hand, BCS-7 and 8 cows had greater (P<.05) weights and yields of large intestines and mesenteric fat than cows given a BCS of 6 or lower. The feet, trachea, lungs, and bone meal from BCS-2 cows had the greatest (P<.05) value, whereas the value of the large intestine, oxtail, and mesenteric fat was highest (P<.05) for BCS-7 and 8 cows. Weight, yield, and value of the skull, head meat, and feet were greater (P<.05) for Continental cows than British cows. Total by-product value was quadratically (P<.001) related to BCS. Cows assigned a BCS of 5 had lower (P<.05) total by-product values than either BCS-2 or BCS-7 and 8 cows. Drop credit for BCS-2 cows was greater (P<.05) than BCS-3, 4, 5, and 6 cows, with cows assigned a BCS of 7 and 8 having intermediate drop credit values. Continental cows tended to have greater (P<.10) total by product and drop credit values than British cows. Information from this study indicated that the BCS of cows at the time of slaughter had a profound influence on by-product yields and, more importantly, values of by-products that are credited back against the cost of production to the beef cattle producer. PMID- 10521027 TI - Evaluation of porcine fat with fiber-optic spectroscopy. AB - Spectroscopy using four types of fiberoptic probes and a sensor at wavelengths of 400 to 1,100 nm was evaluated to assess porcine fat quality. The shapes of the spectrum for the leaf fat with white color and various firmnesses differed with the type of probe: surface, contact, insertion, or transmittance. The internal reflectance ratio using the insertion probe at wavelengths from approximately 600 to 1,000 nm was positively correlated with the hardness, melting point, and saturated fatty acid content of the fat, but it was negatively correlated with the refractive index and polyunsaturated fatty acid content. The correlations between the internal reflectance ratio using the insertion probe and the monounsaturated fatty acid content were strong only near 1,100 nm. Surface reflectance at more than 650 nm was negatively correlated with refractive index. Transmittance at almost all wavelengths showed positive correlations with monounsaturated fatty acid content, but it was negatively correlated with hardness, melting point, and saturated fatty acid content. The interactance using the contact probe did not have a significant correlation with any physiochemical characteristics. The strongest relationships for hardness, refractive index, saturated fatty acid content, monounsaturated fatty acid content, polyunsaturated fatty acid content, and melting point were obtained at 650 nm (r = .88), 660 nm (r = -.91), 645 nm (r = .73), 1,095 nm (r = .68), and 930 nm (r = -.76), respectively, using the insertion probe and 1,050 nm (r = -.79) using the transmittance probe (P<.05). These results indicated that fiber-optic methods were rapid and useful techniques for the evaluation of porcine fat quality. PMID- 10521028 TI - Effect of calpastatin on degradation of myofibrillar proteins by mu-calpain under postmortem conditions. AB - To improve our understanding of the regulation of mu-calpain activity in situ during postmortem storage of muscle, the effect of different calpastatin levels on proteolysis of myofibrillar proteins by mu-calpain in a system closely mimicking postmortem conditions was studied. Increasing the amount of calpastatin in the incubations limited both the rate and extent of proteolysis of myofibrillar proteins and autolysis of mu-calpain. Excess calpastatin (i.e., a mu calpain:calpastatin ratio of 1:4) did not inhibit proteolysis completely. Western blot analysis revealed that proteolysis of myofibrillar proteins virtually ceased after 7 d of incubation, despite the presence of partly autolyzed, therefore seemingly active, mu-calpain. A series of incubations of autolyzed mu-calpain revealed that the autolyzed form of this enzyme is unstable at an ionic strength observed in postmortem muscle. The possible significance of these results in terms of the regulation of mu-calpain activity in postmortem muscle is discussed. PMID- 10521029 TI - Evaluation of slice shear force as an objective method of assessing beef longissimus tenderness. AB - Experiments were conducted to develop an optimal protocol for measurement of slice shear force (SSF) and to evaluate SSF as an objective method of assessing beef longissimus tenderness. Whereas six cylindrical, 1.27-cm-diameter cores are typically removed from each steak for Warner-Bratzler shear force (WBSF) determination, a single 1-cm-thick, 5-cm-long slice is removed from the lateral end of each longissimus steak for SSF. For either technique, samples are removed parallel to the muscle fiber orientation and sheared across the fibers. Whereas WBSF uses a V-shaped blade, SSF uses a flat blade with the same thickness (1.016 mm) and degree of bevel (half-round) on the shearing edge. In Exp. 1, longissimus steaks were acquired from 60 beef carcasses to determine the effects of belt grill cooking rate (very rapid vs. rapid) and conditions of SSF measurement (hot vs cold) on the relationship of SSF with trained sensory panel (TSP) tenderness rating. Slice shear force was more strongly correlated with TSP tenderness rating when SSF measurement was conducted immediately after cooking (r = -.74 to -.76) than when steaks were chilled (24 h, 4 degrees C) before SSF measurement (r = .57 to -.72). When SSF measurement was conducted immediately after cooking, the relationship of SSF with TSP tenderness rating did not differ among the belt grill cooking protocols used to cook the SSF steak. In Exp. 2, longissimus steaks were acquired from 479 beef carcasses to compare the ability of SSF and WBSF of 1.27-cm-diameter cores to predict TSP tenderness ratings. Slice shear force was more strongly correlated with sensory panel tenderness rating than was WBSF (r = .82 vs -.77). In Exp. 3, longissimus steaks were acquired from 110 beef carcasses to evaluate the repeatability (.91) of SSF over a broad range of tenderness. Slice shear force is a more rapid, more accurate, and technically less difficult technique than WBSF. Use of the SSF technique could facilitate the collection of more accurate data and should allow the detection of treatment differences with reduced numbers of observations and reduced time requirements, thereby reducing research costs. PMID- 10521030 TI - Effects of omitting vitamin and trace mineral premixes and(or) reducing inorganic phosphorus additions on growth performance, carcass characteristics, and muscle quality in finishing pigs. AB - Three experiments were conducted to determine the effects of omitting vitamin and trace mineral premixes and(or) reducing inorganic phosphorus additions to finishing diets on growth performance, carcass characteristics, and muscle quality in pigs. In Exp. 1, a corn-soybean meal-based diet (.70% lysine, .65% Ca, and .55% P) was used as the control. Pigs (n = 128; average initial BW of 85.7 kg) were fed the control diet or the control diet without 1) the vitamin premix, 2) the trace mineral premix, or 3) both premixes. Omitting the premixes had no effect on ADG (P>.39); gain/feed (P>.17); carcass backfat thickness (P>.42); and marbling, color, and firmness of the longissimus muscle (P>.11). In Exp. 2, pigs (n = 128; average initial BW of 86.2 kg) were fed the control diet (.65% Ca and .53% P) used in Exp. 1 and the control diet without 1/3 (.56% Ca and .46% P), 2/3 (.51% Ca and .40% P), or all (.47% Ca and .31% P) of the added monocalcium phosphate (MCP). Omitting up to 2/3 of the MCP increased ADG (quadratic effect, P<.02) and had no effect on meat quality (P>.12), but backfat thickness increased slightly (quadratic effect, P<.02). In Exp. 3, pigs (n = 160; average initial BW of 86.6 kg) were fed the control diet used in Exp. 1 or the control without 1) the vitamin and trace mineral premixes, 2) 2/3 of the MCP, or 3) the premixes and 2/3 of the MCP. Treatment had no effects on ADG (P>.23), gain/feed (P>.94), stomach lesions (P>.37), or serum gamma globulins (P>.08). In conclusion, vitamin and trace mineral premixes and up to 2/3 of the supplemental MCP can be omitted during late finishing (i.e., approximately the final 30 d) to reduce nutrient excesses that increase cost of feeding and nutrients excreted in waste material. PMID- 10521031 TI - Effects of diet and housing density on growth and stomach morphology in pigs. AB - Two experiments were conducted to evaluate the impact of housing density on the stomach morphology of growing pigs and determine whether there was an interaction between housing density and diet. All diets were corn-soybean meal based. In Exp. 1, 42 barrows (41.0+/-.95 kg BW) were allotted either individually or three pigs per pen to evaluate the effects of crowding on stomach lesions. Pen space per pig was 1.54 and .51 m2, respectively. All pigs were fed a finely ground and pelleted diet (610 microm) for 6 wk. The ADG decreased (P<.05) for the pigs housed three per pen during wk 4 to 6 only. There was no effect of housing density on feed intake or gain/feed ratio. Neither visual nor histological ulcer score differed between the two treatment groups. No stomachs were graded as normal. In Exp. 2, 80 barrows (39.8+/-.9 kg BW) were allotted either two or four pigs per pen. Pen space per pig was .77 and .39 m2, respectively. Half of the pigs in each housing situation were fed a coarse meal diet (1,050 microm), and half of the pigs were fed a finely ground and pelleted diet (577 microm) throughout the 49-d experimental period. Throughout the trial, pigs housed two per pen gained at a greater rate (P<.05) than pigs housed four per pen. From d 14 to the end of the trial, pigs consuming the finely ground and pelleted diet gained at a greater rate (P<.05) than pigs fed the coarse meal diet. The differences in ADG were reflected in final body weight. Stomach weight as a percentage of body weight was higher for animals on the coarse meal diet. Visual and histological ulcer scores were similar, and both were higher (P<.001) on the finely ground and pelleted diet, indicating greater damage. There was no effect of space restriction on stomach morphology. These data show the major effect of diet type on stomach lesions with no interaction with space restriction. PMID- 10521032 TI - Effects of diets differing in propensity to promote gastric lesions on defense systems in gastric mucosae. AB - The objectives were to characterize biochemical changes, focusing on the antioxidant defense system, in stratified squamous and oxyntic mucosae in pigs fed diets with differing propensity to promote gastric lesions. Barrows (n = 24; 48.7+/-1.0 kg BW) housed in individual pens were used in the experiment. Barrows were fed a corn-soybean meal diet. Half of the animals were fed the diet as a coarsely ground meal (CGM; average particle size = 886 microm), and half were fed the diet as a finely ground pelleted (FGP; average particle size = 528 microm) feed. Initiation and termination of the experiment were staggered over a 3-wk period. Diets were fed for 6 wk. Visual evaluation of the stratified squamous mucosa of the proximal stomach showed increased (P<.001) damage in animals fed the FGP diet. These results were supported by histological evaluation. Thiobarbituric acid-reacting substances (TBARS), indicative of peroxide generation, relative to amount of protein were higher (P<.001) in stratified squamous than in oxyntic mucosa, and, per unit of tissue, TBARS were highest in stratified squamous mucosa of animals fed the FGP diet. Glutathione peroxidase activity followed a pattern similar to that of peroxides. Prostaglandin E2 was higher (P<.004) in stratified squamous than in oxyntic mucosa. In contrast, the activity of catalase was higher (P<.001) in oxyntic mucosa and was not affected by diet. The data show differences in the production of peroxides, the antioxidant defense system, and PGE2 between stratified squamous and oxyntic mucosae. Generation of prooxidants and the antioxidant defense system may play a role in the predilection of ulcers for the stratified squamous mucosal region of the pig stomach. PMID- 10521033 TI - Changes in gastric contents in pigs fed a finely ground and pelleted or coarsely ground meal diet. AB - The objective was to characterize the change in stomach contents in relation to time after feeding between pigs consuming a restricted amount of a finely ground and pelleted (FGP) or coarsely ground meal (CGM) diet. Particular interest was placed on the concentration of organic acids and ammonia, the products of microbial fermentation. Thirty barrows were ranked by weight and assigned to a postfeeding time of 2, 4, 6, 8, or 12 h and either the FGP or CGM diet. Initiation and termination of the experiment were staggered over a 2-wk period. The treatment period was 42 d. Percentage of dry matter was higher (P<.01) in the stomach contents of pigs on the CGM diet. Concentrations of pepsin and protein were higher (P<.05) and ammonia tended to be higher (P = .10) in the proximal stomach of pigs fed the FGP diet. In contrast, concentrations of acetate and L lactate were higher (P<.05) in the proximal stomach of pigs fed the CGM diet. All pigs on the CGM diet had stomachs that graded as normal on visual inspection. There was variable damage to the stomachs of pigs on the FGP diet. Measurement of chromium concentration in the stomach after an oral dose of Cr-EDTA clearly demonstrated the mixing that occurs between the proximal and distal stomach by 2 h after feeding in pigs consuming the FGP diet, whereas a gradient was maintained in pigs consuming the CGM diet. Thus, components normally secreted in the distal stomach return to the proximal stomach. These data show that components secreted in the distal region, such as acid and pepsin, may play a role in initiating damage to the stratified squamous mucosa. High concentrations of organic acids in the stomach of pigs on the CGM diet were not associated with damage to the stratified squamous mucosa in the esophageal region. PMID- 10521034 TI - Bioavailability of zinc in several sources of zinc oxide, zinc sulfate, and zinc metal. AB - Three zinc depletion-repletion assays were carried out with chicks to determine Zn bioavailability in five sources of ZnO, three sources of ZnSO4.H2O, and two sources of Zn metal. A standard 23% CP corn-soybean meal diet was fed during the first 3 d posthatching, after which it was replaced with a Zn-deficient soy concentrate diet (13.5 mg Zn/kg) until d 7. On d 8 after an overnight period of feed withdrawal, chicks were fed for 12 d the Zn-deficient basal diet containing 0, 4.76, and 9.90 (Assay 1); 0, 5.06, or 10.12 (Assay 2); or 0, 4.73, or 9.13 (Assay 3) mg/kg supplemental Zn from analytical grade (AG) ZnSO4.7H2O (22.7% Zn) to generate a standard response curve. The AG and feed-grade (FG) Zn sources being evaluated were then provided at a level that would fall within the standard curve. Weight gain (Assays 1, 2, and 3) and total tibia Zn (Assay 1) responded linearly (P<.01) to Zn supplementation from ZnSO4.7H2O. Weight gain regressed on supplemental Zn intake gave standard-curve equations with fits (r2) ranging from .94 to .97. In Assay 1, regression of total tibia Zn (Y, in micrograms) on supplemental Zn intake (X, in milligrams/12 d) gave the equation Y = 13.2+6.74X (r2 = .90). Standard-curve methodology was used to estimate relative Zn bioavailability (RBV), with RBV of Zn in the ZnSO4.7H2O standard set at 100%. Four sources of FG ZnO were evaluated: Source 1 (78.1% Zn, hydrosulfide process, U.S.), Source 2 (74.1% Zn, Waelz process, Mexico), Source 3 (69.4% Zn, China), and Source 4 (78.0% Zn, French process, Mexico). Analytical-grade ZnO (80.3% Zn) was also evaluated. Feed-grade ZnO Sources 1 and 4 as well as AG ZnO produced average RBV values that were not different (P>.10) from the standard, but average RBV values for FG Source 2 and FG Source 3 were only 34 (P<.05) and 46% (P<.05), respectively. All sources of ZnSO4.H2O, which included two FG sources (source 1, 36.5% Zn; source 2, 35.3% Zn) and one food-grade source (36.5% Zn), were not different (P>.10) in RBV from the ZnSO4.7H2O standard. Two Zn metal products, Zn metal dust (100% Zn) and Zn metal fume (91.5% Zn), were also evaluated, and they were found to have Zn RBV values of 67 (P<.05) and 36% (P<.05), respectively. Feed-grade sources of ZnO vary widely in color, texture, Zn content, and Zn bioavailability. PMID- 10521035 TI - Predicting bull growth performance and carcass composition from growth hormone response to growth hormone-releasing hormone. AB - Development of practical, physiologically based methods that provide an early, yet accurate, evaluation of a bull's genetic merit could benefit the beef industry. The use of GH response to a single, acute dose of GHRH was evaluated as a predictor of future growth performance and carcass characteristics of weanling bulls. Fifty-six Angus bulls averaging 229 d (SD = 27) of age were administered three doses i.v. (0, 1.5, and 4.5 microg/100 kg BW) of human GHRH (1-29) analog in a Latin square design balanced for residual effects. Blood samples were collected via jugular catheter at -60, -45, -30, -15, 0, 5, 10, 15, 30, 45, 60, 90 and 120 min relative to GHRH injection. Serum concentrations of GH were plotted over time. Response to GHRH was calculated as the area under the GH response curve (AUC-GH) using the trapezoidal approximation. Relationships between AUC-GH, weaning weight adjusted to 205 d of age (205-d WW), and direct weaning weight EPD (WWEPD) versus age-adjusted BW (BWadj), ADG, and carcass measurements from a 140-d growth performance test were evaluated using simple linear regression. A positive correlation between AUC-GH and ADG and an inverse relationship between AUC-GH and carcass fat were observed. The present study provides evidence that AUC-GH is a better predictor of future growth performance in beef bulls than 205-d WW or WWEPD values. Thus, GH response to GHRH is associated with subsequent growth and may be a useful tool for sire selection in beef production. PMID- 10521036 TI - Effect of estrus synchronization with norgestomet on the integrity of oocytes from persistent follicles in beef cattle. AB - Our objective was to determine whether oocyte integrity is compromised when oocytes are recovered from progestogen-induced persistent follicles. Beef cows were presynchronized using PGF2alpha (PGF). Cows detected in estrus after PGF were assigned to either NOR (one 6-mg norgestomet implant for 10 d starting on d 16 of cycle; day 0 = estrus; n = 112) or CON (control, no implant [n = 128] and presynchronized 8 d later than NOR). All cows received 25 mg of PGF at the end of treatment (NOR, d 26; CON, d 18). Treatments produced persistent preovulatory follicles (NOR) or normal preovulatory-size follicles (CON), which were measured via ultrasonography 1 d before slaughter. Ovaries were collected from all animals (NOR, d 27; CON, d 19) along with random (RAN) ovaries from cattle slaughtered on the same days. Cumulus oocyte complexes (COC) were aspirated from the preovulatory follicles with recovery rates of 63% across treatments. Small follicles (2 to 7 mm diameter) from NOR, CON, and RAN cows were also aspirated to recover COC. Preovulatory follicles were larger (19.5+/-.9 vs. 13.6+/-.4 mm, P<.05), serum P4 was lower (.4+/-.1 vs. 3.9+/-.2 ng/mL, P<.05), and serum E2 was higher (28.7+/-1.6 vs. 7.6+/-.8 pg/mL, P<.05) in NOR than in CON cows. Cumulus oocyte complexes recovered from preovulatory follicles (62 NOR, 64 CON) were matured, fertilized, and cultured in vitro for comparison of embryonic development. A subset (24 NOR, 34 CON) of COC were assigned morphological quality grades. A separate set of recovered COC (10 NOR, 15 CON) was fixed within 1 h after recovery for assessment of the stage of meiosis. Treatments did not differ for oocyte quality grade or stage of meiosis. However, COC from NOR cows had more layers of cumulus cells (P<.05), and more of those COC had undergone cumulus expansion (29.2 vs. 5.9%, P<.05 for NOR vs. CON, respectively). Development of cleaved embryos to the morula and blastocyst stages from preovulatory follicles (22.6% NOR, 18.9% CON) or small follicles (42% NOR, 40% CON, 42% RAN) did not differ with treatment. Oocyte quality and in vitro developmental competence were not compromised for oocytes from induced persistent follicles compared with oocytes from normal preovulatory follicles. Increased expansion of cumulus cells associated with oocytes from progestogen-induced persistent follicles may be relevant to the reduction of in vivo fertility associated with such follicles. PMID- 10521037 TI - Effects of induced hypothyroidism on ovarian response to superovulation in Brahman (Bos indicus) cows. AB - To evaluate the effects of hypothyroidism on ovarian function, multiparous, nonlactating Brahman cows (n = 18) were assigned randomly to dietary treatments containing either 0 (C; n = 9) or 4 mg x kg BW(-1) x d(-1) 6-n-propyl-2 thiouracil (PTU; n = 9), to suppress thyroid function, in the feed concentrate. Weekly changes in BW and body condition score (BCS) were recorded. Dietary treatments began on d 10 of the estrous cycle. Ten days after the first treatment estrus, all cows received daily i.m. injections of 25 IU of porcine FSH over a 3 d period. Seven days after AI, embryos were collected nonsurgically, and the ovaries were removed via midflank laparotomy. Based on thyroxine (T4) concentrations after 49 d of treatment, five cows were hypothyroid (H-PTU) and four were partially suppressed (P-PTU). Cows in the PTU group had greater (P<.01) ADG, (P<.05) ovarian weights, and numbers of large (> or =8 mm) (P<.05) follicles. Cows in the PTU group had lower embryo recovery rate (P<.001), fertilization rate (P<.001), and percentage of blastocysts (P<.1) than C cows. The H-PTU cows had greater numbers of luteinized follicles (P<.06), greater concentrations of progesterone (P4) in the follicular fluid at all size categories (P<.1), and greater numbers of corpora lutea (P<.05) than C cows. The ratio of luteal to serum P4 on d 7 was greater (P<.05) in hypothyroid cows. Induced hypothyroidism improved weight gain and BCS, increased ovarian response to FSH, and affected ovulation, fertility, and P4 secretion in superovulated Brahman cows. PMID- 10521038 TI - Effects of level of energy intake and energy demand on growth hormone, insulin, and metabolites in Targhee and Suffolk ewes. AB - Yearling ewes (n = 32) were used in a 2x2x2 factorial experiment to determine effects of breed (Targhee vs. Suffolk), energy intake (1x vs. 3x NEm requirements, and physiological status (nonpregnant, nonlactating vs. lactating) on serum GH, insulin, NEFA, glucose, and blood urea nitrogen (BUN) concentrations. Blood collections were made in two periods that began 21 and 32 d after ewes lambed. Lactating ewes had more GH peaks (P<.10), higher (P<.01) mean GH concentration, and greater (P<.01) area under the GH curve (AUC) than nonlactating ewes. The AUC was greater (P<.01) in ewes fed 1x NEm than in ewes fed 3x NEm. Energy intake had no effect on serum GH before feeding (P>.23) when evaluated within physiological statuses. After feeding, GH concentrations were greater (P<.10) for ewes fed 1x NEm than for those fed 3x NEm. Insulin and glucose did not differ (P>.23) between energy intake levels. Insulin and glucose were greater (P<.001) in nonlactating than in lactating ewes when evaluated within breed. Lactating and Targhee ewes fed 1x NEm had greater (P<.001) NEFA concentration than nonlactating and Targhee ewes fed 3x NEm, respectively. Ewes fed 3x NEm and Targhee ewes had greater (P<.005) BUN concentrations than ewes fed 1x NEm and Suffolk ewes, respectively. Physiological status seems to play a more important role in the regulation of GH than does energy intake. Higher BUN concentrations in Targhee than in Suffolk ewes demonstrates one metabolic event that distinguishes a breed's adaptation to the environment in which it originated. PMID- 10521039 TI - Voluntary intake, digestibility, and subsequent selection of Matua bromegrass, coastal bermudagrass, and alfalfa hays by yearling horses. AB - Matua bromegrass (Bromus willdenowii Kunth. cv. Grasslands Matua) was introduced in 1973, but little information exists concerning its potential as a hay for horses. Thus, voluntary intake and apparent digestibility of OM, CP, and fiber components of Matua by 18 Quarter Horse yearlings (mean initial BW 354 kg; SE 5.8) were compared with alfalfa (Medicago sativa L.) and coastal bermudagrass (Cynodon dactylon L.) as hays in a randomized block design. A 15-d adjustment period was followed by a 5-d collection period during which the hays were consumed ad libitum. Voluntary intake of DM was greater (P<.01) for alfalfa (10.9 kg/d) than for the mean of the grasses, and intake of Matua (10.0 kg/d) was greater (P<.001) than that of bermudagrass (7.4 kg/d). Apparent digestibility of OM was greater (P<.001) for alfalfa (74%) than for the mean of the grasses but did not differ between Matua (64%) and bermudagrass (60%). At the end of the digestion trial, each yearling was offered each of the three forage hays during an 11-d period to determine subsequent preference and effect of previous hay experience. Yearlings preferred alfalfa over the grass hays and generally selected more Matua than bermudagrass. All yearlings consumed less of the forage species to which they had been previously exposed compared with unadapted yearlings. The Matua hay fed in this trial was palatable and met most of the nutritional needs for yearling horses. PMID- 10521040 TI - Low- and high-quality forage utilization by heifers and mature beef cows. AB - Eight cows (7 to 9 yr old, 522 kg) and six heifers (10 mo old, 169 kg) were fed either alfalfa hay (18.7% CP) or mature brome hay (5.1% CP) to determine the effect of cattle age on apparent forage utilization. Cattle were fitted with ruminal and duodenal cannulas and were individually fed once daily (ad libitum intake, 1000). The split-plot design consisted of age (whole-plot) and two sampling periods feeding alfalfa or brome hay (subplot). Each period consisted of 28 d: d 1 to 13 for adaptation, d 13 to 20 for feed intake determination, and d 20 to 28 for sampling. Nylon bags containing NDF substrate from alfalfa or brome hay were incubated ruminally for 0, 3, 6, 12, 24, 48, 96, and 192 h to determine the rate and extent of fiber degradation. Ruminal liquid dilution rate and fermentation characteristics were conducted on d 27. Ruminal fill was determined by total evacuation at 0800 on d 28. Cows consumed more feed (BW.75; P<.01) and had greater ruminal OM fill (P = .04) but had similar fluid fill (P = .88) compared with heifers. Ruminal liquid dilution rate was greater in cows than in heifers (P<.01). The rate of in situ NDF degradation was 3 and .5% per hour greater in cows than in heifers when alfalfa and brome hay were fed, respectively (age x hay, P<.01). Ruminal NDF digestibility as a percentage of intake was greater in cows than in heifers (P<.01). Numbers of ruminal cellulolytic bacteria were not affected by treatment (P>.21). These data indicate that mature cows have a smaller ruminal fluid fill that turns over more rapidly, and this may be responsible for a faster rate of ruminal fiber degradation in cows than in young heifers. PMID- 10521041 TI - Influence of postruminal supplementation of methionine and lysine, isoleucine, or all three amino acids on intake and chewing behavior, ruminal fermentation, and milk and milk component production. AB - Four multiparous Holstein cows were fed a basal diet balanced with the Cornell Net Protein and Carbohydrate System (CNCPS). Diets were formulated to be co limiting in intestinally absorbable supplies of methionine, lysine, and isoleucine. Cows were supplemented with no amino acids (control); lysine and methionine in a ruminally protected form; isoleucine by abomasal infusion; or lysine, methionine, and isoleucine in a 4x4 Latin square arrangement of treatments with 28-d periods. Performance of cows on all treatments was lower than expected due to low intake of DM that could have been caused by the high fiber level of the basal diet. This high fiber level was likely responsible for the high daily chewing times for cows fed all diets, which was consistent with the high ruminal pH values. Intake of DM and its components were not influenced by any treatment. Milk protein percentage tended to be higher when cows were fed diets supplemented with ruminally protected lysine and methionine; however, production of milk, milk fat, and milk lactose were not affected by any treatment. Cows tended to have a higher milk lactose proportion and tended to produce more milk and milk lactose when they were abomasally infused with isoleucine alone. However, when cows were supplemented with all three amino acids, milk production and composition did not differ from that of cows fed the unsupplemented diet. Use of the CNCPS to evaluate the performance of the cows fed the unsupplemented diet suggested that these cows may have been colimited by intestinally absorbable supplies of lysine, isoleucine, and methionine in addition to metabolizable protein. Evaluation of the unsupplemented diet with an alternate model, Shield, suggested that cows fed the unsupplemented diet may have been colimited by intestinally absorbable supplies of lysine, isoleucine, and arginine. Results suggest that enhanced delivery of intestinally absorbable isoleucine may stimulate milk lactose synthesis. PMID- 10521042 TI - Effects of different supplemental sugars and starch fed in combination with degradable intake protein on low-quality forage use by beef steers. AB - Twenty ruminally fistulated steers (Exp. 1, 448 kg and Exp. 2, 450 kg) were used in two consecutive randomized complete block experiments with five treatments in each experiment. The purpose was to evaluate the impact of feeding different supplemental sugars or starch in combination with supplemental degradable intake protein (DIP) on the utilization of low-quality tallgrass-prairie hay. In Exp. 1, steers were given ad libitum access to forage and, except for the negative control (NC), received a limited supply (insufficient to maximize forage use) of supplemental DIP (.031% BW/d, DM basis). In addition to the NC, this experiment included four supplementation treatments in which one of four carbohydrate (CHO) sources (starch, glucose, fructose, or sucrose) was fed at .30% BW of DM/d. In Exp. 2, the treatment structure was identical except that the supplemental DIP level (.122% BW, DM basis) was near the level needed to maximize forage use. Forage OM intake (FOMI) was not affected (P> or =.26) by supplementation in Exp. 1 but was increased (P = .05) in Exp. 2. However, no difference (P> or =.46) in FOMI occurred among CHO sources in either experiment. Total OM and digestible OM intakes were increased (P<.01) by supplementation in both experiments. In Exp. 1, no difference (P>.26) in OM digestion (OMD) occurred among treatments. In Exp. 2, supplementation increased (P<.01) OMD. Additionally, sugars yielded a higher (P = .04) OMD than starch, and the monosaccharides yielded a higher (P = .02) OMD than sucrose. In Exp. 1, NDF digestion (NDFD) was decreased (P = .02) by supplementation, but no differences (P> or =.21) occurred among CHO sources. In Exp. 2, NDFD was increased (P = .03) by supplementation. Additionally, sugars led to higher (P = .05) NDFD than starch, and the monosaccharides led to higher (P = .03) NDFD than sucrose. In both experiments, discernible patterns were observable with regard to the effects of supplementation and type of supplemental CHO on ruminal fermentation characteristics. In conclusion, even though some consistency in fermentation profiles for different carbohydrate sources was evident in both experiments, forage intake and digestion responses were not consistent across experiments. This raises the possibility that carbohydrate source may interact with the amount of supplemental DIP fed and, as such, deserves additional investigation. PMID- 10521043 TI - In situ disappearance of neutral detergent insoluble nitrogen from alfalfa and eastern gamagrass at three maturities. AB - In situ digestion kinetics of neutral detergent insoluble nitrogen (NDIN) from alfalfa (Medicago sativa L.) harvested at one-tenth bloom and eastern gamagrass (Tripsacum dactyloides L.) harvested at the boot (GGB), anthesis (GGA), and physiological maturity (GGM) stages of growth were determined with nonlinear regression techniques. Whole-plant tissue and associated leaf and stem fractions were incubated in the ventral rumen simultaneously. On a wholeplant basis, potential extents of degradation were particularly high (> or =904 g/kg NDIN) for GGB and GGA, relative to those of GGM and alfalfa (772 and 658 g/kg NDIN, respectively). For all plant parts, degradation rates of NDIN were faster (P<.05) for alfalfa than for all gamagrass forages. Degradation rate of NDIN did not differ (P>.05) across maturities for any gamagrass tissue type. These results indicate 1) that phenological development and lignification do not limit the rate of NDIN degradation in gamagrass forages but do markedly limit the potential extent of NDIN availability and 2) that most of the NDIN in these forages is potentially available in the rumen and can contribute to the ruminal N supply. Our secondary objective was to compare estimates of N escaping ruminal degradation that were determined on the basis of NDIN degradation kinetics (NDIN method) with those determined traditionally, on the basis of total residual N. The NDIN method mathematically eliminates all neutral detergent soluble N from consideration as part of the pool of dietary N potentially escaping the rumen intact. Estimates of rumen escape nitrogen determined on the basis of degradation rates of NDIN were consistently less than corresponding estimates that were determined on the basis of total residual N. When ruminal escape N that was determined with the NDIN method was regressed on corresponding estimates with the total residual N method, the slopes of the regression lines were .53 and .66 for assumed passage rates of .02 and .06 h(-1), respectively. For the forages evaluated in this study, these results indicate that neutral detergent soluble N may make important contributions to the pool of N escaping ruminal degradation. PMID- 10521044 TI - Evaluation of wheat-based thin stillage as a water source for growing and finishing beef cattle. AB - Two trials were conducted to evaluate the nutritional value of wheat-based thin stillage as a water source for cattle. In Trial 1, 20 large-framed steers were fed a basal diet based primarily on barley grain and barley silage, with ad libitum access to water or thin stillage at one of three DM concentrations (2, 4, and 6.7%) in a completely randomized design. The trial consisted of a 70-d growing period and a finishing phase. In Trial 2, total-tract nutrient digestibility coefficients of the basal diet and water treatments fed in the growing period were determined in a randomized complete block design using 12 medium-framed steers. The results showed that when only DMI from the basal diet was considered, there was a linear reduction (P<.01) in DMI and a linear improvement (P<.01) in the gain:feed ratio with no effect on daily gain as thin stillage DM concentration increased. No differences were detected in DMI or efficiency of gain when total DMI (basal diet and thin stillage) was considered. Carcass traits indicated a trend toward increased (P<.06) carcass fat with increasing thin stillage DM concentration. Results of Trial 2 indicated a linear improvement (P<.05) in apparent digestibility of DM, CP, NDF, and energy of the total diet (basal diet and thin stillage) as thin stillage DM concentration increased. We concluded that supplementing growing and finishing cattle with thin stillage reduced the amount of the basal diet required for gain and improved nutrient utilization. PMID- 10521045 TI - Feeding value of wheat-based thin stillage: in vitro protein degradability and effects on ruminal fermentation. AB - Two experiments were conducted to evaluate the nutritive value of wheat-based thin stillage as a fluid source for ruminants. In vitro CP degradability of thin stillage was estimated relative to canola meal and heated canola meal in a completely randomized design. Four ruminally cannulated steers were used in a double cross-over design to determine the effects of consuming thin stillage or water as drinking sources on ruminal fermentation traits. The in vitro CP degradability of thin stillage (55.4%) was lower (P<.05) than that of canola meal (59.4%) and higher than that of heated canola meal (31.6%). Ruminal pH for steers consuming thin stillage was higher (P<.05) at 1000 and 1100 and lower (P<.05) at 1900 and 2000 than that for steers consuming water. Total VFA followed a pattern that was the reverse of that reported for pH. Ruminal NH3 N levels were higher (P<.05) for steers fed thin stillage than for water-fed steers through most of the collection period. Ruminal fluid and particulate matter passage rates were not affected by treatment and averaged .165 and .06 /h, respectively. The amount of thin stillage and water that did not equilibrate with the ruminal fluid and, thus, was considered to bypass the rumen was estimated to be 51.9 and 59.2% of total fluid consumed, respectively. Feeding wheat-based thin stillage had no adverse effects on ruminal metabolism. PMID- 10521046 TI - Steam-processed corn and sorghum grain flaked at different densities alter ruminal, small intestinal, and total tract digestibility of starch by steers. AB - Crossbred steers (n = 7; 400 kg BW), fitted with T-type cannulas in the duodenum and ileum, were used to examine the effects of processing method, dry-rolled (DR) vs. steam-flaked (SF) sorghum grain, and degree of processing (flake density; FD) of SF corn (SFC) and SF sorghum (SFS) grain on site and extent of DM, starch, and N digestibilities and to measure extent of microbial N flow to the duodenum. In Exp. 1, diets contained 77% DRS or 77% SFS with FD of 437, 360, and 283 g/L (SF34, SF28, and SF22). In Exp. 2, diets contained 77% SFC with FD of SF34 or SF22. For sorghum and corn diets, respective average daily intakes were as follows: DM, 6.7 and 8.1 kg; starch, 3.8 and 4.7 kg; N, 136 and 149 g. Steers fed SFS vs. DRS increased (P = .01) starch digestibilities (percentage of intake) in the rumen (82 vs. 67%) and total tract (98.9 vs. 96.5%) and decreased digestibilities in the small intestine (16 vs. 28%; P = .01) and large intestine (.5 vs 1.2%; P = .05). As a percentage of starch entering the segment, digestibility was increased (P = .01) within the small intestine (91 vs. 85%) but was not altered within the large intestine by steers fed SFS vs. DRS. Decreasing FD of SFS and of SFC, respectively, linearly increased starch digestibilities (percentage of intake) in the rumen (P = .03, .02) and total tract (P = .03, .09) and linearly diminished starch digestibilities in the small intestine (P = .04, .09). Starch digestibilities (percentage of entry) within the small or large intestine were not changed by FD. The percentage of dietary corn or sorghum starch digested in the large intestine was very small, less than 2% of intake. Microbial N flow to the duodenum was not altered by SFS compared to DRS, or by decreasing FD of SFS and SFC. Reducing FD of SFS, but not of SFC, tended to decrease (P = .07) microbial efficiency linearly and tended to increase (P = .06) total tract N digestibilities linearly. Steam flaking compared to dry rolling of sorghum grain and decreasing FD of SFC and SFS grain consistently increased starch digestibility in the rumen and total tract of growing steers. The greatest total digestibility of dietary starch occurred when the proportion digested in the rumen was maximized and the fraction digested in the small intestine was minimized. These changes in sites of digestion account, in part, for the improved N conservation and greater hepatic output of glucose by steers fed lower FD of SFS reported in our companion papers. PMID- 10521048 TI - Adverse effects of excess DL-methionine in calves with different body weights. AB - Adverse effects of excess methionine were examined using 12 Holstein bull calves trained to maintain reflex closure of the reticular groove even after weaning at 5 wk of age. Two nitrogen balance experiments were conducted for 2 wk each from 6 wk (Stage 1; BW = 62 kg) and 12 wk of age (Stage 2; BW = 103 kg) by dividing the calves into three groups at each stage. Calves were fed a corn-soybean meal diet at 62 g/kg of metabolic BW at both stages. At Stage 1, feed efficiency (gain:feed intake) and nitrogen retention did not differ between the group supplemented with .333 g of DL-methionine and .111 g of L-lysine monohydrochloride/kg BW per day and the group supplemented with isonitrogenous diammonium citrate, although the level of DL-methionine was considered to be enough to induce toxicity. Conversely, administration of isonitrogenous casein increased nitrogen retention. At Stage 2, administration of the same levels of methionine and lysine resulted in reduced feed intake, depressed nitrogen retention, and BW loss. Conversely, administration of the isonitrogenous casein did not increase nitrogen retention compared with the supplement of isonitrogenous diammonium citrate. Administration of excess methionine and lysine increased plasma methionine concentrations up to 230 (Stage 1) or 190 micromol/dL (Stage 2). Plasma lysine concentrations were less than 24 micromol/dL at every stage. Administration of the amino acid mixture decreased plasma concentrations of branched-chain amino acids and phenylalanine more obviously at Stage 2 than at Stage 1. These results indicated that abomasal administration of .333 g of DL-methionine/kg BW per day induced methionine toxicity at Stage 2 but methionine imbalance at Stage 1. PMID- 10521047 TI - Effect of dietary phosphorus on finishing steer performance, bone status, and carcass maturity. AB - Yearling crossbred steers (n = 60; 386 kg) were individually fed in a completely randomized experimental design to determine their P requirement. Treatments were in a factorial arrangement with two levels of Ca (.35 or .70% of DM) and five concentrations of P (.14, .19, .24, .29, or .34% of DM). The finishing diet consisted of 34.5% dry-rolled corn, 22.5% brewers grits, 22.5% corn bran, 7.5% ground corncobs, 5% molasses, 3% fat, and 5% supplement. Supplemental P was provided as monosodium phosphate and Ca as limestone. Ash content was determined on the first phalanx bone from the lower front legs following slaughter, and rib bone breaking strength was determined with an Instron Universal Testing Machine. Carcass maturity and shear force were also evaluated on wholesale rib cuts. Because no interactions between Ca and P levels were detected, only main effects are presented. Daily gain, DMI, and feed efficiency were not affected by dietary P concentration or P intake. Bone ash (g or g/100 kg BW) and rib bone breaking strength were also unaffected by dietary P. Feeding .7% Ca decreased (P<.06) ADG and efficiency compared with feeding .35% Ca. Neither dietary Ca nor P had a significant effect on tenderness (shear force), skeletal maturity, or overall maturity. These results indicate that the P requirement for finishing yearlings is .14% of diet DM or less and that supplementing P above levels supplied by basal ingredients in many grain-based finishing diets is not necessary. PMID- 10521050 TI - Rapid communication: mapping of the glutathione-peroxidase-5 (GPX5) gene to pig chromosome 7. PMID- 10521049 TI - Effects of level and source of carbohydrate and level of degradable intake protein on intake and digestion of low-quality tallgrass-prairie hay by beef steers. AB - Ruminally fistulated steers (n = 13; 263 kg) were used in an incomplete Latin square with 13 treatments and four periods to evaluate the effects of level and source of supplemental carbohydrate (CHO) and level of degradable intake protein (DIP) on the utilization of low-quality, tallgrass-prairie hay. Steers were given ad libitum access to forage (5.7% CP, 2.6% DIP, and 74.9% NDF). The supplementation treatments were fashioned as a 2x3x2 factorial arrangement plus a negative control (NC; no supplement). The factors included two DIP levels (.031 and .122% BW) and three CHO sources (starch, glucose, and fiber) fed at two levels (.15 and .30% BW) within each level of DIP supplementation. The effect of supplementation on forage OM intake (FOMI) was dependent (P<.01) on level and source of CHO and level of DIP fed. When DIP was low, forage, total, and digestible OM intakes were generally greater for the starch treatment than for the nonstarch treatments. However, when the DIP level was high, intakes were greater for the nonstarch (i.e., fiber and glucose) treatments. Generally, FOMI decreased (P<.01) when more supplemental CHO was provided. Supplementation typically increased fiber digestion, but the response was dependent (P<.01) on level and source of CHO and level of DIP. Generally, supplements with low levels of CHO improved NDF digestion (NDFD). However, supplements with the high level of CHO decreased NDFD, except for fiber at the high level of DIP. Organic matter digestion was increased by supplementation, but the impact of increasing CHO was dependent (P<.01) on source of CHO and level of DIP. Supplementation treatments had significant impact on ruminal pH, NH3 N, and the total concentration of organic acids as well as their relative proportions. In conclusion, supplemental DIP enhanced the use of low-quality forage; however, the impact of supplemental CHO on low-quality forage use was dependent on source and level of CHO offered, as well as the level of DIP provided. PMID- 10521051 TI - Rapid communication: mapping of the titin (TTN) gene to pig chromosome 15. PMID- 10521052 TI - Impact of the mode of detection on outcome in breast cancer patients treated with breast-conserving therapy. AB - The impact of the mode of detection on outcome in patients with early stage breast cancer treated with breast-conserving therapy (BCT) was reviewed. Between January 1980 and December 1987, 400 cases of stage I and II breast cancer were treated with BCT. All patients underwent an excisional biopsy, external beam irradiation (RT) to the whole breast (45-50 Gy), and a boost to 60 Gy to the tumor bed. One hundred twenty-four cases (31%) were mammographically detected, whereas 276 (69%) were clinically detected. Median follow-up was 9.2 years. Patients whose cancers were detected by mammography more frequently had smaller tumors (90% T1 vs. 62%, p < 0.0001), lower overall disease stage (78% stage I vs. 47%, p < 0.0001), were older at diagnosis (78% >50 years vs. 54%, p < 0.001), less frequently received chemotherapy (8% vs. 21%, p = 0.001), and had an improved disease-free survival (DFS) (80% vs. 70%, p = 0.014), overall survival (OS) (82% vs. 70%, p = 0.005), and cause-specific survival (CSS) (88% vs. 77%, p = 0.003) at 10 years. However, controlling for tumor size, nodal status, and age, no statistically significant differences in the 5- and 10-year actuarial rates of local recurrence (LR), DFS, CSS, or OS were seen based on the mode of detection. Initial mode of detection was the strongest predictor of outcome after a LR. The 3-year DFS rate after LR was significantly better in initially mammographically detected versus clinically detected cases (100% vs. 61%, p = 0.011). Patients with mammographically detected breast cancer generally have smaller tumors and lower overall disease stage at presentation. However, the mode of detection does not independently appear to affect the success of BCT in these patients. PMID- 10521053 TI - Early onset of breast carcinoma in African American women with poor prognostic factors. AB - The purpose of this study was to determine prognostic significance of age and race as independent variables and to see role of age at the onset of breast carcinoma. A retrospective study was conducted of African American and white women with breast cancer treated at SUNY-Health Science Center Brooklyn and Kings County Hospital Center from 1983 to 1993. The objective was to analyze the differences in patterns of disease onset, as related to age and prognostic factors. A total of 738 patients were analyzed for race-adjusted comparison of stage, grade, disease-free survival, and median survival. Age at the time of diagnosis was analyzed to conduct age-specific comparisons of African American (AA) and white patients. The multivariate analysis indicated that AA women develop breast cancer 10 years earlier than white women (p = 0.00001). Corrected by stage and grade, i.e., chi2 test for stage-by-stage and grade-by-grade analysis has revealed that the AA women present with higher stage (p = 0.009), increased number of positive nodes (p = 0.00007), and more estrogen receptor/ progesterone receptor-negative tumors (p = 0.005). Further studies are required to probe into the etiologic possibilities of this significant difference. The important contributing factors could be hormonal, genetic, environmental, and socioeconomic. Obesity and dietary factors also need to be evaluated. Further studies to explore genetic susceptibility by ploidy is recommended to explain this significant difference. We conclude that the onset of breast cancer among AA women occurs at a significantly younger age than in white women, and their prognostic factors are poorer. PMID- 10521054 TI - High-dose hemithorax irradiation in a patient with recurrent thymoma: a study of pulmonary and cardiac radiation tolerance. AB - Malignancy spread throughout a hemithorax without distant metastasis poses a difficult therapeutic challenge. Irradiation is often not considered because of the risk of pulmonary and cardiac toxicity. We report on a patient with thymoma recurrent throughout the left pleural cavity. Disease progressed despite chemotherapy, and subsequently a radical course of radiotherapy (6,600 cGy) was delivered to the entire hemithorax. Tumor regressed markedly by the completion of radiotherapy. Although tumor regrowth was noted 1 year after radiotherapy, the patient remained markedly improved symptomatically until shortly before her death 2 years after radiotherapy. Pulmonary function tests at 1 year (forced expiratory volume of the first second and forced vital capacity) were similar to pretreatment values, and cardiac function at 2 years remained essentially normal. High-dose hemithorax irradiation may be a consideration in select cases. PMID- 10521055 TI - 'Neo-FAC' (5-fluorouracil, doxorubicin, and cyclophosphamide) for poor-prognosis stage IV breast cancer: a Southwest Oncology Group Phase II Study. AB - The authors report a phase II pilot investigation in the Southwest Oncology Group examining a combination of 5-fluorouracil, doxorubicin, and cyclophosphamide (FAC) incorporating modulated 5-FU in patients with poor-prognosis stage IV breast cancer. Patients with poor-prognosis stage IV breast cancer were treated with this "neo-FAC" as front-line therapy. The regimen consisted of 5 fluorouracil by continuous ambulatory infusion pump at 200 mg/m2/day for 42 days, repeated at 56-day intervals; doxorubicin at 20 mg/m2/week intravenously to a maximum cumulative total dose (including adjuvant therapy, if any) of 500 mg/m2; cyclophosphamide 60 mg/m2/day taken orally; methotrexate 15 mg/m2/week intravenously beginning 1 week after termination of doxorubicin; and oral prednisone decreasing from 60 mg/day on a tapering schedule for a total of 7 weeks of treatment. Treatment was continued until progression, unacceptable toxicity, or patient refusal. Twenty-four patients were accrued to this study. Of these, two were ineligible, and the remaining 22 were evaluable for response. Ten patients experienced grade 3 toxicity, and six had grade 4. There were no treatment-associated deaths. Best responses were a complete response in one patient (5%) and partial responses in 6, for an overall response rate of 32% (7/22 evaluable patients). Overall survival in five pilot studies in the Southwest Oncology Group in this poor-prognosis population are relatively superimposable. The present regimen, with its relatively poor outcome and the expense and inconvenience of administering chemotherapy by ambulatory infusion pump, will not be pursued further. PMID- 10521056 TI - Gemcitabine salvage chemotherapy for patients with gynecologic malignancies of the ovary, fallopian tube, and peritoneum. AB - The efficacy and toxicity of gemcitabine salvage chemotherapy was evaluated in 27 heavily pretreated patients with recurrent and progressive ovarian, fallopian tube, or peritoneal cancer. At least one platinum-based chemotherapeutic regimen had failed in each patient. The median number of previous chemotherapy regimens and cycles of chemotherapy was 4 and 23, respectively. A total of 124 cycles of gemcitabine were delivered (median, 3 cycles). Hematologic toxicity included four patients with grade 3/4 thrombocytopenia and two patients with grade 3/4 neutropenia. Thrombocytopenia and neutropenia resulted in eight dose reductions and a single 1-week treatment delay. Nonhematologic side effects were well tolerated and largely self-limiting. No complete responses were observed. Three patients (11%) demonstrated partial responses to therapy. The duration of response was 7 months for two of the responders and 5 months for the third responder. Stable disease was observed in 14 patients (52%), in whom the median progression-free interval was 5 months. In conclusion, among heavily pretreated patients, gemcitabine has limited antitumor activity in platinum-resistant carcinomas of the ovary, fallopian tube, and peritoneum. The role of gemcitabine in the treatment of gynecologic malignancies of the ovary, fallopian tube, and peritoneum will be determined by studies that define the efficacy of multiagent regimens of chemotherapy that include gemcitabine and by studies that include patients who have been less heavily pretreated. PMID- 10521057 TI - Clinical factors and prognosis in non-small cell lung cancer. AB - We evaluated the relationship of clinical characteristics and survival in 1,635 patients with non-small cell lung cancer (NSCLC) treated in Brazil. The following variables were included: sex, age, smoking, Karnofsky's performance status (PS), weight loss, symptoms at diagnosis (cough, dyspnea, hemoptysis, chest pain, wheezing, and hoarseness), presence of superior vena cava syndrome (SVCS), histologic type, TNM stage, and therapeutic modality (surgery, chemotherapy [CT] and radiotherapy [RT]). Multivariate prognostic models were obtained by Cox regression. Patients unsuitable for surgery or who had recidivant disease were elected to further RT and/or CT, and long-term results in this group were equivalent to those in the group treated only by surgery. A diagnosis of bronchioloalveolar carcinoma, small tumors, absence of hoarseness, treatment by surgery, and RT were independent factors related to good overall survival in stage I and II. Weight loss and clinical signs of SVCS were related to poor prognosis in stage III. PS, diagnosis of adenocarcinoma or undifferentiated carcinoma, absence of weight loss and dyspnea, NO or N1 disease, ability to receive RT, CT, and to perform some palliative surgical procedure were good prognostic factors in stage IV. Clinical features of patients with NSCLC at diagnosis offer additional information to estimate their prognosis. PMID- 10521058 TI - Primary appendiceal adenocarcinoma. AB - Adenocarcinoma of the appendix is rarely encountered and is usually discovered at the pathology examination of the surgical specimen. Adenocarcinoma of the vermiform appendix is a rare neoplasm and constitutes <0.5% of all gastrointestinal neoplasms. There is no symptom of appendiceal cancer, and it is very difficult to diagnose preoperatively. Most female patients are diagnosed as having a gynecologic disease. Second primary synchronous and metachronous neoplasms, especially in the gastrointestinal tract, are found in up to 35% of patients with appendix adenocarcinoma. We report a case of adenocarcinoma in a 56 year-old woman misdiagnosed as having right ovarian carcinoma, and we review the literature. PMID- 10521059 TI - Primary cardiac sarcoma in pregnancy: a case report and review of the literature. AB - Primary cardiac sarcoma (PCS) is a rare disease with a poor prognosis, because of diagnostic delay, therapeutic difficulties, and high metastatic potential. Surgery is the standard treatment. A case of PCS in pregnancy is reported, with a review of published surgical series of PCSs, focusing on the role of surgery and adjuvant therapy. Prompt surgery improved cardiac function and patients' outcome in comparison with untreated cases. The role of adjuvant treatment was analyzed only in a few series, mainly without distinction between postoperative chemotherapy and radiotherapy; adjuvant therapy improved survival in the larger series of resected PCSs. Only three other cases of PCS in pregnancy were reported. In the present case, resection was performed with no major complication for the mother and the infant. Even if the patient's survival was short, cardiac surgery allowed prolonging of pregnancy until an acceptable possibility of fetal survival was reached. Although resection is not curative in most cases, surgery remains the treatment of choice for PCS and has a definite palliative significance. The role of postoperative chemotherapy and radiotherapy is difficult to ascertain; however, adjuvant chemotherapy seems advisable in high grade tumors. PMID- 10521060 TI - Severe cardiotoxicity during 5-fluorouracil chemotherapy: a case and literature report. AB - The chemotherapeutic agent 5-fluorouracil (5-FU) is a widely accepted part of many cancer treatment protocols. Its cardiotoxic potential is known, but considered uncommon and usually not life threatening, although some cases of severe cardiotoxicity related to 5-FU have been reported. The pathogenesis of cardiotoxicity caused by 5-FU is not clear. We report a case of sudden onset of severe cardiac failure, without ischemic symptoms or signs, during 5-FU treatment with serious consequences, in a previously healthy 23-year-old patient with squamous cell carcinoma of the tongue. Endomyocardial biopsy showed proliferation of the sarcoplasmic reticulum with marked vacuolization, similar to that found with doxorubicin cardiotoxicity. Because 5-FU cardiotoxicity is unpredictable and can have potentially fatal consequences, it requires, in our opinion, further clarification. With this well-documented case, including an endomyocardial biopsy, we hope to encourage additional efforts to investigate the pathophysiologic mechanisms of 5-FU cardiotoxicity. PMID- 10521061 TI - Combination of epirubicin and cisplatin in hormone-refractory metastatic prostate cancer. AB - Anthracyclines and cisplatin have been shown separately to have modest activity in prostate cancer. The synergism between anthracyclines and cisplatin, with the lack of overlapping toxicities, led to the conduct of this phase II trial of the combination of epirubicin and cisplatin in hormone-refractory metastatic prostate cancer. Twenty-one evaluable patients with hormone-refractory metastatic prostate cancer received epirubicin 100 mg/m2 followed by cisplatin 80 mg/m2 with prehydration and mannitol diuresis. Epirubicin and cisplatin produced a biochemical response (>50% decrease in tumor marker) in 32% of patients, symptomatic improvement in 38%, and a response in measurable and evaluable disease sites in 14%. Toxicities were mainly hematologic, with 77% and 41% >grade 2 neutropenia and thrombocytopenia, respectively. Greater than grade 2 toxicities were: cardiac (three), renal secondary to sepsis (one), nausea and vomiting (two), weakness (one), mucositis (one), and diarrhea (one). The combination of epirubicin and cisplatin was associated with manageable toxicities in this elderly population; however, antitumor activity was marginal in this disease. Participation in clinical trials should continue to be offered to patients with hormone-refractory metastatic prostate cancer. PMID- 10521062 TI - Oral fluoropyrimidines: a closer look at their toxicities. AB - Patient preferences, quality of life issues, and economic considerations are driving the development of orally administered chemotherapy. Oral fluorinated pyrimidines, which have been used in Japan, are attracting increasing interest as a means to provide convenient, less toxic treatment without compromising efficacy. The oral fluoropyrimidines provide prolonged 5-fluorouracil (5-FU) exposure at lower peak concentrations than those observed with bolus intravenous administration. Moreover, depending on the dose schedule, the pharmacokinetics of the oral fluoropyrimidines may mimic the pharmacokinetics of continuous-infusion 5-FU. This review focuses on the toxicity profiles of five emerging oral fluoropyrimidine antineoplastic drugs: combined uracil and tegafur (UFT), capecitabine, eniluracil, S-1, and emitefur (BOF-A2). Different patterns of toxicities emerge from an analysis of the clinical trials of these agents relative to 5-FU administered as an intermittent intravenous bolus or as continuous infusion. The results of ongoing phase III trials comparing the oral fluoropyrimidines with conventional regimens of 5-FU plus leucovorin and 5-FU by continuous intravenous infusion are necessary before their therapeutic role in the management of colorectal carcinoma can be defined. PMID- 10521063 TI - Significance of cytoreductive surgery including bowel resection for patients with advanced ovarian cancer. AB - The aim of this study was to determine the significance of bowel resection in advanced ovarian cancer. A total of 64 women with stage IIIc or IV epithelial ovarian cancer, who consecutively received primary treatment between 1991 and 1995, were entered in this prospective study. The outcome of the patients undergoing bowel resection was evaluated. Thirty-nine patients underwent cytoreductive surgery at initial surgery. Of them, 16 patients could undergo optimal operation without bowel resection. Twenty-three patients received bowel resection at initial surgery. Of these 23 patients, 16 underwent optimal operation and 7 did not. Among 25 patients judged as inoperable cases at initial surgery, 21 responded to chemotherapy and underwent second surgery. Of 21 patients receiving second surgery, 15 underwent optimal operation (7 without bowel resection and 8 with bowel resection). The 3-year survival rate for 24 patients undergoing optimal operation with bowel resection (46.8%) was not significantly different from that for 23 patients without bowel resection (59.1%). Postoperative complications were seen in 8 patients (21.6%) of the patients receiving bowel resection and 3 (13.0%) of those without bowel resection. Cytoreductive surgery including bowel resection is effective for an improvement of the survival in patients with advanced ovarian cancer, if an optimal operation can be performed. PMID- 10521064 TI - Impact of oral submucous fibrosis on chemotherapy-induced mucositis for head and neck cancer in a geographic area in which betel quid chewing is prevalent. AB - In Southeast Asia and Taiwan, betel quid chewing is prevalent. Patients with head and neck cancer who chewed betel quid habitually seem to experience more severe chemotherapy-induced mucositis in our clinical practice. To validate this issue, patients with untreated head and neck cancer who received cisplatin (cDDP) plus a 5-fluorouracil (5-FU)-based neoadjuvant chemotherapy were included in this analysis. Information on the consumption of betel quid, tobacco, and alcohol were recorded before chemotherapy. Oral submucous fibrosis (OSF) was diagnosed clinically according to the fibrotic appearance of the mucosa and trismus. Mucositis was scored according to the World Health Organization criteria, and the mucositis score of the first course of chemotherapy was used for analysis. From December 1993 to April 1996, 120 patients were enrolled in this trial. Neither the betel quid chewing nor the cancer of the oral cavity was to be a significant factor for mucositis. However, clinically diagnosed OSF was found to display a significant correlation with more severe mucositis (p = 0.02). We concluded that in betel quid chewing-prevalent areas, OSF was a risk factor of more severe mucositis in head and neck cancer patients treated by CDDP and 5-FU-based regimens. PMID- 10521065 TI - Resected pancreatic cancer treated with adjuvant radiotherapy with or without 5 fluorouracil: treatment results and patterns of failure. AB - There are relatively little data regarding patterns of recurrence after curative resection and postoperative radiotherapy with or without 5-fluorouracil (5-FU) for patients with adenocarcinonima of the pancreas. Between 1978 and 1997, 41 patients underwent postoperative radiotherapy (RT) at Loyola-Hines Department of Radiotherapy. Of the 38 evaluable patients, 30 had RT + 5-FU and 8 had RT alone. Twenty-nine patients (76.3%) had a Whipple's resection, seven (18.4%) had distal pancreatectomy, and two (5.2%) had total pancreatectomy. Thirty-three (86.8%) of the 38 patients received > or =4,500 cGy to the tumor bed. Median survival for all patients was 21 months. The median survivals for patients who received RT + 5 FU and RT alone were 26 months and 5.5 months (p = 0.004). The most common site of failure was the liver, as seen in 79.2% of all recurrences. The peritoneum, other distant sites (lungs, bone, distant lymph nodes), and locoregional tumor bed were components of failure in 33.3%, 29.2%, and 25.0%, respectively. Locoregional failure alone was found in only one patient. Our median survival with postoperative RT + 5-FU is consistent with results reported by the Gastrointestinal Tumor Study Group and Mayo Clinic. Although patients who had RT + 5-FU had a better median survival than those who received RT alone, our RT alone group had an inferior survival outcome compared to other published reports and may represent patient selection bias. Efforts in controlling this disease should be directed to prevention of intraabdominal relapse. PMID- 10521066 TI - Phase II trial of a novel platinum analog, SKI 2053R, in patients with previously untreated extensive-stage small-cell lung cancer. AB - A phase II trial of a novel platinum analog, SKI 2053R, was performed in patients with previously untreated extensive-stage disease (ED) small-cell lung cancer (SCLC). SKI 2053R was administered at the dose of 400 mg/m2 every 3 to 4 weeks as a 1-h infusion. After the first cycle, the dose was escalated to 440 mg/m2 based on toxicity. Thirty-eight patients (31 male) were enrolled between June 1995 and August 1997. The median age was 61 years (range, 36-70 years). Six of 37 evaluable patients achieved a partial response (16.2%; 95% confidence interval [CI], 4.4-28.0%). The durations of response were 1.1, 1.5, 1.7, 1.9, 3.4, and 4.6 months. The estimated median survival time was 7.4 months (95% CI, 5.1-9.7 months). Grade 3 or 4 toxicities were not observed. Grade 1 to 2 leukopenia, anemia, and thrombocytopenia were seen in 5 of 68 cycles, 16 of 68, and 2 of 68, respectively. Nonhematologic toxicities included grade 1 to 2 nausea or vomiting (30 of 68 cycles), nephrotoxicity (27 of 68), and hepatotoxicity (13 of 68). SKI 2053R showed a modest antitumor activity with limited toxicities in patients with ED SCLC. Further clinical trials are warranted in SCLC with a higher dose of SKI 2053R. PMID- 10521067 TI - Protocols for the preoperative selection of palpable thyroid nodules: review and progress. AB - Modern protocols for the management of patients with palpable thyroid nodules agree that fine-needle aspiration is the first examination to be performed. However, they differ very much in the role attributed to scintigraphy and ultrasound examinations. In some protocols, these two techniques are not considered, whereas in others they are recommended at the end of the diagnostic workup to select for surgery those nodules with nondiagnostic or suspect fine needle aspiration biopsy results. We report original data and literature showing that such use of scintigraphy and ultrasonography is not cost effective. Furthermore, we report original data showing that large-needle aspiration biopsy can be used to select for surgery those nodules with nondiagnostic or suspect results after fine-needle aspiration. Consequently, we suggest a new protocol for the preoperative selection of palpable thyroid nodules. PMID- 10521068 TI - North Central Cancer Treatment Group Phase II study of 5-fluorouracil and high dose levamisole for gastric and gastroesophageal cancer using survival as the primary endpoint of efficacy. AB - At present there is no established standard chemotherapy for advanced gastric cancer. Combination regimens have yielded response rates at times exceeding 50% but with no improvement in survival compared to single agents. This study examined the role of 5-fluorouracil and high-dose levamisole in a phase II setting using survival as the main endpoint. Patients with advanced carcinomas of the stomach or gastroesophageal junction were treated with 5-fluorouracil, 450 mg/m2 IV days 1 to 5, and levamisole, 100 mg/m2 orally three times daily on days 1 to 3, and 50 mg/m2 tid days 4 to 5 every 5 weeks. To allow more rapid accrual and to study a population that more accurately reflects the makeup of patients treated in clinical practice, patients with both measurable and nonmeasurable disease were entered in this study. Two of fifteen (13%) patients with measurable disease experienced a partial response to treatment. The adjusted 1-year survival rate for the 44 patients entered was 29.6%, which is similar to the historical 1 year survival of 30% observed in a group of nearly 400 patients treated in prior North Central Cancer Treatment Group studies. This regimen offers no improvement in therapeutic activity for advanced gastric cancer. This study design, however, allows rapid screening of phase II regimens in patients who would usually be candidates for phase III trials. PMID- 10521069 TI - Phase II study of 2-chlorodeoxyadenosine in combination with chlorambucil in previously untreated B-cell chronic lymphocytic leukemia. AB - The objective was to determine the safety and efficacy of adding a maximally tolerated dose of 2-chlorodeoxyadenosine (2-CdA) to standard chlorambucil (CLB) therapy in previously untreated B-cell chronic lymphocytic leukemia (CLL). Thirty patients with CLL (median age, 64 years) received two courses of 2-CdA given intravenously (2 mg/m2 daily for 7 days) added to biweekly administration of CLB at 30 mg/m2 given orally. The diagnosis of CLL, treatment indications, and response criteria were according to the National Cancer Institute established guidelines. Sixteen patients (53%) had advanced-stage disease, and four (13%) had trisomy 12 abnormality. The overall remission rate was 80%, including 20% complete remission (CR), 30% nodular partial remission (nPR), and 30% partial remission (PR). Minimal residual disease was detected phenotypically in two of five patients with CR and in eight of nine with nPR. Overall, CR, nPR, and PR rates were not influenced significantly by the presence of cytogenetic abnormalities or advanced clinical stage. With a median follow-up of 33 months, 58% of patients who had a response had relapse. Median time to progression in all 30 patients was 30 months, and time to progression and progression-free survival were not significantly different for the different response groups, clinical stages, or cytogenetic groups. Severe neutropenia and thrombocytopenia occurred in 33% and 7% of patients, respectively. Only two patients had documented bacterial infections, and four had herpetic infections. Concurrent combination chemotherapy with abbreviated doses of 2-CdA and standard-dose CLB is feasible and safe in previously untreated CLL. Antitumor activity may be superior to that of CLB alone given in conventional doses. Whether a different schedule of combining these two agents would result in improved outcome is being investigated. PMID- 10521070 TI - Paclitaxel and G-CSF in previously untreated patients with extensive stage small cell lung cancer: a phase II study of the North Central Cancer Treatment Group. AB - Paclitaxel is an antimicrotubule agent that interferes with cell division. It has demonstrated promising single-agent activity against non-small-cell lung cancer. The purpose of this study was to evaluate the therapeutic effectiveness of paclitaxel in previously untreated patients with extensive stage small-cell lung cancer (SCLC). The study was designed as a two-stage phase II trial. All patients who entered received paclitaxel by intravenous infusion at a dose of 250 mg/m2 during 24 hours. Granulocyte colony stimulating factor was also provided to ameliorate neutropenia. Cycles were repeated at 21-day intervals. Patients who achieved a complete response received a maximum of 10 cycles of treatment, whereas those who achieved a partial response/regression continued treatment until progression or undue toxicity developed. Patients who progressed or maintained stable disease for six cycles were crossed over to cisplatin and etoposide. Forty-three patients entered the study and all were evaluable for analysis. Responses were observed in 23 (53%) of the patients. There was no significant difference in the response rates in patients with measurable or evaluable disease (13/23 versus 10/20, p = 0.76). At the time of analysis, 39 patients had progressed with a median time to progression of 95 days, and 39 patients had died with a median survival of 278 days. The 1-year achieved survival rate was 24%. Significant neutropenia (absolute neutrophil count <1,000/microl) occurred in 24 (56%) of the patients, but only 2 patients experienced severe infection (grade > or = 3), and there were no septic deaths. The results indicate that paclitaxel is active against SCLC. Myelosuppression was the main side effect in this patient population. Response duration was short (median = 3.4 months), which suggests that paclitaxel is not sufficient as a single agent. Further studies of paclitaxel in combination with other agents against SCLC are currently in progress within the North Central Cancer Treatment Group and other cancer treatment groups. Key Words: Paclitaxel-G-CSF-Small-cell lung cancer-North Central Cancer Treatment Group. PMID- 10521071 TI - Large proportion of Epstein-Barr virus-associated small noncleaved cell lymphomas among children with non-Hodgkin's lymphoma at a single institution in Moscow, Russia. AB - To evaluate clinical and biologic features of children with non-Hodgkin's lymphoma (NHL) treated in Moscow, Russia, we studied 53 cases treated in the Republican Children's Hospital from 1990 to 1994. Histologic examination disclosed a predominance of the small noncleaved cell subtype (32 cases, 60%); a smaller percentage of the lymphoblastic and large-cell subtypes (14 cases, 26% and 7 cases, 13%, respectively) were identified. The frequencies of primary sites of involvement in descending order included 60% in abdomen, 19% in mediastinum, 15% in head/neck, and 4% in peripheral nodes. The majority of children presented with advanced stage disease (St. Jude stage III/IV/B-ALL, 92.5%). Of interest, 8 of 15 (53%) small noncleaved cell NHL cases examined contained Epstein-Barr virus (EBV). The high frequency of EBV association in this study suggests that this virus may play a more global role in NHL pathogenesis than previously recognized. PMID- 10521072 TI - Radiation response of Rosai-Dorfman disease presenting with involvement of the orbits. AB - The purpose of this study was to review an interesting case of recurrent orbital sinus histiocytosis. The patient initially failed surgery, chemotherapy, and steroid therapy, only to have a durable response to low-dose radiation therapy of the orbits, lasting 6 and 11 years, respectively. Because there are few documented responses to radiotherapy, we present a case report in conjunction with the clinical, radiographic, and histopathologic information as well as a literature review of similar cases. PMID- 10521073 TI - Tamoxifen-induced thrombocytopenia. AB - Thrombocytopenia (platelet count < 100,000/microl) is a rare side effect of tamoxifen (Nolvadex). We report a case of thrombocytopenia that developed 6 months after tamoxifen was given as an adjuvant treatment for breast cancer. The patient received no concurrent cytotoxic chemotherapy or radiation therapy. The thrombocytopenia resolved after the tamoxifen was stopped, reappeared promptly when the tamoxifen was restarted, and resolved again after the second withdrawal, at which time anastrozole (Arimidex) was substituted for the tamoxifen. The patient's platelet count remained normal for more than 6 months thereafter. PMID- 10521074 TI - Paraneoplastic hyperpigmentation secondary to lung cancer resolving after chemotherapy. PMID- 10521075 TI - Is TD50 correlated with organ weight? PMID- 10521077 TI - Thrombosis in sickle cell disease. PMID- 10521076 TI - Efficacy of docetaxel in the second-line treatment of locally advanced or metastatic soft tissue sarcoma after failure of chemotherapy with anthracycline or ifosfamide. PMID- 10521078 TI - Evaluation of wound repair mechanisms with an in vitro cell culture model. PMID- 10521079 TI - Role of DNA methylation in the regulation of cell function. AB - The methylation of DNA helps stabilize chromatin in an inactive configuration and inhibits gene transcription. This mechanism of gene regulation is involved in essential genetic events including differentiation, genomic imprinting, and X chromosome inactivation. The alteration of methylation patterns can result in abnormal gene expression, with significant pathologic effects including carcinogenesis, autoimmunity, and some of the changes in gene expression associated with aging. The mechanisms establishing, maintaining, and modifying methylation patterns in normal and pathologic states are only now becoming understood, as are the mechanisms relating DNA methylation to gene expression and chromosome inactivation. Further characterization of these mechanisms holds promise for delaying or preventing the changes in methylation patterns that contribute to cancer, autoimmunity, and aging. PMID- 10521080 TI - Extracellular proteolysis: new paradigms for an old paradox. PMID- 10521081 TI - Antiphospholipid antibodies, proteins C and S, and coagulation changes in sickle cell disease. AB - The significance, interactions, and sources of coagulation abnormalities and their relationship to clinical severity and painful episodes in sickle cell disease are not clear. To evaluate this, we have examined various measures of coagulation in 37 patients with sickle cell disease (20 patients with HbSS disease and 17 patients with HbSC disease). Measurements have included isotypes of antiphospholipid antibodies (IgG, IgM, IgA) to specific phospholipids; proteins C (activity, total antigen) and S (activity, total and free antigen); measures of coagulation activation (prothrombin fragment 1.2, thrombin antithrombin, fibrinopeptide A, d-dimers); indicators of clinical severity; and studies obtained during steady states and painful episodes. Results in HbSS disease showed that antiphospholipid antibodies were increased, with IgG phosphatidylserine showing the highest and most frequently increased levels (37% of patients). Protein C (activity) and protein S (activity, total, free antigen) were decreased (P<.01), and all measures of coagulation activation were increased (P<.001). In HbSC disease, antiphospholipid antibodies were normal, protein C (activity) and protein S (free antigen) were decreased (P<.001), and all measures of coagulation activation were increased (P<.02). A strong correlation was observed in HbSS disease between IgG-PS and d-dimers. Moderate correlations occurred between protein C activity and thrombin-antithrombin and fibrinopeptide A, between protein S activity and prothrombin fragment 1.2 and d-dimers, and between protein C and protein S activity. In HbSC disease, moderate and fewer correlations occurred. Significant differences between HbSS disease and HbSC disease were observed in aPLs, proteins C and S, and measures of coagulation activation. Measurements during steady states and during painful episodes were not significantly different. We conclude that the antiphospholipid antibody IgG PS may contribute to coagulation activation in HbSS disease and that IgG-PS, protein C, and protein S relate to each other and jointly to measures of coagulation activation. The increased level of IgG-PS in HbSS disease most likely reflects exposure of the procoagulant phosphatidylserine on the surfaces of red cell-shed vesicles and sickle red cells, which would further affect coagulation activation. The significant differences in coagulation measures between HbSS disease and HbSC disease are consistent with differences in clinical severity between the diseases. The development of painful episodes does not appear to be related to the coagulation changes. PMID- 10521082 TI - Basic fibroblast growth factor stimulates repair of wounded hepatocyte monolayer: modulatory role of protein kinase A and extracellular matrix. AB - The two important repair mechanisms after hepatocyte injury are proliferation and migration of the nearby healthy hepatocytes. Although previous studies have shown that basic fibroblast growth factor (bFGF) levels are markedly elevated after liver injury, the role of bFGF in the repair of the wounded hepatocytes is not well understood. The aim of this study was to delineate the role of bFGF in the repair of the wounded hepatocyte monolayers. Specifically, we examined the role of bFGF in cellular proliferation and migration of hepatocytes with an in vitro wound model. Standardized excisional wounds were created in clone 9 rat hepatocyte monolayers by a razor blade, and the extent of epithelial proliferation and migration was measured. After wound formation, bFGF (30 ng/mL) significantly stimulated proliferation of hepatocytes at the wound margin. bFGF also stimulated the migration of hepatocytes at the wound front. bFGF stimulation of hepatocyte migration correlated with increased formation of actin stress fibers and bFGF-receptor protein level. The bFGF stimulation of hepatocyte migration was abolished by various protein kinase A activating agents including 3 isobutyl-1-methylxanthine, 8-bromoadenosine-3', 5'-cyclic monophosphate, forskolin, and cholera toxin. In addition, protein kinase A activating agents almost completely prevented bFGF-induced actin stress fiber formation in the cells at the wound front. Varying the basement membrane composition of the extracellular matrix had a selective enhancing effect on the basal rates of hepatocyte migration (collagen IV > or = laminin > collagen I > fibronectin > control (plastic)). bFGF treatment resulted in a similar additive increase in hepatocyte migration across all coated surfaces studied. We conclude that bFGF promotes hepatocyte wound repair by stimulating both proliferation and migration of the hepatocyte at the margin of the wound. PMID- 10521083 TI - Monochloramine-induced cytolysis to cultured rat gastric mucosal cells: role of glutathione and iron in protection and injury. AB - Helicobacter pylori (H. pylori) infection plays a role in the pathogenesis of peptic ulceration as well as active chronic gastritis. Possible mechanisms of H. pylori-induced mucosal injury include the generation of toxic monochloramine (NH2Cl) from oxidant (HOCl)--which is a product of activated neutrophils-and ammonia (NH3), which is a metabolite of H. pylori urease. To clarify mechanisms by which NH2Cl induces cytolysis, we determined the effects of glutathione (GSH) alteration and iron chelation on NH2Cl-induced damage in cultured rat gastric mucosal cells, because these are involved in oxidant injury. Cytotoxicity was quantified by chromium 51 release from prelabeled cells that were exposed to NH2Cl or hydrogen peroxide (H2O2). Although both NH2Cl and H2O2 caused a time related and dose-dependent increase in 51Cr release, NH2Cl was more cytotoxic than H2O2. Pretreatment with extracellular GSH caused a right shift of the dose response curve for NH2Cl, whereas pretreatment with diethyl maleate (a depletor of cellular GSH) rendered cells less resistant to NH2Cl. Iron chelation with 1,10 phenanthroline or deferoxamine failed to influence NH2Cl injury, whereas such treatment was protective against H2O2. Intracellular GSH seems to play an important role as a potent defense system against NH2Cl, as observed in H2O2 induced damage. However, the mechanisms of NH2Cl-induced damage seem to be distinctly different from cytolysis by H2O2 in terms of the mediation of cellular iron. PMID- 10521085 TI - Endothelin-1 urinary excretion, but not endothelin-1 plasma concentration, is increased in renovascular hypertension. AB - Animal experiments have shown an increase in prepro-endothelin-1 (prepro-ET-1) mRNA expression in the clipped kidney but none in the aortic and mesenteric arteries in 2-kidney, 1-clip Goldblatt hypertensive rats. The present study was aimed at investigating whether plasma and renal endothelin-1 (ET-1) systems are differently activated in patients with renovascular hypertension (RH). The plasma concentration and urinary excretion of ET-1 were measured in 5 patients with RH (before and after successful renal angioplasty), in 7 patients with essential hypertension (EH), and in 8 normotensive control subjects. Immediately before renal angioplasty, plasma samples for ET-1 and plasma renin activity (PRA) measurements were withdrawn from the aorta and both renal veins. Unlike the PRA, the plasma ET-1 concentration did not significantly differ between the involved and the uninvolved sides. The urinary ET-1 excretion level (Fig 1) was markedly increased in patients with RH (30+/-4 ng/g urinary creatinine (UC) vs. 2.5+/-0.2 ng/g UC and 2.6+/-0.5 ng/g UC in control subjects and patients with EH, respectively; P<.001), whereas the plasma ET-1 concentration was normal (0.8+/ 0.2 pg/mL vs. 0.65+/-0.3 pg/mL and 0.8+/-0.2 pg/mL in control subjects and EH, respectively, not significant). Renal angioplasty was followed in all patients by normalization of blood pressure and PRA. One week after angioplasty, urinary ET-1 decreased to one fourth of baseline (8.04+/-5.23 ng/g UC, P<.001 vs. values before angioplasty and P<.04 vs. control subjects) and normalized 1 month thereafter (3.13+/-1.62 ng/g UC, not significant vs. control subjects), whereas plasma ET-1 remained steady. The present findings clearly indicate that in patients with RH, urinary ET-1 excretion is increased, whereas plasma ET-1 concentration remains normal. Successful percutaneous transluminal renal angioplasty induced a notable reduction in ET-1 urinary excretion, whereas it did not affect ET-1 plasma concentration. PMID- 10521084 TI - Increased nuclear factor-kappaB activation in colitis of interleukin-2-deficient mice. AB - Recent studies support nuclear factor-kappaB (NF-kappaB) as a critical transcription factor in inflammatory bowel disease. We examined NF-kappaB and its inhibitors, IkappaB-alpha and IkappaB-beta, in the colitis of interleukin-2 deficient (IL-2-/-) mice at the ages of 5, 10, and 15 weeks and compared them with those of age-matched wild-type mice. Colon levels of nuclear NF-kappaB and mRNA for NF-kappaB responsive cytokines interleukin-1beta and tumor necrosis factor-alpha were markedly increased in interleukin-2-/-mice. Colon interleukin 1beta protein levels were significantly elevated, consistent with increased interleukin-1beta mRNA, whereas tumor necrosis factor-alpha protein levels were either lower than those of the control group or did not differ. Protein levels of the immunomodulatory cytokine interleukin-10 were diminished. The NF-kappaB responsive IkappaB-alpha was also increased, mirroring NF-kappaB activation. In contrast, IkappaB-beta levels did not differ from those of wild-type mice in the 5- and 10-week groups and were only mildly increased in the 15-week group. Serum amyloid A, an acute phase protein that also is NF-kappaB-responsive, was dramatically elevated in the serum of interleukin-2-/- mice and correlated with the severity of the colitis. These data support a role for NF-kappaB in the pathogenesis of intestinal inflammation in interleukin-2-/- mice. The measurement of NF-kappaB in colon tissue samples may provide a sensitive means of assessing the state of activation of the mucosal immune response, and serum amyloid A appears to be a reliable biochemical marker of disease activity. PMID- 10521086 TI - Circulating levels of thrombopoietic and inflammatory cytokines in patients with clonal and reactive thrombocytosis. AB - The regulation of megakaryocytopoiesis and thrombopoiesis appears to be under the control of an array of hematopoietic growth factors. To determine the relationship between endogenous cytokine levels and circulating platelet counts, we measured the serum levels of both thrombopoietic and inflammatory cytokines in the peripheral blood and bone marrow samples from 70 patients with clonal thrombocytosis (CT) caused by myeloproliferative disorders, 28 patients with reactive thrombocytosis (RT), and 35 normal control subjects. The levels of thrombopoietin (TPO), interleukin-6 (IL-6), soluble IL-6 (sIL-6) receptor, IL-11, stem cell factor (SCF), IL-3, and IL-8 were determined by enzyme-linked immunosorbent assay (ELISA). Platelet counts were significantly higher in both CT and patients with RT (699+/-399x10(9)/L, P<.001; 642+/-200 x 10(9)/L, P<.001; respectively) as compared with the normal control subjects (240+/-47x10(9)/L). The concentrations of cytokines in the bone marrow correlated well with those in the peripheral blood. The endogenous levels of TPO, IL-6, and sIL-6 receptor were significantly higher in both CT and patients with RT than those in normal control subjects. The median level of IL-6 was significantly higher in patients with RT than in patients with CT (40 pg/mL vs. 5 pg/mL; P<.001); however, there was no detectable difference in TPO and sIL-6 receptor levels between the two groups. Significantly higher levels of SCF and IL-8 were also found in patients with CT as compared with those found in normal control subjects (median 2460 pg/mL vs 1995 pg/mL, P<.05; 20 ng/mL vs. 5 ng/mL, P = .001; respectively). Finally, IL-11 and IL-3 levels were undetectable in most patients with thrombocytosis. Our results reveal that the endogenous levels of TPO, IL-6, sIL-6 receptor, IL-8, and SCF are elevated in patients with CT or RT. These cytokines appear to be active mediators involved in the regulation of thrombopoiesis during clonal and reactive thrombocytosis. PMID- 10521088 TI - Evaluation of interleukin-6 in bronchoalveolar lavage fluid and serum of patients with lung cancer. AB - Increased levels of serum interleukin 6 (IL-6) are found in patients with lung cancer, and it has been shown that this is part of a systemic inflammatory response syndrome. This study was designed to measure IL-6 levels in bronchoalveolar lavage (BAL) fluid of patients with lung cancer and to describe the relationship of BAL fluid IL-6 to the known systemic increase in IL-6. Increased levels of BAL fluid IL-6 can be found in patients with lung cancer as compared with patients with chronic obstructive pulmonary disease who have acute infection (P = .007). In patients with cancer, no correlation between BAL fluid IL-6 and serum IL-6 was found (P = .8). BAL fluid IL-6 did not correlate with the number of lymphocytes or macrophages found in this fluid. BAL fluid IL-6 does not correlate with tumor size. Although serum IL-6 was higher in patients with extensive stage small cell lung cancer as compared with levels in patients with limited stage disease (P = .06), their corresponding BAL fluid levels were not different (P = .9). Serum IL-6 correlated with other acute phase reactants. This study thus demonstrates the feasibility of utilizing BAL fluid analysis for local cytokine/tumor marker production in lung carcinoma. It also shows that a local increase in IL-6 in the BAL fluid is independent of the systemic inflammatory response syndrome, whereas the serum increase in IL-6 is part of this syndrome. PMID- 10521087 TI - Maintenance of GPIIb-IIIa avidity supporting "irreversible" fibrinogen binding is energy-dependent. AB - The interaction between fibrinogen and GPIIb-IIIa on stimulated platelets is multiphasic, progressing from reversible to irreversible ligand binding, associated with stabilization of platelet aggregates and clot retraction. Because fibrinogen binding to platelets has been linked to "outside-in" signaling events such as postreceptor occupancy protein tyrosine kinase and phosphatidylinositol-3 kinase activation, this study examined intracellular signaling requirements involved in stabilizing 125I-fibrinogen binding to adenosine diphosphate-treated platelets with selective inhibitors of protein tyrosine kinase (herbimycin A) (10 micromol/L) and phosphatidylinositol-3 kinase (Wortmannin) (10 nmol/L) and metabolic inhibitors antimycin A (7.3 micromol/L) and 2 deoxyglucose (6 mmol/L). Preincubation of platelets with herbimycin A or Wortmannin inhibited fibrinogen binding by 80% to 92% and was accompanied by markedly decreased tyrosine phosphorylation of a range of proteins migrating between 60 kDa and 125 kDa. The addition of inhibitors 5 minutes after adenosine diphosphate-induced fibrinogen binding also resulted in decreased tyrosine phosphorylation and dissociation of approximately 50% of bound fibrinogen within 60 minutes but failed to cause dissociation of irreversibly bound fibrinogen. In contrast, platelet exposure to metabolic inhibitors 5 minutes or 60 minutes after fibrinogen binding resulted in complete, spontaneous fibrinogen dissociation. These data suggest that the maintenance of GPIIb-IIIa avidity supporting irreversible fibrinogen binding to intact platelets is not affected by inhibitors of protein tyrosine kinase or phosphatidylinositol-3 kinase but involves other energy-dependent pathways. PMID- 10521089 TI - Therapeutic effect of glucocorticoid on experimental crescentic glomerulonephritis. AB - Crescentic glomerulonephritis shows active and progressive glomerular changes with rapid deterioration in kidney function. A large dose of glucocorticoid (pulse therapy) is clinically used for the treatment, but its efficacy has not been fully estimated. In this study we assessed the therapeutic effect of a large dose of methyl-prednisolone (MP) on a rat model of crescentic glomerulonephritis that had been induced in WKY rats by an injection of anti-glomerular basement membrane antibody. The infiltration of CD8+ cells and monocytes was manifest by day 3, proteinuria appeared on days 4 and 5, and cellular crescents were diffusely formed by day 7. The gene expression of MCP-1, a chemokine for monocytes and T lymphocytes, was enhanced within 4 hours and peaked on day 3. Daily administration of MP (30 mg/kg/d) from day 3 through day 6 reduced the gene expression of MCP-1 and the numbers of glomerular leukocytes and largely prevented both crescent formation and proteinuria. When daily MP treatment started on day 7, the numbers of glomerular CD8+ cells and monocytes, crescents, and urinary protein were significantly reduced by day 11. In addition, continuing treatment with a small dose of MP (3 mg/kg/d) begun on day 11 completely prevented the increase in blood urea nitrogen and serum creatinine levels. These results indicate that treatment with a large dose of MP histologically and clinically ameliorates crescentic glomerulonephritis in a rat model, supporting the efficacy of pulse MP therapy for the treatment of the disease in human subjects. PMID- 10521090 TI - Aerosol delivery of diethylenetriamine/nitric oxide, a nitric oxide adduct, causes selective pulmonary vasodilation in perinatal lambs. AB - Postnatal adaptation of the pulmonary circulation is mediated partly by endothelium-derived nitric oxide (NO). Recent studies have demonstrated that inhaled NO causes selective and sustained vasodilation in infants with persistent pulmonary hypertension of the newborn. Because the short half-life of NO limits its clinical application, we hypothesized that aerosol delivery of an NO-adduct, diethylenetriamine (DETANO), can cause sustained and selective pulmonary vasodilation. To test the acute effects of DETANO, we studied the pulmonary vascular response of late-gestation fetal lambs (n = 8; age = 138 days; term = 147) to aerosolized DETANO in the presence of an endothelium-derived NO inhibitor, nitro-L-arginine. To determine whether DETANO has a sustained effect, fetal lambs were ventilated with FiO2 0.10 before and 15 minutes after they were treated with aerosolized DETANO. Fetal lambs were acutely prepared. Nitro-L arginine (1 mg/min x 30 minutes) was infused into the left pulmonary artery before ventilation with FiO2 1.00 for 30 minutes, followed by continued ventilation with FiO2 0.10 for 10 minutes. This represented the control period. Ventilation was continued with FiO2 1.00, and aerosolized DETANO was given in doses of 0.1, 0.4, and 1.0 mg. Fifteen minutes after the last dose of DETANO was administered, animals were ventilated with FiO2 0.10. In the control period, during ventilation with FiO2 0.10, left pulmonary artery flow was 122+/-33 mL/min and decreased to 104+/-22 mL/min. Aerosol delivery of DETANO increased left pulmonary artery flow to 176+/-26 mL/min (P<.05) and had no effect on aortic pressure or heart rate. After DETANO was administered, ventilation with FiO2 0.10 did not cause any change in left pulmonary artery flow. We conclude that DETANO can cause selective fetal pulmonary vasodilation. Aerosol delivery of DETANO may increase the clinical applications of NO. PMID- 10521091 TI - The role of nutrition and nutritional supplements in women's health. AB - OBJECTIVE: To review the current literature on nutrition to provide a basis for counseling patients. DESIGN: Literature review. RESULT(S): Studies on nutrition and nutritional supplements in women's health are found primarily in literature not typically read by reproductive endocrinologists and gynecologist/obstetricians. A surprising number of people do not receive the vitamins and minerals that they need. Soy and soy isoflavones should be considered an important part of the diet. CONCLUSION(S): A better understanding of nutrition and nutritional supplements may reduce or prevent illness, saving the health care system millions of dollars each year. PMID- 10521092 TI - Assisted reproduction for couples infected with the human immunodeficiency virus type 1. PMID- 10521093 TI - Sex selection and preimplantation genetic diagnosis. The Ethics Committee of the American Society of Reproductive Medicine. PMID- 10521094 TI - Elective transfer of embryos selected on the basis of first polar body morphology is associated with increased rates of implantation and pregnancy. AB - OBJECTIVE: To determine the relationship between first polar body morphology and implantation rate and pregnancy rate (PR), to facilitate decision making concerning elective ET. DESIGN: Prospective, randomized study. SETTING: Fertility center. PATIENT(S): One hundred fifty-eight consecutive patients (173 intracytoplasmic sperm injection cycles) resulting in embryo transfers. INTERVENTION(S): In our study group, priority in ET was given to embryos derived from well-shaped first polar bodies, whereas selection of embryos for transfer in the control group was based exclusively on the degree of embryo fragmentation. MAIN OUTCOME MEASURE(S): Total numbers of implantations and pregnancies, PR and implantation rate, and rates of multiple pregnancy and miscarriage. RESULT(S): In the study cohort, 212 embryos were transferred. In the control group, 313 embryos were transferred. The implantation rate and PR were significantly lower in the control group than in the study cohort. In addition, the rate of multiple pregnancy was significantly higher in the study group. CONCLUSION(S): Elective transfer of embryos selected on the basis of first polar body morphology results in higher implantation and pregnancy rates. Multiple pregnancy can be avoided by transferring a reduced number of embryos selected on the basis of first polar body morphology. PMID- 10521095 TI - Blastocyst culture and transfer: analysis of results and parameters affecting outcome in two in vitro fertilization programs. AB - OBJECTIVE: To determine whether previously described advanced blastocyst development and high implantation rates are confirmed in an expanded multicenter trial. DESIGN: Retrospective review. SETTING: Two private assisted reproductive technology units. PATIENT(S): One hundred seventy-four patients who underwent blastocyst culture and transfer. INTERVENTION(S): Culture of all pronucleate embryos in sequential media to the blastocyst stage (day 5) followed by ET. MAIN OUTCOME MEASURE(S): The number and percentage of blastocysts developed, implantation rates, pregnancy rates, and parameters that affected outcome were analyzed. RESULT(S): Only 3 of 174 patients failed to achieve blastocyst-stage ET. The mean blastocyst development rate was 48%. The ongoing pregnancy rate was 66.3% per oocyte retrieval, with a mean (+/-SE) of 2.2 +/- 0.05 blastocysts transferred and an implantation rate of 48% per blastocyst transferred. CONCLUSION(S): Blastocyst culture and transfer is an effective means of treating patients who respond well to gonadotropins. High pregnancy rates can be accomplished with low numbers of embryos transferred. Patients who failed to achieve ET were rare. PMID- 10521096 TI - How useful is cervical dilatation in patients with cervical stenosis who are participating in an in vitro fertilization-embryo transfer program? The Bourn Hall experience. AB - OBJECTIVE: To evaluate the place of cervical dilatation performed at the initial visit in an IVF-ET cycle in patients with known cervical stenosis. DESIGN: Retrospective study. SETTING: A tertiary care assisted conception unit. PATIENT(S): Fifty-seven patients who failed to conceive after a previous ET attempt and in whom the ET was classified as "difficult." INTERVENTION(S): Cervical dilatation under general anesthesia after pituitary suppression and before gonadotropin stimulation. MAIN OUTCOME MEASURE(S): Ease of the ET procedure and clinical pregnancy rate. RESULT(S): Eighteen (31.6%) of 57 women who failed to conceive after a previous attempt at IVF-ET achieved a clinical pregnancy after cervical dilatation. In 40 patients (70.2%), the subsequent ET was classified as "easy," whereas in the other 17 (29.8%), it remained difficult. The pregnancy rate was significantly higher when the ET was easy than when it was difficult (40% versus 11.8%, P<.05). CONCLUSION(S): In patients with cervical stenosis and a previous difficult ET, cervical dilatation during the initial visit leads to an easier subsequent ET and improves the pregnancy rate. PMID- 10521098 TI - Efficacy of double intrauterine insemination in controlled ovarian hyperstimulation cycles. AB - OBJECTIVE: To investigate the effectiveness of double IUI and to determine the optimal timing of IUI in relation to hCG administration. DESIGN: Prospective randomized study. SETTING: Infertility Center, Department of Obstetrics and Gynecology, University of Milan. PATIENT(S): Patients with male factor and unexplained infertility undergoing controlled ovarian hyperstimulation (COH) and IUI. INTERVENTION(S): After COH with clomiphene citrate and gonadotropins, patients were randomly assigned to one of the following groups: group A received a single IUI 34 hours after hCG administration, group B received a double IUI 12 hours and 34 hours after hCG administration, and group C received a double IUI 34 hours and 60 hours after hCG administration. MAIN OUTCOME MEASURE(S): Number of follicles > 15 mm in diameter on the day of hCG administration, number of motile spermatozoa inseminated, clinical pregnancy rate. RESULT(S): Two hundred seventy three patients underwent 449 treatment cycles: 90 patients were treated for 156 cycles in group A, 92 patients for 144 cycles in group B, and 91 patients for 149 cycles in group C. The overall pregnancies rates for groups A, B, and C were 13 (14.4% per patient and 8.3% per cycle), 28 (30.4% per patient and 19.4% per cycle), and 10 (10.9% per patient and 6.7% per cycle), respectively. There was a statistically significant difference between group B and groups A and C. CONCLUSION(S): Our data indicate that two IUIs performed 12 hours and 34 hours after hCG administration is the most cost-effective regimen for women undergoing COH cycles with clomiphene citrate and gonadotropins. Although the second insemination adds up to a slightly higher cost, it significantly increases the chance of pregnancy. PMID- 10521097 TI - Risk factors for multiple gestation in women undergoing intrauterine insemination with ovarian stimulation. AB - OBJECTIVE: To identify whether sperm characteristics after washing and/or ovulation induction cycle characteristics can predict the occurrence of multiple conception in patients undergoing ovarian stimulation and IUI. DESIGN: Retrospective study. SETTING: A gynecology clinic and an andrology laboratory at a tertiary care facility. PATIENT(S): One hundred patients with single pregnancies and 22 patients with multiple pregnancies. INTERVENTION(S): Patients underwent ovarian stimulation and IUI with their partner's sperm. MAIN OUTCOME MEASURE(S): Relation of patient characteristics, ovarian stimulation, and sperm characteristics after washing to the occurrence of multiple pregnancy. RESULT(S): The mean serum E2 level on the day of hCG injection was significantly higher in the multiple conception group, but the number of follicles was not. The total sperm count, total motile sperm count, and sperm motility after washing did not differ between the groups. However, couples with multiple pregnancies had sperm with a significantly higher amplitude of lateral head movement than couples with single pregnancies. A peak E2 level of >583 pg/mL on the day of hCG injection and sperm with an ALH of >4 microm after washing predicted the occurrence of multiple pregnancy. CONCLUSION(S): Sperm with an amplitude of lateral head movement of >4 microm and a peak E2 level of >583 pg/mL are significant risk factors for multiple pregnancy in patients undergoing IUI. PMID- 10521099 TI - Development of the fertility adjustment scale. AB - OBJECTIVE: To develop a standardized measure of psychological adjustment to infertility. DESIGN: A cross-sectional two-group comparison study. SETTING: Two specialized fertility clinics in large teaching hospitals. PATIENT(S): Fifty men and 50 women undergoing evaluation and/or treatment of fertility problems. INTERVENTION(S): The Fertility Adjustment Scale was administered with the Hospital Anxiety and Depression Scale as a measure of concurrent validity. MAIN OUTCOME MEASURE(S): Scores on the Fertility Adjustment Scale and the Hospital Anxiety and Depression Scale. RESULT(S): Scores on the Fertility Adjustment Scale were distributed normally. Split-half and internal consistency were high. A significant correlation with measures of mood, anxiety, and distress provided evidence of concurrent validity. CONCLUSION(S): Preliminary results suggest that this measure will be a useful tool in assessing psychological reactions to fertility problems and outcomes of treatment. PMID- 10521100 TI - The presence of the 21-hydroxylase deficiency carrier status in hirsute women: phenotype-genotype correlations. AB - OBJECTIVE: To determine the role of heterozygosity for mutations in the 21 hydroxylase gene (CYP21) in the pathogenesis of hyperandrogenism. DESIGN: Controlled clinical study. SETTING: Tertiary care institutional hospital. PATIENT(S): Forty hirsute women and 13 healthy control women. INTERVENTION(S): The source of androgen excess was determined by the changes in serum testosterone levels in response to a single 3.75-mg i.m. dose of triptorelin. MAIN OUTCOME MEASURE(S): CYP21 molecular genetic analysis and serum 17-hydroxyprogesterone levels. RESULT(S): Eight patients and one control were heterozygous carriers of CYP21 mutations. Two patients with adrenal hyperandrogenism and one patient with ovarian hyperandrogenism, who carried the V281L mutation had an increased ACTH stimulated 17-hydroxyprogesterone level (>4.1 ng/mL) that persisted during gonadal suppression. Another patient with adrenal hyperandrogenism carried the V281L mutation, and her ACTH-stimulated 17-hydroxyprogesterone level was elevated only during gonadal suppression. Four patients (three with idiopathic hirsutism, one with ovarian hyperandrogenism) and one control were carriers of CYP21 mutations typically associated with classic congenital adrenal hyperplasia but had normal basal and ACTH-stimulated 17-hydroxyprogesterone levels. Nine patients without CYP21 mutations had increased ACTH-stimulated 17-hydroxyprogesterone levels; these decreased to normal in six of the patients during gonadal suppression. CONCLUSION(S): The response of serum 17-hydroxyprogesterone to ACTH does not predict CYP21 carrier status. No clear concordance was found between the CYP21 genotype and the functional origin of androgen excess. PMID- 10521101 TI - Pregnancies resulting from in vitro matured oocytes retrieved from patients with polycystic ovary syndrome after priming with human chorionic gonadotropin. AB - OBJECTIVE: To describe pregnancies that resulted from in vitro matured oocytes derived from two unstimulated, anovulatory patients with polycystic ovary syndrome after priming with hCG before oocyte retrieval. DESIGN: Case series. SETTING: McGill Reproductive Center, Royal Victoria Hospital, McGill University. PATIENT(S): Two women with polycystic ovary syndrome. INTERVENTION(S): Progesterone induction of a withdrawal bleeding episode. The administration of subcutaneous hCG 36 hours before oocyte retrieval in a natural menstrual cycle. In vitro maturation of immature oocytes, fertilization, and ET. Luteal support with oral estrogen and progesterone. MAIN OUTCOME MEASURE(S): Pregnancy outcome. RESULT(S): Two clinical pregnancies were confirmed after oocyte maturation in vitro, fertilization with intracytoplasmic sperm injection, and ET. CONCLUSION(S): The administration of hCG 36 hours before harvesting of immature oocytes may improve the maturational and developmental competence of the oocytes and the pregnancy rates of unstimulated patients with polycystic ovary syndrome. PMID- 10521102 TI - Are gestational age and endometrial thickness alternatives for serum human chorionic gonadotropin as criteria for the diagnosis of ectopic pregnancy? AB - OBJECTIVE: To compare gestational age and endometrial stripe thickness measurement with serum hCG measurement as criteria for the diagnosis of ectopic pregnancy (EP). DESIGN: Prospective study. SETTING: Two large teaching hospitals in Amsterdam, The Netherlands. PATIENT(S): Three hundred fifty-four consecutively seen pregnant patients who presented between September 1993 and April 1996 with suspected EP and in whom transvaginal ultrasonogram showed no intrauterine pregnancy or EP. Ultrasonography was performed by one of the study investigators or, during shifts, by the resident on call. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): The accuracy of gestational age, endometrial stripe thickness, and serum hCG measurement in the diagnosis of EP was evaluated with receiver operating characteristic curve analysis. RESULT(S): Gestational age and endometrial stripe thickness could not discriminate between patients with EP and patients without EP, whereas serum hCG had an acceptable area under the receiver operating characteristic curve. CONCLUSION(S): Gestational age and endometrial thickness are not useful in the diagnosis of EP. Serum hCG measurement is the diagnostic instrument of choice in patients with suspected EP when transvaginal ultrasonography does not reveal a diagnosis. PMID- 10521103 TI - Screening for factor V Leiden mutation before prescribing combination oral contraceptives. AB - OBJECTIVE: To evaluate the cost-effectiveness of screening for factor V Leiden mutation in women in the United States who use combination oral contraceptives. DESIGN: Cost-effectiveness analysis. SETTING: A national research reference laboratory, a university medical center, and an academic health center managed care organization. PATIENT(S): Women of reproductive age in the United States. INTERVENTION(S): Baseline risk estimates of venous thromboembolic disease in the general population and in carriers of factor V Leiden mutation were calculated using available data. MAIN OUTCOME MEASURE(S): The number of women who would require factor V Leiden testing and the cost of identifying this cohort to prevent one death caused by venous thromboembolic disease before prescribing combination oral contraceptives. RESULT(S): To prevent one venous thromboembolic death attributable to the use of oral contraceptives in women with factor V Leiden mutation, >92,000 carriers would need to be identified and stopped from using these pills. The estimated charge to prevent this one death would exceed $300 million. If the price of testing were discounted to 34.5% of current charges, the cost still would be between $105 million and $130 million. CONCLUSION(S): Screening for factor V Leiden mutation before prescribing combination oral contraceptives is not a cost-effective use of U.S. health care dollars. The best and most cost-effective screening tool we have is taking a thorough personal and family history related to venous thromboembolic events. PMID- 10521104 TI - Oral contraceptives that contain ethinyl estradiol (0.035 mg) and cyproterone acetate (2 mg) inhibit leukocyte transmigration through endothelial cell monolayers. AB - OBJECTIVE: To investigate the influence of ethinyl estradiol and cyproterone acetate in oral contraceptives on leukocyte migration through endothelial cell monolayers. DESIGN: Experimental in vitro prospective study. SETTING: An academic research laboratory. INTERVENTION(S): Endothelial cells were cultured on microporous membranes to produce monolayers. Polymorphonuclear leukocytes were used in a previously described migration assay (n = 7). The amount of untreated polymorphonuclear leukocytes that migrated through untreated endothelial cell monolayers was used as a control and set at 100%. In addition, a leukocyte adhesion assay was used. MAIN OUTCOME MEASURE(S): Leukocyte adhesion to and transmigration through endothelial cell monolayers. RESULT(S): Ethinyl estradiol and cyproterone acetate inhibited the migration of polymorphonuclear leukocytes through endothelial cell monolayers significantly (67% +/- 6.4%) when both cell types were treated to simulate in vivo conditions. The adhesion assay produced similar results. CONCLUSION(S): Ethinyl estradiol and cyproterone acetate were identified as potent inhibitors of leukocyte migration through endothelial cell monolayers. PMID- 10521105 TI - Leptin levels vary significantly during the menstrual cycle, pregnancy, and in vitro fertilization treatment: possible relation to estradiol. AB - OBJECTIVE: To evaluate the influence of sex steroids on leptin levels in patients with conditions in which the steroid levels are increased. DESIGN: Prospective study. SETTING: A hospital unit for reproductive medicine and a maternal care unit affiliated with the hospital and hospital staff. PATIENT(S): Thirteen women with regular menstrual cycles, 29 women with normal pregnancies, and 25 women undergoing IVF treatment. INTERVENTION(S): Blood samples were obtained during days 1-3, 6-8, 13-15, and 22-25 of the menstrual cycle in regularly cycling women and during gestational weeks 13, 20, 28, 32, and 36 and 7-13 weeks after birth in pregnant women. In women undergoing IVF treatment, blood samples were collected after E2 suppression, after ovarian stimulation, and at the time of ovum pickup. MAIN OUTCOME MEASURE(S): Serum levels of leptin, E2, and progesterone. RESULT(S): Leptin levels varied during the menstrual cycle and were elevated during pregnancy, with a peak during week 28. In the IVF group, leptin levels increased throughout the treatment cycle. Body mass index correlated positively with leptin levels in all three groups, and the maternal weight gain from weeks 13-32 tended to correlate with the rise in leptin levels. Estradiol levels correlated positively with leptin levels during E2 suppression. Negative correlations existed between the pregnancy-induced increases in E2 and leptin levels from weeks 13-32, and between the levels after birth. Leptin levels and progesterone levels did not correlate in any of the groups. CONCLUSION(S): Modest elevations of leptin levels were observed during IVF treatment and pregnancy. The increase in the IVF group indicates that factors other than body fat mass (possibly E2) also are of importance for the regulation of leptin levels. PMID- 10521106 TI - Nonfunctioning pituitary macroadenoma presenting with mild hyperprolactinemia and amenorrhea. AB - OBJECTIVE: To describe a patient with a clinically nonfunctioning pituitary macroadenoma who presented with mild hyperprolactinemia and amenorrhea. DESIGN: Case report. SETTING: Tertiary care medical facility. PATIENT(S): A 44-year-old woman with a 6-month history of amenorrhea. INTERVENTION(S): Pituitary testing, magnetic resonance imaging of the sella turcica, and transsphenoidal surgery. MAIN OUTCOME MEASURE(S): Pituitary function testing, magnetic resonance imaging, and return of menstrual cycles. RESULT(S): Baseline laboratory data revealed a serum prolactin level of 34 ng/mL (normal range, 3-20 ng/mL), normal thyroid function test results, and an FSH level of 6.7 mIU/mL. A second fasting prolactin level was 48 ng/mL. Magnetic resonance imaging of the sella turcica revealed a pituitary macroadenoma measuring 1.4 x 3.2 cm. Further testing of baseline pituitary function revealed normal findings. The patient underwent an uncomplicated transsphenoidal resection of the pituitary tumor and maintained normal pituitary function. Pathologic evaluation revealed a pituitary adenoma that stained positive for FSH and focally for the alpha subunit. The adenoma stained negative for GH, prolactin, ACTH, LH, and TSH. CONCLUSION(S): This patient had a nonsecreting gonadotroph macroadenoma that resulted in hypogonadotropic hypogonadism along with mild hyperprolactinemia, presumably secondary to interruption of normal transport down the pituitary stalk. PMID- 10521107 TI - Clinical application of intracytoplasmic sperm injection using in vitro cultured testicular spermatozoa obtained the day before egg retrieval. AB - OBJECTIVE: To study the effect of in vitro culture on the quality of human testicular sperm and the efficiency of intracytoplasmic sperm injection with in vitro cultured testicular sperm. DESIGN: Clinical study. SETTING: A private IVF center. PATIENT(S): Twenty consecutively seen IVF patients undergoing testicular biopsies for ICSI. INTERVENTION(S): The testicular specimens were cultured in vitro for 24 hours and the isolated spermatozoa were microinjected. MAIN OUTCOME MEASURE(S): Preincubation and postculture sperm motility, and fertilization, implantation, and pregnancy rates after intracytoplasmic sperm injection. RESULT(S): Motility increased from initial nonmotile or twitching sperm to free motile sperm in 18 of 20 cases. The injection of in vitro cultured testicular sperm resulted in a fertilization rate of 58%, an implantation rate of 20%, and a pregnancy rate of 45%. CONCLUSION(S): A testicular biopsy procedure can be performed the day before egg retrieval. Despite the low initial sperm quality, a high percentage of the prepared testicular sperm showed increased motility after 24 hours of culture. The injection of in vitro cultured testicular sperm into matured oocytes resulted in fertilization, implantation, and pregnancy rates comparable to those obtained with ejaculated sperm. PMID- 10521108 TI - The role of purified follicle stimulating hormone therapy in the male partner before intracytoplasmic sperm injection. AB - OBJECTIVE: To evaluate the impact of long-term purified FSH (pFSH) therapy in male partners before intracytoplasmic sperm injection (ICSI). DESIGN: Prospective, randomized, controlled study. SETTING: Large university-based IVF unit. PATIENT(S): Seventy-eight patients made up the study and control groups (39 patients each). All patients had severe male factor infertility. INTERVENTION(S): Induction of ovulation, oocyte retrieval, ICSI, and ET were carried out in both groups. In the study group, male partners received pFSH (75 IU FSH, <1 IU LH) for > or = 50 days before oocyte retrieval. MAIN OUTCOME MEASURE(S): Fertilization rate, embryo quality, implantation and pregnancy rates. RESULT(S): Fertilization and pregnancy rates were higher in the study group (68% and 35.9%, respectively) were higher than in the control group (59% and 17.9%, respectively), although the differences did not reach statistical significance. The implantation rate was significantly higher in the study group (15.5% versus 6.5%). The study group showed a trend toward a higher number of better-quality embryos per transfer (mean +/- SD, 2.2 +/- 1.6 versus 1.6 +/- 1.6). CONCLUSION(S): Purified FSH therapy in male partners before ICSI improves implantation rate. Improved embryo quality may be a contributory factor. PMID- 10521109 TI - Prognostic factors in patients continuing in vitro fertilization or intracytoplasmic sperm injection treatment and dropouts. AB - OBJECTIVE: To investigate whether cumulative pregnancy rates using life table analysis but without considering dropouts are representative of the whole population of patients entering an in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) program. DESIGN: Retrospective study. SETTING: University hospital-based infertility center. PATIENT(S): One thousand one hundred sixty-nine patients entering our IVF/ICSI program from January 1993 to December 1996. INTERVENTION(S): Comparison of prognostic factors between pregnant and nonpregnant patients, and between patients continuing IVF/ICSI treatment and dropouts. MAIN OUTCOME MEASURE(S): Prognostic factors, such as patient age, cancellation of oocyte retrieval because of poor response to ovarian stimulation, number of oocytes retrieved, fertilization rate, number and quality of embryos transferred. RESULT(S): No statistical differences in prognostic factors were found between patients continuing IVF/ICSI treatment and dropouts. CONCLUSION(S): Cumulative pregnancy rates using life table analysis can be considered representative of the whole population of patients for at least the first three treatment cycles. PMID- 10521110 TI - Embryo implantation in in vitro fertilization and intracytoplasmic sperm injection: impact of cleavage status, morphology grade, and number of embryos transferred. AB - OBJECTIVE: The aims of this study were to compare preimplantation embryo quality in intracytoplasmic sperm injection (ICSI) with standard IVF and to examine the impact of age and number and quality of embryos transferred on implantation and pregnancy. DESIGN: Retrospective, controlled clinical study. SETTING: Academic tertiary center. PATIENT(S): We examined 211 consecutive couples undergoing ICSI who were matched with 211 couples undergoing IVF therapy during the same time frame. INTERVENTION(S): In vitro embryo culture. MAIN OUTCOME MEASURE(S): Day 3 embryo quality as judged by the number of blastomeres and morphology scoring. RESULT(S): Patients undergoing ICSI had a significantly reduced number of embryos with good morphology and cleavage compared with IVF cases. Nevertheless, pregnancy and abortion rates were similar when adjusted by age and number of embryos transferred. Average cleavage status and age were significant predictors of implantation. Women of advanced age had significantly lower embryo cleavage and implantation rates. CONCLUSION(S): [1] The cleaving status of day 3 embryos is a valuable, although limited, indicator of implantation outcome. [2] In vitro fertilization-derived embryos had better cleavage rates and morphology scores than ICSI-derived embryos; however, the implantation potential was similar for both groups. [3] The age-related decline in implantation rate was associated with impaired embryo growth rates. PMID- 10521111 TI - The antiviral agents, MAP30 and GAP31, are not toxic to human spermatozoa and may be useful in preventing the sexual transmission of human immunodeficiency virus type 1. AB - OBJECTIVE: To investigate the effects of two virucidal compounds, MAP30 (Momordica anti-human immunodeficiency virus [HIV] protein; molecular weight, 30 kd) and GAP31 (Gelonium anti-HIV protein; molecular weight, 31 kd), obtained from Momordica charantia and Gelonium multiflorum, respectively, on the motility and vitality of human spermatocytes. DESIGN: Prospective, controlled study. SETTING: New York University School of Medicine. PATIENT(S): Ten healthy men undergoing evaluation for infertility provided 10 semen specimens. INTERVENTION(S): Human sperm were treated with the anti-HIV agents, MAP30 and GAP3 1. Nonoxynol-9, a commonly used spermicide, and phosphate-buffered saline were used as the positive and negative controls, respectively. MAIN OUTCOME MEASURE(S): The motility and vitality of human spermatocytes treated with MAP30 and GAP31 at doses that inhibit HIV-1 and herpes simplex virus. RESULT(S): MAP30 and GAP31 did not inhibit the motility or vitality of human sperm cells over a dose range of 100 0.1 microg/mL, whereas nonoxynol-9 demonstrated spermicidal action on all 10 samples over the same dose range. CONCLUSION(S): The antiviral agents, MAP30 and GAP31, were not toxic to human sperm cells at the doses at which they inhibit HIV 1 and herpes simplex virus. They had no effect on the motility of spermatozoa, even at a dose of 1,000 times the maximum effective concentration. These results indicate that MAP30 and GAP31 may be useful as nonspermicidal protection against sexually transmitted diseases. PMID- 10521112 TI - Identification of the human sperm protein that interacts with sperm-immobilizing antibodies in the sera of infertile women. AB - OBJECTIVE: To identify the target antigen of sperm-immobilizing antibodies present in the circulation of infertile women. DESIGN: Laboratory research. SETTING: Academic research laboratory. PATIENT(S): Twenty-nine infertile women with sperm-immobilizing antibodies, 22 infertile women with other disorders, and 20 fertile women. INTERVENTION(S): Titers of antibodies to the sperm protein, rSMP-B, were determined by ELISA using as substrate the synthetic peptide segment (rSMP-230) that corresponds with the hydrophilic domain of rSMP-B. Tests for sperm immobilization and zona pellucida penetration were performed using the human IVF system. MAIN OUTCOME MEASURE(S): Human sera with sperm-immobilizing activity were assayed for the presence of antibodies to rSMP-230. Polyclonal antibodies to rSMP-230 were assessed for the same biologic activities as sperm immobilizing antibodies. RESULT(S): Antibodies to rSMP-230 were detected in 10 (34%) of 29 sera obtained from women with immunologic infertility. In contrast, only one serum sample (2%) from women without sperm-immobilizing activity had a low titer of antibodies to rSMP-230. Polyclonal antibodies to rSMP-230 completely immobilized human sperm in the presence of complement and blocked sperm penetration across the zona pellucida. CONCLUSION(S): The human sperm protein, rSMP-B, probably is the target antigen of sperm-immobilizing antibodies. PMID- 10521113 TI - Screening for abnormalities of chromosomes X, Y, and 18 and for diploidy in spermatozoa from infertile men participating in an in vitro fertilization intracytoplasmic sperm injection program. AB - OBJECTIVE: To evaluate the frequency of disomy (for chromosomes X, Y, and 18) and of diploidy in the spermatozoa of infertile men undergoing intracytoplasmic sperm injection (ICSI). DESIGN: Prospective analysis of sperm nuclei by fluorescence in situ hybridization (FISH). SETTING: University-affiliated IVF-ICSI program. PATIENT(S): Semen samples from 19 patients participating in an IVF-ICSI program. INTERVENTION(S): Semen samples were analyzed and prepared for FISH. MAIN OUTCOME MEASURE(S): Semen parameters were evaluated. The frequency of disomy for chromosomes X, Y, and 18 and the frequency of diploidy were analyzed by FISH. RESULT(S): A total of 9,373 spermatozoa from 19 infertile patients were analyzed and compared with spermatozoa from a control group of 5 healthy men. No differences in the frequency of disomy 18 were found, but statistically significant differences in the incidence of sex chromosome disomy and of diploidy were observed. CONCLUSION(S): The study of sperm nuclei by FISH is useful to improve genetic counseling in infertile patients selected for ICSI. PMID- 10521114 TI - Birth after the injection of sperm and the cytoplasm of tripronucleate zygotes into metaphase II oocytes in patients with repeated implantation failure after assisted fertilization procedures. AB - OBJECTIVE: To assess the technique of injecting a single sperm and cytoplasm obtained from tripronucleate zygotes into metaphase II oocytes for the treatment of patients with repeated implantation failure after intracytoplasmic sperm injection or IVF. DESIGN: Clinical study. SETTING: Private infertility clinic. PATIENT(S): Patients with repeated implantation failure after intracytoplasmic sperm injection or IVF. INTERVENTION(S): The metaphase II oocytes of recipients were injected with their husbands' spermatozoa and cytoplasm aspirated from the tripronucleate zygotes of donors. MAIN OUTCOME MEASURE(S): Fertilization after cytoplasm and sperm injection, embryo development, and successful pregnancy. RESULT(S): In total, 62 metaphase II oocytes from nine recipients were injected. Of the 62 injected oocytes, 3 (5%) degenerated and 43 (69%) had two pronuclei 18 hours after injection. Thirty-nine oocytes with two pronuclei cleaved to the two cell to six-cell stage after another 24 hours of culture. All cleaved embryos were transferred into the uteruses of recipients. Four clinical pregnancies occurred in four recipients. No abnormal chromosomes were observed after amniocentesis and karyotyping in all pregnancies. Five healthy infants were born. CONCLUSION(S): Injection of the cytoplasm of tripronucleate zygotes may enhance the clinical pregnancy rate in patients with repeated implantation failure after intracytoplasmic sperm injection or IVF. PMID- 10521115 TI - Perifollicular blood flow Doppler indices, but not follicular pO2, pCO2, or pH, predict oocyte developmental competence in in vitro fertilization. AB - OBJECTIVE: To assess the relationships among perifollicular blood flow; follicular fluid pO2, pCO2, and pH; oocyte developmental capacity; preimplantation embryo quality. DESIGN: Prospective study. SETTING: Academic, tertiary care institution. PATIENT(S): Unselected, gonadotropin-stimulated IVF cycles. INTERVENTION(S): Color, pulsed Doppler analysis of perifollicular blood flow, and follicular pO2, pCO2, and pH determinations of randomly designated, mapped ovarian follicles. MAIN OUTCOME MEASURE(S): Fertilization and day 3 embryo cleavage and morphology. RESULT(S): Perifollicular vascularity indices were significantly and negatively correlated with day 3 embryo cleavage. Pulsatility index and S-D ratio also were significantly and negatively correlated with follicular pO2. The same correlation was found between resistance index and the fertilization rate of preovulatory oocytes. No relationship existed between follicular metabolic analysis and fertilization or embryo quality. The resistance index had a sensitivity of 0.57 and a specificity of 0.71 for the prediction of advanced embryo cleavage status. CONCLUSION(S): Results confirm and extend previous reports demonstrating that color, pulsed Doppler ultrasound analysis of individual preovulatory follicles during IVF therapy may provide an indirect index of the developmental competence of the corresponding oocyte. Although these methods may provide means to select embryos for transfer with the highest implantation potential, the moderate predictive power showed so far may limit their clinical applicability. PMID- 10521116 TI - Expression of leukemia inhibitory factor and its receptor in preimplantation embryos. AB - OBJECTIVE: To examine the expression of leukemia inhibitory factor (LIF) and its receptor (LIF-R) transcripts in human and murine preimplantation embryos. DESIGN: Prospective study. SETTING: University medical center. PATIENT(S): Human oocytes were obtained from patients undergoing IVF treatment. Two-cell murine embryos were obtained from ICR strain mice. INTERVENTION(S): Second-day intracytoplasmic sperm injection procedures were performed on oocytes that failed to be fertilized by IVF. Embryos were cultured to various stages and collected for study. MAIN OUTCOME MEASURE(S): The transcript levels of LIF and LIF-R in these embryos were examined and semiquantitated using single-cell reverse transcription-polymerase chain reaction methodology. RESULT(S): Leukemia inhibitory factor and LIF-R transcripts were detectable in most human preimplantation embryos (30 of 34 and 31 of 34 embryos showed LIF and LIF-R messenger RNA, respectively). There was a trend toward decreased expression of both transcripts in embryos at the four-cell stage and in embryos in which growth had been arrested for 24-48 hours. The expression of LIF and LIF-R genes in murine embryos was inconsistent. CONCLUSION(S): Preimplantation human embryos express LIF and LIF-R messenger RNA. It is suggested that LIF may be able to affect embryo development through its action at stages before implantation in an autocrine or paracrine manner. PMID- 10521117 TI - Superoxide dismutase in normal cycling human ovaries: immunohistochemical localization and characterization. AB - OBJECTIVE: To investigate the expression of manganese (Mn) and copper-zinc (Cu,Zn) superoxide dismutase (SOD) in normal cycling human ovaries throughout the menstrual cycle. DESIGN: Descriptive, controlled study. SETTING: Tohoku University School of Medicine. PATIENT(S): Twenty-four normal cycling human ovaries were obtained from patients who underwent oophorectomy and hysterectomy for squamous cell carcinoma of the uterine cervix. INTERVENTION(S): Immunohistochemistry for Mn-SOD and Cu,Zn-SOD. MAIN OUTCOME MEASURE(S): Immunostaining. RESULT(S): In the follicular stage, Mn-SOD immunoreactivity was detected in granulosa and theca interna cells of steroid-producing follicles, that is, preantral, nondominant, dominant, and atretic follicles, whereas Cu,Zn SOD was detected in theca interna cells of these follicles and in granulosa cells of dominant follicles. In the luteal stage, immunoreactivity for Mn-SOD and Cu,Zn SOD was observed in both luteinized granulosa and theca cells of the functioning corpus luteum. In the early degenerating corpus luteum, both Mn-SOD and Cu,Zn-SOD were positive in steroid-producing luteinized theca cells. Mn-SOD immunoreactivity was also detected in nonsteroid-producing luteinized granulosa cells and macrophages. CONCLUSION(S): Our results suggest that the expression of Mn-SOD and Cu,Zn-SOD closely correlates with steroidogenesis in the human ovary. In addition, Mn-SOD may play an important role in the process of luteal regression. PMID- 10521118 TI - Phantom endometriosis of the sciatic nerve. AB - OBJECTIVE: To assess the efficacy and diagnostic value of GnRH agonist (GnRH-a) therapy in cases of hidden sciatic nerve endometriosis. DESIGN: Case report. SETTING: Academic tertiary referral center for endometriosis treatment. PATIENT(S): Three patients with cyclic, catamenial sciatica associated with pelvic endometriosis who had electromyographic evidence of sciatic nerve damage but negative computed tomography and magnetic resonance imaging findings. INTERVENTION(S): Monthly administration of the GnRH-a leuprolide acetate plus daily transdermal E2 (25 microg). MAIN OUTCOME MEASURE(S): Relief of pain symptoms and improvement in motor function. RESULT(S): All three patients had clear decreases in pain and partial amelioration of claudication. CONCLUSION(S): Endometriosis of the sciatic nerve may be hard to diagnose with the use of current imaging techniques but may be proved by clinical response to GnRH analogue treatment and may be more frequent than previously thought. PMID- 10521119 TI - Bacterial vaginosis and past chlamydial infection are strongly and independently associated with tubal infertility but do not affect in vitro fertilization success rates. AB - OBJECTIVE: To determine the incidence of active vaginal infection in women undergoing IVF, relate it to the cause of infertility, and investigate a relation with the outcome of fresh ET. DESIGN: Cross-sectional study. SETTING: Tertiary care infertility referral center. PATIENT(S): Two hundred eighty-six women who underwent 344 oocyte recovery procedures for IVF cycles between March 1997 and January 1998. INTERVENTION(S): High vaginal swab specimens and endocervical swab specimens were obtained and ELISA serology was performed for detection of Chlamydia species on samples taken immediately before oocyte recovery. The results were related to the cause of infertility and the outcome parameters. MAIN OUTCOME MEASURE(S): Pregnancy rates. RESULT(S): Seropositivity for Chlamydia species and the presence of bacterial vaginosis both were strongly and independently associated with tubal disease. There was no difference in pregnancy rates in any of the groups regardless of their serologic status for chlamydial infection or current infection with bacterial vaginosis. CONCLUSION(S): This study provides further evidence of the pelvic pathogenicity of bacterial vaginosis. However, it shows that women who have bacterial vaginosis or who have been treated for chlamydial infection in the past achieve pregnancy rates with IVF treatment similar to those of women who have no evidence of such infections. PMID- 10521120 TI - Hysteroscopic selective salpingography. AB - OBJECTIVE: To evaluate the effectiveness of hysteroscopic selective salpingography (HSS) as a method for diagnosing the tubal proximal occlusion shown by hysterosalpingography (HSG). DESIGN: Prospective study. SETTING: Outpatient Department of Obstetrics and Gynecology, Social Insurance Saitama Chuo Hospital, Urawa, Japan. PATIENT(S): A total of 572 infertile women underwent HSG. Forty-seven of 50 women with unilateral or bilateral proximal tubal occlusion demonstrated by HSG underwent HSS. INTERVENTION(S): Hysteroscopic selective salpingography was performed for the diagnosis of tubal occlusion in cases in which the proximal tubal occlusion was shown by HSG. MAIN OUTCOME MEASURE(S): Number of patients who underwent HSS and pregnancy rate after HSS. RESULT(S): Twenty-seven (79.4%) of 34 patients with unilateral occlusion diagnosed by HSG were shown to have normal patency by HSS. Of 12 women with bilaterally normal patent tubes confirmed by HSS, 8 (66.7%) achieved normal pregnancies within 1 year. Seven (53.8%) of 13 patients with bilateral occlusion found by HSG were shown to have normally patent tubes by HSS. CONCLUSION: The simple method of HSS was clinically effective for evaluating the presence of proximal tubal occlusion. PMID- 10521121 TI - Computerized assessment of facial hair growth. AB - OBJECTIVE: To evaluate a computer-assisted technique for objective and sensitive monitoring of facial hair growth. DESIGN: Prospective study. SETTING: Department of Gynecological Endocrinology and Reproductive Medicine and Clinic for Ear, Nose, and Throat, General Hospital, University of Vienna, Vienna, Austria. PATIENT(S): Four men, three hirsute women, and three nonhirsute women. INTERVENTION(S): Using video equipment and computer software, we were able to document, analyze, and store data regarding hair growth in specific areas of interest. For digital image analysis, we used the Digi Trace System (Olympus, Vienna, Austria; Imatec, Munich, Germany). MAIN OUTCOME MEASURE(S): Hair growth within 20 days in well-defined regions of interest on the faces of hirsute and nonhirsute women and of men. RESULT(S): Hair growth on day 21 was significantly different between hirsute and nonhirsute women as well as in men. The scores for individual hair growth between day 0 and day 21 also were significantly different in hirsute women and in men. No statistically significant difference in hair growth was found within the group of nonhirsute women. CONCLUSION(S): With digital image analysis, facial hair growth, especially in hirsute women, can be calculated in a sensitive and objective manner. PMID- 10521122 TI - Intrauterine surgery using a new coaxial bipolar electrode in normal saline solution (Versapoint): a pilot study. AB - OBJECTIVE: To determine the feasibility and clinical outcome of using a new hysteroscopic bipolar electrosurgical system. DESIGN: Pilot feasibility study. SETTING: A university teaching hospital. PATIENT(S): Eight women with intrauterine lesions. Six women with impaired fertility and two with abnormal bleeding were selected. INTERVENTION(S): Three septoplasties, two adhesiolysis procedures, two polypectomies, one myomectomy, and one endometrial ablation were performed with the Versapoint system (Gynecare Inc., Menlo Park, CA). MAIN OUTCOME MEASURE(S): Fertility, obstetric outcome, and menstrual blood loss. RESULT(S): Five of six patients conceived six normal fetuses. Menstrual blood loss improved significantly in the remaining two patients. CONCLUSION(S): The Versapoint system is an effective alternative in the treatment of intrauterine lesions. PMID- 10521123 TI - A case of successful conservative chemotherapy for intramural pregnancy. AB - OBJECTIVE: To describe a rare case of intramural pregnancy diagnosed with the use of magnetic resonance imaging (MRI) and conservatively managed. DESIGN: Case report. SETTING: Department of obstetrics and gynecology in a university hospital. PATIENT: A 33-year-old healthy patient with a history of a partial mole after 3 years of primary unexplained infertility. INTERVENTION(S): Laparoscopic and transvaginal local injections of methotrexate. MAIN OUTCOME MEASURE(S): Transvaginal ultrasound (US) and MRI findings. RESULT: Treatment was successful, with no complications, and the patient's reproductive potential was preserved. CONCLUSION(S): Early detection of intramural pregnancy with the use of transvaginal US is important, and MRI is a useful, noninvasive imaging modality. Chemotherapy with methotrexate is an effective treatment that allows preservation of reproductive potential. PMID- 10521124 TI - Sperm freezing--species specific? PMID- 10521125 TI - The debate continues on "money back guarantees". PMID- 10521126 TI - Angiotensin-converting enzyme--a role in reproduction? Questions to be answered. PMID- 10521127 TI - Important or "much ado about nothing?". PMID- 10521128 TI - Sex ratios--the wisdom of a lifetime. PMID- 10521129 TI - Aspirin and reproductive performance? PMID- 10521130 TI - Aspirin and reproductive performance? PMID- 10521131 TI - Aspirin and reproductive performance? PMID- 10521132 TI - Aspirin and reproductive performance? PMID- 10521133 TI - Assessment factors for human health risk assessment: a discussion paper. AB - The general goal of this discussion paper is to contribute toward the further harmonization of human health risk assessment. It first discusses the development of a formal, harmonized set of assessment factors. The status quo with regard to assessment factors is reviewed, that is, the type of factors to be identified, the range of values assigned, as well as the presence or absence of a scientific basis for these values. Options are presented for a set of default values and probabilistic distributions for assessment factors based on the state of the art. Methods of combining default values or probabilistic distributions of assessment factors are also described. Second, the effect parameter, the no-observed-adverse effect level (NOAEL), is discussed. This NOAEL as selected from the toxicological database may be a poor substitute for the unknown, true no-adverse-effect level (NAEL). New developments are presented with respect to the estimation of the NAEL. The already widely discussed Benchmark Dose concept can be extended to obtain an uncertainty distribution of the Critical Effect Dose (CED). This CED distribution can be combined with estimated uncertainty distributions for assessment factors. In this way the full distribution of the Human Limit Value will be derived and not only a point estimate, whereas information on dose response relations is taken into account. Finally, a strategy is proposed for implementation of the new developments into human health risk assessments. PMID- 10521134 TI - Tumor necrosis factor alpha and toxicology. AB - The molecular cloning of a group of proteins, collectively referred to as cytokines, and including interleukins, chemokines, growth factors, colony stimulating factors, and tumor necrosis factors, has allowed for the increased understanding of the mechanisms for many disease processes as well as provided strategies for the development of novel therapies. Conceptually similar to hormones and peptides, this group of phylogenetically related molecules are also involved in various toxicological processes, including apoptosis, cell repair, and in particular inflammation. In this review, we offer a description of what many believe represents the primary regulatory cytokine, tumor necrosis factor (TNF)alpha and its role in toxicological processes. For over a decade it has been suspected that this molecule helps mediate the shock state induced by bacterial endotoxin and the wasting diathesis that typifies chronic diseases. Advances in molecular biology that have provided tools to modulate TNFalpha regulation and synthesis have allowed for the identification of additional roles for TNFalpha in homeostasis, cellular damage, and repair. This review provides a brief summary of our understanding of TNFalpha biology followed by a discussion of its role in toxicological responses. This is followed by specific examples of organ-specific and tissue-specific responses to chemical damage where TNFalpha has been implicated. The review concludes with a review of its implication in human risk assessment, particularly as it relates to genetic polymorphisms of TNFalpha expression and disease susceptibility. PMID- 10521136 TI - Posttraumatic cerebral vasospasm: clinical and morphological presentations. AB - Head injury is one of the leading causes of morbidity and mortality in the young population. Many factors complicate head injury and worsen an outcome. One of these factors is posttraumatic cerebral vasospasm. We studied 75 patients admitted to the University of Mississippi Medical Center with head injury. Their ages ranged from 14 to 67 years (mean 30 years, SD 11.63). Eighty percent of the patients were men, and 20% were women. Of these patients, 53 (70.6%) suffered severe blunt trauma, and 4 patients suffered gunshot wounds to the head. Four patients had mild head injury, and 14 had moderate head injury. Posttraumatic vasospasm was detected in 24 (32%) patients. Among these patients, 19 (79.2%) had severe closed head injury, 3 patients had moderate head injury, and 2 suffered gunshot wounds. The severity of the patient's respective condition was correlated with the development of posttraumatic cerebral vasospasm: 50% of the patients with Glasgow Coma Scale (GCS) 3-4 developed PTV, and only 30% with GCS 9-11, and none of the patients with GCS > 12 developed PTV. Overall, posttraumatic vasospasm started earlier and had a shorter course than did aneurysmal vasospasm. Morphologically, posttraumatic vasospasm resembled the features of aneurysmal vasospasm. We found increased corrugation of the internal elastic lamina and increased amounts of connective tissue in the subendothelial layer. These findings show that posttraumatic vasospasm, although clinically more mild, demonstrates the same morphological changes as aneurysmal vasospasm. PMID- 10521135 TI - Calpain activation and cytoskeletal protein breakdown in the corpus callosum of head-injured patients. AB - Calpain-mediated breakdown of the cytoskeleton has been proposed to contribute to brain damage resulting from head injury. We examined the corpus callosum from patients who died after a blunt head injury in order to determine if there was evidence of these pathophysiological events in a midline myelinated commissure that is susceptible to damage after human head injury. Western blotting revealed marked reductions in the levels of neurofilament triplet proteins 200 and 68kDa in the corpus callosum of head-injured patients compared with control subjects. Neurofilament 200kDa levels were significantly reduced as detected by either phosphorylation-dependent or -independent antibodies. In contrast, there were minimal changes in the levels of beta-tubulin or the microtubule-associated protein, tau, in the head-injured patients, although amyloid precursor protein immunostaining demonstrated axonal damage in 9 of the 10 patients. The inactive 800kDa and active 76kDa subunits of mu-calpain were present in control subjects and head-injured patients. However, there was a significant increase in the levels of calpain-mediated spectrin breakdown products in head-injured patients compared with the control subjects. The results demonstrate that following human blunt head injury, there is a significant degradation of neurofilament proteins and increased levels of calpain-mediated spectrin breakdown products within the corpus callosum. Therefore, our data support the hypothesis that calpain-mediated breakdown of the cytoskeleton may contribute to axonal damage after head injury. PMID- 10521137 TI - Secondary neurologic injury resulting from nonhypotensive hemorrhage combined with mild traumatic brain injury. AB - Although the emergency physician often treats patients with multiple injuries, there are relatively few clinically relevant models that mimic these situations. To describe the changes after a hemorrhagic insult superimposed on traumatic brain injury (TBI), anesthetized and ventilated juvenile pigs were assigned to 35% hemorrhage (35H), TBI (via fluid percussion); TBI + 35H, and TBI + 40H (40% hemorrhage). Animals were resuscitated with shed blood and crystalloid. Hemodynamic, metabolic, behavioral, and histologic parameters were assessed for 48 h. In TBI, mean arterial pressure (MAP) was not significantly different from baseline. For TBI + 40H, MAP fell by 60% (p < 0.05). This was corrected with resuscitation. Interestingly, TBI + 35H did not show a fall in MAP, while in 35H, MAP was reduced similarly to the TBI + 40H group. ICP was elevated only initially in the TBI group. In TBI + 40H and TBI + 35H, ICP increased markedly with resuscitation, remaining elevated for 60 min. ICP remained at baseline with 35 H. Hemorrhagic focal cerebal contusions at the gray-white interface were observed in 3/5 of TBI + 40H and 5/7 of TBI + 35H. Despite the presence of subarachnoid hemorrhage (SAH) in all the animals in the TBI alone group, none of these animals demonstrated grossly discernible intraparenchymal injury. There was no evidence of intracranial injury in the 35H group. Only in animals receiving a secondary insult of hemorrhage following the primary TBI were cerebral contusions found. These experiments demonstrate the evolution of cerebral contusions as a form of secondary neurologic injury following resuscitation from traumatic brain injury and hemorrhage, even in the absence of significant blood pressure changes. PMID- 10521138 TI - Cyclosporin A significantly ameliorates cortical damage following experimental traumatic brain injury in rodents. AB - Experimental traumatic brain injury (TBI) results in a rapid and significant necrosis of cortical tissue at the site of injury. In the ensuing hours and days, secondary injury exacerbates the original damage, resulting in significant neurological dysfunction. Young adult animals were treated either 5 min before or immediately after a cortical injury with the immunosuppressant cyclosporin A (CsA). All animals treated with CsA demonstrated a significant reduction in the amount of cortical damage 7 days following TBI. The effect was observed in adult rats and in two different strains of adult mice following systemic administration of the drug. Cyclosporin A has known effects on mitochondria by inhibiting the opening of the permeability transition pore and maintaining calcium homeostasis. These results with a clinically approved drug demonstrate an almost 50% reduction in lesion volume and suggest that the mechanisms responsible for tissue necrosis following TBI are amenable to manipulation. Since CsA also has known interactions with calcineurin and may be providing neuroprotection through that mechanism, additional animals were treated with the immunosuppressant FK 506. FK 506 failed to protect against the cortical damage. Amelioration of cortical damage with CsA indicates that pharmacological therapies can be devised that will significantly alter neurological outcome after injury. PMID- 10521139 TI - The novel pyrrolopyrimidine PNU-101033-E improves facial motor neuron survival following intracranial axotomy of the facial nerve in the adult rat. AB - Neuronal survival is important to functional restitution following axotomy. Proximal lesions of the facial nerve, due to head trauma or tumor growth, for example, may cause long-standing or even permanent facial nerve palsy. Betamethasone has been used by several neurosurgical clinics for the treatment of postoperative facial nerve palsy; however, this practice is based only on clinical experience. The aim of the present study was to explore the putative effect on facial motor neuron survival of a novel lazaroid (pyrrolopyrimidine, PNU-101033-E) and furthermore to compare the effects with those of betamethasone, following intracranial transection of the facial nerve in adult rats. Both agents are known to inhibit lipid peroxidation by free radical scavenging. The lesion model used has recently been reported to induce massive neuronal cell death with a relative survival of 26.8 +/- 11.3% 1 month after lesion. Oral administration of lazaroids or daily injections of betamethasone followed surgery for 1 month, after which quantification of motor neuronal profiles was performed in the facial nucleus. Lazaroid-treated animals showed a significantly enhanced neuronal survival (68.0 +/- 9.8%), whereas no significant difference was found in betamethasone-treated animals (33.1 +/- 11.7%). The microglial and astrocytic responses in the facial nucleus were intense on the operated sides in betamethasone-treated as well as lazaroid-treated animals, and no differences in comparison with untreated animals were found. In conclusion, we found that the novel pyrrolopyrimidine PNU-101033-E, but not betamethasone, significantly enhanced nerve cell survival. This agent may therefore serve as a useful neuroprotective agent following intracranial trauma to the facial nerve and should be further evaluated for clinical use. PMID- 10521140 TI - Age, outcome, and rehabilitation costs after paraplegia caused by traumatic injury of the thoracic spinal cord, conus medullaris, and cauda equina. AB - The object of this study was to investigate the relationships of age on neurologic and functional outcome, hospitalization length of stay (LOS), and hospital charges after spinal cord injury (SCI). At 20 medical centers, 2,169 consecutive adult patients with paraplegia SCI were assessed in acute care and inpatient rehabilitation. Outcome and treatment measures included the ASIA motor index score, functional independence measure, discharge to community ratio, LOS, and hospital charges. Age differences were examined by separating the sample into 11 age categories and conducting one-way analyses of variance on treatment, medical expense, and outcome measures that included the Functional Independence Measure (FIM) and ASIA motor index scores. Cramer's statistic was used to derive a chi-square value that indicated whether variables differed significantly in terms of age. Post-hoc Tukey tests were also performed. Age-related differences were found with multiple demographic variables. Significant differences between age categories were found with regard to the following treatment measures: ASIA motor index scores at acute-care admission and at discharge, rehabilitation LOS, inpatient rehabilitation hospitalization charges, total LOS, total hospitalization charges, FIM scores at inpatient rehabilitation admission and discharge, FIM change, and FIM efficiency. In conclusion, in patients with paraplegia, age appears to adversely affect functional outcome, rehabilitation LOS, and hospital costs. However, neurologic recovery as defined by the ASIA motor scores does not appear to be related to age. PMID- 10521141 TI - Basic fibroblast growth factor (bFGF) enhances tissue sparing and functional recovery following moderate spinal cord injury. AB - The rapid increase in basic fibroblast growth factor (bFGF) production following spinal cord injury (SCI) in rats is thought to serve a role in the cellular processes responsible for the functional recovery often observed. In this study, bFGF was intrathecally administered continuously for 1 week beginning 30 min after a moderate (12.5 mm) spinal cord contusion in adult rats using the New York University impactor device. Osmotic minipumps were implanted into the lateral ventricle and lumbar thecal sac to deliver bFGF at a rate of 3 microg or 6 microg per day versus control vehicle. Animals were behaviorally tested for 6 weeks using the Basso, Beattie, Bresnahan locomotor rating scale and histologically assessed for both tissue sparing and glial reactivity rostral and caudal to the lesion. Rats treated with bFGF regained coordinated hindlimb movements earlier than controls and demonstrated consistent coordination from 4 to 6 weeks. Vehicle treated rats showed only modest improvements in hindlimb function. The amount of spared tissue was significantly higher in bFGF-treated rats than in controls. Astrocyte and microglial reactivity was more pronounced in bFGF-treated animals versus controls. In summary, intrathecal infusion of exogenous bFGF following SCI significantly reduces tissue damage and enhances functional recovery. Early pharmacological intervention with bFGF following SCI may serve a neuroprotective role and/or create a proregenerative environment, possibly by modulating the neuroglial response. PMID- 10521142 TI - Effect of lactacidosis on cell volume and intracellular pH of astrocytes. AB - Acute traumatic or ischemic cerebral lesions are associated with tissue acidosis leading to cytotoxic brain edema, predominantly affecting astrocytes. Glial swelling from acidosis is believed to be the attempt of cells to maintain a physiological intracellular pH (pHi). However, this concept, potentially important for the development of new treatment strategies for cytotoxic brain edema, has not been validated experimentally. In the present study, cell volume and pHi of astrocytes were measured simultaneously in vitro. Exposure of suspended astrocytes to levels of acidosis found in vivo during ischemia and trauma (pH 6.8-6.2) led to a maximal increase in cell volume of 121.2% after 60 min (n = 5, p < 0.05) and to immediate intracellular acidification close to extracellular levels (pH 6.2, n = 5, p < 0.05). Inhibition of membrane transporters responsible for pHi regulation (0.1 mM amiloride for the Na+/H+ antiporter or 1 mM SITS for HCO3- -dependent transporters) inhibited cell swelling from acidosis but did not affect the profound intracellular acidification. In addition, acidosis-induced cell swelling and intracellular acidification were partly prevented by the addition of ZnCl2 (0.1 mM), an inhibitor of selective proton channels not yet described in astrocytes (n = 5, p < 0.05). In conclusion, these data demonstrate that glial swelling from acidosis is not a cellular response to defend the normal pHi, as had been thought. If these results obtained in vitro are transferable to in vivo conditions, the development of blood-brain barrier-permeable agents for the inhibition of acidosis-induced cytotoxic edema might be therapeutically useful, since they do not enhance intracellular acidosis and thus cell damage. PMID- 10521143 TI - First observations of the safety and tolerability of a competitive antagonist to the glutamate NMDA receptor (CGS 19755) in patients with severe head injury. AB - A dose escalation, safety, and tolerability study of a competitive antagonist to the N-methyl-D-aspartate (NMDA) glutamate receptor (CGS 19755, Selfotel) in patients with severe head injury is reported. The drug was administered i.v. on two separate occasions, 24 h apart, to 31 patients. The dosage was escalated during the study from 1 mg/kg to 6 mg/kg. Continuous monitoring of mean arterial pressure (MABP), intracranial pressure (ICP), cerebral pressure (CPP), arterial oxygen saturation (SaO2), jugular bulb oxygen saturation (SjO2), and temperature was performed. Intermittent measurements of middle cerebral artery (MCA) velocity via transcranial Doppler ultrasound were also made 2 h before drug administration and continued for 24 h after dosing. The patients were ventilated and sedated with morphine and either midazolam or propofol. There were no behavioral changes during or after administration of the drug, and disorders of perception were reported by only three subjects, several days after relatively low doses; these were transient and were not recalled at later follow-up. We did not detect consistent changes in any of the hemodynamic parameters monitored, up to dosages of 3 mg/kg. After higher doses, some patients showed changes in MABP, ICP, and temperature during the 4 to 8-h period following the first bolus of the drug, with a return toward baseline afterwards. No consistent, serious, adverse events were considered to be due to drug effects, and death, in the one patient who died, was due to the effects of the injury. Our results indicate that CGS 19755 may be given at dosages < or = 3-5 mg/kg with acceptable safety and tolerability in stable, ventilated, and carefully monitored severe head-injured patients. PMID- 10521144 TI - Dietary beta-carotene protects lung and liver parenchyma of rats treated with monocrotaline. AB - Some studies have indicated that the injury induced by the hepato- and pneumotoxin monocrotaline (MCT) is in part mediated by oxidation. Because beta carotene is a potent antioxidant, we hypothesized that it would protect the lung and liver parenchyma against MCT-induced injury. Twenty rats were assigned randomly to four groups. All rats were fed a standard AIN93G diet with or without beta-carotene. After 1 week on the purified diets, half of the rats fed the control (standard) diet and half of the rats fed the beta-carotene-supplemented diet were injected subcutaneously with 60 mg MCT/kg body weight or its vehicle (water). All rats were sacrificed at 4 weeks. Histological examination showed that beta-carotene alone did not affect lung or liver structure. On the other hand, lungs of MCT-treated rats had severe focal pneumonia, extensive deposition of collagen in the septa, marked inflammation of the small arteries and arterioles, and arterialization of the small venules. Livers of MCT-treated rats showed some fatty infiltration and diffuse hemorrhages, more prominent sometimes in the centrilobular area and sometimes in the periportal region. Concomitant treatment with beta-carotene protected the lung parenchyma from the inflammatory reaction and the septal fibrosis, but did not prevent cardiac right ventricular hypertrophy and only slightly reduced the thickening of the wall of small arteries and arterioles. Incidence of steatosis and hemorrhages was decreased in the liver. These results indicate that MCT-induced pulmonary vascular remodeling occurs in the absence of inflammatory cell infiltration. Furthermore, beta carotene prevented inflammation and protected the lung and liver parenchyma of MCT-treated rats. PMID- 10521145 TI - An in vitro and in vivo study of some biological and biochemical effects of Sistrurus Malarius Barbouri venom. AB - Some possible biological and biochemical effects of Sistrurus Malarius Barbouri (SMB) crude venom were investigated. The acute median lethal doses of the venom under investigation were found to be 14.4 and 9.72 microg/g body weight (b.w.), respectively, in rats on i.p. administration. The possible neurotoxicity of acute, subchronic and chronic doses was investigated in-vivo and in-vitro. The venom at a dose level of 2 microg/g b.w. significantly impaired motor coordination, learning and retention, spontaneous activity and produced behavioural changes, muscle weakness and loss of righting reflex in mice. The same dose also produced a significant decrease in body temperature and inhibited acetylcholine-induced contraction of the isolated smooth (rabbit intestine) and skeletal (frog rectus abdominis) muscles and impaired transmission at the nerve muscle synapse of the rat phrenic nerve diaphragm preparation. The effects of the acute sublethal and chronic doses on carbohydrate metabolism revealed a hyperglycemic effect associated with a diminution of liver and muscle glycogen, while its effects on blood electrolytes (sodium and potassium) showed a significant elevation in the blood sodium level and a significant reduction in that of potassium. Serum enzymes were also affected. Levels of alkaline phosphatase (ALP), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities were moderately increased. The crude venom had an aggregatory effect on platelets and had also a phospholipase A2 activity while, on the other hand, it showed no L-amino acid oxidase activity. Testing of the effect of the venom on the plasma recalcification time showed that the venom had an anticoagulant effect in case of high dose (200 microg), while a coagulant effect was produced at a low dose of the venom (2.5 microg). SMB venom at a dose level of 1.94 microg/g b.w. (LD10) was found to exhibit no significant inhibitory effect on tumor growth when injected into mice. PMID- 10521146 TI - Strain-dependent disposition of inorganic arsenic in the mouse. AB - Recent studies have suggested that polymorphisms in the methylation of inorganic arsenic (iAs) exist in animals and humans. Methylation of iAs is an important step in the elimination of arsenic. The objective of this study was to examine whether there are differences in iAs disposition, and hence methylation, between three strains of mice. Ninety-day-old female mice (strains: C3H/HeNCrlBR, C57BL/6NCrlBR, and B6C3F1/CrlBR) were administered [73As]arsenate or [73As]arsenite orally at dose levels of 0.5 or 5.0 mg As/kg. Another group of mice were administered [73As]arsenate (5.0 mg As/kg) intraperitoneally (i.p.). Disposition of [73As] was assessed by whole-body counting, and analysis of urine, feces and tissues for radioactivity. Urine was analyzed by chromatography for arsenic metabolites. Several strain- and dose-related effects in the disposition of [73As] were observed with both arsenicals. After oral administration, the clearance of [73As]arsenate, measured by whole-body counting, was dependent on the strain. However, because there was no strain dependence on clearance of [73As]arsenate administered i.p., the effect after oral administration may be due to a difference in absorption of arsenate between the strains. With increased oral dose of arsenate and arsenite, the clearance of [73As] was slower and there was higher tissue retention of [73As]. The percentage of metabolites excreted in urine also was affected by the administered dose. With increased dose, the percentage of arsenite and monomethylarsonic acid were significantly increased, and dimethylarsinic acid decreased. However, our results suggest there is no overall difference between these strains of mice with respect to disposition of iAs. A better understanding of the role of phenotype in the disposition and toxicity of iAs would reduce the uncertainty in arsenic risk assessment. PMID- 10521147 TI - Acute moderate hypoxia reduces ethanol elimination in the conscious rabbit. AB - Hypoxic states decrease the catalytic activity of certain enzymes of the cytochrome P450, leading to decreased clearance of selected xenobiotics. It is not known whether hypoxia can affect the activity of cytosolic enzymes such as liver alcohol dehydrogenase (LADH). This study was carried out to examine the effect of experimentally induced acute moderate hypoxia on the kinetics of ethanol and on the catalytic activity of LADH in the conscious rabbit. To this purpose, the kinetics of elimination of ethanol (70 mg/kg intravenously) were studied in male rabbits (n = 6) kept in an atmosphere with a fractional concentration of O2 (FiO2) of 0.12 for 24 h, and in rabbits (n = 6) breathing room air with a FiO2 approximately 0.21 (controls). Compared with controls, the clearance of ethanol was reduced by 32% (P < 0.05) in rabbits exposed to acute moderate hypoxia. In rabbits that consumed approximately 6 g/kg/day ethanol in their drinking water for 14 days (n = 6), acute moderate hypoxia reduced the clearance of ethanol to the same extent as observed in rabbits receiving a single dose of ethanol. Hypoxia did not change the in vitro Vmax or Km of LADH. This study demonstrates that acute moderate hypoxia reduces the clearance of ethanol following single or multiple administrations. PMID- 10521148 TI - Time- and dose-dependent digoxin redistribution by digoxin-specific antigen binding fragments in a rat model. AB - To study the influence of the interval between digoxin intake and digoxin specific antigen binding fragment (DSFab) administration, we developed a rat kinetic model. 3H-digoxin (0.77 nmol/kg) was injected by intravenous route and DSFab was injected at different times (12, 30 or 60 min) corresponding to different levels of 3H-digoxin distribution (50, 83 and 100%). The effect of increasing the molar DSFab/3H-digoxin ratio from 1 to 5 was also investigated. To evaluate DSFab effect on the 3H-digoxin pharmacokinetics, we also investigated the pharmacokinetics of the 125I-DSFab and DSFab-3H-digoxin complex. 3H-digoxin and DSFab-3H-digoxin complex pharmacokinetics showed that DSFab altered immunoreactive 3H-digoxin pharmacokinetics. In redistribution studies performed 12, 30 or 60 min after 3H-digoxin injection, DSFab bound immunoreactive 3H digoxin including native 3H-digoxin and active metabolites of 3H-digoxin. This binding induced a redistribution process of immunoreactive 3H-digoxin in the DSFab distribution compartment and was evaluated by the redistribution fraction (F(R)). F(R) was 23% lower at 60 min than at 12 and 30 min, and by increasing the DSFab/3H-digoxin ratio from 1 to 5, F(R) increased by 60%. In conclusion, the longer the time interval between digoxin intake and DSFab administration, the lower the efficacy of the redistribution process. This effect could be reduced by increasing the DSFab dose. PMID- 10521149 TI - Apoptosis in the human embryo. AB - Preimplantation human embryos are characterized by various degrees of cytoplasmic fragmentation, and a high incidence of developmental arrest before the blastocyst stage. This review examines the current morphological and biochemical evidence that apoptosis plays a role in early human development and embryonic loss. Embryos examined 24 h or more after arrest often show characteristic features of apoptosis including cytoplasmic, nuclear and DNA fragmentation. In contrast, embryos of good morphology that appear to be developing normally show no evidence of apoptosis before compaction. However, at the morula and blastocyst stages, scattered cells with fragmented nuclei and DNA characteristic of cells undergoing apoptosis are common features. Apoptosis may result from suboptimal culture conditions, or may be involved in the elimination of abnormal cells. However, the causes, role and regulation of apoptosis in the human preimplantation embryo remain to be determined. PMID- 10521150 TI - Gamete recognition: sperm proteins that interact with the egg zona pellucida. AB - The gamete recognition and initial binding processes that are crucial for the success of mammalian fertilization are mediated by moieties associated with the extracellular matrix of the egg (the zona pellucida) and the head of the fertilizing spermatozoon. The zona proteins involved have been characterized in some detail, with ZP3 and ZP2 generally acknowledged to be responsible for the initial (primary) and secondary interactions, respectively. However, the identity of the complementary molecules on the sperm surface is highly contentious and remains unresolved. This review summarizes the current knowledge and controversies in this research area. The credentials of some of the major candidates and the probability of the involvement of multiple sperm receptors with different binding characteristics are assessed. Resolving this very important gap in our understanding is an essential prerequisite to understanding fully the molecular and signal transduction events that cause sperm acrosomal exocytosis. Such fundamental information is also imperative for the development of novel forms of contraception (or sterilization) targeted against specific sperm epitopes. Moreover, this information may contribute to our understanding of certain types of male infertility. PMID- 10521151 TI - Role of reproductive technologies and genetic resource banks in animal conservation. AB - In combination with modem reproductive technologies, there is potential to use frozen and stored germplasm (genetic resource banks) to support conservation measures for the maintenance of genetic diversity in threatened species. However, turning this idea into reality is a complex process, requiring interdisciplinary collaboration and clearly defined goals. As the number of species deserving the attention of conservation scientists is overwhelmingly large, yet detailed knowledge of reproductive physiology is restricted to relatively few of them, choosing which species to conserve is one of the most difficult issues to be tackled. Besides the direct application of technologically advanced reproductive procedures, modern approaches to non-invasive endocrine monitoring play an important role in optimizing the success of natural breeding programmes. Through the analysis of urine and faecal samples, this type of technology provides invaluable management information about the reproductive status of diverse species. For example, it is possible to diagnose pregnancy and monitor oestrous cycles in elephants and rhinos without causing stress through restraint for sample collection. In this review, we identify the potential contribution of reproductive biology and genetic resource banks to animal conservation, but also highlight the complexity of issues determining the extent to which this potential can be achieved. PMID- 10521152 TI - Protein kinases in mammalian sperm capacitation and the acrosome reaction. AB - Binding to the zona pellucida of an egg stimulates the spermatozoon to undergo the acrosome reaction, a process that enables it to penetrate the egg. Before this binding, the spermatozoon undergoes a series of biochemical transformations in the female reproductive tract, collectively called capacitation. Only capacitated spermatozoa can bind to the zona pellucida and undergo the acrosome reaction. Protein kinases may be involved in the regulation of intracellular Ca2+ during capacitation and the acrosome reaction. The first event in capacitation is the increase in intracellular calcium, bicarbonate and hydrogen peroxide, which collectively activate adenylyl cyclase to produce cyclic AMP, which activates protein kinase A to phosphorylate certain proteins. During capacitation, there is an increase in membrane-bound phospholipase C, and this binding is highly stimulated by the addition of epidermal growth factor to the cells. The capacitated spermatozoon binds to the zona pellucida of the egg via specific receptors and it is suggested that the zona pellucida binds to at least two different receptors in the sperm head plasma membrane. One is a Gi-coupled receptor that can activate phospholipase Cbeta1 and may regulate adenylyl cyclase to further increase cyclic AMP concentrations. The cyclic AMP activates protein kinase A to open a calcium channel in the outer acrosomal membrane, resulting in a relatively small increase in cytosolic calcium. This increase in Ca2+ leads to activation of phospholipase Cgamma, which is coupled to the second tyrosine kinase receptor. The products of phosphatidyl-inositol bisphosphate hydrolysis by phospholipase C, diacylglycerol and inositol-trisphosphate, induce the activation of protein kinase C and a calcium channel in the outer acrosomal membrane, respectively. Protein kinase C opens a calcium channel in the plasma membrane and, together with the inositol-trisphosphate-activated calcium channel, leads to a second and higher increase in cytosolic calcium. In addition, the depletion of calcium in the acrosome activates a capacitative calcium entry mechanism in the plasma membrane, leading to a rapid increase in cytosolic calcium (300-500 nmol l(-1)). This increase in intracellular calcium concentration (and pH) leads to membrane fusion and the acrosome reaction. PMID- 10521153 TI - Role of insulin-like growth factor binding protein 1 in human pregnancy. AB - Insulin-like growth factors and their binding proteins are key regulators of fetal and maternal tissue growth and development during human pregnancy. Insulin like growth factors, particularly IGF-II, are produced in abundance by the trophoblast cells of the placenta, whereas one of the insulin-like growth factor binding proteins, IGFBP-1, is the major secretory product of the maternal decidualized endometrium. This spatial (and temporal expression) of the insulin like growth factor axis infers a sophisticated paracrine regulatory mechanism for controlling insulin-like growth factor function. This paper reviews the potential roles of IGFBP-1 in human pregnancy by examining its effects on growth, metabolism and migration at the maternal-fetal interface and how these might be influenced by autocrine-paracrine post-translational modifications. PMID- 10521154 TI - Influence of social factors on immune function and reproduction. AB - Animals are presented with continuous energy demands that vary seasonally. For example, during the winter many small mammals and birds inhibit reproduction and growth and funnel energy into thermogenesis or cellular maintenance. As energy shortages become more severe, survival may become compromised because processes such as immune function and thermogenesis are impaired. Thus, there are trade offs between energetically expensive processes such as reproduction and immune function. In this review, the immune function and reproduction of seasonally breeding species are evaluated in relation to social interactions. It is proposed that individuals maintain the highest degree of immune function that is energetically possible within the constraints of other survival needs, as well as growth and reproduction, in habitats in which energy requirements and availability often fluctuate. It is hypothesized that extrinsic factors, such as social environment, modulate energy allocation to reproductive and immune function and that hormonal mechanisms underlie the partitioning of energy to various physiological components. PMID- 10521155 TI - Male and female isoenzymes of steroid 5alpha-reductase. AB - There are two steroid 5alpha-reductase isoenzymes, designated type 1 and type 2, in mammals and recent experiments show that each plays a unique physiological role. In this article, the hypothesis is developed that the type 1 gene specifies a female isoenzyme, whereas the type 2 gene specifies a male isoenzyme. This idea results from the following observations. First, mutation of the 5alpha-reductase type 1 gene in mice affects reproduction in females by decreasing fecundity and blocking parturition, but has no effect on reproduction in males. Second, mutation of the 5alpha-reductase type 2 gene in mice and men prevents proper virilization but does not affect development or reproductive function in females. Analyses of these diverse phenotypes indicate that the isoenzymes catalyse both anabolic and catabolic reactions in steroid hormone metabolism. PMID- 10521156 TI - Teenage fertility and life chances. AB - Teenage mothers and their children face poorer prospects in life than do women who delay motherhood until later in life. Moreover, patterns of early childbearing tend to be repeated in subsequent generations. Therefore, an understanding of the factors associated with early fertility is important for the prevention of adverse consequences. This paper uses data from the National Survey of Sexual Attitudes and Lifestyles to explore these associations. Early sexual intercourse is an important predictor of early fertility, as is poor educational attainment, although it is not clear to what extent pregnancy acts to thwart academic ambitions, or to what extent poor educational performance leads to a need to seek personal fulfilment in other than academic goals. Thus, interventions designed to influence age at first intercourse and to improve educational performance both have potential in terms of impacting on teenage pregnancy rates. Family background also exerts a powerful influence on teenage fertility. Young people for whom one or both parents are absent are more likely to become parents early in life. However, the most important factor of family life determining the chances of teenage motherhood appear to be the quality of communication about sexual matters with the home. In terms of outcomes, teenage mothers are more likely to live in social housing, are less likely to be in paid employment and have larger than average sized families. Certain areas of the country, notably the older, run-down industrial areas, have higher rates of teenage motherhood than the newer, more prosperous areas. Because most of these effects are independent of one another, there is potential merit in intervening to prevent unintended conception at several points in a young woman's life. Primary preventive efforts are needed to reduce the rates at which teenage pregnancy occurs in this country. Yet, if the cycle of deprivation that means the children of young mothers themselves enter parenthood early is to be broken, then efforts must also be made to mitigate the effects of teenage fertility for both mother and child. PMID- 10521158 TI - Aging and peaking. PMID- 10521159 TI - Update of geriatric psychiatry practices among American psychiatrists: Analysis of the 1996 National Survey of Psychiatric Practice. AB - Using data from the 1996 National Survey of Psychiatric Practice from the American Psychiatric Association (APA), the authors updated information on psychiatrists who are high geriatric providers (HGPs). In 1996, HGPs comprised 18% of the sample. Only 23% reported no geriatric patients in their practice, a 51% reduction from 1988-89; the proportion of HGPs is increasing. HGPs were more often male, minority, international medical school graduates, certified in geriatric psychiatry, and not medical school-affiliated. HGPs worked longer hours/week in direct patient care, had more patient visits/week, and saw more new patients/month, spending more time in hospitals and nursing homes and less time in office-based practice, and seeing more patients with mood disorders, psychotic disorders, and other disorders. Medicare was a proportionally higher payment source. Older psychiatrists were likely to have more patients over age 65. Tracking practice activities of HGPs may help inform policy discussion regarding staffing needs for geriatric patients with late-life mental disorders. PMID- 10521160 TI - Age and suicidal ideation in older depressed inpatients. AB - Age-related patterns of symptom presentation may complicate the recognition of suicide risk. The authors sought to determine whether there is a relationship between age and reported suicidal ideation in depressed suicide attempters (DSAs) and depressed nonattempters (DNs) 50 years of age and over. Regression analyses revealed that increasing age is significantly associated with the absence of suicidal ideation in both DSAs and DNs. Because of their lower rates of depressed mood and suicidal ideation, the depressions of older adults may more readily escape detection. Preventive or treatment measures initiated after the onset of the suicidal state may be insufficient, and other preventive strategies ought to be considered. PMID- 10521161 TI - Cost analysis, policy development, and Alzheimer's disease. PMID- 10521162 TI - The economic cost of Alzheimer's disease and related dementias to the California Medicaid program ("Medi-Cal") in 1995. AB - Although state Medicaid programs may bear a large portion of the costs of Alzheimer's disease (AD), current information on spending is not available. Using a health insurance claims database for a 10% random sample of California Medicaid ("Medi-Cal") recipients 60+ years of age, the authors estimated the excess cost of AD to Medi-Cal in 1995 as the difference in expenditures between an AD cohort (those with AD or related dementias) and an age- and sex-matched cohort without AD. Among 62,450 recipients, 2,575 (4.1%) were found to have AD or related dementias, and their average payments were approximately $7,700 higher (P<0.01) than those for the comparison group. These estimates suggest that Medi-Cal spends about $200 million on AD and related dementias annually, a burden that represents nearly 10% of state spending on elderly patients. PMID- 10521163 TI - Clinical and neuroradiologic features associated with chronicity in late-life depression. AB - The authors conducted a 6-year follow-up of 16 patients with late-life depression to evaluate the relationships between clinical and neuroradiologic variables and disease outcome. Patients had a comprehensive neuropsychiatric evaluation and magnetic resonance imaging (MRI) at baseline and follow-up. Eight of the 16 developed a chronic course of unremitting major depression sufficient to cause significant psychosocial impairment. Six patients with a chronic course and four patients with a non-chronic course of depression had white matter hyperintensities (WMH) on MRI at baseline. Four patients whose WMH increased in size over time developed a chronic unremitting course of depression. No patients with non-chronic depression had large areas of WMH at baseline or exhibited increased WMH size over time. Chronic depression was associated with severity of cerebrovascular risk factors, apathy, and poor quality of life. Treatment and prevention of cerebrovascular disease may improve the outcome of late-life depression. PMID- 10521164 TI - Attempted suicide in older depressed patients: effect of cognitive functioning. AB - The authors explored cognitive functioning and suicidal behavior in older depressed patients. Inpatients age 50 years or older (N=103) with major depression, 45 of whom had attempted suicide, were evaluated within 1 week of their hospital admission. Measures of suicidal behavior included suicide attempter status, the Suicide Intent Scale (SIS), ratings of method used (violent/nonviolent), and seriousness of injuries sustained (lethality). The Mini Mental State Exam (MMSE) score measured cognitive impairment; covariates were age, gender, and living arrangement. The MMSE score was not associated with suicide attempter status, but for attempters, MMSE score showed a positive association with SIS score, but not method or lethality. Findings suggest that although cognitive disturbance may be associated with less-deliberate acts among older depressed suicide attempters, it does not appear to influence the potential lethality of their behavior. PMID- 10521165 TI - Can caregivers independently rate cognitive and behavioral symptoms in Alzheimer's disease patients?: A longitudinal analysis. AB - To examine whether informant-based assessments of patients with Alzheimer's disease (AD) can be used longitudinally to track patient functioning, the authors followed AD patients (N=153) and their caregivers over 1 year with the Relative's Assessment of Global Symptomatology-Elderly (RAGS-E) and the Alzheimer's Disease Assessment Scale-Cognitive Subscale (ADASc). Factor analysis of the RAGS-E yielded two subscales, Cognitive Functioning and Mood Disturbance. The cognitive subscale and ADASc correlated at all visits, whereas the mood subscale did not. After 12 months (n=62), the cognitive scale worsened at a rate similar to the ADASc, suggesting concurrent validity. Therefore, informant-based measures appear to be reliable and valid methods of identifying cognitive change in AD patients. PMID- 10521166 TI - Alpha(1)-acid glycoprotein, age, and sex in mood disorders. AB - Serum alpha(1)-acid glycoprotein (AAG) reportedly can increase with age in the normal population, especially among women. In 33 patients with unipolar major depression or bipolar disorder, serum AAG was measured by age. The authors noted a positive association with age, particularly in depressed female patients (n=18; r(s)=0.61; P<0.01). The authors discuss implications for drug pharmacokinetics. Investigation of AAG in mood disorders needs to take age and sex into consideration. PMID- 10521167 TI - Prognostic value of electrophysiological markers in Alzheimer's disease. AB - The authors assessed the usefulness of event-related potentials (ERP) and flash visual evoked potentials (FVEP) as prognostic markers in a sample of 25 subjects with probable Alzheimer's disease (AD). ERP and FVEP recordings were obtained from 21 and 23 subjects, respectively. Ranking of the annual rate of change in cognitive scales vs. baseline ERP/FVEP latencies was compared, and those with longer FVEP N(2) component latencies showed slower subsequent rates of decline (tau=0.32; z=2.16; P=0.015), suggesting that further study of FVEP responses as prognostic markers in AD is warranted. PMID- 10521168 TI - Estrogen therapy and aggressive behavior in elderly patients with moderate-to severe dementia: results from a short-term, randomized, double-blind trial. AB - The authors used a randomized, double-blind, placebo-controlled clinical trial study design to investigate the efficacy and safety of short-term estrogen therapy in decreasing aggressive behaviors in elderly patients with moderate-to severe dementia. Estrogen therapy was associated with lower total aggression scores (P<0.030) and with decreased frequency of physical aggression (P<0.019) over the 4-week trial. Verbally aggressive behaviors were decreased relative to control subjects, although this effect was not statistically significant. No drug vs.-placebo differences were found for resistive, sexual, or self-directed aggressive behaviors. No adverse effects from the estrogen were observed during the course of the study. PMID- 10521170 TI - Appreciation to reviewers PMID- 10521169 TI - The effects of donepezil on the P300 auditory and visual cognitive evoked potentials of patients with Alzheimer's disease. AB - Donepezil is a cholinesterase inhibitor used for the treatment of patients with mild to moderately severe Alzheimer's disease (AD). The purpose of this study was to determine the effect of treatment with donepezil 5 mg qd on cognitive evoked potentials (EPs) of patients with AD. Although treatment with donepezil did not normalize EP latencies, treatment was associated with a significant decrease in the auditory P300 latency (mean latency pretreatment=401. 5 msec; posttreatment=392.7 msec.; P=0.04), and the visual P300 latency (mean latency pretreatment=605.7 msec; posttreatment=580.3 msec; P=0.04). Treatment with donepezil had no discernible effect on auditory or visual P300 EP amplitudes. PMID- 10521171 TI - Does stress cause cancer? There's no good evidence of a relation between stressful events and cancer. PMID- 10521172 TI - Only a minor part of cerebral palsy cases begin in labour. But still room for controversial childbirth issues in court. PMID- 10521173 TI - The National Service Framework: a scaffold for mental health. Implementation is key to determining whether it's a support or a gallows. PMID- 10521174 TI - Accidents that should never have happened. When technology to prevent accidents exists it should be used. PMID- 10521175 TI - Radiation risks. Appropriate decisions come from valid data, not inaccurate perceptions of risk. PMID- 10521176 TI - Alan milburn returns to UK health department PMID- 10521177 TI - Patients' bill of rights passes first US hurdle. PMID- 10521178 TI - Patients' complaints: GMC could do better PMID- 10521179 TI - JAMA appoints woman editor PMID- 10521180 TI - In brief PMID- 10521181 TI - Cell biologist wins Nobel prize. PMID- 10521182 TI - BMA opens door on human reproductive cloning. PMID- 10521183 TI - Study leads to a call for an end to spanking PMID- 10521184 TI - Dobson backed NICE ruling on flu drug. PMID- 10521185 TI - Tories give a "patient's guarantee" PMID- 10521187 TI - High PSA levels may show body is fighting cancer PMID- 10521186 TI - Irish nurses to strike. PMID- 10521188 TI - Prodi proposes food agency for the EU. PMID- 10521190 TI - Health department backs flexible working for NHS in england PMID- 10521189 TI - Doctor accused of killing patients "for enjoyment". PMID- 10521191 TI - Three jailed in bribery and prescription fraud scandal. PMID- 10521192 TI - Stressful life events and difficulties and onset of breast cancer: case-control study. AB - OBJECTIVE: To determine the relation between stressful life events and difficulties and the onset of breast cancer. DESIGN: Case-control study. SETTING: 3 NHS breast clinics serving west Leeds. PARTICIPANTS: 399 consecutive women, aged 40-79, attending the breast clinics who were Leeds residents. MAIN OUTCOME MEASURES: Odds ratios of the risk of developing breast cancer after experiencing one or more severe life events, severe difficulties, severe 2 year non-personal health difficulties, or severe 2 year personal health difficulties in the 5 years before clinical presentation. RESULTS: 332 (83%) women participated. Women diagnosed with breast cancer were no more likely to have experienced one or more severe life events (adjusted odds ratio 0.91, 95% confidence interval 0.47 to 1. 81; P=0.79); one or more severe difficulties (0.86, 0.41 to 1.81; P=0.69); a 2 year severe non-personal health difficulty (0.53, 0.12 to 2.31; P=0.4); or a 2 year severe personal health difficulty (2.73, 0.68 to 10.93; P=0.16) than women diagnosed with a benign breast lump. CONCLUSION: These findings do not support the hypothesis that severe life events or difficulties are associated with onset of breast cancer. PMID- 10521193 TI - Ecological analysis of ethnic differences in relation between tuberculosis and poverty. AB - OBJECTIVE: To examine the effect of ethnicity on the relation between tuberculosis and deprivation. DESIGN: Retrospective ecological study comparing incidence of tuberculosis in white and south Asian residents of the 39 electoral wards in Birmingham with ethnic specific indices of deprivation. SETTING: Birmingham, 1989-93. SUBJECTS: 1516 notified cases of tuberculosis. MAIN OUTCOME MEASURES: Rates of tuberculosis and measures of deprivation. RESULTS: Univariate analysis showed significant associations of tuberculosis rates for the whole population with several indices of deprivation (P<0.01) and with the proportion of the population of south Asian origin (P<0.01). All deprivation covariates were positively associated with each other but on multiple regression, higher level of overcrowding was independently associated with tuberculosis rates. For the white population, overcrowding was associated with tuberculosis rates independently of other variables (P=0.0036). No relation with deprivation was found for the south Asian population in either single or multivariable analyses. CONCLUSIONS: Poverty is significantly associated with tuberculosis in the white population, but no such relation exists for those of Asian ethnicity. These findings suggest that causal factors, and therefore potential interventions, will also differ by ethnic group. PMID- 10521194 TI - Ecological study of social fragmentation, poverty, and suicide. AB - OBJECTIVES: To investigate the association between suicide and area based measures of deprivation and social fragmentation. DESIGN: Ecological study. SETTING: 633 parliamentary constituencies of Great Britain as defined in 1991. MAIN OUTCOME MEASURES: Age and sex specific mortality rates for suicide and all other causes for 1981-92. RESULTS: Mortality from suicide and all other causes increased with increasing Townsend deprivation score, social fragmentation score, and abstention from voting in all age and sex groups. Suicide mortality was most strongly related to social fragmentation, whereas deaths from other causes were more closely associated with Townsend score. Constituencies with absolute increases in social fragmentation and Townsend scores between 1981 and 1991 tended to have greater increases in suicide rates over the same period. The relation between change in social fragmentation and suicide was largely independent of Townsend score, whereas the association with Townsend score was generally reduced after adjustment for social fragmentation. CONCLUSIONS: Suicide rates are more strongly associated with measures of social fragmentation than with poverty at a constituency level. PMID- 10521196 TI - Prevalence of overweight and obesity in British children: cohort study. PMID- 10521195 TI - Does publicity about cancer screening raise fear of cancer? Randomised trial of the psychological effect of information about cancer screening. PMID- 10521197 TI - A case of tetany due to exposure to the sun PMID- 10521200 TI - Ingenious healing PMID- 10521198 TI - Is Helicobacter pylori associated with non-ulcer dyspepsia and will eradication improve symptoms? A meta-analysis. AB - OBJECTIVES: To examine the association between Helicobacter pylori infection and non-ulcer dyspepsia, and to assess the effect of eradicating H pylori on dyspeptic symptoms in patients with non-ulcer dyspepsia. DESIGN: Systematic review and meta-analysis of (a) observational studies examining the association between Helicobacter pylori infection and non-ulcer dyspepsia (association studies), and (b) therapeutic trials examining the association between eradication of H pylori and dyspeptic symptoms in patients with non-ulcer dyspepsia (eradication trials). DATA SOURCES: Randomised controlled trials and observational studies conducted worldwide and published between January 1983 and March 1999. MAIN OUTCOME MEASURES: Summary odds ratios and summary symptom scores. RESULTS: 23 association studies and 5 eradication trials met the inclusion criteria. In the association studies the summary odds ratio for H pylori infection in patients with non-ulcer dyspepsia was 1.6 (95% confidence interval 1.4 to 1.8). In the eradication trials the summary odds ratio for improvement in dyspeptic symptoms in patients with non-ulcer dyspepsia in whom H pylori was eradicated was 1.9 (1.3 to 2.6). CONCLUSIONS: Some evidence shows an association between H pylori infection and dyspeptic symptoms in patients referred to gastroenterologists. An improvement in dyspeptic symptoms occurred among patients with non-ulcer dyspepsia in whom H pylori was eradicated. PMID- 10521201 TI - Home PMID- 10521199 TI - Fortnightly review. Treatment of schizophrenia. PMID- 10521202 TI - Tetanus in an immunised patient. PMID- 10521203 TI - ABC of complementary medicine: herbal medicine. PMID- 10521204 TI - Some hiding places of the tubercle bacillus PMID- 10521205 TI - A template for defining a causal relation between acute intrapartum events and cerebral palsy: international consensus statement. PMID- 10521206 TI - Impact of legislation on nursing home care in the United States: lessons for the United Kingdom. PMID- 10521207 TI - Is bigger better? Concentration in the provision of secondary care. PMID- 10521208 TI - The whole idea was so disgusting PMID- 10521209 TI - Mozart on death PMID- 10521210 TI - Sunlight and health. Use of sunscreens does not risk vitamin D deficiency. PMID- 10521211 TI - Intervention trial for late life depression defended. PMID- 10521212 TI - Design of CRASH trial. Evidence shows that quality of trial by Faupel et al is good and therefore should not be excluded. PMID- 10521214 TI - Influence of data display formats on decisions to stop clinical trials. Paper is misleading, like a sheep dressed in a wolf's clothing. PMID- 10521213 TI - Routine antenatal HIV testing. Is justified in areas of low HIV prevalence. PMID- 10521215 TI - Fungal infections of skin and nails of feet. Pragmatic clinical trial is now needed. PMID- 10521216 TI - Teaching medical students about bereavement is hard. PMID- 10521217 TI - Cycle helmets. BMA report does not give the whole picture. PMID- 10521218 TI - The coroner service. Coroner service could indeed be improved. PMID- 10521219 TI - Attribution of time lag theory to explain French paradox. PMID- 10521221 TI - Obituaries PMID- 10521220 TI - Pens are certainly more portable than computers. PMID- 10521222 TI - BMA establishes racial equality committee PMID- 10521223 TI - Biohazard PMID- 10521225 TI - Making use of guidelines in clinical practice PMID- 10521224 TI - Making use of guidelines in clinical practice; implementing clinical guidelines: A practical guide PMID- 10521226 TI - Breast problems PMID- 10521227 TI - This time it was not a drill PMID- 10521228 TI - Society column PMID- 10521229 TI - All a matter of tone PMID- 10521230 TI - Life stress is not associated with the onset of breast cancer PMID- 10521232 TI - Social fragmentation is associated with high suicide risk PMID- 10521231 TI - Poverty does not predict tuberculosis in asians in UK PMID- 10521233 TI - Obesity among british children is rising PMID- 10521234 TI - Dyspeptic symptoms may improve after eradication of helicobacter pylori PMID- 10521236 TI - Steve Vatner's editorship of circulation research, 1991-1999 : A note of appreciation PMID- 10521235 TI - How to determine whether cerebral palsy occurs during labour PMID- 10521237 TI - A role for the sarcolemmal Na(+)/H(+) exchanger in the slow force response to myocardial stretch. PMID- 10521238 TI - Gap junctional conductance in cardiomyopathic hamsters: the role of c-Src. PMID- 10521239 TI - Induction of heme oxygenase-1 suppresses venular leukocyte adhesion elicited by oxidative stress: role of bilirubin generated by the enzyme. AB - This study aimed to examine whether an elevated activity of heme oxygenase (HO)-1 in the tissue attenuates endothelial cell-leukocyte interactions microvessels in vivo. When rats were pretreated with an intraperitoneal injection of hemin, an HO 1 inducer, mesenteric tissues, including their microvessels, displayed a marked induction of HO-1 concurrent with an increase in plasma concentrations of bilirubin-IXalpha, the product of HO-catalyzed degradation of protoheme IX. In these rats, oxidative stress such as superfusion with H(2)O(2) and ischemia reperfusion of the tissues neither induced rolling nor exhibited adherent responses of leukocytes in venules. In contrast, the oxidative stresses evoked marked rolling and adhesion of leukocytes in the control rats without HO-1 induction. The HO-1 induction also downregulated leukocyte adhesion elicited by other pro-oxidant stimuli such as N(omega)-nitro-L-arginine methyl ester. The decreases in the oxidant-elicited leukocyte adhesive responses under HO-1 inducing conditions were restored by perfusion with zinc protoporphyrin-IX, an HO inhibitor, but not with copper protoporphyrin-IX, which did not inhibit the enzyme. Furthermore, the effects of zinc protoporphyrin-IX were repressed by superfusion with bilirubin or biliverdin at the micromolar level, but not by the same concentration of carbon monoxide, another product of HO. These results indicate that induction of the HO-1 activity serves as a potential stratagem to prevent oxidant-induced microvascular leukocyte adhesion through the action of bilirubin, a product of HO reaction. PMID- 10521240 TI - Functional role of c-Src in gap junctions of the cardiomyopathic heart. AB - Given the essential role played by gap junctions in the coordination of cardiac muscle contraction, it is plausible that down-regulation of gap junctional conduction is in part responsible for the contractile dysfunction observed in hypertrophied and failing hearts. In the present study, we analyzed the expression and function of the gap junction protein, connexin43, in the ventricular myocardium of hereditary cardiomyopathic, Syrian BIO 14.6 hamsters. Immunoprecipitation and immunoblot analyses revealed that levels of tyrosine phosphorylated connexin43 were increased in BIO 14.6 hamsters at the late stage of congestive heart failure. Furthermore, the increased tyrosine phosphorylation was correlated with increased c-Src activity. The functional consequences of tyrosine phosphorylation of connexin43 in gap junction were assessed using transfected cells expressing constitutively active c-Src. It was found that constitutively active c-Src diminished propagation of Ca(2+) waves in HEK293 cells and reduced gap junctional conductance between pairs of cardiac myocytes. We, therefore, conclude that during the progression of cardiac dysfunction in the cardiomyopathic heart, gap junctional communication is reduced via c-Src-mediated tyrosine phosphorylation of connexin43. PMID- 10521241 TI - Simulated ischemia in flow-adapted endothelial cells leads to generation of reactive oxygen species and cell signaling. AB - We have previously shown that increased reactive oxygen species (ROS) generation occurs with ischemia in the oxygenated lung and have hypothesized that mechanotransduction is the initiating event. In the present study, we developed an in vitro model of oxygenated ischemia by interrupting medium flow to flow adapted bovine pulmonary artery endothelial cells in an artificial capillary system. Cellular oxygenation during the "ischemic" period was maintained by perfusing medium over the abluminal surface of porous capillaries. Cells were assessed for ROS generation, nuclear factor-kappaB (NF-kappaB) and activator protein-1 (AP-1) binding activities, and DNA synthesis using dichlorofluorescein fluorescence by flow cytometry and spectrofluorometry, electrophoretic mobility shift assay of nuclear extracts with NF-kappaB-specific or AP-1-specific (32)P labeled oligonucleotides, and (3)H-thymidine incorporation into DNA. Cells that were flow adapted for 2 to 7 days with 1 to 2 dyne/cm(2) shear stress exhibited a 1.6- to 1.9-fold increase in ROS generation during 1 hour of simulated ischemia compared with continuously perfused cells. This effect was abolished by diphenyleneiodonium chloride (DPI), indicating a role for a flavoprotein such as NADPH oxidase. The increase in ROS generation with ischemia was similar for cells from low and high passages. With ischemia, flow-adapted cells exhibited increases of 1.7-fold in nuclear NF-kappaB and 1.5-fold in nuclear AP-1; these changes were abolished by pretreatment with N-acetylcysteine or DPI. Ischemia for 24 hours resulted in a 1.8-fold increase of (3)H-thymidine incorporation into DNA and a significant increase of cells entering the cell cycle, as indicated by flow cytometry with propidium iodide. We conclude that flow-adapted endothelial cells generate ROS with ischemia that results in activation of NF-kappaB and AP-1 and an increase of DNA synthesis. This effect is not mediated by hypoxia, implicating a role for mechanotransduction in ischemia-mediated cell signaling. PMID- 10521242 TI - Self-protection by cardiac myocytes against hypoxia and hyperoxia. AB - Cardiac muscle must maintain a continuous balance between its energy supply and work performed. An important mechanism involved in achievement of this balance is cross talk via chemical signals between cardiac myocytes and the cardiac muscle vascular system. This has been demonstrated by incubating isolated cardiac myocytes in different concentrations of oxygen and then assaying the conditioned media for vasoactive substances on isolated aortic rings and small-resistance arteries. With increasing oxygen concentrations above 6%, cardiac myocytes produce increasing amounts of angiotensin I, which is converted to angiotensin II by the blood vessel. The angiotensin II stimulates vascular endothelial cells to secrete endothelin and increase vascular tone. Below 6% oxygen, cardiac myocytes secrete adenosine, which acts directly on vascular smooth muscle to block the effect of alpha-adrenergic agonists and reduce vascular tone. In an intact heart, the net effect of these 2 regulatory systems would be the maintenance of oxygen concentration within a narrow range at the cardiac myocytes. By acting as oxygen sensors, cardiac myocytes modulate vascular tone according to the needs of the myocytes and reduce potential problems of hypoxia and extensive formation of reactive oxygen species. PMID- 10521243 TI - Adenosine receptor activation induces vascular endothelial growth factor in human retinal endothelial cells. AB - Adenosine, released in increased amounts by hypoxic tissues, is thought to be an angiogenic factor that links altered cellular metabolism caused by oxygen deprivation to compensatory angiogenesis. Adenosine interacts with 4 subtypes of G protein-coupled receptors, termed A(1), A(2A), A(2B), and A(3). We investigated whether adenosine causes proliferation of human retinal endothelial cells (HRECs) and synthesis of vascular endothelial growth factor (VEGF) and, if so, which adenosine receptor subtype mediates these effects. The nonselective adenosine receptor agonist 5'-N-ethylcarboxamidoadenosine (NECA), in a concentration dependent manner, increased both VEGF mRNA and protein expression by HRECs, as well as proliferation. This proliferative effect of NECA was inhibited by the addition of anti-human VEGF antibody. NECA also increased insulin-like growth factor-I and basic fibroblast growth factor mRNA expression in a time-dependent manner and cAMP accumulation in these cells. In contrast, neither the A(1) agonist N(6)-cyclopentyladenosine nor the A(2A) agonist 2-p-(2-carboxyethyl) phenethylamino-NECA caused any of the above effects of NECA. The effects of NECA were not significantly attenuated by either the A(2A) antagonist SCH58261 or the A(1) antagonist 8-cyclopentyl-1, 3-dipropylxanthine. However, the nonselective adenosine receptor antagonist xanthine amine congener completely inhibited the effects of NECA. Addition of antisense oligonucleotide complementary to A(2B) adenosine receptor mRNA inhibited VEGF protein production by HRECs after NECA stimulation. Thus, the A(2B) adenosine receptor subtype appears to mediate the actions of adenosine to increase growth factor production, cAMP content, and cell proliferation of HRECs. Adenosine activates the A(2B) adenosine receptor in HRECs, which may lead to neovascularization by a mechanism involving increased angiogenic growth factor expression. PMID- 10521244 TI - A novel K(ATP) current in cultured neonatal rat atrial appendage cardiomyocytes. AB - The functional and pharmacological properties of ATP-sensitive K(+) (K(ATP)) channels were studied in primary cultured neonatal rat atrial appendage cardiomyocytes. Activation of a whole-cell inward rectifying K(+) current depended on the pipette ATP concentration and correlated with a membrane hyperpolarization close to the K(+) equilibrium potential. The K(ATP) current could be activated either spontaneously or by a hypotonic stretch of the membrane induced by lowering the osmolality of the bathing solution from 290 to 260 mOsm/kg H(2)O or by the K(+) channel openers diazoxide and cromakalim with EC(50) approximately 1 and 10 nmol/L, respectively. The activated atrial K(ATP) current was highly sensitive to glyburide, with an IC(50) of 1.22+/-0.15 nmol/L. Recorded in inside-out patches, the neonatal atrial K(ATP) channel displayed a conductance of 58.0+/-2.2 pS and opened in bursts of 133.8+/-20.4 ms duration, with an open time duration of 1.40+/-0.10 ms and a close time duration of 0.66+/-0.04 ms for negative potentials. The channel had a half-maximal open probability at 0.1 mmol/L ATP, was activated by 100 micromol/L diazoxide, and was inhibited by glyburide, with an IC(50) in the nanomolar range. Thus, pending further tests at low concentrations of K(ATP) channel openers, the single-channel data confirm the results obtained with whole-cell recordings. The neonatal atrial appendage K(ATP) channel thus shows a unique functional and pharmacological profile resembling the pancreatic beta-cell channel for its high affinity for glyburide and diazoxide and for its conductance, but also resembling the ventricular channel subtype for its high affinity for cromakalim, its burst duration, and its sensitivity to ATP. Reverse transcriptase-polymerase chain reaction experiments showed the expression of Kir6.1, Kir6.2, SUR1A, SUR1B, SUR2A, and SUR2B subunits, a finding supporting the hypothesis that the neonatal atrial K(ATP) channel corresponds to a novel heteromultimeric association of K(ATP) channel subunits. PMID- 10521245 TI - Mechanisms underlying the increase in force and Ca(2+) transient that follow stretch of cardiac muscle: a possible explanation of the Anrep effect. AB - Myocardial stretch produces an increase in developed force (DF) that occurs in two phases: the first (rapidly occurring) is generally attributed to an increase in myofilament calcium responsiveness and the second (gradually developing) to an increase in [Ca(2+)](i). Rat ventricular trabeculae were stretched from approximately 88% to approximately 98% of L(max), and the second force phase was analyzed. Intracellular pH, [Na(+)](i), and Ca(2+) transients were measured by epifluorescence with BCECF-AM, SBFI-AM, and fura-2, respectively. After stretch, DF increased by 1.94+/-0.2 g/mm(2) (P<0.01, n = 4), with the second phase accounting for 28+/-2% of the total increase (P<0.001, n = 4). During this phase, SBFI(340/380) ratio increased from 0.73+/-0.01 to 0.76+/-0.01 (P<0.05, n = 5) with an estimated [Na(+)](i) rise of approximately 6 mmol/L. [Ca(2+)](i) transient, expressed as fura-2(340/380) ratio, increased by 9.2+/-3.6% (P<0.05, n = 5). The increase in [Na(+)](i) was blocked by 5-(N-ethyl-N-isopropyl)-amiloride (EIPA). The second phase in force and the increases in [Na(+)](i) and [Ca(2+)](i) transient were blunted by AT(1) or ET(A) blockade. Our data indicate that the second force phase and the increase in [Ca(2+)](i) transient after stretch result from activation of the Na(+)/H(+) exchanger (NHE) increasing [Na(+)](i) and leading to a secondary increase in [Ca(2+)](i) transient. This reflects an autocrine-paracrine mechanism whereby stretch triggers the release of angiotensin II, which in turn releases endothelin and activates the NHE through ET(A) receptors. PMID- 10521246 TI - Role of Na(+)/H(+) exchanger during ischemia and preconditioning in the isolated rat heart. AB - The role of the Na(+)/H(+) exchanger in ischemia, reperfusion, and preconditioning was investigated in isolated perfused rat hearts. Contractile function, [Na(+)](i), and pH(i) were measured; ischemic damage was assessed by the recovery of developed pressure (DP) on reperfusion. After 30 minutes of ischemia, DP recovered to only 14+/-4% of preischemic control. In contrast, after preconditioning (3x5-minute periods of ischemia) followed by 30 minutes of ischemia, DP recovered to 75+/-4%. Hearts treated with the Na(+)/H(+) exchange inhibitor 5-(N-methyl-N-isobutyl)amiloride (MIA) also showed an enhanced recovery after ischemia (DP 62+/-9%). Treatment with a low concentration of tetrodotoxin (TTX, 100 nmol/L), which blocks the persistent component of the Na(+) current, had a small beneficial effect on recovery (DP 37+/-8%). Thirty minutes of ischemia caused a small [Na(+)](i) rise (3.2+/-0.9 mmol/L); reperfusion resulted in a further [Na(+)](i) increase (+11.9+/-2.5 mmol/L), which partially recovered over 30 minutes. Preconditioning did not change the [Na(+)](i) rise during ischemia but abolished the large [Na(+)](i) rise on reperfusion, and [Na(+)](i) instead fell (-3.6+/-1.3 mmol/L). In the presence of MIA, the [Na(+)](i) rise was unchanged from ischemia only; on reperfusion, [Na(+)](i) fell (-3.7+/-0.9 mmol/L), similar to the preconditioned hearts. TTX abolished the [Na(+)](i) rise during ischemia (+0.3+/-0.7 mmol/L), and the increase on reperfusion was similar to ischemia only. We conclude that the rise of [Na(+)](i) during ischemia is caused by Na(+) entry through persistent Na(+) channels. The rise of [Na(+)](i) on reperfusion is caused by activation of the Na(+)/H(+) exchanger and is blocked by MIA and by preconditioning. It is known that the Na(+)/H(+) exchanger is inhibited during ischemia; the present result suggests that this inhibition is prolonged into the early part of reperfusion by preconditioning. To test this hypothesis, we measured the time course of pH(i) recovery after ischemia and preconditioning. Preconditioning slowed the rate of pH(i) recovery after ischemia, providing further support for the hypothesis that preconditioning inhibits the Na(+)/H(+) exchanger during early reperfusion. This inhibition of the Na(+)/H(+) exchanger during reperfusion prevents Na(+) entry, and therefore Ca(2+) loading, and is part of the protective pathway involved in preconditioning. PMID- 10521247 TI - Activation of mitochondrial ATP-sensitive K(+) channel for cardiac protection against ischemic injury is dependent on protein kinase C activity. AB - Protein kinase C (PKC) is involved in the second messenger signaling cascade during ischemic and Ca(2+) preconditioning. Given that the pharmacological activation of mitochondrial ATP-sensitive K(+) (mitoK(ATP)) channels also mimics preconditioning, the mechanisms linking PKC activation and mitoK(ATP) channels remain to be established. We hypothesize that PKC activity is important for the opening of the mitoK(ATP) channel. To examine this, a specific opener of the mitoK(ATP) channel, diazoxide, was used in conjunction with subcellular distribution of PKC in a model of ischemia/reperfusion (I/R). Langendorff perfused rat hearts were subjected to 40-minute ischemia followed by 30-minute reperfusion. Effects of activation of the mitoK(ATP) channel and other interventions on functional, biochemical, and pathological changes in ischemic hearts were assessed. In hearts treated with diazoxide, left ventricular end diastolic pressure and coronary flow were significantly improved after I/R; lactate dehydrogenase release was also significantly decreased. The morphology was well preserved in diazoxide-treated hearts compared with nontreated ischemic control hearts. The salutary effects of diazoxide on the ischemic injury were similar to those of Ca(2+) preconditioning. Administration of sodium 5 hydroxydecanoate, an effective blocker of the mitoK(ATP) channel, or chelerythrine or calphostin C, an inhibitor of PKC, during diazoxide pretreatment or during continuous presence of diazoxide in the ischemic period, completely abolished the beneficial effects of the diazoxide on the I/R injury. Blockade of Ca(2+) entry during diazoxide treatment by inhibiting the L-type Ca(2+) channel with verapamil also completely reversed the beneficial effect of diazoxide during I/R. PKC-alpha was translocated to sarcolemma, whereas PKC-delta was translocated to the mitochondria and intercalated disc, and PKC-epsilon was translocated to the intercalated disc of the diazoxide-pretreated hearts. Colocalization studies for mitochondrial distribution with tetramethylrhodamine ethyl ester (TMRE) and PKC isoforms by immunoconfocal microscopy revealed that PKC-delta antibody specifically stained the mitochondria. ATP was significantly increased in the diazoxide-treated hearts. Moreover, the data suggest that activation and translocation of PKC to mitochondria appear to be important for the protection mediated by mitoK(ATP) channel. PMID- 10521248 TI - Oxidative stress as a regulator of gene expression in the vasculature. AB - Oxidative stress and the production of intracellular reactive oxygen species (ROS) have been implicated in the pathogenesis of a variety of diseases. In excess, ROS and their byproducts that are capable of causing oxidative damage may be cytotoxic to cells. However, it is now well established that moderate amounts of ROS play a role in signal transduction processes such as cell growth and posttranslational modification of proteins. Oxidants, antioxidants, and other determinants of the intracellular reduction-oxidation (redox) state play an important role in the regulation of gene expression. Recent insights into the etiology and pathogenesis of atherosclerosis suggest that this disease may be viewed as an inflammatory disease linked to an abnormality in oxidation-mediated signals in the vasculature. In this review, we summarize the evidence supporting the notion that oxidative stress and the production of ROS function as physiological regulators of vascular gene expression mediated via specific redox sensitive signal transduction pathways and transcriptional regulatory networks. Elucidating, at the molecular level, the regulatory processes involved in redox sensitive vascular gene expression represents a foundation not only for understanding the pathogenesis of atherosclerosis and other inflammatory diseases but also for the development of novel therapeutic treatment strategies. PMID- 10521249 TI - Protective role of interleukin-10 in atherosclerosis. AB - The potential role of anti-inflammatory cytokines in the modulation of the atherosclerotic process remains unknown. Interleukin (IL)-10 has potent deactivating properties in macrophages and T cells and modulates many cellular processes that may interfere with the development and stability of the atherosclerotic plaque. IL-10 is expressed in human atherosclerosis and is associated with decreased signs of inflammation. In the present study, we show that IL-10-deficient C57BL/6J mice fed an atherogenic diet and raised under specific pathogen-free conditions exhibit a significant 3-fold increase in lipid accumulation compared with wild-type mice. Interestingly, the susceptibility of IL-10-deficient mice to atherosclerosis was exceedingly high (30-fold increase) when the mice were housed under conventional conditions. Atherosclerotic lesions of IL-10-deficient mice showed increased T-cell infiltration, abundant interferon gamma expression, and decreased collagen content. In vivo, transfer of murine IL 10 achieved 60% reduction in lesion size. These results underscore the critical roles of IL-10 in both atherosclerotic lesion formation and stability. Moreover, IL-10 appears to be crucial as a protective factor against the effect of environmental pathogens on atherosclerosis. PMID- 10521250 TI - Molecular analysis of the rhodopsin gene in southern France: identification of the first duplication responsible for retinitis pigmentosa, c.998999ins4. AB - PURPOSE: Mutations in the gene encoding rhodopsin, the visual pigment in rod photoreceptors, were shown to be the most common cause of autosomal retinitis pigmentosa (RP). In order to determine the prevalence of rhodopsin alterations in southern French populations, we examined 52 unrelated patients/families with autosomal dominant RP (adRP=29), RP simplex (6), or unclassified RP (17). METHODS: The full coding and flanking sequences of the rhodopsin (RHO) gene were scanned using an improved DGGE (denaturing gradient gel electrophoresis) assay, followed by sequencing of abnormal fragments. RESULTS: This study revealed three RHO mutations in patients with adRP (G106R, R135W, and c.998999ins4) and a number of frequent or rare polymorphisms. No disease-causing sequence variation was found in simplex and unclassified RP pedigrees. Mutation c.998999ins4 has not been previously reported, and appears as the first duplication identified so far in the RHO gene. This frameshift mutation, which is associated with a severe RP, alters the carboxy terminus and predicts a 353-amino acid mutant rhodopsin instead of 348. DISCUSSION: Our study demonstrates that rhodopsin mutations are responsible for only 10.3% of adRP in French populations living in the Mediterranean area in contrast to the 25-35% reported in other populations. PMID- 10521251 TI - Role of metal ions in the T- to R-allosteric transition in the insulin hexamer. AB - The role of metal ions in the T- to R-allosteric transition is ascertained from the investigation of the T- to R-allosteric transition of transition metal ions substituted-insulin hexamers, as well as from the kinetics of their dissociation. These studies establish that ligand field stabilization energy (LFSE), coordination geometry preference, and the Lewis acidity of the metal ion in the zinc sites modulate the T- to R-state transition. (1)H NMR, (113)Cd NMR, and UV vis measurements demonstrate that, under suitable conditions, Fe2+/3+, Ni2+, and Cd2+ bind insulin to form stable hexamers, which are allosteric species. (1)H NMR R-state signatures are elicited by addition of phenol alone in the case of Ni(II) and Cd(II)-substituted insulin hexamers. The Fe(II)-substituted insulin hexamer is converted to the ferric analogue upon addition of phenol. For the Fe(III) substituted insulin hexamer, appearance of (1)H NMR R-state signatures requires, additionally to phenol, ligands containing a nitrogen that can donate a lone pair of electrons. This is consistent with stabilization of the R-state by heterotropic interactions between the phenol-binding pocket and ligand binding to Fe(III) in the zinc site. UV-vis measurements indicate that the (1)H NMR detected changes in the conformation of the Fe(III)-insulin hexamer are accompanied by a change in the electronic structure of the iron site. Kinetic measurements of the dissociation of the hexamers provide evidence for the modulation of the stability of the hexamer by ligand field stabilization effects. These kinetic studies also demonstrate that the T- to R-state transition in the insulin hexamer is governed by coordination geometry preference of the metal ion in the zinc site and the compatibility between Lewis acidity of the metal ion in the zinc site and the Lewis basicity of the exogenous ligands. Evidence for the alteration of the calcium site has been obtained from (113)Cd NMR measurements. This finding adds to the number of known conformational changes that occur during the T- to R transition and is an important consideration in the formulation of allosteric mechanisms of the insulin hexamer. PMID- 10521252 TI - Amino-terminal modifications of human parathyroid hormone (PTH) selectively alter phospholipase C signaling via the type 1 PTH receptor: implications for design of signal-specific PTH ligands. AB - Parathyroid hormone (PTH) and PTH-related peptide (PTHrP) activate the PTH/PTHrP receptor to trigger parallel increases in adenylyl cyclase (AC) and phospholipase C (PLC). The amino (N)-terminal region of PTH-(1-34) is essential for AC activation. Ligand domains required for activation of PLC, PKC, and other effectors have been less well-defined, although some studies in rodent systems have identified a core region [hPTH-(29-32)] involved in PKC activation. To determine the critical ligand domain(s) for PLC activation, a series of truncated hPTH-(1-34) analogues were assessed using LLC-PK1 cells that stably express abundant transfected human or rat PTH/PTHrP receptors. Phospholipase C signaling and ligand-binding affinity were reduced by carboxyl (C)-terminal truncation of hPTH-(1-34) but were coordinately restored when a binding-enhancing substitution (Glu(19) --> Arg(19)) was placed within hPTH-(1-28), the shortest hPTH peptide that could fully activate both AC and PLC. Phospholipase C, but not AC, activity was reduced by substituting Gly(1) for Ser(1) in hPTH-(1-34) and was eliminated entirely by removing either residue 1 or the alpha-amino group alone. These changes did not alter binding affinity. These findings led to design of an analogue, [Gly(1),Arg(19)]hPTH-(1-28), that was markedly signal-selective, with full AC but no PLC activity. Thus, the extreme N-terminus of hPTH constitutes a critical activation domain for coupling to PLC. The C-terminal region, especially hPTH-(28-31), contributes to PLC activation through effects upon receptor binding but is not required for full PLC activation. The N-terminal determinants of AC and PLC activation in hPTH-(1-34) overlap but are not identical, as subtle modifications in this region may dissociate activation of these two effectors. The [Gly(1),Arg(19)]hPTH-(1-28) analogue, in particular, should prove useful in dissociating AC- from PLC-dependent actions of PTH. PMID- 10521253 TI - Membrane topology of the beta-subunit of the oxaloacetate decarboxylase Na+ pump from Klebsiella pneumoniae. AB - The topology of the beta-subunit of the oxaloacetate Na+ pump (OadB) was probed with the alkaline phosphatase (PhoA) and beta-galactosidase (lacZ) fusion technique. Additional evidence for the topology was derived from amino acid alignments and comparative hydropathy profiles of OadB with related proteins. Consistent results were obtained for the three N-terminal and the six C-terminal membrane-spanning alpha-helices. However, the two additional helices that were predicted by hydropathy analyses between the N-terminal and C-terminal blocks did not conform with the fusion results. The analyses were therefore extended by probing the sideness of various engineered cysteine residues with the membrane impermeant reagent 4-acetamido-4'-maleimidylstilbene-2, 2'-disulfonate. The results were in accord with those of the fusion analyses, suggesting that the protein folds within the membrane by a block of three N-terminal transmembrane segments and another one with six C-terminal transmembrane segments. The mainly hydrophobic connecting segment is predicted not to traverse the membrane fully, but to insert in an undefined manner from the periplasmic face. According to our model, the N-terminus is at the cytoplasmic face and the C-terminus is at the periplasmic face of the membrane. PMID- 10521255 TI - Reaction mechanism of glyoxalase I explored by an X-ray crystallographic analysis of the human enzyme in complex with a transition state analogue. AB - The structures of human glyoxalase I in complexes with S-(N-hydroxy-N-p iodophenylcarbamoyl)glutathione (HIPC-GSH) and S-p nitrobenzyloxycarbonylglutathione (NBC-GSH) have been determined at 2.0 and 1.72 A resolution, respectively. HIPC-GSH is a transition state analogue mimicking the enediolate intermediate that forms along the reaction pathway of glyoxalase I. In the structure, the hydroxycarbamoyl function is directly coordinated to the active site zinc ion. In contrast, the equivalent group in the NBC-GSH complex is approximately 6 A from the metal in a conformation that may resemble the product complex with S-D-lactoylglutathione. In this complex, two water molecules occupy the liganding positions at the zinc ion occupied by the hydroxycarbamoyl function in the enediolate analogue complex. Coordination of the transition state analogue to the metal enables a loop to close down over the active site, relative to its position in the product-like structure, allowing the glycine residue of the glutathione moiety to hydrogen bond with the protein. The structure of the complex with the enediolate analogue supports an "inner sphere mechanism" in which the GSH-methylglyoxal thiohemiacetal substrate is converted to product via a cis-enediolate intermediate. The zinc ion is envisioned to play an electrophilic role in catalysis by directly coordinating this intermediate. In addition, the carboxyl of Glu 172 is proposed to be displaced from the inner coordination sphere of the metal ion during substrate binding, thus allowing this group to facilitate proton transfer between the adjacent carbon atoms of the substrate. This proposal is supported by the observation that in the complex with the enediolate analogue the carboxyl group of Glu 172 is 3.3 A from the metal and is in an ideal position for reprotonation of the transition state intermediate. In contrast, Glu 172 is directly coordinated to the zinc ion in the complexes with S-benzylglutathione and with NBC-GSH. PMID- 10521254 TI - Suppression of ATP diphosphohydrolase/CD39 in human vascular endothelial cells. AB - Vascular ATP diphosphohydrolase/CD39 is an endothelial cell membrane protein with both ecto-ATPase and ecto-ADPase activities. Suppression of constitutive CD39 expression may result in elevated concentrations of ATP and ADP at the vascular interface that could predispose to thrombosis and inflammation. To study the effects of suppression of CD39 synthesis, stable 25-base antisense chimeric oligonucleotides targeting sequences at the 5' region of CD39 were designed. Transfection of these stable oligomers into cultured human endothelial cells resulted in dramatic decreases in levels of CD39 mRNA transcripts. Following transfection with antisense oligonucleotides, total ADPase activity fell from 26.0 +/- 3.1 in control cultures to 9.5 +/- 3.4 nmol of P(i) min(-1) (mg of protein)(-1) (p < 0.005); suppression of CD39 protein expression was also observed by Western blotting. Decreases in ATP diphosphohydrolase activity were associated with increases in concentrations of extracellular purine nucleotides released following stimulation of endothelial cells. Rates of initial hydrolysis of extracellular ATP released from purinergic agonist-stimulated endothelial cells decreased from 17.9 +/- 5.0 to 4.8 +/- 0.5 pmol min(-1) per 10(6) cells (p < 0.005) in antisense transfected cells. Therefore, CD39 regulates extracellular ATP concentrations and may be an important modulator of purinergic receptor activity in vascular endothelial cells. PMID- 10521256 TI - Solution structure of an oligodeoxynucleotide containing the malondialdehyde deoxyguanosine adduct N2-(3-oxo-1-propenyl)-dG (ring-opened M1G) positioned in a (CpG)3 frameshift hotspot of the Salmonella typhimurium hisD3052 gene. AB - The refined solution structure for the ring-opened N2-(3-oxo-1-propenyl)-dG derivative of the malondialdehyde deoxyguanosine adduct M(1)G [3-(2'-deoxy-beta-D erythro-pentofuranosyl)pyrimido[1, 2-a]purin-10(3H)-one] in d(ATCGCXCGGCATG) x d(CATGCCGCGCGAT) [X being N(2)-(3-oxo-1-propenyl)-dG], containing the d(CpG)(3) frameshift hotspot of the Salmonella typhimurium hisD3052 gene, is presented. When inserted into this duplex, M(1)G underwent spontaneous ring opening to N2-(3 oxo-1-propenyl)-dG. NMR analysis revealed that N2-(3-oxo-1-propenyl)-dG induced minor structural perturbations in the hisD3052 oligodeoxynucleotide. However, the stability of the duplex DNA was reduced; the N2-(3-oxo-1-propenyl)-dG-modified hisD3052 oligodeoxynucleotide exhibited a 14 degrees C decrease in T(m) relative to that of the native oligodeoxynucleotide. The modified guanine maintained stacking interactions with neighboring bases but was not Watson-Crick hydrogen bonded. A total of 13 NOEs were observed from the 3-oxo-1-propenyl moiety protons of N2-(3-oxo-1-propenyl)-dG to DNA protons. Molecular dynamics calculations, restrained by 602 distance restraints derived from experimental NOE measurements and 23 empirical distance restraints, converged with pairwise rmsd differences of <0.90 A. The sixth-root residual factor with the NMR data was 9.1 x 10(-2). The cytosine complementary to N2-(3-oxo-1-propenyl)-dG was pushed toward the major groove but maintained partial stacking interactions with its neighboring bases. The modified guanine remained in the anti conformation, while the 3-oxo-1 propenyl moiety was positioned in the minor groove of the duplex. Possible correlations between the relatively small structural perturbations induced in this DNA duplex by N2-(3-oxo-1-propenyl)-dG and the mutagenic spectrum of M(1)G are discussed. PMID- 10521257 TI - Probing the two-gate mechanism of DNA gyrase using cysteine cross-linking. AB - Cross-linking a pair of novel cysteine residues on either side of the bottom dimer interface of DNA gyrase blocks catalytic supercoiling. Limited strand passage is allowed, but release of the transported DNA segment (T segment) via opening of the bottom dimer interface is prevented. In contrast, ATP-independent relaxation of negatively supercoiled DNA is completely abolished, suggesting that T-segment entry via the bottom gate is blocked. These findings support a two-gate model for supercoiling by DNA gyrase and suggest that relaxation by gyrase is the reverse of supercoiling. Cross-linking a truncated version of gyrase (A64(2)B2), which lacks the DNA wrapping domains, does not block ATP-dependent relaxation. This indicates that passage of DNA through the bottom dimer interface is not essential for this reaction. The mechanistic implications of these results are discussed. PMID- 10521258 TI - Atomic resolution structures of the core domain of avian sarcoma virus integrase and its D64N mutant. AB - Six crystal structures of the core domain of integrase (IN) from avian sarcoma virus (ASV) and its active-site derivative containing an Asp64 --> Asn substitution have been solved at atomic resolution ranging 1.02-1.42 A. The high quality data provide new structural information about the active site of the enzyme and clarify previous inconsistencies in the description of this fragment. The very high resolution of the data and excellent quality of the refined models explain the dynamic properties of IN and the multiple conformations of its disordered residues. They also allow an accurate description of the solvent structure and help to locate other molecules bound to the enzyme. A detailed analysis of the flexible active-site region, in particular the loop formed by residues 144-154, suggests conformational changes which may be associated with substrate binding and enzymatic activity. The pH-dependent conformational changes of the active-site loop correlates with the pH vs activity profile observed for ASV IN. PMID- 10521259 TI - Role of conserved Arg40 and Arg117 in the Na+/proline transporter of Escherichia coli. AB - The Na+/proline transporter of Escherichia coli (PutP) is a member of a large family of Na+/solute symporters. To investigate the role of Arg residues which are conserved within this family, Arg40 at the cytoplasmic end of transmembrane domain (TM) II and Arg117 in cytoplasmic loop 4 of PutP are subjected to amino acid substitution analysis. Removal of the positive charge at position 40 (PutP R40C, Q, E) leads to a dramatic decrease of the V(max) of Na(+)-coupled proline uptake (1-10% of PutP-wild-type). The reduced transport rates are accompanied by decreased apparent affinities of the transporter for Na+ and Li+ while the apparent affinity for proline is only slightly altered. Furthermore, single Cys PutP-R40C reacts with N-ethylmaleimide (NEM), and this reaction is partially inhibited by proline and more efficiently by Na+ ions. Remarkably, NEM modification of Cys40 inhibits Na(+)-driven proline uptake almost completely while facilitated influx of proline into deenergized cells is stimulated by this reaction, suggesting an at least partially uncoupled phenotype under these conditions. These results suggest that Arg40 is located close to the site of ion binding and is important for the coupling of ion and proline transport. The observations confirm the functional importance of TM II described in earlier studies [M. Quick and H. Jung (1997) Biochemistry 36, 4631-4636]. In contrast to Arg40, Arg117 is apparently not important for function of the mature protein. The low transport rates observed upon substitution of Arg117 (PutP-R117C, K, Q) can at least partially be attributed to reduced amounts of PutP in the membrane. However, once inserted into the membrane, PutP containing Arg117 replacements shows a stability comparable to the wild-type as indicated by pulse-chase experiments. These observations suggest that Arg117 plays a crucial role at a stage prior to complete functional insertion of PutP into the membrane, i. e., by stabilizing a folding intermediate. PMID- 10521260 TI - Expression, folding, and thermodynamic properties of the bovine oxytocin neurophysin precursor: relationships to the intermolecular oxytocin-neurophysin complex. AB - Earlier thermodynamic studies of the intermolecular interactions between mature oxytocin and neurophysin, and of the effects of these interactions on neurophysin folding, raised questions about the intramolecular interactions of oxytocin with neurophysin within their common precursor. To address this issue, the disulfide rich precursor of oxytocin-associated bovine neurophysin was expressed in Escherichia coli and folded in vitro to yield milligram quantities of purified protein; evidence of significant impediments to yield resulting from damage to Cys residues is presented. The inefficiency associated with the refolding of reduced mature neurophysin in the presence of oxytocin was found not to be alleviated in the precursor. Consistent with this, the effects of pH on the spectroscopic properties of the precursor and on the relative stabilities of the precursor and mature neurophysin to guanidine denaturation indicated that noncovalent intramolecular bonding between oxytocin and neurophysin in the precursor had only a small thermodynamic advantage over the corresponding bonding in the intermolecular complex. Loss of the principal interactions between hormone and protein, and of the enhanced stability of the precursor relative to that of the mature unliganded protein, occurred reversibly upon increasing the pH, with a midpoint at pH 10. Correlation of these results with evidence from NMR studies of structural differences between the precursor and the intermolecular complex, which persist beyond the pH 10 transition, suggests that the covalent attachment of the hormone in the precursor necessitates a conformational change in its neurophysin segment and leads to properties of the system that are distinct from those of either the liganded or unliganded mature protein. PMID- 10521261 TI - Isomerization of all-trans-9- and 13-desmethylretinol by retinal pigment epithelial cells. AB - Photoisomerization of 11-cis-retinal to all-trans-retinal triggers phototransduction in the retinal photoreceptor cells and causes ultimately the sensation of vision. 11-cis-Retinal is enzymatically regenerated through a complex set of reactions in adjacent retinal pigment epithelial cells (RPE). In this study using all-trans-9-desmethylretinol (lacking the C(19) methyl group) and all-trans-13-desmethylretinol (lacking the C(20) methyl group), we explored the effects of C(19) and C(20) methyl group removals on isomerization of these retinols in RPE microsomes. The C(19) methyl group may be involved in the substrate activation, whereas the C(20) methyl group causes steric hindrance with a proton in position C(10) of 11-cis-retinol; thus, removal of this group could accelerate isomerization. We found that all-trans-9-desmethylretinol and all trans-13-desmethylretinol are isomerized to their corresponding 11-cis-alcohols, although with lower efficiencies than isomerization of all-trans-retinol to 11 cis-retinol. These findings make the mechanism of isomerization through the C(19) methyl group unlikely, because in the case of 9-desmethylretinol, the isomerization would have to progress by proton abstraction from electron-rich olefinic C(9). The differences between all-trans-retinol, all-trans-9 desmethylretinol, and all-trans-13-desmethylretinol appear to be a consequence of the enzymatic properties, and binding affinities of the isomerization system, rather than differences in the chemical or thermodynamic properties of these compounds. This observation is also supported by quantum chemical calculations. It appears that both methyl groups are not essential for the isomerization reaction and are not likely involved in formation of a transition stage during the isomerization process. PMID- 10521262 TI - Functional expression and characterization of the wild-type mammalian renal cortex sodium/phosphate cotransporter and an 215R mutant in Saccharomyces cerevisiae. AB - The wild-type and an R215E mutant of the rat renal cortex sodium/phosphate cotransporter type 2 (NaPi-2) were functionally expressed in the yeast Saccharomyces cerevisiae strain MB192, a cell line lacking the high-affinity endogenous H+/P(i) cotransporter. The expression of the mRNA molecules and corresponding proteins was confirmed by Northern and Western blot analysis, respectively. As detected by indirect immunofluorescence and antibody capture assay, both wild-type and mutant NaPi-2 proteins are expressed in the yeast plasma membrane in comparable amounts. In the presence of 5 microM phosphate, Na+ promotes phosphate uptake into yeast cells expressing the wild-type NaPi-2 with a K(0.5) of 5.6 +/- 1.1 mM. The maximum uptake of phosphate (649 +/- 30 pmol/10 min) is approximately 8-fold higher than the uptake obtained with nontransformed cells (76.8 +/- 8 pmol/10 min). Yeast cells expressing the R215E mutant of NaPi-2 accumulate 213 +/- 9 pmol of phosphate/10 min under the same conditions. The K(0.5) for the stimulation of phosphate uptake by Na+ is 4.2 +/- 0.8 mM for the R215E mutant and thus not significantly different from the value obtained with cells expressing the wild-type cotransporter. The reduced level of accumulation of phosphate in yeast cells expressing the R215E mutant is probably due to a reduction of the first-order rate constant k for phosphate uptake: while cells expressing wild-type NaPi-2 accumulate phosphate with a k of 0.06 min(-1), the rate for phosphate uptake into cells expressing the R215E mutant (k) is 0.016 min(-1) and therefore about 4-fold lower. In comparison, the rate for phosphate uptake into nontransformed cells (k) is 0.0075 min(-1). Phosphate uptake into yeast cells that express the wild-type NaPi-2 in the presence of 150 mM NaCl is promoted by extracellular phosphate with a K(0.5) of 45 +/- 4 microM. A phosphate dependent phosphate accumulation is also observed with cells expressing the R215E mutant, but the K(0.5) is twice as high (86 +/- 5 microM) as that obtained with the wild-type cotransporter. We conclude that the yeast expression system is a useful tool for the investigation of structure-function relationships of the renal sodium/phosphate cotransporter and that (215)R, although not involved in Na+ recognition, is a part of the structure involved in phosphate recognition and considerably influences the rate of phosphate uptake by the NaPi-2 cotransporter. PMID- 10521263 TI - The most pathogenic transthyretin variant, L55P, forms amyloid fibrils under acidic conditions and protofilaments under physiological conditions. AB - The L55P transthyretin (TTR) familial amyloid polyneuropathy-associated variant is distinct from the other TTR variants studied to date and the wild-type protein in that the L55P tetramer can dissociate to the monomeric amyloidogenic intermediate and form fibril precursors under physiological conditions (pH 7.0, 37 degrees C). The activation barrier associated with L55P-TTR tetramer dissociation is lower than the barrier for wild-type transthyretin dissociation, which does not form fibrils under physiological conditions. The L55P-TTR tetramer is also very sensitive to acidic conditions, readily dissociating to form the monomeric amyloidogenic intermediate between pH 5.5-5.0 where the wild-type TTR adopts a nonamyloidogenic tetrameric structure. The formation of the L55P monomeric amyloidogenic intermediate involves subtle tertiary structural changes within the beta-sheet rich subunit as discerned from Trp fluorescence, circular dichroism analysis, and ANS binding studies. The assembly of the L55P-TTR amyloidogenic intermediate at physiological pH (pH 7.5) affords protofilaments that elongate with time. TEM studies suggest that the entropic barrier associated with filament assembly (amyloid fibril formation) is high in vitro, amyloid being defined by the laterally assembled four filament structure observed by Blake upon isolation of "fibrils" from the eye of a FAP patient. The L55P-TTR protofilaments formed in vitro bind Congo red and thioflavin T (albeit more weakly than the fibrils produced at acidic pH), suggesting that the structure observed probably represents an amyloid precursor. The structural continuum from misfolded monomer through protofilaments, filaments, and ultimately fibrils must be considered as a possible source of pathology associated with these diseases. PMID- 10521264 TI - Inhibition of cathepsin K by nitric oxide donors: evidence for the formation of mixed disulfides and a sulfenic acid. AB - The cysteine protease cathepsin K is believed to play a key role in bone resorption as it has collagenolytic activity and is expressed predominantly and in high levels in bone resorbing osteoclast cells. The addition of nitric oxide (NO) and NO donors to osteoclasts in vitro results in a reduction of bone resorption, although the mechanism of this effect is not fully understood. The S nitroso derivatives of glutathione (GSNO) and N-acetylpenicillamine (SNAP) and the non-thiol NO donors NOR-1 and NOR-3 all inhibited the activity of purified cathepsin K in a time- and concentration-dependent manner (IC(50) values after 15 min of preincubation at pH 7.5 of 28, 105, 0.4, and 10 microM, respectively). Cathepsin K activity in Chinese hamster ovary cells stably transfected with cathepsin K was also inhibited by the above NO donors with similar potencies. GSNO at 100 microM also completely inhibited the autocatalytic maturation at pH 4.0 of procathepsin K to cathepsin K. The inhibition of cathepsin K by GSNO was rapidly reversed by DTT, but inhibition by NOR-1 was not reversed by DTT, and analysis of the inhibited cathepsin K for S-nitrosylation using the Greiss reaction gave negative results in both cases. Analysis of the protein by electrospray liquid chromatography/mass spectrometry showed that the inhibition of cathepsin K by GSNO resulted in a mass increase of 306 +/- 2 Da, consistent with the formation of a glutathione adduct. Prior inhibition of cathepsin K by the active site thiol-modifying inhibitor E-64 blocked the modification by GSNO, indicating that the glutathione adduct is likely formed at the active site cysteine. Treatment of cathepsin K with NOR-1 resulted in a mass increase of between 30 and 50 Da, corresponding to the oxidation of a cysteine to sulfinic and sulfonic acids. Cotreatment of cathepsin K with NOR-1 plus the sulfenic acid reagent dimedone resulted in a mass increase of approximately 141 Da, which is consistent with the formation of a dimedone adduct. This result demonstrates that the NOR-1-dependent formation of cathepsin K sulfinic and sulfonic acids occurs via a sulfenic acid. These results show that inhibition of cathepsin K activity and its autocatalytic maturation represent two potential mechanisms by which NO can exert its inhibitory effect on bone resorption. This work also shows that oxidative thiol modifications besides S-nitrosylation should be considered when the effects of NO and NO donors on critical thiol-containing proteins are investigated. PMID- 10521265 TI - Partial glycosylation of Asn2181 in human factor V as a cause of molecular and functional heterogeneity. Modulation of glycosylation efficiency by mutagenesis of the consensus sequence for N-linked glycosylation. AB - Coagulation factor V (FV) circulates in two forms, FV1 and FV2, having slightly different molecular masses and phospholipid-binding properties. The aim was to determine whether this heterogeneity is due to the degree of glycosylation of Asn(2181). FVa1 and FVa2 were isolated and digested with endoglycosidase PNGase F. As judged by Western blotting, the FVa2 light chain contained two N-linked carbohydrates, whereas FVa1 contained three. Wild-type FV and three mutants, Asn(2181)Gln, Ser(2183)Thr, and Ser(2183)Ala, were expressed in COS1 cells, activated by thrombin, and analyzed by Western blotting. Wild-type FVa contained the 71 kDa-74 kDa doublet, whereas the Asn(2181)Gln and Ser(2183)Ala mutants contained only the 71 kDa light chain. In contrast, the Ser(2183)Thr mutant gave a 74 kDa light chain. This demonstrated that the third position in the Asn-X Ser/Thr consensus affects glycosylation efficiency, Thr being associated with a higher degree of glycosylation than Ser. The Ser(2183)Thr mutant FVa was functionally indistinguishable from plasma-purified FVa1, whereas Asn(2181)Gln and Ser(2183)Ala mutants behaved like FVa2. Thus, the carbohydrate at Asn(2181) impaired the interaction between FVa and the phospholipid membrane, an interpretation consistent with a structural analysis of a three-dimensional model of the C2 domain and the position of a proposed phospholipid-binding site. In conclusion, we show that the FV1-FV2 heterogeneity is caused by differential glycosylation of Asn(2181) related to the presence of a Ser rather than a Thr at the third position in the consensus sequence of glycosylation. PMID- 10521266 TI - Analysis of the binding of hydroxamic acid and carboxylic acid inhibitors to the stromelysin-1 (matrix metalloproteinase-3) catalytic domain by isothermal titration calorimetry. AB - Matrix metalloproteinases (MMPs) are implicated in diseases such as arthritis and cancer. Among these enzymes, stromelysin-1 can also activate the proenzymes of other MMPs, making it an attractive target for pharmaceutical design. Isothermal titration calorimetry (ITC) was used to analyze the binding of three inhibitors to the stromelysin catalytic domain (SCD). One inhibitor (Galardin) uses a hydroxamic acid group (pK(a) congruent with 8.7) to bind the active site zinc; the others (PD180557 and PD166793) use a carboxylic acid group (pK(a) congruent with 4.7). Binding affinity increased dramatically as the pH was decreased over the range 5.5-7.5. Experiments carried out at pH 6.7 in several different buffers revealed that approximately one and two protons are transferred to the enzyme inhibitor complexes for the hydroxamic and carboxylic acid inhibitors, respectively. This suggests that both classes of inhibitors bind in the protonated state, and that one amino acid residue of the enzyme also becomes protonated upon binding. Similar experiments carried out with the H224N mutant gave strong evidence that this residue is histidine 224. DeltaG, DeltaH, DeltaS, and DeltaC(p) were determined for the three inhibitors at pH 6.7, and DeltaC(p) was used to obtain estimates of the solvational, translational, and conformational components of the entropy term. The results suggest that: (1) a polar group at the P1 position can contribute a large favorable enthalpy, (2) a hydrophobic group at P2' can contribute a favorable entropy of desolvation, and (3) P1' substituents of certain sizes may trigger an entropically unfavorable conformational change in the enzyme upon binding. These findings illustrate the value of complete thermodynamic profiles generated by ITC in discovering binding interactions that might go undetected when relying on binding affinities alone. PMID- 10521267 TI - Zebrafish (Danio rerio) presenilin promotes aberrant amyloid beta-peptide production and requires a critical aspartate residue for its function in amyloidogenesis. AB - Alzheimer's disease (AD) is characterized by the invariable accumulation of senile plaques composed of amyloid beta-peptide (Abeta). Mutations in three genes are known to cause familial Alzheimer's disease (FAD). The mutations occur in the genes encoding the beta-amyloid precursor protein (betaAPP) and presenilin (PS1) and PS2 and cause the increased secretion of the pathologically relevant 42 amino acid Abeta42. We have now cloned the zebrafish (Danio rerio) PS1 homologue (zf PS1) to study its function in amyloidogenesis and to prove the critical requirement of an unusual aspartate residue within the seventh putative transmembrane domain. In situ hybridization and reverse PCR reveal that zf-PS1 is maternally inherited and ubiquitously expressed during embryogenesis, suggesting an essential housekeeping function. zf-PS1 is proteolytically processed to produce a C-terminal fragment (CTF) of approximately 24 kDa similar to human PS proteins. Surprisingly, wt zf-PS1 promotes aberrant Abeta42 secretion like FAD associated human PS1 mutations. The unexpected pathologic activity of wt zf-PS1 may be due to several amino acid exchanges at positions where FAD-associated mutations have been observed. The amyloidogenic function of zf-PS1 depends on the conserved aspartate residue 374 within the seventh putative transmembrane domain. Mutagenizing this critical aspartate residue abolishes endoproteolysis of zf-PS1 and inhibits Abeta secretion in human cells. Inhibition of Abeta secretion is accompanied by the accumulation of C-terminal fragments of betaAPP, suggesting a defect in gamma-secretase activity. These data provide further evidence that PS proteins are directly involved in the proteolytic cleavage of betaAPP and demonstrate that this function is evolutionarily conserved. PMID- 10521268 TI - Synthesis and folding of native collagen III model peptides. AB - Solid-phase synthesis of triple-helical peptides, including native collagen III sequences, was started with a trimeric branch, based upon the lysine dipeptide [Fields, C. G., Mickelson, D. J., Drake, S. L., McCarthy, J. B., and Fields, G. B. (1993) J. Biol. Chem. 268, 14153-14160]. Branch synthesis was modified, using TentaGel R as resin, p-hydroxybenzyl alcohol (HMP) as linker, Dde as N(epsilon) protective group, and HATU/HOAT as coupling reagent. Three homotrimeric sequences, each containing the Gly 606-Gly 618 portion of human type III collagen, were added to the amino functions of the branch. The final incorporation of GlyProHyp triplets, stabilizing the collagen III triple helix, was performed by using protected GlyProHyp tripeptides, each containing tert butylated hydroxyproline [P(tBu)] instead of hydroxyproline (P). Among the protected tripeptides FmocP(tBu)PG, FmocPP(tBu)G, and FmocGPP(tBu), prepared manually on a chlorotrityl resin, incorporation of FmocPP(tBu)Gly was best suited for synthesis of large and stable peptides, such as PPG(8), containing 8 (PPG)(3) trimers (115 residues, 10 610 Da). The structures of five peptides, differing from each other by the type and number of the triplets incorporated, were verified by MALDI-TOF-MS. Their conformations and thermodynamic data were studied by circular dichroism and differential scanning calorimetry. Except for PPG(8), containing 8 (PPG)(3) trimers with hydroxyproline in the X position and adopting a polyproline II structure, all peptides were triple-helical in 0.1 M acetic acid and their thermal stabilities ranged from t(1/2) = 39. 4 to t(1/2) = 62.5 degrees C, depending on the identity and number of the triplets used. Similar values of the van't Hoff enthalpy, DeltaH(vH), derived from melting curves, and the calorimetric enthalpy, DeltaH(cal), obtained from heat capacity curves, indicate a cooperative ratio of CR = DeltaH(vH)/DeltaH(cal) = 1, establishing a two-state process for unfolding of THP(III) peptides. The independence of the transition temperatures t(1/2) on peptide concentration as well as equilibrium centrifugation data indicate monomolecular dimer(f) to dimer(u) (F(2) <--> U(2)) transitions and, in addition, bimolecular dimer(f) to monomer(u) transitions (F(2) <--> 2U). The dominance of the concentration-independent monomolecular reaction over the concentration-dependent bimolecular reaction makes thermal unfolding of THP(III) peptides appear to be monomolecular. If one designates the molecularity described by the term pseudomonomolecular, unfolding of the dimeric peptides PPG(6-8) follows a two-state, pseudomonomolecular reaction. PMID- 10521269 TI - Roles of tyrosine 158 and lysine 165 in the catalytic mechanism of InhA, the enoyl-ACP reductase from Mycobacterium tuberculosis. AB - The role of tyrosine 158 (Y158) and lysine 165 (K165) in the catalytic mechanism of InhA, the enoyl-ACP reductase from Mycobacterium tuberculosis, has been investigated. These residues have been identified as putative catalytic residues on the basis of structural and sequence homology with the short chain alcohol dehydrogenase family of enzymes. Replacement of Y158 with phenylalanine (Y158F) and with alanine (Y158A) results in 24- and 1500-fold decreases in k(cat), respectively, while leaving K(m) for the substrate, trans-2-dodecenoyl-CoA, unaffected. Remarkably, however, replacement of Y158 with serine (Y158S) results in an enzyme with wild-type activity. Kinetic isotope effect studies indicate that the transfer of a solvent-exchangeable proton is partially rate-limiting for the wild-type and Y158S enzymes, but not for the Y158A enzyme. These data indicate that Y158 does not function formally as a proton donor in the reaction but likely functions as an electrophilic catalyst, stabilizing the transition state for hydride transfer by hydrogen bonding to the substrate carbonyl. A conformational change involving rotation of the Y158 side chain upon binding of the enoyl substrate to the enzyme is proposed as an explanation for the inverse solvent isotope effect observed on V/K(DD-CoA) when either NADH or NADD is used as the reductant. These data are consistent with the recently published structure of a C16 fatty acid substrate bound to InhA that shows Y158 hydrogen bonded to the substrate carbonyl group and rotated from the position it occupies in the InhA-NADH binary complex [Rozwarski, D. A., Vilcheze, C., Sugantino, M., Bittman, R., and Sacchettini, J. C. (1999) J. Biol. Chem. 274, 15582-15589]. Finally, the role of K165 has been analyzed using site-directed mutagenesis. Replacement of K165 with glutamine (K165Q) and arginine (K165R) has no effect on the enzyme's catalytic ability or on its ability to bind NADH. However, the K165A and K165M enzymes are unable to bind NADH, indicating that K165 has a primary role in cofactor binding. PMID- 10521270 TI - 8-anilino-1-naphthalene sulfonic acid (ANS) induces folding of acid unfolded cytochrome c to molten globule state as a result of electrostatic interactions. AB - Hydrophobic interaction of 8-anilino-1-naphthalene sulfonic acid (ANS) with proteins is one of the widely used methods for characterizing/detecting partially folded states of proteins. We have carried out a systematic investigation on the effect of ANS, a charged hydrophobic fluorescent dye, on structural properties of acid-unfolded horse heart cytochrome c at pH 2.0 by a combination of optical methods and electrospray ionization mass spectroscopy (ESI MS). ANS was found to induce, a secondary structure similar to native protein and quenching of fluorescence of tryptophan residue, in the acid-unfolded protein. However, the tertiary structure was found to be disrupted thus indicating that ANS stabilizes a molten globule state in acid-unfolded protein. To understand the mechanism of ANS-induced folding of acid-unfolded cytochrome c, comparative ESI MS, soret absorption, and tryptophan fluorescence studies using nile red, a neutral hydrophobic dye, and ANS were carried out. These studies suggested that, at low pH, electrostatic interactions between negatively charged ANS molecules and positively charged amino acid residues present in acid-unfolded cytochrome c are probably responsible for ANS-induced folding of acid-unfolded protein to partially folded compact state or molten globule state. This is the first experimental demonstration of ANS induced folding of unfolded protein and puts to question the usefulness of ANS for characterization/determination of partially folded intermediates of proteins observed under low pH conditions. PMID- 10521271 TI - Molecular mechanisms of interaction of rabbit CAP18 with outer membranes of gram negative bacteria. AB - The mechanism of interaction of the cationic antimicrobial protein (18 kDa), CAP18, with the outer membrane of Gram-negative bacteria was investigated applying transmission electron microscopy and voltage-clamp techniques on artificial planar bilayer membranes. Electron micrographs of bacterial cells exposed to CAP18 showed damage to the outer membrane of the sensitive Escherichia coli strains F515 and ATCC 11775, whereas the membrane of the resistant Proteus mirabilis strain R45 remained intact. Electrical measurements on various planar asymmetric bilayer membranes, one side consisting of a phospholipid mixture and the other of different phospholipids or of lipopolysaccharide (reconstitution model of the outer membrane), yielded information about the influence of CAP18 on membrane integrity. Addition of CAP18 to the side with the varying lipid composition led to lipid-specific adsorption of CAP18 and subsequent induction of current fluctuations due to the formation of transient membrane lesions at a lipid-specific clamp voltage. We propose that the applied clamp voltage leads to reorientation of CAP18 molecules adsorbed to the bilayer into an active transmembrane configuration, allowing the formation of lesions by multimeric clustering. PMID- 10521272 TI - Nonessential role for methionines in the productive association between calmodulin and the plasma membrane Ca-ATPase. AB - To investigate the role of hydrophobic interactions involving methionine side chains in facilitating the productive association between calmodulin (CaM) and the plasma membrane (PM) Ca-ATPase, we have substituted the polar amino acid Gln for Met at multiple positions in both the amino- and carboxyl-terminal domains of CaM. Conformationally sensitive fluorescence signals indicate that these mutations have little effect on the backbone fold of the carboxyl-terminal domain of CaM. The insertion of multiple Gln in either globular domain results in a decrease in the apparent affinity of CaM for the PM-Ca-ATPase. However, despite the multiple substitution of Gln for four methionines at positions 36, 51, 71, and 72 in the amino-terminal domain or for three methionines at positions 124, 144, and 145 in the carboxyl-terminal domain, these mutant CaMs are able to fully activate the PM-Ca-ATPase. Thus, although these CaM mutants have a decreased affinity for the CaM-binding site on the Ca-ATPase, they retain the ability to fully activate the Ca-ATPase at saturating concentrations of CaM. The role of individual methionines in modulating the affinity between the carboxyl terminus and the PM-Ca-ATPase was further investigated through the substitution of individual Met with Gln. Upon substitution of Met(124) and Met(144) with Gln, there is a 5- and 10-fold increase in the amount of CaM necessary to obtain half maximal activation of the PM-Ca-ATPase, indicating that these methionine side chains participate in the high-affinity association between CaM and the PM-Ca ATPase. However, substitution of Gln for Met(145) results in no change in the apparent affinity between CaM and the PM-Ca-ATPase, indicating that in contrast to all other known CaM targets, Met(145) does not participate in the interaction between CaM and the PM-Ca-ATPase. These results emphasize differences in the binding interactions between individual methionines in CaM and different target enzymes, and suggest that hydrophobic interactions between methionines in CaM and the binding site on the PM-Ca-ATPase are not necessary for enzyme activation. Calculation of the binding affinities of individual CaM domains associated with activation of the PM-Ca-ATPase suggests that mutations of methionines located in either domain of CaM can decrease the initial high-affinity association between CaM and the PM-Ca-ATPase, but have little effect upon the subsequent binding of the opposing globular domain. These results suggest that the initial associations between CaM and the CaM-binding sequence in the PM-Ca-ATPase are guided by nonspecific hydrophobic interactions involving both domains of CaM. PMID- 10521273 TI - Functional characterization of a testes-specific alpha-subunit isoform of the sodium/potassium adenosinetriphosphatase. AB - Different isoforms of the sodium/potassium adenosinetriphosphatase (Na,K-ATPase) alpha and beta subunits have been identified in mammals. The association of the various alpha and beta polypeptides results in distinct Na,K-ATPase isozymes with unique enzymatic properties. We studied the function of the Na,K-ATPase alpha4 isoform in Sf-9 cells using recombinant baculoviruses. When alpha4 and the Na pump beta1 subunit are coexpressed in the cells, Na, K-ATPase activity is induced. This activity is reflected by a ouabain-sensitive hydrolysis of ATP, by a Na(+)-dependent, K(+)-sensitive, and ouabain-inhibitable phosphorylation from ATP, and by the ouabain-inhibitable transport of K(+). Furthermore, the activity of alpha4 is inhibited by the P-type ATPase blocker vanadate but not by compounds that inhibit the sarcoplasmic reticulum Ca-ATPase or the gastric H,K-ATPase. The Na,K-ATPase alpha4 isoform is specifically expressed in the testis of the rat. The gonad also expresses the beta1 and beta3 subunits. In insect cells, the alpha4 polypeptide is able to form active complexes with either of these subunits. Characterization of the enzymatic properties of the alpha4beta1 and alpha4beta3 isozymes indicates that both Na,K-ATPases have similar kinetics to Na(+), K(+), ATP, and ouabain. The enzymatic properties of alpha4beta1 and alpha4beta3 are, however, distinct from the other Na pump isozymes. A Na, K ATPase activity with similar properties as the alpha4-containing enzymes was found in rat testis. This Na,K-ATPase activity represents approximately 55% of the total enzyme of the gonad. These results show that the alpha4 polypeptide is a functional isoform of the Na,K-ATPase both in vitro and in the native tissue. PMID- 10521274 TI - Structure-activity relationships in the oxidation of para-substituted benzylamine analogues by recombinant human liver monoamine oxidase A. AB - Monoamine oxidase A (MAO A) plays a central role in the oxidation of amine neurotransmitters. To investigate the structure and mechanism of this enzyme, recombinant human liver MAO A was expressed and purified from Saccharomyces cerevisiae. Anaerobic titrations of the enzyme require only 1 mol of substrate per mole of enzyme-bound flavin for complete reduction. This demonstrates that only one redox-active group (i.e., the covalent FAD cofactor) is involved in catalysis. The reaction rates and binding affinities of 17 para-substituted benzylamine analogues with purified MAO A were determined by steady state and stopped flow kinetic experiments. For each substrate analogue that was tested, the rates of steady state turnover (k(cat)) and anaerobic flavin reduction (k(red)) are similar in value. Deuterium kinetic isotope effects on k(cat), k(red), k(cat)/K(m), and k(red)/K(s) with alpha, alpha-[(2)H]benzylamines are similar for each substrate analogue that was tested and range in value from 6 to 13, indicating that alpha-C-H bond cleavage is rate-limiting in catalysis. Substrate analogue dissociation constants determined from reductive half-reaction experiments as well as from steady state kinetic isotope effect data [Klinman, J. P., and Matthews, R. G. (1985) J. Am. Chem. Soc. 107, 1058-1060] are in excellent agreement. Quantitative structure-activity relationship (QSAR) analysis of dissociation constants shows that the binding of para-substituted benzylamine analogues to MAO A is best correlated with the van der Waals volume of the substituent, with larger substituents binding most tightly. The rate of para substituted benzylamine analogue oxidation and/or substrate analogue-dependent flavin reduction is best correlated with substituent electronic effects (sigma). Separation of the electronic substituent parameter (sigma) into field-inductive and resonance effects provides a more comprehensive treatment of the electronic correlations. The positive correlation of rate with sigma (rho approximately 2.0) suggests negative charge development at the benzyl carbon position occurs and supports proton abstraction as the mode of alpha-C-H bond cleavage. These results are discussed in terms of several mechanisms proposed for MAO catalysis and with previous structure-activity studies published with bovine liver MAO B [Walker, M. C., and Edmondson, D. E. (1994) Biochemistry 33, 7088-7098]. PMID- 10521275 TI - Pre-steady-state kinetic investigation of intermediates in the reaction catalyzed by adenosylcobalamin-dependent glutamate mutase. AB - Glutamate mutase catalyzes the reversible isomerization of L-glutamate to L-threo 3-methylaspartate. Rapid quench experiments have been performed to measure apparent rate constants for several chemical steps in the reaction. The formation of substrate radicals when the enzyme was reacted with either glutamate or methylaspartate was examined by measuring the rate at which 5'-deoxyadenosine was formed, and shown to be sufficiently fast for this step to be kinetically competent. Furthermore, the apparent rate constant for 5'-deoxyadenosine formation was very similar to that measured previously for cleavage of the cobalt carbon bond of adenosylcobalamin by the enzyme, providing further support for a mechanism in which homolysis of the coenzyme is coupled to hydrogen abstraction from the substrate. The pre-steady-state rates of methylaspartate and glutamate formation were also investigated. No burst phase was observed with either substrate, indicating that product release does not limit the rate of catalysis in either direction. For the conversion of glutamate to methylaspartate, a single chemical step appeared to dominate the overall rate, whereas in the reverse direction a lag phase was observed, suggesting the accumulation of an intermediate, tentatively ascribed to glycyl radical and acrylate. The rates of formation and decay of this intermediate were also sufficiently rapid for it to be kinetically competent. When combined with information from previous mechanistic studies, these results allow a qualitative free energy profile to constructed for the reaction catalyzed by glutamate mutase. PMID- 10521276 TI - Structure of the active domain of the herpes simplex virus protein ICP47 in water/sodium dodecyl sulfate solution determined by nuclear magnetic resonance spectroscopy. AB - ICP47 encoded by herpes simplex virus (HSV) is a key factor in the evasion of cellular immune response against HSV-infected cells. By specific inhibition of the transporter associated with antigen processing (TAP), ICP47 prevents peptide transport into the endoplasmic reticulum and subsequent loading of major histocompatibility complex (MHC) class I molecules. Amino acid residues 3-34 have been identified as the active domain. This domain appeared to be unstructured in aqueous solution, whereas after binding to membranes an alpha-helical conformation was observed. Here, we have analyzed the structure of ICP47(2-34) in a lipidlike environment by nuclear magnetic resonance (NMR) spectroscopy. In micellar solution of deuterated sodium dodecyl sulfate, the viral TAP inhibitor adopts an ordered structure. There are two helical regions extending from residues 4 to 15 and from residues 22 to 32. Arg-16 is found on the C-terminus of the first helix, and Gly-33 serves as a terminator of the second helix. A loop between residues 17 and 21 is also evident in the structure. The relative orientation of the helices toward each other, however, could not be determined due to the paucity of NOEs from residues 18-21. PMID- 10521278 TI - Side chain mobility and ligand interactions of the G strand of tear lipocalins by site-directed spin labeling. AB - Side chain mobility, accessibility, and backbone motion were studied by site directed spin labeling of sequential cysteine mutants of the G strand in tear lipocalins (TL). A nitroxide scan between residues 98 and 105 revealed the alternating periodicity of mobility and accessibility to NiEDDA and oxygen, characteristic of a beta-strand. Residue 99 was the most inaccessible to NiEDDA and oxygen. EPR spectra with the fast relaxing agent, K(3)Fe(CN)(6), exhibited two nitroxide populations for most residues. The motionally constrained population was relatively less accessible to K(3)Fe(CN)(6) because of dynamic tertiary contact, probably with side chain residues of adjacent strands. With increasing concentrations of sucrose, the spectral contribution of the immobile component was greater, indicating a larger population with tertiary contact. Increased concentrations of sucrose also resulted in a restriction of mobility of spin-labeled fatty acids which were bound within the TL cavity. The data suggest that sucrose enhanced ligand affinity by slowing the backbone motion of the lipocalin. The correlation time of an MTSL derivative (I) attached to F99C resulted in the lack of side chain motion and therefore reflects the overall rotation of the TL complex. The correlation time of F99C in tears (13.5 ns) was the same as that in buffer and indicates that TL exists as a dimer under native conditions. TL-spin-labeled ligand complexes have a shorter correlation time than the protein alone, indicating that the fatty acids are not rigidly anchored in the cavity, but move within the pocket. This segmental motion of the ligand was modulated by protein backbone fluctuations. Accessibility studies with oxygen and NiEDDA were performed to determine the orientation and depth of a series of fatty acid derivatives in the cavity of TL. Fatty acids are oriented with the hydrocarbon tail buried in the cavity and the carboxyl group oriented toward the mouth. In general, the mobility of the nitroxide varied according to position such that nitroxides near the mouth had greater mobility than those located deep in the cavity. Nitroxides positioned up to 16 carbon units from the hydrocarbon tail of the ligand are motionally restricted and inaccessible, indicating the cavity extends to at least this depth. EPR spectra obtained with and without sucrose showed that the intracavitary position of lauric acid in TL is similar to that in beta-lactoglobulin. However, unlike beta-lactoglobulin, TL binds 16-doxyl stearic acid, suggesting less steric hindrance and greater promiscuity for TL. PMID- 10521277 TI - Resonance Raman studies of cytochrome P450BM3 and its complexes with exogenous ligands. AB - Resonance Raman spectra are reported for both the heme domain and holoenzyme of cytochrome P450BM3 in the resting state and for the ferric NO, ferrous CO, and ferrous NO adducts in the absence and presence of the substrate, palmitate. Comparison of the spectrum of the palmitate-bound form of the heme domain with that of the holoenzyme indicates that the presence of the flavin reductase domain alters the structure of the heme domain in such a way that water accessibility to the distal pocket is greater for the holoenzyme, a result that is consistent with analogous studies of cytochrome P450cam. The data for the exogenous ligand adducts are compared to those previously reported for corresponding derivatives of cytochrome P450cam and document significant and important differences for the two proteins. Specifically, while the binding of substrate induces relatively dramatic changes in the nu(Fe-XY) modes of the ferrous CO, ferric NO, and ferrous NO derivatives of cytochrome P450cam, no significant changes are observed for the corresponding derivatives of cytochrome P450BM3 upon binding of palmitate. In fact, the spectral data for substrate-free cytochrome P450BM3 provide evidence for distortion of the Fe-XY fragment, even in the absence of substrate. This apparent distortion, which is nonexistent in the case of substrate-free cytochrome P450cam, is most reasonably attributed to interaction of the Fe-XY fragment with the F87 phenylalanine side chain. This residue is known to lie very close to the heme iron in the substrate-free derivative of cytochrome P450BM3 and has been suggested to prevent hydroxylation of the terminal, omega, position of long-chain fatty acids. PMID- 10521279 TI - Preparation and characterization of DNA containing a site-specific nonadjacent cyclobutane thymine dimer of the type implicated in UV-induced -1 frameshift mutagenesis. AB - One mechanism for the origin of UV-induced -1 deletion mutations involves the bypass of a nonadjacent cis-syn cyclobutane pyrimidine dimer containing a single intervening nucleotide. To begin to investigate this mechanism, we required a method for obtaining a single, site-specific, nonadjacent dimer. One approach to the preparation of a nonadjacent dimer is to irradiate a DNA duplex containing a centrally located TNT sequence in which the two T's are paired to an AA sequence in an otherwise fully complementary strand. Triplet-sensitized irradiation of the duplex formed between the 13-mer d(GAGTATCTATGAG) and the 12-mer d(CTCATAATACTC) on ice gave a major product that could be reverted to the parent 13-mer by 254 nm irradiation. Proton NMR experiments established the major product to be the nonadjacent cis-syn cyclobutane dimer formed between the two T's of the TCT sequence. Melting temperature studies show that the nonadjacent dimer is more destabilizing to DNA duplex structure than a normal cis-syn dimer and is as stable as the parental bulged DNA duplex. The nonadjacent dimer-containing 13-mer was ligated into a 51-mer and used as a template for primer-extension studies by DNA polymerases. The nonadjacent dimer could not be bypassed by Sequenase Version 2.0 and terminated synthesis primarily prior to and opposite the 3'-T of the dimer. In contrast, approximately 30% of the dimer was bypassed by an exonuclease deficient (exo-) Klenow fragment, and termination occurred primarily opposite the 3'- and 5'-T's of the dimer. Bypass of the nonadjacent dimer by exo(-) Klenow fragment led primarily to a single-nucleotide deletion mutation as well as small amounts of a full-length product and a four-nucleotide deletion that could be explained by a primer misalignment mechanism. PMID- 10521280 TI - tRNA discrimination at the binding step by a class II aminoacyl-tRNA synthetase. AB - Aminoacyl-tRNA synthetases preserve the fidelity of decoding genetic information by accurately joining amino acids to their cognate transfer RNAs. Here, tRNA discrimination at the level of binding by Escherichia coli histidyl-tRNA synthetase is addressed by filter binding, analytical ultracentrifugation, and iodine footprinting experiments. Competitive filter binding assays show that the presence of an adenylate analogue 5'-O-[N-(L-histidyl)sulfamoyl]adenosine, HSA, decreased the apparent dissociation constant (K(D)) for cognate tRNA(His) by more than 3-fold (from 3.87 to 1.17 microM), and doubled the apparent K(D) for noncognate tRNA(Phe) (from 7.3 to 14.5 microM). By contrast, no binding discrimination against mutant U73 tRNA(His) was observed, even in the presence of HSA. Additional filter binding studies showed tighter binding of both cognate and noncognate tRNAs by G405D mutant HisRS [Yan, W., Augustine, J., and Francklyn, C. (1996) Biochemistry 35, 6559], which possesses a single amino acid change in the C-terminal anticodon binding domain. Discrimination against noncognate tRNA was also observed in sedimentation velocity experiments, which showed that a stable complex was formed with the cognate tRNA(His) but not with noncognate tRNA(Phe). Footprinting experiments on wild-type versus G405D HisRS revealed characteristic alterations in the pattern of protection and enhancement of iodine cleavage at phosphates 5' to tRNA nucleotides in the anticodon and hinge regions. Together, these results suggest that the anticodon and core regions play major roles in the initial binding discrimination between cognate and noncognate tRNAs, whereas acceptor stem nucleotides, particularly at position 73, influence the reaction at steps after binding of tRNA. PMID- 10521281 TI - The solution structure of photosystem I accessory protein E from the cyanobacterium Nostoc sp. strain PCC 8009. AB - PsaE is a small basic subunit located on the stromal (cytoplasmic) side of photosystem I. In cyanobacteria, this subunit participates in cyclic electron transport and modulates the interactions of the complex with soluble ferredoxin. The PsaE protein isolated from the cyanobacterium Synechococcus sp. strain PCC 7002 adopts the beta topology of an SH3 domain, with five beta strands (betaA through betaE) and a turn of 3(10) helix between strands betaD and betaE [Falzone, C. J., Kao, Y.-H., Zhao, J., Bryant, D. A., and Lecomte, J. T. J. (1994) Biochemistry 33, 6052-6062]. The primary structure of the PsaE protein is strongly conserved across all oxygen-evolving photosynthetic organisms. However, variability in loop lengths, as well as N- or C-terminal extensions, suggests that the structure of a second representative PsaE subunit would be useful to characterize the interactions among photosystem I polypeptides. In this work, the solution structure of PsaE from the cyanobacterium Nostoc sp. strain PCC 8009 was determined by NMR methods. Compared to PsaE from Synechococcus sp. strain PCC 7002, this PsaE has a seven-residue deletion in the loop connecting strands betaC and betaD, and an eight-residue C-terminal extension. Angular and distance restraints derived from homonuclear and heteronuclear NMR experiments were used to calculate structures by a distance-geometry/simulated-annealing protocol. A family of 20 structures (rmsd of 0.24 A in the regular secondary structure) is presented. Differences between the two cyanobacterial proteins are mostly confined to the CD loop region; the C-terminal extension is disordered. The thermodynamic stability of Nostoc sp. strain PCC 8009 PsaE toward urea denaturation was measured by circular dichroism and fluorescence spectroscopy, and thermal denaturation was monitored by UV absorption spectroscopy. Chemical and thermal denaturation curves are modeled satisfactorily with two-state processes. The DeltaG degrees of unfolding at room temperature is 12.4 +/- 0.3 kJ mol(-1) (pH 5), and the thermal transition midpoint is 59 +/- 1 degrees C (pH 7). Interactions with other proteins in the photosystem I complex may aid in maintaining PsaE in its native state under physiological conditions. PMID- 10521282 TI - DNA triple helix formation at target sites containing several pyrimidine interruptions: stabilization by protonated cytosine or 5-(1-propargylamino)dU. AB - DNase I footprinting has been used to study the formation of parallel triplexes at oligopurine target sequences which are interrupted by pyrimidines at regular intervals. TA interruptions are targeted with third strand oligonucleotides containing guanine, generating G x TA triplets, while CG base pairs are targeted with thymine, forming T x CG triplets. We have attempted to optimize the stability of these complexes by varying the base composition and sequence arrangement of the target sites, and by replacing the third strand thymines with the positively charged analogue 5-(1-propargylamino)dU (U(P)). For the target sequence (AAAT)(5)AA, in which pyrimidines are positioned at every fourth residue, triplex formation with TG-containing oligonucleotides is only detected in the presence of a triplex-binding ligand, though stable triplexes were detected at the target site (AAAAAT)(3)AAAA. Triplex stability at targets containing pyrimidines at every fourth residue is increased by introducing guanines into the duplex repeat unit using the targets (AGAT)(5)AA and (ATGA)(5)AA. In contrast, placing C(+) x GC triplets on the 5'-side of G x TA, using the target (AGTA)(5)TT, produces complexes of lower stability. We have attempted further to increase the stability of these complexes by using the positively charged thymine base analogue U(P), and have shown that (TU(P)TG)(5)TT forms a more stable complex with target (AAAT)(5)AA than the unmodified third strand, generating a footprint in the absence of a triplex-binding ligand. Triplex formation at (AGTA)(5)AA is improved by using the modified oligonucleotide (TCGU(P))(5)TT, generating a complex in which the charged triplets C(+) x GC and U(P) x AT alternate with uncharged triplets. In contrast, placing U(P) x AT triplets adjacent to C(+) x GC, using the third strand oligonucleotide (U(P)CGT)(5)TT, reduces triplex formation, while the third strand with both substitutions, (U(P)CGU(P))(5)TT, produces a complex with intermediate stability. It appears that, although adjacent U(P) x AT triplets form stable triplexes, placing U(P) x AT adjacent to C(+) x GC is unfavorable. Similar results were obtained with fragments containing CG inversions within the oligopurine tract, though triplexes at (AAAAAC)(3)AA were only detected in the presence of a triplex-binding ligand. Placing C(+) x GC on the 5'-side of T x CG triplets also reduces triplex formation, while a 3'-C(+) x GC produces complexes with increased stability. PMID- 10521283 TI - On the stator of rotary ATP synthase: the binding strength of subunit delta to (alpha beta)3 as determined by fluorescence correlation spectroscopy. AB - ATP synthase is conceived as a rotary enzyme. Proton flow drives the rotor (namely, subunits c12 epsilon gamma) relative to the stator (namely, subunits ab2 delta(alpha beta)3) and extrudes spontaneously formed ATP from three symmetrically arranged binding sites on (alpha beta)3 into the solution. We asked whether the binding of subunit delta to (alpha beta)3 is of sufficient strength to hold against the elastic strain, which is generated during the operation of this enzyme. According to current estimates, the elastically stored energy is about 50 kJ/mol. Subunit delta was specifically labeled without impairing its function. Its association with solubilized (alpha beta)3 gamma in detergent-free buffer was studied by fluorescence correlation spectroscopy (FCS). A very strong tendency of delta to dimerize in detergent-free buffer was apparent (K(d) C transition that leads to an I1061T substitution in three patients. The mutation, located in exon 21, affects a putative transmembrane domain of the protein. PCR-based tests with genomic DNA were used to survey 115 unrelated patients from around the world with all known clinical and biochemical phenotypes of the disease. The I1061T allele constituted 33 (14.3%) of the 230 disease-causing alleles and was never found in controls (>200 alleles). The mutation was particularly frequent in patients with NPC from Western Europe, especially France (11/62 alleles) and the United Kingdom (9/32 alleles), and in Hispanic patients whose roots were in the Upper Rio Grande valley of the United States. The I1061T mutation originated in Europe and the high frequency in northern Rio Grande Hispanics results from a founder effect. All seven unrelated patients who were homozygous for the mutation and their seven affected siblings had a juvenile-onset neurological disease and severe alterations of intracellular LDL-cholesterol processing. The mutation was not found (0/40 alleles) in patients with the severe infantile neurological form of the disease. Testing for this mutation therefore has important implications for genetic counseling of families affected by NPC. PMID- 10521298 TI - High frequency of large intragenic deletions in the Fanconi anemia group A gene. AB - Fanconi anemia (FA) is an autosomal recessive disorder exhibiting chromosomal fragility, bone-marrow failure, congenital abnormalities, and cancer. At least eight complementation groups have been described, with group A accounting for 60% 65% of FA patients. Mutation screening of the group A gene (FANCA) is complicated by its highly interrupted genomic structure and heterogeneous mutation spectrum. Recent reports of several large deletions of FANCA, coupled with modest mutation detection rates, led us to investigate whether many deletions might occur in the heterozygous state and thus fail to be detected by current screening protocols. We used a two-step screening strategy, in which small mutations were detected by fluorescent chemical cleavage of the FANCA transcript, and heterozygosity for gross deletions was detected by quantitative fluorescent multiplex PCR. We screened 26 cell lines from FA complementation group A for FANCA mutations and detected 33 different mutations, 23 of which were novel. Mutations were observed in all 26 cell lines and included 43 of a possible 52 mutant alleles (83%). Of the mutant alleles, 40% were large intragenic deletions that removed up to 31 exons from the gene, indicating that this may be the most prevalent form of mutation in FANCA. Several common deletion breakpoints were observed, and there was a highly significant correlation between the number of breakpoints detected in a given intron and the number of Alu repeats that it contained, which suggests that Alu-mediated recombination may explain the high prevalence of deletions in FANCA. The dual screening strategy that we describe may be useful for mutation screening in other genetic disorders in which mutation-detection rates are unexpectedly low. PMID- 10521299 TI - Constitutional mutations of the hSNF5/INI1 gene predispose to a variety of cancers. AB - Biallelic, truncating mutations of the hSNF5/INI1 gene have recently been documented in malignant rhabdoid tumor (MRT), one of the most aggressive human cancers. This finding suggests that hSNF5/INI1 is a new tumor-suppressor gene for which germline mutations might predispose to cancer. We now report the presence of loss-of-function mutations of this gene in the constitutional DNA from affected members but not from healthy relatives in cancer-prone families. Furthermore, a constitutional mutation is documented in a patient with two successive primary cancers. In agreement with the two-hit model, the wild-type hSNF5/INI1 allele is deleted in the tumor DNA from mutation carriers. In all tested cases, DNA from parents demonstrated normal hSNF5/INI1 sequences, therefore indicating the de novo occurrence of the mutation, which was shown to involve the maternal allele in one case and the paternal allele in two other cases. These data indicate that constitutional mutation of the hSNF5/INI1 gene defines a new hereditary syndrome predisposing to renal or extrarenal MRT and to a variety of tumors of the CNS, including choroid plexus carcinoma, medulloblastoma, and central primitive neuroectodermal tumor. This condition, which we propose to term "rhabdoid predisposition syndrome," may account for previous observations of familial and multifocal cases of the aforementioned tumor types. It could also provide the molecular basis for cases of Li-Fraumeni syndrome without p53 germline mutations. PMID- 10521300 TI - The A1555G mutation in the 12S rRNA gene of human mtDNA: recurrent origins and founder events in families affected by sensorineural deafness. AB - The mtDNA variation of 50 Spanish and 4 Cuban families affected by nonsyndromic sensorineural deafness due to the A1555G mutation in the 12S rRNA gene was studied by high-resolution RFLP analysis and sequencing of the control region. Phylogenetic analyses of haplotypes and detailed survey of population controls revealed that the A1555G mutation can be attributed to >/=30 independent mutational events among the 50 Spanish families and that it occurs on mtDNA haplogroups that are common in all European populations. This indicates that the relatively high detection rate of this mutation in Spain is not due to sampling biases or to a single major founder event. Moreover, the distribution of these mutational events on different haplogroups is compatible with a random occurrence of the A1555G mutation and tends to support the conclusion that mtDNA backgrounds do not play a significant role in the expression of the mutation. Overall, these findings appear to indicate that the rare detection of this mutation in other populations is most likely due to inadequacy in patient ascertainment and molecular screening. This probable lack of identification of the A1555G mutation in subjects affected by sensorineural hearing loss implies that their maternally related relatives are not benefiting from presymptomatic detection and information concerning their increased risk of ototoxicity due to aminoglycoside treatments. PMID- 10521301 TI - Methylation imprinting of H19 and SNRPN genes in human benign ovarian teratomas. AB - In humans, studies of female germ cells are very limited by ethics. The current study investigated the usefulness of benign ovarian teratomas as a substitute for ova in analyses of imprinted genes. Twenty-five human benign ovarian teratomas were typed with 45 microsatellite DNA markers and classified according to their genotypic features. Two oppositely imprinted genes, H19 and SNRPN, were then chosen for analysis of their methylation states in these tumors. These analyses revealed that benign ovarian teratomas consist of a mixture of genetically and epigenetically heterogeneous cell populations. In contrast to previous reports, we could document only one case rising from germ cells by meiosis-II nondisjunction. H19 and SNRPN were methylated in individual teratomas to various degrees, ranging from normal somatic cell to expected ovum levels. The allele with residual methylation of H19 was consistent with that methylated in the patient's blood DNA, thus being of paternal origin. Degrees of H19 hypomethylation and SNRPN hypermethylation increased as the cellular origin of the tumors advanced in oogenesis and were closely correlated in individual teratomas. These results could be best explained by the assumption that the primary imprinting is a progressively organized process and suggest that the establishment of primary imprints on different genes might be mechanistically linked, even when those genes are oppositely imprinted. PMID- 10521302 TI - Transfection of BLM into cultured bloom syndrome cells reduces the sister chromatid exchange rate toward normal. AB - The gene BLM, mutated in Bloom syndrome (BS), encodes the nuclear protein BLM, which when absent, as it is from most BS cells, results in genomic instability. A manifestation of this instability is an excessive rate of sister-chromatid exchange (SCE). Here we describe the effects on this abnormal cellular phenotype of stable transfection of normal BLM cDNAs into two types of BS cells, SV40 transformed fibroblasts and Epstein-Barr virus (EBV)-transformed lymphoblastoid cells. Clones of BLM-transfected fibroblasts produced normal amounts of BLM by western blot analysis and displayed a normal nuclear localization of the protein by immunofluorescence microscopy. They had a mean of 24 SCEs/46 chromosomes, in contrast to the mean of 69 SCEs in controls transfected only with the vector. BLM transfected fibroblast clones that expressed highest levels of the BLM protein had lowest levels of SCE. The lymphoblastoid cells transfected with BLM had SCE frequencies of 22 and 42 in two separate experiments in which two different selectable markers were used, in contrast to 57 and 58 in vector-transfected cells; in this type cell, however, the BLM protein was below the level detectable by western blot analysis. These experiments prove that BLM cDNA encodes a functional protein capable of restoring to or toward normal the uniquely characteristic high-SCE phenotype of BS cells. PMID- 10521304 TI - Chromosome breakage hotspots and delineation of the critical region for the 9p deletion syndrome. AB - The clinical features of the 9p-deletion syndrome include dysmorphic facial features (trigonocephaly, midface hypoplasia, upward-slanting palpebral fissures, and a long philtrum) and mental retardation. The majority of these patients appear to have similar cytogenetic breakpoints in 9p22, but some cases show phenotypic heterogeneity. To define the breakpoints of the deleted chromosomes, we studied 24 patients with a deletion of 9p, by high-resolution cytogenetics, FISH with 19 YACs, and PCR using 25 different sequence-tagged sites. Of 10 different breakpoints identified, 9 were localized within an approximately 5-Mb region, in 9p22-p23, that encompasses the interval between D9S1869 (telomeric) and D9S162 (centromeric). Eight unrelated patients had a breakpoint (group 1) in the same interval, between D9S274 (948h1) and D9S285 (767f2), suggesting a chromosome-breakage hotspot. Among 12 patients, seven different breakpoints (groups 3-9) were localized to a 2-Mb genomic region between D9S1709 and D9S162, which identified a breakpoint-cluster region. The critical region for the 9p deletion syndrome maps to a 4-6-Mb region in 9p22-p23. The results from this study have provided insight into both the heterogeneous nature of the breakage in this deletion syndrome and the resultant phenotype-karyotype correlations. PMID- 10521303 TI - Hypomethylation of an expanded FMR1 allele is not associated with a global DNA methylation defect. AB - The vast majority of fragile-X full mutations are heavily methylated throughout the expanded CGG repeat and the surrounding CpG island. Hypermethylation initiates and/or stabilizes transcriptional inactivation of the FMR1 gene, which causes the fragile X-syndrome phenotype characterized, primarily, by mental retardation. The relation between repeat expansion and hypermethylation is not well understood nor is it absolute, as demonstrated by the identification of nonretarded males who carry hypomethylated full mutations. To better characterize the methylation pattern in a patient who carries a hypomethylated full mutation of approximately 60-700 repeats, we have evaluated methylation with the McrBC endonuclease, which allows analysis of numerous sites in the FMR1 CpG island, including those located within the CGG repeat. We report that the expanded-repeat region is completely free of methylation in this full-mutation male. Significantly, this lack of methylation appears to be specific to the expanded FMR1 CGG-repeat region, because various linked and unlinked repetitive-element loci are methylated normally. This finding demonstrates that the lack of methylation in the expanded CGG-repeat region is not associated with a global defect in methylation of highly repeated DNA sequences. We also report that de novo methylation of the expanded CGG-repeat region does not occur when it is moved via microcell-mediated chromosome transfer into a de novo methylation competent mouse embryonal carcinoma cell line. PMID- 10521305 TI - Identification of a new locus for generalized epilepsy with febrile seizures plus (GEFS+) on chromosome 2q24-q33. AB - We report the identification of a new locus for generalized epilepsy with febrile seizures plus (GEFS+). Six family members manifested isolated typical febrile seizures (FS), and five had typical FS associated with generalized epilepsy (FS+, generalized tonic/clonic seizures). Afebrile seizures occurred from childhood until the teenage years. The maximum two-point LOD score was 3.99 for markers D2S294 and D2S2314. Flanking markers place the GEFS+ locus between D2S141 and D2S116, with multipoint analysis favoring the 13-cM interval spanned by D2S294 and D2S364. This locus is the second GEFS+ locus to be reported, which suggests that this syndrome is genetically heterogeneous. PMID- 10521306 TI - An autosomal dominant thrombocytopenia gene maps to chromosomal region 10p. AB - The increasing number of diagnosed cases of inherited thrombocytopenias, owing to the routine practice of including platelet counts in blood tests, suggests that this condition is not so rare as expected. In the majority of cases, the molecular basis of the disease is unknown, although the defect is likely to affect thrombocytopoiesis and regulation of the normal platelet count. Here we report a genomewide search in a large Italian family affected by autosomal dominant thrombocytopenia. Patients showed a moderate thrombocytopenia with minimal symptoms characterized by normocellular bone marrow, normal medium platelet volume, and positive aggregation tests. Microsatellite analysis demonstrated that the disease locus (THC2) is linked to chromosome 10p11.1-12, within a candidate region of 6 cM between markers D10S586 and D19S1639. A maximum LOD score of 8.12 at recombination fraction.00 was obtained with the microsatellite D10S588. These data localized the first locus of an autosomal dominant thrombocytopenia, and the subsequent identification of the gene will provide new insight into the basic mechanism of megakaryocytopoiesis disorders. PMID- 10521307 TI - A new neurological syndrome with mental retardation, choreoathetosis, and abnormal behavior maps to chromosome Xp11. AB - Choreoathetosis is a major clinical feature in only a small number of hereditary neurological disorders. We define a new X-linked syndrome with a unique clinical picture characterized by mild mental retardation, choreoathetosis, and abnormal behavior. We mapped the disease in a four-generation pedigree to chromosome Xp11 by linkage analysis and defined a candidate region containing a number of genes possibly involved in neuronal signaling, including a potassium channel gene and a neuronal G protein-coupled receptor. PMID- 10521308 TI - Pseudoautosomal linkage of Hodgkin disease. AB - Heritable factors appear to account for much of the risk for Hodgkin disease (HD). There is evidence for an HLA-linked gene, but other predisposing loci remain unaccounted for. The observation of a family coinheriting both HD and Leri Weill dyschondrosteosis (LWD) suggests that a gene conferring risk for HD resides adjacent to the LWD locus. The gene responsible for LWD, SHOX, localizes to the short-arm pseudoautosomal region (PAR) of the X and Y chromosomes. A unique segregation pattern for PAR-linked genes has been predicted-that affected sibs will tend to be same sex. An excess of sex-concordant affected sib pairs with HD has been noted but has been attributed to an environmental etiology. These two observations-sex concordance in sib pairs with HD and cosegregation of HD and LWD impelled a test of the hypothesis that there is a PAR-localized gene for HD. By first scoring recombinations dissociating sex from phenotype in individuals from pedigrees with LWD, we determined a male maximum recombination frequency (thetamax) of.405. This places SHOX near the short-arm telomeres of the sex chromosome and supports the prediction that PAR recombination is obligatory for spermatogenesis. By inferring recombinations between HD and sexual phenotype in sib pairs, we predict, for the postulated HD gene, a male thetamax as high as .254, which places it in proximity to SHOX. Morton's nonparametric affected-sib pair "beta" model was used in the evaluation of linkage between HD and phenotypic sex and gave a LOD score of 2.41. Using this approach, we reevaluated evidence for HLA linkage in HD in haplotyped sib pairs and found a LOD score of 2.00. The resulting beta values indicate that the putative PAR- and HLA-linked loci account for 29% and 40%, respectively, of the heritability of HD in an American population. PMID- 10521309 TI - Interindividual variation in mitotic recombination. AB - Mitotic recombination (MR) between homologous chromosomes is a mutational event that results in loss of heterozygosity in half of the segregants at mitosis. Loss of heterozygosity may have important biological consequences. The purpose of this study was to describe human variation in the spontaneous frequency of MR. Using an immunoselection technique for isolating the somatic mutations that result in loss of expression of one of the codominant alleles at the HLA-A locus, we have measured the frequency and molecular basis of somatic mutations in lymphocytes from a population of young adults. Mutations were classified as being the result of intragenic changes, major deletions, or MR. Here we show that the MR mutation frequency in females was significantly greater than that in males but that intragenic mutation frequency showed no association with sex. Individual variation in MR frequency ranged over more than two orders of magnitude and was not normally distributed. Furthermore, the observed number of individuals from whom no mutants resulting from MR were obtained was significantly greater than was expected. The endogenous level of MR may be under genetic control. Given the association of loss of heterozygosity with cancer initiation and progression, low endogenous MR may confer a reduced lifetime risk of cancer, and the converse may apply. PMID- 10521310 TI - A complete genome screen in sib pairs affected by Gilles de la Tourette syndrome. The Tourette Syndrome Association International Consortium for Genetics. AB - Gilles de la Tourette syndrome is a neuropsychiatric disorder characterized by waxing and waning multiple motor and phonic tics with a complex mode of inheritance. Previous attempts, which used large multigenerational families to localize susceptibility loci, have been unsuccessful. In this report, the results of the first systematic genome scan, using 76 affected-sib-pair families with a total of 110 sib pairs, are summarized. While no results reached acceptable statistical significance, the multipoint maximum-likelihood scores (MLS) for two regions (4q and 8p) were suggestive (MLS > 2.0). Four additional genomic regions also gave multipoint MLS scores between 1.0 and 2.0. PMID- 10521312 TI - On a randomization procedure in linkage analysis. AB - Although much theoretical work has been undertaken to derive thresholds for statistical significance in genetic linkage studies, real data are often complicated by many factors, such as missing individuals or uninformative markers, which make the validity of these theoretical results questionable. Many simulation-based methods have been proposed in the literature to determine empirically the statistical significance of the observed test statistics. However, these methods either are not generally applicable to complex pedigree structures or are too time-consuming. In this article, we propose a computationally efficient simulation procedure that is applicable to arbitrary pedigree structures. This procedure can be combined with statistical tests, to assess the statistical significance for genetic linkage between a locus and a qualitative or quantitative trait. Furthermore, the genomewide significance level can be appropriately controlled when many linked markers are studied in a genomewide scan. Simulated data and a diabetes data set are analyzed to demonstrate the usefulness of this novel simulation method. PMID- 10521311 TI - Recent male-mediated gene flow over a linguistic barrier in Iberia, suggested by analysis of a Y-chromosomal DNA polymorphism. AB - We have examined the worldwide distribution of a Y-chromosomal base-substitution polymorphism, the T/C transition at SRY-2627, where the T allele defines haplogroup 22; sequencing of primate homologues shows that the ancestral state cannot be determined unambiguously but is probably the C allele. Of 1,191 human Y chromosomes analyzed, 33 belong to haplogroup 22. Twenty-nine come from Iberia, and the highest frequencies are in Basques (11%; n=117) and Catalans (22%; n=32). Microsatellite and minisatellite (MSY1) diversity analysis shows that non-Iberian haplogroup-22 chromosomes are not significantly different from Iberian ones. The simplest interpretation of these data is that haplogroup 22 arose in Iberia and that non-Iberian cases reflect Iberian emigrants. Several different methods were used to date the origin of the polymorphism: microsatellite data gave ages of 1,650, 2,700, 3,100, or 3,450 years, and MSY1 gave ages of 1,000, 2,300, or 2,650 years, although 95% confidence intervals on all of these figures are wide. The age of the split between Basque and Catalan haplogroup-22 chromosomes was calculated as only 20% of the age of the lineage as a whole. This study thus provides evidence for direct or indirect gene flow over the substantial linguistic barrier between the Indo-European and non-Indo-European-speaking populations of the Catalans and the Basques, during the past few thousand years. PMID- 10521313 TI - About the "Pathological" role of the mtDNA T3308C mutationellipsis. PMID- 10521314 TI - Diaphragmatic spinal muscular atrophy with respiratory distress is heterogeneous, and one form Is linked to chromosome 11q13-q21. PMID- 10521315 TI - Further evidence for a susceptibility locus on chromosome 20q13.11 in families with dominant transmission of Graves disease. PMID- 10521316 TI - Primary autosomal recessive microcephaly: homozygosity mapping of MCPH4 to chromosome 15. PMID- 10521317 TI - Association of RET protooncogene codon 45 polymorphism with Hirschsprung disease. PMID- 10521318 TI - The sex ratio in familial persistent stuttering. PMID- 10521320 TI - Embryonic temperature affects metabolic compensation and thyroid hormones in hatchling snapping turtles. AB - Temperature acclimation of adult vertebrates typically induces changes in metabolic physiology. During early development, such metabolic compensation might have profound consequences, yet acclimation of metabolism is little studied in early life stages. We measured the effect of egg incubation temperature on resting metabolic rate (RMR) and blood thyroid hormone levels of hatchling snapping turtles (Chelydra serpentina). Like many reptiles, snapping turtles have temperature-dependent sex determination (TSD), in which embryonic temperature determines sex. Therefore, we designed the experiments to separately measure effects of temperature and of sex on the response variables. We incubated eggs in the laboratory at 21. 5 degrees, 24.5 degrees, 27.5 degrees, and 30.5 degrees C, producing both sexes, all males, both sexes, and all females, respectively. Hatchling RMR, when measured at a common temperature (either 25 degrees or 31 degrees C), was negatively correlated with egg temperature in both males and females, such that RMR of turtles from 21.5 degrees C-incubated eggs averaged 160% that of turtles from 30.5 degrees C-incubated eggs. These results indicate that egg temperatures induced positive metabolic compensation in both sexes. Thyroid hormone levels of hatchlings showed similar correlations with egg temperature; thyroxine level of turtles from 21.5 degrees C-incubated eggs averaged 220% that of turtles from 30.5 degrees C-incubated eggs. To examine the possibility that thyroid hormones contribute to positive metabolic compensation, we added triiodothyronine to eggs during mid-incubation. RMR of hatchlings from these treated eggs averaged 131% that of controls, consistent with the previous possibility. Moreover, the effects of embryonic temperature on metabolic physiology, in combination with effects on sex, can result in differences in RMR and thyroid hormone levels between male and female hatchling turtles. Such differences may be important to the ecology and evolution of TSD. PMID- 10521319 TI - Effects of temperature and physical activity on blood flow shunts and intracardiac mixing in the toad Bufo marinus. AB - Blood flow in systemic (.Qsys) and pulmocutaneous (.Qpul) arteries was measured as a function of body temperature (10 degrees, 20 degrees, and 30 degrees C) at rest and following enforced physical activity in conscious, adult cane toads (Bufo marinus). Arterial and mixed venous hemoglobin concentration (CHb) and total oxygen content (Co2, tot) were measured in a separate group under identical conditions. Heart rate (fH) and total flow (.Qtot) increased significantly (P<0.001) with elevated temperature and with activity, whereas stroke volume (VS) increased (P<0.001) only with activity. .Qtot ranged about 10-fold, from 10 degrees C (rest) to 30 degrees C (activity); increases in both fH and VS contributed to the increase in .Qtot. The overall distribution of blood to the pulmocutaneous circuit (net L-R shunt) increased with both temperature and activity and was significantly correlated with .Qtot. These data indicate that blood flow distribution in toads is a direct function of cardiac output, and this is linked to relative changes in resistance in the major outflow vessels. Arterial O2 saturation (Sa) was high (mean=93%) in all conditions except activity at 30 degrees C, when it decreased to 74% and contributed to a decrease in the arteriovenous O2 difference. Venous O2 saturation (Sv) was high at rest (76%) and dropped significantly during activity to about 30% at all temperatures. Intracardiac arterial-venous mixing (systemic mixing index) showed the strongest correlation with variation in fH with minimal mixing (17%) occurring at about 50 beats min-1. The most mixing occurred at the lowest fH (13 beats min-1) and at the highest fH (103 beats min-1). The results indicate that the heart of a 0.25 kg toad becomes more efficient from an oxygen transport perspective from low fH to 50 beats min-1 and then less efficient at higher fH, contributing to an uncoupling of blood flow and metabolic rates at these high rates. PMID- 10521321 TI - Ontogeny of intracellular isosmotic regulation in the european lobster Homarus gammarus (L.). AB - Intracellular free amino acids were measured in the abdominal muscle of the three larval instars, postlarvae, and juveniles of the lobster Homarus gammarus, acclimated to seawater (35 per thousand) and to a dilute medium (22 per thousand), to study intracellular isosmotic regulation throughout the development of this species. Transfer to low salinity was followed by a highly significant drop of free amino acids level in all developmental stages. The main regulated amino acids were glycine, proline, and alanine. The level of regulation of total free amino acids changed at metamorphosis: the decrease in total free amino acids at low salinity was 46% in the three larval instars, but it was only 29% in postlarvae and 20% in juveniles. These results suggest that free amino acids, mainly glycine, proline, and alanine, are involved in intracellular isosmotic regulation in the lobster, with different levels of involvement in pre- and postmetamorphic stages. The ontogenetic changes in intracellular isosmotic regulation are discussed in relation to the changes in extracellular regulation (osmoregulation) in the lobster. PMID- 10521322 TI - Seasonal and interindividual variation in field water metabolism of female meadow voles Microtus pennsylvanicus. AB - We analyzed variations in water flux rates on a large sample of meadow voles (Microtus pennsylvanicus) to quantify the effect of season on water metabolism of individuals and to examine patterns of intra- and interindividual variability. Voles were nonreproductive females maintained in outdoor enclosures where they fed on natural vegetation. They were injected one to three times with doubly labeled water, which resulted in one to six measures of daily water flux rate per individual. Summer water flux rates of voles were 258% of the predicted values for herbivorous eutherian mammals of similar size. To date, very few studies have focused on mammals with such high water flux rates. Body water volume of individuals was higher in summer than in winter (75.6% vs. 72.5%), and water flux rate of animals was 12.5% higher in the winter season (0.99 vs. 0.88 mL H2O g-1 d 1). Between-season differences in water fluxes were proportional to differences in energy expenditures, hence the water economy index remained constant across seasons (0.30 mL H2O kJ-1). Intraindividual variability of water flux rate was high compared to interindividual variability (repeatability, r<0.30), which will make it difficult to study natural selection of water metabolism in a microevolutionary framework, at least in meadow voles. PMID- 10521323 TI - Discontinuous gas-exchange cycles in Scarabaeus dung beetles (Coleoptera: Scarabaeidae): mass-scaling and temperature dependence. AB - Although discontinuous gas exchange cycles (DGC) are known from many insects, the effects of body size and temperature on DGC have not been widely examined. Here, these effects are investigated in five Scarabaeus dung beetle species from mesic and xeric habitats. The investigation tests two hypotheses: that previous estimates of the scaling exponents for the DGC and its characteristics are more broadly applicable to insects, and that, in response to temperature, both DGC frequency and the quantity of CO2 emitted during the open (O) phase (O-phase emission volume) are modulated. Like previous workers, we find that V&d2;co2 scaled as mass0.968 and that O-phase emission volume scaled as mass0.833. However, temperature-associated increases in .Vco2 (Q10's of 2.19-2.65) were modulated mostly by increases in DGC frequency since O-phase volumes remained constant across temperature. Flutter (F)-phase and O-phase durations were closely coupled to DGC duration, although the relationship between closed (C)-phase duration and DGC duration was less pronounced. We show that ventilation phase coefficients, previously considered a measure of the proportional duration of each phase of the DGC, calculated from the slopes of these relationships are a measure of change in phase duration with change in DGC duration and not a measure of the way in which total DGC duration is apportioned among phases. We suggest that proportions be used to estimate the contribution of each of the phases to the total duration of the DGC. PMID- 10521324 TI - Evidence for a proximate influence of winter temperature on metabolism in passerine birds. AB - The roles of ultimate and proximate factors in regulating basal and summit metabolic rates of passerine birds during winter have received little study, and the extent to which winter temperatures affect these variables is unknown. To address this question, we measured basal and summit (maximum cold-induced) metabolic rates in black-capped chickadees (Poecile atricapillus), dark-eyed juncos (Junco hyemalis), and American tree sparrows (Spizella arborea) during winters from 1991/1992 to 1997 in southeastern South Dakota. Both temperature and these metabolic rates varied within and among winters. Least-squares regression revealed significant negative relationships for normalized basal and summit metabolism against mean winter temperature for all species pooled (R2=0.62 to 0.69, P3-fold lower the initial rate and 1.5- to >2.5-fold lower the extent of water uptake seen in colder-acclimated conspecifics. Both the incubation temperature sensitivity and the acclimation effects are consistent with transmembrane water permeation. Calcium-free incubations (permitting paracellular water movement) also indicated that interfacial cell membranes contribute to gill permeability characteristics; without calcium, trout gill osmotic water uptake values increased 1.5- to 2-fold, and the temperature dependence of water uptake decreased (initial rate) or was eliminated (extent). The specific contribution of cholesterol to restricting barrier membrane water permeability was indicated by concentration-dependent increases in water uptake in the presence of either nystatin (a cholesterol-complexing, pore-forming agent) or methyl-beta cyclodextrin (which selectively depletes membrane cholesterol). In addition, a cholesterol-specific cytochemical probe (filipin) intensely labeled the apical surface membranes of trout and tilapia gill epithelium. In summary, these studies implicate membrane cholesterol in determining water permeability in fish gills. PMID- 10521331 TI - The tau of continuous feeding on simple foods. AB - Chemical reactor theory under the premise of maximization of net rate of nutrient absorption has been used to predict throughput time, tau, of digesta in animals. Animals that feed on hexoses, such as many vertebrate fruit and nectar eaters, are of central interest in testing reactor theory because they use no hydrolysis before absorption and, hence, should provide valuable, simplified test cases. Graphical methods based on batch reactors and used to make such predictions in the past can give optimal gut throughput times (tauopt) identical with predictions from continuous plug-flow reactor models derived here: in animals with passive, linear uptake alone, tauopt should decline as hexose concentration of food increases. If saturating active uptake is involved, however, a minimum in tauopt (maximum in ingestion rate) is predicted at intermediate hexose concentration, the exact location of this minimum depending on costs of ingestion as well as on uptake kinetics. That is, tauopt first falls to a minimum with increasing hexose concentration and then increases. Optimal throughput time rises as uptake sites become saturated because there is little gross gain and no net gain from increased ingestion rate when uptake already is nearly saturated. It also rises with increasing costs of ingestion. The continuous-time analytic solutions provided here further make the novel and very general prediction of high sensitivity to decreasing tau below tauopt. PMID- 10521332 TI - On the comparative ecological and evolutionary significance of total and mass specific rates of metabolism. PMID- 10521333 TI - Genomic views of human history. AB - New tools of genomic analysis shed light on historical puzzles. Migrations of ancient peoples, differences in migration patterns of males and females, historical demography of cultures with ancient roots, and patterns of human genetic diversity are increasingly the focus of integrated analysis by historians, anthropologists, and geneticists. PMID- 10521334 TI - Biosequence exegesis. AB - Annotation of large-scale gene sequence data will benefit from comprehensive and consistent application of well-documented, standard analysis methods and from progressive and vigilant efforts to ensure quality and utility and to keep the annotation up to date. However, it is imperative to learn how to apply information derived from functional genomics and proteomics technologies to conceptualize and explain the behaviors of biological systems. Quantitative and dynamical models of systems behaviors will supersede the limited and static forms of single-gene annotation that are now the norm. Molecular biological epistemology will increasingly encompass both teleological and causal explanations. PMID- 10521335 TI - The mammalian gene collection. AB - The Mammalian Gene Collection (MGC) project is a new effort by the NIH to generate full-length complementary DNA (cDNA) resources. This project will provide publicly accessible resources to the full research community. The MGC project entails the production of libraries, sequencing, and database and repository development, as well as the support of library construction, sequencing, and analytic technologies dedicated to the goal of obtaining a full set of human and other mammalian full-length (open reading frame) sequences and clones of expressed genes. PMID- 10521336 TI - The promise of comparative genomics in mammals. AB - Dense genetic maps of human, mouse, and rat genomes that are based on coding genes and on microsatellite and single-nucleotide polymorphism markers have been complemented by precise gene homolog alignment with moderate-resolution maps of livestock, companion animals, and additional mammal species. Comparative genetic assessment expands the utility of these maps in gene discovery, in functional genomics, and in tracking the evolutionary forces that sculpted the genome organization of modern mammalian species. PMID- 10521337 TI - Epigenetics: regulation through repression. AB - Epigenetics is the study of heritable changes in gene expression that occur without a change in DNA sequence. Epigenetic phenomena have major economic and medical relevance, and several, such as imprinting and paramutation, violate Mendelian principles. Recent discoveries link the recognition of nucleic acid sequence homology to the targeting of DNA methylation, chromosome remodeling, and RNA turnover. Although epigenetic mechanisms help to protect cells from parasitic elements, this defense can complicate the genetic manipulation of plants and animals. Essential for normal development, epigenetic controls become misdirected in cancer cells and other human disease syndromes. PMID- 10521338 TI - Pharmacogenomics: translating functional genomics into rational therapeutics. AB - Genetic polymorphisms in drug-metabolizing enzymes, transporters, receptors, and other drug targets have been linked to interindividual differences in the efficacy and toxicity of many medications. Pharmacogenomic studies are rapidly elucidating the inherited nature of these differences in drug disposition and effects, thereby enhancing drug discovery and providing a stronger scientific basis for optimizing drug therapy on the basis of each patient's genetic constitution. PMID- 10521339 TI - A cyclic antimicrobial peptide produced in primate leukocytes by the ligation of two truncated alpha-defensins. AB - Analysis of rhesus macaque leukocytes disclosed the presence of an 18-residue macrocyclic, tridisulfide antibiotic peptide in granules of neutrophils and monocytes. The peptide, termed rhesus theta defensin-1 (RTD-1), is microbicidal for bacteria and fungi at low micromolar concentrations. Antibacterial activity of the cyclic peptide was threefold greater than that of an open-chain analog, and the cyclic conformation was required for antimicrobial activity in the presence of 150 millimolar sodium chloride. Biosynthesis of RTD-1 involves the head-to-tail ligation of two alpha-defensin-related nonapeptides, requiring the formation of two new peptide bonds. Thus, host defense cells possess mechanisms for synthesis and granular packaging of macrocyclic antibiotic peptides that are components of the phagocyte antimicrobial armamentarium. PMID- 10521340 TI - Rapid and reversible effects of activity on acetylcholine receptor density at the neuromuscular junction in vivo. AB - Quantitative fluorescence imaging was used to study the regulation of acetylcholine receptor (AChR) number and density at neuromuscular junctions in living adult mice. At fully functional synapses, AChRs have a half-life of about 14 days. However, 2 hours after neurotransmission was blocked, the half-life of the AChRs was now less than a day; the rate was 25 times faster than before. Most of the lost receptors were not quickly replaced. Direct muscle stimulation or restoration of synaptic transmission inhibited this process. AChRs that were removed from nonfunctional synapses resided for hours in the perijunctional membrane before being locally internalized. Dispersed AChRs could also reaggregate at the junction once neurotransmission was restored. The rapid and reversible alterations in AChR density at the neuromuscular junction in vivo parallel changes thought to occur in the central nervous system at synapses undergoing potentiation and depression. PMID- 10521341 TI - Role of metal-oxide interface in determining the spin polarization of magnetic tunnel junctions AB - The role of the metal-oxide interface in determining the spin polarization of electrons tunneling from or into ferromagnetic transition metals in magnetic tunnel junctions is reported. The spin polarization of cobalt in tunnel junctions with an alumina barrier is positive, but it is negative when the barrier is strontium titanate or cerium lanthanite. The results are ascribed to bonding effects at the transition metal-barrier interface. The influence of the electronic structure of metal-oxide interfaces on the spin polarization raises interesting fundamental problems and opens new ways to optimize the magnetoresistance of tunnel junctions. PMID- 10521342 TI - Emergence of scaling in random networks AB - Systems as diverse as genetic networks or the World Wide Web are best described as networks with complex topology. A common property of many large networks is that the vertex connectivities follow a scale-free power-law distribution. This feature was found to be a consequence of two generic mechanisms: (i) networks expand continuously by the addition of new vertices, and (ii) new vertices attach preferentially to sites that are already well connected. A model based on these two ingredients reproduces the observed stationary scale-free distributions, which indicates that the development of large networks is governed by robust self organizing phenomena that go beyond the particulars of the individual systems. PMID- 10521343 TI - Osmium isotope constraints on ore metal recycling in subduction zones AB - Veined peridotite xenoliths from the mantle beneath the giant Ladolam gold deposit on Lihir Island, Papua New Guinea, are 2 to 800 times more enriched in copper, gold, platinum, and palladium than surrounding depleted arc mantle. Gold ores have osmium isotope compositions similar to those of the underlying subduction-modified mantle peridotite source region, indicating that the primary origin of the metals was the mantle. Because the mantle is relatively depleted in gold, copper, and palladium, tectonic processes that enhance the advective transport and concentration of these fluid soluble metals may be a prerequisite for generating porphyry-epithermal copper-gold deposits. PMID- 10521345 TI - Slope water current over the laurentian fan on interannual to millennial time scales AB - The strength and position of surface and deep currents in the slope water south of Newfoundland are thought to vary as a coupled system in relation to the dipole in atmospheric sea level pressure known as the North Atlantic oscillation (NAO). Paleoceanographic data from the Laurentian Fan, used as a proxy for sea surface temperature, reveal that surface slope waters north of the Gulf Stream experienced warming during the Little Ice Age of the 16th to 19th centuries and support the notion of an NAO-driven coupled system. The NAO may be a useful model for millennial-scale ocean variability during interglacial climate states. PMID- 10521344 TI - Continuous deformation versus faulting through the continental lithosphere of new zealand AB - Seismic anisotropy and P-wave delays in New Zealand imply widespread deformation in the underlying mantle, not slip on a narrow fault zone, which is characteristic of plate boundaries in oceanic regions. Large magnitudes of shear wave splitting and orientations of fast polarization parallel to the Alpine fault show that pervasive simple shear of the mantle lithosphere has accommodated the cumulative strike-slip plate motion. Variations in P-wave residuals across the Southern Alps rule out underthrusting of one slab of mantle lithosphere beneath another but permit continuous deformation of lithosphere shortened by about 100 kilometers since 6 to 7 million years ago. PMID- 10521346 TI - Multiple ink nanolithography: toward a multiple-Pen nano-plotter AB - The formation of intricate nanostructures will require the ability to maintain surface registry during several patterning steps. A scanning probe method, dip pen nanolithography (DPN), can be used to pattern monolayers of different organic molecules down to a 5-nanometer separation. An "overwriting" capability of DPN allows one nanostructure to be generated and the areas surrounding that nanostructure to be filled in with a second type of "ink." PMID- 10521347 TI - Beta-defensins: linking innate and adaptive immunity through dendritic and T cell CCR6. AB - Defensins contribute to host defense by disrupting the cytoplasmic membrane of microorganisms. This report shows that human beta-defensins are also chemotactic for immature dendritic cells and memory T cells. Human beta-defensin was selectively chemotactic for cells stably transfected to express human CCR6, a chemokine receptor preferentially expressed by immature dendritic cells and memory T cells. The beta-defensin-induced chemotaxis was sensitive to pertussis toxin and inhibited by antibodies to CCR6. The binding of iodinated LARC, the chemokine ligand for CCR6, to CCR6-transfected cells was competitively displaced by beta-defensin. Thus, beta-defensins may promote adaptive immune responses by recruiting dendritic and T cells to the site of microbial invasion through interaction with CCR6. PMID- 10521348 TI - A new primate from the Middle Eocene of Myanmar and the Asian early origin of anthropoids. AB - A new genus and species of anthropoid primate, Bahinia pondaungensis gen. et sp. nov., is described from the Yashe Kyitchaung locality in the Late Middle Eocene Pondaung Formation (Myanmar). It is related to Eosimias, but it is represented by more complete remains, including upper dentition with associated lower jaw fragment. It is interpreted as a new representative of the family Eosimiidae, which corresponds to the sister group of the Amphipithecidae and of all other anthropoids. Eosimiidae are now recorded from three distinct Middle Eocene localities in Asia, giving support to the hypothesis of an Asian origin of anthropoids. PMID- 10521349 TI - Molecular classification of cancer: class discovery and class prediction by gene expression monitoring. AB - Although cancer classification has improved over the past 30 years, there has been no general approach for identifying new cancer classes (class discovery) or for assigning tumors to known classes (class prediction). Here, a generic approach to cancer classification based on gene expression monitoring by DNA microarrays is described and applied to human acute leukemias as a test case. A class discovery procedure automatically discovered the distinction between acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) without previous knowledge of these classes. An automatically derived class predictor was able to determine the class of new leukemia cases. The results demonstrate the feasibility of cancer classification based solely on gene expression monitoring and suggest a general strategy for discovering and predicting cancer classes for other types of cancer, independent of previous biological knowledge. PMID- 10521350 TI - Sequencing complex polysaccharides. AB - Although rapid sequencing of polynucleotides and polypeptides has become commonplace, it has not been possible to rapidly sequence femto- to picomole amounts of tissue-derived complex polysaccharides. Heparin-like glycosaminoglycans (HLGAGs) were readily sequenced by a combination of matrix assisted laser desorption ionization mass spectrometry and a notation system for representation of polysaccharide sequences. This will enable identification of sequences that are critical to HLGAG biological activities in anticoagulation, cell growth, and differentiation. PMID- 10521351 TI - Stability and variability in competitive communities. AB - Long-term variability in the abundance of populations depends on the sensitivity of species to environmental fluctuations and the amplification of environmental fluctuations by interactions among species. Although competitive interactions and species number may have diverse effects on variability measured at the individual species level, a combination of theoretical analyses shows that these factors have no effect on variability measured at the community level. Therefore, biodiversity may increase community stability by promoting diversity among species in their responses to environmental fluctuations, but increasing the number and strength of competitive interactions has little effect. PMID- 10521352 TI - CFTR chloride channel regulation by an interdomain interaction. AB - The cystic fibrosis gene encodes a chloride channel, CFTR (cystic fibrosis transmembrane conductance regulator), that regulates salt and water transport across epithelial tissues. Phosphorylation of the cytoplasmic regulatory (R) domain by protein kinase A activates CFTR by an unknown mechanism. The amino terminal cytoplasmic tail of CFTR was found to control protein kinase A-dependent channel gating through a physical interaction with the R domain. This regulatory activity mapped to a cluster of acidic residues in the NH(2)-terminal tail; mutating these residues proportionately inhibited R domain binding and CFTR channel function. CFTR activity appears to be governed by an interdomain interaction involving the amino-terminal tail, which is a potential target for physiologic and pharmacologic modulators of this ion channel. PMID- 10521353 TI - Neurogenesis in the neocortex of adult primates. AB - In primates, prefrontal, inferior temporal, and posterior parietal cortex are important for cognitive function. It is shown that in adult macaques, new neurons are added to these three neocortical association areas, but not to a primary sensory area (striate cortex). The new neurons appeared to originate in the subventricular zone and to migrate through the white matter to the neocortex, where they extended axons. These new neurons, which are continually added in adulthood, may play a role in the functions of association neocortex. PMID- 10521354 TI - Yeast gene for a Tyr-DNA phosphodiesterase that repairs topoisomerase I complexes. AB - Covalent intermediates between topoisomerase I and DNA can become dead-end complexes that lead to cell death. Here, the isolation of the gene for an enzyme that can hydrolyze the bond between this protein and DNA is described. Enzyme defective mutants of yeast are hypersensitive to treatments that increase the amount of covalent complexes, indicative of enzyme involvement in repair. The gene is conserved in eukaryotes and identifies a family of enzymes that has not been previously recognized. The presence of this gene in humans may have implications for the effectiveness of topoisomerase I poisons, such as the camptothecins, in chemotherapy. PMID- 10521355 TI - ADP receptors and clinical bleeding disorders. AB - ADP plays a key role in hemostasis and thrombosis. Despite its early identification in 1961 as the first known aggregating agent, the molecular basis of ADP-induced platelet activation is only beginning to be understood. The present review proposes a model of 3 purinergic receptors contributing separately to the complex process of ADP-induced platelet aggregation: the P2X(1) ionotropic receptor, responsible for rapid influx of ionized calcium into the cytosol; the P2Y(1) metabotropic receptor, responsible for mobilization of ionized calcium from internal stores, which initiates aggregation; and an as-yet-unidentified P2Y receptor coupled to G(alphai2), which is essential for the full aggregation response to ADP. It is probable that this as-yet-unidentified receptor is the molecular target of the ADP-selective antiaggregating drugs ticlopidine and clopidogrel. In addition, it is probably defective in patients with a bleeding diathesis that is characterized by selective impairment of platelet responses to ADP. PMID- 10521356 TI - Histamine induces tyrosine phosphorylation of endothelial cell-to-cell adherens junctions. AB - Endothelial adherens junctions (AJ) promote intercellular adhesion and may contribute to the control of vascular permeability. These structures are formed by a transmembrane and cell-specific adhesive protein, vascular endothelial (VE) cadherin, which is linked by its cytoplasmic tail to intracellular proteins called catenins (alpha-catenin, beta-catenin, and plakoglobin) and to the actin cytoskeleton. Little is known about the functional regulation of AJ in endothelial cells. In this study, we analyzed the effect of histamine on AJ organization in cultured endothelial cells. We first observed that histamine induced detectable intercellular gaps only in loosely-confluent cells, whereas this effect was strongly reduced or absent in long-confluent cultures. Despite this difference, in vitro permeability was augmented by histamine in both conditions. In resting conditions, tyrosine phosphorylation of AJ components and permeability values were higher in recently-confluent cells as compared with long confluent cells. Histamine did not affect the phosphorylation state of AJ in recently-confluent cells but strongly increased this parameter in long-confluent cultures. In addition, in long-confluent cells, histamine caused dissociation of VE-cadherin from the actin cytoskeleton measured by a decrease of the amount of the molecule in the detergent-insoluble fraction of the cell extracts. Dibutyryl cAMP was able to prevent the effect of histamine on both tyrosine phosphorylation of AJ components and on endothelial permeability. The effect of histamine was specific for VE-cadherin because the phosphorylation state of neural (N) cadherin, the other major endothelial cadherin, was unchanged by this agent. Hence AJ components are a target of histamine activation cascade; we suggest that induction of tyrosine phosphorylation of VE-cadherin and catenins contributes to the histamine effect on permeability, even in absence of frank intercellular gaps and cell retraction. PMID- 10521357 TI - Adaptive remodeling of internal elastic lamina and endothelial lining during flow induced arterial enlargement. AB - Gaps in the internal elastic lamina (IEL) have been observed in arteries exposed to high blood flow. To characterize the nature and consequences of this change, blood flow was increased in the carotid arteries of 56 adult, male, Japanese white rabbits by creating an arteriovenous fistula between the common carotid artery and the external jugular vein. The common carotid artery proximal to the arteriovenous fistula was studied at intervals from 1 hour to 8 weeks after exposure to high flow. In the controls, the IEL showed only the usual, small, physiological holes, 2 to 10 microm in diameter. At 3 days, some of the holes in the IEL had become enlarged, but they could not be detected by scanning electron microscopy, despite manifest endothelial cell proliferation. At 4 days, gaps in the IEL appeared as small, luminal surface depressions, 15 to 50 microm wide. At 7 days, the gaps in the IEL had enlarged and formed circumferential, luminal depressions occupying 15+/-5% of the lumen surface. Endothelial cell proliferation persisted in the gaps while proliferative activity decreased where the IEL remained intact. At 4 weeks, as the artery became elongated and dilated, the gaps in the IEL widened as intercommunicating circumferential and longitudinal luminal depressions occupying 64+/-5% of the lumen surface. At 8 weeks, the rate of elongation and dilatation of the artery slowed and the widening of the gaps in the IEL diminished. Endothelial cells covered the gaps throughout. We conclude that flow-induced arterial dilatation is accompanied by an adaptive remodeling of the intima. The gaps in the IEL permit an increase in lumen surface area while endothelial cell proliferation assures a continuous cell lining throughout. PMID- 10521358 TI - Influence of elastin gene polymorphism on the elastin content of the aorta: A study in 2 strains of rat. AB - The elastin content in the thoracic aorta of male Brown-Norway (BN) rats is 31.4+/-1.2% (dry weight), whereas that of male LOU rats is 37.2+/-1.0%. A similar difference in the elastin content of the thoracic aorta is also observed in female animals. Furthermore, in the thoracic aorta of young, growing rats as well as in cultured aortic smooth muscle cells, the steady-state level of elastin mRNA is significantly lower in the BN than in the LOU strain. These results suggested that 1 or more genes control the elastin mRNA level and the elastin content in the aortas of BN and LOU rats. A possible relationship between a polymorphism in the elastin gene and the elastin content of the aorta was tested. For this purpose, the aortic elastin content was measured in F(1) and F(2) generations bred from LOU and BN rats and was compared with that of the F(0) (parental) generation. A polymorphic marker located in intron 25 of the elastin gene has been used to genotype the F(2) rats. The degree of genetic determination of aortic elastin content was estimated to be 73% in the F(2) cohort, but the elastin locus accounts for only 3. 9% of the total variance in aortic elastin content. Other genes are thus responsible for the major part of the observed interstrain difference by regulating the transcription of the gene, the stability of elastin mRNA, and/or posttranslational events. PMID- 10521359 TI - Eicosapentaenoic acid and docosahexaenoic acid block serotonin-induced smooth muscle cell proliferation. AB - Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) present in fish oils have been ascribed as having significant antithrombotic and antiatherosclerotic effects. Vascular smooth muscle cell (SMC) proliferation plays an important role in the pathogenesis of atherosclerosis and restenosis. Recent studies have indicated that serotonin at concentrations present at sites of vascular injury stimulates SMC proliferation and may contribute to the restenotic process. In the present study we demonstrate that among the fatty acids tested, only EPA and DHA could block the mitogenic effect of serotonin on vascular SMC. Further, when added together these fatty acids act synergistically in blocking the mitogenic effect of serotonin. EPA and DHA blocked the 5HT-induced increase in the 5-HT(2) receptor mRNA. This antimitogenic effect of EPA and DHA may partially explain some of the beneficial effects of fish oils. PMID- 10521360 TI - Autocrine FGF-2 is responsible for the cell density-dependent susceptibility to apoptosis of HUVEC : A role of a calpain inhibitor-sensitive mechanism. AB - To elucidate the factors affecting endothelial susceptibility to apoptosis, we studied the effects of cell density on endothelial cell apoptosis induced by deprivation of serum and fibroblast growth factor-2 (FGF-2/basic FGF). On deprivation, more cells became apoptotic in a dense culture (5 x 10(2) cells/mm(2)) than in a sparse culture (1 x 10(2) cells/mm(2)) of human umbilical vein endothelial cells. FGF-2, hepatocyte growth factor, and vascular endothelial cell growth factor, but not insulin-like growth factor-I, decreased apoptosis in the dense culture to a level similar in the sparse culture. An anti-FGF-2 antibody significantly increased the apoptosis in the sparse culture, suggesting that the sparse culture was resistant to apoptosis because of the greater autocrine production of FGF-2. Western blot analysis and metabolic labeling revealed that the sparse culture has, in fact, more FGF-2 than the dense culture. The steady state level of mRNA for FGF-2 was not significantly different between the dense and sparse cultures. Among a panel of inhibitors for 2 major cytoplasmic proteolytic enzymes, calpain inhibitors increased FGF-2 in the dense culture, but proteasome inhibitors did not. Our findings demonstrate that cell density affects endothelial survival by regulating autocrine FGF-2 production through a calpain inhibitor-sensitive mechanism. PMID- 10521361 TI - An in vitro coculture model of transmigrant monocytes and foam cell formation. AB - To analyze in vitro the migration of monocytes to the subendothelial space, their differentiation into macrophages, and the subsequent formation of foam cells in vitro, we have developed a 2-coculture system with rabbit aortic endothelial cells (AECs), aortic smooth muscle cells (SMCs), and a mixture of matrix proteins on polyethylene filters in chemotaxis chambers. AECs were seeded on a mixture of type I and IV collagen with or without various types of serum lipoproteins (method 1) or on matrix proteins secreted by SMCs (method 2). In these coculture systems, rabbit AECs can maintain a well-preserved monolayer for up to 2 weeks. When human CD14-positive monocytes were added to the upper medium of the system, with monocyte chemotactic protein-1 treatment approximately 60% of the monocytes transmigrated within 24 hours and were retained for up to 7 days, whereas without MCP-1 treatment, <30% of monocytes transmigrated. On day 1, transmigrant monocytes were negative for immunostaining of type I and II macrophage scavenger receptors but by day 3, became positive for scavenger receptors as well as other macrophage markers. When oxidized low density lipoprotein was added to the matrix layer of the method I coculture, on day 4 transmigrant cells exhibited lipid deposit droplets, and by day 7, they had the appearance of typical foam cells. Some of the transmigrant cells recovered in the lower medium on day 7 also appeared to be foam cells, indicating foam cell motility and escape from the coculture layer through the filter. In summary, this coculture system is a useful in vitro tool to dissect the cellular and molecular events that make up the process of foam cell formation. PMID- 10521362 TI - Expression of the angiogenic protein, platelet-derived endothelial cell growth factor, in coronary atherosclerotic plaques: In vivo correlation of lesional microvessel density and constrictive vascular remodeling. AB - Recent information indicates that platelet-derived endothelial cell growth factor (PD-ECGF), a 45-kDa angiogenic protein, is expressed in the endothelium of various tissues and that its level of expression is correlated with the number of microvessels in human tumors. Because the formation of neovessels is also thought to play a role in atherosclerotic vascular remodeling, we analyzed PD-ECGF expression in fresh, coronary plaque tissues obtained by directional coronary atherectomy. Specimens from 31 patients were collected and analyzed by reverse transcription-polymerase chain reaction, histochemical staining, immunohistochemistry, and in situ hybridization with the use of PD-ECGF-specific primers and probes. Lesional vascular remodeling was assessed by intravascular ultrasound. PD-ECGF immunoreactivity and mRNA were found in plaque macrophages, endothelial cells of plaque neovessels, and stellate smooth muscle cells of 20 atherectomy specimens (64.5%). PD-ECGF immunoreactivity was correlated with the number of lesional microvessels and mast cells. Double-staining experiments revealed a close spatial proximity of PD-ECGF-positive cells and mast cells. Furthermore, the numbers of microvessels and mast cells were significantly higher in lesions lacking compensatory enlargement. The data indicate that PD-ECGF is expressed within cells of the atherosclerotic plaque and may be involved in driving angiogenesis in concert with mast cells. PMID- 10521363 TI - Complement and atherogenesis: binding of CRP to degraded, nonoxidized LDL enhances complement activation. AB - Complement activation occurs in temporal correlation with the subendothelial deposition of LDL during early atherogenesis, and complement also plays a pathogenetic role in promoting lesion progression. Two lesion components have been identified that may be responsible for complement activation. First, enzymatic degradation of LDL generates a derivative that can spontaneously activate complement, and enzymatically degraded LDL (E-LDL) has been detected in the lesions. Second, C-reactive protein (CRP) colocalizes with complement C5b-9, as evidenced by immunohistological studies of early atherosclerotic lesions, so the possibility exists that this acute phase protein also fulfills a complement activating function. Here, we report that addition of LDL and CRP to human serum did not result in significant C3 turnover. Addition of E-LDL provoked complement activation, which was markedly enhanced by CRP. Binding of CRP to E-LDL was demonstrated by sucrose flotation experiments. Binding was Ca(2+)-dependent and inhibitable by phosphorylcholine, and the complement-activating property of E-LDL was destroyed by treatment with phospholipase C. These results indicated that CRP binds to phosphorylcholine groups that become exposed in enzymatically degraded LDL particles. Immunohistological studies complemented these findings in showing that CRP colocalizes with E-LDL in early human atherosclerotic lesions. Thus enzymatic, nonoxidative modification of tissue-deposited LDL can be expected to confer CRP-binding capacity onto the molecule. The ensuing enhancement of complement activation may be relevant to the development and progression of the atherosclerotic lesion. PMID- 10521364 TI - Systemic inflammatory parameters in patients with atherosclerosis of the coronary and peripheral arteries. AB - Plasma concentration of markers of inflammation are increased in patients with atherosclerosis. However, it is unclear whether the pattern and magnitude of this increase vary with the site and extent of disease. In 147 patients undergoing semiquantitative coronary angiography, we measured the acute-phase reactants C reactive protein (CRP) or serum amyloid A (SAA); the proinflammatory cytokine interleukin 6 (IL-6); the active and total fractions of the anti-inflammatory cytokine transforming growth factor-beta (TGF-beta); the macrophage activation marker neopterin; and the infection marker procalcitonin. Compared with 62 patients without either coronary artery disease (CAD) or peripheral artery disease (PAD), 57 patients with CAD but no PAD showed greater median CRP (0. 4 versus 0.2 mg/dL, P=0.004) and IL-6 (3.8 versus 1.6 pg/mL, P=0. 007) levels and a lower level of active-TGF-beta (57 versus 100 ng/mL, P=0.038). Moreover, CRP, IL 6, and neopterin levels showed a positive and the active TGF-beta level a negative correlation with the extent of coronary atherosclerosis. Compared with these 57 patients with CAD alone, 15 patients with PAD and CAD had higher median levels of SAA (17 versus 7 mg/mL, P=0.008), IL-6 (12 versus 4 pg/mL, P=0.002), neopterin (14 versus 11 mg/dL, P=0.006), and total TGF-beta (11834 versus 6417 ng/L, P=0.001). However, these strong univariate associations of markers of inflammation and atherosclerosis were lost in multivariate analysis once age, sex, and high density lipoprotein cholesterol or fibrinogen were taken into account. Increased plasma levels of CRP, SAA, IL-6, TGF-beta, neopterin, and procalcitonin constitute an inflammatory signature of advanced atherosclerosis and are correlated with the extent of disease but do not provide discriminatory diagnostic power over and above established risk factors. PMID- 10521365 TI - Interleukin-6 exacerbates early atherosclerosis in mice. AB - Acute-phase proteins, which respond to systemic proinflammatory cytokines such as interleuken-6, are elevated in cardiovascular disease and are predictive markers of future ischemic events, even over decades. This suggests a role for proinflammatory cytokines and/or acute phase proteins in early lesion development. To explore this issue, we fed C57Bl/6 and nonobese diabetic male mice high-fat (20% total fat, 1.5% cholesterol) diets and ApoE-deficient male mice both high-fat and normal chow diets for 6 to 21 weeks, injecting them weekly with either 5000 U recombinant interleukin-6 (rIL-6) or saline buffer. Blood was collected when animals were euthanized and assayed for cytokines, acute-phase proteins, and cholesterol. Across all mice, IL-6 injection resulted in significant increases in proinflammatory cytokines (IL-6, 4.6-fold; IL-1beta, 1.6 fold; and tissue necrosis factor-alpha, 1.7-fold) and fibrinogen (1.2-fold) and with decreased concentrations of albumin (0.9-fold) in plasma. Total cholesterol levels were unchanged between rIL-6-treated and nontreated groups. Serial sections through the aortic sinus were stained with oil red O to detect fatty streaks, and area of the lesions was determined by image analysis. Although no fatty streaks were detected in the nonobese diabetic mice with or without rIL-6 treatment, rIL-6 treatment increased lesion size in C57Bl/6 and ApoE-deficient mice 1.9- to 5.1-fold over lesions in saline-treated animals. These results suggest that under the appropriate circumstances changes in circulating proinflammatory cytokines and acute-phase proteins may be more than just markers of atherosclerosis but actual participants in early lesion development. PMID- 10521366 TI - Low-cholesterol and high-fat diets reduce atherosclerotic lesion development in ApoE-knockout mice. AB - We have investigated the effect of most common oils used in human nutrition on the development of atherosclerosis in apoE-knockout mice. Seven groups of animals, separated according to sex, were fed for 10 weeks either chow diet or the chow diet 10% (wt/wt) enriched with different oils (palm, coconut, 2 types of olive oil, and 2 types of sunflower oil) without addition of cholesterol. At the end of this period, plasma lipid parameters were measured and vascular lesions scored. None of the diets induced changes in plasma cholesterol concentrations, whereas plasma triglycerides were uniformly reduced in all diet groups. Some diets caused significant reductions in the size of atherosclerotic lesions in males and others in females; males responded most to sunflower oils and females to palm oil and one olive oil (II). The lesion reduction in males consuming sunflower oils was associated with the decrease of triglycerides in triglyceride rich lipoproteins, whereas the decrease in females consuming olive oil II or palm oil was accompanied by an increase in plasma apoA-I. The increase in plasma apoA I in the latter condition, is mainly due to overexpression of hepatic message elicited by a mechanism independent of apoE ligand. The data suggest that the different diets modulate lesion development in a gender specific manner and by different mechanisms and that the development of atherosclerosis, due to genetic deficiencies, may be modulated by nutritional maneuvers that may be implemented in human nutrition. PMID- 10521367 TI - Two patterns of lipid deposition in the cholesterol-fed rabbit. AB - A central feature of arterial lipid deposition is its nonuniform and variable distribution. In immature human and rabbit aortas, spontaneous lesions occur most frequently downstream of branch points, but they tend to occur upstream of the same branches at later ages. In cholesterol-fed rabbits, the juvenile pattern has been seen regardless of age. These distributions may be determined by transport properties of the arterial wall, because uptake of plasma macromolecules is elevated downstream of aortic branches in immature rabbits and upstream in mature ones, except during cholesterol feeding, when the juvenile pattern is seen in adult vessels. The effect of cholesterol could reflect its inhibitory influence on the nitric oxide (NO) pathway because the adult transport pattern is NO dependent. Using protocols expected to preserve NO function and the mature pattern of transport during hypercholesterolemia, we made 2 attempts to induce upstream disease in rabbits. In trial I, plasma concentrations of cholesterol were kept within the normal human range for 15 weeks by using dietary levels of 0.05% to 0.2%. Although disease patterns reverse with age in human vessels exposed to these concentrations, lesions in both immature and mature rabbits occurred downstream of intercostal branch ostia. Trial II used older rabbits, a different base diet containing more vitamin E (96 mg/kg rather than 57 mg/kg), and higher levels of cholesterol (1%, administered for 8 weeks). For some animals, extra vitamin E (2000 mg/kg) was added to the diet. The mature pattern of lipid deposition was apparent around intercostal branches in the first group and was accentuated by the additional vitamin E, a change that was associated with a significant increase in the plasma concentration of NO metabolites. Spontaneous lesions, assessed on the base diet, were too rare to have influenced these distributions. This is the first report of upstream disease in the cholesterol-fed rabbit. The results support but do not prove the view that NO and transport are important in atherogenesis. PMID- 10521368 TI - Vitamin C protects human vascular smooth muscle cells against apoptosis induced by moderately oxidized LDL containing high levels of lipid hydroperoxides. AB - Vascular cell death is a key feature of atherosclerotic lesions and may contribute to the plaque "necrotic" core, cap rupture, and thrombosis. Oxidatively modified low-density lipoproteins (LDLs) are implicated in the pathogenesis of atherosclerosis, and dietary antioxidants are thought to protect the vasculature against LDL-induced cytotoxicity. Because LDL oxidative modification may vary within atherosclerotic lesions, we examined the effects of defined, oxidatively modified LDL species on human arterial smooth muscle cell apoptosis and the cytoprotective effects of vitamin C. Moderately oxidized LDL (0 to 300 microg protein/mL), which has the highest content of lipid hydroperoxides, induced smooth muscle cell apoptosis within 6 hours, whereas native LDL and mildly and highly oxidized LDL had no effect. Moderately oxidized LDL increased cellular DNA fragmentation, release of fragmented DNA into the culture medium, and annexin V binding and decreased mitochondrial dehydrogenase activity and expression of the antiapoptotic mediator Bcl-x(L). Treatment of cells with native LDL together with the lipid hydroperoxide 13(S)-hydroperoxyoctadeca-9Z,11E dienoic acid (HPODE, 200 micromol/L, 6 to 24 hours) also induced apoptotic cell death. Pretreatment of smooth muscle cells with vitamin C (0 to 100 micromol/L, 24 hours) attenuated the cytotoxicity and apoptosis induced by both moderately oxidized LDL and HPODE. Our findings suggest that moderately oxidized LDL, with its high lipid hydroperoxide content, rather than mildly or highly oxidized LDL, causes apoptosis of human smooth muscle cells and that vitamin C supplementation may provide protection against plaque instability in advanced atherosclerosis. PMID- 10521369 TI - Cholesterol-induced changes of type VIII collagen expression and distribution in carotid arteries of rabbit. AB - Lipoproteins play a major role in cardiovascular disease and atherosclerosis. In the vascular wall, they strongly influence the organization of extracellular matrix. The present study set out to investigate the changes in the extracellular matrix of the vessel wall induced by atherogenic diet, focusing on type VIII collagen, a vascular collagen that has not previously been investigated in detail. The influence of cholesterol diet on the expression, distribution, and deposition of type VIII collagen was examined in carotid arteries of New Zealand White rabbits. Carotid arteries of rabbits receiving diet supplemented with 1% cholesterol for 6 weeks and those on the same regimen followed by normal chow for 1 day, 10 days, 5 weeks, and 12 weeks were studied and compared with controls not exposed to the cholesterol diet. Carotid arteries of normocholesterolemic rabbits contained type VIII collagen-expressing cells in all layers, with focal accumulations of expressing cells in the subendothelial areas, the outer medial zone, and the adventitia. In response to cholesterol diet, type VIII collagen synthesis was reduced in media and adventitia and the distribution patterns changed. Expressing cells were found predominantly in the endothelium, and type VIII collagen accumulated in the intimal space. Immunogold labeling for electron microscopy revealed that type VIII collagen in the intima is associated with microfibrils extending from the internal elastic lamina. Withdrawal of cholesterol resulted in reestablishment of the normal distribution pattern. Northern and Western blot analyses supported the immunoconfocal and in situ hybridization data, demonstrating decreased type VIII collagen expression in response to cholesterol diet and progressive recovery to normal levels with time after withdrawal of cholesterol. Our study demonstrates that type VIII collagen is modulated in the presence of cholesterol. The data indicate that type VIII collagen is specifically remodeled during early experimental atherosclerosis, implying a role for this extracellular matrix component in neointimal growth. PMID- 10521370 TI - Regulatory effects of HDL on smooth muscle cell prostacyclin release. AB - One mechanism by which high density lipoproteins (HDLs) exert their protective effect against coronary artery disease could be related to the induction of prostacyclin (PGI(2)) release in the vessel wall. We have recently shown that HDL increases PGI(2) production in rabbit smooth muscle cells (RSMCs) and that this increase is dependent on cyclooxygenase-2 (Cox-2). Here we analyze the mechanism by which rabbit HDL induces PGI(2) release in RSMCs. Our results show that although HDL(2) and HDL(3) share a similar capacity to induce Cox-2 protein levels, HDL(3) stimulates a higher PGI(2) release than does HDL(2), probably because of their relative arachidonate contents. Acetylsalicylic acid pretreatment (300 micromol/L, 30 minutes) significantly reduced the HDL-induced PGI(2) release, suggesting that both preexisting and induced Cox-2 activities were involved in the HDL effect. Ca(2+)-dependent cytosolic phospholipase A(2) (cPLA(2)) and Cox-1 protein levels were not altered by HDL. Dexamethasone (2 micromol/L), which also inhibited the HDL-induced PGI(2) release, reduced significantly both Cox-2 mRNA and protein levels without affecting cPLA(2) and Cox-1 protein levels. In addition, methylarachidonyl fluorophosphonate, a potent inhibitor of cPLA(2), did not produce any effect on HDL-induced PGI(2) release. In the presence of cycloheximide, Cox-2 mRNA levels were induced by HDL and inhibited by dexamethasone, suggesting that HDL and dexamethasone work in the absence of de novo protein synthesis. These results indicate an early effect of HDL on PGI(2) biosynthesis, specifically increasing Cox-2. PD98059, an inhibitor of mitogen-activated protein kinase kinase, completely inhibited HDL-induced PGI(2) release, whereas GF109203X, a protein kinase C inhibitor, had no effect. Thus, HDL induces PGI(2) synthesis by a mechanism dependent on the mitogen activated protein kinase pathway but independent of protein kinase C. PMID- 10521371 TI - Evidence that lipoproteins are carriers of bioactive factors. AB - We recently demonstrated that the mitogenic effect of LDL (100 microg/mL) as well as its early intracellular signaling pathway are mediated by a pertussis-toxin (PTX)-sensitive G(i) protein-coupled receptor that is independent from its classical receptor and involves activation of extracellular response kinases (ERK1/2) (also known as p44(mapk)/p42(mapk)). In the present study we examined whether LDL-adherent factors may be responsible for some of the effects of LDL. The term "signaling activity" is used to characterize fractions that cause an increase in intracellular free Ca(2+) concentration or stimulate ERK1/2 and c-fos mRNA expression. LDL, HDL, and VLDL stimulate ERK1/2 with the following order of potency: LDL>HDL>VLDL. After delipidation of LDL with chloroform/methanol/water mixtures a PTX-sensitive signaling activity was found in one fraction arbitrarily called LDL-F. After further analysis of LDL-F compounds by high pressure liquid chromatography, a PTX-sensitive signaling activity was detected only in the fraction with a retention time of 33 minutes (arbitrarily called LDL-F33). Similarly, after separation of sphingosine-1-phosphate (SPP) and sphingosylphosphorylcholine (SPC) by high pressure liquid chromatography, a PTX sensitive signaling activity was found in the fractions 33 and 33 to 35, respectively. These findings demonstrate that the effects of LDL-F33 are mimicked by similar fractions collected from SPP/SPC, hence suggesting that these LDL adherent molecules are possibly closely related to SPP/SPC. A PTX-sensitive signaling activity was also detected in HDL and HDL-F33. Therefore, LDL and other lipoproteins may function as carriers for bioactive phospholipids thereby contributing to the development of coronary artery disease. Our findings support a new research concept that may contribute in elucidating cellular mechanisms promoting coronary artery disease. PMID- 10521372 TI - LDL particle size distribution is associated with carotid intima-media thickness in healthy 50-year-old men. AB - Results of cross-sectional and prospective studies have suggested that small, dense low-density lipoprotein (LDL) particles predispose to coronary heart disease. We investigated the relationships between plasma concentrations of LDL subfractions and intima-media thickness (IMT) of the common carotid artery (CCA), quantified by B-mode ultrasound, in 94 healthy, 50-year-old men, all of whom were homozygous for the apolipoprotein E3 allele. A novel 3% to 7.5% polyacrylamide gradient gel was developed to provide separation of LDL subfractions with high resolution, as was a procedure to quantify plasma concentrations of these LDL subspecies. The LDL particle size distribution pattern obtained by the gradient gel electrophoresis procedure was in good agreement with the one obtained by a well-established, single-spin density gradient ultracentrifugation technique. LDL II (particle size, 23.5 to 25.0 nm) was the most abundant subfraction, and its plasma concentration correlated closely with the total LDL cholesterol concentration (r=0. 61, P<0.001) but not with CCA IMT (r=-0.13, NS). In contrast, the plasma concentration of the predominant small, dense LDL particle subfraction (LDL-III; particle size, 22.5 to 23.5 nm) correlated strongly with CCA IMT (r=0.42, P<0.001). In multivariate analysis, the plasma concentration of the LDL III subfraction contributed significantly to the variation in CCA IMT (R(2)=0.19). When plasma triglycerides and LDL cholesterol were forced into the multivariate model, 10% of the variation in CCA IMT was still accounted for by the LDL-III subfraction. In summary, use of a novel and sensitive gradient gel electrophoresis method for evaluation of LDL heterogeneity provided the basis for demonstrating an independent relation between the plasma concentration of small LDL and IMT of the CCA in healthy, middle-aged men. PMID- 10521373 TI - Role of plasmalogens in the enhanced resistance of LDL to copper-induced oxidation after LDL apheresis. AB - Extracorporeal reduction of plasma low density lipoproteins (LDLs) by LDL apheresis was shown to attenuate the proatherogenic influences of LDL, such as impairment of vasodilation and increased monocyte adhesion to the endothelium. In 16 patients with familial hypercholesterolemia, we analyzed whether LDL apheresis by the heparin precipitation procedure affected the oxidative resistance of LDL. Plasma LDL cholesterol concentrations were reduced by 65% after the apheresis. The lag time of copper-mediated LDL oxidation was increased from 103 to 117 minutes (P<0.0005). The LDL contents of alpha-tocopherol and beta-carotene, as well as the ratio of monounsaturated to polyunsaturated fatty acids in LDL, were not altered. However, the LDL apheresis induced a 15% increase in the LDL contents of plasmalogen phospholipids (P<0.0005), a class of ether phospholipids that were recently shown to prevent lipid oxidation. The phosphatidylcholine (PC) to lysoPC ratio was elevated by 16% after the apheresis (P<0.0005). The percent increase in LDL plasmalogen phospholipids showed a close association with the increased lag time after apheresis (P<0.0005). The LDL plasmalogen contents of the blood samples from patients and from normolipidemic donors were also positively related to the lag time (P<0.005). In vitro loading of LDL with plasmalogen phospholipids resulted in a prolongation of the lag time and an increase in the PC/lysoPC ratio. In conclusion, the rapid rise in LDL contents of plasmalogen phospholipids most probably causes the increase in lag time after LDL apheresis. Plasmalogens appear to play an important role in the oxidation resistance of LDL in vivo. PMID- 10521374 TI - Human growth hormone increases apo(a) expression in transgenic mice. AB - Levels of Lp(a), an atherogenic lipoprotein that circulates in human plasma, are increased by the administration of growth hormone (GH). Many of the physiological effects of GH are mediated through insulin-like growth factor-1 (IGF-1), but ironically, IGF-1 treatment of humans is associated with a fall in plasma Lp(a) levels. To glean insight into the mechanism responsible for the GH-associated increase in plasma levels of Lp(a), we administered recombinant human GH (rhGH) to mice expressing a 370-kb human genomic fragment containing the apo(a) gene, 40 kb of 5'-, and 200 kb of 3'-flanking sequence [YAC-apo(a) transgenic mice]. The plasma levels of apo(a) and hepatic levels of apo(a) mRNA rose dramatically in the post-pubertal male mice in response to rhGH treatment. To determine whether the increase in plasma apo(a) was mediated by IGF-1, we treated castrated and noncastrated YAC-apo(a) transgenic mice with a continuous infusion of IGF-1 (100 microg/d) for 2 weeks, and plasma levels of apo(a) fell by approximately 50%. Thus the effects of rhGH and IGF-1 administration on plasma levels of apo(a) in the YAC-apo(a) transgenic mice simulate those seen in humans. The coordinate changes in apo(a) mRNA and plasma levels of apo(a) in response to rhGH and IGF-1 strongly suggest that these 2 hormones have independent effects on the transcription of the apo(a) gene. PMID- 10521375 TI - Gender difference in postprandial lipemia : importance of visceral adipose tissue accumulation. AB - Insulin resistance, hyperinsulinemia, hypertriglyceridemia, and low HDL cholesterol concentrations are common features of a plurimetabolic syndrome, which increases the risk of coronary artery disease. Although it has been proposed that the development of atherosclerosis through alterations in plasma lipid levels could be a postprandial phenomenon, most studies on gender differences in plasma lipoprotein-lipid concentrations have reported fasting levels. Therefore, the aim of our study was to examine the response of postprandial triglyceride-rich lipoproteins to a standardized meal in 63 men and 25 women. In addition to the measurement of fasting and postprandial plasma lipid levels, numerous physical and metabolic variables were assessed, including body composition by underwater weighing and body fat distribution by computed tomography. Although no gender difference was noted in total body fat mass, men were characterized by a preferential accumulation of abdominal adipose tissue as revealed by an increased waist circumference and a greater visceral adipose tissue accumulation (50% difference) compared with women (P<0.001). Men also showed a greater plasma triglyceride response (P<0.005) as well as increased postprandial insulin and free fatty acid levels compared with women (P<0.01). Visceral adipose tissue was significantly associated with the postprandial triglyceride response in both genders (men: r=0.49, P<0. 0001; women: r=0.43, P<0.05). Finally, when men and women were matched for visceral adipose tissue accumulation, the gender difference in postprandial plasma triglyceride response was eliminated. Thus results of the present study suggest that the well known gender difference in visceral adipose tissue accumulation is an important contributing factor involved in the exaggerated postprandial triglyceride-rich lipoprotein response noted in men compared with women. PMID- 10521376 TI - Pleiotropic genetic effects on LDL size, plasma triglyceride, and HDL cholesterol in families. AB - The interrelationships among low density lipoprotein (LDL) particle size, plasma triglyceride (TG), and high density lipoprotein cholesterol (HDL-C) are well established and may involve underlying genetic influences. This study evaluated common genetic effects on LDL size, TG, and HDL-C by using data from 85 kindreds participating in the Genetic Epidemiology of Hypertriglyceridemia (GET) Study. A multivariate, maximum likelihood-based approach to quantitative genetic analysis was used to estimate the additive effects of shared genes and shared, unmeasured nongenetic factors on variation in LDL size and in plasma levels of TG and HDL-C. A significant (P<0.001) proportion of the variance in each trait was attributable to the additive effects of genes. Maximum-likelihood estimates of heritability were 0.34 for LDL size, 0.41 for TG, and 0.54 for HDL-C. Significant (P<0.001) additive genetic correlations (rho(G)), indicative of the shared additive effects of genes on pairs of traits, were estimated between all 3 trait pairs: for LDL size and TG rho(G)=-0.87, for LDL size and HDL-C rho(G)=0.65, and for HDL-C and TG rho(G)=-0.54. A similar pattern of significant environmental correlations between the 3 trait pairs was also observed. These results suggest that a large proportion of the well-documented correlations in LDL size, TG, and HDL-C are likely attributable to the influence of the same gene(s) in these families. That is, the gene(s) that may contribute to decreases in LDL size also contribute significantly to higher plasma levels of TG and lower plasma levels of HDL-C. These relationships may be useful in identifying genes responsible for the associations between these phenotypes and susceptibility to cardiovascular disease in these families. PMID- 10521377 TI - Independent effects of Apo E phenotype and plasma triglyceride on lipoprotein particle sizes in the fasting and postprandial states. AB - LDL particle sizes and Apo E phenotypes were determined in 212 subjects of whom 51 had angina. LDL diameter was significantly less in subjects with an epsilon2 allele (24.76+/-0.08 vs 24.94+/-0.02 nm, P=0.02), and this was evident for both E2/E3 (24.77+/-0.09 nm) and E2/E4 (24.69+/-0.08 nm) phenotypes. Although there was a negative relation between LDL diameter and plasma triglyceride, the effect of apo E2 was still evident with adjustment for triglyceride. In multiple regression analysis, the significant determinants of LDL diameter were gender (with females having larger particles than males), body mass index, and the presence (or absence) of E2. HDL particle sizes and compositions were determined on fasting samples and, additionally, 5 and 8 hours after a fat-rich meal for 48 coronary heart disease cases and 49 control subjects. Fasting HDL particle sizes were not related to the presence of E2 but were significantly smaller for subjects possessing an epsilon4 allele (8. 09+/-0.08 vs 8.39+/-0.05 nm, P=0.003) and were negatively related to plasma triglyceride. However, the effect of E4 persisted after adjustment for triglyceride. In a multiple regression analysis, the only significant determinant of fasting HDL diameter was the presence (or absence) of E4 with fasting plasma triglyceride just failing to reach significance (P=0.06). There was a postprandial increase in HDL diameter that was less marked in subjects with coronary heart disease. The postprandial increase in HDL diameter was of sufficient magnitude to result in size reclassification of HDL particles. The influence of E4 was also evident at both postprandial time points. Compositional analysis demonstrated that the increase in HDL diameters postprandially could be attributed to triglyceride enrichment, with an accompanying fall in cholesterol ester content. Phospholipid changes postprandially were biphasic with an initial fall followed by a rise in concentration. The increase in triglyceride content was significantly less in those subjects with angina despite an equivalent rise in plasma triglyceride. The present study demonstrates significant, but different, effects of variation in apo E phenotype on the particle sizes of both HDL and LDL. Such effects were still evident with adjustment for differences in plasma triglyceride and suggests that variation in apo E phenotype exerts effects on lipoprotein particle sizes by mechanisms additional to those dependent on change in plasma triglyceride. PMID- 10521378 TI - Detection, quantification, and characterization of potentially atherogenic triglyceride-rich remnant lipoproteins. AB - Triglyceride-rich lipoprotein (TRL) remnants are formed in the circulation when apolipoprotein (apo) B-48-containing chylomicrons of intestinal origin or apoB 100-containing VLDL of hepatic origin are converted by lipoprotein lipase, and to a lesser extent by hepatic lipase, into smaller and more dense particles. Compared with their nascent precursors, TRL remnants are depleted of triglyceride, phospholipid, and C apolipoproteins and are enriched in cholesteryl esters and apoE. They can thus be identified, separated, and/or quantified in plasma according to their density, charge, size, specific lipid components, apolipoprotein composition, and/or apolipoprotein immunospecificity. Each of these approaches has contributed to our current understanding of the compositional characteristics of TRL remnants and their potential to promote atherosclerosis. An ongoing search is nevertheless under way for more accurate and clinically applicable remnant lipoprotein assays that will be able to better define coronary artery disease risk in patients with hypertriglyceridemia. PMID- 10521379 TI - Westernization of Chinese adults and increased subclinical atherosclerosis. AB - Cardiovascular event rates are much lower in China compared with developed countries. "Westernization" of diet and lifestyle in the Chinese, however, may lead to an increased prevalence of atherosclerosis-related diseases. Because carotid intima-media thickness (IMT) is a marker of subclinical atherosclerosis, we examined IMT and vascular risk profile in community-based groups of rural Chinese, Westernized urban Chinese, and urban whites. Mean IMT of the common carotid artery was measured in 348 healthy adults, aged 42+/-13 years (range 21 to 71 years); 116 subjects from rural China, 116 urban Chinese subjects living in Hong Kong or in Australia, and 116 urban Caucasians living in Australia. These 3 groups were matched for age, sex, and cigarette smoke exposure. Urban Chinese subjects had slightly better risk factor profile (higher HDL-cholesterol and lower blood pressure) compared with rural Chinese subjects. Despite this, however, the mean IMT was lowest in rural Chinese (0.50+/-0.10 mm), intermediate in urban Chinese (0.56+/-0.12 mm), and highest in urban whites (0.64+/-0.13 mm) (P<0.001 for comparisons between all groups). These differences in IMT were not altered after adjustment for the major traditional cardiovascular risk factors (serum lipids, smoking, and blood pressure or for body mass index). The influence of vascular risk factors on atherosclerosis between urban versus rural Chinese subjects was studied by multivariate regression models and by comparing the steepness of regression slopes between risk factors and IMT in the subject groups. The effects of smoking, HDL-cholesterol, and triglycerides on IMT were significantly greater in the urban compared with the rural Chinese (P<0.01). These data suggest that Westernization of Chinese subjects is associated with greater susceptibility to the pro-atherogenic effects of traditional vascular risk factors, such as lipids and smoking, and with evidence of increased IMT as a marker of subclinical atherosclerosis. PMID- 10521380 TI - Anderson's disease: exclusion of apolipoprotein and intracellular lipid transport genes. AB - Anderson's disease is a rare, hereditary hypocholesterolemic syndrome characterized by chronic diarrhea, steatorrhea, and failure to thrive associated with the absence of apo B48-containing lipoproteins. To further define the molecular basis of the disease, we studied 8 affected subjects in 7 unrelated families of North African origin after treatment with a low-fat diet. Lipid loading of intestinal biopsies persisted, but the pattern and extent of loading was variable among the patients. Electron microscopy showed lipoprotein-like particles in membrane-bound compartments, the densities (0.65 to 7.5 particles/mu(2)) and the mean diameters (169 to 580 nm) of which were, in general, significantly larger than in a normal fed subject (0.66 particles/mu(2), 209 nm mean diameter). There were also large lipid particles having diameters up to 7043 nm (average diameters from 368 to 2127 nm) that were not surrounded by a membrane. Rarely, lipoprotein-like particles 50 to 150 nm in diameter were observed in the intercellular spaces. Intestinal organ culture showed that apo B and apo AIV were synthesized with apparently normal molecular weights and that small amounts were secreted in lipid-bound forms (density <1.006 g/mL). Normal microsomal triglyceride transfer protein (MTP) and activity were also detected in intestinal biopsies. Segregation analyses of 4 families excluded, as a cause of the disease, significant regions of the genome surrounding the genes for apo AI, AIV, B, CI, CII, CIII, and E, as were the genes encoding 3 proteins involved in intracellular lipid transport, MTP, and fatty acid binding proteins 1 and 2. The results suggest that a factor other than apoproteins and MTP are important for human intestinal chylomicron assembly and secretion. PMID- 10521381 TI - Impaired endothelium-dependent vascular responses of retinal and intrarenal arteries in patients with type 2 diabetes. AB - Endothelial dysfunction has been implicated in the pathogenesis of diabetic microangiopathies such as retinopathy and nephropathy as well as macrovascular diseases. The aim of the current study was to determine whether endothelial function in the retinal and renal arteries is impaired in type 2 diabetes mellitus. We examined the effects of an intravenous infusion of L-arginine and a sublingual administration of nitroglycerin on the brachial, retinal, and interlobar arterial hemodynamics in 20 type 2 diabetic patients (10 with normoalbuminuria and 10 with microalbuminuria) and 10 aged-matched control subjects. Despite no difference in the nitroglycerin-induced vascular response of the brachial or retinal artery among the 3 groups, the L-arginine-induced vascular response of each artery was significantly lower in both the normoalbuminuric and microalbuminuric patients than in the control subjects and the microalbuminuric patients showed the lowest value among the 3 groups (P<0.01, each artery, respectively). The L-arginine-induced vascular response of each artery was significantly correlated with HbA1c levels (brachial artery, r=0.617, P=0.0003; retinal artery, r=0.599, P=0.0005; interlobar artery, r=0.636, P=0.0002). In addition, stepwise multiple regression analysis of all subjects showed that HbA1c level was an independent determinant for the L-arginine-induced vascular response of each artery. The results showed that the endothelium dependent vascular responses of the retinal and intrarenal arteries as well as the brachial artery were impaired in diabetic patients before the clinical manifestation of diabetic nephropathy, and suggest that endothelial dysfunction in these arteries is associated with hyperglycemia in these patients. PMID- 10521382 TI - Endotoxin-induced activation of the coagulation cascade in humans: effect of acetylsalicylic acid and acetaminophen. AB - During Gram-negative septic shock, lipopolysaccharide (LPS, endotoxin) induces tissue factor (TF) expression. TF expression is mediated by nuclear factor kappaB and amplified by activated platelets. TF forms a highly procoagulant complex with activated coagulation factor VII (FVIIa). Hence, we hypothesized that aspirin, which inhibits LPS-induced, nuclear factor kappaB-dependent TF expression in vitro and platelet activation in vivo, may suppress LPS-induced coagulation in humans. Therefore, we studied the effects of aspirin on systemic coagulation activation in the established and controlled setting of the human LPS model. Thirty healthy volunteers were challenged with LPS (4 ng/kg IV) after intake of either placebo or aspirin (1000 mg). Acetaminophen (1000 mg) was given to a third group to control for potential effects of antipyresis. Neither aspirin nor acetaminophen inhibited LPS-induced coagulation. However, LPS increased the percentage of circulating TF(+) monocytes by 2-fold. This increase was associated with a decrease in FVIIa levels, which reached a minimum of 50% 24 hours after LPS infusion. Furthermore, LPS-induced thrombin generation increased plasma levels of circulating polymerized, but not cross-linked, fibrin (ie, thrombus precursor protein), whereas levels of soluble fibrin were unaffected. In summary, a single 1000-mg dose of aspirin did not decrease LPS-induced coagulation. However, our study showed, for the first time, that LPS increases TF(+) monocytes, substantially decreases FVIIa levels, and enhances plasma levels of thrombus precursor protein, which may be a useful marker of fibrin formation in humans. PMID- 10521383 TI - Intimal deposition of functional von Willebrand factor in atherogenesis. AB - During the formation of intimal thickening in normocholesterolemic rabbits, von Willebrand factor (vWF) is increased in the endothelial cells (ECs) and deposited in the intima. We investigated whether this also occurs during cholesterol induced plaque formation, whether the synthesis of vWF increases, and whether this influences platelet adhesion. Rabbits were fed a cholesterol-rich (0.3%) diet for 26 weeks. Thereafter, half of the animals received a normal diet for another 26 weeks (cholesterol withdrawal). To induce intimal thickening in normocholesterolemic rabbits, collars were positioned around the carotid artery. Arterial segments were studied using immunohistochemistry, reverse transcription polymerase chain reaction, electron microscopy, and platelet adhesion tests. Cholesterol treatment induced plaque formation in the aorta. The ECs had a cuboidal aspect, showed a dense immunoreactivity for vWF, a pronounced rough endoplasmic reticulum, and numerous Weibel-Palade bodies. There were subendothelial vWF deposits in the plaques and vWF mRNA was significantly increased as compared with controls. Similar changes were seen after collar induced intimal thickening. After cholesterol withdrawal, both vWF mRNA and the ultrastructural morphology of the ECs normalized, and the vWF deposits disappeared from the plaque. Perfusion studies with anticoagulated rabbit blood over cross-sections of atherosclerotic aortas revealed increased vWF-mediated platelet adhesion in the subendothelial plaque region. Whereas rabbit platelets perfused through the lumen adhered to the same extent to de-endothelialized aortas of normocholesterolemic and atherosclerotic rabbits, vWF mediated platelet adhesion to endothelium was observed in atherosclerotic but not in normal aortas. Our results show an increased synthesis and (sub)endothelial presence of vWF after vascular injury, with functional consequences for platelet deposition on the vessel wall. PMID- 10521384 TI - Intravenous and oral antithrombotic efficacy of the novel platelet GPIIb/IIIa antagonist roxifiban (DMP754) and its free acid form, XV459. AB - Currently used antiplatelet drugs, including aspirin, ticlopidine, and others, are effective against certain but not all of the many endogenous platelet activators. Because of their limited efficacy, a significant number of serious thromboembolic complications still occur, highlighting the need for a more effective therapy. DMP754 (roxifiban), a prodrug of XV459, is a recently discovered, potent antiplatelet agent with high affinity and specificity for platelet GPIIb/IIIa receptors that blocks platelet aggregate formation regardless of the agonist (IC(50)=0.030 to 0.05 micromol/L) or anticoagulant used for blood collection. DMP754 rapidly converts to its active free-acid form, XV459, which has a comparable high affinity for both resting and activated platelets (K(d)=1 to 2 nmol/L) and a relatively slow rate of dissociation from resting platelets. The present study was undertaken to determine intravenous and oral antithrombotic efficacies of DMP754 and XV459 and to compare them with those of other antiplatelet and anticoagulant agents in canine models of arterial thrombosis. In these models, thrombosis was induced either electrolytically (200-microA anodal current) in the carotid artery or mechanically by external clamping of the femoral artery along with stenosis, which resulted in either total occlusive thrombus formation or cyclic flow reduction, respectively. DMP754 and XV459 were given either intravenously (0.1 mg/kg bolus) or orally (0.1 to 0.4 mg/kg). Additionally, the antithrombotic efficacies of DMP754, aspirin, heparin, and ticlopidine in the canine carotid artery electrolytic injury model were compared. DMP754 demonstrated oral bioavailability of 20.8% in dogs after administration at different doses and prevented cyclic flow reduction (ED(90-100)=<0.1 mg/kg IV or PO). Additionally, both DMP754 and XV459 (0.1 mg/kg IV or 0.3 to 0.4 mg/kg PO) demonstrated maximal antithrombotic efficacy in preventing electrically induced carotid and coronary artery thrombosis and significant antithrombotic efficacy (P<0.001) at relatively low doses in different settings of arterial thrombosis in the canine model. DMP754 resulted in a significant reduction in thrombus mass and sustained arterial blood flow with 100% prevention of occlusive and nonocclusive thrombosis. In contrast, administration of aspirin (10 mg/kg PO for 2 days), heparin (10 IU/kg IV bolus followed by 90 IU/kg IV infusion over 3 hours), or ticlopidine (300 mg/kg PO for 3 days) before initiation of arterial thrombosis did not reduce the incidence of electrolytic injury-induced occlusive arterial thrombosis. These studies demonstrated a distinct antithrombotic efficacy of DMP754 as compared with existing strategies and suggest potential intravenous and oral antithrombotic uses of DMP754 in the prevention and treatment of thromboembolic disorders. PMID- 10521385 TI - Vitamin E inhibits collagen-induced platelet activation by blunting hydrogen peroxide. AB - In this study, we investigated whether vitamin E at concentrations achievable in blood after supplementation inhibits platelet function in humans. Gel-filtered platelets were incubated 30 minutes with scalar concentrations (50 to 250 mmol/L) of vitamin E and then stimulated with collagen. Compared with controls, vitamin E inhibited collagen-induced platelet aggregation and thromboxane A2 formation in a dose-dependent manner. Furthermore, vitamin E inhibited, in a dose-dependent manner, Ca(2+) mobilization and formation of inositol 1,4,5-triphosphate. Because it was previously shown that hydrogen peroxide formation mediates arachidonic acid metabolism and phospholipase C activation in collagen-induced platelet activation, we investigated whether vitamin E was able to blunt hydrogen peroxide. In experiments performed in unstimulated platelets supplemented with hydrogen peroxide and in collagen-stimulated platelets, vitamin E was able to blunt hydrogen peroxide. In 6 healthy subjects given vitamin E for 2 weeks (600 mg/d), we found a significant decrease of collagen-induced H(2)O(2) formation, platelet aggregation, and calcium mobilization. This study demonstrated in vitro and ex vivo that vitamin E inhibits collagen-induced platelet activation by blunting hydrogen peroxide formation. PMID- 10521386 TI - Activation of the contact system of coagulation does not contribute to the hemostatic imbalance in hypertriglyceridemia. AB - In vitro, triglyceride-rich lipoproteins may act as a surface to initiate the contact system of coagulation. Therefore, we studied the activation of factor XII (FXII), prekallikrein, and FXI and the generation of thrombin in 52 hypertriglyceridemic patients before and after 12 weeks of triglyceride-lowering treatment with gemfibrozil or n-3 polyunsaturated fatty acids. Thrombin generation was assessed by measuring the levels of prothrombin fragment F1+2 and thrombin-antithrombin (TAT) complexes. Contact activation was assessed by measuring FXIIa, kallikrein, and FXIa in complex with their major inhibitor, C1 inhibitor, and FXIa was also determined as part of a complex with alpha(1) antitrypsin. Triglyceride and cholesterol levels decreased equally in both treatment groups. In the gemfibrozil group, there was a significant decrease in F1+2, while TAT complexes did not change. FXIIa- and kallikrein-C1 inhibitor complexes were elevated in 13% and 9% of the patients before treatment, respectively, and no changes were observed on triglyceride-lowering therapy. Also, no significant changes in regard to FXIa-C1 inhibitor and FXIa-alpha(1) antitrypsin complexes were seen. FXIa-alpha(1)-antitrypsin complexes were present in 70% of the patients before therapy and were positively correlated with the level of TAT complexes. In conclusion, we did not detect an effect on activation markers of the contact coagulation system in hypertriglyceridemic patients after triglyceride-lowering therapy. Therefore, contact activation is not likely to contribute to the hypercoagulability seen in these patients. PMID- 10521387 TI - Antithrombotic efficacy of a novel murine antihuman factor IX antibody in rats. AB - A murine antihuman factor IX monoclonal antibody (BC2) has been generated and evaluated for its capacity to prolong the activated partial thromboplastin time (aPTT) in vitro and ex vivo and to prevent arterial thrombosis in a rat model in vivo. BC2 extended aPTT to a maximum of 60 to 80 seconds at 100 to 1000 nmol/L in vitro (rat and human plasma, respectively) and ex vivo (rat) after dosing of rats up to 6 mg/kg in vivo. BC2, administered as bolus (1 to 6 mg/kg) followed by infusion (0.3 to 2 mg x kg(-1) x h(-1)), dose-dependently prevented thrombosis of an injured rat carotid artery (FeCl(3)-patch model), increased time to artery occlusion, and reduced incidence of vessel occlusion. BC2 efficacy in preventing arterial thrombosis exceeded that of heparin (bolus 15 to 120 U/kg followed by infusion 0.5 to 4.0 U x kg(-1) x min(-1)), whereas the latter rendered the blood incoagulable (aPTT>1000 seconds). BC2 demonstrated complete antithrombotic efficacy also as a single bolus given either as prevessel or postvessel injury as evidenced by reduction of thrombus mass (from 4.18+/-0.49 to 1.80 +/-0.3 mg, P<0.001), increasing vessel patency time (from 14.9+/-0.9 minutes to 58.3+/-1.7 minutes, P<0.001) and decreasing incidence of vessel occlusion from 100% to 0% in vehicle- versus BC2-treated rats, respectively. BC2 (3 mg/kg, IV) administered in a single bolus resulted in 50% reduction in thrombus mass (P<0.01), extended vessel patency time (P<0.001), extended aPTT only 4-fold, and had no effect on blood loss via a tail surgical wound; heparin, at doses that reduced thrombus mass to a similar extent, extended aPTT beyond 1000 seconds (over 500-fold) and increased blood loss from 1.8+/-0.7 to 3.3 +/-0.6 mL (P<0.001). These data suggest that BC2 may provide enhanced therapeutic efficacy in humans at lesser interference with blood hemostasis than heparin. PMID- 10521388 TI - Structural requirements for TFPI-mediated inhibition of neointimal thickening after balloon injury in the rat. AB - The intimal thickening that follows vascular injury is inhibited by periprocedural tissue factor pathway inhibitor (TFPI) treatment in animal models. TFPI is a multivalent Kunitz-type protease inhibitor that inhibits factor Xa via its second Kunitz domain and the factor VIIa/tissue factor (TF) complex via its first Kunitz domain. The basic C-terminus of TFPI is required for the binding of TFPI to cell surfaces and cell-bound TFPI mediates the internalization and degradation of factor X and the down regulation of surface factor VIIa/TF activity. The C-terminus of TFPI is also required for its reported direct inhibition of smooth muscle cell proliferation in vitro. To examine the structural requirements for the inhibition of neointimal formation by TFPI, several TFPI-related proteins were tested in the rat carotid angioplasty model: 1) XK(1), a hybrid protein containing the N-terminal portion of factor X and the first Kunitz domain of TFPI that directly inhibits factor VIIa/TF; 2) TFPI(WT), the full-length TFPI molecule that inhibits factor Xa and factor VIIa/TF and binds cell surfaces; 3) TFPI(K36I), an altered form of TFPI that inhibits factor Xa, but not factor VIIa/TF, and binds cell surfaces; 4) TFPI(13-161), a truncated form of TFPI that inhibits factor VIIa/TF but interacts with factor Xa poorly and does not bind to cell surfaces. Seven day infusions of XK(1), TFPI(WT), and high levels of TFPI(K36I) begun the day before balloon-induced vascular injury produced a significant reduction in the intimal hyperplasia measured 28 days after angioplasty. The infusion of high concentrations of TFPI(13-161) was ineffective in this model. These in vivo results directly mirror the ability of each TFPI-related protein to inhibit tissue thromboplastin-induced coagulation in rat plasma: XK(1) approximately TFPI(WT)>TFPI(K36I)>>TFPI(13-161). The studies confirm the important role of TF-mediated coagulation in the smooth muscle proliferation and neointimal thickening that follows vascular injury and suggest that the anticoagulant effect alone of TFPI and TFPI-related proteins is sufficient to explain their therapeutic action. PMID- 10521389 TI - Prothrombin G20210A gene mutation and further prothrombotic risk factors in childhood thrombophilia. AB - Risk factors for venous thrombosis in adults are the prothrombin G20210A and the factor V (FV) G1691A mutations and hereditary deficiencies of protein C, protein S and antithrombin. However, data are limited on the relevance of these risk factors for thrombosis in children and adolescents. We therefore investigated 261 patients aged 0 to 18 (median 5.7 years, 48% male) with venous thrombosis and controls (n=370) for the presence of prothrombotic risk factors including the prothrombin G20210A mutation. The following frequencies of hereditary risk factors (patients versus controls), odds ratios (OR) and 95% confidence intervals (CI), or results of Fisher's exact test, respectively, were found: prothrombin G20210A, 4.2% versus 1.1%, OR/CI 4.1/1.3 to 12.8; FV G1691A, 31.8% versus 4. 1%, OR/CI 11.0/6.2 to 19.7; protein C deficiency, 9.2% versus 0.8%, OR/CI 12.4/3.7 to 41.6, protein S deficiency, 5.7% versus 0.8%, OR/CI 7.5/2.1 to 26.0; antithrombin deficiency in 3.4% in the patients, but not in the controls, P=0.0003. The prothrombin mutation was combined with the heterozygous FV G1691A mutation (2. 3%) or protein C deficiency (0.3%) in the patients, but not in the controls (prothrombin and FV mutation, P=0.0048; prothrombin and protein C deficiency, not significant). The carrier frequencies and ORs of all hereditary risk factors showed a non-significant trend toward higher prevalences in patients suffering spontaneous thrombosis, compared with those with an additional underlying disease. In conclusion, the prothrombin G20210A and the FV G1691A mutation, deficiencies of protein C, protein S, and antithrombin are important risk factors for venous thrombosis during childhood and adolescence. PMID- 10521390 TI - Glycoprotein IIIa Pl(A) polymorphism associates with progression of coronary artery disease and with myocardial infarction in an autopsy series of middle-aged men who died suddenly. AB - Glycoprotein IIIa (GPIIIa) has a key role in the aggregation of thrombocytes, and it also mediates intimal hyperplasia after endothelial injuries; the possible association of the Pl(A1/A2) polymorphism of the gene for GPIIIa with coronary thrombosis and with the progression of coronary artery disease (CAD) is still to be confirmed. Therefore, the association of the Pl(A) polymorphism with the development of coronary atherosclerosis, coronary narrowing, and myocardial infarction (MI) was studied in a prospective, consecutive autopsy series of 300 middle-aged, white Finnish men (33 to 69 years) suffering sudden out-of-hospital or violent death. Coronary atherosclerosis was measured morphometrically and the coronary stenosis percentage determined from a cast rubber model of the coronary tree. We found a significant inverse relation (P=0.01) between the Pl(A2) positive genotype and coronary artery stenosis. The frequency of possessing the Pl(A2) allele was significantly (odds ratio [OR] 0.45, 95% confidence interval [CI] 0.22 to 0.98) lower among men with >50% coronary stenosis (18.3%) than among those with <25% stenosis (32.9%). Although the Pl(A) polymorphism was not directly associated with MI, the Pl(A2) allele was present in 11 of the 22 men (50%) with MI and coronary thrombosis (OR 6.6, 95% CI 2.1 to 22.8) but in only 6 of the 47 (12.8%) with MI associated with severe stenosis in the absence of thrombosis. In line with this result, men possessing the Pl(A2) allele also had a larger area of fissured and ulcerated complicated lesions in their coronary arteries (P<0.05). The present results suggest that the Pl(A) polymorphism is involved in the development of CAD and MI. Men with the Pl(A2) allele may harbor more thin-walled, vulnerable coronary plaques, plaques prone to rupture, leading to massive, fatal thrombosis. In contrast, men homozygous for the Pl(A1) allele may more often show stable plaques and present with infarction caused by progressive coronary stenosis. PMID- 10521391 TI - Looping versus linking: toward a model for long-distance gene activation. PMID- 10521392 TI - Multilineage embryonic hematopoiesis requires hypoxic ARNT activity. AB - Although most cells undergo growth arrest during hypoxia, endothelial cells and placental cytotrophoblasts proliferate in response to low O(2). We demonstrate that proliferation of embryonic multilineage hematopoietic progenitors is also regulated by a hypoxia-mediated signaling pathway. This pathway requires HIF-1 (HIF-1alpha/ARNT heterodimers) because Arnt(-/-) embryoid bodies fail to exhibit hypoxia-mediated progenitor proliferation. Furthermore, Arnt(-/-) embryos exhibit decreased numbers of yolk sac hematopoietic progenitors. This defect is cell extrinsic, is accompanied by a decrease in ARNT-dependent VEGF expression, and is rescued by exogenous VEGF. Therefore, "physiologic hypoxia" encountered by embryos is essential for the proliferation or survival of hematopoietic precursors during development. PMID- 10521393 TI - TFIID-specific yeast TAF40 is essential for the majority of RNA polymerase II mediated transcription in vivo. AB - Many questions remain concerning the role of TFIID TBP-associated factors (TAFs) in transcription, including whether TAFs are required at most or only a small subset of promoters. It was shown previously that three histone-like TAFs are broadly required for transcription, but interpretation of this observation is complicated because these proteins are components of both TFIID and the SAGA histone acetyltransferase complex. Here we show that mutations in the yeast TFIID specific protein Taf40 lead to a general cessation of transcription, even in the presence of excess TBP, suggesting that the TFIID complex is required at most promoters in vivo. PMID- 10521394 TI - Transcriptional repression by wild-type p53 utilizes histone deacetylases, mediated by interaction with mSin3a. AB - There is growing evidence that the p53 tumor suppressor protein not only can function to activate gene transcription but also to repress the expression of specific genes. Although recent studies have implicated the transcriptional repression function of p53 in the pathway of apoptosis, the molecular basis of this activity remains poorly understood. This study takes a first step toward elucidating this mechanism. We report that trichostatin A (TSA), an inhibitor of histone deacetylases (HDACs), abrogates the ability of p53 to repress the transcription of two genes that it negatively regulates, Map4 and stathmin. Consistent with this finding, we report that p53 physically associates in vivo with HDACs. This interaction is not direct but, rather, is mediated by the corepressor mSin3a. Both wild-type p53 and mSin3a, but not mutant p53, can be found bound to the Map4 promoter at times when this promoter preferentially associates with deacetylated histones in vivo. Significantly, inhibition of p53 mediated transcriptional repression with TSA markedly inhibits apoptosis induction by p53. These data offer the first mechanistic insights for p53 mediated transcriptional repression and underscore the importance of this activity for apoptosis induction by this protein. PMID- 10521395 TI - Functional interactions between a phage histone-like protein and a transcriptional factor in regulation of phi29 early-late transcriptional switch. AB - Protein p6 is a nonspecific DNA-binding protein occurring in high abundance in phage phi29-infected cells. Here, we demonstrate a novel role for this versatile histone-like protein: its involvement in regulating the viral switch between early and late transcription. p6 performs this role by exhibiting a reciprocal functional interaction with the regulatory protein p4, also phage encoded, which is required for repression of the early A2b and A2c promoters and activation of the late A3 promoter. On the one hand, p6 promotes p4-mediated repression of the A2b promoter and activation of the A3 promoter by enhancing binding of p4 to its recognition site at PA3; on the other, p4 promotes p6-mediated repression of the A2c promoter by favoring the formation of a stable p6-nucleoprotein complex that interferes with RNA polymerase binding to PA2c. We propose that the observed interplay between proteins p6 and p4 is based on their DNA architectural properties. PMID- 10521396 TI - Cleavage of the death domain kinase RIP by caspase-8 prompts TNF-induced apoptosis. AB - Although the molecular mechanisms of TNF signaling have been largely elucidated, the principle that regulates the balance of life and death is still unknown. We report here that the death domain kinase RIP, a key component of the TNF signaling complex, was cleaved by Caspase-8 in TNF-induced apoptosis. The cleavage site was mapped to the aspartic acid at position 324 of RIP. We demonstrated that the cleavage of RIP resulted in the blockage of TNF-induced NF kappaB activation. RIPc, one of the cleavage products, enhanced interaction between TRADD and FADD/MORT1 and increased cells' sensitivity to TNF. Most importantly, the Caspase-8 resistant RIP mutants protected cells against TNF induced apopotosis. These results suggest that cleavage of RIP is an important process in TNF-induced apoptosis. Further more, RIP cleavage was also detected in other death receptor-mediated apoptosis. Therefore, our study provides a potential mechanism to convert cells from life to death in death receptor mediated apoptosis. PMID- 10521397 TI - Conserved requirement for EGF-CFC genes in vertebrate left-right axis formation. AB - Specification of the left-right (L-R) axis in the vertebrate embryo requires transfer of positional information from the node to the periphery, resulting in asymmetric gene expression in the lateral plate mesoderm. We show that this activation of L-R lateral asymmetry requires the evolutionarily conserved activity of members of the EGF-CFC family of extracellular factors. Targeted disruption of murine Cryptic results in L-R laterality defects including randomization of abdominal situs, hyposplenia, and pulmonary right isomerism, as well as randomized embryo turning and cardiac looping. Similarly, zebrafish one eyed pinhead (oep) mutants that have been rescued partially by mRNA injection display heterotaxia, including randomization of heart looping and pancreas location. In both Cryptic and oep mutant embryos, L-R asymmetric expression of Nodal/cyclops, Lefty2/antivin, and Pitx2 does not occur in the lateral plate mesoderm, while in Cryptic mutants Lefty1 expression is absent from the prospective floor plate. Notably, L-R asymmetric expression of Nodal at the lateral edges of the node is still observed in Cryptic mutants, indicating that L R specification has occurred in the node but not the lateral plate. Combined with the previous finding that oep is required for nodal signaling in zebrafish, we propose that a signaling pathway mediated by Nodal and EGF-CFC activities is essential for transfer of L-R positional information from the node. PMID- 10521398 TI - Peripheral nervous system defects in erbB2 mutants following genetic rescue of heart development. AB - The ErbB2 tyrosine kinase functions as coreceptor for the neuregulin receptors ErbB3 and ErbB4 and can participate in signaling of EGF receptor (ErbB1), interleukin receptor gp130, and G-protein coupled receptors. ErbB2(-/-) mice die at midgestation because of heart malformation. Here, we report a genetic rescue of their heart development by myocardial expression of erbB2 cDNA that allows survival of the mutants to birth. In rescued erbB2 mutants, Schwann cells are lacking. Motoneurons form and can project to muscle, but nerves are poorly fasciculated and disorganized. Neuromuscular junctions form, as reflected in clustering of AChR and postsynaptic expression of the genes encoding the alpha AChR, AChE, epsilon-AChR, and the RI subunit of the cAMP protein kinase. However, a severe loss of motoneurons on cervical and lumbar, but not on thoracic levels occurs. Our results define the roles of Schwann cells during motoneuron and synapse development, and reveal different survival requirements for distinct motoneuron populations. PMID- 10521399 TI - Genes regulating dendritic outgrowth, branching, and routing in Drosophila. AB - Signaling between neurons requires highly specialized subcellular structures, including dendrites and axons. Dendrites exhibit diverse morphologies yet little is known about the mechanisms controlling dendrite formation in vivo. We have developed methods to visualize the stereotyped dendritic morphogenesis in living Drosophila embryos. Dendrite development is altered in prospero mutants and in transgenic embryos expressing a constitutively active form of the small GTPase cdc42. From a genetic screen, we have identified several genes that control different aspects of dendrite development including dendritic outgrowth, branching, and routing. These genes include kakapo, a large cytoskeletal protein related to plectin and dystrophin; flamingo, a seven-transmembrane protein containing cadherin-like repeats; enabled, a substrate of the tyrosine kinase Abl; and nine potentially novel loci. These findings begin to reveal the molecular mechanisms controlling dendritic morphogenesis. PMID- 10521400 TI - A PP2A regulatory subunit positively regulates Ras-mediated signaling during Caenorhabditis elegans vulval induction. AB - We describe evidence that a regulatory B subunit of protein phosphatase 2A (PP2A) positively regulates an RTK-Ras-MAP kinase signaling cascade during Caenorhabditis elegans vulval induction. Although reduction of sur-6 PP2A-B function causes few vulval induction defects in an otherwise wild-type background, sur-6 PP2A-B mutations suppress the Multivulva phenotype of an activated ras mutation and enhance the Vulvaless phenotype of mutations in lin-45 raf, sur-8, or mpk-1. Double mutant analysis suggests that sur-6 PP2A-B acts downstream or in parallel to ras, but likely upstream of raf, and functions with ksr-1 in a common pathway to positively regulate Ras signaling. PMID- 10521401 TI - The SIR2/3/4 complex and SIR2 alone promote longevity in Saccharomyces cerevisiae by two different mechanisms. AB - The SIR genes are determinants of life span in yeast mother cells. Here we show that life span regulation by the Sir proteins is independent of their role in nonhomologous end joining. The short life span of a sir3 or sir4 mutant is due to the simultaneous expression of a and alpha mating-type information, which indirectly causes an increase in rDNA recombination and likely increases the production of extrachromosomal rDNA circles. The short life span of a sir2 mutant also reveals a direct failure to repress recombination generated by the Fob1p mediated replication block in the rDNA. Sir2p is a limiting component in promoting yeast longevity, and increasing the gene dosage extends the life span in wild-type cells. A possible role of the conserved SIR2 in mammalian aging is discussed. PMID- 10521402 TI - Meiotic DNA replication checkpoint control in fission yeast. AB - In eukaryotes, the DNA replication checkpoint prevents entry into mitosis when DNA replication is incomplete and is crucial for maintaining genomic integrity. Much less is known about equivalent controls that operate during meiosis. Here, we show that a DNA replication checkpoint control operates during meiosis in fission yeast. The mitotic checkpoint Rad genes and the Cds1 protein kinase are required for the DNA replication checkpoint during meiosis, with Cds1 playing a more prominent role than it does during mitosis. When DNA replication is blocked, the checkpoint maintains Cdc2 tyrosine 15 phosphorylation keeping Cdc2 protein kinase activity low and preventing onset of meiosis I. Additionally, there is a second checkpoint acting during meiosis that is revealed if cells are prevented from maintaining Cdc2 tyrosine 15 phosphorylation when DNA replication is blocked. Such cells arrest with high Cdc2 protein kinase activity and separated spindle pole bodies, an arrest state similar to that observed in mitotic budding yeast cells when DNA replication is incomplete. This second checkpoint is meiosis specific and may reflect processes occurring only during meiosis such as increased recombination rates, an extended duration of nuclear division, or homolog chromosome pairing. PMID- 10521403 TI - Reconstitution of a minimal RNA degradosome demonstrates functional coordination between a 3' exonuclease and a DEAD-box RNA helicase. AB - The RNA degradosome is a multiprotein complex required for the degradation of highly structured RNAs. We have developed a method for reconstituting a minimal degradosome from purified proteins. Our results demonstrate that a degradosome like complex containing RNase E, PNPase, and RhlB can form spontaneously in vitro in the absence of all other cellular components. Moreover, ATP-dependent degradation of the malEF REP RNA by the reconstituted, minimal degradosome is indistinguishable from that of degradosomes isolated from whole cells. The Rne protein serves as an essential scaffold in the reconstitution process; however, RNase E activity is not required. Rather, Rne coordinates the activation of RhlB dependent on a 3' single-stranded extension on RNA substrates. A model for degradosome-mediated degradation of structured RNA is presented with its implications for mRNA decay in Escherichia coli. PMID- 10521405 TI - Signal transduction pathways that regulate eukaryotic protein synthesis. PMID- 10521404 TI - Suppression of epithelial apoptosis and delayed mammary gland involution in mice with a conditional knockout of Stat3. AB - Mammary gland involution is characterized by extensive apoptosis of the epithelial cells. At the onset of involution, Stat3 is specifically activated. To address the function of this signaling molecule in mammary epithelial apoptosis, we have generated a conditional knockout of Stat3 using the Cre-lox recombination system. Following weaning, a decrease in apoptosis and a dramatic delay of involution occurred in Stat3 null mammary tissue. Involution is normally associated with a significant increase in IGFBP-5 levels. This was observed in control glands, but not in the absence of Stat3. IGFBP-5 has been suggested to induce apoptosis by sequestering IGF-1 to casein micelles, thereby inhibiting its survival function. Our findings suggest that IGFBP-5 is a direct or indirect target for Stat3 and its upregulation is essential to normal involution. No marked differences were seen in the regulation of Stat5, Bcl-x(L), or Bax in the absence of Stat3. Precocious activation of Stat1 and increases in levels of p53 and p21 occurred and may act as compensatory mechanisms for the eventual initiation of involution observed in Stat3 null mammary glands. This is the first demonstration of the importance of a Stat factor in signaling the initiation of physiological apoptosis in vivo. PMID- 10521406 TI - NF-kappaB/Rel proteins are required for neuronal differentiation of SH-SY5Y neuroblastoma cells. AB - The expression, cellular localization, and activation of the NF-kappaB/Rel transcription factors are altered during neuronal differentiation, but the significance is unclear. Here we investigate the requirement for NF-kappaB/Rel proteins in neuronal differentiation. SH-SY5Y neuroblastoma cells were induced to differentiate with retinoic acid (RA) or 12-O-tetradecanoylphorbol 13-acetate (TPA), and differentiation was demonstrated by morphological criteria and the enhanced expression of Bcl-2. NF-kappaB was transiently activated after the addition of the differentiation inducers before the morphological signs of differentiation and the enhanced Bcl-2 synthesis. The onset of NF-kappaB activation coincided with a significant reduction in the amount of only one of four NF-kappaB-inhibitory proteins examined (I-kappaBbeta). In contrast, NF kappaB activation and the reduction in I-kappaBbeta failed to occur in SH-SY5Y cells transformed with I-kappaBalphaM, a dominant-negative inhibitor of NF kappaB/Rel proteins. These I-kappaBalphaM-expressing cells failed to differentiate into neuronal cell types when treated with RA or TPA, and the increased Bcl-2 synthesis was blocked. Therefore, NF-kappaB/Rel proteins are required for neuronal differentiation of SH-SY5Y neuroblastoma cells. PMID- 10521407 TI - Regulation of the amiloride-sensitive epithelial sodium channel by syntaxin 1A. AB - The first step in transepithelial sodium absorption lies at the apical membrane where the amiloride-sensitive, epithelial sodium channel, ENaC, facilitates sodium entry into the cell. Here we report that the vesicle traffic regulatory (SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor)) protein, syntaxin 1A (S1A), inhibits ENaC mediated sodium entry. This inhibitory effect is selective for S1A and is not reproduced by syntaxin 3. The inhibition does not require the membrane anchoring domain of syntaxin 1A. It was reversed by the S1A-binding protein, Munc-18, but not by a Munc-18 mutant, which lacks syntaxin affinity. Immunostaining of epitope-tagged ENaC subunits showed that syntaxin 1A decreases ENaC current by reducing the number of ENaC channels in the plasma membrane; S1A does not interfere with ENaC protein expression. Immunoprecipitation of syntaxin 1A from the sodium-transporting epithelial cell line, A6, co-precipitates ENaC. These findings indicate that syntaxin 1A and other members of the SNARE machinery are involved in the control of plasma membrane ENaC content, and they suggest that SNARE proteins participate in the regulation of sodium absorption in relation to agonist mediated vesicle insertion retrieval processes. PMID- 10521408 TI - Identification of a cytoplasmic-retention sequence in ERK2. AB - A key step in the signaling mechanism of the mitogen-activated protein kinase/extracellular signal-responsive kinase (ERK) cascade is its translocation into the nucleus where it regulates transcription and other nuclear processes. In an attempt to characterize the subcellular localization of ERK2, we fused it to the 3'-end of the gene expressing green fluorescent protein (GFP), resulting in a GFP-ERK2 protein. The expression of this construct in CHO cells resulted in a nuclear localization of the GFP-ERK2 protein. However, coexpression of the GFP ERK2 with its upstream activator, MEK1, resulted in a cytosolic retention of the GFP-ERK2, which was the result of its association with MEK1, and was reversed upon stimulation. We then examined the role of the C-terminal region of ERK2 in its subcellular localization. Substitution of residues 312-319 of GFP-ERK2 to alanine residues prevented the cytosolic retention of ERK2 as well as its association with MEK1, without affecting its activity. Most important for the cytosolic retention are three acidic amino acids at positions 316, 319, and 320 of ERK2. Substitution of residues 321-327 to alanines impaired the nuclear translocation of ERK2 upon mitogenic stimulation. Thus, we conclude that residues 312-320 of ERK2 are responsible for its cytosolic retention, and residues 321-327 play a role in the mechanism of ERK2 nuclear translocation. PMID- 10521409 TI - IkappaB kinases phosphorylate NF-kappaB p65 subunit on serine 536 in the transactivation domain. AB - Recent investigations have elucidated the cytokine-induced NF-kappaB activation pathway. IkappaB kinase (IKK) phosphorylates inhibitors of NF-kappaB (IkappaBs). The phosphorylation targets them for rapid degradation through a ubiquitin proteasome pathway, allowing the nuclear translocation of NF-kappaB. We have examined the possibility that IKK can phosphorylate the p65 NF-kappaB subunit as well as IkappaB in the cytokine-induced NF-kappaB activation. In the cytoplasm of HeLa cells, the p65 subunit was rapidly phosphorylated in response to TNF-alpha in a time dependent manner similar to IkappaB phosphorylation. In vitro phosphorylation with GST-fused p65 showed that a p65 phosphorylating activity was present in the cytoplasmic fraction and the target residue was Ser-536 in the carboxyl-terminal transactivation domain. The endogenous IKK complex, overexpressed IKKs, and recombinant IKKbeta efficiently phosphorylated the same Ser residue of p65 in vitro. The major phosphorylation site in vivo was also Ser 536. Furthermore, activation of IKKs by NF-kappaB-inducing kinase induced phosphorylation of p65 in vivo. Our finding, together with previous observations, suggests dual roles for IKK complex in the regulation of NF-kappaB.IkappaB complex. PMID- 10521410 TI - UV resonance raman spectra of ligand binding intermediates of sol-gel encapsulated hemoglobin. AB - We report for the first time specific conformational changes for a homogeneous population of ligand-bound adult deoxy human hemoglobin A (HbA) generated by introducing CO into a sample of deoxy-HbA with the effector, inositol hexaphosphate, encapsulated in a porous sol-gel. The preparation of ligand-bound deoxy-HbA results from the speed of ligand diffusion relative to globin conformational dynamics within the sol-gel (1). The ultraviolet resonance Raman (UVRR) difference spectra obtained reveal that E helix motion is initiated upon ligand binding, as signaled by the appearance of an alpha14beta15 Trp W3 band difference at 1559 cm(-1). The subsequent appearance of Tyr (Y8a and Y9a) and W3 (1549 cm(-1)) UVRR difference bands suggest conformational shifts for the penultimate Tyralpha140 on the F helix, the "switch" region Tyralpha42, and the "hinge" region Trpbeta37. The UVRR results expose a sequence of conformational steps leading up to the ligation-induced T to R quaternary structure transition as opposed to a single, concerted switch. More generally, this report demonstrates that sol-gel encapsulation of proteins can be used to study a sequence of specific conformational events triggered by substrate binding because the traditional limitation of substrate diffusion times is overcome. PMID- 10521411 TI - Mildly oxidized low density lipoprotein induces contraction of human endothelial cells through activation of Rho/Rho kinase and inhibition of myosin light chain phosphatase. AB - Mildly oxidized low density lipoprotein (mox-LDL) is critically involved in the early atherogenic responses of the endothelium and increases endothelial permeability through an unknown signal pathway. Here we show that (i) exposure of confluent human endothelial cells (HUVEC) to mox-LDL but not to native LDL induces the formation of actin stress fibers and intercellular gaps within minutes, leading to an increase in endothelial permeability; (ii) mox-LDL induces a transient decrease in myosin light chain (MLC) phosphatase that is paralleled by an increase in MLC phosphorylation; (iii) phosphorylated MLC stimulated by mox LDL is incorporated into stress fibers; (iv) cytoskeletal rearrangements and MLC phosphorylation are inhibited by C3 transferase from Clostridium botulinum, a specific Rho inhibitor, and Y-27632, an inhibitor of Rho kinase; and (v) mox-LDL does not increase intracellular Ca(2+) concentration. Our data indicate that mox LDL induces endothelial cell contraction through activation of Rho and its effector Rho kinase which inhibits MLC phosphatase and phosphorylates MLC. We suggest that inhibition of this novel cell signaling pathway of mox-LDL could be relevant for the prevention of atherosclerosis. PMID- 10521412 TI - Interaction-induced redox switch in the electron transfer complex rusticyanin cytochrome c(4). AB - The blue copper protein rusticyanin isolated from the acidophilic proteobacterium Thiobacillus ferrooxidans displays a pH-dependent redox midpoint potential with a pK value of 7 on the oxidized form of the protein. The nature of the alterations of optical and EPR spectra observed above the pK value indicated that the redox linked deprotonation occurs on the epsilon-nitrogen of the histidine ligands to the copper ion. Complex formation between rusticyanin and its probable electron transfer partner, cytochrome c(4), induced a decrease of rusticyanin's redox midpoint potential by more than 100 mV together with spectral changes similar to those observed above the pK value of the free form. Complex formation thus substantially modifies the pK value of the surface-exposed histidine ligand to the copper ion and thereby tunes the redox midpoint potential of the copper site. Comparisons with reports on other blue copper proteins suggest that the surface exposed histidine ligand is employed as a redox tuning device by many members of this group of soluble electron carriers. PMID- 10521413 TI - Conformational flexibility of the acetylcholinesterase tetramer suggested by x ray crystallography. AB - Acetylcholinesterase, a polymorphic enzyme, appears to form amphiphilic and nonamphiphilic tetramers from a single splice variant; this suggests discrete tetrameric arrangements where the amphipathic carboxyl-terminal sequences can be either buried or exposed. Two distinct, but related crystal structures of the soluble, trypsin-released tetramer of acetylcholinesterase from Electrophorus electricus were solved at 4.5 and 4.2 A resolution by molecular replacement. Resolution at these levels is sufficient to provide substantial information on the relative orientations of the subunits within the tetramer. The two structures, which show canonical homodimers of subunits assembled through four helix bundles, reveal discrete geometries in the assembly of the dimers to form: (a) a loose, pseudo-square planar tetramer with antiparallel alignment of the two four-helix bundles and a large space in the center where the carboxyl-terminal sequences may be buried or (b) a compact, square nonplanar tetramer that may expose all four sequences on a single side. Comparison of these two structures points to significant conformational flexibility of the tetramer about the four helix bundle axis and along the dimer-dimer interface. Hence, in solution, several conformational states of a flexible tetrameric arrangement of acetylcholinesterase catalytic subunits may exist to accommodate discrete carboxyl-terminal sequences of variable dimensions and amphipathicity. PMID- 10521414 TI - Identification of a heparin-binding region of rat thyroglobulin involved in megalin binding. AB - We recently showed that thyroglobulin (Tg) is a heparin-binding protein and that heparin inhibits binding of Tg to its endocytic receptor megalin (gp330). Here we have identified a heparin-binding region in the carboxyl-terminal portion of rat Tg and have studied its involvement in megalin binding. Rat thyroid extracts, obtained by ammonium sulfate precipitation, were separated by column fractionation into four Tg polypeptides, with apparent masses of 660, 330, 210, and 50 kDa. As assessed by enzyme-linked immunoadsorbent assays and ligand blot binding assays, megalin bound to intact Tg (660 and 330 kDa) and, to a even greater extent, to the 210-kDa Tg polypeptide. Furthermore, the 210-kDa Tg polypeptide inhibited megalin binding to intact Tg by approximately 70%. Solid phase assays showed binding of biotin-labeled heparin to intact Tg and to the 210 kDa Tg polypeptide. We characterized the 210-kDa Tg polypeptide by matrix assisted laser desorption/ionization mass spectrometry analysis and found that it corresponds to the carboxyl-terminal portion of rat Tg. We developed a synthetic peptide corresponding to a 15-amino acid sequence in the carboxyl-terminal portion of rat Tg (Arg(689)-Lys(703)), containing a heparin-binding consensus sequence (SRRLKRP) and demonstrated heparin binding to this peptide. A rabbit antibody raised against the peptide recognized intact Tg in its native conformation and under denaturing conditions. This antibody markedly reduced heparin-binding to intact Tg, indicating that the region of native Tg corresponding to the peptide is involved in heparin binding. Furthermore, the anti-Tg peptide antibody almost completely inhibited binding of megalin to Tg, suggesting that the Tg region containing the peptide sequence is required for megalin binding. Physiologically, Tg binding to megalin on thyroid cells may be facilitated by Tg interaction with heparin-like molecules (heparan sulfate proteoglycans) via adjacent binding sites. PMID- 10521415 TI - Stoichiometry of proton release from the catalytic center in photosynthetic water oxidation. Reexamination by a glass electrode study at ph 5.5-7.2. AB - The catalytic center (CC) of water oxidation in photosystem II passes through four stepwise increased oxidized states (S(0)-S(4)) before O(2) evolution takes place from 2H(2)O in the S(4) --> S(0) transition. The pattern of the release of the four protons from the CC cannot be followed directly in the medium, because proton release from unknown amino acid residues also takes place. However, pH independent net charge oscillations of 0:0:1:1 in S(0):S(1):S(2):S(3) have been considered as an intrinsic indicator for the H(+) release from the CC. The net charges have been proposed to be created as the charge difference between electron abstraction and H(+) release from the CC. Then the H(+) release from the CC is 1:0:1:2 for the S(0) --> S(1) --> S(2) --> S(3) --> S(0) transition. Strong support for this conclusion is given in this work with the analysis of the pH dependent pattern of H(+) release in the medium measured directly by a glass electrode between pH 5.5 and 7.2. Improved and crystallizable photosystem II core complexes from the cyanobacterium Synechococcus elongatus were used as material. The pattern can be explained by protons released from the CC with a stoichiometry of 1:0:1:2 and protons from an amino acid group (pK approximately 5.7) that is deprotonated and reprotonated through electrostatic interaction with the oscillating net charges 0:0:1:1 in S(0):S(1):S(2):S(3). Possible water derivatives that circulate through the S states have been named. PMID- 10521416 TI - An RNA binding motif in the Cbp2 protein required for protein-stimulated RNA catalysis. AB - The fifth and terminal intron of yeast cytochrome b pre-mRNA (a group I intron) requires a protein encoded by the nuclear gene CBP2 for splicing. Because catalysis is intrinsic to the RNA, the protein is believed to promote formation of secondary and tertiary structure of the RNA, resulting in a catalytically competent intron. In vitro, this mitochondrial intron can be made to self-splice or undergo protein-facilitated splicing by varying the Mg(2+) and monovalent salt concentrations. This two-component system, therefore, provides a good model for understanding the role of proteins in RNA folding. A UV cross-linking experiment was initiated to identify RNA binding sites on Cbp2 and gain insights into Cbp2 intron interactions. A 12-amino acid region containing a presumptive contact site near the amino terminus was targeted for mutagenesis, and mutant proteins were characterized for RNA binding and stimulation of splicing. Mutations in this region resulted in partial or complete loss of function, demonstrating the importance of this determinant for stimulation of RNA splicing. Several of the mutations that severely reduced splicing did not significantly shift the overall binding isotherm of Cbp2 for the precursor RNA, suggesting that contacts critical for activity are not necessarily reflected in the dissociation constant. This analysis has identified a unique RNA binding motif of alternating basic and aromatic residues that is essential for protein facilitated splicing. PMID- 10521417 TI - Conservation and divergence of the yeast and mammalian unfolded protein response. Activation of specific mammalian endoplasmic reticulum stress element of the grp78/BiP promoter by yeast Hac1. AB - Yeast Hac1 (yHac1), the transcription factor that binds and activates the unfolded protein response element of endoplasmic reticulum (ER)-chaperone gene promoters, only accumulates in stressed cells after an unconventional splicesosome-free mRNA processing step and escape from translation block. In determining whether the novel regulatory mechanisms for yHac1 are conserved in mammalian cells, we discovered a unique unfolded protein response element-like sequence within the endoplasmic reticulum stress element 163, one of the three ER stress elements recently identified in the rat grp78 promoter. The unspliced form of yHac1 is stably expressed in nonstressed mammalian cells and is as active as the spliced form in stimulating the promoter activities of grp genes. Further, the yHac1 mRNA is not processed in response to ER stress in mammalian cells. We identified a CCAGC motif as the yHac1 binding site, which is contained within a YY1 binding site previously shown to be important for mammalian UPR. Dissection of the yHac1 and the YY1 binding sites uncovered specific contact points for an activator protein predicted to be the mammalian homolog of yHac1, the activity of which can be stimulated by YY1. A model of the conserved and unique features of the yeast and mammalian unfolded protein response transcription machinery is proposed. PMID- 10521418 TI - Involvement of NH(2)-terminal sequences in the negative regulation of Vav signaling and transforming activity. AB - Deletion of the NH(2)-terminal 65 amino acids of proto-Vav (to form onco-Vav) activates its transforming activity, suggesting that these sequences serve a negative regulatory role in Vav function. However, the precise role of these NH(2)-terminal sequences and whether additional NH(2)-terminal sequences are also involved in negative regulation have not been determined. Therefore, we generated additional NH(2)-terminal deletion mutants of proto-Vav that lack the NH(2) terminal 127, 168, or 186 amino acids, and assessed their abilities to cause focus formation in NIH 3T3 cells and to activate different signaling pathways. Since Vav mutants lacking 168 or 186 NH(2)-terminal residues showed a several 100 fold greater focus forming activity than that seen with deletion of 65 residues, residues spanning 66 to 187 also contribute significantly to negative regulation of Vav transforming activity. The increase in Vav transforming activity correlated with the activation of the c-Jun, Elk-1, and NF-kappaB transcription factors, as well as increased transcription from the cyclin D1 promoter. Tyrosine 174 is a key site of phosphorylation by Lck in vitro and Lck-mediated phosphorylation has been shown to be essential for proto-Vav GEF function in vitro. However, we found that an NH(2)-terminal Vav deletion mutant lacking this tyrosine residue (DeltaN-186 Vav) retained the ability to be phosphorylated by Lck in vivo and Lck still caused enhancement of DeltaN-186 Vav signaling and transforming activity. Thus, Lck can stimulate Vav via a mechanism that does not involve Tyr(174) or removal of NH(2)-terminal regulatory activity. Finally, we found that NH(2)-terminal deletion enhanced the degree of Vav association with the membrane-containing particulate fraction and that an isolated NH(2)-terminal fragment (residues 1-186) could impair DeltaN-186 Vav signaling. Taken together, these observations suggest that the NH(2) terminus may serve as a negative regulator of Vav by intramolecular interaction with COOH-terminal sequences to modulate efficient membrane association. PMID- 10521419 TI - Suppression of glycogen synthase kinase activity is not sufficient for leukemia enhancer factor-1 activation. AB - Glycogen synthase kinase-3 (GSK) can be regulated by different signaling pathways including those mediated by protein kinase Akt and Wnt proteins. Wnt proteins are believed to activate a transcription factor leukemia enhancer factor-1 (LEF-1) by inhibiting GSK, and Akt was shown to phosphorylate GSK and inhibit its kinase activity. We investigated the effect of an activated Akt on the accumulation of cytosolic beta-catenin and LEF-1-dependent transcription. Although the activated Akt, mAkt, clearly inhibited the kinase activity of GSK, mAkt alone did not induce accumulation of cytosolic beta-catenin or activate LEF-1-dependent transcription. On the contrary, coexpressed Wnt-1 and Frat activated LEF-1 but did not show significant inhibition of GSK-mediated phosphorylation of a peptide substrate. However, mAkt could act synergistically with Wnt-1 or Frat to activate LEF-1. In addition, the interaction of GSK for Axin appeared to decrease in the presence of mAkt, whereas the interaction for Frat remained unchanged. Consistently, a GSK mutant with substitution of a Phe residue for residue Tyr 216, which showed one-fifth of kinase activity of the wild-type GSK, exhibited a reduced association for Axin than the wild-type GSK. These results suggest that inhibition of GSK kinase activity is not sufficient for activation of LEF-1 but may facilitate the activation by reducing the interaction of GSK for Axin. The additional mechanism for LEF-1 activation may require dissociation of GSK from Axin as Frat facilitates the dissociation of GSK from Axin. PMID- 10521420 TI - Post-transcriptional regulation of the arginine transporter Cat-1 by amino acid availability. AB - The regulation of the high affinity cationic amino acid transporter (Cat-1) by amino acid availability has been studied. In C6 glioma and NRK kidney cells, cat 1 mRNA levels increased 3.8-18-fold following 2 h of amino acid starvation. The transcription rate of the cat-1 gene remained unchanged during amino acid starvation, suggesting a post-transcriptional mechanism of regulation. This mechanism was investigated by expressing a cat-1 mRNA from a tetracycline regulated promoter. The cat-1 mRNA contained 1.9 kilobase pairs (kb) of coding sequence, 4.5 kb of 3'-untranslated region, and 80 base pairs of 5'-untranslated region. The full-length (7.9 kb) mRNA increased 5-fold in amino acid-depleted cells. However, a 3.4-kb species that results from the usage of an alternative polyadenylation site was not induced, suggesting that the cat-1 mRNA was stabilized by cis-acting RNA sequences within the 3'-UTR. Transcription and protein synthesis were required for the increase in full-length cat-1 mRNA level. Because omission of amino acids from the cell culture medium leads to a substantial decrease in protein synthesis, the translation of the increased cat-1 mRNA was assessed in amino acid-depleted cells. Western blot analysis demonstrated that cat-1 mRNA and protein levels changed in parallel. The increase in protein level was significantly lower than the increase in mRNA level, supporting the conclusion that cat-1 mRNA is inefficiently translated when the supply of amino acids is limited, relative to amino acid-fed cells. Finally, y(+) mediated transport of arginine in amino acid-fed and -starved cells paralleled Cat-1 protein levels. We conclude that the cat-1 gene is subject to adaptive regulation by amino acid availability. Amino acid depletion initiates molecular events that lead to increased cat-1 mRNA stability. This causes an increase in Cat-1 protein, and y(+) transport once amino acids become available. PMID- 10521421 TI - Cysteine proteinase activity regulation. A possible role of heparin and heparin like glycosaminoglycans. AB - Papain is considered to be the archetype of cysteine proteinases. The interaction of heparin and other glycosaminoglycans with papain may be representative of many mammalian cysteine proteinase-glycosaminoglycan interactions that can regulate the function of this class of proteinases in vivo. The conformational changes in papain structure due to glycosaminoglycan interaction were studied by circular dichroism spectroscopy, and the changes in enzyme behavior were studied by kinetic analysis, monitored with fluorogenic substrate. The presence of heparin significantly increases the alpha-helix content of papain. Heparin binding to papain was demonstrated by affinity chromatography and shown to be mediated by electrostatic interactions. The incubation of papain with heparin promoted a powerful increase in the affinity of the enzyme for the substrate. In order to probe the glycosaminoglycan structure requirements for the papain interaction, the effects of two other glycosaminoglycans were tested. Like heparin, heparan sulfate, to a lesser degree, was able to decrease the papain substrate affinity, and it simultaneously induced alpha-helix structure in papain. On the other hand, dermatan sulfate was not able to decrease the substrate affinity and did not induce alpha-helix structure in papain. Heparin stabilizes the papain structure and thereby its activity at alkaline pH. PMID- 10521423 TI - Characterization of an 8-oxoguanine DNA glycosylase from Methanococcus jannaschii. AB - A thermostable 8-oxoguanine (oxoG) DNA glycosylase from Methanococcus jannaschii has been expressed in Escherichia coli, purified, and characterized. The enzyme, which has been named mjOgg, belongs to the same diverse DNA glycosylase superfamily as the 8-oxoguanine DNA glycosylases from yeast (yOgg1) and human (hOgg1) but is substantially different in sequence. In addition, unlike its eukaryotic counterparts, which have a strong preference for oxoG.C base pairs, mjOgg has little specificity for the base opposite oxoG. mjOgg has both DNA glycosylase and DNA lyase (beta-elimination) activity, and the combined glycosylase/lyase activity occurs at a rate comparable with the glycosylase activity alone. Mutation of Lys-129, analogous to Lys-241 of yOgg1, abolishes glycosylase activity. PMID- 10521422 TI - A novel calcium signaling pathway targets the c-fos intragenic transcriptional pausing site. AB - In many cell types, increased intracellular calcium gives rise to a robust induction of c-fos gene expression. Here we show that in mouse Ltk(-) fibroblasts, calcium ionophore acts in synergy with either cAMP or PMA to strongly induce the endogenous c-fos gene. Run-on analysis shows that this corresponds to a substantial increase in active polymerases on downstream gene sequences, i.e. relief of an elongation block by calcium. Correspondingly a chimeric gene, in which the human metallothionein promoter is fused to the fos gene, is strongly induced by ionophore alone, unlike a c-fos promoter/beta-globin coding unit chimeric construct. Internal deletions in the hMT-fos reporter localize the intragenic calcium regulatory element to the 5' portion of intron 1, thereby confirming and extending previous in vitro mapping data. Ionophore induced cAMP response element-binding protein phosphorylation on Ser(133) without affecting the extracellular signal-regulated kinase cascade. Surprisingly, induction involved neither CaM-Ks nor calcineurin, while the calmodulin antagonist W7 activated c-fos transcription on its own. These data suggest that a novel calcium signaling pathway mediates intragenic regulation of c-fos expression via suppression of a transcriptional pause site. PMID- 10521424 TI - Cloning and characterization of AOEB166, a novel mammalian antioxidant enzyme of the peroxiredoxin family. AB - Using two-dimensional electrophoresis, we have recently identified in human bronchoalveolar lavage fluid a novel protein, termed B166, with a molecular mass of 17 kDa. Here, we report the cloning of human and rat cDNAs encoding B166, which has been renamed AOEB166 for antioxidant enzyme B166. Indeed, the deduced amino acid sequence reveals that AOEB166 represents a new mammalian subfamily of AhpC/TSA peroxiredoxin antioxidant enzymes. Human AOEB166 shares 63% similarity with Escherichia coli AhpC22 alkyl hydroperoxide reductase and 66% similarity with a recently identified Saccharomyces cerevisiae alkyl hydroperoxide reductase/thioredoxin peroxidase. Moreover, recombinant AOEB166 expressed in E. coli exhibits a peroxidase activity, and an antioxidant activity comparable with that of catalase was demonstrated with the glutamine synthetase protection assay against dithiothreitol/Fe3+/O(2) oxidation. The analysis of AOEB166 mRNA distribution in 30 different human tissues and in 10 cell lines shows that the gene is widely expressed in the body. Of interest, the analysis of N- and C terminal domains of both human and rat AOEB166 reveals amino acid sequences presenting features of mitochondrial and peroxisomal targeting sequences. Furthermore, human AOEB166 expressed as a fusion protein with GFP in HepG2 cell line is sorted to these organelles. Finally, acute inflammation induced in rat lung by lipopolysaccharide is associated with an increase of AOEB166 mRNA levels in lung, suggesting a protective role for AOEB166 in oxidative and inflammatory processes. PMID- 10521425 TI - Identification of glucagon-like peptide-2 (GLP-2)-activated signaling pathways in baby hamster kidney fibroblasts expressing the rat GLP-2 receptor. AB - Glucagon-like peptide-2 (GLP-2) promotes the expansion of the intestinal epithelium through stimulation of the GLP-2 receptor, a recently identified member of the glucagon-secretin G protein-coupled receptor superfamily. Although activation of G protein-coupled receptors may lead to stimulation of cell growth, the mechanisms transducing the GLP-2 signal to mitogenic proliferation remain unknown. We now report studies of GLP-2R signaling in baby hamster kidney (BHK) cells expressing a transfected rat GLP-2 receptor (BHK-GLP-2R cells). GLP-2, but not glucagon or GLP-1, increased the levels of cAMP and activated both cAMP response element- and AP-1-dependent transcriptional activity in a dose-dependent manner. The activation of AP-1-luciferase activity was protein kinase A (PKA) dependent and markedly diminished in the presence of a dominant negative inhibitor of PKA. Although GLP-2 stimulated the expression of c-fos, c-jun, junB, and zif268, and transiently increased p70 S6 kinase in quiescent BHK-GLP-2R cells, GLP-2 also inhibited extracellular signal-regulated kinase 1/2 and reduced serum-stimulated Elk-1 activity. Furthermore, no rise in intracellular calcium was observed following GLP-2 exposure in BHK-GLP-2R cells. Although GLP-2 stimulated both cAMP accumulation and cell proliferation, 8-bromo-cyclic AMP alone did not promote cell proliferation. These findings suggest that the GLP-2R may be coupled to activation of mitogenic signaling in heterologous cell types independent of PKA via as yet unidentified downstream mediators of GLP-2 action in vivo. PMID- 10521426 TI - The human cytotoxic T cell granule serine protease granzyme H has chymotrypsin like (chymase) activity and is taken up into cytoplasmic vesicles reminiscent of granzyme B-containing endosomes. AB - Serine proteases (granzymes) contained within the cytoplasmic granules of cytotoxic T cells and natural killer cells play a variety of roles including the induction of target cell apoptosis, breakdown of extracellular matrix proteins and induction of cytokine secretion by bystander leukocytes. Different granzymes display proteolytic specificities that mimic the activities of trypsin or chymotrypsin, or may cleave substrates at acidic ("Asp-ase") or at long unbranched amino acids such as Met ("Met-ase"). Here, we report that recombinant granzyme H has chymotrypsin-like (chymase) activity, the first report of a human granzyme with this proteolytic specificity. Recombinant 32-kDa granzyme H expressed in the baculovirus vector pBacPAK8 was secreted from Sf21 cells and recovered by Ni-affinity chromatography, using a poly-His tag encoded at the predicted carboxyl terminus of full-length granzyme H cDNA. The granzyme H efficiently cleaved Suc-Phe-Leu-Phe-SBzl (v = 185 nM/s at [S] = 0.217 mM) and also hydrolyzed Boc-Ala-Ala-X-SBzl (X = Phe, Tyr, Met, Nle, or Nva) with slower rates but had little tryptase or Asp-ase activity. Enzymatic activity was inhibited completely by 0.1 mM 3,4-dichloroisocoumarin and 84% by 1.0 mM phenylmethylsulfonyl fluoride. Fluoresceinated granzyme H was internalized in a temperature-dependent manner by Jurkat cells into endosome-like vesicles, suggesting that it can bind to cell surface receptors similar to those that bind granzyme B. This suggests a hitherto unsuspected intracellular function for granzyme H. PMID- 10521427 TI - 1.68-A crystal structure of endopolygalacturonase II from Aspergillus niger and identification of active site residues by site-directed mutagenesis. AB - Polygalacturonases specifically hydrolyze polygalacturonate, a major constituent of plant cell wall pectin. To understand the catalytic mechanism and substrate and product specificity of these enzymes, we have solved the x-ray structure of endopolygalacturonase II of Aspergillus niger and we have carried out site directed mutagenesis studies. The enzyme folds into a right-handed parallel beta helix with 10 complete turns. The beta-helix is composed of four parallel beta sheets, and has one very small alpha-helix near the N terminus, which shields the enzyme's hydrophobic core. Loop regions form a cleft on the exterior of the beta helix. Site-directed mutagenesis of Asp(180), Asp(201), Asp(202), His(223), Arg(256), and Lys(258), which are located in this cleft, results in a severe reduction of activity, demonstrating that these residues are important for substrate binding and/or catalysis. The juxtaposition of the catalytic residues differs from that normally encountered in inverting glycosyl hydrolases. A comparison of the endopolygalacturonase II active site with that of the P22 tailspike rhamnosidase suggests that Asp(180) and Asp(202) activate the attacking nucleophilic water molecule, while Asp(201) protonates the glycosidic oxygen of the scissile bond. PMID- 10521428 TI - Metal ligation by Walker homology B aspartate betaD262 at site 3 of the latent but not activated form of the chloroplast F(1)-ATPase from Chlamydomonas reinhardtii. AB - Site-directed mutations D262C, D262H, D262N, and D262T were made to the beta subunit Walker Homology B aspartate of chloroplast F(1)-ATPase in Chlamydomonas. Photoautotrophic growth and photophosphorylation rates were 3-14% of wild type as were ATPase activities of purified chloroplast F(1) indicating that betaD262 is an essential residue for catalysis. The EPR spectrum of vanadyl bound to Site 3 of chloroplast F(1) as VO(2+)-ATP gave rise to two EPR species designated B and C in wild type and mutants. (51)V-hyperfine parameters of species C, present exclusively in the activated enzyme state, did not change significantly by the mutations examined indicating that it is not an equatorial ligand to VO(2+), nor is it hydrogen-bonded to a coordinated water at an equatorial position. Every mutation changed the ratio of EPR species C/B and/or the (51)V-hyperfine parameters of species B, the predominant conformation of VO(2+)-nucleotide bound to Site 3 in the latent (down-regulated) state. The results indicate that the Walker Homology B aspartate coordinates the metal of the predominant metal nucleotide conformation at Site 3 in the latent state but not in the conformation present exclusively upon activation and elucidates one of the specific changes in metal ligation involved with activation. PMID- 10521429 TI - Cloning the promoter for transforming growth factor-beta type III receptor. Basal and conditional expression in fetal rat osteoblasts. AB - Transforming growth factor-beta binds to three high affinity cell surface molecules that directly or indirectly regulate its biological effects. The type III receptor (TRIII) is a proteoglycan that lacks significant intracellular signaling or enzymatic motifs but may facilitate transforming growth factor-beta binding to other receptors, stabilize multimeric receptor complexes, or segregate growth factor from activating receptors. Because various agents or events that regulate osteoblast function rapidly modulate TRIII expression, we cloned the 5' region of the rat TRIII gene to assess possible control elements. DNA fragments from this region directed high reporter gene expression in osteoblasts. Sequencing showed no consensus TATA or CCAAT boxes, whereas several nuclear factors binding sequences within the 3' region of the promoter co-mapped with multiple transcription initiation sites, DNase I footprints, gel mobility shift analysis, or loss of activity by deletion or mutation. An upstream enhancer was evident 5' proximal to nucleotide -979, and a silencer region occurred between nucleotides -2014 and -2194. Glucocorticoid sensitivity mapped between nucleotides -687 and -253, whereas bone morphogenetic protein 2 sensitivity co mapped within the silencer region. Thus, the TRIII promoter contains cooperative basal elements and dispersed growth factor- and hormone-sensitive regulatory regions that can control TRIII expression by osteoblasts. PMID- 10521431 TI - Biochemical analysis of SopE from Salmonella typhimurium, a highly efficient guanosine nucleotide exchange factor for RhoGTPases. AB - RhoGTPases are key regulators of eukaryotic cell physiology. The bacterial enteropathogen Salmonella typhimurium modulates host cell physiology by translocating specific toxins into the cytoplasm of host cells that induce responses such as apoptotic cell death in macrophages, the production of proinflammatory cytokines, the rearrangement of the host cell actin cytoskeleton (membrane ruffling), and bacterial entry into host cells. One of the translocated toxins is SopE, which has been shown to bind to RhoGTPases of the host cell and to activate RhoGTPase signaling. SopE is sufficient to induce profuse membrane ruffling in Cos cells and to facilitate efficient bacterial internalization. We show here that SopE belongs to a novel class of bacterial toxins that modulate RhoGTPase function by transient interaction. Surface plasmon resonance measurements revealed that the kinetics of formation and dissociation of the SopE.CDC42 complex are in the same order of magnitude as those described for complex formation of GTPases of the Ras superfamily with their cognate guanine nucleotide exchange factors (GEFs). In the presence of excess GDP, dissociation of the SopE.CDC42 complex was accelerated more than 1000-fold. SopE-mediated guanine nucleotide exchange was very efficient (e.g. exchange rates almost 10(5) fold above the level of the uncatalyzed reaction; substrate affinity), and the kinetic constants were similar to those described for guanine nucleotide exchange mediated by CDC25 or RCC1. Far-UV CD spectroscopy revealed that SopE has a high content of alpha-helical structure, a feature also found in Dbl homology domains, Sec7-like domains, and the Ras-GEF domain of Sos. Despite the lack of any obvious sequence similarity, our data suggest that SopE may closely mimic eukaryotic GEFs. PMID- 10521430 TI - Protein kinase C-zeta and phosphoinositide-dependent protein kinase-1 are required for insulin-induced activation of ERK in rat adipocytes. AB - The mechanisms used by insulin to activate the multifunctional intracellular effectors, extracellular signal-regulated kinases 1 and 2 (ERK1/2), are only partly understood and appear to vary in different cell types. Presently, in rat adipocytes, we found that insulin-induced activation of ERK was blocked (a) by chemical inhibitors of both phosphatidylinositol 3-kinase (PI3K) and protein kinase C (PKC)-zeta, and, moreover, (b) by transient expression of both dominant negative Deltap85 PI3K subunit and kinase-inactive PKC-zeta. Further, insulin effects on ERK were inhibited by kinase-inactive 3-phosphoinositide-dependent protein kinase-1 (PDK-1), and by mutation of Thr-410 in the activation loop of PKC-zeta, which is the target of PDK-1 and is essential for PI3K/PDK-1-dependent activation of PKC-zeta. In addition to requirements for PI3K, PDK-1, and PKC zeta, we found that a tyrosine kinase (presumably the insulin receptor), the SH2 domain of GRB2, SOS, RAS, RAF, and MEK1 were required for insulin effects on ERK in the rat adipocyte. Our findings therefore suggested that PDK-1 and PKC-zeta serve as a downstream effectors of PI3K, and act in conjunction with GRB2, SOS, RAS, and RAF, to activate MEK and ERK during insulin action in rat adipocytes. PMID- 10521432 TI - Nuclear DEAF-1-related (NUDR) protein contains a novel DNA binding domain and represses transcription of the heterogeneous nuclear ribonucleoprotein A2/B1 promoter. AB - Nuclear DEAF-1-related (NUDR) protein is a novel transcriptional regulator with sequence similarity to developmental and oncogenic proteins. NUDR protein deletions were used to localize the DNA binding domain between amino acids 167 and 368, and site-specific DNA photocross-linking indicated at least two sites of protein-DNA contact within this domain. The DNA binding domain contains a proline rich region and a region with similarity to a Myc-type helix-loop-helix domain but does not include the zinc finger motif at the C terminus. Deoxyribonuclease I protection assays confirmed the presence of multiple NUDR binding motifs (TTC(C/G)G) in the heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1) promoter and also in the 5'-untranslated region (UTR) of hNUDR cDNA. NUDR produced a 65-70% repression of the hnRNP A2/B1 promoter activity, and NUDR binding motifs in the 5'-UTR were found to mediate this repression. NUDR dependent repression was also observed when the 5'-UTR of NUDR was placed onto a heterologous thymidine kinase promoter in an analogous 5'-UTR position but not when placed upstream of transcription initiation. These results suggest that NUDR may regulate the in vivo expression of hnRNP A2/B1 and NUDR genes and imply that inactivation of NUDR could contribute to the overexpression of hnRNP A2/B1 observed in some human cancers. PMID- 10521433 TI - The Candida albicans phospholipomannan is a family of glycolipids presenting phosphoinositolmannosides with long linear chains of beta-1,2-linked mannose residues. AB - In a series of studies, we have shown that Candida albicans synthesizes a glycolipid, phospholipomannan (PLM), which reacted with antibodies specific for beta-1,2-oligomannosides and was biosynthetically labeled by [(3)H]mannose, [(3)H]palmitic acid, and [(32)P]phosphorus. PLM has also been shown to be released from the C. albicans cell wall and to bind to and stimulate macrophage cells. In this study, we show by thin layer chromatography scanning of metabolically radiolabeled extracts that the C. albicans PLM corresponds to a family of mannose and inositol co-labeled glycolipids. We describe the purification process of the molecule and the release of its glycan fraction through alkaline hydrolysis. Analysis of this glycan fraction by radiolabeling and methylation-methanolysis confirmed the presence of inositol and of 1, 2 linked mannose units. NMR studies evidenced linear chains of beta-1,2 oligomannose as the major PLM components. Mass spectrometry analysis revealed that these chains were present in phosphoinositolmannosides with degrees of polymerization varying from 8 to 18 sugar residues. The PLM appears as a new type of eukaryotic inositol-tagged glycolipid in relationship to both the absence of glucosamine and the organization of its glycan chains. This first structural evidence for the presence of beta-1, 2-oligomannosides in a glycoconjugate other than the C. albicans phosphopeptidomannan may have some pathophysiological relevance to the adhesive, protective epitope, and signaling properties thus far established for these residues. PMID- 10521434 TI - The human PICD gene encodes a cytoplasmic and peroxisomal NADP(+)-dependent isocitrate dehydrogenase. AB - Human PICD was identified by homology probing the data base of expressed sequence tags with the protein sequence of Saccharomyces cerevisiae Idp3p, a peroxisomal NADP(+)-dependent isocitrate dehydrogenase. The human PICD cDNA contains a 1242 base pair open reading frame, and its deduced protein sequence is 59% identical to yeast Idp3p. Expression of PICD partially rescued the fatty acid growth defect of the yeast idp3 deletion mutant suggesting that PICD is functionally homologous to Idp3p. Kinetic studies on bacterially expressed PICD demonstrated that this enzyme catalyzed the oxidative decarboxylation of isocitrate to 2-oxoglutarate with a specific activity of 22.5 units/mg and that PICD displayed K(M) values of 76 microM for isocitrate and 112 microM for NADP(+). In subcellular fractionation experiments, we found PICD in both peroxisomes and cytoplasm of human and rat liver cells, with approximately 27% of total PICD protein associated with peroxisomes. The presence of PICD in mammalian peroxisomes suggests roles in the regeneration of NADPH for intraperoxisomal reductions, such as the conversion of 2, 4-dienoyl-CoAs to 3-enoyl-CoAs, as well as in peroxisomal reactions that consume 2-oxoglutarate, namely the alpha-hydroxylation of phytanic acid. As for cytoplasmic PICD, the phenotypes of patients with glucose-6-phosphate dehydrogenase deficiency (Luzzatto, L., and Mehta, A. (1995) in The Metabolic and Molecular Bases of Inherited Disease (Scriver, C. R., Beaudet, A. L., Sly, W. S., and Valle, D., eds) Vol. 3, 7th Ed., pp. 3367-3398, McGraw-Hill Inc., New York) suggest that PICD serves a significant role in cytoplasmic NADPH production, particularly under conditions that do not favor the use of the hexose monophosphate shunt (Luzzatto et al.). PMID- 10521435 TI - Structural features required for the interaction of the Hsp70 molecular chaperone DnaK with its cochaperone DnaJ. AB - Hsp70 family members together with their Hsp40 cochaperones function as molecular chaperones, using an ATP-controlled cycle of polypeptide binding and release to mediate protein folding. Hsp40 plays a key role in the chaperone reaction by stimulating the ATPase activity and activating the substrate binding of Hsp70. We have explored the interaction between the Escherichia coli Hsp70 family member, DnaK, and its cochaperone partner DnaJ. Our data show that the binding of ATP, subsequent conformational changes in DnaK, and DnaJ-stimulated ATP hydrolysis are all required for the formation of a DnaK-DnaJ complex as monitored by Biacore analysis. In addition, our data imply that the interaction of the J-domain with DnaK depends on the substrate binding state of DnaK. PMID- 10521436 TI - Inactivation of the open reading frame slr0399 in Synechocystis sp. PCC 6803 functionally complements mutations near the Q(A) niche of photosystem II. A possible role of Slr0399 as a chaperone for quinone binding. AB - The Synechocystis sp. PCC 6803 triple mutant D2R8 with V247M/A249T/M329I mutations in the D2 subunit of the photosystem II is impaired in Q(A) function, has an apparently mobile Q(A), and is unable to grow photoautotrophically. Several photoautotrophic pseudorevertants of this mutant have been isolated, each of which retained the original psbDI mutations of D2R8. Using a newly developed mapping technique, the site of the secondary mutations has been located in the open reading frame slr0399. Two different nucleotide substitutions and a deletion of about 60% of slr0399 were each shown to restore photoautotrophy in different pseudorevertants of the mutant D2R8, suggesting that inactivation of Slr0399 leads to photoautotrophic growth in D2R8. Indeed, a targeted deletion of slr0399 restores photoautotrophy in D2R8 and in other psbDI mutants impaired in Q(A) function. Slr0399 is similar to the hypothetical protein Ycf39, which is encoded in the cyanelle genome of Cyanophora paradoxa; in the chloroplast genomes of diatoms, dinoflagellates, and red algae; and in the nuclear genome of Arabidopsis thaliana. Slr0399 and Ycf39 have a NAD(P)H binding motif near their N terminus and have some similarity to isoflavone reductase-like proteins and to a subunit of the eukaryotic NADH dehydrogenase complex I. Deletion of slr0399 in wild type Synechocystis sp. PCC 6803 has no significant phenotypic effects other than a decrease in thermotolerance under both photoautotrophic and photomixotrophic conditions. We suggest that Slr0399 is a chaperone-like protein that aids in, but is not essential for, quinone insertion and protein folding around Q(A) in photosystem II. Moreover, as the effects of Slr0399 are not limited to photosystem II, this protein may also be involved in assembly of quinones in other photosynthetic and respiratory complexes. PMID- 10521437 TI - Casein kinase II activity is required for transferrin receptor endocytosis. AB - The effect of protein kinase inhibitors on transferrin receptor (TR) internalization was examined in HeLa, A431, 3T3-L1 cells, and primary chicken embryo fibroblasts. We show that TR endocytosis is not affected by tyrosine kinase or protein kinase C inhibitors, but is inhibited by one serine/threonine kinase inhibitor, H-89. Inhibition occurred within 15 min, was completely reversible after H-89 withdrawal, and was specific for endocytosis rather than pinocytosis since a TR mutant lacking an internalization signal was not affected. Interestingly, H-89 also inhibited the internalization of a TR chimera containing the major histocompatibility complex class II invariant chain cytoplasmic tail, indicating that the effect was not specific for the TR. Since H-89 inhibits a number of kinases, we employed a permeabilized cell endocytosis assay to further characterize the kinase. In permeabilized 3T3-L1 cells, addition of pseudosubstrate inhibitor peptides of casein kinase II (CKII) blocked TR internalization by more than 50%, whereas pseudosubstrates of cyclic AMP dependent kinase A, protein kinase C, and casein kinase I had no effect. Furthermore, addition of purified CKII to the cell-free reactions containing CKII pseudosubstrates reversed the endocytosis block, suggesting that CKII or a CKII like activity is required for constitutive endocytosis. PMID- 10521438 TI - Molecular cloning of brain-specific GD1alpha synthase (ST6GalNAc V) containing CAG/Glutamine repeats. AB - A novel member of the mouse CMP-NeuAc: beta-N-acetylgalactosaminide alpha2,6 sialyltransferase (ST6GalNAc) subfamily, designated ST6GalNAc V, was identified by BLAST analysis of expressed sequence tags. The sequence of the longest cDNA clone of ST6GalNAc V encoded a type II membrane protein with 8 amino acids comprising the cytoplasmic domain, 21 amino acids comprising the transmembrane region, and 306 amino acids comprising the catalytic domain. The predicted amino acid sequence showed homology to the previously cloned ST6GalNAc III and IV, with common amino acid sequences in sialyl motifs L and S among these three enzymes. Eleven CAG repeats were found in the stem region. A fusion protein with protein A and extracts from L cells transfected with ST6GalNAc V in a expression vector showed enzyme activity of alpha2,6-sialyltransferase almost exclusively for GM1b, but not toward glycoproteins. Sialidase treatment and thin layer chromatography immunostaining revealed that the product was GD1alpha. Northern blotting revealed that three transcripts of the gene were expressed specifically in brain tissues. It is concluded that this enzyme is involved in the synthesis of GD1alpha in the nervous tissues, and the CAG repeats may have implications in neurodegenerative diseases. PMID- 10521439 TI - Specific sequence elements are required for the expression of functional tumor necrosis factor-alpha-converting enzyme (TACE). AB - The tumor necrosis factor-alpha-converting enzyme (TACE) is a membrane-anchored zinc metalloprotease involved in precursor tumor necrosis factor-alpha secretion. We designed a series of constructs containing full-length human TACE and several truncate forms for overexpression in insect cells. Here, we demonstrate that full length TACE is expressed in insect cells inefficiently: only minor amounts of this enzyme are converted from an inactive precursor to the mature, functional form. Removal of the cytoplasmic and transmembrane domains resulted in the efficient secretion of mature, active TACE. Further removal of the cysteine-rich domain located between the catalytic and transmembrane domains resulted in the secretion of mature catalytic domain in association with the precursor (pro) domain. This complex was inactive and function was only restored after dissociation of the complex by dilution or treatment with 4-aminophenylmercuric acetate. Therefore, the pro domain of TACE is an inhibitor of the catalytic domain, and the cysteine-rich domain appears to play a role in the release of the pro domain. Insect cells failed to secrete a deletion mutant encoding the catalytic domain but lacking the inhibitory pro domain. This truncate was inactive and extensively degraded intracellularly, suggesting that the pro domain is required for the secretion of functional TACE. PMID- 10521440 TI - Kinetics of ternary complex formation with fusion proteins composed of the A(1) adenosine receptor and G protein alpha-subunits. AB - High affinity agonist binding to G protein-coupled receptors depends on the formation of a ternary complex between agonist, receptor, and G protein. This process is too slow to be accounted for by a simple diffusion-controlled mechanism. We have tested if the interaction between activated receptor and G protein is rate-limiting by fusing the coding sequence of the human A(1) adenosine receptor to that of Galpha(i-1) (A(1)/Galpha(i-1)) and of Galpha(o) (A(1)/Galpha(o)). Fusion proteins of the expected molecular mass were detected following transfection of HEK293 cells. Ternary complex formation was monitored by determining the kinetics for binding of the high affinity agonist (-)-N(6) 3[(125)I](iodo-4-hydroxyphenylisopropyl)adenosine; these were similar in the wild type receptor and the fusion proteins over the temperature range of 10 to 30 degrees C. Agonist dissociation may be limited by the stability of the ternary complex. This assumption was tested by creating fusion proteins in which the Cys(351) of Galpha(i-1) was replaced with glycine (A(1)/Galpha(i-1)C351G) or isoleucine (A(1)/Galpha(i-1)C351I) to lower the affinity of the receptor for the G protein. In these mutated fusion proteins, the dissociation rate of the ternary complex was accelerated; in contrast, the rate of the forward reaction was not affected. We therefore conclude that (i) receptor activation per se rather than its interaction with the G protein is rate-limiting in ternary complex formation; (ii) the stability of the ternary complex is determined by the dissociation rate of the G protein. These features provide for a kinetic proofreading mechanism that sustains the fidelity of receptor-G protein coupling. PMID- 10521441 TI - Mass spectrometric identification of increased C16 ceramide levels during apoptosis. AB - A variety of molecular changes occur during the process of apoptosis. Much of the recent work has focused on changes in critical cellular proteins, proteins necessary for the initiation and continuation of the apoptotic process. Given the fact that numerous membrane changes occur throughout the apoptotic process, we initiated an investigation aimed at determining the major lipid changes that occurred during programmed cell death. When ionizing radiation was used to initiate the apoptotic process in Jurkat cells, one of the major changes that occurred within 24 h was an increase in a species with a m/z of 572 as determined by negative ion electrospray mass spectrometry. This particular mass ion displayed high performance liquid chromatography characteristics of a neutral lipid species. Further analysis by collision-induced-dissociation tandem mass spectrometry indicated only one daughter species indicative of a Cl adduct and therefore a parental mass of 537. Comparison to a commercial C16 ceramide yielded identical spectra by mass spectrometry (MS) and MS/MS analysis in the negative ion mode. Increases in C16 ceramide levels occurred 2 h after initiation of apoptosis by ionizing radiation, and its accumulation paralleled apoptosis as determined by cellular morphology. Interestingly, radiation-sensitive Jurkat cells displayed increased levels of long term C16 ceramide accumulation, whereas radiation-resistant K562 cells did not. These findings were supported by increases in caspase-3 activity in Jurkat cells, whereas caspase-3 activity in K562 cells remained unchanged. C16 ceramide accumulation and sensitivity to ionizing radiation was investigated further in a melanoma cell line. Only those cells that were radiation sensitive (approximately 70-75%) displayed increases in long term ceramide accumulation. Taken together, these results indicated a correlation between increases in C16 ceramide accumulation and radiation sensitivity. Increases in long term C16 ceramide accumulation were also seen in Fas-induced apoptosis, which occurred at time points greater than 2 h. Analysis of mitochondrial modifications using the mitochondrial probe nonyl acridine orange (NAO) indicated that initial increases in C16 ceramide levels closely paralleled the decrease in mitochondrial mass during Fas or radiation-induced apoptosis. Taken together, these results support a role for C16 ceramide in the effector (mitochondrial) phase of apoptosis. PMID- 10521442 TI - The 42-amino acid insert in the FMN domain of neuronal nitric-oxide synthase exerts control over Ca(2+)/calmodulin-dependent electron transfer. AB - The neuronal and endothelial nitric-oxide synthases (nNOS and eNOS) differ from inducible NOS in their dependence on the intracellular Ca(2+) concentration. Both nNOS and eNOS are activated by the reversible binding of calmodulin (CaM) in the presence of Ca(2+), whereas inducible NOS binds CaM irreversibly. One major divergence in the close sequence similarity between the NOS isoforms is a 40-50 amino acid insert in the middle of the FMN-binding domains of nNOS and eNOS. It has previously been proposed that this insert forms an autoinhibitory domain designed to destabilize CaM binding and increase its Ca(2+) dependence. To examine the importance of the insert we constructed two deletion mutants designed to remove the bulk of it from nNOS. Both mutants (Delta40 and Delta42) retained maximal NO synthesis activity at lower concentrations of free Ca(2+) than the wild type enzyme. They were also found to retain 30% of their activity in the absence of Ca(2+)/CaM, indicating that the insert plays an important role in disabling the enzyme when the physiological Ca(2+) concentration is low. Reduction of nNOS heme by NADPH under rigorous anaerobic conditions was found to occur in the wild type enzyme only in the presence of Ca(2+)/CaM. However, reduction of heme in the Delta40 mutant occurred spontaneously on addition of NADPH in the absence of Ca(2+)/CaM. This suggests that the insert regulates activity by inhibiting electron transfer from FMN to heme in the absence of Ca(2+)/CaM and by destabilizing CaM binding at low Ca(2+) concentrations, consistent with its role as an autoinhibitory domain. PMID- 10521443 TI - Insulin antiapoptotic signaling involves insulin activation of the nuclear factor kappaB-dependent survival genes encoding tumor necrosis factor receptor associated factor 2 and manganese-superoxide dismutase. AB - We recently showed that the antiapoptotic function of insulin requires nuclear factor kappaB (NF-kappaB) activation (Bertrand, F., Atfi, A., Cadoret, A., L'Allemain, G., Robin, H., Lascols, O., Capeau, J., and Cherqui, G. (1998) J. Biol. Chem. 273, 2931-2938). Here we sought to identify the NF-kappaB-dependent survival genes that are activated by insulin to mediate this function. Insulin increased the expression of tumor necrosis factor receptor-associated factor 2 (TRAF2) mRNA and protein in Chinese hamster ovary cells overexpressing insulin receptors (IRs). This effect required (i) IR activation since it was abrogated by IR mutation at tyrosines 1162 and 1163 and (ii) NF-kappaB activation since it was abolished by overexpression of dominant-negative IkappaB-alpha(A32/36) and mimicked by overexpression of the NF-kappaB c-Rel subunit. TRAF2 contributed to insulin protection against serum withdrawal-induced apoptosis since TRAF2 overexpression mimicked insulin protection, whereas overexpression of dominant negative TRAF2-(87-501) reduced this process. Along with its protective effect, overexpressed TRAF2 increased basal and insulin-stimulated NF-kappaB activities. All effects were inhibited by IkappaB-alpha(A32/36), suggesting that an amplification loop involving TRAF2 activation of NF-kappaB is implicated in insulin antiapoptotic signaling. We also show that insulin increased manganese superoxide dismutase (Mn-SOD) mRNA expression through NF-kappaB activation and that Mn-SOD contributed to insulin antiapoptotic signaling since expression of antisense Mn-SOD RNA decreased this process. This study provides the first evidence that insulin activates the NF-kappaB-dependent survival genes encoding TRAF2 and Mn-SOD and thereby clarifies the role of NF-kappaB in the antiapoptotic function of insulin. PMID- 10521444 TI - Tumor necrosis factor-related apoptosis-inducing ligand receptors signal NF kappaB and JNK activation and apoptosis through distinct pathways. AB - Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a member of the TNF family that interacts with several receptors, including TRAIL-R1, TRAIL-R2, and TRAIL-R4. TRAIL-R1 and TRAIL-R2 can induce apoptosis of cancer cells and activate the transcription factor NF-kappaB. TRAIL-R4 can activate NF kappaB and protect cells from TRAIL-induced apoptosis. Here we show that TRAIL-R1 , TRAIL-R2-, and TRAIL-R4-induced NF-kappaB activation are mediated by a TRAF2 NIK-IkappaB kinase alpha/beta signaling cascade but is MEKK1 independent. TRAIL receptors also activate the protein kinase JNK. JNK activation by TRAIL-R1 is mediated by a TRAF2-MEKK1-MKK4 but not the TRAF2-NIK/IkappaB kinase alpha/beta signaling pathway. We also show that activation of NF-kappaB or overexpression of TRAIL-R4 does not protect TRAIL-R1-induced apoptosis. Moreover, inhibition of NF kappaB by IkappaBalpha sensitizes cells to tumor necrosis factor- but not TRAIL induced apoptosis. These findings suggest that TRAIL receptors induce apoptosis, NF-kappaB and JNK activation through distinct signaling pathways, and activation of NF-kappaB is not sufficient for protecting cells from TRAIL-induced apoptosis. PMID- 10521445 TI - Freshly fractured crystalline silica induces activator protein-1 activation through ERKs and p38 MAPK. AB - The transcription factor activator protein-1 (AP-1) reportedly plays an important role in the induction of neoplastic transformation and multiple genes involved in cell proliferation, differentiation, and inflammation. To investigate the mechanisms of silica-induced carcinogenesis, AP-1-luciferase reporter transgenic mice were used as an in vivo model, whereas the JB6 mouse epidermal cell line and a rat lung epithelial cell line were employed as in vitro models to study the effects of silica at the molecular level. Freshly fractured silica caused an 8 fold increase in AP-1 activity in JB6 cells and a 2.5-fold increase in rat lung epithelial cells. The induction of AP-1 activity in cultured cell lines was time- and dose-dependent. Intratracheal administration of silica was also able to induce AP-1 transactivation in transgenic mice. AP-1 activation was first observed at 2 days after silica administration and reached its maximum at 3 days post-exposure of the mice to silica. The signal transduction pathways for AP-1 activation were also investigated using these cell lines. The results demonstrate that freshly fractured silica stimulates mitogen-activated protein kinase (MAPK) family members, as determined by the phosphorylation of p38 MAPK and extracellular signal-regulated protein kinases (ERKs). Inhibition of ERKs with PD98059 or of p38 with SB203580 significantly inhibited silica-induced AP-1 activation. These findings demonstrate for the first time that freshly fractured silica induces AP-1 activation, which may be mediated through p38 MAPK and ERK pathways. Unraveling the complex mechanisms associated with these events may provide insights into the initiation and progression of silica-induced carcinogenesis. PMID- 10521446 TI - Platelet-derived growth factor receptor-induced feed-forward inhibition of excitatory transmission between hippocampal pyramidal neurons. AB - Growth factor receptors provide a major mechanism for the activation of the nonreceptor tyrosine kinase c-Src, and this kinase in turn up-regulates the activity of N-methyl-D-aspartate (NMDA) receptors in CA1 hippocampal neurons (1). Unexpectedly, applications of platelet-derived growth factor (PDGF)-BB to cultured and isolated CA1 hippocampal neurons depressed NMDA-evoked currents. The PDGF-induced depression was blocked by a PDGF-selective tyrosine kinase inhibitor, by a selective inhibitor of phospholipase C-gamma, and by blocking the intracellular release of Ca(2+). Inhibitors of cAMP-dependent protein kinase (PKA) also eliminated the PDGF-induced depression, whereas a phosphodiesterase inhibitor enhanced it. The NMDA receptor-mediated component of excitatory synaptic currents was also inhibited by PDGF, and this inhibition was prevented by co-application of a PKA inhibitor. Src inhibitors also prevented this depression. In recordings from inside-out patches, the catalytic fragment of PKA did not itself alter NMDA single channel activity, but it blocked the up regulation of these channels by a Src activator peptide. Thus, PDGF receptors depress NMDA channels through a Ca(2+)- and PKA-dependent inhibition of their modulation by c-Src. PMID- 10521447 TI - AML3/CBFalpha1 is required for androgen-specific activation of the enhancer of the mouse sex-limited protein (Slp) gene. AB - A complex 120-base pair enhancer, derived from the mouse sex-limited protein (Slp) gene, is activated solely by the androgen receptor (AR) in specific tissues, although it contains a hormone response element recognized by several steroid receptors. The generation of this transcriptional specificity has been ascribed to the interactions of the receptor with tissue-specific nonreceptor factors bound to accessory sites within the enhancer. Protein-DNA interaction assays revealed two factors binding the 5' part of the enhancer that differ widely in abundance between cells showing AR-specific activation of the Slp element compared with those that also permit activation by glucocorticoid receptor (GR). The factor designated B formed a complex centered on the sequence TGTGGT, a core motif recognized by members of the AML/CBFalpha transcription factor family. This complex was competed by a high affinity binding site specific for AML/CBFalpha and was specifically supershifted by an antibody to AML3/CBFalpha1, placing factor B within the AML3/CBFalpha1 subclass. Interestingly, this factor was shown to bind to a second site in the 3' part of the enhancer, positioned between the two critical AR binding sites. Transfection studies revealed that AML1-ETO, a dominant-negative AML/CBFalpha construct, abrogated AR induction of the enhancer, but not of simple hormone response elements. Furthermore, overexpression of AML3/CBFalpha1 could rescue the AML1-ETO repression. Finally, glutathione S-transferase-AML/CBFalpha fusion proteins demonstrated direct interaction between AML/CBFalpha and steroid receptors. Although this interaction was equivalent between AML1/CBFalpha2 and AR or GR, AML3/CBFalpha1 showed stronger interaction with AR than with GR. These data demonstrate that AML3/CBFalpha1 is functionally required for hormonal induction of the Slp enhancer and that direct, preferential protein-protein interactions may contribute to AR-specific activation. These results demonstrate an intriguing role of AML3/CBFalpha1 in steroid- as well as tissue-specific activation of target genes. PMID- 10521448 TI - Common binding sites for beta-amyloid fibrils and fibroblast growth factor-2 in heparan sulfate from human cerebral cortex. AB - Heparan sulfate found in the cerebral plaques of Alzheimer's disease binds to beta-amyloid (Abeta) fibrils. This interaction has been proposed to enhance fibril deposition and mediate Abeta-induced glia activation and neurotoxicity. On the other hand, heparan sulfate augments signaling of fibroblast growth factor-2 (FGF-2), a neuroprotective factor that antagonizes the neurotoxic effects of Abeta. We defined structures in heparan sulfate from human cerebral cortex that bind Abeta fibrils. The minimal binding site is found in N-sulfated hexasaccharide domains and contains critical 2-O-sulfated iduronic acid residues. By contrast, binding of Abeta monomers requires, in addition, 6-O-sulfate groups on glucosamine residues. The binding specificity of fibrillar Abeta is shared by FGF-2, and we here show that cerebral heparan sulfate domains selected for binding to Abeta-(1-40) fibrils bind also to FGF-2. These data suggest that neurotoxic and neuroprotective signals may converge by competing for the same binding sites on the heparan sulfate chain. PMID- 10521449 TI - Regulated migration of epidermal growth factor receptor from caveolae. AB - In quiescent fibroblasts, epidermal growth factor (EGF) receptors (EGFR) are initially concentrated in caveolae but rapidly move out of this membrane domain in response to EGF. To better understand the dynamic localization of EGFR to caveolae, we have studied the behavior of wild-type and mutant receptors expressed in cells lacking endogenous EGFR. All of the receptors we examined, including those missing the first 274 amino acids or most of the cytoplasmic tail, were constitutively concentrated in caveolae. By contrast, migration from caveolae required EGF binding, an active receptor kinase domain, and at least one of the five tyrosine residues present in the regulatory domain of the receptor. Movement appears to be modulated by Src kinase, is blocked by activators of protein kinase C, and occurs independently of internalization by clathrin-coated pits. Two mutant receptors previously shown to induce an oncogenic phenotype lack the ability to move from caveolae in response to EGF, suggesting that a prolonged residence in this domain may contribute to abnormal cell behavior. PMID- 10521450 TI - Syk and Bruton's tyrosine kinase are required for B cell antigen receptor mediated activation of the kinase Akt. AB - Activation of Akt by multiple stimuli including B cell antigen receptor (BCR) engagement requires phosphatidylinositol 3-kinase and regulates processes including cell survival, proliferation, and metabolism. BCR cross-linking activates three families of non-receptor protein tyrosine kinases (PTKs) and these are transducers of signaling events including phospholipase C and mitogen activated protein kinase activation; however, the relative roles of PTKs in BCR mediated Akt activation are unknown. We examined Akt activation in Lyn-, Syk- and Btk-deficient DT40 cells and B cells from Lyn(-/-) mice. BCR-mediated Akt activation required Syk and was partially dependent upon Btk. Increased BCR induced Akt phosphorylation was observed in Lyn-deficient DT40 cells and Lyn(-/-) mice compared with wild-type cells suggesting that Lyn may negatively regulate Akt function. BCR-induced tyrosine phosphorylation of the phosphatidylinositol 3 kinase catalytic subunit was abolished in Syk-deficient cells consistent with a receptor-proximal role for Syk in BCR-mediated phosphatidylinositol 3-kinase activation; in contrast, it was maintained in Btk-deficient cells, suggesting Btk functions downstream of phosphatidylinositol 3-kinase. Calcium depletion did not influence BCR-induced Akt phosphorylation/activation, showing that neither Syk nor Btk mediates its effects via changes in calcium levels. Thus, BCR-mediated Akt stimulation is regulated by multiple non-receptor PTK families which regulate Akt both proximal and distal to phosphatidylinositol 3-kinase activation. PMID- 10521451 TI - Caspase-3 is the primary activator of apoptotic DNA fragmentation via DNA fragmentation factor-45/inhibitor of caspase-activated DNase inactivation. AB - Caspase-3 initiates apoptotic DNA fragmentation by proteolytically inactivating DFF45 (DNA fragmentation factor-45)/ICAD (inhibitor of caspase-activated DNase), which releases active DFF40/CAD (caspase-activated DNase), the inhibitor's associated endonuclease. Here, we examined whether other apoptotic proteinases initiated DNA fragmentation via DFF45/ICAD inactivation. In a cell-free assay, caspases-3, -6, -7, -8, and granzyme B initiated benzoyloxycarbonyl-Asp-Glu-Val Asp (DEVD) cleaving caspase activity, DFF45/ICAD inactivation, and DNA fragmentation, but calpain and cathepsin D failed to initiate these events. Strikingly, only the DEVD cleaving caspases, caspase-3 and caspase-7, inactivated DFF45/ICAD and promoted DNA fragmentation in an in vitro DFF40/CAD assay, suggesting that granzyme B, caspase-6, and caspase-8 promote DFF45/ICAD inactivation and DNA fragmentation indirectly by activating caspase-3 and/or caspase-7. In vitro, however, caspase-3 inactivated DFF45/ICAD and promoted DNA fragmentation more effectively than caspase-7 and endogenous levels of caspase-7 failed to inactivate DFF45/ICAD in caspase-3 null MCF7 cells and extracts. Together, these data suggest that caspase-3 is the primary inactivator of DFF45/ICAD and therefore the primary activator of apoptotic DNA fragmentation. PMID- 10521452 TI - Negative regulation of PYK2/related adhesion focal tyrosine kinase signal transduction by hematopoietic tyrosine phosphatase SHPTP1. AB - Related adhesion focal tyrosine kinase (RAFTK) (also known as PYK2) is a cytoplasmic tyrosine kinase related to the focal adhesion kinase (FAK) p125(FAK). RAFTK is rapidly phosphorylated on tyrosine residues in response to various stimuli, such as tumor necrosis factor-alpha, changes in osmolarity, elevation in intracellular calcium concentration, lysophosphatidic acid, and bradykinin. Overexpression of RAFTK induces activation of c-Jun amino-terminal kinase (also known as stress-activated protein kinase), mitogen-activated protein kinase (MAPK), and p38 MAPK. The present studies demonstrate that RAFTK binds constitutively to the protein tyrosine phosphatase SHPTP1. In contrast to PTP1B, overexpression of wild-type SHPTP1 blocks tyrosine phosphorylation of RAFTK. The results further demonstrate that RAFTK is a direct substrate of SHPTP1 in vitro. Moreover, treatment of PC12 cells with bradykinin is associated with inhibition in tyrosine phosphorylation of RAFTK in the presence of SHPTP1. Furthermore, in contrast to the phosphatase-dead SHPTP1 C453S mutant, overexpression of wild-type SHPTP1 blocks interaction of RAFTK with the SH2-domain of c-Src and inhibits RAFTK-mediated MAPK activation. Significantly, cotransfection of RAFTK with SHPTP1 did not inhibit RAFTK-mediated c-Jun amino-terminal kinase activation. Taken together, these findings suggest that SHPTP1 plays a negative role in PYK2/RAFTK signaling by dephosphorylating RAFTK. PMID- 10521454 TI - Crystal structures of intermediates in the dehalogenation of haloalkanoates by L 2-haloacid dehalogenase. AB - The L-2-haloacid dehalogenase from the 1,2-dichloroethane-degrading bacterium Xanthobacter autotrophicus GJ10 catalyzes the hydrolytic dehalogenation of small L-2-haloalkanoates to their corresponding D-2-hydroxyalkanoates, with inversion of the configuration at the C(2) atom. The structure of the apoenzyme at pH 8 was refined at 1.5-A resolution. By lowering the pH, the catalytic activity of the enzyme was considerably reduced, allowing the crystal structure determination of the complexes with L-2-monochloropropionate and monochloroacetate at 1.7 and 2.1 A resolution, respectively. Both complexes showed unambiguous electron density extending from the nucleophile Asp(8) to the C(2) atom of the dechlorinated substrates corresponding to a covalent enzyme-ester reaction intermediate. The halide ion that is cleaved off is found in line with the Asp(8) Odelta1-C(2) bond in a halide-stabilizing cradle made up of Arg(39), Asn(115), and Phe(175). In both complexes, the Asp(8) Odelta2 carbonyl oxygen atom interacts with Thr(12), Ser(171), and Asn(173), which possibly constitute the oxyanion hole in the hydrolysis of the ester bond. The carboxyl moiety of the substrate is held in position by interactions with Ser(114), Lys(147), and main chain NH groups. The L 2-monochloropropionate CH(3) group is located in a small pocket formed by side chain atoms of Lys(147), Asn(173), Phe(175), and Asp(176). The size and position of the pocket explain the stereospecificity and the limited substrate specificity of the enzyme. These crystallographic results demonstrate that the reaction of the enzyme proceeds via the formation of a covalent enzyme-ester intermediate at the nucleophile Asp(8). PMID- 10521453 TI - The T-superfamily of conotoxins. AB - We report the discovery and initial characterization of the T-superfamily of conotoxins. Eight different T-superfamily peptides from five Conus species were identified; they share a consensus signal sequence, and a conserved arrangement of cysteine residues (- -CC- -CC-). T-superfamily peptides were found expressed in venom ducts of all major feeding types of Conus; the results suggest that the T-superfamily will be a large and diverse group of peptides, widely distributed in the 500 different Conus species. These peptides are likely to be functionally diverse; although the peptides are small (11-17 amino acids), their sequences are strikingly divergent, with different peptides of the superfamily exhibiting varying extents of post-translational modification. Of the three peptides tested for in vivo biological activity, only one was active on mice but all three had effects on fish. The peptides that have been extensively characterized are as follows: p5a, GCCPKQMRCCTL*; tx5a, gammaCCgammaDGW(+)CCT( section sign)AAO; and au5a, FCCPFIRYCCW (where gamma = gamma-carboxyglutamate, W(+) = bromotryptophan, O = hydroxyproline, T( section sign) = glycosylated threonine, and * = COOH terminal amidation). We also demonstrate that the precursor of tx5a contains a functional gamma-carboxylation recognition signal in the -1 to -20 propeptide region, consistent with the presence of gamma-carboxyglutamate residues in this peptide. PMID- 10521455 TI - Methanopyrus kandleri glutamyl-tRNA reductase. AB - The initial reaction of tetrapyrrole formation in archaea is catalyzed by a NADPH dependent glutamyl-tRNA reductase (GluTR). The hemA gene encoding GluTR was cloned from the extremely thermophilic archaeon Methanopyrus kandleri and overexpressed in Escherichia coli. Purified recombinant GluTR is a tetrameric enzyme with a native M(r) = 190,000 +/- 10,000. Using a newly established enzyme assay, a specific activity of 0.75 nmol h(-1) mg(-1) at 56 degrees C with E. coli glutamyl-tRNA as substrate was measured. A temperature optimum of 90 degrees C and a pH optimum of 8.1 were determined. Neither heme cofactor, nor flavin, nor metal ions were required for GluTR catalysis. Heavy metal compounds, Zn(2+), and heme inhibited the enzyme. GluTR inhibition by the newly synthesized inhibitor glutamycin, whose structure is similar to the 3' end of the glutamyl-tRNA substrate, revealed the importance of an intact chemical bond between glutamate and tRNA(Glu) for substrate recognition. The absolute requirement for NADPH in the reaction of GluTR was demonstrated using four NADPH analogues. Chemical modification and site-directed mutagenesis studies indicated that a single cysteinyl residue and a single histidinyl residue were important for catalysis. It was concluded that during GluTR catalysis the highly reactive sulfhydryl group of Cys-48 acts as a nucleophile attacking the alpha-carbonyl group of tRNA-bound glutamate with the formation of an enzyme-localized thioester intermediate and the concomitant release of tRNA(Glu). In the presence of NADPH, direct hydride transfer to enzyme-bound glutamate, possibly facilitated by His-84, leads to glutamate-1-semialdehyde formation. In the absence of NADPH, a newly discovered esterase activity of GluTR hydrolyzes the highly reactive thioester of tRNA(Glu) to release glutamate. PMID- 10521456 TI - Activation of interferon regulatory factor 3 in response to DNA-damaging agents. AB - Genotoxic stress triggers signal transduction pathways that mediate either the protection or apoptosis of affected cells. The interferon regulatory factors (IRFs) are involved in a wide range of host defense mechanisms against environmental stresses. Treatment with DNA-damaging agents, including doxorubicin and UV radiation, caused phosphorylation of the IRF3 transcription factor. Phosphorylation of IRF3 induced its interaction with the transcriptional co activator cAMP-response element binding protein-binding protein. Furthermore, genotoxic stress-induced phosphorylation of IRF3 resulted in its movement from the cytoplasm to the nucleus, where it activated transcription from its binding site. These observations suggest that IRF3 plays a role in the defensive responses induced by genotoxic stress. PMID- 10521457 TI - Yeast topoisomerase II is inhibited by etoposide after hydrolyzing the first ATP and before releasing the second ADP. AB - Topoisomerase II-catalyzed DNA transport requires coordination between two distinct reactions: ATP hydrolysis and DNA cleavage/religation. To further understand how these reactions are coupled, inhibition by the clinically used anticancer drug etoposide was studied. The IC(50) for perturbing the DNA cleavage/religation equilibrium is nucleotide-dependent; its value is 6 microM in the presence of ATP, 25 microM in the presence of a nonhydrolyzable ATP analog, and 45 microM in the presence of ADP or no nucleotide. This inhibition was further characterized using steady-state and pre-steady-state ATPase and decatenation assays. Etoposide is a hyperbolic noncompetitive inhibitor of the ATPase activity with a K(i)(app) of 5.6 microM no inhibition of ATP hydrolysis is seen in the absence of DNA cleavage. In order to determine which steps of the ATPase mechanism etoposide inhibits, pre-steady-state analysis was performed. These results showed that etoposide does not reduce the rate of binding two ATP, hydrolyzing the first ATP, or releasing the second ADP. Inhibition is therefore associated with the first product release step or hydrolysis of the second ATP, suggesting that DNA religation normally occurs at one of these two steps. Multiple turnover decatenation is inhibited when etoposide is present; however, single turnover decatenation occurs normally. The implications of these results are discussed in terms of their contribution to our current model for the topoisomerase II mechanism. PMID- 10521458 TI - The COOH terminus of aminoglycoside phosphotransferase (3')-IIIa is critical for antibiotic recognition and resistance. AB - The aminoglycoside phosphotransferases (APHs) are widely distributed among pathogenic bacteria and are employed to covalently modify, and thereby detoxify, the clinically relevant aminoglycoside antibiotics. The crystal structure for one of these aminoglycoside kinases, APH(3')-IIIa, has been determined in complex with ADP and analysis of the electrostatic surface potential indicates that there is a large anionic depression present adjacent to the terminal phosphate group of the nucleotide. This region also includes a conserved COOH-terminal alpha-helix that contains the COOH-terminal residue Phe(264). We report here mutagenesis and computer modeling studies aimed at examining the mode of aminoglycoside binding to APH(3')-IIIa. Specifically, seven site mutants were studied, five from the COOH-terminal helix (Asp(261), Glu(262), and Phe(264)), and two additional residues that line the wall of the anionic depression (Tyr(55) and Arg(211)). Using a molecular modeling approach, six ternary complexes of APH(3')-IIIa.ATP with the antibiotics, kanamycin, amikacin, butirosin, and ribostamycin were independently constructed and these agree well with the mutagenesis data. The results obtained show that the COOH-terminal carboxylate of Phe(264) is critical for proper function of the enzyme. Furthermore, these studies demonstrate that there exists multiple binding modes for the aminoglycosides, which provides a molecular basis for the broad substrate- and regiospecificity observed for this enzyme. PMID- 10521459 TI - Cell surface localization of tissue transglutaminase is dependent on a fibronectin-binding site in its N-terminal beta-sandwich domain. AB - Increasing evidence indicates that tissue transglutaminase (tTG) plays a role in the assembly and remodeling of extracellular matrices and promotes cell adhesion. Using an inducible system we have previously shown that tTG associates with the extracellular matrix deposited by stably transfected 3T3 fibroblasts overexpressing the enzyme. We now show by confocal microscopy that tTG colocalizes with pericellular fibronectin in these cells, and by immunogold electron microscopy that the two proteins are found in clusters at the cell surface. Expression vectors encoding the full-length tTG or a N-terminal truncated tTG lacking the proposed fibronectin-binding site (fused to the bacterial reporter enzyme beta-galactosidase) were generated to characterize the role of fibronectin in sequestration of tTG in the pericellular matrix. Enzyme linked immunosorbent assay style procedures using extracts of transiently transfected COS-7 cells and immobilized fibronectin showed that the truncation abolished fibronectin binding. Similarly, the association of tTG with the pericellular matrix of cells in suspension or with the extracellular matrix deposited by cell monolayers was prevented by the truncation. These results demonstrate that tTG binds to the pericellular fibronectin coat of cells via its N-terminal beta-sandwich domain and that this interaction is crucial for cell surface association of tTG. PMID- 10521460 TI - Differential expression of multiple transglutaminases in human brain. Increased expression and cross-linking by transglutaminases 1 and 2 in Alzheimer's disease. AB - The transglutaminase (TGase) family of enzymes, of which seven different members are known in the human genome, participate in many biological processes involving cross-linking proteins into large macromolecular assemblies. The TGase 2 enzyme is known to be present in neuronal tissues and may play a role in neuronal degenerative diseases such as Alzheimer's disease (AD) by aberrantly cross linking proteins. In this paper, we demonstrate by reverse transcriptase polymerase chain reaction and immunological methods with specific antibodies that in fact three members, the TGase 1, TGase 2, and TGase 3 enzymes, and are differentially expressed in various regions of normal human brain tissues. Interestingly, the TGase 1 and 3 enzymes and their proteolytically processed forms are involved in terminal differentiation programs of epithelial cell development and barrier function. In addition, we found that the levels of expression and activity of the TGase 1 and 2 enzymes were both increased in the cortex and cerebellum of AD patients. Furthermore, whereas normal brain tissues contain approximately 1 residue of cross-link/10,000 residues, AD patient cortex and cerebellum tissues contain 30-50 residues of cross-link/10,000 residues. Together, these findings suggest that multiple TGase enzymes are involved in normal neuronal structure and function, but their elevated expression and cross linking activity may also contribute to neuronal degenerative disease. PMID- 10521461 TI - Protein kinase C recognizes the protein kinase A-binding motif of nonstructural protein 3 of hepatitis C virus. AB - The nonstructural protein 3 (NS3) of hepatitis C virus (HCV) inhibits the nuclear transport and the enzymatic activity of the catalytic subunit of protein kinase A. This inhibition is mediated by an arginine-rich domain localized between amino acids 1487-1500 of the HCV polyprotein. The data presented here indicate that the arginine-rich domain, when embedded in recombinant fragments of NS3, interacts with the catalytic site of protein kinase C (PKC) and inhibits the phosphorylation mediated by this enzyme in vitro and in vivo. Furthermore, a direct binding of PKC to the NS3 fragments leads to an inhibition of the free shuttling of the kinase between the cytoplasm and the particulate fraction. In contrast, a peptide corresponding to the arginine-rich domain (HCV (1487-1500)), despite also being a PKC inhibitor, did not influence the PKC shuttling process and was transported to the particulate fraction by the translocating kinase upon activation with tetradecanoylphorbol-13-acetate. Using the tetradecanoylphorbol 13-acetate -stimulated respiratory burst of NS3-introduced neutrophils as a model system, we could demonstrate that NS3 is able to block PKC-mediated functions within intact cells. Our data support the possibility that NS3 disrupts the PKC mediated signal transduction. PMID- 10521462 TI - TNIK, a novel member of the germinal center kinase family that activates the c Jun N-terminal kinase pathway and regulates the cytoskeleton. AB - Germinal center kinases (GCKs) compose a subgroup of the Ste20 family of kinases. Here we describe the cloning and characterization of a novel GCK family kinase, Traf2- and Nck-interacting kinase (TNIK) that interacts with both Traf2 and Nck. TNIK encodes a polypeptide of 1360 amino acids with eight spliced isoforms. It has 90% amino acid identity to the Nck-interacting kinase in both the N-terminal kinase domain and the C-terminal germinal center kinase homology region. The homology drops to 53% in the intermediate region. TNIK specifically activates the c-Jun N-terminal kinase pathway when transfected into Phoenix-A cells (derivatives of 293 cells), similar to many GCKs. However, in contrast to other GCKs, this activation is mediated solely by the GCK homology region of TNIK. In addition, in Phoenix-A, NIH-3T3, and Hela cells, overexpression of wild type TNIK, but not the kinase mutant form of TNIK, results in the disruption of F actin structure and the inhibition of cell spreading. Furthermore, TNIK can phosphorylate Gelsolin in vitro. This is the first time that a GCK family kinase is shown to be potentially involved in the regulation of cytoskeleton. PMID- 10521463 TI - Divergent signaling pathways link focal adhesion kinase to mitogen-activated protein kinase cascades. Evidence for a role of paxillin in c-Jun NH(2)-terminal kinase activation. AB - Stimulation of a number of cell surface receptors, including integrins and G protein-coupled receptors, results in the activation of a non-receptor tyrosine kinase known as focal adhesion kinase (FAK). In turn, this kinase is believed to play a critical role in signaling to intracellular kinase cascades controlling gene expression such as extracellular signal-regulated kinases (ERKs), by a yet poorly defined mechanism. Furthermore, whether this tyrosine kinase also mediates the activation of other mitogen-activated protein kinase family members, such as c-Jun NH(2)-terminal kinases (JNKs), is still unclear. We show here that the activation of FAK by anchoring to the cell membrane is itself sufficient to stimulate potently both ERK and JNK. These effects were found to be phosphatidylinositol 3-kinase-independent, as FAK effectively stimulated Akt, and wortmannin suppressed Akt but not ERK or JNK activation. As previously reported by others, activation of ERK correlated with the ability of FAK to induce tyrosine phosphorylation of Shc. Surprisingly, however, stimulation of JNK was not dependent on the kinase activity of FAK or on the ability to induce tyrosine phosphorylation of FAK substrates. Instead, we provide evidence that FAK may stimulate JNK through a novel pathway involving the recruitment of paxillin to the plasma membrane and the subsequent activation of a biochemical route dependent on small GTP-binding proteins of the Rho family. PMID- 10521464 TI - Mononuclear leukocytes preferentially bind via CD44 to hyaluronan on human intestinal mucosal smooth muscle cells after virus infection or treatment with poly(I.C). AB - Pathological changes in inflammatory bowel disease include an increase in intestinal mucosal mononuclear leukocytes and hyperplasia of the muscularis mucosae smooth muscle cells (M-SMCs). Because virus infections have correlated with disease flare, we tested the response of cultured M-SMCs to respiratory syncytial virus, measles virus, and the viral analogue, poly(I.C). Adhesion of U937 cells and peripheral blood mononuclear cells was used to measure the leukocyte-interactive potential of M-SMCs. Untreated M-SMCs, only minimally adhesive for leukocytes, bound U937 cells after treatment with respiratory syncytial virus or measles virus. Mononuclear leukocytes also bound to poly(I.C) treated M-SMCs. Although both vascular cell adhesion molecule-1 mRNA and protein increased 3-4-fold in poly(I.C)-treated M-SMC cultures, U937 cell adhesion was not blocked by an anti-vascular cell adhesion molecule-1 monoclonal antibody. However, hyaluronidase digestion of poly(I.C)- or virus-treated M-SMCs dramatically reduced leukocyte adhesion ( approximately 75%). Fluorophore assisted carbohydrate electrophoresis demonstrated a approximately 3-fold increase in surface-bound hyaluronan on poly(I.C)-treated M-SMCs compared with untreated controls. In addition, pretreatment of mononuclear cells with a blocking anti-CD44 antibody, greatly decreased adhesion to poly(I.C)-treated M SMCs. Recognition of this virus-induced hyaluronan/CD44 mechanism of mesenchymal cell/leukocyte interaction introduces a new avenue in the research of gut inflammation. PMID- 10521465 TI - The fibronectin extra domain A activates matrix metalloproteinase gene expression by an interleukin-1-dependent mechanism. AB - The extra domain-A (EDA), present in fibronectin (FN) molecules arising from alternatively spliced transcripts, appears only during specific biological and pathogenic processes. However, its function is poorly understood. To define the physiologic role of this domain in joint connective tissue, the biological effects on rabbit cartilage explants, chondrocytes, and synovial cells were studied. A recombinant EDA protein (rEDA) increased proteoglycan release (3. 6 fold) in cartilage explant cultures and markedly induced production of matrix metalloproteinase (MMP)-1 in chondrocytes. In addition, rEDA induced MMP-1, MMP 3, and MMP-9 in synovial cells. These effects were elicited only by rEDA, while its neighboring type III repeats, III(11) or III(12), scarcely had any such effects. Interestingly, reorganization of F-actin stress fibers accompanied MMP-1 expression in synovial cells treated with rEDA, suggesting alteration of cellular phenotype. Subsequent Northern blotting revealed expression of pro-inflammatory cytokines, including interleukin (IL)-1alpha and IL-1beta, was induced by rEDA prior to MMP-1 expression. Delayed MMP-1 expression suggests that rEDA-induced IL 1s promote MMP-1 expression in an autocrine manner. This hypothesis is supported by the reduction of EDA-induced MMP-1 production by IL-1 receptor antagonist. The effect of EDA on MMP-1 production was reduced by connection with an adjacent type III repeat on either the NH(2) or COOH side of EDA and was abolished by connection on both sides of EDA, suggesting that exposure of either the NH(2) or COOH terminus of EDA domain by proteolytic cleavage releases the inducing activity. In agreement with these results, full-length cellular FN did not induce MMP-1 production. Furthermore, a 160-kDa EDA-positive FN fragment, which was purified from human placental tissue and corresponds to the region from NH(2) terminus through the EDA, induced MMP-1 production. Taken together, these results suggest that the EDA in FN fragments triggers alterations of cell physiology and plays a role in matrix degradation in joint connective tissue. PMID- 10521466 TI - Presenilin 1 protein directly interacts with Bcl-2. AB - Presenilin proteins are involved in familial Alzheimer's disease, a neurodegenerative disorder characterized by massive death of neurons. We describe a direct interaction between presenilin 1 (PS1) and Bcl-2, a key factor in the regulation of apoptosis, by yeast two-hybrid interaction system, by co immunoprecipitation, and by cross-linking experiments. Our data show that PS1 and Bcl-2 assemble into a macromolecular complex, and that they are released from this complex in response to an apoptotic stimulus induced by staurosporine. The results support the idea of cross-talk between these two proteins during apoptosis. PMID- 10521467 TI - Characterization of the active site of ADP-ribosyl cyclase. AB - ADP-ribosyl cyclase synthesizes two Ca(2+) messengers by cyclizing NAD to produce cyclic ADP-ribose and exchanging nicotinic acid with the nicotinamide group of NADP to produce nicotinic acid adenine dinucleotide phosphate. Recombinant Aplysia cyclase was expressed in yeast and co-crystallized with a substrate, nicotinamide. x-ray crystallography showed that the nicotinamide was bound in a pocket formed in part by a conserved segment and was near the central cleft of the cyclase. Glu(98), Asn(107) and Trp(140) were within 3.5 A of the bound nicotinamide and appeared to coordinate it. Substituting Glu(98) with either Gln, Gly, Leu, or Asn reduced the cyclase activity by 16-222-fold, depending on the substitution. The mutant N107G exhibited only a 2-fold decrease in activity, while the activity of W140G was essentially eliminated. The base exchange activity of all mutants followed a similar pattern of reduction, suggesting that both reactions occur at the same active site. In addition to NAD, the wild-type cyclase also cyclizes nicotinamide guanine dinucleotide to cyclic GDP-ribose. All mutant enzymes had at least half of the GDP-ribosyl cyclase activity of the wild type, some even 2-3-fold higher, indicating that the three coordinating amino acids are responsible for positioning of the substrate but not absolutely critical for catalysis. To search for the catalytic residues, other amino acids in the binding pocket were mutagenized. E179G was totally devoid of GDP-ribosyl cyclase activity, and both its ADP-ribosyl cyclase and the base exchange activities were reduced by 10,000- and 18,000-fold, respectively. Substituting Glu(179) with either Asn, Leu, Asp, or Gln produced similar inactive enzymes, and so was the conversion of Trp(77) to Gly. However, both E179G and the double mutant E179G/W77G retained NAD-binding ability as shown by photoaffinity labeling with [(32)P]8-azido-NAD. These results indicate that both Glu(179) and Trp(77) are crucial for catalysis and that Glu(179) may indeed be the catalytic residue. PMID- 10521468 TI - DECAY, a novel Drosophila caspase related to mammalian caspase-3 and caspase-7. AB - Caspases are key effectors of programmed cell death in metazoans. In Drosophila, four caspases have been described so far. Here we describe the identification and characterization of the fifth Drosophila caspase, DECAY. DECAY shares a high degree of homology with the members of the mammalian caspase-3 subfamily, particularly caspase-3 and caspase-7. DECAY lacks a long prodomain and thus appears to be a class II effector caspase. Ectopic expression of DECAY in cultured cells induces apoptosis. Recombinant DECAY exhibited substrate specificity similar to the mammalian caspase-3 subfamily. Low levels of decay mRNA are ubiquitously expressed in Drosophila embryos during early stages of development but its expression becomes somewhat spatially restricted in some tissues. During oogenesis decay mRNA was detected in egg chambers of all stages consistent with a role for DECAY in apoptosis of nurse cells. Relatively high levels of decay mRNA are expressed in larval salivary glands and midgut, two tissues which undergo histolysis during larval/pupal metamorphosis, suggesting that DECAY may play a role in developmentally programmed cell death in Drosophila. PMID- 10521469 TI - Identification of a new member of the tryptase family of mouse and human mast cell proteases which possesses a novel COOH-terminal hydrophobic extension. AB - Mapping of the tryptase locus on chromosome 17 revealed a novel gene 2.3 kilobase 3' of the mouse mast cell protease (mMCP) 6 gene. This 3.7-kilobase gene encodes the first example of a protease in the tryptase family that contains a membrane spanning segment located at its COOH terminus. Comparative structural studies indicated that the putative transmembrane tryptase (TMT) possesses a unique substrate-binding cleft. As assessed by RNA blot analyses, mTMT is expressed in mice in both strain- and tissue-dependent manners. Thus, different transcriptional and/or post-transcriptional mechanisms are used to control the expression of mTMT in vivo. Analysis of the corresponding tryptase locus in the human genome resulted in the isolation and characterization of the hTMT gene. The hTMT transcript is expressed in numerous tissues and is also translated. Analysis of the tryptase family of genes in mice and humans now indicates that a primordial serine protease gene duplicated early and often during the evolution of mammals to generate a panel of homologous tryptases in each species that differ in their tissue expression, substrate specificities, and physical properties. PMID- 10521470 TI - Involvement of small GTPases in Mycoplasma fermentans membrane lipoproteins mediated activation of macrophages. AB - Mycoplasma fermentans lipoproteins (LAMPf) are capable of activating macrophages and inducing the secretion of proinflammatory cytokines. We have recently reported that mitogen-activated protein kinase (MAPK) pathways and NF-kappaB and activated protein 1 (AP-1) play a crucial role in the activation induced by this bacterial compound. To further elucidate the mechanisms by which LAMPf mediate the activation of macrophages, we assessed the effects of inhibiting small G proteins Rac, Cdc42, and Rho. The Rho-specific inhibitor C3 enzyme completely abolished the secretion of tumor necrosis factor alpha by macrophages stimulated with LAMPf and also inhibited the activation of extracellular signal-regulated kinase (ERK), c-Jun NH(2)-terminal kinase (JNK), and p38 kinase. In addition, we have shown that LAMPf stimulate Cdc42 and that inhibition of Cdc42 or Rac by dominant negative mutants abrogates LAMPf-mediated activation of JNK and transactivation of NF-kappaB and AP-1 in the murine macrophage cell line RAW 264.7. These results indicate that small G proteins Rho, Cdc42, and Rac are involved in the cascade of events leading to the macrophage activation by mycoplasma lipoproteins. PMID- 10521471 TI - The pharmacological and functional characteristics of the serotonin 5-HT(3A) receptor are specifically modified by a 5-HT(3B) receptor subunit. AB - While homomers containing 5-HT(3A) subunits form functional ligand-gated serotonin (5-HT) receptors in heterologous expression systems (Jackson, M. B., and Yakel, J. L. (1995) Annu. Rev. Physiol. 57, 447-468; Lambert, J. J., Peters, J. A., and Hope, A. G. (1995) in Ligand-Voltage-Gated Ion Channels (North, R., ed) pp. 177-211, CRC Press, Inc., Boca Raton, FL), it has been proposed that native receptors may exist as heteromers (Fletcher, S., and Barnes, N. M. (1998) Trends Pharmacol. Sci. 19, 212-215). We report the cloning of a subunit 5-HT(3B) with approximately 44% amino acid identity to 5-HT(3A) that specifically modified 5-HT(3A) receptor kinetics, voltage dependence, and pharmacology. Co-expression of 5-HT(3B) with 5-HT(3A) modified the duration of 5-HT(3) receptor agonist induced responses, linearized the current-voltage relationship, increased agonist and antagonist affinity, and reduced cooperativity between subunits. Reverse transcriptase-polymerase chain reaction in situ hybridization revealed co localization of both 5-HT(3B) and 5-HT(3A) in a population of neurons in the amygdala, telencephalon, and entorhinal cortex. Furthermore, 5-HT(3A) and 5 HT(3B) mRNAs were expressed in spleen and intestine. Our data suggest that 5 HT(3B) might contribute to tissue-specific functional changes in 5-HT(3)-mediated signaling and/or modulation. PMID- 10521472 TI - Inhibitor binding studies on enoyl reductase reveal conformational changes related to substrate recognition. AB - Enoyl acyl carrier protein reductase (ENR) is involved in fatty acid biosynthesis. In Escherichia coli this enzyme is the target for the experimental family of antibacterial agents, the diazaborines, and for triclosan, a broad spectrum antimicrobial agent. Biochemical studies have suggested that the mechanism of diazaborine inhibition is dependent on NAD(+) and not NADH, and resistance of Brassica napus ENR to diazaborines is thought to be due to the replacement of a glycine in the active site of the E. coli enzyme by an alanine at position 138 in the plant homologue. We present here an x-ray analysis of crystals of B. napus ENR A138G grown in the presence of either NAD(+) or NADH and the structures of the corresponding ternary complexes with thienodiazaborine obtained either by soaking the drug into the crystals or by co-crystallization of the mutant with NAD(+) and diazaborine. Analysis of the ENR A138G complex with diazaborine and NAD(+) shows that the site of diazaborine binding is remarkably close to that reported for E. coli ENR. However, the structure of the ternary ENR A138G-NAD(+)-diazaborine complex obtained using co-crystallization reveals a previously unobserved conformational change affecting 11 residues that flank the active site and move closer to the nicotinamide moiety making extensive van der Waals contacts with diazaborine. Considerations of the mode of substrate binding suggest that this conformational change may reflect a structure of ENR that is important in catalysis. PMID- 10521473 TI - Crystal structure of chitosanase from Bacillus circulans MH-K1 at 1.6-A resolution and its substrate recognition mechanism. AB - Chitosanase from Bacillus circulans MH-K1 is a 29-kDa extracellular protein composed of 259 amino acids. The crystal structure of chitosanase from B. circulans MH-K1 has been determined by multiwavelength anomalous diffraction method and refined to crystallographic R = 19.2% (R(free) = 23.5%) for the diffraction data at 1.6-A resolution collected by synchrotron radiation. The enzyme has two globular upper and lower domains, which generate the active site cleft for the substrate binding. The overall molecular folding is similar to chitosanase from Streptomyces sp. N174, although there is only 20% identity at the amino acid sequence level between both chitosanases. However, there are three regions in which the topology is remarkably different. In addition, the disulfide bridge between Cys(50) and Cys(124) joins the beta1 strand and the alpha7 helix, which is not conserved among other chitosanases. The orientation of two backbone helices, which connect the two domains, is also different and is responsible for the differences in size and shape of the active site cleft in these two chitosanases. This structural difference in the active site cleft is the reason why the enzymes specifically recognize different substrates and catalyze different types of chitosan degradation. PMID- 10521474 TI - Post-transcriptional adenylation of signal recognition particle RNA is carried out by an enzyme different from mRNA Poly(A) polymerase. AB - A fraction of the signal recognition particle (SRP) RNA from human, rat, Xenopus, and Saccharomyces cerevisiae cells contains a single post-transcriptionally added adenylic acid residue on its 3'-end; in the case of human SRP RNA, over 60% of the SRP RNA molecules contain a nontemplated adenylic acid residue on their 3' ends (Sinha, K. M., Gu, J., Chen, Y., and Reddy, R. (1998) J. Biol. Chem. 273, 6853-6859). In this study, we investigated the enzyme that is involved in this 3' end adenylation of SRP RNA. A U1A protein peptide conjugated to albumin completely inhibited the polyadenylation of a SV40 mRNA by HeLa cell nuclear extract in vitro; however, the 3'-end adenylation of human SRP RNA or Alu RNA, which corresponds to 5' and 3'-ends of SRP RNA, was not affected by this U1A peptide conjugate. SRP RNA from mutant strains of S. cerevisiae with a temperature-sensitive mRNA poly(A) polymerase grown at a restrictive temperature of 37 degrees C also contained a post-transcriptionally added adenylic acid residue just like SRP RNA from wild-type cells and mutant cells grown at permissive temperature of 23 degrees C. In addition, binding of SRP 9/14-kDa protein heterodimer was required for adenylation of Alu RNA in vitro. These lines of evidence, along with other data, show that post-transcriptional adenylation of SRP and Alu RNAs is carried out by a novel enzyme that is distinct from the mRNA poly(A) polymerase, CCA-adding enzyme, and nonspecific terminal transferase. PMID- 10521475 TI - Nuclear protein binding at the beta-myosin heavy chain A/T-rich element is enriched following increased skeletal muscle activity. AB - In adult mouse skeletal muscle, beta-myosin heavy chain (betaMyHC) gene expression is primarily restricted to slow-type I fibers but can be induced in fast-type II fibers by mechanical overload (MOV). Our previous transgenic analyses have delimited an 89-base pair (bp) MOV-responsive region (-293 to 205), and shown that mutation of the MCAT and C-rich elements within this region did not abolish betaMyHC transgene induction by MOV. In this study we describe an A/T-rich element (betaA/T-rich; -269 5'-GGAGATATTTTT-3' -258) located within this 89-bp region that, only under MOV conditions, revealed enriched binding as characterized by electrophoretic mobility shift assays and dimethyl sulfate and diethyl pyrocarbonate interference footprinting. Direct, competition, and supershift electrophoretic mobility shift assays revealed highly enriched specific binding activity at the betaA/T-rich element that was antigenically distinct from GATA-4, MEF2A-D, SRF, and Oct-1, nuclear proteins that were previously shown to bind A/T-rich elements. In vitro translated GATA-4, MEF2C, SRF, and Oct-1 bound to consensus GATA, MEF2, SRE, and Oct-1 elements, respectively, but not to the betaA/T-rich element. Two-dimensional UV cross linking of the bromodeoxyuridine-substituted betaA/T-rich element with mechanically overloaded plantaris (MOV-P) nuclear extract detected two proteins (44 and 48 kDa). Our results indicate that the betaA/T-rich element may function in vivo as a betaMyHC MOV-inducible element during hypertrophy of adult skeletal muscle by binding two distinct proteins identified only in MOV-P nuclear extract. PMID- 10521476 TI - Embryonic lethality and defective neural tube closure in mice lacking squalene synthase. AB - Squalene synthase (SS) catalyzes the reductive head-to-head condensation of two molecules of farnesyl diphosphate to form squalene, the first specific intermediate in the cholesterol biosynthetic pathway. We used gene targeting to knock out the mouse SS gene. The mice heterozygous for the mutation (SS+/-) were apparently normal. SS+/- mice showed 60% reduction in the hepatic mRNA levels of SS compared with SS+/+ mice. Consistently, the SS enzymatic activities were reduced by 50% in the liver and testis. Nevertheless, the hepatic cholesterol synthesis was not different between SS+/- and SS+/+ mice, and plasma lipoprotein profiles were not different irrespective of the presence of the low density lipoprotein receptor, indicating that SS is not a rate-limiting enzyme in the cholesterol biosynthetic pathway. The mice homozygous for the disrupted SS gene (SS-/-) were embryonic lethal around midgestation. E9.5-10.5 SS-/- embryos exhibited severe growth retardation and defective neural tube closure. The lethal phenotype was not rescued by supplementing the dams either with dietary squalene or cholesterol. We speculate that cholesterol is required for the development, particularly of the nervous system, and that the chorioallantoic circulatory system is not mature enough to supply the rapidly growing embryos with maternal cholesterol at this developmental stage. PMID- 10521477 TI - Glycine insertion in the hinge region of lactose repressor protein alters DNA binding. AB - Amino acid alterations were designed at the C terminus of the hinge segment (amino acids approximately 51-59) that links two functional domains within lactose repressor protein (LacI). Gly was introduced between Gly(58) and Lys(59) to generate Gly(58+1); Gln(60) was changed to Gly or Pro, and up to three additional glycines were inserted following Gln(60) --> Gly. All mutant proteins exhibited purification behavior, CD spectra, assembly state, and inducer binding properties similar to wild-type LacI and only small differences in trypsin proteolysis patterns. In contrast, significant differences were observed in DNA binding properties. Gly(58+1) exhibited a decrease of approximately 100-fold in affinity for O(1) operator, and sequential Gly insertion C-terminal to Gln(60) - > Gly resulted in progressively decreased affinity for O(1) operator, approaching nonspecific levels for insertion of >/=2 glycines. Where sufficient affinity for O(1) operator existed, decreased binding to O(1) in the presence of inducer indicated no disruption in the allosteric response for these proteins. Collectively, these results indicate that flexibility and/or spacing between the core and N-terminal domains did not significantly affect folding or assembly, but these alterations in the hinge domain profoundly altered affinity of the lactose repressor protein for its wild-type target sequence. PMID- 10521478 TI - The p38 mitogen-activated protein kinase is required for NF-kappaB-dependent gene expression. The role of TATA-binding protein (TBP). AB - Endotoxin-induced cytokine gene transcription in monocytes and macrophages is regulated in part by NF-kappaB. We have previously shown that the p38 mitogen activated protein (MAP) kinase is necessary for endotoxin-induced cytokine gene transcription. Due to the fact that most cytokine promoter sequences have active NF-kappaB sites, we hypothesized that the p38 MAP kinase was necessary for NF kappaB-dependent gene expression. We found that NF-kappaB-dependent gene expression was reduced to near control levels with either SB 203580 or a dominant negative p38 MAP kinase expression vector. Inhibition of the p38 MAP kinase did not alter NF-kappaB activation at any level, but it significantly reduced the DNA binding of TATA-binding protein (TBP) to the TATA box. The dominant-negative p38 MAP kinase expression vector interfered with the direct interaction of native TFIID (TBP) with a co-transfected p65 fusion protein. Likewise, this dominant negative plasmid also interfered with the direct interaction of a co-transfected TBP fusion protein with the native p65 subunit. The p38 kinase also phosphorylated TFIID (TBP) in vitro, and SB 203580 inhibited phosphorylation of TFIID (TBP) in vivo. Thus, the p38 MAP kinase regulates NF-kappaB-dependent gene transcription, in part, by modulating activation of TFIID (TBP). PMID- 10521479 TI - Differences in signaling properties of the cytoplasmic domains of the insulin receptor and insulin-like growth factor receptor in 3T3-L1 adipocytes. AB - Insulin and insulin-like growth factors (IGFs) elicit distinct but overlapping biological effects in vivo. To investigate whether differences in intrinsic signaling capacity of receptors contribute to biological specificity, we constructed chimeric receptors containing the extracellular portion of the neurotrophin receptor TrkC fused to the intracellular portion of the insulin or IGF-I receptors. Chimeras were stably expressed in 3T3-L1 adipocytes at levels comparable to endogenous insulin receptors and were efficiently activated by neurotrophin-3. The wild-type insulin receptor chimera mediated approximately 2 fold greater phosphorylation of insulin receptor substrate 1 (IRS-1), association of IRS-1 with phosphoinositide 3-kinase, stimulation of glucose uptake, and GLUT4 translocation, compared with the IGF-I receptor chimera. In contrast, the IGF-I receptor chimera mediated more effective Shc phosphorylation, association of Shc with Grb2, and activation of mitogen-activated protein kinase compared with the insulin receptor chimera. The two receptors elicited similar activation of protein kinase B, p70S6 kinase, and glycogen synthesis. We conclude that the insulin receptor mediates some aspects of metabolic signaling in adipocytes more effectively than the IGF-I receptor, as a consequence of more efficient phosphorylation of IRS-1 and greater recruitment/activation of phosphoinositide 3 kinase. PMID- 10521480 TI - The PEST domain of IkappaBalpha is necessary and sufficient for in vitro degradation by mu-calpain. AB - Polypeptide sequences enriched in proline (P), glutamate (E), serine (S), and threonine (T), dubbed PEST domains, are proposed to expedite the degradation of proteins. The proteolysis of one PEST-containing protein, IkappaBalpha, is prerequisite to the activation of the transcription factor NF-kappaB. Two mechanisms of IkappaBalpha degradation in vivo have been described, one well characterized through the ubiquitin-proteasome pathway, and another less characterized through calpain. In this report, a mutational analysis was done to identify any regions of IkappaBalpha that facilitate its recognition and proteolysis by calpain in vitro. These studies revealed that the PEST sequence of IkappaBalpha is critical for its calpain-dependent degradation. Furthermore, the IkappaBalpha-PEST domain binds to the calmodulin-like domain of the large subunit of mu-calpain (muCaMLD). Transfer of the IkappaBalpha-PEST domain to a protein incapable of either binding to or being degraded by mu-calpain allowed for the interaction of the chimeric protein with muCaMLD and resulted in its susceptibility to calpain proteolysis. Moreover, the muCaMLD of calpain acts as a competitive inhibitor of calpain-dependent IkappaBalpha degradation. Our data demonstrate that the IkappaBalpha-PEST sequence acts as a modular domain to promote the physical association with and subsequent degradation by mu-calpain and suggest a functional role for PEST sequences in other proteins as potential calpain-targeting units. PMID- 10521482 TI - Leucine zipper-mediated homodimerization of the adaptor protein c-Cbl. A role in c-Cbl's tyrosine phosphorylation and its association with epidermal growth factor receptor. AB - The 120-kDa proto-oncogenic protein c-Cbl is a multidomain adaptor protein that is phosphorylated in response to the stimulation of a broad range of cell surface receptors and participates in the assembly of signaling complexes that are formed as a result of the activation of various signal transduction pathways. Several structural features of c-Cbl, including the phosphotyrosine-binding domain, proline-rich domain, and motifs containing phosphotyrosine and phosphoserine residues, mediate the association of c-Cbl with other components of these complexes. In addition to those domains that have been demonstrated to play a role in the binding of c-Cbl to other signaling molecules, c-Cbl also contains a RING finger motif and a putative leucine zipper. In this study, we demonstrate that the previously identified putative leucine zipper mediates the formation of Cbl homodimers. Using the yeast two-hybrid system, we show that deletion of the leucine zipper domain is sufficient to abolish Cbl homodimerization, while Cbl mutants carrying extensive N-terminal truncations retain the ability to dimerize with the full-length Cbl. The requirement of the leucine zipper for the homodimerization of Cbl was confirmed by in vitro binding assays, using deletion variants of the C-terminal half of Cbl with and without the leucine zipper domain, and in cells using Myc and green fluorescent protein (GFP) N-terminal tagged Cbl variants. In cells, the deletion of the leucine zipper caused a decrease in both the tyrosine phosphorylation of Cbl and its association with the epidermal growth factor receptor following stimulation with epidermal growth factor, thus demonstrating a role for the leucine zipper in c-Cbl's signaling functions. Thus, the leucine zipper domain enables c-Cbl to homodimerize, and homodimerization influences Cbl's signaling function, modulating the activity of Cbl itself and/or affecting Cbl's associations with other signaling proteins in the cell. PMID- 10521481 TI - Engagement of tumor necrosis factor (TNF) receptor 1 leads to ATF-2- and p38 mitogen-activated protein kinase-dependent TNF-alpha gene expression. AB - Engagement of the tumor necrosis factor-alpha (TNF-alpha) receptors by the TNF alpha ligand results in the rapid induction of TNF-alpha gene expression. The study presented here shows that autoregulation of TNF-alpha gene transcription by selective signaling through tumor necrosis factor receptor 1 (TNFR1) requires p38 mitogen-activated protein (MAP) kinase activity and the binding of the transcription factors ATF-2 and Jun to the TNF-alpha cAMP-response element (CRE) promoter element. Consistent with these findings, TNFR1 engagement results in increased p38 MAP kinase activity and p38-dependent phosphorylation of ATF-2. Furthermore, overexpression of MADD (MAP kinase-activating death domain protein), an adapter protein that binds to the death domain of TNFR1 and activates MAP kinase cascades, results in CRE-dependent induction of TNF-alpha gene expression. Thus, the TNF-alpha CRE site is the target of TNFR1 stimulation and mediates the autoregulation of TNF-alpha gene transcription. PMID- 10521483 TI - Identification of Tek/Tie2 binding partners. Binding to a multifunctional docking site mediates cell survival and migration. AB - The Tek/Tie2 receptor tyrosine kinase plays a pivotal role in vascular and hematopoietic development. To study the signal transduction pathways that are mediated by this receptor, we have used the yeast two-hybrid system to identify signaling molecules that associate with the phosphorylated Tek receptor. Using this approach, we demonstrate that five molecules, Grb2, Grb7, Grb14, Shp2, and the p85 subunit of phosphatidylinositol 3-kinase can interact with Tek in a phosphotyrosine-dependent manner through their SH2 domains. Mapping of the binding sites of these molecules on Tek reveals the presence of a multisubstrate docking site in the carboxyl tail of Tek (Tyr(1100)). Mutation of this site abrogates binding of Grb2 and Grb7 to Tek in vivo, and this site is required for tyrosine phosphorylation of Grb7 and p85 in vivo. Furthermore, stimulation of Tek expressing cells with Angiopoietin-1 results in phosphorylation of both Tek and p85 and in activation of endothelial cell migration and survival pathways that are dependent in part on phosphatidylinositol 3-kinase. Taken together, these results demonstrate that Angiopoietin-1-induced signaling from the Tek receptor is mediated by a multifunctional docking site that is responsible for activation of both cell migration and cell survival pathways. PMID- 10521484 TI - Mutations in the heparin binding domain of fibronectin in cooperation with the V region induce decreases in pp125(FAK) levels plus proteoglycan-mediated apoptosis via caspases. AB - Intact fibronectin (FN) protects cells from apoptosis. When FN is fragmented, specific domains induce proteinase expression in fibroblasts. However, it is not known whether specific domains of FN can also regulate apoptosis. We exposed fibroblasts to four recombinant FN fragments and then assayed for apoptosis using criteria of cellular shape change, condensed nuclear morphology, and DNA fragmentation. The fragments extended from the RGD-containing repeat III10 to III15; they included (V(+)) or excluded (V(-)) the alternatively spliced V region and contained either a mutated (H(-)) or an unmutated (H(+)) heparin binding domain. Only the V(+)H(-) fragment triggered decreases in pp125(FAK) levels and apoptosis, which was rescued by intact FN and inhibitors of caspase-1 and caspase 3. This apoptotic mechanism was mediated by a chondroitin sulfate proteoglycan, since treating cells with chondroitin sulfate or chondroitinase reversed the apoptotic cell shape changes. The alpha4 integrin receptor may also be involved, since using a blocking antibody to alpha4 alone induced apoptotic cell shape changes, whereas co-treatment with this antibody plus V(+)H(+) reversed these effects. These results demonstrate that the V and heparin binding domains of FN modulate pp125(FAK) levels and regulate apoptosis through a chondroitin sulfate proteoglycan- and possibly alpha4 integrin-mediated pathway, which triggers a caspase cascade. PMID- 10521485 TI - nArgBP2, a novel neural member of ponsin/ArgBP2/vinexin family that interacts with synapse-associated protein 90/postsynaptic density-95-associated protein (SAPAP). AB - Postsynaptic density (PSD)-95/synapse-associated protein (SAP) 90 and synaptic scaffolding molecule (S-SCAM) are synaptic membrane-associated guanylate kinases. Both the proteins interact with SAP90/PSD-95-associated protein (SAPAP) (also called guanylate kinase-associated protein/Dlg-associated protein). SAPAP is a protein highly enriched in the PSD fraction and may link PSD-95/SAP90 and S-SCAM to Triton X-100-insoluble structures. We found here a novel SAPAP-interacting protein, which was specifically expressed in neural tissue and was present in the postsynaptic density fraction in brain. This protein had a sorbin homology domain in the N terminus, a zinc finger motif in the middle region, and three src homology (SH) 3 domains in the C terminus and was homologous to the ponsin/ArgBP2/vinexin family proteins. We named this protein nArgBP2 because it was the most homologous to ArgBP2. nArgBP2 is a neural member of a growing family of SH3-containing proteins. nArgBP2 bound to the proline-rich region of SAPAP via its third SH3 domain and was coimmunoprecipitated with SAPAP from the extract of rat brain. Furthermore, nArgBP2 was colocalized with SAPAP at synapses in cerebellum. nArgBP2 bound to not only SAPAP but also vinculin and l-afadin, known to bind to ponsin and vinexin. nArgBP2 may be implicated in the protein network around SAPAP in the PSD. PMID- 10521487 TI - Heart 6-phosphofructo-2-kinase activation by insulin results from Ser-466 and Ser 483 phosphorylation and requires 3-phosphoinositide-dependent kinase-1, but not protein kinase B. AB - Previous studies have shown that (i) the insulin-induced activation of heart 6 phosphofructo-2-kinase (PFK-2) is wortmannin-sensitive, but is insensitive to rapamycin, suggesting the involvement of phosphatidylinositol 3-kinase; and (ii) protein kinase B (PKB) activates PFK-2 in vitro by phosphorylating Ser-466 and Ser-483. In this work, we have studied the effects of phosphorylation of these residues on PFK-2 activity by replacing each or both residues with glutamate. Mutation of Ser-466 increased the V(max) of PFK-2, whereas mutation of Ser-483 decreased citrate inhibition. Mutation of both residues was required to decrease the K(m) for fructose 6-phosphate. We also studied the insulin-induced activation of heart PFK-2 in transfection experiments performed in human embryonic kidney 293 cells. Insulin activated transfected PFK-2 by phosphorylating Ser-466 and Ser 483. Kinase-dead (KD) PKB and KD 3-phosphoinositide-dependent kinase-1 (PDK-1) cotransfectants acted as dominant negatives because both prevented the insulin induced activation of PKB as well as the inactivation of glycogen-synthase kinase 3, an established substrate of PKB. However, the insulin-induced activation of PFK-2 was prevented only by KD PDK-1, but not by KD PKB. These results indicate that the insulin-induced activation of heart PFK-2 is mediated by a PDK-1 activated protein kinase other than PKB. PMID- 10521486 TI - Interleukin-6 increases rat metalloproteinase-13 gene expression through stimulation of activator protein 1 transcription factor in cultured fibroblasts. AB - The role of IL-6 in collagen production and tissue remodeling is controversial. In Rat-1 fibroblasts, we measured the effect of IL-6 on matrix metalloproteinase 13 (MMP-13), c-jun, junB, and c-fos gene expression, binding of activator protein 1 (AP1) to DNA, amount of AP1 proteins, immunoreactive MMP-13 and TIMP-1 proteins, and Jun N-terminal kinase activity. We show that IL-6 increased MMP-13 mRNA and MMP-13 protein. These effects were exerted by acting on the AP1-binding site of the MMP-13 promoter, as shown by transfecting cells with reporter plasmids containing mutations in this element. Mobility shift assays demonstrated that IL-6 induced the DNA binding activity of AP1. This effect was accompanied by a marked increase in c-Jun, JunB, and c-Fos mRNA, as well as in c-Jun protein and its phosphorylated form. The latter is not due to increased Jun N-terminal kinase activity but to a decreased serine/threonine phosphatase activity. We conclude that IL-6 increases interstitial MMP-13 gene expression at the promoter level. This effect seems to be mediated by the induction of c-jun, junB, and c-fos gene expression, by the binding of AP1 to DNA, by increasing phosphorylated c-Jun, and by the inhibition of serine/threonine phosphatase activity. These effects of IL-6 might contribute to remodeling connective tissue. PMID- 10521488 TI - Fibroblast growth factor receptor 3 gene transcription is suppressed by cyclic adenosine 3',5'-monophosphate. Identification of a chondrocytic regulatory element. AB - Signaling through fibroblast growth factor receptors (FGFRs) is critical for the development and patterning of the vertebrate skeleton. Gain-of-function alleles of fgfr2 and fgfr3 have been linked to several dominant skeletal disorders in humans, while null mutations in fgfr3 result in the overgrowth of long bones in a mouse model system. Interestingly, the expression pattern of fgfr3 in growth plate chondrocytes overlaps that of the parathyroid hormone (PTH)-related peptide (PTHrP) receptor, a signaling molecule that also regulates endochondral ossification. The coincident expression of these two receptors suggests that their signaling pathways may also interact. To gain insight into the regulatory mechanism(s) that govern the expression of the fgfr3 gene in chondrocytes, we have identified a cell-specific transcriptional regulatory element (CSRh) by measuring the activity of various promoter fragments in FGFR3-expressing (CFK2) and nonexpressing (RCJ) chondrocyte-like cell lines. Furthermore, we demonstrate that activation of PTH/PTHrP receptors, either by stimulation with PTH or through the introduction of activating mutations, represses CSRh-mediated transcriptional activity. Finally, the transcriptional repression of the CSRh element was mimicked by treatment with forskolin, 8-bromo-cAMP, and 3-isobutyl-1 methylxanthine or by overexpression of the catalytic subunit of protein kinase A. Together, these data suggest that protein kinase A activity is a critical factor that regulates fgfr3 gene expression in the proliferative or prehypertrophic compartment of the epiphyseal growth plate. Furthermore, these results provide a possible link between PTHrP signaling and fgfr3 gene expression during the process of endochondral ossification. PMID- 10521489 TI - Sp1 and NF-Y are necessary and sufficient for growth-dependent regulation of the hamster thymidine kinase promoter. AB - Thymidine kinase (TK) genes from different species are growth- and cell cycle regulated in a very similar manner; still, the promoter regions of these genes show little homology to each other. It was previously shown that the murine TK gene is growth-regulated by Sp1 and E2F. Here we have characterized cis regulatory elements in the hamster promoter that are essential and sufficient to confer efficient and serum-responsive expression. The TK promoter was isolated from baby hamster kidney cells. DNase I protection experiments revealed a protected region from positions -24 to -99 relative to the transcription start site. Within this region, binding sites for the transcription factor Sp1 and a CCAAT box, which interacts with the transcription factor NF-Y, were identified. An E2F-like sequence was found not to bind protein, and its removal did not affect promoter activity. This was supported by the observation that cotransfection of a hamster TK reporter gene construct with E2F-1 does not lead to transactivation of the promoter. A 122-base pair region that contains a single Sp1 site, a CCAAT box, and a TATA element was found to be sufficient for serum responsive expression of a reporter gene. Mutations that inactivate any one of these three elements caused a strong reduction or a loss of promoter activity. PMID- 10521490 TI - Conformational change in the pheromone-binding protein from Bombyx mori induced by pH and by interaction with membranes. AB - The pheromone-binding protein (PBP) from Bombyx mori was expressed in Escherichia coli periplasm. It specifically bound radiolabeled bombykol, the natural pheromone for this species. It appeared as a single band both in native and SDS polyacrylamide gel electrophoresis and was also homogeneous in most chromatographic systems. However, in ion-exchange chromatography, multiple forms sometimes appeared. Attempts to separate them revealed that they could be converted into one another. Analysis of the protein by circular dichroism and fluorescence spectroscopy demonstrated that its tertiary structure was sensitive to pH changes and that a dramatic conformational transition occurred between pH 6.0 and 5.0. This high sensitivity to pH contrasted markedly with its thermal stability and resistance to denaturation by urea. There was also no significant change in CD spectra in the presence of the pheromone. The native protein isolated from male antennae displayed the same changes in its spectroscopic properties as the recombinant material, demonstrating that this phenomenon is not an artifact arising from the expression system. This conformational transition was reproduced by interaction of the protein with anionic (but not neutral) phospholipid vesicles. Unfolding of the PBP structure triggered by membranes suggests a plausible mechanism for ligand release upon interaction of the PBP pheromone complex with the surface of olfactory neurons. This pH-linked structural flexibility also explains the heterogeneity reported previously for B. mori PBP and other members of this class of proteins. PMID- 10521491 TI - Distinct domains of mouse dishevelled are responsible for the c-Jun N-terminal kinase/stress-activated protein kinase activation and the axis formation in vertebrates. AB - Recent studies have shown that Drosophila Dishevelled (Dsh), an essential component of the wingless signal transduction, is also involved in planar polarity signaling through the c-Jun N-terminal kinase (JNK)/stress-activated protein kinase (SAPK) pathway in Drosophila. Here, we show that expression of a mouse homolog of Dsh (mDvl-1) in NIH3T3 cells activates JNK/SAPK, and its activator MKK7. A C-terminal half of mDvl-1 which contains the DEP domain was sufficient for the activation of JNK/SAPK, whereas an N-terminal half of mDvl-1 as well as the DEP domain is required for stimulation of the TCF/LEF-1-dependent transcriptional activation, a beta-catenin-dependent process. A single amino acid substitution (Met for Lys) within the DEP domain (mDvl-1 (KM)) abolished the JNK/SAPK-activating activity of mDvl-1, but did not affect the activity to activate the LEF-1-dependent transcription. Ectopic expression of mDvl-1 (KM) or an N-terminal half of mDvl-1, but not the C-terminal, was able to induce secondary axis in Xenopus embryos. Because the secondary axis formation is dependent on the Wnt/beta-catenin signaling pathway, these results suggest that distinct domains of mDvl-1 are responsible for the two downstream signaling pathways, the beta-catenin pathway and the JNK/SAPK pathway in vertebrates. PMID- 10521492 TI - The ubiquitin-conjugating enzymes UbcH7 and UbcH8 interact with RING finger/IBR motif-containing domains of HHARI and H7-AP1. AB - Ubiquitinylation of proteins appears to be mediated by the specific interplay between ubiquitin-conjugating enzymes (E2s) and ubiquitin-protein ligases (E3s). However, cognate E3s and/or substrate proteins have been identified for only a few E2s. To identify proteins that can interact with the human E2 UbcH7, a yeast two-hybrid screen was performed. Two proteins were identified and termed human homologue of Drosophila ariadne (HHARI) and UbcH7-associated protein (H7-AP1). Both proteins, which are widely expressed, are characterized by the presence of RING finger and in between RING fingers (IBR) domains. No other overt structural similarity was observed between the two proteins. In vitro binding studies revealed that an N-terminal RING finger motif (HHARI) and the IBR domain (HHARI and H7-AP1) are involved in the interaction of these proteins with UbcH7. Furthermore, binding of these two proteins to UbcH7 is specific insofar that both HHARI and H7-AP1 can bind to the closely related E2, UbcH8, but not to the unrelated E2s UbcH5 and UbcH1. Although it is not clear at present whether HHARI and H7-AP1 serve, for instance, as substrates for UbcH7 or represent proteins with E3 activity, our data suggests that a subset of RING finger/IBR proteins are functionally linked to the ubiquitin/proteasome pathway. PMID- 10521493 TI - Estrogen inducibility of c-Ha-ras transcription in breast cancer cells. Identification of functional estrogen-responsive transcriptional regulatory elements in exon 1/intron 1 of the c-Ha-ras gene. AB - Although mutation of ras gene is rare in human breast cancer, overexpression of normal c-Ha-ras gene is frequently observed. Using a mouse mammary metastasis model consisting of genetically related mammary tumor sublines with variant metastatic potential, we have previously (i) demonstrated a direct correlation between c-Ha-ras mRNA and protein levels and metastatic potential and (ii) identified a novel hormone-responsive transcriptional regulatory element in intron 1 of the mouse c-Ha-ras gene that contains the consensus half-site of a glucocorticoid response element and flanking consensus half-sites for estrogen response element. Here, we have examined the functionality of intron 1 sequence in context of upstream sequences by using transient transfection assays with plasmids expressing chloramphenicol acetyltransferase. Intron 1 sequence and sequences similar to intron 1 element located in exon 1 function as transcriptional regulatory elements that confer hormonal inducibility to chloramphenicol acetyltransferase gene expression both independently and in context of 5'-flanking sequences. Measurement of c-Ha-ras transcription rates and protein expression by nuclear run-on and metabolic labeling assays showed a 5-12 fold enhancement, respectively, following treatment with 17beta-estradiol that was blunted by ICI 182,780 in the nonmetastatic variant. In contrast, constitutive overexpression of c-Ha-ras transcripts and protein in the metastatic subline was unaffected by estrogen and ICI 182,780. Gel shift assays demonstrated specific interaction of c-Ha-ras exon 1 sequence with nuclear proteins of human breast cancer MCF-7 cells with formation of two complexes, one of which contains estrogen receptor. Our data demonstrate a direct (i) interaction of c-Ha-ras sequence with estrogen receptor and (ii) stimulatory effect of estrogen on c-Ha ras gene transcription and suggest that alteration in transcriptional regulation of c-Ha-ras gene by estrogen may play an important role in progression of breast cancer. PMID- 10521494 TI - The lipid products of phosphoinositide 3-kinase contribute to regulation of cholangiocyte ATP and chloride transport. AB - ATP stimulates Cl(-) secretion and bile formation by activation of purinergic receptors in the apical membrane of cholangiocytes. The purpose of these studies was to determine the cellular origin of biliary ATP and to assess the regulatory pathways involved in its release. In Mz-Cha-1 human cholangiocarcinoma cells, increases in cell volume were followed by increases in phophoinositide (PI) 3 kinase activity, ATP release, and membrane Cl(-) permeability. PI 3-kinase signaling appears to play a regulatory role because ATP release was inhibited by wortmannin or LY294002 and because volume-sensitive current activation was inhibited by intracellular dialysis with antibodies to the 110 kDa-subunit of PI 3-kinase. Similarly, in intact normal rat cholangiocyte monolayers, increases in cell volume stimulated luminal Cl(-) secretion through a wortmannin-sensitive pathway. To assess the role of PI 3-kinase more directly, cells were dialyzed with the synthetic lipid products of PI 3-kinase. Intracellular delivery of phosphatidylinositol 3, 4-bisphosphate, and phosphatidylinositol 3,4,5 trisphosphate activated Cl(-) currents analogous to those observed following cell swelling. Taken together, these findings indicate that volume-sensitive activation of PI 3-kinase and the generation of lipid messengers modulate cholangiocyte ATP release, Cl(-) secretion, and, hence, bile formation. PMID- 10521495 TI - Isolation and characterization of photoautotrophic mutants of Chlamydomonas reinhardtii deficient in state transition. AB - In photosynthetic cells of higher plants and algae, the distribution of light energy between photosystem I and photosystem II is controlled by light quality through a process called state transition. It involves a reorganization of the light-harvesting complex of photosystem II (LHCII) within the thylakoid membrane whereby light energy captured preferentially by photosystem II is redirected toward photosystem I or vice versa. State transition is correlated with the reversible phosphorylation of several LHCII proteins and requires the presence of functional cytochrome b(6)f complex. Most factors controlling state transition are still not identified. Here we describe the isolation of photoautotrophic mutants of the unicellular alga Chlamydomonas reinhardtii, which are deficient in state transition. Mutant stt7 is unable to undergo state transition and remains blocked in state I as assayed by fluorescence and photoacoustic measurements. Immunocytochemical studies indicate that the distribution of LHCII and of the cytochrome b(6)f complex between appressed and nonappressed thylakoid membranes does not change significantly during state transition in stt7, in contrast to the wild type. This mutant displays the same deficiency in LHCII phosphorylation as observed for mutants deficient in cytochrome b(6)f complex that are known to be unable to undergo state transition. The stt7 mutant grows photoautotrophically, although at a slower rate than wild type, and does not appear to be more sensitive to photoinactivation than the wild-type strain. Mutant stt3-4b is partially deficient in state transition but is still able to phosphorylate LHCII. Potential factors affected in these mutant strains and the function of state transition in C. reinhardtii are discussed. PMID- 10521496 TI - Characterization of the LolA-LolB system as the general lipoprotein localization mechanism of Escherichia coli. AB - The major outer membrane lipoprotein (Lpp) of Escherichia coli requires LolA for its release from the cytoplasmic membrane, and LolB for its localization to the outer membrane. We examined the significance of the LolA-LolB system as to the outer membrane localization of other lipoproteins. All lipoproteins possessing an outer membrane-directed signal at the N-terminal second position were efficiently released from the inner membrane in the presence of LolA. Some lipoproteins were released in the absence of externally added LolA, albeit at a slower rate and to a lesser extent. This LolA-independent release was also strictly dependent on the outer membrane sorting signal. A lipoprotein-LolA complex was formed when the release took place in the presence of LolA, whereas lipoproteins released in the absence of LolA existed as heterogeneous complexes, suggesting that the release and the formation of a complex with LolA are distinct events. The release of LolB, an outer membrane lipoprotein functioning as the receptor for a lipoprotein LolA complex, occurred with a trace amount of LolA, and therefore was extremely efficient. The LolA-dependent release of lipoproteins was found to be crucial for the specific incorporation of lipoproteins into the outer membrane, whereas lipoproteins released in the absence of LolA were nonspecifically and inefficiently incorporated into the membrane. The outer membrane incorporation of lipoproteins including LolB per se was dependent on LolB in the outer membrane. From these results, we conclude that lipoproteins in E. coli generally utilize the LolA-LolB system for efficient release from the inner membrane and specific localization to the outer membrane. PMID- 10521497 TI - Role of factor VIII C2 domain in factor VIII binding to factor Xa. AB - Factor VIII (FVIII) is activated by proteolytic cleavages with thrombin and factor Xa (FXa) in the intrinsic blood coagulation pathway. The anti-C2 monoclonal antibody ESH8, which recognizes residues 2248-2285 and does not inhibit FVIII binding to von Willebrand factor or phospholipid, inhibited FVIII activation by FXa in a clotting assay. Furthermore, analysis by SDS polyacrylamide gel electrophoresis showed that ESH8 inhibited FXa cleavage in the presence or absence of phospholipid. The light chain (LCh) fragments (both 80 and 72 kDa) and the recombinant C2 domain dose-dependently bound to immobilized anhydro-FXa, a catalytically inactive derivative of FXa in which dehydroalanine replaces the active-site serine. The affinity (K(d)) values for the 80- and 72 kDa LCh fragments and the C2 domain were 55, 51, and 560 nM, respectively. The heavy chain of FVIII did not bind to anhydro-FXa. Similarly, competitive assays using overlapping synthetic peptides corresponding to ESH8 epitopes (residues 2248-2285) demonstrated that a peptide designated EP-2 (residues 2253-2270; TSMYVKEFLISSSQDGHQ) inhibited the binding of the C2 domain or the 72-kDa LCh to anhydro-FXa by more than 95 and 84%, respectively. Our results provide the first evidence for a direct role of the C2 domain in the association between FVIII and FXa. PMID- 10521498 TI - Transforming growth factor beta1 enhances platelet aggregation through a non transcriptional effect on the fibrinogen receptor. AB - Upon activation, platelets store and release large amounts of the peptide transforming growth factor beta1 (TGFbeta1). The released TGFbeta1 can then act on nearby vascular cells to mediate subsequent vessel repair. In addition, TGFbeta1 may circulate to bone marrow and regulate megakaryocyte activity. It is not known what effect, if any, TGFbeta1 has on platelets. Adult TGFbeta1 deficient mice exhibit thrombocythemia and a mild bleeding disorder that is shown to result from faulty platelet aggregation. TGFbeta1-deficient platelets are shown to contain functional receptors, and preincubation with recombinant TGFbeta1 improves aggregation, demonstrating that TGFbeta1 plays an active role in platelet aggregation. TGFbeta1-deficient platelets fail to retain bound fibrinogen in response to aggregation agonists, but they possess normal levels of the alpha(IIb)/beta(3) fibrinogen receptor. Signaling from agonist receptors is normal because the platelets change shape, produce thromboxane A(2), and present P-selectin in response to stimulation. Consequently, activation and maintenance of alpha(IIb)/beta(3) into a fibrinogen-binding conformation is impaired in the absence of TGFbeta1. 4-Phorbol 12-myristate 13-acetate treatment and protein kinase C activity measurements suggest a defect downstream of protein kinase C in its activation cascade. Because platelets lack nuclei, these data demonstrate for the first time a non-transcriptionally mediated TGFbeta1 signaling pathway that enhances the activation and maintenance of integrin function. PMID- 10521499 TI - Activation of the beta(2)-adrenergic receptor-Galpha(s) complex leads to rapid depalmitoylation and inhibition of repalmitoylation of both the receptor and Galpha(s). AB - Palmitoylation is unique among lipid modifications in that it is reversible. In recent years, dynamic palmitoylation of G protein alpha subunits and of their cognate receptors has attracted considerable attention. However, very little is known concerning the acylation/deacylation cycle of the proteins in relation to their activity status. In particular, the relative contribution of the activation and desensitization of the signaling unit to the regulation of the receptors and G proteins palmitoylation state is unknown. To address this issue, we took advantage of the fact that a fusion protein composed of the stimulatory alpha subunit of trimeric G protein (Galpha(s)) covalently attached to the beta(2) adrenergic receptor (beta(2)AR) as a carboxyl-terminal extension (beta(2)AR Galpha(s)) can be stimulated by agonists but does not undergo rapid inactivation, desensitization, or internalization. When expressed in Sf9 cells, both the receptor and the Galpha(s) moieties of the fusion protein were found to be palmitoylated via thioester linkage. Stimulation with the beta-adrenergic agonist isoproterenol led to a rapid depalmitoylation of both the beta(2)AR and Galpha(s) and inhibited repalmitoylation. The extent of depalmitoylation induced by a series of agonists was correlated (0.99) with their intrinsic efficacy to stimulate the adenylyl cyclase activity. However, forskolin-stimulated cAMP production did not affect the palmitoylation state of beta(2)AR-Galpha(s), indicating that the agonist-promoted depalmitoylation is linked to conformational changes and not to second messenger generation. Given that, upon activation, the fusion protein mimics the activated receptor-G protein complex but cannot undergo desensitization, the data demonstrate that early steps in the activation process lead to the depalmitoylation of both receptor and G protein and that repalmitoylation requires later events that cannot be accommodated by the activated fusion protein. PMID- 10521500 TI - Increases in acidic phospholipid contents specifically restore protein translocation in a cold-sensitive secA or secG null mutant. AB - Both the secAcsR11 and DeltasecG::kan mutations cause cold-sensitive growth, although the growth defect due to the latter mutation occurs in a strain-specific manner. Overexpression of pgsA encoding phosphatidylglycerophosphate synthase suppresses the growth defects of the two mutants. We investigated the mechanism underlying the pgsA-dependent suppression of the two mutations using purified mutant SecA and inverted membrane vesicles (IMVs) prepared from pgsA overexpressing cells. The acidic phospholipid content increased by about 10% upon pgsA overexpression. This increase resulted in the stimulation of proOmpA translocation only when mutant SecA or SecG-depleted IMVs were used. The translocation-coupled ATPase activity of SecA was significantly defective with the mutant SecA or SecG-depleted IMVs, but it recovered to a near normal level when the acidic phospholipid level was increased. The stimulation of ATPase activity was observed only at low temperature. The steady-state level of membrane inserted SecA was low with the mutant SecA or SecG-depleted IMVs, and it decreased further upon the increase in the acidic phospholipid content. However, the level of SecA insertion markedly increased upon the inhibition of SecA deinsertion by the addition of beta,gamma-imido adenosine 5'-triphosphate (AMP PNP), especially with IMVs containing increased levels of acidic phospholipids. These results indicate that the increase in the level of acidic phospholipids stimulates the SecA cycle in the two mutants by facilitating both the insertion and deinsertion of SecA. PMID- 10521501 TI - Arsenite exposure of cultured airway epithelial cells activates kappaB-dependent interleukin-8 gene expression in the absence of nuclear factor-kappaB nuclear translocation. AB - Airway epithelial cells respond to certain environmental stresses by mounting a proinflammatory response, which is characterized by enhanced synthesis and release of the neutrophil chemotactic and activating factor interleukin-8 (IL-8). IL-8 expression is regulated at the transcriptional level in part by the transcription factor nuclear factor (NF)-kappaB. We compared intracellular signaling mediating IL-8 gene expression in bronchial epithelial cells cultured in vitro and exposed to two inducers of cellular stress, sodium arsenite (As(III)), and vanadyl sulfate (V(IV)). Unstimulated bronchial epithelial cells expressed IL-8, and exposure to both metal compounds significantly enhanced IL-8 expression. Overexpression of a dominant negative inhibitor of NF-kappaB depressed both basal and metal-induced IL-8 expression. Low levels of nuclear NF kappaB were constitutively present in unstimulated cultures. These levels were augmented by exposure to V(IV), but not As(III). Accordingly, V(IV) induced IkappaBalpha breakdown and NF-kappaB nuclear translocation, whereas As(III) did not. However, both As(III) and V(IV) enhanced kappaB-dependent transcription. In addition, As(III) activation of an IL-8 promoter-reporter construct was partially kappaB-dependent. These data suggested that As(III) enhanced IL-8 gene transcription independently of IkappaB breakdown and nuclear translocation of NF kappaB in part by enhancing transcription mediated by low levels of constitutive nuclear NF-kappaB. PMID- 10521502 TI - The role of the Escherichia coli mug protein in the removal of uracil and 3,N(4) ethenocytosine from DNA. AB - The human thymine-DNA glycosylase has a sequence homolog in Escherichia coli that is described to excise uracils from U.G mismatches (Gallinari, P., and Jiricny, J. (1996) Nature 383, 735-738) and is named mismatched uracil glycosylase (Mug). It has also been described to remove 3,N(4)-ethenocytosine (epsilonC) from epsilonC.G mismatches (Saparbaev, M., and Laval, J. (1998) Proc. Natl. Acad. Sci. U. S. A. 95, 8508-8513). We used a mug mutant to clarify the role of this protein in DNA repair and mutation avoidance. We find that inactivation of mug has no effect on C to T or 5-methylcytosine to T mutations in E. coli and that this contrasts with the effect of ung defect on C to T mutations and of vsr defect on 5-methylcytosine to T mutations. Even under conditions where it is overproduced in cells, Mug has little effect on the frequency of C to T mutations. Because uracil-DNA glycosylase (Ung) and Vsr are known to repair U.G and T.G mismatches, respectively, we conclude that Mug does not repair U.G or T.G mismatches in vivo. A defect in mug also has little effect on forward mutations, suggesting that Mug does not play a role in avoiding mutations due to endogenous damage to DNA in growing E. coli. Cell-free extracts from mug(+) ung cells show very little ability to remove uracil from DNA, but can excise epsilonC. The latter activity is missing in extracts from mug cells, suggesting that Mug may be the only enzyme in E. coli that can remove this mutagenic adduct. Thus, the principal role of Mug in E. coli may be to help repair damage to DNA caused by exogenous chemical agents such as chloroacetaldehyde. PMID- 10521503 TI - Asparagine-linked oligosaccharides protect Lamp-1 and Lamp-2 from intracellular proteolysis. AB - Lysosomes contain several integral membrane proteins (termed Lamps and Limps) that are extensively glycosylated with asparagine-linked oligosaccharides. It has been postulated that these glycans protect the underlying polypeptides from the proteolytic environment of the lysosome. Previous attempts to test this hypothesis have been inconclusive because they utilized approaches that prevent initial glycosylation and thereby impair protein folding. We have used endoglycosidase H to remove the Asn-linked glycans from fully folded lysosomal membrane proteins in living cells. Deglycosylation of Lamp-1 and Lamp-2 resulted in their rapid degradation, whereas Limp-2 was relatively stable in the lysosome in the absence of high mannose Asn-linked oligosaccharides. Depletion of Lamp-1 and Lamp-2 had no measurable effect on endosomal/lysosomal pH, osmotic stability, or density, and cell viability was maintained. Transport of endocytosed material to dense lysosomes was delayed in endoglycosidase H treated cells, but the rate of degradation of internalized bovine serum albumin was unchanged. These data provide direct evidence that Asn-linked oligosaccharides protect a subset of lysosomal membrane proteins from proteolytic digestion in intact cells. PMID- 10521504 TI - Vascular endothelial growth factor effect on endothelial cell proliferation, migration, and platelet-activating factor synthesis is Flk-1-dependent. AB - Vascular endothelial growth factor (VEGF) is a potent inducer of endothelial cell (EC) proliferation and migration in vitro as well as inflammation in vivo. We showed recently that VEGF effect on vascular permeability was dependent on the synthesis of platelet-activating factor (PAF) by EC. Consequently, we sought to evaluate by antisense knockdown of gene expression the contribution of VEGF receptors (Flt-1 and Flk-1) on these events. VEGF (10(-11) to 10(-8) M) elicited a dose-dependent increase of bovine aortic EC proliferation, migration, and PAF synthesis by up to 2.05-, 1.31- and 35.9-fold above basal levels, respectively. A treatment with two modified antisense oligomers (1-5 x 10(-7) M) directed against Flk-1 mRNA blocked by 100, 91, and 85% the proliferation, migration, and PAF synthesis mediated by VEGF, respectively. A treatment with two antisense oligomers directed against Flt-1 mRNA failed to modulate these activities. The use of placenta growth factor (up to 10(-8) M), an Flt-1-specific agonist, induced only a slight increase (0.6-fold) of PAF synthesis. These data illustrate the crucial role of Flk-1 in EC stimulation by VEGF. The capacity to inhibit the protein synthesis of Flt-1 and Flk-1 by antisense oligonucleotides provides a new approach to block VEGF pathological effects in vivo. PMID- 10521505 TI - Serine phosphorylation and negative regulation of Stat3 by JNK. AB - STATs are activated by various cytokines and growth factors via tyrosine phosphorylation, which leads to sequential dimer formation, nuclear translocation, binding to specific DNA sequences, and regulation of gene expression. Recently, serine phosphorylation of Stat3 on Ser-727 by ERK has been identified in response to epidermal growth factor (EGF). Here, we report that Ser 727 phosphorylation of Stat3 can also be induced by JNK and activated either by stress or by its upstream kinase and that various stress treatments induce serine phosphorylation of Stat3 in the absence of tyrosine phosphorylation. Inhibitors of ERK and p38 did not inhibit UV-induced Stat3 serine phosphorylation, suggesting that neither of them is involved. We further demonstrate that JNK1, activated by its upstream kinase MKK7, negatively regulated the tyrosine phosphorylation and DNA binding and transcriptional activities of Stat3 stimulated by EGF. Correspondingly, pretreatment of cells with UV reduced the EGF stimulated tyrosine phosphorylation and phosphotyrosine-dependent activities of Stat3. The inhibitory effect was not observed for Stat1. Our results suggest that Stat3 is a target of JNK that may regulate Stat3 activity via both Ser-727 phosphorylation-dependent and -independent mechanisms. PMID- 10521506 TI - Platelet-derived growth factor stimulation of monocyte chemoattractant protein-1 gene expression is mediated by transient activation of the phosphoinositide 3 kinase signal transduction pathway. AB - Platelet-derived growth factor (PDGF) stimulates transcription of an immediate early gene set in Balb/c 3T3 cells. One cohort of these genes, typified by c-fos, is induced within minutes following activation of PDGF receptors. A second cohort responds to PDGF only after a significant time delay, although induction is still a primary response to receptor activation as shown by "superinduction" in the presence of the protein synthesis inhibitor cycloheximide. PDGF-receptor activated signaling pathways for the "slow" immediate-early genes are poorly resolved. Using gain-of-function mutations together with small molecule inhibitors of kinase activity, we show that activation of PI 3-kinase is both necessary and sufficient for the induction of the prototype slow immediate-early gene, monocyte chemoattractant-1 (MCP-1). Following activation of PDGF receptors, MCP-1 mRNA does not begin to accumulate for at least 90 min. However, only a brief (10 min) interval of PI 3-kinase activity is required to trigger this delayed response. The serine/threonine protein kinase, Akt/PKB, likely functions as a downstream affector of PI 3-kinase for this induction. PMID- 10521507 TI - Characterization of a mammalian gene related to the yeast CCR4 general transcription factor and revealed by transposon insertion. AB - Murine intracisternal A-particles (IAPs) are reiterated retrovirus-like transposable elements that can act as insertional mutagens. Accordingly, we previously identified a chimeric transcript initiated at an IAP promoter and extending through a 3'-located open reading frame with significant similarity to the C-terminal domain of the yeast CCR4 general transcription factor. In this report, we characterize the corresponding murine gene, mCCR4, and its human homologue, thus providing the first description of CCR4-like factors in mammals. cDNA cloning revealed two mCCR4 mRNAs of 2.7 and 3.1 kilobases, differing by their transcription start sites within the native mCCR4 gene promoter, and encoding a putative 430-amino acid protein. The mCCR4 gene contains three exons and two introns spanning almost 27 kilobases. The IAP insertion, detected only in some laboratory mouse strains, is recent and lies within the first intron. The 5' region of the gene has features of housekeeping gene promoters. It lacks a TATA box but contains a CpG island and Sp1 sites. This region discloses strong promoter activity in transient transfection assays and also stimulates transcription in the reverse orientation, a feature common to other CpG island containing promoters. Transcripts were detected in all the organs tested, although at a variable level, and displayed no strain-dependent differences relative to the IAP insertion, suggesting the existence of mechanisms preserving mCCR4 transcription from the usually deleterious effects of intronic transposition. The strong amino acid conservation between the human, murine, and the previously identified Xenopus CCR4-like proteins, is consistent with an important and conserved role for this protein in vertebrates. PMID- 10521508 TI - Trafficking of the HIV coreceptor CXCR4. Role of arrestins and identification of residues in the c-terminal tail that mediate receptor internalization. AB - The G protein-coupled chemokine receptor CXCR4 serves as the primary coreceptor for entry of T-cell tropic human immunodeficiency virus. CXCR4 undergoes tonic internalization as well as internalization in response to stimulation with phorbol esters and ligand (SDF-1alpha). We investigated the trafficking of this receptor, and we attempted to define the residues of CXCR4 that were critical for receptor internalization. In both COS-1 and HEK-293 cells transiently overexpressing CXCR4, SDF-1alpha and phorbol esters (PMA) promoted rapid internalization of cell surface receptors as assessed by both enzyme-linked immunosorbent assay and immunofluorescence analysis. Expression of GRK2 and/or arrestins promoted modest additional CXCR4 internalization in response to both PMA and SDF. Both PMA- and SDF-mediated CXCR4 internalization was inhibited by coexpression of dominant negative mutants of dynamin-1 and arrestin-3. Arrestin was also recruited to the plasma membrane and appeared to colocalize with internalized receptors in response to SDF but not PMA. We then evaluated the ability of CXCR4 receptors containing mutations of serines and threonines, as well as a dileucine motif, within the C-terminal tail to be internalized and phosphorylated in response to either PMA or SDF-1alpha. This analysis showed that multiple residues within the CXCR4 C-terminal tail appear to mediate both PMA- and SDF-1alpha-mediated receptor internalization. The ability of coexpressed GRK2 and arrestins to promote internalization of the CXCR4 mutants revealed distinct differences between respective mutants and suggested that the integrity of the dileucine motif (Ile-328 and Leu-329) and serines 324, 325, 338, and 339 are critical for receptor internalization. PMID- 10521509 TI - Regulators of G protein signaling 6 and 7. Purification of complexes with gbeta5 and assessment of their effects on g protein-mediated signaling pathways. AB - Regulators of G protein signaling (RGS) proteins that contain DEP (disheveled, EGL-10, pleckstrin) and GGL (G protein gamma subunit-like) domains form a subfamily that includes the mammalian RGS proteins RGS6, RGS7, RGS9, and RGS11. We describe the cloning of RGS6 cDNA, the specificity of interaction of RGS6 and RGS7 with G protein beta subunits, and certain biochemical properties of RGS6/beta5 and RGS7/beta5 complexes. After expression in Sf9 cells, complexes of both RGS6 and RGS7 with the Gbeta5 subunit (but not Gbetas 1-4) are found in the cytosol. When purified, these complexes are similar to RGS11/beta5 in that they act as GTPase-activating proteins specifically toward Galpha(o). Unlike conventional G(betagamma) complexes, RGS6/beta5 and RGS7/beta5 do not form heterotrimeric complexes with either Galpha(o)-GDP or Galpha(q)-GDP. Neither RGS6/beta5 nor RGS7/beta5 altered the activity of adenylyl cyclases types I, II, or V, nor were they able to activate either phospholipase C-beta1 or -beta2. However, the RGS/beta5 complexes inhibited beta(1)gamma(2)-mediated activation of phospholipase C-beta2. RGS/beta5 complexes may contribute to the selectivity of signal transduction initiated by receptors coupled to G(i) and G(o) by binding to phospholipase C and stimulating the GTPase activity of Galpha(o). PMID- 10521510 TI - The dimerization domain of the b subunit of the Escherichia coli F(1)F(0)-ATPase. AB - In this study a series of N- and/or C-terminal truncations of the cytoplasmic domain of the b subunit of the Escherichia coli F(1)F(0) ATP synthase were tested for their ability to form dimers using sedimentation equilibrium ultracentrifugation. The deletion of residues between positions 53 and 122 resulted in a strongly decreased tendency to form dimers, whereas all the polypeptides that included that sequence exhibited high levels of dimer formation. b dimers existed in a reversible monomer-dimer equilibrium and when mixed with other b truncations formed heterodimers efficiently, provided both constructs included the 53-122 sequence. Sedimentation velocity and (15)N NMR relaxation measurements indicated that the dimerization region is highly extended in solution, consistent with an elongated second stalk structure. A cysteine introduced at position 105 was found to readily form intersubunit disulfides, whereas other single cysteines at positions 103-110 failed to form disulfides either with the identical mutant or when mixed with the other 103-110 cysteine mutants. These studies establish that the b subunit dimer depends on interactions that occur between residues in the 53-122 sequence and that the two subunits are oriented in a highly specific manner at the dimer interface. PMID- 10521512 TI - Regulation of bad phosphorylation and association with Bcl-x(L) by the MAPK/Erk kinase. AB - Phosphorylation of the Bcl-2 family protein Bad may represent an important bridge between survival signaling by growth factor receptors and the prevention of apoptosis. Bad phosphorylation was examined following cytokine stimulation, which revealed phosphorylation on a critical residue, serine 112, in a MEK-dependent manner. Furthermore, Bad phosphorylation also increased on several sites distinct from serine 112 but could not be detected on serine 136, previously thought to be a protein kinase B/Akt-targeted residue. Serine 112 phosphorylation was shown to be absolutely required for dissociation of Bad from Bcl-x(L). These results demonstrate for the first time in mammalian cells the involvement of the Ras-MAPK pathway in the phosphorylation of Bad and the regulation of its function. PMID- 10521511 TI - Calcineurin enhances MEF2 DNA binding activity in calcium-dependent survival of cerebellar granule neurons. AB - Myocyte enhancer factor 2 (MEF2) has been shown recently to be necessary for mediating activity-dependent neuronal survival. In this study, we show that calcium signals regulate MEF2 activity through a serine/threonine phosphatase calcineurin. In cultured primary cerebellar granule neurons, the electrophoretic mobility of MEF2A protein was sensitive to the level of extracellular potassium chloride (KCl) and depolarizing concentrations of KCl led to hypophosphorylation of the protein. The specific inhibitors of calcineurin cyclosporin A (CsA) and FK506 could overcome KCl-dependent MEF2A hypophosphorylation. The effects of CsA and FK506 were KCl specific as they had little effect on MEF2A phosphorylation when granule neurons were cultured in the presence of full media. Hyperphosphorylation of MEF2A led to the loss of its DNA binding activity as determined by DNA mobility shift assay. Consistent with this, CsA/FK506 also inhibited MEF2-dependent reporter gene expression. These findings demonstrate that regulation of MEF2A by calcium signals requires the action of protein phosphatase calcineurin. By maintaining MEF2A in a hypophosphorylated state, calcineurin enhances the DNA binding activity of MEF2A and therefore maximizes its transactivation capability. The identification of MEF2 as a novel target of calcineurin may provide in part a biochemical explanation for the therapeutic and toxic effects of immunosuppressants CsA and FK506. PMID- 10521513 TI - More than just a surface thing. Rice infection by magnaporthe grisea PMID- 10521514 TI - A weed reaches new heights down under PMID- 10521515 TI - Function search in a large transcription factor gene family in Arabidopsis: assessing the potential of reverse genetics to identify insertional mutations in R2R3 MYB genes. AB - More than 92 genes encoding MYB transcription factors of the R2R3 class have been described in Arabidopsis. The functions of a few members of this large gene family have been described, indicating important roles for R2R3 MYB transcription factors in the regulation of secondary metabolism, cell shape, and disease resistance, and in responses to growth regulators and stresses. For the majority of the genes in this family, however, little functional information is available. As the first step to characterizing these genes functionally, the sequences of >90 family members, and the map positions and expression profiles of >60 members, have been determined previously. An important second step in the functional analysis of the MYB family, through a process of reverse genetics that entails the isolation of insertion mutants, is described here. For this purpose, a variety of gene disruption resources has been used, including T-DNA-insertion populations and three distinct populations that harbor transposon insertions. We report the isolation of 47 insertions into 36 distinct MYB genes by screening a total of 73 genes. These defined insertion lines will provide the foundation for subsequent detailed functional analyses for the assignment of specific functions to individual members of the R2R3 MYB gene family. PMID- 10521516 TI - Multiple independent defective suppressor-mutator transposon insertions in Arabidopsis: a tool for functional genomics. AB - A new system for insertional mutagenesis based on the maize Enhancer/Suppressor mutator (En/Spm) element was introduced into Arabidopsis. A single T-DNA construct carried a nonautonomous defective Spm (dSpm) element with a phosphinothricin herbicide resistance (BAR) gene, a transposase expression cassette, and a counterselectable gene. This construct was used to select for stable dSpm transpositions. Treatments for both positive (BAR) and negative selection markers were applicable to soil-grown plants, allowing the recovery of new transpositions on a large scale. To date, a total of 48,000 lines in pools of 50 have been recovered, of which approximately 80% result from independent insertion events. DNA extracted from these pools was used in reverse genetic screens, either by polymerase chain reaction (PCR) using primers from the transposon and the targeted gene or by the display of insertions whereby inverse PCR products of insertions from the DNA pools are spotted on a membrane that is then hybridized with the probe of interest. By sequencing PCR-amplified fragments adjacent to insertion sites, we established a sequenced insertion-site database of 1200 sequences. This database permitted a comparison of the chromosomal distribution of transpositions from various T-DNA locations. PMID- 10521517 TI - A two-component enhancer-inhibitor transposon mutagenesis system for functional analysis of the Arabidopsis genome. AB - A modified Enhancer-Inhibitor transposon system was used to generate a series of mutant lines by single-seed descent such that multiple I insertions occurred per plant. The distribution of original insertions in the population was assessed by isolating transposon-flanking DNA, and a database of insertion sites was created. Approximately three-quarters of the identified insertion sites show similarity to sequences stored in public databases, which demonstrates the power of this regimen of insertional mutagenesis. To isolate insertions in specific genes, we developed three-dimensional pooling and polymerase chain reaction strategies that we then validated by identifying mutants for the regulator genes APETALA1 and SHOOT MERISTEMLESS. The system then was used to identify inserts in a class of uncharacterized genes involved in lipid biosynthesis; one such insertion conferred a fiddlehead mutant phenotype. PMID- 10521518 TI - Tonoplast intrinsic protein isoforms as markers for vacuolar functions AB - Plant cell vacuoles may have storage or lytic functions, but biochemical markers specific for the tonoplasts of functionally distinct vacuoles are poorly defined. Here, we use antipeptide antibodies specific for the tonoplast intrinsic proteins alpha-TIP, gamma-TIP, and delta-TIP in confocal immunofluorescence experiments to test the hypothesis that different TIP isoforms may define different vacuole functions. Organelles labeled with these antibodies were also labeled with antipyrophosphatase antibodies, demonstrating that regardless of their size, they had the expected characteristics of vacuoles. Our results demonstrate that the storage vacuole tonoplast contains delta-TIP, protein storage vacuoles containing seed-type storage proteins are marked by alpha- and delta- or alpha- and delta- plus gamma-TIP, whereas vacuoles storing vegetative storage proteins and pigments are marked by delta-TIP alone or delta- plus gamma-TIP. In contrast, those marked by gamma-TIP alone have characteristics of lytic vacuoles, and results from other researchers indicate that alpha-TIP alone is a marker for autophagic vacuoles. In root tips, relatively undifferentiated cells that contain vacuoles labeled separately for each of the three TIPs have been identified. These results argue that plant cells have the ability to generate and maintain three separate vacuole organelles, with each being marked by a different TIP, and that the functional diversity of the vacuolar system may be generated from different combinations of the three basic types. PMID- 10521519 TI - An Arabidopsis cell cycle -dependent kinase-related gene, CDC2b, plays a role in regulating seedling growth in darkness. AB - The Arabidopsis CDC2b gene has been defined as a plant-specific cell cycle dependent kinase-related gene, although it lacks the conserved cyclin binding motif, and its exact function is not known. Here, we report that in etiolated seedlings, the expression of the CDC2b gene is correlated with elongation rate of the hypocotyl. Inhibition of CDC2b gene expression by using an inducible antisense construct resulted in short-hypocotyl and open-cotyledon phenotypes when transgenic seedlings were grown in the dark. The severity of these phenotypes in dark-grown seedlings could be correlated with the level of the antisense gene expression. The short hypocotyl of seedlings underexpressing CDC2b was a result of inhibition of cell elongation rather than a reduction in cell number, whereas in cotyledons, inhibition of CDC2b expression resulted in large, open cotyledons with amyloplasts rather than etioplasts. Although the nuclear DNA was less compact in the antisense hypocotyl cells, DNA content and endoreduplication were not affected. Cell division of the shoot apical meristem also was not affected by antisense expression. The short-hypocotyl phenotype of these transgenic plants was partially rescued by the addition of brassinolide. Brassinolide can only induce CDC2b expression in darkness. These results suggest a role for the CDC2b gene in seedling growth via regulation of hypocotyl cell elongation and cotyledon cell development. PMID- 10521520 TI - ABI1 protein phosphatase 2C is a negative regulator of abscisic acid signaling. AB - The plant hormone abscisic acid (ABA) is a key regulator of seed maturation and germination and mediates adaptive responses to environmental stress. In Arabidopsis, the ABI1 gene encodes a member of the 2C class of protein serine/threonine phosphatases (PP2C), and the abi1-1 mutation markedly reduces ABA responsiveness in both seeds and vegetative tissues. However, this mutation is dominant and has been the only mutant allele available for the ABI1 gene. Hence, it remained unclear whether ABI1 contributes to ABA signaling, and in case ABI1 does regulate ABA responsiveness, whether it is a positive or negative regulator of ABA action. In this study, we isolated seven novel alleles of the ABI1 gene as intragenic revertants of the abi1-1 mutant. In contrast to the ABA resistant abi1-1 mutant, these revertants were more sensitive than the wild type to the inhibition of seed germination and seedling root growth by applied ABA. They also displayed increases in seed dormancy and drought adaptive responses that are indicative of a higher responsiveness to endogenous ABA. The revertant alleles were recessive to the wild-type ABI1 allele in enhancing ABA sensitivity, indicating that this ABA-supersensitive phenotype results from a loss of function in ABI1. The seven suppressor mutations are missense mutations in conserved regions of the PP2C domain of ABI1, and each of the corresponding revertant alleles encodes an ABI1 protein that lacked any detectable PP2C activity in an in vitro enzymatic assay. These results indicate that a loss of ABI1 PP2C activity leads to an enhanced responsiveness to ABA. Thus, the wild-type ABI1 phosphatase is a negative regulator of ABA responses. PMID- 10521522 TI - The Arabidopsis CLAVATA2 gene encodes a receptor-like protein required for the stability of the CLAVATA1 receptor-like kinase. AB - The CLAVATA2 (CLV2) gene regulates both meristem and organ development in Arabidopsis. We isolated the CLV2 gene and found that it encodes a receptor-like protein (RLP), with a presumed extracellular domain composed of leucine-rich repeats similar to those found in plant and animal receptors, but with a very short predicted cytoplasmic tail. RLPs lacking cytoplasmic signaling domains have not been previously shown to regulate development in plants. Our prior work has demonstrated that the CLV1 receptor-like kinase (RLK) is present as a disulfide linked multimer in vivo. We report that CLV2 is required for the normal accumulation of CLV1 protein and its assembly into protein complexes, indicating that CLV2 may form a heterodimer with CLV1 to transduce extracellular signals. Sequence analysis suggests that the charged residue in the predicted transmembrane domain of CLV2 may be a common feature of plant RLPs and RLKs. In addition, the chromosomal region in which CLV2 is located contains an extremely high rate of polymorphism, with 50 nucleotide and 15 amino acid differences between Landsberg erecta and Columbia ecotypes within the CLV2 coding sequence. PMID- 10521521 TI - A defect in beta-oxidation causes abnormal inflorescence development in Arabidopsis. AB - The abnormal inflorescence meristem1 (aim1) mutation affects inflorescence and floral development in Arabidopsis. After the transition to reproductive growth, the aim1 inflorescence meristem becomes disorganized, producing abnormal floral meristems and resulting in plants with severely reduced fertility. The derived amino acid sequence of AIM1 shows extensive similarity to the cucumber multifunctional protein involved in beta-oxidation of fatty acids, which possesses l-3-hydroxyacyl-CoA hydrolyase, l-3-hydroxyacyl-dehydrogenase, d-3 hydroxyacyl-CoA epimerase, and Delta(3), Delta(2)-enoyl-CoA isomerase activities. A defect in beta-oxidation has been confirmed by demonstrating the resistance of the aim1 mutant to 2,4-diphenoxybutyric acid, which is converted to the herbicide 2,4-D by the beta-oxidation pathway. In addition, the loss of AIM1 alters the fatty acid composition of the mature adult plant. PMID- 10521523 TI - Anticipating endoplasmic reticulum stress. A novel early response before pathogenesis-related gene induction AB - When it is attacked by a pathogen, a plant produces a range of defense-related proteins. Many of these are synthesized by the rough endoplasmic reticulum (RER) to be secreted from the cell or deposited in vacuoles. Genes encoding endoplasmic reticulum (ER)-resident chaperones, such as the lumenal binding protein (BiP), are also induced under these conditions. Here, we show that BiP induction occurs systemically throughout the plant. Furthermore, this induction occurs rapidly and precedes expression of genes encoding pathogenesis-related (PR) proteins. The underlying signal transduction pathway was shown to be independent of the signaling molecule salicylic acid and the unfolded protein response pathway. In addition, BiP induction was independent of PR gene induction. Overproduction of BiP alone was not sufficient to cause induction of PR gene expression; however, limiting the amount of BiP in the ER lumen via superimposed ER stress inhibited the induction of PR gene expression. We propose that the induction of BiP expression during plant-pathogen interactions is required as an early response to support PR protein synthesis on the RER and that a novel signal transduction pathway exists to trigger this rapid response. PMID- 10521524 TI - Imprinting of the MEDEA polycomb gene in the Arabidopsis endosperm. AB - In flowering plants, two cells are fertilized in the haploid female gametophyte. Egg and sperm nuclei fuse to form the embryo. A second sperm nucleus fuses with the central cell nucleus that replicates to generate the endosperm, which is a tissue that supports embryo development. MEDEA (MEA) encodes an Arabidopsis SET domain Polycomb protein. Inheritance of a maternal loss-of-function mea allele results in embryo abortion and prolonged endosperm production, irrespective of the genotype of the paternal allele. Thus, only the maternal wild-type MEA allele is required for proper embryo and endosperm development. To understand the molecular mechanism responsible for the parent-of-origin effects of mea mutations on seed development, we compared the expression of maternal and paternal MEA alleles in the progeny of crosses between two Arabidopsis ecotypes. Only the maternal MEA mRNA was detected in the endosperm from seeds at the torpedo stage and later. By contrast, expression of both maternal and paternal MEA alleles was observed in the embryo from seeds at the torpedo stage and later, in seedling, leaf, stem, and root. Thus, MEA is an imprinted gene that displays parent-of origin-dependent monoallelic expression specifically in the endosperm. These results suggest that the embryo abortion observed in mutant mea seeds is due, at least in part, to a defect in endosperm function. Silencing of the paternal MEA allele in the endosperm and the phenotype of mutant mea seeds supports the parental conflict theory for the evolution of imprinting in plants and mammals. PMID- 10521525 TI - Alteration of enod40 expression modifies medicago truncatula root nodule development induced by sinorhizobium meliloti AB - Molecular mechanisms involved in the control of root nodule organogenesis in the plant host are poorly understood. One of the nodulin genes associated with the earliest phases of this developmental program is enod40. We show here that transgenic Medicago truncatula plants overexpressing enod40 exhibit accelerated nodulation induced by Sinorhizobium meliloti. This resulted from increased initiation of primordia, which was accompanied by a proliferation response of the region close to the root tip and enhanced root length. The root cortex of the enod40-transformed plants showed increased sensitivity to nodulation signals. T(1) and T(2) descendants of two transgenic lines with reduced amounts of enod40 transcripts (probably from cosuppression) formed only a few and modified nodulelike structures. Our results suggest that induction of enod40 is a limiting step in primordium formation, and its function is required for appropriate nodule development. PMID- 10521526 TI - Arabidopsis cop8 and fus4 mutations define the same gene that encodes subunit 4 of the COP9 signalosome. AB - The pleiotropic constitutive photomorphogenic/deetiolated/fusca (cop/det/fus) mutants of Arabidopsis exhibit features of light-grown seedlings when grown in the dark. Cloning and biochemical analysis of COP9 have revealed that it is a component of a multiprotein complex, the COP9 signalosome (previously known as the COP9 complex). Here, we compare the immunoaffinity and the biochemical purification of the COP9 signalosome from cauliflower and confirm its eight subunit composition. Molecular cloning of subunit 4 of the complex revealed that it is a proteasome-COP9 complex-eIF3 domain protein encoded by a gene that maps to chromosome 5, near the chromosomal location of the cop8 and fus4 mutations. Genetic complementation tests showed that the cop8 and fus4 mutations define the same locus, now designated as COP8. Molecular analysis of the subunit 4-encoding gene in both cop8 and fus4 mutants identified specific molecular lesions, and overexpression of the subunit 4 cDNA in a cop8 mutant background resulted in complete rescue of the mutant phenotype. Thus, we conclude that COP8 encodes subunit 4 of the COP9 signalosome. Examination of possible molecular interactions by using the yeast two-hybrid assay indicated that COP8 is capable of strong self association as well as interaction with COP9, FUS6/COP11, FUS5, and Arabidopsis JAB1 homolog 1, the latter four proteins being previously defined subunits of the Arabidopsis COP9 signalosome. A comparative sequence analysis indicated that COP8 is highly conserved among multicellular eukaryotes and is also similar to a subunit of the 19S regulatory particle of the 26S proteasome. PMID- 10521527 TI - The role of opaque2 in the control of lysine-degrading activities in developing maize endosperm. AB - We have isolated a cDNA clone, designated ZLKRSDH, encoding the bifunctional enzyme lysine-ketoglutarate reductase/saccharopine dehydrogenase (LKR/SDH) from maize. The predicted polypeptide has an N-terminal LKR domain and a C-terminal SDH domain that are similar to the yeast LYS1 and LYS9 monofunctional proteins, respectively. The maize LKR/SDH protein is located in the cytoplasm of subaleurone endosperm cell layers. Transcripts and polypeptides as well as enzyme activities showed an upregulation and downregulation during endosperm development. The developmental expression of ZLKRSDH was examined in normal and opaque2 seeds. In the mutant endosperm, mRNA levels were reduced by >90%, with concomitant reductions in polypeptide levels and LKR/SDH activity. These results suggest that lysine levels in the endosperm are likely to be controlled at the transcriptional level by the Opaque2 transcription factor. PMID- 10521528 TI - Glutamine synthetase in the phloem plays a major role in controlling proline production AB - To inhibit expression specifically in the phloem, a 274-bp fragment of a cDNA (Gln1-5) encoding cytosolic glutamine synthetase (GS1) from tobacco was placed in the antisense orientation downstream of the cytosolic Cu/Zn superoxide dismutase promoter of Nicotiana plumbaginifolia. After Agrobacterium-mediated transformation, two transgenic N. tabacum lines exhibiting reduced levels of GS1 mRNA and GS activity in midribs, stems, and roots were obtained. Immunogold labeling experiments allowed us to verify that the GS protein content was markedly decreased in the phloem companion cells of transformed plants. Moreover, a general decrease in proline content in the transgenic plants in comparison with wild-type tobacco was observed when plants were forced to assimilate large amounts of ammonium. In contrast, no major changes in the concentration of amino acids used for nitrogen transport were apparent. A (15)NH(4)(+)-labeling kinetic over a 48-hr period confirmed that in leaves of transgenic plants, the decrease in proline production was directly related to glutamine availability. After 2 weeks of salt treatment, the transgenic plants had a pronounced stress phenotype, consisting of wilting and bleaching in the older leaves. We conclude that GS in the phloem plays a major role in regulating proline production consistent with the function of proline as a nitrogen source and as a key metabolite synthesized in response to water stress. PMID- 10521529 TI - Magnaporthe grisea pth11p is a novel plasma membrane protein that mediates appressorium differentiation in response to inductive substrate cues. AB - Mutagenesis of Magnaporthe grisea strain 4091-5-8 led to the identification of PTH11, a pathogenicity gene predicted to encode a novel transmembrane protein. We localized a Pth11-green fluorescent protein fusion to the cell membrane and vacuoles. pth11 mutants of strain 4091-5-8 are nonpathogenic due to a defect in appressorium differentiation. This defect is reminiscent of wild-type strains on poorly inductive surfaces; conidia germinate and undergo early differentiation events, but appressorium maturation is impaired. Functional appressoria are formed by pth11 mutants at 10 to 15% of wild-type frequencies, suggesting that the protein encoded by PTH11 (Pth11p) is not required for appressorium morphogenesis but is involved in host surface recognition. We assayed Pth11p function in multiple M. grisea strains. These experiments indicated that Pth11p can activate appressorium differentiation in response to inductive surface cues and repress differentiation on poorly inductive surfaces and that multiple signaling pathways mediate differentiation. PTH11 genes from diverged M. grisea strains complemented the 4091-5-8 pth11 mutant, indicating functional conservation. Exogenous activation of cellular signaling suppressed pth11 defects. These findings suggest that Pth11p functions at the cell cortex as an upstream effector of appressorium differentiation in response to surface cues. PMID- 10521530 TI - Analysis of the nucleus-encoded and chloroplast-targeted rieske protein by classic and site-directed mutagenesis of Chlamydomonas. AB - Three mutants of the alga Chlamydomonas reinhardtii affected in the nuclear PETC gene encoding the Rieske iron-sulfur protein 2Fe-2S subunit of the chloroplast cytochrome b(6)f complex have been characterized. One has a stable deletion that eliminates the protein; two others carry substitutions Y87D and W163R that result in low accumulation of the protein. Attenuated expression of the stromal protease ClpP increases accumulation and assembly into b(6)f complexes of the Y87D and W163R mutant Rieske proteins in quantities sufficient for analysis. Electron transfer kinetics of these complexes were 10- to 20-fold slower than those for the wild type. The deletion mutant was used as a recipient for site-directed mutant petC alleles. Six glycine residues were replaced by alanine residues (6G6A) in the flexible hinge that is critical for domain movement; substitutions were created near the 2Fe-2S cluster (S128 and W163); and seven C-terminal residues were deleted (G171och). Although the 6G6A and G171och mutations affect highly conserved segments in the chloroplast Rieske protein, photosynthesis in the mutants was similar to that of the wild type. These results establish the basis for mutational analysis of the nuclear-encoded and chloroplast-targeted Rieske protein of photosynthesis. PMID- 10521531 TI - Independent signaling pathways regulate cellular turgor during hyperosmotic stress and appressorium-mediated plant infection by Magnaporthe grisea. AB - The phytopathogenic fungus Magnaporthe grisea elaborates a specialized infection cell called an appressorium with which it mechanically ruptures the plant cuticle. To generate mechanical force, appressoria produce enormous hydrostatic turgor by accumulating molar concentrations of glycerol. To investigate the genetic control of cellular turgor, we analyzed the response of M. grisea to hyperosmotic stress. During acute and chronic hyperosmotic stress adaptation, M. grisea accumulates arabitol as its major compatible solute in addition to smaller quantities of glycerol. A mitogen-activated protein kinase-encoding gene OSM1 was isolated from M. grisea and shown to encode a functional homolog of HIGH OSMOLARITY GLYCEROL1 (HOG1), which encodes a mitogen-activated protein kinase that regulates cellular turgor in yeast. A null mutation of OSM1 was generated in M. grisea by targeted gene replacement, and the resulting mutants were sensitive to osmotic stress and showed morphological defects when grown under hyperosmotic conditions. M. grisea deltaosm1 mutants showed a dramatically reduced ability to accumulate arabitol in the mycelium. Surprisingly, glycerol accumulation and turgor generation in appressoria were unaltered by the Deltaosm1 null mutation, and the mutants were fully pathogenic. This result indicates that independent signal transduction pathways regulate cellular turgor during hyperosmotic stress and appressorium-mediated plant infection. Consistent with this, exposure of M. grisea appressoria to external hyperosmotic stress induced OSM1-dependent production of arabitol. PMID- 10521532 TI - Conserved domains of glycosyltransferases. AB - Glycosyltransferases catalyze the synthesis of glycoconjugates by transferring a properly activated sugar residue to an appropriate acceptor molecule or aglycone for chain initiation and elongation. The acceptor can be a lipid, a protein, a heterocyclic compound, or another carbohydrate residue. A catalytic reaction is believed to involve the recognition of both the donor and acceptor by suitable domains, as well as the catalytic site of the enzyme. To elucidate the structural requirements for substrate recognition and catalytic reactions of glycosyltransferases, we have searched the databases for homologous sequences, identified conserved amino acid residues, and proposed potential domain motifs for these enzymes. Depending on the configuration of the anomeric functional group of the glycosyl donor molecule and of the resulting glycoconjugate, all known glycosyltransferases can be divided into two major types: retaining glycosyltransferases, which transfer sugar residue with the retention of anomeric configuration, and inverting glycosyltransferases, which transfer sugar residue with the inversion of anomeric configuration. One conserved domain of the inverting glycosyltransferases identified in the database is responsible for the recognition of a pyrimidine nucleotide, which is either the UDP or the TDP portion of a donor sugar-nucleotide molecule. This domain is termed "Nucleotide Recognition Domain 1 beta," or NRD1 beta, since the type of nucleotide is the only common structure among the sugar donors and acceptors. NRD1 beta is present in 140 glycosyltransferases. The central portion of the NRD1 beta domain is very similar to the domain that is present in one family of retaining glycosyltransferases. This family is termed NRD1 alpha to designate the similarity and stereochemistry of sugar transfer, and it consists of 77 glycosyltransferases identified thus far. In the central portion there is a homologous region for these two families and this region probably has a catalytic function. A third conserved domain is found exclusively in membrane-bound glycosyltransferases and is termed NRD2; this domain is present in 98 glycosyltransferases. All three identified NRDs are present in archaebacterial, eubacterial, viral, and eukaryotic glycosyltransferases. The present article presents the alignment of conserved NRD domains and also presents a brief overview of the analyzed glycosyltransferases which comprise about 65% of all known sugar-nucleotide dependent (Leloir-type) and putative glycosyltransferases in different databases. A potential mechanism for the catalytic reaction is also proposed. This proposed mechanism should facilitate the design of experiments to elucidate the regulatory mechanisms of glycosylation reactions. Amino acid sequence information within the conserved domain may be utilized to design degenerate primers for identifying DNA encoding new glycosyltransferases. PMID- 10521533 TI - God must love galectins; he made so many of them. PMID- 10521534 TI - Structural characterization of gangliosides isolated from mullet milt using electrospray ionization-tandem mass spectrometry. AB - Electrospray ionization (ESI) coupled with tandem mass spectrometry has been used in conjunction with microwave-mediated saponification, periodate oxidation, and clostridial sialidase hydrolysis to enable detailed structural characterization of gangliosides and their derivatives present in mullet milt. The gangliosides extracted from mullet milt were determined to be GM3, GM3 lactone, GM3 methyl ester, and 9-O-acetyl GM3. For the major ganglioside GM3 and all GM3 derivatives, the ceramide composition was revealed to be C18:1/C16:0. GM3 with a C18:0/C16:0 ceramide was also found as a minor ganglioside. Both the ganglioside intramolecular ester and the ganglioside methyl ester (lacking carboxylic acid groups) showed dominant chloride attachment peaks (M + Cl)- in negative ion ESI MS in addition to low intensity peaks corresponding to (M-H)-. GM3 and O-acetyl GM3 bearing carboxylic acid functions showed only (M-H)-. In positive ion ESI, GM3 and O-acetyl GM3 revealed (M + 2Na-H)+ peaks in addition to (M + Na)+, indicating free exchange of the carboxylic acid proton with a sodium cation, while the ganglioside intramolecular ester and ganglioside methyl ester with no acidic protons yielded only (M + Na)+. The strategy of employing ESI-MS to detect products of established wet chemical reactions represents a general approach for elucidation of ganglioside structural details. PMID- 10521535 TI - The sugar binding activity of MR60, a mannose-specific shuttling lectin, requires a dimeric state. AB - MR60 is an intracellular membrane protein which has been shown to act as a mannoside specific lectin and to be identical to ERGIC-53, a protein characteristic of the endoplasmic reticulum-Golgi apparatus-intermediate compartment, acting as a shuttle. According to its primary sequence, this MR60/ERGIC-53 protein contains a luminal domain including the carbohydrate recognition domain, a stem, a transmembrane segment and a cytosolic domain. The endogenous MR60/ERGIC-53 protein is spontaneously oligomeric, (dimers and hexamers). In this paper, we study the relationship between the oligomerization state and the sugar binding capacity by using recombinant proteins. The expression of the recombinant proteins was evidenced by immunocytochemistry and by immunoprecipitation followed by SDS-PAGE analysis. The full size recombinant protein binds mannosides and is oligomeric, up to the hexameric form. Two truncated proteins lacking the transmembrane and the cytosolic domains were prepared and characterized. A long one, containing the cysteine 466 close to the C-terminal end of the recombinant protein but lacking the cysteine 475, close to the C-terminal end of the native protein, does bind mannosides and forms dimers but no higher oligomeric forms. A shorter one, lacking both the cysteines 466 and 475, does not bind mannosides and does not form dimers or higher polymers. The two cysteines in the carbohydrate recognition domain (C190 and C230) are not involved in the stabilization of oligomers. In conclusion, this study shows that the luminal moiety of MR60/ERGIC-53 contains a device allowing both its oligomeric pattern and its sugar binding capability. PMID- 10521536 TI - Hepatic acute phase induction of murine beta-galactoside alpha 2,6 sialyltransferase (ST6Gal I) is IL-6 dependent and mediated by elevation of exon H-containing class of transcripts. AB - Hepatic expression of CMP-NeuAc:Gal beta 1,4GlcNAc alpha 2,6-sialyltransferase (ST6Gal I) is induced as part of the acute phase response in mammals by mechanisms that remain poorly understood. Previous work suggests that murine liver ST6Gal I mRNA contains an additional and novel region that is not found on ST6Gal I mRNA from human HepG2 hepatoma cells and from rat liver. This novel region, residing 5' of the common Exon I sequence, is encoded by a discrete upstream exon, Exon H. Here we provide evidence that the Exon H-containing transcript is the murine counterpart of the human and rat ST6Gal I mRNAs transcribed from the hepatic-specific promoter, P1. Exon H-containing ST6Gal I mRNA is expressed in all three mice strains examined: balb/c, C57B46, and 129Sv. Furthermore, murine RNA tissue survey indicates that presence of Exon H containing transcripts is restricted to the liver. When mice are subjected to subcutaneous injection of turpentine to elicit the hepatic acute phase response, greater than 4-fold elevation in liver ST6Gal I mRNA was observed. Consistent with the view that Exon H-containing transcripts is regulated by the murine P1 promoter, 5'-RACE analysis indicates that the majority of these transcripts contains the Exon H sequence. This is consistent with the view that Exon H containing transcripts are regulated by the murine P1 region. To assess the mechanism of ST6Gal I response in the hepatic acute phase reaction, mice harboring lesions in both alleles of the IL-6 gene were examined. IL-6(-/-) animals expressed normal levels of ST6Gal I mRNA in liver, with Exon H-containing transcripts remaining the predominant mRNA isoform. However, hepatic ST6Gal I is not elevated upon turpentine injection in the IL-6(-/-) animals. These results indicate that ST6Gal I induction in mouse liver during the acute phase reaction is mediated predominantly by the IL-6 pathway, and results in the induction of the Exon H-containing class of ST6Gal I mRNA that is specific to the liver. PMID- 10521537 TI - Scanning the available Dictyostelium discoideum proteome for O-linked GlcNAc glycosylation sites using neural networks. AB - Dictyostelium discoideum has been suggested as a eukaryotic model organism for glycobiology studies. Presently, the characteristics of acceptor sites for the N acetylglucosaminyl-transferases in Dictyostelium discoideum, which link GlcNAc in an alpha linkage to hydroxyl residues, are largely unknown. This motivates the development of a species specific method for prediction of O-linked GlcNAc glycosylation sites in secreted and membrane proteins of D. discoideum. The method presented here employs a jury of artificial neural networks. These networks were trained to recognize the sequence context and protein surface accessibility in 39 experimentally determined O-alpha-GlcNAc sites found in D. discoideum glycoproteins expressed in vivo. Cross-validation of the data revealed a correlation in which 97% of the glycosylated and nonglycosylated sites were correctly identified. Based on the currently limited data set, an abundant periodicity of two (positions-3, -1, +1, +3, etc.) in Proline residues alternating with hydroxyl amino acids was observed upstream and downstream of the acceptor site. This was a consequence of the spacing of the glycosylated residues themselves which were peculiarly found to be situated only at even positions with respect to each other, indicating that these may be located within beta-strands. The method has been used for a rapid and ranked scan of the fraction of the Dictyostelium proteome available in public databases, remarkably 25-30% of which were predicted glycosylated. The scan revealed acceptor sites in several proteins known experimentally to be O-glycosylated at unmapped sites. The available proteome was classified into functional and cellular compartments to study any preferential patterns of glycosylation. A sequence based prediction server for GlcNAc O-glycosylations in D. discoideum proteins has been made available through the WWW at http://www.cbs.dtu.dk/services/DictyOGlyc/ and via E-mail to DictyOGlyc@cbs.dtu.dk. PMID- 10521538 TI - Interaction between collagens and glycosaminoglycans investigated using a surface plasmon resonance biosensor. AB - The interactions of glycosaminoglycans with collagens and other glycoproteins in extracellular matrix play important roles in cell adhesion and extracellular matrix assembly. In order to clarify the chemical bases for these interactions, glycosaminoglycan solutions were injected onto sensor surfaces on which collagens, fibronectin, laminin, and vitronectin were immobilized. Heparin bound to type V collagen, type IX collagen, fibronectin, laminin, and vitronectin; and chondroitin sulfate E bound to type II, type V, and type VII collagen. Heparin showed a higher affinity for type IX collagen than for type V collagen. On the other hand, chondroitin sulfate E showed the highest affinity for type V collagen. The binding of chondroitin sulfate E to type V collagen showed higher affinity than that of heparin to type V collagen. These data suggest that a novel characteristic sequence included in chondroitin sulfate E is involved in binding to type V collagen. PMID- 10521539 TI - Mice infected with Schistosoma mansoni generate antibodies to LacdiNAc (GalNAc beta 1-->4GlcNAc) determinants. AB - Schistosoma mansoni is a parasitic trematode infecting humans and animals. We reported previously that adult S. mansoni synthesizes complex type biantennary N glycans bearing the terminal sequence GalNAc beta 1-->4GlcNAc-R (lacdiNAc or LDN). We now report that mice infected with S. mansoni generate antibodies to LDN, as assessed by ELISA using a synthetic neoglycoconjugate containing LDN sequences. Sera of infected mice, but not uninfected mice, contained primarily IgM and low levels of IgG toward LDN. Interestingly, these antibodies also recognize bovine milk glycoproteins, which are known to express LDN sequences. The anti-LDN in sera of infected mice were affinity purified on immobilized bovine milk glycoproteins and shown to specifically bind LDN. An IgM monoclonal antibody (SMLDN1.1) was derived from the spleens of S. mansoni infected mice and shown to specifically bind LDN determinants. Immunoblots with affinity purified anti-LDN and SMLDN1.1 demonstrate that LDN sequences occur primarily on N-glycans of numerous glycoproteins of adult S. mansoni. LDN sequences are also expressed in many glycoproteins from S. japonicum and S. haematobium. The availability of antibody to LDN determinants should aid in defining the roles of these glycans in helminth and vertebrate biology. PMID- 10521540 TI - Characterization of the oligosaccharides assembled on the Pichia pastoris expressed recombinant aspartic protease. AB - Aspartic protease, widely used as a milk-coagulating agent in industrial cheese production, contains three potential N-glycosylation sites. In this study, we report the characterization of N-linked oligosaccharides on recombinant aspartic protease secreted from the methylotrophic yeast Pichia pastoris using a combination of mass spectrometric, 2D chromatographic, chemical and enzymatic methods. The carbohydrates from site I (Asn79) were found to range from Man6 17GlcNAc2 with 50% bearing a phospho-diester-motif, site II (Asn113) was not occupied and site III (Asn188) contained mostly uncharged species ranging from Man-13GlcNAc2. These charged groups are not affecting the transport through the secretion pathway of the recombinant glycoprotein. Changes from a molasses-based medium to a minimal salts-based medium led to a clear reduction of the degree of phosphorylation of the N-glycan population. PMID- 10521541 TI - Mnt2p and Mnt3p of Saccharomyces cerevisiae are members of the Mnn1p family of alpha-1,3-mannosyltransferases responsible for adding the terminal mannose residues of O-linked oligosaccharides. AB - The genome of Saccharomyces cerevisiae contains five genes that encode type II transmembrane proteins with significant amino acid similarity to the alpha-1,3 mannosyltransferase Mnn1p. The roles of the three genes most closely related to MNN1 were examined in mutants carrying single and multiple combinations of the disrupted genes. Paper chromatographic analysis of [2-3H]mannose-labeled O-linked oligosaccharides released by beta-elimination showed that the MNT2 (YGL257c) and MNT3 (YIL014w) genes in combination with MNN1 have overlapping roles in the addition of the fourth and fifth alpha-1,3-linked mannose residues to form Man4 and Man5 oligosaccharides whereas MNT4 (YNR059w) does not appear to be required for O-glycan synthesis. PMID- 10521542 TI - Characterization of mammalian UDP-GalNAc:glucuronide alpha 1-4-N acetylgalactosaminyltransferase. AB - We previously reported that cultured cells incubated with beta-xylosides synthesized alpha-GalNAc-capped GAG-related xylosides, GalNAc alpha GlcA beta Gal beta Gal beta Xyl beta-R and GalNAc alpha GlcA beta GalNAc beta GlcA beta Gal beta Gal beta Xyl beta-R, where R is 4-methylumbelliferyl or p-nitrophenyl (Manzi et al., 1995; Miura and Freeze, 1998). In this study, we characterized an alpha-N acetylgalactosaminyltransferase (alpha-GalNAc-T) that probably adds the alpha GalNAc residue to the above xylosides. Microsomes from several animal cells and mouse brain contained the enzyme activity which requires divalent cations, and has a relatively broad pH optimal range around neutral. The apparent K(m) values were in the submillimolar range for the acceptors tested, and 19 microM for UDP GalNAc. 1H-NMR analysis of the GlcA-beta-MU acceptor product showed the GalNAc residue is transferred in alpha 1,4-linkage to the glucuronide, which is consistent with previous results reported on alpha-GalNAc-capped Xyl-MU (Manzi et al., 1995). Various artificial glucuronides were tested as acceptors to assess the influence of the aglycone. Glucuronides with a bicyclic aromatic ring, such as 4-methylumbelliferyl beta-D-glucuronide (GlcA-beta-MU) and alpha-naphthyl beta D-glucuronide, were the best acceptors. Interestingly, a synthetic acceptor that resembles the HNK-1 carbohydrate epitope but lacking the sulfate group, GlcA beta 1,3Gal beta 1,4GlcNAc beta-O-octyl (delta SHNK-C8), was a better acceptor for alpha-GalNAc-T than the glycosaminoglycan-protein linkage region tetrasaccharyl xyloside, GlcA beta 1,3Gal beta 1,3Gal beta 1,4Xyl beta-MU. GlcA-beta-MU and delta SHNK-C8 competed for the alpha-GalNAc-T activity, suggesting that the same activity catalyzes the transfer of the GalNAc residue to both acceptors. Taken together, the results show that the alpha-GalNAc-T described here is not restricted to GAG-type oligosaccharide acceptors, but rather is a UDP GalNAc:glucuronide alpha 1-4-N-acetylgalactosaminyltransferase. PMID- 10521543 TI - High-level expression of the Neisseria meningitidis lgtA gene in Escherichia coli and characterization of the encoded N-acetylglucosaminyltransferase as a useful catalyst in the synthesis of GlcNAc beta 1-->3Gal and GalNAc beta 1-->3Gal linkages. AB - We have expressed the Neisseria meningitidis lgtA gene at a high level in Escherichia coli. The encoded beta-N-acetylglucosaminyltransferase, referred to as LgtA, which in the bacterium is involved in the synthesis of the lacto-N-neo tetraose structural element of the bacterial lipooligosaccharide, was obtained in an enzymatically highly active form. This glycosyltransferase appeared to be unusual in that it displays a broad acceptor specificity toward both alpha- and beta-galactosides, whether structurally related to N- or O-protein-, or lipid linked oligosaccharides. Product analysis by one- and two-dimensional 400 MHz 1H- and 13C-NMR spectroscopy reveals that LgtA catalyzes the introduction of GlcNAc from UDP-GlcNAc in a beta 1-->3-linkage to accepting Gal residues. The enzyme can thus be characterized as a UDP-GlcNAc:Gal alpha/beta-R beta 3-N acetylglucosaminyltransferase. Although lactose is a highly preferred acceptor substrate the recombinant enzyme also acts efficiently on monomeric and dimeric N acetyllactosamine revealing its potential value in the synthesis of polylactosaminoglycan structures in enzyme assisted procedures. Furthermore, LgtA shows a high donor promiscuity toward UDP-GalNAc, but not toward other UDP sugars, and can catalyze the introduction of GalNAc in beta 1-->3-linkage to alpha- or beta-Gal in the acceptor structures at moderate rates. LgtA therefore shows promise to be a useful catalyst in the preparative synthesis of both GlcNAc beta 1-->3Gal and GalNAc beta 1-->3Gal linkages. PMID- 10521544 TI - Cloning and expression of a specific human alpha 1,2-mannosidase that trims Man9GlcNAc2 to Man8GlcNAc2 isomer B during N-glycan biosynthesis. AB - We report the isolation of a novel human cDNA encoding a type II membrane protein of 79.5 kDa with amino acid sequence similarity to Class I alpha 1,2 mannosidases. The catalytic domain of the enzyme was expressed as a secreted protein in Pichia pastoris. The recombinant enzyme removes a single mannose residue from Man9GlcNAc and [1H]-NMR analysis indicates that the only product is Man8GlcNAc isomer B, the form lacking the middle-arm terminal alpha 1,2-mannose. Calcium is required for enzyme activity and both 1-deoxymannojirimycin and kifunensine inhibit the human alpha 1,2-mannosidase. The properties and specificity of this human alpha 1,2-mannosidase are identical to the endoplasmic reticulum alpha 1,2-mannosidase from Saccharomyces cerevisiae and differ from those of previously cloned Golgi alpha 1,2-mannosidases that remove up to four mannose residues from Man9GlcNAc2 during N-glycan maturation. Northern blot analysis showed that all human tissues examined express variable amounts of a 3 kb transcript. This highly specific alpha 1,2-mannosidase is likely to be involved in glycoprotein quality control since there is increasing evidence that trimming of Man9GlcNAc2 to Man8GlcNAc2 isomer B in yeast cells is important to target misfolded glycoproteins for degradation. PMID- 10521545 TI - CD80 (B7-1) and CD86 (B7-2) expression in multiple sclerosis patients: clinical subtype specific variation in peripheral monocytes and B cells and lack of modulation by high dose methylprednisolone. AB - Autoimmune activation of T cells by central nervous system (CNS)-derived antigens is hypothesised to underlie neural damage in multiple sclerosis (MS) patients. The role of coreceptor mediated signalling is currently under investigation in order to further elucidate the immunopathogenic mechanisms implicated and to determine possible targets for immune modulation. We have investigated whether differential coreceptor (B7-1/CD80; B7-2/CD86; CD28) expression on circulating lymphocytes and monocytes is (i) a feature of distinctive clinical subtypes of MS (relapsing-remitting in remission/stable-RRMS; relapsing-remitting in relapse/relapsing-RRMS; primary progressive/PPMS), (ii) related to disease activity, and (iii) altered by high dose corticocosteroid treatment. CD80(+) B cells were significantly reduced (P<0.05) in PPMS (4.0+/-0.8%) compared with normal subjects (CON) (9.1+/-1.1%), stable-RRMS (6.7+/-0.7%) and relapsing-RRMS (7.8+/-0.9%) patients. Comparatively fewer monocytes from relapsing-RRMS patients expressed CD86 (relapsing-RRMS 50+/-4.9% vs. stable-RRMS 75.1+/-3.4%, PPMS 77. 7+/-3.2%, CON 72.1+/-3.6%/P<0.05). Otherwise expression of coreceptors did not vary significantly between the groups. A 3-day course of methylprednisolone therapy did not alter coreceptor expression, but did suppress monocyte and B cell HLA-DR expression. There is evidence for differential coreceptor expression on circulating B cells and monocytes in MS disease subtypes. The biological significance of these findings is discussed in relation to alternative theories regarding coreceptor functioning. PMID- 10521547 TI - Motor and somatosensory evoked potential findings in HTLV-I associated myelopathy. AB - The central motor and sensory conduction times were measured using the motor and somatosensory evoked potentials in 21 patients with HTLV-I associated myelopathy/tropical spastic paraparesis (HAM/TSP) to evaluate the functions of the corticospinal tract and posterior column. The control subjects were closely matched for gender, age and height with the patients in the central sensory conduction time (CSCT) and central motor conduction time (CMCT) studies. An abnormal upper limb (UL) CMCT was present in six of 34 limbs (17.6%) while the lower limb (LL) CMCT was abnormal in 31 of 42 limbs (73.8%). No patients had an abnormal UL CSCT while the LL CSCT was abnormal in 10 of 32 limbs (31.3%). The frequency of abnormal LL CMCT was significantly higher than that for an abnormal LL CSCT. The duration of the disease and the disability score did not significantly influence the evoked potential findings. These results indicate that major lesions of the spinal cord in HAM/TSP patients tend to be localized more in the descending corticospinal pathways than in the ascending posterior columns at the thoracic spinal cord level. This finding also closely agrees with the clinico-pathological findings of HAM/TSP. PMID- 10521546 TI - X-linked dominant Charcot-Marie-Tooth neuropathy: clinical, electrophysiological, and morphological phenotype in four families with different connexin32 mutations(1). AB - The sensorimotor neuropathy of the Charcot-Marie-Tooth type (CMT) is the most common hereditary disorder of the peripheral nervous system. The X-linked dominant form of CMT (CMTX) is associated with mutations in the gene for the gap junction protein connexin32. We examined four CMTX pedigrees two of which had potentially novel mutations in the only coding exon of connexin32. One previously unreported missense mutation, Ala39Val, was found in a family displaying a CMT phenotype with additional upper limb postural tremor reminiscent of a Roussy-Levy syndrome. A novel single base insertion, 679insT, is among the first mutations found in the fourth transmembrane domain of connexin32. Frameshift and premature stop of translation are supposed to result in a non-functional carboxy-terminus. Two further families had the known missense mutations Arg15Trp and Arg22Gln. Several female carriers were found normal on clinical presentation, however, the genotype was paralleled by decreased nerve conduction velocities (NCV) and slowed central conduction of brain stem auditory evoked responses (BAER). Median motor NCVs showed mild (in women) to intermediate (in males) reduction, indicating a peripheral neuropathy with a predominating axonal component. Nerve biopsy findings were consistent with the electrophysiological data showing a marked loss of large myelinated fibres and clusters of regenerating axons. Electron microscopy revealed various alterations of the axoglial attachment zone. This suggests defective axon-Schwann cell interactions which may induce the axonopathy in CMTX. PMID- 10521548 TI - Relationship between the clinical manifestations, computed tomographic findings and the outcome in 80 patients with primary pontine hemorrhage. AB - The relationship between the clinical manifestations, computed tomographic (CT) findings and the outcome in 80 patients with primary pontine hemorrhage (PPH) was analyzed to clarify factors predicting the outcome. Patients were 58 males and 22 females (60. 5+/-12.9 years old) with PPH. Correlations between the clinical manifestations, CT findings and the outcome was assessed with multivariate regression analysis. The initial level of consciousness and the transaxial size of the hematoma on CT were strongly related to the outcome. In patients with small hematoma (with the transaxial size 85% of the variability in kyphosis. There was a trend for a more pronounced anterior wedge configuration of the midthoracic vertebral bodies and discs. Higher associations between variables were also noted in this region. CONCLUSIONS: The normal kyphosis of the thoracic spine reflects the morphological adaptation of both the vertebral bodies and intervertebral discs. RELEVANCE: This study contributes new data on the thoracic spine, particularly the characteristics of thoracic discs and their contribution to kyphosis genesis. Future directions for morphology studies should encompass more detailed examination of the thoracic discs and greater emphasis on the midthoracic segments, considering the prevalence of osteoporosis related fractures and subsequent deformity at these levels. PMID- 10521628 TI - The influence of trunk modelling in 3D biomechanical analysis of simple and complex lifting tasks. AB - OBJECTIVE: The purpose of this study was to evaluate different methods of estimating the body segment parameters and different methods of partitioning the trunk in order to reduce errors in the inverse dynamic analysis of lifting tasks. DESIGN: The same data set was used to evaluate moment errors associated to five linked models differing by the way the trunk was modelled. BACKGROUND: The inverse dynamic analysis of complex lifting tasks involving significant lower limb displacements requires the use of upper body linked models. However, the estimation of the body segment parameters of trunk segments and the flexible properties of the trunk can lead to errors when using this modelling approach. METHODS: Twenty-one male subjects performed four lifting tasks. Five cameras, two force platforms and a dynamometric box provided inputs to five tridimensional (3D) dynamic linked models. Three modelling parameters of the trunk were tested in these models: (1) the use of a geometric or a proportional anthropometric model to estimate the body segment parameters of the trunk, (2) the location of the antero-posterior position of the centre of mass of the trunk segments (at a percentage of the trunk depth vs on a line between hips and shoulders), and (3) the partitioning of the trunk in two or three segments. The behavior of these linked models was assessed with three different error analyses. RESULTS: The results revealed that all three modelling parameters of the trunk can reduce moment errors, especially when applied to subjects characterized by a larger abdomen. CONCLUSIONS: The inverse dynamic analysis of lifting tasks using an upper body modelling approach should take into consideration the interindividual variability inherent to the trunk morphology and non-rigidity. RELEVANCE: The trunk geometry and flexibility shows considerable variability among individuals. The use of upper body linked models requires adequate modelling of this segment to minimize errors in 3D inverse dynamic analysis of lifting tasks. PMID- 10521629 TI - Normal global motion of the cervical spine: an electrogoniometric study. AB - OBJECTIVE: Establishment of a normal database and clinical reference of active global cervical spine motion ranges and patterns using a commercial electrogoniometer. DESIGN: Three-dimensional cervical motion ranges and patterns were analyzed in 250 asymptomatic volunteers. BACKGROUND: In vivo out-of-plane motion patterns of the cervical spine have not yet been reported in large populations, but could be of clinical interest. METHODS: In 250 subjects (aged 14 70 yr), motion range and patterns between the first thoracic vertebra and the head were analyzed for flexion-extension, lateral bending, rotation in neutral sagittal plane position and in full flexion using the CA 6000 Spine Motion Analyzer. RESULTS AND CONCLUSIONS: Average motion range in the sagittal plane was 122 degrees (SD: 18 degrees ). Flexion was slightly more important than extension. Out-of-plane components were negligible. Global bending range averaged 88 degrees (SD: 16 degrees ), left and right bending being comparable. Homolateral rotation was associated to lateral bending. Its extent was approximately 40% of the bending range. Global rotation range in neutral sagittal plane position was 144 degrees (SD: 20 degrees ), without significant difference between right and left rotations. Associated motion components were small. During rotation in flexed head position, global range (134 degrees, SD: 24 degrees ) was comparable to the one in neutral flexion. But heterolateral bending, averaging 60% of the primary motion, was associated to flexed rotation. Significant reduction of all primary (but not conjunct) motions with age were obtained. Sex had no influence on cervical motion range. RELEVANCE: Our results agreed with previous observations, validating the methodology used. They thus constitute reference data of cervical out-of-plane motion for clinical applications. PMID- 10521630 TI - Age related biomechanical properties of the glenoid-anterior band of the inferior glenohumeral ligament-humerus complex. AB - OBJECTIVE: To quantify the biomechanical properties of the glenoid-anterior band of the inferior glenohumeral ligament-humerus complex for the two age groups. DESIGN: In vitro human cadaver study evaluating the biomechanical properties of the glenoid-anterior band of the inferior glenohumeral ligament-humerus complex for a younger group (n=5, mean age 38.5, SD 0.5 years) and an older group (n=7, mean age 74.8, SD 5.3 years). BACKGROUND: Glenohumeral instability is more of a problem in younger than in older individuals, primarily because recurrence is much more common at a young age. METHODS: Tensile testing was performed on the glenoid-anterior band of the inferior glenohumeral ligament-humerus complex in the shoulder apprehension position using a custom jig, Instron machine and a video digitizing system. RESULTS: In the younger individuals disruption of the complex most often occurred at the glenoid-labrum region of the glenoid insertion site. In the older individual, disruption most often occurred at the midsubstance region. The load and the stress at failure of the glenoid-anterior band of the inferior glenohumeral ligament-humerus complex showed that the older group was only 61% and 46% of the younger group, respectively. CONCLUSIONS: The structural properties of the glenoid-anterior band of the inferior glenohumeral ligament humerus complex and the material characteristics of the anterior band of the inferior glenohumeral ligament for the younger group were significantly superior than the older group. RELEVANCE: A stronger and more extensive repair, such as the traditional open technique, may be necessary for younger individuals with glenohumeral instability whereas in older individuals, a different repair technique, such as an arthroscopic technique, may be sufficient. PMID- 10521631 TI - A simulation of muscle force and internal kinematics of extensor carpi radialis brevis during backhand tennis stroke: implications for injury. AB - OBJECTIVE: The purpose of this work was use a computer simulation of the action of extensor carpi radialis brevis during a typical backhand tennis stroke of novice and advance players to examine a potential mechanism of injury. DESIGN: This study uses established kinematic data in conjunction with a computer model to give a time varying description of muscle force and length changes. BACKGROUND: Lateral epicondylitis or tennis elbow has been attributed to over exertion of extensor carpi radialis brevis with novice tennis players being particularly susceptible. METHODS: We used a simple Hill-type muscle model to predict muscle force and internal kinematics based on activation and joint angle changes as inputs. Magnetic resonance images were used to determine the morphometric dimensions of extensor carpi radialis brevis which were used to scale the mechanical properties determined from in vivo contractions of flexor pollicis longus. RESULTS: The simulation indicated that the novice group generated considerably less force and the muscle was subjected to a substantial eccentric contraction as a result of racquet-ball impact. This eccentric contraction occurred with the muscle at a very long length with diminishing tension capabilities. CONCLUSION: The observed pattern of activation and joint kinematics of novice tennis players results in substantial eccentric contractions which are likely the cause of repetitive microtrauma leading to tennis elbow injuries. Adopting the technique seen in advanced players would limit the eccentric contractions and reduce the likelihood of injury. RELEVANCE: Lateral epicondylitis can be extremely problematic because of its chronic nature and relatively high incidence. This study offers one aetiology of the condition that results from improper kinematics during the tennis backhand stroke. PMID- 10521632 TI - Flat foot functional evaluation using pattern recognition of ground reaction data. AB - OBJECTIVE: Main purpose of this study was to apply quantitative gait analysis and statistical pattern recognition as clinical decision-making aids in flat foot diagnosis and post-surgery monitoring. DESIGN: Statistical pattern recognition techniques were applied to discriminate between normal and flat foot populations through ground reaction force measurements; ground reaction forces time course was assumed as a sensible index of the foot function. BACKGROUND: Gait analysis is becoming recognized as an important clinical tool in orthopaedics, in pre surgery planning, post-surgery monitoring and in a posteriori evaluation of different treatment techniques. Statistical pattern recognition techniques have been utilized with success in this field to identify the most significant variables of selected motor functions in different pathologies, and to design classification rules and quantitative evaluation scores. METHODS: Ground reaction forces were recorded during free speed barefoot walks on 28 healthy subjects, and 28 symptomatic flexible flat foot children selected for surgical intervention. A new feature selection algorithm, based on heuristic optimization, was applied to select the most discriminant ground reaction forces time samples. A two-stage pattern recognition system, composed by three linear feature extractors, one for each ground reaction force component, and a linear classifier, was designed to classify the feet of each subject using the selected features. The output of the classifier was used to define a functional score. RESULTS: The classifier assigned the ground reaction force patterns performed by each subject into the right class with an estimated error of 15%, corresponding to an assignment error for each subject's foot of 9%. The most discriminant ground reaction forces time samples selected are in full agreement with the pathophysiology of the symptomatic flexible flat foot. The obtained score was utilized to monitor the 1 and 2 years post-operative functional recovery of two differently treated subgroups of 32 flexible flat foot subjects. CONCLUSIONS: Statistical pattern recognition techniques are promising tools for clinical gait analysis; the obtained score provides important functional information that could be used as a further aid in the clinical evaluation of flat foot and different surgical treatment techniques. RELEVANCE: Symptomatic flexible flat foot surgical decision making is frequently difficult because of the lack of objective criteria to assess functional abnormalities of the foot/ankle complex. Gait analysis and statistical pattern recognition can give us parameters with which to characterize "functional" flat foot. Moreover, we can objectively follow up the recovery of the foot/ankle complex function after surgical treatment. PMID- 10521633 TI - Adaptation of the human calcaneus to variations of impact forces during running. AB - OBJECTIVE: The influence of varied forces under the heel induced by changes in midsole hardness on adaptations of the human calcaneus during running training was investigated. DESIGN: A longitudinal study was conducted over a period of 20 weeks with subjects training for 50 km per week on average. BACKGROUND: The skeletal systems' metabolism acts highly dynamic, governed by mechanical factors. The amount of running training has been shown to increase the bone mineral density in the calcaneus. Mechanical factors have not been controlled in former investigations. METHODS: Bone quality parameters were determined before and after the training by use of an ultrasound system and quantitative MRI while the mechanics of foot-ground contact were controlled. The total group of 26 subjects was divided into three subgroups based upon different magnitude of forces under the heel inside the shoe. RESULTS: The biomechanical testing demonstrate no relationship between midsole hardness and external or in-shoe impacts. Bone parameters showed specific differences for all groups which are pronounced in runners with intermediate impacts. CONCLUSIONS: The observed variations reflect metabolic changes in bone marrow which appear to be effected by the impact magnitude and cannot be characterised as negative. RELEVANCE: The current data imply that no negative changes of impacts on calcaneal bone were produced by high amounts of training in distance running. The mechanical testing indicates that the potential of modifying calcaneal adaptation directly by varying midsole hardness is limited. PMID- 10521634 TI - Variability in spine loading model performance. AB - OBJECTIVE: To assess the sources of variability associated with an EMG-assisted model of spine loading. DESIGN: In vivo measurements of trunk dynamics, lifting moments and muscle activities were used as inputs into an EMG-assisted spine loading model. BACKGROUND: Several types of variability are inherent in biomechanical assessments of risk associated with trunk bending motions during lifting. Variability may occur as a function of variations in spine loading due to either subject variations in motion profiles (kinematics) or biomechanical model performance. METHODS: Twelve experienced and inexperienced materials handlers performed 10 repeated lifts where load weight, asymmetry, and velocity were varied. The experiment was replicated on a second day to assess day to day variability. RESULTS: These model performance variables indicated that variability was mainly a function of subject characteristics and experience. Minor variations in variability were associated with the task asymmetry and weight lifted. Advanced analyses suggested that experienced workers had a greater range of back motion compared to inexperienced workers which would affect the length-strength component of the model calibration. CONCLUSIONS: This study indicates that for the results of an EMG-assisted model to be accurate, it is important to ensure that the model reflects a realistic relationship between the trunk muscle length and the muscle force production capacity. Underestimation if this relationship can degrade model fidelity and robustness. RELEVANCE: These results imply that by properly calibrating the model it is then reasonable to assume that the vast majority of variations observed in repeated exertions of a particular trial are due to kinematic and kinetic differences inherent in the muscle control system and not a function of model randomness. PMID- 10521635 TI - The displacement, velocity and frequency profile of the frontal plane motion produced by the cervical lateral glide treatment technique. AB - OBJECTIVE: To investigate the specificity of linear and angular displacement, peak velocity and frequency of oscillation of the frontal plane motion produced by the lateral glide treatment technique of the cervical spine. DESIGN: A within subjects design was used. BACKGROUND: The lateral glide treatment technique of the cervical spine has previously been shown to produce specific neurophysiological effects, whereas the technique's biomechanical effects remain uninvestigated. A relationship may exist between a technique's biomechanical effects and its clinical outcome. METHODS: Eight asymptomatic subjects participated in the study. Retroreflective markers were placed dorsally at the occiput, and the C5, T1, T3 and T5 vertebrae. Movement of these markers was recorded by a video processor. The same physiotherapist applied the treatment technique to each subject. RESULTS: Linear displacement (3.336 cm) and peak velocity (13.643 cm/s) between C5 and T1 markers was greater than between T1 and T3, Occiput and C5, and T3 and T5. Angular displacement about C5 (26.5 degrees ) was twice that about T1 and T3, but unlike T1 and T3 occurred in the opposite direction to that of the treatment technique. Frequency of oscillation was 1. 255 Hz. CONCLUSION: The lateral glide treatment technique produced a characteristic ipsilateral displacement rather than side flexion, predominantly at its region of application. RELEVANCE: This study provides the basis for further investigations of the mechanisms of action of the lateral glide treatment technique. It also provides clinicians with guidelines for the application of this technique. PMID- 10521636 TI - Identification of feigned grip effort using isokinetic dynamometry. AB - OBJECTIVE: To investigate the feasibility of applying isokinetic dynamometry for identifying submaximal grip strength. DESIGN: Measurement of maximal and feigned concentric and eccentric strength at high and low contraction velocities. BACKGROUND: Identification of feigned grip strength is a highly problematic issue which has been challenged using various techniques, invariably related to isometric efforts. This study is based on recent research which has indicated that isokinetic dynamometry was highly efficient in identifying feigned efforts in other major muscle groups. METHODS: Seventeen healthy women aged 20-25 took part in the study. Prior to executing the feigned effort, subjects were told to exert lower than the maximal grip strength in an attempt to obtain financial compensation for a simulated injury to hand musculature which in fact did not result in weakness of grip. RESULTS: Findings indicated that based on a parameter termed DEC which was defined as the difference between the ratios of the eccentric to concentric strength at the high and low velocities, feigned efforts could very effectively be identified (P<0.0001). Furthermore, a multivariate model enabled this identification to be described in terms of the level of confidence by which a claim concerning weakness of grip may be proclaimed as genuine or insincere. CONCLUSIONS: Though the neuromotor mechanisms responsible for grip strength may differ from those acting with respect to other muscle groups, the inability to adjust the eccentric and concentric force components during submaximal efforts is probably a general feature. RELEVANCE: Isokinetic dynamometry is a powerful method for quantifying various aspects of muscle performance. However, its application in the medicolegal area of muscle weakness has only recently been explored. Combined with previous research, this study strongly indicates that if certain trauma or pathology-related weakness does not result in variations in the force-velocity characteristics of the affected muscles, than this technique has the potential to validly differentiate between patients who have a genuine reason for compensation and those ('symptom magnifiers') who do not. PMID- 10521637 TI - An anatomically based protocol for the description of foot segment kinematics during gait. AB - OBJECTIVE: To design a technique for the in vivo description of ankle and other foot joint rotations to be applied in routine functional evaluation using non invasive stereophotogrammetry. DESIGN: Position and orientation of tibia/fibula, calcaneus, mid-foot, 1st metatarsal and hallux segments were tracked during the stance phase of walking in nine asymptomatic subjects. Rigid clusters of reflective markers were used for foot segment pose estimation. Anatomical landmark calibration was applied for the reconstruction of anatomical landmarks. BACKGROUND: Previous studies have analysed only a limited number of joints or have proposed invasive techniques. METHODS: Anatomical landmark trajectories were reconstructed in the laboratory frame using data from the anatomical calibration procedure. Anatomical co-ordinate frames were defined using the obtained landmark trajectories. Joint co-ordinate systems were used to calculate corresponding joint rotations in all three anatomical planes. RESULTS: The patterns of the joint rotations were highly repeatable within subjects. Consistent patterns between subjects were also exhibited at most of the joints. CONCLUSION: The method proposed enables a detailed description of ankle and other foot joint rotations on an anatomical base. Joint rotations can therefore be expressed in the well-established terminology necessary for their clinical interpretation. RELEVANCE: Functional evaluation of patients affected by foot diseases has recently called for more detailed and non-invasive protocols for the description of foot joint rotations during gait. The proposed method can help clinicians to distinguish between normal and pathological pattern of foot joint rotations, and to quantitatively assess the restoration of normal function after treatment. PMID- 10521638 TI - Optimum length of muscle contraction. AB - OBJECTIVE: The purpose of this study was to develop a mathematical method to determine optimum muscle length and muscle stress based on the measurable physiological and biomechanical data. DESIGN: The values of optimum muscle length and muscle stress are investigated. BACKGROUND: Understanding the characteristics of muscle function in vivo is important for assisting the design of the tendon transfer and other rehabilitation procedures. In vivo determination of the physiological and anatomical parameters of muscle contraction is difficult but not impossible. Optimum muscle length and muscle stresses are important parameters for understanding muscle function. METHODS: A Cybex dynamometer was used to measure isometric elbow flexion torque in eight different joint positions in seven subjects. Then the optimization method was used to determine optimum muscle length and muscle stress of three major elbow flexors, the biceps brachii, the brachialis, and the brachioradialis based on the model and joint torque data. RESULTS: The calculated muscle stress for each subject was on average 109 N/cm(2), while the optimum muscle length for the biceps brachii, the brachialis, and the brachioradialis was on average 14.05, 6.53, 17.24 cm, respectively. The joint angles corresponding to these optimum muscle lengths are 110 degrees, 100 degrees and 50 degrees of elbow flexion, respectively. CONCLUSIONS: Optimum muscle length and muscle stress can be properly predicted using an analytical mathematical model along with an experimentally measured joint torque. RELEVANCE: The estimate of optimum muscle length is important for muscle modeling and tendon transfer surgery by taking advantage of length-tension relationship of individual muscles. PMID- 10521639 TI - Damage to rabbit femoral articular cartilage following direct impacts of uniform stresses: an in vitro study. AB - OBJECTIVE: To determine the acute gross and histologic damage resulting to femoral cartilage from an in vitro direct impact of uniform stress. DESIGN: Gross and histologic evaluations were performed on rabbit femoral condyles impacted by a drop-tower device. BACKGROUND: It is thought that impacts above a given threshold stress may initiate post-traumatic arthritis. The extent of damage following impacts of specific stress has not been previously studied. METHODS: 12 New Zealand White rabbit medial femoral condyles were divided into three groups by impact type and magnitude. A drop tower was used to strike the femoral condyle with a flat impactor, or with a custom contoured impactor. Gross and histological grades were given depending on the depth and number of fissures and cracks in the impacted condyle. RESULTS: The degree of damage correlated best with the type of impactor used and with the impact force; correlation between damage and impact stress was less significant. Contoured impactors tended to produce superficial fibrillation, while flat impactors tended to produce deep cracks. Impact forces above 500 N tended to create more severe damage than impact forces below 500 N. Subchondral bone remained intact in all cases and deep cartilage damage did not occur without disruption of more superficial layers. Poor correlation was found between damage as graded by gross examination versus damage graded histologically. CONCLUSIONS: Acute damage corresponds best with type of impactor and impact force, and not as well with impact stress. Micro structural injuries may be present in the absence of gross findings. RELEVANCE: Post-traumatic arthritis is a disabling disease thought to occur when a blow of stress above a given threshold is delivered to articular cartilage. Current animal models of post-traumatic arthritis are unable to characterize the impact stress applied to an articular surface. This study examines grossly and histologically the structural damage occurring as a result of impacts of given stress and force. PMID- 10521640 TI - novel Award First Prize Paper. Orthotic management of plantar pressure and pain in rheumatoid arthritis. AB - OBJECTIVE: To investigate the effectiveness of foot orthoses in the management of plantar pressure and pain in subjects with rheumatoid arthritis. DESIGN: A repeated measures study in which the independent variable was orthosis design. Dependent variables, including pressure, gait and pain parameters, were examined using analysis of variance and correlation statistics. BACKGROUND: The aim of orthotic management of the rheumatoid foot is to relieve metatarsalgia through the reduction of metatarsal head pressure. Few studies have investigated the relative effectiveness of different orthosis designs. To date, no studies have examined the relationship between plantar pressure and second metatarsal head pain in rheumatoid arthritis subjects. METHODS: Twelve rheumatoid arthritis subjects with foot involvement and second metatarsal head pain were tested. Four styles of foot orthosis (prefabricated, standard custom moulded, custom with metatarsal bar, custom with metatarsal dome) were compared to a shoe only control. An EMED Pedar system was used to measure plantar pressure during repeated trials of comfortable cadence walking and quiet standing. Reports of subjective pain were recorded for each orthosis as were orthosis preferences. RESULTS: All orthoses significantly reduced pressure beneath the first and second metatarsal heads compared to the shoes only control. The custom moulded orthosis with metatarsal dome was the most effective orthosis for reducing subjective ratings of pain. A significant correlation (r=0.562) was found between ratings of pain and average pressure beneath the second metatarsal head. CONCLUSIONS: Results from this study suggest that average pressure measurement may be a useful indicator in the management of metatarsalgia in RA. Further study is required to improve understanding of the relationship between rheumatoid foot mechanics and pain. RELEVANCE: Appropriate foot orthosis design can substantially improve comfort in RA patients with symptomatic feet. A custom moulded foot orthosis incorporating a metatarsal dome was the most effective design for subjects with painful second metatarsal heads. Foot pressure measurement technology can be a useful adjunct to research and clinical management of the painful rheumatoid foot. PMID- 10521641 TI - novel Award Second Prize Paper. Functional monitoring during rehabilitation following anterior cruciate ligament reconstruction. AB - OBJECTIVE: It was hypothesized that testing of ambulatory function and more demanding activities were more appropriate predictors of dynamic knee function before and after reconstruction of the anterior cruciate ligament than conventional measures of functional evaluation. It was assumed that assessment of dynamic plantar pressure distribution would represent a practical tool for guidance of the rehabilitation process after anterior cruciate ligament reconstruction. DESIGN: In a prospective study, 10 patients with isolated anterior cruciate deficiency were examined before and after replacement of the anterior cruciate (6, 12, 24 weeks) in a standardized technique. BACKGROUND: Today, functional assessment following anterior cruciate ligament reconstruction relies on clinical examination supplemented by instrumented testing of knee laxity and on isokinetic evaluation of muscle performance. Gait analysis has not been used as a quantitative measure of rehabilitation progress after surgery. METHODS: All patients were subjected to the same physiotherapy protocol. The clinical results were documented using the International Knee Documentation Committee (IKDC) protocol and the degree of knee laxity by an instrumented anterior drawer test. Muscular performance was evaluated by isokinetic testing. Dynamic pedography (EMED-SF 4) was performed to compare the non-injured and the operated leg during level walking and while descending stairs. RESULTS: Gait performance six weeks after surgery tended to be inferior to preoperative and late postoperative values. While the slight increase of maximum knee extensor torque in the operated leg and the improvement of the IKDC score during the rehabilitation period were not statistically significant, a significantly decreased gait asymmetry could be observed 12 weeks after surgery. The descending stairs test revealed functional deficits better than level walking. The latter test exhibited a strong correlation with the preoperative IKDC level and the maximum knee extensor deficit at 60 degrees /s. CONCLUSIONS: Dynamic pedography during level walking and while descending stairs is a valuable tool for monitoring the rehabilitation process after anterior cruciate ligament reconstruction. RELEVANCE: Due to the better resolution of functional deficits compared with indirect measures of function (isokinetic testing) assessment of the plantar pressure distribution may provide a more individualized adaptation for the rehabilitation program. PMID- 10521642 TI - novel Award Third Prize Paper. Assessment of the horizontal,fore-aft component of the ground reaction force from insole pressure patterns by using artificial neural networks. AB - OBJECTIVE: In this study it was investigated whether an artificial neural network can be used to determine the horizontal, fore-aft component of the ground reaction force from insole pressure patterns. DESIGN: An artificial neural network was applied to map insole pressures and ground reaction forces. METHOD: To train an artificial neural network insole pressure patterns and ground reaction force data were simultaneously determined for a wide range of different speeds (0.9-2.3 m s(-1)) for five subjects. Both intrasubject and intersubject generalizability were evaluated. RESULTS: At the intrasubject level generalizability was good when the speed for which the force was to be predicted was within the range of speeds from which data were used to train the network. Besides in some cases, generalizability to a condition outside the range of training conditions could be demonstrated. At the intersubject level the quality of generalization differed widely over subjects, from poor to good. CONCLUSIONS: It was found that an artificial neural network is able to map the relationship between insole pressure patterns and the fore-aft component of the ground reaction force. RELEVANCE: Good intrasubject generalization of 'knowledge' obtained by an artificial neural network will allow the assessment of the fore aft component of ground reaction force in condition that cannot be evaluated with force plates, e.g. activities of daily living or real sport situations. Additionally, intersubject generalization will allow shear-force recordings in subjects that are not able to complete a great number of runs to acquire enough force-plate hits. PMID- 10521643 TI - Biomechanics of the knee: methodological considerations in the in vivo kinematic analysis of the tibiofemoral and patellofemoral joint. AB - The purpose of this review article is twofold: to report on the use of intracortical pins to measure three-dimensional tibiofemoral and patellofemoral joint kinematics and highlight methodological concerns associated with this procedure. Tibiofemoral and patellofemoral kinematics has been extensively investigated using reflective markers attached to the surrounding soft tissue of the calf and thigh. However, surface markers may not adequately represent true anatomical locations and skin movement artefacts present the most critical source of measurement error. Consequently, knowledge about skeletal tibiofemoral kinematics is limited, in particular abduction-adduction and internal-external rotations. Considerable questions remain regarding what constitutes normal motion of the knee. A way to avoid the problem of surface markers is use invasive markers to directly measure skeletal motion. To date, many co-ordinate systems have been used to describe three-dimensional skeletal kinematics of the lower limb in vivo. They include helical axes, finite helical axes, instantaneous helical axes, and the joint co-ordinate system based on local anatomic landmarks. Although each method accurately describes the relative motion in 6 d. of f., the differences in how the motion is partitioned may account for the differences across investigations. Additionally, the problem of defining the anatomical co ordinate system makes comparisons across subjects and studies difficult since subtle differences may be caused by small deviations in the anatomical reference alignment. Cross talk is also a primarily a concern. For joints that articulate principally about one axis, the primary flexion/extension that is registered will be cross-talked into ab/adduction and internal/external rotations. PMID- 10521644 TI - A comparison of two motion analysis devices used in the measurement of lumbar spinal mobility. AB - OBJECTIVE. The aim of the study was to compare lumbar range of motion determined using two computerised dynamic motion analysis devices. BACKGROUND: Measures of spinal motion are currently used in biomechanical, epidemiological and clinical studies of the low back. It is essential that the various devices used to measure mobility yield similar results, particularly when the absolute values are to be used to assess job suitability, the extent of injury or the need for rehabilitation. METHODS: Eleven volunteers took part. The ranges of lumbar flexion, extension, lateral bending and axial rotation were measured using the CA6000 Spine Motion Analyser and the Polhemus Fastrak system, using standardised protocols. RESULTS: Each device showed good test-retest reliability in itself (R0.82). The absolute values for range of flexion in a standing posture were significantly higher with the CA6000 than with the Fastrak (though well correlated); those recorded in sitting were comparable for the two devices. Values for lateral bending using the two devices were well correlated, although small (but significant) differences in the absolute values were found. For extension and axial rotation, the devices gave significantly different values that were also poorly correlated. The 'limits of agreement' for the two devices (calculated to examine whether they could be used interchangeably) were rather wide, especially for extension and axial rotation. CONCLUSION: The two devices do not always yield comparable measures for spinal mobility. The accuracy of each, in relation to true angular movements of the vertebrae, remains unknown. RELEVANCE: The two computerised motion analysis devices can each be used reliably in longitudinal studies. However, if 'normal' values for spinal mobility are to be established, they must be considered device-specific. PMID- 10521645 TI - Patterns of stiffness during clinical examination of the glenohumeral joint. AB - OBJECTIVE: The purpose of our study was to develop a quantitative technique for performing clinical laxity tests, and to characterize the force-displacement response patterns in normal shoulders during two commonly applied clinical tests in different arm positions. DESIGN: The study was an in vivo clinical experiment.Background. We developed a method to objectively quantify the effects of clinical laxity testing at the shoulder. No previous studies have measured the applied force during clinical testing along with the displacement so that glenohumeral joint stiffness could be determined in vivo. METHODS: Manually applied forces were measured and shoulder displacement was recorded using electromagnetic tracking sensors during clinical stability testing in 21 subjects with normal shoulders. End-range stiffness was calculated and compared across all test conditions using repeated measures analysis of variance. RESULTS: The maximum force applied by the examiner to reach clinical end-point across all tests ranged from 101-113 N The stiffest position for posterior drawer test was at 180 degrees of abduction with the arm in internal rotation. This position was the most compliant position for the anterior drawer test. Only by internally rotating the arm did the force-displacement pattern change significantly (P<0.05). For anterior drawer tests, the patterns changed significantly (P<0.05) only when the arm was in external rotation. CONCLUSIONS: Results showed that end range stiffness was predominantly dependent on humeral rotation angle and not effected by arm abduction angle for the three angles studied. Stiffness from anterior drawer tests was highest with the arm in external rotation, and stiffness from posterior drawer tests was highest with the arm in internal rotation. RELEVANCE: This study has several clinically relevant implications for quantification of a clinical shoulder examination, and as a valuable teaching tool. Our findings also question some accepted notions about clinical "closed packed" joint positions. The study provides normal patterns of force-displacement and normative stiffness values that can be compared to patients with shoulder pathology for similar testing. PMID- 10521646 TI - Identification of optimal strategies for increasing whole arm strength using Karush-Kuhn-Tucker multipliers. AB - OBJECTIVE: The purpose of this study was to develop a computer model for identifying muscles critical to improving functional upper extremity strength. DESIGN: A three-dimensional biomechanical model of the upper extremity was developed, and the predictions were compared to maximal arm strength data collected from healthy volunteers. BACKGROUND: Although several optimization based mathematical models of the shoulder have been developed, none have utilized the mathematical properties of the Karush-Kuhn-Tucker multipliers to efficiently estimate the effect of strengthening individual muscles on functional strength of the whole arm. METHODS: A static three-dimensional biomechanical model of the glenohumeral, radio-humeral, ulno-humeral and wrist joints was developed for predicting maximal hand exertion forces. The model was formulated as a linear program. Constraints consisted of moment equilibrium conditions and limits on maximum and minimum allowable muscle forces. Predicted arm strengths were compared to maximal pull strength measurements made on 10 subjects (5 male; 5 female). The task involved pulling toward the mid-sagittal plane of the body with the arm flexed 45 degrees. The Karush-Kuhn-Tucker variables associated with the maximal limits on muscle force were computed to estimate the effect of altering the strength of individual muscles on functional arm strength. RESULTS: Maximum pull strengths were predicted well by the model. Karush-Kuhn-Tucker values ranged from 0 (for muscles not at their upper force limits) to 0.11 for the flexor carpi radialis and pectoralis major muscles. Karush-Kuhn-Tucker multipliers were found to be insensitive to the assumed specific tension of muscle. CONCLUSIONS: Upper extremity strength can be predicted from musculoskeletal geometry and physiology using linear programming. RELEVANCE: Karush-Kuhn-Tucker multipliers associated with the muscle force upper limits give insight into the effect of strengthening individual muscles on whole arm exertion strength. Such an analysis may provide insight into the development of optimal rehabilitation protocols. PMID- 10521647 TI - Changes in geometry of the finger flexor tendons in the carpal tunnel with wrist posture and tendon load: an MRI study on normal wrists. AB - OBJECTIVES: (1) To develop a methodology to determine the trajectories of the digital flexor tendons using MRI. (2) To examine changes in tendon trajectories due to wrist posture, with and without pinch force. (3) To calculate the radius of curvature of the flexor tendons and note implications for contact forces on the median nerve. (4) To assess the use of Landsmeer's models at the wrist. DESIGN: Finger flexor tendon centroids were digitized from magnetic resonance images of the carpal tunnel and the tendon paths were determined analytically. Radii of curvature were calculated from the tendon paths. BACKGROUND: Landsmeer's models of joint-tendon interaction (Landsmeer, 1961) have been used to determine moment arms and radius of curvature of the tendon paths about articulations. An explanation for a biomechanical cause of work-related carpal tunnel syndrome originated from these models. METHODS: Three healthy male participants had their right wrist scanned while splinted in four wrist postures (flexed to 20 degrees, 45 degrees, neutral, extended to 20 degrees ) with and without maintaining a 10 N pinch grip. 20-24 cross-sectional images were used for each condition. RESULTS: Volar movement of the tendons was seen with wrist flexion and the opposite was true with extension. Tendon intersection angles were calculated between the tendon as it entered the carpal tunnel and as it exited the tunnel and were 50 65% of the wrist angle (R(2)=0.81-0.96). The radius of curvature was smallest (mean=82-127 mm) with an active pinch grip with the wrist splinted at 45 degrees of flexion (mean actual wrist angle 37 degrees ). CONCLUSIONS: The radius of flexor tendon curvature is not constant as previously assumed and is larger than previous estimates. The addition of tendon force with the wrist flexed acts to reduce the radius of curvature which further increases the contact stress on the median nerve and other wrist structures. The use of MRI to determine the tendon paths has provided new insight into the relationships between the finger flexor tendons and other structures at the wrist. RELEVANCE: These findings provide data for biomechanical models of the carpal tunnel and predict the possible pathophysiology of work-related carpal tunnel syndrome. PMID- 10521648 TI - Measurement of external three-dimensional interphalangeal loads applied during activities of daily living. AB - OBJECTIVE: To measure the external three-dimensional loads applied to the interphalangeal joints during activities of daily living. DESIGN: A six-degree-of freedom force transducer was used in conjunction with motion analysis studies. BACKGROUND: There is a lack of accurate three-dimensional load data available for input into biomechanical models of the hand. METHODS: A new force transducer has been incorporated into several housings representing objects in domestic use: a jar, a tap, a key in a lock and a jug kettle. Three-dimensional kinematic data were acquired using a six-camera VICON motion analysis system. Twelve healthy volunteers took part in the study, which compared power and precision grips in 'opening' and 'closing' activities. RESULTS: Large external forces and moments are applied to the middle and distal phalanges in sagittal, coronal and axial directions. Average inter-segmental forces of up to 25 N and average moments of up to 1.8 Nm are experienced at the proximal interphalangeal joint. CONCLUSIONS: The results show that complex loading patterns are associated with routine activities of daily living. RELEVANCE: Biomechanical models of the interphalangeal joints are limited in their ability to accurately predict tendon and joint forces by the quality of the input data obtained by conventional measurement techniques. Models have tended to rely on hypothetical values of external forces acting on the hand and are over-simplified or limited to two dimensions. The results from the current study challenge the validity of these simplified models and offer a more complete picture of the complex loading system applied to the finger during daily life. PMID- 10521649 TI - Passive dynamics of the knee joint in healthy children and children affected by spastic paresis. AB - OBJECTIVE: The purposes of this study were (1) to evaluate how changes in biomechanical parameters affect segment dynamics in children and (2) to determine whether the biomechanical parameters were changed in children with spastic paresis. DESIGN: In vivo measurements were collected of knee viscoelastic properties. Background. It is unknown if the inertial and viscoelastic properties of a human growing limb should be considered in motor performance. Also unclear are whether changes in passive dynamics might be responsible for abnormal control in human spastic paresis. METHOD: Small oscillation techniques were used to measure moment of inertia of lower leg, stiffness and viscous damping of the knee joint. Eighty seven healthy children and 32 children with spastic paresis participated. RESULTS: Moment of inertia, stiffness and the damping changed with the fifth power of child's height. Dynamic equation of motion parameters were a constant, independent of the child's height. Passive viscoelastic parameters were not changed in spastic patients. CONCLUSIONS: Inertial and viscoelastic properties of a growing limb segment should not be considered in motor performance. Passive viscoelastic properties were not changed in patients with spastic paresis and, therefore, cannot be responsible for abnormal control in human spastic paresis. RELEVANCE: There is no need to adapt control patterns in children (ages 6-18). Passive viscoelastic parameters cannot be used as a descriptor of spasticity. PMID- 10521650 TI - Changes in the tibialis anterior tendon moment arm from rest to maximum isometric dorsiflexion: in vivo observations in man. AB - OBJECTIVE: In the present study, we examined the hypothesis that the tibialis anterior tendon moment arm increases during maximum isometric dorsiflexion as compared with rest. BACKGROUND: In musculoskeletal modelling applications, moment arms from passive muscles at rest are assumed representative of those measured during isometric muscle contraction. The validity of this assumption is questionable in musculotendon actuators enclosed by retinacular systems as in tibialis anterior. DESIGN AND METHODS: Sagittal-plane magnetic resonance images of the right ankle were taken in six subjects at rest and during maximum isometric dorsiflexion at six ankle angles between dorsiflexion and plantarflexion having the body placed in the supine position and the knee flexed at 90 degrees. Instant centres of rotation in the tibio-talar joint, tibialis anterior tendon action lines and moment arms were identified in the sagittal plane at ankle angles of -15 degrees, 0 degrees,+15 degrees and +30 degrees at rest and during maximum isometric dorsiflexion. RESULTS: At any given ankle angle, the tibialis anterior tendon moment arm during maximum isometric dorsiflexion increased by 0.9-1.5 cm (P<0.01) compared with rest. This was attributed to a displacement of both tibialis anterior tendon action line by 0.8 1.2 cm (P<0.01) and all instant centres of rotation by 0.3-0.4 cm (P<0. 01) distally in relation to their corresponding resting positions. CONCLUSIONS AND IMPLICATIONS: The assumption that the tibialis anterior tendon moment arm does not change from rest to maximum isometric dorsiflexion is invalid. Erroneous tendon forces, muscle stresses and joint moments by as much as 30% would be calculated using resting tibialis anterior tendon moment arms in the moment equilibrium equation around the ankle joint during maximum isometric dorsiflexion. RELEVANCE: A substantial increase in the tibialis anterior tendon moment arm occurs from rest to maximum isometric dorsiflexion. This needs to be taken into consideration when using planimetric musculoskeletal modelling for analysing maximal static ankle dorsiflexion loads. PMID- 10521651 TI - Quantifying a relationship between tactile and vibration sensitivity of the human foot with plantar pressure distributions during gait. AB - OBJECTIVE: To quantify the relationship between the tactile and vibration sensitivity thresholds of the sole of the human foot with plantar pressure distribution while walking and running. DESIGN: Cross-sectional study performed in a laboratory setting. BACKGROUND: Results of previous studies of human locomotion have identified potentially dangerous variations in locomotion patterns. A common approach to manage these variations is with the use of orthotics. Individual responses to differences in the construction and shape of orthotics cannot be fully explained with a mechanical model. It has been suggested that sensory feedback from the receptors in the feet may play an important role in regulating gait patterns. METHODS: Fifteen subjects were recruited for this study. Pressure (tactile) and vibration thresholds were determined from each subject. Plantar pressure distributions were obtained while walking at 1.5 m s(-1) and running at 3.5 m s(-1). Sensitivity measurements were correlated to pressure measurements under the foot. RESULTS: Significant negative correlation exists between the vibration threshold of the hallux at 125 Hz and peak pressures under the hallux while walking (P=0.02) and running (P=0.01). A significant negative relationship was shown between the foot mean vibration threshold at 125 Hz with peak force during running (P=0.038). A similar trend was seen at the heel, lateral arch and first metatarsal head. CONCLUSIONS: The results from this study support recent hypotheses that suggest that the body can detect and respond to external stimuli. The relationship between plantar sensitivity and peak pressures at the hallux, and the relationship between sensitivity to higher frequency vibrations and peak force during running suggests that neurological feedback should be incorporated in to any model that attempts to explain gait patterns. RELEVANCE: It is suggested that the body is able to detect small biomechanical changes in the external environment and alter gait patterns as a defensive mechanism. Understanding the relationship between neural feedback and gait patterns will help in the development of criteria for the proper application of inserts, and the prevention of lower extremity injuries. PMID- 10521652 TI - Nonlinear analysis of cartilage in unconfined ramp compression using a fibril reinforced poroelastic model. AB - OBJECTIVE: To develop a biomechanical model for cartilage which is capable of capturing experimentally observed nonlinear behaviours of cartilage and to investigate effects of collagen fibril reinforcement in cartilage. DESIGN: A sequence of 10 or 20 steps of ramp compression/relaxation applied to cartilage disks in uniaxial unconfined geometry is simulated for comparison with experimental data. BACKGROUND: Mechanical behaviours of cartilage, such as the compression-offset dependent stiffening of the transient response and the strong relaxation component, have been previously difficult to describe using the biphasic model in unconfined compression. METHODS: Cartilage is modelled as a fluid-saturated solid reinforced by an elastic fibrillar network. The latter, mainly representing collagen fibrils, is considered as a distinct constituent embedded in a biphasic component made up mainly of proteoglycan macromolecules and a fluid carrying mobile ions. The Young's modulus of the fibrillar network is taken to vary linearly with its tensile strain but to be zero for compression. Numerical computations are carried out using a finite element procedure, for which the fibrillar network is discretized into a system of spring elements. RESULTS: The nonlinear fibril reinforced poroelastic model is capable of describing the strong relaxation behaviour and compression-offset dependent stiffening of cartilage in unconfined compression. Computational results are also presented to demonstrate unique features of the model, e.g. the matrix stress in the radial direction is changed from tensile to compressive due to presence of distinct fibrils in the model. RELEVANCE: Experimentally observed nonlinear behaviours of cartilage are successfully simulated, and the roles of collagen fibrils are distinguished by using the proposed model. Thus this study may lead to a better understanding of physiological responses of individual constituents of cartilage to external loads, and of the roles of mechanical loading in cartilage remodelling and pathology. PMID- 10521653 TI - Regulation of E2F: a family of transcription factors involved in proliferation control. AB - Members of the E2F family of transcription factors are key participants in orchestration of the cell cycle, cell growth arrest and apoptosis. Therefore, an understanding of the regulation of E2F activity is essential for an understanding of the control of cellular proliferation. E2F activity is regulated by the retinoblastoma family of tumor suppressors and by multiple other mechanisms. This review will describe our current knowledge of these mechanisms which together constitute a highly complex network by which the cell cycle and cellular proliferation can be controlled. PMID- 10521654 TI - Ribozymes and the anti-gene therapy: how a catalytic RNA can be used to inhibit gene function. AB - Ribozymes are RNA molecules that possess the dual properties of RNA sequence specific recognition and site-specific cleavage of other RNA molecules. These properties provide powerful tools for studies requiring gene inhibition, when the DNA sequence is known. The use of these molecules goes beyond basic research, with a potential impact in therapeutical practice in medicine in the near future. In this review, we briefly describe the progress towards developing this class of molecules and its applications for the control of gene expression. PMID- 10521656 TI - The genes responsible for O-antigen synthesis of vibrio cholerae O139 are closely related to those of vibrio cholerae O22. AB - Several studies have shown that the emergence of the O139 serogroup of Vibrio cholerae is a result of horizontal gene transfer of a fragment of DNA from a serogroup other than O1 into the region responsible for O-antigen biosynthesis of the seventh pandemic V. cholerae O1 biotype El Tor strain. In this study, we show that the gene cluster responsible for O-antigen biosynthesis of the O139 serogroup of V. cholerae is closely related to those of O22. When DNA fragments derived from O139 O-antigen biosynthesis gene region were used as probes, the entire O139 O-antigen biosynthesis gene region could be divided into five classes, designated as I-V based on the reactivity pattern of the probes against reference strains of V. cholerae representing serogroups O1-O193. Class IV was specific to O139 serogroup, while classes I-III and class V were homologous to varying extents to some of the non-O1, non-O139 serogroups. Interestingly, the regions other than class IV were also conserved in the O22 serogroup. Long and accurate PCR was employed to determine if a simple deletion or substitution was involved to account for the difference in class IV between O139 and O22. A product of approx. 15kb was amplified when O139 DNA was used as the template, while a product of approx. 12.5kb was amplified when O22 DNA was used as the template, indicating that substitution but not deletion could account for the difference in the region between O22 and O139 serogroups. In order to precisely compare between the genes responsible for O-antigen biosynthesis of O139 and O22, the region responsible for O-antigen biosynthesis of O22 serogroup was cloned and analyzed. In concurrence with the results of the hybridization test, all regions were well conserved in O22 and O139 serogroups, although wbfA and the five or six genes comprising class IV in O22 and O139 serogroups, respectively, were exceptions. Again the genes in class IV in O22 were confirmed to be specific to O22 among the 155 'O' serogroups of V. cholerae. These data suggest that the gene clusters responsible for O139 O-antigen biosynthesis are most similar to those of O22 and genes within class IV of O139, and O22 defines the unique O antigen of O139 or O22. PMID- 10521655 TI - PAX3 gene structure, alternative splicing and evolution. AB - PAX3 is a member of the paired box family of transcription factors that function during embryogenesis and cancer epigenesis. Mutations in PAX3 cause Waardenburg syndrome (types 1 and 3), Craniofacial-deafness-hand syndrome and alveolar rhabdomyosarcoma in humans and the Splotch phenotype in mice. In this study, we describe the genomic structure of PAX3, including novel coding sequences and the complete 3' UTR. Alternative transcripts of PAX3 were identified in various tissues, including human adult skeletal muscle and mouse embryos. One of the novel alternative transcripts is evolutionarily conserved in quail and can transactivate a reporter construct containing the mouse c-met promoter. The sequences and alternative transcripts reported herein extend our understanding of the function and evolution of PAX3 in vertebrates and enable a comprehensive mutation screen for individuals with Waardenburg syndrome. PMID- 10521657 TI - pET-prof, a plasmid for high-level expression of recombinant peptides fused to a birch profilin-derived hexadecapeptide tag: a system for the detection and presentation of recombinant antigens. AB - We have previously identified a birch pollen profilin hexadecapeptide (Bp36/51), which was recognized by a monoclonal antibody (moAb 4A6) with high affinity. Here, we report the construction of a T7 RNA polymerase-driven high-level plasmid expression system, pET-prof, capable of producing proteins and peptides containing the Bp36/51 birch profilin-derived peptide fused to their N-terminus. As examples, the cDNAs coding for two major timothy grass (Phleum pratense) pollen allergens, Phl p 2 and Phl p 6, as well as for an alder (Alnus glutinosa) pollen allergen, Aln g 4, were overexpressed in Escherichia coli as BP36/51 tagged proteins. All three recombinant allergens were readily detected in nitrocellulose-blotted E. coli extracts by the Bp36/51-specific moAb 4A6. We demonstrate comparable IgE recognition of Bp36/51-tagged and untagged recombinant allergens by immunoblotting. A sandwich ELISA was developed using plate-bound moAb 4A6 to immobilize and present Bp36/51-tagged recombinant allergens to IgE antibodies of allergic patients. Using immunoelectronmicroscopy, we demonstrate that even under harsh fixation conditions, tagged allergens can be localized simultaneously in situ by moAb 4A6 and allergen-specific antisera. We suggest the use of the pET-prof system for the high-level expression of Bp36/51-tagged polypeptides that can be rapidly detected in total protein extracts, immunolocalized in situ, immobilized and presented to other antigen-specific antibodies (e.g. IgE), even when they occur in minute concentrations. PMID- 10521658 TI - Sequence and expression patterns of a human EMAP-related protein-2 (HuEMAP-2). AB - Until recently, the microtubule-associated protein, EMAP, was identified only in echinoderms such as sea urchin, starfish and sand dollar. Sea urchin EMAP localizes to the mitotic apparatus in vivo and modifies the assembly dynamics of microtubules in vitro. To identify domains important for EMAP function, we cloned and sequenced cDNAs for an EMAP-related protein in human. The nucleotide sequence of a human EMAP-related protein-2 (HuEMAP-2) encodes a protein of 649 amino acids in length. The translated polypeptide sequence and domain structure of sea urchin EMAP and HuEMAP-2 are highly conserved, with greater than 57% identity and 77% similarity at the translated amino acid level. Southern blot analysis is consistent with the presence of a single HuEMAP-2 gene in the human genome. Moreover, HuEMAP-2 is a member of a larger protein family with at least four HuEMAP sequences in the NCBI databases. One of these, HuEMAP-1, is identified as the candidate gene for the Usher syndrome 1 a locus (Genomics 43:104-106, 1997). Northern blot analysis indicates that HuEMAP-1, and HuEMAP-2 are expressed in different human tissues. In addition, these RNA blots indicate that HuEMAP-2 transcripts may be differentially spliced in neuronal tissues. PMID- 10521659 TI - Promoter analysis of the murine T-cell protein tyrosine phosphatase gene. AB - The T-cell protein tyrosine phosphatase (TC PTP) is expressed ubiquitously at all stages of mammalian development. However, mRNA levels fluctuate in a cell-cycle dependent manner, reaching peak levels in late G1, and rapidly decreasing in S phase. Furthermore, TC PTP being present in higher amounts in lymphoid tissues, we have recently shown that it is essential for proper maintenance of both the bone marrow micro-environment and B- and T-cell functions. In order to better understand the elements controlling the expression pattern of this gene, we have isolated and characterized approx. 4kb of the murine TC PTP promoter. DNA sequencing of the proximal 5' region revealed the absence of both TATAA and CAAT boxes. Primer extension analysis and S1 nuclease mapping techniques identified multiple transcription initiation sites. Functional promoter activity was determined using transfection experiments of promoter deletion constructs fused to a CAT reporter construct. Our results indicate that the minimal promoter sequence required for functional expression is contained within the first 147bp of the TC PTP promoter. In addition, consistent with the cell-cycle-dependent expression of TC PTP, we localized a domain between 492 and 1976bp from the transcription initiation site through which repression occurs. In conclusion, although initiator-driven transcription allows for ubiquitous expression of TC PTP, we define general transcription motifs present within the promoter that may mediate specific modulations of the TC PTP gene. PMID- 10521660 TI - Construction and use of low-copy number T7 expression vectors for purification of problem proteins: purification of mycobacterium tuberculosis RmlD and pseudomonas aeruginosa LasI and RhlI proteins, and functional analysis of purified RhlI. AB - Purification of proteins from Escherichia coli under native conditions is often hampered by inclusion-body formation after overexpression from T7 promoter-based expression vectors. This is probably due to the relatively high copy number of the ColE1-based expression vectors. To circumvent these problems, the low-copy number pViet and pNam expression vectors were constructed. These vectors contain the pSC101 origin of replication and allow the expression of oligohistidine and intein chitin-binding domain fusion proteins, respectively. Since pViet and pNam do not replicate in E. coli B strains, an E. coli K-12 host strain [SA1503(DE3)] was constructed. This strain is defective in the Lon and OmpT proteases and allows IPTG-inducible expression of recombinant proteins from the T7 promoter. The new vectors were successfully tested by purification of three very insoluble proteins (RmlD, LasI and RhlI) under non-denaturing conditions, and all three proteins retained enzymatic activity. The purified hexahistidine (His6)-tagged Pseudomonas aeruginosa RhlI protein was subjected to more detailed analyses, which indicated that (1) only butyryl-acyl carrier protein (ACP) and S adenosylmethionine (SAM) were required for synthesis of N-butyryl-L-homoserine lactone; (2) when present at physiological concentrations, butyryl-coenzyme A and NADPH were not substrates for RhlI; (3) RhlI was able to synthesize N-hexanoyl-L homoserine lactone from hexanoyl-ACP and SAM; (4) RhlI was able to direct synthesis of N-butyryl-L-homoserine lactone from crotonyl-ACP in a reaction coupled to purified P. aeruginosa FabI (enoyl-ACP reductase). PMID- 10521661 TI - Developmental expression of specific genes detected in high-quality cDNA libraries from single human preimplantation embryos. AB - We describe an improved highly sensitive method for generating cDNA libraries containing a high proportion of cDNAs enriched with 5'-coding sequences from single human preimplantation embryos and a 10 week old whole foetus. The embryonic mRNA was isolated using oligo-(dT) linked to magnetic beads. First strand cDNA synthesis was carried out directly on the bound mRNA, followed by PCR designed to amplify the cDNA molecules synthesized in their entirety. The complexities of the libraries are between 10(5) and 10(6) independent clones. The average cDNA size is 1.0 kb, and the size range is 0.5-3.0 kb. PCR analysis of the embryonic libraries for specific genes has revealed transcripts for genes known to be transcribed in preimplantation stages, such as the imprinted gene SNRPN, developmental genes WNT11, HOX, OCT-1 and the embryonic OCT-4, cytoskeletal genes keratin-18 and beta-actin, the cell cycle gene C-MOS, and housekeeping genes GAPDH and HPRT. Sequencing of random clones showed the presence of a variety of sequences, such as human chorionic gonadotrophin, ubiquitin, TFIIA, guanine nucleotide-binding protein (beta-subunit), annexin I, a gene encoding a kinesin-like protein, and TWIST, which encodes a basic helix-loop helix (bHLH) transcription factor implicated in Saethre-Chotzen syndrome (characterized by craniofacial and limb anomalies). Approximately 40% of these randomly analysed clones were full length. In addition to cDNAs matching known ESTs (Expressed Sequence Tags) in the GenBank and dbEST databases, novel sequences were detected at a frequency of 16% of randomly picked clones. The libraries are a valuable resource, providing longer cDNAs representing genes expressed during human preimplantation development. PMID- 10521662 TI - A complex population of RNAs exists in human ejaculate spermatozoa: implications for understanding molecular aspects of spermiogenesis. AB - The presence of mRNAs in human ejaculate spermatozoa is well established, yet little is known of the representation or function of these transcripts. To address these issues, the complexity of spermatozoal RNA was examined. As expected, testis-expressed mRNAs were detected by RT-PCR in mature human spermatozoa. Interestingly, when a testis cDNA library was probed with total spermatozoal RNA, less than 2% of plaques gave a strong hybridization signal, suggesting a rather unique sperm-derived population. To further define the sequence distribution, 18 strongly hybridizing clones were selected at random for end-sequence analysis. Twelve matched unique sequences in the EST, STS and NR databases, whereas five showed no similarity to any of the sequences in the databases. In addition, one clone belonged to the SINE repetitive element family. As demonstrated by sequencing randomly primed cloned inserts, short (SINE/MER) or long (LINE/ORF2) interspersed repeat-like sequences are also contained as part of the spermatozoal RNA fraction. It is now evident that human spermatozoa contain a rich repertoire of both known and unknown protein-encoding and non-coding RNAs. This provides a unique opportunity to identify and investigate the many genes responsible for the structure and function/dysfunction of the male gamete using spermatozoal RNA as the template. PMID- 10521663 TI - Cloning, structural characterization, and chromosomal localization of the gene encoding the human prostaglandin E(2) receptor EP2 subtype. AB - Northern blot analysis of human placental RNA using a probe to the 5' end of the human prostaglandin E(2) (PGE(2)) EP2 receptor subtype coding region revealed the existence of a high abundance, low molecular weight transcript. To investigate the origin of this transcript, and its possible relationship to the human EP2 mRNA, we have cloned and characterized the gene encoding the human PGE(2) EP2 receptor subtype, identified transcriptional initiation and termination sites in two tissues (spleen and thymus), and determined its chromosomal localization. The human EP2 gene consists of two exons separated by a large intron, utilizes a common initiation site in both spleen and thymus at 1113 bp upstream of the translation initiation site, and has 3' transcript termini at 1140 bp and 1149 bp downstream of the translation stop site in spleen and thymus respectively. Southern and fluorescence in situ hybridization analysis demonstrated the human EP2 gene to be a single copy gene located in band 22 of the long arm of chromosome 14 (14q22). Though our initial interest in this gene was to investigate potential differential splicing of the human EP2 gene in placenta, this work demonstrates that the atypical transcript observed in placenta probably arises from a distinct, yet related, gene. Knowledge of the sequence, structure, and transcription events associated with the human EP2 gene will enable a broader understanding of its regulation and potential role in normal physiology and disease. PMID- 10521664 TI - Zwittermicin A biosynthetic cluster. AB - The goal of this study was to identify the biosynthetic cluster for zwittermicin A, a novel, broad spectrum, aminopolyol antibiotic produced by Bacillus cereus. The nucleotide sequence of 2.7kb of DNA flanking the zwittermicin A self resistance gene, zmaR, from B. cereus UW85 revealed three open reading frames (ORFs). Of these ORFs, two had sequence similarity to acyl-CoA dehydrogenases and polyketide synthases, respectively. Insertional inactivation demonstrated that orf2 is necessary for zwittermicin A production and that zmaR is necessary for high-level resistance to zwittermicin A but is not required for zwittermicin A production. Expression of ZmaR was temporally associated with zwittermicin A production. The results suggest that zmaR is part of a cluster of genes that is involved in zwittermicin A biosynthesis, representing the first biosynthetic pathway for an aminopolyol antibiotic. PMID- 10521665 TI - Sequence analysis and expression of the aspartokinase and aspartate semialdehyde dehydrogenase operon from rifamycin SV-producing amycolatopsis mediterranei. AB - A approximately 4.8 kb KpnI fragment, from the upstream region of the methylmalonyl-CoA mutase gene (mutAB) of rifamycin SV-producing Amycolatopsis mediterranei, was cloned and partially sequenced. Codon preference analysis showed three complete ORFs. ORF2 is internal to ORF1, and encodes a polypeptide corresponding to 172 amino acids, whereas ORF1 encodes a polypeptide of 421 amino acids. They were identified as the encoding genes of aspartokinase alpha- and beta-subunits by comparing the amino acid sequences with those in the database. The downstream ORF3, whose start codon was overlapped with the stop codon of both ORF1 and ORF2 by 1 bp, was identified as the aspartate semialdehyde dehydrogenase gene (asd), encoding a polypeptide of 346 amino acids. Subclones containing either the ask gene or the asd gene were constructed, in which the genes could be expressed under Lac promoters. Two subclones could transform E. coli CGSC 5074 (ask-) and E. coli X6118 (asd-) to prototrophy, supporting the functional assignments. Southern hybridisation indicated that the approximately 4.8 kb sequenced region represented a continuous segment in the A. mediterranei chromosome. It is concluded that ask and asd genes are present in an operon in A. mediterranei, and therefore that organisation of these two genes is the same as in most gram-positive bacteria, such as Mycobacteria, Corynebacterium glutamicum and Bacillus subtilis, but is different from Streptomyces akiyoshiensis. PMID- 10521666 TI - SIN, a novel Drosophila protein that associates with the RNA binding protein sex lethal. AB - Sex-lethal (SXL) is an RNA binding protein that acts as a regulator of both alternative pre-mRNA splicing and translation. Because SXL must sometimes function at some distance from its binding sites, it is believed that it must interact with other proteins. We used a yeast two-hybrid screen to isolate a novel Drosophila protein, SIN (SXL interactor), that interacts specifically with SXL. A direct physical association was demonstrated in vitro, and a single SXL RNA binding domain was sufficient for the interaction. SIN shows a high degree of similarity to a mammalian protein of unknown function. The cytogenetic location of Sin is 78A2-4. The transcript, which is abundant in early embryos, appears to be of maternal origin. PMID- 10521667 TI - Target organ and time-course in the mutagenicity of five carcinogens in MutaMouse: a summary report of the second collaborative study of the transgenic mouse mutation assay by JEMS/MMS. AB - We studied five carcinogens for (a) organ-specific mutagenicity and expression time in the transgenic (TG) mouse mutation assay and (b) clastogenicity in the peripheral blood micronucleus assay in the same mice. Groups of mice were injected intraperitoneally (ip) with N-nitroso-di-n-propylamine (NDPA), propylnitrosourea (PNU), 7, 12-dimethylbenz[a]anthracene (DMBA), 4-nitroquinoline 1-oxide (4NQO), or procarbazine (PCZ); 4NQO was also administered orally. LacZ mutant frequencies (MF) of various organs, sampled 7, 14 and 28 days after treatment, were analyzed by galE positive selection. At least 5 organs were analyzed in each experiment. Bone marrow, liver, and testis were always analyzed, as were each chemical's target organs. All chemicals, except NDPA, induced micronuclei. All chemicals increased lacZ MF in all of their target organs for carcinogenesis and, to a lesser extent, in some non-target organs. That suggests that an organ that has a positive response to a chemical in the TG mouse mutation assay is likely to develop tumors on exposure to that chemical, but it does not always happen. The time-course of MF increases (7-28 days) differed among tissues. In general, time-dependent increase in MF occurred in organs with a low cell proliferation rate whereas no increase, or even a decrease, occurred in organs with a high proliferation rate. Our results demonstrated that the TG mouse mutation assay is effective for the detection of chemical mutagenesis in the target organs for carcinogenesis, and organ and time-course variations in chemical mutagenesis are important issues for the establishment of an optimal protocol for the assay. PMID- 10521668 TI - Procarbazine genotoxicity in the MutaMouse; strong clastogenicity and organ specific induction of lacZ mutations. AB - Procarbazine, a drug used for cancer chemotherapy, is carcinogenic in rodent bioassays. We analyzed the mutagenicity of procarbazine in various organs and the clastogenicity of the drug in hematopoietic cells of the lacZ transgenic MutaMouse. This was part of the second collaborative study of the Mammalian Mutagenesis Study Group of the Japanese Environmental Mutagen Society on the transgenic mouse mutation assay. At 50 mg kg(-1), procarbazine induced micronuclei in hematopoietic cells, but it did not increase the lacZ mutant frequency (MF) in bone marrow. It was also negative in liver, testis, spleen, kidney, and lung. Five daily administrations of 150 mg kg(-1) yielded highly positive responses in the drug's target organs for carcinogenesis (lung, bone marrow, and spleen). Lower positive responses were detected in kidney, which is a minor target organ. Liver showed only a slight increase in lacZ MF and brain showed no increase. The testis MF more than doubled which suggest that procarbazine is mutagenic to germ cells. Thus, we demonstrated that procarbazine has a strong clastogenic effect in hematopoietic cells and is mutagenic in a variety organs after high dose treatment. The induced MF was especially high in procarbazine's target organs for carcinogenesis, which supports the relevance of the transgenic mouse mutation assay for the assessment of potential genotoxins in vivo. PMID- 10521669 TI - Induction of lacZ mutation by 7,12-dimethylbenz[a]anthracene in various tissues of transgenic mice. AB - The induction of gene mutations was examined in MutaMouse after an intraperitoneal injection of 7, 8-dimethylbenz[a]anthracene (DMBA) at 20 mg/kg in a collaborative study participated by four laboratories. Although the DMBA dose used was lower than the level that has been reported to induce micronucleated erythrocytes maximally in several mouse strains, a killing effect appeared after day 9 of the post-treatment interval. Mutations in lacZ transgene were detected by the positive selection assay following in vitro packaging of phage lambda from the genomic DNA of the transgenic animals that survived. The mutant induction was evaluated in the bone marrow, liver, skin, colon, kidney, thymus, and testis 7 to 28 days after the treatment. In the bone marrow, the mutant frequency reached a maximum, approximately a 30-fold increase, 14 days after the treatment and the increased frequency persisted at least up to day 28 of the post-treatment. Induction of mutants was detected in the liver, colon, thymus, and skin to lesser extents. Marginal responses were obtained in the kidney and testis. The slight increases in the mutant frequencies in the kidney and testis observed in some laboratories were within laboratory-to-laboratory or animal-to-animal variations. In contrast to the gene mutation induction in the bone marrow, the frequency of micronucleated reticulocytes increased transiently 3 days after the treatment and returned to a control level before day 8 of the post-treatment. It was suggested that DMBA induced gene mutation is fixed in stem cells depending on cell proliferation while DNA damages responsible for chromosome breakage are not transmitted to progeny cells. PMID- 10521670 TI - Mutation induction by N-propyl-N-nitrosourea in eight MutaMouse organs. AB - As a part of the 2nd Collaborative Study for the Transgenic Mouse Mutation Assay, we studied the organ specificity and the temporal changes in mutant frequency (MF) of the lacZ gene following intraperitoneal injection of 250 mg/kg N-propyl-N nitrosourea into male MutaMouse. We used a positive selection system and examined eight organs, i.e., bone marrow, liver, kidney, lung, spleen, brain, heart, and testis. The chemical caused a significant increase in MF in all organs except for brain, and the bone marrow was the most sensitive organ, exhibiting a MF on day 7 that was 10 times that of the control. The MF increased from day 7 to day 28 in liver, kidney, and testis, while it decreased in bone marrow. The relationship between the results of this study and the target organs of carcinogenesis, and the cause of the temporal changes in MF, are discussed. PMID- 10521671 TI - N-Nitrosodi-n-propylamine induces organ specific mutagenesis with specific expression times in lacZ transgenic mice. AB - The mutagenic and clastogenic effects of N-nitrosodi-n-propylamine (NDPA) in lacZ transgenic mice (MutaMouse) were investigated as a part of the second collaborative study of the transgenic mouse mutation assay by a subgroup of the Mammalian Mutagenesis Study Group, a suborganization of the Environmental Mutagen Society of Japan. Male MutaMouse mice were administered NDPA intraperitoneally at a dose of 250 mg/kg, which is half of the LD(50) of the compound. The clastogenicity of NDPA was examined by the peripheral blood micronucleus test just before and at 24, 48 and 72 h after the treatment. The mutant frequencies in the bone marrow, liver, lung, kidney and urinary bladder were examined by the positive selection method for lacZ kidney. These findings demonstrate that NDPA induces organ-specific mutagenesis with specific expression times, and that the mutagenicity of NDPA in lacZ transgenic mice is consistent with its carcinogenicity. PMID- 10521672 TI - Mutagenicity of 4-nitroquinoline 1-oxide in the MutaMouse. AB - As part of a collaborative study, the Mammalian Mutagenesis Study Group (MMS), a sub-organization of the Environmental Mutagen Society of Japan (JEMS) conducted mutagenicity tests in MutaMouse. Using a positive selection method, we studied the organ-specificity and time dependence of mutation induction by 4 nitroquinoline 1-oxide (4NQO). A single dose of 4NQO was administered intraperitoneally (7.5 or 15 mg/kg) or orally (200 mg/kg) to groups of male mice. On days 7, 14 and 28 after treatment, we isolated the liver, kidney, lung, spleen, bone marrow, testis and stomach in the intraperitoneal administration experiment and the liver, lung, bone marrow, testis and stomach in the oral administration experiment. In addition, we performed the peripheral blood micronucleus test to evaluate clastogenicity. In the mice treated intraperitoneally at 7.5 mg/kg, we found increased mutant frequency (MF) only in the lung, where the MF did not vary with expression time. In the mice treated at 15 mg/kg, we found increased MF in the liver, bone marrow and lung. In orally treated mice, the MF was high in the lung and liver and very high in the bone marrow and stomach while the increase in the testis was negligible. As the expression time was prolonged, the MF tended to increase in the liver, decrease in the bone marrow, and remain stable in the lung, testis and stomach. The incidence of micronucleus induction in peripheral blood cells was significantly increased (p<0.01) in the 4NQO groups when compared with the vehicle control group by intraperitoneal treatment. Thus, these assay systems appeared to be of use in detecting not only genetic mutation but also chromosomal aberration. PMID- 10521673 TI - Spontaneous sister chromatid exchange and chromosome aberration frequency in humans: the familial effect. AB - The possible effects of environmental and genetic factors on spontaneous frequencies of sister chromatid exchanges (SCEs) and cells with chromosome aberrations (CAs) in human lymphocytes were investigated by analysing 177 completed families (mother, father and at least one child). After removing the effects of methodological, biological and life-style factors by the use of multifactor analysis of variance (MANOVA), SCEs and CAs residuals were analysed by simple correlation analysis and principal component analysis. SCEs and CAs inter-familiar variability was higher than that found within families. A significant correlation was found between the average SCE frequencies shared by parents (the so-called 'midpoint parents', or 'midparent') and offspring (linear slope b=0.26+/-0.07, p<0.05), but also between mother and father (b=0.23+/-0.11, p<0.05) suggesting the presence of an effective environmental factor. The midparent-offspring correlation was found to be sustained by the mother-offspring relationship (b=0.28+/-0.08, p<0.05), being the father-offspring correlation not significant (b=0.16+/-0.11, p0.05). Concerning CAs, no statistically significant correlation between parents was found, but the strong relationship between mother and offspring was confirmed (b=0.468+/-0.11, p<0.001). The SCEs correlation between mother vs. offspring disappeared for older offspring (over 23 years old). The obtained findings strongly showed that the genetic make-up is barely detectable in the presence of domestic environment factors which are shown to play the major role in determining the interfamilial variability of SCE and CA in a general population. These results strengthen the suitability of the use of SCEs and CAs analysis in human cytogenetic surveillance for the detection of effective environmental factors. PMID- 10521674 TI - Development of a mouse cell line containing the PhiX174 am3 allele as a target for detecting mutation. AB - Transgenic mice containing multiple copies of the PhiX174 am3 allele are being developed as a model for detecting tissue-specific in vivo mutation. In order to derive an analogous system for measuring am3 mutation in vitro, cells were cultured from 15-day-old C57Bl/6J mouse embryos that were homozygous for the transgene and these cells were transfected with a plasmid expressing the SV40 large T-antigen. Two G418-resistant colonies were isolated from this culture and expanded to continuously proliferating cell lines (PX-1 and PX-2). Line PX-2 was treated with up to 1.0 mg/ml of N-ethyl-N-nitrosourea (ENU), assayed for survival by cloning efficiency after overnight culture, and assayed for am3 mutations after 5 days of culture. Survival decreased to 31% at the highest dose of ENU, while mutant frequency increased with dose from approximately 2 x 10(-7) in the untreated cells to 13 x 10(-7) in cultures treated with 0.6 mg/ml of ENU. PX-2 cells also were treated with 0 and 0.6 mg/ml of ENU and mutant frequency assays were performed after 5, 24, 48 and 72 h of growth. The mutant frequency in the treated culture increased to 20 x 10(-7) at 48 h and remained approximately the same at 72 h. These results indicate that PX-2 cells should be a useful resource for developing the in vivo am3 mutant assay and for evaluating the sensitivity of the am3 allele to various classes of mutagens. PMID- 10521675 TI - The synergistic effects of vanillin on recombination predominate over its antimutagenic action in relation to MMC-induced lesions in somatic cells of Drosophila melanogaster. AB - The wing Somatic Mutation And Recombination Test (SMART) in Drosophila melanogaster was used to study the modulating action of vanillin (VA) in combination with the alkylating agents mitomycin C (MMC), methylmethanesulphonate (MMS) and the bifunctional nitrogen mustard (HN2). Two types of treatments with VA and each of the three genotoxins were performed: chronic co-treatments of three-day-old larvae of the standard cross as well as post-treatments after acute exposure with the genotoxins. This allowed the study of the action of VA not only in the steps that precede the induction of DNA lesions but also in the repair processes. The overall findings from the co-treatment series suggest that ingestion of VA with MMS or MMC can lead to significant protection against genotoxicity; but this is not the case with HN2. Antioxidant activity, suppression of metabolic activation or interaction with the active groups of these two alkylating agents could be mechanisms by means of which VA exerts its desmutagenic action. In contrast, when evaluated in the post-treatment procedure, VA causes two antagonistic effects on the genotoxicity of MMC: (i) synergism on recombination (172.8%) and (ii) protection against mutation (79.0%). Consequently, both activities together lead to a considerable increase in mitotic recombination. In spite of being separate events, recombination and gene mutation are correlated during mitosis since the fate of a DNA lesion depends on the repair pathway followed. Our results may suggest that VA is a modifying factor that blocks the mutagenic pathway and consequently directs the MMC-induced lesions into a recombinational repair. Furthermore, VA did not modify the genotoxicity when administered after treatments with HN2 or MMS. Therefore, the major finding of the present study, namely the co-recombinagenic activity of VA on MMC-induced lesions, seems to be related to the type of induced lesion and consequently to the repair processes involved in its correction. PMID- 10521676 TI - Radioprotective effect of melatonin assessed by measuring chromosomal damage in mitotic and meiotic cells. AB - This study was taken to evaluate the radioprotective effects of melatonin. Male adult albino mice were treated (intraperitoneal, i.p.) with 10 mg/kg melatonin either 1 h before or 1/2 h after exposure to 1.5 Gy of gamma-irradiation. Control, melatonin, irradiated and melatonin plus irradiation groups were sacrificed 24 h following treatment. The incidence of micronuclei (MN) in bone marrow cells was determined in all groups. The results show that melatonin caused a significant reduction in micronuclei polychromatic erythrocytes (MNPCE) when animals were treated with melatonin before and not after exposure to radiation. Mitotic and meiotic metaphases were prepared from spermatogonial and primary spermatocytes, respectively. Examination and analysis of metaphases showed no mutagenic effect of melatonin on chromosomal aberration (CA) frequency in spermatogonial chromosomes. Administration of one single dose of melatonin to animals before irradiation lowered total CA from 46 to 32%. However, no significant effect was observed when melatonin was given after irradiation. Similarly, the frequency of CA in meiotic metaphases decreased from 43.5% in the irradiated group to 31.5% in the irradiated group treated with melatonin 1 h before irradiation, but no change was observed when melatonin was administered after irradiation. The data obtained in this study suggest that melatonin administration confers protection against damage inflicted by radiation when given prior to exposure to irradiation and not after, and support the contention that melatonin radioprotection is achieved by its ability as a scavenger for free radicals generated by ionizing radiation. PMID- 10521677 TI - Mutagenicity monitoring of airborne particulate matter (PM10) in the Czech Republic. AB - The mutagenic activities associated with inhalable airborne particulate matter (PM10) collected over a year in four towns (Czech Republic) have been determined. The dichloromethane extracts were tested for mutagenicity using the Ames plate incorporation test and the Kado microsuspension test both with Salmonella typhimurium TA98 and its derivative YG1041 tester strains in the presence and absence of S9 mixture. The aim of this study was to assess the suitability of both bacterial mutagenicity tests and to choose the appropriate indicator strain for monitoring purposes. To elucidate the correlation between mutagenicity and polycyclic aromatic hydrocarbons (PAHs), the concentration of PAHs in the air samples were determined by GC/MS. In general, the significant mutagenicity was obtained in organic extracts of all samples, but differences according to the method and tester strain used were observed. In both mutagenicity tests, the extractable organic mass (EOM) exhibited higher mutagenicity in the YG1041 strain (up to 97 rev/microg in the plate incorporation and 568 rev/microg in the microsuspension tests) than those in TA98 (up to 2.2 rev/microg in the plate incorporation and 14.5 rev/microg in the microsuspension tests). In the plate incorporation test, the direct mutagenic activity in YG1041 was on average 60 fold higher and in microsuspension assay 45-fold higher with respect to strain TA98. In the presence of S9 mix, the mutagenic potency in YG1041 declined (P<0.001) in summer, but increased in TA98 (P<0.05) in samples collected during the winter season. The microsuspension assay provided higher mutagenic responses in both tester strains, but in both strains a significant decrease of mutagenic potency was observed in the presence of S9 mix (P<0.001 for YG1041, P<0.05 for TA98 in winter). The mutagenic potencies detected with both indicator strains correlated well (r=0.54 to 0.87) within each mutagenicity test used but not (for TA98) or moderately (r=0.44 to 0. 66 for YG1041) between both of the tests. The mutagenic activity (in rev/m(3)) likewise the concentration of benzo[a]pyrene and sum of carcinogenic PAHs showed seasonal variation with distinctly higher values during winter season. A correlation between the PAH concentrations and the mutagenicity results for the plate incorporation, but not for the microsuspension tests was found. In samples from higher industrial areas, the higher mutagenicity values were obtained in plate incorporation test with TA98 and in both tests with YG1041 in summer season (P<0.05). According to our results, plate incorporation test seems to be more informative than microsuspension assay. For routine ambient air mutagenicity monitoring, the use of YG1041 tester strain without metabolic activation and the plate incorporation test are to be recommended. PMID- 10521678 TI - Epoxide hydrolase activity in human blood mononuclear leukocytes: individual differences in native and mitogen-stimulated cells. AB - Epoxide hydrolase (EH; EC 3.3.2.3) activity was measured in whole-cell sonicates of native and cultured peripheral blood mononuclear cells (PBMCs) from 19 healthy unrelated Caucasian donors (age, 28-55 years). We used 1,2-epoxy-3-(p nitrophenoxy)propane (0. 34 mM) as the substrate and, for the diol assay, a quantitative HPLC method with spectrophotometric detection. One portion of the PBMCs was frozen immediately, while the other portion was PHA-stimulated and cultivated for 36 h. In native leukocytes, the EH activity varied from 2.2 to 8.2 pmol/min per 10(6) cells (3.8-fold), the mean+/-SD was 5.6+/-1.4 pmol/min per 10(6) cells. In most of the samples from different donors, the specific activity increased in cultivation, varying from 2.4 to 15.4 pmol/min per 10(6) cells (6. 3 fold), the mean+/-SD being 8.5+/-3.8 pmol/min per 10(6) cells. From a methodological point of view, enzyme measurement in native cells is simple to perform and may provide a better index of the specific activity, as the accuracy of electronic cell counting is better for cell samples taken before than after cultivation. The differences in the EH activity of PBMCs indicate that significant interindividual variation may occur in the detoxification of epoxides produced in the human lymphocyte test systems commonly used for genotoxicity screening of chemicals in vitro. Further studies are needed to determine the extent to which the reproducibility and thus also the sensitivity of such assays could be improved by analyses carried out to control the donors for their EH phenotype. PMID- 10521679 TI - The frequency of illegitimate TCRbeta/gamma gene recombination in human lymphocytes: influence of age, environmental exposure and cytostatic treatment, and correlation with frequencies of t(14;18) and hprt mutation. AB - Chromosome translocations in lymphoid malignancies often involve V(D)J recombinase mediated events giving rise to aberrant T-cell receptor (TCR) and immunoglobulin genes, which have been suggested to be useful as markers of genomic instability, genotoxic exposure and cancer risk. Illegitimate rearrangements involving the TCRbeta/gamma loci on chromosome 7 create TCRbeta/gamma hybrid genes which occur at low frequency in peripheral blood lymphocytes (PBLs) of normal healthy individuals. To evaluate the utility of this marker, we studied the possible effects of age and genotoxic exposures on the TCRbeta/gamma gene variant frequency (VF), and compared the frequencies of hypoxanthine guanine phosphoribosyl transferase (hprt) mutation, hprt exon 2/3 deletion, t(14;18) and TCRbeta/gamma gene rearrangements in cells from the same donors. The TCRbeta/gamma VF ranged five-fold among 16 middle aged blood donors with a mean of 0.74+/-0.29/10(5) PBLs, which is consistent with our previous estimate in healthy subjects. The TCRbeta/gamma VF was found to increase from birth until early adult life, and then to decrease with increasing age. Four testis cancer patients, who 6 years earlier had been treated with etoposide and other cytostatic drugs, showed TCRbeta/gamma VF similar to that in healthy controls. No increase of the TCRbeta/gamma VF was found among non-smoking PAH exposed aluminum smelter workers compared to non-smoking controls. Smoking smelter workers showed decreased TCRbeta/gamma VF compared to non-smoking workers and controls, but in a follow-up study 2 years later the difference was no longer statistically significant, although the smoking smelter workers still showed a lower TCRbeta/gamma VF than the controls. No correlation was obtained between the TCRbeta/gamma VF and the t(14;18) or hprt mutant frequency (MF) in a group of healthy individuals. However, there was a statistically significant correlation between the TCRbeta/gamma VF and the hprt exon 2/3 deletion frequency in PBL DNA from the same donors. These results show that the TCRbeta/gamma VF in healthy individuals changes with age and correlates with the frequency of hprt exon 2/3 deletion, another marker of aberrant V(D)J recombination in T-cells. However, no effect of smoking or present or previous exposure to genotoxic agents on TCRbeta/gamma VF was observed in this study. Thus, further studies are needed to prove the utility of TCRbeta/gamma gene rearrangement as a marker of genotoxic exposure. PMID- 10521680 TI - Impact of dimethyl sulfoxide and examples of combined genotoxicity in the SOS chromotest. AB - The bacterial SOS chromotest with Escherichia coli PQ37 was used for the assessment of genotoxicity of combined xenobiotic treatments. The modulation of test compound genotoxicity by dimethyl sulfoxide (DMSO), a common solvent for test compounds, was assessed as well. It was shown that DMSO modulated SOS chromotest genotoxicity of several xenobiotics: in comparison to test compound dissolution in water, the commonly used addition of 3.2% (v/v) DMSO as solvent lead to a significant increase in the genotoxicity of K(2)RhCl(5) and beta propiolactone (BPL). However, the effects of cisplatin decreased significantly when DMSO was added. Thus, albeit DMSO is not genotoxic in this test itself, it can interfere with SOS chromotest responses. Further experiments were performed in the absence of DMSO. BPL and cisplatin in combination showed an over-additive synergism in SOS genotoxicity as well as K(2)RhCl(5) and cisplatin did. Addition of Pd(NH(3))(4)Cl(2) and NaAsO(2), which are non-genotoxic in the SOS chromotest, did not enhance the K(2)RhCl(5)- or BPL-mediated SOS sfiA induction. Nevertheless, at the highest subcytotoxic dose of NaAsO(2) tested (200 microM), a slight yet significant suppression of BPL-mediated SOS genotoxicity was observed. These results confirm that the SOS chromotest is a useful tool for the rapid evaluation of the combined genotoxicity of compound mixtures. However, the use of DMSO as test solvent has to be taken with caution. PMID- 10521681 TI - Mutation and DNA modification in Salmonella exposed to N-nitrosodimethylamine under UVA- and sunlight-irradiation. AB - Previously, we reported that when Salmonella typhimurium and Escherichia coli were treated with N-nitrosodimethylamine (NDMA) under irradiation with ultraviolet-A (UVA), mutagenesis of the bacteria took place without externally added activation enzymes. We also observed the formation of O(6)-methylguanine (O(6)-meG), N(7)-methylguanine (N(7)-meG) and 7,8-dihydro-8-oxodeoxyguanosine (8 oxodG) in calf thymus DNA treated with NDMA plus UVA. In this study, we observed the mutagenicity of NDMA under irradiation of natural sunlight in S. typhimurium. Furthermore, we detected the formation of O(6)-meG, N(7)-meG and 8-oxodG in calf thymus DNA treated with NDMA plus simulated sunlight. Regarding the mutagenesis of S. typhimurium by NDMA plus UVA, we have now identified and quantified O(6) meG formed in the genomic DNA of the bacteria under conditions of the mutagenesis. PMID- 10521682 TI - No clastogenic activity of a senna extract in the mouse micronucleus assay. AB - In previous studies, an analytically well-defined senna extract, commonly used as a laxative, gave positive responses in vitro in the Ames test and in the CHO assay. Therefore, the objective of this study was to investigate the genotoxic activity of the same senna extract in an in vivo genotoxicity assay by means of the generally acknowledged MNT. After administration of an oral dose of 2000 mg senna extract/kg to NMRI mice of both genders, which is equivalent to 119 mg potential rhein/kg, 5.74 mg potential aloeemodin/kg and 0. 28 mg potential emodin/kg, there were no elevated levels of micronuclei in bone marrow cells. Kinetic studies were performed in parallel to demonstrate target organ availability. Highest concentrations in the plasma were reached after 1 h with 3.4 microg rhein/ml and 0.065 microg aloeemodin/ml. In all cases, emodin was below the limit of quantification. From the results, the in vitro clastogenic activity of the senna extract could not be confirmed in the mouse micronucleus assay. Together with further negative in vivo genotoxicity studies with anthranoids, the conclusion can be drawn that there is no indication so far demonstrating a genotoxic risk for patients taking senna laxatives. PMID- 10521683 TI - Biomarkers of DNA damage in marine mammals. AB - Certain environmental contaminants found in marine mammals have been shown to cause DNA damage and cancer. The micronuclei (MN), sister chromatid exchange (SCE) and/or chromosome aberration (CA) assays were used to assess baseline (spontaneous) levels of DNA damage in blood lymphocytes of individuals of the relatively healthy and lightly contaminated Arctic beluga whale (Delphinapterus leucas), Sarasota Bay, FL, bottlenose dolphin (Tursiops truncatus) and Northwestern Atlantic grey (Halichoerus grypus) and harp (Phoca groenlandicus) seal populations. MN cell (MNC) frequencies ranged between 2 and 14/1000 binucleated (BN) cells and were statistically similar between species. In bottlenose dolphins, MNC frequency was correlated with age and was significantly higher in females than in males. No intraspecific variation in MNC frequency was found in beluga whales. Intraspecific variation was not tested in seals due to the small sample size. Frequencies of SCEs and total CAs, excluding gaps, ranged, respectively, between 1 and 15 SCE(s)/per cell and 4-6 CAs/100 cells in beluga whales. SCE and CA frequencies did not vary with age or sex in beluga whales. The MN, SCE and CA assays were found to be practical tools for the detection of DNA damage in marine mammals and could be used in the future to compare DNA damage between relatively lightly and highly contaminated populations. PMID- 10521684 TI - Polymorphisms of the GSTM1 and CYP2D6 genes associated with susceptibility to lung cancer in Chinese. AB - The case-control study was conducted to examine the association between GSTM1 null and CYP2D6Ch (T(188)/T) genotypes and lung cancer risk among Chinese of Han nationality living in Guangdong. All 191 subjects were investigated with unitary questionnaire and their DNAs were isolated from peripheral lymphocytes by standard procedures with proteinase K digestion and phenol/chloroform extraction. GSTM1(-) was detected with polymerase chain reaction (PCR) in all 191 subjects, involving 59 lung cancer cases, 59 hospital controls and 73 healthy controls. The frequencies of GSTM1(-) were not significantly different between the cases and the two controls overall. However, among adenocarcinoma of lung, the frequency of GSTM1(-) (76.9%) appeared to be higher than that in controls (49.2%), and the odd radios were 3.42-3.45. The results suggested an elevated risk for adenocarcinoma of lung would be shown by GSTM1(-). Using polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLP) to detect CYP2D6 T(188)/T genotype in 59 lung cancer patients and 59 hospital controls, it showed no significant difference between the two groups. However, non-smokers with non-T(188)/T (C(188)/C or C(188)/T) genotype showed 3.78-folds increased risk of lung cancer compared with those with T(188)/T genotype (P=0.036). The data did not suggest a substantial interaction effect between GSTM1 and CYP2D6 polymorphisms and the risk of lung cancer. Additionally, among Chinese (Han) of Guangdong, the frequency of CYP2D6 T(188) allele appeared to be 57.2%, and GSTM1(-) to be 51.8%. PMID- 10521685 TI - Protection against the bacterial mutagenicity of heterocyclic amines by purpurin, a natural anthraquinone pigment. AB - Purpurin (1,2,4-trihydroxy-9,10-anthraquinone) is a naturally occurring anthraquinone pigment found in species of madder root. We have found that the presence of purpurin in bacterial mutagenicity assays is responsible for a marked inhibition of mutagenicity induced by food-derived heterocyclic amines. Purpurin was found to be a better inhibitor of Trp-P-2-dependent mutagenicity than either epigallocatechin gallate or chlorophyllin both of which are well-established anti mutagenic components of diet. Inhibition of Trp-P-2(NHOH) mutagenicity by purpurin was dependent upon pH. It was a better inhibitor in neutral than acidic conditions. Purpurin was protective against the direct mutagen Trp-P-2(NHOH) in both the presence and the absence of hepatic S9 but required pre-incubation. Finally, purpurin was responsible for the inhibition of human CYP1A2 and human NADPH-cytochrome P450 reductase and a decrease in the bioactivation of Trp-P-2 by these enzymes when they were expressed in Salmonella typhimurium TA1538ARO. However, inhibition of Trp-P-2(NHOH)-dependent mutations suggests purpurin also has a direct effect on this mutagen in addition to inhibiting its formation by CYP1A2. PMID- 10521686 TI - Genotoxicity assessment of new synthesized acridine derivative--3,6-diamino-10 methyl-9,10-dihydroacridine. AB - A new synthesized acridine derivative, 3,6-diamino-10-methyl-9, 10 dihydroacridine (AcrH), was tested for in vitro reverse mutations with Salmonella TA strains, chromosome aberrations and sister chromatid exchanges (SCE) in human lymphocytes, and for in vivo chromosome aberrations in bone marrow of mice. Using the classic plate incorporation method, mutagenicity of AcrH in bacterial cells (TA97a, TA98, TA100 and TA102) was observed in the experiments performed with, and without, rat liver S9 metabolic activation. The reverse mutation assay showed no difference in mutagenic activity between AcrH and acriflavine (Acr(+)) in the test with TA97. The results of in vitro chromosome aberrations assay revealed potential clastogenicity. The test using macroculture of human lymphocytes induced mainly chromatid gaps. The experiments with human lymphocytes revealed SCE-inducing effect of AcrH and Acr(+). In an in vivo study, AcrH given intraperitoneally to Balb/c mice did not cause any significant increase in the percentage of cells with aberrations compared to the negative control. PMID- 10521687 TI - Comparative antimutagenic effects of D- and L-centchroman and their comparison with tamoxifen in Salmonella assay. AB - Centchroman (CC)--a contraceptive and a candidate drug for breast cancer has been developed by the Central Drug Research Institute. It has been successfully marketed as a contraceptive for last several years. CC has also been reported to exhibit partial to complete remission of lesions in 40.5% breast cancer patients. Recently, we have reported the antimutagenic effects of CC in Ames Salmonella assay and in vivo and in vitro mammalian cells in multiple mutational assay. The potent antimutagenic activity of CC and its anti-breast cancer activity prompted us to evaluate the antimutagenic effects of its enantiomers, i.e., D-centchroman (DC) and L-centchroman (LC) in the Ames Salmonella strains TA97a, TA98, TA100 and TA102 against known bacterial mutagens. Attempts have also been made to compare the results of antimutagenicity assays of CC and its enantiomers with the known breast cancer drug tamoxifen (TM). The main objective was to identify the best suitable form of CC having antimutagenic effects with anticancer profile similar to TM, would replace the latter for toxicity reasons. When mutagenicity assays were carried out with these compounds as expected like CC, none of these enantiomers or TM showed any mutagenic effects in these Salmonella strains. In the antimutagenicity assay a significantly reduced number of bacterial histidine revertant colonies were observed when positive compounds were co-incubated with certain concentrations of LC compared with bacterial plates treated with respective positive compound. This was observed in some concentrations in all the four strains in both plate incorporation and preincubation tests. The protective effects of LC in preincubation tests were slightly more than in plate incorporation tests. Both the DC and TM showed protective effects only in certain concentrations in some strains in either plate or preincubation tests. Thus the above results indicate that LC showed more protective effects in Salmonella strains TA97a, TA98, TA100 and TA102 than either DC or TM. PMID- 10521688 TI - The protective activity of alpha-hederine against H2O2 genotoxicity in HepG2 cells by alkaline comet assay. AB - This study was designed to evaluate the protective effect of alpha-hederine (alpha-hed) against H2O2-mediated DNA damage on HepG2 cell line by the alkaline comet assay. For the protective effect of alpha-hed study, cells were treated according to three protocols: pre-treatment, simultaneous treatment and post treatment. The effect of alpha-hed on catalase activity was evaluated after treating the cells with 3.36 mg/ml of 3-amino-1,2,4-triazole (AMT) singly or in combination with alpha-hed (1.5 or 3 microg/ml) and H2O2 (8.8 microM) during 1 h. The catalase activity was also biochemically measured after treating cells with alpha-hed at 1.5, 3, or 15 microg/ml during 1 h. Additionally, the influence of alpha-hed on membrane RedOx potential, pool of reduced glutathione and total protein content was evaluated by flow cytometry. In the pre-treatment, the two concentrations of alpha-hed (1.5 and 3 microg/ml) decreased the lesions induced by H2O2 (8.8 microM) significantly. This decrease was about 57.2% and 66.1%, respectively. Similar results were observed when cells were treated with alpha hed and H2O2 simultaneously. The decrease of H2O2-induced lesions was about 78.2% and 83.2% (alpha-hed 1.5 and 3 microg/ml, respectively). In the post-treatment protocol, this decrease was not significant. The combination of AMT and H2O2 induced more DNA damage than H2O2 alone (tail moment (TM) means was 31.4% and 21.8%, respectively). When alpha-hed was added to this mixture, TM means were reduced significantly (17.4% for alpha-hed 1. 5 microg/ml and 15.5% for alpha-hed 3 microg/ml). Up to 6.9 microg/ml, alpha-hed enhanced catalase activity (60.5%), followed by a decrease of the activity. Total protein content and membrane RedOx potential were slightly increased up to 11 microg/ml (14% and 3.6%, respectively) followed by a drop and a plateau. Pool of reduced glutathione remained unchanged up to 10 microg/ml then dropped and reached a plateau. In conclusion, alpha-hed could exert its protective effect against H2O2-mediated DNA damage by scavenging free radicals or by enhancing the catalase activity. PMID- 10521690 TI - DNA damage by nitrogen mustard in a gene containing multiple Sp1-binding sites. AB - The human cytochrome c(1) gene TATA-less promoter contains 10 Sp1-binding elements that regulate the activation of transcription of this gene. Quantitative PCR was used to show that nitrogen mustard induces DNA lesions within this Sp1 binding region following exposure of HeLa cells to clinical levels of the drug. Alkylation of the cytochrome c(1) gene in HeLa cells increased with reaction time up to 4 h following exposure to nitrogen mustard, with 50% of the lesions (approximately 0.8/kb) forming within 1 h. An Sp1 competition assay showed that nitrogen mustard inhibited the binding of Sp1 to the promoter region of the cytochrome c(1) gene in HeLa cells. These results show that nitrogen mustard induced damage to Sp1-binding sites may contribute to the toxicity of this compound by interfering with the activation of specific genes. PMID- 10521689 TI - The DNA repair inhibitors hydroxyurea and cytosine arabinoside enhance the sensitivity of the alkaline single-cell gel electrophoresis ('comet') assay in metabolically-competent MCL-5 cells. AB - We have found previously that the metabolically-competent human MCL-5 cell line did not appear to be usefully sensitive to the DNA-damaging effects of several carcinogens, as measured by the alkaline single-cell gel electrophoresis ('comet') assay. We therefore sought to increase its sensitivity by inhibiting DNA repair during exposure to test compounds, using 10 mM hydroxyurea (HU) and 1.8 mM cytosine arabinoside (ara-C), which inhibit DNA resynthesis during nucleotide excision repair. The following compounds were tested, using a 30-min exposure, in the absence or presence of HU/ara-C: 2-amino-3,8-dimethylimidazo[4,5 f]quinoxaline (8-MeIQx), 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (4, 8 DiMeIQx), 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-9H-pyrido[2,3 b]indole (A[alpha]C), 2-amino-3-methyl-9H-pyrido[2,3-b]indole (MeA[alpha]C), 2 amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), benzo[a]pyrene (B[a]P), 3 methylcholanthrene (3-MCA), 7, 12-dimethylbenz[a]anthracene (DMBA), 1-nitropyrene (1-NP), 2-nitrofluorene (2-NF), aniline, o-toluidine, benzene, lindane, bleomycin, cisplatin, N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), sodium chromate, chromic chloride, and diethylstilboestrol (DES). We made the following observations. The background level of comet formation was reasonably constant over several months and was increased only slightly, but significantly, in the presence of the DNA-repair inhibitors. All compounds that induced comet formation did so without appreciable cytotoxicity as assessed by trypan blue exclusion. Of the compounds tested, the heterocyclic amines and polycyclic aromatic hydrocarbons (with the exceptions of PhIP and B[a]P) failed to induce convincing levels of comet formation in the absence of repair inhibitors. In their presence the heterocyclic amines tested induced comet formation (with the exception of 8 MeIQx), with widely differing potencies. 1-NP failed to elicit marked comet formation even in the presence of HU/ara-C. Aniline and o-toluidine produced significant levels of comet formation in the absence of HU/ara-C, but in their presence comet formation was markedly increased. Benzene, lindane, bleomycin, cisplatin, MNNG, sodium chromate and chromic chloride induced comet formation in the absence of HU/ara-C, but, with the exception of cisplatin, their presence enhanced comet formation. Neither sucrose nor DES elicited comet formation under the conditions used in this study. Many more agents need to be tested in order to determine how well the comet assay using MCL-5 cells (or modified versions of it) can distinguish genotoxins from non-genotoxins. PMID- 10521692 TI - Direct analysis by small-pool PCR of MS205 minisatellite mutation rates in sperm after mutagenic therapies. AB - We have used small-pool PCR to analyse mutation in samples of sperm taken from men after mutagenic therapy. Small-pool PCR uses direct analysis of germline DNA at a highly unstable tandem-repeated "minisatellite" locus to measure rates of length-change mutation in individual sperm samples. The advantages of this approach are that the normal mutation rate is extremely high (about 0.4% per gamete at the locus analysed here), so that relatively small increases in mutation rate can be detectable in individual samples. It is known from work on sperm from untreated individuals that different alleles at this locus have different mutation rates. For this reason, we have analysed the germline mutation rates in sperm samples from two men, in each case comparing a post-treatment sample with a pre-treatment sample from the same individual. We find no evidence for altered mutation in the post-treatment sample, suggesting that the repopulation of the germ-cell compartment after treatment may be subject to stringent mechanisms for the detection and elimination of germ-cell damage. PMID- 10521691 TI - The application of the micronucleus test in Chinese hamster V79 cells to detect drug-induced photogenotoxicity. AB - Recent reports on the photochemical carcinogenicity and photochemical genotoxicity of fluoroquinolone antibacterials led to an increasing awareness for the need of a standard approach to test for photochemical genotoxicity. In this study the micronucleus test using V79 cells was adapted to photogenotoxicity testing. Results of using different UVA/UVB relationships enabled us to identify a suitable irradiation regimen for the activation of different kinds of photosensitizers. Using this regimen, 8-methoxypsoralen and the fluoroquinolones lomefloxacin, grepafloxacin and Bay Y 3118 were identified to cause micronuclei and toxicity upon photochemical activation. Among the phenothiazines tested, chlorpromazine and 2-chlorophenothiazine, were positive for both endpoints, whereas triflupromazine was only slightly photoclastogenic in the presence of strong phototoxicity. Among the other potential human photosensitizers tested (oxytetracycline, doxycycline, metronidazole, emodin, hypericin, griseofulvin), only hypericin was slightly photogenotoxic. Photochemical toxicity in the absence of photochemical genotoxicity was noted for doxycycline and emodin. With the assay system described, it is possible to determine photochemical toxicity and photochemical genotoxicity concomitantly with sufficient reliability. PMID- 10521693 TI - SOS induction of selected naturally occurring substances in Escherichia coli (SOS chromotest). AB - Naturally occurring substances were tested for genotoxicity using a modified laboratory protocol of the Escherichia coli PQ37 genotoxicity assay (SOS chromotest) in the presence and in the absence of an exogenous metabolizing system from rat liver S9-mix. Aristolochic acid I, II, the plant extract aristolochic acid and psoralene were genotoxic; cycasine, emodine, monocrotaline and retrorsine were classified as marginal genotoxic in the SOS chromotest in the absence of S9-mix. In the presence of an exogenous metabolizing system from rat liver S9-mix aristolochic acid I, the plant extract, beta-asarone, cycasin, monocrotaline, psoralen and retrorsine showed genotoxic effects; aristolochic acid II marginal genotoxic effects. Arecoline, benzyl acetate, coumarin, isatidine dihydrate, reserpine, safrole, sanguinarine chloride, senecionine, senkirkine, tannin and thiourea revealed no genotoxicity in the SOS chromotest either in the presence or in the absence of an exogenous metabolizing system from rat liver S9-mix. For 17 of 20 compounds, the results obtained in the SOS chromotest could be compared to those obtained in the Ames test. It was found that 12 (70.6%) of these compounds give similar responses in both tests (6 positive and 6 negative responses). The present investigation and those reported earlier, the SOS chromotest, using E. coli PQ37, was able to detect correctly most of the Salmonella mutagens and non-mutagens. PMID- 10521694 TI - Delayed transfection of DNA after riboflavin mediated photosensitization increases G:C to C:G transversions of supF gene in Escherichia coli mutY strain. AB - We have previously reported that the majority of base substitution mutations of the Escherichia coli supF gene induced by riboflavin mediated photosensitization were G:C to C:G changes, in addition to G:C to T:A changes which were probably caused by 8-hydroxyguanine (oh(8)Gua), in wild type and mutM mutator mutant strains. This implies that lesions other than oh(8)Gua are produced by riboflavin photosensitization. G:C to C:G base substitutions have been found in the mutations induced by ionizing radiation and reactive oxygen species, as well as spontaneous mutation. To characterize the G:C to C:G mutation, riboflavin- photosensitized plasmid DNA carrying the supF gene was left at room temperature for 5 h in the dark before transfection. The delayed transfection gave a mutational spectrum different from that for immediate transfection. G:C to C:G transversions significantly increased in mutY mutator strain, in which the transversion was not detected in the immediate transfection. Lesions causing G:C to C:G changes increased during 5-h holding after photosensitization and MutY protein presumably takes part in this type of base change mutation. PMID- 10521696 TI - A cytogenetic study of nuclear power plant workers using the micronucleus centromere assay. AB - A cytogenetic study was performed in 215 nuclear power plant workers occupationally exposed to radiation using the micronucleus-centromere assay for peripheral blood lymphocytes. As control population served administrative staff with yearly doses below 1 mSv. The increase of the micronucleus frequency with age, observed in the non-smoking control population, is mainly due to an enhanced number of centromere-positive micronuclei, pointing to an increased chromosome loss. No differences in the number of micronuclei, centromere-positive and centromere-negative micronuclei between smokers and non-smokers are observed. An analysis of the micronucleus data vs. the dose accumulated over the 10 years preceding the venepuncture shows no significant clastogenic or aneuploidogenic effects of the exposure in the studied population which is representative for workers in the nuclear industry at present. According to the linear fits to our data an increase of the micronucleus frequency pro rata 0.5 per 1000 binucleated cells per year, related to the centromere-negative micronuclei, may be expected for workers with the maximal tolerable dose of 20 mSv/year. PMID- 10521695 TI - Anticlastogenic effects of Ginkgo biloba extract (EGb 761) and some of its constituents in irradiated rats. AB - In a previous study we reported that radiation-induced clastogenic factors (CF) are found in the plasma of Chernobyl accident recovery workers and that their chromosome damaging effects are inhibited by antioxidant treatment with a Ginkgo biloba extract (EGb761). In the present study, we induced CF in rats with a radiation dose of 4.5 Gy. The protective effects of the complete extract were compared to those obtained with the extract devoid of its terpene fraction (CP205), with isolated ginkgolides A+B and bilobalide at the concentrations present in EGb761. The pretreatment samples were taken at day 22 postirradiation, the posttreatment samples the day following arrest of the 3-week treatment. The adjusted clastogenic score (ACS) were reduced from 11.71+/-3.55 to 2.00+/-2.83 after treatment with 100 mg/kg and from 13.43+/-2.23 to 4.29+/-2.14 with 50 mg/kg of the complete extract (p<0.0001). Similar protective effects were observed with CP205, ginkgolides and bilobalide (p<0. 001), while the reduction of ACS in placebo-treated rats was not statistically significant (12.80+/-1.79 and 9.20+/ 2.68). However, if the efficacy of the treatment was compared to placebo, only the complete extract was significantly protective. While all components exerted anticlastogenic effects at the concentrations present in the complete extract, the comparison of the different groups by analysis of variance did not reveal significant differences. This may be due to to the small number of animals available in each treatment group. The complete extract reduced the ACS by 83% at the dose of 100 mg/kg, while the lower dose of 50 mg/kg and the three components reached only 66%-68% reduction. The better protection provided by the complete extract is due to synergistic rather than to additive effects. PMID- 10521697 TI - Evidence for oxidative metabolism in the genotoxicity of the atmospheric reaction product 2-nitronaphthalene in human lymphoblastoid cell lines. AB - 2-Nitronaphthalene (2NN) has been identified as a mutagenic atmospheric reaction product of naphthalene in the Ames bacterial reversion assay. Recent experiments have shown this nitroarene to be genotoxic in a human lymphoblastoid cell line (MCL-5) transfected with plasmids encoding epoxide hydrolase and four cytochrome P450 monooxygenase activities. The present study investigated the genotoxicity of 2NN in two related human B-lymphoblastoid cell lines, h1A1v2 containing a single P450 isozyme (cytochrome P450 1A1) and L3 cells which are isogenic with MCL-5 cells and are distinguished only by the absence of transfected plasmids. The results indicate that 2NN-induced mutagenesis at the heterozygous thymidine kinase (tk) locus was dependent on metabolic activities provided by the transfected plasmids in MCL-5; no significant induction of mutants was observed in L3 cells studied in parallel. A similar induction of mutation was observed in h1A1v2 and MCL-5 cell lines at the tk locus and no induction was observed at the hemizygous hypoxanthine phosphoribosyl transferase (hprt) locus. The induction of mutations in h1A1v2 cells suggests that cytochrome P450 1A1 alone can activate 2NN to a mutagenic species, however, this interpretation may be confounded by differences between the h1A1v2 and MCL-5 cell lines. The observed genotoxic activity induced by 2NN prompted testing of the amino analogue, beta naphthylamine (betaNA), to investigate potential similarities in the metabolic activation pathways of the two compounds. The negative response of betaNA in all cell lines suggests that 2NN and betaNA are not activated in these human cells by similar metabolic pathways. PMID- 10521698 TI - Mutation spectrum of 4-nitroquinoline N-oxide in the lacI transgenic Big Blue Rat2 cell line. AB - This paper describes the spectrum of mutations induced by 4-nitroquinoline N oxide (4-NQO) in the lacI target gene of the transgenic Big Blue Rat2 cell line. There are only a few report for the mutational spectrum of 4-NQO in a mammalian system although its biological and genetic effects have been well studied. Big Blue Rat2 cells were treated with 0.03125, 0.0625 or 0.125 microg/ml of 4-NQO, the highest concentration giving 85% survival. Our results indicated that the mutant frequency (MF) induced by 4-NQO was dose-dependent with increases from three- to seven-fold. The DNA sequence analysis of lacI mutants from the control and 4-NQO treatment groups revealed an obvious difference in the spectra of mutations. In spontaneous mutants, transition (60%) mutations, especially G:C- >A:T transition (45%), were most frequent. However, the major type of base substitution after treatment of 4-NQO was transversions (68.8%), especially G:C- >T:A (43.8%), while only 25% of mutants were transitions. These results are consistent with those produced by 4-NQO in other systems and the transgenic assay system will be a powerful tool to postulate more accurately the mechanism of chemical carcinogenesis involved. PMID- 10521700 TI - A novel glycosphingolipid from gram-negative aquatic bacteria. AB - The chloroform-methanol extractable lipids of the Gram-negative fresh-water bacteria Arcocella aquatica NO-502 and Flectobacillus major FM were found to contain an unusual ninhydrin-positive glycolipid. It was purified by two-stage silica gel-column chromatography. By the use of IR and (1)H-NMR spectroscopy, mass spectrometry and chemical-degradation experiment, the lipid was established to be 1-O-monoglycosyl ceramide, the carbohydrate moiety of which was the alpha pyranose-ring form of 7-desoxy-7-amino-D-manno-heptulosonic acid, or 1 hydroxycarbonyl-6-deoxy-6-amino-alpha-D-mannopyranose. The ceramide portion consisted mainly (by 95% in the A. aquatica glycolipid and 80% in the F. major glycolipid) of 2-N-(2'-D-hydroxy-13'-methyltetradecanoyl)-15-methyl-4(E)-hexad ecasph ingenine. The minor molecular species differed from the major one only in fatty acid structure. The glycolipid accounted for 8 and 11% of the total lipids extracted from A. aquatica NO-502 and F. major FM cells, respectively. PMID- 10521699 TI - Interactions of retinoid binding proteins and enzymes in retinoid metabolism. AB - Naturally occurring retinoids (vitamin A or retinol and its active metabolites) are vital for vision, controlling the differentiation program of epithelial cells in the digestive tract and respiratory system, skin, bone, the nervous system, the immune system, and for hematopoiesis. Retinoids are essential for growth, reproduction (conception and embryonic development), and resistance to and recovery from infection. The functions of retinoids in the embryo begin soon after conception and continue throughout the lifespan of all vertebrates. Both naturally occurring and synthetic retinoids are used in the therapy of various skin diseases, especially acne, for augmenting the treatment of diabetes, and as cancer chemopreventive agents. Retinol metabolites serve as ligands that activate specific transcription factors in the superfamily of steroid/retinoid/thyroid/vitamin D/orphan receptors and thereby control gene expression. Additionally, retinoids may also function through non-genomic actions. Various retinoid binding proteins serve as partners in retinoid function. These binding proteins show high specificity and affinity for specific retinoids and seem to control retinoid metabolism in vivo qualitatively and quantitatively by reducing 'free' retinoid concentrations, protecting retinoids from non-specific interactions, and chaperoning access of metabolic enzymes to retinoids. Implementation of the physiological effects of retinoids depends on the spatial-temporal expressions of binding proteins, receptors and metabolic enzymes. This review will discuss current understanding of the enzymes that catalyze retinol and retinoic acid metabolism and their unique and integral relationship to retinoid binding proteins. PMID- 10521701 TI - Phytanic acid alpha-oxidation: identification of 2-hydroxyphytanoyl-CoA lyase in rat liver and its localisation in peroxisomes. AB - Phytanic acid is broken down by alpha-oxidation in three steps finally yielding pristanic acid. The first step occurs in peroxisomes and is catalysed by phytanoyl-CoA hydroxylase. We have studied the second step in the alpha-oxidation pathway, which involves conversion of 2-hydroxyphytanoyl-CoA to pristanal catalysed by 2-hydroxyphytanoyl-CoA lyase. To this end, we have developed a stable isotope dilution gas chromatography-mass spectrometry assay allowing activity measurements in rat liver homogenates. Cell fractionation studies demonstrate that in rat liver 2-hydroxyphytanoyl-CoA lyase is localised in peroxisomes. This finding may have important implications for inherited diseases in man characterised by impaired phytanic acid alpha-oxidation. PMID- 10521702 TI - IL-13 induces serine phosphorylation of cPLA2 in mouse peritoneal macrophages leading to arachidonic acid and PGE2 production and blocks the zymosan-induced serine phosphorylation of cPLA2 and eicosanoid production. AB - In a recent investigation, we demonstrated that long-term treatment of macrophages with IL-13 enhances cPLA2 expression and modulates zymosan-stimulated AA mobilization. In the present study, we examine the ability of IL-13 to modify the cPLA2 activity and the AA mobilization of macrophages after a short-period of treatment. We demonstrate that in resting macrophages, IL-13 induces, through a MAP kinase-dependent process, (1) an increase of free AA release within 15 min, followed by increased PGE2 production and (2) a time-dependent serine phosphorylation of cPLA2. Conversely, in macrophages stimulated by zymosan, IL-13 added 30 min before zymosan inhibited the AA release and the serine phosphorylation of cPLA2 induced by the phagocytic agonist. In conclusion, these findings show for the first time that a Th2-type cytokine can upregulate cPLA2 activity and downregulate zymosan-induced AA metabolism. Thus, establishment of the connection between these two events may help to understand the complex regulatory role of IL-13 on the macrophage AA metabolism. PMID- 10521703 TI - Occurrence of lysophosphatidic acid and its alkyl ether-linked analog in rat brain and comparison of their biological activities toward cultured neural cells. AB - Rat brain was found to contain substantial amounts of potent bioactive lipids lysophosphatidic acid (acyl LPA) (3.73 nmol/g tissue) and lysoplasmanic acid (alkyl LPA) (0.44 nmol/g tissue). The presence of alkyl LPA was confirmed by mild alkaline hydrolysis analysis and by gas chromatography/mass spectrometry analysis of the trimethylsilyl derivative. This is the first clear evidence of the occurrence of an alkyl LPA in nature. The predominant molecular species of acyl LPA are 18:1-, 18:0- and 16:0-containing species (46. 9, 22.5 and 18.8%, respectively). A significant amount of a 20:4-containing species (7.2%) was also detected in the acyl LPA fraction. We also confirmed that rat brain alkyl LPA consists of 16:0-, 18:0- and 18:1-containing species. Noticeably, either acyl or alkyl LPA is capable of stimulating neuroblastomaxglioma hybrid NG108-15 cells to elicit a Ca(2+) transient, the potencies being almost the same. Both acyl and alkyl LPAs also induce cell rounding upon addition to the cells. These results suggest that acyl and alkyl LPAs play important physiological roles as intercellular signaling molecules as well as the roles as metabolic intermediates in the nervous system. PMID- 10521704 TI - Equilibrium in the hydrolysis and synthesis of cannabimimetic anandamide demonstrated by a purified enzyme. AB - Anandamide, an endogenous ligand for cannabinoid receptors, loses its biological activities when it is hydrolyzed to arachidonic acid and ethanolamine by anandamide amidohydrolase. We overexpressed a recombinant rat enzyme with a hexahistidine tag in a baculovirus-insect cell expression system, and purified the enzyme with the aid of a Ni-charged resin to a specific activity as high as 5.7 micromol/min/mg protein. The purified recombinant enzyme catalyzed not only the hydrolysis of anandamide and palmitoylethanolamide, but also their reverse synthetic reactions. In order to attain an equilibrium of the anandamide hydrolysis and its reverse reaction within 10 min, we utilized a large amount of the purified enzyme. The equilibrium constant ([arachidonic acid][ethanolamine])/([anandamide][water]) was calculated as 4x10(-3) (37 degrees C, pH 9.0). These experimental results with a purified enzyme preparation quantitatively confirmed the reversibility of the enzyme reaction previously observed with crude enzyme preparations. PMID- 10521705 TI - Can the fatty acid selectivity of plant lipases be predicted from the composition of the seed triglyceride? AB - To address the question can the fatty acid selectivity of plant lipases be predicted from the composition of the seed triglyceride, we have characterised the selectivity of lipases from a wide range of oilseeds with diverse fatty acid compositions. For this study, a novel hydrolysis assay using a fully randomised oil, was developed. From some seed sources (e.g. Cinnamomum camphora), lipases show high preference for particular fatty acids, whilst from others (e.g. Brassica napus, Theobroma cacao80% saturated or 'unusual' fatty acids may contain lipases which exhibit selectivity. It therefore follows that since the majority of seeds are composed of unsaturated fatty acids, that highly selective lipases will be unusual in nature. However lipases from some species of the Cuphea genera show exceptionally high preference for particular fatty acids. For example, lipase from seeds of Cuphea procumbans has over 20-fold selectivity for C10:0. PMID- 10521706 TI - Specific protein targets of 13-oxooctadecadienoic acid (13-OXO) and export of the 13-OXO-glutathione conjugate in HT-29 cells. AB - The linoleic acid metabolite, 13-oxooctadecadienoic acid (13-OXO), is reactive with cellular thiols. In the present report, incubations of HT-29 or CaCo-2 homogenates with 13-OXO and GSH indicate that HT-29 cell homogenates produce a 13 OXO-GSH conjugate. The conjugate formed was likely of enzymatic origin as chiral phase HPLC showed the major product consisted of only one of two possible diastereomers. The glutathione transferase activity (GST), using chlorodinitrobenzene, was found to be 126 nmol/mg/min in HT-29 cells and 21 nmol/mg/min in CaCo-2 cells. These levels of activity are consistent with the relative ability of the two cell lines to conjugate GSH to 13-OXO. Incubation of intact HT-29 cells with either 13-OXO, or the metabolic precursor 13 hydroxyoctadecadienoic acid (13-HODE), showed detectable 13-OXO-GSH conjugate in the media, but none in the cells. The stereochemistry of the extracellular conjugate suggested an enzymatic origin. In additional experiments, the labeling of cellular protein by 13-HODE was much more specific than the labeling of protein by 13-OXO suggesting that in situ generation of 13-OXO from 13-HODE confers selectivity on the reactions between cellular thiols and 13-OXO. These results demonstrate that in HT-29 cells, 13-HODE is converted to 13-OXO which then either reacts with cellular protein or is conjugated to GSH by GST. The 13 OXO-GSH conjugate is then exported from the cell. PMID- 10521707 TI - Overexpression of phospholipase C-gamma1 suppresses UVC-induced apoptosis through inhibition of c-fos accumulation and c-Jun N-terminal kinase activation in PC12 cells. AB - Ultraviolet-C (UVC) irradiation induces DNA damage and UVC-irradiated cells undergo cell growth arrest to repair the damaged DNA or the induction of apoptosis to prevent the risk of neoplastic transformation. Phospholipase C gamma1 (PLC-gamma1) is a mediator of growth factor induced-signal cascade, catalyzing the hydrolysis of phosphatidyl 4,5-bisphosphate to generate second messengers, diacylglycerol and inositol 1,4,5-trisphosphate (IP(3)). PLC-gamma1 is activated by phosphorylation of tyrosine residues upon occupation of cell surface receptors by growth factors and plays an important role in controlling cellular proliferation and differentiation. In this study, we found that PLC gamma1 was tyrosine phosphorylated within 2.5 min after UVC irradiation. To investigate the role of UVC-induced tyrosine phosphorylation of PLC-gamma1, we compared the effect of UVC between PLC-gamma1 overexpressing cells and empty vector transfected cells. Overexpression of PLC-gamma1 inhibited UVC-induced sub diploid peak and DNA fragmentation. Northern blot analysis revealed that UVC induced c-fos mRNA accumulation was inhibited in PLC-gamma1 overexpressing cells, while c-jun expression was not affected. In addition, UVC-induced activation of c Jun N-terminal kinase (JNK) was significantly suppressed in PLC-gamma1 overexpressing cells. These results suggest that PLC-gamma1 may associate with the protective function against the UVC-induced cell death progression via the inhibition of accumulation of c-fos mRNA and the inhibition of JNK kinase activity. PMID- 10521708 TI - Liquid chromatography/mass spectrometry analysis of mixtures of rhamnolipids produced by Pseudomonas aeruginosa strain 57RP grown on mannitol or naphthalene. AB - Liquid chromatography/mass spectrometry using electrospray ionisation was used to analyse rhamnolipids produced by a Pseudomonas aeruginosa strain with mannitol or naphthalene as carbon source. Identification and quantification of 28 different rhamnolipid congeners was accomplished using a reverse-phase C(18) column and a 30 min chromatographic run. Isomeric rhamnolipids that were not chromatographically resolved could be identified by interpretation of their mass spectra and their relative proportions estimated. The most abundant rhamnolipid produced on mannitol contained two rhamnoses and two 3-hydroxydecanoic acid groups. The most abundant rhamnolipid produced from naphthalene contained two rhamnoses and one 3-hydroxydecanoic acid group. PMID- 10521709 TI - Manipulation of cholesterol and cholesteryl ester synthesis has multiple effects on the metabolism of apolipoprotein B and the secretion of very-low-density lipoprotein by primary hepatocyte cultures. AB - Inhibition of esterified and non-esterified cholesterol synthesis by lovastatin in primary rat hepatocytes suppressed the net synthesis and very-low-density lipoprotein (VLDL) secretion of apolipoprotein B (apoB)-48 and apoB-100. Lovastatin did not alter the rates of apoB-48 and apoB-100 post-translational degradation. 25-Hydroxycholesterol, which inhibited non-esterified cholesterol synthesis but increased the synthesis of cholesteryl ester, showed differential effects on the metabolism of apoB-48 and apoB-100. Whereas the secretion of apoB 48 VLDL was suppressed there was no effect on the secretion of apoB-100 VLDL. The post-translational degradation of apoB-48, but not of apoB-100, was enhanced by 25-hydroxycholesterol. The net synthesis rates of apoB-48 and apoB-100 were unaffected by 25-hydroxycholesterol. The inhibitory effect of lovastatin alone on the net synthesis of apoB-48 and apoB-100 was reversed by the simultaneous presence of 25-hydroxycholesterol, suggesting a role for newly synthesised cholesteryl ester. Prevention of the reversal effect by the acyl-CoA: cholesterol acyltransferase (ACAT) inhibitor YM 17E supported this interpretation. In the presence of lovastatin, restoration of the net synthesis of apoB by 25 hydroxycholesterol was not accompanied by an increased VLDL output of apoB-48 and apoB-100. However, under these conditions there was an increased post translational degradation of apoB-48 and apoB-100. These results suggest that interference with intracellular cholesterol and cholesteryl ester metabolism interrupts VLDL assembly at sites of both apoB net synthesis and post translational degradation. PMID- 10521710 TI - Analogues and homologues of N-palmitoylethanolamide, a putative endogenous CB(2) cannabinoid, as potential ligands for the cannabinoid receptors. AB - The presence of CB(2) receptors was reported in the rat basophilic cell line RBL 2H3 and N-palmitoylethanolamide was proposed as an endogenous, potent agonist of this receptor. We synthesized a series of 10 N-palmitoylethanolamide homologues and analogues, varying by the elongation of the fatty acid chain from caproyl to stearoyl and by the nature of the amide substituent, respectively, and evaluated the affinity of these compounds to cannabinoid receptors in the rat spleen, RBL 2H3 cells and CHO-CB(1) and CHO-CB(2) receptor-transfected cells. In rat spleen slices, CB(2) receptors were the predominant form of the cannabinoid receptors. No binding of [(3)H]SR141716A was observed. [(3)H]CP-55,940 binding was displaced by WIN 55,212-2 and anandamide. No displacement of [(3)H]CP-55,940 or [(3)H]WIN 55,212-2 by palmitoylethanolamide derivatives was observed in rat spleen slices. In RBL-2H3 cells, no binding of [(3)H]CP-55,940 or [(3)H]WIN 55,212-2 could be observed and conversely, no inhibitory activity of N-palmitoylethanolamide derivatives and analogues was measurable. These compounds do not recognize the human CB(1) and CB(2) receptors expressed in CHO cells. In conclusion, N palmitoylethanolamide was, in our preparations, a weak ligand while its synthesized homologues or analogues were essentially inactive. Therefore, it seems unlikely that N-palmitoylethanolamide is an endogenous agonist of the CB(2) receptors but it may be a compound with potential therapeutic applications since it may act via other mechanisms than cannabinoid CB(1)-CB(2) receptor interactions. PMID- 10521711 TI - A structural comparison of the total polar lipids from the human archaea Methanobrevibacter smithii and Methanosphaera stadtmanae and its relevance to the adjuvant activities of their liposomes. AB - Mice were immunized with bovine serum albumin (BSA) entrapped within archaeosomes (i.e. liposomes) composed of the total polar lipids (TPL) from the two methanogenic archaea common to the human digestive tract. Methanobrevibacter smithii archaeosomes boosted serum anti-BSA antibody to titers comparable to those achieved with Freund's adjuvant, whereas Methanosphaera stadtmanae archaeosomes were relatively poor adjuvants. An explanation for this difference was sought by analysis of the polar lipid composition of each archaeobacterium. Fast atom bombardment mass spectrometry and NMR analyses of the purified lipids revealed a remarkable similarity in the ether lipid structures present in each TPL extract. However, the relative amounts of each lipid species varied dramatically. The phospholipid fraction in M. stadtmanae TPL was dominated by archaetidylinositol (50 mol% of TPL) and the glycolipid fraction by beta-Glcp (1,6)-beta-Glcp-(1,1)-archaeol (36 mol%), whereas in M. smithii extracts, both caldarchaeol and archaeol lipids containing a phosphoserine head group were relatively abundant. Liposomes prepared from purified archaetidylinositol and from M. stadtmanae TPL supplemented with increasing amounts of phosphatidylserine elicited poor humoral responses to encapsulated BSA. A dramatic loss in the adjuvanticity of M. smithii archaeosomes was seen upon incorporation of 36 mol% of the uncharged lipid diglucosyl archaeol and, to a lesser extent, of 50 mol% of archaetidylinositol. Interestingly, the relative rates of uptake of M. smithii and M. stadtmanae archaeosomes by phagocytic cultures in vitro were similar. Thus, the lipid composition may influence archaeosome adjuvanticity, particularly a high diglucosyl archaeol and/or archaetidyl inositol content, resulting in a low adjuvant activity. PMID- 10521712 TI - Troponin: structure, properties, and mechanism of functioning. AB - This review discusses the structure and properties of the isolated components of troponin, their interaction, and the mechanisms of regulation of contractile activity of skeletal and cardiac muscle. Data on the structure of troponin C in crystals and in solution are presented. The Ca2+-induced conformational changes of troponin C structure are described. The structure of troponin I is analyzed and its interaction with other components of actin filaments is discussed. Data on phosphorylation of troponin I by various protein kinases are presented. The role of troponin I phosphorylation in the regulation of contractile activity of the heart is analyzed. The structural properties of troponin T and its interaction with other components of thin filaments are described. Data on the phosphorylation of troponin T are presented and the effect of troponin T phosphorylation on contractile activity of different muscles is discussed. Modern models of the functioning of troponin are presented and analyzed. PMID- 10521713 TI - Trypsin-like proteinase and its endogenous inhibitor from Yersinia pseudotuberculosis. Biological activity. AB - A trypsin-like proteinase (YPTP) and its endogenous inhibitor (ITYP) were isolated from the culture filtrate of the pathogenic bacterium Yersinia pseudotuberculosis, and their biological activities were studied. YPTP was found to be highly toxic for random-bred white mice. Under in vitro conditions the proteolytic enzyme destroyed protective proteins of the immune system of the animals--IgG, IgA, and proteins of the complement system (CIq, C3, and C5)--and, consequently, was a pathogenetic factor in yersinioses. The inhibitor ITYP was shown to manifest antibacterial activity against virulent forms of Yersinia pseudotuberculosis, Escherichia coli, and Salmonella typhimurium. The ITYP preparation was harmless and nontoxic. PMID- 10521714 TI - Hydrolysis of tripolyphosphate by purified exopolyphosphatase from Saccharomyces cerevisiae cytosol: kinetic model. AB - The kinetics of hydrolysis of tripolyphosphate by purified exopolyphosphatase from Saccharomyces cerevisiae cytosol has been studied in the presence of Mg2+. Two kinetic models suggesting the formation of complexes of tripolyphosphate and the enzyme with Mg2+ are compared. Both models suggest that only enzyme- substrate complexes containing Mg2+ and tripolyphosphate simultaneously are able to hydrolyze the tripolyphosphate. The first model suggests that the enzyme is able to bind to Mg2+ independently from substrate binding. The second model does not consider this possibility, but suggests that both complexes containing tripolyphosphate and Mg2+ in proportion 1:1 and 1:2 can serve as the reaction substrates. The description of the experimental data by both models is essentially the same. The complex containing tripolyphosphate and Mg2+ in proportion 1:1 is optimal for the enzyme activity, the complex containing tripolyphosphate and Mg2+ in proportion 1:2 being hydrolyzed at a lower rate. PMID- 10521715 TI - Subcellular localization and properties of glyoxylate cycle enzymes in the liver of rats with alloxan diabetes. AB - Key enzymes of the glyoxylate cycle (isocitrate lyase and malate synthetase) were found in the liver and kidney of rats suffering from alloxan diabetes. The activities of these enzymes in the liver were 0.080 and 0.0430 U/mg protein, respectively. Isocitrate lyase activity in the kidney was 0.030 U/mg protein, and that of the malate synthetase was 0.018 U/mg protein. Peroxisomal localization of the enzymes was shown. A novel malate dehydrogenase isoform was found in a liver of rats suffering from the alloxan diabetes. The isocitrate lyase was isolated by selective (NH4)2SO4 precipitation and DEAE-Toyopearl chromatography. The resulting enzyme preparation had specific activity 6.1 U/mg protein, corresponding to 76.25-fold purification with 32.6% yield. The isocitrate lyase was found to follow the Michaelis--Menten kinetic scheme (Km for isocitrate, 0.08 mM) and to be competitively inhibited by glucose 1-phosphate (Ki = 1. 25 mM), succinate (Ki = 1.75 mM), and citrate (Ki = 1.0 mM); the pH optimum of the enzyme was 7.5 in Tris-HCl buffer. PMID- 10521716 TI - Study of the structure of the thylakoid membrane-bound chloroplast coupling factor CF1. AB - The structure of thylakoid membrane-bound chloroplast coupling factor CF1 was studied by limited proteolysis followed by sodium dodecylsulfate polyacrylamide gel electrophoresis and N-terminal sequence analysis. The N-terminal fragment of the alpha-subunit was shown to have an exposed area including the peptide bond R21-E22. The cleavage of this peptide bond caused the alphaK24-V25 bond to be exposed to the outside. In the N-terminal fragment of the beta-subunit, the L14 E15 bond was identified and found to be subject to trypsinolysis. Also, the alphaR140-S141, alphaG160-R161, and betaG102-G103 bonds were accessible to the proteolytic attack. In general, the beta-subunit of membrane-bound CF1 is more sensitive to proteolysis than that of solubilized CF1. The products of proteolysis of the alpha-subunit did not contain the polypeptides typical of the reaction of cleavage of the alphaE17-G18 and alphaE22-V23 bonds in isolated CF1. These results suggest a significant structural difference between soluble and membrane-bound CF1. A number of peptide bonds, alphaG160-R161 in particular, were shown to be shielded from proteolytic attack by papain in illuminated thylakoid membranes, probably as a result of membrane energization. In contrast, the light induced reduction of the gamma-subunit caused an increase in the accessibility of some peptide bonds to this protease, including the alphaG160-R161 bond. PMID- 10521717 TI - Conformational analysis of the biologically active cyclic analog of beta casomorphin H-Tyr-cyclo[D-OrnPheProGly]. AB - A biologically active analog of beta-casomorphin, H-Tyr-cyclo[D-OrnPheProGly], was studied by theoretical conformational analysis. Random sampling was used to search the conformational space of allowed dihedral angles. Among 53 low-energy conformers with different folding of the peptide cyclic moiety, only those were selected which correspond to the low-energy area of the model linear tripeptide Ac-D-AlaAlaPro-NHMe. This peptide was used as the main chain "template" for the D OrnPheProGly fragment of the studied cyclopeptide molecule. Only 15 conformers were chosen as potentially biologically active structures. The conformational possibilities of the Phe residue were constrained to the combined (A + G) region of the Ala residue phi,psi-map for linear peptides. PMID- 10521718 TI - Enhancement of the affinity of cellobiohydrolase I and its catalytic domain to cellulose in the presence of the reaction product--cellobiose. AB - The catalytic domain of cellobiohydrolase I from Trichoderma reesei has been obtained by papain treatment of the native enzyme adsorbed onto the surface of microcrystalline cellulose. Both the intact and the truncated enzyme are almost equally active toward soluble fluorogenic derivatives of cellobi-, -tri-, -tetra , and -pentaose, the fastest and the slowest fluorophore liberation being observed for MUF-cellopenta- and -tetraose, respectively. Titration of the active centers of the intact enzyme and its catalytic domain with MUF-cellotetraose showed their molecular masses to be 49 and 39 kD, respectively, the dissociation constants of the enzyme-soluble ligand complexes being almost equal (65 and 70 nM at 20 degrees C, respectively). In contrast, the intact enzyme and its catalytic core have been shown to significantly (50-60 times) differ in their affinity to insoluble microcrystalline cellulose at low enzyme loading (up to 10 mg per g of the substrate). At 20 degrees C the dissociation constants for the two forms of the enzyme are estimated to be 10 and 500 nM, respectively. Surprisingly, under these conditions the reaction product and inhibitor, cellobiose (Ki = 10 microM), at the concentration 10 mM, increased 3-4-fold the affinity of both the intact cellobiohydrolase and its catalytic domain to cellulose. PMID- 10521719 TI - Interaction of mutant alkaline phosphatase precursors with membrane phospholipids in vivo and in vitro. AB - Positively charged amino acid residues in the N-terminal domain of the signal peptides of secreted proteins are thought to interact with negatively charged anionic phospholipids during the initiation of secretion. To test this hypothesis, substitutions of the uncharged Ala or the negatively charged Glu residue for the positively charged Lys-20 of the N-terminus of the signal peptide of Escherichia coli alkaline phosphatase were introduced using a modified method of oligonucleotide-directed mutagenesis. We found that Lys-20 is involved in the interaction of the signal peptide with anionic phospholipids in vivo and effects the efficiency of insertion of the signal peptide of isolated precursor into model phospholipid membranes in vitro. We also show that the efficiency of signal peptide insertion into the lipid bilayer depends on the fluidity of the bilayer. PMID- 10521720 TI - A new subtilisin-like proteinase from roots of the dandelion Taraxacum officinale Webb S. L. AB - A serine proteinase from roots of Taraxacum officinale Webb S. L. was isolated by affinity chromatography and gel-filtration on Superose 6R using FPLC. The enzyme is a 67-kD glycoprotein containing 54% carbohydrate which we have named taraxalisin. The substrate specificity of taraxalisin toward synthetic peptides and oxidized insulin B-chain is comparable with that of cucumisin from Cucumis melo and the subtilisin-like serine proteinase macluralisin from Maclura pomifera. The proteinase is inactivated by DFP and PMSF. Taraxalisin exhibits maximal activity at pH 8.0. The pH range for stability of the enzyme is narrow- 6.0-9.0. The temperature optimum for the subtilisin-like activity is 40 degrees C. The N-terminal sequence of taraxalisin has 40% of its residues identical to those of subtilisin Carlsberg. Thus, the serine proteinase from dandelion roots is a member of the subtilisin family, which is evidently widespread in the plant kingdom. PMID- 10521721 TI - Probability description of ligand-receptor interactions. Evaluation of reliability of events with small and supersmall doses. I. Kinetics of ligand receptor interactions. AB - We have developed mathematical methods for describing ligand-receptor interactions (LRI) using Markov chains. Under some conditions, the mean value of ligand-receptor complexes obtained using Markov chains coincides with that obtained from the law of mass action. Using the calculated ratio of standard deviation to mean number of ligand-receptor complexes, we show that with small concentrations of ligand-receptor complexes LRI must be described using probability methods. Using data from the literature, we show that LRI description using the mass-action law under these conditions can cause significant errors in interpretation of experimental data. PMID- 10521722 TI - Effect of hydration degree of aerosol OT reversed micelles and surfactant concentration in heptane on spectral and catalytic properties of catalase. AB - The influence of micelle hydration degree (w0) and AOT concentration on fluorescence, circular dichroism (CD), catalytic activity, and stability of catalase in Aerosol OT (AOT) reversed micelles in heptane was investigated. The quantitative parameters--differential fluorescence of catalase (DeltaI), protein molar ellipticity ([theta]lambda), initial rate of catalytic reaction, catalase efficiency (kcat/Km), and rate constant of enzyme inactivation (kin, sec-1)- decreased with increasing AOT concentration in micellar systems, reflecting the interaction of solubilized catalase with the AOT micellar aggregates in heptane. The dependences of all these parameters on increasing hydration degree of micelles (w0) were characterized by the appearance of maxima at w0 of 8, 15-18, and 26-30. These maxima are suggested to reflect three different states of catalase in the micellar system, distinguished by their conformations and catalytic activity, which is determined by the micellar microenvironment of the enzyme. PMID- 10521723 TI - Adaptation to salt stress in a salt-tolerant strain of the yeast Yarrowia lipolytica. AB - We have studied the cellular mechanisms underlying adaptation to salt stress in a newly isolated osmo- and salt-tolerant strain of the yeast Yarrowia lipolytica. When cells are incubated in the presence of 9% NaCl, a rapid change in their size and shape is observed. Salt stress is accompanied by an increase in the intracellular level of glycerol, free amino acids (notably proline and aliphatic amino acids), and Na+, as well as by changes in lipid and fatty acid composition. PMID- 10521724 TI - Cellulase complex from Chaetomium cellulolyticum: isolation and properties of major components. AB - Four major components of the cellulase complex of Chaetomium cellulolyticum have been isolated by gel-filtration, ion-exchange chromatography on DEAE-Toyopearl and Macro Prep Q, and chromatofocusing on Mono P. These components include three endoglucanases (19, 35, and 40 kD) and a cellobiohydrolase (45 kD). The isoelectric points of the enzymes vary from 3.8 to 4.2. The optimal pH values for catalytic activity are in the range 4.5-6.0, and the optimal temperatures are in the range 60-70 degrees C. Of these enzymes the 19 kD endoglucanase is the most stable; it retained high activity within a broad pH range (from 5.0 to 9.6) at 50 degrees C for 3 h. This enzyme also had the highest topolytic activity determined by the efficiency of removal of indigo from the surface of cotton. PMID- 10521725 TI - Reactive sites of the 21-kD protein inhibitor of serine proteinases from potato tubers. AB - The effect of modifications of Met, Arg, and Lys residues on the inhibitory activity of a serine proteinase-inhibiting 21-kD protein from potato tubers has been studied. The data indicate that the 21-kD protein has two independent reactive sites for human leukocyte elastase (or chymotrypsin) and trypsin. It is concluded that the 21-kD inhibitor has Met and Arg residues in the P1 position of the reactive sites responsible for interactions with elastase (or chymotrypsin) and trypsin. It is shown that the 21-kD protein is capable of forming a triple complex binding simultaneously one molecule of trypsin and one molecule of chymotrypsin. PMID- 10521726 TI - Comparative structural-functional characterization of recombinant and natural adrenodoxin. Interaction with cytochrome P450scc. AB - The conditions for heterologous expression of recombinant bovine adrenodoxin in E. coli have been optimized, thus reaching expression levels up to 12-14 micromoles per liter of culture medium. A highly efficient method for purification of this recombinant ferredoxin from the E. coli cells has been developed. The structural-functional properties of the highly purified recombinant protein have been characterized and compared to those of natural adrenodoxin purified from bovine adrenocortical mitochondria. In contrast to natural adrenodoxin, which is characterized by microheterogeneity, the recombinant adrenodoxin is homogeneous as judged by N- and C-terminal amino acid sequencing, and its sequence corresponds to the full-length mature form of adrenodoxin containing 128 amino acid residues. The interactions of the natural and recombinant adrenodoxins with cytochrome P450scc have been studied and compared with respect to: the efficiency of their enzymatic reduction of cytochrome P450scc in a reconstituted system; the ability of the immobilized adrenodoxins to bind cytochrome P450scc; the ability of the adrenodoxins to induce a spectral shift of cytochrome P450scc and to effect the average polarity of exposed tyrosines in the low-spin cytochrome P450scc. The recombinant adrenodoxin is functionally active and in the reduced state as well as at low ionic strength it displays higher affinity to cytochrome P450scc as compared to the natural bovine adrenocortical adrenodoxin. The possible role of the C terminal sequence of the adrenodoxin molecule in its interaction with cytochrome P450scc as well as the advantages of using the recombinant protein instead of the natural one are discussed. PMID- 10521728 TI - Transperineal versus endovaginal ultrasonographic examination of the cervix in the midtrimester: a blinded comparison. AB - OBJECTIVE: Shortening of the cervix that occurs early in gestation as determined by ultrasonography has been correlated with subsequent spontaneous preterm birth. Because previous studies have compared transperineal and endovaginal ultrasonographic assessment of the cervix over a wide gestational age range, we compared these methods in the midtrimester. STUDY DESIGN: We performed a prospective, blinded comparison of endovaginal and transperineal cervical ultrasonographic assessment. Unselected gravid women at 15-23 weeks' gestation were sequentially examined by 2 experienced sonographers using an endovaginal probe and a curvilinear probe. For each patient the initial sonographer and examination method were randomly assigned. The sonographer who performed the second examination was blinded to the results and images from the first examination. RESULTS: One hundred two women were studied at a mean of 19.6 weeks' gestation. The overall intermethod correlation was poor (R = 0.38). In 12 cases a transperineal measurement could not be obtained because of poor visualization of the required landmarks. In 33% of cases the intermethod difference in cervical length was >/=20%. There was no correlation in the identification of funneling at the internal os and very small correlation for the identification of a poorly developed lower uterine segment. CONCLUSION: Transperineal ultrasonographic imaging of the cervix is an unsatisfactory alternative to an endovaginal assessment in the midtrimester. PMID- 10521727 TI - A systemic fetal inflammatory response and the development of bronchopulmonary dysplasia. AB - OBJECTIVE: The purpose of this study was to test the hypothesis that a systemic fetal inflammatory response is a risk factor for the subsequent development of bronchopulmonary dysplasia in preterm neonates. STUDY DESIGN: The relationship between interleukin 6 concentrations in umbilical cord plasma at birth and the occurrence of bronchopulmonary dysplasia was examined in 203 preterm births (25 34 weeks). Ninety-six patients underwent transabdominal amniocentesis within 5 days of birth. The relationship between umbilical cord plasma interleukin 6 concentration and bronchopulmonary dysplasia was compared with the relationship between amniotic fluid interleukin 6 concentration and bronchopulmonary dysplasia. Interleukin 6 was measured by specific immunoassay. Logistic regression was used for statistical analysis. RESULTS: Bronchopulmonary dysplasia was diagnosed in 17% (34/203) of the infants. Neonates in whom bronchopulmonary dysplasia developed had a significantly higher median interleukin 6 concentration in umbilical cord plasma at birth than did those in whom bronchopulmonary dysplasia did not develop (median, 68.3 pg/mL and range, 0.3-6150.0 pg/mL vs median, 6.9 pg/mL and range 0-19,230.0 pg/mL; P <.001). This difference remained significant after adjustment for gestational age at birth (odds ratio, 4.2; 95% confidence interval, 1.6-11.2). Logistic regression analysis indicated that an elevated umbilical cord plasma interleukin 6 concentration was a better predictor of the development of bronchopulmonary dysplasia than was an elevated amniotic fluid interleukin 6 concentration (P <.005). CONCLUSION: An elevated interleukin 6 concentration in umbilical cord plasma at birth is an independent risk factor for the development of bronchopulmonary dysplasia. These data support the concept that the injury responsible for bronchopulmonary dysplasia in a subset of neonates may begin before birth and is associated with the development of a fetal systemic inflammatory response, as determined by plasma concentrations of interleukin 6. PMID- 10521729 TI - Association of oligohydramnios in women with preterm premature rupture of membranes with an inflammatory response in fetal, amniotic, and maternal compartments. AB - OBJECTIVE: This study was undertaken to examine whether oligohydramnios in women with preterm premature rupture of membranes is associated with evidence of fetal, amniotic, and maternal inflammatory responses. STUDY DESIGN: Amniotic fluid index was measured before the performance of amniocentesis in patients with preterm premature rupture of membranes. Fifty-nine patients who were delivered of preterm neonates (gestational age 5 cm (positive amniotic fluid culture result, 79% [15/19] vs 30% [12/40]; clinical chorioamnionitis, 37% [7/19] vs 5% [2/40]; histologic chorioamnionitis, 100% [17/17] vs 69% [24/35]; median amniotic fluid interleukin 6 concentration, 13.5 ng/mL; range, 0.2-142.2 ng/mL vs 3.0 ng/mL and 0.001-115.2 ng/mL; median amniotic fluid interleukin 1beta concentration, 348.0 pg/mL; range, 0.7->80, 000 pg/mL vs 36.6 pg/mL and 0-2075 pg/mL; median amniotic fluid tumor necrosis factor alpha concentration, 132.0 pg/mL; range, 0-1600 pg/mL vs 11.2 pg/mL and 0-1305 pg/mL; median cord plasma interleukin 6 concentration, 49.7 pg/mL; range, 4.4-7400 pg/mL vs 9. 1 pg/mL and 0-5211 pg/mL; P <.05 for each). There was no significant difference between the 2 groups of patients in the mean umbilical artery pH at birth. CONCLUSION: Oligohydramnios in women with preterm premature rupture of membranes is associated with an inflammatory response in the fetal, amniotic, and maternal compartments. PMID- 10521730 TI - A five-year experience with fragile X screening of high-risk gravid women. AB - OBJECTIVE: We sought to compare our 5-year program of fragile X screening of high risk gravid women with our program of fragile X testing of affected individuals (probands). STUDY DESIGN: All women referred to the prenatal genetics clinic from 1994 to 1998 who had a family history of unspecified mental retardation or learning or behavioral disorders (known fragile X families excluded) were offered fragile X screening. Results were compared with those of probands with the same diagnoses who underwent fragile X testing during the same time period. RESULTS: We counseled 12,349 prenatal patients from 1994-1998, of whom 263 (2.1%) had a positive family history and underwent fragile X screening. No mutations or premutations were identified. In contrast, 31 (1.9%) of 1637 affected probands who underwent fragile X testing during the same time period had positive results, which was a significant difference (0/263 vs 31/1637; P <.05). CONCLUSIONS: Testing the affected proband is superior to screening the pregnant relative of the proband for identification of families at risk for fragile X syndrome. PMID- 10521731 TI - The relationship between placental histology and cervical ultrasonography in women at risk for pregnancy loss and spontaneous preterm birth. AB - OBJECTIVE: Our objective was to determine whether there were any differences in the placental lesions of high-risk patients with versus without ultrasonographic evidence of cervical shortening between 15 and 24 weeks' gestation. STUDY DESIGN: Women who were at risk for pregnancy loss and spontaneous preterm birth were followed by serial transvaginal cervical ultrasonography with transfundal pressure between 15 and 24 weeks' gestation. Two groups of women were identified: those in whom progressive cervical shortening developed to below 2 cm, either spontaneously or induced by transfundal pressure, and those in whom it did not. A perinatal pathologist who was blinded to the pregnancy outcome retrospectively examined placental histologic slides. The histologic placental lesions were categorized as acute or chronic inflammatory lesions, decidual vascular lesions, and coagulation-related lesions. RESULTS: There were 278 women who were followed during the study. Placentas were submitted for histologic examination in 189 cases (125 singleton, 45 twin, and 19 triplet gestations). There were 72 pregnancies with and 117 pregnancies without an ultrasonographic diagnosis of cervical shortening, respectively. Overall, there were significantly more acute inflammatory lesions in patients in whom cervical shortening developed, as determined by ultrasonographic examination. However, there were significantly more decidual vascular lesions in women in whom cervical shortening did not develop. When we examined the distribution of the placental histologic lesions in the 64 cases of multiple gestations, the only significant finding was again a greater frequency of acute inflammatory lesions in patients in whom cervical shortening developed. There was no difference in the distribution of the placental histologic lesion categories among women treated with bed rest versus cervical cerclage because of the ultrasound diagnosis of cervical shortening. CONCLUSION: Acute inflammatory lesions of the placenta were more frequent in patients with second-trimester cervical shortening. These findings support that patients with cervical shortening in the second trimester are prone to acute placental inflammation. PMID- 10521732 TI - Universal versus selective gestational diabetes screening: application of 1997 American Diabetes Association recommendations. AB - OBJECTIVE: We sought to evaluate the impact of the 1997 American Diabetes Association gestational diabetes mellitus screening guidelines applied to a universally screened population. STUDY DESIGN: A retrospective analysis of 18,504 women universally screened for gestational diabetes mellitus at Mayo Clinic, Rochester, between January 1, 1986, and December 31, 1997, was performed. Diabetic screening consisted of plasma glucose determination 1 hour after a 50-g oral glucose challenge. Diagnosis of gestational diabetes mellitus was based on National Diabetes Data Group criteria. RESULTS: Of 564 cases of gestational diabetes mellitus diagnosed during the study period, 17 (3.0%) would have been missed under the 1997 American Diabetes Association selective screening guidelines while exempting only 10% of this predominantly white population from screening. Screening only women >/=25 years old would have detected 90.4% of gestational diabetes mellitus cases, whereas the addition of the remaining 3 screening criteria combined would have detected only an additional 6.6% of cases. CONCLUSIONS: The proportion of patients with gestational diabetes mellitus that would remain undiagnosed under the 1997 American Diabetes Association screening guidelines would be relatively small in our population. However, implementation of these guidelines would decrease the number of screens by only 10% while adding significant complexity to the screening process. Youth appears to be the most significant protective factor for gestational diabetes mellitus in our population. PMID- 10521733 TI - A randomized trial of conjugated group B streptococcal type Ia vaccine in a rabbit model of ascending infection. AB - OBJECTIVE: Maternal vaccination may become a central strategy in the prevention of early-onset group B Streptococcal sepsis. Unlike earlier group B streptococcal polysaccharide vaccines that were poorly immunogenic, newer vaccines conjugated to tetanus toxoid have been developed and have improved immunogenicity. We sought to evaluate a conjugated vaccine using our rabbit model of ascending infection. STUDY DESIGN: Rabbit does were randomized to receive either conjugated group B streptococcal type Ia (Ia-tetanus toxoid) or conjugated group B streptococcal type III (III-tetanus toxoid) vaccine. Does were vaccinated 7 days before conception and 7 and 21 days after conception. On days 28 to 30 of a 30-day gestation, does were inoculated intracervically with 10(6) colony-forming units of type Ia group B Streptococcus. Labor was induced if does were undelivered after 72 hours. Does were observed up to 7 days after inoculation. Offspring were observed up to 4 days. We obtained maternal cultures from the uterus, peritoneum, and blood and offspring cultures from the mouth, anus, and blood. Antibody levels were also determined. RESULTS: Offspring survival was significantly improved in the group receiving Ia-tetanus toxoid (P =.047). Outcomes such as maternal sepsis and severe illness, although not reaching statistical significance, showed a trend toward improved outcomes in the Ia-tetanus toxoid group. CONCLUSIONS: This is the first study to evaluate the conjugated group B streptococcal vaccine by using any model of ascending infection. The Ia-tetanus toxoid vaccine led to improved survival and was immunogenic but fell short of its expected efficacy in preventing ascending group B streptococcal disease under these experimental conditions. PMID- 10521734 TI - Prediction of preterm delivery with transvaginal ultrasonography of the cervix in patients with high-risk pregnancies: does cerclage prevent prematurity? AB - OBJECTIVES: We sought to determine the predictive accuracy for preterm delivery of transvaginal ultrasonography of the cervix between 14 and 24 weeks' gestation in high-risk patients and to determine whether cerclage prevents preterm delivery in patients with ultrasonographic cervical changes. STUDY DESIGN: Patients with asymptomatic singleton pregnancies at high risk for preterm delivery were followed prospectively from 14 weeks' to 23 weeks 6 days' gestation with transvaginal ultrasonography of the cervix. The subgroup of patients with either a cervical length of <25 mm or funneling of >25% or both was offered McDonald salvage cerclage, which was performed at the discretion of the patient and the obstetrician. The 2 groups (with and without cerclage) were compared for the primary outcome of preterm delivery at <35 weeks' gestation. RESULTS: One hundred sixty-eight women were followed, including 97 (58%) with >/=1 prior 14- to 34 week preterm deliveries. Of 63 (37. 5%) patients identified as having cervical changes, 23 (37%) had preterm delivery; of 105 patients with no cervical changes, 8 (8%) had preterm delivery (relative risk, 4.8; 95% confidence interval, 2. 3 10.1). The sensitivity, specificity, and positive and negative predictive values of either a short cervix of <25 mm or funneling of >25% or both were 74%, 70%, 37%, and 92%, respectively. Of 63 pregnancies in which there were cervical changes, 39 underwent cerclage and 24 did not. These 2 groups were similar for demographic characteristics, risk factors, and transvaginal ultrasonographic cervical length and funneling but dissimilar for gestational age at identification of cervical changes (18.3 vs 21.2 weeks' gestation in the groups with and without cerclage, respectively; P <.001). Multivariate logistic regression analysis after adjustment for gestational age at cervical changes showed no difference in the rate of preterm delivery between the groups with and without cerclage (odds ratio, 1.1; 95% confidence interval, 0.3-4.6). Stratified analysis of patients identified between 18 and 24 weeks revealed 22 pregnancies with cerclage and 22 pregnancies without cerclage, which was similar for all characteristics studied. The incidence of preterm delivery remained similar (27% vs 23%, respectively; P =.7), as did days from cervical changes to delivery (111 vs 96, respectively; P =.2). CONCLUSIONS: Transvaginal ultrasonography of the cervix between 14 and 24 weeks' gestation is a good predictor of preterm delivery in high-risk pregnancies. Cerclage may not prevent preterm delivery in patients identified to be at high risk for this outcome by transvaginal ultrasonography. PMID- 10521735 TI - Antibiotic use in pregnancy and drug-resistant infant sepsis. AB - OBJECTIVE: We sought to evaluate the effect of antepartum and intrapartum antibiotic use on antimicrobial-resistant neonatal sepsis. STUDY DESIGN: We analyzed perinatal outcomes for 8474 pregnancies (8593 live births) delivered at 6 hospitals. Data were collected regarding maternal antibiotic use and perinatal course, neonatal cultures, and outcomes. The diagnosis of confirmed neonatal sepsis required at least one positive blood or cerebrospinal fluid culture. Neonatal cultures were evaluated on the basis of the occurrence and timing of maternal antibiotic exposure. RESULTS: There were 96 neonates with confirmed sepsis (11.2/1000 live births). Sepsis was 19.3-fold more common after preterm birth (57 vs 3. 1/1000; P <.001), with 76% of septic infants being delivered preterm. Forty-five percent of pathogens were ampicillin resistant. Ampicillin resistance increased with preterm birth (50% vs 26%; P =. 04), antepartum antibiotics (57% vs 34%; P =.03), intrapartum antibiotics (55% vs 28%; P <.01), and any prenatal antibiotic exposure (52% vs 22%; P =.01). Infection with an organism resistant to at least one maternal antibiotic was more common with intrapartum antibiotic exposure than with antepartum exposure only (57% vs 17%; P =.01). Regarding early-onset sepsis (n = 55), ampicillin resistance was more common with intrapartum antibiotics (50% vs 16%; P <.01), and resistance to at least one maternally administered antibiotic was more frequent with intrapartum exposure (56.7% vs 0%; P <.01). CONCLUSIONS: Maternal antibiotic treatment is associated with neonatal sepsis by organisms resistant to ampicillin and to maternally administered antibiotics. PMID- 10521736 TI - Maintenance oral nifedipine for preterm labor: a randomized clinical trial. AB - OBJECTIVE: This study was undertaken to evaluate the efficacy of maintenance oral nifedipine in patients initially treated with intravenous magnesium sulfate for preterm labor. STUDY DESIGN: Patients with a diagnosis of preterm labor between 24 and 33.9 weeks' gestation were randomly assigned to receive either maintenance tocolytic therapy with oral nifedipine (20 mg every 4-6 hours) or no treatment (control) after discontinuation of magnesium tocolysis. Pregnancy and neonatal outcomes were evaluated. A sample size of 50 patients was required to detect a 10 day difference in mean time gained (beta =.2, alpha =.05). Statistical analyses were based on intent to treat. The t, chi(2), and Fisher exact tests were performed. RESULTS: Seventy-four patients were randomly assigned to receive either oral nifedipine (n = 37) or no treatment (n = 37). There were no statistically significant differences in age, race, parity, preterm delivery risk factors, enrollment gestational age, results of cervical examination, delivery gestational age, time gained, or neonatal complications between the groups. Delivery gestational age (mean +/- SD) was 35.4 +/- 3.2 weeks for patients randomly assigned to receive nifedipine and 35.3 +/- 3.2 weeks for patients who received no treatment (P =.9). Time gained during pregnancy was 37 +/- 23.9 days in the nifedipine group and 32.8 +/- 20.4 days in the control group (P =.4). CONCLUSION: Maintenance therapy with oral nifedipine does not significantly prolong pregnancy in patients initially treated with intravenous magnesium sulfate for preterm labor. PMID- 10521737 TI - Fetal rat brain damage caused by maternal seizure activity: prevention by magnesium sulfate. AB - OBJECTIVE: Our purpose was to determine whether maternal rat seizure activity was associated with fetal histopathologic brain changes and whether magnesium sulfate reduced these changes. STUDY DESIGN: Electrodes were stereotaxically implanted into the hippocampus of nonpregnant rats 1 week before breeding. Pregnant rats were randomly assigned to 1 of 4 groups: (1) sodium chloride solution and no seizure (n = 2), (2) magnesium sulfate and no seizure (n = 2), (3) sodium chloride solution and seizure (n = 5), and (4) magnesium sulfate and seizure (n = 5). On gestational days 9, 11, 13, 15, 17, and 19, subcutaneous doses of sodium chloride solution or magnesium sulfate were administered to all rats every 20 minutes for 4 hours (loading-maintenance-loading), followed by seizure induction. On gestational day 20, the rats were perfused with formalin and fetuses were delivered via cesarean. Fetuses were perfused with formalin, brains were obtained and embedded in paraffin, and the forebrain and hindbrain were sectioned in the coronal plane and stained with hematoxylin and eosin. A neuropathologist masked to the protocol performed histopathologic grading of each section, including extent and nature of cellular damage. Eleven brain regions were examined in each section. Scores were expressed as mean +/- SD. Kruskal-Wallis analysis of variance was used, and P <.05 was considered significant. RESULTS: We evaluated 26 fetal brains in group 1, 9 in group 2, 72 in group 3, and 45 in group 4. Fetuses in the sodium chloride solution-and-seizure group (group 3) presented significantly higher grades of neuronal damage in the hippocampus (group 1, 0.50 +/- 0. 88; group 2, 0.22 +/- 0.66; group 3, 1.01 +/- 1.17; and group 4, 0. 48 +/- 0.72) and in the tegmentum region (group 1, 1.0 +/- 1.0; group 2, 0.8 +/- 1.0; group 3, 1.7 +/- 0.7; and group 4, 1.5 +/- 0. 8) (P <.05, group 3 compared with others). Isolated and patchy neuronal injury with shrinkage of cells, nuclear pyknosis, and karyorrhexis were the main histologic findings. CONCLUSIONS: Maternal rat seizure activity was associated with histologic brain injury in the fetus. Maternal administration of magnesium sulfate before seizure prevented or significantly decreased this effect. PMID- 10521738 TI - Prediction by maternal risk factors of neonatal intensive care admissions: evaluation of >59,000 women in national managed care programs. AB - OBJECTIVE: Managed care plans have adopted risk assessment tools as part of pregnancy disease state management strategies to assist in reducing poor pregnancy outcomes and related costs. We evaluated the relationships of maternal risk factors to determine which pregnancy risk factors were associated with neonatal intensive care unit (levels II and III) admission. STUDY DESIGN: Risk assessments were performed through perinatal telephone interviews of nurses with 59, 861 pregnant women during 1996 and 1997 calendar years as part of managed care maternity risk screening and education programs. A series of 3 interviews was conducted, at 17 weeks and 28 weeks average gestational age and at 2 weeks post partum. Univariate chi(2) analysis was performed on >50 historical and pregnancy risk factors to determine the associations with neonatal intensive care unit admission. Significant factors were included in a stepwise logistic regression model. Receiver operating curves were generated for the use of significant factors in a risk scoring system in the prediction of neonatal intensive care unit admission, and the percentages of neonatal intensive care unit days attributable to significant risk factors were calculated. RESULTS: Among the participants most women (90%) had their prenatal visit during the first trimester. The mean maternal age was 30.2 +/- 5.2 years, with 74% of women reportedly of white ethnicity, 86% married, and 44.3% primigravid. The mean gestational age at birth decreased with increasing number of fetuses from singletons to quadruplets. The chi(2) analysis identified 26 significant risk factors associated with neonatal intensive care unit admission. Of these, 14 remained significant by logistic regression. Multiple gestation, preterm premature rupture of membranes, diabetes, abruptio placentae, pregnancy-induced hypertension, and preterm labor were independently associated with at least a 3 fold risk of neonatal intensive care unit admission. A modeled risk scoring system that used these and other significant factors was poorly predictive of neonatal intensive care unit admission. However, an analysis of neonatal intensive care unit length of stay attributable to significant risk factors concluded that 19% of all neonatal intensive care unit days in this population were associated with multiple gestations. Furthermore, 85% of neonatal intensive care unit days were the result of infant lengths of stay >/=1 week. CONCLUSION: This analysis of a managed care population showed similar risk factors to those traditionally associated with neonatal intensive care unit admission. Although many of these risk factors are not preventable, identification of neonatal intensive care unit admission risks with a screening program may be of use for focusing interventions, and earlier identification of these factors may allow maximum impact of interventions. Importantly, a reduction in the incidence of higher-order multiple gestations might help to reduce neonatal intensive care unit admissions and costs. PMID- 10521739 TI - Prenatal diagnosis of congenital toxoplasmosis: a multicenter evaluation of different diagnostic parameters. AB - OBJECTIVE: Our purpose was to evaluate different methods of diagnosing congenital toxoplasmosis prenatally by amniocentesis and cordocentesis. STUDY DESIGN: In a retrospective multicenter study, we investigated consecutive women who had seroconversion for Toxoplasma gondii during pregnancy and who underwent either amniocentesis or cordocentesis or both to obtain a prenatal diagnosis of fetal toxoplasmosis. Data were obtained from 122 patients recruited in 6 different European Toxoplasma reference centers. Infants born to these mothers were followed up until 1 year of age to confirm or exclude congenital toxoplasmosis. Sensitivity, specificity, positive predictive value, and negative predictive value were measured for the following parameters: (1) detection of the parasite in amniotic fluid by mouse inoculation, (2) detection of the parasite in amniotic fluid by in vitro cell culture, (3) detection of Toxoplasma deoxyribonucleic acid in amniotic fluid by a polymerase chain reaction assay, (4) detection of the parasite in fetal blood by mouse inoculation, (5) detection of specific immunoglobulin M antibodies in fetal blood, and (6) detection of specific immunoglobulin A antibodies in fetal blood. RESULTS: The polymerase chain reaction test performed on amniotic fluid had the highest level of sensitivity (81%) and also a high level of specificity (96%). The combination of the polymerase chain reaction test and mouse inoculation of amniotic fluid increased sensitivity to 91%. The sensitivity of immunoglobulins M and A in fetal blood was 47% and 38%, respectively. In congenitally infected fetuses a negative correlation was observed between positive serologic parameters and gestational age at the time of maternal infection and at prenatal diagnosis. CONCLUSION: Congenital toxoplasmosis is best predicted by prenatal examination with the combination of T gondii polymerase chain reaction and mouse inoculation of amniotic fluid. The role of cordocentesis in the diagnosis of congenital toxoplasmosis is limited. PMID- 10521740 TI - Ultrasonographically guided direct gene transfer in utero: successful induction of beta-galactosidase in a rabbit model. AB - OBJECTIVE: We sought to determine whether the transfer of enzyme-encoding genes in utero can be detected after birth. STUDY DESIGN: An adenoviral vector carrying the gene for beta-galactosidase was injected under ultrasonographic guidance into the livers of 4 rabbit fetuses per litter (3 litters total) at 27 days' gestation. On delivery of the pups 2 to 3 days later, the livers were analyzed for beta-galactosidase activity by using 5-bromo-4-chloro-3-indolyl-beta-D galactopyranoside (X-gal) staining. Polymerase chain reaction was also performed on liver extracts as an additional independent measure of successful vector delivery. RESULTS: Successful targeting of the livers of fetal rabbits was demonstrated by beta-galactosidase activity in the nuclei of liver serosal cells, parenchymal hepatocytes, or columnar cells of the gallbladder in 7 (58%) of 12 injected pups and by polymerase chain reaction in liver extracts from 10 (83%) of 12 injected pups. CONCLUSIONS: These results suggest that vectors that carry genes for specific enzymes can be delivered to fetal organs in utero and that expression of the enzyme can be detected after delivery. PMID- 10521741 TI - Human chorionic gonadotropin exhibits potent inhibition of preterm delivery in a small animal model. AB - OBJECTIVE: The purpose of this study was to test the capability of human chorionic gonadotropin to inhibit prostaglandin-induced preterm delivery in a murine model. STUDY DESIGN: A preterm delivery model was developed by using intraperitoneal injection of 20 microgram of prostaglandin F(2)(alpha) to induce preterm labor in C3H/HeN inbred mice. Mice were then pretreated with human chorionic gonadotropin 4 hours before administration of prostaglandin F(2)(alpha), and time to delivery of the first pup was recorded. After initial promising results, mice were then given increasing intraperitoneal doses of human chorionic gonadotropin (100 IU, 250 IU, or 1000 IU or sodium chloride solution vehicle) 4 hours after administration of prostaglandin F(2)(alpha). The specificity of the human chorionic gonadotropin effect was assessed by treating mice with whole human chorionic gonadotropin, an equal mass dose of the beta subunit or the alpha-subunit of human chorionic gonadotropin, or an equal mass dose of luteinizing hormone 4 hours after administration of prostaglandin F(2)(alpha). Delivery times between groups were compared by using the Mann Whitney U test and the log-rank test. Survival estimates were computed by using the Kaplan-Meier method. RESULTS: Pilot studies in 52 mice confirmed that a single intraperitoneal injection of 20 microgram of prostaglandin F(2)(alpha) on day 16 (80% gestation) consistently induced preterm delivery compared with the effect of sodium chloride solution on control mice (prostaglandin F(2)(alpha), 19.3 +/- 2.9 hours; sodium chloride solution, 53.5 +/- 13.6 hours; P <.0001). Mice pretreated with human chorionic gonadotropin (1000 IU) demonstrated significant delays in delivery times compared with the prostaglandin-only group (prostaglandin F(2)(alpha) only, 21.9 +/- 2. 0 hours; human chorionic gonadotropin pretreatment plus prostaglandin F(2)(alpha), 48.5 +/- 20 hours; P <.0001; n = 17). Mice treated with human chorionic gonadotropin (100 IU, 250 IU, 1000 IU) 4 hours after administration of prostaglandin F(2)(alpha) demonstrated significant dose-dependent inhibition of preterm delivery compared with the prostaglandin-only group (P <.00005; n = 34). Mice treated with the alpha-subunit or the beta-subunit of human chorionic gonadotropin after prostaglandin administration did not demonstrate delays in delivery times (P =.46; n = 27). Administration of luteinizing hormone delayed delivery compared with the effect of prostaglandin F(2)(alpha) on control animals (P <.05; n = 17); however, the effect was less pronounced than that seen with a mass equivalent of human chorionic gonadotropin. CONCLUSIONS: Human chorionic gonadotropin exhibits potent inhibition of prostaglandin-induced preterm delivery in mice. The effect is dose dependent, and whole human chorionic gonadotropin is required to elicit inhibition. Further studies are needed to determine the safety and efficacy of human chorionic gonadotropin as a potential therapy for preterm labor inhibition in human pregnancy. PMID- 10521742 TI - A randomized, double-blind trial of oral nifedipine and intravenous labetalol in hypertensive emergencies of pregnancy. AB - OBJECTIVE: We sought to compare the efficacies of oral nifedipine and intravenous labetalol in the acute management of hypertensive emergencies of pregnancy. STUDY DESIGN: We performed a randomized double-blind trial of oral nifedipine (10 mg) and intravenous labetalol (20 mg) in 50 peripartum patients with sustained systolic blood pressure of >/=170 mm Hg or diastolic blood pressure of >/=105 mm Hg. Both agents were repeated at sequentially escalating dosages every 20 minutes until a therapeutic goal of systolic blood pressure of <160 mm Hg and diastolic blood pressure of <100 mm Hg was achieved. Crossover occurred if the treatment goal was not achieved after 5 doses. Primary outcome was time to achievement of the therapeutic goal. Secondary outcome variables were agent failure, urinary output, and adverse effects. Data were analyzed by unpaired t test, Mann-Whitney U test, and analysis of variance for repeated measures. RESULTS: The time to achieve the blood pressure goal was significantly shorter with nifedipine (mean +/- SD, 25 +/- 13.6 minutes) than with labetalol (43.6 +/- 25.4 minutes; P =.002). No patients required crossover therapy. Urine output was significantly increased (P <.001) at 1 hour after nifedipine dosing (99 +/- 99 mL) compared with labetalol (44.8 +/- 19.1 mL) and remained significantly increased at 2, 6, 12, 18, and 24 hours after initial administration. Adverse effects were infrequent. There were no differences in maternal age, gestational age, number of antepartum patients, or enrollment blood pressures between groups. CONCLUSIONS: Both oral nifedipine and intravenous labetalol are effective in the management of acute hypertensive emergencies of pregnancy; however, nifedipine controls hypertension more rapidly and is associated with a significant increase in urinary output. PMID- 10521743 TI - A randomized, double-blind, hemodynamic evaluation of nifedipine and labetalol in preeclamptic hypertensive emergencies. AB - OBJECTIVE: Our purpose was to compare the hemodynamic effects of orally administered nifedipine and intravenously administered labetalol in preeclamptic hypertensive emergencies. STUDY DESIGN: Our study was a randomized, double-blind evaluation of nifedipine and labetalol in women with preeclampsia and a systolic blood pressure >170 mm Hg or a diastolic blood pressure >105 mm Hg. Nifedipine or labetalol and placebo were given, so patients received both tablet and intravenous solution. Hemodynamic parameters at dosing and at 15, 30, 60, and 120 minutes were recorded. Outcome measures were cardiac index, systemic vascular resistance index, mean arterial pressure, and heart rate. Data were analyzed by repeated-measures analysis of variance (Friedman test) with Dunn posttests, the Mann-Whitney U test, and the chi(2) test with the Yates correction. Significance was set at P <.05. RESULTS: At dosing, the nifedipine group (n = 6) had a cardiac index of 3.08 +/- 0.51 L/min per square meter. There was a 43% increase in the cardiac index after nifedipine administration (P =.0008). There was no significant effect in the labetalol group (P =.697). There was a significant decrease in the systemic vascular resistance index after nifedipine dosing (P =.002) but no significant effect on this index after labetalol use (P =.479). The mean arterial pressure was significantly affected in both groups as follows: nifedipine, P =. 001; labetalol, P =.004. The postanalysis showed significance at 60 minutes for both. An insignificant increase in heart rate with nifedipine (P =.147) and a significant decrease with labetalol (P =. 034) were noted. CONCLUSIONS: Nifedipine increases cardiac index, whereas labetalol may not do so. PMID- 10521744 TI - Association between umbilical blood gas parameters and neonatal morbidity and death in neonates with pathologic fetal acidemia. AB - OBJECTIVE: Our purpose was to correlate umbilical artery blood gas parameters with neonatal death and indicators of morbidity in neonates with pathologic fetal acidemia (pH <7.0). STUDY DESIGN: We reviewed maternal and neonatal charts of 93 neonates with an umbilical artery pH <7.0 who were delivered at 2 university based centers. The relationships between umbilical artery pH, PO (2), PCO (2), bicarbonate, base deficit, and neonatal variables-death, need for intubation, cardiopulmonary resuscitation, seizures, hypoxic-ischemic encephalopathy, respiratory distress syndrome, intraventricular hemorrhage, meconium, sepsis, and intrauterine growth restriction-were determined with the Student t test, Mann Whitney U test, and multiple logistic regression analysis. Data are presented as either median with 25th-75th percentiles or mean +/- SD. RESULTS: The mean gestational age at delivery was 37.9 +/- 3. 6 weeks, and the mean birth weight was 3003 +/- 866 g. There was no relationship between neonatal death, respiratory distress syndrome, intraventricular hemorrhage, necrotizing enterocolitis, patent ductus arteriosus, meconium, sepsis, and any umbilical artery blood gas parameter. The PO (2) was not related to any of the variables studied. A lower umbilical artery pH was associated with hypoxic-ischemic encephalopathy (6.69 vs 6.93, P =.03), cardiopulmonary resuscitation (6.83 vs 6.93, P =.03), seizure (6.75 vs 6.93, P =.02), intubation (6.83 vs 6.94, P <.001), and intrauterine growth restriction (6.72 vs 6.93, P =.01). Greater mean base deficit was associated with seizure (20.6 vs 15, P =.01), intubation (18.0 vs 13.7, P <.001), cardiopulmonary resuscitation (18.5 vs 15.0, P =.03), intrauterine growth restriction (22.0 vs 14. 0, P =.02), and hypoxic-ischemic encephalopathy (24.0 vs 14.5, P =. 03). Arterial PCO (2) was higher only in infants with hypoxic-ischemic encephalopathy (138 vs 95.5, P =.048), intubation (106.0 vs 90.5, P =.003), and cardiopulmonary resuscitation (106.5 vs 93.0, P =.04). After control for birth weight and gestational age in the multivariate analysis, base deficit and bicarbonate were independently related to death or morbidity. CONCLUSION: Our data suggest that "pathologic" fetal acidemia is indicated by an umbilical artery pH <7.00 with a metabolic component. The metabolic component of fetal acidemia (ie, base deficit and bicarbonate) is the most important variable in subsequent neonatal morbidity. As expected, the umbilical artery PO (2) has no apparent clinical utility. The ability to predict more accurately which newborn infants with fetal acidemia are at risk of having complications may lead to a more efficient implementation of preventive measures. PMID- 10521745 TI - Rate of uterine rupture during a trial of labor in women with one or two prior cesarean deliveries. AB - OBJECTIVE: We sought to determine whether there is a difference in the rate of symptomatic uterine rupture after a trial of labor in women who have had 1 versus 2 prior cesarean deliveries. STUDY DESIGN: The medical records of all women with a history of either 1 or 2 prior cesarean deliveries who elected to undergo a trial of labor during a 12-year period (July 1984-June 1996) at the Brigham and Women's Hospital were reviewed. Rates of uterine rupture were compared for these 2 groups. Potential confounding variables were controlled by using logistic regression analyses. RESULTS: Women with 1 prior cesarean delivery (n = 3757) had a rate of uterine rupture of 0.8%, whereas women with 2 prior cesarean deliveries (n = 134) had a rate of uterine rupture of 3.7% (P =.001). In a logistic regression analysis that was controlled for maternal age, use of epidural analgesia, oxytocin induction, oxytocin augmentation, the use of prostaglandin E(2) gel, birth weight, gestational age, type of prior hysterotomy, year of trial of labor, and prior vaginal delivery, the odds ratio for uterine rupture in those patients with 2 prior cesarean deliveries was 4.8 (95% confidence interval, 1.8 13. 2) CONCLUSIONS: Women with a history of 2 prior cesarean deliveries have an almost 5-fold greater risk of uterine rupture than those with only 1 prior cesarean delivery. PMID- 10521746 TI - Incidence of uterine rupture among women with mullerian duct anomalies who attempt vaginal birth after cesarean delivery. AB - OBJECTIVE: The purpose of this study was to determine and compare the incidences of uterine rupture among women with and without mullerian duct anomalies who were attempting vaginal birth after cesarean delivery. STUDY DESIGN: There were 1813 attempts at vaginal birth after cesarean delivery between 1992 and 1997 at the Foothills Hospital in Calgary, Alberta, Canada. Of the patients 25 had known mullerian duct anomalies and 1788 did not. The records of these 1813 women were reviewed with respect to uterine rupture, other complications, mode of delivery, and characteristics of the trial of labor. Comparisons were made with the Fisher exact test. RESULTS: The rates of uterine rupture were 8% (2/25) in the group with mullerian duct anomalies and 0.61% (11/1788) in the group without mullerian duct anomalies (P =.013). The cesarean delivery rates were 20% (5/25) and 25.1% (448/1788), respectively. All cesarean deliveries among women with mullerian duct anomalies were performed urgently in response to severe fetal heart rate abnormalities. The rates of fetal heart rate abnormalities necessitating immediate delivery (60% vs 14.1%, P =.013), operative vaginal delivery (40% vs 19.6%, P =.02), and cord prolapse (8% vs 0.45%, P =.0076) were significantly greater in the group with mullerian duct anomalies. CONCLUSIONS: Vaginal delivery is common among women with mullerian duct anomalies who attempt vaginal birth after cesarean delivery, but the rates of uterine rupture and other complications are high. PMID- 10521747 TI - Uterine rupture during induced or augmented labor in gravid women with one prior cesarean delivery. AB - OBJECTIVE: Our purpose was to examine the risk of uterine rupture during induction or augmentation of labor in gravid women with 1 prior cesarean delivery. STUDY DESIGN: The medical records of all gravid women with history of cesarean delivery who attempted a trial of labor during a 12-year period at a single center were reviewed. The current analysis was limited to women at term with 1 prior cesarean delivery and no other deliveries. The rate of uterine rupture in gravid women within that group undergoing induction was compared with that in spontaneously laboring women. The association of oxytocin induction, oxytocin augmentation, and use of prostaglandin E(2) gel with uterine rupture was determined. Logistic regression analysis was used to examine these associations, with control for confounding factors. RESULTS: Of 2774 women in the analysis, 2214 had spontaneous onset of labor and 560 women had labor induced with oxytocin or prostaglandin E(2) gel. The overall rate of rupture among all patients with induction of labor was 2.3%, in comparison with 0.7% among women with spontaneous labor (P =.001). Among 1072 patients receiving oxytocin augmentation, the rate of uterine rupture was 1.0%, in comparison with 0.4% in nonaugmented, spontaneously laboring patients (P =.1). In a logistic regression model with control for birth weight, use of epidural, duration of labor, maternal age, year of delivery, and years since last birth, induction with oxytocin was associated with a 4.6-fold increased risk of uterine rupture compared with no oxytocin use (95% confidence interval, 1.5-14.1). In that model, augmentation with oxytocin was associated with an odds ratio of 2.3 (95% confidence interval, 0.8-7.0), and use of prostaglandin E(2) gel was associated with an odds ratio of 3.2 (95% confidence interval, 0.9-10.9). These differences were not statistically significant. CONCLUSION: Induction of labor with oxytocin is associated with an increased rate of uterine rupture in gravid women with 1 prior uterine scar in comparison with the rate in spontaneously laboring women. Although the rate of uterine rupture was not statistically increased during oxytocin augmentation, use of oxytocin in such cases should proceed with caution. PMID- 10521748 TI - Prospective evaluation of free beta-subunit of human chorionic gonadotropin and dimeric inhibin A for aneuploidy detection. AB - OBJECTIVE: Our goal was to prospectively evaluate the use of the free beta subunit of human chorionic gonadotropin and dimeric inhibin A for the detection of fetal Down syndrome and other aneuploidies. STUDY DESIGN: Women who had a second-trimester multiple-marker screening test (alpha-fetoprotein, unconjugated estriol, human chorionic gonadotropin) and genetic amniocentesis from August 1996 to August 1998 were included. Serum was also analyzed for inhibin and the free beta-subunit of human chorionic gonadotropin. Detection and false-positive rates for 4 analyte combinations at 5 different screening risk cutoff points for Down syndrome were determined and compared. RESULTS: We evaluated 1256 patients, including 23 with aneuploidy (13 with Down syndrome, 10 others). The maternal age was 35.9 +/- 4.6 years (mean +/- SD). At the optimal risk cutoff point for Down syndrome detection (1:190; false-positive rate, 19%), the multiple-marker screening test plus inhibin was superior, detecting 85% of Down syndrome cases, in comparison with 69% when the multiple-marker screening test alone was used and 62% when the other 2 combinations were used. The multiple-marker screening test plus inhibin also detected 60% of the other aneuploidies. CONCLUSIONS: When evaluated prospectively in a high-risk population, the multiple-marker screening test plus inhibin was superior to the traditional multiple-marker screening test and 2 other analyte combinations, with a lower false-positive rate and increased detection of all aneuploidies in a high-risk population. PMID- 10521750 TI - Utility of minor ultrasonographic markers in the prediction of abnormal fetal karyotype at a prenatal diagnostic center. AB - OBJECTIVE: This study was undertaken to assess the value of minor ultrasonographic markers in predicting significant karyotypic abnormalities. STUDY DESIGN: A total of 2743 fetuses (14-24 weeks' gestation) prospectively underwent a detailed ultrasonographic survey before genetic amniocentesis. Criteria for 8 minor ultrasonographic markers were established. Odds ratios for significant karyotypic abnormalities in the presence of minor ultrasonographic markers were calculated with the chi(2) and Fisher exact tests. RESULTS: Of the fetuses, 14.6% had a single minor ultrasonographic marker, 2.1% had >/=2 minor ultrasonographic markers, and 2.7% had >/=1 major ultrasonographic abnormality. One hundred four fetuses (3.8%) had an abnormal karyotype. Compared with a normal ultrasonographic examination result a single minor ultrasonographic marker increased the risk of karyotypic abnormality 5.7-fold (95% confidence interval, 3.5-9.3), whereas multiple minor markers increased the risk of an abnormal karyotype 12-fold (95% confidence interval, 5.5-26.5). When they were identified ultrasonographically in isolation, echogenic bowel, 2-vessel umbilical cord, echogenic intracardiac foci, choroidal separation, and choroid plexus cysts were statistically associated with an abnormal karyotype. When minor markers were identified in clusters of >/=2, echogenic bowel, short femur, 2-vessel umbilical cord, echogenic intracardiac foci, and mild ventriculomegaly were significantly predictive of karyotypic abnormality. With respect to the a priori aneuploidy risk of 1:26 and the a priori Down syndrome risk of 1:50, a normal ultrasonographic examination result reduced the risks to 1:67 and 1:120, respectively. The use of minor ultrasonographic markers in addition to major ultrasonographic abnormalities increased the detection of karyotypic abnormality from 27.9% to 68.3%. For trisomy 21 the sensitivity rose from 16.4% to 67. 3%. CONCLUSIONS: Significant karyotypic abnormality risk assessment by ultrasonography was greatly enhanced by the addition of minor ultrasonographic markers. Further, clusters of minor ultrasonographic markers greatly increased the likelihood of karyotypic abnormality compared with a single minor marker. A completely normal ultrasonographic examination result reduced the risk of an abnormal karyotype by 62%. Inclusion of minor ultrasonographic markers in the genetic sonogram in a high-risk population will allow the detection of 68% of fetuses with karyotypic abnormalities with a false-positive rate of 17%. PMID- 10521749 TI - Selective termination for structural, chromosomal, and mendelian anomalies: international experience. AB - OBJECTIVE: Our purpose was to evaluate the outcomes of selective termination for fetal anomalies at 8 centers with the largest known experiences worldwide. STUDY DESIGN: Outcomes in 402 cases of selective termination in pregnancies with dizygotic twins from 8 centers in 4 countries were analyzed by year, gestational age at procedure, and indication. Reductions of fetuses were as follows: 2 to 1, n = 345; 3 to 2, 39; >/=4 to 2 or 3, n = 18. Potassium chloride was used in all procedures. RESULTS: Selective termination resulted in delivery of a viable infant or infants in >90% of cases. Loss up to 24 weeks occurred in 7.1% of cases in which the final result was a singleton fetus and in 13.0% of cases in which the final result was twins. Loss was 6.6% as a result of structural abnormalities, 7.0% for chromosomal abnormalities, and 10% for mendelian abnormalities (difference not statistically significant). Loss rates for procedures were as follows: 9-12 weeks, 5.4%; 13-18 weeks, 8.7%; 19-24 weeks, 6.8%; and >/=25 weeks, 9.1% (difference not statistically significant). Mean gestational age at delivery was 35.7 weeks. No differences were seen in outcomes by maternal age. The rate of very early premature deliveries has fallen in recent years. There were no known cases of disseminated intravascular coagulation or serious maternal complications. CONCLUSION: (1) Selective termination, in the most experienced hands, can be technically performed in all 3 trimesters with good outcomes in >90% of cases. (2) The previously observed increase in second- versus first-trimester losses has diminished. (3) Third-trimester procedures, where legal, can be performed with a good outcome for the surviving fetus. PMID- 10521751 TI - When is fasting really fasting? The influence of time of day, interval after a meal, and maternal body mass on maternal glycemia in gestational diabetes. AB - OBJECTIVE: The object of the study was to determine whether time of day, interval after a standard meal, and maternal body mass influence plasma glucose concentrations in women with gestational diabetes mellitus. STUDY DESIGN: Identical mixed meals were administered on 2 separate occasions 1 week apart to 30 women with dietarily treated gestational diabetes and pregnancies between 28 and 38 weeks' gestation. One meal was administered at 7 AM (morning meal) and the other was administered at 9 PM (evening meal), each after a fast of >/=5 hours. The order of the meals (morning first versus evening first) was assigned randomly. Sixteen of the women had a body mass index >/=27 kg/m(2) (overweight) and 14 women had a body mass index <27 kg/m(2) (lean). Venous plasma concentrations of glucose, insulin, free fatty acids, beta-hydroxybutyrate, and bound and free cortisol were measured hourly for 9 hours after each of the test meals. RESULTS: When all women were considered together glucose concentrations after the morning meal were significantly greater at 1 hour, were not different at 2 hours, and were significantly lower from 3 through 9 hours postprandially than those at corresponding times after the evening meal. Plasma beta hydroxybutyrate and free fatty acid concentrations were higher between 5 and 9 hours after the morning meal than at the same times after the evening meal. Total and free cortisol levels were higher for the first 7 hours after the morning feeding, reflecting known diurnal variation in cortisol concentrations. Overweight patients' glucose values were significantly greater than those of lean subjects during the last 4 hours of the overnight fast. CONCLUSIONS: Among women with dietarily treated gestational diabetes the glucose concentrations were significantly higher from 3 to 9 hours after an evening meal, whereas suppression of free fatty acids and beta-hydroxybutyrate was less sustained after a morning feeding. The mechanisms underlying these differences remain to be determined but may involve diurnal influences of counterregulatory hormones. The relationships between measurements of maternal glycemia and maternal and perinatal outcomes in pregnancies complicated by gestational diabetes may be clarified by establishing a uniform duration of a fast and by developing meal-specific preprandial and postprandial maternal glucose targets for these patients. PMID- 10521752 TI - Gestational diabetes mellitus: metabolic and blood glucose parameters in singleton versus twin pregnancies. AB - OBJECTIVES: This study compared the frequency, glucose tolerance test results, and parameters of blood glucose control in twin and singleton pregnancies associated with gestational diabetes mellitus and carbohydrate intolerance. STUDY DESIGN: Twin and singleton pregnancies associated with gestational diabetes mellitus and carbohydrate intolerance were compared as follows: frequency, maternal age, weight, 1-hour screen, glucose tolerance test results, posttreatment blood glucose values, insulin requirement, and insulin dose. Statistical analysis included the chi(2) and Student t tests. RESULTS: Gestational diabetes mellitus was increased in twins (7.7% vs 4.1%; P <.05). The maternal weight at first visit was significantly less, and the 3-hour glucose tolerance test value was significantly greater than that for singletons. The other parameters were not different. CONCLUSIONS: There is a significant increase in the incidence of gestational diabetes mellitus and disturbance of the 3-hour glucose tolerance test in twin pregnancies. However, insulin requirements were not different, suggesting a mild disturbance of carbohydrate tolerance that was effectively managed by the strategies used to achieve blood glucose control in singletons. PMID- 10521753 TI - Expression of the placental cytokines tumor necrosis factor alpha, interleukin 1beta, and interleukin 10 is increased in preeclampsia. AB - OBJECTIVE: We sought to determine whether placental cytokine expression is altered in patients with preeclampsia. STUDY DESIGN: Whole placental tissue was collected at cesarean delivery, and total ribonucleic acid was extracted. Reverse transcriptase-polymerase chain reaction was performed to determine cytokine expression. Product bands were quantitated by scanning densitometry, and results were expressed as a ratio of cytokine/housekeeping gene (cytokine expression index). Statistical analysis was performed by the Student t test and the Mann Whitney U test. RESULTS: Placentas from 6 patients with preeclampsia and 4 normotensive patients were analyzed. Placental expression of interleukin 1beta and interleukin 10 was greater in preeclamptic women than in normotensive subjects (median interleukin 1beta cytokine expression index, 0.675; range, 0.394 0. 953; vs 0.106; range, 0.084-0.166; P =.011; median interleukin 10 cytokine expression index, 1.042; range, 0.672-1.192; vs 0.126; range, 0.062-0.398; P <.011). Tumor necrosis factor alpha messenger ribonucleic acid was detected in placentas of preeclamptic subjects but not in normotensive control subjects. CONCLUSION: Placentas from preeclamptic patients demonstrated increased expression of interleukin 1beta, interleukin 10, and tumor necrosis factor alpha. This may be in association with placental hypoxia and may contribute to the global endothelial dysfunction observed in preeclampsia. PMID- 10521754 TI - Neuropeptide Y and nitrite levels in preeclamptic and normotensive gravid women. AB - OBJECTIVES: Vascular tone is controlled largely by the sympathetic nervous system and is modulated by neuropeptide Y. Preeclampsia is linked to sympathetic overactivity. Nitric oxide can cause vasorelaxation of vessels or decrease sympathetic outflow by activating the baroreceptor reflex. Our purpose in this study was to compare serum levels of neuropeptide Y and nitrite levels in normotensive and preeclamptic gravid women. STUDY DESIGN: Twelve preeclamptic and 12 normotensive women matched for race, body mass index, parity, and gestational age were studied. Neuropeptide Y was measured by using a commercial radioimmunoassay. Nitric oxide was converted to nitrite by using metallic cadmium, and nitrite levels were determined spectrophotometrically by using a colorimetric assay. Data are presented as mean +/- SEM and were compared by using a t test. RESULTS: Neuropeptide Y levels were similar among preeclamptic and normotensive gravid women (33.8 +/- 3.0 and 32.2 +/- 3 pg/mL, respectively). Similarly, there were no differences in nitrite concentrations between preeclamptic and normotensive patients (11.6 +/- 0.8 vs 11.2 +/- 0.4 micromol/L, respectively). We also examined the ratios of neuropeptide Y and nitrite and found no correlation between preeclamptic and normotensive women. CONCLUSION: Peripheral levels of neuropeptide Y or nitrite do not correlate with preeclampsia. Assessment of sympathetic overactivity in preeclampsia requires an alternate model. PMID- 10521755 TI - Maternal mortality associated with HELLP (hemolysis, elevated liver enzymes, and low platelets) syndrome. AB - OBJECTIVE: The aim of this study was to determine factors contributing to deaths among women with HELLP (hemolysis, elevated liver enzymes, and low platelets) syndrome. STUDY DESIGN: Information from multiple sources was scrutinized to distinguish and profile maternal deaths associated with HELLP syndrome. RESULTS: Information was available regarding 54 maternal deaths. According to HELLP syndrome classification 60.0% had class 1 disease, 35.6% had class 2 disease, and 4.4% had class 3 disease. Events associated with maternal deaths included cerebral hemorrhage (45%), cardiopulmonary arrest (40%), disseminated intravascular coagulopathy (39%), adult respiratory distress syndrome (28%), renal failure (28%), sepsis (23%), hepatic hemorrhage (20%), and hypoxic ischemic encephalopathy (16%). Delay in diagnosis of HELLP syndrome was implicated in 22 of 43 patients' deaths (51.1%). CONCLUSIONS: It appears that (1) most maternal deaths occurred among women with class 1 HELLP syndrome, (2) delay in diagnosis was associated with mortal consequences, and (3) hemorrhage in the hepatic or central nervous system or vascular insult to the cardiopulmonary or renal system were associated with increased mortality risk. PMID- 10521756 TI - Elevated maternal urine level of beta-core fragment of human chorionic gonadotropin versus serum triple test in the second-trimester detection of down syndrome. AB - OBJECTIVE: This study was undertaken to compare the Down syndrome screening efficiency of elevated maternal urine level of the beta-core fragment of human chorionic gonadotropin with that of the traditional serum triple test. STUDY DESIGN: Urinary beta-core fragment and serum analyte levels were measured prospectively in women with singleton pregnancies who were undergoing second trimester genetic amniocentesis. Urinary analyte levels were measured within a week of specimen collection. In some cases only alpha-fetoprotein was measured initially and human chorionic gonadotropin and unconjugated estriol levels were subsequently determined from the stored serum specimens. The Down syndrome screening efficiency of urinary concentration of beta-core fragment plus maternal age was compared with that of the traditional triple test. Receiver operating characteristic curves were generated for each algorithm and the areas under the curves were compared to determine which algorithm was superior. RESULTS: There were a total of 926 study patients, of whom 21 (2.3%) carried fetuses with Down syndrome. The mean (+/-SD) gestations at amniocentesis were 16.6 +/- 1.5 weeks for the fetuses without Down syndrome and 17.7 +/- 2.3 for the fetuses with Down syndrome. A total of 539 women (4 of whom carried fetuses with Down syndrome) had serum alpha-fetoprotein alone measured initially. Urinary concentration of beta core fragment had a 61.9% detection rate with a 4.9% false-positive rate for Down syndrome, whereas the values for the triple screen were 57. 1% and 11.2%, respectively. The areas under the receiver-operating characteristic curves were 0.8744 for elevated urinary beta-core fragment level and 0.7504 for the triple screen (P =.1116). When the false-positive rate was fixed at an ideal threshold value (3000 IU/L. CONCLUSIONS: Although systemic methotrexate administration is safe and effective for the treatment of tubal pregnancy, it does not necessarily reduce costs. Systemic methotrexate therapy could reduce costs if administered to patients with low initial serum human chorionic gonadotropin concentrations without confirmatory laparoscopy. PMID- 10521760 TI - The pattern of infertility diagnoses in women of advanced reproductive age. AB - OBJECTIVE: Our intention was to determine whether there is a unique pattern of infertility diagnoses in older infertile couples. STUDY DESIGN: The design of this study was a retrospective chart review study. It was performed in a tertiary referral reproductive medicine unit. There were 2 groups of patients-couples: group 1, female partner aged 20-29 (n = 105) at presentation; group 2, female partner aged 40-45 (n = 112) at presentation. All women underwent infertility evaluations between 1989 and 1994. There were no interventions. The prevalence of standard infertility diagnoses was the main outcome measure. RESULTS: The prevalence of 8 major infertility diagnoses in the younger and older groups (each couple could have >/=1 diagnosis) was as follows: (1) ovulatory factor-younger group, 56%; older group, 30%; (2) tubal factor-younger group, 34%; older group, 29%; (3) endometriosis-younger group, 13%; older group, 17%; (4) uterine factor younger group, 1%; older group, 5%; (5) cervical factor-younger group, 4%; older group, 1%; (6) luteal deficiency-younger group, 4%; older group, 10%; (7) male factor-younger group, 32%; older group, 45%; (8) unexplained-younger group, 5%; older group, 10%. The only significant difference was an increase in ovulatory factor in the younger group. CONCLUSIONS: There is no unique pattern of infertility diagnoses in women of advanced reproductive age as seen at a tertiary referral center. We speculate that a high false-positive rate associated with standard infertility tests and a different referral pattern for older couples obscures any real differences in the etiology of infertility in older couples. PMID- 10521761 TI - Hemostatic responses to maximal exercise in oral contraceptive users. AB - OBJECTIVE: This study examined the effect of exercise on markers of fibrinolysis and coagulation in users and nonusers of oral contraceptives. STUDY DESIGN: Fourteen oral contraceptive users and 14 nonusers performed a maximal exercise test on a cycle ergometer. Blood samples were collected before and immediately after the completion of the test. A repeated-measures analysis of variance was used for statistical analysis with values considered significant at P =.05. RESULTS: Acute maximal exercise resulted in significant increases in tissue plasminogen activator activity in both groups. There was a trend toward a smaller increase in tissue plasminogen activator activity in oral contraceptive users, but the difference between groups was not statistically significant. Plasminogen activator inhibitor 1 activity was reduced with exercise in both groups but with a significantly greater decrease observed in the nonusers (P <.0001). Prothrombin fragment 1+2 was significantly higher (P <.0001) in the oral contraceptive group but did not change with exercise. Epinephrine levels before and after exercise were similar between the 2 groups, but postexercise norepinephrine concentrations were significantly lower (P =.026) in the oral contraceptive users. CONCLUSION: These data suggest that oral contraceptive use blunts the fibrinolytic response to exercise. This, together with increased coagulation activation in oral contraceptive users, may alter the hemostatic balance during exercise. PMID- 10521762 TI - Vaginitis, cervicitis, and cervical length in pregnancy. AB - OBJECTIVE: We sought to determine the possible association among vaginitis, cervicitis, and cervical length in pregnancy. STUDY DESIGN: Primigravid volunteers, between 20 and 36 weeks' gestation (n = 210), were examined. Vaginitis was diagnosed by pH determination and wet mount smear, cervicitis was diagnosed by cervicography, and cervical length was diagnosed by vaginal ultrasonographic measurement. Patients with both vaginitis and cervicitis (n = 70) were compared with those without any trace of infection (n = 23). The remainder (n = 117) had variable degrees of infection and were excluded. RESULTS: The mean gestational age was 28.3 weeks. No significant association was found among vaginitis, cervicitis, and cervical length. In the infection group (n = 70), however, a significant association between an elevated vaginal pH (>5) and a shortened cervical length (r = 0.29) was noted. CONCLUSION: No significant association exists among vaginitis, cervicitis, and cervical length, but in patients with clinical signs of infection, an elevated pH appears to be associated with a decreased cervical length. PMID- 10521763 TI - Second-trimester maternal serum marker screening: maternal serum alpha fetoprotein, beta-human chorionic gonadotropin, estriol, and their various combinations as predictors of pregnancy outcome. AB - OBJECTIVE: We evaluated the value of all 3 common biochemical serum markers, maternal serum alpha-fetoprotein, beta-human chorionic gonadotropin, and unconjugated estriol, and combinations thereof as predictors of pregnancy outcome. STUDY DESIGN: A total of 60,040 patients underwent maternal serum screening. All patients had maternal serum alpha-fetoprotein measurements; beta human chorionic gonadotropin was measured in 45,565 patients, and 24,504 patients had determination of all 3 markers, including unconjugated estriol. The incidences of various pregnancy outcomes were evaluated according to the serum marker levels by using clinically applied cutoff points. RESULTS: In confirmation of previous observations, increased maternal serum alpha-fetoprotein levels (>2.5 multiples of the median) were found to be significantly associated with pregnancy induced hypertension, miscarriage, preterm delivery, intrauterine growth restriction, intrauterine fetal death, oligohydramnios, and abruptio placentae. Increased beta-human chorionic gonadotropin levels (>2.5 multiples of the median [MoM]) were significantly associated with pregnancy-induced hypertension, miscarriage, preterm delivery, and intrauterine fetal death. Finally, decreased unconjugated estriol levels (<0.5 MoM) were found to be significantly associated with pregnancy-induced hypertension, miscarriage, intrauterine growth restriction, and intrauterine fetal death. As with increased second-trimester maternal serum alpha-fetoprotein levels, increased serum beta-human chorionic gonadotropin and low unconjugated estriol levels are significantly associated with adverse pregnancy outcomes. These are most likely attributed to placental dysfunction. CONCLUSION: Multiple-marker screening can be used not only for the detection of fetal anomalies and aneu-ploidy but also for detection of high-risk pregnancies. PMID- 10521764 TI - Preeclampsia and genetic risk factors for thrombosis: a case-control study. AB - OBJECTIVE: Recently, it has been proposed that hereditary coagulation abnormalities leading to an increased venous thrombosis risk may play a role in the development of preeclampsia. We tested this hypothesis in women who have had preeclampsia compared with matched control subjects. STUDY DESIGN: We conducted a case-control study of 163 women with preeclampsia during 1991-1996. Control subjects were matched for age and delivery date. Patients and control subjects were tested for the presence of factor V Leiden, prothrombin 20210A allele, protein C, protein S, and antithrombin deficiency. Logistic regression methods were used for data analysis. RESULTS: The prevalence of these genetic risk factors was similar in the patient group (12.9%) and the control group (12.9%; odds ratio, 1.0; 95% confidence interval, 0.5-3.9). Unexpectedly, we found a high prevalence of factor V Leiden in the control group (9.2%). CONCLUSION: We found no differences in the prevalence of genetic risk factors of thrombosis in women with preeclampsia compared with control subjects. PMID- 10521765 TI - Effect of nitric oxide and carbon monoxide on uterine contractility during human and rat pregnancy. AB - OBJECTIVE: We sought to study the effects of authentic nitric oxide and carbon monoxide on the contractile activity of pregnant human and rat myometrium. STUDY DESIGN: Strips were prepared from uterine biopsy specimens of 10 pregnant, nonlaboring women at term gestation undergoing cesarean delivery. In addition, rings were prepared from the uteri of pregnant rats at midterm (day 14) and at term (day 22) gestation (n = 10-12). The tissues were mounted in organ chambers filled with Krebs-Henseleit solution continuously aerated with 5% carbon dioxide in air (37 degrees C, pH approximately 7.4) for isometric tension recording. The effects of nitric oxide and carbon monoxide gases on spontaneous contractile activity were studied. Responses to hemin (hemoxygenase substrate), which produces endogenous carbon monoxide, were also examined. Responses to nitric oxide and carbon monoxide were also studied in aortic and tail artery rings from pregnant rats after contraction with phenylephrine. RESULTS: Nitric oxide significantly inhibited contractility of human myometrium at term (area under the concentration-response curve, 145.36 +/- 30.02 vs 40.56 +/- 22.81 in controls; P <.05) and rat myometrium at midterm gestation (264.23 +/- 47.86 vs 121.82 +/- 23.50; P <.05) but not at term. No statistically significant inhibition was induced in human or rat myometrium by carbon monoxide, whereas hemin significantly attenuated contractility in human myometrium at term and in rat myometrium at midterm gestation (P <. 05). Nitric oxide, carbon monoxide, and hemin relaxed aortic and tail artery rings. CONCLUSIONS: Authentic nitric oxide inhibits rat uterine contractile activity at midterm gestation but not at term. However, nitric oxide inhibits human myometrium activity at term. Authentic carbon monoxide does not appear to modulate uterine contractility, whereas hemin may have some inhibitory properties. PMID- 10521766 TI - A role for the novel cytokine RANTES in pregnancy and parturition. AB - OBJECTIVE: RANTES (regulated on activation, normal T cell expressed and secreted), a potent and versatile chemokine, is capable of attracting monocytes, lymphocytes, basophils, and eosinophils. This cytokine has been implicated in the regulation of the inflammatory response and in the recruitment of macrophages to the implantation site in early pregnancy. RANTES messenger ribonucleic acid and protein have been detected in fetal tissue and first-trimester trophoblast in response to bacterial endotoxin. The purpose of this study was to determine whether intrauterine infection, parturition (preterm and term), and gestational age affect the amniotic fluid concentrations of RANTES in human pregnancy. STUDY DESIGN: A cross-sectional study was designed to examine the relationship between labor, microbial invasion of the amniotic cavity, gestational age, and RANTES expression in amniotic fluid. Amniotic fluid was obtained from 214 women in the following groups: (1) midtrimester (n = 22), (2) preterm labor with intact membranes in the presence (n = 20) or absence (n = 74) of microbial invasion of the amniotic cavity, (3) term, not in labor (n = 44) and term, in labor in the presence (n = 27) and absence (n = 27) of microbial invasion of the amniotic cavity. Microbial invasion of the amniotic cavity was defined as a positive amniotic fluid culture for microorganisms. RANTES concentrations were determined by use of a sensitive and specific immunoassay. RESULTS: (1) Amniotic fluid RANTES concentrations decrease with advancing gestational age (r = 0. 43; P <.01). (2) Labor at term was associated with an increase in median concentrations of RANTES (labor-median, 8.4 pg/mL; range, <1.3-94.4 vs no labor-median, <1.3 pg/mL; range, <1.3-230.3; P <.01). (3) Women with preterm labor who delivered preterm (no microbial invasion of the amniotic cavity) had a higher median concentration of amniotic fluid RANTES than those who delivered at term (median, 12.7 pg/mL; range, <1.3-928 vs median, <1.3 pg/mL; range, <1.3-127. 5; P <.001). (4) Microbial invasion of the amniotic cavity was associated with a significant increase in median amniotic fluid RANTES in both preterm and term labor (preterm labor with microbial invasion of the amniotic cavity-median, 51.6 pg/mL; range, <1.3-2290 vs preterm labor without microbial invasion of the amniotic cavity median, 12.7 pg/mL; range, <1.3-928 and vs preterm labor with delivery at term median, <1.3 pg/mL; range, <1.3-127.5; P <.001 for each; term labor with microbial invasion of the amniotic cavity-median, 16.8 pg/mL; range, <1.3-171.4 vs term labor without microbial invasion of the amniotic cavity-median, 8.4 pg/mL; range, <1.3-94.4; P <.05 and vs no labor and no microbial invasion of the amniotic cavity-median, 1.4 pg/mL; range, <1.3-230.3; P <.001 and P <.05, respectively). CONCLUSION: These results support a role for RANTES in the mechanisms of human parturition and in the regulation of the host response to intrauterine infection. PMID- 10521767 TI - Gelatin sponge plug to seal fetoscopy port sites: technique in ovine and primate models. AB - Amniotic fluid leakage and rupture of membranes are common complications of fetoscopy. We describe a plug technique for leakproof removal of endoscopic cannulas. Thirty gelatin sponge plugs were introduced in 5 pregnant ewes and 5 pregnant rhesus monkeys. In the primate model no evidence of amniorrhexis was noted on postoperative ultrasonography. Myometrium and membranes at the port sites were intact at term. A gelatin sponge plug technique may facilitate leakproof port removal after fetoscopy. PMID- 10521768 TI - Labor and delivery characteristics and risks of cranial ultrasonographic abnormalities among very-low-birth-weight infants. The Developmental Epidemiology Network Investigators. AB - OBJECTIVE: The objective of this study was to evaluate the relationships between 3 labor and delivery characteristics (duration of labor, interval between membrane rupture and delivery, and route of delivery) and 4 neonatal cranial ultrasonographic abnormalities (intraventricular hemorrhage, ventriculomegaly, echodensity of cerebral white matter, and periventricular leukomalacia). STUDY DESIGN: We prospectively gathered data on 1588 very low birth weight infants including neonatal cranial ultrasonographic studies, maternal interview, and maternal and infant chart reviews. We performed univariate and multivariate analyses. RESULTS: In univariate analysis vaginal delivery was associated with an increased risk of all 4 cranial ultrasonographic abnormalities. In multivariate analysis, however, vaginal delivery was no longer associated with periventricular leukomalacia. Moreover, the risks of intraventricular hemorrhage, ventriculomegaly, and echodensity attributable to vaginal delivery were no longer elevated when the sample was limited to infants born within 1 hour after membrane rupture and adjustment was made for fetal vasculitis and for other potential confounders. CONCLUSION: Vaginal delivery was the only obstetric characteristic consistently associated with intracranial hemorrhage and white matter disease in these preterm infants. Because its relationship to brain lesions was markedly reduced when placental inflammation was accounted for, however, vaginal delivery may simply have acted as a marker for antecedent inflammation or infection and not as a direct contributor to brain disorders. PMID- 10521769 TI - Antenatal corticosteroids and cranial ultrasonographic abnormalities. AB - OBJECTIVE: This study was undertaken to determine whether very-low-birth-weight infants whose mothers received a course of antenatal corticosteroids were at decreased risk for 3 cranial ultrasonographic entities that predict neurodevelopmental dysfunction. STUDY DESIGN: This retrospective cohort study evaluated 1604 infants weighing 500 to 1500 g who underwent >/=1 of 3 cranial ultrasonographic scans required by design at specified postnatal intervals and whose own and mother's hospital charts were reviewed. Infants were classified according to mother's course of antenatal corticosteroids (none, partial, or complete). RESULTS: In the total sample the risks of intraventricular hemorrhage and of an echolucent image in the cerebral white matter were only modestly (and not statistically significantly) reduced after a full course of antenatal corticosteroids, whereas antenatal corticosteroids appeared to significantly reduce the risk of ventriculomegaly after even a partial course. Antenatal corticosteroids appeared to halve the risk of ventriculomegaly and echolucent image among the gestationally youngest infants and those with intraventricular hemorrhage, hypothyroxinemia, or vasculitis of the umbilical cord or chorionic plate of the placenta. CONCLUSION: These observations are consistent with the hypothesis that antenatal corticosteroids protect very-low-birth-weight infants, especially those who are most vulnerable, against the risk of cranial ultrasonographic abnormalities. PMID- 10521770 TI - Acute cerebral redistribution in response to invasive procedures in the human fetus. AB - OBJECTIVES: We sought to investigate the fetal hemodynamic response to the acute stress of invasive procedures. STUDY DESIGN: The middle cerebral artery pulsatility index was measured by Doppler ultrasonography before and after 136 invasive procedures (fetal blood sampling, transfusion, shunt insertion, tissue biopsy, and ovarian cyst aspiration). The response of fetuses submitted to invasive procedures involving transgression of the fetal body, such as intrahepatic vein blood sampling, was compared with that of control procedures at the placental cord insertion. RESULTS: The middle cerebral artery pulsatility index value fell with fetal blood sampling performed at the intrahepatic vein (median, -0.26; 95% confidence interval, -0.35 to -0.15) but not at the placental cord insertion (median, 0.05; 95% confidence interval, -0.04 to 0.19). With transfusions, the middle cerebral artery pulsatility index also fell with procedures at the intrahepatic vein (mean, -0.51; 95% confidence interval, -0.66 to -0.35) but not at the placental cord insertion (mean, -0.04; 95% confidence interval, -0.23 to 0.14). The magnitude of the response was greater with transfusions than with blood sampling alone. The middle cerebral artery pulsatility index value also fell with non-fetal blood sampling procedures involving transgression of the fetal body (mean, -0.32; 95% confidence interval, 0.56 to -0.09) but not with control non-fetal blood sampling procedures. The change in the middle cerebral artery pulsatility index was not related to gestational age, with the youngest fetus showing a fall in the middle cerebral artery pulsatility index value being at 16 weeks' gestation. Although the degree of response was weakly correlated with the duration of needling (y = -0.21 - 0.00014x; R (2) = 0.08; P =.02), multiple logistic regression demonstrated that this was instead a function of the type of the procedure. A response was seen within 70 seconds of fetal puncture. The fetal heart rate did not change significantly with procedures in any of the above-mentioned groups. CONCLUSIONS: The human fetus mounts a cerebral hemodynamic response to invasive procedures involving transgression of the fetal body, which is consistent with the brain sparing effect. PMID- 10521771 TI - Cigarette smoking during pregnancy and risk of preeclampsia: a systematic review. AB - In this systematic review of the existing evidence regarding the relationship between cigarette smoking during pregnancy and preeclampsia, studies were found through searches of MEDLINE (1966-October 31, 1998), Embase, Popline, CINAHL, Lilacs, bibliographies of identified studies, and proceedings of meetings on preeclampsia, and also through contact with relevant researchers. No language restrictions were imposed. Only cohort and case-control studies dealing with the relationship between cigarette smoking and preeclampsia were considered. Assessment of methodologic quality and data extraction of each study were carried out by 2 authors working independently. Typical relative risks and odds ratios with 95% confidence intervals were calculated for cohort and case-control studies, respectively, with both fixed and random effects models. Twenty-eight cohort studies and 7 case-control studies including a total of 833,714 women were included. All cohort studies reported an inverse association between cigarette smoking during pregnancy and incidence of preeclampsia (typical relative risk, 0.68; 95% confidence interval, 0.67-0.69). The findings were similar for case control studies (typical odds ratio, 0.68; 95% confidence interval, 0.57-0.81). An inverse dose-response relationship was also found. Pooled data from cohort and case-control studies showed a lower risk of preeclampsia associated with cigarette smoking during pregnancy. PMID- 10521772 TI - Selective estrogen receptor modulators. PMID- 10521775 TI - A flawed experiment, again. PMID- 10521776 TI - Unusual cases of severe thrombotic episodes during the peripartum period. PMID- 10521780 TI - Co-localization of Trk neurotrophin receptors and regulatory peptides in the endocrine cells of the teleostean stomach. AB - Recently it has been observed that a subpopulation of gut endocrine cells in vertebrates express Trk-like proteins, suggesting that neurotrophins could regulate the synthesis and storage of amines and peptides of these cells. Nevertheless, the peptides and amines present in the endocrine cells that express Trks have not been characterized. In this study we used immunohistochemistry to investigate the occurrence of Trk-like proteins (TrkA-like, TrkB-like and TrkC like) and the possible co-localization of these with peptides and/or biogenic amines in the endocrine cells of the stomach of three teleost (bass, gilt-head and scorpionfish). No TrkA-like immunoreactivity (IR) was detected in the stomach of these species, whereas TrkB-like IR and TrkC-like IR were observed in numerous cells of the gastric epithelium. TrkB-like immunoreactive cells were present in all three species examined, and were particularly abundant in the blind sac. Conversely, TrkC-like immunoreactive cells were found only in the bass stomach, apparently co-localized with TrkB-like IR. TrkB-like IR was found co-localized with somatostatin IR in scorpionfish, and with somatostatin and CGRP IR in gilt head and bass. Gastric endocrine cells expressing 5-HT, glucagon, insulin, met-, leu-enkephalin, substance P, PYY, VIP, CCK, NPY, bombesin and motilin were unreactive for Trk-like proteins. The present results provide direct evidence for the occurrence of Trk-like neurotrophin receptor proteins in a subpopulation of the teleostean gastric endocrine cells and suggest that neurotrophins could regulate, as in neurons, the expression of some neuropeptides such as somatostatin and CGRP. PMID- 10521781 TI - Inside-out vs. standard vein graft to repair a sensory nerve in rats. AB - Nerve regeneration in a sensory nerve was obtained by the application of different techniques: inside-out vein graft (IOVG group) and standard vein graft (SVG group). These techniques provide a good microenvironment for axon regeneration in motor nerves, but their efficiency for regeneration of sensory nerves is controversial. The saphenous nervtce was sectioned and repaired by the inside-out and standard vein graft techniques in rats. After 4, 12, and 20 weeks the graft and the distal stump were observed under electron microscopy. In each studied period, the pattern, diameters, and thickness of the myelin sheaths of the regenerated axons were measured in the graft and distal stump. A comparative study about the regenerated nerve fibers by these two different techniques was performed. Regenerated nerve fibers were prominent in both vein grafts 4 weeks after the surgical procedures. On the other hand, in the distal stump, regenerated nerve fibers were observed only from 12 weeks. In both inside-out vein graft and standard vein graft statistical difference was not observed about the diameters and thickness of the myelinated fibers after 20 weeks. On the other hand, the inside-out group had greater regenerated axon number when compared to the standard group. There is a capillary invasion in both graft and distal stump, especially in the IOVG group. The regenerated axons follow these capillaries all the time like satellite microfascicles. After 20 weeks, the diameters of regenerated fibers repaired by the standard vein graft technique were closer to the normal fibers compared to the inside-out vein graft. On the other hand, the pattern of these regenerated axons was better in the IOVG group. PMID- 10521782 TI - Post-traumatic migration and emergence of a novel cell line upon the ependymal surface of the third cerebral ventricle in the adult mammalian brain. AB - This investigation describes the migration and emergence of significant numbers of what appear to be neuron-like cells upon the surface of the median eminence of the adult rodent neurohypophyseal system of the endocrine hypothalamus following the trauma of hypophysectomy. These cells appear to migrate through the neuropil of the underlying median eminence and emerge in large numbers upon the surface of the third cerebral ventricle within 7 days following hypophysectomy (axotomy) of supraoptic (SON) and paraventricular neurites (PVN) of the adult neurohypophyseal system. Previous investigations have demonstrated regeneration of the neural stem and neural lobe in a variety of mammalian species (Adams et al., J Comp Neurol, 1969;135:121-144; Beck et al., Neuroendocrinology, 1969;5:161-182; Scott et al., Exp Neurol, 1995;131-1:23-39; Scott and Hansen, Vir Med 1997;124:249-261). It also has been demonstrated that the process of regeneration is invariably accompanied by the up-regulation of nitric oxide synthase (NOS), the enzyme that catalyzes arginine to nitric oxide (NO) and that both neurohypophyseal regeneration, as well as migration and emergence of neuron-like cells upon the surface of the adjacent third cerebral ventricle, is associated with the up regulation of NOS and increased expression of NO. It also has been amply demonstrated that this entire process of neurohypophyseal regeneration and cell migration is completely inhibited by the introduction of the antagonist of nitric oxide, namely, nitroarginine (Scott et al., Exp Neurol, 1995;131-1:23-39; Scott and Hansen, Vir Med, 1997;124:249-261). The emergence and migratory dynamics of this novel cell line upon the floor of the rodent third cerebral ventricle are discussed with respect to the role of the ubiquitous free radical NO and the implications and potential clinical applications of neuronal migration following trauma in the human central nervous system (CNS). PMID- 10521783 TI - An autoradiographic assessment of epithelial cell proliferation and post-natal maturation of the tracheal epithelium in infant ferrets. AB - The tracheal epithelium of infant ferrets undergoes rapid postnatal maturation over the first month of life to achieve the pseudostratified columnar configuration characteristic of the large airways of other mammals. We have used in vivo pulsing with tritiated thymidine ((3)HT) to elicit autoradiographic labeling of cells synthesizing nucleic acids in order to characterize more fully the contribution to development of different cell types comprising the nascent epithelial layer during this period of rapid growth. These studies indicate that two distinct populations of epithelial cells possess proliferative potential and contribute to the establishment of the mature adult epithelial layer. These investigations further confirm the mitotic potential of basal cells during a period of rapid postnatal growth and development of the tracheal epithelial layer. These studies also document the contribution to early airway development by non-ciliated cells, which predominate on the luminal border of the ferret trachea at birth. The temporal and histologic patterns of airway epithelial maturation during post-natal life in the ferret as contained in this study exhibit similarities to those which occur with recovery from injury by infection and irritant exposure in mature airways. Thus, the characterization of epithelial cell compartments having proliferative potential may provide insights into the mechanisms whereby normal airway epithelial organization is established and maintained during development as well as the possible recapitulation of these mechanisms during times of epithelial regeneration following injury. PMID- 10521785 TI - Urokinase regulates embryonic cardiac cushion cell migration without converting plasminogen. AB - Urokinase-type plasminogen activator (uPA) activation of plasminogen is an important mediator of cell migration in many cell types. In the developing avian heart, uPA has been implicated as a mediator of atrioventricular (AV) cushion cell migration; however, the role of the plasminogen/plasmin system has not been examined. The purpose of this study was to test the hypothesis that uPA conversion of plasminogen to plasmin mediates AV cushion cell migration in vitro. Stage 17/18 chicken atrioventricular tissue lysates converted plasminogen into plasmin through uPA activity but no tissue-type plasminogen activator activity was detected. Zymograms on living cultured AV explants also activated plasminogen producing plasmin that degraded extracellular protein. The migratory capacity of cushion cells was assessed in the presence or absence of various test reagents known to alter the plasminogen/plasmin system. Addition of either human or chicken plasminogen or aprotinin (an inhibitor of plasmin) had no effect on cell migration. However, an anti-catalytic uPA antibody that blocked AV uPA activity, significantly decreased cell migration at all concentrations tested. These results showed that uPA mediated a portion of cushion cell migration in vitro. Although AV segments activated plasminogen and degraded extracellular proteins, uPA's functional role in cushion cell migration did not involve the plasminogen/plasmin system. PMID- 10521784 TI - Submandibular gland morphogenesis: stage-specific expression of TGF-alpha/EGF, IGF, TGF-beta, TNF, and IL-6 signal transduction in normal embryonic mice and the phenotypic effects of TGF-beta2, TGF-beta3, and EGF-r null mutations. AB - Branching morphogenesis of the mouse submandibular gland (SMG) is dependent on cell-cell conversations between and within epithelium and mesenchyme. Such conversations are typically mediated in other branching organs (lung, mammary glands, etc.) by hormones, growth factors, cytokines, and the like in such a way as to translate endocrine, autocrine, and paracrine signals into specific gene responses regulating cell division, apoptosis, and histodifferentiation. We report here the protein expression in embryonic SMGs of four signal transduction pathways: TGF-alpha/EGF/EGF-R; IGF-II/IGF-IR/IGF-IIR; TGF-betas and cognate receptors; TNF, IL-6, and cognate receptors. Their in vivo spatiotemporal expression is correlated with specific stages of progressive SMG development and particular patterns of cell proliferation, apoptosis, and mucin expression. Functional necessity regarding several of these pathways was assessed in mice with relevant null mutations (TGF-beta2, TGF-beta(3), EGF-R). Among many observations, the following seem of particular importance: (1) TGF-alpha and EGF R, but not EGF, are found in the Initial and Pseudoglandular Stages of SMG development; (2) ductal and presumptive acini lumena formation was associated with apoptosis and TNF/TNF-R1 signalling; (3) TGF-beta2 and TGF-beta3 null mice have normal SMG phenotypes, suggesting the presence of other pathways of mitostasis; (4) EGF-R null mice displayed an abnormal SMG phenotype consisting of decreased branching. These and other findings provide insight into the design of future functional studies. PMID- 10521787 TI - Estrogen-induced microvilli and microvillar channels and entrapment of surfactant lipids by alveolar type I cells of bovine lung. AB - The ATI cells are simple, flat squamous epithelial cells, which are evolved to function as a component of the alveolar-capillary membrane, ideally designed for gaseous exchange. They inherently lack an active metabolic machinery and lead a precarious existence in the face of hostile environment. On the other hand, the ATI cells of the lung of ruminating animals are endowed with structure-functional properties which enable them to exert a selective barrier function against a wide range of osmotic pressure gradients at their luminal surface. Such gradients are created by a complex gaseous homeostasis due to expectoration of several gases and volatile fatty acids originating from the complex stomach of the ruminants. The purpose of this study is to examine the effect of estradiol propionate on the ultrastructure of the ATI cells and their interaction with the surfactant lipids. The lungs of estrogen and dexamethasone treated male calves were harvested for electromicroscopic examination. The evidence is presented that estradiol induced the formation of microvilli and microvillar channels at the luminal surface. At these regional modifications, intense interactions with the surfactant lipids and their entrapment into the pathways of endocytosis, took place in the squamous part of the ATI cells. Concurrently, large basal protrusions ended up as long lamellipods deep into the alveolar interstitium. The filamentous cytoskeletal network and microtubules intermixed with the translocated organelles such as Golgi apparatus and associated coated and uncoated vesicles. The results of this study support the hypothesis that estrogen regulate the selective barrier function of the ATI cells. The entrapment of surfactant lipids under the influence of estrogen by ATI cells is a significant change perhaps in response to extracellular stimuli and expression of transmembrane receptors. It implies that these epithelial cells are specially evolved to adapt to a complex gaseous homeostasis in the lung of the ruminating ungulates. PMID- 10521786 TI - Implantation in the marmoset monkey: expansion of the early implantation site. AB - This study was initiated to examine the early stages of trophoblast adhesion and invasion during implantation in the marmoset. Seven implantation sites were found in the uteri of four marmosets taken between days 13 and 15 of gestation. Three implantation sites in two uteri were examined in detail by electron microscopy. Between days 13 and 15, the marmoset implantation site expanded peripherally by adding areas where syncytial trophoblast penetrated between uterine luminal epithelial cells. Such penetrating masses often bridged openings of endometrial glands, shared junctional complexes with the uterine epithelial cells between which they are infiltrating, and subsequently reached the residual basal lamina of the uterine luminal epithelium. Centripetal to the peripheral region was an intermediate region in which syncytial trophoblast overlay individual clusters of epithelial cells and rested along the basal lamina. In this region there was some evidence of fusion of syncytial trophoblast with uterine epithelial cells. In the central region of the implantation site near the inner cell mass and amnion the trophoblast formed elaborate lamellipodia in relation to the basal lamina. In one of the three specimens examined with electron microscopy there were two foci where trophoblast penetrated through the basal lamina. It was also in the central region that trophoblast penetrated farthest into the uterine glands. The gland cells closest to trophoblast were less closely associated and lost their columnar shape, forming large round cells similar to the epithelial plaque cells of other primates. Where two blastocysts implanted on the same side of the uterus a conjoint membrane was formed which in regions consisted solely of syncytial trophoblast with two basal surfaces and two basal laminas. The prolonged period of time when the implantation site expands within the plane of the uterine epithelium (trophoblastic plate stage) and the peripheral to central sequence in extent of development make this primate a particularly useful animal for studies of trophoblast adhesion to and penetration of the uterine luminal epithelium. PMID- 10521788 TI - Structural abnormalities underlying alveolar hypoventilation and fluid imbalance in the dystrophic hamster lung. AB - Bio 14.6 dystrophic hamsters exhibit alveolar hypoventilation and increased lung hydration. This study evaluated whether age- and genotype-related morphometric differences in lungs exist and correlate with the development of lung pathophysiology. Morphometry was used to characterize lungs of young (Y) and mature (M) control (C) and dystrophic (D) hamsters. With age, both C and D had increased barrier surface area [S(a-b,p)] and morphometric diffusing capacity index [mdci], and decreased harmonic thickness. In C but not D, mean capillary diameter [d(c)] and parenchymal volume density [V(v)(p,L)] increased with age, whereas barrier arithmetic thickness decreased. Chord length increased with age, whereas the ratio of parenchymal surface area to airspace volume [S/V] and the intersection density of the air-blood interface [I(v)(a-b,s)] decreased in D but not C. At both ages, lung volume relative to body mass was greater in D than C. With that exception, no genotype differences were found in young hamsters. Mature D displayed lower V(v)(p,L), S/V, d(c), I(v)(a-b,s), S(a-b,p), and mdci than mature C. Independent of age, chord length was greater but arithmetic thickness, airspace surface density, frequency of type II cells, and lamellar body area and volume density were lower in D than C. We conclude: 1) lung volume relative to body growth was greater in dystrophics than controls; 2) parenchymal remodeling was delayed or abnormal in dystrophics; 3) lower diffusing capacity in mature dystrophics may effect alveolar hypoventilation; 4) lower tissue volume, surface area, and the type II cell abnormalities in dystrophics could reduce sodium and water transport leading to greater lung hydration. PMID- 10521789 TI - Comparative tyrosine-kinase profiles in colorectal cancers: enhanced arg expression in carcinoma as compared with adenoma and normal mucosa. AB - There is strong evidence that tyrosine kinases are involved in the regulation of cellular growth and tumor progression. Over-expressions of tyrosine kinases have been documented in a number of neoplasms. To study the roles of tyrosine kinases in colon cancer, we developed a tyrosine-kinase-expression profile for each of the four different stages of colon carcinogenesis, using normal colon mucosa, adenomatous polyps, primary carcinoma and hepatic metastases collected from the same patient. We identified 30 tyrosine kinases expressed in these tissues: they include 10 non-receptor tyrosine kinases (yes, fyn, lyn, brk, abl, arg, jak1, jak3, tyk2 and itk), 17 receptor tyrosine kinases (erbB2, PDGF-Ralpha, PDGF Rbeta, kit, c-fms, met, ron, FGF-R1, FGF-R2, FGF-R3, FGF-R4, cek5, tie-1, tkt, axl, sky and Ins-R), 2 dual kinases (mek and sek) and one possible novel kinase. Among these kinases, arg kinase appears to be expressed at a higher level in primary carcinoma and metastatic tumor than in adjacent normal mucosa or adenomatous polyp. This result was confirmed by extensive analysis of 50 additional matched sets of normal colon and colon-tumor specimens, using arg specific primers and RT-PCR reactions. This study identifies a possible role for arg tyrosine kinase in colon carcinogenesis, especially in the transition from adenoma to carcinoma. PMID- 10521790 TI - Fatty-acid composition in serum phospholipids and risk of breast cancer: an incident case-control study in Sweden. AB - The study of the relationship between dietary intake of fatty acids and the risk of breast cancer has not yielded definite conclusions with respect to causality, possibly because of methodological issues inherent to nutritional epidemiology. To evaluate the hypothesis of possible protection of n-3 polyunsaturated fatty acids (PUFA) against breast cancer in women, we examined the fatty-acid composition of phospholipids in pre-diagnostic sera of 196 women who developed breast cancer, and of 388 controls matched for age at recruitment and duration of follow-up, in a prospective cohort study in Umea, northern Sweden. Individual fatty acids were measured as a percentage of total fatty acids, using capillary gas chromatography. Conditional logistic-regression models showed no significant association between n-3 PUFA and breast-cancer risk. In contrast, women in the highest quartile of stearic acid had a relative risk of 0.49 (95% confidence interval, 0.22-1.08) compared with women in the lowest quartile (trend p = 0.047), suggesting a protective role of stearic acid in breast-cancer risk. Besides stearic acid, women in the highest quartile of the 18:0/18:1 n-9c ratio had a relative risk of 0.50 (95% confidence interval, 0.23-1.10) compared with women in the lowest quartile (trend p = 0.064), suggesting a decrease in breast cancer risk in women with low activity of the enzyme delta 9-desaturase (stearoyl CoA desaturase), which may reflect an underlying metabolic profile characterized by insulin resistance and chronic hyper-insulinemia. PMID- 10521791 TI - Mechanisms associated with abnormal E-cadherin immunoreactivity in human bladder tumors. AB - The involvement of E-cadherin in the progression of carcinoma is supported by a large number of studies showing an inverse relationship between E-cadherin immunoreactivity and tumor aggressiveness. However, the mechanisms leading to decreased E-cadherin immunoreactivity are still unclear. Comparison of Northern blotting and immunohistochemistry in a series of 49 frozen bladder tumors revealed that, in 16 of 23 tumors with abnormal staining, clear mRNA down regulation occurred. In the 7 cases without mRNA down-regulation, no structural anomalies of E-cadherin could be detected by Southern blotting, Western blotting or PCR-SSCP. Western blotting confirmed that, in 6 of these tumors, E-cadherin was down-regulated at the protein level. This down-regulation was accompanied by down-regulation of alpha-catenin and, to a lesser extent, of beta- or gamma catenin. However, Northern-blot analysis indicated that expression of the 3 catenins is maintained at the mRNA level. Thus our data show that, in bladder tumors, mRNA down-regulation accounts for about two thirds (16/23) of tumors with abnormal staining and that post-transcriptional down-regulation of E-cadherin occurs in 6/23 of these tumors. PMID- 10521792 TI - Organochlorines and breast cancer: a case-control study in Brazil. AB - Breast cancer is an important cause of morbidity and mortality in Brazil. Some studies have analyzed the potential role of organochlorine compounds in breast cancer etiology. These chemical compounds have been widely used in agriculture and in vector-control programs in Brazil. A case-control study was carried out in the main cancer hospital of the Instituto Nacional de Cancer in Rio de Janeiro to investigate exposure to organochlorinated pesticides as a risk factor for breast cancer. We investigated 177 cases of invasive breast cancer at the hospital, between May 1995 and July 1996, and 350 controls selected among female visitors at the same hospital. In addition to information obtained from interviews, blood samples were taken, to analyze residual amounts of organochlorine by gas chromatography using an electron capture detector. [1,1-Dichloro-2, 2-bis(p chlorophenyl)ethylene] (DDE) was determined in sera of 457 women from a total of 493 participants who had serum samples available. Residues of hexachlorobenzene were found in 11 women only. No statistically significant association was found between breast cancer risk and serum level of DDE or history of exposure to pesticides. Serum levels of DDE (ng/ml) were similar in patients (median = 3.1, mean = 5.1) and controls (median = 3.1, mean = 4.8) (p = 0.93). The age-adjusted odds ratio of breast cancer for women in the upper quintile compared with those in the lowest quintile was 0.90 (95% confidence interval 0.47-1.73). When adjusted for other variables, the risk remained not statistically significant (upper quintile odds ratio = 0.79, 95% confidence interval 0.39-1.60). In our hands, exposure to organochlorinated pesticides measured by history or serum analysis was thus not a risk factor for breast cancer. PMID- 10521793 TI - Nutrient intake and gastric cancer in Mexico. AB - In contrast to the decreasing trends observed in most countries, gastric-cancer mortality has remained at about the same level in Mexico throughout the last 40 years. As part of a study carried out in the metropolitan area of Mexico City, an assessment of nutrient intake and gastric cancer is presented here. The study population comprised 220 cases of gastric cancer and 752 population-based controls. Our results showed 70 to 80% reduction in the risk of developing this tumor, associated with the intake of polyunsaturated fat, fiber and vitamin E; and this effect was independent of the histological type of the tumor (i.e., intestinal or diffuse). On the other hand, an increased risk of gastric cancer was related to the consumption of saturated fat (OR(Q4vs.Q1) = 4.37, 95% CI 1.89 10.12) and, cholesterol (OR(Q4vs.Q1) = 2.39, 95% CI 1.23-4.64), but such effects were restricted to the intestinal type of gastric cancer. In the whole study population, mono-unsaturated fat intake increased the risk for gastric cancer, and a marginally significant increasing trend was observed for protein consumption. The findings from this study add information about the role of specific nutrients in the etiology of gastric cancer. PMID- 10521794 TI - Genetic polymorphisms of tobacco- and alcohol-related metabolizing enzymes and oral cavity cancer. AB - Both genetic and environmental factors are involved in the development of cancer. Oral cavity cancer has been reported to be epidemiologically associated with tobacco and alcohol consumption. We examined genetic polymorphisms of the glutathione-S-transferase (GST) M1/T1, cytochrome P-450 (CYP) 1A1/2E1 and aldehyde dehydrogenase 2 (ALDH2) genes in 92 Japanese patients with oral cavity cancer and 147 unrelated non-cancer Japanese controls. There was a significant association between cigarette smoking and cancer risk but no significant association between alcohol consumption and cancer risk. The frequency of the GSTM1 null genotype was significantly higher in cancers (58.7%) compared with controls (46. 3%). However, there were no significant differences between controls and patients with oral cavity cancer in the polymorphisms of the GSTT1, CYP1A1, CYP2E1 and ALDH2 genes. From statistical evaluation on various combinations of genotypes, we did not observe any gene combinations associated with cancer risk. There were also no genetic polymorphisms associated with increased risk of oral cavity cancer among smokers and drinkers. These results imply that the GSTM1 null genotype has a weak correlation, but another 4 genetic polymorphisms are unlikely to be associated, with oral cavity cancer among Japanese. PMID- 10521795 TI - High frequency of deletion on chromosome 9p21 may harbor several tumor-suppressor genes in human prostate cancer. AB - Chromosome 9p has been reported to be a critical region of loss in various cancers. Our present study was designed to determine the frequency of deletions at different loci of chromosome 9p in microdissected samples of normal prostatic epithelium and carcinoma from the same patients. For this purpose, DNA was extracted from the microdissected sections of normal and tumor cells of 40 prostate specimens, amplified by PCR and analyzed for loss of heterozygosity (LOH) on chromosome 9p using 15 microsatellite markers. Only 6 of 15 microsatellite markers exhibited LOH in prostate cancer specimens (D9S162, D9S1748, D9S171, D9S270, D9S273 and D9S153). LOH on chromosome 9p was identified in 29 of 40 cases (72.5%) with at least 1 marker. The main deletion was found on 9p21, at loci D9S1748 (50%), D9S171 (51.4%) and D9S270 (21.8%). There was also a deletion on 9p22 at locus D9S162 (8.3%), on 9p13 at locus D9S273 (13.8%) and on 9p11 at locus D9S153 (7.7%). LOH data were correlated with stage of prostate cancer and revealed a high frequency of LOH at 3 or more loci in samples with stage T(3)N(0)M(0) (46%) compared with stage T(2)N(0)M(0) (15%), which suggests a higher incidence of LOH in the advanced stage of prostate cancer. One of the candidate target tumor-suppressor genes, p16 (MTS-1/CDKN2), has been identified within the 9p21 deleted region in tumor cell lines. Expression of P16 protein was either absent or very low in prostate cancer samples, suggesting that loss of the p16 gene may be involved in prostatic carcinogenesis. PMID- 10521796 TI - Evolution of precancerous lesions in a rural Chinese population at high risk of gastric cancer. AB - The pathogenesis of gastric cancer (GC), particularly of the intestinal type, is thought to involve a multistep and multifactorial process. Our objective was to determine the rates of transition from early to advanced gastric lesions in a population in Linqu County, China, where the GC rates are among the highest in the world. An endoscopic screening survey was launched in 1989-1990 among 3,399 residents aged 34-64 years with precancerous lesions diagnosed from biopsies taken from 7 standard locations in the stomach and from any suspicious sites. The cohort was subsequently followed, with endoscopic and histopathologic examinations conducted in 1994. Logistic regression analysis was used to estimate odds ratios (ORs) of progression to advanced lesions of various levels of severity as a function of age, sex and baseline pathology. The rates of progression were higher among older subjects, among men and among subjects with more extensive gastric lesions. 34 incident GCs were identified during the follow up period. The ORs of GC, adjusted for age and sex, varied from 17.1, for those with baseline diagnoses of superficial intestinal metaplasia (IM), to 29.3, for those with deep IM or mild dysplasia (DYS) or IM with glandular atrophy and neck hyperplasia, to 104.2, for those with moderate or severe DYS, as compared with subjects with superficial gastritis (SG) or chronic atrophic gastritis (CAG) at baseline. Our prospective study of a high-risk population revealed sharp increases in the risk of GC and advanced precursor lesions according to the severity of lesions diagnosed at the start of follow-up. Int. J. Cancer, 83:615 619, 1999. Published 1999 Wiley-Liss, Inc. PMID- 10521797 TI - Ecological study on the risks of esophageal cancer in Ci-Xian, China: the importance of nutritional status and the use of well water. AB - Our purpose was to determine the environmental risks in the development of esophageal cancer in Ci-Xian, which has one of the highest incidences of esophageal cancer in China. The subjects included 404, 352 and 400 inhabitants living in high-, medium-, and low-incidence areas of esophageal cancer, as well as 301 esophageal cancer patients. A food intake-frequency survey using a 7-day weighted inventory questionnaire was conducted on these individuals. Questions on occupation, working conditions, income per year, family disease history, medical complaints, and demographic features were also included in the questionnaire. The levels of nitrogen compounds in selected samples of well water were also measured in each of the 3 areas. Clear-cut differences in food intake were seen among inhabitants living in the 3 different areas, suggesting that regional differences in nutritional styles do exist. In both males and females, the intake of potatoes, fruit, vegetables, and meat were significantly lower in inhabitants living in the high-incidence area than in the other inhabitants, much the same as that of cancer patients. A low intake of carotene, and vitamins A and C was also seen in populations living in the high-incidence area of esophageal cancer. The well water polluted with nitrogen compounds was significantly related to the high incidence of esophageal cancer. In contrast, tobacco, alcohol consumption, and the intake of pickled vegetables and moldy foods did not relate to the different incidence rates. Our results suggest that low intake of fruit, vegetables, potatoes and meat, and the quality of well water may be important factors in the development of esophageal cancer in Ci-Xian. PMID- 10521798 TI - Mitochondrial gene mutation, but not large-scale deletion, is a feature of colorectal carcinomas with mitochondrial microsatellite instability. AB - We have shown that microsatellite instability (MSI) occurs in mitochondrial DNA (mtDNA) of colorectal carcinomas. To determine whether such mitochondrial microsatellite instability (mtMSI) is associated with certain forms of mitochondrial gene alterations, we extended the screening in the same series of 45 carcinomas. Analysis by whole mtDNA amplification (16.5 kb) and digestion revealed no detectable large-scale change in these carcinomas. In contrast, single-strand conformation polymorphism (SSCP) analysis demonstrated NADH dehydrogense (ND) gene alterations in 7 carcinomas (16%), including 3 mononucleotide repeat alterations, 2 missense mutations and 1 small (15 bp) deletion. Six of these 7 carcinomas also exhibited mtMSI of the (C)n sequence in the displacement-loop (D-loop) region. Thus, frameshift or missense mutations rather than large-scale changes in the mtDNA were more common features in colorectal carcinomas with mtMSI. By analogy to mutational features of nuclear MSI, mtMSI most likely results from certain repair deficiencies in the mtDNA and probably plays a role in the tumor development of certain colorectal carcinomas. PMID- 10521799 TI - Incidence of testicular germ-cell malignancies in England and Wales: trends in children compared with adults. AB - The incidence of testicular cancer has been increasing markedly in most industrialised countries. This rise is known to have affected young adults, but it is less clear whether it has affected other age groups, particularly children. We used data from the National Cancer Registry file at the Office of National Statistics (ONS) and the National Registry of Childhood Tumours to examine trends in testicular germ-cell malignancies overall in England and Wales from 1962 to 1990 and in children from 1962 to 1995. The incidence of testicular cancer at all ages rose by 3.4% (95% CI 3.3-3.6%) per annum from 1962 to 1990. A similar rise in the incidence of germ-cell malignancies occurred during the years for which histological information was available in the ONS files, 1971-1989 (3.4%; 3.1 3.6%), to which both seminomas and non-seminomas contributed equally. The incidence of non-seminomas in adults rose in men under age 55 years and declined in older men, whereas there were increases in the incidence of seminomas in both young and older men. Cohort analysis at young ages showed a marked rise in the risk of germ-cell malignancies up to the cohort born in 1955-1959 but no further rise for those born subsequently. The rise in the incidence of these tumours in young adults was paralleled by a similar trend, although less marked, in children aged under 15 years (1.3% per annum; 0.2-2.5%). The increase in risk for children in this very large data set alongside the rise in young adults is compatible with the hypothesis that childhood and adult testicular germ-cell malignancies may have some common risk factors, presumably pre-natal. PMID- 10521800 TI - Exposure to environmental tobacco smoke and risk of adenocarcinoma of the lung. AB - We conducted a case-control study of adenocarcinoma of the lung and exposure to environmental tobacco smoke (ETS) in 7 countries. We interviewed 70 cases of adenocarcinoma of the lung and 178 population or hospital controls. All subjects had smoked fewer than 400 cigarettes in their lifetimes. Ever exposure to ETS from the parents during childhood was associated with a decreased risk [odds ratio (OR) 0.6, 95% confidence interval (CI) 0.3-1.2], and there was a suggestion of a decreasing trend in risk with increasing duration of exposure. Ever exposure to ETS from the spouse was not associated with an increased risk (OR 1.0, 95% CI 0.5-1.8), while the OR of ever exposure to ETS at the workplace was 1.5 (95% CI 0.8-3.0). For both exposure sources, an increased risk was observed among the highly exposed, and the OR among those with the highest duration of exposure to ETS from the spouse or at the workplace was 1.8 (95% CI 0.5-6.2). A similar risk was estimated for current exposure to ETS from either source. Our results confirm previous reports of a weak effect of adult ETS exposure on risk of adenocarcinoma of the lung. Bias and confounding cannot be excluded as explanations for the apparent decrease in risk from childhood exposure. PMID- 10521801 TI - HGF induces FAK activation and integrin-mediated adhesion in MTLn3 breast carcinoma cells. AB - Expression of hepatocyte growth factor (HGF) and its tyrosine kinase receptor, c Met, is positively correlated with breast carcinoma progression. We found that in invasive and metastatic MTLn3 breast carcinoma cells, HGF stimulated both initial adhesion to and motility on the extracellular matrix (ECM) ligands laminin 1, type I collagen, and fibronectin. Next, analysis with function-perturbing antibodies showed that adhesion to the different ECM proteins was mediated through specific beta1 integrins. In MTLn3 cells, HGF induced rapid tyrosine phosphorylation and activation of both c-Met and focal adhesion kinase (FAK). Cell anchorage and adhesion to the ECM substrates was required for HGF-induced FAK activation, since HGF failed to trigger tyrosine phosphorylation of FAK in suspended cells. Our results provide evidence that the 2 signaling pathways, integrin/ECM and c-Met/HGF, cooperate synergistically to induce FAK activation in an adhesion-dependent manner, leading to enhanced cell adhesion and motility. Moreover, we found that a FRNK (the FAK-related non-kinase)-like molecule is expressed in MTLn3 cells. Since FRNK acts as a competitive inhibitor of FAK function, our results suggest that a FRNK-like protein could facilitate disassembly of focal adhesions and likely be responsible for the HGF-induced scattering and motility of MTLn3 cells. PMID- 10521802 TI - Protein-kinase-C iso-enzymes support DNA synthesis and cell survival in colorectal-tumor cells. AB - Protein-kinase-C signalling has been blocked in colorectal tumor cells by kinase inhibitors, by TPA down-regulation or by exposure to anti-sense oligonucleotides. This resulted in growth inhibition in all cell lines used. The kinase inhibitors H7 and calphostin induced apoptosis, demonstrated by the appearance of cells with characteristically condensed chromatin and the induction of stand-breaks in the DNA. A cell-death-inducing concentration of 15 microgram/ml H7 down-regulated the bcl-2 levels after 9 hr, while bak levels were not affected. Go6976,-an inhibitor of Ca(++)-dependent PKC iso-enzymes, was not active in growth inhibition or induction of apoptosis. Analysis of DNA synthesis in inhibitor-treated cultures indicated that H7 caused strong inhibition in all cell lines, while the more specific inhibitor calphostin was effective only in VACO235 adenoma cells. When down-regulation by TPA or anti-sense oligonucleotides was used to block PKC, effects on cell numbers were smaller and delayed. However, induction of apoptosis was significantly increased in SW480 carcinoma cells 4 days after exposure to anti-epsilon and anti-zeta oligonucleotides in SW480 and T84 carcinoma cells. Apoptosis was preceeded by loss of PKC protein and of bcl-2 from day 1 after addition of the oligonucleotides. In VACO235 adenoma cells, no induction of apoptosis could be observed when anti-epsilon and anti-zeta oligonucleotides were used. On the other hand, the adenoma cells were more responsive to anti-alpha and anti-beta oligonucleotides, which strongly inhibited DNA-synthesis 3 days after addition to the culture medium. Our results indicate that the Ca(++)-dependent PKCs alpha and beta are involved in proliferation signals, while the Ca(++) independent PKCs epsilon and zeta are involved in survival pathways of colorectal tumor cells. PMID- 10521803 TI - Pre-clinical evaluation of [(111)In-DTPA-Pro(1), Tyr(4)]bombesin, a new radioligand for bombesin-receptor scintigraphy. AB - Bombesin (BN) is a 14-amino-acid neuropeptide with a high affinity for the gastrin-releasing peptide receptor. This receptor has been found to be expressed in a variety of tumours, including lung, breast, prostate and pancreas. A newly synthesized BN analogue, [DTPA-Pro(1),Tyr(4)]BN, was shown to be a high-affinity BN-receptor (BNR) agonist, stimulating prolactin secretion from 7315b cells with an IC(50) of 8 nM. The (111)In-labelled analogue was found to bind with high affinity to rat BNR in vitro and in vivo. The radioligand is internalized by BNR expressing cells, in contrast to DTPA-conjugated BN antagonists. Therefore, we further studied the biodistribution of i.v. injected [(111)In-DTPA Pro(1),Tyr(4)]BN in rats. High and specific uptake was found in tissues of the gastrointestinal tract, notably pancreas. Uptake of radioactivity was blocked by pre- or co-injection of 100 microgram [Tyr(4)]BN, but not when this was administered 30 min after the radioligand. This suggests BNR-mediated internalization of the radioligand within 30 min. The percentage injected dose (ID) taken up by BNR-positive tissues was a bell-shaped function of the amount (0.01-0.1 microgram) of injected ligand. Next to the pancreas, highest uptake was observed in the kidneys, which was not blocked by excess [Tyr(4)]BN. Dynamic gamma camera studies showed rapid clearance of radioactivity from the blood compartment. Urinary excretion amounted to about 35% ID after 1 hr and to 70% ID after 24 hr, with a total body retention of 10% ID. Specific uptake was found in the BNR-positive CA20948 pancreas tumour and CC531 colon carcinoma in tumour bearing rats. The CA20948 tumour, inoculated in the hindleg, was also visualized scintigraphically. [(111)In-DTPA-Pro(1), Tyr(4)]BN appears to be a promising radioligand for scintigraphy of BNR-expressing tumours. PMID- 10521804 TI - Quantitative evaluation of the expression of MAGE genes in tumors by limiting dilution of cDNA libraries. AB - The MAGE-A genes are expressed in tumor cells but not in healthy tissues, except in male germ line cells and in placenta. They encode tumor-specific antigens recognized by autologous cytolytic T lymphocytes (CTLs). On the basis of semi quantitative reverse transcription-polymerase chain reaction (RT-PCR) assays, 6 of the 12 members of the MAGE-A family, including MAGE-A1, were previously reported to have a high level of expression in tumors, whereas 5 other members, including MAGE-A10, were expressed at a much lower level, deemed to be insufficient for CTL recognition. However, analysis with antibodies has shown that some melanoma cell lines contain equivalent amounts of MAGE-A1 and MAGE-A10 proteins. This discrepancy appeared to be due to the low efficacy of the primers that had been used for the previous MAGE-A10 RT-PCR assays. This led us to develop a method that is independent of the efficacy of the PCR primers to evaluate MAGE-A gene expression. cDNA libraries from tumor cell lines were introduced into bacteria, of which 200 pools of about 500 bacteria were maintained in microcultures. The frequencies of the MAGE-A cDNA clones in each library were evaluated by performing PCR assays on each of these pools. The abundance of MAGE-A10 cDNAs was found to be similar to that of MAGE-A1 in 3 of the libraries that were analyzed, including 2 with high expression (1/6,400), confirming that MAGE-A10 is expressed at a high level. MAGE-A2, A3, A4, A6 and A12 cDNAs were also confirmed often to be present at a frequency of more than 1/10,000, a level of expression that should suffice for recognition of antigenic peptides encoded by these genes by cytolytic T cells. The remaining MAGE genes are either not expressed in tumors or are expressed at a very low level, with the exception of MAGE-A8 and 11, which show high expression in a very small number of tumors. This method also allowed us to isolate 5 MAGE-A cDNAs that we had not obtained previously, enabling us to delineate the exons in the sequences of genes MAGE-A5, A8, A9, A10 and A11. PMID- 10521806 TI - Genetics of susceptibility to radiation-induced lymphomas, leukemias and lung tumors studied in recombinant congenic strains. AB - The genetic control of susceptibility to radiation-induced tumors in mice has been tested using the series of 20 CcS/Dem (CcS) Recombinant Congenic Strains, each carrying a different random set of 12.5% of genes of the resistant strain STS/A (STS) on the genetic background of the susceptible strain BALB/cHeA (BALB/c). Two classes of tumors were frequently observed: tumors of the haematopoietic system (lymphomas, myelocytic leukemias) and lung tumors. The results indicate that the genes controlling various aspects of tumor development were segregated in the CcS strain series. Large inter-strain differences were observed in the incidence of lung tumors. With lymphomas and leukemias, we not only observed strain differences in the incidence of tumors and in the latency of their development but also in the type of tumors (T- vs. B-cell lymphomas, myelocytic tumors) and in the frequency of their localized or disseminated (leukemic) form. Surprisingly, the myelocytic tumors, which occur very rarely or not at all in the parental strains BALB/c and STS or in their crosses, developed with high frequency in one of the CcS strains (CcS-2), indicating a unique combination of genes in this strain, which facilitates the development of myelocytic tumors. The effect of these genes is suppressed in the genetic composition of the parental strains. Tests of crosses of the resistant-strain CcS 13 with BALB/c indicated a suggestive linkage of a susceptibility gene for lymphomas to chromosome 5. These tests of the CcS strains illustrate the genetic complexity of the control of radiation-induced tumors in mice and suitability of these model systems to study their different facets. PMID- 10521805 TI - Inhibition by rat C-erbB-2/neu antisense oligonucleotide of gastric carcinogenesis induced by N-methyl-N'-nitro-N-nitrosoguanidine in Wistar rats. AB - The effect of prolonged administration of a rat C-erbB-2/neu (C-erbB-2) antisense oligonucleotide on gastric carcinogenesis induced by N-methyl-N'-nitro-N nitrosoguanidine (MNNG) and on the labeling and apoptotic indices of gastric cancer was examined in Wistar rats After oral treatment with MNNG for 25 weeks, the rats received intraperitoneal injections of a C-erbB-2 antisense-liposome complex or a sense-liposome complex at a dose of 50 microgram oligonucleotide/kg body weight every other day until the end of the experiment in week 52. In week 52, the incidence of gastric cancers was significantly lover in rats treated with the C-erbB-2 antisense oligonucleotide than in rats treated with the sense oligonucleotide. Administration of the C-erbB-2 antisense oligonucleotide also significantly decreased the bromodeoxyuridine-labeling index and significantly increased the apoptotic index of gastric cancers. The mean cellular fluorescence of gastric antral cells in MNNG-treated rats was positively correlated with the dose of FITC-labeled C-erbB-2 antisense oligonucleotide. Our findings indicate that the antisense oligonucleotide inhibits gastric carcinogenesis through decreased cell proliferation and increased apoptosis induction and suggest that antisense strategies may provide new treatment for gastric cancer. PMID- 10521807 TI - Long-term carcinogenicity of pan masala in Swiss mice. AB - Carcinogenicity of pan masala, a dry powdered chewing mixture of areca nut, catechu, lime, spices and flavoring agents was evaluated by means of the long term animal bio-assay 6- to 7-week old male and female S/RVCri mice were divided randomly into intermediate and lifetime exposure groups and fed normal diet without pan masala-(zero dose) or diet containing 2.5% and 5% pan masala. Animals in the intermediate-exposure group (n = 10/gender/dose group) were killed after 6, 12 or 18 months of treatment, while those in the lifetime-exposure group (n = 54/gender/dose group) were killed when moribund or at the termination of the experiment at 24 months. Several tissues were processed for histopathological examination. The body weight and survival rate of mice fed pan masala were lower than that of the controls. Histopathological observations of tissues from control animals did not reveal any neoplastic alterations. However, lifetime feeding of pan masala induced adenoma of the liver, stomach, prostate and sebaceous glands, also forestomach papilloma, liver hamartoma, hepatoma and hemangioma, carcinoma of the forestomach, adenocarcinoma of the lung and liver, and testicular lymphoma. Neoplastic lesions appeared mainly in the liver (n = 13), stomach (n = 3) and lung (n = 8). Lung adenocarcinoma, the most frequent malignant tumor type, was observed in 2/120 mice in the intermediate-exposure group and in 8/216 animals in the lifetime-exposure group. Statistical analysis of tumor-induction data revealed a significant dose-related increase in lung adenocarcinomas but not in liver and stomach neoplasms indicating that lung is the major target tissue for the carcinogenic action of pan masala. PMID- 10521808 TI - Suppression by ganglioside GD1A of migration capability, adhesion to vitronectin and metastatic potential of highly metastatic FBJ-LL cells. AB - Ganglioside GD1a, which is highly expressed in poorly metastatic FBJ-S1 cells, has been shown to inhibit the serum-induced migration capability of highly metastatic FBJ-LL cells. In the present study, the capacity of FBJ-S1 cells to adhere to vitronectin was found to be about half that of FBJ-LL cells. Pre treatment of FBJ-LL cells with GD1a decreased this capacity by 30% that of the control, whereas GM1-pre-treatment caused only a 10% decrease, indicating that GD1a specifically inhibits FBJ-LL cell adhesion to vitronectin. Since FBJ-LL cells contain almost no GD1a, transfectants capable of expressing GD1a to varying degrees were produced in this study by transfection of FBJ-LL cells with GM2/GD2 synthase cDNA. Decrease in the serum-induced migration capacity of these transfectants was accompanied by an increment in GD1a expression. Adhesion of the transfectants to vitronectin decreased by 30% as compared with mock-transfected cells. Within 4 to 5 weeks after GD1a-expressing transfectant and mock transfected cells were transplanted into mice, metastatic nodules were observed in liver, lung, kidney and adrenal glands of mock-transplanted mice, but not in those with GD1a-expressing transfectants, indicating that GD1a suppresses the metastasis of FBJ-osteosarcoma cells, possibly by inhibiting cell migration and cell adhesion. The involvement of the ganglioside in the suppression of metastasis is clearly demonstrated in the present study. PMID- 10521809 TI - N-cadherin-mediated adhesion and aberrant catenin expression in anaplastic thyroid-carcinoma cell lines. AB - The role of cadherins and catenins in the progression of thyroid carcinoma is unclear. We have investigated alpha-, beta- and gamma-catenins and p120(ctn) in relation to the expression of cadherins in human anaplastic thyroid-carcinoma cell lines (HTh7, HTh74, C643 and SW1736) with Western blotting and immunofluorescence. E-cadherin was lacking except in SW1736, which consisted of E cadherin-positive (approx. 5%) and -negative cells. The alpha- and beta-catenin levels were similar to those of primary cultured non-neoplastic (E-cadherin positive) human thyrocytes. In contrast, the expression of gamma-catenin was low and variable, correlating with the different levels of cytokeratin in the same cells (HTh74 > SW1736 > C643 > HTh7). p120(ctn) resolved as a doublet in Western blots; the approximately 100-kDa band also found in non-neoplastic epithelial cells was reduced whereas the approximately 115-kDa band, corresponding to the fibroblast-type isoform of p120(ctn), was neo-expressed. A DNA-demethylating agent, 5-aza-2'-deoxycytidine, up-regulated E-cadherin in SW1736 and gamma catenin in SW1736 and C643, whereas the other cell lines were unresponsive; other catenins were not affected. The catenins were generally distributed along the cell borders. Immunostaining, cell-surface biotinylation and co immunoprecipitation revealed that all cell lines expressed N-cadherin in connection with beta-catenin at the plasma membrane. Incubation with an N cadherin antibody disrupted cell-cell adhesion. We conclude that E-cadherin negative anaplastic thyroid-carcinoma cell lines display functional N cadherin/beta-catenin complexes, partial or complete loss of gamma-catenin, and isoform shift of p120(ctn). The unequal expression of E-cadherin and gamma catenin and the variable response to DNA de-methylation suggest that anaplastic thyroid carcinoma is not a uniform entity. PMID- 10521810 TI - A functional and quantitative mutational analysis of p53 mutations in yeast indicates strand biases and different roles of mutations in DMBA- and BBN-induced tumors in rats. AB - In order to analyze the mutational events and to understand the biological significance of the p53 gene in chemical carcinogenesis, we applied a new yeast based p53 functional assay to ovarian tumors induced by 7, 12 dimethylbenz[a]anthracene (DMBA), as well as to transitional cell carcinomas of the urinary bladder induced by N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) in rats. The assay demonstrated that 15 of 19 DMBA induced tumors harbored clonal p53 mutations, which is consistent with the expectations of the "clonal expansion" hypothesis. The majority of the mutations were purine (AG) to pyrimidine (CT) transversions (12/19) on the non-transcribed (sense) strand (NTS), which is likely to be due to depurination created by DMBA adduct formation on the NTS. In contrast, we found no pyrimidine to purine [corrected] transversion on the NTS. After cessation of BBN treatment, BBN-induced multifocal lesions in the bladder contained heterogeneous p53 mutations at an early stage. In the later stage, however, clonal p53 mutations were identified in 4 out of 7 bladders analyzed, conforming with the concept of "field cancerization". The observed base substitutions were G-->A (1/6) or C -->T transitions (2/6), and mutations at T (3/6) on the NTS in clonal mutations, together with non-clonal mutations, showing a preference of C-->T to G-->A (17 vs. 0). Thus, preferential repair was found in the transcribed strand of the p53 gene, whether modified by DMBA or by BBN carcinogens. Very similar mutation patterns were observed between clonal and non-clonal mutations in the DMBA- and BBN-induced tumors, indicating that the rat yeast p53 functional assay can be a potential tool for the characterization of in vivo mutation patterns of p53, when modified by chemical carcinogens. PMID- 10521811 TI - Directory of on-going research in cancer prevention. PMID- 10521812 TI - DNA polymerase beta gene mutation in human breast cancer. PMID- 10521813 TI - The proform of eosinophil major basic protein: a new maternal serum marker for Down syndrome. AB - The proform of eosinophil major basic protein (proMBP), the most abundant protein in the eosinophil specific granule, is synthesized by the placenta and secreted into the maternal circulation, where it is found complex-bound to pregnancy associated plasma protein-A (PAPP-A) and other proteins. We examined the potential of proMBP as a maternal serum marker for fetal Down syndrome (DS) by determining its maternal serum concentration (MSpMBP) in 25 Down syndrome (DS) pregnancies and 152 control pregnancies in the first trimester, and in 105 DS pregnancies and 156 control pregnancies in the second trimester. The median (95 per cent confidence interval) MSpMBP MoM in DS pregnancies (n=15) was 0.66 (0.49 0.79) in gestational weeks 5-9; 1.06 (0.71-1.97) in weeks 10-12 (n=10) and 1.62 (1.18-1.98) in weeks 14-20 (n=105). Using parameterized receiver operator characteristics analysis for proMBP as a single marker for DS, detection rates (DRs) of 22 per cent and 38 per cent, for false-positive rates (FPRs) of 5 per cent, were found in weeks 5-9 (using MSpMBP/=cut-off), respectively. When age and MSpMBP were used as markers in combination, a DR of 36.8 per cent for an FPR of 5.5 per cent was obtained in weeks 5-9 using a risk cut-off of 1:250. In weeks 14-20 the DR was 48.4 per cent for an FPR of 5.3 per cent using the same risk cut-off. This makes proMBP a marker comparable in diagnostic efficiency to human chorionic gonadotrophin (hCG), and exceeding that of alpha-fetoprotein (AFP) and unconjugated oestriol (uE3), in the second trimester. PMID- 10521815 TI - Maternal urine hyperglycosylated hCG in pregnancies with Down syndrome. AB - Stored maternal urine samples were used to determine the distribution of hyperglycosylated human chorionic gonadotrophin (H-hCG) levels in pregnancies with Down syndrome. A total of 349 samples from singleton pregnancies, including 45 with Down syndrome, were tested at 10-19 weeks' gestation. Urinary concentration was allowed for by expressing H-hCG in ng per mmol creatinine. The median level in Down syndrome was 3.63 multiples of the gestation-specific median in unaffected pregnancies (p<0.0001, Wilcoxon rank-sum test, two-tail). However, creatinine levels were relatively low in cases and creatinine did not fully correct for concentration. When this bias was allowed for, the median level was 3.34 multiples of the normal median (MoM). The H-hCG elevation in affected pregnancies was more marked at 14 weeks' gestation or later: a median of 4.64 MoM and allowing for creatinine bias 4.46 MoM. PMID- 10521814 TI - Collaborative study of maternal urine beta -core human chorionic gonadotrophin screening for Down syndrome. AB - Several studies have shown that second-trimester maternal urine beta-core human chorionic gonadotrophin (hCG) levels are raised on average in Down syndrome pregnancies. However, in all but one, testing was retrospective after extended sample storage and so we carried out a large international multicentre prospective study. 16 centres provided 6730 samples from 14-19 week pregnancies: 39 with Down syndrome, 12 with Edwards' syndrome, 42 with other aneuploidies, 52 unaffected twins and 6585 singleton unaffected pregnancies. Samples were from those having routine maternal serum screening in 6 centres and invasive prenatal diagnosis for reasons unrelated to maternal serum screening in 10 centres. Normalized levels of beta-core hCG (nmom/mmol creatinine) were expressed as multiples of the gestation-specific normal median (MoMs). The median beta-core hCG level in Down syndrome was 1.70 MoM (95 per cent confidence interval, 1.26 2.30); 14 (36 per cent) exceeded the normal 90th centile and 9 (23 per cent) the 95th centile. The median level in Edwards' syndrome was 0.23 MoM. On the basis of our results alone it is unlikely that urinary beta-core hCG will be a useful marker in Down syndrome screening practice. But the considerable variability in results between studies means that further research is needed before a reliable conclusion can be drawn. PMID- 10521816 TI - Fetal muscle biopsy as a diagnostic tool in Duchenne muscular dystrophy. AB - Duchenne muscular dystrophy (DMD) is a relentless progressive disorder, leading to severe disability during childhood and death in adolescence or early adulthood. In most families, prenatal diagnosis is readily achieved by molecular detection of DNA deletions using chorionic villi or amniocytes, or by linkage analysis. In some cases, however, molecular methods fail to provide a definitive diagnosis and in such cases in utero fetal muscle biopsy may serve as a diagnostic option. We describe three families in whom fetal muscle biopsy was performed, focusing on the prenatal diagnostic dilemmas, the indications and timing for in utero fetal muscle biopsy, and the difficulties encountered. PMID- 10521817 TI - Accuracy of the haemoglobin alkaline denaturation test for detecting maternal blood contamination of fetal blood samples for prenatal karyotyping. AB - The haemoglobin alkaline denaturation test was routinely performed in 183 fetal blood samples obtained by cordocentesis for prenatal karyotyping by adding 0.1 ml of the blood into a glass test tube containing 5 ml of water and 0.3 ml of 10 per cent KOH as the alkali reagent. The mixture was agitated gently and read at 2 minutes, at which time it was interpreted as a pure fetal blood sample or contaminated with maternal blood according to the change in colour. In order to determine the accuracy of this test to detect maternal blood contamination, the results were compared with the number of fetal and maternal cells detected by standard cytogenetic techniques in those blood samples obtained from male fetuses (n=97). Among these samples, the haemoglobin alkaline denaturation test gave an adult haemoglobin reaction in two cases (2.1 per cent); both samples showed different degrees of maternal 46,XX cells in the metaphases examined (29 of 30 cells in one case and 2 of 31 cells in the other). Conversely, of the 95 samples which gave a fetal haemoglobin reaction, the cytogenetic analysis did not reveal any maternal cells in the metaphases analysed (median 30 cells, range 20-65). We concluded that the haemoglobin alkaline denaturation test is an accurate method for excluding clinically significant maternal blood contamination of fetal blood samples obtained for prenatal karyotyping. This simple, inexpensive technique provides immediate information and, therefore, can be safely incorporated as a bedside test for analysis during fetal blood sampling procedures. PMID- 10521818 TI - Prenatal diagnosis of lysosomal storage diseases using fetal blood. AB - Lysosomal storage diseases are a rare but significant cause of non-immune hydrops fetalis (NIHF). In 17 cases of NIHF detected by ultrasound, the activity of five lysosomal enzymes was measured in leukocytes or plasma of 1 ml of fetal blood obtained by cordocentesis. By this approach seven lysosomal storage diseases known to present with hydrops fetalis can be diagnosed. In this series one case of mucopolysaccharidosis VII (M. Sly) was diagnosed at 20 weeks' gestation. The other samples allowed the establishment of reference ranges for lysosomal enzymes associated with NIHF in fetal blood. We conclude that, also in view of the poor prognosis of lysosomal storage diseases presenting with hydrops fetalis, the use of fetal blood for the early and fast biochemical diagnosis of these diseases is a valuable supplement in the diagnostic work-up and the management of NIHF. PMID- 10521819 TI - Prenatal diagnosis on fetal cells obtained from maternal peripheral blood: report of 66 cases. AB - The potential use of fetal cells circulating in maternal blood for a non-invasive prenatal diagnosis has been widely described. Several authors have developed different methods for the enrichment of fetal cells from maternal peripheral blood. The aim of this study was to make a practical valuation of this new prenatal diagnosis technique, using those methods described as efficient and easy to carry out in a prenatal diagnosis unit. These methods consist of the double density gradient and the positive selection by magnetic activated cell sorting (MACS) of the fetal erythroblasts, and the posterior study of the cells applying the FISH interphasic technique. Once the technique was ready, we obtained results from the study of 66 venous blood samples from women coming for prenatal diagnosis. Using a specific staining for fetal haemoglobin, fetal cells were identified in 63 cases. Fetal sex was well determined in 56 cases, 23 females and 33 males; in 7 cases the sex determination failed. All the aneuploidies found in a previous prenatal diagnosis were confirmed. PMID- 10521820 TI - Cloning of multiple keratin 16 genes facilitates prenatal diagnosis of pachyonychia congenita type 1. AB - Pachyonychia congenita type 1 (PC-1) is an autosomal dominant ectodermal dysplasia characterized by severe nail dystrophy, focal non-epidermolytic palmoplantar keratoderma (FNEPPK) and oral lesions. We have previously shown that mutations in keratin K16 cause fragility of specific epithelia resulting in phenotypes of PC-1 or FNEPPK alone. These earlier analyses employed an RT-PCR approach to avoid amplification of K16-like pseudogenes. Here, we have cloned the K16 gene (KRT16A) and two homologous pseudogenes (psiKRT16B and psiKRT16C), allowing development of a genomic mutation detection strategy based on a long range PCR, which is specific for the functional K16 gene. We report a novel heterozygous 3 bp deletion mutation (388del3) in K16 in a sporadic case of PC-1. The mutation was detected in genomic DNA and confirmed at the mRNA level by RT PCR, showing that our genomic PCR system is reliable for K16 mutation detection. Using this system, we carried out the first prenatal diagnosis for PC-1 using CVS material, correctly predicting a normal fetus. This work will greatly improve K16 mutation analysis and allow predictive testing for PC-1 and the related phenotype of FNEPPK. PMID- 10521821 TI - A retrospective evaluation of second-trimester serum screening for fetal trisomy 18: experience of two laboratories. AB - A retrospective study on screening methods for fetal trisomy 18 has been carried out in two different laboratories using the serum parameters: total human chorionic gonadotropin (hCG), unconjugated oestriol (uE3), and alpha-fetoprotein (AFP) in different combinations and in single marker protocols. Laboratory A (L(A)) utilized a radio-immunoassay to examine 38 fetal trisomy 18 cases and laboratory B (L(B)) utilized an enzyme-immunoassay to examine 33 trisomy 18 cases. As unaffected references the whole routine cohorts of each laboratory were used (L(A): 29 043; L(B): 4264). In both trisomy 18 study groups the median hCG and uE3 multiples of the median (MoM) values were markedly declined (L(A): 0.21 MoM, 0.37 MoM; L(B): 0.31 MoM, 0.44 MoM). Even after exclusion of trisomy 18 cases with combined neural tube or ventral wall defects the medians of AFP MoM values were only moderately declined (L(A): 0.73 MoM; L(B): 0.8 MoM). Receiver operator characteristic (ROC) curves after multivariate discriminance analysis and single marker evaluation demonstrated that the difference of efficiency between a combination of hCG, uE3 and AFP, and a combination of hCG and uE3 is small but that any of these combinations are more efficient than a combination of hCG and AFP or single marker protocols, respectively. At a risk cut-off generating a false-positive rate of one per cent the most effective marker combination detected 31 of 38 (81.6 per cent) affected pregnancies in L(A) and 25 of 33 (75.8 per cent) in L(B). The differences in sensitivity and specificity seem to be due to the different analytical systems being utilized by the two laboratories. PMID- 10521823 TI - Prenatal diagnosis for facioscapulohumeral muscular dystrophy (FSHD). AB - This study outlines the molecular DNA findings derived from 12 separate prenatal diagnoses offered to families with a history of facioscapulohumeral muscular dystrophy. A high risk of the fetus being affected was identified in five pregnancies. Several practical problems are discussed, particularly those arising from the quality and quantity of DNA made available for molecular diagnosis. Evidence of the 4q35 and 10q26 telomeric exchanges is present in 20 per cent of the general population and the specificity of the test is 95 per cent. The eventual isolation and functional characterization of the FSHD gene should allow us to unravel many of the complexities currently associated with the molecular diagnosis of this disorder. PMID- 10521822 TI - Elevated second-trimester maternal serum hCG in patients undergoing haemodialysis. AB - Prenatal Down syndrome screening with maternal serum alpha-fetoprotein (AFP) and human chorionic gonadotrophin (hCG) has become common. High levels of maternal serum hCG and low levels of AFP have been associated with an increased risk of fetal Down syndrome. In this paper, we report five pregnancies in patients undergoing long-term haemodialysis, all of whom had false-positive second trimester Down syndrome screening results. All of our five patients had extremely high levels of maternal serum hCG, but normal AFP values for their gestational age, and all had serious complications during pregnancy. PMID- 10521824 TI - Non-lethal arthrogryposis multiplex congenita presenting with cystic hygroma at 13 weeks gestational age. AB - Arthrogryposis is defined as multiple joint contractures, the aetiology of which is variable. Prenatal diagnosis has focused on diminshed fetal movement and detection of joint contractures on ultrasound. These findings usually do not become evident until 16 to 18 weeks of gestation. Although others (Baty, 1989; Hyett et al., 1997) have reported the diagnosis of arthrogryposis in the first and early second trimester by the presence of nuchal oedema, these reports have all focused on lethal conditions. We report on two female siblings with non lethal arthrogryposis multiplex congenita. The diagnosis was suspected in the second pregnancy at 13.5 weeks when a large cystic hygroma was detected on ultrasound. Multiple joint contractures became evident at 18 weeks. We hypothesize that the aetiology may be secondary to delay in lymphatic maturation with development of a large cystic hygroma resulting in restriction of fetal movement during early joint formation. Further, the fact that the two female siblings had a similar pattern of facial and joint development, and that their parents are second cousins, suggests an autosomal recessive basis for this form of AMC. PMID- 10521825 TI - The cardiac echogenic focus. PMID- 10521826 TI - Prenatal diagnosis of mosaicism for partial trisomy 8: a case report including fetal pathology. AB - A case of prenatally diagnosed partial trisomy 8 is described. The 'syndrome' is associated with skeletal and cardiac anomalies, as well as hepatic calcification. Differing proportions of 47,XY, +der(8) and 46 XY were present in the different fetal tissues sampled. The highest proportion of 47,XY,+der(8) cells was found in the placenta. PMID- 10521827 TI - Transplacental exposure to antipsychotic drugs during pregnancy and megacystis in the fetus. AB - Urinary retention is an adverse effect of antipsychotic drugs that has not been previously reported in the fetus. We have diagnosed megacystis in a fetus possibly caused by transplacental exposure to the antipsychotic drugs being administered to the mother. Two weeks after the mother stopped taking the drugs, the size of the enlarged fetal urinary bladder returned to normal. Sonographic examination of the neonate revealed an anatomically normal urinary tract, with a normal bladder capacity and urination. PMID- 10521828 TI - Sonographic diagnosis of limb reduction defects in a fetus with haemoglobin Bart's disease at 12 weeks of gestation. AB - Limb reduction defect is a rare event but is found in eight per cent of fetuses affected by haemoglobin Bart's disease. We present a case of haemoglobin Bart's disease with terminal transverse limb reduction defects of all four limbs diagnosed by abdominal ultrasound examination at 12 weeks of gestation. The pregnancy was terminated by suction curettage. Just prior to the procedure, transabdominal needle embryoscopy was performed and this confirmed the sonographic diagnosis. The present case demonstrates the need and feasibility of a detailed anatomic survey of a fetus affected by haemoglobin Bart's disease at 12 weeks. This is particularly relevant if the patient is considering the option of intra-uterine therapy. PMID- 10521829 TI - Prenatal diagnosis of partial trisomy 3p(3p23-->pter) and monosomy 7q(7q36- >qter) in a fetus with microcephaly alobar holoprosencephaly and cyclopia. AB - We report the prenatal diagnosis of partial trisomy 3p(3p23-->pter) and monosomy 7q(7q36-->qter) in a fetus with microcephaly, alobar holoprosencephaly and cyclopia. A 26-year-old primigravida woman was referred for genetic counselling at 23 gestational weeks due to sonographic findings of intra-uterine growth retardation and cranio-facial abnormalities. Level II ultrasonograms further demonstrated alobar holoprosencephaly, a proboscis above the eye and a single median orbit consistent with cyclopia. Genetic analysis and fluorescence in situ hybridization on cells obtained from amniocentesis showed distal 3p trisomy (3p23 ->pter) and 7q36 deletion, 46,XX,der(7)t(3;7)(p23;q36), resulting from a paternal t(3;7) reciprocal translocation. The pregnancy was terminated. Autopsy further confirmed the presence of arrhinencephaly, agenesis of the corpus callosum and a single ventricle of the brain. The phenotype of this antenatally diagnosed case is compared with those observed in 10 previously reported cases with simultaneous occurrence of partial trisomy 3p and terminal deletion 7q. All cases are associated with severe forms of holoprosencephaly and facial dysmorphism. This delineates an autosomal imbalance syndrome or a dosage effect involving duplication of distal 3p/deficiency of terminal 7q and dysmorphogenesis of the forebrain and mid-face. PMID- 10521830 TI - Fetus with long QT syndrome manifested by tachyarrhythmia: a case report. AB - We encountered a fetus who exhibited transient (at most 30 s), repeated episodes of tachyarrhythmia (240 bpm). This female neonate was born at 36 weeks of gestation and showed a markedly prolonged QT interval and transient, repeated episodes of polymorphic ventricular tachycardia. Congenital long QT syndrome was diagnosed. Retrospective analysis of the videotape showing fetal cardiac movement revealed that atrio-ventricular dissociation was present prenatally and thus, the fetal tachyarrhythmia was due to ventricular tachycardia. To our knowledge, there are few reports of a fetus with the long QT syndrome who exhibited ventricular tachycardia in utero. In the presence of unexplained fetal tachyarrhythmia, long QT syndrome should be considered as a possible underlying cause disorder. The presence of atrio-ventricular dissociation may be useful in prenatal diagnosis of long QT syndrome. PMID- 10521831 TI - Prenatal diagnosis of sanfilippo type A syndrome in a family with S66W mutant allele. PMID- 10521832 TI - Fetal cells in maternal blood: NIFTY clinical trial interim analysis. DM-STAT. NICHD fetal cell study (NIFTY) group. PMID- 10521833 TI - Nuchal translucency and trisomy 18. PMID- 10521834 TI - Preconceptional use of folic acid amongst women of advanced maternal age. PMID- 10521835 TI - Genetic amniocentesis: gestation-specific pregnancy outcome and comparison of outcome following early and traditional amniocentesis. AB - Amniocentesis remains the most common prenatal diagnostic invasive procedure for fetal karyotyping. During counselling prior to this procedure miscarriage rates are often quoted as a single figure. In this review of 2924 amniocenteses, we report that miscarriage rates vary with the gestational age at which the procedure is performed. The total miscarriage rate was 1.0 per cent after early amniocenteses (11 + 0-14 + 6 weeks) and 1.2 per cent after traditional mid trimester amniocenteses (15 + 0-18 + 6 weeks). The rate was greatest (3.1 per cent) for amniocenteses performed after 18 + 6 weeks' gestation. The cumulative miscarriage risk increased from 0.03 per cent one week after the procedure to plateau at 1.1 per cent five weeks after the procedure. The preterm and still birth rates following amniocenteses were similar in early and traditional mid trimester amniocenteses but were significantly higher when amniocenteses were performed after 19 weeks' gestation. Although the incidence of talipes equinovarus was higher after early amniocentesis compared with traditional mid trimester amniocenteses (1.4 per cent versus 0.2 per cent), none of the affected infants required corrective surgery. We conclude that counselling for this procedure should be tailored to each unit's unintended fetal loss rate based on cumulative rates. Such figures should be available to parents to assist them in their decision-making. PMID- 10521836 TI - Termination rates after prenatal diagnosis of Down syndrome, spina bifida, anencephaly, and Turner and Klinefelter syndromes: a systematic literature review. European Concerted Action: DADA (Decision-making After the Diagnosis of a fetal Abnormality). AB - The aims of this systematic literature review are to estimate termination rates after prenatal diagnosis of one of five conditions: Down syndrome, spina bifida, anencephaly, and Turner and Klinefelter syndromes, and to determine the extent to which rates vary across conditions and with year of publication. Papers were included if they reported (i) numbers of prenatally diagnosed conditions that were terminated, (ii) at least five cases diagnosed with one of the five specified conditions, and (iii) were published between 1980 and 1998. 20 papers were found which met the inclusion criteria. Termination rates varied across conditions. They were highest following a prenatal diagnosis of Down syndrome (92 per cent; CI: 91 per cent to 93 per cent) and lowest following diagnosis of Klinefelter syndrome (58 per cent; CI: 50 per cent to 66 per cent). Where comparisons could be made, termination rates were similar in the 1990s to those reported in the 1980s. PMID- 10521837 TI - Second-trimester maternal serum analyte levels associated with fetal trisomy 13. AB - The aim of this study was to determine whether pregnancies affected by fetal trisomy 13 are associated with second-trimester maternal serum analyte levels different from those typical of the unaffected population. Pregnancies with trisomy 13 were identified through cytogenetics laboratories. Those which had second-trimester maternal serum screening analyte measurements were further evaluated. Maternal serum analyte levels for each case and five matched controls were statistically analysed by matched ranked-sum analysis. 28 cases of fetal trisomy 13 were identified. The median AFP, uE3 and hCG levels were 1.35 MoM, 0.71 MoM and 0.90 MoM, respectively. Only uE3 levels were statistically different (p < 0.01) from those for the unaffected population. These data suggest that second-trimester maternal serum AFP, uE3 and hCG levels are not useful in detecting fetal trisomy 13 and protocols already existing for Down syndrome or trisomy 18 screening will not detect the majority of cases of this aneuploidy. PMID- 10521838 TI - The predictive value of findings of the common aneuploidies, trisomies 13, 18 and 21, and numerical sex chromosome abnormalities at CVS: experience from the ACC U.K. Collaborative Study. Association of Clinical Cytogeneticists Prenatal Diagnosis Working Party. AB - We report 611 non-mosaic and 91 mosaic findings of trisomies 13, 18 and 21, and numerical sex chromosome abnormalities in a series of 20,527 CVS, in the Association of Clinical Cytogeneticists U.K. Collaborative Study, the majority with analysis of both direct preparations and cultured cells. No false-positive results were encountered among the 611 non-mosaic cases, making these findings a very reliable indicator of the fetal karyotype. One false-negative case was reported. In contrast, the 91 mosaic abnormalities were unreliable predictors of fetal abnormality. Many were associated with normal outcomes, but a significant proportion of cases of each individual aneuploidy proved genuine. Mosaicism for 45,X, and trisomies 13 and 18 was disproportionately common. 17 of the mosaic cases showed complete discordance between the karyotype from direct preparations and that from cultured cells. All would have resulted in either a false-positive or a false-negative finding if only one technique had been used. Based on our experience, and that of others, we believe that the highest level of predictive accuracy using CVS can only be achieved if both direct preparation and cell culture are performed. In addition, we continue to recommend that all pregnancies demonstrating mosaicism for these aneuploidies at CVS undergo amniocentesis or fetal blood sampling to differentiate between confined placental mosaicism and true fetal karyotypic abnormality. PMID- 10521839 TI - Mohr syndrome in two sisters: prenatal diagnosis in a 22-week-old fetus with post mortem findings in both. AB - Oral-facial digital syndrome type II (OFP syndrome II; orofaciodigital syndrome II) is a rare autosomal recessive syndrome, first described by Mohr (1941). We present two sisters with Mohr syndrome from a consanguineous family. One is a three-day-old female patient, the other is 22-week-old fetus. Polydactyly with bifid thumbs in both hands, bilateral polysyndactyly of halluces, lateral polysyndactyly and bilateral pes equinovarus were demonstrated in the fetus sonographically. Corpus callosum agenesis, congenital heart disease, bilateral bifid thumbs and halluces and polydactyly were seen in both patients. In addition, post-mortem findings showed absence of olfactory nerve, single atrium. VSD, abnormal lung lobulation and natal teeth in the fetus. Absence of olfactory nerve and natal teeth have not been reported previously in Mohr syndrome. PMID- 10521840 TI - Risk of false-positive prenatal diagnosis using interphase FISH testing: hybridization of alpha-satellite X probe to chromosome 19. AB - FISH analysis of uncultured interphase amniotic fluid cells from a male fetus revealed two signals using an alpha-satellite X-chromosome DNA probe. One of the signals was much smaller than the other. It was subsequently shown that the normal sized signal was located on the X chromosome and the smaller signal was located at the centromere of chromosome 19. This hybridization pattern was confirmed in the newborn infant and in his phenotypically normal father. The use of alpha-satellite DNA probes on interphase cells could result in false-positive errors due to rare variants such as the X-chromosome alpha-satellite found on chromosome 19 in our patient. PMID- 10521841 TI - Prenatal diagnosis of glycogen storage disease type IV using PCR-based DNA mutation analysis. AB - Deficiency of glycogen branching enzyme activity causes glycogen storage disease type IV (GSD-IV). Clinically, GSD-IV has variable clinical presentations ranging from a fatal neonatal neuromuscular disease, to a progressive liver cirrhosis form, and to a milder liver disease without progression. Current methods for prenatal and postnatal diagnosis are based on an indirect method of measuring the enzyme activity, which has a limited sensitivity and cannot be used to distinguish patients with these variable clinical phenotypes. In this study, a GSD-IV family with a non-progressive hepatic form of the disease requested prenatal diagnosis. Determination of the branching enzyme activity in cultivated amniocytes showed 20 per cent residual activity overlapping with the level detected in the heterozygotes. Mutation analysis revealed that the fetus carried two mutant alleles, L224P and Y329S, the same as the proband of this family. The fetus was predicted to be affected and postnatally his clinical presentation is consistent with the diagnosis. We conclude that DNA mutation analysis should be used in the prenatal diagnosis of GSD-IV, especially in the situation of high residual enzyme activity. PMID- 10521842 TI - Latent class analysis applied to patterns of fetal sonographic abnormalities: definition of phenotypes associated with aneuploidy. AB - The aim of the present study was to generate different latent variables that classify the major chromosome aneuploidies using frequency and patterns of fetal sonographic abnormalities in a large database. A total of 1867 fetuses with sonographic abnormalities recorded in a database at New England Medical Center from January 1995 to March 1998 were available for the statistical analysis. Included within this group were 61 aneuploid fetuses, including 11 with 45,X, 30 with trisomy 21, 14 with trisomy 18 and 6 with trisomy 13, 40 structural malformations and/or sonographic markers were detected in these 61 aneuploid fetuses. The ability of malformations and sonographic markers to generate different groups of phenotypes was evaluated by means of latent class analysis, using the 61 affected cases. Four different classes were generated with the hypothetical assumption that each of them could satisfactorily identify a respective fetal aneuploidy represented in the study group. Among 40 fetal malformations and/or sonographic markers, the most important findings in generating specific karyotypic groups were cystic hygroma (class 1), duodenal atresia (class 2), holoprosencephaly (class 3) and omphalocele (class 4), respectively. Accuracy of the classification was 72 per cent for Turner syndrome (class 1), 74 per cent for Down syndrome (classes 1 and 2), 88 per cent for trisomy 13 (class 3) and 93 per cent for trisomy 18. The frequency of associated malformations detected sonographically can help to define a phenotype that is likely to be representative of a specific aneuploidy. Before the definitive karyotype is available or, in cases in which patients refuse an invasive prenatal diagnostic procedure, this may improve antenatal clinical management. PMID- 10521843 TI - Detection of aneuploidy in single cells using comparative genomic hybridization. AB - The ability of comparative genomic hybridization (CGH) to detect aneuploidy following universal amplification of DNA from a single cell, or a small number of cells, was investigated with a view to preimplantation diagnosis following in vitro fertilization, and prenatal diagnosis using fetal erythroblasts obtained from maternal blood. The DNA obtained from lysed single cells was amplified using degenerate oligonucleotide-primed PCR (DOP-PCR). This product was labelled using nick translation and hybridized together with normal reference genomic DNA. The CGH fluorescent ratio profiles obtained could be used to determine aneuploidy with cut-off thresholds of 0.75 and 1.25. Deviation in the profiles in the heterochromatic regions was reduced by using, as a reference sample, normal genomic DNA that had also undergone DOP-PCR. Single cells known to be trisomic for chromosomes 13, 18 or 21 were analysed using this technique. The resolution of CGH with amplified DNA from a single cell is of the order of 40 Mb, sufficient for the diagnosis of trisomy 21, and possibly segmental aneuploidy of equivalent size. These results, and those of others, demonstrate that diagnosis of chromosomal aneuploidy in single cells is possible using CGH with DOP-PCR amplified DNA. PMID- 10521844 TI - Management of Rh-immunized pregnancies. PMID- 10521845 TI - Laryngeal atresia presenting as fetal ascites, olygohydramnios and lung appearance mimicking cystic adenomatoid malformation in a 25-week-old fetus with Fraser syndrome. AB - We describe a 25-week-old female fetus of consanguineous parents with ultrasonographic findings of increased echogenicity of lungs mimicking CAM (cystic adenomatoid malformation) type III, olygohydramnios and fetal ascites. A therapeutic abortion was performed and unilateral cryptophthalmos, laryngeal atresia and bilateral syndactyly of the hands and feet were observed at post mortem. These findings confirmed the diagnosis of Fraser syndrome after abortion. PMID- 10521846 TI - Histological findings in a case of alobar holoprosencephaly diagnosed at 10 weeks of pregnancy. AB - A case of alobar holoprosencephaly diagnosed at 10 + 3 weeks' gestation by transabdominal and transvaginal ultrasound examination followed by histological confirmation is presented. The diagnosis was based on two sonographic criteria: intracranial finding of a single ventricle with a mantle and no visible midline structures but fusion of the thalami and corpus striatum, and facial abnormalities, including hypotelorism and proboscis. The fetal karyotype was triploidy. The ultrasound findings were confirmed by pathological examination. The histological findings of proboscis, single lens and single ventricle with neural tissue remnants are presented. PMID- 10521847 TI - Prenatal detection and mapping of a distal 8p deletion associated with congenital heart disease. AB - We report the prenatal diagnosis, at 18 weeks' gestational age of a del(8)(p23.1- >pter) in a fetus with an atrio-ventricular canal, persistent left superior vena cava and hypoplastic right ventricle detected by sonographic imaging. We further refine the breakpoints associated with this defect using fluorescent in situ hybridization analysis (FISH). Our findings correlate with recent reports of the localization and importance of GATA4 (a zinc finger transcription factor) in cardiac development. Though microcephaly, mental retardation and typical behavioural features are well described in various deletions in 8p, the absence of notable microcephaly in this case raises the possibility for a separate genetic aetiology for some of these features. Indeed, primary autosomal recessive microcephaly (MCPH1) was recently mapped to a nearby region and may be the cause for this frequent observation in some cases of 8p deletions. These observations illustrate the role of FISH in prenatal diagnosis and refinement of chromosomal breakpoints. In addition, mappings of loci significant for cardiac development are presented. Our findings suggest that some features of the 8p deletion syndrome may ultimately be uncoupled from one another, and underscore the need for further study of this region of chromosome 8, in order to achieve adequate information for genetic counselling. PMID- 10521848 TI - A paternally inherited terminal deletion, del(8)(p23.1)pat, detected prenatally in an amniotic fluid sample: a review of deletion 8p23.1 cases. AB - A subtle terminal deletion of the short arm of chromosome 8 with a breakpoint in p23.1 was detected in amniocytes. Parental chromosome studies revealed a similar deletion in the father. The fetus did not have any abnormalities in a level II ultrasound. The pregnancy was continued and resulted in the birth of a baby girl. The child was normal at six months of age and no heart murmur was detected. In a retrospective review of cases in our laboratory, four other cases with a deletion del(8)(p23.1) were found. Three were paediatric cases with microcephaly, developmental delay, ASD, VSD, pulmonic stenosis, congenital and behavioural abnormalities. One was a 29-year-old woman with a mosaic karyotype. She had a history of spontaneous abortions and no known cardiac defect. Using conventional cytogenetics and/or FISH studies with 8p telomere probe and 8 painting probe, the 8p23.1 deletions were shown to be either terminal or interstitial. The karyotype from the prenatal case was compared with the other cases of 8p23.1 deletions in our laboratory to see if there was a discernible difference in the size of the deletion. The deletion in the proband seemed to involve a more distal 8p23.1 breakpoint. In the father's high resolution chromosomes (550-850 band level) the breakpoint appeared to be 8p23.1 approximately 23.2 and FISH studies using an 8p telomeric probe confirmed a terminal deletion. Interstitial deletion of sub-band 8p23.1 was associated with phenotypic abnormalities and distal 8p23.2pter deletion was found in apparently normal individuals, therefore, 8p23.1 appears to be the critical region for clinical abnormalities. PMID- 10521849 TI - Prenatal diagnosis of osteogenesis imperfecta type I by COL1A1 null-allele testing. AB - Osteogenesis imperfecta (OI) type I is caused by a reduction of type I collagen resulting from the presence of a non-functional COL1A1 allele (null-allele). Owing to the lack of mutant mRNA, genomic screening of the COL1A1 and COL1A2 genes is required to identify a causal mutation, which is a costly and time consuming endeavour. We have developed an alternative approach for confirmation of a suspected diagnosis of OI type I based on the detection of a COL1A1 null allele. Here we report the application of this COL1A1 null-allele detection test for prenatal diagnosis in a patient with OI type I in which it was shown that the fetus had inherited the normal COL1A1 allele from his affected mother and would not be affected with OI. PMID- 10521850 TI - Prenatal diagnosis of a fetus with distal 10q trisomy. AB - Distal 10q trisomy is a well-defined but rare syndrome. Most cases are diagnosed in infancy or in childhood and rarely include prenatal findings. We present a case of fetal distal 10q trisomy with abnormal prenatal sonographic findings. A 19-year-old primigravida was referred for genetic counselling at 18 gestational weeks because her husband had a familial history of congenital anomalies. Genetic amniocentesis was thus performed and showed fetal distal 10q trisomy (10q24.1- >qter), 46,XX,der(22)t(10;22)(q24.1;p11.2)pat, resulting from paternal t(10;22) reciprocal translocation. Level II ultrasonograms further demonstrated bilateral hydronephrosis, ventricular septal defect and facial dysmorphism ascertained by three-dimensional ultrasound. The pregnancy was terminated at 22 gestational weeks. Post-mortem autopsy confirmed the sonographic findings. We suggest that abnormal prenatal sonographic findings such as cardio-vascular, renal and facial malformations should alert cytogeneticists to search for subtle chromosomal abnormalities. PMID- 10521851 TI - Ellis-van Creveld syndrome: examination at 15 weeks' gestation. AB - In 1940, Ellis and van Creveld defined a syndrome they referred to as chondro ectodermal dysplasia. This autosomal recessive condition, now usually referred to as Ellis-van Creveld syndrome (EVC), comprises bilateral postaxial polydactyly, a chondrodysplasia, characterized by shortness of limbs, and ectodermal dysplasia. Congenital heart defects are also common. There are many reports in medical literature describing affected newborns and even, older children. Here, we report the clinical, radiological and histological findings in a 15-week-old affected fetus. The diagnosis of Ellis-van Creveld syndrome in this fetus is based on a positive family history (an affected sib) and shortness of long bones as well as hexadactyly diagnosed by prenatal ultrasonography. On post-mortem examination, bilateral postaxial hexadactyly and symmetrical shortness of the long bones was noted. Histologically, there was too short a zone of cartilagineous columns in the metaphyses, a reduced number of chondrocytes and an irregularly structured spongiosa within the ossification zone. In addition, the fetus was found to have an atrio-ventricular canal. This heart defect is presumably rare in this syndrome. Other characteristic features such as small and dysplastic nails, sparse hair and abnormalities of the teeth were, of course, not yet present in this early developmental stage. In addition to EVC, the fetus had a 47,XXY chromosome constitution. PMID- 10521852 TI - Segregation of digital number with partial monosomy or trisomy of 13q in familial 5;13 translocation. AB - It has been postulated that the deletion of band 13q22 may be associated with digital malformations, especially thumb and big toe anomalies. We report a family where the mother is carrying a balanced translocation between chromosomes 5p15 and 13q22. The offspring have a specific and well-defined phenotype depending on which is the unbalanced chromosome in the karyotype. When a partial trisomy of 13q22-->qter is present, the fetuses have polydactyly in the four limbs, and when the fetus is carrying a partial monosomy of this portion, an oligodactyly in all members can be observed. PMID- 10521853 TI - A case of discordant related abnormal karyotypes from chorionic villi and amniocytes. AB - A case of three discordant cell lines in prenatal diagnosis is described, of which two were abnormal related structural abnormalities of chromosome 11. One of the abnormal cell lines was seen in all metaphases examined from direct preparations of chorionic villi, the cultured preparations showing an apparently normal male karyotype; the other abnormal cell line was seen in conjunction with a normal cell line in cultured amniocytes. Prenatal diagnosis offered solely on chorionic villus sampling would have yielded a mistakenly normal result on the basis of established criteria for distinguishing true mosaicism from confined placental mosaicism. PMID- 10521854 TI - False-negative findings in chorionic villi. PMID- 10521855 TI - Author's response. PMID- 10521856 TI - Inhibin-A regression. PMID- 10521857 TI - Nothing ventured, nothing gained! PMID- 10521858 TI - Another look at two phase I clinical trial designs. AB - This note is a response to a recent paper by Korn et al. in which two phase I trial designs were compared, the designs in question being the standard design and the CRM design. The authors concluded that: (i) CRM designs will take longer to complete than standard designs; and (ii) CRM designs are less safe than the standard designs. These conclusions followed from a set of simulations for three different dose toxicity situations. The first purpose of this note is to point out that these conclusions lean on false assumptions. The claims are in error. In their comparisons Korn et al. never in fact used CRM, as defined in O'Quigley, Pepe and Fisher, but a modified version. The second purpose of this note is to look at the same cases studied by Korn et al. but this time using a correctly defined CRM model. Using the same comparison tools of Korn et al. we will see that for these situations, and indeed for a very wide class of other situations not presented here, not only are (i) and (ii) not true but that the exact opposite holds. PMID- 10521859 TI - Commentary PMID- 10521860 TI - Explaining heterogeneity in meta-analysis: a comparison of methods. AB - Exploring the possible reasons for heterogeneity between studies is an important aspect of conducting a meta-analysis. This paper compares a number of methods which can be used to investigate whether a particular covariate, with a value defined for each study in the meta-analysis, explains any heterogeneity. The main example is from a meta-analysis of randomized trials of serum cholesterol reduction, in which the log-odds ratio for coronary events is related to the average extent of cholesterol reduction achieved in each trial. Different forms of weighted normal errors regression and random effects logistic regression are compared. These analyses quantify the extent to which heterogeneity is explained, as well as the effect of cholesterol reduction on the risk of coronary events. In a second example, the relationship between treatment effect estimates and their precision is examined, in order to assess the evidence for publication bias. We conclude that methods which allow for an additive component of residual heterogeneity should be used. In weighted regression, a restricted maximum likelihood estimator is appropriate, although a number of other estimators are also available. Methods which use the original form of the data explicitly, for example the binomial model for observed proportions rather than assuming normality of the log-odds ratios, are now computationally feasible. Although such methods are preferable in principle, they often give similar results in practice. PMID- 10521861 TI - Autoregressive models for describing non-linear changes in biological parameters fitted using BUGS. AB - Many biological processes give outcome data which show a curvilinear association with time which tends to an asymptote. We show how autoregressive models can be used to describe this association within individual subjects. We also present a Bayesian approach implemented using statistical software, BUGS, to fit these models in a multi-level (hierarchical) setting that describes variation in the association between subjects. Peak expiratory flow data from a clinical trial involving subjects with asthma are used to illustrate the methods. PMID- 10521862 TI - Classification to ordinal categories using a search partition methodology with an application in diabetes screening. AB - A method is proposed for classification to ordinal categories by applying the search partition analysis (SPAN) approach. It is suggested that SPAN be repeatedly applied to binary outcomes formed by collapsing adjacent categories of the ordinal scale. By a simple device, whereby successive binary partitions are constrained to be nested, a partition for classification to the ordinal states is obtained. The approach is applied to ordinal categories of glucose tolerance to discriminate between diabetes, impaired glucose tolerance and normal states. The results are compared with analysis by ordinal logistic regression and by classification trees. PMID- 10521863 TI - Applying the Cox proportional hazards model for analysis of latency data with interval censoring. AB - The latency time of an infectious disease is defined as the time from infection to disease onset. This paper applies the proportional hazards model to estimate the effect of covariates on latency when the time of disease onset is exact or right-censored but the time of infection is interval-censored. We use a Monte Carlo EM algorithm to estimate parameters of the joint distribution of infection times and latency times. At each EM iteration, exact infection times are multiply imputed from the density determined by the parameters of the infection and latency time distributions. The methodology is tested using a simulation study and is applied to data from a cohort of haemophiliacs with HIV disease. PMID- 10521865 TI - Approximating the power of Wilcoxon's rank-sum test against shift alternatives. AB - Three methods of approximating the power of Wilcoxon's rank-sum test against shift alternatives are studied. They are obtained by using a Gaussian assumption, Edgeworth expansion, or bootstrap. It is assumed that a historical data set is available to use in estimating the shape of the distribution. The methods are compared through simulation across several different distributional types. The results indicate that the bootstrap generally gives the most reliable approximation, however the Edgeworth expansion has the practical advantage that a lower bound on the power can be roughly approximated. The methods are illustrated on muscle strength data from patients with osteogenesis imperfecta. Published in 1999 by John Wiley & Sons, Ltd. This is US Government work and is in the public domain in the United States. PMID- 10521864 TI - A Wald test comparing medical costs based on log-normal distributions with zero valued costs. AB - Medical cost data often exhibit strong skewness and sometimes contain large proportions of zero values. Such characteristics prevent the analysis of variance (ANOVA) F-test and other frequently used standard tests from providing the correct inferences when the comparison of means is of interest. One solution to the problem is to introduce a parametric structure based on log-normal distributions with zero values and then construct a likelihood ratio test. While such a likelihood ratio test possesses excellent type I error control and power, its implementation requires a rather complicated iterative optimization program. In this paper, we propose a Wald test with simple computation. We then conduct a Monte Carlo simulation to compare the type I error rates and powers of the proposed Wald test with those of the likelihood ratio test. Our simulation study indicates that although the likelihood ratio test slightly outperforms the Wald test, the performance of the Wald test is also satisfactory, especially when the sample sizes are reasonably large. Finally, we illustrate the use of the proposed Wald test by analysing a clinical study assessing the effects of a computerized prospective drug utilization intervention on in-patient charges. PMID- 10521866 TI - Instrumental variables when evaluating screening trials: estimating the benefit of detecting cancer by screening. AB - When evaluating the benefit of detecting cancer by screening we try to answer the question, 'what would a screen detected subject's outcome have been if his/her cancer had progressed to clinical detection'. By 'outcome' we mean survival time, cancer size and stage, lead time effects and more. Because only an unethical study can answer it directly, researchers have attempted to answer the question indirectly using data from randomized cancer screening studies (subjects randomized to study (screened) or control (not screened)). Inferences are made by first selecting the cancer cohort (those subjects who are found to have cancer), then comparing subjects having screen detected cancers to subjects having clinically detected cancers. However, there are two difficulties with this approach: (i) because screening (intends to) detect cancers early, at the trial's end the study group contains more cancer cases than the control group and so the cancer cohort has some unidentified control subjects missing (that is, subjects having cancer during the screening period that have not yet been clinically detected); (ii) because screen detected cancers (may) differ from clinically detected cancers, the comparison group should include only a (non-identified) subset of the cancer cohort's control subjects (that is, only those control subjects having cancers that would have been screen detected). Statistical literature acknowledges these difficulties and attempts to solve them separately, but without success; those methods do not yield meaningful causal inferences and admit substantial bias. Recently, Angrist, Imbens and Rubin and Imbens and Rubin provide a framework for instrumental variable methods that we interpret as allowing us to make causal inferences with incompletely identified comparison groups. We apply their framework to evaluating cancer screening trials and find that we may simultaneously accommodate both difficulties while giving a meaningful answer to the question posed above. Using data from a breast cancer screening trial we demonstrate the general method with a variety of outcome measures and extensions. PMID- 10521867 TI - Evaluating the exposure and disease relationship with adjustment for different types of exposure misclassification: a regression approach. AB - Misclassification of exposure can lead to biased results in the epidemiologic research. Available methods accounting for misclassification often require the use of a gold standard or assume non-differential misclassification of exposure. We present a regression approach which can detect and account for different types of misclassification when estimating the exposure and disease relationship. This approach uses two imperfect measures of a dichotomous exposure and does not require a gold standard. Standard statistical packages with a logistic regression module can be used for estimation of parameters through the EM algorithm process. Two examples are used to illustrate the methodology. PMID- 10521868 TI - Extra-medullary relapse of leukemia following allogeneic bone marrow transplantation. AB - The curative effect of allogeneic bone marrow transplantation (BMT) for acute and chronic leukemia is attributed to the intensive conditioning chemotherapy with or without radiotherapy, as well as an immune-mediated graft versus leukemia (GVL) effect. A different pattern of relapse has been observed after allogeneic BMT for patients with leukemia. Compared with treatment using conventional chemotherapy alone, isolated extra-medullary relapse of disease appears to be seen more commonly after allogeneic BMT. While a full donor's hematopoiesis may be retained, prolonged morphological remission has been observed in the recipient's bone marrow. There appears to be a population of leukemic cells with an affinity to extra-medullary tissues. The failure of the leukemic clone to repopulate the recipient's marrow suggests the presence of a more profound GVL effect in the marrow environment. The optimal treatment for extra-medullary relapse of leukemia following allogeneic BMT remains uncertain. In the case of isolated extra medullary relapses following BMT, the leukemia may still be responsive to further treatment with chemotherapy and/or radiotherapy. The prognosis is poor in general, but prolonged survival has been observed in some of these patients. With the preservation of donor's hematopoiesis in the recipient's marrow, the use of intensive chemotherapy followed by donor lymphocyte or stem cell re-infusion is a promising option. PMID- 10521869 TI - Lymphoma at uncommon sites. AB - Lymphoma can often present in unusual situations. This article provides a comprehensive review of the literature in which both non-Hodgkin's lymphoma and Hodgkin's disease are discussed. PMID- 10521870 TI - Lectin histochemistry of the hyaline layer around the larvae of Patiriella species (Asteroidea) with different developmental modes. AB - Larvae of sea stars are surrounded by an extracellular matrix called the hyaline layer. The lectin-binding properties of this matrix were investigated in an ultrastructural study of Patiriella species having different modes of development. The planktonic bipinnaria and brachiolaria of P. regularis and the planktonic brachiolaria of P. calcar demonstrated the same labeling of the hyaline layer for three lectins: Con A, SBA, and WGA. In both species the outer coarse meshwork stained for all three lectins, whereas the intervillous layer displayed patchy labeling. In the benthic brachiolaria of P. exigua, the outer coarse meshwork displayed heavy labeling for all three lectins. The heavy labeling of the outer coarse meshwork of P. exigua compared with that of the other species suggests an increased number of lectin binding sites in the hyaline layer of this species. The similar ultrastructure and histochemistry of the hyaline layer of P. regularis and P. calcar may reflect similar requirements of their extracellular cover in their planktonic environment. Lectin labeling shows that hypertrophy of the hyaline layer of P. exigua, in particular the outer coarse meshwork, involves elaboration of the carbohydrate composition of the matrix. Modifications seen in the ultrastructure and histochemistry of the hyaline layer of P. exigua appear to be associated with the evolution of benthic development. PMID- 10521871 TI - Multiple acrosomal vesicles and their differentiation during spermiogenesis in Ascidia zara and Ascidia gemmata (Ascidiacea, tunicata). AB - A fully differentiated spermatozoon of both Ascidia zara and Ascidia gemmata is approximately 35 microM long. It contains a head and a tail lacking a midpiece. The head (approximately 4 microM long for A. zara and 5 microM long for A. gemmata) contains an elongated nucleus and a single mitochondrion that flanks the nucleus. Multiple acrosomal vesicles (three or four in number) are present at the apex of the sperm head in both species. Each vesicle is approximately 50 x 50 x 60 nm, and contains moderately electron-dense material. During spermiogenesis of A. zara, three or four vesicles appear in a blister of an early stage spermatid. These vesicles transform into multiple acrosomal vesicles without fusing with each other. Spermiogenesis and acrosome differentiation are similar in A. gemmata and A. zara. Three types of acrosome differentiation in ascidians are described. PMID- 10521872 TI - Comparison of isometric contractile properties of the tongue muscles in three species of frogs, Litoria caerulea, Dyscophus guinetti, and Bufo marinus. AB - Previous studies show that anurans feed in at least three different ways. Basal frogs have a broad tongue that shortens during protraction and emerges only a short distance from the mouth. Some frogs have long, narrow tongues that elongate dramatically due primarily to inertia from mouth opening, which is transferred to the tongue. A few species have a hydrostatic mechanism that produces tongue elongation during protraction. This functional diversity occurs among frogs that share the same two pairs of tongue muscles. Our study compares the isometric contractile properties of these tongue muscles among three frog species that represent each feeding mechanism. Nerves to the paired protractors and retractors were stimulated electrically in each species to record the force properties, contraction speeds, and fatigabilites of these muscles. Few differences were found in the isometric contractile properties of tongue muscles, and the greatest differences were found in the retractors, not the protractors. We propose that the unique arrangement of the tongue muscles in frogs results in a retractor that may also be coactivated with the protractor in order to produce normal tongue protraction. Inertial effects from body, head, and jaw movements, along with clear differences that we found in passive resistance of the tongues to elongation, may explain much of the behavioral variation in tongue use among species. PMID- 10521873 TI - Gill-cleaning mechanisms of a dendrobranchiate shrimp, Rimapenaeus similis (Decapoda, penaeidae): description and experimental testing of function. AB - Observations on functional morphology and results from experiments demonstrate that setiferous epipods compose the major gill-cleaning mechanism in a penaeoid shrimp, Rimapenaeus similis. Epipods on the second maxillipeds and on pereopods 1 3 are equipped with long setae bearing an array of digitate scale setules. These multidenticulate setae reach to most gills and are jostled among them during limb movements. Experiments were performed in which epipods were removed from the gill chamber on one side (experimental) but not the other (control); treated animals were exposed to fouling in a recirculating water system for 2 weeks. Particulate fouling, measured by reduction in relative gill transparency, was significantly greater on experimental than control gills. The pereopodal exopods, not previously implicated in gill cleaning in any decapod, were similarly identified as important gill-cleaning structures. Equipped with long multidenticulate setae like those on the epipods, exopods sweep back and forth over the gill filaments just under the gill cover, areas not reached by the epipods. Exopod-ablation experiments were conducted that showed that exopods prevent particulate fouling on gill surfaces over which they sweep. The similarity in action of the passive gill-cleaning system of R. similis to that of crayfish (Bauer [1998] Invert Biol 117:29-143) suggests the hypothesis that the epipodal and exopodal cleaning setae of R. similis are ineffective against epibionts. The reduction in epipodal and exopodal cleaning systems that occurs in the Penaeoidea is hypothesized to be compensated for by increased development of gill-cleaning setae on the branchiostegite, scaphognathite, or other structures. PMID- 10521874 TI - Development of the genital ducts and spermathecae in the Rhyacodrilines rhyacodrilus coccineus and Monopylephorus rubroniveus (Oligochaeta, tubificidae). AB - The male genital duct in Tubificidae consists of a funnel, a vas deferens, an atrium, and, frequently, a copulatory structure. There may also be a diffuse or compact prostate gland in association with the duct. The morphogenesis of this duct is described for Rhyacodrilus coccineus and Monopylephorus rubroniveus (Rhyacodrilinae). The funnel and vas deferens in both species originate from peritoneal (mesodermal) cells in the posterior septum in the testis segment. The atrium in R. coccineus develops from a primary epidermal (ectodermal) invagination. A typical atrium is not formed in M. rubroniveus; the entire duct is of mesodermal origin. In the latter species, a shallow epidermal invagination occurs, into which both male ducts open, but it bears resemblance to a copulatory structure, which usually forms from a secondary invagination, rather than to a proper atrium. We therefore conclude that M. rubroniveus lacks an atrium. The copulatory structure is termed the male bursa. Both species have diffuse prostate glands that differentiate from peritoneal (mesodermal) cells surrounding the male duct. In R. coccineus the cells cover the atrium, whereas in M. rubroniveus they cover only a part of the vas deferens. The development of the spermathecae and female ducts is also examined. The spermatheca is of ectodermal origin in both studied species, i.e., it forms as an invagination of the epidermis. The female duct develops from peritoneal (mesodermal) cells in the posterior septum of the ovary segment. However, in M. rubroniveus the first sign of the duct disappears and a proper duct never develops. PMID- 10521875 TI - Proportions of slow myosin heavy chain-positive fibers in muscle spindles and adjoining extrafusal fascicles, and the positioning of spindles relative to these fascicles. AB - Chicken leg muscles were examined to calculate the percentages of slow myosin heavy chain (MHC)-positive fibers in spindles and in adjacent extrafusal fascicles, and to clarify how the encapsulated portions of muscle spindles are positioned relative to these fascicles. Unlike mammals, in chicken leg muscles slow-twitch MHC and slow-tonic MHC are expressed in intrafusal fibers and in extrafusal fibers, suggesting a close developmental connection between the two fiber populations. In 8-week-old muscles the proportions of slow MHC-positive extrafusal fibers that ringed muscle spindles ranged from 0-100%. In contrast, proportions of slow MHC-positive intrafusal fibers in spindles ranged from 0-57%. Similar proportions in fiber type composition between intrafusal fibers and surrounding extrafusal fibers were apparent at embryonic days 15 and 16, demonstrating early divergence of extrafusal and intrafusal fibers. Muscle spindles were rarely located within single fascicles. Instead, they were commonly placed where several fascicles converged. The frequent extrafascicular location of spindles suggests migration of intrafusal myoblasts from developing clusters of extrafusal fibers toward the interstitium, perhaps along a neurotrophic gradient established by sensory axons that are advancing in the connective tissue matrix that separates adjoining fascicles. PMID- 10521877 TI - Meeting announcements PMID- 10521878 TI - A message from the editor PMID- 10521876 TI - Craniofacial sutures: morphology, growth, and in vivo masticatory strains. AB - The growth and morphology of craniofacial sutures are thought to reflect their functional environment. However, little is known about in vivo sutural mechanics. The present study investigates the strains experienced by the internasal, nasofrontal, and anterior interfrontal sutures during masticatory activity in 4-6 month-old miniature swine (Sus scrofa). Measurements of the bony/fibrous arrangements and growth rates of these sutures were then examined in the context of their mechanical environment. Large tensile strains were measured in the interfrontal suture (1,036 microepsilon +/- 400 SD), whereas the posterior internasal suture was under moderate compression (-440 microepsilon +/- 238) and the nasofrontal suture experienced large compression (-1,583 microepsilon +/- 506). Sutural interdigitation was associated with compressive strain. The collagen fibers of the internasal and interfrontal sutures were clearly arranged to resist compression and tension, respectively, whereas those of the nasofrontal suture could not be readily characterized as either compression or tension resisting. The average linear rate of growth over a 1-week period at the nasofrontal suture (133.8 micrometer, +/- 50.9 S.D) was significantly greater than that of both the internasal and interfrontal sutures (39.2 micrometer +/- 11.4 and 65. 5 micrometer +/- 14.0, respectively). Histological observations suggest that the nasofrontal suture contains chondroid tissue, which may explain the unexpected combination of high compressive loading and rapid growth in this suture. PMID- 10521879 TI - The capacity to make decisions in dementia: some contemporary issues. PMID- 10521880 TI - Depressive symptoms, cognitive impairment and functional impairment in a rural elderly population in India: a Hindi version of the geriatric depression scale (GDS-H). AB - OBJECTIVE: To measure depressive symptomatology in a largely illiterate elderly population in India, using a new Hindi version of the Geriatric Depression Scale (GDS-H), and to examine its distribution and associations with age, gender, literacy, cognitive impairment and functional impairment. DESIGN: A Hindi version of the Geriatric Depression Scale was developed and administered to participants along with measures of demographic characteristics, cognitive functioning and functional ability. SETTING: The rural community of Ballabgarh in northern India. PARTICIPANTS: A community sample of 1554 mostly illiterate Hindi-speaking residents of Ballabgarh aged 55+. MEASURES: The Hindi version of the Geriatric Depression Scale (GDS-H); the Hindi Mental State Exam (HMSE); the Everyday Abilities Scale for India (EASI); age, gender and literacy. RESULTS: The GDS-H had high internal consistency and a factor structure comparable to the original English language version. The overall distribution of scores was higher than reported from other populations. Greater numbers of depressive symptoms, as measured by higher scores on the GDS-H, were associated with older age and illiteracy. Among the illiterate, there was no gender difference while among the literate, higher GDS-H scores were found among women. Cognitive impairment and functional disability were independently associated with higher scores on the GDS H after adjustment for age, gender and literacy. CONCLUSION: A reliable and valid Hindi version of the GDS has been developed. Depressive symptoms as measured by the GDS-H were prominent in this elderly illiterate northern Indian population and strongly associated with both cognitive and functional impairment. PMID- 10521881 TI - A meta-analysis of epidemiological studies in depression of older people in the People's Republic of China. AB - BACKGROUND: There has been little information about depression in Chinese elderly people. In order to investigate whether or not there is an excess of depression among the Chinese elderly, we have performed a meta-analysis of the published epidemiological studies. METHODS: Papers published in the literature from The People's Republic of China included in the Chinese medical databases were obtained. Some additional papers collected from other sources were also included. The fixed/random effects model and Poisson model were employed for data analysis. RESULTS: There were 10 cross-sectional studies (23 samples divided according to men/women and urban/rural subjects) giving sufficient prevalence data on depression (13 565 subjects) or depressive mood (8476 subjects). The pooled prevalence of depression was 3.86% (95%CI 3.37-4.42%), while that of depressive mood was 14.81% (14.20-15.64%). The risk of depression in the rural communities (5.07%, 3.61-7.13%) was higher than in the urban (2.61%, 2.22-3.08%). The same trends were observed for depressive mood. The patterns of risk factors were similar to those in western countries. CONCLUSIONS: Chinese tradition and culture may be explanatory factors for the low prevalence, provided the methodological issues have not seriously biased the results. PMID- 10521882 TI - Longitudinal predictors of non-aggressive agitated behaviors in the elderly. AB - Longitudinal predictors of physically and verbally non-aggressive inappropriate behaviors were examined in 200 community-dwelling elderly persons attending senior day care centers. Models based on ratings obtained from staff members and family caregivers were compared. Multiple factors contributed simultaneously to the prediction of non-aggressive behaviors. Similar to previous cross-sectional results, physically non-aggressive behaviors were predicted mainly by good health and cognitive impairment. In addition, depression emerged consistently as a predictor of physically non-aggressive behaviors in all models. Verbally non aggressive behaviors were predicted by depressed affect and pain, confirming previous suggestions that these behaviors are related to discomfort. The relationship of these behaviors with cognitive functioning was relatively weak. Understanding the etiologies of non-aggressive problem behaviors can aid in developing appropriate care for this population. PMID- 10521883 TI - Cross-cultural differences in demented geropsychiatric inpatients with behavioral disturbances. AB - OBJECTIVE: Cross-cultural differences in treatment and diagnosis exist in several psychiatric disorders. This study examines phenomenological and treatment differences between Caucasian and African-American patients presenting to a geropsychiatric unit for treatment of behavioral disturbances associated with dementia. METHODS: One hundred and forty-one Caucasian patients were compared to 56 African-American patients consecutively admitted to a VA geropsychiatric inpatient unit. At admission, differences in behavior disturbances between the two groups were examined using the Mini-Mental State Examination (MMSE), Cohen Mansfield Agitation Inventory (CMAI), Hamilton Rating Scale for Depression (HAM D), Brief Psychiatric Rating Scale (BPRS) and the Positive and Negative Syndrome Scale for Schizophrenia (PANSS). Differences in treatment were assessed by comparing medication types and doses between the two groups. RESULTS AND CONCLUSION: Results showed that Caucasian and African-American patients with dementia and behavioral disturbances presented and responded similarly to like treatment on an inpatient geropsychiatric unit. The similarity between the two groups may be explained by the multi-ethnic make-up of the interdisciplinary treatment team and by the use of standardized scales to measure symptomatology and response. PMID- 10521884 TI - The use of non-prescription sleep products in the elderly. AB - While sleep disorders are common in the elderly, the use of non-prescription products for sleep in this population has not been fully evaluated. The objectives of this project were to assess the use, perceived effectiveness and toxicity of non-prescription sleep products in an ambulatory elderly population. METHODS: A self-administered 20-question survey was distributed to seniors, aged 60 years or more, during hospital or pharmacy visits. RESULTS: Of the total respondents (N=176, mean age 74+/-7 years, 59% female), 84 (48%) indicated that they had used one or more therapies for sleep within the past year. These included non-prescription products (50% of therapies), prescription products (17%) and non-drug activities such as walking or drinking milk (34%). For those individuals who had used a non-prescription product in the past year (N=47, 27% of total respondents), the most frequently used products were: dimenhydrinate (21%), acetaminophen (19%), diphenhydramine (15%), alcohol (13%) and herbal products (11%). Most took them at least 1 day per week (79%) and 32% took them daily. These products subjectively improved sleep latency (mean 32 vs 61 minutes, p<0.001), number of nocturnal awakenings (mean 2 vs 3 awakenings, p<0.001) and total hours of sleep (mean 6.6 vs 5.4 hours, p<0.001). Mild side-effects were reported by 35 respondents (75%), the most common being dry mouth (N=22) and daytime drowsiness (N=13). Respondents were taking an average of four (SD+/-3, range 0-10) other medications currently. CONCLUSIONS: Non-prescription products are widely used by this population of ambulatory elderly for sleep disturbances. Most of the products were not marketed for sleep; however, they were perceived to be efficacious with low toxicity. The potential for drug interaction is high. Further research is warranted to evaluate the safety and effectiveness of non prescription sleep products in the elderly. PMID- 10521885 TI - Short versions of the geriatric depression scale: a study of their validity for the diagnosis of a major depressive episode according to ICD-10 and DSM-IV. AB - OBJECTIVE: To determine the validity of short Geriatric Depression Scale (GDS) versions for the detection of a major depressive episode according to ICD-10 criteria for research and DSM-IV. DESIGN: Cross-sectional evaluation of depressive symptoms in a sample of elderly subjects with short GDS versions. Different GDS cutoff points were used to estimate the sensitivity, specificity, positive predictive value and negative predictive value for the diagnosis of major depressive episode. Internal consistency of the scales was estimated with the Cronbach's alpha coefficient. SETTING: Mental Health Unit for the Elderly of 'Santa Casa' Medical School in Sao Paulo, Brazil. PARTICIPANTS: Sixty-four consecutive outpatients aged 60 or over who met criteria for depressive disorder (current or in remission). Subjects with severe sensory impairment, aphasia or Mini-Mental State score lower than 10 were excluded from the study. MEASUREMENTS: ICD-10 Checklist of Symptoms, GDS with 15, 10, 4 and 1 items, Montgomery-Asberg Depression Rating Scale (MADRS), ICD-10 diagnostic criteria for research and DSM IV diagnostic criteria. RESULTS: The use of the cutoff point 4/5 for the GDS-15 produced sensitivity and specificity rates of 92.7% and 65.2% respectively, and positive and negative predictive values of 82.6% and 83.3% respectively when ICD 10 diagnostic criteria for major depressive episode were used as the 'gold standard'. Similarly, rates of 97.0%, 54.8%, 69.6% and 94.4% were found when DSM IV was the comparing diagnostic criteria. Sensitivity, specificity and positive and negative predictive values for the cutoff point 6/7 were 80.5%, 78. 3%, 86.8% and 69.2% according to ICD-10, and 84.8%, 67.7%, 73.7% and 80.8% respectively according to DSM-IV. Intermediate values were found for the cutoff point 5/6. The best fit for GDS-10 was the cutoff point 4/5, which produced a sensitivity rate of 80.5%, specificity of 78.3%, positive predictive value of 86.8% and negative predictive value of 60.2% according to ICD-10 diagnosis of a major depressive episode. Similarly, rates of 84.8%, 67.7%, 73.7% and 80.8% were found when DSM-IV criteria for major depression were used. GDS-4 cutoff point of 2/3 was associated with a sensitivity rate of 80.5%, specificity of 78.3%, positive predictive value of 86. 8% and negative predictive value of 69.2% when compared to ICD-10. Again, rates of 84.8%, 67.7%, 73.7% and 80.8% respectively were found when the criteria used were based on DSM-IV. GDS-1 had low sensitivity (61.0% and 63.6% for ICD-10 and DSM-IV respectively) and negative predictive value (56.7% and 67.6% for ICD 10 and DSM-IV respectively), suggesting that this question is of limited clinical utility in screening for depression. GDS-15 (rho=0.82), GDS-10 (rho=0.82) and GDS 4 (rho=0.81) scores were highly correlated with subjects' scores on the MADRS. Reliability coefficients were 0.81 for GDS-15, 0.75 for GDS-10 and 0.41 for GDS 4. CONCLUSION: GDS-15, GDS-10 and GDS-4 are good screening instruments for major depression as defined by both the ICD-10 and DSM-IV. The shorter four- and one item versions are of limited clinical value due to low reliability and failure to monitor the severity of the depressive episode. General practitioners may benefit from the systematic use of short GDS versions to increase detection rates of depression among the elderly. (c) 1999 John Wiley & Sons, Ltd. PMID- 10521886 TI - Mental symptoms in Parkinson's disease are important contributors to caregiver distress. AB - OBJECTIVE: To determine the emotional and social distress of caring for a patient with Parkinson's disease and to explore the impact of motor and mental symptoms in subjects with Parkinson's disease on their caregivers' situation. DESIGN: Cross-sectional, population-based study using self-report questionnaires to measure caregiver distress and rating scales to assess patient symptomatology. SETTING: Neurology and old age psychiatry services in Stavanger, Norway. SUBJECTS: Caregivers of 94 home-dwelling patients with Parkinson's disease. Two control groups (patients with diabetes mellitus and healthy elderly). OUTCOME MEASURES: Measures of social and emotional distress in caregivers, including the Relative Stress Scale, Beck Depression Inventory and the General Health Questionnaire. RESULTS: Caregivers, in particular spouses, had more severe depression and a higher proportion reporting tiredness, sadness and less satisfaction with life compared with healthy elderly subjects. Using linear regression analysis, patient predictors of caregiver distress were depression, functional and cognitive impairment, agitation, aberrant motor behaviour and delusions. CONCLUSIONS: Caring for a spouse with Parkinson's disease is associated with emotional and social distress, underlining the importance of also assessing the needs of carers. Mental symptoms of parkinsonian patients were the most consistent and powerful predictors of caregiver distress, suggesting that identification and treatment of mental symptoms may reduce distress in caregivers of subjects with Parkinson's disease. PMID- 10521887 TI - Depressogenic medication as an aetiological factor in major depression: an analysis in a clinical population of depressed elderly people. AB - OBJECTIVE: To study the role of depressogenic medication in the aetiology of major depression in the elderly. BACKGROUND: Depression can be caused, provoked or sustained by drugs prescribed for other reasons. The evidence for this statement is based on case-reports, not on investigations in relevant populations. METHOD: In the geriatric wards of three Dutch psychiatric hospitals, 195 patients with a DSM-III-R diagnosis of major depression (MDD) were studied. In the first week after admission the following data were recorded: age, gender, personal psychiatric history, family psychiatric history, Montgomery-Asberg Depression Rating Scale, Mini-Mental State Examination, history of stroke, use of medication and number of different medications used. Subjects using depressogenic medication were contrasted with subjects not using depressogenic medication on all variables. RESULTS: There was a significant negative relationship, adjusted for the other variables, between the use of depressogenic medication and a previous admission for depression. No other significant relationships between the use of depressogenic medication and aetiological variables were found. Patients with a first-time admission for MDD use depressogenic medication 2.44 times more often than patients with previous admissions for depression. CONCLUSION: The use of depressogenic medication is an independent and clinically relevant aetiological factor in MDD. PMID- 10521888 TI - The cognitive abilities screening instrument (CASI): data from a cohort of 2524 cognitively intact elderly. AB - OBJECTIVES: To describe the effects of age and education for the Cognitive Abilities Screening Instrument (CASI), a 25-item test of cognitive function. DESIGN: Cross-sectional descriptive study of the initial enrollment in a community-based prospective cohort study. PARTICIPANTS: A total of 2524 cognitively intact older adults over age 65 who were members of a major health maintenance organization, and who consented to participate in a longitudinal study. MEASUREMENTS: Summary scores for the CASI are given in the form of mean, median and percentile distributions specific for age and educational level. RESULTS: Based upon maximum likelihood analyses, age and education were significant (p<0.0001) predictors of total CASI score. Increased age and lower education were associated with a lower CASI score, as well as an increased spread in score distribution. Gender was also significantly related (p<0.01) to total CASI, with women having a slightly higher distribution of scores. Mean total scores ranged from CASI=82.2 (SD=9.0) in subjects aged 90-95 who had less than a high school degree to CASI=94.8 (SD=3. 8) in subjects aged 65-69 with at least a high school education. CONCLUSIONS: Like most cognitive screening instruments, performance on the CASI in non-demented persons is influenced by age and education. The reference values for 5-year age categories described in this article should be useful for clinicians and research investigators when using the CASI as a measure of cognitive function. PMID- 10521889 TI - Violent crime in an elderly demented patient. AB - I present here a case of an elderly, severely demented gentleman who is being charged with the violent murder of his wife. These patients, although uncommon, represent a sensitive and difficult area of management for the old age psychiatrist. I examine what literature there is available in this area and draw the conclusion that it is an important and understudied area of psychiatry. PMID- 10521890 TI - Problems in cross-cultural psychogeriatric research: contacting older Turkish immigrants in the UK. PMID- 10521891 TI - Capgras syndrome and animals. PMID- 10521892 TI - Announcement PMID- 10521893 TI - Announcement PMID- 10521894 TI - Calcified aortic valves. A warning sign. PMID- 10521895 TI - Statins and C-reactive protein. New connection. PMID- 10521897 TI - Heart lines. Winter: the dangerous season for heart failure. PMID- 10521896 TI - Heart lines. Flexible arteries and folic acid. PMID- 10521898 TI - Heart lines. Uric acid: an innocent bystander. PMID- 10521899 TI - Ask the doctor. I tried Viagra for impotence and it didn't work. Is there anything else that I can try? PMID- 10521901 TI - Flu treatment. A new member of the armamentarium. PMID- 10521900 TI - Colon cancer prevention. Worth the trouble. PMID- 10521902 TI - Eyesight. Living with low vision. PMID- 10521903 TI - Acupuncture. Pains and needles. PMID- 10521904 TI - Cholesterol tests. The good, the bad, and what's healthy. PMID- 10521906 TI - The whole is greater than the sum of the parts. I. Whole grain. PMID- 10521905 TI - Puzzling through a case. Life sentence gets commuted. PMID- 10521907 TI - General review: cocaine abuse and addiction - Part I. PMID- 10521908 TI - Insights: ambiguous loss: living with frozen grief. PMID- 10521909 TI - THC for Tourette's syndrome. PMID- 10521910 TI - Depression as a female illness. PMID- 10521912 TI - Coffee and cholesterol. PMID- 10521911 TI - Forum: what are the best ways to treat sexual problems caused by SSRIs? PMID- 10521913 TI - Treating prostate cancer. Part IV: radiotherapy. PMID- 10521914 TI - Depression and coronary artery disease: a heartbreak for men. PMID- 10521915 TI - Preventing "unpreventable" cancers. PMID- 10521916 TI - Walking. The ideal exercise? PMID- 10521917 TI - Hormones. Phytoestrogens. PMID- 10521918 TI - Health care. What blood tests tell Us. PMID- 10521919 TI - Was that a heart attack? PMID- 10521920 TI - By the way, doctor. I'm 47 and my menstrual periods are quite irregular, so my doctor started me on birth control pills. How will I know when I enter menopause and when to switch to HRT? PMID- 10521922 TI - Apoptosis and appearance of Trp53-positive micronuclei in murine tumors with different radioresponses in vivo. AB - The purpose of this paper is to determine the relationship between the response to radiation and the appearance of apoptosis and micronuclei with Trp53 protein in murine tumors after irradiation. Two murine tumors, EL4, which was derived from a mouse lymphoma, and FM3A, which was derived from a mouse mammary carcinoma, were locally irradiated with 15 Gy and sections were stained with H&E and an anti-Trp53 antibody. The response to radiation was greater in EL4 tumors than in FM3A tumors. The frequency of apoptotic cells in EL4 tumors was 6.1 +/- 1.2% at time zero, reached a peak of 36.3 +/- 3. 8% at 6 h, and then decreased with time through 72 h to 2.5 +/- 1.5% after 15 Gy irradiation. In FM3A tumors, no apoptotic cells were detected at 0, 1, 3, 6 or 24 h after exposure. At 48 and 72 h, the frequency was only 3.0 +/- 0.6% and 1.3 +/- 0.3%. Apoptotic cells increased significantly at 3, 6 and 24 h after irradiation in EL4 tumors (P < 0.008) and at 48 and 72 h in FM3A tumors (P < 0.006). The frequency of Trp53 positive cells was 17.9 +/- 2.2 and 15.2 +/- 2.3% at time zero in EL4 and FM3A tumors, respectively, increased to 74.5 +/- 4.5% in EL4 cells (P = 0.001), and increased to 33.9 +/- 1. 1% in FM3A cells (P = 0.005) 1 h after irradiation. Trp53-positive micronuclei appeared in cells in both tumors from 24 to 72 h after irradiation. The frequency of Trp53-positive micronuclei was 3.8 +/- 0.5 and 13.5 +/- 1.3% at 24 h in EL4 and FM3A tumors, respectively, and gradually decreased by 72 h. After exposure to 15 Gy, Trp53-positive micronuclei increased significantly in FM3A tumors compared to EL4 tumors at both 24 and 48 h (P < 0.02). The frequency of these micronuclei increased with increasing dose in FM3A tumors, and the difference between these percentages after 3 Gy and after 5, 10 and 15 Gy was significant (P < 0.02). Many apoptotic cells were observed in the radiosensitive EL4 tumor after irradiation. Death by apoptosis may be related to an early response to radiation in these tumors. The appearance of micronuclei may be an important mechanism of cell death in FM3A tumors in which no apoptosis was induced. PMID- 10521921 TI - A novel human stress response-related gene with a potential role in induced radioresistance. AB - We have isolated a novel gene, DIR1, from L132 cells that is transiently repressed after exposure to low radiation doses and has a potential role in induced radioresistance. Molecular and cellular characterization of this gene reveals that it is unique but has similarities to a family of heat-shock-related proteins known as immunophilins. These have been implicated in various cellular functions including general stress responses and control of the cell cycle. Antisense strategies have demonstrated that the DIR1 gene also appears to have some involvement in the control of the cell cycle. Furthermore, there appears be a potential role for this gene product in the phenomenon of induced radioresistance through a mechanism that increases the rate of DNA repair in cells exposed to X rays and subsequently increases the cells' resistance to radiation. This is the first description of an immunophilin-like gene that has a possible role in adaptive/inducible responses to X rays in mammalian cells. PMID- 10521923 TI - Ascorbic acid inhibits apoptosis induced by X irradiation in HL60 myeloid leukemia cells. AB - Exposure of cells to ionizing radiation can cause apoptosis. Since antioxidants have been shown to protect against radiation-induced apoptosis, in this study we have evaluated the putative protective effect of ascorbate against radiation induced apoptosis as well as the production of peroxides in the cells. HL60 cells transport the oxidized form of ascorbic acid, dehydroascorbic acid (DHA), and accumulate reduced ascorbate. Exposure of the cells to 5-40 Gy X radiation resulted in induction of apoptosis. Preincubation of the cells with DHA reduced the level of apoptosis after exposure to 5-20 Gy. Exposure of the cells to 5 or 20 Gy X radiation did not affect the intracellular concentration of peroxides, while phorbol myristate acetate (PMA), which is known to induce production of H(2)O(2) in cells (and served as a control), resulted in an increase in peroxides and a decrease in intracellular ascorbate. Irradiation of the cells with 1-3 Gy resulted in up-regulation of expression of BCL2 without affecting the level of apoptosis. At higher doses of radiation, enhanced BCL2 expression did not prevent radiation-induced apoptosis. Loading of the cells with ascorbate prior to their exposure to 1-3 Gy X radiation did not affect the enhanced BCL2 expression observed in the irradiated cells. At higher doses of radiation, ascorbate decreased apoptosis and restored the level of BCL2 in the cells. Exposure of the cells to 3-20 Gy X radiation enhanced the cell surface expression of TNFRSF6 (formerly known as Fas/APO-1) antigen and enhanced anti-TNFRSF6 antibody-induced apoptosis of the cells. Ascorbate loading did not affect expression of TNFRSF6 and did not overcome the anti-TNFRSF6 antibody-induced apoptosis. In conclusion, our data demonstrate that exposure of HL60 cells to radiation enhanced BCL2 and TNFRSF6 expression. Ascorbate did not affect BCL2 or TNFRSF6 expression. We therefore conclude that it protects HL60 cells against radiation-induced apoptosis, although the mechanisms of protection must still be elucidated. PMID- 10521924 TI - The vitamin D3 analog EB 1089 enhances the response of human breast tumor cells to radiation. AB - Previous studies from this laboratory as well as others have demonstrated that breast tumor cells fail to undergo primary apoptosis in response to agents which induce DNA damage such as ionizing radiation and the topoisomerase II inhibitor adriamycin. Similarly, the primary response of breast tumor cells to vitamin D(3) [1,25-(OH)(2)-D(3)] and its analogs such as EB 1089 is growth inhibition, with apoptosis occurring in only a small fraction of the cell population. The possibility that the combination of vitamin D(3) compounds with radiation might promote cell death (i.e. through a differentiation stimulus plus DNA damage) was investigated by exposing both TP53 (formerly known as p53) wild-type and TP53 mutated breast tumor cells to 1,25-(OH)(2)-D(3) or EB 1089 for 48 h prior to irradiation. This combination resulted in enhanced antiproliferative effects in the TP53 wild-type MCF-7 cells based on both a clonogenic assay and the determination of numbers of viable cells. The combination of EB 1089 with radiation increased DNA fragmentation based on both the terminal transferase end labeling (TUNEL) and bisbenzamide spectrofluorometric assays, suggesting the promotion of apoptosis. The observed increase in DNA fragmentation was not due to an enhancement of the extent of initial DNA damage induced by radiation. These findings suggest that vitamin D compounds may be useful in combination with radiation in the treatment of breast cancer. PMID- 10521925 TI - Regulation of transforming growth factor beta1 by radiation in cells of two human breast cancer cell lines. AB - We have investigated the mechanisms by which radiation inhibits proliferation of human breast cancer cells in culture. Radiation, within the dose range used for treatment of humans, decreased the rate of proliferation of estrogen-independent MDA-MB-231 cells more effectively than it did that of estrogen-dependent MCF-7 cells. The rate of proliferation of MDA-MB-231 cells was also inhibited to a greater extent than that of MCF-7 cells by purified TGFB1. Using an ELISA specific for activated TGFB1, we found that conditioned medium from irradiated MDA-MB-231 or MCF-7 cells contained twofold more TGFB1 than that from nonirradiated cells. Conditioned medium from irradiated breast cancer cells, but not from nonirradiated cells, inhibited the growth of untreated MDA-MB-231 cells. The inhibitory activity was blocked by an anti-TGFB1 neutralizing antibody. An approximately twofold increase in the TGFB1 mRNA in irradiated cells compared to controls was found using semiquantitative reverse-transcriptase PCR. Last, the mRNA for insulin-like growth factor binding protein 3, a reported target of the cell inhibitory activity of TGFB1, was increased threefold upon irradiation. Our results demonstrate that the TGFB1 is increased after irradiation and that the activation of the TGFB1 signaling pathway may sensitize cells to the effects of radiation. PMID- 10521926 TI - Sensitizing human cervical cancer cells In vitro to ionizing radiation with interferon beta or gamma. AB - Human cervical cancer is often associated with human papilloma virus (HPV). HPV products, such as the oncoproteins E6 and E7, are known to disrupt the function of TP53 (formerly known as p53). The protein encoded by the TP53 gene plays a central role in managing cellular damage. Interferons are known to down-regulate E6/E7 and may therefore restore TP53 function and influence radiation sensitivity. We investigated whether IFNB or IFNG, at various concentrations (2- 300 IU/ml) and for a range of durations of exposure (from 48 h before to 8 h after irradiation), were able to modify the radiation response of HeLa, C4-1, Me 180, C33-A and SiHa cells. In parallel to the clonogenic assays, we analyzed the effect on the mRNA that encodes IFNB and E6 by Northern blotting in the same experimental conditions. A significant change in the initial slope of the dose response curve was observed more consistently with IFNB than with IFNG. No changes in the mRNA or protein level of TP53 and E6 could be detected. Thus other mechanisms of action need to be investigated to explain radiosensitization with recombinant IFNB in cells of human cervical cancer cell lines. PMID- 10521927 TI - Combined radiation and enzyme/prodrug treatment for head and neck cancer in an orthotopic animal model. AB - In an effort to improve the therapeutic outcome for squamous cell cancer of the head and neck, we have used the enzyme cytosine deaminase (CD) and the prodrug 5 fluorocytosine (5-FC) as a means to deliver the chemotherapeutic agent 5 fluorouracil (5-FU) in a tumor-specific manner and have evaluated the use of this treatment in combination with external-beam radiation. Infection of SCCVII cells in culture with a CD-expressing retrovirus and treatment with 5-FC was cytotoxic depending on the time of treatment and dose of 5-FC. An orthotopic model of squamous cell cancer of the head and neck was used in vivo to study the CD/5-FC system both alone and with concurrent radiation due to the radiosensitizing properties that 5-FU generates in situ. Treated mice were imaged using magnetic resonance imaging (MRI), and their survival was evaluated. Neither 5-FU nor radiation either alone or combined provided a survival advantage. In contrast, 5 FC treatment prolonged survival and decreased tumor burden compared to control animals, but the tumors recurred after the treatment ceased. Finally, combined treatment with concurrent administration of 5-FC and radiation resulted in a synergistic decrease in tumor growth and enhanced survival over treatment with 5 FC or radiation alone. PMID- 10521928 TI - Intact alveolar epithelial permeability and transalveolar fluid absorption after thoracic irradiation in rats. AB - We have addressed the question of how the alveolar space stays relatively free of fluid when thoracic irradiation injures the pulmonary capillary endothelium and plasma fluid leaks into the interstitium. A single dose of 15 Gy to the thorax of rats significantly increased the pulmonary capillary filtration coefficient and the lung wet/dry weight ratio 2 h after irradiation. However, there was no significant increase in the release of lactose dehydrogenase or leaking of Evans blue dye into the alveolar space, indicating that alveolar epithelial permeability remained intact. We found no significant difference in the basal alveolar fluid clearance between control and irradiated animals. There was also no significant difference in blockage of alveolar fluid clearance by amiloride. This indicates that the function of the alveolar epithelial Na(+) channels is not impaired and that alveolar epithelium absorbs fluid normally. Examination of lung tissue by light microscopy demonstrated accumulation of fluid in the perivascular region but not in the alveolar space. Our data appear to indicate that the alveolar epithelial barrier function is more resistant to radiation than that of the pulmonary capillary endothelium. We conclude that intact alveolar epithelial permeability and normal transalveolar epithelial fluid absorption ability are of critical importance in keeping the alveolar space relatively free of fluid during acute radiation lung injury. PMID- 10521929 TI - Metabolic correction of cerebral radiation syndrome. AB - DNA strand breaks that occur after irradiation activate the repair enzyme adenosine diphosphoribosyl transferase, which consumes NAD as a substrate and causes depletion first of neuronal NAD and then of the ATP pool. This is considered to be the crucial link in the mechanism underlying the cerebral radiation syndrome (CRS). In this study, two ways to correct the CRS metabolically were examined: (a) prevention of depletion of NAD after irradiation by administration of the enzyme inhibitor nicotinamide and (b) shunting the NAD dependent oxidative phosphorylation pathway of ATP resynthesis by administration of a substrate of NAD-independent oxidation, succinate. Cerebral lesions induced by radiation were modeled by irradiation of rats or rat brain homogenates with 150 Gy of X rays. The manifestations of CRS in rats (excitement, convulsions, etc.) closely resembled those seen after acute hypoxia. In brain homogenates, pyruvate tetrazolium-reductase activity decreased after irradiation and could be corrected by addition of NAD after irradiation. Succinate tetrazolium-reductase activity was not affected by irradiation. Oxygen consumption by brain homogenates after irradiation in vitro and in situ decreased, as did oxygen consumption by rats in vivo after cranio-caudal irradiation. Administration of nicotinamide or succinate prevented both the postirradiation decrease in respiration (in both rats in vivo and brain homogenates in vitro) and the development of cerebral radiation syndrome. These results help to clarify the mechanisms underlying CRS and its metabolic correction. PMID- 10521930 TI - Strand breaks in plasmid DNA after positional changes of Auger electron-emitting iodine-125: direct compared to indirect effects. AB - To elucidate the nature and kinetics of DNA strand breaks caused by low-energy Auger electron emitters, we compared the yields of DNA breaks in supercoiled pUC19 DNA in the presence of the (.)OH scavenger dimethyl sulfoxide (DMSO) after the decay of (125)I (1) in proximity to DNA after minor-groove binding ((125)I iodoHoechst 33342, (125)IH) and (2) at a distance from DNA ((125)I iodoantipyrine, (125)IAP). DMSO is efficient at protecting supercoiled plasmid DNA from the decay of (125)I free in solution (dose modification factor, DMF = 59 +/- 4) and less effective when the (125)I decays occur close to DNA (DMF = 3.8 +/ 0.3). This difference is due mainly to the inability of DMSO to protect DNA from the double-strand breaks produced by groove-bound (125)I (DMF = 1.0 +/- 0.2). Additionally, the fragmentation of plasmid DNA beyond the production of single strand and double-strand breaks that is seen after the decay of (125)IH and not (125)IAP (Kassis et al., Radiat. Res. 151, 167-176, 1999) cannot be modified by DMSO. These results demonstrate that the mechanisms underlying double-strand breaks caused by the decay of (125)IH differ in nature from those caused by the decay of (125)IAP. PMID- 10521931 TI - Decreased proportion of CD4 T cells in the blood of atomic bomb survivors with myocardial infarction. AB - Epidemiological studies of the atomic bomb survivors have suggested dose-related increases in mortality from diseases other than cancer. Cardiovascular disease is one such noncancer disease for which increases in both mortality and incidence have been found to be associated with radiation dose. Immunological studies have revealed long-term impairment of T-cell-mediated immunity, especially involving deficiencies of CD4 helper T cells, in atomic bomb survivors. In the present study, we investigated whether decreases in CD4 T cells were associated with myocardial infarction in atomic bomb survivors. Of 1,006 survivors examined to determine the proportion of CD4 T cells in peripheral blood lymphocytes, 18 persons had a history of myocardial infarction. The proportion of CD4 T cells was significantly decreased with increased radiation dose [corrected]. Further, the prevalence of myocardial infarction was significantly greater in individuals with a lower proportion of CD4 T cells. These results suggest that myocardial infarction in atomic bomb survivors may be associated with defects in CD4 helper T cells. PMID- 10521932 TI - Gap junction intercellular communication mediates the competitive cell proliferation disadvantage of irradiated mouse preimplantation embryos in aggregation chimeras. AB - Gap junction intercellular communication (GJIC) is thought to play a role in the growth modulation that occurs within cell populations. An example of heterologous growth inhibition (competitive cell proliferation disadvantage) occurs within mouse aggregation chimeras comprised of irradiated and nonirradiated cleavage stage embryos. The goal of this investigation was to test the hypothesis that GJIC participates in the competitive cell proliferation disadvantage that is expressed by the irradiated embryo in aggregation chimeras. Specifically, we tested the capacity of the GJIC inhibitor 18 alpha-glycyrrhetinic acid (AGA) to inhibit competitive cell proliferation disadvantage in heterologous aggregation chimeras that were comprised of one embryo that was irradiated with 1.0 Gy of (137)Cs gamma rays and then paired with one nonirradiated embryo. We found that AGA successfully inhibited fluorescent dye transfer between irradiated and nonirradiated embryos in heterologous chimeras. Chronic exposure to AGA prevented competitive cell proliferation disadvantage in these radiation chimeras, while exposure to AGA for the first 15 h of culture (prior to gap junction development) did not prevent competitive cell proliferation disadvantage. An unexpected observation was the apparent lack of any effect of inhibiting GJIC by exposure to AGA on blastocyst formation and cell number allocation in the two principal stem cell lineages of the preimplantation mammalian embryo, trophectoderm and inner cell mass. PMID- 10521933 TI - Unexpected sensitivity to the induction of mutations by very low doses of alpha particle radiation: evidence for a bystander effect. AB - We examined the induction of HPRT mutations in CHO cells exposed to low fluences of (238)Pu alpha particles from a specially constructed irradiator. The dose response relationship was linear over the dose range of 5 cGy-1.2 Gy. However, unexpected sensitivity, leading to a significantly higher frequency of mutations than would be predicted by a back extrapolation from the data for higher doses, was observed in the dose range below 5 cGy, where the mean number of alpha particle traversals per nucleus was significantly less than one (0.05-0.3). The frequency of mutations induced by a single alpha particle traversing the nucleus of a cell was increased nearly fivefold at the lowest fluence studied. The data are consistent with the conclusion that the enhanced efficiency of each nuclear traversal at low particle fluences is the result of mutations arising in nonirradiated, bystander cells. PMID- 10521935 TI - Does the limiting F value at very low doses depend systematically on linear energy transfer? AB - The debate on the validity of the ratios of radiation-induced yields of chromosome aberrations, in particular the F value (dicentrics/ring chromosomes), as a chromosomal fingerprint of radiation quality is still in progress. From a recent analysis of their experimental data, Sasaki et al. (Radiat. Res. 150, 253 258, 1998) noted that despite a considerable variability in the data, the limiting F value at the lowest doses, or the F(0) value, obviously decreased with increasing LET, indicating that the LET could be a factor that determines the F value. We have reassessed here our own 13 cytogenetic data sets that cover a range of dose-averaged LET of 0.5 to 150 keV/microm in terms of this F(0)-value approach, but we could not confirm such a dependence on LET at very low doses. The validity of the F value as a biomarker therefore remains questionable. For a final evaluation, scoring of a far greater number of cells at low doses would be necessary to reduce the large error ranges of F values. PMID- 10521934 TI - The F value for chromosome aberrations in atomic bomb survivors does not provide evidence for a primary contribution of neutrons to the dose in Hiroshima. AB - Brenner and Sachs (Radiat. Res. 140, 134-142, 1994) proposed that the ratio of interchromosomal to intrachromosomal exchanges, termed the F value, can be a cytogenetic fingerprint of exposure to radiations of different linear energy transfer (LET). Using published data, they suggested that F values are over 10 for low-LET radiations and approximately 6 for high-LET radiations. Subsequently, as F values for atomic bomb survivors were reported to be around 6, Brenner suggested that the biological effects of atomic bomb radiation in Hiroshima are due primarily to neutrons. However, the F values used for the survivors were means from individuals exposed to various doses. As the F-value hypothesis predicts a radiation fingerprint at low doses, we analyzed our own data for the survivors in relation to dose. G-banding data for the survivors showed F values varying from 5 to 8 at DS86 doses of 0.2 to 5 Gy in Hiroshima and around 6 in Nagasaki with no evidence of a difference between the two cities. The results are consistent with our in vitro data that the F values are invariably around 6 for X and gamma rays at doses of 0.5 to 2 Gy as well as two types of fission-spectrum neutrons at doses of about 0.2 to 1 Gy. Thus, apart from a possible effect at even lower doses, current data do not provide evidence to support the proposition that the biological effects of atomic bomb radiation in Hiroshima are caused mainly by neutrons. PMID- 10521936 TI - Heart lines. Angioplasty versus clot busters for older heart-attack patients. PMID- 10521937 TI - Heart lines. The third blood pressure number. PMID- 10521938 TI - The whole is greater than the sum of the parts. II. Beta carotene. PMID- 10521939 TI - Treatment for misbehaving children. PMID- 10521940 TI - Birth complications and schizophrenia. PMID- 10521941 TI - Aging and body composition. PMID- 10521942 TI - By the way, doctor. I take 10 mg of Prilosec about four times each week to control my heartburn. Does taking this drug on a continuing basis affect my body's ability to utilize nutrients in food? PMID- 10521943 TI - False-negative finding in rapid interphase FISH analysis of uncultured amniotic cells. PMID- 10521945 TI - Etiology and pathogenesis of preeclampsia: current concepts. PMID- 10521946 TI - Minilaparotomy hysterectomy-comments. PMID- 10521948 TI - Neurobiology of infants born to women who exercise regularly throughout pregnancy. PMID- 10521950 TI - The clinical significance of intermittent sinusoidal fetal heart rate. PMID- 10521951 TI - Ductus venosus blood flow evaluation: its importance in the screening of chromosomal abnormalities. PMID- 10521952 TI - Outcomes of aggressive treatment of stage IV gallbladder cancer and predictors of survival. AB - BACKGROUND/AIMS: Stage IV gallbladder carcinoma patients are rarely considered treatable by resection. They resign themselves to palliation because there is no long-term survival data available on the risks of morbidity and mortality following aggressive treatment. The aim of this study was to evaluate predictors of survival following aggressive resection surgery for stage IV gallbladder carcinoma. METHODOLOGY: In this retrospective study, we examined 93 patients with stage IV gallbladder carcinoma who had undergone resections. Of the 93 patients, 69 had undergone liver resection to various extents together with hepaticocholedochus resection (HCR); 2 had undergone pancreaticoduodenectomy (PD) both with and without HCR; 31 had undergone hepatopancreaticoduodenectomy (HPD); 7 had undergone cholecystectomy together with HCR; 12 had undergone cholecystectomy; and 3 had undergone extended cholecystectomy. Fifty of the 93 patients had also undergone adjuvant radiotherapy. Using univariate and multivariate analyses, 13 clinicopathologic risk factors were analyzed to predict survival. RESULTS: Operative morbidity and mortality rates were 17.2% and 5.4%, respectively. Overall, the 5-year survival rate and median survival time were 9.8% and 243 days, respectively. The 5-year survival rate was significantly higher in stage IVA (n = 17) than in stage IVB (n = 76), at 42.8% and 4.9%, respectively. Multivariate analysis revealed that sex, histopathologic type, lymph node involvement (N), subgroup of stage IV, post-resection residual tumors, and adjuvant radiotherapy were significant predictors of survival. CONCLUSIONS: Long-term survival, with acceptable mortality and morbidity, can be expected in female patients who have stage IVA gallbladder cancer consisting of well differentiated adenocarcinoma and who undergo either complete microscopic resection or grossly complete resection followed by adjuvant radiotherapy. PMID- 10521953 TI - Proposal on the extent of lymph node dissection for gallbladder carcinoma. AB - BACKGROUND/AIMS: To evaluate the value of performing extended regional lymph node dissection for gallbladder carcinoma, the mode of recurrence after curative resection was analyzed. METHODOLOGY: Records of 45 patients who underwent surgical resection for gallbladder carcinoma from 1973 to August 1997 were reviewed. RESULTS: Thirty-three cases underwent a curative resection and 12 received a non-curative resection. Among the 32 patients who survived the curative resection, cancer recurred in 7 with lymph node metastasis, whereas recurrence was found in only 1 of the remaining 25 patients without lymph node metastasis (p < 0.0001). At the 1st diagnosis of recurrence in these 8 patients, lymph node recurrence was detected in 7, and the site of recurrence was limited to the lymph nodes, which were confined to the peripancreatic region and the interaortocaval nodes near the left renal vein in 4 cases. CONCLUSIONS: In view of the site of the metastatic lymph nodes and the lymphatic drainage system of the gallbladder, it was considered that lymph node dissection was inadequate in 5 of the 8 patients and that 2 might have been cured by extended regional lymph node dissection, including complete resection of the retroportal, posterior pancreatoduodenal, right celiac and interaortocaval nodes. PMID- 10521954 TI - Does aggressive surgical resection improve the outcome in advanced gallbladder carcinoma? AB - BACKGROUND/AIMS: Patients with advanced gallbladder carcinoma have usually been considered nonresectable, leading to a very poor outcome. This study was aimed to evaluate the results of our aggressive surgical approaches in certain cases of advanced gallbladder carcinoma. METHODOLOGY: Ninety-one patients with advanced gallbladder carcinoma of stages pT3 and pT4 who underwent surgery at our institution were the subjects of this study. Fifty-eight of 91 patients had surgical excision; 44 by hepatic resection and 14 by hilar resection. Post operative outcome was evaluated. Advanced gallbladder carcinomas were classified according to our previously reported classification: type I hepatic; type II biliary; type III hepatobiliary; type IV others. RESULTS: Curative resection was obtained at a more increased rate in type I tumor patients than in types II and III (91% vs. 29%, p < 0.01). The surgical mortality rate was 17%. Survival rates of resected patients were significantly higher that those of nonresected patients: 45%, 31%, 22%, 17%, 13% at 1, 2, 3, 4, 5 years vs. 9%, 9%, 0% at 1, 2, 3 years (p < 0.01). Survival rates of type I tumor patients after curative resection were remarkably higher than those of type II and III tumor patients, (69%, 64%, 56%, 48%, 39% at 1, 2, 3, 4, 5 years vs. 17%, 17%, 0% at 1, 2, 3 years). CONCLUSIONS: Aggressive surgical approaches might bring about improved prognosis in advanced gallbladder carcinoma, especially for patients with type I tumors. PMID- 10521955 TI - Should the bile duct be preserved or removed in radical surgery for gallbladder cancer? AB - BACKGROUND/AIMS: It is unclear whether resection of the extrahepatic bile duct in radical surgery for gallbladder cancer should be performed when direct infiltration into the hepatoduodenal ligament is absent. METHODOLOGY: The results of radical surgery with or without bile duct resection were compared in 55 patients with gallbladder cancer without direct extension to the hepatoduodenal ligament. Lymph node dissection and combined resection of involved organs were carried out according to the extent of the tumor. RESULTS: Nodal involvement was present in 43% of patients with tumors more advanced than pT1. Survival rates were similar between patients with or without bile duct resection in stages I III, while significantly better survival was observed with bile duct resection in stage IV. CONCLUSIONS: Considering the adverse effect of bilioenteric anastomosis, preservation of the extrahepatic bile duct is recommended in radical surgery for gallbladder cancer when the tumor is less advanced than stage IV and does not extend to the hepatoduodenal ligament. PMID- 10521957 TI - Acute emphysematous cholecystitis. Report of twenty cases. AB - BACKGROUND/AIMS: Our aim is to present our experience with acute emphysematous cholecystitis (AEC), a severe variety of acute cholecystitis characterized by early gangrene and perforation of the gallbladder. METHODOLOGY: We reviewed the clinical records of 20 patients with AEC, analyzing age, sex, past medical history, symptoms, laboratory tests, X-rays, ultrasounds, operative and microbiological findings, morbidity and mortality. RESULTS: Our study included 13 men and 7 women (mean age 59 years). Associated factors were diabetes mellitus (11 cases) and gallstones (6 cases, 3 of them with common bile duct stones). Clinical symptom presentation included: right hypochondrial pain and fever in all cases, vomiting in 9, septic shock in 3, jaundice in 7, and peritonitis in 8. Hyperbilirubinemia was present in 7 cases. Plain abdominal X-rays or ultrasounds led to diagnosis in 95% of the cases. Surgical findings were AEC in all cases, pericholecystic abscess in 8, gallbladder necrosis in 7 and bile peritonitis in 3. C perfringens, E coli and B fragilis were the most frequent pathogens. Mortality rate was 25%, and morbidity 50%. CONCLUSIONS: AEC predominantly affects elderly diabetic men. Abdominal X-rays or ultrasounds are good diagnostic techniques, and emergency surgery is needed due to the high incidence of gangrene and perforation Despite all the efforts made, morbidity and mortality are still high. PMID- 10521956 TI - Outcome of extensive surgery for TNM stage IV carcinoma of the gallbladder. AB - BACKGROUND/AIMS: Whether extensive surgery improves the prognosis of patients with advanced gallbladder carcinoma remains unclear. Twenty-three patients with stage IV carcinoma underwent resection of tumors at our department over a period of 19 years. The outcome of extensive surgery was analyzed in this group. METHODOLOGY: Resection of segments 4a plus 5, extended right lobectomy or right trisegmentectomy, and hepatopancreatoduodenectomy (HPD) were performed in 5 patients, 7 patients, and 11 patients, respectively. Extrahepatic bile duct resection, regional lymph node dissection and paraaortic node dissection were performed in all patients. The survival rate after operation was compared to that of 39 patients with unresectable tumors. RESULTS: The morbidity rate was high (60.8%), but there were no post-operative deaths. The 1-, 3- and 5-year survival rates were 51%, 17%, and 11%, respectively. The longest survival time of stage IV carcinoma patients was 18 years. The 5-year survival rate of 6 patients undergoing curative resection was significantly better than that of 17 patients undergoing noncurative surgery (p < 0.05). The survival of 11 patients undergoing HPD and 7 patients with para-aortic nodal metastasis were both significantly improved compared to that of patients with unresectable tumors (p < 0.05). The quality of life and the out-of-hospital period were also better. CONCLUSIONS: Extensive surgery for stage IV gallbladder carcinoma has a good palliative effect. PMID- 10521958 TI - Effects of gallstone-promoting diet and vagotomy on the mouse gallbladder epithelium. AB - BACKGROUND/AIMS: An increased incidence in gallstone disease has been suggested in patients subjected to vagotomy, therefore we studied whether vagotomy influences the gallbladder epithelium and the secretion of glucoprotein granules in mice fed a gallstone-inducing diet. METHODOLOGY: Ten mice were given a lithogenic diet for 7 weeks. Five of them were subjected to left truncal vagotomy. As controls, 5 mice were sham-operated and 5 were subjected to vagotomy. Seven weeks after the operation, the morphological changes in the gallbladder epithelium were quantified with an electron microscopic morphometric method. RESULTS: In mice on a lithogenic diet, an increase in the volume density of the secretory granules and the profile area of the principal cells of the epithelium were found. The morphological changes were identical in animals subjected to truncal vagotomy and given a lithogenic diet. CONCLUSIONS: The effect of a gallstone-promoting diet in mice, seen as an increase in the volume density of the secretory granules and in the profile area of the principal cells of the gallbladder epithelium, was not influenced by deprivation of vagal innervation. PMID- 10521959 TI - Percutaneous transhepatic cholangioscopic lithotripsy and change of biliary manometry patterns. AB - BACKGROUND/AIMS: Percutaneous transhepatic cholangioscopic lithotripsy (PTCSL) is used to remove bile duct stones. This work aims to evaluate the clinical usefulness of PTCSL and the reversibility of the terminal bile duct dysfunctions after PTCSL. METHODOLOGY: Thirty patients who underwent PTCSL using mechanical and/or electrohydraulic lithotripsy over the past 10 years (20 patients with common bile duct stones and 10 with intrahepatic bile duct stones) were evaluated. Terminal bile ductal pressure was measured using the percutaneous transhepatic biliary drainage (PTBD) tube prior to and after lithotripsy by means of variable-load cholangiomanometry. RESULTS: Complete stone extraction was possible in 26 patients (86.7%). The other 4 patients had intrahepatic stones. Complications included 2 cases of hemobilia, one of pneumonia, and 3 of localized peritonitis. Of 26 patients without residual stones, only 4 patients had a linear pressure flow (P-F) pattern which indicates normal biliary tract function prior to lithotripsy. In 17 of 22 patients with other type P-F patterns, however, these types also changed to a linear pattern after complete removal of stones. The P-F pattern of the other 5 patients remained unchanged. CONCLUSIONS: PTCSL is a safe and efficient method treating biliary tract lesions while preserving the function of the sphincter of Oddi. The terminal biliary tract function normalized after stone removal. Thus, PTCSL was useful for patients with complicated bile duct stones not accessible to endoscopic retrograde management. PMID- 10521960 TI - Obstructive jaundice promotes bacterial translocation in humans. AB - BACKGROUND/AIMS: Significant bacterial translocation was demonstrated following experimental biliary obstruction, however very little is known about the importance and the prevalence of gut-origin sepsis in obstructive jaundice patients. Therefore, the aim of this study was to investigate the concept of gut origin sepsis in obstructive jaundiced patients and its clinical importance. METHODOLOGY: Twenty-one patients requiring laparotomy for obstructive jaundice (group I) and thirty patients operated on electively mainly for chronic cholecystitis (group II) were studied. Peritoneal swab, mesenteric lymph node, portal venous blood, liver wedge biopsy and bile were sampled for culture immediately after opening the peritoneum. Additionally, peripheral blood samples were taken pre- and post-operatively from all patients. Post-operatively, patients were monitored for infectious complications. RESULTS: The mean serum bilirubin concentration, gamma glutamyl transferase and alkaline phosphatase levels in jaundiced patients before therapeutic intervention were significantly higher than in control patients. Five patients demonstrated bacterial translocation in group I (24%), whereas only one did so in group II (3.5%, p < 0.05). Septic complications were detected in three patients, but only in two with bacterial translocation in group I. There was one patient with bacterial translocation who had septic complication in group II. CONCLUSIONS: The present study demonstrated that obstructive jaundice significantly promotes bacterial translocation in humans, however, its clinical importance has yet to be defined. PMID- 10521961 TI - The role of insulin and glucagon in experimental obstructive jaundice. AB - BACKGROUND/AIMS: The oxidative phosphorylation of liver mitochondria is regulated by the amount of portal insulin available to the hepatocytes. Thus, hepatic energy is mediated by the values of blood sugar and insulin. Insulin and glucagon are the main fuel homeostats in the liver. This study was performed to investigate the concept of energy mediated by glucose, during the process of obstructive jaundice and its recovery. METHODOLOGY: Experimental Wistar rats were studied, with bile duct tied for 4, 7 and 14 days respectively. The serum concentration and relative tissue concentration of insulin and glucagon were measured. And the common bile duct was tied for 4, 7 and 14 days, then relieved by time sequences for 4, 7 and 14 days. Serum concentration and relative tissue concentration of insulin and glucagon were also measured. RESULTS: When the common bile duct was tied for 4, 7, and 14 days respectively, the serum concentration and relative tissue concentration of insulin declined (p < 0.05) and glucagon concentration was elevated (p < 0.05). When the common bile duct was tied for 4, 7 and 14 days, then relieved by time sequences for 4, 7 and 14 days, the concentrations of insulin in both groups appeared to decline at first (p < 0.05) and then progressively increase (p < 0.05). The concentrations of glucagon exhibit the reverse behavior. Both serum and tissue concentration are elevated at first (p < 0.05), then progressively decline (p < 0.05). CONCLUSIONS: These studies indicated that, during obstructive jaundice, more fuel is demanded to make up for the energy deficiency. In spite of surgical or non-surgical relief of obstructive jaundice, the energy reserve is still not sufficiently recovered. The recovery of the hepatic energy reserve takes longer than we expected. PMID- 10521962 TI - Rate of bilirubin decrease as a risk predictor in hepato-biliary-pancreatic surgery. AB - BACKGROUND/AIMS: Although percutaneous transhepatic biliary drainage is widely performed for jaundice reduction, the clinical significance and mechanism responsible for delayed decrease of the bilirubin level remains unclarified. METHODOLOGY: The rate of bilirubin decrease was estimated in 104 consecutive patients who underwent drainage. Morbidity and mortality after major and minor operations for hepato-biliary-pancreatic diseases in groups showing slow and rapid bilirubin decrease were estimated. The risk factors for slow bilirubin decrease were also examined by uni- and multivariate analyses. RESULTS: A statistically significant difference between the slow and rapid bilirubin decrease groups was found only in the morbidity rates of major surgery (73% vs 28%, p < 0.05). Univariate analysis showed that the longer interval from onset of jaundice to drainage, the use of multiple catheters for jaundice reduction, and advanced age were significant risk factors for slow bilirubin decrease. These factors were found to be independent by multivariate analysis (p < 0.05) CONCLUSIONS: A slow rate of jaundice reduction is a predictor of high risk in major surgery. It can be attributed to 3 factors; longer term of undrained jaundice, multiple biliary drainage, and age. Careful post-operative management is required when major surgery is scheduled for patients with these risk predictive factors. PMID- 10521963 TI - Mirizzi's syndrome must be ruled out in the differential diagnosis of any patients with obstructive jaundice. AB - BACKGROUND/AIMS: Mirizzi syndrome is a rare benign complication of long-standing cholelithiasis and neither diagnostic modality nor clinical feature has a 100% sensitivity and specificity. The objective of our study was to call attention to the importance of this rare syndrome with its miscellaneous treatments. METHODOLOGY: Between January 1992 and June 1997, a total of 8 (4 females and 4 males) patients, who were operated and diagnosed as Mirizzi syndrome, were retrospectively evaluated. RESULTS: The mean age was 53.75 years. During the same time period 0.98% of the total 812 cholelithiasis patients were Mirizzi syndrome. The ultrasound was used in 7, computed tomography (CT) in 4 and endoscopic retrograde cholangiopancreatography (ERCP) in 2 cases. Ultrasound allowed the detection of cholelithiasis in all, but proximal bile duct dilatation in only 71% of cases. CT detected the non-specific findings of syndrome in 75% of cases. In 2 patients, because of the difficulties due to the patients themselves and the technical management problems. ERCP could not detect the pathology properly. In 2 of 5 type I patients, we performed only cholecystectomy and in another 2 cholecystectomy plus T-tube drainage. In 1 case, due to major hepatic duct injury during surgery, cholecystectomy plus hepaticojejunostomy over the Y-stent was performed. Biliary fistula developed in 1 patient with T-tube drainage and was successfully managed with conservative treatment. In all type II patients we preferred cholecystectomy plus choledochoduodenostomy and all of them were free of complications. CONCLUSIONS: If there is no question about the security of the common bile duct at surgery in type I patients, we recommended cholecystectomy, otherwise cholecystectomy plus exploration of common bile duct and/or drainage should be the procedure of choice. However, in type II patients cholecystectomy plus choledochoduodenostomy is a safe and effective procedure to perform. PMID- 10521964 TI - The influence of intravenous omeprazole on intragastric pH and outcomes in patients with peptic ulcer bleeding after successful endoscopic therapy--a prospective randomized comparative trial. AB - BACKGROUND/AIMS: The role of omeprazole in preventing rebleeding in patients with peptic ulcer bleeding after successful endoscopic therapy has been controversial. In this study, we used 3 different formulas of intravenous omeprazole in the above patients. We wished to compare the intragastric pH and outcomes among them. METHODOLOGY: Between July 1996 and May 1997, after having obtained initial hemostasis with endoscopic therapy, a total of 20 patients with peptic ulcer bleeding (spurting/oozing/non-bleeding visible vessel: 6/4/10) received intravenous bolus of omeprazole 20 mg every 3 hours; 20 patients (3/5/12) received intravenous bolus of omeprazole 40 mg every 6 hours; and, 20 patients (5/4/11) received intravenous bolus of omeprazole 80 mg every 12 hours for 3 days. One intragastric pH meter (Gastrograph Mark III, Medical Instruments Corp. Switzerland) was used to record 24-hour intragastic pH. RESULTS: The intragastric pH in the patients receiving omeprazole 20 mg every 3 hours was 6.1, 6.0-6.2 (mean: 95% CI); in patients receiving omeprazole 40 mg every 6 hours it was 6.4, 6.2-6.5; and, in patients receiving omeprazole 80 mg every 12 hours it was 5.8, 5.7-5.9. The duration of intragastric pH > 6.0 in omeprazole 20 mg every 3 hours was 70.9%, 57.3%-84.4% (mean: 95% CI); in omeprazole 40 mg every 6 hours it was 83.1%, 73.1%-93.1%; and, in omeprazole 80 mg every 12 hours it was 66%, 51.5% 80.4%. Patients with peptic ulcers receiving omeprazole 40 mg intravenous bolus every 6 hours had the highest intragastric pH as compared with the other 2 groups (p < 0.0001). There were no significant differences concerning rebleeding rates, volume of blood transfusion, hospital stay, numbers of operation and mortality among the 3 groups. CONCLUSIONS: After initial hemostasis had been obtained, patients with peptic ulcer bleeding receiving 40 mg intravenous bolus every 6 hours had the highest intragastric pH. However, they had similar outcomes with the other 2 groups. PMID- 10521965 TI - A case of classical carcinoid tumor of the gallbladder: review of the Japanese published works. AB - A 58 year-old man was admitted to Kimitsu Chuo Hospital complaining of epigastralgia. Abdominal ultrasound and computed tomography revealed a polypoid lesion at the neck of the gallbladder. Given the pre-operative diagnosis of gallbladder carcinoma, we resected the gallbladder along with the extrahepatic bile duct. There was a papillary tumor (25 x 16 mm) at the neck of the gallbladder. Histopathological examinations showed a subserosal nodular proliferation of uniform small tumor cells. Grimelius staining was slightly positive and Fontana-Masson staining was negative. Most of the tumor cells stained positively for chromogranin A and neuron-specific enolase (NSE), and some of the tumor cells were positive for pancreatic polypeptide. The presence of neurosecretory intracytoplasmic granules was proven ultrastructurally. It was diagnosed as a classical carcinoid tumor of the gallbladder. We reviewed the Japanese reported cases and discussed the difference in clinicopathological findings between classical and atypical carcinoid tumors of the gallbladder. Classical carcinoids of the gallbladder have neither a metastatic nor invasive character, and an extremely favorable prognosis compared with atypical carcinoids. The difference in character between classical and atypical carcinoids of the gallbladder is thought to be derived from their histogenetic origin. PMID- 10521966 TI - A case of bilioduodenal fistula treated with a self-expandable metallic stent. AB - We report the case of a 72 year-old female patient who suffered from biliary fistulae. The biliobiliary and bilioduodenal fistulae appeared after an operation for biliary bleeding. Conventional therapy for biliary fistula would be the disconnection of the fistula by either conservative or operative treatment. In the present case, however, it was preferable to enlarge the fistula to drain bile juice into the duodenum, rather than to close the fistula because it would have been difficult to achieve a tight adhesion with this operation. The enlargement by a plastic tube stent failed to drain the bile juice into the duodenum, because the sludge made the tube stenotic. Therefore, a self-expandable metallic stent was applied in this case. An expandable stent was used because a large final caliber is necessary to prevent stenosis of the fistula by sludge and mucosal hyperplasia. After insertion of a self-expandable metallic stent by the percutaneous transhepatic biliary drainage route, the patient has not suffered from cholestasis and cholangitis for the last 30 months. It can therefore be concluded that enlargement of the fistula by a self-expandable metallic stent is a convenient therapy for such biliointestinal fistulae. PMID- 10521967 TI - Percutaneous and endoscopic management of bile leak following endoscopic stone retrieval--a case report. AB - Endoscopic sphincterotomy with stone removal is the method of choice for the treatment of choledocholithiasis. The main complications of this procedure are bleeding, pancreatitis, intestinal perforation and cholangitis. Herein, we report on a case of bile peritonitis in a patient who underwent sphincterotomy and stone retrieval. The literature regarding the etiology and management of bile peritonitis is also reviewed. PMID- 10521968 TI - Management of a patient with hepatic-thoracic-pelvic and omental hydatid cysts and post-operative bilio-cutaneous fistula: a case report. AB - In humans, most hydatid cysts occur in the liver and 75% of these are single. Our patient was a 31 year-old male. His magnetic resonance imaging (MR) showed one cyst (15 x 20 cm) in the right lobe and three cysts (5 x 6 cm, 8 x 6 cm, and 5 x 5 cm) in the left lobe of the liver, two cysts (4 x 5 cm and 5 x 5 cm) on the greater omentum, and two cysts (15 x 10 and 10 x 10 cm) in the pelvis. The abdomen was entered first by a bilateral subcostal incision and then by a Phennenstiel incision. Partial cystectomy + capitonnage was done on the liver cysts; the cysts on the omentum were excised, and the pelvic cysts were enucleated. The cyst in the right lobe of the liver was in communication with a thoracic cyst. An air leak developed from the thoracic cyst which had underwater drainage and bile drainage from the drain in the cavity of the right lobe cyst. Sphincterotomy was done on the seventh post-operative day by endoscopic retrograde cholangiopancreatography (ERCP). No significant effect on mean bile output from the fistula occurred. Octreotide therapy was initiated, but due to abdominal pain and gas bloating the patient felt and could not tolerate, it was stopped on the fourth day; besides, it had no decreasing effect on bile output during the 4 days. Because air and bile leak continued and he had bile stained sputum, he was operated on on post-operative day 18. By right thoracotomy, the cavity and the leaking branches were closed. By right subcostal incision, cholecystectomy and T-tube drainage of the choledochus were done. On post operative day 30, he was sent home with the T-tube and the drain in the cavity. After 3 months post-operatively, a second T-tube cholangiography was done, and a narrowing in the distal right hepatic duct and a minimal narrowing in the distal left hepatic duct were exposed. Balloon dilatation was done by way of a T-tube. Bile drainage ceased. There was no collection in the cavity in follow-up CT scanning, so the drain in the cavity, and the drainage catheter in the right hepatic duct were extracted. Evaluation of the biliary ductal system is important in bilio-cutaneous fistulas, and balloon dilatation is very effective in fistulas due to narrowing of the ducts. PMID- 10521969 TI - Acute acalculous cholecystitis in a patient on total parenteral nutrition: case report and review of the Japanese literature. AB - Acute acalculous cholecystitis (AAC) is a rare and dangerous complication of various medical and surgical conditions. We report on a male patient with bile panperitonitis caused by gangrenous AAC, which developed while he was on total parenteral nutrition (TPN) for ileus related to obstructive colon cancer. We also review the relevant Japanese literature on AAC associated with TPN. Our patient suddenly developed right hypochondrial pain after 3 days of TPN while waiting for colon cancer surgery. We diagnosed acute AAC by ultrasonography, and salvaged the patient by cholecystectomy plus left colectomy. Early diagnosis by ultrasound is important for this critical condition. Knowledge of the risk of acute gangrenous cholecystitis during TPN may allow the physician to provide an appropriate diagnosis and treatment. PMID- 10521970 TI - Pre-operative imaging can diagnose torsion of the gallbladder: report of a case. AB - Torsion of the gallbladder is a rare disease and pre-operative diagnosis of the disease is uncommon. About 400 cases have been reported, but only 4 were diagnosed by pre-operative imaging. We report on a case of gallbladder volvulus diagnosed pre-operatively using pre-operative imaging with ultrasound and computed tomography. PMID- 10521971 TI - New frontiers in liver surgery. Two-stage liver surgery for the management of advanced metastatic liver disease. AB - BACKGROUND/AIMS: To assess the value and the safety of main portal branch transection combined with transarterial targeting locoregional neo and adjuvant immunochemotherapy, 32 patients suffering from advanced metastatic liver disease underwent two-stage hepatectomy. METHODOLOGY: From September 1995 to June 1999, 32 consecutive patients underwent two-stage surgery for advanced metastatic liver disease. Firstly we performed ligation and transection of the main portal branch corresponding to the liver lobe occupied by the tumor and introduction of an arterial jet port catheter towards the hepatic artery. After a locoregional transarterial targeting immunochemotherapy regimen the patient had a 2nd laparotomy for hemihepatectomy. Following surgery, locoregional targeting immunochemotherapy was carried out in all patients via the arterial port of the gastroduodenal artery as an adjuvant treatment. RESULTS: There were no operative deaths. Mean survival was 27 +/- 8 months. CONCLUSIONS: Two-stage liver surgery is an appealing alternative that increases the resectability rate and overall survival in patients with advanced metastatic liver disease and is associated with excellent quality of post-operative life. PMID- 10521972 TI - Consecutive re-explorations for final resection of initially unresectable pancreatic head carcinoma. AB - BACKGROUND/AIMS: The lack of high surgical expertise and specialization of the practicing surgeon may lead some patients with pancreatic cancer to die. This study also investigates the role of combined neo and adjuvant locoregional immunochemotherapy in patients considered initially as non-amenable to resection. METHODOLOGY: 32 patients underwent re-exploration aiming at pancreatic resection. After the initial diagnostic work-up 22 of them underwent pancreatic resection during the first re-exploration. The remaining 10 patients were judged again as unresectable. All 32 patients had 2 catheters introduced into a side arterial branch of the jejunal artery and vein for locoregional immunochemotherapy. Seven out of 10, considered as unresectable initially, had pancreatic resection after immunochemotherapy regimen. RESULTS: All patients survived surgery. Early morbidity included wound infection in 3, bleeding in 1 and leakage of gastric stump in 1 patient. Treatment related toxicity included leukopenia in 4 patients, anemia in 3 and fever and chills in 21. Mean follow-up was 62 +/- 1.2 months. One , 2-, 3- and 5-year survival was 100, 80, 70 and 48% respectively. CONCLUSIONS: Our results strongly support the necessity for neo and adjuvant locoregional immunochemotherapy and its contribution to prolongation of survival. PMID- 10521973 TI - Sialylated MUC1 mucin expression in normal pancreas, benign pancreatic lesions, and pancreatic ductal adenocarcinoma. AB - BACKGROUND/AIMS: Pancreatic cancer has the poorest prognosis of all gastrointestinal cancers. Because sialylated mucin influences the biologic behavior of carcinoma cells, we investigated sialylated MUC1 mucin expression in patients with pancreatic ductal adenocarcinoma. METHODOLOGY: The expression of sialylated MUC1 mucin was examined in 55 pancreatic ductal adenocarcinomas, 2 normal pancreas specimens, 3 chronic pancreatitis specimens, 1 ductal hyperplasia of the pancreas, 3 mucinous cystadenomas, and 2 liver metastases from pancreatic ductal adenocarcinoma. Expression was assessed by immunohistochemistry with a new monoclonal antibody (mAb) (MY.1E12). RESULTS: Sialylated MUC1 mucin was expressed in the cancer cell membrane in all the ductal carcinomas. The reaction product was seen at the apical aspect of cells when these were in tubule formation. This pattern was also detected in mucinous cystadenomas. However, it was seen diffusely in the cell membrane in single cancer cells or small clusters of cells without tubule formation and in metastatic liver tumors. Namely, invading or metastatic cancer cells expressed this type of mucin throughout the entire cell membrane. The expression of sialylated MUC1 mucin was not observed in specimens from normal pancreas, chronic pancreatitis, or ductal hyperplasia of the pancreas. In normal pancreas and these lesions, expression of sialylated Mession of sialylated MUC1 was limited to acini and secreted mucin. CONCLUSIONS: Sialylated MUC1 mucin, which is expressed throughout the cancer cell membrane, may be a factor in the metastatic potential of pancreatic ductal adenocarcinoma. PMID- 10521974 TI - Abdominal wall-lifting laparoscopic simple closure for perforated peptic ulcer. AB - BACKGROUND/AIMS: To develop and simplify the laparoscopy-assisted simple closure of perforated peptic ulcers by using conventional instruments and techniques. METHODOLOGY: An abdominal wall-lifting method was applied for laparoscopic simple closure with omental patch on 6 perforated peptic ulcer patients without using the pneumoperitonium. Our procedure added a 2 cm long right upper quadrant (RUQ) window over the perforation hole to enable the use of conventional instruments and techniques for simple closure, as well as thorough normal saline irrigation of the abdominal cavity. RESULTS: Satisfactory results were attained without mortality or morbidity. Benefits included small operation wounds and less need for analgesics. CONCLUSIONS: The RUQ window simplifies the suture/ligation technique by enabling the use of conventional instruments in laparoscopic surgery for perforated peptic ulcer. This procedure can be an alternative to the laparoscopic closure of perforated peptic ulcer. PMID- 10521975 TI - Ultrasonographic evaluation of portal blood flow using transhepatic carbon dioxide injection. AB - BACKGROUND/AIMS: The evaluation of portal blood flow in hepatic mass is important for the diagnosis of hepatocellular carcinoma. We have designed a new method to easily evaluate portal blood flow in hepatic mass using ultrasonography with injection of carbon dioxide into the intrahepatic portal vein by direct puncture with a fine needle. METHODOLOGY: We evaluated 29 masses in the liver of 20 patients ultrasonically with injection of carbon dioxide into the intrahepatic portal vein. RESULTS: Of 29 space-occupying lesions (SOLs), 13 were found to have portal blood flow and 16 were found to have no portal flow by this method. All 15 SOLs which had no portal flow were histologically confirmed to be hepatocellular carcinoma. Of 13 SOLs with portal flow, 5 were confirmed to be hepatocellular carcinoma. For 7 of 9 SOLs in which both this method and arterial portographic computed tomography were performed, the results were in agreement. CONCLUSIONS: Ultrasonographic evaluation of portal blood flow using transhepatic carbon dioxide injection appears to be useful for the evaluation of portal flow in mass and may aid in the diagnosis and management of mass in patients with liver disease. PMID- 10521976 TI - Relationship between serum levels of soluble interleukin-2 receptor and various disease parameters in patients with squamous cell carcinoma of the esophagus. AB - BACKGROUND/AIMS: Soluble interleukin-2 receptor (sIL-2R) is a useful biomarker for the management of hematologic malignancies. We determined the significance of serum sIL-2R levels in patients with squamous cell carcinoma of the esophagus. METHODOLOGY: The correlation between serum sIL-2R levels and a variety of clinicopathologic factors in 51 patients with squamous cell carcinoma of the esophagus was evaluated. We also investigated the expression of IL-2R in the resected specimen using immunohistochemical staining. RESULTS: Expression of IL 2R was detected in primary esophageal carcinoma cells as well as infiltrating mononuclear cells. Serum sIL-2R levels in the 51 patients with esophageal cancer were significantly higher than those in the 18 healthy volunteers (p < 0.01). Multivariate analysis showed that pM, tumor size, and lymph node metastasis was significantly correlated with serum sIL-2R levels. Univariate analysis demonstrated that tumor size, pM, pTNM stage, and resectability were parameters which were significantly correlated with serum sIL-2R levels. There was no relationship between serum sIL-2 levels and tumor depth, lymph node metastasis, pT, histologic type, or curability. CONCLUSIONS: These results suggest that the serum sIL-2R levels may be an indicator of the extent of disease and resectability in patients with esophageal squamous cell carcinoma. Immunohistochemical staining suggests that esophageal cancer cells and infiltrating mononuclear cells may be the source of sIL-2R in the serum. PMID- 10521977 TI - Diagnostic possibilities and clinical indications of policlonal labeled Ig in the bowel inflammatory disease. AB - BACKGROUND/AIMS: Ulcerative colitis (UC) and Crohn's disease (CD) represent the more common forms of idiopathic inflammatory bowel disease. They have a peculiar course, which is characterized by exacerbation and submissions, and a symptomatology that changes in relation to severity and extension of the lesions. The aim of this study is to establish the diagnostic usefulness of 99m Tc human policlonal immunoglobulin (HIG) imaging in patients with inflammatory bowel disease (IBD). METHODOLOGY: During the period between September 1995 and September 1997 we submitted a group of 32 patients affected by UC (n = 22) and CD (n = 10), to Human Immunoglobulin labeled with Technetium 99m (99m Tc-HIG) scintigraphy. The diagnosis of IBD was obtained in all cases by endoscopy with biopsy. RESULTS: Scintigraphic examination with labeled HIG showed an abnormal intestinal fixation (ileal or colic) of the marked immunoglobulins in 17 patients (77.2%) affected by UC and in 8 patients (80%) affected by CD. In 5 (22.7%) and 2 (20%) patients, respectively, the scintigraphic examination revealed a normal distribution of the immunoglobulins without appreciable focal accumulation. Among the patients with a negative scintigraphic examination, only one presented an endoscopic and histologic report that documented UC in an active stage. The histologic and endoscopic findings observed in all cases of CD were similar to that of HIG scintigraphy in 7 of the 8 scintigraphic-positive cases and in 1 of the 2 scintigraphic-negative patients. CONCLUSIONS: The authors, on the basis of their results, suggest that this diagnostic strategy may have a significant role in the diagnostic protocol and in the follow-up of chronic inflammatory bowel disease. PMID- 10521979 TI - Helicobacter pylori and prurigo nodularis. AB - BACKGROUND/AIMS: Numerous data have suggested that there may be a relationship between Helicobacter pylori (H. pylori) infection and extragastric diseases, including dermatological pathologies. We studied some cases of Hide's Prurigo Nodularis (NP), a very itchy skin disease of unknown origin, in order to asses whether there is a pathogenic correlation between PN and H. pylori infection. METHODOLOGY: Prurigo Nodularis had been diagnosed clinically and histologically in 42 outpatients (27 females and 15 males with mean age of 62 +/- 5 years). All patients were screened for H. pylori infection by esophagogastroduodenoscopy, histologic examination and specific immuno-enzymatic determination. Specific pharmacological treatment was administered to all patients with H. pylori infection. RESULTS: H. pylori colonization was observed in 40/42 patients examined and 32/40 patients presented some immunologic alterations. After the pharmacological treatment, endoscopy and rapid urease test confirmed eradication of H. pylori in 39/40 cases; itching was greatly reduced in the latter and microscopic examination of repeated skin biopsies revealed an improved histologic picture in patients affected by PN associated with H. pylori infection. CONCLUSIONS: The concomitant presence of skin disease, H. pylori infection and immune disorders infers that there may be a pathogenic connection between bacterial infection and the inflammatory alteration of PN. We believe that the pharmacologic treatment induced remission of the skin lesions by direct control of H. pylori chronic infection; in fact, H. pylori may have triggered or enhanced the vasculitis which, in turn, may have enhanced the clinical signs and inflammatory histologic component of PN. PMID- 10521978 TI - High-dose intravenous cyclosporine in steroid refractory attacks of inflammatory bowel disease. AB - BACKGROUND/AIMS: To determin whether cyclosporine is effective in inducing remission in patients with severe active inflammatory bowel disease, refractory to steroids. METHODOLOGY: Twenty-three patients with severe and steroid refractory inflammatory bowel disease (15 ulcerative colitis and 8 Crohn's disease) were included. The Mayo Clinic Score and the CDAI were used to evaluate activity. Cyclosporine (4 mg/kg/day) was administered for a maximum of ten and a minimum of 7 days. RESULTS: Ten of the 15 ulcerative colitis patients achieved remission with a mean response lag time to onset improvement of 8 days. Seven of these patients remained stable with mesalazine 4 months after cyclosporine treatment. Two patients relapsed and underwent colectomy on the 50th and 200th day after treatment. Five patients presented no response and required urgent colectomy. Six of the 8 Crohn's disease patients achieved remission with a mean response lag time to onset improvement of 7 days. The 6 patients remained stable with mesalazine four months after cyclosporine treatment. The other 2 developed reversible renal failure and had to be released from the study. CONCLUSIONS: Intravenous high dose cyclosporine is effective and can be used as a rapid onset treatment for acute steroid refractory IBD. PMID- 10521980 TI - Bactericidal/permeability-increasing protein in colonic mucosa in ulcerative colitis. AB - BACKGROUND/AIMS: Increased mucosal concentration of bactericidal/permeability increasing protein (BPI) has been shown in inflammatory bowel diseases. The purpose of the present study was to investigate the relationship between the mucosal concentration of BPI and the grade of mucosal inflammation in ulcerative colitis. METHODOLOGY: Samples of colonic mucosa from 12 patients with ulcerative colitis and from 8 control patients were studied. The concentration of BPI in tissue extracts was measured by a time-resolved fluoroimmunoassay. The concentration of BPI was compared between samples with histological inflammatory changes of different severity. BPI was localized in tissue sections by immunohistochemistry. RESULTS: The concentration of BPI was higher (p < 0.001) in samples of colonic mucosa from patients with ulcerative colitis (median: 3.2 micrograms/g, range: 0.3-22.6 micrograms/g) than in control samples (0.4 microgram/g, 0.1-0.6 microgram/g,). Moreover, the concentration of BPI was higher (p = 0.015) in samples with severe inflammation (2.5 mu/g, 0.3-22.6 micrograms/g) than in those with mild inflammation (0.5 mu/g, 0.3-2.5 micrograms/g). The concentration of BPI in mucosal samples correlated well with the degree of histological inflammation (Spearman R = 0.70, p = 0.01). BPI was localized in polymorphonuclear leukocytes in the mucosa and stroma of the colonic wall. CONCLUSIONS: The concentration of BPI is increased in the colonic mucosa of patients with ulcerative colitis. The increase in the concentration of BPI in colonic mucosa seems to be closely associated with the inflammatory activity of ulcerative colitis. PMID- 10521981 TI - Vascular reconstruction of the hepatic artery using the gastroepiploic artery: a case report. AB - A 59 year-old woman with obstructive jaundice secondary to proximal bile duct carcinoma underwent percutaneous transhepatic biliary drainage (PTDB). This revealed complete obstruction of the bifurcation of the hilar hepatic duct and encasement of the right hepatic artery. Wedged hilar hepatectomy with combined resection of the extrahepatic bile duct, gallbladder, and the encased right hepatic artery was performed. The hepatic artery was reconstructed using an in situ right gastroepiploic artery (GEA) pedicle graft. The anastomosis was protected with fatty tissue from the greater omentum. This technique can be used to reconstruct the hepatic artery after radical surgery for malignant hepatobiliary and pancreatic disease. PMID- 10521982 TI - Prurigo nodularis (Hyde's prurigo) disclosing celiac disease. AB - A case of prurigo nodularis (Hyde's prurigo) preceding by 15 years the diagnosis of celiac disease is presented. In particular, the association between the clinical course of prurigo nodularis and the compliance with gluten-free diet is reported and discussed. PMID- 10521984 TI - Hepatic and retroperitoneal tumor resection for late metastases of a Wilms' tumor in an adult patient--a case report. AB - Hepatic metastases after a Wilms' tumor in adult patients are seen extremely rarely. A 21 year-old male patient developed liver metastases 13 years after resection of a primary left extrarenal Wilms' tumor. In this case, without any other metastases, extended right curative hepatic lobectomy was performed. The patient was re-admitted 4 months after the hepatic lobectomy for a resection of a new Wilms' tumor metastatic mass in the area of the pancreatic tail. The patient received adjuvant high dose systemic chemotherapy with ordinary bone marrow cell rescue after the 2nd operation. He is alive and well with no signs of new metastases 18 months after surgery and adjuvant chemotherapy. PMID- 10521983 TI - Hepatic artery aneurysm associated with upper gastrointestinal bleeding after intrahepatic artery chemotherapy. AB - We present the course of illness of a 56 year-old patient with acute gastrointestinal bleeding after penetration of an aneurysm of the hepatic artery in the duodenal bulb. A diffuse intrahepatic metastasis of a carcinoid was treated with a loco-regional intraarterial chemotherapy via a catheter system implanted in the gastroduodenal artery. Five months after the catheter implantation melena occurred. The gastrointestinal arterial bleeding from the penetrating aneurysm presented a life-threatening situation. This cause of bleeding could not be brought under control endoscopically. Immediate surgical management became necessary. PMID- 10521985 TI - The experience of endoscopic tissue glue injection in the treatment of hepatic sarcoidosis related gastric variceal bleeding: report of a case. AB - A case of hepatic sarcoidosis complicated with portal hypertension and gastric variceal bleeding is described. A 53 year-old male suffered from persistent fever and massive hematemesis. Acute gastric variceal bleeding was diagnosed. Endoscopic tissue glue injection stopped this acute episode and ablated the varices after another two sessions of endoscopic tissue glue injection treatment. Subsequent administration of corticosteroids improved the symptoms and liver function. This was probably the first case of hepatic sarcoidosis associated with gastric variceal bleeding which was successfully treated by endoscopic tissue glue injection to be reported. PMID- 10521986 TI - Right hepatic lobectomy for recurrent cholangitis after bile duct and hepatic artery injury during laparoscopic cholecystectomy: report of a case. AB - A patient is reported who required a right hepatic lobectomy for recurrent cholangitis due to injury of the major bile ducts and the right hepatic artery during laparoscopic cholecystectomy. A 39 year-old woman with acute cholecystitis underwent laparoscopic cholecystectomy. A laparotomy was performed due to a bile duct injury at the hepatic bifurcation. After surgery, she suffered from recurrent cholangitis due to inadequate biliary reconstruction. A right hepatic lobectomy and reconstruction of the left hepatic duct was required because of right hepatic lobe atrophy and recurrent cholangitis. After the 2nd operation, she was active and exhibited no evidence of recurrence at 22 months. PMID- 10521987 TI - Urokinase-type plasminogen activator and plasminogen activator inhibitor-1 as a prognostic factor in human colorectal carcinomas. AB - BACKGROUND/AIMS: The urokinase pathway of plasminogen activation is known to be involved in proteolytic degradation of the extracellular matrix during carcinoma invasion. METHODOLOGY: We immunohistochemically examined 97 colorectal carcinomas for the expression of urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor-1 (PAI-1) to investigate whether uPA and PAI-1 expressions could serve as prognostic parameters; the gene expression of uPA and PAI-1 in human colon cancer tissues was also analyzed using reverse transcription polymerase chain reaction (RT-PCR). RESULTS: The relative expression levels of uPA and PAI-1 mRNAs were well correlated with immunoreactivities of uPA and PAI 1, respectively (p < 0.05). In immunohistochemical staining, diffuse specific staining was observed in the cytoplasm of carcinoma cells. uPA expression was detected in 57 carcinoma specimens (58.8%) and PAI-1 expression was detected in 36 specimens (37.1%). With regard to 5-year overall survival rate, patients whose tumors had positive uPA and negative PAI-1 immunoreactivities had a significantly poorer prognosis (p < 0.05). In multivariate analysis, the combined variable of uPA and PAI-1 was shown to be an independent prognostic indicator. CONCLUSIONS: Our results suggest that immunohistochemical combined analysis of uPA and PAI-1 may be a useful prognostic factor in colorectal carcinoma patients. PMID- 10521988 TI - Individual fecal alpha 1-antitrypsin excretion reflects clinical activity in Crohn's disease but not in ulcerative colitis. AB - BACKGROUND/AIMS: The natural course of fecal alpha 1-antitrypsin (AAT) excretion was assessed in patients with inflammatory bowel disease (IBD) to evaluate its role in monitoring their clinical disease activity. METHODOLOGY: A prospective cohort pilot study was performed in 9 patients with Crohn's disease (CD) and 3 individuals with ulcerative colitis (UC). Subjects were investigated at regular monthly intervals for about one year for (a) parallel AAT stool and serum concentrations by standard immunonephelometry, and (b) for clinical disease activity by Crohn's Disease Activity Index (CDAI) in CD patients, and by Clinical Activity Index (CAI) in UC subjects. Absolute results during follow-up were each referred to individual findings at study entry as relative results. RESULTS: While absolute fecal AAT concentration did not correlate with disease activity indices (p > 0.26), relative fecal AAT concentration significantly correlated to concurrent relative CDAI score in CD patients (p < 0.001, r = 0.67), but not to relative CAI score in UC subjects (p = 0.92). Monthly intraindividual variation of fecal AAT excretion did not predict development of either disease activity index (p > 0.14). CONCLUSIONS: Individual fecal AAT excretion closely reflects clinical course in CD subjects, but not in UC patients. It does not predict symptomatic deterioration in these individuals, at least on a short-term basis. PMID- 10521989 TI - Neutrophil-elastase in chronic inflammatory bowel disease: a marker of disease activity? AB - BACKGROUND/AIMS: Neutrophil elastase is a proteinase which exists in granulocytes and plays an important role in the pathogenesis of inflammatory disorders. In inflammatory bowel disease there is a leukocyte infiltration of the bowel mucosa. The purpose of this study was to examine whether plasma elastase represents a reliable laboratory marker for establishing the activity of chronic inflammatory bowel disease. METHODOLOGY: We measured plasma elastase concentrations in 61 patients suffering from either Crohn's disease or ulcerative colitis and compared these data with other clinical and laboratory findings and with elastase concentrations in 40 healthy controls. The sensitivity and specificity of the elastase values in chronic IBD were calculated with the use of concomitant measurements of CRP and ESR. RESULTS: Plasma levels were found to be significantly higher in patients (49 micrograms/l) compared with healthy controls (23 micrograms/l). Patients with active disease had higher plasma levels than patients in remission. In general, the sensitivity of elastase to detect active inflammatory bowel disease was about 60%; the specificity was 65%. For patients in remission, the sensitivity was higher than 80%. However, there was a wide range of overlapping values between chronic inactive patients and those with moderately active disease. CONCLUSIONS: We conclude that plasma elastase is a useful independent marker of disease activity in inflammatory bowel disease. Especially for identifying patients in remission, the measurements of elastase seem to be more suitable than other parameters of inflammation, like CRP or ESR. PMID- 10521990 TI - The role of surgery in the treatment of liver metastases for colorectal cancer patients. AB - BACKGROUND/AIMS: Liver metastases deriving from colorectal cancer can be treated with curative intention in a select number of patients. Controversy does, however, persist pertaining to the impact of adjuvant treatment strategies. The aim of this study is to elucidate upon the various treatment modalities for patients suffering from liver metastases of colorectal primary tumor as well as to provide a rationale for surgical and adjuvant treatment. METHODOLOGY: From November 1987 to September 1998, a total of 449 consecutive patients suffering from liver metastases deriving from a colorectal cancer were documented at our institution in a prolective study. Prognostic factors providing the most beneficial outcome (whether with surgical and/or adjuvant treatment modalities) were analyzed by univariate and multivariate analysis. RESULTS: Whenever possible, curative (R0) surgical resection of colorectal liver metastases provides the most benefit to the patient. Multivariate analysis revealed tumor infiltration of the lymph nodes of the hepatoduodenal ligament and metachronous occurrence of liver metastases as most independent factors related to survival. CONCLUSIONS: Adjuvant post-operative chemotherapy fails to significantly improve survival following resection of liver metastases when compared to the liver resection only group. In patients with unresectable metastases, regional arterial chemotherapy did not improve survival significantly when compared with systemic chemotherapy. PMID- 10521991 TI - Evaluation of local excision for sessile-type lower rectal tumors. AB - BACKGROUND/AIMS: A study was undertaken to evaluate the surgical morbidity and risk of recurrence in patients undergoing local excision for rectal carcinoma. METHODOLOGY: Twenty patients with well or moderately differentiated lesions less than 6 cm in diameter, which were difficult to remove endoscopically but had no clinically involved regional nodes, were eligible for local excision. RESULTS: The transanal approach was simple and had few complications. After posterior parasacral excision, 3 patients developed rectocutaneous fistulas, and 1 of them died. None of the 12 patients with cancer extension limited to the lesser depth of the submucosa died of rectal cancer. Among 8 patients with cancer invasive into the greater depth of the submucosa or deeper, 5 underwent additional radical resection, and regional lymph node metastasis was revealed in 2. CONCLUSIONS: Transanal excision is suitable for curative resection of lower rectal tumors when the cancer extends only to the lesser depth of the submucosa. The procedure should be defined as a total excisional biopsy until the results of histologic examination are obtained. PMID- 10521992 TI - Platelet activating factor in stool from patients with ulcerative colitis and Crohn's disease. AB - BACKGROUND/AIMS: Platelet activating factor (PAF) is a potent endogenous mediator in inflammatory processes. The role of this mediator, especially in connection with the unknown etiology of chronic inflammatory bowel diseases, remains poorly understood. A determination of PAF in stool may be helpful in recognizing quiescent inflammations in chronic inflammatory bowel diseases. A simple and reliable method for the determination of PAF in stool seems to be necessary to achieve this goal. METHODOLOGY: PAF analysis was performed with the help of a commercial PAF radioimmunoassay (RIA) kit after solid phase extraction (SPE) of ethanolic stool extracts. PAF was determined in stool from 10 healthy volunteers (m = 4; f = 6), 13 patients with ulcerative colitis (m = 7; f = 6) and 15 patients with Crohn's disease (m = 9; f = 6). Fecal PAF concentrations were compared with activity index of disease, endoscopic index, localization of lesions, leukocyte count, erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), medical prednisolone treatment, sex and age of the patients. RESULTS: In healthy volunteers, no PAF was detectable in stool. In patients with Crohn's disease 319.2 +/- 143.5 pg PAF/g stool and in patients with ulcerative colitis 824.9 +/- 408.7 pg PAF/g stool could be determined. A significant correlation (p < 0.05) was found between PAF-content in stool and the endoscopical index and intestinal localization of inflammatory lesions. No further correlations could be detected in our patients. CONCLUSIONS: Fecal PAF assessment may be used clinically as a non-invasive method to estimate severity of mucosal inflammation in patients with inflammatory bowel disease (IBD). PMID- 10521993 TI - Mucosal and plasma prostaglandin E2 in ulcerative colitis. AB - BACKGROUND/AIMS: Despite extensive studies, the role of prostaglandins in the course of inflammatory bowel diseases and their possible usefulness as predictive indicators of inflammation, remain largely speculative. The aim of this study was to determine whether mucosal and plasma concentrations of prostaglandin E2 (PGE2) are affected by the clinical course and degree of colonic injury in patients with ulcerative colitis. METHODOLOGY: PGE2 concentration was measured with an enzyme immunoassay (EIA) in biopsies of rectal mucosa and in the plasma of 38 patients with ulcerative colitis and 12 controls. Patients were divided into groups according to mild or severe clinical course of the disease, and with respect to scored endoscopical picture. RESULTS: Ulcerative colitis resulted in an increase of mucosal and plasma concentrations of PGE2, that was significantly elevated in patients with a severe clinical course of the disease. These concentrations increased depending on degree of mucosal injury. A significant, positive correlation with endoscopical score regarding plasma and mucosal PGE2 concentration, as well as between them, was found. CONCLUSIONS: Plasma and mucosal PGE2 rise simultaneously with degree of colonic injury. Because of a good correlation with mucosal injury and PGE2 content, measurement of plasma PGE2 could be considered as a possible surrogate marker of bowel inflammation. PMID- 10521994 TI - Practicality of 5-aminosalicylic suppositories for long-term treatment of inactive distal ulcerative colitis. AB - BACKGROUND/AIMS: Topical 5-aminosalicylic acid (5-ASA) is regarded as an effective form of therapy for distal ulcerative colitis. Unfortunately, experience about acceptability and tolerance of long-term treatment with 5-ASA suppositories is still meager. METHODOLOGY: We have evaluated the performance of 5-ASA suppositories as maintenance treatment in 34 patients with inactive distal colitis. Prior to entering the study, all patients were in clinical remission for 1-12 months. A single 500 mg 5-ASA suppository was administered nightly for 12 months in every patient and all other treatments were discontinued. Clinical evaluation was performed at 3, 6 and 12 months. RESULTS: Four patients withdrew from the study within the first 3 months because of local intolerance to the suppositories. Score of both comfort and tolerance was significantly higher in patients who continued treatment than in those who withdrew. Two patients who stayed in remission withdrew, one at 7 months because of thrombocytopenia and the other at 9 months for personal reasons. Of the 28 remaining patients, 7 relapsed and 21 remained in clinical remission at the end of the 12-month study period. Eleven of the 21 responder patients agreed to a sigmoidoscopic examination, that confirmed remission in all of them. Eight out of the 9 patients who had previously received 5-ASA enemas preferred suppositories to enemas. Apart from the 4 patients who did not tolerate suppositories, 26 patients considered this form of therapy quite comfortable. CONCLUSIONS: 5-ASA suppositories are generally well tolerated and considered comfortable for treatments of at least one year. PMID- 10521995 TI - Role of prostaglandin E2 in rat colon carcinoma. AB - BACKGROUND/AIMS: It has previously been reported that prostaglandin E2 (PGE2) promotes colon carcinogenesis. We therefore studied the effects of long-term administration of prostaglandin E2 (PGE2) on colon carcinogenesis in rats. METHODOLOGY: Rats received intrarectal n-methyl-n-nitrosourea (MNU) or n-methyl-n nitrosoguanidine (MNNG) to induce the formation of colonic tumors. Rates then received indomethacin (IND) and/or PGE2, or nothing. After 44 weeks (MNU group) or 46 weeks (MNNG group), colon lesions were identified histologically and colonic mucosa PGE2 concentrations were measured by radioimmunoassay. RESULTS: The incidence of colon carcinoma in the control, MNU, MNU + IND, MNU + PGE2, and MNU + IND + PGE2 groups was 0/14, 5/15, 0/14, 4/10, and 2/9, respectively. In the MNNG group, no tumors were observed. Induction of colon carcinomas by MNU was completely inhibited by IND, while exogenous PGE2 blocked the inhibitory effect of IND. However, PGE2 administration did not accelerate colon carcinogenesis. Neither MNU nor MNNG alone resulted in increased colonic mucosal PGE2 concentrations, whereas exogenous PGE2 administration significantly increased mucosal PGE2 concentrations. IND significantly decreased the mucosal concentration of PEG2 with or without PGE2 administration. CONCLUSIONS: These results suggest that endogenous PGE2 in colon mucosa may be adequate to promote colon carcinoma. To block colon carcinogenesis, PGE2 levels in colonic mucosa must be decreased to less than endogenous levels. PMID- 10521996 TI - A case of MALT (mucosa-associated lymphoid tissue) lymphoma occurring in the rectum. AB - A case of mucosa-associated lymphoid tissue (MALT) lymphoma of the rectum is reported. A 64 year-old woman was referred to us for the evaluation of occult blood in the stool. A hard mass was palpable on digital examination. Biopsy specimens revealed a histologic picture compatible with MALT lymphoma. Abdominoperineal excision of the rectum was carried out. Chemotherapy was not performed, and the post-operative course was uneventful, with no evidence of recurrence for 2 years and 11 months. Surgical resection is an effective therapy for MALT lymphoma of the rectum. PMID- 10521997 TI - Carcinoma of the colon with synchronous hepatic metastasis in a cirrhotic liver harboring a hepatocellular carcinoma. AB - Tumor metastasis to a cirrhotic liver is rare. It has been suggested that colorectal cancer does not metastasize to the cirrhotic liver. We reported a 65 year-old man, a known carrier of hepatitis B surface antigen, diagnosed to have hepatocellular carcinoma with routine screening. A partial hepatectomy with resection of segments VI and VII was performed. The hepatectomy specimen revealed a 4.5 cm diameter HCC in a cirrhotic liver. Incidentally, 0.8 cm diameter ulcer at the descending colon. Histological examination of the left hemicolectomy specimen showed a moderately differentiated adenocarcinoma. PMID- 10521998 TI - Effect of omeprazole and amoxicillin plus metronidazole on the eradication of Helicobacter pylori and the healing of duodenal ulcer: comparison with a historical control. AB - BACKGROUND/AIMS: To test the hypothesis of equivalence of an omeprazole 7-day triple therapy without subsequent acid suppression and a historical ranitidine 12 day triple therapy (recruiting phase 1989-91) with subsequent acid suppression in their effect on the eradication of Helicobacter pylori (H. pylori) and the healing of duodenal ulcer. METHODOLOGY: Seventy-seven patients with H. pylori positive duodenal ulcers received a 7-day treatment with amoxicillin 750 mg tid and metronidazole 500 mg tid. Additional omeprazole 20 mg or 40 mg once daily was given to 39 and 38 of the patients, respectively. Endoscopy was performed before treatment and four weeks after cessation of therapy. RESULTS: The cumulative intention-to-treat (ITT) H. pylori-eradication rate was 66% (51/77) as compared to 89% (46/52) for the historical control (p < 0.05). The corresponding ulcer healing rates were 90% (69/77) and 92% (48/52). Primary metronidazole resistance (PMR) had escalated from 10% to 27% within 6 years resulting in eradication rates of 84% for sensitive and 19% for resistant strains (p < 0.001). PMR could be demonstrated in 45% of all female, but only in 17% of the male patients (p < 0.05). In the patients with H. pylori eradication, the ulcers healed in 98% (50/51) as compared to 73% (19/26) in those with persistent infection (p < 0.005). Analysis based on the presence of PMR showed ulcer healing rates of 95% (53/56) for sensitive and 76% (16/21) for resistant strains (p < 0.05). Improvement of pain also showed a significant correlation with successful eradication. H. pylori-eradication, healing and symptom relief were similar in the omeprazole 20 mg and 40 mg groups. CONCLUSIONS: The effect of amoxicillin plus metronidazole plus antisecretory agent on the eradication of H. pylori has decreased markedly during the past 6 years due to the escalation of PMR. Doubling of the omeprazole dose does not affect outcome. Cure of the infection as well as metronidazole susceptibility enhance duodenal ulcer healing and symptom relief. Acid suppression following a successful 1-week anti-HP therapy is not required for duodenal ulcer treatment. PMID- 10521999 TI - A three-day course of intravenous omeprazole plus antibiotics for H. pylori positive bleeding duodenal ulcer. AB - BACKGROUND/AIMS: This prospective trial aimed to test the efficacy of 3-day intravenous omeprazole plus antibiotics for Helicobacter pylori (H. pylori) eradication rate, and to see whether individualized response to omeprazole in intragastric pH elevation will alter the success of eradication. METHODOLOGY: One hundred and thirty-eight cases with H. pylori-positive duodenal ulcer bleeding were randomized into four therapy groups: Group 1 (n = 32) received a 3-day course of intravenous omeprazole (80 mg loading then 40 mg q 9 am & 9 pm) plus ampicillin/salbactum (1.5 gm i.v. loading then 750 mg q 9 am, 3 pm, & 9 pm); Group 2 (n = 35) followed protocol as for Group 1 except the antibiotics were metronidazole and erythromycin (both 500 mg i.v. q 9 am, 3 pm, & 9 pm). Group 3 (n = 31) followed protocol as for Group 1 and further added with erythromycin (both 500 mg i.v. q 9 am, 3 pm, & 9 pm). Group 4 served as a control group (n = 40) receiving oral dual therapy after leaving the emergency room (omeprazole 20 mg and amoxycillin 1 g bid x 2 weeks). In each case, three gastric biopsies were done for total histologic density of H. pylori (THPD) (range: 0-15) before, 1 day and 6 weeks after completion of therapy. Except for the control group, the 24 hour ambulatory intragastric pH meter (MIC Inc, Gastrograph Spark III, Swiss) was inserted as possible on the 2nd day of therapy. RESULTS: The 3-day intravenous regimens achieved high clearance rates of H. pylori (Group 1: 93.8%; Group 2: 93.9%; Group 3: 100%). The eradication rates of H. pylori in Groups 1-4 were 43.8%, 57.1%, 58.1%, and 72.8%, respectively. In Groups 1-3, the H. pylori eradicated cases had lower pre-treatment THPD than non-eradicated cases (6.01 vs. 9.24, p < 0.001). Among 72 cases with pH meter insertion, the percentage of intragastric pH > 5.3 during 24-hour was not different among 35 H. pylori non eradicated and 37 eradicated cases (78.7 vs. 76.7%, p > 0.05). CONCLUSIONS: The 3 day intravenous regimens may achieve clearance of H. pylori quickly. However, they were not so effective for eradication, especially in cases with higher bacterial loads. The interindividual response to omeprazole in intragastric pH elevation under the study dosage had insignificant variations to alter the success of eradication. PMID- 10522000 TI - The efficacy of the third pump inhibitor--pantoprazole--in the short-term treatment of Chinese patients with duodenal ulcer. AB - BACKGROUND/AIMS: To study the efficacy and tolerability of pantoprazole 40 mg once daily before breakfast compared with ranitidine 300 mg once daily at bedtime in Chinese patients with duodenal ulcer, and to evaluate the relationship between Helicobacter pylori (H. pylori) clearance and ulcer healing rate. METHODOLOGY: A total of 160 patients (80 in each group) with endoscopically diagnosed, active duodenal ulcers were studied in this randomized double-blind trial. Endoscopy was performed after 2 weeks of treatment. If unhealed, then the patients were re endoscoped after an additional 2 weeks of similar treatment. RESULTS: The healing rates after 2 and 4 weeks were 61.3% and 97.3%, respectively in the pantoprazole group, and 50.7% and 76.9% in the ranitidine group. The difference between the two groups was significant at 4 weeks (p < 0.01, per protocol analysis). The rate of pain free ulcer was higher in the pantoprazole group than in the ranitidine group at 2 weeks (84.2% vs. 59.6%, p < 0.01). Higher clearance of H. pylori was also observed in the pantoprazole group compared with the ranitidine group at 4 weeks (20% vs. 0%, p = 0.05). The healing rate tended to be higher in patients who were H. pylori-cleared at 2 weeks (p = 0.07) in the pantoprazole group. Both medications were well tolerated without any serious adverse effects. CONCLUSIONS: Pantoprazole 40 mg daily is superior to ranitidine 300 mg daily in the short-term treatment of acute duodenal ulcer in Chinese patients, in terms of ulcer healing and pain relief, and appears to be well-tolerated. PMID- 10522001 TI - A case of abscess caused by a penetrating duodenal ulcer. AB - A case of abscess caused by a penetrating duodenal ulcer in a 34 year-old female patient is presented. She had a past history of duodenal ulcer and presented with a low grade fever which had persisted for 1 month. Abdominal ultrasound confirmed a hypoechoic mass and computed tomography revealed a low density area in the posterior side of the hepatoduodenal ligament. The common bile duct and portal vein were compressed. Mild peripheral enhancement was detected. Laparotomy was performed and an abscess in the posterior side of the hepatoduodenal ligament was confirmed. The abscess was firmly adhered to the lesser curvature side of the bulbus and a penetrating duodenal ulcer scar was noted. In conclusion, this report describes a rare event where penetrating duodenal ulcer formed an abscess with only mild complaints. PMID- 10522002 TI - Free jejunal pouch graft reconstruction after a resection of hypopharyngeal or cervical esophageal cancer. AB - BACKGROUND/AIMS: Pharyngoesophageal reconstruction using the free vascularized jejunal graft sometimes results in dysphagia and this may be caused by anastomotic stenosis at either the distal or proximal anastomotic site, graft contractility and the entrapment of food in the blind loop after an end-to-side pharyngojejunostomy. We therefore applied pouch procedures to the free jejunal graft in order to improve the ability for such patients to consume normal food. METHODOLOGY: We performed this procedure on 4 patients with pharyngoesophageal cancer located within the cervical regions. RESULTS: As a result, the following post-operative complications occurred in 1 case each: anastomotic leakage at the pharyngojejunostomy (proximal anastomosis) which healed spontaneously, and anastomotic stenosis in jejunoesophagostomy (distal anastomosis) which improved after performing endoscopic dilatation. CONCLUSIONS: However, these complications were not thought to be due to the pouch procedures and the passage of food was found to be excellent in all cases at the time of discharge. PMID- 10522003 TI - Preserved esophagogastric manometric motility in patients after distal gastrectomy. AB - BACKGROUND/AIMS: A prospective study resolved whether commonly employed distal gastrectomies could influence the esophagogastric manometric measurements and the role of vasoactive intestinal polypeptide in mediating motility after surgery. METHODOLOGY: Studied groups consisted of 20 patients following radical subtotal gastrectomy for gastric cancer, 20 patients after subtotal gastrectomy for duodenal ulcer and 20 controls. Fasting blood was obtained to measure serum vasoactive intestinal polypeptide levels. A pneumohydraulic infusion system measured esophagogastric motility parameters. RESULTS: Measured lower esophageal sphincter pressures in subjects of gastric cancer surgery, duodenal ulcer surgery and controls were 15.3 +/- 4.7, 13.1 +/- 5.3 and 12.6 +/- 5.0 mmHg, respectively (NS), while the sphincter lengths were 3.15 +/- 0.81, 3.22 +/- 0.79 and 2.86 +/- 0.85 cm, respectively (NS). In addition, other parameters including lower esophageal body remained unchanged. The serum vasoactive intestinal polypeptide levels of three groups were 24.1 +/- 10.8, 22.5 +/- 9.5 and 21.3 +/- 7.8 pg/ml, respectively (NS). CONCLUSIONS: Neither gastric cancer nor duodenal ulcer in distal stomach removal can alter the lower esophageal body and LES manometric motilities. Unchanged serum VIP levels after gastric surgery are likely one of the mechanisms preserving esophagogastric integrity. PMID- 10522004 TI - Pattern of recurrence after esophageal resection for cancer. AB - BACKGROUND/AIMS: Surgery is still the main treatment option for esophageal cancer; however, long-term survival has remained poor, even when a curative operation is performed. The present study was undertaken to analyze the pattern and time of recurrence after a curative esophagectomy. METHODOLOGY: We studied 53 patients who underwent curative esophageal resection for cancer between 1985 and 1994. We examined number and pattern of recurrences, time after surgery, and any factor with contribution to carcinoma recurrence. RESULTS: During the follow-up period, 34 patients had tumor recurrence. The disease-free interval was 12.7 months (SD = 9.8). Twenty patients (58.9%) developed extrathoracic tumor recurrence and 23 patients (67.6%) intrathoracic. In 3 cases an esophageal stump recurrence was presented. Thirteen patients were considered for palliative treatment after recurrence. The 5-year survival rate was 13%, with median survival time between recurrence and death, 4.1 months. The recurrence of disease was always before 40 months after surgery. Any significant difference related with recurrence was observed between the analyzed factors. CONCLUSIONS: The majority of recurrences are developed before 2 years. Neoplastic recurrence is most common at the mediastinum. Palliative treatments after recurrence do not modify the progression of tumor. PMID- 10522005 TI - Analysis of macroscopic appearance and p53 expression in superficial esophageal carcinoma. AB - BACKGROUND/AIMS: The purpose of the present study was to investigate the relationship between macroscopic appearance of superficial esophageal carcinoma, with particular attention to the horizontal and vertical extent of tumor growth, clinicopathologic findings and p53 expression. METHODOLOGY: Eighty-seven patients with superficial esophageal carcinoma were divided into three groups: 1) group A, patients with protruding or distinct depressed lesions (n = 28); 2) group B, patients with superficial and flat lesions > or = 5 cm in length (n = 45); and, group C, patients with superficial and flat lesions (5 cm in length (n = 14). Tumors were examined immunohistochemically for p53 expression. RESULTS: The incidence of submucosal invasion, lymph node metastasis and lymphatic invasion was significantly higher in group A than in groups B and C. The rate of p53 expression was significantly lower in group B than in the other two groups. The prognosis in groups B and C was better than that in group A. CONCLUSIONS: Vertical extent was more strongly associated with tumor depth, lymph node metastasis and prognosis than was horizontal extent, although p53 overexpression was related to both the vertical and horizontal extent of tumors. Analysis of the macroscopic appearance of superficial esophageal carcinoma is useful in choosing treatment strategies. PMID- 10522006 TI - Congenital esophageal cysts--two cases in adult patients. AB - Esophageal cysts are a rare clinicopathological condition. They usually cause respiratory symptoms in children, while they are often asymptomatic in adults. Two cases of esophageal cysts in adults, recently diagnosed and treated in our department, are reported. In the 1st case (a 52 year-old woman) dysphagia was the main symptom. In the 2nd one (a 39 year-old woman) the patient was asymptomatic. Both were surgically excised by enucleation, with no post-operative complications. The histological study showed both cysts to be lined with ciliated cylindrical epithelium, and they were therefore considered to be congenital. Smooth muscle was only seen in the cyst wall in the 2nd case, but it was not organized in 2 layers, as is typical of duplication cysts. Cartilage or respiratory glands, the pathognomonic features of bronchogenic cysts, were not identified in either of them. Therefore, the diagnosis was inclusion cysts in both cases. PMID- 10522007 TI - A case of complete response to esophageal cancer by a novel concurrent chemoradiation therapy. AB - BACKGROUND/AIMS: A combination of chemotherapy and radiotherapy (chemoradiation therapy) has recently been developed for the treatment of non-operable esophageal cancer patients. Chemoradiation therapy is based on the concept of biochemical modulation effects and radiosensitizing effects of chemotherapeutic agents. However, the optimal choice of chemotherapeutic agents and their doses, as well as chemotherapy and radiotherapy regimens have not been precisely established. METHODOLOGY: Based on our recent experience of an effective chemoradiation therapy protocol which consisted of 5-fluorouracil (5-FU) combined with daily concurrent cisplatin and radiation, in addition to the recent knowledge on the biochemical modulation between 5-FU and cisplatin or between leucovorin and 5-FU, we have developed a complete daily concurrent chemoradiation therapy for non operated esophageal cancer as a treatment modality with curative intent. RESULTS: We report here a novel intensive concurrent chemoradiation protocol by which complete response could be achieved. A low dose of 5-FU (250 mg/body/day) was continuously infused over 24 hours followed by leucovorin (30 mg/body), and a low dose of daily cisplatin (5 mg/body) was administered 1 hour before radiotherapy (2 Gy/day). The administration schedule was 5 consecutive days, followed by a 2 day withdrawal, and then repeated weekly for 4 weeks. CONCLUSIONS: Our completely daily concurrent chemoradiation therapy is rational, effective, and safe, because an endoscopically and pathologically complete response without severe side effects was able to be achieved and this has continued for at least 6 months after chemoradiation therapy. Therefore, we believe that our completely daily concurrent chemoradiation therapy can be recommended for non-operated esophageal cancer patients. PMID- 10522008 TI - Acute reversible dysphagia and dysphonia as initial manifestations of sarcoidosis. AB - A 60 year-old white woman presented with sudden painless dysphagia, hoarseness and dysphonia. A diagnosis of sarcoidosis was made based on bilateral hilar lymphadenopathy, gallium uptake, elevated serum angiotensin-converting enzyme levels, as well as non-caseating granulomatous lymphadenitis in a prescalenic node. Symptoms were attributed to isolated vagus neuropathy, a rare form of presentation of neurosarcoidosis. PMID- 10522009 TI - Submucosal esophageal dissection--a rare case report. AB - The severity of submucosal dissection is intermediate between transmural rupture and mucosal tear in the esophagus. We describe a case of submucosal dissection of the esophagus with characteristic features of mucosal bridge endoscopically and "double-barreled" in radiography. The patient was successfully treated by intermittent esophageal tamponade of 5-day duration using a Sengstaken-Blackmore tube, and total parenteral nutrition. His course was uneventful in a follow-up period of 5 years. PMID- 10522010 TI - Leptin levels in nonalcoholic steatohepatitis and chronic hepatitis C. AB - BACKGROUND/AIMS: Leptin is a peptide which regulates food intake and energy expenditure. Moreover, it is involved in the homeostasis of body composition and is linked to the regulation of insulin signaling, thus playing an important role in liver fat storage. Steatosis is a common finding in chronic hepatitis C, and both viral and metabolic factors have been suggested to explain the presence of this histological characteristic. In order to study leptin in chronic liver disease characterized by the presence of steatosis, we evaluated its serum levels in patients with nonalcoholic steatohepatitis (NASH), in chronic hepatitis C (CHC) patients with no histological findings of steatosis, and CHC patients with steatosis but no risk factors for its development. METHODOLOGY: We studied 6 male patients with NASH whose diagnosis was made on the basis of histological findings and clinical criteria. From among a cohort of 170 CHC patients we put together 2 groups of 6 male patients each (with or without steatosis at liver biopsy examination), who had no risk factors for NASH. Male patients were chosen in order to avoid gender influence on leptin levels. Further criteria for admission were similar impairment of liver metabolic function as assessed by the monoethylglycinexylidide (MEGX) test and, in patients with CHC, similar body mass index (BMI) and histological staging. Moreover, we evaluated leptin/BMI ratio, in order to rule out BMI influence on leptin levels. Leptin levels were assessed by means of radioimmunoassay. RESULTS: We found that BMI was higher in NASH compared to CHC (27.2 +/- 2.9 vs. 23.9 +/- 1.8; p = 0.01). Analysis of serum leptin levels showed an increasing trend starting from patients with CHC without steatosis (3.2 +/- 0.4 ng/ml), through CHC patients with steatosis (4.2 +/- 0.7 ng/ml) up to patients with NASH (5.7 +/- 2 ng/ml), although the differences observed were not statistically significant. Moreover, the ratio of leptin to BMI also followed the same trend showing increasing values (CHC without steatosis = 0.04 +/- 0.02; CHC patients with steatosis = 0.17 +/- 0.03; NASH = 0.203 +/- 0.07). Leptin levels and BMI showed a significant relationship (n = 18; r = 0.60; p < 0.01). CONCLUSIONS: The increasing trend observed in leptin serum levels among the different groups of patients showed that in chronic liver disease characterized by the presence of steatosis, leptin signaling is preserved. Moreover, CHC factors different from the metabolic ones should be investigated in order to explain the presence of steatosis. Further studies on broader groups of patients are needed to verify these preliminary results. PMID- 10522011 TI - Percutaneous microwave coagulation therapy for superficial hepatocellular carcinoma. AB - We report a 67 year-old man with residual hepatocellular carcinoma after arterial embolization therapy, which was located on both the anterior and inferior surfaces of segment 6 of the liver. Percutaneous microwave coagulation therapy could be performed safely and the treated tumor became non-enhancing on contrast computed tomography. Two years after treatment, the tumor remains non-enhancing on contrast computed tomography and has decreased in size. Percutaneous microwave coagulation therapy appears to be useful even in patients who have superficial liver tumors associated with cirrhosis. PMID- 10522012 TI - Resection of liver metastases after pancreatoduodenectomy: report of seven cases. AB - There has been no English report of a long survivor after hepatectomy for metastasis from periampullary malignancies, who had previously undergone pancreatoduodenectomy (PD) for primary disease. Herein, we report 7 patients of liver metastases who underwent 8 hepatectomies after PD for peri-ampullary malignancies. One patient whose liver metastasis was neuroendocrine tumor, survived 2 years and 6 months without recurrence after hepatectomy. Another patient who had 2 hepatectomies for metastasis from duodenal leiomyosarcoma survived for 3 years and 20 days after the first hepatectomy. Procedure of hepatectomy comprised 4 limited resections (including 1 second hepatectomy) and 4 lobectomies at the first resection. Hepatic inflow clamp was used in 6 out of 8 hepatectomies and 4 out of 8 hepatectomies did not require allogeneic blood transfusion. Bacterial contamination of the drained discharge from the cut surface of the liver, mostly representative of enteric organisms, was identified in all but 2 patients who were not examined. Subphrenic abscesses developed in 2 patients after removal of the drains. Thus, prophylactic use of abdominal drain is indispensable after hepatectomy for the patients with bilioenteric anastomosis. PMID- 10522013 TI - Volvulus of the ileum in adult diagnosed pre-operatively by helical 3-dimensioned computed-tomography. AB - A 46 year-old male who was strictly diagnosed as having volvulus of the ileum based on the pre-operative information brought by computed tomography (CT) and helical (spiral) 3-dimensioned computed tomography was surgically treated. The post-operative course was satisfactory and the patient is now under observation without any exacerbation of symptoms one year after surgery. PMID- 10522014 TI - Perforation of jejunal lymphoma--ultrasonographic diagnosis of free air over left flank area. AB - Acute abdomen due to perforation of one of the hollow organs is one of the major challenges for clinicians. Traditionally, pneumoperitoneum shown on X-ray film taken of the decubitus view or in the standing position, is the major key to making a diagnosis of perforation. However, free air is not shown on X-ray film in about one third of cases and sometimes, a standing X-ray cannot be taken in weak patients or for various reasons. In such conditions, abdominal ultrasonography (US) plays a complementary role. Free air is usually detected between the anterior surface of the liver and the anterior abdominal wall by US. However, if free air is not detected on an erect X-ray or not demonstrated over the anterior surface of the liver by US, the diagnosis of perforation of the hollow organ will be difficult. We treated a patient with perforation of a small intestinal lymphoma, which presented as free air over the left flank area by US rather than the anterior surface of liver as is usually the case. Moreover, we located the perforated site pre-operatively by US, which detected focal thickening of a segment of small intestine with intramural slits. Lymphoma of the jejunum with perforation was finally diagnosed after surgery. The value of US is justified in such a condition. PMID- 10522015 TI - Hyperplastic foci as a prognostic factor after resection of hepatocellular carcinoma. AB - BACKGROUND/AIMS: Since chronic liver disease is generally considered a pre malignant condition, recurrence in the residual liver after hepatic resection for hepatocellular carcinoma may be related to the malignant potential of the underlying liver disease. METHODOLOGY: We studied non-cancerous regions of hepatocellular carcinomas that were 3 cm or smaller in diameter from 170 patients who underwent curative hepatic resection. The presence of clusters of hyperplastic small cells, which we called hyperplastic foci, was investigated microscopically. The nuclei of the cells in hyperplastic foci were normal, but the nuclear/cytoplasmic ratio was high, cellularity was increased compared with neighboring regions, and the hepatic trabeculae were somewhat thick. We also calculated the labeling index in hyperplastic foci areas by proliferating cell nuclear antigen and compared this to that of hepatocytes without cellular atypia. RESULTS: In 59 of 170 patients (35%), hyperplastic foci were found. The labeling index in hyperplastic foci was significantly higher than in control regions (p = 0.0016). As of December 1995, 113 patients (63%) were found to have a recurrence. When hyperplastic foci were found, the tumor-free survival rate was significantly lower (p = 0.0443). CONCLUSIONS: Hyperplastic foci represent an important predictor of recurrence after hepatic resection for a small hepatocellular carcinoma. PMID- 10522016 TI - A comparison of hepatocellular carcinoma associated with HBV or HCV infection. AB - BACKGROUND/AIMS: Close relationships between hepatocellular carcinoma (HCC) and hepatitis virus infection have been elucidated. However, clinical differences between HBV- and HCV-associated HCC remain unclear. METHODOLOGY: Four hundred and sixteen patients with HCC were examined for both HBsAg and HCV-Ab. Sixty-nine of the 416 patients (16.6%) were positive for HBsAg and negative for HCV-Ab (B-HCC), while 290 patients (69.7%) were negative for HBsAg and positive for HCV-Ab (C HCC). Various comparisons were made between the 2 groups. RESULTS: B-HCC patients were younger in age (p < 0.0001), and had significantly better liver function than C-HCC patients. The motivation of HCC detection was different between the 2 groups (p < 0.0001), and the tumor size of B-HCC was significantly larger when comparing groups with regard to symptoms (p < 0.05). Although B-HCC demonstrated better survival in Stage I/II treated by surgery (p < 0.05), it was associated with poorer survival in Stage III/IV receiving hepatic arterial infusion chemotherapy when compared to C-HCC (p < 0.01). CONCLUSIONS: These results suggest that clinical differences between B-HCC and C-HCC may depend upon the difference of the natural course between HBV and HCV infection, and B-HCC may be more resistant to treatment than C-HCC in the advanced stage. This also illustrates the need for early tumor detection in B-HCC. PMID- 10522017 TI - Ten year follow-up of patients with chronic hepatitis C treated with interferon. AB - BACKGROUND/AIMS: The impact of the treatment with interferon (IFN) on the natural history of chronic hepatitis C is not defined. The aim of this study was to evaluate the long term effect of the treatment in patients with chronic hepatitis C. METHODOLOGY: In 31 patients with chronic hepatitis C (9 with cirrhosis) consecutively treated with recombinant alpha 2a interferon (r alpha 2a IFN), the evolution of the disease at 10 years from the therapy was evaluated by means of upper endoscopy, liver ultrasonography (US), liver function tests and hepatitis C virus (HCV) viremia. RESULTS: Among 10/31 patients previously classified as responders, only 1 has signs of evolution to cirrhosis; HCV-RNA is still present in 2. Among 21 non-responder patients, 5 developed hepatocarcinoma (HCC) and 4 died during the follow-up; HCV-RNA is present in all the patients still alive. The 6 patients already cirrhotic when treated have clinical signs of progression to Child class B and C. The biochemical, ultrasonographical and endoscopical evaluation shows onset of cirrhosis in 7 of the others. CONCLUSIONS: Patients with chronic hepatitis C who respond to treatment with interferon have good outcome and rare evolution to cirrhosis. The treatment does not seem to influence the natural history of the disease in non-responders. PMID- 10522018 TI - Treatment of spontaneous ruptured hepatocellular carcinoma. AB - BACKGROUND/AIMS: Spontaneous rupture with bleeding is a potentially life threatening complication of hepatocellular carcinoma (HCC). We review our experience with treatments of ruptured HCC. METHODOLOGY: Between January 1988 and December 1997, 18 patients with ruptured HCC were admitted. The patients were divided into 4 groups according to the treatment type of ruptured HCC. Group 1 consisted of 10 patients treated by transarterial embolization (TAE) followed by elective hepatectomy. Group 2 consisted of 2 patients treated by only TAE. Group 3 consisted of 3 patients treated by emergency operation. Group 4 consisted of 3 patients who could not be treated by TAE or surgery. RESULTS: In Group 1, 4 of the 10 patients died; 3 from recurrent HCC and 1 from cerebral hemorrhage, and hospital mortality was absent. The 1-year survival rate was 87.5%. In Group 2, both patients recovered sufficiently well to be discharged. The 1-year survival rate was 50%. In Groups 3 and 4, hospital mortality rate was 100%. CONCLUSIONS: TAE followed by elective hepatectomy was an effective treatment in patients with ruptured HCC. PMID- 10522019 TI - The significance of measuring the serum total bile acids levels during orthotopic liver transplantation. AB - BACKGROUND/AIMS: Bile acid is confined to the enterohepatic circulation and consists of intestinal absorption and hepatic elimination. We investigated whether measuring the serum total bile acids (TBA) levels was useful for both evaluating the function of the grafted liver and predicting the outcome in porcine orthotopic liver transplantation (OLT). METHODOLOGY: Twenty-two female Yorkshire pigs undergoing OLT were divided into 2 groups as follows: Group A consisted of 11 pigs which survived over 7 days with an uneventful early post operative course, while Group B consisted of 11 pigs which died within 5 days due to hepatic failure. The serum TBA levels were measured before and after reperfusion in the recipients. RESULTS: Between Groups A and B, no significant difference was observed in the operative backgrounds including the operation time as well as the cold and warm ischemic time. In Group A, the levels of serum TBA rapidly increased during the anhepatic phase, and thereafter promptly decreased after the reperfusion of the grafted liver. A significant difference was observed in the levels of serum TBA before and after reperfusion (p < 0.01), whereas no significant difference was seen in Group B. The delta TBA, which represents the difference in the levels of serum TBA between just prior to reperfusion and 10 min after reperfusion, was 71.8 +/- 43.5 mumol/L in Group A and 20.1 +/- 34.5 mumol/L in Group B, and demonstrated a significant difference between these 2 groups (p < 0.01). On the other hand, no significant differences were seen in the levels of serum AST, ALT, ALP and TB at each time point between Groups A and B. CONCLUSIONS: The level of serum TBA was found to be a more sensitive parameter and also reflected the developing grafted liver function earlier than the conventional parameters for liver function. Moreover, delta TBA thus appeared to be a valuable predictor for the post-operative outcome. PMID- 10522020 TI - Antioxidant status of erythrocytes from patients with cirrhosis. AB - BACKGROUND/AIMS: The aim of the present study was to investigate possible involvement of oxidant stress in the anemia of cirrhotic patients and to assess blood antioxidant status of these patients. METHODOLOGY: Superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) enzyme activities were measured in the erythroctes from patients with liver cirrhosis and from controls. Levels of thiobarbituric acid reagent substance were also measured in the erythrocyte (TBARSe) and plasma (TBARSp) samples of the groups. RESULTS: Lower activities of SOD and GSH-Px were established in the erythrocytes from patients compared with control subjects. No differences were found between erythrocyte TBARS levels of control and patient groups. Plasma TBARS levels were, however, found to be significantly higher in the patient groups compared with controls. CONCLUSIONS: Results suggest that although enzymatic antioxidant defense system is significantly reduced in the erythrocytes from cirrhotic patients, this does not lead to further peroxidative reactions in the erythrocytes, possible due to preoxidation of some cellular structures sensitive to peroxidative attacks. There was, however, an important indication of accelerated peroxidative reactions in the plasma of the cirrhotic patients, which possibly resulted from extracellular oxidant stress in these patients. PMID- 10522021 TI - Sensitive measurement of serum abnormal prothrombin (PIVKA-II) as a marker of hepatocellular carcinoma. AB - BACKGROUND/AIMS: The usefulness of abnormal prothrombin (PIVKA-II) for diagnosis of small HCC has been limited by its low sensitivity, despite a high specificity. METHODOLOGY: The serum concentration of PIVKA-II was determined by using a new sensitive enzyme immunoassay (EIA) kit in patients with hepatocellular carcinoma (HCC), liver cirrhosis (LC), or chronic hepatitis (CH) and normal controls (NC). alpha-Fetoprotein (AFP) was simultaneously determined in same patients. RESULTS: This kit has made it possible to detect low concentrations of PIVKA-II in the NC. The serum PIVKA-II concentration (mean +/- SE) was 15.7 +/- 1.1 mAu/ml, 16.1 +/- 2.0 mAu/ml, 26.3 +/- 7.2 mAu/ml and 5420.3 +/- 3960.0 mAu/ml in NC, CH, LC and HCC, respectively. Among 106 patients with HCC, 74 patients (69.8%) were positive for PIVKA-II (> or = 40 mAu/ml), while only 9 patients out of 68 patients with LC were positive (13.2%) and only 2 out of 90 patients with CH were positive (2.2%). No significant correlation was observed between AFP and PIVKA-II levels. With combined assay of AFP and PIVKA-II, the positive rate for HCC was increased to 78.3%. Among 14 patients with HCC < 20 mm in diameter. 7 were positive for PIVKA II, and 6 out of 10 patients with HCC between 20 and 30 mm in diameter were positive for PIVKA-II. There was a correlation between tumor size and the PIVKA II level. CONCLUSIONS: Determination of PIVKA-II by this new EIA kit could be useful for the diagnosis of HCC, especially combined with determination of AFP. PMID- 10522022 TI - Risk factors of the recurrence of hepatocellular carcinoma originating from residual cancer cells after hepatectomy. AB - BACKGROUND/AIMS: Little has been documented to differentiate between recurrence originating from microscopic residual tumor cells and recurrence due to metachronous multicentric origin of hepatocellular carcinoma (HCC). The aim of this study was to clarify the risk factors of HCC recurrence closely related to residual tumor cells. METHODOLOGY: A retrospective review of hepatic resections for HCC during the period between April 1985 and April 1997 was undertaken at a University Hospital with a long history of hepatectomy for HCC. Three hundred and thirteen HCC patients without any definite multicentric recurrence, who underwent hepatectomy, were retrospectively investigated. Main outcome measures were: (Study 1) Risk factors for recurrence were univariately and multivariately investigated among various clinicopathological variables, including the vi factor as a new indicator of the potential malignancy of HCC (i.e., the presence of both microscopic portal vein invasion and intrahepatic metastasis). (Study 2). The risk factors for recurrence were then analyzed according to the period of recurrence. RESULTS: (Study 1) Independent risk factors were: (tumor factors) a positive vi factor, alpha-fetoprotein > 100 ng/ml, and poorly differentiated histology; (host factors) albumin < 3.8 g/dl, the presence of diabetes mellitus, platelet count < 14 x 10(4)/microliter, Y-globulin fraction > 20%. In those risk factors, the relative risk of the vi factor (2.6) was the largest. (Study 2) Within 1 year after hepatectomy, only tumor factors, including the vi factor and poorly differentiated histology, were significant risk factors, tumor factors were significant only up to 2 years after hepatectomy, and thereafter only host factors were significant. CONCLUSIONS: The risk factors for non-multicentric recurrence of HCC are considered to be a positive vi factor, alpha-fetoprotein, and poorly differentiated histology, and the vi factor is considered to be a new prognostic indicator expressing the potential malignancy of HCC such as invasion and metastasis. PMID- 10522023 TI - Immunological function and nutritional status in patients with hepatocellular carcinoma. AB - BACKGROUND/AIMS: Infection is a major complication associated with increased morbidity and mortality in patients with hepatocellular carcinoma. We compared the immunological function and nutritional status in 16 patients with hepatocellular carcinoma (13 patients had liver cirrhosis) with those of 21 normal healthy subjects. METHODOLOGY: The immunological function was assessed by chemotaxis and superoxide anion production by neutrophils, phagocytosis and killing activities of neutrophils and monocytes, absolute and relative number of peripheral blood lymphocytes, the percentage of peripheral lymphocyte subsets and serum concentrations of immunoglobulins. RESULTS: Although the phagocytic and bactericidal activities of monocytes and superoxide production of neutrophils were not different between the groups, the phagocytic and bactericidal activities of neutrophils and the percentage of natural killer cells were significantly reduced in patients with hepatocellular carcinoma. In the latter group, the prognostic nutrition index was significantly high compared with normal subjects, indicating a poor nutritional status. The phagocytic and bactericidal activities of neutrophils were low in patients with a poor nutritional status compared to those with a good nutritional status. CONCLUSIONS: Our results suggest that impaired immunological competence and undernourishment may be one of the mechanisms causing increased susceptibility of patients with hepatocellular carcinoma to infection. PMID- 10522024 TI - Extracorporeal bypass using a centrifugal pump during resection of malignant liver tumors. AB - BACKGROUND/AIMS: Total hepatic vascular exclusion (THVE) during extracorporeal bypass is used for hepatic resection in patients with malignant liver tumors. The aim of this study was to determine the efficacy of hepatectomy during total hepatic vascular exclusion using a centrifugal pump (Bio-pump). METHODOLOGY: Fourteen patients with malignant liver tumors who underwent hepatectomy during total hepatic vascular exclusion using the Bio-pump were studied retrospectively. RESULTS: In 3 of 14 patients, insufficient hepatic vascular exclusion was achieved. Six patients underwent tumor resection during total hepatic vascular exclusion, without extracorporeal bypass. In the remaining 5 patients, flow exclusion averaging 1500 ml was achieved with the Bio-pump, and hepatectomy was performed during the procedure. In these 5 patients, the mean operative time and blood loss were 11 hours 38 minutes and 6850 +/- 2451 ml. The Bio-pump bypass time, the excluded blood flow and the mean blood pressure were 82 minutes, 1650 ml and 108/53 mmHg, respectively. The arterial ketone body ratio (AKBR) decreased from a pre-operative value of 1.85-0.32 during total hepatic vascular exclusion. CONCLUSIONS: Total hepatic vascular exclusion was useful for hepatectomy in patients with tumor invasion into the hepatic vein and inferior vena cava, or tumor thrombus in the inferior vena cava and right atrium. However, this technique did not decrease blood loss or improve outcome in patients undergoing hepatectomy. PMID- 10522025 TI - Influence of the extent of hepatectomy on the portal hypertensive state in patients with hepatoma. AB - BACKGROUND/AIMS: Portal hypertensive symptoms, such as esophageal varices and hypersplenism, are frequently observed in patients with hepatocellular carcinoma (HCC). We investigated whether or not the extent of hepatectomy for HCC has an influence on the deterioration of the portal hypertensive state. METHODOLOGY: Fifty-four patients who underwent curative hepatectomy for HCC at our institute were retrospectively studied. The 54 patients were classified in two groups according to the extent of hepatectomy: Group A patients (n = 38) underwent minor hepatectomy (subsegmentectomy or less) and Group B patients (n = 16) underwent major hepatectomy (segmentectomy or more). On the basis of the endoscopic findings for the esophageal varices and the blood platelet counts, the alterations of portal hypertensive state were evaluated before and after hepatectomy. RESULTS: The number of patients whose esophageal varices deteriorated post-operatively, amounted to 9 (23.7%) in Group A and 1 (6.3%) in Group B (not significant). No significant differences were found in the platelet counts between pre- and post-operative states in each Group (A and B). In all of the 6 patients whose esophageal varices first came about after hepatectomy, the advent of the varices occurred more than 1 year after surgery. CONCLUSIONS: These results suggest that in the patients undergoing hepatectomy for HCC, the clinical deterioration of the portal hypertensive state in not caused by the extent of hepatectomy, but by the advance of the coexisting chronic hepatic diseases or tumor recurrence. PMID- 10522026 TI - Hepatocellular carcinoma with tumor thrombi in the bile duct. AB - BACKGROUND/AIMS: Hepatocellular carcinoma (HCC), presenting as obstructive jaundice caused by tumor thrombi in the bile duct, is rare. The authors report on clinical experiences and evaluate the results of different treatment modalities for this disease. METHODOLOGY: We experienced 549 cases of HCC at Ajou University Hospital from June 1994 through January 1998. Among them, 10 cases with gross evidence of tumor thrombi in the bile duct were treated with different resection methods and interventions, and then compared with those receiving short-term results. RESULTS: Eight out of 10 patients underwent exploratory laparotomy: right lobectomy with extrahepatic bile duct resection in 2 cases; right lobectomy with tumor thrombectomy in 2 cases; left lobectomy and caudate lobectomy with extra-hepatic bile duct resection in 2 cases: T-tube drainage in 1 case and biopsy only with post-operative internal biliary stent, in 1 case. Survival times of these patients were 39 months (still alive); 38 months (still alive); 8 months (died); 8 months (died); 8 months (still alive); 1 month (still alive); 14 months (died); 8 months (died), respectively. Of the 2 non-surgical cases, 1 underwent PTBD only and the other had endoscopic removal of the thrombi. Their survival times were 18 days (died) and 24 months (still alive with recurrence), respectively. The 4 cases, with right lobectomy or left lobectomy including extrahepatic bile duct resection, had relatively long-term disease-free survival (39 months, 38 months, 8 months and 1 month after operation, respectively). However, there were no differences in survival between the partial hepatectomy procedure with removal of tumor thrombi and the simple drainage procedure without tumor resection. CONCLUSIONS: Although the number of patients in this study is small, our results suggest that: 1) For the improvement of survival, it seems necessary to perform major hepatic resection with removal of the extrahepatic bile duct. 2) If hepatic resection cannot be accomplished with bile duct resection due to limited liver function, non-surgical modalities should be considered instead of surgery because no differences in prognosis between the 2 groups exist. PMID- 10522027 TI - Hepatocarcinoma: considerations on surgical treatment in a personal series of 23 patients. AB - BACKGROUND/AIMS: Surgical treatment of primary liver tumors has undergone significant changes in recent years because of improved surgical and anesthesiological techniques and better pre- and post-operative care. We review our personal series from 1987-1995. METHODOLOGY: Of 31 cases of hepatocellular carcinoma (HCC) observed in the years 1987-1995, 23 underwent curative resective surgery for a total of 24 liver resections: 6 hepatectomies; 10 segmentectomies; 4 atypical subsegmentectomies; 2 extended resections, with excision of neoplastic thrombi within the portal vein; 1 orthotopic liver transplantation in another institution, and 1 limited segmental resection for tumor recurrence. In 7 recent cases, pre-operative transcatheter arterial chemoembolization (TAE) was used. RESULTS: The mean survival of the 13 patients that are known to be deceased is 27 months (range: 7-114 months). Perioperative mortality was nil. Actuarial 5-year survival rate is 27%. Pre-operative TAE was used in 7 patients: 4 out of 7 lesions were significantly reduced at computed tomography (CT) scan control 21 days following TAE, while in 3 the tumor size was unchanged. CONCLUSIONS: Liver surgery, even major resections, has become safe with no perioperative mortality in our series. In our experience, pre-operative TAE has often produced significant reduction of the mass, but its real efficacy is still the subject of debate. TAE and percutaneous ethanol injection (PET) should be evaluated as part of combined multimodality treatment in the therapy of large lesions previously considered inoperable. PMID- 10522028 TI - Clinical evaluation of hepatic arterial infusion of low dose-CDDP and 5-FU with hyperthermotherapy: a preliminary study for liver metastases from esophageal and gastric cancer. AB - BACKGROUND/AIMS: The prognosis for gastric and esophageal cancer patients with liver metastases remains very poor. In most cases, liver metastasis is unresectable because of its number, size and location and therefore, other approaches need to be considered. METHODOLOGY: In this study we examined 4 patients. We showed the therapeutic benefits of employing hepatic arterial infusion of low-dose CDDP and 5-FU combined with hyperthermia for the treatment of liver metastases of gastric and esophageal cancer. RESULTS: All patients showed partial response, and bone marrow toxicities and gastrointestinal toxicities were extremely slight while liver toxicities were not observed at all. Moreover, 3 of the patients excluding patient 3 who had metastatic lesions other than liver metastases have still been alive for more than 17 months (17-28 months) maintaining a good quality of life. CONCLUSIONS: Therefore, it is suggested that the merits of both low dose-FP and hepatic arterial infusion chemotherapy contribute to ideal clinical effects, and that hyperthermotherapy could enhance clinical responses without potentiating any toxicities. However, this is just a preliminary study, and therefore, a prospective randomized control study is necessary to evaluate the efficiency of this therapy. PMID- 10522029 TI - Caudal left hepatic duct in relation to the umbilical portion of the portal vein. AB - BACKGROUND/AIMS: Intrahepatic ducts from the left lateral segment of the liver are generally considered to run from the cranial to the umbilical portion of the left portal vein. This understanding often limits surgical exploration of the left hepatic ducts during both curative and palliative surgical procedures. The prevalence of ducts from the lateral segment running caudally to the umbilical portion was studied retrospectively on computed tomography (CT) films, and its clinical implications were evaluated. METHODOLOGY: One hundred and sixty-six DIC (drip infusion cholangiography) CT and 30 post-PTCD (percutaneous transhepatic cholangio-drainage) CT films taken during 4 consecutive years were reviewed giving special consideration to whether the ducts from the left lateral segment of the liver run caudal to the umbilical portion of the left portal vein. RESULTS: Corresponding variation was found in 3.6% of the DIC-CT examinations and in 3.3% of the CT examination after PTCD, respectively. CONCLUSIONS: The incidence of caudal left hepatic ducts with respect to the umbilical portion of the left portal vein was higher than expected from clinical experience. These results emphasize that more careful planning is necessary prior to hepatobiliary surgery. PMID- 10522030 TI - Tissue immunodetection of c100 hepatitis C virus antigen in major thalassemic patients. AB - BACKGROUND/AIMS: Hepatitis C Virus (HCV) detection in the livers of chronically infected patients remains a debatable issue. To determine the significance of hepatic expression of hepatitis C viral antigen c100, an immunohistochemical assay was performed in 113 young thalassemics with chronic HCV infection. METHODOLOGY: One hundred and thirteen patients were seropositive for antibody to HCV by second-generation testing. The monoclonal antibody TORDJI-22 was used in an alkaline phosphatase 3-step staining method, and any possible association between the results of HCV immunodetection and various clinicopathologic variables was investigated by univariate and multivariate statistical analysis. In 36 cases, post-therapy liver biopsy specimens were also studied. RESULTS: HCV c100 antigen was detected in 62% of all pretherapy samples, exclusively in the cytoplasm of rather few hepatocytes. Its expression was positively associated with male gender (p = 0.02) as well as with rather advanced age (p = 0.03) and was frequently accompanied by low necroinflammatory scores (according to the modified HAI grading). At the end of interferon-alpha (IFN-alpha) therapy, the immunoreactive prevalence of c100 antigen decreased significantly (pF = 0.002). CONCLUSIONS: We conclude that hepatic expression of c100 antigen is detected in a considerable percentage of thalassemics but it is not likely to provide information concerning the viral load in the infected liver. IFN therapy appears to reduce the hepatic expression of this viral antigen in thalassemic patients. PMID- 10522031 TI - Anastomotic recurrence due to tumor implantation using the double stapling technique. AB - The double stapling technique is indispensable in low anterior resection for colorectal cancer. However, to prevent local recurrences due to the driving of cancer cells into the anastomosis. Intraluminal lavage should be performed. PMID- 10522032 TI - Adrenal metastasis from hepatocellular carcinoma (HCC): report of 3 cases. AB - Although autopsy reports show that the adrenal gland is the second most common organ of hematogeneous metastasis from hepatocellular carcinoma (HCC), paradoxically there is found to be a very scarce number of the adrenal metastasis in clinical practice. We have recently experienced rare patients with right adrenal metastasis from HCC. Case 1: A 51 year-old man with a 5-year history of chronic hepatitis was admitted with hematemesis to Nippon Medical School Hospital. CT revealed a main tumor associated with a few daughter tumors in the hepatic posterior segment and in addition another tumor located between the right hepatic lobe and right kidney. The diagnosis of HCC with a right adrenal gland metastasis was made, and hepatectomy and right adrenalectomy was performed. Twenty months after operation he was alive and free of disease. Case 2: A 78 year old man underwent resection of the lateral segment of the left hepatic lobe for HCC. Twelve months later, recurrent foci in the residual liver were found and those were treated with transarterial embolization (TAE). Right adrenal metastasis was found on CT 26 months after hepatectomy. TAE was done for the hepatic recurrent tumors and adrenal metastasis. Twelve months after, he survived in good condition. Case 3: A 47 year-old man presented with liver cirrhosis with a long history. He was diagnosed as having HCC with multiple intrahepatic metastases and was treated with TAE 4 times. Follow-up CT revealed right adrenal metastasis. TAE was done for hepatic recurrent tumor and right adrenal metastasis. Three months later the patient died of liver failure. PMID- 10522033 TI - Combined interferon, famciclovir and GM-CSF treatment of HBV infection in an individual with periarteritis nodosa. AB - Treatment of chronic hepatitis B virus (HBV) infection in an individual with periarteritis nodosum is described. A combination of famciclovir, granulocyte macrophage colony stimulating factor (GM-CSF) and interferon alpha 2b was utilized. The periarteritis, but not the HBV infection, responded to immunosuppressive therapy consisting of cyclophosphamide and glucocorticoids. Moreover, the patient failed to clear this HBV infection, despite a full year of interferon therapy at 5 MU daily. With the addition of famciclovir and GM-CSF, the HBV infection rapidly resolved and he converted from HBsAg and eAg positive to HBsAb and eAb positive. No exacerbation of his periarteritis nodosum occurred during the course of his antiviral therapy. PMID- 10522034 TI - A case of partial autotransplantation of the liver in advanced hepatocellular carcinoma. AB - Hepatocellular carcinoma in Japan is frequently complicated by chronic hepatic disease such as chronic hepatitis and liver cirrhosis, and it is often impossible to decide the range to be resected only based on clinical stage and other tumor factors. We experienced a case with advanced hepatocellular carcinoma complicated by liver cirrhosis that directly infiltrated into the right and middle hepatic vein. Right trisegmentectomy was performed, the tumor site was extracorporeally removed and the hepatic posterior segment was autotransplanted. An anastomosis of the right hepatic vein and the inferior vena cava was performed with a vascular prosthesis. The patencies of the anastomosed vessels in the vascular reconstructions were confirmed by Doppler sonography, which was very useful, providing an easy and exact evaluation of hepatic blood flow at the patient's bedside. Throughout the post-operative course before the patient's discharge, no abnormal hepatic function was found. Though cases for which partial hepatic autotransplantation is appropriate may be few, this operation procedure, which applies hepatic transplantation techniques, is significant in that it increases the resectability and achieves curative resection of hepatocellular carcinoma. PMID- 10522035 TI - Successful resection of renal cell carcinoma with tumor thrombi extending into the right atrium: a case report. AB - Renal cell carcinoma (RCC) is often associated with an extension of tumor thrombi into the inferior vena cava (IVC) and occasionally up to the right atrium. RCC with IVC involvement has a relatively favorable prognosis when it is completely resected. We present a successfully resected case of RCC with tumor thrombi extending into the right atrium. We performed radical right nephrectomy with lymph node dissection and removed the tumor thrombi en bloc under total hepatic vascular exclusion with the veno-venous bypass between the IVC and the right atrium using an active centrifugal force pump. The patient has been in good condition for 3 years since surgery with no evidence of recurrence. PMID- 10522036 TI - Laparoscopic partial pericystectomy of Echinococcus granulosus cysts in the liver. AB - Laparoscopic partial pericystectomy is a promising new therapeutical approach in surgery of hydatid liver disease. In combination with a review of the published results of laparoscopic therapy for hydatid disease the actual relevance of this technique should be defined. Together with our own experience with this technique we evaluated all patients with hydatid liver disease from Echinococcus granulosus published in literature operated either by pericystectomy or by partial pericystectomy. The review was projected as a search over DIMDI data access. This technique is practicable without increasing the risk of intraabdominal spillage of scolices if well-known security criteria are respected. Additional training is not necessary. Laparoscopic treatment of Echinococcus multilocularis is not possible yet, as complicated liver resections may be required for these patients. Hydatid hepatic cysts of E. granulosus however may be operated upon laparoscopically and do not necessarily require open surgery. While working under visual control minimal invasiveness is achievable and post-operative hospital stay can be reduced. This new technique is a feasible method, especially regarding obese patients, but on the other hand it is limited by a laparoscopically inaccessible intrahepatic localization (Segments IVa, VII, VIII and small centrally located cysts). PMID- 10522037 TI - Post-operative blood tests and multicentric recurrence of hepatocellular carcinoma. AB - Second hepatic resections (SHR) were performed in 2 patients with recurrent hepatocellular carcinoma (HCC) with hepatitis C virus (HCV) more than 10 years after initial curative resections. Appearance on imaging studies of the late recurrences thought to be multicentric primary tumors, was preceded by fluctuating laboratory abnormalities such as increased alanine aminotransferase activity (ALT), increased serum alpha-fetoprotein (AFP), and decreased platelet counts by as long as 2 years. Therefore, serial changes of blood tests after resection of HCC with HCV might be useful predictors of late multicentric recurrence. PMID- 10522038 TI - Primary hepatic carcinoid tumors confirmed with long-term follow-up after resection. AB - Primary carcinoid tumor of the liver is very rare. Only 30 cases have been reported in the English literature. However, most of those cases were diagnosed only on the basis of diagnostic imaging and surgical exploration, their follow-up periods being up to 5 years. Considering the slow progression of the tumor, long term follow-up is required to exclude occult extrahepatic primaries which may manifest afterwards, and to determine the clinical course of this disease. We experienced 3 patients with primary hepatic carcinoid tumors who underwent total resections. They all survived more than 7 years and were clinically confirmed as hepatic primary by failing to detect other primaries during their courses. One patient is disease-free for more than 7 years after resection. The other 2 were found to have recurrence in the remnant liver; 1 underwent transcatheter arterial embolization several times and is alive more than 4 years after recurrence, and the other underwent 2 more operations and survived 8 years after the 1st recurrence. Surgery might offer a possible chance of cure for primary hepatic carcinoid tumor, and transcatheter arterial embolization might be a good treatment option when an unresectable disease is confined to the liver. PMID- 10522039 TI - Palmaz-Schatz stent for hepatic artery stenosis during hepatic arterial infusion chemotherapy. AB - Hepatic arterial infusion chemotherapy using an implantable port system was performed on a 40 year-old man with advanced hepatocellular carcinoma. When the in-dwelling catheter was inserted into the common hepatic artery (CHA), intimal dissection occurred as a result of the catheterization causing severe stenosis. On day 55 after intimal dissection, an in-dwelling Palmaz-Schatz stent was inserted after percutaneous transluminal angioplasty (PTA). CHA blood flow was shown to have improved on Digital subtraction angiography (DSA) and Doppler ultrasound after the in-dwelling Palmaz-Schatz stent. Thus a partial response was shown. The DSA from the implantable port system showed adequate patency 6 months after. This is the first report describing the usefulness of a Palmaz-Schatz stent for the severe stenosis of the CHA caused by the technique of catheterization. PMID- 10522040 TI - Hepatocellular carcinoma developed in a patient with chronic hepatitis C after the disappearance of hepatitis C virus due to interferon therapy. AB - A 62 year-old man was admitted to Asahikawa Medical College Hospital. Injection therapy of natural interferon-alpha was performed against chronic active hepatitis with hepatitis C virus infection. He successfully responded to interferon therapy with normalization of serum transaminases and disappearance of serum hepatitis C virus RNA. The liver function test remained within normal limits and serum hepatitis C virus RNA was not detected throughout the observation period. Three years later, CT examination demonstrated 2 small hepatic masses. Ultrasound-guided biopsy of the hepatic mass demonstrated well differentiated hepatocellular carcinoma histologically. Laparoscopic examination revealed chronic hepatitis, but neither active inflammation nor cirrhotic changes were noted as an underlying liver disease. In the liver specimen, hepatitis C virus RNA was not detected by RT-PCR. Percutaneous ultrasound-guided ethanol injection therapy achieved complete necrosis of the hepatocellular carcinoma and there was no recurrence of hepatic cancer during the follow-up period. This case suggests that patients with chronic hepatitis C infection, who have complete disappearance of serum hepatitis C virus RNA by interferon therapy, should be followed-up carefully for the potential development of hepatocellular carcinoma. PMID- 10522041 TI - Generalized intraperitoneal seeding of hepatocellular carcinoma after microwave coagulation therapy: a case report. AB - We first describe a case of generalized intraperitoneal seeding of hepatocellular carcinoma (HCC) after microwave coagulation therapy (MCT). A 61 year-old man underwent operative MCT for an exophytic HCC, 60 mm in diameter, in segment IV of his cirrhotic liver. Despite successful tumor ablation, the serum alpha fetoprotein levels continuously rose after MCT. Five months later, radiographic examinations delineated several perihepatic masses with hypervascularity, and the patient presented with constipation. At the second laparotomy, there were numerous small peritoneal metastases involving the entire peritoneal cavity and slightly bloody ascites. An omental mass, 50 mm in diameter, involved the transverse colon. Most of these intraabdominal masses were removed together with the involved colon. Histologically, the initial tumor was a moderately differentiated HCC, and the peritoneal masses were poorly differentiated HCCs. The patient died of rapid tumor progression and bleeding 2 months later. In conclusion, we should be aware of the possible occurrence of peritoneal seeding after MCT for HCC. Every effort should be made to prevent this serious complication, particularly in cases of superficial, large, and less differentiated HCCs. PMID- 10522042 TI - Time course of plasma soluble intercellular adhesion molecule-1 (sICAM-1) is related to severity of acute pancreatitis. AB - BACKGROUND/AIMS: In severe acute pancreatitis the release of cytokines indicates a key step from local to systemic inflammation. Increased plasma concentrations of circulating soluble intercellular adhesion molecule-1 (sICAM-1), a marker of leukocyte activation, were detected in necrotizing pancreatitis at the time of diagnosis, however, the exact role of sICAM-1 in the development of complications such as shock or organ dysfunction is unclear. Therefore, we investigated in what manner the time course of plasma sICAM-1 is associated with the development of severe pancreatitis and whether these results are of any predictive value for the further course of the disease. METHODOLOGY: In a medical intensive care unit we studied 29 consecutive patients admitted for acute pancreatitis. Plasma levels of sICAM-1 were measured serially over a period of 6 days and the time courses were assigned either to a group of patients with uncomplicated, mild disease or to patients who developed complications including multiple organ failure. RESULTS: In mild pancreatitis, decreasing and peak sICAM-1 concentrations were found in 88% of the patients with a mean maximal level of 574 +/- 59 ng/ml (SE) (upper limit of normal: 400 ng/ml) on day 1. Partial pancreatic necrosis was present in 24% and no deaths were observed. In severe pancreatitis an increase of sICAM-1 levels or an initial fall followed by an increase (relapsing response) was the predominant pattern (92%). Maximal values of 1453 +/- 136 ng/ml occurred on day 6, significantly different when compared to mild disease (p < 0.0001). Necrotizing pancreatitis was diagnosed in 75% and the mortality rate was 58%. The sensitivity in predicting severe pancreatitis using sICAM-1 plasma levels with an increasing or relapsing pattern was much higher (92%) when compared with serial C reactive protein measurements (42%). CONCLUSIONS: In acute pancreatitis, increasing or relapsing plasma levels of sICAM-1 over 6 days after admission to hospital are associated with a high rate of pancreatic necrosis and a high mortality. Daily measurements of sICAM-1 would allow early recognition of patients prone to develop complications and follow a severe course. PMID- 10522043 TI - Mortality in acute pancreatitis in Turku University Central Hospital 1971-1995. AB - BACKGROUND/AIMS: The role of surgical procedures in the treatment of acute pancreatitis is still unclear. The aim of the present study was to analyze the mortality in acute pancreatitis in Turku University Central Hospital during the last quarter century with special reference to the prevailing surgical treatment trends. METHODOLOGY: A total of 3921 patients with acute pancreatitis were treated 1971-1995. We analyzed the mortality in acute pancreatitis and the number of patients treated in the intensive care unit as well as the number of various surgical procedures used in the treatment of acute pancreatitis in each year 1971 1995. RESULTS: The most conspicuous finding was that the mortality in acute pancreatitis has not decreased any more during the last 15 years. Neither pancreatic resections nor peritoneal lavages seem to decrease the mortality. CONCLUSIONS: We conclude that despite various surgical procedures used in the treatment of acute pancreatitis the mortality in acute pancreatitis has not decreased any more during the last 15 years. Because of the retrospective nature of the current study the present results do not justify drawing any strict conclusions concerning the treatment of acute pancreatitis. However, the present results support the view that conservative treatment in the intensive care unit is justified as an initial therapy even in the fulminant attacks of acute pancreatitis. PMID- 10522044 TI - Ki-ras codon 12 point mutation and p53 mutation in pancreatic diseases. AB - BACKGROUND/AIMS: The Ki-ras gene located at 12p, encodes the GTP binding protein involving the signal transduction system and concerns cell proliferation and differentiation. METHODOLOGY: Pancreatic tissues were obtained from 37 patients with various pancreatic diseases. Ki-ras codon 12 point mutation and p53 (exon 5 8) mutation were examined in 3 patients with chronic pancreatitis, 9 mucinous adenoma of the pancreas (2 with mucinous cystadenoma and 7 with intraductal papillary-mucinous adenoma), 22 pancreatic ductal carcinoma, and 3 serous cystadenoma. RESULTS: On usual pancreatic exocrine ductal lesions, Ki-ras point mutation was evident in 0% (0/3) of chronic pancreatitis, in 56% (5/9) of mucinous adenoma, and in 57% (12/21) of ductal carcinoma, the mutation being located in the second letter in 18 and in the 1st letter in 2. One Ki-ras codon 12 positive pancreatic cancer showed Ki-ras codon 12 point mutation in the surrounding pancreas (2nd letter mutation in both areas). p53 mutation was present in 0% (0/1) of chronic pancreatitis, in 0% (0/8) of mucinous adenoma, while it was evident in 29% (6/21) of pancreatic ductal carcinoma, the mutation being situated in exon 5 in 3, in exon 6 in 1, and in exon 7 in 2. In 3 patients with serous cystadenoma, there was no mutation in Ki-ras codon 12 or p53 (exon 5 8). CONCLUSIONS: These findings suggest that Ki-ras point mutation is involved in the early events of pancreatic ductal carcinoma, while p53 mutation is intricated in the late phase of pancreatic ductal carcinogenesis and the histogenesis of serous cystadenoma is different from that of pancreatic exocrine ductal lesions including mucinous adenoma and ductal carcinoma. PMID- 10522045 TI - Portal hypertension as a complication of chronic pancreatitis. AB - BACKGROUND/AIMS: Intractable abdominal pain, duodenal stenosis and common bile duct stenosis are considered the main reasons for surgery in cases of chronic pancreatitis. The aim of the study was to discover the influence of the disease on the portal pressure. METHODOLOGY: Blood pressure was measured in the superior mesentric vein before and after resection of the head of the pancreas in 17 patients by direct method. RESULTS: Venous pressure was lower after resection of the head of the pancreas in all 17 measured patients. CONCLUSIONS: Chronic pancreatitis increases not only left-sided portal pressure but also right-sided portal pressure. PMID- 10522046 TI - Segmental pancreatectomy for mucin-producing pancreatic tumors. AB - BACKGROUND/AIMS: Segmental pancreatectomy for benign tumors of the neck of the pancreas was reported in 1993. Mucin-producing carcinomas are generally regarded as low-grade malignancies as compared with ductal cell carcinomas of the pancreas. We report herein our experience with a segmental pancreatectomy for mucin-producing pancreatic tumors. METHODOLOGY: Three patients with mucin producing tumors of the pancreatic body underwent a segmental pancreatectomy. After the pancreatic tumor had been located with intra-operative ultrasonography (US), the medial pancreas centered on the tumor was resected. The margin of the retained pancreas was submitted for histopathologic inspection intra-operatively to prevent retained disease. A conduit for draining the pancreatic juice consisted of a jejunal Roux-en-Y loop between the left and cephalic portions of the pancreas. Histologically, the 3 tumors were identified as a cystadenocarcinoma, an intraductal papillary adenocarcinoma, and a cystadenoma with a focus of borderline malignancy. The functional result was evaluated with oral glucose tolerance and pancreatic function diagnostic (PFD) testing. Pancreatic juice drainage was confirmed using magnetic resonance cholangiopancreatography (MRCP). RESULTS: Neither technical failure nor operative death occurred in any of the patients. The patients have been followed-up for between 33 months and 77 months after surgery and all are disease free. The oral glucose tolerance test and PFD test results were all within the normal range. MRCP showed good pancreatic juice drainage in the 2 patients examined. CONCLUSIONS: Segmental pancreatectomy may be an appropriate surgical procedure for mucin-producing pancreatic tumors, to prolong survival and to preserve endocrine and exocrine function. PMID- 10522047 TI - Is age a risk factor for major pancreatic surgery? An analysis of 300 resections. AB - BACKGROUND/AIMS: The aim of this study was to analyze if age alone is a risk factor in major pancreatic surgery. METHODOLOGY: From September 1, 1985 to December 31, 1997, 806 patients underwent surgery for malignant and benign diseases of the pancreas in a prospective case control study performed at the Department of Surgery, Johannes Gutenberg University Hospital Mainz. In 228 patients (men: n = 139; women: n = 89; mean age: 61 years; range: 23-83 years) we performed partial (n = 178) or total (n = 50) pancreaticoduodenectomy, which was combined with portal vein resection in 16 cases. Left pancreatic resection was carried out in 72 patients (men: n = 40; women: n = 32; mean age: 65 years; range: 28-86 years). RESULTS: Surgical complications after pancreaticoduodenectomy occurred in 22.1% of patients < or = 70 years and in 30.2% of patients > 70 years, however, less than half of them had severe complications ranging below 50%. General complications developed in 16.1% of patients < or = 70 years and in 27.9% of patients > 70 years (p < 0.001). The mortality rates 30 and 90 days after surgery were 3.2% (< or = 70 years) and 2.3% (> 70 years), and 6.0% (< 70 years) and 6.9% (> 70 years), respectively. Regression analysis showed the following factors to exert an independent influence on mortality: Pre-operative serum bilirubin, the diameter of the pancreatic duct, intra-operative blood loss and the occurrence of surgical and nonsurgical complications. Age did not exert an independent influence on the prognosis of either morbidity or mortality. However, general complications developed significantly more often in elderly patients. After left pancreatic resection surgical complications developed in 29.3% (< or = 70 years) and 21.4% (> 70 years) of patients, however the rate of severe complications was below 10%. General complications occurred in 10.3% (< or = 70 years) and 28.6% (> 70 years) (p < 0.001). Mortality rates 30 and 90 days after operation were 1.7% (< or = 70 years) and 14.2% (> 70 years), and 3.4% (< or = 70 years) and 14.2% (> 70 years) (p = n.s.), respectively. Regression analysis showed the intra-operative blood loss to exert an independent influence on post-operative morbidity and mortality. Age had no independent influence on either morbidity or mortality. CONCLUSIONS: Results obtained by this study show that, although general complications develop significantly more often in elderly patients, age is not an independent risk factor for post-operative mortality after major pancreatic resection. Factors of importance in improving the outcome of this operation include the experience of the surgeon in selecting patients eligible to undergo the procedure, his operative skills in performing major pancreatic resections, as well as better anticipation and management of post-operative complications. PMID- 10522048 TI - Immunohistochemical analysis of expression of molecular biologic factors in intraductal papillary-mucinous tumors of pancreas--diagnostic and biologic significance. AB - BACKGROUND/AIMS: In this study we investigated the expressions of molecular biologic factors, p53, rasp21, bcl-2, c-erbB-2, and Ki-67 by immunohistochemical method in intraductal papillary-mucinous tumor of the pancreas to identify their diagnostic values and to determine their relations to the degree of histopathologic abnormalities. METHODOLOGY: Thirty-eight different histologic lesions from 28 patients of intraductal papillary-mucinous tumor of the pancreas, comprising normal pancreatic duct (n = 6), intraductal papillary hyperplasia (n = 6), intraductal adenoma (n = 15), and intraductal carcinoma (n = 11) were immunostained by the avidin-biotin peroxidase conjugate method. RESULTS: p53 and Ki-67 expressions were significantly greater in malignant intraductal papillary mucinous tumor than in their benign counterpart (p = < 0.0001), while rasp21 showed gradual increase in the frequency of expression from normal pancreatic duct (0%), to intraductal hyperplasia (16.7%), to intraductal adenoma (26.7%), and ultimately to intraductal carcinoma (63.6%). bcl-2 and c-erbB-2 were not expressed in any lesions. CONCLUSIONS: These results suggest that p53 and Ki-67 expressions have significant diagnostic values in differentiating benign intraductal papillary-mucinous tumors from malignant ones and thus can facilitate in the pre-operative planning of treatment in individual cases. Secondly, gradual stepwise increase in the frequency of rasp21 expression with increasing degree of cellular atypia supports the presence of adenoma-carcinoma sequence in the carcinogenesis of this tumor. PMID- 10522049 TI - Surgical palliation in pancreatic head carcinoma and gastric cancer: the role of laparoscopy. AB - BACKGROUND/AIMS: In all patients with pancreatic and gastric cancer we always make a laparoscopic exploration to complete the staging. Lately we have adopted the following technique for nonresectable cancers of the head of the pancreas: following endoscopic retrograde cholangiography we position a biliary stent to restore bile flow and obtain regression of jaundice, a laparoscopic-assisted gastroentero-anastomosis (GEA) is then performed as an antecolic isoperistaltic side-to-side gastrojejunostomy. Also in case of nonresectable gastric cancer we perform a laparoscopic-assisted gastrojejunostomy. METHODOLOGY: From January 1994 February 1998 we performed a total of 25 laparoscopic assisted gastrojejunostomies. We adopted this minimally invasive technique for 11 out of 20 patients (55%) with nonresectable cancers of the head of the pancreas, 7 men and 4 women, whose median age was 73 (range: 60-89). A video-assisted gastrojejunostomy was also performed in 14 patients out of 28 (50%), 10 men and 3 women, with a median age of 70 (range: 58-76), with nonresectable distal gastric cancers and 1 woman with non-resectable and obstructing duodenal cancer. The operative time of the video-assisted procedure was 35 min (range: 25-40 min). RESULTS: There were no intra-operative complications and no mortality. All the patients had a very satisfactory post-operative course, with only 1 (4%) with post-operative complications (hyperpyrexia in a patient due to an infection of the biliaryendoprosthesis, with precocious regression after replacement of the prosthesis) and minimal post-operative pain. Median post-operative hospital stay was 3 days (range: 2-4). Median survival after operation was 6 months (range: 2 12) for gastric cancer and 9 months (range: 5-15 months) for pancreatic head carcinoma. CONCLUSIONS: We believe that this technique, for the obstructive syndrome of the pylorus and duodenum, offers these patients the best results/trauma ratio. Two currently remaining types of indications for a GEA, namely non-malignant ulcer and unresectable duodenal or antropyloric obstructive cancer. PMID- 10522050 TI - Experience with intraarterial infusion of styrene maleic acid neocarzinostatin (SMANCS)-lipiodol in pancreatic cancer. AB - A 54 year-old man was admitted to our hospital and diagnosed with inoperable cancer in the body and tail of the pancreas. The spleen was embolized at its hilum with a coil to infuse an anti-tumor agent selectively into the pancreatic parenchyma and transcatheter intraarterial infusion (TAI) of styrene maleic acid neocarzinostatin (SMANCS)-Lipiodol, 3 mg, was performed. The computed tomography (CT) scan taken immediately after TAI revealed the incorporation of SMANCS Lipiodol into the site of the pancreatic tail. At 2 weeks, a small amount of SMANCS-Lipiodol remained and clearness of the tumor margin was lacking in the pancreatic tail, but no remarkable change was noted in the body. As for the laboratory data, pancreatic enzyme level was not elevated immediately after TAI. At 2 weeks, tumor markers showed improvement in CEA (3.9-->2.6 ng/ml) and Elastase 1 (370-->230 ng/ml), but little change was seen in CA 19-9 (1600 U/ml: no change) and DUPAN-2 (730-->740 U/ml). In pancreatic cancer, SMANCS-Lipiodol could be infused from the splenic artery into the pancreatic parenchyma by the splenic arterial embolization. PMID- 10522051 TI - Intra-pancreatic rupture of the bile duct. AB - Injury of the bile duct after blunt trauma is rare but injury and rupture of the intrapancreatic portion of the bile duct is extremely rare. Injury is very difficult to recognize even if explorative laparotomy is done. Elevation of the liver tests, especially gamma GT and alkaline phosphatase, is very often the first sign of injury. When jaundice occurs the diagnostic procedures of ultrasound (US), computed tomography (CT) and endoscopic retrograde cholangiopancreatography (ERCP) must be done. If there is no sign of great injury to the head of the pancreas we have to perform normal biliary flow. Here we report a rare case of intrapancreatic rupture of the common bile duct without high degree of pancreatic injury after blunt injury. Cholecystectomy and choledochojejunostomy were performed. PMID- 10522053 TI - Abdominal aortic aneurysm compression is probably responsible for the recurrent episodes of acute pancreatitis: case report. AB - An aged male with a known history of abdominal aortic aneurysm suffered from epigastralgia, vomiting and cold sweating for one day. According to the physical examination, serum amylase level and computed tomographic examination, acute pancreatitis was diagnosed. Surgical intervention for the abdominal aortic aneurysm was not performed because of his age, and finally this patient died after three recurrent episodes. Acute pancreatitis co-existing with an intact abdominal aortic aneurysm has never been reported before. The possible pathogenesis of this recurrent acute pancreatitis was discussed. PMID- 10522052 TI - A case of pancreatic cancer achieving symptomatic improvement with systemic chemotherapy. AB - We present a case of pancreatic cancer demonstrating symptomatic improvement with systemic chemotherapy using 5-fluorouracil and cisplatin (FP therapy). A 43 year old man had pancreatic cancer with para-aortic lymph node metastases. He received FP therapy and achieved a partial response. After the initiation of chemotherapy, his symptoms such as severe pain and fatigue improved remarkably. Serum interleukin-6 levels correlated with these symptoms; the level was high before chemotherapy, but the levels were decreased during the courses of FP therapy. It is important to achieve symptomatic improvement in patients with pancreatic cancer, and serum interleukin-6 levels may be useful to evaluate a symptomatic response to chemotherapy. PMID- 10522054 TI - Portal vein resection without reconstruction during Appleby operation in a patient with pancreatic body carcinoma with cavernous transformation. AB - The prognosis of pancreatic body carcinoma has been poor due to cancerous invasion of major vessels. Resection of the involved vessels may improve resectability and prognosis. We report a patient who had a pancreatic body carcinoma with cavernous transformation of the portal vein, in whom the portal vein was resected without reconstruction during an Appleby operation. A 67 year old man was admitted for evaluation of back pain. Enhanced computed tomography showed no main trunk of the portal vein but a developed collateral circulation. Celiac angiography revealed encasement of the common hepatic, splenic and celiac artery. Venous angiography revealed obstruction of the portal and splenic veins with cavernous transformation surrounding these veins. Pre-operative diagnosis was carcinoma in the pancreatic body, which invaded the portal vein, the celiac and common hepatic arteries. The Appleby operation combined with resection of the portal vein without reconstruction could be performed, by preserving collateral vessels and monitoring hepatic venous oxygen saturation (ShvO2) to prevent hepatic ischemia caused by occlusion of the portal vein. The post-operative course was uneventful. PMID- 10522055 TI - Laparoscopic splenectomy for splenic artery aneurysm. AB - Two cases undergoing a laparoscopic splenectomy for the treatment of a splenic artery aneurysm are herein reported. This lesion is relatively rare. Surgical treatment is indicated for such cases since approximately 10% of these aneurysms tend to rupture which thus results in fatal hemorrhaging. Both cases demonstrated aneurysms measuring more than 2 cm in diameter based on the ultrasonography, computed tomography and celiac angiography findings and, as a result, a laparoscopic splenectomy was thus prophylactically performed. This procedure is the preferred technique for high risk patients, such as those with chronic renal failure, as observed in case 1, since patients can be spared the disadvantages of undergoing a laparotomy. PMID- 10522056 TI - Significance of para-aortic lymph node dissection in advanced gastric cancer. AB - BACKGROUND/AIMS: Since surgical results in advanced gastric cancer remain poor and para-aortic lymph node dissection may contribute to survival, it is useful to determine the significance of para-aortic lymph node dissection. METHODOLOGY: Para-aortic lymph node dissection was provisionally indicated for patients with invasion depth deeper than the subserosal layer. Clinicopathologic variables were retrospectively analyzed using univariate analysis and multivariate analysis to predict para-aortic lymph node metastasis. Similarly, they were analyzed using univariate analysis and the Cox's proportional hazards regression model to estimate the prognostic factor in 120 patients who underwent para-aortic lymph node dissection. Surgical results and post-operative complications were compared between para-aortic lymph node dissection and D2 dissection. RESULTS: Univariate analysis revealed that the mean diameter, the degree of lymph node metastasis, and the invasion depth were significant predictors of para-aortic lymph node metastasis. Multivariate analysis showed that n2 was the only independent predictive factor as to para-aortic lymph node metastasis. Univariate analysis revealed tumor site, tumor diameter, lymph node metastasis, number of positive lymph nodes, INF, and stage were significantly associated with 5-year survival. The Cox's proportional hazards regression model showed that the number of positive lymph nodes and the number of positive para-aortic lymph nodes were independent prognostic factors. Patients with < or = 10 positive lymph nodes in any stage or < or = 3 positive para-aortic lymph nodes in stage IVb had significantly better surgical results. Surgical results for patients who underwent para-aortic lymph node dissection with n2 or invasion depth deeper than the exposed serosa were significantly higher than those in D2. As to post operative complications, pancreatic fistula and respiratory complications were significantly frequent after para-aortic lymph node dissection. CONCLUSIONS: n2 is helpful in predicting para-aortic lymph node metastasis. Whereas, post operative morbidity such as pancreatic fistula and respiratory complications after para-aortic lymph node dissection were significantly higher, they were controllable. Para-aortic lymph node dissection should be indicated in advanced gastric cancer patients in which lymph node metastasis is over n2 or invasion depth is deeper than the exposed serosa. But the number of positive para-aortic lymph nodes must be less than three. PMID- 10522057 TI - Surgical therapy of gastric stump carcinoma--a retrospective analysis of 109 patients. AB - BACKGROUND/AIMS: Although gastric stump carcinoma has been described as early as 1922, knowledge regarding best treatment is still insufficient. Therefore, we analyzed our results of the surgical therapy of gastric stump carcinoma. METHODOLOGY: Between May 1968 and November 1996, 109 patients were operated upon because of gastric stump carcinoma, and the data of these cases were retrospectively analyzed. Survival rates were calculated with the Kaplan-Meier method (Log-rank-test; p < 0.05). RESULTS: A distal Billroth II gastrectomy was the most frequent type of prior operation in 95.4% of the patients. Resectability was 67% (n = 73), and in 64 cases total gastrectomy with systematic lymphadenectomy was performed. Overall post-operative morbidity and mortality were 33.9% and 13.8% respectively. These figures were significantly reduced to 13.8% and 2.8% in the last decade. The 5-year survival rate after radical resection was 40.7%, and prognosis was influenced by R-classification and tumor stage. CONCLUSIONS: Improvements of surgical technique and intensive care management enable resections of gastric stump carcinoma with a low peri-operative morbidity and mortality. Total gastrectomy with systematic lymphadenectomy should be the goal of surgical therapy to obtain a curative resection. Long-term prognosis is similar to that of primary gastric carcinomas. PMID- 10522058 TI - Mutated p53 protein expression and proliferative activity in advanced gastric cancer. AB - BACKGROUND/AIMS: When the DNA of cells is damaged, wild-type p53 protein induces the expression of p21 (Waf1/Cip1/Sdi1), and regulates the progression of the cell cycle by inducing G1 arrest. Thus, wild type p53 or p21 protein negatively regulates cancer cell proliferation. However, in tumor with loss of expression of functional wild-type p53 protein by mutation or allelic deletion of the gene for p53, whether the proliferative activity of cancer cells might be accelerated or not is unclear. In this study, we investigated the correlation between the level of expression of mutated p53 protein and the proliferative activity of cancer cells in advanced gastric cancer. METHODOLOGY: Ninety-seven samples from patients with gastric cancer that had invaded the serosa without lymph node metastasis (t3, n0, stage II) were investigated by immunohistochemical staining with a monoclonal antibody against p53 and against p21, and with the monoclonal antibody Ki-67. DNA ploidy patterns were analyzed by flow cytometry. The immunoreactivity against p53 and the proliferative activity of cancer cells were scored in terms of a labeling index (LI; percentage of immunostained cells) in each case. Moreover, the prognostic values for 93 surviving patients were evaluated by univariate and multivariate analysis. RESULTS: The mean p53 LI was 24% (range: 0 82.4%) and the mean Ki-67 LI was 23.1% (range: 0-70.7%) in 97 tumors. The expression of p21 protein was detected in 30 of 97 tumors (30.9%) and DNA aneuploidy was detected in 36 of 97 tumors (37.1%). There was significant correlation between the p53 LI and the Ki-67 LI (r = 0.61, t = 7.456, p < 0.001) in 97 tumors. Although, no significant difference was detected, the mean p53 LI (18.3%) of 30 tumors with expression of p21 protein was lower than that of 67 tumors without expression of p21 protein (26.6%, p = 0.096). However, no significant correlation between expression of p21 protein and Ki-67 LI was observed. The p53 LI was not an independent prognostic factor in 93 surviving patients by multivariate survival analysis (p = 0.069). However, the 5-year survival rate of 50 patients with a low level of p53 LI (p53 LI (< or = 10%, 78.3%) was significantly better than that of 43 patients with a high level of p53 LI (p53 LI > 10%, 62.1%, p = 0.045). CONCLUSIONS: Accumulation of mutated p53 protein might suppress the expression of p21 protein in gastric adenocarcinoma, and cancer cells with overexpression of mutated p53 protein might have a high proliferative activity. PMID- 10522059 TI - Gastric exclusion for unresectable gastric cancer. AB - BACKGROUND/AIMS: Conventional gastrojejunostomy is sometimes performed for unresectable gastric cancer, but it is not fully effective. To improve the patient's quality of life, we performed gastric exclusion. METHODOLOGY: Twenty seven patients who received gastrojejunostomy (11 conventional, 16 gastric exclusion) were retrospectively examined as to post-operative quality of life and outcome. RESULTS: No stomal strictures were observed, and gastrointestinal bleeding was significantly reduced in the gastric exclusion group. These advantages enabled the gastric exclusion group to achieve better quality of life, as indicated by longer oral intake (244 days vs. 98 days) and home stay (211 days vs. 91 days). The prognosis also improved. The 50% survival period in the gastric exclusion group was 229 days, whereas, that of the conventional gastrojejunostomy group was 131 days. CONCLUSIONS: The quality of life and prognosis of the gastric exclusion group significantly improved, and we believe that the improvement of the quality of life yielded a better prognosis. We recommend gastric exclusion as a standard procedure for unresectable gastric cancer. PMID- 10522060 TI - Perigastric lymph nodes with metastasis in gastric cancer. AB - BACKGROUND/AIMS: We analyzed the significance of metastasis to the subdivided perigastric lymph node stations according to the distance from the primary gastric cancer, and the appropriateness of the recent change in the Union Internacional Contra la Cancrum (UICC) tumor node metastasis (TNM) system. METHODOLOGY: Gastrectomy was performed in 753 patients with gastric cancer. The perigastric lymph nodes were divided into 6 stations according to the Japanese classification. These were subdivided into 2 categories according to the distance from the primary tumor: -1, nodes within 3 cm of the edge of the tumor; and -2, nodes more than 3 cm from the edge of the tumor. Survival rates were calculated with the Kaplan-Meier method, and the difference between each group was evaluated by the log-rank method. RESULTS: The frequency of metastasis to the subdivided perigastric lymph node stations, numbered 1-1 to 6-2, varied between 10.0% and 41.1%. The 5-year survival rate of the patients with positive 6-1 lymph node was higher than that of the patients with positive 6-2 lymph node (31.5% and 17.5%, P = 0.0032). There were no statistically significant differences in survival between subgroups of patients who had metastatic lymph node in the other 5 stations. The frequency of metastasis to other regional lymph nodes in patients with N2 perigastric lymph nodes was higher than that in patients with N1 perigastric lymph nodes. CONCLUSIONS: Subdivision of the perigastric lymph nodes had little advantage. Elimination of the old system of classifying perigastric lymph nodes according to distance from the tumor is appropriate. PMID- 10522061 TI - A multi-institutional prospective study of lentinan in advanced gastric cancer patients with unresectable and recurrent diseases: effect on prolongation of survival and improvement of quality of life. Kanagawa Lentinan Research Group. AB - BACKGROUND/AIMS: Lentinan is one of the host-mediated anti-cancer drugs which has been shown to affect host defense immune systems. Although the mechanisms involved in the antitumor effects of lentinan have been reported experimentally, the clinical outcome on prolongation of survival and improvement of quality of life in gastric cancer patients with unresectable or recurrent diseases has yet to be clarified. The aim of the present study was to investigate whether administration of lentinan prolonged survival or improved quality of life in these patients. METHODOLOGY: A multi-institutional randomized prospective protocol, consisting of patients administered tegafur and cisplatin (control group), and patients administered lentinan, tegafur and cisplatin (lentinan group), was performed. Quality of life was investigated using a questionnaire survey. RESULTS: Median survival was significantly longer in the lentinan group than in the control group (297 days vs. 199 days, p = 0.028). One-year survival rate was greater in the lentinan group than in the control group (49.1% vs. 0%). Total QOL score, especially appetite and sleep quality, was significantly improved with the administration of lentinan. CONCLUSIONS: Lentinan is considered to prolong survival and improved quality of life when gastric cancer patients with unresectable or recurrent diseases are treated in combination with other chemotherapeutic agents. PMID- 10522062 TI - Prognostic factors of advanced gastric carcinoma without serosal invasion (pT2 gastric carcinoma). AB - BACKGROUND/AIMS: The purpose of this study was to investigate the prognostic factors of advanced gastric carcinoma without serosal invasion (pT2 gastric carcinoma), for the planning of therapeutic strategy. METHODOLOGY: Prognostic factors were evaluated by univariate and multivariate analysis in a total of 304 curatively resected pT2 gastric carcinoma patients in whom the tumor invaded the muscularis propria or the subserosa. RESULTS: Macroscopic type, depth of invasion, lymph node metastasis and venous invasion were significantly related to outcome, using univariate analysis. Lesions resembling early gastric carcinoma had better prognosis than lesions belonging to one of the Borrmann types. Multivariate analysis (Cox's proportional hazards model) demonstrated that macroscopic type, lymph node metastasis and venous invasion, but not depth of invasion, were significant prognostic factors. CONCLUSIONS: Macroscopic appearance, lymph node metastasis and venous invasion were the important prognostic factors of pT2 gastric carcinoma. Extensive lymph node dissection and aggressive post-operative chemotherapy should be performed, especially in Borrmann type lesions with lymph node metastasis. PMID- 10522063 TI - An experimental study of intramural blood supply network of the stomach wall. AB - BACKGROUND/AIMS: A fundamental experiment was undertaken with reference to local resection with lymphadenectomy for gastric cancer. The intramural blood supply network of the stomach wall, about which no reports have previously appeared, was surgically investigated. METHODOLOGY: Five pigs were used. The left gastric and the right gastroepiploic vessels and their branches were removed from all the animals. The right gastric vessels were also removed, while the short gastric and the left gastroepiploic vessels were preserved in the 1st pig. Only the short gastric vessels were preserved in the 2nd, and the right and the short gastric vessels in the 3rd. After resection of the whole stomach with the spleen, angiography was performed through the preserved arteries. The 4th and 5th pigs underwent the same procedures as the 1st, with additional local resection of the stomach in the 5th. Both pigs were maintained for 1 year and then, angiography was performed. RESULTS: When the short gastric and the left gastroepiploic vessels, or the short and the right gastric vessels were preserved, the whole stomach continued to receive blood supply through the intramural network. Stomach wall resection therefore can be performed with no complications after these procedures. CONCLUSIONS: The present results confirm the possibility of local resection with lymphadenectomy as a treatment for gastric cancer. PMID- 10522064 TI - Efficacy of extended lymphadenectomy in the noncurative gastrectomy for advanced gastric cancer. AB - BACKGROUND/AIMS: We retrospectively analyzed clinicopathologic data on 83 patients with advanced gastric cancer who underwent noncurative gastrectomy, with respect to the relation between the extent of lymphadenectomy and survival benefit. METHODOLOGY: These 83 patients were divided into 44 patients with limited or simple lymph node dissection (D0 in 14 and D1 in 30: Group A) and 39 patients with extended lymph node dissection (D2: Group B). RESULTS: The 1-year survival rate in Group B (82.1%) was significantly higher than in Group A (49.0%). However, the 3-year and 5-year survival rates did not significantly differ between Group A versus Group B, 39.7% versus 25.7% and 39.7% versus 20.5%, respectively. Median survival time after surgery with and without distant metastasis in Group B (21.5 months) was longer than in Group A (16.4 months), although not significant. CONCLUSIONS: While gastrectomy with extended lymphadenectomy did not contribute to improve long-term survival in patients with noncurable advanced gastric cancer, the utility of extended lymph node dissections may be relevant to improved locoregional control, at least in the prognosis within 1 year after surgery. Not only extended lymphadenectomy but also aggressive chemotherapy may be needed to improve the long-term survival for such patients. PMID- 10522065 TI - Expression of facilitative glucose transporters in gastric tumors. AB - BACKGROUND/AIMS: Although cancer cells are known to have an increased rate of glucose metabolism, the complete mechanism for increased glucose uptake in tumor cells is unknown. METHODOLOGY: The presence of mRNA for 5 facilitative glucose transporter (GLUT) isoforms was evaluated by reverse transcription-polymerase chain reaction (RT-PCR) in paired samples of normal gastric mucosa and gastric tumor from 20 individuals. Expression of GLUT proteins was immunohistochemically determined in 70 resected gastric cancer specimens. RESULTS: By using RT-PCR, GLUT2 mRNA was detected in 80% of normal gastric mucosal samples, while GLUT4 mRNA was seen in only 40%, GLUT1 mRNA was not detected in normal gastric mucosa. In gastric carcinoma samples, GLUT1 mRNA was detected in 19 out of 20 cases (95%) and GLUT2, GLUT3, and GLUT4 mRNAs in all samples. By immunohistochemistry, GLUT1 protein was detected in 19% of the tumors. A majority of tumors (61%) expressed 1 or more transporter protein. The presence of GLUT1 protein in a tumor was positively correlated with the tumor's invasion into the gastric wall, lymphatics or blood vessels and with lymph node metastases. The post-operative survival of patients with tumor expressing GLUT1 protein was significantly worse than those with tumor without GLUT1 protein. CONCLUSIONS: Gastric cancer cells may acquire the ability to produce GLUT1 mRNA by malignant transformation. Increased expression of the high-affinity glucose transporters, GLUT1 and/or GLUT4, in tumor cells may drain glucose preferentially to the tumor at the expense of the tumor-bearing host. PMID- 10522066 TI - Poorer prognosis in young patients with gastric cancer? AB - BACKGROUND/AIMS: Although the relationship between prognosis and age of patients with gastric cancer is controversial, a poorer prognosis in young patients has been suggested by most investigators. To further examine the hypothesis, a retrospective study was undertaken to analyze a large series of patients with gastric cancer in Taiwan. METHODOLOGY: A total of 1,642 consecutive patients diagnosed with gastric cancer and receiving further management at one medical center from 1988 to 1993 were reviewed. The gender, TNM tumor stage, rate of curative resection and survival of the patients were compared in the young age group (< or = 39 years) and the old age group (> 39 years). Survival was estimated with the product-limit method and difference in survival was tested by the log-rank test. Multivariate analysis was done by the Cox proportional hazard model. RESULTS: Among the 1,642 patients, 61 patients were in the young age group and 1,581 patients were in the old age group. There was no significant difference for the 2 groups of patients in the distribution of TNM stage (stage I: 20%; II: 8%; III: 13%; IV: 59% vs. 19%, 11%, 25% and 45% respectively, in the old age group, p = 0.098) and rate of curative resection (38% vs 51% in the old age group, p = 0.059). The overall 5-year survival showed no significant difference between the 2 groups (25% vs. 29% in the old). Subgroup analyses showed that survival after curative resection and survival without curative treatment (including palliative resection and no resection) also had no difference in the 2 groups. Multivariate analysis also showed that age was not an independent factor. CONCLUSIONS: Although most reports suggested a dismal prognosis in young patients with gastric cancer, based on our findings, young patients (< or = 39 years) do not have a worse prognosis than older patients. PMID- 10522067 TI - Lymph node involvement rate in low-grade gastric mucosa-associated lymphoid tissue lymphoma--too high to be neglected. AB - BACKGROUND/AIMS: Anti-Helicobacter pylori (H. pylori) treatment for low-grade gastric mucosa-associated lymphoid tissue (MALT) lymphoma has been the subject of attention. The aim of this study was to determine the proportion of such cases which could be suitable candidates for H. pylori eradication for the purpose of cure; we focused on gross morphology and lymph node metastasis. METHODOLOGY: We retrospectively reviewed the medical records of 53 patients diagnosed and treated for gastric MALT lymphoma at Seoul National University Hospital between 1992 and 1996. RESULTS: According to Isaacson's classification, 60% of cases were low grade, and H. pylori was detected in 88% of them. In low-grade disease, gastroscopy revealed superficial lesions in 56% of cases, ulcerofungating lesions were found in as much as 19%, and ulceroinfiltrating in 25%. Even in low-grade disease, invasion of proper muscle, or deeper, was seen in 28% of patients, and lymph node involvement in 36%; even in low-grade disease confined to mucosa and submucosa, the rate of lymph node involvement was 40%. All cases which, on gastroscopy, appeared to be gastritis or benign ulcer-like lesions were free of lymph node metastasis, but in low-grade disease, this proportion was only 16%. In 33% of cases, pre-operative clinical stage I--as shown by abdominal CT--was found post-operatively to be stage II. The negative predictive value of lymph node detection by CT was 68%. CONCLUSIONS: In low-grade gastric MALT lymphoma, the lymph node involvement rate was too high to be neglected. In detecting lymph node metastasis, the diagnostic accuracy of CT was too low. The proportion of suitable candidates for anti-H. pylori treatment for low-grade gastric MALT lymphoma was not high, and in clinical practice, anti-H. pylori treatment in such cases should at present be very carefully applied. PMID- 10522068 TI - Pre-operative angiography in gastric cancer surgery with extended lymphadenectomy. AB - BACKGROUND/AIMS: The value of pre-operative angiographic evaluation in patients undergoing gastric cancer surgery with extended lymphadenectomy was assessed in a prospective study comparing exposed and unexposed groups of patients. METHODOLOGY: During the period from July 1991 to October 1997, 76 patients (Group A--exposed) were pre-operatively submitted to a digital subtraction angiography (DSA) after informed consent. Concurrently, 94 patients (Group B--unexposed) were included as an unexposed reference group. All patients underwent total or subtotal gastrectomy with D2 lymphadenectomy according to the guidelines proposed by the Japanese Research Society for Gastric Cancer (JRSGC). RESULTS: In 34 (45%) exposed patients (Group A), DSA detected an atypical vascular anatomy. Major anatomical variations of the celiac axis, its branches and the superior mesenteric artery were discovered in 4 subjects (5%). Vascular anomalies affecting the surgical tactics of lymphadenectomy were detected in less than 8% of patients. Five post-operative deaths (6.6%) were registered between patients of the Group A, exposed to pre-operative angiography, 8 in the unexposed Group B (8.5%). Post-operative morbidity was significantly higher (P = 0.038) in the Group B (34%) in comparison to Group A (20%) but no difference in risk of individual complications was detected. CONCLUSIONS: Although useful in the presence of major vascular anomalies, it appears that pre-operative angiography did not significantly reduce intra- and post-operative complications associated with radical gastrectomy combined with extended lymphadenectomy. Arteriography is therefore not routinely recommendable but its use is mandatory in specific operations for gastric cancer. PMID- 10522069 TI - Gastric cyto-secretory correlations in peptic ulcer. AB - BACKGROUND/AIMS: Maximal acid output, parietal cell mass, serum pepsinogen A (PGA) and total peptic activity (TPA) in gastric juice were studied and compared in duodenal ulcer and in different gastric ulcer sites. METHODOLOGY: 152 peptic ulcer patients were studied. 64 cases of gastric ulcer (GU) were subdivided according to Johnsons's classification and compared with 88 duodenal ulcer (DU) patients diagnosed for the first time. 40 normal subjects were studied as controls. RESULTS: Duodenal ulcer is characterized by normo-hyperparietalism, normo-hyperchloridria and an increase in peptic activity. In cases of GU, such correlation is not only conditioned by the topographic seat of the ulcer, but by the histological condition of the gastric mucosa too. Body GU is characterized by hypoparietalism, hypochloridria, hyper-PGA and hyper-TPA. Pre-pyloric GU is characterized by normo-hyperparietalism, normo-hyperchloridria, hyper-PGA and hyper-TPA. In GU the cyto-secretory behavior is characterized by the histology of the body mucosa with prevalence of preatrophic-atrophic gastritis in case of body GU and prevalence of superficial gastritis in case of GU type II and III. CONCLUSIONS: The results confirm the anatomic-functional analogy between DU and type II and III GU. If considered from the functional point of view, these conditions differ considerably from those that are characteristic of type I GU (as they closely follow the chronic gastritis pattern). PMID- 10522070 TI - The effect of intragastric acidity on Helicobacter pylori eradication with bismuth-metronidazole-amoxicillin. AB - BACKGROUND/AIMS: Adding an acid secretion inhibitor to anti-H. pylori regimens may be potentially valuable for enhancing the effectiveness of antimicrobials that exhibit markedly reduced activity at low pH. This study was conducted to evaluate intragastric acidity as a factor in H. pylori eradication with bismuth based triple therapy. METHODOLOGY: Forty patients with duodenal ulcer and H. pylori infection were included. The patients were divided into 2 groups--normacid (n = 20) and hyperacid (n = 20)--based on the amount of time that 24-hour intragastric pH took to reach the level pH > or = 3. All patients received bismuth subsalicylate (600 mg 3 times daily), metronidazole (500 mg 3 times daily) and amoxicillin (500 mg 3 times daily) for 2 weeks. Then, all patients continued treatment with ranitidine (150 mg twice daily) for 8 weeks prior to the follow-up examination. Blood samples were collected before treatment for measurement of fasting gastrin and pepsinogen-I. RESULTS: Nine patients (45%) in the normacid group and 8 patients (40%) in the hyperacid group reported side effects. However, there were only 2 patients (10%) in each group who withdrew from the study due to intolerance of side-effects. There was no difference in the H. pylori eradication rate between the normacid and hyperacid groups (16/18, 88.9% vs. 15/18, 83.3%). CONCLUSIONS: Without co-administration of anti-secretary agents, intragastric acid is not a significant factor in the effectiveness of H. pylori eradication with bismuth-based triple therapy. PMID- 10522072 TI - Management of erectile dysfunction in general practice. PMID- 10522071 TI - Intra-abdominal gastric volvulus. An indication for gastropexy through laparoscopy. AB - Intra-abdominal gastric volvulus is a chronic or acute condition that may be cured by conventional surgery through laparotomy. We report a case of subacute intra-abdominal volvulus who benefited from a gastropexy through laparoscopy. We show that this technique is feasible, safe and efficient. Technical details are described to perform the procedure easily and rapidly. PMID- 10522073 TI - Childsafe Ireland--can we do it? PMID- 10522074 TI - Management of acne vulgaris. PMID- 10522075 TI - How reliable is memory recall? PMID- 10522076 TI - Restless leg syndrome. PMID- 10522077 TI - Increasing incidence of ectopic pregnancy: is it iatrogenic? AB - The incidence of ectopic pregnancy at the National Maternity hospital in 1996 was double that of the previous year. The rise in incidence coincided with the introduction at the hospital of both serum quantitative bHCG (beta human chorionic gonadotrophin) testing and a clear protocol for the early diagnosis of ectopic pregnancy. The aim of this study was to determine whether the increased incidence in 1996 was due to an increase in pre-existing risk factors for the development of ectopic pregnancy or to increased diagnosis of the condition. For the years 1986, 1995 and 1996, the incidence of ectopic pregnancy at the National Maternity Hospital was 1.8, 4.8 and 8.3 respectively per thousand pregnancies. There was no significant difference between the three years in terms of maternal age, parity or risks factors for ectopic pregnancy. The median gestational age at diagnosis decreased from 8 weeks gestation in 1995 to 6 weeks in 1996 (p < 0.001). However, the incidence of ruptured ectopic and blood transfusion was similar in both years. This study suggests that the two-fold increase noted in the incidence of ectopic pregnancy is related to earlier diagnosis of the condition. This is aided by the availability of serum quantitative bHCG testing. These findings have important implications for the management of women with ectopic pregnancy. PMID- 10522078 TI - The diagnostic value, parental and patient acceptability of micturating cysto urethrography in children. AB - OBJECTIVES: To evaluate the medical indications and outcome including psychological and physical consequences of micturating cysto urethrography. METHODS: A prospective study of 165 consecutive children undergoing MCUG during a 3 month period. Medical data, including outcome was recorded. The distress of the child was recorded by the radiographer at the time of the examinations. Postal questionnaires were sent to parents one week after the test to obtain information on their own and their child's perception of the test, and any physical and/or behavioural changes. RESULTS: Age distribution for first and subsequent MCUG. [table: see text] 73% of first MCUG's were requested because of urinary tract infection. 52% of first MCUG's in infants were abnormal compared with 13% in older children aged 1 to 4 years. 29 children aged 4 and over underwent a second or subsequent MCUG (53% abnormal). These children would be suitable for indirect cystography. One quarter of children experienced difficulty in passing urine following the test, haematuria was experienced by four. Radiographers recorded severe distress in 27% of children and 27% of parents also recorded distress. CONCLUSIONS: A high incidence of distress was detected for both parent and child. Units should establish special guidelines for the use of this invasive procedure. A clear explanation of the investigation to parents and children should be standard practice. Routine employment of sedation for patients may be advisable. Alternative methods of imaging should be considered and evaluated, and indirect isotope cystography employed where appropriate. PMID- 10522080 TI - Lyme disease and glomerulonephritis. PMID- 10522079 TI - Premixed insulin preparations and glycaemic control in type 1 diabetes mellitus. AB - The role of premixed insulin preparations in Type 1 DM remains unresolved. The degree of glycaemic control achieved with the use of premixed insulin preparations in an unselected group of subjects with Type 1 DM has not previously been reported. We abstracted and reviewed data on 600 subjects with Type 1 diabetes mellitus in our computer data base. 134 (23%) were taking two injections of premixed insulin daily. In these subjects glycaemic control as assessed by the most recent HbA1c measurement was 7.6 +/- 0.1% vs 7.4 +/- 0.1% in those subjects on separate insulins, p = N.S. In those subjects aged < 35 years, 62/220 (28%) used premixed insulin. HbA1c was higher in those on premixed insulin 7.8 +/- 0.2% and 7.2 +/- 0.1%, p < 0.05. This difference in HbA1c was seen only in those subjects with diabetes duration of 3 to 8 years (n = 78) where HbA1c was higher in those subjects on premixed insulin, 8.4 +/- 0.5% versus 6.9 +/- 0.2%, p < 0.01. In subjects aged > 35 years, 74/318 (23%) used premixed insulin. HbA1c did not differ between those using premixed insulin and those using separate insulin preparations, 7.5 +/- 0.2% and 7.5 +/- 0.1% respectively. We conclude that use of premixed insulin preparations achieved a similar level of glycaemic control as use of individual insulin preparations with the exception of patients aged less than 35 years who were 3 to 8 years following diagnosis. The relatively good glycaemic control achieved may make these preparations a useful alternative to multiple injection therapy for many patients with Type 1 DM. PMID- 10522081 TI - An unusual soft tissue mass: extraosseous fracture of a metallic implant. PMID- 10522082 TI - Stevens-Johnson syndrome after exposure to a pesticide in a patient with AIDS. PMID- 10522083 TI - DOs need more training in diagnosing and treating addiction. PMID- 10522084 TI - Perceptions and reported practices of osteopathic physicians in diagnosing and treating addiction. AB - The objective of this study was to assess the perceptions and reported practices of osteopathic physicians in the diagnosis and treatment of addiction. Copies of survey questions were sent to the 344 members of the West Virginia Osteopathic Society. A total of 176 (51.2%) physicians responded; of these responses, 166 surveys were used for analysis. Respondents included 130 practicing physicians and 36 physicians in internship or residency training programs. Of those responding, 133 were men and 33 were women, and ages ranged from 24 to 81 years with a mean of 41.6 years. Respondents who were graduates of the West Virginia School of Osteopathic Medicine numbered 132 (79.5%), and 99 (59.6%) were in family practice. Characteristics most commonly attributed to addiction were a chronic nature and psychological or physical dependence. More than half of the test subjects did not consider addiction to be a primary disease independent of other factors or psychiatric conditions. Respondents reported a mean addiction prevalence of 20.4%, with the most common substances reported as tobacco, alcohol, and benzodiazapines, respectively. Individual prevalence reports varied from 0% to 95% (SD +/- 20.4%). The most commonly used diagnostic tools were the CAGE (Cut down, Annoyed, Guilty, and Eye-opener) test, DSM III-R (Diagnostic and Statistical Manual of Mental Disorders, 3rd edition, revised) or DSM-IV (Diagnostic and Statistical Manual of Mental Disorders, 4th edition) criteria, and quantity and frequency questions. Medical sequelae such as jaundice or emphysema were the most likely reasons for the respondents to address a substance abuse problem. For referral resources, respondents were most likely to use inpatient or outpatient treatment. A mean success rate of 27.7% was reported by the 133 physicians responding. The wide variance in reported prevalence and the low success rate reported in comparison to that demonstrated in published treatment studies indicate that there is a need for further education of both physicians in training and those presently in practice. Medical sequelae are frequently irreversible signs of late-stage addiction, and physicians should be urged to include such tools as the CAGE test in each regular physical to facilitate earlier intervention. PMID- 10522085 TI - A review of alternative birthing positions. AB - This article reviews a number of nonevidenced-based studies that have been conducted on the different physical positions of labor and delivery. A review of the literature disclosed that the traditional supine position appeared to be associated with a prolonged second stage of labor and persistence of occiput posterior presentations. However, the supine and left lateral positions are excellent for providing anesthesia and access, although there may be a little added benefit for the parturient's comfort. The sitting, squatting, and hands-and knees positions offer superior patient participation. The squatting position held for a long time may be physically stressful. The sitting and standing positions are known for occasional excessive hemorrhage and added expense. The hands-and knees position offers the advantages of the gravitation effect of the upright positions and may be associated with less perineal damage. Overall, there has been no report of any harm to the infant when alternative positions are used. PMID- 10522086 TI - ["Load and go" or "stay and play"? Measures at the accident site must be adapted according to the needs of the injured]. PMID- 10522088 TI - [A discovery that can result in new therapies. The enzyme telomerase is an important factor in the development of cancer]. PMID- 10522087 TI - [A new antibacterial agent can stop resistance development. Expectations are connected to linezolid, an oxazolidinone with a new pharmacological action]. PMID- 10522089 TI - [Future tasks of physicians in charge of school health. Support to student with problems instead of health control]. PMID- 10522090 TI - [The physician should guide the patient to avoid the worst therapeutic pitfalls]. PMID- 10522091 TI - [Quackery or physicians--a question of reliance]. PMID- 10522092 TI - [The wandering myths about vaccination against whooping cough]. PMID- 10522093 TI - [Ehrlichiosis is common among animals but can also occur in humans]. AB - Granulocytic ehrlichiosis is attracting increasing attention as a disease of both humans and animals. Pasture fever in cattle is a form long known in Sweden, and a similar disease was observed in Swedish dogs and horses in the late 1980s. The first cases of human granulocytic ehrlichiosis (HGE) reported from the USA in 1994 proved to have been caused by a species of Erhlichia identical or very similar to the species found in Swedish dogs and horses. Presence of the HGE agent has now been demonstrated in Ixodes ricinus ticks in Sweden. Serological studies among residents in areas of Sweden with a high incidence of ticks have shown the presence of antibodies against the HGE agent in over 10 per cent of cases. To date, there have been about 10 documented cases of HGE in Sweden, a figure to be compared with those for dogs and horses among which ehrlichiosis is relatively common. PMID- 10522094 TI - [Three Swedish cases of African tick-bite fever. Can our native Rickettsia species cause disease in humans?]. AB - The article consists in a report of three cases of African tick-bite fever in Swedish tourists returning from brief visits to South Africa. The clinical course included eschar, regional lymphadenopathy, fever and, in two cases, maculopapular rash. Two cases were characterised by significant increases in anti-Rickettsia conorii IgG and IgM antibody titres. However, the aetiological agent was assumed to be Rickettsia africae, based on reports by others and the widespread serological cross-reactivity among spotted fever Rickettsia spp. The third case was diagnosed on clinical grounds. During the past ten years, 50 per cent (41/80) of cases diagnosed serologically as rickettsial (R. conorii antigen) spotted fever at the Swedish Institute for Infectious Disease Control were associated with travel to South Africa. Parallels are drawn to the recent finding of R. helvetica in Swedish ticks (Ixodes ricinus), and the possibility of its pathogenicity to humans is discussed, though no such clinical cases have been reported to date. PMID- 10522095 TI - [Aggravated uncontrolled hemorrhage induced by intravenous fluid administration]. AB - A model of uncontrolled haemorrhage where a 0.5 mm laceration is made in the porcine abdominal aorta has shown outcome to be impaired by conventional fluid therapy given to restore blood volume. Findings in recent studies where the difference in blood flow rates, proximal vs. distal to the site of vascular lesion, was used as a measure of bleeding suggest the adverse effect of fluid therapy to be strongly associated with re-bleeding after primary haemostasis has occurred. Optimal survival is dependent on a fluid infusion rate ensuring balance between the risk of re-bleeding and the beneficial effects of fluid therapy on oxygen consumption. PMID- 10522096 TI - [More precise indications for fluid therapy during transportation to hospital are required]. AB - Although early intravenous fluid therapy for haemorrhage and shock is usually given before arrival at the hospital, its value is unclear and more precise indications are needed. The indications will take into account such factors as transport time, volume and type of bleeding, and the presence or absence of concomitant head injury. Fluid resuscitation can be omitted if transport time is less than 30 min, but may be beneficial if it is more than 30 min. Choice of infusion rate should be guided by the estimated risk of re-bleeding when haemorrhage is uncontrolled, and by cerebral perfusion where severe head injury is present. PMID- 10522097 TI - [Should patients with cystic fibrosis become parents? Patient's own choice following matter-of-fact information is significant]. PMID- 10522098 TI - [Medical research and outlook on mankind: studies on the experience of disease--a task for phenomenology]. PMID- 10522099 TI - [A trauma course for medical students prepares physicians for emergency service]. PMID- 10522100 TI - [Influenza vaccine caused problems in joints. Compensation from the drug insurance authority]. PMID- 10522101 TI - [Health economics analysis of diabetes is necessary. It facilitates decision making and international comparison]. AB - Cost-of-illness studies have shown diabetes to be associated with substantial direct and indirect costs, accounting for 5-6 percent of total health care expenditure. In a Swedish study, where total costs were divided into costs due to management of diabetes and costs due to complications, the total annual cost to the community was estimated to be SEK 5.7 billion in 1994, costs due to complications being the major item, accounting for over 75 per cent of the total. There have been few other Swedish studies of costs for diabetes or diabetes related complications. The most widely studied category of complications is diabetes-related foot ulcers, with an estimated annual cost of SEK 1-2 billion. However, earlier studies were marred by shortcomings: costs estimated for the main diagnosis only, without breakdown into categories or distinction between type 1 and type 2 diabetes, sources of data other than official data-bases ignored, etc. Diabetes care in Sweden is of high quality, and substantial clinical, epidemiological and health economics research has been carried out. It is important that Sweden contributes to international research on health economics aspects of diabetes. PMID- 10522102 TI - [Commercialization of genes--breast cancer gene patent is a pilot case. Sweden must play a part in the development]. PMID- 10522103 TI - [Obstetrics and gynecology in Sweden during a century]. PMID- 10522104 TI - [Goals of epilepsy treatment--a multidimensional model. The description of goals as a basis for efficiency measurement]. AB - Review of relevant literature suggests at least seven distinct goals of epilepsy treatment to exist: to reduce seizure frequency and severity, to improve function, to enhance quality of life, to promote coping, to improve external circumstances, to prevent premature death, and (in children) to promote growth and development. However, these goals are subject to certain qualifications. For example, seizure frequency and severity is not to be reduced under all conditions, but only when such reduction can be expected to have beneficial effects on quality of life or on life expectancy (the two ultimate goals). The different qualifications suggest that the proposed goals can not, and should not be considered in isolation, but incorporated in a multidimensional model. PMID- 10522105 TI - [There is time to defuse the issue of sedation in terminal care]. PMID- 10522106 TI - [The hand-arm vibration syndrome: (I) the clinical picture, exposure-response relationship and exposure limits]. AB - Part I of this paper presents an overview of the medical aspects of the hand-arm vibration syndrome, as well as the relationship between occupational exposure to hand-transmitted vibration and the vascular, neurological, and musculoskeletal disorders occurring in the upper limbs of workers who use vibrating tools. There is epidemiologic evidence for an increased occurrence of peripheral sensorineural and vascular disorders in occupational groups using a great variety of vibrating tools. An excess risk for wrist osteoarthrosis and elbow arthrosis and osteophytosis has been reported in workers exposed to shocks and low frequency vibration of high magnitude from percussive tools. To date, the available epidemiologic data are insufficient to outline an exposure-response relationship for both sensorineural disturbances and bone and joint disorders caused by hand transmitted vibration. The association between vibration white finger (VWF) and exposure to hand-transmitted vibration has been clearly established in both cross sectional and longitudinal studies of vibration-exposed workers. A proposal of exposure-response relationship for VWF is included in an annex to ISO 5349 (1986). However, the shape of the relationship between vibration exposure and VWF is not yet fully understood. The results of several epidemiologic studies seem to indicate that the current ISO frequency-weighting may be inappropriate for all types of vibration and for all kinds of vibration injury. Alternative exposure response relationship for VWF have been suggested in recent epidemiologic investigations. Regarding exposure limits for hand-transmitted vibration, the findings of clinical and epidemiologic studies have shown that the vibration exposure levels proposed by the European Directive for physical agents are sufficiently protective for the safety and health of workers exposed to hand transmitted vibration. PMID- 10522107 TI - [A system for the active surveillance of occupational bronchial asthma: the results of 2 years of activity of the PRiOR program]. AB - A surveillance programme of work-related hazards and diseases (PRiOR) was developed in the Piedmont region of Italy during 1996-97, including an active surveillance system for asthma. Incidence estimation, early information to the patient, identification of responsible workplaces were the main objectives of the programme. A network of 18 clinical centers (allergologists, chest physicians, occupational physicians) from whole the Piedmont region provides information on newly diagnosed cases to a surveillance center that receives and analyzes reports and disseminates surveillance results to clinicians and occupational health and safety services, responsible for intervention. Between 1 March 1996 and 31 December 1997 67 new cases were reported, which corresponds to an incidence rate of 24 (C.I. 95%: 18-30) per million workers per year. The incidence was higher among bakers (540 per million), among workers employed in the leather and shoe industry (214 per million), in health care services (205 per million), in the pharmaceutical industry (199 per million) and among hairdressers (196 per million). The agents to which workers were exposed at the time of diagnosis were generally well recognised (latex 51%; flour 27%). Procedures and results of the PRiOR programme are comparable with those from surveillance systems operating in other countries. Nevertheless, continued efforts are needed to increase the proportion of collaborating physicians, to standardize the use of tests for diagnosis and to improve the communication between clinical centers and occupational health and safety services. Like most surveillance systems based on similar methods, PRiOR underestimates the true incidence of work-related asthma in the Piedmont Region. Despite this limitation, the PRiOR active surveillance system of work-related asthma has proven successful in identifying more new cases of asthma than official sources do. It can therefore be permanently implemented in the prevention system regional. PMID- 10522108 TI - Measurement of surface contamination by certain antineoplastic drugs using high performance liquid chromatography: applications in occupational hygiene investigations in hospital environments. AB - Within the context of continuing interest in occupational hygiene of hospitals as workplaces, the authors report the results of a preliminary study on surface contamination by certain antineoplastic drugs (ANDs), recently performed in eight cancer departments of two large general hospitals in Milan, Italy. Since reliable quantitative information on the exposure levels to individual drugs is mandatory to establish a strong interpretative framework for correctly assessing the health risks associated with manipulation of ANDs and rationally advise intervention priorities for exposure abatement, two automated analytical methods were set up using reverse-phase high-performance liquid chromatography for the measurement of contamination by 1) methotrexate (MTX) and 2) the three most important nucleoside analogue antineoplastic drugs (5-fluorouracil 5FU, Cytarabin CYA, Gemcytabin GCA) on surfaces such as those of preparation hoods and work-benches in the pharmacies of cancer wards. The methods are characterized by short analysis time (7 min) under isocratic conditions, by the use of a mobile phase with a minimal content of organic solvent and by high sensitivity, adequate to detect surface contamination in the 5-10 micrograms/m2 range. To exemplify the performance of the analytical methods in the assessment of contamination levels from the target analyte ANDs, data are reported on the contamination levels measured on various surfaces (such as on handles, floor surfaces and window panes, even far from the preparation hood). Analyte concentrations corresponding to 0.8-1.5 micrograms of 5FU were measured on telephones, 0.85-28 micrograms/m2 of CYA were measured on tables, 1.2-1150 micrograms/m2 of GCA on furniture and floors. Spillage fractions between 1-5% of the used ANDs (daily use 5FU 7-13 g; CYA 0.1-7.1 g; GCA 0.2-5 g) were measured on the disposable polythene-backed paper cover sheet of the preparation hood. PMID- 10522109 TI - [Work accidents in Brazil. A study in Sao Paulo state, Brazil, the Botucatu region, using the causal tree method]. AB - This paper discusses, within the prevailing Brazilian situation, the possibility of applying "causal tree" (CT) method in investigating occupational accidents by safety personnel in the public health services and workers' unions. The method was developed during the seventies in France, for use by plant safety personnel. The authors used this method in Botucatu, state of Sao Paulo, Brazil, in order to investigate 40 serious occupational accidents that occurred in industrial plants during the second half of 1993, that had been registered by social security. In these cases, the predominance of situations in which the lack of safety measures were identified by inspection indicates that in most instances, the use of CT is unnecessary. However, the authors discuss its use by safety personnel from the public health services and workers' unions to investigate certain accidents to contribute to the knowledge base and help overcome the cultural based guilt which, in Brazil, has turned the victim into the person responsible for the accident. PMID- 10522110 TI - Occupational health and safety in Morocco: present and future. AB - Occupational health and safety in Morocco remain the poor link in our health system despite the existence since several decades of regulations concerning the protection of workers. This legislation is interesting but unfortunately not implemented and not updated. Our study shows failures at all levels: three occupational medical inspectorates with nine occupational inspection physicians for the whole of Morocco; 1,322 occupational medical services for 4,600 firms required to have such services. Occupational medical services cover only 7% of the urban working population; more than 9 workers out of 10 do not benefit from any medical protection. Only one occupational medical service out of four submits its annual medical report to the occupational medical inspectorate; 683 physicians practice occupational medicine while our theoretical needs are for about 3,000; among the 300 doctors holding a diploma of occupational medicine, only 100 practice in their speciality; of the 1,200 nurses employed in work environments, few hold state diplomas as provided for by legislation; safety engineers, prevention experts and ergonomists are rare; several exposed sectors do not have occupational safety and health services: civil servants, handicraft workers, small firms, rural areas, temporary and occasional workers, etc.; no serious study on occupational hazards (occupational accidents and diseases) has been undertaken. A reorganization of occupational safety and health is required: at the level of the Council of physicians and occupational medical inspectorate; a commission should control "who does what"; at the national level: extension of occupational safety and health services to all the working population (political will); meeting of the Consultative Medical Council and the establishment of a National Institute of Occupational Health; at the international level: the fight against the introduction of dangerous substances and technologies, originating in industrialized countries. Only correct and generalized occupational safety and medicine can ensure a true health protection of the population, particularly those working. PMID- 10522111 TI - Improved respiratory CO2-He analysis for peripheral airways impairment detection. Utilization possibilities of a new computerized system in preventive medicine studies. AB - The authors illustrate the application possibilities in occupational and preventive medicine of a recently computerized system for the evaluation of peripheral ventilatory non-homogeneities with or without alveolar air trapping. The method consists of an improved individual multiple breath by breath test utilizing the analysis of the respiratory gases CO2 and He. It is based on the results of previous experimental measurements performed simultaneously in different pulmonary sectors an depends on the technical possibility of signalizing out-of-phase expired CO2-He mixtures at the end of the inert gas clearance when in subjects with peripheral respiratory disorders the two gases CO2 and He arrive at the mouth at different times, during the same expiration and cause different, opposite signals. The applied physiological reliability of the measurements was experimentally verified, their technical specificity was recently established. The use of a new computerized system allows actually automatic, on-line measurements under the control of the operator, following a standardized method. The resolution power of the signals is very high. The calculation of the results is performed by the software. The system is easy to transport and can be used in the field for screening on large groups of persons in preventive medicine inquiries and during occupational medicine checks. Quite recently it has been usefully employed during an investigation on 200 subjects, occupationally exposed (policemen of the city traffic department of the town of Bologna). PMID- 10522112 TI - [Letters to the editor]. PMID- 10522114 TI - [In Process Citation] PMID- 10522113 TI - [A clinical case of possible screen dermatitis]. PMID- 10522116 TI - Call for an international ban on asbestos. PMID- 10522115 TI - [The biocidal capacity of polyphenols and their use in the hospital environment]. PMID- 10522117 TI - An object-oriented framework for the development of computer-based guideline implementations. AB - Clinical practice guidelines provide a means of directing medical care towards clinically appropriate and cost-effective interventions. A direct relationship exists between the integration of a guideline into clinical workflow and the effectiveness of the guideline in influencing clinicians' behavior. Computer based guideline implementations, used at the point-of-care, accomplish this integration. Employing object-oriented technologies, we propose a framework of reusable components for the development of guideline implementation systems. We have identified eight information management services that are common to such systems. Our framework integrates these services and their respective reusable components into clinical workflow to promote the development of comprehensive guideline implementation systems, which should ultimately enhance guideline compliance and the overall quality of care. PMID- 10522118 TI - An electronic study form to support collaborating agents in the management of clinical knowledge. AB - When dealing with biological organisms, one has to take into account some peculiarities which significantly affect the representation of knowledge about them. These are complemented by the limitations in the representation of propositional knowledge, i.e. the majority of clinical knowledge, by artificial agents. Thus, the opportunities to automate the management of clinical knowledge are widely restricted to closed contexts and to procedural knowledge. Therefore, in dynamic and complex real-world settings such as health care provision to HIV infected patients human and artificial agents must collaborate in order to optimize the time/quality antinomy of services provided. If applied to the implementation level, the overall requirement ensues that the language used to model clinical contexts should be both human- and machine-interpretable. The eXtensible Markup Language (XML), which is used to develop an electronic study form, is evaluated against this requirement, and its contribution to collaboration of human and artificial agents in the management of clinical knowledge is analyzed. PMID- 10522119 TI - Toward a framework for computer-mediated collaborative design in medical informatics. AB - The development and implementation of enabling tools and methods that provide ready access to knowledge and information are among the central goals of medical informatics. The need for multi-institutional collaboration in the development of such tools and methods is increasingly being recognized. Collaboration involves communication, which typically involves individuals who work together at the same location. With the evolution of electronic modalities for communication, we seek to understand the role that such technologies can play in supporting collaboration, especially when the participants are geographically separated. Using the InterMed Collaboratory as a subject of study, we have analyzed their activities as an exercise in computer- and network-mediated collaborative design. We report on the cognitive, sociocultural, and logistical issues encountered when scientists from diverse organizations and backgrounds use communications technologies while designing and implementing shared products. Results demonstrate that it is important to match carefully the content with the mode of communication, identifying, for example, suitable uses of E-mail, conference calls, and face-to-face meetings. The special role of leaders in guiding and facilitating the group activities can also be seen, regardless of the communication setting in which the interactions occur. Most important is the proper use of technology to support the evolution of a shared vision of group goals and methods, an element that is clearly necessary before successful collaborative designs can proceed. PMID- 10522120 TI - Evaluation of general practice computer templates. Lessons from a pilot randomised controlled trial. AB - We conducted a pilot randomised trial of computerised templates for the management of asthma and diabetes in general practice in six general practices in North London. Uptake of the guidelines by general practitioners and practice nurses was assessed using qualitative (semi-structured interviews designed to assess the users' views) and quantitative (change in use of the template during the study period) outcome measures. The practice nurses used the templates frequently but general practitioners rarely used them. Several reasons were offered for non-use of the templates, such as the length of the template and non involvement in the care of asthma or diabetes. Despite this, however, health professionals were favourably disposed to the use of templates for general clinical care. Pilot investigations of computerised templates are best achieved by observational or quasi-experimental methods rather than a randomised controlled trial. The use of both qualitative and quantitative methods in this study allowed exploration of the barriers to use of computers. PMID- 10522121 TI - Graphic representation of sequential Bayesian analysis. AB - Previously undescribed algebraic transforms of Bayes' theorem define families of operating points in the receiver operating characteristic (ROC) space which, at given pre-test probability, produce constant post-test probabilities. These isopredictive operating points form straight lines in the ROC space. The lines can be used to emulate Bayesian sequential analysis in a strictly graphic procedure, which can be applied in clinical work and medical education. PMID- 10522122 TI - Design elements for a computerized patient record. AB - Computerized medical record systems have to present user- and problem-oriented views of a patient record to health-care professionals. Presentation and manipulation of data must be easily adaptable to current and future demands of medical specialties and specific settings. During the definition, development and evaluation of a prototype of a computerized patient record system, design elements were elaborated to support physicians and other health-care professionals. Our approach shows a high degree of flexibility and adaptability to specific needs, problem orientation and connectivity to other systems, via a hospital information network. The explicit description of the contents of a patient record allows to augment the number of items that can be recorded without modifying the data structure. New views on patient data can be added to the system without interfering with the routine use of the system. Application in several medical specialties proved the feasibility of our prototype. PMID- 10522123 TI - Internet-based physician's workbench as user interface for a central medical case repository. AB - Two World Health Organization Radiation Medical Emergency Preparedness and Assistance Network centers have constructed a standardized central repository of acute radiation syndrome case histories. The case histories are stored on a database server. Radiation protection centers can remotely access the database by user-friendly client software over the Internet. Physicians can use the medical information system to retrieve similar case histories for decision support, to improve their medical knowledge by inspecting real case histories, and for research on the acute radiation syndrome. The system architecture is presented and it is shown in detail how the information system can be employed to deliver medical decision support. Dialogue-response times over narrow-bandwidth Internet connections are better than when using conventional World-Wide-Web technology. However, the latter does not require the installation of client software other than a browser. A Java applet as client could combine the advantages of the two approaches. PMID- 10522124 TI - Status and perspective of hospital information systems in Japan. AB - Questionnaire surveys were sent to hospital managers, designed to shape the policy for future hospital information systems in Japan. The answers show that many hospitals use dedicated management systems, especially for patient registration and accounting, and personnel, food control, pharmacy and financial departments. In many hospitals, order-entry systems for laboratory tests and prescriptions are well developed. Half of the hospitals have patient databases used for inquiries of basic patient information, history of outpatient care and hospital care. The most obvious benefit is the reduction of office work, due to effective hospital information system. Many hospital managers want to use the following sub systems in the future for automatic payment, waiting time display, patient records search, automatic prescription verification, drug side-effect monitoring, and graphical display of patient record data. PMID- 10522125 TI - Quantifying stenosis in renal arteriograms: a fuzzy syntactic analysis. AB - The introduction of fuzzy logic improves a system for the automatic quantification of renal artery lesions seen in digital subtraction angiograms. A two-step approach has been followed. An earlier system based on non-fuzzy syntactic analysis provided a clear symbolic description of the stenotic lesions. Although this system worked correctly, it did not take into account the variability and uncertainty inherent to image processing and to knowledge on the reference diameter. This system has been improved by the introduction of fuzzy logic in the representation of the reference diameter. It provides a description of the stenosis in terms of fuzzy quantities. To illustrate the benefits of the fuzzy approach, the results of the two systems have been compared by plotting the differences of an index of variability. It appears that the differences are statistically different when using a two-tailed paired t-test (t = 2.37; p = 0.025). The result shows that the fuzzy approach is better than a non-fuzzy approach in the sense that the index of variability is reduced significantly. PMID- 10522126 TI - Adaptable preprocessing units and neural classification for the segmentation of EEG signals. AB - In this contribution, a methodology for the simultaneous adaptation of preprocessing units (PPUs) for feature extraction and of neural classifiers that can be used for time series classification is presented. The approach is based upon an extension of the backpropagation algorithm for the correction of the preprocessing parameters. In comparison with purely neural systems, the reduced input dimensionality improves the generalization capability and reduces the numerical effort. In comparison with PPUs with fixed parameters, the success of the adaptation is less sensitive to the choice of the parameters. The efficiency of the developed method is demonstrated via the use of quadratic filters with adaptable transmission bands as preprocessing units for the segmentation of two different types of discontinuous EEG: discontinuous neonatal EEG (burst interburst segmentation) and EEG in deep stages of sedation (burst-suppression segmentation). PMID- 10522127 TI - [Evidence-based medicine in anesthesia]. PMID- 10522128 TI - Continuing medical education in anaesthesiology. PMID- 10522129 TI - [Continuing medical education in anesthesiology...the search for methods]. PMID- 10522130 TI - [General anesthesia with remifentanil in a case of "sleep apnea syndrome"]. AB - The authors report the case of a patient suffering from central sleep apnea (CSA) who underwent neurosurgery for ventriculo-peritoneal derivation under general anesthesia. Given the risk of postoperative hypoventilation in CSA, intraoperative anesthesia was induced using remifentanyl, an opiate with a plasma half-life of less than 5 minutes. Propofol (2 mg/kg) and remifentanyl at a dose of 0.5 microgram/kg was used during induction. The patient was curarised with vecuronium bromide, intubated and ventilated with a mixture of O2/N2O. During the operation, remifentanyl was administered as a continuous infusion at a starting dose of 0.2 microgram/kg/min, subsequently modified according to changes in arterial pressure and heart rate. At the end of surgery, which lasted approximately 120", decurarisation was carried out using prostigmin, and the infusion of remifentanyl was suspended, together with N2O. Reawakening times were recorded. Extubation took place 8' and 30" after the suspension of remifentanyl. Postoperative monitoring of SpO2 continued for 1 h and blood-gas analysis was satisfactory. No hypoventilation episodes were reported throughout the postoperative period and the patient was discharged from hospital after 7 days. The authors consider that remifentanyl should be the drug of choice to guarantee intraoperative analgesia in patients suffering from CSA requiring general anesthesia. PMID- 10522131 TI - Haemodynamic changes during anaesthesia in knee-elbow position. AB - BACKGROUND: To evaluate hemodynamic alterations during surgical procedures performed in the knee-elbow position. METHODS: DESIGN OF THE STUDY: prospective evaluation of the aortic blood flow (ABF) and other cardiovascular variables measured with a transesophageal Doppler (TED) in 2 groups of patients free of previous cardiovascular diseases undergoing lumbar discectomy. ENVIRONMENT: Operating theatre of a neurosurgical department. Beside TED, the standard monitoring included continuous ECG surveillance, capnometry, noninvasive measurement of blood pressure and pulsoxymetry. PATIENTS: Overall, 24 ASA 1 patients have been enrolled. In 12 patients (Group A) the intervention was performed without dobutamine. In the other 12 patients (Group B) dobutamine (2.5 3 micrograms/kg/min) was started from the beginning of the intervention; the two groups did not differ in terms of age, body size, duration of surgery or anesthetic technique used; hemodynamic measurements were obtained at the beginning of the intervention (T1), with the patients still supine, during the intervention in the knee-elbow position (T2) and finally at the end of the procedure (T3) being the patients still anesthetized but lying supine again. RESULTS: In both groups the ABF; the systolic volume and the ETCO2 significantly decreased in the knee-elbow position; concomitantly, peripheral vascular resistances increased. In Group B, the hemodynamic variables were significantly better than in the other group. CONCLUSIONS: In the patients enrolled, the perioperative administration of low-dose dobutamine was associated with better cardiovascular performance. PMID- 10522132 TI - [The effect on breastfeeding rate of regional anesthesia technique for cesarean and vaginal childbirth]. AB - BACKGROUND: To evaluate the influence of regional techniques of anesthesia and analgesia on breastfeeding rate after cesarean section and vaginal delivery. METHODS: STUDY DESIGN: prospective, area-based. SETTING: Obstetrics and Pediatrics Department at Aosta Valley Regional Hospital. SUBJECTS: all the mothers and their newborns during a three-year period (1993-1995). Maternal wish to breastfeed was the main inclusion criterion. Data recorded: feeding modality at discharge, anesthesia and analgesia modality, maternal/neonatal socio demographic and clinical data. RESULTS: 2725 records were examined, among them 1920 vaginal deliveries and 355 cesarean sections were statistically analyzed. chi 2 analysis showed a significant greater incidence of breastfeeding after cesarean section under regional anesthesia (spinal or epidural) versus general anesthesia: 95% vs 85.5%, p = 0.002. Breastfeeding rate was not different after vaginal delivery with epidural analgesia versus delivery without analgesia: 96.5% vs 97.8%. Logistic regression confirmed the positive role of regional anesthesia and few other maternal and neonatal variables on breastfeeding rate after cesarean section. CONCLUSIONS: Regional anesthesia seems to be advantageous for breastfeeding after cesarean section, probably because of a faster neonatal maternal bonding if compared with general anesthesia. Epidural analgesia for vaginal delivery does not adversely affects breastfeeding if compared with delivery without analgesia. PMID- 10522133 TI - [Severe infection from invasive beta-hemolytic streptococcus group A. Three cases of toxic shock observed in resuscitation]. AB - Group A streptococci (GAS) may cause a variety of infections, some of which are severe and may be life-threatening. Patients affected by severe invasive GAS infections may develop, early in the course of the infection, a syndrome characterized by circulatory insufficiency with multiple organ failure: Streptococcal Toxic Shock-Like syndrome (Strep-TSLS). The presence of shock and organ failure differentiate it from other types of invasive GAS infections. Three cases patients presenting with the Strep-TSLS, over a period of 16 months in our multidisciplinary 10-bed ICU are described. The Strep-TSLS was of nosocomial origin in the first case, due to poststernotomy wound infection, caused from erysipela in the second patient, and associated to a puerperal sepsis in the third case, respectively. In this small series the primary sources of streptococcal infection associated with the syndrome are confirmed to be in soft tissues and skin. One patient died early after the admission to the ICU, whereas the other two patients completely recovered with appropriate antibiotic and supportive treatment although early diagnosis and radical operative debridement may have been conclusive in one case. All 3 observed cases fulfilled the consensus case definition of "certain case of Strep-TSLS", whose criteria have been recently revised. PMID- 10522134 TI - Evaluation of the utility of serological tests in the diagnosis of candidemia. AB - BACKGROUND: It has been observed that the incidence and prevalence of candida infections in critically ill non-immunocompromised patients has increased. This study aims to evaluate the utility of the use of serological tests (double immunodiffusion and Cand-Tec Test) for the determination of candidemia. METHODS: A retrospective evaluation is made of 214 patients admitted to the Intensive Care Unit (ICU) of the Agostino Gemelli University Polyclinic during a period of 42 months. The double immunodiffusion technique was utilized for the determination of Candida antibodies. The Cand-Tec latex agglutination test was performed to evaluate the presence of Candida antigen. Four hundred and fifty-five antigen and antibody tests were performed. RESULTS: Thirty-six patients (16.8%) developed an invasive candidiasis. The sensitivity and specificity of antibody detection tests was 29 and 67 respectively; the positive predictive value was 15 and the negative predictive value was 82. The sensitivity and specificity of the antigen detection test ranged between 82 and 3 and between 8 and 98 respectively according to different cut-off titre; the positive predictive value was low (13-17%) and the negative predictive value decreased from 70 to 29. CONCLUSIONS: The utility of the use of serological tests in the diagnosis of candidemia is extremely limited. The gold standard for the determination of Candida sepsis remains the demonstration of candida in blood or in tissues. PMID- 10522135 TI - Perioperative management of glucose 6 phosphate dehydrogenase deficiency. A review of the literature. AB - Glucose 6 phosphate dehydrogenase (G6PDH) deficiency is the most frequent cause of hemolytic anemias due to enzyme abnormality. Perioperative management must be careful to avoid the onset of hemolytic crisis. We present a complete review of the literature on this illness and describe the perioperative management of an adult with known G6PD deficiency and the pathogenesis and clinical manifestations of the disorder and its possible anesthetic implications are discussed. A 49-year old patient had undergone varum osteotomy in her left knee due to genu valgum. She had been diagnosed as having G6PDH deficiency sixteen years earlier provoked by ingesting beans. The perioperative circumstances capable of causing autohemolysis are described and discussed. In spite of the fact that the pattern is self-limited, it provokes the onset of jaundice and anemia which can complicate the recovery. Simple elimination of those elements which precipitate with oxyhemoglobin will allow an uneventful anesthetic procedure. PMID- 10522137 TI - [Continuous epidural anesthesia with a double catheter for sedation for surgery of the vertebral dorsolumbar column]. AB - The case of presented of an obese 42 year-old female patient, undergoing a second spinal cord decompression for a large dorsolumbar fibrous-scar mass, having a small-bore Montgomery tracheal stent (T-tube) on site. Stent replacement with a tracheotomy tube was impossible because of strong accretions hindering stent removal, as well as insertion of a suitable tracheal tube through the external stent branch, because of its very small lumen. So, general anaesthesia was not administered and surgery was performed under continuous epidural block, with light sedation, using two catheters introduced up and down the compression. Surgical and anaesthetic outcomes were optimal and confirm the effectiveness and safety of epidural anaesthesia for dorsolumbar spinal surgery, even in the obese patient. Moreover, with the continuous double-catheter technique it was possible to achieve a good and homogeneous spread of the analgesic solution, with low volume, despite the hard compression and deformation of the epidural space due to the fibrous-scar mass. PMID- 10522136 TI - [Anesthetics management in patients with epidermolysis bullosa]. AB - The anesthetic management used in a 4-yr-old boy with dystrophic epidermolysis bullosa (DEB) submitted to long-lasting urological surgery is reported. Medical treatment of cutaneous bullae in various stages of healing was undertaken preoperatively. Other preoperative therapies were performed to treat sepsis and to improve poor nutritional state. Different measures were used during monitoring and inhalation induction and maintenance of anesthesia in order to prevent frictional trauma to the skin and oropharyngeal mucosa, leading to blistering. Anesthesia and surgery proceeded uneventfully, while surgical complications and sepsis occurred postoperatively. However the patient was discharged from intensive care unit to surgical ward on 35 postoperative days without evidence of oropharyngeal bullae and with a good improvement of his cutaneous condition. The conclusions are drawn that, even if the patient with DEB represents a challenge to the anesthesiologist, a long-lasting surgical intervention can be performed successfully. The main aspect in the anesthetic management of these patients is the detailed knowledge of all problems that anesthesiologist must face and solve to preserve skin and mucosa integrity, avoiding the risk of severe complications. PMID- 10522138 TI - [Anaphylactic reaction to thiopental. A case documented by tryptase values and RAST]. AB - During general anesthesia adverse reactions are not uncommon, and the hypothesis of an anaphylactic response cannot be excluded. In these situations, a differential diagnosis from other events which often present identical clinical manifestations is necessary. For this purpose measurement of serum tryptase, as a biochemical marker of the release of mast-cell granules, is considered a valuable and specific method, especially if it is carried out on several hematic samples, obtained in successive times, for pointing out the progressive reduction of the values. If an anapyhlactic pathogenesis is confirmed, the identification of the responsible drug is necessary for a safer approach of the patient in view of a further anesthesia. A severe and protracted reaction has been observed after a standard induction of anesthesia, in which measurement of serum tryptase has shown high values of this protease 2 hours after the reaction which a subsequent decrease in the samples repeated after 5, 8, 11 e 20 hours, suggesting an anaphylactic etiology of the reaction. The specific RAST for the substances employed has excluded a role for muscle relaxant, plasma expander and latex, while it was positive for tiopenthal, suggesting in this case a true anaphylactic reaction caused by the hypnotic drug. PMID- 10522139 TI - [Peripartum dilatative cardiomyopathy. Case report with literature review]. AB - Peripartum cardiomyopathy (PPCM) is a rare form of heart failure affecting women in the last month of pregnancy or the first six months post-partum. The etiology of PPCM remains poorly understood although some risk factors were described. Diagnosis is often difficult and is always necessary to exclude other prior heart disease and other cause of left ventricular dysfunction in pregnancy. Medical therapy for PPCM is similar to that for other forms of congestive heart failure; prognosis is better than in idiopathic cardiomyopathy but many authors observed that women who have had one episode of PPCM are likely to have recurrences in subsequent pregnancies. The present report describes the case of a woman presenting with severe cardiac failure immediately after cesarean section for twin pregnancy. The patient is a 35-year-old nulliparous white woman, with history of anorexia, subsequent amenorrhea, sterility and pregnancy induced with Gn-Rh. The diagnosis of PPCM was difficult for the presence of preeclampsia and acute pulmonary edema occurred four hours after delivery. The successful outcome was possible with an intensive treatment (mechanical ventilation, Swan-Ganz catheter). The whole resolution of the heart failure, six months post-partum, was demonstrated by ultrasonography. PMID- 10522141 TI - [Scientific knowledge and the methodologic problem of pain in medicine]. AB - The paper discusses the epistemological question of pain and the way this concept forms part of scientific knowledge. The first part gives a rapid overview of the main aspects of modern scientific knowledge. In earlier times Bacon, Galileo and Descartes felt that the new form of knowledge could head to a knowledge of truth and certainty. Today, these ideals have proved unattainable and science only takes the form of well founded, rigorous and objective knowledge. Moreover, the objectivity of science is based on intersubjectivity. The second part of the paper examines pain and underlines that it is a subjective phenomenon which cannot be ascertained by the doctor in the same way as commonplace anatomic and physiological phenomena. Our scientific knowledge of pain is based on operations for which it is possible to achieve an intersubjective knowledge: these operations constitute the protocol criteria on which it is possible to base a scientific knowledge of pain. In the last part of this paper, having stressed that pain is a concept belonging to many disciplines--scientific, psychological, philosophical, religious--the emphasis is placed on pain itself as opposed to suffering. While the former is a concept that belongs to scientific medicine, the latter belongs to that area of medicine which lies beyond the boundaries of science and concerns the activity of a man who helps a fellow human who is suffering. PMID- 10522140 TI - [Acute pulmonary edema from dislocation of a laryngeal mask]. AB - A case of pulmonary edema secondary to dislocation of laryngeal mask during awakening from anesthesia is described. The complication has been ascribed to physiopathological changes of the alveolo-capillary membrane caused by inspiratory efforts secondary to dislocation of laryngeal mask during awakening from anesthesia. Other possible causes of pulmonary edema are discussed, especially the cardiogenic one, and among the non cardiogenic edema, the Mendelson's syndrome. The quick identification and the intensive care of post obstructive pulmonary edema lead to a rapid resolution of the pathology. PMID- 10522142 TI - [Acute pain service warning signs for perioperative intensive care]. PMID- 10522143 TI - [Phlebodynamometry in the study of venous insufficiency of the lower limbs]. AB - BACKGROUND: The significance of ambulatory venous pressure measurement in the diagnosis of chronic venous insufficiency of the lower limbs is evaluated. METHODS: 190 limbs of 120 outpatients were investigated from 1991 through 1997; the study was performed whenever clinical and ultrasound evaluation did not allow to establish the diagnosis of "simple varices". The technique standardized according to Nicolaides was followed. RESULTS AND CONCLUSIONS: The results of the study confirm the importance of ambulatory venous pressure measurement in the differential diagnosis of chronic venous hypertension of the lower limbs, especially when caused by vein compression, as well as in the follow-up of these patients when submitted to surgical treatment. PMID- 10522145 TI - Acute cardiovascular diseases and respiratory sleep disorders. AB - Respiratory sleep disorders are a risk factor, sometimes independent, for acute cardiovascular diseases which are the most frequent cause of death among populations of industrialized countries. Snoring and obstructive sleep apnea (OSA) are generally involved, while the pathogenetic role of acute exacerbation of COPD seems less evident. The most important acute cardiovascular events related to sleep respiratory disorders are angina pectoris, acute myocardial infarction, cardiac arrhythmias (in some instances as paroxysmal attacks), systemic hypertension with hypertensive crisis, ischemic stroke. A respiratory sleep disorder should be suspected in all obese, cigarette smokers, alcoholics, hypertensives, who present symptoms of obstructive sleep apnea, where snoring may be a marker, and in patients with COPD. The diagnosis is readily established by performing polysomnography and, when needed, by 24-hour Holter monitoring and blood pressure ambulatory recording. Therapy aims at correcting risk factors with particular attention to weight reduction in obese patients. Furthermore, upper airway anatomic abnormalities should be eliminated. In obstructive sleep apnea, nasal continuous positive airway pressure during sleep is to be used, when necessary, while tracheostomy must be performed only in more severe cases. PMID- 10522144 TI - Prevention of embolism using calciparine during electrical cardioversion in atrial fibrillation. AB - On the basis of a review of literature data on cardioversion of AF, a protocol of fast decoagulation using calciparine in non-valvular AF is examined. Pre cardioversion patients were sedated by administering Diazepam at a personalized dose and without significant subjective disturbances for the patients; this meant not having to involve the anaesthetist. PMID- 10522146 TI - [The results of large clinical trials comparing primary angioplasty and thrombolysis]. AB - One of the main cardiological debate is about which one, between primary angioplasty (PTCA) and thrombolysis, is to prefer for the therapy of acute myocardial infarction. The data available in the literature do not show that one of these two therapeutical choices is definitely better than the other one. Since the main therapeutical goal in patients with acute myocardial infarction is the early and persisting recovery of the anterograde coronary flow, the best therapy for every patient is the one that can be performed more quickly and safely. Therefore, PTCA has to be preferred whenever it can be done quickly, by expert personnel and with cardiosurgical support, especially in patients considered to be at high risk or with contraindications to thrombolysis. Otherwise, thrombolytic therapy should be better. Stent implantation seems to be better than conventional angioplasty, in particular for the reduction of restenosis and reocclusion. These conclusions, however, derive from small studies and require further evidences. Moreover, there are not trials directly comparing primary PTCA and stent implantation with thrombolysis. Rescue PTCA, after failure of thrombolytic therapy, is useful when the coronary flow of the culprit lesion is TIMI 0 or 1, but not when the flow is TIMI 2; there are neither indications to early, but not rescue, angioplasty in all the patients already thrombolyzed. Finally, for patients with acute myocardial infarction and cardiogenic shock, the data currently available, derived more from observational than from randomized studies, suggest revascularization by PTCA. PMID- 10522147 TI - [The determination of the thermal threshold in subjects with Raynaud's disease. A preliminary report of 2 clinical cases]. AB - BACKGROUND: The onset of Raynaud's phenomenon is mainly provoked by cold stimuli which, in patients affected by Raynaud's disease, are milder than those able to provoke the same phenomenon in healthy subjects. Therefore, it is reasonable to theorize the existence of a "thermal threshold", below which it is more likely for a Raynaud's phenomenon to be provoked. The aim of this investigation is to determine whether a thermal threshold in Raynaud's disease exists and if it can be used as a practical tool for preventing the complications of Raynaud's disease (digital gangrene and ulcers), whose onset is strictly related to the frequency of the phenomena. METHODS: Two patients affected, respectively, by slight Raynaud's phenomenon secondary to peripheral neuropathy (Case 1), by severe Raynaud's phenomenon associated with mixed connective tissue disease (Case 2), were asked to write a note about the phenomena and the days of their onset, within a period of three months. Afterwards, all data were related to the daily ground maximum temperature occurred in the period of study. RESULTS AND CONCLUSIONS: These preliminary results show an interesting correlation between this new parameter and the clinical data, the thermal threshold being higher in more severe cases. It was equal to 22 degrees C in the patient affected by slight phenomenon (Case 1), and equal to 27 degrees C in the severe phenomenon (Case 2). This method for evaluating the thermal threshold in these two cases of Raynaud's disease (the greater the figure, the more severe the case) allowed both the clinical assessment of the pathology and the prevention of secondary complications. PMID- 10522148 TI - [Edema and skin ulcers of the lower limbs as a collateral effect of nifedipine. A clinical case report]. AB - Nifedipine, as most calcium-antagonist drugs, is widely used for the treatment of angina pectoris and primary hypertension. Among its side effects, edema of lower limbs may expose patients to traumatic ulcers which become chronic. A case with painful ulcers and remarkable swelling of the ankles is reported. In this case, misdiagnosed as chronic venous insufficiency, good results were obtained with topical treatment and discontinuation of nifedipine. Leg ulcers and edema resolved in four months after ten years of disease history. PMID- 10522149 TI - [The evaluation by video capillaroscopy of the efficacy of a Ginkgo biloba extract with L-arginine and magnesium in the treatment of trophic lesions in patients with stage-IV chronic obliterating arteriopathy]. AB - BACKGROUND AND AIMS: This study aimed to evaluate the influence of Ginkgo biloba extract with arginine and magnesium used for the treatment of trophic lesions in the lower limbs caused by both diabetic and non-diabetic microangiopathy. METHODS: A comparative study was carried out in 20 patients who were divided into two groups: 10 were treated with ASA plus Ginkgo biloba extract with arginine and magnesium and 10 with ASA plus conventional hemorheology. The observation time was extended to 6 months, taking into consideration patients with trophic lesions to the lower limbs suffering from diabetic and non-diabetic peripheral arterial occlusive disease. The evaluation was performed by clinical, ultrasonographic and perilesional video capillaroscopy monitoring. Ultrasonographic and video capillaroscopy instrumental methods were used because they provide a full picture of macro and microcirculatory conditions around lesions. RESULTS: The study showed the undoubted efficacy of Ginkgo biloba extract with magnesium and arginine in relation to the following points: reduced healing times for the trophic lesion compared to the control group, improved painful symptoms, increased perilesional neoangiogenesis. No significant differences were observed from a Doppler ultrasonographic point of view or with regard to the claudication free interval. CONCLUSIONS: Ginkgo biloba extract with arginine and magnesium can improve the dynamics of cutaneous trophism in lesions caused by diabetic and non diabetic microangiopathy. PMID- 10522150 TI - Phosphorus balance and prostate cancer. AB - Prostatic diseases affect men over the age of 45 and increase in frequency with age so that by the eighth decade more than 90% of men have Benign Prostatic Hyperplasia, (BPH), of which some progress to Carcinoma of Prostate (CaP). CaP, the most common malignancy in men, is also the second most common cause of death in men. Over the last three decades the mortality rate for CaP has steadily increased. There, however, are scant clues to the aetiology/pathogenesis of CaP. As treatment failures of advanced carcinoma continue to frustrate clinicians, emphasis has recently been focused on possible preventive strategies. Several studies support the view that higher levels of 1,25-(OH)2D, the active metabolite of vitamin D, reduce the risk of BPH and CaP. Men with high serum levels of 1,25 (OH)2D have a reduced risk of poorly differentiated and clinically advanced CaP. Hypercalcemic activity of 1,25-(OH)2D or its analogues, however, thwart their use for therapy in humans. Incidentally, a low dietary intake of phosphorus has been reported to increase serum levels of 1,25-(OH)2D. In addition, dietary fructose reduces plasma phosphate levels by 30 to 50% for more than 3 hr. Fruit intake has, indeed, been shown to be associated with reduced risk of CaP, particularly the advanced type. These observations, taken together, support that dietary determinants of hypophosphatemia, leading to increased plasma levels of 1,25 (OH)2D, could reduce the risk of ageing men to develop prostatic diseases, both BPH and/or carcinoma of Prostate. PMID- 10522151 TI - Chicken egg yolk anti-asialoGM1 immunoglobulin (IgY): an inexpensive glycohistochemical probe for localization of T-antigen in human colorectal adenocarcinomas. AB - A egg yolk polyclonal IgY has been prepared by immunization of white leghorn chickens with small unilamellar liposomal asialoGM1. The newly prepared anti asialoGM1 IgY has been characterized to be specific toward the terminal carbohydrate moiety of asialoGM1, and has no cross reactivity to its sialylated counterpart (ganglioside, GM1) as evidenced by immunochromatographic studies. General glycohistochemical methods along with antigen specific lectin and immunohistochemical staining using anti-asialoGM1 IgY were used to study the expression of Thomsen-Friedenreich (T-) disaccharide antigen in human colorectal adenocarcinoma tissues. The expression of T-antigen in colon cancer tissue was detected by two T-disaccharide specific probes, chicken anti-T-yolk antibody (IgY) and Artocarpus integrifolia lectin (AIL) and was found to be more pronounced in both the secreted mucin as well as the cytoplasmic mucin deposits. These immunochemical detection methods for T-antigen showed a weaker correlation with other glycostaining methods using, alcian-blue/periodic acid-Schiff (AB-PAS) and high iron diamine (HID). However, a general enzymatic staining for galactose and galactosamine containing glycoconjugates, by galactose oxidase-Schiff method, showed a good correlation with T-antigen detection. While the T-beta specific anti-asialoGM1 could localize T-antigen in 11 of 13 (84%) human colorectal adenocarcinoma tissue sections tested, the T-alpha specific AIL could localize the T-antigen in only 6 of the tissues (46%). These observations confirm previously reported findings, of the prevalence of T-beta conformation in colon cancer, that binds significantly more with the anti-asialoGM1 IgY than with the T alpha specific AIL. Hence, both anti-T IgY and the AIL immunohistochemical probes may have useful diagnostic value because of the ease of preparation and cost effectiveness, but the T-beta specific anti-asialoGM1 probe (IgY) would have a better prognostic value in colon adenocarcinomas. PMID- 10522152 TI - Monoclonal antibodies against bovine growth hormone. AB - A number of mouse x mouse hybridomas producing monoclonal antibodies (MAbs) against bovine growth hormone (bGH) were prepared by fusion of spleen cells from bGH-primed mice (Balb/c) with non-secretory mouse myeloma cells (PAIOP3) and characterized. MAbs obtained from three fusion experiments belonged to IgM, IgG1 and IgG2b class/subclass of antibodies. Cross-reaction studies indicated that generated antibodies were against three different epitopes of bGH. VIA6E8 (IgG1) and VIIB2E11C9 (IgM) did not cross-react with ovine prolactin (oPRL), ovine leutinizing hormone (oLH) and porcine follicle stimulating hormone. Antibody VIB3C9E8 (IgM) exhibited cross-reaction with oPRL and oLH. Antibody VIC1F9 (IgG2b) cross reacted with oPRL. All MAbs were against conformational epitopes of bGH. PMID- 10522153 TI - Identification of bovine sperm specific polypeptides reactive with antisperm antibodies. AB - The present study was taken to characterize molecular weights of sperm specific polypeptides antigenic to rabbits and calf with the aim to assess their immunoreactivity with IgG antibodies in sera from immuno-infertile cows. Seropositivity for antisperm IgG antibodies in 75 repeat breeder and 15 pregnant control cattle was tested by cellular ELISA using washed spermatozoa antigen from 4 bulls. Molecular weights of bovine sperm polypeptides antigenic to rabbit and calf were determined by 10% SDS-PAGE and Western blotting. Molecular weights of sperm peptides reactive with sera from immuno-infertile cows were also determined. Seropositivity of antisperm IgG antibodies for bull I, II, III and IV was 23.6, 14.6, 26.6 and 20%, respectively. A total of 16 polypeptides were discernible on gel. Out of these, 7 polypeptides were immunoreactive with sera from hyperimmunized rabbits as compared to 3 poly-peptides which reacted with sera from hyper-immunized calf. Only two polypeptides were reactive with sera from immuno-infertile cows. Variable number of sperm polypeptides and their immunoreactivity have been reported in different species. Antigenicity of different polypeptides in sperm needs further investigations. PMID- 10522154 TI - Reduced nociceptive responses in mice with alloxan induced hyperglycemia after garlic (Allium sativum Linn.) treatment. AB - Administration of ethanol (95%) extract (45 mg/kg body wt/day for 28 days) of garlic (A. sativum) to alloxan induced diabetic (ALX-D) mice significantly lowered the serum glucose levels, nociceptive response in tail-flick, hotplate, allodynia, formalin test and relative thickness, weight of hind paw in formalin induced Paw oedema test, over 28 days, thus, showing the reversal trend in hyperglycemia and hyperalgesia compared to ALX-D mice. The reversal of hyperglycemia and hyperalgesia was progressive and more effective as duration of extract administration increased. The results suggest therapeutic potential of ethanol extract of garlic for anti-hyperglycemic and anti-nociceptive effects in diabetes. PMID- 10522155 TI - Molecular mechanics aided design of antineoplastic agents from ruthenium coordinate complexes. AB - Through energy minimization using molecular mechanics force field four ruthenium cordinate complexes have been synthesized. Compound I to IV showed antineoplastic activity with varying degree on EAC bearing mice. Mode of action may be through inhibition of antioxidant property of tumor cell as evident from lipid peroxidase activity. Among the complexes Bis pyridine tetrachloro ruthenium exhibits highest order of activity with respect to increase mean survival time, inhibition of tumour volume, total blood count, hemoglobin and lipid peroxidase activity. PMID- 10522156 TI - Potentiality of tricyclic compound thioridazine as an effective antibacterial and antiplasmid agent. AB - Thioridazine (Th), which is therapeutically used in psychiatric patients, was found to possess conspicuous antimicrobial activity when tested against 316 strains belonging to a number of Gram positive and Gram negative bacteria. Although Staphylococcus aureus, Vibrio chloerae and V. parahaemolyticus were found to be most sensitive, Th was highly bactericidal against S. aureus and bacteriostatic for vibrios and other Gram negative organisms. In the study of antiplasmid/curing effect of Th on twelve multiply antibiotic and Th resistant bacteria, it was observed that elimination of R plasmids was facilitated by choice of optimal concentration of Th. Significant elimination of single and combined antibiotic resistance occurred in E. coli and Shigella flexneri and not in S. aureus. PMID- 10522157 TI - Antioxidant activity of active tannoid principles of Emblica officinalis (amla). AB - The antioxidant activity of tannoid active principles of E. officinalis consisting of emblicanin A (37%), emblicanin B (33%), punigluconin (12%) and pedunculagin (14%), was investigated on the basis of their effects on rat brain frontal cortical and striatal concentrations of the oxidative free radical scavenging enzymes, superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX), and lipid peroxidation, in terms of thiobarbituric acid reactive products. The results were compared with effects induced by deprenyl, a selective monoamine oxidase (MAO) B inhibitor with well documented antioxidant activity. The active tannoids of E. officinalis (EOT), administered in the doses of 5 and 10 mg/kg, i.p., and deprenyl (2 mg/kg, i.p.), induced an increase in both frontal cortical and striatal SOD, CAT and GPX activity, with concomitant decrease in lipid peroxidation in these brain areas when administered once daily for 7 days. Acute single administration of EOT and deprenyl had insignificant effects. The results also indicate that the antioxidant activity of E. officinalis may reside in the tannoids of the fruits of the plant, which have vitamin C-like properties, rather than vitamin C itself. PMID- 10522158 TI - Purification and characterization of Klebsiella pneumoniae cytotoxins. AB - Isolation, purification and characterization of 3 new cytotoxins of a K. pneumoniae strain isolated from ready to eat pork sausage are reported. Purification process involved extraction of cytotoxins with polymyxin B sulphate, salt precipitation, gel filtration and anion exchange chromatography. Klebsiella cytotoxin (KCT) I, a glycoprotein of about 65 kDa was verocytotoxic, enterotoxic and dermonerotic. KCT II was erythemogenic, verocytotoxic and enterotoxic protein of co 55 kDa, while KCT III was about double in MW (110 kDa) hadverocytotoxicity but neither enterotoxicity nor dermatotoxicity. KCT I and II caused granulation, conglomeration, shrinkage, detachment and lysis of MDBK and Vero cells, while KCT III induced enlargement, vacuolation, granulation, multinucleolation and syncytia formation in exposed cells. All the three cytotoxins induced specific neutralizing antibodies and cytotoxins were detectable in nanogram quantities with enzyme-linked immunosorbant assay using homologous antibodies. None of the anticytotoxin cross-reacted with either heterologous Klebsiella cytotoxins or with verocytotoxic preparations of Shigella dysenteriae. PMID- 10522159 TI - Induction of stable benomyl-tolerant phenotypic mutants of Trichoderma pseudokoningii MTCC 3011, and their evaluation for antagonistic and biocontrol potential. AB - Trichoderma pseudokoningii MTCC 3011 is a very useful strain for biological control of the plant pathogen Sclerotium rolfsii under post-harvest conditions. In the present investigation, several benomyl-tolerant phenotypic mutants of this strain have been generated using a two step mutagenesis-chemical followed by gamma irradiation. The mutants differed from the wild type strain in antibiotic and disease control potential. Some of the mutants are superior to the wild type in biocontrol potential on S. rolfsii. PMID- 10522160 TI - Latitudinal variation in eclosion rhythm among strains of Drosophila ananassae. AB - Eclosion rhythm parameters of D. ananassae strains originating between 8 degrees 34 degrees N were highly variable and latitude dependent. In the field under naturally fluctuating light intensity, temperature and R.H., the amplitude of the rhythm was high and the eclosion gate was narrow; however, under the naturally fluctuating light intensity but at constant temperature and R.H., the amplitude of the rhythm was lowered and the width of eclosion gate was widened. The eclosion rhythm entrained to light-dark (LD) cycles ranging from LD 6:18 to LD 18:6, the width of the eclosion gate was decreased and increased in the short and long photoperiods respectively. Among the strains, both the phase angle difference (psi, the time from lights-off in a 24 hr LD cycle to the eclosion median) and the period of free-running rhythm (tau) in constant darkness varied by about 3 hr and the amplitude of the rhythmicity (Amp) by about 10%. Lower latitude was correlated with late psi (r = -0.69), long tau (r = -0.88) and high Amp value (r = -0.95). PMID- 10522161 TI - Use of yeast respiratory adaptation test system to detect chemical mutagens/carcinogens in mammals. AB - An approach to follow distribution of injected DNA-acting chemicals (mutagens/carcinogens) in animal tissues has been described. This is based on the use of respiratory adaptation (mitochondrial biogenesis) process in Saccharomyces cerevisiae during transition from anaerobic to aerobic state. By virtue of specific interaction of such chemicals with mitochondrial DNA associated with promitochondrial structures this process is extremely sensitive to DNA-acting chemicals. Solutions of berylium sulphate, aflatoxin G1, aflatoxin B1, and carbaryl (all known DNA-acting agents) were injected to rats at low concentrations and, after 24 hr, distribution of these chemicals or their metabolites was studied by determining the inhibitory action of appropriately diluted urine and tissue homogenates on respiratory adaptation in S.cerevisiae. Detectable amounts of the chemicals and their DNA-acting metabolites could be analyzed in urine, liver, lungs, kidney and spleen. PMID- 10522162 TI - In vitro studies on peroxidative changes leading to hemolysis of erythrocytes infested with malarial parasite Plasmodium vivax. AB - Blood erythrocytes of 25 confirrhed malarial patients infested with P. vivax were analyzed for peroxidation and hemolysis and results compared with 10 uninfected normal control samples. Results indicated significant increase in peroxide formation measured as malondialdehyde, both in presence and absence of H2O2, in parasite infested erythrocytes. These changes induced hemolysis of infected erythrocytes which was increased manifold in presence of H2O2 and could probably be the reason for extensive anemia reported in malaria. PMID- 10522163 TI - Production of calcium gluconate by fermentation. AB - Calcium gluconate production by Aspergillus niger was investigated in shake flask, rolling shaker, air-lift reactor and stirred reactor. Growth pattern of the organism and fermentation conditions determined the yield of the product. High calcium gluconate production was achieved in air-lift reactor with pellet form of cell growth at moderate specific growth rate and biomass concentration. In another variation of air-lift reactor, when calcium carbonate was confined to a cellulose membrane, calcium gluconate production was maximum (149 g/L). At higher specific growth rate, obtained in shake flask, despite the formation of cell pellets, product formation was low. Physical separation of particulate calcium carbonate and growing cells favoured product formation. In stirred reactor pulpy mycelial growth was obtained and calcium gluconate production was poor. PMID- 10522164 TI - When is a medical product too risky? An interview with FDA's top drug official. Interview by Tamar Nordenberg. PMID- 10522165 TI - Campylobacter. Low-profile bug is food poisoning leader. PMID- 10522166 TI - Keeping food safety surveys honest. Video checks up on consumer meal preps. PMID- 10522167 TI - When machines do the breathing. PMID- 10522168 TI - Crohn's disease. New drug may help when others fail. PMID- 10522169 TI - Taking time to use medicines wisely. PMID- 10522170 TI - Maker of growth hormone feels long arm of law. PMID- 10522171 TI - Sales of contaminated animal drugs halted. PMID- 10522172 TI - Repair of the mandibular nerve by autogenous grafting after ablative surgery of the mandible. AB - In this paper, we introduce our original method of autologous nerve grafting for substitution of the mandibular nerve after mandibular resection in three subjects. The lateral cutaneous nerve of the forearm served as a donor nerve, and the graft was imbedded microsurgically at the junctures using an epineurial nerve suture technique. Oversized grafts were chosen in order to insert them without tension in a space between the stumps of the recipient nerve and the regeneration zone of the bone. In all three subjects, sensibility of the lower lip and chin recovered completely after about ten months. PMID- 10522173 TI - A study of the changes in facial profile after orthodontic treatment. Part 1. Comparison between the improved group and unimproved group. AB - The purpose of this study was to investigate the harmonious facial profile before and after orthodontic treatment on permanent dentition and to determine the factors which influence lateral facial harmony. Materials were lateral Roentgen cephalograms from 150 subjects (25 males and 25 females each in 3 groups- maxillary protrusion, mandibular protrusion and crowding--) taken before and after treatment. The average age before treatment was 11 years and 6 months and, after treatment, was 14 years and 3 months. The term of active treatment was 2 years and 9 months. Lateral facial evaluation based on the external profile line was performed by a group of 40 persons which included dental students and the general public. In the 5 stage evaluation, each subject could receive from 40 to 200 points. The subjects with more than 121 points were classified in the harmonious group; those with less than 120 points were classified in the disharmonious group. Subjects who had been classified as disharmonious before treatment but became harmonious after treatment constituted the improved group. Subjects classified disharmonious before treatment who remained disharmonious after treatment constituted the unimproved group. The value of the overall harmonious group before treatment was 8.6% and, after treatment, was 36.6%. The increase in the percentage of cases classified as harmonious varied among the different occlusal types: for the mandibular protrusion, the value before treatment was 2% and after treatment was 32%, for crowding it was 18% before and 46% after treatment, and for maxillary protrusion, it was 6% before and 32% after treatment. The ratio of percentage in the harmonious group increased on every malocclusion group after treatment, suggesting the importance of orthodontic treatment for improvement of lateral facial harmony. Before treatment, the morphological conditions in the improved group were more advantageous than those in the unimproved group. PMID- 10522174 TI - The DYS19 system in the Japanese population and its detection using teeth as a source of DNA. AB - The Y-specific short tandem repeat (STR) polymorphism of the locus DYS19 was amplified by PCR and analyzed by denaturing polyacrylamide gel electrophoresis followed by silver staining. Among 119 DNA samples from Japanese males, five alleles were observed. When sequences of the products were compared, each allelic segment contained 13 to 17 GATA tetranucleotide repeats, and revealed no differences from the known allele (GenBank X77751) other than the number of tetranucleotide repeats. The most common allele in the Japanese population was allele 15, and the distribution of the alleles did not differ from the data from other regions in Japan but did differ from those of Caucasians. Amplification of the locus using 12 tooth samples as a source of DNA matched the patterns obtained from blood samples. PMID- 10522175 TI - Pleomorphic adenoma with nuclear palisading arrangement of modified myoepithelial cells: histopathologic and immunohistochemical study. AB - Pleomorphic adenomas with a nuclear palisading arrangement of spindle-shaped modified myoepithelial cells (MMCs), suggesting the appearance of palisading leiomyoma or Antoni's A type of neurilemmoma, are quite rare, and its cytologic nature has been poorly understood. This paper reports histologic and immunohistochemical findings of palisading MMCs in two cases of pleomorphic adenoma. Histologically, foci of spindle-shaped MMCs with nuclei in a palisading arrangement were scattered in the myxoid areas. Near the large foci of spindle shaped MMCs with nuclear palisading arrangements, tiny foci of spindle-shaped MMCs forming nuclear palisading or rosette-like arrangements were seen. Such nuclear palisading arrangements of MMCs were suggestive of differentiation or transformation of MMCs into cells that were more smooth muscle in nature, supported by occasional existence of palisading leiomyoma in the myometrium and gastrointestinal tract. However, immunohistochemical findings of palisading MMCs in pleomorphic adenoma were similar to those of non-palisading MMCs, and showed no evidence of smooth muscle differentiation; neither palisading nor non palisading MMCs in pleomorphic adenoma expressed desmin, muscle specific actin (HHF-35), alpha smooth muscle actin, or myoglobin. The biologic significance and formative mechanism of nuclear palisading arrangement of MMCs in pleomorphic adenoma could not be determined in the present study. However, if the MMCs with nuclear palisading arrangements in pleomorphic adenoma, presented here, are aspirated for cytologic diagnosis or are included in a small biopsy specimen, the correct diagnosis of pleomorphic adenoma may be confused by a suspicion of myogenic or neurogenic tumor. PMID- 10522176 TI - A study of the harmonious profile in facial esthetics. Part 1. Descriptive statistics. AB - The purpose of this study was to investigate the relationship between a harmonious profile and normal occlusion, differences between good and poor groups, and the characteristics of a good group from normal occlusion group and after orthodontic treatment groups. The sample included 60 subjects with normal occlusion (normal subjects) and 88 after orthodontic treatment subjects (orthodontic subjects). Evaluators were 20 students and 20 orthodontists from Tokyo Dental College. For the purpose of profile evaluation, normal and orthodontic subjects were classified into 3 groups (good, mediocre, and poor) based in their external profile lines. Lateral Roentgen-cephalograms were used to measure hard and soft tissues. Mean values and Student's t-test were calculated statistically. RESULTS: 1. The frequencies of the good and poor subjects were similar in both normal subjects and orthodontic subjects. When considering the mediocre group, however, the normal subjects were more likely to have a better profile than the other subjects. 2. There were differences in hard tissues between the good group and the poor group in both the normal subjects and the orthodontic subjects, but only slight differences in soft tissues. 3. In the good groups of both normal and orthodontic subjects (all subjects), the edges of the upper and lower central incisors and upper and lower lips retruded, the ratio of upper facial height to total facial height was greater and the chin region was thicker than in the poor group. Additionally, in the good group of normal subjects, the maxilla protruded and the anteroposterior difference between the maxilla and mandible was larger. PMID- 10522177 TI - Microhardness evaluations of resin-dentin bonding areas by nano-indentation. AB - The purpose of this experiment was to determine the hardness values of the hybrid layer and its surroundings through the continuous use of a microhardness measuring device. Black's Class V cavities were prepared in nine dog teeth. The cavities were divided into four groups according to the dentin adhesive system applied. The adhesive systems were: "Bond One System", "Liner Bond II sigma System", "One Step System", and "Single Bond System". The treated teeth were observed at seven days post-application. Specimens were cross-sectioned perpendicularly or horizontally to the resin-dentin interface and embedded in epoxy resin. Their surfaces were polished. The microhardness of the resin-dentin bonding area was measured with a nano-indentation tester. The hardness values at a point of 10 microns distant from the interface in the direction of the dentin differed between systems. It appeared that this was influenced by the presence of the decalcified dentin not impregnated by resin, differences in the chemistry forming the hybrid layer, and the composition of the bonding resin. The hardness of the dentin-bonding interface and its surroundings was determined, and these areas were observed using SEM. Three layers were confirmed the healthy dentin layer, the composite resin layer, and the hybrid layer, (in which decalcified dentin impregnated by resin and that not impregnated by resin are considered to be mix). In the hybrid layer, no impression was found by SEM although the hardness in the bonding interface was significantly different. These layers appear to be more elastic and softer than the healthy dentin. PMID- 10522178 TI - Implant supported single-tooth replacements compared to contralateral natural teeth. Crown and soft tissue dimensions. AB - The aim of this study was to make a comparative evaluation of crown and soft tissue dimensions between implant-supported single-tooth replacements and the contralateral natural tooth. Twenty patients, who had been treated with an implant-supported single-tooth replacement in the esthetic zone of the maxillary jaw and had i) a non-restored contralateral natural tooth and ii) completed the implant-supported crown restoration at least 6 months prior to the scheduled follow-up examination, were included in the study. At the re-examination various variables describing crown form, soft tissue dimensions and soft tissue conditions were assessed. In addition, the patient's overall satisfaction with the esthetic outcome of the implant-supported single crown was scored using a Visual Analogue Scale (VAS). In 12 of the subjects clinical photographs were available from the time of crown insertion for evaluation of longitudinal alterations of the papilla height. The results revealed that, in comparison to the contralateral natural crown, the implant supported crown i) was longer, ii) had a smaller facio-lingual width, iii) was bordered by a thicker facial mucosa, iv) had a lower height of the distal papilla, v) showed a higher frequency of mucositis and bleeding on probing and vi) showed greater probing depths. The longitudinal evaluation of the papillae adjacent to the implant crown showed an improved proximal soft tissue fill at the follow-up examination. The VAS scoring of the patients' satisfaction with the appearance of their single implant supported restorations revealed a median value of 96% with a range from 70 to 100%. Hence, observed differences in clinical crown height and soft tissue topography between implant-supported single-tooth replacements and the contralateral natural tooth may in most patients be of minor importance for the appreciation of the esthetic outcome of implant therapy. PMID- 10522179 TI - Resolution of peri-implantitis following treatment. An experimental study in the dog. AB - The aim of the present experiment was i) to study the effect of anti-microbial therapy of experimentally induced peri-implantitis lesions and ii) to assess features of bone regrowth following treatment. Four beagle dogs were used. Three titanium fixtures (Branemark System) were installed in each quadrant of the mandible (premolars previously extracted). Abutment connection was performed 5 months later and ligature induced breakdown was initiated. The ligatures were removed when approximately 50% of the initial bone support was lost. A 3-week antibiotic regimen (amoxicillin and metronidazole) was initiated 1 month later. Two days after the start of the antibiotic administration, the experimental implant sites were exposed to local therapy. The abutments were removed and the exposed fixture surfaces were treated with an abrasive (pumice) administered via a rotating brush (left side) or cleaned with cotton pellets soaked in saline (right side). Cover screws were attached to the fixtures and the implants were submerged. Fluorochromes were injected intravenously 2 weeks, 4 weeks and 12 weeks after surgery. The animals were killed 7 months after surgery and block biopsies of each implant site were dissected and prepared for histological analysis. The findings of the examinations disclosed that the inflammatory lesion was resolved and new bone formation had occurred in the previous defect following antimicrobial and local therapy. The amount of "re-osseointegration" that had taken place, however, was small. Indeed, at all experimental implant sites, a thin connective tissue capsule was found to separate the implant surface from the newly formed bone. PMID- 10522180 TI - Chemical treatment of machined titanium surfaces. An in vitro study. AB - Microbial plaque accumulation on titanium dental implant surfaces can result in an inflammatory condition of the surrounding tissues. Cleaning of such a contaminated surface, in vivo, by means of a solution of amino-alcohol, following surgical exposure, has been proposed. However, the tissue healing following treatment resulted in formation of a fibrous capsule at the tissue-implant interface, i.e. improper implant re-integration. The present experiment was designed to investigate the possible influence of an amino-alcohol solution on machined titanium surface properties. Titanium samples with topography and chemical composition similar to the clinically used Branemark implant surfaces were used in this experimental in-vitro study to investigate the adsorption of amino-alcohol to such surfaces, and the possibilities to chemically remove the adsorbed alcohols in order to recover a pristine titanium surface. The amino alcohol solution was supplied to the sample surfaces and four different methods were subsequently used in order to remove the adsorbed alcohol molecules. It was shown that rinsing in water, saline solution, and 5% H2O2 did not remove the amino-alcohol from the surface. However, exposure to ozone produced by using a commercial mercury lamp in ambient air resulted in complete removal of the adsorbed amino-alcohol. The results show that the amino-alcohol used forms a stable and dense film at the implant surface in vitro. Presence of such a film most likely prevents re-integration to occur at the implant-tissue interface in vivo. PMID- 10522181 TI - BMP stimulation of bone response adjacent to titanium implants in vivo. AB - The objective of this study was to evaluate the effect of bone morphogenetic protein (BMP) on the bond strength of titanium (Ti) implants at the bone-implant interface. Thirty-six Branemark screw implants (3.75 mm diameter by 15 mm long) were implanted for 3 and 12 weeks. At 3 weeks after implantation, the mean reverse torque value for implants stimulated with BMP delivered using an atelopeptide type-I collagen carrier (74.2 +/- 5.2 N cm) was observed to be statistically greater (P < 0.000016) than the mean reverse torque value for control Ti implants (32.8 +/- 1.1 N cm). Similarly, at 12 weeks after implantation, the mean reverse torque value for implants stimulated with BMP (89.2 +/- 2.7 N cm) was statistically greater (P < 0.0042) than the mean reverse torque value for control Ti implants (75.8 +/- 2.4 N cm). In addition, histomorphometric evaluations indicated more bone contact with the BMP stimulated implant surfaces compared to the controls after 3 weeks implantation. It was concluded from this study that the use of BMP-atelopeptide type-I collagen mixture is an effective means of obtaining greater bond strength at the bone implant interface within a shorter time period than Ti implants without BMP. PMID- 10522182 TI - Three-dimensional bone structure around hydroxyapatite and titanium implants in rabbits. AB - Long-term support of dental implants requires adequate bone thickness in the area surrounding the implant. A three-dimensional examination, including calculations of percent bone-implant contact and percent bone volume, was conducted to clarify the bone structure around pure titanium (Ti) and dense hydroxyapatite (HA) implants. The implants were installed in the tibiae of rabbits, and the bone structure was examined after 2, 4, 6 and 8 weeks. The bone formation following implantation differed in the cortical bone and cancellous bone areas. In the cortical bone area, the percent bone-implant contact and percent bone volume were comparatively consistent for both Ti and HA implants during the observation period. In the cancellous bone area, both findings were influenced by the implantation period, and the chronological bone structure in the cancellous bone area also differed between Ti and HA implants. The percent bone-implant contact and percent bone volume in the Ti implant increased over 8 weeks, whereas the dense HA implant increased for the first 4 weeks and then decreased. The implant materials, Ti and HA, affected the bone remodeling in the cancellous bone area. PMID- 10522183 TI - Comparison of bioabsorbable and bioinert membranes for guided bone regeneration around non-submerged implants. An experimental study in the mongrel dog. AB - The aim of this clinical investigation was to evaluate the effect of guided bone regeneration around non-submerged implants using different barrier membranes. Five adult mongrel dogs were used in this investigation. After having all premolars extracted and implant osteotomies performed in the regions of the former premolars, buccal bone defects were created. Subsequently, 3 implants were placed and the defects treated with 1 of the following 3 modalities: a) guided bone regeneration using an expanded polytetrafluoroethylene membrane, b) guided bone regeneration using a bioabsorbable membrane made from a synthetic copolymer of glycolide and lactide and c) no membrane application. Following implant and membrane placement, the mucoperiosteal flaps were repositioned and tightly sutured around the neck of the implants allowing for a non-submerged healing. After a healing period of 6 months, the animals were sacrificed and the specimens processed for histologic evaluation. The clinical pre-treatment defects between the different treatment groups were not statistically different (bioinert membrane group: 4.9 mm; control group: 4.8 mm; bioabsorbable membrane group: 4.5 mm). The remaining histological defects after 6 months of healing amounted to approximately 2.5 mm in the bioinert membrane group, 5.7 mm in the control group and 6.0 mm in the bioabsorbable membrane group. A significant difference was observed between the bioinert membrane group and the other 2 groups. The mineralized bone-to-implant contact in the bioinert membrane group was 51.5%, in the control group 46.3% and in the bioabsorbable membrane group 37.5%. The values between the bioinert membrane group and the bioabsorbable membrane group were statistically different. The results of this study indicate that bone regeneration with bioinert e-PTFE membranes around non-submerged implants is possible. The utilized absorbable polyglycolic/polylactid membrane did not show any bone regenerative effect and the results did not differ from the control group without membrane application. PMID- 10522184 TI - Implant-retained mandibular overdentures compared with complete dentures; a 5 years' follow-up study of clinical aspects and patient satisfaction. AB - The aim of this prospective randomized controlled clinical trial was to evaluate and compare clinical aspects and satisfaction during the first year following treatment and consecutively the change in treatment during the next 4 years of follow-up. Patients were allocated to one of the following treatment modalities: an implant-retained overdenture (IRO-group, 2 endosseous implants, n = 61) or a complete denture (CD-group, n = 60). One year after placement of the denture, unsatisfied patients of the CD-group got the opportunity for a retreatment including an implant-retained overdenture. In the IRO-group 4 implants were lost during the first year and again 4 implants were lost during the next 4 years (survival rate: 93%). All patients could be re-operated successfully. In the CD group 14 patients (23%) chose an implant-retained overdenture after 1 year. Patients of the IRO-group were significantly more satisfied than patients of the CD-group after 1 year (satisfaction score 8.3 versus 6.6, scale 1-10) and after 5 years (7.4 versus 6.4). From this study it can be concluded that endosseous implants have a high survival rate after 5-years' follow-up. Satisfaction score of the IRO-group is diminishing in time, probably because patients get used to an improved situation. After 5 years, the mean satisfaction score of the CD-group (including patients who got implants) was still lower than of the IRO-group, in spite of the opportunity to a retreatment and have implant-retained overdentures. PMID- 10522185 TI - The tooth-worm: historical aspects of a popular medical belief. AB - The concept of a tooth-worm, which according to popular belief, caused caries and periodontitis, has existed in diverse cultures and across the ages. During the Enlightenment, however, the theory of the tooth-worm was assigned by medical doctors almost exclusively to superstition. Even so, the idea that toothache was caused by gnawing worms held on even into this century. There were many different ideas with regard to the appearance of tooth-worms. In England, for instance, it was thought that the tooth-worm looked like an eel. In Northern Germany, people supposed the tooth-worm to be red, blue, and gray and in many cases the worm was compared to a maggot. The gnawing worm was held responsible for many evils and, in particular, was blamed for toothache provoked by caries. The question is discussed of how the belief in the existence of the tooth-worm in former times can be explained. In popular medicine, numerous therapies were applied in order to eradicate the tooth-worm. In addition to the fumigations with henbane seeds, which allowed the "tooth-worm" to develop in the form of burst seeds, there were also magical formulas and oaths. PMID- 10522187 TI - Fatigue behaviour of different dentin adhesives. AB - The aim of this in vitro study was to compare quasistatic and cyclic fatigue dentin bond strength of modern adhesive systems representing different generations. One hundred and fifty cavities were made in discs of freshly extracted human third molars and filled with direct resin composite restorations. Dentin adhesives of different generations (SY = Syntac Classic, multi-step system with self-etching primer; SE = Syntac Classic with additional phosphoric acid etching prior to application of the self-etching primer; SB = Scotchbond Multi Purpose Plus, multi-step system with total etching; PE = Prime & Bond 2.1, single step system with and without [PB] total etching) were used in combination with one hybrid composite. After 21 days of storage, 10 specimens for each adhesive system were subjected to thermocycling (1150 cycles) for 24 h and were afterwards debonded in a push-out test. Another 20 specimens were tested with cyclic fatigue according to the staircase method with 5000 cycles for each specimen. Static and cyclic push-out bond strengths, respectively, for each group were (MPa): SY 16.9 +/- 0.9 and 14.2 +/- 1.7, SE 17.5 +/- 1.8 and 14.8 +/- 3.4, SB 18.5 +/- 1.7 and 13.9 +/- 2.1, PB 14.6 +/- 2.2 and 7.2 +/- 2.4, PE 13.4 +/- 2.2 and 6.8 +/- 1.8. Both quasistatic and dynamic bond strengths revealed better values for the multi step systems (P < 0.05). All adhesive systems tested revealed a significant fatigue behaviour which was more pronounced for the one-bottle system with a decrease of about 50% independent of additional dentin etching. PMID- 10522186 TI - Salivary mutans streptococci and lactobacilli in 9- and 13-year-old Italian schoolchildren and the relation to oral health. AB - The prevalence and levels of mutans streptococci (MS) and lactobacilli (LB) in saliva and its possible correlation with dental caries and periodontal conditions was investigated in 473 Italian schoolchildren, 9 and 13 years of age. A clinical examination and sampling of stimulated whole saliva was carried out in the school and oral health was assessed as DMFT and CPITN using the WHO criteria. The saliva samples were frozen in liquid nitrogen and after thawing, cultivated on selective media. To test the influence of cryopreservation, fresh samples from 20 subjects were cultivated. Thirty-five percent of the children were caries-free with a mean DMFT of 1.9 at the age of 13. The majority exhibited healthy periodontal conditions. Salivary MS and LB were identified in 52% and 21% of the children, respectively. The prevalence of MS was higher among the 13-year-olds than the 9 year-olds while no such difference was found regarding LB. There was a statistically positive relationship (P < 0.01) between the levels of MS and LB and both were significantly correlated to caries (P < 0.01). The correlation coefficient of microbial recovery between frozen and unfrozen samples was 0.99. In conclusion, the data provided cross-sectional information of a clear positive relationship between selected micro-organisms in saliva and caries in 9- and 13 year-old children in spite of a relatively low prevalence of the disease. The findings are discussed in a risk selection perspective. PMID- 10522188 TI - Transverse changes after surgical closure of complete cleft lip, alveolus and palate. AB - Surgery for patients with unilateral (UCLP) and bilateral (BCLP) complete cleft lip, alveolus and palate has a considerable influence upon craniofacial growth. With respect to this, the cleft team at Hannover Medical School has attempted to reduce necessary surgical interventions to labioplasty, palatoplasty and veloplasty. Still, the effects of these operations influence maxillary growth to an extent which requires orthodontic treatment in all patients. This study focuses upon the transverse alterations of the alveolar arch and the deciduous dentition after lip and palate surgery. Dental casts prior to any surgical intervention and after labioplasty and complete palaotoplasty of the hard and soft palate were measured for transverse changes by using anatomical landmarks. The results indicate a significant occurrence of anterior relative to posterior arch width loss for both UCLP and BCLP patients. Orthodontic treatment should be planned and performed with respect to these findings in order to support craniofacial growth and prevent maxillary dental arch deficiency. PMID- 10522189 TI - Cytopathologic effects of arecoline on human gingival fibroblasts in vitro. AB - Arecoline, a major betel nut alkaloid, has been detected in saliva obtained during betel nut chewing in concentrations up to 140 micrograms/ml, corresponding to 0.9 mM. Arecoline in the millimolar concentration range might participate in the initiation and/or progression of periodontal disease during the long-term effects of betel nut chewing. In this study, cell growth, cell proliferation, assessment of cytoplasmic enzyme lactate dehydrogenase (LDH) and collagen synthesis were used to investigate the effects of human gingival fibroblasts exposed to arecoline levels of 0-200 micrograms/ml. Control culture exhibited a normal monolayer of long spindle-shaped fibroblast morphology. Arecoline-treated human gingival fibroblasts showed a more rounded appearance and detached at the higher concentrations. At concentrations higher than 75 micrograms/ml, many cells had detached from the surface of the petri dish and numerous floating cells could be seen under the inverted microscope. At a concentrations higher than 25 micrograms/ml, arecoline inhibited cell growth, proliferation and collagen synthesis and increased LDH leakage in a dose-dependent manner (P < 0.05). These results indicate that arecoline is a cytotoxic agent to human gingival fibroblasts. Repeated and long-term exposure to arecoline could impair gingival fibroblast function. Betel quid chewers might be more susceptible to destruction of the periodontium and less responsive to a regeneration procedures during periodontal therapy. PMID- 10522190 TI - Tooth-colored restorations of posterior teeth in German dental education. AB - Optimizing the quality of tooth-colored restorations is one of the main topics of current research. But there is only little information available about university education in this field. The aim of this study was to collect and evaluate data about the different aspects of dental education in Germany concerning tooth colored restorations. Based on the response to a questionnaire from 90% of all German dental schools in the fall of 1997 a detailed survey is given of the utilization, indications, practical procedure, problems and limitations of both direct and indirect tooth-colored restorations done by students. The results indicate a wide-spread use of directly inserted composite for posterior teeth in the different education programs. Indeed, the preferred preparation of the cavity margin differs from school to school. Rebuilding an adequate proximal contact and a precise fit at the gingival margin are looked upon as the main problems of class II composite fillings. Ceramic inlays are mainly inserted by students in advanced clinical courses with the insertion procedure being claimed as the main problem of this technique. The findings of this study mostly show the same limitations and difficulties of tooth-colored restorations in education as found by research. Partly different teaching concepts are reflected in the differing scientific results. PMID- 10522191 TI - Caries pattern and oral health habits in 2- to 6-year-old children exhibiting differing levels of caries. AB - The aim of the present study was to describe in detail the distribution of caries lesions among tooth types and surfaces in the primary dentition of young children with different levels of disease. A total of 125 children (between 2 and 6 years old) attending the pediatric dental clinic of the University Hospital of Leuven was allocated to three groups based on their caries experience: dmft = 1-5 formed the low caries experience group (LC, n = 27), dmft = 6-9 the moderate caries experience group (MC, n = 61) and those with dmft > = 10 the high caries experience group (HC, n = 37). Caries experience (at patient level, tooth and tooth surface level) and oral hygiene were evaluated. Oral health habits were recorded using a questionnaire (completed by parents). Caries lesions were not evenly distributed among different tooth types. Teeth most frequently affected were lower (first and second) primary molars. Primary incisors were rarely found to be carious. The distribution of the lesions followed a comparable pattern, irrespective of the disease level of the child. Decay on primary canines and buccal/lingual surfaces of primary molars was strongly indicative of a high caries experience. There was a clear link with reported oral hygiene habits and the use of a pacifier and baby bottle with sugared content. PMID- 10522193 TI - Large band spectral analysis and harmful risks of dental turbines. AB - Previous studies have attributed harmful effects to ultrasonic frequencies and recently many have addressed the problem of early deafness among dentists caused by high-speed air turbines. In this study, we measured the spectra of the sounds generated by high-speed air turbines, ranging from audible to ultrasonic frequencies (0-70 kHz). The hypothesis advanced is that the ultrasound spectrum of high-speed air turbines reaches amplitudes that may be noxious. We performed continuous analyses of the spectra of three bands of turbines (n = 17). Measurements of frontal incidence were made using a Bruel & Kjaer microphone and sonometer. For spectral analysis we used the autoregressive parametric method. Using Akaike's criteria the model weights were fixed at 12. The method used in the present work led to an accuracy in the results of 10(-2) kHz within the same brand of turbine. Results show, in terms of frequencies, the presence of four main peaks: 5.6 kHz +/- 0.73 in the audible range, and 20.1 kHz +/- 2.16, 35.7 kHz +/- 2.56 and 46.5 kHz +/- 0.71 in the ultrasonic range. In a normalized spectrum, the amplitude of the ultrasonic component reaches 115 dBspl for 46.5 kHz and is 76% greater than that of the audible component. Such values, both in terms of frequencies and amplitude, reach levels which may provoke short- or long term negative physiological disturbances and hearing-damage risk. Further research should be directed to determine to what extent they might induce noxious effects for the dental team. PMID- 10522192 TI - Cytocompatibility of periodontal dressing materials in fibroblast and primary human osteoblast-like cultures. AB - Purpose of this investigation was to determine the cytocompatibility of various periodontal dressing materials by means of human primary gingival fibroblasts (HGF), human osteoblast-like cells (HObl) derived from the alveolar bone, and permanent 3T3 mouse fibroblasts (3T3). Cell culture medium extracts (time periods of extraction: day 1 and between day 2 and day 8 after setting) as well as solid specimens of the following materials were investigated: Coe-pak, Voco pac, Peripac, and Barricaid. Responses of cultures exposed for 24 h and 48 h to these materials were monitored by the fluorescent dyes H33342 and sulforhodamin 101 as well as by light microscopy. It was found that most extracts of Voco pac, Peripac, and Barricaid did not inhibit growth of HGF. Coe-Pak, however, clearly reduced the proliferation of HGF compared to untreated controls. Peripac decreased growth of HObl whereas Coe-Pak, Voco pac, and Barricaid caused no cytotoxic alterations in any of the test assays. Contrary to HGF and HObl, 3T3 cells were much more irritated by the test materials. But the light-curing resinous material Barricaid reduced proliferation of 3T3-fibroblasts only slightly. Our data indicate that Barricaid is exceedingly cytocompatible, whereas all other materials revealed moderate or severe cytotoxic effects according to the cell type. PMID- 10522194 TI - The pneumatic spaces of the temporal bone: relationship to the temporomandibular joint. AB - OBJECTIVES: To assess the topographic relationship between the pneumatic spaces of the temporal bone and the temporomandibular joint (TMJ) using high-resolution CT. METHODS: Findings from 100 consecutive patients who had undergone high resolution axial CT of the base of the skull were reviewed on a digital imaging workstation. Additional multiplanar reformatted images were created in the sagittal and coronal planes through the TMJ. The extension of the pneumatic spaces of the temporal bone and their relation to the TMJ were determined on both sides. RESULTS: The extent of pneumatisation of the temporal bone varied considerably. The roof of the TMJ fossa was pneumatised in 51 patients. The articular eminence contained air spaces in 12 patients, the root of the zygomatic process in five patients. Air cells in the peritubal area extended into the medial wall of the glenoid fossa 53 patients. In approximately 25% the extent of pneumatisation showed marked asymmetry. CONCLUSIONS: Pneumatisation of the temporal bone frequently extends close to the TMJ. Knowledge of these air spaces is helpful for the interpretation of imaging studies such as panoramic radiographs and to understand the spread of pathological processes into the joint. PMID- 10522196 TI - Effects of developer exhaustion on the sensitometric properties of four dental films. AB - OBJECTIVES: To examine the effects of exhaustion of five different processing solutions on the sensitometric properties of four dental X-ray films: Ektaspeed Plus and Ultra-speed (Kodak Eastman Co. Rochester, USA) and new and previous Dentus M2 (Agfa-Gevaert, Mortsel, Belgium). METHODS: An aluminum stepwedge was used to construct characteristic curves for the four films. All films were processed manually using three sets of chemicals for manual processing: Agfa (Heraeus Kulzer, Dormagen, Germany), Kodak (Kodak-Pathe, Paris, France) and Demat (Viscopac, Athens, Greece) and two sets for automatic processing: Durr XR and Periomat (Durr Dental, Bietigheim-Bissingen, Germany). Film speed and gradient were evaluated until the chemicals were completely exhausted. An analysis of variance was performed separately for each set of chemicals for manual and automatic processing. RESULTS: Ektaspeed Plus was the fastest film in the manual processing solutions. The new Dentus M2 and Ektaspeed Plus films had similar speed using the chemicals for automatic processing. Ultra-speed had the lowest speed in all solutions, but it had the greatest consistency. Exhaustion of the developer caused a comparable decrease in speed of Ektaspeed Plus and the two Dentus M2 films. In fresh chemistry Agfa was the strongest manual processing solution, but it had the highest exhaustion rate. The Durr XR chemical was stronger than Periomat over the whole test period. CONCLUSIONS: The combination of film and processing solution is an important factor for achieving constant sensitometric properties. Ektaspeed Plus and the new Dentus M2 film should be used in dental practice, as they require lower exposure and have equivalent or superior properties compared with Ultra-speed. PMID- 10522195 TI - Sensitometric evaluation of four dental X-ray films using five processing solutions. AB - OBJECTIVES: To compare the sensitometric properties of four dental films: Ektaspeed Plus and Ultra-speed (Kodak Eastman Co, Rochester, USA) and new and previous Dentus M2 (Agfa-Gevaert, Mortsel, Belgium) in five different processing solutions. METHODS: Characteristic curves were constructed for four dental X-ray films using five different processing solutions. All films were processed manually in three sets of chemicals for manual processing: Agfa (Heraeus Kulzer, Dormagen, Germany), Kodak (Kodak-Pathe, Paris, France) and Demat (Viscopac, Athens, Greece) and two sets of chemicals for automatic processing: Durr XR and Periomat (Durr Dental, Bietigheim-Bissingen, Germany). Analysis of variance and regression analysis was performed to examine the factors significantly affecting film density. RESULTS: The new Dentus M2 film had comparable gradient, higher speed and lower base-plus-fog than its predecessor. It had comparable speed with Ektaspeed Plus in chemicals for automatic processing. All films had a higher speed and lower gradient when processed in the chemicals for automatic processing. The highest film speed was achieved using Durr XR chemicals. CONCLUSIONS: The new Agfa Dentus M2 film is an E-speed film and can be considered an alternative to Ektaspeed Plus: both can be recommended for use in dental practice as they contribute to dose reduction. PMID- 10522197 TI - Cleidocranial dysplasia: radiological appearances on dental panoramic radiography. AB - OBJECTIVE: To report the effectiveness of dental panoramic radiography in identifying features pathognomonic for cleidocranial dysplasia. METHODS: Panoramic radiographs of nine male Caucasian patients with cleidocranial dysplasia are analysed. RESULTS: In addition to the established dental complications of failure of eruption of the permanent dentition and multiple supernumerary teeth, morphological abnormalities of the maxilla and mandible, particularly in the ascending ramus and coronoid process were present. CONCLUSION: Dental panoramic radiography is a valuable adjunct in confirming the diagnosis of cleidocranial dysplasia. PMID- 10522198 TI - Dental students' ability for three-dimensional perception of two-dimensional images using natural stereopsis: its impact on radiographic localization. AB - OBJECTIVES: To investigate the ability of dental students to perceive 2D images in 3D using natural stereopsis and to evaluate whether the use of stereovision influences their interpretation of radiographic depth, with particular reference to intra-oral radiographs of third molars compared with scanograms. METHODS: Seventy-two dental students (37 from sixth semester and 35 from fourth semester) who had all completed a course in the use of the buccal-object rule participated in the study. Lectures in 3D perception were given to the sixth semester students. They were trained in 3D perception using a 'single image colour field' stereogram and classical stereograms of circles. Both semester students diagnosed nine scanograms and nine intra-oral radiographs for: (1) the position of the crown of an impacted third molar in relation to the second molar and (2) root deviations in the bucco-lingual plane. The sixth semester students recorded whether they were able to achieve a 3D image in these cases. RESULTS: Five students could not achieve a depth image from the training stereograms. Fifty-six per cent agreed on which two objects in the circles were nearest to the viewer and which were most distant, whereas 38% agreed on the opposite. Root deviation was a more difficult diagnostic task than crown position, and the sixth semester students made significantly more correct diagnoses than the fourth semester students (P < 0.001). Irrespective of semester, there was no significant difference between number of correct diagnoses of crown alignment in intra-oral radiographs and scanograms (P = 0.57), whereas root deviations were diagnosed significantly more accurately from the intra-oral radiographs (P < 0.001). CONCLUSIONS: Most dental students can be taught to use natural stereovision to achieve a 3D image from two radiographs at different angles. 3D image is achieved equally well from intra-oral radiographs with a free-hand tube shift and scanograms. Those students who achieved a 3D image in a particular case, made more correct diagnoses than those who could not. PMID- 10522199 TI - Predictability of a three-dimensional planning system for oral implant surgery. AB - OBJECTIVES: To compare 2D CT alone with 2D + 3D reconstruction for pre-operative planning of implant placement. METHODS: Spiral CT scans of 33 consecutive patients were used for both reformatted 2D and 3D computer-assisted planning. The number, site and size of implants and the occurrence of anatomical complications during planning and implant placement were statistically compared using the percentage agreement and the Kendall's correlation coefficients (tau). Although planning was performed in 33 patients, implants were only placed in 21 patients. In 11 patients surgery was based on 2D + 3D imaging and in ten patients on 2D planning. RESULTS: Agreement between planning and placement of implants was highly significant for the implant sites selected. For 2D based planning and placement, agreement reached 68% (tau = 0.94). For 2D + 3D based planning and placement, agreement attained 73% (tau = 0.89). For planning and placement of implant size based on 2D images, agreement was 31% and not significant (tau = 0.23). When based on 2D + 3D images, agreement for implant size was 44% (tau = 0.5). Agreement was not significant for anatomical complications: 69% for 2D planning and 71% for 2D + 3D planning (tau = 0.24 for 2D and tau = 0.21 for 2D + 3D). CONCLUSIONS: The 3D planning system is a reliable tool for pre-operative assessment of implant placement. Both 2D and 2D + 3D planning have a good predictability for the number and site of the implants but less so for anatomical complications. However, the 2D + 3D planning provides a better pre-operative assessment of implant size. PMID- 10522200 TI - A simplified method to obtain perceptibility curves for direct dental digital radiography. AB - OBJECTIVES: To derive and test a simplified method to construct Perceptibility Curves (PCs) for dental digital detectors. METHODS: Mathematical expressions were derived to make it possible to construct PCs from viewer data obtained at two exposures, one low and one high. PCs were constructed applying these expressions and compared with data previously obtained employing the conventional method. RESULTS: PCs constructed according to the simplified method agree extremely well with conventionally obtained data. CONCLUSIONS: Reliable PCs may be constructed according to the simplified method. PMID- 10522201 TI - Zygomatic air cell defect (ZACD). Prevalence and characteristics in a dental clinic outpatient population. AB - OBJECTIVES: To determine the prevalence and characteristics of zygomatic air cell defect (ZACD) among a general dental clinic population. METHODS: The panoramic radiographs of 2734 dental clinic outpatients were examined for the presence of ZACD. ZACD was defined as a nonexpansile, nondestructive cyst-like radiolucency in the zygomatic process of the temporal bone which appears similar to the mastoid air cells and which does not extend further anteriorly than the zygomaticotemporal suture. RESULTS: ZACD was found in 40 patients (1.5%) with a mean age of 49.6 (s.d. 18.0) years. Twenty cases (50%) each occurred in males and females. Meta-analysis of three large case series comprising 4579 patients revealed a total of 76 cases of ZACD (1.7% prevalence) occurring over an age range of 15-83 years. Thirty-four (44.7%) occurred in males while 42 (55.3%) occurred in females. Bilateral ZACD were found in 17 patients (22.4%). CONCLUSIONS: ZACD is not a rare anatomical variant, and clinicians planning eminectomy or other surgical procedures involving the zygomatic arch are advised to obtain appropriate presurgical imaging studies to avoid the need for creative intra-operative reconstruction. PMID- 10522202 TI - Development of a system for craniofacial analysis from monitor-displayed digital images. AB - OBJECTIVES: To develop a program for clinical craniofacial analysis directly on monitor-displayed digital images. METHODS: The program was developed in VisualBasic (Microsoft, Redwood WA, USA), using Access (Microsoft) database, for landmark identification analysis and data storage. It runs on Windows95 (Microsoft). RESULTS: The following established cephalometric analyses were included in the program: Bjork, Down, Frontal, Jarabak, Occlusogram, Rickett and Tweed. In addition, an orthodontist can simply define his own analyses. If he wants to identify a particular landmark, he clicks on its name at the top of the screen, and the definition including a drawing will be shown. If specified in the analysis selected, reference lines or templates are drawn using the identified landmarks. Soft tissues can be drawn either freehand or using the 'soft tissue detection' that generates an outline from the grey-level histogram. The program has been implemented at the Royal Dental College and at the Department of Maxillofacial Surgery and at the Cleft Palate Centre, Aarhus University Hospital. CONCLUSION: A program (PorDiosW) has been developed which has facilitated cephalometric analysis from digital lateral and frontal skull radiographs and digital photographs. The program has been successfully applied in patients with cleft lip and palate and other developmental craniofacial anomalies before and after surgery. PMID- 10522203 TI - Molecular pathogenesis of root dentin caries. AB - Human dentin has a higher content of organic matrix and more non-ideal hydroxyapatite than human enamel. Ultrastructural studies indicate that root caries involves both mineral dissolution and breakdown of the organic matrix. Factors involved in the root caries process seem more complicated than those in enamel caries. Moreover, the distinct roles of acids and enzymes and the sequence of events in the root caries process are not well-understood. Although Streptococcus mutans and Actinomyces viscosus are considered to be major pathogenic micro-organisms of root caries, their roles in degradation of the organic matrix components of root dentin need clarification. The purpose of this paper is to review the basic composition of root dentin and the roles of acids and both endogenous and bacterial enzymes in the root caries process. PMID- 10522204 TI - Influence of lead on the cariostatic effect of fluoride co-crystallized with sucrose in desalivated rats. AB - OBJECTIVE: Results from previous studies have shown that pre- and perinatal exposure to lead enhances susceptibility of rats to development of dental caries. A possible explanation for this phenomenon may be that lead complexes with fluoride and renders F insoluble and unable to exert its cariostatic effects. MATERIALS AND METHODS: Thus, to explore this hypothesis, 48 desalivated Sprague Dawley rats were placed in a Konig-Hofer programmed feeder and received 17 meals of powdered sucrose daily, and water ad libitum as follows: group (1) plain sucrose and sterile distilled water (SDW); (2) sucrose containing 15 ppm F and SDW; (3) sucrose containing 15 ppm F and 10 ppm Pb water; (4) sucrose containing 15 ppm F and 25 ppm Pb water. RESULTS: The highest smooth-surface, sulcal surface caries and severity scores were observed in group I. Animals that were exposed to fluoride showed reduced smooth-surface caries and severity scores. S. sobrinus counts did not differ among the groups. CONCLUSION: Lead did not interfere with the protective effect of fluoride in the conditions of the present study. PMID- 10522205 TI - The protein phosphatase inhibitors, okadaic acid and calyculin A, induce apoptosis in human submandibular gland ductal cell line HSG cells. AB - OBJECTIVE: To investigate a possible relationship between protein phosphorylation or dephosphorylation status and apoptosis in salivary gland cells, we examined the effects of okadaic acid and calyculin A, the protein phosphatase inhibitors, on cultured human submandibular gland ductal cell line, HSG cells. METHODS: HSG cells at subconfluent stages were exposed to varying concentrations of okadaic acid or calyculin A. Apoptoses were analysed in HSG cells by phase-contrast microscopy, WST-1 cytotoxicity assay, Hoechst 33342 staining, and DNA ladder formation. RESULT: Both okadaic acid and calyculin A induced cell death in HSG cells in a dose-dependent fashion. Marked nuclear condensation and fragmentation of chromatin was observed in HSG cells. DNA ladder formation was also detected in HSG cells by treatment with okadaic acid or calyculin A. The induced DNA ladder formation was dose-dependent with maximal effect at concentrations of 50 nM okadaic acid and 2 nM calyculin A, respectively, and were time-dependent from 14 h to 48 h. To further determine if new gene transcription and protein synthesis regulate okadaic acid-induced apoptosis in HSG cells, the cells were treated with cycloheximide or actinomycin D in the presence of 20 nM okadaic acid. Neither inhibitor protected the cells against okadaic acid-induced apoptosis. CONCLUSION: Based on the known selectivity of okadaic acid and calyculin A, our results indicate that the pathway of the apoptosis in the cultured salivary gland cells is regulated by protein phosphatase type 1 or type 2A. Our results also suggest that new protein synthesis and/or mRNA expression are not involved in okadaic acid-induced apoptosis in HSG cells. PMID- 10522206 TI - An immunohistochemical study of keratin expression in ameloblastoma from a Kenyan population. AB - OBJECTIVES: Ameloblastomas appear to exhibit biological heterogeneity and, except in the case of malignancy, histological appearances that do not always allow their behaviour to be predicted. The aim of this study was to assess keratin expression in African ameloblastomas and to correlate this with their clinical and histological features. MATERIALS AND METHODS: Expression of simple keratins 7, 8, 18 and 19; cornification keratins 1 and 10; basal and differentiation keratins 5 and 14 and hyperproliferation-related keratins 6 and 16 in 14-39 cases of ameloblastoma was assessed by immunohistochemical methods. RESULTS: There was patchy expression of keratin 7 in the suprabasal and stellate reticulum-like cells in some cases. All cases showed similar weak expression for keratins 8 and 18 in suprabasal and stellate reticulum-like cells but none showed keratin 1 or 10 expression. There was intense expression of keratins 5, 14 and 19 by all tumour cells suggesting that they may retain basal cell characteristics with a potential for proliferation. No consistent relationship was seen between histological types and keratin expression pattern. However, keratins 6 and 16, expressed by suprabasal and stellate reticulum-like cells, showed a marked variation within and between cases, with the highest levels of expression in squamous strands. CONCLUSIONS: We propose that squamous strands may represent the sites of most active growth within individual tumours and expression of keratins 6, 16 and 19 may be predictors of rapid growth. There is a need for further investigation of this in longitudinal clinical studies. PMID- 10522207 TI - Apoptosis-induced and -suppressed cells in salivary gland adenoid cystic carcinoma: correlation with histological growth patterns. AB - OBJECTIVE: It is well known that adenoid cystic carcinoma (ACC) arising from salivary glands shows a correlation between prognosis and histological growth patterns. The aim of the study was to evaluate whether three growth patterns of ACC are related to the distributions of apoptosis-induced and -suppressed tumor cells. MATERIALS AND METHODS: We examined 77 cases of ACC including tubular (18 cases), glandular (50) and solid (9) patterns. In order to visualize the apoptotic cells, terminal deoxynucleotidyl transferase (TdT)- mediated dUTP biotin nick end labeling (TUNEL) and avidin-biotin complex staining using Lewis Y (LeY) antibody were applied to paraffin sections. For detection of the apoptosis suppressed cells, immunohistochemistry employing bcl-2 antibody was utilized. RESULTS: Apoptosis index (AI) based on the TUNEL-stained specimens were tubular, 7.0; glandular 2.4; solid, 5.1. In tubular type, apoptotic cells were frequently located in the inner tubular layer rather than the outer layer. Solid type had scattered apoptotic cells in the nests. Bcl-2 expression was found in 61% of tubular, 20% of glandular and 22% of solid cases. The localization of bcl-2 protein was noticed in outer tubular cells, and peripheral cells or undifferentiated cells in solid pattern. CONCLUSIONS: The peculiar distribution of apoptotic cells may result from the various proportions and distinctive arrangement of neoplastic ductal cells and neoplastic myoepithelial cells in ACC. Apoptotic cells and bcl-2 positive apoptosis-suppressed cells may participate in the construction of characteristic histological appearances of ACC. PMID- 10522208 TI - The effect of saliva specimen collection, handling and storage protocols on hepatitis C virus (HCV) RNA detection by PCR. AB - OBJECTIVES: Commercial assays can now be adapted to detect salivary anti hepatitis C virus (HCV) antibodies, increasing the potential of saliva as a non invasive diagnostic specimen suited to surveillance and epidemiological studies. However, current diagnostic algorithms involve confirmation of HCV infection by RT-PCR. Manipulation and storage conditions of serum can influence the stability of viral RNA. This study examined whether varying specimen collection, handling and storage protocols also affected subsequent HCV RNA detection by RT-PCR applied to saliva specimens. METHODS: Whole unstimulated saliva, together with saliva samples collected in two commercially available devices (Salivette and Omnisal) were obtained from 50 HCV seropositive intravenous drug users. The specimens were subjected to a number of handling and storage conditions, including heat treatment and prolonged storage, then examined for HCV RNA by RT PCR using primers derived from the 5' non-coding region (5'NCR). RESULTS: HCV RNA was detected in saliva samples from 25 (50%) of the patients. No single collection device or handling procedure identified all the subjects with HCV RNA positive saliva though whole saliva yielded the greatest number of positive results. CONCLUSIONS: Collection and processing of saliva specimens for RT-PCR analysis is complex. At present, detection of salivary HCV RNA by PCR is not sufficiently sensitive for use as a diagnostic tool in epidemiological studies. PMID- 10522209 TI - Primary Sjogren's syndrome: salivary gland function and clinical oral findings. AB - OBJECTIVE: To evaluate salivary gland function, saliva composition and oral findings in patients with primary Sjogren's syndrome (pSS) subdivided into patients with and without focus score > or = 1 (FS) and/or antibodies to SSA/SSB (AB) as well as in healthy controls. SUBJECTS AND METHODS: Unstimulated (UWS) and chewing stimulated (SWS) whole saliva, and stimulated parotid saliva (SPS) were collected in 16 patients fulfilling the European classification criteria for pSS subdivided into those with FS and/or AB (n = 8) and those without FS and AB (n = 8), and in age-matched (n = 14) and young healthy controls (n = 13). UWS and SWS were analysed for Na+ and K+. SPS was analysed for Na+, K+, statherin, and proline-rich proteins (PRPs). Sicca symptoms, DMFT/DMFS, plaque (PI) and gingival (GI) scores, periodontal pocket depth (PPD), and mucosal status were recorded. RESULTS: The young healthy controls had lower UWS as compared to the aged controls (P = 0.03). However, the aged controls had higher DMFT/DMFS (P < 0.001) and PI, GI and PPD (P < 0.01). Patients with FS and/or AB generally had lower saliva secretory rates than patients without FS and/or AB (P = 0.01 for UWS and SPS) and age-matched healthy controls (P = 0.001). There was no significant difference in the content of Na+ and K+, statherin and PRPs between groups. Patients with FS and/or AB had the highest frequency of oral mucosal changes and higher DMFT/DMFS than patients without FS and/or AB and healthy controls (P < 0.01). However, PI, GI, and PPD did not differ significantly. CONCLUSION: Patients with FS and/or AB had lower salivary secretory rates, higher DMFT/DMFS, and more oral mucosal changes than patients without FS and/or AB. Additionally, data suggest that salivary gland function in healthy individuals do not decrease with age. PMID- 10522211 TI - International Association for Dental Research. Symposium on Noma. Nice, June 1998. PMID- 10522210 TI - Defensin-1, an antimicrobial peptide present in the saliva of patients with oral diseases. AB - OBJECTIVES AND DESIGN: A preceding paper has noted a detection of defensin-1 (HNP 1), a peptide with antimicrobial and cytotoxic properties, in the saliva of patients with oral squamous cell carcinoma. The present study deals with the presence of HNP-1 in the saliva of patients with various oral diseases. METHODS: Whole saliva samples were obtained from the patients. HNP-1 in the saliva was isolated and purified by HPLC and the amino acid sequence of the peptide was determined. The molecular weight of HNP-1 was measured by mass spectrometry. The concentration of HNP-1 in saliva was determined by comparing the height of eluted HNP-1 with that of a synthetic HNP-1 standard. RESULTS: The concentrations of HNP 1 in the saliva of patients with oral lichen planus (n = 5), leukoplakia (n = 4), and glossitis associated with iron deficiency (n = 4) were 8.3 +/- 4.3 micrograms ml-1, 13.2 +/- 7.9 micrograms ml-1, and 11.4 +/- 4.9 micrograms ml-1, (mean +/- s.d.), respectively. These concentrations were significantly higher than those in healthy subjects (0.8 microgram ml-1) (P < 0.01). In contrast, salivary HNP-1 concentrations in patients with glossodynia (n = 4) and oral discomfort (n = 4) were similar to those in healthy subjects. CONCLUSIONS: Since HNP-1 is a non specific defensive peptide present in neutrophils, it may play an important role in the protection against diseases such as oral lichen planus, leukoplakia, and glossitis associated with iron deficiency. PMID- 10522212 TI - Noma (cancrum oris): questions and answers. AB - Noma (cancrum oris) is an infectious disease which destroys the oro-facial tissues and other neighboring structures in its fulminating course. It affects predominantly children aged 2-16 years in sub-Saharan Africa where the estimated frequency in some communities may vary from one to seven cases per 1000 children. The key risk factors are poverty, malnutrition, poor oral hygiene, deplorable environmental sanitation, close residential proximity to livestock, and infectious diseases, particularly measles. Malnutrition acts synergistically with endemic infections in promoting an immunodeficient state, and noma results from the interaction of general and local factors with a weakened immune system as the common denominator. Acute necrotizing gingivitis (ANG) is considered the antecedent lesion. Current studies suggest that evolution of ANG to noma requires infection by a consortium of microorganisms with Fusobacterium necrophorum and Prevotella intermedia as the suspected key players. Without appropriate treatment, mortality rate is 70-90%. Survivors suffer the two-fold affliction of oro-facial disfigurement and functional impairment. Reconstructive surgery of the resulting deformity is time-consuming and financially prohibitive for the victims who are poor. PMID- 10522213 TI - Microbiological understandings and mysteries of noma (cancrum oris). AB - The microbiologic history of noma was reviewed. Studies have associated the disease process with large numbers of fusiform bacilli and spirochetal organisms. In order to study the microbiology of the staging and infection periods of noma 62 Nigerian children, aged 3-14 years, 22 children had acute necrotizing ulcerative gingivitis (ANUG) and were also malnourished, 20 exhibited no acute necrotizing ulcerative gingivitis but were malnourished and 20 were free of acute necrotizing ulcerative gingivitis and in good nutritional state) were evaluated for the presence of viruses and oral microorganisms. The ANUG cases in the malnourished children had a higher incidence of Herpesviridae, the main virus being detected was cytomegalovirus. There were more anaerobic microorganisms recovered, with Prevotella intermedia as the predominant isolate, in the malnourished children as compared to the healthy children. A study of the predominant microflora in active sites of noma lesions was carried out in eight noma patients, 3-15 years of age, in Sokoto State, northwestern Nigeria. Fusobacterium necrophorum was recovered from 87.5% of the noma lesions. Oral microorganisms isolated included Prevotella intermedia, alpha-hemolytic streptococci and Actinomyces spp. which were isolated from 75.0, 50.0 and 37.5% of the patients, respectively. Peptostreptococcus micros, Veillonella parvula, Staphylococcus aureus and Pseudomonas spp. were each recovered from one lesion. All strains were observed to be sensitive to all of the antibiotics tested with the exception of one strain of P. intermedia which showed resistance to penicillin. The pathogenic mechanisms of F. necrophorum as a trigger organism were discussed. The isolation from human noma lesions of F. necrophorum, a pathogen primarily associated with animal diseases, may have important etiologic and animal transmission implications. PMID- 10522214 TI - Living conditions of children at risk for noma: Nigerian experience. AB - The study reported in this paper was carried out in the Northwestern and Southwestern regions of Nigeria, between October 1996 and April 1998. The study examined the possible contributory role of living conditions in the development of acute necrotizing gingivitis (ANG) or noma from oral lesions. Questionnaire data obtained from 42 fresh noma cases seen in the Northwest and four fresh cases seen in the Southwest were examined. In addition 46 cases of advanced ANG from the Southwest were included. The main focus was to compare some of the environmental living conditions of cases with advanced ANG and those with noma in these regions. All the noma and ANG cases were seen in children aged 2-12 years. The level of good oral hygiene practices and general environmental living conditions were significantly higher in the Southwest than in the Northwest. Data also showed that living in close proximity with livestock was significantly higher in the Northwest than in the Southwest (P < 0.05). The environmental living conditions of children in the Northwest were further compounded by poor sanitary faecal disposal practices as well as minimal access to potable water. The overall data indicated that living in substandard accommodations, exposure to debilitating childhood diseases, living in close proximity to livestock, poor oral hygiene, limited access to potable water and poor sanitary disposal of human and animal faecal waste could have put the children in the Northwest at higher risk for noma than the children in the Southwest. These could have been responsible for the higher prevalence of noma in the Northwest than in the Southwest. PMID- 10522215 TI - Noma: public health problem in Senegal and epidemilogical surveillance. AB - Noma or cancrum oris is currently a real public health problem for developing countries. In Senegal, awareness of the disease has led the country to be included in the noma programme initiated by the WHO as early as 1994. The objectives are to evaluate the incidence of necrotizing ulcerative gingivitis (NUG) among children, to evaluate the prevalence of noma and infantile diseases, and to promote prevention strategies among vulnerable populations. Data processing was carried out in two phases: manual processing consisted of checking the questionnaires by nurses, and computer processing started as early as the first collection of data. Noma occurs owing to fever and similar cases. Successfully fighting against malnutrition would allow us to reduce the noma rate. PMID- 10522216 TI - Surgical treatment of noma. AB - In the acute stage of noma the role of surgery is a minor one: wound care and, very occasionally, treatment of haemorrhage. However in patients who survive noma, and develop a mutilated and disabled face (trismus, leakage of saliva, impaired speech), reconstructive surgery may improve their fate significantly. Because of economic and educational reasons reconstructive surgery in noma patients should be performed preferably in their own country. Treatment consists of excision of all scar tissue, correction of the trismus and closure of the tissue defects with local, pedicled or free flaps. Because of the large variety of tissue defects and the many surgical options, systematization and subsequently standardization of the reconstructive surgical approach to patients with the sequelae of noma is needed. PMID- 10522217 TI - The World Health Organization initiative on noma. AB - Oro-facial noma is a worldwide scourge in a context where the World Health Report 1998 gives a global incidence of 140,000 cases, a prevalence in 1997 of 770,000 persons surviving with heavy sequelae. The background and the five steps of the WHO oral health programme to control noma including: (i) ensuring training and awareness on early diagnosis and treatment for each public health structure, (ii) raising awareness and informing populations, (iii) promoting epidemiological research, (iv) promoting aetiological research, (v) setting up an African regional centre for the treatment of after-effects, are developed in this paper. PMID- 10522219 TI - Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis in periodontal disease. PMID- 10522218 TI - Intravascular papillary endothelial hyperplasia of the mouth: report of six cases and literature review. AB - OBJECTIVE: Oral intravascular papillary endothelial hyperplasia (IPEH) is an uncommon, reactive vascular lesion with no specific clinical features. Microscopically two subtypes have been described: a pure and a mixed form. The importance of IPEH is in its resemblance to angiosarcoma. The aim of this study was to evaluate both the prevalence of IPEH in a consecutive series of oral vascular malformations and the clinico-pathological features. MATERIALS AND METHODS: Histological sections of 103 consecutive cases filed as vascular malformations were reviewed for histopathological criteria of IPEH. RESULTS AND CONCLUSION: We found six cases of oral IPEH, of which three were diagnosed previously. Five of these cases were in males, and the mean age was 58 years. It was usually described as a blue or reddish nodule. The lower lip mucosa was the most common site followed by the tongue and the upper lip. Histologically, thrombi were always present and five out of the six lesions appeared in a mixed form, while only one was in a pure form. Three cases were diagnosed at an early stage and in the other three, the lesions were well-established. In this series, IPEH associated with vascular malformation was more common than reported in previous studies. Lesions at early stages, especially in the mixed form, may be unnoticed. PMID- 10522220 TI - Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis in periodontal disease: introduction. PMID- 10522221 TI - Taxonomy and biochemical characteristics of Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis. PMID- 10522222 TI - Microbiological tests for Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis. PMID- 10522223 TI - Oral ecology and person-to-person transmission of Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis. AB - The ecological characteristics of the oral cavity are dissimilar for A. actinomycetemcomitans and for P. gingivalis, as judged by differences in their colonization preferences and patterns, associations with periodontal disease parameters, relationships with the subgingival microbiota and the type of periodontitis and their clonal persistence in the oral cavity. These features also suggest that as a periodontal pathogen, A. actinomycetemcomitans is different from P. gingivalis. Probably in most infected individuals, low levels of A. actinomycetemcomitans can persist for years in equilibrium with the host and the resident oral microbiota. However, it is well established that A. actinomycetemcomitans can cause disease in some individuals or in some circumstances when the regulatory mechanisms are unable to maintain homeostasis in the ecosystem. Elevated A. actinomycetemcomitans proportions of the biota can be regarded as a sign of ecological imbalance, leading to increased risk of periodontal destruction. There is also evidence showing elevated pathogenic potential of certain A. actinomycetemcomitans clones. Although A. actinomycetemcomitans seems to be relatively rarely transmitted between cohabiting adults, transmission can occur to periodontally healthy children of A. actinomycetemcomitans-positive parents. Parents and children may share factors that promote successful oral colonization of A. actinomycetemcomitans, or the window of opportunity is in childhood. Therefore, to prevent parent-child transmission of A. actinomycetemcomitans, bacterium-positive parents of young children are optimal targets for enhanced information and treatment. In selected populations, screening for specific clones of A. actinomycetemcomitans has been employed in prevention of peridontitis. Future research aiming at finding the reasons which cause the changes in the oral homeostasis to allow the growth of A. actinomycetemcomitans may give insight into novel prevention strategies for A. actinomycetemcomitans-associated periodontitis. Compared with A. actinomycetemcomitans, P. gingivalis shows a different pattern of coexistence with the host. In periodontal health or in children, P. gingivalis is absent or only rarely detected. When present, P. gingivalis is commonly recovered in high numbers from dentitions exhibiting inflamed periodontitis and poor oral hygiene. Contrary to A. actinomycetemcomitans, the data on the vertical transmission of P. gingivalis are limited. The major infection route of P. gingivalis seems to be between adults, indicating that P. gingivalis commonly colonizes in an established oral microbiota. These characteristics suggest that the degree of tolerance between P. gingivalis and the host is inferior to that between A. actinomycetemcomitans and the host. It appears that the association of P. gingivalis with disease is a rule rather than an accidental incident. On these grounds, it seems that the host-P. gingivalis relationship approaches antibiosis. Since P. gingivalis infection is related to a typical periodontal eco-pathology, the susceptibility to person-to-person transmission of this pathogen may be controlled by periodontal treatment and emphasizing the significance of high standard oral hygiene. PMID- 10522224 TI - Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis in human periodontal disease: occurrence and treatment. PMID- 10522225 TI - Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis in nonoral infections. PMID- 10522226 TI - Virulence factors of Actinobacillus actinomycetemcomitans. AB - A. actinomycetemcomitans has clearly adapted well to its environs; its armamentarium of virulence factors (Table 2) ensures its survival in the oral cavity and enables it to promote disease. Factors that promote A. actinomycetemcomitans colonization and persistence in the oral cavity include adhesins, bacteriocins, invasins and antibiotic resistance. It can interact with and adhere to all components of the oral cavity (the tooth surface, other oral bacteria, epithelial cells or the extracellular matrix). The adherence is mediated by a number of distinct adhesins that are elements of the cell surface (outer membrane proteins, vesicles, fimbriae or amorphous material). A. actinomycetemcomitans enhances its chance of colonization by producing actinobacillin, an antibiotic that is active against both streptococci and Actinomyces, primary colonizers of the tooth surface. The fact that A. actinomycetemcomitans resistance to tetracyclines, a drug often used in the treatment of periodontal disease, is on the rise is an added weapon. Periodontal pathogens or their pathogenic products must be able to pass through the epithelial cell barrier in order to reach and cause destruction to underlying tissues (the gingiva, cementum, periodontal ligament and alveolar bone). A. actinomycetemcomitans is able to elicit its own uptake into epithelial cells and its spread to adjacent cells by usurping normal epithelial cell function. A. actinomycetemcomitans may utilize these remarkable mechanisms for host cell infection and migration to deeper tissues. A. actinomycetemcomitans also orchestrates its own survival by elaborating factors that interfere with the host's defense system (such as factors that kill phagocytes and impair lymphocyte activity, inhibit phagocytosis and phagocyte chemotaxis or interfere with antibody production). Once the organisms are firmly established in the gingiva, the host responds to the bacterial onslaught, especially to the bacterial lipopolysaccharide, by a marked and continual inflammatory response, which results in the destruction of the periodontal tissues. A. actinomycetemcomitans has at least three individual factors that cause bone resorption (lipopolysaccharide, proteolysis-sensitive factor and GroEL), as well as a number of activities (collagenase, fibroblast cytotoxin, etc.) that elicit detrimental effects on connective tissue and the extracellular matrix. It is of considerable interest to know that A. actinomycetemcomitans possesses so many virulence factors but unfortunate that only a few have been extensively studied. If we hope to understand and eradicate this pathogen, it is critical that in-depth investigations into the biochemistry, genetic expression, regulation and mechanisms of action of these factors be initiated. PMID- 10522227 TI - Virulence factors of Porphyromonas gingivalis. PMID- 10522228 TI - The role of the cell-mediated immune response to Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis in periodontitis. PMID- 10522229 TI - Humoral immune response to Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis in periodontal disease. PMID- 10522230 TI - Ecological considerations in the treatment of Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis periodontal infections. PMID- 10522231 TI - Evidence-based implementation of evidence-based medicine. AB - BACKGROUND: The slow and haphazard process of translating research findings into clinical practice compromises the potential benefits of clinical research. Most quality improvement (QI) initiatives are based on the beliefs of decision makers rather than on the growing theoretical and empirical knowledge about organizational and provider behavior change. If future QI activities are to improve the translation of evidence into practice, they should be based on an understanding of the different models and strategies for implementing research evidence and the evidence base supporting their use. Evidence-based medicine should be complemented by evidence-based implementation. THE EVIDENCE FOR DIFFERENT STRATEGIES OF IMPLEMENTING CHANGE: A general framework for changing practice based on theoretical perspectives and research evidence considers a variety of theoretical approaches and their contribution to an understanding of provider behavior change. The framework summarizes evidence from systematic reviews of provider behavior change, which suggest the potential of several dissemination and implementation strategies that are effective under certain conditions. Passive dissemination approaches are largely ineffective; specific strategies to implement research-based recommendations appear to be necessary to ensure practice change. Multifaceted interventions that address specific barriers to change are more likely to lead to changes in practice. PRACTICAL, FIVE-STAGE FRAMEWORK: A practical, five-stage framework for changing practice, which is illustrated with experiences from a comprehensive program on implementing evidence-based clinical guidelines in primary care, includes development of a concrete proposal for change; analysis of the target setting and group to identify obstacles to change; linking interventions to needs, facilitators, and obstacles to change; development of an implementation plan; and monitoring progress with implementation. PMID- 10522232 TI - Appraising clinical practice guidelines in England and Wales: the development of a methodologic framework and its application to policy. AB - BACKGROUND: Clinical guidelines are pervading all aspects of health care. Their potential benefits are vast--from decreasing unjustified variation in treatment, to improving outcomes for patient, to containing escalating costs. However, there is increasing concern that many of the guidelines produced may be of low quality and recommend ineffective treatment. A framework to appraise the methodologic quality of clinical guidelines, commissioned by the NHS in 1997, was developed and validated in the United Kingdom (UK) under the auspices of the National Research and Development Programme. THE INDEPENDENT APPRAISAL SERVICE: This framework is now used to assess all national guidelines funded by the National Health Service (NHS) in the UK as part of an independent appraisal service. The appraisal provides a basis for policymakers to decide which guidelines should be commended for use in the NHS. Each guideline is appraised by a minimum of six appraisers. Twenty-one guidelines had been appraised as of July 1999. The mean time for completion of the appraisals, from receipt of the guidelines to dispatch of the reports, was just over ten weeks. There has been a marked improvement in the quality of documentation for national guidelines--including the search strategy and synthesis of evidence--in the past three years, although some areas of development remain inadequately reported. POLICY IMPLICATIONS: Ensuring that the clinical guidelines are sound before recommending their use is essential to policymakers responsible for guideline programs, and a formal appraisal should be an integral part of those programs. PMID- 10522234 TI - Musings on using evidence to guide CQI efforts toward success: the computerized firm system as primary care microunit. AB - BACKGROUND: The attempt to transfer classic industrial CQI (continuous quality improvement) theory into the clinical arena has proved to be more difficult than originally promised. A new "computerized firm system" approach to incorporating CQI efforts into mainstream practice settings, which has been able to obviate many of these shortcomings, is described. METHODS: To make it easier for CQI efforts to be successful, the scope of activities undertaken in completing the Shewhart cycle popularly referred to as PDSA (plan change, do change, study results, act on results) was delimited. Rather than plan the intervention themselves, staff worked with experts on tailoring a preselected change idea with already established efficacy--a computerized reminder system. Because the clinic was divided into two small group practices known as firms, a controlled time series trial (CTST) design was used by initially turning the reminders on for one firm but not the other. The clinic was thereby also relieved of the responsibility of conducting a study to determine whether the intended improvement in quality had been achieved. In essence, one clinic was asked to do just DA (that is, do-act). RESULTS: This approach engendered the successful completion of a streamlined Shewhart cycle in a busy clinic setting at remarkably low cost. The compelling nature of controlled evaluation results aided leadership in rapidly disseminating the reminder system to the remaining 11 primary care clinics associated with the university's 2 academic medical centers. CONCLUSION: Computerized firm systems can be developed to conduct CTSTs as part of streamlined CQI cycles guided by both published and local evidence, and they are worth developing. PMID- 10522233 TI - The role of evidence reports in evidence-based medicine: a mechanism for linking scientific evidence and practice improvement. AB - STUDY QUESTIONS: In this article two related questions are considered: (1) Why isn't evidence-based medicine (EBM) more consistently implemented? and (2) Using the EBM paradigm, by what mechanism can we link evidence reports to concrete practice improvement activities? STUDY DESIGN: To motivate a systematic analysis, answers to these questions are framed within the context of a general conceptual model for practice improvement, using as an example the application of this general model to the question of improving anticoagulation. CONCLUSIONS: The potential role of evidence reports is quite broad and to be most effective, they should (1) be considered as part of a comprehensive strategy for practice improvement and (2) be designed with their customers in mind. A system-based method for using the information from evidence reports involves ultimately suggesting specific practice improvement strategies in which the strategies are defined in terms of (1) a set of functional specifications and (2) a toolbox of implementation options. Such an approach brings to bear the specialized expertise and generalized fund of scientific knowledge used to produce the evidence report, but does so in a way that facilitates local tailoring. That is, while information synthesis should be global, implementation must be local. PMID- 10522235 TI - Discipline-based approaches to evidence-based practice: a view from nursing. AB - BACKGROUND: Research carried out by nurses or by others on patient problems of concern to nurses is contributing to the development of evidence-based nursing practice. In the past few decades, there has been a dramatic increase in clinical research, in health services research, and in the content and process of informatics, all focused on nursing care. The translation of findings of this research into clinical practice and the organization of nursing is less dramatic. The opportunity to implement research-based practice is great, but requires attention, methods, and resources. Also required are a database and an information system which include terms essential to nursing practice. DIMENSIONS OF NURSES' INVOLVEMENT IN EVIDENCE-BASED PRACTICE: The importance of nurses' involvement in evidence-based practice (EBP) can be viewed from three perspectives: (1) nurses' participation in medical problems and medical interventions, (2) nursing problems and nursing interventions, and (3) development and use of a standardized language that describes the problems, interventions, and outcomes important to nursing. APPLYING EBP TO COMBINED MEDICAL AND NURSING PROBLEMS: The best outcomes for a specific patient population are achieved through a combination of the medical and nursing problems and evidence-based interventions. Examples of problems of importance to nursing practice and research include pain, dehydration, incontinence, lifestyle change, confusion, immobility, knowledge deficit, noncompliance, anxiety, skin breakdown, inappropriate use of restraints, and falls. Interventions for prevention and treatment of the individual problem or combination of problems comprise the focus of nursing research and EBP. PMID- 10522236 TI - From book to bedside: putting evidence to use in the care of the elderly. AB - BACKGROUND: Infusion of research findings into clinical practice is a challenging part of the research process. Because the length of time between discovery and use of knowledge averages 20 years, methods are needed to speed translation of research findings into practice. Few efforts have been made to coordinate the generation of new knowledge with the dissemination of findings from research to improve care of the elderly. RESEARCH-BASED PRACTICE PROTOCOLS: The Research Development and Dissemination Core (RDDC) of the Gerontological Nursing Interventions Research Center (GNIRC) at the University of Iowa emphasizes development of research-based (RB) protocols, which requires collecting relevant literature, critiquing studies, and synthesizing research findings for practice. GNIRC-generated research is disseminated to nurses in practice, and the RDDC links nurses who identify clinical problems in care of the elderly with GNIRC scientists. Currently, 19 RB protocols are offered for dissemination through the RDDC, and 5 protocols are under development. Implementation and evaluation of research-based practices on "Split Thickness Skin Graft Donor Site Care" and "Nasogastric/Nasointestinal Tube Placement" are described. CONCLUSIONS: Lessons learned on the basis of experience in disseminating and implementing research based practices include the necessity of tailoring them to the local needs of various clinical settings in which they are used, reinfusing them periodically to keep staff motivated, and making them consumer friendly. The challenge remains to integrate these practices into the fiber of organizations and to keep staff educated and motivated to carry out research-based practices to improve the care of the elderly. PMID- 10522237 TI - Amino acid similarity matrix for homology modeling derived from structural alignment and optimized by the Monte Carlo method. AB - In this paper, we obtained a similarity matrix for homology modeling based on the structure of proteins in a structural alignment. The alignment procedure was executed within dynamic programming generally used in alignment methods. An initial matrix derived from the structural alignment was optimized by the Markov chain Monte Carlo method at low temperature to fit its sequence alignment to the structural alignment. Structural alignment was performed on the basis of the superposition of C alpha atoms for two protein structures. The objective function in the Monte Carlo procedure was defined by entropy in the information theory, allowing us to show that the amino acid similarity matrix aligned accurately. When compared with the structural alignment, the average number of incorrect amino acid residues in the sequence alignment was 22.6 for all residues and about 3.7 for residues in structurally conserved regions. The alignment with our matrix was more similar to structural alignment than to sequence alignments using other amino acid substitution matrices. PMID- 10522238 TI - A comparative study of isodensity surfaces using ab initio and ASA density functions. AB - In this article, we report a visual comparison between several of the available methods for constructing electronic density functions. The density forms studied include ab initio, atomic shell approximation, and promolecular densities. A graphical comparison is made for six different molecules at different levels of density function values. The differences between the various density functions are analysed by considering a molecular quantum self-similarity measure and the required computational time for all molecules at all computation levels is considered. PMID- 10522240 TI - Atomic-level molecular modeling of the nonionic surfactant Triton X-100: the OPE9 component in vacuum and water. AB - The commercially available nonionic surfactant Triton X-100 is a mixture of polyoxyethylene tert-octylphenyl ethers (OPEn) with an average of n = 9.5 oxyethylene (OE) units in the molecules, and the population maximum at n = 9. Thus, the OPEn = 9 component was chosen to be studied by atomic level molecular modeling, using second-generation force fields. The 1,000 conformers generated via random sampling of torsional angles around single bonds yielded 11 clusters based on geometrical similarity. Representatives of geometrically distinctly different clusters with significant populations were chosen from a narrow energy range around the most probable energy to be analyzed for interaction with water. The effect of water on the conformation of the OE chain was found to be modest, similar to the situation that had been reported earlier for the anionic surfactant Aerosol-OT (AOT). The number of bound water molecules is strongly dependent on the conformation of the OE chain and is affected by electrostatic as well as steric effects. Unlike the case of AOT, for which the length of the hydrophobic tail was found to govern the size of reverse micelles in CCl4, the size of reverse micelles of OPEn = 9 cannot be predicted from the dimensions of the hydrophilic tail. PMID- 10522239 TI - MANIP: an interactive tool for modelling RNA. AB - Large RNA structures can be viewed as assemblies of smaller units or modules that are usually clearly identified (helices, hairpin loops, other recurrent motifs, etc.). We have developed a program, MANIP, which allows the rapid assembly of separate motifs (each with a specified sequence) into a complex three-dimensional architecture. The already determined modules are present in a database from which they can be extracted with the appropriate sequence. Their assembly is performed in real time on the computer screen with buttons and dials that command rotation and translation of any chosen fragment with respect to the chosen pivot, or that generate all possible variations of any torsion angle within a specified segment either in the 5' or in the 3' direction. The possible in-built manipulations follow the general stereochemical rules of RNA structure. MANIP automatically recognizes and displays the allowed and nonallowed hydrogen bonds between the residues. The program is interfaced with a rapid and automatic online refinement tool of partial or full assemblies, NUCLIN-NUCLSQ. The refinement protocol incorporates canonical as well as noncanonical base pairing constraints together with restraints imposed by covalent geometry, stereochemistry, and van der Waals contacts. The computer package runs on UNIX Silicon Graphics workstations and is written in C with OpenGL and X11/Motif libraries. PMID- 10522241 TI - Q: a molecular dynamics program for free energy calculations and empirical valence bond simulations in biomolecular systems. AB - A new molecular dynamics program for free energy calculations in biomolecular systems is presented. It is principally designed for free energy perturbation simulations, empirical valence bond calculations, and binding affinity estimation by linear interaction energy methods. Evaluation of ligand-binding selectivity and free energy profiles for nucleophile activation in two protein tyrosine phosphatases as well as absolute binding affinity estimation for a lysine-binding protein are given as examples. PMID- 10522242 TI - Temperature dependence of distributions of conformations of a small peptide. AB - Multicanonical Monte Carlo simulations of the pentapeptide Met-enkephalin were used to study its low-energy conformations in detail. The resulting conformations are classified into six categories of similar structures based on the pattern of intrachain hydrogen bonds. Several thermodynamic quantities such as the distributions of hydrogen bonds and those of backbone dihedral angles were obtained as a function of temperature. From these results, it was concluded that at least four of the six categories are well-defined local minimum energy states. These four categories are in agreement with our prior results based on root-mean square interatomic distances. PMID- 10522243 TI - DISSIM: a program for the analysis of chemical diversity. AB - As interest in database searching and compound selection has grown, there has been a concomitant growth in interest in the quantification of chemical similarity. Described here is a computer program called DISSIM, which addresses the problem of selecting diverse subsets from larger collections of chemical compounds. It is a pragmatic solution combining a maximum dissimilarity search algorithm and a general multidimensional measure of chemical similarity based on the combination of different molecular descriptors. The problem of correlation between descriptors is addressed and appropriate schemes for weighting and normalisation are described. The specific application of these techniques to the comparative analysis of topological indices and their use in the area of chemical diversity analysis and compound selection are also described. PMID- 10522244 TI - [Psychobiological aspects of posttraumatic stress disorder]. AB - The exposure to a traumatic event may result in posttraumatic stress disorder (PTSD), which is characterized by a complex symptomatology, clustered into three groups of symptoms, i.e. intrusive memories, avoidance behavior, and hyperarousal. Since PTSD is a stress reaction, alterations of stress-responses of neurobiological systems have been examined in patients suffering from PTSD. The investigation of biological parameters refers to the hypothalamic-pituitary adrenal axis (HPA), studies of the noradrenergic and the endogenous opiate system as well as psychophysiological and neuroimaging studies. Besides others, the observed biological dysregulations refer to hypocortisolism with an enhanced negative feedback of the HPA axis, enhanced noradrenergic activity, and neuroanatomical changes. To elucidate the specificity of this findings for PTSD, the dysregulations will be discussed with reference to findings in major depression and somatoform disorders. PMID- 10522245 TI - [Serotonergic dysfunction caused by social isolation. Implications for the development of aggressiveness and alcoholism]. AB - Dysfunction of serotonergic neurotransmission has been associated with two different psychopathological syndromes--impulsive aggressivity resulting form a lack of stimulation of the "behavior inhibition system" on the one hand and the manifestation of clinical depression and compulsive syndromes on the other. The examination of primate behavior provides a model which may reconciliate these seemingly contradictory hypotheses. According to primate experiments, monoaminergic depletion results in anxious and desperate behavior only if the individual has previously been exposed to social isolation stress, which in turn induces a decrease in the central serotonin turnover rate. Young non-human primates who experience early social separation stress are anxious and fearful, while as adults they tend to be aggressive, consume excessive amounts of alcohol and are less intoxicated by alcohol intake. These observation indicate the importance of social separation stress in the pathogenesis of alcoholism and antisocial behavior and may point to prophylactic and pharmacological treatment strategies. PMID- 10522246 TI - [Therapeutic experiences with alcohol-related Korsakoff syndrome]. AB - Korsakoff's psychosis (Wernicke-Korsakoff-Syndrome) following severe, long-term alcoholism is considered to be widely resistant to treatment. We report two cases with beneficial effects of treatment by a combination of clonidine and fluvoxamine. PMID- 10522247 TI - [Buprenorphine vs. methadone as maintenance treatment for opioid dependence]. AB - The efficacy of buprenorphine in opioid dependent patients (n = 20) was compared to methadone maintained subjects (n = 20) in a randomized comparison trial. Sublingual application of buprenorphine as an alternative synthetical opioid is being compared to methadone during a 24 week study period. A trend (p = 0.06) could be found in the retention rate of investigated patients being maintained on a mean dosage of 63 mg oral applicable methadone (racemat of L- and D-methadone) in comparison to the group on a mean dosage of 7.3 mg buprenorphine (sublingual tablets). The dropout-rate of 11 subjects at the end of the study in the buprenorphine group was higher when compared to the dropout-rate of 5 in the methadone group. There was no significant difference between the two groups over the treatment period in respect to additional consumption of opiates, benzodiazepines and cocaine as evaluated through urine toxicology. The result in regard to compliance over the study period demonstrates that methadone appears to be the more successful oral opioid (p = 0.04). Nevertheless, efficacy of buprenorphine in maintenance could be demonstrated in the remaining subjects, and further studies with higher daily doses and a higher number of subjects have to be performed. PMID- 10522248 TI - ["Pathological intoxication."Diagnostic artefact or a reliable psychiatric diagnosis?]. AB - "Pathological intoxication" has been a matter of controversial discussions during the last years. On one hand diagnosis of "pathological intoxication" in forensic expertise often is associated with the assumption of legal insanity. On the other hand it has been suggested that the term "pathological intoxication" should be abandoned in favor of more accurate clinical diagnosis, and recently "alcoholic idiosyncratic intoxication" was canceled as a separate diagnosis in DSM-IV. In order to register the diagnostic habits connected with "pathological intoxication" we had sent a questionnaire to all psychiatric institutes (n = 541) in Germany. We were then able to evaluate 338 questionnaires (62.5%) relating to number of diagnoses and diagnostic criteria. In synopsis diagnostic habits turned out to be very inhomogeneous. Nearly two third of psychiatric institutes reported not to have used the diagnostic category between 1991 and 1993, whereas nearly 50% of diagnoses (210 out of 456) were reported by only 13 (3.9%) institutes. Diagnosis of "pathological intoxication" is based predominantly on vague and non distinct criteria, such as "violent excitation" or "strange unusual behavior". In accordance to DSM-IV we therefore suggest that the diagnostic term "pathological intoxication" should be abandoned, especially in the context of forensic psychiatric expertise. Instead, assessment of intoxication must be based on individual somatic and psychopathological symptoms. PMID- 10522249 TI - [Does psychiatric comorbidity increase length of stay in medical, surgical and gynecological departments?]. AB - Several studies from Anglo-American countries indicate that in non-psychiatric hospital departments mentally ill patients have a longer length of hospital stay than mentally well, while in Austria and Germany, until now, no studies concerning this question exist. Therefore, we investigated the influence of psychiatric comorbidity on the length of stay in 608 patients of medical, surgical and gynecological departments in Vienna and Tyrol. Based on the Clinical Interview Schedule, 28.1% of the patients in this sample suffered from psychiatric disorders. The presence of psychiatric disorders, as well as type of hospital department (medical department), higher age, more previous non psychiatric treatment periods, and more somatic diagnoses predicted a longer duration of inpatient treatment. To avoid the influence of cofounding variables, psychiatric cases were matched with psychiatric non-cases. The mentally ill group was treated for a markedly longer period as inpatients than the mentally well. Patients with a diagnosis of dementia or of substance abuse showed a significantly increased length of stay, while we could not confirm this for other psychiatric diagnoses. PMID- 10522250 TI - [Is there a disproportional increase in health care costs for schizophrenic patients?]. AB - Empirical data on costs of care for chronically mentally ill patients in Germany are rare. There is a lack of cross-sectional studies as well as of any analysis of the long-term course of mental health care costs. This study combines data from two cost-studies on patients with schizophrenia, conducted in the same catchment area at an interval of 15 years, to draw conclusions about the long term course of these costs. The direct costs of comprehensive community based mental health care had increased by 77.0% during 15 years, while the costs of a permanent stay in a long-term ward of a psychiatric hospital had increased by 78.5% in the same period. This increase was 27 and 28.5 per cent higher than the general rise in cost of living in Germany during the interval between the two studies. This higher-than-average increase in mental health care costs was accompanied by a much higher cost-effectiveness. Thus, this rise of costs must be seen as a consequence of the great deficits and the resulting basic improvement of care for chronically mentally ill patients in Germany, which was demanded by an expert-commission of the government in the mid-seventies. PMID- 10522252 TI - [Psycho-neurogenic factors as a cause of life-threatening arrhythmias]. AB - Tachyarrhytmias often occur during increased emotional arousal or mental excitation. The implantable cardioverter defibrillator (ICD) allows the exact documentation of arrhythmic episodes and their time of onset. Therefore, the type of arrhythmia can be differentiated as well as the circumstances surrounding the event. These features allow the assessment of possible psychic arrhythmogenic factors in the natural environment. We analyzed the ICD-protocols of three male patients (in ages between 60 and 68), whose devices had successfully terminated a ventricular tachycardia and compared the onset of the episodes with the patients' detailed descriptions of the corresponding life situations. The analysis of the circumstances at the time of arrhythmia-onset revealed a relationship between the occurrence of life-threatening arrhythmias in natural environment and emotional stress. The stressors could be defined as situations of increased vulnerability leading to sympathetic excitation. The induction of tachyarrhythmia was promoted in case 1 by acute psychic distress (public speaking), by the increasing panic attack-like vicious circle of the cognitive anticipation of an unfavorable outcome (case 2), and an adverse anger reaction superimposed on persistent feelings of help- and hopelessness (case 3). These findings are in line with several experimental and epidemiological studies providing evidence for a relationship between psychic arousal and the induction of tachyarrhythmias. The knowledge of emotional and mental factors that function as a trigger for arrhythmias may lead to new therapeutic approaches in the prevention of sudden cardiac death. PMID- 10522251 TI - [Osteoporosis in anorexia nervosa. New aspects of pathogenesis and therapy]. AB - Osteoporosis has been shown to be a relatively common complication of anorexia nervosa (AN). So far the exact mechanisms which are implicated are not fully clarified. Several factors such as malnutrition, reduced body weight, amenorrhea, and hypercortisolaemia seem to be involved. There is a strong relationship with the duration of amenorrhea and--in some studies--with the age of onset. Osteoporosis is for a long time a "silent" disease and the first symptoms such as back pain, loss of height, kyphosis, and fractures are late complications. Therefore, routine screening methods for bone density measurements should be established. The most accurate is the dual energy X-ray absorptiometry (DEXA). Therapeutically the primary aim should be to reach restoration of both normal body weight and regular menses. As AN is a chronic disease clinicians should be aware of the dangers of osteoporosis and start with the treatment and/or prevention of osteoporosis early. However, at this stage it is difficult to provide an evidence-based management plan for osteoporosis in AN. Hormone replacement therapy (HRT) as well as calcium and vitamin D-supplementation are under discussion, however, further controlled investigations are warranted. PMID- 10522253 TI - [Hyperglycemia and ketoacidosis associated with olanzapine]. AB - Two psychotic patients developed hyperglycaemia several weeks after starting olanzapine. In one case the elevated glucose concentrations returned to normal soon after withdrawal of olanzapine. In the second case severe ketoacidosis with lethal outcome occurred. PMID- 10522254 TI - [Leukopenia associated with butyrophenones. A successful treatment of psychoses with pulsed administration of neuroleptics]. AB - In the monotherapeutic treatment of an acute schizophrenic psychoses with butyrophenones (haloperidol and benperidol) we saw immediate leucopenic reactions when the medication was given every day. In literature the risk of butyrophenone induced blood dyscrasias seems to be underestimated. Therefore we decided to report this single case evaluation. When benperidol (short elimination--half life period) was given every other day ("interval-therapy") the blood cell count of the leucocytes normalized. Furthermore, the amount of psychopathological abnormalities measured by the Brief Psychiatric Rating Scale decreased under this intermittent treatment. PMID- 10522255 TI - [Child sexual abuse, false-memory syndrome and Huntington chorea]. AB - We report on a female inpatient who at the age of 47 years presented depressive and anxiety symptoms and alcohol abuse and who suffered from cognitive, personality, and discrete movement disturbances later on. In the course of the long-term psychotherapy which was supported by the technique of creative drawings previously forgotten memories of a very severe sexual abuse in childhood emerged. The recovery of these memories was followed by an intensification of the anxiety which lead to several psychiatric rehospitalizations. During the last hospital stay the diagnosis of Huntington's chorea was verified explaining well the rich psychopathology of the patient. The recollections of sexual abuse were for the most part qualified as a false memory syndrome. In addition to other factors the chorea-inherent cognitive impairment will have contributed to the occurrence of false memory syndrome. The new diagnosis dictated a change of the therapeutic procedure which, at least in the medium-term, proved to be successful. PMID- 10522256 TI - [Investigation of personal niche as basis of supportive psychotherapy]. AB - According to the ecological theory of Willi 1996, people create and maintain their psychic wellbeing through constant shaping of their environment, i.e. setting up a personal niche. The personal niche in turn responds to, guides and challenges further personal development. This study explores what kind of solutions single patients with psychiatric invalidity can find in order to form their environment as a niche in which they may develop effectiveness and use it for their psychic regulation. 30 people were questioned with a semi-structured interview: they were patients of a psychiatric out-patient's clinic, living alone and psychiatrically invalidated (2/3 with schizophrenia of the residual type). The results were ranked after their increasing demands in a relational and in an activity inventory. The more relationship is based on mutuality expectations of others, personal disclosure and binding the more the subjects feel overstrained. The investigation of these inventories serve as a technical aid in supportive psychotherapy. PMID- 10522257 TI - [Molecular and cellular determinants of arterial stiffness: role of cell-matrix connections]. AB - Increased large artery stiffness is believed to be a cardiovascular risk factor independent from mean arterial pressure. The mechanical properties of large arteries depend not only on the amounts of their main constituents (elastin, collagen, and smooth muscle cells) but also on the spatial organization and mechanical interactions among these components. These interactions may be mediated by extracellular matrix adhesion proteins and their membrane receptors or integrins. From a mechanical viewpoint, a key element may be the dense plaque, which is composed of cytoskeletal proteins linked to matrix proteins via membrane integrin receptors. Integrin expression in normal and diseased blood vessels is currently the focus of active research. In humans, hypertension-related arterial hypertrophy is not associated with an increase in intrinsic arterial wall stiffness. Aortic fibronectin expression is increased in spontaneously hypertensive rats (SHRs). By increasing cell-matrix anchoring, fibronectin may contribute to protect arterial wall components from the increased mechanical loads associated with hypertension. In atherosclerosis, the increase in cell matrix anchoring plays a key role in preventing atheroma plaque rupture. To determine the exact role of adhesion molecules in arterial stiffness, there is a need for studies involving use of specific anti-integrin agents and of transgenic animal models. PMID- 10522258 TI - Arterial calcification. Mechanisms, consequences and animal models. AB - The arterial extracellular matrix undergoes many profound age-related changes leading to an increase in wall stiffness. In this review the evidence suggesting that calcium--and more importantly "calcification" of elastin fibres--is involved in the age-related increase in arterial stiffness is examined. PMID- 10522259 TI - Noninvasive evaluation of arterial abnormalities in hypertensive patients. AB - Morbidity and mortality in hypertension are mainly determined by arterial lesions which may occur in different regional circulations: kidney, cerebral, coronary ..., causing respectively nephroangiosclerosis, stroke or myocardial infarction... Despite the arteries heterogeneity, structural and functional abnormalities are usually observed at an early stage of hypertension in both large and small arteries. These alterations modify arterial wall physiological and mechanical properties which can be expressed clinically by increasing arterial pulsatility or pulse pressure; they facilitate establishment and progression of atherosclerosis and arteriosclerosis. Since arteries constitute the target, site and common denominator of hypertension cardiovascular complications, several noninvasive techniques may be usefull to assess their haemodynamic: casual and ambulatory blood pressure measurements can evaluate pulse pressure which can be also directly measured in different sites of the arterial tree using the "Tonometer" device; ultrasound techniques can be applied: Doppler signal to assess the arterial flow, video-echo signal to analyse the arterial structure such as intima-media thickness, or echo-tracking systems for direct measurements of arterial wall distension and thickness; pulse wave velocity is widely used as index of arterial distensibility; its assessment, using the Complior device showed that hypertensive patients present a decrease of arterial distensibility and that antihypertensive treatment do not always reverse this abnormality. Since cardiovascular morbidity and mortality are due to arterial lesions, it is important to evaluate the effect of cardiovascular prevention on the arterial wall. Large therapeutical trials, including arterial evaluation, are necessary to assess whether this consideration may particularize patients with high cardiovascular risk and contribute to their treatment and prognostic improvement. PMID- 10522260 TI - Mechanical properties of the aorta determined by the pressure-diameter relation. AB - Elastic properties of the aorta represent an important determinant of left ventricular function and coronary blood flow. Aortic pressure-diameter relation provides insights into the mechanism, involved in the alterations of the elastic properties of the aorta. Recently, we described a new method to obtain aortic pressure-diameter relation in conscious humans. Aortic strain, distensibility, pressure-diameter relation, stiffness constant, augmentation index and aortic energy loss are indexes of aortic elastic properties derived by this method. Coronary artery disease is a potent independent factor associated with aortic elastic performance. Smoking produced an acute decrease of the elastic properties of the aorta in habitual smokers. Passive smoking has a similar unfavourable effect on aortic elastic properties. Elastic properties of the aorta are deteriorated in hypertensive patients compared to normotensive subjects and energy loss due to aortic wall viscosity is increased in the first group. Tachycardia induced by ventricular pacing produces an acute increase of the aortic distensibility in humans. Intravenous 17 beta-estradiol produces an improvement of the elastic properties of the aorta in menopausal women both with and without coronary artery disease. A decrease of aortic stiffness is demonstrated during intraaortic balloon pumping, associated with significant increase of the cardiac index and significant reduction of myocardial oxygen consumption. Thus, therapeutic interventions alter the elastic properties of the aorta and improvement of the elastic properties of the aorta may be beneficial in altering the natural history of diseases. PMID- 10522261 TI - High resolution angiometers for the assessment of the elastic modulus. AB - Distensibility and compliance are vessel wall properties of large arteries that determine their dynamic behaviour and play a role in cardiovascular disease. At present, arterial distensibility and compliance can be accurately investigated using noninvasive ultrasound techniques. Echo-tracking devices in particular are suited to study local distensibility and compliance of several superficial large arteries. The best-known echo-tracking devices (Nius02 and Wall Track System) are briefly outlined with respect to their advantages and disadvantages. These techniques have shown a good reproducibility and accuracy. The two devices are complementary and can be applied to different parts of the arterial tree. The results of the studies done with echo-tracking devices give insight in the pathophysiologic mechanisms underlying cardiovascular diseases in different arterial territories. In to the future it may be possible to use local arterial wall properties together with cardiac wall hypertrophy as important markers of future cardiovascular risk and as relevant parameters for the evaluation of pharmacological interventions. PMID- 10522262 TI - Noninvasive measurements of aortic stiffness: methodological considerations. AB - This article overviews methods currently available for the non-invasive determination of aortic stiffness and critically appraises their strengths and weaknesses. Approaches are divided into indirect and direct (Windkessel models are not reviewed). Indirect techniques rely on the measurement of pulse wave velocity (PWV) to obtain information about the average stiffness of the vessel pathway being studied. Typically "foot-to-foot" transit time measurements are combined with transcutaneous length measurements to calculate PWV (as length/transit time). Applanation tonometry, acoustic transducers, Doppler ultrasound and magnetic resonance imaging (MRI) have all been used to measure transit times. Direct techniques rely on assessing the relative change in aortic diameter (or area) between systole and diastole (strain) and combine these data with pulse pressure (stress) measurements to calculate vessel stiffness (as stress/strain). Transthoracic echocardiography and MRI have been used to assess aortic strain. All the approaches--both direct and indirect--however have their limitations. For indirect techniques the greatest errors are likely to be introduced by the transcutaneous estimation of the aortic path length. For direct techniques stress and strain are generally measured in different portions of the vasculature. These and other methodological issues are considered and areas where further work is needed are highlighted. PMID- 10522263 TI - Intravascular ultrasound studies of arterial elastic mechanics. AB - A number of new methods are available for the measurement of large artery elastic properties in human subjects in vivo. One powerful tool which has recently been applied to the study of large artery mechanics is intravascular ultrasound (IVUS). IVUS studies are performed using a high frequency ultrasound transducer mounted on the tip of a catheter. This catheter is inserted into a blood vessel and detailed cross-sectional images of the vessel, are obtained from within the lumen. IVUS techniques have been used to study wall mechanics of the human aorta, as well as peripheral and coronary arteries in normal human subjects and in patients with vascular disease. This paper reviews IVUS studies of human arterial elasticity and discusses the strengths and weaknesses of this emerging technique as it is applied to the understanding of arterial mechanical properties. PMID- 10522264 TI - Large arteries in hypertension. Pharmacological and therapeutic aspects. AB - The blood pressure curve may be divided into two components: mean arterial pressure, the steady component, and pulse pressure, the pulsatile component. Whereas pharmacological interventions in hypertension have been focused mainly on mean arterial pressure, little has been done to reduce selectively pulse pressure. However, increased pulse pressure with nearly normal mean arterial pressure is the classical hallmark of systolic hypertension in the elderly and a strong independent predictor of cardiac mortality, mainly for myocardial infarction. In this report, after the analysis of the factors influencing pulse pressure, a review of the pharmacological agents susceptible to reduce this hemodynamic factor is performed. PMID- 10522265 TI - Pharmacological improvement of large arteries properties. AB - Arterial distensibility reflects mechanical properties of the arterial wall and have, thus a clearcut clinical relevance. This because an arterial distensibility reduction is associated with an increased pulse pressure, an increased cardiac work and a reduced diastolic vital organ perfusion. In recent years it has been demonstrated that arterial distensibility is reduced in marked and mild hypercholesterolemia, in a manner independent from arterial blood pressure values. Arterial mechanical properties are also impaired in heart failure, this leading to a further cardiac damage. This important vascular properties however, can be improved by appropriate treatment, while the time needed for that can be extremely long. PMID- 10522266 TI - Clinical studies and therapeutic trials in systolic hypertension. AB - A disproportionate increase in SBP over DBP has been recognized for many years as a frequent accompaniment of aging. Initially this was considered to be benign, risk free and potentially dangerous to treat. Study over the years has shown that it is not benign and that antihypertensive therapy can reduce the risks of stroke, myocardial infarction, congestive heart failure and cardiovascular death. Currently, the drugs most widely recommended for this purpose are the thiazide diuretics, long acting dihydropiridine calcium channel antagonists, ACE inhibitors, and beta blocking agents. There may be a special place for nitrates, since these agents are very effective in increasing arterial distensibility--a primary abnormality of the disorder--but a formal study of their effectiveness has not been done. Concern about diastolic hypotension during therapy suggests that treatment to lower the blood pressure in this disorder should be carried out gradually with the aim of reducing the SBP toward normal while avoiding diastolic hypotension. PMID- 10522267 TI - Quantitative analysis of pharmacokinetic study samples by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). AB - The basis of all pharmacokinetic evaluations are powerful assays to quantify drugs and/or metabolites in biological matrices using modern sensitive instrumental analytical techniques, such as capillary gas chromatography and high performance liquid chromatography (HPLC). Being both specific and universal, mass spectrometry (MS) is an ideal chromatographic detector. Due to recent exciting achievements in the interfacing of liquid chromatography (LC) and MS, LC-MS, like the successfully preceding hyphenated technique gas chromatography-mass spectrometry (GC-MS), has now become a valuable technique in the analyst's toolbox. The key features of LC-MS are explained and four examples demonstrating its potential for highly specific and sensitive routine drug assays with the option of high sample throughput in pharmacokinetic investigations are presented. PMID- 10522268 TI - Naphthannelated azepinones: synthesis and antitumor activity. AB - 5H-Benzo[b]naphth[2,3-e]azepine-6,13-diones 4a, 4b and 4H-naphtho[2,3 e]thieno[3,2-b]azepine-5,12-dione (6) were prepared by aldol condensation of phthalic dialdehyde (3) with the fused azepinediones 2a, 2b and 5, respectively. The Schmidt reaction of naphthacene-5,12-quinone (7) yielded 6H benzo[e]naphth[2,3-b]azepine-7,12-dione (10). Several derivatives of the heterocyclic basic scaffolds 4, 6 and 10 were prepared by standard procedures, e.g. Grignard reaction, deoxygenation with triethylsilane, and sodium borohydride reduction. Evaluation of the synthesized compounds in the NCI in vitro cell line screening revealed a modest antitumor activity without marked cell line selectivity for the majority of the derivatives. The 2-bromo-5H benzo[b]naphth[2,3-e]azepin-6(13H)-one (19) was the only representative in this series exhibiting a noteworthy growth inhibitory effect for human tumor cells. PMID- 10522269 TI - Synthesis and some reactions of 2-acetylimidazo[4,5-b]pyridine. Antituberculotic activity of the obtained compounds. AB - 2-Acetylimidazo[4,5-b]pyridine was prepared and its reactions with some aromatic amines and sulfur (Willgerodt-Kindler reaction), some aromatic aldehydes, some carboxylic acid hydrazides as well as thiourea were investigated. New imidazo[4,5 b]pyridine derivatives with different substituents in 2-position (N arylthioamides, imines, alpha, beta-unsaturated ketones, hydrazido-hydrazones and aminothiazole) were obtained. Most of the synthesized compounds were tested in vitro for their antituberculotic activity. PMID- 10522270 TI - Synthesis of some new heterobicyclic nitrogen systems bearing the 1,2,4-triazine moiety as anti-HIV and anti-cancer drugs, Part II. AB - Some new heterobicyclic nitrogen systems 5-18 and/or thioethers 20, 22 bearing the 1,2,4-triazine moiety have been synthesized via condensation of 3-formylamino 1,2,4-triazine 3 with nitrogen compounds followed by heterocyclization with oxygen reagents. Thioether analogs 20, 22 have been obtained from fusion of compound 19 with 4-chlorothiophenol. The structure of the products have been established by elemental analysis and spectral data. The anti-HIV and anticancer activities of some products have also been investigated where compounds 20a, c and 22b exhibited a moderate activity. The biocidal-structures activity correlation was also studied. PMID- 10522271 TI - Characterization of urinary metabolites of a new benzofuroquinoline derivative, 3,9-bis(N,N-dimethylcarbamoyloxy)-5 H-benzofuro[3,2-c]-quinoline-6-one (KCA-098), in dogs. AB - The metabolism of 3,9-bis(N,N-dimethylcarbamoyloxy)-5 H-benzofuro[3,2-c] quinoline-6-one (KCA-098), a new inhibitor of bone resorption and stimulator of bone formation, was examined after oral administration to dogs. Nine metabolites and the unchanged KCA-098 were isolated by extraction and HPLC from dog urine. The structures of these metabolites were characterized by LC/MS or LC/MS/MS, and/or were confirmed by comparison with corresponding authentic standards. The presumed main metabolic pathways were hydrolysis, hydroxylation, and N demethylation of the N,N-dimethyl-carbamate ester group. PMID- 10522272 TI - Free versus liposome-encapsulated lignocaine hydrochloride topical applications. AB - The influence of encapsulating lignocaine hydrochloride, in a liposomal delivery system on its release from a topical formulation was studied. Liposomes were prepared from a mixture of L-alpha-dipalmitoylphosphatidyl choline (DPPC) and cholesterol in different molar ratios with or without charge inducing agents. The swelling time affording maximum entrapment was tested from 0-48 h at 53 degrees C. The percentage of drug entrapped in liposomes was found to be charge dependent and more pronounced for negatively charged liposomes. The amount of drug entrapped increased by increasing the ratio of cholesterol. The selected formulations were evaluated in vivo using the pin prick method. The results revealed localized and prolonged activity of lignocaine contained in liposomes when compared with an equivalent conventional topical application. PMID- 10522273 TI - The cytotoxicity and mode of action of 2,3,4-trisubstituted pyrroles and related derivatives in human Tmolt4 leukemia cells. AB - 4-Carbethoxy-1-methyl-2-phenacyl-3-phenylpyrrole (9), 4-carbethoxy-2-(4 methoxybenzoyl)-3-(4-methoxyphenyl)pyrrole (10) and 2-(4-methoxybenzoyl)-3,4-bis (4-methoxyphenyl)pyrrole (11) proved to be potent cytotoxic agents against the growth of murine and human leukemias and lymphomas. Selective toxicity was demonstrated against the growth of solid tumors, e.g., human adenocarcinoma of the colon SW480 and ileum HCT-8, glioma U-87-MG, and rat UMR-106 osteosarcoma. A mode of action study in Tmolt4 leukemia cells demonstrated that the agents inhibited de novo purine synthesis at the regulatory sites PRPP-amido transferase, IMP dehydrogenase as well as dihydrofolate reductase resulting in significant inhibition of DNA synthesis in 60 min. Other biochemical sites which were affected significantly were thymidylate synthetase, DNA polymerase alpha, RNA polymerases, nucleoside kinase and ribonucleoside reductase. PMID- 10522274 TI - Development of a predictive statistical model for the analgesic activity of a family of imides. AB - Quantitative structure-activity relationship (QSAR) studies were done with a family of cyclic imides. Promising efforts to create a QSAR model with substantial predictive power for the design of novel cyclic imides with improved analgesic activity are reported. PMID- 10522275 TI - Synthesis and anticonvulsant activity of new 4-aminopyrazoles and 5-aminopyrazol 3-ones. PMID- 10522276 TI - [Analysis of elements in psoriatic and non-psoriatic skin]. PMID- 10522277 TI - Effect of dianthrones and their precursors from Hypericum perforatum L. on lipoxygenase activity. PMID- 10522278 TI - Inhibition of granulocyte elastase activity by caffeic acid derivatives. PMID- 10522279 TI - Stereoisomers of 4-fluoroglutamic acid: influence on Escherichia coli glutamate decarboxylase. PMID- 10522280 TI - Detecting bouts of physical activity in a field setting. AB - The purposes of this study were to assess the TRITRAC and CSA for: (a) interaccelerometer agreement; (b) agreement in detecting patterns of moderate intensity physical activity; and (c) agreement in detecting walking patterns recorded in a diary. Thirty-one women wore both the TRITRAC and CSA accelerometers for three consecutive days. Interaccelerometer agreement (measured with generalizability coefficients) ranged from .88 to .99. In total, 71.3% of the accelerometers' patterns agreed in length, with CSA patterns being on average significantly longer. Interaccelerometer agreement in detecting patterns of brisk walking, as recorded in a diary, was comparable (69.4%). Interaccelerometer discrepancies may be related in part to the threshold employed by each instrument for classifying moderate intensity patterns. PMID- 10522281 TI - Manipulating visual informational constraints during practice enhances the acquisition of catching skill in children. AB - Previous motor learning studies examining the effects of practicing to catch one handed under varying informational constraints on subsequent skill acquisition are equivocal, perhaps due to the use of relatively inexperienced adult participants. Ecological theory predicts that directing the learner's search for information in the perceptual-motor workspace can enhance skill acquisition. This study manipulated visual informational constraints on novice children (ages 9-10 years) learning to catch one-handed. A crossover transfer design was implemented in which one group acted as controls while two other groups practiced either without visual restrictions before transferring to full vision, or vice versa. The data indicated that learners forced to seek additional information sources under restricted viewing conditions demonstrated a greater positive, accumulative residual effect on acquiring a catching skill. The findings contradict current work on the specificity of practice hypothesis and suggest that varying visual informational constraints to encourage exploratory practice may represent a significant pedagogical approach to motor learning in sport. PMID- 10522282 TI - Expert-novice differences in performance skills and problem representations of youth and adults during tennis competition. AB - Expert and novice tennis players selected from three different age groups (i.e., 10-11 years, 12-13 years, and collegiate adults) were examined for differences in performance skills (i.e., behavioral analyses of video recordings) and problem representations (i.e., verbal report analyses of tape recordings) during matched competition. Factorial analyses of variance on behavioral measures indicated that experts' performances exhibited higher levels of decision and execution than novices, regardless of age. Kruskal-Wallis tests on verbal report measures indicated that experts generated more total, varied, and sophisticated condition and action concepts than novices. Within experts, adults accessed more sophisticated problem representations than youth. Both current event and action plan profiles guided and mediated adult experts' response selections and executions, respectively. Youth experts primarily used action plan profiles to guide their response selections. Novices, regardless of age, accessed weak problem representations. PMID- 10522283 TI - Spatial assimilation effects and error detection in rapid overlapping and sequential bimanual movements. AB - This study verified earlier anecdotal evidence indicating that spatial assimilations could be reduced by offsetting movements in time. In Experiment 1, 40 right-handed participants (ages 18-23 years) made single and dual quick lever reversals of 20 degrees and 60 degrees with the left and right limbs, respectively. Participants were assigned to either the Overlapping (O) group, in which one limb began when the other limb reached the reversal point, or to the Sequential (S) group in which one limb followed the other with a delay of 114 ms, on average. The shorter-distance limb of the O group overshot relative to the S group. Short-distance spatial assimilations were also shown in the S group in Experiment 2, when the delay was increased to 250, 500, or 1,500 ms (N = 30), suggesting that assimilation effects can be caused by command interactions at both the planning and the execution levels. PMID- 10522284 TI - Physical assistance devices in complex motor skill learning: benefits of a self controlled practice schedule. AB - This study examines the effects of a self-controlled use of physical assistance devices on learning a complex motor skill (i.e., producing slalom-type movements on a ski simulator). Physical assistance was provided by ski poles. One group of learners (self-control) was provided with the poles whenever they requested them, whereas another (yoked) group had no influence on the pole/no-pole schedule. While there were no group differences during the practice phase (Days 1 and 2), clear group differences emerged in the retention test without poles (Day 3). The self-control group produced significantly larger amplitudes than the yoked group. These results extend previous findings by showing learning advantages of the self controlled use of physical assistance devices in complex motor skill learning. PMID- 10522285 TI - Creating a sense of family in urban schools using the "sport for peace" curriculum. AB - Urban students often have difficulty engaging in the learning process and affiliating with others. A three-phase research design was used to examine the effectiveness of a high school physical education curriculum reform initiative entitled "Sport for Peace" to enhance student engagement and willingness to interact positively with others. Ten physical educators in six urban schools taught a traditional soccer unit (Phase I) followed by instruction and mentoring in the Sport for Peace curriculum (Phase II). In the third phase of the research, teachers developed and taught a Sport for Peace unit to their students. Data were collected using observation and interview methods and analyzed with constant comparison. Results suggested that the Sport for Peace curricular structures fostered shared responsibility for learning, trust, respect, and a sense of family. Both high- and low-skilled girls and boys felt successful and responded positively, creating a class community more conducive to engagement and participation. PMID- 10522286 TI - An investigation into teaching games for understanding: effects on skill, knowledge, and game play. AB - The purpose of this study was to test the validity of the games for understanding model by comparing it to a technique approach to instruction and a control group. The technique method focused primarily on skill instruction where the skill taught initially was incorporated into a game at the end of each lesson. The games for understanding approach emphasized developing tactical awareness and decision making in small game situations. Two physical education specialists taught field hockey using these approaches for 15 lessons (45 min each). The control group did not receive any field hockey instruction. Data were collected from 71 middle school children. Pretests and posttests were administered for hockey knowledge, skill, and game performance. Separate analyses of variance or analyses of covariance were conducted to examine group differences for cognitive and skill outcomes. The games for understanding group scored significantly higher on passing decision making than the technique and control groups during posttest game play and significantly higher than the control group for declarative and procedural knowledge. The games for understanding group scored significantly higher on control and passing execution than the other groups during posttest game play. For hockey skill, there were no significant differences among the treatment groups for accuracy, but the technique group recorded faster times than the control group on the posttest. PMID- 10522287 TI - The relationship between salivary adrenocortical hormones changes and personality in elite female athletes during handball and volleyball competition. AB - Salivary cortisol, androstenedione delta 4, and dehydroepiandrosterone (DHEA) in 20 elite sportswomen were measured using radioimmunoassay in samples taken 5 min before and after a handball or volleyball competition. Three psychometric tests- State Trait Anxiety Inventory, Bortner, and Questionnaire de Personnalite pour Sportifs (questionnaire of personality for sports participants)--were used to evaluate the participants' personalities. Results indicated higher concentrations of cortisol and lower concentrations of delta 4 and DHEA in handball players before and after the competition. Cortisol values increased significantly during the competition in both groups. No changes were observed in androgen levels. The state of anxiety was higher in handball players, characterized by Pattern A behavior, whereas Pattern B behavior defined the volleyball players. The results suggest that adrenocortical changes during handball and volleyball competition are influenced by the different energy systems required by the two activities, individual personality characteristics, and the athlete's anxiety level relative to winning or losing. PMID- 10522288 TI - Women, disability, and sport and physical fitness activity: the intersection of gender and disability dynamics. AB - This study explores the often overlooked experiences of women with physical disabilities in the sport and physical fitness activity domain. Interviews with 16 women with a physical disability (age range of 19-54 years) revealed the following major themes: (a) participation in fitness-related as opposed to sport related activities, (b) participation to maintain the functional level of the body and preserve existing capabilities, (c) intrinsic nature of gains derived from participation (perceived competence, enhanced view of body, motivational outlet, control in life), and (d) perceived differences in the sport and physical fitness activity experiences of men and women with disabilities. Findings support the notion that gender and disability interact in the sport and physical fitness context for women with physical disabilities. PMID- 10522289 TI - Can the anticipatory skills of experts be learned by novices? PMID- 10522290 TI - Home and recess physical activity of Hong Kong children. PMID- 10522291 TI - [Free access of patients to their medical records. Letter of Professor Pierre Godeau to Mr. Lionel Jospin, Prime Minister]. PMID- 10522292 TI - [Cancer. 35th Congress of the American Society of Clinical Oncology (ASCO), Atlanta, 15-18 May 1999]. PMID- 10522293 TI - [Atherosclerosis. Congress of the European Atherosclerosis Society (EAS), Athens (Greece), 26-29 May 1999]. PMID- 10522294 TI - [Horton's disease and rhizomelic pseudopolyarthritis. First International Congress on Horton's Disease and rhizomelic pseudopolyarthritis, Prato (Italy), 25-26 May 1999]. PMID- 10522295 TI - [Prevention and treatment of vitamin D deficiency in aged patients: a proven efficacy]. PMID- 10522296 TI - [Value of the biopsy of accessory salivary glands in amyloidosis]. AB - PURPOSE: To assess the value of accessory salivary gland biopsy for the diagnosis of amyloidosis, a study was conducted in the nephrology and hemodialysis department at Ibn Rochd University Hospital from February 1996 to January 1998. METHODS: Renal amyloidosis was confirmed by renal biopsy accompanying accessory salivary gland biopsy. RESULT: The patient's mean age was 39 years old (range 15 80), with a 4:1-male/female ratio. An infectious cause (either tuberculosis or superinfection and dilatation of the bronchi) was the most frequent (70% of the cases) etiology. All the patients presented renal symptomatology. Nephrotic syndrome predominated. Amyloid deposits were observed in 100% of renal needle biopsies and in 80% of accessory salivary gland biopsies. CONCLUSION: Renal biopsy led to more positive cases than the other biopsies. It may be accompanied by severe complications. Furthermore, biopsy of the accessory salivary glands is a simple and very reliable technique for the diagnosis of amyloidosis. It is currently the best diagnostic test. PMID- 10522297 TI - [Low prevalence of anti-annexin V antibodies in antiphospholipid syndrome with fetal loss]. AB - PURPOSE: Annexin V, a protein with potent anticoagulant activity has a calcium dependent binding affinity for phospholipids. Annexin V is distributed in many organs, especially in the placenta and endothelium. Various studies have shown that placental annexin V is decreased in women with anti-phospholipid syndrome. It has been suggested that annexin V might be a target of anti-phospholipid antibodies and that the subsequent decrease in annexin V might be associated with obstetrical complications. We investigated the presence of anti-annexin V antibodies in the plasma of women with anti-phospholipid syndrome and obstetrical complications. METHODS: Twenty-three patients with at least one spontaneous abortion were included in the study. Anti-cardiolipin antibodies and lupus anticoagulant were present in 87% and 30% of the patients, respectively. A group of 40 healthy women were included in the control group. Anti-annexin V IgG and IgM antibodies were measured by ELISA. RESULTS: The IgG mean OD was 0.07 +/- 0.013 in patients and 0.042 +/- 0.06 in the control group. There was no significant difference between the two groups (P = NS). Only two out of the 23 patients and two out of the 40 healthy women were positive for IgG (OD > 0.25). The sensitivity of the assay was poor (8.7%). Even when the threshold was adjusted according to the mean OD in control subjects +2 SD, the sensitivity was still poor, reaching only 13%. CONCLUSION: The prevalence of anti-annexin V was low in patients with anti-phospholipid syndrome and repetitive spontaneous abortions. Anti-annexin V assay does not appear to be sensitive enough for the identification of anti-phospholipid antibodies that might be involved in the decrease in annexin V leading subsequently to thrombosis risk. PMID- 10522298 TI - [Hepatitis C virus infection and thyroid diseases]. AB - INTRODUCTION: The combination of hepatitis C virus (HCV) infection and thyroid diseases raises several issues that are the prevalence of thyroid autoimmunity in patients with chronic hepatitis C, the prevalence of HCV infection in patients with autoimmune thyroid diseases, and the effects of interferon alpha treatment on thyroid function in chronic HCV hepatitis. CURRENT KNOWLEDGE AND KEY POINTS: The prevalence of anti-thyroid auto-antibodies ranges from 4.6 to 15% in HCV infection, which is considered as significant by various authors. Results have to be interpreted according to the following: the type of auto-antibodies detected, the age, sex, ethnic origin of the population studied, and characteristics of the control population. Recent data are suggestive of a high prevalence of anti thyroid auto-antibodies in females with HCV infection. An increased prevalence of HCV infection in patients with Hashimoto's thyroiditis is not confirmed. During treatment of chronic hepatitis C, interferon alpha induces thyroid dysfunctions (3 to 15% of the cases) with various clinical presentations. Hypothyroidism is more common (two out of three cases) than hyperthyroidism (one out of three cases). Hyperthyroidism followed by hypothyroidism has also been described. Clinical symptoms vary, ranging from subclinical to severe manifestations. Thyroid dysfunction may be delayed after discontinuation of the interferon treatment. Hypothyroidism is easily cured by L-thyroxine replacement therapy when necessary, and regression may be observed following discontinuation of interferon treatment. Each case of hyperthyroidism has to be precisely evaluated. Development of anti-thyroid antibodies or an increase in anti-thyroid antibodies titers is often observed during interferon alpha treatment, thus suggesting the existence of immunological mechanisms at the origin of thyroid dysfunction. Furthermore, interferon would directly act on iodine. FUTURE PROSPECTS AND PROJECTS: Clinical studies are still necessary to better clarify the links between HCV infection and thyroid autoimmunity, and to determine risk factors for the development of thyroid dysfunction during interferon alpha therapy. The effects of HCV and interferon alpha on thyroid autoimmunity and function have to be investigated in basic research. PMID- 10522299 TI - [Ion-channel related muscular diseases]. AB - INTRODUCTION: Though ion channel-related muscular disorders were described long ago, better understanding of their underlying mechanisms has been more recently achieved. These mechanisms include myotonic syndromes that may be caused by mutations in sodium and chloride channels, as well as periodic paralysis which is due to mutations in sodium and calcium channels. CURRENT KNOWLEDGE AND KEY POINTS: Knowledge of the involved pathophysiological mechanisms has led to better clinical description of these disorders, as well as more efficacious treatment. In some cases, it is now possible to establish the diagnosis, using genetic tests. FUTURE PROSPECTS AND PROJECTS: Other neuromuscular disorders might be related to ion channel mutations. PMID- 10522300 TI - [Transfusion-related lung injury edema]. AB - INTRODUCTION: Transfusion-related acute lung injury (TRALI) is a relatively rare but potentially severe complication of blood transfusion that should be diagnosed. EXEGESIS: The origin of this complication is most often an immunological leucocyte conflict due to transfused plasma HLA antibodies. Prevention (anti-HLA antibody screening in blood donors and elimination of blood products from immunized donors) is discussed. CONCLUSION: Transfusion-related acute lung injury should be suspected in transfusion-related respiratory distress, after elimination of potential pulmonary edema due to overloading. An immunologic cause should always be searched for. Leukodepletion of blood products and harmonization in blood donor selection should prevent this rare complication. PMID- 10522301 TI - [Improvement in prevention of venous thromboembolism in medical environment. Mission impossible?]. AB - PURPOSE: In France, low molecular weight heparins are widely used for prophylaxis of venous thromboembolism. This practice is not based on strong evidence supporting the use of this technique, thus leading to important difficulties in defining accurate guidelines. Improvement in the prophylaxis must focus on the process leading to prescription or non-prescription. Our primary objective was to identify and describe this process. We also analyzed the causes for dysfunction and implemented corrective actions. METHODS: Basic tools and methods specific to improvement of quality were used including: flow charting for process description, causes-effect diagram for updating and analysis of dysfunction, weighted vote for decisions regarding corrective actions, and agenda and designation of leaders for the implementation of corrective actions. RESULTS: The theoretical process beginning with the recognition of a risk and ending with the adequate treatment or prevention of this risk was demonstrated. The most important dysfunction was the lack of epidemiological and clinical trials. Key corrective actions were the following: standardization of practices where evidence was available; if not, collaborative efforts to collect valid epidemiological data through multicenter surveys and clinical trials; systematic appraisal of the quality of published data; assessment of the risk for venous thromboembolism during hospitalization. To achieve real improvement in practice, priority was given to repeated measures of the risk for thromboembolism. The use of low molecular weight heparins has been so wide however, that it has led to some difficulties. We must draw lessons from this experience. CONCLUSION: Improvement of quality in terms of healthcare is not only based on auditing the practices or modifying the defective processes, but also on our ability not to use new drugs before fully assessing their validity. PMID- 10522302 TI - [Muscle, fatigue, sports and infection]. AB - PURPOSE: Skeletal muscle can be considered as motors which convert chemical energy into mechanical energy. We can evaluate the intracellular pH and energy state of phosphate-containing metabolites in skeletal muscle of patients complaining fatigue or asthenia, using phosphorus MRS. MAIN POINTS: Acute infectious disease and extreme endurance exercise may induce a loss of oxidative capacity of muscle tissue. Muscle fatigue is not due only to an insufficient supply of ATP to the energy consuming mechanisms. Phosphorus MRS show a muscle production of toxic metabolites such as lactates, protons and ammonia. These metabolic features induced excessive intracellular acidosis of skeletal muscle and systemic hyperammonia, responsible of fatigue and asthenia. PERSPECTIVES: Reversal of the excessive acidosis and improvement of the capacity for oxidative ATP synthesis might help to relieve the symptoms of exhaustion/fatigue in these patients. PMID- 10522303 TI - [Sinus dysfunction in Basedow's disease. A case report]. AB - INTRODUCTION AND EXEGESIS: The authors report a case of sinus node dysfunction that occurred in a 22-year-old patient with Graves' disease and disappeared after thyroidectomy. CONCLUSION: Bradycardia is uncommon in hyperthyroidism, the sinus node dysfunction would be due to thyrotoxicity-related myocarditis. PMID- 10522304 TI - [Pulmonary lymphangioleiomyomatosis: an often fortuitous diagnosis. A case report]. AB - INTRODUCTION: Lymphangioleiomyomatosis is an uncommon disorder of unknown origin, which exclusively occurs in women of reproductive age. The condition is characterized by proliferation of immature smooth muscle cells throughout the lungs, i.e., in the peribronchial, perilymphatic, and perivascular areas. This results in obliteration of the respiratory tract and in the development of cysts. Lymphangioleiomyomatosis has a poor prognosis due to both numerous lung complications and progression of the disease to respiratory failure. EXEGESIS: We report the case of a patient in whom lymphangioleiomyomatosis was fortuitously diagnosed from chest CT scan, itself performed for the diagnosis of pulmonary embolism. This case is therefore of particular interest. CONCLUSION: Our results suggest that the prevalence of lymphangioleiomyomatosis is probably underestimated due to its clinical latency and the absence of specific laboratory tests. Therefore, the development of non-invasive radiological methods should permit early diagnosis of the disease. PMID- 10522305 TI - [Adenocarcinoma of the exocrine pancreas: management and therapeutic hopes]. AB - INTRODUCTION: Pancreatic carcinoma is a major public health concern, as it kills more than 6,000 people each year in France. CURRENT KNOWLEDGE AND KEY POINTS: The main risk factor demonstrated by concordant case-control studies is cigarette smoking. Pancreatic carcinoma is generally diagnosed at an advanced stage. Results of radical surgery are still poor. In most of the reported series, less than 25% of the patients survive at five years. FUTURE PROSPECTS AND PROJECTS: Postoperative radiochemotherapy slightly increases the hope of cure. In locally advanced tumors, radiochemotherapy, sometimes preoperative, allows some patients to survive more than two years. Though results of palliative chemotherapy remain very poor, some clinical benefit has been observed in randomized trials comparing this treatment with the currently best supportive treatment. PMID- 10522306 TI - [Contribution of gemcitabine in the treatment of advanced pancreatic cancer]. AB - INTRODUCTION: Pancreatic cancer is one of the most common tumor of the gastrointestinal tract. CURRENT KNOWLEDGE AND KEY POINTS: Because this malignancy is usually diagnosed at an advanced stage, its prognosis is poor, and patients are generally considered incurable at diagnosis. The traditional palliative approach to management of this tumor is chemotherapy. The most widely used agent is 5-FU, alone or in combination. Benefits of the treatment are still poor: the overall survival time rarely exceeds 5 months, and no study has shown a response rate greater than 20%. FUTURE PROSPECTS AND PROJECTS: Gemcitabine, a new antinucleoside agent, has led to promising results, as several phase II and III studies have demonstrated an increase in survival as compared with 5-FU, the overall 1-year survival rates being 18% and 2%, respectively (p < 0.002). Furthermore, even if only discrete results in terms of objective response rate have been achieved, gemcitabine decreases disease-related symptoms, thus benefiting to the patient's quality of life. The concept of clinical benefit therefore appears to be an important judgement criteria in the assessment of chemotherapy efficacy, and will certainly be extended to other malignant neoplasms. PMID- 10522308 TI - [Corticoid-sensitive recurrent multifocal superficial thrombophlebitis in an 84 year-old man]. PMID- 10522307 TI - [Multiple arterial aneurysms]. PMID- 10522309 TI - [Temporal arteritis, hypereosinophilia and cryoglobulinemia]. PMID- 10522310 TI - [Giant cell arteritis of the ovary: a rare localization of Horton's disease]. PMID- 10522311 TI - [Preclinical Cushing's syndrome: diagnostic difficulties]. PMID- 10522313 TI - [Orphan diseases and society (III). Therapeutic use of intravenous immunoglobulins in internal medicine]. PMID- 10522312 TI - [Intracellular signalization and inflammation: variations for an intracellular symphony. 1st movement]. AB - Inflammatory response involves complex signalling pathways at cellular and molecular levels. During the cellular activation, intracellular substrates are subsequently phosphorylated for inducing transcription and synthesis of various inflammatory and/or anti-inflammatory mediators. This review focuses on the description of the main mechanisms involved in the activation signalling pathways that occur during inflammatory processes. PMID- 10522314 TI - [Value of intravenous immunoglobulins during antiphospholipid syndrome]. AB - BACKGROUND: The antiphospholipid syndrome was individualized 12 years ago. Treatment was initially based on steroids, immunosuppressive drugs and intravenous immunoglobulin therapy. More recently, several retrospective studies have established that in most clinical conditions therapeutic doses of oral vitamin K antagonists (INR > or = 3) are sufficient to control the disease. THE ROLE OF IMMUNOGLOBULIN THERAPY: However, high dose immunoglobulin therapy is still indicated in a few cases, especially in life-threatening immune peripheral thrombocytopenia, and in recurrent foetal loss: in the latter indication, immunoglobulin therapy alone is efficient in 80% of cases. FUTURE PROSPECTS: Prospective studies are needed to assess the efficacy of intravenous immunoglobulin therapy in neurological complications occurring in spite of anticoagulant therapy, and in the context of repeated foetal losses when antithrombotic therapy with aspirin and subcutaneous heparin has failed. PMID- 10522315 TI - [Polyvalent intravenous immunoglobulins in systemic lupus]. AB - PURPOSE: To evaluate intravenous immunoglobulins (IVIG) treatment, which is immunomodulatory but not immunosuppressive, in SLE. MAIN ISSUES: IVIG indications in SLE could be categorized as already validated as for chronic polyradiculoneuropathy or thrombocytopenia; failure of classical treatment in threatening active disease; undetermined manifestation as reactive hemophagocytic syndrome which could be both disease-specific or iatrogenic, infection-related. To date, no published study has firmly established the efficacy of IVIG in SLE. CONCLUSION: IVIG therapy in SLE should be evaluated in prospective trials. PMID- 10522316 TI - [Still's disease in adults: treatment with intravenous immunoglobulins]. AB - INTRODUCTION: Adult onset Still's disease is a rare systemic disorder of unknown etiology occurring in young adults. The diagnosis is based upon Yamaguchi criteria. Treatment is difficult and not well codified. CURRENT KNOWLEDGE AND KEY POINTS: Non steroidal anti-inflammatory drugs (salicylates, indomethacin) are used as first-line therapy but are not efficient. Steroids are needed in 80% of cases to control systemic manifestations of adult onset Still's disease. Immunosuppressive agents, such as methotrexate, are necessary when a high dose of steroids are required. The use of intravenous immunoglobulin was rarely reported, in particular in patients refractory to non steroidal anti-inflammatory drugs. Intravenous immunoglobulin was administered at 2 g/kg of body weight during two or five days. Infusion was given monthly for four-six cycles. Long-term remission was obtained in half of the patients. Precise mechanisms of action of intravenous immunoglobulin in adult onset Still's disease remain unclear. FUTURE PROSPECTS AND PROJECTS: Intravenous immunoglobulin may represent a new treatment, particularly in patients refractory to non steroidal anti-inflammatory drugs before the use of steroids. Further prospective works are needed to confirm these preliminary optimistic data. PMID- 10522317 TI - [Immunomodulatory effects of intravenous immunoglobulins in autoimmune diseases]. AB - PURPOSE: Intravenous immunoglobulins (IVIg) therapy has been reported to be beneficial in a large number of autoantibody-mediated or self-reactive T cells associated autoimmune diseases. Thus, the beneficial effect of IVIg is probably due to multiple distinct mechanisms. MAIN POINTS: The immunoregulatory effect of IVIg in autoimmune diseases is dependent on: interaction of the Fc portion of IVIg with Fc receptors on leucocyte surfaces; interaction of infused IgG with complement components; modulation of synthesis and release of cytokines produced by lymphocytes and monocytes: V region-dependent neutralization of circulating autoantibodies by infused IgG; selection of immune repertoires; modulation of cell proliferation, particularly through modulation of Fas-induced apoptosis; interaction of IVIg with numerous other molecules onto the surface of T and B cells. PERSPECTIVES: Better understanding of these mechanisms should allow a better definition of the spectrum of diseases likely to benefit from IVIg treatment. PMID- 10522318 TI - [Treatment of ANCA-positive systemic vasculitis with intravenous immunoglobins]. AB - PURPOSE: ANCA positive systemic vasculitis comprises Wegener granulomatosis (WG), microscopic polyangiitis (MPA) and Churg Strauss syndrome (CSS). In WG, anti-Pr3 ANCA are present in 90% of the cases, whereas in MPA and CSS ANCA are present in 40 to 80% and 25 to 60% of the cases, respectively, with anti-MPO specificity. The treatment of WG and MPA associates prednisone and cyclophosphamide, and clinical remission is obtained in 70 to 90% of the cases. However, relapses occur in 10 to 30% of the patients and tolerance of corticosteroids and immunosuppressive therapy is not always good. STATE OF THE ART: Intravenous immunoglobulins (IVIg) are well tolerated and indicated in a large number of autoimmune and systemic inflammatory diseases. In vitro IVIg specifically inhibit ANCA activity through V-region dependent interaction of anti-ANCA anti-idiotypic antibodies with ANCA. Several open studies and a few case reports of patients with WG or MPA treated with IVIg were published in the past seven years, with various results. PROJECTS: A prospective controlled multicenter study is necessary to evaluate the efficacy of IVIg in the treatment of ANCA positive vasculitides. PMID- 10522319 TI - [Intravenous immunoglobulins in polymyositis and dermatomyositis]. AB - PURPOSE: Polymyositis (PM) and dermatomyositis (DM) are inflammatory idiopathic myopathies of dysimmune origin. Studies have shown two different pathogenic mechanisms: a primitive vascular mechanism mediated mainly by a humoral mechanism responsible for a muscular ischemia in juvenile DM; and a primitive injury of muscle cells mediated by a cytotoxic cellular mechanism, directed against the myofibrillae, in PM. CURRENT KNOWLEDGE AND KEY POINTS: Oral corticosteroids are the treatment of first choice in patients with PM/DM. This therapy has transformed the prognosis of these diseases. The efficacy of polyvalent human intravenous immunoglobulin (IVIG) was first described in an open study with severe refractory PM and DM, and confirmed by a controlled study in DM. FUTURE PROSPECTS AND PROJECTS: We present our experience about the interest of intravenous immunoglobulin (IVIG) in refractory PM and DM and discuss the different mechanisms of action of this new immunotherapy. PMID- 10522320 TI - [Intravenous immunoglobulins and autoimmune thrombopenic purpura]. AB - OBJECTIVE: To determine the usefulness of intravenous immunoglobulin in the treatment of autoimmune thrombocytopenic purpura (AITP). RESULTS: IVIg infusion is an important agent in managing AITP because most patients experienced a rapid increased platelet count. Its precise place in the management of these patients is however controversial, particularly on account of its high cost and transient effect. It is indicated in the severe form of the disease or before surgery in corticoresistant patients. PERSPECTIVE: Prospective studied are mandatory to compare the efficacy of IVIg and high dose corticosteroids in this indication. PMID- 10522321 TI - [Eckelt's technic: an evaluation of the access route used at Salpetriere Hospital]. AB - AIM: We report a prospective study undertaken to investigate the morbidity connected with the approach of lag screw osteosynthesis for the treatment of subcondylar fractures in the department of Maxillo-Facial Surgery at the Salpetriere University Hospital. METHOD: 24 patients have been operated between November 1997 and December 1998. We estimated the accessibility, the size and quality of scars the satisfaction index, and the complication rate. RESULTS: The average size of scars was 3 cm (2.5-4). Their quality was satisfactory, except for an hypertrophic and an invaginated one. The satisfaction index was always excellent and often superior to the surgeon one. COMPLICATIONS: three paresis in the mental territory of the facial nerve regressive in three months, a salivary fistula regressive in three weeks, an hematoma and a subcutaneous infection. These different criteria improved during the study. CONCLUSION: The approach is not free of risks but these ones seem to decrease with surgeon's experience. PMID- 10522322 TI - [The architectural balance of the face: a 3D cephalometric concept]. AB - The use of the c2000 software led us to develop a new 3D cephalometric construction, based on the selection on the axial CT Scans of 8 anatomical landmarks and of teeth, all of which were situated along the trigeminal neuro matriciel facial growth axes. The analysis of this construction is based on the use of an original mathematical tool in biology: the axes of inertia. Using the selection of both mental foramen, both infra and supra orbital foramen and the head of both malleus, the C 2000 software creates a geometrical construction called: "the maxillo-facial frame", as well as, a 3D cephalometric analysis: angles, distances, areas, volumes center of gravity and axes of inertia. Using the selection of teeth, the C 2000 software calculates the axes of inertia of each tooth or of groups of teeth. The use of the axes of inertia allow us to create of a hierarchy of anatomical levels the teeth, the half arches, the arches, both arches and the maxillo-facial frame. In addition, for each of these anatomical levels, the axes of inertia create a 3D landmark which allows the calculation of the orientation of each of these elements in relation to the others. The study of 28 orthomorphic people using this analysis revealed the existence of a maxillo-facial balance that is unique for each individual. PMID- 10522323 TI - [A new application of extraoral implants: the permanent percutaneous electrical connection. Apropos a case]. AB - The principle of EOI is an implant fixed to the bone supporting a percutaneous abutment. Placed through the skin, the abutment allows a permanent communication between inside and outside of the organism. A rigorous very precise surgical protocol has allowed good results of the technique for over 20 years and shown the reliability of this two-piece device. This surgical protocol and device concept have been used by the authors to design a percutaneous electrical connector experimented in a rabbit model since 1996. A percutaneous electrical connector called PEP (percutaneous electrical plug) directly steming from the Sweden and German extra oral implants as been design for human applications. Authors describe the surgical placement and loading of there first PEP, under Huriet law, in an otologic application: the treatment of sever, drug-resistant tinnitus by electrical stimulation of the internal ear. The anatomical implantation area of these new extra-oral implants is retro-auricular. So the implants placements should be realized by ENT and maxillo-facial surgeons. PMID- 10522324 TI - [Stomatologic and maxillofacial pathology in a medieval population (10th-12th centuries) of southwestern France]. AB - We present an assessment of the dental and maxillofacial pathology in a medieval population in southwestern France. One hundred and ninety eight mandibles and 29 craniofacial complexes were analysed. Dental and periodontal infectious pathology predominated. Third molar agenesia was quite frequent, concerning 25% of the mandibles. Third molar eruption was almost constant and in a normal position. Condylar process degeneration concerned 6% of the population. Three cases of traumatic pathology were observed, one case of long mandible was noted, and two cases of hypertrophic inferior alveolar process. Dento-mandibular maladjustment was uncommon. No unwedging of the maxillo-mandibular bone basis was observed. PMID- 10522325 TI - [Lithiasis of a minor salivary gland. Apropos a case]. AB - Sialolithiasis of minor salivary glands is generally considered to be extremely rare, particularly in younger persons. In addition, the symptomatology of this entity is not always typical and, for this reason, clinical misdiagnosis is possible. A case of sialolithiasis of minor salivary glands of the cheek in a 20 year-old woman is reported. The clinical diagnosis is verified by the histopathological examination after surgical excision of the lesion. PMID- 10522326 TI - [The clinical and economic assessment of surgery in the treatment of the obstructive sleep apnea syndrome. The Working Group assembled by the ANAES. L'Agence Nationale d'Accreditation et d'Evaluation en Sante. A synthesis]. PMID- 10522327 TI - [Apropos the report on the clinical and economic assessment of surgery in the treatment of the obstructive sleep apnea syndrome]. PMID- 10522328 TI - Neonatal seizures: unusual causes. AB - Seizures are the most common manifestation of neurological dysfunction in the newborn. The causes of newborn seizures are manifold with etiological determination important with respect to treatment, prognosis, and recurrence risk perspectives. This article highlights two cases with unusual, genetically based causes for newborn seizures. These cases are used to highlight the diagnostic approach to this clinical problem. PMID- 10522329 TI - A 5-month-old with intractable epilepsy. AB - The nosology of early infantile seizures and epilepsy syndromes is controversial. There are two age-related early infantile epileptic encephalopathy syndromes that are characterized by early onset of spasms, a burst-suppression EEG pattern, poor response to treatment, and poor prognosis. This paper examines the early myoclonic epilepsies. PMID- 10522330 TI - A 13-year-old with an acute change in mental status. AB - A 13-year-old boy with Lennox-Gastaut syndrome characterized by absence, myoclonic, complex-partial, and secondarily generalized tonic-clonic seizures, presents with progressive obtundation and loss of motor and verbal skills over a 2-day period. Initial evaluation revealed therapeutic phenytoin serum concentrations. This article discusses the differential diagnosis and management approach used in this setting, as well as the appropriate interpretation of antiepileptic drug serum concentrations. PMID- 10522331 TI - A retarded boy with seizures precipitated by stepping into the bath water. AB - This report describes the unusual case of a boy with partial deletion in the proximal region of a long arm of chromosome 2 and reflex epilepsy. Seizures with identical onset were precipitated predictably by two independent triggers, micturition and immersion of the feet in tepid or hot water. A seizure precipitated by immersion could be reproduced under video and EEG monitoring. The EEG seizure onset was in the central midline region. PMID- 10522332 TI - Hypertonia, hyperreflexia, and excessive startle response in a neonate. AB - Following an uneventful gestation, a newborn girl presented with hypertonia, hyperreflexia, tremor, and excessive startle response. Nose tap elicited a dramatic head recoil. Her mother had similar symptoms beginning as a child that improved but persisted into adulthood. In addition, several members of mother's family died unexpectedly in infancy. Hypertonia in the newborn period indicates central nervous system dysfunction of several possible causes, most of which are associated with severe cognitive deficits and limited neurological development. PMID- 10522333 TI - A 4-year-old with pica, progressive incoordination, and decreased responsiveness. AB - This article reports a typical case of subacute sclerosing panencephalitis (SSPE). The patient contracted measles as an infant during the 1989 to 1991 United States measles epidemic. At 4 1/2 years of age, he developed behavioral changes and quickly progressed through the typical clinical stages of SSPE. His EEG was characteristic. Serum and CSF measles immunoglobulin G were markedly elevated. He remains alive but is vegetative. To our knowledge, this is the first case of SSPE stemming from the 1989 to 1991 measles epidemic. Because infants- the group at highest risk to develop SSPE--were most severely affected by this measles outbreak, other cases of SSPE stemming from this epidemic may occur. PMID- 10522334 TI - Subacute encephalopathy in a 5-year-old boy. AB - A 5-year-old boy presented with an acute ataxia and altered mental status. Although he initially recovered from these symptoms, he presented a second time with myoclonus and seizures and rapidly became vegetative. Cerebrospinal fluid studies, magnetic resonance imaging, and brain biopsy all confirmed the presence of subacute sclerosing panencephalitis. Despite courses of therapy with cimetidine, amantadine, ribavirin, and inosine, no clinical improvement has been seen. Clinicians need to be alert to the possibility of subacute sclerosing panencephalitis even in the vaccinated child in the appropriate clinical setting. PMID- 10522335 TI - An adolescent with complicated migraine. AB - During an evaluation for complicated migraine, a 14-year-old adolescent female was found to have a left frontoparietal cortical infarction on magnetic resonance imaging study. A transthoracic echocardiogram was normal, but a transesophageal echocardiogram, with contrast study, showed occasional right to left shunting through a patent foramen ovale. The role of cardiac anomalies in the pathogenesis of migraine-associated stroke is discussed. PMID- 10522336 TI - New-onset headache in an adolescent with MASS syndrome. AB - A 15-year-old girl with the "MASS" phenotype (meeting several of the minor criteria for Marfan syndrome) presents with a new onset low-pressure postural headache. Clinical features and magnetic resonance imaging suggested intracranial hypotension, which was confirmed with lumbar puncture. The pathophysiology and treatment of spontaneous intracranial hypotension are discussed. PMID- 10522337 TI - An 8-year-old girl with unilateral facial and ear pain and isolated frontal headaches. AB - An 8 1/2-year-old with chronic but fluctuating unilateral facial pain, earache, frontal headache and facial swelling is presented. Her journey through the health care system provides an instructional lesson for all who deal with patients with unusual or difficult to recognize conditions. PMID- 10522338 TI - A 15-year-old with back pain, fever, and leg numbness. AB - Spinal epidural abscess (SEA) is an uncommon entity. We report an adolescent presenting with fever and back pain beginning 3 months after a leg abscess. This case highlights several important aspects of the diagnosis and care of patients with SEA. As illustrated by this case, plain radiographs and computed tomography of the spine can miss the diagnosis, thus when spinal epidural abscess is suspected, magnetic resonance imaging is the imaging modality of choice. Epidural abscesses most commonly arise from hematological dissemination, with Staphylococcus aureus being the most often cultured organism. Surgical intervention early combined with the administration of proper antibiotics leads to the best outcome. PMID- 10522339 TI - Focal seizures with single small ring-enhancing lesion. AB - An 8-year-old girl presented with simple partial seizures. The differential diagnosis and evaluation point out the fact that in most of the world, conditions considered rare in the United States are important diagnostic considerations. PMID- 10522340 TI - An 11-year-old girl with syndrome of inappropriate antidiuretic hormone secretion. AB - An 11-year-old girl presented with a syndrome of inappropriate antidiuretic hormone secretion, which was transitory and, initially, of obscure origin. Subsequently, the patient's hypothalamic disorder emerged as a component of a steroid-responsive relapsing encephalomyelitis with cerebral pathology restricted to the basal ganglia and brainstem. Where such a disorder fits in the spectrum from acute disseminating encephalomeylitis to multiple sclerosis is discussed. PMID- 10522341 TI - A 16-year-old with episodic hemisdystonia. AB - The paroxysmal dyskinesias are a subset of the hyperkinetic movement disorders characterized by their episodic nature. Classification based on precipitating factors is helpful in considering treatment and prognosis. The clinical similarities with partial seizures are discussed. An approach to differential diagnosis, diagnostic evaluation, and treatment options are presented. PMID- 10522342 TI - Acute onset of chorea and dystonia following a febrile illness in a 1-year-old boy. AB - A 12-month-old boy with acute onset hemichorea and dystonia following a gastroenteritis has abnormal signal intensities of his basal ganglia on brain magnetic resonance imaging (MRI). A rigorous laboratory investigation is successful in diagnosing his rare condition. A discussion of the differential of abnormal basal ganglia on MRI is presented to help illustrate this case. PMID- 10522343 TI - A child with severe head banging. AB - We present a 7-year-old boy with a developmental disorder presenting with severe head banging. Clinical evolution was consistent with diagnosis of autistic spectrum disorder, obsessive compulsive disorder, stuttering, and Tourette's syndrome. This report emphasizes the overlap between developmental disorder phenotypes. There is a need to understand the natural history and relationship of specific symptoms that occur in developmental disorders to devise effective and appropriate intervention strategies. PMID- 10522344 TI - An unresponsive infant in the emergency room. AB - The physician must be aware of the computed tomography appearance of an acute hyperacute subdural hematoma in child abuse and not mistake it for chronic subdural hematoma with "spontaneous" rebleeding. As always, the imaging findings must be correlated with the clinical findings. Clinical and imaging findings of injury out of proportion to the history, and injuries of different ages are the key indicators to the possibility of child abuse, particularly when encountered in a young infant. PMID- 10522345 TI - Acute encephalopathy and intractable seizures in a 10-year-old boy. AB - We report a 10-year-old child with Robinow's syndrome who had a 2-week history of headaches and dizziness. On the day of admission, he developed a focal onset seizure with rapid secondary generalization. The seizures were intractable despite adequate doses of benzodiazepine, phenytoin, and phenobarbital, requiring a pentobarbital drip. Continuous electroencephalogram (EEG) monitoring showed persistence of the epileptiform discharges for 13 days. Cerebrospinal fluid and brain biopsy studies were unrevealing. Mycoplasma pneumonia titers showed elevation of both immunoglobulins G and M that doubled during the tenth hospital day. High-dose methylprednisolone was begun, and within 12 hours of initiation the patient sat up and began to follow commands appropriately. The overall EEG background markedly improved. Central nervous system Mycoplasma pneumoniae infection should be suspected in patients with an encephalopathy of unclear etiology. PMID- 10522346 TI - A child with reading impairment and a family history of adrenoleukodystrophy. AB - The clinical course of a 6-year-old boy with adrenoleukodystrophy (ALD) who underwent allogeneic stem-cell transplantation during an early clinical stage is described. Twenty-three months after transplant, he remains neurologically stable, but with moderate neurological sequelae; the serum very long chain fatty acid profile has improved, but not normalized. The indications, mechanism of action, and complications of bone marrow transplantation in ALD are discussed briefly, along with other potential therapies. PMID- 10522347 TI - Hypothalamic dysfunction with polydactyly and hypoplastic nails. AB - Attention to a careful diagnostic work-up should be given to patients with postaxial polydactyly or apparently isolated hypothalamic hamartomas. Early recognition of the condition is critical for management, particularly for anticipating and preventing endocrine emergencies. Conservative management of the hypothalamic hamartoma should be stressed. Finally, detailed genetic counseling should be provided to the family regarding autosomal-dominant inheritance as well as the wide range of interfamilial and intrafamilial variability. PMID- 10522348 TI - The utility of the determination of CTG trinucleotide repeat length in hypotonic infants. AB - A 9-month-old male infant was floppy from birth with nonprogressive facial and distal limb weakness and apparently normal mother and father. The facial characteristics and distribution of involvement suggested congenital myotonic dystrophy and the infant, but not the mother, had insertional myotonia in one of four muscles tested. Had the number of CTG trinucleotide repeats been tested when the presence of a congenital myotonic dystrophy-like clinical picture was first appreciated, the proper diagnosis could have been made several months earlier. The application of new molecular genetic techniques is changing the usual sequence of studies performed in the evaluation of the hypotonic infant. PMID- 10522349 TI - [Analysis of survey data about the effect of vaccination of sows against blue pig abortion (PRRS) in the Netherlands]. AB - Questionnaires were used to collect information on the experience of pig breeders with the vaccination of sows against PRRS. There was a 1-year interval between the two questionnaires. Sixty-two pig breeders returned the first questionnaire and 36 returned the second questionnaire. Before they started vaccination, 61% of the pig breeders sent samples to the laboratory of the Animal Health Service for investigation. At least one blood sample was positive for PRRS antibodies, but PRRS virus was not detected in any of the samples. Repeat breeding was the most important reason to vaccinate sows. On 54 farms (77%) sows were first vaccinated during the suckling period, on one farm (2%) sows were vaccinated during gestation, and on 7 farms (11%) all sows were vaccinated irrespective of their phase of reproduction. Repeat vaccinations were given during the suckling period on all farms. Mild side effects were reported on 16 farms (25%). Repeat breeding was the most important adverse reaction, followed by anoestrus. After 1 year, 58% of the pig breeders expressed satisfaction with vaccination. Twenty-eight pig breeders stopped vaccinating their sows in the first year, mainly because vaccination had no effect on the technical results. The effect of PRRS on the technical results of eight farms before and a year after vaccination was analysed. Although technical results were slightly better after vaccination, it was not clear whether this improvement was due to vaccination. From the results of this study it can be concluded that, without good diagnostic investigations, the routine vaccination of sows against PRRS is not economically viable in herds infected with PRRS virus. PMID- 10522351 TI - [Predecessors: veterinarians from earlier times (35). Gerardus Johannes Hengeveld (1814-1894)]. PMID- 10522352 TI - [Reply of Slappendel to the commentary, "From the Editor." No word wrong or too much]. PMID- 10522350 TI - [Navicular disease in the hind limb of a Warmblood horse]. AB - A 12-year-old Dutch Warmblood mare was admitted to the clinic with a 1-month history of lameness of the left hind leg. After clinical and radiological examinations and an bursascopy, the diagnosis navicular disease was made. The therapy consisted of stallrest, non-steroidal anti-inflammatory drugs, and orthopaedic shoeing plus intrabursal injections of short-acting corticosteroids and hyaluronic acid. The therapy was repeated following recurrence of the lameness. According to the owner the horse is currently performing at its previous level. PMID- 10522353 TI - [Does sonography have a ranking in sports injuries of the hand?]. PMID- 10522354 TI - Diagnosis of digital flexor tendon annular pulley disruption: comparison of high frequency ultrasound and MRI. AB - PURPOSE: The objective of this study is to assess accuracy of ultrasound (US) and MRI to diagnose completely disrupted annular pulley ligaments (APL) of the flexor tendons of the fingers. PATIENTS AND METHODS: In a prospective study 32 patients (all males 18-42 years, mean age 25 years) with clinically suspected annular pulley disruption were studied with US and MRI. As a control group we investigated 40 fingers of 10 healthy volunteers and a non-injured finger of the patients with US. A 10 MHz linear array was used for US examinations and a 1.5 Tesla unit for MRI. Imaging was performed between the accident and surgical repair within 6 weeks. Examinations were performed in an extended and forced flexed finger position. RESULTS: Complete disruption of the APL was diagnosed by US and MRI in 14 cases confirmed by surgery. The typical sign of disruption was a marked volar displacement of the flexor tendon in the area of the torn annular pulley during forced flexion. A distance between tendon and bone at the area of the ruptured pulley of 3 mm during extension and a distance of 5 mm at flexion was typical for complete disruption. CONCLUSION: Both diagnostic modalities, US and MRI are valuable techniques in diagnosing complete disrupted annular pulleys of the flexor tendons. PMID- 10522355 TI - [Area reduction in carotid stenosis of the internal carotid artery]. AB - AIM: In patients with atherosclerotic extracranial internal carotid artery (ICA-) stenosis the diagnostic value of colour Doppler energy (CDE)-coded duplexsonography was compared to three other methods: continuous wave (cw) Doppler peak systolic frequency (pF), pulsed wave (pw) Doppler peak systolic velocity (pV), and intraarterial digital subtraction angiography. METHODS: In 58 patients who suffered from 60 moderate to severe ICA stenoses, B-mode sonography combined with CDE-coded duplex sonography was applied to measure the extent of the stenosis by determining the residual lumen width. Results were correlated to pF and pV and with various angiographic indices. RESULTS: The determined values of the degree of stenosis were correlated to the measurement of pV (r = 0.441, p < 0.01), but not to pF (r = 0.122, n.s.). The best correlation to angiography was obtained when the linear ICA diameter was compared to the distal common carotid artery (common carotid artery index) (r = 0.214, n.s.). Sensitivity, specificity and diagnostic accuracy were comparable to the different frequency-based measurements, but the positive predictive value was lower. CONCLUSIONS: Determination of the degree of stenosis based on CDE alone is not reliable enough to allow correct diagnosis of severe carotid artery stenosis. In combination with the peak frequency method is's diagnostic value could be improved. This requires verification in a separate study. PMID- 10522357 TI - [Soft-tissue sonography in the follow-up care of breast cancer: indications of liver metastases caused by lymphatic spread]. AB - AIM: During follow-up care of breast cancer patients a prospective study was carried out to answer two questions: 1. Can soft-tissue sonography improve early detection of locoregional metastases? 2. Can a combination of soft-tissue sonography and ultrasound liver examination provide information about the lymphogenous spread of metastases? METHOD: 312 unselected breast cancer patients were examined sonographically. The examinations included 9 lymph node regions of the efferent lymphatic ducts, along with the chest wall and the liver. Further examinations of the soft-tissue metastases were carried out with a different imaging technique or histologically, or with a combined sonographic and clinical follow-up, as a control. RESULTS: Sonography revealed 112 locoregional metastases in 49 patients (15.7%); 62 of these metastases (55%) were non-palpable. Especially metastases in the axillary level II and III areas, interpectoral and intramuscular recurrences and internal mammary lymph node metastases were found to be non-palpable, except in a very advanced stage. Liver metastases often occurred in conjunction with internal mammary lymph node metastases. Liver metastases were the only manifestation of distant metastasis in three patients with internal mammary lymph node metastases, all of whom were in the fifth year after mastectomy. CONCLUSION: Soft-tissue sonography greatly improves early detection of locoregional metastases of breast cancer. There seems to be the possibility that liver metastases of breast cancer can develop lymphogenously via internal mammary lymph node metastases. PMID- 10522356 TI - [Intima media thickness in patients with vertebrobasilar and carotid stenosis/occlusions]. AB - AIM: The purpose of this study was to investigate the degree of atherosclerosis in patients with stenoses or occlusions in the vertebrobasilar system (VBS) and the carotid system (CS). METHOD: The Intima-Media-Thickness (IMT) and the diameter of the common carotid artery (CCA) (as parameters of the proliferative and dilatative form of atherosclerosis) were measured on both sides on areas of vessel wall without stenoses and plaques. We examined of the neck 134 people, including 32 normal healthy counds, 57 patients with stenoses or occlusions in the CS and 16 patients with macroangiopathy exclusively in the VBS (following doppler- and duplex sonographical criteria). RESULTS: In control persons, a IMT of 0.67 +/- 0.2 mm and a CCA-diameter of 5.8 +/- 1.2 mm was found. Patients with CS-macroangiopathy exhibited a statistically significant increase in IMT with 0.97 +/- 0.2 mm (p < 0.001) and dilatation of the CCA (6.6 +/- 1.2 mm, p < 0.01). In comparison to the controls we also found a significant increase in IMT (0.86 +/- 0.2 mm, p < 0.001) and in diameter (6.8 +/- 0.9 mm, p < 0.01) in patients with VBS-macroangiopathy. There was no significant difference between both groups, despite a tendency of less severe changes in patients suffering from VBS macroangiopathy. Patients with diabetes (1.1 +/- 0.2 mm, p < 0.001), with hypertension (0.99 +/- 0.2 mm, p < 0.05) and with coronary heart disease (0.96 +/ 0.2 mm, p < 0.05) showed a significant increase in IMT. CONCLUSION: Parameters of generalized atherosclerosis do not significantly differ between patients with stenoses or occlusions in the VBS and patients with changes in the CS and are correlated to the classical risk factors. PMID- 10522358 TI - [Three-dimensional ultrasonography and intraoperative navigation:a new application of ultrasonograms in osteotomy of the proximal femur]. AB - AIM: As precise operative control is difficult to achieve, the accuracy of femoral osteotomies can only be estimated. Therefore, a computer-assisted ultrasound navigation system was developed in order to apply an on-line control for femoral osteotomies. METHOD: Three ultrasound emitters were fixed on a triangle. The exact position of triangles could be determined by measuring the time the ultrasound beam takes to reach microphones positioned in a frame. With a reference triangle fixed distally to the osteotomy and a second triangle fixed on the surgical chisel the exact correction angle can be determined three dimensionally. RESULTS: A high degree of accuracy was found in both laboratory trials and in simulation trials using pig femurs. The deviation of measured values compared to a laser beam control was less than 0.5 degrees. CONCLUSION: The system was introduced into our operating theatre as an optimised control device that can provide excellent support to the surgical procedure. PMID- 10522359 TI - [Follow-up control of ultrasonographic neonatal screening of the hip]. AB - 1656 newborns were examined by ultrasound for early diagnosis of CDH in a screening program. In 2.5% of the cases, dysplasia or dislocation of the hip was diagnosed and required treatment. In all cases the final outcome after adequate treatment showed a hip joint without any abnormalities. 11.3% of type IIa-hips (using Graf's classification) did not reach an alpha-angle of 60 degrees until the 12th week. They were then therefore classified as type IIb. 97% of all hip joints were judged as normal in the course of the 4th month. This study emphasizes the necessity of a general ultrasound screening program. It also demonstrated that early and adequate treatment leads to a normal development of the joint. PMID- 10522360 TI - [Classification of intracranial hemorrhage in premature infants]. AB - The most common classification of intracranial haemorrhage in premature infants into four degrees of severity is based on the results of CT-scans. However, this classification does not adequately account for some pathophysiological and morphological changes. For this reason, the paediatric section of DEGUM developed a new method of classification. This classification distinguishes more precisely between the bleeding itself and secondary changes, such as posthaemorrhagic ventricular dilation, which were excluded from the revised classification. The new system contains three levels: Grade I: subependymal haemorrhage, grade II: intraventricular haemorrhages taking up < 50% of the ventricular volume, grade III: intraventricular haemorrhages of > 50% of ventricular volume. Areas of increased echo levels within the brain tissue (formerly grade IV) which are caused by haemorrhagic infarction are now taken as a separate entity. The morphological description lists the side and the location of the haemorrhagic infarction as well as its size, which is classified into 'small' (< or = 1 cm in diameter), 'medium' (1 < or = 2 cm) and large (> 2 cm). Bleeding into the basal ganglia, cerebellum and brainstem are separate entities. In post-haemorrhagic ventricular dilation the distinction is made between self-limiting transient dilation and hydrocephalus requiring treatment. PMID- 10522361 TI - [Intact unilateral twin pregnancy--sonographic diagnosis and laparoscopic treatment]. AB - Ectopic twin pregnancy is rare. Most frequently it occurs as a heterotopic pregnancy located simultaneously in the the fallopian tube and in the uterus. An unusual case of an intact unilateral ectopic twin pregnancy in the 7th week of gestation was diagnosed at this early stage by TVS (transvaginal ultrasound). The patient was successfully treated by laparoscopic salpingectomy. A review of the current literature is discussed. PMID- 10522362 TI - [Can the function of the transjugular intrahepatic portosystemic shunt be evaluated noninvasively by Doppler sonography?]. AB - Transjugular intrahepatic portosystemic shunt (TIPS) is an effective treatment of complications due to portal hypertension. Possible shunt stenosis or shunt occlusion make periodical assessment of stent function necessary. Direct portal venography is the gold standard in morphologic and functional surveillance of TIPS. Controversially discussed is whether Doppler ultrasonography is effective in hemodynamical evaluation of TIPS and sufficient in prediction of shunt dysfunction. In 39 patients, 147 Doppler ultrasonographical examinations were performed and correlated with the results obtained by direct portal venography in TIPS follow-up, 43 of 47 hemodynamically relevant stenoses, including six shunt occlusions, were being diagnosed correctly by Doppler ultrasonography, by assessing maximal flow velocity in portal vein proximal to the TIPS (sensitivity 91.5%; specificity 97%). If Doppler ultrasonographical measurement of maximal flow velocity was performed within the proximal stent itself, sensitivity was only 70.4% and specificity 27%. In conclusion, assessment of portal maximal flow velocity more accurately represents hemodynamical TIPS function than Doppler ultrasonographical measurement within the proximal stent tract itself. Since, according to data presented. Doppler ultrasonography appears to be able to detect hemodynamically significant TIPS stenoses, it might reduce the number of invasive angiographies and thus contribute to more cost-effective follow-up of TIPS patients. PMID- 10522363 TI - [Treatment of Barrett esophagus with argon plasma coagulation with acid suppression--a prospective study]. AB - 26 patients with a Barrett's esophagus of at least 2 cm length (medium 4.9 cm) were treated with a combination of repeated argon-plasma-coagulation (APC) and a long-term acid suppression using proton pump inhibitors controlled by a 2 h pH monitoring. Eleven out of 26 patients (42%) showed endoscopically and histologically a complete eradication of the metaplastic cylindric epithelium (intention--to treat analysis). Nine patients (35%) had an endoscopic complete remission but remnants of the cylindric epithelium were found at the histologic examination. On an average one APC-session was necessary for 1 cm of initial length of the Barrett's esophagus. No serious complications were seen. It remains unclear if this therapy can reduce the long-term risks for adenocarcinoma of the esophagus in patients with Barrett's esophagus. PMID- 10522365 TI - [Implantation metastasis at the exit site after percutaneous endoscopic gastrostomy in esophageal carcinoma]. AB - In patients with esophageal cancer causing obstruction percutaneous endoscopic gastrostomy (PEG) is a well-established procedure with a low complication rate to provide sufficient enteral nutrition. We report on a 68-year-old patient suffering from inoperable squamous cell cancer of the proximal esophagus, who underwent PEG insertion prior to a combined radiochemotherapy. Initially bougienage was performed because of subtotal esophageal stenosis. Four months later a metastasis was found at the PEG exit site with involvement of the gastric wall, most likely caused by spread of tumor cells during insertion of the PEG. PMID- 10522364 TI - [Aorto-duodenal fistula--a rare cause of upper gastrointestinal hemorrhage]. AB - Upper gastrointestinal hemorrhage is the most frequent indication for emergency endoscopy. In most of the cases the bleeding is primarily treated endoscopically. The duodenal penetration of an aortic graft implant is a rare condition, only accidentally diagnosed by endoscopy. This condition represents a difficult situation for the endoscopist since it is usually not included in the differential diagnosis. Additionally hemostasis can not be achieved by endoscopic intervention. Therefore the instant realization of this dangerous diagnosis is extremely important, because only surgical therapy in a center of vascular surgery may save the live of the patient. PMID- 10522366 TI - [35-year-old patient with metastasized carcinoid of Vater's ampulla--case report and review of the literature]. AB - We report the case of a 35-year-old female patient with a metastasized carcinoid of the papilla of Vater which is a rare lesion. 96 cases have been published in world literature previously. The carcinoid of the papilla of Vater appears typically as a hormone inactive tumor. It becomes symptomatic by cholestasis and jaundice in most cases and not by carcinoid-syndrome. An association with von Recklinghausen's disease as described in 25% of cases was not given in our patient. In contrast to the duodenal carcinoid there is no linear relationship between primary tumor size and incidence of metastases. The correct diagnosis was proven by histologic and immunohistochemical methods on specimen taken after endoscopic papillotomy. In spite of sensitive diagnostic methods like endosonography and somatostatin-receptor-scintigraphy exact staging was made intraoperatively in this case. Three months after pylorus pancreatoduodenal resection with lymphadenectomy the patient remained well with no evidence of tumor recurrence. PMID- 10522367 TI - Gastrointestinal involvement in patients with diabetes mellitus: Part I (first of two parts). Epidemiology, pathophysiology, clinical findings. AB - In patients with type 1 or type 2 diabetes mellitus disturbances of the gastrointestinal transit are well recognized. In decreasing order of frequency, transit disturbance through the colon, stomach, small intestine and esophagus as well as altered motility of the gallbladder occur. Acute changes of blood glucose concentrations have a major, however, reversible influence on motility in various parts of the intestinal tract. Long-term hyperglycemia may influence the incidence of gastrointestinal involvement via the occurrence of neuropathic changes of the autonomic nervous system. Early satiety, nausea, vomiting, weight loss, constipation, diarrhea and epigastric pain are often reported. These symptoms and recurrent episodes of hypoglycemia or prolonged hyperglycemia can result from intestinal transit disturbances. PMID- 10522368 TI - Gastrointestinal involvement in patients with diabetes mellitus: Part II (second of two parts). Diagnostic procedures, pharmacological and nonpharmacological therapy. AB - Diagnostic work-up in patients with diabetes mellitus, in whom gastrointestinal involvement is suspected comprises the assessment of gastrointestinal transit times, endoscopy, esophageal pH-metry or manometry, sonography and lactulose- or glucose-H2-breath tests. Prokinetic agents such as metoclopramide, cisapride or erythromycin and substances like loperamide, octreotide or clonidine are used to improve gastrointestinal dysfunction. Furthermore, in recent trials which aimed for the modulation of gastrointestinal transit times, cholecystokinin, proteinase inhibitors or amylin were administered in patients with diabetes mellitus. Nonpharmacological interventions include pancreas and kidney transplantation, gastric pacemakers and biofeedback training. PMID- 10522369 TI - [Carbohydrate-deficient transferrin (CDT) is modified by iron status: further reservations regarding the value of a new alcohol marker?]. PMID- 10522370 TI - [Hepatorenal syndrome: a new therapeutic approach?]. PMID- 10522371 TI - [Ulcerative colitis: fate of pediatric ileoanal pouches]. PMID- 10522372 TI - [Persistent follicle syndrome: a forgotten clinical entity?]. AB - There is no evidence that morphologic alterations of the ovaries cause symptoms of hormonal imbalance and a deficit in ovarian hormones in a case of micro-cystic degeneration of the ovaries. Similarly, fertility does not have to be impeded, since follicular development may be unaffected. In absence of functional disturbances, such as amenorrhea or anovulation, no therapy is needed. A different approach is, however, required when the ovaries contain larger, hormone producing cysts which can persist for a long time. In contrast to micro-cystic ovaries the presence of persistent ovarian follicles can entail marked risks of endometrial hyperplasia, severe uterine bleeding, and anemia. PMID- 10522373 TI - [Glutamic acid decarboxylase autoantibody concentration and glucose tolerance in late pregnancy]. AB - Genetic and immunological factors may play a role as possible causes for gestational diabetes. Autoantibodies to glutamic acid decarboxylase (GADA) are frequently found in patients with insulin dependent diabetes, but have only rarely been analyzed with regard to the carbohydrate tolerance in pregnancy. An oral glucose tolerance test (oGTT) with 75 g glucose was performed in 110 pregnant patients during the third trimenon. Glucose (glucose dehydrogenase method) and insulin (RIA) concentrations were measured after 0, 30, 60, 120, and 180 minutes. Patients were divided into five groups of increasing glucose intolerance based on the highest glucose concentration reached during the oGTT. GADA were measured using a quantitative enzyme-immunoassay. Only a single patient showed pathologically elevated GADA, and her oGTT results were within the normal range. GADA in subjects with normal pathological glucose tolerance showed no significant difference (276.6 +/- 151.6 and 263.0 +/- 107.1 mU/ml respectively). There was a tendency of positive correlations between high GADA-levels and higher concentrations of insulin as well as an increased insulin-glucose-index. These findings suggest that pregnant patients with higher GADA-levels may have an increased insulin resistance. In conclusion, the concentration of GADA was not found to be helpful in evaluating the current metabolic situation in gestational diabetes. It remains unclear whether elevated GADA during pregnancy have a prognostic value regarding the manifestation of overt diabetes mellitus later in life. PMID- 10522374 TI - [Investigation of adhesion molecules (sVCAM-1) in serum of nonpregnant women, normotensive pregnant women and in patients with pregnancy complications]. AB - OBJECTIVE: An increased expression of endothelial adhesion molecules combined with neutrophil activation in the placental bed is to be assumed aetiopathogenetically relevant in preeclampsia. MATERIAL AND METHODS: Ranges of sVCAM-1 serum concentrations of both control persons (29 nonpregnant and 25 normotensive pregnant women) and patients with different complications of pregnancy (HELLP-syndrome n = 10, preeclampsia n = 12, gestational hypertension n = 38, diabetes n = 24, growth retardation n = 21) were determined by means of ELISA. Frozen placental samples of 5 normotensive and 10 hypertensive pregnant women were investigated immunhistochemically to study the distribution of VCAM-1 in the placenta. RESULTS: Significantly elevated sVCAM-1 serum levels (p < 0.05) were identified in samples of patients with HELLP syndrome, preeclampsia, diabetes and gestational hypertension compared with serum levels of normotensive pregnant women. The cut-off level (97.5% percentile of normotensive serum levels) was calculated (775 ng/ml). VCAM-1 was localized immunhistochemically at capillaries of villi and main villi. CONCLUSIONS: There are closed relations between elevated serum levels of sVCAM-1 during pregnancy and diseases with vasculopathies of placental bed. PMID- 10522375 TI - [Renal morphometric investigations in pre-eclampsia-like syndrome in a Wistar rat model]. AB - OBJECTIVE: Etiology and pathogenesis of renal changes in pregnancy-induced hypertension (PIH) remain somewhat controversial. Reducing the uterine perfusion leads to the development of systemic hypertension in animal models. Aim of this study was to investigate the effect of systemic hypertension on the kidney during pregnancy. MATERIAL AND METHODS: A rat model was used and the degree of the renal changes was quantified by morphometry. Normotensive pregnant and virgin animals served as a control. RESULTS: Hypertensive animals showed a decrease of the intravascular space in comparison to the cellular component. This difference was significant not only in pregnant (p = 0.0026) but also in virgin (p = 0.001) animals. CONCLUSIONS: These findings indicate that renal changes are usually found in normotensive pregnancies, while PIH strongly aggravates these changes. PMID- 10522376 TI - [Different techniques of reduction mammaplasty comparing clinical and esthetic complications with patient satisfaction]. AB - OBJECTIVE: The purpose of this study was to evaluate whether the use of one singleton and safe surgical procedure could be replaced by modified procedures adapted to the individual pathology without increase of complications rates. MATERIAL AND METHODS: 225 consecutive reduction mammaplasties were evaluated retrospectively, comparing the inferior central pedicle (n = 89) to the superior central pedicle (n = 12), the superior medial pedicle (n = 13), the central gland pedicle (n = 17) and the free areola nipple transplantation (n = 34). We checked for surgical and aesthetic complications of different techniques and satisfaction of the patients by questionnaire and personal interview. RESULTS: There was no difference in the incidence of different complications, satisfaction of the patients was comparable. The most frequent perioperative complication was hematoma formation, long-term problems were scar formations. CONCLUSIONS: As different techniques of reduction mammaplasties are mainly variations of surgical procedures it seems to be possible to adapt the procedure to the individual situation. With thorough indication and technique this approach does not increase complication rates. PMID- 10522377 TI - [Puerperal sepsis due to infected episiotomy wound]. AB - A healthy 31-year-old woman showed a severe septic shock syndrome a few days after vaginal delivery. In the episiotomy wound were found Group A Streptococci and E. coli. Although an antibiotic therapy was instituted immediately, the condition of the patient worsened. Platelet counts fell below 5000/microliter and she developed respiratory, cardiocirculatory and renal insufficiency, so that mechanical ventilation, high-dose-catecholamine therapy and continuous venovenous hemodiafiltration had to be performed. In the course of the disease the patient showed a reversible cardiomegaly with pulmonary hypertension and an extensive desquamation of the skin. Fever persisted in spite of the fact that in all following clinical and laboratory examinations no septic focus could be revealed any longer. She recovered slowly and could not be weaned from the respirator for four weeks because of a severe critical illness polyneuromyopathy. PMID- 10522378 TI - [Fertilization of cryopreserved and thawed human oocytes (Cryo-Oo) by injection of spermatozoa (ICSI)--medical management of sterility and case report of a twin pregnancy]. AB - It is reported on a twin pregnancy and parturition of a healthy girl and a healthy boy following the treatment of the mother by intracytoplasmic injection of spermatozoa (ICSI) into cryopreserved and thawed human oocytes (Cryo-Oo). The couple suffered from a primary sterility because of a severe subfertility of the husband and oligomenorrhea with anovulation of the wife. Before the couple contacted our center for the first time, several pretreatments had been carried out, e.g. intrauterine inseminations (IUI) with husband's and donor sperm and one cycle of in vitro fertilization (IVF) with intracytoplasmic sperm injection. This IVF/ICSI-cycle resulted in a tubal pregnancy. It must be noted that the patient showed in each treatment cycle a remarkable tendency to develop ovarian hyperstimulation syndromes (OHSS), even under stimulation with 50 mg clomiphen citrate. In 1997 we started the first treatment cycle for IVF/ICSI with GnIZH analogues and FSH according to the long protocol. In spite of a low dosage of gonadotropins the patient again developed a impressive multifollicular growth that we decided, together with the couple, only to perform the oocyte retrieval and cryopreserve the collected eggs. For ethic considerations the couple did not consent in the freezing of pronuclear stages. Two months later we performed the transfer-cycle: after thawing of four oocytes, the intracytoplasmic injection of native spermatozoa led to the fertilization of three oocytes and a total of three preimplantation embryos was transferred one day later. Two weeks later the blood level of hCG was 518 IU/ml and an intact twin pregnancy developed. In July 1998, after 36 weeks of gestation, a healthy girl (2540 g) and an healthy boy (2375 g) were delivered by cesarean section. This case report and the experience with further 23 patients (6 pregnancies) demonstrate that ICSI with cryopreserved and thawed oocytes is an effective approach to avoid repetitive oocyte retrievals. The achievable pregnancy rates so far seem to be similar to frozen pronuclear stages, possibly even better. Under the specific regulations of the German Embryo Protection Act (ESchG)--i.e. freezing of embryos not allowed--this technique is worth being pursued with attention. PMID- 10522379 TI - [Vaginal delivery after two previous cesarean sections]. AB - We report on a patient, who, after two previous caesarean sections, normally delivered vaginally without complications. The obstetric approach to, or better, the management of a vaginal delivery after caesarean section, our experience in other vaginal deliveries after previous caesarean section is discussed. This case report shows that, after two previous caesarean sections, the next child can normally be delivered vaginally without complications. PMID- 10522380 TI - [Hospital information systems or data processing in gynecology? Strategic reflections towards a comprehensive information concept]. AB - Hospital information systems are essential tools for efficient hospital management and improvement of medical procedures. Structural differences between the management of hospital information systems and the management of other huge, distributed and heterologous information systems cannot be found. However, there are differences concerning special tasks in patient care and medical research with high demands in data protection and modularity. The management of information processing has to be performed systematically. PMID- 10522381 TI - Biocontrol of Rhizoctonia solani and Pythium ultimum on Capsicum by Trichoderma koningii in potting medium. AB - Two isolates of Trichoderma koningii were evaluated for efficacy in control of damping-off diseases in seedlings of Capsicum annuum grown in pasteurized potting medium in a glasshouse. A selected isolate of binucleate Rhizoctonia and two fungicides were also included as standards for control of Rhizoctonia solani and Pythium ultimum var. sporangiiferum. Both isolates of T. koningii reduced seedling death caused by R. solani in one of two experiments, and by P. u. sporangii-ferum in two of three experiments. Neither isolate of T. koningii suppressed damping-off caused by either pathogen as consistently as the binucleate Rhizoctonia or fungicides. The implications of these results for commercial disease management are discussed. PMID- 10522382 TI - Detection of mycobacteria in clinical specimen using the mycobacteria growth indicator tube (MGIT) and the Lowenstein Jensen medium. AB - The recovery rates of mycobacteria strains isolated from 1200 clinical specimens using the mycobacteria growth indicator tube (MGIT) system and the conventional Lowenstein Jensen medium (LJ) were assessed. Of the 87 mycobacterial isolates recovered, 54 belonged to the M. tuberculosis complex (MTB) and 33 to the non tuberculosis complex (NTM). MGIT recovered 78 (89.65%) mycobacteria isolates (51 MTB (94.44%) and 27 NTM (81.81%) and LJ recovered 70 (80.46%) mycobacteria isolates (49 MTB (90.74%) and 21 NTM (63.63%). Sixty one (70.1%) of the total mycobacteria isolates were recovered with both systems (46 (85.2%) MTB and 15 (45.5%) NTM). No significant difference was found between MGIT and LJ (p > 0.05) in both MTB and NTM recoveries. The average detection time for MTB was significantly shorter with MGIT than with LJ, in both the smear-positive specimens (8 vs 30 days: p < 0.0001) and smear-negative specimens (15 vs 30 days: p < 0.001). The average detection time of NTM was also shorter for MGIT (15 vs 30 days: p < 0.0001). However, the contamination rate was higher in MGIT (8.5%) than in LJ (3%). The results suggest that the use of MGIT contributes to a more rapid and effective diagnosis of mycobacterial infections particularly when combined with the classical LJ. PMID- 10522383 TI - Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis and plasmid profile of soil and clinical isolates of Nocardia. AB - The aim of this study was to develop a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay for generic and species-specific differentiation of Nocardia from other morphologically similar bacterial pathogens. To examine the utility of the PCR-RFLP approach in species identification, genomic DNA was prepared from 40 soil isolates, 10 clinical isolates and 8 reference strains of Nocardia. A set of oligonucleotide primers was designed from the consensus sequence of the highly conserved groEL gene that encodes the 65-kDa heat shock protein (hsp 65). The primers selectively amplified 422 bp DNA from the genomic DNA of all Nocardia species and isolates. The digestion of the amplicons with the restriction enzyme MspI produced DNA fragments that could differentiate between different Nocardia species regardless of their origin. Additionally, the RFLP patterns obtained with restriction enzymes MspI and BsaHI resulted in the differentiation of six Nocardia species which were earlier identified by biochemical tests. Apart from soil isolates of N. asteroides, which had shown some degree of genotypic polymorphism with BsaHI, the remaining taxa yielded more consistent results. Our results on the isolation of plasmids indicated that their occurrence is not a consistent feature in Nocardia species. It is neither related to the source of origin (clinical versus saprobic), nor to virulence, anti-microbial resistance or species specificity. PMID- 10522384 TI - Phylogenetic analysis of Leptospira strains of pathogenic serovars using 23S rDNA gene sequences. AB - The 23S ribosomal DNAs were amplified from 11 strains of Leptospira interrogans sensu lato by polymerase chain reaction (PCR) and sequenced. The PCR products of about 290-bp DNA fragments indicated more than 97% sequence similarity to each other. The phylogenetic tree based on the 23S ribosomal DNAs obtained in this study revealed that 11 strains of L. interrogans examined composed a cluster distinct to that of L. weilii and L. borgpetersenii, confirming that these strains were similar to strain Moulton of L. interrogans serovar canicola in 23S rDNA sequence. PMID- 10522385 TI - The damage-inducible (din) genes of Escherichia coli are induced by various genotoxins in a different way. AB - The SOS response of Escherichia coli strains carrying the lacZ gene fused to the polB (dinA), dinB or dinD gene were investigated after treatment with several chemical agents and gamma-radiation. The induction levels of polB::lacZ reached levels between 4.0- and 9.0-fold 120 min after treatment with nalidixic acid, H2O2 or ethanol. Pentachlorophenol did not significantly induce any din genes. gamma-Irradiation is not an inducer of polB and ethanol failed to induce dinB::lacZ and dinD::lacZ. Following irradiation with a dose of 10 Gy the responses of dinB and dinD were induced about 2.5-3.0-fold above non-irradiated dinB and dinD. We found that the responses of din::lacZ fusion genes to these genotoxins are induced in a dose-dependent manner. The polB gene showed antagonistic responses to the simultaneous treatment of nalidixic acid and H2O2 or nalidixic acid and ethanol. In addition, dinB and dinD in the presence of both nalidixic acid and H2O2 at the same time showed no synergistic responses. PMID- 10522386 TI - Purification and amino acid sequence of lactocin 705, a bacteriocin produced by Lactobacillus casei CRL 705. AB - Lactobacillus casei CRL 705, isolated from a dry fermented sausage, produces an antibacterial peptide which is active against Listeria monocytogenes. Previous studies have shown that this compound is potentially useful to control food-borne pathogens in ground meat. In view of the potential application of this antimicrobial substance in food fermentation, a detailed biochemical analysis of this peptide is required. In this work, the purification and amino acid sequence of this bacteriocin is presented. The adsorption-desorption pH-dependent property of lactocin 705 was exploited for purification. The active extract was further subjected to RP-HPLC and SDS-PAGE. The active antimicrobial band was electroeluted from an SDS-PAGE gel and its amino acid sequence determined. Lactocin 705 had an estimated molecular weight of 3357.80 and an isoelectric point of 10.03. The peptide contains a high ratio of glycine residues and does not show any modified amino acids, like lanthionine or beta-methyllanthionine. The sequence was unique when compared to several databases. PMID- 10522387 TI - Stabilization of cellobiase by covalent coupling to soluble polysaccharide. AB - Cellobiase from Aspergillus niger was glycosylated by covalent coupling to cyanogen bromide activated dextran. The conjugated enzyme retained 62% of the original specific activity exhibited by the native cellobiase. The optimum pH as well as the pH stability of the conjugated form remain almost the same as for the native enzyme. Compared to the native enzyme, the conjugated form exhibited a higher optimal reaction temperature and energy of activation, a higher K(m) (Michaelis constant) and lower Vmax (maximal reaction rate), and improved thermal stability. The thermal deactivation of the native and conjugated cellobiase obeyed the first-order kinetics. The calculated half-life values of heat inactivation at 60, 70 and 80 degrees C was 10.7, 6.25, and 4.05 h, respectively, whereas at these temperatures the native enzyme was less stable (half-life of 3.5, 1.69, and 0.83 h, respectively). The deactivation rate constant at 80 degrees C for the conjugated cellobiase is about 7.9 x 10(-2) h-1, which is lower than that of the native enzyme (36.0 x 10(-2) h-1). The activation energy for denaturation of the native enzyme is about 10.58 kcal/mol, which is 7.25 kcal/mol lower than that of the conjugated enzyme. The effect of different surfactants and some metal ions on the activity of the conjugated cellobiase has been investigated. PMID- 10522388 TI - Proliferation, apoptosis and thymic involution. AB - The thymus reaches its maximal size at the age of 1 month in ICR mice and thereafter, the thymic cortex undergoes an exponential decline. This study was designed to compare the proliferation and apoptosis of thymocytes in different parts of the thymus of ICR female mice at the beginning and after the rapid phase of decline of the thymic cortical cellularity. The pattern of proliferation and apoptosis of the thymus was studied in situ in 1-month-old ICR female mice (10 mice) compared to mice at 7 months of age (10 mice). Staining for argyrophylic nucleolar organizer region by histochemistry was used to determine the proportion of type 2 thymocytes, which are considered as cells at S phase of the cell cycle. The mean number of type 2 cells in four random samples of 50 cells in each part of the thymus was defined as the proliferation index of this part of the thymus. In situ detection of apoptosis of thymocytes was carried out using the Apoptag kit, which can detect a single cell apoptosis. The mean number of apoptotic cells in five randomly selected fields of each part of the thymus was defined as the apoptotic index of this part of the thymus. The proliferation index of the peripheral cortex of the 1-month-old mice was 3.6 times higher than the proliferation index of the deep cortex and 5.8 times higher than the proliferation index of the medulla (P < 0.0001). The proliferation index of the peripheral cortex of the 7-month-old mice was reduced by 45% compared to the 1 month-old mice (P < 0.005). The apoptotic index of the corticomedullary junction of the 1-month-old mice was six times higher than the apoptotic index of the cortex and 18 times higher than the apoptotic index of the medulla. The apoptotic index of the thymic cortex was elevated by 66% in the 7-month-old mice compared to the 1-month-old mice (P < 0.0001). We conclude that there is a reduction of the proliferation index and an elevation of the apoptotic index of the thymic cortex in adult mice compared to young mice. These changes might account for the reduction of thymic cortical cellularity during thymic involution. PMID- 10522390 TI - Seasonal variations in serotonin immunoreactivity and ultrastructure in the pineal organ of the Japanese grass lizard, with special reference to environmental temperature. AB - The seasonal variations in serotonin immunoreactivity and ultrastructure of the secretory rudimentary photoreceptor cells (SRPC) were studied in the pineal organ of the Japanese grass lizard, Takydromus tachydromoides in relation to the environmental temperature. Our results clearly demonstrated that serotonin immunoreactivity in the lizard pineal organ displayed seasonal variations under an artificial photoperiod of LD 12:12 and natural temperature in the laboratory. Immunoreactivity became intense with increase in temperature from spring to summer, showing the strongest reaction in the summer, and subsequently became weak with the drop in temperature to winter. Also, the SRPC of the lizard showed distinct seasonal variations in number and size of the dense-cored vesicles correlated to the serotonin immunoreactivity. In contrast, the changes in size of the lysosomes and nucleoli of the SRPC were inversely proportional to that of the dense-cored vesicles. Furthermore, the lysosomes ingested some dense-cored vesicles after the autumn, and they coalesced to form huge autophagosomes or residual bodies during the winter. The present study provided serotonin immunohistochemical and ultrastructural evidence for seasonal variations in the secretory activity of the lizard pineal organ in accordance with changes in the environmental temperature. However, there may be few functional relationships between the pineal gland and the reproductive organs in the male Japanese lizard in relation to environmental temperature. PMID- 10522389 TI - Effects of IGF-I, IGF-II, bFGF and PDGF on the initiation of mRNA translation in C2C12 myoblasts and differentiating myoblasts. AB - In order to study the mechanisms by which growth factors stimulate protein synthesis, C2C12 myogenic cells were treated with a variety of growth factors and the recruitment of free ribosomes to polysomes was quantified. All experiments were conducted on C2C12 myoblasts (24 h prior to induction of fusion) and differentiating myoblasts (24 h after induction of fusion). After the 2 h incubation, cells were rinsed with phosphate buffered saline and quickly frozen at -80 degrees C. Cell lysates were fractionated on 15-60% sucrose gradients by centrifugation at 200,000 x g for 1 h. Absorbance at 254 nm was recorded continuously across the gradient. The response to each of the four growth factors, IGF-I and-II, basic fibroblast growth factor (FGF), and platelet-derived growth factor was a decrease (P < 0.05) in monosome peak height and a increase (P < 0.05) in polysome percentage (P < 0.05). All responses were linear, except IGF I, and the monosome peak height response to FGF which were quadratic (P < 0.05). None of the growth factors had a significant effect (P > 0.05) on RNA concentrations over the 2-h incubation. Protein content did not vary due to growth factor or level of treatment. This corroborates the hypothesis that the acute increase of protein synthesis exhibited by growth factor treated cells is due to an increase in the activity of existing ribosomes rather than an increase in ribosome synthesis. These results suggest that we can study the mechanisms regulating protein synthesis in muscle cells effectively by studying shifts in ribosomal activity. This method gave more consistent results than the H3-tyrosine incorporation and has the added benefit of not requiring the use of radioactivity. The strong correlation between monosome peak heights and percentage polysomes will allow researchers to measure total protein synthetic activity in a culture from the free or cytoplasmic fraction and to reserve the polysomes for other uses. The similarity of response among the various growth factors may indicate a common mechanism for increasing the initiation of protein synthesis. PMID- 10522391 TI - Effects of serum deprivation, insulin and dexamethasone on polysome percentages in C2C12 myoblasts and differentiating myoblasts. AB - An increase in the rate of protein synthesis in living cells can be achieved by regulating the quantity of mRNA, ribosomes, and enzymes available for translation or by regulating the efficiency at which existing components are used. Efficiency can be measured by comparing the number of ribosomes actively engaged in the synthesis of protein (polysomes) to the pool of free ribosomes. The objective of this study was to determine the percentage of ribosomes found as polysomes in C2C12 cells deprived of serum or exposed to insulin or dexamethasone 24 h before and after being stimulated to differentiate. Individual 60 mm culture dishes were exposed to serum-free control medium, medium containing serum, insulin, or dexamethasone for a period of 1 h or 2 h and then quickly frozen. The ribosomes and polysomes from these cells were separated by ultracentrifugation on 15 to 60% sucrose gradients and the absorbance across the gradient at 254 nm was recorded. Polysome percentages were determined as the area under the polysome peak divided by the total area under the curve. Serum deprivation caused a 12% decline in the percentage of ribosomes found as polysomes (P < 0.01). Dexamethasone caused a quadratic decline (P < 0.05) in polysome percentage, while insulin yielded a quadratic increase (P < 0.05). Protein synthesis assays measuring 3H-tyrosine uptake showed similar responses. These changes occurred in the absence of any differences in total RNA concentration. It was concluded that differentiation and the absence of serum in the media reduced the rate of recruitment of ribosomes for protein synthesis. Insulin increased ribosome recruitment which was also observed by a similar increase in incorporation of radio-labeled tyrosine. PMID- 10522392 TI - [In vitro evaluation of mercury leakage from dental amalgam using atomic absorption spectrophotometry]. AB - OBJECTIVE: The use of silver amalgam as a tooth filler is under constant critical review because of its mercury content. After a review of the literature on this subject, in vitro spectrophotometry was used to assay the release of mercury by these amalgams in basal conditions. METHODS: The experiment was conducted in two phases using standard doses of amalgam. In Phase 1 Black Class I cavities were created in extracted teeth that were big enough to take the required dose of amalgam. In some cases the material was inserted in a single operation, while in others the amalgam was introduced in three stages. Some of the cavities were cleansed with cotton buds bathed in ethyl alcohol, while others were not. All the drilled teeth were imbued in a bath of artificial saliva held at a constant temperature. In the second experiment, crushed amalgam was immersed in a similar bath of artificial saliva but otherwise untreated. The saliva was assayed after different time lapses using atomic absorption spectrophotometry and the FIAS technique. RESULTS: The results showed extremely variable but always modest quantities of free mercury in the artificial saliva. CONCLUSIONS: The findings suggest that not all the mercury available in the amalgamation phase is involved in the formation of the crystalline reticulum and that the percentage of mercury bonded is different every time. While the titre of free mercury encountered was always extremely low and hard to predict, it cannot be ignored. PMID- 10522393 TI - Radiographic evaluation of alveolar bone height in HIV-positive patients. AB - BACKGROUND: This prospective case-control study was performed to assess alveolar bone height in HIV-positive and HIV-negative patients. METHODS: Twenty-three HIV positive patients, 16 men and 7 women, aged 24 to 40 years (mean age: 33 years), consecutively referred to the Dental Clinic, University of Ferrara, for clinical and radiographic assessment of oral conditions, were included in the study (test group). All patients had undergone laboratory evaluation to assess HIV-infection status and were classified according to CDC diagnostic criteria. Nineteen patients were intravenous drug abusers. Thirty-four HIV-negative subjects were matched for demographic characteristics and smoking status as a control group. Radiographic evaluation was based on panoramic radiography and bone measurements were limited to premolars and molars. Alveolar bone height was measured mesially and distally to each tooth and determined as the distance from the apex of the root to a point where the lamina dura became continuous with the compact bone of the interdental septum. Alveolar bone height was recorded as well as the ratio between alveolar bone height and tooth length. RESULTS: The results indicated a tendency for a difference in alveolar bone height between groups, lower in the test group compared to controls. However, this difference only reached statistical significance on a tooth-specific basis. CONCLUSIONS: In conclusion, the results show a greater trend for alveolar bone loss of posterior teeth in HIV positive patients compared to HIV-negative patients. PMID- 10522394 TI - [Odontogenic kertocysts. Review of a series of cases and long-term clinical control]. AB - BACKGROUND AND AIMS: The aim of this retrospective study was to identify and evaluate the clinical characteristics and incidence of recidivation in a series of 40 cases of keratocyst consisting of 31 primary lesions, 7 cases of primary recidivation (17.5%) and 2 cases of secondary recidivation (5%). All the lesions were large sized and were treated using surgery. All patients underwent an annual clinical control for a minimum follow-up of seven years. METHODS: The authors present a series of 40 odontogenic keratocysts treated between 1985 and 1996 by the Division of Maxillofacial Surgery at Turin University. The series consisted of 40 keratocysts including 31 primary lesions, 7 cases of primary recidivation (17.5%) and 2 cases of secondary recidivation (5%); 12 patients were female (38.7%) and 19 were male (61.3%). The mean age of patients was 42 years old. The clinical records and enclosed X-ray documentation (OPT X-ray, head X-ray, face X ray) were examined for each patient, together with histological findings. Each case then underwent an annual follow-up. RESULTS: The review of clinical data and the examination of X-ray documentation showed that 28 lesions developed in the mandible and only 3 cases in the upper jaw. From a therapeutic point of view, keratocysts localised in a mandibular site were managed using cystectomy in 19 cases and in 18 cases this was followed by marsupialisation. Caldwell-Luc's operation was used in 2 cases of intrasinusal maxillary development, whereas the single case of extrasinusal development underwent cystectomy. Recidivation always involved the mandible and occurred in 22% of cases within 5 years of surgery. Of the 19 cases undergoing simple cystectomy, 8 cases (42%) revealed recidivation. Only one (5.5%) of 18 cases treated using cystectomy and marsupialisation showed recurrence. CONCLUSIONS: On the basis of their experience and in view of the specific characteristics of keratocysts, in particular the tendency to undergo recidivation, the authors affirm that annual clinical and radiological controls are indispensable and important, including biopsy where necessary, in order to diagnose new lesions promptly irrespective of the surgical technique used. PMID- 10522395 TI - [Diagnostic evaluation of abnormally erupted maxillary canines]. AB - Impaction of the maxillary canine is not a rare affection. A correct treatment can be appointed only after a serious evaluation of the case. In this work, the Authors describe clinical and radiographic methods which give the possibility to make a correct diagnostic evaluation of the maxillary canine impaction. A correct radiographic analysis could allow to three-dimensionally locate the impacted teeth and to show resorptions or ankylosis. Literature reports a great number of radiographic analysis; conventional methods can be used in simple cases, but, in difficult cases, some Authors suggest to use five different projections. New technologies in digital radiology allow to have high quality images, using which it is possible to make a diagnosis of great accuracy, also in difficult cases. Bidimensional images can show little particulars; three-dimensional images give the possibility to understand immediately the problems of the case also to not expert operators. The rate of radiation absorbed by the patient is similar to a panoramic radiograph. These characteristics make CT the method of election in difficult cases of maxillary canine impaction. PMID- 10522396 TI - [Necrotizing sialometaplasia. Report of a case]. AB - Necrotizing sialometaplasia is a benign lesion of the minor and major salivary glands. It occurs with a palatal swelling which rapidly becomes a deep ulcer. Histopathological aspects include ulceration of the surface epithelium, squamous metaplasia and acinar necrosis. In general no treatment is necessary and the lesion heals spontaneously in 4-10 weeks. This condition must be distinguished from more serious diseases such as squamous cell carcinoma and mucoepidermoid carcinoma. PMID- 10522397 TI - Stevens-Johnson syndrome. Description of an unusual clinical case due to glucocorticoid therapy for oral lichen planus. AB - Erythema multiforme (EM) is an acute inflammatory disease with an autoimmune pathogenesis clinically expressing in a wide variety of mucocutaneous illnesses. It is usually described in a minor form (Von Hebra) characterized by classical cutaneous lesions, and in major form (Stevens-Johnson), involving mucosal damage, while a clinical type restricted to the oral mucosa is described in oral pathology. A considerable number of factors of different nature have been reported as etiologic agents of EM, but most of them are not well documented; however, a certain relationship with EM is recognized for different classes of systemic drugs. This paper describes a case of Stevens-Johnson syndrome with initial oral involvement, in which the precipitating factor was due to the administration of systemic glucocorticoids, prescribed for the therapeutic treatment of an erosive form of oral lichen planus. PMID- 10522398 TI - [Von Recklinghausen osteitis fibrosa cystica. A clinical case]. AB - In Recklinghausen's disease the skeleton lesions are often the first signal of the pathology. The main clinical manifestations are represented by bony lesions which appear as multicystic lesions with loss of the hard lamina and skull malformation and asymmetry. In this disease there is a relevant osteoclastic activity which prevails over the osteoblastic one associated to the fibrous substitution of the marrow, sometimes producing micro or macro cysts. The typical alteration consists of an increase along the endosteal and trabecular surfaces in the number of osteoclasts which can be found in small reabsorption gaps. This is the cause of a cortical and trabecular reduction which can appear as interrupted. Histologic lesions consist in the replacement of bone tissue with fibrous and osteoid tissue. These bony lesions are not characteristic of the disease but to be distinguished from other pathologies such as for example Paget's disease and other forms of fibrous dysplasia (Gardner's syndrome, Leontiasis ossea). A case personally observed is described: a women, 29 years old, suffering from Recklinghausen's disease with face and skull asymmetry, condyles and glenoid cavity deformation, abnormal face reduction. Observing the planigraphy on the right side of the temporomandibular articulation, flattened glenoid cavity and condyles with irregular outlines can be noticed, aplastic coronoid cuts, altered jaw. The patient was submitted to surgery for dental extraction followed by a biopsy which showed some regressive alterations on cellular level of the bony structure. PMID- 10522399 TI - [Description of a severe and rare case of tinea barbae in the mental region]. AB - The authors report a case of tinea barbae which initially presented clinical and microscopic symptoms that led to the suspected diagnosis of a rapidly developing malignant tumour. A more detailed diagnosis and multi-specialist collaboration enabled the pathology to be correctly diagnosed and treated. PMID- 10522400 TI - [Telomerase in diseases of the digestive system]. AB - Synthesis of dezoxyrybonucleic acid at the ends of chromosomes by telomerase is the main process leading to indefinite cell proliferation. In contrary to majority of malignant tissues the telomerase activity is not detected in the most somatic cells. Detection of telomerase may be than an alternative to widely used diagnostic markers. It is proved that detection of telomerase activity is a useful method supporting cytology. Recent data confirm the correlation between the value of enzyme activity and progress of some cancers of digestive system. Therefore telomerase could play an important role in screening programs. Actually, the methods of detecting the enzyme activity are expensive and time consuming so their application in everyday clinical practice is limited. Simple immunohistochemical and immunoenzymatic methods are tested. New therapeutic procedures become available after describing the role of telomerase in carcinogenesis. The telomerase inhibitors are supposed to play an important role in the therapy of cancer. Such therapy could be efficacious in majority of cancers in which carcinogenesis depends on telomerase activation. Telomerase inhibitors are likely to be used in connection with surgery and followed by radio or/and chemotherapy but the potentials of antitelomerase therapy should be defined more accurately. PMID- 10522401 TI - [Echinococcus granulosus cysts in the liver. Ultrasonography findings during medical treatment]. AB - Since the sensitivity and specificity of serologic tests for echinococcosis is low, the role of imaging techniques is increasing. The goal of this study was to determine the usefulness of ultrasonography in diagnostics and classification of liver hydatid cysts and in the follow up after antiparasitic treatment. The study group of 35 persons, 26 women (mean age 42 years) and 9 men (mean age 39 years) was selected. The echography diagnosis was made using Toschiba scanner (SSH 140 A) with 3.75 and 7.5 MHz transducers. All 35 patients had positive results of serological tests for echinococcosis (EIA and/or indirect hemagglutination). The liver hydatid cysts (h.c.) are classified into seven types according to Caremani et all: I--simple h.c., II--multiple h.c., III--h.c. with detachment of the wall, IV--h.c. with mixed pattern with or without septations, V--heterogenous h.c., VI- hyperechoic h.c., VII--calcified h.c. The antiparasitic chemiotherapy with albendazole (10 mg/kg) was applied to 23 patients with total number of 59 h.c. of types I, II, III, IV, V. The observation points that cysts types I, II, III, IV, V respond to antiparasitic treatment. Our findings show that the sonographic patterns of cyst degeneration are seen with greater frequency in treated cysts than in nontreated ones. PMID- 10522402 TI - [The neuropsychiatric side effects of the interferon therapy in patients with chronic hepatitis B and C]. AB - Interferons are usage in therapy patients with chronic viral hepatitis (ch). Independently apart from therapy effects, lots of side effects are observed. The neuropsychiatrical side effects are most often and maybe hinder of interferon therapy. Among patients with ch B and C were estimated efficiency, type and frequency of side effects after interferon therapy. Especially symptoms disorders of central nervous system were observed between all side effects. The 67 patients: 20 with ch B, 45 with ch C and 2 with ch B retherapy by interferon were estimated. The lymphoblastoid interferon: alpha n1, Wellferon (Wellcome/Great Britain) and recombinant interferon alpha 2b, Roferon (Roche/Switzerland) and Intron-A (Schering-Plough/USA) were administrated. The efficiencies of treatment were observed after 6 month from finished interferon therapy. Elimination of polymerase HBV-DNA, antigens: HBs, HBe and antibody HBcIgM and normalization of aminotransferase activity in serum, were evaluated in patients with ch B. Elimination of HCV-RNA and normalization of aminotransferase activity in serum was evaluated in patients with ch C. Among neuropsychiatrical side effects was study. The frequently of irritability, fatigue neurosis, disorders of psychomotion activity, vegetative symptoms, somatic symptoms and disorder of higher mind processes. Grow excitability of sensorimotor, affective, irritability of vegetative system, were examined among irritability symptoms. Deficiency activity of polymerase HBV-DNA was observed in 40% patients with ch B. Deficiency of RNA-HCV in serum was observed in 31% patients with ch C. The flu-likely syndrome was observed at the beginning interferon therapy in 85% patients. Among patients treated with interferon were observed side effects: hair loss (29% patients), disorder of vision (29%), thrombocytopenia (< 50,000 mm3) (29%), leukopenia (< 2,000 mm3) (16%). The neurasthenia was detected in 60% patients with ch B and 50% patients with ch C. Neurasthenia most often manifested by irritable and quickly exhaustion of strength. 53% patients with ch B and 42% patients with ch C manifested disorders of psychomotion activity. Disorders of higher mind processes were in 50% patients with ch B and 33% with ch C. The neurological side effects were not influence on modification therapy. The flu likely syndrome was observed at the beginning interferon therapy in majority patients treat with interferon. The neuropsychiatrical disorders are frequently watching of interferon therapy in patients with ch B and C. PMID- 10522403 TI - [Lipoprotein and plasminogen serum levels in patients with diabetes mellitus type 2: risk factors for macroangiopathy? Preliminary study]. AB - Lipoprotein(a) [Lp(a)] is the specific lipoprotein which physiological role has not been explained. Similarity between apolipoprotein(a) and plasminogen structure suggests, that Lp(a) can be the bridge connecting the lipid metabolism and the coagulation system. The aim of the study was to evaluate if there is any correlation between Lp(a) and plasminogen serum concentrations in patients with diabetes t.2. 20 males and females with diabetes t.2 in the age 19-75 years (mean: 54.9 years). The control group consisted of 15 healthy men and woman in the age of 23 years (medical students). Lp(a) was estimated by ELISA, plasminogen by turbidimetric methods. Lp(a) serum level in the examined group was: 40.06 + 59.45 mg%. In 7 patients it was over 30 mg%. In the control group: Lp(a) concentration was: 13.23 + 10.5 mg%, no one was above 30 mg%. The different was significant. Plasminogen concentration was: in patients: 94.08 + 53.08%, in the control group: 89.08 + 40.7%. There was no significant difference. Correlation index between Lp(a) and plasminogen concentrations was in the patient's group: 0.3, in the control group: 0.2. In patients with diabetes we can find increased Lp(a) commentation in serum and normal plasminogen concentration. Concentration of Lp(a) didn't demonstrate significant correlation with plasminogen serum level. PMID- 10522404 TI - [The role of thoracoscopy in traumatic diaphragmatic rupture]. AB - The aim of this study was assess the role of thoracoscopy in the diagnosis of diaphragmatic rupture. Eight patients: six male and two female with blunt thoracic trauma underwent thoracoscopy. The chest X-ray and CT findings, pleural ultrasound indicated a diaphragmatic rupture only in three cases. Five of eight diaphragmatic injuries were confirmed by thoracoscopy. Authors suggests that thoracoscopy is a safe, accurate method for the diagnosis of diaphragmatic rupture. PMID- 10522405 TI - [The alcohol dependence in the patients of the Department of Orthopedics and the Department of Endocrinology of the clinical hospital in Bialystok]. AB - The authors analyse problem of alcohol dependence in the patients of the traumatic department (orthopedics) and the internal department (endocrinology). The evaluation of the dependence was made by the help of MAST. This test is easy to perform by the patients themselves. As authors predicted, the probability of alcohol dependence was higher among the patients of the orthopaedics department. Although the difference was not statistically significant, the authors confirm that the alcohol dependence may be related to physical traumas, especially in men. PMID- 10522406 TI - [Percutaneous drainage of recto-cecal abscess due to appendectomy]. AB - The authors presented 26 year old women suffered from lumbal and hypogastric pain, fever and recurrence diarrhea. She was performed appendectomy 3 years previously. From that time she has had abdominal pain, lack of appetite and moderate anemia. Based on ultrasound examination the diagnosis was established abscessus of retrocecalis. The diagnostic and therapeutic difficulties resulted from the situation of the abscessus were discussed. PMID- 10522407 TI - [Chronic hepatitis C in a 12-year-old girl coinfected with HGV]. AB - In this study we present a 12-year-old girl with chronic C hepatitis coinfected with HGV, in which severe life-threatening side effects from alfa interferon therapy occurred after 3 months of injection and required definite IFN withdrawal. It seems, that infection HGV may predispose patients with chronic C hepatitis treated with alpha interferon to this severe side effect. PMID- 10522408 TI - [Efficacy of low-dose albendazole therapy in hydatic liver disease: a case report]. AB - Hydatid liver disease is the most common form of human echinococcosis in Poland. Epidemiological data show a rising number of cases recognised last years. It is probably because of easier availability of ultrasound examination and computed tomography in the diagnostic procedures. While surgery remains the only approach for a radical cure, the chemotherapy with albendazole has became a useful advance in the management of cystic echinococcosis for the problem patient (inoperable disease, widely disseminated cysts, high surgical risk, etc.). The authors describe a patient with hydatid liver disease successfully treated with albendazole as a alternative treatment to surgery. A cyclic treatment of 28 days with 2 week break in between for 9 months with low doses of albendazole (6 mg/kg) resulted in clinical improvement and involution of hepatic cysts. PMID- 10522409 TI - [Surgical treatment of squamous cell carcinoma of the thoracic esophagus]. AB - The authors present modern approach of the squamous cell esophageal cancer's therapy. They emphasize the role of the surgical treatment with Akiyama method and the value of the preoperative chemotherapy and/or radiotherapy in the treatment of esophageal cancer. PMID- 10522410 TI - [Pharmacokinetic aspects of geriatric therapy]. AB - Some pharmacokinetic aspects of geriatric therapy were showed. Influence of alterations of physiological processes (changes of protein and lipid metabolism, failure of blood circulation and of renal function) in old age, on pharmacokinetics of drugs applied in geriatric pharmacotherapy (antibiotics, antiarrhythmic drugs, hypotensive drugs) were discussed. In this context, principles of rational pharmacotherapy of geriatric patients were proposed. PMID- 10522411 TI - [Vitamin D3]. AB - Pharmacokinetic and pharmacodynamic properties of vitamin D3 were described. Clinical aspects of application were also revealed. PMID- 10522412 TI - [The role of endorphins in the etiopathogenesis of circulatory diseases]. AB - At the beginning of the 70-ties various studies showed the existence of endogenous opiates in central nervous system. The chemical structure of first endogenous pentapeptides called enkephalins (met-enkephalin and leu-enkephalin) and their receptor was already known in 1975. Further investigations confirmed the existence of opiates (endorphins) in brain, pituitary gland, spinal cord and other tissues. Endorphins may exert various effects: analgesic (especially dynorphin and b-endorphin), antidiuretic, depressive on respiratory center, constipative, they also cause physical and mental dependence. These peptides, which are not known thoroughly may play a big part in the regulation of many biochemical and hemodynamic processes. Many of these mechanisms are already described and changes in concentrations of endorphins, for example in patients with silent myocardial ischaemia, are basic in the pathogenesis of this disease. Many of these mechanisms are controversial, the role of endogenous opiates in pathogenesis of hypertension or congestive heart failure is still unknown. This study summerizes the present knowledge endorphins and tries to answer whether changes in the concentration of endorphins are primary or secondary to the biochemical and hemodynamic disturbances in 2 diseases. PMID- 10522413 TI - [Do cytokines have any value in the patients with chronic blood circulation insufficiency?]. AB - Heart failure is a common and increasing public problem. Neurohormonal activation plays a role in the pathophysiology of heart failure, but is probably also affected by cytokines. We studied 75 patients with heart failure NYHA functional class II and III-IV, who were treated with angiotensin converting enzyme inhibitor (enarenal), diuretics (furosemide) and digoxine. Their mean age was 63.9 years/range 65-86/, left ventricular ejection fraction in the patients NYHA functional class II and III-IV classes was 68.9% and 47.3% respectively; 12 were females. Significant improvements in NYHA classification were shown. The levels plasma TNF-alpha (tumor necrosis factor-alpha) and interleukin-6 (IL-6) were analysed before and after therapy. The authors showed increased plasma levels TNF alpha and IL-6 in patients with chronic heart failure. After the treatment the plasma IL-6 levels decreased only in the patients III-IV NYHA functional classes, whereas the treatment had no effect on the plasma TNF-alpha levels. PMID- 10522414 TI - [The influence of digoxin treatment on the statistical distribution of R-R intervals in atrial fibrillation]. AB - Functional refraction of atrio-ventricular node and the phenomenon of concealed conduction are basic factors which determine the frequency of ventricular beats as well as the duration of R-R intervals in atrial fibrillation. R-R intervals may be ordered according to their size and presented graphically in the form of histograms. The morphology of histograms depends on the ability of A-V node. Digoxin influences the morphology of histograms of R-R intervals, which may have a diagnostic value and become a therapeutic indicator. The purpose of the study was to evaluate the correlation between the morphology of histograms and digoxin serum concentration, as well as the clinical treatment with digoxin. In the study a classification was used which divided R-R intervals into 4 types and several sub-types. The study covers 91 patients treated with digoxin upon whom 161 histograms were made. Two groups of patients were studied: A-patients in whom digoxin concentration was determined, B-patients in whom treatment with digoxin was evaluated on the basis of their clinical condition. The study confirmed a hypothesis that treatment with digoxin causes specific changes in the statistical distribution of R-R intervals even though no strict correlation was found between the direction of these changes and the digoxin serum concentration. The dependence, however, was observed between histograms of R-R intervals in atrial fibrillation and the clinical course of treatment with digoxin. PMID- 10522415 TI - [Influence of treatment with metoprolol or enalapril on recovery of contractile function of the left ventricle in patients after acute myocardial infarction treated by thrombolytics]. AB - Regional left ventricular contractility caused by myocardial stunning as a result of transient ischemia and postreperfusion injury is a reversible state it can however persist even for several month. It seems reasonable to shorten this period as much as possible. The aim of the study was to estimate the influence of treatment with metoprolol or enalapril on the recovery of contractile function of left ventricle in patients after acute myocardial infarction treated thrombolytically. Investigations were carried out in 127 patients (mean age 62.3 +/- 11.9 years). Metoprolol was used in 37 patients in dose 0.02-0.05 g b.i.d., enalapril in 48 patients 0.0025-0.01 g b.i.d. 42 patients were not treated with any beta-blocker or ACE inhibitor. In all patients echocardiographic study was performed 3 times: on 2-3rd day following acute myocardial infarction immediately before introducing the treatment with metoprolol or enalapril, after 1 month and after 3 months. Echocardiographic study wall motion index (WMI) was calculated basing on. Significant decrease in WMI was observed after 1 month compared to its value on 2-3rd day acute myocardial infarction and after 3 months compared to 1 month after myocardial infarction in each of 3 subgroups of patients. No statistically significant differences in WMI were found out between studied subgroups. Neither metoprolol nor enalapril started on 2-3rd after thrombolytic treatment of acute myocardial infarction do not affect the recovery of contractile function of stunned myocardium. PMID- 10522416 TI - [The effect of intravenous magnesium on the arrhythmias in patients after electrical cardioversion]. AB - The aim of this study was to examine the effect of magnesium, applied by intravenous infusion, on supraventricular and ventricular cardiac arrhythmias in patients after effective electric cardioversion. 30 patients examined were: 4 women and 26 men aged from 24 to 84 years (the average age being 56.1). All patients had previously been treated successfully by electric cardioversion. Patients were divided into two groups: group 1 of 15 patients aged 24 through 74 years, group 2 of 15 patients aged 25 through 76 years. After effective cardioversion, in the first group of patients, the infusion of 500 ml 0.9% NaCl was applied; whereas the other group of patients received 500 ml 0.9% NaCl with 5g MgSO4 and 20 mEq KCl. After electric cardioversion all patients had 24-hour Holter monitoring. After successful electric cardioversion in all the examined patients, supraventricular and ventricular cardiac arrhythmias were observed. However, in group 2 numerous premature supraventricular beats, episodes of atrial fibrillation, numerous premature ventricular beats, multifocal premature ventricular beats and pairs of ventricular beats were rarer than in the 1st group. In conclusion, it should be stated that the intravenous application of magnesium can be an effective method of treating supraventricular et ventricular cardiac arrhythmias occurring with patients after electric cardioversion. PMID- 10522417 TI - [The level of plasma endothelin-1 in patients with essential hypertension]. AB - Endothelin-1 (ET-1) as a potent vasoconstrictor, has mitogenic and inotropic properties, stimulates the renin-angiotensin-aldosterone system and sympathetic nervous system. The results of recent studies suggest that overall hemodynamic effects of ET-1 may play a part in the control of blood pressure and the pathophysiology of hypertension. Several investigators have invoked increase level of ET-1 in human essential hypertension but the results of studies concerning hypertensive patients with normal renal function have shown that they have similar concentrations of ET-1 to those in normotensives. To establish whether ET-1 may elevate of blood pressure, the value of plasma ET-1 activity was determined in peripheral blood in 101 patients with essential hypertension. There was no significant difference between in mean level of ET-1 in patients with essential hypertension and in control group. The plasma ET 1 was significantly higher in patients with severe hypertension than that of patients with mild and moderate hypertension, and in patients with severe hypertension there was a correlation between the level of ET-1 and microalbuminuria. There was no correlation between the plasma ET-1 level and systolic blood pressure and diastolic blood pressure in the patients with essential hypertension as a whole, not was there any correlation between ET-1 and noradrenaline, aldosterone level and renin plasma activity. PMID- 10522418 TI - [The study of the activity of creatine kinase in diagnosis of coronary reperfusion in patients with acute myocardial infarction after thrombolysis]. AB - Early estimation of the efficacy of thrombolysis in acute myocardial infarction is of great clinical importance because the appearance of coronary reperfusion changes therapeutic and diagnostic procedures and decreases the mortality rate. Previous studies showed that the analysis of activity of creatinine kinase (CPK) measured in regular, short periods of time after thrombolysis night be useful in the diagnosis of reperfusion equally to coronary angiography. The aim of the study was to estimate the usefulness of the analysis of creatine kinase (CPK) and its isoenzyme (CK-MB) in the diagnosis of coronary reperfusion in patients with acute myocardial infarction after thrombolytic therapy. The study was performed in 50 patients with acute myocardial infarction admitted to our Cardiology Department, of these 42 were men aged from 34 to 68 and 8 were women aged from 43 to 70. 28 patients had acute inferior myocardial infarction, 22 patients--acute anterior myocardial infarction. All patients were administered 300 mg of aspirin after admission and then 150 mg of aspirin daily and 1,500,000 IU of streptokinase i.v. within 1 hour. Venous blood samples for determination of CPK and CK-MB were obtained every 3 hours during the first 48 h and once a day at 8 a.m from 3rd to 11th day. All patients underwent coronary angiography 2-4 weeks after thrombolysis. The study showed that in patients with reperfusion, activities of CPK and CK-MB three hours after thrombolysis were higher than 30% of later peak. These findings show the usefulness of this criterion in early, non invasive estimation of efficacy of thrombolysis. Determination of activity of isoenzyme CK-MB during thrombolytic therapy is not necessary, because it evaluates similarly to CPK. We showed that electrocardiographic and enzymatic criteria are comparable in estimation of efficacy of thrombolytic therapy. PMID- 10522419 TI - [The significance of the Doppler techniques -- "color Doppler" and "power Doppler" -- in the diagnostic ultrasonography of the eyeball]. AB - The eyeball is perfectly suited for ultrasound examination due to its location and structure. The ultrasound examination including the "color Doppler" and "power Doppler" option was performed in 59 patients. Ophthalmologists requested ultrasound examination in those patients mainly in order to confirm and differentiate retinal ablation or because of opacity of vitreous body of the eye. The ultrasound examination was performed with a Logiq 500 f-y GE unit equipped with a 7.5 MHz linear transducer and a 7.5 MHz linear transducer and a Hitachi 415 EUB applied. Primary retinal ablation was diagnosed in 27 cases, and in 14 cases retinal ablation was caused by a proliferative process. In 18 cases a different pathology was demonstrated including 6 patients in whom we detected an intraocular foreign body. PMID- 10522420 TI - [The evaluation of the bioavailability of isosorbide 5-mononitrate]. AB - The aim of this study was to investigate the bioavailability of isosorbide mononitrate (IS-5MN) after oral administration of Monocard 20 mg-capsules, made in "Synteza" a Pharmaceutical-Chemical Company in Poznan. Effox 20 mg, coated tablets from Schwarz Pharma, was used as an counterpart of Monocard 20 mg. The concentrations of IS-5MN in healthy volunteers' plasma were determined by using Hewlett Packard gas chromatography. CONCLUSIONS: The bioavailability of IS-5MN after oral administration of Monocard 20 mg is the same as after oral administration of Effox 20 mg, whose clinical efficacy was tested before. This conclusion confirms the same value of AUC, tmax, cmax and other pharmacokinetic parameters of Monocard 20 mg and Effox 20 mg. PMID- 10522421 TI - [Coronary artery bypass grafting in patients with left main coronary artery occlusion]. AB - The paper describes 3 men aged 43, 62 and 66 years with left main coronary artery occlusion. In all patients well-developed collateral circulation from right to left coronary artery was present. They were operated in cardiopulmonary bypass and two bypass grafts were implanted in each of them. There were no deaths in the perioperative period. Actually all the patients are alive and have no angina. PMID- 10522423 TI - [Stress echocardiography: methods, indications, clinical application]. AB - Stress echocardiography is an accepted alternative method for non-invasive assessment of coronary artery disease--diagnosis, risk stratification and prognosis. Myocardial ischaemia triggers a cascade of events resulting first in regional relaxation abnormalities (or diastolic dysfunction) followed by regional motion abnormalities (or diastolic dysfunction). The basic principle in stress echocardiography is to provoke myocardial ischemia by exercise, pharmacologic interventions (like dobutamine, dipirydamole, enoximone, adenosine) or less often atrial pacing and subsequently to evaluate regional wall motion abnormalities in segments vascularized by coronaries with flow--limiting stenosis. Myocardial viability can be identified at the low dose dobutamine or dipirydamole stage as a functional improvement in regions with rest dyssynergy and myocardial ischaemia can be recognized at high doses as well motion dysfunction. The ideal test for evaluation of the patient with CAD remains exercise stress testing, pharmacologic stress should be reserved only for those patients in whom optimal workload of stress cannot be obtained. This article reviews the current status of stress echocardiography in clinical practice and assesses the possible indications for the tests in a modern cardiac department. PMID- 10522422 TI - [Drug-related hyperkalemia resulted from spironolactone and angiotensin converting enzyme inhibitors therapy]. AB - A case of drug-related hyperkaliemia linked to treatment with angiotensin converting enzyme inhibitors and spironolactone simultaneously. The paper presents the case of drug-related hiperkaliemia induced by captopril and spironolactone combined treatment in a patient with early stage of renal failure. PMID- 10522424 TI - [Contrast agents in echocardiography]. AB - Contrast echocardiography is new imaging modality dynamically entering to clinical practice. The efficient application of the contrast agents needs some knowledge of microbubbles physics. In this paper the basic principles of echo contrast effects are presented and ultrasound microbubble fluid dynamics in the ultrasonic pressure field is addressed briefly. The recent developments in harmonic imaging end the transient and non-linear acoustic phenomena allow ultrasound to detect of blood flow in the microcirculation. In the second part of the paper it was demonstrated a potential for myocardial opacification by contrast echocardiography and the more recent application such as myocardial perfusion. The authors also reported intracardiac shunts imaging, enhancing endocardial border definition and bringing out of weak Doppler signals. The review of new contrast agents coming to the clinical practice is presented. PMID- 10522425 TI - [Hematologic disorders in patients with cyanotic congenital heart disease]. AB - Erythrocytosis, thrombocytopenia, platelets function defects, coagulation factors deficiencies are the main haematologic disorders in patients with cyanotic congenital heart disease. They are responsible for increased bleeding tendency and contraindication of anticoagulation. Phlebotomy in this group of patients should be recommended when erythrocytosis is accompanied by clinical symptoms. Excessive phlebotomy can lead to iron deficiency and cause hyperviscosity symptoms. The iron supplementation should be closely monitored. PMID- 10522426 TI - [Lipodystrophy and antiretroviral agents]. PMID- 10522427 TI - [Heart transplantation at the end of the 20th century]. PMID- 10522428 TI - [Infectious complications of heart transplantation. A prospective study for the first 6 years of a transplantation program]. AB - OBJECTIVE: To study the infectious complications, mortality, and associated factors in heart transplant recipients. METHODS: Prospective study of the first 69 heart transplantations performed from January 1991 until December 1996 in a university hospital. Description of clinical features of infectious complications during the first year after transplantation. Univariate and multivariate analyses of the risk factors associated with mortality and development of infectious complications. RESULTS: Seventy-three percent of patients had at least one infectious complication; the incidence was 1.13 episodes per patient-year. The etiology of complications was bacterial (50%), viral (31%), Pneumocystis carinii (5%), fungal (4%), and protozoal (4%). The opportunist organisms accounted for 42% of cases. Pneumonia was the most common complication (28%), followed by mucocutaneous herpetic reactivation (19%), bacteremia (13%), urinary tract infection (13%), cytomegalovirus disease (11.5%), pleural empyema (5%) and surgical wound infection (5%). Nosocomial pneumonia accounted for 50% of cases. Gram-negative rods accounted for 41% of pneumonia cases. A total of 62.5% of deaths were directly related to infectious complications. Factors independently associated with mortality were hospital origin at transplantation (RR = 4.5 [2 9], p = 0.034), development of infectious complications in the post-heart transplantation period (RR = 3.2 [1.2-12], p = 0.04) and a more prolonged hospital stay at ICU (p = 0.0004). The factor which was independently associated with the development of infectious complications was one or more severe episodes of acute rejection (RR = 1.5 [1.1-2.2], p = 0.04). Patients who developed infectious complications had a more prolonged accumulated annual hospital stay (p = 0.004) than those without infectious complications. CONCLUSIONS: Infectious complications are very common, prolong hospital stay, and are the first cause of mortality during the first year after transplantation. Bacteria are the most common etiology and pneumonia is the most common infection. PMID- 10522429 TI - [Clinico-radiological correlation of pulmonary complications of pediatric HIV infection]. AB - Fifty-five percent of children with HIV infection, aged two months to ten years, were admitted at our hospitals because of respiratory conditions. Pulmonary complications found at admission in these children were lymphoid interstitial pneumonitis, Pneumocystis carinii pneumonia, fungal over-infection, tuberculosis, and bacterial complications. Also, non-specific infectious bronchial conditions, probably of viral origin. The most representative chest-X rays of these pulmonary conditions were analyzed; together with data from clinical records a clinico radiological diagnosis was obtained. PMID- 10522430 TI - [Bacteremias in a university hospital: study of etiologic agents and their sensitivity patterns]. AB - OBJECTIVE: To determine the bacterial etiology of bacteremic episodes recorded at our hospital during 1995 and their antimicrobial susceptibility patterns. METHODS: The microbiological records of all bacteremic episodes detected at our hospital from January to December 1995 were analysed. The susceptibility patterns of the 334 gram-positive aerobic isolates to 11 antimicrobials and of 236 gram negative aerobic isolates to 16 antimicrobial agents were determined. The reference agar dilution method was used for these determinations. RESULTS: The incidence of bacteremia was 19.3/1,000 admissions. Gram-positive aerobic bacteria accounted for 56.6% of monomicrobial bacteremias; the microorganisms recovered most frequently were coagulase-negative staphylococci (22.4%), Escherichia coli (16.5%) and Staphylococcus aureus (14.2%); 75 polymicrobial episodes were recorded. Over half of bacteremic episodes occurred at medical services. Hematologic diseases and solid tumours were the most common underlying diseases. No resistance to glycopeptides was observed among the staphylococci studied. The incidence of resistance to vancomycin in enterococci was small (1.5%). The aminoglycosides tested and some beta lactams showed good activity against the gram-negative bacilli studied. CONCLUSIONS: To carry out an epidemiologic surveillance of bacteremic episodes occurring at every hospital it is necessary to provide information on trends observed in the etiology of such infections, possible outbreaks, antimicrobial resistance, and uncommon pathogens. PMID- 10522431 TI - [Persistence of hypercoagulability state after hip and knee arthroplasty: what is the optimal duration of antithrombotic guidelines in this surgery?]. AB - Among 79 patients candidates to hip (53) and knee (26) replacement an evaluation was made of the influence of a two-week program with LMWH on the evolution of hypercoagulability markers: D-D, TAT, and F1 + 2. Measurements were performed by ELISA preoperatively and on days 1, 7 and about 45 postoperatively; in the latter, two extraction intervals were considered: < or = 45 days and > 45 days. With both surgical modalities, D-D and F1 + 2 peaked at 7th day postoperatively, whereas TAT peaked on day 1. Among D-D and F1 + 2 values quantitated on day 7th and the extraction interval < or = 45 days, no significant differences were obtained (Z < 2.64). The hypercoagulative chronicity exhibited by D-D and F1 + 2 during the first month and a half after this surgery, might require in some cases a more prolonged thromboprophylaxis. PMID- 10522432 TI - [Liver abscess caused by Klebsiella pneumoniae in diabetic patients]. AB - Pyogenic liver abscess are macroscopic collections of pus within the hepatic parenchyma after a bacterial infection. These infections are usually polymicrobial in nature, and in most occasions due to biliary tract diseases or cryptogenetic in origin. Monomicrobial hepatic abscess caused by Klebsiella pneumoniae are uncommon lesions in western countries. These lesions are associated with underlying diseases, particularly diabetes mellitus, and are frequently complicated with septic metastasis. We report here three cases of monomicrobial liver abscess caused by Klebsiella pneumoniae in diabetic patients, without septic metastasis and a favourable outcome. PMID- 10522433 TI - [Langerhans cell histiocytosis in the central nervous system: meningeal involvement]. AB - The clinical manifestations of Langerhans cell histiocytosis in the central nervous system include not only the classical involvement of the hypothalamus hypophyseal axis, but also of other anatomic locations, with the corresponding variability of symptoms. We report here two patients with meningeal lesions. In both patients, the onset of the disease was systemic, and neurological symptoms developed some years later. Imaging studies, particularly magnetic resonance, are very sensitive tools for the diagnosis and follow-up of these lesions. The natural history of this disease is still poorly understood. In this paper we related the clinical manifestations to the morphological changes observed in imaging studies. PMID- 10522434 TI - [Relationship between essential arterial hypertension and osteoporosis:impact of antihypertensive treatment on extracellular metabolism of calcium]. PMID- 10522435 TI - [Physicians and Justice (I): aid to the Justice System]. PMID- 10522436 TI - [A 36-year-old woman with HIV infection and abdominal distension]. PMID- 10522437 TI - [Asymptomatic pulmonary mass]. PMID- 10522438 TI - [A 62-year-old man with oligosymptomatic pulmonary mass]. PMID- 10522439 TI - [Prolonged fever and liver tumor]. PMID- 10522440 TI - [Current treatment of cystic fibrosis]. PMID- 10522441 TI - [Peripheral lipodystrophy associated with protease inhibitor therapy]. PMID- 10522442 TI - [Impact of protease inhibitors on hospitalization in an Infectious Disease Unit]. PMID- 10522443 TI - [Prevention of malaria]. PMID- 10522444 TI - [Total hip and knee arthroplasty: do obese patients or smokers require a more prolonged thrombosis prophylaxis?]. PMID- 10522445 TI - [Monocytosis and disseminated intravascular coagulation as clinical manifestations in a case of hepatosplenic tuberculosis]. PMID- 10522446 TI - [Spontaneous rupture of the spleen: a rare form of onset of non-Hodgkin's lymphoma]. PMID- 10522447 TI - [Harmonic imaging--what is it]. PMID- 10522448 TI - [Power Doppler and contrast-enhanced duplex ultrasound: what is practically relevant at the time?]. PMID- 10522449 TI - [New technical aspects of B-image ultrasound]. PMID- 10522450 TI - [Emergency ultrasound diagnosis of the musculoskeletal system]. PMID- 10522451 TI - [Pitfalls in ultrasound diagnosis of acute abdominal trauma]. PMID- 10522452 TI - [Possible applications of the newest "hig end" ultrasound equipment in pediatrics]. PMID- 10522453 TI - [Color-coded duplex ultrasound in diagnosis of acute deep venous thrombosis of the leg: how reliable is the diagnosis in routine practice?]. PMID- 10522454 TI - [Ultrasound diagnosis of acute appendicitis--really an additional source of information?]. PMID- 10522455 TI - [Angiosarcoma of the spleen]. PMID- 10522456 TI - [What is your diagnosis? Carotid stenosis and ulcerated atherosclerotic plaque induced the formation of a thrombus resulting in a cerebrovascular stroke complicated by hemiplegia of the right side and motor aphasia]. PMID- 10522457 TI - [Incidence of benign prostatic hyperplasia and hepatic cirrhosis]. AB - We studied the incidence of benign hyperplasia of the prostate and hepatic cirrhosis in an ambulatory cohort. Undisturbed androgen production is one of the prerequisites for the development of benign prostatic hyperplasia. The disturbed metabolic activity of the cirrhotic liver causes often hypoandrogenism. Therefore a decreased incidence of prostatic hyperplasia is expected for patients with cirrhosis. All male patients of european origin over 40 newly admitted for treatment at the medical outpatient clinic in Zurich during a period of 6 months were enrolled. From a cohort of 2038 males over 40 a benign prostatic hyperplasia was diagnosed in 124. Nine had cirrhosis of the liver. In 6 patients cirrhosis was found in addition to benign prostatic hyperplasia. The study also confirmed an increased incidence of benign prostatic hyperplasia with age. The incidence of cirrhosis, however, decreased with age in our study, probably as a consequence of reduced life expectancy of the cirrhotic patient. The combined incidence of benign prostatic hyperplasia and hepatic cirrhosis became less common with age than benign prostatic hyperplasia alone. This corresponded to our expectations. This collection of epidemiologic data on benign prostatic hyperplasia and hepatic cirrhosis disclosed several difficulties that should be considered for future studies. PMID- 10522458 TI - [Clinical efficacy and tolerance of the hypericum special extract LI 160 in depressive disorders--a drug monitoring study]. AB - PATIENTS AND METHODS: 647 patients suffering from mild to moderate depression treated for 6 weeks with hypericum extract LI 160 (Jarsin 300), 1 tablet t.i.d. RESULTS: The condition of the patients improved in 75% (primary endpoint). The von Zerssen depression score decreased from 19.8-21.2 (95%-CI) at base-line to 8.1-9.3 at week 6 (p < 0.001). All symptoms were improved at week 3 and further at week 6. The condition improved somewhat slower in patients older than 65 years. The severity of the depression did not appear to have an effect on the outcome. Adverse events were reported by 17% of the patients (gastrointestinal 10%). These were mostly mild or moderate. Tolerance was rated in 89% as satisfactory or better. PMID- 10522459 TI - [Neuro-axial regional anesthesia: rational choice between spinal and epidural anesthesia]. PMID- 10522460 TI - [Headache, painful eyes, fever and weight loss]. AB - A 64-year-old patient with herpetic keratouveitis was hospitalized because of fatigue, fever, headache and confusion. Three days before admittance keratouveitis was diagnosed. He reported a recent onset of aversion against meat consumption and weight loss of 11 kg over the last 4 months. Clinical investigation revealed a slightly confused patient with conjunctivitis and reduced vision of the left eye. Laboratory tests showed anemia, hyponatremia, and increased carcinoembryonic antigen (CEA). In the cerebrospinal fluid examination protein concentration was increased, glucose concentration was decreased. CT-scan of the brain revealed multiple, hyperintense, circular lesions. Biopsy showed lymphoplasmacellular infiltration with increased number of glial and oligodendroglial cells with central necrosis. Despite therapy with tuberculostatic and antiviral drugs and corticosteroids the condition of the patient progressively deteriorated. The patient died 42 days after admission. Autopsy revealed a high grade B-cell non-Hodgkin's lymphoma of the jejunum. Septic shock was the cause of death with the lymphoma of the jejunum as a possible nidus of infection. The multiple brain lesions with central necrosis were probably caused by thromboembolization or by a previous viral meningoencephalitis. PMID- 10522461 TI - [Delay of the diagnosis of an amelanotic malignant melanoma due to CO2-laser treatment--case report and discussion]. AB - The incidence of malignant melanoma is steadily increasing. The best prognostic indicator of melanoma is the vertical tumor thickness. Early recognition of melanoma by the naked eye is often difficult, because many lesions simulate its clinical presentation. We report on a 47-year-old man with a red papular skin lesion treated by CO2 laser. The histological diagnosis of an amelanotic malignant melanoma was thus delayed for one year. Clinically ambiguous skin tumors should never be treated by laser prior to a diagnostic biopsy. PMID- 10522462 TI - [Insufficient immune response to hepatitis B-vaccination--causes? Patient: a 52 year-old housewife]. PMID- 10522463 TI - [Dyslipidemia and personality--an investigation of 376 aged athletes]. AB - There exists much evidence for a close connection between "Type A" personality and a high cardiovascular risk. In addition, there seems to be a strong association between high lipids and personality as well. In this investigation we addressed the question of this relationship. METHODS AND RESULTS: In a sample of n = 376 male life-time athletes, born 1910 to 1940 a retrospective longitudinal study aiming at motor development and influencing factors was conducted including for instance data concerning health status, personality, locus-of-control. In the sample (medium age of 65 years) the prevalence for hypercholesterin /triglyceridemia equals 35%. The objective of the present study was the identification of sub-samples with significantly higher prevalences defining personality features, locus-of-control and closely related constructs as independent variables. The data analysis was performed using CHAID. The sample is separated into 14 subgroups by the combination of 10 predictors. Above average prevalences were found in people with the key personality features of "tension", "neuroticism", "aggressiveness", and "extraversion" in several combinations. If a low personal-best or many statements "sport becomes less important" gain relevance, the prevalence rises to 100%. A trustworthy occupation and "neuroticism" predict a percentage of 83%. "Aggression" and "extraversion" are related to an unhealthy, "worrying about ones health" to a healthy lifestyle. The latter as well as low "life-satisfaction" and feeling "dependent upon others" correspond to compensatory sport engagement. Many results therefore support findings concerning the protective power of sport and the negative impact of tension, neuroticism, and inactivity. Our results showed a close association between features of "Type A" personality and high lipids in older life-time athletes. It is only possible to speculate about causality. A plausible idea is to consider a common patho-physiological basis of changes in metabolism and personality (e.g. sympathic activity or "drive"). In any case, the above mentioned personality features moderate unhealthy behavior patterns leading to hypercholesterin-/triglyceridemia. PMID- 10522465 TI - [Modern instrument-assisted gait analysis]. PMID- 10522464 TI - [Some aspects of tinnitus]. AB - Tinnitus is the sensation of sound, a sensation generated by the auditory system because of a pathology, without any external acoustic or electrical stimulation. Most often, it is associated with a sensorineural hearing loss. Tinnitus is still one of the most frequent symptoms encountered by the otorhinolaryngologist and other doctors. Diagnosis and therapy are demanding due to complex etiology and secondary symptoms. Tinnitus is a symptom and not a disease. Therefore a thorough diagnosis is necessary. First of all one has to evaluate whether there is a treatable underlying organic disease possibly responsible for symptoms like tinnitus. The evaluation of the patient includes the history, ENT-status, audiological and vestibular findings, investigative imaging and examinations by other specialists. The therapeutic aim is the compensation of tinnitus. There is no universal medical or surgical treatment. Acute tinnitus is treated like sudden deafness. For chronic forms, the analysis of the causes is particularly important for developing an individual consultation and therapy plan. Providing information to the patient is the first step for a sensible treatment of the symptoms. Supportive therapy includes a psychosomatic therapy and the use of medication or instrumentation. PMID- 10522466 TI - [Round lesion. Churg-Strauss Syndrome (CSS)]. PMID- 10522467 TI - [Fever with immunodeficiency. Hodgkins lymphoma with Reed-Sternberg cells]. PMID- 10522468 TI - Need in the midst of plenty. PMID- 10522469 TI - Risperidone in the treatment of choreiform movements and aggressiveness in a child with "PANDAS". AB - The acronym 'PANDAS' (pediatric autoimmune neuropsychiatric disorders associated with streptococcal infections) is used to describe neuropsychiatric symptoms putatively resulting from autoimmune responses to streptococcal infection in vulnerable children. A case of 'PANDAS' is presented to increase physician awareness of this disorder and to document effectiveness of risperidone in chorea and treatment-resistant disruptive behavior associated with this syndrome. To our knowledge, this is the first case report on risperidone in pediatric chorea, although studies on effectiveness of this agent in Tourette's disorders have been previously published. PMID- 10522470 TI - The use of quinolones in pediatric patients. PMID- 10522471 TI - [Dangers of safety]. PMID- 10522472 TI - [Interferon-beta in multiple sclerosis--who is going to be treated?]. PMID- 10522473 TI - [Should dosing practice in aminoglycosides therapy be changed?]. PMID- 10522474 TI - [Organ transplantation]. PMID- 10522476 TI - [Raeder's syndrome]. AB - Raeder's syndrome was first described by the Norwegian ophthalmologist J.G. Raeder in 1918 and again in 1924 by the same author. The seminal report was a description of a young male patient with unilateral periocular pain combined with ipsilateral miosis and ptosis, and with slight objective signs of trigeminal nerve involvement. Autopsy demonstrated a tumour at the base of the skull in the middle cranial fossa. Raeder coined the term "paratrigeminal" for the reported clinical picture. Later case reports by Raeder and other authors have included patients experiencing a more benign clinical course, some with spontaneous remissions, with unilateral periocular pain and ipsilateral signs of oculosympathetic paresis as the common denominator. This article is a chronological survey of the main contributions to the medical literature. Various definitions of the syndrome are outlined, including the more recent classification, as well as some pathophysiological and prognostic considerations. PMID- 10522475 TI - [Johan Georg Raeder and Raeder's syndrome]. PMID- 10522477 TI - [Surgical treatment of ulcerative colitis]. AB - Continence-preserving coloproctectomy for ulcerative colitis is technically demanding and is relatively often afflicted with complications. We have retrospectively reviewed the files of all patients being operated for ulcerative colitis at Ulleval Hospital from 1992 to 1997 (n = 53). Most of the patients (n = 50) were examined clinically; 12 patients had anal manometry before and after operation. 44 patients were operated with continence-preserving coloproctectomy with J-pouch and handsewn anastomosis; of these, 42 were followed more than six months. Eight had pouchitis, four perianal abscess/fistula, three septicaemia and three were operated for ileus. Two had anastomotic leakage and pelvic abscess that required transanal drainage. One had the pouch removed six years after operation due to chronic pouchitis and pouch-vaginal fistula. There was no deterioration of anal maximal resting and squeezing pressures on pre- and post operative anal manometry. Mean number of stools from the reservoir per 24 hours were 6.2 (range 3-11); 11 patients had leakage of air and loose stool, three at day-time and eight at night. Two patients (4%) died from colorectal cancer and three (7%) had Crohn's disease. Nine patients were unfit for pouch surgery and underwent coloproctectomy (n = 7) or subtotal colectomy (n = 2). Our results indicate that pouch surgery for ulcerative colitis is a good option for most patients. PMID- 10522478 TI - [Polychondritis--a rare and difficult diagnosis]. AB - We present two patients with polychondritis. Polychondritis is a rare inflammatory disease of unknown origin, but immunological mechanisms are essential in the pathogenesis. Histological features are inflammation and destruction of cartilage. The disease is systemic, may have a remitting course, and it can be primary or associated with several other diseases. We also review the literature with emphasis on symptoms, clinical presentation, diagnostic procedures and treatment. The literature consists mainly of case reports, summary of case reports, no epidemiological and only few clinical studies. Two sets of diagnostic criteria with great similarities have been proposed. Non-steroidal antiinflammatory drugs or immunosuppressive drugs like glucocorticoids and cyclophosphamide are the drugs of choice, depending on the stage and severity of the disease. PMID- 10522479 TI - [Cryptococcal meningitis in a patient without known predisposing disease]. AB - Cryptococcal meningitis is a rare disease. It may occur as a superinfection in AIDS patients or other immunosuppressed patients. We describe a case of cryptococcal meningitis in a non-immunosuppressed patient. Initial symptoms were fatigue, depression and headache. A correct diagnosis was made after two weeks based on microscopic examination of cerebrospinal fluid. The patient died after six days on antimycotic therapy. Cryptococcosis is a difficult diagnosis, as our case illustrates. Psychiatric symptoms are often the first clinical manifestations. Early diagnosis is crucial for the outcome. A short overview on cryptococcosis is given. PMID- 10522480 TI - [Sick leave while on a waiting list]. AB - Evidence suggest that waiting for hospital treatment leeds to unnecessary sick leaves. However, we know little about the extent of the problem. During a six week period, 1,061 patients attending a total of ten hospital departments for the first time for outpatient treatment or examination received a self-administered questionnaire about sick leaves during the waiting period. 901 patients responded. Great variations were found between the ten departments. The average waiting period was 86 days (32-226 days). 26% (8-61%) of the patients had had a sick leave during the waiting period caused by the disease for which they were seeking treatment; average duration was 48 days (17-89 days); total sick leave for all patients responding in this survey was 11,000 days. Of patients with a sick leave during the waiting period, 44% thought that the examination or treatment which they attended would increase their ability to work, while their doctors were somewhat less optimistic (36%). The study shows that a large proportion of patients on waiting lists for hospital treatment are incapable of work and that cost savings achieved by cutting down waiting period may be considerable. PMID- 10522481 TI - [Interferon therapy of multiple sclerosis]. AB - The use of beta-interferon in the relapsing-remitting phase of multiple sclerosis is a new though fairly established therapy. The beneficial effect of this treatment is thought to be due to immune modulation, with inhibitory action on proliferation of leukocytes and antigen presentation, an increased amount of interleukins, but no change in the proinflammatory cytokine interferon-gamma. In Norway, one interferon-beta 1b and two interferon-beta 1a products have recently been introduced for administration by either subcutaneous or intramuscular injection. Approximately 30% reduction in the relapse rate is reported for both products,but differences appear in the effect on the disability progression, and also with regard to adverse reactions. Patients less strongly affected by the illness seem to have the best outcome. Neutralizing antibodies are often produced during this treatment, but the significance of this is still unclear. Results from international studies on interferon therapy of multiple sclerosis are discussed. Initial results from studies on interferon treatment of secondary progressive multiple sclerosis are now available. A further expansion, also in Norway, of the indications for beta-interferon therapy of multiple sclerosis is expected in the near future. PMID- 10522482 TI - [Nerve biopsy]. AB - Nerve biopsy, if undertaken at centres with appropriate expertise, may be a valuable diagnostic procedure in selected patients with peripheral neuropathy. Sural nerve biopsy and skin biopsy for evaluation of epidermal nerves are described. Indications, surgical procedure, preparation and transport are reviewed. Usually the patient has to be admitted to a hospital with the necessary resources for a comprehensive battery of nerve tissue processing, including teasing and electron microscopical examination, and other investigative procedures. Another option, when practically and economically feasible, is to obtain sural nerve biopsy from several (2-4) patients performed by a skilled surgeon at the local (central) hospital. A technician from a neuromuscular centre or laboratory of neuropathology should be at the site to provide immediate and proper preparation and transport of the tissue samples. An example of peripheral nerve vasculitis with teased nerve fibers in early and late axonal degeneration is presented as an illustration. Sural nerve biopsy is useful in selected cases, particularly in order to demonstrate peripheral nerve vasculitis. For detection of small fiber neuropathy in the investigation of painful neuropathies, skin biopsy for evaluation of epidermal nerves may be an appropriate alternative. PMID- 10522483 TI - [Once-daily dosage of aminoglycosides--a therapeutic simplification and an economical benefit]. AB - Conventionally, aminoglycosides have been administered in two or three daily doses. Numerous in-vitro and animal experiments and several clinical trials favour a once-daily dosage regimen of aminoglycosides, which provides a more rapid concentration-dependent bacterial killing and is probably less toxic than two or three dosage regimens. In this article the pharmacological and microbiological background for once-daily aminoglycoside administration is reviewed, and some controlled trials are discussed. Recommendations for clinical practice are given. PMID- 10522484 TI - [Blood supply readiness in case of war or disasters]. AB - The supply and use of blood during war or disaster require efficient cooperation between national and regional civilian authorities and blood banks as well as the military chain of command and medical units. Tasks and responsibilities are regulated by a directive issued by the Norwegian Armed Forces Joint Medical Services, specifying that during crises, civilian blood banks will be ordered to provide blood to military units. Based upon information obtained by a questionnaire survey, blood banks have an adequate capacity even if the normal power supply or computerised systems fail temporarily. However, plans are presently lacking for the organisation and operation of the civilian blood supply during a crisis requiring cooperation with military units. Key roles and responsibilities for personnel on the civilian side must be defined. Normal and alternative routes of communication must be established and tested regularly. PMID- 10522485 TI - [Kidney transplantation in Norway--a historical perspective]. AB - Until the late 1950s, few treatment modalities for patients with chronic renal failure were available. The first successful renal transplantation was performed in Boston, USA in 1954 when a young man with renal failure received a kidney from his monozygotic twin brother. Tissue typing was not available, there were few options for immunosuppressive treatment, and dialysis could be offered for short term treatment only. At the Surgical Department A of the National Hospital, Oslo, Norway, treatment of acute renal failure by hemodialysis was started in 1956 and the first renal transplantation in Scandinavia performed the same year. In 1963, the first renal transplantation at Ulleval Hospital, Oslo was performed, the first in Norway with a long-term function transplant. Since 1983, all renal transplantations in Norway have been performed at the National Hospital. In 1969, Scandiatransplant was established as an organisation for organ exchange between the Scandinavian countries, and a national program for renal transplantation in Norway was launched. The Norwegian renal transplant program is based on co operation between regional and central hospitals and the transplant centre at the National Hospital. In recent years, the number of renal transplantations per year has been 180-200, i.e. 40-45 per million population. Approximately 40% of the transplanted kidneys come from living related or spouse donors. The results in terms of renal graft survival are good. PMID- 10522486 TI - [Immunologic reactions in transplantation]. AB - Transplantation is the therapy of choice for an increasing number of patients. Organ transplantation is used to treat irreversible kidney, heart, lung and liver failure. Stem cell transplantation (previously called bone marrow transplantation) is used to treat leukaemia, bone marrow failure, severe combined immunodeficiencies and various congenital metabolic disorders. After organ transplantation, the recipient's immune system will recognise and reject the transplanted organ. T-lymphocytes transferred along with the stem cells in stem cell transplantation will attack the recipient. This paper describes how immunocompetent cells (T- and B-lymphocytes) recognise and destroy foreign cells. Strategies for the induction of specific immunologic tolerance are briefly explained. PMID- 10522487 TI - [Kidney biopsies and video conferences]. PMID- 10522488 TI - [Should the shape of child's head be standardized?]. PMID- 10522489 TI - [Why does the prevalence of allergy increase more in industrialized countries than in developing countries?]. PMID- 10522490 TI - [Safety guarantee and the medical profession]. PMID- 10522491 TI - [Disinfection of air with ultraviolet rays]. PMID- 10522493 TI - [The rules of the drug industry association is contempt of physicians]. PMID- 10522492 TI - [What do we do in connection with failed competence?]. PMID- 10522494 TI - Lack of promotional effects of groundwater contaminant mixtures on the induction of preneoplastic foci in rat liver. AB - F344 rats were exposed to drinking water mixtures of seven of the most common groundwater contaminants associated with hazardous waste sites [arsenic, benzene, chloroform, chromium, lead, phenol, and trichloroethylene (TCE)] as the full mixture or submixtures of the organic and/or inorganic chemicals. The lowest concentrations (1x) of the individual chemicals were environmentally realistic and below what would be expected to induce significant short-term toxicity. This study was intended to determine if previously reported increases in localized hepatocellular proliferation in response to these chemicals might be correlated with increased risk for hepatocarcinogenesis. Rats were exposed via a drinking water solution to the full seven- chemical mixture (at 1x and 10x concentrations), submixtures of the organic or inorganic chemicals (at 10x concentrations), a mixture of TCE, lead, and chloroform (TLC submixture at 10x and 100x concentrations), or deionized water as a control. The rats were evaluated for promotion of placental glutathione-S-transferase (GST-P) positive preneoplastic liver cell foci after diethylnitrosamine (DEN) initiation and partial hepatectomy. Focus formation, cell proliferation, and apoptosis were evaluated after exposure to DEN or saline controls, the chemical mixtures or deionized water controls, or combinations of these treatments. The total number and area of GST-P positive foci in DEN-treated rats exposed to the full seven chemical mixture was increased as compared with the DEN-water controls, but this was statistically significant only for total focus area in the 1x dose group. In DEN-treated rats, the inorganic or TLC submixtures resulted in a significant reduction in number and area of GST-P positive foci. Focus area also was decreased in the organic submixture-treated group, but not significantly. Hepatocellular proliferation was not significantly changed in the chemical mixture saline groups as compared with the mixture water controls. After DEN treatment, however, cell proliferation was significantly decreased after the 10x seven-chemical and organic mixture treatments and the 100x TLC mixture treatment. Different groups showed either increased or decreased apoptotic rates which did not correlate well with proliferation rates or focus formation. Mixtures of these seven chemicals, therefore, did not appear to act as promoters of hepatic foci at environmentally relevant concentrations, and some mixture combinations appeared to decrease promotional activity. PMID- 10522495 TI - Effects of pretreatment with cytochrome P-450 inducers, especially phenobarbital on triphenyltin metabolism and toxicity in hamsters. AB - The effects of cytochrome P-450 (CYP) induction by phenobarbital (PB), CYP 2B, 2C, and 3A inducer in mammalians, on triphenyltin metabolism and toxicity in hamsters were studied. A single dose of 50 mg/kg of triphenyltin chloride was given by gavage to hamsters after pretreatment with or without PB for 3 days continuously at a daily dose of 80 mg/kg intraperitoneally (i.p.). Although the triphenyltin produced marked but reversible hyperglycemia and hypertriglyceridemia in PB-untreated hamsters, the pretreatment of hamsters with PB, which increased levels of CYP, suppressed the diabetogenic effects compared with PB-untreated hamsters. Furthermore, we investigated whether the mitigation of triphenyltin-induced diabetogenic toxicity by PB pretreatment is due to an alteration of triphenyltin metabolism. Triphenyltin and its metabolites in liver, kidneys, pancreas and brain were determined by gas chromatography periodically for 96 h after triphenyltin administration in both groups of hamsters. The initial triphenyltin levels in the tissues of PB-pretreated hamsters were about half of those in the tissues of PB-untreated hamsters and PB pretreatment accelerated metabolism of triphenyltin at early stage in hamsters. We also examined the other CYP 1A and 2A inducers, beta-naphthoflavone (B-NF) and 3 methylcholanthrene (MC). The PB pretreatment showed the strongest suppression of the toxicity at 24 h after the triphenyltin intubation, compared with the effects of B-NF and MC. In addition, the maximum proportion of diphenyltin to parent triphenyltin in pancreas was observed in PB-treated hamsters. These findings suggest that the induction of CYP system enzymes affects the metabolism and toxicity of triphenyltin in hamsters. Especially, based on effects of PB and other CYP inducers, PB induction has a key role in suppressing the diabetogenic action of triphenyltin, i.e. by decreasing triphenyltin accumulation in the hamsters. PMID- 10522496 TI - Chelation therapy in aluminum-loaded rats: influence of age. AB - The influence of age at which aluminum (Al) exposure was initiated on the efficacy of chelation therapy in mobilizing Al was investigated in two groups of male rats exposed to this element at two different stages of the life cycle. Young (21 days old) and old (18 months) rats were exposed to 0 and 50 mg Al/kg/day administered as Al nitrate in drinking water for a preliminary period of 14 days followed by a period of 100 days, in which Al-exposed animals received 100 mg Al/kg/day. At the end of the period of exposure, Al-loaded rats in each age group were given one of the following treatments: s.c. deferoxamine (DFO), oral 1,2-dimethyl-3-hydroxypyrid-4-one (L1) and 1-(p-methylbenzyl)-2-ethyl-3 hydroxypyrid-4-one (MeBzEM) at doses of 0.89 mmol/kg/day for 5 consecutive days. Another group of Al-exposed rats received a concurrent administration of s.c. DFO and oral L1 both at 0.45 mmol/kg/day. During chelation therapy urines were collected daily. Control groups included rats exposed and unexposed to Al. Oral administration of L1 was the most effective treatment in enhancing urinary Al excretion in both age groups of Al-loaded rats. This beneficial effect was similar for old and young animals. Concurrent administration of DFO and L1 had no advantages over the use of either single agent, while MeBzEM was not effective in mobilizing Al from Al-exposed rats. PMID- 10522497 TI - Protein binding of mercury in milk and plasma from mice and man--a comparison between methylmercury and inorganic mercury. AB - Inorganic mercury has previously been shown to be excreted to milk from plasma to a higher extent than methylmercury. Protein binding of mercury as methylmercury and inorganic mercury in whey and plasma from mouse and man was studied in order to get a better understanding of the transport of mercury into milk. Mice were administered a single i.v. dose of 0.25 mg Hg/kg body weight labelled with (CH3)203HgCl or 203HgCl2, resulting in 11 ng Hg/g milk and 38 ng Hg/g milk after 1 h, respectively. Milk and plasma from mice and man were also incubated with the respective radiolabelled compound (150 ng Hg/g milk or plasma). Casein, fat and whey fractions in milk from methylmercury treated mice were found to contain 11, 39 and 34%, respectively, and from inorganic mercury treated mice 31, 15 and 41%, respectively, of the total amount of mercury in milk. Serum albumin was a major mercury binding protein in whey and plasma from mice for both methylmercury and inorganic mercury, as demonstrated by FPLC gel filtration and anion-exchange chromatography and further characterised by SDS-PAGE for whey. In addition, anion exchange chromatography indicated that inorganic mercury, but not methylmercury, in whey from mouse milk formed a dimer of serum albumin. The unbound fraction of mercury in whey and plasma from mice was very small (<0.7%), and somewhat higher in plasma and whey from man. It is concluded, that the unbound fraction in plasma cannot be a determining factor for the observed differences in milk excretion between the two mercury compounds. Instead, it is suggested that methylmercury and to some extent inorganic mercury are transferred from plasma into milk using albumin as a passive carrier. PMID- 10522498 TI - The American Board of Otolaryngology, 1924-1999: 75 years of excellence. PMID- 10522499 TI - Biochemoprevention for dysplastic lesions of the upper aerodigestive tract. AB - OBJECTIVES: To evaluate the efficacy and secondarily the toxic effects of biochemopreventive therapy (high-dose isotretinoin [13-cis-retinoic acid], alpha tocopherol, and interferon alfa) in the reversal of advanced premalignant lesions of the upper aerodigestive tract and to correlate the therapeutic events with modulation of biomarkers. DESIGN: Prospective, nonrandomized chemoprevention trial. SETTING: Tertiary cancer care referral center and ambulatory care. PARTICIPANTS: Thirty-six patients with advanced premalignant lesions of the upper aerodigestive tract, without cancer during the 2 years before the intervention, with evaluable lesions, and without retinoid therapy for 3 months before the trial. INTERVENTION: Administration of oral isotretinoin (100 mg/m2 per day), oral alpha-tocopherol (1200 IU/d), and subcutaneous interferon alfa (3 megaunits per square meter twice weekly) for 12 months, with serial biopsies and clinical examination at 0, 6, 12, and 18 months from study start. MAIN OUTCOME MEASURES: Clinical and histologic responses to the intervention. RESULTS: Of the 36 patients, evaluation was possible in 30 for response at 6 months and in 21 at 12 months. At 6 months, there were 10 pathologic complete responses and 7 partial responses; at 12 months, 7 complete and 3 partial responses. A striking difference in response was observed in favor of laryngeal lesions (9/19 [47%] complete response rate at 6 months and 7/14 [50%] at 12 months vs 1/11 [9%] and 0/7 [0%], respectively, for oral lesions). Toxic effects were acceptable and did not exceed grade 3. CONCLUSION: Biochemoprevention is a promising biologic approach for laryngeal dysplasia and needs to be investigated further. PMID- 10522500 TI - Use of topical ascorbic acid and its effects on photodamaged skin topography. AB - OBJECTIVE: To determine the efficacy of topical ascorbic acid application in treating mild to moderate photodamage of facial skin using an objective, computer assisted image analysis of skin surface topography and subjective clinical, photographic, and patient self-appraisal questionnaires. DESIGN: A 3-month, randomized, double-blind, vehicle-controlled study. SETTING: Facial plastic surgery private practice. PATIENTS: Nineteen evaluable volunteer sample patients aged between 36 and 72 years with Fitzpatrick skin types I, II, and III who were in good physical and mental health with mild to moderately photodamaged facial skin were considered for analysis. INTERVENTION: Coded, unmarked medications were randomly assigned to the left and right sides of each subject's face, one containing the active agent, topical ascorbic acid (Cellex-C high-potency serum; Cellex-C International, Toronto, Ontario), the other, the vehicle serum (Cellex-C International). Three drops (0.5 mL) of each formulation were applied daily to the randomly assigned hemifaces over the 3-month study period. Treatment assignments were not disclosed to subjects, clinicians, or personnel involved in analyzing skin replicas. MAIN OUTCOME MEASURES: Specific clinical parameters were evaluated and graded on a 0- to 9-point scale (0, none; 1-3, mild; 4-6, moderate; and 7-9, severe). Reference photographs were used to standardize grading criteria. Overall investigator scores were compared with baseline and graded as excellent (much improved), good (improved), fair (slightly improved), no change, or worse. Patient self-appraisal questionnaires rated the degree of improvement (much improved, improved, slightly improved, no change, or worse) and reported adverse effects (burning, stinging, redness, peeling, dryness, discoloration, itching, and rash). Standard photographs were taken at baseline, including anteroposterior and left and right oblique views to facilitate subsequent clinical evaluations, and at the end of therapy for comparison. Optical profilometry analysis was performed on the skin surface replicas of the lateral canthal (crow's feet) region, comparing baseline to end-of-study specimens. Using this computer-based system, the resulting image was digitally analyzed, and numeric values were assigned to reflect surface features. The parameters obtained included Rz, Ra, and shadows. These values provided objective data that document pretreatment and posttreatment texture changes proportional to the degree of wrinkling, roughness, and other surface irregularities. RESULTS: Optical profilometry image analysis demonstrated a statistically significant 73.7% improvement in the Ra and shadows north-south facial axis values with active treatment greater than vehicle control, as well as a trend for improvement in the Rz north-south facial axis parameter, showing a 68.4% greater improvement of active treatment vs vehicle control. Clinical assessment demonstrated significant improvement with active treatment greater than control for fine wrinkling, tactile roughness, coarse rhytids, skin laxity/tone, sallowness/yellowing, and overall features. Patient questionnaire results demonstrated statistically significant improvement overall, active treatment 84.2% greater than control. Photographic assessment demonstrated significant improvement, active treatment 57.9% greater than control. CONCLUSIONS: A 3-month daily regimen of topical ascorbic acid provided objective and subjective improvement in photodamaged facial skin. Skin replica optical profilometry is an objective method for quantification of the skin surface texture changes. PMID- 10522501 TI - The role of image-guidance systems for head and neck surgery. AB - BACKGROUND: Although image-guidance systems have gained widespread acceptance for neurosurgical procedures, their role for extracranial surgery of the head and neck is yet to be defined. OBJECTIVES: To describe the authors' experience with image-guidance systems and to measure the effects of image-guided technology on the performance of minimally invasive otolaryngological procedures. DESIGN: Prospective cohort study. METHODS: Optical- and electromagnetic-based image guidance systems were used during the performance of endoscopic surgery on patients with disease of the paranasal sinuses, orbit, skull base, and temporal bone (n = 79). Results were compared with those in control patients who underwent similar surgery without image guidance during the same period (n = 42). RESULTS: Intraoperative anatomical localization was accurate to within 2 mm at the start of surgery in all cases. Accuracy degraded by 0.89 +/- 0.20 mm (mean +/- SE) during the operative procedure. The use of an image-guidance system increased operating room time by a mean of 17.4 minutes per case (image-guidance group, 137.3 +/- 6.0 minutes [mean +/- SE]; control group, 119.9 +/- 5.7 minutes; P=.006) and increased hospital charges by approximately $496 per case. Intraoperative blood loss (image-guidance group, 178.4 +/- 18.0 mL [mean +/- SE]; control group, 149.4 +/- 20.1 mL) and complication rates (image-guidance group, 2.7%; control group, 4.7%) did not differ significantly between groups. CONCLUSIONS: Image-guidance systems can provide the head and neck surgeon with accurate information regarding anatomical localization in cases with poor surgical landmarks caused by extensive disease or prior surgery; however, the use of such systems is associated with increased operative time and expense. PMID- 10522502 TI - Electroglottography in the pediatric population. AB - OBJECTIVE: To establish normative electroglottography (EGG) data in the pediatric population. DESIGN: Clinical study with EGG data gathered on children with normal voices. SETTING: Major children's hospital and specialty eye and ear hospital. PATIENTS: A total of 164 children, 79 girls and 85 boys, aged 3 to 16 years. METHODS: Children with normal voices, determined through subjective evaluation and a voice use history questionnaire, underwent EGG recording. The EGG data were analyzed with commercially available software for fundamental frequency, jitter, open quotient, closing quotient, and opening quotient. RESULTS: Normative EGG data were established for children aged 3 to 16 years. Jitter, open quotient, closing quotient, and opening quotient were all found to have no significant dependence on age. CONCLUSIONS: Children as young as 3 years can easily tolerate EGG, making it possible to establish this initial set of normative pediatric EGG data. These preliminary results suggest that EGG may have potential to assist clinicians with noninvasive documentation of vocal function in the pediatric population. This maybe particularly important for tracking treatment-related changes in the vocal function of children who are difficult to examine endoscopically. PMID- 10522503 TI - Surgery for cervicofacial nontuberculous mycobacterial adenitis in children: an update. AB - OBJECTIVE: To assess optimal surgical treatment with excision or curettage techniques in children with cervicofacial nontuberculous mycobacterial (NTM) adenitis. DESIGN: Retrospective case series. SETTING: Tertiary university-based pediatric referral center. PATIENTS: Patients younger than 18 years diagnosed as having cervicofacial NTM adenitis by positive mycobacterial cultures or stains, or by histopathologic evaluation. INTERVENTIONS: Fine-needle aspiration biopsy for diagnosis, surgical excision and/or curettage of head and neck lesions for treatment. MAIN OUTCOME MEASURES: Number of procedures per patient, complications, resolution of mass. RESULTS: A total of 32 surgical procedures were performed in 25 children with cervicofacial NTM adenitis (mean, 1.3 procedures per patient; range, 1-3): 19 excisional and 13 curettage procedures. The 14 children who had excision as an initial procedure required no additional surgery. Of 11 children who had curettage as an initial procedure, 6 (55%) required additional procedures. Three of these children had additional surgery as planned staged procedures. Excisional surgery after initial curettage (5 patients) was simplified by initial debridement and secondary healing. No complications of curettage were noted. Transient marginal mandibular nerve weakness was seen in 4 patients who had excision. Fourteen of 16 fine-needle aspiration biopsy specimens were diagnostic for NTM adenitis. CONCLUSIONS: Cervicofacial NTM adenitis can be treated with excision or curettage. Excision remains the treatment of choice because of the high cure rate with a single procedure. We now consider curettage as a staged procedure for lesions in proximity to the facial nerve or with extensive skin necrosis, with initial curettage simplifying subsequent excision and wound closure. Preoperative counseling should include discussion of planned or unplanned revision surgery after curettage. Fine-needle aspiration biopsy allows early diagnosis of NTM adenitis. PMID- 10522504 TI - The microbiology of recurrent rhinosinusitis after endoscopic sinus surgery. AB - OBJECTIVE: To determine the microbiology of recurrent sinus infections occurring in patients after endoscopic sinus surgery (ESS). DESIGN: Retrospective review of sinus cultures obtained over a 4-year period from a consecutive series of patients who underwent ESS. SETTING: An academic general otolaryngology practice. RESULTS: A total of 290 cultures were performed in 125 patients after ESS. The female-male ratio of cultures was 2.5:1 with an average patient age of 47.3 years. This group of patients represents 14.5% of 860 patients who underwent ESS during the same period. A total of 65 patients had 1 culture performed, and 60 patients had multiple cultures. Of the 290 culture specimens, 87 (30.0%) demonstrated no growth. Gram-positive cocci predominated, accounting for 37.9% of culture results. Gram-negative rods constituted 14.8% of the isolates. Of the cultures yielding gram-negative rods, 90.7% occurred in patients who had multiple cultures (P = .03). Fungal forms were cultured in 1.7% of the specimens. None of the Streptococcus pneumoniae isolates demonstrated penicillin-based resistance. The percentages of beta-lactamase-producing strains for Haemophilus influenzae and Branhamella (Moraxella) catarrhalis were 45.4% and 81.8%, respectively. Staphylococcal species also exhibited significant antibiotic resistance patterns, but no statistical association with multiple cultures was noted (P = .23). CONCLUSIONS: A wide range of bacteria may be present in the infected post-ESS sinus cavity, with a considerable population of gram-negative organisms, including Pseudomonas species. Beta-Lactamase-producing organisms continue to be prevalent in postoperative sinus infections. Culture and sensitivity analyses of pathologic secretions may identify drug-resistant organisms or organisms related to difficult-to-treat infections in exacerbations of chronic rhinosinusitis in the postoperative setting. PMID- 10522505 TI - IgE-mediated reactions and hyperreactivity in pregnancy rhinitis. AB - OBJECTIVES: To determine whether respiratory allergy or hyperreactive nasal mucosa is exceptionally common in women with pregnancy rhinitis, and to evaluate other possible risk factors such as clinical asthma or rhinitis, smoking, age, parity, and sex of the child. PATIENTS AND METHODS: From an antenatal questionnaire study, 165 women, 83 (50%) of whom had had pregnancy rhinitis, were examined 6 months after delivery, and multiple antigen simultaneous testing chemiluminescent assay (MAST CLA) (10 airborne allergens) was performed. After histamine provocations, rhinostereometry and acoustic rhinometry were performed in 25 of them. Serum levels of soluble intercellular adhesion molecule-1 were determined 4 times during and once after pregnancy in 5 women with pregnancy rhinitis and 17 without pregnancy rhinitis. RESULTS: Thirty-nine women (24%) were sensitized to 1 or more allergen. The pregnancy rhinitis group showed significantly higher levels of IgE to house dust mites. There were also more smokers in the pregnancy rhinitis group. Clinical asthma or rhinitis, age, parity, and sex of the child did not differ significantly between the 2 groups. Mucosal swelling increased with rising concentrations of histamine, as measured with rhinostereometry, but there was no significant difference between the 2 groups in any of the variables. Serum soluble intercellular adhesion molecule-1 was not elevated in the pregnancy rhinitis group. CONCLUSIONS: This study found no increased frequency of allergy in general in women who have had pregnancy rhinitis. However, IgE against house dust mite was more frequent in the pregnancy rhinitis group. Smoking seems to be a risk factor, but age, parity, sex of the child, and hyperreactive nasal mucosa do not. Soluble intercellular adhesion molecule-1 was not elevated during pregnancy rhinitis. PMID- 10522506 TI - Ten days' use of oxymetazoline nasal spray with or without benzalkonium chloride in patients with vasomotor rhinitis. AB - CONTEXT: In most countries, the use of topical nasal decongestants is limited to a maximum of 10 days because of the risk of developing rebound mucosal swelling and rhinitis medicamentosa. OBJECTIVE: To determine whether topical nasal decongestants can be safely used for 10 days in patients with chronic inflammation of the nasal mucosa. DESIGN: Double-blind, randomized, controlled, parallel study. PATIENTS: Thirty-five patients with vasomotor rhinitis selected from our outpatient department. INTERVENTION: Eighteen patients received oxymetazoline hydrochloride (0.5 mg/mL) nasal spray containing the preservative benzalkonium chloride (0.1 mg/mL), and the other 17 were treated with oxymetazoline nasal spray without benzalkonium chloride. Before and after the treatment, recordings of the nasal mucosa and minimal cross-sectional area were made with rhinostereometry and acoustic rhinometry, followed by histamine hydrochloride challenge tests. Symptoms of nasal stuffiness were estimated on visual analog scales (0-100) in the morning and the evening, just before the nasal spray was used. RESULTS: No rebound swelling was found after the 10-day treatment in the 2 groups with either of the methods or as estimated by symptom scores. In the group receiving oxymetazoline containing benzalkonium chloride, but not in the other group, the histamine sensitivity was significantly reduced after treatment (P<.001). CONCLUSIONS: It is safe to use topical nasal oxymetazoline with or without benzalkonium chloride for 10 days in patients with vasomotor rhinitis. However, this study indicates that benzalkonium chloride in nasal decongestant sprays affects the nasal mucosa also after short-term use. PMID- 10522507 TI - Evolution of the bacteriologic features of persistent acute otitis media compared with acute otitis media: a 15-year study. AB - OBJECTIVES: To define the epidemiologic features of persistent acute otitis media (PAOM) and modifications of these features during the past 15 years and to investigate for possible differences in bacterial resistance between acute otitis media (AOM) and PAOM. DESIGN: Retrospective patient series. SETTING: Academic tertiary care center. PATIENTS AND METHODS: Persistent acute otitis media was defined as AOM lasting longer than 3 weeks despite 1 or several courses of antibiotic therapy, with the persistence of clinical and otoscopic signs of AOM. From 1982 to 1997, 475 children with PAOM were seen in our department. Every patient had 1 or several specimens of aspirations or swabs of spontaneous otorrhea (or both). Microbiologic characteristics of the isolated strains (including antibiotic susceptibility) were analyzed. Four successive series of specimens were analyzed-group 1: from October 1, 1982, to June 30, 1986 (136 patients); group 2: from January 1, 1987, to December 31, 1989 (165 patients); group 3: from January 1, 1992, to April 30, 1993 (73 patients); and group 4: from January 1, 1994, to January 31, 1997 (101 patients). During the same study periods, the bacteriologic results of patients with AOM in the same geographic region were recorded. MAIN OUTCOME MEASURES: A longitudinal comparison between the groups of patients with PAOM and a cross-comparison within each group between patients with PAOM and those with AOM. RESULTS: Obtaining repeated and multiple specimens from patients with PAOM led to a progressive decrease in the rate of sterile specimens, from 35.3% (group 1, 48 patients) to 14.9% (group 4, 15 patients) (P<.01). During this period, the prevalence of Streptococcus pneumoniae increased in patients with positive culture results, from 18.2% (group 1, 16 of 88 patients) to 44.2% (group 4, 38 of 86 patients) (P<.001). These strains rapidly and dramatically became resistant to penicillin (amoxicillin) (0% through 1989, 76.2% [16 of 21 patients] in 1993, and 97.4% [37 of 38 patients] in 1996) (P = .01). The overall prevalence of Haemophilus influenzae remained stable (between 31.4% [27 of 86 patients] and 45.4% [40 of 88 patients]), but the proportion of beta-lactamase-producing strains increased from 30.0% (group 1, 12 patients) to 55.6% (group 4, 15 patients) (P=.04). The prevalences of Pseudomonas aeruginosa and Staphylococcus aureus did not vary significantly (from 23.1% [group 2, 30 patients] to 10.7% [group 3, 6 patients] and from 10.2% [group 1, 9 patients] to 4.6% [group 4, 4 patients], respectively). Comparing data from patients with PAOM with those with AOM revealed that the increased resistance of H influenzae and, in particular, of S pneumoniae was more rapid and more marked in patients with PAOM than in those with AOM (highest rate of resistance in AOM: 36.0% [271 of 753 specimens] and 50.6% [398 of 787 specimens] for H influenzae and S pneumoniae, respectively; P<.001 for S pneumoniae). CONCLUSIONS: The increase in bacterial resistance frequently encountered during otitis media is even more marked in patients with PAOM. The identification of the organism is essential when the otitis does not resolve, especially in patients with PAOM. Obtaining repeated specimens helps to decrease the rate of sterile cultures. PMID- 10522508 TI - Intralesional cidofovir for recurrent respiratory papillomatosis in children. AB - OBJECTIVE: To assess the potential benefit of intralesional administration of cidofovir, an acyclic nucleoside phosphonate with activity against several DNA viruses, for treating severe respiratory papillomas in pediatric patients. DESIGN: Prospective case series. SETTING: Tertiary care children's hospitals. PATIENTS: Five pediatric patients with severe recurrent respiratory papillomatosis requiring laryngoscopy with carbon dioxide laser therapy more frequently than once a month to maintain airway patency. Each patient underwent between 12 and 33 laryngoscopies with laser treatment prior to being injected with cidofovir. INTERVENTION: Microsuspension laryngoscopy with intralesional injection of cidofovir (Vistide) in conjunction with mechanical debulking and carbon dioxide laser of papillomas. MAIN OUTCOME MEASURE: Papilloma stage at time of serial laryngoscopies. RESULTS: One patient was disease free and 3 patients demonstrated a dramatic response to adjuvant therapy with cidofovir at the 9 month follow-up visit after the last injection of cidofovir. One patient showed an improvement in papilloma stage that was possibly related to concurrent therapy with interferon. CONCLUSIONS: Intralesional injection of cidofovir seems to be of benefit in the treatment of severe respiratory papillomatosis in pediatric patients. Larger prospective studies with longer follow-up will be required before cidofovir can be considered an accepted means of managing this difficult disease. PMID- 10522509 TI - Venous valves in the neck: implications for microvascular free flap reconstruction. AB - Successful microvascular free flap reconstruction requires adequate arterial inflow and venous outflow. We report 4 cases that demonstrate the not uncommon occurrence of locating valves in veins during microvascular head and neck reconstructive procedures. Failure to recognize these valves could have compromised the venous anastomosis. The anatomical literature states that veins in the head and neck lack valves, allowing bidirectional blood flow. As a result, there is potential significant flexibility in the selection of recipient veins for the microvascular anastomosis during free flap reconstruction. The unrecognized presence of a venous valve, however, could cause thrombosis of the venous anastomosis and, ultimately, flap failure. This report of venous valves should speak caution to the head and neck microvascular surgeon when he or she is selecting recipient veins in the neck. PMID- 10522510 TI - Cricoid cartilage necrosis after arytenoidectomy in a previously irradiated larynx. AB - Several open and endoscopic surgical techniques are available to provide an adequate airway for patients with bilateral vocal cord paralysis. Transoral laser arytenoidectomy has repeatedly been reported to be a reliable and effective minimally invasive procedure for airway restoration. To our knowledge, there have been no previous reports of serious complications, other than poor vocal results, aspiration, and failed decannulation in individual patients, that have resulted from this intervention. We report a case in which arytenoidectomy led to severe complications and death. Prior irradiation is suspected to be a causative factor. To prevent such an outcome, we believe that operative settings should be chosen that avoid deep thermal injury of the laryngeal framework. PMID- 10522511 TI - Quiz case 1. Squamous cell carcinoma. PMID- 10522512 TI - Quiz case 2. Coccidioidomycosis. PMID- 10522513 TI - Removal of the inferior half of the superficial lobe is sufficient to treat pleomorphic adenoma in the tail of the parotid gland. PMID- 10522514 TI - Conservative vs superficial parotidectomy. PMID- 10522515 TI - Conservative vs superficial parotidectomy for benign lesions of the parotid tail. PMID- 10522516 TI - Diagnosis of allergic fungal sinusitis vs a mucocele. PMID- 10522517 TI - Caution advised when using diluted phenylephrine hydrochloride. PMID- 10522518 TI - Extracerebral biopsies in neurodegenerative diseases of childhood. AB - Among the numerous neurodegenerative diseases in children few may allow morphological diagnosis by extracerebral biopsy. These encompass neurometabolic conditions, foremost lysosomal disorders, but also peroxisomal and mitochondrial diseases marked by disease- or group-specific organelles. Largely, these neurometabolic conditions can also be diagnosed by biochemical and increasingly by molecular genetic techniques. However, there are a few neurodegenerative diseases which do not allow either biochemical or molecular genetic diagnosis and, thus, rely on biopsy of extracerebral tissues, so-called 'essential' biopsies to achieve a diagnosis during the patient's life. Among these few disorders only Lafora disease, as other polyglucosan disorders, may be considered a neurometabolic disease, whereas in the others, neuroaxonal dystrophies, giant axonal neuropathy and neuronal intranuclear inclusion disease no metabolic abnormalities are known, but these disorders share the peripheral nervous system as a common site of their disease-specific morphological lesions. With the progress of molecular genetics and the fact that many neurodegenerative diseases are familial, it is expected that the number of neurodegenerative disorders and the number of patients afflicted with these diseases, currently subject to diagnostic extracerebral biopsies, will be continuously reduced. Thus, it is foreseeable that within the next few years or decades diagnostic electron microscopy and the related knowledge of respective ultrastructural pathology may become outmoded, and, possibly, unknown to future generations of neuropathologists and other members of the neuroscience community. PMID- 10522519 TI - Single-photon emission computed tomography: ictal perfusion in childhood epilepsies. PMID- 10522520 TI - The natural history of somatic morbidity in disintegrative psychosis and infantile autism: a validation study. AB - In order to study the validity of disintegrative psychosis (DP) as defined in ICD 9, we compared the natural history of somatic morbidity of 13 patients given this diagnosis in childhood with a control group of 39 patients with infantile autism (IA) matched for gender, age, IQ and social class. Average follow-up time was 22 and 23 (11-33) years, respectively. Significantly more DP patients (85 versus 41%) had been admitted to a non-psychiatric hospital during the follow-up period. They also had significantly more admissions (3.6 versus 1.0) and stayed longer in hospital (78 versus 4 days) than patients with IA. Three of the DP individuals had an associated medical disorder and made extensive use of somatic services during the follow-up period. Altogether the DP group had utilised the medical health care system more than patients with IA suggesting that they had more medical symptoms than the IA group. On the whole our findings suggest that individuals with DP and IA should be conceptualised as essentially distinct and should be studied separately as regards aetiology, pathophysiology, course and treatment. PMID- 10522521 TI - Neurodevelopmental outcomes of infants with birth weights of less than 1000 g: comparison between periods before and after the introduction of surfactant. AB - In order to assess the effect of surfactant therapy on the neurodevelopmental outcome of infants with a birth weight of less than 1000 g, we evaluated the outcome of 169 infants in the period after introduction of the surfactant (January 1989 to May 1993) by a follow-up of 6 years on the average, and compared these results to those of 107 infants in our previous study before the surfactant period (October 1981 to March 1986) by a follow-up of 7.5 years on the average. Nursery survival rate was significantly increased from 69.3% in the presurfactant period to 80% in the surfactant period (P < 0.05). However, there were no substantial differences in the neurodevelopmental outcomes between the two periods. This enhanced survival rate without the improvement of outcome resulted in a proportionate increase in normal and impaired survivors. PMID- 10522522 TI - Clinical and immunological significance of neopterin measurement in cerebrospinal fluid in patients with febrile convulsions. AB - Neopterin is synthesized mainly by monocytes/macrophages and is considered to be a marker for activation of the cellular immune system. It has been reported that cerebrospinal fluid (CSF) neopterin levels are significantly higher in patients with bacterial meningitis than in those with aseptic meningitis or non pleocytotic CSF. In this study levels of neopterin and interferon-gamma (IFN gamma) were measured in children with non-pleocytotic CSF. The CSF neopterin levels were significantly higher in patients with typical febrile convulsions (FCs) (15.0 +/- 4.5 nmol/l) than in those with pyrexia without convulsions (6.5 +/- 2.7 nmol/l) or convulsions without pyrexia, namely, epilepsy (4.8 +/- 2.4 nmol/l). The CSF neopterin/serum neopterin ratio (C/S ratio) was also higher in patients with typical FCs (1.54 +/- 0.83) than in those with pyrexia without convulsions (0.32 +/- 0.18) or convulsions without pyrexia (0.77 +/- 0.28). Patients with prolonged FCs tended to have higher CSF neopterin levels than those with typical FCs. There was also a tendency for CSF IFN-gamma levels to be higher in patients with FCs than in those with pyrexia without convulsions or convulsions without pyrexia. The results of the present study suggest that some immune activation in the central nervous system (CNS) compartment may be related to the mechanisms of FCs. PMID- 10522523 TI - A comparative study of high-dose and low-dose ACTH therapy for West syndrome. AB - PURPOSE: A prospective randomized controlled study was conducted for the purpose of identifying the lowest effective ACTH dose, with the fewest adverse effects, for the treatment of West syndrome (WS). SUBJECTS AND METHODS: Twenty-five subjects with cryptogenic (CWS, n = 9) or symptomatic (SWS, n = 16) WS were enrolled in this study. They were randomly assigned to receive either low-dose (0.005 mg/kg per day = 0.2 IU/kg per day) or high-dose (0.025 mg/kg per day = 1 IU/kg per day) synthetic ACTH therapy. ACTH was administered every morning for 2 weeks and tapered to zero over the subsequent 2 weeks. Both effectiveness and adverse effects were compared between the two treatment regimens in each type of WS. RESULT: After completion of the treatment protocol in the CWS group, spasms and hypsarrhythmia were completely suppressed in 3/4 (75%) given the low-dose and 5/5 (100%) given the high-dose treatment. In the SWS group, the spasms and hypsarrhythmia disappeared in 6/8 (75%) in each dose group. There were no significant differences in initial responses between the low-dose and high-dose treatments for either type of WS (P > 0.05). Long-term seizure and developmental outcomes, assessed in the 17 responders who were followed up for longer than 1 year after the completion of ACTH therapy, were also essentially the same. We did not recognize differences in side effect profiles between the two treatment regimens with the exceptions of sleepiness and brain shrinkage estimated by CT scan, both of which were significantly milder in the low-dose than in the high dose group (P < 0.05). CONCLUSION: Unexpectedly, this prospective randomized controlled study demonstrated the dose of ACTH required for spasm cessation and disappearance of the hypsarrhythmic EEG pattern to be lower than previously believed. A low-dose regimen should thus be considered for CWS, and for SWS associated with significant cerebral atrophy. PMID- 10522524 TI - Emerging and entraining patterns of the sleep-wake rhythm in preterm and term infants. AB - It has been repeatedly reported that the sleep-wake rhythm in infants entrains around 3-4 months of age after a transient free-run rhythm. To clarify the emerging and entraining patterns of the sleep-wake rhythm, the sleep and wakefulness of 84 infants (44 preterm and 40 term infants) were longitudinally recorded at home for more than 16 weeks by the day-by-day plot method. Our results showed that the entrained sleep-wake rhythm emerged after transient manifestation of either ultradian or irregular sleep-wake patterns for 3-4 weeks in 75% of the infants. Only 7% of the infants showed a free-running sleep-wake rhythm before the entrainment. These facts suggest that most infants would be entrained to an ordinary daily schedule of mothers without expression of overt free-running rhythm of the biological clock. The mean age of the entrainment was 44.8 postconceptional weeks. There were no significant differences in either frequency of each pattern or the mean age of the entrainment, between preterm and term infants. In conclusion, the entrained sleep-wake rhythm emerges around 1 corrected month, after ultradian patterns in the majority of infants. PMID- 10522525 TI - Application of rapid random stimulation (RRS) to visual evoked potentials in children. AB - The present study was performed to examine the effects of regular (1 Hz) and modified rapid random stimulation (RRS) (6 and 12 Hz) on visual evoked potentials (VEPs), by simultaneously recording negative waves around 100 ms, wave IV latency, positive waves around 60 ms, wave III-latency, and amplitudes calculated from peak to peak, without causing impairment of visual acuity, in 44 patients aged 5-17 years. The wave IV-latencies of VEPs obtained by 6 and 12 Hz RRS were easily determined, and the latencies were not significantly changed compared to those obtained by previous 1 Hz regular stimulation. On the other hand, the amplitudes decreased in a frequency-dependent manner (1 < 6 < 12 Hz). These results were found to be similar in both preschool and school children. The examination time of VEPs determined by RRS is one-tenth shorter than that of 1 Hz regular stimulation. Thus, this method has the benefit of shortening the examination time, which decreases fatigue and inattention of the subjects, suggesting that modified RRS is a practically useful method for children. PMID- 10522526 TI - A clinical study of a large inbred kindred with pure familial spastic paraplegia. AB - BACKGROUND AND OBJECTIVES: Spastic paraplegia, an uncommon neurodegenerative disorder with phenotypic and genotypic heterogeneity, is mainly characterized by progressive weakness and spasticity of the lower limbs. We here present a large inbred family with pure familial spastic paraplegia outlining the clinical picture, the age at onset and the possible mode of inheritance. METHODS: This family was ascertained through two probands after which we structured an extended 10 generation pedigree. We examined 43 available family members to identify affected individuals based on fixed criteria. The clinical presentation and phenotypic specifics of this disease were studied in the affected members. We analyzed the possible mode of inheritance and the age at onset in this family. RESULTS: This 10 generation family reported about 50 affected individuals distributed over 5 consecutive generations. We identified 13 affected individuals out of the examined 43 and five individuals were classified as probably affected. We noticed the clinical specifics of this disorder in this family and identified some unique features not described in previous reports. DISCUSSION AND CONCLUSION: The mode of inheritance is either autosomal recessive or autosomal dominant with incomplete penetrance or variable expression of the age at onset. The age at onset seems to decrease with successive generations, either due to a true anticipatory phenomenon or to increased awareness. The unique features of this disorder in this family are discussed. PMID- 10522527 TI - Dipole analysis in a case with tumor-related epilepsy. AB - In order to evaluate the effectiveness of presurgical dipole analysis of interictal spikes as a non-invasive technique for the determination of epileptogenic area, we compared the results of this method with those of electrocorticography (ECoG) localization in the diagnosis of a patient with tumor related epilepsy. A preoperative MRI revealed a temporal lobe tumor on the right side. The individual dipoles estimated from the interictal spikes were located mainly in the anterolateral region of the right temporal lobe, although some were located in the mesial side. The ECoG recorded frequent spikes in the anterolateral region of the right temporal lobe consistent with the location estimated by dipole analysis. After surgery, the patient suffered from residual seizures. Therefore, the residual epileptogenic area was examined by dipole analysis using a four-layered head model instead of the previous three-layered head model. As a result, the dipole analysis was able to pinpoint the epileptic focus in the area directly adjacent to the resected area, and in the mesial temporal lobe. In conclusion, EEG dipole analysis appears to hold promise as a non-invasive presurgical evaluation technique for locating epileptogenic areas as well as for postsurgical evaluation of residual epileptic focus. PMID- 10522528 TI - Coexistence of oto-palato-digital syndrome type II and Arnold-Chiari I malformation in an infant. AB - A Taiwanese infant with clinically apparent oto-palato-digital syndrome type II had Arnold-Chiari I malformation. Arnold-Chiari I malformation has not been reported previously to occur in association with oto-palato-digital type II syndrome. The pathogenesis of both conditions has remain unclear although the Arnold-Cliari I malformation is most likely due to a developmental abnormality of improperly times or incomplete closure of the neural tube. We propose the physician who care for children with OPD type II must be aware of one more condition. PMID- 10522529 TI - Bibliography of congenital muscular dystrophies-cobblestone lissencephalies: Series II (1998). PMID- 10522530 TI - Proceedings of the second annual meeting of the Study Group on Seizures in Infancy and Early Childhood, Tokyo, April 29, 1999. PMID- 10522531 TI - Head-controlled laparoscopy: experiment, prototype, and preliminary results. AB - Depth perception is closely linked to the ability to explore. Previously described laboratory experiments showed the advantage of linking the motions of the laparoscope directly to the head movements of the surgeon. Additionally, it was found that the laparoscope should be mechanically supported instead of manually (by an assistant). This article describes three stages in the design and evaluation of a practical mechanism with which to link the motions of the laparoscope directly to the head movements of the surgeon. First, a description is given of the perceptual and technical considerations. Second, a laboratory experiment is described investigating the validity of the proposed suggestions. Last, a prototype was built that was tested in a practical setting. PMID- 10522532 TI - Quantitative analysis of the functionality and efficiency of three surgical dissection techniques: a time-motion analysis. AB - The increasing technological complexity of surgery demands objective evaluation of surgical techniques. In particular, alternatives for laparoscopic ligation, such as monopolar coagulation and the relatively new bipolar scissors combining dissection with coagulation, should be analyzed and compared. This study tests the efficacy of quantitative time-motion analysis in evaluating and comparing the functionality and efficiency of dissection and ligation techniques in a clinical setting. Standard dissection with ligation of vessels, bipolar scissors, and monopolar coagulation were consecutively applied to dissect 4 of the small bowel mesentery of pigs, in random order. All actions performed were recorded and analyzed, using a standard action list. The efficiency of each technique was expressed in mean dissection time and number of actions, and the safety in occurrence of complications and severity of microscopic damage. Time-motion analysis evaluated the efficiency objectively and reproducibly (ICC 0.98). Bipolar scissors were significantly more efficient (time 7 +/- 2 min, actions 129 +/- 33) than the standard technique (28 +/- 6, 771 +/- 185) and monopolar coagulation (14 +/- 5, 368 +/- 32) (p < 0.01). Furthermore, bipolar coagulation needed significantly less recoagulation of an oozing vessel (0.5% of the total dissected vessels) than did monopolar coagulation (10.4%), and the damaged zone was significantly smaller (p < 0.05). Significantly less time was spent waiting or exchanging instruments with bipolar scissors than with the standard technique (p < 0.05). This time-motion analysis objectively compared the efficiency and functionality of three surgical dissection techniques during clinical use. Bipolar scissors were more efficient than were both other techniques, and they coagulated vessels more safely than did monopolar coagulation. PMID- 10522533 TI - Endoscopic neck surgery: expanding horizons. AB - There have now been several attempts at neck exploration using minimally invasive surgery. These encouraging reports paved the way for the authors to attempt endoscopic neck surgery. Having the necessary technical expertise in minimally invasive surgery with an experience of more than 6000 laparoscopic procedures, they attempted endoscopic parathyroidectomy in three patients with hyperparathyroidism. Of these, two had a hyperfunctioning adenoma and one had parathyroid hyperplasia. The hyperfunctioning tissue was accurately localized using a 99Tc-thallium subtraction scan. It was possible to localize and dissect the parathyroid tissue in two of the three patients. One patient required an open hemithyroidectomy before the adenoma could be localized and excised. The total operative time averaged 113 min. The working space was found to be adequate provided good hemostasis was maintained. The magnification proved excellent in identifying and defining important neck structures. Sufficient mobilization of the lateral thyroid lobe for access to the tracheoesophageal groove was found to be technically very difficult. No subcutaneous emphysema was observed beyond the neck region, and none lasted beyond 24 h. Cosmesis was acceptable to both the patient and the surgeon. PMID- 10522534 TI - Laparoscopic cystogastrostomy for pancreatic pseudocyst is safe and effective. AB - Between March 1997 and March 1998, three consecutive patients underwent laparoscopic cystogastrostomy for persistent giant retrogastric pancreatic pseudocyst complicating an attack of acute pancreatitis. The mean cyst diameter was 15 +/- 1 cm (range 14-16). The procedure was performed with four trocars. The anterior wall of the stomach was opened longitudinally. The pseudocyst was entered through the posterior wall of the stomach. A cystogastrostomy was created by suturing the margins of the communication by interrupted nonabsorbable sutures. The mean operative time was 123 +/- 15 min, and there were no postoperative complications. The mean postoperative hospital stay was 4 +/- 1 days. Computed tomography demonstrated complete resolution of the pseudocyst. Laparoscopic cystogastrostomy represents a good therapeutic option for persistent retrogastric pancreatic pseudocyst. PMID- 10522535 TI - Laparoscopic management of gastric diverticula. AB - Gastric diverticular are rare and usually are diagnosed incidentally on radiographic examination. Surgical treatment, consisting of simple excision or inversion of the diverticulum, has been reserved for patients with proven symptoms or complications. These procedures have typically required laparotomy, but with the development of advanced endoscopic techniques, a minimally invasive approach may be appropriate. The authors report two cases of gastric diverticula managed laparoscopically and review the literature related to this entity. Between 1993 and 1996, two patients were evaluated for dyspepsia-like gastrointestinal complaints. Both patients were found to have a gastric diverticulum on a contrast study, and one diverticulum was also seen on upper endoscopy. Laparoscopic resection was undertaken in both cases. Flexible gastroscopy was performed intraoperatively to help localize the diverticulum, which was resected with an endoscopic stapling device. Nissen fundoplication was performed in conjunction with the diverticulectomy in the second patient for gastroesophageal reflux. Both procedures were completed laparoscopically without complications. The postoperative course was uneventful in both patients. At long term follow-up, the patients are asymptomatic. This experience indicates that laparoscopic resection of symptomatic gastric diverticula is a feasible alternative to laparotomy. A prospective analysis to verify the safety and efficacy of this procedure should be done. PMID- 10522536 TI - Laparoscopic excision of an omental cyst. AB - An omental cyst is a rare intra-abdominal tumor. The authors describe a case of omental cyst that was diagnosed correctly with abdominal magnetic resonance imaging (MRI) and successfully resected completely by use of minimal-access surgical techniques. A sagittal or coronal MRI view may precisely reveal the tumor position. The authors consider MRI to be very useful in the diagnosis of abdominal cystic masses. Laparoscopic surgical techniques are replacing or complementing open abdominal surgical procedures. PMID- 10522537 TI - Laparoscope-assisted minimally invasive treatment for choledochal cyst. AB - The principle of treatment of choledochal cysts is total cyst excision with hepaticojejunostomy because of the high rate of associated malignancy of the biliary system. The authors used a minimally invasive laparoscopic procedure to treat a patient with nonmalignant choledochal cyst. Although a large median laparotomy is usually used for cyst excision and hepaticoenterostomy, laparoscope assisted total cystectomy and hepaticojejunostomy were performed with minimal skin incision. To avoid gas embolism during dissection around the hepatic hilus the surgical procedure was divided into two stages: CO2 insufflation and abdominal lifting without pneumoperitoneum. This combination of procedures was as safe and technically adequate as conventional surgery. No abnormalities were observed in liver function, and the patient could sit up in bed the first day postoperatively. Thirteen days after surgery, he was discharged from the hospital uneventfully. PMID- 10522538 TI - Laparoscopic splenectomy at caesarean section. AB - The role of minimally invasive surgery continues to expand in all the specialties and subspecialties of surgery. At one time, obesity and pregnancy were considered a relative contraindication to laparoscopic surgery. With improved technology and skills, surgeons are able to operate on patients who were once considered to be at too high a risk for laparoscopic surgery. Herein, laparoscopic splenectomy performed at the time of Caesarean section is reported. This case demonstrates the safety and efficacy of laparoscopic splenectomy in a morbidly obese female in the immediate postpartum time. PMID- 10522539 TI - Creation of neovagina by laparoscopic Vecchietti operation. AB - The Vecchietti operation is a surgical technique that creates a neovagina by dilation in 7-9 days. The authors report a case of a 17-year-old woman with mullerian agenesis, Mayer-Rokitansky-Kuster-Hauser syndrome, in whom the Vecchietti operation was performed. A neovagina with a depth of 11 cm was created in 7 days. There were no complications, and the functional result was excellent. PMID- 10522540 TI - Laparoscopic resection of an insulinoma. AB - Surgical excision of an insulinoma results in dramatic reversal of hypoglycemic symptoms. Laparoscopic resection, if feasible, could reduce postoperative discomfort. Successful laparoscopic resection of an insulinoma is described. Preoperative studies failed to visualize the tumor; however, it was visualized and then imaged successfully during surgery. The operative approach described allows access to the entire pancreas and, therefore, may be useful for islet cell tumors at most sites in the gland. Based on this case and others in the literature, further efforts at laparoscopic pancreatic surgery are warranted. PMID- 10522541 TI - Laparoscopy and mesothelioma. AB - Malignant mesothelioma is a well-recognized long-term sequela of chronic asbestos exposure. Asbestos use in the United States began in the 1950s and was widespread until the mid-1970s. Although currently only 2.2 cases per million population per year are diagnosed, disease incidence is increasing because of the long latency of this neoplasm. A latency of 15-50 years means that a higher incidence of this neoplasm can be anticipated in the future. The authors report a patient with peritoneal mesothelioma and no known prior exposure to asbestos. The diagnosis was confirmed by exploratory laparoscopy, which entailed biopsies of the diaphragm and of the peritoneal and abdominal walls, and by cytologic evaluation of 700 ml ascitis fluid. At present, exploratory laparoscopy offers the quickest, safest, and least invasive way to confirm the clinical diagnosis of peritoneal malignant mesothelioma. PMID- 10522542 TI - Laparoscopic splenectomy in pregnancy. AB - A laparoscopic splenectomy during pregnancy is described in this case report. The operation took place at 18 weeks' gestation for life-threatening thrombocytopaenia secondary to antiphospholipid syndrome that had failed to respond to medical therapy. The patient made a full and rapid recovery and was delivered of a healthy baby girl at term. PMID- 10522543 TI - Recurrent common bile duct stones containing metallic clips following laparoscopic common bile duct exploration. AB - A case of recurrent common bile duct stones 2 years following laparoscopic cholecystectomy and laparoscopic common bile duct exploration in a 52-year-old man is reported. Surgical material as a nidus for recurrent stone formation has been reported and occurred in the present case. Factors influencing metallic clip migration after biliary surgery are discussed, with recommendations for decreasing recurrent stones caused by foreign material. PMID- 10522544 TI - Delayed presentation of spilled gallstones. AB - The authors describe a case of a cutaneous sinus at the umbilical port site following spillage of gallstones during laparoscopic cholecystectomy. The sinus tract was explored using a flexible cystoscope, the stones found within were removed, and the tract itself was curetted. The consequences of spillage of gallstones and its prevention are discussed. PMID- 10522545 TI - Laparoscopic-assisted Roux-en-Y gastric bypass. AB - Eight patients underwent laparoscopic Roux-en-Y gastric bypass from May 1998 to September 1998 in which a hand-assist technique was used. The operation consisted of a 7.5-cm periumbilical midline incision along with three trocars placed in the upper abdomen. The operative times ranged from 2.25 to 4.5 h. The average preoperative body mass index was 44 kg/m2. Three-month postoperative follow-up revealed an average weight loss of 59 lb. Cosmetic results to date have been excellent even when compared with those of a total laparoscopic operation. The hand-assist technique allows the surgeon to have more control over the most difficult part of the case, which is manipulation of the small bowel in a morbidly obese abdomen. PMID- 10522546 TI - Comparison of wounds created by non-bladed trocars and pyramidal tip trocars in the pig. AB - Wounds made by the Endopath nonbladed obturator, the Step trocar, and conventional pyramidal tip trocars were compared. The endopath nonbladed obturator and the Step trocar made wounds by separating tissue fibers, whereas the pyramidal tip trocar cut tissue fibers. The wounds of the Endopath nonbladed obturator and the Step trocar were similar in length but were narrower than wounds made by the pyramidal tip trocar. Further studies are needed to determine whether the wounds made by the Endopath nonbladed obturator and the Step trocar will have fewer complications than conventional pyramidal tip trocars. PMID- 10522547 TI - Mesh-plug hernioplasty. PMID- 10522548 TI - Author's reply to letter to the editor by Arthur P. Fine. Re: Laparoscopic resection of gastric diverticulum. PMID- 10522549 TI - New perspectives on iron: an introduction. AB - Iron is an essential nutritional element for all life forms. Iron plays critical roles in electron transport and cellular respiration, cell proliferation and differentiation, and regulation of gene expression. Two emerging new functions for iron are its necessary role in supporting transcription of certain key genes required for cell growth and function [eg, nitric oxide synthase, protein kinase C-beta, p21 (CIP1/WAF1)] and its complex role in hematopoietic cell differentiation. However, iron is also potentially deleterious. Reactive oxygen species generated by Fenton chemistry may contribute to major pathological processes such as cancer, atherosclerosis, and neurodegenerative diseases. Iron generated reactive oxygen species may also function in normal intracellular signaling. Therefore, roles of iron are both essential and extraordinarily diverse. This symposium explores this diversity by covering topics of iron absorption and transport, the regulation of gene expression by iron responsive proteins, the cellular biology of heme, hereditary hemochromatosis, and clinical use of serum transferrin receptor measurements. PMID- 10522550 TI - Iron absorption and transport. AB - Iron is vital for living organisms because it is essential for multiple metabolic processes to include oxygen transport, DNA synthesis, and electron transport. However, iron must be bound to proteins to prevent tissue damage from free radical formation. Thus, its concentrations in body organs must be regulated carefully. Intestinal absorption is the primary mechanism regulating iron concentrations in the body. Three pathways for intestinal iron uptake have been proposed and reported. These are the mobilferrin-integrin pathway, the divalent cation transporter 1 (DCT-1) [or natural resistance-associated macrophage protein (Nramp2)] pathway, and a separate pathway for uptake of heme by absorptive cells. Each of these pathways are incompletely described. However, studies with blocking antibodies, observations in rodents with disorders of iron metabolism, and studies in tissue culture cells suggest that the DCT-1 pathway is dominant in embryonic cells and is involved with cellular uptake of ferrous iron, whereas the mobilferrin-integrin pathway facilitates absorption of dietary inorganic ferric iron. Thus, there are separate pathways for cellular uptake of ferric and ferrous inorganic iron. Body iron can enter intestinal cells from plasma via basolateral membranes containing the classical transferrin receptor pathway with a high affinity for holotransferrin. This keeps the absorptive cell informed of the state of iron repletion of the host. Intestinal mucosal cell iron seems to exit the cell via a distinct apotransferrin receptor and a newly described protein named hephaestin. Unlike the absorptive surface of intestinal cells, most other cells possess transferrin receptors on their surfaces and the vast majority of iron entering these cells is transferrin associated. There seem to be 2 distinct pathways by which transferrin iron enters nonintestinal cells. In the classical clathrin-coated pitendosome pathway, iron accompanies transferrin into the cell to enter a vesicle, which releases the iron to the cytosol with acidification (high affinity, low capacity). Under physiological conditions, a second transferrin associated pathway (low affinity, high capacity) exists which has been named the transferrin receptor independent pathway (TRIP). How the TRIP delivers iron to cells is incompletely described. In addition, tissue culture studies show that nonintestinal cells can accept iron from soluble iron salts. This occurs via the mobilferrin-integrin and probably the DCT-1 pathways. Cellular uptake of iron from iron salts probably occurs in iron overloading disorders and may be responsible for free radical damage when the iron binding capacity of plasma is exceeded. Radioiron entering the cell via the heme and transferrin associated pathways can be found in isolates of mobilferrin/paraferritin and hemoglobin. This interaction probably occurs to permit NADPH dependent ferrireduction so iron can be used for synthesis of heme proteins. Production of heme from iron delivered via these routes indicates functional specificity for the pathways. PMID- 10522551 TI - Regulation of genes of iron metabolism by the iron-response proteins. AB - Iron is an essential nutrient, yet excess iron can be toxic to cells. The uptake of iron by mammalian cells is post-transcriptionally regulated by the interaction of iron-response proteins (IRP1 and IRP2) with iron-response elements (IREs) found in the mRNAs of genes of iron metabolism, such as ferritin, the transferrin receptor, erythroid aminolevulinic acid synthase, and mitochondrial aconitase. The IRPs are RNA binding proteins that bind to the IRE (found in the mRNAs of the regulated genes) in an iron- dependent manner. Binding of IRPs to the IREs leads to changes in the expression of the regulated genes and subsequent changes in the uptake, utilization, or storage of intracellular iron. Recent work has demonstrated that the binding of the IRPs to the IREs can also be modulated by changes in the redox state or oxidative stress level of the cell. These findings provide an important link between iron metabolism and states of oxidative stress. PMID- 10522552 TI - Cell biology of heme. AB - Heme is a complex of iron with protoporphyrin IX that is essential for the function of all aerobic cells. Heme serves as the prosthetic group of numerous hemoproteins (eg, hemoglobin, myoglobin, cytochromes, guanylate cyclase, and nitric oxide synthase) and plays an important role in controlling protein synthesis and cell differentiation. Cellular heme levels are tightly controlled; this is achieved by a fine balance between heme biosynthesis and catabolism by the enzyme heme oxygenase. On a per-cell basis, the rate of heme synthesis in the developing erythroid cells is at least 1 order of magnitude higher than in the liver, which is in turn the second most active heme producer in the organism. Differences in iron metabolism and in genes for 5-aminolevulinic acid synthase (ALA-S, the first enzyme in heme biosynthesis) are responsible for the differences in regulation and rates of heme synthesis in erythroid and nonerythroid cells. There are 2 different genes for ALA-S, one of which is expressed ubiquitously (ALA-S1), whereas the expression of the other (ALA-S2) is specific to erythroid cells. Because the 5'-untranslated region of the erythroid specific ALA-S2 mRNA contains the iron-responsive element, a cis-acting sequence responsible for translational induction of erythroid ALA-S2 by iron, the availability of iron controls protoporphyrin IX levels in hemoglobin-synthesizing cells. In nonerythroid cells, the rate-limiting step of heme production is catalyzed by ALA-S1, whose synthesis is feedback-inhibited by heme. On the other hand, in erythroid cells, heme does not inhibit either the activity or the synthesis of ALA-S but does inhibit cellular iron acquisition from transferrin without affecting its utilization for heme synthesis. This negative feedback is likely to explain the mechanism by which the availability of transferrin iron limits heme synthesis rate. Moreover, in erythroid cells heme seems to enhance globin gene transcription, is essential for globin translation, and supplies the prosthetic group for hemoglobin assembly. Heme may also be involved in the expression of other erythroid-specific proteins. Furthermore, heme seems to play a role in regulating either transcription, translation, processing, assembly, or stability of hemoproteins in nonerythroid cells. Heme oxygenase, which catalyzes heme degradation, seems to be an important enzymatic antioxidant system, probably by providing biliverdin, which is an antioxidant agent. PMID- 10522553 TI - New developments in hereditary hemochromatosis. AB - The iron content of the body is normally tightly controlled by regulation of iron absorption. In hereditary hemochromatosis, mutation of an HLA class 1 gene, designated HFE, results in excessive iron absorption. Over many years, accumulating iron produces tissue damage, most notably cirrhosis, cardiomyopathy, diabetes, and arthropathies. Hereditary hemochromatosis is the most common hereditary disease of Northern Europeans with a prevalence of approximately 5 per 1000. The most sensitive screening test for hemochromatosis is saturation of the transferrin with iron; a fasting value greater than 50% is strongly suggestive of the disease. Confirmation of increased iron storage can be achieved most readily by serial phlebotomy. We do not regard liver biopsy to be indicated, except in unusual circumstances. Early diagnosis and treatment by phlebotomy before tissue damage has occurred is essential, because life span seems to be normal in treated patients but markedly shortened in those who are not. Therefore, genetic counseling with evaluation of first-degree relatives is mandatory. PMID- 10522554 TI - The measurement of serum transferrin receptor. AB - The concentration of the soluble fragment of transferrin receptor in serum is an important new hematological parameter. Clinical and laboratory studies have shown that this serum form of the receptor reflects the total body mass of cellular transferrin receptor, 80% of which is contained in the erythroid marrow. The two disorders that result in an elevation in the serum transferrin receptor are anemias associated with enhanced erythropoiesis and tissue iron deficiency. The concentration of soluble transferrin receptor provides a useful quantitative measure of the erythroid marrow mass and thereby assists clinically in categorizing the type of anemia. The most important clinical use of the serum transferrin receptor is in determining the cause of iron deficient erythropoiesis (that is, identifying iron deficiency anemia whether it occurs alone or in the presence of the anemia of chronic disease). Present evidence supports the routine use of the serum transferrin receptor in the clinical evaluation of anemic patients. PMID- 10522555 TI - Blood volumes and renal function in overt and subclinical primary hypothyroidism. AB - INTRODUCTION: Thyroid dysfunction is associated with marked alterations in cardiovascular and renal functions. In hypothyroidism, myocardial contractility, cardiac output, and oxygen consumption are decreased, whereas peripheral resistance is increased. METHODS: We assessed blood volumes and effective renal plasma blood flow (ERPF) and glomerular filtration rate (GFR) in 17 patients with overt primary hypothyroidism and in 15 of these patients when in euthyroid state after substitutive therapy. We performed the same measurements in eight patients with subclinical hypothyroidism. RESULTS: In the hypothyroid state, the plasma volume measured by dilution of 125I-albumin (APV) was higher than the calculated plasma volume (CPV) from packed red cell mass, suggesting an extravascular escape of albumin. After substitutive therapy, the CPV showed a statistical increase (P < 0.05), whereas APV remained unchanged. Both ERPF and GFR increased after thyroxine therapy (p < 0.05). In the subclinical group, blood volumes and renal function were similar to those found in the other group of patients when in the euthyroid state. CONCLUSIONS: We conclude that in primary hypothyroidism, ERPF and GFR are low, but that these values improve with substitutive therapy. CPV is a better index of the current plasma volume than APV. The difference between these two parameters suggests that the escape of albumin into the extravascular space in primary hypothyroidism is terminated by treatment. There are no clear abnormalities either in blood volumes or in renal function in subclinical hypothyroidism. PMID- 10522556 TI - Use of coronary flow reserve to evaluate the physiologic significance of coronary artery disease. PMID- 10522557 TI - Ascites and pleural effusion secondary to extramedullary hematopoiesis. AB - Extramedullary hematopoiesis in the pleura and peritoneum is rare. It is usually asymptomatic and generally is diagnosed on post mortem examination. Herein we describe a 33-year-old woman with long-standing myelofibrosis who presented with symptomatic ascites and pleural effusion. After complete evaluation, these were found to have been caused by extramedullary hematopoietic implants to the pleura and peritoneum. The pleural effusion responded to low-dose radiotherapy. PMID- 10522558 TI - Assessment of ultra low frequency band power of heart rate variability: validation of alternative methods. AB - Alternative methods for assessing ULF spectral power using data from commercial Holter analysers were studied. Different heuristics for ULF calculation were compared with standard research software-based determination of ULF. SETTING: University Hospital. PATIENTS: 43 patients in NYHA classes I-IV heart failure and seven normals of similar ages. METHODS: SDNN, SDANN, ULF, VLF, LF, HF calculated from 24 h Holter monitoring using Oxford scanner software (method 1). ULF power also calculated by subtracting the sum of VLF. LF and HF powers obtained from the Holter scanner from the total variance (method 2) from 2 x ln(SDANN) (method 3), and by performing a standard, research-quality 24-h EFT analysis on the beat files (standard). Results of methods 1-3 were compared with standard using two way ANOVA with repeated measures, regression analysis and a graphical technique. RESULTS: ULF calculated by method 1 correlated r=0.66 with standard but means differed substantially. In contrast, ULF calculated by method 2 correlated r=0.99 with standard with no significant differences between means. ULF calculated from SDANN (method 3) correlated r=0.983 with standard but means, while similar, were significantly lower (P=0.005). CONCLUSION: ULF reported by commercial HOLTER software is not equivalent to ULF power derived from 24 h FFT analysis. ULF calculated by method 2 can be considered equivalent to the ULF derived by standard 24-h FFT. ULF estimated by method 3 offers direct ULF power estimation from a temporal measure of HRV and can be useful when spectral values are not available. PMID- 10522559 TI - A comparison of the cold pressor test and the diving test or coronary and systemic hemodynamics in patients with and without coronary artery disease. PMID- 10522560 TI - Is hyperuricemia a risk factor of stroke and coronary heart disease among Africans? AB - BACKGROUND: Uric acid stabilizes platelet aggregation and enhances thrombotic tendency. OBJECTIVE: To examine the relationship between raised serum uric acid and subsequent cardiovascular events (mortality, myocardial infarction, stroke). METHODS: This is a longitudinal study in a small random number (418) of patients in Kinshasa, Congo. In this hospital-based study, uricemia was determined with respect to gender, obesity and hypertension as well correlated with traditional cardiovascular risk factors. A univariate regression model was used to investigate the association of serum uric acid with the incidence of mortality, stroke and myocardial infarction. RESULTS: Serum uric acid was higher (P<0.05) in obese women and men as well in hypertensives, than in their controls. The significant (P<0.05 and 0.001) highest frequency of hyperuricemia was observed in both diabetic and hypertensive patients. Blood pressure (BP) was higher (P<0.001) in patients with hyperuricemia than in those with normal serum uric acid. Serum uric acid was significantly correlated with body weight, BP, glucose, fibrinogen, urea, creatinin and total cholesterol. In men, hyperuricemia was significantly (P<0.01) associated with a twofold increased risk of both myocardial infarction and stroke incidence. However, hyperuricemia was significantly related to a double risk of all mortality and stroke onset. CONCLUSION: Our results indicate that hyperuricemia among african patients is a strong predictor of myocardial infarction in men, stroke in both sexes and all causes of mortality in women. PMID- 10522561 TI - Effect of exercise on left and right ventricular ejection fraction and wall motion. AB - OBJECTIVE: We evaluated the diagnostic value of response of left and right ventricular ejection fraction and wall motion to exercise using electron beam computed tomography. METHODS AND RESULTS: We attempted to determine the value of exercise electron beam computed tomography for detecting coronary artery disease, including evaluation of the right ventricular ejection fraction and wall motion abnormalities. A study of 35 patients undergoing electron beam tomography exercise cine studies and coronary artery angiography for the evaluation of chest pain was performed. Of the 18 patients with significant coronary disease (> or = 50% luminal diameter stenosis in at least one coronary artery), 17 (94%) had failure to increase global left ventricular ejection fraction with exercise. Fourteen of 18 (78%) developed a wall motion abnormality during peak exercise, and eight (44%) developed a regional right ventricular wall motion abnormality during peak exercise. Of the 17 patients without obstructive disease, 14 (82%) had a increase in ejection fraction > or = 5% and none had an abnormal response in left ventricular wall motion during peak exercise (specificity = 100%). The change in right ventricular ejection fraction with exercise was not a significant predictor of obstructive coronary disease in this study (P=NS). Using different criteria during stress to predict coronary disease, the accuracy was 89% (31/35) using an increase of <5% in ejection fraction, 89% (31/35) using the development of a new or worsened wall motion abnormality, and 91% (32/35) using both left ventricular criteria. CONCLUSION: Our study suggests that exercise electron beam computed tomography appears to be a useful tool for the detection of coronary disease. A increase of <5% in ejection fraction and abnormal left ventricular response to exercise were important predictors, while the exercise induced changes of right ventricular ejection fraction was not a significant predictor of obstructive disease. Both left and right ventricular wall motion abnormalities are useful and important parameters in identifying patients with obstructive disease from those with normal coronary arteries. PMID- 10522562 TI - Right bundle branch block: varying electrocardiographic patterns. Aetiological correlation, mechanisms and electrophysiology. AB - Ten dissimilar electrocardiographic (ECG) patterns associated with right bundle branch block (RBBB) are presented. Electrophysiologic basis of the changes is discussed and possible causes for such diversity outlined. We have not found any aetiological association to this variation. The morphological diversity in RBBB patterns is likely to be related to multiple factors--site of block, nature of defect (functional, necrosis, fibrosis), degree of conduction delay, and associated pathologies with their own ECG patterns. Distinguishing RBBB from a normal ECG-variant like rsr' is particularly important when associated with left hemiblocks as the latter situation warrants extensive cardiac evaluation. PMID- 10522563 TI - Circadian patterns of heart rate variability in normals, chronic stable angina and diabetes mellitus. AB - The purpose of our investigation was to compare circadian patterns of heart rate variability as assessed by 24-h ambulatory electrocardiographic (ECG) recordings in normal subjects, chronic stable angina, and Type 1 diabetes mellitus. The study population consisted of three groups: 12 normal subjects, 23 chronic angina patients, and 23 Type 1 diabetics. For purposes of analyzing circadian variation, the ECG recordings were divided into daytime (08:00-00.00 h) and night-time (00:00-08:00 h) periods. Analysis was performed for all time and frequency domain measures of heart rate variability, attempting to identify differences in day-to night variability among these three groups. All time domain parameters except standard deviation of all 5-min mean RR intervals, and all frequency domain indices maintain significant circadian variations (P<0.0001), with the greatest day to night variation seen in normals, the least in diabetics, and intermediate values in chronic angina. These changes in heart rate variability circadian rhythms reflect significant reductions in cardiac parasympathetic activity with the most marked reduction in nocturnal vagal activity. Given the circadian pattern of myocardial ischemia and infarction, these data suggest that quantification of the magnitude of circadian variation in heart rate variability may have the potential to further risk stratify chronic angina and diabetes for future cardiac events. PMID- 10522564 TI - Significance of exercise-induced simultaneous ST-segment changes in lead aVR and V5. AB - This study was undertaken to investigate the ability of the exercise-induced ST depression in lead V5 and concomitant ST elevation in lead aVR for the identification of the significantly narrowed coronary artery in patients with single vessel disease. We studied 229 consecutive patients who developed the aforementioned exercise-induced electrocardiographic changes. All underwent Thallium-201 scintigraphy and coronary arteriography. Patients were divided into three groups. In group A, 58 patients with ST depression in V5 and ST elevation in aVR, in group B 149 patients with ST depression in V5 without ST elevation in aVR, and in group C 22 patients with ST elevation in aVR without ST depression in V5 induced with exercise, were included. In group A, 81% of the patients while in group B, 29% and in group C only 18% of the patients had left anterior descending artery disease. According to Thallium-201 scintigraphy, 80% of the group A, 27% of the group B and 12% of the group C patients developed myocardial ischemia in areas supplied by the left anterior descending artery. Thus, exercise-induced ST depression in V5 and concomitant ST elevation in aVR, may detect left anterior descending artery significant stenosis in patients with single vessel disease. PMID- 10522565 TI - Left ventricular geometry and function in patients with aortic stenosis: gender differences. AB - BACKGROUND: Gender differences in cardiac size have been described in normal and pathological conditions in human and animals. Sex determination of a pattern of hypertrophy as a response to pressure overload has not been extensively evaluated and is still poorly understood in humans. METHODS AND RESULTS: To investigate the influence of gender in the left ventricle remodelling and preservation of the left ventricle function 195 adults (140 men and 55 women) with isolated aortic stenosis were evaluated. The mean age was 52 +/- 11 years for men and 53 +/- 13 years for women. All the patients had similar degree of aortic stenosis finally treated with valve replacement, similar clinical status and no signs of coronary artery disease in coronary angiograms. On echocardiography the left ventricle of women had a smaller the end systolic (30.5 +/- 7.8 vs. 39.4 +/- 11.2, P<0.001) and the end diastolic (49.4 +/- 9 vs. 57.3 +/- 11, P<0.001) chamber size. The female left ventricle generated a higher relative wall thickness (0.65 +/- 0.21 vs. 0.52 +/- 0.12, P<0.01), a greater fractional shortening (35.3 +/- 8.5 vs. 32.0 +/- 9.0, P<0.01) and a higher ejection fraction (64.4 +/- 12.7 vs. 57.5 +/- 14.6, P<0.001). The left ventricle posterior wall thickness and the septal thickness indexes were similar in both groups. There were also significant differences between the two groups in the left ventricle mass index. CONCLUSIONS: Gender has an important influence on the left ventricle adaptation pattern to pressure overload due to aortic stenosis. Women developed a greater degree of left ventricle hypertrophy documented as changes in left ventricle geometry (increased relative wall thickness, left ventricular mass) and left ventricle function (fractional shortening and ejection fraction). PMID- 10522566 TI - Transvenous internal cardioversion for atrial fibrillation: a randomized study on defibrillation threshold and tolerability of asymmetrical compared with symmetrical shocks. AB - The aim of the study was to compare, according to a randomized cross-over design, two different biphasic waveforms (6.5/2.5 ms and 3.0/3.0 ms phases duration, respectively) for low energy internal atrial cardioversion with regard to energy requirements for cardioversion and shock induced discomfort. METHODS: Nineteen patients with chronic persistent atrial fibrillation (AF)(mean duration 16+/-20 months) were submitted to internal atrial cardioversion (shock delivery between catheters in right atrium and coronary sinus, respectively) and were randomly allocated to baseline cardioversion with an asymmetrical biphasic shock (6.5/2.5 ms) or with a symmetrical biphasic shock (3.0/3.0 ms), according to a step up protocol. After baseline cardioversion, a sustained AF was reinduced and the patients crossed to the alternative waveform. The procedure was performed without routine administration of sedatives and shock induced discomfort was monitored by a subjective score (1 to 5). Sedatives or anesthetics were administered at patient's request. RESULTS: The procedure was effective in all the patients and was performed without need for sedatives/anesthetics in 17/19 patients (89%). Leading edge voltage of effective shocks resulted lower for asymmetrical shocks compared to symmetrical shocks (290+/-76 vs. 337+/-104 V, P<0.001) with no statistically significant differences in delivered energy (7.74+/-4.25 vs. 8.65+/ 5.94 J). Moreover shock induced discomfort resulted lower for asymmetrical shocks compared to symmetrical (pain score=4.18+/-0.73 vs. 4.59+/-0.62, P<0.02). Shock impedence of effective shocks was 59+/-10 ohms for both waveforms. No significant complications occurred during the procedure and no ventricular arrhythmia was observed after atrial cardioversion. Transient bradycardia requiring support ventricular pacing was observed in one patient. CONCLUSIONS: Delivery of biphasic asymmetrical shocks (6.5/2.5 ms) results in lower leading edge voltage of effective shocks and better patients tolerability compared with conventional biphasic symmetrical shocks (3.0/3.0 ms). These findings are of interest both for transvenous internal cardioversion of chronic persistent AF and for implantable atrial defibrillators. PMID- 10522567 TI - Limited internal shocks for atrial fibrillation refractory to external cardioversion. AB - We investigated the feasibility and long-term results of low-energy internal defibrillation using a limited number of shocks in patients with persistent atrial fibrillation resistant to external cardioversion. A relatively high number of shocks of lower energy are usually required in those cases and can be poorly tolerated. METHODS AND RESULTS: Twenty-five patients with persistent atrial fibrillation underwent internal defibrillation, using biphasic R wave synchronous shocks between two catheters in the high right atrium and the coronary sinus. Conversion to sinus rhythm was obtained in all patients, with a median of two shocks. Early recurrence of atrial fibrillation (AF) occurred in eight cases (32%). Seven patients (41%) out of 17 discharged in sinus rhythm remained free of AF after a median follow-up of 8.9 months. Severe mitral insufficiency (P=0.05) and low left ventricle ejection fraction (P=0.002) were correlated with earlier recurrence. Amiodarone significantly favored (P=0.019) maintenance of sinus rhythm. CONCLUSION: Internal defibrillation using a limited number of shocks equal to or less than 30 Joules is effective in terminating refractory atrial fibrillation and could be more acceptable for patients. However, the recurrence rate remains high, particularly in patients with severe mitral insufficiency or poor ventricular function. Amiodarone delays recurrences of atrial fibrillation. PMID- 10522568 TI - Anthropometric evaluation of cachexia in chronic congestive heart failure: the role of tricuspid regurgitation. AB - Cardiac cachexia has recently been identified as an independent risk factor for mortality in chronic congestive heart failure. The aims of our study were to further identify the clinical or biochemical predictors or correlates of the cachexia, and to quantitate the magnitude of wasting. We undertook an anthropometric comparison of 30 patients with congestive heart failure, aged 56 (13) years, with ten age- and sex-matched healthy volunteers and 16 patients with essential hypertension. In comparison to the healthy volunteers, the heart failure patients exhibited a trend towards a lower body mass index, 21 (2.7) versus 23 (3.8) kg/m2, the 95% confidence interval for the difference being -0.54 to 5.4. However, the mid-upper arm circumference, of 24 (3.8) cm in the heart failure patients, was significantly (P<0.02) lower than the 27 (2.0) cm in the healthy volunteer group, with a 95% confidence interval for the difference being 1.18 to 4.82 cm. The triceps, mid-thigh, scapula and abdominal skinfold thicknesses were separately and significantly (P<0.05) diminished in the heart failure patients compared to the healthy controls. The sum of the four skin fold thicknesses, with a value of 68 (13) mm in the healthy volunteers, was highly significantly greater (P<0.001) than the value of 35.6 (9) mm in the heart failure patients. The 95% confidence interval for this difference was 22.7 to 41.3 mm. The patients with essential hypertension differed significantly from the heart failure patients in all of these parameters (P<0.01), but were not statistically different from the healthy controls in the anthropometric parameters. Among the heart failure patients, those with tricuspid regurgitation (n = 12) had a worse clinical, biochemical and cachexia profile compared to patients without the tricuspid regurgitation (n = 18). The values (tricuspid regurgitation versus no regurgitation) were New York Heart Association Class, 3.5 (0.65) versus 2.7 (0.75), P<0.01; ejection fraction of 34 (9) versus 43 (13)%, not significant; greater hepatomegaly of 159 (31) versus 135 (29) mm, P<0.05; more severe hypoalbuminemia, 24.5 (2.7) versus 28.5 (6.8) g/l, P<0.05; and worse hyponatremia, 128 (4) versus 133 (5) mmol/l, P<0.05. The tricuspid regurgitation group had a significantly more severe reduction in abdominal and scapula skin fold thickness (P<0.01) than that found in patients without tricuspid regurgitation. The sum of the four skin fold thicknesses was significantly lower (P<0.05) in tricuspid regurgitation, 30.9 (8) mm, than in heart failure without associated regurgitation, 38.0 (9.6). The 95% confidence interval for the difference was 0.8 to 13.4 mm. It is concluded that significant diminution of muscle bulk and subcutaneous fat occurs in chronic heart failure. Tricuspid regurgitation may be an accentuating and accelerating risk factor for cardiac cachexia, on account of a greater hypoalbuminemia and hyponatremia, which, presumably, results from the associated protein-losing enteropathy. PMID- 10522569 TI - Manual assembly of a retrieval system for inadvertently retained objects in the coronary circulation. PMID- 10522570 TI - Spontaneous coronary dissection by coronary stenting. PMID- 10522571 TI - Isolated infundibuloarterial inversion and fifth aortic arch in an infant: a newly recognized cardiovascular phenotypes with chromosome 22q11 deletion. PMID- 10522572 TI - Recognition of Berry syndrome in a 4-day-old neonate by echocardiography and transvenous angiocardiography. PMID- 10522573 TI - Ganciclovir resistance in a heart transplant recipient infected by cytomegalovirus. PMID- 10522574 TI - Recurrent acute coronary events in a patient with primary antiphospholipid syndrome: successful management with intracoronary stenting. AB - Patients with Antiphospholipid syndrome usually present with recurrent deep vein thrombosis, pulmonary thromboembolism and thromboembolic stroke. Recurrent coronary events, though reported, are rare. We describe an unusual case of Antiphospholipid syndrome who presented with recurrent acute ischaemic events in two different coronary territories, who was managed successfully with intracoronary stenting. PMID- 10522575 TI - Consequences of inadequate Canadian physician resource planning. PMID- 10522576 TI - Femoral nerve block and ketorolac in patients undergoing anterior cruciate ligament reconstruction. AB - PURPOSE: The primary objective was to evaluate the analgesic effectiveness of femoral nerve block and ketorolac following ACL reconstruction. The secondary objective was to examine their effects on recovery milestones. METHODS: Prior to standard general anesthesia, 90 patients were randomized into three groups of preoperative treatment: 1) femoral nerve block (15 mL bupivacaine 0.5%) and 1 mL normal saline i.v. (FNB group); 2) placebo femoral nerve block (15 mL normal saline) and 30 mg (1 mL) ketorolac i.v. (KT group); 3) placebo femoral nerve block (15 mL normal saline) and 1 mL normal saline i.v. (PL group). Postoperatively, pain was assessed by visual analogue score, demand and consumption of morphine via patient-controlled analgesia pump. The times for patients to tolerate oral fluid, food, sit up, ambulate and void were also noted. RESULTS: Morphine consumption within one hour, three hours and until POD 1 in the FNB group was lower than the PL group (7 +/- 6, 11 +/- 9, 27 +/- 23 mg vs 13 +/- 5, 20 +/- 9, 49 +/- 28 mg respectively), whereas only that within one hour in the KT group was lower than the PL group. Pain score was lower in FNB and KT groups in the first postoperative hour than in the PL group (P < 0.05). There were no differences among the three groups in the times to meet recovery milestone and discharge criteria. CONCLUSION: Femoral nerve block provides superior analgesia than placebo for ACL reconstruction but was insufficient to facilitate early recovery. PMID- 10522577 TI - Tourniquet inflation during arthroscopic knee ligament surgery does not increase postoperative pain. AB - PURPOSE: A double-blind clinical trial was conducted to determine the effect of inflation of a thigh tourniquet during anterior cruciate ligament repair on arthroscopic visibility, duration of procedure, postoperative pain and opioid consumption. METHODS: Thirty patients were randomly allocated into two groups; Group I had the thigh tourniquet inflated during surgery whereas the tourniquet was not inflated in Group II patients. All patients received standardized general anesthesia and postoperative pain management. Supplemental analgesia was provided with i.v. morphine via a patient-controlled analgesia (PCA) apparatus. Verbal pain rating scores (0-10) were obtained after surgery. RESULTS: Arthroscopic visibility was impaired in Group II patients (P < 0.0001), but this was ameliorated by increased irrigation flow or addition of epinephrine. Duration of surgery was similar in both groups. There was no difference between groups in postoperative morphine consumption (9.8 +/- 7.1 mg in Group I vs 11.4 +/- 10.2 mg in Group II) or in postoperative pain scores between groups. CONCLUSION: Inflation of a thigh tourniquet did not result in increased pain or opioid consumption after arthroscopic ACL surgery. Arthroscopic visibility was somewhat impaired in some patients without the use of tourniquet. Finally, the duration of the surgical procedure was not increased in patients where the tourniquet was not inflated during the ACL repair. PMID- 10522578 TI - Saline volume and local anesthetic concentration modify the spread of epidural anesthesia. AB - PURPOSE: To examine the effects of the volume of saline and the concentration of local anesthetic on the quality of anesthetic level. METHODS: One hundred and fifty two patients received thoracic epidural anesthesia were allocated into two groups; mepivacaine 1% (75 patients) and 1.5% (77 patients). Each group was randomly divided into three subgroups depending on epidural saline volumes of 1 ml, 5 ml, or 10 ml. Fifteen minutes after the injection of 10 ml mepivacaine, the dermatome levels of hypesthesia to cold and pinprick were determined by an individual blinded to the saline volume. RESULTS: The number of spinal segments with hypesthesia to cold in the three subgroups in the mepivacaine 1% group were 12.5 [6-20], 13 [8.5-20.5] and 12.5 [6.5-22], respectively (median [range]). The segments in the mepivacaine 1.5% group were 12 [7-18.5], 14 [8.5-19]* and 15 [6 23]*, respectively (*P < 0.05 vs 1-ml group). The number of spinal segments with hypesthesia for pinprick in the three subgroups in the 1% mepivacaine group were 10.5 [2-22], 10.5 [4-17] and 11 [4-19], respectively. The segments in the mepivacaine 1.5% group were 12 [7.5-16], 12 [7.5-17] and 11.5 [5-22.5], respectively. Saline volume did not alter the anesthetic level of the mepivacaine 1%, although it did affect the anesthetic spread of the mepivacaine 1.5%. In both groups, a differential nerve block was elicited in the 5 ml and 10 ml saline subgroups. CONCLUSION: When a large volume of saline is administered prior to local anesthetic, more differential blockade and a greater extent of anesthesia may be elicited. PMID- 10522579 TI - Saline-anesthetic interval and the spread of epidural anesthesia. AB - PURPOSE: To examine the effect of modifying the interval between administration of saline used during the loss of resistance (LOR) method and local anesthetic on epidural anesthetic level and its quality. METHODS: Seventy-three patients who received thoracic epidural anesthesia were randomly allocated into three groups; the 2, 5 and 10 min groups, according to the interval between the administration of saline and 8 ml mepivacaine 1.5%. Fifteen minutes after the mepivacaine injection, the dermatome level of hypesthesia was determined by an individual blinded to the interval. RESULTS: When the saline-anesthetic interval was prolonged, the hypesthetic levels for coldness and pinprick were decreased. The number of spinal segments with hypesthesia for coldness were 15 [12-20]#, 12.5 [10.5-22.5]## and 10.5 [6.5-15.5]### in the 2, 5 and 10 min groups, respectively (median [range], # P < 0.05 vs the 5 min group, ## P < 0.05 vs the 10 min group, ### P < 0.05 vs the 2 min group). The number of spinal segments with hypesthesia for pinprick were 13.5 [11-18]#, 11 [7.5-20.5]## and 10 [5.5-13]### in the 2, 5 and 10 min groups, respectively. There were differences in all groups between the number of segments with hypesthesia for coldness and pinprick elicited. CONCLUSION: The interval between the administration of saline and local anesthetic alters the anesthetic level and quality of epidural analgesia. PMID- 10522580 TI - Non-alkalinized and alkalinized 2-chloroprocaine vs lidocaine for intravenous regional anesthesia during outpatient hand surgery. AB - PURPOSE: Chloroprocaine should be an ideal agent for intravenous regional anesthesia (IVRA) because of its rapid onset and ester hydrolysis. Raising the pH of local anesthetics may increase the speed of onset and the intensity of nerve blocks. We compared plain and alkalinized 2-chloroprocaine 0.5% with lidocaine for IVRA. METHODS: In two separate double-blind studies, 78 patients scheduled for daycare hand surgery were randomized to receive 40 mL plain 2-chloroprocaine 0.5%, alkalinized 2-chloroprocaine 0.5% or lidocaine 0.5% for IVRA. Time to sensory and motor block, need for supplemental analgesia, and side effects were compared. RESULTS: There was no difference in time to sensory or motor block in either group. Patients who received plain chloroprocaine required more supplemental opioid and had a higher incidence of metallic taste and of hives than patients who received lidocaine (P < 0.05). Comparing alkalinized chloroprocaine with lidocaine, there was no difference found with respect to opioid supplementation, CNS side effects, or incidence of hives. CONCLUSION: In conclusion, alkalinized chloroprocaine was found to be an effective agent for IVRA but no benefit over lidocaine was detected. Plain chloroprocaine for IVRA produced more minor side effects than lidocaine. PMID- 10522581 TI - A multicentre trial of ropivacaine 7.5 mg x ml(-1) vs bupivacaine 5 mg x ml(-1) for supra clavicular brachial plexus anesthesia. AB - PURPOSE: To compare the efficacy of ropivacaine 7.5 mg x ml(-1) with bupivacaine 5.0 mg x ml(-1) for subclavian perivascular brachial plexus block. METHODS: After informed consent, 104 ASA I-III adults participated in a randomized, double blind, multi-center trial to receive 30 ml of either ropivacaine 7.5 mg x ml(-1) or bupivacaine 5.0 mg x ml(-1) for subclavian perivascular brachial plexus block prior to upper limb surgery. Onset and duration of sensory and motor block in the distribution of the axillary, median, musculo-cutaneous, radial and ulnar nerves were assessed. RESULTS: Onset times and duration of sensory and motor block were similar between groups. Mean duration of analgesia for the five nerves was between 11.3 and 14.3 hr with ropivacaine and between 10.3 and 17.1 hr with bupivacaine. Quality of muscle relaxation judged as excellent by the investigators was not significantly different (ropivacaine - 35/49, bupivacaine - 30/49). The median time to first request for analgesia was comparable between the two groups (11-12 hr). One patient developed a grand mal seizure shortly after receiving bupivacaine and recovered consciousness within 30 min. There were no serious adverse events in the ropivacaine group. CONCLUSIONS: Thirty ml ropivacaine 7.5 mg x ml(-1) (225 mg) produced effective and well tolerated brachial plexus block of long duration by the subclavian perivascular route. In this study, the results were similar to those of 30 ml bupivacaine 5.0 mg x ml( 1). PMID- 10522582 TI - Combined diltiazem and lidocaine reduces cardiovascular responses to tracheal extubation and anesthesia emergence in hypertensive patients. AB - PURPOSE: Hypertensive patients exhibit exaggerated cardiovascular responses to tracheal extubation. This study was undertaken to compare the efficacy of combined diltiazem and lidocaine with each drug alone in suppressing the hemodynamic changes during tracheal extubation. METHODS: Sixty hypertensive patients (ASA II), defined as systolic blood pressure > 160 mmHg and/or diastolic blood pressure > 95 mmHg (WHO), undergoing elective orthopedic surgery received, in a randomized, double-blind manner, 0.2 mg x kg(-1) diltiazem, 1.0 mg x kg(-1) lidocaine, or 0.2 mg x kg(-1) diltiazem plus 1.0 mg x kg(-1) lidocaine (n=20 of each) i.v. before tracheal extubation. Changes in heart rate (HR), mean arterial pressure (MAP) and rate-pressure product (RPP) were measured before and after tracheal extubation. RESULTS: Hemodynamic changes during tracheal extubation were less in patients receiving diltiazem plus lidocaine than in those receiving diltiazem or lidocaine as a sole medicine (RPP; 10322 +/- 1674 (combined) vs 11532 +/- 1802 (diltiazem), 15388 +/- 2050 (lidocaine), mean +/- SD, P < 0.05). CONCLUSION: Combined diltiazem and lidocaine is more effective prophylaxis than diltiazem or lidocaine alone for attenuating the cardiovascular responses to tracheal extubation and emergence from anesthesia in hypertensive patients. PMID- 10522583 TI - Target controlled infusion of ketamine as analgesia for TIVA with propofol. AB - PURPOSE: To determine the accuracy of a target controlled infusion system for ketamine and to assess its suitability for the provision of analgesia when used in conjunction with a propofol infusion in spontaneously breathing patients. METHODS: Nineteen, adult, ASA I-III patients scheduled for elective surgery were studied. After premedication with 20 mg temazepam an appropriate plasma concentration of ketamine was selected and, when the target controlled infusion (TCI) system indicated that this had been achieved, anesthesia was induced and maintained using a propofol infusion. The plasma ketamine concentration was measured at predetermined intervals and cardiovascular and respiratory parameters recorded at 10 min intervals. Patients were reviewed in recovery and 24 hr postoperatively to assess the adequacy of their recovery and the presence of any undesirable side effects. RESULTS: The TCI system had a median performance error against predicted plasma concentrations of 18.9% (SE 2.5%) and a median absolute performance error of 23.3% (SE 2.3%). Divergence was 20.3% (SE 30.1%) and wobble was 12.9% (SE 2.1%). There was a mean decrease in arterial pressure of 6.4% (SD 19.7%) and a mean increase in heart rate of 4.3% (SD 17.4%). Little respiratory depression occurred and all patients made a rapid postoperative recovery with none describing unpleasant dreams or hallucinations. CONCLUSION: The TCI system provided a clinically acceptable degree of control of the plasma ketamine concentration although some further improvement should be possible by amending the pharmacokinetic model. Clinically the combination with a propofol infusion proved to be a satisfactory anesthetic technique. PMID- 10522584 TI - Canadian anesthesia physician resources: 1996 and beyond. AB - PURPOSE: To report physician resource information from the 1996 national anesthesia physician and residency programme surveys in Canada. The findings are used to discuss the potential effects on availability of future specialist anesthesia services in Canada. METHODS: Twenty-six hundred and ninety-three physicians (2,206 specialists, 487 family physicians) providing anesthesia services were surveyed. Information on demographics and patterns of clinical practice were sought. Anesthesia programme directors provided trainee information. Projections of the potential number of practicing anesthesiologists to 2026 were made based on the number of available training positions and age distribution of anesthesiologists. RESULTS: There was a 58.3% response rate to the national survey. Since 1986 there has been a 10% increase in the number of specialist anesthesiologists. Marked regional variations in age distribution and changes in the number of specialist anesthesiologists were noted. Most specialists remain in the region or province of postgraduate training. Sixty percent of specialists were either re-entry trainees or international medical graduates. Changes in anesthesia practice patterns have resulted in 40% of the anesthesiologist's work now occurring outside of the operating room. Anesthesia training positions have decreased by at least 15%. The population of Canada is projected to increase by 33.8% between 1996 and 2026. If current government and position allocation policies continue, it is projected there will be 0% increase in the number of specialist anesthesiologists over the same time period. CONCLUSIONS: Changes in anesthesia practices have exacerbated the current shortages of anesthesiologists. These shortages will worsen if the number of, and restrictions to, available residency positions is unchanged. PMID- 10522585 TI - Management of an infant with diffuse bullous pulmonary lesions using high frequency oscillatory ventilation. AB - PURPOSE: To describe the anesthetic and ventilatory management of an infant with diffuse pulmonary bullous lesions. CLINICAL FEATURES: Four successive operations were scheduled for an infant with diffuse pulmonary bullous lesions. At the age of seven weeks, conventional positive pressure ventilation during laparotomy for intestinal occlusion led to arterial desaturation. This was corrected by returning to spontaneous respiration and deep inhalation anesthesia with halothane. Based on our ICU experience and due to a potential impaired oxygenation during conventional ventilation, we chose high-frequency oscillatory ventilation (HFOV) for bilateral sequential thoracotomies for bullectomies at the age of five months. We elected the same ventilatory mode for laparotomy for intestinal obstruction secondary to a polyp at the age of six months. This ventilatory mode was combined with total intravenous anesthesia and epidural analgesia and provided optimal oxygenation and ventilation as well as vital signs stability. CONCLUSION: High frequency oscillatory ventilation is a safe technique that may be used in the operating room in cases where conventional ventilation failed to provide satisfactory gas exchange. PMID- 10522587 TI - Etiology of preoperative anemia in patients undergoing scheduled cardiac surgery. AB - PURPOSE: Ten percent of our cardiac surgical patients have preoperative anemia. Anemia diagnosed before scheduled cardiac surgery is a strong predictor of the need for homologous blood transfusion (RBC) perioperatively but the cause of this preoperative anemia is not known. The purpose of this study was to evaluate the etiology of preoperative anemia. METHODS: Seventy-five consecutive anemic cardiosurgical patients (Hb = < 120 g x L(-1) the day before surgery) were studied prospectively. All had multiple diagnostic blood tests done in the preoperative period to diagnose the cause of the anemia and subsequently underwent non-emergency cardiac surgery. Anesthesia and RBC transfusion were standardized according to the protocol. Data in respect to operation, RBC and other blood product transfusion during operation and hospital stay were recorded. RESULTS: Hospital-acquired anemia was present in 37.3% of anemic patients (hemoglobin decrease during hospitalization before surgery > or =9 g x l(-1)). The second most common diagnosis was iron deficiency anemia (29.3% patients) followed by anemia of chronic renal disease (10.7% patients). When coronary angiography was performed close to operation time, patients had a higher decrease in hemoglobin concentration during hospitalization --suggesting that blood loss during angiography was, in part, responsible for anemia. Seventy-five percent of anemic patients were transfused with RBC perioperatively compared with our overall transfusion rate of 30% of cardiac surgery patients. CONCLUSIONS: In the majority of patients, preoperative anemia is potentially preventable. Investigation and treatment of anemia before cardiac surgery should be a priority in preparing the patient for surgery. PMID- 10522586 TI - Airway obstruction due to late-onset angioneurotic edema from angiotensin converting enzyme inhibition. AB - PURPOSE: Angioneurotic edema is a well-documented complication of angiotensin converting enzyme inhibitors (ACEI). We report a case of acute airway obstruction from a late-onset, probable ACEI-related angioneurotic edema and its subsequent management. CLINICAL FEATURES: A 48-yr-old obese man presented for transurethral resection of a bladder tumour (TURBT). His past medical history included hypertension controlled with hydrochlorothiazide and quinapril which had been started 13 mo earlier. Previous surgery was uncomplicated. Midazolam was used for premedication and for intraoperative sedation together with fentanyl and propofol. After uneventful spinal anesthesia with bupivacaine, operation and recovery, he was transferred to the floor. Five hours later he developed severe edema of his face, tongue and neck, with drooling, that progressed into airway obstruction and respiratory arrest. Ventilation was restored via immediate cricothyroidotomy, and a subsequent tracheotomy was completed uneventfully in the operating room. His serum C1 esterase inhibitor levels at 1, 5 and 23 days later were normal. The angioneurotic edema was attributed to the ACEI treatment. The edema resolved after 48 hr, and further follow-up was unremarkable. CONCLUSION: This observation is consistent with other reports that angioneurotic edema from ACEI can occur many months after the initiation of treatment. This can involve the airway and may produce life-threatening respiratory compromise. Physicians should be aware of this association and the possible need for immediate surgical intervention for the establishment of an airway in case of worsening edema or respiratory arrest. PMID- 10522588 TI - Catheter entrapment by atrial suture during minimally invasive port-access cardiac surgery. AB - PURPOSE: The port-access approach allows surgeons to perform heart operations through small intercostal openings, or "ports". This technique requires new skills for anesthesiologists. A pulmonary artery venting (PAV) catheter and, in some cases, a coronary sinus catheter (for administration of retrograde cardioplegia) are positioned with the aid of fluoroscopy and transesophageal echography (TEE). Both catheters have a wider diameter than the more commonly used conventional PA catheter and present distinctive features. We report a case in which a pulmonary artery venting catheter was entrapped by a suture during a port-access procedure. CLINICAL FEATURES: A 35-yr-old man with severe mitral valve insufficiency was scheduled for valve repair. After a successful bypass procedure, resistance was felt while attempting to withdraw the PAV catheter. On fluoroscopy, fixation of the catheter at the heart level was established and perforation by suture was confirmed after injection of a contrast agent. Because of the risk of cardiac wall rupture and tamponade, the thorax was reopened. After release of some atrial sutures, the catheter could be withdrawn easily. Transfixion by a suture was confirmed by visual examination. CONCLUSION: The more frequent use of a PAV catheter in minimally invasive cardiac surgery with the port-access technique should remind the anesthesiologist of the higher risk of entrapment by surgical sutures. Surgeons should be aware of the risk of accidentally transfixing this catheter during closure of the atriotomy via the port. PMID- 10522589 TI - Assessment of laryngeal view: percentage of glottic opening score vs Cormack and Lehane grading. AB - PURPOSE: To examine the intra- and inter-rater reliability of two methods that categorize laryngeal view during direct laryngoscopy, the Cormack-Lehane grading system and a new scale, the percentage of glottic opening (POGO) scale. METHODS: Seven anesthesiologists from the University of Pennsylvania Health System viewed 25 identical pairs of slides of laryngeal views during direct laryngoscopy. Each anesthesiologist rated the 50 slides for both Cormack-Lehane grades and POGO scores. The latter CL replaces grades 1 and 2 C-L grades with a percentage of glottic opening: the POGO score. Inter and intra-physician reliability for the Cormack-Lehane grades were determined using the kappa statistic analysis, comparison of POGO scores was performed using the intraclass correlation coefficients (rI). RESULTS: The POGO score had a better inter and intra-physician reliability than the Cormack-Lehane grading system. The intra-physician reliability for the POGO score was very good with an average interclass rI value of 0.88. The inter-physician score was good with a rI of 0.73. The Cormack-Lehane grading system had excellent intra-physician concordance (average kappa = 0.83.) but the inter-physician reliability was poor (kappa = 0.16.) CONCLUSION: The Cormack-Lehane grading system has very poor inter-physician reliability. The lack of inter-physician reliability with Cormack-Lehane grading calls into question the results of previous studies in which different laryngoscopists used this method to assess laryngeal view. The POGO score appears to have good intra and inter-rater reliability. It has several theoretical advantages and may prove to be more useful for research studies in direct laryngoscopy. PMID- 10522590 TI - Ramosetron vs granisetron for the prevention of postoperative nausea and vomiting after laparoscopic cholecystectomy. AB - PURPOSE: To compare the efficacy of ramosetron with granisetron for the prevention of postoperative nausea and vomiting (PONV) after laparoscopic cholecystectomy. METHODS: In a randomized, double-blind study, 80 female inpatients received 3 mg granisetron or 0.3 mg ramosetron i.v. (n=40 of each) at the completion of surgery The standardized anesthetic included isoflurane and nitrous oxide in oxygen. RESULTS: Complete response, defined as no PONV, during the first 24 hr (0-24 hr) after anesthesia was 85% with granisetron and 93% with ramosetron, respectively (P=0.241); the corresponding incidence during the next 24 hr (24-48 hr) after anesthesia was 63% and 90% (P=0.004). No clinically important adverse events due to the study drug were observed in any of the groups. CONCLUSION: Ramosetron was more effective than granisetron for prevention of PONV during 0-48 hr after anesthesia for laparoscopic cholecystectomy. PMID- 10522591 TI - Obstetric anesthesia sites on the Internet. PMID- 10522592 TI - Consent in anesthesia research: the pre-admission phone call. PMID- 10522593 TI - Use of the laryngeal mask after tracheal extubation. PMID- 10522594 TI - Misleading end-tidal CO2 tensions. PMID- 10522595 TI - Every endotracheal tube needs a Murphy eye! PMID- 10522596 TI - Similar invasive procedures, but different techniques. (A potential disaster.) PMID- 10522597 TI - Unexpected left atrial occlusion secondary to esophageal tumour terminates anesthesia. PMID- 10522598 TI - Leukocyte reduction filter problems. PMID- 10522599 TI - Developments in serologic detection of human pancreatic adenocarcinoma. PMID- 10522600 TI - Fish oil reduces gastric acid secretion. AB - BACKGROUND: The aim of the present investigation was to study gastric acid secretion and release of gastrin, cholecystokinin (CCK), and secretin during intraduodenal perfusion of either fish oil or trioleate. METHODS: Seven healthy volunteers were stimulated on two separate days in random order with intraduodenal perfusates of either fish oil or trioleate. RESULTS: Intravenous infusion with gastrin-17 was used as a background stimulation in doses mimicking a postprandial situation (39.9 +/- 4.8 pmol/l fish oil and 43.6 +/- 3.8 pmol/l trioleate). Gastric acid secretion increased significantly from a basal level of 0.7 +/- 0.1 meq/15 min to 4.0 +/- 0.6 meq/15 min (P < 0.05) before perfusion of fish oil, which reduced gastric acid secretion to 1.9 +/- 0.4 meq/15 min (P < 0.01). After termination of fish oil perfusion gastric acid secretion increased to preperfusion concentrations (P < 0.01). Perfusion of trioleate did not influence gastric acid secretion. Plasma concentrations of CCK rose significantly during perfusion of fish oil (from 2.8 +/- 0.6 pmol/l to 4.4 +/- 0.7pmol/l, P<0.01), whereas trioleate only tended to increase CCK concentrations. Plasma concentrations of secretin did not change during perfusion of fish oil; however, concentrations were significantly lower during and after perfusion of trioleate (P < 0.01). CONCLUSION: The present study shows that intraduodenal perfusion of fish oil is associated with a significant reduction of the gastric acid secretion stimulated by gastrin in healthy humans. PMID- 10522601 TI - Therapy and diagnostic tests used for Helicobacter pylori infection in the Scandinavian countries in 1998. AB - BACKGROUND: We wanted to ascertain how Helicobacter pylori infection is managed in Scandinavia. METHODS: A one-page questionnaire with seven questions was mailed in April 1998 to 1718 gastroenterologists in Finland, Denmark, Norway, and Sweden (excluding Swedish surgeons). RESULTS: The questionnaire was returned by 36%. Antimicrobials were used by 99% for peptic ulcer associated with H. pylori, by 67% for mucosa-associated lymphoid tissue lymphoma, by 27% before long-term therapy with a proton-pump inhibitor (PPI), by 16% for non-ulcer dyspepsia, by 11% for reflux disease, and by 11% for other indications. In Finland several conditions other than ulcer were treated more frequently than in the other countries. The commonest primary therapy is PPI triple therapy (94%), followed by bismuth-based (11%), 'other' (2%), and PPI dual therapy (0.2%). Primary bismuth based therapy was almost completely limited to Norway. The commonest secondary therapy for failures was also PPI triple therapy (71%), followed by bismuth-based (41%), 'other' (10%), and PPI dual therapy (1%). Clarithromycin for primary therapy was used much less frequently in Finland than in the other countries. Follow-up to ascertain whether eradication is successful was done always or often by 90% in Finland, 63% in Norway, 62% in Sweden, and 21% in Denmark and by 61% of the internists and 42% of the surgeons. The commonest method to confirm eradication was gastroscopy (69%), followed by the breath test (52%) and serology (11%). CONCLUSIONS: In Scandinavia H. pylori associated with peptic ulcer disease is treated with antimicrobials by virtually all gastroenterologists. PPI triple therapy is the commonest regimen for primary and secondary eradication. PPI dual therapy has essentially disappeared. Fifty-four per cent confirm eradication always or often, with gastroscopy being the commonest method. PMID- 10522602 TI - Serologic screening before endoscopy: the value of Helicobacter pylori serology, serum recognition of the CagA and VacA proteins, and serum pepsinogen I. AB - BACKGROUND: We wanted to assess the diagnostic value of pre-endoscopy screening by Helicobacter pylori serology, serum recognition of the CagA and VacA proteins, and serum pepsinogen I levels (sPGI) in patients up to 55 years of age with uncomplicated simple dyspepsia. METHODS: Consecutive dyspeptic patients referred for open-access endoscopy, excluding patients with alarm symptoms, recent intake of acid suppressants, or ingestion of non-steroidal anti-inflammatory drugs. H. pylori status was determined by histology and urease testing. H. pylori serologic status was determined with the enzyme-linked immunosorbent assay (ELISA) and Western blotting, serum recognition of CagA and VacA with Western blot, and sPGI levels by radioimmunoassay. RESULTS: One hundred and fifteen patients were studied (mean age, 40 years: range, 20-55 years), of whom 58 were H. pylori positive in biopsy-based tests. Twenty-one patients (18%) had significant gastroduodenal lesions (erosions, ulcers, or cancer). The sensitivity (specificity) of the ELISA (optimized) and Western blot in determining H. pylori status was 94.8% (89.5%) and 100% (96.4%), respectively. Screening strategies based on the ELISA or Western blot for determining H. pylori serologic status would have detected 95% or 100% of significant lesions, respectively, and each 'saved' 47% of endoscopies for simple dyspepsia. Serum recognition of the CagA protein would have detected 95% of significant lesions and 'saved' 55% of endoscopies, whereas recognition of the VacA protein would have detected only 81% of the lesions. Screening by H. pylori serology plus a 'low' (<55 ng/ml) or 'high' sPGI (>125 ng/ml) would detect only 57% of significant lesions, although the only case of cancer was included in the hypopepsinogenaemic subgroup of just 11 patients. CONCLUSIONS: In patients with uncomplicated, simple dyspepsia up to 55 years of age, screening by H. pylori serology identified 95%-100% of patients with significant gastroduodenal lesions while potentially saving 46.9% of endoscopies. Serum recognition of the CagA protein identified 95% of lesions and would have saved an additional number of endoscopies (7.9%) compared with basic serology. Measurement of sPGI was of limited diagnostic value. PMID- 10522603 TI - An 18-year follow-up study of chronic gastritis and Helicobacter pylori association of CagA positivity with development of atrophy and activity of gastritis. AB - BACKGROUND: We wanted to evaluate the course of chronic gastritis and its association with Helicobacter pylori and CagA seropositivity in an adult sample from Saaremaa (Estonia) during an 18-year follow-up. METHODS: Seventy persons (31 men, 39 women; median age, 57.5 years) from a primary sample of 304 subjects endoscoped in 1979 were reinvestigated by endoscopy and biopsy in 1997. The state of the gastric mucosa and the presence of H. pylori in histologic sections from the antrum and corpus were assessed both in 1979 and 1997 in 66 subjects in accordance with the Sydney system, and H. pylori status in all 70 subjects was determined with the enzyme-linked immunosorbent assay (ELISA). Anti-CagA IgGs were determined with the ELISA, using the recombinant fragment of CagA. RESULTS: During an 18-year follow-up 11% of the subjects developed atrophy in the antrum, whereas 35% developed it in the corpus. Development of atrophy in the corpus and the appearance of intestinal metaplasia in the antrum were associated with increased activity of gastritis both in the initial and last follow-up biopsies. Anti-CagA positivity was found in 71% of H. pylori-seropositive persons (94% of subjects). There was a significant association between CagA positivity and the activity of gastritis, the presence of atrophy or damage to surface epithelial cells in the antrum and in corpus mucosal biopsy specimens at the last follow-up endoscopy. CONCLUSION: The CagA-positive strains of H. pylori enhance the development of atrophic gastritis compared with CagA-negative strains. PMID- 10522604 TI - Psychometric validation of a constipation symptom assessment questionnaire. AB - BACKGROUND: Clinical management of constipation is complicated by the lack of a gold standard for evaluation of symptoms. A constipation symptom assessment instrument, the PAC-SYM, was developed to address the patient perspective on the disorder. Instrument content was based on literature review and results of focus groups. METHODS: Two hundred and sixteen patients at nine sites participated in a 6-week psychometric evaluation of the PAC-SYM. The final instrument contained 12 items assigned to 3 subscales: stool symptoms, rectal symptoms, and abdominal symptoms. The psychometric properties of this final instrument were assessed. RESULTS: Internal consistency and test-retest reliability of the final instrument was high (Cronbach's alpha = 0.89; intraclass correlation = 0.75). Concurrent validity was supported by the correlation with both subject and investigator constipation severity ratings (r= 0.68 and 0.72, respectively; P < 0.0001). Scores were moderately correlated with instruments measuring quality of life. Comparison of treatment responders with nonresponders showed the ability of the instrument to differentiate between groups on the basis of clinical severity (t = -6.12, P < 0.0001 ). Scores changed significantly over time among responders, indicating instrument responsiveness. CONCLUSIONS: The PAC-SYM is internally consistent, reproducible under stable conditions, valid, and responsive to change and provides a comprehensive means to assess the effectiveness of treatment for constipation. PMID- 10522605 TI - Pharyngo-esophageal motility disturbances in patients with myotonic dystrophy. AB - BACKGROUND: Esophageal motility is often disturbed in patients with myotonic dystrophy. The esophageal motor derangement pattern and its correlation with esophageal and peripheral motor symptoms is not well defined. Our aims were to evaluate: 1) pharyngo-esophageal motor abnormalities in these patients; 2) the relationship between motor involvement and clinical manifestations; and 3) the correlation between pharyngo-esophageal motility abnormalities and peripheral neuromuscular involvement. METHODS: We compared data from 18 patients and 18 healthy controls. Neuromuscular affectation was quantified with a five-point muscular disability rating scale. Pharyngo-esophageal symptoms were assessed with a directed questionnaire, whereas motility was evaluated by means of manometry. RESULTS: Myotonic dystrophy patients had diminished pharyngeal contraction amplitude, upper esophageal sphincter basal pressure, and esophageal body contraction amplitude compared with the control group (P < 0.001). No signs of esophageal myotony were evident. Simultaneous esophageal waves after more than 40% of liquid swallows were found in 80% of patients. No relationship between esophageal manometric alteration and esophageal or peripheral motility symptoms was elicited. CONCLUSION: In patients with myotonic dystrophy pharyngo-esophageal motility is severely deranged in both amplitude and coordination. These abnormalities may be present even if symptoms referred by the patient or the severity of the disease is not remarkable. PMID- 10522606 TI - Gastric myoelectric activity in patients with end-stage liver disease. AB - BACKGROUND: Abnormalities of gastrointestinal motility and transit time have been reported in association with end-stage liver disease. Motility abnormalities could be routinely studied if a simple noninvasive test were available. The electrogastrogram is a cutaneous measure of gastric myoelectric activity and correlates well with serosal recordings of gastric myoelectric activity. The aim of this study was to evaluate gastric myoelectric activity in patients with end stage liver disease. METHODS: Fourteen patients with end-stage liver disease had gastric myoelectric activity measured with the electrogastrogram. An electrogastrogram was considered abnormal when normal gastric slow waves were seen less than 70% of the time or there was no increase in the electrogastrogram amplitude after a meal. RESULTS: Abnormal electrogastrograms were present in 8 of 14 (57%) end-stage liver disease patients. CONCLUSIONS: Abnormal gastric myoelectric activity is common in end-stage liver disease. PMID- 10522607 TI - Small-bowel involvement in systemic lupus erythematosus: a morphometric and immunohistochemical study. AB - BACKGROUND: We have investigated the intestinal mononuclear cell subpopulations in patients with systemic lupus erythematosus (SLE) and correlated these with the disease activity. METHODS: Eighteen female outpatients were studied; in 10 of them lupus activity was measured with the Lupus Activity Criteria Count and the SLE Disease Activity Index. Eight patients were in lupus remission. The control group consisted of 10 healthy volunteers. Peroral jejunal biopsy was performed in all individuals, at the angle of Treitz, using a Watson capsule, under X-ray control. Histologic studies analysed the villous to crypt ratio, lamina propria cells, and intraepithelial lymphocyte count. Immunohistochemical evaluation was carried out with the indirect immunoperoxidase technique, using monoclonal antibodies against CD3, CD4, CD8, D1, D7, D9, and M1. RESULTS: Lamina propria CD3+, CD8+, D7+, and M1+ cells from patients with SLE did not differ significantly from those of controls. CD4+ cells were decreased in all patients with SLE, especially in the clinically inactive patients. D1+ and D9+ cells were also decreased in all patients. CONCLUSION: The finding of quantitative abnormalities in the cell-mediated immunity of the intestinal mucosa may reflect systemic defects of the immune system in SLE. PMID- 10522608 TI - Fat and mesenteric blood flow. AB - BACKGROUND: The introduction of fat to the duodenum leads to an increase in mesenteric blood flow. The exact mechanism, however, is unknown. In this study we investigate the influence of the terminal carboxy group of the oleic acid molecule on superior mesenteric artery blood flow. METHODS: In six healthy male volunteers duplex ultrasound was used to evaluate the effects of duodenal perfusion of 48 mmol oleic acid and 48 mmol oleyl alcohol on superior mesenteric artery blood flow variables and diameter. RESULTS: The blood flow variables time average velocity and maximal diastolic velocity increased significantly, and the resistance index decreased significantly during perfusion with oleic acid, but during oleyl alcohol perfusion no changes were found. No significant changes in vessel diameter were observed at any time. CONCLUSION: The carboxy group of the oleic acid molecule has an important role in the duodenum in mediating the postprandial increase in superior mesenteric artery blood flow. PMID- 10522609 TI - Low trehalase activity is associated with abdominal symptoms caused by edible mushrooms. AB - BACKGROUND: The purpose of the study was to evaluate whether maldigestion of trehalose causes abdominal symptoms and which available diagnostic method best distinguishes intolerant from tolerant subjects. METHODS: A 25-g oral trehalose load test was performed in 64 subjects. The 19 experiencing clear symptoms constituted the trehalose-intolerant subjects. Changes from base-line levels of blood glucose, breath hydrogen, and methane and symptoms were recorded after the test. Trehalase activity was determined in serum and on a duodenal biopsy specimen obtained by endoscopy. RESULTS: Intolerant subjects were best differentiated from tolerant subjects by changes in breath gases (hydrogen and methane) and duodenal trehalase to sucrase ratio. The change in breath gases correlated inversely with duodenal trehalase activity, duodenal trehalase to sucrase ratio, and plasma trehalase activity. The correlation between serum and duodenal trehalase activities was on the order of 0.6. Two subjects were found to have trehalase deficiency. CONCLUSIONS: It is obvious that trehalose maldigestion can cause symptoms similar to those of lactose maldigestion and intolerance. Three factors control the genesis of symptoms: 1) the activity of small-bowel trehalase: if it is low, trehalose is maldigested and more trehalose is passed into the colon; 2) the maldigested trehalose, which causes osmotic water flow into the colon, resulting in loose stools and diarrhea; and 3) most importantly, the microflora of the colon, from which symptoms will arise if there are bacteria capable of producing gases from maldigested trehalose. If colonic bacteria cannot produce gases, then distention of the abdomen and intestinal gas expulsion as eructations and flatus will not occur. PMID- 10522610 TI - Low circulating insulin-like growth factor I in coeliac disease and its relation to bone mineral density. AB - BACKGROUND: Patients with coeliac disease have low bone mineral density (BMD), but the underlying mechanisms are unclear. Our aim was to study circulating insulin-like growth factor I (IGF-I) and its possible relationship to BMD in adults with untreated coeliac disease and after 1 year on a gluten-free diet. METHODS: In 29 consecutive adult coeliac patients fasting IGF-I and BMD (n = 28) were examined before and 1 year after starting a gluten-free diet. Intact parathyroid hormone (PTH) was measured (n = 20) before the gluten-free diet was started. RESULTS: Untreated coeliac patients had lower IGF-I values than controls matched for age and sex, and their BMD was low. A relationship was observed between BMD and IGF-I but not independent of age and body mass index. During the 1st year on a gluten-free diet BMD increased (P < 0.001), as did the circulating IGF-I levels in 21 of the 29 patients (P = 0.078). In the subgroup of 14 patients with normal initial PTH the increase in IGF-I correlated positively with the increase in BMD (femoral trochanter, r = 0.62, P < 0.05, and lumbar spine, r = 0.70, P < 0.02). CONCLUSIONS: BMD and circulating IGF-I levels are low in adults with untreated coeliac disease. In patients with normal initial PTH level there is an association between the change in BMD and circulating IGF-I, although this parallel increase may not be causally connected. PMID- 10522611 TI - Persistent mucosal abnormalities in coeliac disease are not related to the ingestion of trace amounts of gluten. AB - BACKGROUND: It is expected that in patients with coeliac disease the small-bowel mucosal mucosa will return to normal if they adhere to a gluten-free diet (GFD). However, in many this is not the case. This study aims to determine whether this persistent villous atrophy (VA) could be due to continued ingestion of the trace amounts of gluten in 'gluten-free' foods, as defined by the WHO/FAO Codex Alimentarius. METHODS: Duodenal biopsy specimens from 89 adults with long standing coeliac disease were examined, and the findings correlated with their form of gluten-free diet. RESULTS: In 51 subjects the duodenal specimen was normal, whereas in 38 there was villous atrophy (partial, 28; subtotal, 8; total, 2). There was no relationship between the presence or absence of VA and ingestion of either a GFD as defined by the Codex Alimentarius (Codex-GFD; 39 patients) or a GFD that contained no detectable gluten (NDG diet: 50 patients). Intraepithelial lymphocyte counts were higher, and lactase levels lower, in subjects with an abnormal biopsy specimen than in those in whom it was normal. However, within each of these biopsy groups there was no difference in these variables between patients on a Codex-GFD and those on an NDG-GFD. IgA antigliadin antibody was detected in 4 of 29 patients on a Codex-GFD and in 3 of 13 on a NDG-GFD (NS). CONCLUSION: The persistent mucosal abnormalities seen in patients with coeliac disease on a GFD are not due to the ingestion of trace amounts of gluten. The consequences of these abnormalities have yet to be determined. PMID- 10522612 TI - Ursodeoxycholic acid increases the activities of alkaline sphingomyelinase and caspase-3 in the rat colon. AB - BACKGROUND: Ursodeoxycholic acid (UDCA) has been found to inhibit the development of colon carcinoma induced by chemical carcinogens with unidentified mechanisms. Sphingomyelin metabolism has emerged as a novel signal transduction pathway closely related to cell proliferation and apoptosis. We recently found that alkaline sphingomyelinase (SMase) activity was decreased in human colon cancer. The present study is to investigate whether UDCA has effect on the levels of SMase and whether the activity of caspase-3, a key regulatory protease in apoptosis that can be activated by sphingomyelin breakdown products, is also influenced by UDCA. METHODS: Rats were fed UDCA in amounts ranging from 37.5 to 300 mg/kg/day for 10 days by gavage. The colonic mucosa was scraped, homogenized, and sonicated. The activities of acid, neutral and alkaline SMases, and caspase-3 were determined. RESULTS: UDCA dose-dependently increased alkaline SMase activity in colonic mucosa and faeces, slightly increased acid SMase activity in the mucosa, and had no effect on neutral SMase. UDCA also dose-dependently increased caspase-3 activity in the colonic mucosa, and the increase correlated significantly with the changes in alkaline but not that in acid or neutral SMase activity. CONCLUSIONS: UDCA increases alkaline sphingomyelinase and caspase-3 activities, which might be a mechanism involved in its anticarcinogenic effect on colon cancer development. PMID- 10522613 TI - The effect of increasing blood pressure with dopamine on systemic, splanchnic, and lower extremity hemodynamics in patients with acute liver failure. AB - BACKGROUND: Arterial hypotension occurs frequently in patients with acute liver failure (ALF). Treatment with epinephrine and norepinephrine in patients with ALF has been associated with a decrease in whole-body (systemic) oxygen consumption. We aimed to investigate the effect of increasing blood pressure with dopamine on whole-body (systemic), splanchnic, and lower extremity hemodynamics and oxygen consumption in patients with acute liver failure and hepatic encephalopathy grade III or IV. METHODS: In seven patients with ALF cardiac output (CO) was measured with the thermodilution technique, and hepatic blood flow (HBF) was estimated with infusion of sorbitol as test compound, liver vein catheterization, and calculations on the basis of Fick's principle. Lower-extremity blood flow was measured with strain-gauge plethysmography. RESULTS: During infusion of dopamine (5 +/- 2 microg kg(-1) min(-1)) mean arterial pressure (MAP) increased from 68 +/ 5 to 85 +/- 8 mmHg. CO increased from 6.8 +/- 0.8 to 9.0 +/- 2.4 l/min (P < 0.05), systemic oxygen delivery from 45 +/- 7 to 63 +/- 19 mmol/min (P < 0.05), systemic oxygen consumption from 10.2 +/- 2.0 to 11.5 +/- 3.3 mmol/min (NS). HBF increased from 2.2 +/- 0.7 to 2.7 +/- 1.0 l/ min (P < 0.05), splanchnic oxygen delivery from 14.4 +/- 5.3 to 18.5 +/- 7.2 mmol/min (P < 0.01), and splanchnic oxygen consumption decreased from 3.9 +/- 1.1 to 2.9 +/- 0.6 mmol/min (P < 0.05). No significant changes in lower extremity flow and oxygenation variables were found. CONCLUSIONS: The use of dopamine in patients with ALF to increase MAP was associated with increases in systemic and splanchnic oxygen delivery. A concomitant decrease in splanchnic oxygen consumption was observed. PMID- 10522614 TI - Intravenous recombinant interferon-beta versus interferon-alpha-2b and ribavirin in combination for short-term treatment of chronic hepatitis C patients not responding to interferon-alpha. Multicenter Interferon Beta Italian Group Investigators. AB - BACKGROUND: Little is known about the therapeutic role of intravenous interferon beta in chronic hepatitis C patients unresponsive to a previous treatment with interferon-alpha. METHODS: Two hundred interferon-alpha non-responders were randomized to receive either intravenous recombinant interferon-beta or interferon-alpha-2b and ribavirin for 12 weeks. The responders in both groups were followed up for a further 48 weeks. RESULTS: At week 12 a biochemical and virologic response was documented in 42% of the patients treated with interferon beta and in 22% of the patients treated with combination therapy. A sustained response was observed in 21% of the patients treated with interferon-beta and in 13% of those treated with combination therapy, with similar differences on intention-to-treat analysis. CONCLUSIONS: Short-term treatment with intravenous interferon-beta seems to offer a chance for sustained response in a subset of interferon-alpha non-responders. The role of long-term therapy in these patients still remains to be explored. PMID- 10522615 TI - Detection of hepatitis B virus DNA in tissues of hepatocellular carcinomas related to hepatitis C virus which are negative for hepatitis B virus surface antigen. AB - BACKGROUND: Uniform conclusions have not yet been drawn as to the frequency and copy number of hepatitis B virus (HBV) DNA in hepatocellular carcinoma (HCC) tissues from patients who are negative for hepatitis B surface antigen (HBsAg). METHODS: The existence of HBV DNA was investigated with Southern blot hybridization and polymerase chain reaction (PCR) in HCC tissues from 55 patients who were negative for HBsAg and positive for hepatitis C virus antibody (HCVAb). RESULTS: Southern blot hybridization showed that two tissues (4%) contained HBV DNA at a frequency of more than 1 HBV copy per 10 cells. Another 5 tissues were found to be positive for HBV DNA only by PCR analysis but contained less than 3 HBV DNA copies per 10(5) cells. CONCLUSIONS: HCC tissues from patients seronegative for HBsAg and seropositive for HCVAb contain HBV DNA less frequently than has generally been claimed. PMID- 10522616 TI - Complete recovery after spontaneous drainage of pancreatic abscess into the stomach. AB - Pancreatic abscess is a dreaded complication of acute pancreatitis, with a high death rate even with aggressive surgical treatment. We report two cases in which recovery followed spontaneous drainage into the stomach. A 75-year-old woman with biliary pancreatitis and a 63-year-old man with ethanol-induced pancreatitis both developed pancreatic abscess, diagnosed by computed tomography scans and ultrasound. The spontaneous gastric fistula was heralded by a large emesis of purulent and necrotic material in one case and copious nasogastric tube secretions of a similar material in the other. Defervescence was immediate, and both patients went on to complete recovery without any further interventions. Contrast studies showed the fistulae. It is concluded that in the event that a pancreatic pseudocyst spontaneously drains into the stomach a 'wait and see' policy should be adopted, and a favorable outcome can be expected. PMID- 10522617 TI - 13C-urea breath test for helicobacter pylori infection: stability of samples over time. PMID- 10522618 TI - Helicobacter pylori and human leukocyte antigens. PMID- 10522619 TI - Evaluating tinnitus. PMID- 10522620 TI - One-year vestibular and balance outcomes of Oklahoma City bombing survivors. AB - This multisite investigation assessed subjective, behavioral, and objective balance function in 30 blast survivors. Subjects with vertigo, dizziness, or imbalance were screened (n = 6) or evaluated (n = 27) during 1 year. Tests included a questionnaire, electronystagmography (ENG), and computerized dynamic posturography (CDP). Ninety-seven percent of subjects were located inside a building during the blast, and 63 percent of subjects experienced dysequilibrium within 48 hours. Forty-three percent of symptoms could not be attributed to head injury. Sixty percent of subjects had abnormal ENG and/or CDP; ENG abnormalities mostly were peripheral or nonlocalizing, whereas CDP patterns were "vestibular," "surface dependent," and "physiologically inconsistent." At 1-year postblast, 55 percent of initially abnormal CDP results were normal, and 72 percent of subjects said symptoms were unchanged or occurred intermittently. A serial, test battery approach is recommended to assess symptoms. Blast-related dysequilibrium had clinically significant manifestations and should be considered a valid component of aural blast injury. PMID- 10522621 TI - Auditory neuropathy and a mitochondrial disorder in a child: case study. AB - A child was referred for an audiologic evaluation, to include auditory brainstem evoked response testing, due to inconsistent responses to sound and delayed speech and language development. Results were characteristic of auditory neuropathy. In view of subsequent decline in motor function, a genetics evaluation was conducted, revealing a mitochondrial disorder. A brief overview of mitochondrial genetics in association with hearing loss is presented. The patient's audiologic profile is described and the implications for management are discussed. PMID- 10522622 TI - Tinnitus Handicap Inventory (THI) as a hearing aid outcome measure. AB - This study assessed the effects of hearing aids on the perception of tinnitus using the Tinnitus Handicap Inventory (THI). THI benefit scores (unaided-aided) were examined in relation to hearing aid benefit as measured with the Abbreviated Profile of Hearing Aid Benefit (APHAB) inventory. The THI benefit was also related to the users' ratings of overall satisfaction with their hearing aids. Thirty-four novice hearing aid users with complaints of hearing loss and tinnitus participated in the study. Outcome measures were obtained 6 weeks after the hearing aid fittings. The results showed that hearing aid use reduced tinnitus handicap significantly, but, typically, the effect was small. The association between overall satisfaction ratings and THI benefit scores was weak. In contrast, the overall satisfaction ratings were strongly related to benefit on the speech subscales (average of Ease of Communication, Reverberation, and Background Noise) but not on the Aversiveness subscale of the APHAB. The weak relationship observed between THI benefit and benefit on the speech subscales of the APHAB suggested that the two inventories were not redundant. The results of the study suggest that the THI can make a useful contribution to the overall profile of hearing aid benefit for new hearing aid users with tinnitus. PMID- 10522623 TI - Efficacy of a modified politzer apparatus in management of eustachian tube dysfunction in adults. AB - This study evaluates the efficacy of politzeration on eustachian tube dysfunction following airplane travel using an automated, hand-held device that controls the volume velocity of air flow. Fourteen adults with eustachian tube dysfunction following airplane travel comprised the experimental group. They received politzeration over a period of 6 weeks on a twice-a-week basis. Fourteen adults with eustachian tube dysfunction following airplane travel comprised the control group. They were untreated. Complete audiologic and otolaryngologic evaluations were performed at the pretest and after politzeration treatment was completed. The results revealed a substantial, significant improvement in the mean tympanometric peak pressure from pretest to final post-treatment retest in the experimental, but not the control, subjects. The mean air-bone gap increased significantly from pretest to final post-treatment retest in the control, but not the experimental, subjects. Of the experimental subjects with abnormal tympanometric peak pressures at the pretest, resolution to within normal limits occurred in 71 percent of the experimental subjects versus 21 percent of the control subjects. These results suggest the potential feasibility of treatment of aerotitis media in adults with this modified Politzer method using the automated apparatus. PMID- 10522624 TI - Reliability of tinnitus loudness matches under procedural variation. AB - Repeated tinnitus loudness matches (LMs) were obtained to determine response reliability using a computer-automated technique with two procedural variations, fixed or random step sizes, to increase output level during the initial ascending series of tones at each frequency. Twenty subjects with stable, tonal tinnitus were evaluated with both methods during each of two sessions. Response instructions were displayed on a portable computer, and a pen device was used to make response choices that appeared on the touch-sensitive video monitor. For each method, hearing thresholds and LMs were obtained, with 1-dB resolution, at 1/3-octave frequencies from 1 to 16 kHz. Analyses revealed reliability of LMs to be equivalent between methods. LM data are reported in both dB SPL and dB SL, with the SPL values providing greater reliability both within and between sessions (all r's > or = .889, p's < or = .0001). PMID- 10522625 TI - Fair, honest, ethical, and last. PMID- 10522626 TI - Exercise in old age: time to unwrap the cotton wool. PMID- 10522627 TI - Perpetuating ignorance: intravenous fluid therapy in sport. PMID- 10522628 TI - You can run but you can't hide: the role of concussion severity scales in sport. PMID- 10522629 TI - Oxidative stress and exercise: need for antioxidant supplementation? PMID- 10522630 TI - Tobacco sponsorship of sport. PMID- 10522631 TI - Sport injuries of the elbow. PMID- 10522632 TI - Epidemiology of injury in elite and subelite female gymnasts: a comparison of retrospective and prospective findings. AB - OBJECTIVES: An 18 month prospective injury survey was conducted on 64 Australian elite and subelite female gymnasts. The aims were to determine the rate of injury, anatomical location, and types of injury incurred by female competitive gymnasts, and to compare the findings with data collected retrospectively from the same sample of gymnasts. METHODS: The gymnasts recorded (weekly) in an injury record booklet the number of hours trained and information on any injuries suffered over that week. RESULTS: The sample reported 349 injuries, a rate of 5.45 per person (6.29 for the elite and 4.95 for subelite gymnasts) over the 18 month survey. Injuries to the ankle and foot (31.2%) were the most commonly reported, followed by the lower back (14.9%). The most prevalent type of injury were sprains (29.7%), followed by strains (23.2%), and growth plate injuries (12.3%). The elite gymnasts reported that, for each injury, they missed fewer training sessions (p = 0.01), but modified more sessions (p = 0.0001) than their subelite counterparts. Further, the elite gymnasts spent 21.0% of the year training at less than full capacity because of injury. Although a significantly higher number of injuries were recorded in the prospective study (p = 0.0004), no differences were found between the distribution of injury by anatomical location or type between the two methods of data collection. CONCLUSIONS: The findings have important implications in terms of training procedures and periodic screening of gymnasts. PMID- 10522633 TI - Electrocardiographic changes in 1000 highly trained junior elite athletes. AB - OBJECTIVES: To evaluate the spectrum of electrocardiographic (ECG) changes in 1000 junior (18 or under) elite athletes. METHODS: A total of 1000 (73% male) junior elite athletes (mean (SD) age 15.7 (1.4) years (range 14-18); mean (SD) body surface area 1.73 (0.17) m2 (range 1.09-2.25)) and 300 non-athletic controls matched for gender, age, and body surface area had a 12 lead ECG examination. RESULTS: Athletes had a significantly higher prevalence of sinus bradycardia (80% v 19%; p<0.0001) and sinus arrhythmia (52% v 9%; p<0.0001) than non-athletes. The PR interval, QRS, and QT duration were more prolonged in athletes than non athletes (153 (20) v 140 (18) milliseconds (p<0.0001), 92 (12) v 89 (7) milliseconds (p<0.0001), and 391 (27) v 379 (29) milliseconds (p = 0.002) respectively). The Sokolow voltage criterion for left ventricular hypertrophy (LVH) and the Romhilt-Estes points score for LVH was more common in athletes (45% v 23% (p<0.0001) and 10% v 0% (p<0.0001) respectively), as were criteria for left and right atrial enlargement (14% v 1.2% and 16% v 2% respectively). None of the athletes with voltage criteria for LVH had left axis deviation, ST segment depression, deep T wave inversion, or pathological Q waves. ST segment elevation was more common in athletes than non-athletes (43% v 24%; p<0.0001). Minor T wave inversion (less than -0.2 mV) in V2 and V3 was present in 4% of athletes and non athletes. Minor T wave inversion elsewhere was absent in non-athletes and present in 0.4% of athletes. CONCLUSIONS: ECG changes in junior elite athletes are not dissimilar to those in senior athletes. Isolated Sokolow voltage criterion for LVH is common; however, associated abnormalities that indicate pathological hypertrophy are absent. Minor T wave inversions in leads other than V2 and V3 may be present in athletes and non-athletes less than 16 but should be an indication for further investigation in older athletes. PMID- 10522634 TI - Leisure physical activity and various pain symptoms among adolescents. AB - OBJECTIVES: To investigate the association between leisure physical activity and various pain symptoms in adolescents. METHODS: In this nationwide cohort based cross sectional study in Finland, 698 schoolchildren, 344 girls and 354 boys, aged 10 to 17 years responded to a questionnaire on pain symptoms (neck and shoulder pain, upper back pain, low back pain, upper limb pain, lower limb pain, headache, and abdominal pain) and physical activity habits and also participated in a fitness test. RESULTS: Reported physical activity correlated with measured fitness. Musculoskeletal pains (p = 0.013) (in particular low back pain (p = 0.022), upper limb pain (p<0.001), and lower imb pain (p<0.001)) were found more often in subjects participating in large amounts of leisure physical activity, while non-musculoskeletal pains (p = 0.065) (in particular headache among boys (p = 0.004)) tended to be less common. Co-occurrence of different musculoskeletal pains was common in subjects participating in sports. CONCLUSIONS: In addition to its likely long term health benefits, vigorous physical activity causes musculoskeletal pains during adolescence. This should be considered when tailoring health promotion programmes to adolescents. Also, co-occurrence of musculoskeletal pains may occur as the result of sports activity, which should be considered as a confounder in epidemiological studies on fibromyalgia and related issues. PMID- 10522635 TI - Injuries to polo riders: a prospective evaluation. AB - OBJECTIVE: To assess prospectively the incidence, nature, and severity of injuries to polo riders competing in the 1996 Argentine High Polo season. METHODS: Assessment, documentation, and provision of care for all injuries sustained during the 1996 season by one of the authors. Riders were also surveyed retrospectively for their previous polo injuries. RESULTS: 34 riders took part in the study. Nine injuries were sustained prospectively and 55 injuries were reviewed retrospectively (64 total). The injuries were categorised as minor (10), moderate (13), and major (41). Twenty five (39%) injuries occurred in the arms, 20 (31%) in the legs, 12 (19%) in the head, 3 (5%) in the back, and 4 (6%) in the face. A fracture occurred in 25 (39%) injuries as most resulted from a fall from the horse. Additionally, facial lacerations occurred prospectively in five riders but did not result in missed play. An overall injury rate of 7.8/1000 player-game hours was calculated. CONCLUSIONS: Although many sports have injury rates much greater than 8/1000 player-game hours, the severity of most injuries occurring in polo was classified as major, with fractures and facial lacerations common. The use of a helmet with a face protector is recommended to decrease injury to players. A doctor experienced in the management of serious trauma should be present at all polo matches. PMID- 10522636 TI - A major sporting event does not necessarily mean an increased workload for accident and emergency departments. Euro96 Group of Accident and Emergency Departments. AB - AIM: To determine whether there were any changes in attendance at accident and emergency departments that could be related to international football matches (Euro96 tournament). METHOD: Fourteen accident and emergency departments (seven adjacent to and seven distant from a Euro96 venue) provided their daily attendance figures for a nine week period: three weeks before, during, and after the tournament. The relation between daily attendance rates and Euro96 football matches was assessed using a generalised linear model and analysis of variance. The model took into account underlying trends in attendance rates including day of the week. RESULTS: The 14 hospitals contributed 172 366 attendances (mean number of daily attendances 195). No association was shown between the number of attendances at accident and emergency departments and the day of the football match, whether the departments were near to or distant from stadia or the occurrence of a home nation match. The only observed independent predictors of variation were day of the week and week of the year. Attendance rates were significantly higher on Sunday and/or Monday; Monday was about 9% busier than the daily average. Increasing attendance was observed over time for 86% of the hospitals. CONCLUSION: Large sports tournaments do not increase the number of patients attending accident and emergency departments. Special measures are not required for major sporting events over and above the capacity of an accident and emergency department to increase its throughput on other days. PMID- 10522637 TI - Reliability of ratings of perceived exertion during progressive treadmill exercise. AB - OBJECTIVE: To assess the test-retest reliability (repeatability) of Borg's 6-20 rating of perceived exertion (RPE) scale using a more appropriate statistical technique than has been employed in previous investigations. The RPE scale is used widely in exercise science and sports medicine to monitor and/or prescribe levels of exercise intensity. The "95% limits of agreement" technique has recently been advocated as a better means of assessing within-subject (trial to trial) agreement than traditional indicators such as Pearson and intraclass correlation coefficients. METHODS: Sixteen male athletes (mean (SD) age 23.6 (5.1) years) completed two identical multistage (incremental) treadmill running protocols over a period of two to five days. RPEs were requested and recorded during the final 15 seconds of each three minute stage. All subjects successfully completed at least four stages in each trial, allowing the reliability of RPE responses to be examined at each stage. RESULTS: The 95% limits of agreement (bias +/- 1.96 x SDdiff) were found to widen as exercise intensity increased: 0.88 (2.02) RPE units (stage 1), 0.25 (2.53) RPE units (stage 2), -0.13 (2.86) RPE units (stage 3), and -0.13 (2.94) RPE units (stage 4). Pearson correlations (0.81, 0.72, 0.65, and 0.60) and intraclass correlations (0.82, 0.80, 0.77, and 0.75) decreased as exercise intensity increased. CONCLUSIONS: These findings question the test-retest reliability of the RPE scale when used to monitor subjective estimates of exercise intensity in progressive (or graded) exercise tests. PMID- 10522638 TI - Hip and ankle range of motion and hip muscle strength in young female ballet dancers and controls. AB - OBJECTIVES: To compare the hip and ankle range of motion and hip muscle strength in 8-11 year old novice female ballet dancers and controls. METHODS: Subjects were 77 dancers and 49 controls (mean (SD) age 9.6 (0.8) and 9.6 (0.7) years respectively). Supine right active hip external rotation (ER) and internal rotation (IR) were measured using an inclinometer. A turnout protractor was used to assess standing active turnout range. The measure of ER achieved from below the hip during turnout (non-hip ER) was calculated by subtracting hip ER range from turnout range, and hip ER:IR was derived by dividing ER range by IR range. Range of right weight bearing ankle dorsiflexion was measured in a standing lunge using two methods: the distance from the foot to the wall (in centimetres) and the angle of the shank to the vertical via an inclinometer (in degrees). Right calf muscle range was measured in weight bearing using an inclinometer. A manual muscle tester was used to assess right isometric hip flexor, internal rotator, external rotator, abductor, and adductor strength. RESULTS: Dancers had less ER (p<0.05) and IR (p<0.01) range than controls but greater ER:IR (p<0.01). Although there was no difference in turnout between groups, the dancers had greater non hip ER. Dancers had greater range of ankle dorsiflexion than controls, measured in both centimetres (p<0.01) and degrees (p<0.05), but similar calf muscle range. After controlling for body weight, controls had stronger hip muscles than dancers except for hip abductor strength which was similar. Regression analyses disclosed a moderate relation between turnout and hip ER (r = 0.40). There were no significant correlations between range of motion and training years and weekly training hours. CONCLUSIONS: Longitudinal follow up will assist in determining whether or not hip and ankle range in young dancers is genetically fixed and unable to be improved with further balletic training. PMID- 10522639 TI - Correlation of medial/lateral rotation of the humerus with glenohumeral translation. AB - OBJECTIVES: To correlate glenohumeral translation in the anterior/posterior direction with medial and lateral rotation of the humerus. In addition, the length of the anterior and posterior component of the glenohumeral capsuloligamentous complex was varied in order to gain insight into the contribution of each component to limiting translation. All measurements were made with the humerus positioned at 90 degrees of abduction and 0 degrees of flexion/ extension. METHODS: Six fresh cadaveric shoulders were used. Each scapula was mounted in a cement pot to rest it in its correct anatomical position. Seven tests were carried out on each shoulder. A series of measurements of translation of the humerus in the anterior direction and posterior direction were taken at 20 degrees intervals of lateral rotation and then at 20 degrees intervals of medial rotation until the limit of lateral or medial rotation had clearly been reached (test 1). The capsuloligamentous complex was then incised and a beaded chain and catches were sutured across the joint to mimic the capsuloligamentous complex at different lengths (tests 2 to 7). RESULTS/CONCLUSIONS: (a) When the glenohumeral capsuloligamentous complex is intact, the humerus translates maximally in the glenoid (between 20 and 30 mm) when the humerus is between 40 degrees and 100 degrees of lateral rotation. (b) As the glenohumeral capsuloligamentous complex increases in length, so does the extent of translation. (c) In medial rotation, the length of the posterior capsule, rather than the length of the anterior capsule, has the greater effect on anterior/posterior translation. (d) In lateral rotation the length of the anterior capsule, rather than the length of the posterior capsule, has the greater effect on anterior/posterior translation. (e) The glenohumeral ligamentous complex acts more as a cuff, enclosing the joint, rather than as a sling, as is commonly thought. PMID- 10522640 TI - Prediction of peak oxygen uptake in chronic fatigue syndrome. AB - OBJECTIVES: To establish a simple, valid, and acceptable method of predicting peak oxygen uptake (VO2peak) in patients with chronic fatigue syndrome (CFS), which could provide a basis for subsequent exercise prescription at an appropriate intensity as part of a clinical rehabilitation programme. METHODS: A total of 130 patients who met UK research criteria for CFS were taken from consecutive referrals for chronic fatigue to the University Department of Medicine at Withington Hospital, Manchester. VO2peak was determined using an incremental graded exercise test to exhaustion. Respiratory gas exchange, work rate, and heart rate were monitored throughout. RESULTS: In all patients, VO2peak was found to correlate strongly and significantly with peak work rate (WRpeak) during testing (r2 = 0.88, p<0.001). In patients who exercised for longer than two minutes (n = 119), regression analysis established the relation as Vo2peak = 13.1 x WRPpeak + 284, where VO2 is given in ml/min and WR in W. The mean error between the measured VO2peak and the predicted value was 10.7%. The relation between increase in work rate and oxygen uptake across the group was highly significant (r2 = 0.87, p<0.001), and given as VO2increase = 12.0 x WRincrease, this value being similar to that expected for healthy individuals. Almost all (97%) subjects reported no exacerbation of symptoms after maximal exercise testing. CONCLUSIONS: Using a simple to administer maximal exercise test on a cycle ergometer, it is possible to predict accurately the VO2peak of a patient with CFS from peak work rate alone. This value can then be used as an aid to setting appropriate exercise intensity for a rehabilitation programme. The increase in VO2 per unit increase in workload was consistent with that expected in healthy individuals, suggesting that the physiological response of the patients measured here was not abnormal. Contrary to the belief of many patients, maximal exercise testing to the point of subjective exhaustion proved to be harmless, with no subjects suffering any lasting deterioration in their condition after assessment. PMID- 10522641 TI - Sledging related spinal injuries and fracture patterns: a report on five cases. AB - The cases are reported of five patients who presented to The Queens Medical Centre, Nottingham after a sledging accident. All five patients presented consecutively during the first weekend in 1997 having sustained the accident in the same public park. The mechanism and subsequent fracture type is described for each. These injuries are preventable, and increasing public awareness of the risk of sledging in public places may reduce the incidence. PMID- 10522642 TI - General practitioners' training for, interest in, and knowledge of sports medicine and its organisations. AB - OBJECTIVES: To assess the training and interest of a group of general practitioners in the area of sport and exercise medicine, and the organisations representing the specialty. DESIGN: A postal questionnaire using a Likert scale in a previously piloted set of questions. SUBJECTS: 275 general practitioners registered with the Northampton Regional Health Authority. MAIN OUTCOME MEASUREMENTS: Responses to questions designed to assess training and interest in sport and exercise medicine. RESULTS: A response rate of 87.6% was achieved. It was found that 72.7% of the responding general practitioners felt inadequately trained to practice sport and exercise medicine. Some 76.0% would welcome more training and 36.4% felt that their undergraduate orthopaedic training was of no use in primary care. Many (63.6%) of the general practitioners believed that the current NHS cannot sustain sport and exercise medicine, and there was uncertainty as to whether it is currently a recognised specialty, although 60.4% felt that it should be. General practitioners listed lack of facilities (53.1%), lack of training (42.9%), and lack of time (38.2%) as the main problems in practicing sport and exercise medicine in primary care within the current NHS. CONCLUSIONS: General practitioners feel undertrained in sport and exercise medicine at both undergraduate and post-graduate level; they have a perceived need for more training and show an interest in the subject. There is scope for improving the value of undergraduate orthopaedic training. General practitioners wish to see sport and exercise medicine recognised as an NHS specialty but fear that this is not sustainable under current conditions. There is confusion among general practitioners about the current sport and exercise medicine organisations. PMID- 10522643 TI - Heart rate of a rhinoceros running a marathon. PMID- 10522644 TI - Polychlorinated dibenzo-p-dioxins, polychlorinated dibenzofurans, and coplanar polychlorinated biphenyls in breast milk from two cities in Ukraine. AB - Substantial environmental pollution has been alleged in Ukraine, but little information is available to allow an assessment of the possible impact on humans. To help remedy this lack of information, it was of interest to investigate whether certain polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), or coplanar polychlorinated biphenyls (PCBs) were elevated in people from Ukraine. Samples of breast milk were obtained from 200 women from the cities of Kyiv and Dniprodzerzhinsk; Kyiv is the capital and Dniprodzerzhinsk is a highly industrialized city. The samples were combined into four pools by city and age, and analyzed for 7 PCDDs, 10 PCDFs, and 2 coplanar PCBs (126 and 169). The total of the measured PCDDs, expressed as toxic equivalent, ranged from 5.1 to 7.6 pg/g lipid; for PCDFs from 3.6 to 5.2, and for PCBs from 11 to 18 pg/g lipid. Results from the two cities were similar; older women had slightly higher concentrations than did younger women. Levels of these compounds seen in Ukraine were similar to or lower than those seen in other recent studies from European and Asian countries. PMID- 10522645 TI - Increased airway hyperresponsiveness and inflammation in a juvenile mouse model of asthma exposed to air-pollutant aerosol. AB - Asthma and its exacerbation by air pollution are major public health problems. This investigation sought to more precisely model this disorder, which primarily affects children, by using very young mice. The study first attempted to create allergic airway hypersensitivity in neonatal mice and to determine if physiologic testing of airway function was possible in these small animals. Neonatal mice were sensitized by i.p. injection of ovalbumin (OVA, 5 microg) and alum (1 mg) at 3 and 7 d of age. One week later, mice were challenged by allergen nebulization (3% OVA in PBS, 10 min/d, d 14-16). OVA-exposed mice showed: (1) increased airway hyperresponsiveness (AHR) to methacholine by whole-body plethysmography; (2) eosinophilia in bronchoalveolar lavage (BAL) fluid; (3) airway inflammation using histopathology techniques; and (4) elevated serum anti-OVA immunoglobulin E. Hence, these neonatal mice were successfully sensitized and manifested "asthmatic" responses after allergen challenge. Experiments were conducted to investigate the effect of one surrogate for ambient air particles, residual oil fly ash (ROFA), on this juvenile asthma model. Aerosolized ROFA leachate (supernatant of 50 mg/ml, 30 min, on d 15) had no marked effect alone, but caused a significant increase in AHR and airway inflammation in OVA-sensitized and challenged mice. This synergistic effect was abrogated by the antioxidant dimethylthiourea (DMTU, 3 mg/kg mouse, i.p.). This model may be useful to study air pollution-mediated exacerbation of asthma in children. PMID- 10522646 TI - Increased proinflammatory cytokines production by ergotamine in male BALB/c mice. AB - The ergopeptine alkaloid ergotamine (ET) mimics the effects of ergopeptine alkaloids found in endophyte-infected (E+) fescue forage considered causative for fescue toxicosis. Altered immune capacity, compromised intake and thermoregulation, and inflammatory changes are observed in fescue toxicosis. Taken together, these suggest the cytokine pattern may be altered by ergot alkaloids. Thus, the objective of this study was to determine whether major splenocyte-derived cytokines--interleukin 2 (IL-2), interleukin 4 (IL-4), interferon gamma (IFN-gamma)--and macrophage-derived cytokines--interleukin 1beta, (IL-1beta), interleukin 6 (IL-6), and tumor necrosis factor alpha (TNF alpha)--were affected by ergotamine. Two sets of male BALB/c mice (n = 5/treatment) were treated with ergotamine tartrate (s.c.) for 10 d at doses of 0 (control), 0.4, 2, 10, or 50 mg/kg body weight. Twenty-four hours after the last treatment, splenocytes (S) were isolated from one set of animals and macrophages (Mphi) from the other set for determination of IL-2, IL-4, INF-gamma, and IL 1beta, IL-6, TNF-alpha, respectively. Following activation with 5 microg/ml concanavalin A (Con A) (S) and 10 microg/ml lipopolysaccharide (LPS) (Mphi), cells were incubated for 48 and 24 h, respectively, and supernatants were collected and assayed for respective cytokines by enzyme-linked immunosorbent assay (ELISA). Additionally, differential white blood cell (WBC) counts were performed and the neutrophil (N):lymphocyte (L) ratio calculated. Ergotamine treatment significantly increased IL-6 levels at the 2.0 mg/kg dose and greater and TNF-alpha at the highest dose. There was no treatment effect on IL-1beta, IL 2, IL-4, and IFN-gamma. Also, no effect was observed upon total and differential WBC counts as well as N:L ratio. Ergotamine affected the proinflammatory cytokine IL-6, and this increase may contribute to fescue tosicosis. PMID- 10522647 TI - Penetration of chloroform, trichloroethylene, and tetrachloroethylene through human skin. AB - In vitro dermal absorption was measured for three volatile organic compounds in dilute aqueous solution through freshly prepared and previously frozen human skin. The permeability coefficients at 26 degrees C for chloroform (0.14 cm/h) and trichloroethylene (0.12 cm/h) were similar but much larger than that for tetrachloroethylene (0.018 cm/h). Storage of the skin at -20 degrees C did not significantly affect the penetration of these chemicals. The dermal absorption of chloroform through freshly prepared human skin was not changed significantly by pretreatment of the skin with commonly used consumer products (moisturizer, baby oil, insect repellent, sunscreen); however, the permeability coefficient was found to increase from 0.071 cm/h at 11 degrees C to 0.19 cm/h at 50 degrees C. These data suggest that exposure estimates for chloroform and other contaminants in water should consider the appropriate exposure scenario to properly assess the dermal dose. PMID- 10522649 TI - Massive outbreak of Escherichia coli O157:H7 infection in schoolchildren in Sakai City, Japan, associated with consumption of white radish sprouts. AB - In July 1996, an outbreak of Escherichia coli O157:H7 infection occurred among schoolchildren in Sakai City, Osaka, Japan. This outbreak developed in 13 North East District and 34 Middle-South District elementary schools in the city. All children hospitalized on July 17-19 had presented on July 8 (North-East District) and July 9 (Middle-South District). School lunches served on July 1 and 8 in the North-East District and on July 1, 4, 8, and 9 in the Middle-South District were proposed by a food consumption study to be associated with infection. White radish sprouts from a single farm were the only uncooked food common to the most highly implicated meals on the involved days in two school districts (sweet and sour chicken with lettuce on July 8 in the North-East District and chilled Japanese noodles on July 9 in the Middle-South District). Two incidents of E. coli O157:H7 in neighboring areas were also related to white radish sprouts from the farm. The pulse-field gel electrophoresis patterns of isolates from patients in these two districts and the neighboring areas were identical. Thus, it was concluded that the cause of the outbreak was the white radish sprouts shipped on July 7-9 from one particular farm. PMID- 10522648 TI - The PPAR activator docosahexaenoic acid prevents acetaminophen hepatotoxicity in male CD-1 mice. AB - Acetaminophen (APAP)-induced hepatocellular necrosis can be prevented by treatment with peroxisome proliferators. This protection is associated with lowered protein arylation and glutathione depletion in mice. Peroxisome proliferators have been shown to activate nuclear receptors. These receptors, termed peroxisome proliferator activated receptors (PPARs), can also be activated by free fatty acids. This study was designed to determine if treatment with the PPAR activator docosahexaenoic acid (DHA) would also lower APAP toxicity. Male CD 1 mice received 250 mg DHA/kg or 500 mg clofibrate (CFB)/kg, i.p., for 5 d. Controls received corn oil vehicle, i.p. After overnight fasting, mice received 800 mg APAP/kg, p.o. At 24 h after APAP, hepatotoxicity was evident in control mice by elevated plasma sorbitol dehydrogenase activity (SDH) and histologic evidence of hepatic degeneration and necrosis. As expected, CFB pretreatment significantly decreased this. Similarly, DHA protected against APAP-induced hepatotoxicity at 24 h after challenge. However, treatment with DHA did not increase hepatic glutathione prior to APAP, as previously shown with CFB. Interestingly, DHA did not increase palmitoyl coenzyme A (CoA) oxidase activity or other biochemical parameters associated with peroxisome proliferation after 5 d of treatment at 250 mg/kg. No significant alterations in microsomal APAP glucuronidation or cytochrome P-450-mediated bioactivation were detected either. Collectively, these results show that DHA also prevents APAP-induced hepatotoxicity at 24 h after challenge. However, the association between resistance against APAP-induced liver injury, PPAR activation, and peroxisome proliferation is not clearly understood. PMID- 10522650 TI - Public health in crisis: outbreaks of Escherichia coli O157:H7 infections in Japan. PMID- 10522652 TI - Exposure to natural fluoride in well water and hip fracture: a cohort analysis in Finland. AB - In the retrospective cohort study based on record linkage, the authors studied a cohort of persons born in 1900-1930 (n = 144,627), who had lived in the same rural location at least from 1967 to 1980. Estimates for fluoride concentrations (median, 0.1 mg/liter; maximum, 2.4 mg/liter) in well water in each member of the cohort were obtained by a weighted median smoothing method based on ground water measurements. Information on hip fractures was obtained from the Hospital Discharge Registry for 1981-1994. No association was observed between hip fractures and estimated fluoride concentration in the well water in either men or women when all age groups were analyzed together. However, the association was modified by age and sex so that among younger women, those aged 50-64 years, higher fluoride levels increased the risk of hip fractures. Among older men and women and younger men, no consistent association was seen. The adjusted rate ratio was 2.09 (95% confidence interval: 1.16, 3.76) for younger women who were the most exposed (>1.5 mg/liter) when compared with those who were the least exposed (< or =0.1 mg/liter). The results suggest that fluoride increases the risk of hip fractures only among women. PMID- 10522651 TI - Prospective study of snoring and risk of hypertension in women. AB - Whether snoring increases the risk of hypertension remains unclear. The authors examined the association between snoring and risk of hypertension in a cohort of 73,231 US female nurses aged 40-65 years and without diagnosed cardiovascular disease or cancer in 1986. Blood pressure levels and physician-diagnosed hypertension were self-reported through validated questionnaires. During 8 years of follow-up, 7,622 incident cases of physician-diagnosed hypertension were reported. Older age, smoking, body mass index, waist circumference, waist-hip ratio, weight gain, less physical activity, and sleeping on the back were directly associated with regular snoring. After adjustment for age, body mass index, waist circumference, and other covariates, snoring was associated with a significantly higher prevalence of hypertension at baseline (odds ratio = 1.22, 95% confidence interval (CI): 1.16, 1.27 for occasional snoring and odds ratio = 1.43, 95% CI: 1.33, 1.5 for regular snoring). In prospective analyses using incident cases of hypertension as the outcome, the multivariate relative risks of hypertension were 1.29 (95% CI: 1.22, 1.37) for occasional snoring and 1.55 (95% CI: 1.42, 1.70) for regular snoring. In addition, snoring was associated with significantly higher systolic and diastolic blood pressure levels. These data suggest that snoring may increase risk of hypertension in women, independent of age, body mass index, waist circumference, and other lifestyle factors. PMID- 10522653 TI - Risk factors for back injury in 31,076 retail merchandise store workers. AB - Risk factors for work-associated strain or sprain back injuries were investigated in a cohort of 31,076 material handlers from 260 retail merchandise stores in the United States. The workers studied were those with significant material-handling responsibilities--daily lifting and movement of merchandise. Workers in jobs with the greatest physical work requirements had an injury rate of 3.64 per 100 person years versus 1.82 in workers with lesser work requirements. The unadjusted injury rate for males was 3.67 per 100 person-years compared with 2.34 per 100 person years for females, but the excess for males was confounded by higher physical work requirements for men in the stocker/receiver job category. The injury rate ratio for short versus long duration of employment was 3.53 (95% confidence interval: 2.90, 4.30); for medium versus long duration of employment, it was 1.38 (95% confidence interval: 1.18, 1.62). The elevated rate ratios were maintained when the data were stratified by subsets with different rates of turnover. The results suggest that workers with the greatest physical work requirements and those with the shortest duration of employment are at the highest risk of back injuries. However, selection forces causing worker turnover within this cohort of active workers are not well characterized and have the potential to bias the measures for time-related factors such as duration of employment. PMID- 10522654 TI - Do elderly women have more physical disability than men do? AB - This study investigated whether the commonly observed higher prevalence of physical disability among women is due to higher incidence rates or to other factors such as selective mortality or poor recovery. Methods included observed measures of prevalent lower body physical disability and potential risk factors at baseline (1989-1991) and 4-year follow-up of 2,025 community-dwelling adults aged 55 years and older in Marin County, California. Incidence, recovery, and mortality rates were determined at the follow-up examination. Results indicated that women had higher age-specific and age-adjusted prevalence rates at both examinations (odds ratios = 1.66 and 1.60, p<0.001) but that incidence rates were not significantly different (odds ratio = 1.12, 95% confidence interval: 0.77, 1.64). In the classic formulation, prevalence = incidence x duration, the higher prevalence rates in women could not be due to a higher incidence rate, but could be explained by longer duration due to lower recovery and mortality rates in women. Incident physical disability was predicted by prevalent chronic illnesses, poor vision, obesity, physical inactivity, poor memory, fewer social activities, and higher depression scores, but not by sex. Prevention efforts should concentrate on reducing known risk factors in both men and women and on promoting higher recovery rates among women. PMID- 10522655 TI - Influence of education on risk of hysterectomy before age 45 years. AB - In a population of 4,278 women aged 36-44 years identified from Massachusetts Town Books between 1995 and 1997, relative to more highly educated women, those who completed only their high school education were about four times more likely (95% confidence interval: 1.8, 10.8) to have undergone hysterectomy, regardless of smoking status, body mass index, or medical indications for the hysterectomy. Possible explanations are that less educated women may delay seeking health services for gynecologic problems resulting in hysterectomy as the last treatment option or may be offered hysterectomy as the primary treatment option by their physicians. Future studies should assess diagnoses that lead to hysterectomy and the interval between onset of the condition and delivery of medical care. PMID- 10522656 TI - Role of transurethral resection of the prostate in population-based prostate cancer incidence rates. AB - The extensive pool of asymptomatic prostate disease in the population, which increases substantially with age, suggests that the frequent use of transurethral resection of the prostate (TURP) in recent decades has had a large effect on prostate cancer incidence. The authors identified the effect of TURP-detected prostate cancer on the observed incidence rates between 1973 and 1993 for men aged 65 years and older. They linked population-based cancer registry data from the Surveillance, Epidemiology, and End Results Program to Medicare records between 1986 and 1993 to determine whether a TURP occurred sufficiently close to the time of a prostate cancer diagnosis for them to assume that it led to the diagnosis. TURP-detected cases prior to 1986 were calculated using an indirect method that involved multiplying the TURP procedure rate in the general population (from the National Hospital Discharge Survey) by estimates of the proportion of TURPs resulting in a prostate cancer diagnosis (from Medicare data and the literature). TURP explained much of the observed increase in overall prostate cancer incidence between 1973 and 1986 and possibly all of it in men aged 70 years and older. However, its influence on the trend and overall magnitude of the rates diminished between 1987 and 1993. The changing role of TURP in detecting prostate cancer is attributed to changes in medical technology and screening practices. The declining influence of TURP on prostate cancer incidence is likely to have continued beyond the study period due to the recent introduction and increasing use of medications for treating obstructive uropathy. PMID- 10522657 TI - Risk of breast cancer according to use of antidepressants, phenothiazines, and antihistamines. AB - In laboratory studies, some antidepressants caused increased growth of mammary tumors. The relation of use of these drugs to the development of breast cancer was examined in a hospital-based case-control study. Information, including lifetime medication history, was collected by interview from 5,814 women with primary breast cancer diagnosed within the previous year, 5,095 women with primary malignancies of other sites, and 5,814 women with other conditions. Relative risks were estimated by using unconditional multiple logistic regression for regular use (> or =4 days per week for > or =4 weeks beginning > or =1 year before admission) of antidepressants and structurally similar drugs. With reference to never use of each drug, relative risks were statistically compatible with 1.0 for selective serotonin reuptake inhibitors (SSRI), tricyclics, other antidepressants, phenothiazines, and antihistamines; results were very similar using both control groups. There were no significant increases in risk for any category of regular use, stratified according to cumulative duration of use or time interval since the most recent use or for any individual drug within the broader classes. However, the estimate for regular SSRI use in the previous year, 1.8, was of borderline statistical significance (95% confidence interval: 1.0, 3.3). The findings do not support an overall association between the use of antidepressants, phenothiazines, or antihistamines and breast cancer. However, the results for SSRIs are not entirely reassuring. PMID- 10522659 TI - Potential misinterpretation of the case-only study to assess gene-environment interaction. AB - Novel epidemiologic study designs are often required to assess gene-environment interaction. A design using only cases, without controls, is one of several approaches that have been proposed as more efficient alternatives to the typical random sampling of cases and controls. However, it has not been pointed out that a case-only analysis estimates a different interaction parameter than does a traditional case-control analysis: The latter typically estimates departure from multiplicative population odds or rate ratios, depending on the method of control selection, while the former estimates departure from multiplicative risk ratios if genotype and environmental exposure are not associated in the population. These parameters are approximately equal if the disease risk is small at all levels of the study variables. The authors quantify the impact of allowing for higher disease risk among gene carriers, a relevant situation when the gene under study is highly penetrant. Their findings show that the cross-product ratio computed from case-only data may be substantially smaller than the odds ratio computed from case-control data and may therefore underestimate either the population odds or the rate ratio. Thus, to avoid misinterpretation of interaction parameters estimated from case-only data, the definition of multiplicative interaction should be made explicit. PMID- 10522658 TI - Lifestyle and colon cancer: an assessment of factors associated with risk. AB - Studies of the etiology of colon cancer indicate that it is strongly associated with diet and lifestyle factors. The authors use data from a population-based study conducted in northern California, Utah, and Minnesota in 1991-1995 to determine lifestyle patterns and their association with colon cancer. Data obtained from 1,993 cases and 2,410 controls were grouped by using factor analyses to describe various aspects of lifestyle patterns. The first five lifestyle patterns for both men and women loaded heavily on dietary variables and were labeled: "Western," "moderation," "calcium/low-fat dairy;" "meat and mutagens," and "nibblers, smoking, and coffee." Other important lifestyle patterns that emerged were labeled "body size," "medication and supplementation," "alcohol," and "physical activity." Among both men and women, the lifestyle characterized by high levels of physical activity was the most marked lifestyle associated with colon cancer (odds ratios = 0.42, 95% confidence interval: 0.32, 0.55 and odds ratio = 0.52, 95% confidence interval: 0.39, 0.69, for men and women, respectively) followed by medication and supplementation (odds ratio = 1.68, 95% confidence interval: 1.29, 2.18 and odds ratio = 1.63, 95% CI 1.23, 2.16, respectively). Other lifestyles that were associated with colon cancer were the Western lifestyle, the lifestyle characterized by large body size, and the one characterized by calcium and low-fat dairy. Different lifestyle patterns appear to have age- and tumor site-specific associations. PMID- 10522660 TI - Recovering true risks when multilevel exposure and covariables are both misclassified. AB - The authors extend previous results on nondifferential exposure misclassification to the situation in which multilevel exposure and covariables are both misclassified. They show that if misclassification is nondifferential and the predictive value matrices are independent of other predictor variables it is possible to recover the true relative risks as a function of the biased estimates and the misclassification matrices alone. If the covariable is a confounder, the true relative risks may be recovered from the apparent relative risks derived from misclassified data and the misclassification matrix for the exposure variable with respect to its surrogate. If the covariable is an effect modifier, the true relative risk matrix may be recovered from the apparent relative risk matrix and misclassification matrices for both the exposure variable with respect to its surrogate and the covariable with respect to its surrogate. By varying the misclassification matrices, the sensitivity of published relative risk estimates to different patterns of misclassification can be analyzed. If it is not possible to design a study protocol that is free of misclassification, choosing surrogate variables whose predictive value is constant with respect to other predictors appears to be a desirable design objective. PMID- 10522661 TI - Re: "Use of census-based aggregate variables to proxy for socioeconomic group: evidence from national samples". PMID- 10522662 TI - Topical imiquimod therapy for chronic giant molluscum contagiosum in a patient with advanced human immunodeficiency virus 1 disease. PMID- 10522663 TI - Mohs micrographic surgery: a pathologist's view. PMID- 10522664 TI - A systematic review of treatment modalities for primary basal cell carcinomas. AB - OBJECTIVE: To systematically review the literature for studies reporting on recurrence rates of basal cell carcinomas (BCCs) after different therapies. DESIGN: We reviewed all studies published in English, French, German, Dutch, Spanish, or Italian between 1970 and 1997 that prospectively examined recurrence rates for at least 50 patients with primary BCCs observed for at least 5 years after treatment with Mohs micrographic surgery, surgical excision, curettage and electrodesiccation, cryosurgery, radiotherapy, immunotherapy with interferon or fluorouracil, or photodynamic therapy. SETTING: Department of Dermatology, University Hospital Maastricht, Maastricht, the reference center for dermatologic oncology and Mohs micrographic surgery in the Netherlands. MAIN OUTCOME MEASURES: The recurrence rates after different therapies for BCCs, resulting in the development of guidelines for the treatment of these disorders. RESULTS: Of 298 studies found in several electronic databases, only 18 met the requirements and could be used for analysis. Tumors treated with Mohs micrographic surgery show the lowest recurrence rates after 5 years, followed in order by those treated with surgical excision, cryosurgery, and curettage and electrodesiccation. CONCLUSIONS: Recurrence rates for different therapies could not be compared because of a lack of uniformity in the method of reporting, so evidence-based guidelines could not be developed. We surmise that Mohs micrographic surgery should be used mainly for larger, morphea-type BCCs located in danger zones. For smaller BCCs of the nodular and superficial types, surgical excision remains the first treatment of choice. Other treatment modalities can be used in patients in whom surgery is contraindicated. Immunotherapy and photodynamic therapy are still investigative. PMID- 10522665 TI - Lack of consensus among experts on the choice of UV therapy for psoriasis. AB - CONTEXT: Each year tens of thousands of patients in the United States are treated with UV-B radiation or psoralen plus UV-A radiation (PUVA) for a variety of skin disorders. Although PUVA is generally considered more effective, it is also more toxic and more expensive. The degree of consensus among experts in prescribing these alternative treatments has not been quantified. OBJECTIVES: To quantify variation among specialty clinics in the type of ultraviolet therapy used to treat specific skin conditions and assess factors associated with the use of specific treatments. DESIGN: Survey conducted during two 2-week periods in the late fall of 1994 and early spring of 1995. SETTING: Thirty-nine specialty clinics in 17 US geographic areas in 14 states and Washington, DC. PARTICIPANTS: A total of 3401 patients treated with UV radiation one or more times. OUTCOME MEASURES: Type of UV therapy used and indications for treatment, age, sex, number of patients treated, and geographic location of each clinic. RESULTS: The proportion of patients at each center treated with PUVA ranged from 0% to 93% (mean, 41%). Clinic size and geographic location, demographic characteristics of the patients, and diagnosis did not explain these large intercenter differences. CONCLUSIONS: Among specialized clinics, there is little consistency in the use of alternative therapies, which differ substantially in safety and cost, but whose relative efficacy is not well quantified. There is a lack of consensus among experts about the circumstances in which the greater risks and costs of PUVA are outweighed by its possibly greater efficacy, especially in the treatment of psoriasis. PMID- 10522666 TI - Birt-Hogg-Dube syndrome: a novel marker of kidney neoplasia. AB - BACKGROUND: Birt-Hogg-Dube syndrome (BHD) is a dominantly inherited predisposition for development of fibrofolliculomas, trichodiscomas, and acrochordons. Concurrent internal tumors, such as colonic polyps and renal carcinoma, have been described in patients with BHD. OBJECTIVE: To evaluate kindreds with familial renal tumors for cutaneous manifestations of BHD. DESIGN: One hundred fifty-two patients from 49 families underwent complete oral and skin examination. Skin lesions were identified by their clinical appearance, and the diagnosis was confirmed by results of histologic examination. Individuals underwent screening for familial renal neoplasms. SETTING: A tertiary referral research hospital. PATIENTS: Individuals with familial renal tumors and their asymptomatic at-risk relatives. MAIN OUTCOME MEASURE: We determined whether any form of renal cancer is associated BHD. RESULTS: We identified 3 extended kindreds in whom renal neoplasms and BHD appeared to segregate together. Two kindreds had renal oncocytomas and a third had a variant of papillary renal cell carcinoma. Thirteen patients exhibited BHD. Seven individuals, including a set of identical twins, had renal neoplasms and BHD. An additional 4 patients (3 deceased and not examined) in these families had renal neoplasms but not BHD. Birt-Hogg-Dube syndrome without renal neoplasms was present in 6 individuals. Thirteen patients with fibrofolliculomas and trichodiscomas presented clinically with multiple smooth skin-colored to grayish-white papules located on the face, auricles, neck, and upper trunk. Oral papules were present in 9 of 28 and achrochordons in 11 of 28 patients. Features of BHD not previously appreciated included deforming lipomas in 5, collagenomas in 4, and pulmonary cysts in 4 of 28 patients. Families with BHD did not display germline mutations in the von Hippel-Lindau gene or in the tyrosine kinase domain of the MET proto-oncogene. CONCLUSIONS: Birt-Hogg-Dube syndrome may be associated with familial renal tumors. Birt-Hogg-Dube and renal tumors segregate together in an autosomal dominant fashion. Patients with BHD and their relatives are at risk for development of renal tumors. Therefore, patients with BHD and their relatives should undergo abdominal computed tomography and renal ultrasound screening for renal tumors. PMID- 10522667 TI - Tumor burden index as a prognostic tool for cutaneous T-cell lymphoma: a new concept. AB - OBJECTIVES: To introduce a prognostic tool for cutaneous T-cell lymphoma that takes into account the tumor burden and to compare the prognostic value of this tumor burden index (TBI) with that of other prognostic factors. DESIGN: Retrospective clinical and statistical study. PATIENTS: One hundred sixteen patients with cutaneous T-cell lymphoma. METHODS: A TBI was designed that takes into account the types, numbers, and severity of skin lesions with the use of the Cox proportional hazard model. RESULTS: Models of the TBI were developed to test the relative contributions of patches, plaques, and tumors to the total tumor burden and, hence, survival time. Weighting factors reflecting the severity of each skin lesion were tested and incorporated. The best prognostic correlation was a dichotomized index with the following formula: TBI = 1 + (patches x 2) + (plaques x 2) + (tumor x 1.3), where the patches factor equals 0 if 30% or less of the skin area is involved and 1 if greater than 30% of the skin area is involved and where the plaque or tumor factor equals 1 if plaques or tumors are present. Both the TBI and TNM provided predictive information. Discrimination of survival curves and significance of differences was better for TBI (P < .001) than for TNM (P = .009). Sex was also statistically related to survival (males had a better prognosis; P < .04), whereas neither age at first symptoms (P = .35) nor age at time of diagnosis (P = .36) was of prognostic value. CONCLUSIONS: The TBI offers a simple prognostic index for the evaluation of cutaneous T-cell lymphoma. It may become a valuable tool for designing therapeutic strategies for patients according to their specific survival expectancies. However, this model is preliminary and has to be validated on a larger number of patients. PMID- 10522668 TI - Long- and short-term histological observations of congenital nevi treated with the normal-mode ruby laser. AB - OBJECTIVE: To evaluate histologically the long- and short-term changes associated with cosmetic improvement or failure of normal-mode ruby laser treatment of patients with congenital nevi. DESIGN: A biopsy of the laser-treated lesions of 10 patients with good or poor cosmetic results was performed at periods up to 8 years 10 months after treatment (mean, 4 years 9 months). Short-term findings were evaluated in 3 patients. SETTING: Ueda Setsuko Clinic and the Dermatology Unit of the Kyushu University, Fukuoka, Japan. PATIENTS: Of the 85 Japanese patients with relatively large congenital nevi who had been treated with the normal-mode ruby laser since 1990, 13 gave informed consent for biopsy and histological examination of the treated area. RESULTS: A long-term follow-up study of the 8 patients with good cosmetic results showed the presence of residual nevus cells 1.11 +/- 0.35 mm (mean +/- SD) (range, 0.63-2.05 mm) below the skin surface. Above these cells was a layer of connective tissue that formed a subtle microscopic scar that preserved the normal structure of the papillary dermis. Hair follicles were damaged at the base, and the hairs were attenuated. However, in the 2 patients with poor cosmetic results, nests of pigmented cells were commonly seen in the epidermis, and melanin was relatively abundant in basal keratinocytes. No malignant changes were observed in any patient. A short-term study in 3 patients showed damage to pigmented cells in the epidermis and upper dermis as observed following electrodesiccation. CONCLUSIONS: Multiple treatments with the normal-mode ruby laser produced immediate thermal damage to the superficial nests of nevus cells and a subsequent remodeling of the superficial connective tissue. When the thickness of the subtle microscopic scar reached 1 mm, it masked the underlying residual nevus cells and achieved a good cosmetic result. Follow-up for at least 8 years after laser treatment showed no evidence of malignant change in the treated areas. PMID- 10522669 TI - The use of tissue-engineered skin (Apligraf) to treat a newborn with epidermolysis bullosa. AB - BACKGROUND: Inherited epidermolysis bullosa (EB) is a mechanobullous disorder. The Dowling-Meara variant, a subtype of EB, is characterized by widespread blister formation that may include the oral cavity and nails. Many patients with the Dowling-Meara phenotype are at increased risk of sepsis and death during infancy. The treatment of EB is generally supportive. The tissue-engineered skin used (Apligraf) is a bilayered human skin equivalent developed from foreskin. It is the only Food and Drug Administration-approved skin equivalent of its kind. It is approved for the treatment of venous ulcers of the lower extremities. It has also been used to treat acute wounds, such as graft donor sites and cancer excision sites. OBSERVATION: To our knowledge, we describe the first case in which a newborn with EB, Dowling-Meara variant, was treated with bilayered tissue engineered skin. The areas treated with the tissue-engineered skin healed faster than the areas treated with conventional therapy. Most of the areas treated with tissue-engineered skin have remained healed, without developing new blisters. These areas appear to be more resistant to trauma. CONCLUSIONS: Our early success with tissue-engineered skin in this patient may have a significant impact on the future treatment of neonates with EB simplex. Future studies are needed to determine if the beneficial effects of tissue-engineered skin are reproducible in other neonates with EB simplex and in patients of all ages with different subtypes of EB. PMID- 10522670 TI - Acquired progressive kinking of the hair: clinical features, pathological study, and follow-up of 7 patients. AB - BACKGROUND: Acquired progressive kinking of the hair (APKH) is a relatively rare condition, with fewer than 20 cases reported in the literature. Whether APKH is a separate entity or a variety of androgenetic alopecia is still controversial. This study reviews the clinical and pathological features and long-term follow-up of 7 patients with APKH. OBSERVATIONS: Since January 1989, we have diagnosed APKH in 7 males aged 15 to 22 years. All patients had strong family history for androgenetic alopecia. Hair kinking affected the frontotemporal region and/or the vertex where the hair appeared curly, frizzy, and lusterless. The pathological features of the affected scalp were consistent with the diagnosis of the early stages of androgenetic alopecia. In all patients, APKH evolved into androgenetic alopecia during the follow-up period. Mean follow-up was 4.5 years (range, 2-9 years). Treatment with topical minoxidil did not prevent development of hair thinning in the scalp areas affected by hair kinking. CONCLUSIONS: The term acquired progressive kinking of the hair encompasses a number of conditions characterized by acquired curling of the scalp hair. Acquired hair kinking on the androgen-dependent areas of the scalp represents a modality of onset of androgenetic alopecia associated with poor prognosis. PMID- 10522671 TI - Multiple hereditary infundibulocystic basal cell carcinomas: a genodermatosis different from nevoid basal cell carcinoma syndrome. AB - BACKGROUND: Infundibulocystic basal cell carcinoma is a recently described distinctive clinicopathologic variant of basal cell carcinoma. Histopathologic differential diagnosis among infundibulocystic basal cell carcinoma, trichoepithelioma, and basaloid follicular hamartoma has generated controversy in the literature. OBSERVATIONS: Members of 2 families with multiple infundibulocystic basal cell carcinomas are described. Each patient showed multiple papular lesions, mostly located on the face. No patient showed palmar pits or jaw cysts. Forty-two cutaneous lesions from 5 patients were studied histopathologically. Thirty-nine lesions were infundibulocystic basal cell carcinomas. This clinicopathologic variant of basal cell carcinoma consists of a relatively well-circumscribed basaloid neoplasm composed of buds and cords of neoplastic cells arranged in anastomosing fashion and with scant stroma. Some of the neoplastic cords contain tiny infundibular cysts filled by cornified cells with abundant melanin. Linkage analysis in family 2 was performed using polymorphic markers (D9S196, D9S280, D9S287, and D9S180), and the affected members shared the same haplotype. Loss of heterozygosity analysis was performed in 2 affected members of this family from whom tumoral DNA was available, and although these individuals were constitutively heterozygous for D9S196, they did not show loss of heterozygosity for this marker in their neoplasms. CONCLUSIONS: Multiple hereditary infundibulocystic basal cell carcinomas represent a distinctive genodermatosis different from multiple hereditary trichoepitheliomas and nevoid basal cell carcinoma syndrome. We propose clinical and histopathologic criteria to distinguish infundibulocystic basal cell carcinoma from trichoepithelioma, basaloid follicular hamartoma, and folliculocentric basaloid proliferation. PMID- 10522672 TI - Migratory ichthyosiform dermatosis with type 2 diabetes mellitus and insulin resistance. AB - BACKGROUND: In addition to the well-defined hereditary primary ichthyoses, many sporadic or less well-defined keratinization disorders with or without systemic manifestations have been reported. Herein we describe ichthyosiform dermatosis associated with type 2 diabetes mellitus. OBSERVATIONS: The patients were members of a large Arab family with heavy consanguinity. Eighteen members were affected with a variously severe scaly disorder. They showed migratory polycyclic keratotic scaly plaques evolving into diffuse generalized scaling or complete remission. Acanthosis nigricans-like lesions were also noted, and there was an association with type 2 diabetes mellitus. A scarcity of intercorneocyte lamellae and reduction in lamellar body contents were observed. CONCLUSIONS: We could not find a report of a similar dermatosis. Furthermore, an association between ichthyosis and diabetes has not been documented. Therefore, we believe that this may constitute a new entity. PMID- 10522673 TI - Outcome after surgical repair of junctional epidermolysis bullosa-pyloric atresia syndrome: a report of 3 cases and review of the literature. AB - BACKGROUND: Junctional epidermolysis bullosa-pyloric atresia syndrome is recognized as a distinct autosomal recessive entity. Affected infants present with skin fragility and inability to feed due to intestinal obstruction. Despite successful surgical repair of the anatomical defect, the outcome is poor owing to poor feeding, malabsorption, failure to thrive, and sepsis. OBSERVATIONS: In 70 cases of intestinal obstruction and epidermolysis bullosa reported in the medical literature and the 3 reported here, surgical intervention was attempted 51 times. In all except 16 infants, death occurred before age 11 months (mean age, 70 days). Renal involvement and continued failure to thrive accompanied the skin disease in survivors, who ranged in age from 30 days to 16 years (mean age, 4.0 years). CONCLUSIONS: The poor prognosis of this condition must be considered when decisions are made regarding surgical correction. Attempting surgical correction may be warranted in individual circumstances, but withholding surgical intervention and providing palliative support is an acceptable alternative. PMID- 10522674 TI - A glossary of modern health care economics: platitudes and principles gleaned from the first year's education of a new academic health care delivery center department head. PMID- 10522675 TI - Mohs surgery: the informed view. PMID- 10522676 TI - Dermatology, the academic medical center, and the new millennium. PMID- 10522677 TI - Blistering, scarring, and photosensitivity in a male teenager. PMID- 10522679 TI - Facial eruptions in a child. PMID- 10522678 TI - Malar rash in a child. PMID- 10522680 TI - Periorificial dermatitis and irritability in an infant. PMID- 10522681 TI - On standard dermatology definitions. PMID- 10522682 TI - Zinc in wound repair. PMID- 10522683 TI - Therapy-resistant erythrodermia-related pruritus: effective treatment with ketamine hydrochloride. PMID- 10522684 TI - Methotrexate as an adjuvant treatment for pemphigus vulgaris. PMID- 10522685 TI - Regression of in-transit melanoma of the scalp with intralesional recombinant human granulocyte-macrophage colony-stimulating factor. PMID- 10522687 TI - Correlating skin type and minimum erythema dose. PMID- 10522686 TI - Localized lipoatrophy after glatiramer acetate injection in patients with remitting-relapsing multiple sclerosis. PMID- 10522689 TI - Rubella outbreak--Westchester County, New York, 1997-1998 PMID- 10522688 TI - Low-dose 2-chlorodeoxyadenosine for the treatment of mycosis fungoides. PMID- 10522691 TI - A direct method for the formation of peptide and carbohydrate dendrimers. AB - Two new methods for the modification of PAMAM dendrimers have been developed which allow the covergent synthesis of either peptide or carbohydrate-bearing dendrimer molecules. Both methods involve condensation between hydroxylamino nucleophiles and appropriate carbonyl-bearing reaction partners. PMID- 10522690 TI - Structural basis of the dynamic mechanism of ligand binding to cyclooxygenase. AB - Molecular modeling studies performed on the two cyclooxygenase (COX) isozymes suggest that the cavity at the mouth of the active site on the membrane domain that may act as an actual binding site of COX ligands. Actual docking of different inhibitors at this site provides a structural basis to explain the dynamics of COX inhibition. PMID- 10522692 TI - Bifunctional 2-naphthyl propargylic sulfones exhibiting high DNA intercalating and alkylating activity. AB - A number of novel 2-naphthyl propargylic sulfones were synthesized as nucleic base alkylating agents. Extremely high DNA cleavage activity was observed for the sulfones with a free omega-hydroxyl group in the carbon chain in contrast to the ester conjugates possessing an additional intercalating unit. PMID- 10522693 TI - Binding affinities of 3-(3-phenylisoxazol-5-yl)methylidene-1-azabicycles to acetylcholine receptors. AB - A series of 3-(3-phenylisoxazol-5-yl)methylidene-1-azabicycles synthesized showed different binding characteristics to acetylcholine receptors depending on the substituents on the phenyl ring. Small polar substituents gave preferential binding affinity to nicotinic receptors, and large hydrophobic substituents to muscarinic receptors. PMID- 10522694 TI - The synthesis of a new class of oxytocin antagonists. AB - The synthesis of a new class of oxytocin antagonists, with significantly modified C-terminal part, is described. The chemistry of the Mitsunobu reaction was applied to obtain the key derivatives. In spite of the extensive modifications of previously described compound F792, the peptides retain biological activity as oxytocin antagonists. PMID- 10522695 TI - Synthesis and evaluation of analogs of Efavirenz (SUSTIVA) as HIV-1 reverse transcriptase inhibitors. AB - Efavirenz (SUSTIVA) is a potent non-nucleoside reverse transcriptase inhibitor. Due to the observation of breakthrough mutations of the reverse transcriptase enzyme during Efavirenz therapy, we sought to develop an optimized second generation series. To that end, SAR of the substituents on the aromatic ring was undertaken and the results are summarized here. The 5,6-difluoro (4f) and the 6 methoxy (4m) substituted benzoxazinones were determined to be equipotent, and as a result such substitution patterns will be incorporated in second generation scaffolds. PMID- 10522696 TI - Stereospecific synthesis of a GS 4104 metabolite: determination of absolute stereochemistry and influenza neuraminidase inhibitory activity. AB - The total synthesis for the determination of the absolute stereochemistry of a GS 4104 metabolite 3 is described. In addition, the influenza neuraminidase inhibitory activity of 3 and related intermediates are reported. PMID- 10522697 TI - Discovery of subtype-selective NMDA receptor ligands: 4-benzyl-1 piperidinylalkynylpyrroles, pyrazoles and imidazoles as NR1A/2B antagonists. AB - 4-Benzyl-1-[4-(1H-imidazol-4-yl)but-3-ynyl]piperidine (8) has been identified as a potent antagonist of the NR1A/2B subtype of the NMDA receptor. When dosed orally, this compound potentiates the effects of L-DOPA in the 6-hydroxydopamine lesioned rat, a model of Parkinson's disease. PMID- 10522698 TI - Synthesis and pharmacological profile of 1-aryl-3-substituted pyrrolo[3,2 c]quinolines. AB - A series of 1-aryl-3-substituted pyrrolo[3,2-c]quinolines were synthesized and evaluated for their anti-ulcer activity. While 3-substituents of pyrrolo[3,2 c]quinolines mostly affected the in vitro H+/K+ ATPase activity, 1-aryl substituents of pyrrolo[3,2-c]quinolines affected the in vivo gastric acid secretion. In addition, the compounds with good in vivo activity protected from ethanol-induced ulcer. PMID- 10522699 TI - Design, combinatorial chemical synthesis and in vitro characterization of novel urea based gelatinase inhibitors. AB - A novel series of hydroxamate/urea-based inhibitors of gelatinases has been discovered via solid-phase combinatorial chemistry. SAR of P1', P2', and P3' has been exploited and structures different from traditional succinate-based MMP inhibitors have been found. PMID- 10522700 TI - Parallel solid-phase synthesis of a model library of 7alpha-alkylamide estradiol derivatives as potential estrogen receptor antagonists. AB - The C17-THP derivative of 7alpha-(11-azidoundecanyl)-estradiol (4) was synthesized and coupled to an aminomethyl resin via a photolabile o-nitrobenzyl linker. Reduction of the azide by the Staudinger reaction to its corresponding amine followed by acylation using four activated NFmoc protected amino acids gave a first level of diversity. Subsequent deprotection of the Fmoc followed by a second acylation with five activated carboxylic acids produced, after photocleavage, a model library of twenty antiestrogen-related 7alpha-alkylamide estradiol derivatives in acceptable overall yields and very good purities. PMID- 10522701 TI - Synthesis and biological evaluation of 1,1-difluoro-2-(tetrahydro-3 furanyl)ethylphosphonic acids possessing a N9-purinylmethyl functional group at the ring. a new class of inhibitors for purine nucleoside phosphorylases. AB - 1,1-Difluoro-2-(tetrahydro-3-furanyl)ethylphosphonic acids cis-3 and trans-3 possessing a N9-purinylmethyl functionality at the ring were synthesized and tested as "multi-substrate analogue" inhibitors for purine nucleoside phosphorylases. Radical cyclization of allyic alpha,alpha-difluorophosphonate (E) 7 was applied to construct the alpha,alpha-difluorophosphonate-functionalized tetrahydrofuranyl moiety. The IC50 values of cis-3 and trans-3 for human erythrocyte PNP-catalyzed phosphorylation of inosine were determined to be 88 and 320 nM, respectively. The stereochemistry of the inhibitors was found to affect significantly the inhibitory potency. PMID- 10522702 TI - Design, synthesis and testing of amino-bicycloaryl based orally bioavailable thrombin inhibitors. AB - Replacement of the highly basic benzamidine moiety with moderate basic amino bicycloaryl moieties in a series of thrombin inhibitors related to NAPAMP provided potent enzyme inhibition and significant improvements in membrane transport and oral bioavailability. PMID- 10522703 TI - Design and synthesis of novel dihydropyridine alpha-1a antagonists. AB - A series of analogs of SNAP 5150 containing heteroatoms at C2 or C6 positions is described. Herein, we report that the presence of alkyl substituted heteroatoms at the C2(6)-positions of the dihydropyridine are well tolerated. In addition, 15 inhibited the phenylephrine induced contraction of dog prostate tissue with a Kb of 1.5 nM and showed a Kb (DBP, dogs, microg/kg)/Kb (IUP, dogs, microg/kg) ratio of 14.8/2.5. PMID- 10522704 TI - Generation of an Ugi library of phosphate mimic-containing compounds and identification of novel dual specific phosphatase inhibitors. AB - The four-component Ugi reaction was utilized to prepare a library of dipeptidic compounds in order to explore the binding requirements of the key cell cycle phosphatase, Cdc25. Several phosphate surrogates were incorporated into the Ugi product to mimic either the mono- or bis-phosphorylated substrate. PMID- 10522705 TI - Expedient synthesis of a series of N-acetyllactosamines. AB - A series of poly-N-acetyllactosamines representative of those found on complex N glycans was synthesized for use in the kinetic characterization of recently cloned glycosyltransferases. The syntheses involved the iterative addition of a selectively protected N-acetyllactosaminyl donor to an octyl alpha-D mannopyranosyl-1,6-beta-D-mannopyranoside acceptor, followed by deprotection. In addition, non-reducing galactosyl residues were removed with beta-galactosidase to furnish GlcNAc terminated compounds. In this manner tetra- to octasaccharides were efficiently produced. PMID- 10522706 TI - Interaction of tyrosyl aryl dipeptides with S. aureus tyrosyl tRNA synthetase: inhibition and crystal structure of a complex. AB - Tyrosyl aryl dipeptide inhibitors of S. aureus tyrosyl tRNA synthetase have been identified with IC50 values down to 0.5 microM. A crystal structure of the enzyme complexed to one of the inhibitors shows occupancy of the tyrosyl binding pocket coupled with inhibitor interactions to key catalytic residues. PMID- 10522707 TI - 2-chloro-3-substituted-1,4-naphthoquinone inactivators of human cytomegalovirus protease. AB - A random screening approach has identified 2-chloro-3-substituted-1,4 naphthoquinones as potent inactivators of HCMV protease. Enzyme inactivation is due to modification of Cys202. Two of the most potent compounds maintain activity against HCMV in a plaque reduction assay. PMID- 10522708 TI - Antagonism of the TXA2 receptor by seratrodast: a structural approach. AB - The crystal structure of seratrodast (AA-2414), a potent thromboxane A2 (TXA2) receptor antagonist, served as starting point to docking studies with the modeled human TXA2 receptor. This structural approach provides rational basis for the design of new antagonists within the aryl sulfonamide family. PMID- 10522709 TI - A novel method to highly versatile monomeric PNA building blocks by multi component reactions. AB - A novel approach to monomeric PNA building blocks by a solution phase Ugi multi component reaction (MCR) is described. The reaction is easily performed in 96 well plates. The products precipitate from the reaction solution and are thus obtained in high yields and purity. Those products are not amenable by another multi step synthesis in such a diversity. PMID- 10522710 TI - Structure-activity relationship in two series of aminoalkyl substituted coumarin inhibitors of gyrase B. AB - Two series of aminosubstituted coumarins were synthesised and evaluated in vitro as inhibitors of DNA gyrase and as potential antibacterials. Novel novobiocin like coumarins, 4-(dialkylamino)methylcoumarins and 4-((2 alkylamino)ethoxy)coumarins, were discovered as gyrase B inhibitors with promising antibacterial activity in vitro. PMID- 10522711 TI - Synthesis and biological evaluation of coumarincarboxylic acids as inhibitors of gyrase B. L-rhamnose as an effective substitute for L-noviose. AB - A series of novobiocin-like coumarincarboxylic acids has been prepared bearing the L-rhamnosyl moiety as the sugar portion of the molecule. The similar DNA gyrase inhibitory activity of the novel class of coumarins to that of novobiocin demonstrates that L-rhamnose can effectively replace L-noviose. Introduction of alkyl side-chains at C-5 of coumarin leads to improved in vitro antibacterial properties in the novel series. PMID- 10522712 TI - The synthesis and biological evaluation of non-peptidic matrix metalloproteinase inhibitors. AB - Novel sulfonamide matrix metalloproteinase inhibitors of general formula (9) were synthesised by a route involving a stereoselective conjugate addition reaction. Enzyme selectivity was found to be dependant on the nature of the sulfonamide substituents. Compounds (9f, 9q) are potent selective collagenase inhibitors with good oral bioavailability. PMID- 10522713 TI - 2-Naphthylcarbapenems: broad spectrum antibiotics with enhanced potency against MRSA. AB - A regioisomeric set of 2-naphthylcarbapenems featuring cationic substituents was synthesized. Optimal placement of the cationic group was found to markedly improve activity against methicillin-resistant staphylococci while maintaining a good spectrum of gram-negative activity. PMID- 10522714 TI - 15N NMR and electronic properties of S-nitrosothiols. AB - Investigation of the 15N NMR of S-nitrosothiols showed that primary and tertiary RSNOs have distinct 15N chemical shifts around 730 and 790 ppm, respectively. Using 15N NMR technique, the equilibrium constant of NO transfer between SNAP and GSH was found to be 0.74. For primary RSNOs, linear relationships exist among 15N NMR chemical shifts, reduction potentials, and the pK(a)s of their parent thiols. PMID- 10522716 TI - Six-month assessment of a phase I trial of angiogenic gene therapy for the treatment of coronary artery disease using direct intramyocardial administration of an adenovirus vector expressing the VEGF121 cDNA. AB - OBJECTIVE: To summarize the 6-month follow-up of a cohort of patients with clinically significant coronary artery disease who received direct myocardial injection of an E1-E3- adenovirus (Ad) gene transfer vector (Ad(GV)VEGF121.10) expressing the human vascular endothelial growth factor (VEGF) 121 cDNA to induce therapeutic angiogenesis. BACKGROUND: Therapeutic angiogenesis describes a novel approach to the treatment of vascular occlusive disease that uses the administration of growth factors known to induce neovascularization of ischemic tissues. METHODS: Direct myocardial injection of Ad(GV)VEGF121.10 into an area of reversible ischemia was carried out in 21 patients as an adjunct to conventional coronary artery bypass grafting (group A, n = 15) or as sole therapy using a minithoracotomy (group B, n = 6). RESULTS: No evidence of systemic or cardiac related adverse events related to vector administration was observed up to 6 months after therapy. Trends toward improvement in angina class and exercise treadmill testing at 6-month follow-up in the sole therapy group suggest the effects of this therapy are persistent for > or =6 months. CONCLUSIONS: This study suggests that direct myocardial administration of Ad(GV)VEGF121.10 appears to be well tolerated in patients with clinically significant coronary artery disease. Initiation of phase II evaluation of this therapy appears warranted. PMID- 10522715 TI - Validation of the accuracy of intraoperative lymphatic mapping and sentinel lymphadenectomy for early-stage melanoma: a multicenter trial. Multicenter Selective Lymphadenectomy Trial Group. AB - OBJECTIVE: To evaluate the multicenter application of intraoperative lymphatic mapping, sentinel lymphadenectomy, and selective complete lymph node dissection (LM/SL/SCLND) for the management of early-stage melanoma. SUMMARY BACKGROUND DATA: The multidisciplinary technique of LM/SL/SCLND has been widely adopted, but not validated in a multicenter trial. The authors began the international Multicenter Selective Lymphadenectomy Trial (MSLT) 5 years ago to evaluate the survival of patients with early-stage primary melanoma after wide excision alone versus wide excision plus LM/SL/SCLND. This study examined the accuracy of LM/SL/SCLND in the MSLT, using the experience of the organizing center (John Wayne Cancer Institute [JWCI]) as a standard for comparison. METHODS: Before entering patients into the randomization phase, each center in the MSLT was required to finish a 30-case learning phase with complete nuclear medicine, pathology, and surgical review. Selection of MSLT patients in the LM/SL/SCLND treatment arm was based on complete pathologic and surgical data. The comparison group of JWCI patients was selected using these criteria: primary cutaneous melanoma having a thickness > or =1 mm with a Clark level > or =III, or a thickness <1 mm with a Clark level > or =IV (MSLT criterion); LM/SL performed between June 1, 1985, and December 30, 1998; and patient not entered in the MSLT. The accuracy of LM/SL/SCLND was determined by comparing the rates of sentinel node (SN) identification and the incidence of SN metastases in the MSLT and JWCI groups. RESULTS: There were 551 patients in the MSLT group and 584 patients in the JWCI group. In both groups, LM performed with blue dye plus a radiocolloid was more successful (99.1 %) than LM performed with blue dye alone (95.2%) (p = 0.014). After a center had completed the 30-case learning phase, the success of SN identification in the MSLT group was independent of the center's case volume or experience in the MSLT. CONCLUSIONS: Lymphatic mapping and sentinel lymphadenectomy can be successfully learned and applied in a standardized fashion with high accuracy by centers worldwide. Successful SN identification rates of 97% can be achieved, and the incidence of nodal metastases approaches that of the organizing center. A multidisciplinary approach (surgery, nuclear medicine, and pathology) and a learning phase of > or =30 consecutive cases per center are sufficient for mastery of LM/SL in cutaneous melanoma. Lymphatic mapping performed using blue dye plus radiocolloid is superior to LM using blue dye alone. PMID- 10522717 TI - Alcohol interventions in a trauma center as a means of reducing the risk of injury recurrence. AB - OBJECTIVE: Alcoholism is the leading risk factor for injury. The authors hypothesized that providing brief alcohol interventions as a routine component of trauma care would significantly reduce alcohol consumption and would decrease the rate of trauma recidivism. METHODS: This study was a randomized, prospective controlled trial in a level 1 trauma center. Patients were screened using a blood alcohol concentration, gamma glutamyl transpeptidase level, and short Michigan Alcoholism Screening Test (SMAST). Those with positive results were randomized to a brief intervention or control group. Reinjury was detected by a computerized search of emergency department and statewide hospital discharge records, and 6- and 12-month interviews were conducted to assess alcohol use. RESULTS: A total of 2524 patients were screened; 1153 screened positive (46%). Three hundred sixty six were randomized to the intervention group, and 396 to controls. At 12 months, the intervention group decreased alcohol consumption by 21.8+/-3.7 drinks per week; in the control group, the decrease was 6.7+/-5.8 (p = 0.03). The reduction was most apparent in patients with mild to moderate alcohol problems (SMAST score 3 to 8); they had 21.6+/-4.2 fewer drinks per week, compared to an increase of 2.3+/-8.3 drinks per week in controls (p < 0.01). There was a 47% reduction in injuries requiring either emergency department or trauma center admission (hazard ratio 0.53, 95% confidence interval 0.26 to 1.07, p = 0.07) and a 48% reduction in injuries requiring hospital admission (3 years follow-up). CONCLUSION: Alcohol interventions are associated with a reduction in alcohol intake and a reduced risk of trauma recidivism. Given the prevalence of alcohol problems in trauma centers, screening, intervention, and counseling for alcohol problems should be routine. PMID- 10522718 TI - Recent clinical experience with left heart bypass using a centrifugal pump for repair of traumatic aortic transection. AB - OBJECTIVE: To analyze the indications, results, and limitations of using left atrial to femoral artery (LA-FA) bypass to provide distal perfusion during repair of traumatic aortic injuries. SUMMARY BACKGROUND DATA: There is no consensus about the best method for repair of traumatic aortic transection. Distal aortic perfusion with LA-FA bypass and a centrifugal pump has been the authors' preferred technique for injuries to the aortic isthmus and descending thoracic aorta. METHODS: From 1988 to 1998, the authors operated on 30 patients with traumatic aortic transection using LA-FA bypass. The mean age of the group was 36+/-2 years. The mechanism of injury was from a motor vehicle accident in 97% of the cases. Distal aortic perfusion was maintained at 50 to 75 mm Hg with flow rates of 1.5 and 3 L/min. The mean aortic cross-clamp time was 38+/-2 minutes, and the mean bypass time was 49+/-2 minutes. RESULTS: No complications related to cannulation, arterial thromboembolism, renal failure, mesenteric ischemia, or hepatic insufficiency occurred. There were no cases of postoperative paraplegia and no deaths. CONCLUSION: Left atrial to femoral artery bypass is a safe, simple, and effective adjunct to the repair of traumatic injuries to the thoracic aorta. Active distal aortic perfusion preserves spinal cord, mesenteric, and renal blood flow and eliminates the potential catastrophic consequence of spinal cord ischemia from an unexpectedly prolonged aortic cross-clamp time. PMID- 10522719 TI - Decreased acute rejection in kidney transplant recipients is associated with decreased chronic rejection. AB - OBJECTIVE: To determine whether a recent decrease in the rate of acute rejection after kidney transplantation was associated with a decrease in the rate of chronic rejection. SUMMARY BACKGROUND DATA: Single-institution and multicenter retrospective analyses have identified acute rejection episodes as the major risk factor for chronic rejection after kidney transplantation. However, to date, no study has shown that a decrease in the rate of acute rejection leads to a decrease in the rate of chronic rejection. METHODS: The authors studied patient populations who underwent transplants at a single center during two eras (1984 1987 and 1991-1994) to determine the rate of biopsy-proven acute rejection, the rate of biopsy-proven chronic rejection, and the graft half-life. RESULTS: Recipients who underwent transplantation in era 2 had a decreased rate of biopsy proven acute rejection compared with era 1 (p < 0.05). This decrease was associated with a decreased rate of biopsy-proven chronic rejection for both cadaver (p = 0.0001) and living donor (p = 0.08) recipients. A trend was observed toward increased graft half-life in era 2 (p = NS). CONCLUSIONS: Development of immunosuppressive protocols that decrease the rate of acute rejection should lower the rate of chronic rejection and improve long-term graft survival. PMID- 10522720 TI - Sequential accumulation of K-ras mutations and p53 overexpression in the progression of pancreatic mucinous cystic neoplasms to malignancy. AB - OBJECTIVE: Pancreatic mucinous cystic neoplasms (MCNs) provide a spectrum of neoplastic changes ranging from benign to malignant. The authors have correlated K-ras mutations and p53 overexpression with the evolution of these tumors. METHODS: Areas of mild, moderate, or severe dysplasia were microdissected from paraffin-embedded tissue sections of 28 different MCNs (10 benign, 9 borderline, 9 malignant). Nonneoplastic pancreatic ducts were also microdissected from tissues adjacent to the tumors. Ten serous cystadenomas served as negative controls. K-ras codon 12 mutations were identified by a mutant-enriched nested polymerase chain reaction-restriction fragment length polymorphism assay and confirmed by sequencing. p53 overexpression was demonstrated by immunohistochemistry. RESULTS: K-ras mutations were detected in 20% of benign, 33% of borderline, and 89% of malignant MCNs. Histologically, mutations were found in 26% (7/27) of MCN epithelia with mild dysplasia, 38% (5/13) of MCN epithelia with moderate dysplasia, and 89% (8/9) of MCN epithelia with severe dysplasia or carcinoma. Ten percent (4/39) of nonneoplastic pancreatic ducts at the margins of MCN harbored mutations, all associated with borderline or malignant tumors. Overexpression of p53 occurred in none of the benign or borderline MCNs but in 44% (4/9) of the malignant tumors (p = 0.006 benign/borderline vs. malignant). p53 immunoreactivity was concentrated in areas of severe dysplasia/carcinoma or invasion, where K-ras mutation had been detected. CONCLUSION: These findings demonstrate a sequential accumulation of genetic changes in the carcinogenesis of MCN. K-ras mutations appear early and increase in proportion with increasing dysplasia. Overexpression of p53 is a late finding observed only in carcinomas, and in combination with mutated K-ras genes. The presence of K-ras mutations in nonneoplastic ducts supports formal pancreatic resection over enucleation for treatment. Mucinous cystic neoplasms may be a useful model to study the evolution of pancreatic ductal adenocarcinomas, in which precursor lesions remain unknown. PMID- 10522721 TI - Duodenum-preserving head resection in chronic pancreatitis changes the natural course of the disease: a single-center 26-year experience. AB - OBJECTIVE: To present preoperative and early postoperative data for 504 patients who underwent duodenum-preserving pancreatic head resection (DPPHR) for severe chronic pancreatitis (CP). BACKGROUND: The pancreatic head is considered to be the pacemaker of the disease in alcohol-induced CP. Indications for surgery in CP are intractable pain and local complications. DPPHR offers the advantage of treating the complications related to the inflammatory process in the head, relieving the pain syndrome, and preserving the bilioduodenal anatomy, and it may have the potential to change the natural course of chronic pancreatitis. METHODS: Between November 1972 and December 1998, 504 patients with chronic pancreatitis and an inflammatory mass in the pancreatic head were treated surgically after medical pain treatment for a median of 3.6 years. The procedure resulted in a hospital mortality rate of 0.8%. A continuous follow-up investigation lasting up to 26 years was conducted, during which the patients were reevaluated four times (1983, 1987, 1994, 1996). Between November 1982 and October 1996, 388 patients treated surgically were reinvestigated to evaluate the late outcome; the follow up rate was 94% (25 patients were lost to follow-up). The reinvestigation evaluation included glucose tolerance test, exocrine pancreatic function test, pain status, physical status, professional and social rehabilitation, and quality of life. RESULTS: After an observation period of up to 14 years, 78.8% of the patients were completely pain-free and 12.5% had (yearly) pain. 91.3% were considered as pain-free; 8.7% had continuing abdominal pain; 12% had abdominal complaints. During the 14 years of follow-up, only 9% were admitted to the hospital for acute episodes of chronic pancreatitis. Endocrine function was improved in 11%; in 21%, diabetes developed de novo. The rate of hospital admission for acute episodes decreased from 69% before surgery to 9% after surgery. In the clinical management period of 9 years (median), the frequency of hospital admission dropped from 5.4 per patient before surgery to 2.7 after surgery. Fourteen years after surgery, 69% of the patients were professionally rehabilitated; in 72%, the quality of life index (Karnofsky criteria) was 90 to 100 and in 18%, it was <80. CONCLUSION: In patients with alcoholic chronic pancreatitis in whom an inflammatory mass has developed in the pancreatic head, DPPHR results in a change in the natural course of the disease in terms of pain status, frequency of acute episodes, need for further hospital admission, late death, and quality of life. PMID- 10522722 TI - Outcome of renal artery reconstruction: analysis of 687 procedures. AB - OBJECTIVE: To evaluate the short- and long-term results of surgical reconstruction of the renal arteries, the authors review their experience with more than 600 reconstructions performed over a 12-year period. SUMMARY BACKGROUND DATA: Reconstruction of the renal arteries, whether for primary renal indications or concomitantly with aortic reconstruction, has evolved over the past 40 years. There is concern that renal artery reconstructions carry significant rates of mortality and morbidity and may fare poorly compared with less-invasive procedures. METHODS: From 1986 to 1998, 687 renal artery reconstructions were performed in 568 patients. Of these, 105 patients had simultaneous bilateral renal artery reconstructions. Fifty-six percent of the patients were male; 11% had diabetes; 35% admitted to smoking at the time of surgery. Mean age was 67 (range, 1 to 92). One hundred fifty-six (23%) were primary procedures and the remainder were adjunctive procedures with aortic reconstructions; 406 were abdominal aortic aneurysms and 125 were aortoiliac occlusive disease. Five hundred procedures were bypasses, 108 were endarterectomies, 72 were reimplantation, and 7 were patch angioplasties. There were 31 surgical deaths (elective and emergent) in the entire group for a mortality rate of 5.5%. Predictors of increased risk of death were patients with aortoiliac occlusive disease and patients undergoing bilateral simultaneous renal artery revascularization. Cause of death was primarily cardiac. Other nonfatal complications included bleeding (nine patients) and wound infection (three patients). There were 9 immediate occlusions (1.3%) and 10 late occlusions (1.5%). Thirty-three patients (4.8%) had temporary worsening of their renal function after surgery. CONCLUSION: Renal artery revascularization is a safe and durable procedure. It can be performed in selected patients for primary renovascular pathology. It can also be an adjunct to aortic reconstruction with acceptable mortality and morbidity rates. PMID- 10522723 TI - Work loads and practice patterns of general surgeons in the United States, 1995 1997: a report from the American Board of Surgery. AB - OBJECTIVE: To characterize the work loads and practice patterns of general surgeons in the United States over a 3-year period (1995 to 1997). METHODS: The surgical operative logs of 2434 "generalist" general surgeons recertifying in surgery form the basis of this report. Selected demographics of the group are as follows: location: 50% Northeast and Southeast, 21 % Midwest, 29% West and Southwest; practice type: 45% solo, 40% group, 9% academics; size of practice community: 46% highly urban, 19% rural. Parameters evaluated were the average number of procedures and their distribution by category related to geographic area, practice type, community size, and other parameters. Statistical analysis was accomplished using analysis of variance. RESULTS: No significant year-to-year differences were observed between cohorts. The average numbers of procedures per surgeon per year was 398, distributed as follows: abdomen 102, alimentary tract 63, breast 54, endoscopic 51, vascular 39, trauma 6, endocrine 4, and head and neck, 3. Eleven percent of the 398 procedures were performed laparoscopically. Major index cases were largely concentrated with small groups of surgeons representing 5% to 10% of the total. Significant differences were as follows: surgeons in the Northeast and West performed far fewer procedures than those elsewhere. Urban surgeons performed a few more tertiary-type procedures than did rural ones; however, rural surgeons performed many more total procedures, especially in endoscopy, laparoscopy, gynecology, genitourinary, and orthopedics. Academic surgeons performed substantially fewer total procedures as a group than did nonacademic ones and in all categories except liver, transplant, and pancreas. Male surgeons performed more procedures than did female surgeons, except those involving the breast. More procedures were done by surgeons in group practice than by those in solo practice. U.S. medical graduates and international medical graduates had similar work loads but with a different distribution. CONCLUSIONS: This unique database will be useful in tracking trends over time. More importantly, it demonstrates that general surgery practice in the United States is extremely heterogeneous, a fact that must be acknowledged in any future workforce deliberations. PMID- 10522724 TI - Safety and efficacy of low anterior resection for rectal cancer: 681 consecutive cases from a specialty service. AB - OBJECTIVE: To determine perioperative morbidity, survival, and local failure rates in a large group of consecutive patients with rectal cancer undergoing low anterior resection by multiple surgeons on a specialty service. The primary objective was to assess the surgical complications associated with preoperative radiation sequencing. SUMMARY BACKGROUND DATA: The goals in the treatment of rectal cancer are cure, local control, and preservation of sphincter, sexual, and bladder function. Surgical resection using sharp perimesorectal dissection is important for achieving these goals. The complications and mortality rate of this surgical strategy, particularly in the setting of preoperative chemoradiation, have not been well defined. METHODS: There were 1233 patients with primary rectal cancer treated at the authors' cancer center from 1987 to 1995. Of these, 681 underwent low anterior resection and/or coloanal anastomosis for primary rectal cancer. The surgical technique used the principles of sharp perimesorectal excision. Morbidity and mortality rates were compared between patients receiving preoperative chemoradiation (Preop RT, n = 150) and those not receiving preoperative chemoradiation (No Preop RT, n = 531). Recurrence and survival data were determined in patients undergoing curative resection (n = 583, 86%) among three groups of patients: those receiving Preop RT (n = 131), those receiving postoperative chemoradiation (Postop RT, n = 110), and those receiving no radiation therapy (No RT, n = 342). RESULTS: The perioperative mortality rate was 0.6% (4/681). Postoperative complications occurred in 22% (153/681). The operative time, estimated blood loss, and rate of pelvic abscess formation without associated leak were higher in the Preop RT group than the No Preop RT group. However, the overall complication rate, rate of wound infection, anastomotic leak, and length of hospital stay were no different between Preop RT and No Preop RT patients. With a median follow-up of 45.6 months, the overall actuarial 5-year recurrence rate for patients undergoing curative resection (n = 583) was 19%, with 4% having local recurrence only, 12% having distant recurrence, and 3% having both local and distant recurrence, for an overall local recurrence rate of 7%. The actuarial 5-year overall survival rate was 81%; the disease-free survival rate was 75% and the local recurrence rate was 10%. The overall survival rate was similar between Preop RT (85%), Postop RT (72%), and No RT (83%) patients (p = 0.10), whereas the disease-free survival rate was significantly worse for Postop RT (65%) patients compared with Preop RT (79%) and No RT (77%) patients (p = 0.04). CONCLUSION: The use of preoperative chemoradiation results in increased operative time, blood loss, and pelvic abscess formation but does not increase the rate of anastomotic leaks or the length of hospital stay after low anterior resection for rectal cancer. The 5 year actuarial overall survival rate for patients undergoing curative resection exceeded 80%, with a local recurrence rate of 10%. PMID- 10522725 TI - Results of transcervical thymectomy for myasthenia gravis in 100 consecutive patients. AB - OBJECTIVE: To review the results of the authors' most recent 100 consecutive cases of transcervical thymectomy for myasthenia gravis (MG) in terms of complications and outcome in comparison with other reported techniques. SUMMARY BACKGROUND DATA: Myasthenia gravis is believed to be an autoimmune disorder characterized by increasing fatigue with exertion. The role of thymectomy in the management of the disease remains unproven, but there is widespread acceptance of the notion that complete thymectomy improves the course of the disease. Complete excision of the thymus is the goal in all cases; however, the best technique to achieve complete thymectomy remains controversial. The authors favor a transcervical approach through a small collar incision aided by a specially designed sternal retractor. Others prefer a transsternal, a combined transcervical and transsternal ("maximal"), or a video-assisted thoracoscopic surgical approach. METHODS: A retrospective review of the authors' most recent 100 consecutive transcervical thymectomies for nonthymoma-associated MG was performed using medical records and telephone interviews. Patients' symptoms were graded before surgery and at the most recent (within the last 6 months) postoperative time point, using the modified Osserman classification: 0 = asymptomatic, 1 = ocular signs and symptoms, 2 = mild generalized weakness, 3 = moderate generalized weakness, bulbar dysfunction, or both, and 4 = severe generalized weakness, respiratory dysfunction, or both. RESULTS: There were 61 female patients and 39 male patients with a mean age of 38 years (range, 14 to 84). The median hospital stay was 1 day. There were no deaths and no significant complications. Seventy-eight patients who had undergone surgery >12 months ago were available for analysis. In these patients, with a mean follow-up time of 5 years (median 5.3; range, 12 months to 10 years), the median preoperative Osserman grade improved from 3.0 (mean 2.73) before surgery to 1.0 after surgery (mean 0.94). CONCLUSIONS: The transcervical approach for thymectomy for the treatment of MG produces results similar to those of other surgical approaches, with the added benefits of shortened hospital stay, decreased complications, reduced cost, and broader physician and patient acceptance of surgical treatment. PMID- 10522726 TI - Clinical and pathologic predictors of survival in patients with thymoma. AB - OBJECTIVE: To evaluate the Johns Hopkins Hospital experience with 136 thymomas over the past 40 years. This number of patients allowed quantitative estimation of the independent influence of common clinicopathologic risk factors using multivariate analysis. SUMMARY BACKGROUND DATA: Thymomas vary widely in terms of recurrence and influence on overall survival. Several series have indicated the importance of initial tumor invasion, as well as the extent of surgical resection, as predictors of recurrence and survival after thymoma resection. However, findings have been equivocal when other predictors of prognosis were examined. METHODS: The authors evaluated 136 patients seen at the Johns Hopkins Hospital between 1957 and 1997 with a pathologic diagnosis of thymoma. Demographic information, clinical staging data, surgical and adjuvant treatment details, and patient follow-up data were obtained from the patient record and from detailed patient or family interviews. Microscopic sections of all 136 patients were reviewed by two pathologists blinded to the clinical data. All data were analyzed by multivariate Cox regression analysis, which allowed the quantification of the independent predictive value of 12 putative clinicopathologic prognostic indicators. RESULTS: Completeness of follow-up was 99%, 99%, and 98% of eligible patients at 5, 10, and 15 years, respectively. Forty percent of the patients had associated myasthenia gravis and 27% had a secondary primary malignancy. Overall patient survival rates were 71%, 56%, 44%, 38%, and 33% at 5, 10, 15, 20, and 25 years, respectively. Overall, the thymoma related mortality rate was 14%; the nonthymoma-related mortality rate was 26%. Incomplete resection, preoperative absence of myasthenia gravis, and advanced Lattes/Bernatz pathologic class were found to be independent predictors of poorer overall survival. CONCLUSIONS: These findings support a policy of aggressive, complete surgical resection of all thymomas when feasible. Thymoma behaves as a rather indolent tumor, with most deaths from causes unrelated to thymoma or its direct treatment. Clinicians should have an increased awareness of the possibility of second primary malignancies in patients with thymoma. PMID- 10522727 TI - Long-term functional outcome and quality of life after stapled restorative proctocolectomy. AB - OBJECTIVE: To evaluate prospectively long-term quality of life and functional outcome after restorative proctocolectomy (RPC) with ileal pouch-anal anastomosis, and to evaluate and validate a novel quality-of-life indicator in this group of patients. SUMMARY BACKGROUND DATA: Restorative proctocolectomy with ileal pouch-anal anastomosis is now the preferred option when total proctocolectomy is required for ulcerative colitis or familial adenomatous polyposis, but long-term data on functional outcome and quality of life after the procedure are lacking. METHODS: Patients (n = 977) who underwent RPC with stapled anastomosis for colitis or polyposis coli and who were followed for > or =12 months were included. Quality of life, fecal incontinence, and satisfaction with surgery were prospectively evaluated by structured interview or questionnaire for 1 to 12 years after surgery (median 5.0). Quality of life was scored using the Cleveland Global Quality of Life (CGQL) instrument (Fazio Score). This is a novel score developed over the past 15 years by the senior author. Quality of life was also evaluated in a subgroup of patients with the Short Form 36 (SF-36). The CGQL was validated by determining its reliability, responsiveness, and validity as well as its correlation with the SF-36 score. RESULTS: Postoperative quality of life as measured by SF-36 was excellent and compared well with published norms for the general U.S. population. The CGQL was found to be reliable, responsive, and valid, and there was a high correlation with the SF-36 scores. Using the CGQL, quality of life was shown to increase after the first 2 years after surgery, and there was no deterioration thereafter. The prevalence of perfect continence increased from 75.5% before surgery to 82.4% after surgery, and although this deteriorated somewhat >2 years after surgery, it was no worse than preoperative values. Ninety-eight percent of patients would recommend the surgery to others. CONCLUSIONS: Long-term quality of life after ileal pouch surgery is excellent and the level of continence is satisfactory. This surgery is an excellent long-term option in patients requiring total proctocolectomy. The CGQL is a simple, valid, and reliable measure of quality of life after pelvic pouch surgery and may well be applicable in many other clinical conditions. PMID- 10522728 TI - Minimally invasive surgery for achalasia: an 8-year experience with 168 patients. AB - BACKGROUND: Seven years ago, the authors reported on the feasibility and short term results of minimally invasive surgical methods to treat esophageal achalasia. In this report, they describe the evolution of the surgical technique and the clinical results in a large group of patients with long follow-up. PATIENTS AND METHODS: Between January 1991 and October 1998, 168 patients (96 men, 72 women; mean age 45 years, median duration of symptoms 48 months), who fulfilled the clinical, radiographic, endoscopic, and manometric criteria for a diagnosis of achalasia, underwent esophagomyotomy by minimally invasive techniques. Forty-eight patients had marked esophageal dilatation (diameter >6.0 cm). Thirty-five patients had a left thoracoscopic myotomy, and 133 patients had a laparoscopic myotomy plus a partial fundoplication. Follow-up to October 1998 was complete in 145 patients (86%). RESULTS: Median hospital stay was 72 hours for the thoracoscopic group and 48 hours for the laparoscopic group. Eight patients required a second operation for recurrent or persistent dysphagia, and two patients required an esophagectomy. There were no deaths. Good or excellent relief of dysphagia was obtained in 90% of patients (85% after thoracoscopic and 93% after laparoscopic myotomy). Gastroesophageal reflux developed in 60% of tested patients after thoracoscopic myotomy and in 17% after laparoscopic myotomy plus fundoplication. Laparoscopic myotomy plus fundoplication corrected reflux present before surgery in five of seven patients. Patients with a dilated esophagus had excellent relief of dysphagia after laparoscopic myotomy; none required an esophagectomy. CONCLUSIONS: Minimally invasive techniques provided effective and long-lasting relief of dysphagia in patients with achalasia. The authors prefer the laparoscopic approach for three reasons: it more effectively relieved dysphagia, it was associated with a shorter hospital stay, and it was associated with less postoperative reflux. Laparoscopic Heller myotomy and partial fundoplication should be considered the primary treatment for esophageal achalasia. PMID- 10522729 TI - Laparoscopic fundoplication failures: patterns of failure and response to fundoplication revision. AB - OBJECTIVE: To determine rates and mechanisms of failure in 857 consecutive patients undergoing laparoscopic fundoplication for gastroesophageal reflux disease or paraesophageal hernia (1991-1998), and compare this population with 100 consecutive patients undergoing fundoplication revision (laparoscopic and open) at the authors' institution during the same period. SUMMARY BACKGROUND DATA: Gastroesophageal fundoplication performed through a laparotomy or thoracotomy has a failure rate of 9% to 30% and requires revision in most of the patients who have recurrent or new foregut symptoms. The frequency and patterns of failure of laparoscopic fundoplication have not been well studied. METHODS: All patients undergoing fundoplication revision were included in this study. Symptom severity was scored before and after surgery by patients on a 4-point scale. Evaluation of patients included esophagogastroscopy, barium swallow, esophageal motility, 24-hour ambulatory pH, and gastric emptying studies. Statistical analysis was performed with multiple chi-square analyses, Fisher exact test, and analysis of variance. RESULTS: Laparoscopic fundoplication was performed in 758 patients for gastroesophageal reflux disease and in 99 for paraesophageal hernia. Median follow-up was 2.5 years. Thirty-one patients (3.5%) have undergone revision for fundoplication failure. The mechanism of failure was transdiaphragmatic herniation of the fundoplication in 26 patients (84%). In 40 patients referred from other institutions, after laparoscopic fundoplication, only 10 (25%) had transdiaphragmatic migration (p < 0.01); a slipped or misplaced fundoplication occurred in 13 patients (32%), and a twisted fundoplication in 12 patients (30%). The failure mechanisms of open fundoplication (29 patients) followed patterns previously described. Fundoplication revision procedures were initiated laparoscopically in 65 patients, with six conversions (8%). The morbidity rate was 4% in laparoscopic procedures and 9% in open ones. There was one death, from aspiration and adult respiratory distress syndrome after open fundoplication. A year or more after revision operation, heartburn, chest pain, and dysphagia were rare or absent in 88%, 78%, and 91%, respectively, after laparoscopic revision, and were rare or absent in 91%, 83%, and 70%, respectively, after open revision, but 11 patients ultimately required additional operations for continued or recurrent symptoms, 3 after open revision (17%), and 8 after laparoscopic fundoplication (11%). CONCLUSIONS: Laparoscopic fundoplication failure is infrequent in experienced hands; the rate may be further reduced by extensive esophageal mobilization, secure diaphragmatic closure, esophageal lengthening (applied selectively), and avoidance of events leading to increased intraabdominal pressure. When revision is required, laparoscopic access may be used successfully by the laparoscopically experienced esophageal surgeon. PMID- 10522730 TI - Overall survival or other clinical benefits from adjuvant selective intraarterial chemotherapy in patients undergoing curative liver resection for metastatic colorectal tumor. PMID- 10522731 TI - Results on debridement and closed packing with stuffed Penrose drains for necrotizing pancreatitis. PMID- 10522732 TI - Second International Copenhagen Pain Symposium: aspects of chronic pain. PMID- 10522733 TI - Pain--an overview. AB - The neuromatrix theory of pain proposes that pain is a multidimensional experience produced by characteristic "neurosignature" patterns of nerve impulses generated by a widely distributed neural network--the "body-self neuromatrix"--in the brain. These neurosignature patterns may be triggered by sensory inputs, but they may also be generated independently of them. Pains that are evoked by noxious sensory inputs have been meticulously investigated by neuroscientists, and their sensory transmission mechanisms are generally well understood. In contrast, chronic pain syndromes, which are often characterized by severe pain associated with little or no discernible injury or pathology, remain a mystery. The neuromatrix theory of pain, however, provides a new conceptual framework that is consistent with recent clinical evidence. It proposes that the output patterns of the neuromatrix activate perceptual, homeostatic and behavioral programs after injury or pathology, or as a result of multiple other inputs that act on the neuromatrix. Pain, then, is produced by the output of a widely distributed neural network in the brain rather than directly by sensory input evoked by injury, inflammation or other pathology. The neuromatrix, which is genetically determined and modified by sensory experience, is the primary mechanism that generates the neural pattern that produces pain. Its output pattern is determined by multiple influences, of which the somatic sensory input is only a part, that converge on the neuromatrix. PMID- 10522734 TI - The role of psychological factors in chronic pain. AB - The traditional view conceptualizes pain as being directly associated with the extent of physical pathology. The observations that there are a number of patients who report pain in the absence of physical pathology, the converse, asymptomatic individuals who evidence objective physical pathology, the inconsistency in response of patients with identical diagnoses, and the low association between impairments and disability suggest that factors other than physical pathology contribute to the reports of pain. The role of behavioral, cognitive, and affective factors have each been shown to have direct effects on the report of pain, adaptation, and response to treatment, as well as indirect effects by influencing sympathetic nervous system and neurochemical factors associated with nociception. The direct and indirect effects of behavioral (operant), cognitive, and affective factors in chronic pain are described. PMID- 10522735 TI - Psychiatric approaches to non-cancer pain. AB - Most pain clinic therapists would agree as to the importance of including a psychiatrist and/or a psychologist on the team. Because of the psychiatrist's training in medicine he/she may have a useful role as a bridge between the somatically orientated clinicians and those dealing with psychosocial issues such as psychologists and social workers. This paper highlights the psychiatric and psychological approaches important for treatment of chronic non-cancer pain conditions. PMID- 10522736 TI - Health-related quality of life measurements in the assessment of pain clinic results. AB - Following the World Health Organization (WHO) classification of impairment, disability and subjective quality of life, chronic pain disorder has been examined. In this overview impairment has been restricted to non-malignant pain. Disability has been subdivided into disorder-oriented and ailment-oriented symptoms. The disorder-oriented symptoms are pain and depression, the latter emphasizing that pain is an unpleasant sensation. Both pain and depression are essentially subjectively reported symptoms which are unidimensionel, implying that they often are measured by use of visual analog scales. However, the Major Depression Inventory (MDI) and the Symptom Checklist 90 (SCL-90) are appropriate questionnaires itemizing the respective dimensions. The ailment-oriented symptoms have negative social well-being as their core-symptoms, including reduced role functioning. The Short-Form (SF-36) is an appropriate scale in this respect. Subjective quality of life is essentially measuring positive well-being and the most appropriate measurement instrument is the Psychological Well-Being Schedule (PGWB) or its subscales, among which is the WHO (Five) well-being index. PMID- 10522737 TI - Pain measurement. AB - Increasing evidence from laboratory methods in humans and animals indicates that pain arises from, and is modulated by, a number of mechanisms. In addition, these mechanisms are not static but change as pain persists. Recent human studies have demonstrated new aspects of pain processing at all levels of the central nervous system. Studies of the influence of analgesic agents on a large number of experimental pain measures have shown a preferential effect of opioids for attenuating the central integration of prolonged stimuli while local anesthetics may be more effective for brief stimulation. Studies of NK1 antagonists in man have shown results similar to those found with animals. There is little effect on brief stimulation of A delta and C-fiber nociceptors, including conditions that can evoke central summation. However, these antagonists, which block the effects of substance P, are effective in more persistent states such as postsurgical pain. Persistent pain can also alter the function of the large diameter A beta touch afferents, ranging from increased tactile sensitivity in inflammatory conditions to frank allodynia following nerve injury or focal nociceptor stimulation. Recent advances in evaluation of supraspinal pain processing in humans have demonstrated pain-related activation using both methods that assess synchronized neural activity and methods that infer this activity in the whole brain by local changes in regional cerebral blood flow. These methods have begun to identify brain regions associated with the multiple dimensions and processing of painful stimulation and the modulation of these processes by pharmacological agents and non-pharmacological interventions. PMID- 10522738 TI - Opioid therapy of chronic pain: assessment of consequences. AB - This paper will review what is known about key issues of importance in the clinical use of opioids for the treatment of intractable non-cancer related pain, and will attempt to describe the evolving areas of consensus among clinicians who treat pain and addiction regarding various aspects of use of opioids for the treatment of chronic non-cancer pain. PMID- 10522739 TI - Opioid rotation in chronic non-malignant pain patients. A retrospective study. AB - BACKGROUND: The clinical advantage of opioid rotation is probably due to incomplete cross-tolerance favouring analgesia more than adverse effects. The objectives of opioid rotation in chronic non-malignant patients are 1): rotation between different long-acting opioids (LAO) to improve analgesia and reduce side effects, and 2): rotation from short-acting opioids (SAO) to LAO to establish stable analgesia in order to minimise withdrawal symptoms, risk of tolerance and addiction. METHODS: A retrospective analysis of 37 rotations between different LAO and 59 rotations from SAO to LAO. RESULTS: The main reason for opioid rotation was insufficient pain relief. Opioid rotations resulted in significantly better pain control in 59% (CI95=49-76%) of the patients rotated between different LAO and 73% (CI95=60-84%) of the patients rotated from SAO to LAO. During rotations symptoms of withdrawal and overdosing were relatively frequent in both groups. No significant dose changes were seen when rotating between different LAO. However, the consequence of rotation from SAO to LAO was a 74% increase in the opioid dose (CI95=30-117%). CONCLUSION: Opioid rotations between different LAO resulted in better pain control and fewer side-effects at dose levels predicted to be equianalgesic. The majority of the patients rotated from SAO to LAO obtained improved analgesia, but the cost was a 74% increase in the opioid dose. PMID- 10522740 TI - Opioid use in cancer pain. Is a more liberal approach enhancing toxicity? AB - The majority of cancer patients develop pain before death. This pain has been shown to be underdiagnosed and undertreated. Opioid use has increased in the past 20 years in both developing and developed countries. The changing pattern in opioid use has resulted in the emergence of neurotoxicity as a major side effect of the treatment of cancer pain. The syndrome of opioid-induced neurotoxicity (OIN) encompasses delirium, hallucinosis, myoclonus/seizures and hyperalgesia. Increased vigilance can lead to the timely diagnosis of OIN, and strategies for its treatment can be implemented with encouraging results. Identification and modification of risk factors for the development of OIN can help in its prevention and improve the quality of life in advanced cancer patients. PMID- 10522741 TI - Fatigue. Measures and relation to pain. AB - Fatigue describes reduced capacity to sustain force or power output, reduced capacity to perform multiple tasks over time and simply a subjective experience of feeling exhausted, tired, weak or having lack of energy. Pain and fatigue have several components in common, such as being subjective, prevalent in most patients with cancer and caused by multiple factors of both a physical and psychological nature. In order to explore the relationship between fatigue and pain, data from five studies were used: two random samples from the Norwegian population (n=2323 and n=1965), Hodgkin's disease survivors (n=459), palliative care patients (n=434) and patients with bone metastases (n=94). All patients had completed one or more of the following instruments: EORTC QLQ-C30, SF-36 and/or Fatigue Questionnaire. The level of fatigue was much higher in the two palliative care populations (54.4 and 63.2) as compared to the normal population samples (25.0). Patients with bone metastases had significantly more pain (72.0) than the patients in the palliative care trial (47.4) and norms (20.5). In the two palliative care and bone metastases populations fatigue was almost unchanged over time, while pain was reduced. In the palliative care population a high level of fatigue (80.3) and pain (57.8) was reported 0-1 month before death. The relationship between pain, fatigue and the health-related quality of life domains should be explored in more detail, especially in follow-up studies in order to assess possible changes over time. In addition, the validity of the existing instruments measuring fatigue should be investigated for use in patients with advanced disease and short life expectancy. PMID- 10522742 TI - Methodological issues in the assessment of health-related quality of life in palliative care trials. AB - Palliative care aims at improving the patient's quality of life. Clinical trials, therefore, often include the patient's subjective evaluation of symptoms and psychosocial problems, so-called health-related quality of life (HRQL), as end points. Unfortunately, there are frequently methodological weaknesses in the assessment of HRQL. This paper discusses four criteria which can be used in the evaluation of the quality of such studies: I. The authors should document that they have included the relevant HRQL issues in the questionnaire. If a trial misses important issues, its results may be misleading. II. The sample size should be sufficient to detect meaningful differences. Many trials are too small. III. The assessment of HRQL should have the appropriate timing, reflecting the research questions. The symptoms and the benefit resulting from treatments are not constant over time and often have cyclic patterns. The results may therefore be dependent on the timing of the administration of questionnaires to patients and on the time frames specified in the instructions. IV. The data must be reasonably complete. Incomplete data cannot be avoided in palliative care research, but missing data due to administrative failures or unrealistic schedules must be avoided. A pilot study may show whether a study is feasible. Missing data are likely to bias results. Many published palliative care studies are suboptimal with regard to one or more of these four criteria. This should be considered when reading published studies and when new trials are planned. PMID- 10522743 TI - Ethical issues in palliative care. AB - The relief of suffering is one of the aims of health care. Pain relief is a moral obligation in health care, not an optional extra. Doctors have moral obligations to strive to relieve pain, to be competent in basic pain control, and to endeavour to give patients an adequate understanding of their illness and painkillers. The most common moral problem in pain control in terminally ill patients is the conflict between the obligation to relieve suffering and the obligation to prolong life. The law prohibits intentionally causing the death of another person. Debates follow as to what constitutes an intention to cause death, and what actually constitutes a cause of death. At present, doctors are legally permitted to give sedatives and analgesics to terminally ill patients with the intention of relieving suffering, even if life is shortened. The moral principle of the 'double effect' relates to this and is explained. It relies on a distinction between intended and foreseen effects of treatment. Some people dispute the distinction between intended and foreseen effects and claim that the principle of double effect allows doctors who intend euthanasia to carry it out under cover of the law. This debate is explored in the article. Finally, is it ever morally justifiable to end the patient's life on the grounds that this is the only way to end pain? Even if it is, should euthanasia be legalised? A brief comment on these issues, and the roles of law and morality, are made. PMID- 10522744 TI - Economic implications of pain management. AB - BACKGROUND: Economic issues in pain management affect the patient, the provider and society. This paper will review some of the data on the costs to society of chronic pain and its associated disability. It will also look at the costs to patient and provider of alternative economic models. Conceptual issues that underlie health care delivery and the attendant costs must be addressed if society is to gain control over runaway health care costs and reduce the economic burden of chronic pain and disability for the patient as well as the provider. METHODS: Literature review and synthesis. RESULTS: Chronic pain is the primary cause of health care consumption and disability in the working years. Multidisciplinary pain clinics have proven utility. Data on efficacy of most other kinds of care is lacking. Disability costs are related to conceptual inadequacies and the medicalization of post-industrial societies. CONCLUSION: To control inappropriate care and escalating costs, we must change concepts of pain and disability and the methods of funding both of these in relation to chronic pain. The outcome of the continuing struggle between the profession of medicine, the state and capitalists will determine how and whether pain management is a part of medical care. PMID- 10522745 TI - A non-thoracic operation for a patient with single lung transplantation. AB - The number of lung transplants performed annually is increasing. It is becoming more likely that these patients will present post transplantation to anesthesiologists who have little experience in dealing with the physiological, pharmacological, medical and surgical problems that these patients present. This article discusses the management of a patient presenting for surgery after lung transplantation. PMID- 10522746 TI - Relief of postural post dural puncture headache by an epidural blood patch 12 months after dural puncture. AB - A 20-year-old previously healthy male presented at the pain clinic with chronic headache of about one year duration. Clinical examination revealed no pathological manifestations. During the consultation the patient was drinking Coca-Cola. On direct questioning he told that drinking Coca-Cola gave partial relief from the headache, and that the headache started after he had received two spinal anaesthetics for treatment of a lower leg fracture. Postural post dural puncture headache was now suspected and an epidural blood patch performed. Despite an interval of nearly 12 months since the dural punctures, a single epidural blood patch completely relieved the headache. This case history demonstrates that an epidural blood patch should be tried if a chronic post dural puncture headache is suspected. PMID- 10522747 TI - Bevel orientation and postdural puncture headache. A new possible explanation? PMID- 10522748 TI - Glomerular effects of cholera toxin in isolated perfused rat kidney: a potential role for platelet activating factor. AB - Cholera toxin (MW 84 kDa) is now considered a pharmacological tool to study the adenylyl cyclase system and a stimulus to generate platelet activating factor in the intestinal tract. We used this toxin to evaluate the renal haemodynamics, glomerular filtration function, tubular sodium transport and toxicity in isolated perfused rat kidney. Kidneys from adult male Wistar rats were isolated for perfusion. The perfusion fluid was modified Krebs-Henseleit solution and the samples were analyzed for sodium, potassium, inulin and osmolality. Clearance techniques were used to calculate physiological parameters. Cholera toxin (1.0 microg/ml) caused a significant time-dependent reduction of glomerular filtration rate and urinary flow. This toxin also caused a small, but consistent reduction in fractional proximal sodium reabsortion (toxin = 67.43+/-2.42% versus control = 79.26+/-5.80%; P<0.025). WEB 2086, a platelet activating factor receptor antagonist at 100 microg/ml completely blocked the effects induced by cholera toxin on glomerular filtration rate, fractional proximal sodium reabsortion and urinary flow. In contrast to cholera toxin, dibutyryl-cyclic AMP (10(-5) M) significantly increased glomerular filtration rate (Db-cyclic AMP = 0.651+/-0.035 versus control = 0.514+/-0.043 ml x g(-1) x min(-1); P<0.025) in isolated perfused kidneys. Db-cyclic AMP caused a similar, but more severe reduction in fractional proximal sodium reabsortion (Db-cyclic AMP = 54.21+/-2.35% versus control = 70.10+/-3.24%; P<0.025). In addition Db-cyclic AMP increased significantly the urinary flow (Db-Cyclic AMP = 0.290+/-0.018 versus control = 0.179+/-0.026 ml x g(-1) x min.(-1); P<0.025). WEB 2086+ Db-cyclic AMP also caused a significant increase in the urinary flow with maximal effect at 90 min. (WEB+Db-cyclic AMP = 0.26+/-0.01 versus control = 0.15+/-0.01 ml x g(-1) x min.( 1); n = 8, P<0.025). Cholera toxin caused a decrease of urinary flow (toxin = 0.034+/-0.004 versus control = 0.145+/-0.02 ml x g(-1) x min.(-1); P<0.025), this effect was also completely abolished by WEB 2086 when it was injected previously to toxin. When only WEB 2086 was injected, the functional parameters remained stable throughout the perfusion time. Cholera toxin had no effect on renal vascular resistance, renal perfusate flow or tissue potassium, suggesting renal integrity in kidneys treated with this toxin. The results suggest that cholera toxin effects in the perfused rat kidney are primarily mediated by platelet activating factor. PMID- 10522749 TI - Effects of propofol on contractile response and electrophysiological properties in single guinea-pig ventricular myocyte. AB - Effects of propofol on contractile response, action potential, resting membrane potential and L-type voltage-dependent calcium channel current were examined in guinea-pig single cardiac myocyte. Propofol (10(-4) M) inhibited contractile response induced by electrical stimulation (83.6% of control, n = 5), but did not change the resting membrane potential. On the other hand, propofol reduced the overshoot of action potential (10(-4) M), and shortened the duration of action potential (10(-5) and 10(-4) M). Whole-cell voltage clamp experiment showed inhibition of L-type calcium channel current (ICa, 10(-5) M: 90.8+/-1.39, 10(-4) M: 83.4+/-1.53% of control, n = 5). In addition, propofol showed use-dependent block of ICa. It is concluded that negative inotropic effect of propofol is caused by suppression of action potential, and that inhibition of ICa plays a role in shortening of the duration of action potential. PMID- 10522750 TI - Threshold concentrations of endothelin-1: the effects on contractions induced by 5-hydroxytryptamine in isolated rat cerebral and mesenteric arteries. AB - This study compares the effects of threshold concentrations of endothelin-1 in isolated rat basilar arteries with those in mesenteric arterial branches and investigates the mechanisms of inhibitory and potentiating endothelin-1-effects. In basilar arteries, endothelin-1 reduces the contractions induced by 5 hydroxytryptamine (5-HT), by the thromboxane A2 agonist U46619, and by vasopressin. The inhibitory effect of endothelin-1 on the contraction induced by 5-HT is abolished by deendothelialization, by the endothelin ET(B) receptor antagonist RES 701-1, by indomethacin, or by glibenclamide. In mesenteric arteries, endothelin-1 potentiates the contractile effects of 5-HT, U46619, and vasopressin. The potentiation of the contractile effect induced by 5-HT is only somewhat modified by deendothelialization, but abolished by the thromboxane A2 receptor antagonists GR32191 and ridogrel. U46619 potentiates the 5-HT-effect in mesenteric arteries. Thus, though the contractile endothelin ET(A) receptors were not blocked, threshold concentrations of endothelin-1 inhibited contractile effects in the rat basilar artery via activation of endothelial ET(B) receptors. Prostaglandins and ATP-sensitive K+ channels are involved in this inhibitory action. In contrast, endothelin-1 potentiates contractile actions in mesenteric arteries via the release of endogeneous thromboxane A2 from non-endothelial cells. The study points out the completely different role of the endothelium in combined effects of endothelin-1 between cerebral and mesenteric arteries. PMID- 10522751 TI - Extensive biliary excretion of the sulfasalazine analogue, susalimod, but different concentrations in the bile duct in various animal species correlating to species-specific hepatobiliary toxicity. AB - Studies on biliary concentrations of susalimod were conducted in rat, dog and monkey to clarify the interspecies differences observed in toxicology studies with respect to hepatobiliary toxicity after long-term administration of the compound. Dose-related bile duct hyperplasia appeared only in dogs at doses > or =75 mg/kg/day, while in rats and monkeys it did not appear at doses up to 1500 and 2000 mg/kg/day respectively. Biliary excretion was investigated after intraduodenal administration of susalimod in anaesthetised animals. In addition excretion routes were determined by collecting urine and faeces following a radiolabelled intravenous dose. Susalimod was extensively excreted via the bile in all animal species, > or =90%, mainly as non-conjugated parent compound. However, the local concentrations in bile varied between the species. Highest concentrations were obtained in the dog. The bile/plasma concentration ratio was 3400 in the dog, 300 in the monkey and 50 in the rat. In the dog, bile duct concentrations of susalimod about 30,000 micromol/l was obtained at plasma concentrations approximately similar to those at which hepatobiliary toxicity occurred, while in rat and monkey the levels were < or =7000 micromol/l at plasma concentrations similar to those obtained at the highest doses in the toxicology studies. From these results supported by a previous biliary excretion study in conscious dogs with chronic bile fistula receiving repeated administration of susalimod (Pahlman et al. 1999), it is likely that the hepatotoxic findings in dog are induced by the high concentrations of susalimod in the bile duct. PMID- 10522752 TI - The effect of cimetidine on the pharmacodynamics of theophylline-induced seizures and ethanol hypnotic activity. AB - Due to its availability as an over-the-counter drug, the use of cimetidine is increasing, thus adverse interactions with other commonly used agents may also increase. The aim of this study was to investigate whether acute administration of cimetidine could alter the pharmacodynamics of theophylline neurotoxicity and the hypnotic action of ethanol. To examine these questions, rats received a dose of 77 mg/kg cimetidine followed by a constant infusion of either theophylline (1.2 mg/min.) or ethanol (16.3 mg/min.) until the onset of the pharmacological end point, maximal seizure or loss of righting reflex, where samples of blood and brain were obtained and assayed for either theophylline or ethanol. We report that cimetidine in doses that may cause pharmacokinetic interactions did not affect the concentration-effect relationship of either the stimulating action of theophylline or the depressant activity of ethanol. These outcomes emphasize the relative safety which patients using cimetidine in self-medication rely on. PMID- 10522753 TI - Hydroxyl radical formation following methamphetamine administration to rats. AB - Administration of neurotoxic doses of methamphetamine (8 mg/kg, intraperitoneally x 4 times, at 2 hr intervals) caused a significant decrease in dopamine and 3,4 dihydroxyphenylacetic acid and an increase in 3-methoxytyramine levels in the striatum along with a decrease in serotonin and 5-hydroxyindoleacetic acid levels in the striatum and hippocampus. In addition, the methamphetamine treatment caused an increase in rat rectal temperature. Intraventricular injection of salicylate 105 min. after the last injection of methamphetamine produced an increase in 2,3- and 2,5-dihydroxybenzoic acid in the striatum and hippocampus. Moreover, the ratio of 2,3-dihydroxybenzoic acid to salicylate was significantly increased in the striatum, but not in the hippocampus. These results indicate that the hydroxyl radical may play an important role in methamphetamine-induced neurotoxicity in rat striatum and that its formation may be the result of methamphetamine-induced release of dopamine. PMID- 10522754 TI - Plasma and brain levels of oxindole in experimental chronic hepatic encephalopathy: effects of systemic ammonium acetate and L-tryptophan. AB - It has previously been shown that the neurodepressant L-tryptophan metabolite oxindole is increased in the blood and brain of rats with fulminant hepatic failure and in the blood of cirrhotic patients affected by chronic hepatic encephalopathy. In the present investigation, we found that oxindole levels were significantly increased in the blood and brain of portacaval-shunted rats, an animal model of chronic hepatic encephalopathy, compared with sham-operated controls. A further increase in plasma and brain oxindole content was found after oral administration of L-tryptophan (300 mg/kg) to both portacaval-shunted or sham-operated animals, while intraperitoneal injection of the amino acid did not modify oxindole content either in brain or blood. Ammonium acetate administration (4.0 mmol/kg, intraperitoneal) reversibly deteriorated the neurological status of portacaval-shunted animals, but did not modify, in a directly related manner, plasma and brain oxindole content. The present findings are in line with the possibility that oxindole may be an additional L-tryptophan-related candidate in the pathogenesis of chronic hepatic encephalopathy. PMID- 10522755 TI - Long-term alterations in benzodiazepine, muscarinic and alpha-amino-3-hydroxy-5 methylisoxazole-4-propionic acid (AMPA) receptor density following continuous cocaine administration. AB - Animal models of clinical phenomena, such as stimulant-induced psychosis have focused primarily on persisting alterations that develop in brain after chronic stimulant administration. The present study utilized autoradiographic measures to examine changes in the density of benzodiazepine ([3H] flunitrazepam), muscarinic ([3H] quinuclidinyl benzilate), and non-NMDA glutamatergic (3H alpha-amino-3 hydroxy-5-methylisoxazole-4-propionic acid: AMPA) receptor binding in rats 21 days following two exposures to cocaine administered continuously for 5 days via subcutaneous pellets. A marked, selective increase in [3H] flunitrazepam binding in both the lateral and medial habenula nucleus was observed. Reduced [3H] quinuclidinyl benzilate binding was observed in various brain areas, including large decreases in the anterior cingulate cortex and ventral thalamus. A reduction in [3H]AMPA binding was observed in the ventral striatum and was suggested in the nucleus accumbens. [3H] Flunitrazepam binding was also examined 12 hr following a single 5 day cocaine exposure to determine if the long-term habenular changes were evident at acute withdrawal. No alterations in [3H] flunitrazepam binding were observed in the habenula or any other structure analyzed at this time point. The relation of these results to persisting alterations in mesocorticolimbic pathways and previous findings of cocaine induced degeneration in lateral habenula circuitry is discussed. PMID- 10522756 TI - Progress in cancer gene therapy. AB - Many technical difficulties have to be overcome before effective gene therapy can be achieved. Strategies for gene therapy include 'suicide' gene therapy, transfer of a tumor suppressor gene, inhibition of activated oncogenes by antisense mechanisms, and cytokine gene transfer and tumor cell vaccination. Gene therapy will have a major impact on the healthcare of our population only when vectors are developed that can safely and efficiently be injected directly into patients as drugs. One of the most promising areas of vector development is that of non viral vectors, which consist of liposomes, molecular conjugates, and naked DNA delivered by mechanical methods. Future research should be focused on modifying viral vectors to reduce toxicity and immunogenicity, increasing the transduction efficiency of non-viral vectors, enhancing vector targeting and specificity, regulating gene expression, and identifying synergies between gene-based agents and other cancer therapeutics. The evaluation of gene therapy combinations is another important area for future research. The identification of tumor rejection antigens from a variety of cancers and the immune response that is defective in cancer patients are important topics for future studies. A universal gene delivery system has yet to be identified, but the further optimization of each of these vectors should result in each having a unique application. Gene therapy has still a long way to go and requires the efforts of investigators in the basic and clinical sciences. Despite substantial progress, a number of key technical issues need to be resolved before gene therapy can be effectively applied in the clinic. PMID- 10522757 TI - Innovations and dilemmas in psychosocial oncology. Contributions from the 10th conference of the European Society for Psychosocial Oncology. PMID- 10522758 TI - Cancer prevention. AB - Over 70% of human cancers are associated with lifestyle and about half of cancer deaths could be prevented by relatively simple individual actions: no smoking, moderate consumption of alcohol, increased consumption of fruit and vegetables, avoidance of sunbathing, obesity and a too high consumption of saturated lipids. Most of these efforts would also markedly decrease the incidence of cardiovascular and respiratory diseases. However, the concept of prevention is currently neither well accepted nor understood by the medical community and the general public. It is often felt that it restricts freedom, imposes a choice between pleasure and duty, and that passing judgement on lifestyle is a form of intolerance. The case of tobacco illustrates the difficulties encountered by prevention, notably among adolescents. The fight against smoking requires information, a societal approach (ban on advertising, increase in price), and a reduction of the example given by adult smoking (parents, peers, teachers, physicians, TV presenters, movie stars, have a great influence on adolescents), while tobacco cessation programs must be promoted. The various approaches should be integrated into a global program of health prevention, including health education at school from 5 to 12 years of age. The efficacy of each of the global program's components should be evaluated. Misconceptions such as overestimation of the impact of pollution on health should also be corrected. Health is created and experienced by people within the setting of their daily lives, in particular during childhood. Prevention is the responsibility of individual members of the community but also of the community as a whole. PMID- 10522759 TI - Using cross-cultural input to adapt the Functional Assessment of Chronic Illness Therapy (FACIT) scales. AB - Cross-cultural quality of life measurement and psychosocial assessment in oncology have become reality with the translation and international validation of quality of life questionnaires. The Functional Assessment of Chronic Illness Therapy (FACIT) Measurement System, under development since 1987, produced its 4th version in November 1997. The FACIT system includes the Functional Assessment of Cancer Therapy (FACT), the Functional Assessment of Human Immunodeficiency Virus Infection (FAHI) and the Functional Assessment of Multiple Sclerosis (FAMS). These questionnaires were developed in North America and, to date, many have been translated into almost 30 languages. One of the strengths of this ongoing translation project is its use of input from patients, linguists, psychologists and physicians internationally to assure that the wording of Version 4 is more cross-culturally relevant and more sensitive to measuring the psychosocial impact of illness in cultures outside the United States. Statistical analysis, aggregation of patient comments, and comments from linguists and users of the translated versions are used as needed to revise item wording to ensure clarity and consistency across languages. A 'decentering' approach is also used to adapt the source (English language) document in response to problems encountered during its translation. PMID- 10522760 TI - Interface between research and practice in psycho-oncology. AB - Examination of the interface between research and practice in any field inevitably raises questions over whether the most important issues are addressed by researchers, or indeed whether the findings of studies have sufficient relevance to practice. As a field of study develops its own research methods and language, a chasm often opens between the producers of research findings and the consumers. Psychosocial oncology is no different. Early work which highlighted the psychosocial impact of a cancer diagnosis, and how health professionals helped or hindered coping with the disease, was ground breaking, and highly relevant to the way cancer services subsequently developed. However, as psychosocial oncology has evolved into an established research discipline, it has become increasingly oriented around measurement (e.g., quality of life, psychopathology, communication skills). The paradox here is that the more reliable psychosocial measures become, the less direct relevance they appear to have for everyday practice in cancer treatment centres. Solutions to this problem could be found through reintegrating psychological and physical aspects of cancer; by changing the orientation of research from measurement of the disruption imposed by cancer and its treatment, to evaluations of more clinically relevant models of care; and by using collaborative models of research in studies in order to promote closer involvement of health professionals and people who have cancer. PMID- 10522761 TI - Revealing response shift in longitudinal research on fatigue--the use of the thentest approach. AB - In this study we examined whether response shift resulting from changes in internal standards occurs in cancer patients undergoing radiotherapy. Ninety-nine newly diagnosed patients undergoing radiotherapy were administered two standardized self-report measures of fatigue prior to receiving radiotherapy. After completion of radiotherapy, patients filled out these questionnaires as a conventional posttest and in reference to how they perceived themselves as they were prior to radiotherapy (a so-called 'thentest'). A transition (direct change) score on fatigue was used as a stratification measure. Patients were subsequently interviewed about their responses. The pattern of mean scores indicative of response-shift effects was found in two distinct subgroups: patients experiencing diminishing levels of fatigue and patients facing early stages of adaptation to increased levels of fatigue. Since response shift may adversely affect the results of self-reported outcomes in clinical trials or other longitudinal research, further research is very much needed. PMID- 10522762 TI - Behavioral interventions in the diagnosis, treatment and rehabilitation of children with cancer. AB - Behavioral interventions used to reduce distress and increase cooperation in children undergoing cancer treatment incorporate: contingency management, cognitive/attentional distraction, hypnosis, systematic desensitization, modeling and behavioral rehearsal. In most cases clinical interventions integrate these procedures into a multimodal intervention package. Although in most behavioral interventions the 'therapist' is a nurse, social worker or child psychologist; parents often take an active role in behavioral intervention. Early return to school can 'normalize' the child's life in the midst of coping with cancer and can promote optimal rehabilitation. More research is needed on the integration of behavioral methods with other therapeutic methods (e.g., pharmacologic). Indeed, research in this area of pediatric oncology must be continuously updated as advances in other areas may affect clinical decisions regarding preferred psychosocial intervention methods. PMID- 10522763 TI - Knowledge of no return--getting and giving information about genetic risk. AB - New genetic information can provide constructive preventive possibilities for individuals and for society but can also create new dilemmas for them. In consultations dealing with the risk of inheriting cancer, many problems connected with the notion of information exchange come to the surface. Individuals have to deal not only with the information given by the doctor, or how to give information to the doctor, but also with the problem of informing others, close kin with a similar risk potential, or getting information from them. In all these information exchanges concerning risk of cancer, different notions of 'information' are being invoked and used as resources in the talks, implying communicative problems at different levels. Some of these problems are discussed in this article. PMID- 10522764 TI - Will cancer risk assessment and counseling services survive genetic testing? AB - With the availability of genetic testing to detect increased hereditary susceptibility to breast and other cancers, Cancer Risk Assessment and Counseling services have come to be viewed by many primarily as a means of obtaining genetic testing and hereditary risk information. The public and healthcare professionals need to be aware that even when genetic testing is not used or is uninformative, families with and without a strong family history of cancer will benefit from Cancer Risk Assessment and Counseling if the process includes assessment of cancer risk, information about cancer etiology, help in dealing with the psychosocial consequences of the cancer experience, and development of emotional and medical coping strategies. Risk assessment services can best serve concerned individuals and their families when sufficient time is allotted for these primary aims, and when genetic testing is seen as one of the useful tools, not the primary goal of such services. PMID- 10522765 TI - Illness-related distress: does it mean the same for men and women? Gender aspects in cancer patients' distress and adjustment. AB - Gender differences were investigated in a sample of 149 married cancer patients (82 males, 67 females) undergoing outpatient chemotherapy. A cross-sectional design was used and evaluation included medical assessments and self-rating questionnaires. Tumour sites varied, and advanced stages of disease were predominant. Overall, the results suggest gender differences as well as some similarities. Although female patients reported symptoms and higher overall distress because of illness more frequently than male patients did, general satisfaction with life did not differ between genders, suggesting comparable adjustment. From the results of multivariate analyses physical impairment, such as older age, primarily explained female patients' distress, whereas men's distress was closely linked to their psychological condition. Men and women also differ in the way they make use of social support. Assessment of the distinctive aspects contributing to male and female cancer patients' distress could improve the provision of adequate support adapted to gender-specific requirements. PMID- 10522766 TI - Mental distress--gender aspects of symptoms and coping. AB - The article examines men's and women's views on their reasons for mental distress and on their coping styles, respectively. The data were taken from written statements given on two open-ended questions from a survey questionnaire returned by 43 men and 57 women who were self-reported, long-term users of these drugs, and from taped interviews with 10 respondents. Men's accounts (n = 25) expressed a layered theory of mental health: alcohol was a remedy to alleviate temporary strain caused by external pressure, while the use of psychotropic drugs indicated a loss of a men's assumed self-regulatory powers and autonomy. Women's accounts (n = 31) were stories of emotional pain related to their caring work in the private sphere, and psychotropics restored their capacity to carry out emotional labor. PMID- 10522767 TI - Effects of a nursing intervention on subjective distress, side effects and quality of life of breast cancer patients receiving curative radiation therapy--a randomized study. AB - The purpose of this randomized study was to investigate whether a nursing intervention using Orem's self-care theory as a framework would affect subjective distress, side effects and quality of life as perceived by breast cancer patients receiving curative radiation therapy. The intervention consisted of five 30-min sessions once a week during the treatment period and two follow-up sessions after completion of treatment. The experimental group consisted of 67 patients, as did the control group. Measurements were collected five times: at baseline before commencement of treatment, at weeks 3 and 5 (completion of treatment) and follow up periods of 2 weeks and 3 months. No measurable effect of the nursing intervention was found for side effects or quality of life but nursing intervention proved to have a positive effect in minimizing stress reactions (p = < 0.05). It is suggested that a nursing intervention should be implemented for breast cancer patients receiving curative radiation therapy. PMID- 10522768 TI - Disentangling ethical and psychological issues--a guide for oncologists. AB - The rapid growth of bioethics has injected a new style of analysis into medicine. It requires philosophical rigor, yet is deeply embedded in human situations that frustrate abstract thinking and are laced with subjective factors. These interlaced ethical and psychological components can lead to conflicts and dilemmas. Doctors, as experts and decision-makers, play a key role, but will benefit from additional skills to disentangle these situations. This paper notes ways in which patients, families and caregivers are newly vulnerable and delineates how ethical dilemmas and psychological issues mold or frustrate decision-making. To help physicians manage such cases, a method of systematic analysis, the 'situational diagnosis', and a related hierarchy of interventions, is described and illustrated with case examples. PMID- 10522769 TI - Small-cell lung cancer in the elderly--is age of patient a relevant factor? AB - Different management procedures for diagnosis and treatment of small-cell lung cancer (SCLC) and other tumours in the elderly have been reported, but there is a lack of data from a communal hospital perspective. Information on clinical parameters such as weight loss, co-morbidity, performance status and investigative procedures for staging of disease and inclusion in clinical trials was recorded for patients in the province of Albacete (Spain). Patients' ages were categorized in two groups: under 70 years and 70 years or more, and a comparison of treatment variables, toxicities, response and time to event measures was carried out. Ninety-five patients were referred to our Unit for treatment. Of these patients, 62% were under 70 years of age and 38% were in the older age category. Clinical variables and staging procedures did not differ between groups. Trial assignment showed a bias in favour of younger patients (11 vs. 1, p = 0.02). No differences in the number of patients without treatment were found, but the older group presented fewer cases of optimal (> or = 4 cycles) therapy, less chemotherapy delivery (smaller mean total doses of cisplatin and etoposide) and smaller mean total dose of radiotherapy (57/45 Gy). The response to treatment (46%/50%) toxicity registered and overall survival did not differ between age categories. Age does not seem to be a relevant prognostic factor in this disease. Carefully calculated dose reductions for chemotherapy in elderly patients based on initial performance status and/or toxicity during treatment may be a useful policy without detrimental implications on outcome. PMID- 10522770 TI - Optimization of the dose level for a given treatment plan to maximize the complication-free tumor cure. AB - During the past decade, tumor and normal tissue reactions after radiotherapy have been increasingly quantified in radiobiological terms. For this purpose, response models describing the dependence of tumor and normal tissue reactions on the irradiated volume, heterogeneity of the delivered dose distribution and cell sensitivity variations can be taken into account. The probability of achieving a good treatment outcome can be increased by using an objective function such as P+, the probability of complication-free tumor control. A new procedure is presented, which quantifies P+ from the dose delivery on 2D surfaces and 3D volumes and helps the user of any treatment planning system (TPS) to select the best beam orientations, the best beam modalities and the most suitable beam energies. The final step of selecting the prescribed dose level is made by a renormalization of the entire dose plan until the value of P+ is maximized. The index P+ makes use of clinically established dose-response parameters, for tumors and normal tissues of interest, in order to improve its clinical relevance. The results, using P+, are compared against the assessments of experienced medical physicists and radiation oncologists for two clinical cases. It is observed that when the absorbed dose level for a given treatment plan is increased, the treatment outcome first improves rapidly. As the dose approaches the tolerance of normal tissues the complication-free cure begins to drop. The optimal dose level is often just below this point and it depends on the geometry of each patient and target volume. Furthermore, a more conformal dose delivery to the target results in a higher control rate for the same complication level. This effect can be quantified by the increased value of the P+ parameter. PMID- 10522771 TI - Positron emission tomography with 18-fluorodeoxyglucose in the staging and follow up of lymphoma in the chest. AB - The purpose of this retrospective study was to evaluate the accuracy of positron emission tomography (PET) using 18-F-fluorodeoxyglucose (FDG) in predicting lymphomatous involvement in the hilar and mediastinal regions in the staging and follow-up of patients with malignant lymphoma. One hundred forty-seven thoracic PET studies in 89 consecutive lymphoma patients were reviewed. Static FDG-PET imaging was performed following application of 270 MBq FDG (mean). Results of FDG PET were compared with the findings of computed tomography (CT) in all patients and clinical follow-up examination. Eighty-nine of 147 (60%) PET studies showed no FDG uptake in the hilar or mediastinal regions, while 58 (40%) studies did detect FDG uptake in these regions. In 52 of 58 abnormal studies (90%), lymphomatous involvement of the hilar and/or mediastinal regions seen by CT was present. In the remaining six abnormal PET studies (10%), FDG uptake was considered as false-positive because of missing lesions on corresponding CT scans. In four patients false-positive FDG uptake was observed before treatment, in two patients after completion of therapy. In these two patients FDG uptake after therapy was caused by thymus hyperplasia. The remaining four cases before treatment remained unresolved. Sensitivity of FDG-PET was 96%, specificity 94%, positive predictive value 90%, and negative predictive value 98%, respectively. The present study suggests that FDG-PET has potential value in predicting lymphomatous involvement in the hilar and mediastinal regions. FDG-PET may obviate invasive diagnostic procedures in patients with lymphoma. PMID- 10522772 TI - Immunoradiometric measurement of pS2 in breast cancer--correlation with steroid receptors and plasminogen activators. AB - pS2 was measured by radioimmunometric assay in tumour extracts from 197 breast cancer patients. Values ranged from 0 to 50 ng/mg protein (mean 9.6 and median 3 ng/mg). We found no correlation with age, menopausal status, nodal metastases, disease stage or tumour histology. There was, however, a linear relationship with both ER (p < 0.0001) (particularly nuclear ER) and PR (p < 0.0001) expression determined by enzyme immunoassay (ELISA), as well as a good correlation when high and low expressors were stratified on the basis of combined ER/PR expression using consensus cut-off points. Only 15% of ER - ve/PR - ve patients were classified as pS2 + ve compared with 83% of those who were ER + ve/PR + ve. pS2 was also directly correlated with high expression of tPA and inversely with uPA. Comparison with previous studies showed that the current ELISA method produced consistent results, in contrast to other methods, particularly those based on immunohistochemical detection. The close relationship between pS2 and both steroid receptors suggests that pS2 may be important in terms of defining hormone responsive patients who are likely to benefit from endocrine therapy. PMID- 10522773 TI - Bone tissue reaction around implants placed in a compromised jaw. AB - The present experiment was carried out to examine bone tissue alterations that occurred around implants at which the marginal level of bone support at fixture installation was different at buccal and lingual surfaces. 8 beagle dogs were randomly divided into one test group and one control group. The mandibular premolars in the left side of the mandible (P1, P2, P3, P4) were extracted. In the 4 dogs of the test group, the buccal bone plate in the mandibular premolar region was removed to establish a bone defect that was about 25 mm long, about 5 6 mm high, and about 4 mm wide. In the 4 dogs of the control group, no bone resection was performed. 8 months after tooth extraction, 3 fixtures (Astra Tech AB, Molndal, Sweden:TiO-blast: 8x3.5 mm) were installed in each dog. In the 4 dogs of the test group, the implants were positioned in the defect sites in such a way that (i) mechanical stability was achieved and (ii) their lingual surfaces were entirely invested in bone. At the buccal and approximal surfaces of the fixtures, however, the unthreaded portion (2 mm) and the 3 marginal threads remained exposed. In the control group, all implants were following installation entirely surrounded by bone tissue. After a healing period of 3 months, abutment connection was performed and a plaque control program initiated. 4 months later, the dogs were sacrificed. The mandibles were removed and placed in a fixative. Each implant region was dissected, the tissue samples were dehydrated, embedded, sectioned in a bucco-lingual plane and used for light microscopic examination. The findings demonstrated that osseointegration occurred at implants, placed in a chronic defect with large discrepancies between the buccal and lingual bone. During the process of healing and function, however, marked modeling and remodeling of the bone tissue took place. Thus, at the buccal surface, some bone regrowth and osseointegration occurred while at the lingual wall, there was a substantial resorption of the marginal bone and an enhanced number of bone multicellular units. Concomitant with the bone tissue alterations described, there was some recession of the peri-implant mucosa. PMID- 10522774 TI - The effects of nicotine and age on replication and viability of human gingival fibroblasts in vitro. AB - The aim of the present study was to examine: (1) the effects of nicotine on human gingival fibroblasts (HGF); (2) differences between smokers (> or = 10 cigarettes/day at least for 5 years) and non-smokers; (3) differences between patients of different age. HGF were obtained, through biopsies during periodontal surgical procedures, from 15 patients which were divided in 4 groups: 4 patients, smokers aged < or = 25 years; 4 patients, non-smokers aged < or = 25 years; 3 patients, smokers aged > or = 40 years; 4 patients, non-smokers aged > or = 40 years. Nicotine has been tested in 3 different concentrations: 6 microg/ml; 60 microg/ml; 600 microg/ml. To assess cells viability, the neutral red (NR) test was used; to evaluate cell proliferation, the Hoechst test was employed. After 48 h of nicotine exposure, it was found that 600 microg/ml nicotine was strongly cytotoxic to HGF of all groups, with a significant reduction of both proliferation and viability of cells versus control. Comparison between groups of the same age: when comparing untreated HGF (i.e., control values) of smokers < or = 25 years versus non-smokers < or = 25 years, cell proliferation, but not viability, was found to be increased in smokers. Both viability and proliferation of control cells of smokers > or = 40 years were increased versus non-smokers > or = 40 years. HGF of non-smokers < or = 25 years, when exposed to nicotine 600 microg/ml, have less viability and proliferation than HGF of smokers of the same age. Comparison between groups of different age: In the smoker group, untreated HGF (i.e., control values) had similar viability and proliferation, irrespective of age, but nicotine 600 microg/ml kills more HGF in smokers < or = 25 years than in smokers > or = 40 years. In non-smokers, untreated HGF < or = 25 years replicate less, but are not less viable than HGF > or = 40 years. When challenged with nicotine 600 microg/ml, HGF < or = 25 years were less viable than HGF > or = 40 years. From this study, it appears that the smoking history and the patient age could be relevant for final evaluation of the results, since HGF from smokers are less sensitive to nicotine than HGF from non-smokers, and cells from older donors are more resistant to nicotine than cells from younger donors. PMID- 10522775 TI - Bone formation in furcation defects. An experimental study in the dog. AB - The aim of the present investigation was to study bone formation in an experimentally-produced furcation defect in the dog. 15 foxhound dogs (group A) and 4 large mongrel dogs (group B) were used. The 2nd and 4th mandibular premolars were extracted and the 3rd lower premolars (3P3) were assigned as experimental teeth. "Through and through" furcation defects, about 4 mm high and 3 mm wide, were first produced in the experimental teeth of the dogs in group A. Reconstructive surgery was subsequently performed in group A using a GTR technique. The dogs of group A were scheduled for biopsy 2 weeks (2 dogs), 1 month (2 dogs), 2 months (2 dogs), 4 months (3 dogs), 5 months (3 dogs) and 6 months (3 dogs) after GTR. The dogs in group B (4 animals) represented healthy, untreated pristine furcations and served as positive controls. Biopsies from the 3P3 regions were harvested, embedded in paraffin and prepared for histological analysis. Mesio-distal sections were cut with the microtome set at 7 microm. The sections were stained in hematoxylin and eosin, and Van Gieson. 3 sections, about 50 microm apart, and representing the central portion of the furcation site were selected for histological measurements. In group A, the proportions of various structures in the newly formed bone and marrow were assessed. In addition, the proportions of primary and secondary osteons, and the number of bone multicellular units (BMU)/mm2 mineralized bone tissue were determined. In the pristine furcations (group B), the histological analyses were performed in a corresponding area to that of the healing furcations. The results demonstrated that the process of bone formation in a large "suprabony" furcation defect can be divided into 3 different phases, namely, (i) the formation of a provisional connective tissue, (ii) the development of a primary bone spongiosa (including mainly woven bone), (iii) the replacement of the spongiosa by lamellar bone and bone marrow through processes of modeling and remodeling. The newly-formed trabeculae of woven bone were reinforced by the deposition of parallel-fibered bone and lamellar bone, a finding which substantiates the validity of the concept that woven bone possesses poor weight-bearing properties and, hence, needs to be re-inforced by a more mature type of bone. The modeling of the newly-formed bone resulted in the formation of (i) one large marrow space in the center of the furcation and, in addition, (ii) a smaller bone marrow space in the most coronal portion of the defect. At the end of the study (6 months), the bone marrow occupied a much larger space than in the bone tissue of pristine furcations. It was suggested that the process of modeling or remodeling of bone tissue in the furcation defect was not completed at the end of the study. PMID- 10522776 TI - Calprotectin in gingival crevicular fluid correlates with clinical and biochemical markers of periodontal disease. AB - Clinical and biochemical markers of periodontal disease have been used for precise objective diagnosis of periodontal inflammation. Interleukin 1beta (IL 1beta) and prostaglandin E2 (PGE2), inflammatory factors, levels in gingival crevicular fluid (GCF) of patients with periodontal disease are elevated and have been studied as biochemical markers. The levels of calprotectin, a leukocyte protein, in body fluids of patients with some inflammatory diseases are raised. Recently, we detected calprotectin in GCF and its concentrations in periodontal pockets were higher than those in healthy gingival crevices. In this study, we investigated the correlations between GCF calprotectin levels and clinical indicators (probing depth and bleeding on probing, BOP), and the IL-1beta or PGE2 levels in GCE Probing depth and BOP at 130 sites of 110 subjects with periodontal or other oral diseases were examined, then GCF samples were collected and their calprotectin, IL-1beta and PGE2 were determined by ELISA. The calprotectin level correlated positively with the probing depth and was significantly higher at BOP positive than BOP-negative sites. There were significant, positive correlations between the calprotectin and IL-1beta or PGE2 concentrations. These results indicate that the calprotectin level in GCF correlates well with clinical and biochemical markers of periodontal disease and suggest that calprotectin may be useful for evaluating the extent of periodontal inflammation. PMID- 10522777 TI - The barrier between the keratinized mucosa and the dental implant. An experimental study in the dog. AB - The present study was performed in order to examine the composition of the connective tissue that forms an attachment to a dental implant. 6 beagle dogs were used. All mandibular premolars were extracted. After 3 months of healing, 6 fixtures--3 in each side of the mandible--(Astra Tech Implants, Dental System TiO blast; Astra Tech AB, Molndal, Sweden) were installed. After another 3 months of healing, abutment (Uni-abutment 45; Astra Tech AB, Molndal, Sweden) connection was performed and a plaque control program was initiated. The animals were sacrificed and perfused with a fixative through the carotid arteries. Each implant site, including the implant and the soft and hard peri-implant tissues, was dissected, decalcified in EDTA and further processed using a "fracture technique". The specimens were subsequently embedded in EPON, cut with the microtome set at 3 microm and the sections stained in PAS and toluidine blue. From the EPON-embedded blocks, ultra-thin sections were cut and electron micrographs were prepared. The detailed histologic and morphometrical examinations were restricted to a 200 microm wide zone of connective tissue interposed between the apical border of the junctional epithelium and the bone tissue. In the analysis, this zone was further subdivided into 2 different units; (i) one central, 40 microm wide unit (zone A) located immediately next to the implant surface, and (ii) one lateral, 160 microm wide unit (zone B) that was continuous with the central unit. The implant surface apical of the junctional epithelium and coronal of the bone crest appeared to be in direct contact with a connective tissue. Zone A of this connective tissue was characterized by its (i) absence of blood vessels and (ii) abundance of fibroblasts which were interposed between thin collagen fibers. The more lateral zone B contained comparatively fewer fibroblasts, but more collagen fibers and blood vessels. There are reasons to assume that the fibroblast rich barrier tissue next to the titanium surface plays a role in the maintenance of a proper seal between the oral environment and the peri-implant bone. PMID- 10522778 TI - Associations of medical status and physical fitness with periodontal disease. AB - To determine the possible associations of medical status and physical fitness with periodontal disease, a cross-sectional study was conducted. The subjects were 517 males and 113 females aged 23 to 83 years who participated in a multiphasic health test at the Aichi Prefectural Center of Health Care, Japan, from 1992 to 1997. Their periodontal status was assessed by means of the CPITN scoring system. To assess the strength of associations between the examined factors and the score, odds ratios were computed using ordinal logistic models. Conventional risk factors such as old age, smoking habits, and higher fasting plasma glucose and simplified debris index increased the risk of periodontal disease. Hypertension, hematuria, leucocytosis or thrombocytosis, positive C reactive protein and higher serum alkaline phosphatase were positively associated with the score, whereas higher serum high-density lipoprotein cholesterol was related to a lower risk. Poor physical fitness affecting aerobic capacity, foot balance and reaction was associated with a higher CPITN score. These associations were independent of the conventional risk factors. Although these new potential risk factors should be further investigated for their causal relationship, our findings suggested a close relationship of oral health to medical status and physical fitness. PMID- 10522779 TI - Factors influencing the success of GBR. Smoking, timing of implant placement, implant location, bone quality and provisional restoration. AB - The aim of this retrospective clinical study was to evaluate the influence of different factors on the outcome of GBR treatment. 75 patients, who were not randomly assigned to the investigated parameters for clinical reasons, were included in the study. They presented with defect sites around implants and were treated with a xenogenic grafting material and a resorbable collagen membrane. The defect morphology was described, its dimension was measured and calculated at the time of implant installation and at re-entry. The success of GBR treatment was related to several clinical variables and possible correlations were evaluated. Defect sites around maxillary implants showed significantly more bone fill (96%) compared to those in the mandible (78%). The insertion of a provisional restoration during the healing period was also associated with significantly better results than when no provisional was inserted. Immediate and short-term delayed implant placements showed the best results both with 92% bone fill, when compared with long-term delayed placements with 80% bone fill (n.s.). In sites with type I bone quality (compact bone), a reduced bone fill was observed (64%). The results indicate that successful bone fill can be achieved with GBR; this is more feasible in the maxilla, when a provisional restoration is used. Early implant placement timings seem to be preferable due to the alveolar ridge preservation, more favorable defect morphologies and a higher regenerative capacity. PMID- 10522781 TI - Quantification of root surface plaque using a new 3-D laser scanning method. AB - There are no published reports in the literature objectively quantifying thickness of plaque on teeth. The aim of this study was to quantify plaque on a tooth surface and assess if this quantification correlates with a clinical index of plaque from each of 51 patients. Patients were instructed not to perform any oral hygiene on the day of the assessment. The Silness and Loe plaque index was scored and replicas were scanned using a co-ordinate measuring machine (CMM) and laser scanning probe. A replica was obtained from this surface before and after toothbrushing. Plaque adjacent to the gingival margin had a mean thickness of 0.106+/-0.118 mm (mean+/-SD) whilst mean plaque thickness 250 microm from the gingival margin was 0.053+/-0.052 mm (mean+/-SD). There was a significant correlation between the plaque index and the plaque thickness (p < or = 0.002). The finding that plaque is present in the greatest amount adjacent to the gingival margin supports a previously reported hypothesis that primary root carious lesions (PRCL's) may initiate adjacent to the gingival margin. This method quantifies plaque thickness on exposed root surfaces which correlates with the plaque index as well as illustrating how the morphological characteristics of teeth, gingivae and plaque can be studied in vivo from replicas recorded. PMID- 10522780 TI - Effects of smoking on local delivery of controlled-release doxycycline as compared to scaling and root planing. AB - This paper examines the effects of smoking on the treatment outcomes of two nonsurgical therapies: (1) scaling and root planing alone (SRP) or (2) controlled release of subgingivally delivered doxycycline hyclate in a polylactic acid based polymer gel. Subjects from 2 9-month multicenter studies were classified as nonsmokers (never smoked: 100 subjects), former smokers (137 subjects), and current smokers (> or = 10 cigarettes/day: 121 subjects). Clinical parameters were analyzed for treated sites with baseline probing depths > or = 5 mm and for a subset of treated sites with baseline probing depths of > or = 7 mm. Clinical parameters (plaque levels, clinical attachment levels, pocket depths, and bleeding on probing) were analyzed at baseline, 4, 6, and 9 months. In the doxycycline treated group in general, there were neither marked significant differences in clinical attachment gain nor differences in probing depth reduction among the 3 smoking groups. On the other hand, in the scaling and root planing treated group in general, there were significant differences in clinical attachment gain and pocket depth reduction, with non-smokers responding better than former smokers and current smokers at 6 and 9 months. These differences in clinical response between scaling and root planing alone versus controlled release of locally-delivered doxycycline hyclate among these 3 smoking groups are discussed in relation to treatment implications for smokers. PMID- 10522782 TI - The effect of steroidal and non-steroidal anti-inflammatory drugs on the cellular immunity of calves with experimentally-induced local lung inflammation. AB - We examined the effect of a single intravenous dose of flumetasone (SAID) and meloxicam (NSAID) treatment of calves with experimentally-induced localized lung inflammation on immunological and hematological variables such as total protein, gamma globulin, hemoglobin (Hb) concentrations, alkaline phosphatase activity, packed red cell volume (PCV), red blood cell (RBC) and white blood cell (WBC) counts. The influence of drug treatment on the phagocytic activity of WBC and bronchoalveolar lavage (BAL) cells and their ex vivo ability to produce interferon (IFN) and tumor necrosis factor (TNF) after induction with Newcastle disease virus (NDV), as well as on the development of PHA-induced skin delayed hypersensitivity reaction was also determined. Two days after the treatment of calves with experimentally-induced local lung inflammation with flumetasone (5 mg per calf), we observed a significant increase in WBC count, especially neutrophils, and a decrease in gamma globulin concentration, in the percent of blood phagocytic cells and their random migration. Flumetasone treatment also inhibited the development of skin delayed hypersensitivity reaction. In contrast, the treatment of calves with meloxicam (50 mg per calf) did not influence any hematological parameters or skin reactivity. Both drugs, flumetasone and meloxicam, influenced TNF production in ex vivo cultures of blood and BAL cells, inhibiting excessive TNF production induced by local lung inflammation. Contrary to TNF, the treatment of calves with meloxicam and flumetasone enhanced ex vivo IFN production in blood and BAL cells. Histological examination of lung tissue revealed that in control calves (those not treated with anti-inflammatory drugs) and in calves treated with flumetasone, symptoms of stromo-purulent inflammation of pulmonary tissue developed. However, in calves treated with meloxicam, only interstitial inflammation with a slight thickening of interalveolar septa and infiltration of lymphoid cells was observed. These results suggest that in this model of pneumonia, it is more appropriate to use a single dose of meloxicam, rather than flumetasone, to modulate lung inflammation. PMID- 10522783 TI - Characterization of T-lymphocytes in the anterior uvea of eyes with chronic equine recurrent uveitis. AB - Equine recurrent uveitis (ERU), a chronic, recurrent inflammation primarily of the anterior uveal tract, is the most common cause of blindness in horses. Recently, T-lymphocytes have been found to be the most numerous cell type to infiltrate the anterior uveal of horses with ERU. In the present study, we characterized the T-lymphocyte population in the anterior uveal tract of eyes of horses with chronic ERU by evaluating the microscopic appearance (histopathologic features), the T-lymphocyte subsets, and the relative levels and amounts of T lymphocyte cytokine mRNA in the anterior uvea. Seven inflamed eyes (from six horses with chronic ERU) and 5 normal eyes (from five horses with nonocular problems) were studied. After clinical examination, the eyes were removed, ocular fluids were aspirated, and anterior uveal tissues (iris and ciliary body) were processed for histologic and molecular (RNA isolation) analyses. Histologic examination by hematoxylin and eosin (H and E) staining and immunohistochemistry evaluating T-lymphocyte subsets (anti-CD4, CD8, CD5) were performed for each sample. RNA samples were analyzed for levels of messenger (m) RNA specific for interleukin (IL)-2, 4, and interferon-gamma (IFNgamma) by quantitative reverse transcriptase polymerase chain reaction (QRT-PCR). Eyes with ERU exhibited characteristic clinical signs, including corneal edema, aqueous flare, posterior synechia, corpora nigra degeneration, and cataract formation. Histologically, infiltration of the uveal tract with lymphocytes, plasma cells, and macrophages was most evident in the ciliary body and base of the iris. Loss of tissue structure (destruction) was most evident in the ciliary processes. Infiltrating lymphocytes were predominantly CD4+ T-cells (e.g. 48% CD4+ and 18% CD8+ in the ciliary body stroma), as determined by immunohistochemistry. Few inflammatory cells were observed in the normal eyes. The QRT-PCR results revealed increased transcription of IL-2 and IFNgamma and low IL-4 mRNA expression in eyes with chronic ERU compared to normal eyes, demonstrating a Thelper (Th) 1-like inflammatory response in eyes with ERU. PMID- 10522784 TI - Comparison of a maedi-visna virus CA-TM fusion protein ELISA with other assays for detecting sheep infected with North American ovine lentivirus strains. AB - A maedi-visna virus CA-TM fusion protein ELISA (MVV ELISA) was evaluated for the detection of antibody in sheep infected with North American ovine lentivirus (OvLV). The results of the MVV ELISA were compared with other assays for OvLV antibody and with viral infection in an intensively studied group of 38 sheep with a high prevalence of OvLV infection and disease. The sensitivity, specificity, and concordance of assays for OvLV antibody (MVV ELISA, indirect ELISA, Western blot, and AGID), virus (virus isolation, PCR, antigen ELISA), and OvLV-induced disease in each animal were compared with OvLV infection status as defined by a positive result in two or more of the assays. Five sheep met the criteria for absence of OvLV infection. The sensitivity of the MVV ELISA in detecting OvLV infected sheep was 64%, whereas the sensitivity of the other three tests for antibody ranged from 85 to 94%. All the antibody assays were 100% specific in this group of animals. Of the assays for virus, the PCR test had the highest sensitivity and the best concordance with OvLV infection, but it also had the lowest specificity of any of the virus or antibody assays. Among the antibody tests, the concordance of the MVV ELISA compared most favorably with the AGID test for detecting OvLV-infected sheep. Analysis of serum samples from 28 lambs experimentally-infected with one of three North American strains of OvLV suggested that there were no significant strain differences detectable by antibody assay. Twenty virus-inoculated lambs were positive by both the MVV ELISA and the AGID test, five lambs were MVV ELISA negative and AGID test positive, and three lambs were MVV ELISA positive and AGID test negative. No pre-inoculation samples were positive by either assay. In a longitudinal study involving seven lambs, antibodies to OvLV were detected by AGID 3-5 weeks post-inoculation, but were not detected by MVV ELISA until 5-10 weeks post-inoculation. Among 128 naturally and experimentally-infected sheep that were seropositive in the AGID test, the overall sensitivity of the MVV ELISA was higher in the naturally infected sheep (84%) than in the experimentally infected sheep (69%). The data indicated that the MVV ELISA represents a less sensitive, but specific alternative for the detection of OvLV antibodies. PMID- 10522785 TI - Detection of IgM responses to bovine respiratory syncytial virus by indirect ELISA following experimental infection and reinfection of calves: abolition of false positive and false negative results by pre-treatment of sera with protein-G agarose. AB - The IgM responses in three panels of sera generated by infection and reinfection of calves with bovine respiratory syncytial virus (BRSV) were measured by indirect ELISA (I-ELISA). The effect of depleting serum IgG by pre-treatment with protein G agarose (PGA) was evaluated. Following primary infection a weak IgM response was detected in the untreated sera of 3 out of 4 calves with maternally derived antibody (MDA). Both the magnitude and duration of the specific IgM responses in these calves were increased by pre-treatment with PGA. In addition, the fourth infected calf tested gave a single positive IgM result following PGA treatment. Transient or persistent IgM responses which were abolished by pre treatment of sera with PGA were detected in 4/8 calves following reinfection. These were considered to be false positive results, consistent with the influence of IgM rheumatoid factor (IgM-RF). One of these calves and two additional calves showed transient increases in IgM which were resistant to PGA treatment. These were considered to represent specific IgM responses to reinfection. The results indicate the ability of PGA treatment to eliminate both false positive and false negative results and emphasise the necessity for controlling the influence of IgM RF in IgM-specific indirect ELISAs. PMID- 10522787 TI - Analysis of immune responses in dogs with canine visceral leishmaniasis before, and after, drug treatment. AB - The incidence of canine visceral leishmaniasis (CVL) is increasing in the Mediterranean region. Many drugs have been tested for treatment of CVL, but little is known regarding their effect on test immune responses. In our study, three dogs naturally infected with Leishmania infantum and five dogs experimentally infected with the same strain, were treated with dimethasulfonate pentamidine (Lomidine) and the immune response evaluated before, and after, treatment. After the last injection, animals began to gain weight and the major clinical signs disappeared. Antibody titers gradually decreased to low levels, six months after treatment. At the same time, antigen specific lymphoproliferation reappeared in the sampled animals. This study shows that, after treatment, immune cellular responses to leishmanial antigens, involved in protection against Leishmania infection, were established. PMID- 10522786 TI - Effect of oral rotavirus/iscom vaccines on immune responses in gnotobiotic lambs. AB - A comparison of the effect on the immune responses in gnotobiotic lambs was made between an iscom vaccine prepared from recombinant rotavirus VP6 protein, an inactivated rotavirus/iscom-matrix vaccine and a vaccine comprising inactivated rotavirus alone. All three vaccines induced immunological priming and some degree of protection was observed after a single oral dose. However, different immune responses were induced in response to a virulent infection. The group vaccinated with the rotavirus/iscom-matrix vaccine showed a Th2-like response characterised by rotavirus-specific antibodies and a down-regulation of IFNgamma in jejunal Peyer's patches. Both Th1-like and Th2-like immune responses were induced in the group receiving the VP6 vaccine as seen by significantly increased expressions of IFNgamma and IL-6 in the jejunal Peyer's patch together with an increased percentage of CD8+ T cells in the intestine and rotavirus-specific antibodies at mucosal surfaces. Iscom vaccines given orally have the ability to induce both Th1 like and Th2-like immune responses in a ruminant model. PMID- 10522788 TI - Structural protection of the myocardial capillary endothelium by different forms of cardiac arrest and subsequent global ischemia at 5 degrees C. AB - BACKGROUND: In transplantation surgery sufficient myocardial protection achieved by cardioplegic cardiac arrest and deep hypothermia is a prerequisite for successful resumption of donor heart function. Intraischemic damage of the endothelium combined with capillary compression may lead to the "no-reflow phenomenon" during reperfusion, resulting in insufficient cardiac resuscibility. METHODS: We evaluated the endothelial ultrastructure after various common forms of cardiac arrest and subsequent ischemia in deep hypothermia. Canine hearts were arrested by aortic cross clamping and surface cooling with Tutofusin' (ACC) or by coronary perfusion with Custodiol (histidine tryptophane ketoglutarate, HTK solution), with University of Wisconsin solution (UW), or with St. Thomas' Hospital solution. After extirpation the hearts were incubated at 5 degrees C in the solution used for cardiac arrest. Myocardial samples were taken immediately after cardiac arrest and after 2h, 4h, 6h, and 10 h of global ischemia. The degree of structural damage was evaluated by a scoring system. Endothelial swelling was determined as the mean barrier thickness of the capillary endothelium. RESULTS: At all selected time points our results show that 1) after cardioplegia with St. Thomas' solution, the degree of endothelial cell swelling was higher than after aortic cross clamping; 2) using HTK or UW solution, the endothelial ultrastructure was better preserved than after aortic cross clamping or using St. Thomas' solution, whereby HTK was slightly better than UW; 3) using UW solution, endothelial cell swelling was a little (up to 10%) but significantly less than after HTK perfusion. CONCLUSIONS: With respect to the intraischemic structural preservation of endothelial cells, UW or HTK solution combined with deep hypothermia promises adequate protection, compared with other clinically used methods tested. PMID- 10522789 TI - Management of symptomatic hypertrophic obstructive cardiomyopathy--long-term results after surgical therapy. AB - BACKGROUND: The natural history of Hypertrophic Obstructive Cardiomyopathy (HOCM), is well known from earlier investigations. The yearly death rate of medically or non-treated patients with HOCM is between 1.7% and 4%. After conservative management with beta-blockers and/or calcium antagonist, early improvement is followed in many patients by a symptomatic and clinical impairment, which today may lead to surgical or interventional treatment. METHODS: From 1963 to 12/1998 a total of 519 patients were operated by transaortic subvalvular myectomy (TSM). The mean age was 49 +/- 11 years (range 3 months - 82 years) in 292 males and 227 females. RESULTS: The early risk was related to the clinical class (NYHA) and the need for additional cardiac procedures during the same intervention. Total early mortality was 4.4% (n=23), in isolated myectomy 3.6% (n= 11). During the last 10 years it could be reduced to 1.9%. The first complete (100%) reinvestigation of 346 patients up to 26 years after surgery (1963-1991) demonstrated a disease-related mortality rate of 5.2% (n=20). The analysis of late deaths showed that disease-related lethal complications (sudden death, life-threatening arrhythmias, valve endocarditis, secondary LV dilatation) were relatively rare, the age-related death rate nearly followed the natural course because of other causes. The cumulative survival rate after 10 years was 88%, after 20-26 years 72%. The yearly disease-related death rate could be reduced to 0.6%. The long-lasting, symptomatic clinical improvement (NYHA I-II), and also the physical and mental capacity with enlargement of the acitivity radius and improvement of quality of life were remarkable. The positive effects of surgical enlargement of the LVOT could be confirmed in the meantime by hemodynamic, rhythmological, echocardiographic investigations as well as endurance tests. CONCLUSION: We have examined the outcome of a large series of patients treated surgically for HOCM since 1963. The majority of patients were in NYHA class III and came to surgery after long-term medical, but finally insufficient, management. The perioperative risk could be reduced considerably during recent years, despite the advanced cardiomyopathy status. The long-term postoperative observation of the patients demonstrated an unexpectedly continuing good outcome. Therefore these results may serve as a standard for assessing the results after the less invasive alcohol-induced transcoronary ablation of septal hypertrophy. PMID- 10522790 TI - Special cerebral perfusion in surgery for the ruptured thoracic aortic aneurysm. AB - BACKGROUND: For surgical treatment of the ruptured thoracic aortic aneurysm (TAA), it is important to control bleeding and to protect the brain, spinal cord, and myocardium. We have developed and performed a new procedure on 6 patients with a ruptured TAA, a true aneurysm in 3 patients and a type A dissection in the other 3. METHOD: Cardiopulmonary bypass is installed with cannulations to the iliac artery and vein and to the common carotid arteries on both sides of the neck before the sternum is divided. For control of bleeding, venous drainage is accelerated, whereas cerebral perfusion is maintained via the carotid arteries. After insertion of the occlusion catheters into the descending aorta and the left subclavian artery following the aortotomy, the bypass flow to the iliac artery is increased. RESULTS: The arch replacement was performed in 4 patients and hemiarch replacement in two. Five patients are alive without neurologic deficits; one patient died of multi-organ failure on the 24th postoperative day. CONCLUSIONS: We conclude that our procedure may be advantageous for patients with a ruptured TAA, a large retrosternal aneurysm, or reoperation of the thoracic aorta. PMID- 10522791 TI - Protective effect of thiopental against cerebral ischemia during circulatory arrest. AB - BACKGROUND: One of the most important disadvantages of the hypothermic circulatory arrest technique is the limited time allowable for circulatory arrest. Thiopental is usually used to protect the brain against ischemic injuries. However, it remains uncertain how well thiopental reduces cerebral metabolism. We investigated its effectiveness by comparing outcomes after different doses. METHODS: Fifty patients who underwent aortic arch repair with hypothermic circulatory arrest had their records reviewed. Electroencephalograms (EEG) and partial pressures of oxygen in the internal jugular vein (PjO2) were monitored. Following confirmation of total disappearance of EEG activity, 15 or 30 mg/kg thiopental was administered before circulatory arrest Th duration of circulatory arrest ranged from 16 to 77 min. RESULTS: Hospital mortality rate was 10% and 4 (8%) patients developed neu-rologic complications, but 3 of them were transient. After thiopental infusion, PjO2 increased significantly from 430 to 499mmHg (p <0.01), indicating that thiopental reduces cerebral oxygen consumption. The rate of the decrease in PjO2 during circulatory arrest was slower with the higher thiopental dose, suggesting that thiopental lowered the cerebral metabolic rate of oxygen during circulatory arrest. CONCLUSION: It appears that thiopental has protective effects against cerebral ischemia under profound hypothermia. PMID- 10522792 TI - Valve-preserving treatment of Ebstein's anomaly: perioperative and follow-up results. AB - BACKGROUND: Ebstein's anomaly is a rare congenital cardiac defect of the tricuspid valve (TV) leading to severe tricuspid insufficiency (TI). METHODS: In ten patients, 6 adults (39-53 years) and 4 children (5-10 years), operated between 1989 and 1995 echocardiography was performed pre and post repair and at follow-up. Patients were assessed in our institution at two cut-off points, resulting in a mean first follow-up of 17 +/-15 months and a mean second follow up of 53+/-23 months. All patients had additional congenital cardiac defects (ASD,VSD). In all patients the TV was repaired by techniques described by Carpentier et al. with some modifications. The goal of this reparative attempt is to mobilize restricted leaflet tissue and aid coaptation through implantation of a ring. RESULTS: Echocardiographically we were able to identify significant characteristics for the successful repair of Ebstein's anomaly. The severity of the disease is represented by the size and function of the right ventricle and the atrialized chamber, the most advanced cases exhibiting a dilated right ventricle with poor contractility. There was severe preoperative TI ( mean grade 3.2 +/- 0.3). Postoperatively TI was significantly reduced to a mean grade of 2+/ 0.2. 60% of the patients demonstrated an improvement in the ratio of atrialized chamber to functional right ventricle. Right-ventricular function was improved, the mean score being 2.8+/-0.1. At follow-up I and II right-ventricular function and tricuspid insufficiency was improved in most patients and all patients benefited in quality of life. CONCLUSION: These results suggest that surgical correction should not be delayed until severe right heart failure develops as, particularly in children, good results are achieved, improving the quality of life. PMID- 10522793 TI - Radicality and prognosis of surgical treatment of thoracal carcinoid tumors: a review of 152 operated cases. AB - BACKGROUND: Experience of thoracal (bronchial and thymic) carcinoid tumors is discussed to add some remarkable diagnostic and therapeutic details for their treatment, based on a retrospective clinico-pathological analysis of 152 consecutive patients operated on at the Thoracic Surgery unit in Budapest between 1974 and 1988. METHODS: Prior to surgery 70 patients were symptom free, obstructive symptoms dominated in 65 patients, and hemoptysis occurred in 23 cases. In 68 patients a peripheral coin lesion was visible in radiographs and in 81 cases the tumor could be seen by bronchoscopy. In 3 patients the neoplasm appeared as mediastinal thymic-carcinoid. Pathological confirmation was based on routine light-microscopic sections, Grimelius technique, and immunohistochemical stainings for NSE and chromogranin. Bronchoplastic procedures were performed in 28 patients and limited (wedge or segmental) resection in 21 cases. RESULTS: Atypical carcinoids were diagnosed in 18 cases, microscopic vascular invasion could be seen in 70 tumors (46%), and 12 patients had a single hilar lymph-node metastasis. Immunostaining for NSE was evident in all carcinoids and 82% of the tumors presented positive reaction for chromogranin staining. Hospital mortality was 1.3%. The 5-year-survival rate amounted to 93% and the 10-year-survival rate to 83% (126/ 152). The early postoperative deaths were among 49 patients operated on by parenchyma-sparing methods; the rest of these 49 are alive and free of symptoms. Local recurrence occurred after a lobectomy and following removal of a mediastinal carcinoid. The tumors of 23 of the 26 dead patients showed vascular invasion, but 19 neoplasms among them had neither atypia nor regional lymph-node involvement. In the group of patients having tumors with signs of microscopic vascular invasion the 10-year-survival rate was 67%, while in the others it amounted to 96%. CONCLUSIONS: Bronchial carcinoids require anatomic resection with lymph-node dissection. On the other hand, however, parenchyma-sparing methods have to be encouraged because of excellent late results. In our experience, immunohistochemistry for chromogranin can give some help in separation on the neuroendocrine tumor scale, and the presence of microscopic vascular invasivity is the main prognostic factor. PMID- 10522794 TI - Influence of several solutions used in bypass surgery on the permeability of the endothelium of carotid arteries in New Zealand rabbits. AB - BACKGROUND: Alteration of endothelial permeability by perfusion solutions used may influence the outcome of bypass grafts. METHOD: Carotid arteries of New Zealand rabbits were locally perfused in situ for 20 or 60 min with various solutions used in bypass surgery. After restoring normal circulation, horseradish peroxidase was injected in the ear vein. Endothelial permeability was measured by electronmicroscopy as the peroxidase accumulation in the subendothelial space during 6min circulation. RESULTS: The density indices (mean standard deviation) as a parameter for permeability in comparison to the control vessels were significantly greater than 100% for all solutions: for physiological saline 254+/ 22% and 358+/-15%, for Ringer's lactate 206+/-26% and 302+/-17%, for St. Thomas' Hospital solution 163+/-15 % and 252+/-29%, and for Bretschneider's HTK solution 130+/-15% (p=0.003) and 169+/-26%, after 20 and 60 min perfusion. Addition of heparin (50IU/ml) caused a significant increase in endothelial permeability (p<0.05). CONCLUSIONS: Bretschneider's is the most suitable of the solutions studied as a graft storage medium in bypass and cardiothoracic surgery, but a solution causing even less damage is desireable. PMID- 10522795 TI - Time-dependent efficacy of initial reperfusion with 2,3 butanedione monoxime (BDM) on release of cytosolic enzymes and ultrastructural damage in isolated hearts. AB - BACKGROUND: Reperfusion injury after cardioplegia may not be sufficiently addressed by conventional cardioplegic techniques in open heart surgery. 2,3 butanedione monoxime (BDM) has the potential to reduce myocardial reperfusion injury by uncoupling myocyte contraction from the intracellular calcium concentration, thus reducing reperfusion contracture. The aim of this study was to investigate the effects of different application periods of BDM during initial reperfusion on myocardial tissue injury after cardioplegia. METHODS: Isolated guinea-pig hearts underwent 50 min of cardioplegic arrest in St. Thomas' Hospital II solution at 37 C. Control hearts (n = 8) were immediately reperfused with normal Krebs-Henseleit solution for 30 min. In the therapy groups BDM-5, BDM-20, and BDM-40 (n = 8, each), hearts were initially reperfused with BDM (20mmol/L) for either 5, 20, or 40 min, respectively, followed by 30 min of reperfusion with normal Krebs-Henseleit solution. Coronary venous effluent was collected to estimate myocardial tissue damage through release of cytosolic enzymes (LDH and CK) and cardiac troponin 1. Ultrastructural alterations were qualitatively assessed by electron microscopy. RESULTS: Initial reperfusion with BDM markedly reduced LDH and CK release, as long as BDM was present. After washout of the protective agent a rebound of enzyme release occurred in BDM-5 hearts which was effectively reduced in BDM-20 and BDM-40 hearts. Troponin I release was similarly increased in all groups at the onset of reperfusion and rapidly decreased thereafter. Myocardial ultrastructural damage was most pronounced in control hearts, intermediate in BDM-5 and BDM-40 hearts, but markedly attenuated in BDM 20 hearts. CONCLUSIONS: Both 20 and 40 min of initial reperfusion effectively protected the hearts from reperfusion damage as indicated by cytosolic enzyme release, while 5 min of treatment were clearly insufficient. Toxic effects of BDM during the longer treatment period of 40 min or induction of edema by the long term perfusion of non-beating hearts in this group may account for the worse preservation of myocardial ultrastructure in BDM-40 hearts. Thus, contraction uncoupling during initial reperfusion by BDM or similarly acting drugs may prove a viable principle for reduction of myocardial reperfusion injury. However, the ideal duration of treatment for the best therapeutic effect must be carefully evaluated. PMID- 10522796 TI - Extracardiac conduit replacement with autologous tissue only, via a left anterior small thoracotomy. AB - A 13-year-old boy, who had undergone a Rastelli procedure 8 years before, developed severe extracardiac conduit stenosis and infectious endocarditis. Sternal re-entry appeared to be dangerous and to require extensive adhesiolysis. Through a left anterior small thoracotomy, however, good exposure was obtained and the operation was performed safely and much less invasively. The main pulmonary artery was reconstructed with autologous tissues only, i.e. fibrous tissue bed and broadly based pericardial pedicle. There was no residual stenosis. The patient convalesced rapidly. PMID- 10522797 TI - Accessory mitral valve tissue causing severe subaortic stenosis with dextrocardia in a premature newborn. AB - We report an unusual case of left-ventricular outflow obstruction caused by accessory mitral valve tissue associated with dextrocardia and ventricular septal defect in a seven-day-old, 2200 grams premature infant, who was referred with a heart murmur. The diagnosis was made by two-dimensional and Doppler echocardiography which demonstrated the accessory tissue as well as a 100 mmHg peak systolic gradient between the left ventricle and the aorta. Ten days after the presentation the infant underwent emergency surgery after respiratory arrest and recurrent episodes of syncope. The accessory mitral valve tissue and its fibrous extension were excised and the ventricular septal defect was closed. We believe that surgical treatment of patients with accessory mitral valve tissue should be performed early because of the possibility of acute deterioration. Combined aortotomy and interatrial approach is very helpful in evaluating the anatomy and the mitral valve function as well as delineating the tissue to be excised. PMID- 10522798 TI - Value of video-assisted thoracic surgery in traumatic extrapleural hematoma. AB - The current article reports on a rare case recently experienced, in which a medially displaced extrapleural fat layer with parietal pleura, revealed by CT scan of the chest, was a sign of traumatic extrapleural hematoma. Video-assisted thoracic surgery was not suited to approaching and managing the extrapleural hematoma, so that a limited thoracotomy was mandatory. Extrapleural hematoma should be considered a relative major contraindication to video-assisted thoracic surgery. PMID- 10522799 TI - Successful removal of a primitive neuroectodermal tumor in the lung with gross extension into the left atrium. AB - We report here the successful multi-model treatment of a 31-year-old female demonstrating a primitive neuroectodermal tumor originating in the lower lobe of the right lung with gross extension into the left atrium via the inferior pulmonary vein. The tumor was markedly reduced by combination chemotherapy consisting of vincristine, doxorubicin, cyclophosphamide, and ifosfamide. The residual tumor was completely removed through a combined left atrial resection and right middle and lower lobectomy, using a percutaneous cardiopulmonary support system. PMID- 10522800 TI - Cardiac surgery in Germany during 1998. A report by the German Society for Thoracic and Cardiovascular Surgery. PMID- 10522801 TI - Co-regulation of mucosal nitric oxide and prostaglandin in gastric adaptive cytoprotection. AB - OBJECTIVE: The correlation between mucosal generation of nitric oxide (NO) and prostaglandin E2 (PGE2) in gastric adaptive cytoprotection was investigated. MATERIALS AND TREATMENT: Male Sprague-Dawley rats were pretreated with either N(w)-nitro-L-arginine methyl ester (L-NAME, 12.5 mg/kg i.v.) or indomethacin (5 mg/kg s.c.). Following that, mild irritant 20% ethanol was administered, 15 min prior to 100% ethanol challenge. METHODS: Macroscopic gastric mucosal damage, NO synthase activity, mucosal PGE2 and leukotriene C4 (LTC4) levels were measured. RESULTS: Administration of L-NAME and indomethacin significantly reduced the protective action of 20% ethanol against 100% ethanol-induced gastric mucosal damage. Besides, mucosal activity of constitutive NO (cNO) synthase, but not of the inducible isozyme (iNO synthase), was elevated following 20% ethanol treatment. This was accompanied by a reduction in mucosal leukotriene C4 level. Indomethacin significantly inhibited mucosal PGE2 biosynthesis but increased cNO synthase activity. Nevertheless, L-NAME reduced both cNO and iNO formation and prevented the increase in cNO formation caused by 20% ethanol, while enhancing mucosal PGE2 production. Combined L-NAME and indomethacin treatment markedly potentiated ethanol-induced mucosal damage, and completely prevented the increase in cNO or PGE2 biosynthesis when either compound was given alone. CONCLUSIONS: These findings suggest a co-regulatory relationship between mucosal NO and PG in the gastric defense system, which will be released after activation by the mild irritants to induce cytoprotection. PMID- 10522802 TI - Inhibition of neutrophil derived lysosomal enzymes and reactive oxygen species by a novel tetrapeptide. AB - OBJECTIVE AND DESIGN: The role of a tetrapeptide derivative PEP 1261 {Boc Lys(Boc)-Arg-Asp-Ser(tBu)-OtBu}, corresponding to residues 39-42 of human lactoferrin, has been tested in vitro in the modulation of neutrophil function. MATERIAL AND SUBJECTS: The level of non-enzymatic mediators of inflammation such as reactive oxygen species (ROS), enzymatic mediators such as myeloperoxidase (MPO) and lysosomal enzymes have been assessed in the presence or absence of PEP 1261 in phorbol 12-myristate 13 acetate (PMA) stimulated human neutrophils (n = 6) and also in neutrophils isolated from adjuvant induced arthritic rats (AIA) (n = 4). TREATMENT: PEP 1261, at a concentration of 0.14 mM, was added to the neutrophil cultures. STATISTICAL METHOD: The results were analysed by nonparametric statistics using Mann Whitney U test. RESULTS: Addition of PEP 1261 effectively blocked the H2O2 and O2*- release, decreased the levels of MPO levels (p< 0.01) and lysosomal enzymes (p < 0.05) as compared to PMA stimulated human neutrophils. PEP 1261 was also observed to inhibit the levels of H2O2, O2*-, MPO and lysosomal enzymes (p < 0.05) as compared to PMA stimulated control rat neutrophils and neutrophils from arthritic rats. CONCLUSIONS: The results of this study indicate that PEP 1261 could serve as an excellent antiinflammatory agent. PMID- 10522803 TI - Periarthritis promotes gait disturbance in zymosan-induced arthritis in rats. AB - OBJECTIVE AND DESIGN: We studied the contribution of periarthritis and synovitis to gait disturbance in zymosan (Zy)-induced arthritis. METHODS: Sixty Wistar rats were subjected to injection of Zy (1 mg) into their right knee joints. A first group of animals (GI) had Zy injected through the intact skin. A second group (GII) had Zy injected directly into the articular cavity after excision of the skin and subcutaneous tissue surrounding the joint. Gait disturbance was evaluated using the rat-knee joint incapacitation test. Increase in vascular permeability and cell influx were assessed in joint fluids and joint histology was performed. RESULTS: Zy injection induced a dose-dependent gait disturbance which was maximal at the third/fourth hour of arthritis, being significantly greater in GI rats, whereas cell influx (neutrophils > or = 80%) was maximal at the sixth hour. Cell influx and increase in vasopermeability did not differ between both groups. Histology revealed no significant difference between GI and GII. A third group (GIII), subjected to immune-complex arthritis, that received anti-bovine serum albumin (BSA) antibodies intra-articularly and BSA i.v., did not present gait disturbance, despite the increase in cell counts. CONCLUSIONS: Vascular permeability increase and cell influx are phenomena independent of gait disturbance. Neutrophils do not seem to contribute to development of gait disturbance in Zy arthritis. Sensitization of specific pain receptors in periarticular rather than in synovial tissue is responsible for gait disturbance in Zy-induced arthritis. PMID- 10522804 TI - Periarticular tissue levels of cytokine- and endothelin-1-like immunoreactivity during the course of antigen-induced arthritis in the rat. AB - OBJECTIVE AND DESIGN: To correlate the changes in periarticular tissue levels of pro-inflammatory cytokines and endothelin-1 (ET-1) to local pathophysiology in rats with antigen-induced arthritis (AIA). MATERIALS: The periarticular soft tissue was excised from sixty-six rats at different stages of AIA. METHODS: The samples were homogenized and the levels of immune like (LI) reactivities were determined. The levels in the arthritic joints were compared to those in non arthritic tissue. Statistical significance was determined by ANOVA followed by Fisher PLSD. RESULTS: Compatible with a role in an early alarm reaction some mediators (tumor necrosis factor alpha-LI, interleukin-2-LI and interferon-gamma LI) were elevated prior to the vascular inflammatory activity. The curve for ET-1 LI closely followed the intensity of the inflammation whereas these parameters preceeded the elevation of interleukin-1beta-LI. CONCLUSIONS: Transient waves of different mediators appear during the course of the arthritis. This indicates the presence of active downregulatory mechanisms. Here the model could differ from human rheumatoid arthritis. PMID- 10522805 TI - The beta-adrenoceptor agonist clenbuterol is a potent inhibitor of the LPS induced production of TNF-alpha and IL-6 in vitro and in vivo. AB - OBJECTIVE AND DESIGN: To investigate the suppressive effects of the beta-agonist clenbuterol on the release of TNF-alpha and IL-6 in a lipopolysaccharide (LPS) model of inflammation, both in vitro and in vivo. MATERIAL AND SUBJECTS: Human U 937 cell line (monocyte-derived macrophages), and male Wistar rats (200-250 g). TREATMENT: U-937 macrophages were incubated with LPS at 1 microg/ml, with or without 1.0 mM-0.1 nM test drugs (clenbuterol and other cAMP elevating agents) for 1-24 h. Rats were administered either 1 or 10 microg/kg clenbuterol (or saline) orally, 1 h before intraperitoneal administration of 2 mg/kg LPS. METHODS AND RESULTS: TNF-alpha and IL-6 time-concentration profiles were determined both in culture media and plasma, using ELISA' s and bioassays. LPS-mediated release of both cytokines was significantly suppressed by clenbuterol. CONCLUSIONS: The beta-agonist clenbuterol very potently suppresses the LPS-induced release of the pro-inflammatory cytokines TNF-alpha and IL-6 both in vitro and in vivo. PMID- 10522806 TI - Cyclooxygenase-2-dependent generation of 8-epiprostaglandin F2alpha by lipopolysaccharide-activated J774 macrophages. AB - OBJECTIVE: The generation of 8-epiprostaglandin F2alpha (8-epi-PGF2alpha) by arachidonic acid (AA)- and lipopolysaccharide (LPS)- stimulated J774 macrophages has been investigated. MATERIAL: Murine monocyte/macrophage J774 cell line. METHODS: Cells were incubated with AA or LPS and the amount of 8-epi-PGF2alpha, 6 ketoprostaglandin F1alpha (6-keto-PGF1alpha) and prostaglandin E2 (PGE2) released in the incubation media measured by radioimmunoassay (RIA) or, in some experiments, by enzyme immunoassay (EIA). The effect of dexamethasone (DXM), cycloheximide (CXM) and 5,5 dimethyl-3-(3-fluorophenyl)-4-(4 methylsulfonyl)phenyl-2(5H)-furanone (DFU), a cyclooxygenase-2 (COX-2) selective inhibitor, on LPS-induced generation of AA metabolites was assessed. RESULTS: AA induced a significant production of 6-ketoPGF1alpha and PGE2, whereas LPS caused a concentration- and time-dependent increase of 8-epi-PGF2alpha, 6-keto-PGF1alpha and PGE2. DXM (2 microM) as well as CXM (1 microM) significantly decreased (p<0.001; n = 4) the LPS-stimulated production of 8-epi-PGF2alpha (by 86% and 82%, respectively), 6-ketoPGF1alpha (by 78% and 74%, respectively) and PGE2 (by 83% and 78%, respectively). Immunostimulated production of AA metabolites was also inhibited by DFU (IC50 0.3+/-0.04 microM; 0.16 +/- 0.02 microM and 0.11 +/- 0.05 microM for 8-epi-PGF2alpha, 6-keto-PGF1alpha and PGE2, respectively. CONCLUSIONS: These results demonstrate the role of COX-2 in the generation of 8 epi-PGF2alpha by LPS-stimulated J774 macrophages. The relevance of these findings requires further elucidation. PMID- 10522807 TI - Autoimmunity and endocrinology. AB - The etiology and pathogenesis of autoimmune endocrinopathies are now much clearer as a result of advances in our understanding of basic immunology, and particularly the development of novel animal models. Also crucial has been the molecular characterisation of target autoantigens, although this still remains elusive for some disorders, such as hypoparathyroidism and premature ovarian failure, retarding progress. The application of new genetic techniques, the detailed structural analysis of autoantigen epitopes and the creation of new experimental animal disorders (by DNA immunisation or knockouts) will ensure still more rapid progress into the new millennium. PMID- 10522808 TI - Environmental factors in autoimmune thyroid disease. PMID- 10522809 TI - Pathogenic and clinical aspects of Graves' disease. PMID- 10522810 TI - Pathogenetic and clinical aspects of endocrine ophthalmopathy. PMID- 10522811 TI - Pathogenetic and clinical aspects of autoimmune thyroiditis. PMID- 10522812 TI - Prophylactic levothyroxine therapy in patients with Hashimoto's thyroiditis. PMID- 10522813 TI - Pathophysiology of type 1 diabetes mellitus. PMID- 10522814 TI - Genetic susceptibility to type 1 diabetes: clinical and molecular heterogeneity of IDDM1 and IDDM12 in a german population. AB - Type 1 Diabetes mellitus (IDDM) results from an immune-mediated destruction of the pancreatic b-cells. The genetic predisposition is mainly confered by variations within MHC class II region on chromosome 6p as well as the CTLA4 gene located on chromosome 2q33. We analysed the transmission of HLA DQA1, DQB1, DRB1*04 alleles as well as an endogenous retroviral element (DQLTR3) in 130 families with a type 1 diabetic offspring in order to evaluate their role in genetic susceptibility to IDDM. Also the combined transmission of HLA and CTLA4 haplotypes was investigated. MHC class II alleles were typed using sequence specific primer analysis. The presence or absence of DQLTR3 was defined by a nested PCR approach and CTLA4 microsatellite polymorphisms were detected with fluorescence-labeled primer on an automated sequencing system. By transmission distortion test we confirm the linkage of HLA DQA1*0501 DQB1*0201 (DR3 DQ2) as well as DQA1*0301 DQB1*0302 (DR4 DQ8) with IDDM. Whereas the combination with CTLA4 risk markers leads to the highest transmission rate on DR3 positive haplotypes, the predisposing CTLA4 variant does not modulate the risk on DR4 haplotypes. However, the absence of DQLTR3 on DR3, but its presence on DR4 haplotypes significantly increases the genetic risk for type 1 diabetes. Therefore predisposing MHC class II haplotypes are defined by distinct loci which differentially control genetic susceptibility. The combined transmission of protective CTLA4 and HLA DR3 as well as DR4 haplotypes confirms the dominant role of HLA class II polymorphisms in defining disease susceptibility to type 1 diabetes mellitus. PMID- 10522815 TI - Environmental factors in the pathogenesis of type 1 diabetes mellitus. AB - Type 1 diabetes mellitus is perceived as a chronic autoimmune disease with a subclinical prodrome characterized by selective loss of insulin producing beta cells in the pancreatic islets in genetically predisposed subjects. Less than 10% of those with increased genetic susceptibility progress to clinical disease suggesting a strong environmental modification of the prediabetic process. Various exogenous triggers, such as certain dietary factors and viruses, are thought to induce the autoimmune process leading in some individuals to extensive beta-cell destruction and ultimately to the clinical manifestation of type 1 diabetes. In addition to their role as triggers, environmental factors are also likely affecting the outcome of the process and the rate of progression to clinical disease in those who do develop Type 1 diabetes. The present review focuses on relatively recent data on environmental factors potentially involved in the pathogenesis of Type 1 diabetes with an emphasis on dietary factors, and cow's milk (CM) proteins in particular. The CM hypothesis has remained controversial for a decade, and therefore an intervention trial should be performed to settle the issue. Recent prospective studies have indicated that enterovirus infections may induce beta-cell autoimmunity and potentiate the humoral immune response to beta-cell antigens in subjects with an ongoing process. There are also very preliminary data suggesting a similar role for rotavirus infections. Although there may be a single trigger of beta-cell autoimmunity in a given individual, it is highly unlikely that there is only one exogenous determinant of Type 1 diabetes. Rather we have a complicated interaction between a series of environmental factors and between environmental factors and genetic disease predisposition resulting in progression to clinical Type 1 diabetes in those genetically susceptible individuals who experience an unfortunate timing and/or clustering of diabetogenic exogenous culprits and/or a lack of protective environmental modifiers. Ongoing prospective studies starting from birth provide an optimal setting for the identification of environmental factors affecting the risk of progression to clinical Type 1 diabetes. PMID- 10522816 TI - Prevention studies in type 1 diabetes. PMID- 10522817 TI - Macrophage and lymphocyte homing in experimental diabetes. AB - Diabetes mellitus was induced in C57BL/6 mice by multiple low doses of streptozotozin (STZ). By transferring lymphocytes from these diabetic animals to healthy recipient mice insulitis can be induced in healthy recipient mice. On day 21 after the start of STZ-treatment splenic lymphocytes were separated in vitro and stained with the fluorochrom acridine red for adoptive transfer. The recipient mouse had been pretreated with a subdiabetogenic dose of STZ (5 mg/kg) i.p. 24 h prior to the lymphocyte transfer. With in vivo microscopy we measured the lymphocyte adherence to the endothelium of islets of the recipient mouse. After the administration of mAbs directed against LFA-1, ICAM-1, murine VCAM-1, VLA-4, MadCAM or alpha4,beta7-integrin prior to the cell transfer we could demonstrate a significant decrease of donor lymphocyte adherence in islets (P<0.01). The pretreatment of the recipient mice with STZ 24 h before the transfer of lymphocytes might attract macrophages to the islets. Therefore we pretreated the recipients with antibodies to cytokines or silica. mAb IFN-gamma, pAb TNF-alpha, pAb IL-1alpha or silica reduced lymphocyte adherence to islet endothelium significantly (P<0.01). The presented results support the following interpretation: The pretreatment of the recipient mice with the islet specific toxin STZ in subdiabetogenic doses attracts macrophages to the islets. The macrophages release cytokines leading to an increased expression of adhesion molecules on the endothelium of the islets. The consequence is lymphocyte homing selectively to the islets. The cascade of lymphocyte homing is complex and various pairs of adhesion molecules are involved. The discussion on the importance of the single pairs of adhesion molecules is irrelevant, since the cascade of lymphocyte homing can be disturbed by the application of a mAb to any one adhesion molecule involved in the cascade of lymphocyte homing. PMID- 10522818 TI - New developments in the treatment of type 1 diabetes mellitus. AB - Treatment of type 1 diabetes mellitus has made tremendous advances within the last decades. With concern to insulin delivery there are two promising new approaches. One is the intrapulmonary insulin delivery which has become feasible by the development of new inhalation devices which provide a sufficient degree of intrapulmonary drug retention. Also oral insulin delivery seems feasible when surface active substances are used to cross the mucosal membrane in the gut. Clinical research has also focussed on coatings for the insulin molecules to solve the problem raised by the proteolytic activity of the digestive system. A very new agent produced by a fungus called Pseudomassaria has been demonstrated to reverse the clinical signs of diabetes mellitus in mice. The compound diffuses through the cell membrane, binds to the inner part of the insulin receptor and activates the insulin typical biological effects. Nowadays a variety of insulin analogs are designed and tested for their clinical use. By shifting the isoelectric point towards to a slightly acidic pH, HOE 901 precipitates at physiologic pH resulting in a constant and peakless insulin delivery. NN 304 is a 14-carbon aliphatic fatty acid acylated analog that binds to serum albumin resulting in a flatter time-action profile than NPH insulin. Also rapid acting insulin analogs are or will be launched in the near future aiming to ensure an improved postprandial glucose regulation. Glucagon-like peptide-1 (GLP-1) improves metabolic control by a variety of effects, e.g. the enhancement of insulin secretion and inhibition of glucagon secretion. Moreover, GLP-1 reduces food and water intake controlled by the brain, and inhibits gastric emptying. A disadvantage of GLP-1 is its very short half-life. Novel derivatives with the beneficial effects of GLP-1 but a better resistance against degradation have been designed. In addition substances have been developed inhibiting GLP-1 degradation or augmenting GLP-1 release from its abundant endogenous pool. Finally, there is a variety of interesting approaches aiming to improve or ease blood glucose self monitoring. One is the development of subcutaneous catheters for continuous blood glucose control. In another system reverse iontophoresis is used for sampling interstitial fluid which reflects capillary blood glucose levels. Instead of using an electric current, a brandnew system creates micropores in the skin by a laser ablation system. Through these micropores a specific device performs a mild suction to obtain intersitial fluid. Further systems which measure blood glucose by near infrared spectroscopy are still investigated in order to improve their technical function and to reduce their weight. This article intends to give an overview over the new developments in the treatment and management of type-1 diabetes mellitus. PMID- 10522819 TI - A new method for simultaneous demonstration of anterograde and retrograde connections in the brain: co-injections of biotinylated dextran amine and the beta subunit of cholera toxin. AB - In studying reciprocally connected brain networks, it is advantageous to use techniques that allow simultaneous visualization of both efferent and afferent connections from a single injection site. We report on a new technique to achieve this using pressure injections of a mixture of biotinylated dextran amine (BDA) and the beta subunit of cholera toxin (Ctb). Adult male hamsters (n = 12) received 20-30-nl injections of either a 1:1 mixture of BDA (Sigma, 10%) and Ctb (List Biological, 0.5%), or each tracer by itself, into the medial amygdala. Adult female sheep (n = 4) received 200-300 nl of the combined tracer into the A15 region of the hypothalamus. After 1 (hamster) or 2 weeks' (sheep) survival, animals were perfused with 4% paraformaldehyde. Sections were double-labeled, first for BDA histochemistry using nickel-enhanced DAB, then for Ctb using a PAP technique and unenhanced DAB. In all animals, combined injections resulted in clear and consistent patterns of both anterograde and retrograde labeling. Ctb immunoreactivity was distinct and easily distinguished from BDA labeling. There was no evidence for loss of sensitivity of either tracer due to the combined delivery; no differences were seen between combined or single tracer injections in numbers of retrogradely-labeled cells or in the distribution of anterogradely labeled fibers. In summary, the combined delivery of BDA and Ctb is an easy and reliable technique for simultaneous afferent and efferent tract tracing in both small and large animals; it could potentially be combined with immunocytochemistry to determine the neurochemical content of labeled cells or fibers. PMID- 10522820 TI - Intraparenchymal infusions of 192 IgG-saporin: development of a method for selective and discrete lesioning of cholinergic basal forebrain nuclei. AB - The immunotoxin 192 IgG-saporin has a high degree of selectivity for cholinergic neurons within the basal forebrain (CBF). Intracerebroventricular delivery of 192 IgG-saporin results in a diffuse and massive depletion of choline acetyltransferase (ChAT) activity in projections of the CBF, and non-selective loss of Purkinje cells. To dissociate the basal-cortical and septo-hippocampal cholinergic systems and to minimize non-specific effects, we developed intraparenchymal parameters to deliver 192 IgG-saporin discretely to either the nucleus basalis magnocellularis (NBM) or the medial septum (MS). Intraparenchymal administration of the immunotoxin into the NBM or MS resulted in a dose-dependent depletion of ChAT activity in the corresponding projection areas and a concomitant loss of ChAT immunoreactive neurons in both nuclei. Both lesions were regionally restricted, having a minimal diffusion into adjacent CBF nuclei. Control infusions did not result in non-specific parenchymal damage. In addition, immunotoxic infusions had no effect on monoamine neurotransmitter systems. By optimizing the dosages for both CBF nuclei, we maximized ChAT depletion while minimizing diffusion into the adjacent CBF nuclei. This study delineated injection parameters enabling a selective dissociation of two cholinergic subpopulations in the basal forebrain for further functional characterization. PMID- 10522821 TI - A 100-channel system for real time detection and storage of extracellular spike waveforms. AB - As extracellular electrode arrays with 100 or more active recording sites become more widely used for simultaneous recording of neural ensembles, practical data acquisition systems that can efficiently accommodate high electrode counts are needed. To reduce the high data rates associated with extracellular recordings from these arrays, various algorithms and systems have been designed to provide complete online detection and classification of extracellular spike waveforms. However, many of these algorithms require significant user supervision to ensure accurate performance. In this paper, we discuss the design and validation of a 100-channel PC-based system that can be used with arrays of extracellular electrodes such as the Utah Electrode Array. Instead of comprehensive online spike analysis, the system performs online detection and storage of the spike waveforms for offline classification. This strategy preserves the data of interest, reduces system complexity, and requires less user supervision during experiments. PMID- 10522822 TI - Validation of a new antiserum directed towards the synthetic c-terminus of the FOS protein in avian species: immunological, physiological and behavioral evidence. AB - In the past 10 years, the study of the expression of immediate early genes, such as c-fos, in the brain has become a common method for the identification of brain areas involved in the regulation of specific physiological and behavioral functions. The use of this method in avian species has been limited by the paucity of suitable antibodies that cross-react with the FOS protein in birds. We describe in this paper the preparation of an antibody directed against a synthetic fragment of the protein product of the c-fos gene in chickens (Gallus domesticus). We demonstrate that this new antibody can be used in several avian species to study FOS expression induced by a variety of pharmacological, physiological and behavioral stimuli. Western blot studies indicated that this antibody recognizes a protein of the expected size (47 kDa) but also cross reacts to some extent with proteins of lower molecular weight that share sequence homology with FOS (Fos-related antigens). FOS immunocytochemistry was performed with this antibody in four species of birds in three different laboratories utilizing diverse variants of the immunocytochemical procedure. In all cases the antibody provided a reliable identification of the FOS antigen. The new antibody described here appears to be suitable for the study of FOS expression in several different avian species and situations. It is available in substantial amounts and will therefore make it possible to use FOS expression as a tool to map brain activity in birds as has now been done for several years in mammalian species. PMID- 10522823 TI - A new Alamar Blue viability assay to rapidly quantify oligodendrocyte death. AB - We developed a rapid fluorometric viability assay for primary cultures of OL precursors (preOLs) or mature OLs that utilized the oxidation/reduction indicator dye Alamar Blue (AB). PreOLs had a lower rate of AB reduction than did mature OLs (0.02 +/- 0.01 units/min per cell versus 0.07 +/- 0.01). The assay was tested under two conditions toxic to preOLs: oxidative stress induced by glutathione depletion or kainate excitotoxicity. When glutathione was depleted by a 24-h exposure to cystine-depleted medium, the EC50 values for the dependence upon cystine for survival did not differ significantly when determined by AB reduction (2 +/- 2 microM), by the trypan blue exclusion method (3 +/- 3 microM) or by MTT histochemistry (1 +/- 0.4 microM). Quantification of preOL viability with AB was unaffected by the presence of free radical scavengers (alpha-tocopherol or idebenone) or the antioxidant enzymes Cu,Zn-superoxide dismutase and catalase. There was no difference in preOL viability as determined by AB or MTT after a 24 h exposure to kainate at concentrations up to 1 mM. AB offers a rapid objective measure of OL viability in primary culture and is a valid means to quantify OL death. PMID- 10522824 TI - Development and characterization of an adult model of obstructive hydrocephalus. AB - While hydrocephalus is common in adults its pathophysiology is not fully understood and its treatment remains problematic. Previous animal models have been acute, developmental, or involved non-specific blockage or inflammation and are not suitable for study of chronic adult-onset hydrocephalus. In this study, we describe the development of a canine model which allows basic physiological studies along with diagnostic and treatment procedures via surgical occlusion of the fourth ventricle with a bolus injection of cyanoacrylic gel glue. A total of 26 adult male canine mongrels were used for the induction of chronic hydrocephalus and were monitored for 1-12 weeks post-induction using magnetic resonance imaging (MRI), intracranial pressure measurements, and neurological fitness assessments. Of these, 81% (21/26) developed hydrocephalus that was mild (N = 6), moderate (N = 7), or severe (N = 8). Pressures were mild and transiently elevated, and brain compliance decreased. Clinical symptoms were also mild and transient. This model is unique in its focal obstruction without local compression or general inflammation and should facilitate the study of the pathophysiology and treatment of chronic adult-onset hydrocephalus. PMID- 10522825 TI - Reversible mechanical fixation of eye position in awake head-restrained pigeons (Columba livia). AB - Here we describe a method to fix gaze positions and to significantly reduce saccadic oscillations in pigeons. The procedure consists of a mechanical immobilization of the eye through the use of an electromagnet that exerts a radial force upon a small metal rectangle glued to the dorsal part of the eye. The method can be used in avian visual neurophysiology in order to hold the eye immobilized for periods of time, long enough to map the properties of visual receptive fields and investigate the possible functions of saccadic oscillations. PMID- 10522827 TI - A statistical method for correcting distortions of amplitude distribution histograms due to collisions of synaptic events. AB - The waveform of spontaneous synaptic potentials or currents comprising synaptic noise can be significantly distorted when these events are closely spaced due to a high frequency activity in the presynaptic inputs that generate them. It is essential to correct these alterations prior to measurements of overlapping miniature and/or postsynaptic potentials, in order to provide reliable information about their true amplitude distributions, and to avoid spurious peaks in the resulting histograms. In this paper we describe a statistical method for making these corrections, its range of application, and its theoretical background. Its use becomes necessary when the frequency of events is of the order of 8-50 Hz, depending upon their time to peak, which ranges from 6 to 1 ms in most synaptic potentials recorded in the central nervous system. PMID- 10522826 TI - Recording of intracellular Ca2+, Cl-, pH and membrane potential in cultured astrocytes using a fluorescence plate reader. AB - A multi-well fluorescence plate reader was used to determine changes in intracellular ionic concentrations and changes in transmembrane voltage in primary cortical or hippocampal astrocytes. Recordings were compared to those obtained using a traditional microscope-based imaging setup that utilizes a monochromatic light source for excitation and an intensified CCD camera for detection. Measurement of pHi with the ratiometric dye BCECF provided resolution similar to that of a microscopic approach. We also demonstrate using Fluo-3 that the measurement of glutamate-induced [Ca2+]i fluctuations are comparable to recordings on the microscope-based system when 25 cells were averaged. This is expected because the plate reader averages responses from many cells. Voltage changes induced by changes in K+(o) from 3 to 5 mM were readily resolvable with the plate reader using the potentiometric dye bis-oxynol, and overall sensitivity was similar to that of microscopically-determined voltage changes. We also found that the plate reader was capable of resolving GABA-induced Cl-(i) fluctuations using the Cl(-)-sensitive indicator MEQ. From these experiments we conclude that multi-well fluorescence plate readers can be used to effectively record changes in intracellular ion concentrations and transmembrane voltage of populations of cells affording time and amplitude resolution approaching that of conventional fluorescence imaging methods. In addition, plate reader-based fluorescence studies demonstrate the added capability to rapidly screen large numbers of samples. PMID- 10522828 TI - A chemiluminescent catecholamine assay: its application for monitoring adrenergic transmitter release. AB - A chemiluminescent procedure for measuring catecholamines (dopamine, norepinephrine, epinephrine) is described. It is based on the observation that lactoperoxidase catalyses both the oxidation of catecholamines, and the chemiluminescent reaction of luminol with their oxidation product. The assay has been adapted for continuously monitoring the release of catecholamines from adrenergic tissues, from cell suspensions and from cells loaded in culture with dopamine. PMID- 10522829 TI - New technique for vagal nerve stimulation. AB - INTRODUCTION: The vagus nerve travels in a neurovascular bundle with the carotid artery and internal jugular vein. The present study was designed to assess whether transvascular stimulation through the carotid artery of the dog can be used to directly stimulate the vagus nerve and increase parasympathetic tone. METHODS: In five anesthetized dogs, a steerable electrode catheter was positioned under fluoroscopic guidance in the right carotid artery in the mid neck via the femoral artery. Multipolar catheters were positioned transvenously through the femoral vein in the right atrium, across the tricuspid valve to record a His bundle electrogram, and in the right ventricle. RESULTS: In all five animals, vagal nerve stimulation was successfully achieved with outputs ranging between 10 and 30 mA. Sinus cycle length increased from 473 +/- 113 ms at baseline to 894 +/ 315 ms (P < 0.025) during stimulation from the right carotid artery. There was an increase in the AH interval from 55 +/- 14 to 77 +/- 23 ms (P < 0.03), a shortening of the atrial effective refractory period from 136 +/- 8 to 126 +/- 6 ms (P < 0.01), and a fall in the systolic blood pressure from 135 +/- 20 to 117 +/- 20 mmHg (P < 0.005) with stimulation from the right carotid artery. A prolongation of the AV and VA block cycle lengths and the AV nodal effective refractory period was also noted with stimulation from the right carotid artery. Atrial fibrillation was not induced at baseline in any animal. During stimulation from the right carotid artery, atrial fibrillation was induced in three of five animals and persisted for the duration of stimulation from the right carotid artery. CONCLUSION: Cardiac parasympathetic stimulation can be achieved by positioning a catheter in the neurovascular bundle in the neck adjacent to the vagus nerve with resultant effects on cardiac electrophysiology. PMID- 10522830 TI - A new video/computer method to measure the amount of overall movement in experimental animals (two-dimensional object-difference method). AB - Evaluation of the amount of overall animal movement is important for investigations of motor control mechanisms in the central nervous system. We describe a new method to quantify overall free movements of an animal without any markers using a video camera and a personal computer equipped with a video capture board. The operating principle is that the amount of overall movement of an object can be expressed by the difference in total area occupied by the object in two consecutive picture frames. The software for this application operates in real-time. Using this method and with proper setting for the cage and recording view, we can estimate three-dimensional movements of animals. The major advantages are low cost, easy operation and high sensitivity. The experimental results indicate that this method can be applied to various fields of motion analysis. PMID- 10522831 TI - Automatic detection of bursts in spike trains recorded from the thalamus of a monkey performing wrist movements. AB - In a previous paper (Churchward PR, Butler EG, Finkelstein DI, Aumann TD, Sudbury A, Horne MK. J Neurosci Methods 1997;76:203-210), we showed that a simple back propagation neural network could reliably model visual inspection by human observers in detecting the point of change of neuronal discharge patterns. The data for that study was deliberately chosen so that the point of change was readily detected and there would be high concordance between human observers. We wished to extend this investigation by comparing a variety of automatic analysis methods on more complex data sets. Two automatic analysis methods have been discussed in this paper. The knowledge based spike train analysis (KBSTA) was designed to emulate the detection of bursts by human observers. The self organizing feature map (SOFM) spike train analysis determined a burst by classifying the patterns of neuronal discharge. Neuronal discharge was recorded from the motor thalamus and nucleus ventralis posterior lateralis caudalis (VPLc) of a monkey performing consecutive trials of skilled wrist movements. Recordings were made from 36 neurons whose discharge patterns were related to wrist movement. Three hundred and sixty trials performed during the recording of these 36 neurons were chosen at random and used to compare the three methods, KBSTA, SOFM, and visual inspection. The main results of this study show that for the 360 trials the three detection methods have very similar results in detecting the onset and offset of neuronal bursts. The SOFM method is not the best first approach for detecting a burst, but it does provides independent evidence to support the KBSTA and visual inspection methods. In conclusion we propose the KBSTA method as a practical, automatic technique to identify bursts of neuronal discharge. PMID- 10522832 TI - A focusing image probe for assessing neural activity in vivo. AB - We describe a compact, focusing image probe to record rapid optical changes from neural tissue. A gradient index (GRIN) lens served as a relay lens from tissue to a microscope objective which projected an image onto a CCD camera. The microscope objective and camera assembly was adjusted independently from the GRIN lens, allowing focus changes without disturbing the probe/tissue interface; firm contact minimized movement and specular reflectance. Fiber optics around the probe perimeter provided diffuse illumination from a 780 nm laser, or 660 and 560 nm light emitting diodes. To characterize depth-of-field, we imaged a black suture through increasing tissue thicknesses. Light modulation by the suture remained detectable down to 900 microm using 780 nm illumination. We acquired images from cardiorespiratory areas of the rat dorsal medulla, at different depths and illumination wavelengths. Images illuminated at 560 nm were dominated by vasculature flow patterns, while 660 nm illumination revealed different spatial patterns which preceded vascular flow by 40 ms and may represent cardiac related neural activity. Using 780 nm light, image sequences triggered by the cardiac R-wave showed vascular perfusion changes with delayed and broader responses at deeper levels. Electrical stimulation within the vagal bundle caused fast optical changes which track the electrical response, with a different spatial distribution from hemodynamic signals. PMID- 10522833 TI - High-speed CCD imaging system for monitoring neural activity in vivo and in vitro, using a voltage-sensitive dye. AB - We have designed and constructed a high-speed CCD imaging system for optically detecting neural activity from preparations stained externally with a voltage sensitive dye, and have used this system to image evoked and epileptiform neural activity in the rat somatosensory cortex. The imaging system uses a commercially available 1/3-in. CCD chip, and it can continuously capture images for more than 8 s, at 1000 frames/s, with a spatial resolution of 128 x 62 pixels. The spatial/temporal resolution of the CCD sensor is variable by changing the geometry of on-chip binning pixels, which can be controlled by a PC/AT computer. Dye bleaching correction was not necessary for long-term imaging of epileptiform neural events, since the sensitivity of the CCD sensor was increased by combining the signal from adjacent pixels. PMID- 10522834 TI - Rapid-acting insulin secretagogues: a clinical need? AB - The history of achieving adequate insulin secretion in Type 2 diabetes has interesting parallels with Type 1 diabetes. Initial insulin secretagogue therapy involved short-acting sulphonylureas--tolbutamide being introduced 40 years ago- and early insulin therapy used unmodified (soluble) insulin. Subsequently, long acting sulphonylureas and insulin were introduced, but more recently short-acting agents have become popular again. This approach was endorsed by the European non insulin-dependent diabetes mellitus guidelines of 1989/1993. The trend at present is to match modern rapid-acting agents (such as repaglinide or the rapid-acting insulin analogues) to physiological meal-time insulin requirements in both types of diabetes. The fundamental reasons for tailoring therapy to meal-times fall into two categories: a pathophysiological rationale and a behavioural rationale. The pathophysiological rationale for tailoring treatment to mealtimes is based on the importance of restoring the mealtime insulin secretion profiles of patients with Type 2 diabetes to physiological levels to re-establish the tight control of blood glucose levels seen in healthy individuals. The behavioural rationale is derived from the observation that most people with Type 2 diabetes are overweight and would like to reduce calorie consumption, but that the risk of hypoglycaemia does not allow them to be flexible over their day-to-day calorie intake. These observations suggest that the matching of a meal-time pharmacodynamic profile to a patient's eating habits would provide a knowledge-based rationale for achieving good blood glucose control once dietary means alone are inadequate. Evidently, this can be usefully combined with other pharmacological approaches (insulin sensitizers or basal insulin). The need for combination therapy is increasingly likely as the defect in insulin secretion progresses. PMID- 10522835 TI - Cardiovascular risk factors in type 2 diabetes: the role of hyperglycaemia. AB - Macrovascular complications are the most important causes of morbidity, mortality and disability in people with Type 2 diabetes mellitus. Although other known risk factors for macrovascular disease (e.g. dyslipidaemia, hypertension, obesity) often co-exist, diabetes itself is an important risk factor for accelerated development of atherosclerosis. Hyperglycaemia, hyperinsulinaemia and insulin resistance may each play a major role in the onset and development of atherosclerotic disease, which causes arterial wall dysfunction, haematological disturbances and lipid abnormalities through two mechanisms: oxidative stress and non-enzymatic glycation. Hyperglycaemia induces damage to the endothelium through activation of mitogen-activated protein kinase, protein kinase C and transcription factor nuclear factor (NF)-kappaB and through increased levels of pro-adhesion proteins such as intracellular adhesion molecule (ICAM)-1. The arterial wall tone is shifted towards vasoconstriction by hyperglycaemia, which is also associated with vascular smooth muscle cell proliferation and increased intimal wall thickness. Alteration of the coagulation system towards thrombophilia is observed in Type 2 diabetes and a series of lipid abnormalities that facilitate the development of atherosclerosis is evident. In Type 2 diabetes, undiagnosed disease and unrecognized postprandial hyperglycaemia are becoming the most relevant issues in reducing the risk of vascular complications and cardiovascular mortality; improved glycaemic control may reduce the incidence of macrovascular complications. PMID- 10522836 TI - Evolution of beta-cell dysfunction in impaired glucose tolerance and diabetes. AB - Patients with overt Type 2 diabetes consistently have alterations in insulin secretion, including reduced insulin secretory responses to glucose, delayed and blunted meal-induced insulin secretion, increased pro-insulin and abnormal insulin secretory oscillations. More recently, it has become evident that abnormal insulin secretion antedates the onset of overt hyperglycaemia and is present in people with impaired glucose tolerance, i.e. normal fasting glucose and glycohaemoglobin concentrations. These defects are subtle and include shifts to the right in the dose-response curves that relate to glucose and insulin secretion, reduced ability of the beta-cell to detect and respond to oscillatory changes in the plasma glucose concentration and impaired beta-cell compensation for insulin resistance. In order to define the factors responsible for these defects in secretion, we have infused human patients with a lipid emulsion and heparin to raise plasma free fatty acid concentrations. This is associated with reduced ability of the beta-cell to detect and respond to small changes in the plasma glucose concentration. In summary, defects in insulin secretion are consistently present in patients with impaired glucose tolerance and play a critical role in the progression from impaired glucose tolerance to diabetes. PMID- 10522837 TI - Approaches to the management of postprandial hyperglycaemia. AB - Increasing evidence suggests that tight metabolic control slows or prevents the development of diabetic complications. Various degrees of peripheral insulin resistance and inadequate glucose-induced insulin secretory profiles are seen in Type 2 diabetes. The main deficit in insulin release occurs early in the development of the disease and can be measured in association with meals, when the increase in meal-related insulin secretion is delayed and sluggish (first peak/early-phase defect). The consequences of this are high postprandial blood glucose concentrations and overall loss of glycaemic control. Improved prandial glucose metabolism will lead to an overall improvement in glycaemic control. The new concept of prandial glucose regulation, aimed at improving or restoring meal related insulin secretion, has led to the development of repaglinide, a carbamoylmethyl benzoic acid derivative, in the treatment of Type 2 diabetes. Patients treated with repaglinide using the concept 'one meal, one dose; no meal, no dose' experience improved insulin secretion, long-term metabolic control and quality of life and reduced postprandial hyperglycaemia. Treatment of Type 2 diabetes will be improved further through the use of combinations of drugs with different modes of action, which will prevent or reduce acute and chronic complications. New training programmes for physicians and patients must be developed to optimize the available pharmacological options for treating the extremely heterogeneous group of patients with Type 2 diabetes with a flexible oral therapeutic concept. An interdisciplinary approach to the care of these patients is urgently needed. PMID- 10522838 TI - Repaglinide as monotherapy in Type 2 diabetes. AB - The action of repaglinide, a carbamoylmethyl benzoic acid derivative, mimics the physiological insulin secretion that is deficient in Type 2 diabetes mellitus. Repaglinide stimulates insulin release from beta-cells only in the presence of glucose. Two placebo-controlled studies were performed to establish the effective dose range of repaglinide. In one study, repaglinide (0.25-4.0 mg preprandially) caused a dose-dependent decrease in blood glucose and a non-dose-dependent increase in insulin over 4 weeks (all doses p < 0.001 vs. placebo). In the second study, repaglinide (0.25-8.0 mg preprandially) was titrated over 6 weeks to obtain the optimum response (fasting plasma glucose < 8.9 mmol/L). The titration period was followed by a 12-week dose-maintenance period. At the end of the study, repaglinide had decreased fasting plasma glucose by 3.4 mmol/L (p < 0.05) and 2-h postprandial blood glucose by 5.8 mmol/L (p < 0.001) versus placebo. Glycated haemoglobin (HbA1c) decreased significantly from 8.5% to 7.9% in the repaglinide group and increased significantly from 8.1% to 9.2% in the placebo group (p < 0.001 between groups). In five 1-year, multicentre, randomized, double blind, phase III trials, repaglinide (0.5-4.0 mg preprandially) was compared with the sulphonylureas glibenclamide, glipizide and gliclazide. Repaglinide was more effective than glipizide at maintaining glycaemic control and was equivalent to glibenclamide and gliclazide on the basis of change in HbA1c. Hypoglycaemic events were reported in 16% of repaglinide-treated patients and 15-20% of sulphonylurea-treated patients. These data indicate that repaglinide monotherapy, with diet and exercise, is effective in patients with Type 2 diabetes. PMID- 10522839 TI - Repaglinide in combination therapy with metformin in Type 2 diabetes. AB - Results are presented of a double-blind, Australian multicentre study of the efficacy and safety of adjunctive repaglinide in patients with Type 2 diabetes who were controlled inadequately on metformin monotherapy. Patients had to have been on metformin for at least 6 months and to have glycated haemoglobin (HbA1c) levels of more than 7.1%. Patients were randomized to one of three treatment regimens: metformin plus placebo (MET, n = 27), repaglinide plus placebo (REP, n = 29) and metformin plus repaglinide (MET/REP, n = 27). The metformin dose remained unchanged from the prestudy dose, whereas repaglinide was titrated from 0.5 mg to 4.0 mg preprandially, depending on fasting capillary blood glucose concentration. Maintenance treatment was continued for 3 months. In the MET and REP groups, the HbA1c level decreased from 8.6% to 8.3% and from 8.6% to 8.2%, respectively; in the MET/REP group, HbA1c decreased from 8.3% to 6.9% (p < 0.001 vs. baseline; p < 0.05 vs. each monotherapy group). Overall, 59% of patients in the MET/REP group achieved a HbA1c level of less than 7.1% by the end of the study, compared with 20% and 22% in the MET and REP groups, respectively. No serious adverse events occurred that were considered to be related to study medication. Mild symptoms of hypoglycaemia were seen in the REP and MET/REP groups, in many cases during the titration phase. The combination of repaglinide with metformin was safe and well tolerated and produced a greater improvement in glycaemic control than that seen by the sum of the changes with the two agents alone. PMID- 10522840 TI - Mechanism of action of a new class of insulin secretagogues. AB - Repaglinide, a carbamoylmethyl benzoic acid (CMBA) derivative, belongs to a new class of antidiabetic agents structurally related to meglitinide (previously known as the non-sulphonylurea moiety of glibenclamide). Repaglinide and glibenclamide exert reciprocal competitive effects on their respective binding to insulinoma cells. Repaglinide does not affect the metabolism of D-glucose or endogenous nutrients or the biosynthesis of peptides in isolated rat pancreatic islets. Repeated intragastric administration of repaglinide to normal rats increases basal and glucose-stimulated peptide biosynthesis in isolated islets. The major primary action of repaglinide in islets is the closure of ATP-sensitive K+ channels. This agent decreases 86Rb outflow from prelabelled islets perifused in the absence of any exogenous nutrient and protects beta-cells against the inhibitory action of a diazoxide analogue on glucose-stimulated insulin release. The decrease in K+ conductance coincides with stimulation of Ca2+ influx into the islet cells. Meglitinide and its analogues stimulate insulin release more efficiently in the presence of D-glucose or other nutrient secretagogues than in their absence. They are efficient insulinotropic agents in animal models of Type 2 diabetes or in animals infused for 48 h with a hypertonic solution of D glucose. Repaglinide augments somatostatin secretion, without affecting glucagon release, in isolated perfused rat pancreases exposed to D-glucose. The insulinotropic action of repaglinide was documented in vivo after intravenous or oral administration in normal or Goto-Kakizaki rats. These preclinical investigations suggest that these new insulin secretagogues are well suited as potential insulinotropic tools in the treatment of Type 2 diabetes. PMID- 10522841 TI - Preclinical and clinical studies on safety and tolerability of repaglinide. AB - Repaglinide, an insulinotropic benzoic acid derivative, has been tested extensively in the preclinical and clinical setting for safety and tolerability. In preclinical safety trials, no clinically relevant laboratory or histopathological changes were found and repaglinide was not found to be genotoxic, immunogenic, teratogenic or tumorigenic at levels 50 times greater than the exposure in humans. In an ascending-dose tolerability study in Type 2 diabetic patients, no clinically relevant changes in liver enzymes or ECG occurred during the trial. A total of five active-controlled, long-term trials of identical design have been carried out (with 12 months of maintenance on a fixed, pretitrated dose), including 1228 patients on repaglinide and 597 on sulphonylureas, 417 of whom were on glibenclamide. The most common adverse event in the phase II studies was hypoglycaemia. The frequency of hypoglycaemia was identical for repaglinide and sulphonylureas; however, fewer nocturnal hypoglycaemic events were observed with repaglinide and no increase in the frequency of hypoglycaemic events occurred in elderly patients (> 65 years) compared with younger patients. Severe reactions (assistance required) were approximately twice as frequent in the comparator group. During these studies, two serious hypoglycaemic reactions were reported (coma, seizure or hospitalization), both occurring in patients on glibenclamide. The frequency of serious adverse events and serious cardiovascular events was comparable between repaglinide and sulphonylureas. PMID- 10522842 TI - Prediction of poorer prognosis by infection with antibiotic-resistant gram positive cocci than by infection with antibiotic-sensitive strains. AB - HYPOTHESES: Surgical patients with antibiotic-resistant gram-positive coccal (GPC) infections have a poorer prognosis than those with antibiotic-sensitive GPC infections, and colonization with resistant GPC predisposes to the development of resistant GPC infections. DESIGN: All infections among surgical patients from December 1, 1996, to December 1, 1998, were followed up prospectively. Patients with antibiotic-sensitive and antibiotic-resistant GPC infections were compared. Cohorts were also subdivided on the basis of GPC species, colonization status, and immunosuppression. SETTING: The surgical wards and intensive care units of a tertiary care, university hospital. MAIN OUTCOME MEASURES: In-hospital mortality, inhospital mortality during antibiotic therapy, length of stay, and length of stay from the time of initiation of antibiotics to discharge. RESULTS: Antibiotic resistant GPC infection compared ki4th antibiotic-sensitive GPC infection was associated with a higher mortality and previous colonization rate (25.8% and 31.0% vs 17.6% and 8.8%, respectively; P = .04 and P<.001, respectively) and a markedly longer length of stay (55.0 +/- 3.3 vs 31.0 +/- 2.0 days; P<.001). Length of stay and treatment to discharge times were longer after resistant Staphylococcus aureus infections than after resistant Staphylococcus epidermidis infections. The mortality and length of stay of patients with gentamicin resistant or vancomycin-resistant enterococcal infections were equivalently higher than those with antibiotic-sensitive enterococcal infections. Transplant recipients with resistant enterococcal infection had the highest mortality (41.9%). CONCLUSIONS: Surgical patients who develop antibiotic-resistant GPC infections have a significantly higher mortality rate, longer length of stay, and longer treatment to discharge time than patients with antibiotic-sensitive GPC infections. Colonization with resistant GPC predisposes to resistant GPC infection. Gentamicin-resistant enterococcus appears to be as virulent as vancomycin-resistant enterococcus. PMID- 10522843 TI - Six years of surgical wound infection surveillance at a tertiary care center: review of the microbiologic and epidemiological aspects of 20,007 wounds. AB - HYPOTHESES: (1) Antibiotic restriction policies result in alteration of microbiologic features of surgical site infections (SSIs) and (2) reported SSI rates are underestimated when postdischarge surveillance is not included in SSI surveillance efforts. DESIGN: Retrospective analysis of prospectively collected SSI surveillance data. PATIENTS AND METHODS: We compared initial microbial isolates from SSIs between (1) January 1, 1993, and December 31, 1995, and (2) January; 1, 1996, and December 31, 1998. Antibiotic restriction policies were implemented at Fairview-University Medical Center, Minneapolis, Minn, on March 1, 1995. For the combined periods (January 1, 1993, to December 31, 1998), we determined SSI rates for 20007 operations according to the extent of bacterial contamination at surgery (wound class). Then, we analyzed SSI rates for 10559 of these operations (selected based on availability of Anesthesia Society of America score and type of procedure) using the surgical wound risk index (wound class, Anesthesia Society of America score, and length of operation). We categorized SSI rates by 17 procedures for comparison with SSI rates reported by 286 hospitals that contributed data confidentially and voluntarily to the National Nosocomial Infections Surveillance System in 1998. We compared SSI rates with and without postdischarge surveillance. RESULTS: Coagulase-negative staphylococcus and group D enterococcus were the 2 most frequent isolates before and after antibiotic restriction policies were implemented. Candida albicans isolates decreased from 7.9% (1993-1995) to 6.5% (1996-1998; P=.46). Methicillin-resistant Staphylococcus aureus (1.8% of isolates) and vancomycin-resistant enterococcus (2.4% of isolates) organisms were first identified between 1996 and 1998. Our SSI rates were 2.6% for class I wounds, 3.6% for class II wounds, and 10.5% for class III/IV wounds; 53.9% of SSIs were identified after hospital discharge. CONCLUSIONS: Antibiotic restriction policies did not alter the microbial spectrum of SSIs during the observation period. Reporting SSI rates in the absence of postdischarge surveillance dramatically underestimates actual SSI rates, especially in tertiary care hospitals that provide care for large populations of elderly and immunosuppressed patients. PMID- 10522844 TI - Role of granulocyte-macrophage colony-stimulating factor and its receptor in the genesis of acute respiratory distress syndrome through an effect on neutrophil apoptosis. AB - HYPOTHESIS: That granulocyte-macrophage colony-stimulating factor (GM-CSF) and its receptor modulate the suppression of apoptosis (Ao) of normal neutrophils incubated in the plasma of patients with postraumatic acute respiratory distress syndrome (ARDS). DESIGN: Experimental study using cultured human neutrophils. SETTING: University hospital, level I trauma center. PARTICIPANTS: Plasma was obtained from 14 patients with early, fulminant posttraumatic ARDS (mean Injury Severity Score, 22). All samples were drawn within 24 hours after injury. Plasma was also taken from up to 21 healthy control subjects. These volunteers were also used as sources of polymorphonuclear leukocytes (PMNs). MAIN OUTCOME MEASURES: (1) Effect of early, fulminant ARDS and normal plasma on spontaneous Ao and GM CSF receptor expression in PMNs in vitro. (2) Effect of ligation of either GM-CSF or its receptor with a neutralizing monoclonal antibody (mAb) on PMN Ao in ARDS and normal plasma. (3) Correlation of extracellular GM-CSF concentration with rate of PMN Ao. (4) Levels of GM-CSF in ARDS and normal plasma and in culture supernatant of normal PMNs incubated in early, fulminant ARDS and normal plasma. RESULTS: Plasma from patients with ARDS enhanced PMN viability at 24 hours (data are given as mean +/- SEM) 52% +/- 3% control vs 60% +/- 3% ARDS, P<.05). Binding of the GM-CSF receptor with a neutralizing mAb significantly reduced PMN viability in ARDS plasma, but not in normal plasma (60% +/- 3% ARDS vs 53% +/- 3% ARDS + mAb, P<.05). Ligation of GM-CSF with mAb had no significant effect on PMN viability in either plasma. Only 1% of PMNs expressed detectable levels of the GM CSF receptor when incubated for 24 hours in either ARDS or normal plasma. The GM CSF levels were undetectable (>7 pg/mL) in both ARDS and normal plasma and in culture supernatants taken after 24 hours of incubation in both plasma types. Levels of GM-CSF ranging from 0 to 50000 pg/mL had no effect on PMN Ao in plasma free medium. CONCLUSIONS: The antiapoptotic effect of ARDS plasma appears to be mediated by the GM-CSF receptor. This effect occurs at both low levels of plasma GM-CSF and surface expression of its PMN receptor. Ligation of GM-CSF had no effect of PMN Ao, suggesting that Ao is triggered by Fc portion-mediated receptor cross-linking. These results provide the theoretical basis for alphaGM-CSF receptor mAb therapy as a novel modality of treatment for ARDS. PMID- 10522845 TI - Route of nutrition influences intercellular adhesion molecule-1 expression and neutrophil accumulation in intestine. AB - HYPOTHESIS: The levels of intestinal interleukin 10 and interleukin 4, inhibitors of intercellular adhesion molecule-1 (ICAM-1) expression, decline with total parenteral nutrition (TPN). These cytokine changes induced by lack of enteral nutrition may increase ICAM-1 expression, resulting in polymorphonuclear neutrophil accumulation in intestine. DESIGN: Prospective randomized experimental trials. SETTING: Laboratory. MATERIALS: Male mice. INTERVENTIONS: Sixty-three mice were randomized to chow, intravenous TPN, or intragastric TPN. MAIN OUTCOME MEASURES: Experiment 1: After diet manipulation, iodine 125-labeled anti-ICAM-1 antibody and iodine 131-labeled nonbinding antibody were injected to quantify ICAM-1 expression on endothelial cells in the lung, liver, kidney, and small intestine. Measurement of myeloperoxidase was used to quantify polymorphonuclear neutrophil accumulation in the organs. Experiment 2: Intestine was harvested for both ICAM-1 and myeloperoxidase levels after chow refeeding of mice in the intravenous TPN group. RESULTS: In experiment 1, uninjured mice fed intravenous TPN showed significantly increased intestinal ICAM-1 expression and polymorphonuclear neutrophil accumulation with no significant changes in the lung, liver, or kidney. No changes occurred in mice fed chow or intragastric TPN. In experiment 2, reinstitution of enteral feeding returned intestinal ICAM-1 and myeloperoxidase levels to normal. CONCLUSION: Gut changes associated with lack of enteral feeding induce endothelial changes and an immunologic response, which may influence subsequent responses to injury. PMID- 10522846 TI - Enough is enough. PMID- 10522847 TI - Radiofrequency ablation of breast cancer: first report of an emerging technology. AB - HYPOTHESIS: Radiofrequency (RF) energy applied to breast cancers will result in cancer cell death. DESIGN: Prospective nonrandomized interventional trial. SETTING: A university hospital tertiary care center. PATIENTS: Five women with locally advanced invasive breast cancer, aged 38 to 66 years, who were undergoing surgical resection of their tumor. One patient underwent preoperative chemotherapy and radiation therapy, 3 patients received preoperative chemotherapy, and 1 had no preoperative therapy. All patients completed the study. INTERVENTIONS: While patients were under general anesthesia and just before surgical resection, a 15-gauge insulated multiple-needle electrode was inserted into the tumor under sonographic guidance. Radiofrequency energy was applied at a low power by a preset protocol for a period of up to 30 minutes. Only a portion of the tumor was treated to evaluate the zone of RF ablation and the margin between ablated and nonablated tissue. Immediately after RF ablation, the tumor was surgically resected (4 mastectomies, 1 lumpectomy). Pathologic analysis included hematoxylin-eosin staining and enzyme histochemical analysis of cell viability with nicotinamide adenine dinucleotide-diaphorase (NADH diaphorase) staining of snap-frozen tissue to assess immediate cell death. MAIN OUTCOME MEASURE: Cancer cell death as visualized on hematoxylin-eosin-stained paraffin section and NADH-diaphorase cell viability stains. RESULTS: There was evidence of cell death in all patients. Hematoxylin-eosin staining showed complete cell death in 2 patients. In 3 patients there was a heterogeneous pattern of necrotic and normal-appearing cells within the ablated tissue. The ablated zone extended around the RF electrode for a diameter of 0.8 to 1.8 cm. NADH-diaphorase cell viability stains of the ablated tissue showed complete cell death in 4 patients. The fifth patient had a single focus of viable cells (<1 mm) partially lining a cyst. There were no perioperative complications related to RF ablation. CONCLUSIONS: Intraoperative RF ablation results in invasive breast cancer cell death. Based on this initial report of the use of RF ablation in breast cancer, this technique merits further investigation as a percutaneous minimally invasive modality for the local treatment of breast cancer. PMID- 10522848 TI - Importance of dissection of the hernial sac in laparoscopic surgery for large hiatal hernias. AB - HYPOTHESIS: Laparoscopic repair of large hiatal hernia is an appropriate management strategy. DESIGN: A prospective patient series. SETTING: A university teaching hospital. PATIENTS: All patients with hiatal hernias 10 cm or greater in diameter repaired laparoscopically between February 1, 1992, and September 30, 1998. INTERVENTIONS: Two operative strategies were used for laparoscopic repair: the first, which was used until early 1996, entailed initial esophageal dissection while leaving the sac in the mediastinum. The second involved preliminary dissection of the hernial sac from the mediastinum before dissecting the esophagus. MAIN OUTCOME MEASURES: Successful completion of the procedure using a laparoscopic technique, postoperative complication rate, reoperation rate, and clinical outcome. RESULTS: Eighty-six patients with a large hiatal hernia underwent attempted repair using laparoscopic methods. The median age was 63 years (range, 30-91 years), and 45 patients (52%) were women. There were 30 sliding, 10 rolling, and 46 mixed hiatal hernias. Operating times ranged from 48 to 240 minutes (median, 90 minutes), and 20 procedures (23%) were converted to an open operation. Conversion was significantly more common in the first half of our experience (16 [40%] of 40 patients vs 4 [9%] of 46 patients) before the operative strategy was changed. Esophageal-lengthening procedures were not carried out for any patient. At follow-up of a median of 2 years, 1 patient has moderate dysphagia, 4 patients have reflux symptoms, and 1 patient has undergone further surgery for a recurrent paraesophageal hernia. An overall satisfactory outcome was achieved in 81 patients (94%). CONCLUSIONS: Large hiatal hernias can be treated effectively laparoscopically. Dissecting the sac fully from the mediastinum before dissecting the esophagus helps to safely mobilize the esophagus, and we think changing to this strategy is the main reason for the improved laparoscopic success rate reported in the latter half of this series. PMID- 10522849 TI - Extracellular signal-related kinase 1/2 and p38 mitogen-activated protein kinase pathways serve opposite roles in neutrophil cytotoxicity. AB - BACKGROUND: Inflammatory stimuli rapidly activate mitogen-activated protein kinases (MAPKs) in neutrophils (PMNs). However, their role in cytotoxic function remains unknown. Elucidating the signals involved in release of cytotoxic agents from PMNs may provide new avenues for therapy in diseases of diminished or excessive PMN function. HYPOTHESIS: The p38 MAPK and extracellular signal-related kinase 1/2 (ERK1/2) modulate superoxide generation and elastase release in activated human PMNs. STUDY DESIGN: Isolated human PMNs were incubated with specific inhibitors of MAPK pathways, or vehicle control solution, before activation with the bacterial peptide f-Met-Leu-Phe. MAIN OUTCOME MEASURES: The rate of superoxide release from activated PMNs was measured by the superoxide dismutase-inhibitable reduction of cytochrome-c. Elastase release from PMNs was determined by cleavage of the substrate Ala-Ala-Pro-Val-pNA. RESULTS: Superoxide release from activated PMNs was inhibited by blockade of p38 MAPK activation but unaffected by blockade of ERK1/2. Conversely, elastase release was unaffected by p38 MAPK inhibition and increased by ERK1/2 inhibition. CONCLUSIONS: Activation of p38 MAPK promotes superoxide release from PMNs activated by f-Met-Leu-Phe. The ERK1/2 pathway may serve as a negative feedback mechanism for granule exocytosis. PMID- 10522850 TI - L-selectin and leukocyte function in skeletal muscle reperfusion injury. AB - HYPOTHESIS: Treatment with anti-L-selectin monoclonal antibody will reduce venular neutrophil-endothelial rolling (flux and velocity) and adhesion associated with ischemia reperfusion injury in rat skeletal muscle. DESIGN: Prospective, randomized experimental trials. SETTING: Basic science research laboratory. MATERIALS: Male Wistar rats weighing 109 +/- 5 g (mean +/- SEM). INTERVENTIONS: Gracilis pedicle muscle flaps were elevated and microcirculation was observed by intravital microscopy. Two groups were evaluated: (1) the control group, which received 4 hours of global ischemia, and (2) the experimental group, which received 4 hours of global ischemia, plus treatment with anti-L-selectin monoclonal antibody 30 minutes before reperfusion. MAIN OUTCOME MEASURES: The number of rolling and adherent leukocytes in postcapillary venules were counted in the 2 groups at baseline and at 1 through 5, 10, 15, 20, 30, 45, and 60 minutes of reperfusion. RESULTS: Treatment with the monoclonal antibody to L selectin significantly reduced the number of rolling leukocytes (flux) at 2 through 5, 20, 30, 45, and 60 minutes of reperfusion compared with controls (P<.05). Use of the monoclonal antibody significantly reduced the number of adherent neutrophils at 5, 10, 15, 20, 30, 45, and 60 minutes of reperfusion (P<.05). There was no significant difference in leukocyte velocity. CONCLUSION: L Selectin plays a significant role in leukocyte rolling and adherence to venular endothelium in rat skeletal muscle ischemia reperfusion injury. PMID- 10522851 TI - Intra-abdominal hypertension is an independent cause of postoperative renal impairment. AB - HYPOTHESIS: Intra-abdominal hypertension exerts an effect on renal function independent of other confounding variables. DESIGN: A prospective study of all patients admitted to an intensive care unit following abdominal surgery was undertaken between September 1, 1994, and July 31, 1997, in a single university hospital. MAIN OUTCOME MEASURES: Intra-abdominal pressure (IAP) was measured every 8 hours (normal IAP, 0-17 mm Hg); 18 mm Hg or higher was considered increased. Forward stepwise logistic regression determined whether intra abdominal hypertension is an independent cause of renal impairment. RESULTS: A total of 263 patients (174 after emergency surgery), whose mean +/- SD age was 61.0 +/- 18.7 years and Acute Physiology and Chronic Health Evaluation II score was 14.6 +/- 7.7, were studied. Intra-abdominal pressure was increased in 107 (40.7%) of the 263 patients. Renal impairment occurred in 35 (32.7%) of the 107 patients with intra-abdominal hypertension and in 22 (14.1%) of the 156 with a normal IAP (odds ratio, 1.62-5.42). Using the Wald maximizing model, renal impairment was independently associated with 4 antecedent factors: hypotension (P= .09), sepsis (P = .006), age older than 60 years (P = .03), and increased IAP (P = .004). CONCLUSIONS: To our knowledge, for the first time in a large clinical study, IAP has been shown to be an independent cause of renal impairment, and it ranks in importance after hypotension, sepsis, and age older than 60 years. Surgeons need to be aware of the importance of intra-abdominal hypertension in patients postoperatively. PMID- 10522852 TI - Blood supply to the pancreatic head, bile duct, and duodenum: evaluation by computed tomography during arteriography. AB - HYPOTHESIS: Blood supply to the peripancreatic region is derived from the celiac and superior mesenteric arteries, complementary to each other. DESIGN: Cohort analytic study. SETTING: Tertiary care public hospital. PATIENTS AND METHODS: Computed tomography (CT) during superior mesenteric artery arteriography (SMAA CT) and during celiac artery arteriography (CEAA-CT), in which a catheter tip was inserted into the common hepatic or gastroduodenal artery, was performed in 25 patients. MAIN OUTCOME MEASURE: Distribution and correlation of these areas of marked enhancement on SMAA-CT and CEAA-CT were analyzed. RESULTS: The right cephalic part of the pancreatic head that is derived from the dorsal bud was enhanced on CEAA-CT, and the left caudal part of the pancreatic head that is derived from the ventral bud was enhanced on SMAA-CT. Blood supply to the intrapancreatic bile duct, including the ampulla of Vater, is derived from the CEA. The boundary between the areas of the duodenum supplied from the CEA and SMA was in the second or third portion. CONCLUSION: The pancreatic head can be separated into 2 segments by the arterial supply, and each segment may be removed separately. PMID- 10522853 TI - Cardiovascular effect of 7.5% sodium chloride-dextran infusion after thermal injury. AB - HYPOTHESIS: Clinical study can help determine the safety and cardiovascular and systemic effects of an early infusion of 7.5% sodium chloride in 6% dextran-70 (hypertonic saline-dextran-70 [HSD]) given as an adjuvant to a standard resuscitation with lactated Ringer (RL) solution following severe thermal injury. DESIGN: Prospective clinical study. SETTING: Intensive care unit of tertiary referral burn care center. PATIENTS: Eighteen patients with thermal injury over more than 35% of the total body surface area (TBSA) (range, 36%-71%) were studied. INTERVENTIONS: Eight patients (mean +/- SEM, 48.2% +/- 2% TBSA) received a 4-mL/kg HSD infusion approximately 3.5 hours (range, 1.5-5.0 hours) after thermal injury in addition to routine RL resuscitation. Ten patients (46.0% +/- 6% TBSA) received RL resuscitation alone. MAIN OUTCOME MEASURES: Pulmonary artery catheters were employed to monitor cardiac function, while hemodynamic, metabolic, and biochemical measurements were taken for 24 hours. RESULTS: Serum troponin I levels, while detectable in all patients, were significantly lower after HSD compared with RL alone (mean +/- SEM, 0.45 +/- 0.32 vs 1.35 +/- 0.35 microg/L at 8 hours, 0.88 +/- 0.55 vs 2.21 +/- 0.35 microg/L at 12 hours). While cardiac output increased proportionately between 4 and 24 hours in both groups (from 5.79 +/- 0.8 to 9.45 +/- 1.1 L/min [mean +/- SEM] for HSD vs from 5.4 +/- 0.4 to 9.46 +/- 1.22 L/min for RL), filling pressure (central venous pressure and pulmonary capillary wedge pressure) remained low for 12 hours after HSD infusion (P = .048). Total fluid requirements at 8 hours (2.76 +/- 0.7 mL/kg per each 1% TBSA burned [mean +/- SEM] for HSD vs 2.67 +/- 0.24 mL/kg per each 1% TBSA burned for RL) and 24 hours (6.11 +/- 4.4 vs 6.76 +/- 0.75 mL/kg per each 1% TBSA burned) were similar. Blood pressure remained unchanged, and serum sodium levels did not exceed 150 +/- 2 mmol/L (mean +/- SD) in either group. CONCLUSIONS: The absence of deleterious hemodynamic or metabolic side effects following HSD infusion in patients with major thermal injury confirms the safety of this resuscitation strategy. Postburn cardiac dysfunction was demonstrated in all burn patients through the use of cardiospecific serum markers and pulmonary artery catheter monitoring. Early administration of HSD after a severe thermal injury may reduce burn-related cardiac dysfunction, but it had no effect on the volume of resuscitation or serum biochemistry values. PMID- 10522854 TI - Attenuation of the acute-phase response in thermally injured rats by cholesterol containing cationic liposomes used as a delivery system for gene therapy. AB - HYPOTHESIS: Cholesterol-containing cationic liposomes alone modulate the acute phase response and cytokine expression in thermally injured rats and are an effective delivery system for gene therapy in trauma. SETTING: Laboratory. INTERVENTION: Fifty-six adult male Sprague-Dawley rats with a full-thickness scald burn covering 60% of total body surface area were randomly divided into 2 groups to receive either intravenous injections of cholesterol-containing cationic liposomes or saline (control). MAIN OUTCOME MEASURES: Body weights, muscle and liver dry-wet weights, serum levels of constitutive hepatic proteins, acute-phase protein levels, and cytokine levels were determined at 1, 2, 5, and 7 days after thermal injury. RESULTS: Rats receiving cholesterol-containing cationic liposomes had less body weight loss, increased serum transferrin levels, and decreased serum alpha1-acid glycoprotein levels when compared with controls (P<.05). Serum interleukin 1beta and tumor necrosis factor alpha levels were decreased in rats receiving liposomes at 1 and 2 days after burn compared with controls (P<.05). CONCLUSIONS: These results suggest that cholesterol-containing cationic liposomes alone may have a beneficial effect in modulating the hypermetabolic response after burn injury by decreasing type 1 acute-phase proteins and the expression of the proinflammatory cytokines interleukin 1beta and tumor necrosis factor alpha. Therefore, cholesterol-containing cationic liposomes appear to be suitable as a delivery system for gene therapy in trauma. PMID- 10522855 TI - Spontaneous rupture of hepatocellular carcinoma: conservative management and selective intervention. AB - HYPOTHESIS: A conservative approach using selective intervention is better than an aggressive approach using nonselective intervention for ruptured hepatocellular carcinoma. DESIGN: Nonrandomized controlled trial. SETTING: A university hospital. PATIENTS AND INTERVENTIONS: From 1984 to 1990, an aggressive approach was adopted in which 29 and 8 of a total of 40 patients underwent surgical intervention or attempted transarterial embolization (TAE), respectively. From 1991 to 1997, a more conservative approach was used. The initial treatment for 72 patients was conservative with close monitoring. Additional hemostatic procedures consisting of TAE (n = 13) or surgical intervention (n = 9) were given, depending on the clinical progress, disease status, and liver function of the patients. MAIN OUTCOME MEASURES: In-hospital mortality, survival. RESULTS: In-hospital mortality rate was 62% (25 of 40 patients) in the first period and 51% (37 of 72 patients) in the second period. The respective median survival times were 7 and 12 days. If 36 patients with end stage malignant neoplasms were excluded, the in-hospital mortality rate became 60% (18 of 30 patients) in the first period and 35% (16 of 46 patients) in the second period (P = .03, chi2 test). The respective median survival times became 8 and 72 days (P = .02, log rank test). In the second period, 7 (54%) of 13 patients who underwent TAE and 1 (11%) of 9 patients who underwent surgical intervention died within the same hospital admission (P = .07, Fisher exact test). CONCLUSIONS: Selective intervention was cost-effective and gave better results than an aggressive approach. When intervention was indicated for hemostasis, surgery seemed better than TAE although the difference was not statistically significant. PMID- 10522856 TI - General weakness as an indication for parathyroid surgery in patients with secondary hyperparathyroidism. AB - HYPOTHESES: There are factors that affect patients with general weakness owing to secondary hyperparathyroidism and as reported by results noted after parathyroidectomy and autotransplantation. DESIGN: Case series and consecutive samples. SETTING: Tertiary care center. PATIENTS: From July 1996 to June 1998, 56 patients with secondary hyperparathyroidism underwent total parathyroidectomy and autotransplantation. Their ages were 45 +/- 13 years (mean +/- SD) and preoperative duration of dialysis was 75 +/- 37 months. Prior to surgery the patients were divided into 2 groups: group A comprised 2 men and 19 women who had some general weakness; and group B, 15 men and 20 women who reported no general weakness. The etiologies of renal failure, such as diabetic nephropathy (n = 3) or hypertensive nephropathy (n = 3), were found only in group A patients. INTERVENTIONS: Serum levels of calcium, phosphorus, alkaline phosphatase, and parathyroid hormone (intact) were checked preoperatively and 1 day, 1 week, and 3 months after surgery. Extension force of the quadriceps femoris muscle was measured at 60 degrees of right knee flexion preoperatively and 3 months after surgery. The extension force was expressed as newtons (N) in 2 different quantities: peak force and average force. The degree of general weakness was classified into 4 groups: 0, no weakness; 1, some subjective weakness and/or walking with assistance; 2, the patient was wheelchair bound; and 3, the patient was bedridden. MAIN OUTCOME MEASURES: The t test was used for paired and unpaired data; Wilcoxon signed rank and Fisher exact tests were incorporated for nonparametric data. Any values of P<.05 were considered significant. RESULTS: Prior to surgery, 2 patients in group A reported degree 3 weakness; 5, degree 2 weakness; and 14, degree 1 weakness. Three months after surgery, the peak force was noticed to have increased from 185 +/- 56 N to 249 +/- 82 N (mean +/- SD) (n = 11, P = .003), and the average force showed an increase from 136 +/- 45 N to 202 +/- 69 N (n = 11, P = .003). Postoperatively, only 5 patients had degree 1 weakness, 1 had degree 2 weakness, and none had degree 3 weakness. The patient with degree 2 weakness after surgery had diabetes mellitus and a femoral neck fracture prior to parathyroidectomy. Improvement in condition of general weakness was found (P<.001) between preoperative and postoperative periods. Serum levels of calcium were higher in group A (2.82 +/- 0.23 mmol/L [11.3 +/- 0.9 mg/dL]) than in group B (2.64 +/- 0.27 mmol/L [10.6 +/- 1.1 mg/dL]) (P = .013), and serum levels of parathyroid hormone (intact) were lower in group A (108.9 +/- 39.2 pmol/L) than in group B (139.8 +/- 39.6 pmol/L) (P = .006). Except for sex, other data such as phosphorus and alkaline phosphatase levels, age, and duration of dialysis were not significantly different between the 2 groups. CONCLUSIONS: General weakness that is commonly observed in patients with secondary hyperparathyroidism is found more frequently in women and only in patients with diabetic nephropathy or hypertensive nephropathy. Patients with general weakness had relatively higher levels of calcium and lower levels of parathyroid hormone (intact). We found that improvement of muscle power and general weakness can be achieved by parathyroidectomy and autotransplantation. In addition to itchy skin, bone pain, and soft tissue calcification, general weakness that may cause disability is also an indication for surgery in secondary hyperparathyrodism. PMID- 10522857 TI - Cardiovascular and respiratory changes and convalescence in laparoscopic colonic surgery: comparison between carbon dioxide pneumoperitoneum and gasless laparoscopy. AB - HYPOTHESIS: Gasless laparoscopy produces smaller cardiopulmonary and systemic changes than carbon dioxide (CO2) laparoscopy during colonic surgery. DESIGN: Prospective randomized trial. SETTING: Department of Surgery in a university hospital. PATIENTS: Twenty-two patients scheduled for laparoscopic colonic resection; 5 patients were excluded because of conversion to open surgery (N = 17). INTERVENTIONS: Patients were randomized to either gasless (n = 9) or conventional CO2 (n = 8) surgery. MAIN OUTCOME MEASURES: Intraoperative assessment of hemodynamic factors and pulmonary function, and postoperative assessment of pain, pulmonary function, convalescence, and various injury factors were done several times until 30 days after surgery. Surgical complications were noted. RESULTS: Descending aorta blood flow after 30 minutes (P=.03) and heart rate after 150 minutes were higher in the CO2 group (P=.009). Central venous pressure, PaCO2 inspiration pressure, and end tidal CO2 level were significantly higher in the CO2 group (P = .05, .03, .04, and .01, respectively). Patients in the CO2 group had less pain during mobilization and coughing (P = .008 and .006, respectively), and were significantly more fatigued (P = .04). No other important differences were observed in intraoperative hemodynamic factors, postoperative convalescence, immunocompetence, or pulmonary function. CONCLUSION: No clinically important differences in cardiovascular and systemic response were observed between patients undergoing CO2 or gasless laparoscopy for colonic disease. PMID- 10522858 TI - Osteoporosis in multiple endocrine neoplasia type 1: severity, clinical significance, relationship to primary hyperparathyroidism, and response to parathyroidectomy. AB - BACKGROUND: Sporadic primary hyperparathyroidism (PHPT) occurs most frequently in postmenopausal women. Multiple endocrine neoplasia type 1 (MEN 1) is an autosomal dominant disease in which mild to moderate PHPT develops in most gene carriers by 20 years of age. Primary hyperparathyroidism associated with MEN 1 is typically recurrent, despite initially successful subtotal parathyroidectomy. Osteoporosis is considered a complication of sporadic PHPT and an indication for parathyroidectomy. In the setting of MEN 1, however, the relationship of bone mass to PHPT, fracture risk, and parathyroidectomy is unknown. HYPOTHESIS: Parathyroidectomy improves bone mineral density for patients with primary hyperparathyroidism in the setting of MEN 1. DESIGN: Case series. SETTING: Tertiary referral center. PATIENTS: Twenty-nine women with MEN 1 belonging to a single family with a history of MEN 1. INTERVENTIONS: Parathyroidectomy. MAIN OUTCOME MEASURES: Bone mineral density (BMD) and history of skeletal fracture. RESULTS: Osteopenia and osteoporosis were diagnosed in 41% and 45% of patients, respectively. Forty-four percent of patients with uncontrolled PHPT had severe osteopenia (T score, <-2.0) by 35 years of age. Reduction in BMD was greatest at the femoral neck. Reduced BMD was associated with an increased likelihood of skeletal fracture (P = .05). Patients with uncontrolled PHPT had lower femoral neck and lumbar spine BMDs than those in whom PHPT was controlled by parathyroidectomy (P = .005 and .02, respectively). Successful parathyroidectomy improved femoral neck and lumbar spine BMDs by a mean +/- SEM of 5.2% +/- 2.5% and 3.2% +/- 2.9%, respectively. CONCLUSIONS: Osteoporosis is a frequent and early complication of PHPT in MEN 1. Despite difficulty in achieving a cure of PHPT in MEN 1, parathyroidectomy has an important role in the optimization of BMD for patients with MEN 1. PMID- 10522859 TI - Hepatectomy for hepatocellular carcinoma: the surgeon's role in long-term survival. AB - HYPOTHESIS: The surgeon can contribute substantially to the long-term survival rate of patients undergoing hepatectomy for hepatocellular carcinoma (HCC). DESIGN: The long-term survival rate of patients with HCC undergoing hepatectomy has improved, but the contribution of the surgeon to the improved survival rate is unknown. We surveyed 211 consecutive patients undergoing hepatectomy for HCC. The clinical, operative, and pathological factors were analyzed to identify factors that were important in affecting long-term survival. SETTING: A tertiary referral center. PATIENTS: From April 1989 to December 1995, 211 consecutive patients with HCC underwent 153 major and 58 minor hepatectomies. MAIN OUTCOME MEASURES: Disease-free and overall cumulative survival rate. RESULTS: The 5-year disease-free survival rate was 27%. By Cox regression analysis, blood transfusion (relative risk [RR], 1.21; 95% confidence interval [CI], 1.05-1.40) and TNM stage (RR, 1.90; 95% CI, 1.47-2.47) were shown to be independent prognostic factors in the 5-year disease-free survival rate. The 5-year overall cumulative survival rate was 37%. By Cox regression analysis, the preoperative indocyanine green retention value at 15 minutes after injection (RR, 1.03; 95% CI, 1.01-1.06), blood transfusion (RR, 1.191; 95% CI, 1.078-1.316), tumor rupture (RR, 1.48; 95% CI, 1.08-2.04), and TNM stage (RR, 1.62; 95% CI, 1.27-2.07) were shown to be significant independent factors that influenced cumulative survival rate. CONCLUSIONS: The long-term survival of patients with HCC after hepatectomy depends on tumor staging, preoperative hepatic functional reserve, history of blood transfusion, and rupture of HCC. Preoperative liver function and tumor staging cannot be altered; however, the surgeon can play an important role in improving the prognosis if blood transfusion and iatrogenic tumor rupture can be avoided and if function of the liver remnant can be preserved. PMID- 10522860 TI - Intraductal papillary mucinous tumors of the pancreas comprise 2 clinical subtypes: differences in clinical characteristics and surgical management. AB - HYPOTHESIS: Intraductal papillary mucinous tumors (IPMTs) of the pancreas may be meaningfully construed as representing 2 clinically distinct subtypes: main duct tumors (MDT) and branch duct tumors (BDT). DESIGN: Retrospective study. SETTING: University hospital from January 1988 through December 1994. PATIENTS AND INTERVENTION: We reviewed diagnostic findings and late results of surgical treatment in 30 patients with IPMT. RESULTS: The tumor was located in the head of the pancreas more often in BDT than in MDT (65% [11/17] and 23% [3/13], respectively). Of the 13 patients with MDTs, 12 (92%) had intraductal papillary adenocarcinoma (noninvasive and minimally invasive types) and/or carcinoma in situ (carcinoma in situ: low papillary and/or flat tumor cells), and 3 (23%) had stromal invasion. Of the 17 patients with BDTs, 5 (29%) had intraductal papillary adenocarcinoma and/or carcinoma in situ. Two pancreatoduodenectomies and 8 pylorus-preserving pancreatoduodenectomies were performed in 10 of the 17 patients with BDTs, distal pancreatectomy in 7 patients with MDTs, and total pancreatectomy in 4 patients with MDTs. The 5-year survival rates were 47% for MDT and 90% for BDT. Four of 6 patients with MDTs who died had local recurrence. One patient died of liver metastasis and 1 of esophageal cancer. Only 1 patient with BDT of the 2 who died had recurrent disease. CONCLUSIONS: Intraductal papillary mucinous tumors may be composed of 2 clinically distinct subtypes: MDTs and BDTs. Initially, although distal pancreatectomy can be recommended for most MDTs, the need for cancer-free margins in this more aggressive type may necessitate total pancreatectomy. Pylorus-perserving pancreatoduodenectomies is recommended for most BDTs, but, because these tumors are more often adenomas, a good prognosis can be expected. PMID- 10522861 TI - Effect of multiple gene transfers of insulinlike growth factor I complementary DNA gene constructs in rats after thermal injury. AB - HYPOTHESIS: Multiple subcutaneous injections of cholesterol-containing cationic liposomes encapsulating the complementary DNA (cDNA) gene for insulinlike growth factor I (IGF-I) increase the rate of transfected skin cells and result in increased IGF-I protein levels in the skin with subsequent improvement in wound healing when compared with a single injection. SETTING: Laboratory. INTERVENTION: Twenty-four adult male Sprague-Dawley rats (350-375 g) received a full-thickness scald burn on 60% of their body surface. These rats were randomly divided to receive either 1 injection of liposomes containing 2.2 microg-cytomegalovirus driven cDNA coding for IGF-I and 0.2 microg of the Lac Z gene cDNA construct, or 2 injections of liposomes containing 2.2 microg cytomegalovirus-driven cDNA coding for IGF-I and 0.2 microg of the Lac Z gene cDNA construct. MAIN OUTCOME MEASURES: Transfection rates and IGF-I protein levels in the skin and physiological responses to the IGF-I gene therapy, evaluated from changes in body weight, protein content in serum and liver, and the rate of burn wound healing. RESULTS: There was a significant decrease in transfection rate and IGF-I protein expression distal from the injection site in animals receiving 1 injection, as compared with a consistent increase in rats receiving multiple injections. Multiple injections improved the response to thermal trauma by increasing the extent of the healed burn wound 33 days after thermal injury (single injection, 31% +/- 1% vs multiple injections, 38% +/- 2%), total serum protein (single injection, 52 +/- 0.5 g/L vs multiple injections, 55 +/- 0.6 g/L), and total liver protein (single injection, 82.0 +/- 0.3 mg/mL vs multiple injections, 91.0 +/- 3.8 mg/mL), P<.05. CONCLUSIONS: Gene transfer rates can be increased by multiple injections of liposomes encapsulating IGF-I cDNA constructs. Increased transfer results in greater IGF-I protein skin concentrations, accelerated wound healing, and increased serum and liver protein concentrations. The clinical relevance of these findings is that liposomal gene constructs should be applied in well-defined distances to improve gene transfer in the skin, and thus clinical outcome after thermal injury. PMID- 10522862 TI - Localization of atherosclerosis: role of hemodynamics. AB - Atherosclerosis is a chronic disease attributed to risk factors that are systemic in nature. Yet the lesions involved do not occur in random fashion. The coronary arteries, the major branches of the aortic arch, and the abdominal aorta and its visceral and major lower extremity branches are particularly susceptible sites. Hemodynamic forces interacting with an active vascular endothelium are responsible for localizing lesions in a nonrandom pattern of distribution. Shear stress and cyclic circumferential strain are the predominant forces that have been characterized. The modification of endothelial cell structure and function by these mechanical forces sheds insight into the vasculature's propensity for atherogenesis. PMID- 10522863 TI - Incomplete pancreas divisum with anomalous choledochopancreatic duct junction with choledochal cyst. AB - The coexistence of incomplete pancreas divisum, an anomalous choledochopancreatic duct junction, and a choledochal cyst is an extremely rare condition, described in only 3 patients in the available medical literature. The symptoms may be similar to any of these 3 distinct pathological conditions. Magnetic resonance cholangiopancreatography or endoscopic retrograde cholangiopancreatography is usually required for diagnosis. Management of symptomatic pancreas divisum may be accomplished with open accessory duct sphincteroplasty or endoscopic papillotomy with or without stenting. Treatment of choledochal cyst is by complete excision of the cyst whenever possible, with hepaticodochoenterostomy being the treatment of choice. Here, we describe a patient with this complex disorder who was successfully managed with endoscopic minor duct papillotomy with accessory pancreatic duct stenting and resection of the choledochal cyst with hepaticodochojejunostomy. PMID- 10522864 TI - A surgical sponge and medical malpractice in 1856. PMID- 10522865 TI - Repeats may not be everything in anticipation. PMID- 10522866 TI - Why drugs fail in postpolio syndrome: lessons from another clinical trial. PMID- 10522868 TI - Federal funding for graduate medical education. PMID- 10522867 TI - Carbamazepine in comparative trials: pharmacokinetic characteristics too often forgotten. AB - OBJECTIVES: To compare the use of carbamazepine (CBZ) in comparative monotherapy trials with its use in practice. BACKGROUND: CBZ is often the drug of first choice for partial onset seizures and is frequently used as the reference drug in comparative trials with newly developed antiepileptic compounds. There are several issues related to CBZ that may have an impact on the final outcome of these trials. CBZ has nonlinear time-dependent kinetics due to autoinduction, and this may cause adverse reactions on starting treatment; somnolence and rash usually occur in the first weeks and may be related to the dose and titration schedule. Total daily dose or the serum levels of the drug do not correlate with therapeutic efficacy. METHODS: Review of the current literature on the pharmacokinetics properties of CBZ and of comparative trials of new antiepileptic drugs that used CBZ as the reference drug. CONCLUSIONS: The use of CBZ differed in comparative clinical trials and in clinical practice, with slower and lower titration rates used in the latter. This may be disadvantageous for CBZ because its pharmacokinetics peculiarities are ignored. If CBZ is to be used in these trials, titration rates and dosages should resemble those used in clinical practice because this is likely to improve efficacy and tolerability. Otherwise, the results of these trials may support regulatory applications but have little relevance to clinical practice. We suggest an initial CBZ dose of 100 mg/d and a slow titration schedule with dose incremental steps of 100 mg/d every week up to 600 mg/d. Blood levels should determine the initial target dose, but after this has been achieved titration should be based on seizure control and tolerability rather than by blood levels. PMID- 10522869 TI - Genetic localization of the familial adult myoclonic epilepsy (FAME) gene to chromosome 8q24. AB - OBJECTIVE: To identify the genetic locus for the familial adult myoclonic epilepsy (FAME) gene. BACKGROUND: Idiopathic generalized epilepsy (IGE) represents a collection of disorders in which affected individuals present with recurring seizures that have diffuse onset on EEG. These individuals have no known structural cerebral lesions or other identifiable etiology. IGE accounts for approximately 40% of all epilepsies. FAME is a type of IGE characterized by autosomal dominant inheritance, adult onset, varying degrees of myoclonus in the limbs, rare tonic-clonic seizures, and a benign course. METHODS: We investigated four previously reported Japanese kindreds and performed a genome-wide screen with genetic linkage analysis. RESULTS: Clinical characterization and sampling of 30 individuals in four families revealed that 21 had the FAME phenotype. We defined a 4.6-cM region on chromosome 8q24 (maximum lod score of 4.86 at theta = 0) that contains the FAME gene. CONCLUSIONS: The identification and characterization of the FAME gene allows us to better understand the molecular basis of FAME. Such knowledge may provide clues to understanding the molecular basis of the clinically similar, and more common, juvenile myoclonic epilepsies, and other generalized seizure disorders that have thus far eluded genetic approaches. PMID- 10522870 TI - Multisequence MRI in clinically isolated syndromes and the early development of MS. AB - OBJECTIVE: To apply multisequence MRI techniques to patients with clinically isolated syndromes, to document the pattern and frequency of abnormalities at baseline and early follow-up, and to determine their predictive values for the early development of clinical MS. BACKGROUND: Disseminated lesions on T2-weighted brain MRI confer an increased risk of progression to clinically definite MS. Newer MRI techniques increase detection of lesions in both brain and spinal cord, and clarify further their pathology. The predictive value of such techniques for the development of clinical MS needs to be defined. METHODS: Brain and spinal MRI were performed on 60 patients after their first demyelinating event. A total of 50 patients were followed for 1 year, and 49 underwent repeat brain MRI 3 months after the initial scan. RESULTS: At baseline, 73% of patients had lesions on T2 weighted fast spin-echo (FSE) brain images and 42% had asymptomatic spinal cord lesions. Fast fluid-attenuated inversion-recovery brain did not improve detection of brain lesions. Repeat brain MRI demonstrated new FSE lesions in 43% of patients. After 1 year, 26% of patients developed MS. The MRI features that provided the best combination of sensitivity and specificity for the development of MS were the presence of new FSE lesions at follow-up and enhancing lesions at baseline. The frequency of developing clinical MS was higher for those with both brain and spinal cord lesions at baseline (48%) than brain lesions alone (18%). CONCLUSIONS: The combination of baseline MRI abnormalities and new lesions at follow-up, indicating dissemination in space and time, was associated with a high sensitivity and specificity for the early development of clinical MS. These data suggest a potential role for new diagnostic criteria for MS based on early MRI activity. Such criteria may be useful in selecting patients for therapeutic trials at this early clinical stage. PMID- 10522871 TI - General neurologist and subspecialist care for multiple sclerosis: patients' perceptions. AB - OBJECTIVE: To compare general neurologists and MS specialists on patients' clinical characteristics and MS care as perceived by patients with MS. METHODS: We sampled all adult patients with MS having physician visits over a 2-year period from a Midwestern managed-care organization and from the fee-for-service portion of 23 randomly selected California neurologists' practices. In mid-1996, 694 subjects were mailed questionnaires; 532 (77%) responded. Sociodemographic/clinical characteristics, recent utilization of services/treatments, unmet needs, symptom care, and research participation were measured. Of 502 subjects (94%) who indicated their usual physician providing MS care was a neurologist, 217 (43%) reported having a general neurologist and 285 (57%) reported having an MS specialist. Comparisons between these two groups were adjusted for comorbidity and disease severity. RESULTS: General neurologist and MS specialist patient groups did not differ on any sociodemographic or clinical characteristic except age (p<0.05). Although health care utilization generally was similar, higher proportions of the MS specialist group were aware of or had discussed interferon beta-1b (IFNbeta-1b) with their physician (p<0.05) and were currently taking it (p<0.05); a smaller proportion of the MS specialist group reported stopping it because of side effects (p<0.01). Overall, levels of unmet need and care for recent symptoms were similar, but the MS specialist group reported more confidence in their physician/carefulness in listening (p<0.05). Twice as many MS specialist subjects had participated in nondrug research (p<0.05); drug study participation was similar. CONCLUSIONS: Patients' perceptions of their care were similar in most ways for those who designated their main MS provider as a general neurologist compared to an MS specialist; however, care differed in potentially important areas. Prospective, longitudinal studies are needed to measure and relate neurologists' training, experience, knowledge, and MS patient volume with both process and outcome measures of quality of MS care. PMID- 10522872 TI - Incubation period of Creutzfeldt-Jakob disease in human growth hormone recipients in France. AB - OBJECTIVE: To estimate the statistical distribution of the incubation period of Creutzfeldt-Jakob disease (CJD) in human growth hormone (hGH) recipients in France. BACKGROUND: Published papers suggest that the median incubation period of hGH-related CJD is approximately 15 years, but there are as yet no statistical data that support this assertion. METHODS: Of the 1,361 hGH recipients who were included in this study, 55 had developed CJD at the time of the study. Individual data on hGH treatment history were available. Different mathematical models were used to estimate the statistical distribution of the incubation period. One main feature of the models was to take into account the occurrence of future CJD cases. RESULTS: Models showed that the mean incubation period was 9 to 10 years, and the 95th percentile of the distribution was 15 to 16 years. Data and models indicated that the incubation period was significantly shorter in homozygotes at codon 129 of the prion protein gene than in heterozygotes. CONCLUSIONS: The short mean incubation period of CJD in French hGH recipients may be due to high infectivity in hormone lots. Estimates of the 95th percentile indicate that the number of hGH-related CJD cases may continue to increase in the coming years. PMID- 10522873 TI - Changes in excitability of motor cortical circuitry in primary restless legs syndrome. AB - OBJECTIVE: To investigate the excitability of segmental and suprasegmental systems in patients with primary restless legs syndrome (pRLS) by measuring the cortical silent period (C-SP) and the peripheral silent period (P-SP). BACKGROUND: There is some evidence that RLS may be the motor manifestation of normal CNS periodicity that becomes disinhibited under certain conditions. The mechanism of this disinhibition is unclear. DESIGN/METHODS: Ten patients with pRLS and 10 normal age-matched subjects were studied. The mixed nerve P-SP produced by electrical stimulation of the median and common peroneal nerves was recorded during maximal contraction of the abductor pollicis brevis (APB) and tibialis anterior (TA) muscles. The C-SP produced by a single magnetic shock to motor cortex at 150% of resting threshold was also measured during maximal contraction of the APB and TA muscles. The average of 5 to 10 trials at each site was obtained and compared using Student's t-test. RESULTS: Resting central motor threshold was not significantly different between pRLS patients and the control group. The average duration of the C-SP was shorter in the APB (74.5+/-37.7 versus 129.56+/-35.95 msec, p<0.05) and TA (66.81+/-25.63 versus 136.1+/-40.35 msec, p<0.05) in patients with pRLS. The P-SP duration, however, was not significantly different between two groups in either limb. CONCLUSION: The supraspinal inhibitory system is impaired in pRLS. PMID- 10522874 TI - Clinical and therapeutic observations in aromatic L-amino acid decarboxylase deficiency. AB - OBJECTIVES: To elucidate the phenotype in aromatic L-amino acid decarboxylase (AADC) deficiency, a rare autosomal recessive disorder of neurotransmitter synthesis, and report preliminary treatment observations with directed therapy of the associated neurotransmitter deficiencies. BACKGROUND: AADC is a required enzyme in dopamine, norepinephrine, epinephrine, and serotonin biosynthesis. Five patients have been previously reported. Responses to treatment interventions in these patients have been mixed. METHODS: Clinical and biochemical evaluation and therapeutic trials were performed in two children over a 26-month period. RESULTS: Characteristic features included axial hypotonia, hypokinesia, and athetosis, with superimposed episodes of ocular convergence spasm, oculogyric crises, dystonia, and limb rigidity. Catecholamine deficiency was manifest by ptosis, nasal congestion, paroxysmal diaphoresis, temperature instability, and blood pressure lability. Abnormal sleep, feeding difficulties, and esophageal reflux were typical. Significant therapeutic benefit was observed in one child with a combination of pergolide, trihexyphenidyl, and tranylcypromine. Preliminary trials using serotonin receptor agonists or reuptake inhibitors resulted in adverse effects. CONCLUSIONS: The movement disorder in AADC deficiency, particularly the characteristic eye movement abnormalities, should facilitate the identification of patients with this rare but possibly underrecognized disorder. Directed therapy of the underlying dopamine and norepinephrine deficiency may be beneficial in some cases. PMID- 10522875 TI - FluoroDOPA PET shows the nondopaminergic as well as dopaminergic destinations of levodopa. AB - OBJECTIVE: To evaluate the visible and quantitative anatomic distribution of fluorine-18-labeled L-DOPA in the healthy human brain, to thereby expand the understanding of extrastriatal sites of levodopa function, and to provide a broader foundation for clinical and research studies of fluoroDOPA accumulation in patients. METHODS: The authors performed dynamic three-dimensional fluoroDOPA PET imaging in 10 healthy volunteers and analyzed the images visually and quantitatively. Twenty-eight regions of interest were applied to parametric images of the uptake rate constant (using the multiple-time graphic plot method with cortical input function) and also were used to quantitate regional radioactivity at 80 to 90 minutes. The authors correlated the uptake constants with published human regional neurotransmitter and decarboxylation data. RESULTS: PET imaging with fluoroDOPA demonstrates trapping of labeled dopamine or its metabolites in substantial quantities in many areas of the brain other than the mesostriatal pathways, including considerable uptake in the serotonergic and noradrenergic areas of the hypothalamus and brainstem as well as in extrastriatal cerebral sites. Total fluoroDOPA uptake correlates best with the sum of catecholamine and indolamine concentrations in the brain and moderately well with regional activity of aromatic L-amino acid decarboxylase, but correlates poorly with extrastriatal dopamine concentration. CONCLUSION: Neither L-DOPA nor its radiolabeled analog fluoroDOPA is metabolized or accumulates specifically in dopaminergic or even catecholaminergic neurons. Substantial dopamine production within serotonin and norepinephrine neurons may play a role in either therapeutic effects or adverse effects of therapy with L-DOPA. PMID- 10522876 TI - The tau gene A0 allele and progressive supranuclear palsy. AB - BACKGROUND: Recent studies have shown an association between a polymorphic tandem repeat allele, located in intron 9, of the tau gene and progressive supranuclear palsy (PSP). OBJECTIVE: To investigate this tau polymorphism in individuals with a clinical diagnosis of sporadic or familial PSP as well as in cases confirmed by pathology. METHODS: We analyzed the frequency of tau intronic polymorphism, the presence of linkage in two families with multiple cases of PSP, the splicing of exon 10, and direct sequence of the tau gene. RESULTS: We found that patients with a clinical diagnosis of sporadic or familial PSP and individuals with PSP confirmed by neuropathology have greater prevalence of the A0 allele and A0/A0 genotype than controls. This finding, however, was also true for asymptomatic relatives of individuals with PSP. Linkage analysis in familial PSP excluded the location of the gene in the region 17q21. Furthermore, no significant differences were found in the level of expression of exon 10 in PSP, A0/A0 brain with respect to Alzheimer A3/A3 brain. We found no mutations in the tau gene in individuals with familial PSP. CONCLUSIONS: A mutation in the tau gene was not the primary cause of familial PSP. The role of tau and the tau A0 allele in white PSP patients remains unknown, although it may represent a genetic risk factor for several neurodegenerative disorders. PMID- 10522877 TI - A multicenter, randomized, double-blinded trial of pyridostigmine in postpolio syndrome. AB - BACKGROUND: Postpoliomyelitis syndrome (PPS) is likely due to degeneration and dysfunction of terminal axons of enlarged postpolio motor units. Age-related decline in growth hormone and insulin-like growth factor (IGF-I) may be a contributing factor. Neuromuscular junction abnormalities and decreased IGF-I levels may respond to the anticholinesterase pyridostigmine, with consequent improvement in strength, fatigue, and quality of life. OBJECTIVES: To determine the effect of pyridostigmine in PPS on health-related quality of life, isometric muscle strength, fatigue, and serum IGF-I levels; and to assess the safety of pyridostigmine in PPS. METHODS: The study was a multicenter, randomized, double blinded, placebo-controlled trial of a 6-month course of pyridostigmine 60 mg three times per day in 126 PPS patients. The primary data analysis compared mean changes of outcomes between treatment and control groups at 6 months using an intention to treat approach. Secondary analyses included a comparison of outcomes at 6 and 10 weeks, and in compliant patients. RESULTS: The study showed no significant differences in pyridostigmine and placebo-treated patients with regard to changes in quality of life, isometric strength, fatigue, and IGF-I serum levels at 6 months in the primary analysis and in compliant patients. There were no differences in outcomes at 6 and 10 weeks between groups. However, very weak muscles (1 to 25% predicted normal at baseline) were somewhat stronger (p = 0.10, 95% CI of difference -9.5 to 73.3%), and in compliant patients IGF-I was somewhat increased (p = 0.15, 95% CI of difference -6.4 to 44.8 ng/mL) at 6 months with the medication. Pyridostigmine was generally well tolerated. CONCLUSIONS: This study showed no significant differences between pyridostigmine and placebo-treated PPS patients on measures of quality of life, isometric strength, fatigue, and serum IGF-I. PMID- 10522878 TI - Myasthenia, thymoma, presynaptic antibodies, and a continuum of neuromuscular hyperexcitability. AB - BACKGROUND: Autoantibodies specific for the nicotinic acetylcholine receptor (AChR) of skeletal muscle impair neuromuscular transmission in myasthenia gravis (MG). Autoantibodies specific for alpha3 neuronal AChRs or voltage-gated potassium channels have been reported in patients with Isaacs syndrome, an acquired disorder of continuous muscle fiber activity characterized by neuromyotonia. OBJECTIVE: To report the neuromuscular autoantibody profiles of three patients with a syndrome of MG and neuromuscular hyperexcitability. RESULTS: All three patients reported here had clinical and electrophysiologic evidence of MG and neuromuscular hyperexcitability. None had neuromyotonia. Thymoma was proven in two patients and suspected in the third. One had MG and thymoma and subsequently developed cramp-fasciculation syndrome; MG and rippling muscle syndrome appeared simultaneously in the other two. All patients had muscle and neuronal AChR binding antibodies and striational antibodies. Only one had antibodies reactive with alpha-dendrotoxin-complexed potassium channels. CONCLUSIONS: The coexistence of cramp-fasciculation syndrome and acquired rippling muscle syndrome with MG, thymoma, and neuronal AChR autoantibodies suggests that there is a continuum of autoimmune neuromuscular hyperexcitability disorders related pathogenically to Isaacs syndrome. Manifestations of neuromuscular hyperexcitability may be altered and less apparent in the context of MG because of the coexisting defect of neuromuscular transmission. PMID- 10522879 TI - Mice lacking cytosolic copper/zinc superoxide dismutase display a distinctive motor axonopathy. AB - OBJECTIVE: To characterize the motor neuron dysfunction in two models by performing physiologic and morphometric studies. BACKGROUND: Mutations in the gene encoding cytosolic superoxide dismutase 1 (SOD1) account for 25% of familial ALS (FALS). Transgenes with these mutations produce a pattern of lower motor neuron degeneration similar to that seen in patients with FALS. In contrast, mice lacking SOD1 develop subtle motor symptoms by approximately 6 months of age. METHODS: Physiologic measurements, including motor conduction and motor unit estimation, were analyzed in normal mice, mice bearing the human transgene for FALS (mFALS mice), and knockout mice deficient in SOD1 (SOD1-KO). In addition, morphometric analysis was performed on the spinal cords of SOD1-KO and normal mice. RESULTS: In mFALS mice, the motor unit number in the distal hind limb declined before behavioral abnormalities appeared, and motor unit size increased. Compound motor action potential amplitude and distal motor latency remained normal until later in the disease. In SOD1-KO mice, motor unit numbers were reduced early but declined slowly with age. In contrast with the mFALS mice, SOD1 KO mice demonstrated only a modest increase in motor unit size. Morphometric analysis of the spinal cords from normal and SOD1-KO mice showed no significant differences in the number and size of motor neurons. CONCLUSIONS: The physiologic abnormalities in mFALS mice resemble those in human ALS. SOD1-deficient mice exhibit a qualitatively different pattern of motor unit remodeling that suggests that axonal sprouting and reinnervation of denervated muscle fibers are functionally impaired in the absence of SOD1. PMID- 10522880 TI - Neonatal Guillain-Barre syndrome: blocking antibodies transmitted from mother to child. AB - OBJECTIVE: To investigate the role of blocking antibodies in neonatal Guillain Barre syndrome (GBS) occurring 12 days postpartum in a child born to a mother with ongoing GBS. METHODS: We studied plasma filtrate, purified IgG, and monovalent Fab fragments from the affected mother and serum from the neonate as well as serum samples after recovery from disease 3 months later. Experiments were performed on the hemidiaphragms of adult mice and neonatal and juvenile rats. Quantal endplate currents were recorded with the perfused macro-patch clamp electrode. RESULTS: A dual effect was seen. Serum from mother and infant depressed quantal content by approximately 90% and reduced the amplitude of postsynaptic currents by 30 to 40%. The antibody nature of the blockade could be confirmed by showing that monovalent Fab fragments were similarly effective as purified immunoglobulin (Ig) G. No IgG antibodies to gangliosides, fetal or adult nicotinic acetylcholine receptor, or voltage-gated calcium channels could be detected, but IgM antibodies to the ganglioside GM1 were present. After recovery from GBS no blocking activity was seen in the sera of mother and infant. To elucidate why neonatal disease onset was delayed we examined the possible influence of early developmental changes in functional properties of the neuromuscular junction and applied the mother's active serum to postnatal rats. Although blockade was present in 23-day-old rats, it was absent in 5-day-old rats. CONCLUSION: Transplacentally transferred blocking antibodies may be specifically directed at epitopes of the mature but not the fetal neuromuscular junction. PMID- 10522881 TI - Primary megalencephaly at birth and low intelligence level. AB - OBJECTIVES: To evaluate the association between primary megalencephaly (PMG) at birth and psychosensory conditions and to determine mother-child similarity for PMG at birth. BACKGROUND: PMG is defined as a head circumference (HC) above the 98th percentile that most likely is due to brain enlargement and is not secondary to disease. Previously, PMG in children was reported to be associated with learning disabilities. Contradictory results regarding an association between PMG and intelligence in children exist. Many believe PMG expressed in childhood and adulthood may be inherited. METHODS: Birth records on HC from 144,273 boys, of whom 732 had PMG, were linked to data concerning intelligence level, mental retardation, and impairment of vision and hearing. A potential association between PMG at birth and mental retardation was also examined in 3,204 mentally retarded boys and girls. Parent-offspring similarity for PMG at birth was determined in 13,585 mother-child pairs. RESULTS: PMG was significantly associated with low intelligence level (odds ratio, 1.32; 95% confidence interval [CI], 1.11 to 1.38). The estimated odds ratio for mental retardation in PMG cases versus controls was 1.31 (95% CI, 0.80 to 2.02). There were no differences in frequencies of vision and hearing impairments between PMG cases and controls. Associations between mother's and child's birth weight-normalized HC (r = 0.14; p<0.0001) and PMG (odds ratio, 2.55; CI, 2.00 to 3.25) were found, supporting a multifactorial inheritance of PMG. CONCLUSIONS: PMG at birth is a risk factor for low intelligence level but not for vision and hearing impairments. The heritability of HC and PMG is moderate. PMID- 10522882 TI - Structural and functional brain asymmetries in human situs inversus totalis. AB - OBJECTIVE: To determine whether individuals with situs inversus totalis (SI), a condition in which there is a mirror-image reversal of asymmetric visceral organs, have alterations in brain asymmetries. BACKGROUND: The human brain is asymmetric in structure and function. Although correlations between anatomic asymmetries and functional lateralization in human brain have been demonstrated, it has been difficult to further analyze them. Characterization of asymmetries of brain structure and function in SI might advance the understanding of these relationships. METHODS: Using anatomic and functional MRI techniques, we analyzed asymmetries in the brains of three individuals with SI. RESULTS: Two major anatomic asymmetries of the cerebral hemispheres, the frontal and occipital petalia, were reversed in individuals with SI. In contrast, SI subjects had left cerebral hemisphere language dominance on functional MRI analysis as well as strong right-handedness. CONCLUSION: These observations suggest that the developmental factors determining anatomic asymmetry of the cerebral petalia and viscera are distinct from those producing the functional lateralization of language. PMID- 10522883 TI - Mapping of autosomal dominant progressive external ophthalmoplegia to a 7-cM critical region on 10q24. AB - OBJECTIVE: To map the gene responsible for autosomal dominant progressive external opthalmoplegia. BACKGROUND: The pathogenesis of progressive external ophthalmoplegia (PEO) can be associated with multiple deletions of mitochondrial DNA (mtDNA). PEO may show autosomal dominant (adPEO) or autosomal recessive (arPEO) patterns of inheritance, indicating that the genetic defect has a Mendelian basis and most likely involves a nuclear gene encoding a protein that interacts with the mitochondrial genome. adPEO is heterogeneous genetically, and thus far disease loci have been identified on chromosomes 3 and 10. The locus on chromosome 10q23-q25 was assigned by linkage analysis in a single Finnish family. METHODS: Samples from a large Pakistani family with adPEO, in which clinical symptoms are bilateral ptosis, limitations of eye movements, and varying degrees of proximal muscle weakness, were collected. Muscle biopsy and mtDNA rearrangement analysis was used to confirm the diagnosis. Genomewide linkage analysis was set up using a set of 391 microsatellite markers. RESULTS: The muscle biopsy from an affected member showed ragged red fibers, increased succinic dehydrogenase staining, lack of cytochrome oxidase activity, and multiple deletions of mtDNA. The disease locus was mapped to 10q23.31-q25.1 by linkage analysis, and a maximum lod score of 5.72 was obtained with D10S1267. CONCLUSION: By analysis of meiotic recombinations in affected individuals, the critical region was restricted to the 7-cM interval between D10S198 and D10S1795. PMID- 10522884 TI - Pediatric Chiari I malformations: do clinical and radiologic features correlate? AB - BACKGROUND: Although Chiari I malformation is increasingly recognized in children, little is known about its clinical presentation in this age group. OBJECTIVE: To evaluate the relationship between clinical and MRI features of pediatric Chiari I malformations. METHODS: We performed a chart review and MRI analysis of 49 children with Chiari I malformation. The degree of tonsillar ectopia was compared with age at onset, presence of syringomyelia, and a neurologic severity score as follows: asymptomatic = 0, symptomatic with normal neurologic examination = 1, and symptomatic with abnormal examination = 2. RESULTS: Age at onset of symptoms ranged from 10 months to 14 years. Fifty-seven percent of patients were asymptomatic. Headache and neck pain were the most frequent complaints. Syringomyelia was detected in 14% of patients and skull base abnormalities in 50%. The magnitude of tonsillar ectopia (5 to 23 mm) correlated with severity score (p = 0.04) but not with other clinical measures. CONCLUSIONS: The clinical symptoms of Chiari I malformations in children are nearly identical to those seen in adults. Children with greater amounts of tonsillar ectopia on MRI are more likely to be symptomatic. PMID- 10522885 TI - A study of mortality after temporal lobe epilepsy surgery. AB - OBJECTIVE: To determine early and late mortality in a cohort of 305 consecutive patients who had temporal lobe epilepsy (TLE) surgery over a 20-year period. METHODS: Survival status, cause of death, and postoperative clinical details of those who died were ascertained in a cohort of 305 patients who had TLE surgery. Mortality was related to postoperative seizure status, operative pathology, and side of resection. RESULTS: The survival status of 299 patients was established. Twenty deaths occurred. Mortality was 1 per 136 person-years, with a standardized mortality ratio (SMR) of 4.5 (95% confidence interval [CI], 3.2 to 6.6). Six deaths were sudden and unexpected (SUDEP). The SUDEP rate was 1 per 455 person years. The overall death and SUDEP rates were lower than those reported for similar patient populations with chronic epilepsy. Mortality in patients who had right-sided resections for mesial temporal sclerosis (MTS) remained considerably elevated with a mortality rate of 1 per 54 person-years, an SMR of 32.0 (95% CI, 24.7 to 40.5), and a SUDEP rate of 1 per 134 person-years. These patients had significantly lower seizure remission rates than left-sided patients, but the excess mortality was not simply explained by those patients whose partial seizures were uninfluenced by surgery. Patients who died had more severe or convulsive seizures despite an overall reduction in seizure frequency. CONCLUSIONS: The present findings confirm previous reports that TLE surgery lowers but does not normalize the overall mortality associated with chronic epilepsy. In patients with right-sided MTS, however, the postoperative mortality has remained similar to other groups with medically intractable seizures. PMID- 10522886 TI - Clinical and quantitative pathologic correlates of dementia with Lewy bodies. AB - OBJECTIVES: To examine the distribution of cortical Lewy bodies (LB) and their contribution to the clinical syndrome in dementia with LB (DLB) and to address their relationship to the pathologic markers of AD and PD. METHODS: We studied 25 cases meeting neuropathologic criteria for DLB: 13 cases without AD (Braak stage I or II) and 12 cases with concomitant AD changes (Braak stages III to V). Age at onset, disease duration, and clinical symptoms were reviewed for each case. We quantified the regional distribution of LB in substantia nigra, paralimbic areas (cingulate gyrus, insula, entorhinal cortex, and hippocampus), and neocortex (frontal and occipital association areas) using anti-alpha-synuclein immunostaining. We compared the LB pathology between groups of patients with different symptoms at onset or with specific clinical phenotypes. RESULTS: There were no significant differences in clinical symptoms or LB density between cases with or without concomitant AD. LB density showed a consistent gradient as follows: substantia nigra > entorhinal cortex > cingulate gyrus > insula > frontal cortex > hippocampus > occipital cortex. LB density in substantia nigra and neocortex was not significantly different in cases that started with parkinsonism compared with those that started with dementia. There were no significant differences in LB density in any region among patients with or without cognitive fluctuations, visual hallucinations, delusions, recurrent falls, or parkinsonism. Duration of the disease correlated with a global LB burden for each case (p = 0.02) but did not correlate with LB density in any individual area. Paralimbic and neocortical LB density were highly correlated with each other (p<0.0001), but neither of these correlated well with the number of LB in substantia nigra. LB density did not correlate with Braak stage or frequency of neuritic plaques. CONCLUSIONS: There is a consistent pattern of vulnerability to LB formation across subcortical, paralimbic, and neocortical structures that is similar for DLB cases with or without concomitant AD. Paralimbic and neocortical LB do not correlate with LB in substantia nigra, suggesting that DLB should not be considered just a severe form of PD. LB density correlates weakly with clinical symptoms and disease duration. PMID- 10522887 TI - Accuracy of four clinical diagnostic criteria for the diagnosis of neurodegenerative dementias. AB - OBJECTIVE: To evaluate the inter-rater reliability and validity of clinical diagnostic criteria for neurodegenerative dementias. BACKGROUND: Inter-rater accuracy of the diagnosis of AD has been explored, but there are few accuracy studies for progressive supranuclear palsy (PSP) and frontotemporal lobe dementia (FTD). Furthermore, there have been no simultaneous accuracy studies in a mixed sample of patients with cortical and subcortical neurodegenerative processes. METHODS: Four experienced clinicians reviewed first-visit clinical data abstracted from the records of 40 pathologically diagnosed demented subjects. They were asked to apply the NINCDS-ADRDA criteria for AD, the NINDS-SPSP clinical criteria for PSP, the Lund and Manchester criteria for FTD, and the Consensus Guidelines for the Clinical Diagnosis of Dementia with Lewy Bodies (DLB). RESULTS: The generalized K for AD was 0.73, for PSP 0.82, for FTD 0.75, and for DLB 0.37. The K pool test showed a statistically significant difference between DLB and the other disease processes, and no differences were observed among AD, FTD, and PSP. The mean sensitivity for AD was 95%, for PSP 75%, for FTD 97%, and for DLB 34%. The mean specificity for AD was 79%, for PSP 98.5%, for FTD 97%, and for DLB 94%. CONCLUSIONS: We found improved inter-rater reliability for the diagnosis of AD among clinicians compared with earlier studies. Similarly, there was a near-perfect and substantial inter-rater agreement for the diagnosis of PSP and FTD. The sensitivity for the diagnosis of AD was high, although clinicians overdiagnosed this condition. However, there was a reasonable accuracy for the diagnosis of PSP and FTD. Heterogeneity of the clinical presentation of DLB significantly affected inter-rater agreement and accuracy. The use of multiple diagnostic criteria for cortical and subcortical dementia increases the level of clinical diagnostic accuracy. PMID- 10522888 TI - Brain activation during working memory 1 month after mild traumatic brain injury: a functional MRI study. AB - OBJECTIVE: To assess patterns of regional brain activation in response to varying working memory loads shortly after mild traumatic brain injury (MTBI). BACKGROUND: Many individuals complain of memory difficulty shortly after MTBI. Memory performance in these individuals can be normal despite these complaints. METHODS: Brain activation patterns in response to a working memory task (auditory n-back) were assessed with functional MRI in 12 MTBI patients within 1 month of their injury and in 11 healthy control subjects. RESULTS: Brain activation patterns differed between MTBI patients and control subjects in response to increasing working memory processing loads. Maximum intensity projections of statistical parametric maps in control subjects showed bifrontal and biparietal activation in response to a low processing load, with little additional increase in activation associated with the high load task. MTBI patients showed some activation during the low processing load task but significantly increased activation during the high load condition, particularly in the right parietal and right dorsolateral frontal regions. Task performance did not differ significantly between groups. CONCLUSION: MTBI patients differed from control subjects in activation pattern of working memory circuitry in response to different processing loads, despite similar task performance. This suggests that injury related changes in ability to activate or to modulate working memory processing resources may underlie some of the memory complaints after MTBI. PMID- 10522889 TI - APOE genotype as a risk factor for ischemic cerebrovascular disease: a meta analysis. AB - OBJECTIVE: To determine whether a specific apolipoprotein E (APOE) polymorphism is a risk factor for ischemic cerebrovascular disease (CVD; stroke or TIA). BACKGROUND: The APOE epsilon4 allele is overrepresented in AD, atherosclerosis, and ischemic heart disease. In addition, epsilon4 carriers have higher plasma cholesterol levels than non-epsilon4 carriers. METHODS: Using Medline (OVID and PubMed), a search was performed for all studies that examined APOE in ischemic CVD. The authors identified nine case-control studies that were suitable for analysis. RESULTS: There were 926 patients with ischemic stroke or TIAs and 890 age- and sex-matched control subjects. Overall analysis revealed a significantly higher APOE-epsilon4 allelic frequency in affected patients compared with control subjects (0.14 versus 0.09; odds ratio, 1.68; 95% CI, 1.36 to 2.09; p<0.001). There was a significant excess of the epsilon3 allele (0.85 versus 0.80) but not the epsilon2 allele (0.06 versus 0.06) in the control subjects compared with the ischemic CVD patients. Seven studies had data on APOE genotypes. Carriers of epsilon4 were more frequent among ischemic CVD patients than control subjects (27% versus 18%; odds ratio, 1.73; 95% CI, 1.34 to 2.23; p<0.001). CONCLUSIONS: The APOE-epsilon4 allele and carriers of epsilon4 are more frequent among patients with ischemic CVD compared with control subjects. The epsilon2 allele does not appear to be protective for ischemic CVD. These findings imply a role for the APOE genotype in the pathogenesis of some cases of ischemic CVD. PMID- 10522890 TI - Mechanisms and clinical features of posterior border-zone infarcts. AB - BACKGROUND: Previous studies link posterior border-zone cerebral infarcts between the middle cerebral artery (MCA) and the posterior cerebral artery (PCA) to hemodynamic causes, not embolism. OBJECTIVE: To study the cause of these infarcts. METHODS: We studied 21 patients (unilateral = 18, bilateral = 3) with acute, symptomatic posterior border-zone infarcts shown on CT or MRI to clarify stroke mechanisms. Patients were identified by review of CT and MRI logs and medical records during a 35-month period. An embolic mechanism was assigned when a source of embolism from either the heart, aorta, or parent large artery was present in the absence of intrinsic MCA or PCA disease. A hemodynamic mechanism was assigned when systemic hypotension was present. RESULTS: Among patients with unilateral lesions, 10 were embolic (7 cardiac, 3 carotid), 7 were unknown, and one patient had vasospasm from a ruptured aneurysm. Visual field abnormalities predominated over motor, sensory, and language abnormalities. All patients with bilateral posterior border-zone lesions had perioperative hypotension. Prolonged lethargy, bilateral limb weakness, and cortical blindness were common. CONCLUSIONS: Embolism, either cardiac or from the parent carotid artery, is the predominant stroke mechanism in unilateral posterior border-zone infarcts, not distal field perfusion failure. Bilateral posterior border-zone infarcts have a distinctive clinical presentation and are caused by systemic hypotension. Variability of irrigation of the major arteries, passage of emboli to border-zone areas, and decreased clearance of emboli in these areas explain the findings in the patients with unilateral lesions. PMID- 10522891 TI - Major bleeding during anticoagulation after cerebral ischemia: patterns and risk factors. Stroke Prevention In Reversible Ischemia Trial (SPIRIT). European Atrial Fibrillation Trial (EAFT) study groups. AB - OBJECTIVE: To assess independent predictors of hemorrhage in 651 anticoagulated patients. BACKGROUND: An excess incidence of major bleeding (7% per year) in patients with nondisabling cerebral ischemia of presumed arterial origin treated with oral anticoagulation led to early termination of the Stroke Prevention In Reversible Ischemia Trial (SPIRIT). METHODS: The relationship between known risk factors and hemorrhage was assessed by univariate and multivariate analyses. We compared the risk factors with those in 225 patients anticoagulated because of cerebral ischemia with atrial fibrillation in the European Atrial Fibrillation Trial (EAFT). RESULTS: Leukoaraiosis (hazard ratio [HR] 2.7, 95% confidence interval [CI] 1.4 to 5.3) and age older than 65 years (HR 1.9, 95% CI 1.0 to 3.4) were independent predictors of all anticoagulation-related hemorrhages in SPIRIT. The incidence of intracranial bleeding in SPIRIT was 3.7% per year; this incidence increased by a factor of 1.37 for each 0.5 unit international normalized ratio (INR). Patients with cerebral ischemia of presumed arterial origin had a 19 times (95% CI 2.4 to 150) higher risk of intracranial hemorrhages than those with atrial fibrillation after correcting for baseline differences between SPIRIT and EAFT patients. CONCLUSIONS: In addition to the intensity of anticoagulation, leukoaraiosis and age older than 65 years are independent risk factors for bleeding in patients anticoagulated because of cerebral ischemia of presumed arterial origin. These patients have a higher inherent risk of anticoagulation-related intracranial hemorrhages than patients with atrial fibrillation. PMID- 10522892 TI - Preliminary evidence for anticipation in genetic E200K Creutzfeldt-Jakob disease. AB - Creutzfeldt-Jakob disease (CJD) linked to the E200K mutation of the prion protein (PrP) gene presents within a wide range of phenotypic heterogeneity, including the age at disease onset. We report an earlier disease onset for mutation carriers of the offspring generation when compared with that of their parents, suggesting the possibility of anticipation. A still unidentified environmental or genetic element may affect the age at onset in mutation carriers of different generations. PMID- 10522893 TI - Evidence for the GluR6 gene associated with younger onset age of Huntington's disease. AB - Huntington's disease (HD) is attributed to a triplet CAG repeat mutation, and about half of the variation in onset age can be explained by the size of the repeat expansion. Recently, a TAA repeat polymorphism in close linkage to the kainate receptor, GluR6, was reported related to onset age in HD. We examined this polymorphism in 258 unrelated HD-affected persons (172 from a clinic sample and 86 from a postmortem series). This study confirms that the 155 allele is associated with younger onset age of HD and suggests that it is in linkage disequilibrium with a variant of the GluR6 gene or another gene in this region. PMID- 10522894 TI - Moyamoya disease in Italian monozygotic twins. AB - We report white monozygotic twins with moyamoya disease (MMD) (adult ischemic type). Both had cerebral angiography, MRI, magnetic resonance angiography, SPECT, EEG, human leukocyte antigen (HLA) typing, evaluation of thrombophilia, and immunologic and karyotype analysis. The clinical features and HLA phenotypes described in Asian monozygotic twins with MMD were not found in our patients. However, genetic analysis revealed a homozygous state for C-->T (Ala-->Val substitution) in position 677 of the methylenetetrahydrofolate reductase-encoding gene. PMID- 10522895 TI - An inverse correlation of HTLV-I viral load in CSF and intrathecal synthesis of HTLV-I antibodies in TSP/HAM. AB - The authors measured human T-cell lymphotrophic virus type I (HTLV-I) proviral load and intrathecal synthesis of antibodies to HTLV-I in CSF of 13 Brazilian patients with tropical spastic paraparesis/ HTLV-I-associated myelopathy (HAM). The authors also measured HTLV-I proviral load in peripheral blood mononuclear cells of five of these patients and found that it was 10- to 100-fold higher than that in CSF cells. The combination of HTLV-I proviral load and intrathecal synthesis of antibodies to HTLV-I appears to be a useful marker of disease progression. Patients with high viral load and no intrathecal synthesis of antibodies to HTLV-I had more rapidly progressing, serious clinical disease. PMID- 10522896 TI - Absence of Borrelia burgdorferi-specific immune complexes in chronic fatigue syndrome. AB - Chronic fatigue syndrome (CFS) and Lyme disease often share clinical features, especially fatigue, contributing to concern that Borrelia burgdorferi (Bb), the cause of Lyme disease, may underlie CFS symptoms. We examined 39 CFS patients and 40 healthy controls with a Bb immune complex test. Patients and controls were nonreactive. Centers for Disease Control and Prevention-defined CFS patients lacking antecedent signs of Lyme disease--erythema migrans, Bell's palsy, or large joint arthritis--are not likely to have laboratory evidence of Bb infection. PMID- 10522897 TI - Autopsy patterns for Parkinson's disease and related disorders in Olmsted County, Minnesota. AB - Using a records-linkage system, we determined the frequency and distribution of brain autopsies in residents of Olmsted County, Minnesota, in whom parkinsonism developed during 1976 through 1990. Of the 364 incident cases identified, 235 patients were deceased at the time of record abstraction. The overall autopsy rate was low (23%). Diagnostic certainty (for PD), diagnostic type (PD versus other parkinsonism), sex, age at death, and location at death were important selection factors for autopsy. PMID- 10522898 TI - Corpus callosum measurements in girls with Tourette syndrome. AB - The objective of this study was to examine whether abnormalities in corpus callosum morphology, previously reported in boys with Tourette syndrome (TS), were also present in girls with this disorder. Among the three groups of girls TS, TS and attention deficit hyperactivity disorder, and controls-there were no significant differences in the size of the corpus callosum or any of five individual subregions. The findings suggest that abnormalities in corpus callosum morphology in TS are a gender-related phenomenon. PMID- 10522899 TI - Cortical excitability and recovery curve analysis in generalized epilepsy. AB - Seven untreated patients with idiopathic generalized epilepsy were studied with recovery curve analysis using transcranial magnetic stimulation and compared with 16 controls. Patients had a shorter period of inhibition of the test response following a conditioning stimulus and demonstrated a period of increased facilitation of the test response at interstimulus intervals of 200 to 300 msec, which was similar to the mean interdischarge interval of spike-wave activity on EEG. This provides further evidence of cortical hyperexcitability in idiopathic generalized epilepsy. PMID- 10522900 TI - HLA-DQ1 associated with reflex sympathetic dystrophy. AB - Reflex sympathetic dystrophy (RSD) is a relatively common disabling disorder of unknown pathophysiology. From a cohort of 52 patients, carefully selected to fulfill the recently formulated diagnostic criteria for RSD, venous blood samples were taken for typing of class I and II major histocompatibility antigens. The frequency of HLA-DQ1 was found to be significantly increased compared with control frequencies. The association provides an indication for an organic basis of RSD. PMID- 10522901 TI - Myophosphorylase gene transfer in McArdle's disease myoblasts in vitro. AB - McArdle's disease is due to a genetic deficiency of glycogen phosphorylase and results in a lack of glucose mobilization from glycogen during anaerobic exercise. A genetic defect in Merino sheep produces a similar picture. We constructed a first-generation adenoviral recombinant containing the full-length human phosphorylase cDNA under the control of the Rous sarcoma virus promoter. Primary myoblast cultures from phosphorylase-deficient human and sheep muscle were efficiently transduced with this vector, resulting in restoration of the phosphorylase activity. A similar correction of the genetic defect in muscles of McArdle's patients in vivo appears feasible, preferably with the use of an adeno associated viral vector. PMID- 10522902 TI - Clinical and MRI findings in spinocerebellar ataxia type 5. AB - Spinocerebellar ataxia type 5 (SCA5), one of the genetically heterogeneous autosomal dominant cerebellar ataxias, was assigned to chromosome 11 in a single family descending from the grandparents of President Abraham Lincoln. We report a second, apparently unrelated, SCA5 family of French origin. The overall clinical picture was a slowly progressive cerebellar syndrome beginning mostly in the third decade (27+/-10 years, range 14 to 40). MRI showed a marked global cerebellar atrophy similar to SCA6. PMID- 10522903 TI - A modified Epley's procedure for self-treatment of benign paroxysmal positional vertigo. AB - We compared the efficacy of a modified Epley's procedure (MEP) and Brandt-Daroff exercises (BDE) for self-treatment of benign paroxysmal positional vertigo of the posterior semicircular canal (PC-BPPV) in 54 patients. PC-BPPV resolved within 1 week in 18 of 28 patients (64%) using the MEP and in 6 of 26 patients (23%) performing BDE (p<0.01). Type and adequate performance of the maneuver predicted treatment outcome in the multivariate analysis. The frequency of side effects was not significantly different between the two groups. The MEP is more suitable for self-treatment of PC-BPPV than conventional BDE. PMID- 10522904 TI - Primary association of a TNF gene polymorphism with susceptibility to multiple sclerosis. AB - The associations of three promoter polymorphisms in the tumor necrosis factor (TNFA) gene have been studied in 238 patients and 324 control subjects. A significant correlation was found between MS susceptibility and the TNFA-376 polymorphism. This association was independent of the human leukocyte antigen (HLA) class II association and the combined inheritance of HLA-DRB1*1501 and the TNFA-376A allele more than additively increased susceptibility to MS. PMID- 10522905 TI - Interferon-beta1a in chronic inflammatory demyelinating polyneuropathy. PMID- 10522906 TI - SIADH as the first symptom of Guillain-Barre syndrome. PMID- 10522907 TI - Fulminant neuropathy and lactic acidosis associated with nucleoside analog therapy. PMID- 10522908 TI - Multisystem hypersensitivity reaction to lamotrigine. PMID- 10522909 TI - Absence of human herpesvirus 6 and 7 from spinal fluid and serum of multiple sclerosis patients. PMID- 10522910 TI - Chronic "brain death": meta-analysis and conceptual consequences. PMID- 10522911 TI - Chronic "brain death": meta-analysis and conceptual consequences. PMID- 10522912 TI - Chronic "brain death": meta-analysis and conceptual consequences. PMID- 10522913 TI - Chronic "brain death" meta analysis and conceptual consequences. PMID- 10522914 TI - Posterior leukoencephalopathy without severe hypertension: utility of diffusion weighted MRI. PMID- 10522915 TI - Phenotypic variation in leukoencephalopathy with vanishing white matter. PMID- 10522916 TI - Cerebral dopamine concentrations during levodopa treatment. PMID- 10522917 TI - Pathologic and biochemical studies of juvenile parkinsonism linked to chromosome 6q. PMID- 10522918 TI - Wallerian degeneration of the pyramidal tract does not affect stroke rehabilitation outcome. PMID- 10522919 TI - Esthesioneuroblastoma: reflections of a 21-year experience. AB - OBJECTIVES: To evaluate the results of standardized treatment of esthesioneuroblastoma at a single institution during a 21-year period and calculate pertinent parameters, i.e., metastatic disease (local, regional, distant), disease-free survival, and complications of treatment. STUDY DESIGN: A retrospective review was conducted of all patients treated at a single institution from September 1976 through May 1998. METHODS: Only those patients who received their complete evaluation and treatment at our institution were included in this analysis. Thirty-five patients met this criterion. In addition, results of epidemiological, pathological, and molecular analyses were evaluated to seek accurate indicators for clinical outcomes. RESULTS: Six percent of patients presented with cervical metastatic disease, but ultimately 25.7% developed at least one episode of cervical metastases; 14.3% of patients developed a local recurrence an average of 6 years after diagnosis; and 37% of the patients ultimately developed at least one episode of metastatic disease. The disease-free survival for this cohort of 35 patients was 80.4% at 8 years. CNS complications occurred in 25.7% of the patients, 22.9% had orbital complications, 20% had systemic posttreatment problems, 18.2% had chemotoxic sequelae, 8.6% had infectious complications, and 14.3% had cosmetic sequelae. No epidemiological, pathological, or molecular factors appeared to be more accurate clinical indicators than the Kadish staging system. CONCLUSIONS: This series of esthesioneuroblastoma patients (N=35) reflects an 8-year disease-free survival of 80.4%, representing a significant number of patients treated and followed at one institution for an extended period of time. No valuable pathological or molecular indicators to predict aggressive clinical behavior were found. The average time interval before recurrent disease developed was more than 6 years, far greater than that expected for other sinonasal malignancies. Therefore, extended follow up is necessary for this patient group. PMID- 10522921 TI - Increased expression of keratinocyte growth factor represents a stereotypic response to tracheal lumenal insult independent of injury mechanism. AB - HYPOTHESIS: The author hypothesized that keratinocyte growth factor may play a role in mucosal epithelial repair in large conducting airway, and that the nature of the injury, denudation with disruption of the basement membrane by either mechanical or chemical means, versus exfoliation (loss of suprabasal cells only), would be reflected in the growth factor response. STUDY DESIGN: This study evaluated the interdependent relationship between respiratory epithelial cells and the surrounding extracellular matrix in response to injury. In human cutaneous models, both keratinocyte growth factor and basic fibroblast growth factor are important in repair process. METHOD: A sterile tracheal explant culture system was developed that allows for controlled wounding of the epithelial surface using either 1) NaOH, 2) mechanical debridement, or 3) suprabasal exfoliation. Control and injured bovine tracheas were studied by immunohistochemistry up to 24 hours after injury. RESULTS: Keratinocyte growth factor expression was strongly upregulated in epithelium and submucosa of denuded, but not exfoliation cultures, irrespective of means of injury. Disruption of the basement membrane by any means triggers a stereotypic growth factor response paradigm. This growth factor response has a consistent spatial, but variable, temporal relationship that is influenced by the type of injury mechanical versus chemical. CONCLUSION: This work demonstrates that the type of injury (denudation vs. exfoliation) affects the expression patterns of growth factors that may be important for airways repair. PMID- 10522920 TI - Head and neck cancer in nonsmokers: a distinct clinical and molecular entity. AB - OBJECTIVES/HYPOTHESIS: Clinical and molecular patterns of head and neck squamous cell carcinoma (HNSCC) in nonsmokers and smokers may be different. Analysis of these patterns may improve understanding and management of this disease. STUDY DESIGN: three hundred five subjects were included (46 nonsmokers, 29 former smokers, and 230 smokers). Subsets were analyzed for p53 mutation, human papillomavirus (HPV), and loss of heterozygosity (LOH) at 10 chromosomal loci. METHODS: Clinical information was analyzed for common patterns of disease among the groups. The p53 gene was sequenced, and polymerase chain reaction was used to detect HPV DNA and LOH at two microsatellite markers for each loci. The chi2 test was used to assess differences in genetic alterations between groups, logistic regression to examine the independence of association of tobacco exposure with each alteration, and Kaplan Meier estimates and the log-rank statistic to assess differences in survival. RESULTS: Nonsmokers included a disproportionate number of women, oral cavity (especially tongue) tumors, and very young or old individuals. Smokers had more tumors of the larynx, hypopharynx, and floor of mouth. The rate of p53 mutation was much greater in smokers; the percentage with HPV was marginally lower. The percentage of LOH at 3p, 4q, and 11q13, and the overall average number of chromosomal losses, was significantly lower in nonsmokers' tumors. Survival did not vary with smoking or age. CONCLUSIONS: The clinical and genetic features of HNSCC are distinct between groups defined by tobacco use. Tumors of nonsmokers contain a lower frequency of common genetic alterations, suggesting that the underlying changes in these tumors may remain undiscovered. PMID- 10522922 TI - A method for quantitative assessment of vestibular otopathology. AB - BACKGROUND: Quantitative studies of the vestibular system using serial sections from human temporal bones have been limited because it has been generally difficult to reliably differentiate hair cells from supporting cells and type I from type II hair cells. OBJECTIVES: 1. To develop a new method to overcome the above limitations and permit quantitative assessments of types I and II vestibular hair cells in archival temporal bone sections. 2. To demonstrate that this method is reliable, valid, and repeatable. 3. To describe the advantages of this method compared with other traditional techniques. 4. To discuss the potential of this method to provide new insight into the etiology, pathology, and pathophysiology of vestibular disorders. STUDY DESIGN: Examination of archival human temporal sections prepared for conventional light microscopy. METHODS: The method used Nomarski (differential interference contrast) microscopy to permit visualization of the cuticular plate and stereociliary bundle, to allow unambiguous identification of hair cells. Types I and II hair cells were distinguished by their morphological characteristics. The method was used to measure the density of types I and II hair cells in each vestibular sense organ. Raw-density counts were corrected for potential double counting using Abercrombie's formula. RESULTS: Intrarater and interrater reliability was strong as judged by high Pearson and Spearman correlation values (P < .01). Abercrombie's formula was shown to be valid by comparison with counts made by an unbiased calibration technique using the optical disector principle (correlation coefficients > 0.9, P < .01). CONCLUSIONS: The method described in this report has several advantages when compared with alternative techniques such as surface preparations. The method is applicable to archival bones, permits simultaneous evaluation of the rest of the labyrinth, is relatively inexpensive, and does not preclude other techniques of study (e.g., polymerase chain reaction and immunostaining). Case studies of temporal bones with aminoglycoside ototoxicity and Meniere's disease are used to show how this method has the potential to provide new insight into the pathology and pathophysiology of vestibular disorders. PMID- 10522923 TI - Head and neck computed tomography virtual endoscopy: evaluation of a new imaging technique. AB - OBJECTIVE: To evaluate a new radiographic imaging technique: computed tomography virtual endoscopy (CTVE) for head and neck tumors. STUDY DESIGN: Twenty-one patients presenting with head and neck masses who underwent axial computed tomography (CT) scan with contrast were evaluated by CTVE. Comparisons were made with video-recorded images and operative records to evaluate the potential utility of this new imaging technique. METHODS: Twenty-one patients with aerodigestive head and neck tumors were evaluated by CTVE. One patient had a nasal cylindrical cell papilloma; the remainder, squamous cell carcinomas distributed throughout the upper aerodigestive tract. Patients underwent complete head and neck examination, flexible laryngoscopy, axial CT with contrast, CTVE, and in most cases, operative endoscopy. Available clinical and radiographic evaluations were compared and correlated to CTVE findings. RESULTS: CTVE accurately demonstrated abnormalities caused by intraluminal tumor, but where there was apposition of normal tissue against tumor, inaccurate depictions of surface contour occurred. Contour resolution was limited, and mucosal irregularity could not be defined. There was very good overall correlation between virtual images, flexible laryngoscopic findings, rigid endoscopy, and operative evaluation in cases where oncological resections were performed. CTVE appears to be most accurate in evaluation of subglottic and nasopharyngeal anatomy in our series of patients. CONCLUSION: CTVE is a new radiographic technique that provides surface-contour details. The technique is undergoing rapid technical evolution, and although the image quality is limited in situations where there is apposition of tissue folds, there are a number of potential applications for this new imaging technique. PMID- 10522924 TI - Repair of a rodent nasal critical-size osseous defect with osteoblast augmented collagen gel. AB - OBJECTIVES/HYPOTHESIS: Facial skeletal defects are a common challenge for the otolaryngologist. Type I collagen gels have shown promise in the repair of nonhealing critical size defects (CSDs) of facial bone by providing scaffolding for new bone growth by osteoblasts at the defect perimeter. The objective of the present study was to evaluate the effect that suspending osteoblasts within a type I collagen gel has on the repair of a rodent facial CSD. STUDY DESIGN: Randomized controlled trial using a rodent model. METHODS: A previously described facial CSD was created by removing the nasalis bones with a cutting burr to the level of the nasal mucosal membranes on 18 Sprague-Dawley rats. Groups of six animals were treated with an implant containing either 300 microg of type I collagen gel, 12 x 10(5) osteoblasts suspended within type I collagen gel, or 12 x 10(5) fibroblasts suspended within type I collagen gel for comparison. After 30 days the animals-were examined at necropsy with planimetry, histological analysis of new bone growth, and radiodensitometric analysis of bone thickness. RESULTS: All animals had complete coverage with a thin layer of bone. Histological sectioning revealed an increased thickness in the osteoblast augmented group. Radiodensitometric measurements revealed a statistically significant increase in bone repair in the osteoblast group compared with the collagen-only group (P < or = .0005) and the fibroblast group (P < or = .04). CONCLUSION: Type I collagen gels augmented with an osteoblastic suspension significantly enhance the repair of nasal CSDs in a rodent model. The use of cultured bone precursor cells represents a leap forward in osteoengineering. PMID- 10522925 TI - Repair of an osseous facial critical-size defect using augmented fibrin sealant. AB - OBJECTIVE: Osseous defects of the head and neck are a common challenge for the otolaryngologist. To develop improved reconstructive options, osteoconductive engineering experiments are being conducted. A nasal critical-size defect (CSD) model has previously been described in which less than 7% bone healing is observed over 6 months. An implant containing fibrin sealant with and without osteoprogenitor cells is evaluated in this model. STUDY DESIGN: Randomized controlled trial using a rodent model. METHODS: A nasal CSD was surgically created in 18 male retired breeder Sprague-Dawley rats. Six animals were not implanted with any material, six received fibrin sealant consisting of fibrin (25 mg/mL) and thrombin (1000 U/mL), and six were implanted with fibrin sealant and rat calvarial osteoprogenitor cells (1.8 x 10(6) cells/mL). Thirty days later, the animals were examined at necropsy by planimetry, histological analysis of new bone growth, and radiodensitometric analysis of bone thickness. RESULTS: A thin layer of bone covered the defect in all of the treated animals. A statistically significant increase in bone density (P < .05) between fibrin sealant plus osteoprogenitor cells and each of the other groups was shown using radiodensitometric analysis. Histological analysis also confirmed this difference. CONCLUSION: Osteoprogenitor cells contained within fibrin sealant result in a greater augmentation of bone regeneration than controls or fibrin sealant alone. PMID- 10522926 TI - Laryngeal contact granuloma. AB - OBJECTIVE: To report outcomes of treatment for laryngeal contact granuloma. STUDY DESIGN: Prospective treatment of 21 patients with laryngeal contact granulomas using proton-pump inhibitor (PPI) medication. METHODS: Patients were diagnosed and followed by office endoscopy and patient interview. RESULTS: Three patients did not tolerate PPI medication and were managed by treatment with type 2 histamine (H2) blockers. The lesion completely resolved in 14 of the 18 patients maintained on PPI medication, and significantly regressed in the other 4. Residual granulomas were surgically excised in one patient. Lesions resolved in two patients following injection of botulinum toxin into one thyroarytenoid muscle. One patient had a residual lesion, but symptoms were controlled by medication, and he declined treatment with botulinum toxin. Of the three patients treated with H2-blocker medication, the lesion resolved in only one. CONCLUSION: PPI medication is effective in the treatment of laryngeal contact granuloma, even in the absence of identifiable symptoms of gastroesophageal reflux. PMID- 10522927 TI - Can topical mitomycin prevent laryngotracheal stenosis? AB - OBJECTIVES/HYPOTHESIS: Early topical application of mitomycin to a laryngotracheal lesion may prevent or reduce laryngotracheal stenosis (LTS). STUDY DESIGN: Prospective controlled animal study. METHODS: LTS was induced in 60 dogs randomly assigned to four groups. Controls received an immediate topical application of normal saline. The suction-control group received an immediate application of normal saline followed by suction of secretions on day 2. The mitomycin group received immediate application of 0.7 mL mitomycin (0.2 mg/mL). The repeat-mitomycin group received an immediate application of mitomycin and a second application on day 2, after secretions were suctioned. The laryngeal lumens were measured endoscopically at baseline, day 12, and day 21. Animals were euthanatized if stenosis approximated 95% or at day 21. RESULTS: All dogs in the mitomycin groups survived to day 21, compared with 12 in the suction group and only 2 controls. No side effects of mitomycin were observed. At day 21, surviving controls had 85% and 95% stenosis. In the mitomycin group, median stenosis was 27% (interquartile range, 29% to 42%); in the repeat-mitomycin group, 30% (22% to 40%); and in the suction-control group, 84.5% (72.5% to 93.5%). The mitomycin group differed significantly from controls on day 12 (median difference = 85%, 95% CI = 80%-94%, P < .0001) and day 21 (difference = 63.9%, 95% CI = 58%-85%, P = .031). CONCLUSION: A single topical application of mitomycin significantly reduces the severity of LTS in dogs. Reapplication after 2 days does not improve results. Prospective clinical studies are warranted to assess the efficacy in humans. PMID- 10522928 TI - Endoscopic surgical management of sphenoid sinus disease. AB - OBJECTIVES: To assess the outcome of functional endoscopic sphenoid sinus surgery, and to determine the predictors of outcome. STUDY DESIGN: Retrospective chart review of 651 consecutive endoscopic sinus procedures performed between 1992 and 1997. SETTING: USC University Hospital, University of Southern California, Los Angeles. MATERIALS AND METHODS: Seventy-four patients (11.4% of all endoscopic procedures) with sphenoid sinus disease were selected. All 74 patients were mailed a sinusitis-specific questionnaire, and 46 of them (62.2%) responded. Outcome measures derived from clinician ratings were applied to all 74 patients, and those derived from self-report were applied to 46. Outcome measures were determined from patient questionnaires at a minimum of 6-month postoperative follow-up, operative complications, and clinician perceptual ratings. Patient questionnaires addressed general patient satisfaction, symptom score, and medication usage. A statistical analysis was performed using chi2 test, linear regression, and one-way nonparametric ANOVA. RESULTS: Favorable surgical outcomes based on general patient satisfaction (84.8%, n = 39) and clinician perceptual rating (78.4%, n = 58) were noted. Minor postoperative complications were noted in 10 patients (13.5%) and 8 patients (10.8%) needed revision endoscopic procedures during follow-up. Of the complications, eight (80%) occurred in revision endoscopic procedures. The use of an expanded, sinus-specific symptom score revealed far fewer favorable outcomes (56.5%, n = 26). Seven outcome predictors were established, although none of the predictors held for more than one of the six outcome measures used. CONCLUSION: Endoscopic sphenoid sinus surgery is safe and effective. An expanded symptom score is recommended to assess the outcome of this procedure. PMID- 10522929 TI - Margins of partial cricotracheal resection in children. AB - OBJECTIVE: To review the surgical margins of partial cricotracheal resection in our series of patients. This includes specific anatomic detail as to each superior and inferior resection margin. To apply this information and access the utility of partial cricotracheal resection for the treatment of subglottic stenosis. STUDY DESIGN/METHODS: A retrospective review was performed of 38 children with severe subglottic stenosis who underwent partial cricotracheal resection. Information was obtained with regard to the specific anatomic location of the superior and inferior resection margins, the grade of subglottic stenosis preoperatively, the type of stenting material used postoperatively, and other surgical details specific to each procedure. RESULTS: The superior resection margins were generally to the superior aspect of the cricoid cartilage but as high as the undersurface of the true vocal folds in a minority of patients. Inferior resection margins were generally to the second tracheal ring. Length of resection varied, but was as long as 3.0 cm in one patient. Overall surgical success based on decannulation was > 86%. CONCLUSION: Partial cricotracheal resection is a safe and successful procedure for the treatment of subglottic stenosis. The margins and length of resection should be tailored specifically for each patient; and special considerations must be taken when extensive resection to the level of the true vocal folds is required. Safe airway management in the postoperative period is essential. PMID- 10522930 TI - Surgical management of obstructive sleep apnea in children with cerebral palsy. AB - OBJECTIVES: To evaluate the surgical management of obstructive sleep apnea in children with cerebral palsy. STUDY DESIGN: Retrospective review of 27 children with cerebral palsy who underwent surgical treatment for obstructive sleep apnea. METHODS: Charts were reviewed. Data gathered included primary complaint, coexisting illnesses, initial procedure performed, age at initial surgery, number of days the child was monitored postoperatively in the intensive care unit, notation of postoperative respiratory distress and management, and outcome. RESULTS: Nineteen children underwent adenotonsillectomy for initial treatment of obstructive sleep apnea. Three of these children also had a uvulectomy. Six children had an adenoidectomy alone as their initial procedure. Neither uvulopalatopharyngoplasty nor tracheostomy was performed as an initial procedure. Mean follow-up was 34 months. Seventy-six percent of these children have not required any further surgery. Of the six children who have undergone further surgery, one has required a revision adenoidectomy, and another underwent a tonsillectomy and uvulectomy 2 months after the initial adenoidectomy. Four children ultimately required a tracheotomy. CONCLUSIONS: Eighty-four percent of these children were successfully managed without a tracheotomy. We recommend tonsillectomy and/or adenoidectomy for initial surgical treatment of obstructive sleep apnea in children with cerebral palsy. PMID- 10522931 TI - Management of lateral sinus thrombosis. AB - OBJECTIVES: Review the clinical signs and symptoms, management, bacteriology and outcomes of patients treated for lateral sinus thrombosis. STUDY DESIGN: A retrospective review of six patients, treated from 1993 through 1998, with an intraoperatively confirmed diagnosis of lateral sinus thrombosis. METHODS: All charts from 1993 through 1998 coded for sinus thrombosis, meningitis, brain abscess, otitic hydrocephalus, subdural abscess, and mastoidectomy were reviewed. Operative reports, radiological examinations, laboratory data, culture data and other pertinent data were reviewed. RESULTS: The presenting symptoms ranged from headache to mental status changes. All patients had a history of chronic ear disease and all had at least one additional intracranial complication. The range of additional intracranial complications included otitic hydrocephalus, epidural abscess, and brain abscess. All of the infections were polymicrobial, with a predominance of anaerobes. There were no mortalities; morbidities included anacusis, acute respiratory distress syndrome, reoperation, seizures, septic cardiomyopathy, transfusion, ventriculoperitoneal shunt and nutritional supplementation. CONCLUSION: In patients with otologic disease, complaints of headache, earache or photophobia should warrant an evaluation. The presence of lateral sinus thrombosis mandates further investigation for additional intracranial complications. Conservative surgical intervention, consisting of removal of all perisinus infection and needle aspiration of the sinus, has been found to be effective. Lateral sinus thrombosis is an uncommon complication of otitis media, with potentially significant morbidities, necessitating a high index of suspicion. PMID- 10522932 TI - Ossiculoplasty in young children with the Applebaum incudostapedial joint prosthesis. AB - OBJECTIVE: To evaluate the performance of the Applebaum incudostapedial joint prosthesis in young children in terms of hearing results and long-term stability despite continuing eustachian tube dysfunction and otitis media. STUDY DESIGN: Retrospective review of all Applebaum prostheses placed in children at our institution from June 1993 to June 1998. RESULTS: In 1993 Applebaum proposed the use of a hydroxylapatite ossicular prosthesis as an alternative to incus interposition for the repair of incudostapedial discontinuity. We have used this prosthesis exclusively for the repair of such defects in children over the past 5 years. Among 12 operated ears, all healed, all prostheses remain in place (average duration, 2.6 y), and all children have excellent hearing (mean air-bone gap, 15 dB; range, 5-25 dB). CONCLUSIONS: The Applebaum incudostapedial joint prosthesis restores conductive hearing even in young children. It has been stable in the face of recurrent otitis media and has not interfered with revision surgery. Placement of the prosthesis at primary cholesteatoma surgery should be considered in children. PMID- 10522933 TI - Diagnosis and management of intralabyrinthine schwannomas. AB - OBJECTIVES/HYPOTHESIS: Describe the symptoms, signs, radiographic findings, and treatment results for four patients with intralabyrinthine schwannoma beginning either primarily within the labyrinth or extending secondarily into the labyrinth from the internal auditory canal. STUDY DESIGN: Retrospective review. METHODS: Review of clinic records, operative records, imaging studies with follow-up telephone interview, and when possible, repeat examination. RESULTS: Four patients with intralabyrinthine schwannoma treated by the first author were identified. Episodic vertigo, indistinguishable from Meniere's disease, was present in all but one of the patients in this study. A progressive unilateral hearing loss was also found in all of the patients. Magnetic resonance imaging revealed tumor isolated to the vestibule in two patients with the cochlea primarily involved in the other two patients. Intracochlear tumor extending into the internal auditory canal had been missed on preoperative imaging in one patient and was found during a translabyrinthine vestibular nerve section. In another patient with an intracanalicular schwannoma, tumor extending into the basal turn of the cochlea was not removed during a translabyrinthine approach to the internal auditory canal. The tumor subsequently recurred, necessitating a transotic approach for removal. A transmastoid/translabyrinthine approach was used to successfully remove tumor in one patient. Another patient with good hearing and no vestibular symptoms at time of this writing is being followed with serial imaging studies. As expected, the three patients who underwent surgery have anacusis in the operated ear and are free of vertigo at follow-up intervals of 12, 26, and 65 months. CONCLUSIONS: Intralabyrinthine schwannomas are rare tumors with optimal treatment being determined by the symptoms, tumor location, and hearing. Findings of an intralabyrinthine schwannoma on magnetic resonance imaging may be easily overlooked and attributed to inflammatory changes. PMID- 10522934 TI - Pierre Robin sequences: secondary respiratory difficulties and intrinsic feeding abnormalities. AB - OBJECTIVE: There is considerable variation in opinion regarding the optimal management of patients with Pierre Robin sequence (PRS). No single method of airway intervention or feeding strategy is universally appropriate and effective. This study was performed to examine methods used for airway and feeding management and to identify specific problems encountered. STUDY DESIGN: A retrospective study of 252 patient charts between 1989 and 1997 at Children's Hospital of Wisconsin. METHODS: Patient information was collected regarding perinatal history, genetics evaluation, and airway and feeding evaluations and intervention. A group of 47 patients was determined as having PRS. RESULTS: Secondary respiratory difficulties, defined as respiratory abnormalities in addition to the expected PRS obstruction, were identified in 23% of patients. Also, intrinsic feeding abnormalities not associated with airway obstruction were identified in 11% of patients. Analysis by Fisher's Exact Test revealed patients with a syndromic diagnosis to have a significantly higher rate for tracheotomies and gastrostomy tube placement (P = .041, and P = .0004, respectively). Syndromic patients were also found to have significantly lower Apgar scores and longer hospital stays. Positioning techniques, tongue-lip adhesion, and tracheotomy were also employed effectively with specific indications and specific difficulties that need to be considered. CONCLUSION: Patients with PRS require thorough airway and feeding evaluation. Those with additional syndromic diagnoses demonstrate higher rates of more invasive interventions. Patients with PRS must undergo individualized approaches with consideration of multiple factors for successful management. PMID- 10522935 TI - Recovery of laryngeal sensation after superior laryngeal nerve anastomosis. AB - OBJECTIVES/HYPOTHESIS: Reliable motor reinnervation has been show in multiple laryngeal transplant studies; however, sensory reinnervation of the larynx after nerve anastomosis has yet to be demonstrated. The role of sensory nerve anastomosis in the transplanted larynx in unknown, but is thought to be necessary to provide airway protection. A canine model was developed to examine the possibility of reformation of sensory pathways in the larynx after nerve section and anastomosis. STUDY DESIGN: Randomized controlled experiment. METHODS: Ten canines were randomly assigned to two groups. Hydrochloric acid-induced laryngospasm was demonstrated in every dog. All dogs then had their necks explored, and the internal branch of the superior laryngeal nerve was identified and transected bilaterally. Following nerve section all dogs were retested for an acid-induced laryngospasm reflex. The control group had their wounds closed and were then awakened from anesthesia. The study group underwent microscopic anastomosis of their sensory nerves. Following a 6-month period the two groups of dogs were compared for the presence of the laryngospasm reflex. RESULTS: No dog in the control group had a response to the acid. All dogs in the study group had some response to the acid, although none of them had return of true laryngospasm. CONCLUSION: We concluded that sensory reinnervation does occur after nerve anastomosis, but the recovery of sensation may be incomplete or altered. PMID- 10522936 TI - Diagnostic yield of high-resolution computed tomography for pediatric sensorineural hearing loss. AB - OBJECTIVES/HYPOTHESIS: In recent years, relatively subtle inner ear anomalies have become apparent using high-resolution computed tomography (CT). The purpose of this study was to determine the diagnostic yield of high-resolution CT for pediatric sensorineural hearing loss (HL) (SNHL). METHODS: A review was performed on the records of all children (<18 y of age) who had undergone CT of the temporal bones over a 5-year period, since the introduction of current CT techniques. RESULTS: Three hundred eighty-three studies were performed in 351 subjects. The indication for the CT was SNHL or mixed HL in 157 children. Forty nine (31%) of these studies revealed significant inner ear findings. Large vestibular aqueducts (LVAs) were reported in 15%, commonly in association with cochlear modiolar deficiencies. Modiolar deficiencies (11%) and other cochlear dysplasias (12%) followed LVA in frequency. The incidence of inner ear dysplasia in children with perinatal or postnatal risk factors was only slightly lower than those without (22% vs. 32%, P > .05). The rate of dysplasias did not correlate with SNHL severity, pattern of HL, or type of HL (mixed vs. sensorineural). CONCLUSIONS: These findings suggest that radiographic imaging has a relatively high diagnostic yield in children with SNHL. These findings may be of value in counseling patients and guiding the management of their SNHL. PMID- 10522937 TI - A comparative model: reaction time performance in sleep-disordered breathing versus alcohol-impaired controls. AB - OBJECTIVES/HYPOTHESIS: Patients with sleep-disordered breathing have reaction time deficits that may lead to catastrophic accidents and loss of life. Although safety guidelines do not exist for unsafe levels of sleepiness, they have been established for unsafe levels of alcohol consumption. Since reaction time performance is altered in both, we prospectively used seven measures of reaction time performance as a comparative model in alcohol-challenged normal subjects with corresponding measures in subjects with sleep-disordered breathing. STUDY DESIGN: Institutional Review Board-approved, nonrandomized prospective controlled study. METHODS: Eighty healthy volunteers (29.1+/-7.5 y of age, 56.3% female subjects) performed four reaction time trials using a psychomotor test at baseline and at three subsequent rising alcohol-influenced time points. The same test without alcohol was given to 113 subjects (47.2+/-10.8 y of age, 19.3% female subjects) with mild to moderate sleep-disordered breathing. RESULTS: Mean blood alcohol concentrations (BACs) in the alcohol-influenced subjects at baseline and three trials were 0, 0.057, 0.080, and 0.083 g/dL. The sleep disordered subjects had mean respiratory disturbance indices of 29.2 events per hour of sleep. On all seven reaction time measures, their performance was worse than that of the alcohol subjects when BACs were 0.057 g/dL. For three of the measures, the sleep-disordered subjects performed as poorly as or worse than the alcohol subjects when alcohol levels were 0.080 g/dL. These results could not be explained by sex or age differences. CONCLUSION: The data demonstrate that sleep disordered subjects in this study (with a mean age of 47 y) with mild to moderate sleep-disordered breathing had worse test reaction time performance parameters than healthy, nonsleepy subjects (with a mean age of 29 y) whose BAC is illegally high for driving a commercial motor vehicle in California. This comparative model points out the potential risks of daytime sleepiness in those with sleep disordered breathing relative to a culturally accepted standard of impairment. PMID- 10522938 TI - Morphological assessment of the soft palate in habitual snoring using image analysis. AB - OBJECTIVES: Define differences in palatal and uvular dimensions between habitual snorers and healthy nonsnoring control subjects. Document the changes in palatal configuration after different types of palatoplasty. STUDY DESIGN: A prospective controlled clinical study was performed analyzing video recordings of the soft palate and oropharynx of 251 subjects (121 habitual snorers, 79 patients after laser-assisted uvulopalatoplasty ([LAUP], and 51 healthy volunteers). METHODS: The recordings were captured using a rigid endoscope with a reference measure applied to the soft palate and a mark at the junction of the soft and hard palate. Four parameters were studied in the captured pictures after correction for the distortion deformity in fiberoptic endoscopic images: 1) length of soft palate, 2) length of uvula, 3) width of uvula, and 4) distance between posterior pillars. RESULTS: Analysis showed that habitual snorers, compared with healthy volunteers have significantly increased soft palate length (P = .00001), increased uvula length (P = .0002) and width (P = .00001), and narrowed oropharyngeal isthmus (distance between the posterior pillars) (P = .04). In patients studied after LAUP, the length of the soft palate is significantly shorter (P = .00001) than in the preoperative cohort, and the oropharyngeal isthmus is significantly narrower (P = .00001). Moreover, this latter distance is significantly narrower (P = .00001) when compared with healthy volunteers. CONCLUSIONS: Habitual snorers have a long soft palate, a long wide uvula, and a narrowed oropharyngeal isthmus. LAUP shortens and tightens the elongated palate and causes a further reduction in the space between the posterior pillars. PMID- 10522939 TI - Cochlear fluid space dimensions for six species derived from reconstructions of three-dimensional magnetic resonance images. AB - OBJECTIVES: To establish the dimensions and volumes of the cochlear fluid spaces. STUDY DESIGN: Fluid space volumes, lengths, and cross-sectional areas were derived for the cochleas from six species: human, guinea pig, bat, rat, mouse, and gerbil. METHODS: Three-dimensional reconstructions of the fluid spaces were made from magnetic resonance microscopy (MRM) images. Consecutive serial slices composed of isotropic voxels (25 microm3) representing the entire volume of fixed, isolated cochleas were obtained. The boundaries delineating the fluid spaces, including Reissner's membrane, were resolved for all specimens, except for the human, in which Reissner's membrane was not consistently resolved. Three dimensional reconstructions of the endolymphatic and perilymphatic fluid spaces were generated. Fluid space length and variation of cross-sectional area with distance were derived by an algorithm that followed the midpoint of the space along the length of the spiral. The total volume of each fluid space was derived from a voxel count for each specimen. RESULTS: Length, volume, and cross sectional areas are provided for six species. In all cases, the length of the endolymphatic fluid space was consistently longer than that of either perilymphatic scala, primarily as a result of a greater radius of curvature. For guinea pig specimens, the measured volumes of the fluid spaces were considerably lower than those suggested by previous reports based on histological data. CONCLUSIONS: The quantification of cochlear fluid spaces provided by this study will enable the more accurate calculation of drug and other solute movements in fluids of the inner ear during experimental or clinical manipulations. PMID- 10522940 TI - Aural symptoms and signs of temporomandibular disorder in association with treatment need and visits to a physician. AB - OBJECTIVES: To analyze associations between aural symptoms and clinical signs of and treatment need for temporomandibular disorders, as well as visits to a physician, in a Finnish population. STUDY DESIGN: A longitudinal study of a random sample of adults. METHODS: Four hundred eleven subjects (203 men and 208 women aged 25, 35, 45, 55, or 65 years at baseline) were examined and interviewed at three consecutive examinations at 12-month intervals, and a questionnaire on visits to physician during the preceding 12 months was completed. RESULTS: The aural symptoms were common. The prevalence of otalgia without infection varied between 12% and 16%, while the prevalence of tinnitus and fullness of ears was 12% to 17% and 5% to 9%, respectively. Women had more aural symptoms than men. When compared with the other subjects, the subjects with aural symptoms more often had masticatory muscles that were tender to palpation or temporomandibular joint signs. Subjects with obvious treatment need for temporomandibular disorders had more aural symptoms than subjects in the other treatment need subgroups. They also visited a physician more often because of otalgia than the subjects with otalgia in the other treatment need subgroups. CONCLUSIONS: In patients with otalgia, infectious otolaryngologic diseases should be ruled out. Then the patients without infection should be remitted to a dentist with stomatognathic experience. In the absence of temporomandibular disorders, further medical consultations (e.g., otorhinolaryngological, neurological, physiatric, and psychiatric) are indicated. PMID- 10522941 TI - Effects of depletion of complement in the development of labyrinthitis ossificans. AB - HYPOTHESIS: Labyrinthitis ossificans results in part from the intense inflammatory response to Streptococcus pneumoniae cell wall components. Depletion of complement in Mongolian gerbils following induction of meningitis will reduce the degree of inflammation and subsequent cochlear fibrosis. STUDY DESIGN: Random prospective study. Histological evaluations were performed with the researcher blinded to the experimental group METHODS: S. pneumoniae meningitis was induced in 10 control and 18 experimental Mongolian gerbils with an intrathecal injection of the bacteria. Both groups of animals received treatment with penicillin. The experimental group was also treated with cobra venom factor to deplete complement in the animals. Three months after the induction of meningitis, the animals' temporal bones were harvested for histological evaluation. RESULTS: The decomplemented animals developed significantly less intracochlear fibrosis (P < .01). The mortality rate for the experimental group was 11% compared with 40% in the control group (P = .14). CONCLUSIONS: Reduction of the intense inflammatory response to the S. pneumoniae cell wall components in suppurative labyrinthitis secondary to bacterial meningitis reduced the degree of labyrinthitis ossificans. PMID- 10522942 TI - Histopathological changes of the eustachian tube cartilage and the tensor veli palatini muscle with aging. AB - OBJECTIVES: The eustachian tube (ET) and the tensor veli palatini muscle (TVPM) are thought to play an important role in ventilatory function. Calcification of the ET cartilage and the replacement of TVPM by fat tissue are often observed histologically in elderly patients. To our knowledge, however, there are no quantitative studies of these pathological findings in relation to age. STUDY DESIGN: The calcification of the ET cartilage and the atrophy of the TVPM in 36 normal human temporal bones obtained from 36 individuals with ages ranging from 2 days to 88 years were investigated. METHODS: The number of calcified chondrocytes in the midportion of the ET cartilage was quantified as the average number of cells per square millimeter. Atrophy of the TVPM was evaluated at the midportion of the site where the TVPM is attached to the tip of lateral lamina of ET cartilage. A grade of 0, 1, 2, 3, or 4 was assessed for each section, which indicated approximately 0% to 5%, 5% to 30%, 30% to 70%, 70% to 95%, or 95% to 100% of the TVPM replacement by fat tissue, respectively. RESULTS: A statistically significant correlation was found between the number of the calcified cells and aging (P < .001). A statistically significant correlation was also found between the degree of the atrophy of TVPM and aging (P < .001). CONCLUSIONS: The calcification of the ET cartilage and the atrophy of the TVPM are closely associated with aging. Therefore, it is suggested that these two findings may be a predisposing factor for ET dysfunction in elderly adults. PMID- 10522943 TI - Lamina propria of the mucosa of benign lesions of the vocal folds. AB - OBJECTIVE/HYPOTHESIS: To demonstrate a correlation between the duration and specific pattern of trauma of benign lesions of the vocal folds and their histopathologic appearance. Benign lesions of the vocal folds have various macroscopic appearances. Investigations demonstrate characteristic histopathologic features for three clinically well-defined lesions: 1) vocal fold polyps, 2) Reinke edema, and 3) vocal fold nodules. It is expected that additional histological stainings can contribute to additional insight into the pathophysiology of these lesions. STUDY DESIGN: Retrospective. METHODS: Histological stainings were used to study the constitution of the lamina propria of the mucosa: Verhoeff-van Gieson, Masson trichrome, alcian blue and alcian blue after pretreatment with hyaluronidase. RESULTS: Estimation of the age of a lesion was not possible. Specific observations: 1) accumulation of hyaluronic acid around vessels occurred uniquely in polyps and 2) transverse orientation of elastic fibers was more often seen in vocal fold nodules. Combinations of histopathologic findings were specific to the lesion. CONCLUSIONS: The additional stainings support our previous observations, but had no additional discriminating value in making a histopathologic diagnosis. PMID- 10522944 TI - Ho:YAG laser treatment of hyperplastic inferior nasal turbinates. AB - OBJECTIVES: Although preliminary studies about the successful use of the Ho:YAG laser in nasal turbinate surgery have been reported, no clinical study has been performed on this procedure. The aim of this prospective clinical study was to assess the long-term effect of Ho:YAG laser in the treatment of hyperplastic inferior nasal turbinates. METHODS: Eighty-five patients with nasal obstruction who did not respond to conservative medical treatment were treated with a pulsed Ho:YAG laser (wavelength of =2080 nm). Fifty-two of these patients were included in this clinical study and were followed for 1 year. RESULTS: Within the first 2 weeks, nasal obstruction was correlated to the extent of nasal crusting. Six months after laser treatment, the mucociliary function test showed no variation compared with the preoperative measurements. One year after laser treatment 77% of the patients demonstrated improved nasal airflow on rhinomanometry and questionnaire. CONCLUSIONS: Ho:YAG-laser treatment of hyperplastic turbinates can be performed as outpatient surgery under local anesthesia and offers controllable ablation of soft tissue in a short operation time with satisfactory results and excellent patient acceptance. PMID- 10522945 TI - Eosinophilic cationic protein as a marker of nasal inflammation in patients with cystic fibrosis. AB - OBJECTIVES: Evaluation was made of eosinophilic cationic protein (ECP) in nasal secretion for measuring the degree of nasal inflammation and monitoring response to therapy in cystic fibrosis (CF) patients with chronic rhinosinusitis. Symptoms and findings in regard to ECP levels before and after treatment were described. STUDY DESIGN: Study was prospective, with 21 CF patients aged 4 to 19 years; 20 healthy volunteers served as controls. Collection of nasal secretion by a sponge was performed, and blood samples were obtained for serum. Cystic fibrosis (CF) patients were classified according to nasal symptoms and findings. METHODS: ECP was measured by fluoroimmunoassay. Age, sex, nasal symptoms, and endoscopic and histological findings were obtained, and examinations were conducted before and after treatment; recurrences were recorded. RESULTS: In CF patients with chronic nasal inflammation, increased nasal levels of ECP were detected when compared with asymptomatic CF patients or healthy nonatopic subjects. ECP concentrations were strongly related to the extent of nasal disease; patients with nasal polyps had higher levels than those without. Checked at 1 and 4 months after treatment, ECP levels declined with regression of symptoms, and in patients with exacerbation of nasal disease, ECP levels rose. CONCLUSIONS: According to our study, there is a relationship between levels of ECP in nasal secretions and the degrees of nasal inflammation. In addition, the measurement of ECP could be useful in monitoring nasal disease in CF patients. PMID- 10522946 TI - Abrasive esophageal cytology for the oncological follow-up of patients with head and neck cancer. AB - OBJECTIVES: The occurrence of a second primary cancer in the esophagus in patients with head and neck squamous cell carcinoma is frequent and is associated with a poor prognosis. The aim of this study was to evaluate the yield of abrasive esophageal cytology as a means of screening for metachronous cancer of the upper aerodigestive tract. STUDY DESIGN: We retrospectively reviewed the results of abrasive esophageal cytology performed twice yearly for the screening of patients with prior head and neck cancer. METHODS: From 1987 to 1996, 320 patients treated for head and neck cancer underwent 1,673 abrasive cytology examinations of the esophagus during a mean follow-up period of 4 years. Cytological results were classified as negative, suspect, or positive for malignancy. RESULTS: Twenty-five patients without symptoms had one or more suspect or positive cytologic findings, leading to 29 endoscopic examinations. These revealed 20 premalignant or early malignant lesions of the esophagus (2 dysplasias, 18 squamous cell carcinomas), 2 glandular carcinomas, and 10 clinically unsuspected oral or pharyngeal carcinomas. In seven patients, positive cytological results were associated with clinically visible head and neck cancer. Of the 34 patients with suspect cytological results for malignancy, 10 had no evidence of tumor at endoscopy and 24 had no endoscopic examination because of refusal or because suspected cells were not found in additional examinations. Negative results on cytological examination were found for 254 patients throughout their follow-up, and none of them developed esophageal cancer during a mean follow-up period of 3 years. CONCLUSIONS: For patients with head and neck cancer, abrasive sponge cytology is useful for detecting esophageal cancer at an early stage. In addition, it may reveal unsuspected second primaries or recurrences in the head and neck region. PMID- 10522947 TI - Noninvasive imaging of human oral mucosa in vivo by confocal reflectance microscopy. AB - OBJECTIVES/HYPOTHESIS: To study the microscopic anatomy of normal oral tissues in vivo using confocal reflectance microscopy (CRM). This novel and noninvasive imaging modality can define and characterize healthy oral mucosa and thus this work serves as the foundation for studying oral diseases in vivo. STUDY DESIGN: This was a pilot observational cohort study comparing noninvasive CRM images with histology. MATERIALS AND METHODS: Lip and tongue mucosa were imaged by CRM in six healthy human subjects. In CRM living tissue is illuminated by a laser source and backscattered (or reflected) light is collected by a detector. Image contrast is determined by natural differences in refractive indices of organelles and other subcellular structures within the tissues. Gray-scale images were displayed in real-time on a video monitor and represented horizontal (en face) optical sections through the tissue. Motion of the oral tissue relative to the objective lens was minimized with a tissue stabilizer. After imaging, biopsies were taken from the same site of lip mucosa to correlate noninvasive confocal images with conventional histology. RESULTS: Confocal images correlated well with conventional histology, both qualitatively (visual analysis) and quantitatively (stereology). Imaging was possible up to depths of 490 and 250 microm in the lip and tongue, respectively. Cells and organelles including nuclei, circulating blood cells, and extracellular matrix were clearly observed. CONCLUSION: CRM provides details of normal human oral mucosa at the cellular level without the artifacts of histological processing, and thus has the potential for further development and use in clinical practice as a diagnostic tool for the early detection of oral cancer and precancer. PMID- 10522948 TI - Enhanced cognitive performance with estrogen use in nondemented community dwelling older women. AB - OBJECTIVE: To examine the association between history of postmenopausal estrogen use and cognitive function in a large sample of nondemented community-dwelling older women. SETTING: A community of older residents in Cache County, Utah. PARTICIPANTS: A total of 2338 nondemented women aged 65 and older. MEASUREMENTS: All subjects were administered the Modified Mini-Mental State Examination (3MSE). Self-reported information on current and past use of estrogen after menopause was also obtained using a structured interview. Estrogen use was trichotomized as: no use, past use, and current use. Apolipoprotein E (APOE) genotype was determined and was dichotomized by the presence of an epsilon4 allele. A series of variance/covariance models was conducted with the 3MSE score as the dependent variable, first considering estrogen use alone, then adding, sequentially as covariates, education, age, health status, APOE genotype, current depression status, and history of head injury. RESULTS: In the simplest bivariate model, the 3MSE means (and confidence intervals) were 92.1 (91.7-92.4), 93.5 (93.1-93.9), and 94.4 (94.0-94.7) for never-, past-, and current users, respectively. In the final model (R2 = 0.28), no use of estrogen replacement therapy (P = .006), lower education (P < .001), poorer perceived health status (P = .035), current depression (P = .014), and presence of at least one APOE epsilon4 allele (P < .001) each independently predicted lower 3MSE score. Both current and past estrogen users had significantly higher 3MSE scores than never-users (P = .0063 and P = .0096, respectively). CONCLUSIONS: In this large community study, women who had used estrogen after menopause scored higher on the 3MSE. This finding remained, even after controlling for the effects of age, education, APOE genotype, and other variables that may affect cognition. These data support studies reporting a beneficial role of estrogen on cognition in postmenopausal women, particularly among current estrogen users. PMID- 10522949 TI - Association between bone mineral density and cognitive decline in older women. AB - OBJECTIVE: To test the hypothesis that bone mineral density (BMD), a marker of cumulative estrogen exposure, is associated with cognitive function in nondemented older women. DESIGN: A prospective cohort study. SETTING: Clinical centers in Baltimore, Maryland, Minneapolis, Minnesota, the Monongahela Valley near Pittsburgh, Pennsylvania, and Portland, Oregon. PARTICIPANTS: We evaluated 8333 older community-dwelling women enrolled in the Study of Osteoporotic Fractures who were not taking estrogen replacement. MEASUREMENTS: Calcaneal and hip BMD were measured at baseline and at follow-up (4-6 years later); vertebral fractures were ascertained radiologically at year 6. Women were administered a modified Mini-Mental State Exam, Trails B, and Digit Symbol at baseline and at follow-up. RESULTS: Compared with women with higher bone mineral density, women with low baseline BMD had up to 8% worse baseline cognitive scores (P = .001) and up to 6% worse repeat cognitive scores (P = .001), even after multivariate adjustments. For 1 SD decrease in baseline hip BMD or calcaneal BMD, women had a 32% (95% CI, 19-47%) or a 33% (95% CI, 20-48%) greater odds of cognitive deterioration (worst 10th percentile of change). Women with vertebral fractures had lower cognitive test scores and a greater odds of cognitive deterioration than those without fractures (OR = 1.29; 95% CI, 1.03-1.60). CONCLUSIONS: Women with osteoporosis, whether measured by baseline BMD, reductions in BMD, or vertebral fractures, have poorer cognitive function and greater risk of cognitive deterioration. Our findings suggest a link between two of the most common conditions affecting older women. Further understanding of this association may be important for new treatment and prevention directions. PMID- 10522950 TI - Firearm presence in households of patients with Alzheimer's disease and related dementias. AB - OBJECTIVE: Attention has recently been drawn to the potential dangers of firearm use among patients with dementia. However, little is known about the actual prevalence of firearms in households with demented family members. This study seeks to determine the prevalence and loaded status of firearms in households with a demented family member in a sample of outpatients at a University memory disorders clinic. DESIGN: Utilizing a cross-sectional design, subjects underwent a structured NINCDS-ADRDA criteria comprehensive evaluation to assess dementia and were also administered a questionnaire to assess level of mood disturbance. Family members were administered a behavioral checklist and surveyed about the number and loaded status of firearms in the patient's household. SETTING: The study took place in an outpatient Medical University memory disorders clinic in the Southern United States. PATIENTS: Subjects were 106 consecutive outpatients referred for symptoms suggestive of dementia. MAIN OUTCOME MEASURES: Firearm presence was coded as "present," "not present," and "unsure." In cases where firearms were present, the number and loaded status were collected. Other outcome measures included the Clinical Dementia Rating of each patient, the Yesavage Mood Inventory, and the Revised Memory and Behavior Problems Checklist. RESULTS: A high prevalence of firearm prevalence in households with demented family members was revealed (60.4%). Gun ownership was equally prevalent in households regardless of the severity of the dementia (chi-square, P = .426), severity of behavioral disturbance (ANOVA P = .88), or depressive symptoms (ANOVA P = .37). In households with firearms, 44.6% of the families reported that the guns were kept loaded; 38% reported that they did not know whether the guns were loaded. Only 16.9% of the families reported that guns were maintained in an unloaded state. CONCLUSIONS: This study suggests that many family members living in households in which there are demented patients do not take appropriate action to remove or unload firearms in their households, regardless of the severity of dementia, behavioral disturbance, or depression. These findings suggest that clinicians need to ask families specifically about the presence of firearms and advocate for their removal. PMID- 10522951 TI - Factors contributing to dehydration in nursing homes: inadequate staffing and lack of professional supervision. AB - OBJECTIVE: To investigate the factors that influenced fluid intake among nursing home residents who were not eating well. DESIGN: A prospective, descriptive, anthropological study. SETTING: Two proprietary nursing homes with 105 and 138 beds, respectively. PARTICIPANTS: Forty nursing home residents. MEASUREMENTS: Participant observation, event analysis, bedside dysphagia screening, mental and functional status evaluation, assessment of level of family/advocate involvement, and chart review were used to collect data. Data were gathered on the amount of liquid served and consumed over a 3- day period. Daily fluid intake was compared with three established standards: Standard 1 (30 mL/kg body weight), Standard 2 (1 mL/kcal/energy consumed), and Standard 3 (100 mL/kg for the first 10 kg, 50 mL/kg for the next 10 kg, 15 mL/kg for the remaining kg). RESULTS: The residents' mean fluid intake was inadequate; 39 of the 40 residents consumed less than 1500 mL/day. Using three established standards, we found that the fluid intake was inadequate for nearly all of the residents. The amount of fluid consumed with and between meals was low. Some residents took no fluids for extended periods of time, which resulted in their fluid intake being erratic and inadequate even when it was resumed. Clinical (undiagnosed dysphagia, cognitive and functional impairment, lack of pain management), sociocultural (lack of social support, inability to speak English, and lack of attention to individual beverage preferences), and institutional factors (an inadequate number of knowledgeable staff and lack of supervision of certified nursing assistants by professional staff) contributed to low fluid intake. During the data collection, 25 of the 40 residents had illnesses/conditions that may have been related to dehydration. CONCLUSIONS: When staffing is inadequate and supervision is poor, residents with moderate to severe dysphagia, severe cognitive and functional impairment, aphasia or inability to speak English, and a lack of family or friends to assist them at mealtime are at great risk for dehydration. Adequate fluid intake can be achieved by simple interventions such as offering residents preferred liquids systematically and by having an adequate number of supervised staff help them to drink while properly positioned. PMID- 10522952 TI - Falls relate to vitamin D and parathyroid hormone in an Australian nursing home and hostel. AB - OBJECTIVES: To determine whether falling relates to serum levels of vitamin D and parathyroid hormone. DESIGN: A cross-sectional study with retrospective analysis. SETTING: An aged-care institution in Melbourne Australia. PARTICIPANTS: Ambulant nursing home and hostel residents (n = 83). MEASUREMENTS: Frequency of falling, frequency of going outdoors, use of cane or walker, age, sex, weight, type of accommodation, and duration of residence. Serum concentrations of 25 hydroxyvitamin D, 1,25-dihydroxyvitamin D, and parathyroid hormone (PTH). Plasma concentrations of albumin, calcium, phosphate, and creatinine. Use of furosemide or non-benzodiazepine anticonvulsants. RESULTS: Median age of residents was 84 years. The cohort was vitamin D deficient with a median (interquartile range) 25 hydroxyvitamin D level of 27 (18-37) nmol/L (one-third the reference range median), P < .001. The median (interquartile range) PTH of 5.2 (3.8-7.7) pmol/L exceeded the reference range median, P < .001. Residents who fell (n = 33) had lower serum 25-hydroxyvitamin D levels than other residents (medians 22 vs 29 nmol/L, P = .02) and higher serum PTH levels (medians 6.2 vs 4.8 pmol/L, P < .01). Sixty residents lived in the hostel (72%), and 41 (49%) walked without any walking aid. In a multiple logistic regression for falling, higher serum PTH remained independently associated with falling, with an odds ratio (95% confidence interval) for falling of 5.6 (1.7-18.5) per unit of the natural logarithm of serum PTH. Other terms in the regression were hostel accommodation, odds ratio .04 (.01-.25), and ability to walk without aids, odds ratio .07 (.01 .37). CONCLUSIONS: In ambulant nursing home and hostel residents, residents who fall have lower serum 25-hydroxyvitamin D and higher serum parathyroid hormone levels than other residents. The association between falling and serum PTH persists after adjustment for other variables. PMID- 10522953 TI - Restraint reduction reduces serious injuries among nursing home residents. AB - OBJECTIVES: To describe how removing physical restraints affected injuries in nursing home settings. DESIGN: A 2-year prospective study of an educational intervention for physical restraint reduction. SETTING: Sixteen diverse nursing homes with 2075 beds in California, Michigan, New York, and North Carolina. PARTICIPANTS: Study A: 859 residents who were physically restrained at the onset of the intervention on October 1, 1991. Study B: all residents who occupied the 2075 beds in the 16 facilities 3 months before the intervention and 3 months after its completion. INTERVENTION: Educational program for nursing home staff followed by quarterly site consultations to participating nursing homes. MAIN OUTCOME MEASURES: Rate of physical restraint use and injuries. RESULTS: Study A: Serious injuries declined significantly among the 859 residents restrained initially when restraint orders were discontinued (X2 = 6.2, P = .013). Study B: During the intervention period, physical restraint use among the 2075 residents decreased from 41% to 4%, a 90% reduction. The decrease in the percentage of injuries of moderate to serious severity was significant (i.e., 7.5% vs 4.4%, P2 tail = .0004) as was the rate of moderate and serious injuries combined (Rate Ratio = 1.580, P2-tail = .0033). CONCLUSIONS: A substantial decrease in restraint use occurred without an increase in serious injuries. Although minor injuries and falls increased, restraint-free care is safe when a comprehensive assessment is done and restraint alternatives are used. PMID- 10522954 TI - Once-weekly resistance exercise improves muscle strength and neuromuscular performance in older adults. AB - OBJECTIVE: To determine the effect of frequency of resistive training on gain in muscle strength and neuromuscular performance in healthy older adults. DESIGN: A randomized controlled trial with subjects assigned either to high-intensity resistance training 1 (EX1), 2 (EX2), or 3 (EX3) days per week for 24 weeks or to a control group (CO). SETTING: An exercise facility at an academic medical center. SUBJECTS: Forty-six community-dwelling healthy men (n = 29) and women (n = 17) aged 65 to 79 years. INTERVENTION: Progressive resistance training consisting of three sets of eight exercises targeting major muscle groups of the upper and lower body, at 80% of one-repetition maximum (1-RM) for eight repetitions, either 1, 2, or 3 days per week. MEASURES: Dynamic muscle strength (1-RM) using isotonic equipment every 4 weeks, bone mineral density and body composition by dual energy X-ray absorptiometry (DXA), and neuromuscular performance by timed chair rise and 6-meter backward tandem walk. RESULTS: For each of the eight exercises, muscle strength increased in the exercise groups relative to CO (P < .01), with no difference among EX1, EX2 and EX3 groups at any measurement interval. Percent change averaged 3.9 +/- 2.4 (CO), 37.0 +/- 15.2 (EX1), 41.9 +/- 18.2 (EX2), and 39.7 +/- 9.8 (EX3). The time to rise successfully from the chair 5 times decreased significantly (P < .01) at 24 weeks, whereas improvement in the 6-meter backward tandem walk approached significance (P = .10) in the three exercise groups compared with CO. Changes in chair rise ability were correlated to percent changes in quadriceps strength (r = -0.40, P < .01) and lean mass (r = -0.40, P < .01). CONCLUSIONS: A program of once or twice weekly resistance exercise achieves muscle strength gains similar to 3 days per week training in older adults and is associated with improved neuromuscular performance. Such improvement could potentially reduce the risk of falls and fracture in older adults. PMID- 10522955 TI - Strength training normalizes resting blood pressure in 65- to 73-year-old men and women with high normal blood pressure. AB - OBJECTIVE: To determine the effects of heavy resistance strength training (ST) on resting blood pressure (BP) in older men and women. DESIGN: Prospective intervention study. SETTING: University of Maryland Exercise Science Laboratory. PARTICIPANTS: Twenty-one sedentary, healthy older men (69 +/- 1 year, n = 11) and women (68 +/- 1 year, n = 10) served as subjects for the study. INTERVENTION: Six months of progressive whole body ST performed 3 days per week using Keiser K-300 air-powered resistance machines. MEASUREMENTS: One-repetition maximum (1 RM) strength was measured for seven different exercises before and after the ST program. Resting BP was measured on six separate occasions before and after ST for each subject. RESULTS: Substantial increases in 1 RM strength were observed for upper body (UB) and lower body (LB) muscle groups for men (UB: 215 vs 265 kg; LB: 694 vs 838 kg; P < .001) and women (UB: 128 vs 154 kg; LB: 441 vs 563 kg; P < .001). The ST program led to reductions in both systolic (131 +/- 2 vs 126 +/- 2 mm Hg, P < .010) and diastolic (79 +/- 2 vs 75 +/- 1 mm Hg, P < .010) BP. Systolic BP was reduced significantly in men (134 +/- 3 vs 127 +/- 2 mm Hg, P < .01) but not in women (128 +/- 3 vs 125 +/- 3 mm Hg, P < .01), whereas diastolic BP was reduced following training in both men (81 +/- 3 vs 77 +/- 1, mm Hg, P = .054) and women (78 +/- 2 vs 74 +/- 2 mm Hg, P = .055). CONCLUSIONS: Six months of heavy resistance ST may reduce resting BP in older persons. According to the latest guidelines from the Joint National Committee for the Detection, Evaluation, and Treatment of Hypertension, the changes in resting BP noted in the present study represent a shift from the high normal to the normal category. PMID- 10522956 TI - Delayed thrombolytic treatment of older patients with acute myocardial infarction. AB - OBJECTIVE: To determine demographic and clinical factors associated with delayed thrombolysis in patients with acute myocardial infarction. DESIGN: A retrospective cohort. SETTING: 37 Minnesota hospitals during the time periods October 1992-July 1993 and July 1995-April 1996. PATIENTS: We reviewed the medical records of 776 older patients aged 65 or older hospitalized with an admission diagnosis of acute myocardial infarction, suspected acute myocardial infarction, or rule-out acute myocardial infarction, who were treated with a thrombolytic agent. MEASUREMENT: We used multivariate logistic regression models to examine the association between selected study characteristics and time between hospital presentation and administration of thrombolytic treatment. Early thrombolysis was defined as less than 60 minutes after hospital presentation and late thrombolysis as 60+ minutes. RESULTS: Of 776 study patients, 57.5% (n = 446) received early thrombolysis. Of the remaining 330 patients receiving late treatment, 12.1% (n = 94) were thrombolyzed more than 2 hours after hospital presentation. After controlling for other factors, the odds of delayed thrombolysis among patients aged 75 or older were 1.48 compared with younger individuals (95% CI, 1.17-1.88). The odds of delayed thrombolysis among patients with severe comorbidity were 1.46 (95% CI, 1.10-1.94) compared with individuals without severe comorbidity. Predictors of early thrombolytic treatment included hospital arrival via emergency transport (ORdelay = 0.46; 95% CI, 0.34-0.63) and chest discomfort at admission (ORdelay = 0.40; 95% CI, 0.18-0.86). CONCLUSIONS: The present study indicates that patients of advanced age and with severe comorbidity are more likely to experience delayed thrombolytic treatment after hospital presentation. These are the patients who suffer the highest morbidity from acute myocardial infarction and for whom expeditious treatment may enhance therapeutic benefit. PMID- 10522957 TI - Detection of older people at increased risk of adverse health outcomes after an emergency visit: the ISAR screening tool. AB - OBJECTIVES: To develop a self-report screening tool to identify older people in the emergency department (ED) of a hospital at increased risk of adverse health outcomes, including: death, admission to a nursing home or long-term hospitalization, or a clinically significant decrease in functional status. DESIGN: Prospective (6-month) follow-up study of a cohort of ED patients aged 65 and older. SETTING: The EDs of four acute-care hospitals in Montreal, Quebec, Canada. PARTICIPANTS: Community-dwelling patients aged 65 and older who came to the EDs during the weekday shift over a 3-month recruitment period. Patients were excluded if they could not be interviewed either because of their medical condition or because of cognitive impairment and no other informant was available. MEASUREMENTS: Measures ascertained at the ED visit included: 27 self report screening questions on social, physical, and mental risk factors; medical history; use of hospital services, medications, and alcohol; and the Older American Resources and Services (OARS) activities of daily living (ADL) scale. At follow-up, the OARS scale was readministered by telephone, and other adverse health outcomes were ascertained. RESULTS: Among 1673 patients who completed the follow-up measures, 488 (29.2%) had an adverse health outcome. Scale development and selection methods included logistic regression, receiver operating characteristic curves, and expert judgment. The proposed screening tool (ISAR) comprises six self-report questions on functional dependence (premorbid and acute change), recent hospitalization, impaired memory and vision, and polymedication. The tool performed well in the total cohort aged 65 and older, and in sub-groups defined by disposition (admitted or released from ED), language of questionnaire administration (French or English), information source (patient or other), and other characteristics. CONCLUSIONS: The ISAR is a short self-report questionnaire that can quickly identify older patients in the ED at increased risk of several adverse health outcomes and those with current disability. PMID- 10522958 TI - Thyroid function, morphology and prevalence of thyroid disease in a population based study of Danish centenarians. AB - OBJECTIVES: To investigate thyroid function, morphology, and autoimmunity in relation to physical function in an unselected population of centenarians. DESIGN: A population-based survey. SETTING: Denmark. PARTICIPANTS: All persons living in Denmark who celebrated their 100th anniversary during the period April 1, 1995 to May 31, 1996, a total of 276 persons. MAIN OUTCOME MEASUREMENTS: Thyroid hormones (TSH, T4, FT4I, T3, FT3I, and T3RU), thyroid autoantibodies (TPOab and Tgab), thyroid volume, activities of daily living according to the Katz Index of ADL. RESULTS: In all, 207 (75%) of the 276 eligible subjects participated, and 148 agreed to blood tests. Among the participants, 2.9% had previously known hyperthyroidism, and the same proportion had previously known hypothyroidism. The blood tests did not reveal any undiagnosed cases of overt thyroid dysfunction. However 7.2% had a subnormal serum TSH, and 2.9% had an elevated serum TSH; all had normal serum T3 and serum T4 levels. Thyroid autoantibodies were detected in 26 (17.6%) centenarians (11.5% had Tgab and 9.5% had TPOab). Among relatively independent centenarians, low serum T3 was significantly associated with high comorbidity (P = .029), whereas both low serum T3 and thyroid autoantibodies were significantly associated with ADL-dependency (P < .001 and P = .030, respectively). Ultrasonography (n = 50) revealed a small gland with a median volume of 8.3 mL (range 3.2-27.9) compared with an expected volume of 20 mL (14-26) (P < .001). There was no significant relationship to body weight. When examined by ultrasound, only 26% had significant morphological alterations. CONCLUSIONS: Thyroid dysfunction does not seem to be more prevalent among centenarians than among younger old people. Low serum T3 is related to poor physical function and co-morbidity, whereas thyroid autoimmunity is related only to poor physical functioning. Despite atrophy of the thyroid gland, these findings suggest that thyroid function is well preserved in centenarians. PMID- 10522959 TI - An in-home nurse-administered geriatric assessment for hypoalbuminemic older persons: development and preliminary experience. AB - BACKGROUND: Although malnutrition in older persons is a common, potentially treatable condition, few data indicate that treatments for this disorder can be effective. OBJECTIVE: To develop and preliminarily evaluate a two-component intervention that includes a nurse-administered, in-home assessment to identify potentially remediable causes of hypoalbuminemia and protocols to treat these problems. DESIGN: A pre-test post-test case-series. SETTING: An academic geriatrics practice. PARTICIPANTS: Seventeen persons aged 65 and older with serum albumin levels < or = 3.8 g/dL; eight of the participants received pre-and post test outcome measures. INTERVENTION: Nurse-administered standardized assessment and intervention protocols. MEASUREMENTS: Serum albumin, Medical Outcome Study (MOS) SF-36, serum IL-1a and b, TNF alpha, IL-6, and lymphocyte markers of immune function. RESULTS: The assessment took 87 minutes, on average, and generated a mean 4.2 recommendations. Among the eight subjects with pre- and post-test measures, serum albumin increased by 0.2 g/dL (P = .035). Compared with baseline, two T cell markers of immune function demonstrated changes consistent with better function. CONCLUSIONS: These preliminary data support the potential benefit of a nurse-administered assessment coupled with protocols to address remediable contributors to hypoalbuminemia. PMID- 10522960 TI - Predictors of five-year mortality in older Canadians: the Canadian Study of Health and Aging. AB - OBJECTIVE: Based on the Canadian Study of Health and Aging (CSHA), to determine the importance of cognitive status, sociodemographic factors, functional status, and other health related factors as predictors of 5-year overall mortality in older Canadians. DESIGN, SETTING AND PARTICIPANTS: Two partially overlapping groups from the Canadian Study of Health and Aging (1991) were identified: (1) older people living in the community (n = 8949) who had a screening interview (larger sample, fewer variables) and (2) older people who underwent an extensive clinical examination (smaller sample, more objective variables; n = 2914). Deaths in the subsequent 5 years were determined from death certificates and interviews with the caregivers. Multivariate logistic regression models, with death within 5 years as the outcome, were developed separately for men and women. Predictor variables were introduced in the following groups: sociodemographic factors, physical and cognitive status, and physical illnesses and life style factors. Parallel models were developed for the screening sample and for the clinical sample. RESULTS AND DISCUSSION: Five-year mortality ranged from 10.0% (women aged 65-74 living in the community) to 88.1% (men aged 85 and older living in institutions). Multivariate models showed that the odds of death within 5 years increased with age. This effect remained after adjustment for all other variables. Odds of death increased with institutionalization and with increasing cognitive and physical impairment. Although vision and hearing problems and the presence of heart disease, stroke, and diabetes were all strongly related to 5 year mortality in univariate, unadjusted analyses, their contributions were minimal in the multivariate analyses. Increased Body Mass Index was associated with lower mortality in both univariate and multivariate analyses. CONCLUSIONS: This population-based study supported the importance of gender, age, functional status, cognition, and health status in predicting 5-year mortality, and after accounting for cognitive status, physical status, and specific disease variables, the difference in mortality between older people in the community and in institutions was reduced. Knowledge about survival and prognosis is important not only for the planning of long-term facilities and home care, but it can also be helpful for clinical decision-making and for family and caregivers. PMID- 10522961 TI - Prevalence of coexistence of coronary artery disease, ischemic stroke, and peripheral arterial disease in older persons, mean age 80 years, in an academic hospital-based geriatrics practice. AB - OBJECTIVE: To investigate the prevalence of coronary artery disease (CAD), ischemic stroke, and peripheral arterial disease (PAD), alone and in combination, in older persons. DESIGN: A retrospective analysis of charts from all older persons seen from April 1, 1998, through December 31, 1998, at an academic hospital-based geriatrics practice. SETTING: An academic hospital-based geriatrics practice staffed by fellows in a geriatrics training program and full time faculty geriatricians. PATIENTS: A total of 474 men and 1328 women, mean age 80 +/- 9 years (range 60 to 102 years) were included in the study. MEASUREMENTS AND MAIN RESULTS: Of 1802 persons studied, 612 (34%) had CAD, 351 (19%) had ischemic stroke, 236 (13%) had PAD, and 816 (45%) had either CAD, stroke, or PAD. Three hundred twenty-eight (18%) of the 1802 persons had CAD alone, 128 (7%) had stroke alone, 50 (3%) had PAD alone, 123 (7%) had CAD + stroke and no PAD, 86 (5%) had CAD + PAD and no stroke, 25 (1%) had PAD + stroke and no CAD, 75 (4%) had CAD + stroke + PAD, and 986 (55%) had no CAD, PAD, or stroke. If CAD was present, coexistent PAD was present in 26% and coexistent stroke in 32% of persons studied. If stroke was present, coexistent CAD was present in 56% and coexistent PAD in 28%. If PAD was present, coexistent CAD was present in 68% and coexistent stroke in 42% of persons studied. CONCLUSIONS: These data showed that if CAD was present, ischemic stroke was also present in 32% and PAD in 26% of the population. If ischemic stroke was present, CAD was also present in 56% and PAD in 28% of the population. If PAD was present, CAD was also present in 68% and ischemic stroke in 42% of the population. PMID- 10522962 TI - Walking speed and stride length predicts 36 months dependency, mortality, and institutionalization in Chinese aged 70 and older. AB - BACKGROUND: Increasing emphasis is being placed on physical performance measures as an outcome predictor. It is uncertain whether one or two simple measurements will have predictive value compared with a battery of tests. OBJECTIVES: To assess whether simple performance measures such as walking speed and stride length will predict dependency, mortality, and institutionalization. DESIGN: A 3 year longitudinal study of a random sample of subjects. SETTING: Older people living in the community in Hong Kong, Special Administrative Region, China. SUBJECTS: A total of 2032 Chinese subjects aged 70 years and older were recruited territory-wide by proportional random sampling and followed for 3 years. MEASUREMENTS: Functional status was measured using the Barthel Index at baseline and follow-up. The time taken to walk a distance of 16 feet and the number of steps taken were measured at baseline. Stride length is estimated by dividing 16 by the average number of steps needed to complete the walk. Outcomes regarding dependency, mortality, and institutionalization at 36 months were recorded. RESULTS: After excluding subjects lost to follow-up and those who had died, data were available for 559 men and 612 women. Univariate analysis showed that reduced walking speed and stride length were associated with increased risk of dependency, mortality, and institutionalization. In multivariate analysis for dependency and mortality, stride length, walking speed, age, and sex were included in the best prediction model (ROC = 0.798 and 0.707, respectively), whereas only stride length was included in the prediction for institutionalization (ROC = 0.764). CONCLUSIONS: In terms of prevention or modifying outcomes, these two simple performance measures may be used as indicators for checking for occult disease and for interventional measures such as exercise prescription. PMID- 10522963 TI - Attempting resuscitation in nursing homes: policy considerations. PMID- 10522964 TI - Can falls and hip fracture be prevented in frail older adults? PMID- 10522965 TI - Cognitive performance and estrogen use in nondemented older women. PMID- 10522966 TI - Fluid deprivation and research ethics. PMID- 10522967 TI - Functional and demographic predictors of health and human services utilization: a community-based study. PMID- 10522968 TI - Obstructive sleep apnea syndrome may be a significant cause of gastroesophageal reflux disease in older people. PMID- 10522969 TI - Sertraline-induced hyponatremia in an older patient. PMID- 10522970 TI - Age effect on pupil dilation among Alzheimer's patients. PMID- 10522971 TI - Medicare expenditures for beneficiaries with dementia of the Alzheimer's type. PMID- 10522972 TI - Clinical review 110: Cardiovascular actions of estrogens in men. PMID- 10522973 TI - Alzheimer's disease in males: endocrine issues and prospects. PMID- 10522974 TI - Anabolic interventions for aging-associated sarcopenia. PMID- 10522975 TI - Osteoporosis in men. PMID- 10522976 TI - Male pseudohermaphroditism caused by mutations of the human androgen receptor. PMID- 10522977 TI - Infertility in men: recent advances and continuing controversies. PMID- 10522978 TI - Hypertension in men. PMID- 10522979 TI - Gender assignment and reassignment in 46,XY pseudohermaphroditism and related conditions. PMID- 10522980 TI - Selective androgen receptor modulators (SARMs): a novel approach to androgen therapy for the new millennium. PMID- 10522981 TI - Hormonal signaling in prostatic hyperplasia and neoplasia. PMID- 10522982 TI - Pharmacokinetics, efficacy, and safety of a permeation-enhanced testosterone transdermal system in comparison with bi-weekly injections of testosterone enanthate for the treatment of hypogonadal men. AB - The pharmacokinetics, efficacy, and safety of the Androderm testosterone (T) transdermal system (TTD) and intramuscular T enanthate injections (i.m.) for the treatment of male hypogonadism were compared in a 24-week multicenter, randomized, parallel-group study. Sixty-six adult hypogonadal men (22-65 years of age) were withdrawn from prior i.m. treatment for 4-6 weeks and then randomly assigned to treatment with TTD (two 2.5-mg systems applied nightly) or i.m. (200 mg injected every 2 weeks); there were 33 patients per group. Twenty-six patients in the TTD group and 32 in the i.m. group completed the study. TTD treatment produced circadian variations in the levels of total T, bioavailable T, dihydrotestosterone, and estradiol within the normal physiological ranges. i.m. treatment produced supraphysiological levels of T, bioavailable T, and estradiol (but not dihydrotestosterone) for several days after each injection. Mean morning sex hormone levels were within the normal range in greater proportions of TTD patients (range, 77-100%) than i.m. patients (range, 19-84%). Both treatments normalized LH levels in approximately 50% of patients with primary hypogonadism; however, LH levels were suppressed to the subnormal range in 31% of i.m. patients vs. 0% of TTD patients. Both treatments maintained sexual function (assessed by questionnaire and Rigiscan) and mood (Beck Depression Inventory) at the prior treatment levels. Prostate-specific antigen levels, prostate volumes, and lipid and serum chemistry parameters were comparable in both treatment groups. Transient skin irritation from the patches was reported by 60% of the TTD patients, but caused only three patients (9%) to discontinue treatment. i.m. treatment produced local reactions in 33% of patients and was associated with significantly more abnormal hematocrit elevations (43.8% of patients) compared with TTD treatment (15.4% of patients). Gynecomastia resolved more frequently during TTD treatment (4 of 10 patients) than with i.m. treatment (1 of 9 patients). Although both treatments seem to be efficacious for replacing T in hypogonadal men, the more physiological sex hormone levels and profiles associated with TTD may offer possible advantages over i.m. in minimizing excessive stimulation of erythropoiesis, preventing/ameliorating gynecomastia, and not over-suppressing gonadotropins. PMID- 10522983 TI - Modest hormonal effects of soy isoflavones in postmenopausal women. AB - Soy isoflavones have been hypothesized to exert hormonal effects in postmenopausal women. To test this hypothesis, we studied the effects of three soy powders containing different levels of isoflavones in 18 postmenopausal women. Isoflavones were consumed relative to bodyweight [control: 0.11 +/- 0.01; low isoflavone (low-iso): 1.00 +/- 0.01; high isoflavone (high-iso): 2.00 +/- 0.02 mg/kg/day] for 93 days each in a randomized crossover design. Blood was collected on day 1 of the study (baseline) and days 36-38, 64-66, and 92-94 of each diet period, for analysis of estrogens, androgens, gonadotropins, sex hormone binding globulin (SHBG), prolactin, insulin, cortisol, and thyroid hormones. Vaginal cytology specimens were obtained at baseline and at the end of each diet period, and endometrial biopsies were performed at baseline and at the end of the high-iso diet period, to provide additional measures of estrogen action. Overall, compared with the control diet, the effects of the low-iso and high-iso diets were modest in degree. The high-iso diet resulted in a small but significant decrease in estrone-sulfate (E1-S), a trend toward lower estradiol (E2) and estrone (E1), and a small but significant increase in SHBG. For the other hormones, the few significant changes noted were also small and probably not of physiological importance. There were no significant effects of the low-iso or high-iso diets on vaginal cytology or endometrial biopsy results. These data suggest that effects of isoflavones on plasma hormones per se are not significant mechanisms by which soy consumption may exert estrogen-like effects in postmenopausal women. These data also show that neither isoflavones nor soy exert clinically important estrogenic effects on vaginal epithelium or endometrium. PMID- 10522984 TI - Relationship between insulin resistance, soluble adhesion molecules, and mononuclear cell binding in healthy volunteers. AB - The relationship between insulin resistance, soluble adhesion molecules E selectin (sE-selectin), intracellular adhesion molecule-1 (sICAM-1), and vascular adhesion molecule-1 (sVCAM-1), mononuclear cell binding to cultured endothelium, and lipoprotein concentrations were evaluated in 28 healthy, nondiabetic, and normotensive individuals. The mean (+/-SEM) lipid and lipoprotein concentrations were within the normal rage: cholesterol (199 +/- 18 mg/dL); triglyceride (128 +/ 12 mg/dL); low-density cholesterol (127 +/- 8 mg/dL; and high-density cholesterol (47 +/- 3 mg/dL). The results indicated that degree of insulin resistance was significantly correlated with concentrations of sE-selectin (r = 0.54, P < 0.005), sICAM-1 (r = 0.67, P < 0.001), and sVCAM-1 (r = 0.41, P < 0.05). Furthermore, the relationship between insulin resistance and both sE selectin and sI-CAM-1 remained statistically significant when adjusted for differences in age, gender, body mass index, and all measures of lipoprotein concentrations. Finally, mononuclear cell binding correlated significantly with concentrations of sE-selectin (r = 0.54, P < 0.005) and sICAM-1 (r = 0.47, P < 0.01). These findings raise the possibility that previously described relationships between soluble adhesion molecules in patients with hypertension, type 2 diabetes, and dyslipidemia may be due to the presence of insulin resistance in these clinical syndromes and suggests that insulin resistance may predispose individuals to coronary heart disease by activation of cellular adhesion molecules. PMID- 10522985 TI - In vivo semiquantification of hypothalamic growth hormone-releasing hormone (GHRH) output in humans: evidence for relative GHRH deficiency in aging. AB - GH secretion declines with aging. The neuroendocrine mechanisms of somatopause are uncertain. To semiquantify endogenous hypothalamic GHRH output, we measured the suppressibility of spontaneous and GHRH-stimulated GH secretion by graded doses of a specific competitive GHRH receptor antagonist (N-Ac-Tyr1,D-Arg2)GHRH (1-29) in healthy young and elderly men. Nocturnal GH was about 30% lower in the elderly than in the young. Graded boluses of GHRH elicited dose-dependent GH responses, with no difference between the two age groups. Graded infusions of GHRH antagonist suppressed GH responses to GHRH in a dose-dependent manner, but with similar potency in both groups. The degree of inhibition depended on the magnitude of GHRH bolus: the dose-inhibition curves for the low GHRH boluses were shifted to the left compared to those with the high GHRH bolus (P = 0.01). Similarly, the dose-inhibition curve for spontaneous GH secretion was shifted to the left for the elderly compared to the young men (P = 0.01). Thus, the model of graded infusions of GHRH antagonist differentiates between different amounts of GHRH presented to the pituitary, permitting semiquantification of the endogenous hypothalamic GHRH output in vivo. Our data suggest that there is an age-dependent decrease in the endogenous hypothalamic GHRH output contributing to the age associated GH decline. PMID- 10522986 TI - Disruption of the young-adult synchrony between luteinizing hormone release and oscillations in follicle-stimulating hormone, prolactin, and nocturnal penile tumescence (NPT) in healthy older men. AB - The healthy human male hypothalamo-pituitary-gonadal axis exhibits age-dependent loss of coordinate LH-testosterone secretion. A putative independent defect in Leydig-cell steroidogenesis with aging would confound the attribution of such LH testosterone asynchrony to a hypothalamo-pituitary locus per se. Accordingly, here we appraise by sampling every 2.5 min overnight the joint synchrony of moment-to-moment LH release with simultaneously monitored pituitary FSH secretion, prolactin release, and nocturnal penile tumescence (NPT) oscillations, as a neurophysiological correlate of sleep regulation) in 10 young (ages 21-34) and 8 older (ages 62-72) healthy men. Joint synchrony for paired LH-FSH, LH prolactin, and LH-NPT observations in young vs. older individuals was quantified by the cross-approximate entropy (cross-ApEn) statistic, with larger cross-ApEn values indicating greater two-variable asynchrony. Concomitantly, we assessed (possible) univariate changes with age for each of LH, FSH, prolactin, and NPT, as quantified by approximate entropy (ApEn). Hormone assays were performed by random-access direct chemiluminescence analyzer. Overnight mean (+/- SEM) serum LH concentrations (IU/L) were equivalent in older (3.1 +/- 0.31 IU/L) and younger (2.9 +/- 0.29) men, as were their serum total testosterone concentrations; viz., 425 +/- 48 (older) and 523 +/- 40 (younger) ng/dL. However, all three sets of paired time-series were significantly more asynchronous in the older cohort. First, cross-ApEn of paired LH-FSH release was significantly higher (or more asynchronous) in older subjects; viz., 1.902 +/- 0.022 in older men vs. 1.607 +/- 0.058 in younger individuals (P = 0.0005). Second, cross-ApEn of paired LH and prolactin release was 1.744 +/- 0.085 in older volunteers vs. 1.346 +/- 0.084 in younger subjects (P = 0.0046). Third, and most notably, cross-ApEn for the joint LH-NPT observation time-series was significantly greater in older subjects at 1.771 +/- 0.056 vs. 1.223 +/- 0.086 (young) (P = 0.0001), thereby denoting loss of coordination between (neural) signals directing intermittent LH secretion and those governing sleep-associated penile tumescence in older men. Among one variable results, only ApEn of LH release was significantly higher in older individuals at 1.323 +/- 0.058 vs. 0.897 +/- 0.089 in younger subjects (P = 0.0019), signifying greater disorderliness of the LH secretory process in aged men. Individual ApEn values of FSH and prolactin release and NPT were age invariant. In ensemble, the present clinical experiments indicate that, within the aging male reproductive axis, bihormonal network disruption is more pronounced than individual signal disruption. We suggest that abrogation of joint synchrony among hypothalamically directed pituitary hormones and a neurogenically organized sexual response (nocturnal penile tumescence) can be unified thematically under an hypothesis of disrupted central nervous system hypothalamo pituitary network coordination in human aging. Such implicit disarray of multinodal communication is of consequence both scientifically and clinically, especially in proposing aging theories and intervention strategies. PMID- 10522987 TI - Joint basal and pulsatile hypersecretory mechanisms drive the monotropic follicle stimulating hormone (FSH) elevation in healthy older men: concurrent preservation of the orderliness of the FSH release process: a general clinical research center study. AB - To appraise the neuroendocrine mechanisms that underlie a selective (monotropic) elevation of serum FSH concentrations in healthy older men, we sampled blood in 11 young (ages 21-34) and 8 older men (ages 62-72) men every 2.5 min overnight. Serum FSH concentrations were quantitated in an automated, high-sensitivity, chemiluminescence-based assay. Rates of basal and pulsatile FSH secretion were estimated by deconvolution analysis, and the orderliness of the FSH release process via quantitated the approximate entropy statistic. Statistical analysis revealed that healthy older men manifest dual neuroendocrine hypersecretory mechanisims; specifically, a 2-fold increase in the basal (nonpulsatile) FSH secretion rate, and a concurrent 50% amplification of FSH secretory burst mass (and amplitude). The regularity or orderliness of ad seriatim FSH release is preserved in older individuals. We postulate that higher basal FSH secretion in older men is a consequence of reduced testosterone negative feedback, whereas amplified FSH secretory burst mass reflects net enhanced stimulation of gonadotrope cells by endogenous FSH secretagogues (e.g. GnRH and/or activin). The foregoing specific mechanisms driving heightened FSH secretion in older men contrast with the lower-amplitude pulsatility and more disorderly patterns of LH release in the same individuals. Thus, the present data illuminate an age dependent disparity in the disruption of FSH neuroregulation in the aging male. PMID- 10522988 TI - Inactivity amplifies the catabolic response of skeletal muscle to cortisol. AB - Severe injury or trauma is accompanied by both hypercortisolemia and prolonged inactivity or bed rest (BR). Trauma and BR alone each result in a loss of muscle nitrogen, albeit through different metabolic alterations. Although BR alone can result in a 2-3% loss of lean body mass, the effects of severe trauma can be 2- to 3-fold greater. We investigated the combined effects of hypercortisolemia and prolonged inactivity on muscle protein metabolism in healthy volunteers. Six males were studied before and after 14 days of strict BR using a model based on arteriovenous sampling and muscle biopsy. Fractional synthesis and breakdown rates of skeletal muscle protein were also directly calculated. Each assessment of protein metabolism was conducted during a 12-h infusion of hydrocortisone sodium succinate (120 microg/kg x h), resulting in blood cortisol concentrations that mimic severe injury (approximately 31 microg/dL). After 14 days of strict BR, hypercortisolemia increased phenylalanine efflux from muscle by 3-fold (P < 0.05). The augmented negative amino acid balance was the result of an increased muscle protein breakdown (P < 0.05) without a concomitant change in muscle protein synthesis. Muscle efflux of glutamine and alanine increased significantly after bed rest due to a significant increase in de novo synthesis (P < 0.05). Thus, inactivity sensitizes skeletal muscle to the catabolic effects of hypercortisolemia. Furthermore, these effects on healthy volunteers are analogous to those seen after severe injury. PMID- 10522989 TI - A novel case of multiple endocrine neoplasia type 2A associated with two de novo mutations of the RET protooncogene. AB - We report a novel case of multiple endocrine neoplasia type 2A (MEN 2A) associated with two mutations of the protooncogene RET. One affects codon 634 and causes a cysteine to arginine substitution; the second at codon 640 causes an alanine to glycine substitution in the transmembrane region. The two mutations were present on the same RET allele and were detected in germline and tumor DNA. Both mutations were de novo, i.e. they were not found in the DNA of the parents or relatives. Immunohistochemical and RT-PCR analysis showed that the pheochromocytoma expressed calcitonin as well as both RET alleles. A cell line established from the tumor and propagated in culture sustained the expression of RET and calcitonin, as did the original pheochromocytoma. Because the patient presented with medullary thyroid carcinoma and pheochromocytoma without parathyroid gland involvement, we speculate that this clinical picture could be correlated with the two RET mutations and to the unusual calcitonin production. This is the first report of a MEN 2A case due to two mutations of the RET gene and associated with a calcitonin-producing pheochromocytoma. PMID- 10522990 TI - Short term cardiovascular effects of aldosterone in healthy male volunteers. AB - Clinical evidence of rapid, nongenomic aldosterone effects in the cardiovascular system has been provided by clinical studies; an increase in systemic vascular resistance (SVR) was shown by invasive techniques within 3 min after injection of aldosterone. Here, we study the dose dependency and the later course of the rapid aldosterone effects by noninvasive techniques. In 12 healthy male volunteers, SVR and heart rate variability were determined by impedance cardiography and digital electrocardiography, respectively, for 8 h after the injection of 0.05 or 0.5 mg aldosterone in a double blind, placebo-controlled, 3-fold cross-over study. No significant differences were observed for baseline values among the three treatments. The area under the curve of SVR during the first 45 min after injection was significantly different between the periods with the highest areas under the curve seen after the injection of 0.5 mg aldosterone (mean +/- SD, 40.4 +/- 12.8 vs. 36.8 +/- 10.3 for 0.05 mg aldosterone and 36.8 +/- 10.4 for placebo; P = 0.05). Individual comparisons showed significant differences at 6 and 30 min between placebo and the 0.5 mg aldosterone period (P < 0.05), with values for the 0.05 mg aldosterone period similar to those for the placebo period. From 330-390 min, opposite changes occurred; SVR was depressed during the 0.05 mg (P < 0.05) and 0.5 mg aldosterone periods compared with that during the placebo period. These delayed effects may reflect an increased vagal tone in the aldosterone groups, as demonstrated by higher values of the time domain parameter of heart rate variability pNN50. This study provides further evidence for clinically detectable rapid cardiovascular aldosterone effects in vivo obtained by noninvasive techniques. The data are consistent with the view of aldosterone as a rapid modulator of cardiovascular responses acting through nongenomic mechanisms. PMID- 10522991 TI - Plasma cholesterol esterification and transfer, the menopause, and hormone replacement therapy in women. AB - With the onset of the menopause, plasma lipids and lipoprotein metabolism changes toward a more atherogenic profile that is improved by HRT. To determine whether cholesterol esterification rate (CER) and transfer of cholesteryl esters from high density lipoproteins to apolipoprotein B-containing lipoproteins are affected by menopause and HRT, plasma newly synthesized cholesteryl ester transfer (NCET) activity, CER and plasma lipids, lipoproteins, and apolipoprotein concentrations were measured in perimenopausal women (age range: 40-55 yr), including 49 premenopausal women and 32 postmenopausal women who were subsequently randomized to receive either placebo or 17-beta estradiol/norethisterone for 6 months. Plasma NCET (P = 0.03) and CER (P = 0.008) were significantly higher in postmenopausal women. Plasma low density lipoprotein cholesterol concentration, high density lipoprotein concentration, and body mass index were independent predictors of plasma NCET in premenopausal women, and plasma triglyceride and apolipoprotein B concentrations were corresponding predictors in postmenopausal women. When data were adjusted for plasma triglyceride, plasma NCET activity was no longer significantly different (P = 0.81) between premenopausal and postmenopausal women. Plasma NCET and CER did not change significantly in postmenopausal women during HRT. These data suggest that the determinants of plasma NCET activity after menopause and increased levels of triglyceride-rich lipoprotein acceptors of cholesteryl esters may lead to increased plasma NCET that is not reduced by HRT in postmenopausal women. PMID- 10522992 TI - Diagnostic value of fluorometric assays in the evaluation of precocious puberty. AB - To establish normative data and determine the value of fluorometric AutoDELFIA assays (Wallac Oy) in the investigation of precocious puberty, we determined serum levels of LH, FSH, testosterone, and estradiol under basal and GnRH stimulated conditions in 277 normal subjects at various pubertal stages and in 77 patients with precocious puberty. A substantial overlap was observed in basal and GnRH-stimulated gonadotropin levels in normal individuals of both sexes with pubertal Tanner stages 1 and 2. The 95th percentile of the normal prepubertal population was the cut-off limit between prepubertal and pubertal levels. These limits were 0.6 IU/L in both sexes for basal LH, 9.6 IU/L in boys and 6.9 IU/L in girls for peak LH after GnRH stimulation, 19 ng/dL in boys for basal testosterone, and 13.6 pg/mL in girls for basal estradiol. Basal and peak LH exceeding these limits were considered positive tests for the diagnosis of gonadotropin-dependent precocious puberty. According to these criteria, the sensitivities of basal and peak LH for the latter diagnosis were 71.4% and 100% in boys, and 62.7% and 92.2% in girls. The specificity and positive predicted value were 100% in both sexes for basal and peak LH levels. The negative predicted values for basal and peak LH were 62.5% and 100% in boys, and 40.6% and 76.5% in girls. Basal and GnRH-stimulated FSH levels overlapped among the various pubertal stages in normal subjects and were, in general, not helpful in the differential diagnosis of precocious puberty. In conclusion, basal LH levels were sufficient to establish the diagnosis of gonadotropin-dependent precocious puberty in 71.4% of boys and 62.7% of girls. In the remaining patients, a GnRH stimulation test was still necessary to confirm this diagnosis. Finally, suppressed LH and FSH levels after GnRH stimulation indicate gonadotropin independent sexual steroid production. PMID- 10522993 TI - Serum parathyroid hormone-related protein levels and response to bisphosphonate treatment in hypercalcemia of malignancy. AB - The pathogenesis of hypercalcemia of malignancy comprises increased net bone resorption and enhanced renal tubular reabsorption of calcium (Ca). To evaluate the prevalence of an increased renal tubular reabsorption of Ca index [tubular reabsorption of calcium index (TRCaI)] in cancer patients with hypercalcemia and of elevated circulating levels of PTH-related protein (PTHrP), which is recognized as a major mediator of this syndrome, we investigated 315 well rehydrated patients, aged 58.1 +/- 0.7 yr (mean +/- SEM), with hypercalcemia [albumin-corrected plasma Ca (pCa), >2.7 mmol/L] secondary to histologically proven malignancy. Changes in pCa and, therefore, various Ca filtered loads were obtained by different degrees of bone resorption inhibition achieved with a single infusion of the bisphosphonate ibandronate, given at various doses on a randomized, double blind basis. PTHrP was determined at baseline in 147 of the patients and 7 days after bisphosphonate therapy in 73. Before ibandronate therapy, pCa was 3.36 +/- 0.02 mmol/L, mean TRCaI was increased at 3.09 +/- 0.03 mmol/L glomerular filtration rate (GFR; normal, 2.40-2.90), and 65% of patients had TRCaI above 2.90 mmol/L GFR. Mean serum PTHrP levels were 4.9 +/- 0.5 pmol/L (normal, <2.5) and values above the normal range were found in 53% of the patients (76% in lung and upper respiratory tract malignancies). By 7 days after the infusion of ibandronate, a decrease in pCa of 0.69 +/- 0.03 mmol/L (20.0 +/- 0.7%; P < 0.001) and in bone resorption [mean change in fasting urinary Ca, 0.09 +/- 0.04 mmol/L GFR (47.6 +/- 8.6%; P < 0.001) and 14.4 +/- 1.7 nmol/mmol (27.6 +/- 10.6%; P < 0.01) in deoxypyridinoline] was observed. TRCaI was slightly lowered by 0.30 +/- 0.09 mmol/L GFR. Mean changes in PTHrP, 1,25-dihydroxyvitamin D3, and PTH were +0.7 +/- 0.4 (P = NS), +27.6 +/- 3.0 (P < 0.001), and +2.9 +/- 0.8 (P < 0.005) pmol/L, respectively. After ibandronate treatment, the relative risk of relapsing hypercalcemia was particularly increased (3.43-fold) in lung and upper respiratory tract malignancies. These results obtained in a large cohort of patients indicate a significant prevalence of an increased renal tubular reabsorption of calcium index in hypercalcemia of malignancy and a substantial proportion of patients with detectable PTHrP. PMID- 10522994 TI - Direct postoperative and follow-up results of transsphenoidal surgery in 19 acromegalic patients pretreated with octreotide compared to those in untreated matched controls. AB - In this study 19 patients were preoperatively treated with octreotide for 1-17 months (mean, 5 months), with doses from 150-1500 microg daily, and those patients were matched to 19 untreated patients with comparable tumor classification and preoperative serum GH concentrations. Octreotide was started at 300 microg daily by s.c. injections or continuous sc infusion using a pump in increasing doses, depending on the responses of the serum GH and insulin-like growth factor I (IGF-I) concentrations. During pretreatment, seven patients achieved a serum GH concentration below 5 mU/L, whereas six patients normalized their serum IGF-I. Postoperatively, a serum GH concentration below 5 mU/L was achieved in 15 pretreated and 14 untreated patients, a normal serum IGF-I level (<2 SD) was achieved in 10 pretreated and 15 untreated patients, and normal serum GH suppression during GTT was reached in 12 treated and 14 control patients. No differences were found in complication rate or incidence of hypopituitarism caused by surgery. Adjuvant therapy was required in 7 treated and 5 untreated patients. At follow-up examination, 5.7 and 4 yr postoperatively, 10 pretreated and 12 control patients could be considered cured by surgery only, according to our criteria for remission (serum GH, <5 mU/L; normal GH suppression and normal serum IGF-I). In summary, we found no difference in direct postoperative and follow-up results of transsphenoidal surgery between pretreated patients and untreated patients. This finding is in discordance with other studies, which have claimed a beneficial effect of octreotide pretreatment. PMID- 10522995 TI - 7Alpha-methyl-19-nortestosterone maintains sexual behavior and mood in hypogonadal men. AB - The synthetic steroid 7alpha-methyl-19-nortestosterone (MENT) is a potent androgen that is resistant to 5alpha-reductase. It thus has decreased activity at the prostate and may have advantages over testosterone-based regimens in long term treatment or as part of a male contraceptive. Administration to eugonadal men results in suppression of gonadotropins, but its ability to support androgen dependent behavior has not been investigated. For sustained release administration, MENT acetate was used, because its diffusion characteristics were more suitable for use in implants. However, upon release the acetate is rapidly hydrolyzed, and MENT is the biologically active moiety in circulation. We studied the effects of MENT on sexual interest and activity, spontaneous erection, and mood states in comparison with testosterone enanthate (TE) in 20 Caucasian and Chinese hypogonadal men recruited in Edinburgh and Hong Kong (n = 10 in each center). Outcomes were measured using a combination of daily diaries, semistructured interviews, and questionnaires. Nocturnal penile tumescence (NPT) was also recorded in the Edinburgh group. After withdrawal of androgen replacement treatment (wash-out phase) for a minimum of 6 weeks, subjects were randomized to two groups in a cross-over design. Drug treatment regimens were of 6-week duration and consisted of two implants, each containing 115 mg MENT acetate, inserted s.c. into the upper arm and removed after 6 weeks and two injections of TE (200 mg, i.m.) 3 weeks apart. MENT treatment resulted in stable plasma MENT concentrations of 1.4 +/- 0.1 nmol/L after 3 weeks and 1.3 +/- 0.1 nmol/L after 6 weeks (mean +/- SEM; all men). Nadir testosterone concentrations were 3.6 +/- 0.6 nmol/L at the end of the wash-out phase and 9.4 +/- 0.6 nmol/L 3 weeks after each injection. There were no differences in hormone concentrations between centers. There were no adverse toxicological effects. There were only minor differences between the two treatments. Both MENT and TE treatment resulted in significant increases in sexual interest and activity, spontaneous erection (both by self-report and NPT measurement), and increases in positive moods, with decreases in negative moods in the Edinburgh group. In the Hong Kong group, both treatments increased waking erection, with a trend toward increased sexual interest and activity. Mood states appeared to be less affected during the wash out phase than in Edinburgh men and showed no significant response to either treatment. These results demonstrate that MENT has similar effects on sexual activity and mood states as testosterone in hypogonadal men. As NPT is a physiological androgen-dependant outcome, these data provide further evidence for the androgenicity of MENT. The lack of detected effect of either androgen in Hong Kong men other than on waking erection illustrates the importance of the cultural context of symptomatology and its measurement. The appropriate dose of MENT remains to be determined, but these results support its development as a potential androgen replacement therapy. PMID- 10522996 TI - Combined hypothalamic-pituitary-gonadal defect in a hypogonadic man with a novel mutation in the DAX-1 gene. AB - We have studied a 20-yr-old male patient with adrenal hypoplasia congenita and hypogonadotropic hypogonadism (HH) due to a C to A transversion at nucleotide 825 in the DAX-1 gene, resulting in a stop codon at position 197. The same mutation was detected in his affected first cousin (adrenal hypoplasia congenita and HH) and in a heterozygous state in their carrier mothers. The patient had had acute adrenal insufficiency at the age of 2 yr and 6 months, bilateral cryptorchidism corrected surgically at the age of 12 yr, and failure of spontaneous puberty. Plasma testostereone (T) was undetectable (<0.30 nmol/L), gonadotropin levels were low (LH, <0.4 IU/L; FSH, 1.5 IU/L) and not stimulated after i.v. injection of 100 microg GnRH. The endogenous LH secretory pattern was apulsatile, whereas free alpha-subunit (FAS) levels depicted erratic pulses, suggesting an incomplete deficiency of hypothalamic GnRH secretion. During i.v. pulsatile GnRH administration (10 microg/pulse every 90 min for 40 h), each GnRH pulse induced a LH response of low amplitude (0.54 +/- 0.05 UI/L), whereas mean LH (0.45 +/- 0.01 IU/L) and FAS (63 +/- 8 mU/L) levels remained low. Amplitude of LH peaks (0.83 +/ 0.09 IU/L), mean LH (0.53 +/- 0.02 IU/L), and FAS (161 +/- 18 mU/L) levels increased (P < 0.01), whereas the T concentration remained low (0.75 nmol/L) when the pulsatile GnRH regimen was raised to 20 microg/pulse for a 40-h period, suggesting a partial pituitary resistance to GnRH. Thereafter, plasma T levels remained in prepubertal value after three daily im injections of 5000 IU hCG (3.6 nmol/L) and after 1-yr treatment with weekly i.m. injections of 1500 IU hCG (1.2 nmol/L), implying Leydig cell resistance to hCG. The patient had a growth spurt, bone maturation, progression of genital and pubic hair stages, and normalization of plasma T level (15.8 nmol/L) after a 12-month treatment with twice weekly injections of hCG and human menopausal gonadotropin (75 IU International Reference Preparation 2) preparations, suggesting that, in presence of FSH, a Sertoli cell-secreted factor stimulated Leydig cell production of T. In conclusion, we report a novel mutation in the DAX-1 gene in patients with AHC and HH. Our results suggest that the hypogonadism is due to a combined hypothalamic pituitary-gonadal defect and imply that the DAX-1 gene may play a critical role in human testicular function. PMID- 10522997 TI - Apparent cortisone reductase deficiency: a functional defect in 11beta hydroxysteroid dehydrogenase type 1. AB - A 36-yr-old woman was referred to the endocrine clinic for investigation of oligomenorrhea, hirsutism, and acne. She was plethoric and overweight with central fat distribution. Plasma cortisol was normal, but her adrenal glands were enlarged (CT scan). Urinary tetrahydrocortisone excretion rate was consistently high, raising the possibility of 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) deficiency. In addition, 5beta- reduction of cortisol and cortisone was markedly enhanced. The levels of all cortisol metabolites were suppressed normally with dexamethasone, but conversion of oral cortisone acetate to plasma cortisol was delayed and subnormal compared with that of healthy volunteers. This was accompanied by a larger than normal increase in plasma cortisone concentration. Thus, the defect appears to be in 11beta-HSD1 activity and not in 5beta-reductase activity. Three close relatives of the subject showed no comparable abnormalities, and analysis of the coding region and exon/intron boundaries of the 11beta-HSD1 gene of the case revealed no differences from the consensus sequence. The defect may lie outside the coding region. Alternatively, some other inherited or acquired defect may lead to inhibition of this enzyme system. PMID- 10522998 TI - Lack of effect of GnRH agonists on final height in girls with advanced puberty: a randomized long-term pilot study. AB - GnRH agonists improve final height in girls with "true" precocious puberty. To test if a comparable effect can be obtained in older girls, we performed a long term controlled study in 30 caucasian girls whose puberty started between 8.4 and 10 yr (9.4 +/- 0.1 yr), a variant of normal called "advanced" puberty. At entry into trial, these girls had clinical, biological, and sonographic manifestations of puberty and a bone age greater than 10.9 yr. They were randomized 2:1 to receive 3.75 mg triptorelin im every 4 weeks for 2 yr (n = 20, group I) or no treatment (n = 10, group II). Mean height at inclusion was 135.2 +/- 4.3 cm (+0.6 SDS) in group I, 136.1 +/- 4.2 cm (+0.8 SDS) in group II, with target height 157.6 +/- 4.3 cm (group I) and 157.8 +/- 4.7 cm (group II), and predicted height (Bayley-Pinneau) 154.1 +/- 3.9 cm and 155.2 +/- 3.7 cm. Although GnRH agonists transiently delayed sexual maturation as well as bone age and growth rate, they had no clear-cut long-standing effect, and final height was comparable in treated (157.6 +/- 4.0 cm) and untreated girls (156.1 +/- 5.3 cm) (NS). PMID- 10522999 TI - Cerebrospinal fluid and plasma leptin measurements: covariability with dopamine and cortisol in fasting humans. AB - Leptin (OB protein) is an important signal in the regulation of energy balance. Leptin levels correlate with adiposity, but also decrease acutely with caloric restriction and increase with refeeding. The brain is an established critical site of leptin function, yet little is known about leptin concentrations in the central nervous system relative to plasma levels, psychiatric diagnoses, and other endocrine parameters. Therefore, using a novel ultrasensitive leptin assay, we explored relationships of human plasma and cerebrospinal fluid (CSF) leptin levels to body mass index, smoking, posttraumatic stress disorder diagnosis, and levels of dopamine, monoamine metabolites, beta-lipotropin, glucocorticoid, and thyroid and cytokine hormones. A strong linear relation between CSF and plasma leptin levels in the am (r = 0.63; P < 0.002) and afternoon (r = 0.90; P < 0.0001) was revealed. CSF and plasma leptin concentrations decreased during a 12- to 20-h period of fasting. A strong association was found between plasma leptin and CSF dopamine levels (r = 0.74; P < 0.01) as well as between CSF leptin levels and urinary free cortisol (r = 0.73; P < 0.01). Both of these parameters covaried with leptin independently of adiposity, as estimated by body mass index. Implications for leptin transport, regulation, and its potential role in therapeutic strategies for obesity and diabetes are discussed. PMID- 10523000 TI - Serum leptin levels correlate with growth hormone secretion and body fat in children. AB - The aim of this study was to investigate the relationship among GH secretion, leptin concentrations, and body composition measured with x-ray absorptiometry (DXA) in children. In total, 71 children were investigated, 51 males and 20 females. Their mean chronological age was 10.8 yr (range, 6.2-17.7 ys), and their mean height (SD) was -2.1 (0.63) SD scores. Their mean weight for height SD scores (WH(SDS)) was 0.2 (1.18). Body composition was investigated using DXA. Blood samples were taken for analysis of leptin, insulin-like growth factor I (IGF-I), IGF-binding protein-3, and 24-h GH secretion. A positive correlation was found between leptin and total body fat (r = 0.83; P < 0.0001) and when fat was expressed as a percentage of body weight (r = 0.86; P < 0.0001). There were significant (P < 0.0001) relationships between leptin and WH(SDS) (r = 0.45) and between leptin and body mass index (r = 0.69). A significant gender difference in leptin levels was found, but this disappeared after adjustment for body fat, as measured by DXA. There were significant (P < 0.001) inverse correlations between leptin and the AUCb for GH (r = -0.41), leptin, and GHmax (r = -0.38), where AUCb is the area under the curve above the calculated baseline, and GHmax is the maximum peak during the 24-h GH profile (percent fat and AUCb for GH, r = -0.43; percent fat and GHmax, r = -0.39). In a multiple stepwise forward regression analysis with leptin as the dependent variable, the percent trunk fat accounted for 77.7% of the leptin variation. With AUCb for GH as the dependent variable, the percent trunk fat accounted for 20.3% of the variation. With GHmax as the dependent variable, the percent trunk fat accounted for 18.8% of the variation, IGF-binding protein-3 for another 8.5%, and the percentage of fat from arms and legs for another 4.4%. We demonstrated a strong positive correlation between leptin levels and body fat, a significant negative correlation between leptin levels and GH secretion, and a significant negative correlation between body fat and GH secretion. We have also shown that specific regional fat depots have different relationships with leptin and particular markers of GH secretion. PMID- 10523001 TI - Responses of the growth hormone (GH) and insulin-like growth factor axis to exercise, GH administration, and GH withdrawal in trained adult males: a potential test for GH abuse in sport. AB - GH abuse by elite athletes is currently undetectable. To define suitable markers of GH doping, we assessed the effects of acute exercise, GH administration, and GH withdrawal on the GH/insulin-like growth factor (IGF) axis in athletic adult males. Acute endurance-type exercise increased serum GH, GH-binding protein (GHBP), total IGF-I, IGF-binding protein (IGFBP)-3, and acid-labile subunit (ALS), each peaking at the end of exercise. IGFBP-1 increased after exercise was completed. Free IGF-I did not change with exercise. Recombinant human GH treatment (0.15 IU/kg x day) for 1 week increased serum total IGF-I, IGFBP-3, and ALS, exaggerating the responses to exercise. IGFBP-2 and IGFBP-1 were trivially suppressed. After GH withdrawal, the GH response to identical exercise was suppressed. Total IGF-I, IGFBP-3, and ALS returned to baseline over 3-4 days. In summary, 1) acute exercise transiently increased all components of the IGF-I ternary complex, possibly due to mobilization of preformed intact complexes; 2) GH pretreatment augmented the exercise-induced changes in ternary complexes; 3) postexercise IGFBP-1 increments may protect against delayed onset hypoglycemia; 4) serum total IGF-I, IGFBP-3, and ALS may be suitable markers of GH abuse; and 5) differences in disappearance times altered the sensitivity of each marker for detecting GH abuse. PMID- 10523002 TI - A possible role of immunoglobulin E in patients with hyperthyroid Graves' disease. AB - To investigate the possible participation of immunoglobulin E (IgE) in the autoimmune process of Graves' disease, incidence of elevation of serum IgE level, TSH receptor antibody (TRAb), and thyroid status were studied in 66 patients with hyperthyroid Graves' disease, 54 patients with Hashimoto's thyroiditis, 19 patients with bronchial asthma, and 15 patients with pollen allergy. In hyperthyroid Graves' patients, elevation of serum IgE levels (> or = 170 U/mL) was found in 19 of 66 patients (29%), 11 of whom had hereditary and/or allergic conditions. Elevations of serum IgE levels were found in 63% of patients with bronchial asthma and in 40% of patients with pollen allergy. Mean values of serum IgE were the same in patients with hyperthyroid Graves' disease and with bronchial asthma. During methimazole treatment TRAb decreased without fluctuation of IgE levels in both groups. The decrease in TRAb was significantly greater in patients with normal IgE than in patients with IgE elevation. After prednisone administration, reduction in TRAb was greater in patients with normal IgE than that in patients with IgE elevation. High incidence of IgE elevation in hyperthyroid Graves' disease and slower reduction in TRAb in association with IgE elevation suggest a difference in the autoimmune processes in Graves' disease with and without elevation of IgE. PMID- 10523003 TI - A high frequency of Y chromosome deletions in males with nonidiopathic infertility. AB - Microdeletions of the long arm of the human Y chromosome are associated with spermatogenic failure and have been used to define three regions of Yq (AZFa, AZFb, and AZFc) that are recurrently deleted in infertile males. In a blind study we screened 131 infertile males (46 idiopathic and 85 nonidiopathic) for Y chromosome microdeletions. Nineteen percent of idiopathic males, with an apparently normal 46,XY chromosome complement had microdeletions of either the AZFa, AZFb, or AZFc region. There was no strict correlation between the extent or location of the deletion and the phenotype. The AZFb deletions did not include the active RBM gene. Significantly, a high frequency of microdeletions (7%) was found in patients with known causes of infertility and a 46,XY chromosome complement. These included deletions of the AZFb and AZFc regions, with no significant difference in the location or extent of the deletion compared with the former group. It is recommended that all males with reduced or absence sperm counts seeking assisted reproductive technologies be screened for deletions of the Y chromosome. PMID- 10523004 TI - Ethnicity and migration as determinants of human prostate size. AB - The roles of ethnicity and migration in determining the size of human prostate zones during midlife were explored. Prostate size was measured by planimetric ultrasound in 163 men residing in Sydney who were either Australian non-Chinese (AR; n = 116) or Chinese migrants (ACM; n = 47) and had lived in Australia for a median of 7.3 yr (range, 0.2-25 yr). These were compared with Chinese men residing in China (CR; n = 210). Central and total prostate volumes were estimated by a single observer using the same equipment at both sites. After adjustment for age, central and total prostate volumes were significantly smaller, and plasma prostate-specific antigen and 5alpha-dihydrotestosterone (DHT) concentrations and International Prostate Syndrome Scores were significantly lower, in CR compared with either ACM or AR, whereas the scores of the latter two groups were similar. Almost all of the population difference in total prostate volumes could be accounted for by differences in central prostate volumes. The strongest correlates of age-adjusted prostate volume were prostate specific antigen and DHT, the latter presumably reflecting the quantitative importance of prostatic stromal type II 5alpha-reductase activity to circulating DHT concentrations. Sex hormone-binding globulin concentrations were significantly higher in CR and significantly lower in ACM compared with those in AR, but the significance of these observations is unclear. These findings highlight the importance of the central zone of the prostate as well as provide evidence for an environmental factor influencing prostate growth. This factor operates over a relatively short time period compared with the evolution of prostate disease. Hence, this study provides evidence that ethnicity and geographical factors, such as migration, can influence the growth of the normal human prostate during midlife and may facilitate future studies of the origins and pathogenesis of human prostate disease. PMID- 10523005 TI - A soluble PC-1 circulates in human plasma: relationship with insulin resistance and associated abnormalities. AB - An increased tissue content of PC-1, an inhibitor of insulin receptor signaling, may play a role in insulin resistance. Large scale prospective studies to test this hypothesis are difficult to carry out because of the need for tissue biopsies. The aim of this study was to investigate whether PC-1 is measurable in human plasma and whether its concentration is related to insulin sensitivity. A soluble PC-1, with mol wt and enzymatic activity similar to those of tissue PC-1, was measurable in human plasma by a specific enzyme-linked immunosorbent assay and was inversely correlated to skeletal muscle PC-1 content (r = -0.5; P < 0.01). The plasma PC-1 concentration was decreased (P < 0.05) in insulin resistant (22.7 +/- 3.0 ng/mL; n = 25) compared to insulin-sensitive (36.7 +/- 4.5; n = 25) nondiabetic subjects and was correlated negatively with the waist/hip ratio (r = -0.48; P < 0.001) and mean blood pressure (r = -0.3; P < 0.05) and positively with high density lipoprotein/total cholesterol (r = 0.38; P < 0.01) and both the M value and the plasma free fatty acid level decrement at clamp studies (r = 0.28; n = 50; P = 0.05 and r = 0.43; n = 22; P < 0.05, respectively). A plasma PC-1 concentration of 19 ng/mL or less identified a cluster of insulin resistance-related alterations with 75% accuracy. In conclusion, PC-1 circulates in human plasma, and its concentration is related to insulin sensitivity. This may help to plan studies aimed at understanding the role of PC-1 in insulin resistance. PMID- 10523006 TI - Hormonal and biochemical parameters in the determination of osteoporosis in elderly men. AB - The extent to which changes in several hormonal and biochemical parameters are involved in the pathogenesis of osteoporosis in men remains controversial. This study examined the roles of free testosterone (T), estradiol (E2), sex hormone binding globulin (SHBG), 25-hydroxyvitamin D, PTH, and insulin-like growth factor I in the determination of bone mineral density (BMD) in 437 community-dwelling elderly men. Age, height, weight, quadriceps strength, and femoral neck (FN) and lumbar spine (LS) BMD were also obtained. In multiple regression analysis, after adjusting for age and weight, low E2 (P = 0.01), and high SHBG (P = 0.0002) levels were common determinants of FN and LS BMD. In addition, high PTH (P = 0.03) was an independent predictor of FN BMD, and low free T (P = 0.02) was an independent predictor of LS BMD. Low free T was associated with FN BMD in univariate analysis only. The hormonal measurements collectively accounted for 5% and 7% of the age- and weight-adjusted variance of FN and LS BMD, respectively. The sex steroids, SHBG and insulin-like growth-I were found to be interrelated using a technique of path analysis that examines the intercorrelation between these variables. A subject with any one abnormal serum parameter had a 4-fold increase in the risk of osteoporosis, whereas three abnormal parameters were associated with an 11-fold increased risk, although the latter group only applied to 1% of the study population. Although the precise causal effects these biochemical parameters may have on the development of osteoporosis remains to be determined, the present findings support an important interrelated role for these hormonal and biochemical parameters on changes in bone density in elderly men. PMID- 10523007 TI - The feasibility of high dose iodine 131 treatment as an alternative to surgery in patients with a very large goiter: effect on thyroid function and size and pulmonary function. AB - Some patients with very large goiters (>150 mL) are not candidates for surgery. We evaluated the feasibility of high dose 131I in such patients. Twenty-three patients (2 men and 21 women; median age, 67 yr; range, 42-86 yr) with very large goiter (8 toxic) were treated with calculated high dose 131I [median, 2281 megabecquerels (61.6 mCi); range, 988-4620 megabecquerels (26.7-124.9 mCi)]. During the 12-month observation period, goiter reduction and tracheal anatomy were monitored by magnetic resonance imaging, and the respiratory capacity was monitored by pulmonary function tests. Five patients (22%) developed hypothyroidism. Thyroid volumes were at baseline, after 1 week, and after 1 yr [mean +/- SEM, 311 +/- 28, 314 +/- 26 (P = NS), and 215 +/- 26 (P < 0.01) mL]. The relative changes 1 week after therapy ranged from -14.1% to 15.3%. After 1 yr the mean size was reduced by 33.9% (range, 13.5-61.4%). Only the initial goiter size showed a significant negative correlation to the percent reduction. The smallest cross-sectional area of the trachea decreased 9.2% within 1 week after treatment, but eventually emerged with a 17.9% larger area [mean +/- SEM, 84.3 +/ 4.8, 75.5 +/- 5.1 (P < 0.01), and 98.2 +/- 6.0 (P < 0.01) mm2]. The inspiratory parameter, FIF50%, improved after an initial insignificant decline [baseline therapy, after 1 week, after 3 months, and after 1 yr (mean +/- SEM), 2.37 +/- 0.24, 2.20 +/- 0.21 (P = NS), 2.51 +/- 0.23 (P = NS), and 2.76 +/- 0.25 (P = 0.01) L/s]. FIF50% correlated significantly with the smallest cross-sectional tracheal area (baseline, 1 week, and 1 yr: r = 0.74; P < 0.001, r = 0.63; P < 0.005, and r = 0.46; P < 0.05). Changes in tracheal anatomy did not correlate with changes in either lung dynamics or goiter size. In conclusion, very large goiters can be reduced by a third, on the average, with high dose 131I therapy without any initial clinically significant tracheal compression. Tracheal cross sectional area as well as pulmonary inspiratory capacity improve. No serious adverse effects are seen. PMID- 10523008 TI - A clinical trial of injectable testosterone undecanoate as a potential male contraceptive in normal Chinese men. AB - This is a pilot dose-finding study of spermatogenic suppression using testosterone undecanoate (TU) injections alone in normal Chinese men. Thirty-two healthy men were recruited. Volunteers underwent pretreatment evaluation, then a treatment period in which group I (n = 13) received 500 mg TU, group II (n = 12) received 1000 mg TU, and group III (n = 7) received placebo, respectively, at monthly intervals during the treatment period (or until azoospermia was achieved). Thereafter, they underwent a recovery period until all parameters returned to pretreatment levels. Eleven of 12 volunteers in the 500-mg TU group, and all volunteers in the 1000-mg TU group became azoospermic. Faster suppression of spermatogenesis was achieved in the 1000-mg TU group. Serum testosterone increased significantly in the higher dose group at weeks 8 and 12, but remained within the normal range. Mean serum LH and FSH were profoundly suppressed by both doses to undetectable levels at week 16. TU injections did not cause a significant change in high density lipoprotein cholesterol levels. No serious side-effects were found. We conclude that both dosages of TU can effectively, safely, and reversibly suppress spermatogenesis in normal Chinese men. PMID- 10523009 TI - Comparison of adrenocorticotropin (ACTH) stimulation tests and insulin hypoglycemia in normal humans: low dose, standard high dose, and 8-hour ACTH-(1 24) infusion tests. AB - The efficacy of the standard high dose ACTH stimulation test (HDT), using a pharmacological 250-microg dose of synthetic ACTH-(1-24), in the diagnosis of central hypoadrenalism is controversial. The insulin hypoglycemia test is widely regarded as the gold standard dynamic stimulation test of the hypothalamo pituitary-adrenal (HPA) axis that provides the most reliable assessment of HPA axis integrity and reserve. Alternatively, a prolonged infusion of ACTH causes a continuing rise in plasma cortisol levels that may predict the adrenals' capacity to respond to severe ongoing stress. In nine normal subjects, we compared plasma ACTH and cortisol levels produced by three i.v. bolus low doses of ACTH-(1-24) (0.1, 0.5, and 1.0 microg/1.73 m2; LDTs) with those stimulated by hypoglycemia (0.15 U/kg insulin) and with the cortisol response to a standard 250-microg dose of ACTH-(1-24). The normal cortisol response to an 8-h ACTH-(1-24) infusion (250 microg at a constant rate over 8 h) was determined using three modern cortisol assays: a high pressure liquid chromatography method (HPLC), a fluorescence polarization immunoassay (FPIA), and a standard RIA. In the LDTs, stepwise increases in mean peak plasma ACTH were observed (12.4 +/- 2.0, 48.2 +/- 7.2, 120.2 +/- 15.5 pmol/L for the 0.1-, 0.5-, and 1.0-microg LDTs, respectively; P values all <0.0022 when comparing peak values between tests). The peak plasma ACTH level after insulin-induced hypoglycemia was significantly lower than that produced in the 1.0-microg LDT (69.6 +/- 9.3 vs. 120.2 +/- 15.5 pmol/L; P < 0.0002), but was higher than that obtained during the 0.5-microg LDT (69.6 +/- 9.3 vs. 48.2 +/- 7.2 pmol/L; P < 0.02). In the LDTs, statistically different, dose-dependent increases in peak cortisol concentration occurred (355 +/- 16, 432 +/- 13, and 482 +/- 23 nmol/L; greatest P value is 0.0283 for comparisons between all tests). The peak cortisol levels achieved during the LDTs were very different from those during the HDT (mean peak cortisol, 580 +/- 27 nmol/L; all P values <0.00009. However, the mean 30 min response in the 1.0-microg LDT did not differ from that in the HDT (471 +/- 22 vs. 492 +/- 22 nmol/L; P = 0.2). In the 8-h ACTH infusion test, plasma cortisol concentrations progressively increased, reaching peak levels much higher than those in the HDT [995 +/- 50 vs. 580 +/- 27 nmol/L (HPLC) and 1326 +/- 100 vs 759 +/- 31 nmol/L (FPIA)]. Significant differences in the basal, 1 h, and peak cortisol levels as determined by the three different assay methods (HPLC, FPIA, and RIA) were observed in the 8-h infusion tests. Similarly, in the HDTs there were significant differences in the mean 30 and 60 min cortisol levels as measured by HPLC compared with those determined by FPIA. We conclude that up to 30 min postinjection, 1.0 microg/1.73 m2 ACTH-(1-24) stimulates maximal adrenocortical secretion. Similar lower normal limits at 30 min may be applied in the 1.0-microg LDT and the HDT, but not when lower doses of ACTH-(1-24) are administered. The peak plasma ACTH level produced in the 1.0 microg LDT is higher than in the insulin hypoglycemia test, but is of the same order of magnitude. The peak cortisol concentration obtained during an 8-h synthetic ACTH-(1-24) infusion is considerably higher than that stimulated by a standard bolus 250-microg dose, potentially providing a means of evaluating the adrenocortical capacity to maintain maximal cortisol secretion. Appropriate interpretation of any of these tests of HPA axis function relies on the accurate determination of normal response ranges, which may vary significantly depending on the cortisol assay used. PMID- 10523010 TI - Sex-dependent association of a genetic polymorphism of cholesteryl ester transfer protein with high-density lipoprotein cholesterol and macrovascular pathology in type II diabetic patients. AB - Cholesterol ester transfer protein (CETP) is a key regulating factor of lipid metabolism, and the polymorphism of its gene may be a candidate for modulating the lipid parameters in type 2 diabetic subjects. In a group of 406 type 2 diabetic patients aged 59.5 +/- 10.8 y, with a body mass index of 28.9 +/- 5.3 kg/m2 and HbA1c = 8.2 +/- 1.9%, we studied the B polymorphism at the CETP locus detectable with the restriction enzyme TaqI. Patients were separated into groups, 231 males (78 B1B1, 108 B1B2, 45 B2B2) and 175 females (48 B1B1, 94 B1B2, 33 B2B2), and compared on the basis of their lipid parameters (total cholesterol, triglycerides, high-density lipoprotein-cholesterol (HDL-C), ApoA1 ApoB, and low density lipoprotein-cholesterol), their micro and macrovascular complications. HDL-C was significantly higher in man with the B2B2 genotype (respectively, 1.31 +/- 0.44 mmol/L vs. 1.13 +/- 0.32 mmol/L, P < 0.05), together with a lower incidence of coronary heart disease (9 vs. 25% for B1B1 and B1B2 together). Women displayed a higher HDL-C than men and a equally high incidence of coronary heart disease in B2 homozygotes as in other genotypes (26 vs. 27%). Thus, in type 2 diabetic patients, Taq1b polymorphism seems to exert a modulating role in males only. This may contribute to the loss of macrovascular protection in type 2 diabetic females. PMID- 10523011 TI - Y chromosome microdeletions in cryptorchidism and idiopathic infertility. AB - To clarify whether cryptorchidism might be the expression of an intrinsic congenital testicular abnormality, we investigated the frequency of Y chromosome long arm (Yq) microdeletions in unilateral excryptorchid subjects manifesting an important bilateral testiculopathy. Microdeletion analysis of Yq was performed by polymerase chain reaction in the following subjects: 40 unilateral excryptorchid patients with azoospermia or severe oligozoospermia due to a bilateral severe testiculopathy (Sertoli cell-only syndrome or severe hypospermatogenesis); 20 unilateral excryptorchid men with moderate oligozoospermia and a normal testicular cytological picture in the contralateral testis; 110 patients affected by idiopathic severe primary testiculopathies; 20 patients affected by idiopathic moderate testiculopathy; and, as controls, 50 patients affected by known causes of testiculopathy and 100 fertile men. Eleven of 40 (27.5%) unilateral excryptorchid patients affected by bilateral testiculopathy and 28 of 110 (25.4%) patients affected by idiopathic severe primary testiculopathy showed Yq microdeletions, whereas no microdeletions were found in all the other subjects, nor in male relatives of patients with deletions. Microdeletions were located in different parts of Yq, including known regions involved in spermatogenesis (DAZ and RBM, AZFa, b, and c) and other loci still poorly defined. No difference in localization of deletions was evident between cryptorchid and idiopathic patients. Microdeletions in Yq may be responsible for severe bilateral testicular damage that could be phenotypically expressed by unilateral cryptorchidism, as well as by idiopathic infertility. PMID- 10523012 TI - A critical evaluation of simple methods for the estimation of free testosterone in serum. AB - The free and nonspecifically bound plasma hormone levels generally reflect the clinical situation more accurately than total plasma hormone levels. Hence, it is important to have reliable indexes of these fractions. The apparent free testosterone (T) concentration obtained by equilibrium dialysis (AFTC) as well as the fraction of serum T not precipitated by 50% ammonium sulfate concentration (non-SHBG-T; SHBG, sex hormone-binding globulin), often referred to as bioavailable T, appear to represent reliable indexes of biologically readily available T, but are not well suited for clinical routine, being too time consuming. Several other parameters have been used without complete validation, however: direct immunoassay of free T with a labeled T analog (aFT), calculation of free T (FT) from total T and immunoassayed SHBG concentrations (iSHBG), and the free androgen index (FAI = the ratio 100T/iSHBG). In the view of substantial discrepancies in the literature concerning the free or bioavailable T levels, we compared AFTC, FT, aFT, FAI, and non-SHBG-T levels in a large number of sera with SHBG capacities varying from low, as in hirsute women, to extremely high as in hyperthyroidism. All these indexes of bioavailable T correlated significantly with the AFTC concentration; AFTC and FT values were almost identical under all conditions studied, except during pregnancy. Values for aFT, however, were only a fraction of either AFTC or FT, the fraction varying as a function of SHBG levels. Also, the FAI/AFTC ratio varied as a function of the SHBG levels, and hence, neither aFT nor FAI is a reliable index of bioavailable T. The FT value, obtained by calculation from T and SHBG as determined by immunoassay, appears to be a rapid, simple, and reliable index ofbioavailable T, comparable to AFTC and suitable for clinical routine, except in pregnancy. During pregnancy, estradiol occupies a substantial part of SHBG-binding sites, so that SHBG as determined by immunoassay overestimates the actual binding capacity, which in pregnancy sera results in calculated FT values that are lower than AFTC. The nonspecifically bound T, calculated from FT, correlated highly significantly with and was almost identical to the values of non-SHBG-T obtained by ammonium sulfate precipitation, testifying to the clinical value of FT calculated from iSHBG. PMID- 10523013 TI - Leptin and androgens in male obesity: evidence for leptin contribution to reduced androgen levels. AB - Leptin circulates in plasma at concentrations that parallel the amount of fat reserves. In obese males, androgen levels decline in proportion to the degree of obesity. Recently, we have shown that in rodent Leydig cells leptin inhibits hCG stimulated testosterone (T) production via a functional leptin receptor isoform; others have found that leptin inhibits basal and hCG-induced T secretion by testis from adult rats. In this study, we further investigated the relationship linking leptin and androgens in men. Basal and hCG-stimulated leptin and sex hormone levels were studied in a large group of men ranging from normal weight to very obese (body mass index, 21.8-55.7). Initial cross-sectional studies showed that circulating leptin and fat mass (FM) were inversely related with total and free T (r = -0.51 and r = -0.38, P < 0.01 and P < 0.05, respectively). Multiple regression analysis indicated that the correlation between leptin or FM and T was not lost after controlling for SHBG and/or LH and/or estradiol (E2) levels and that leptin was the best hormonal predictor of the lower androgen levels in obesity. Dynamic studies showed that in obese men the area under the curve of T and free T to LH/hCG stimulation (5000 IU i.m.) was 30-40% lower than in controls and inversely correlated with leptin levels (r = -0.45 and r = -0.40, P < 0.01 and P < 0.05, respectively). Also, LH/hCG-stimulation caused higher increases in 17-OH-progesterone to T ratio in obese men than in controls, whereas no differences were observed between groups either in stimulated E2 levels or in the E2/T ratio. In all subjects, the percentage increases from baseline in the 17-OH progesterone to T ratio were directly correlated with leptin levels or FM (r = 0.40 and r = 0.45, P < 0.01), but not with E2 or other hormonal variables. In conclusion, our studies, together with previous in vitro findings, indicate that excess of circulating leptin may be an important contributor to the development of reduced androgens in male obesity. PMID- 10523014 TI - Endogenous sex hormones and cognitive function in older men. AB - The objective of this study was to determine whether endogenous sex hormone levels predict cognitive function in older men. Our study design was an exploratory analysis in a population-based cohort in Rancho Bernardo, California. The study participants were 547 community-dwelling men 59-89 yr of age at baseline who were not using testosterone or estrogen therapy. Between 1984 and 1987, sera were collected for measurement of endogenous total and bioavailable testosterone and estradiol levels. Between 1988 and 1991, 12 standard neuropsychological instruments were administered, including two items from the Blessed Information-Memory-Concentration (BIMC) Test, three measures of retrieval from the Buschke-Fuld Selective Reminding Test, a category fluency test, immediate and delayed recall from the Visual Reproduction Test, the Mini-Mental State Examination with individual analysis of the Serial Sevens and the "World" Backwards components, and the Trail-Making Test Part B. In age- and education adjusted analyses, men with higher levels of total and bioavailable estradiol had poorer scores on the BIMC Test and Mini-Mental State Examination. Men with higher levels of bioavailable testosterone had better scores on the BIMC Test and the Selective Reminding Test (long-term storage). Five associations were U-shaped: total testosterone and total and bioavailable estradiol with the BIMC Test; bioavailable testosterone with the "World" test; and total estradiol with the Trail-Making Test. All associations were relatively weak but independent of age, education, body mass index, alcohol use, cigarette smoking and depression. In these older men, low estradiol and high testosterone levels predicted better performance on several tests of cognitive function. Linear and nonlinear associations were also found, suggesting that an optimal level of sex hormones may exist for some cognitive functions. PMID- 10523015 TI - Human leptin deficiency caused by a missense mutation: multiple endocrine defects, decreased sympathetic tone, and immune system dysfunction indicate new targets for leptin action, greater central than peripheral resistance to the effects of leptin, and spontaneous correction of leptin-mediated defects. AB - We have previously demonstrated that genetically based leptin deficiency due to a missense leptin gene mutation in a highly consanguineous extended Turkish pedigree is associated with morbid obesity and hypogonadism. We have now performed detailed assessments of endocrine, sympathetic, and immune function. We have also identified a new adult female homozygous patient in this extended family who is severely obese and amenorrheic. In this family all wild-type and heterozgous individuals have normal body weight. Seven obese members of this family, whom we presume to have been leptin deficient, died during childhood. There are several findings that indicate potentially novel targets for leptin action in humans. Four homozygous patients (1 adult male, 2 adult females, and 1 child) have sympathetic system dysfunction, whereas all heterozygous subjects have normal sympathetic system function. Despite sympathetic system dysfunction and postural hypotension, 1 of 3 homozygous adult patients has impaired renin aldosterone function. The patients also exhibit alterations in GH and PTH-calcium function, and 1 of them has decreased bone mineral density. Despite their obesity, these patients do not have risk factors for cardiovascular disease, such as hypertension, impairments in lipid metabolism, or hyperleptinemia [corrected]. These data support the hypothesis that the obese may have central, but not peripheral, resistance to the effects of leptin and that hyperleptinemia [corrected] may mediate the cardiovascular morbidity of the obese who are not leptin deficient. Furthermore, these data indicate that there may be several new targets for leptin action in human physiology. Such new targets may lead to novel pharmacological strategies for the use of leptin agonists and antagonists in the treatment of human disease. All 19 normal weight individuals in this family are alive, whereas 7 of 11 obese individuals died in childhood after infections. The odds ratio for mortality in the context of this obesity phenotype is 25.4, indicating that this mutation severely impairs key biological functions during childhood, negatively impacting on survival. We found that only the obese child in this family had thyroid function abnormalities. The oldest homozygous female patient started to menstruate, albeit with a luteal phase defect, 7 months ago, after a delay of over 20 yr, whereas the younger adult subjects are still hypogonadic. Thus, we conclude that due to their long life span, humans who survive the negative effects of leptin deficiency during childhood can, in contrast to ob/ob mice, over decades compensate some of the effects of leptin deficiency on immunity and endocrine function through mechanisms that remain to be elucidated. PMID- 10523016 TI - Recovery of hypopituitarism after neurosurgical treatment of pituitary adenomas. AB - Surgery is the treatment of choice for many pituitary tumors; pituitary function may suffer after operation, but relief of pressure on the normal pituitary may also favor postoperative recovery of hypopituitarism. The aim of this study was to investigate the frequency of new appearance and recovery of hypopituitarism after neurosurgery and try to identify features associated with it. Pre- and postoperative anterior pituitary functions were investigated in 234 patients with pituitary adenomas (56 nonfunctioning, 71 PRL-secreting, 66 GH-secreting, 39 ACTH secreting, 1 LH/FSH-secreting, and 1 TSH-secreting tumors). Eighty-eight new postoperative pituitary hypofunctions appeared in 52 patients (12 NF, 14 PRL secreting, 15 GH-secreting, 10 ACTH-secreting, and 1 LH/FSH-secreting adenomas). They corresponded to 27% ACTH deficiencies (in 29 of the 107 patients with normal preoperative ACTH in whom postoperative evaluation was complete), 14.5% (15 of 103) new GH deficiencies, 10.5% (15 of 143; P < 0.0005, significantly less than ACTH deficiency) new TSH deficiencies, 16.5% (20 of 121) new gonadotropin deficiencies, and 13% (9 of 71) new PRL deficiencies. Preoperatively, 93 were deficient in at least 1 pituitary hormone; after surgery, 45 (48%) recovered between 1 and 3 hormones. The 2 patients with LH/FSH- and TSH-secreting macroadenomas did not recover pituitary function. Factors associated with a higher probability of postoperative pituitary function recovery were: no tumor rests on postoperative pituitary imaging (P = 0.001) and no neurosurgical (P = 0.001) or pathological evidence (P = 0.049) of an invasive nature. Tumor size did not differ significantly between those who did and those who did not recover pituitary function after surgery. Even if clear hypofunction is observed at initial work-up, patients should be reassessed after surgery without substitution therapy, because practically half the preoperative pituitary hormone deficiencies recover postoperatively, eliminating the need for life-long substitution therapy. PMID- 10523017 TI - Microsatellite polymorphism of the MHC class I chain-related (MIC-A and MIC-B) genes marks the risk for autoimmune Addison's disease. AB - The major histocompatibility complex class I chain-related MIC-A and MIC-B genes are located on chromosome 6 between the histocompatibility leucocyte antigen (HLA)-B and the B-associated transcript genes. The presence of 21-hydroxylase autoantibodies is a sensitive and specific marker of autoimmune Addison's disease. We studied the polymorphism of exon 5 of the MIC-A gene, of intron 1 of the MIC-B gene, and of HLA-DRB1, -DQA1, and -DQB1 genes in 28 autoimmune (21 hydroxylase autoantibody positive) Addison's disease patients and in 75 healthy subjects from central Italy. The MIC-A5.1 allele was significantly more frequent in Addison's disease patients (79%) than in healthy subjects (36%) [odds ratio (OR) = 6.52, corrected P (Pc) = 0.0015], whereas MIC-A6 was significantly reduced in affected subjects (15% vs. 56%, OR = 0.13, Pc = 0.002). The A5.1/A5.1 genotype had an OR for autoimmune Addison's disease as high as 18.0 and an absolute risk of 1 per 1131. In the presence of MIC-A5.1, MICB-CA-25 was significantly increased in Addison's disease patients (25% vs. 4%, OR = 8.0, P = 0.0039, Pc = 0.047). The MICB-CA-17 allele was absent in Addison's disease patients, but present in more than 25% healthy individuals (OR = 0.10, P = 0.0025, Pc = 0.03). Among HLA-DR and -DQ haplotypes, only DRB1*03-DQA1*0501-DQB1*0201 (DR3/DQ2) was significantly more frequent in Addison's disease patients than in healthy subjects, but only in the presence of MIC-A5.1. The frequency of MIC-A5.1 was significantly increased in Addison's disease patients only in the presence of HLA DR3-DQ2. Our study demonstrates that susceptibility to autoimmune Addison's disease is linked to the MIC-A microsatellite allele 5.1 and that both MIC-A5.1 and HLA-DR3/DQ2 are necessary to confer increased genetic risk for Addison's disease. PMID- 10523018 TI - Two polymorphisms in the peroxisome proliferator-activated receptor-gamma gene are associated with severe overweight among obese women. AB - Peroxisome proliferator-activated receptor-gamma (PPARgamma) is a nuclear receptor that regulates adipocyte differentiation. Variations in the PPARgamma gene may affect the function of the PPARgamma and, therefore, body adipocity. We investigated the frequencies of the Pro12Ala polymorphism in exon B and the silent CAC478CAT polymorphism in exon 6 of the PPARgamma gene and their effects on body weight, body composition, and energy expenditure in obese Finns. One hundred and seventy obese subjects [29 men and 141 women; body mass index (BMI), 35.7 +/- 3.8 kg/m2; age, 43 +/- 8 yr; mean +/- SD) participated in the study. The frequencies of the Ala12 allele in exon B and CAT478 allele in exon 6 were not significantly different between the obese and population-based control subjects (0.14 vs. 0.13 and 0.19 vs. 0.21, respectively). The polymorphisms were associated with increased BMI [Pro12Pro, 34.5 +/- 3.8; Pro12Ala, 34.8 +/- 3.1; Ala12Ala, 39.2 +/- 4.6 kg/m2 (P = 0.011); CAC478CAC, 34.5 +/- 3.8; CAC478CAT, 34.5 +/- 3.3; CAT478CAT, 37.7 +/- 4.1 kg/m2 (P = 0.046)]. In addition, the women with both Ala12Ala and CAT478CAT genotypes (n = 5) were significantly more obese compared with the women having both Pro12Pro and CAC478CAC genotypes (n = 85; BMI, 40.6 +/- 3.3 vs. 34.4 +/- 3.9 kg/m2; P = 0.001), and they had increased fat mass (46.8 +/- 9.1 vs. 36.8 +/- 7.5 kg; P = 0.005). In conclusion, the Pro12Ala and CAC478CAT polymorphisms in the PPARgamma gene are associated with severe overweight and increased fat mass among obese women. PMID- 10523019 TI - A frame-shift mutation in the type I parathyroid hormone (PTH)/PTH-related peptide receptor causing Blomstrand lethal osteochondrodysplasia. AB - Blomstrand osteochondrodysplasia (BOCD) is a rare lethal skeletal dysplasia characterized by accelerated endochondral and intramembranous ossification. Comparison of the characteristics of BOCD with type I PTH/PTH-related peptide (PTHrP) receptor-ablated mice reveals striking similarities that are most prominent in the growth plate. In both cases, the growth plate is reduced in size due to a strongly diminished zone of resting cartilage and the near absence of columnar arrangement of proliferating chondrocytes. This overall similarity suggested that an inactivating mutation of the PTH/PTHrP receptor might be the underlying genetic defect causing BOCD. Indeed, inactivating mutations of the PTH/PTHrP receptor have been recently identified in two cases of BOCD. We describe here a novel inactivating mutation in the PTH/PTHrP receptor. Sequence analysis of all coding exons of the type I PTH/ PTHrP receptor gene and complementary DNA of a case with BOCD identified a homozygous point mutation in exon EL2 in which one nucleotide (G at position 1122) was absent. The mutation was inherited from both parents, supporting the autosomal recessive nature of the disease. The missense mutation resulted in a shift in the open reading frame, leading to a truncated protein that completely diverged from the wild-type sequence after amino acid 364. The mutant receptor, therefore, lacked transmembrane domains 5, 6, and 7; the connecting intra- and extracellular loops; and the cytoplasmic tail. Functional analysis of the mutant receptor in COS-7 cells and of dermal fibroblasts obtained from the case proved that the mutation was indeed inactivating. Neither the transiently transfected COS-7 cells nor the dermal fibroblasts responded to a challenge with PTH or PTHrP with a rise in intracellular cAMP levels, in sharp contrast to control cells. Our results provide further evidence that BOCD is caused by inactivating mutations of the type I PTH/PTHrP receptor and underscore the importance of this receptor in mammalian skeletal development. PMID- 10523020 TI - Human leukocyte antigen-A24 and -DQA1*0301 in Japanese insulin-dependent diabetes mellitus: independent contributions to susceptibility to the disease and additive contributions to acceleration of beta-cell destruction. AB - The aim of this study is to identify insulin-dependent diabetes mellitus (IDDM) susceptible HLA antigens in IDDM patients who do not have established risk allele, HLA-DQA1*0301, and analyze relationship of these HLA antigens and the degree of beta-cell destruction. In 139 Japanese IDDM patients and 158 normal controls, HLA-A, -C, -B, -DR and -DQ antigens were typed. Serum C-peptide immunoreactivity response (deltaCPR) to a 100-g oral glucose load < or = 0.033 nmol/l was regarded as complete beta-cell destruction. All 14 patients without HLA-DQA1*0301 had HLA-A24, whereas only 35 of 58 (60.3%) normal controls without HLA-DQA1*0301 and only 72 of 125 (57.6%) IDDM patients with HLA-DQA1*0301 had this antigen (Pc = 0.0256 and Pc = 0.0080, respectively). DeltaCPR in IDDM patients with both HLA-DQA1*0301 and HLA-A24 (0.097 +/- 0.163 nmol/L, mean +/- SD, n = 65) were lower than in IDDM patients with HLA-DQA1*0301 only (0.219 +/- 0.237 nmol/L, n = 45, P < 0.0001) and in IDDM patients with HLA-A24 only (0.187 +/- 0.198 nmol/L, n = 14, P = 0.0395). These results indicate that both HLA DQA1*0301 and HLA-A24 contribute susceptibility to IDDM independently and accelerate beta-cell destruction in an additive manner. PMID- 10523021 TI - Leptin response to insulin in humans is related to the lipolytic state of abdominal subcutaneous fat. AB - Insulin-induced leptinemia in humans appears to be blunted by insulin resistance. We therefore examined the relationship between insulin action and plasma leptin by monitoring regional and whole body lipolysis and plasma leptin levels in 15 premenopausal women (body fat range, 14-59%) during a two-stage euglycemic clamp (insulin was infused 90 min each at 6-10 and 12-20 mU/m2 x min). Microdialysis probes were placed in abdominal and femoral sc adipose tissue. Subjects were given a primed, constant infusion of a stable isotope tracer (2H5-glycerol), and plasma glycerol isotope enrichments were analyzed by mass spectrometry to determine glycerol kinetics. Although there was no mean change in plasma leptin during the clamp (baseline, 16.6 +/- 4.5 ng/mL; final, 16.3 +/- 4.3 ng/mL), there was large interindividual variability in the changes in plasma leptin (range, 18% to +19%). Changes in plasma leptin during the clamp stages were correlated with abdominal dialysate glycerol concentrations (r = -0.44; P < 0.05), but not femoral dialysate glycerol concentrations (r = -0.15), the rate of appearance of glycerol in plasma (r = 0.005), or plasma insulin levels (r = 0.16). The results suggest that insulin-induced changes in plasma leptin are more related to the lipolytic state (i.e. low leptin response when lipolysis is high) of abdominal sc adipose tissue than that of other fat depots. PMID- 10523022 TI - Corticotropin-releasing hormone stimulates P450 17alpha-hydroxylase/17,20-lyase in human fetal adrenal cells via protein kinase C. AB - CRH directly stimulates dehydroepiandrosterone sulfate (DHEAS) production in human fetal adrenal cells. In the human fetal and adult pituitary, CRH acts via protein kinase A (PKA). We determined the CRH signal transduction pathway in fetal adrenal cells, i.e. whether CRH modulates human fetal adrenal steroidogenesis via PKA and/or protein kinase C (PKC). In primary cultures, CRH increased inositol trisphosphate. After CRH treatment, inositol tris-, bis-, and monophosphates increased within 1 min, reaching maximal levels at 5 min. In contrast, PGF2alpha, known to act via PKC, induced a sustained response for up to 20 min. The response to CRH was dose dependent, maximal at 1 micromol/L at both 1 and 5 min. CRH increased DHEAS production, with a much lesser effect on cortisol. CRH did not stimulate inositol phospholipid in adult adrenal glands, suggesting that this pathway is unique to the fetal adrenal. CRH increased messenger ribonucleic acid encoding 17alpha-hydroxylase/17,20 lyase (P450c17), but not 3beta-hydroxysteroid dehydrogenase/delta(4-5) isomerase. However, 3betaHSD expression was stimulated by ACTH. PKC, but not PKA, inhibitors blocked CRH stimulated P450c17 induction, whereas PKA inhibitors blocked ACTH-stimulated cortisol. Thus, CRH is coupled to the phospholipase C-inositol phosphate second messenger system and preferentially induces the expression of P450c17 and DHEAS, suggesting a unique role of CRH regulating human fetal adrenal function via PKC. PMID- 10523023 TI - Regulatory effects of 1alpha,25-dihydroxyvitamin D3 on the cytokine production of human peripheral blood lymphocytes. AB - We studied the possible regulatory effects of 1alpha,25-dihydroxyvitamin D3 [1alpha,25-(OH)2D3] on cytokine production and differentiation of subsets of CD4+ [T helper 1 (Th1) and Th2] and CD8+ [T cytotoxic 1 (Tc1) and Tc2] lymphocytes at the single cell level. PBMC from healthy donors were cultured with or without 1alpha,25-(OH)2D3 for up to 21 days. On days 0, 7, 14, and 21, the percentage of cytokine-producing T lymphocytes was analyzed by intracellular cytokine detection with mAb and flow cytometry. Simultaneous staining for cell surface markers allowed discrimination of CD4+ and CD8+ T cell subsets. After culture with 1alpha,25-(OH)2D3 (10(-8) mol/L), no significant effects on the proportion of interferon-gamma (IFNgamma)- or interleukin-4 (IL-4)-producing cells were detected, whereas reduced frequencies of IL-2-producing cells in the CD4+ as well as in the CD8+ population were found. An increase in the low percentage of CD4+ and CD8+ T cells producing the Th2 cytokine IL-13 was noticed. Most interestingly, IL-6-producing CD4+ and CD8+ T cells could only be detected in cultures with 1alpha,25-(OH)2D3, reaching a plateau after 14 days. The percentage of IL-6-producing T cells induced by 1alpha,25-(OH)2D3 after a given time period remained stable for at least 7 weeks. Studies of cytokine coexpression revealed that about 70% of IL-6-producing CD4+ and CD8+ cells were also positive for IL-2, but more than 90% were negative for IFNgamma, IL-4, or IL-13, respectively. This suggests that the IL-6-producing population does not match the Th1/Tc1-like (IFNgamma+) or Th2/Tc2-like (IL-4+ or IL-13+) subset. The influence of 1alpha,25 (OH)2D3 on cytokine production by lymphocytes is probably an important point of intersection between the endocrine and the immune system. PMID- 10523025 TI - Linkage of familial euthyroid goiter to the multinodular goiter-1 locus and exclusion of the candidate genes thyroglobulin, thyroperoxidase, and Na+/I- symporter. AB - Iodine deficiency is the most important etiological factor for euthyroid endemic goiter. However, family and twin pair studies also indicate a genetic predisposition for euthyroid simple goiter. In hypothyroid goiters several molecular defects in the thyroglobulin (TG), thyroperoxidase (TPO), and Na+/I- symporter (NIS) genes have been identified. The TSH receptor with its central role for thyroid function and growth is also a strong candidate gene. Therefore, we investigated a proposita with a relapsing euthyroid goiter and her family, in which several members underwent thyroidectomy for euthyroid goiter. Sequence analysis of the complementary DNA (cDNA) of the TPO and TSH receptor genes revealed several previously reported polymorphisms. As it is not possible to exclude a functional relevance for all polymorphisms, we opted for linkage analysis with microsatellite markers to investigate whether the candidate genes are involved in the pathogenesis of euthyroid goiter. The markers for the genes TG, TPO, and NIS gave two-point and multipoint logarithm of odds score analysis scores that were negative or below 1 for all assumed recombination fractions. As no significant evidence of linkage was found, we conclude that these candidate genes can be excluded as a major cause of the euthyroid goiters in this family. In contrast, we have found evidence for linkage of familial euthyroid goiter to the recently identified locus for familial multinodular nontoxic goiter (MNG-1) on chromosome 14q. The haplotype cosegregates clearly with familial euthyroid goiter. Our results provide the first confirmation for MNG-1 as a locus for nontoxic goiter. PMID- 10523024 TI - Molecular basis of human salt sensitivity: the role of the 11beta-hydroxysteroid dehydrogenase type 2. AB - Salt-sensitive subjects (SS) increase their blood pressure with increasing salt intake. Because steroid hormones modulate renal sodium retention, we hypothesize that the activity of the 11beta-hydroxysteroid dehydrogenase type 2 (11betaHSD2) enzyme is impaired in SS subjects as compared with salt-resistant (SR) subjects. The 11betaHSD2 enzyme inactivates 11-hydroxy steroids in the kidney, thus protecting the nonselective mineralocorticoid receptor from occupation by glucocorticoids. We performed an association study using a recently identified single AluI polymorphism in exon 3 and a polymorphic microsatellite marker of the HSD11B2 gene in 149 normotensive white males (37 SS and 112 SR). The activity of the enzyme 11betaHSD2 was assessed by determining the urinary ratio of cortisol (THF+5alphaTHF) to cortisone (THE) metabolites by gas chromatography in all the 37 SS subjects and in 37 age- and body habitus-matched SR volunteers. Mean (THF+5alphaTHF)/THE ratio was markedly elevated in SS subjects compared with SR subjects (1.51 +/- 0.34 vs. 1.08 +/- 0.26, P < 0.00001), indicating enhanced access of glucocorticoids to the mineralocorticoid receptor in SS subjects. In 58% of SS subjects this ratio was higher than the maximum levels in SR subjects. The salt-induced elevation in arterial pressure increased with increasing (THF+5alphaTHF)/THE ratio (r2 = 0.51, P < 0.0001). A total of 12 alleles of the polymorphic microsatellite marker were detected. Homozygosity for the allele A7 was higher in SS subjects than in SR subjects (41 vs. 28%, P < 0.005), whereas the occurrence of the allele A7 with allele A8 was lower in SS subjects than in SR subjects (8 vs. 15%, P < 0.03). The prevalence of salt sensitivity was 35% in subjects with allele A7/A7, whereas salt sensitivity was present in only 9% of the subjects with allele A7/A8. The (THF+5alphaTHF)/THE ratio was higher in subjects homozygous for the A7 microsatellite allele as compared with the corresponding control subjects. The prevalence of the AluI allele was 8.0% in SR subjects and 5.4% in SS subjects and did not correlate with blood pressure. The decreased activity of the 11betaHSD2 in SS subjects indicates that this enzyme is involved in salt-sensitive blood pressure response in humans. The association of a polymorphic microsatellite marker of the gene with a reduced 11betaHSD2 activity suggests that variants of the HSD 11B2 gene contribute to enhanced blood pressure response to salt in humans. PMID- 10523026 TI - Daily treatment with human recombinant parathyroid hormone-(1-34), LY333334, for 1 year increases bone mass in ovariectomized monkeys. AB - PTH stimulates bone formation to increase bone mass and strength in rats and humans. The aim of this study was to determine the skeletal effects of recombinant human PTH-(1-34) [rhPTH-(1-34)] in monkeys, as monkey bone remodeling and structure are similar to those in human bone. Adult female cynomolgus monkeys were divided into sham-vehicle (n = 21), ovariectomized (OVX)-vehicle (n = 20), and OVX groups given daily s.c. injections of rhPTH-(1-34) at 1 (n = 39) or 5 (n = 41) microg/kg for 12 months. Whole body bone mineral content was measured, as was bone mineral density (BMD) in the spine, proximal tibia, midshaft radius, and distal radius. Serum and urine samples were also analyzed. rhPTH-(1-34) treatment did not influence serum ionized Ca levels or urinary Ca excretion, but depressed endogenous PTH while increasing serum calcitriol levels. Compared to that in the OVX group, the higher dose of rhPTH-(1-34) increased spine BMD by 14.3%, whole body bone mineral content by 8.6%, and proximal tibia BMD by 10.8%. Subregion analyses suggested that the anabolic effect of rhPTH-(1-34) on the proximal tibia was primarily in cancellous bone. Similar, but less dramatic, effects on BMD were observed with the lower dose of rhPTH-(1-34). Daily s.c. rhPTH-(1-34) treatment for 1 yr increases BMD in ovariectomized monkeys without inducing sustained hypercalcemia or hypercalciuria. PMID- 10523027 TI - Beta-adrenergically stimulated fat oxidation is diminished in middle-aged compared to young subjects. AB - The effect of aging on beta-adrenergically mediated substrate utilization was investigated in nine young (25.2 +/- 1.7 yr old) and eight older males (52.9 +/- 2.1 yr old), matched for body weight and body composition. In a first experiment, the nonselective beta-agonist isoprenaline (ISO) was infused in increasing standardized doses, and during each infusion period energy expenditure and substrate utilization were determined by indirect calorimetry. In a second experiment, forearm skeletal muscle metabolism was studied during a standardized infusion dose of ISO (19 ng/kg fat-free mass x min). During beta-adrenergic stimulation there was an increased carbohydrate oxidation (at an ISO infusion dose of 24 ng/kg fat-free mass x min, 31% vs. 21% of total energy expenditure; P < 0.05) and a decreased fat oxidation (51 vs. 62 of total energy expenditure; P < 0.05) in older compared to young subjects. Skeletal muscle lactate release significantly increased in the older subjects (from -175 +/- 32 to -366 +/- 66 nmol/100 mL forearm tissue x min), whereas there was no change in young subjects (from -32 +/- 21 to 23 +/- 57 nmol/100 mL forearm tissue x min; interaction group x ISO, P < 0.01). Additionally, there was a tendency toward a blunted ISO-induced increase in nonesterified fatty acid uptake in the older subjects (interaction group x ISO, P = 0.062). Thus, middle-aged subjects have a blunted ability to oxidize fat during beta-adrenergic stimulation compared to young subjects. This diminished fat oxidation may be an important etiological factor in the age related increase in body fatness and obesity by favoring fat storage above oxidation. PMID- 10523028 TI - Influence of low density lipoprotein from insulin-dependent diabetic patients on platelet functions. AB - In the present work we studied in vitro the action of low density lipoproteins (LDL) isolated from normolipemic insulin-dependent diabetic (IDDM) patients on transmembrane cation transport, nitric oxide synthase (NOS) activity, and aggregating response to stimuli of platelets from healthy subjects to elucidate whether the modified interaction between circulating lipoproteins and cells might be one of the pathogenetic mechanisms of the increased platelet activation in IDDM. LDL were obtained by discontinuous gradient ultracentrifugation from 15 IDDM out-patients and 15 sex- and age-matched healthy subjects and used for incubation experiments with control platelets. Lipid composition and hydroperoxide concentrations were studied in LDL. Platelet aggregation responses to ADP, NOS activity, cytosolic Ca2+ concentrations, and platelet membrane Na+/K+ adenosine triphosphatase (Na+/K+-ATPase) and Ca2+-ATPase activities were measured after incubation. IDDM LDL showed an increased lysophosphatidylcholine content compared with that of control LDL. IDDM LDL significantly increased the platelet aggregating response to ADP, cytosolic Ca2+ concentrations, and plasma membrane Ca2+-ATPase activity and significantly reduced NOS activity and platelet membrane Na+/K+-ATPase activity compared with those of platelets incubated in buffer or cells incubated with control LDL. The effects exerted by IDDM LDL on platelet suspensions from healthy subjects mimic the alterations observed in platelets from diabetic subjects in basal conditions. Both the decreased activity of NOS and the higher cytoplasmic concentrations of Ca2+ might cause increased platelet activation, as observed in IDDM. In conclusion, the present study suggests a new mechanism with a potential role in the early development of atherosclerosis in diabetic patients, i.e. an altered interaction between circulating lipoproteins and platelets. PMID- 10523029 TI - Alternative genetic pathways in parathyroid tumorigenesis. AB - In this study 44 parathyroid tumors from 26 sporadic cases, 10 cases previously given irradiation to the neck, and 8 familial cases were screened for sequence copy number alterations by comparative genomic hybridization. In the sporadic adenomas, commonly occurring minimal regions of loss could be defined to chromosome 11 (38%), 15q15-qter (27%), and 1p34-pter (19%), whereas gains preferentially involved 19p13.2-pter (15%) and 7pter-qter (12%). Multiple aberrations were found in sporadic tumors with a somatic mutation and/or loss of heterozygosity of the MEN1 gene. The irradiation-associated tumors also showed multiple comparative genomic hybridization alterations and frequent losses of 11q (50%), and subsequent analysis of the MEN1 gene demonstrated mutations in 4 of 8 cases (50%). The adenomas from familial cases showed few alterations, and in 3 of these tumors a gain of 19p13.2-pter was seen as the only aberration. In this study numerical copy number alterations were frequently detected in sporadic and irradiation-associated parathyroid adenomas, although these tumors are benign. The majority of these alterations were found in tumors with confirmed involvement of the MEN1 gene locus in agreement with a role of the MEN1 gene in genomic stability. Furthermore, the frequent occurrence of MEN1 mutations (50%) in irradiation-associated parathyroid tumors suggests that inactivation of the MEN1 gene is an important genetic alteration involved in the development of parathyroid tumors in postirradiation patients. PMID- 10523031 TI - A novel mutation of the signal peptide of the preproparathyroid hormone gene associated with autosomal recessive familial isolated hypoparathyroidism. AB - We report a novel mutation of the signal peptide of the prepro-PTH gene associated with autosomal recessive familial isolated hypoparathyroidism. The proposita presented with neonatal hypocalcemic seizures. Serum calcium was 1.5 mmol/L (normal, 2.0-2.5); phosphate was 3.6 mmol/L (normal, 0.9-1.5). She was born to consanguineous parents. A few years later, 2 younger sisters and her niece presented with neonatal hypocalcemic seizures. Their intact PTH levels were undetectable during severe hypocalcemia. Genomic DNA from the proposita was sequenced all exons of the prepro-PTH gene. A replacement of thymine with a cytosine was found in the first nucleotide of position 23 in the 25-amino acid signal peptide. This results in the replacement of the normal Ser (TCG) with a Pro (CCG). Genotyping of family members was carried out by identification of a new MspI site created by the mutation. Only affected family members were homozygous for the mutant allele, whereas the parents were heterozygous, supporting autosomal recessive inheritance. As this mutation is at the -3 position in the signal peptide of the prepro-PTH gene, we hypothesized that the prepro-PTH mutant might not be cleaved by signal peptidase at the normal position, and it might be degraded in rough endoplasmic reticulum. PMID- 10523030 TI - Activation of central neuropeptide Y Y1 receptors potently stimulates food intake in male rhesus monkeys. AB - The orexigenic role of central neuropeptide Y (NPY) in nonhuman primates has been questioned. Therefore, we have studied the effect of central NPY on feeding in ad libitum-fed male rhesus macaques. NPY dose-dependently increased food intake, with the maximal effect obtained by 50 microg (960 min food intake +/- SEM, 104 +/- 5 to 188 +/- 11 g; vehicle vs. NPY; n = 6). Blood glucose levels were unaffected by intracerebroventricular administration of NPY, but animals receiving either 20 or 50 microg displayed increased plasma levels of insulin and cortisol at few time points. To assess the pharmacological specificity of this response, a novel Y1 antagonist, [(Ile,Glu,Pro,Daba,Tyr,Arg,Leu,Arg,Tyr-NH2)2 cyclic (2,4'),(2',4)-diamide] (Y1ANT), was synthesized. Receptor binding experiments demonstrated that Y1ANT preferentially binds to Y1 and Y4 receptors (pKi 10.12 +/- 0.06 and 9.11 +/- 0.05 nmol/L, respectively). Functional analysis revealed that Y1ANT is a Y1 antagonist and a partial Y4 agonist. Central administration of Y1ANT blocked NPY-induced feeding. In food-deprived monkeys, Y1ANT attenuated the feeding response. However, Y1ANT had no effect on food intake in satiated monkeys. Thus, endogenous NPY is likely to be involved in the regulation of food intake in the nonhuman primate, and this effect is at least partially mediated via Y1-like receptors. PMID- 10523033 TI - Evidence for genetic heterogeneity in male pseudohermaphroditism due to Leydig cell hypoplasia. AB - Leydig cell aplasia or hypoplasia is a rare form of male pseudohermaphroditism resulting from inadequate fetal testicular Leydig cell differentiation. Affected individuals presented a wide phenotypic spectrum, ranging from complete female external genitalia to males with micropenis. Recessive mutations in the LH receptor gene have been identified as responsible for the condition. The majority of these mutations are point mutations and have been located in exon 11 of the gene. In this study, we report the molecular characterization of the LH receptor gene in three siblings with Leydig cell hypoplasia. After sequencing the 11 exons of the gene, no deleterious mutations were detected in any patient. However, we identified a previously described polymorphism in exon 11. In patients 1 and 3 DNA sequencing revealed a C to T substitution at nucleotide 1065; both patients were homozygous GAT/GAT at codon 355. In contrast, patient 2 was homozygous GAC/GAC, whereas the father and one unaffected sister were heterozygous GAC/GAT at this polymorphic site. These results exclude that Leydig cell hypoplasia in this family is due to a mutation in the LH receptor gene and provide evidence that defects in other loci may also result in failure of Leydig cell differentiation, demonstrating, for the first time, that Leydig cell hypoplasia is a genetically heterogeneous condition. PMID- 10523032 TI - The interaction of TSH receptor autoantibodies with 125I-labelled TSH receptor. AB - Detergent-solubilized porcine TSH receptor (TSHR) has been labeled with 125I using a monoclonal antibody to the C-terminal domain of the receptor. The ability of sera containing TSHR autoantibody to immunoprecipitate the labeled receptor was then investigated. Sera negative for TSHR autoantibody (as judged by assays based on inhibition of labeled TSH binding to detergent-solubilized porcine TSHR) immunoprecipitated about 4% of the labeled receptor, whereas sera with high levels of receptor autoantibody immunoprecipitated more than 25% of the labeled receptor. The ability to immunoprecipitate labeled TSHR correlated well with ability of the sera to inhibit labeled TSH binding to the receptor (r = 0.92; n = 63), and this is consistent with TSHR autoantibodies in these samples being directed principally to a region of the receptor closely related to the TSH binding site. Preincubation of labeled TSHR with unlabeled TSH before reaction with test sera inhibited the immunoprecipitation reaction, providing further evidence for a close relationship between the TSHR autoantibody binding site(s) and the TSH binding site. This was the case whether the sera had TSH agonist (i.e., thyroid stimulating) or TSH antagonist (i.e., blocking) activities, thus, providing no clear evidence for different regions of the TSHR being involved in forming the binding site(s) for TSHR autoantibodies with stimulating and with blocking activities. The ability of TSHR autoantibodies to stimulate cyclic AMP production in isolated porcine thyroid cells was compared with their ability to immunoprecipitate labeled porcine TSHR. A significant correlation was observed (r = 0.58; n = 50; P < 0.001) and the correlation was improved when stimulation of cyclic AMP production was compared with inhibition of labeled TSH binding to porcine TSHR (r = 0.76). Overall, our results indicate that TSHR autoantibodies bind principally to a region on the TSHR closely related to the TSH binding site, and this seems to be the case whether the autoantibodies act as TSH agonists or antagonists. PMID- 10523034 TI - Analysis of meiosis in intratesticular germ cells from subjects affected by classic Klinefelter's syndrome. AB - Azoospermic subjects affected by Klinefelter's syndrome may occasionally show the presence of intratesticular residual foci of spermatogenesis, and the retrieval of mature spermatozoa from the testis may permit fertility and paternity by means of intracytoplasmic sperm injection. Previous studies have demonstrated that these subjects show the presence of an increased incidence of hyperaploid spermatozoa. Here we analyzed, by fluorescence in situ hybridization using specific probes for chromosomes 8, X, and Y, the spermatogenic process and the meiotic progression of 47,XXY germ cells retrieved by fine needle aspiration of the testis in ten azoospermic patients affected by classic Klinefelter's syndrome. All patients had lower testicular volume, higher gonadotropins, and lower testosterone plasma levels compared with control subjects. Cytological analysis of the testicular cells retrieved by fine needle aspiration showed the presence of Sertoli cells only in eight subjects, while germ cells were observed in two patients. In each patient Sertoli cells showed a 47,XXY karyotype, and the same chromosome pattern was observed in spermatogonia and primary spermatocytes of patients presenting a residual spermatogenesis. Secondary spermatocytes, spermatids, and mature spermatozoa showed different sex chromosome patterns, reflecting their origin from 47,XXY spermatogonia. In conclusion, this study demonstrated that, in subjects affected by Klinefelter's syndrome, residual germ cells may be present in the testis and that 47,XXY spermatogonia are able to undergo and complete the spermatogenic process leading to mature spermatozoa. These data further suggest the need to evaluate the sex chromosome status of sperm from patients affected by Klinefelter's syndrome undergoing assisted reproductive techniques. PMID- 10523035 TI - Complete hypogonadotropic hypogonadism associated with a novel inactivating mutation of the gonadotropin-releasing hormone receptor. AB - In this study, we describe a patient with a phenotype of complete hypogonadotropic hypogonadism who presented primary failure of pulsatile GnRH therapy, but responded to exogenous gonadotropin administration. This patient bore a novel point mutation (T for A) at codon 168 of the gene encoding the GnRH receptor (GnRH-R), resulting in a serine to arginine change in the fourth transmembrane domain of the receptor. This novel mutation was present in the homozygous state in the patient, whereas it was in the heterozygous state in both phenotypically normal parents. When introduced into the complementary DNA coding for the GnRH-R, this mutation resulted in the complete loss of the receptor mediated signaling response to GnRH. In conclusion, we report the first mutation of the GnRH-R gene that can induce a total loss of function of this receptor and is associated with a phenotype of complete hypogonadotropic hypogonadism. PMID- 10523036 TI - Food-dependent Cushing's syndrome: possible involvement of leptin in cortisol hypersecretion. AB - Stimulation ofcortisol secretion by food intake has been implicated in the pathogenesis of some cases of ACTH-independent Cushing's syndrome, via an aberrant response of the adrenal glands to gastric inhibitory polypeptide (GIP). We report here a novel case of food-dependent Cushing's syndrome in a patient with bilateral macronodular adrenal hyperplasia. In this patient we were able to confirm a paradoxical stimulation of cortisol secretion by GIP in vivo as well as in vitro on dispersed tumor adrenal cells obtained at surgery. In addition to GIP, in vitro stimulation of these cultured tumor adrenal cells with leptin, the secreted product of the adipocyte, induced cortisol secretion. By comparison, no such stimulation was observed in vitro in adrenal cells obtained from another patient with bilateral macronodular adrenal hyperplasia and Cushing's syndrome that did not depend on food intake, in tumor cells obtained from a solitary cortisol-secreting adrenal adenoma, and in normal human adrenocortical cells. These results demonstrate that as in previously described cases of food-dependent Cushing's syndrome, GIP stimulated cortisol secretion from the adrenals of the patient reported here. Therefore, they indicate that such a paradoxical response probably represents the hallmark of this rare condition. In addition, they suggest that leptin, which normally inhibits stimulated cortisol secretion in humans, participated in cortisol hypersecretion in this case. Further studies in other cases of food-dependent Cushing's syndrome, however, will be necessary to better ascertain the pathophysiological significance of this finding. PMID- 10523037 TI - Low expression of the cell cycle inhibitor p27Kip1 in normal corticotroph cells, corticotroph tumors, and malignant pituitary tumors. AB - The cell cycle is regulated by a number of inhibitors, including p27Kip1 (p27), which belongs to the kip1 family. By binding to the cyclin/cyclin-dependent kinase complexes, it regulates progression of G1 to S phase in the cell cycle. It has been reported that p27 knockout mice develop multiorgan hyperplasia and intermediate lobe pituitary tumors secreting ACTH. Previously, we and others have been unable to show any consistent change in messenger RNA expression or genomic mutations for p27 in human corticotroph adenomas. However, dysregulation at the protein level has been reported in nonendocrine tumors, and we, therefore, investigated the expression of p27 in a range of benign and metastatic pituitary tumors. We studied a total of 107 pituitaries, including normal pituitary (n = 20), Cushing's disease (n = 21), acromegaly (n = 19), nonfunctioning adenomas (n = 18), prolactinomas (n = 7), TSH-omas (n = 2), FSH-omas (n = 6), aggressive tumors showing invasiveness and recurrence (n = 9), and metastatic pituitary carcinomas (n = 5). Using standard immunohistochemical techniques with a highly specific monoclonal antibody, p27 expression was determined quantitatively as the percentage of cells showing strongly positive, weak, or negative staining. In each sample, approximately 500 cells were analyzed. We also analyzed normal pituitaries using double-labeling for p27 and each of the pituitary hormones to characterize the expression of p27 in each cell type. p27 was expressed in normal pituitary cells; in tumors expressing GH, prolactin, TSH, and FSH; and in aggressive tumors, but markedly reduced expression of p27 was seen in corticotroph tumors and pituitary carcinomas. In the normal pituitary, somatotroph, lactotroph, and thyrotroph cells showed strong p27 staining, whereas normal corticotroph cells showed a much lower level of p27 staining (P < 0.001). Somatotroph, lactotroph, gonadotroph, and thyrotroph adenomas showed a lower level of p27 expression compared with normal somatotrophs (P = 0.02), lactotrophs (P = 0.03), gonadotrophs (P = 0.01), and thyrotrophs, respectively, whereas the lower level of p27 expression present in normal corticotrophs virtually disappeared in corticotroph adenomas (P = 0.001). We conclude that pituitary adenomas show a lower level of p27 protein expression than the normal cells from which they are derived, with malignant transformation leading to complete loss of p27 immunoreactivity. Corticotrophs are quite different to other pituitary cell types in terms of p27 immunoreactivity because both normal and tumorous corticotrophs have low p27 staining, and we speculate that this may relate to their inherent control mechanisms. PMID- 10523038 TI - Development of the baboon fetal adrenal gland: regulation of the ontogenesis of the definitive and transitional zones by adrenocorticotropin. AB - Throughout gestation, the primate fetal adrenal gland is comprised of the fetal zone, which expresses the P-450 17alpha-hydroxylase-C17,20 lyase (P-450c17) enzyme that catalyzes the synthesis of C19 steroids used for placental estrogen production. The development of the transitional zone comprised of cortical cells that express the P-450c17 and the 3beta-hydroxysteroid dehydrogenase-isomerase (3betaHSD) enzymes for cortisol production, and the definitive zone, which expresses 3betaHSD, but not P-450c17, for mineralocorticoid synthesis, does not occur until relatively late in gestation. Although ACTH is considered essential to fetal adrenal growth and function, the role that ACTH has in the development of the transitional and definitive zones, is less clear. To answer this question, the width of these zones was determined by immunocytochemical expression of P 450c17 and/or 3betaHSD in fetal adrenal glands obtained on day 100 (mid) of gestation (term = day 184) from baboons in which ACTH was administered to the fetus on days 95-99 of gestation or on day 165 (late) of gestation from baboons in which fetal ACTH was suppressed by treatment of the mother and fetus with betamethasone on days 150-164 of gestation. At midgestation, the fetal adrenal was comprised almost exclusively of fetal zone cells and a small definitive zone (38 +/- 2 microm in width), but was essentially devoid of a transitional zone (7 +/- 2 microm). Treatment with ACTH enhanced (P < 0.05) the width of the transitional zone (67 +/- 4 microm), but not the size of the definitive zone (10 +/- 4 microm). In late gestation, the width of the definitive zone, although 2 fold greater than that on day 100, was smaller (P < 0.05) than that of the transitional zone (120 +/- 15 microm), which greatly exceeded that at midgestation. Treatment with betamethasone in late gestation eliminated the transitional zone, but had no effect on the size of the definitive zone (120 +/- 8 microm). These findings indicate that the development of the baboon fetal adrenal transitional zone late in gestation is dependent on fetal pituitary ACTH. In contrast, the ontogenesis of the definitive zone at midgestation and its growth in late gestation occur in the relative absence of ACTH. PMID- 10523039 TI - Expression of anti-Mullerian hormone during normal and pathological gonadal development: association with differentiation of Sertoli and granulosa cells. AB - The ontogeny of expression of anti-Mullerian hormone (AMH) was examined by immunohistochemistry in 135 human gonadal tissue specimens of various developmental age, ranging from 6 weeks of fetal development to 38 yr of postnatal age. The series included specimens from normal testes and ovaries and from individuals either with pathological conditions affecting gonadal development or with idiopathic infertility manifested as azoospermia or severe oligozoospermia. AMH expression was found only in Sertoli and granulosa cells. A 6-week-old fetal testis at the indifferent gonad stage did not yet express AMH. The protein was first visible at 8.5 weeks of development, when sex cords have not yet been formed. Afterward, a majority of testicular specimens, including those from pathological conditions, strongly expressed AMH through fetal development and childhood until puberty. Markedly prolonged expression of AMH was observed in a 20-yr-old 46,XY female with androgen insensitivity syndrome, who retained prepubertal testicular morphology. In normal testes, the switch-off of AMH expression was usually associated with the appearance of primary spermatocytes, suggesting that their presence had an inhibitory effect on AMH. However, in adolescent boys lacking germ cells because of cancer treatment and in a majority of infertile adult men with idiopathic germ cell aplasia, AMH expression was also down-regulated despite the complete lack of spermatogenesis. The decrease in AMH expression thus reflects the terminal differentiation of Sertoli cells and is probably only partially dependent upon a regulatory factor associated with the onset of meiosis. In fetal ovaries, AMH was first detected at 36 weeks gestation in granulosa cells of preantral follicles. Thus, the onset of ovarian expression is at the end of fetal life and not in infancy as previously reported. PMID- 10523041 TI - Comment on analysis of mutations in genes of the follicle-stimulating hormone receptor in ovarian granulosa cell tumors. PMID- 10523040 TI - The vascular endothelial growth factor/fms-like tyrosine kinase system in human ovary during the menstrual cycle and early pregnancy. AB - In human ovaries, angiogenesis is known to be associated with the development of follicles and the formation of the corpus luteum (CL). A complex vascular network is formed within the thecal cell layer during follicular growth, and rapid neovascularization occurs toward the granulosa cell layer after ovulation. Vascular endothelial growth factor (VEGF) is a multifunctional cytokine, stimulating endothelial cell growth and enhancing microvascular permeability. A specific receptor for VEGF, fms-like tyrosine kinase (Flt-1), is expressed in vascular endothelial cells that mediates the action of VEGF. We examined the localization and expression of VEGF and Flt-1, using an immunohistochemical technique and RT-PCR analysis, in human follicles and corpora lutea during the normal menstrual cycle and early pregnancy. We measured concentrations of VEGF in extracts of human CL using an enzyme-linked immunosorbent assay during the luteal phase and early pregnancy. Immunostaining for VEGF was observed in granulosa cells from small antral follicles to preovulatory follicles. The staining was detected in thecal cells from medium-sized to preovulatory follicles. The intensity of the staining was gradually increased as a follicle grew. Flt-1 was localized in granulosa and thecal cells of preovulatory follicles as well as in endothelial cells. In the human CL, the intense staining for VEGF was observed in granulosa and thecal lutein cells, especially in the midluteal phase. The immunostaining for Flt-1 was faint in endothelial cells in the CL, whereas it was distinct in granulosa and thecal lutein cells. The concentrations of VEGF in lutein extracts were high in the early and midluteal phases and tended to decrease toward the late luteal phase. During early pregnancy, a measurable amount of VEGF was detected. RT-PCR analysis demonstrated that messenger ribonucleic acids encoding VEGF121, VEGF165, and Flt-1 were expressed in the CL. These results suggest that VEGF might have an autocrine role in the ovulatory process and luteal function as well as a paracrine role in angiogenesis. PMID- 10523042 TI - Individualized testing and outcome measurements are needed for successful hormone replacement therapy. PMID- 10523043 TI - Evidence for striatal modulation in the presence of fixed cortical injury in obsessive-compulsive disorder (OCD). AB - A patient with obsessive-compulsive disorder (OCD) onset resulting from a traumatic head injury underwent longitudinal brain imaging evaluation. Structural and functional brain imaging studies were repeatedly performed before and after treatment. Computerized tomography (CT) demonstrated bilateral prefrontal contusions immediately following the trauma and prior to the onset of OCD. Magnetic resonance imaging (MRI) demonstrated bilateral cortical abnormalities in the prefrontal and anterior-temporal regions a few months following the onset of OCD. Almost concurrently, single photon emission computerised tomography (SPECT) demonstrated bilateral perfusion deficits in fronto-temporal regions, and asymmetric increased perfusion in the anterior striatum. Six months later, after clinical improvement, a second SPECT study demonstrated improvement of brain perfusion, mostly in the striatum. The reflection of these results on a possible model of brain pathogenesis in OCD, and the role of brain imaging in neuropsychiatric evaluation, are demonstrated. PMID- 10523044 TI - Effects of abused drugs on thresholds and breaking points of intracranial self stimulation in rats. AB - The present study aimed to compare the effects of various abused drugs on threshold current intensities and the breaking points of intracranial self stimulation. Effects of morphine (1-10 mg/kg, s.c.), d-amphetamine (0.3-3.2 mg/kg, i.p.), nicotine (0.1-3.2 mg/kg, s.c.), ethanol (0.6-2.4 g/kg, p.o.), caffeine (1-20 mg/kg, i.p.) and phencyclidine (0.3-5.6 mg/kg, i.p.) were studied in male Wistar rats trained to lever-press for electrical stimulation of ventral tegmental area. Morphine and d-amphetamine were the only two drugs that both decreased threshold currents and increased the maximal ratio of reinforced and non-reinforced responses. Nicotine (1 mg/kg) and ethanol (1.2 g/kg) lowered threshold currents while both decreases and increases in threshold current were seen after administration of low (5 mg/kg) and high (20 mg/kg) doses of caffeine, respectively. Nicotine, ethanol and caffeine had no effects in the progressive ratio procedure. Effects of phencyclidine did not reach levels of statistical significance in either procedure although high doses of phencyclidine disrupted performance in the progressive ratio procedure. PMID- 10523045 TI - Interactions of lithium and drugs that affect signal transduction on behaviour in rats. AB - The therapeutic mechanism of the action of lithium in the treatment of bipolar affective disorder is not known, in spite of a burgeoning number of biochemical studies linking lithium to signal transduction processes. This article reviews a decade of studies examining the behavioural manifestations of manipulating inositol, cyclic adenosine monophosphate (cAMP) and G proteins in rats. Inositol, forskolin, dibutyryl cAMP and pertussis toxin all interacted with lithium when rearing behavior was measured. Lithium potentiated the increase in locomotion induced by injections of cholera toxin into the nucleus accumbens, consistent with the hypothesis that it inactivates inhibitory G proteins. More specific interactions were found between lithium and inositol following cholinergic and serotonergic stimulation. Inositol, but not forskolin, attenuated lithium pilocarpine seizures and the enhancement of the serotonin syndrome; however, inositol had no effect on lithium-induced attenuation of wet dog shakes following an injection of 5-hydroxytryptophan. Behavioural evidence supports biochemical findings suggesting that lithium's interactions with the phoshphatidyl inositol and cyclic AMP signal transduction systems may be relevant to its therapeutic effects in bipolar disorder. Further research on more specific behaviours may elucidate the relevant pharmacological mechanisms underlying the therapeutic effect of lithium. PMID- 10523046 TI - Reduced cerebrospinal HVA concentrations and HVA/5-HIAA ratios in suicide attempters. Monoamine metabolites in 120 suicide attempters and 47 controls. AB - Dysfunctions of central monoaminergic systems are important elements of the leading biological hypotheses of suicide and depression. The purpose of the present paper was to study the levels and the relationships between the serotonin metabolite 5-hydroxyindoleacetic acid (5-HIAA), the dopamine metabolite homovanillic acid (HVA) and the norepinephrine metabolite 3-methoxy-4 hydroxyphenylglycol (MHPG) in the cerebrospinal fluid (CSF) in 120 hospitalised suicide attempters and 47 controls (healthy volunteers or patients admitted for minor surgery). The suicide attempters showed significantly lower HVA levels (174+/-82 vs. 216+/-96 nmol/L, P=0.004), HVA/5HIAA ratios (1.6+/-0.5 vs. 2.1+/ 0.6, P=0.0001) and HVA/MHPG ratios (4.2+/-2.1 vs. 4.8+/-1.7, P=0.02) than the controls. The correlations between the monoamine metabolites were markedly lower in patients than in controls. CSF 5-HIAA showed no significant differences between patients and controls (107+/-40 vs. 108+/-51 nmol/L) or between violent and non-violent attempters (112+/-58 vs. 105+/-33 nmol/L). The monoamine metabolites showed no significant differences between survivors and patients who subsequently completed suicide, or between suicide attempters subgrouped by psychiatric diagnoses. The results suggest that low HVA levels and altered relationships between the monoamine metabolites are associated with suicidal behaviour. PMID- 10523047 TI - Orbitofrontal cortex dysfunction in obsessive-compulsive disorder? I. Alternation learning in obsessive-compulsive disorder: male-female comparisons. AB - BACKGROUND: We have previously reported a significant negative correlation between severity of symptoms and performance of an alternation learning task in female obsessive-compulsive disorder (OCD) patients. The present study was aimed at exploring this relationship between alternation learning and OCD symptom severity in male OCD patients. METHODS AND RESULTS: Eighteen female obsessive compulsive disorder patients and 14 male non-depressed, drug free, OCD patients participated in the study. Measures of dorsolateral prefrontal function (Wisconsin Card Sorting Test) and orbitofrontal cortex function (object alternation learning) showed no significant differences between the sexes. The relationship between orbitofrontal cortex function and severity of OC symptoms was significantly different between the sexes (z=2.44. P=0.007). While this correlation was negative in the females it was positive in the males. CONCLUSIONS: These results may indicate sexual dimorphism in OCD. PMID- 10523048 TI - Orbitofrontal cortex dysfunction in obsessive-compulsive disorder? II. Olfactory quality discrimination in obsessive-compulsive disorder. AB - BACKGROUND: Olfactory quality discrimination is a putative marker of orbitofrontal cortex function in mammals. As this portion of the cerebral cortex was repeatedly implicated in the pathophysiology of obsessive-compulsive disorder (OCD) this study was designed in an attempt to quantify this behavioural function in OCD patients. METHODS AND RESULTS: Olfactory quality discrimination was compared in OCD patients and healthy controls. Thirty two subjects participated in the study: 16 (13 women and 3 men) medication free OCD outpatients and 16 sex and age matched healthy controls. Olfactory tests consisted of determination of detection thresholds to isoamyl acetate, and a three way forced choice quality discrimination task, using isoamyl acetate, citral and eugenol as stimuli. No significant differences in sensitivity and performance of the quality discrimination task between the two groups were found. Within the OCD group the more severely affected patients (Y-BOCS>29) performed significantly better than the less severely affected (Y-BOCS<30) patients on the more difficult part of the quality discrimination task. Within this subgroup of patients the correlation between performance on the olfactory task and a previously reported alternation task tended to be negative as compared to a significantly positive correlation in the control group. CONCLUSIONS: It seems that olfactory quality discrimination may prove to be a useful noninvasive marker of prefrontal cortex function in OCD. Furthermore, the organization of functional modules within the orbitofrontal cortex, rather than a simple dysfunction, may prove to characterize OCD. PMID- 10523049 TI - Some behavioural effects of risperidone in rats: comparison with haloperidol. AB - Risperidone is a dopaminergic as well as a 5-HT2 antagonist. The drug was found to exert beneficial effects on both positive and negative symptoms of schizophrenia. Since recently, schizophrenia is regarded as a composite of not only positive and negative but also affective and cognitive symptoms, in this study the effects of risperidone compared with typical neuroleptic haloperidol, on affective and cognitive functions were investigated in rats (anxiolytic, antidepressive and memory tests). We found, that in contrast to haloperidol, risperidone had antidepressive, anxiolytic and memory enhancing effects. The results obtained correspond with favourable effects of risperidone on mood disturbances and cognitive functions of schizophrenic patients observed under clinical conditions. PMID- 10523051 TI - Comments on donepezil open label trial. PMID- 10523050 TI - Effect of sustained administration of the 5-HT1A receptor agonist flesinoxan on rat 5-HT neurotransmission. AB - A short-term treatment with flesinoxan (2.5 and 5 mg/kg/day x 2 days, s.c., delivered using osmotic minipumps) decreased significantly the spontaneous firing activity of dorsal raphe serotonin (5-HT) neurons of male Sprague-Dawley rats. This firing was still decreased following 1 week of treatment with flesinoxan (5 mg/kg/day) but was back to normal after a treatment of 2 weeks. This recovery of firing was associated with a 3-fold shift to the right of the dose-response curve of the effect of the 5-HT autoreceptor agonist lysergic acid diethylamide on the firing activity of 5-HT neurons, indicating a desensitization of somatodendritic 5-HT1A autoreceptors. At the postsynaptic level, long-term treatment with flesinoxan (5 mg/kg/day x 14 days) did not modify the responsiveness of dorsal hippocampus CA3 pyramidal neurons to microiontophoretic applications of 5-HT and flesinoxan nor to endogenous 5-HT released by the electrical stimulation of the ascending 5-HT pathway, indicating an unchanged sensitivity of postsynaptic 5 HT1A receptors. Finally, in rats treated with flesinoxan for 2 weeks, the administration of the selective 5-HT1A receptor antagonist (N-{2-[4(2 methoxyphenyl)-1-piperazinyl]ethyl}-N-(2-pyridinyl)cyclohe xanecarboxamide trihydroxychloride (WAY 100635, 100 and 500 microg/kg, i.v.) did not increase the firing activity of dorsal hippocampus CA3 pyramidal neurons, thus failing to reveal an enhanced tonic activation of postsynaptic 5-HT1A receptors as for other antidepressant drugs, including the 5-HT1A receptor agonist gepirone. The marked potency and the long dissociation constant of flesinoxan for the 5-HT1A receptors may account for the latter discrepancy. In conclusion, as for selective 5-HT re uptake inhibitors, monoamine oxidase inhibitors and 5-HT1A receptor agonists, flesinoxan produced most of the adaptive changes exerted by these antidepressant drugs on the 5-HT system. PMID- 10523052 TI - Clinical investigation of medicinal products in the treatment of Parkinson's disease (CPMP note for guidance). European Agency for Evaluation of Medicinal Products. AB - These notes are intended to provide guidance for the evaluation of drugs in the treatment of Parkinson's disease. They should be read in conjunction with the Directive 75/318/EEC and 83-570/EEC and current and future EC and ICH guidelines, especially those on: Studies in support of special populations: geriatrics (ICH E7). The extent of population exposure to assess clinical safety for drugs intended for long-term treatment in non life threatening conditions (ICH E1). General considerations for clinical trials (ICH-E8). Statistical principles for clinical trials (ICH-E9). Pharmacokinetic studies in man Clinical testing of prolonged action forms, with special reference to extended release forms. Dose response information to support product authorisation PMID- 10523053 TI - Vitamins and brain development. AB - Effects of deficiency of vitamins on early development of brain have been reviewed. Unusual developmental problems in neurogenesis specific for the brain and impairment of its functional capacities due to vitamin deficiency have been discussed. The species-specific "critical periods" in development of various systems have been mentioned. Indices such as reflex activity, locomotion, special senses, cognition and adaptive behavior were used for assessing brain maturation in experimental models and humans. Significant examples include brain anomalies in humans and other mammals caused by retinoid excess or deficit; increase in calbindin D28K, a vitamin D dependent calcium-binding protein during postnatal period in rat; hydrocephalus and exencephaly in prenatal rats and subarachnoidal or intracerebral hemorrhage in infants caused by vitamin E deficiency. Peripheral neuropathic lesions leading to infantile beriberi is caused by thiamine deficiency. Impaired growth in retinal layers leading to delay in maturation of electroretinogram and depth-perception in postnatal rats occur due to pyridoxine deficiency. Infants of severely vitamin B12 deficient mothers show abnormalities in behavior involving basal ganglia and pyramidal tract. Folic acid deficiency results in delayed maturation of the basic electroencepalographic patterns. In addition, vitamin-interactions leading to developmental errors have been pointed out. Vitamin B6 deficiency impairs vitamin B12 absorption and biotin deficiency may be aggravated by pantothenic acid deficiency. Vitamin C deficiency resulting in impaired metabolism may produce symptoms of deficiency of folic acid. Another characteristic examples is that iron absorption from dietary sources is dependent on ascorbic acid. PMID- 10523054 TI - Pre- and postprandial expression of the leptin receptor splice variants OB-Ra and OB-Rb in murine peripheral tissues. AB - Leptin receptors (OB-R) are widely distributed in peripheral tissues. However, the RT-PCR data published on the distribution of OB-R are not always consistent. The present study was undertaken in order to test whether the different muscle fiber type profile or the acute nutritional status in which tissue samples were excised from animals may influence OB-R expression. Six 12-week-old male Swiss Webster mice were killed by decapitation either 1 h after feeding or after a 16-h fast, and the kidneys, testes, brown adipose tissue, gastrocnemius (G), soleus (SOL) and extensor digitorum longus (EDL) muscles were dissected out. In parallel, muscle fibers obtained from other animals were classified on the basis of differences in the staining intensity for myofibrillar adenosinetriphosphatase. The expression of OB-R isoforms was assessed by RT-PCR and ethidium bromide staining. The signal for OB-Ra and OB-Rb was detected in all tissues examined. No differences were observed in samples obtained from either fed or fasted mice. G, SOL and EDL muscles showed the same pattern of OB-R expression. Neither the short-term nutritional changes of the animal as regards to the pre- versus the postprandial-state nor differences in muscle fiber type had any influence on the qualitative expression of the OB-R splice variants a and b in the murine tissues studied. However, quantitative differences cannot be ruled out. PMID- 10523055 TI - Serum leptin levels in patients with anorexia nervosa before and after partial refeeding, relationships to serum lipids and biochemical nutritional parameters. AB - Leptin is a protein hormone produced by adipocytes that provide information about the body fat content. It was previously reported that serum leptin levels were decreased in patients with anorexia nervosa in comparison with healthy control subjects. The aim of our study was to compare serum leptin levels in patients with anorexia nervosa (n=11, initial mean BMI=15.4 kg/m2) before and after partial recovery with control age-matched subjects (n=11, mean BMI= 20.3 kg/m2) and to study the relationships of leptin levels, serum lipids and biochemical nutritional parameters. We found that serum leptin concentrations in patients with anorexia nervosa were significantly reduced in comparison with control subjects (3.61 vs 9.37 ng.ml(-1), p<0.01). Serum cholesterol, triglycerides, total protein and albumin in patients with anorexia nervosa either before or after partial recovery did not differ from the control group. After partial recovery, a significant increase in serum leptin was observed (4.83 vs 3.61 ng.ml(-1), p<0.05), but the values still remained significantly lower than in the control group (p<0.01) Leptin levels correlated positively with the body mass index in the control group and anorexia nervosa group before recovery. The correlation with BMI in the anorexia nervosa group after refeeding was not significant. No significant correlation was found between leptin concentrations and serum lipids, total protein, albumin and prealbumin, respectively. Serum leptin thus represents a sensitive parameter that reflects the nutritional status in patients with anorexia nervosa suitable for long-term follow up during refeeding therapy. PMID- 10523056 TI - Oxidative stress in normal and diabetic rats. AB - Parameters related to oxidative stress were studied in a group of 10 Wistar diabetic rats and 10 control rats. The levels of total erythrocyte catalase activity in the diabetic animals were significantly (p<0.001) greater than the control levels. The diabetic animals presented an amount of vitamin E far greater (p<0.0001) than the controls, as was also the case for the vitaminE/polyunsaturated fatty acid (PUFA) and vitaminE/linoleic acid (C18:2) ratios. Greater vitaminE/triglyceride (TG) ratio, however, appeared in the control group. The corresponding vitamin A ratios (vitaminA/TG, vitaminA/PUFA, vitaminA/C 18:2) were higher in the control group. Our work corroborates the findings that fatty acid metabolism presents alterations in the diabetes syndrome and that the antioxidant status is affected. PMID- 10523057 TI - Estrogen and gender do not affect fatigue resistance of extensor digitorum longus muscle in rats. AB - The effects of estrogen on skeletal muscle fatigue are controversial. To determine the effects of estrogen and gender on rat extensor digitorum longus (EDL) muscle, we either injected 40 microg beta-estradiol 3/benzoate.kg BW(-1) to female rats or sham injected male or female rats for 14 days. Subsequently a 90 min fatigue protocol consisting of electrical stimulation at 10 Hz delivered in 500 ms trains was administered. Force was recorded for a 5 s period at the start of the protocol (0 min) and at 5 min intervals until completion following 90 min of stimulation. After 90 min, EDL force generation at 10 Hz stimulation declined in all groups to between 50-60 % of initial values. However, no significant difference in fatigue rate or final 10 Hz stimulated force was seen between females administered estrogen, sham injected females or males. Hence, estrogen administration and gender did not significantly affect EDL muscle fatigue in this model. PMID- 10523058 TI - Anoxia/induced membrane changes in human red blood cells. AB - The cation-osmotic hemolysis was studied in human red blood cells incubated under anoxic conditions. In relation to the time course of anoxia, two phases of hemolysis were distinguished. A significant decrease of hemolysis was found between 3 and 24 h of incubation. On the other hand, hemolysis was significantly increased after prolonged incubation (48-72 h). Using the method of cation osmotic hemolysis, the properties of two membrane constituents, spectrine membrane skeleton and membrane bilayer, were studied. The relation between cation osmotic hemolysis and erythrocyte deformability is being discussed. PMID- 10523059 TI - Bioassay of cadmium and its effect on differential distribution of dehydrogenases in different brain regions in Labeo rohita (HAM). AB - The sublethal effect of cadmium on the specific activities of lactic, malic and succinic dehydrogenases in different brain regions in Labeo rohita (HAM) was assessed with reference to acute, chronic and recovery conditions. Cadmium enhanced succinic, malic and lactic dehydrogenases to a marked extent in the cerebrum from 0 to 12 h exposure. However, a subsequent fall of the above enzymes in some regions was recorded from 12 to 24 h. In chronic studies, the greatest decrease in succinic dehydrogenase was noted in the cerebrum (0 to 15 days) and the least reduction in the cerebellum (30 to 45 days) in comparison with malic and lactic dehydrogenase. In recovery studies an optimum rise in lactic, malic and succinic dehydrogenase was found in the cerebrum (30-45 days). In general, cadmium accumulation was highest in the cerebrum (12 h and 15 days) and least in the cerebellum (24 h and 45 days). This was markedly above the safety level in acute and chronic situations. PMID- 10523060 TI - Two-phase response of rat pineal melatonin to lethal whole-body irradiation with gamma rays. AB - Male Wistar rats adapted to artificial light:dark (LD) regimen 12:12 h were whole body irradiated with a single dose of 9.6 Gy of gamma rays and sham/irradiated in the night in darkness. The rats were examined 60 min, 1, 3 and 5 days after exposure between 22:00 and 01:30 h in the darkness. The results obtained indicate a two-phase reaction of pineal melatonin after the lethal irradiation of rats: the decline of melatonin concentration early after the exposure (at 60 min) with unchanged serotonin N-acetyltransferase (NAT) activity followed by an increase of melatonin synthesis, accompanied by an increase of pineal and serum melatonin on day 5 after the exposure. NAT activity was increased on day 3 after the exposure. Serum corticosterone concentrations in irradiated rats were increased 60 min and 3 days after exposure. With respect to the antioxidant, immunomodulating and stress-diminishing properties of melatonin, we consider the increase in melatonin synthesis during later periods after irradiation as part of adaptation of the organism to overcome radiation stress. PMID- 10523061 TI - Daily rhythm in rat pineal catecholamines. AB - A daily rhythm in the oscillations of pineal dopamine, norepinephrine and epinephrine content was found in male Wistar: Han rats. The acrophases of the oscillations were localized in the first half of the dark period and generally higher values were found in the dark part of the day. PMID- 10523062 TI - Mechanisms of action of selective serotonin reuptake inhibitors in the treatment of psychiatric disorders. AB - Selective serotonin reuptake inhibitors (SSRIs) have demonstrated efficacy in depression and anxiety disorders. This raises the question of how the single action of serotonin reuptake inhibition can improve several psychiatric conditions. In order to understand this apparent paradox it is necessary to consider where SSRIs act in the pathogenic process underlying depression or anxiety disorders. Tryptophan depletion has been used extensively in research into depression and has shown that, in patients receiving an SSRI whose depression is in remission, depleting serotonin leads to recurrence of the disorder. Similar results have been found for panic disorder. This suggests that increased levels of serotonin are necessary in the synapse for the SSRI to be effective in the treatment of depression and panic disorder. In obsessive compulsive disorder, depletion of serotonin in patients recovered on an SSRI does not cause relapse; receptor adaptation may be more important. Variations within the SSRI drug class, such as the selectivity ratios for serotonin versus noradrenaline uptake, elimination half-life, and affinity for the 5-HT2 receptor have been identified and may be important determinants of efficacy, side effects and clinical use. PMID- 10523063 TI - Anxiety with depression: a treatment need. AB - In the community, depression and anxiety are highly prevalent. The relationship between these two conditions is often unclear and has been the subject of much discussion. The classical viewpoint considered depression and anxiety to be two disorders with separate treatment options: anxiolytic agents for anxiety and antidepressant agents for depression. Depression and anxiety may occur together representing comorbid 'pure' conditions, or anxiety may predispose depression (or vice versa), or symptoms of anxiety and depression may be external manifestations of one underlying cause, or there may be an overlap of classifications. It is also possible that a mixed anxiety and depressive disorder exists: the association of subthreshold depressive symptoms and subthreshold depressive anxiety. This review examines the relationship of co-occurring depression and anxiety. The socioeconomic burden and the need for effective treatment of comorbid anxiety and depression are highlighted. The SSRIs, e.g. paroxetine, are discussed in particular as appropriate pharmacological therapy for anxiety with depression. PMID- 10523064 TI - Facing the challenge of social anxiety disorder. AB - Social anxiety disorder is at serious and prevalent disorder that leads to significant disability in the social and professional lives of patients. It is a chronic condition that frequently coexists with other psychiatric conditions such as depression or alcoholism. Pharmacotherapy with benzodiazepines, monoamine oxidase inhibitors and selective serotonin reuptake inhibitors (SSRIs) has been assessed. Alprazolam has modest efficacy, while a more robust response to clonazepam has been reported. Benzodiazepines, however, are not suitable for long term treatment of a chronic condition such as social anxiety disorder and are ineffective against comorbid depression. Phenelzine has demonstrated efficacy but the need for dietary restrictions has limited its use. Conflicting results have been reported in placebo-controlled trials for moclobemide, with two studies showing moclobemide to be more effective than placebo while recent trials have reported less robust results. Based on clinical evidence, SSRIs are the first line treatment in social anxiety disorder. The most extensive database for the treatment of social anxiety disorder exists for the SSRI, paroxetine. Three large multicentre, placebo-controlled, trials have been completed. In all three studies, a significantly greater proportion of patients responded to paroxetine treatment compared with placebo. Paroxetine is currently the only SSRI licensed for use in this condition. PMID- 10523065 TI - Where are we going with SSRIs? AB - During the past decade the selective serotonin reuptake inhibitors (SSRIs) have become established as the treatment of choice for depression. As newer antidepressants become available on the market, it is important to reappraise the position of the SSRIs in the management of depression. This review will address the question: where are we going with the SSRIs? The continued establishment of the SSRIs as first-line treatment for depression will be discussed, focusing on the more rapid onset of antidepressant efficacy seen with pindolol augmentation and the use of SSRIs for treatment of depression in patients with physical illnesses, particularly ischaemic heart disease. The SSRIs have well-documented efficacy in panic disorder and obsessive-compulsive disorder, and paroxetine has recently been licensed for social anxiety disorder/social phobia in some countries. Results will be presented from studies with the SSRIs in new therapeutic areas, including post-traumatic stress disorder and generalized anxiety disorder. PMID- 10523066 TI - Hypoxic pulmonary vasoconstriction. AB - Hypoxic vasoconstriction is unique to pulmonary circulation. The pulmonary response is part of a self-regulatory mechanism by which pulmonary capillary blood flow is automatically adjusted to alveolar ventilation for maintaining the optimal balance of ventilation and perfusion. In pathological conditions, hypoxic pulmonary vasoconstriction may occur as an acute episode or as a sustained response with pulmonary hypertension and vascular remodeling. Vasoactive substances produced from the endothelial cells (prostanoids, nitric oxide, or endothelin) or other mediators such as 5 hydroxytryptamine have been examined as possible mediators of hypoxic vasoconstriction. These appear more likely to be modulators than mediators of the vasoconstrictor response to hypoxia. Recent hypotheses have emerged indicating that O2 levels per se can regulate ion channel activity. The modulation of both K+ and Ca2+ channels differs according to the conduit or resistance pulmonary vessel type, tending to extend the former and contract the latter, thereby opposing the ventilation to perfusion mismatching. In the absence of drugs that act selectively on pulmonary circulation, inhaled therapy is an alternative in the treatment of pulmonary hypertension. According to its short half-life and to its potential cytotoxicity, nitric oxide is only of value in the management of patients with acute respiratory disease. Aerosolized prostacyclin and iloprost result in a sustained efficacy of the inhaled vasodilator regimen in patients with severe pulmonary hypertension and offer a new strategy for treatment of this disease. At the moment, therapy aimed at reversing the structural remodeling and matrix deposition in pulmonary arteries remains experimental. New drugs such as potassium channel openers or endothelin receptor antagonists warrant further investigations as possible therapeutic candidates in the treatment of pulmonary hypertension. PMID- 10523067 TI - The effects of chronic L-arginine treatment on vascular responsiveness of streptozotocin-diabetic rats. AB - In this study, the protective effects of L-arginine treatment in vivo on vascular reactivity of streptozotocin (STZ)-induced 12-week-old diabetic rats were examined. Loss of weight, polydipsia, polyphagia, hyperglycemia, hypoinsulinemia, and elevated levels of plasma cholesterol and triglyceride were observed in diabetic rats. L-arginine treatment (1 mg/mL in drinking water) did not significantly affect these metabolic and biochemical abnormalities. Plasma malondialdehyde (MDA) levels in untreated diabetic rats were also significantly higher than untreated controls. However, L-arginine treatment prevented the increase in MDA level of plasma of diabetic rats. Contractile responses, but not sensitivity to noradrenaline (NA), were significantly increased in diabetic rats compared to controls. Treatment of diabetic rats with L-arginine completely prevented the increase in NA responses. Relaxation response to acetylcholine (ACh), but not to sodium nitroprusside (SNP), in diabetic aorta has been found to be significantly decreased as compared with controls. However, there were no significant differences in pD2 values of acetylcholine in either of the groups. L arginine treatment increased the ACh responses to the control level. All effects of L-arginine on vascular reactivity were found to be specific for diabetic rats and not controls. These results suggest that functional abnormalities occurred in aorta from diabetic rat might at least in part result from L-arginine deficiency, and the lipid peroxidation-lowering effect of L-arginine may account for its protective effect on vascular reactivity of diabetic rats. PMID- 10523068 TI - The contribution of nitric oxide and endothelins to angiotensin: II. Evoked responses in the rat isolated uterus smooth muscle. AB - The present study was designed to determine the role of endogenous endothelin peptides and nitric oxide on angiotensin II (All) responses in the isolated nonpregnant rat uterine smooth muscle. AII (10, 20, or 50 ng/ml) increases rhythmic oscillations dose dependently (32.7 +/- 8.9, 55.96 +/- 10.3, and 62.78 +/- 17.7% increase, respectively). L-arginine methyl ester (L-NAME; 10(-5) M) did not affect the increase in rhythmic oscillations induced by All (10, 20, or 50 ng/ml) (17.5 +/- 12.1, 31.5 +/- 18.3, and 52.5 +/-11.8% increase, respectively, n = 6, p > 0.05). It reduced the contractile responses to AII (10 ng/ml: from 4.63 +/- 0.6 to 1.8 +/-0.7 cm2, p < 0.05: and 20 ng/ml: from 5.59 +/- 0.8 to 2.11 +/- 0.4 cm2, p < 0.05, n = 6). L-arginine (10 mM) decreased the contractile response obtained by AII (10 or 20 ng/ml) (1.93 +/- 1.05, p < 0.05 and 2.14 +/- 0.7 cm2, p < 0.05, respectively, n = 6). BQ 485 (50 ng/ml) decreased both the number of rhythmic oscillations and the contractility increased by AII. Bosentan (10(-5) M) induced an increase in the number of rhythmic oscillations but decreased the contractile responses to the higher concentrations of All. These data show that endogenous NO and endothelin peptides contribute to the motility changes induced by AII and may play an important role in the pathophysiological events of the uterine function. PMID- 10523069 TI - Cardiac effects of non-depolarizing neuromuscular blocking agents pancuronium,vecuronium, and rocuronium in isolated rat atria. AB - Pancuronium, vecuronium, and rocuronium produce different cardiac effects. Using spontaneously beating right and electrically stimulated left rat atria, while measuring developed force, effective refractory period, and heart rate, we determined and compared the concentration-dependent cardiac effects of the compounds. The preparations were exposed to five progressively increasing concentrations of these compounds (10(-9), 10(-8), 10(-7), 10(-6), and 10(-5) mol/L). Pancuronium increased heart rate; vecuronium and rocuronium produced positive inotropic effects; and vecuronium shortened refractoriness. These effects may be the result of a blockade of the M2 muscarinic receptors. However, the concentrations required to produce changes were higher than those observed in patients under neuromuscular blockade. PMID- 10523070 TI - Structure-activity relationships of new thienothiazine derivatives in isolated heart and smooth muscle preparations of guinea pigs. AB - New thienothiazine derivatives that differ in their side chains on the nitrogen atom of the thienothiazine ring were investigated regarding structure-activity relationships and calcium antagonistic and/or potassium channel opening properties. Isometric contraction force was measured in guinea pig papillary muscles, aortic strips, and terminal ilea. Chronotropic activity was studied in right atria of guinea pigs. The derivatives with a dimethylaminoethylcarboxamide side chain (HO4) and with a dimethoxyphenylethyl-N-methylaminoethylcarboxamide side chain (HO7) had the most potent negative inotropic effects on papillary muscles and spontaneously beating right atria. The negative inotropic and chronotropic effects of the compounds with a methylpiperazinylcarbonyl side chain (HO5) or a diethylaminopropylcarboxamide side chain (HO6) were less pronounced. The negative inotropic action was reversed by increasing the extracellular calcium concentration. It was also reversed by glibenclamide, for concentrations of the compounds up to the EC50, but not at higher compound concentrations. Among all the compounds studied, HO 7 had the strongest relaxing effect on aortic strips and terminal ilea. The effects of the derivatives on the smooth muscles could not be reversed by glibenclamide. The calcium antagonistic effect of the thienothiazine derivatives is more pronounced than the potassium channel opening activity, at least in high drug concentrations. Compounds with an aromatic or heterocyclic ring in the side chain have the weakest negative inotropic and negative chronotropic effects on papillary muscles and right atria. However, HO7 showed a tissue specificity with the most potent relaxing effect on aortic strips and terminal ilea. PMID- 10523071 TI - Hydrogen peroxide-induced endothelium-dependent relaxation of rat aorta involvement of Ca2+ and other cellular metabolites. AB - In phenylephrine-precontracted rings, H2O2 produced an endothelium-dependent relaxation at concentrations of 4.4 x 10(-7) to approximately 4.4 x 10(-5) M. Removal of extracellular Ca2+ ([Ca2+]0) markedly attenuated the relaxant effects of H2O2. Complete inhibition of the H2O2 relaxant action was obtained after buffering intracellular Ca2+ ([Ca2+]i) in endothelial cells, with 10 microM acetyl methyl ester of bis (o-aminophenoxy) ethane-N,N,N',N'-tetraacetic acid (BAPTA-AM). These relaxant effects of H2O2 were nearly abolished by 15 x 10(-5)M N(G)-monomethyl-arginine (L-NMMA) or 5 x 10(-5) M N(G)-nitro-L-arginine (L-NAME) and were attenuated markedly by the presence of either 10(-6) M Fe2+, 10(-6) M Fe3+, or 5 x 10(-6) M methylene blue. These inhibitory effects of L-NMMA or L NAME could be reversed partly by 5 x 10(-5) M L-arginine. These Fe(2+)- and Fe(3+)-induced inhibitions of H2O2-stimulated relaxation were reduced significantly by either 1.0 mM deferoxamine (a Fe2+ chelator) or 100 microM dimethyl sulfoxide (DMSO). In addition, 17-octadecynoic acid (2.5 microM) or proadifen (10 microM) (both antagonists of cytochrome P450 metabolism of fatty acids) markedly decreased the H2O2 relaxant effects. Proadifen (10 microM) produced concentration-dependent impairment of vasorelaxation to acetylcholine. A variety of amine antagonists and a cyclo-oxygenase inhibitor all fail to interfere with or attenuate the H2O2-induced relaxations. Our observations suggest that, at suitable pathophysiologic concentrations, H2O2 could induce release of an endothelium-derived relaxing factor, probably nitric oxide, from endothelial cells. The H2O2 relaxant effects are clearly Ca(2+)-dependent and require formation of cyclic guanosine monophosphate (cGMP). These vasorelaxing effects of H2O2 appear to be induced by H2O2 itself. Hydrogen peroxide may stimulate production of some unknown metabolites metabolized by cytochrome P450 dependent enzymes. PMID- 10523072 TI - Fatty acid compositions in local sea cucumber, Stichopus chloronotus, for wound healing. AB - Fatty acid profile from crude extracts of local sea cucumber Stichopus chloronotus was determined using gas chromatography (GC) technique. The extracts were prepared separately in methanol, ethanol, phosphate buffer saline (PBS), and distilled water as part of our study to look at the affinity of these solvents in extracting the lipid from sea cucumber. The PBS and distilled water extractions indicate water-soluble components, while the organic fractions are extracted in methanol and ethanol as organic solvents. Furthermore, water extraction is the conventional method practiced in Malaysia. In our analysis the C14:0 (myristic), C16:0 (palmitic), C18:0 (stearic), C18:2 (linoleic), C20:0 (arachidic), and C20:5 (eicosapentaenoic, EPA) were significantly different (p < 0.01) in the four solvent extractions. However, the PBS extraction contained a much higher percentage of EPA (25.69%) compared to 18.89% in ethanol, 7.84% in distilled water, and only 5.83% in methanol, and variances were significantly different (p < 0.01 ). On the other hand, C22:6 (docosahexaenoic acid or DHA) is much higher in water extraction (57.55%), in comparison to the others where only 3.63% in PBS and 1.20% in methanol, and this difference is significant at p < 0.01. No DHA was detected in ethanol extractions. Subsequently, C18:1 (oleic acid) was only detected in PBS (21.98%) and water extraction (7.50%). It is interesting that palmitic acid, C16:() was higher in methanol (20.82%) and ethanol (2.18%), while 12.55% was detected in PBS and only 2.20% in water extraction: and again this was significantly different at p < 0.01. Although our results have shown that all four solvents were different in terms of their ability to extract fatty acids, the major component for tissue repair was well preserved. Probably this is one of the important precocious steps when working with a delicate sea cucumber, in both experimental and/or at the preparative stages. Freshness of the sea cucumber samples is important when undertaking this type of experiment. Finally, we believe that the local sea cucumber S. chloronotus contains all the fatty acids required to play a potential active role in tissue repair. PMID- 10523073 TI - The interaction of 6-mercaptopurine (6-MP) and methotrexate (MTX). AB - The antimetabolites 6-mercaptopurine (6-MP) and methotrexate (MTX) are the cornerstones in the maintenance treatment of children's acute lymphoblastic leukemia (ALL). The biochemical mechanisms underlying the increased therapeutic efficacy of the combination of these drugs have not yet been elucidated. However, both drugs interact with important pathways. such as purine de novo synthesis (PDNS), purine salvage, and methylation reactions. A review of the mechanistic aspects of the interactions between 6-MP and MTX is given. PMID- 10523074 TI - Age-related change of the role of alpha1L-adrenoceptor in canine urethral smooth muscle. AB - To examine age-related alteration of the role of alpha1L-adrenoceptor in the urethra, young non-parous and aged parous female dogs were used. In a functional study, we evaluated phenylephrine-induced contraction and antagonistic effects of JTH-601, a newly synthesized alpha1-adrenoceptor antagonist, and prazosin; in a localization survey using autoradiographic technique, we investigated specific [3H]JTH-601 and [3H]tamsulosin binding. Concentration-response curves were obtained for phenylephrine (pD2 = 5.0-5.3). JTH-601 and prazosin antagonized this contraction with pA2 values of 8.2-8.3 and 8.0-8.1, respectively. Specific binding of both [3H]JTH-601 and [3H]tamsulosin were observed in the bladder neck and proximal section of urethra. There were no significant differences of the pD2, pA2, and radio ligand binding between young non-parous and aged parous dogs. PMID- 10523075 TI - Role of hypoxia and constitutionally different resistance to hypoxia/stress as the determiners of individual profile of cytochrome P450 isozyme activity. AB - The hepatic cytochromes P450 1A1, 1A2, 2B1, and 2E1 activities have been investigated in the sublines of Wistar rats with principally different (high or low) resistance to hypoxia/stress. Repeated measurements in normoxic conditions showed a significant prevalence of total cytochrome P450 content, CYP 1A1, and CYP 2E1 activities in rats with low-resistance (LR) to hypoxia compared to rats with high-resistance to hypoxia (HR), whereas in HR rats the CYP 1A2 activity was 63% higher (p < 0.001) than in LR rats. In the conditions of acute hypobaric hypoxia these differences were manifested distinctly: in HR rats an enhancement of CYP 1A2 activity by 49% of aerobic value (p < 0.01) was observed, in LR rats the total P450 content, CYP 1A1 and 2E1 activities (p < 0.05-0.001) were increased. The 30-min total liver ischemia formed an individual response of the drug-metabolizing system: in HR rats CYP 1A2 and 2B1 activities were decreased in the early postischemic period and were not restored by the 21st day, whereas in LR rats CYP 1A2 activity was not affected and was induced more than 2-fold of aerobic value in the late post-ischemic period. The CYP 2B1 activity was induced almost 1.5-fold during the whole postischemic period. These data suggest that acute hypoxia and individual resistance to hypoxia/stress, one of the cardinal constitutional features, provide an individual reaction of drug-metabolizing system and enzymes of P450, in particular. The individual constitutional resistance to hypoxia/stress may be a serious criterion for an individual approach in pharmacotherapy of hypoxic states, diseases, as well as for prognosis and prevention of early and distant complications of irrational pharmacotherapy. PMID- 10523076 TI - Recent references. PMID- 10523077 TI - Negative contrast imaging of mitochondria by confocal microscopy. PMID- 10523078 TI - [Cancer reports and cancer incidence from regional cancer registries in Japan]. PMID- 10523079 TI - Deglycosylation of hen ovotransferrin under mild conditions: effect on the immunoreactivity and biological activity. PMID- 10523080 TI - Combining the two neonatal examinations. Midwives perform a neonatal examination, so was this counted? PMID- 10523081 TI - Proposed appraisal system and political correctness. Correspondent rather than the royal college may have lost touch with reality. PMID- 10523082 TI - Cyclosporin neurotoxicity after chemotherapy. Cyclosporin causes reversible posterior leukoencephalopathy syndrome. PMID- 10523083 TI - Funding long term care for older people. Redistribution of wealth from rich to poor is not function of a care service. PMID- 10523084 TI - Funding long term care for older people. Taxes ensure equal quality care for all at time it is needed. PMID- 10523085 TI - Commentary: Helicobacter pylori--the story so far. PMID- 10523086 TI - The end of the heparin pump? Epidural haematoma may occur after epidural and spinal regional anaesthesia. PMID- 10523087 TI - The end of the heparin pump? Low molecular weight heparins have practical advantages, but clinical advantages are small. PMID- 10523088 TI - Pleasing both authors and readers. Summary of electronic responses. PMID- 10523089 TI - Structuring the discussion of scientific papers. Wouldn't structured discussions be taking things too far? PMID- 10523090 TI - Structuring the discussion of scientific papers. Randomised controlled trial of structured discussions is needed. PMID- 10523091 TI - What is allodynia? PMID- 10523092 TI - When doctors might kill their patients. Concept of intent is being defined inconsistently by courts. PMID- 10523093 TI - When doctor's might kill their patients. Moral character of clinicians and best interests of patients cannot be separated. PMID- 10523094 TI - When doctors might kill their patients. Patients must never be left to suffer so that doctors "stay out of trouble". PMID- 10523095 TI - Effectiveness of rivastigmine in Alzheimer's disease. Patients' view on quality of life should be assessed. PMID- 10523096 TI - Effectiveness of rivastigmine in Alzheimer's disease. Guidelines do not ignore clinically relevant end points. PMID- 10523097 TI - Screening and mortality from cervical cancer. Study shows importance of centralised organisation in screening. PMID- 10523098 TI - Form 990. Disclosure as PR. PMID- 10523099 TI - Medicare HMOs. Risk retreat. PMID- 10523100 TI - Hospital libraries. Small print. PMID- 10523101 TI - Medical foods? Label loophole. PMID- 10523102 TI - Diagnostic imaging. Pictures of health. PMID- 10523103 TI - ER diverts. Going nowhere fast. PMID- 10523104 TI - Spend tobacco settlement funds outside health care? PMID- 10523105 TI - Call centers. No-show showdown. PMID- 10523106 TI - Patient pleasers. Baby's first keeper. PMID- 10523107 TI - Weighing capitation's cost. PMID- 10523108 TI - Start-ups. Power to the pad. PMID- 10523109 TI - Teen line. Cyber pals. PMID- 10523110 TI - [Lung volume reducing operations in the surgical treatment of pulmonary emphysema]. PMID- 10523112 TI - Reflex anoxic seizures and anaesthesia. PMID- 10523111 TI - Masked bleeding posttonsillectomy with ondansetron. PMID- 10523113 TI - Heart failure and ST segment depression in a child aged 6 weeks. PMID- 10523114 TI - A novel technique for securing lumbar epidural catheters in children undergoing hip surgery. PMID- 10523115 TI - West Point cardiology. PMID- 10523116 TI - Copper associated liver diseases in infancy and childhood. Proceedings of the 4th Emma-Thaler Symposium held at the Department of Paediatrics, Munich University Medical School, Dr. von Hauner'sches Kinderspital, Munich, Germany. PMID- 10523117 TI - Copper associated liver diseases in infancy and childhood. Proceedings of the 4th Emma-Thaler Symposium held at the Department of Paediatrics, Munich University Medical School, Dr. von Hauner'sches Kinderspital, Munich, Germany. PMID- 10523118 TI - Copper associated liver diseases in infancy and childhood. Proceedings of the 4th Emma-Thaler Symposium held at the Department of Paediatrics, Munich University Medical School, Dr. von Hauner'sches Kinderspital, Munich, Germany. PMID- 10523119 TI - Comment: Further evidence for the genetic basis of copper associated pathological conditions. PMID- 10523120 TI - Interictal and postictal cognitive changes in migraine. PMID- 10523121 TI - Primary care in a health maintenance organization. PMID- 10523122 TI - Ethics and professional responsibility. PMID- 10523123 TI - Jerome M. Vaeth, M.D. 1925-1998. President of the American Radium Society in 1974. PMID- 10523124 TI - Fibrinolysis in patients with fulminant hepatic failure. PMID- 10523125 TI - Office of Research Integrity: a reflection of disputes and misunderstandings. AB - Each year, the U.S. Public Health Service (PHS) provides billions of dollars to support over 30,000 extramural research grants to more than 2,000 institutions in the U.S. and other countries. The Office of Research Integrity (ORI) is responsible for protecting the integrity of the research supported by the grants awarded for the PHS extramural research program. One of its responsibilities includes monitoring investigations into alleged or suspected scientific misconduct by institutions that receive the PHS funds. However, not all of the alleged or suspected scientific misconduct meet the the PHS definition of scientific misconduct. Among the wide range of allegations that the ORI receives are those that are ultimately determined to be authorship disputes. This article will report on ORI's functions and review some of the commonly reported allegations that do not constitute scientific misconduct according to the PHS definition. PMID- 10523127 TI - Metered dose inhaler (MDI) systems. PMID- 10523126 TI - Migration of a PEJ tube following insertion: a bizarre complication. PMID- 10523128 TI - Bibliography. Current world literature. Perioperative nutrition. PMID- 10523129 TI - Update treatment of multidrug-resistant tuberculosis. Introduction, discussion and summary. PMID- 10523130 TI - Environmental tobacco smoke epidemiology. PMID- 10523131 TI - New cotton clothes are healthier. PMID- 10523132 TI - The plane truth about disinsection. PMID- 10523133 TI - The coast is cleaner. PMID- 10523134 TI - Protecting schools from pesticides. PMID- 10523135 TI - IUPAC-IUBMB Joint Commission on Biochemical Nomenclature (JCBN) and Nomenclature Committee of IUBMB (NC-IUBMB), newsletter 1999. PMID- 10523137 TI - Generalized onycholysis associated with sodium valproate therapy. PMID- 10523136 TI - Classification of the epilepsies: time for a change? A critical review of the International Classification of the Epilepsies and Epileptic Syndromes (ICEES) and its usefulness in clinical practice and epidemiological studies of epilepsy. AB - The Commission on Classification and Terminology of the International League against Epilepsy (ILAE) first devised a comprehensive classification for the epilepsies and epileptic syndromes nearly 30 years ago. Despite subsequent revisions, the classification remains too complicated to be of utility in clinical practice and epidemiological research. Recent developments in neuro imaging and neurogenetics have also contributed to the limited usefulness of the current International Classification of the Epilepsies and Epileptic Syndromes (ICEES). This review examines the evolution, advantages, and notable disadvantages of the ICEES and assesses its previous application in several population-based studies of epilepsy. The important need for a new, simplified, and aetiologically orientated classification which is amenable to use outside of the tertiary epilepsy centre is discussed. PMID- 10523138 TI - A case of cerebral fat embolism demonstrating no pathophysiological involvement of lung dysfunction. PMID- 10523139 TI - H+ -PPases: a tightly membrane-bound family. AB - The earliest known H+-PPase (proton-pumping inorganic pyrophosphatase), the integrally membrane-bound H+-PPi synthase (proton-pumping inorganic pyrophosphate synthase) from Rhodospirillum rubrum, is still the only alternative to H+-ATP synthase in biological electron transport phosphorylation. Cloning of several higher plant vacuolar H+-PPase genes has led to the recognition that the corresponding proteins form a family of extremely similar proton-pumping enzymes. The bacterial H+-PPi synthase and two algal vacuolar H+-PPases are homologous with this family, as deduced from their cloned genes. The prokaryotic and algal homologues differ more than the H+-PPases from higher plants, facilitating recognition of functionally significant entities. Primary structures of H+-PPases are reviewed and compared with H+-ATPases and soluble PPases. PMID- 10523140 TI - A prospective randomized comparative trial showing that omeprazole prevents rebleeding in patients with bleeding peptic ulcers after successful endoscopic therapy. PMID- 10523141 TI - "Salvage" transjugular intrahepatic portosystemic shunt: gastric fundal compared with esophageal variceal bleeding. PMID- 10523142 TI - Endoscopic palliation of inoperable cancer of the esophagus by argon electrocoagulation. PMID- 10523143 TI - Does hybrid lethality depend on sex or genotype? PMID- 10523145 TI - Helping others help themselves. PMID- 10523144 TI - Chest volume and shape and intrapleural pressure in microgravity. PMID- 10523146 TI - Maternal psychological distress and parenting stress after the birth of a very low-birth-weight infant. PMID- 10523147 TI - Neuropsychological sequelae of haemolytic uraemic syndrome. PMID- 10523148 TI - Buccal midazolam and rectal diazepam for treatment of prolonged seizures in childhood and adolescence: a randomised trial. PMID- 10523149 TI - Liposomal amphotericin B for empirical therapy in patients with persistent fever and neutropenia. PMID- 10523150 TI - Endogenous virus of BHK-21 cells complicates electron microscopy studies of foamy virus maturation. PMID- 10523151 TI - Problems with new criteria for diagnosis of diabetes mellitus. PMID- 10523152 TI - Hospital in the home: a randomised controlled trial. PMID- 10523153 TI - Transplantation of thymus tissue in complete DiGeorge syndrome. AB - BACKGROUND: The DiGeorge syndrome is a congenital disorder that affects the heart, parathyroid glands, and thymus. In complete DiGeorge syndrome, patients have severely reduced T-cell function. METHODS: We treated five infants (age, one to four months) with complete DiGeorge syndrome by transplantation of cultured postnatal thymus tissue. Follow-up evaluations included immune phenotyping and proliferative studies of peripheral-blood mononuclear cells plus biopsy of the thymus allograft. Thymic production of new T cells was assessed in peripheral blood by tests for T-cell-receptor recombination excision circles, which are formed from excised DNA during the rearrangement of T-cell-receptor genes. RESULTS: After the transplantation of thymus tissue, T-cell proliferative responses to mitogens developed in four of the five patients. Two of the patients survived with restoration of immune function; three patients died from infection or abnormalities unrelated to transplantation. Biopsies of grafted thymus in the surviving patients showed normal morphologic features and active T-cell production. In three patients, donor T cells could be detected about four weeks after transplantation, although there was no evidence of graft-versus-host disease on biopsy or at autopsy. In one patient, the T-cell development within the graft was demonstrated to accompany the appearance of recently developed T cells in the periphery and coincided with the onset of normal T-cell function. In one patient, there was evidence of thymus function and CD45RA+CD62L+ T cells more than five years after transplantation. CONCLUSIONS: In some infants with profound immunodeficiency and complete DiGeorge syndrome, the transplantation of thymus tissue can restore normal immune function. Early thymus transplantation - before the development of infectious complications - may promote successful immune reconstitution. PMID- 10523154 TI - The chances for health care reform. PMID- 10523155 TI - The chances for health care reform. PMID- 10523156 TI - The chances for health care reform. PMID- 10523157 TI - The chances for health care reform. PMID- 10523158 TI - Ultrasound therapy for calcific tendinitis of the shoulder. PMID- 10523159 TI - Prospective evaluation of a patient with Trypanosoma cruzi infection transmitted by transfusion. PMID- 10523160 TI - Endocarditis due to Streptococcus mitis with high-level resistance to penicillin and cefotaxime. PMID- 10523161 TI - Effect of highly active antiretroviral therapy on thrombocytopenia in patients with HIV infection. PMID- 10523162 TI - Tissue plasminogen activator for acute ischemic stroke. PMID- 10523163 TI - HHV-8 peripheral-blood viral load and the titer of antibodies against HHV-8. PMID- 10523164 TI - Interventional sialendoscopy. PMID- 10523165 TI - Building on the past to create nursing's future. PMID- 10523166 TI - News from NINR. PMID- 10523167 TI - Systems thinking, archetypes and interventions. PMID- 10523168 TI - Has foreign nurse recruitment impeded African American access to nursing education and practice? PMID- 10523169 TI - Too sleepy for surgery. PMID- 10523170 TI - Do we need to wake up to the sleep apnea problem? PMID- 10523171 TI - Assessment of sleep complaints and sleep-disordered breathing in a consecutive series of obese patients. AB - BACKGROUND/OBJECTIVE: The prevalence of sleep-related complaints (SRC) and the frequency of sleep-disordered breathing (SDB) in obese patients has not been studied extensively. We investigated SRC and SDB in a group of obese persons as part of a preoperative workup for weight reduction (bariatric) surgery. METHODS: All consecutive patients attending a weight-loss clinic for evaluation for bariatric surgery were asked to complete a questionnaire. The questionnaire consisted of a section on SRC and a validated general sleep questionnaire (Sleep Wake Experience List). The patients underwent sleep studies in which an Edentrace recorder registered heart rate, chest wall movements by impedance, airflow and oxygen saturation. RESULTS: Fifty-one patients (14 men, 37 women) were evaluated. Mean body mass index (BMI) was 45 kg/m2 (range 33-61). Eighteen patients (35%) demonstrated SDB, defined as (a) an apnea/hypopnea index 5, and/or (b) more than 2% of registration time with an oxygen saturation below 90%. There was no difference between these 18 patients and patients who did not exhibit SDB in age, sex, BMI or SRC. Seven patients had SDB of a severity warranting closer investigation and perioperative monitoring. CONCLUSION: Both SRC and SDB are common in obese patients. Limited nocturnal respiratory monitoring is indicated as part of the preoperative workup for weight reduction surgery. PMID- 10523172 TI - Chronic persistent cough due to Gilles de la Tourette's syndrome. PMID- 10523173 TI - Global warming and efficacy of therapeutic agents for respiratory diseases. PMID- 10523174 TI - Pharyngeal cross-sectional area and compliance in normal males and females. PMID- 10523175 TI - The ozone layer. Burnt by the sun down under. PMID- 10523176 TI - Research lab to surrender chimps. PMID- 10523178 TI - View from the top of a biomedical empire. Interview by Eliot Marshall. PMID- 10523177 TI - Cancer research. A new way to combat therapy side effects. PMID- 10523179 TI - A threat to biomedical research. PMID- 10523180 TI - Early plant history: something borrowed, something new? PMID- 10523181 TI - Infants learning algebraic rules. PMID- 10523182 TI - Likelihood of NIH extramural funding. PMID- 10523183 TI - Overlooked control. PMID- 10523184 TI - Who's balancing the federal research portfolio and how? PMID- 10523185 TI - Perspectives: signal transduction. Proteins in motion. PMID- 10523186 TI - Ribozymes in the nucleolus. PMID- 10523187 TI - Making brain circuits listen. PMID- 10523188 TI - Team wrapping up sequence of first human chromosome. PMID- 10523189 TI - Turnover at the top at Cell and NEJM. Cell editor step down. PMID- 10523190 TI - Turnover at the top at Cell and NEJM. NEJM publisher resigns. PMID- 10523191 TI - Ambitious clinical trial stirs debate. PMID- 10523193 TI - AIDS researchers blast NIH peer review plan. PMID- 10523192 TI - On the way to a better immunosuppressant? PMID- 10523194 TI - University of Illinois. Chancellor quits after research shutdown. PMID- 10523195 TI - The race to the ribosome structure. PMID- 10523196 TI - Meeting. American Chemical Society. Raising a glass to health and nanotubes. PMID- 10523197 TI - Ethical dilemmas and stem cell research. PMID- 10523198 TI - Smallpox vaccine. PMID- 10523199 TI - Testing the genetics of behavior in mice. PMID- 10523200 TI - Testing the genetics of behavior in mice. PMID- 10523201 TI - Testing the genetics of behavior in mice. PMID- 10523202 TI - Testing the genetics of behavior in mice. PMID- 10523203 TI - Testing the genetics of behavior in mice. PMID- 10523204 TI - Policy forum: human genetics. Ethical considerations in leaping from bench to bedside. PMID- 10523205 TI - Narcolepsy genes wake up the sleep field. PMID- 10523206 TI - Function is structure. PMID- 10523208 TI - Prospects for in utero human gene therapy. AB - Gene therapy for the treatment of disease in children and adults is being actively pursued at many medical centers. However, a number of genetic disorders result in irreversible damage to the fetus before birth. In these cases, as well as for those with genetic diseases who may benefit from therapy before symptoms are manifested, in utero gene therapy (IUGT) could be beneficial. Although some successes with in utero gene transfer have been reported in animals, significant questions remain to be answered before IUGT clinical trials would be acceptable. This review analyzes the state of the art and delineates the studies that still need to be performed before it would be appropriate to consider human IUGT. PMID- 10523207 TI - Perspectives: cell biology. All creatures great and small. PMID- 10523209 TI - [The effect of alcohol on the cardiovascular system]. PMID- 10523210 TI - [Platelet function]. PMID- 10523211 TI - Beta agonist dose reduction in asthma. PMID- 10523212 TI - Effect of inhaled corticosteroid therapy on bone markers and bone density. PMID- 10523213 TI - Summer tuberculosis. PMID- 10523214 TI - Ischemic stroke and tissue hypodensity on computed tomography. PMID- 10523215 TI - Transmission of tuberculosis in a jail. AB - BACKGROUND: Outbreaks of tuberculosis are uncommonly recognized in jails. In 1996, an increase in active tuberculosis cases was noted among inmates of a large urban jail. OBJECTIVES: To determine the source and extent of a tuberculosis outbreak in an urban jail and to recommend control measures. DESIGN: Retrospective cohort study. SETTING: Urban jail. PATIENTS: Inmates and guards with tuberculosis. INTERVENTION: Outbreak evaluation and control. MEASUREMENTS: Medical records of inmates and guards with tuberculosis were reviewed, and inmates were interviewed. DNA fingerprinting was performed on Mycobacterium tuberculosis isolates. RESULTS: From 1 January 1995 through 31 December 1997, active tuberculosis was diagnosed in 38 inmates and 5 guards from the jail. Nineteen (79%) of the 24 culture-positive inmates had isolates with DNA fingerprints matching those of other inmates. Isolates from both culture-positive guards matched the predominant inmate strain; only 6 (14%) of 43 isolates from infected persons in the community had this pattern. The median length of incarceration of all inmates in the jail was 1 day; the median length of continuous incarceration before diagnosis of tuberculosis in inmates was 138 days. Inmates with tuberculosis had been incarcerated a median of 15 times. Forty three percent of persons in this city with tuberculosis diagnosed from January 1995 through July 1997 had been incarcerated in the jail at some time before diagnosis. CONCLUSIONS: Traditional and molecular epidemiologic investigations suggest that tuberculosis was transmitted among inmates and guards in an urban jail. Aggressive measures to screen for active tuberculosis upon incarceration are important for preventing spread of disease in jails and to the surrounding community. PMID- 10523216 TI - Cardiac and arterial target organ damage in adults with elevated ambulatory and normal office blood pressure. AB - BACKGROUND: Ambulatory blood pressure may be higher or lower than clinic blood pressure. Attention has focused on "white coat hypertension" (normal ambulatory blood pressure elevated in the clinic). The converse phenomenon of high ambulatory blood pressure but normal office blood pressure-"white coat normotension"-has not been studied. OBJECTIVE: To assess whether white coat normotension (awake ambulatory blood pressure > 134/90 mm Hg and clinic blood pressure < 140/90 mm Hg) is associated with target organ damage. DESIGN: Cross sectional observational study. SETTING: University hospital hypertension center and participant work sites. PATIENTS: 295 clinically normotensive adults and 64 patients with sustained hypertension (elevated clinic and ambulatory blood pressure). MEASUREMENTS: Target organ abnormalities were measured by echocardiography and arterial ultrasonography in 61 patients with white coat normotension, 234 with sustained normotension (normal clinic and ambulatory blood pressure), and 64 with sustained hypertension. RESULTS: Patients with white coat normotension were older; had higher body mass indices, serum creatinine concentrations, and glucose levels; and a higher prevalence of current smokers. Left ventricular mass index and relative wall thickness were higher by 13 g/m2 (CI, 8 to 18 g/m2) and by 0.03 (CI, 0.01 to 0.04), respectively, in patients with white coat normotension compared with those who had sustained normotension. Patients with white coat normotension and those with sustained hypertension did not differ significantly for left ventricular mass index (4 g/m2 [CI, - 3 to 10 g/m2) or relative wall thickness (0.01 [CI, -0.01 to 0.03]). The prevalence of discrete atherosclerotic plaques was similar in patients with white coat normotension (17 of 61, or 28% [CI, 17% to 39%]) and those with sustained hypertension (17 of 64, or 27% [CI, 16% to 38%]), but the difference lost significance after adjustment for age. CONCLUSIONS: White coat normotension is associated with left ventricular mass and carotid wall thickness similar to those in sustained hypertension. The association of white coat normotension with prognostically important target organ damage may partly explain the ability of high normal left ventricular mass and high normal clinic blood pressure to predict subsequent hypertension and cardiovascular events in patients with clinical normotension. PMID- 10523217 TI - Baseline IgG antibody titers to Chlamydia pneumoniae, Helicobacter pylori, herpes simplex virus, and cytomegalovirus and the risk for cardiovascular disease in women. AB - BACKGROUND: Results of cross-sectional and retrospective studies have suggested that chronic infection may be a risk factor for cardiovascular disease. However, prospective data evaluating the relation between baseline antibody titers against various plausible agents and risk for cardiovascular disease are sparse, particularly among women. OBJECTIVE: To determine whether previous exposure to Chlamydia pneumoniae, Helicobacter pylori, herpes simplex virus, or cytomegalovirus is associated with increased risk for cardiovascular events. DESIGN: Prospective, nested, case-control study. SETTING: Women's Health Study. PARTICIPANTS: Apparently healthy postmenopausal women. MEASUREMENTS: IgG antibody titers against C. pneumoniae, H. pylori, herpes simplex virus, and cytomegalovirus were measured in baseline blood samples obtained from 122 study participants who subsequently reported a first cardiovascular event (case patients) and 244 participants matched for age and smoking status who did not report a cardiovascular event (controls) during 3 years of follow-up. RESULTS: Little evidence was found of an association between risk for cardiovascular events and baseline IgG seropositivity for antibodies against C. pneumoniae (rate ratio, 1.1 [95% CI, 0.7 to 1.8]), H. pylori (rate ratio, 0.90 [CI, 0.6 to 1.4]), herpes simplex virus (rate ratio, 1.2 [CI, 0.6 to 2.1]), and cytomegalovirus (rate ratio, 0.9 [CI, 0.6 to 1.5]). In addition, there was little evidence of an association between a participant's total number of infections and subsequent cardiovascular risk (P > 0.2). CONCLUSION: In apparently healthy postmenopausal women, little evidence was found that previous infection, as measured by IgG antibody titers to C. pneumoniae, H. pylori, herpes simplex virus, and cytomegalovirus, is associated with subsequent risk for cardiovascular disease. PMID- 10523218 TI - A qualitative analysis of how physicians with expertise in domestic violence approach the identification of victims. AB - BACKGROUND: Physicians have been called upon to identify victims of domestic violence, but few studies provide insight into how physicians can navigate around the barriers to identification. OBJECTIVE: To describe how physicians who are committed to helping battered patients identify victims of domestic violence in health care encounters. DESIGN: Six focus groups were conducted. SETTING: Focus group research facilities. PARTICIPANTS: 45 emergency department, obstetrician/ gynecologist, and primary care physicians in the San Francisco Bay Area who identify and intervene with victims of domestic violence. MEASUREMENTS: Through constant comparison, a template of open codes was constructed to identify themes that emerged from the data. Data were analyzed according to the conventions of qualitative research. RESULTS: The data revealed five major themes: 1) how physicians framed screening questions to reduce patient discomfort; 2) patient signs that "switched on a light bulb" for physicians to suspect abuse; 3) direct and indirect approaches to identification, with an emphasis on facilitating patient trust and disclosure over time; 4) the rarity of direct patient disclosure; and 5) how physicians redefined successful outcomes of universal screening. Physicians also described two new barriers to screening: mandatory reporting and "burnout" due to lack of direct disclosure. CONCLUSIONS: Identifying domestic abuse is difficult even for physicians committed to helping victims. Physician reports illustrate the need to frame questions and develop indirect approaches that foster patient trust. Given the many barriers to screening and the rarity of direct patient disclosure, it may be more productive to redefine the goals of universal screening so that compassionate asking in and of itself constitutes the first step in helping battered patients. PMID- 10523219 TI - Paradoxical response to dexamethasone in the diagnosis of primary pigmented nodular adrenocortical disease. AB - BACKGROUND: Primary pigmented nodular adrenocortical disease causes the Cushing syndrome in children and young adults and is most frequently associated with the Carney complex. OBJECTIVE: To evaluate diagnostic tests for primary pigmented nodular adrenocortical disease. DESIGN: Retrospective cohort study. SETTING: Tertiary care center. PATIENTS: 21 patients with primary pigmented nodular adrenocortical disease. The control groups consisted of 9 patients with macronodular adrenocortical disease and 15 patients with primary unilateral adrenocortical disease (single adenomas). MEASUREMENTS: Clinical characteristics, radiologic imaging, and a 6-day Liddle test with determination of urinary free cortisol and 17-hydroxycorticosteroid excretion. RESULTS: Adrenal imaging and other tests were of limited value for the diagnosis of primary pigmented nodular adrenocortical disease. The Liddle test, however, distinguished patients with this disorder from those with other primary adrenocortical lesions. An increase of 50% or more in urinary free cortisol levels on day 6 of the Liddle test identified 9 of 13 patients (69.2% [95% CI, 46.6% to 91.8%]) with primary pigmented nodular adrenocortical disease, excluded all patients with macronodular adrenocortical disease, and was present in only 3 of the 15 patients with single adrenocortical adenomas (20% [CI, 0% to 40.2%]). An increase in urinary free cortisol excretion of 100% or more on day 6 of the Liddle test identified only patients with primary pigmented nodular adrenocortical disease. CONCLUSIONS: Patients with primary pigmented nodular adrenocortical disease responded to dexamethasone with a paradoxical increase in glucocorticoid excretion during the Liddle test. This feature distinguishes such patients from those who have the Cushing syndrome caused by other primary adrenal disorders and may lead to timely detection of the Carney complex (a potentially fatal disorder) in asymptomatic patients. PMID- 10523220 TI - Partial-liver transplantation to treat familial amyloid polyneuropathy: follow-up of 11 patients. AB - BACKGROUND: Recently, liver transplantation has been used to treat patients with familial amyloid polyneuropathy (FAP). OBJECTIVE: To describe the clinical course of patients with FAP who received partial-liver transplantation from living donors. DESIGN: Case series. SETTING: University hospital in Matsumoto, Japan. PATIENTS: 11 patients with FAP who underwent partial-liver transplantation. The transthyretin gene abnormality in all 11 patients was the substitution of methionine for valine at position 30. INTERVENTION: Partial liver transplantation from living donors. MEASUREMENTS: Preoperative and follow-up (3 to 64 months) clinical data, including routine laboratory data, nerve conduction velocity tests, and sural nerve histology. RESULTS: All 7 patients who had severe gastrointestinal autonomic disorders or polyneuropathy localized to the lower limbs for less than 4 years showed improvement. Three of 4 patients with polyneuropathy involving both the upper and lower limbs had adverse outcomes, including two deaths. The preoperative duration of their illness was more than 6 years. These 3 patients also had marked decreases in creatinine clearance and nerve conduction velocities and severe loss of myelinated fibers in sural nerves. CONCLUSION: Preoperative clinical severity and duration of illness are associated with outcomes after liver transplantation for FAP. PMID- 10523221 TI - Update in pulmonary diseases. PMID- 10523222 TI - Prescribing hormone replacement therapy for menopausal symptoms. AB - This paper addresses the clinical presentation of menopause, pretreatment assessment for hormone replacement therapy, benefits and risks of this treatment, common hormone replacement regimens and their side effects, and patient management. The case-based discussion focuses on the clinical management of a patient who is considering hormone replacement therapy for menopausal symptoms. PMID- 10523223 TI - On target: a tuberculosis control strategy whose time has come. PMID- 10523224 TI - Integrating routine inquiry about domestic violence into daily practice. PMID- 10523225 TI - Influenza prevention and treatment: current practices and new horizons. PMID- 10523226 TI - The sharer. PMID- 10523227 TI - A visit to the doctor. PMID- 10523228 TI - Prediction equation for glomerular filtration rate. PMID- 10523229 TI - Prediction equation for glomerular filtration rate. PMID- 10523230 TI - Prediction equation for glomerular filtration rate. PMID- 10523231 TI - Biliary sludge. PMID- 10523232 TI - Biliary sludge. PMID- 10523233 TI - When doctors marry doctors. PMID- 10523234 TI - Subjective compared with objective sleepiness. PMID- 10523235 TI - White coat pockets. PMID- 10523236 TI - Ocular venous occlusion and hyperhomocysteinemia. PMID- 10523237 TI - Late clonal complications in older patients receiving immunosuppressive therapy for aplastic anemia. PMID- 10523238 TI - Autoimmune skin rashes associated with etanercept for rheumatoid arthritis. PMID- 10523239 TI - Beyond HIV viral load testing. PMID- 10523240 TI - There's no place like home: the home health care alternative. PMID- 10523241 TI - Perception of need for emergency admission to hospital. PMID- 10523242 TI - Clinical and laboratory findings in referrals for mitochondrial DNA analysis. PMID- 10523243 TI - Improving communication between doctors and patients. PMID- 10523244 TI - Cancer in Sotos syndrome. PMID- 10523245 TI - Influence of five years of antenatal screening on the paediatric cystic fibrosis population in one region. PMID- 10523246 TI - Serum bicarbonate and the severity of dehydration in gastroenteritis. PMID- 10523247 TI - The risks and benefits of cisapride. PMID- 10523248 TI - Echocardiography by a neonatologist. PMID- 10523249 TI - Empyema thoracis: a role for open thoracotomy and decortication. PMID- 10523250 TI - Guidelines for managing acute gastroenteritis. PMID- 10523251 TI - Guidelines for managing acute gastroenteritis. PMID- 10523252 TI - Management of tuberculosis in Wales: 1986-92. PMID- 10523253 TI - Reduction of plasma concentrations of large neutral amino acids in patients with maple syrup urine disease during crises. PMID- 10523254 TI - The vitamin K debacle. PMID- 10523255 TI - Chlamydia pneumoniae infection in children with persistent cough. PMID- 10523256 TI - Chronic cough: is it asthma? PMID- 10523257 TI - Risks and benefits of cisapride. PMID- 10523258 TI - Colonic wall thickness and pancreatic enzymes in cystic fibrosis. PMID- 10523259 TI - Unnatural sudden infant death. PMID- 10523260 TI - Unnatural sudden infant death. PMID- 10523261 TI - Unnatural sudden infant death. PMID- 10523262 TI - J.B. Georg W. Fresenius and the description of the species Aspergillus fumigatus in 1863. PMID- 10523263 TI - The genus Aspergillus with special regard to the Aspergillus fumigatus group. PMID- 10523264 TI - Pathogenesis and clinical presentation of aspergillosis. PMID- 10523265 TI - Epidemiology and molecular basis of the involvement of Aspergillus fumigatus in allergic diseases. PMID- 10523266 TI - Host defense mechanism in Aspergillus fumigatus infections. PMID- 10523267 TI - Antigen and DNA patterns characteristic of Aspergillus fumigatus. PMID- 10523268 TI - Laboratory diagnosis of Aspergillus fumigatus-associated diseases. PMID- 10523269 TI - Therapy of Aspergillus fumigatus-related diseases. PMID- 10523270 TI - Animal models of A. fumigatus infections. PMID- 10523271 TI - Problems of antifungal in vitro testing in Aspergillus fumigatus. PMID- 10523272 TI - Transformation systems of Aspergillus fumigatus. New tools to investigate fungal virulence. PMID- 10523273 TI - Interactions between Aspergillus fumigatus and host matrix proteins. PMID- 10523274 TI - Aspergillus fumigatus-secreted proteases as antigenic molecules and virulence factors. PMID- 10523275 TI - Chitin synthase genes of Aspergillus species. PMID- 10523276 TI - Pigment biosynthesis and virulence. PMID- 10523277 TI - The influence of microgravity and spaceflight on columella cell ultrastructure in starch-deficient mutants of Arabidopsis. AB - The ultrastructure of root cap columella cells was studied by morphometric analysis in wild-type, a reduced-starch mutant, and a starchless mutant of Arabidopsis grown in microgravity (F-microgravity) and compared to ground 1g (G 1g) and flight 1g (F-1g) controls. Seedlings of the wild-type and reduced-starch mutant that developed during an experiment on the Space Shuttle (both the F microgravity samples and the F-lg control) exhibited a decreased starch content in comparison to the G-1g control. These results suggest that some factor associated with spaceflight (and not microgravity per se) affects starch metabolism. Elevated levels of ethylene were found during the experiments on the Space Shuttle, and analysis of ground controls with added ethylene demonstrated that this gas was responsible for decreased starch levels in the columella cells. This is the first study to use an on-board centrifuge as a control when quantifying starch in spaceflight-grown plants. Furthermore, our results show that ethylene levels must be carefully considered and controlled when designing experiments with plants for the International Space Station. PMID- 10523279 TI - Frequency of somaclonal variation in plants of black spruce (Picea mariana, Pinaceae) and white spruce (P. glauca, Pinaceae) derived from somatic embryogenesis and identification of some factors involved in genetic instability. AB - Plants of black spruce (Picea mariana, N = 7047 individuals) and white spruce (P. glauca, N = 3995 individuals) were regenerated from a total of 87 clones over a 5 yr period by somatic embryogenesis to study factors that might be associated with the appearance of variant phenotypes. Morphological evaluation of the plants showed several types of variation. These variations were grouped into nine types: dwarfism (type A), reduced height with various form anomalies (types B, C, and D), needle fasciation (type E), abnormality in tree architecture (type F), variegata phenotype (type G), and plants with an overall regular morphology but smaller than normal plants (type H). Plagiotropic plants were also observed (type I). Each plant from types A to H (except type C where no plants survived more than 6 mo) had retained its phenotype over 4-5 yr of growth. Some of the variant types could be related to chromosomic instability: chromosome counts showed aneuploid cells for type-A and type-D plants. The type I (plagiotropism) was not related to genetic instability but rather to physiological disorders. In total, spruce variants of types A-H were obtained at relatively low frequencies, i.e., 1.0% (39/3995) for white spruce and 1.6% (110/7047) for black spruce. Statistical analyses, conducted with family, clone, and time in maintenance as variables, showed that clone was the most important source of genetic instability followed by time in maintenance. PMID- 10523278 TI - The relationship between xylem conduit diameter and cavitation caused by freezing. AB - The centrifuge method for measuring the resistance of xylem to cavitation by water stress was modified to also account for any additional cavitation that might occur from a freeze-thaw cycle. A strong correlation was found between cavitation by freezing and mean conduit diameter. On the one extreme, a tracheid bearing conifer and diffuse-porous angiosperms with small-diameter vessels (mean diameter <30 MUm) showed no freezing-induced cavitation under modest water stress (xylem pressure = -0.5 MPa), whereas species with larger diameter vessels (mean >40 MUm) were nearly completely cavitated under the same conditions. Species with intermediate mean diameters (30-40 MUm) showed partial cavitation by freezing. These results are consistent with a critical diameter of 44 MUm at or above which cavitation would occur by a freeze-thaw cycle at -0.5 MPa. As expected, vulnerability to cavitation by freezing was correlated with the hydraulic conductivity per stem transverse area. The results confirm and extend previous reports that small-diameter conduits are relatively resistant to cavitation by freezing. It appears that the centrifuge method, modified to include freeze-thaw cycles, may be useful in separating the interactive effects of xylem pressure and freezing on cavitation. PMID- 10523280 TI - Structure and development of the pitchers from the carnivorous plantNepenthes alata (Nepenthaceae). AB - The pitchers of the tropical carnivorous plant Nepenthes alata are highly specialized organs for the attraction and capture of insects and absorption of nutrients from them. This study examined the structure and development of these pitchers, with particular focus on the nectaries and digestive glands. Immature pitchers developed at the tips of tendrils and were tightly sealed by a lid structure that opened during the end of pitcher elongation. Opened pitchers exposed a ridged peristome containing large nectaries. Like other members of the genus, a thick coating of epicuticular waxy scales covered the upper one-third of the pitcher. Scattered within this zone were cells resembling a stomatal complex with a protruding ridge. Cross sections showed that this ridge was formed by asymmetric divisions of the epidermal cells and lacked an underlying pore. The basal region of the trap had large multicellular glands that developed from single epidermal cells. These glands were closely associated with underlying vascular traces and provided a mechanism for supplying fluid to closed immature pitchers. PMID- 10523281 TI - "Lycostrobus" chinleana, an equisetalean cone from the Upper Triassic of the southwestern United States and its phylogenetic implications. AB - Detailed study of the cone Lycostrobus chinleana Daugherty shows that the fossil was incorrectly attributed to the Lycopodiales by the author and to the quillworts by Retallack and that it actually should be assigned to the Equisetales. The cone, which occurs in the Upper Triassic Chinle Formation at several localities in the southwestern United States, is ~2.5 cm wide and nearly 6 cm long and consists of a stout axis bearing whorls of peltate sporangiophores. Each sporangiophore is composed of a slender stalk and a hexagonal disk, which typically bears a single, generally long, lanceolate, forward-directed leaf-like umbo tip on the outer surface and several recurrent sporangia on the inner surface. Small round to oval trilete spores occur in the sporangia. Since the leaf-like umbo tip is similar to the sterile leaves found in certain calamite cones and the recurrent sporangia are equisetalean-like, it appears that the cone may represent a intermediate stage between Calamites and modern Equisetum. According to this hypothesis, the nonbracteate Equisetum cone could have developed from a bracteate calamite cone, through reduction and fusion of the bracts and the sporangiophores, rather than by the loss of whorls of bracts of the Calamites cone as suggested earlier by others. As a result of this study the cone is assigned to the new Equisetalean genus Equicalastrobus and redescribed under the name E. chinleana (Daugherty) Grauvogel-Stamm and Ash, n. comb. PMID- 10523282 TI - Reproductive biology and conservation genetics of Goodyera procera (Orchidaceae). AB - Goodyera procera is an endangered terrestrial orchid in Hong Kong. Information on its reproductive biology and pattern of genetic variation is needed to develop efficient conservation strategies. Pollination experiments showed that the species is self-compatible, but dependent on pollinators for fruit set. Bagged plants produced no fruits. Artificial pollinations resulted in 92% fruit set through selfing, 94% with geitonogamous pollination, and 95% following xenogamous pollination. Fruit set in the open-pollinated control was 75% at the same sites. Allozyme electrophoresis and random amplified polymorphic DNA (RAPD) were used to evaluate genetic variation and structure of 15 populations of Goodyera procera. Despite its outbreeding system, allozyme data revealed low variation both at the population (P = 21.78%, A = 1.22, and H = 0.073) and species (P = 33%, A = 1.33, and H = 0.15) levels, in comparison with other animal-pollinated outbreeding plant species. However, RAPD variation was relatively high (P = 55.13% and H = 0.18 at the population level, and P = 97.03% and H = 0.29 at the species level). G(ST) estimates indicated high levels of genetic differentiation among populations (G(ST) = 0.52 and I = 0.909 +/- 0.049 based on allozyme data, and G(ST) = 0.39 and I = 0.859 +/- 0.038 based on RAPD data), much above the average for outcrossing species, suggesting that gene flow was limited in this species. Based on these data, suitable strategies were developed for the genetic conservation and management of the species. PMID- 10523284 TI - Cryptic dioecy and leaky dioecy in endemic species of Dombeya (Sterculiaceae) on La Reunion. AB - The high frequency of dioecy on oceanic islands such as Hawaii and New Zealand has attracted a great deal of attention from plant evolutionary biologists. One reason suggested for the high prevalence of dioecy on oceanic islands is that taxa considered truly dioecious may have occasional hermaphrodite flowers, i.e., show leaky dioecy. In this study, we quantified the presence and distribution of leaky dioecy in a group of congeneric endemic species of the genus Dombeya (Sterculiaceae) on La Reunion island (Indian Ocean). All eight species show cryptic dioecy. Five species show strict dioecy and three species show leaky dioecy due to the presence of male trees that set fruit. Species with strict dioecy and large populations tend to occur in mid- to high-altitude moist tropical cloud forest, whereas species in smaller populations at lower altitude and in semidry tropical forest tend to show leaky dioecy. Two reasons for this differential distribution of strict dioecy and leaky dioecy are discussed. First, environmental variation along the altitudinal gradient, biotic and/or abiotic, may influence the breeding system. Second, leaky dioecy may be favored in lowland populations due to the small size and disturbed nature of such populations. PMID- 10523283 TI - Plant reproductive phenology over four years including an episode of general flowering in a lowland dipterocarp forest,Sarawak, Malaysia. AB - The first systematic observation of a general flowering, a phenomenon unique to lowland mixed-dipterocarp forests in Southeast Asia, is presented. During general flowering, which occurs at irregular intervals of 3-10 yr, nearly all dipterocarp species together with species of other families come heavily into flower. We monitored reproductive phenology of 576 individual plants representing 305 species in 56 families in Sarawak, Malaysia. Observations continued for 53 mo from August 1992 and covered one episode of a general flowering cycle. Among 527 effective reproductive events during 43 mo, 57% were concentrated in the general flowering period (GFP) of 10 mo in 1996. We classified 257 species into flowering types based on timing and frequency of flowering. The most abundant type was "general flowering" (35%), which flowered only during GFP. The others were "supra annual" (19%), "annual" (13%), and "sub-annual" (5%) types. General flowering type and temporal aggregation in reproductive events were commonly found among species in various categories of taxonomic groups, life forms, pollination systems, and fruit types. Possible causes for general flowering, such as promotion of pollination brought about by interspecific synchronization and paucity of climatic cues suitable for flowering trigger, are proposed, in addition to the predator satiation hypothesis of Janzen (1974). PMID- 10523285 TI - Chloroplast DNA evidence for the evolution of Microseris (Asteraceae) in Australia and New Zealand after long-distance dispersal from western North America. AB - Restriction site mutations and trnL(UAA)-trnF(GAA) intergenic spacer length variants in the chloroplast genome were used to investigate the phylogenetic relationships among 53 Australian and New Zealand Microseris populations and to assess their position within their primarily North American genus. The study was performed to enhance understanding of evolutionary processes within this unique example of intercontinental dispersal and subsequent adaptive radiation. A southern blot method using four-base restriction enzymes and fragment separation on polyacryamide gels resulted in 55 mutations of which 30 were potentially phylogenetically informative. Most mutations were small indels of <162 bp, 80% of which were <20 bp. The small indels were useful for phylogenetic reconstruction of Australasian Microseris as judged by the high consistency indexes. The results confirmed the monophyly of the Australian and New Zealand Microseris. The occurrence of "hard" basal polytomies in the most parsimonious trees indicated that rapid radiation has occurred early in the history of the taxon. The monophyly of M. lanceolata, which includes the self-incompatible ecotypes of the Australian mainland, was confirmed. Within this species three clades were found that reflect more geographic distribution than morphological entities, suggesting that migration and possibly introgression between different ecotypes, or parallel evolution of similar adaptations, has occurred. One of the three clades was supported by a 162-bp deletion in the trnL-trnF spacer, while a subgroup of this exhibited also a tandemly repeated trnF exon. The data were inconclusive about the monophyly of the second Australasian species, M. scapigera, which comprises the New Zealand, Tasmanian, and autofertile ecotypes of Australia. PMID- 10523286 TI - Light environment, sapling architecture, and leaf display in six rain forest tree species. AB - Architecture and leaf display were compared in saplings of six rain forest tree species differing in shade tolerance. Saplings were selected along the whole light range encountered in a forest environment. Species differed largely in realized height and crown expansion per unit support biomass, but this could not be related to differences in shade tolerance. The results demonstrate that there exist various solutions to an effective expansion of plant height and crown area. It is argued that choice of the study species and the ontogenetic trajectory regarded determine to a large extent the outcome of interspecific comparisons. No evidence was found that pioneers were characterized by a multilayered and shade tolerants by a monolayered leaf distribution. Yet, sun plants had a similar crown area, a deeper crown, and a higher leaf area index compared to shade plants and their leaves were more evenly distributed along the stem. This suggests that differences in leaf layering are found between plants growing in different light environments, rather than between species differing in shade tolerance. PMID- 10523287 TI - Phylogeny of the core Malvales: evidence from ndhF sequence data. AB - The monophyly of the group comprising the core malvalean families, Bombacaceae, Malvaceae, Sterculiaceae, and Tiliaceae, was recently confirmed by molecular studies, but the internal structure of this clade is poorly understood. In this study, we examined sequences of the chloroplast ndhF gene (aligned length 2226 bp) from 70 exemplars representing 35 of the 39 putative tribes of core Malvales. The monophyly of one traditional family, the Malvaceae, was supported in the trees resulting from these data, but the other three families, as traditionally circumscribed, are nonmonophyletic. In addition, the following relationships were well supported: (1) a clade, /Malvatheca, consisting of traditional Malvaceae and Bombacaceae (except some members of tribe Durioneae), plus Fremontodendron and Chiranthodendron, which are usually treated as Sterculiaceae; (2) a clade, /Malvadendrina, supported by a unique 21-bp (base pair) deletion and consisting of /Malvatheca, plus five additional subclades, including representatives of Sterculiaceae and Tiliaceae, and Durionieae; (3) a clade, /Byttneriina, with genera traditionally assigned to several tribes of Tiliaceae, plus exemplars of tribes Byttnerieae, Hermannieae, and Lasiopetaleae of Sterculiaceae. The most striking departures from traditional classifications are the following: Durio and relatives appear to be more closely related to Helicteres and Reevesia (Sterculiaceae) than to Bombacaceae; several genera traditionally considered as Bombacaceae (Camptostemon, Matisia, Phragmotheca, and Quararibea) or Sterculiaceae (Chiranthodendron and Fremontodendron) appear as sister lineages to the traditional Malvaceae; the traditional tribe Helictereae (Sterculiaceae) is polyphyletic; and Sterculiaceae and Tiliaceae, as traditionally circumscribed, represent polyphyletic groups that cannot sensibly be maintained with their traditional limits for purposes of classification. We discuss morphological characters and conclude that there has been extensive homoplasy in characters previously used to delineate major taxonomic groups in core Malvales. The topologies here also suggest that /Malvatheca do not have as a synapormophy monothecate anthers, as has been previously supposed but, instead, may be united by dithecate, transversely septate (polysporangiate) anthers, as found in basal members of both /Bombacoideae and /Malvoideae. Thus, "monothecate" anthers may have been derived at least twice, independently, within the /Bombacoideae (core Bombacaceae) and /Malvoideae (traditional Malvaceae). PMID- 10523288 TI - Polyploid evolution and biogeography in Chelone (Scrophulariaceae): morphological and isozyme evidence. AB - Chelone is a genus of perennial herbs comprising three diploid species (C. cuthbertii, C. glabra, and C. lyonii) and a fourth species (C. obliqua) that occurs as tetraploid and hexaploid races. To assess patterns of isozyme and morphological variation, and to test hypotheses of hybridization and allopolyploidy, we analyzed variation among 16 isozyme loci from 61 populations and 16 morphological characters from 33 populations representing all taxa and ploidy levels. Based on morphological analyses using clustering (unweighted pair group method using an arithmetic average) and ordination (principal components analysis and canonical variance analysis) methods, we recognize three diploid species without infraspecific taxa. Polyploids in the C. obliqua complex were most similar morphologically to diploid populations of C. glabra and C. lyonii. Patterns of isozyme variation among polyploids, which included fixed heterozygosity and recombinant profiles of alleles present in diploids, suggested polytopic origins of tetraploids and hexaploids. Our data indicate independent origins of polyploids in or near the southern Blue Ridge, Interior Highlands and Plains, and Atlantic Coastal Plain regions from progenitors most similar to C. glabra and C. lyonii. Extant tetraploids were not implicated in evolution of hexaploids, and plants similar to C. cuthbertii appeared unlikely as diploid progenitors for polyploids. We propose multiple differentiation and hybridization/polyploidization cycles in different geographic regions to explain the pattern of allopatry and inferred polytopic origins among polyploids. PMID- 10523289 TI - Membrane trafficking and the cytoskeleton: an integrated view. ASCB/EMBO/Dudley Wright Summer Research Conference, Santa Maria Imbaro, Italy, June 26-30, 1999. PMID- 10523290 TI - Crystal structure of gingipain R: an Arg-specific bacterial cysteine proteinase with a caspase-like fold. AB - Gingipains are cysteine proteinases acting as key virulence factors of the bacterium Porphyromonas gingivalis, the major pathogen in periodontal disease. The 1.5 and 2.0 A crystal structures of free and D-Phe-Phe-Arg-chloromethylketone inhibited gingipain R reveal a 435-residue, single-polypeptide chain organized into a catalytic and an immunoglobulin-like domain. The catalytic domain is subdivided into two subdomains comprising four- and six-stranded beta-sheets sandwiched by alpha-helices. Each subdomain bears topological similarities to the p20-p10 heterodimer of caspase-1. The second subdomain harbours the Cys-His catalytic diad and a nearby Glu arranged around the S1 specificity pocket, which carries an Asp residue to enforce preference for Arg-P1 residues. This gingipain R structure is an excellent template for the rational design of drugs with a potential to cure and prevent periodontitis. Here we show the binding mode of an arginine-containing inhibitor in the active-site, thus identifying major interaction sites defining a suitable pharmacophor. PMID- 10523291 TI - The structure of the 2A proteinase from a common cold virus: a proteinase responsible for the shut-off of host-cell protein synthesis. AB - The crystal structure of the 2A proteinase from human rhinovirus serotype 2 (HRV2 2A(pro)) has been solved to 1.95 A resolution. The structure has an unusual, although chymotrypsin-related, fold comprising a unique four-stranded beta sheet as the N-terminal domain and a six-stranded beta barrel as the C-terminal domain. A tightly bound zinc ion, essential for the stability of HRV2-2A(pro), is tetrahedrally coordinated by three cysteine sulfurs and one histidine nitrogen. The active site consists of a catalytic triad formed by His18, Asp35 and Cys106. Asp35 is additionally involved in an extensive hydrogen-bonding network. Modelling studies reveal a substrate-induced fit that explains the specificity of the subsites S4, S2, S1 and S1'. The structure of HRV2-2A(pro) suggests the mechanism of the cis cleavage and its release from the polyprotein. PMID- 10523292 TI - Receptor-mediated regulation of peroxisomal motility in CHO and endothelial cells. AB - The regulation of peroxisomal motility was investigated both in CHO cells and in cells derived from human umbilical vein endothelium (HUE). The cells were transfected with a construct encoding the green fluorescent protein bearing the C terminal peroxisomal targeting signal 1. Kinetic analysis following time-lapse imaging revealed that CHO cells respond to simultaneous stimulation with ATP and lysophosphatidic acid (LPA) by reducing peroxisomal movements. When Ca(2+) was omitted from the extracellular medium or the cells were incubated with inhibitors for heterotrimeric G(i)/G(o) proteins, phospholipase C, classical protein kinase C isoforms (cPKC), mitogen-activated protein kinase kinase (MEK) or phospholipase A(2) (PLA(2)), this signal-mediated motility block was abolished. HUE cells grown to confluency on microporous membranes responded similarly to ATP-LPA receptor co stimulation, but only when the ligands had access to the basolateral membrane region. These data demonstrate that peroxisomal motility is subject to specific modulation from the extracellular environment and suggest a receptor-mediated signaling cascade comprising Ca(2+) influx, G(i)/G(o) proteins, phospholipase C, cPKC isoforms, MEK and PLA(2) being involved in the regulation of peroxisomal arrest. PMID- 10523293 TI - Reaper-induced dissociation of a Scythe-sequestered cytochrome c-releasing activity. AB - Reaper is a potent apoptotic inducer critical for programmed cell death in the fly Drosophila melanogaster. While Reaper homologs from other species have not yet been reported, ectopic expression of Reaper in cells of vertebrate origin can also trigger apoptosis, suggesting that Reaper-responsive pathways are likely to be conserved. We recently reported that Reaper-induced mitochondrial cytochrome c release and caspase activation in a cell-free extract of Xenopus eggs requires the presence of a 150 kDa Reaper-binding protein, Scythe. We now show that Reaper binding to Scythe causes Scythe to release a sequestered apoptotic inducer. Upon release, the Scythe-sequestered factor(s) is sufficient to induce cytochrome c release from purified mitochondria. Moreover, addition of excess Scythe to egg extracts impedes Reaper-induced apoptosis, most likely through rebinding of the released factors. In addition to Reaper, Scythe binds two other Drosophila apoptotic regulators: Grim and Hid. Surprisingly, however, the region of Reaper which is detectably homologous to Grim and Hid is dispensable for Scythe binding. PMID- 10523294 TI - Dual targeting of cytochrome P4502B1 to endoplasmic reticulum and mitochondria involves a novel signal activation by cyclic AMP-dependent phosphorylation at ser128. AB - We have investigated mechanisms of mitochondrial targeting of the phenobarbital inducible hepatic mitochondrial P450MT4, which cross-reacts with antibody to microsomal P4502B1. Results show that P4502B1 and P450MT4 have identical primary sequence but different levels of phosphorylation and secondary structure. We demonstrate that P4502B1 contains a chimeric mitochondrial and endoplasmic reticulum (ER) targeting signal at its N-terminus. Inducers of cAMP and protein kinase A-mediated phosphorylation of P4502B1 at Ser128 activate the signal for mitochondrial targeting and modulate its mitochondrial or ER destination. S128A mutation inhibits in vitro mitochondrial transport and also in vivo mitochondrial targeting in COS cells. A fragment of P4502B1 containing the N-terminal signal and the phosphorylation site could drive the transport of dihydrofolate reductase (DHFR) into mitochondria. Ser128 phosphorylation reduced the affinity of 2B1 protein for binding to SRP, but increased the affinity of the 2B1-DHFR fusion protein for binding to yeast mitochondrial translocase proteins, TOM40 and TIM44, and matrix Hsp70. We describe a novel regulatory mechanism by which cAMP modulates the targeting of a protein to two distinct organelles. PMID- 10523295 TI - A rectifying ATP-regulated solute channel in the chloroplastic outer envelope from pea. AB - Phosphorylated carbohydrates are the main photoassimilated export products from chloroplasts that support the energy household and metabolism of the plant cell. Channels formed by the chloroplastic outer envelope protein OEP21 selectively facilitate the translocation of triosephosphate, 3-phosphoglycerate and phosphate, central intermediates in the source-sink relationship between the chloroplast and the cytosol. The anion selectivity and asymmetric transport properties of OEP21 are modulated by the ratio between ATP and triosephosphates, 3-phosphoglycerate and phosphate in the intermembrane space. Conditions that lead to export of triosephosphate from chloroplasts, i.e. photosynthesis, result in outward-rectifying OEP21 channels, while a high ATP to triosephosphate ratio, e.g. dark metabolism, leads to inward-rectifying OEP21 channels with a less pronounced anion selectivity. We conclude that solute exchange between plastids and cytosol can already be regulated at the level of the organellar outer membrane. PMID- 10523296 TI - Formation of anion-selective channels in the cell plasma membrane by the toxin VacA of Helicobacter pylori is required for its biological activity. AB - The vacuolating toxin VacA, a major determinant of Helicobacter pylori-associated gastric diseases, forms anion-selective channels in artificial planar lipid bilayers. Here we show that VacA increases the anion permeability of the HeLa cell plasma membrane and determines membrane depolarization. Electrophysiological and pharmacological approaches indicated that this effect is due to the formation of low-conductance VacA pores in the cell plasma membrane and not to the opening of Ca(2+)- or volume-activated chloride channels. VacA-dependent increase of current conduction both in artificial planar lipid bilayers and in the cellular system was effectively inhibited by the chloride channel blocker 5-nitro-2-(3 phenylpropylamino) benzoic acid (NPPB), while2-[(2-cyclopentenyl-6,7dichloro-2, 3 dihydro-2-methyl-1-oxo-1H-inden-5-yl)oxy]acetic acid (IAA-94) was less effective. NPPB inhibited and partially reversed the vacuolation of HeLa cells and the increase of ion conductivity of polarized Madine Darby canine kidney cell monolayers induced by VacA, while IAA-94 had a weaker effect. We conclude that pore formation by VacA accounts for plasma membrane permeabilization and is required for both cell vacuolation and increase of trans-epithelial conductivity. PMID- 10523297 TI - The polo-like protein kinases Fnk and Snk associate with a Ca(2+)- and integrin binding protein and are regulated dynamically with synaptic plasticity. AB - In order to stabilize changes in synaptic strength, neurons activate a program of gene expression that results in alterations of their molecular composition and structure. Here we demonstrate that Fnk and Snk, two members of the polo family of cell cycle associated kinases, are co-opted by the brain to serve in this program. Stimuli that produce synaptic plasticity, including those that evoke long-term potentiation (LTP), dramatically increase levels of both kinase mRNAs. Induced Fnk and Snk proteins are targeted to the dendrites of activated neurons, suggesting that they mediate phosphorylation of proteins in this compartment. Moreover, a conserved C-terminal domain in these kinases is shown to interact specifically with Cib, a Ca(2+)- and integrin-binding protein. Together, these studies suggest a novel signal transduction mechanism in the stabilization of long-term synaptic plasticity. PMID- 10523298 TI - Oncogenic potential of EAG K(+) channels. AB - We have investigated the possible implication of the cell cycle-regulated K(+) channel ether a go-go (EAG) in cell proliferation and transformation. We show that transfection of EAG into mammalian cells confers a transformed phenotype. In addition, human EAG mRNA is detected in several somatic cancer cell lines, despite being preferentially expressed in brain among normal tissues. Inhibition of EAG expression in several of these cancer cell lines causes a significant reduction of cell proliferation. Moreover, the expression of EAG favours tumour progression when transfected cells are injected into immune-depressed mice. These data provide evidence for the oncogenic potential of EAG. PMID- 10523300 TI - HLS7, a hemopoietic lineage switch gene homologous to the leukemia-inducing gene MLF1. AB - Hemopoietic lineage switching occurs when leukemic cells, apparently committed to one lineage, change and display the phenotype of another pathway. cDNA representational difference analysis was used to identify myeloid-specific genes that may be associated with an erythroid to myeloid lineage switch involving the murine J2E erythroleukemic cell line. One of the genes isolated (HLS7) is homologous to the novel human oncogene myeloid leukemia factor 1 (MLF1) involved in the t(3;5)(q25.1;q34) translocation associated with acute myeloid leukemia. Enforced expression of HLS7 in J2E cells induced a monoblastoid phenotype, thereby recapitulating the spontaneous erythroid to myeloid lineage switch. HLS7 also inhibited erythropoietin- or chemically-induced differentiation of erythroleukemic cell lines and suppressed development of erythropoietin responsive colonies in semi-solid culture. However, intracellular signaling activated by erythropoietin was not impeded by ectopic expression of HLS7. In contrast, HLS7 promoted maturation of M1 monoblastoid cells and increased myeloid colony formation in vitro. These data show that HLS7 can influence erythroid/myeloid lineage switching and the development of normal hemopoietic cells. PMID- 10523299 TI - Blue light activates the plasma membrane H(+)-ATPase by phosphorylation of the C terminus in stomatal guard cells. AB - The opening of stomata, which is driven by the accumulation of K(+) salt in guard cells, is induced by blue light (BL). The BL activates the H(+) pump; however, the mechanism by which the perception of BL is transduced into the pump activation remains unknown. We present evidence that the pump is the plasma membrane H(+)-ATPase and that BL activates the H(+)-ATPase via phosphorylation. A pulse of BL (30 s, 100 micromol/m(2)/s) increased ATP hydrolysis by the plasma membrane H(+)-ATPase and H(+) pumping in Vicia guard cell protoplasts with a similar time course. The H(+)-ATPase was phosphorylated reversibly by BL, and the phosphorylation levels paralleled the ATP hydrolytic activity. The phosphorylation occurred exclusively in the C-termini of H(+)-ATPases on both serine and threonine residues in two isoproteins of H(+)-ATPase in guard cells. An endogenous 14-3-3 protein was co-precipitated with H(+)-ATPase, and the recombinant 14-3-3 protein bound to the phosphorylated C-termini of H(+)-ATPases. These findings demonstrate that BL activates the plasma membrane H(+)-ATPase via phosphorylation of the C-terminus by a serine/threonine protein kinase, and that the 14-3-3 protein has a key role in the activation. PMID- 10523301 TI - Cell-type specific phosphorylation of threonines T654 and T669 by PKD defines the signal capacity of the EGF receptor. AB - In Rat-1 fibroblasts epidermal growth factor (EGF), but not platelet-derived growth factor (PDGF) stimulates the activity of the c-Jun N-terminal kinase (JNK). Moreover, PDGF induced suppression of EGF-mediated JNK activation, apparently through protein kinase C (PKC) activation. Further analysis revealed that PKD was specifically activated by PDGF but not EGF in Rat-1 cells. In SF126 glioblastoma cells, however, EGF and PDGF synergistically activated JNK, while neither PDGF nor EGF stimulated PKD activity. In this cell line, overexpression of PKD blocked EGF- and PDGF-induced JNK activation. Mutational analysis further revealed that the EGFR mutant (T654/669E) was incapable of activating JNK and provided evidence that PKD-mediated dual phosphorylation of these critical threonine residues leads to suppression of EGF-induced JNK activation. Our results establish a novel crosstalk mechanism which allows signal integration and definition in cells with many different RTKs. PMID- 10523302 TI - A novel regulator of G protein signalling in yeast, Rgs2, downregulates glucose activation of the cAMP pathway through direct inhibition of Gpa2. AB - We have characterized a novel member of the recently identified family of regulators of heterotrimeric G protein signalling (RGS) in the yeast Saccharomyces cerevisiae. The YOR107w/RGS2 gene was isolated as a multi-copy suppressor of glucose-induced loss of heat resistance in stationary phase cells. The N-terminal half of the Rgs2 protein consists of a typical RGS domain. Deletion and overexpression of Rgs2, respectively, enhances and reduces glucose induced accumulation of cAMP. Overexpression of RGS2 generates phenotypes consistent with low activity of cAMP-dependent protein kinase A (PKA), such as enhanced accumulation of trehalose and glycogen, enhanced heat resistance and elevated expression of STRE-controlled genes. Deletion of RGS2 causes opposite phenotypes. We demonstrate that Rgs2 functions as a negative regulator of glucose induced cAMP signalling through direct GTPase activation of the Gs-alpha protein Gpa2. Rgs2 and Gpa2 constitute the second cognate RGS-G-alpha protein pair identified in yeast, in addition to the mating pheromone pathway regulators Sst2 and Gpa1. Moreover, Rgs2 and Sst2 exert specific, non-overlapping functions, and deletion mutants in Rgs2 and Sst2 are complemented to some extent by different mammalian RGS proteins. PMID- 10523303 TI - The Aspergillus nidulans sfaD gene encodes a G protein beta subunit that is required for normal growth and repression of sporulation. AB - flbA encodes an Aspergillus nidulans RGS (regulator of G protein signaling) domain protein that antagonizes FadA (G(i)alpha-subunit of heterotrimeric G protein)-mediated growth signaling to allow asexual development. We previously defined and characterized five suppressors of flbA (sfa) loss-of-function mutations and showed that one suppressor (sfaB) resulted from a novel dominant negative allele of fadA. In this report we show that a second suppressor gene (sfaD) is predicted to encode the beta subunit of a heterotrimeric G protein. Deletion of sfaD suppressed all defects resulting from complete loss-of-flbA function mutations, caused a hyperactive sporulation phenotype and severely reduced vegetative growth. However, the sfaD deletion could not suppress the growth activation caused by dominant-activating fadA alleles, indicating that constitutively active FadA can cause proliferative growth in the absence of Gbetagamma signaling. We propose that SfaD and FadA are both positive growth regulators with partially overlapping functions and that FlbA has an important role in controlling the activities of both proteins. Inactivation of signaling events stimulated by both components of the heterotrimeric G protein is essential for both sexual and asexual sporulation. PMID- 10523305 TI - Ref-1 regulates the transactivation and pro-apoptotic functions of p53 in vivo. AB - Ref-1 is a multifunctional protein that stimulates DNA binding by a number of transcription factors and serves as the abasic (A/P) endonuclease in base excision repair. Ref-1 was discovered to be a potent activator of p53 DNA binding in vitro. To address the physiological significance of the effects of Ref-1 on p53, we have analyzed its role in regulating p53 function in vivo. We found that Ref-1 over-expression enhances the ability of p53 to transactivate a number of p53 target promoters and increases the ability of p53 to stimulate endogenous p21 and cyclin G expression. Additionally, it was observed that Ref-1 associates with p53 in vivo and in vitro. Importantly, downregulation of Ref-1 (by antisense) causes a marked reduction in p53 induction of p21 mRNA and protein, as well as diminished ability of p53 to transactivate the p21 and Bax promoters. Moreover, Ref-1 levels are correlated with the extent of apoptosis induced by p53. Finally, we observed that Ref-1 cooperates with a DNA-damaging compound, camptothecin, to stimulate the transcriptional activity of p53. Together these data indicate that Ref-1 is a key cellular regulator of p53. PMID- 10523304 TI - p38 MAP kinase is required for STAT1 serine phosphorylation and transcriptional activation induced by interferons. AB - Activation of cytosolic phospholipase A(2 )(cPLA(2)) is a prerequisite for the formation of the transcription factor complex interferon-stimulated gene factor 3 (ISGF3) in response to interferon-alpha (IFN-alpha). Here we show that p38 mitogen-activated protein kinase (MAPK), an activator of cPLA(2), is essential for both IFN-alpha and IFN-gamma signalling. SB203580, a specific inhibitor of p38, was found to inhibit ISGF3 formation but had no apparent effects on signal transducer and activator of transcription (STAT)1 homodimer formation. Regardless of this, the antiviral activities of both IFN-alpha and IFN-gamma were attenuated by SB203580. Treatment with either IFN led to rapid and transient activation of p38. Both IFNs induced STAT1 Ser727 phosphorylation, which was inhibited by SB203580 but not by an extracellular signal related kinase (ERK)1/2 inhibitor (PD98059). In an inducible 3T3-L1 clone, expression of dominant-negative p38 led to defective STAT1 serine phosphorylation and diminished IFN-gamma-mediated protection against viral killing. Reporter activity mediated by ISGF3 or STAT1 homodimer was diminished by SB203580 and enhanced by a constitutively active mutant of MKK6, the upstream activator of p38. Therefore, p38 plays a key role in the serine phosphorylation of STAT1 and transcriptional changes induced by both IFNs. PMID- 10523306 TI - Functional interaction between GCN5 and polyamines: a new role for core histone acetylation. AB - Polyamines are organic polycations essential for a wide variety of cellular functions, including nuclear integrity and chromosome condensation. Here we present genetic evidence that depletion of cellular polyamines partially alleviates the defects in HO and SUC2 expression caused by inactivation of the GCN5 histone acetyltransferase. In addition, the combination of polyamine depletion and a sin(-) allele of the histone H4 gene leads to almost complete bypass of the transcriptional requirement for GCN5. In contrast, polyamine depletion does not alter the transcriptional requirements for the SWI/SNF chromatin remodeling complex nor does depletion lead to global defects in transcriptional regulation. In addition to these genetic studies, we show that polyamines facilitate oligomerization of nucleosomal arrays in vitro, and that polyamine-mediated condensation requires intact core histone N-terminal domains and is inhibited by histone hyperacetylation. Our studies suggest that polyamines are repressors of transcription in vivo, and that one role of histone hyperacetylation is to antagonize the ability of polyamines to stabilize highly condensed states of chromosomal fibers. PMID- 10523308 TI - RNA elements required for RNA recombination function as replication enhancers in vitro and in vivo in a plus-strand RNA virus. AB - RNA replication requires cis-acting elements to recruit the viral RNA-dependent RNA polymerase (RdRp) and facilitate de novo initiation of complementary strand synthesis. Hairpins that are hot spots for recombination in the genomic RNA of turnip crinkle virus (TCV) and satellite (sat)-RNA C, a parasitic RNA associated with TCV infections, stimulate RNA synthesis 10-fold from a downstream promoter sequence in an in vitro assay using partially purified TCV RdRp. Artificial hairpins had an inhibitory effect on transcription. RNA accumulation in single cells was enhanced 5- to 10-fold when the natural stem-loop structures were inserted into a poorly accumulating sat-RNA. The effect of the stem-loop structures on RNA replication was additive, with insertion of three stem-loop RNA elements increasing sat-RNA accumulation to the greatest extent (25-fold). These stem-loop structures do not influence the stability of the RNAs in vivo, but may serve to recruit the RdRp to the template. PMID- 10523307 TI - p300 stimulates transcription instigated by ligand-bound thyroid hormone receptor at a step subsequent to chromatin disruption. AB - We investigate the role of the transcriptional coactivator p300 in gene activation by thyroid hormone receptor (TR) on addition of ligand. The ligand bound TR targets chromatin disruption independently of gene activation. Exogenous p300 facilitates transcription from a disrupted chromatin template, but does not itself disrupt chromatin in the presence or absence of ligand-bound receptor. Nevertheless, the acetyltransferase activity of p300 is required to facilitate transcription from a disrupted chromatin template. Expression of E1A prevents aspects of chromatin remodeling and transcriptional activation dependent on TR and p300. E1A selectively inhibits the acetylation of non-histone substrates. E1A does not prevent the assembly of a DNase I-hypersensitive site induced by TR, but does inhibit topological alterations and the loss of canonical nucleosome arrays dependent on the addition of ligand. Mutants of E1A incompetent for interaction with p300 partially inhibit chromatin disruption but still allow nuclear receptors to activate transcription. We conclude that p300 has no essential role in chromatin disruption, but makes use of acetyltransferase activity to stimulate transcription at a subsequent step. PMID- 10523310 TI - Resolution of head-on collisions between the transcription machinery and bacteriophage phi29 DNA polymerase is dependent on RNA polymerase translocation. AB - The outcome of collisions between Bacillus subtilis phage Phi29 DNA polymerase and oppositely oriented transcription complexes has been studied in vitro. We found that the replication fork was unable to go past a transcription ternary complex stalled head-on. However, head-on collisions did not lead to a deadlock. Both DNA and RNA polymerase remained bound to the template and, when the halted transcription complex was allowed to move, the replication machinery resumed normal elongation. These results suggested that a replication fork that encounters an RNA polymerase head-on whose movement is not impeded would bypass the transcription machinery. Our results for head-on collisions between concurrently moving replication and transcription complexes are indeed consistent with the existence of a resolving mechanism. The ability of Phi29 DNA polymerase to resolve head-on collisions with itself during symmetrical replication of Phi29 DNA in vivo is likely to be related to its ability to pass a head-on oriented RNA polymerase. PMID- 10523309 TI - The mechanism of phosphorylation-inducible activation of the ETS-domain transcription factor Elk-1. AB - Protein phosphorylation represents one of the major mechanisms for transcription factor activation. Here we demonstrate a molecular mechanism by which phosphorylation by mitogen-activated protein (MAP) kinases leads to changes in transcription factor activity. MAP kinases stimulate DNA binding and transcriptional activation mediated by the mammalian ETS-domain transcription factor Elk-1. Phosphorylation of the C-terminal transcriptional activation domain induces a conformational change in Elk-1, which accompanies the stimulation of DNA binding. C-terminal phosphorylation is coupled to activation of DNA binding by the N-terminal DNA-binding domain via an additional intermediary domain. Activation of DNA binding is mediated by an allosteric mechanism involving the key phosphoacceptor residues. Together, these results provide a molecular model for how phosphorylation induces changes in Elk-1 activity. PMID- 10523311 TI - Organization of DNA replication origins in the fission yeast genome. AB - Eukaryotic DNA replication initiates at multiple points along the chromosomes known as replication origins (ORIs). We have developed a strategy to identify ORIs directly from replication intermediates in the fission yeast Schizosaccharomyces pombe. Mapping of a selection of the novel ORIs onto the genome reveals their preferential localization at intergenic regions upstream from genes. These results are supported by the observation that a large proportion of regions overlapping gene promoters contain active ORIs. Mapping of the genomic ars1 replication origin at nucleotide resolution shows that replication initiates at a defined position immediately upstream from the hus5(+) promoter. Deletion analysis indicates that the regulatory elements required to initiate transcription and replication lie in close proximity, suggesting a possible relationship between both processes in vivo. PMID- 10523312 TI - Mammalian homologues of the plant Tousled gene code for cell-cycle-regulated kinases with maximal activities linked to ongoing DNA replication. AB - The Tousled (TSL) gene of the plant Arabidopsis thaliana encodes a serine/threonine kinase that is essential for proper flower development. Here we report the cloning and characterization of two human putative homologues of the Arabidopsis TSL gene, termed TLK1 and TLK2 (Tousled-like kinase). At the protein level, the two human Tlks share 84% sequence similarity with each other and almost 50% with Arabidopsis Tsl. Furthermore, nuclear localization signals and predicted coiled-coil regions are conserved in the N-terminal domains of all three kinases. The mammalian Tlks share several functional properties with plant Tsl, including a broad expression, a propensity to dimerize and autophosphorylate, and a preference for similar substrates. Most interestingly, human Tlks are cell-cycle-regulated enzymes, displaying maximal activities during S phase. Whereas protein levels are virtually constant throughout the cell cycle, both Tlks appear to be regulated by cell-cycle-dependent phosphorylation. Drug induced inhibition of DNA replication causes a rapid loss of Tlk activity, indicating that Tlk function is tightly linked to ongoing DNA replication. These findings provide the first biochemical clues as to the possible molecular functions of Tlks, a highly conserved family of kinases implicated in the development of multicellular organisms. PMID- 10523313 TI - Mammalian Cdc7-Dbf4 protein kinase complex is essential for initiation of DNA replication. AB - The Cdc7-Dbf4 kinase is essential for regulating initiation of DNA replication in Saccharomyces cerevisiae. Previously, we identified a human Cdc7 homolog, HsCdc7. In this study, we report the identification of a human Dbf4 homolog, HsDbf4. We show that HsDbf4 binds to HsCdc7 and activates HsCdc7 kinase activity when HsDbf4 and HsCdc7 are coexpressed in insect and mammalian cells. HsDbf4 protein levels are regulated during the cell cycle with a pattern that matches that of HsCdc7 protein kinase activity. They are low in G(1), increase during G(1)-S, and remain high during S and G(2)-M. Purified baculovirus-expressed HsCdc7-HsDbf4 selectively phosphorylates the MCM2 subunit of the minichromosome maintenance (MCM) protein complex isolated by immunoprecipitation with MCM7 antibodies in vitro. Two-dimensional tryptic phosphopeptide-mapping analysis of in vivo (32)P labeled MCM2 from HeLa cells reveals that several major tryptic phosphopeptides of MCM2 comigrate with those of MCM2 phosphorylated by HsCdc7-HsDbf4 in vitro, suggesting that MCM2 is a physiological HsCdc7-HsDbf4 substrate. Immunoneutralization of HsCdc7-HsDbf4 activity by microinjection of anti-HsCdc7 antibodies into HeLa cells blocks initiation of DNA replication. These results indicate that the HsCdc7-HsDbf4 kinase is directly involved in regulating the initiation of DNA replication by targeting MCM2 protein in mammalian cells. PMID- 10523314 TI - The Saccharomyces cerevisiae MER3 gene, encoding a novel helicase-like protein, is required for crossover control in meiosis. AB - The MER3 gene is identified as a novel meiosis-specific gene, whose transcript is spliced in an MRE2/MER1-dependent manner. The predicted Mer3 protein contains the seven motifs characteristic of the DExH-box type of helicases as well as a putative zinc finger. Double strand breaks (DSBs), the initial changes of DNA in meiotic recombination, do not disappear completely and are hyperresected late in mer3 meiosis, indicating that MER3 is required for the transition of DSBs to later intermediates. A mer3 mutation reduces crossover frequencies, and the remaining crossovers show random distribution along a chromosome, resulting in a high incidence of non-disjunction of homologous chromosomes at the first meiotic division. MER3 appears to be very important for both the DSB transition and crossover control. PMID- 10523315 TI - FtsK-dependent and -independent pathways of Xer site-specific recombination. AB - Homologous recombination between circular chromosomes generates dimers that cannot be segregated at cell division. Escherichia coli Xer site-specific recombination converts chromosomal and plasmid dimers to monomers. Two recombinases, XerC and XerD, act at the E. coli chromosomal recombination site, dif, and at related sites in plasmids. We demonstrate that Xer recombination at plasmid dif sites occurs efficiently only when FtsK is present and under conditions that allow chromosomal dimer formation, whereas recombination at the plasmid sites cer and psi is independent of these factors. We propose that the chromosome dimer- and FtsK-dependent process that activates Xer recombination at plasmid dif also activates Xer recombination at chromosomal dif. The defects in chromosome segregation that result from mutation of the FtsK C-terminus are attributable to the failure of Xer recombination to resolve chromosome dimers to monomers. Conditions that lead to FtsK-independent Xer recombination support the hypothesis that FtsK acts on Holliday junction Xer recombination intermediates. PMID- 10523316 TI - TRF1 binds a bipartite telomeric site with extreme spatial flexibility. AB - TRF1 is a key player in telomere length regulation. Because length control was proposed to depend on the architecture of telomeres, we studied how TRF1 binds telomeric TTAGGG repeat DNA and alters its conformation. Although the single Myb type helix-turn-helix motif of a TRF1 monomer can interact with telomeric DNA, TRF1 predominantly binds as a homodimer. Systematic Evolution of Ligands by Exponential enrichment (SELEX) with dimeric TRF1 revealed a bipartite telomeric recognition site with extreme spatial variability. Optimal sites have two copies of a 5'-YTAGGGTTR-3' half-site positioned without constraint on distance or orientation. Analysis of binding affinities and DNase I footprinting showed that both half-sites are simultaneously contacted by the TRF1 dimer, and electron microscopy revealed looping of the intervening DNA. We propose that a flexible segment in TRF1 allows the two Myb domains of the homodimer to interact independently with variably positioned half-sites. This unusual DNA binding mode is directly relevant to the proposed architectural role of TRF1. PMID- 10523317 TI - The replication factory targeting sequence/PCNA-binding site is required in G(1) to control the phosphorylation status of DNA ligase I. AB - The recruitment of DNA ligase I to replication foci in S phase depends on a replication factory targeting sequence that also mediates the interaction with proliferating cell nuclear antigen (PCNA) in vitro. By exploiting a monoclonal antibody directed at a phospho-epitope, we demonstrate that Ser66 of DNA ligase I, which is part of a strong CKII consensus site, is phosphorylated in a cell cycle-dependent manner. After dephosphorylation in early G(1), the level of Ser66 phosphorylation is minimal in G(1), increases progressively in S and peaks in G(2)/M phase. The analysis of epitope-tagged DNA ligase I mutants demonstrates that dephosphorylation of Ser66 requires both the nuclear localization and the PCNA-binding site of the enzyme. Finally, we show that DNA ligase I and PCNA interact in vivo in G(1) and S phase but not in G(2)/M. We propose that dephosphorylation of Ser66 is part of a novel control mechanism to establish the pre-replicative form of DNA ligase I. PMID- 10523318 TI - Enhancement of hepatitis C virus replication by Epstein-Barr virus-encoded nuclear antigen 1. AB - Based on our recent observation that Epstein-Barr virus (EBV) is detected in 37% of the tissues of hepatocellular carcinoma, and especially frequently in cases with hepatitis C virus (HCV), the effect of EBV infection on the replication of HCV was investigated. EBV-infected cell clones and their EBV-uninfected counterparts in cell lines MT-2 (a human T-lymphotropic virus type I-infected T cell line), HepG2 (a hepatoblastoma cell line) and Akata (a Burkitt's lymphoma cell line) were compared in terms of their permissiveness for HCV replication following inoculation of HCV derived from patients who were HCV carriers. The results indicated that EBV-infected cell clones, but not their EBV-uninfected counterparts, promoted HCV replication. EBV-encoded nuclear antigen 1 (EBNA1), which is invariably expressed in EBV-infected cells, supported HCV replication. Deletion analysis of the EBNA1 gene showed good correlation between transactivation activity and the activity supporting HCV replication. The present findings suggest that EBV acts as a helper virus for HCV replication. PMID- 10523319 TI - Rat8p/Dbp5p is a shuttling transport factor that interacts with Rat7p/Nup159p and Gle1p and suppresses the mRNA export defect of xpo1-1 cells. AB - In a screen for temperature-sensitive mutants of Saccharomyces cerevisiae defective for mRNA export, we previously identified the essential DEAD-box protein Dbp5p/Rat8p and the nucleoporin Rat7p/Nup159p. Both are essential for mRNA export. Here we report that Dbp5p and Rat7p interact through their Nterminal domains. Deletion of this portion of Rat7p (Rat7pDeltaN) results in strong defects in mRNA export and eliminates association of Dbp5p with nuclear pores. Overexpression of Dbp5p completely suppressed the growth and mRNA export defects of rat7DeltaN cells and resulted in weaker suppression in cells carrying rat7-1 or the rss1-37 allele of GLE1. Dbp5p interacts with Gle1p independently of the N terminus of Dbp5p. Dbp5p shuttles between nucleus and cytoplasm in an Xpo1p dependent manner. It accumulates in nuclei of xpo1-1 cells and in cells with mutations affecting Mex67p (mex67-5), Gsp1p (Ran) or Ran effectors. Overexpression of Dbp5p prevents nuclear accumulation of mRNA in xpo1-1 cells, but does not restore growth, suggesting that the RNA export defect of xpo1-1 cells may be indirect. In a screen for high-copy suppressors of the rat8-2 allele of DBP5, we identified YMR255w, now called GFD1. Gfd1p is not essential, interacts with Gle1p and Rip1p/Nup42p, and is found in the cytoplasm and at the nuclear rim. PMID- 10523321 TI - Another nobel laureate honored by the council for high blood pressure research PMID- 10523320 TI - A doughnut-shaped heteromer of human Sm-like proteins binds to the 3'-end of U6 snRNA, thereby facilitating U4/U6 duplex formation in vitro. AB - We describe the isolation and molecular characterization of seven distinct proteins present in human [U4/U6.U5] tri-snRNPs. These proteins exhibit clear homology to the Sm proteins and are thus denoted LSm (like Sm) proteins. Purified LSm proteins form a heteromer that is stable even in the absence of RNA and exhibits a doughnut shape under the electron microscope, with striking similarity to the Sm core RNP structure. The purified LSm heteromer binds specifically to U6 snRNA, requiring the 3'-terminal U-tract for complex formation. The 3'-end of U6 snRNA was also co-precipitated with LSm proteins after digestion of isolated tri snRNPs with RNaseT(1). Importantly, the LSm proteins did not bind to the U-rich Sm sites of intact U1, U2, U4 or U5 snRNAs, indicating that they can only interact with a 3'-terminal U-tract. Finally, we show that the LSm proteins facilitate the formation of U4/U6 RNA duplices in vitro, suggesting that the LSm proteins may play a role in U4/U6 snRNP formation. PMID- 10523322 TI - The nature of intracrine peptide hormone action. AB - Current theory holds that peptide hormone action results from hormone binding to cell-surface receptors, with the generation of intracellular second messengers. However, a growing body of evidence suggests that intracellular peptide hormone, either internalized or synthesized in situ, can exert physiologically relevant effects. These effects are diverse and poorly understood. I propose that such intracrine action can serve to modulate cellular function over time and thereby play a role in biological memory of various sorts, in the maintenance of hormonal responsiveness, and in cellular differentiation. PMID- 10523323 TI - Endothelial function in hypertension: the role of superoxide anion. AB - Much attention has been focused on the role of nitric oxide in hypertension and cardiovascular disease. More recently, the role of superoxide anion and its interaction with nitric oxide has been investigated in this context. This review will concentrate on the role of superoxide in human and experimental hypertension, paying particular attention to the potential sources of superoxide within the vasculature and discussing some of the molecular mechanisms surrounding its production and dismutation. We discuss what is known about the human superoxide dismutase enzymes. We conclude that the balance between nitric oxide and superoxide is more important than the absolute levels of either alone. PMID- 10523324 TI - Pathogenic role of oxidative stress in vascular angiotensin-converting enzyme activation in long-term blockade of nitric oxide synthesis in rats. AB - Inhibition of nitric oxide (NO) synthesis with N(omega)-nitro-L-arginine methyl ester (L-NAME) activates vascular angiotensin-converting enzyme (ACE) and causes oxidative stress. We investigated the role of oxidative stress in the pathogenesis of ACE activation in rats. Studies involved aortas of rats receiving no treatment, L-NAME, L-NAME plus L-arginine, or L-NAME plus an antioxidant drug (N-acetylcysteine, allopurinol, or ebselen) for 7 days. L-NAME significantly increased oxidative stress (O(2)(-)) and ACE activity. The increased O(2)(-) production was normalized by removal of endothelium. Immunohistochemistry showed the increased ACE activity in the endothelial layer. Treatment with antioxidant drugs did not affect the L-NAME-induced increase in systolic arterial pressure but did prevent increases in vascular O(2)(-) production and ACE activity. These results implicate oxidative stress in the pathogenesis of vascular ACE activation in rats with long-term inhibition of NO synthesis. The observed effects of antioxidant drugs on ACE activation do not appear to involve the hypertension induced by L-NAME. PMID- 10523325 TI - Nitric oxide synthase isotype expression in salt-sensitive and salt-resistant Dahl rats. AB - Previous studies have suggested that salt-sensitive hypertension in humans and experimental animals may in part be due to dysregulation of the L-arginine/nitric oxide system. This study was conducted to determine the endothelial, inducible, and neuronal nitric oxide synthase expressions in the kidney, heart, aorta, and brain of salt-sensitive and salt-resistant Dahl rats. We studied salt-sensitive and salt-resistant Dahl rats maintained on high- (8%) and regular- (0.2%) salt diets for 3 weeks. Blood pressure was modestly elevated in both Dahl salt sensitive and salt-resistant rats consuming regular diet and severely increased in sensitive but not resistant rats consuming the high-salt diet. The Dahl salt sensitive animals showed a significant reduction in kidney, heart, and aorta inducible nitric oxide synthase protein abundance on the regular diet, with further reductions on the high-salt diet. In addition, the high-salt diet markedly downregulated endothelial nitric oxide synthase expression in the kidney and aorta but not in the heart of the Dahl salt-sensitive animals. The rise in blood pressure in the Dahl salt-sensitive rats on the high-salt diet was accompanied by a significant elevation of brain neuronal nitric oxide synthase protein. In contrast, salt-resistant animals showed no change in heart, kidney, and aorta endothelial or brain neuronal nitric oxide synthase and considerably less intense changes in inducible isotype than that seen in the salt-sensitive group in response to the high-salt diet. In conclusion, the study revealed a marked downregulation of inducible nitric oxide synthase in the Dahl salt sensitive rats on the regular diet, with further reductions on the high-salt diet. Furthermore, Dahl salt-sensitive rats consuming the high-salt diet showed significant reductions of kidney and aorta endothelial nitric oxide synthase and an upregulation of brain neuronal nitric oxide synthase expression. PMID- 10523326 TI - Nitric oxide synthase expression in the course of lead-induced hypertension. AB - We recently showed elevated reactive oxygen species (ROS), reduced urinary excretion of NO metabolites (NOx), and increased NO sequestration as nitrotyrosine in various tissues in rats with lead-induced hypertension. This study was designed to discern whether the reduction in urinary NOx in lead induced hypertension is, in part, due to depressed NO synthase (NOS) expression. Male Sprague-Dawley rats were randomly assigned to a lead-treated group (given lead acetate, 100 ppm, in drinking water and regular rat chow), a group given lead and vitamin E-fortified chow, or a normal control group given either regular food and water or vitamin E-fortified food for 12 weeks. Tail blood pressure, urinary NOx excretion, plasma malondialdehyde (MDA), and endothelial and inducible NOS (eNOS and iNOS) isotypes in the aorta and kidney were measured. The lead-treated group exhibited a rise in blood pressure and plasma MDA concentration, a fall in urinary NOx excretion, and a paradoxical rise in vascular and renal tissue eNOS and iNOS expression. Vitamin E supplementation ameliorated hypertension, lowered plasma MDA concentration, and raised urinary NOx excretion while significantly lowering vascular, but not renal, tissue eNOS and iNOS expression. Vitamin E supplementation had no effect on either blood pressure, plasma MDA, or NOS expression in the control group. The study also revealed significant inhibition of NOS enzymatic activity by lead in cell-free preparations. In conclusion, lead-induced hypertension in this model was associated with a compensatory upregulation of renal and vascular eNOS and iNOS expression. This is, in part, due to ROS-mediated NO inactivation, lead associated inhibition of NOS activity, and perhaps stimulatory actions of increased shear stress associated with hypertension. PMID- 10523327 TI - Role of nitric oxide in the control of cardiac oxygen consumption in B(2)-kinin receptor knockout mice. AB - The aim of this study was to determine whether bradykinin, the angiotensin converting enzyme inhibitor ramiprilat, and the calcium-channel antagonist amlodipine reduce myocardial oxygen consumption (MV(O2)) via a B(2)-kinin receptor/nitric oxide-dependent mechanism. Left ventricular free wall and septum were isolated from normal and B(2)-kinin receptor knockout (B(2) -/-) mice. Myocardial tissue oxygen consumption was measured in an airtight chamber with a Clark-type oxygen electrode. Baseline MV(O2) was not significantly different between normal (239+/-13 nmol of O(2). min(-1). g(-1)) and B(2) -/- (263+/-24 nmol of O(2). min(-1). g(-1)) mice. S-nitroso-N-acetyl-penicillamine (10(-7) to 10(-4) mol/L) reduced oxygen consumption in a concentration-dependent manner in both normal (maximum, 36+/-3%) and B(2) -/- mice (28+/-3%). This was also true for the endothelium-dependent vasodilator substance P (10(-10) to 10(-7) mol/L; 22+/-7% in normal mice and 20+/-4% in B(2) -/- mice). Bradykinin (10(-7) to 10( 4) mol/L), ramiprilat (10(-7) to 10(-4) mol/L), and amlodipine (10(-7) to 10(-5) mol/L) all caused concentration-dependent decreases in MV(O2)in normal mice. At the highest concentration, tissue O(2) consumption was decreased by 18+/-3%, 20+/ 5%, and 28+/-3%, respectively. The reduction in MV(O2) to all 3 drugs was attenuated in the presence of N(G)-nitro-L-arginine-methyl ester. However, in the B(2) -/- mice, bradykinin, ramiprilat, and amlodipine had virtually no effect on MV(O2). Therefore, nitric oxide, through a bradykinin-receptor-dependent mechanism, regulates cardiac oxygen consumption. This physiological mechanism is absent in B(2) -/- mice and may be evidence of an important therapeutic mechanism of action of angiotensin-converting enzyme inhibitors and amlodipine. PMID- 10523328 TI - Early upregulation of endothelial adhesion molecules in obese hypertensive men. AB - Upregulation of endothelial adhesion molecules is the earliest step of atherogenesis. Whether obesity induces endothelial adhesin upregulation is unknown. To address this topic, circulating vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), E-selectin, and von Willebrand factor (vWF) concentrations were evaluated in 22 obese hypertensive (51.4+/-4.6 years [mean+/-SD age]), 19 obese normotensive (50.6+/-3.8 years), 18 nonobese hypertensive (52.3+/-3.9 years), and 16 nonobese normotensive (52. 4+/ 3.5 years) men without other risk factors or overt atherosclerosis. All measurements were repeated in the obese subgroups after weight loss induced by 12 weeks of caloric restriction. Basal circulating VCAM-1 levels were similar between the 2 obese groups but were higher (P<0.0001) than in the 2 nonobese groups. No differences were found between nonobese hypertensives and normotensives. Serum low density lipoprotein cholesterol was weakly correlated with plasma soluble VCAM-1 levels in pooled, obese subjects (r=0.362, P=0.02). Plasma soluble adhesin and vWF concentrations decreased significantly after weight loss in obese hypertensives (VCAM-1 P=0.03, ICAM-1 P=0.004, E-selectin P<0.0001, and vWF P=0.003) and normotensives (VCAM-1 P=0.04, ICAM-1 P=0.003, E selectin P<0.0001, and vWF P<0.0001). Body mass index was correlated with plasma E-selectin concentrations at baseline and after weight loss in obese hypertensives (r=0.501, P=0.018 and r=0. 466, P=0.03, respectively) and obese normotensives (r=0.523, P=0.021 and r=0.460, P=0.05, respectively). In conclusion, our data show that obesity per se induces early endothelial activation in hypertensive and normotensive men. Weight loss counteracted endothelial activation in both obese hypertensive and normotensive men. PMID- 10523329 TI - N-acetyl-L-cysteine enhances interleukin-1beta-induced nitric oxide synthase expression. AB - The effect of N-acetyl-L-cysteine on interleukin-1beta-induced nitric oxide synthase expression was studied in rat vascular smooth muscle cells to determine if the reduction/oxidation state would modulate cytokine-induced changes. Interleukin-1beta induced the production of nitrite, a stable metabolite of nitric oxide in a time- and dose-dependent manner. Cytokine-induced nitrite production was enhanced by the addition of N-acetyl-L-cysteine in a dose dependent manner, with a >50% increase produced by the addition of 1 mmol/L N acetyl-L-cysteine. There was no influence on nitrite production when the cells were treated with N-acetyl-L-cysteine alone. Northern and Western blot analyses revealed that the upregulation of interleukin-1beta-induced nitric oxide production by N-acetyl-L-cysteine resulted from an enhanced expression of inducible nitric oxide synthase. Interferon-gamma or tumor necrosis factor-alpha when used alone had no influence on nitrite production in the absence or presence of N-acetyl-L-cysteine. Nitrite accumulation was higher by the cells treated with interleukin-1beta combined with either interferon-gamma or tumor necrosis factor alpha compared with those treated with interleukin-1beta alone. N-Acetyl-L cysteine upregulated nitrite production and inducible nitric oxide synthase expression induced by combination treatment with interleukin-1beta and either interferon-gamma or tumor necrosis factor-alpha. However, N-acetyl-L-cysteine had no significant influence in cytokine-induced activation of nuclear factor-kappaB or signal transducer and activator of transciption-1, as assessed by electrophoretic mobility shift assays. These results demonstrate that N-acetyl-L cysteine possibly acted as a thiol-containing reducing agent and facilitated the expression of inducible nitric oxide synthase in rat vascular smooth muscle cells by cytokines through a mechanism that is independent of nuclear factor-kappaB or signal transducer and activator of transciption-1. PMID- 10523330 TI - Hypotensive effect of low-fat, high-carbohydrate diet can be independent of changes in plasma insulin concentrations. AB - To examine the relationship between diet, blood pressure, and plasma insulin concentrations, we studied 14 healthy males who were prescribed low-fat and high fat diets. The low-fat diet contained 25% (of energy intake) fat and 54% carbohydrate; the high-fat diet was 45% fat (predominantly saturated fat) and 36% carbohydrate. The diets were consumed over consecutive 2-week periods in random sequence, separated by a 2-week washout period. Resting supine systolic and diastolic blood pressures decreased significantly by 7 and 3 mm Hg, respectively, and plasma total cholesterol, LDL cholesterol, and HDL cholesterol concentrations all fell (by 21.6%, 25.7%, and 18.0%, respectively; all P<0.001) on the low-fat compared with the high-fat diet. Fasting glucose and the glucose area under the curve during the frequently sampled intravenous glucose tolerance test (300 mg/kg glucose load with blood sampling for 180 minutes) were significantly lower, and the glucose disappearance rate tended to be faster after the low-fat diet. In contrast, fasting insulin concentrations and the insulin response (insulin area under the curve) to glucose challenge were unchanged. Insulin sensitivity (defined as the rate of glucose disappearance per unit of insulin increase during the period 0 to 40 minutes after the glucose load) was significantly higher on the low-fat diet. These results suggest that the hypotensive effects of a low fat, high-carbohydrate diet, although associated with an improvement in insulin sensitivity, are not mediated by changes in plasma insulin concentration. PMID- 10523331 TI - Effects of sympathectomy and nitric oxide synthase inhibition on vascular actions of insulin in humans. AB - Insulin exerts cardiovascular actions by stimulating nitric oxide (NO) release and sympathetic neural outflow. It is unclear, however, whether insulin stimulates muscle blood flow (and NO release) by a direct action at the vasculature and/or by stimulating neural vasodilator mechanisms. In these studies we used patients with regional sympathectomy to examine the vascular actions of insulin in the presence and absence of sympathetic vasoconstrictor and vasodilator innervation. A 2-hour insulin (6 pmol/kg per minute)/glucose clamp increased muscle blood flow in both innervated and denervated limbs by roughly 40% (P<0.01 versus baseline for both limbs). The vasodilation reached its maximum within the first 30 to 45 minutes of insulin/glucose infusion in sympathetically denervated limbs, but only at the end of the infusion in innervated limbs (P<0. 01, denervated versus innervated limb). Infusion of a NO synthase inhibitor (N(G) monomethyl-L-arginine [L-NMMA]) increased baseline arterial pressure, abolished the vasodilation in the denervated limb, and led to a significant additional increase in arterial pressure during the clamp, but did not alter whole body glucose uptake. Our data indicate that insulin stimulates blood flow in sympathectomized limbs by a direct action at the vasculature. This effect is mediated by stimulation of NO release and appears to be masked by the sympathetic vasoconstrictor tone in innervated limbs. PMID- 10523332 TI - Sex steroids, insulin, and arterial stiffness in women and Men. AB - Arterial stiffness may be influenced by sex steroids and insulin; the association with fasting insulin level may be stronger in women than in men. Therefore, we analyzed the effects of sex steroid administration on (1) arterial stiffness and (2) the relationship between fasting insulin level and arterial stiffness. Twelve male-to-female transsexuals were treated with ethinyl estradiol and cyproterone acetate, and 18 female-to-male transsexuals were treated with testosterone esters, with assessments made at baseline and after 4 and 12 months. Changes in distensibility and compliance coefficients (DC and CC, respectively) of the common carotid artery, femoral artery (FA), and brachial artery (BA) were analyzed in relation to changes in fasting plasma levels of glucose, insulin, HDL cholesterol, and triglycerides. After 4 months of estrogens and antiandrogens in men, significant reductions in the CC and DC of the FA (P=0.006 and P=0.04, respectively) and BA (P=0.04 and P=0.04, respectively) were observed. In women, testosterone, on average, did not affect DC or CC, but the changes in fasting insulin level were strongly negatively associated with changes in the CC and DC, especially in the FA and BA. These associations were significantly less strong in genetic men and were independent of age, mean arterial pressure, and glucose and lipid levels. This experimental study shows (1) that short-term administration of estrogens and antiandrogens increases FA and BA stiffness in men and (2) that the fasting insulin level is a stronger determinant of arterial stiffness in women than in men. PMID- 10523333 TI - Heart rate-dependent stiffening of large arteries in intact and sympathectomized rats. AB - In the anesthetized rat, acute increases in heart rate are accompanied by a reduction in arterial distensibility, which is a significant phenomenon in elastic-type vessels such as the common carotid but much less evident in muscle type vessels such as the femoral artery. Because the sympathetic nervous system importantly reduces arterial distensibility, the present study aimed to determine whether sympathetic influences (1) are involved in the heart rate-dependent changes in arterial distensibility and (2) exert differential effects on elastic type versus muscle-type arteries. To address this issue, 9 sympathectomized (6 hydroxydopamine) and 10 vehicle-treated, 12-week-old, pentobarbitone-anesthetized Wistar-Kyoto rats were subjected to atrial pacing via a transjugular catheter at 5 different randomly sequenced rates (280, 310, 340, 370, and 400 bpm). After each step, spontaneous sinus rhythm was allowed to return to normal. Common carotid and femoral artery diameters were measured by an echo Doppler device (NIUS 01), and blood pressure was measured via catheter inserted into the contralateral vessel. Arterial distensibility was calculated over the systolic diastolic pressure range according to the Langewouters formula. In the common carotid artery, progressive increases in heart rate determined progressive and marked reductions of distensibility (range, 15% to 43%) in sympathectomized and intact rats. In the femoral artery, the stiffening effect of tachycardia was present in sympathectomized rats (range, 21% to 42%), at variance with the inconsistent changes observed in intact rats. In conclusion, our experiments support the notions (1) that in predominantly elastic-type arteries, the stiffening effect of tachycardia is exerted independently of sympathetic modulation of the vessel wall properties and (2) that in predominantly muscle type arteries, removal of sympathetic influences unmasks the stiffening effect of tachycardia. PMID- 10523334 TI - Angiotensin II-stimulated induction of sis-inducing factor is mediated by pertussis toxin-insensitive G(q) proteins in cardiac myocytes. AB - The Janus kinase-signal transducers and activators of transcription (JAK-STAT) pathway is stimulated by angiotensin II (Ang II) via the type 1 receptor after acute pressure overload in the heart. The purpose of this study was to determine whether activation of the JAK-STAT pathway by Ang II is dependent on G proteins. Ang II (100 nmol/L for 120 minutes) caused formation of sis-inducing factor (SIF) complexes and tyrosine phosphorylation of STAT proteins in neonatal rat ventricular myocytes. The percentage of change in Ang II-stimulated SIF induction was not affected by pertussis toxin (PTX) or GP antagonist-2A, compounds that inhibit activation of G(i) and G(o) proteins. In contrast, GP antagonist-2A, a peptide that selectively inhibits activation of G(q) proteins, completely abolished Ang II-stimulated SIF induction and STAT3 tyrosine phosphorylation. Pretreatment of cardiac myocytes with U73122, an inhibitor of phosphatidylinositol-specific phospholipase C (PLC) activity, decreased Ang II stimulated SIF induction and STAT3 tyrosine phosphorylation in a dose-dependent manner. Chelation of intracellular Ca(2+) with BAPTA-AM did not alter Ang II stimulated SIF induction. In contrast, pretreatment of cardiac myocytes with Ro 31-8220, a potent and specific inhibitor of protein kinase C (PKC), decreased Ang II-stimulated SIF induction in a dose-dependent manner. Ang II-stimulated SIF induction was abolished in cardiac myocytes after downregulation of PKC by treatment with PMA. From these data, we conclude that Ang II-stimulated SIF induction and STAT3 tyrosine phosphorylation is mediated by PTX-insensitive G proteins through a G(q)-PLC-PKC-mediated pathway in neonatal rat ventricular myocytes. PMID- 10523335 TI - Apoptosis, coronary arterial remodeling, and myocardial infarction after nitric oxide inhibition in SHR. AB - This study was designed to investigate the relationship between apoptosis (programmed cell death) and coronary arterial remodeling in spontaneously hypertensive rats (SHR) following prolonged nitric oxide synthesis inhibition. In addition, we evaluated whether the development of coronary arterial smooth muscular cell apoptosis was related to hemodynamics or to vascular hypertrophy. Three groups of 20-week-old male SHR were investigated: controls, and two groups that received two doses of N(G)-nitro-L arginine (L-NAME, 50 mg/L and 80 mg/L) each for 3 weeks. Mean arterial pressure and total peripheral resistance index increased whereas cardiac index diminished with L-NAME. Pathohistological study demonstrated increased pericardiac fibrosis and coronary arterial injury score in the L-NAME group in a dose-dependent manner. The high dose of L-NAME (Group 3) produced myocardial infarction in 78% of the rats. The wall:lumen ratio of epicardial coronary arteries was greater in L-NAME treated SHR (0.23+/-0.02 versus 0.16+/-0.02; P<0.05) and was associated with markedly increased apoptosis (15.3+/-6 versus 1. 9+/-1; P<0.05) without smooth muscle cell proliferation (PCNA positive cells). Apoptosis occurred predominantly in hypertrophic coronary arterial smooth muscular cells; myocardial infarction and ventricular fibrosis were exacerbated by impaired hemodynamics induced by L-NAME. These data suggest that coronary endothelial dysfunction and myocardial ischemic disease induced by L-NAME were responsible for apoptosis of coronary arterial smooth muscle cells, myocardial fibrosis, and infarction, all pathological findings that are consistent with what may be found in clinical hypertensive heart disease. PMID- 10523336 TI - In vivo study of AT(1) and AT(2) angiotensin receptors in apoptosis in rat blood vessels. AB - In vitro experiments suggest that angiotensin II (Ang II) may cause growth via angiotensin type 1 (AT(1)) receptors and apoptosis via angiotensin type 2 (AT(2)) receptors. To answer the question of whether AT(1) or AT(2) receptor activation could induce apoptosis in the vasculature in vivo, Wistar rats were infused for 7 days with Ang II (120 ng. kg(-1). min(-1) subcutaneously) and treated with the AT(2) receptor antagonist PD 123319 (30 mg. kg(-1). d(-1) subcutaneously) or the AT(1) receptor antagonist losartan (10 mg. kg(-1). d(-1) orally). Apoptosis in thoracic aorta was quantified by radiolabeled DNA laddering and by terminal deoxynucleotide transferase-mediated dUTP nick end-labeling. The expression of p53, bax, bcl-2, and caspase-3, which play critical roles in apoptotic signaling, was examined by Western blot analysis. The mRNA expression of AT(1) and AT(2) receptors was determined by reverse transcription-polymerase chain reaction. The increase in systolic blood pressure and aortic growth induced by Ang II infusion was completely prevented by losartan alone or losartan given with PD 123319, whereas PD 123319 resulted in a greater increase in systolic blood pressure and aortic growth than Ang II alone. Radiolabeled DNA laddering showed that Ang II infusion+/-losartan or PD 123319 significantly increased apoptosis (147+/-8%, 178+/-20%, and 238+/-41%, respectively, P<0.05 compared with control). Expression of bax and active forms of caspase-3 was increased in the Ang II+PD 123319 group, whereas the expression of p53 and bcl-2 was not significantly different in all groups. The expression of AT(1) and AT(2) receptor mRNA was downregulated by losartan and PD 123319, respectively. Thus, when AT(1) or AT(2) receptors are stimulated in vivo, apoptosis is enhanced in the media of blood vessels. In the case of AT(1) receptor stimulation, this may occur secondary to vascular growth and modulate the latter. Both bax and caspase-3 participate in the pathways of apoptosis triggered by in vivo AT(1) receptor stimulation. PMID- 10523337 TI - Genome scan for blood pressure loci in mice. AB - Hypertension is a complex trait of unknown cause in humans. Mice of the inbred strain BPH/2 serve as a rodent model of human hypertension and display elevated blood pressure compared with the hypotensive strain BPL/1. An F2 intercross of BPH/2 and BPL/1 and 2 backcrosses of BPL/1 with Mus spretus were used to perform interval linkage mapping for systolic blood pressure in a genome scan. Significant linkage was observed in the F2s on chromosome 10 (logarithm of the odds score [LOD]=4.9) and on chromosome 13 in the M spretus backcross (LOD=3.3), with additional suggestive LODs on chromosomes 2, 6, 8, and 18. In addition, several suggestive linkages were observed for phenotypes associated with human hypertension. Our study is the first reported genome-wide linkage scan for blood pressure genes in the mouse. PMID- 10523338 TI - Genetic variants in the epithelial sodium channel in relation to aldosterone and potassium excretion and risk for hypertension. AB - Renin and aldosterone secretion is often lower in blacks than in whites, characteristics that resemble a milder form of Liddle syndrome in which a mutation in the amiloride-sensitive epithelial sodium channel (ENaC) of the kidney results in enhanced resorption of sodium. In the present study, we looked for evidence that the intrinsic level of ENaC activity is indeed higher in blacks than in whites. In overnight urine samples collected from young people (249 white and 181 black subjects, mean age 13.4 years), the urinary aldosterone/potassium ratio, which is typically very low in Liddle syndrome, was lower in blacks than in whites: 0.421+/-0.024 (mean+/-SE) versus 0.582+/-0.016 nmol/mmol (P<0.0001). In addition, all but 1 of 5 molecular variants in ENaC were much more common in blacks than in whites. G442V in the beta-subunit, present in 16% of the blacks and in only 1 white, was associated with parameters reflective of a greater Na retention and potentially a higher ENaC activity: a lower plasma aldosterone concentration (P=0.070), a lower urinary aldosterone excretion rate (P=0.052), a higher potassium excretion rate (P=0.048), and a lower urinary aldosterone/potassium ratio (P=0.027). In a second cohort consisting of 126 black and 161 white normotensive subjects and 232 black and 188 white hypertensive subjects, betaG442V did not show a significant association with hypertension (P=0.089). On the other hand, a variant that was twice as common in whites, alphaT663A, was associated with being normotensive both in blacks (P=0.018) and in whites (P=0.034). Expression of either betaG442V or alphaT663A in Xenopus oocytes did not result in a change in basal Na current, consistent with the variants being in linkage disequilibrium with alleles at active loci. In conclusion, several lines of evidence are presented to suggest that ENaC activity is higher in blacks than in whites, which could contribute to racial differences in Na retention and the risk for hypertension. PMID- 10523340 TI - Congenic substitution mapping excludes Sa as a candidate gene locus for a blood pressure quantitative trait locus on rat chromosome 1. AB - Previously, linkage analysis in several experimental crosses between hypertensive rat strains and their contrasting reference strains have identified a major quantitative trait locus (QTL) for blood pressure on rat chromosome 1 (Chr 1) spanning the Sa gene locus. In this study, we report the further dissection of this Chr 1 blood pressure QTL with congenic substitution mapping. To address whether the Sa gene represents a candidate gene for the Chr 1 blood pressure QTL, congenic strains were developed by introgressing high blood pressure QTL alleles from the stroke-prone spontaneously hypertensive rat (SHRSP) into the normotensive Wistar-Kyoto (WKY-1) reference strain. Congenic animals carrying a chromosomal segment from stroke-prone spontaneously hypertensive rats between genetic markers Mt1pa and D1Rat200 (including the Sa gene locus) show a significant increase in basal systolic and diastolic blood pressure compared with their normotensive Wistar-Kyoto progenitors (P<0.001, respectively), whereas congenic animals carrying a subfragment of this Chr 1 region defined by markers Mt1pa and D1Rat57 (also spanning the Sa gene) do not show elevated basal blood pressure levels (P=0.83 and P=0.9, respectively). Similar results were obtained for NaCl-induced blood pressure values. Thus, the blood pressure QTL on Chr 1 is located centromeric to the Sa gene locus in a region that is syntenic to human chromosome 11p15.4-p15.3. This region excludes the Sa as a blood pressure elevating candidate gene locus on the basis of congenic substitution mapping approaches. PMID- 10523339 TI - Mutants of 11beta-hydroxysteroid dehydrogenase (11-HSD2) with partial activity: improved correlations between genotype and biochemical phenotype in apparent mineralocorticoid excess. AB - Mutations in the kidney isozyme of human 11-hydroxysteroid dehydrogenase (11 HSD2) cause apparent mineralocorticoid excess, an autosomal recessive form of familial hypertension. We studied 4 patients with AME, identifying 4 novel and 3 previously reported mutations in the HSD11B2 (HSD11K) gene. Point mutations causing amino acid substitutions were introduced into a pCMV5/11HSD2 expression construct and expressed in mammalian CHOP cells. Mutations L179R and R208H abolished activity in whole cells. Mutants S180F, A237V, and A328V had 19%, 72%, and 25%, respectively, of the activity of the wild-type enzyme in whole cells when cortisol was used as the substrate and 80%, 140%, and 55%, respectively, of wild-type activity when corticosterone was used as the substrate. However, these mutant proteins were only 0.6% to 5.7% as active as the wild-type enzyme in cell lysates, suggesting that these mutations alter stability of the enzyme. In regression analyses of all AME patients with published genotypes, several biochemical and clinical parameters were highly correlated with mutant enzymatic activity, demonstrated in whole cells, when cortisol was used as the substrate. These included the ratio of urinary cortisone to cortisol metabolites (R(2)=0.648, P<0.0001), age at presentation (R(2)=0.614, P<0.0001), and birth weight (R(2)=0.576, P=0.0004). Approximately 5% conversion of cortisol to cortisone is predicted in subjects with mutations that completely inactivate HSD11B2, suggesting that a low level of enzymatic activity is mediated by another enzyme, possibly 11-HSD1. PMID- 10523341 TI - The role of alpha-adducin polymorphism in blood pressure and sodium handling regulation may not be excluded by a negative association study. AB - The basic requirement for declaring an association study positive is that the "hypertension-favoring" allele is more frequent in hypertensive cases than in normotensive controls. However, both positive and negative associations with hypertension have been found for the same polymorphism when studied in different populations. In the present study, we addressed the question of the possible cause(s) of this discrepancy among populations by using the alpha-adducin polymorphism as a paradigm. Four hundred ninety hypertensives and 176 normotensives enrolled in Sassari, Italy, and 468 hypertensives and 181 normotensives enrolled in Milano, Italy, were genotyped for the alpha-adducin Gly460Trp polymorphism. The blood pressure response to 2 months of hydrochlorothiazide therapy could be evaluated in 143 (85 in Sassari and 58 in Milano) hypertensives with and without the 460Trp alpha-adducin allele. The alpha adducin 460Trp allele was not significantly more frequent in hypertensives in the Sassari population but was more frequent in hypertensives than in normotensives in Milano (P=0.019). Basal plasma renin activity was lower and blood pressure fall after diuretic therapy more pronounced (P<0.01) in hypertensives carrying at least one 460Trp allele than in Gly460Gly homozygotes, irrespective of their membership in the Sassari or Milano cohort. The effect of alpha-adducin genotype in predicting basal plasma renin activity and blood pressure decrease with diuretic treatment is similar in Sassari and Milano, despite the lack of association of the alpha-adducin genotype with hypertension in Sassari. PMID- 10523342 TI - Rarefaction of skin capillaries in borderline essential hypertension suggests an early structural abnormality. AB - We recently showed that rarefaction of skin capillaries in the dorsum of the fingers of patients with essential hypertension is due to the structural (anatomic) absence of capillaries rather than functional nonperfusion. It is not known whether this rarefaction is primary (ie, antedates the onset of hypertension) or secondary (ie, as a consequence of sustained and prolonged elevation of blood pressure [BP]). The aim of the present investigation was to study skin capillary density in a group of patients with mild borderline hypertension to assess whether rarefaction antedates the onset of sustained elevation of BP. The study group included 18 patients with mild borderline hypertension (mean supine BP, 136/83 mm Hg), 32 normotensive controls (mean BP, 126/77 mm Hg), and 45 patients with established essential hypertension (mean BP, 156/98 mm Hg). The skin of the dorsum of the fingers was examined by intravital capillary videomicroscopy before and after venous congestion at 60 mm Hg for 2 minutes. Patients with borderline essential hypertension had the lowest resting capillary density when compared with normotensive controls and patients with established hypertension. Maximal capillary density with venous congestion in the borderline group remained the lowest. The study confirmed that patients with borderline essential hypertension have skin capillary densities that are equally low as or even lower than patients with established hypertension. Both groups had significantly lower capillary densities than normal controls. One explanation for the results is that capillary rarefaction may be due to an early structural abnormality in essential hypertension. PMID- 10523343 TI - Activation of AT(2) receptors by endogenous angiotensin II is involved in flow induced dilation in rat resistance arteries. AB - Pressure-induced tone (myogenic, MT) and flow (shear stress)-induced dilation (FD) are potent modulators of resistance artery tone. We tested the hypothesis that locally produced angiotensin II interacts with MT and FD. Rat mesenteric resistance arteries were perfused in situ. Arterial diameter was measured by intravital microscopy after a bifurcation on 2 distal arterial branches equivalent in size (150 microm, n=7 per group). One was ligated distally and thus submitted to pressure only (MT, no FD). The second branch was submitted to flow and pressure (MT and FD). The difference in diameter between the 2 vessels was considered to be FD. Flow-diameter-pressure relationship was established in the absence and then in the presence of 1 of the following agents. In the nonligated segment (MT+FD), angiotensin II type 1 (AT(1)) receptor blockade (losartan) had no significant effect, whereas angiotensin II type 2 (AT(2)) receptor blockade (PD123319) or saralasin (AT(1)+AT(2) blocker) decreased the diameter significantly, by 9+/-1 and 10+/-0.8 microm, respectively. Angiotensin II in the presence of losartan increased the diameter by 18+/-0.6 microm (inhibited by PD123319). PD123319 or saralasin had no effect after NO synthesis blockade or after endothelial disruption. In the arterial segment ligated distally (MT only), AT(1) or AT(2) receptor blockade had no significant effect. AT(2)-dependent dilation represented 20% to 39% of FD (shear stress from 22 to 37 dyn/cm(2)), and AT(2)-receptor mRNA was found in mesenteric resistance arteries. Thus, resistance arterial tone was modulated in situ by locally produced angiotensin II, which might participate in flow-induced dilation through endothelial AT(2) receptor activation of NO release. PMID- 10523345 TI - Contrasting effects of selective T- and L-type calcium channel blockade on glomerular damage in DOCA hypertensive rats. AB - Mibefradil and amlodipine are calcium antagonists with different channel selectivities. Mibefradil blocks both L- and T-type calcium channels; although in the usual pharmacological doses, it predominantly blocks the T-type channels. In contrast, amlodipine selectively blocks L-type channels. The goal of the present study was to assess whether this differential selectivity would result in different effects on end-organ damage in experimental hypertension. For this purpose, deoxycorticosterone acetate (DOCA)-salt hypertensive rats were treated either with equipotent doses of mibefradil or amlodipine (30 mg. kg(-1). d(-1) as food admix). Despite the fact that both drugs decreased systolic arterial pressure to the same extent (140+/-5 mm Hg in the mibefradil group and 144+/-3 mm Hg in the amlodipine group versus 225+/-5 mm Hg in the untreated-DOCA group), only mibefradil decreased proteinuria (35. 5+/-6.5 versus 103.3+/-14.1 mg/24 h in untreated DOCA-salt animals) and prevented glomerular lesions. Both drugs, however, prevented the occurrence of vascular renal lesions. To elucidate the mechanism responsible for this difference, we evaluated in an additional series of experiments the effects of mibefradil and amlodipine on plasma and renal renin concentrations, as well as the effects of the addition of enalapril, an ACE inhibitor, given on top of both drugs on proteinuria. Amlodipine, in contrast to mibefradil, markedly stimulated the plasma (17.8+/-2.6 ng Ang I. mL(-1). h(-1) in the amlodipine group versus 3.9+/-0.4 ng Ang I. mL(-1). h(-1) in the mibefradil group and 3.2+/-0.3 ng Ang I. mL(-1). h(-1) in the untreated-DOCA group) and renal (2.42+/-0.37 ng Ang I. mL(-1). h(-1) in the amlodipine group versus 0.36+/ 0.04 ng Ang I. mL(-1). h(-1) in the mibefradil group and 0.26+/-0.08 ng Ang I. mL(-1). h(-1) in the untreated-DOCA group) renin concentrations. Stimulation of the renin-angiotensin system could explain the absence of a renal protective effect of amlodipine. This was also suggested by the fact that enalapril given in addition to amlodipine could decrease proteinuria. In conclusion, T-type channel blockade by mibefradil decreases blood pressure without stimulation of the renin angiotensin system and therefore prevents most of the glomerular damage in DOCA hypertensive rats. PMID- 10523344 TI - Aging, high salt intake, and renal dopaminergic activity in Fischer 344 rats. AB - The present study examined renal dopaminergic activity and its response to high salt (HS) intake in adult (6-month-old) and old (24-month-old) Fischer 344 rats. Daily urinary excretion of L-3, 4-dihydroxyphenylalanine (L-DOPA), dopamine, and its metabolites 3, 4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid was similar in adult and old rats; by contrast, daily urinary excretion of norepinephrine in old rats was almost twice that in adult animals. HS intake (1% NaCl) over a period of 24 hours resulted in a 2-fold increase in the urinary excretion of dopamine, DOPAC, and norepinephrine in adult animals but not in old animals. Norepinephrine and L-DOPA plasma levels did not change during HS intake and were similar in both groups of rats. The natriuretic response to an HS intake in old rats (from 4.7+/-0.4 to 10.7+/-2.0 nmol. kg(-1). d(-1); Delta=6.0+/-0.9 nmol. kg(-1). d(-1)) was less than in adult rats (from 5.2+/-0.4 to 13.5+/-2.5 nmol. kg(-1). d(-1); Delta=8.3+/-0.8 nmol. kg(-1). d(-1)). A diuretic response to HS intake was observed in adult rats (from 20.9+/-2.3 to 37.6+/-2.8 mL. kg(-1). d(-1)) but not in old rats (from 37.7+/-5.7 to 42.3+/-6. 0 mL. kg(-1). d(-1)). Dopamine levels and dopamine/L-DOPA ratios in the renal cortex of old rats were greater than in adult rats. HS intake increased both dopamine levels and dopamine/L-DOPA ratios in the renal cortex of adult rats but not in old rats. Aromatic L-amino acid decarboxylase activity was higher in old rats than in adult rats; HS intake increased L-amino acid decarboxylase activity (nmol. mg protein( 1). l5 min(-1)) in adult rats (from 67+/-1 to 93+/-1) but not in old rats (from 86+/-2 to 87+/-2). Dopamine inhibited Na(+),K(+)-ATPase activity in proximal tubules obtained from adult rats, but it failed to exert such an inhibitory effect in old rats. It is concluded that renal dopaminergic tonus in old rats is higher than in adult rats but fails to respond to HS intake as observed in adult rats. This may be due in part to the inability of dopamine to inhibit Na(+),K(+) ATPase activity in old rats. PMID- 10523346 TI - Increased availability of nitric oxide leads to enhanced nitric oxide dependency of tubuloglomerular feedback in the contralateral kidney of rats with 2-kidney, 1 clip Goldblatt hypertension. AB - The contralateral kidney of 2-kidney, 1-clip hypertensive (2K1C) rats is unable to escape the renal vasoconstrictive and sodium-retaining effects of increased circulating angiotensin II levels. Evidence is accumulating that renal function is relatively preserved by enhanced influence of NO in the contralateral kidney. In this study, we investigated (1) whether the high NO dependency of renal hemodynamics in the contralateral kidney is due to increased availability of NO or increased sensitivity to NO and (2) whether elevated NO activity dampens the actions of angiotensin II to enhance tubuloglomerular feedback (TGF) responses in the nonclipped kidney of 2K1C rats. To estimate whether the available NO is increased, the NO clamp technique was applied in rats that underwent sham operation (n=6) and in the contralateral kidney of 2K1C Sprague-Dawley rats (3 weeks old; 0.25-mm silver clip; n=6). During systemic infusion of nitro-L arginine (L-NNA; 50 microg/kg. min(-1)), sodium nitroprusside (SNP) was infused in the renal artery and the rate was adjusted so that renal vascular resistance (RVR) was restored to baseline levels. In sham rats, RVR increased during L-NNA treatment from 17.2+/-2.0 to 33.0+/-3.6 U (P<0.01) and was restored to baseline values during SNP infusion (17.1+/-2.3 U); 9. 2+/-1.8 nmol/min of SNP was needed to restore RVR to baseline values. In 2K1C rats, RVR increased during L-NNA treatment from 16.7+/-1.1 to 53.4+/-3.5 U (P<0.01). This increase of RVR was significantly larger than in sham rats. RVR was restored to baseline values during SNP infusion (17.4+/-0.9 U); 26.0+/-4.3 nmol/min of SNP was needed to restore RVR to baseline values (P<0.05 versus sham). Furthermore, maximum TGF responses were assessed before and during late proximal tubular infusion of L-NNA in the kidneys of sham rats and the nonclipped kidneys of 2K1C rats. Control maximum TGF responses were 4.7+/-0.7 and 5.1+/-0.4 mm Hg in sham and 2K1C rats, respectively. During intraluminal L-NNA infusion, maximum TGF responses were 15. 4+/-0.9 mm Hg in sham rats and 22.2+/-2.5 mm Hg in 2K1C rats (P<0.05 versus sham). Finally, urinary NO(2)+NO(3) excretion in the nonclipped kidney was significantly higher than in the clipped kidney (P<0.05). In conclusion, (1) as assessed using the NO clamp, ambient intrarenal NO levels are increased in the contralateral kidney of 2K1C rats and (2) the NO dependency of the TGF system is enhanced. These experiments indicate that adaptations in NO activity lead to relatively low TGF responsiveness, which will offset the simultaneous sodium retaining actions of angiotensin II on proximal tubular reabsorption and TGF responsiveness. PMID- 10523347 TI - Ambulatory physical activity as a determinant of diurnal blood pressure variation. AB - There are reports that indicate that diurnal blood pressure (BP) variation, in addition to high BP per se, is related to target organ damage and the incidence of cardiovascular events. However, the determinants of diurnal BP variation are not adequately understood. We used actigraphy and ambulatory BP monitoring to study the diurnal variation of BP and physical activity in 160 adults. Within individuals, activity was more strongly related to pulse rate than to BP. The correlation between BP and activity was stronger during sleep than when awake, but the correlation between activity and pulse rate was higher during the awake period than during sleep. Between individuals, the sleep/awake ratio of systolic BP (SBP) was correlated with mean sleep activity (r=.17, P<0.05), mean awake activity (r=-0.16, P<0.05), and, especially, the ratio of sleep/awake activity (r=.24, P<0.01). Awake BP variability (SD of awake SBP) was positively correlated with awake activity (r=.16, P<0. 05). In regard to the effect of position, the standing-supine SBP difference was negatively correlated with the sleep/awake SBP ratio (r=-0.39, P<0.01) and positively correlated with awake SBP variability (r=.33, P<0.01). When we divided the subjects into 3 groups, 19 extreme dippers (with a sleep SBP decrease of >/=20% of awake SBP), 102 dippers (with decreases of >/=10% to <20%), and 39 nondippers (with decreases of <10%), no significant differences existed in awake activity among the groups. However, the nondippers exhibited greater sleep activity than extreme dippers (P<0.05) and an increased sleep/awake activity ratio compared with extreme dippers and dippers (P<0.01). Extreme dipping may also be associated with increased BP variability (P=0.08). Individual SBP responses to activity (the within-person slope of awake SBP regressed on activity) did not differ significantly among the 3 subgroups. In conclusion, physical activity is one of the determinants of ambulatory BP and its diurnal variation. We hypothesize that the association of sleep activity to sleep BP and dipping reflects differences in sleep quality. PMID- 10523348 TI - State-of-the-Art lecture: influence of exercise training on neurogenic control of blood pressure in spontaneously hypertensive rats. AB - Exercise training plays an important role in the reduction of high blood pressure. In this review, we discuss the effect of distinct intensities of exercise training on the reduction of high blood pressure in spontaneously hypertensive rats (SHR). In addition, we present some hemodynamic mechanisms and associated neural controls by which exercise training attenuates hypertension in SHR. Low-intensity exercise training is more effective in reducing high blood pressure than is high-intensity exercise training in SHR. The decrease in blood pressure is due to resting bradycardia, and in consequence, lower cardiac output. Sympathetic attenuation to the heart is the major explanation for the resting bradycardia. Recovery of the sensitivity of baroreflex control of heart rate, which is usually impaired in SHR, is an important neurogenic component involved in the benefits elicited by exercise training. PMID- 10523349 TI - Sympathetic activation in the pathogenesis of hypertension and progression of organ damage. AB - Although animal models of hypertension have clearly shown that high blood pressure is associated with and is probably caused by an increase in sympathetic cardiovascular influences, a similar demonstration in humans has been more difficult to obtain for methodological reasons. There is now evidence, however, of increased sympathetic activity in essential hypertension. This article will review this evidence by examining data showing that plasma norepinephrine is increased in essential hypertension and that this is also the case for systemic and regional norepinephrine spillover, as well as for the sympathetic nerve firing rate in the skeletal muscle nerve district. Evidence will also be provided that sympathetic activation is a peculiar feature of essential hypertension, particularly in its early stages, with secondary forms of high blood pressure not usually characterized by an increased central sympathetic outflow. Humoral, metabolic, reflex, and central mechanisms are likely to be the factors responsible for the adrenergic activation characterizing hypertension, which may also promote the development and progression of the cardiac and vascular alterations that lead to hypertension-related morbidity and mortality, independent of blood pressure values. This represents the rationale for considering sympathetic deactivation one of the major goals of antihypertensive treatment. PMID- 10523350 TI - L-arginine restores the effect of ouabain on baroreceptor activity and prevents hypertension. AB - In spontaneously hypertensive rats, ouabain exerts an excitatory effect on baroreceptor nerve activity (BNA). The aim of this study was to determine the effects of ouabain on BNA in other experimental models of hypertension and its interaction with nitric oxide. Rats were made hypertensive using the procedures for N(omega)-nitro-L-arginine methyl ester (L-NAME), deoxycorticosterone acetate (DOCA) salt, and 2-kidney, 1 clip (2K1C) hypertension models. In these groups, systolic arterial pressure was 195+/-7, 149+/-6, and 148+/-4 mm Hg, respectively, compared with 110+/-4 mm Hg in normotensive rats. Acute ouabain administration had an excitatory effect on BNA in normotensive rats (37+/-4%), an inhibitory effect in L-NAME hypertensive rats (-60+/-7%), and no effect in DOCA-salt and 2K1C hypertensive rats. The effects of ouabain were not related to arterial pressure levels, and no excitatory effect on BNA was observed in prehypertensive DOCA-salt rats. Long-term administration of L-arginine (3 g x kg(-1) x day(-1)) prevented DOCA-salt (121+/-8 mm Hg) and 2K1C (104+/-4 mm Hg) hypertension, markedly attenuated L-NAME (130+/-9 mm Hg) hypertension, and restored the excitatory effect of ouabain on BNA in these groups to levels similar to the normotensive rats and their respective control groups. We conclude that ouabain has a diverse effect on BNA in experimental models of hypertension, and it can be normalized by L-arginine. The data also indicate that nitric oxide may play a pivotal role in mediating the excitatory effect of ouabain on BNA, and we speculate that a therapeutic combination of ouabain and L-arginine may be beneficial in secondary hypertension. PMID- 10523351 TI - Digoxin prevents ouabain and high salt intake-induced hypertension in rats with sinoaortic denervation. AB - Digoxin prevents ouabain-induced hypertension in rats. In the present study, we tested whether this effect of digoxin depends on its sensitizing effect on baroreflex function or is due to an antagonistic action on exogenous ouabain or endogenous ouabain-like activity ("ouabain") in the brain. In Wistar rats, resting mean arterial pressure (MAP) was significantly increased by long-term subcutaneous (SC) ouabain (75 microg/d) plus high salt (8%) intake for 12 days (but not after only 5 days). In rats with chronic sinoaortic denervation (SAD), MAP was increased within 5 days of ouabain treatment to the same extent as MAP after 12 days of treatment in intact rats. The effect of ouabain and high salt was prevented when digoxin was given SC concomitantly via osmotic minipump (200 microg x kg(-1) x d(-1)). Resting MAP was not changed in rats treated with digoxin alone. In a second set of rats with chronic SAD or sham surgery, high salt intake was given for 14 days, with or without SC digoxin (200 microg x kg( 1) x d(-1)) or intracerebroventricular (ICV) antibody Fab fragments (200 microg/d), which bind "ouabain" with high affinity. On day 14, MAP, central venous pressure, heart rate, and renal sympathetic nerve activity were recorded in conscious rats at rest and in response to air-jet stress, IV phenylephrine and nitroprusside, and acute volume expansion with 5% dextrose IV. In rats with SAD versus sham surgery, high salt significantly increased resting MAP as well as excitatory responses of MAP, heart rate, and renal sympathetic nerve activity to air stress. These effects of high salt in rats with SAD were prevented by digoxin or Fab fragments. Arterial baroreflex function was blunted but cardiopulmonary baroreflex function was not affected in rats with SAD. Digoxin and Fab fragments had no effects on either function. In an in vitro assay for the inhibitory effects on Na+, K(+)-ATPase activity, 20 ng of ouabain caused 29% inhibition, but 20 ng of ouabain plus 13 or 53 ng of digoxin caused only 16% or 4% inhibition, respectively. These data indicate that the arterial baroreflex opposes sympathoexcitatory responses to ouabain and "ouabain" in the brain, thereby delaying ouabain- and preventing high salt-induced hypertension in Wistar rats. In addition to possible effects on the arterial baroreflex, digoxin appears to act centrally to prevent the sympathoexcitatory and pressor effects of increased brain "ouabain" or ouabain. PMID- 10523352 TI - Commissural NTS lesions and cardiovascular responses in aortic baroreceptor denervated rats. AB - Both acute (1 day) lesions of the commissural nucleus of the solitary tract (commNTS) and aortic baroreceptor denervation increase pressor responses to bilateral common carotid occlusion (BCO) during a 60-second period in conscious rats. In this study, we investigated the following: (1) the effects of commNTS lesions on basal mean arterial pressure (MAP) and heart rate (HR) of aortic denervated (ADNx) rats; (2) the effects of acute commNTS lesions on pressor responses to BCO in ADNx rats; and (3) the effects of chronic (10 days) commNTS lesions on the pressor response to BCO. ADNx increased basal MAP and HR in sham lesioned rats. Acute commNTS lesions abolished the MAP and HR increases observed in ADNx rats. Acute commNTS lesions increased the pressor responses to BCO in rats with intact-baroreceptor innervation but produced no additional change in the pressor response to BCO in ADNx rats. Chronic commNTS lesions did not change the pressor responses to BCO in rats with intact-baroreceptor innervation. The data show that acute commNTS lesions abolish the MAP increase produced by aortic baroreceptor denervation. They also suggest that acute commNTS lesions enhance the pressor response to BCO by partial withdrawal of aortic baroreceptor inputs into the NTS. Chronically, reorganization in the remaining aortic baroreceptor or in the baroreflex function as a whole might produce normalization of the cardiovascular responses to BCO. PMID- 10523353 TI - Rostral ventrolateral medulla : A source of sympathetic activation in rats subjected to long-term treatment with L-NAME. AB - The major aim of the present study was to evaluate the role of the rostral ventrolateral medulla (RVLM) in the maintenance of hypertension in rats subjected to long-term treatment with N(G)-nitro-L-arginine methyl ester (L-NAME) (70 mg/kg orally for 1 week). We inhibited or stimulated RVLM neurons with the use of drugs such as glycine, L-glutamate, or kynurenic acid in urethane-anesthetized rats (1.2 to 1.4 g/kg IV). Bilateral microinjection of glycine (50 nmol, 100 nL) into the RVLM of hypertensive rats produced a decrease in mean arterial blood pressure (MAP) from 158+/-4 to 71+/-4 mm Hg (P<0.05), which was similar to the decrease produced by intravenous administration of hexamethonium. In normotensive rats, glycine microinjection reduced MAP from 106+/-4 to 60+/-3 mm Hg (P<0.05). Glutamate microinjection into the RVLM produced a significant increase in MAP in both hypertensive rats (from 157+/-3 to 201+/-6 mm Hg) and normotensive rats (from 105+/-5 to 148+/-9 mm Hg). No change in MAP was observed in response to kynurenic acid microinjection into the RVLM in either group. These results suggest that hypertension in response to long-term L-NAME treatment is dependent on an increase in central sympathetic drive, mediated by RVLM neurons. However, glutamatergic synapses within RVLM are probably not involved in this response. PMID- 10523354 TI - Nitric oxide-dependent guanylyl cyclase participates in the glutamatergic neurotransmission within the rostral ventrolateral medulla of awake rats. AB - A well-known action of nitric oxide (NO) is to stimulate the soluble form of guanylyl cyclase, evoking an accumulation of cyclic GMP in target cells. The aim of the present study was to examine the effects of inhibition of guanylyl cyclase dependent on NO during cardiovascular responses induced by L-glutamate and S nitrosoglutathione (SNOG) microinjected into the rostral ventrolateral medulla (RVLM) of awake rats. Three days before the experiments, adult male Wistar rats (280 to 320 g) were anesthetized for implantation of guide cannulas to the desired stereotaxic position (AP=-2.5 mm, L=1.8 mm) in relation to lambda. The cannulas were fixed to the skull with acrylic cement. Twenty-four hours before the experiments, a femoral artery and vein were cannulated for recording arterial pressure (AP) and heart rate (HR) and injection of anesthetic. Unilateral microinjections (100 nL) of L-glutamate (5 nmol/L) and SNOG (2.5 nmol/L) were made into the histologically confirmed RVLM. The cardiovascular responses to these drugs were evaluated before and after microinjection (3 nmol/L, 200 nL) of either methylene blue or oxodiazoloquinoxaline (ODQ). The hypertensive effect of L-glutamate was attenuated by 74% after methylene blue (DeltaAP=49+/-8 to 13+/-4 mm Hg) and by 80.5% after ODQ (DeltaAP=30+/-2 to 6+/-2 mm Hg). The increase in AP produced by SNOG was fully blocked by ODQ (DeltaAP=39+/-8 to 1+/-2 mm Hg). These data indicate that cyclic GMP mechanisms have a key role in glutamatergic neurotransmission in the RVLM of awake rats, and it is most probable that NO participates in this response. PMID- 10523355 TI - Importance of glycinergic and glutamatergic synapses within the rostral ventrolateral medulla for blood pressure regulation in conscious rats. AB - In this study we used a method that permits bilateral or unilateral microinjections of drugs into the rostral ventrolateral medulla (RVLM) of conscious, freely moving rats. There is only limited information about how sympathetic vasomotor tone is maintained by premotor RVLM neurons in conscious animals. It has long been known that glycine microinjection into the RVLM region leads to a decrease in blood pressure (BP) in anesthetized animals. In the present study we show that both unilateral and bilateral microinjection of glycine at the same dose used for anesthetized rats (50 nmol, 50 nL) into the RVLM increases BP in conscious animals. A similar response was also observed when the excitatory amino acid L-glutamate was microinjected into the RVLM. The microinjection of kynurenic acid into the RVLM did not change the basal level of BP but blocked the increase in BP after glycine or glutamate microinjection. A decrease in BP was only observed when low doses of glycine were used (1 to 10 nmol). We conclude that, in conscious animals, the hypertension occurring in response to high doses of glycine into the RVLM is dependent on glutamatergic synapses within the RVLM. A decrease in BP observed when low doses of glycine were used shows that in conscious animals, the RVLM, in association with other premotor neurons, is probably responsible for the maintenance of sympathetic vasomotor tone, because glycine is less effective in decreasing BP under these circumstances than in anesthetized animals. PMID- 10523357 TI - Effects of acute AV3V lesions on renal and hindlimb vasodilation induced by volume expansion. AB - The role of the anteroventral third ventricle (AV3V) region in the cardiovascular adjustments to volume expansion (VE) with 4% Ficoll (1% body weight, 1.4 mL/min) was studied in urethane-anesthetized rats. In sham-lesioned animals, VE produced a transitory (/=60 years) patients. Pulse pressure decreased in parallel with stroke index from age >30 to 40 to 49 years. Upright posture, however, eliminated this difference through a larger orthostatic fall in stroke index and pulse pressure in the youngest patients. After age 50 years, pulse pressure exhibited a progressive widening despite the further age-related decrease in stroke index. Supine, upright, and 24-hour pulse pressure fitted a curvilinear correlation with age (r=0.55, 0.56, and 0.68, respectively, P<0.001), with a transition at age 50 years. Before age 50 years, 24-hour pulse pressure correlated positively with stroke volume (r=0.5, P<0.001) and negatively with arterial compliance (SV/PP ratio, r=-0.37, P<0.01). In contrast, in men >/=50 years old, 24-hour pulse pressure correlated negatively with the SV/PP ratio (r=-0.5; P<0.01), without significant influence of stroke volume. Thus, in hypertensive men, the age-related change in stroke volume significantly accounted for the change in clinic and ambulatory pulse pressure during young adulthood, but its contribution decreased after the fifth decade. PMID- 10523366 TI - Cardiopulmonary reflex impairment in experimental diabetes in rats. AB - The aim of the present study was to evaluate the sensitivity of the cardiopulmonary receptors in experimental diabetes induced by streptozotocin by the use of 2 different methods: (1) administration of increasing doses of serotonin to analyze peak changes of arterial pressure and heart rate for each given dose in conscious intact normal and diabetic rats; (2) expanding blood volume with the use of dextran (6%) to produce similar increases in left ventricular end-diastolic pressure to quantify the arterial pressure, heart rate, and renal sympathetic nerve activity in sinoaortic, denervated, anesthetized normal and diabetic rats. Blood samples were collected to measure blood glucose. Diabetic rats showed hyperglycemia (22+/-0. 7 versus 7+/-0.2 mmol/L), reduced body weight (226+/-12 versus 260+/-4 g) and heart rate (294+/-14 versus 350+/-10 bpm), and similar arterial pressure (104+/-4 versus 113+/-4 mm Hg) when compared with control rats. Serotonin induced significant bradycardia and hypotension, which were similar and proportional to the dose injected in both groups. Mean arterial pressure and heart rate decreases in response to volume overload were significantly lower in diabetic than in control rats. The reflex reduction of the renal sympathetic nerve activity as expressed by percentage changes in nerve activity in response to increasing left end-diastolic pressure was abolished in diabetic animals (1.9+/-0.8% versus -14+/-4%/mm Hg in controls). These results showed an impairment of cardiopulmonary reflex control of circulation in diabetes during acute volume expansion. The normal responses to serotonin administration indicated that the cardiopulmonary reflex is still preserved in diabetic rats. PMID- 10523367 TI - Ambulatory blood pressure: normality and comparison with other measurements. Hypertension Working Group. AB - Previous studies have reported results on 24-hour ambulatory blood pressure (ABP) in Europe and Japan, but no data exists from South America. In this study, we conducted a population survey to identify reference values and to compare ambulatory blood pressure with clinic, home, and self-measured values. A random sample of 2650 adults was selected among 190 000 people covered by our prepaid healthcare institution. Clinic (physician and nurse) and home (nurse) blood pressure measurements were performed 3 times each, with semiautomatic electronic equipment. Self-measurements were performed by the subjects manually activating the ambulatory device. We analyzed 1573 individuals who were not receiving antihypertensive therapy from 1921 participants. Self-measurement was available in a subgroup of 577 participants younger than the whole sample. Normal ambulatory blood pressure limits were estimated as those that best correlated with 140/90 mm Hg at clinic. Estimated values were 125/80 mm Hg for 24-hour ambulatory (range: 122 to 128 and 77 to 83 mm Hg) and 129/84 mm Hg for daytime ambulatory (range: 127 to 132 and 81 to 86) blood pressure, depending on gender and age. Ambulatory and clinic blood pressures increased with age. The age dependent increase in ABP was similar in women and men. Average blood pressure at clinic was 124/79 mm Hg by physician and 123/78 mm Hg by nurse. Nurse measurement at home was 125/78 mm Hg, daytime ambulatory was 121/77 mm Hg, and 24-hour ambulatory was 118/74 mm Hg. The values of the subgroup with self-measurement were physician 119/77 mm Hg; nurse at clinic 118/77 mm Hg; nurse at home 121/78 mm Hg; self-measured 115/72 mm Hg; daytime ambulatory 119/77 mm Hg; and 24-hour ambulatory 115/73 mm Hg. This study shows that a 24-hour ABP average value of 125/80 mm Hg and a daytime ABP average value of 129/84 mm Hg are suitable upper limits for normality. Higher limits would yield an artificially higher prevalence of white coat hypertension. Most subjects showed higher blood pressure levels when measurements were performed by healthcare personnel at a clinic or at home than when self-measured at home. PMID- 10523368 TI - Kinin B2 receptors mediate blockade of atrial natriuretic peptide natriuresis induced by glucose or feeding in fasted rats. AB - We have shown previously that the kininogen-derived peptides bradykinin, prokinins, and PU-D1, given intravenously or into the duodenal lumen, block the atrial natriuretic peptide (ANP)-induced diuretic-natriuretic effect in fasting, anesthetized rats infused with isotonic glucose. HOE-140, an inhibitor of bradykinin B2 receptors, completely suppresses this ANP blockade. When intravenous glucose infusion is omitted, the above-described inhibition of ANP does not take place. Therefore, to clarify the role of glucose and/or feeding in this phenomenon, we used fasted, anesthetized rats to test how the ANP excretory response was affected by (1) short-term feeding before anesthesia, (2) 1 mL of isotonic glucose introduced into the stomach, and (3) the interaction of HOE-140 with these treatments. In addition, we tested the effects of 1 mL of intragastric glucose administration and HOE-140 on urinary excretion in awake rats. In anesthetized rats, both glucose administration and feeding significantly inhibited the diuretic-natriuretic effect of ANP for up to 90 minutes. Similarly, intragastric glucose delayed spontaneous sodium and water excretion for 90 minutes in awake rats. In all 3 cases, pretreatment with HOE-140 (2.5 microg IV) fully prevented the inhibition of ANP excretory action, ruling out osmotic effects as the cause of reduced diuresis. These results indicate that the presence of glucose in the digestive tract triggers an inhibitory effect on ANP renal actions that requires activation of kinin B2 receptors, providing strong support to our hypothesis that during the early prandial period, gastrointestinal signals elicit a transient blockade of renal excretion with a mechanism involving the kallikrein-kinin system. PMID- 10523369 TI - Responses to acute changes in arterial pressure on renal medullary nitric oxide activity in dogs. AB - A direct relationship between renal arterial pressure (RAP) and cortical tissue nitric oxide (NO) activity in the canine kidney was reported earlier. We have conducted further experiments to evaluate medullary NO responses to alterations in RAP with the use of a NO-selective microelectrode inserted into the renal medulla of 6 anesthetized, sodium-replete dogs. Graded reductions in RAP (from 140 to 80 mm Hg) elicited decreases in medullary tissue NO concentration, with a slope of 10.2+/-4.5 nmol x L(-1) x mm Hg(-1). These changes in NO levels were associated with decreases in urinary excretion rate of nitrate and nitrite (U(NOx)V; control value, 1.7+/-0.03 nmol x min(-1) x g(-1); slope, 0.02+/-0.004 nmol x min(-1) x g(-1) x mm Hg(-1)) and sodium excretion (U(Na)V; control, 3.2+/ 0.7 micromol x min(-1) x g(-1); slope, 0.06+/-0.02 micromol x min(-1) x g(-1) x mm Hg(-1)) without changes in glomerular filtration rate control (0.84+/-0.06 mL x min(-1) x g(-1)). Intra-arterial administration of the NO synthase inhibitor N(omega)-nitro-L-arginine (NLA; 50 microg x kg(-1) x min(-1)) decreased medullary NO concentration by 218+/-55 nmol x L(-1) (n=5) and attenuated the relationship between RAP and NO concentration (slope, 2.7+/-2.2 nmol x L(-1) x mm Hg(-1)). NLA infusion decreased U(NOx)V (0.8+/-0.06 nmol x min(-1) x g(-1)) and U(Na)V (1.1+/ 0.2 micromol x min(-1) x g(-1)) without changes in glomerular filtration rate and attenuated RAP versus U(Nox)V and U(Na)V relationships. Total and regional blood flows, as measured by electromagnetic and laser Doppler needle flow probes, respectively, remained autoregulated both before and during NLA infusion. These data support the hypothesis that acute changes in RAP elicit changes in intrarenal NO production, which may participate in the mediation of pressure natriuresis. PMID- 10523370 TI - Plasma homocysteine, aortic stiffness, and renal function in hypertensive patients. AB - Hyperhomocysteinemia has been associated with both vascular structure alterations and vascular clinical end points. To assess the relation between plasma homocysteine, structure and function of large arteries, and the presence of clinical vascular disease, we investigated a population of 236 hypertensive patients. We estimated arterial stiffness by measuring the carotid-femoral pulse wave velocity. Total plasma homocysteine was determined by fluorometric high performance liquid chromatography. The presence of cardiovascular disease was defined on the basis of clinical events, including coronary heart disease, cerebrovascular disease, and peripheral vascular disease. In this population, pulse wave velocity was positively correlated with homocysteine, even after adjustments for age, mean blood pressure, extent of atherosclerosis, and creatinine clearance (P=0.016). Analysis of variance showed statistically significant differences between the mean values of homocysteine, creatinine clearance, and pulse wave velocity according to the extent of atherosclerosis, with an increase in these 3 parameters concomitant with an increase in the number of vascular sites involved with atherosclerosis. In conclusion, in hypertensive patients the levels of homocysteine are strongly and independently correlated to arterial stiffness measured by aortic pulse wave velocity. Plasma homocysteine, creatinine clearance, and aortic pulse wave velocity are higher in patients presenting with clinical vascular disease. These results suggest that the evaluation of aortic distensibility and homocysteine levels can help in cardiovascular risk assessment in hypertensive populations. PMID- 10523371 TI - Cyclooxygenase-2 modulates afferent arteriolar responses to increases in pressure. AB - This study was designed to examine the contribution of cyclooxygenase-2 (COX-2) in the afferent arteriolar autoregulatory responses to increases in perfusion pressure and its relationship with neuronal nitric oxide synthase (nNOS). In rat kidneys, afferent arteriolar diameter responses to increases in perfusion pressure were assessed in vitro with the blood-perfused juxtamedullary nephron technique. Basal afferent arteriolar diameter at 100 mm Hg averaged 21.0+/-1.2 microm (n=7), and the vasoconstrictor response to increasing perfusion pressure to 160 mm Hg averaged 18.4+/-1.2%. Superfusion with the COX-2 inhibitor NS398 (10 micromol/L) did not influence basal diameters, but it did significantly enhance the vasoconstrictor response to the increase in perfusion pressure (32.9+/-4.0%). In contrast to previous findings that the nNOS inhibitor S-methyl-L thiocitrulline (10 micromol/L) enhanced afferent arteriolar autoregulatory responses in normal rat kidneys, in this study, administration of 10 micromol/L S methyl-L-thiocitrulline did not further modulate the vasoconstrictor response to increases in perfusion pressure in the NS398-treated kidneys of normal rats (31.8+/-4.7%). When tubuloglomerular feedback activity was interrupted by papillectomy and the addition of 50 micromol/L furosemide to the blood perfusate (n=5 for each), the afferent arteriolar constrictor responses to increasing perfusion pressure to 160 mm Hg averaged 7.9+/-0.9% and 10.7+/-0.7%, respectively, and they were significantly attenuated compared with the responses observed in control kidneys. NS398 treatment did not modulate the afferent arteriolar autoregulatory responses in papillectomized or furosemide-treated kidneys. These results indicate that COX-2-derived metabolites contribute to the nNOS modulation of pressure-mediated afferent arteriolar autoregulatory responses. PMID- 10523372 TI - Effect of cyclooxygenase-2 inhibition on renal function after renal ablation. AB - Kidney failure is the common end of hypertension and renal diseases. Several authors have suggested that vasodilatory prostaglandins participate in the hemodynamic mechanism responsible for the development of kidney failure. However, the mechanism by which prostaglandins are increased in renal disease is not clear. Recently, 2 isoforms of the enzyme responsible for prostaglandin synthesis, cyclooxygenase, have been described as cyclooxygenase-1 (COX-1), a constitutive isoform, and cyclooxygenase-2 (COX-2), an inducible isoform. In the present study, we investigated whether COX-2-dependent prostaglandins participate in the evolution of renal functional changes after renal ablation. We inhibited prostaglandin synthesis by COX-1 and COX-2 with indomethacin (3 mg/kg) and prostaglandin synthesis by COX-2 with NS-398 (3 mg/kg) and tested the effect of these inhibitors on the renal functional changes elicited by renal ablation. Renal ablation produced an increase in urinary volume, protein, and prostaglandin E(2), whereas urinary sodium and potassium were not affected and urinary osmolarity decreased; treatment with indomethacin or NS-398 partially prevented the renal functional changes elicited by renal ablation. Immunoblots for COX showed an increase in the expression of COX-2 protein 2 days after renal ablation. Furthermore, COX-2 mRNA expression was increased 1 day after renal ablation. These data suggest that COX-2-dependent prostaglandins participate in the renal mechanisms associated with the development of renal functional changes after renal ablation. PMID- 10523373 TI - Effects of amlodipine on tubulointerstitial lesions in normotensive hyperoxaluric rats. AB - Although controversial, a number of reports have suggested that calcium antagonists can retard or prevent the progression of various renal diseases in experimental models. Nevertheless, there are few data related to tubulointerstitial changes in these studies. On the other hand, hyperoxaluria is a recognized cause of tubulointerstitial lesions, and this could contribute to the development of hypertension and chronic renal failure. The aim of the present study was to evaluate a possible beneficial effect of amlodipine, a 1,4 dihydropyridine class of calcium antagonist, in a model of primary tubulointerstitial lesion produced by hyperoxaluria. Two-month-old male Sprague Dawley rats were separated into 4 groups for a 4-week period: G1 (control; tap water only); G2 (hyperoxaluric); G3 (hyperoxaluric plus amlodipine treatment); and G4 (amlodipine treatment). G2 and G3 rats were given 1% ethylene glycol (a precursor for oxalates) in drinking water, and G3 and G4 rats were given amlodipine 2 mg. kg(-1). d(-1) by gavage. At the end of the study, we evaluated by semiquantitative scores (0 to 4) the different renal tubulointerstitial lesions, urinary albumin excretion, renal function by creatinine clearance, and blood pressure. Rats belonging to the hyperoxaluric group treated with amlodipine (G3) had fewer tubulointerstitial lesions, as follows: (1) inflammatory infiltrate score: 3.31+/-0.07 versus 0.23+/-0.12; P<0.05; (2) tubular atrophy score: 3.33+/-0.33 versus 0.50+/-0.22, P<0.05; (3) interstitial fibrosis score: 2.76+/-0.34 versus 0.31+/-0. 16, P<0.05; (4) oxalate deposits score: 3.66+/-0.33 versus 0.09+/-0. 08, P<0.05; (5) lower urinary albumin excretion (11.3+/-2 versus 27+/-4.5 mg/d, P<0.01); and (6) higher creatinine clearance (1. 22+/-0.08 versus 1.13+/-0.08, P<0.01) compared with the hyperoxaluric group untreated with amlodipine (G2). On the other hand, there were no significant changes in blood pressure in any group. In view of these data, we suggest that amlodipine, probably by nonhemodynamic mechanisms of action, can provide an important benefit in the prevention of epithelial tubular cell injury and inflammatory response and therefore in the prevention of the progressive tubulointerstitial fibrosis caused by oxalates. PMID- 10523374 TI - In vivo renal vascular and tubular function in experimental hypercholesterolemia. AB - Hypercholesterolemia (HC) is often associated with impaired peripheral and coronary vascular responses to endothelium-dependent vasodilators, which are probably due to low bioavailability of nitric oxide. To examine the effect of HC on renal vascular and tubular function, 22 domestic pigs were studied after being fed a 12-week normal (n=11) or HC (n=11) diet. Renal regional perfusion and intratubular contrast media concentration in each nephron segment (representing fluid reabsorption) were quantified in vivo with electron-beam computed tomography before and after a suprarenal infusion of either acetylcholine (6 pigs of each diet) or sodium nitroprusside (SNP; 5 pigs of each diet). An increase in cortical perfusion, observed in normal pigs with acetylcholine (+35+/-6%, P=0. 002) and SNP (+12+/-4%, P=0.005), was blunted in the HC group (+8. 8+/-4.0, P=0.01, and -4.6+/-4.0%, P=0.1, respectively, P=0.003 and P=0.005 compared with normal) as was an increase in medullary perfusion (+58+/-21 in normal versus +24+/-11% in HC, P=0.04). A decrease in the intratubular contrast media concentration in the distal tubule and collecting duct of normal pigs was observed in all tubular segments (and was significantly enhanced in the proximal tubule and Henle's loop) in the HC group, which was associated with increased sodium excretion. The tubular and renal excretory responses to SNP were similar between the groups. In conclusion, early experimental HC in the pig attenuates renal perfusion response to both endothelium-dependent and -independent vasodilators possibly because of decreased bioavailability or decreased vascular responsiveness to nitric oxide. This vascular impairment may play a role in maladjusted renovascular responses and contribute to renal damage in later stages of atherosclerosis. PMID- 10523375 TI - Long-term nitric oxide synthase inhibition in rat pregnancy reduces renal kallikrein. AB - This study was performed to test the hypothesis that long-term nitric oxide synthase (NOS) inhibition during pregnancy may alter the predominance of the vasodilator kallikrein system. Sprague-Dawley rats were treated with the competitive inhibitor of NOS N(omega)-nitro-L-arginine (L-NNA, 50 mg. kg(-1). d( 1), dissolved in water) from days 7 to 21 of pregnancy. Rats were studied before treatment (day 5), at days 11, 17, and 21 of pregnancy (during treatment), and at postpartum days 7 and 21 (after the drug was withdrawn at delivery). Each group (n=5 to 8) had its corresponding control group (C) that received only vehicle. Additional rats were treated with N(G)-nitro-L-arginine methyl ester (L-NAME) alone or with an excess of L-arginine. At each study day, we measured blood pressure, collected urine overnight, obtained blood samples, and processed the kidneys for conventional histology and immunohistochemistry. In L-NNA rats, fetal and placental weights were reduced at days 17 and 21. Blood pressure was higher at days 17 and 21, returning to normal after L-NNA was removed. Urinary kallikrein activity was lower at days 11 and 17 (L-NNA=1147+/-213 and C=2317+/ 146 nmol/16 h, P<0.001). Plasma renin activity was reduced at day 21 (L-NNA=9.6+/ 2.1 and C=25.9+/-5 ng x mL(-1) x h(-1), P<0.05) and remained lower at postpartum day 7 x L-NNA rats exhibited glomerular lesions and tubular atrophy, particularly of connecting tubules that displayed reduced kallikrein staining. Tubulointerstitial infiltrating macrophages (ED1+) were also observed. Renal lesions were present as early as day 11 and persisted at day 7 postpartum. L-NAME rats exhibited similar alterations that were attenuated with an excess of L arginine. We postulate that the reduction in renal kallikrein may contribute to the hemodynamic alterations described in this model. PMID- 10523376 TI - Identification of diadenosine hexaphosphate in human erythrocytes. AB - Diadenosine polyphosphates have been identified as important regulators of vascular tone and blood pressure. In reference to the background of the well known vasoconstriction induced by hemolysate, we questioned whether this action may be due in part to the presence of diadenosine polyphosphates in human erythrocytes. Therefore, erythrocytes were separated from other blood cells and deproteinated. To concentrate and purify nucleotides, the extract was chromatographed by anion exchange, affinity, and reversed-phase columns. In one of the purified fractions, diadenosine hexaphosphate (diadenosine 5', 5'-P(1), P(6) hexaphosphate [AP(6)A]) was identified by matrix-assisted laser desorption/ionization mass spectrometry, ultraviolet spectroscopy, and enzymatic analysis. Hemolysate of erythrocytes injected into the isolated perfused rat kidney induced a dose-dependent vasoconstriction, which was partially inhibited by P(2)-purinoceptor antagonist. The data document the existence of AP(6)A in erythrocytes. AP(6)A may be involved in the pathogenesis of vasospasm induced by free hemoglobin. PMID- 10523377 TI - State-of-the-Art lecture. Role of endothelin-1 in hypertension. AB - Endothelin-1 (ET-1) is overexpressed in the vascular wall in certain models of experimental hypertension: deoxycorticosterone acetate salt-treated rats, deoxycorticosterone acetate salt-treated spontaneously hypertensive rats (SHR), stroke-prone SHR, Dahl salt-sensitive rats, angiotensin II-infused rats, and 1 kidney 1 clip Goldblatt rats; it is not overexpressed in SHR, 2-kidney 1-clip hypertensive rats, or L-NAME-treated rats. In hypertensive rats without generalized vascular overexpression, however, expression of ET-1 was often enhanced in intramyocardial coronary arteries, suggesting a role of endothelin in myocardial ischemia in hypertension. In rats overexpressing ET-1, ET(A/B) and ET(A)-selective receptor antagonists lowered blood pressure and reduced vascular growth, particularly in small arteries, beyond what could be attributed to blood pressure lowering, suggesting a direct effect of ET-1 on growth. Hypertensive rats treated with endothelin antagonists are protected from stroke and renal injury. The ET(A/B) antagonist bosentan induced blood-pressure reductions in mildly hypertensive patients similar to those achieved with an angiotensin converting enzyme inhibitor. Moderately to severely hypertensive patients presented with enhanced expression of prepro-ET-1 mRNA in the endothelium of subcutaneous resistance arteries, suggesting that these stages of hypertension may respond particularly well to endothelin antagonism. Hypertensive patients with coronary artery disease have increased arterial expression of ET-1, and increased plasma levels of immunoreactive endothelin have been described in black patients. ET-1 plays an important role in atherosclerosis, for which hypertension is an important risk factor. Thus, ET-1 may be involved in experimental and human hypertension. Endothelin antagonists may prove effective as disease-modifying agents if they are shown clinically, as they are experimentally, to offer target organ protection and reduce long-term complications of hypertension. This remains to be demonstrated in humans. PMID- 10523378 TI - Enhanced vascular reactivity and Ca2+ entry with low-salt diet: effect of obesity. AB - Salt moderation is often recommended as a nonpharmacological therapy for hypertension, particularly in overweight individuals; however, the effects of low dietary salt on the Ca(2+)-dependent mechanisms of vasoconstriction are unclear. The purpose of this study was to investigate the effect of low salt diet on vascular reactivity and Ca2+ mobilization mechanisms and the modulation of these effects with obesity. Active stress and (45)Ca2+ influx were measured in deendothelialized aortic strips isolated from lean (3.74 kg) and obese (5.51 kg) female rabbits on a normal (0.75%) or low (0.23%) salt (sodium chloride) diet for 18 weeks. Both phenylephrine (Phe, 10(-5) mol/L) and membrane depolarization by 96 mmol/L KCl caused extracellular Ca(2+)-dependent increases in active stress and (45)Ca2+ influx. In lean rabbits, the Phe- and KCl-induced stress and Ca2+ influx were significantly greater with the low-salt versus the normal-salt diet. The Phe-induced Ca2+ influx-stress relationship was significantly greater than that induced by KCl with low-salt diet. In obese rabbits on a normal-salt diet, the Phe- and KCl-induced stress and Ca2+ influx were significantly less than that in lean rabbits but the Ca2+ influx-stress relationship was not significantly altered. Feeding the obese rabbits a low-salt diet was associated not only with significant increases in Phe- and KCl-induced active stress and Ca2+ influx but also with significant enhancement in the Ca2+ influx-stress relationship. In Ca(2+)-free (2 mmol/L EGTA) Krebs solution, stimulation of intracellular Ca2+ release by Phe or caffeine (25 mmol/L) caused a transient contraction that was not significantly different in all groups of rabbits. Thus, with normal salt intake, obesity is associated with a reduction in Ca2+ entry and vascular reactivity. Low-salt diet is associated with an increase in Ca2+ entry and vascular reactivity in both obese and lean rabbits. The enhancement of the Ca2+ influx-stress relationship with low-salt diet, particularly in the obese rabbits, suggests activation of other contractile mechanisms in addition to Ca2+ entry. PMID- 10523379 TI - The aging process modifies the distensibility of elastic but not muscular arteries. AB - Aging decreases the distensibility of large elastic arteries; however, the effects of age on the functional parameters of muscular, medium-sized arteries are not well determined. This study evaluated the consequences of aging on the functional parameters of the carotid and radial arteries in normotensive men. A total of 62 elderly subjects (aged 74+/-2 years) were compared with 87 young subjects (aged 35+/-3 years). Internal diameter and intima-media thickness (IMT) were measured by a high-resolution echo-tracking system to calculate distensibility and incremental elastic modulus (Einc). Although in the normal range, systolic and diastolic blood pressure levels were statistically different in the 2 groups at 128+/-19 and 74+/-13 mm Hg versus 121+/-27 and 71+/-18 mm Hg in the young and elderly subjects, respectively (P<0.05). At the carotid artery level, elderly subjects exhibited a greater IMT (742+/-144 versus 469+/-132 microm; P<0.01) and internal diameter (7067+/-828 versus 6062+/-1026 microm; P<0.01) than young subjects; elderly subjects also had lower distensibility (12+/ 2 versus 21+/-2 kPa(-1) x 10(-3); P<0.01) and higher Einc (0.9+/-0.2 versus 0.7+/ 0.3 kPa x 10(3); P<0.01). At the radial artery level, both IMT (240+/-42 versus 218+/-51 microm; P<0.01) and internal diameter (2685+/-432 versus 2491+/-444 microm; P<0.01) were greater in elderly subjects, but no differences in distensibility and Einc were observed between the 2 groups. All differences remained significant, even after adjusting for mean blood pressure. These results indicate that the increase of the internal diameter and IMT observed during the aging process can have opposite effects on the functional parameters of large elastic or medium-sized muscular arteries. PMID- 10523380 TI - Proteoglycan production by vascular smooth muscle cells from resistance arteries of hypertensive rats. AB - Extracellular matrix (ECM) modifications in the vascular wall contribute to the narrowing of arteries in hypertension. Because direct evidence for the role of proteoglycans (PGs) in the pathological process of resistance-sized arteries has not already been demonstrated, we examined the effect of growth factors on secreted and membrane-bound PG synthesis by cultured mesenteric vascular smooth muscle cells (VSMC) from spontaneously hypertensive rats (SHR) and Wistar rats. After 48 hours of stimulation with angiotensin II (Ang II), platelet-derived growth factor (PDGF-BB), and 10% fetal calf serum (FCS) or 0.1% FCS as control, PG synthesis (in dpm/ng DNA) was evaluated in the medium (M-ECM) and in the cell layer (P-ECM) by a double-isotopic label method with both [(3)H]-glucosamine and [(35)S]-sodium sulfate, which are incorporated into all complex carbohydrates or only into sulfated disaccharides, respectively. VSMC from SHR displayed a significantly lower level of synthesis of M-ECM [(3)H]-PGs than those of Wistar rats in all the experimental groups, including the control group (0. 1% FCS), but no differences in M-ECM [(35)S] uptake were found in any case. In the P-ECM, Ang II was the only factor that produced a lesser effect on [(3)H]-glucosamine and a greater effect on [(35)S]-sodium sulfate uptakes in VSMC from SHR than from Wistar rats. The most prominent change seen in VSMC from SHR was an increased sulfation, assessed by [(35)S]/[(3)H] ratio, in nonstimulated cells and in response to 10% FCS and Ang II but not to PDGF-BB compared with VSMC from Wistar rats. These data indicate the existence of changes in PG modulation in the resistance vessels of SHR, which suggests that PGs may contribute to the development of structural and functional modifications in hypertensive states. PMID- 10523381 TI - Effect of cholecalciferol treatment on the relaxant responses of spontaneously hypertensive rat arteries to acetylcholine. AB - We studied the effect of oral cholecalciferol treatment on the endothelium dependent vascular relaxation and hyperpolarization induced by acetylcholine (ACh), which is impaired in spontaneously hypertensive rats (SHR). Adult female SHR and normotensive Wistar-Kyoto rat (WKY) controls received 125 microg of cholecalciferol per kilogram body weight per day for 6 weeks. The responses to ACh of the isolated mesenteric vascular bed and mesenteric artery rings were measured, as well as the smooth muscle cell membrane potential. After cholecalciferol treatment, the systolic blood pressure and basal perfusion pressure of the mesenteric vascular bed of the SHR fell to control levels. The relaxant and hyperpolarizing effects of ACh, which are reduced in SHR, were also brought to control levels after cholecalciferol treatment. These effects of ACh were inhibited by N(omega)-nitro-L-arginine in SHR and by apamin in WKY. After cholecalciferol treatment, SHR hyperpolarizing responses showed the same inhibition pattern as those of WKY. This indicates that, after cholecalciferol treatment, SHR vascular mesenteric preparation responses to ACh are mediated by endothelium-derived hyperpolarizing factor, which induces activation of Ca(2+) dependent K(+) channels, as in WKY. In untreated SHR, the ACh-mediated response is entirely due to ACh acting via the release of nitric oxide. PMID- 10523382 TI - Effects of aging on serum ionized and cytosolic free calcium: relation to hypertension and diabetes. AB - Elevated cytosolic free calcium (Ca(i)) and reciprocally reduced, extracellular ionized calcium (Ca-ion) levels are observed in both hypertension and non-insulin dependent diabetes mellitus (NIDDM). Because the changes of vascular function and insulin sensitivity in these conditions resemble the changes associated with "normal" aging, we wondered to what extent similar alterations in calcium metabolism occur with aging per se in the absence of overt hypertension or diabetes. We therefore measured platelet Ca(i) levels by spectrofluorometry and serum Ca-ion levels in normotensive, nondiabetic, healthy, normal, elderly (>65 years old) subjects, mean age +/-SEM, 72.2+/-1.5 years old (n=11); in healthy, normal, young (<65 years old) adults, 46.1+/-2.3 years old (n=12); in 10 young adult hypertensives, 48.6+/-1.9 years old; and in 10 normotensive NIDDM subjects, 49.2+/-1.6 years old. Platelet Ca(i) levels were higher (104.5+/-4.9 versus 80.2+/-1.8 nmol/L, P<0.01) and Ca-ion levels lower (1.212+/-0.010 versus 1.236+/ 0.011 mmol/L, P<0.05) in normal elderly compared with young control subjects, but normal elderly Ca(i) and Ca-ion levels were indistinguishable from those in hypertensive (Ca(i) 107.5+/-3.6 nmol/L, Ca-ion 1.210+/-0.009 mmol/L) and NIDDM (Ca(i) 110.7+/-4.7 nmol/L, Ca-ion 1.204+/-0.014 mmol/L) subjects. In normal subjects, significant correlations were found between platelet Ca(i) levels and age (r=0.655, P<0.01) and between Ca(i) levels and systolic blood pressure (r=0.733, P<0.001). We conclude that aging is associated with alterations of Ca(i) and Ca-ion levels resembling those changes present at any age in hypertension and type 2 diabetes. We hypothesize that these alterations of calcium metabolism underlie the predisposition to the alterations of blood pressure and insulin sensitivity characteristic of "normal" aging. The data also suggest that studies of the aging process should be limited to subjects with normal blood pressure and glucose tolerance. PMID- 10523383 TI - Blood pressure and small arteries in DOCA-salt-treated genetically AVP-deficient rats: role of endothelin. AB - Hypertension is associated with structural and mechanical abnormalities of resistance arteries. We have recently reported that vasopressin may be involved in the blood pressure elevation and remodeling of resistance arteries in deoxycorticosterone acetate (DOCA)-salt hypertension, perhaps by modulating vascular endothelin-1 expression. We tested this hypothesis further by examining DOCA-salt hypertension in homozygous vasopressin-deficient Brattleboro (BB) rats in comparison with Long-Evans (LE; control) rats. Mesenteric resistance arteries (lumen <300 microm) were studied on pressurized myographs. After 5 weeks, systolic blood pressure was greater in LE DOCA-salt-treated rats (189+/-5 mm Hg) compared with uniephrectomized (UNx) LE control rats (117+/-4 mm Hg; P<0.01). The increase in blood pressure induced by DOCA-salt treatment was attenuated in vasopressin-deficient rats, such that BB DOCA-salt-treated rats exhibited only a slight elevation of blood pressure (134+/-6 mm Hg) compared with BB-UNx rats (111+/-4 mm Hg; P<0.05). Resistance arteries in LE DOCA-salt-treated rats had a smaller lumen diameter and a larger media width, media cross-sectional area, and media-lumen ratio compared with LE-UNx rats. Isobaric stiffness was unaltered in resistance arteries from LE DOCA-salt-treated rats, despite stiffening of the arterial wall components as indicated by a significant increase in the slope of the media stress-incremental elastic modulus relationship. DOCA-salt treatment in the absence of endogenous vasopressin, ie, in homozygous di/di BB rats, failed to alter vascular structure or wall component stiffness and resulted in a lesser degree of blood pressure elevation. Reverse transcription-polymerase chain reaction analysis revealed that DOCA-salt treatment enhanced endothelin gene expression in LE rats but failed to do so in BB rats. These data indicate that vasopressin plays a critical role in modulating vascular structure and mechanics, as well as blood pressure, in DOCA-salt-induced hypertension. Moreover, these effects of vasopressin are in part mediated by enhancement of endothelin expression. PMID- 10523384 TI - Influence of female sex hormones on endothelium-derived vasoconstrictor prostanoid generation in microvessels of spontaneously hypertensive rats. AB - Although female sex hormones may attenuate endothelial dysfunction in spontaneously hypertensive rats (SHR) by increasing endothelium-derived relaxing factors (EDRFs), the influence of ovarian hormones on the generation of endothelium-derived contracting factors (EDCFs) remains unknown. The aim of this study was to evaluate the effect of estrogen and progesterone on the generation of vasoconstrictor prostanoids and superoxide anion (O2(-)) by microvessels from SHR. Vascular reactivity to norepinephrine (NE), acetylcholine (ACh), and sodium nitroprusside (SNP) were evaluated in the mesenteric arteriolar bed from estrous (OE) and ovariectomized (OVX) SHR. OVX-SHR were treated for 24 hours or 15 days with estradiol and for 15 days with estradiol+progesterone. The vascular reactivity was evaluated in the absence or presence of indomethacin (INDO, 10 micromol/L) and sodium diclofenac (DIC, 10 micromol/L), ridogrel (RID, 50 micromol/L), dazoxiben (DAZ, 10 micromol/L), or superoxide dismutase (SOD, 100 U/mL). Prostanoid levels in the arteriolar perfusate of mesenteries with or without endothelium were measured by enzyme immunoassay. An increased reactivity to NE and reduced sensitivity to ACh were observed in microvessels from OVX-SHR compared with OE-SHR. There were no differences in the responses to SNP. Treatments with estradiol and estradiol+progesterone similarly restored these altered responses. INDO, DIC, RID, and SOD also restored the NE and ACh responses in OVX-SHR. DAZ had no effect on the vascular reactivities. The release of PGF(2alpha), but not of TXB(2) and 6-keto-PGF(1alpha), was greater in OVX-SHR than in OE-SHR microvessels with endothelium when stimulated by NE. This response was normalized by hormonal treatments. Neither NE nor ACh stimulated prostanoid production by microvessels without endothelium. These results suggest that estrogen may protect female SHR against severe hypertension partly by decreasing the synthesis of EDCFs such as PGH(2)/PGF(2alpha) and O2(-). PMID- 10523385 TI - Gender differences in hypertension in spontaneously hypertensive rats: role of androgens and androgen receptor. AB - Males are at greater risk of cardiovascular and renal disease than are females. For example, male spontaneously hypertensive rats (SHR) have higher blood pressures than females. Androgens have been strongly implicated in the hypertension of male SHR, because castration attenuates the hypertension. This study determined whether the androgen receptor plays a role in hypertension in male SHR and whether testosterone alone can cause the hypertension or whether conversion to dihydrotestosterone is necessary. Male SHR, aged 10 weeks, were given the androgen receptor antagonist flutamide (8 mg/kg SC; n=8) or the 5alpha reductase inhibitor finasteride (30 mg x kg(-1) x d(-1) SC; n=11) daily for 5 to 6 weeks. Control rats (n=10) received vehicle (20% benzyl benzoate or ethanol in castor oil). After 5 to 6 weeks, blood pressure (mean arterial pressure) and glomerular filtration rate were measured. Long-term flutamide treatment caused a reduction in mean arterial pressure (control 178+/-5 mm Hg; flutamide 159+/-3 mm Hg; P<0.01), but finasteride had no effect (180+/-5 mm Hg). There were no differences in glomerular filtration rate among the groups. These data indicate that hypertension in male SHR is mediated via the androgen receptor and does not require conversion of testosterone to dihydrotestosterone. PMID- 10523386 TI - Ca(2+)-insensitive vascular protein kinase C during pregnancy and NOS inhibition. AB - Pregnancy-induced hypertension is associated with increased vascular resistance; however, the cellular mechanisms involved are unclear. We have previously found that the relation between Ca(2+) entry and the developed force in vascular smooth muscle is altered during normal pregnancy and in a rat model of pregnancy-induced hypertension produced by long-term treatment with the nitric oxide synthase inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME). The purpose of this study was to investigate whether the pregnancy-associated changes in the vascular Ca(2+) entry-force relation reflect changes in the amount and/or activity of Ca(2+)-insensitive protein kinase C (PKC) isoforms. Active stress and the amount and activity of PKC were measured in deendothelialized aortic strips from nonpregnant and pregnant rats untreated or treated with L-NAME and incubated in Ca(2+)-free (2 mmol/L EGTA) Krebs solution. In nonpregnant rats, the PKC activator phorbol 12,13-dibutyrate (PDBu, 10(-6) mol/L) and the alpha-adrenergic agonist phenylephrine (Phe, 10(-5) mol/L) caused significant, maintained increases in active stress and PKC activity that were inhibited by the PKC inhibitors staurosporine and calphostin C. Western blots in aortic strips of nonpregnant rats revealed the Ca(2+)-insensitive delta-PKC and zeta-PKC isoforms. Both PDBu and Phe caused translocation of delta-PKC from the cytosolic to the particulate fraction. Compared with nonpregnant rats, the amount of delta-PKC and zeta-PKC and the PDBu-stimulated and Phe-stimulated stress, PKC activity and translocation of delta-PKC were significantly reduced in late pregnant rats but significantly enhanced in pregnant rats treated with L-NAME. The PDBu-induced and Phe-induced responses in nonpregnant rats treated with L-NAME were not significantly different from nonpregnant rats, whereas the responses in pregnant rats treated with L-NAME+L-arginine were not significantly different from pregnant rats. These results provide evidence that a signaling pathway in vascular smooth muscle possibly involving the Ca(2+)-insensitive delta-PKC and zeta-PKC isoforms is reduced in late pregnancy and enhanced during long-term inhibition of nitric oxide synthesis. The changes in the amount and activity of vascular PKC isoforms may, in part, explain the changes in vascular resistance during normal pregnancy and pregnancy-induced hypertension. PMID- 10523387 TI - Gender differences in Ca(2+) entry mechanisms of vasoconstriction in Wistar-Kyoto and spontaneously hypertensive rats. AB - We investigated whether putative vascular protection against hypertension in females reflects differences in the Ca(2+) mobilization mechanisms of vasoconstriction depending on the gender and the status of the gonads. Active stress and (45)Ca(2+) influx were measured in aortic strips isolated from intact and gonadectomized male and female Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR). In aortic strips of intact male WKY incubated in normal Krebs' solution (2.5 mmol/L Ca(2+)), both phenylephrine (10(-5) mol/L) and membrane depolarization by 96 mmol/L KCl caused significant increases in active stress and (45)Ca(2+) influx. In intact female WKY, the phenylephrine- and KCl induced stress and (45)Ca(2+) influx were significantly reduced. In Ca(2+)-free (2 mmol/L EGTA) Krebs' solution, stimulation of aortic strips with phenylephrine or caffeine (25 mmol/L) to induce Ca(2+) release from intracellular stores caused a transient increase in stress that was not significantly different between males and females. In SHR, the phenylephrine- and KCl-induced stress and (45)Ca(2+) influx were significantly greater than those in WKY in all groups of rats. The reduction in stress and Ca(2+) entry in intact females compared with intact males was greater in SHR than in WKY. The contractile responses and Ca(2+) entry in castrated male and ovariectomized female WKY or SHR were not significantly different from the respective responses in intact males. The contractile responses and Ca(2+) entry in ovariectomized female WKY or SHR with 17beta estradiol implant were not significantly different from the respective responses in intact females. Thus, the phenylephrine- and depolarization-induced vascular reactivity and Ca(2+) entry in vascular smooth muscle are dependent on gender and on the presence or absence of functional female gonads. Ca(2+) release from intracellular stores is not affected by gender or gonadectomy. The gender specific changes in vascular reactivity and Ca(2+) entry are augmented in hypertension. PMID- 10523388 TI - Increased acetylcholine-induced vasodilation in pregnant rats: A role for gap junctional communication. AB - We have tested the hypothesis that increased gap junctional communication contributes to the augmented endothelium-dependent vasodilation in pregnancy. Contractile force and connexin43 expression were measured in aortic rings from nonpregnant and pregnant rats. Norepinephrine-constricted aortas from pregnant rats were more sensitive to acetylcholine, but not to sodium nitroprusside, compared with those from nonpregnant rats. Vessels from pregnant rats, constricted either with 45 mmol/L KCl or with norepinephrine + 10(-4) mol/L N(G) monomethyl-L-arginine (L-NMMA), an inhibitor of nitric oxide synthase, also exhibited greater relaxation to acetylcholine. Heptanol, an uncoupler of gap junctional communication, inhibited acetylcholine responses in norepinephrine constricted aortas from nonpregnant rats but greatly impaired acetylcholine relaxation in aortas from pregnant rats. Heptanol also inhibited in both groups acetylcholine responses in vessels constricted with KCl, only minimally affected acetylcholine relaxation in arteries constricted with norepinephrine + L-NMMA, and did not change sodium nitroprusside-induced relaxation. Tetraethylammonium chloride induced greater contractions in control aortas compared with aortas from pregnant rats. Increased connexin43 mRNA levels were found in the uterus and in the mesenteric, uterine, and thoracic aortic arteries, but not in the heart and brain, from pregnant rats. These results suggest that increased gap junctional communication, possibly due to increased gap junction protein expression, may facilitate the effects of endothelium-derived relaxing factors, contributing to the augmented endothelium-dependent relaxation in arteries from pregnant rats. PMID- 10523389 TI - State-of-the-Art lecture. Role of angiotensin and oxidative stress in essential hypertension. AB - In this review, we examine the possibility that small increments in angiotensin II are responsible for an increase in blood pressure and maintenance of hypertension through the stimulation of oxidative stress. A low dose of angiotensin II (2 to 10 ng x kg(-1) x min(-1), which does not elicit an immediate pressor response), when given for 7 to 30 days by continuous intravenous infusion, can increase mean arterial pressure by 30 to 40 mm Hg. This slow pressor response to angiotensin is accompanied by the stimulation of oxidative stress, as measured by a significant increase in levels of 8-iso-prostaglandin F(2alpha) (F(2)-isoprostane). Superoxide radicals and nitric oxide can combine chemically to form peroxynitrite, which can then oxidize arachidonic acid to form F(2)-isoprostanes. F(2)-isoprostanes exert potent vasoconstrictor and antinatriuretic effects. Furthermore, angiotensin II can stimulate endothelin production, which also has been shown to stimulate oxidative stress. In this way, a reduction in the concentration of nitric oxide (which is quenched by superoxide) along with the formation of F(2)-isoprostanes and endothelin could potentiate the vasoconstrictor effects of angiotensin II. We hypothesize that these mechanisms, which underlie the development of the slow pressor response to angiotensin II, also participate in the production of hypertension when circulating angiotensin II levels appear normal, as occurs in many cases of essential and renovascular hypertension. PMID- 10523391 TI - Protective effects of captopril against ischemic stress: role of cellular Mg. AB - Magnesium (Mg) deficiency enhances tissue sensitivity to ischemic damage, an effect reversed not only by Mg, but also by sulfhydryl (SH)-containing compounds. We therefore created an in vitro model of red blood cell ischemia to investigate whether the protective effects of these compounds might be related to effects on intracellular free Mg (Mg(i)) content. (31)P-nuclear magnetic resonance (NMR) spectroscopy was used to measure the high-energy metabolites ATP and 2,3 diphosphoglycerate (DPG) and Mg(i) and inorganic phosphate (P(i)) levels in erythrocytes before and for 6 hours after progressive oxygen depletion in the presence or absence of SH-compounds, including captopril, N-acetyl-L-cysteine (NAC), penicillamine, and N-(2-mercaptopropionyl)-glycine (MPG). Under basal aerobic conditions, captopril increased Mg(i) in a dose- and time-dependent fashion (174.5+/-5.3 to 217.1+/-5.1 micromol/L, P<0. 05 at 100 micromol/L, 60 minutes). The SH compounds NAC, penicillamine, and MPG but not the non-SH compound enalaprilat also significantly raised Mg(i) in erythrocytes (P<0.05). With oxygen deprivation, a consistent decrease occurred in both ATP and 2,3-DPG levels associated with a rise in P(i) and in the P(i)/2,3-DPG ratio used as an index of high-energy metabolite depletion. Captopril, compared with control, retarded the rise in P(i) and reduced the P(i)/2,3-DPG ratio (P<0.008 and P<0.025 at 4 and 6 hours, respectively). Furthermore, the higher the initial Mg(i) and the greater the captopril-induced rise in Mg(i), the greater the metabolite protective effect (r=0.799 and r=0.823, respectively; P<0. 01 for both). Altogether, the data suggest that Mg influences the cellular response to ischemia and that the ability of SH compounds such as captopril to ameliorate ischemic injury may at least in part be attributable to the ability of such compounds to increase cytosolic free Mg levels. PMID- 10523390 TI - State-of-the-Art lecture. Antiproliferative actions of angiotensin-(1-7) in vascular smooth muscle. AB - Hemodynamic factors, circulating hormones, paracrine factors, and intracrine factors influence vascular smooth muscle growth and plasticity. The well characterized role of angiotensin II in the modulation of vascular tone and cell function may be critically involved in the mechanisms by which vascular smooth muscle responds to signals associated with vascular endothelial dysfunction and increases in oxidative stress. Studies from this laboratory suggest that the trophic actions of angiotensin II may be intrinsically regulated by angiotensin (1-7), a separate product of the angiotensin system derived from the common substrate, angiotensin I. Exposure of cultured vascular smooth muscle cells to angiotensin-(1-7) inhibited the trophic actions of angiotensin II and reduced the expression of the mitogenic effects of both normal serum and platelet-derived growth factor. The growth-inhibitory actions of angiotensin-(1-7) were blocked by the selective D-alanine(7)-angiotensin-(1-7) antagonist and the nonselective angiotensin receptor blocker sarcosine(1)-threonine(8)-angiotensin II. In contrast, subtype-selective antagonists for the AT(1) and AT(2) receptors had no effect on the inhibitory actions of angiotensin-(1-7), a finding that is consistent with the pharmacological characterization of a high-affinity (125)I labeled angiotensin-(1-7) binding site in the vasculature by use of selective and nonselective angiotensin II receptor antagonists. The relevance of these findings to the proliferative response of vascular smooth muscle cells after endothelial injury was confirmed by assessment of the effect of a 12-day infusion of angiotensin-(1-7) on neointimal formation. In these experiments, the proliferative response produced by injuring the carotid artery was inhibited by angiotensin-(1-7) through a mechanism that could not be explained by changes in arterial pressure. Because plasma angiotensin-(1-7) increased to levels comparable to those found in animals and human subjects given therapeutic doses of angiotensin-converting enzyme inhibitors, angiotensin-(1-7) may be one factor participating in the reversal of vascular proliferation during inhibition of angiotensin II formation or activity. PMID- 10523392 TI - Cardiovascular effects of angiotensin-(1-7) in conscious spontaneously hypertensive rats. AB - In the present study, we reassessed whether angiotensin (Ang)-(1-7) can exert short- and long-term cardiovascular effects because there has been a resurgence of interest in this N-terminal heptapeptide fragment of Ang II. In particular, we studied 3 aspects relating to the reported cardiovascular effects of Ang-(1-7): does this peptide (1) potentiate the hypotensive effect of bradykinin in normotensive Wistar-Kyoto rats and spontaneously hypertensive rats (SHR), (2) cause a depressor effect after long-term treatment in SHR, and (3) contribute to the antihypertensive effects of angiotensin-converting enzyme inhibitors? In the first series of experiments, Ang-(1-7) failed to enhance the dose-related hypotensive responses evoked by bradykinin in SHR (n=11) and Wistar-Kyoto (n=5) rats. In the second series of experiments, a 7-day intravenous infusion of Ang-(1 7) (24 microg x kg(-1) x h(-1)) decreased blood pressure in SHR (n=12) on days 4 and 5, although this effect waned despite continual Ang-(1-7) infusion. However, a new finding was that the Ang-(1-7) antagonist A-779 (24 microg x kg(-1) x h(-1) for 7 days) attenuated the depressor effect of Ang-(1-7) when given concurrently in a separate group of SHR (n=8). In the third series of novel experiments, the angiotensin-converting enzyme inhibitor perindopril was given in drinking water for 7 days (0.3 mg. kg(-1) x day(-1)), either alone (n=6) or combined with an intravenous infusion of A-779 (24 microg x kg(-1) x h(-1) for 7 days, n=8). Although this dose of A-779 attenuated the depressor effect of Ang-(1-7), it did not alter the antihypertensive effect caused by perindopril. Thus, the present results contrast with a number of previous studies and argue against Ang-(1-7) playing a major role in blood pressure regulation. PMID- 10523393 TI - AT(1) receptor antagonism reduces endothelial dysfunction and intimal thickening in atherosclerotic rabbits. AB - The effects of angiotensin (AT)(1) receptor antagonists on functional and morphological alterations associated with atherosclerosis are not well known. The current study was performed to examine the long-term effects of valsartan (3 or 10 mg/kg per day for 10 weeks) on endothelial function and structural changes in aorta from rabbits fed with either a control diet or a cholesterol-enriched diet. Rabbits fed with the cholesterol-rich diet showed higher (P<0.05) plasma levels of cholesterol than did controls. Treatment with valsartan (3 or 10 mg/kg per day) did not alter plasma cholesterol levels or systolic arterial pressure in any group. Contractions induced by angiotensin II were comparable in both control and hypercholesterolemic rabbits and were markedly reduced by treatment with valsartan. Relaxations induced by acetylcholine were lower in hypercholesterolemic rabbits than in controls. Treatment with valsartan (3 or 10 mg/kg per day) enhanced (P<0.05) this response in hypercholesterolemic rabbits but not in controls. Lumen and media cross-sectional areas were comparable in control and hypercholesterolemic rabbits. Vessel area was higher (P<0.05) in hypercholesterolemic rabbits than in controls. Intimal lesion was 29.5+/-6% in cholesterol-fed rabbits and nonexistent in control rabbits. Treatments with 3 and 10 mg/kg per day valsartan reduced (P<0.05) intimal lesion to 2.4+/-0.7% and 2.7+/-0.9%, respectively, and increased lumen area in hypercholesterolemic rabbits. No changes in either vessel or media cross-sectional areas were observed in these animals. In summary, angiotensin II, through AT(1) receptors, appears to play a key role in the development of the vascular functional and structural changes associated with hypercholesterolemia. AT(1) receptor antagonists, besides their antihypertensive effects, could be an important therapeutic tool to reduce the development of atherosclerosis. PMID- 10523394 TI - Ang II-stimulated superoxide production is mediated via phospholipase D in human vascular smooth muscle cells. AB - Intracellular signaling events that mediate the long-term effects of Ang II in vascular smooth muscle cells are unclear, but oxidative stress may play an important role. This study examined the ability of Ang II to generate reactive oxygen species and investigated the putative role of phospholipase D (PLD) dependent signaling pathways for its production in human vascular smooth muscle cells. In addition, we assessed whether redox-sensitive pathways influence Ang II stimulated cell growth. Primary and low-passage cells (passages 1 to 4) derived from resistance arteries of subcutaneous gluteal biopsies from healthy subjects were studied. Oxidative stress was measured with the fluorescent probe 5-(and 6) chloromethyl-2', 7'-dichlorodihydrofluorescein diacetate (CM-H(2)DCFDA) (8 micromol/L), and the role of PLD was assessed with the PLD inhibitor D-erythro sphingosine, dihydro (sphinganine) (10 micromol/L). To determine whether NADH/NADPH oxidase contributes to production of reactive oxygen species, Ang II stimulated cells were pretreated with the specific flavoprotein inhibitor diphenylene iodinium (DPI) (10 micromol/L). DNA and protein synthesis were determined by [(3)H]thymidine and [(3)H]leucine incorporation, respectively. Ang II increased CM-H(2)DCFDA fluorescence, and this was inhibited by catalase (350 U/mL), indicating that the fluorescence signal was derived predominantly from H(2)O(2). Ang II dose-dependently increased H(2)O(2) production (E(max)=57.6+/ 1.7 nmol/L, pD(2)=7.7+/-0.06) and PLD activation (E(max)=207+/-3.3% of control, pD(2)=7.7+/-0.5). H(2)O(2) effects were evident within 1 hour, and maximal PLD activation occurred within 40 minutes after stimulation. DPI inhibited (P<0.01) Ang II-stimulated responses. PLD inhibition significantly attenuated (P<0.05) Ang II-elicited H(2)O(2) production (E(max)=29+/-5 nmol/L). DPI and sphinganine inhibited Ang II-induced DNA and protein synthesis. These data indicate that in vascular smooth muscle cells from human peripheral resistance arteries, Ang II increases H(2)O(2) generation via PLD-dependent, NADH/NADPH oxidase-sensitive pathways. These cascades may function as second messengers in long-term Ang II mediated growth-signaling events. PMID- 10523395 TI - Low-dose angiotensin II increases free isoprostane levels in plasma. AB - Chronic intravenous infusion of subpressor doses of angiotensin II causes blood pressure to increase progressively over the course of several days. The mechanisms underlying this response, however, are poorly understood. Because high dose angiotensin II increases oxidative stress, and some compounds that result from the increased oxidative stress (eg, isoprostanes) produce vasoconstriction and antinatriuresis, we tested the hypothesis that a subpressor dose of angiotensin II also increases oxidative stress, as measured by 8-epi prostaglandin F(2alpha) (isoprostanes), which may contribute to the slow pressor response to angiotensin II. To test this hypothesis, we infused angiotensin II (10 ng/kg per minute for 28 days via an osmotic pump) into 6 conscious normotensive female pigs (30 to 35 kg). We recorded mean arterial pressure continuously with a telemetry system and measured plasma isoprostanes before starting the angiotensin II infusion (baseline) and again after 28 days with an enzyme immunoassay. Angiotensin II infusion significantly increased mean arterial pressure from 121+/-4 to 153+/-7 mm Hg (P<0. 05) without altering total plasma isoprostane levels (180.0+/-24.3 versus 147.0+/-29.2 pg/mL; P=NS). However, the plasma concentrations of free isoprostanes increased significantly, from 38.3+/ 5.8 to 54.7+/-10.4 pg/mL (P<0.05). These results suggest that subpressor doses of angiotensin II increase oxidative stress, as implied by the increased concentration of free isoprostanes, which accompany the elevation in mean arterial pressure elevation. Thus, isoprostane-induced vasoconstriction and antinatriuresis may contribute to the hypertension induced by the slow pressor responses of angiotensin II. PMID- 10523396 TI - State-of-the-Art lecture. Statins and blockers of the renin-angiotensin system: vascular protection beyond their primary mode of action. AB - In addition to their primary mode of action, statins and blockers of the renin angiotensin system possess common additional properties that are under active investigation. The inhibition of cellular proliferation, the restoration of endothelial activity, the inhibition of platelet reactivity, and an antioxidant potential are only a few examples of shared effects that target the arterial wall. These and other properties may eventually become exploited for the improved treatment of cardiovascular diseases and of other diseases apparently unrelated to the cardiovascular field, including inflammation and cancer. This review analyzes the current knowledge on the pleiotropic properties of these classes of drugs. Direct comparison indicates that study of the associations among these drugs may eventually disclose additive or synergistic effects that, perhaps even at lower dosages, may provide improved vascular protection and a strong alliance against several atherogenic mechanisms. PMID- 10523397 TI - Influence of verapamil and diclofenac on leukocyte migration in rats. AB - Nonsteroidal anti-inflammatory drugs and calcium channel blockers can reduce inflammatory responses. Leukocytes play an important role in these responses. An increased expression of adhesion molecules may increase leukocyte migration. Verapamil and diclofenac are known to reduce leukocyte-endothelium interaction. To investigate a possible synergism between these drugs that could be beneficial in cardiovascular diseases, we studied leukocyte behavior by using intravital microscopy. Venules of the spermatic fascia of anesthetized Wistar rats were observed with a closed-circuit TV coupled to an optical microscope. The number of leukocytes rolling along the venular endothelium ("rollers"), sticking after application of a stimulus such as leukotriene B(4) or zymozan-activated plasma ("stickers"), or migrating after a carrageenan stimulus was reduced by verapamil at the dose of 10 mg/kg IP and by diclofenac at the dose of 2.5 mg/kg IP. The combination of both did not augment the effect of each agent alone. Verapamil, diclofenac, or their combination did not interfere with vessel diameter, number of circulating leukocytes, blood pressure levels, or heart rate. Verapamil alone or together with diclofenac reduced venular blood flow velocity and in consequence, the venular shear rate. Our data allow us to suggest that these drugs might interfere with the expression of adhesion cell molecules to reduce cell migration in inflammation. The lack of synergism between the drugs might be explained by the reduction in venular shear rate induced by verapamil, which might not be sufficient to hinder the effect of verapamil alone but hindered the summation effects of both. PMID- 10523398 TI - Effects of vitamin E and glutathione on glucose metabolism: role of magnesium. AB - Vitamin E is an antioxidant that has been demonstrated to improve insulin action. Glutathione, another natural antioxidant, may also be important in blood pressure and glucose homeostasis, consistent with the involvement of free radicals in both essential hypertension and diabetes mellitus. Our group has recently suggested that the effects of reduced glutathione on glucose metabolism may be mediated, at least in part, by intracellular magnesium levels (Mg([i])). Recent evidence suggests that vitamin E enhances glutathione levels and may play a protective role in magnesium deficiency-induced cardiac lesions. To directly investigate the effects of vitamin E supplementation on insulin sensitivity in hypertension, in relation to the effects on circulating levels of reduced (GSH) and oxidized (GSSG) glutathione and on Mg([i]), we performed a 4-week, double-blind, randomized study of vitamin E administration (600 mg/d) versus placebo in 24 hypertensive patients and measured whole-body glucose disposal (WBGD) by euglycemic glucose clamp, GSH/GSSG ratios, and Mg([i]) before and after intervention. The relationships among WBGD, GSH/GSSG, and Mg([i]) in both groups were evaluated. In hypertensive subjects, vitamin E administration significantly increased WBGD (25.56+/-0.61 to 31.75+/-0.53 micromol/kg of fat-free mass per minute; P<0.01), GSH/GSSG ratio (1.10+/-0.07 to 1.65+/-0.11; P<0.01), and Mg([i]) (1.71+/-0.042 to 1.99+/-0.049 mmol/L; P<0.01). In basal conditions, WBGD was significantly related to both GSH/GSSG ratios (r=0.58, P=0.047) and Mg([i]) (r=0.78, P=0.003). These data show a clinical link between vitamin E administration, cellular magnesium, GSH/GSSG ratio, and tissue glucose metabolism. Further studies are needed to explore the cellular mechanism(s) of this association. PMID- 10523399 TI - Potential role of glycerol leading to rat fructose hypertension. AB - A fructose-enriched diet promotes hypertension in rats. We thought that an enhancement of the glycolytic and/or lipid disorder (s) that raise blood pressure could be the cause. Therefore, we studied 4 groups of Sprague-Dawley rats (+/-200 g): (1) control rats received a standard diet and tap water; (2) the glycerol group of rats received a standard diet and 0.54 mol/L glycerol in tap water; (3) the fructose group was given a fructose-enhanced diet (chow had 55% fructose instead of dextrin) and tap water; and (4) the fructose-glycerol group was given the fructose-enhanced diet and 0. 54 mol/L glycerol in drinking water. At the end of the second week, the findings were as follows. Blood pressure was 149+/-2 mm Hg in the fructose-glycerol group versus 129+/-2 (P<0.001), 131+/-2 (P<0. 001), and 140+/-3 (P<0.005) mm Hg in the control, glycerol, and fructose groups, respectively. Insulinemia was higher in the fructose-glycerol group than the control (P<0.001), glycerol (P<0. 001), and fructose groups (P<0.001); triglyceridemia was higher in the fructose-glycerol (P<0.02), fructose (P<0.05), and glycerol groups (P<0.02) than the control group. Thoracic aorta rings showed a lower ED(50) to 12,13-phorbol dibutyrate in the fructose-glycerol group than in the control (P<0.001), glycerol (P<0.002), and fructose groups (P<0.001). In conclusion, glycerol-fructose administration resulted in hypertriglyceridemia, hyperinsulinemia, and increased vascular sensitivity to 12,13-phorbol dibutyrate (with respect to the control group), and significantly greater expression of protein kinase C alpha and betaII (with respect to the glycerol group). PMID- 10523400 TI - Cardiovascular effects of clonidine-like drugs in pithed rabbits. AB - Administration (3 to 100 microg/kg IV) of clonidine, rilmenidine, and an imidazoline derivative, 2-(2-chlorophenylamino)imidazoline, in pithed nonstimulated rabbits caused a dose-dependent increase in mean arterial pressure without affecting heart rate. Prazosin (0.1 mg/kg IV) almost abolished the pressor responses to 2-(2-chlorophenylamino)imidazoline, partially inhibited those induced by clonidine, but failed to affect those elicited by rilmenidine. In contrast, yohimbine (1 mg/kg IV) blunted the pressor responses of the 3 drugs. In sympathetically stimulated pithed rabbits, 2-(2-chlorophenylamino)imidazoline induced only pressor effects, whereas clonidine and rilmenidine caused a transient pressure increase followed by a dose-dependent depressor effect. Yohimbine abolished the depressor effect of both drugs, which may have involved presynaptic alpha(2)-adrenoceptors. In conclusion, peripheral effects of 2-(2 chlorophenylamino)imidazoline and clonidine involved at least alpha(1)- and alpha(2)-adrenoceptor activation, whereas pressor and depressor effects of rilmenidine were mediated by alpha(2)-adrenoceptors. PMID- 10523401 TI - Administration time-dependent effects of aspirin in women at differing risk for preeclampsia. AB - This study extends previous results on the effects of low-dose aspirin on blood pressure in pregnant women at differing risk of developing hypertension in pregnancy and who received aspirin at different times according to their rest activity cycle. A double-blind, randomized, placebo-controlled trial was conducted in 240 pregnant women randomly assigned to 1 of 6 groups according to treatment (placebo or aspirin, 100 mg/d, starting at 12 to 16 weeks of gestation) and the time of treatment: on awakening (time 1), 8 hours after awakening (time 2), or before bedtime (time 3). Blood pressure and heart rate for each subject were automatically monitored for 2 days every 4 weeks from the day of recruitment until delivery, as well as at puerperium (6 to 8 weeks after delivery). Subjects were further divided for comparative purposes according to the results of the tolerance-hyperbaric test for early identification of those with a higher risk for developing hypertensive complications in pregnancy. Results indicated that there was no effect of aspirin on blood pressure at time 1 (compared with placebo). A blood pressure reduction was, however, highly statistically significant at time 2 and, to a greater extent, at time 3 (mean reductions of 14.2 and 9.6 mm Hg in 24-hour means for systolic and diastolic blood pressure, respectively, at the time of delivery compared with placebo given at the same time). Effects of aspirin on blood pressure were significantly larger for women with a positive test at the time of recruitment (P<0.001). Differences in blood pressure among pregnant women receiving aspirin at different times in the circadian cycle disappeared at puerperium (P>0.212). There was no effect of aspirin or placebo on heart rate. This study corroborates the statistically significant, time-dependent effect of low-dose aspirin on blood pressure in pregnant women with differing risk of developing hypertensive complications in pregnancy. Although the mechanism involved in the administration-time-dependent responsiveness of blood pressure to aspirin still remains uncertain, the use of doses of aspirin <80 mg/d that do not affect placental thromboxane, initiation of the use of aspirin after 16 weeks' gestation, and the lack of circadian timing for aspirin administration could all explain the lack of positive results in previous clinical trials. PMID- 10523402 TI - Salivary carbonic anhydrase isoenzyme VI. AB - The carbonic anhydrases (CAs) participate in the maintenance of pH homeostasis in various tissues and biological fluids of the human body by catalysing the reversible reaction CO2 + H2O HCO3- + H+ (Davenport & Fisher, 1938; Davenport, 1939; Maren, 1967). Carbonic anhydrase isoenzyme VI (CA VI) is the only secretory isoenzyme of the mammalian CA gene family. It is exclusively expressed in the serous acinar cells of the parotid and submandibular glands, from where it is secreted into the saliva. In this review, we will discuss recent advances in research focused on the physiological role of salivary CA VI in the oral cavity and upper alimentary canal. PMID- 10523403 TI - Effects of mutations causing hypokalaemic periodic paralysis on the skeletal muscle L-type Ca2+ channel expressed in Xenopus laevis oocytes. AB - 1. A truncated form of the rabbit alpha1S Ca2+ channel subunit (alpha1SDeltaC) was expressed with the beta1b, alpha2delta and gamma auxiliary subunits in Xenopus laevis oocytes. After 5-7 days, skeletal muscle L-type currents were measured (469 +/- 48 nA in 10 mM Ba2+). All three of the auxiliary subunits were necessary to record significant L-type current. A rapidly inactivating, dihydropyridine-insensitive endogenous Ba2+ current was observed in oocytes expressing the auxiliary subunits without an exogenous alpha subunit. Expression of full-length alpha1S gave 10-fold smaller currents than the truncated form. 2. Three missense mutations causing hypokalaemic periodic paralysis (R528H in domain II S4 of the alpha1S subunit; R1239H and R1239G in domain IV S4) were introduced into alpha1SDeltaC and expressed in oocytes. L-type current was separated from the endogenous current by nimodipine subtraction. All three of the mutations reduced L-type current amplitude ( approximately 40 % for R528H, approximately 60 70 % for R1239H and R1239G). 3. The disease mutations altered the activation properties of L-type current. R528H shifted the G(V) curve approximately 5 mV to the left and modestly reduced the voltage dependence of the activation time constant, tauact. R1239H and R1239G shifted the G(V) curve approximately 5-10 mV to the right and dramatically slowed tauact at depolarized test potentials. 4. The voltage dependence of steady-state inactivation was not significantly altered by any of the disease mutations. 5. Wild-type and mutant L-type currents were also measured in the presence of (-)-Bay K8644, which boosted the amplitude approximately 5- to 7-fold. The effects of the mutations on the position of the G(V) curve and the voltage dependence of tauact were essentially the same as in the absence of agonist. Bay K-enhanced tail currents were slowed by R528H and accelerated by R1239H and R1239G. 6. We conclude that the domain IV mutations R1239H and R1239G have similar effects on the gating properties of the skeletal muscle L-type Ca2+ channel expressed in Xenopus oocytes, while the domain II mutation R528H has distinct effects. This result implies that the location of the substitutions is more important than their degree of conservation in determining their biophysical consequences. PMID- 10523404 TI - Amino acid substitutions in the pore of rat glutamate receptors at sites influencing block by polyamines. AB - 1. The effect on polyamine block of mutations at the Q/R site and the conserved negative charge +4 site in AMPA and kainate receptors was studied using the rat glutamate receptor GluR6 expressed in Xenopus oocytes and human embryonic kidney (HEK) cells. 2. Introduction of negative charge at the Q/R site increased the equilibrium dissociation constant at 0 mV (Kd(0)) for spermine from 1.3 to 4.0 microM (Q590E); the smaller side chains Q590D and Q590N had Kd(0) values of 47 and 20 microM. Reductions in spermine affinity were also obtained for the small hydrophobic residues Q590V and Q590A, with Kd(0) values of 3.6 and 8.8 microM. Positively charged side chains produced outward rectifying responses similar to those recorded for GluR6(Q) with polyamine-free conditions, suggesting a complete absence of voltage-dependent block by spermine. 3. Substitution of tryptophan at the Q/R site produced high-affinity block with a Kd(0) of 190 pM. In Xenopus oocytes no outward current was observed at potentials up to +200 mV. A much smaller increase in affinity was observed for Q590F and Q590Y, which had Kd(0) values of 0.28 and 0.83 microM respectively. 4. The Q590H mutant gave weakly birectifying responses strikingly different from those for other mutants. When ionization of the His group was increased by raising the external hydrogen ion concentration, responses became outward rectifying. The ratios of the conductance at 100 mV over that at -100 mV for Q590H were 0.52 at pH 8.3 and 2.5 at pH 5.3. 5. Neutralization of charge or aromatic residues at the +4 site produced a large reduction of spermine affinity, with Kd(0) values for E594N, E594Q and E594W of 109, 1020 and 2150 microM, respectively. In the absence of polyamines, E594K and E594R produced strongly inward rectifying responses while E594Q, E594A and E594W were birectifying. 6. A model for permeant block allowed quantitative comparisons between mutants. Despite large changes in well depth and barrier heights, there was little change in the voltage dependence of block for both Q/R and +4 site mutants. We propose a model with a distributed binding site for polyamines in which the +4 site is located near the entrance to the channel. PMID- 10523405 TI - Functional expression of tagged human Na+-glucose cotransporter in Xenopus laevis oocytes. AB - 1. High-affinity, secondary active transport of glucose in the intestine and kidney is mediated by an integral membrane protein named SGLT1 (sodium glucose cotransporter). Though basic properties of the transporter are now defined, many questions regarding the structure- function relationship of the protein, its biosynthesis and targeting remain unanswered. In order to better address these questions, we produced a functional hSGLT1 protein (from human) containing a reporter tag. 2. Six constructs, made from three tags (myc, haemaglutinin and poly-His) inserted at both the C- and N-terminal positions, were thus tested using the Xenopus oocyte expression system via electrophysiology and immunohistochemistry. Of these, only the hSGLT1 construct with the myc tag inserted at the N-terminal position proved to be of interest, all other constructs showing no or little transport activity. A systematic comparison of transport properties was therefore performed between the myc-tagged and the untagged hSGLT1 proteins. 3. On the basis of both steady-state (affinities for substrate (glucose) and inhibitor (phlorizin) as well as expression levels) and presteady-state parameters (transient currents) we conclude that the two proteins are functionally indistinguishable, at least under these criteria. Immunological detection confirmed the appropriate targeting of the tagged protein to the plasma membrane of the oocyte with the epitope located at the extracellular side. 4. The myc-tagged hSGLT1 was also successfully expressed in polarized MDCK cells. alpha Methylglucose uptake studies on transfected cells showed an exclusively apical uptake pathway, thus indicating that the expressed protein was correctly targeted to the apical domain of the cell. 5. These comparative studies demonstrate that the myc epitope inserted at the N-terminus of hSGLT1 produces a fully functional protein while other epitopes of similar size inserted at either end of the protein inactivated the final protein. PMID- 10523406 TI - Expression and polarized distribution of an inwardly rectifying K+ channel, Kir4.1, in rat retinal pigment epithelium. AB - 1. In the eye, different substances and ions including potassium (K+) are transported between neural retina and choroid via the subretinal space. Inwardly rectifying K+ channels (Kir) on the apical membrane of retinal pigment epithelial (RPE) cells are thought to play an essential role in K+ transport in the subretinal space. 2. Single-channel recordings from the apical membrane of RPE cells exhibited functional expression of a Kir channel with properties identical to those of Kir4.1, while recordings from the basolateral membrane showed no detectable Kir channel currents. 3. The expression of Kir4.1 mRNA in RPE cells was confirmed by RT-PCR analysis and in situ hybridization. Furthermore, using immunohistochemistry, we found that Kir4.1 was prominently expressed in RPE cells and localized specifically on the processes on their apical membrane. 4. Developmental studies revealed that expression of Kir4.1 started to appear 10 days or later after birth in RPE cells, in parallel with the maturation of retinal neuronal activity as represented by the a- and b-waves of the electroretinogram. 5. These data suggest that Kir4.1 is one of the Kir channels involved in RPE-mediated control of K+ ions in the subretinal space. PMID- 10523408 TI - C-terminal interactions modulate the affinity of GLAST glutamate transporters in salamander retinal glial cells. AB - 1. Proteins that interact with the intracellular carboxy termini of neurotransmitter- and voltage-gated ion channels are known to control the subcellular localization of the channels, localize other proteins near those channels, and modulate channel activity. By contrast, little is known about the control of neurotransmitter transporter function by interacting proteins. 2. To competitively disrupt interactions of the C- and N-termini of the GLAST glutamate transporter with other proteins, we dialysed whole-cell patch-clamped retinal glia with peptides identical to the eight amino acids at the C- or N-termini of the transporter, and compared the effect on transporter-mediated currents with dialysis of scrambled versions of the same peptides. 3. Dialysis with the N terminus peptide had no effect on the maximum glutamate-evoked current nor on the glutamate affinity of the transporter. Dialysis with the C-terminus peptide had no effect on the maximum current, but increased the affinity of the transporter for glutamate (compared with scrambled C-terminus peptide, and with N- and scrambled N-terminus peptides: Km decreased from 16 to 11 microM)). 4. These data suggest that disruption of an interaction between an intracellular protein and the last eight amino acids of the GLAST C-terminus, which have some similarity to the PDZ binding domain of ion channel C-termini, increases the glutamate affinity of GLAST. Thus, the interacting protein decreases the affinity of GLAST transporters. 5. Removing the GLAST C-terminus interaction increases the transporter current by 40 % at low glutamate concentrations. Thus, this interaction may significantly slow the removal of low concentrations of glutamate from the extracellular space, and affect the kinetics of retinal cell light responses. PMID- 10523407 TI - Functional MRI localisation of central nervous system regions associated with volitional inspiration in humans. AB - 1. Functional magnetic resonance imaging (fMRI) provides a means of studying neuronal circuits that control respiratory muscles in humans with better spatial and temporal resolution than in previous positron emission tomography (PET) studies. 2. Whole brain blood oxygenation level-dependent (BOLD) changes determined by fMRI were used to identify areas of neuronal activation associated with volitional inspiration in five healthy men. Four series of scans of each subject were acquired during voluntary breathing (active task) and mechanical ventilation (passive task). Ventilation and end-tidal PCO2 were similar between tasks. Scan data were re-aligned to correct for movement artefacts and cross referenced breath by breath to respiratory data for selective averaging of inspiratory and expiratory images. 3. Group analysis identified significant increases in the fMRI signal with volitional inspiration in the superior motor cortex, premotor cortex and supplementary motor area at loci similar to those detected in earlier studies that used PET. Additional regions activated by volitional inspiration included inferolateral sensorimotor cortex, prefrontal cortex and striatum (these foci were only revealed by PET under significant inspiratory load). 4. This study represents the first synchronised breath-by breath analysis of respiratory-related neuronal activity with whole brain imaging in humans. Temporal resolution is sufficient to distinguish individual breaths at a normal breathing frequency. PMID- 10523409 TI - Induction of cyclooxygenase-2 expression in human myometrial smooth muscle cells by interleukin-1beta: involvement of p38 mitogen-activated protein kinase. AB - 1. Human myometrial smooth muscle cells (HMSMCs) in culture were exposed to recombinant human interleukin-1beta (IL-1beta, 10 ng ml-1) for 1 to 24 h. Cyclooxygenase-2 (COX-2) mRNA and protein were rapidly induced, with expression sustained at 24 h. 2. Cycloheximide (10 microg ml-1, 6 h) blocked IL-1beta induced COX-2 protein expression and super-induced COX-2 mRNA expression. Induction of COX-2 mRNA and protein was blocked by dexamethasone (1 microm, 6 h). 3. IL-1beta-induced COX-2 expression was accompanied by a 3-fold increase of prostaglandin E2 release into the culture medium. 4. IL-1beta induced a transient (5-30 min) activation of p42/44 and p38 mitogen-activated protein kinase (MAPK) enzymes in HMSMCs. Activity of p38 MAPK was monitored by in-gel activity of its substrate MAP kinase-activated protein kinase-2 (MAPKAP kinase-2). Induction of MAPKAP kinase-2 activity was prevented by the p38 MAPK inhibitor SB 203580 (10 microm, 5-30 min). 5. COX-2 protein expression detected after 6 h IL-1beta stimulation was blocked by SB 203580 (10 microM). Exposure of HMSMCs to 10 ng ml 1 IL-1beta for only 30 min induced a level of COX-2 protein expression at 6 h culture similar to that detected in cells exposed to the cytokine for 6 h. 6. Exposure of cells to SB 203580 (10 microM during only the first 30 min of IL 1beta stimulation was effective in blocking COX-2 protein expression assayed after 6 h in culture. 7. This study has established that a transient activation of the p38 MAPK cascade is involved in IL-1beta-stimulated COX-2 expression in human myometrial smooth muscle cells. Induction of COX-2 by IL-1beta in HMSMCs provides support for the hypothesis that autocrine prostaglandin signalling in the myometrium, initiated by elevated intrauterine cytokine concentrations, plays a role in regulating myometrial contractility during labour. PMID- 10523410 TI - On the characterisation of the mechanism underlying passive activation of the Ca2+ release-activated Ca2+ current ICRAC in rat basophilic leukaemia cells. AB - 1. Tight-seal whole-cell patch clamp experiments were performed to investigate the mechanism whereby passive depletion of stores activates the Ca2+ release activated Ca2+ current (ICRAC) in rat basophilic leukaemia (RBL) cells. 2. Passive depletion of stores was achieved by dialysing cells with different concentrations of Ca2+ chelators. Low concentrations generally evoked a submaximal ICRAC, which developed slowly and monophasically. Higher concentrations resulted in a biphasic current in which the initial slow monophasic component developed into a faster and bigger second phase. 3. The kinetics of ICRAC as well as its final amplitude were not affected by Ca2+ chelators that had different affinities or speeds of binding. 4. Exogenous Ca2+ binding ratios > or = 16,670 were necessary to fully activate ICRAC. Because the Ca2+ binding ratio within the stores is presumably low, this indicates that other factors like Ca2+ transport across the stores membrane are rate limiting for passive store depletion. 5. Heparin and Ruthenium Red both failed to affect passive Ca2+ leak from the intracellular stores. 6. Treatment with sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) pump blockers dramatically altered the kinetics of activation of biphasic currents, and increased the amplitude of monophasic ones. 7. Our results suggest that SERCA pumps are very effective in preventing ICRAC from activating passively, and are responsible for the phasic nature of the current, its time course of development and its overall extent. PMID- 10523411 TI - Intracellular calcium signals measured with indo-1 in isolated skeletal muscle fibres from control and mdx mice. AB - 1. Intracellular free calcium concentration ([Ca2+]i) was measured with the fluorescent indicator indo-1 in single skeletal fibres enzymatically isolated from the flexor digitorum brevis and interosseus muscles of control and dystrophic mdx C57BL/10 mice. Measurements were taken from a portion of fibre that was voltage clamped to allow detection of depolarization-induced changes in [Ca2+]i. 2. The mean (+/- s.e.m.) initial resting [Ca2+]i from all control and mdx fibres tested was 56 +/- 5 nM (n = 72) and 48 +/- 7 nM (n = 57), respectively, indicating no significant overall difference between the two groups. However, when comparing a batch of control and mdx fibres obtained from mice older than approximately 35 weeks, resting [Ca2+]i was significantly lower in mdx (16 +/- 4 nM, n = 11) than in control fibres (71 +/- 10 nM, n = 14). 3. Changes in [Ca2+]i elicited by short (5-35 ms) depolarizing pulses from -80 to 0 mV showed similar properties in control and mdx fibres. After a 5 ms duration pulse the mean time constant of [Ca2+]i decay was, however, significantly elevated in mdx as compared to control fibres, by a factor of 1.5-2. For longer pulses, no significant difference could be detected. 4. In response to 50 ms duration depolarizing pulses of various amplitudes the threshold for detection of an [Ca2+]i change and the peak [Ca2+]i reached for a given potential were similar in control and mdx fibres. 5. Overall results show that mdx skeletal muscle fibres are quite capable of handling [Ca2+]i at rest and in response to membrane depolarizations. PMID- 10523412 TI - Local regulation of the threshold for calcium sparks in rat ventricular myocytes: role of sodium-calcium exchange. AB - 1. To determine whether Na+-Ca2+ exchange modulates Ca2+ sparks, we studied enzymatically isolated patch clamped rat ventricular myocytes loaded with the Ca2+-sensitive indicator fluo-3, using confocal microscopy at 20-22 C. Two dimensional images of Ca2+ sparks were recorded at 240 Hz using a laser scanning confocal microscope, allowing observation of a large area of the cell (820 microm2) at one time. 2. At a holding potential of -75 mV, spontaneous sparks were infrequent. Removal of extracellular Na+ for 520 ms, which in the absence of pipette Na+ should block Na+-Ca2+ exchange bidirectionally, was associated with a fourfold increase in spark frequency, without a significant change in cytoplasmic [Ca2+], sarcoplasmic reticulum (SR) Ca2+ content, or spark intensity, size or time course. 3. These findings are consistent with a model of excitation contraction coupling in which Na+-Ca2+ exchange locally regulates the resting Ca2+ concentration in the diadic cleft (T-tubule-SR junction), thereby modulating the threshold for triggering Ca2+ sparks. PMID- 10523413 TI - Protein kinase C regulation of K+ currents in rat ventricular myocytes and its modification by hormonal status. AB - 1. The effects of protein kinase C (PKC) activation on cardiac K+ currents were studied in rat ventricular myocytes, using whole-cell voltage clamp methods. Control rats were compared to hypothyroid or diabetic rats, in which PKC expression and activity were enhanced. 2. In control myocytes, two calcium independent outward K+ currents, the transient It and the sustained Iss, were attenuated by 18.9 +/- 2.0 and 16.8 +/- 3.5 %, respectively (mean +/- s.e.m.), following addition of a synthetic analogue of diacylglycerol, DiC8 (20 microM). In myocytes from hypothyroid or diabetic rats, It and Iss were not affected by DiC8. 3. The effects of DiC8 were restored in myocytes from thyroidectomized rats by injection of physiological doses of tri-iodothyronine (T3; 10 microg kg-1 for 6-8 days). Incubating cells from diabetic rats with 100 nM insulin for 5-9 h also restored the ability of DiC8 to attenuate It and Iss. 4. The attenuation of K+ currents by DiC8 in control cells was absent in the presence of a peptide known to inhibit the translocation of the isoform PKCepsilon (EAVSKPLT, 24 microM introduced through the recording pipette). A scrambled peptide (LSETKPAV) was without effect. 5. Under hypothyroid conditions the inhibitory peptide restored the effects of DiC8 on It and Iss. These currents were attenuated by 11.9 +/- 1. 5 and 9.8 +/- 1.5 %, respectively, which was significantly (P < 0. 001) more than without the peptide or with the scrambled peptide. 6. These results show that the PKC-mediated suppression of cardiac K+ currents is normally mediated by PKCepsilon translocation. This effect is absent under hypothyroid and diabetic conditions, presumably due to prior PKC activation and translocation. A PKCepsilon translocation inhibitor restores the ability of DiC8 to attenuate K+ currents under hypothyroid conditions. This presumably reflects a (partial) reversal of a chronic translocation and a shift in the balance between PKC and its anchoring proteins. PMID- 10523414 TI - Nitric oxide increases persistent sodium current in rat hippocampal neurons. AB - 1. The effects of nitric oxide (NO) donors on whole-cell, TTX-sensitive sodium currents and single sodium channels in excised patches were examined in rat hippocampal neurons. The whole-cell sodium current consisted of a large transient component (INa,t) and a smaller, inactivation-resistant, persistent component (INa,p). 2. In acutely dissociated neurons, the amplitude of the whole-cell INa, p increased by 60-80 % within a few minutes of exposure to either of two NO donors, sodium nitroprusside (SNP, 100 microM) or S-nitroso-N-acetyl-DL penicillamine (SNAP, 100 microM). 3. The amplitude of INa,t was not changed significantly by the same concentrations of SNP and SNAP, indicating that NO had a selective effect on INa,p. 4. Both NO donors significantly increased the mean persistent current in excised inside-out patches from cultured hippocampal neurons. SNP at 10-100 microM increased average mean persistent current at a pipette potential (Vp) of +30 mV from -0.010 +/- 0.014 pA (control) to -2.91 +/- 1.41 pA (n = 10). SNAP at 3-100 microM increased the average mean inward current in six inside-out patches from -0.07 +/- 0.02 to -0.30 +/- 0.08 pA (Vp = +30 mV). 5. The increase in persistent Na+ channel activity recorded in inside-out patches in the presence of SNP or SNAP could be reversed by the reducing agent dithiothreitol (DTT, 2-5 mM) or by lidocaine (1-10 microM). 6. The average mean current recorded in the presence of SNP was 10-fold higher than that elicited by SNAP. The time delay before an increase was observed was shorter with SNP (4.0 +/ 0.8 min, n = 8) than with SNAP (8.4 +/- 1.6 min, n = 7). 7. A component of the SNP molecule added on its own, 5 mM sodium cyanide (NaCN), increased mean current in excised inside-out patches (Vp = +30 mV) from -0.06 +/- 0.04 to -0.58 +/- 0.21 pA (n = 19). This increase in channel activity could be blocked by 10 microM lidocaine and 2-5 mM DTT. 8. These results suggest that NO may directly increase the activity of neuronal persistent Na+ channels, but not transient Na+ channels, through an oxidizing action directly on the channel protein or on a closely associated regulatory protein in the plasma membrane. PMID- 10523416 TI - Effects of calcium buffering on glucose-induced insulin release in mouse pancreatic islets: an approximation to the calcium sensor. AB - 1. The properties of the calcium sensor for glucose-induced insulin secretion have been studied using cell-permeant Ca2+ buffers with distinct kinetics and affinities. In addition, submembrane cytosolic Ca2+ distribution has been modelled after trains of glucose-induced action potential-like depolarizations. 2. Slow Ca2+ buffers (around 1 mmol l-1 intracellular concentration) with different affinities (EGTA and Calcium Orange-5N) did not significantly affect glucose-induced insulin release. Modelling showed no effect on cytosolic Ca2+ concentrations at the outermost shell (0.05 microm), their effects being observed in the innermost shells dependent on Ca2+ affinity. 3. In contrast, fast Ca2+ buffers (around 1 mmol l-1 intracellular concentration) with different affinities (BAPTA and Calcium Green-5N) caused a 50 % inhibition of early insulin response and completely blocked the late phase of glucose-induced insulin response, their simulations showing a decrease of [Ca2+]i at both the inner and outermost shells. 4. These data are consistent with the existence in pancreatic beta-cells of a higher affinity Ca2+ sensor than that proposed for neurons. Moreover, these data are consistent with the proposed existence of two distinct pools of granules: (i) 'primed' vesicles, colocalized with Ca2+ channels and responsible of the first phase of insulin release; and (ii) 'reserved pool' vesicles, not colocalized and responsible for the second phase. PMID- 10523415 TI - In vitro motility speed of slow myosin extracted from single soleus fibres from young and old rats. AB - 1. Isolated soleus muscle fibres from aged rats contract more slowly than those from young rats. To determine whether this effect is due to a difference between the myosin molecules, we measured the rate at which actin filaments are driven over a myosin coated surface in the presence of ATP by using a novel in vitro motility assay where myosin is extracted from single muscle fibre segments. 2. Motility was dependent on the myosin density on the coverslip. In regions of high myosin density, actin motility was orientated parallel and anti-parallel to the direction of flow during myosin adhesion to the coverslip. In contrast, in regions of lower myosin density, actin motility was more random. The speed was about 20 % higher in the high density regions (P < 0.001). Further, the speed of filaments in the high density region, moving away or towards the fibre was less variable (P < 0.05) than that of more randomly moving filaments in the low density region. 3. The speed with myosin from slow soleus fibres of young adult rats (3-6 months old; v = 1.43 +/- 0.23 microm s-1; mean +/- s.d.) was faster (P < 0.001) than with myosin from aged rats (20-24 months old; v = 1.27 +/- 0.23 microm s-1). 4. No difference in myosin isoforms between young adult and aged fibres could be detected using electrophoretic and immunocytochemical techniques. Fibres of both ages expressed the beta/slow myosin heavy chain (MyHC) isoform and slow isoforms of essential and regulatory myosin light chains (MyLCs). 5. It is concluded that an age-related alteration in myosin contributes to the slowing of the maximum shortening velocity (V0) observed in soleus muscle fibres expressing the beta/slow MyHC isoform. PMID- 10523417 TI - Calcium dynamics and buffering in motoneurones of the mouse spinal cord. AB - 1. A quantitative analysis of endogenous calcium homeostasis was performed on 65 motoneurones in slices of the lumbar spinal cord from 2- to 8-day-old mice by simultaneous patch-clamp and microfluorometric calcium measurements. 2. Somatic calcium concentrations were monitored with a temporal resolution in the millisecond time domain. Measurements were performed by using a monochromator for excitation and a photomultiplier detection system. 3. Somatic calcium signalling was investigated during defined voltage-clamp protocols. Calcium responses were observed for membrane depolarizations positive to -50 mV. A linear relation between depolarization time and free calcium concentrations ([Ca2+]i) indicated that voltage-dependent calcium influx dominated the response. 4. Endogenous calcium homeostasis was quantified by using the 'added buffer' approach. In the presence of fura-2 and mag-fura-5, calcium transients decayed according to a monoexponential function. Decay-time constants showed a linear dependence on dye concentration and the extrapolated constant in the absence of indicator dye was 371 +/- 120 ms (n = 13 cells, 21 C). 5. For moderate elevations (< 1 microM), recovery kinetics of depolarization-induced calcium transients were characterized by a calcium-independent, 'effective' extrusion rate gamma = 140 +/- 47 s-1 (n = 13 cells, 21 C). 6. The endogenous calcium binding ratio for fixed buffers in spinal motoneurones was kappaB' = 50 +/- 17 (n = 13 cells), indicating that less than 2 % of cytosolic calcium ions contributed to [Ca2+]i. 7. Endogenous binding ratios in spinal motoneurones were small compared to those found in hippocampal or cerebellar Purkinje neurones. From a functional perspective, they provided motoneurones with rapid dynamics of cytosolic [Ca2+]i for a given set of influx, extrusion and uptake mechanisms. 8. With respect to pathophysiological conditions, our measurements are in agreement with a model where the selective vulnerability of spinal motoneurones during excitotoxic conditions and motoneurone disease partially results from low endogenous calcium buffering. PMID- 10523418 TI - Role of calcium channels in catecholamine secretion in the rat adrenal gland. AB - 1. We elucidated the contribution of voltage-dependent Ca2+ channels to cholinergic control of catecholamine secretion in the isolated perfused rat adrenal gland. 2. Nifedipine (0.3-3 microM) inhibited increases in noradrenaline output induced by transmural electrical stimulation (1-10 Hz) and acetylcholine (6-200 microM), whereas it only slightly inhibited the adrenaline output responses. Nifedipine also inhibited the catecholamine output response induced by 1, 1-dimethyl-4-phenyl-piperazinium (DMPP; 5-40 microM) but not by methacholine (10-300 microM). 3. omega-Conotoxin MVIIC (10-1000 nM) inhibited the catecholamine output responses induced by electrical stimulation but not by acetylcholine, DMPP and methacholine. 4. omega-Conotoxin GVIA (50-500 nM) had no inhibitory effect on the catecholamine output responses. 5. These results suggest that L-type Ca2+ channels are responsible for adrenal catecholamine secretion mediated by nicotinic receptors but not by muscarinic receptors, and that their contribution to noradrenaline secretion may be greater than that to adrenaline secretion. P/Q-type Ca2+ channels may control the secretion at a presynaptic site. PMID- 10523419 TI - Analysis of NMDA-independent long-term potentiation induced at CA3-CA1 synapses in rat hippocampus in vitro. AB - 1. Excitatory postsynaptic currents (EPSCs) were evoked at synapses formed by Schaffer collaterals/commissural (CA3) axons with CA1 pyramidal cells using the rat hippocampal slice preparation. Long-term potentiation (LTP) was induced at these synapses using a pairing protocol, with 50 microM d,l-APV present in the artificial cerebrospinal fluid (ACSF). 2. Quantal analysis of the amplitudes of the control and conditioned EPSCs showed that the enhancement of synaptic strength was due entirely to an increase in quantal content of the EPSC. No change occurred in the quantal current. 3. These results were compared with those obtained from a previous quantal analysis of LTP induced in normal ACSF, where both quantal current and quantal content increased. The results suggest that calcium entering via NMDA receptors initiates the signalling cascade that results in enhanced AMPA currents because it is adding to cytoplasmic calcium from other sources to reach a threshold for this signalling pathway, or because calcium entering via NMDA receptors specifically activates this signalling pathway. PMID- 10523420 TI - Quantal evoked depolarizations underlying the excitatory junction potential of the guinea-pig isolated vas deferens. AB - 1. The effects of a putative gap junction uncoupling agent, heptanol, on the intracellularly recorded junction potentials of the guinea-pig isolated vas deferens have been investigated. 2. After the stimulation-evoked excitatory junction potentials (EJPs) had been suppressed by heptanol (2.0 mM) to undetectable levels, a different pattern of evoked activity ensued. This consisted of transient depolarizations that were similar to EJPs in being stimulus locked and in occurring at a fixed latency, but differed from EJPs in that they occurred intermittently and had considerably briefer time courses. 3. Analysis of the amplitudes and temporal parameters of the rapid residual depolarizations revealed a close similarity with spontaneous EJPs (SEJPs). There was no statistically significant difference between the rise times, time constants of decay and durations of the rapid residual depolarizations and of SEJPs. 4. Selected evoked depolarizations were virtually identical to SEJPs occurring in the same cell. Evoked depolarizations of closely similar amplitude and time course also occurred, usually within a few stimuli of each other. 5. These depolarizations appear to represent the individual quantal depolarizations that normally underlie the EJP and are therefore termed 'quantal excitatory junction potentials' (QEJPs) to distinguish them from both EJPs and SEJPs. 6. We examined the possibility that heptanol revealed QEJPs by disrupting electrical coupling between cells in the smooth muscle syncytium. Heptanol (2.0 mM) had no effect on the amplitude distribution, time courses, or the frequency of occurrence of SEJPs. Intracellular input impedance (Rin) of smooth muscle cells was left unaltered by heptanol. 7. 'Cable' potentials of the vas deferens, recorded using the partition stimulation method, also remained unchanged in the presence of heptanol. Thus, heptanol appeared not to modify syncytial electrical properties of the smooth muscle in any significant way. 8. Our observations show directly that the quantal depolarizations underlying the EJP in syncytial smooth muscle are SEJP-like events. However, no unequivocal statement can be made about the mechanism by which heptanol unmasks QEJPs from EJPs. PMID- 10523421 TI - The endothelial component of cannabinoid-induced relaxation in rabbit mesenteric artery depends on gap junctional communication. AB - 1. We have shown that the endocannabinoid anandamide and its stable analogue methanandamide relax rings of rabbit superior mesenteric artery through endothelium-dependent and -independent mechanisms that are unaffected by blockade of NO synthase and cyclooxygenase. 2. The endothelium-dependent component of the responses was attenuated by the gap junction inhibitor 18alpha-glycyrrhetinic acid (18alpha-GA; 50 microM), and a synthetic connexin-mimetic peptide homologous to the extracellular Gap 27 sequence of connexin 43 (43Gap 27, SRPTEKTIFII; 300 microM). By contrast, the corresponding connexin 40 peptide (40Gap 27, SRPTEKNVFIV) was inactive. 3. The cannabinoid CB1 receptor antagonist SR141716A (10 microM) also attenuated endothelium-dependent relaxations but this inhibition was not observed with the CB1 receptor antagonist LY320135 (10 microM). Furthermore, SR141716A mimicked the effects of 43Gap 27 peptide in blocking Lucifer Yellow dye transfer between coupled COS-7 cells (a monkey fibroblast cell line), whereas LY320135 was without effect, thus suggesting that the action of SR141716A was directly attributable to effects on gap junctions. 4. The endothelium-dependent component of cannabinoid-induced relaxation was also attenuated by AM404 (10 microM), an inhibitor of the high-affinity anandamide transporter, which was without effect on dye transfer. 5. Taken together, the findings suggest that cannabinoids derived from arachidonic acid gain access to the endothelial cytosol via a transporter mechanism and subsequently stimulate relaxation by promoting diffusion of an to adjacent smooth muscle cells via gap junctions. 6. Relaxations of endothelium-denuded preparations to anandamide and methanandamide were unaffected by 43Gap 27 peptide, 18alpha-GA, SR141716A, AM404 and indomethacin and their genesis remains to be established. PMID- 10523422 TI - Excitatory effect of P2X receptor activation on mesenteric afferent nerves in the anaesthetised rat. AB - 1. We examined the effects of P2X purinoceptor agonists and P2 purinoceptor antagonists on mesenteric afferent nerves supplying the jejunum in the pentobarbitone sodium-anaesthetised rat. 2. ATP (0. 01-10 mg kg-1, i.a.) and alpha,beta-methylene-ATP (1-30 microg kg-1, i.a.) each induced dose-dependent increases in afferent nerve discharge and intrajejunal pressure. The effect on afferent nerves comprised an early (< 2 s after administration) intense burst of activity followed by a later increase (> 2 s after administration), less pronounced in comparison, which coincided with elevated intrajejunal pressure. 3. Pyridoxalphosphate-6-azophenyl-2', 4'-disulphonic acid (20 mg kg-1, i.v.) and suramin (80 mg kg-1, i.v. ) each antagonised both the early and later increases in afferent nerve discharge elicited by alpha,beta-methylene-ATP (30 microg kg-1, i.a.). 4. Co-administration of omega-conotoxin MVIIA and omega-conotoxin SVIB (each at 25 microg kg-1, i.v.), or treatment with the selective 5-HT3 receptor antagonist alosetron (30 microg kg-1, i.v.), did not affect the rapid burst of afferent nerve activity elicited by alpha,beta-methylene-ATP (30 microg kg-1, i.a.). However, toxin treatment did attenuate the elevations in intrajejunal pressure and the corresponding later phases of evoked afferent discharge, while alosetron inhibited basal afferent nerve activity. 5. In summary, ATP and alpha,beta-methylene-ATP each evoke excitation of mesenteric afferent nerves in the anaesthetised rat. We propose that the early increase in mesenteric afferent nerve activity represents a direct effect on the nerve ending, mediated by P2X receptors, whereas the later increase reflects activation of mechanosensitive fibres secondary to elevated intrajejunal pressure. PMID- 10523423 TI - Changes in somatic action potential shape in guinea-pig nociceptive primary afferent neurones during inflammation in vivo. AB - We have examined whether there are changes during inflammation in the membrane properties of nociceptive primary afferent neurones in the guinea-pig that might contribute to hyperalgesia. Inflammation was induced by intradermal injections of complete Freund's adjuvant (CFA) in the left leg. Intracellular voltage recordings were made from the somata of ipsilateral L6 and S1 dorsal root ganglion neurones in anaesthetised untreated guinea-pigs at 2 or 4 days after CFA treatment. 2. Units were classified as C, Adelta or Aalpha/beta on the basis of their dorsal root conduction velocities (CVs). Units with receptive fields on the left leg were characterized as nociceptive, low- threshold mechanoreceptive (LTM) or unresponsive according to their responses to mechanical and thermal stimuli. The shapes of their somatic action potentials (APs) evoked by dorsal root stimulation were recorded. 3. Comparisons of data from nociceptive neurones recorded in CFA treated animals after 2 and 4 days with data from CFA untreated (control) animals showed the following significant changes: in C-fibre nociceptors, decreased AP duration at base, AP rise time and AP fall time, and increased maximum rates of AP rise and fall with no change in afterhyperpolarization measured to 80 % recovery (AHP80); in Adelta-fibre nociceptors, decreased AP duration at base, AP fall time and a reduction in AHP80; and in Aalpha/beta-fibre nociceptors, a decreased AHP80 but no change in AP duration. Apart from a more negative membrane potential and AHP depth below 0 mV in Aalpha/beta nociceptors at 4 days compared with 2 days post-CFA, none of the above variables differed significantly between units recorded 2 or 4 days after CFA. Therefore the two groups were pooled and called CFA2 + 4d. 4. The reduction in AP duration in C-fibre nociceptors was apparent both in high threshold mechanoreceptor and polymodal nociceptors and also in units with either cutaneous or subcutaneous receptive fields. 5. No significant changes in AP duration at base or AHP80 were seen 2 or 4 days after CFA compared with control in either LTM or unresponsive neurones, although some of the latter may have become classified as nociceptors after CFA treatment. 6. The alterations in membrane properties of nociceptors should permit higher discharge frequencies, thus contributing to inflammatory hyperalgesia. They suggest active changes in the expression or activation of cation channels during peripheral inflammation. PMID- 10523424 TI - Metabolic and thermodynamic responses to dehydration-induced reductions in muscle blood flow in exercising humans. AB - 1. The present study examined whether reductions in muscle blood flow with exercise-induced dehydration would reduce substrate delivery and metabolite and heat removal to and from active skeletal muscles during prolonged exercise in the heat. A second aim was to examine the effects of dehydration on fuel utilisation across the exercising leg and identify factors related to fatigue. 2. Seven cyclists performed two cycle ergometer exercise trials in the heat (35 C; 61 +/- 2 % of maximal oxygen consumption rate, VO2,max), separated by 1 week. During the first trial (dehydration, DE), they cycled until volitional exhaustion (135 +/- 4 min, mean +/- s.e.m.), while developing progressive DE and hyperthermia (3.9 +/- 0.3 % body weight loss and 39.7 +/- 0.2 C oesophageal temperature, Toes). On the second trial (control), they cycled for the same period of time maintaining euhydration by ingesting fluids and stabilising Toes at 38.2 +/- 0.1 degrees C. 3. After 20 min of exercise in both trials, leg blood flow (LBF) and leg exchange of lactate, glucose, free fatty acids (FFA) and glycerol were similar. During the 20 to 135 +/- 4 min period of exercise, LBF declined significantly in DE but tended to increase in control. Therefore, after 120 and 135 +/- 4 min of DE, LBF was 0.6 +/- 0.2 and 1.0 +/- 0.3 l min-1 lower (P < 0.05), respectively, compared with control. 4. The lower LBF after 2 h in DE did not alter glucose or FFA delivery compared with control. However, DE resulted in lower (P < 0.05) net FFA uptake and higher (P < 0.05) muscle glycogen utilisation (45 %), muscle lactate accumulation (4.6-fold) and net lactate release (52 %), without altering net glycerol release or net glucose uptake. 5. In both trials, the mean convective heat transfer from the exercising legs to the body core ranged from 6.3 +/- 1.7 to 7.2 +/- 1.3 kJ min-1, thereby accounting for 35-40 % of the estimated rate of heat production ( approximately 18 kJ min-1). 6. At exhaustion in DE, blood lactate values were low whereas blood glucose and muscle glycogen levels were still high. Exhaustion coincided with high body temperature ( approximately 40 C). 7. In conclusion, the present results demonstrate that reductions in exercising muscle blood flow with dehydration do not impair either the delivery of glucose and FFA or the removal of lactate during moderately intense prolonged exercise in the heat. However, dehydration during exercise in the heat elevates carbohydrate oxidation and lactate production. A major finding is that more than one-half of the metabolic heat liberated in the contracting leg muscles is dissipated directly to the surrounding environment. The present results indicate that hyperthermia, rather than altered metabolism, is the main factor underlying the early fatigue with dehydration during prolonged exercise in the heat. PMID- 10523425 TI - Voluntary and reflex control of human back muscles during induced pain. AB - 1. Back pain is known to change motor patterns of the trunk. The purpose of this study was to examine the motor output of the erector spinae (ES) muscles during pain in the lumbar region. First, their voluntary activation was assessed during flexion and re-extension of the trunk. Second, effects of cutaneous and muscle pain on the ES stretch reflex were measured, since increased stretch reflex gain has been suggested to underlie increased muscle tone in painful muscles. 2. The trunk movement and electromyographical (EMG) signals from the right and left ES during pain were compared with values before pain. Controlled muscle pain was induced by infusion of 5 % saline into the right lumbar ES. Cutaneous pain was elicited by mechanical or electrical stimulation of the dorsal lumbar skin. The stretch reflex was evoked by rapidly indenting the right lumbar ES with a servo motor prodder. 3. The results from the voluntary task show that muscle pain decreased the modulation depth of ES EMG activity. This pattern was associated with a decreased range and velocity of motion of the painful body segment, which would normally serve to avoid further injury. Interestingly, when subjects overcame this guarding tendency and made exactly the same movements during pain as before pain, the EMG modulation depth was still reduced. The results seem to reconcile the controversy of previous studies, in which both hyper- and hypoactivity of back muscles in pain have been reported. 4. In the tapped muscle, the EMG response consisted of two peaks (latency 19.3 +/- 2.1 and 44.6 +/- 2.5 ms, respectively) followed by a trough. On the contralateral side the first response was a trough (26.2 +/- 3.2 ms) while the second (46.4 +/- 4.3 ms) was a peak, similar to the second peak on the tapped side. Cutaneous pain had no effect on the short-latency response but significantly increased the second response on the tapped side. Surprisingly, deep muscle pain had no effect on the stretch reflex. A short-latency reciprocal inhibition exists between the right and left human ES. 5. It is concluded that deep back pain does not influence the stretch reflexes in the back muscles but modulates the voluntary activation of these muscles. PMID- 10523427 TI - Training-induced adaptations in the central command and peripheral reflex components of the pressor response to isometric exercise of the human triceps surae. AB - 1. The effect of calf raise training of the dominant limb on the pressor response to isometric exercise of the triceps surae was examined in the trained dominant limb and the contralateral untrained limb. Blood pressure and heart rate responses to electrically evoked and voluntary exercise at 30 % maximum voluntary contraction (MVC), followed by post-exercise circulatory occlusion (PECO), were compared before and after a 6 week training period. 2. In the trained limb the diastolic blood pressure rise seen during electrically evoked exercise was reduced by 27 % after training. However, the response during PECO was not significantly affected. 3. During voluntary exercise of the trained limb, diastolic blood pressure rise was reduced by 28 %, and heart rate rise was significantly attenuated after training. During PECO no significant effects of training were observed. 4. Voluntary exercise of the untrained limb resulted in a 24 % reduction in diastolic blood pressure rise after the training period, and a significant attenuation of the heart rate increase during exercise. Responses to electrically evoked exercise and PECO of the untrained limb remained unaltered after training. 5. Attenuation of blood pressure and heart rate responses, in the contralateral untrained limb, during voluntary but not electrically evoked exercise, indicates a training-induced alteration in central command. PMID- 10523426 TI - Modulation of reciprocal inhibition between ankle extensors and flexors during walking in man. AB - 1. The modulation of disynaptic reciprocal inhibition between antagonistic ankle muscles during walking was investigated in 17 healthy human subjects. Inhibition from ankle dorsiflexors to ankle plantar flexors was evoked by stimulation of the common peroneal nerve (CPN) and evaluated as the stimulus-induced depression of rectified soleus EMG activity (latency approx. 40 ms) or the short-latency depression of the soleus H-reflex (conditioning-test intervals around 2-3 ms). In some experiments the inhibition from ankle plantar flexors to ankle dorsiflexors was investigated. In these experiments the tibial nerve was stimulated and the amount of inhibition was evaluated from the short-latency depression of the voluntary rectified tibialis anterior (TA) EMG. 2. The short-latency inhibition of the soleus H-reflex following the CPN stimulation (1.1 x motor threshold; MT) was strongly modulated during walking, being large in the swing phase and absent in the stance phase. 3. A smaller amount of EMG depression following the CPN stimulation (1. 1-1.2 x MT) was observed in the stance phase of walking as compared to tonic or dynamic plantar flexion at a similar background EMG activity level in standing or sitting subjects. 4. In four subjects a depression of the TA EMG activity was produced by stimulation of the tibial nerve (1.1-1.2 x MT). In all subjects a smaller amount of inhibition was observed in the swing phase of walking as compared to tonic dorsiflexion at a comparable EMG activity level. 5. It is concluded that the transmission in the disynaptic Ia reciprocal pathway between ankle plantar flexors and dorsiflexors is modulated during walking. Inhibition from dorsiflexors to plantar flexors seems to be large in swing and small in stance, whereas inhibition from plantar flexors to dorsiflexors seems to be small in swing. PMID- 10523428 TI - Psychotherapy research: new findings and implications for training and practice. AB - The last decade has seen progress in psychotherapy research, despite the methodological complexity in this field. However, empirical research has influenced training and clinical practice to only a limited extent. This article is a brief evaluation of trends and some findings in modern psychotherapy research that may influence professional psychotherapy training and practice. PMID- 10523429 TI - Measurement of transference interpretations. AB - The authors present a cost-efficient process rating scale for detailed measurement of how much transference interpretations and related therapist interventions are used in brief dynamic psychotherapy. Theoretical and methodological considerations on how to operationalize and quantify such therapeutic interventions are discussed. The scale had highly satisfactory interrater reliability for three raters, who rated 60 whole sessions from an ongoing randomized study of two manualized forms of brief dynamic psychotherapy. In one treatment group, moderate emphasis on transference analysis was intended. In the other, minor or no use of the studied component was intended. The two treatment groups differed significantly in the use of transference interpretations and related interventions. There was no significant difference in therapists' general therapeutic skill or use of supportive interventions. The treatment differentiation was consistent with the manuals. PMID- 10523430 TI - Presence and enactment as a vehicle of psychotherapeutic change. AB - This article addresses an aspect of psychoanalytic and psychotherapeutic process that leads to change. Focusing on an aspect of the patient-therapist interaction that the author calls "presence" of the therapist, it demonstrates how the experience of this may lead the patient to unconscious enactment of early wishful fantasies concerning the good parent. The gratification of these wishes implicit in the interaction influences the therapist-patient relationship and plays a significant role in change. PMID- 10523431 TI - Sexual misconduct and enactment. AB - Sexual misconduct remains a significant problem in the behavioral health professions. Although it is tempting to view sexual misconduct as perpetrated by "bad" clinicians against patients who are "victims," this is an oversimplification of a complex problem. In this article, the author explores the psychoanalytic concept of enactment as a mechanism that can lead well-meaning clinicians to engage in sexual misconduct; defines enactment and differentiates it from near neighbor phenomena; uses case examples to illustrate how enactments may lead to sexual misconduct or may offer opportunities to deepen and enhance psychotherapeutic work; and offers recommendations for prevention of sexual misconduct. PMID- 10523432 TI - Psychodynamic perspective on therapeutic boundaries: creative clinical possibilities. AB - Discussion of boundaries in therapeutic work most often focuses on boundary maintenance, risk management factors, and boundary violations. The psychodynamic meaning and clinical management of boundaries in therapeutic relationships remains a neglected area of discourse. Clinical vignettes will illustrate a psychodynamic, developmental-relational perspective using boundary dilemmas to deepen and advance the therapeutic process. This article contributes to the dialogue about the process of making meaning and constructing therapeutically useful and creative boundaries that further the psychotherapeutic process. PMID- 10523434 TI - What Do We know about master therapists? PMID- 10523435 TI - Appreciation to reviewers PMID- 10523433 TI - Psychotherapies in residency training. PMID- 10523436 TI - Skin anatomy, physiology, and pathophysiology. AB - Human skin is the largest multifunctional organ of the body, and knowledge of its structure and function is essential to clinicians and researchers. The skin has two layers, the epidermis and dermis, separated by a basement membrane zone. It provides protection, sensation, thermoregulation, biochemical/metabolic, and immune functions. Key and emerging concepts important to understanding pathophysiological mechanisms for practicing clinicians are: knowledge of differences between acute and chronic wounds; ability to evaluate depth and extent of injury; understanding stages of healing versus zones of activity; and knowledge of ischemic-reperfusion injury, the skin immune system, cytokines, growth factors and other biomolecules, and matrix synthesis and degradation. These concepts are addressed in this article. PMID- 10523437 TI - Chronic wound assessment. AB - Chronic wounds are a drain on health care resources, and as such, continue to challenge health care providers to define and create more effective intervention strategies. Wound assessment is the foundation for maintaining and evaluating a therapeutic plan of care. Without adequate baseline wound assessment and valid interpretation of the assessment data, the plan of care for the wound may be inappropriate or ineffective. This article discusses comprehensive assessment of the total patient as well as assessment of wound severity and wound status. PMID- 10523438 TI - Pain management of wound care. AB - Children and adults still suffer pain during wound dressing changes despite national guidelines. Assessing and managing pain are essential components of comprehensive wound care. Developmentally sensitive pain assessment tools are available to measure verbal, behavioral, and physiologic responses to pain. Holistic pain assessment includes pain intensity, location, description, relief measures, cultural background, and the patient's developmental level and anxiety. Pharmacologic and nonpharmacologic interventions should be combined to manage pain based upon patient's response and nursing assessment. Nurses with a fundamental knowledge of pain assessment and management provide their patients with pain and symptom relief during wound care. PMID- 10523440 TI - Surgical alternatives for wounds. AB - Surgical alternatives in wound care are a primary consideration for the treatment of nonhealing and traumatic wounds. Using the Reconstructive Ladder as an outline, this article provides an overview of preoperative wound care and the indications for surgical options in wound care. An overview of nursing care is highlighted as each option is reviewed to include perioperative care of the patient. PMID- 10523439 TI - Interrupting the pressure ulcer cycle. AB - Pressure ulcers represent a significant health care problem for older adults and patients with chronic disease. The National Pressure Ulcer Advisory Panel (NPUAP) estimates that over 1 million people in America suffer from pressure ulcers. Reported pressure ulcer prevalence varies widely and pressure ulcer incidence is difficult and labor-intensive to track; however, nursing has both a responsibility and an opportunity to help decrease the number of individuals who suffer from pressure ulcers. This article suggests ways that nurses can assist with improving care for patients at risk for pressure ulceration. PMID- 10523441 TI - Trauma wound management. AB - This article provides a review of wound healing and management after injury. Wound management includes cleansing, primary and delayed closure, and a discussion of gunshot wounds. Forensic issues for evidence collection and the accurate documentation of wounds are reviewed. PMID- 10523442 TI - Prevention and management of infant skin breakdown. AB - There are anatomic and physiologic differences in the skin of both premature and full-term infants that place them at increased risk for skin injury and breakdown. This article reviews these differences and discusses some of the infant skin care practices that can cause injury to infant skin with preventive strategies identified. Care of skin breakdown is outlined, and topical treatments reviewed in detail. Diaper dermatitis is also discussed with treatment of the underlying causes as well as the goals of treating diaper dermatitis described. PMID- 10523443 TI - Meeting the challenges of healing chronic wounds in older adults. AB - Older Americans are living longer. With aging, the skin becomes thinner, more fragile, and more susceptible to injury. Progressive normal and pathological aging changes increase the risk for chronic health problems that may lead to open wounds and delayed healing. Optimum care of chronic wounds supports the older person's specific health and care needs, progresses to cleansing and debridement of the wound, then uses an appropriate cover and other protective interventions to promote healing. PMID- 10523445 TI - Evidence-based practice: what is it and how is it used in wound care? AB - Evidence-based practice is an approach to providing clinical care based on research and scientific knowledge. Nurses who work with patients with wounds need to understand and apply evidence-based principles. This article provides guidelines for critical analysis of the literature using the principles of evidence-based practice. PMID- 10523444 TI - Nursing management of chronic wounds: best practices across the continuum of care. AB - This article highlights the nurse's role within a holistic, interdisciplinary approach to chronic wound management. Best practices for chronic wound care are discussed, drawing on evidence-based science when it is available. The fundamentals of chronic wound care, including cleansing, irrigation, debridement, infection control, and topical treatment are addressed. New devices and technologies are briefly reviewed. Implementing these best practices across the continuum of care will result in greater advances in the management of chronic wounds. PMID- 10523446 TI - The Marion Downs National Center for Infant Hearing: developing comprehensive state systems. AB - The Marion Downs National Center for Infant Hearing was established in 1996 through a Maternal and Child Health Grant awarded to the University of Colorado. The goals of the grant are to implement statewide systems of newborn hearing screening, audiologic assessment, and early intervention in 19 states. Newborn hearing screening alone will not assure early identification or positive outcomes for the development of communication and language. Therefore, the staff at the Marion Downs National Center developed comprehensive goals for all participating states. These goals are described in this article. PMID- 10523447 TI - Hear early: New Mexico's universal newborn hearing screening program. AB - Hear Early, New Mexico's newborn screening program, was established in 1996. Thirty-one of the state's 32 birthing hospitals participate in the Hear Early program that now screens 95% of New Mexico's 27, 500 births. The authors' experience in developing this statewide system is described and the critical components and the rationale behind this program are addressed. The development of Hear Early required input from a wide variety of professionals and consumers. Screening is only the first step in a comprehensive state program, which must also include assessment and intervention. Systems development, equipment issues, follow-up procedures, data management, and a tracking system are all pertinent factors. The various accomplishments and challenges that Hear Early has encountered are discussed. PMID- 10523448 TI - Automated auditory brainstem response testing for universal newborn hearing screening. AB - The goal of universal newborn hearing screening is to identify infants with hearing loss by 3 months of age and to provide appropriate intervention by 6 months of age. Automated auditory brainstem response helps accomplish this goal by providing a test that is easy to use, fast, cost-effective, and accurate. Universal newborn hearing screening with automated auditory brainstem response ensures that children with hearing loss are given the opportunity for early intervention and an improved quality of life. PMID- 10523449 TI - Audiologic evaluation of hearing-impaired infants and children. AB - With the advent of auditory brainstem response (ABR) and otoacoustic emission (OAE) testing, as well as the widespread use of universal hearing screening programs, the audiologic assessment of hearing-impaired infants and young children has become more frequent in recent years. When assessing a hearing impaired infant, it is necessary to obtain as much frequency and ear specific information as possible prior to selection of hearing amplification. An overview of the battery of tests and the need for a diagnostic center that is well versed in both physiologic and behavioral tests will be addressed. PMID- 10523450 TI - Clinical evaluation of the hearing-impaired infant. AB - Early identification of hearing loss and rapid rehabilitative intervention are the two key elements that will give an infant the best chance to develop normal speech and allow the family members to make appropriate adjustments that will enhance communication in the home environment. Hearing loss in an infant may be only one feature of a syndromic disorder, and the physician must aspire to diagnose and treat the entire dysfunction. While a limited laboratory evaluation and appropriate referrals to related specialties are helpful, a comprehensive history and a complete physical examination are the most effective means to identify syndromic hearing loss and to direct future evaluation and therapy. PMID- 10523451 TI - Counseling families with a hearing-impaired child. AB - Parents are the linchpin of the family, and counseling for the young, newly diagnosed child with a hearing impairment needs to be directed at them. The operative rule for the counseling model described in this article is that feelings are neither good nor bad; they just are, and they need acknowledgment and acceptance, never judgment. Behavior can be judged as to whether or not it is productive, but parents never have to be responsible for how they feel. Armed with this nonjudgmental attitude toward emotions, the clinician can elicit them by empathetic listening. PMID- 10523452 TI - Amplification for infants: selection and verification. AB - Hearing aid fittings for children under the age of 1 year provide many challenges to the audiologist. This article offers guidelines for assessment, hearing aid selection, verification, and follow-up for infants with hearing loss. The importance of early intervention and the consideration of new technologies are emphasized in this article. Case studies illustrate various aspects of infant hearing aid fittings. PMID- 10523453 TI - Genetics and molecular biology of deafness. AB - With increased emphasis on early detection of hearing impairment, more babies are likely to be referred at younger ages to otolaryngologists for evaluation. With a diminution in the number of infants who have hearing impairment as a result of such factors as maternal infection, neonatal sepsis, or ototoxicity, the relative importance of detecting a genetic cause of newborn hearing impairment is likely to increase. Therefore, the otolaryngologist must become familiar with common causes of hereditary hearing impairment and the ways in which the newborn should be evaluated for hereditary hearing impairment. Advancements are rapidly being made in the ability to detect genes that cause hearing impairment, and we are now on the threshold of discovering ways to use gene therapy to prevent or treat hereditary deafness. PMID- 10523455 TI - Communication methodologies: options for families. AB - When parents are told their child has a hearing loss they not only have to cope with the loss, they must also decide which communication and educational option is best for them. The purpose of this article is to discuss the different communication modalities available for hearing-impaired children. Whether parents choose auditory/oral, sign language, or cued speech they must be committed to the chosen modality. It is important that families are presented with all the communication options available in an unbiased manner. Parents need to research each option carefully and decide which is best, not only for the child, but also for the entire family. PMID- 10523454 TI - Benefits of early intervention for children with hearing loss. AB - This article discusses the relationship between language development and degree of hearing loss and how these factors relate to the age of identification of hearing loss. An interesting effect caused by the interaction of these variables has been discussed in this article. The relationship of speech production, degree of hearing loss, language development, and age of identification are also presented. Social-emotional development of deaf and hard-of-hearing children and the variables that impact this development, in particular the strong relationship with language development, is included. PMID- 10523456 TI - Cochlear implantation in the very young child. AB - Children younger than 2 years of age were initially excluded from cochlear implant candidacy for a variety of reasons. Reasons ranged from concerns about the reliability of the diagnosis of a profound hearing loss in very young children, to concerns about surgical safety and long-term durability of the device in a growing child. Results from several recent studies have shown that children younger than 2 years of age can safely and successfully be implanted. Provided this success, and the general agreement that early remediation of a hearing loss provides a greater potential for speech and language development, implantation of very young children may soon become the norm rather than the exception. This article discusses the issues related to the implantation of young children and the need for special tools and protocols to use with this population. PMID- 10523457 TI - Speech and language acquisition in young children after cochlear implantation. AB - Auditory and speech intelligibility scores of 22 prelingually profoundly deaf children who had used cochlear implants for between 1 and 7 years in an intensive auditory/oral educational program greatly exceeded those previously obtained from similar samples of deaf children using hearing aids. Half of the children obtained language quotient scores within the average range for normal-hearing children and the majority obtained reading quotients within 80% of normal levels. Normal levels of language and reading development were associated with early implantation and open set speech perception. PMID- 10523458 TI - The diagnosis and classification of diabetes mellitus in nonpregnant adults. AB - This article reviews the definition, diagnosis, and classification of diabetes and focuses on the rationale for and the limitations of the new diagnostic criteria for diabetes adopted by the American Diabetes Association in 1997. It also reviews the impact of the new fasting criterion for diabetes on both the prevalence of diabetes and the phenotype of individuals identified. Finally, it describes the revised classification scheme for diabetes and explains how it simplifies and clarifies some of the confusion created by the previous scheme. PMID- 10523459 TI - The natural history of type 2 diabetes. Implications for clinical practice. AB - This article reviews the natural history of type 2 diabetes and emphasizes that the disease is a continuum from impaired glucose tolerance and impaired fasting glucose to frank diabetes. Understanding this natural history aids the clinician in identifying those individuals most at risk for developing type 2 diabetes. Early intervention may significantly limit the severity of the microvascular and macrovascular complications of diabetes. Additionally, this article discusses oral antidiabetic agents and reviews their potential effectiveness at the different stages in the continuum to aid the clinician in developing a treatment strategy. PMID- 10523460 TI - Prevention and treatment of microvascular and neuropathic complications of diabetes. AB - The most important factors that influence development of the eye, kidney, and nerve complications of diabetes are the duration and degree of exposure to hyperglycemia. Hypertension is also very important. The risk of complications appears to be similar for all patients with diabetes; differences in complication rates between patients with type 1 and type 2 diabetes are largely due to differences in duration of diabetes and glycemic control. The benefits of lowering blood sugar are also similar for patients with diabetes, regardless of the type. Significant reductions in complications can be seen with relatively short-term treatment of hyperglycemia. PMID- 10523461 TI - Implications of the United Kingdom prospective diabetes study. AB - The United Kingdom Prospective Diabetes Study (UKPDS) is the largest ever study to clarify the role of glycemia and blood pressure control in improving morbidity and mortality in patients with type 2 diabetes. The study examined the impact of glycemia and blood pressure control on microvascular and macrovascular risk, and if any particular therapy was advantageous for these patients. This articles discusses the background, methodology and results of the UKPDS and develops recommendations for treatment and management of patients with type 2 diabetes in the clinical setting. PMID- 10523462 TI - Hyperglycemia as a risk factor for cardiovascular disease in type 2 diabetes. AB - According to several prospective population-based studies, glycemic control influences the risk for cardiovascular disease, including coronary heart disease, independently of conventional risk factors in patients with type 2 diabetes. Recent clinical trials, particularly the United Kingdom Prospective Diabetes Study, have shown that microvascular complications and macrovascular complications, although to a lesser extent, can be prevented in patients with type 2 diabetes with correction of glycemic control. However, in the treatment and prevention of cardiovascular disease in type 2 diabetes, all known cardiovascular risk factors should be attacked simultaneously. PMID- 10523463 TI - Defining and measuring quality of diabetes care. AB - The literature on diabetes mellitus has increasingly focused on the quality of diabetes care and its measurement. Serious and widespread quality problems exist throughout American medicine. Current efforts to improve will not succeed unless we undertake a major, systematic effort to overhaul how we deliver health care services, educate and train clinicians, and assess and improve quality. This article defines the components of quality of diabetes care provision and discusses approaches to their measurement individually and globally. PMID- 10523464 TI - Nutrition management and physical activity as treatments for diabetes. AB - Diabetes is essentially a self-management disease in which patients must learn to integrate blood glucose monitoring with nutrition management and physical activity and, if needed, oral agents or insulin. This almost always requires behavior change. The American Diabetes Association diet is no longer a meaningful prescription, because recommendations are now based on individualized nutrition assessment and treatment goals. Physical activity recommendations, as an adjunct to nutrition management, should also be individualized. Using a team approach, focusing on individualization of nutrition management and physical activity, applying behavior change concepts, and providing frequent follow-up can improve self-management and result in improved metabolic control. Primary care practitioners are in a unique position to oversee this process. PMID- 10523465 TI - The treatment of obesity in type 2 diabetes mellitus. AB - Obesity has a critical role in the pathophysiology of type 2 diabetes mellitus, and prevention of weight gain and treatment after onset of obesity is crucial to the management of the disease. A recent National Institutes of Health (NIH) report on the evaluation and treatment of overweight and obesity serves as a model for managing obese individuals with type 2 diabetes. Lifestyle intervention is the fundamental approach and should be implemented by a multidisciplinary team of health professionals. Adjunctive therapies, such as anti-obesity pharmacotherapy and surgery, should only be considered if at least 6 months of lifestyle intervention has produced suboptimal results. If adjunctive therapy is indicated, it will only be successful long term if lifestyle therapy remains central to treatment. The objective of this review is to integrate the recommendations of the NIH into the primary care management of the obese individual with type 2 diabetes. PMID- 10523466 TI - Special problems with children and adolescents with diabetes. AB - Treatment of children and adolescents with insulin-dependent diabetes mellitus (type 1) is different in many ways than it is for adults. Physical, cognitive, and emotional development changes affect therapeutic goals and modalities. Neonatal, early childhood, school-age, and adolescent patients all have unique needs. Further, diabetes can affect psychosocial maturation and the likelihood of difficulties with mood. PMID- 10523467 TI - The use of insulin secretagogues in the treatment of type 2 diabetes. AB - Secretatogues are a class of agents that achieve their hypoglycemic effects through stimulating insulin release. They include the sulfonylureas, repaglinide, and the investigational agent glucagon-like peptide. The secretagogue agents have been studied extensively as monotherapy and in conjunction with other classes of oral agents, including alpha-glucosidase inhibitors, bijuanides, and thiazolidinediones, for the treatment of type 2 diabetes. This article reviews the pharmacodynamic and pharmacokinetic differences of the secretagogues, as well as the most recent clinical trials. Such information should be helpful when deciding which agent or agents will yield the best glycemic control for an individual patient. PMID- 10523469 TI - The treatment of hypertension and dyslipidemia in diabetes mellitus. AB - The relationship between macrovascular complications of diabetes mellitus, hypertension, and dyslipidemia is explored. Management strategies for patients with lipid abnormalities are given, along with indications and use of available drug classes. Pharmacologic and nonpharmacologic approaches to hypertension management are reviewed. The controversies concerning the use of various classes of anti-hypertensive drugs including calcium channel blockers and diuretic therapy are also discussed. PMID- 10523468 TI - The use of insulin alone and in combination with oral agents in type 2 diabetes. AB - To achieve optimal outcomes in patients with type 2 diabetes, clinical trial data suggest that near normal glycemic control should be targeted. Insulin is arguably the most effective treatment available for diabetes and yet many patients remain poorly controlled without the benefit of insulin therapy. Discussion of its putative risks and benefits as well as the barriers to its wider use both in the context of monotherapy and in combination with oral antidiabetic agents is provided. Application of the strategies and principles of insulin therapy in type 2 diabetes should minimize the burden of complications in patients with the disease. PMID- 10523470 TI - To quebec! PMID- 10523471 TI - The changing face of heart disease and stroke in Canada - release of the fifth report from the Canadian Heart and Stroke Surveillance system. PMID- 10523472 TI - Unforgettable voyages [directory]. PMID- 10523473 TI - Access to cardiac resources. PMID- 10523474 TI - Access to cardiac resources - reply PMID- 10523475 TI - Circulatory support for cardiogenic shock due to acute myocardial infarction: a Canadian experience. AB - BACKGROUND: Cardiogenic shock due to acute myocardial infarction (AMI) is associated with high mortality. Circulatory support devices may be used to assist these patients while they await cardiac transplantation. METHODS AND RESULTS: From 1986 to 1997, 25 patients in cardiogenic shock complicating AMI within 3.6+/ 0.7 days of the event were supported with artificial hearts. Of the 25 patients, 21 were men with a mean age of 48.4 +/- 1.8 years. The age range was 26 to 62 years. Patients were considered for a device when the following criteria were met: cardiac index less than 1.8 L/min/m2, wedge pressure greater than 20 mmHg despite one or two inotropes and/or intra-aortic balloon support. They received either a CardioWest total artificial heart (n=13), a Thoratec biventricular assist device (n=6) or left ventricular assist device (LVAD) (n=6). Three patients were not considered transplant candidates and died while on the devices (two with multiorgan failure and one found to have a bronchogenic carcinoma after implant), with 22 undergoing cardiac transplantation within 8.6+/-2.2 days of device implant. Six patients died in hospital after the transplants (27.3% mortality). Complications included bleeding or tamponade in seven (28%), pneumonia in six (24%) and right ventricular failure in three LVAD patients (12%). Post-transplant actuarial one-, two- and five-year survival rates were 71.4%, 71.4% and 51%, respectively. CONCLUSIONS: Circulatory support devices offer a means to maintain organ perfusion in patients who develop cardiogenic shock due to AMI. Patients can then undergo transplantation with a reasonable expectation for survival when the alternative is death. Eventually the availability of permanent support devices may obviate the need for transplant in these patients. PMID- 10523476 TI - Higher rates of coronary angiography and revascularization following myocardial infarction may be associated with greater survival in the United States than in Canada. The CARS Investigators (Coumadin/Aspirin Reinfarction Study). AB - BACKGROUND: Significant differences are known to exist between the United States and Canada with respect to coronary catheterization and intervention. In a post hoc analysis, it was hypothesized that these differences may have the greatest impact on outcome in patients at risk for recurrent events such as those following myocardial infarction (MI). PATIENTS AND METHODS: The hypothesis was tested in a nonrandomized comparison of the catheterization and revascularization patterns for patients following acute MI in 7029 patients in the United States and 1774 patients in Canada who participated in the Coumadin/Aspirin Reinfarction Study (CARS). CARS tested the effectiveness of low dose warfarin in combination with acetylsalicylic acid (ASA) versus ASA alone in reducing cardiovascular morbidity and mortality. RESULTS: Before study enrollment (median day 7 to 8), 84.5% of the American patients underwent coronary angiography compared with only 7.7% in Canada (P<0.01). CARS patients in the United States underwent significantly more frequent angioplasty during their hospital admission before study enrollment than their Canadian counterparts (55.8% versus 3.0%, respectively, P<0.01), and there was a more frequent use of intravenous heparin among American CARS patients (96% versus 88%, respectively, P<0.01) but less frequent administration of thrombolytic therapy (44% versus 49%, respectively, P<0.01). For follow-up of up to 32 months, American CARS patients had significantly fewer primary events (cardiovascular deaths, nonfatal MI, nonfatal ischemic stroke) than Canadian patients. Cumulative estimate of a primary end point comparing American with Canadian patients was, respectively, 2.0% versus 3.1% at one month, 8.0% versus 11.3% at one year and 11.6% versus 15.0% at two years. Thus, time to the primary event was significantly longer in American patients (P=0.0001). All-cause mortality estimates at 12 months were 2.2% and 4.0%, respectively, for the American and Canadian CARS subgroups. When management for the index MI (ie, angiography and angioplasty) is included in the model, the risk of death for Canadian versus American subgroups of CARS is not statistically different (0.9, 95% CI 0.6 to 1.2, P=0.40). CONCLUSION: Among study participants, American patients experienced a better outcome than Canadian patients, which may be attributable to more aggressive management based on early coronary angiography and angioplasty. However, angioplasty before study enrollment in American patients may have resulted in enrollment of lower risk patients. This selection bias limits the scope of the comparison. PMID- 10523477 TI - Effectiveness of nifedipine GITS in combination with atenolol in chronic stable angina. AB - Nifedipine gastrointestinal therapeutic system (GITS) is a once-daily formulation of nifedipine that provides stable plasma concentrations over the entire 24 h dosing interval. Two-hundred and one patients with Canadian Cardiovascular Society class II to III angina who were on 50 mg of atenolol yet still experiencing angina symptoms were randomized to receive either placebo or nifedipine GITS 30, 60 or 90 mg/day. After four weeks of treatment, the changes in time from baseline to onset of 1 mm ST segment depression in the 183 eligible patients were 26.7+/-10.2 s, 40.9+/-11.3 s, 63.2+/-12.9 s and 70.3+/-12.6 for the placebo, and 30, 60 and 90 mg/day groups, respectively. These differences were significant (P<0.05) for the 60 and 90 mg/day groups compared with placebo and for the 60 mg/day group compared with the 30 mg/day group. The times to onset of pain and termination of exercise showed similar prolongation but did not achieve statistical significance. During the one-year open label phase of the study, patients exhibited statistically significant improvements in the time to onset of ST segment depression, time to anginal pain and time to termination of exercise at a mean dose of 52.3 mg/day of nifedipine GITS. Adverse events were primarily vasodilatory in nature. This study supports the use of nifedipine GITS in patients with chronic stable angina inadequately controlled on beta-blocker alone. PMID- 10523478 TI - Valve surgery in octogenarians. AB - OBJECTIVE: To review the outcomes of octogenarians undergoing valve operations. PATIENTS AND METHODS: One hundred and twenty-five consecutive patients aged 80 years and over received valve operations between 1990 and 1996 at the Toronto General Hospital, Toronto, Ontario. All hospital survivors were prospectively followed for a mean of 36.6 months (range 0.1 to 89.9). RESULTS: One hundred and two patients received aortic valve operations, 18 patients received mitral procedures and five patients underwent double valve operations. Significant aortic stenosis was present in 95 of 102 patients (93%) receiving isolated aortic valve surgery, and mitral regurgitation was present in 16 of 18 patients (89%) undergoing mitral valve operations. Overall in-hospital mortality was 6.4% (n=8) and the perioperative infarction rate was 1.6% (n=2). In-hospital mortality was higher for mitral valve patients at 17% (n=3) than for aortic valve patients at 4% (n=4) (P=0.06). For the group overall, the six-year actuarial survival rate was 71.6+/-6%. The actuarial freedom from valve-related death was 97.1+/-2% at three years. Concomitant coronary artery disease was not significantly associated with perioperative mortality. Survivors had significantly improved New York Heart Association functional class status. CONCLUSION: In carefully selected patients aged 80 years and over, aortic valve surgery carries a low perioperative mortality with good intermediate term survival and benefits. Octogenarians undergoing mitral valve procedures experience higher perioperative mortality. Although the number of participants was small for this study, it does appear that coexistent coronary artery disease should not be the sole reason for denial of surgery because it has less of an impact on short and intermediate term survival than other factors. PMID- 10523479 TI - Cardiac KATP channels and ischemic preconditioning: current perspectives. AB - Ischemic preconditioning (IPC) is a naturally occurring protective mechanism in the heart and is triggered by brief ischemic insults preceding a sustained ischemic period. The artificial induction of the protective effects of IPC are potentially of great benefit to postinfarct patients and would serve to augment traditional surgical and thrombolytic treatment regimens. As yet, no agent has been developed to trigger IPC specifically. The mechanisms underlying IPC have been the focus of intense study, and many components of the IPC framework have been elucidated. Adenosine, noradrenalin, angiotensin II and endothelin are several of the key inducers of IPC. Signal transduction events downstream of receptor activation converging on protein kinase C stimulation are thought to trigger the intracellular pathways leading to 'preconditioning'. Although the complete sequence of events beneath IPC are not clear, several of the end effectors for the initiation of IPC are. For instance, activation of a class of potassium channels that are sensitive to internal ATP (KATP channels) can induce IPC and may be at least one of the targets for the action of protein kinase C. The precise mechanisms by which stimulation of KATP channels protects the myocardium are still under debate and are discussed in this review. It seems likely that both the plasma membrane and mitochondrial KATP channel populations are involved in IPC. Because KATP channels are rich and varied pharmacologically, it may be possible to target therapeutic agents at the cardiac channel isoform. Such compounds would likely meet the pharmacological criteria for a 'conditioning' agent. PMID- 10523480 TI - Stenting of an unprotected left main coronary artery stenosis in a cardiac transplant patient. AB - Cardiac allograft vasculopathy is the leading cause of death in cardiac transplant patients who survive the first year. Retransplantation is limited by shortage of donors and reduced survival rates compared with the initial transplant. Recent reports of successful stenting in these patients may offer some hope, although randomized trials are lacking. Successful stenting of an 'unprotected' left main coronary artery stenosis under cardiopulmonary support is presented in a cardiac transplant patient. A 16-month follow-up angiogram demonstrated a patent stent without restenosis and no interim clinical events. PMID- 10523481 TI - Aortic dissection: a rare complication of osteogenesis imperfecta. AB - Osteogenesis imperfecta (OI) is an inherited connective tissue disorder, a group that includes Ehlers-Danlos syndrome, Marfan's syndrome and pseudoxanthoma elasticum. OI is a heterogeneous disease of collagen I biosynthesis characterized by variable clinical phenotypes, including skeletal and cardiovascular manifestations. A 65-year-old man with OI who had extensive prior successful cardiac valve surgeries is described. He survived for 18 years after his initial valve surgery, but died of multiorgan failure and sepsis after repair of a spontaneous type A aortic dissection. This is the fourth reported case of aortic dissection secondary to OI and illustrates the extensive cardiovascular pathology associated with OI. Aggressive management of arterial dissection risk factors, such as systemic arterial hypertension, is advocated for patients with OI. PMID- 10523482 TI - Right ventricular ischemia mimicking acute anterior myocardial infarction. AB - Isolated right ventricular ischemia in combination with myocardial infarction (MI) is uncommon, accounting for fewer than 3% of all MI cases. A young man who presented with acute right ventricular ischemia from occlusion of a codominant right coronary artery proximal to an acute marginal branch is presented. His presenting electrocardiogram (ECG) showed ST segment elevation in V1 to V4 mimicking acute anterior MI. ECG criteria for isolated right ventricular ischemia are discussed. PMID- 10523483 TI - Intervention rates and outcomes following acute myocardial infarction. PMID- 10523484 TI - On call. I really enjoy eating nuts but I've been told to avoid cashews (my favorites) because they have too much saturated fat. Is it true? PMID- 10523485 TI - On call. My husband subscribes to Harvard Men's Health Watch, but I read the newsletter as much as he does. I'm writing to ask about our son who just made his college lacrosse team. I remember reading about a lacrosse player in your area who died during a game. Is lacrosse safe for our boy? PMID- 10523486 TI - The Red Queen and Fluctuating Epistasis: A Population Genetic Analysis of Antagonistic Coevolution. AB - Host-parasite coevolution has been shown to provide an advantage to recombination, but the selective mechanism underlying this advantage is unclear. One possibility is that recombination increases the frequency of advantageous genotypes that are disproportionately rare because of fluctuating epistasis. However, for this mechanism to work, epistasis for fitness must fluctuate over a very narrow timescale: two to five generations. Alternatively, recombination may speed up the response to directional selection by breaking up linkage disequilibria that decrease additive genetic variance. Here we analyze the results of a numerical simulation of host-parasite coevolution to assess the importance of these two mechanisms. We find that linkage disequilibria may tend to increase, rather than decrease, additive genetic variance. In addition, the sign of epistasis changes every two to five generations under several of the parameter values investigated, and epistasis and linkage disequilibrium are frequently of opposite signs. These results are consistent with the idea that selection for recombination is mediated by fluctuating epistasis. Finally, we explore the conditions under which an allele causing free recombination can spread in a nonrecombining host population and find general agreement between the predictions of a population genetic model of fluctuating epistasis and our simulation model. PMID- 10523487 TI - Patterns of Food Chain Length in Lakes: A Stable Isotope Study. AB - Food web structure is paramount in regulating a variety of ecologic patterns and processes, although food web studies are limited by poor empirical descriptions of inherently complex systems. In this study, stable isotope ratios (delta15N and delta13C) were used to quantify trophic relationships and food chain length (measured as a continuous variable) in 14 Ontario and Quebec lakes. All lakes contained lake trout as the top predator, although lakes differed in the presumed number of trophic levels leading to this species. The presumed number of trophic levels was correlated with food chain length and explained 40% of the among-lake variation. Food chain length was most closely related to fish species richness ([Formula: see text]) and lake area ([Formula: see text]). However, the two largest study lakes had shorter food chains than lakes of intermediate size and species richness, producing hump-shaped relationships with food chain length. Lake productivity was not a powerful predictor of food chain length ([Formula: see text]), and we argue that productive space (productivity multiplied by area) is a more accurate measure of available energy. This study addresses the need for improved food web descriptions that incorporate information about energy flow and the relative importance of trophic pathways. PMID- 10523488 TI - Queen-Worker Conflict over Sexual Production and Colony Maintenance in Perennial Social Insects. AB - An important idea from kin selection theory as applied to life-history evolution in perennial insect societies suggests that potential conflict exists between the queen and workers over the relative allocation of resources to colony reproduction and colony growth. This prediction assumed a colony with one singly mated queen, sterile workers, and independent colony foundation by dispersing queens. We argue that this prediction is mistaken because queen and workers under these circumstances can only invest in sexuals (new queens and males) derived from the colony queen. Assuming population sex ratio equilibrium, potential conflict is absent because both parties maximize fitness by maximizing the colony's total output of these sexuals. However, a similar queen-worker conflict is predicted in facultatively polygynous species in which existing queens are superseded. We hypothesize that queens favor the production of relatively more new workers to prolong their lives as reproductives, whereas workers favor raising relatively more new queens as possible replacements. We tested this hypothesis using productivity data from the ant Leptothorax acervorum. As predicted, queen number and worker number were, respectively, positively and negatively associated with the investment ratio of new workers to new queens. These findings imply a queen-worker conflict over caste determination in ants. PMID- 10523489 TI - Immigration and the Maintenance of Local Species Diversity. AB - Explaining the maintenance of high local species diversity in communities governed by competition for space has been a long-standing problem in ecology. We present a simple theoretical model to explore the influence of immigration from an external source on local coexistence, species abundance patterns, and ecosystem processes in plant communities. The model is built after classical metapopulation models but is applied to competition for space between individuals and includes immigration by a propagule rain and an extinction threshold for rare species. Our model shows that immigration can have a huge effect on local species diversity in competitive communities where competition for space would lead to the exclusion of all but one species if the community were closed. Local species richness is expected to increase strongly when immigration intensity increases beyond the threshold required for the successful establishment of one or a few individuals. Community structure and species relative abundances are also expected to change markedly with immigration intensity. Increasing immigration causes total space occupation by the community to increase but primary productivity on average to either decrease or stay constant with increasing diversity, depending on the relation between immigration and local reproduction rates. These results stress the need for a regional perspective to understand the processes that determine species diversity, species abundance patterns, and ecosystem functioning in local communities. PMID- 10523490 TI - Carotenoid Plasma Concentration, Immune Profile, and Plumage Ornamentation of Male Barn Swallows (Hirundo rustica). AB - Carotenoids exert immunomodulating, immunostimulating, and antioxidant actions in mammals and are major determinants of coloration in animals. Honest advertisement models of sexual selection propose that male ornaments, including coloration, are reliable indicators of male quality. Because of their simultaneous effects on male coloration and immunity, carotenoids might mediate the hypothesized relationship between the expression of epigamic coloration and parasitism in vertebrates. We analyzed the relationship between immune profile and concentration of lutein, the most abundant carotenoid in the plasma of male barn swallows (Hirundo rustica). Consistent with our predictions, lutein plasma concentration was negatively correlated with gamma-globulin plasma levels and concentration of selected leukocyte types in peripheral blood, suggesting that, to exert immune function, carotenoids are taken up from plasma, thus becoming unavailable for epigamic signaling. The coloration of red feathers of the throat of adult males was positively related to plasma concentration of lutein, but not with immunologic variables, consistent with the idea that more brightly colored males do not pay a larger immunological cost for their coloration compared with less brightly colored males. Length of male tail ornaments, which is currently under directional sexual selection, was positively correlated with lutein plasma levels. In species where carotenoids limit immune function, demands for pigments for sexual signaling might compete with those for immunity, thus generating a mechanism that enforces honesty on the signal. PMID- 10523491 TI - Patterns in the Fate of Production in Plant Communities. AB - I examine, through an extensive compilation of published reports, the nature and variability of carbon flow (i.e., primary production, herbivory, detrital production, decomposition, export, and biomass and detrital storage) in a range of aquatic and terrestrial plant communities. Communities composed of more nutritional plants (i.e., higher nutrient concentrations) lose higher percentages of production to herbivores, channel lower percentages as detritus, experience faster decomposition rates, and, as a result, store smaller carbon pools. These results suggest plant palatability as a main limiting factor of consumer metabolical and feeding rates across communities. Hence, across communities, plant nutritional quality may be regarded as a descriptor of the importance of herbivore control on plant biomass ("top-down" control), the rapidity of nutrient and energy recycling, and the magnitude of carbon storage. These results contribute to an understanding of how much and why the trophic routes of carbon flow, and their ecological implications, vary across plant communities. They also offer a basis to predict the effects of widespread enhancement of plant nutritional quality due to large-scale anthropogenic eutrophication on carbon balances in ecosystems. PMID- 10523492 TI - Seed Mass and the Evolution of Early-Seedling Etiolation. AB - The influence of seed mass on the evolution of seedling-foraging strategies for light acquisition under deep shade was assessed in a comparative study of etiolation behavior. This was done across 50 Australian species varying in seed reserve mass by eight orders of magnitude during the first week after germination. Proportional increase in hypocotyl length in shade compared to light was similar across the range of seed reserve mass. Etiolation did not lead to increase in hypocotyl length per wet mass, in other words, etiolated hypocotyls were not thinner. However, hypocotyl length per dry mass did increase, more so in smaller-seeded species. Thus, part of the hypocotyl elongation was because of increased water content, which would increase vulnerability to loss of turgor. There was also reallocation of dry matter from root to hypocotyl, again more so in smaller-seeded species, which would decrease anchorage strength and increase vulnerability to soil drying. Results were very similar when considered as correlated evolutionary divergences, compared to the cross-species patterns. The higher-risk etiolation behavior of smaller-seeded species can be understood through their having little to lose. Because they hold less reserve resource uncommitted and attempt a faster initial growth rate, their chances of sustained longevity in shade below the compensation point are very low. PMID- 10523493 TI - How Common Are Meiotically Driving Sex Chromosomes in Insects? PMID- 10523494 TI - Floral Symmetry and Its Influence on Variance in Flower Size. PMID- 10523495 TI - Inferring Host-Parasitoid Stability from Patterns of Parasitism among Patches. PMID- 10523496 TI - Aurora/Ipl1p-related kinases, a new oncogenic family of mitotic serine-threonine kinases. AB - During the past five years, a growing number of serine-threonine kinases highly homologous to the Saccharomyces cerevisiae Ipl1p kinase have been isolated in various organisms. A Drosophila melanogaster homologue, aurora, was the first to be isolated from a multicellular organism. Since then, several related kinases have been found in mammalian cells. They localise to the mitotic apparatus: in the centrosome, at the poles of the bipolar spindle or in the midbody. The kinases are necessary for completion of mitotic events such as centrosome separation, bipolar spindle assembly and chromosome segregation. Extensive research is now focusing on these proteins because the three human homologues are overexpressed in various primary cancers. Furthermore, overexpression of one of these kinases transforms cells. Because of the myriad of kinases identified, we suggest a generic name: Aurora/Ipl1p-related kinase (AIRK). We denote AIRKs with a species prefix and a number, e.g. HsAIRK1. PMID- 10523497 TI - Metalloprotease-disintegrins: modular proteins capable of promoting cell-cell interactions and triggering signals by protein-ectodomain shedding. AB - Metalloprotease-disintegrins (ADAMs) have captured our attention as key players in fertilization and in the processing of the ectodomains of proteins such as tumor necrosis factor (&agr;) (TNF(&agr;)), and because of their roles in Notch mediated signaling, neurogenesis and muscle fusion. ADAMs are integral membrane glycoproteins that contain a disintegrin domain, which is related to snake-venom integrin ligands, and a metalloprotease domain (which can contain or lack a catalytic site). Here, we review and critically discuss current topics in the ADAMs field, including the central role of fertilin in fertilization, the role of the TNF(&agr;) convertase in protein ectodomain processing, the role of Kuzbanian in Notch signaling, and links between ADAMs and processing of the amyloid precursor protein. PMID- 10523498 TI - Ca(2+)/calmodulin and p85 cooperatively regulate an initiation of cytokinesis in Tetrahymena. AB - Tetrahymena p85 differs in mobility in two-dimensional SDS-polyacrylamide gel electrophoresis between wild-type and temperature-sensitive cell-division-arrest mutant cdaA1 cell extracts, and is localized to the presumptive division plane before the formation of the division furrow. The p85 contained three identical sequences which show homology to the calmodulin binding site of Ca(2+)/calmodulin dependent protein kinase Type II in Saccharomyces cerevisiae. We found the p85 directly interacts with Tetrahymena calmodulin in a Ca(2+)-dependent manner, using a co-sedimentation assay. We next examined the localization of p85 and calmodulin during cytokinesis using indirect immunofluorescence. The results showed that both proteins colocalize in the division furrow. This is the first observation that calmodulin is localized in the division furrow. Moreover, the direct interaction between p85 and Ca(2+)/calmodulin was inhibited by Ca(2+)/calmodulin inhibitor N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide HCl. When the cells were treated with the drug just before the beginning of cytokinesis, the drug also inhibited the localization of p85 and calmodulin to the division plane, and the formation of the contractile ring and division furrow. Therefore, we propose that the Ca(2+)/calmodulin signal and its target protein p85 cooperatively regulate an initiation of cytokinesis and may be also concerned with the progression of cytokinesis in Tetrahymena. PMID- 10523499 TI - Substitution of flight muscle-specific actin by human (beta)-cytoplasmic actin in the indirect flight muscle of Drosophila. AB - The human (beta)-cytoplasmic actin differs by only 15 amino acids from Act88F actin which is the only actin expressed in the indirect flight muscle (IFM) of Drosophila melanogaster. To test the structural and functional significance of this difference, we ectopically expressed (beta)-cytoplasmic actin in the IFM of Drosophila that lack endogenous Act88F. When expression of the heterologous actin was regulated by approximately 1.5 kb of the 5' promoter region of the Act88F gene, little (beta)-cytoplasmic actin accumulated in the IFM of the flightless transformants. Including Act88F-specific 5' and 3' untranslated regions (UTRs) yielded transformants that expressed wild-type amounts of (beta)-cytoplasmic actin. Despite the assembly of (beta)-cytoplasmic actin containing thin filaments to which endogenous myosin crossbridges attached, sarcomere organization was deficient, leaving the transformants flightless. Rather than affecting primarily actin-myosin interactions, our findings suggest that the (beta)-cytoplasmic actin isoform is not competent to interact with other actin-binding proteins in the IFM that are involved in the organization of functional myofibrils. PMID- 10523500 TI - Targeted inactivation of the type VII collagen gene (Col7a1) in mice results in severe blistering phenotype: a model for recessive dystrophic epidermolysis bullosa. AB - Dystrophic forms of epidermolysis bullosa (DEB) are associated with mutations in the type VII collagen gene (Col7a1) which encodes the major component of anchoring fibrils. To develop a DEB animal model, type VII collagen deficient mice were generated by targeted homologous recombination. The targeting vector replaced exons 46-69 of Col7a1 with the neomycin-resistance gene, in reverse transcriptional orientation, resulting in elimination of most of the collagenous domain 1. Col7a1 heterozygous (+/-) mice were phenotypically normal. Mating of Col7a1 +/- mice revealed that Col7a1 null (-/-) mice, which were born with extensive cutaneous blistering, died during the first two weeks of life probably due to complications arising from the blistering. Transmission electron microscopy revealed subepidermal blistering below the lamina densa and absence of anchoring fibrils. Immunohistochemical staining with anti-human type VII collagen antibody stained the dermal-epidermal junction in control mice, but did not stain the skin of Col7a1 null mice. Collectively, the DEB mice recapitulate the clinical, genetic, immunohistochemical and ultrastructural characteristics of recessive DEB in humans. These mice provide an animal model to study the pathomechanisms of DEB and serve as a system to test therapeutic approaches, including gene replacement, towards the cure of this devastating skin disease. PMID- 10523501 TI - Cell cycle dependent localization of the telomeric PARP, tankyrase, to nuclear pore complexes and centrosomes. AB - Tankyrase is a human poly(ADP-ribose) polymerase that was initially identified through its interaction with the telomeric protein TRF1, a negative regulator of telomere length. In vitro poly(ADP-ribosyl)ation by tankyrase inhibits TRF1 binding to telomeric DNA suggesting a role for tankyrase in telomere function. We previously demonstrated that tankyrase co-localizes with TRF1 at the ends of human chromosomes in metaphase. Here we show that tankyrase localizes to additional subcellular sites in a cell cycle dependent manner. In interphase, tankyrase co-localized with TRF1 to telomeres, but in addition was found to reside at nuclear pore complexes, as evidenced by indirect immunofluorescence, subcellular fractionation and immunoelectron microscopy. At mitosis, concomitant with nuclear envelope breakdown and nuclear pore complex disassembly, tankyrase was found to relocate around the pericentriolar matrix of mitotic centrosomes. This complex staining pattern along with the observation that tankyrase did not contain a nuclear localization signal suggested that its telomeric localization might be regulated, perhaps by TRF1. Indeed, localization of exogenously expressed tankyrase to telomeres was dependent upon co-transfection with TRF1. These data indicate that the subcellular localization of tankyrase can be regulated by both the cell cycle and TRF1. PMID- 10523502 TI - Inhibition of intravacuolar acidification by antisense RNA decreases osteoclast differentiation and bone resorption in vitro. AB - The role of proton transport and production in osteoclast differentiation was studied in vitro by inhibiting the transcription/translation of carbonic anhydrase II (CA II) and vacuolar H(+)-ATPase (V-ATPase) by antisense RNA molecules. Antisense RNAs targeted against CA II, or the 16 kDa or 60 kDa subunit of V-ATPase were used to block the expression of the specific proteins. A significant decrease in bone resorption rate and TRAP-positive osteoclast number was seen in rat bone marrow cultures and fetal mouse metacarpal cultures after antisense treatment. Intravacuolar acidification in rat bone marrow cells was also significantly decreased after antisense treatment. The CA II antisense RNA increased the number of TRAP-positive mononuclear cells, suggesting inhibition of osteoclast precursor fusion. Antisense molecules decreased the number of monocytes and macrophages, but increased the number of granulocytes in marrow cultures. GM-CSF, IL-3 and IL-6 were used to stimulate haematopoietic stem cell differentiation. The 16 kDa V-ATPase antisense RNA abolished the stimulatory effect of GM-CSF, IL-3 and IL-6 on TRAP-positive osteoclast formation, but did not affect the formation of monocytes and macrophages after IL-3 treatment, or the formation of granulocytes after IL-6 treatment. These results suggest that CA II and V-ATPase are needed, not only for the actual resorption, but also for osteoclast formation in vitro. PMID- 10523503 TI - Endosome fusion in living cells overexpressing GFP-rab5. AB - CHO and BHK cells which overexpress either wild-type rab5 or rab5:Q79L, a constitutively active rab5 mutant, develop enlarged cytoplasmic vesicles that exhibit many characteristics of early endosomes including immunoreactivity for rab5 and transferrin receptor. Time-lapse video microscopy shows the enlarged endosomes arise primarily by fusion of smaller vesicles. These fusion events occur mostly by a 'bridge' fusion mechanism in which the initial opening between vesicles does not expand; instead, membrane flows slowly and continuously from the smaller to the larger endosome in the fusing pair, through a narrow, barely perceptible membranous 'bridge' between them. The unique aspect of rab5 mediated 'bridge' fusion is the persistence of a tight constriction at the site where vesicles merge and we hypothesize that this constriction results from the relatively slow disassembly of a putative docking/fusion complex. To determine the relation of rab5 to the fusion 'bridge', we used confocal fluorescence microscopy to monitor endosome fusion in cells overexpressing GFP-rab5 fusion proteins. Vesicle docking in these cells is accompanied by recruitment of the GFP rab5 into a brightly fluorescent spot in the 'bridge' region between fusing vesicles that persists throughout the entire length of the fusion event and which often persist for minutes following endosome fusion. Other endosomal membrane markers, including FM4-64, are not concentrated in fusion 'bridges'. These results support the idea that the GFP-rab5 spots represent the localized accumulation of GFP-rab5 between fusing endosomes and not simply overlap of adjacent membranes. The idea that the GFP-rab5 spots do not represent membrane overlap is further supported by experiments using photobleaching techniques and confocal imaging which show that GFP-rab5 localized in spots between fusion couplets is resistant to diffusion while GFP-rab5 on endosomal membranes away from these spots rapidly diffuses with a rate constant of about 1.0 (+/-0.3) x10( )(9)cm(2)/second. PMID- 10523504 TI - A targeted gene silencing technique shows that Drosophila myosin VI is required for egg chamber and imaginal disc morphogenesis. AB - We report that Drosophila unconventional myosin VI, encoded by Myosin heavy chain at 95F (Mhc95F), is required for both imaginal disc and egg chamber morphogenesis. During oogenesis, Mhc95F is expressed in migrating follicle cells, including the border cells, which migrate between the nurse cells to lie at the anterior of the oocyte; the columnar cells that migrate over the oocyte; the centripetal cells that migrate between the oocyte and nurse cells; and the dorsal anterior follicle cells, which migrate to secrete the chorionic appendages. Its function during development has been studied using a targeted gene silencing technique, combining the Gal4-UAS targeted expression system and the antisense RNA technique. Antibody staining shows that the expression of myosin 95F is greatly decreased in follicle cells when antisense Mhc95F RNA is expressed. Interfering with expression of Drosophila myosin VI at various developmental stages frequently results in lethality. During metamorphosis it results in adult flies with malformed legs and wings, indicating that myosin VI is essential for imaginal disc morphogenesis. During oogenesis, abnormal follicle cell shapes and aberrant follicle cell migrations are observed when antisense Mhc95F is expressed in follicle cells during stages 9 to 10, suggesting that the Drosophila myosin VI is required for follicle cell epithelial morphogenesis. PMID- 10523505 TI - The suppression of testis-brain RNA binding protein and kinesin heavy chain disrupts mRNA sorting in dendrites. AB - Ribonucleoprotein particles (RNPs) are thought to be key players in somato dendritic sorting of mRNAs in CNS neurons and are implicated in activity-directed neuronal remodeling. Here, we use reporter constructs and gel mobility shift assays to show that the testis brain RNA-binding protein (TB-RBP) associates with mRNPs in a sequence (Y element) dependent manner. Using antisense oligonucleotides (anti-ODN), we demonstrate that blocking the TB-RBP Y element binding site disrupts and mis-localizes mRNPs containing (alpha)-calmodulin dependent kinase II (alpha)-CAMKII) and ligatin mRNAs. In addition, we show that suppression of kinesin heavy chain motor protein alters only the localization of (alpha)-CAMKII mRNA. Thus, differential sorting of mRNAs involves multiple mRNPs and selective motor proteins permitting localized mRNAs to utilize common mechanisms for shared steps. PMID- 10523506 TI - The fission yeast origin recognition complex is constitutively associated with chromatin and is differentially modified through the cell cycle. AB - The origin recognition complex (ORC) binds to the well defined origins of DNA replication in budding yeast. Homologous proteins in other eukaryotes have been identified but are less well characterised. We report here the characterisation of a fission yeast ORC complex (SpORC). Database searches identified a fission yeast Orc5 homologue. SpOrc5 is essential for cell viability and its deletion phenotype is identical to that of two previously identified ORC subunit homologues, SpOrc1 (Orp1/Cdc30) and SpOrc2 (Orp2). Co-immunoprecipitation experiments demonstrate that SpOrc1 forms a complex with SpOrc2 and SpOrc5 and gel filtration chromatography shows that SpOrc1 and SpOrc5 fractionate as high molecular mass complexes. SpORC subunits localise to the nucleus in a punctate distribution which persists throughout interphase and mitosis. We developed a chromatin isolation protocol and show that SpOrc1, 2 and 5 are associated with chromatin at all phases of the cell cycle. While the levels, nuclear localisation and chromatin association of SpORC remain constant through the cell cycle, one of its subunits, SpOrc2, is differentially modified. We show that SpOrc2 is a phosphoprotein which is hypermodified in mitosis and is rapidly converted to a faster migrating isoform as cells proceed into G(1) in preparation for S-phase. PMID- 10523507 TI - Decreasing oncoprotein 18/stathmin levels reduces microtubule catastrophes and increases microtubule polymer in vivo. AB - Oncoprotein 18/stathmin (Op18) has been identified recently as a protein which destabilizes microtubules. To characterize the function of Op18 in living cells, we used microinjection of anti-Op18 antibodies or antisense oligonucleotides to block either Op18 activity or expression in interphase newt lung cells. Anti tubulin staining of cells microinjected with anti-Op18 and fixed 1-2 hours after injection showed an increase in total microtubule polymer. In contrast, microinjection of either non-immune IgG or anti-Op18 preincubated with bacterially-expressed Op18 had little effect on microtubule polymer level. Cells treated with Op18 antisense oligonucleotides for 4 days had (greater than or equal to)50% reduced levels of Op18 with no change in the soluble tubulin level. Measurement of MT polymer level in untreated, antisense or nonsense oligonucleotide treated cells demonstrated that reduced Op18 levels resulted in a 2.5-fold increase in microtubule polymer. Next, the assembly dynamics of individual microtubules at the peripheral regions of living cells were examined using video-enhanced contrast DIC microscopy. Microinjection of antibodies against oncoprotein 18 resulted in a 2.2-fold reduction in catastrophe frequency and a slight reduction in plus end elongation velocity compared to uninjected cells or cells microinjected with non-immune IgG. Preincubation of anti-Op18 antibody with recombinant Op18 greatly diminished the effects of the antibody. Similarly, treatment of cells with antisense oligonucleotides reduced catastrophes 2.5- to 3-fold compared to nonsense oligonucleotide treated or untreated cells. The other parameters of dynamic instability were unchanged after reducing Op18 with antisense oligonucleotides. These studies are consistent with Op18 functioning to regulate microtubule catastrophes during interphase in vivo. PMID- 10523508 TI - VAP-33 localizes to both an intracellular vesicle population and with occludin at the tight junction. AB - Tight junctions create a regulated intercellular seal between epithelial and endothelial cells and also establish polarity between plasma membrane domains within the cell. Tight junctions have also been implicated in many other cellular functions, including cell signaling and growth regulation, but they have yet to be directly implicated in vesicle movement. Occludin is a transmembrane protein located at tight junctions and is known to interact with other tight junction proteins, including ZO-1. To investigate occludin's role in other cellular functions we performed a yeast two-hybrid screen using the cytoplasmic C terminus of occludin and a human liver cDNA library. From this screen we identified VAP-33 which was initially cloned from Aplysia by its ability to interact with VAMP/synaptobrevin and thus was implicated in vesicle docking/fusion. Extraction characteristics indicated that VAP-33 was an integral membrane protein. Antibodies to the human VAP-33 co-localized with occludin at the tight junction in many tissues and tissue culture cell lines. Subcellular fractionation of liver demonstrated that 83% of VAP-33 co-isolated with occludin and DPPIV in a plasma membrane fraction and 14% fractionated in a vesicular pool. Thus, both immunofluorescence and fractionation data suggest that VAP-33 is present in two distinct pools in the cells. In further support of this conclusion, a GFP-VAP-33 chimera also distributed to two sites within MDCK cells and interestingly shifted occludin's localization basally. Since VAP-33 has previously been implicated in vesicle docking/fusion, our results suggest that tight junctions may participate in vesicle targeting at the plasma membrane or alternatively VAP-33 may regulate the localization of occludin. PMID- 10523509 TI - A key function of non-planar membranes and their associated microtubular ribbons in contractile vacuole membrane dynamics is revealed by electrophysiologically controlled fixation of Paramecium. AB - The contractile vacuole complex of the fresh water protozoan Paramecium multimicronucleatum exhibits periodic exocytotic activity. This keeps cytosolic osmolarity at a constant value. The contractile vacuole, the central exocytotic vesicle of the complex, becomes disconnected from its surrounding radial arms and rounds before its fluid content is expelled. We previously proposed a hypothesis that the rounding of the contractile vacuole corresponds to an increase in its membrane tension and that a periodic increase in membrane tension governs the exocytotic cycle. We also proposed a hypothesis that transformation of excess planar membrane of the contractile vacuole into 40 nm diameter tubules, that remain continuous with the contractile vacuole membrane, is a primary cause for the tension development in the planar membrane. In order to investigate tension development further, we have examined electron microscopically the contractile vacuole membrane at the rounding phase. To do this, we developed a computer-aided system to fix the cell precisely at the time that the contractile vacuole exhibited rounding. In this system a decrease in the electrical potential across the contractile vacuole membrane that accompanied the vacuole's rounding was monitored through a fine-tipped microelectrode inserted directly into the in vivo contractile vacuole. A decrease in membrane potential was used to generate an electric signal that activated an injector for injecting a fixative through a microcapillary against the cell at the precise time of rounding. Subsequent electron micrographs of the contractile vacuole membrane clearly demonstrated that numerous approximately 40 nm membrane-bound tubules formed in the vicinity of the vacuole's microtubule ribbons when the vacuole showed rounding. This finding suggested that membrane tubulation was the cause for topographical isolation of excess membrane from the planar membrane during the periodic rounding of the contractile vacuole. This together with stereo-pair images of the contractile vacuole complex membranes suggested that the microtubule ribbons were intimately involved in enhancing this membrane tubulation activity. Electron micrographs of the contractile vacuole complexes also showed that decorated tubules came to lie abnormally close to the contractile vacuole in these impaled cells. This suggested that the contractile vacuole was capable of utilizing the smooth spongiome membrane that lies around the ampullae and the collecting canals to increase its size. PMID- 10523510 TI - Phosphatase 2A and polo kinase, two antagonistic regulators of cdc25 activation and MPF auto-amplification. AB - The auto-catalytic activation of the cyclin-dependent kinase Cdc2 or MPF (M-phase promoting factor) is an irreversible process responsible for the entry into M phase. In Xenopus oocyte, a positive feed-back loop between Cdc2 kinase and its activating phosphatase Cdc25 allows the abrupt activation of MPF and the entry into the first meiotic division. We have studied the Cdc2/Cdc25 feed-back loop using cell-free systems derived from Xenopus prophase-arrested oocyte. Our findings support the following two-step model for MPF amplification: during the first step, Cdc25 acquires a basal catalytic activity resulting in a linear activation of Cdc2 kinase. In turn Cdc2 partially phosphorylates Cdc25 but no amplification takes place; under this condition Plx1 kinase and its activating kinase, Plkk1 are activated. However, their activity is not required for the partial phosphorylation of Cdc25. This first step occurs independently of PP2A or Suc1/Cks-dependent Cdc25/Cdc2 association. On the contrary, the second step involves the full phosphorylation and activation of Cdc25 and the initiation of the amplification loop. It depends both on PP2A inhibition and Plx1 kinase activity. Suc1-dependent Cdc25/Cdc2 interaction is required for this process. PMID- 10523511 TI - The rough deal protein is a new kinetochore component required for accurate chromosome segregation in Drosophila. AB - Mutations in the rough deal (rod) gene of Drosophila greatly increase the missegregation of sister chromatids during mitosis, suggesting a role for this gene product in spindle or kinetochore function. The activity provided by rod also appears to be necessary for the recruitment of two known kinetochore components, Zw10 and cytoplasmic dynein. In this paper we describe the cloning of rough deal and an initial cytological characterization of its product. The Rod protein shares no identifiable structural motif with other known proteins, although apparent homologs exist in the genomes of nematode and man. By immunocytochemistry we show that Rod displays a dynamic intracellular staining pattern, localizing first to kinetochores in prometaphase, but moving to kinetochore microtubules at metaphase. Early in anaphase the protein is once again restricted to the kinetochores, where it persists until the end of telophase. This behavior is in all respects similar to that described for Zw10, and suggests that the proteins function together. PMID- 10523512 TI - Protein transport and flagellum assembly dynamics revealed by analysis of the paralysed trypanosome mutant snl-1. AB - The paraflagellar rod (PFR) of Trypanosoma brucei is a large, complex, intraflagellar structure that represents an excellent system in which to study flagellum assembly. Molecular ablation of one of its major constituents, the PFRA protein, in the snl-1 mutant causes considerable alteration of the PFR structure, leading to cell paralysis. Mutant trypanosomes sedimented to the bottom of the flask rather than staying in suspension but divided at a rate close to that of wild-type cells. This phenotype was complemented by transformation of snl-1 with a plasmid overexpressing an epitope-tagged copy of the PFRA gene. In the snl-1 mutant, other PFR proteins such as the second major constituent, PFRC, accumulated at the distal tip of the growing flagellum, forming a large dilation or 'blob'. This was not assembled as filaments and was removed by detergent extraction. Axonemal growth and structure was unmodified in the snl-1 mutant and the blob was present only at the tip of the new flagellum. Strikingly, the blob of unassembled material was shifted towards the base of the flagellum after cell division and was not detectable when the daughter cell started to produce a new flagellum in the next cell cycle. The dynamics of blob formation and regression are likely indicators of anterograde and retrograde transport systems operating in the flagellum. In this respect, the accumulation of unassembled PFR precursors in the flagellum shows interesting similarities with axonemal mutants in other systems, illustrating transport of components of a flagellar structure during both flagellum assembly and maintenance. Observation of PFR components indicate that these are likely to be regulated and modulated throughout the cell cycle. PMID- 10523513 TI - Profilin is predominantly associated with monomeric actin in Acanthamoeba. AB - We used biochemical fractionation, immunoassays and microscopy of live and fixed Acanthamoeba to determine how much profilin is bound to its known ligands: actin, membrane PIP(2), Arp2/3 complex and polyproline sequences. Virtually all profilin is soluble after gentle homogenization of cells. During gel filtration of extracts on Sephadex G75, approximately 60% of profilin chromatographs with monomeric actin, 40% is free and none voids with Arp2/3 complex or other large particles. Selective monoclonal antibodies confirm that most of the profilin is bound to actin: 65% in extract immunoadsorption assays and 74-89% by fluorescent antibody staining. Other than monomeric actin, no major profilin ligands are detected in crude extracts. Profilin-II labeled with rhodamine on cysteine at position 58 retains its affinity for actin, PIP(2) and poly-L-proline. When syringe-loaded into live cells, it distributes throughout the cytoplasm, is excluded from membrane-bounded organelles, and concentrates in lamellapodia and sites of endocytosis but not directly on the plasma membrane. Some profilin fluorescence appears punctate, but since no particulate profilin is detected biochemically, these spots may be soluble profilin between organelles that exclude profilin. The distribution of profilin in fixed human A431 cells is similar to that in amoebas. Our results show that the major pool of polymerizable actin monomers is complexed with profilin and spread throughout the cytoplasm. PMID- 10523515 TI - Kinesin-mediated transport of neurofilament protein oligomers in growing axons. AB - We examined cytoskeleton-associated forms of NF proteins during axonal neuritogenesis in cultured dorsal root ganglion (DRG) neurons and NB2a/d1 neuroblastoma. In addition to filamentous immunoreactivity, we observed punctate NF immunoreactivity throughout perikarya and neurites. Immuno-electron microscopy revealed this punctate immunoreactivity to consist of non-membrane-bound 75 nm round/ovoid structures consisting of amorphous, fibrous material. Endogenous and microinjected NF subunits incorporated into dots prior to their accumulation within filaments. A transfected GFP-conjugated NF-M incorporated into dots and translocated at a rate consistent with slow axonal transport in real-time video analyses. Some dots converted into a filamentous form or exuded filamentous material during transport. Dots contained conventional kinesin immunoreactivity, associated with microtubules, and their transport into axons was blocked by anti kinesin antibodies and nocodazole. These oligomeric structures apparently represent one form in which NF subunits are transported in growing axons and may utilize kinesin as a transport motor. PMID- 10523514 TI - Expression of human (beta)3-adrenergic receptor induces adipocyte-like features in CHO/K1 fibroblasts. AB - It is reported here that CHO/K1 cells stably transfected with the human (beta)3 AR gene (CHO/K1-(beta)3), grown in the presence of differentiation-stimulating agents accumulate triglycerides. This lipid formation is mediated through the (beta)3 AR, since non-transfected CHO/K1 cells, or cells expressing the human (beta)2 AR, accumulate no significant amount of lipids when grown in supplemented medium. Moreover, lipid production can be inhibited significantly by the (beta) AR antagonist bupranolol. CHO/K1 cells expressing the W64R polymorphism (Trp to Arg polymorphism at position 64 of the human (beta)3 AR), which has been associated with morbid obesity, show increased lipid accumulation as compared to CHO/K1 cells expressing the wild-type (beta)3 AR. Semi-quantitative RT-PCR experiments reveal that a major gene regulating adipocyte differentiation, peroxisome-proliferator-activated-receptor (gamma) (PPAR(gamma)), is expressed in CHO/K1 cells. Concomitantly with the formation of lipid droplets, the expression of PPAR(gamma) mRNA is increased in CHO/K1-(beta)3 cells, but not in non transfected CHO/K1 cells. We furthermore detected constitutive expression of another adipocyte-associated protein: hormone sensitive lipase, while leptin or uncoupling protein-1 transcripts were not expressed. These data suggest that the frequently used CHO/K1 fibroblasts display several preadipocyte-like features, and that the sole expression of the (beta)3 AR modifies the expression of PPAR(gamma) mRNA in these cells, and induces lipid formation under certain culture conditions. PMID- 10523517 TI - Rac GTPases localize at sites of actin reorganization during dynamic remodeling of the cytoskeleton of normal embryonic fibroblasts. AB - Rac GTP-binding proteins are implicated in the dynamic organization of the actin cytoskeleton, and the mechanisms utilized for this purpose are not understood yet. In this paper we have analysed the effects of the expression of Rac proteins on the organization of the cytoskeleton, and their subcellular distribution in chicken embryo fibroblasts. In these cells, overexpression of wild-type Rac GTPases induces disassembly of stress fibers, and production of long, highly branched actin-rich protrusions, with consequent dramatic changes in cell morphology. The formation of these protrusions is mediated by adhesion to the substrate, and is prevented by incubation with anti-(beta)1 function-blocking antibodies. Rac-mediated cell shape changes require a wild-type GTPase, since expression of constitutively active V12-Rac proteins affects actin organization differently in these cells, without causing alterations in their morphology. Localization studies performed on ventral plasma membranes from fibroblasts transfected with wild-type or mutant GTPases show codistribution of Rac along stress fibers, before their disassembly and the formation of the actin-rich protrusions. These data show a link between Rac protein distribution, and their effects on the actin cytoskeleton. Altogether, our results are indicative of an active role of Rac proteins in stress fiber disassembly, and show that Rac, which can cycle its bound nucleotide, produces unique dynamic effects on actin organization. PMID- 10523516 TI - A secreted frizzled related protein, FrzA, selectively associates with Wnt-1 protein and regulates wnt-1 signaling. AB - The Wnt gene family encodes proteins that serve key roles in differentiation and development. Wnt proteins interact with seven transmembrane receptors of the Frizzled family and activate a signaling pathway leading to the nucleus. A primary biochemical effect of Wnt-1 signaling is the stabilization of cytoplasmic (beta)-catenin which, in association with transcription factors of the Lef/tcf family, regulates gene expression. The recent identification of a new class of secreted proteins with similarity to the extracellular, ligand-binding domain of Frizzled proteins, soluble Frizzled related proteins (sFRP), suggested that additional mechanisms could regulate Wnt signaling. Here we demonstrate that FrzA, a sFRP that is highly expressed in vascular endothelium and a variety of epithelium, specifically binds to Wnt-1 protein, but not Wnt-5a protein, and modulates Wnt-1 signaling. FrzA associated with Wnt-1 either when expressed in the same cell or when soluble FrzA was incubated with Wnt-1-expressing cells. FrzA efficiently inhibited the Wnt-1 mediated increase in cytoplasmic (beta) catenin levels as well as the Wnt-1 induction of transcription from a Lef/tcf reporter gene. The effects of FrzA on (beta)-catenin levels could be demonstrated when co-expressed with Wnt-1 or when individual cells expressing FrzA and Wnt-1 were co-cultured. These data demonstrate the existence of a negative regulatory mechanism mediated by the selective binding of FrzA to Wnt-1 protein. PMID- 10523518 TI - Characterization of Dictyostelium discoideum cathepsin D. AB - Previous studies using magnetic purification of Dictyostelium discoideum endocytic vesicles led us to the identification of some major vesicle proteins. Using the same purification procedure, we have now focused our interest on a 44 kDa soluble vesicle protein. Microsequencing of internal peptides and subsequent cloning of the corresponding cDNA identified this protein as the Dictyostelium homolog of mammalian cathepsins D. The only glycosylation detected on Dictyostelium cathepsin D (CatD) is common antigen 1, a cluster of mannose 6 sulfate residues on N-linked oligosaccharide chains. CatD intracellular trafficking has been studied, showing the presence of the protein throughout the entire endocytic pathway. During the differentiation process, the catD gene presents a developmental regulation, which is also observed at the protein level. catD gene disruption does not alter significantly the cell behaviour, either in the vegetative form or the differentiation stage. However, modifications in the SDS-PAGE profiles of proteins bearing common antigen 1 were detected, when comparing parental and catD(-) cells. These modifications point to a possible role of CatD in the maturation of a few Dictyostelium lysosomal proteins. PMID- 10523519 TI - Extreme reduction of chromosome-specific alpha-satellite array is unusually common in human chromosome 21. AB - Human centromeres contain large arrays of alpha-satellite DNA that are thought to provide centromere function. The arrays show size and sequence variation, but the extent to which extremely low levels of this DNA can occur on normal centromeres is unclear. Using a set of chromosome-specific alpha-satellite probes for each of the human chromosomes, we performed interphase fluorescence in situ hybridization (FISH) in a population-screening study. Our results demonstrate that extreme reduction of chromosome-specific alpha satellite is unusually common in chromosome 21 (screened with the alphaRI probe), with a prevalence of 3.70%, compared to < or =0.12% for each of chromosomes 13 and 17, and 0% for the other chromosomes. No analphoid centromere was identified in >17,000 morphologically normal chromosomes studied. All of the low-alphoid centromeres are fully functional as indicated by their mitotic stability and binding to centromere proteins CENP-B, CENP-C, and CENP-E. Sensitive metaphase FISH analysis of the low alphoid chromosome 21 centromeres established the presence of residual alphaRI as well as other non-alphaRI alpha-satellite DNA suggesting that centromere function may be provided by (1) the residual alphaRI DNA, (2) other non-alphaRI alpha satellite sequences, (3) a combination of 1 and 2, or (4) an activated neocentromere DNA. The low-alphoid centromeres, in particular those of chromosome 21, should provide unique opportunities for the study of the evolution and the minimal DNA requirement of the human centromere. PMID- 10523520 TI - Caenorhabditis elegans has scores of hedgehog-related genes: sequence and expression analysis. AB - Previously, we have described novel families of genes, warthog (wrt) and groundhog (grd), in Caenorhabditis elegans. They are related to Hedgehog (Hh) through the carboxy-terminal autoprocessing domain (called Hog or Hint). A comprehensive survey revealed 10 genes with Hog/Hint modules in C. elegans. Five of these are associated with a Wart domain in wrt genes, and three with multiple copies of the Ground domain in grd genes. Both the Wart domain and the Ground domain occur also in genes encoding no Hog domain. Further, we define a new group of genes related to the grd genes, called ground-like (grl). Overall, C. elegans has more than 50 genes belonging to these gene families. Phylogenetic and sequence analysis shows that the wrt, grd, and grl genes are derived from each other. Further examination reveals a sequence motif with similarity to the core of the amino-terminal-signaling domain of Hh proteins. Our data suggest that the wrt, grd, grl, and hh genes are derived from a single ancestral gene. wrt, grd, and grl genes are also present in other nematodes, but so far not in any other phyla. Conversely, hh is not found presently in C. elegans nor other nematodes. Thus, the nematode genes could be the homologs of Hh molecules in other phyla. The membrane molecule Patched has been shown previously to be a receptor of Hh. Many Patched-related proteins are present in C. elegans, which may be targets of the hh-related genes. No Hedgehog-interacting protein (Hip) was found. We analyzed the expression patterns of eight wrt and eight grd genes. The results show that some closely related genes are expressed in the same tissues, but, overall, the expression patterns are diverse, comprising hypodermis, seam cells, the excretory cell, sheath and socket cells, and different types of neurons. PMID- 10523521 TI - Genomic analysis of Caenorhabditis elegans reveals ancient families of retroviral like elements. AB - Retrotransposons are the most abundant and widespread class of eukaryotic transposable elements. The recent genome sequencing of Caenorhabditis elegans has provided an unprecedented opportunity to analyze the evolutionary relationships among the entire complement of retrotransposons within a multicellular eukaryotic organism. In this article we report the results of an analysis of retroviral-like long terminal repeat retrotransposons in C. elegans that indicate that this class of elements may be even more abundant and divergent than previously expected. The unexpected presence, in C. elegans, of an element displaying a number of characteristics previously thought to be unique to vertebrate retroviruses suggests an ancient lineage for this important class of infectious agents. PMID- 10523522 TI - A multilocus genotyping assay for candidate markers of cardiovascular disease risk. AB - A number of chronic diseases, including cardiovascular disease, appear to have a multifactorial genetic risk component. Consequently, techniques are needed to facilitate evaluation of complex genetic risk factors in large cohorts. We have designed a prototype assay for genotyping a panel of 35 biallelic sites that represent variation within 15 genes from biochemical pathways implicated in the development and progression of cardiovascular disease. Each DNA sample is amplified using two multiplex polymerase chain reactions, and the alleles are genotyped simultaneously using an array of immobilized, sequence-specific oligonucleotide probes. This multilocus assay was applied to two types of cohorts. Population frequencies for the markers were estimated using 496 unrelated individuals from a family-based cohort, and the observed values were consistent with previous reports. Linkage disequilibrium between consecutive pairs of markers within the apoCIII, LPL, and ELAM genes was also estimated. A preliminary analysis of single and pairwise locus associations with severity of atherosclerosis was performed using a composite cohort of 142 individuals for whom quantitative angiography data were available; evaluation of the potentially interesting associations observed will require analysis of an independent and larger cohort. This assay format provides a research tool for studies of multilocus genetic risk factors in large cardiovascular disease cohorts, and for the subsequent development of diagnostic tests. PMID- 10523523 TI - Large-scale statistical analyses of rice ESTs reveal correlated patterns of gene expression. AB - Large, publicly available collections of expressed sequence tags (ESTs) have been generated from Arabidopsis thaliana and rice (Oryza sativa). A potential, but relatively unexplored application of this data is in the study of plant gene expression. Other EST data, mainly from human and mouse, have been successfully used to point out genes exhibiting tissue- or disease-specific expression, as well as for identification of alternative transcripts. In this report, we go a step further in showing that computer analyses of plant EST data can be used to generate evidence of correlated expression patterns of genes across various tissues. Furthermore, tissue types and organs can be classified with respect to one another on the basis of their global gene expression patterns. As in previous studies, expression profiles are first estimated from EST counts. By clustering gene expression profiles or whole cDNA library profiles, we show that genes with similar functions, or cDNA libraries expected to share patterns of gene expression, are grouped together. Promising uses of this technique include functional genomics, in which evidence of correlated expression might complement (or substitute for) those of sequence similarity in the annotation of anonymous genes and identification of surrogate markers. The analysis presented here combines the application of a correlation-based clustering method with a graphical color map allowing intuitive visualization of patterns within a large table of expression measurements. PMID- 10523524 TI - Generation and analysis of 25 Mb of genomic DNA from the pufferfish Fugu rubripes by sequence scanning. AB - We have generated and analyzed >50,000 shotgun clones from 1059 Fugu cosmid clones. All sequences have been minimally edited and searched against protein and DNA databases. These data are all displayed on a searchable, publicly available web site at. With an average of 50 reads per cosmid, this is virtually nonredundant sequence skimming, covering 30%-50% of each clone. This essentially random data set covers nearly 25 Mb (>6%) of the Fugu genome and forms the basis of a series of whole genome analyses which address questions regarding gene density and distribution in the Fugu genome and the similarity between Fugu and mammalian genes. The Fugu genome, with eight times less DNA but a similar gene repertoire, is ideally suited to this type of study because most cosmids contain more than one identifiable gene. General features of the genome are also discussed. We have made some estimation of the syntenic relationship between mammals and Fugu and looked at the efficacy of ORF prediction from short, unedited Fugu genomic sequences. Comparative DNA sequence analyses are an essential tool in the functional interpretation of complex vertebrate genomes. This project highlights the utility of using the Fugu genome in this kind of study. PMID- 10523525 TI - G-protein beta3 subunit gene splice variant and body fat distribution in Nunavut Inuit. AB - The GNB3 825T allele encodes a product that has enhanced activation of heterotrimeric G proteins in vitro and could play a role in adipogenesis. We therefore evaluated the possibility that the GNB3 825T allele was associated with obesity in a sample of 213 healthy Canadian Inuit. We found that body weight, body mass index, waist girth, hip girth, subscapular skinfold thickness, and triceps skinfold thickness were significantly higher in subjects with the GNB3 825T/T genotype than in subjects with other genotypes. Furthermore, two anthropometric ratios, namely that of waist to hip circumference and that of subscapular to triceps skinfold thickness, were not significantly different across genotypes. This suggested that the increased deposition of fat in subjects with the GNB3 825T/T genotype was generalized and not localized to particular subregions. There was no association of this genetic variation with blood pressure. The GNB3 825T/T genotype accounted for between 1.6% and 3.3% of the total variation (< or =13% of attributable variation) of the obesity-related traits. The potential for a genetic marker of obesity creates opportunities for future studies in the Inuit, not just to confirm the associations, but also to examine prospectively the influence of interventions and possible relationships between GNB3 825T and longer term complications of obesity. PMID- 10523527 TI - A resource of mapped human bacterial artificial chromosome clones. AB - To date, despite the increasing number of genomic tools, there is no repository of ordered human BAC clones that covers entire chromosomes. This project presents a resource of mapped large DNA fragments that span eight human chromosomes at approximately 1-Mb resolution. These DNA fragments are bacterial artificial chromosome (BAC) clones anchored to sequence tagged site (STS) markers. This clone collection, which currently contains 759 mapped clones, is useful in a wide range of applications from microarray-based gene mapping to identification of chromosomal mutations. In addition to the clones themselves, we describe a database, GenMapDB (http://genomics.med.upenn.edu/genmapdb), that contains information about each clone in our collection. PMID- 10523526 TI - A comprehensive view of human chromosome 1. AB - Comprehensive representations of human chromosomes combining diverse genomic data sets, localizing expressed sequences, and reflecting physical distance are essential for disease gene identification and sequencing efforts. We have developed a method (CompView) for integrating genomic information derived from available cytogenetic, genetic linkage, radiation hybrid, physical, and transcript-based mapping approaches. CompView generates chromosome representations with substantially higher resolution, coverage, and integration than current maps of the human genome. The CompView process was used to build a representation of human chromosome 1, yielding a map with >13,000 unique elements, an effective resolution of 910 kb, and a marker density of 50 kb. CompView creates comprehensive and fully integrated depictions of a chromosome's clinical, biological, and structural information. PMID- 10523529 TI - Using quality measures to facilitate allele calling in high-throughput genotyping. AB - Currently, the main limitation in high-throughput microsatellite genotyping is the required manual editing of allele calls. Even though programs for automated allele calling have been available for several years, they have limited capability because accurate data could only be assured by manual inspection of the electropherograms for confirmation. Here we describe the development of a parametric approach to allele call quality control that eliminates much of the time required for manual editing of the data. This approach was implemented in an editing tool, Decode-GT, that works downstream of the allele calling program, TrueAllele (TA). Decode-GT reads the output data from TA, displays the underlying electropherograms for the genotypes, and sorts the allele calls into three categories: good, bad, and ambiguous. It discards the bad calls, accepts the good calls, and suggests that the user inspect the ambiguous calls, thereby reducing dependence on manual editing. For the categorization we use the following parameters: (1) the quality value for each allele call from TrueAllele; (2) the peak height of the alleles; and (3) the size of the peak shift needed to move peaks into the nearest bin. Here we report how we optimized the parameters such that the size of the ambiguous category was minimized, and both the number of miscalled genotypes in the good category and the useable genotypes in the bad category were negligible. This approach reduces the manual editing time and results in <1% miscalls. PMID- 10523530 TI - Detection of Brucellae in blood cultures. PMID- 10523528 TI - Human genome anatomy: BACs integrating the genetic and cytogenetic maps for bridging genome and biomedicine. AB - Human genome sequencing is accelerating rapidly. Multiple genome maps link this sequence to problems in biology and clinical medicine. Because each map represents a different aspect of the structure, content, and behavior of human chromosomes, these fundamental properties must be integrated with the genome to understand disease genes, cancer instability, and human evolution. Cytogenetic maps use 400-850 visible band landmarks and are the primary means for defining prenatal defects and novel cancer breakpoints, thereby providing simultaneous examination of the entire genome. Recent genetic, physical, and transcript maps use PCR-based landmarks called sequence-tagged sites (STSs). We have integrated these genome maps by anchoring the human cytogenetic to the STS-based genetic and physical maps with 1021 STS-BAC pairs at an average spacing of approximately 1 per 3 Mb. These integration points are represented by 872 unique STSs, including 642 polymorphic markers and 957 bacterial artificial chromosomes (BACs), each of which was localized on high resolution fluorescent banded chromosomes. These BACs constitute a resource that bridges map levels and provides the tools to seamlessly translate questions raised by genomic change seen at the chromosomal level into answers based at the molecular level. We show how the BACs provide molecular links for understanding human genomic duplications, meiosis, and evolution, as well as reagents for conducting genome-wide prenatal diagnosis at the molecular level and for detecting gene candidates associated with novel cancer breakpoints. PMID- 10523531 TI - Phenotypic and phylogenetic characterization of a new Corynebacterium species from dogs: description of Corynebacterium auriscanis sp. nov. AB - Six strains of a previously undescribed catalase-positive coryneform bacterium isolated from clinical specimens from dogs were characterized by phenotypic and molecular genetic methods. Biochemical and chemotaxonomic studies revealed that the unknown bacterium belonged to the genus Corynebacterium sensu stricto. Comparative 16S rRNA gene sequencing showed that the six strains were genealogically highly related and constitute a new subline within the genus Corynebacterium; this subline is close to but distinct from C. falsenii, C. jeikeium, and C. urealyticum. The unknown bacterium from dogs was distinguished from all currently validated Corynebacterium species by phenotypic tests including electrophoretic analysis of whole-cell proteins. On the basis of phylogenetic and phenotypic evidence, it is proposed that the unknown bacterium be classified as a new species, Corynebacterium auriscanis. The type strain of C. auriscanis is CCUG 39938(T). PMID- 10523532 TI - Prevalence of Borrelia burgdorferi and granulocytic and monocytic ehrlichiae in Ixodes ricinus ticks from southern Germany. AB - A total of 287 adult Ixodes ricinus ticks, collected in two regions of southern Germany (Frankonia and Baden-Wurttemberg) where Borrelia burgdorferi infections are known to be endemic, were examined for the presence of 16S ribosomal DNA specific for the Ehrlichia phagocytophila genogroup, E. chaffeensis, E. canis, and B. burgdorferi by nested PCR. Totals of 2.2% (6 of 275) and 21.8% (65 of 275) of the ticks were positive for the E. phagocytophila genogroup and B. burgdorferi, respectively. Two ticks (0.7%) were coinfected with both bacteria. Of 12 engorged I. ricinus ticks collected from two deer, 8 (67%) were positive for the E. phagocytophila genogroup and one (8%) was positive for B. burgdorferi. There was no evidence of infection with E. canis or E. chaffeensis in the investigated tick population. The nucleotide sequences of the 546-bp Ehrlichia PCR products differed at one or two positions from the original sequence of the human granulocytic ehrlichiosis (HGE) agent (S.-M. Chen, J. S. Dumler, J. S. Bakken, and D. H. Walker, J. Clin. Microbiol. 32:589-595, 1994). Three groups of sequence variants were detected; two of these were known to occur in other areas in Europe or the United States, whereas one has not been reported before. Thus, in the German I. ricinus tick population closely related granulocytic ehrlichiae are prevalent, which might represent variants of E. phagocytophila or the HGE agent. PMID- 10523533 TI - Emergence and spread in French hospitals of methicillin-resistant Staphylococcus aureus with increasing susceptibility to gentamicin and other antibiotics. AB - Oxacillin (methicillin) resistance in methicillin-resistant Staphylococcus aureus (MRSA) is associated with an increased incidence of resistance to other antibiotics, which has increased since it was first reported in 1969. In 1992 a new phenotype of MRSA arose in France; this was characterized by a heterogeneous expression of resistance to oxacillin and susceptibility to various antibiotics, including gentamicin but also tetracycline, minocycline, lincomycin, pristinamycin, co-trimoxazole, rifampin, and fusidic acid. In French hospitals a longitudinal nationwide surveillance of antibiotic resistance in S. aureus has allowed for the detection of changes in antibiotic susceptibility profiles. Seven French clinical laboratories (six from the mainland and one from the West Indies) reported the results of susceptibility testing of 57,347 S. aureus strains isolated in their institutes between 1992 and 1998. Over a 7-year period the incidence of isolation of gentamicin-susceptible MRSA (GS-MRSA) strains has steadily increased to represent, in 1998, 46.8 to 94.4% of the MRSA strains, irrespective of the overall incidence of MRSA. Two predominant types recognized by pulsed-field gel electrophoresis (PFGE) accounted for the majority of the GS MRSA in different mainland hospitals, both differing from the predominant type observed in the French West Indies. Some GS-MRSA and gentamicin-resistant MRSA (GR-MRSA) strains had closely related PFGE profiles, and hybridization studies confirmed the lack in GS-MRSA of the aac6'-aph2" gene, which confers resistance to all aminoglycosides, with conservation of the ant4' gene, which confers resistance to kanamycin, tobramycin, and amikacin. Thus, it is likely that certain GS-MRSA strains could have emerged from GR-MRSA strains by excision or deletion of the aac6'-aph2" gene. PMID- 10523535 TI - Value of PCR for detection of Toxoplasma gondii in aqueous humor and blood samples from immunocompetent patients with ocular toxoplasmosis. AB - Toxoplasma gondii infection is an important cause of chorioretinitis in the United States and Europe. Most cases of Toxoplasma chorioretinitis result from congenital infection. Patients are often asymptomatic during life, with a peak incidence of symptomatic illness in the second and third decades of life. Diagnosis is mainly supported by ophthalmological examination and a good response to installed therapy. However, establishment of a diagnosis by ophthalmological examination alone can be difficult in some cases. To determine the diagnostic value of PCR for the detection of T. gondii, 56 blood and 56 aqueous humor samples from 56 immunocompetent patients were examined. Fifteen patients with a diagnosis of ocular toxoplasmosis had increased serum anti-T. gondii immunoglobulin G levels but were negative for anti-T. gondii immunoglobulin M (group 1), and 41 patients were used as controls (group 2). Samples were taken before antiparasitic therapy was initiated, and only one blood sample and one aqueous humor sample were obtained for each patient. Single nested PCRs and Southern blot hybridization were performed with DNA extracted from these samples. The results obtained showed sensitivity and specificity values of 53. 3 and 83%, respectively. Interestingly, among all patients with ocular toxoplasmosis, a positive PCR result with the aqueous humor sample was accompanied by a positive PCR result with the blood sample. This result suggests that ocular toxoplasmosis should not be considered a local event, as PCR testing of blood samples from patients with ocular toxoplasmosis yielded the same result as PCR testing of aqueous humor samples. PCR testing may be useful for discriminating between ocular toxoplasmosis and other ocular diseases, and also can avoid the problems associated with ocular puncture. PMID- 10523534 TI - Case-control study of enteropathogens associated with childhood diarrhea in Dhaka, Bangladesh. AB - The International Centre for Diarrhoeal Disease Research, Bangladesh, is a major center for research into diarrheal diseases. The center treats more than 100,000 patients a year. To obtain useful information representative of all patients, a surveillance system in which a 4% systematic sample of all patients is studied in detail, including etiological agents of diarrhea, was installed in October 1979. The first paper on etiology for the surveillance patients was published in 1982, which identified a potential enteric pathogen in 66% of patients. In subsequent years, several new agents of diarrhea have been identified. To assess the importance of a broader spectrum of diarrheal agents including the ones identified relatively recently, we studied 814 children with diarrhea. The children were up to 5 years of age and were part of the surveillance system. They were matched with an equal number of community controls without diarrhea. The study was conducted from February 1993 to June 1994. A potential enteric pathogen was isolated from 74.8% of diarrheal children and 43.9% of control children (P = 0.0001). Even though the first study was not a case-control study, it identified rotavirus, Campylobacter jejuni, enterotoxigenic Escherichia coli, Shigella spp. , and Vibrio cholerae O1 as major pathogens. The present study identified these pathogens as being significantly associated with diarrhea. In addition, the study also identified six additional agents, including enteropathogenic E. coli, Aeromonas spp., V. cholerae O139, enterotoxigenic Bacteroides fragilis, Clostridium difficile, and Cryptosporidium parvum, as being significantly associated with diarrhea. Plesiomonas shigelloides, Salmonella spp., diffusely adherent E. coli, enteroaggregative E. coli, Entamoeba histolytica, and Giardia lamblia were not significantly associated with diarrhea. Enteroinvasive E. coli, enterohemorrhagic E. coli, and Cyclospora cayetanensis were not detected in any of the children. The major burden of diseases due to most pathogens occurred in the first year of life. As in the previous study, seasonal patterns were seen for diarrhea associated with rotavirus, V. cholerae, and enterotoxigenic E. coli, and infections with multiple pathogens were common. With a few exceptions, these findings are in agreement with those from other developing countries. This knowledge of a broader spectrum of etiological agents of diarrhea in the surveillance patients will help us plan studies into various aspects of diarrheal diseases in this population. PMID- 10523536 TI - Invasive and noninvasive group A streptococcal isolates with different speA alleles in The Netherlands: genetic relatedness and production of pyrogenic exotoxins A and B. AB - Streptococcal pyrogenic exotoxin A (SPE-A) and SPE-B have been implicated in the pathogenesis of severe group A streptococcal (GAS) disease. We studied 31 invasive GAS strains including 18 isolates from patients with toxic shock syndrome and 22 noninvasive strains isolated in The Netherlands between 1994 and 1998. These strains were associated with the different allelic variants of the gene encoding SPE-A. We selected endemic strains with speA-positive M and T serotypes: speA2-associated M1T1 and M22-60T12 strains, speA3-associated M3T3 strains, and speA4-associated M6T6 strains. Since speA1-positive isolates were not frequently encountered, we included speA1 strains of different serotypes. The GAS strains were compared genotypically by pulsed-field gel electrophoresis and phenotypically by the in vitro production of SPE-A and SPE-B. All strains within one M and T type appeared to be of clonal origin. Most strains produced SPE-A and SPE-B, but only a minority of the speA4-positive isolates did so. Among our isolates, speA1- and speA3-positive strains produced significantly more SPE-A than speA2- and speA4-carrying strains, while SPE-B production was most pronounced among speA1- and speA2-containing strains. There was a marked degree of variability in the amounts of exotoxins produced in vitro by strains that shared the same genetic profile. We conclude that the differences in the in vitro production of SPE-A and SPE-B between our selected strains with identical M and T types were not related to either genetic heterogeneity or the clinical course of GAS disease in the patient from whom they were isolated. PMID- 10523537 TI - Cloning and expression of a 48-kilodalton Babesia caballi merozoite rhoptry protein and potential use of the recombinant antigen in an enzyme-linked immunosorbent assay. AB - A cDNA expression library prepared from Babesia caballi merozoite mRNA was screened with a monoclonal antibody BC11D against the rhoptry protein of B. caballi merozoite. A cDNA encoding a 48-kDa protein of B. caballi was cloned and designated BC48. The complete nucleotide sequence of the BC48 gene had 1,828 bp and was shown to contain no intron. Southern blotting analysis indicated that the BC48 gene contained more than two copies in the B. caballi genome. Computer analysis suggested that this sequence contained an open reading frame of 1,374 bp with a coding capacity of approximately 52 kDa. The recombinant protein expressed by the vaccinia virus vector in horse cells had an apparent molecular mass of 48 kDa, which was the same as that of the native B. caballi 48-kDa protein. Moreover, recombinant proteins expressed by the pGEX4T expression vector in Escherichia coli as glutathione S-transferase fusion proteins were used for antigen in an enzyme-linked immunosorbent assay (ELISA). The ELISA was able to differentiate very clearly between B. caballi-infected horse sera and B. equi infected horse sera or noninfected normal horse sera. These results suggest that this simple and highly sensitive test might be applicable to the detection of B. caballi-infected horses in the field. PMID- 10523538 TI - Biotyping and virulence properties of skin isolates of Candida parapsilosis. AB - The biotype and virulence of skin isolates of Candida parapsilosis were compared with blood isolates of the same fungus. Morphotype, resistotype, and electrophoretic karyotype determinations did not reveal any special cluster with a unique or dominant pathogenic feature among all of the isolates, regardless of their source. However, all cutaneous isolates had uniformly elevated secretory aspartyl-protease (Sap) activity, more than four times higher than the enzyme activity of the blood isolates. They were also highly vaginopathic in a rat vaginitis model, being significantly more virulent than blood isolates in this infection model. In contrast, skin isolates were nonpathogenic in systemic infection of cyclophosphamide-immunodepressed mice, while some blood isolates were, in this model, highly pathogenic (median survival time, 2 days, with internal organ invasion at autopsy). Finally, skin isolates did not differ, as a whole, from blood isolates in their adherence to plastic. This property was associated with a morphotype, as defined by a colony with continuous fringe, which was present among both skin and blood isolates. While confirming the genetic heterogenicity of C. parapsilosis, our data strongly suggest that the potential of this fungus to cause mucosal disease is associated with Sap production and is substantially distinct from that of systemic invasion. PMID- 10523539 TI - Detection of specific immunoglobulin E during maternal, fetal, and congenital toxoplasmosis. AB - Toxoplasma immunoglobulin E (IgE) antibodies in 664 serum samples were evaluated by using an immunocapture method with a suspension of tachyzoites prepared in the laboratory in order to evaluate its usefulness in the diagnosis of acute Toxoplasma gondii infection during pregnancy, congenital infection, and progressive toxoplasmosis. IgE antibodies were never detected in sera from seronegative women, from patients with chronic toxoplasma infection, or from infants without congenital toxoplasmosis. In contrast, they were detected in 86.6% of patients with toxoplasmic seroconversion, and compared with IgA and IgM, the short kinetics of IgE was useful to date the infection precisely. For the diagnosis of congenital toxoplasmosis, specific IgE detected was less frequently than IgM or IgA (25 versus 67.3%), but its detection during follow-up of children may be interesting, reflecting an immunological rebound. Finally, IgE was detected early and persisted longer in progressive toxoplasmosis with cervical adenopathies, so it was also a good marker of the evolution of toxoplasma infection. PMID- 10523540 TI - Escherichia coli O157:H7 and O157:H(-) strains that do not produce Shiga toxin: phenotypic and genetic characterization of isolates associated with diarrhea and hemolytic-uremic syndrome. AB - We have isolated one sorbitol-nonfermenting (SNF) Escherichia coli O157:H7 isolate and five sorbitol-fermenting (SF) E. coli O157:H(-) isolates that do not contain Shiga toxin (Stx) genes (stx). Isolates originated from patients with diarrhea (n = 4) and hemolytic-uremic syndrome (HUS) (n = 2). All isolates harbored a chromosomal eae gene encoding gamma-intimin as well as the plasmid genes E-hly and etp. The E. coli O157:H7 isolate was katP and espP positive. Respective sera obtained from the patient with HUS contained antibodies to the O157 lipopolysaccharide antigen. The stx-negative E. coli O157:H7 isolate is genetically related to stx-positive SNF E. coli O157:H7. All stx-negative SF E. coli O157:H(-) isolates belong to the same genetic cluster and are closely related to stx-positive SF E. coli O157:H(-) isolates. Our data indicate that stx negative E. coli O157:H7/H(-) variants may occur at a low frequency and cannot be recognized by diagnostic methods that target Stx. PMID- 10523541 TI - Development of a PCR assay for rapid detection of enterococci. AB - Enterococci are becoming major nosocomial pathogens, and increasing resistance to vancomycin has been well documented. Conventional identification methods, which are based on culturing, require 2 to 3 days to provide results. PCR has provided a means for the culture-independent detection of enterococci in a variety of clinical specimens and is capable of yielding results in just a few hours. However, all PCR-based assays developed so far are species specific only for clinically important enterococci. We have developed a PCR-based assay which allows the detection of enterococci at the genus level by targeting the tuf gene, which encodes elongation factor EF-Tu. Initially, we compared the nucleotide sequences of the tuf gene from several bacterial species (available in public databases) and designed degenerate PCR primers derived from conserved regions. These primers were used to amplify a target region of 803 bp from four enterococcal species (Enterococcus avium, E. faecalis, E. faecium, and E. gallinarum). Subsequently, the complete nucleotide sequences of these amplicons were determined. The analysis of a multiple alignment of these sequences revealed regions conserved among enterococci but distinct from those of other bacteria. PCR primers complementary to these regions allowed amplification of genomic DNAs from 14 of 15 species of enterococci tested (E. solitarius DNA could not be amplified). There was no amplification with a majority of 79 nonenterococcal bacterial species, except for 2 Abiotrophia species and several Listeria species. Furthermore, this assay efficiently amplified all 159 clinical isolates of enterococci tested (61 E. faecium, 77 E. faecalis, 9 E. gallinarum, and 12 E. casseliflavus isolates). Interestingly, the preliminary sequence comparison of the amplicons for four enterococcal species demonstrated that there were some sequence variations which may be used to generate species-specific internal probes. In conclusion, this rapid PCR-based assay is capable of detecting all clinically important enterococci and has potential for use in clinical microbiology laboratories. PMID- 10523542 TI - Improved multiplex PCR using conserved and species-specific 16S rRNA gene primers for simultaneous detection of Actinobacillus actinomycetemcomitans, Bacteroides forsythus, and Porphyromonas gingivalis. AB - Among putative periodontal pathogens, Actinobacillus actinomycetemcomitans, Bacteroides forsythus, and Porphyromonas gingivalis are most convincingly implicated as etiological agents in periodontitis. Therefore, techniques for detection of those three species would be of value. We previously published a description of a multiplex PCR that detects A. actinomycetemcomitans and P. gingivalis. The present paper presents an improvement on that technique, which now allows more sensitive detection of all three periodontal pathogens. Sensitivity was determined by testing serial dilutions of A. actinomycetemcomitans, B. forsythus, and P. gingivalis cells. Primer specificity was tested against (i) all gene sequences from the GenBank-EMBL database, (ii) six A. actinomycetemcomitans, one B. forsythus, and four P. gingivalis strains, (iii) eight different species of oral bacteria, and (iv) supra- and subgingival plaque samples from 20 healthy subjects and subgingival plaque samples from 10 patients with periodontitis. The multiplex PCR had a detection limit of 10 A. actinomycetemcomitans, 10 P. gingivalis, and 100 B. forsythus cells. Specificity was confirmed by the fact that (i) none of our forward primers were homologous to the 16S rRNA genes of other oral species, (ii) amplicons of predicted size were detected for all A. actinomycetemcomitans, B. forsythus, and P. gingivalis strains tested, and (iii) no amplicons were detected for the eight other bacterial species. A. actinomycetemcomitans, B. forsythus, and P. gingivalis were detected in 6 of 20, 1 of 20, and 11 of 20 of supragingival plaque samples, respectively, and 4 of 20, 7 of 20, and 13 of 20 of subgingival plaque samples, respectively, from periodontally healthy subjects. Among patients with periodontitis, the organisms were detected in 7 of 10, 10 of 10, and 7 of 10 samples, respectively. The simultaneous detection of three periodontal pathogens is an advantage of this technique over conventional PCR assays. PMID- 10523543 TI - Near absence of vancomycin-resistant enterococci but high carriage rates of quinolone-resistant ampicillin-resistant enterococci among hospitalized patients and nonhospitalized individuals in Sweden. AB - Rates of colonization with enterococci with acquired resistance to vancomycin (vancomycin-resistant enterococci [VRE]) and ampicillin (ampicillin-resistant enterococci [ARE]) were determined by using fecal samples from 670 nonhospitalized individuals and 841 patients in 27 major hospitals. Of the hospitalized patients, 181 (21.5%) were carriers of ARE and 9 (1.1%) were carriers of VRE. In univariate analyses, length of hospital stay (odds ratio [OR], 4.6; 95% confidence interval [CI], 2.5 to 8.9) and antimicrobial therapy (OR, 4.7; 95% CI, 3.3 to 6.7) were associated with ARE colonization, as were prior treatment with penicillins (OR, 3.1; 95% CI, 1.8 to 5. 5), cephalosporins (OR, 2.9; 95% CI, 1.7 to 5.0), or quinolones (OR, 2.7; 95% CI, 1.5 to 4.7). In logistic regression analysis, antimicrobial therapy for at least 5 days was independently associated with ARE carriage (adjusted OR, 3.8; 95% CI, 2.6 to 5.4). Over 90% of the ARE isolates were fluoroquinolone resistant, whereas 14% of the ampicillin-susceptible Enterococcus faecium isolates were fluoroquinolone resistant. ARE carriage rates correlated with the use of fluoroquinolones (P = 0.04) but not with the use of ampicillin (P = 0.68) or cephalosporins (P = 0.40). All nine VRE isolates were E. faecium vanB and were found in one hospital. Seven of these isolates were related according to their types as determined by pulsed field gel electrophoresis. Among the nonhospitalized individuals, the ARE carriage rate was lower (6%; P < 0.05), and only one person, who had recently returned from Africa, harbored VRE (E. faecium vanA). The absence of VRE colonization in nonhospitalized individuals reflects an epidemiological situation in Sweden radically different from that in countries in continental Europe where glycopeptides have been widely used for nonmedical purposes. PMID- 10523544 TI - Analysis by multiplex PCR of the physical status of human papillomavirus type 16 DNA in cervical cancers. AB - Integration of human papillomavirus (HPV) DNA occurs early in cancer development and is an important event in malignant transformation of cervical cancer. Integration of HPVs preferentially disrupts or deletes the E2 open reading frame, which results in the loss of its expression. The preferential disruption of the E2 gene causes the absence of the E2 gene sequences in the PCR product following integration. Twenty-two carcinomas positive for HPV type 16 (HPV-16) DNA were first tested for the disruption of the E2 gene by PCR. A specific fragment of the E2 gene was not amplified in 10 cases, suggesting integration of HPV DNA into the host genome. Next, multiplex PCR for the HPV E2 and E6 genes was carried out in the remaining 12 cases. Copy numbers of both genes should be equivalent in episomal forms, while the E2 gene copy number will be smaller than that for E6 following the preferential disruption of the E2 gene in concomitant forms. Although relative ratios of HPV E2 to E6 PCR products (E2/E6 ratios) ranged from 1.40 to 2.34 in 10 of 12 cases, multiplex PCR products from 2 cases displayed extremely low ratios of 0.69 and 0.61. Southern blot hybridization with an HPV-16 probe revealed that only in these two cases was both episomal and integrated HPV DNA being carried simultaneously. Thus, multiplex PCR for the E2 and E6 genes of HPV-16 DNA following PCR for the E2 gene can distinguish the pure episomal form from a mixed form of episomal and integrated HPV DNA. Clinical application of this technique will help researchers to understand the implication of the integration of HPV DNA for cervical carcinogenesis and cervical cancer progression. PMID- 10523545 TI - Amplification of reiterated sequences of herpes simplex virus type 1 (HSV-1) genome to discriminate between clinical HSV-1 isolates. AB - Herpes simplex virus type 1 (HSV-1)-related disease ranges from a localized, self limiting illness to fatal disease in immunocompromised individuals. The corneal disease herpetic keratitis may develop after reactivation of a latent virus or reinfection with an exogenous herpesvirus. Molecular analysis of the virus involved may allow distinction between these two options. The HSV-1 genome contains several hypervariable regions that vary in numbers of reiterating regions (reiterations I to VIII [ReI to ReVIII]) between individual strains. Twenty-four HSV-1 clones, derived by subcloning of HSV-1 (strain F) twice in limiting dilutions, were tested in a PCR-based assay to analyze the stabilities of ReI, ReIII, ReIV, and ReVII. ReI and ReIII proved to vary in size upon subcloning, whereas ReIV and ReVII were stable. Subsequently, 37 unrelated isolates and 10 sequential isolates from five patients, all with HSV-1-induced keratitis, were genotyped for ReIV and ReVII. Of the 37 unrelated samples, 34 (92%) could be discriminated, while the genotypes of the viruses in sequential samples were identical for each individual. Conclusively, the data show that the approach presented allows the rapid and accurate discrimination of HSV-1 strains in studies that address the transmission and pathogenesis of HSV-1 infections. PMID- 10523546 TI - Evaluation of reverse transcription-PCR and a bacteriophage-based assay for rapid phenotypic detection of rifampin resistance in clinical isolates of Mycobacterium tuberculosis. AB - New rapid phenotypic assays for the detection of rifampin resistance in Mycobacterium tuberculosis have recently been described, but most of these require liquid cultures, which reduces the utility of many tests in terms of turnaround times. In the United Kingdom, over 90% of rifampin-resistant isolates are also resistant to isoniazid, so rifampin resistance can be used as a sensitive marker for multidrug-resistant tuberculosis. In this study, two new rapid phenotypic assays were compared to the standard resistance ratio method on 91 clinical isolates of M. tuberculosis. One, the phage amplified biologically (PhaB) assay, has been described previously and is based on the inability of susceptible isolates of M. tuberculosis to support the replication of bacteriophage D29 in the presence of inhibitory doses of rifampin. The other employed reverse transcription (RT)-PCR to demonstrate a reduction in inducible dnaK mRNA levels in susceptible isolates treated with rifampin. After incubation for 18 h with 4 microg of rifampin per ml, the PhaB assay showed concordance with the resistance ratio method for 46 of 46 (100%) susceptible and 31 of 31 (100%) resistant isolates, while RT-PCR showed concordance for 46 of 48 (96%) susceptible and 35 of 36 (97%) resistant isolates. We believe these assays provide a reliable rapid means of susceptibility testing with a total turnaround time of only 48 h, although the PhaB assay is better in terms of its lower technical demand and cost and its applicability to tuberculosis susceptibility testing in developing countries. PMID- 10523547 TI - Evaluation of a bacteriophage-based assay (phage amplified biologically assay) as a rapid screen for resistance to isoniazid, ethambutol, streptomycin, pyrazinamide, and ciprofloxacin among clinical isolates of Mycobacterium tuberculosis. AB - Rapid molecular assays for the detection of mutations associated with rifampin resistance in Mycobacterium tuberculosis are commercially available. However, they are complex and expensive and have predictive values of 90 to 95%. Molecular assays for other drugs are less predictive of resistance. Ideally, assays based on phenotypic markers should be used for susceptibility testing, but these can take weeks to complete. We previously described a rapid phenotypic assay, the phage amplified biologically (PhaB) assay, for the rapid determination of rifampin and isoniazid susceptibility in clinical isolates of M. tuberculosis. In this study, we extended the assay to the study of ethambutol, pyrazinamide, streptomycin, and ciprofloxacin. After the optimization of antibiotic concentrations and incubation conditions, the assay was applied to each drug for a total of 157 isolates. The correlations between the results of the PhaB assay and the resistance ratio method were 94% for isoniazid, 96% for streptomycin, 100% for ciprofloxacin, 88% for ethambutol, and 87% for pyrazinamide. For ciprofloxacin, ethambutol, and pyrazinamide, significantly better correlations were found when a 90% reduction in plaque count was used as the cutoff. Turnaround times for the PhaB assay were 2 to 3 days, compared with 10 days for the resistance ratio method. We believe that this low-cost assay may have widespread applicability for the rapid screening of drug resistance in M. tuberculosis isolates, especially in developing countries. PMID- 10523548 TI - Rapid identification of Candida dubliniensis with commercial yeast identification systems. AB - Candida dubliniensis is a newly described species that is closely related phylogenetically to Candida albicans and that is commonly associated with oral candidiasis in human immunodeficiency virus-positive patients. Several recent studies have attempted to elucidate phenotypic and genotypic characteristics of use in separating the two species. However, results obtained with simple phenotypic tests were too variable and tests that provided more definitive data were too complex for routine use in the clinical laboratory setting. The objective of this study was to determine if reproducible identification of C. dubliniensis could be obtained with commercial identification kits. The substrate reactivity profiles of 80 C. dubliniensis isolates were obtained by using the API 20C AUX, ID 32 C, RapID Yeast Plus, VITEK YBC, and VITEK 2 ID-YST systems. The percentages of C. dubliniensis isolates capable of assimilating or hydrolyzing each substrate were compared with the percentages from the C. albicans profiles in each kit's database, and the results were expressed as percent C. dubliniensis and percent C. albicans. Any substrate that showed >50% difference in reactivity was considered useful in differentiating the species. In addition, assimilation of methyl-alpha-D-glucoside (MDG), D-trehalose (TRE), and D-xylose (XYL) by the same isolates was investigated by the traditional procedure of Wickerham and Burton (L. J. Wickerham and K. A. Burton, J. Bacteriol. 56:363-371, 1948). At 48 h (the time recommended by the manufacturer for its new database), we found that the assimilation of four carbohydrates in the API 20C AUX system could be used to distinguish the species, i.e., glycerol (GLY; 88 and 14%), XYL (0 and 88%), MDG (0 and 85%), and TRE (15 and 97%). Similarly, results with the ID 32 C system at 48 h showed that XYL (0 and 98%), MDG (0 and 98%), lactate (LAT; 0 and 96%), and TRE (30 and 96%) could be used to separate the two species. Phosphatase (PHS; 9 and 76%) and alpha-D-glucosidase (23 and 94%) proved to be the most useful for separation of the species in the RapID Yeast Plus system. While at 24 h the profiles obtained with the VITEK YBC system showed that MDG (10 and 95%), XYL (0 and 95%), and GLY (26 and 80%) could be used to separate the two species, at 48 h only XYL (6 and 95%) could be used to separate the two species. The most useful substrates in the VITEK 2 ID-YST system were TRE (1 and 89%), MDG (1 and 99%), LAT (4 and 98%), and PHS (83 and 1%). While the latter kit was not yet commercially available at the time of the study, it would appear to be the most valuable for the identification of C. dubliniensis. Although assimilation of MDG, TRE, and XYL proved to be the most useful for species differentiation by the majority of commercial systems, the results with these carbohydrates by the Wickerham and Burton procedure were essentially the same for both species, albeit following protracted incubation. Thus, it is the rapidity of the assimilation achieved with the commercial systems that allows the differentiation of C. dubliniensis from C. albicans. PMID- 10523549 TI - Serodiagnosis of human granulocytic ehrlichiosis by a recombinant HGE-44-based enzyme-linked immunosorbent assay. AB - Current antibody testing for human granulocytic ehrlichiosis relies predominantly on indirect fluorescent-antibody assays and immunoblot analysis. Shortcomings of these techniques include high cost and variability of test results associated with the use of different strains of antigens derived from either horses or cultured HL-60 cells. We used recombinant protein HGE-44, expressed and purified as a maltose-binding protein (MBP) fusion peptide, as an antigen in a polyvalent enzyme-linked immunosorbent assay (ELISA). Fifty-five normal serum samples from healthy humans served as a reference to establish cutoff levels. Thirty-three of 38 HGE patient serum samples (87%), previously confirmed by positive whole-cell immunoblotting, reacted positively in the recombinant ELISA. In specificity analyses, serum samples from patients with Lyme disease, syphilis, rheumatoid arthritis, and human monocytic ehrlichiosis (HME) did not react with HGE-44-MBP antigen, except for one sample (specificity, 98%). We conclude that recombinant HGE-44 antigen is a suitable antigen in an ELISA for the laboratory diagnosis and epidemiological study of HGE. PMID- 10523550 TI - Degenerate and nested PCR: a highly sensitive and specific method for detection of human papillomavirus infection in cutaneous warts. AB - The role of human papillomavirus (HPV) in anogenital carcinogenesis is firmly established, but evidence that supports a similar role in skin remains speculative. Immunosuppressed renal transplant recipients have an increased incidence of viral warts and nonmelanoma skin cancer, and the presence of HPV DNA in these lesions, especially types associated with the condition epidermodysplasia verruciformis (EV), has led to suggestions that HPV may play a pathogenic role. However, differences in the specificities and sensitivities of techniques used to detect HPV in skin have led to wide discrepancies in the spectrum of HPV types reported. We describe a degenerate nested PCR technique with the capacity to detect a broad spectrum of cutaneous, mucosal, and EV HPV types. In a series of 51 warts from 23 renal transplant recipients, this method detected HPV DNA in all lesions, representing a significant improvement over many previously published studies. Cutaneous types were found in 84.3% of warts and EV types were found in 80.4% of warts, whereas mucosal types were detected in 27.4% of warts. In addition, the method allowed codetection of two or more distinct HPV types in 94.1% of lesions. In contrast, single HPV types were detected in all but 1 of 20 warts from 15 immunocompetent individuals. In summary, we have established a highly sensitive and comprehensive degenerate PCR methodology for detection and genotyping of HPV from the skin and have demonstrated a diverse spectrum of multiple HPV types in cutaneous warts from transplant recipients. Studies designed to assess the significance of these findings to cutaneous carcinogenesis are under way. PMID- 10523551 TI - Evaluation of protein A gene polymorphic region DNA sequencing for typing of Staphylococcus aureus strains. AB - Three hundred and twenty isolates of Staphylococcus aureus were typed by DNA sequence analysis of the X region of the protein A gene (spa). spa typing was compared to both phenotypic and molecular techniques for the ability to differentiate and categorize S. aureus strains into groups that correlate with epidemiological information. Two previously characterized study populations were examined. A collection of 59 isolates (F. C. Tenover, R. Arbeit, G. Archer, J. Biddle, S. Byrne, R. Goering, G. Hancock, G. A. Hebert, B. Hill, R. Hollis, W. R. Jarvis, B. Kreiswirth, W. Eisner, J. Maslow, L. K. McDougal, J. M. Miller, M. Mulligan, and M. A. Pfaller, J. Clin. Microbiol. 32:407-415, 1994) from the Centers for Disease Control and Prevention (CDC) was used to test for the ability to discriminate outbreak from epidemiologically unrelated strains. A separate collection of 261 isolates form a multicenter study (R. B. Roberts, A. de Lencastre, W. Eisner, E. P. Severina, B. Shopsin, B. N. Kreiswirth, and A. Tomasz, J. Infect. Dis. 178:164-171, 1998) of methicillin-resistant S. aureus in New York City (NYC) was used to compare the ability of spa typing to group strains along clonal lines to that of the combination of pulsed-field gel electrophoresis and Southern hybridization. In the 320 isolates studied, spa typing identified 24 distinct repeat types and 33 different strain types. spa typing distinguished 27 of 29 related strains and did not provide a unique fingerprint for 4 unrelated strains from the four outbreaks of the CDC collection. In the NYC collection, spa typing provided a clonal assignment for 185 of 195 strains within the five major groups previously described. spa sequencing appears to be a highly effective rapid typing tool for S. aureus that, despite some expense of specificity, has significant advantages in terms of speed, ease of use, ease of interpretation, and standardization among laboratories. PMID- 10523552 TI - Comparison of direct and concentrated acid-fast smears to identify specimens culture positive for Mycobacterium spp. AB - Microscopic examination of respiratory specimens for acid-fast bacilli (AFB) plays a key role in the initial diagnosis of tuberculosis, monitoring of treatment, and determination of eligibility for release from isolation. The objective of this study was to compare the sensitivity obtained with smears for detection of AFB (AFB smears) made directly from respiratory specimens (direct AFB smears) to that obtained with parallel smears made from concentrates of the specimens (concentrated AFB smears). A total of 2,693 specimens were evaluated; 1,806 were from the University of California Irvine Medical Center Medical Microbiology Laboratory (UCIMC), which serves a tertiary-care hospital with outpatient clinics, and 887 were from the Microbial Disease Laboratory at the California Department of Public Health (MDL), which receives specimens from outpatient facilities and clinics on Pacific islands. Of the 353 AFB culture positive specimens at UCIMC, there was a statistically significant difference in the sensitivity of the direct AFB smear (34%) and that of the smear made from the concentrated specimen (58%) (P < 0.05). This was also true for the 208 specimens positive for Mycobacterium tuberculosis, for which the sensitivity of the direct smear was 42% (87 of 208) and that for the smear made from the concentrated specimen was 74% (154 of 208). At MDL, where all but 1 of the 45 culture-positive specimens grew M. tuberculosis, the sensitivity of the smear made from the concentrated specimen was 93% (42 of 45) and was not significantly higher than the sensitivity of the direct smear, which was 82% (37 of 45). By combining the results from both laboratories, 42 patients from whom at least three specimens were received were culture positive for M. tuberculosis. The cumulative results for the initial three specimens from these patients showed that the direct smear detected M. tuberculosis in 81% of these patients, whereas the smear made from the concentrate detected M. tuberculosis in 91% of these patients. In summary, when all culture-positive specimens are considered, the sensitivity of the direct smear compared to that of a smear made from the concentrated specimen was significantly different overall in the two different laboratory settings. However, this difference was reduced only if the cumulative results for the initial three specimens received from patients who were culture positive for M. tuberculosis were evaluated. PMID- 10523553 TI - Evaluation of the prototype Roche DNA amplification kit incorporating the new SSK145 and SKCC1B primers in detection of human immunodeficiency virus type 1 DNA in Zimbabwe. AB - We assessed the sensitivity and specificity of a newly developed DNA PCR kit (Roche Diagnostic Corporation, Indianapolis, Ind.) that incorporates primers for all the group M viruses for the detection of human immunodeficiency virus (HIV) type 1 (HIV-1) infection in Zimbabwe. A total of 202 whole-blood samples from adults whose HIV status was known were studied. This included 100 HIV-1-positive and 102 HIV-1-negative samples selected on the basis of concordant results obtained with two enzyme-linked immunosorbent assay kits. The prototype Roche DNA PCR assay had a 100% sensitivity for the detection of HIV-1 DNA and a specificity of 100%. We conclude that the new Roche DNA PCR kit is accurate for the detection of HIV DNA in Zimbabwean samples, in which HIV-1 subtype C dominates. PMID- 10523554 TI - Detection of decreased fluoroquinolone susceptibility in Salmonellas and validation of nalidixic acid screening test. AB - We evaluated 1,010 Salmonella isolates classified as fluoroquinolone susceptible according to the National Committee for Clinical Laboratory Standards guidelines for susceptibility to nalidixic acid and three fluoroquinolones. These isolates were divided into two distinct subpopulations, with the great majority (n = 960) being fully ciprofloxacin susceptible and a minority (n = 50) exhibiting reduced ciprofloxacin susceptibility (MICs ranging between 0.125 and 0.5 microg/ml). The less ciprofloxacin-susceptible isolates were uniformly resistant to nalidixic acid, while only 12 (1.3%) of the fully susceptible isolates were nalidixic acid resistant. A similar association was observed between resistance to nalidixic acid and decreased susceptibility to ofloxacin or norfloxacin. A mutation of the gyrA gene could be demonstrated in all isolates for which the ciprofloxacin MICs were >/= 0.125 microg/ml and in 94% of the nalidixic acid-resistant isolates but in none of the nalidixic acid-susceptible isolates analyzed. Identification of nalidixic acid resistance by the disk diffusion method provided a sensitivity of 100% and a specificity of 87.3% as tools to screen for isolates for which the MICs of ciprofloxacin were >/= 0.125 microg/ml. We regard it as important that microbiology laboratories endeavor to recognize these less susceptible Salmonella strains, in order to reveal their clinical importance and to survey their epidemic spread. PMID- 10523555 TI - Use of BACTEC MGIT 960 for recovery of mycobacteria from clinical specimens: multicenter study. AB - The BACTEC MGIT 960 instrument is a fully automated system that exploits the fluorescence of an oxygen sensor to detect growth of mycobacteria in culture. Its performance was compared to those of the radiometric BACTEC 460 instrument and egg-based Lowenstein-Jensen medium. An identical volume of sample was inoculated in different media, and incubation was carried out for 6 weeks with the automatic systems and for 8 weeks on solid media. A total of 2,567 specimens obtained from 1,631 patients were cultured in parallel. Mycobacteria belonging to nine different taxa were isolated by at least one of the culture systems, with 75% of them being represented by Mycobacterium tuberculosis complex. The best yield was obtained with the BACTEC 460 system, with 201 isolates, in comparison with 190 isolates with the BACTEC MGIT 960 system and 168 isolates with Lowenstein-Jensen medium. A similar but not significant difference was obtained when the most represented organisms, the M. tuberculosis complex, Mycobacterium xenopi, and the Mycobacterium avium complex, were analyzed separately and when combinations of a solid medium with the BACTEC MGIT 960 system and with the BACTEC 460 system were considered. The shortest times to detection were obtained with the BACTEC MGIT 960 system (13.3 days); 1.5 days earlier than that with the BACTEC 460 system (14.8 days) and 12 days earlier than that with Lowenstein-Jensen medium (25.6 days). The BACTEC MGIT 960 system had a contamination rate of 10.0%, intermediate between those of the radiometric system (3.7%) and the egg-based medium (17.0%). We conclude, therefore, that the BACTEC MGIT 960 system is a fully automated, nonradiometric instrument that is suitable for the detection of growth of tuberculous and other mycobacterial species and that is characterized by detection times that are even shorter than that of the "gold standard," the BACTEC 460 system. The contamination rate was higher than that for the radiometric BACTEC 460 system and needs to be improved. PMID- 10523556 TI - Comparison of direct inoculation and Copan transport systems for isolation of Neisseria gonorrhoeae from endocervical specimens. AB - Two commercial swab transport systems, Copan Amies gel agar with and without charcoal (Copan Diagnostics, Corona, Calif.), were compared to direct inoculation onto modified Thayer-Martin medium for detection of Neisseria gonorrhoeae in 1,490 endocervical specimens obtained from women attending a sexually transmitted disease clinic. Copan swabs were held in the transport system for 24 h at room temperature prior to inoculation onto modified Thayer-Martin medium. All cultures were incubated at 35 degrees C in 5% CO(2), and bacteria were identified on the basis of Gram stain, oxidase, and biochemical reactions. Copan Amies gel agar transport system without charcoal detected 77 of 81 (95%) direct inoculation culture-positive specimens, and Copan Amies gel agar transport system with charcoal detected 53 of 56 (95%) directly inoculated culture-positive specimens. Copan Amies gel agar without charcoal inoculated after 6 h supported growth of 56 (98%) positive cultures out of only 55 directly inoculated culture-positive specimens. This study demonstrates that Copan swabs represent a reasonable alternative, providing convenience, low cost, and ease of use while still maintaining a satisfactory recovery rate of N. gonorrhoeae from clinical specimens, if specimens can be inoculated onto selective media within a relatively short time period not involving overnight shipment. PMID- 10523557 TI - Identification of Fusarium species involved in human infections by 28S rRNA gene sequencing. AB - Fusarium spp. have emerged as major opportunistic fungal agents. Since new antifungal agents exhibit variable activity against Fusarium isolates depending on the species, rapid identification at the species level is required. Conventional culture methods are difficult, fastidious, and sometimes inconclusive. In this work, we sequenced a 440-bp fragment encoding the 28S rRNA from 33 Fusarium isolates belonging to six Fusarium species associated with human infections. The data were then analyzed by the neighbor-joining method. By using distance matrix analysis and constructing the phylogram, we could easily distinguish the different species for all but one isolate. The method also allowed differentiation between the closely related genera Acremonium and Cylindrocarpon. In contrast to the case with conventional methods, the results could be obtained within 48 h from a 3-day culture and are independent of mycologist experience, making this method rapid and reliable for identification of Fusarium species isolated from patients. PMID- 10523558 TI - Glycopeptide-intermediate Staphylococcus aureus: evaluation of a novel screening method and results of a survey of selected U.S. hospitals. AB - Isolates of Staphylococcus aureus with decreased susceptibilities to glycopeptide antimicrobial agents, such as vancomycin and teicoplanin, have emerged in the United States and elsewhere. Commercially prepared brain heart infusion agar (BHIA) supplemented with 6 microg of vancomycin per ml was shown in a previous study to detect glycopeptide-intermediate S. aureus (GISA) with high sensitivity and specificity; however, this medium, when prepared in-house, occasionally showed growth of vancomycin-susceptible control organisms. This limitation could significantly impact laboratories that prepare media in-house, particularly if they wished to conduct large surveillance studies for GISA. Therefore, a pilot study to detect GISA was performed with vancomycin-containing Mueller-Hinton agar (MHA) prepared in-house in place of commercially prepared BHIA. MHA was selected for this study because this medium is widely available and well standardized. The results of the pilot study showed that supplementation of MHA with 5 microg of vancomycin per ml was both a sensitive and a specific method for screening for GISA isolates. This method was used to screen for GISA among 630 clinical isolates of methicillin-resistant S. aureus collected during 1997 from 33 U.S. hospitals. Although 14 S. aureus isolates grew on the screening agar, all were vancomycin susceptible (MICs were 100 parasites/microl) but drops to 59% when the level is <100 parasites/microl and to 39% when it is <50 parasites/microl. In addition, the OptiMAL test failed to identify four patients whose blood smears contained P. falciparum gametocytes only. We conclude that the sensitivity and specificity of the OptiMAL test are comparable to those of microscopy in detecting malaria infection at a parasitemia level of >100 parasites/microl; however, the test failed to identify more than half of the patients with a parasitemia level of <50 parasites/microl. Thus, the OptiMAL test should be used with great caution, and it should not replace conventional microscopy in the diagnosis of malaria infection. PMID- 10523568 TI - Isolation, identification, and molecular characterization of strains of Photorhabdus luminescens from infected humans in Australia. AB - We describe the isolation of Photorhabdus (Xenorhabdus) luminescens from four Australian patients: two with multiple skin lesions, one with bacteremia only, and one with disseminated infection. One of the patients had multiple skin lesions following the bite of a spider, while the lesions in the other patient were possibly associated with a spider bite. The source of infection for the remaining two patients is unknown. As a member of the family Enterobacteriaceae, P. luminescens is unusual in that it fails to reduce nitrate and ferments only glucose and mannose. It gives negative reactions for lysine decarboxylase, arginine dihydrolase, and ornithine decarboxylase (Moeller). The species is motile, utilizes citrate, hydrolyzes urea, and usually produces a unique type of annular hemolysis on sheep blood agar plates incubated at 25 degrees C. A weak bioluminescence is the defining characteristic. P. luminescens is an insect pathogen and is symbiotically associated with entomopathogenic nematodes. Its isolation from human clinical specimens has been reported previously from the United States. Restriction fragment length polymorphism-PCR analysis of the 16S rRNA gene demonstrated a high level of similarity among the Australian clinical strains and significant differences between the Australian clinical strains and the U.S. clinical strains. However, numerical analyses of the data suggest that the two groups of clinical strains are more similar to each other than they are to the symbiotic strains found in nematodes. This is the first report of the isolation of P. luminescens from infected humans in Australia and the second report of the isolation of this species from infected humans worldwide. PMID- 10523569 TI - Application of different genotyping methods for Pseudomonas aeruginosa in a setting of endemicity in an intensive care unit. AB - Colonization with Pseudomonas aeruginosa was studied by taking serial swab specimens from the oropharynges and anuses and tracheal and gastric aspirates from patients in an intensive care unit during a 10-month period in a setting of endemicity. Nineteen (10%) of the 192 patients included in the study were colonized on admission, while another 30 (16%) patients acquired P. aeruginosa while in the hospital. Typing of 353 isolates was performed by random amplified polymorphic DNA (RAPD) analysis, and 56 strains were selected for further typing by RAPD analysis, pulsed-field gel electrophoresis (PFGE), and amplified fragment length polymorphism (AFLP) analysis. By these methods, 42, 44, and 44 genotypes were found, respectively. Computer-aided cluster analysis indicated that similar groups of related isolates were obtained by each method. By taking admission periods into account, analysis of the typing results suggested cross-acquisition of P. aeruginosa for five patient pairs. The small number of transfers and the large number of genotypes found indicate that most P. aeruginosa strains were derived from the patients themselves. The numbers of observed typing patterns and band differences between related isolates were counted for each typing method. AFLP analysis with primers without a selective base proved to be the most discriminatory method, followed by PFGE, AFLP analysis (with one selective base), and RAPD analysis. On the basis of a comparison with established strain differentiation criteria for PFGE, the criteria for differentiation of P. aeruginosa by AFLP analysis are presented. PMID- 10523570 TI - Rapid identification of Burkholderia pseudomallei in blood cultures by a monoclonal antibody assay. AB - Burkholderia pseudomallei is the causative agent of melioidosis. In northeast Thailand, this gram-negative bacterium is a major cause of mortality from septicemia. The definitive diagnosis of this disease is made by bacterial culture. In this study, we produced a monoclonal antibody (MAb) specific to the 30-kDa protein of B. pseudomallei by in vivo and in vitro immunization of BALB/c mice with a crude culture filtrate antigen. The MAb could directly agglutinate with all 243 clinical isolates of B. pseudomallei but not with other gram negative bacteria, except for one strain of Burkholderia mallei. However, the MAb cross-reacted with the gram-positive Bacillus sp. and Streptococcus pyogenes. B. pseudomallei in brain heart infusion broth (BHIB) subcultured from a BacT/Alert automated blood culture system could be identified by simple agglutination with this MAb assay. The sensitivity and specificity of direct agglutination compared to the "gold standard," the culture method, were 94.12 and 98.25%, respectively. However, the MAb adsorbed to polystyrene beads or latex particles directly identified the bacterium in blood culture specimens and in BHIB subcultured from a BacT/Alert automated blood culture system. The sensitivity of the latex agglutination test was 100% for both blood culture and BHIB specimens. The specificity was 85.96 and 96.49% for the blood culture and BHIB specimens, respectively. The specificity could be increased if the nonspecific materials in the blood culture broths were eradicated by centrifugation at low speeds. Thus, a combination of blood culture and the agglutination method could be used for the rapid diagnosis of melioidosis in the routine bacteriological laboratory. This method could speed up detection of the bacterium in blood culture by at least 2 days, compared to the conventional bacterial culture method. In addition, the MAb is stable at room temperature for 2 weeks and at 4, -20, and -70 degrees C for at least 1 year. The latex reagent was stable for at least 6 months at 4 degrees C. PMID- 10523571 TI - Ability of the digene hybrid capture II test to identify Chlamydia trachomatis and Neisseria gonorrhoeae in cervical specimens. AB - The Digene Hybrid Capture II (HCII CT/GC) test is a combination test designed to detect Chlamydia trachomatis and Neisseria gonorrhoeae in a single specimen. It is a nucleic acid hybridization test which uses signal amplification to increase sensitivity. We compared its performance to that of culture on cervical specimens from 1,370 women. Direct fluorescent-antibody assay was used to resolve discrepant results for C. trachomatis. Samples were collected with a proprietary cervical brush or with endocervical swabs. The HCII CT/GC test proved to be sensitive and specific in detecting these organisms. Compared to N. gonorrhoeae culture, it had a sensitivity of 93% (87/94) and a specificity of 98.5% (1,244/1,263). Compared to C. trachomatis culture, the sensitivity was 97.7% (129/132) and specificity was 98.2% (1,216/1,238). Testing of some specimens with discrepant results by PCR suggested that the test would actually prove to be even more specific if it were compared to a nucleic acid amplification test (NAAT). The sensitivity of C. trachomatis culture was somewhat less, at 88.6% (117/132). The endocervical brush appeared to be better than Dacron swabs for collecting specimens. The HCII CT/GC test offers an attractive format that allows simultaneous detection of C. trachomatis and N. gonorrhoeae with a single specimen. An initial positive result is followed by repeat tests with probes to identify chlamydiae or gonococci. This test is more sensitive than C. trachomatis culture and is at least as sensitive as culture for gonococci. It deserves further evaluation and comparison with NAATs and may well offer an attractive alternative for diagnosis and screening of these infections. PMID- 10523572 TI - Detection of human herpesvirus 6 by reverse transcription-PCR. AB - The role of human herpesvirus 6 (HHV-6) in disease beyond primary infection remains unclear. We have developed and validated a new reverse transcription-PCR (RT-PCR) assay for HHV-6 that can determine the presence of HHV-6 in clinical specimens and differentiate between latent and replicating virus. Peripheral blood mononuclear cells from 109 children were evaluated for HHV-6 by RT-PCR, DNA PCR, and viral culture. Of these samples, 106 were suitable for analysis. A total of 20 samples were positive for HHV-6 by culture and DNA PCR, of which 19 were positive by RT-PCR (sensitivity, 95%). All 28 samples from children that were negative by viral culture, but positive by DNA PCR, were negative for viral transcripts by our RT-PCR assay. One positive RT-PCR result was observed in 56 samples that were negative by tissue culture and DNA PCR. This indicates a low rate of false-positive results (1.2%) and a specificity of 98.8%. This RT-PCR assay can reliably differentiate between latent and actively replicating HHV-6 and should allow insight into the pathogenesis of this ubiquitous virus. PMID- 10523573 TI - Phenotypic and genotypic heterogeneity among cultivable pathogen-related oral spirochetes and Treponema vincentii. AB - Recent findings challenge the assumption that pathogen-related oral spirochetes (PROS) are related to Treponema pallidum. Treponema vincentii, grown in OMIZ-Pat media, cross-reacted with monoclonal antibody H9-2 against T. pallidum, and cultivable PROS had 16S rRNA gene sequences similar to those of T. vincentii (C. B. Choi, C. Wyss, and U. B. Gobel. J. Clin. Microbiol. 34:1922-1925, 1996). Aims of the present study were to determine whether antigen phenotypes of oral treponemas were influenced by growth conditions and to evaluate the genetic relatedness of cultivable PROS to T. pallidum and T. vincentii. Results show that three T. pallidum monoclonal antibodies (H9-1, H9-2, and F5) cross-reacted with whole cells from four Treponema species grown in modified OMIZ-Pat medium, but not with treponemas grown in NOS medium. Only H9-2 reacted in immunoblots with reduced proteins from cultivable PROS and T. vincentii. Three of five PROS isolates were amplified by T. vincentii-specific PCR, and one was amplified by Treponema medium-specific PCR. None were amplified by T. pallidum-specific PCR. Three of five PROS isolates had 16S ribosomal DNA restriction fragment length polymorphism patterns identical to that of T. vincentii, and the patterns of two isolates resembled that of T. medium. Arbitrarily primed-PCR profiles from whole genomic DNA were distinct among five PROS isolates and two T. vincentii strains. Thus, PROS isolates represent a heterogeneous group of treponemas that share some 16S rRNA gene sequences with T. vincentii and T. medium, but not with T. pallidum. It is proposed that the PROS nomenclature be dropped. PMID- 10523574 TI - Genotypic and phenotypic characterization of "Streptococcus milleri" group isolates from a Veterans Administration hospital population. AB - Because identification of the species within the "Streptococcus milleri" group is difficult for the clinical laboratory as the species share overlapping phenotypic characteristics, we wished to confirm biochemical identification with identification by 16S rRNA gene sequence analysis. Ninety-four clinical isolates previously identified as the "Streptococcus milleri" group were reclassified as S. anginosus, S. constellatus, or S. intermedius with the API 20 Strep system (bioMerieux Vikek, Hazelton, Mo.) and the Fluo-card (Key Scientific, Round Rock, Tex.). In addition, we determined the Lancefield group, hemolysis, colony size, colony texture, repetitive extragenic palindromic PCR (rep-PCR) pattern, and cellular fatty acid (CFA) profile (MIDI, Newark, Del.). 16S rRNA gene sequence analysis with 40 selected representative strains showed three distinct groups, with S. constellatus and S. intermedius found to be more closely related to each other than to S. anginosus, and further distinguished a biochemically distinct group of urogenital isolates within the S. anginosus group of isolates. Except for strains unreactive with the Fluo-card (8%), all S. anginosus and S. intermedius strains identified by sequencing were similarly identified by biochemical testing. However, 23% of the selected S. constellatus isolates identified by sequencing (9% of all S. constellatus isolates) would have been identified as S. anginosus or S. intermedius by biochemical tests. Although most S. anginosus strains formed one unique cluster by CFA analysis and most S. constellatus strains showed similar rep-PCR patterns, neither method was sufficiently dependable for identification. Whereas Lancefield group or lactose fermentation did not correspond to sequence or biochemical type, S. constellatus was most likely to be beta-hemolytic and S. intermedius was most likely to have a dry colony type. The most frequent isolate in our population was S. constellatus, followed by S. anginosus. There was an association of S. anginosus with a gastrointestinal or urogenital source, and there was an association of S. constellatus and S. intermedius with both the respiratory tract and upper-body abscesses. PMID- 10523575 TI - Identification of Mycobacterium species by PCR-restriction fragment length polymorphism analyses using fluorescence capillary electrophoresis. AB - We developed a scheme for the rapid identification of Mycobacterium species based upon PCR amplification of polymorphic genetic regions with fluorescent primers followed by restriction and analysis by fluorescence capillary electrophoresis. Mycobacterium species were identified by restriction enzyme analysis of a 439-bp segment of the 65-kDa heat shock protein gene (labeled [both strands] at the 5' end with 4,7,2',7'-tetrachloro-6-carboxyfluorescein) using HaeIII and BstEII and of a 475-bp hypervariable region of the 16S rRNA gene (labeled [both strands] at the 5' end with 6-carboxyfluorescein) using HaeIII and CfoI. Samples were analyzed on an automated fluorescence capillary electrophoresis instrument, and labeled fragments were sized by comparison with an internal standard. DNA templates were prepared with pure cultures of type strains. In all, we analyzed 180 strains, representing 22 Mycobacterium species, and obtained distinctive restriction fragment length polymorphism (RFLP) patterns for 19 species. Three members of the Mycobacterium tuberculosis complex had a common RFLP pattern. A computerized algorithm which eliminates subjectivity from pattern interpretation and which is capable of identifying the species within a sample was developed. The convenience and short preparatory time of this assay make it comparable to conventional methodologies such as high-performance liquid chromatography and hybridization assays for identification of mycobacteria. PMID- 10523576 TI - Simple and rapid identification of the Mycobacterium tuberculosis complex by immunochromatographic assay using anti-MPB64 monoclonal antibodies. AB - A newly developed immunochromatographic assay (MPB64-ICA) for identification of the Mycobacterium tuberculosis complex was evaluated with 20 reference strains of mycobacterial species and 111 clinical isolates. MPB64-ICA displayed a very strong reaction band with organisms belonging to the M. tuberculosis complex but not with mycobacteria other than M. tuberculosis (MOTT bacilli), except for one of four M. marinum strains tested and one M. flavescens strain, both of which gave very weak signals. The effectiveness of MPB64-ICA in combination with two liquid culture systems (MB-REDOX and MGIT) was tested. A total of 108 of 362 sputum specimens processed were positive for acid-fast bacilli. Samples taken from the cultures on the same days when either of the two culture systems became positive for mycobacteria were assayed with MPB64-ICA. Of 108 cultures with mycobacteria, 51 showed a positive signal with the test, in which the presence of the M. tuberculosis complex was demonstrated later by the Accuprobe for M. tuberculosis complex. In addition, MPB64-ICA could correctly detect the M. tuberculosis complex in mixed cultures of the M. tuberculosis complex and MOTT bacilli. These results indicate that MPB64-ICA can be easily used for rapid identification of the M. tuberculosis complex in combination with culture systems based on liquid media without any technical complexity in clinical laboratories. PMID- 10523577 TI - Field evaluation of the Determine rapid human immunodeficiency virus diagnostic test in Honduras and the Dominican Republic. AB - Rapid detection of human immunodeficiency virus (HIV) infection can result in improved patient care and/or faster implementation of public health preventive measures. A new rapid test, Determine (Abbott, Abbott Park, Ill.), detects HIV type 1 (HIV-1) and HIV-2 antibodies within 15 min by using 50 microl of serum or plasma. No specialized equipment or ancillary supplies are required, and results are read visually. A positive result is noted by the appearance of a red line. An operational control (red line) indicates proper test performance. We evaluated the Determine rapid HIV detection test with a group of well-characterized serum samples (CD4 counts and viral loads were known) and serum samples from HIV positive individuals at field sites in Honduras and the Dominican Republic. In the field evaluations, the results obtained by the Determine assay were compared to those obtained by local in-country HIV screening procedures. We evaluated serum from 100 HIV-positive patients and 66 HIV-negative patients. All samples gave the expected results. In a companion study, 42 HIV-positive samples from a Miami, Fla., serum bank were tested by the Determine assay. The samples had been characterized in terms of CD4 counts and viral loads. Fifteen patients had CD4 counts <200 cells/mm(3), while 27 patients had CD4 counts >200 cells/mm(3). Viral loads ranged from 630 to 873,746 log(10) copies/ml. All samples from the Miami serum bank were positive by the Determine test. Combined results from the multicenter studies indicated that the correct results were obtained by the Determine assay for 100% (142 of 142) of the HIV-positive serum samples and 100% (66 of 66) of the HIV-negative serum samples. The Determine test was simple to perform and the results were easy to interpret. The Determine test provides a valuable new method for the rapid identification of HIV-positive individuals, especially in developing countries with limited laboratory infrastructures. PMID- 10523578 TI - Similar signature of the prion protein in natural sheep scrapie and bovine spongiform encephalopathy-linked diseases. AB - It has been suggested that specific molecular features could characterize the protease-resistant prion protein (PrP res) detected in animal species as well as in humans infected by the infectious agent strain that causes bovine spongiform encephalopathy (BSE). Studies of glycoform patterns in such diseases in French cattle and cheetahs, as well as in mice infected by isolates from both species, revealed this characteristic molecular signature. Similar studies of 42 French isolates of natural scrapie, from 21 different flocks in different regions of France, however, showed levels of the three glycoforms comparable to those found in BSE-linked diseases. Moreover, the apparent molecular size of the unglycosylated form was also indistinguishable among all different sheep isolates, as well as isolates from BSE in cattle. Overall results suggest that scrapie cases with features similar to those of BSE could be found more frequently in sheep than previously described. PMID- 10523579 TI - Comparison of selective broth medium plus neomycin-nalidixic acid agar and selective broth medium plus Columbia colistin-nalidixic acid agar for detection of group B streptococcal colonization in women. AB - The combination of neomycin-nalidixic acid (NNA) agar and a selective broth medium (SBM) has recently been shown to improve the sensitivity of screening cultures for group B streptococcal (GBS) carriage in women. Because of the relatively high cost of NNA agar, a study was initiated to determine whether Columbia colistin-nalidixic acid (CNA) agar would be an equally sensitive, more economical alternative. A total of 580 cervical-vaginal and/or rectal specimens submitted for detection of GBS were included in the study. Each was plated onto NNA and CNA agar and then inoculated into SBM. GBS were recovered from 95 of 580 (16.4%) specimens, including 61 isolates from CNA, 74 from NNA, 73 from the CNA SMB combination, and 86 from the NNA-SMB tandem. Of those, 22 isolates were recovered on NNA but not CNA, 9 were cultured on CNA but not NNA, 52 were isolated on both media, and 12 were recovered from subcultures of SBM only. The overall sensitivity of CNA alone (64. 2%) was statistically significantly less than that of NNA agar (77. 9%), as was the sensitivity of combination of CNA plus SBM (76.8%) compared to that of NNA plus SBM (90.5%). Based on these findings, CNA should not be considered an acceptable alternative to NNA for the detection of GBS colonization in women despite potential cost savings. PMID- 10523580 TI - Predicting susceptibility of Streptococcus pneumoniae to ceftriaxone and cefotaxime by cefuroxime and ceftizoxime disk diffusion testing. AB - In this study, disk diffusion testing with ceftizoxime and cefuroxime was evaluated for use in predicting the susceptibility of Streptococcus pneumoniae to ceftriaxone and cefotaxime. Of the 194 isolates included in this study, 138 were susceptible, 34 were intermediate, and 22 were resistant to cefotaxime by MIC testing; 138 isolates were susceptible, 35 were intermediate, and 21 were resistant to ceftriaxone by MIC testing. A zone of inhibition around the cefuroxime disk of >/=32 mm correctly categorized 101 of 138 isolates as susceptible to cefotaxime and ceftriaxone. A zone of inhibition around the ceftizoxime disk of >/=26 mm correctly categorized 111 of 138 isolates as susceptible to cefotaxime and 114 of 138 as susceptible to ceftriaxone. We conclude that disk diffusion can separate S. pneumoniae isolates susceptible to ceftriaxone and cefotaxime from those that are not susceptible. Isolates not falling into the susceptible category by disk diffusion require additional testing to determine the MIC. PMID- 10523581 TI - Comparison of MB/BacT and BACTEC 460 TB systems for recovery of mycobacteria in a routine diagnostic laboratory. AB - MB/BacT, BACTEC 460 TB, and Lowenstein-Jensen (LJ) medium were evaluated in parallel for recovery of mycobacteria from 3,700 continuous clinical specimens in a routine laboratory. Mycobacteria were identified from 123 (3.3%) specimens. The recovery rates for all mycobacteria by the different systems were 91.0, 73.0, and 53.6% for BACTEC 460 TB, MB/BacT, and LJ medium, respectively. The recovery rates for Mycobacterium tuberculosis complex were 97.1, 80. 2, and 67.6%, respectively. The lack of sensitivity of the MB/BacT system was more pronounced with smear negative specimens and resulted in a failure to detect three patients with infectious tuberculosis. PMID- 10523582 TI - Molecular markers reveal exclusively clonal reproduction in Trichophyton rubrum. AB - Genotypic variability among 96 Trichophyton rubrum strains which displayed different colony morphologies and were collected from four continents was investigated. Twelve markers representing 57 loci were analyzed by PCR fingerprinting, amplified fragment length polymorphism, and random amplified monomorphic DNA markers. Interestingly, none of the methods used revealed any DNA polymorphism, indicating a strictly clonal mode of reproduction and a strong adaptation to human skin. PMID- 10523583 TI - Effects of OspA vaccination on Lyme disease serologic testing. AB - This study presents the effects of OspA vaccination on two-step testing for Borrelia burgdorferi antibodies. Although vaccinees developed enzyme-linked immunosorbent assay reactivity, immunoblots did not fulfill Centers for Disease Control and Prevention criteria for positivity. Furthermore, OspA reactivity did not interfere with interpretation of immunoblots with sera from patients who developed early Lyme disease despite vaccination. PMID- 10523584 TI - Isolation of Rickettsia prowazekii from blood by shell vial cell culture. AB - A blood sample from a patient who returned from Algeria with a fever inoculated on human embryonic lung fibroblasts by the shell vial cell culture technique led to the recovery of Rickettsia prowazekii. The last clinical strain was isolated 30 years ago. Shell vial cell culture is a versatile method that could replace the classic animal and/or embryonated egg inoculation. PMID- 10523585 TI - Genotypic survey of recent beta-lactam-resistant pneumococcal nasopharyngeal isolates from asymptomatic children in Chile. AB - To assess pneumococcal strain variability among young asymptomatic carriers in Chile, we used serotyping, antibiotic susceptibility testing, and genotyping to analyze 68 multidrug-resistant pneumococcal isolates recovered from 54 asymptomatic children 6 to 48 months of age. The isolates represented capsular serotypes 19F (43 isolates), 14 (14 isolates), 23F (7 isolates), 6B (3 isolates), and 6A (1 isolate). Genotypic analysis, which included pulsed-field gel electrophoresis (PFGE) of chromosomal digests, penicillin binding protein (PBP) gene fingerprinting, and dhf gene fingerprinting, revealed that the isolates represented six different genetic lineages. Clear circumstantial evidence of capsular switching was seen within each of four of the genetically related sets. The majority of the isolates, consisting of the 43 19F isolates and 2 type 6B isolates, appeared to represent a genetically highly related set distinct from previously characterized pneumococcal strains. Each of three other genetically defined lineages was closely related to one of the previously characterized clones Spain(6B)-2, France(9V)-3, or Spain(23F)-1. A fifth lineage was comprised of four type 23F isolates that, by the techniques used for this study, were genetically indistinguishable from three recent type 19F sterile-site isolates from the United States. Finally, a sixth lineage was represented by a single type 23F isolate which had a unique PFGE type and unique PBP and dhf gene fingerprints. PMID- 10523586 TI - Importance of selective media for recovery of yeasts from clinical specimens. AB - We compared the recovery of yeasts from clinical specimens cultured on routine bacteriological media to the recovery of yeast from specimens cultured on a selective fungal medium (Sabouraud agar). The use of Sabouraud agar was especially important in cases of mixed cultures, since in such cases yeast was recovered on bacteriological media from only 50% of 44 yeast-positive pus specimens and from 22. 5% of 22 yeast-positive throat specimens. The use of a selective fungal medium is therefore necessary to ensure the detection of yeast in specimens containing a mixture of bacteria and yeasts. As a result, clinicians must request yeast isolation when clinically indicated, and the microbiological laboratory must add a selective fungal medium when clinically significant yeasts are likely to be encountered. It is also important that selective fungal media be used in clinical studies of yeast infections. PMID- 10523587 TI - Case of peritonitis caused by Ewingella americana in a patient undergoing continuous ambulatory peritoneal dialysis. AB - Reports of serious infections caused by Ewingella americana have been rare. A case of E. americana peritonitis in a patient receiving continuous ambulatory peritoneal dialysis is described. This is the first report of E. americana causing such an infection. PMID- 10523588 TI - Candida isolates from neonates: frequency of misidentification and reduced fluconazole susceptibility. AB - Systemic candidiasis affects 1.6 to 4.5% of very low birth weight (8 microg/ml. PMID- 10523589 TI - Evaluation of a new commercially available immunoglobulin M capture enzyme-linked immunosorbent assay for diagnosis of Japanese encephalitis infections. AB - A new commercial enzyme-linked immunosorbent assay (ELISA) for the diagnosis of Japanese encephalitis virus infections showed a sensitivity of 88% with sera and 81% with cerebrospinal fluid and a specificity of 97% with sera from patients with primary and secondary dengue virus infections. Specificity was 100% when samples from nonflavivirus infections were tested. PMID- 10523590 TI - Cloning and characterization of a nonhemolytic phospholipase C gene from Burkholderia pseudomallei. AB - We cloned and characterized a phosphatidylcholine-hydrolyzing phospholipase C (PC PLC) gene from Burkholderia pseudomallei. DNA sequence analysis of the gene indicated an open reading frame coding for 700 amino acids with a 34-amino-acid signal peptide. When cleaved, this yields a secreted 73-kDa mature protein. The deduced amino acid sequence exhibited 48% similarity to that of a nonhemolytic PLC from Pseudomonas aeruginosa. The expressed PC-PLC was heat stable, nonhemolytic for sheep erythrocytes, and active between pH 2 and 8. Western blot analysis with sera from melioidosis patients indicated that they produced immunoglobulin M antibodies against this PC-PLC protein. PMID- 10523591 TI - Novel method for rapid determination of clarithromycin sensitivity in Helicobacter pylori. AB - A novel PCR-hybridization assay, performed in single closed capillaries, was developed to detect clarithromycin resistance-associated gene mutations in Helicobacter pylori. Mutations were detected by thermal analysis in 33 of 34 (97%) resistant isolates but not in 66 isolates determined to be sensitive by conventional antibiotic assays. The method was rapid and reproducible, and it reduced PCR product contamination risk. PMID- 10523592 TI - Viability of Trichomonas vaginalis in transport medium. AB - The ability of Amies gel agar transport medium to maintain the viability of Trichomonas vaginalis was determined by comparing transported vaginal specimens to specimens immediately inoculated into culture medium. The prevalence of trichomonosis in the study population was 26% (68 of 260 women). The immediate inoculation method detected infections in 64 of 68 infected women (sensitivity of 94.1%). The transport method detected 62 of 68 infections (sensitivity of 91.2%). There was no significant difference between the two methods. PMID- 10523593 TI - Fatal case of Trichoderma harzianum infection in a renal transplant recipient. AB - We describe the second known case of human infection by Trichoderma harzianum. A disseminated fungal infection was detected in the postmortem examination of a renal transplant recipient and confirmed in culture. The only other reported infection by this fungus caused peritonitis in a diabetic patient. The in vitro antifungal susceptibilities of the clinical strain and three other strains of Trichoderma species to six antifungal drugs are provided. This case illustrates the widening spectrum of opportunistic Trichoderma spp. in immunocompromised patients and emphasizes the problems in diagnosing invasive fungal diseases. PMID- 10523594 TI - Evaluation of the revised MicroScan dried overnight gram-positive identification panel to identify Enterococcus species. AB - The revised MicroScan Dried Overnight Gram-Positive Identification panel was evaluated for its efficacy at identifying Enterococcus species in comparison with conventional biochemical tests. Supplemental testing of ampicillin-susceptible Enterococcus faecium for motility and the ability to acidify methyl-alpha-D glucopyranoside helped recognize E. gallinarum and increased the accuracy of the panel for identifying Enterococcus species to 98.5%. PMID- 10523595 TI - A serotype VIII strain among colonizing group B streptococcal isolates in Boston, Massachusetts. AB - Maternal colonization with group B Streptococcus (GBS) is a risk factor for neonatal GBS disease. Whereas serotypes Ia, Ib, II, III, and V are prevalent in the United States, types VI and VIII predominate in Japan. Recently, a serotype VIII strain was detected among 114 clinical GBS isolates from a Boston, Mass., hospital. PMID- 10523596 TI - Recovery of mycobacteria from patients with cystic fibrosis. AB - Despite decontamination, overgrowth by pseudomonads renders cultural isolation of mycobacteria from respiratory specimens of patients with cystic fibrosis (CF) difficult or impossible. We performed a prospective study by comparing levels of reduction of overgrowth and recovery of mycobacteria using either pretreatment with N-acetyl-L-cysteine (NALC)-NaOH alone or pretreatment with NALC-NaOH and then with oxalic acid. From 406 specimens of 148 CF patients, 11 specimens were positive for mycobacteria, 5 of which grew mycobacteria after decontamination by either procedure. Three specimens grew mycobacteria only after decontamination with NALC-NaOH, whereas three specimens grew mycobacteria only after treatment with NALC-NaOH followed by oxalic acid but were overgrown after decontamination with NALC-NaOH. Thus, inactivation of mycobacteria by the more aggressive oxalic acid treatment offsets its beneficial effect of reducing the proportion of cultures overgrown with microorganisms other than mycobacteria. PMID- 10523597 TI - Transport and storage of fresh and frozen gastric biopsy specimens for optimal recovery of Helicobacter pylori. AB - The viability of Helicobacter pylori in vitro and in gastric biopsy specimens was determined. Recovery rates were 94, 87, and 77% from biopsy specimens in Portagerm pylori in cooled containers after 1, 2, and 3 days of transport, respectively (n = 307), and 97% if stored and shipped in glycerol broth at -70 degrees C (n = 232). PMID- 10523598 TI - PCR assay for detecting porcine cytomegalovirus. AB - This is the first published report of a PCR assay for detecting porcine cytomegalovirus (PCMV), the causative agent of inclusion body rhinitis in pigs. The DNA to be tested was extracted directly from lungs and nasal scrapings of pigs with various clinical syndromes. Fifty-nine percent (74 of 126) of tested pigs with various clinical syndromes were found to be PCR positive for PCMV. It is hoped that veterinary diagnostic laboratories will benefit by using this PCR assay for routine testing and surveillance of PCMV in pigs. PMID- 10523599 TI - Characterization of extended-spectrum beta-lactamase-producing Salmonella typhimurium by phenotypic and genotypic typing methods. AB - During 1994, 10 isolates of extended-spectrum beta-lactamase-producing Salmonella typhimurium were recovered from children transferred to our hospital from two different centers. Two additional isolates were recovered from two nurses from one of these centers. The aim of this study was to determine if there is any relationship between these isolates. The characterization was done by phenotypic and genotypic methods: biotyping, phage typing, antibiotic susceptibility pattern determination, plasmid analysis, ribotyping (by the four endonucleases EcoRI, SmaI, BglII, and PvuII), pulsed-field gel electrophoresis (PFGE) of genome macrorestriction patterns with XbaI, and randomly amplified polymorphic DNA (RAPD) pattern determination (with the three primers 217 d2, B1, and A3). The same biotype, the same serotype, and an identical antibiotype were found. All isolates were resistant to oxyimino-beta-lactams, gentamicin, tobramycin, and sulfamethoxazole-trimethoprim. All isolates showed an indistinguishable pattern by ribotyping and very similar patterns by PFGE and RAPD. The overall results indicated the spread of a closely related strain of S. typhimurium in children and nurses. PMID- 10523600 TI - Spread of a Salmonella typhimurium clone resistant to expanded-spectrum cephalosporins in three European countries. AB - Twelve Salmonella typhimurium strains resistant to broad-spectrum cephalosporins were isolated from cases of gastroenteritis during 1996 to 1998 in Russia, Hungary, and Greece. Resistance was due to the production of CTX-M-type extended spectrum beta-lactamases encoded by similar 12-kb plasmids. By pulsed-field gel electrophoresis, all strains shared the same chromosomal type. These data suggest that an S. typhimurium clone resistant to broad-spectrum cephalosporins is present in at least three European countries. PMID- 10523601 TI - Septicemia in neutropenic patients infected with Clostridium tertium resistant to cefepime and other expanded-spectrum cephalosporins. AB - Clostridium tertium was isolated from two immunocompromised patients with septicemia, fever, and gastrointestinal symptoms. The strains were resistant to ceftazidime, cefepime, and clindamycin; intermediately resistant to penicillin; and susceptible to metronidazole, quinolones, and vancomycin. PMID- 10523602 TI - Pre-T cell receptor (TCR) and TCR-controlled checkpoints in T cell differentiation are set by Ikaros. AB - T cell differentiation relies on pre-T cell receptor (TCR) and TCR signaling events that take place at successive steps of the pathway. Here, we show that two of these T cell differentiation checkpoints are regulated by Ikaros. In the absence of Ikaros, double negative thymocytes can differentiate to the double positive stage without expression of a pre-TCR complex. Subsequent events in T cell development mediated by TCR involving transition from the double positive to the single positive stage are also regulated by Ikaros. Nonetheless, in Ikaros deficient thymocytes, the requirement of pre-TCR expression for expansion of immature thymocytes as they progress to the double positive stage is still maintained, and the T cell malignancies that invariably arise in the thymus of Ikaros-deficient mice are dependent on either pre-TCR or TCR signaling. We conclude that Ikaros regulates T cell differentiation, selection, and homeostasis by providing signaling thresholds for pre-TCR and TCR. PMID- 10523603 TI - Immunoglobulin A1 protease, an exoenzyme of pathogenic Neisseriae, is a potent inducer of proinflammatory cytokines. AB - A characteristic of human pathogenic Neisseriae is the production and secretion of an immunoglobulin (Ig)A1-specific serine protease (IgA1 protease) that cleaves preferentially human IgA1 and other target proteins. Here we show a novel function for native IgA1 protease, i.e., the induction of proinflammatory cytokines such as tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, IL 6, and IL-8 from peripheral blood mononuclear cells. The capacity of IgA1 protease to elicit such cytokine responses in monocytes was enhanced in the presence of T lymphocytes. IgA1 protease did not induce the regulatory cytokine IL-10, which was, however, found in response to lipopolysaccharide and phytohemagglutinin. The immunomodulatory effects caused by IgA1 protease require a native form of the enzyme, and denaturation abolished cytokine induction. However, the proteolytic activity is not required for the cytokine induction by IgA1 protease. Our results indicate that IgA1 protease exhibits important immunostimulatory properties and may contribute substantially to the pathogenesis of neisserial infections by inducing large amounts of TNF-alpha and other proinflammatory cytokines. In particular, IgA1 protease may represent a key virulence determinant of bacterial meningitis. PMID- 10523604 TI - PIM1 reconstitutes thymus cellularity in interleukin 7- and common gamma chain mutant mice and permits thymocyte maturation in Rag- but not CD3gamma-deficient mice. AB - The majority of lymphomas induced in Rag-deficient mice by Moloney murine leukemia virus (MoMuLV) infection express the CD4 and/or CD8 markers, indicating that proviral insertions cause activation of genes affecting the development from CD4(-)8(-) pro-T cells into CD4(+)8(+) pre-T cells. Similar to MoMuLV wild-type tumors, 50% of CD4(+)8(+) Rag-deficient tumors carry a provirus near the Pim1 protooncogene. To study the function of PIM proteins in T cell development in a more controlled setting, a Pim1 transgene was crossed into mice deficient in either cytokine or T cell receptor (TCR) signal transduction pathways. Pim1 reconstitutes thymic cellularity in interleukin (IL)-7- and common gamma chain deficient mice. In Pim1-transgenic Rag-deficient mice but notably not in CD3gamma deficient mice, we observed slow expansion of the CD4(+)8(+) thymic compartment to almost normal size. Based on these results, we propose that PIM1 functions as an efficient effector of the IL-7 pathway, thereby enabling Rag-deficient pro-T cells to bypass the pre-TCR-controlled checkpoint in T cell development. PMID- 10523605 TI - Binding and antigen presentation of ceramide-containing glycolipids by soluble mouse and human CD1d molecules. AB - We have purified soluble mouse and human CD1d molecules to assess the structural requirements for lipid antigen presentation by CD1. Plate-bound CD1d molecules from either species can present the glycolipid alpha-galactosyl ceramide (alpha GalCer) to mouse natural killer T cells, formally demonstrating both the in vitro formation of antigenic complexes, and the presentation of alpha-GalCer by these two CD1d molecules. Using surface plasmon resonance, we show that at neutral pH, mouse CD1 and human CD1d bind to immobilized alpha-GalCer, unlike human CD1b, which requires acidic pH for lipid antigen binding. The CD1d molecules can also bind both to the nonantigenic beta-GalCer and to phosphatidylethanolamine, indicating that diverse lipids can bind to CD1d. These studies provide the first quantitative analysis of monomeric lipid antigen-CD1 interactions, and they demonstrate that the orientation of the galactose, or even the nature of the polar head group, are likely to be more important for T cell receptor contact than CD1d binding. PMID- 10523606 TI - The activated type 1-polarized CD8(+) T cell population isolated from an effector site contains cells with flexible cytokine profiles. AB - The capacity of activated T cells to alter their cytokine expression profiles after migration into an effector site has not previously been defined. We addressed this issue by paired daughter analysis of a type 1-polarized CD8(+) effector T cell population freshly isolated from lung parenchyma of influenza virus-infected mice. Single T cells were activated to divide in vitro; individual daughter cells were then micromanipulated into secondary cultures with and without added IL-4 to assess their potential to express type 2 cytokine genes. The resultant subclones were analyzed for type 1 and 2 cytokine mRNAs at day 6-7. When the most activated (CD44(high)CD11a(high)) CD8(+) subpopulation from infected lung was compared with naive or resting (CD44(low)CD11a(low)) CD8(+) cells from infected lung and from normal lymph nodes (LNs), both clonogenicity and plasticity of the cytokine response were highest in the LN population and lowest in the activated lung population, correlating inversely with effector function. Multipotential cells were nevertheless detected among clonogenic CD44(high)CD11a(high) lung cells at 30-50% of the frequency in normal LNs. The data indicate that activated CD8(+) T cells can retain the ability to proliferate and express new cytokine genes in response to local stimuli after recruitment to an effector site. PMID- 10523607 TI - Allelic exclusion of the T cell receptor beta locus requires the SH2 domain containing leukocyte protein (SLP)-76 adaptor protein. AB - Signaling via the pre-T cell receptor (TCR) is required for the proliferative expansion and maturation of CD4(-)CD8(-) double-negative (DN) thymocytes into CD4(+)CD8(+) double-positive (DP) cells and for TCR-beta allelic exclusion. The adaptor protein SH2 domain-containing leukocyte protein (SLP)-76 has been shown to play a crucial role in thymic development, because thymocytes of SLP-76(-/-) mice are arrested at the CD25(+)CD44(-) DN stage. Here we show that SLP-76(-/-) DN thymocytes express the pre-TCR on their surfaces and that introduction of a TCR-alpha/beta transgene into the SLP-76(-/-) background fails to cause expansion of DN thymocytes or developmental progression to the DP stage. Moreover, analysis of TCR-beta rearrangement in SLP-76(-/-) TCR-transgenic mice or in single CD25(+)CD44(-) DN cells from SLP-76(-/-) mice indicates an essential role of SLP 76 in TCR-beta allelic exclusion. PMID- 10523608 TI - Herpes simplex virus type 1 infection of activated cytotoxic T cells: Induction of fratricide as a mechanism of viral immune evasion. AB - Herpes simplex virus type 1 (HSV1), a large DNA-containing virus, is endemic in all human populations investigated. After infection of mucocutaneous surfaces, HSV1 establishes a latent infection in nerve cells. Recently, it was demonstrated that HSV1 can also infect activated T lymphocytes. However, the consequences of T cell infection for viral pathogenesis and immunity are unknown. We have observed that in contrast to the situation in human fibroblasts, in human T cell lines antigen presentation by major histocompatibility complex class I molecules is not blocked after HSV1 infection. Moreover, HSV1 infection of T cells results in rapid elimination of antiviral T cells by fratricide. To dissect the underlying molecular events, we used a transgenic mouse model of HSV1 infection to demonstrate that CD95 (Apo-1, Fas)-triggered apoptosis is essential for HSV1 induced fratricide, whereas tumor necrosis factor (TNF) also contributes to this phenomenon but to a lesser extent. By contrast, neither TRAIL (TNF-related apoptosis-inducing ligand) nor perforin were involved. Finally, we defined two mechanisms associated with HSV1-associated fratricide of antiviral T cells: (a) T cell receptor-mediated upregulation of CD95 ligand and (b) a viral "competence-to die" signal that renders activated T lymphocytes susceptible to CD95 signaling. We propose that induction of fratricide is an important immune evasion mechanism of HSV1, helping the virus to persist in the host organism throughout its lifetime. PMID- 10523609 TI - Compromised OX40 function in CD28-deficient mice is linked with failure to develop CXC chemokine receptor 5-positive CD4 cells and germinal centers. AB - Mice rendered deficient in CD28 signaling by the soluble competitor, cytotoxic T lymphocyte-associated molecule 4-immunoglobulin G1 fusion protein (CTLA4-Ig), fail to upregulate OX40 expression in vivo or form germinal centers after immunization. This is associated with impaired interleukin 4 production and a lack of CXC chemokine receptor (CXCR)5 on CD4 T cells, a chemokine receptor linked with migration into B follicles. Germinal center formation is restored in CTLA4-Ig transgenic mice by coinjection of an agonistic monoclonal antibody to CD28, but this is substantially inhibited if OX40 interactions are interrupted by simultaneous injection of an OX40-Ig fusion protein. These data suggest that CD28 dependent OX40 ligation of CD4 T cells at the time of priming is linked with upregulation of CXCR5 expression, and migration of T cells into B cell areas to support germinal center formation. PMID- 10523610 TI - In vivo-activated CD4 T cells upregulate CXC chemokine receptor 5 and reprogram their response to lymphoid chemokines. AB - Migration of antigen-activated CD4 T cells to B cell areas of lymphoid tissues is important for mounting T cell-dependent antibody responses. Here we show that CXC chemokine receptor (CXCR)5, the receptor for B lymphocyte chemoattractant (BLC), is upregulated on antigen-specific CD4 T cells in vivo when animals are immunized under conditions that promote T cell migration to follicles. In situ hybridization of secondary follicles for BLC showed high expression in mantle zones and low expression in germinal centers. When tested directly ex vivo, CXCR5(hi) T cells exhibited a vigorous chemotactic response to BLC. At the same time, the CXCR5(hi) cells showed reduced responsiveness to the T zone chemokines, Epstein-Barr virus-induced molecule 1 (EBI-1) ligand chemokine (ELC) and secondary lymphoid tissue chemokine (SLC). After adoptive transfer, CXCR5(hi) CD4 T cells did not migrate to follicles, indicating that additional changes may occur after immunization that help direct T cells to follicles. To further explore whether T cells could acquire an intrinsic ability to migrate to follicles, CD4(-)CD8(-) double negative (DN) T cells from MRL-lpr mice were studied. These T cells normally accumulate within follicles of MRL-lpr mice. Upon transfer to wild-type recipients, DN T cells migrated to follicle proximal regions in all secondary lymphoid tissues. Taken together, our findings indicate that reprogramming of responsiveness to constitutively expressed lymphoid tissue chemokines plays an important role in T cell migration to the B cell compartment of lymphoid tissues. PMID- 10523612 TI - Txk, a nonreceptor tyrosine kinase of the Tec family, is expressed in T helper type 1 cells and regulates interferon gamma production in human T lymphocytes. AB - Differentiation of human T cells into T helper (Th)1 and Th2 cells is vital for the development of cell-mediated and humoral immunity, respectively. However, the precise mechanism responsible for the Th1 cell differentiation is not fully clarified. We have studied the expression and function of Txk, a member of the Tec family of nonreceptor tyrosine kinases. We found that Txk expression is restricted to Th1/Th0 cells with IFN-gamma producing potential. Txk transfection of Jurkat T cells resulted in a several-fold increase of IFN-gamma mRNA expression and protein production; interleukin (IL)-2 and IL-4 production were unaffected. Antisense oligodeoxynucleotide of Txk specifically inhibited IFN gamma production of normal peripheral blood lymphocytes, antigen-specific Th1 clones, and Th0 clones; IL-2 and IL-4 production by the T cells was unaffected. Txk cotransfection led to the enhanced luciferase activity of plasmid (p)IFN gamma promoter/enhancer (pIFN-gamma[-538])-luciferase-transfected Jurkat cells upon mitogen activation. Txk transfection did not affect IL-2 and IL-4 promoter activities. Thus, Txk specifically upregulates IFN-gamma gene transcription. In fact, Txk translocated from cytoplasm into nuclei upon activation and transfection with a mutant Txk expression plasmid that lacked a nuclear localization signal sequence did not enhance IFN-gamma production by the cells, indicating that nuclear localization of Txk is obligatory for the enhanced IFN gamma production. In addition, IL-12 treatment of peripheral blood CD4(+) T cells enhanced the Txk expression, whereas IL-4 treatment completely inhibited it. These results indicate that Txk expression is intimately associated with development of Th1/Th0 cells and is significantly involved in the IFN-gamma production by the cells through Th1 cell-specific positive transcriptional regulation of the IFN-gamma gene. PMID- 10523611 TI - A20 inhibits cytokine-induced apoptosis and nuclear factor kappaB-dependent gene activation in islets. AB - Insulin-dependent diabetes mellitus (IDDM) is an autoimmune disease resulting from apoptotic destruction of beta cells in the islets of Langerhans. Low expression of antioxidants and a predilection to produce nitric oxide (NO) have been shown to underscore beta cell apoptosis. With this perspective in mind, we questioned whether beta cells could mount an induced protective response to inflammation. Here we show that human and rat islets can be induced to rapidly express the antiapoptotic gene A20 after interleukin (IL)-1beta activation. Overexpression of A20 by means of adenovirus-mediated gene transfer protects islets from IL-1beta and interferon gamma-induced apoptosis. The cytoprotective effect of A20 against apoptosis correlates with and is dependent on the abrogation of cytokine-induced NO production. The inhibitory effect of A20 on cytokine-stimulated NO production is due to transcriptional blockade of inducible NO synthase (iNOS) induction; A20 inhibits the activation of the transcription factor nuclear factor kappaB at a level upstream of IkappaBalpha degradation. These data demonstrate a dual antiapoptotic and antiinflammatory function for A20 in beta cells. This qualifies A20 as part of the physiological cytoprotective response of islets. We propose that A20 may have therapeutic potential as a gene therapy candidate to achieve successful islet transplantation and the cure of IDDM. PMID- 10523613 TI - Human dendritic cells mediate cellular apoptosis via tumor necrosis factor related apoptosis-inducing ligand (TRAIL). AB - TRAIL (TNF-related apoptosis-inducing ligand) is a member of the TNF family that induces apoptosis in a variety of cancer cells. In this study, we demonstrate that human CD11c(+) blood dendritic cells (DCs) express TRAIL after stimulation with either interferon (IFN)-gamma or -alpha and acquire the ability to kill TRAIL-sensitive tumor cell targets but not TRAIL-resistant tumor cells or normal cell types. The DC-mediated apoptosis was TRAIL specific, as soluble TRAIL receptor blocked target cell death. Moreover, IFN-stimulated interleukin (IL)-3 receptor (R)alpha(+) blood precursor (pre-)DCs displayed minimal cytotoxicity toward the same target cells, demonstrating a clear functional difference between the CD11c(+) DC and IL-3Ralpha(+) pre-DC subsets. These results indicate that TRAIL may serve as an innate effector molecule on CD11c(+) DCs for the elimination of spontaneously arising tumor cells and suggest a means by which TRAIL-expressing DCs may regulate or eliminate T cells responding to antigen presented by the DCs. PMID- 10523614 TI - Protective T cell-independent antiviral antibody responses are dependent on complement. AB - Complement is part of the innate immune system and one of the first lines of host defense against infections. Its importance was evaluated in this study in virus infections in mice deficient either in soluble complement factors (C3(-/-), C4(-/ )) or in the complement signaling complex (complement receptor [CR]2(-/-), CD19( /-)). The induction of the initial T cell-independent neutralizing immunoglobulin (Ig)M antibody response to vesicular stomatitis virus (VSV), poliomyelitis virus, and recombinant vaccinia virus depended on efficient antigen trapping by CR3 and 4-expressing macrophages of the splenic marginal zone. Neutralizing IgM and IgG antibody responses were largely independent of CR2-mediated stimulation of B cells when mice were infected with live virus. In contrast, immunizations with nonreplicating antigens revealed an important role of B cell stimulation via CR2 in the switch to IgG. The complement cascade was activated after infection with VSV via the classical pathway, and active complement cleavage products augmented the effector function of neutralizing IgM and IgG antibodies to VSV by a factor of 10-100. Absence of the early neutralizing antibody responses, together with the reduced efficiency of neutralizing IgM in C3(-/-) mice, led to a drastically enhanced susceptibility to disease after infection with VSV. PMID- 10523616 TI - The reduced expression of 6Ckine in the plt mouse results from the deletion of one of two 6Ckine genes. AB - 6Ckine is an unusual chemokine capable of attracting naive T lymphocytes in vitro. It has been recently reported that lack of 6Ckine expression in lymphoid organs is a prominent characteristic of mice homozygous for the paucity of lymph node T cell (plt) mutation. These mice show reduced numbers of T cells in lymph nodes, Peyer's patches, and the white pulp of the spleen. The genetic reason for the lack of 6Ckine expression in the plt mouse, however, has remained unknown. Here we demonstrate that mouse 6Ckine is encoded by two genes, one of which is expressed in lymphoid organs and is deleted in plt mice. A second 6Ckine gene is intact and expressed in the plt mouse. PMID- 10523615 TI - CD47 ligation selectively downregulates human interleukin 12 production. AB - Interleukin (IL)-12 plays a key role not only in protective innate and adaptive T helper cell type 1 (Th1) responses but also in chronic inflammatory diseases. We report here that engagement of CD47 by either monoclonal antibody, its natural ligand thrombospondin (TSP), or 4N1K (a peptide of the COOH-terminal domain of TSP selectively binding CD47) inhibits IL-12 release by monocytes. The suppression occurred after T cell-dependent or -independent stimulation of monocytes and was selective for IL-12 inasmuch as the production of tumor necrosis factor (TNF)-alpha, IL-1, IL-6, and granulocyte/macrophage colony stimulating factor was not inhibited. CD47 ligation did not alter transforming growth factor (TGF)-beta and IL-10 production, and the suppressive effect on IL 12 was not due to autocrine secretion of TGF-beta or IL-10. The IL-12 inhibition was not mediated by Fcgamma receptor ligation, did not require extracellular Ca(2+) influx, but was reversed by two phosphoinositide 3-kinase inhibitors (wortmannin and Ly294002). Thus, engagement of CD47 on monocytes by TSP, which transiently accumulates at the inflammatory site, is a novel and unexplored pathway to selectively downregulate IL-12 response. The pathway may be relevant in limiting the duration and intensity of the inflammatory response, and in developing novel therapeutic strategies for Th1-mediated diseases. PMID- 10523617 TI - The Src family tyrosine kinase Fyn regulates natural killer T cell development. AB - T lymphocytes express two Src tyrosine kinases, Lck and Fyn. While thymocyte and T cell subsets are largely normal in fyn(-/-) mice, animals lacking Lck have impaired T cell development. Here, it is shown that Fyn is required for the rapid burst of interleukin (IL)-4 and IL-13 synthesis, which occurs promptly after T cell receptor activation. The lack of cytokine induction in fyn mutant mice is due to a block in natural killer (NK) T cell development. Studies using bone marrow chimeras indicate that the defect behaves in a cell-autonomous manner, and the lack of NK T cells is probably not caused by inappropriate microenvironmental cues. Both NK T cells and conventional T cells express similar levels of Lck, implying that Fyn and Lck have distinct roles in regulating NK T cell ontogeny. The fyn mutation defines the first signaling molecule that is selectively required for NK T cell, but not for T lymphocyte or NK cell development. PMID- 10523619 TI - Sensitivity of an epstein-barr virus-positive tumor line, Daudi, to alpha interferon correlates with expression of a GC-rich viral transcript. AB - The exquisite sensitivity of the Burkitt's lymphoma (BL)-derived cell line Daudi to type I interferons has not previously been explained. Here we show that expression of an Epstein-Barr virus (EBV) transcript, designated D-HIT (Y. Gao et al., J. Virol. 71:84-94, 1997), correlates with the sensitivity of different Daudi cell isolates (or that of other EBV-carrying cells, where known) to alpha interferon (IFN-alpha). D-HIT, transcribed from a GC-rich repetitive region (IR4) of the viral genome, is highly structured, responding to RNase digestion in a manner akin to double-stranded RNA. Comparing EBV-carrying BL cell lines with differing responses to IFN-alpha, we found the protein levels of the dsRNA activated kinase, PKR, to be similar, whereas the levels of the autophosphorylated active form of PKR varied in a manner that correlated with endogenous levels of D-HIT expression. In a classical in vitro kinase assay, addition of either poly(I)-poly(C) or an in vitro-transcribed D-HIT homolog stimulated the autophosphorylation activity of PKR from IFN-alpha-treated cells in both EBV-positive and EBV-negative B lymphocytes. By transfection experiments, these RNAs were shown to reduce cell proliferation and to sensitize otherwise relatively insensitive Raji cells to IFN-alpha. The data lead to a model wherein the D-HIT viral RNA also serves as a possible transcriptional activator of IFN alpha or cellular genes regulated by this cytokine. PMID- 10523620 TI - An open reading frame element mediates posttranscriptional regulation of tropoelastin and responsiveness to transforming growth factor beta1. AB - Elastin, an extracellular component of arteries, lung, and skin, is produced during fetal and neonatal growth. We reported previously that the cessation of elastin production is controlled by a posttranscriptional mechanism. Although tropoelastin pre-mRNA is transcribed at the same rate in neonates and adults, marked instability of the fully processed transcript bars protein production in mature tissue. Using RNase protection, we identified a 10-nucleotide sequence in tropoelastin mRNA near the 5' end of the sequences coded by exon 30 that interacts specifically with a developmentally regulated cytosolic 50-kDa protein. Binding activity increased as tropoelastin expression dropped, being low in neonatal fibroblasts and high in adult cells, and treatment with transforming growth factor beta1 (TGF-beta1), which stimulates tropoelastin expression by stabilizing its mRNA, reduced mRNA-binding activity. No other region of tropoelastin mRNA interacted with cellular proteins, and no binding activity was detected in nuclear extracts. The ability of the exon-30 element to control mRNA decay and responsiveness to TGF-beta1 was assessed by three distinct functional assays: (i) insertion of exon 30 into a heterologous gene conferred increased reporter activity after exposure to TGF-beta1; (ii) addition of excess exon 30 RNA slowed tropoelastin mRNA decay in an in vitro polysome degradation assay; and (iii) a mutant tropoelastin cDNA lacking exon 30, compared to wild-type cDNA, produced a stable transcript whose levels were not affected by TGF-beta1. These findings demonstrate that posttranscriptional regulation of elastin production in mature tissue is conferred by a specific element within the open reading frame of tropoelastin mRNA. PMID- 10523618 TI - Caveolins, liquid-ordered domains, and signal transduction. PMID- 10523621 TI - DNA methylation is the primary silencing mechanism for a set of germ line- and tumor-specific genes with a CpG-rich promoter. AB - A subset of male germ line-specific genes, the MAGE-type genes, are activated in many human tumors, where they produce tumor-specific antigens recognized by cytolytic T lymphocytes. Previous studies on gene MAGE-A1 indicated that transcription factors regulating its expression are present in all tumor cell lines whether or not they express the gene. The analysis of two CpG sites located in the promoter showed a strong correlation between expression and demethylation. It was also shown that MAGE-A1 transcription was induced in cell cultures treated with demethylating agent 5'-aza-2'-deoxycytidine. We have now analyzed all of the CpG sites within the 5' region of MAGE-A1 and show that for all of them, demethylation correlates with the transcription of the gene. We also show that the induction of MAGE-A1 with 5'-aza-2'-deoxycytidine is stable and that in all the cell clones it correlates with demethylation, indicating that demethylation is necessary and sufficient to produce expression. Conversely, transfection experiments with in vitro-methylated MAGE-A1 sequences indicated that heavy methylation suffices to stably repress the gene in cells containing the transcription factors required for expression. Most MAGE-type genes were found to have promoters with a high CpG content. Remarkably, although CpG-rich promoters are classically unmethylated in all normal tissues, those of MAGE-A1 and LAGE-1 were highly methylated in somatic tissues. In contrast, they were largely unmethylated in male germ cells. We conclude that MAGE-type genes belong to a unique subset of germ line-specific genes that use DNA methylation as a primary silencing mechanism. PMID- 10523622 TI - Eukaryotic translation initiation factor 4AIII (eIF4AIII) is functionally distinct from eIF4AI and eIF4AII. AB - Eukaryotic initiation factor 4A (eIF4A) is an RNA-dependent ATPase and ATP dependent RNA helicase that is thought to melt the 5' proximal secondary structure of eukaryotic mRNAs to facilitate attachment of the 40S ribosomal subunit. eIF4A functions in a complex termed eIF4F with two other initiation factors (eIF4E and eIF4G). Two isoforms of eIF4A, eIF4AI and eIF4AII, which are encoded by two different genes, are functionally indistinguishable. A third member of the eIF4A family, eIF4AIII, whose human homolog exhibits 65% amino acid identity to human eIF4AI, has also been cloned from Xenopus and tobacco, but its function in translation has not been characterized. In this study, human eIF4AIII was characterized biochemically. While eIF4AIII, like eIF4AI, exhibits RNA dependent ATPase activity and ATP-dependent RNA helicase activity, it fails to substitute for eIF4AI in an in vitro-reconstituted 40S ribosome binding assay. Instead, eIF4AIII inhibits translation in a reticulocyte lysate system. In addition, whereas eIF4AI binds independently to the middle and carboxy-terminal fragments of eIF4G, eIF4AIII binds to the middle fragment only. These functional differences between eIF4AI and eIF4AIII suggest that eIF4AIII might play an inhibitory role in translation under physiological conditions. PMID- 10523625 TI - Saccharomyces cerevisiae RNA polymerase I terminates transcription at the Reb1 terminator in vivo. AB - We have mapped transcription termination sites for RNA polymerase I in the yeast Saccharomyces cerevisiae. S1 nuclease mapping shows that the primary terminator is the Reb1p terminator located at +93 downstream of the 3' end of 25S rRNA. Reverse transcription coupled with quantitative PCR shows that approximately 90% of all transcripts terminate at this site. Transcripts which read through the +93 site quantitatively terminate at a fail-safe terminator located further downstream at +250. Inactivation of Rnt1p (an RNase III involved in processing the 3' end of 25S rRNA) greatly stabilizes transcripts extending to both sites and increases readthrough at the +93 site. In vivo assay of mutants of the Reb1p terminator shows that this site operates in vivo by the same mechanism as has previously been delineated through in vitro studies. PMID- 10523624 TI - A sampling of the yeast proteome. AB - In this study, we examined yeast proteins by two-dimensional (2D) gel electrophoresis and gathered quantitative information from about 1,400 spots. We found that there is an enormous range of protein abundance and, for identified spots, a good correlation between protein abundance, mRNA abundance, and codon bias. For each molecule of well-translated mRNA, there were about 4,000 molecules of protein. The relative abundance of proteins was measured in glucose and ethanol media. Protein turnover was examined and found to be insignificant for abundant proteins. Some phosphoproteins were identified. The behavior of proteins in differential centrifugation experiments was examined. Such experiments with 2D gels can give a global view of the yeast proteome. PMID- 10523623 TI - Identification of a bidirectional splicing enhancer: differential involvement of SR proteins in 5' or 3' splice site activation. AB - The adenovirus E1A pre-mRNA undergoes alternative splicing whose modulation occurs during infection, through the use of three different 5' splice sites and of one major or one minor 3' splice site. Although this pre-mRNA has been extensively used as a model to compare the transactivation properties of SR proteins, no cis-acting element has been identified in the transcript sequence. Here we describe the identification and the characterization of a purine-rich splicing enhancer, located just upstream of the 12S 5' splice site, which is formed from two contiguous 9-nucleotide (nt) purine motifs (Pu1 and Pu2). We demonstrate that this sequence is a bidirectional splicing enhancer (BSE) in vivo and in vitro, because it activates both the downstream 12S 5' splice site through the Pu1 motif and the upstream 216-nt intervening sequence (IVS) 3' splice site through both motifs. UV cross-linking and immunoprecipitation experiments indicate that the BSE interacts with several SR proteins specifically, among them 9G8 and ASF/SF2, which bind preferentially to the Pu1 and Pu2 motifs, respectively. Interestingly, we show by in vitro complementation assays that SR proteins have distinct transactivatory properties. In particular, 9G8, but not ASF/SF2 or SC35, is able to strongly activate the recognition of the 12S 5' splice site in a BSE-dependent manner in wild-type E1A or in a heterologous context, whereas ASF/SF2 or SC35, but not 9G8, activates the upstream 216-nt IVS splicing. Thus, our results identify a novel exonic BSE and the SR proteins which are involved in its differential activity. PMID- 10523626 TI - A region within the RAP74 subunit of human transcription factor IIF is critical for initiation but dispensable for complex assembly. AB - Human transcription factor IIF (TFIIF) is an alpha(2)beta(2) heterotetramer of RNA polymerase II-associating 74 (RAP74) and RAP30 subunits. Mutagenic analysis shows that the N-terminal region of RAP74 between L155 (leucine at codon 155) and M177 is important for initiation. Mutants in this region have reduced activity in transcription, but none are inactive. Single amino acid substitutions at hydrophobic residues L155, W164, I176, and M177 have similar activity to RAP74(1 158), from which all but three amino acids of this region are deleted. Residual activity can be explained because each of these mutants forms a complex with RAP30 and recruits RNA polymerase II into the preinitiation complex. Mutants are defective for formation of the first phosphodiester bond from the adenovirus major late promoter but do not appear to have an additional significant defect in promoter escape. Negative DNA supercoiling partially compensates for the defects of TFIIF mutants in initiation, indicating that TFIIF may help to untwist the DNA helix for initiation. PMID- 10523627 TI - Functional independence and interdependence of the Src homology domains of phospholipase C-gamma1 in B-cell receptor signal transduction. AB - B-cell receptor (BCR)-induced activation of phospholipase C-gamma1 (PLCgamma1) and PLCgamma2 is crucial for B-cell function. While several signaling molecules have been implicated in PLCgamma activation, the mechanism coupling PLCgamma to the BCR remains undefined. The role of PLCgamma1 SH2 and SH3 domains at different steps of BCR-induced PLCgamma1 activation was examined by reconstitution in a PLCgamma-negative B-cell line. PLCgamma1 membrane translocation required a functional SH2 N-terminal [SH2(N)] domain, was decreased by mutation of the SH3 domain, but was unaffected by mutation of the SH2(C) domain. Tyrosine phosphorylation did not require the SH2(C) or SH3 domains but depended exclusively on a functional SH2(N) domain, which mediated the association of PLCgamma1 with the adapter protein, BLNK. Forcing PLCgamma1 to the membrane via a myristoylation signal did not bypass the SH2(N) domain requirement for phosphorylation, indicating that the phosphorylation mediated by this domain is not due to membrane anchoring alone. Mutation of the SH2(N) or the SH2(C) domain abrogated BCR-stimulated phosphoinositide hydrolysis and signaling events, while mutation of the SH3 domain partially decreased signaling. PLCgamma1 SH domains, therefore, have interrelated but distinct roles in BCR-induced PLCgamma1 activation. PMID- 10523628 TI - Cytokine receptor common beta chain as a potential activator of cytokine withdrawal-induced apoptosis. AB - Growth factors and cytokines play an important role in supporting cellular viability of various tissues during development due to their ability to suppress the default cell death program in each cell type. To date, neither the triggering molecule nor the transduction pathway of these default apoptosis programs is understood. In this study, we explored the possibility that cytokine receptors are involved in modulating cytokine withdrawal-induced apoptosis (CWIA) in hematopoietic cells. Expression of the exogenous cytokine receptor common beta chain (betac), but not the alpha chains, accelerated CWIA in multiple cytokine dependent cell lines. Reduction of the expression level of endogenous betac by antisense transcripts resulted in prolonged survival during cytokine deprivation, suggesting a critical role of betac in modulating CWIA. Fine mapping of the betac subunit revealed that a membrane-proximal cytoplasmic sequence, designated the death enhancement region (DER), was critical to the death acceleration effect of betac. Furthermore, DER accelerated cell death either as a chimeric membrane protein or as a cytosolic protein, suggesting that DER functions independently of the cytokine receptor and membrane anchorage. Cross-linking of the chimeric membrane-bound DER molecules by antibody or of the FK506-binding protein-DER fusion protein by a synthetic dimerizing agent, AP1510, did not abrogate the death acceleration effect. Transient transfection assays further indicated that DER promoted cell death in the absence of serum in the nonhematopoietic 293 cell line. In summary, our data suggest that betac plays an important role in modulating CWIA via an anchorage-independent and aggregation-insensitive mechanism. These findings may facilitate further studies on the signaling pathways of CWIA. PMID- 10523630 TI - Cell-extracellular matrix interactions stimulate the AP-1 transcription factor in an integrin-linked kinase- and glycogen synthase kinase 3-dependent manner. AB - Integrin-mediated interactions of cells with components of the extracellular matrix regulate cell survival, cell proliferation, cell differentiation, and cell migration. Some of these physiological responses are regulated via activation of transcription factors such as activator protein 1 (AP-1). Integrin-linked kinase (ILK) is an ankyrin repeat containing serine-threonine protein kinase whose activity is rapidly and transiently stimulated by cell-fibronectin interactions as well as by insulin stimulation. ILK activates protein kinase B and inhibits the glycogen synthase kinase 3 (GSK-3) activity in a phosphatidylinositol-3 kinase (PI 3-kinase)-dependent manner. We now show that cell adhesion to fibronectin results in a rapid and transient stimulation of AP-1 activity. At the same time, the kinase activity of ILK is stimulated whereas that of GSK-3 is inhibited. This fibronectin-dependent activation of AP-1 activity is inhibited in a dose-dependent manner if the cells are transfected with wild-type GSK-3, and also by inhibitors of PI 3-kinase. Stable or transient overexpression of ILK results in a stimulation of AP-1 activity which is inhibited by cotransfection with wild-type GSK-3 and kinase-deficient ILK. Transient transfection of ILK in HEK-293 cells stimulates complex formation between an AP-1 consensus oligonucleotide and nuclear proteins containing c-jun. The formation of this complex is inhibited by cotransfection with active GSK-3 or kinase-deficient ILK, suggesting that ILK may regulate AP-1 activation by inhibiting GSK-3, which has previously been shown to be a negative regulator of AP-1. In the presence of serum, ILK has no effect on the phosphorylation of Ser-73 in the N-terminal transactivation domain of c-jun. These results demonstrate a novel signaling pathway for the adhesion-mediated stimulation of AP-1 transcriptional activity involving ILK and GSK-3 and the subsequent regulation of the c-jun-DNA interaction. PMID- 10523629 TI - DNA damage and replication checkpoints in fission yeast require nuclear exclusion of the Cdc25 phosphatase via 14-3-3 binding. AB - In fission yeast as well as in higher eukaryotic organisms, entry into mitosis is delayed in cells containing damaged or unreplicated DNA. This is accomplished in part by maintaining the Cdc25 phosphatase in a phosphorylated form that binds 14 3-3 proteins. In this study, we generated a mutant of fission yeast Cdc25 that is severely impaired in its ability to bind 14-3-3 proteins. Loss of both the DNA damage and replication checkpoints was observed in fission yeast cells expressing the 14-3-3 binding mutant. These findings indicate that 14-3-3 binding to Cdc25 is required for fission yeast cells to arrest their cell cycle in response to DNA damage and replication blocks. Furthermore, the 14-3-3 binding mutant localized almost exclusively to the nucleus, unlike wild-type Cdc25, which localized to both the cytoplasm and the nucleus. Nuclear accumulation of wild-type Cdc25 was observed when fission yeast cells were treated with leptomycin B, indicating that Cdc25 is actively exported from the nucleus. Nuclear exclusion of wild-type Cdc25 was observed upon overproduction of Rad 24, one of the two fission yeast 14-3-3 proteins, indicating that one function of Rad 24 is to keep Cdc25 out of the nucleus. In support of this conclusion, Rad 24 overproduction did not alter the nuclear location of the 14-3-3 binding mutant. These results indicate that 14-3-3 binding contributes to the nuclear exclusion of Cdc25 and that the nuclear exclusion of Cdc25 is required for a normal checkpoint response to both damaged and unreplicated DNA. PMID- 10523631 TI - Context-dependent modulation of replication activity of Saccharomyces cerevisiae autonomously replicating sequences by transcription factors. AB - Evidence for transcription factor involvement in the initiation of DNA replication at certain replication origins in Saccharomyces cerevisiae mainly comes from an indirect assay which measures the mitotic stability of plasmids containing an autonomously replicating sequence (ARS), a selectable marker gene, and a centromere. In order to eliminate the effect of transcription factor binding to the selectable marker gene or centromere in such assays, we have adapted the DpnI assay to directly measure ARS replication activity in vivo by using ARS plasmids devoid of extraneous transcription elements. Using this assay, we found that the B3 element of ARS1, which serves as a binding site for the transcription factor Abf1p, does not stimulate ARS activity on plasmids lacking a centromere and a selectable marker gene. We also found with such plasmids that exogenous expression of the strong transcriptional activators Gal4 and Gal4-VP16 inhibited the replication activity of ARS1 when B3 was replaced by the Gal4 binding site, although these activators had previously been shown to stimulate replication activity in the stability assay. Moreover, a chromosomally inactive ARS, ARS301, which was active by itself on a plasmid, was inactivated by placing an Abf1p binding site in its vicinity. These results indicate that the sequences surrounding the ARS as well as properties of the ARS element itself determine its response to transcription factors. PMID- 10523633 TI - Rb and prohibitin target distinct regions of E2F1 for repression and respond to different upstream signals. AB - E2F transcription factor is subject to stringent regulation by a variety of molecules. We recently observed that prohibitin, a potential tumor suppressor protein, binds to the retinoblastoma (Rb) protein and represses E2F transcriptional activity. Here we demonstrate that prohibitin requires the marked box region of E2F for repression; further, prohibitin can effectively inhibit colony formation induced by overexpression of E2F1 in T47D cells. Prohibitin was also found to interact with the signaling kinase c-Raf-1, and Raf-1 could effectively reverse prohibitin-mediated repression of E2F activity. Agents such as E1A, p38 kinase, and cyclins D and E had no effect on prohibitin-mediated repression of E2F1, but all of these molecules could reverse Rb function. Similarly, stimulation of the immunoglobulin M signaling pathway in Ramos cells could inactivate prohibitin, but this had no effect on Rb function. Serum stimulation of quiescent Ramos cells inactivated Rb and prohibitin with different kinetics; further, while the serum-dependent inactivation of Rb was dependent on cyclin-dependent kinase activity, the inactivation of prohibitin was not. We believe that prohibitin is a novel regulator of E2F function which channels specific signaling cascades to the cell cycle regulatory machinery. PMID- 10523632 TI - Targeted disruption of the murine fps/fes proto-oncogene reveals that Fps/Fes kinase activity is dispensable for hematopoiesis. AB - The fps/fes proto-oncogene encodes a cytoplasmic protein-tyrosine kinase that is functionally implicated in the survival and terminal differentiation of myeloid progenitors and in signaling from several members of the cytokine receptor superfamily. To gain further insight into the physiological function of fps/fes, we targeted the mouse locus with a kinase-inactivating missense mutation. Mutant Fps/Fes protein was expressed at normal levels in these mice, but it lacked detectable kinase activity. Homozygous mutant animals were viable and fertile, and they showed no obvious defects. Flow cytometry analysis of bone marrow showed no statistically significant differences in the levels of myeloid, erythroid, or B-cell precursors. Subtle abnormalities observed in mutant mice included slightly elevated total leukocyte counts and splenomegaly. In bone marrow hematopoietic progenitor cell colony-forming assays, mutant mice gave slightly elevated numbers and variable sizes of CFU-granulocyte macrophage in response to interleukin-3 (IL 3) and granulocyte-macrophage colony-stimulating factor (GM-CSF). Tyrosine phosphorylation of Stat3 and Stat5A in bone marrow-derived macrophages was dramatically reduced in response to GM-CSF but not to IL-3 or IL-6. This suggests a distinct nonredundant role for Fps/Fes in signaling from the GM-CSF receptor that does not extend to the closely related IL-3 receptor. Lipopolysaccharide induced Erk1/2 activation was also reduced in mutant macrophages. These subtle molecular phenotypes suggest a possible nonredundant role for Fps/Fes in myelopoiesis and immune responses. PMID- 10523634 TI - Point mutations in yeast CBF5 can abolish in vivo pseudouridylation of rRNA. AB - In budding yeast (Saccharomyces cerevisiae), the majority of box H/ACA small nucleolar RNPs (snoRNPs) have been shown to direct site-specific pseudouridylation of rRNA. Among the known protein components of H/ACA snoRNPs, the essential nucleolar protein Cbf5p is the most likely pseudouridine (Psi) synthase. Cbf5p has considerable sequence similarity to Escherichia coli TruBp, a known Psi synthase, and shares the "KP" and "XLD" conserved sequence motifs found in the catalytic domains of three distinct families of known and putative Psi synthases. To gain additional evidence on the role of Cbf5p in rRNA biosynthesis, we have used in vitro mutagenesis techniques to introduce various alanine substitutions into the putative Psi synthase domain of Cbf5p. Yeast strains expressing these mutated cbf5 genes in a cbf5Delta null background are viable at 25 degrees C but display pronounced cold- and heat-sensitive growth phenotypes. Most of the mutants contain reduced levels of Psi in rRNA at extreme temperatures. Substitution of alanine for an aspartic acid residue in the conserved XLD motif of Cbf5p (mutant cbf5D95A) abolishes in vivo pseudouridylation of rRNA. Some of the mutants are temperature sensitive both for growth and for formation of Psi in the rRNA. In most cases, the impaired growth phenotypes are not relieved by transcription of the rRNA from a polymerase II driven promoter, indicating the absence of polymerase I-related transcriptional defects. There is little or no abnormal accumulation of pre-rRNAs in these mutants, although preferential inhibition of 18S rRNA synthesis is seen in mutant cbf5D95A, which lacks Psi in rRNA. A subset of mutations in the Psi synthase domain impairs association of the altered Cbf5p proteins with selected box H/ACA snoRNAs, suggesting that the functional catalytic domain is essential for that interaction. Our results provide additional evidence that Cbf5p is the Psi synthase component of box H/ACA snoRNPs and suggest that the pseudouridylation of rRNA, although not absolutely required for cell survival, is essential for the formation of fully functional ribosomes. PMID- 10523635 TI - BCR/ABL directly inhibits expression of SHIP, an SH2-containing polyinositol-5 phosphatase involved in the regulation of hematopoiesis. AB - The BCR/ABL oncogene causes chronic myelogenous leukemia (CML), a myeloproliferative disorder characterized by clonal expansion of hematopoietic progenitor cells and granulocyte lineage cells. The SH2-containing inositol-5 phosphatase SHIP is a 145-kDa protein which has been shown to regulate hematopoiesis in mice. Targeted disruption of the murine SHIP gene results in a myeloproliferative syndrome characterized by a dramatic increase in numbers of granulocyte-macrophage progenitor cells in the marrow and spleen. Also, hematopoietic progenitor cells from SHIP(-/-) mice are hyperresponsive to certain hematopoietic growth factors, a phenotype very similar to the effects of BCR/ABL in murine cells. In a series of BCR/ABL-transformed hematopoietic cell lines, Philadelphia chromosome (Ph)-positive cell lines, and primary cells from patients with CML, the expression of SHIP was found to be absent or substantially reduced compared to untransformed cell lines or leukemia cells lacking BCR/ABL. Ba/F3 cells in which expression of BCR/ABL was under the control of a tetracycline inducible promoter showed rapid loss of p145 SHIP, coincident with induction of BCR/ABL expression. Also, an ABL-specific tyrosine kinase inhibitor, CGP57148B (STI571), rapidly caused reexpression of SHIP, indicating that BCR/ABL directly, but reversibly, regulates the expression of SHIP protein. The estimated half-life of SHIP protein was reduced from 18 h to less than 3 h. However, SHIP mRNA also decreased in response to BCR/ABL, suggesting that SHIP protein levels could be affected by more than one mechanism. Reexpression of SHIP in BCR/ABL-transformed Ba/F3 cells altered the biological behavior of cells in culture. The reduction of SHIP due to BCR/ABL is likely to directly contribute to the pathogenesis of CML. PMID- 10523636 TI - In vivo analysis of functional regions within yeast Rap1p. AB - We have analyzed the in vivo importance of different regions of Rap1p, a yeast transcriptional regulator and telomere binding protein. A yeast strain (SCR101) containing a regulatable RAP1 gene was used to test functional complementation by a range of Rap1p derivatives. These experiments demonstrated that the C terminus of the protein, containing the putative transcriptional activation domain and the regions involved in silencing and telomere function, is not absolutely essential for cell growth, a result confirmed by sporulation of a diploid strain containing a C terminal deletion derivative of RAP1. Northern analysis with cells that expressed Rap1p lacking the transcriptional activation domain revealed that this region is important for the expression of only a subset of Rap1p-activated genes. The one essential region within Rap1p is the DNA binding domain. We have investigated the possibility that this region has additional functions. It contains two Myb-like subdomains separated by a linker region. Individual point mutations in the linker region had no effect on Rap1p function, although deletion of the region abolished cell growth. The second Myb-like subdomain contains a large unstructured loop of unknown function. Domain swap experiments with combinations of elements from DNA binding domains of Rap1p homologues from different yeasts revealed that major changes can be made to the amino acid composition of this region without affecting Rap1p function. PMID- 10523637 TI - Multiple Cbfa/AML sites in the rat osteocalcin promoter are required for basal and vitamin D-responsive transcription and contribute to chromatin organization. AB - Three Cbfa motifs are strategically positioned in the bone-specific rat osteocalcin (rOC) promoter. Sites A and B flank the vitamin D response element in the distal promoter and sites B and C flank a positioned nucleosome in the proximal promoter. The functional significance of each Cbfa element was addressed by mutating individual or multiple Cbfa sites within the context of the -1.1-kb rOC promoter fused to a chloramphenicol acetyltransferase reporter gene. Promoter activity was assayed following transient transfection and after stable genomic integration in ROS 17/2.8 osteoblastic cell lines. We show that all three Cbfa sites are required for maximal basal expression of the rOC promoter. However, the distal sites A and B each contribute significantly more (P < 0.001) to promoter activity than site C. In a genomic context, sites A and B can largely compensate for a mutation at the proximal site C, and paired mutations involving site A (mAB or mAC) result in a far greater loss of activity than the mBC mutation. Strikingly, mutation of the three Cbfa sites leads to abrogation of responsiveness to vitamin D. Vitamin D-enhanced activity is also not observed when sites A and B are mutated. Significantly, related to these losses in transcriptional activity, mutation of the three Cbfa sites results in altered chromatin structure as reflected by loss of DNase I-hypersensitive sites at the vitamin D response element and over the proximal tissue-specific basal promoter. These findings strongly support a multifunctional role for Cbfa factors in regulating gene expression, not only as simple transcriptional transactivators but also by facilitating modifications in promoter architecture and chromatin organization. PMID- 10523639 TI - The Rpb4 subunit of fission yeast Schizosaccharomyces pombe RNA polymerase II is essential for cell viability and similar in structure to the corresponding subunits of higher eukaryotes. AB - Both the gene and the cDNA encoding the Rpb4 subunit of RNA polymerase II were cloned from the fission yeast Schizosaccharomyces pombe. The cDNA sequence indicates that Rpb4 consists of 135 amino acid residues with a molecular weight of 15,362. As in the case of the corresponding subunits from higher eukaryotes such as humans and the plant Arabidopsis thaliana, Rpb4 is smaller than RPB4 from the budding yeast Saccharomyces cerevisiae and lacks several segments, which are present in the S. cerevisiae RPB4 subunit, including the highly charged sequence in the central portion. The RPB4 subunit of S. cerevisiae is not essential for normal cell growth but is required for cell viability under stress conditions. In contrast, S. pombe Rpb4 was found to be essential even under normal growth conditions. The fraction of RNA polymerase II containing RPB4 in exponentially growing cells of S. cerevisiae is about 20%, but S. pombe RNA polymerase II contains the stoichiometric amount of Rpb4 even at the exponential growth phase. In contrast to the RPB4 homologues from higher eukaryotes, however, S. pombe Rpb4 formed stable hybrid heterodimers with S. cerevisiae RPB7, suggesting that S. pombe Rpb4 is similar, in its structure and essential role in cell viability, to the corresponding subunits from higher eukaryotes. However, S. pombe Rpb4 is closer in certain molecular functions to S. cerevisiae RPB4 than the eukaryotic RPB4 homologues. PMID- 10523638 TI - An ATP/ADP-dependent molecular switch regulates the stability of p53-DNA complexes. AB - Interaction with DNA is essential for the tumor suppressor functions of p53. We now show, for the first time, that the interaction of p53 with DNA can be stabilized by small molecules, such as ADP and dADP. Our results also indicate an ATP/ADP molecular switch mechanism which determines the off-on states for p53-DNA binding. This ATP/ADP molecular switch requires dimer-dimer interaction of the p53 tetramer. Dissociation of p53-DNA complexes by ATP is independent of ATP hydrolysis. Low-level ATPase activity is nonetheless associated with ATP-p53 interaction and may serve to regenerate ADP-p53, thus recycling the high-affinity DNA binding form of p53. The ATP/ADP regulatory mechanism applies to two distinct types of p53 interaction with DNA, namely, sequence-specific DNA binding (via the core domain of the p53 protein) and binding to sites of DNA damage (via the C terminal domain). Further studies indicate that ADP not only stabilizes p53-DNA complexes but also renders the complexes susceptible to dissociation by specific p53 binding proteins. We propose a model in which the DNA binding functions of p53 are regulated by an ATP/ADP molecular switch, and we suggest that this mechanism may function during the cellular response to DNA damage. PMID- 10523641 TI - The yeast ras/cyclic AMP pathway induces invasive growth by suppressing the cellular stress response. AB - Haploid yeast cells are capable of invading agar when grown on rich media. Cells of the Sigma1278b genetic background manifest this property, whereas other laboratory strains are incapable of invasive growth. We show that disruption of the RAS2 gene in the Sigma1278b background significantly reduces invasive growth but that expression of a constitutively active Ras2p (Ras2(Val19)p) in this strain has a minimal effect on its invasiveness. On the other hand, expression of Ras2(Val19)p in another laboratory strain, SP1, rendered it invasive. These results suggest that a hyperactive Ras2 pathway induces invasive growth and that this pathway might be overactive in the Sigma1278b genetic background. Indeed, cells of the Sigma1278b are defective in the induction of stress-responsive genes, while their Gcn4 target genes are constitutively transcribed. This pattern of gene expression was previously shown to be associated with an active Ras/cyclic AMP (cAMP) pathway. We show that suppression of stress-related genes in Sigma1278b cells is a result of their inability to activate transcription through the stress response element (STRE). Disruption of RAS2, which abolished invasiveness, induced an increase in STRE activity. Further, in the SP1 genetic background, disruption of either the MSN2/4 genes (encoding activators of STRE) or the yAP-1 gene was sufficient to restore invasive growth in ras2Delta cells. We conclude that Ras2-mediated suppression of the stress response is sufficient to induce invasiveness. Accordingly, the fact that the stress response is suppressed in Sigma1278b background explains its invasiveness. It seems that invasiveness is a phenotype related to unregulated growth and is therefore manifested by cells harboring an overactive Ras/cAMP cascade. In this respect, invasiveness in yeast is reminiscent of the property of ras-transformed fibroblasts to invade soft agar. PMID- 10523640 TI - Requirement for Ras/Rac1-mediated p38 and c-Jun N-terminal kinase signaling in Stat3 transcriptional activity induced by the Src oncoprotein. AB - Signal transducers and activators of transcription (STATs) are transcription factors that mediate normal biologic responses to cytokines and growth factors. However, abnormal activation of certain STAT family members, including Stat3, is increasingly associated with oncogenesis. In fibroblasts expressing the Src oncoprotein, activation of Stat3 induces specific gene expression and is required for cell transformation. Although the Src tyrosine kinase induces constitutive Stat3 phosphorylation on tyrosine, activation of Stat3-mediated gene regulation requires both tyrosine and serine phosphorylation of Stat3. We investigated the signaling pathways underlying the constitutive Stat3 activation in Src oncogenesis. Expression of Ras or Rac1 dominant negative protein blocks Stat3 mediated gene regulation induced by Src in a manner consistent with dependence on p38 and c-Jun N-terminal kinase (JNK). Both of these serine/threonine kinases and Stat3 serine phosphorylation are constitutively induced in Src-transformed fibroblasts. Furthermore, inhibition of p38 and JNK activities suppresses constitutive Stat3 serine phosphorylation and Stat3-mediated gene regulation. In vitro kinase assays with purified full-length Stat3 as the substrate show that both JNK and p38 can phosphorylate Stat3 on serine. Moreover, inhibition of p38 activity and thus of Stat3 serine phosphorylation results in suppression of transformation by v-Src but not v-Ras, consistent with a requirement for Stat3 serine phosphorylation in Src transformation. Our results demonstrate that Ras- and Rac1-mediated p38 and JNK signals are required for Stat3 transcriptional activity induced by the Src oncoprotein. These findings delineate a network of tyrosine and serine/threonine kinase signaling pathways that converge on Stat3 in the context of oncogenesis. PMID- 10523642 TI - JSAP1, a novel jun N-terminal protein kinase (JNK)-binding protein that functions as a Scaffold factor in the JNK signaling pathway. AB - The major components of the mitogen-activated protein kinase (MAPK) cascades are MAPK, MAPK kinase (MAPKK), and MAPKK kinase (MAPKKK). Recent rapid progress in identifying members of MAPK cascades suggests that a number of such signaling pathways exist in cells. To date, however, how the specificity and efficiency of the MAPK cascades is maintained is poorly understood. Here, we have identified a novel mouse protein, termed Jun N-terminal protein kinase (JNK)/stress-activated protein kinase-associated protein 1 (JSAP1), by a yeast two-hybrid screen, using JNK3 MAPK as the bait. Of the mammalian MAPKs tested (JNK1, JNK2, JNK3, ERK2, and p38alpha), JSAP1 preferentially coprecipitated with the JNKs in cotransfected COS 7 cells. JNK3 showed a higher binding affinity for JSAP1, compared with JNK1 and JNK2. In similar cotransfection studies, JSAP1 also interacted with SEK1 MAPKK and MEKK1 MAPKKK, which are involved in the JNK cascades. The regions of JSAP1 that bound JNK, SEK1, and MEKK1 were distinct from one another. JNK and MEKK1 also bound JSAP1 in vitro, suggesting that these interactions are direct. In contrast, only the activated form of SEK1 associated with JSAP1 in cotransfected COS-7 cells. The unstimulated SEK1 bound to MEKK1; thus, SEK1 might indirectly associate with JSAP1 through MEKK1. Although JSAP1 coprecipitated with MEK1 MAPKK and Raf-1 MAPKKK, and not MKK6 or MKK7 MAPKK, in cotransfected COS-7 cells, MEK1 and Raf-1 do not interfere with the binding of SEK1 and MEKK1 to JSAP1, respectively. Overexpression of full-length JSAP1 in COS-7 cells led to a considerable enhancement of JNK3 activation, and modest enhancement of JNK1 and JNK2 activation, by the MEKK1-SEK1 pathway. Deletion of the JNK- or MEKK1-binding regions resulted in a significant reduction in the enhancement of the JNK3 activation in COS-7 cells. These results suggest that JSAP1 functions as a scaffold protein in the JNK3 cascade. We also discuss a scaffolding role for JSAP1 in the JNK1 and JNK2 cascades. PMID- 10523643 TI - Heterodimerization between members of the Nur subfamily of orphan nuclear receptors as a novel mechanism for gene activation. AB - We have recently shown that the orphan nuclear receptor Nur77 (NGFI-B) is most active in transcription when it is interacting with a cognate DNA sequence as a homodimer. Further, we have shown that the target for Nur77 dimers, the Nur response element (NurRE), is responsive to physiological stimuli in both endocrine and lymphoid cells, whereas other DNA targets of Nur77 action are not. The Nur77 subfamily also includes two related receptors, Nur-related factor 1 (Nurr1) and neuron-derived orphan receptor 1 (NOR-1). Often, more than one member of this subfamily is induced in response to extracellular signals. We now show that Nur77 and Nurr1 form heterodimers in vitro in the presence or absence of NurRE, and we have documented interactions between these proteins in vivo by using a two-hybrid system in mammalian cells. These heterodimers synergistically enhance transcription from NurRE reporters in comparison to that seen with homodimers. The naturally occurring NurRE from the pro-opiomelanocortin gene preferentially binds and activates transcription in the presence of Nur77 homo- or heterodimers, while a consensus NurRE sequence does not show this preference. Taken together, the data indicate that members of the Nur77 subfamily are most potent as heterodimers and that different dimers exhibit target sequence preference. Thus, we propose that a combinatorial code relying on specific NurRE sequences might be responsible for the activation of subsets of target genes by one of the members of the Nur77 subfamily of transcription factors. PMID- 10523644 TI - Separation-of-function mutations in Saccharomyces cerevisiae MSH2 that confer mismatch repair defects but do not affect nonhomologous-tail removal during recombination. AB - Yeast Msh2p forms complexes with Msh3p and Msh6p to repair DNA mispairs that arise during DNA replication. In addition to their role in mismatch repair (MMR), the MSH2 and MSH3 gene products are required to remove 3' nonhomologous DNA tails during genetic recombination. The mismatch repair genes MSH6, MLH1, and PMS1, whose products interact with Msh2p, are not required in this process. We have identified mutations in MSH2 that do not disrupt genetic recombination but confer a strong defect in mismatch repair. Twenty-four msh2 mutations that conferred a dominant negative phenotype for mismatch repair were isolated. A subset of these mutations mapped to residues in Msh2p that were analogous to mutations identified in human nonpolyposis colorectal cancer msh2 kindreds. Approximately half of the these MMR-defective mutations retained wild-type or nearly wild-type activity for the removal of nonhomologous DNA tails during genetic recombination. The identification of mutations in MSH2 that disrupt mismatch repair without affecting recombination provides a first step in dissecting the Msh-effector protein complexes that are thought to play different roles during DNA repair and genetic recombination. PMID- 10523645 TI - Mutations in VPS16 and MRT1 stabilize mRNAs by activating an inhibitor of the decapping enzyme. AB - Decapping is a rate-limiting step in the decay of many yeast mRNAs; the activity of the decapping enzyme therefore plays a significant role in determining RNA stability. Using an in vitro decapping assay, we have identified a factor, Vps16p, that regulates the activity of the yeast decapping enzyme, Dcp1p. Mutations in the VPS16 gene result in a reduction of decapping activity in vitro and in the stabilization of both wild-type and nonsense-codon-containing mRNAs in vivo. The mrt1-3 allele, previously shown to affect the turnover of wild-type mRNAs, results in a similar in vitro phenotype. Extracts from both vps16 and mrt1 mutant strains inhibit the activity of purified Flag-Dcp1p. We have identified a 70-kDa protein which copurifies with Flag-Dcp1p as the abundant Hsp70 family member Ssa1p/2p. Intriguingly, the interaction with Ssa1p/2p is enhanced in strains with mutations in vps16 or mrt1. We propose that Hsp70s may be involved in the regulation of mRNA decapping. PMID- 10523647 TI - CHOP enhancement of gene transcription by interactions with Jun/Fos AP-1 complex proteins. AB - The transcription factor CHOP (C/EBP homologous protein 10) is a bZIP protein induced by a variety of stimuli that evoke cellular stress responses and has been shown to arrest cell growth and to promote programmed cell death. CHOP cannot form homodimers but forms stable heterodimers with the C/EBP family of activating transcription factors. Although initially characterized as a dominant negative inhibitor of C/EBPs in the activation of gene transcription, CHOP-C/EBP can activate certain target genes. Here we show that CHOP interacts with members of the immediate-early response, growth-promoting AP-1 transcription factor family, JunD, c-Jun, and c-Fos, to activate promoter elements in the somatostatin, JunD, and collagenase genes. The leucine zipper dimerization domain is required for interactions with AP-1 proteins and transactivation of transcription. Analyses by electrophoretic mobility shift assays and by an in vivo mammalian two-hybrid system for protein-protein interactions indicate that CHOP interacts with AP-1 proteins inside cells and suggest that it is recruited to the AP-1 complex by a tethering mechanism rather than by direct binding of DNA. Thus, CHOP not only is a negative or a positive regulator of C/EBP target genes but also, when tethered to AP-1 factors, can activate AP-1 target genes. These findings establish the existence of a new mechanism by which CHOP regulates gene expression when cells are exposed to cellular stress. PMID- 10523646 TI - PBX and MEIS as non-DNA-binding partners in trimeric complexes with HOX proteins. AB - HOX, PBX, and MEIS transcription factors bind DNA through a homeodomain. PBX proteins bind DNA cooperatively as heterodimers with MEIS family members and also with HOX proteins from paralog groups 1 to 10. MEIS proteins cooperatively bind DNA with ABD-B class HOX proteins of groups 9 and 10. Here, we examine aspects of dimeric and higher-order interactions between these three homeodomain classes. The most significant results can be summarized as follows. (i) Most of PBX N terminal to the homeodomain is required for efficient cooperative binding with HOXD4 and HOXD9. (ii) MEIS and PBX proteins form higher-order complexes on a heterodimeric binding site. (iii) Although MEIS does not cooperatively bind DNA with ANTP class HOX proteins, it does form a trimer as a non-DNA-binding partner with DNA-bound PBX-HOXD4. (iv) The N terminus of HOXD4 negatively regulates trimer formation. (v) MEIS forms a similar trimer with DNA-bound PBX-HOXD9. (vi) A related trimer (where MEIS is a non-DNA-binding partner) is formed on a transcriptional promoter within the cell. (vii) We observe an additional trimer class involving non-DNA-bound PBX and DNA-bound MEIS-HOXD9 or MEIS-HOXD10 heterodimers that is enhanced by mutation of the PBX homeodomain. (viii) In this latter trimer, PBX is likely to contact both MEIS and HOXD9/D10. (ix) The stability of DNA binding by all trimers is enhanced relative to the heterodimers. These findings suggest novel functions for PBX and MEIS in modulating the function of DNA-bound MEIS-HOX and PBX-HOX heterodimers, respectively. PMID- 10523648 TI - Structural and functional cross-talk between a distant enhancer and the epsilon globin gene promoter shows interdependence of the two elements in chromatin. AB - We investigated the requirements for enhancer-promoter communication by using the human beta-globin locus control region (LCR) DNase I-hypersensitive site 2 (HS2) enhancer and the epsilon-globin gene in chromatinized minichromosomes in erythroid cells. Activation of globin genes during development is accompanied by localized alterations of chromatin structure, and CACCC binding factors and GATA 1, which interact with both globin promoters and the LCR, are believed to be critical for globin gene transcription activation. We found that an HS2 element mutated in its GATA motif failed to remodel the epsilon-globin promoter or activate transcription yet HS2 nuclease accessibility did not change. Accessibility and transcription were reduced at promoters with mutated GATA-1 or CACCC sites. Strikingly, these mutations also resulted in reduced accessibility at HS2. In the absence of a globin gene, HS2 is similarly resistant to nuclease digestion. In contrast to observations in Saccharomyces cerevisiae, HS2-dependent promoter remodeling was diminished when we mutated the TATA box, crippling transcription. This mutation also reduced HS2 accessibility. The results indicate that the epsilon-globin promoter and HS2 interact both structurally and functionally and that both upstream activators and the basal transcription apparatus contribute to the interaction. Further, at least in this instance, transcription activation and promoter remodeling by a distant enhancer are not separable. PMID- 10523649 TI - A human TATA binding protein-related protein with altered DNA binding specificity inhibits transcription from multiple promoters and activators. AB - The TATA binding protein (TBP) plays a central role in eukaryotic and archael transcription initiation. We describe the isolation of a novel 23-kDa human protein that displays 41% identity to TBP and is expressed in most human tissue. Recombinant TBP-related protein (TRP) displayed barely detectable binding to consensus TATA box sequences but bound with slightly higher affinities to nonconsensus TATA sequences. TRP did not substitute for TBP in transcription reactions in vitro. However, addition of TRP potently inhibited basal and activated transcription from multiple promoters in vitro and in vivo. General transcription factors TFIIA and TFIIB bound glutathione S-transferase-TRP in solution but failed to stimulate TRP binding to DNA. Preincubation of TRP with TFIIA inhibited TBP-TFIIA-DNA complex formation and addition of TFIIA overcame TRP-mediated transcription repression. TRP transcriptional repression activity was specifically reduced by mutations in TRP that disrupt the TFIIA binding surface but not by mutations that disrupt the TFIIB or DNA binding surface of TRP. These results suggest that TFIIA is a primary target of TRP transcription inhibition and that TRP may modulate transcription by a novel mechanism involving the partial mimicry of TBP functions. PMID- 10523650 TI - p57(Kip2) stabilizes the MyoD protein by inhibiting cyclin E-Cdk2 kinase activity in growing myoblasts. AB - We show that expression of p57(Kip2), a potent tight-binding inhibitor of several G(1) cyclin-cyclin-dependent kinase (Cdk) complexes, increases markedly during C2C12 myoblast differentiation. We examined the effect of p57(Kip2) on the activity of the transcription factor MyoD. In transient transfection assays, transcriptional transactivation of the mouse muscle creatine kinase promoter by MyoD was enhanced by the Cdk inhibitors. In addition, p57(Kip2), p21(Cip1), and p27(Kip1) but not p16(Ink4a) induced an increased level of MyoD protein, and we show that MyoD, an unstable nuclear protein, was stabilized by p57(Kip2). Forced expression of p57(Kip2) correlated with hypophosphorylation of MyoD in C2C12 myoblasts. A dominant-negative Cdk2 mutant arrested cells at the G(1) phase transition and induced hypophosphorylation of MyoD. Furthermore, phosphorylation of MyoD by purified cyclin E-Cdk2 complexes was inhibited by p57(Kip2). In addition, the NH2 domain of p57(Kip2) necessary for inhibition of cyclin E-Cdk2 activity was sufficient to inhibit MyoD phosphorylation and to stabilize it, leading to its accumulation in proliferative myoblasts. Taken together, our data suggest that repression of cyclin E-Cdk2-mediated phosphorylation of MyoD by p57(Kip2) could play an important role in the accumulation of MyoD at the onset of myoblast differentiation. PMID- 10523651 TI - RPH1 and GIS1 are damage-responsive repressors of PHR1. AB - The Saccharomyces cerevisiae DNA repair gene PHR1 encodes a photolyase that catalyzes the light-dependent repair of pyrimidine dimers. PHR1 expression is induced at the level of transcription by a variety of DNA-damaging agents. The primary regulator of the PHR1 damage response is a 39-bp sequence called URS(PHR1) which is the binding site for a protein(s) that constitutes the damage responsive repressor PRP. In this communication, we report the identification of two proteins, Rph1p and Gis1p, that regulate PHR1 expression through URS(PHR1). Both proteins contain two putative zinc fingers that are identical throughout the DNA binding region, and deletion of both RPH1 and GIS1 is required to fully derepress PHR1 in the absence of damage. Derepression of PHR1 increases the rate and extent of photoreactivation in vivo, demonstrating that the damage response of PHR1 enhances cellular repair capacity. In vitro footprinting and binding competition studies indicate that the sequence AG(4) (C(4)T) within URS(PHR1) is the binding site for Rph1p and Gis1p and suggests that at least one additional DNA binding component is present in the PRP complex. PMID- 10523652 TI - Dual transforming activities of the FUS (TLS)-ERG leukemia fusion protein conferred by two N-terminal domains of FUS (TLS). AB - The FUS (TLS)-ERG chimeric protein associated with t(16;21)(p11;q22) acute myeloid leukemia is structurally similar to the Ewing's sarcoma chimeric transcription factor EWS-ERG. We found that both FUS-ERG and EWS-ERG could induce anchorage-independent proliferation of the mouse fibroblast cell line NIH 3T3. However, only FUS-ERG was able to inhibit the differentiation into neutrophils of a mouse myeloid precursor cell line L-G and induce its granulocyte colony stimulating factor-dependent growth. We constructed several deletion mutants of FUS-ERG lacking a part of the N-terminal FUS region. A deletion mutant lacking the region between amino acids 1 and 173 (exons 1 to 5) lost the NIH 3T3 transforming activity but retained the L-G-transforming activity. On the other hand, a mutant lacking the region between amino acids 174 and 265 (exons 6 and 7) lost the L-G-transforming activity but retained the NIH 3T3-transforming activity. These results indicate that the N-terminal region of FUS contains two independent functional domains required for the NIH 3T3 and L-G transformation, which we named TR1 and TR2, respectively. Although EWS intrinsically possessed the TR2 domain, the EWS-ERG construct employed lacked the EWS sequence containing this domain. Since the TR2 domain is always found in chimeric proteins identified from t(16;21) leukemia patients but not in chimeric proteins from Ewing's sarcoma patients, it seems that the TR2 function is required only for the leukemogenic potential. In addition, we identified three cellular genes whose expression was altered by ectopic expression of FUS-ERG and found that these are regulated in either a TR1-dependent or a TR2-dependent manner. These results suggest that FUS ERG may activate two independent oncogenic pathways during the leukemogenic process by modulating the expression of two different groups of genes simultaneously. PMID- 10523653 TI - Differential regulation of the cell wall integrity mitogen-activated protein kinase pathway in budding yeast by the protein tyrosine phosphatases Ptp2 and Ptp3. AB - Mitogen-activated protein kinases (MAPKs) are inactivated by dual-specificity and protein tyrosine phosphatases (PTPs) in yeasts. In Saccharomyces cerevisiae, two PTPs, Ptp2 and Ptp3, inactivate the MAPKs, Hog1 and Fus3, with different specificities. To further examine the functions and substrate specificities of Ptp2 and Ptp3, we tested whether they could inactivate a third MAPK, Mpk1, in the cell wall integrity pathway. In vivo and in vitro evidence indicates that both PTPs inactivate Mpk1, but Ptp2 is the more effective negative regulator. Multicopy expression of PTP2, but not PTP3, suppressed growth defects due to the MEK kinase mutation, BCK1-20, and the MEK mutation, MKK1-386, that hyperactivate this pathway. In addition, deletion of PTP2, but not PTP3, exacerbated growth defects due to MKK1-386. Other evidence supported a role for Ptp3 in this pathway. Expression of MKK1-386 was lethal in the ptp2Delta ptp3Delta strain but not in either single PTP deletion strain. In addition, the ptp2Delta ptp3Delta strain showed higher levels of heat stress-induced Mpk1-phosphotyrosine than the wild-type strain or strains lacking either PTP. The PTPs also showed differences in vitro. Ptp2 was more efficient than Ptp3 at binding and dephosphorylating Mpk1. Another factor that may contribute to the greater effectiveness of Ptp2 is its subcellular localization. Ptp2 is predominantly nuclear whereas Ptp3 is cytoplasmic, suggesting that active Mpk1 is present in the nucleus. Last, PTP2 but not PTP3 transcript increased in response to heat shock in a Mpk1-dependent manner, suggesting that Ptp2 acts in a negative feedback loop to inactivate Mpk1. PMID- 10523654 TI - Control of meiotic recombination and gene expression in yeast by a simple repetitive DNA sequence that excludes nucleosomes. AB - Tandem repeats of the pentanucleotide 5'-CCGNN (where N indicates any base) were previously shown to exclude nucleosomes in vitro (Y. -H. Wang and J. D. Griffith, Proc. Natl. Acad. Sci. USA 93:8863-8867, 1996). To determine the in vivo effects of these sequences, we replaced the upstream regulatory sequences of the HIS4 gene of Saccharomyces cerevisiae with either 12 or 48 tandem copies of CCGNN. Both tracts activated HIS4 transcription. We found that (CCGNN)(12) tracts elevated meiotic recombination (hot spot activity), whereas the (CCGNN)(48) tract repressed recombination (cold spot activity). In addition, a "pure" tract of (CCGAT)(12) activated both transcription and meiotic recombination. We suggest that the cold spot activity of the (CCGNN)(48) tract is related to the phenomenon of the suppressive interactions of adjacent hot spots previously described in yeast (Q.-Q. Fan, F. Xu, and T. D. Petes, Mol. Cell. Biol. 15:1679-1688, 1995; Q. Q. Fan, F. Xu, M. A. White, and T. D. Petes, Genetics 145:661-670, 1997; T.-C. Wu and M. Lichten, Genetics 140:55-66, 1995; L. Xu and N. Kleckner, EMBO J. 16:5115 5128, 1995). PMID- 10523655 TI - Antagonistic effects of protein kinase C alpha and delta on both transformation and phospholipase D activity mediated by the epidermal growth factor receptor. AB - Downregulation of protein kinase C delta (PKC delta) by treatment with the tumor promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) transforms cells that overexpress the non-receptor class tyrosine kinase c-Src (Z. Lu et al., Mol. Cell. Biol. 17:3418-3428, 1997). We extended these studies to cells overexpressing a receptor class tyrosine kinase, the epidermal growth factor (EGF) receptor (EGFR cells); like c-Src, the EGF receptor is overexpressed in several human tumors. In contrast with expectations, downregulation of PKC isoforms with TPA did not transform the EGFR cells; however, treatment with EGF did transform these cells. Since TPA downregulates all phorbol ester-responsive PKC isoforms, we examined the effects of PKC delta- and PKC alpha-specific inhibitors and the expression of dominant negative mutants for both PKC delta and alpha. Consistent with a tumor-suppressing function for PKC delta, the PKC delta specific inhibitor rottlerin and a dominant negative PKC delta mutant transformed the EGFR cells in the absence of EGF. In contrast, the PKC alpha-specific inhibitor Go6976 and expression of a dominant negative PKC alpha mutant blocked the transformed phenotype induced by both EGF and PKC delta inhibition. Interestingly, both rottlerin and EGF induced substantial increases in phospholipase D (PLD) activity, which is commonly elevated in response to mitogenic stimuli. The elevation of PLD activity in response to inhibiting PKC delta, like transformation, was dependent upon PKC alpha and restricted to the EGFR cells. These data demonstrate that PKC isoforms alpha and delta have antagonistic effects on both transformation and PLD activity and further support a tumor suppressor role for PKC delta that may be mediated by suppression of tyrosine kinase-dependent increases in PLD activity. PMID- 10523656 TI - The Mre11-Rad50-Xrs2 protein complex facilitates homologous recombination-based double-strand break repair in Saccharomyces cerevisiae. AB - Saccharomyces cerevisiae mre11Delta mutants are profoundly deficient in double strand break (DSB) repair, indicating that the Mre11-Rad50-Xrs2 protein complex plays a central role in the cellular response to DNA DSBs. In this study, we examined the role of the complex in homologous recombination, the primary mode of DSB repair in yeast. We measured survival in synchronous cultures following irradiation and scored sister chromatid and interhomologue recombination genetically. mre11Delta strains were profoundly sensitive to ionizing radiation (IR) throughout the cell cycle. Mutant strains exhibited decreased frequencies of IR-induced sister chromatid and interhomologue recombination, indicating a general deficiency in homologous recombination-based DSB repair. Since a nuclease deficient mre11 mutant was not impaired in these assays, it appears that the role of the S. cerevisiae Mre11-Rad50-Xrs2 protein complex in facilitating homologous recombination is independent of its nuclease activities. PMID- 10523658 TI - The TFIIIC90 subunit of TFIIIC interacts with multiple components of the RNA polymerase III machinery and contains a histone-specific acetyltransferase activity. AB - Human transcription factor IIIC (hTFIIIC) is a multisubunit complex that directly recognizes promoter elements and recruits TFIIIB and RNA polymerase III. Here we describe the cDNA cloning and characterization of the 90-kDa subunit (hTFIIIC90) that is present within a DNA-binding subcomplex (TFIIIC2) of TFIIIC. hTFIIIC90 has no specific homology to any of the known yeast TFIIIC subunits. Immunodepletion and immunoprecipitation studies indicate that hTFIIIC90 is a bona fide subunit of TFIIIC2 and absolutely required for RNA polymerase III transcription. hTFIIIC90 shows interactions with the hTFIIIC220, hTFIIIC110, and hTFIIIC63 subunits of TFIIIC, the hTFIIIB90 subunit of TFIIIB, and the human RPC39 (hRPC39) and hRPC62 subunits of an initiation-specific subcomplex of RNA polymerase III. These interactions may facilitate both TFIIIB and RNA polymerase III recruitment to the preinitiation complex by TFIIIC. We show that hTFIIIC90 has an intrinsic histone acetyltransferase activity with a substrate specificity for histone H3. PMID- 10523657 TI - RelB modulation of IkappaBalpha stability as a mechanism of transcription suppression of interleukin-1alpha (IL-1alpha), IL-1beta, and tumor necrosis factor alpha in fibroblasts. AB - Members of the NF-kappaB/RelB family of transcription factors play important roles in the regulation of inflammatory and immune responses. RelB, a member of this family, has been characterized as a transcription activator and is involved in the constitutive NF-kappaB activity in lymphoid tissues. However, in a previous study we observed an overexpression of chemokines in RelB-deficient fibroblasts. Here we show that RelB is an important transcription suppressor in fibroblasts which limits the expression of proinflammatory mediators and may exert its function by modulating the stability of IkappaBalpha protein. Fibroblasts from relb(-/-) mice overexpress interleukin-1alpha (IL-1alpha), IL 1beta, and tumor necrosis factor alpha in response to lipopolysaccharide (LPS) stimulation. These cells have an augmented and prolonged LPS-inducible IKK activity and an accelerated degradation which results in a diminished level of IkappaBalpha protein, despite an upregulated IkappaBalpha mRNA expression. Consequently, NF-kappaB activity was augmented and postinduction repression of NF kappaB activity was impaired in these cells. The increased kappaB-binding activity and cytokine overexpression was suppressed by introducing RelB cDNA or a dominant negative IkappaBalpha into relb(-/-) fibroblasts. Our findings suggest a novel transcription suppression function of RelB in fibroblasts. PMID- 10523659 TI - Dual lipid modification of the yeast ggamma subunit Ste18p determines membrane localization of Gbetagamma. AB - The pheromone response in the yeast Saccharomyces cerevisiae is mediated by a heterotrimeric G protein. The Gbetagamma subunit (a complex of Ste4p and Ste18p) is associated with both internal and plasma membranes, and a portion is not stably associated with either membrane fraction. Like Ras, Ste18p contains a farnesyl-directing CaaX box motif (C-terminal residues 107 to 110) and a cysteine residue (Cys 106) that is a potential site for palmitoylation. Mutant Ste18p containing serine at position 106 (mutation ste18-C106S) migrated more rapidly than wild-type Ste18p during sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The electrophoretic mobility of wild-type Ste18p (but not the mutant Ste18p) was sensitive to hydroxylamine treatment, consistent with palmitoyl modification at Cys 106. Furthermore, immunoprecipitation of the Gbetagamma complex from cells cultured in the presence of [(3)H]palmitic acid resulted in two radioactive species on nonreducing SDS-PAGE gels, with molecular weights corresponding to Ggamma and Gbetagamma. Substitution of serine for either Cys 107 or Cys 106 resulted in the failure of Gbetagamma to associate with membranes. The Cys 107 substitution also resulted in reduced steady-state accumulation of Ste18p, suggesting that the stability of Ste18p requires modification at Cys 107. All of the mutant forms of Ste18p formed complexes with Ste4p, as assessed by coimmunoprecipitation. We conclude that tight membrane attachment of the wild-type Gbetagamma depends on palmitoylation at Cys 106 and prenylation at Cys 107 of Ste18p. PMID- 10523660 TI - Characterization of a vacuolar pyrophosphatase in Trypanosoma brucei and its localization to acidocalcisomes. AB - Inorganic pyrophosphate promoted the acidification of an intracellular compartment in permeabilized procyclic trypomastigotes of Trypanosoma brucei, as measured by acridine orange uptake. The proton gradient generated by pyrophosphate was collapsed by addition of nigericin or NH(4)Cl. Pyrophosphate driven proton translocation was stimulated by potassium ions and inhibited by KF, by the pyrophosphate analogs imidodiphosphate and aminomethylenediphosphonate (AMDP), and by the thiol reagent p-hydroxymercuribenzoate at concentrations similar to those that inhibit the plant vacuolar H(+)-pyrophosphatase (PPase). The proton translocation activity had a pH optimum around 7.5 and was partially inhibited by 7-chloro-4-nitrobenz-2-oxa-1,3-diazole (10 microM) and unaffected by bafilomycin A(1) (40 nM), concanamycin A (5 nM), sodium o-vanadate (500 microM), oligomycin (1 microM), N-ethylmaleimide (100 microM), and KNO(3). AMDP-sensitive pyrophosphate hydrolysis was detected in both procyclic and bloodstream trypomastigotes. Measurements of acridine orange uptake in permeabilized procyclic trypomastigotes in the presence of different substrates and inhibitors suggested the presence of H(+)-ATPase, H(+)-PPase, and (ADP-dependent) H(+)/Na(+) antiport activity in the same compartment. Separation of bloodstream and procyclic trypomastigote extracts on Percoll gradients yielded fractions that contained H(+)-PPase (both stages) and H(+)/Na(+) exchanger (procyclics) activities but lacked markers for mitochondria, glycosomes, and lysosomes. The organelles in these fractions were identified by electron microscopy and X-ray microanalysis as acidocalcisomes (electron-dense vacuoles). These results provide further evidence for the unique nature of acidocalcisomes in comparison with other, previously described, organelles. PMID- 10523661 TI - The retinoblastoma protein is linked to the activation of Ras. AB - The inner membrane-bound protein Ras integrates various extracellular signals that are subsequently communicated from the cytoplasm to the nucleus via the Raf/MEK/MAPK cascade. Here we show that the retinoblastoma protein pRb, previously reported to be a nuclear target of this pathway, can in turn influence the activation state of Ras. Rb-deficient fibroblasts display elevated levels (up to 30-fold) of activated Ras during G(1). Expression of wild-type pRb or a number of pRb mutants defective in E2F regulation reverses this effect. We provide evidence that the mid-G(1) activation of Ras in Rb-deficient cells, which occurs at the level of guanine nucleotide binding, differs from that of epidermal growth factor-induced stimulation of Ras, being dependent on protein synthesis. The aberrant levels of Ras activity associated with loss of pRb may be responsible for the differentiation defects in Rb-deficient cells, because suppression of Ras activity in Rb(-/-) fibroblasts restores the transactivation function of MyoD and the expression of a late marker of skeletal muscle differentiation. These data suggest that nuclear-cytoplasmic communication between pRb and Ras is bidirectional. PMID- 10523662 TI - Pta1, a component of yeast CF II, is required for both cleavage and poly(A) addition of mRNA precursor. AB - CF II, a factor required for cleavage of the 3' ends of mRNA precursor in Saccharomyces cerevisiae, has been shown to contain four polypeptides. The three largest subunits, Cft1/Yhh1, Cft2/Ydh1, and Brr5/Ysh1, are homologs of the three largest subunits of mammalian cleavage-polyadenylation specificity factor (CPSF), an activity needed for both cleavage and poly(A) addition. In this report, we show by protein sequencing and immunoreactivity that the fourth subunit of CF II is Pta1, an essential 90-kDa protein originally implicated in tRNA splicing. Yth1, the yeast homolog of the CPSF 30-kDa subunit, is not detected in this complex. Extracts prepared from pta1 mutant strains are impaired in the cleavage and the poly(A) addition of both GAL7 and CYC1 substrates and exhibit little processing activity even after prolonged incubation. However, activity is efficiently rescued by the addition of purified CF II to the defective extracts. Extract from a strain with a mutation in the CF IA subunit Rna14 also restored processing, but extract from a brr5-1 strain did not. The amounts of Pta1 and other CF II subunits are reduced in pta1 strains, suggesting that levels of the subunits may be coordinately regulated. Coimmunoprecipitation experiments indicate that the CF II in extract can be found in a stable complex containing Pap1, CF II, and the Fip1 and Yth1 subunits of polyadenylation factor I. While purified CF II does not appear to retain the association with these other factors, this larger complex may be the form recruited onto pre-mRNA in vivo. The involvement of Pta1 in both steps of mRNA 3'-end formation supports the conclusion that CF II is the functional homolog of CPSF. PMID- 10523663 TI - Two new members of the emerging KDWK family of combinatorial transcription modulators bind as a heterodimer to flexibly spaced PuCGPy half-sites. AB - Initially recognized as a HeLa factor essential for parvovirus DNA replication, parvovirus initiation factor (PIF) is a site-specific DNA-binding complex consisting of p96 and p79 subunits. We have cloned and sequenced the human cDNAs encoding each subunit and characterized their products expressed from recombinant baculoviruses. The p96 and p79 polypeptides have 40% amino acid identity, focused particularly within a 94-residue region containing the sequence KDWK. This motif, first described for the Drosophila homeobox activator DEAF-1, identifies an emerging group of metazoan transcriptional modulators. During viral replication, PIF critically regulates the viral nickase, but in the host cell it probably modulates transcription, since each subunit is active in promoter activation assays and the complex binds to previously described regulatory elements in the tyrosine aminotransferase and transferrin receptor promoters. Within its recognition site, PIF binds coordinately to two copies of the tetranucleotide PuCGPy, which, remarkably, can be spaced from 1 to 15 nucleotides apart, a novel flexibility that we suggest may be characteristic of the KDWK family. Such tetranucleotides are common in promoter regions, particularly in activating transcription factor/cyclic AMP response element-binding protein (ATF/CREB) and E box motifs, suggesting that PIF may modulate the transcription of many genes. PMID- 10523664 TI - The glycine-phenylalanine-rich region determines the specificity of the yeast Hsp40 Sis1. AB - Hsp40s are ubiquitous, conserved proteins which function with molecular chaperones of the Hsp70 class. Sis1 is an essential Hsp40 of the cytosol of Saccharomyces cerevisiae, thought to be required for initiation of translation. We carried out a genetic analysis to determine the regions of Sis1 required to perform its key function(s). A C-terminal truncation of Sis1, removing 231 amino acids but retaining the N-terminal 121 amino acids encompassing the J domain and the glycine-phenylalanine-rich (G-F) region, was able to rescue the inviability of a Deltasis1 strain. The yeast cytosol contains other Hsp40s, including Ydj1. To determine which regions carried the critical determinants of Sis1 function, we constructed chimeric genes containing portions of SIS1 and YDJ1. A chimera containing the J domain of Sis1 and the G-F region of Ydj1 could not rescue the lethality of the Deltasis1 strain. However, a chimera with the J domain of Ydj1 and the G/F region of Sis1 could rescue the strain's lethality, indicating that the G-F region is a unique region required for the essential function of Sis1. However, a J domain is also required, as mutants expected to cause a disruption of the interaction of the J domain with Hsp70 are inviable. We conclude that the G-F region, previously thought only to be a linker or spacer region between the J domain and C-terminal regions of Hsp40s, is a critical determinant of Sis1 function. PMID- 10523665 TI - Dependence of Dbl and Dbs transformation on MEK and NF-kappaB activation. AB - Dbs was identified initially as a transforming protein and is a member of the Dbl family of proteins (>20 mammalian members). Here we show that Dbs, like its rat homolog Ost and the closely related Dbl, exhibited guanine nucleotide exchange activity for the Rho family members RhoA and Cdc42, but not Rac1, in vitro. Dbs transforming activity was blocked by specific inhibitors of RhoA and Cdc42 function, demonstrating the importance of these small GTPases in Dbs-mediated growth deregulation. Although Dbs transformation was dependent upon the structural integrity of its pleckstrin homology (PH) domain, replacement of the PH domain with a membrane localization signal restored transforming activity. Thus, the PH domain of Dbs (but not Dbl) may be important in modulating association with the plasma membrane, where its GTPase substrates reside. Both Dbs and Dbl activate multiple signaling pathways that include activation of the Elk-1, Jun, and NF-kappaB transcription factors and stimulation of transcription from the cyclin D1 promoter. We found that Elk-1 and NF-kappaB, but not Jun, activation was necessary for Dbl and Dbs transformation. Finally, we have observed that Dbl and Dbs regulated transcription from the cyclin D1 promoter in a NF-kappaB-dependent manner. Previous studies have dissociated actin cytoskeletal activity from the transforming potential of RhoA and Cdc42. These observations, when taken together with those of the present study, suggest that altered gene expression, and not actin reorganization, is the critical mediator of Dbl and Rho family protein transformation. PMID- 10523666 TI - A role for protein kinase Bbeta/Akt2 in insulin-stimulated GLUT4 translocation in adipocytes. AB - Insulin stimulates glucose uptake into muscle and fat cells by promoting the translocation of glucose transporter 4 (GLUT4) to the cell surface. Phosphatidylinositide 3-kinase (PI3K) has been implicated in this process. However, the involvement of protein kinase B (PKB)/Akt, a downstream target of PI3K in regulation of GLUT4 translocation, has been controversial. Here we report that microinjection of a PKB substrate peptide or an antibody to PKB inhibited insulin-stimulated GLUT4 translocation to the plasma membrane by 66 or 56%, respectively. We further examined the activation of PKB isoforms following treatment of cells with insulin or platelet-derived growth factor (PDGF) and found that PKBbeta is preferentially expressed in both rat and 3T3-L1 adipocytes, whereas PKBalpha expression is down-regulated in 3T3-L1 adipocytes. A switch in growth factor response was also observed when 3T3-L1 fibroblasts were differentiated into adipocytes. While PDGF was more efficacious than insulin in stimulating PKB phosphorylation in fibroblasts, PDGF did not stimulate PKBbeta phosphorylation to any significant extent in adipocytes, as assessed by several methods. Moreover, insulin, but not PDGF, stimulated the translocation of PKBbeta to the plasma membrane and high-density microsome fractions of 3T3-L1 adipocytes. These results support a role for PKBbeta in insulin-stimulated glucose transport in adipocytes. PMID- 10523667 TI - Evidence for distinct substrate specificities of importin alpha family members in nuclear protein import. AB - Importin alpha plays a pivotal role in the classical nuclear protein import pathway. Importin alpha shuttles between nucleus and cytoplasm, binds nuclear localization signal-bearing proteins, and functions as an adapter to access the importin beta-dependent import pathway. In contrast to what is found for importin beta, several isoforms of importin alpha, which can be grouped into three subfamilies, exist in higher eucaryotes. We describe here a novel member of the human family, importin alpha7. To analyze specific functions of the distinct importin alpha proteins, we recombinantly expressed and purified five human importin alpha's along with importin alpha from Xenopus and Saccharomyces cerevisiae. Binding affinity studies showed that all importin alpha proteins from humans or Xenopus bind their import receptor (importin beta) and their export receptor (CAS) with only marginal differences. Using an in vitro import assay based on permeabilized HeLa cells, we compared the import substrate specificities of the various importin alpha proteins. When the substrates were tested singly, only the import of RCC1 showed a strong preference for one family member, importin alpha3, whereas most of the other substrates were imported by all importin alpha proteins with similar efficiencies. However, strikingly different substrate preferences of the various importin alpha proteins were revealed when two substrates were offered simultaneously. PMID- 10523668 TI - Does ribosomal DNA get out of the micronuclear chromosome in Paramecium tetraurelia by means of a rolling circle? AB - The macronuclear genes coding for rRNA (ribosomal DNA [rDNA]) of Paramecium tetraurelia, stock 51, are arranged in polymers consisting of units made up of a transcribed coding region and a nontranscribed spacer region. The whole macronuclear polymer ends with a portion of the spacer on either end followed by a telomere. Six kinds of macronuclear units, or genes, were mapped. Spacers were different, and transcribed regions were the same. These genes are found in markedly different numbers in the macronucleus. The most common gene shows two regions in the spacer where a sequence is followed by a direct repeat. The next most common gene is similar but shows a deletion plus a number of base pair substitutions. Although most cosmid clones contain only a single kind of gene, many contain more than one. These are thought to be produced by somatic crossing over. The four micronuclear genes that have been isolated consist of a single central transcribed region and portions of the spacer on either end. Sequencing indicates that the two ends of the molecule are partially redundant. While the spacer region at the right end of the macronuclear polymer is derived from the micronuclear spacer on the right, the spacer at the left end of the macronuclear polymer is derived from regions of the micronuclear spacer on both the right and the left. To account for this situation, a rolling-circle model for generation of the macronuclear rDNA from the micronuclear DNA is proposed. PMID- 10523669 TI - Saccharomyces cerevisiae pol30 (proliferating cell nuclear antigen) mutations impair replication fidelity and mismatch repair. AB - To understand the role of POL30 in mutation suppression, 11 Saccharomyces cerevisiae pol30 mutator mutants were characterized. These mutants were grouped based on their mutagenic defects. Many pol30 mutants harbor multiple mutagenic defects and were placed in more than one group. Group A mutations (pol30-52, 104, -108, and -126) caused defects in mismatch repair (MMR). These mutants exhibited mutation rates and spectra reminiscent of MMR-defective mutants and were defective in an in vivo MMR assay. The mutation rates of group A mutants were enhanced by a msh2 or a msh6 mutation, indicating that MMR deficiency is not the only mutagenic defect present. Group B mutants (pol30-45, -103, -105, -126, and -114) exhibited increased accumulation of either deletions alone or a combination of deletions and duplications (4 to 60 bp). All deletion and duplication breakpoints were flanked by 3 to 7 bp of imperfect direct repeats. Genetic analysis of one representative group B mutant, pol30-126, suggested polymerase slippage as the likely mutagenic mechanism. Group C mutants (pol30 100, -103, -105, -108, and -114) accumulated base substitutions and exhibited synergistic increases in mutation rate when combined with msh6 mutations, suggesting increased DNA polymerase misincorporation as a mutagenic defect. The synthetic lethality between a group A mutant, pol30-104, and rad52 was almost completely suppressed by the inactivation of MSH2. Moreover, pol30-104 caused a hyperrecombination phenotype that was partially suppressed by a msh2 mutation. These results suggest that pol30-104 strains accumulate DNA breaks in a MSH2 dependent manner. PMID- 10523670 TI - HDAC4, a human histone deacetylase related to yeast HDA1, is a transcriptional corepressor. AB - Histone acetylation plays an important role in regulating chromatin structure and thus gene expression. Here we describe the functional characterization of HDAC4, a human histone deacetylase whose C-terminal part displays significant sequence similarity to the deacetylase domain of yeast HDA1. HDAC4 is expressed in various adult human tissues, and its gene is located at chromosome band 2q37. HDAC4 possesses histone deacetylase activity intrinsic to its C-terminal domain. When tethered to a promoter, HDAC4 represses transcription through two independent repression domains, with repression domain 1 consisting of the N-terminal 208 residues and repression domain 2 containing the deacetylase domain. Through a small region located at its N-terminal domain, HDAC4 interacts with the MADS-box transcription factor MEF2C. Furthermore, HDAC4 and MEF2C individually upregulate but together downmodulate c-jun promoter activity. These results suggest that HDAC4 interacts with transcription factors such as MEF2C to negatively regulate gene expression. PMID- 10523671 TI - The Gal3p-Gal80p-Gal4p transcription switch of yeast: Gal3p destabilizes the Gal80p-Gal4p complex in response to galactose and ATP. AB - The Gal3, Gal80, and Gal4 proteins of Saccharomyces cerevisiae comprise a signal transducer that governs the galactose-inducible Gal4p-mediated transcription activation of GAL regulon genes. In the absence of galactose, Gal80p binds to Gal4p and prohibits Gal4p from activating transcription, whereas in the presence of galactose, Gal3p binds to Gal80p and relieves its inhibition of Gal4p. We have found that immunoprecipitation of full-length Gal4p from yeast extracts coprecipitates less Gal80p in the presence than in the absence of Gal3p, galactose, and ATP. We have also found that retention of Gal80p by GSTG4AD (amino acids [aa] 768 to 881) is markedly reduced in the presence compared to the absence of Gal3p, galactose, and ATP. Consistent with these in vitro results, an in vivo two-hybrid genetic interaction between Gal80p and Gal4p (aa 768 to 881) was shown to be weaker in the presence than in the absence of Gal3p and galactose. These compiled results indicate that the binding of Gal3p to Gal80p results in destabilization of a Gal80p-Gal4p complex. The destabilization was markedly higher for complexes consisting of G4AD (aa 768 to 881) than for full length Gal4p, suggesting that Gal80p relocated to a second site on full-length Gal4p. Congruent with the idea of a second site, we discovered a two-hybrid genetic interaction involving Gal80p and the region of Gal4p encompassing aa 225 to 797, a region of Gal4p linearly remote from the previously recognized Gal80p binding peptide within Gal4p aa 768 to 881. PMID- 10523672 TI - Normal skeletal development of mice lacking matrilin 1: redundant function of matrilins in cartilage? AB - Matrilin 1, or cartilage matrix protein, is a member of a novel family of extracellular matrix proteins. To date, four members of the family have been identified, but their biological role is unknown. Matrilin 1 and matrilin 3 are expressed in cartilage, while matrilin 2 and matrilin 4 are present in many tissues. Here we describe the generation and analysis of mice carrying a null mutation in the Crtm gene encoding matrilin 1. Anatomical and histological studies demonstrated normal development of homozygous mutant mice. Northern blot and biochemical analyses show no compensatory up-regulation of matrilin 2 or 3 in the cartilage of knockout mice. Although matrilin 1 interacts with the collagen II and aggrecan networks of cartilage, suggesting that it may play a role in cartilage tissue organization, studies of collagen extractability indicated that collagen fibril maturation and covalent cross-linking were unaffected by the absence of matrilin 1. Ultrastructural analysis did not reveal any abnormalities of matrix organization. These data suggest that matrilin 1 is not critically required for cartilage structure and function and that matrilin 1 and matrilin 3 may have functionally redundant roles. PMID- 10523673 TI - The levels of the bancal product, a Drosophila homologue of vertebrate hnRNP K protein, affect cell proliferation and apoptosis in imaginal disc cells. AB - We have characterized the Drosophila bancal gene, which encodes a Drosophila homologue of the vertebrate hnRNP K protein. The bancal gene is essential for the correct size of adult appendages. Reduction of appendage size in bancal mutant flies appears to be due mainly to a reduction in the number of cell divisions in the imaginal discs. Transgenes expressing Drosophila or human hnRNP K are able to rescue weak bancal phenotype, showing the functional similarity of these proteins in vivo. High levels of either human or Drosophila hnRNP K protein in imaginal discs induces programmed cell death. Expression of the antiapoptotic P35 protein suppresses this phenotype in the eye, suggesting that apoptosis is the major cellular defect caused by overexpression of K protein. Finally, the human K protein acts as a negative regulator of bancal gene expression. We propose that negative autoregulation limits the level of Bancal protein produced in vivo. PMID- 10523674 TI - Mutagenesis of SNM1, which encodes a protein component of the yeast RNase MRP, reveals a role for this ribonucleoprotein endoribonuclease in plasmid segregation. AB - RNase MRP is a ribonucleoprotein endoribonuclease that has been shown to have roles in both mitochondrial DNA replication and nuclear 5.8S rRNA processing. SNM1 encodes an essential 22.5-kDa protein that is a component of yeast RNase MRP. It is an RNA binding protein that binds the MRP RNA specifically. This 198 amino-acid protein can be divided into three structural regions: a potential leucine zipper near the amino terminus, a binuclear zinc cluster in the middle region, and a serine- and lysine-rich region near the carboxy terminus. We have performed PCR mutagenesis of the SNM1 gene to produce 17 mutants that have a conditional phenotype for growth at different temperatures. Yeast strains carrying any of these mutations as the only copy of snm1 display an rRNA processing defect identical to that in MRP RNA mutants. We have characterized these mutant proteins for RNase MRP function by examining 5.8S rRNA processing, MRP RNA binding in vivo, and the stability of the RNase MRP RNA. The results indicate two separate functional domains of the protein, one responsible for binding the MRP RNA and a second that promotes substrate cleavage. The Snm1 protein appears not to be required for the stability of the MRP RNA, but very low levels of the protein are required for processing of the 5.8S rRNA. Surprisingly, a large number of conditional mutations that resulted from nonsense and frameshift mutations throughout the coding regions were identified. The most severe of these was a frameshift at amino acid 7. These mutations were found to be undergoing translational suppression, resulting in a small amount of full length Snm1 protein. This small amount of Snm1 protein was sufficient to maintain enough RNase MRP activity to support viability. Translational suppression was accomplished in two ways. First, CEN plasmid missegregation leads to plasmid amplification, which in turn leads to SNM1 mRNA overexpression. Translational suppression of a small amount of the superabundant SNM1 mRNA results in sufficient Snm1 protein to support viability. CEN plasmid missegregation is believed to be the result of a prolonged telophase arrest that has been recently identified in RNase MRP mutants. Either the SNM1 gene is inherently susceptible to translational suppression or extremely small amounts of Snm1 protein are sufficient to maintain essential levels of MRP activity. PMID- 10523675 TI - Biological and regulatory properties of Vav-3, a new member of the Vav family of oncoproteins. AB - We report here the identification and characterization of a novel Vav family member, Vav-3. Signaling experiments demonstrate that Vav-3 participates in pathways activated by protein tyrosine kinases. Vav-3 promotes the exchange of nucleotides on RhoA, on RhoG and, to a lesser extent, on Rac-1. During this reaction, Vav-3 binds physically to the nucleotide-free states of those GTPases. These functions are stimulated by tyrosine phosphorylation in wild-type Vav-3 and become constitutively activated upon deletion of the entire calponin-homology region. Expression of truncated versions of Vav-3 leads to drastic actin relocalization and to the induction of stress fibers, lamellipodia, and membrane ruffles. Moreover, expression of Vav-3 alters cytokinesis, resulting in the formation of binucleated cells. All of these responses need only the expression of the central region of Vav-3 encompassing the Dbl homology (DH), pleckstrin homology (PH), and zinc finger (ZF) domains but do not require the presence of the C-terminal SH3-SH2-SH3 regions. Studies conducted with Vav-3 proteins containing loss-of-function mutations in the DH, PH, and ZF regions indicate that only the DH and ZF regions are essential for Vav-3 biological activity. Finally, we show that one of the functions of the Vav-3 ZF region is to work coordinately with the catalytic DH region to promote both the binding to GTP-hydrolases and their GDP-GTP nucleotide exchange. These results highlight the role of Vav-3 in signaling and cytoskeletal pathways and identify a novel functional cross-talk between the DH and ZF domains of Vav proteins that is imperative for the binding to, and activation of, Rho GTP-binding proteins. PMID- 10523677 TI - Adoptive therapy of canine metastatic mammary carcinoma with the human MHC non restricted cytotoxic T-cell line TALL-104. AB - Adoptive transfer of human TALL-104 killer cells into a dog with metastatic mammary adenocarcinoma resulted in 50% reduction of the largest lung metastasis and stabilization of the other lesions for 10 weeks, accompanied by the development of tumor-specific immune responses. Upon halting cell therapy, the dog developed new lung lesions within 10 weeks and died of slowly progressive disease. TALL-104 cell therapy of mice bearing the dog's tumor xenograft induced 65% reduction of local tumor growth and regression of lung metastases in 100% of the animals. The overall findings indicate the therapeutic potential of TALL-104 cells for canine mammary tumors. PMID- 10523676 TI - Molecular architecture of the mouse DNA polymerase alpha-primase complex. AB - The DNA polymerase alpha-primase complex is the only enzyme that provides RNA-DNA primers for chromosomal DNA replication in eukaryotes. Mouse DNA polymerase alpha has been shown to consist of four subunits, p180, p68, p54, and p46. To characterize the domain structures and subunit requirements for the assembly of the complex, we constructed eukaryotic polycistronic cDNA expression plasmids expressing pairwise the four subunits of DNA polymerase alpha. In addition, the constructs contained an internal ribosome entry site derived from poliovirus. The constructs were transfected in different combinations with vectors expressing single subunits to allow the simultaneous expression of three or four of the subunits in cultured mammalian cells. We demonstrate that the carboxyl-terminal region of p180 (residues 1235 to 1465) is essential for its interaction with both p68 and p54-p46 by immunohistochemical analysis and coprecipitation studies with antibodies. Mutations in the putative zinc fingers present in the carboxyl terminus of p180 abolished the interaction with p68 completely, although the mutants were still capable of interacting with p54-p46. Furthermore, the amino terminal region (residues 1 to 329) and the carboxyl-terminal region (residues 1280 to 1465) were revealed to be dispensable for DNA polymerase activity. Thus, we can divide the p180 subunit into three domains. The first is the amino terminal domain (residues 1 to 329), which is dispensable for both polymerase activity and subunit assembly. The second is the minimal core domain (residues 330 to 1279), required for polymerase activity. The third is the carboxyl terminal domain (residues 1280 to 1465), which is dispensable for polymerase activity but required for the interaction with the other three subunits. Taken together, these results allow us to propose the first structural model for the DNA polymerase alpha-primase complex in terms of subunit assembly, domain structure, and stepwise formation at the cellular level. PMID- 10523678 TI - Hypothesis: more mutations to cure cancer? AB - Cancer results from the accumulation of selected mutations. We propose here that some combinations of mutations may be counterselected. To illustrate this, we show an analysis of allelotyping in human breast tumors in which we demonstrate that specific associations of genetic events are statistically under-represented. This emerging concept could be used in a new therapeutical approach of cancer. PMID- 10523679 TI - Effects of oral bromelain administration on the impaired immunocytotoxicity of mononuclear cells from mammary tumor patients. AB - The protease bromelain from pineapple was suggested for adjuvant therapy of malignant diseases. We studied immunological effects of an orally applied bromelain drug on 16 breast cancer patients in comparison with healthy donors. Bromelain was applied for 10 days with a daily dose of 3000 F.I.P. units and the immunocytotoxicity of blood monocytes and lymphocytes against the leukemic K562 and MDA-MB-231 mammary carcinoma target cells was determined in vitro. In addition, the expression of the cell surface markers CD44, CD16, CD11a and CD62L on lymphocytes and the secretion of IL-2 and IL-1beta from monocytes was measured. Patients leukocytes expressed lower bMAK-, MAK-, NK- and LAK-cell activities, compared with those from healthy donors. Orally applied bromelain increased the reduced bMAK- and MAK-cell activity of patients monocytes about 2 fold. When the patients were classified on the basis of bromelain effects on the monocytic cytotoxicity into bromelain responders and nonresponders, about 40% of the patients responded to bromelain with an increase of cytotoxicity from 7.8% to 54% (bMAK-cell activity) and from 16% to 47% (MAK-cell activity). Bromelain was less effective on the higher cytotoxicity of monocytes from healthy donors, but stimulated the secretion of IL-1beta from monocytes. In contrast, patient monocytes secreted no detectable IL-1beta, before, during and after bromelain treatment. Bromelain had no effects on the impaired patients NK- and LAK-cell activity, but reduced the LAK-cell activity of healthy donors. No IL-2 was found in the supernatants of untreated and treated lymphocytes from healthy donors. Bromelain reduced the expression of CD44, but weakly increased CD11a and CD62L expression on patient lymphocytes, whereas CD16 remained unchanged. In vitro bromelain application to lymphocytes had similar effects, with greater reduction rates of CD44 and CD16 expression. As to coagulation parameters in plasma of healthy donors, the activated partial thromboplastin time was increased from 38 to 46 sec, leaving prothrombin time and plasminogen unchanged. These data suggest, that orally applied bromelain stimulates the deficient monocytic cytotoxicity of mammary tumor patients, which may partially explain its proposed antitumor activity. PMID- 10523681 TI - Pre-operative IL-2 immunoprophylaxis of cancer recurrence: long-term clinical results of a phase II study in radically operable colorectal cancer. AB - This study evaluates retrospectively the outcome of 20 colorectal cancer patients radically operated (M/F 13/7; primary/recurrent 15/5; Dukes B=11; C=6; D=3) who received pre-operative IL-2 (18,000, 000 IU/daily s.c. for 3 days) and the outcome of 40 colorectal cancer (primary/recurrent 40/0) patients age, sex and stage-matched radically operated, as control group. After a median follow-up of 72 months, in the IL-2 pre-operative group we observed 6/20 recurrences (30%) vs. 19/40 (47.5%) in controls. Mean and median disease-free period in patients who relapsed were respectively 21 months and 20.5 months (range 6-36) in IL-2 group vs. 14.1 and 12 months in the control group (range 3-34). After a 5-year follow up, 4/20 (20%) IL-2 treated patients were dead vs. 19/40 control patients (47.5%) (log-rank chi2=3.7, p=0.05). Pre-operative IL-2 administration is safe, active in preventing post-operative lymphocytopenia and seems to improve the clinical outcome in radically operated colorectal cancer patients. PMID- 10523680 TI - Xenografts of human solid tumors frequently express cellular-associated isoform of vascular endothelial growth factor (VEGF) 189. AB - Vascular endothelial growth factor (VEGF), a major factor mediating tumor stromal angiogenesis, is expressed as five splice variants encoded by a single gene (VEGF121, VEGF145, VEGF165, VEGF189 and VEGF206). Recently, we demonstrated that the cell-associated isoform, VEGF189, plays important roles in establishment of human colon and esophageal cancer xenografts. We have established 228 xenografts originating from various human solid tumors. In this study, we investigated the expression patterns of VEGF isoforms in those tumor xenografts by RT-PCR. The isoform patterns were VEGF121/VEGF165 in 27 xenografts (11.8%) and VEGF121/VEGF165/VEGF189 in 201 (88.2%). All human solid tumor xenografts expressed VEGF189 more frequently than primary tumors reported previously. These results suggest that VEGF189 contributes to the successful xenotransplantability of various human solid tumors via augmentation of stromal vascularization. PMID- 10523682 TI - The immunopotentiation effects of the bifunctional radiosensitizer KIN-806 in comparison with its analogs KIN-804 and KIN-844. AB - KIN-806 is one of the 2-nitroimidazole derivative hypoxic cell radiosensitizers. Its radiosensitizing effects and the degree of lung metastasis detected were evaluated and compared with its analogs KIN-804 and KIN-844. The immune reactions induced by these radiosensitizers were also analyzed. Female C3H/He mice and SCCVII tumor cells were used. Seventeen days after inoculation of SCCVII tumor cells into the animals, 0.4 g/kg of KIN-806, KIN-804, and KIN-844 was administered to each radiosensitizer group 30 min before 40 Gy was delivered as local irradiation. In each group, KIN-806, KIN-804, and KIN-844 markedly suppressed tumor regrowth in comparison with animals that received irradiation alone. There was no significant difference in the radiosensitizing effects among these three radiosensitizers. A marked suppression of lung metastasis and macrophage/T-lymphocyte infiltration into the tumor were observed only in the KIN 806-administered groups, regardless of the presence or absence of radiation therapy. It therefore appears likely that the lung metastasis suppression was caused by the immune reaction elicited by KIN-806, which is an excellent immunopotentiator as well as an effective radiosensitizer. PMID- 10523683 TI - Heterogeneity and clinical role of thymidine phosphorylase activity in gastric cancer. AB - We examined the dThdPase activity in primary gastric cancer and metastatic lymph nodes from human subjects. The dThdPase activities were significantly higher in primary tumors than in the normal gastric wall, particularly in the center of the tumor rather than in the periphery (P<0.01). Tumors with a high dThdPase activity often had venous invasion (P<0.01). The dThdPase activities were significantly higher in metastatic lymph nodes than in the nodes without metastasis (P<0.01). The intratumoral heterogeneity of dThdPase activity was identified and cancer cells with high dThdPase activity may indicate that metastasis will likely occur. PMID- 10523684 TI - Expression of alpha-smooth muscle actin in small bronchioloalveolar adenocarcinoma of the lung: assessment and comparison with noguchi criteria. AB - The Noguchi criteria are useful in assessing the prognosis of patients with small lung adenocarcinoma. Although there is a significant difference in prognosis between type A or B and type C, it is difficult in some cases to distinguish these types accurately by microscopy. In this study, we used immunohistochemistry to examine alpha-smooth muscle actin (alpha-SMA) produced by active fibroblasts in 25 pulmonary adenocarcinomas less than 2 cm in diameter. Eleven of type C (61%) showed positive staining for alpha-SMA, whereas no positive cases were seen in type A or B. The incidence of cancerous blood vessel and lymphatic invasion were significantly higher in alpha-SMA positive cases than in negative cases, and the positive cases showed poorer prognosis. These findings indicate that immunohistochemical detection of alpha-SMA is useful and essential for histological typing by the Noguchi criteria. PMID- 10523685 TI - Frequent allelic loss/imbalance on the long arm of chromosome 21 in oral cancer: evidence for three discrete tumor suppressor gene loci. AB - Frequent allelic imbalances including loss of heterozygosity (LOH) and microsatellite instability (MI) on the long arm of chromosome 21 (21q) have been found in several types of human cancer. This study was designed to identify tumor suppressor locus (or loci) associated with oral squamous cell carcinoma (SCC) on 21q. Among 38 patients with oral SCC tested, 15 (44%) of 34 informative cases showed LOH at one or more loci. Deletion mapping of these 15 tumors revealed three discrete commonly deleted regions on the chromosome arm. A minimal region with frequent LOH was found at the marker D21S236 mapped on 21q11.1. Another region of frequent deletion was identified between markers D21S11 and D21S1436 on 21q21, and a further commonly deleted region was found at D21S1254 on 21q22.1. In addition, we have detected MI on the chromosome arm in our oral SCC samples with significant correlation with tumor stage. Thus, our results strongly suggest that allelic imbalances on 21q may be involved in the development of oral SCC; and that at least three different putative tumor suppressor genes contributing to the pathogenesis of this disease are present on 21q. PMID- 10523687 TI - Immunohistochemical detection of occult metastases in paraaortic lymph nodes in advanced gastric cancer. AB - Some Japanese surgeons have examined the utility of super-extended paraaortic lymphadenectomy (PAL) as part of the surgical treatment for advanced gastric cancer. However, therapeutic value of this PAL remains controvertial. The purpose of this study was to evaluate appropriate candidates who might benefit from PAL by the immunostaining with cytokeratin (CK) of the macroscopically intact paraaortic nodes. A total of 525 paraaortic nodes from 35 patients was serially sectioned and stained with hematoxylin-eosin (H&E) and CK staining. A total of 17 nodes (3.2%) from 7 patients (20.0%), among 525 macroscopically intact paraaortic nodes, was determined to be immunopositive for CK. In 4 patients, 8 H&E-positive nodes with metastases were all immunopositive and, in addition, 4 H&E-negative nodes were also immunopositive. Furthermore, 3 patients with H&E-negative nodes had five immunopositive nodes. Immunostaining with CK was useful for detection of occult metastases. Survival was prolonged in 3 of these 7 patients. The incidence of CK-positive nodes was significantly higher in patients with gross type of 3 or 4 gastric cancer and in patients with H&E-detected nodal metastasis within group 3 (N3) nodes. It seems that such patients would benefit from prophylactic PAL. PMID- 10523686 TI - Usefulness of Lens culinaris agglutinin A-reactive fraction of alpha-fetoprotein (AFP-L3) as a marker of distant metastasis from hepatocellular carcinoma. AB - The objective of this study was to clarify the relationship between the serum level of the Lens culinaris agglutinin A-reactive fraction of alpha-fetoprotein (AFP-L3) and the clinical features including sex, age, Child's classification, virus markers, tumour size, tumour stage, distant metastasis, histopathologic findings, portal thrombus and outcome of hepatocellular carcinoma (HCC). We measured the AFP-L3 levels in 170 HCC cases at the time of diagnosis using lectin affinity electrophoresis followed by antibody-affinity blotting. The patients were divided into two groups, those who were AFP-L3 positive (n=56; AFP-L3 >/=15% relative to the total alpha-fetoprotein (AFP) concentration) and those who were AFP-L3 negative (n=114; AFP-L3 <15%). Then we examined the association between the serum AFP-L3 level and the clinical features of HCC. No significant differences were found in age, sex, and virus markers between the AFP-L3-positive and -negative groups. However, patients in the positive group had worse liver function and larger tumours compared to the negative group. They also had more advanced cancer with poor tumour histology compared to the negative group. Distant metastasis was diagnosed significantly more often in the positive group than that in the negative group. There was no significant correlation between the AFP-L3 level and portal thrombus. Although the follow-up period was brief the prognosis for the positive group clearly was poor. These results suggest that AFP L3 is a useful indicator of distant metastasis and a poor prognosis for HCC. PMID- 10523688 TI - Enhanced adenoviral transduction efficiency in HER-2/neu-overexpressing human breast cancer cells not induced by an integrin pathway. AB - Adenoviruses are currently widely used as vectors in gene therapy. The steps involved in adenoviral infection have been investigated, but the factors regulating viral entry to the cell have not been clearly identified. We observed a high adenoviral infection rate in HER-2/neu-overexpressing breast cancer cells in vitro (435.eb1 and MCF-7/H18) and in vivo (435.eb1). We used emodin, a tyrosine kinase inhibitor that suppresses autophosphorylation and transphosphorylation activities of the HER-2/neu tyrosine kinase, to test the role of HER-2/neu in adenoviral transduction. Emodin treatment resulted in a marked decrease in the transduction efficiency of HER-2/neu-overexpressing cells but not in the parental cells. Because previous studies have shown that epidermal growth factor and tumor growth factor-alpha increase the expression level of integrin. Because integrin alphav is known as a promotor of viral internalization, penetration, or both, we investigated whether the observed increased transduction rate in HER-2/neu transfectants was mediated through the increased expression of integrin alphav. To test this hypothesis, we examined the level of integrin alphav of in HER-2/neu overexpressing cells. We found that the level of integrin alphav expression detected in HER-2/neu overexpressing cells by immunoblot analysis was similar to the level of integrin alphav found in its parental cells. These results suggest that HER-2/neu expression may have a significant role in the viral transduction efficiency through an integrin alphav independent pathway. PMID- 10523689 TI - Ileo-rectal fistula complicating advanced ovarian carcinoma. AB - Fistula between the bowels and an ovarian carcinoma is recognized but rare complication. Internal malignant fistula of the gastrointestinal tract involving two or more loops of different segments of the bowel and genitourinary structure are rare. The colon is frequently one of the participating loops. In reviewing the literature, however, we were unable to find a previous report of ileo-rectal fistula as a complication of an ovarian carcinoma. A case report and review of the English medical literature are presented with emphasis on the cause, clinical presentation, and management of advanced ovarian cancer with ileo-rectal involvement. PMID- 10523690 TI - Sentinel node biopsy in vulvar malignancies: a preliminary feasibility study. AB - Sentinel lymph node biopsies (SLNB) were investigated in 8 cases (6 squamous cell carcinomas, 2 melanomas) of vulvar malignancy. The sentinel node was detected by patent blue dye injection (1 case), pre operative lymphoscintigraphy with intra operative gamma hand-held probe (2 cases), and combined techniques (5 cases). The procedure was successful in all cases but one (1 invasive squamous cell carcinoma) in which there was medial groin recurrence at 6 months. Nodal invasion was observed in only one case and was confined to the sentinel node. No specific morbidity related to the SLNB procedure occurred. SLNB appears to be a feasible and promising technique, however, requiring further evaluation before being considered as a reliable method to spare inguinofemoral lymphadenectomy in early stage patients free of sentinel node metastasis, or to be substituted in screening elderly clinically node-negative females. PMID- 10523692 TI - A radioimmunoassay for the measurement of paclitaxel and taxanes in biological specimens using commercially available reagents. AB - We have developed a rapid and sensitive radioimmunoassay for the measurement of paclitaxel and related taxanes in biological specimens using commercially available reagents. The presence of paclitaxel competitively inhibited the binding of radioactive paclitaxel to polyclonal anti-taxane antibody or anti taxane monoclonal antibody. When using polyclonal antibody, this method detected paclitaxel in concentrations as low as 0.87 nM and was also effective in 20% methanol. This radioimmunoassay is useful for the measurement of paclitaxel and taxanes in biological specimens like culture medium of paclitaxel-producing microorganisms and may be useful for the measurement of paclitaxel and related compounds in other potential natural sources. PMID- 10523693 TI - Mutational alteration of the p16CDKN2a tumor suppressor gene is infrequent in Ewing's sarcoma. AB - Mutational alteration of the p16CDKN2a tumor suppressor gene has been frequently observed in a variety of human cell lines and tumor tissues. To assess whether alterations of the p16CDKN2a gene play an important role in the pathogenesis of Ewing's sarcoma, we examined the allelic deletion and point mutation of the gene in 27 primary tumors. In quantitative DNA/PCR analysis for three individual exons of the gene, none of the 27 specimens showed detectable reduction in the amplification levels compared to those of normal lymphocytes. To explore the presence of any subtle sequence changes within the protein coding region, we performed a comprehensive screening of sequence alteration in the three exons using DNA/PCR-SSCP analysis. However, no abnormal shift of SSCP bands indicative of sequence change was identified. In addition, no elevation of p16CDKN2a mRNA expression was observed in the RD-ES cell line following 5'-Aza-cytidine treatment, indicating that the promoter of the gene is not abnormally methylated in this cell line. Immunohistochemical study of the same tissue specimens for p16CDKN2a and pRB also revealed grade 2+ or 3+ nuclear staining of both proteins in most of the specimens we examined. Taken together, our results strongly suggest that the mutational inactivation of the p16CDKN2a gene might be a rare event, and thus not play a critical role in the pathogenesis of Ewing's sarcoma. PMID- 10523691 TI - A correlation of APC and c-myc mRNA levels in lung cancer cell lines. AB - One of the functions of adenomatous polyposis coli (APC) in colorectal cancers is regulation of c-myc gene expression. However, the role of APC in lung cancers has not been elucidated. In the present study, the levels of APC and c-myc mRNA were determined in one strain of normal human bronchial epithelial (NHBE) cells, an SV 40-immortalized non-tumorigenic human bronchial epithelial cell line (BEAS-2B), 13 non-small cell lung cancer cell lines, and 4 small cell lung cancer cell lines. To establish a relationship between c-Myc and APC, we determined the ratio of c-myc and APC mRNA levels in different lung cancer cell lines. Out of 19 lung cancer cell lines, we found that 13 exhibited c-myc/APC mRNA ratio of more than two. Among the cell lines CaLu-3, NCI-H82, A427 and SW900 showed a very low level of APC mRNA and a high level of c-myc mRNA. The ratio of c-myc/APC mRNA in these cell lines was 48, 127, 325 and 708, respectively. The results of these analyses revealed an inverse relationship between APC and c-myc mRNA levels, suggesting that APC may regulate c-myc expression in lung cancer cells in a manner analogous to its role in colon cancer. PMID- 10523694 TI - Yeast functional assay of the p53 gene status in 11 cell lines and 26 surgical specimens of human hepatocellular carcinoma. AB - The structural abnormalities of the p53 gene have frequently been detected in hepatocellular carcinomas (HCCs). To ascertain whether or not functional inactivation of this gene also occurs in HCCs, the transactivation of p53 gene products in 11 HCC cell lines maintained in our laboratory and 26 HCC surgical specimens was examined by yeast functional assay (functional analysis of separated alleles in yeast: FASAY), which determines the functional status of the individual p53 alleles. The p53 gene product was inactivated in 8 of 11 (72.7%) HCC cell lines and in 12 of 26 (46.2%) HCC surgical specimens. The inactivation frequency of the gene was 37.5%, 36.4%, and 71.4% in well, moderately, and poorly differentiated HCCs, respectively. In HCC surgical specimens larger than 5 cm in diameter, the inactivation frequency was 83.3% while in those smaller than 2. 5 cm, it was 14.3%. These results show that functional inactivation of p53 gene products often occurs in HCCs and that the inactivation frequency of the gene in HCCs is well correlated with differentiation degree and tumor size, suggesting that the inactivation of p53 gene products plays a role in progression from well to poorly differentiated HCC. PMID- 10523695 TI - Most of the patients with cirrhosis in Japan die from hepatocellular carcinoma. AB - A total of 424 patients with cirrhosis were entered into a registry in Japan. One hundred and seven patients were hepatitis B virus (HBV) associated cirrhosis while 252 were hepatitis C virus (HCV) associated cirrhosis. Patients were followed for a period of 3.3+/-3. 3 years. Fifty-six (80%) of 70 deaths in HBV patients and 151 (90%) of 161 deaths in HCV patients were due to complications associated with hepatocellular carcinoma (HCC). The complication rate of HCC in HCV group was significantly higher than in HBV group. In conclusion, most of the patients with cirrhosis died from HCC. PMID- 10523696 TI - LOH analyses of premalignant and malignant lesions of human breast: frequent LOH in 8p, 16q, and 17q in atypical ductal hyperplasia. AB - The number of breast cancer patients is increasing yearly, and it is important to establish effective methods for clinical management. We investigated loss of heterozygosity (LOH) in tumors derived from 23 patients that harbored synchronous lesions of atypical ductal hyperplasia (ADH), ductal carcinoma in situ (DCIS), and invasive ductal carcinoma (IDC). Fourteen selected microsatellite markers that were mapped to and/or very close to the tumor suppressor genes or regions with frequent LOH in breast cancer. Our results suggested that i) losses of chromosomal regions 8p, 16q, and 17q are early genetic changes, and ii) ADH is a premalignant lesion for the development of breast cancer. PMID- 10523697 TI - Are physiological clonal expansion and maturation pathways a basis for discrepant behaviour of colon tumoral cell growth in response to the thiazolidinedione PPARgamma agonists? AB - Recently, anti-diabetic thiazolidinediones, pharmacological agonist ligands of peroxisome proliferator-activated receptor gamma (PPARgamma), have been shown to induce either protective or permissive effects towards colon epithelium tumoral cell growth. Several attractive explanations have been proposed but no final answer to these is currently provided. It is not the purpose of the authors to bring this final answer but to offer another attractive hypothesis which might help our approach to explore further this exciting field of medical research. PMID- 10523698 TI - Inhibition of ciprofibrate-induced hepatocarcinogenesis in the rat by dimethylthiourea, a scavenger of hydroxyl radical. AB - DNA damage caused by oxidative stress is considered to play an important role in peroxisome proliferator-induced hepatocarcinogenesis in rats and mice. In this study, we investigated the effect of dimethylthiourea (DMTU), a known hydroxyl radical scavenger, on ciprofibrate-induced hepatocarcinogenesis. Male F-344 rats were fed a diet containing 0.025% ciprofibrate and given daily intraperitoneal injections of DMTU (5 days a week) at a dose of 50 or 250 mg/kg body weight for 60 weeks at which time the study was terminated. Livers from all animals were analyzed grossly and microscopically for incidence, number and type of tumors. All rats given ciprofibrate alone developed altered areas, neoplastic nodules (NN) and hepatocellular carcinomas (HCC). Combined administration of ciprofibrate and DMTU resulted in inhibition of tumor development. In the group given higher doses of DMTU the incidence of NN was 100% and HCC 0%. The number of tumors per liver also significantly decreased (p<0.001). At lower dose levels DMTU caused significant reduction in the number of tumors per liver (p<0. 05) and a slight reduction (29%) in the incidence of HCC. The inhibitory effect of DMTU on ciprofibrate-induced hepatocarcinogenesis could be explained by hydroxyl radical scavenging properties of DMTU, resulting in decreased free radical induced DNA damage. PMID- 10523699 TI - Transient rapid growth of uterine leiomyoma in a postmenopausal woman. AB - We report a case of transient rapid growth of uterine leiomyoma in a postmenopausal woman. A 58-year old woman presented with a rapidly enlarging pelvic tumor, from a 14-week to a 20-week gestational size in the one month. Transcervical needle biopsy revealed a smooth muscle tumor with 5 mitotic figures per 10 high power fields. However, surgical specimen obtained 1 month later revealed less than 1 mitotic figure per 100 high power fields. This variation may influence the final histopathological diagnosis because mitotic index is believed to be one of the most reliable histopathological parameters for differentiating between uterine leiomyoma and leiomyosarcoma. PMID- 10523700 TI - Clinicopathological and immunohistochemical study of cancer arising from Barrett's esophagus. AB - In Japan, Barrett's esophageal cancer is a very rare disease. We examined clinicopathologically and immunohistologically 4 patients with Barrett's esophageal cancer who underwent surgical resection in our department. Barrett's esophageal mucosa was classified into 3 types for detailed observation. Specialized columnar epithelium (SCE) remained on the orifice side of carcinoma, and progression to adenocarcinoma was observed in some dysplastic glands. positive findings were detected on p53 immunohistochemical staining, and the ki 67 labeling index was higher than other types. SCE-type Barrett's esophagus may be a precancerous lesion arising prior to the development of adenocarcinoma. PMID- 10523701 TI - Neoadjuvant intraarterial high-dose chemotherapy and autologous peripheral blood stem cell transplantation for advanced breast cancer. AB - Eight locally advanced breast cancer patients were treated with neoadjuvant intraarterial high-dose chemotherapy (epirubicin) plus MPA combined with autologous PBSCT. All patients completed the scheduled treatment, and there were no toxic deaths. Patients were treated with an escalating dose of epirubicin (370 480 mg) and cyclophosphamide (0-6000 mg). The rate of clinical response was 100%. The rate of good histologic response was 87.5% in the main tumor and 75% in diseased lymph nodes. The 2-year survival rate was 100%. Six patients were disease-free at the time of writing. This treatment resulted in higher rates of clinical and histologic response when compared with standard-dose intraarterial chemotherapy. PMID- 10523702 TI - Metachronous multicentric occurrence of hepatocellular carcinoma after surgical treatment - clinicopathological comparison with recurrence due to metastasis. AB - This study investigated clinicopathological features of patients with recurrence of metachronous multicentric occurrence by comparison with patients with recurrence due to metastasis. In 177 patients, recurrences after curative surgical treatment were classified into recurrence due to metastasis according to criteria based on imaging findings. This group consisted of 35 patients. Among the rest of the patients, 59 underwent fine needle biopsies for recurrent tumor and, in these patients, a classification of recurrence of metachronous multicentric occurrence was made based on the histological findings of primary and recurrent tumor. This group consisted of 33 patients. The estimated incidence for recurrence of metachronous multicentric occurrence was 44.8% to total total patients. Metachronous multicentric occurrence frequently developed in patients with anti-HCV antibody and an early stage of primary tumor. In 80% of the patients who had recurrent tumor of multicentric origin, the recurrence developed within 3 postoperative years. The survival rate in patients with metachronous multicentric occurrence was significantly higher than that in patients with recurrence due to metastasis. Conclusively, the incidence of patients with recurrence of metachronous multicentric occurrence was high, but the prognosis for these patients was significantly better than that for patients with recurrence due to metastasis. PMID- 10523703 TI - Potentiation by 2-deoxy-D-glucose tetraacetate of the antitumoral action of 5 fluorouracil in mice inoculated with L1210 ascites-producing cells. AB - Ascites-producing tumoral cells of the L1210 line were inoculated to syngenic mice. The gain in body weight and survival time were measured in untreated animals and mice injected intraperitoneally 1, 5 and 9 days after the administration of the tumoral cells with 5-fluorouracil (20-200 microg/g body wt.) and/or 2-deoxy-D-glucose tetraacetate (75 or 1,350 microg/g body wt.). The two agents failed to affect adversely the growth of control mice not inoculated with the tumoral cells. 5-fluorouracil caused a dose-related decrease in the body weight gain caused by the occurrence of the tumoral ascites and a dose-related prolongation of survival time. Although 2-deoxy-D-glucose tetraacetate, when tested in the low dose of 75 microg/g body weight opposed the effect of 5 fluorouracil on survival time, a higher dose of the ester (1,350 microg/g body weight) potentiated the effect of 5-fluorouracil to both decrease body weight gain and increase survival time. It is proposed, therefore, that 2-deoxy-D glucose tetraacetate could be used to increase the cytostatic action of classical chemotherapeutic agents in the treatment of neoplastic diseases. PMID- 10523704 TI - Oxidative stress is insignificant in N1S1-transplanted hepatoma despite markedly declined activities of the antioxidant enzymes. AB - It has been proposed that persistent oxidative stress accounts for the increased levels of DNA damage in cancer tissues. We have examined the profile of anti oxidant enzymes in a transplanted hepatic tumor model by injecting N1S1 rat hepatoma cells into the liver of Sprague-Dawley rats. The transplanted N1S1 tumors displayed characteristics resembling human hepatocellular carcinoma. The immunoreactivities of catalase (CAT), manganese-superoxide dismutase (Mn SOD), copper/zinc-SOD (Cu/Zn SOD), and glutathione peroxidase (GPx) were found to decrease significantly. The enzyme activity in tumors decreased 26.2-, 4.2-, 4.5 , and 5.4-fold for CAT, Mn SOD, Cu/Zn SOD, and GPx, respectively, relative to those in normal liver tissue from the same animals. In contrast, the mRNA levels of CAT and GPx in tumors decreased only 5- and 2-fold, respectively, and the mRNA levels of Cu/Zn SOD and Mn SOD showed either no change or an increase as compared to those of normal liver tissue. The contents of 8-hydroxy-2'-deoxyguanosine (8 OH-dG) and thiobarbituric acid-reactive substances (TBARS) were comparable to those of normal controls. Furthermore, mitochondrial production of superoxide in tumors was 4 times lower than that in normal tissues. In conclusion, the data indicate that the reduced activities of anti-oxidant enzymes in the N1S1 tumor did not cause significant oxidative stress. PMID- 10523706 TI - High prevalence of HPV18 in correlation with ras gene mutations and clinicopathological parameters in cervical cancer studied from stained cytological smears. AB - The multi-event nature of carcinogenesis has led to extensive studies of oncogenes, onco-suppressor genes and viruses involved in human cancers. The collaboration of ras oncogene with HPV E6/E7 genes inducing full transformation of cervical keratinocytes has also been suggested. The purpose of this study was to detect the presence of codon 12 point mutations of ras genes, as well as to detect and identify the human papillomaviruses in stained smears of cervical malignancies and to correlate them with the clinicopathological parameters of the Greek patients. Specimens were obtained from 88 women, codon 12 point mutations of the K-ras (30%) and H-ras (10%) oncogenes, as well as HPV18 were detected at a higher rate than HPV16 (66% vs 7%) in cervical lesions by PCR-RFLP and PCR analysis, respectively. The statistical analysis of the data demonstrates correlation between the presence of K-ras mutations and FIGO stage and between FIGO stage and survival of the patients. It is suggested that ras activation combined with HPV infection may be an important step in the carcinogenesis of a substantial number of cervical carcinomas. PMID- 10523705 TI - Correlation between nm23-H1 overexpression and clinicopathological variables in human anal canal carcinoma. AB - To improve life expectancy prognostic factors other than TNM have been investigated. It is thought that nm23 protein may play a specific biological role in suppressing tumor metastasis. The purpose of this study was to elucidate the clinical significance of nm23 expression in human anal canal carcinoma. Immunostaining using anti-nm23 monoclonal antibody was performed in 22 anal canal tumors. The results were correlated with clinicopathological variables. Six cases out of 22 (27.3%) were nm23-positive. Significant association was found between nm23-H1 expression and depth of invasion, lymph node involvement and prognosis (p<0.05). There was no significant association between nm23-H1 expression, histologic type and age of the patients. nm23-H1 expression was not seen in our cases with metastasis and this may be related to nm23 gene alterations not being detectable by the monoclonal antibody used or to the presence of a subset of tumors in which nm23 gene abnormalities had not yet occurred at the time of tumor excision or biopsy. Overexpression of nm23-H1 protein in anal canal carcinoma may have implications for its metastatic potential. nm23-H1 expression would provide a more accurate evaluation of outcome for individual patients and thus improve treatment planning. PMID- 10523708 TI - Inhibitory effect of shark liver oil on cutaneous angiogenesis induced in Balb/c mice by syngeneic sarcoma L-1, human urinary bladder and human kidney tumour cells. AB - The effect of shark liver oil on cutaneous angiogenesis induced in mice by intradermal grafting of tumour cells was evaluated. It was shown that this substance (Ecomer) suppressed neovascular response in mice grafted with sarcoma L 1 syngeneic cells, human kidney cancer and human urinary bladder cancer cells. In addition, strong stimulatory effect of this drug on mice blood granulocyte number and their metabolic activity was observed. PMID- 10523707 TI - Inhibition of azoxymethane-induced aberrant crypt foci in rats by natural compounds, caffeine, quercetin and morin. AB - The modifying effects of dietary administration of natural compounds, caffeine, quercetin and morin, which are present in our daily food, on the development of azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) were investigated in rats and compared to that of a metabolic inhibitor of AOM, disulfiram. Male F344 rats were given s. c. injections of AOM (15 mg/kg body weight) once a week for 3 weeks to induce ACF. They also received the experimental diets containing one of test compounds (500 ppm) for 5 weeks, starting one week before the first dosing of AOM. At the termination of the study (week 5), AOM exposure produced 101.0+/ 10.2 ACF/rat. Disulfiram almost completely inhibited ACF development (0.60+/ 0.90, 99% reduction). Dietary administration of test compounds caused significant reduction in the frequency of ACF: caffeine (70.4+/-16.6, 30% reduction), quercetin (53.0+/-8.4, 48% reduction) and morin (37. 6+/-18.1, 63% reduction). Numbers of cells positive for proliferative cell nuclear antigen in ACF and surrounding crypts were lowered by feeding of test compounds. Feeding of these test compounds also suppressed polyamine content in the colonic mucosa and blood as did disulfiram. These findings might indicate possible chemopreventive effects of caffeine, quercetin and morin, through their modulation of cell proliferation activity in crypt cells, on colon tumorigenesis. PMID- 10523709 TI - DNA cytofluorometric analysis using HP/DAPI double staining of parathyroid carcinoma arising in a patient with chronic renal failure and secondary hyperparathyroidism. AB - A 55-year old female patient on long-term hemodialysis began to suffer from pain in her knees and ankles. An ultrasonographic study showed enlargement of all four parathyroid glands. Serum parathyroid hormone and calcium levels were increased. Parathyroidectomy was performed. The right superior gland was enlarged and adherent to surrounding tissues. The other three glands were slightly enlarged. Histologically, the largest gland was a parathyroid carcinoma because capsular and vascular invasion were observed. To our knowledge, there have been only 13 cases of parathyroid carcinomas arising in patients with chronic renal failure reported in the English literature. To evaluate the characteristics of this tumor, we measured nuclear DNA and protein content using stains for HP (hematoporphyrin)/DAPI (4,6-diamidino-2-phenylindole dihydroporphyrin chloride). The nuclear DNA pattern was typically diploid or tetraploid. The cellular protein content was similar to that seen in the controls. The discrepancy between the histology, indicative of malignancy and the cytofluorometrical findings show that it is difficult to determine the prognosis for patients with secondary hyperparathyroidism and parathyroid carcinomas solely from the results of tumor DNA cytometry. PMID- 10523710 TI - A new protocol of iron therapy combined with epoetin alpha as a treatment for preoperative autologous blood donation in gynaecological tumor surgery. AB - The use of epoetin alpha and sodium ferrous gluconate has been shown to be a safe and effective treatment which can be used to avoid allogeneic blood transfusions and to plan short term elective surgery. In this study the authors submitted 20 patients, scheduled to undergo surgery for gynaecological tumors, to a program of pre-operative autologous blood donation. All the patients received both epoetin alpha and sodium ferrous gluconate in the pre- and post-donation period. Epoetin alpha was administered subcutaneously at a dose of 200 IU/kg thrice a week during the week before and after autologous blood donation (400 ml). Sodium ferrous gluconate was administered intravenously shortly before the first and fourth administration of 125 mg epoetin alpha, and shortly before the third and sixth administration of 62.5 mg epoetin alpha. Surgery was scheduled to be performed 10 15 days after the last epoetin alpha administration, i.e. within 15-20 days from blood donation. All the patients were tested for the following blood chemistry parameters: hematocrit, haemoglobin, sideraemia and ferritin at treatment start, before donation, at treatment end, before autologous blood infusion and on the third and seventh day after surgery. No patient receiving epoetin alpha required allogeneic blood transfusion, as both the hematocrit and haemoglobin values remained normal. Epoetin alpha was observed to be a safe and effective treatment to be used in autologous blood donation programs in all patients scheduled to undergo surgery. It limits the decrease of hematocrit values following autologous blood donation thus enabling all the patients, who for a variety of reasons refuse allogeneic blood infusion, to predeposit autologous blood shortly before the date scheduled for surgery. PMID- 10523711 TI - Correlation between histological differentiation and DNA-synthesizing enzymes in rat colorectal tumors induced with 1, 2-dimethylhydrazine. AB - Thymidylate synthase and thymidine kinase are key enzymes involved in de novo and salvage pathways for pyrimidine nucleotide synthesis. Colorectal carcinogenesis induced with 1,2-dimethylhydrazine in rats enhanced mRNA expression levels of both enzymes, resulting in the increase of both enzyme activities and bromodeoxyuridine-immunoreactive S-phase cells. Poorly and well differentiated adenocarcinomas of the colorectum showed the relative elevation of activities of thymidylate synthase and thymidine kinase, respectively. These results indicate that the relationship between de novo and salvage pathways for pyrimidine nucleotide synthesis may depend on the histopathological grades of cell differentiation. PMID- 10523712 TI - Amifostine: A selective cytoprotective agent of normal tissues from chemo radiotherapy induced toxicity (Review). AB - Patients receiving systemic cancer chemotherapy must often have their dose intensity of therapeutic agents reduced, because a broad range of organs are adversely affected. Therefore, research and the development of agents protecting the normal tissues from the toxicity of antineoplastic therapy, without reducing the antitumour efficacy, are very important. Amifostine, a prodrug that forms an activated free thiol, when dephosphorylated by alkaline phosphatase, appears selective in its entry in non-malignant cells, and exerts a protective effect from toxicity induced by chemo- or radiotherapy on normal tissues, through free radical scavenging, hydrogen donation and inhibition of DNA damage. Studies in vitro and experimental models have confirmed the protective properties of amifostine in normal cells. In clinical trials pretreatment with amifostine reduced the frequency of cyclophosphamide induced neutropenia and nephro-, oto- and neurotoxicity of platinum compounds. In some cases the use of amifostine have also potentiated the effects of several drugs, such as alkylating agents and, in recent studies, taxanes. The main potentially dose-limiting adverse effect is hypotension, that is often asymptomatic. Amifostine is thus usefully employed in order to obtain a better quality of life in patients receiving oncologic treatments. PMID- 10523713 TI - The role of protein kinase C and novel phorbol ester receptors in tumor cell invasion and metastasis (Review). AB - Phorbol ester tumor promoters activate protein kinase C (PKC) isozymes and novel non-kinase receptors, suggesting a high degree of complexity in the signaling mechanisms of tumorigenesis. Many studies have shown that PKC isozymes contribute to the progression of malignant phenotype. We review the emerging understanding of the roles of PKC isozymes in the three sequential cellular processes of tumor invasion and metastasis: attachment to extracellular matrix or basement membrane components, matrix degradation by proteolytic enzymes, and migration through the digested matrix. In addition, we discuss the potential role of chimaerins, novel non-kinase phorbol ester receptors, in carcinogenesis. PMID- 10523714 TI - Comparative response of normal and of human papillomavirus-16 immortalized human epithelial cervical cells to benzo[a]pyrene. AB - Laboratory evidence suggests synergism of human papillomavirus (HPV) infection with cigarette smoking behaviors in enhancing the risk of cervical cancer. In this preliminary investigation, we tested the hypothesis that HPV infection may alter the metabolic activation of tobacco smoke carcinogens, such as benzo[a]pyrene (B[a]P), thereby playing a role in the etiology of cervical cancer. We examined in vitro the metabolism and DNA adduct formation of [3H]B[a]P in normal and HPV-16 immortalized human epithelial cervical cells in culture, and investigated the effect of [3H]B[a]P on growth of these cells. Cultures of normal human cervical cells and of HPV-16 immortalized cervical epithelial cells were exposed to 0.2 microM [3H]B[a]P for 24 and 48 h. [3H]B[a]P inhibited growth of both normal and HPV-16 immortalized cervical cells. However, the growth inhibition of normal cells was more profound than that of HPV-16 immortalized cells. Comparison of the metabolism of [3H]B[a]P in these cells indicated that they both metabolize [3H]B[a]P predominantly to [3H]trans-9,10-dihydroxy-9,10 dihydrobenzo[a]pyrene ([3H]B[a]P-9, 10-diol), [3H]r-7,t-8, 9,c-10-tetrahydroxy 7,8,9, 10-tetrahydrobenzo[a]pyrene ([3H]trans-anti-B[a]P-tetraol), and unknown polar products. Enzymatic hydrolysis of water-soluble metabolites indicated that the levels of glucuronide and sulfate conjugates in these cells are negligible. Similarly, both cell lines form similar [3H]B[a]P-DNA adducts. However, the level of the (+)[3H] anti-B[a]P diol epoxide (BPDE)-deoxyguanosine adduct in HPV-16 immortalized cells after 24 and 48 h exposures was 3.8 and 3. 1 pmol/mg DNA, respectively, which is 2.2-fold and 2.6-fold greater than the level of this adduct in normal cells. Under the conditions and within the time frame employed in these assays, both the cell growth and DNA damage induced by [3H]B[a]P appear to be higher in HPV-16 immortalized cells than those detected in normal cells. The results, although preliminary, suggest that HPV-16 immortalized cervical cells are more susceptible to DNA damage by BaP which, in part, may enhance their transformation to malignant cells. PMID- 10523715 TI - Advances in the molecular genetics of endometrial cancer (Review). AB - Recent studies have identified some of the genetic alterations involved in endometrial carcinoma development. Transforming genes, including K-ras and c erbB2/neu oncogenes and the p53, PTEN and hMLH1 tumor suppressor genes, are the most frequently altered. In addition, endometrial carcinomas express high levels of chemoresistance markers, including the MDR-1 or the MRP genes. The genetic background of an endometrial cancer patient may include high-penetrance genes such as the DNA mismatch repair genes causing microsatellite instability, and low penetrance genes such as those involved in estrogen-metabolism. The spectrum of several molecular lesions suggest a model for endometrial tumorigenesis through two divergent pathways, and which may improve the design of more rational therapeutic agents. PMID- 10523716 TI - Growth inhibition of human breast cancer cells by herbs and phytoestrogens. AB - Epidemiologic studies have suggested that consumption of phytoestrogen-rich foods may protect against breast cancer, and phytoestrogens such as genistein have been reported to both inhibit and stimulate the growth of some human breast cancer cells. The phytoestrogens genistein, daidzein, biochanin A, and coumestrol were tested and found to inhibit serum-stimulated growth in both T-47D and MCF-7 breast cancer cells at 10-100 microM. Extracts of several estrogenic herbs, including hops, black cohosh and vitex, inhibited growth of T-47D cells. These in vitro results suggest that certain herbs and phytoestrogens may have potential in the prevention of breast cancer. PMID- 10523717 TI - Toxicity of adjuvant high-dose interferon-alpha-2b in patients with cutaneous melanoma at high risk of recurrence. AB - Interferon-alpha-2b (INF-alpha-2b) has been approved by the FDA as adjuvant treatment for patients with melanoma at high risk of recurrence. INF-alpha-2b is administered at 20 MU/m2/day IV, 5 days per week for 4 weeks, and then 10 MU/m2/day SC, three times weekly for 48 weeks. We investigated the toxicity of this protocol in 30 patients between June 1996 and February 1998. An intensive toxicity evaluation program was developed to monitor side effects. During both induction and maintenance phases, 60% of patients required a dose delay and/or reduction. Twenty percent were unable to complete the treatment plan, and 53% tolerated at least 80% of the scheduled dose. The frequently reported toxicity during induction included constitutional symptoms, myelosuppression, and hepatotoxicity. All were reversible on cessation of treatment or dose modification. During maintenance, toxicity included thyroid dysfunction, hypertriglyceridemia, retinopathy and a combination of mood disturbances, memory loss, cognitive slowing and impaired executive function. Administration of high dose INF-alpha-2b is feasible, with close patient monitoring. PMID- 10523718 TI - Early, late symptoms and histological type of nasopharyngeal carcinoma. AB - After a wide literature review, we retrospectively analyzed the accurately recorded early onset and late symptoms at first diagnosis of nasopharyngeal carcinoma (NPC) in a series of 85 patients. Thirty-seven (44%) and 48 (56%) of cases had a squamous (SCC) and undifferentiated (UCNT) histological NPC subtype, respectively. Thirteen (15%), 21 (25%) and 51 (60%) of cases were T2, T3 and T4, respectively. The involvement of locoregional lymph nodes resulted significantly more frequent in UCNT with respect to SCC (35% versus 18% respectively; p<0.05). Overall, the early onset symptoms were represented by locoregional lymph node enlargement in 35% of cases; nasal symptoms in 32%, otological symptoms in 36%, while neurological symptoms were reported in only 2% of cases. On the contrary, symptoms at first diagnosis were more frequently represented by lymph node enlargement (53%) and nasal symptoms (68%) which were the symptoms effectively conducting the patients to the specialist. A protocol for management of ORL symptomatic patients according to the incidence of described early onset and at first diagnosis symptoms is suggested. PMID- 10523719 TI - TAG-72 expression and clinical outcome in primary breast cancer. AB - Clinical data of 92 patients with primary breast carcinomas previously analysed for the pattern of immunohistochemical expression of three distinct carbohydrate epitopes of the TAG-72 molecule were reviewed. The clinical outcome of the patients after a median follow-up of 66 months was determined in 84 out of 92 patients. Clinicopathological characteristics of the tumours and clinical outcome of the patients were correlated with the TAG-72 epitope expression. TAG-72 was expressed more frequently in patients aged more than 50 years and in tumours of larger size, with lymph nodes metastasis, with low differentiation and with high proliferative activity. A statistical correlation was found with more advanced stages of the disease (35.7% vs 60% in stage I and in stage II-III, respectively, p=0.03). Disease-free survival and overall survival were estimated by the Kaplan Meier method. The survival of the patients with tumours expressing TAG-72 was not statistically different from that of patients with tumours without TAG-72 expression. These data suggest that TAG-72 expression is associated with clinicopathological parameters of aggressiveness in primary breast cancer, but it does not appear to affect the clinical outcome of the patients. PMID- 10523720 TI - Overexpression of CXC-chemokines and CXC-chemokine receptor type II constitute an autocrine growth mechanism in the epidermoid carcinoma cells KB and A431. AB - The CXC-chemokines Groalpha and interleukin-8 (IL-8) are well characterized growth factors for melanoma cells. Here the constitutive expression of Groalpha, IL-8 and their receptors (CXCR1 and CXCR2) as well as their functional involvement in the proliferation response were analyzed in normal keratinocytes and epidermoid carcinoma cell lines A431 and KB. Flow cytometric measurements, ELISA and semi-quantitative RT-PCR revealed low constitutive protein secretion and mRNA expression of both CXC-chemokines as well as CXCR1 and 2 in normal keratinocytes, whereas significant higher levels of CXC-chemokines and CXCR2 were deteced in epidermoid carcinoma cells. Proliferation of epidermoid carcinoma cells could be induced by CXC-chemokines and constitutive proliferation could be inhibited by neutralizing antibodies against CXC-chemokines and CXCR2. These studies indicate that constitutive Groalpha, IL-8 and CXCR2 protein expression enable an autocrine growth mechanism in epidermoid carcinoma cells. PMID- 10523721 TI - Relationship among red blood cell polyamine content, phagocytic activities and plasma endotoxins in untreated colorectal cancer patients. AB - Polyamines are actively involved in immune processes and it is known that patients with cancer often exhibit immune deficits. Twenty-two patients with colorectal cancer were enrolled into this study, before starting conventional treatments. The relationship among the content of polyamines in red blood cells and phagocytosis and killing of monocytes and polymorphonuclear cells, and endotoxemia was investigated. The data show a negative correlation among levels of total polyamines and spermine and monocyte phagocytosis. Higher levels of spermine were present in patients with detectable circulating endotoxins. Our findings suggest a down-modulating effect of polyamines on the monocyte phagocytosis in untreated colorectal cancer patients; this effect could explain the presence of circulating endotoxins in cancer bearing patients. PMID- 10523722 TI - Cell cycle variations in azoxymethane-induced rat colorectal carcinogenesis studied by flow cytometry. AB - Cell cycle variations and DNA aneuploidy, were investigated in different phases of azoxymethane (AOM)-induced colon carcinogenesis in rats by flow cytometry. K ras gene mutations (transitions Gright curved arrow A) were frequently detected in aberrant crypt foci (ACF) initial pre-neoplastic lesions. The fraction of cells in the G2M-phase of the cell cycle was higher in ACF compared to the normal mucosa of control rats. A similar modification of the cell cycle was found in adenomas and adenocarcinomas but, unexpectedly, also in morphologically normal mucosa from AOM-treated animals indicating that AOM treatment permanently modifies cell cycle control in rat colon mucosa. These alterations, however, were not associated with DNA aneuploidy as reported in human sporadic colorectal cancer, suggesting that tumour development in AOM-treated rats is less dependent on aneuploidy. PMID- 10523723 TI - Retrospective evaluation of toxicity in three different schedules of adjuvant chemotherapy for patients with resected colorectal cancer. TTD Spanish Cooperative Group. AB - Adjuvant chemotherapy has been established since 1990 as standard treatment for patients with colon cancer stage III (Dukes' C). Chemotherapeutic schemes combining 5-fluorouracil with levamisole or leucovorin have shown significant advantage over surgery alone. Adjuvant trials are being currently implemented to investigate some relevant questions, such as which is the optimal duration of chemotherapy, as well as the possible advantage of levamisol versus leucovorin schedules, and of high-dose versus low-dose leucovorin. While these trials are ongoing, a retrospective evaluation of the toxicity associated with the different chemotherapeutic schemes might be of help when choosing the most appropriate regimen for individual patients not involved in clinical trials. A total of 519 patients subjected to three different schedules of adjuvant chemotherapy between 1993 and 1996, were evaluated for toxicity according to the NCI-CTC criteria. Chemotherapeutic regimens were: 5-fluorouracil plus levamisole (5-Fu+Lev; Moertel schedule), 5-fluorouracil plus low-dose leucovorin (5-Fu+LVLD; NCCTG schedule) and 5-fluorouracil plus high-dose leucovorin (5-Fu+LVHD; IMPACT-modified schedule). 5-Fu+LVLD is significantly more toxic than the other two regimens in terms of neutropenia, mucositis and diarrhea. delay in chemotherapy and dose reduction of 5-fluorouracil were also more frequent in the 5-Fu+LVLD group. However, the percentage of prematurely discontinued treatments was significantly higher in the 5-Fu+Lev group. Information on toxicity of adjuvant chemotherapy for colon cancer may help medical oncologists to choose the most appropriate regimen for individual patients not involved in clinical trials. PMID- 10523724 TI - Phase II study of induction chemotherapy followed by concomitant chemoradiotherapy in advanced head and neck cancer: clinical response and organ/function preservation. AB - We planned to conduct a trial of induction chemotherapy followed by concomitant chemoradiotherapy with the goal of organ-function preservation in advanced head and neck cancer patients with the response rate and local control of disease as primary endpoints and the assessment of toxicity as secondary endpoint. The overall treatment plan consisted of 3 cycles, each q. 28 days, of induction chemotherapy with cisplatin, 5-FU, leucovorin and interferon alpha2b (PFL-IFN), followed by response evaluation and local therapy with concomitant chemoradiotherapy with 5-FU, hydroxyurea and concomitant radiotherapy (FHX). The evaluation of clinical response was performed during the 2nd week after the 3rd cycle of induction chemotherapy and FHX was initiated 28 days after the 3rd cycle of induction chemotherapy. Hydroxyurea was administered orally at doses of 1 g every 12 h x 11, 5-FU was administered on days 1 through 5 at 800 mg/m2/d for 5 days. Daily fraction of radiotherapy were administered at 2.0 Gy on days 1 through 5. FHX cycles were repeated every 14 days until completion of radiotherapy. Total radiotherapy doses consisted of 70 Gy. Seventeen patients (mean age 56.53 years, range 40-73, male/female 15/2, site: oral cavity 6, 35.29%; oropharynx 3, 17.6%; hypopharynx 3, 17.65%; larynx 2, 11.76%; paranasal sinuses 2, 11.76%; salivary glands 1, 5.88%; ECOG PS 0/1: 10/7, stage: III/IV 3/14) were enrolled from January 1998 to August 1998. All 17 patients initiated induction chemotherapy on this protocol. Twelve patients were analyzed for response (5 patients were not evaluable): 2/12 (16.7%) patients achieved a CR and 10/12 (83.3%) achieved a PR for an ORR of 100%. Concomitant chemoradiotherapy was administered on protocol to 10 patients: 4 patients (40%) had CR, 3 patients (30%) had PR >/=70% for an ORR of 70%, 1 patient (10%) had SD and 2 patients (20%) had PD. As for local therapy, according to treatment plan, of the 8 eligible patients who completed chemoradiotherapy, the 4 patients with CR were submitted to random biopsies, which resulted histologically negative, the 3 patients with PR >/=70% underwent conservative organ-preserving surgery, the patient with SD underwent salvage surgery, preserving voice. Thus, organ preservation was achieved in all 8 patients at the completion of all therapy: 4 patients had no surgical procedure and 3 patients only conservative surgery. Overall, after completion of all therapy, 6/8 (75%) patients were rendered disease-free. Both induction chemotherapy and concomitant chemoradiotherapy resulted in significant toxicity, which consisted mainly of mucositis and thrombocytopenia. In conclusion, in the present study we have achieved a good clinical response and an optimal organ preservation, at the cost of a severe toxicity. PMID- 10523725 TI - Increased hyaluronidase expression in more aggressive prostate adenocarcinoma. AB - Tumor metastasis involves a complex sequence of cellular and biochemical events of which tumor cell penetration through the basement membrane is an essential component. Disruption of basement membrane integrity by hyaluronidase has been implicated in the development of locally advanced and metastatic carcinoma. This investigation correlates hyaluronidase expression with pathologic prognostic variables for prostate adenocarcinoma. Paraffin samples (n=9) of patients receiving prostatectomies for clinical stage B adenocarcinoma were evaluated. RT PCR was utilized to detect the presence of hyaluronidase. These results were correlated with the histological assessment of each sample. Gleason score was significantly higher in patients with RT-PCR detectable hyaluronidase (mean +/- SD 7.25+/-1.8 versus 4.17+/-1.0; p<0.05). Histological evidence of perineural invasion was seen in 83% of the specimens with detectable hyaluronidase, whereas none was found in hyaluronidase negative samples (p<0.05). A trend was seen between hyaluronidase and capsular invasion with 33% hyaluronidase positive tumor exhibiting evidence of capsular invasion, while no evidence in the hyaluronidase negative tumor. These data suggest that hyaluronidase may be involved in prostate adenocarcinoma progression and metastasis. PMID- 10523727 TI - Ferrochelatase, a novel target for photodynamic therapy of cancer. AB - This study was designed to investigate the hypothesis that the inhibition of ferrochelatase will cause in situ build up of high concentrations of protoporphyrin-IX which may act as a putative agent for photodestruction of cancer cells. The parenteral administration of lead acetate, a known inhibitor of ferrochelatase, to mice bearing cutaneous tumors (papillomas and carcinomas) caused a six-fold enhancement in the concentration of protoporphyrin-IX in tumors within a period of one month. Forty-eight hours after the second injection of lead, mice were exposed to visible light, at a light dose of about nine kilo lux for a period of one hour (in four sittings of fifteen minutes each keeping a gap of ten minutes between two exposures). A significant reduction in tumor size was observed starting as early as day one following the treatment. Continuous treatment for six consecutive days resulted in almost complete ablation of the tumor mass in most of the animals. Complete regression of the tumors was observed at two to three days following the first exposure. Our observations on in situ accumulation of protoporphyrin-IX by heme-biosynthesis inhibition represent a novel method for photodynamic therapy of cancer cells. It is important to emphasize that lead is a fairly toxic agent and developing a non-toxic agent is one of our future goals. PMID- 10523726 TI - Germline mutation analysis of BRCA1 and BRCA2 genes in Yugoslav breast/ovarian cancer families. AB - The frequency of germline BRCA1 and BRCA2 mutations was tested in Yugoslav breast and breast/ovarian cancer families using combined heteroduplex/single-strand conformation polymorphism analysis for the entire coding region of both genes. Three different recurrent BRCA1 mutations (one 185delAG, one 3447del4 and two 5382insC) were identified in 4 of 12 families (33%), whereas no definite disease causing alterations of BRCA2 was detected. Genotype analysis revealed a possible common founder effect for each 185delAG and 5382insC. The relatively high frequency of germline BRCA1 mutations determined in this panel of families confirms the important role of BRCA1 in disease predisposition in the Yugoslav population, while the lack of population specific founder and/or unique mutations show the need of further analysis of samples from this yet unexamined region of Europe. PMID- 10523728 TI - Recent physiopathological insights in cutaneous lymphocytic infiltrates. AB - The spectrum of benign cutaneous lymphocytic infiltrates (CLI) includes a variety of entities which are classified on the basis of clinicopathological findings, and of the phenotype of the predominant lymphocytic subset among the skin infiltrate. Major concern has been recelntly given to the clonality status of CLI by using highly sensitive assays based on the PCR amplification of TCR/Ig loci. These studies allowed the characterisation of clonal benighn cutaneous lymphocytic expansions. Other studies have shown evidence of oligoclonal patterns in HIV-associated CD8 cutaneous pseudolymphomas, and functionnal studies of the skin infiltrate further showed that an antigen-driven mechanism was involved in the pathogenesis of this latter entity. Finally, the knowledge in the field of CLI has been improved by the identification of antigens associated with skin homing properties such as the so-called. Cutaneous Lymphocyte-associated Antigen (CLA) whicis expressed at the surface of most memory T cells infiltrating the dermis in inflammatory conditions. PMID- 10523729 TI - A case of dyskeratosis congenita associated with impaired DNA repair as shown by the comet assay. AB - An 18-year old boy with dyskeratosis congenita is presented. To examine the DNA metabolism of our patient, we applied the comet assay, a simple, quick and sensitive method that so far has not been used in this disease. After exposure to UVB, cells originating from the patient present abnormal DNA repair localized in the late step. We consider that such repair deficiency could be related to susceptibility to cancer. The comet assay seems to be a good procedure to investigate dyskeratosis congenita or other genodermatoses. PMID- 10523730 TI - Costello syndrome with decreased glucose tolerance. AB - We report a case of Costello syndrome, which is an uncommon multisystemic condition with cutaneous manifestations on the palms and soles. In the literature there are 29 cases described, all the studies are published in the genetic literature with a few exceptions. We add a further case associated with impaired glucose tolerance. The diagnostic clinical signs are impressive, and highly characteristic. Cutaneous manifestations are: loose skin of the hands and feet "washer woman's hand", hyperkeratosis palmoplantaris, curly or sparse hair, acanthosis nigricans, papillomata nasi. Coarse, progeroid facial features with a bulbous nose, feeding difficulties in infancy, cardiac involvement with cardiomyopathy or conduction defect, and in our case impaired glucose tolerance also presented. Postnatal growth retardation, mental retardation, and a distinctive friendly personality is characteristic. Hyperextensible fingers with broad distal phalanges and joint contractures were observed, and peroneal hypertonicity required treatment by Achilles tendon lengthening. The decreased glucose tolerance is interesting in the view of the acanthosis nigricans. No storage disease and no chromosomal abnormality were observed. Only in one case is a balanced translocation described in the literature. PMID- 10523731 TI - Psoriatic erythroderma and bullous pemphigoid treated successfully with acitretin and azathioprine. AB - We present the case of a 59-year-old male patient who developed lesions of a bullous pemphigoid during the course of a long-lasting severe psoriasis which had been treated for years with different topical treatments as well as with PUVA and UV-B radiations. Our patient was successfully treated with a combination of acitretin and azathioprine (follow up 28 months). Our case shows that it is possible to avoid systemic corticosteroid treatment in this difficult therapeutic situation. PMID- 10523732 TI - Continuous itraconazole treatment for onychomycosis and dermatomycosis: an overview of safety. AB - Over the past 10 years, itraconazole has been used to treat more than 34 million patients worldwide. We present a review of the safety of various continuous itraconazole schedules used in the treatment of dermatomycosis and onychomycosis. Data from controlled clinical trials and extensive post-marketing surveillance show that itraconazole has an impressive safety profile at a dose of 50-200 mg/day for 1-4 weeks for dermatomycosis and 200 mg/day for 3 months for onychomycosis. In addition, itraconazole is safe to use in diabetic patients with dermatomycosis or onychomycosis. Short-term, intermittent itraconazole regimens, which may offer additional benefits in terms of safety and cost, have now been introduced. PMID- 10523733 TI - Acquired partial curly hair. AB - We report 6 adolescent females apparently affected by diffuse partial woolly hair. The patients complained of thinning and curling of some hair of the scalp. Examination revealed two distinct hair populations. The first hair population was straight, normally pigmented and of normal length, thus considered "normal" hair. The second hair population was wavy, curly, thinner and apparently slightly shorter, thus considered abnormal. Direct examination of the abnormal hair population under a light microscope revealed a curled pattern of the hair shaft, and single torsions. With light microscopy the clinically normal hair had no abnormalities. Under scanning electron microscopy (SEM) the abnormal hair had curly and flattened hair shafts of 30-60 mum in diameter, oval shaped sections, canalicular formations, single torsions and cuticular weathering. With SEM, clinically "normal" hair had a diameter between 60-80 mum, with no major relevant abnormalities except for weathering. These results are suggestive of a new variant of acquired kinking (curling) of hair simulating diffuse partial woolly hair. The absence of clinical alterations in the proximal area of the affected hair and the spontaneous improvement with time, suggests that this variant may result from environmental exposure (weathering, cosmetic procedures) in predisposed individuals. PMID- 10523734 TI - Generalized eruptive histiocytoma of childhood associated with rheumatic fever. AB - We describe a widespread papular eruption in a 5-year-old girl with rheumatic fever. Histological examination revealed a dense histiocytic infiltration in the dermis. On immunohistochemical studies, the cells were positive for vimentin, CD68, MAC387, alpha1-antichymotrypsin and lysozyme, but negative for CD1a and S 100 protein. Electron microscopic studies showed no Birbeck granules in their cytoplasm. A diagnosis of generalized eruptive histiocytoma of childhood was established. The skin lesions completely disappeared within 8 months. PMID- 10523735 TI - Scleromyxedema is a scleroderma-like disorder and not a coexistance of scleroderma with papular mucinosis. AB - We present four cases of scleromyxedema with scleroderma-like cutaneous changes mimicking systemic sclerosis and stress the importance of their differentiation from true scleroderma. Scleromyxedema should be recognized as an entity since it differs from scleroderma in the pathogenesis, histopathology of cutaneous lesions, type of visceral involvement (if present), frequent association with paraproteinemia, the course and prognosis. PMID- 10523736 TI - Aggressive verbal behaviour as a function of experimentally induced anger in persons with psoriasis. AB - The importance of psychosocial factors on the etiology and fluctuating disease activity of psoriasis has been discussed in recent years. The present experiment investigated whether psoriatics in an anger-inducing situation show less aggressive verbal behaviour than average person. Twenty-six psoriatics and 26 matched healthy controls were randomly assigned to either an anger-inducing or a non-anger-inducing social situation. The experimental conditions were arranged so that the persons were confronted with either negative, derogatory, or positive, favorable feedback on eight characteristics (intelligence, appearance, maturity, tolerance, honesty, friendliness, humor, and helpfulness). Standardized feedback was given by a confederate of the experimenter. Immediately after the feedback was received by the subjects the photo hand test (PHT) was applied. The PHT is an item-analyzed, validated projective test for aggression. Two independent raters categorized the subjects' responses into six mutually exclusive categories, including a category for responses with aggressive content. 2 x 2 analysis of variance (psoriatics vs controls; anger-induced vs non-anger induced) were calculated for the aggressive responses and the acting-out score (AOS). The results showed a significant interaction, suggesting that psoriatics did indeed exhibit fewer verbal aggression responses under anger-inducing circumstances than the controls. PMID- 10523737 TI - In vitro released interferon-gamma in the diagnosis of drug-induced anaphylaxis. AB - A 17-year-old Japanese male was referred with acute urticaria and anaphylaxis after the administration of PL (salicylamide, acetaminophen, anhydrous caffeine and promethazine methylene disalicylate) and Bufferin (aspirin and dialminate) for headache and a high grade fever. The results of prick test, patch test and drug-induced lymphocyte stimulation test with PL and Bufferin were all negative. The patient's peripheral blood mononuclear cells (PBMC) were cultured with or without PL for 72 hours, and the activity of interferon-gamma (IFN-gamma) in the culture supernatant was measured with EIA. A significantly high level of IFN gamma was detected in PBMC from the patient, but very little in those from healthy control subjects with a history of exposure to PL. This finding may indicate the presence of drug-specific IFN-gamma producing T cells in patients with an anaphylactic shock reaction to medication. Assays that measure the drug induced IFN-gamma production may thus be a useful diagnostic tool not only for identifying delayed-type hypersensitivity (DTH) to drugs, but also for predicting anaphylactic shock reaction to drugs. PMID- 10523738 TI - Cutis laxa acquisita: is there any association with Borrelia burgdorferi? AB - We report the first case of an acquired form of generalized cutis laxa which has positive serology and a positive polymerase chain reaction (PCR) result for lyme borreliosis. A 44-year-old man complained of excessively loose skin for four years and had no family history of any skin disease. Dermatological examination showed lax and wrinkled skin all over the body (especially on the cheeks and the intertriginous areas). Positive serology for lyme borreliosis and the presence of Borrelia burgdorferi DNA which was demonstrated by nested PCR in this acquired form of cutis laxa is interesting since it has not been reported in literature previously. PMID- 10523739 TI - Panniculitis revealing qualitative alpha 1 antitrypsine deficiency (MS variant). AB - A 16-year-old girl presented painful, red, nodular lesions on the abdomen. A cutaneous biopsy showed inflammatory cell infiltrate and fibrosis in the dermis and in the septa with isolated adipocyte lobules. alpha1-antitrypsin level was found to be normal but M1S phenotype of alpha1-antitrypsin was determined by isoelectric focusing in polyacrylamide gel. alpha1-antitrypsin level was normal for her family but M2S phenotype was found in her father. Alpha 1-antitrypsin (alpha1 AT) deficiency is a common hereditary disorder of Caucasians. The locus is pleiomorphic and 75 alleles have been identified. Numerous pathological mutations can be classified by the mechanisms which cause the deficiency. The major clinical importance of this deficiency is emphysema and liver disease. Panniculitis is rarely reported and seems to occur principally for the ZZ or MZ phenotype and for low levels of alpha1 AT. MS phenotype has been more rarely reported and triggering agents such as trauma and infections must be present. However, normal levels of alpha1 AT in the serum have previously been reported as in our case, and we suggest the study of alpha1 AT phenotype even if the plasma level is normal. PMID- 10523740 TI - Metastasising malignant lymphoma mimicking necrotising and hyperplastic gingivostomatitis. AB - This paper presents the case of a 65 year-old woman suffering from recurrent oral aphthoid ulcers which rapidly evolved towards hyperplastic and ulcerated lesions over the entire floor of the mouth. The initial lesions were interpreted as non specific aphthoid ulcers. Later, a tentative diagnosis of necrotising stomatitis with secondary reactive proliferating epithelial hyperplasia was made. The clinical symptoms and the immuno-phenotyping of lymphocytes circulating in the peripheral blood suggested the diagnosis of CD30-positive large cell anaplastic lymphoma. The biopsy showed only a pseudoepitheliomatous hyperplasia, reactive infiltrates and no lymphoma cells. The disease ran a fulminant course leading to death within 4 weeks due to acute gastro-intestinal bleeding. Autopsy revealed infiltrates of CD30+ large cell anaplastic lymphoma in a submandibular lymph node, in a thrombus stenosing the right subclavian vein, in the spleen, the anterior and posterior gastric wall as well as in the depth of the tumour on the floor of the mouth. The clinical and histopathological spectrum of CD30+ large cell anaplastic lymphoma is considerably variable. The particular feature of pseudoepitheliomatous hyperplasia has been reported especially in CD30+ anaplastic large cell lymphomas. An early correct diagnosis is rendered difficult in insufficient biopsy size, becauses this type of lymphoma often simulates other inflammatory or neoplastic skin diseases. Thus, with a necrotising and hyperplastic gingivostomatitis, the diagnosis of a CD30+ anaplastic large cell lymphoma should be considered. PMID- 10523741 TI - Generalized pustular psoriasis with hypoparathyroidism. AB - A 36-year-old Japanese woman with pustular psoriasis associated with hypoparathyroidisum was reported. The patient showed hypocalcemia and was treated with calcium supplements and calcitriol. When the serum calcium level became normal, the pustules disappeared and erythroderma completely resolved. Histopathological features consisted of the formation of intraepidermal pustules including spongiform pustules underneath the stratum corneum, and acantholysis was observed in the epidermis. It was suggested that generalized pustular psoriasis may have been induced by hypocalcemia due to hypoparathyroidism in this case, and that acantholysis may be caused by hypocalcemia, since intercellular junctional components such as cadherins are highly dependent on calcium in the epidermis. PMID- 10523742 TI - Disseminated Trichosporonosis with Trichosporon asahii. AB - Trichosporon asahii fungemia was associated with multiple, purpuric, papular lesions on the abdomen and extremities in a 53-year-old man with acute myeloblastic leukemia. Histologically, budding yeasts were demonstrated in the dermis. The yeast-form fungus was identified as T. asahii. PMID- 10523744 TI - Measurement error in epidemiology: the design of validation studies I: univariate situation. AB - It is becoming standard practice in epidemiology to adjust relative risk estimates to remove the bias caused by non-differential errors in the exposure measurement. Estimation of the correction factor is often based on a validation study incorporating repeated measures of exposure, which are assumed to be independent. This assumption is difficult to verify and often likely to be false. We examine the effect of departures from this assumption on the correction factor estimate, and explore the design of validation studies using two or even three different types of measurement of exposure, where assumption of independence between the measures may be more realistic. The value of good biomarker measures of exposure is demonstrated even if they are feasible to use only in a validation study. PMID- 10523743 TI - Superantigens in skin diseases. AB - Superantigens are strong modulators of the immune system affecting T cells, antigen-presenting cells and other MHC class II-positive cells directly and many other cells indirectly, e.g. mast cells that have bound IgE specific for superantigens. The strong immunomodulatory effects, which are outlined in this article besides the biological properties of superantigens, strongly influence immunologically-mediated disorders including inflammatory skin diseases like atopic dermatitis and psoriasis, which are the focus of this article. PMID- 10523745 TI - Measurement error in epidemiology: the design of validation studies II: bivariate situation. AB - The bias in relative risk estimates caused by errors in measurement of the relevant exposure is being increasingly recognized in epidemiology. Estimation of the necessary correction factor to remove this bias for univariate exposure has been considered in an earlier paper. We consider here the multivariate situation in which non-differential errors in measurement can lead to incorrect identification of the variable most closely associated with disease. Estimation of the necessary correction factor when the true exposure is unobservable necessarily requires assumptions. We explore the robustness of the estimation to departures from a range of assumptions. The value of good biomarkers is demonstrated. We present a bivariate example in which failure to take account of measurement error leads to the incorrect exposure being identified as the important determinant of disease risk. PMID- 10523746 TI - Regression with covariates and outcome calculated from a common set of variables measured with error: estimation using the SIMEX method. AB - In medical research, a situation commonly arises where new variables are calculated from a common set of directly measured variables. When the directly measured variables each contain an error component, the relationship between the observed calculated variables can often be distorted. A source of this distortion is the presence of common measurement error in the observed calculated variables. Often known as coupled error, it is still possible to estimate the relationship between the calculated variables when measurement error is present. This paper presents two general methodologies that account for the presence of correlated measurement error when working with calculated variables. The equivalence of the methods will be established for one case, while the general advantage of the simulation extrapolation technique will be shown for more complicated situations. The performance of the estimators will be examined with examples arising from the medical literature. PMID- 10523747 TI - The problem of measurement error in modelling the effect of compliance in a randomized trial. AB - This paper explores the implications of measurement error in the analysis of compliance-response relationships in data from randomized trials. Given that compliance measures are rarely, if ever, error-free indicators of exposure it is argued that both the designs for the collection of compliance data and the statistical models for their resulting analysis should be changed to take the possibility of measurement error into account. An analysis which ignores measurement error in the compliance measurements will provide biased estimates of compliance-response relationships. Provided that one has two or more indicators of compliance for each subject, more appropriate models can be fitted using covariance structure modelling software. If one wishes to explore interactions from repeated measures data on both compliance and response then it is also important that one recognizes that the response measures are also error-prone and that they too are dealt with appropriately. PMID- 10523748 TI - Correcting for non-compliance in randomized trials: an application to the ATBC Study. AB - Different methods for estimating the effect of treatment actually received in a longitudinal placebo-controlled trial with non-compliance are discussed. Total mortality from the ATBC Study is used as an illustrative example. In the ATBC Study some 25 per cent of the participants dropped out from active follow-up prior to the scheduled end of the study. The 'intention-to-treat' analysis showed an increased death risk in the beta-carotene arm when compared with the no beta carotene arm. Owing to considerable non-compliance it is also of interest to estimate the effect of beta-carotene actually received. We use a simple model for the treatment action and discuss three methods for estimation of the treatment effect under the model - the 'intention-to-treat' approach, the 'as-treated' approach and the g-estimation approach. These approaches are compared in a simulation study under different settings for non-compliance. Finally, the data from the ATBC Study are analysed using the proposed methods. PMID- 10523750 TI - Item response models for longitudinal quality of life data in clinical trials. AB - Assessment of quality of life is becoming standard in clinical trials. A popular method for measuring quality of life is with instruments which utilize multiple item subscales, in which each item is scored on a Likert scale. Most statistical methods for the analysis of quality of life data in clinical trials do not explicity consider the properties and psychometric features which were of interest in scale development. In this regard, the measurement and statistical summarization of quality of life data, along with the clinical interpretation, can be somewhat disjoint from the psychometric concerns of the development process. The aim of this paper is to address the complicated issues present in analysing multiple-item ordinal quality of life data in clinical trials while maintaining fidelity to the psychometrical foundations upon which quality of life instruments are built. Accomplishing this will require the development of item response models which recognize the longitudinal aspects of clinical trial designs as well as the potential problem of informatively missing data. A general item response modeling approach is presented for longitudinal multiple-item quality of life data measured on ordinal scales with model components for missing data mechanisms and latent trait regression on treatment indicators and other covariates. PMID- 10523749 TI - Modelling disease progression in terms of exposure history. AB - We consider the relationship between accumulating exposure to a putative agent and the associated change in physiologic function. This type of problem is common to prospective studies of cognitive, pulmonary and cardiovascular function. A general model is proposed for data from prospective, observational studies with concurrent measures of exposures and continuous outcome measures. This model permits non-linearity in the relationship between exposure and outcome and is designed to describe outcome in terms of one's entire exposure history. As exposure data are often severely right-skewed, we use regression spline estimation methods which localize the influence of extreme points. We illustrate our methodology using data from a longitudinal epidemiologic investigation of the effects of amateur boxing on neuropsychologic function. PMID- 10523751 TI - Simple pattern-mixture models for longitudinal data with missing observations: analysis of urinary incontinence data. AB - In longitudinal studies each subject is observed at several different times. Longitudinal studies are rarely balanced and complete due to occurrence of missing data. Little proposed pattern-mixture models for the analysis of incomplete multivariate normal data. Later, Little proposed an approach to modelling the drop-out mechanism based on the pattern-mixture models. We advocate the pattern-mixture models for analysing the longitudinal data with binary or Poisson responses in which the generalized estimating equations formulation of Liang and Zeger is sensible. The proposed method is illustrated with a real data set. PMID- 10523752 TI - Why not routinely use best linear unbiased predictors (BLUPs) as estimates of cholesterol, per cent fat from kcal and physical activity? AB - Measures of biologic and behavioural variables on a patient often estimate longer term latent values, with the two connected by a simple response error model. For example, a subject's measured total cholesterol is an estimate (equal to the best linear unbiased estimate (BLUE)) of a subject's latent total cholesterol. With known (or estimated) variances, an alternative estimate is the best linear unbiased predictor (BLUP). We illustrate and discuss when the BLUE or BLUP will be a better estimate of a subject's latent value given a single measure on a subject, concluding that the BLUP estimator should be routinely used for total cholesterol and per cent kcal from fat, with a modified BLUP estimator used for large observed values of leisure time activity. Data from a large longitudinal study of seasonal variation in serum cholesterol forms the backdrop for the illustrations. Simulations which mimic the empirical and response error distributions are used to guide choice of an estimator. We use the simulations to describe criteria for estimator choice, to identify parameter ranges where BLUE or BLUP estimates are superior, and discuss key ideas that underlie the results. PMID- 10523753 TI - Genetic mapping of complex traits. AB - Statistical genetic mapping methods are powerful tools for finding genes that contribute to complex human traits. Mapping methods combine knowledge of the biological mechanisms of inheritance and the randomness inherent in those mechanisms to locate, with increasing precision, trait genes on the human genome. We provide an overview of the two major classes of mapping methods, genetic linkage analysis and linkage disequilibrium analysis, and related concepts of genetic inheritance. PMID- 10523754 TI - Both glutamate receptor antagonists and prefrontal cortex lesions prevent induction of cocaine sensitization and associated neuroadaptations. AB - Behavioral sensitization to psychomotor stimulants is accompanied by a number of alterations in the mesoaccumbens dopamine (DA) system, including DA autoreceptor subsensitivity in the ventral tegmental area (VTA) and DA D1 receptor supersensitivity in the nucleus accumbens (NAc). We investigated the role of excitatory amino acid (EAA) transmission in the induction of cocaine sensitization and these accompanying DA receptor alterations. To do so, we used three glutamate receptor antagonists, the noncompetitive NMDA receptor antagonist MK-801 (0.1 mg/kg), the competitive NMDA receptor antagonist CGS 19755 (10.0 mg/kg), and the AMPA receptor antagonist NBQX (12.5 mg/kg). Rats received daily double injections of either one of these antagonists or saline with either cocaine (15.0 mg/kg) or saline for 5 days. Cocaine sensitization was defined as an increase in horizontal locomotor activity in response to cocaine challenge (7.5 mg/kg) on the third day of withdrawal. All three antagonists prevented the induction of cocaine sensitization. Extracellular single cell recordings revealed that these antagonists also prevented the induction of DA autoreceptor subsensitivity in the VTA and DA D1 receptor supersensitivity in the NAc. To determine whether the relevant glutamate receptors were under regulation by medial prefrontal cortex (mPFC) EAA efferents, we next lesioned the mPFC bilaterally with ibotenic acid at least 7 days before repeated cocaine treatment began. These lesions also prevented the induction of cocaine sensitization and the associated neuroadaptations. Our findings indicate that glutamate transmission from mPFC to the mesoaccumbens DA system is critical for the induction of cocaine sensitization and its cellular correlates. PMID- 10523755 TI - Identification of a high-affinity orphanin FQ/nociceptin(1-11) binding site in mouse brain. AB - The presence of pairs of basic amino acids within the orphanin FQ/Nociceptin (OFQ/N) sequence has raised the possibility that truncated versions of the peptide might be physiologically important. OFQ/N(1-11) is pharmacologically active in mice, despite its poor affinity in binding assays (K(i) > 250 nM) for the OFQ/N receptor. Using an analog of OFQ/N(1-11), [(125)I][Tyr(10)]OFQ/N(1-11), we identified a high-affinity binding site (K(D) 234 pM; B(max) 43 fmol/mg protein) with a selectivity profile distinct from the OFQ/N receptor and all the traditional opioid receptors. This site had very high affinity for OFQ/N and its related peptides. The most striking differences between the new site and the OFQ/N receptor previously observed in brain were seen with traditional opioids. Dynorphin A analogs and alpha-neoendorphin competed with [(125)I][Tyr(10)]OFQ/N(1 11) binding in mouse brain with K(i) values below 10 nM, while naloxone benzoylhydrazone (K(i) 3.9 nM) labeled the [(125)I][Tyr(10)]OFQ/N(1-11) binding site as potently as many traditional opioid receptors. Several other opioids, including fentanyl, (-)cyclazocine, levallorphan, naltrindole, and diprenorphine, also displayed moderate affinities for this site. Finally, the [(125)I][Tyr(10)]OFQ/N(1-11) site had a unique regional distribution consistent with a distinct receptor. Thus, [(125)I][Tyr(10)]OFQ/N(1-11) labels a novel site in brain with a selectivity profile intermediate between that of either opioid or OFQ/N receptors. PMID- 10523756 TI - [(125)I]orphanin FQ/nociceptin binding in Raji cells. AB - Western blots using an antibody which recognizes the orphanin FQ/nociceptin (OFQ/N) receptor reveals a band at approximately 69 kD in several cell lines, including the Raji human B cell lymphoma cell line. RT-PCR confirms the presence of this receptor in the Raji cells. Binding studies revealed a high affinity [(125)I][Tyr(14)]OFQ/N site in the Raji cells. The affinity of [(125)I][Tyr(14)]OFQ/N in the Raji cells (K(D) 68.4 pM) was similar to that in the transfected receptor (K(D) 36.7 pM). Its selectivity profile also was quite similar. OFQ/N competed binding quite potently (K(i) 65 pM), as did [Tyr(14)]OFQ/N (K(i) 33 pM). Traditional opioids displayed no appreciable affinity for the binding at any concentration examined, with the exception of naloxone benzoylhydrazone, which had only a very modest affinity. The receptors in the Raji cells were functionally active. OFQ/N inhibited forskolin-stimulated cyclase by 72% with an IC(50) value of approximately 1 nM. PMID- 10523757 TI - Neurochemical changes in the entopeduncular nucleus and increased oral behavior in rats treated subchronically with clozapine or haloperidol. AB - The purpose of the present experiment was to test the possibility that atypical antipsychotics and classical antipsychotics differentially regulate specific neurochemical processes within the entopeduncular nucleus. For these experiments, rats were administered clozapine (25 mg/kg), haloperidol (1 mg/kg), or Tween-80 (control) daily for 21 days. Dopamine D(1)-receptor binding was assessed with in vitro receptor autoradiographic methods and the mRNAs corresponding to the two forms of glutamate decarboxylase (glutamate decarboxylase-65 and glutamate decarboxylase-67) were analyzed using in situ hybridization histochemical methods. In addition, vacuous chewing movements (VCM) were measured throughout the drug administration period as a functional indicator of drug action and changes in striatal dopamine D(2)-receptor binding were measured as a positive control for D(2)-receptor antagonist properties of haloperidol and clozapine. In agreement with previous reports, haloperidol increased D(2)-receptor binding throughout the striatum while clozapine had a more limited impact on D(2) receptors. Behavioral analysis revealed that both haloperidol and clozapine enhanced the display of vacuous chewing movements to a similar extent but with a different postinjection latency. In the entopeduncular nucleus, clozapine increased D(1)-receptor binding compared to controls while haloperidol was without effect. With respect to the regulation of GAD mRNAs, haloperidol increased glutamate decarboxylase-65 and glutamate decarboxylase-67 mRNA levels throughout the entopeduncular nucleus. The effects of clozapine were restricted to increases in glutamate decarboxylase-65 mRNA. These studies show that clozapine and haloperidol, both of which increase the occurrence of VCM, differentially modulate the neurochemistry of the entopeduncular nucleus. PMID- 10523758 TI - Ultrastructural and metabolic changes in the neuropeptide Y-containing striatal neuronal network after thermocoagulatory cortical lesion in adult rat. AB - This study examined the effects of unilateral thermocoagulatory cortical lesion on the pattern of neuropeptide Y immunostaining in the rat ipsilateral striatum at 4 and 21 days post-lesion. Light microscopic analysis showed a significant increase in the number of neuropeptide Y-positive neurons vs. control at both time points; paradoxically, the intraneuronal level of labelling significantly decreased at 4 days post-lesion but increased at 21 days post-lesion. Ultrastructural analysis in control condition showed a higher proportion of dendritic versus axonal labelled processes (3.5 ratio); all the neuropeptide Y synaptic terminals formed symmetrical contacts, mostly onto unlabelled dendrites. At 4 days post-lesion, the neuropeptide Y-positive axon density dramatically increased (+576%) without significant change in the labelled dendrite density, vs. control values; the density of neuropeptide Y synaptic terminals increased in parallel by 233%. In addition, a significant proportion of large neuropeptide Y boutons forming asymmetrical synapses onto unlabelled spines were observed. At 21 days post-lesion, densities of neuropeptide Y dendrites, axons, and synaptic terminals increased by 68, 246 and 125%, respectively, vs. control. But, the morphological features of the neuropeptide Y axonal processes and synaptic specializations of the boutons were similar to those observed in control condition. These data (1) raise an important issue regarding the origin of the terminals forming asymmetrical synapses in the striatum, (2) suggest that adaptative changes in the neuropeptide Y neuronal network may be a main component of striatal remodelling resulting from the progressive loss of cortical inputs, and (3) reinforce the view that neuropeptide Y and excitatory amino acid functions may be tightly linked in the striatum. PMID- 10523759 TI - Regulation of striatal dopamine receptors by estrogen. AB - The ability of estrogen to modulate the expression of ventral and dorsal striatal dopamine receptors D(1), D(2,) and D(3) was examined in vivo using semi quantitative in situ hybridization and ligand binding autoradiography. Two-week treatment with subcutaneous pellets of 17beta-estradiol (25 mg) downregulated D(2) dopamine receptor mRNA in both dorsal and ventral striatum (shell and core regions of nucleus accumbens). No significant changes in D(1) or D(3) mRNA expression were detected. Ligand binding autoradiography did not reveal changes in D(1), D(2,) or D(3) receptor protein expression. We also assessed the ability of 17beta-estradiol to regulate D(2) gene promoter activity in NB41A3 neuroblastoma cells that express this gene endogenously using co-transfections with an estrogen receptor expression vector. While a small fragment of the D(2) promoter could be activated 2.5-fold by estrogen, a larger portion of the D(2) gene was not regulated by this treatment. Estrogens do not appear to have a net effect on striatal dopamine receptor expression. The observed downregulation of D(2) receptor mRNA in the dorsal and ventral striatum in vivo could be secondary to the increased striatal dopamine release induced by estrogen. Synapse 34:222 227, 1999. Published 1999 Wiley-Liss, Inc. PMID- 10523760 TI - Subthalamic responses to amphetamine and apomorphine in the behaving rat with a unilateral 6-OHDA lesion in the substantia nigra. AB - The activity of neurons in the subthalamic nucleus (STN) of the behaving rat, before and after a unilateral 6-OHDA lesion of the substantia nigra, was recorded with the extracellular technique to determine whether it was altered following systemic amphetamine, 5 mg/kg, apomorphine, 3 mg/kg, and apomorphine, 0.3 mg/kg, and whether in cases of altered activity, it was related to the drug-induced motor response expressed concurrently. Activity in the STN of intact rats increased dramatically after amphetamine, 5 mg/kg. This excitatory response had the same latency, similar magnitude, and the same duration as the motor response expressed in terms of locomotion and oral stereotypy. Motor and unit responses were also induced by amphetamine after the lesion with 6-hydroxydopamine (6 OHDA), but now the excitatory response was attenuated while the motor response was not. The effects of the 6-OHDA lesion were the same in all animals with loss of the nigra dopamine neurons, regardless of whether they were rotators or non rotators. Activity in the STN of intact rats also increased after apomorphine, 3 mg/kg, and again, this increase was correlated with the increase in motor behavior, but both responses were of shorter duration than the responses to amphetamine. The increases in unit activity and motor behavior induced by apomorphine in the 6-OHDA-lesioned rats had the same magnitude but lasted longer than in the intact rats. Treatment with apomorphine, 0.3 mg/kg, of the intact rats produced small and very brief increases in the activity of the STN and in motor behavior. The same treatment given the 6-OHDA-lesioned rats produced responses of larger magnitude but no change in duration. These findings demonstrate a role for STN neurons in the mediation of the motor behaviors induced by stimulation of the dopamine receptor. The results also show that a unilateral lesion of the substantia nigra with 6-OHDA did not block these responses but altered them in a manner consistent with a dopaminergic deafferentation of the basal ganglia. The increased activity in the STN during the expression of dopamine-dependent motor behavior conflicts with the current model of basal ganglia function that assumes prejudicial effects of excessive STN activity on the expression of motor behavior. An explanation for this conflict suggests that it is more apparent than real. PMID- 10523761 TI - Regulation of tyrosine hyroxylase gene transcription in ventral midbrain by glial cell line-derived neurotrophic factor. PMID- 10523762 TI - Mass spectral studies of methoxynaphthoflavones AB - The electron impact induced fragmentations of seven methoxynaphthoflavones have been studied with the aid of low- and high-resolution measurements, metastable decompositions and isotope labelling using carbon-13 atoms. The retro Diels-Alder cleavage of the methoxynaphthoflavones is strongly influenced by the substituent position providing in most cases intact A- and B-ring fragments. The intensity ratio of these ring fragments appears to be very sensitive to the charge distribution within the parent ion. Copyright -Copyright 1999 John Wiley & Sons, Ltd. PMID- 10523763 TI - Quantitation of blood and plasma amino acids using isotope dilution electron impact gas chromatography/mass spectrometry with U-(13)C amino acids as internal standards. AB - A method to quantitate blood and plasma amino acids by isotope dilution gas chromatography/mass spectrometry (GC/mS) is described. Samples were spiked with U (13)C amino acids as internal standards and the tert-butyldimethylsilyl derivatives (tBDMS) separated by capillary column gas chromatography. Linear regression curves, generated for individual amino acids, gave correlation coefficients of 0.9999. The reproducibility of the method was assessed from the analysis of 10 replicate blood and plasma samples. For most amino acids a coefficient of variance (CV) of 5% blasts in BM) as well as 3 patients acute lymphoblastic leukemia (ALL) who were in CR. In BM from ALL patients, an abnormal uniform B cell population was observed however antigen expression patterns varied greatly between patients. BM from acute myeloblastic leukemia (AML) patients showed an abnormal distribution of CD34+ cells. In addition, a correlation was observed between pre-BMT cytogenetics and MPFC. Only 2 out of 8 (25%) patients with normal MPFC pre autologous bone marrow transplantation (ABMT) relapsed (AML), while 6 out of 13 (46%) patients with abnormal pre-BMT MPFC relapsed including 2 out of 3 patients who were transplanted in clinical CR. Pre-BMT MPFC may thus be an effective tool for detection of MRD by detection of a pre-transplant MPFC abnormality. PMID- 10523798 TI - A strategy for erythropoietin treatment in myelodysplastic syndromes. PMID- 10523796 TI - The role of busulfan in bone marrow transplantation. AB - High-dose busulfan is an important component in many conditioning protocols for hematopoietic stem cell transplantation (HSCT) or bone marrow transplantation (BMT) in both adults and children. During the past 12y several studies have reported the wide inter-individual variability in busulfan disposition. Age, disease status, hepatic function, circadian rhythmicity, drug interactions and bioavailability, were identified as factors contributing to the high inter individual variability found in busulfan disposition. Traditionally, a standard busulfan dose of 4mg/kg/d for four days is used in most BMT/HSCT protocols. Many investigations have pointed out the pharmacodynamic relationship between a high busulfan systemic exposure and the occurrence of BMT related toxicity including hepatic veno-occlusive disease (VOD), interstitial pneumonia and alopecia in adult patients. However, studies in young patients have shown a high rate of graft failure and subsequently relapse which most probably is due to the low systemic exposure despite the standard dose schedule. In children and infants VOD was not observed with the standard doses. Increasing interest for the drug and new modification strategies for children led to higher rate of VOD and CNS toxicity when busulfan was administered according to the body surface area. More pharmacodynamic studies are required to establish the relation between the systemic exposure to busulfan and the therapeutic efficacy, especially in young children undergoing BMT or HSCT. In the present time an accurate and effective busulfan plasma level monitoring combined with dose adjustment based on the known pharmacological parameters may improve the clinical outcome for patients undergoing BMT. PMID- 10523800 TI - The role of neoadjuvant and adjuvant chemotherapy regimens consisting of different combinations of drugs in the treatment of advanced oral cancer. AB - We performed a retrospective analysis on the effect of neoadjuvant chemotherapy with three cycles of methotrexate (100 mg/m2 on day 1), cisplatin (90 mg/m2 on day 1) and bleomycin (20mg/m2 on day 1-5) with 21 d gap between each cycle in 44 patients with advanced squamous cell carcinoma of the cheek, lip and tongue followed by surgery and adjuvant chemotherapy consisting of cisplatin (90 mg/m2 on day 1), Mitomycin C (6 mg/m2 on day 1) and 5-fluorouracil (1000 mg/m2 120 h continuous infusion from day 1) repeated every 3 weeks for three cycles. Following induction chemotherapy, complete response was observed in 11 out of 44 patients (25%), and a partial response in a further 28 patients (64%). The overall median survival of all patients was 29 months and those in stage III and stage IV were 30 and 15 months respectively (P< 0.001). The median duration of the time to relapse in patients who responded to adjuvant chemotherapy was 28 months. The main toxic effect was vomiting followed by hematological toxicity. No treatment-related deaths occurred. The regimen showed a significant response, encouraging median survival and a good tolerability profile. PMID- 10523799 TI - Topoisomerase II-mediated alterations of K562 drug resistant sublines. AB - In order to further elucidate the roles of DNA topoisomerase II (topo II) subtypes, alpha and beta, as drug targets in chemotherapy, we have determined the enzyme levels in K562 cells selected for resistance to mitoxantrone (K562/Mxn), daunorubicin (K562/Dnr) and idarubicin (K562/Ida 20 and K562/Ida 60), as well as topo II-DNA complex formation, DNA damage and cytotoxicity, induced by topo II interactive agents, for example etoposide, teniposide, mitoxantrone and amsacrine. As compared to the parental cells, topo IIalpha/beta protein levels in K562/Mxn, K562/Dnr, K562/Ida 20 and 60 lines, measured with Western blot, were 17/67%, 85/88, 24/31% and 10/7% respectively. DNA damage, determined by DNA unwinding technique, induced by teniposide and amsacrine correlated with both topo IIalpha/beta protein levels (r2 = 0.8/0.9, P = 0.03/0.01 and r2 = 0.8/0.9, P = 0.04/0.01, respectively). Topo II-DNA complex formation induced by all studied drugs correlated with topo IIbeta protein levels (r2-range 0.8-0.9, P-range 0.01 0.04), while the correlation with topo IIalpha was weaker. Topo IIalpha/beta protein levels tended to show an inverse correlation with the cytotoxicity of etoposide (r2 = -0.9/-0.7, P = 0.01/0.06). The overall topo II-DNA complex formation correlated with drug-induced DNA damage (r2 = 0.9, P = 0.0001), whilst not with the cytotoxicity. Our findings indicate that both topo II isozymes are the targets of the antitumor agents studied, and of potential clinical relevance for prediction of treatment efficacy. They could play a role in tailored chemotherapy. PMID- 10523801 TI - The effect of two different doses of oral clodronate on pain in patients with bone metastases. AB - The aim of this study was to evaluate the efficacy of low dose oral clodronate in palliation of pain arising from bone metastases (BM) and to determine the optimal oral clodronate dose which inhibits osteolysis caused by tumor. Fifty patients with bone pain caused by BM were included in this study. All were receiving antitumor chemotherapy or hormonal therapy. The patients were randomized into three groups according to the dose of clodronate. Groups A and B were given 800 mg/d and 1600 mg/d of oral clodronate respectively for 3 months. Group C was the control group. The effect of clodronate in pain palliation was evaluated with pain score, performance status, and changes in analgesic use. The effect on osteolysis was examined with urinary calcium, hydroxyproline (OHP) and serum cross-linked carboxyterminal telopeptide region of type I collagen (ICTP) levels. Group A contained 16 patients, and groups B and C contained 17 patients each. After 3 months use of oral clodronate, significant decrease in the pain score of groups A and B was noted when compared to group C (P = 0.024 and P = 0.007, respectively). The analgesic use of 11 patients in group A (69%) and 8 patients in group B (47%) was decreased, but only the decrease in group A was statistically significant (P = 0.038). Pain score increased in 5 patients in group C (29%), and 3 patients in groups A (19%) and B (18%) each. Urinary calcium, OHP and serum ICTP levels increased in group C and decreased in groups A and B, but only the decrease of urinary calcium levels of group B was significant (P = 0.003). In conclusion, low dose (800 mg/d) oral clodronate seems to be as effective as standard dose (1600 mg/d) in palliation of bone pain secondary to BM. PMID- 10523802 TI - Prevention of recurrence and prolonged survival in primary central nervous system lymphoma (PCNSL) patients treated with adjuvant high-dose methylprednisolone. AB - Five patients at risk for primary central nervous system lymphoma (PCNSL) recurrence were treated with high-dose methylprednisolone (HDMP) to prevent 'trafficking' of malignant lymphocytes into the central nervous system (CNS). HDMP was chosen because of its ability to stabilize the 'blood brain barrier (BBB)'. Three men with newly diagnosed PCNSL, ages 62, 76 and 78y, whose survival was projected to be 6.6 months, began treatment after achieving complete response (CR) to initial radiation therapy alone and survived 27, 37 and 59 months after treatment. In none was death from recurrent disease in CNS but one patient did die of systemic non-Hodgkin's lymphoma (NHL) five years after PCNSL diagnosis. A 20 y old man was treated with HDMP after successful combined modality therapy and is alive 75+ months after initial diagnosis without evidence of disease recurrence. A 34 y old man relapsed after combined modality initial treatment and failed to respond to HDMP when treatment was begun after unsuccessful salvage therapy; he died of disease 12 months after initial diagnosis. There were no treatment complications. The promising results in this pilot study from the basis for a North Central Cancer Treatment Group (NCCTG) 96-73-51, a Phase 2 clinical trial of brain radiotherapy and HDMP for PCNSL patients 70y of age and older, a group of patients at high risk for toxicity from intensive combined modality therapy. PMID- 10523805 TI - Mouse genomics and informatics offer new tools for psychiatry PMID- 10523803 TI - A case of resectable lung adenocarcinoma associated with sarcoidosis. AB - A 71-year-old woman with uveitis was referred to our hospital for further examination of the possible underlying diseases. In roentgenological examination with plain X-ray and CT scan, hilar and mediastinal lymphadenopathy and a mass shadow in the right upper lung field was observed, whereas fibrotic changes were not obvious in both lung fields. Transbronchial lung biopsy with fiberoptic bronchoscope revealed granulomatous interstitial pneumonia. CD4-positive lymphocytes were increased in bronchoalveolar lavage. The patient was diagnosed as having sarcoidosis. Subsequently, right upper lobectomy was performed, and Stage I lung adenocarcinoma was diagnosed. The patient is under follow up without medication and the disease has been stable for two years. A relationship between epithelioid granulomatosis and malignant diseases is discussed and a review of the literature is given. Since it is still controversial as to the incidence of malignant diseases in sarcoidosis patients, it is important to accumulate data on these associations. PMID- 10523804 TI - Chimeric monoclonal anti-CD20 antibody (rituximab)--an effective treatment for a patient with relapsing hairy cell leukaemia. AB - A case story is presented, describing a 46 y old man, with a relapsing hairy cell leukaemia. After treatment with monoclonal anti CD-20 antibodies (rituximab) 375mg/week, four times, a complete remission was obtained which has lasted >9 months. The rituximab treatment produced a better remission than earlier treatments with alpha-interferon and chlorodeoxyadenosine. In addition, in contrast to other treatments, no initial worsening of the pancytopenia was observed. PMID- 10523806 TI - Mouse models of madness. AB - Psychiatric disorders have important genetic contributions, but it has been very difficult to identify the responsible genes using human populations. Recent developments in mouse genomics hold considerable promise of providing important insights into the genetics of these diseases. PMID- 10523807 TI - Knocking out the stress response. AB - Transgenic animals and knockout mice have been generated with defined defects in various components of the hypothalamic-pituitary-adrenal axis and the autonomic nervous system. These models provide valuable and novel insights into the development, crosstalk, organization, and functioning of the stress system. PMID- 10523808 TI - Searching for molecules mediating glial-neuronal communication. AB - The newly identified axotactin (AXO) protein in Drosophila, a member of the neurexin superfamily, appears to be a key element that mediates the effects of glial cells on neuronal membrane excitability and synaptic plasticity. PMID- 10523810 TI - Waldman and rowe reply PMID- 10523811 TI - Thapar and barrett reply PMID- 10523809 TI - Further lack of association between the 5-HT2A gene promoter polymorphism and susceptibility to eating disorders and a meta-analysis pertaining to anorexia nervosa. PMID- 10523813 TI - Linkage disequilibrium mapping of chromosome Xp11 for a schizophrenia susceptibility locus. PMID- 10523812 TI - Castellanos replies PMID- 10523814 TI - Mechanisms of typical and atypical antipsychotic drug action in relation to dopamine and NMDA receptor hypofunction hypotheses of schizophrenia. AB - Available evidence indicates that clozapine is the most effective antipsychotic currently used for the pharmacotherapy of schizophrenia. Unfortunately, clozapine can cause serious side effects that limit the use of the drug. The therapeutic mechanism of action of clozapine is poorly understood, and accordingly, it has been difficult to design new drugs with the advantageous therapeutic properties of clozapine. Based on hypotheses that dopaminergic and serotonergic receptor blocking properties of clozapine account for its clinical efficacy, several novel antipsychotic drugs have been introduced recently. There is currently insufficient data to reach definitive conclusions regarding the efficacy of the newer 'atypical' antipsychotics in comparison to clozapine. However, most published studies, and general clinical impressions, suggest that none of the newer drugs are as effective as clozapine in treating patients resistant to typical antipsychotic drug therapy. The present paper briefly reviews the clinical experience with the newer 'atypical' antipsychotic drugs and then discusses clinical and preclinical data potentially relevant to mechanisms of action of clozapine in relation to the NMDA receptor hypofunction hypothesis of schizophrenia. PMID- 10523815 TI - Psychiatric research in the 21st century: opportunities and limitations. AB - The year 2000 prompts numerous prognostications of the events to occur during the next century. Except for the Y2K problem, the year 2000 is little different in substance than 1999 or 2001. However it is useful to use the occasion to consider where we have come from and where we are going. Thinking of the future will help clarify goals, define the problems, identify priorities, and outline pathways for progress. PMID- 10523816 TI - Radiation hybrid mapping of genes in the lithium-sensitive wnt signaling pathway. AB - Lithium, an effective drug in the treatment of bipolar disorder, has been proposed to disrupt the Wnt signaling pathway. To facilitate analysis of the possible involvement of elements of the Wnt pathway in human bipolar disorder, a high resolution radiation hybrid mapping (RHM) of these genes was performed. A fine physical location has been obtained for Wnt 7A, frizzled 3, 4 and 5, dishevelled 1, 2 and 3, GSK3beta, axin, alpha-catenin, the Armadillo repeat containing genes (delta-catenin and ARVCF), and a frizzled-like protein (frpHE) using the Stanford Human Genome Center (SHGC) G3 panel. Most of these genes were previously mapped by fluorescence in situ hybridization (FISH). Frizzled 4, axin and frpHE did not have a previous chromosomal assignment and were linked by RHM to chromosome markers, SHGC-35131 at 11q22.1, NIB1488 at 16p13.3 and D7S2919 at 7p15.2, respectively. Interestingly, some of these genes were found to map within potential regions underlying susceptibility to bipolar disorder and schizophrenia as well as disorders of neurodevelopmental origin. This alternative approach of establishing the precise location of selected genetic components of a candidate pathway and determining if they map within previously defined susceptibility loci should help to identify plausible candidate genes that warrant further analysis through association and mutational scanning. PMID- 10523817 TI - Genetic alteration of the alpha2-adrenoceptor subtype c in mice affects the development of behavioral despair and stress-induced increases in plasma corticosterone levels. AB - alpha2-Adrenoceptors (alpha2-AR) modulate many central nervous system functions, such as regulation of sympathetic tone, vigilance, attention, and reactivity to environmental stressors. Three alpha2-AR subtypes (alpha2A, alpha2B, and alpha2C) with distinct tissue-distribution patterns are known to exist, but the functional significance of each subtype is not clear. Since specific, alpha2-AR subtype selective pharmacological probes are not available, mice with genetically altered alpha2C-AR expression were studied in order to investigate the possible involvement of the alpha2C-AR in physiological and behavioral responses to acute and repeated stress. A modified version of Porsolt's forced swimming test was used to assess the possible effects of altered alpha2C-AR expression on the development of behavioral despair. alpha2C-Overexpression increased and the lack of alpha2C-AR (alpha2C-KO) decreased the immobility of mice in the forced swimming test, ie alpha2C-AR expression appeared to promote the development of behavioral despair. In addition, alpha2C-KO was associated with attenuated elevation of plasma corticosterone after different stressors, and overexpression of alpha2C-ARs was linked with increased corticosterone levels after repeated stress. Moreover, the brain dopamine and serotonin balance, but not norepinephrine turnover, was dependent on alpha2C-AR expression, and the expression of c-fos and junB mRNA was increased in alpha2C-KO mice. Since alpha2C KO produced stress-protective effects, and alpha2C-AR overexpression seemed to promote the development of changes related to depression, it is suggested that a yet-to-be developed subtype-selective alpha2C-AR antagonist might have therapeutic value in the treatment of stress-related neuropsychiatric disorders. PMID- 10523818 TI - Identification of female-specific QTLs affecting an emotionality-related behavior in rats. AB - The influence of genetic factors on psychological traits and disorders has been repeatedly demonstrated; however, the molecular mechanisms underlying such an influence remain largely unknown. Anxiety-related disorders constitute the most common class of mental disorder in humans, with women being diagnosed far more frequently than men. A better understanding of the genetic and gender-related mechanisms mediating anxiety traits should enable the development of more rational methods for preventing and treating anxiety disorders. In this study we have aimed to identify, for the first time, quantitative trait loci (QTL) influencing anxiety/emotionality-related traits in rats. To this end, two strains Lewis (LEW) and Spontaneously Hypertensive Rats (SHR)-that differ for several behavioral measures of anxiety/emotionality were intercrossed. A QTL analysis of the F2 population revealed suggestive loci for various traits, including behaviors in the elevated plus-maze and blood pressure. In addition, one major QTL explaining 50.4% of the total variance (LOD = 7.22) was identified on chromosome 4 for the locomotion in the central and aversive area of the open field. Two other relevant QTLs have been recently mapped near this chromosomic region in the rat, which also harbors Tac1r, the gene encoding for the substance P receptor. Our major QTL affected females but not males and its effect depended on the type of cross (LEW or SHR grandmothers). The present results reveal a complex genetic basis underlying emotional behaviors and they confirm the existence of interactions between genetic factors and sex for this kind of trait. Further investigation of the loci identified herein may give clues to the pathophysiology of psychiatric disorders such as anxiety-related ones. PMID- 10523819 TI - Association of the serotonin transporter promoter regulatory region polymorphism and obsessive-compulsive disorder. AB - Although modulation of symptoms of obsessive-compulsive disorder (OCD) by serotonergic agents is well established, it is unclear whether an abnormality in the central serotonergic system is involved in its etiology. The serotonin (5-HT) transporter (5-HTT), which is the key modulator of serotonergic neurotransmission, is the target for serotonin reuptake inhibiting drugs (SRIs) that are uniquely effective in the treatment of OCD. In this preliminary study we report an association of a functional polymorphism in the 5-HTT 5' regulatory region and OCD. Seventy-five OCD Caucasian patients and 397 ethnically-matched individuals from a non-patient control group were genotyped for the 5-HTTLPR. Population-based association analysis revealed that patients with OCD were more likely to carry two copies of the long allele (l) as compared to controls (46.7% vs 32.3%: chi2 = 5.19, P = 0.023). This finding replicates a recent family-based study of this polymorphism in OCD, and thus indicates that the 5-HTTLPR may be associated with susceptibility to OCD. PMID- 10523820 TI - Alterations of hippocampal secreted N-CAM in bipolar disorder and synaptophysin in schizophrenia. AB - Schizophrenia and bipolar disorder have both been linked to structural abnormalities of the hippocampus, which is consistent with a neurodevelopmental anomaly. One isoform of the neural cell adhesion molecule (N-CAM) protein, cytosolic N-CAM 105-115 kDa, was previously shown to be increased in schizophrenia in the hippocampus and prefrontal cortex. Another isoform of N-CAM, the variable alternative spliced exon of N-CAM, was also increased in the hippocampus and prefrontal cortex of bipolar disorder patients. In the present study, the secreted isoform of N-CAM (SEC N-CAM), synaptophysin, and actin proteins were measured in the hippocampus of controls, suicide victims, and patients with bipolar disorder or schizophrenia by quantitative Western immunoblotting. Previous measurements of cytosolic N-CAM (105-115 kDa) protein, from the same hippocampus samples, were used to calculate the N-CAM (105-115 kDa)/synaptophysin ratio. An affinity purified antibody to SEC N-CAM recognized SEC N-CAM (108 kDa and 115 kDa) in brain but SEC N-CAM was not detectable in CSF. In bipolar disorder, but not in schizophrenia, an increased SEC N-CAM 115 kDa/108 kDa ratio was found as compared to controls (P = 0.03). The synaptophysin/actin ratio was significantly decreased in schizophrenia (P = 0.014) as compared to controls. The cytosolic N-CAM 105-115 kDa/synaptophysin ratio was increased in patients with schizophrenia (P= 0.017), but not in bipolar disorder. Thus, bipolar disorder patients show altered expression of SEC N-CAM in the hippocampus. Patients with schizophrenia show a decrease in synaptophysin and an increase in the cytosolic N-CAM 105-115 kDa/synaptophysin ratio. The results offer further evidence of differences in protein expression between bipolar disorder and schizophrenia in the hippocampus, which is consistent with a distinct neuropathology for each neuropsychiatric disorder. PMID- 10523822 TI - Homozygosity at the dopamine DRD3 receptor gene in cocaine dependence. AB - We examined the hypothesis that the dopamine D3 receptor gene (DRD3) is a susceptibility factor for cocaine dependence. The MscI/BalI polymorphism of the DRD3 gene was examined in 47 Caucasian subjects with cocaine dependence and 305 Caucasian controls. Based on prior studies with a range of psychiatric disorders we hypothesized there would be a decrease in the frequency of the 12 genotype in the patient sample (increased homozygosity). We observed a significant decrease in the frequency of 12 heterozygotes in subjects with cocaine dependence (29.8%) vs controls (46.9%) (P 200 ml), recurrent hematuria, bladder stones. New alternative and minimally invasive treatment such as TUNA generate necrotic lesions within the prostate through needle introduced endoscopically. This leads also to marked improvement in patients symptomatology. PMID- 10523897 TI - [Driver's licence: responsibility, ethics and deontology]. AB - The new law enacted on march 23rd 1998 regarding driving-licence, which became effective on october 1st 1998, modifies the method for obtaining the different types of driving-licences. Its 6th annex enumerates a list of physical impairments and pathologies implying the denial, the limitation or the withdrawal of the driving-licence; it determines if a medical advice, and if the case which kind of medical advice, is required. This leads to new situations, for which no jurisprudence has been established to date, where not only the responsibility of the licence's applicant but also that of the examining physician could be suspected. The physicians are also confronted to ethical an deontological problems on which the medical authorities have not always given an opinion. This paper examines them in order to give some bases for reflexion and discussion. PMID- 10523898 TI - [The thromboembolic risk of atrial fibrillation]. AB - The assessment of the thrombo-embolic risk is currently well defined in case of atrial fibrillation, in the general population as well as in several subgroups. Predictive factors of thrombo-embolism have been identified, they are clinical and echocardiographic criteria. They allow to stratify the individual risk of each patient and to establish the therapeutic attitude best suited to its thrombo embolic and haemorrhagic risk profile. Recent clinical trials have demonstrated that oral anticoagulation with coumarinics, adjusted at an INR between 2 and 3, provided a greater protection for patients at higher risk, compared to aspirin, with an acceptable low rate of haemorrhagic complications. When atrial fibrillation is of recent onset, the therapeutic attitude will take into account the time delay between onset of the arrhythmia and the medical consultation, 48 h representing the maximal delay allowed to perform cardioversion without prolonged anticoagulation. PMID- 10523899 TI - [Resistant atrial fibrillation: how far does one go in its treatment?]. AB - Resistant atrial fibrillation consists in recurrent or persistent fibrillation despite a conventional treatment persistent. When conventional treatments (drugs, external defibrillation) fails, we have the following options: internal defibrillation, pacemaker implantation either standard or with biatrial pacing, curative or palliative radiofrequency ablation techniques, antiarrhythmic surgery, atrial defibrillator implantation. These different treatment modalities, isolated or combined will frequently be able to reduce the patient symptoms and some of at them may suppress the majority of atrial fibrillation episodes. Not all these treatments are widely used at the present time but a lot of research is being done about them. The purpose of this article is to describe their use in resistant patients. PMID- 10523900 TI - [Anticoagulation in cardiac pathologies (except atrial fibrillation)]. AB - Anticoagulants, including heparins and antivitamins K are medicines which are largely prescribed in cardiology. Even though the outlines of the treatment have been clearly established for a few decades, new cardiological indications have recently appeared. The long-term treatment of cardiac valvular prosthesis by oral anticoagulants significantly reduces the risk of thrombo-embolic incident among people carrying a cardiac valvular prosthesis. In case of non-operated valvulopathy, treatment indication must be evaluated for each patient; in so doing, the connection benefit/risk must always be taken into account. In case of dilated cardiomyopathy, the treatment prescription must be limited to the patients with a high embolic risk. Other cardiological indications (apart from atrial fibrillation) must be carefully weighed up. PMID- 10523901 TI - [Surgical treatment of Parkinson disease: indications and limitations]. AB - Thalamotomy, a surgical lesion of the ventro-intermediate nucleus of the thalamus, is a well known surgical treatment of tremor in Parkinson's disease. Over the last years, new surgical therapies had been developed. These therapies, instead of making a lesion in the brain, consist in placing electrodes in specific areas of the brain and to inhibit neuronal function by electrical stimulation. Electrical stimulation of the subthalamic nucleus or of the pallidum are effective to treat motor symptoms of Parkinson's disease. The procedure can be done bilaterally, contrary to thalamotomy. A short overview of the indications and contra-indications of this kind of therapy is given. PMID- 10523902 TI - [The role of interferon beta in the treatment of multiple sclerosis]. AB - Because of the demonstration for the first time of a measurable effect on magnetic resonance imaging, the publication in 1993 of the results of the American trial of interferon beta-1b in multiple sclerosis constituted a turning point in the history of multiple sclerosis (MS) treatment. Although many questions subsist concerning its optimal use, it has become a standard in the treatment of relapsing remitting MS to which new and old drugs will have to be compared. It seemed therefore meaningful to review the results obtained with recombinant interferon beta-1b and the more recently developed interferon beta 1a, and to place them in the context of the other immunomodulatory treatments currently offered in MS. PMID- 10523903 TI - [Communication with the aphasic patient]. AB - The consequences of aphasia with reference to the WHO ICIDH classification are evoked. The role of the physician and the behaviour of the relatives are underlined as well as the main parameters to be taken into consideration to communicate with an aphasic patient. As a rule, a dialogue in quiet surroundings will make communication easier with an aphasic patient. Our attitude will be modulated according to the behaviour of the patient, his past and the aphasiological picture. At the early stage of the disease, verbal means of communication will be mainly used, whereas later on, in a pragmatic approach, all means (verbal and non verbal) will be used to improve communication. PMID- 10523904 TI - [Dementia and Alzheimer disease. Early detection and treatment]. AB - Early diagnosis and treatment of dementia are intimately connected following recent progress in these topics and limitation of actual treatments to the initial cause of this ailment. Precise diagnosis alone can lead to adequate treatment and a logical procedure is mandatory to define, as precisely as possible, the exact aetiology. The first step will be to establish if the patient is demented, then if it is a curable dementia and finally if the dementia is treatable. If necessary, work up can be completed by a precise typing of the disease. Beside support which is a basic option for these diseases but out of the scope of this paper, the clinician will aim, if possible, to prescribe an aetiological treatment or at least a symptomatic one. This last can be not only non-specific, based on antidepressants, sedatives and neuroleptics, but also specific. This last therapeutic option has tremendously evolved recently with the coming of acetylcholine aimed treatment, particularly cholinesterase inhibitors which are the first medications to be demonstrated as effective for Alzheimer's disease. Even if these drugs have still significant limitations, therapeutic nihilism which was often the rule has to be abandoned and must be an incentive to keep on with research for better diagnostic tests and therapeutic options. PMID- 10523905 TI - [What degree of freedom is there for living donors?]. AB - The shortage of cadaveric donors has induced a renewed interest in living kidney donation. This paper describes the legal, religious and ethical factors which ensure or restrict the autonomy of the potential donor. We conclude that it is possible with appropriate measures to protect his freedom of choice. PMID- 10523906 TI - [To what extent is organ procurement acceptable? The legal framework and European legislation]. AB - Progress in the field of organs and tissues transplantation contributes today to saving lives and improving their quality. Consequently, the increasing need of organs and tissues and their constant shortage give rise financial benefits and commercial risks. Several European countries have tasted in one or another way organ trading. These ethical and legal important problems implicated preventive measures in European countries and in the whole world. Since 1986, Belgium has a national legislation which forbids in every forms, organs and tissues commerce. In addition, an item imposes penalties in this connection. Meanwhile different international organisations established acts, recommendations and resolutions preventing purchase and sale of human organs and tissues. PMID- 10523907 TI - [To what extent is organ procurement acceptable? From principles to reality]. AB - Organ trade is unlawful; organ gift is promoted on condition it is gratuitous, anonymous and spontaneous. The idea of graft and transplantation is understood differently according to culture, religion and ability of a region to supply to its needs in the field of health. The patient's eagerness can explain his quest of a donor in whatever country he is. Even those physicians who have strict ethical guidelines must pay attention not to be involved in blameable jobs. The only way to avoid rewarded organ gifts is to prohibit transplantation touring and grafting from friends, at the risk to loose true-hearted and noble donors. PMID- 10523908 TI - [Asthma and chronic obstructive bronchopneumopathy: differential diagnosis]. AB - Asthma and COPD are common problems seen by general practitioners. They share several signs and symptoms, so that a differential diagnosis generally needs lung function testing, with at least a spirometry, either performed on several occasions, or completed by bronchodilation or non specific bronchoprovocation tests. Spirometry should become one of the tools used daily by a general practitioner. PMID- 10523909 TI - [The treatment of asthma]. AB - As asthma is a chronic, predominantly inflammatory process, maintenance therapy must be emphasized. It consists in most patients to regularly inhale a steroid agent (fluticasone or budesonide), in combination with non pharmacological interventions and occasional inhalation, as needed, of a short-acting sympathomimetic agent. When single therapy with an inhaled steroid is not sufficiently efficient, adding a long-acting sympathomimetic agent (salmeterol or formeterol) is recommended. Treatment of acute asthma includes repeated administration, at short interval, of an inhaled sympathomimetic agent (salbutamol or terbutaline) combined with oxygen and a systemic corticosteroid. PMID- 10523910 TI - [Treatment of chronic obstructive bronchopneumopathies]. AB - Guidelines and statements from ERS, ATS, BTS and SPLF are now available for adequate treatment of COPD. The recommendations are summarised but the author emphasizes what has changed, what remains unchanged or was adapted and what is new and promising. The therapeutic approach tends also to improve symptoms, especially dyspnea, and quality of life. This is mainly obtained with a multidisciplinary team offering the patient a program of rehabilitation. PMID- 10523911 TI - [Chronic cough]. AB - Cough becomes chronic after three weeks of evolution. Chronic cough is due to four syndromes in 90% of cases: postnasal drip syndrome, asthma, gastroesophageal reflux and chronic bronchitis. Each syndrome needs a specific therapeutic approach. Antitussive drugs like dextromethorphan are prescribed in cases of complicated cough. Cough secondary to angiotensin converting enzyme inhibitors must not be neglected. In case of failure of initial check up or lack of response to specific therapy, a more thorough examination must be conducted in a specialized centre. PMID- 10523912 TI - [A case of secondary cough]. PMID- 10523913 TI - [The aging Joseph]. PMID- 10523914 TI - [Vaccinations: a necessity]. AB - The preventive efficiency of vaccination for numerous infectious diseases remains up to date. Multiple epidemiological data are documenting the underuse of this approach. Risk situations are briefly reviewed, specially those related to unappropriated immunologic functions associated with ageing. Both extremes of life can display insufficient responses to vaccines, either by immaturity or immunosenescence. The diagnostic interest of these situations, by following the vaccinal responses, resides in allowing some prophylactic treatments (antibiotics, gammaglobulins) or consider immunostimulating treatments. This is not to speak, for the older subjects, about the repletion of multiple deficits of their nutritional status. PMID- 10523916 TI - [Influenza vaccination of elderly persons: state of the art]. AB - Influenza is a major health problem among elderly people in industrialized countries. Elderly individuals suffering from chronic illnesses are more prone to suffer complications from influenza infection. In institutions, the outbreaks of influenza may touch about 60% of residents and induce life-threatening complications in more than 25% of cases. In this population--generally very old and handicapped by several chronic physical and mental diseases--the mortality associated with infections is very high. Influenza vaccination has been shown to be effective in 33% of vaccinated elderly persons for preventing clinical infection and in 74% for preventing mortality. This efficacy seems to be age related. The other factors implicated in the immune response are discussed. A better understanding of these relationships is of great importance for public health and could improve the immune responses after influenza vaccination. PMID- 10523915 TI - [Vaccination of children in 1999]. AB - Childhood immunization programs are regularly reevaluated to take into account epidemiologic changes in the diseases covered by the vaccines as well as the development of new vaccines. Among the significant additions brought in Belgium to the childhood immunization program in recent years are immunization against hepatitis B during infancy, as well as the administration of a second dose of measle-mumps-rubella vaccine around the age of 12 years. The switch to inactivated polio vaccine will be proposed until the eradication of this disease which is expected in a few years. Combination vaccines including all injectable vaccines recommended for administration during the first year of life including acellular pertussis are in their final stage of development. PMID- 10523917 TI - [The rights of patients--medical concerns]. AB - The protection of patients rights is neither a new concern nor a limited to Belgium one. The study shows that, at all times, the doctors have been worried about the necessity of respect of the patients. This respect is inherent to the confidence patients have in the doctors. The study also explains that the policy framework on the rights of the patients has been developed in the world and in particular, in Europe, after World War II. In a third part, the belgian legislative initiatives are commented according to the opinions of famous legal writers. Finally, the study emphasizes the fact that it is necessary to stimulate dialogue-structures for doctors and patients and that we have to avoid too rigid rules. PMID- 10523918 TI - [Arterial hypertension of renovascular origin. The value of color Doppler ultrasonography in the detection of stenosis of the renal artery]. AB - Color Doppler sonography is a performant method to detect direct signs of renal artery stenosis with a feasibility rate superior to 90%. The reliability of the method, evaluated with regard to the angiography as reference method, varies, according to the authors, between 80 and 90% with a sensitivity and a specificity between 85-90%. The investigation is above all dedicated to a population where prevalence of renal artery stenosis is high: hypertensive patient with lower limb arteriopathy, or with impairing of the renal function under angiotensin convertase inhibitor, or with hypertension resistant to a multi-therapy, young woman hypertension. The success of the method requires a trained operator, a modern equipment and a precise methodology: a fasting patient, patient positioning adapted to the spectral signal recording, peak velocity measurements at the anatomical sites of stenosis (ostium, first centimeters). Investigation length does not exceed 20 minutes. The exploration is totally atraumatic and does not require any contrast injection. PMID- 10523919 TI - [Spiral computed tomography of the renal arteries]. AB - With the recent advent of spiral or helical CT, the ability to acquire large volume of imaging has become possible. Fast scanning of both kidneys, the aorta and the renal vessels can be accomplished during one breathhold. Early reports of CT angiography in the evaluation of renal artery stenosis indicated a sensitivity of 92% utilizing the MIP projection method and 59% utilizing the shaded surface display method. Specificity for both method was approximately 82%. More recent articles reports for hemodynamically relevant renal artery stenosis a sensitivity/sensibility of 96/99%. The introduction of a new multidetector CT technology will probably increase those results and the indication of CT angiography. PMID- 10523921 TI - [Overall and separate clearance--renography with and without captopril]. AB - The present work briefly describes the radionuclide renal functional tests available for the work up of adult hypertensive patients. The use and specific indications of the different techniques and radiopharmaceuticals are reviewed and summarized. A special attention is given to captopril enhanced renograms: these non invasive procedures contribute to the diagnosis of renovascular hypertension. PMID- 10523920 TI - [Magnetic resonance angiography of the renal arteries]. AB - Initially, the clinical use of magnetic resonance angiography (MRA) in the abdomen has been restricted because of motion and flow related artifacts. The advent of high performance gradient systems made possible the development of 3D gadolinium-enhanced MRA techniques and expanded the clinical applications of MRA into the abdominal area, particularly for the investigation of renal arteries. This technique is safe, because the administered contrast agent (gadolinium) is free of clinically detectable nephrotoxicity and has a low incidence of allergic reactions. Moreover, contrast MRA also eliminates the risks of ionizing radiation which allows repeating the examination without the accumulation of radiation exposure. The main disadvantages of the technique are its low availability and the fact that the use of contrast agents for this procedure is still not reimbursed by the social security. Many studies demonstrated that contrast MRA allows for the reliable assessment of renal artery morphology and pathologic states. Furthermore, within a single MR examination a comprehensive approach including renal artery morphology, hemodynamic significance of any stenosis and kidney perfusion is available. In this paper, we provide a review of the literature concerning the clinical performance of contrast MRA for the renal arteries and suggest its rationale for the investigation of patients suspected of renovascular disease in our specific environment. PMID- 10523922 TI - [Arteriography and treatment using angioplasty]. AB - Renal angiography is certainly not the first examination to be done in case of hypertension. Its interest remains however high. It allows to demonstrate without doubt the reality of a significant stenosis. It can also be associated with a therapeutic procedure. The indications, with practical modalities, results and eventual complications are described below in two different parts: diagnostic and therapeutic. PMID- 10523923 TI - [Imaging and arterial hypertension: algorithms and conclusions]. PMID- 10523924 TI - [Future perspectives in medical imaging]. AB - Medical imaging, less than 105 years after the invention of X-rays in medicine, shows signs of extraordinary and almost exponential rise. Among the new techniques, the stress is laid on the last developments of echography like the imaging in three dimensions or the harmonic imaging. The helical scanner currently allows imaging in real time, from incredibly short times of acquisition and launches out in the virtual endoscopy. The MRI and particularly angio MR allows the visualization of small arterial or venous malformations as for soon the study of the coronary vessels with a reliability which will compete with the coronarography. The diagnostic angiography is undoubtedly destined for disappearing as of many other techniques which will be one day obsolete, or superfluous compared with such relevant and less invasive methods. PMID- 10523925 TI - [Imaging of cancer using positron emission tomography]. AB - Positron emission tomography (PET) is a method making use of short half-life radioactive compounds which allow imaging and quantification of functional and metabolic data at the level of multiple organs. This method has been initially oriented towards neurological and cardiological applications but gets now a more widespread use in oncology. This recent development has been made possible thanks to methodological progresses allowing "whole body" imaging and thanks to the use of a practical tracer, the 2-[18 F]fluoro-2-deoxy-D-glucose (FDG). The uptake of this tracer is enhanced in diverse cancer tissues. Recent studies has clarified the biological processes which lead to this enhanced uptake of FDG in cancers. This new insight allows a rational and helpful usage of PET in diverse aspects of clinical oncology: diagnosis of lesion, staging, follow-up of patients and treatment evaluation. PMID- 10523926 TI - [AIDS: the impact of new molecules]. AB - In recent times important progress has been made in the treatment of HIV. Plasma viral load count is now used in routine practice permitting a accurate evaluation of each patient and in particular the risk of progression to symptomatic AIDS. A better understanding of the natural history of HIV has enabled physicians to recognise the need for commencing treatment early. The availability of more effective therapies like protease inhibitors and their use in tritherapy regimens permit more ambitious therapeutic strategies aimed at prolonging life and perhaps even viral eradication. Already the first positive effects of these treatments have been observed with the reduction of mortality and in hospitalisation rates, etc. The impact of these developments on the prevention and testing for HIV is of course very important with the risk of a demobilizing effect in primary as much as in secondary HIV prevention. Conversely the possibility of very early treatment is a major argument for effective education on early HIV testing. The future politics of prevention and testing must be elaborated taking into account these changes in the public perception of HIV. PMID- 10523927 TI - [New treatments for impotence]. AB - The development of new oral drugs and the poor results of the reconstructive vascular surgery has modified the therapeutical approach of the impotent patient. The doctor has to be aware of these changes in the management of impotence. PMID- 10523928 TI - [New treatments for obesity]. AB - The aim of this short review is to summarize our knowledge on the pharmacological treatment of obesity. We consider first the old molecules, mostly amphetamine derivatives, which tend to be abandoned and second, the more recent ones which just appeared or are just about to become available. We also envision some therapeutical perspectives derived from recent advances in molecular biology. Although, it is "scientifically correct" to deliver an optimistic message regarding the possibility of finding an effective and safe treatment of obesity in the near future, we think that this is, on the contrary, an unrealistic goal. PMID- 10523929 TI - [Rejuvenating hormones]. AB - The clinical and biological syndromes of menopause, andropause, somatopause and adrenopause are presented successively. Various substitutive hormonotherapies (including melatonin) are considered according to their efficacy, risks and cost. PMID- 10523930 TI - [Organochlorine pesticides and polychlorinated biphenyls in the Vistula river water]. AB - The composition and loads of organochlorine pesticides (DDTs, HCBs, HCHs, CHLs) and polychlorinated biphenyls (PCBs) transported with the Vistula River waters to the Gulf of Gdansk in 1991-1992 has been determined. The method of organochlorine compounds measurement was capillary gas chromatography with ECD after adsorption of the analyte on Ambertlite XAD-2, resin and subsequent elution, clean-up and HPLC fraction of the extract. The concentrations of DDTs, HCBs, HCHs, CHLs and PCBs in the Vistula River water ranged between 120-840, 7.6-52, 1600-410,000, 8.1 57 and 120-300 pg/l, respectively. During 12 months period of the study the total load of DDTs, HCBz, HCHs, CHLs and PCBs transported with the Vistula River water to the Gulf of Gdansk was assessed on 10.54, 0.73, 1377, 0.38 and 5.02 kg, respectively. PMID- 10523931 TI - [Dieldrin, aldrin, endrin, isodrin, endosulfan 1 and 2 on fish in the Gulf of Gdansk]. AB - The residues of dieldrin, aldrin, endrin, isodrin, endosulfan 1 and 2 has been determined in a several species of fish caught in the Gulf of Gdansk in 1992. The method of measurement was capillary gas chromatograph and low resolution mass spectrometry (HRGC/LRMS) after a nondestructive extraction and clean-up step with a further fractionation of the extract on Florisil column. Apart from dieldrin no other cyclodiene pesticides studied were found in fishes in detectable amounts, and for dieldrin concentrations ranged from 0.84 to 6.6 ng/g wet weight. PMID- 10523932 TI - [Determination of iodide ions content in the edible salts and medical preparations by spectrophotometric and voltamperometric methods]. AB - In this paper the spectrophotometric and voltamperometric methods for the determination of the iodide ions content are presented. The conformity of the edible table salts iodisation to the Polish Standards and to the contests declared by the producer was verified. PMID- 10523933 TI - [The contents of radon in deep borehole water of hydro-geological region of Gdansk]. AB - Radon 222Rn in deep borehole water of Gdansk Hydrogeological Region has been quantitative determined. This region is located in east part of Gdansk Voivodship and in west part of Elblag Voivodship including Zulawy. The measurements were performed using alpha liquid scintillation counting method. Only in some case the concentrations of 222Rn in investigated samples exceed recommended limit 11 Bq/l. PMID- 10523934 TI - [The influence of cooking on radiocaesium contamination of edible mushrooms]. AB - Radiocaesium concentration in some kinds of edible mushrooms collected in October 1990 has been determined to evaluate the radiocaesium activity 5 years after Chernobyl accident. The highest activity was found in Xerocomus subtomentosus (1080.5 Bq/kg of fresh weight), then in Rozites caperata (768.5 Bq/kg) and Xerocomus badius (562.5 Bq/kg); the lowest--in Suillus luteus (52.0 Bq/kg) and Cantharellus cibarius (63.0 Bq/kg). Studies on the influence of cooking on radiocaesium activity revealed that parboiling and boiling of mushrooms led to high, even 85% losses of radiocaesium in the product. Samples of Xerocomus badius collected in various sites of North-East Poland in 1995 averaged to 195.4 +/- 125.5 Bq/kg of fresh weight. PMID- 10523935 TI - [The use of "sous vide" technology in the packaging of chilled and ready to serve food]. AB - As chilled precooked dishes show limited to 3-5 days shelf life several additional factors have to be applied to extend it up to 21 or even 42 days as is sometimes allowed for sous vide technology products. Those factors comprise high hygienic standards for raw materials and premises as well as technological steps and parameters that efficiently destroy microbial contamination, and do not allow for recontamination or bacterial growth. Such steps include precooking which also means pasteurisation in high vacuum or anaerobic atmosphere in sealed pouches, blast chilling, low temperature storage parameters as well as high temperature of reheating process and quick serving procedures. Paper specifies parameters for each technological steps and presents microbiological requirements for final products. Sous vide technology allows for good quality and high nutritional value in soups, meats in sauces and stewed vegetables. It is used for individual consumer in chilled "ready to eat" line dishes in supermarkets and supplies such dishes for catering units. PMID- 10523936 TI - [Health quality criteria and the analytical procedures for the evaluation of pacifiers]. AB - The health quality criteria for the evaluation of soothers for babies and young children have been developed and the analytical procedures were collected and indicated for the evaluation purposes. They include: sensory analysis global migration migration of metals (Sb, As, Ba, Cd, Pb, Cr, Hg, Se) to 0.07 M HCl analysis of volatile compounds chemical demand for oxygen presence of reaction accelerators presence of antioxidants The requirements concerning mechanical and chemical parameters and also general safety of the product have been developed and the procedures for the conformity checking of these parameters, based on Polish Standard and the draft project of European Standard EN 14000 series concerning soothers for babies and small children have also been presented. These criteria have been accepted by the Ministry of Health and Social Welfare as suitable for issuing of health certificates for placing on the market as well as for the current sanitary surveillance of soothers taken from the market. The criteria will be obligatory until the EN 1400 Standards come into force being introduced into Polish legislation. PMID- 10523937 TI - [Variability of equivalent traffic noise level for the establishment of its standards]. AB - This article includes analysis of variability of equivalent traffic noise level of great intensity together with regard to variable time references. It also concerns normative times applied and proposed in Poland. It was discovered, that short-term measurements--30 or 60 min, usually don't reflect environmental acoustic conditions precisely. Long-term measurements and measurements conducted in the first night hour are characterized by great repeatability. The difference between is merely two dB. The above range of variability is an exact measure of valuation of acoustic conditions. Newly introduced system of regulations in Poland, the concerns protection of environment make the acoustic requirements 2-3 dB less severe with reference to previous ones. According to present system of noise measurement the difference in equivalent noise levels between day and night is 4-6 dB. Less difference occurs in in districts of intense traffic and through traffic. PMID- 10523938 TI - [The evaluation of nutrition quality of adolescents living in residential schools at the province of Bialystok]. AB - The aim of this study was an assessment of nutrition quality of adolescents living in boarding schools at the province of Bialystok. The investigations were carried out in the range of nutritional value and contamination level of lead, cadmium, mercury, copper and zinc in daily food rations given to young people. The nutrition quality was determined by calculating the energy and nutritional components during ten days (decade), using computer programme "Menu". Chemical analysis were made on individual meals, which create daily food ration per estimated day. All the studies (in 1997 and 1998 year) were done according to the methods referred in chapter on this article "Material and methods of research". It was found, (during two years study) that nutrition of young people in boarding schools devites from recommended dietary allowances. High products consumption from groups: "meat and its products", "butter" and "other fats", caused too high energy contribution taken from dietary fats of whole daily energy and high iron intake. The percent proportion from energy supplied from proteins was maintained in recommended value. The consumption deficit in groups "milk and dairy products", "vegetables and fruit rich in vitamin C reflected in the low percent of realization of the requirements for calcium and vitamin C. Exceedation of permissible tolerable weekly intake (PTWI) of cadmium and mercury wasn't stated, (in 1997 year) while 26% of estimated in 1997 year, and 14% in 1998 diets were above PTWI for lead. Daily intake of copper and zinc was lower than maximal tolerable daily intake (MTDI). Some assessed (during 1997 and 1998 year) rations didn't cover the requirements for zinc, while in 81% estimated diets in 1997 and 76% in 1998, the recommended intake level of copper was exceeded. The contents of sodium chloride in daily food rations analyzed in 1997 year ranged from 13.8 g to 27.2 g and the highest source of dietary salt were dinner meals (6.9 g-13.2 g), analogous in current year--from 10.8 g to 38.3 g, with contents of salt in dinner meals--5.7 g to 14.4 g. The investigation from 1997 and 1998 year prove, that nutrition of adolescent in boarding schools isn't correct according to rational diet principles. There is the need of giving systematic training for people who are planing and realising nutrition in boarding schools, and taking up other activity mobilizing personnel for higher engagement in young people nutrition problem. PMID- 10523939 TI - [The evaluation of organizational structure and work hygiene in milk kitchen centers in Poland on the basis of questionnaire]. AB - On the basis of available publications and obligatory legal regulations regarding the organisation of collective feeding patterns in Social Service Health Centres a questionnaire study was established to examine the way milk kitchen function in Poland. The study covered 533 milk kitchen centres (in hospitals, orphanages and creches) from February to December 1997. The afore-mentioned study analysed the general characteristics of the milk kitchen centre, its type of work, supervisory methods and the personnel's hygiene. From the results of this research it can be concluded that the organisational structure of the milk centres was appropriate, appliances and equipment in the majority centres were adequate but basis utensils were insufficient. It is obvious that dietetic feeding methods for this particular group of children are insufficiently applied. Only 22.2% of the centres apply the recommended proposals i.e. without boiling, without pasteurisation, ready for consumption when preparing, infant formulae, follow-up formulae and other mixed products. Work hygiene and health quality of production in the milk kitchen centres is subject to systematic controls by competent individuals from the State Sanitary Inspection services. In the years 1995-96, 72 samples of infant food that mainly came from hospital milk kitchen centres were taken due to the poor microbiological quality of the food. The considerable changes that have taken place over the last few years as far as infant feeding is concerned, for example, the promotion of breast-feeding, taking advantage of prepared dietetic products for infants (infant formulae, follow-up formulae, vegetable products, fruits, vegetable-meat products, cereal products) require changes at the organisational level of infant collective feeding which was confirmed by the afore-mentioned study. PMID- 10523940 TI - [Sensitivity of Klebsiella pneumoniae strains to the disinfectants]. AB - The sensitivity of 2 Klebsiella strains (isolated from hospital environment--Ks and museum--K28) to 7 disinfectants with the sensitivity of referent strain E. coli NCTC 8196 were compared. Suspension method was applied. Determined the sensitivity Klebsiella strains for phenol, septyl, lizol, chloramine, formalin, glutaraldehyde and laurosept in compare with sensitivity of E. coli during 10 minutes of exposure. Certify the insignificant of difference in testing sensitivity of both Klebsiella strains on the majority disinfectants and more sensitive those strains than referent strain E. coli. In the case of chloramine the difference was almost two fold--the value concentration ratio of the solutions giving bactericidal effect for E. coli in comparing the some concentration for Ks was 2.3. Only in the case of formalin the sensitivity of E. coli and Klebsiella pneumoniae was inverse--the value of concentration ratio was 0.51--E. coli strain was 1.9 more sensitive than Ks strain and 1.6 more sensitive than K28 strain. PMID- 10523941 TI - [Cardiology. Future status]. PMID- 10523942 TI - [Chronic heart insufficiency should be treated also with spironolactone]. PMID- 10523943 TI - [Revascularization in the treatment of ischemic heart disease--state of development in Denmark]. PMID- 10523944 TI - [Biochemical diagnosis of acute coronary syndrome]. PMID- 10523945 TI - [Coronary arteriography and coronary angioplasty in stable ischemic heart disease. Value in the prediction and prevention of future acute myocardial infarction]. AB - Acute myocardial infarction (AMI) is usually caused by the sudden formation of an intracoronary thrombus that occludes the coronary artery at the site of a vulnerable atherosclerotic plaque. Coronary angiography (KAG) offers the opportunity to visualize and characterize coronary artery lesions. The demonstration of significant stenoses (> 50%) often leads to mechanical revascularization, including coronary angioplasty (PTCA). Several studies in which serial angiograms were performed on patients who subsequently had AMI have shown that most of these acute events develop from lesions that on the first KAG were nonsignificant (< 50%). The KAG method does not adequately predict the location of the culprit plaque that will subsequently produce AMI PTCA results in less severe angina, but the price may be a higher rate of procedure related acute events. Large scale trials comparing the prognostic effect of an intense medical therapy versus PTCA with and without stenting are required to better define the independent and combined roles of the different therapeutic modalities in stable ischaemic heart disease. PMID- 10523946 TI - [Beta-blockade--a new therapeutic modality in chronic heart insufficiency]. AB - Heart failure due to decreased left ventricular function is a condition with a considerable morbidity and mortality. Until recently beta-blocker treatment has been considered contraindicated in this condition. During the last 20 years a number of investigations have pointed to a possible positive effect of beta blocker treatment in chronic heart failure and recently several major randomized trials have shown a significantly increased survival during beta-blocker treatment. This review summarizes the background for the use of beta-blocker treatment in chronic heart failure patients. PMID- 10523947 TI - [Pediatric heart surgery]. AB - Surgery for congenital heart defects has since its beginnings fifty years ago evolved from being palliative to corrective in many cases. Surgery for the more simple defects is now performed with low surgical mortality and excellent prognoses, but the mortality for some of the more complex defects is still high, and the treatment regarded as palliative. The principles for and the results of the surgical treatment of some more complex defects are reviewed. The importance of follow-up is emphasized. Future possibilities are discussed. PMID- 10523948 TI - [Infections and cardiovascular disease]. AB - Epidemiological studies indicate that respiratory tract and dental infections increase the risk of atherosclerotic cardiovascular disease. Several microorganisms have been claimed as clinically important, especially Chlamydia pneumoniae (Cp) and cytomegalovirus (CMV), Cp is frequently isolated from atherosclerotic plaques, and treatment with macrolide antibiotics may have a beneficial effect on the course of ischaemic heart disease. CMV seems to play a role in the development of coronary stenosis following coronary bypass surgery and heart transplantation. Likewise, inflammatory markers are associated with atherosclerotic cardiovascular disease. The pathogenetic role of microorganisms may be ascribed to the inflammatory response, which is elicited during infection. There is an urgent need for more documentation of the role of microorganisms in cardiovascular disease, and of the possible clinical effect of antibiotic treatment. PMID- 10523949 TI - [Diagnosis of acute myocardial infarction in Denmark]. AB - The introduction of new biochemical markers for myocardial damage in the recent years and different application of these methods in different centres may have an impact on the diagnostic criteria for acute myocardial infarction (AMI). By means of a questionnaire we studied the diagnostic criteria for AMI in relation to the use of different biochemical markers among 78 Danish hospitals. There were large variations with regard to the choice of cardiac markers and diagnostic values for different markers. CK-B is the cardiac marker mostly used followed by CK-MB. Troponin-T test was used by about 20% of the centres. Many centres are planning to use CK-MB and Troponin-T test. A common national and international policy for diagnosis of AMI in relation to different cardiac markers should reduce these improper differences. PMID- 10523951 TI - [Chlamydia pneumoniae and atherosclerosis--the hen, the egg or five feathers?]. PMID- 10523950 TI - [Cardiac adaptation to physical training--physiological and clinical aspects]. AB - Vigorous physical training induces left ventricular hypertrophy (LVH), a so called "athlete's heart". The hypertrophic response represents an adaptation to the increased haemodynamic load on the heart associated with physical exercise. Wall thickness and/or left ventricular internal diameters are specifically increased depending on the type of haemodynamic load (volume- or pressure load). It is a consistent finding that left ventricular mass is increased in elite athletes when compared with sedentary controls, but few athletes have cardiac dimensions exceeding accepted normal values. Most commonly these athletes are male oarsmen. Occasionally, it may be difficult to distinguish physiological LVH from primary heart disease and therefore it is of importance also for physicians to be familiar with the "athlete's heart". Also, the sensitivity of the non invasive methods for detecting coronary heart disease is reduced in athletes with LVH. This should be considered in the planning of the diagnostic strategy in athletes with LVH and suspected ischaemic heart disease. PMID- 10523952 TI - [Cardiac scanning with positron emission tomography--PET. Clinical use and research aspect]. AB - Assessment of regional myocardial glucose metabolism and regional myocardial perfusion has become possible with positron emission tomography (PET). These parameters are of importance in distinguishing viable from fibrotic myocardium in patients with ischaemic heart disease. PET scanning appears to be the method of choice in these patients, which has led to an increased clinical application of PET as a method usable to select patients with severe heart disease before potential revascularisation. In the present review, PET technology is briefly described, together with an overview of the scientific evidence supporting the clinical application of cardiac PET. Finally, its applications in the fields of pathophysiology and pharmacology are briefly described. PMID- 10523953 TI - [Picture of the month. Sleep apnea]. PMID- 10523954 TI - [Price and quality of early medical abortion]. PMID- 10523955 TI - [Glaucomatous optic neuropathy]. AB - Glaucomatous optic neuropathy is the major component in the pathogenesis of all forms of glaucoma. The author describes the anatomy and blood supply to the optic nerve head, the pathohistologic changes in the optic nerve, pathogenesis of glaucomatous optic neuropathy, and the treatment philosophy. The author claims that the existence of numerous little known insufficiently verified methods of therapy of this disease necessitates multicenter controlled studies for evaluation of the efficacy of the treatments. PMID- 10523956 TI - [Sinusotrabeculectomy with regulated filtration in the treatment of secondary glaucoma]. AB - A new method for surgical treatment of secondary glaucoma (neovascular, postuveal, traumatic, and aphakic) is described. The aims of the new method are as follows: regulation of ophthalmic tone in the postoperative period, pain relief, organ preservation, stabilization of the process, and creation of reliable routes of intraocular fluid discharge with formation of a stable diffuse functional pad. Seven patients (8 eyes) aged 13-70 years were operated on and followed up for up to 6 months. Preliminary results are presented. Positive shifts were observed in all cases. PMID- 10523957 TI - [Results of optic reconstructive interventions in children with ocular injuries]. AB - Optic reconstructive interventions were carried out in 36 children with ocular injuries aged 3-14 years. Perforating keratoplasty was combined with measures aimed at repair of the normal anatomic status of the eye and the transparency of its optic media. Microsurgical interventions were performed for penetrating corneal wounds in 27 cases, for penetrating corneoscleral injuries in 8, and for eye contusion in 1 case. The patients were followed up for 1 year after surgery. Taking in of donor cornea, time course of visual acuity, intra- and postoperative complications, and the risk factors of the graft failure were evaluated. Transparent healing of donor cornea was observed after 1 month in 94% cases and after 1 year in 60%. In the immediate postoperative period, vision improved in 32 patients, deteriorated in 1, and did not change in 3 patients. In 56% children visual acuity with correction of 0.05 diopters and higher was attained, while before the intervention the predominant (in 80% children) variant was pr. certae. Among the complications, the most incident were graft rejection crises and secondary glaucoma liable to conservative therapy. Vitreoretinal injury is one of the unfavorable factors fraught with the graft failure. Hence, perforating keratoplasty is an effective and obligatory surgical procedure ensuring sufficient visual rehabilitation in the majority of children with severe ocular injuries. PMID- 10523958 TI - [Organ of vision in preterm infant]. AB - Examinations of the organ of vision in 79 preterm infants showed lack of sensitivity of the conjunctiva and cornea, unstable precorneal membrane, and insufficient lacrimal production; these features may promote the development of diseases of the anterior segment of the eyeball. Decreased reaction to a light stimulus and lack of the pupil reaction to light are characteristic features of a preterm infant. The position of the baby lying in an incubator and poor transparency of ocular media impedes the ophthalmological examination and require special training of the physician. Pathological changes in the fundus oculi of a preterm baby should be regularly checked up by an oculist during the postnatal period. PMID- 10523959 TI - [Complex method for examining children with false myopia]. AB - The authors propose a complex method for examining children with false myopia. The complex includes routine methods for detecting and evaluating the stability of spasm, measurement of the relative accommodation reserve, ophthalmic tone, and anteroposterior axis of the eye. The method allows differentiation between false and true myopia, evaluates its degree, and helps prescribe etiopathogenetic treatment for ruling out the transfer of false myopia into true (axial). PMID- 10523961 TI - [Effect of superphonoelectrophoresis on chorioretinal dystrophy]. AB - The effect of a pharmacophysical method superphonophoresis on the course of chorioretinal dystrophy has been studied in 320 patients (640 eyes) with chorioretinal dystrophies treated by an ultrasonic UZT-104 device and a Nion galvanic device. The "superexposure" was phonophoresis of a bipolar drug 10% dimethylsulfoxide, creating the optimal conditions for penetration of the main agent through tissue barrier. The results (improvement of visual acuity, hemodynamics, trophics, shown by a higher linear rate of the blood flow in the orbital artery and middle cerebral arteries) persisted for up to 12 months, moreover, the rate of cellular and tissue metabolism increased (the time of adaptation to darkness decreased and color sensitivity increased). PMID- 10523960 TI - [Clinical and immunomorphological parallels in posttraumatic uveitis]. AB - Parallels between the clinical diagnosis, immunological parameters of the leukocyte migration inhibition test (LMIT) to eye tissue antigens (uveoretinal, lenticular, and retinal), and morphological picture are studied in patients with posttraumatic uveitis and consequences of grave penetrating injuries to the eye without uveitis symptoms. Cell sensitization to uveoretinal antigen is detected only in posttraumatic uveitis but not in consequences of injuries without uveitis. The authors come to a conclusion that only positive LMIT with uveoretinal or a combination of uveoretinal and lenticular antigens may be considered as an immunological validation of autoimmune posttraumatic uveitis. Further improvement of LMIT with purified fractions of uveoretinal antigens is needed. PMID- 10523962 TI - [Antioxidative activity of histochrome and some other drugs used in ophthalmology]. AB - The antioxidative activity (AOA) of histochrome (2,3,5,6,8-pentahydroxy-7-ethyl 1,4-naphthoquinone) and some other drugs with antioxidant properties used in ophthalmology has been studied in the hemoglobin--hydrogen peroxide--luminol system. The AOA of histochrome, ascorbate, dicinon, rutin, quercetin, and dihydroquercetin is comparable to that of trolox, the reference antioxidant in our studies. The drugs (in physiological concentrations) listed above are characterized by a high AOA. Cerebrolysin was 300 times less active than trolox, and emoxipin showed no antioxidant properties. Lacrimal AOA increased after a parabulbar injection of histochrome. Histochrome is believed to be a perspective antioxidant for the treatment of ocular diseases. PMID- 10523963 TI - [Time course of bioelectrical activity of retina in patients with pigmented abiotrophy treated by deoxinate]. AB - Bioelectrical retinal activity was assessed before and after a course of deoxinate therapy and in a blind placebo test in order to assess the efficacy of this new drug in the treatment of patients with pigmented retinal abiotrophy. Time course of electroretinograms during deoxinate therapy indicated a significant positive effect of the drug on retinal function as regards a variety of electrophysiological parameters in 80-94% patients. Improvement of electroretinography data was paralleled by improvement of visual acuity, extension of visual fields, and decrease of scotomas. Good effect was attained in patients with early and far advanced stages of pigmented retinal abiotrophy. Analysis of bioelectrical activity of the retina after a course of therapy with deoxinate and enkad showed a higher efficacy of the former drug in patients with pigmented retinal abiotrophy. PMID- 10523964 TI - [Effects of repeated courses of electrostimulation and psychophysiologic correction on visual system characteristics in patients with poor vision of different genesis]. AB - The effect of 5 courses of transcutaneous electrostimulation and of a combination of electrostimulation and psychophysiological correction of vision on the visual functions is assessed in 47 patients (90 eyes) with poor sigh of different genesis. Courses of 8-10-days were administered twice a year for 2.5-3 years. Cluster analysis of 13 objective and subjective parameters of vision permitted singling out patients responding to treatment well and poorly (groups 1 and 2). The absolute and differential electric activities of the central retina were increased during and between the courses in the first group while in the second group the shifts in the retina were negligible. Retinal changes are believed to reflect the general processes of adjustment of its activity to the levels needed for maintaining the integral visual functions, increased during treatment. PMID- 10523965 TI - [Vascular architectonics of eye and orbital space in color reflection of doppler spectrum energy]. AB - Super-sharp gray scale image and highly sensitive digital wide-band Doppler, three-dimensional visualization of vessels in the pulsed energy Doppler mode were used in examination of the posterior segment of the eye, optic nerve, and orbital space. Twelve healthy volunteers (24 eyes) aged 25-45 years were examined. The architectonics of the optic disk and the blood filling of the arterial Zinn Haller circle, reticular platelet, choroid, posterior long and posterior short ciliary arteries were studied. Vascular architectonics of the retrobulbar space was studied by the three-dimensional reconstruction in the energy Doppler mode and the borders of the vascular plexus round the optic nerve were defined. PMID- 10523966 TI - [Our method of functional rheography of eye]. AB - The technology of rheoophthalmogram obtaining and processing is modified by creating an automated rheographic complex (ARGC). The test for assessing the function of ocular vessels includes recording of the background rheoophthalmogram, local 10-min hypothermal exposure (+10 +/- 2 degrees C), and repeated recording of rheoophthalmogram 10 and 20 min after hypothermal exposure. The test helps determine the type of neurovascular reaction and the index of ocular vessel elasticity. The studies were carried out in 1050 subjects, 133 of them without ocular disease and 917 patients with myopia, initial open-angle glaucoma, retinal vein thrombosis, central serous chorioretinopathy, central atherosclerotic chorioretinopathy, diabetic retinopathy, toxic degeneration of the optic nerve, and contusions of the eye. PMID- 10523967 TI - [Treatment of eyeball and ocular appendages malignant tumors]. AB - Based on the results of her observations, the author demonstrates the advantages of organ-sparing treatment. PMID- 10523968 TI - [Comparative results evaluation of residual myopia and astigmatism correction after radial keratotomy by photorefraction keratectomy and laser specialized keratomileusis]. AB - The results of correction of residual myopia by photorefraction keratectomy (PRK) (51 eyes) and laser specialized keratomileusis (LASIK) (36 eyes) after radial keratotomy (RK) are compared. The patients were observed for up to 12 months. After PRK, 7.3% patients developed late fleur of the cornea, evaluated by at least 2 points. The incidence of fleur directly depended on the value of residual myopia. After LASIK none of the patients developed such fleur. The best visual acuity (0.5 and higher without correction) was attained in 70.73% after PRK and RK, vs. 100% after LASIK. The results of photorefraction operations and severity of residual myopia after RK correlated. In residual myopia of up to -3 diopters the results of correction by PRK and LASIK were virtually the same. In residual myopia higher than -3 diopters, LASIK is preferable. PMID- 10523969 TI - [Case of lagophthalmos treatment with an expander]. PMID- 10523970 TI - [Three cases with dirofilariasis of eyes]. PMID- 10523971 TI - [Role of fibrinolytic enzymes in corneal ulceration]. PMID- 10523972 TI - [Functional methods of examination in ophthalmology. Moscow, 1998]. PMID- 10523973 TI - [President of the Russian Academy of Medical Sciences V.I.Pokrovskii's opening address]. PMID- 10523974 TI - [Current trends in health of Russia's population]. PMID- 10523975 TI - [Updated technologies of epidemiological surveillance of infectious diseases]. PMID- 10523976 TI - [New research technologies in human ecology and environmental hygiene]. PMID- 10523977 TI - [Conception of healthy nutrition]. PMID- 10523978 TI - [New medical technologies in human health evaluation on the basis of functional systems theory]. PMID- 10523980 TI - [Fundamental sciences in surgery]. PMID- 10523981 TI - [Lipid distress syndrome in atherosclerosis obliterans]. PMID- 10523979 TI - [Current approaches to studying the pathogenesis of diseases]. PMID- 10523982 TI - [New technologies in cardiological care]. PMID- 10523983 TI - [Atherosclerosis: view on solution of this problem]. PMID- 10523984 TI - [Cardiovascular surgery at the turn of the XXI century : achievements and progress]. PMID- 10523985 TI - [Children's health in Russia: scientific and organizational priorities]. PMID- 10523986 TI - [New medical technologies in obstetrics, gynecology and neonatology]. PMID- 10523987 TI - [Future of laparoscopic surgery in gynecology]. PMID- 10523989 TI - [Ways of developing traumatology and orthopedics]. PMID- 10523990 TI - [New technologies in neurosurgery]. PMID- 10523991 TI - [Urology in the XXI century: prospects of development]. PMID- 10523988 TI - [Organ and tissue grafting: present and future]. PMID- 10523993 TI - [The outlook for the development and improvement of ambulatory polyclinic care]. AB - Out-patients medical care development represent one of the main tasks of the military medical service reforming. The essential effort focuses on a pre hospital stage of medical aid, i.e. forming-up of day hospitals, centers of specialist care for out-patients, new personnel policy and medics' training improvement. PMID- 10523992 TI - [The problems in and the means for improving ambulatory surgical care in the Armed Forces]. AB - The article presents some conclusions on ambulatory surgery in the Military Medical Academy and Surgery Centers in St. Petersburg resulting in higher efficiency of surgical aid and its availability for military beneficiaries. Organization frames and staffing principles for day-time ambulatory surgery stations. PMID- 10523994 TI - [The organizational aspects of medical support for the troops in armed conflicts]. AB - The theory of medical maintenance adopted for young military teachers to help them to organize the course of "Medical Maintenance" discipline within educational programmes of military schools. The article focuses on the main principles of troops medical maintenance and supply. PMID- 10523995 TI - [Epidural anesthesia in aortocoronary bypass]. PMID- 10523996 TI - [Experience in treating gunshot wounds of the maxillofacial area]. PMID- 10523998 TI - [The efficacy of treating patients with tunnel neuropathies under ambulatory conditions]. PMID- 10523997 TI - [The prognosis for the development of acute heart failure in patients with a large-focus myocardial infarct]. PMID- 10523999 TI - [The rehabilitation of servicemen after myocardial revascularization]. PMID- 10524001 TI - [Assessing the health of servicemen]. PMID- 10524000 TI - [Typhoid fever in periods of wars and armed conflicts]. AB - Study of military-epidemiological importance of typhoid-paratyphoid infections in military operations of XIX and XX centuries. Despite the research progress, and new means of special prevention, the problem has not been solved yet. In wars water becomes an important factor of typhoid spread in the troops and civil population, so water supply should be closely supervised by the civil authorities and military medics. PMID- 10524002 TI - [Ways to improve prophylactic work in aviation medicine]. PMID- 10524004 TI - [The automation of the management of a medical service for the armed forces abroad]. AB - Based on the contemporary information technologies the optimization of administration in the military medical service of leading industrial states has led to a considerable improvement of medical care in their armed Forces, especially on the units level. Telemedicine is being viewed as the most important factor in further development of military medical care. PMID- 10524003 TI - [The use of modern economic methods for the rational choice of drugs]. PMID- 10524005 TI - [N. S. Molchanov--military field therapist (on the centenary of his birth)]. PMID- 10524006 TI - [The 70th anniversary of the Department of Medical Service Organization and Tactics of the Military Medical Academy (1929-1999)]. PMID- 10524007 TI - [A half century of Solnechnogorsk Military Hospital]. PMID- 10524008 TI - [The 55th anniversary of the 25th Central Polyclinic of the Ministry of Defense of the Russian Federation]. PMID- 10524009 TI - [The healers of the city of "9 banners"]. PMID- 10524010 TI - High-dose irradiation: wholesomeness of food irradiated with doses above 10 kGy. Report of a Joint FAO/IAEA/WHO Study Group. AB - This report presents the recommendations of an international group of experts convened by the World Health Organization, in association with the Food and Agriculture Organization of the United Nations and the International Atomic Energy Agency to consider the implications of food irradiated to doses higher than those recommended in 1980 by the Joint Expert Committee on the Wholesomeness of Irradiated Food. Irradiation ensures the hygienic quality of food and extends shelf-life. The public perception of the safety of food irradiation has generally precluded its widespread use. However, current applications of food irradiation to doses over 10 kGy have been in the development of high-quality shelf-stable convenience foods for specific target groups such as immunosuppressed individuals and those under medical care, astronauts and outdoor enthusiasts. The Study Group reviewed data relating to the toxicological, nutritional, radiation chemical and physical aspects of food irradiated to doses above 10 kGy from a wide range and number of studies carried out over the last forty years. This report presents a comprehensive summary, along with references, of the effectiveness and safety of the irradiation process. It concludes that foods treated with doses greater than 10 kGy can be considered safe and nutritionally adequate when produced under established Good Manufacturing Practice. PMID- 10524011 TI - Development of an infection surveillance project for home healthcare. AB - Individual home care agencies must begin to take responsibility for gathering and analyzing data on infections that occur in the home. This is the only way to determine whether the infections are related to home care services or to other factors in the patient's environment. The Infection Surveillance Project was born out of the need for reliable and useful information on infection rates in home care. The need to be able to identify this information within individual companies and the industry as a whole takes on greater importance with managed care and accrediting bodies. PMID- 10524012 TI - The insight of a good manager. PMID- 10524013 TI - Establishing or enhancing your risk management program, Part II: The key elements of an effective risk management program. PMID- 10524015 TI - Management strategies for implementing OASIS. PMID- 10524014 TI - Making the transition from clinician to manager. AB - The entire program provides a beginning orientation for the transition into a manager's position. The program focuses on basic skills and topics as well as provides the participant resources for continuing development. It is anticipated that the thought processes and viewpoints of a manager emerge during the discussions of class content. As the program evolves, other topics may be important to an agency. The program can be adapted to suit specific groups of managers, such as those in specialty programs or those working in branch offices. Ultimately, managers benefit from the interaction with others who encounter similar problems or concerns. It is highly recommended that each new manager also receive a peer mentor within the agency to ease the transition. PMID- 10524016 TI - Compliance is job one. PMID- 10524017 TI - A quality perspective. PMID- 10524018 TI - New BSN graduates as home health nurses--why not?: an educator's perspective. PMID- 10524019 TI - Hiring new graduates: how one home health agency made the transition. PMID- 10524020 TI - A valuable message. PMID- 10524021 TI - Life's not fair... PMID- 10524022 TI - Unmet needs in the over-75s. PMID- 10524023 TI - Prevention is the key. PMID- 10524024 TI - Caring for the carers. PMID- 10524025 TI - Getting to grips with spirometry. PMID- 10524026 TI - Treatment solutions for nutritional anaemias. PMID- 10524027 TI - What the future holds for diabetes care. PMID- 10524029 TI - Treatments for osteoporosis in postmenopausal women. PMID- 10524028 TI - Hypertension: only the tip of the iceberg. PMID- 10524030 TI - How the community and hospital can work together. PMID- 10524031 TI - Life with venous leg ulcers: the story of one patient. PMID- 10524032 TI - Implementation of nurse prescribing in Scotland. PMID- 10524033 TI - Promises, promises, promises.... PMID- 10524034 TI - The misery of acne: a look at the treatment options. PMID- 10524035 TI - Handwashing in infection control. PMID- 10524036 TI - Management of COPD with oxygen therapy at home. PMID- 10524037 TI - Easing that itch ... treatments for lice and scabies. PMID- 10524038 TI - Cavity wound management. PMID- 10524039 TI - Crown Review II report: the issues facing prescribers. PMID- 10524040 TI - Adherence to standards of care and implications of body temperature measurement in trauma patients. AB - PURPOSE: To identify adherence to the standard that trauma patients have body temperature (T) recorded, range-of-temperature measurements, and the incidence of hypothermia recorded; and to examine the relationship between (T) and Injury Severity Score (ISS). METHODS: A retrospective review of the records of 60 trauma patients was conducted. FINDINGS: Forty percent of the patient records had temperatures recorded with values that ranged from (T) 87-100.6 degrees F; 33% of the patients had hypothermia as defined by a temperature of 96.6 degrees F or less. There appeared to be a significant inverse relationship between (T) and ISS. CONCLUSIONS: Based on the data, temperature was recorded in only 40% of the cases sampled. Adherence to the standard of measuring and recording a value was only intermittently followed. Nursing personnel should be educated to appreciate the potential for unsuspected hypothermia and to respond by following the standards of care. PMID- 10524041 TI - Paediatric head injuries: a literature review. PMID- 10524042 TI - Rehabilitation strategies for the tetraplegic patient in the trauma center. PMID- 10524043 TI - Caught between the rock and the hard place: proving the worth of nursing. PMID- 10524044 TI - Making it count: key factors to consider when assessing continuing professional educational offers. AB - Sustaining an essential level of specific knowledge, skills, and abilities for the competent practice of professional nursing in today's healthcare environment is imperative. Entry into the field of nursing first compels a commitment to lifelong education and lifelong learning--whether voluntary or mandatory. Today, educators most often enhance a practitioner's competency and professional accountability through a variety of continuing professional education offerings. This article offers specific suggestions that can help trauma nurses choose their learning ventures wisely and optimize their ongoing investment of time, effort, and money. PMID- 10524045 TI - Motor vehicle crashes and positive toxicology screens in adolescents and young adults. AB - The purpose of this study was to identify and describe the outcome of motor vehicle crashes for adolescents and young adults with positive toxicology screens. A retrospective design was used to collect data for 134 subjects ranging in age from 15 to 25 at an urban Level II trauma center. Outcomes related to sex, age, injury-severity score, length of stay, and hospital cost were analyzed using multiple regression, and the relationship was significant (p < .0001). Spearman's correlation analysis resulted in a significant relationship between survival outcome, injury severity, discharge, and hospital cost (p < .05). PMID- 10524046 TI - Advice for the trauma coordinator. PMID- 10524048 TI - Nursing assessment in adult trauma patients with nonoperative management of spleen and liver lacerations. AB - PURPOSE: To describe the clinical presentation of patients with blunt abdominal trauma undergoing nonoperative management of spleen or liver lacerations for identification of pertinent assessment findings indicative of the impending need for surgical intervention. METHODS: A retrospective study utilizing medical records for a 5 year period of adult blunt trauma patients with a diagnosis of spleen or liver laceration were reviewed. FINDINGS: Patients who failed nonoperative management of spleen or liver lacerations had statistically different measurements of heart rate, uncontrolled pain, skin color and temperature, numbers of units of blood received, urgent computed tomography scans, and serial hemoglobin levels. CONCLUSIONS: Nursing observations are crucial to the early identification of bleeding in patients with blunt abdominal trauma. PMID- 10524047 TI - "Troo, the Traumaroo" bicycle and playground safety program: a pilot study of kindergarten through second graders in the southwest. AB - "Troo, the Traumaroo" bicycle and playground safety education program created for kindergarten, first, and second grade students, was provided to a convenience sample of seven elementary schools in a Southwestern city. Favorable principal, classroom teacher, and school nurse evaluations indicated that the "Troo, the Traumaroo" program was successful in providing young children with bicycle and playground safety in a fun and entertaining way. Results of this pilot study indicated that kindergarten children pre-tested for bicycle safety knowledge prior to participating in the program, had significantly higher bicycle safety knowledge scores 30 days after participating in the program (p < .0001). PMID- 10524049 TI - Court of appeals overturns EMTALA decision fining physician $100,000 for inappropriate transfer. PMID- 10524051 TI - [Therapy and nursing interventions in venous leg ulcers]. PMID- 10524052 TI - [Care problem: dry skin]. PMID- 10524053 TI - [Better antibiotics therapy for patients with cystic fibrosis]. PMID- 10524050 TI - Trauma registrar training: integrating registry functions into the trauma program -Part I. AB - The University of Pennsylvania Health System Trauma Network views the registry staff as key members of the trauma team and the registry as a vital component to the overall success of our programs. Trauma Registry databases are increasingly complex and require highly skilled and knowledgeable personnel to perform the functions of the job. Although the background and experience level of each registry staff member may differ, this orientation and training process provides a consistent educational experience for all. This guideline ensures that all registry functions are being performed in a standardized and consistent manner. It sets the stage for validation of not only the data, but also an individual's performance relative to abstraction and computer entry. Providing the proper learning environment for registry staff has proven to be most beneficial for our network as we strive to develop and maintain trauma registry databases which have a strong foundation and provide valid, useful and current information. PMID- 10524054 TI - [Why results in the laboratory can not replace the dialog with the patient]. PMID- 10524055 TI - [The difficult disclosure]. PMID- 10524056 TI - [New aspects in the care of chronic wounds]. PMID- 10524057 TI - [Management of infections in chronic wounds. Hydroactive dressings instead of antiseptics?]. PMID- 10524058 TI - [Dismofix--the new cleanser assortment by BODE] [In Process Citation] PMID- 10524059 TI - [Partner for the nursing personnel]. PMID- 10524060 TI - [Ethics. The body]. PMID- 10524061 TI - [Federal legislation for the care of the elderly]. PMID- 10524062 TI - [Beautiful networking]. PMID- 10524063 TI - [Measures against sepsis]. PMID- 10524064 TI - [Care in acute stroke]. PMID- 10524065 TI - [Acute pulmonary embolism in surgical patients--a severe perioperative complication]. PMID- 10524066 TI - [Diseases that threaten our joints. When is the use of artificial joints necessary?]. PMID- 10524067 TI - [Breakthrough in the development of materials. Focus hip implantation]. PMID- 10524068 TI - [Expectations from hormone substitution therapy]. PMID- 10524069 TI - [Can you help me] PMID- 10524070 TI - [Slimness craze or eating disorder? When the soul is hungry and the family is helpless]. PMID- 10524072 TI - [From sayings by the Buddha. The blind men and the elephant] [In Process Citation] PMID- 10524071 TI - [Prenatal medicine and ethics]. PMID- 10524073 TI - [Prototype of caring]. PMID- 10524074 TI - [Nursing visit--a catchword and its transfer into practice]. PMID- 10524075 TI - [Ambulatory care of HIV-infected patients and patients with AIDS]. PMID- 10524077 TI - [Primary and secondary causes of constipation]. PMID- 10524076 TI - [Contribution of psychopharmacotherapy to the control of post-traumatic stress disorders]. PMID- 10524078 TI - [Isomol--an innovation in the therapy of constipation]. PMID- 10524079 TI - [GP11b/111a receptor antagonists. Better prognoses in acute coronary syndromes]. PMID- 10524080 TI - [Enteral nutrition from the viewpoint of nutrition science: more than a supply of energy]. PMID- 10524081 TI - [20 years of Fresubin: a milestone in enteral nutrition]. PMID- 10524082 TI - [Insuman--from the physician's point of view]. PMID- 10524084 TI - [Report of experiences in the use of Clipper 9661 by the Firm 3M. Shaving of the area to be operated]. PMID- 10524083 TI - [Danger from anesthetic gases?]. PMID- 10524085 TI - [The molecule as a symbol for drugs]. PMID- 10524086 TI - [Decubitus ulcers--without end]. PMID- 10524087 TI - [Decisions about ending measures that extend the dying process]. PMID- 10524088 TI - [What is new about diabetes mellitus, type 2?]. PMID- 10524089 TI - [Portrait of a drug. Insulin lispro]. PMID- 10524091 TI - [Of values and valuation. The sources of conscience in our existence]. PMID- 10524090 TI - [Making peace with the inner child]. PMID- 10524092 TI - [Reasons. Moving into an old age home]. PMID- 10524093 TI - [The best health care system in the world--but ... limits of capacity in the delivery of health care]. PMID- 10524094 TI - Central venous lines. PMID- 10524095 TI - Health for all by 2010? PMID- 10524096 TI - Making it all better. PMID- 10524097 TI - A culture of learning. PMID- 10524098 TI - Putting patients first. PMID- 10524099 TI - A new sisterhood. PMID- 10524100 TI - Learning from life. PMID- 10524101 TI - Confidentially speaking. PMID- 10524102 TI - Don't forget nurse teachers. PMID- 10524103 TI - Full complement. PMID- 10524104 TI - Modernizing mental health services. PMID- 10524105 TI - Don't hang up: use of the telephone by people with communication difficulties. AB - Joan Murphy describes a research project funded by the Scottish Office Home and Health Department, that investigated telephone use by people with communication difficulties. PMID- 10524107 TI - Pulse oximetry. AB - Pulse oximetry provides a non-invasive means to measure pulse rate and haemoglobin saturation. Philip Woodrow describes the nurse's role in its use. PMID- 10524106 TI - Meeting the needs of homeless people: the St John Ambulance mobile service. AB - St John Ambulance has a well established history of providing services to the community. This article describes the organisation's first venture in meeting the needs of homeless people, focusing on the implications for qualified nurses. PMID- 10524108 TI - Developing better ways to share bad news. PMID- 10524109 TI - Nurse prescribing--implications in practice. PMID- 10524111 TI - An evaluative approach to selecting mattresses. AB - Gillian Arblaster discusses the process undertaken by the Walsgrave Hospitals NHS Trust to evaluate dynamic pressure-relieving/reducing mattresses before deciding which type to rent or purchase. The study was undertaken over three years ago and, therefore, the products discussed in this article may have been superceded by newer ones. Structural and educational changes might also have taken place within the companies. John Timmons, a tissue viability nurse specialist at Monklands Hospital, Airdrie, evaluates the process and outcomes of this small study. PMID- 10524110 TI - Preventing pressure sores in the seated patient. PMID- 10524113 TI - Get on board. PMID- 10524114 TI - Practice nursing. PMID- 10524112 TI - Managing eczema and dermatitis. PMID- 10524115 TI - Administration of medicines--1. PMID- 10524116 TI - Should nursing unions 'grass' on their members if they make a mistake? PMID- 10524117 TI - Capital's property boom making London a no-go zone for nurses, experts warn. PMID- 10524118 TI - What is a CNO? PMID- 10524119 TI - Time is money for nurse with MS. Interview by Esther Leach. PMID- 10524120 TI - The International Council of Nurses is in need of a major rethink. PMID- 10524121 TI - Go for assistance not resistance. PMID- 10524122 TI - Nurse heroes of the century. PMID- 10524123 TI - Return ticket. PMID- 10524124 TI - Nurses can do it. PMID- 10524126 TI - Nursing abroad. Straight talk. PMID- 10524125 TI - Act on equality. PMID- 10524127 TI - Mental health. Lullaby of Broadmoor. PMID- 10524128 TI - Face to face. Interview by Eileen Fursland. PMID- 10524130 TI - The diminishing role of nurses in hands-on care. PMID- 10524129 TI - Are people facing the facts on the dangers that smoking posed to their and their children's health. PMID- 10524131 TI - Reducing health risks in ethnic communities. PMID- 10524132 TI - Cooperate to innovate. PMID- 10524133 TI - First aid series. Drowning. PMID- 10524134 TI - Wound care. In on a limb. AB - In the light of nurses' increasing role in the assessment and diagnosis of leg ulcers, this article looks at leg ulcer management, focusing on increased aetiological factors, improved assessment techniques and treatment. PMID- 10524135 TI - Wound care. Arguments over iodine. PMID- 10524136 TI - Wound care. Pressure gauge. AB - This article describes the development of a nursing pressure sore manual and presents the results of a risk assessment and point prevalence survey. PMID- 10524138 TI - Nurse consultants. PMID- 10524137 TI - Administration of medicines--2. PMID- 10524139 TI - Where's the beef? PMID- 10524140 TI - Lessons in life. PMID- 10524141 TI - Scots ambition. PMID- 10524142 TI - Starting from scratch. Interview by Clare Harvey. PMID- 10524143 TI - Poaching the world. PMID- 10524144 TI - What is the Public Interest Disclosure Act? PMID- 10524145 TI - The government checks out supermarkets to see what family-friendly policies it can borrow for the NHS. PMID- 10524146 TI - Nowhere in the world has market-led health care been made to work. PMID- 10524147 TI - Putting the public interest first. PMID- 10524148 TI - Madness and the millennium. PMID- 10524149 TI - Not a belly laugh. Interview by Rebecca Coombes. PMID- 10524150 TI - A date with destiny. PMID- 10524151 TI - Community nursing. Where the heart is. PMID- 10524152 TI - Boxed in. PMID- 10524153 TI - Nursing abroad. Odyssey in the outback. PMID- 10524154 TI - Assessment tool promotes continence after childbirth. AB - This article looks at how midwives, health visitors and continence advisers are working together in Sandwell in the West Midlands to promote women's health. By acknowledging the evidence that certain factors associated with childbirth can predispose women to incontinence, a risk-assessment tool has been developed providing a process for women to achieve continence health postnatally. PMID- 10524155 TI - Lessons from the experience of using a video camera for research. AB - This paper discusses the practical and theoretical issues associated with the use of video cameras in a field setting to collect data of nurse-patient communication. In particular, the advantages and difficulties of filming participants are addressed and strategies put forward for overcoming potential problems. This paper aims to provide a useful insight into the use of these methods for other researchers. PMID- 10524156 TI - Infection control. Dazed and confused. AB - This research project aims to uncover the practical concerns of health care staff on a hospital ward while attempting to implement isolation precaution guidelines for patients with Clostridium difficile-associated diarrhoea (CDAD) and methicillin-resistant Staphylococcus aureus (MRSA). The project is still in progress so this article describes the research methods used and some preliminary findings. PMID- 10524157 TI - Infection control. A clean sheet. PMID- 10524158 TI - Infection control. Gunning for the big three. PMID- 10524159 TI - Flexibility in learning. PMID- 10524160 TI - Surgery during pregnancy. AB - The anatomic and physiologic changes of pregnancy complicate a woman's response to elective or emergency surgical intervention. Alterations related to perioperative positioning, fluid volume replacement, effects of medicinal and anesthetic agents, and maternal/fetal assessment are discussed. Emotional and ethical/legal concerns are considered. A postoperative assessment guideline is presented. PMID- 10524161 TI - The obese patient as a surgical risk. AB - Obesity has become a serious problem in the United States. The increasing prevalence of obesity makes the likelihood of clinicians caring for these individuals high. Several considerations for the preoperative care of these patients include appropriate assessment, particularly of the cardiopulmonary systems, and a thorough clinical examination. Intraoperative concerns include appropriate equipment, medication, positioning, and cardiopulmonary monitoring. Postoperative care for the obese patient requires special concern regarding oxygenation and wound healing. PMID- 10524162 TI - Perioperative considerations for patients with musculoskeletal and neuromuscular disorders. AB - This article first presents a brief description of musculoskeletal disease, then the anesthetic drugs that can trigger distress, and finally recommendations for perioperative nursing care based on pathophysiology and preservation of strength and function. The administration of a variety of anesthetic agents can exacerbate symptoms or result in unpredictable untoward effects on patients with musculoskeletal disease. Neuromuscular diseases result in diminished muscle strength. These deviations in structure and function must be considered when drafting a successful plan of care. PMID- 10524163 TI - Cultural considerations in perioperative nursing. AB - This article provides a brief overview of cultural diversity and its significance to health care. LEARN, an illness explanatory model, and its application to a case study of a minority patient's surgical experience are presented to increase surgical nurses' awareness of cultural illness beliefs and treatment expectations. PMID- 10524164 TI - Noncardiac surgery in individuals with coronary heart disease. AB - Atherosclerosis is a pathological condition of the coronary arteries clinically manifested as cardiovascular disease, the major cause of death in the industrialized world. Coronary heart disease (CHD) and cerebrovascular disease (CVD), both of which are atherosclerotic diseases, cause more death, disability, and economic loss in the United States than any other disease. PMID- 10524165 TI - Surgery and smoking. AB - The patient who smokes presents a unique challenge to the operating room nurse. The various components of tobacco impact in a number of important ways on the cardiovascular and pulmonary system. Wound healing also is affected by smoking. It is vital that the nurse recognize the complications that may be caused by smoking. PMID- 10524166 TI - Evidence-based practice and perioperative nursing. AB - The implementation of evidence-based practice in perioperative nursing holds promise of improving quality of care and client outcomes. Several factors within health care have precipitated an emphasis on evidence-based practice. The use of research results in clinical decisions is recommended as the basis of nursing practice of the future. To assist with development of evidence-based practice in nursing, basic steps of the process are presented. In addition, strategies for locating existing evidence-based practice guidelines and resources are described. Perioperative nurse researchers and practice leaders should move this issue into a top priority for the specialty. PMID- 10524167 TI - [Public health. The disquieting numbers in chronic pain]. PMID- 10524168 TI - [Public health. The government plan for the battle against pain]. PMID- 10524169 TI - [Professional activities. Pain, obligation and responsibility]. PMID- 10524170 TI - [Medical strategy. Use of morphine derivatives in chronic pain]. PMID- 10524171 TI - [Anthropology. But, after all, what is pain?]. PMID- 10524172 TI - [Nursing management. And if the nursing care were psychotherapy?]. PMID- 10524173 TI - [Pain within the framework of the nurse's proper role]. PMID- 10524174 TI - [Public health. Principles and limits of caring for patients with pain at home]. PMID- 10524176 TI - [Perspectives. Being able to refer to a pattern for decisions]. PMID- 10524175 TI - [Education. "Auvergne without pain"]. PMID- 10524177 TI - [Restoring capability to the patient]. PMID- 10524178 TI - [Accidents of exposure to blood]. PMID- 10524179 TI - [A synergistic intervention in the service of the patient]. PMID- 10524180 TI - [Patient education. Development of an educational project]. PMID- 10524181 TI - [The nurse's responsibility. 2. Personal responsibility of a public agent]. PMID- 10524182 TI - [What to do in case of cardio-respiratory arrest]. PMID- 10524183 TI - [Attention to the integrity of the skin. 1]. PMID- 10524185 TI - [Professional news. The National Network of Hospital Documentalists, a network to discover] [In Process Citation] PMID- 10524184 TI - [Interview on professional federations] [In Process Citation] PMID- 10524187 TI - [Rehabilitation of psychiatric nursing] [In Process Citation] PMID- 10524186 TI - [Professional news. A new ruling for a "unique" field of action] [In Process Citation] PMID- 10524188 TI - [Stages of scarring]. PMID- 10524189 TI - [Surgical and non-surgical wound debridement]. PMID- 10524190 TI - [For each stage of the wound its own dressing]. PMID- 10524191 TI - [Pain and decubitus ulcers in the aged]. PMID- 10524192 TI - DNA binding, protein interaction and differential expression of the human germ cell nuclear factor. AB - The mouse germ cell nuclear factor (mGCNF) is an orphan nuclear receptor implicated in diverse biological processes, including gametogenesis, embryonic development and embryonal carcinoma cell differentiation. We have examined the binding and regulation of the human orthologue, hGCNF, expressed in the teratocarcinoma-derived cell line NTera-2/clone D1 (NT2/D1). Binding of GCNF to the direct repeat of the sequence -AGGTCA- (DR-0) is conserved in mammalia. The formation of interspecies dimers of the in vitro synthesized proteins suggests that cellular GCNF binding is mediated by homodimers. Both the mouse and the human protein bind in concert with cellular factors to DNA. Treatment of NT2/D1 cells with all-trans retinoic acid (atRA) is accompanied first by an up regulation followed later by a down-regulation of hGCNF and its mRNA. Temporary up-regulation in NT2/D1 cells after treatment with atRA suggests that hGCNF is important for human neural determination and differentiation. PMID- 10524193 TI - p53 is a rate-limiting factor in the repair of higher-order DNA structure. AB - The product of the p53 tumor suppressor gene has been implicated in safeguarding genomic stability by transactivating genes involved in cell cycle arrest, repair of DNA damage or induction of apoptosis. Several properties of p53 suggest that it might be directly involved in DNA repair processes. Eukaryotic DNA is highly organized in supercoiled loops anchored to the nuclear matrix. This organization is very important for cell function and survival, suggesting that repair of DNA damage must include both, the integrity of the double helix and the complex DNA topology. In this work, we studied the kinetics and efficiency of higher-order DNA structure repair in cells with normal and reduced levels of p53, and present evidence suggesting that p53 may be involved in the stabilization and/or repair of higher-order DNA structure. PMID- 10524194 TI - The cartilage-derived, C-type lectin (CLECSF1): structure of the gene and chromosomal location. AB - Cartilage is a tissue that is primarily extracellular matrix, the bulk of which consists of proteoglycan aggregates constrained within a collagen framework. Candidate components that organize the extracellular assembly of the matrix consist of collagens, proteoglycans and multimeric glycoproteins. We describe the human gene structure of a potential organizing factor, a cartilage-derived member of the C-type lectin superfamily (CLECSF1; C-type lectin superfamily) related to the serum protein, tetranectin. We show by Northern analysis that this protein is restricted to cartilage and locate the gene on chromosome 16q23. We have characterized 10.9 kb of sequence upstream of the first exon. Similarly to human tetranectin, there are three exons. The residues that are conserved between CLECSF1 and tetranectin suggest that the cartilage-derived protein forms a trimeric structure similar to that of tetranectin, with three N-terminal alpha helical domains aggregating through hydrophobic faces. The globular, C-terminal domain that has been shown to bind carbohydrate in some members of the family and plasminogen in tetranectin, is likely to have a similar overall structure to that of tetranectin. PMID- 10524195 TI - Carotenoid hydroxylase from Haematococcus pluvialis: cDNA sequence, regulation and functional complementation. AB - A cDNA homologous to beta-carotene hydroxylase from Arabidopsis thaliana was isolated from the green alga Haematococcus pluvialis. The predicted amino acid sequence for this enzyme shows homology to the three known plant beta-carotene hydroxylases from Arabidopsis thaliana and from Capsicum annuum (38% identity) and to prokaryote carotenoid hydroxylases (32-34% identities). Heterologous complementation using E. coli strains which were genetically engineered to produce carotenoids indicated that the H. pluvialis beta-carotene hydroxylase was able to catalyse not only the conversion of beta-carotene to zeaxanthin but also the conversion of canthaxanthin to astaxanthin. Furthermore, Northern blot analysis revealed increased beta-carotene hydroxylase mRNA steady state levels after induction of astaxanthin biosynthesis. In accordance with the latter results, it is proposed that the carotenoid hydroxylase characterized in the present publication is involved in the biosynthesis of astaxanthin during cyst cell formation of H. pluvialis. PMID- 10524196 TI - Genomic structure and promoter analysis of the ecto-phosphodiesterase I gene (PDNP3) expressed in glial cells. AB - PDNP (phosphodiesterase I/nucleotide pyrophosphatase) is one of a series of ectoenzymes that are involved in hydrolysis of extracellular nucleotides. PDNP possesses ATPase (EC 3.6.1.3) and ATP pyrophosphatase (EC 3.6.1.8) activities. Mammalian PDNP consists of three closely related family proteins (PDNP1, -2, and 3), and they are expressed in different cell types and at different developmental stages. Rat PDNP3 is expressed in a subset of immature glial cells and in the alimentary tract. Human PDNP3 is expressed in glioma cells, prostate, and uterus, but not in the alimentary tract. We have cloned genomic DNA containing the whole coding region of the human PDNP3 gene and determined its exon-intron structure. The human PDNP3 gene spans over 60 kb and is organized into 25 exons and 24 introns. We determined the nucleotide sequence of the 5'-flanking region of human and rat PDNP3 genes. The upstream region of both species lacks a canonical TATA box and contains a putative binding site for CCAAT enhancer-binding proteins near the transcription start site. Promoter activity analysis of the 5'-flanking region revealed that the sequence around the CCAAT box is required for its transcriptional activity in 9L rat glioma cells. A gel shift assay demonstrated that 9L nuclear extract contains proteins that bind to this region. PMID- 10524197 TI - Transcription of actin, cyclophilin and glyceraldehyde phosphate dehydrogenase genes: tissue- and treatment-specificity. AB - Studies involving RNA transcription, in varying biological systems, usually necessitate a term of transcriptional reference. Traditionally, the transcription of the gene of interest was compared to a constitutively expressed 'control' gene. Run-on transcription analysis was undertaken to evaluate and compare the transcription of three frequently used 'control genes' (beta-actin, cyclophilin and glyceraldehyde-3-phosphate dehydrogenase), in nine rat tissues. Similarities, but also clear and highly significant differences, were found in the transcription profiles of these three genes. There was significantly greater transcription for uterine glyceraldehyde-phosphate dehydrogenase compared to all other tissues tested, while both cyclophilin and glyceraldehyde-phosphate dehydrogenase were significantly elevated in the adrenal cortex. Upon cholinergic agonist treatment, both beta-actin and glyceraldehyde-phosphate dehydrogenase RNA expression were greatly induced in the adrenal medulla (41- and 94-fold, respectively), while cyclophilin transcription was not altered. In another treatment paradigm, surgical ovariectomy, only uterine glyceraldehyde-phosphate dehydrogenase transcription was significantly reduced. While, all three of these genes are assumed to be constitutively expressed throughout the body and hence used as normalization controls, the current study questions these accepted terms of reference. As cyclophilin transcription was not affected in both treatment paradigms, it should be considered more seriously as a RNA normalization control. PMID- 10524199 TI - Myb and Ets transcription factors cooperate at the myb-inducible promoter of the tom-1 gene. AB - Transformation of myeloid cells by the retroviral oncogene v-myb is thought to be caused by deregulated expression of specific cellular genes that act as targets of v-Myb in myeloid cells. Recently, we have identified the chicken tom-1 gene as a direct target for v-Myb. tom-1 has two promoters, only one of which (the tom-1A promoter) is activated by v-Myb. Here, we show that v-Myb activates the tom-1A promoter by cooperating with Ets-2, a member of the Ets transcription factor family. Interestingly, we find that the ability of v-Myb to cooperate with Ets proteins differs from that of its non-oncogenic cellular counterpart c-Myb. c-Myb cooperates with Ets-1 and Ets-2, whereas v-Myb only cooperates with Ets-2. Truncation of the N-terminus of c-Myb, which is known to activate the oncogenic potential of c-Myb, specifically abrogates the ability of the protein to cooperate with Ets-1. Our findings, therefore, reveal a novel function for the N terminus of c-Myb and raise the possibility that oncogenic activation of c-Myb is linked to the loss of cooperation between Myb and c-Ets-1. PMID- 10524198 TI - Differential regulation of the human insulin gene transcription by GG1 and GG2 elements with GG- and C1-binding factors. AB - Using a human growth hormone reporter system, the introduced mutations in GG1 alone or both GG elements of GG1 and GG2 in the human insulin promoter abolished 94 or 96% of the beta-cell-specific transcriptional activity in a pancreatic islet beta-cell line of MIN6, while the mutations in GG2 or its total deletion abolished 85 or 86% of the transcriptional activity. When linked to the thymidine kinase promoter, mutations in GG1 or both GG elements abolished 74% of the transcriptional activity in MIN6 cells, while the mutations in GG2 or its total deletion abolished 55 or 54%. In the electrophoretic mobility shift assay (EMSA), one nuclear factor was shown to interact with two GG elements, and another C1 binding factor with GG1 and C1. The differential effects of deletions or selective mutations in the GG2 or GG1 sequence in the oligonucleotide probes on the binding activity of GG- or C1-binding factors in EMSA proved the requirement of both GG1 and GG2 or both GG1 and C1, respectively, for the transaction of these two factors. The molecular size of the GG-binding factor was estimated about 30 kDa. Based on these, we conclude that two GG elements contribute, with GG1 more critically than GG2, to the beta-cell-specific transcription of the human insulin gene through transaction with the GG- and C1-binding factors. PMID- 10524201 TI - Isolation and characterization of the distal promoter region of mouse Cbfa1. AB - Cbfa1 is an essential transcription factor for bone formation, and as such little is known about the region responsible for the transcriptional regulation of this gene. Here we report the determination of the transcription start sites, isolation and partial characterization of distal promoter region of this gene. Three transcription start sites were identified by the 5'-Cap site method, recently invented for rapid examination of the 5'-end of genes of interest. A reporter construct containing 1.8 kb of 5' of transcription start sites had approximately 25-fold more luciferase activity than the promoter-less vector in osteoblastic cell lines. Deletion analysis of the reporter construct demonstrated that the minimal region to express promoter activity lies between bp -168 and 99, taking the most downstream transcription start site as +1. By Northern blot analysis, mRNA expression from the distal promoter was detected in the differentiated osteoblastic cell lines, UMR-106, ROS17/2.8 and MC3T3-E1, but not in cell lines of immature phenotype or originated from other organs. Luciferase activity was strongest in UMR-106 and ROS17/2.8, and weakest in COS-1 and HepG2, which are cell lines originating from other organs, corresponding to the level of mRNA expression. These results demonstrated that the distal promoter region examined here is important for tissue- and cell-type-specific gene expression of Cbfa1. PMID- 10524200 TI - Human phosducin-like protein (hPhLP) messenger RNA stability is regulated by cis acting instability elements present in the 3'-untranslated region. AB - Phosducin (Pd) and phosducin-like protein (PhLP) have been shown to regulate G protein signaling by binding G beta gamma subunits. To better define the function and regulation of PhLP, and to begin to investigate its potential role in human pathophysiological states, we have cloned the human PhLP (hPhLP) cDNA. The hPhLP shows 92% identity with the rat PhLP (rPhLP). However, unlike the rPhLP, no evidence of hPhLP isoforms were detected in the human tissues investigated. Additionally, unlike the rPhLP, alternative polyadenylation sites were detected in hPhLP cDNA clones which corresponded with two distinct mRNA transcripts, 1.2 kb and 3.1 kb, respectively. Interestingly, the predominantly expressed long transcript contains multiple AU-rich elements (AREs) in its 3'-untranslated region (3'-UTR) which have been shown to correlate with rapid mRNA turnover and translational control. This study shows that the hPhLP AREs are functional both in vitro and in vivo, with the long transcript exhibiting a much shorter mRNA half-life. We also demonstrate that subcloning of either the full-length 3'-UTR or the ARE-rich region of the long transcript immediately following the stop codon of luciferase reporter gene confers instability to the luciferase mRNA and results in a ninefold reduction of luciferase activity in the cell types investigated. Taken together, these findings suggest that the AREs present in the long hPhLP mRNA may play a critical role in the regulation of hPhLP gene expression. PMID- 10524203 TI - Primary structure, gene expression and chromosomal mapping of rodent homologs of the MEN1 tumor suppressor gene. AB - Mutations of the MEN1 tumor suppressor gene cause the multiple endocrine neoplasia type 1 (MEN1) syndrome in humans, and they are involved in a variety of sporadic human endocrine tumors. We here characterize the MEN1 gene homologs of the mouse and rat. cDNA was isolated from a mouse phage library, and two alternative MEN1 mRNA transcripts containing variant 5' untranslated regions were identified by RT-PCR in several mouse and rat tissues. When compared to the human molecule, mouse and rat MEN1 (611 and 610 amino acids, respectively) show an overall identity of 96.5% and 97.0% at the protein level, delimiting four conservational domains (A-D). Mouse and rat MEN1 mRNA, as studied by template calibrated quantitative RT-PCR, is non-exclusively expressed in hematopoietic and endocrine cells, with similar expression patterns found in fetal and adult tissues. Fluorescent in situ hybridization maps the single murine MEN1 locus to chromosome 19, region B. No MEN1 gene mutations were identified in endocrine islet tumor cell lines RIN 5AH (rat) and NIT-1 (mouse) as compared to wild type cDNA. Our data define mouse and rat MEN1 as widely expressed and highly conserved homologs of the human MEN1 tumor suppressor gene whose role in biology and endocrine tumorigenesis is due for experimental study. PMID- 10524202 TI - Distamycin A selectively inhibits Acanthamoeba RNA synthesis and differentiation. AB - The effects of distamycin A on Acanthamoeba transcription, growth and differentiation were determined. Distamycin A inhibits transcription both in vitro and in vivo and can displace from DNA the transcription activator TATA binding protein promoter binding factor (TPBF). Inhibition in vivo is surprisingly selective for large rRNA precursors, 5S rRNA, profilin, S adenosylmethionine synthetase, and extendin. Transcription from the TATA binding protein (TBP), TPBF, protein disulfide isomerase, tubulin and RNA polymerase II large subunit genes is only slightly inhibited. Moreover the rate of 5S rRNA transcription eventually recovers and exceeds that of untreated cells, while profilin transcription remains inhibited. Distamycin A inhibition is accompanied by a complex pattern of alterations to steady state levels of mRNAs. Actin, profilin and S-adenosylmethionine synthetase mRNAs are degraded, whereas mRNA encoding TBP is increased slightly in abundance. Transcription inhibition is accompanied by cessation of growth and severe morphological changes to Acanthamoeba, which are consistent with loss of production of mRNA encoding cytoskeletal proteins. Distamycin A also prevents starvation-induced differentiation of Acanthamoeba, in part due to complete prevention of cellulose production and cell wall formation. PMID- 10524204 TI - Biochemical analysis of mouse FKBP60, a novel member of the FKPB family. AB - We have identified mouse and human FKBP60, a new member of the FKBP gene family. FKBP60 shares strongest homology with FKBP65 and SMAP. FKBP60 contains a hydrophobic signal peptide at the N-terminus, 4 peptidyl-prolyl cis/trans isomerase (PPIase) domains and an endoplasmic reticulum retention motif (HDEL) at the C-terminus. Immunodetection of HA-tagged FKBP60 in NIH-3T3 cells suggests that FKBP60 is segregated to the endoplasmic reticulum. Northern blot analysis shows that FKBP60 is predominantly expressed in heart, skeletal muscle, lung, liver and kidney. With N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide as a substrate, recombinant GST-FKBP60 is shown to accelerate effectively the isomerization of the peptidyl-prolyl bond. This isomerization activity is inhibited by FK506. mFKBP60 binds Ca2+ in vitro, presumably by its C-terminal EF-hand Ca2+ binding motif, and is phosphorylated in vivo. hFKBP60 has been mapped to 7p12 and/or 7p14 by fluorescence in situ hybridization (FISH). PMID- 10524205 TI - Analysis of cis-acting sequences and trans-acting factors regulating the interleukin-3 response element of the DUB-1 gene. AB - The murine DUB-1 gene is a hematopoietic-specific, immediate-early gene that encodes a growth-regulatory deubiquitinating enzyme. DUB-1 contains an IL-3 inducible enhancer element that is activated in a JAK2-dependent, STAT5 independent manner. In this study, we have further characterized this novel IL-3 response element. Transcriptional reporter assays in Ba/F3 cells revealed that two AP-1 sites, a GATA motif, and an Ets site are required for induction of DUB-1 enhancer activity. Gel shift assays indicated that IL-3 activates the binding of an AP-1 complex containing JunD to the AP-1 sites and the binding of another protein complex to the Ets motif. The latter complex was not detectable in Ba/F3 cells stably transfected with a dominant-negative mutant of JAK2. As previously shown, these cells do not express DUB-1 mRNA or protein. Furthermore, we demonstrated that GATA-1 constitutively binds to the DUB-1 enhancer element. The involvement of GATA-1 may be important for the hematopoietic-restricted expression pattern of DUB-1. This combination of inducible and constitutive elements of the DUB-1 enhancer appears to account for the unique STAT-independent expression characteristics of DUB-1. PMID- 10524206 TI - Alternative splicing of CD45 pre-mRNA is uniquely obedient to conditions in lymphoid cells. AB - The leucocyte common antigen (LCA or CD45) consists of various isoforms generated by alternative splicing of variable exons 4, 5 and 6 (or A, B and C). To follow splicing behaviour in different cell types we developed a human CD45 mini-gene and analysed its expression in transfected cell lines and transgenic mouse tissues. In Cos-1, HeLa and 3T3 cells we found distinct expression patterns which could only be modulated slightly by protein synthesis inhibitors but not by variation in culture conditions like pH, serum concentration and cell density, or by stimulation with phorbol ester (TPA). In all non-lymphoid transgenic tissues the default splicing pattern (CD45R0) was found, while the expression profile in lymphoid cells, where all eight isoforms are present, mimics that of the endogenous mouse LCA gene products. Next, to examine the factors involved in alternative exon use we analysed the expression pattern of members of the family of SR proteins, well known splicing regulators with arginine/serine-rich (R/S) domains. Cell lines expressed variable levels of SRp75, SRp30 and SRp20 and constant amounts of SRp40. Mouse tissues expressed large amounts of SRp75, SRp55 and SRp40, additional expression of SRp30s and SRp20 was restricted to lymphoid tissues. Therefore, SRp30 and SRp20 may contribute to forming the appropriate cellular conditions for alternative use of CD45 exons 4-6 in the haematopoietic compartment. PMID- 10524207 TI - Structure and chromosomal location of mouse and human CD52 genes. AB - Human CD52 (CAMPATH-1 antigen) is an abundant surface molecule on lymphocytes and a favoured target for lymphoma therapy and immunosuppression. It comprises a small glycosylphosphatidylinositol (GPI) anchored peptide to which a large carbohydrate moiety is attached. Structurally similar proteins include the proposed mouse homologue, B7 antigen (B7-Ag; not to be confused with the CD28 ligand), and human and mouse CD24. Sequence similarities between CD52 and B7-Ag precursors are concentrated over the signal peptides and the sequences cleaved during GPI attachment. While the short mature peptides are not apparently homologous, the N-linked glycosylation site is retained in both. We describe similarities in exon-intron organisation, syntenic chromosome positions (human CD52, 1p36; mouse B7-Ag, chromosome 4, between Dsil and D4Nds16) and sequence homology in the promoter regions which strongly suggests that B7-Ag is the mouse homologue of CD52. The structure of these genes is also similar to that of mouse CD24, suggesting a common ancestor. Promoter activities and transcription start sites were also analysed. These results suggest that human CD52 and mouse B7-Ag gene expressions are controlled by TATA-less promoters. PMID- 10524208 TI - The expression of human H2A-H2B histone gene pairs is regulated by multiple sequence elements in their joint promoters. AB - The majority of human H2A and H2B histone genes are organized as gene pairs: 14 H2A-H2B gene pairs, one solitary H2A gene and three solitary H2B genes have been described. Two of the H2A genes and two of the H2B genes arranged within gene pairs are pseudogenes. The gene pairs are organized with divergent transcriptional orientation, and the coding regions of the respective H2A and H2B genes are separated by about 320 nucleotide pairs that form overlapping promoter regions. Comparison of promoters of H2A-H2B gene pairs has previously shown that these belong to two different groups (groups I and II) which are characterized by specific patterns of conserved sequence elements. We have constructed a reporter gene vector that allows the simultaneous analysis of both genes regulated by the divergent promoters belonging to group I or II, respectively. Firefly-luciferase and beta-galactosidase genes were taken as reporter genes. Site directed mutagenesis performed at individual promoter elements revealed that individual sequence elements within both groups of promoters functionally depend on each other and may contribute to a coordinate expression of paired H2A and H2B genes through assembly of their joint promoter into a mutually dependent promoter complex. Group II promoters are characterized by the presence of an E2F binding site upstream of the H2A gene-proximal TATA box. Immediately upstream of the E2F element, we have identified a highly conserved octanucleotide CACAGCTT (RT-1) that exists in all human group II H2A-H2B gene promoters. Protein binding studies at the RT-1 element indicate factor binding to this sequence. Site directed mutagenesis indicates that both the E2F element and the RT-1 motif are essential for full promoter activity. PMID- 10524209 TI - Cloning and functional characterization of the bovine endothelin-converting enzyme-1a promoter. AB - Endothelin-converting enzyme-1 (ECE-1) mRNA is expressed in three isoforms, termed a, b, and c, originating from alternative promoters. In cultured bovine aortic endothelial cells, we detected mRNA isoform expression of ECE-1a and ECE 1b/c, respectively. Investigating transcriptional mechanisms of bovine endothelial ECE-1a expression in more detail, we identified multiple transcription start sites localized 120-415 nucleotides upstream from the presumptive translation start codon by RNase protection assay and 5' RACE. Using luciferase reporter gene assays we found that 1.4 kb of the 5' untranslated region showed strong promoter activity in endothelial cells. Sequence analysis revealed 71% overall homology of the bovine ECE-1a promoter with its human homologue. The proximal 680 base pair promoter region was shown to contain cis elements that are sufficient for basal and serum-induced transcriptional activation. PMID- 10524210 TI - Identification of the second tonicity-responsive enhancer for the betaine transporter (BGT1) gene. AB - When certain cells are exposed to a hypertonic solution, transcription of the BGT1 gene is markedly increased. The ensuing rise in betaine transport leads to cellular accumulation of betaine that protects the cells from the stress of hypertonicity. We have previously identified a tonicity-responsive enhancer (TonE1) in the 5' flanking region of the BGT1 gene. It was recognized, however, that full activation of transcription requires additional sequence upstream from the TonE1. Now we report that there is another TonE (named TonE2) 72 base pairs upstream from the TonE1. TonE1 and TonE2 act synergistically to stimulate transcription of BGT1 in response to hypertonicity. PMID- 10524211 TI - Isolation, molecular characterization, and tissue-specific expression of ECP-51 and ECP-54 (TIP49), two homologous, interacting erythroid cytosolic proteins. AB - We isolated two proteins, ECP-51 and ECP-54, from human erythrocyte cytosol by affinity chromatography using a peptide of the integral membrane protein stomatin as bait. Partial amino acid sequence information obtained by microsequencing allowed us to clone the respective cDNAs. Analysis of the nucleotide sequences revealed that ECP-51 and ECP-54 are homologous (44.2% amino acid identity) and contain ATP-binding sites. ECP-54 was identified as TIP49/RUVBL1/NMP238, which is a component of a large nuclear protein complex, possibly the RNA polymerase II holoenzyme; ECP-51 is a novel protein. Using the two-hybrid system, we showed that these proteins interact with each other. The interaction of ECP-51 and ECP 54 with the stomatin peptide and the localization to the nucleus and cytoplasm suggest an additional function for these proteins as chaperone components. PMID- 10524212 TI - Isolation and characterisation of a cDNA encoding the precursor for a novel member of the acyl-CoA dehydrogenase gene family. AB - A gene encoding the precursor for a novel member of the human acyl-CoA dehydrogenase (ACD) gene family has been isolated which maps to human chromosome 11q25. The cDNA contains an open reading frame of 1248 nucleotides encoding a predicted 415-amino-acid peptide, and shares considerable sequence similarity with other members of the ACD family. PMID- 10524213 TI - The ABC-exporter genes involved in the lipase secretion are clustered with the genes for lipase, alkaline protease, and serine protease homologues in Pseudomonas fluorescens no. 33. AB - In Pseudomonas fluorescens no. 33, the lipase gene is clustered with the genes for alkaline protease, AprDEF exporter, and two homologue proteins of Serratia serine proteases (pspA and pspB). Secretion of the lipase and alkaline protease through AprDEF was shown in the Escherichia coli cells. Interestingly, the E. coli cells carrying the pspA gene secreted PspA to the media AprDEF independently. PMID- 10524214 TI - Identification and expression of a cDNA for human hydroxypyruvate/glyoxylate reductase. AB - The isolation and expression of a human liver cDNA encoding a 40-kDa protein with glyoxylate and hydroxypyruvate reductase activities is described. The cDNA (GLXR) is 1235 bp and consists of a predicted open reading frame of 987 bp with a 225-bp 3'-untranslated region. The 328-amino acid protein has partial sequence similarity to hydroxypyruvate and glyoxylate reductases from a variety of plant and microbial species. PMID- 10524215 TI - Microsomal GST-I: genomic organization, expression, and alternative splicing of the human gene. AB - In this paper we report the genomic organization of the human microsomal GST-I gene. This gene spans 18 kb, and contains seven exons. Sequences that encode the 155 amino acid open reading frame are present in Exons II, III, IV, the 5' untranslated region is present in Exons Ia, Ib, Ic, Id, and II, and the 3' untranslated region is present in Exon IV. Exons Ia, Ib, Ic, Id, and III are alternatively spliced to generate at least six different mGST-I transcripts. The results of EST and PCR analysis show that most mGST-I transcripts terminate within Exon Ib, and primer extension analysis shows these transcripts initiate at three major sites located at 79, 81, and 88 nucleotides upstream of the ATG initiation codon. Sequences surrounding the putative initiation sites are G-C rich, and several Sp1 consensus binding sites were identified. Northern analysis shows that the human GST-I gene is preferentially expressed as a 1.0 kb transcript in liver, and in several other tissues. Finally, a comparison of the mGST-I and PIG12 sequences with those of FLAP, LTC4 synthase, mGST-II, and mGST III suggests that these proteins are the related products of a dispersed microsomal GST gene superfamily. PMID- 10524216 TI - Molecular cloning and expression of human neurochondrin-1 and -2. AB - Human neurochondrins have been cloned from a brain cDNA library. The human neurochondrin-1 and -2 predict leucine-rich (15.8 and 15.9%) proteins of 729 and 712 amino acid residues, with molecular weights of 78.9 and 77.2 kDa, respectively. The deduced amino acid sequence indicates 98% identity among human, mouse and rat species. Northern analysis indicates that about 4 kb human neurochondrin mRNAs are abundant in the fetal and the adult brain. PMID- 10524217 TI - Molecular cloning and nucleotide sequence of a gene encoding a cotton palmitoyl acyl carrier protein thioesterase. AB - A cotton genomic clone containing a 17.4-kb DNA segment was found to encompass a palmitoyl-acyl carrier protein (ACP) thioesterase (Fat B1) gene. The gene spans 3.6 kb with six exons and five introns, and is apparently the first plant FatB acyl-ACP thioesterase gene to be completely sequenced. The six exons are identical in nucleotide sequence to the open reading frame of the corresponding cDNA, and would encode a preprotein of 413 amino acids. The preprotein can clearly be identified as a FatB acyl-ACP thioesterase from its similarity to the deduced amino acid sequences of other FatB thioesterase preproteins. A 5' flanking region of 914 bp was sequenced, with the potential TATA basal promoter 324 bp upstream from the ATG initiation codon. The 5'-flanking sequence also has a putative CAAT box and two presumptive basic region helixloop-helix (bHLH) elements with the consensus motif CANNTG (termed an E box), implicated as being a positive regulatory element in seed-specific gene expression. PMID- 10524218 TI - Genomic cloning, structure, and regulatory elements of the 1 alpha, 25(OH)2D3 down-regulated gene for cyclic AMP-dependent protein kinase inhibitor. AB - The cyclic AMP-dependent protein kinase inhibitor (PKI) mRNA and protein are negatively and tissue-specifically regulated in the kidney by 1 alpha, 25(OH)2D3. A 17-kb PKI clone, isolated from a chick genomic library, revealed that the PKI gene consists of two exons separated by a 4.5-kb intron. A 411-bp upstream region (constituting 93 bp upstream and 318 bp downstream from the transcriptional start site) containing a putative negative VDRE (nVDRE) fused to the luciferase gene was used for transient transfections of primary cultures of chick kidney cells. Luciferase activity was significantly down-regulated in response to 1 alpha, 25(OH)2D3. This result suggests that the promoter region containing the putative nVDRE plays a pivotal role in the negative regulation of PKI gene transcription. PMID- 10524219 TI - The human gene coding for HCN2, a pacemaker channel of the heart. AB - Hyperpolarization-activated, cyclic nucleotide-gated (HCN) channels, underlying 'pacemaker' currents (I(f)/Ih), are involved in pacemaker activity of cardiac sinoatrial node myocytes and central neurons. Several cDNAs deriving from four different genes were recently identified which code for channels characterized by six transmembrane domains and a cyclic nucleotide binding domain. We report here the identification of the human HCN2 gene and show that its functional expression in a human kidney cell line generates a current with properties similar to the native pacemaker f-channel of the heart. The hHCN2 gene maps to the telomeric region of chromosome 19, band p13.3. This is the first identification of a genetic locus coding for an HCN channel. PMID- 10524220 TI - U82, a novel snoRNA identified from the fifth intron of human and mouse nucleolin gene. AB - A novel snoRNA, designated as U82, was found from the sequence analysis of the 5th intron of human and mouse nucleolin gene. The snoRNA U82 has characteristic boxes C, D and D' and 11 nucleotides (nt) antisense complementarity to the 18S rRNA. Presumably U82 functions as a guide in the methylation of residue A1678 in human 18S rRNA. Northern blot analysis with various oligodeoxynucleotide probes showed that human and mouse U82 is expressed as RNA variants with length of 70 (+/- 1) and 67 (+/- 1) nt in HeLa and mouse C127 cells. Most probably, the 70 nt variant of U82 is encoded by nucleolin gene 5th intron. The 67 nt variant of U82 could be a transcript of another gene, the genomic locus of which remains unknown. PMID- 10524221 TI - Cloning of ClC-2 chloride channel from murine duodenum and its presence in CFTR knockout mice. AB - We report the cloning of a murine ClC-2 chloride channel cDNA from duodenal epithelium by reverse transcriptase-polymerase chain reaction (RT-PCR) using degenerate primers and by rapid amplification of cDNA ends (RACE)-PCR. Other than CFTR, this represents the first cloned chloride channel from intact intestine. The ClC-2 cDNA predicts encoding of a 908 amino acid polypeptide with a calculated M(r) of 99,373. The amino acid sequence of the murine ClC-2 chloride channel is over 94% identical to the ClC-2 chloride channel proteins of other species. Of interest is the finding that the ClC-2 mRNA is expressed about the same level in duodena from both CFTR knockout and wild-type mice. This is in keeping with the suggestion that ClC-2 might be a therapeutic target in cystic fibrosis. PMID- 10524222 TI - Molecular cloning, heterologous expression, and characterization of a novel member of CYP2A in the Syrian hamster. AB - The cDNA clone coding for a novel cytochrome P-450 2A subfamily member (CYP2A16) was isolated from a Syrian hamster liver cDNA library. The deduced amino acid sequence of CYP2A16 showed more than 90% identity with those of rat CYP2A3 and mouse CYP2A4/5. The catalytic activity of CYP2A16 was determined by transient expression of its cDNA in transfected COS7 cells and CYP2A16 was found to have the testosterone 2 beta-, 15 alpha-, and 15 beta-hydroxylases, coumarin 7 hydroxylase, and ethoxycoumarin O-deethylase activities. These enzymatic characteristics of CYP2A16 are different from those of other Syrian hamster CYP2A subfamily members, CYP2A8 and CYP2A9. Northern blot analysis showed that CYP2A16 was expressed in kidney and lung while most of the other CYP2A subfamily members have been reported to be expressed in liver and olfactory. These observations indicated that the Syrian hamster CYP2A16 had unique properties compared with those of other CYP2A subfamily members. PMID- 10524223 TI - Structure of Tetrahymena CCT theta gene and its expression under colchicine treatment. AB - We report here the cloning and the characterization of the Tetrahymena pyriformis chaperonin-containing-TCP1 theta gene (TpCCT theta), an orthologue of the mouse chaperonin gene CCT theta. TpCCT theta gene is interrupted by eight introns, ranging in size between 91 and 419 nucleotides, and encodes a protein consisting of 540 amino acid residues (59.1 kDa), with a putative pI of 5.73. The amino acid sequence of TpCCT theta reveals 39.4-46.0% identity with the sequences of Candida albicans and mouse CCT theta subunits and 28.0-32.6% identity with the other TpCCT subunits known so far. We have studied the expression of this gene in exponentially growing Tetrahymena cells and in cells treated with colchicine for different times. The steady-state levels of CCT theta mRNA rapidly decrease in the first 30 min of colchicine treatment. Interestingly, treatment for subsequent 60 min gives expression levels higher than those found in exponentially growing cells. PMID- 10524224 TI - Aberrations of ammonia metabolism in ornithine carbamoyltransferase-deficient spf ash mice and their prevention by treatment with urea cycle intermediate amino acids and an ornithine aminotransferase inactivator. AB - Sparse fur with abnormal skin and hair (spf-ash) mice are deficient in ornithine carbamoyltransferase (OCT) activity, but their OCT protein is kinetically normal. We administered ammonium chloride to spf-ash mice, in order to analyze ammonia metabolism and to find a rationale for the therapy of OCT deficiency. Ammonia concentration in the liver of spf-ash mice increased to a level much higher than in the control. Ammonium chloride injection caused an increase in ornithine (Orn) 5 min after injection and an increase in the sum of Orn, citrulline (Cit) and arginine (Arg) for at least 15 min in the liver of control mice, but no increase in Orn, Cit and Arg in the liver of spf-ash mice. Treatment of spf-ash mice with Arg 5-20 min prior to the injection of ammonium chloride kept the hepatic ammonia concentration at a level comparable to that without the load. A significant reciprocal relationship between ammonia and Orn concentrations in the liver of spf-ash mice 5 min after an ammonium chloride load with or without Arg strongly suggests that ammonia disposal is dependent on the supply of Orn. In spf-ash mice loaded with tryptone as a nitrogen source, Arg supplementation showed a dramatic decrease in urinary orotic acid excretion in a dose-dependent manner. Similar effects were observed with Cit and Orn at the same dose, and a long-lasting effect with an ornithine aminotransferase inactivator, 5-(fluoromethyl)ornithine, at a much lower dose. The rate of urea formation in liver perfused with ammonium chloride was lower in spf-ash mice than in controls, but with the addition of Orn to the medium it increased to a similar level in control and spf-ash mice. These results indicate that OCT is not saturated with Orn in vivo under physiological conditions and that the administration or enrichment of the urea cycle intermediate amino acids enhances the OCT reaction so that the ammonia metabolism of OCT-deficient spf-ash mice is at least partially normalized. PMID- 10524225 TI - Induction of TIMP-1 expression in rat hepatic stellate cells and hepatocytes: a new role for homocysteine in liver fibrosis. AB - Elevated plasma levels of homocysteine have been shown to interfere with normal cell function in a variety of tissues and organs, such as the vascular wall and the liver. However, the molecular mechanisms behind homocysteine effects are not completely understood. In order to better characterize the cellular effects of homocysteine, we have searched for changes in gene expression induced by this amino acid. Our results show that homocysteine is able to induce the expression and synthesis of the tissue inhibitor of metalloproteinases-1 (TIMP-1) in a variety of cell types ranging from vascular smooth muscle cells to hepatocytes, HepG2 cells and hepatic stellate cells. In this latter cell type, homocysteine also stimulated alpha 1(I) procollagen mRNA expression. TIMP-1 induction by homocysteine appears to be mediated by its thiol group. Additionally, we demonstrate that homocysteine is able to promote activating protein-1 (AP-1) binding activity, which has been shown to be critical for TIMP-1 induction. Our findings suggest that homocysteine may alter extracellular matrix homeostasis on diverse tissular backgrounds besides the vascular wall. The liver could be considered as another target for such action of homocysteine. Consequently, the elevated plasma levels of this amino acid found in different pathological or nutritional circumstances may cooperate with other agents, such as ethanol, in the onset of liver fibrosis. PMID- 10524226 TI - Role of neuronal and endothelial nitric oxide synthase in nitric oxide generation in the brain following cerebral ischemia. AB - Nitric oxide (NO) plays an important role in the pathogenesis of neuronal injury during cerebral ischemia. The endothelial and neuronal isoforms of nitric oxide synthase (eNOS, nNOS) generate NO, but NO generation from these two isoforms can have opposing roles in the process of ischemic injury. While increased NO production from nNOS in neurons can cause neuronal injury, endothelial NO production from eNOS can decrease ischemic injury by inducing vasodilation. However, the relative magnitude and time course of NO generation from each isoform during cerebral ischemia has not been previously determined. Therefore, electron paramagnetic resonance spectroscopy was applied to directly detect NO in the brain of mice in the basal state and following global cerebral ischemia induced by cardiac arrest. The relative amount of NO derived from eNOS and nNOS was accessed using transgenic eNOS(-/-) or nNOS(-/-) mice and matched wild-type control mice. NO was trapped using Fe(II)-diethyldithiocarbamate. In wild-type mice, only small NO signals were seen prior to ischemia, but after 10 to 20 min of ischemia the signals increased more than 4-fold. This NO generation was inhibited more than 70% by NOS inhibition. In either nNOS(-/-) or eNOS(-/-) mice before ischemia, NO generation was decreased about 50% compared to that in wild type mice. Following the onset of ischemia a rapid increase in NO occurred in nNOS(-/-) mice peaking after only 10 min. The production of NO in the eNOS(-/-) mice paralleled that in the wild type with a progressive increase over 20 min, suggesting progressive accumulation of NO from nNOS following the onset of ischemia. NOS activity measurements demonstrated that eNOS(-/-) and nNOS(-/-) brains had 90% and < 10%, respectively, of the activity measured in wild type. Thus, while eNOS contributes only a fraction of total brain NOS activity, during the early minutes of cerebral ischemia prominent NO generation from this isoform occurs, confirming its importance in modulating the process of ischemic injury. PMID- 10524227 TI - Biochemical, genetic and immunoblot analyses of 17 patients with an isolated cytochrome c oxidase deficiency. AB - Mitochondrial respiratory chain defects involving cytochrome c oxidase (COX) are found in a clinically heterogeneous group of diseases, yet the molecular basis of these disorders have been determined in only a limited number of cases. Here, we report the clinical, biochemical and molecular findings in 17 patients who all had isolated COX deficiency and expressed the defect in cultured skin fibroblasts. Immunoblot analysis of mitochondrial fractions with nine subunit specific monoclonal antibodies revealed that in most patients, including in a patient with a novel mutation in the SURF1 gene, steady-state levels of all investigated COX subunits were decreased. Distinct subunit expression patterns were found, however, in different patients. The severity of the enzymatic defect matched the decrease in immunoreactive material in these patients, suggesting that the remnant enzyme activity reflects the amount of remaining holo-enzyme. Four patients presented with a clear defect of COX activity but had near normal levels of COX subunits. An increased affinity for cytochrome c was observed in one of these patients. Our findings indicate a genetic heterogeneity of COX deficiencies and are suggestive of a prominent involvement of nuclear genes acting on the assembly and maintenance of cytochrome c oxidase. PMID- 10524228 TI - Role of protein kinase C in cyclic AMP-mediated suppression of T-lymphocyte activation following burn injury. AB - Major burn injury induces T-lymphocyte dysfunction. Previous studies suggest that prostaglandin (PG) E2, which is elevated post-burn, is the causative factor via a cyclic AMP-dependent process. The present study was conducted to elucidate the mechanism by which cAMP induces T-lymphocyte dysfunction following burn injury. Splenocytes were isolated from mice 7 days after receiving a scald burn covering 25% of their total body surface or sham procedure. ConA-induced proliferation by splenocytes from burned mice was significantly suppressed. Macrophage depletion of the splenocyte cultures abrogated the suppression. Concanavalin A-stimulated proliferation by macrophage-depleted splenocytes was suppressed by PGE2 and dibutyryl cAMP in both groups. The IC50 of these cAMP-elevating agents, however, was approximately 100-fold lower for cells from burned mice, indicating an increased sensitivity to cAMP. PGE2 did not suppress PMA/Ca2+ ionophore-induced T lymphocyte activation. Addition of PMA to ConA-stimulated cultures prevented the suppression of proliferative responses by PGE2, whereas Ca2+ ionophore had no effect. Thus, the suppression of T-lymphocyte activation following burn injury is macrophage-dependent, related to an increased sensitivity to cAMP and due to an uncoupling of cell surface receptors from protein kinase C activation. PMID- 10524229 TI - beta-Trace protein in human cerebrospinal fluid: a diagnostic marker for N glycosylation defects in brain. AB - As carbohydrate-deficient glycoprotein syndromes (CDGS) are multisystemic disorders with impaired central nervous function in nearly all cases, we tested isoforms of beta-trace protein (beta TP), a 'brain-type' glycosylated protein in cerebrospinal fluid (CSF) of nine patients with the characteristic CDGS type I pattern of serum transferrin. Whereas the serum transferrin pattern did not discriminate between the various subtypes of CDGS type I (CDGS type Ia, type Ic, and patients with unknown defect), beta TP isoforms of CDGS type Ia patients differed from that of the other CDGS type I patients. The percentage of abnormal beta TP isoforms correlated with the severity of the neurological symptoms. Furthermore, two patients are described, who illustrate that abnormal protein N glycosylation can occur restricted to either the 'peripheral' serum or the central nervous system compartment. This is the first report presenting evidence for an N-glycosylation defect restricted to the brain. Testing beta TP isoforms is a useful tool to detect protein N-glycosylation disorders in the central nervous system. PMID- 10524230 TI - Interferon regulatory factors: the next generation. AB - Interferons are a large family of multifunctional secreted proteins involved in antiviral defense, cell growth regulation and immune activation. Viral infection induces transcription of multiple IFN genes, a response that is in part mediated by the interferon regulatory factors (IRFs). The initially characterized members IRF-1 and IRF-2 are now part of a growing family of transcriptional regulators that has expanded to nine members. The functions of the IRFs have also expanded to include distinct roles in biological processes such as pathogen response, cytokine signaling, cell growth regulation and hematopoietic development. The aim of this review is to provide an update on the novel discoveries in the area of IRF transcription factors and the important roles of the new generation of IRFs- particularly IRF-3, IRF-4 and IRF-7. PMID- 10524231 TI - A novel sperm-specific hypomethylation sequence is a demethylation hotspot in human hepatocellular carcinomas. AB - Certain human DNA regions are strikingly undermethylated at CpG sites in sperm compared to adult somatic tissues. These sperm-specific hypomethylation sequences are thought to function early in embryogenesis or gametogenesis. By using the restriction landmark genomic scanning (RLGS) cloning method, we have isolated a novel sperm-specific hypomethylation sequence, the status of which changes during spermatogenesis, embryonal growth and differentiation. This sequence is a part of a new 'NotI repeat' consisting of a 1.4 kb repetitive unit sequence named DE-1. The sequence is GC-rich and has high homology to a CpG DNA clone that was isolated by a methyl CpG protein binding column, indicating that it was normally highly methylated. We investigated the methylation status of this sequence. In the normal genome the sequence was methylated, but in the human hepatocellular carcinoma (HCC) genome, the target sequence was demethylated at the cytosine residue of the CpG dinucleotides with high frequency (75% in the previous study). These data suggest that this regional DNA hypomethylation may play a role in both cell differentiation and hepatocarcinogenesis. PMID- 10524232 TI - Cloning of human cDNAs for Apg-1 and Apg-2, members of the Hsp110 family, and chromosomal assignment of their genes. AB - In mice, the Hsp110/SSE family is composed of the heat shock protein (Hsp)110/105, Apg-1 and Apg-2. In humans, however, only the Hsp110/105 homolog has been identified as a member, and two cDNAs, Hsp70RY and HS24/p52, potentially encoding proteins structurally similar to, but smaller than, mouse Apg-2 have been reported. To clarify the membership of Hsp110 family in humans, we isolated Apg-1 and Apg-2 cDNAs from a human testis cDNA library. The human Apg-1 was 100% and 91.8% identical in length and amino acid (aa) sequence, respectively, to mouse Apg-1. Human Apg-2 was one aa shorter than and 95.5% identical in sequence to mouse Apg-2. In ECV304, human endothelial cells Apg-1 but not Apg-2 transcripts were induced in 2 h by a temperature shift from 32 degrees C to 39 degrees C. As found in mice, the response was stronger than that to a 37-42 degrees C shift. The human Apg-1 and Apg-2 genes were mapped to the chromosomal loci 4q28 and 5q23.3-q31.1, respectively, by fluorescence in-situ hybridization. We isolated cDNA and genomic clones encompassing the region critical for the difference between Apg-2 and HS24/p52. Although the primer sets used were derived from the sequences common to both cDNAs, all cDNA and genomic clones corresponded to Apg-2. Using a similar approach, the relationship between Apg-2 and Hsp70RY was assessed, and no clone corresponding to Hsp70RY was obtained. These results demonstrated that the Hsp110 family consists of at least three members, Apg-1, Apg-2 and Hsp110 in humans as well as in mice. The significance of HS24/p52 and Hsp70RY cDNAs previously reported remains to be determined. PMID- 10524234 TI - A GC-rich sequence within the 5' untranslated region of human basonuclin mRNA inhibits its translation. AB - By the method of RNase protection, the 5' ends of the basonuclin mRNA were mapped to four sites distributed over 400 bases of the genomic sequence, a result implying four different basonuclin transcripts within the cell. Despite the heterogeneity at the 5' end, all four basonuclin mRNA shared the same translation initiation codon. However, only two transcripts contained, in their 5' untranslated region (UTR), a GC-rich sequence of approx. 180 bases. The ability of this GC-rich sequence to form a large and stable secondary structure was suggested by experimental results from primer extension, RNase resistance, and computer analysis of the sequence. In vitro study showed that translation of basonuclin RNA containing this putative structure could not be initiated efficiently from the first two AUGs, whereas those that lacked it could. PMID- 10524233 TI - The exon-intron organization of the prohormone convertase PC2 gene from the insect Lucilia cuprina. AB - Prohormone or proprotein convertases are members of the subtilisin family of serine proteases. They are involved in the activation of precursor molecules by endoproteolytic cleavage at basic amino acid residues. Among the different members of this prohormone convertase family, the prohormone convertase 2 (PC2) is almost exclusively expressed in endocrine and neuroendocrine tissues and plays an important role in the endoproteolytic processing of prohormones. Here we describe the exon-intron organization of the PC2 gene from the insect Lucilia cuprina by characterization of PCR-amplified genomic DNA fragments. The insect PC2 gene contains 12 exons with an estimated size of over 14.5 kb. The exon sizes range from 38 bp to > 448 bp. All identified intron-exon boundaries are consistent with the GT-AG-rule. A comparison of the genomic structures of the thus far known prohormone convertase genes with that of the insect PC2 gene revealed a conservation of the positions of most introns interrupting the exons coding for the amino-terminal and catalytic domains. This conservation is consistent with the suggestion of a common evolutionary origin for the prohormone convertase gene family. PMID- 10524235 TI - Identification of new sigma K-dependent promoters using an in vitro transcription system derived from Bacillus subtilis. AB - In Bacillus subtilis, the genes that depend on sigma K-RNA polymerase for their transcription are expressed in the mother cell compartment at later stages of sporulation. More than a dozen genes belonging to the sigma K regulon have been identified. Here I describe the identification of two additional promoters under the control of sigma K-RNA polymerase. Using a set of histidine-tagged RNA polymerases prepared from cells harvested at various times during the course of growth and sporulation (Fujita, M., Sadaie, Y., 1998. Gene 221, 185-190), transcription initiated from putative promoter sequences on a number of DNA fragments, as inferred from genome sequencing, was examined in vitro. One of these showed sigma K-dependent transcription. For further characterization of transcription initiated from this site, in vitro transcription analysis was performed using RNA polymerase holoenzyme reconstituted from purified sigma K and core RNA polymerase. Two sigma K-dependent promoters, yfhP P1 and yfhP P2, separated by a distance of about 15 bp, were thereby identified. These promoters are located immediately upstream of the yfhP gene that encodes a protein of unknown function consisting of 327 amino acids residues. The promoter strength, the rate of open complex formation and the RNA polymerase binding affinity were examined for these two promoters in comparison with other known sigma K-dependent promoters, gerE and cotD. The promoter strength displayed was in the order of gerE > cotD > yfhP P2 > yfhP P1. PMID- 10524237 TI - The identification of seven metalloproteinase-disintegrin (ADAM) genes from genomic libraries. AB - Metalloproteinase-disintegrins (ADAMs) are membrane-spanning multi-domain proteins containing a zinc metalloproteinase domain and a disintegrin domain which may serve as an integrin ligand. Based on a conserved sequence within the disintegrin domain, GE(E/Q)CDCG, seven genes were isolated from a human genomic library. Two of these genes lack introns and show testis-specific expression (ADAM20 and ADAM21), while the other two genes contain introns (ADAM22 and ADAM23) and are expressed predominantly in the brain. In addition, three pseudogenes were isolated; one of which evolved from ADAM21. Human chromosomal mapping indicated that ADAM22 and ADAM23 mapped to chromosome 7q21 and 2q33, respectively, while the three pseudogenes 1-2, 3-3, and 1-32 mapped to chromosome 14q24.1, 8p23, and 14q24.1, respectively. An ancestral analysis of all known ADAMs indicates that the zinc-binding motif in the catalytic domain arose once in a common ancestor and was lost by those members lacking this motif. PMID- 10524236 TI - Five different genes, Eif4a1, Cd68, Supl15h, Sox15 and Fxr2h, are clustered in a 40 kb region of mouse chromosome 11. AB - We characterized a region of the mouse genome disrupted by integration of a gene trap (GT) vector in ES cells. On 5' rapid amplification of cDNA ends analysis of the fusion transcripts containing the GT vector, we identified the eukaryotic protein synthesis initiation factor 4A1 gene (Eif4a1) as a promoter-trapped gene. Plasmid rescue was used to show that the other end of the integrated vector disrupted the murine homolog of the human fragile X mental retardation syndrome related protein 2 gene (Fxr2h). Structural analysis of P1 clones, isolated from the wild-type mouse genome by PCR with Eif4a1-specific primers, indicated that the integration of the GT vector was accompanied by the deletion of about 35 kb of genomic DNA and that the disrupted region also included three genes, Cd68, Supl15h and Sox15, the latter two of which are transcribed in opposite directions with overlapping 3' ends. These five different genes at least, Eif4a1, Cd68, Supl15h, Sox15 and Fxr2h, are clustered in a 40 kb region. The chromosomal location of this region was mapped by means of interspecific backcross panel DNAs to the central part of mouse chromosome 11, exhibiting a known region of synteny with human chromosome 17. PMID- 10524238 TI - An initiator element and a proximal cis-acting sequence are essential for transcriptional activation of the complement factor I (CFI) gene. AB - Human complement factor I (CFI) is a serine protease which regulates the complement system by inactivation of C3b and C4b in the presence of appropriate cofactors. In this study, we have analyzed the mechanism controlling the constitutive transcriptional activation of the CFI gene. Using deletion analysis and transient CAT expression assays, we have mapped the minimal promoter to the region located between -46 and +160 bp relative to the major transcription start point (tsp), and also shown that cis-acting elements both upstream and downstream of the tsp are important for promoter activity. A silencer element was also found between -71 and -46 bp. The sequence surrounding the tsp was related to the mouse terminal deoxynucleotidyltransferase initiator element (Inr) and point mutations in this sequence, from -3 to +4, drastically reduced CFI promoter activity. Mutations in a -9 to -5 bp CTGAA sequence immediately upstream of the tsp also reduced promoter activity. Gel mobility shift analysis demonstrated the binding of nuclear factors to a CTGAA repeat located at -9 to -5 and +101 to +105. Our results suggest that CFI promoter contains a functional Inr element that is essential for promoter activity, and the interactions of the CTGAA element located between -9 and +5 with nuclear factor(s) may be part of the machinery required for CFI Inr-dependent transcription. PMID- 10524239 TI - A unique bFGF-responsive transcriptional element. AB - The mitogen-regulated protein/proliferin (mrp/plf) genes encode closely related proteins that stimulate cell proliferation and angiogenesis. Basic fibroblast growth factor (bFGF) increases mrp/plf mRNA and protein production by 3T3 cells. Although the three cloned mrp/plf gene promoters are over 97% identical, only mrp3 is transcriptionally activated by bFGF. A series of truncated mrp3 promoter sequences were tested to determine the minimal promoter sequence necessary for bFGF-responsive transcription. Within the minimal bFGF-responsive mrp3 promoter fragment, a putative FGF-regulatory element (FRE) was identified. Nuclear factors that bind the FRE are present in 3T3 cells. When present upstream of a thymidine kinase basal promoter, the FRE exhibits high transcriptional activity and responds to bFGF. Thus, the FRE is a strong transcriptional element that is regulated by bFGF and that may participate in regulating the mrp3 gene and perhaps other FGF-regulated genes. PMID- 10524240 TI - Molecular characterisation of the Arabidopsis SBP-box genes. AB - The Arabidopsis thaliana SPL gene family represents a group of structurally diverse genes encoding putative transcription factors found apparently only in plants. The distinguishing characteristic of the SPL gene family is the SBP-box encoding a conserved protein domain of 76 amino acids in length, the SBP-domain, which is responsible for the interaction with DNA. We present here characterisation of 12 members of the SPL gene family. These genes show highly diverse genomic organisations and are found scattered over the Arabidopsis genome. Some SPL genes are constitutively expressed, while transcriptional activity of others is under developmental control. Based on phylogenetic reconstruction, gene structure and expression patterns, they can be divided into subfamilies. In addition to the Arabidopsis SPL genes, we isolated and determined the sequences of three SBP-box genes from Antirrhinum majus and seven from Zea mays. PMID- 10524241 TI - Characterization of the rat, mouse, and human genes of growth/differentiation factor-15/macrophage inhibiting cytokine-1 (GDF-15/MIC-1). AB - We have isolated the rat, mouse and human genes of a distant member of the TGF beta superfamily, growth/differentiation factor-15/macrophage inhibiting cytokine 1 (GDF-15/MIC-1) by screening of genomic libraries. All three genes are composed of two exons, and contain one single intron that interrupts the coding sequences at identical positions within the prepro-domain of the corresponding proteins. The predicted proteins contain the structural hallmarks of members of the TGF beta superfamily, including the seven conserved carboxy-terminal cysteine residues that form the cystine knot. The orthologous molecules show the lowest sequence conservation of all members of the TGF-beta superfamily. RT-PCR reveals an abundant expression of GDF-15/MIC-1 mRNA in numerous embryonic and adult organs and tissues. Promoter analysis of the rat promoter indicates the presence of multiple regulatory elements, including a TATA-like sequence as well as several SP1, AP-1 and AP-2 sites. Deletion analysis suggests that a 350 bp region upstream of the start of the open reading frame appears to be the most important for regulation of transcription. PMID- 10524242 TI - How many potentially secreted proteins are contained in a bacterial genome? AB - Artificial neural networks were trained on the prediction of the subcellular location of bacterial proteins. A cross-validated average prediction accuracy of 93% was reached for distinction between cytoplasmic and non-cytoplasmic proteins, based on the analysis of protein amino-acid composition. Principal component analysis and self-organizing maps were used to create graphical representations of amino-acid sequence space. A clear separation of cytoplasmic, periplasmic, and extracellular proteins was observed. The neural network system was applied to predicting potentially secreted proteins in 15 complete genomes. For mesophile bacteria the predicted fractions of non-cytoplasmic proteins agree with previously published estimates, ranging between 15% and 30%. Characteristics of thermophile genomes might lead to an under-estimation of the fraction of secreted proteins by presently available prediction systems. A self-organizing map was constructed from all 15 bacterial genomes. This technique can reveal additional sequence features independent from exhaustive pair-wise sequence alignment. The Treponema pallidum and Mycobacterium tuberculosis data formed separate clusters indicating unusual characteristics of these genomes. PMID- 10524243 TI - Sequence, overproduction and purification of the family 11 endo-beta-1,4-xylanase encoded by the xyl1 gene of Streptomyces sp. S38. AB - The xyl1 gene encoding the Xyl1 xylanase of Streptomyces sp. strain S38 was cloned by screening an enriched DNA library with a specific DNA probe and sequenced. Three short 5 bp -CGAAA- sequences are located upstream of the Streptomyces sp. S38 xyl1 gene 105, 115 and 250 bp before the start codon. These sequences, named boxes 1, 2 and 3, are conserved upstream of the Actinomycetales xylanase genes and are specifically recognized by a DNA-binding protein (Giannotta et al., 1994. FEMS Microbiol. Lett. 142, 91-97) and could be probably involved in the regulation of xylanase production. The Xyl1 ORF encodes a 228 residue polypeptide and the Xyl1 preprotein contains a 38 residue signal peptide whose cleavage yields a 190 residue mature protein of calculated M(r) = 20,585 and basic pI value of 9.12. The molecular mass of the produced and purified mature protein determined by mass spectrometry (20,586 +/- 1 Da) and its pI (9.8) agree with these calculated values. Its N-terminal amino-acid sequence confirmed the proposed cleavage site between the signal peptide and the mature protein. Comparisons between Xyl1 and the 62 other xylanases belonging to family 11 allowed the construction of a phylogenetic tree and revealed its close relationship with Actinomycetales enzymes. Moreover, nine residues were found to be strictly conserved among the 63 xylanases. PMID- 10524244 TI - Developmentally-regulated expression of mNapor encoding an apoptosis-induced ELAV type RNA binding protein. AB - Proteins with RNA recognition motifs (RRMs) participate in many aspects of RNA metabolism, and some of them are required for the accomplishment of normal development. The neuroblastoma apoptosis-related RNA binding protein (NAPOR) is an ELAV-type RNA-binding protein with three characteristic RNP2/RNP1-type RRMs, which we identified as a gene induced during apoptosis of neuroblastoma cells. Here we isolated and characterized the cDNA for mNapor, the mouse homolog of NAPOR. The mNapor encodes mRNA sharing striking homology with that of NAPOR, not only in its open reading frame (98.5%) but also in the 3'-untranslated region (80.1%), and is mapped to chromosome 2 A2-A3, a region syntenic to the human NAPOR locus. In situ hybridization analysis revealed that the expression pattern of mNapor is spatially and temporally coincident with the occurrence of programmed cell death, suggesting its involvement in the development of the central nervous system in which apoptosis plays a crucial role. PMID- 10524245 TI - Evolution of the proximal promoter region of the mammalian growth hormone gene. AB - The evolutionary relationship between the proximal growth hormone (GH) gene promoter sequences of 12 mammalian species was explored by comparison of their trinucleotide composition and by multiple sequence alignment. Both approaches yielded results that were consistent with the known fossil record-based phylogeny of the analysed sequences, suggesting that the two methods of tree reconstruction might be equally efficient and reliable. The pattern of evolution inferred for the mammalian GH gene promoters was found to vary both temporally and spatially. Thus, two distinct regions devoid of any evolutionary changes exist in primates, but only one of these 'gaps' is also observed in rodents, and neither is seen in ruminants. Furthermore, different evolutionary rates must have prevailed during different periods of evolutionary time and in different lineages, with a dramatic increase in evolutionary rate apparent in primates. Since a similar pattern of discontinuity has been previously noted for the evolution of the GH-coding regions, it may reflect the action of positive selection operating upon the GH gene as a single cohesive unit. Strong evidence for the action of gene conversion between primate GH gene promoters is provided by the fact that the human GH1 and GH2 sequences, which are thought to have diverged before the divergence of Old World monkeys from great apes, are more similar to one another than either is to the rhesus monkey GH2 promoter. Finally, it was noted that a number of nucleotide positions in the GH1 gene promoter that are polymorphic in humans appear to be highly conserved in mammals. This apparent conundrum, which could represent a caveat for the interpretation of phylogenetic footprinting studies, is potentially explicable in terms either of reduced genetic diversity in highly inbred animal species or insufficient population data from non-human species. PMID- 10524246 TI - Improved assay sensitivity of an engineered secreted Renilla luciferase. AB - We have previously reported the construction of a functional Renilla luciferase enzyme secreted by mammalian cells when fused to the signal peptide of human interleukin-2. The presence of three predicted cysteine residues in the amino acid sequence of Renilla luciferase suggested that its secreted form could contain oxidized sulfhydryls, which might impair enzyme activity. In this work, four secreted Renilla luciferase mutants were constructed to investigate this possibility: three luciferase mutants in which a different cysteine residue was replaced by an alanine residue, and one luciferase mutant in which all three cysteine residues were replaced by alanine residues. Simian cells were transfected with the genes encoding these mutant luciferases, as well as with the original gene construct, and cell culture media were assayed for bioluminescence activity. Only media containing a mutated luciferase with a cysteine to alanine substitution at position 152 in the preprotein showed a marked increase in bioluminescence activity when compared to media containing the original secreted Renilla luciferase. This increase in light emission was due in part to enhanced stability of the mutant enzyme. This new enzyme represents a significant improvement in the sensitivity of the secreted Renilla luciferase assay for monitoring gene expression. PMID- 10524247 TI - Alternative splicing creates sex-specific transcripts and truncated forms of the furin protease in the parasite Dirofilaria immitis. AB - Many extracellular proteins are activated by specific cleavage with an endoprotease. In nematodes, several proteins are cleaved after RX(K/R)R, the recognition site for the subtilisin-like proprotein convertases, furin and blisterase. To characterize furin in the parasitic nematode Dirofilaria immitis, we determined the sequence of the difur gene and its multiple transcripts. The gene spans 11 kb; encodes 16 exons and has a complex pattern of alternative splicing which generates at least 16 distinct mRNAs. The major transcript is a 4.4 kb mRNA which codes for a protein of 834 aa with an unusually long prodomain of 254 aa. Sex-specific splice variants of difur were observed by RT-PCR. The three female-specific and five male-specific transcripts are the first reported examples of sex-specific splicing in parasitic nematodes. This suggests that nematodes have sex-specific factors which regulate RNA splicing. Other splice variants are predicted to alter the phosphorylation and localization of the protease. Alternative splicing after the prodomain encodes a truncated protein that may be an inhibitor and/or substrate of Difurin. PMID- 10524248 TI - Transcriptional induction of p69 2'-5'-oligoadenylate synthetase by interferon alpha is stimulated by 12-O-tetradecanoyl phorbol-13-acetate through IRF/ISRE binding motifs. AB - Protein kinase C (PKC) is required for transcriptional induction of 2'-5' oligoadenylate (2-5A) synthetases by interferon (IFN)-alpha. Regulatory elements located in the 5'-flanking region of the p69 2-5A synthetase gene have been identified which are required for transcriptional stimulation by PKC. The region from -366 to -117 bp, relative to the translational start site, contains three sequence motifs that resemble interferon stimulated response elements/interferon regulatory factor elements (ISRE/IRF-E), which are required for stimulation of the IFN-alpha-response by the PKC activator, 12-O-tetradecanoyl phorbol-13 acetate (TPA). Constructs which have a deletion of the region containing IRF-Es located at -361 bp to -280 and at -246 to -172 bp do not respond to TPA treatment. Likewise, introduction of point mutations into either of these IRF-Es decreases stimulation of IFN-alpha induction by TPA and constructs containing point mutations in both upstream IRF-Es are nonresponsive to TPA. Binding of the inducible factor to the ISRE is abrogated in cells depleted of PKC by prolonged treatment with TPA. PKC appears to function as a signaling component in an IFN independent pathway that increases the activity of IFN-alpha-regulated transcription factors in the nucleus. PMID- 10524250 TI - Genomic analysis of male germ cell-specific actin capping protein alpha. AB - The Gsg3 gene which expresses specifically in haploid germ cells is a mouse testicular homolog of somatic cell type actin capping protein alpha (ACP alpha). We have obtained a mouse Gsg3 genomic clone using cDNA as a probe. Sequencing data showed that the Gsg3 gene was not interrupted by introns. The transcription initiation site of the gene was preceded not by a TATA box or GC rich promoter motifs, but by two consensus cAMP-response element (CRE) motifs at the putative position. Southern blotting analysis showed that Gsg3 is a single copy gene in the mouse, and conserved in mammals. Phylogenetic analysis showed that Gsg3 is a novel ACP alpha specific for haploid germ cells. PMID- 10524249 TI - The human homolog of Sex comb on midleg (SCMH1) maps to chromosome 1p34. AB - Polycomb group genes were originally identified in Drosophila as repressors required to maintain the silenced state of homeotic loci. About ten Polycomb group genes have been cloned in Drosophila, and mammalian homologs have been identified for most of these. Here, we isolate cDNAs encoding two isoforms of a human homolog of Drosophila Sex comb on midleg (Scm), named Sex comb on midleg homolog-1 (SCMH1). Overall, SCMH1 has 94% identity to its mouse counterpart Scmh1, and 41% identity to Scm, and contains two 1(3)mbt domains, and the SPM domain that are characteristic of Scm. SCMH1 is widely expressed in adult tissues, and maps to chromosome 1p34. PMID- 10524251 TI - Cloning of a cDNA encoding a sequence-specific single-stranded-DNA-binding protein from Rattus norvegicus. AB - In this paper, we report the isolation of a cDNA clone encoding a sequence specific single-stranded-DNA-binding protein (SSDP) from rat (Rattus norvegicus). The full-length nucleotide sequence was determined and encodes a 361 amino acid protein with a predicted molecular mass of 37.7 kDa. This clone has approximately 80% homology to a previously isolated partial cDNA clone for SSDP from chicken (Gallus gallus). Northern blot analysis revealed two transcripts of 2.0 and 3.0 kb. The protein appears to be evolutionarily highly conserved with > 97% identity between chicken, rat, mouse and human. Chicken SSDP has been proposed to be involved in the transcriptional regulation of the alpha 2(I) collagen gene. PMID- 10524252 TI - Cloning and characterization of the human ADP-ribosylation factor 4 gene. AB - ADP-ribosylation factor 4 (ARF4) is a member of a family of approximately 20 kDa guanine nucleotide-binding proteins that were initially identified by their ability to stimulate the ADP-ribosyltransferase activity of cholera toxin in vitro. They have recently been shown to play a role in vesicular trafficking and as activators of phospholipase D. The organization of the human ARF4 gene was determined from a genomic clone isolated from an arrayed PAC genomic library. The gene spans approximately 12 kb and contains six exons and five introns. Translation initiates in exon 1 and terminates in exon 6. Nuclease protection experiments indicated that the major transcription initiation site is located 211 bp 5' to the start of translation. In some cell lines derived from human tissues, however, multiple initiation sites were observed. The proximal 5'-flanking region of the human ARF4 gene lacks a TATA box, is highly GC rich, and contains multiple potential Spl-binding sites. An alignment of the exons for the class I ARF genes (ARF1, ARF2, and ARF3) and class II ARF genes (ARF4 and ARF5) reveals that the members of each class share a common gene organization. The structures of the class I and II ARF genes, however, are quite distinct and support the division of the ARFs into these groups based on deduced amino acid sequence, protein size, phylogenetic analysis, and gene structure. PMID- 10524253 TI - Sequence analysis of the chitin synthase A gene of the Dutch elm pathogen Ophiostoma novo-ulmi indicates a close association with the human pathogen Sporothrix schenckii. AB - Degenerate oligonucleotide primers were designed according to conserved regions of the chitin synthase gene family and used to amplify a 621 basepair (bp) fragment from genomic DNA of Ophiostoma novo-ulmi, the causal agent of Dutch elm disease. The amplification product was used as a hybridization probe to screen a library of genomic DNA sequences and to retrieve a full-length chitin synthase gene (chsA). The putative coding region of the gene was 2619 bp long, lacked introns, and encoded a polypeptide of 873 amino acids. Based on the similarity of the predicted amino acid sequence to the full-length chsC gene of Aspergillus nidulans and chsA gene of Ampelomyces quisqualis, the O. novo-ulmi chsA was classified as a Class I chitin synthase. The phylogenies constructed, according to a subregion of all available chitin synthases, showed that O. novo-ulmi consistently clustered most closely with the human pathogen Sporothrix schenckii, recently classified as a member of the mitosporic Ophiostomataceae. Disruption of the chsA gene locus had no obvious effects on the growth or morphology of the fungus. PMID- 10524254 TI - Domain organization and molecular characterization of 13 two-component systems identified by genome sequencing of Streptococcus pneumoniae. AB - In bacteria, adaptive responses to environmental stimuli are often initiated by two-component signal transduction systems (TCS). The prototypical TCS comprises two proteins: a histidine kinase (HK, hk) and a response regulator (RR rr). Recent research has suggested that compounds that inhibit two-component systems might have good antibacterial activity. In order to identify TCS that are crucial for growth or virulence of Streptococcus pneumoniae, we have examined the genomic sequence of a virulent S. pneumoniae strain for genes that are related to known histidine kinases or response regulators. Altogether 13 histidine kinases and 13 response regulators have been identified. The protein sequences encoded by these genes were compared with sequences deposited in public databases. This analysis revealed that two of the 13 pneumococcal TCSs have been described before (ciaRH and comDE) and two are homologous to the yycFG and the phoRP genes of Bacillus subtilis. All the pneumococcal response regulators contain putative DNA binding motifs within the C-terminal output domain, implying that they are involved in transcriptional control. Two of these response regulators are obviously the first representatives of a new subfamily containing an AraC-type DNA-binding effector domain. To assess the regulatory role of these transcription factors, we disrupted each of the 13 response regulator genes by insertional mutagenesis. All the viable mutant strains with disrupted response regulator genes were further characterized with regard to growth in vitro, competence, and experimental virulence. Two response regulator genes could not be inactivated, indicating that they may regulate essential cellular functions. The possibility of using these systems as targets for the development of novel antibacterials will be discussed. PMID- 10524255 TI - Molecular cloning of a novel tissue-specific gene from human nasopharyngeal epithelium. AB - A novel cDNA of human nasopharyngeal epithelium was cloned. The cDNA fragment of the gene, termed NESG1, was originally isolated by mRNA differential display, and was not homologous to any of the known genes in the database. The complete sequence of NESG1 cDNA, 1850 bp, contains an open reading frame of 1575 nucleotides, and encodes a soluble basic protein of 386 amino acids containing multiple protein kinase phosphorylation sites. The deduced protein has no homology to any of the known proteins in the database. A homologous STS localized NESG1 to the chromosomal region of 1q22-24. Messenger RNA of this gene was expressed only in nasopharynx and trachea. These results suggest that the human NESG1 gene may be a specific gene of columnar ciliated epithelium. PMID- 10524256 TI - Genomic organization of the Oreochromis mossambicus glutamate receptor subunit 2 beta gene (fGluR2 beta): presence of two different 5'-untranslated regions. AB - The AMPA (alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid)-preferring receptor is one of the pharmacologically defined ionotropic glutamate receptors, which mediate fast excitatory synaptic transmission in the central nervous system of vertebrates. Here, we report the mapping of the transcriptional start points and identification of the intron-exon boundaries of the teleost AMPA receptor subunit gene fGluR2 beta. fGluR2 beta and the mouse GluR2 share a similar genomic organization, having identical intron insertion sites and a large intron 2; however, fGluR2 beta has an extra exon encoding an alternate 5'-UTR. Results of RT-PCR and RNase protection analyses indicate that mature fish brain expresses two types of fGluR2 beta transcripts with different 5' ends. Transcriptions of these two fGluR2 beta transcripts started from two chromosomal regions separated by at least 10 kb. Only the transcript starting from the region more upstream on the chromosome was spliced. Moreover, transcript initiated from the downstream region was more abundant than that initiated from the upstream region. PMID- 10524257 TI - Characterization of a brain-specific nuclear LIM domain protein (FHL1B) which is an alternatively spliced variant of FHL1. AB - We have amplified and sequenced a novel, alternatively spliced variant of a human gene coding for the four-and-a-half LIM domain protein 1 (FHL1). This gene is located at chromosome Xq27 and the spliced variant is named FHL1B. The ORF of FHL1B cDNA codes for a LIM-only protein that possesses a zinc finger and three tandem repeats of LIM domains at the N-terminus with an active bipartite nuclear localization signal (NLS) motif and a possible RBP-J binding region at the C terminus. FHL1B and FHL1 have the same N-terminal three-and-a-half LIM domains but different C-terminal protein sequences, due to the presence of an additional alternative exon 4b in FHL1B causing a frame-shift in the 3'coding region. RT-PCR results revealed that the expression of FHL1 is not restricted in skeletal muscle and heart, but is widely distributed in other tissues, including brain, placenta, lung, liver, kidney and pancreas, albeit as a low abundance transcript. In contrast, FHL1B is specifically expressed in brain. The C-terminal alternative region in FHL1B is sufficient to localize FHL1B in the nucleus of mammalian cell. FHL1B is probably related to neural differentiation and certain fragile X syndrome. PMID- 10524258 TI - Regulation of avian fibroblast growth factor receptor 1 (FGFR-1) gene expression during skeletal muscle differentiation. AB - Myogenic cell proliferation and differentiation are regulated by a fibroblast growth factor (FGF) signal transduction cascade mediated by a high-affinity fibroblast growth factor receptor (FGFR). Exogenous FGF added to myogenic cultures has a mitogenic effect promoting myoblast proliferation while repressing differentiation. We have examined the regulation of the FGFR-1 gene (cek-1) in avian myogenic cultures by immunocytochemistry and Northern blot analysis. FGFR-1 protein was readily detected in undifferentiated myoblast cultures and was significantly reduced in differentiated muscle fiber cultures. Similarly, FGFR-1 mRNA was 2.5-fold more abundant in myoblast cultures than in differentiated cultures. To define the molecular mechanism regulating FGFR-1 gene expression in proliferating myoblasts and post-mitotic muscle fibers, we have isolated and partially characterized the avian FGFR-1 gene promoter. Transfection of FGFR-1 promoter-chloramphenicol acetyltransferase gene constructs into myogenic cultures identified two regions regulating expression of this gene in myoblasts. A distal region of 2226 bp conferred a high level of expression in myoblasts. This region functioned in an orientation-dependent manner and interacted with a promoter element(s) in a proximal 1058 bp promoter region to direct transcription. Deletion analysis revealed a 78 bp region that confers a high level of cek1 promoter activity in myoblasts. This DNA segment also contains Spl binding sites and interacts with a component in myoblast nuclear protein extracts. The proximal promoter region alone demonstrated no activity in directing transcription in either myoblasts or muscle fibers. Using the full-length promoter, gene expression was significantly decreased in differentiated muscle fibers relative to undifferentiated myoblasts indicating that the promoter-reporter gene constructs contain elements regulating expression of the endogenous FGFR-1 gene in both myoblasts and muscle fibers. PMID- 10524259 TI - The caa3 terminal oxidase of the thermohalophilic bacterium Rhodothermus marinus: a HiPIP:oxygen oxidoreductase lacking the key glutamate of the D-channel. AB - The respiratory chain of the thermohalophilic bacterium Rhodothermus marinus contains a novel complex III and a high potential iron-sulfur protein (HiPIP) as the main electron shuttle (Pereira et al., Biochemistry 38 (1999) 1268-1275 and 1276-1283). In this paper, one of the terminal oxidases expressed in this bacterium is extensively characterised. It is a caa3-type oxidase, isolated with four subunits (apparent molecular masses of 42, 19 and 15 kDa and a C-haem containing subunit of 35 kDa), which has haems of the A(s) type. This oxidase is capable of using TMPD and horse heart cytochrome c as substrates, but has a higher turnover with HiPIP, being the first example of a HiPIP:oxygen oxidoreductase. The oxidase has unusually low reduction potentials of 260 (haem C), 255 (haem A) and 180 mV (haem A3). Subunit I of R. marinus caa3 oxidase has an overall significant homology with the subunits I of the COX type oxidases, namely the metal binding sites and most residues considered to be functionally important for proton uptake and pumping (K- and D-channels). However, a major difference is present: the putative essential glutamate (E278 in Paraccocus denitrificans) of the D-channel is missing in the R. marinus oxidase. Homology modelling of the R. marinus oxidase shows that the phenol group of a tyrosine residue may occupy a similar spatial position as the glutamate carboxyl, in relation to the binuclear centre. Moreover, sequence comparisons reveal that several enzymes lacking that glutamate have a conserved substitution pattern in helix VI: -YSHPXV- instead of -XGHPEV-. These observations are discussed in terms of the mechanisms for proton uptake and it is suggested that, in these enzymes, tyrosine may play the role of the glutamate in the proton channel. PMID- 10524260 TI - Effect of adenine nucleotide pool size in mitochondria on intramitochondrial ATP levels. AB - Net adenine nucleotide transport into and out of the mitochondrial matrix via the ATP-Mg/Pi carrier is activated by micromolar calcium concentrations in rat liver mitochondria. The purpose of this study was to induce net adenine nucleotide transport by varying the substrate supply and/or extramitochondrial ATP consumption in order to evaluate the effect of the mitochondrial adenine nucleotide pool size on intramitochondrial adenine nucleotide patterns under phosphorylating conditions. Above 12 nmol/mg protein, intramitochondrial ATP/ADP increased with an increase in the mitochondrial adenine nucleotide pool. The relationship between the rate of respiration and the mitochondrial ADP concentration did not depend on the mitochondrial adenine nucleotide pool size up to 9 nmol ADP/mg mitochondrial protein. The results are compatible with the notion that net uptake of adenine nucleotides at low energy states supports intramitochondrial ATP consuming processes and energized mitochondria may lose adenine nucleotides. The decrease of the mitochondrial adenine nucleotide content below 9 nmol/mg protein inhibits oxidative phosphorylation. In particular, this could be the case within the postischemic phase which is characterized by low cytosolic adenine nucleotide concentrations and energized mitochondria. PMID- 10524261 TI - Uncoupling protein 2 from carp and zebrafish, ectothermic vertebrates. AB - Uncoupling protein 1 (UCP1) is of demonstrated importance in mammalian thermogenesis, and early hypotheses regarding the functions of the newly discovered UCP homologues, UCP2, UCP3 and others, have focused largely on their potential roles in thermogenesis. Here we report the amino acid sequences of two new UCPs from ectothermic vertebrates. UCPs from two fish species, the zebrafish (Danio rerio) and carp (Cyprinus carpio), were identified in expressed sequence tag databases at the European Molecular Biology Laboratory. cDNAs from a C. carpio 'peritoneal exudate cell' cDNA library and from a D. rerio 'day 0 fin regeneration' cDNA library were obtained and fully sequenced. Each cDNA encodes a 310 amino acid protein with an average 82% sequence identity to mammalian UCP2s. The fish UCP2s are about 70% identical to mammalian UCP3s, and 60% identical to mammalian UCP1s. Carp and zebrafish are ectotherms--they do not raise their body temperatures above ambient by producing excess heat. The presence of UCP2 in these fish thus suggests the protein may have function(s) not related to thermogenesis. PMID- 10524262 TI - Cloning and sequence analysis of the gene encoding Methylophilus methylotrophus cytochrome c", a unique protein with a perpendicular orientation of the histidinyl ligands. AB - Cytochrome c" from Methylophilus methylotrophus is an unusual monohaem protein that undergoes a major redox-linked spin-state transition: one of the two axial histidines bound to the iron in the oxidised form is detached upon reduction and a proton is taken up. A 3.5-kb DNA fragment, containing the gene encoding cytochrome c" (cycA), has been cloned and sequenced. The cytochrome c" gene codes for a pre-protein with a typical prokaryotic 20-residue signal sequence, suggesting that the protein is synthesised as a precursor which is processed during its secretion into the periplasm. The C-terminus of cytochrome c" has homology with the corresponding region of an oxygen-binding haem protein (SHP) from phototrophically grown Rhodobacter sphaeroides. SHP is similar in size and in the location of its haem-binding site. Immediately downstream from cytochrome c" a second open reading frame (ORF) codes for a 23-kDa protein with similarity to the cytochrome b-type subunit of Ni-Fe hydrogenase. The possibility of coordinated expression of cycA and this ORF is discussed. PMID- 10524263 TI - [Phylogeny of nociceptors]. PMID- 10524264 TI - [Real-time detection of glutamic acid, acetylcholine, and GABA released from cultured neurons using the online enzyme sensor]. PMID- 10524265 TI - [Morphological and enzymatic adaptation of skeletal muscle fibers in stroke-prone hypertensive rats to ACE inhibitors]. PMID- 10524266 TI - [Photoallergic dermatitis]. AB - The most common "allergic" photodermatoses are reported in this review: (a) systemic and topical exogenous photosensitizations with phototoxic and photoallergic reactions and the most frequently chemicals photosensitizers incriminated. (b) idiopathic photodermatoses including benign summer eruption, polymorphous light eruption and solar urticaria. Clinical recognition patterns are stressed as well as differential diagnosis. Photobiological exploration is also detailed. PMID- 10524267 TI - [The Prolair Autohaler in the treatment of adult asthma in current practice]. AB - OBJECTIVE OF THE STUDY: To describe the use of the Prolair Autohaler (Prolair AH) in the conditions of "natural" prescription so as to define its place in the therapeutic arsenal available to practitioners. NATURE OF THE STUDY: Open study conducted in any practice of liberal chest physicians. RESULTS: Three hundred and seventy six patients (56.1%) were treated by Prolair AH and 296 (43.9%) continued their usual treatment with a standard aerosol doser (BDP ADS) that contained the same active principle, used with an inhalation chamber (65 patients, around 22%) or without (231 patients around 78%). The comparisons between the two groups, made at the end of two months of treatment, showed significant differences in efficacy of the two therapies. The percentage of patients who presented a respiratory shortage was significantly lower at day 60 in the Prolair AH group than in the BDP ADS (33.6% vs 41.4%; p < 0.05) though the percentages were comparable at inclusion. Peak problems were significantly more frequent at inclusion in the Prolair AH group (45.5% vs 37.8%; p < 0.05) but at day 60 were significantly less frequent (14.6% vs 22.9%; p < 0.05). Up to day 60, in each treatment group, the same percentage of patients presented whistling (Prolair AH 21.8% vs 22.9%; p: NS) although initially they were significantly more frequent in the patients of the Prolair AH group (60.3% vd 49.8%; p < 0.01). Improvement of DEP measured theoretically was significantly more important (p < 0.05) in the Prolair AH group which passed from 67.6% to 78.8% against 70.5% to 75.6% in the BDP ADS group. CONCLUSION: When prescribed to patients with a more evolutive asthma and poor coordinators, Prolair Ahas produced results that are comparable to or better than those of patients treated with ADS that contains the same active principle, together or not with an inhalation chamber. PMID- 10524268 TI - [History-taking of the allergic patient]. AB - Etiological diagnosis of an allergic condition is assured mostly clinically and in particular by the data from a specific and minute interrogation. Complementary investigations only confirm the diagnostic suspicion produced by this interrogation, which includes several essential steps: The history of the patient from the first symptom to the day of consultation. The common criteria of unity of time, place or action on the different manifestations of hypersensitivity. The provocation factors and improvement of critical episodes. The mode of life of the subject and the factors of environmental imbalance. PMID- 10524269 TI - [Advice for allergic travellers]. AB - Business and tourist journeys by air contribute to exposure of the body to multiple environments. The allergic patient, considered rightly to be a sentry of the environment, has many reasons to care about his journeys and to take precautions that are adapted to his case under the impetus of advice and information from his physician and his specialist. Some advice falls within a simple logic that is enough to remember when planning the journey while the others measures must follow a correct preventative strategy for allergy risks as much as those that concern the modalities before leaving as a drive taken on the ground. It is important therefore to know how to give advice and information on the different risks linked to the allergic condition and to the field of allergy and help the patient to orientate his choice of place of the journey, the methods of lodging, of transport and the programme of the journey. The advice should also include the preventative measures as a function of the known pathology under the form of medical equipment before, during the stay and on return. Finally some advice relative to medical equipment for prevention and cure would appear to be judicious. PMID- 10524270 TI - [Asthma and diving with a cylinder]. AB - Undersea diving is an activity that is practised more and more in holiday clubs. There is no precise legislation on the causes of unfitness of the amateur, in contrast to the professional diver, where the medical criteria are strict and controlled. When diving with a cylinder, on descent, the ventilatory load increases with increase of the ambient pressure and dynamic resistance in the airways increases. "As with an insufficient respiration on the surface, a healthy subject when diving has a ventilatory ability that is drastically reduced". Moreover with cylinder ventilation, the diver has available a reserve of gas under pressure from which he inspires with the aid of a breathing apparatus (regulator): he breathes dry gas that is dried before compression in the reservoirs, chilled by the relief valve on leaving the reservoir. This inhalation of cold, dry air associated with a hyperventilation during the descent produces ideal conditions to trigger exercise induced asthma. All subjects who present a bronchial hyperreactivity have the risk when diving with a cylinder of triggering a bronchospasm that is identical with that of a sporting asthmatic. During surfacing: the re-surfacing diver runs the risk of an accident of pulmonary suppression if he does not expire sufficiently during his return to the surface: the mass of intrapulmonary air of the resurfacer dilates and the excess of volume is exhaled by the diver: a volume of air of 5 l at 10 m depth corresponds to a volume of 10 l on the surface. Therefore the airways must remain free: an obstruction of the peripheral airways associated with an urgent re-surfacing produces a very rapid thoracic dilation which is responsible for pulmonary barotrauma (pulmonary barotrauma is frequently lethal with 30% of accidental deaths). PMID- 10524271 TI - Clonidine action in spontaneously hypertensive rats (SHR) depends on the GABAergic system function. AB - The effect of gamma-aminobutyric acid (GABA)A antagonists (bicuculline, picrotoxin) on clonidine hypotension in spontaneously hypertensive (SHR) and Wistar Kyoto (WKY) rats were examined. The GABA turnover changes after clonidine injection in both strains were also studied. Administration of clonidine alone induced the stronger decrease of systolic blood pressure (SBP) in SHR. Co-dosage of clonidine with these agents reduced its hypotensive effect in dose dependent manner and the effectiveness of both antagonists was higher in SHR. We find that clonidine stimulates GABA synthesis in the hypothalamus and the pons-medulla in both strains but the GABA turnover rate is significantly slower in SHR. Therefore, the differences in inhibitory action of GABAA receptor antagonists between WKY and SHR rats may be explained by central GABAergic system dysfunction in the hypertension. Our results indicate that the down regulation of the GABAergic system observed in hypertension may be compensated by the action of clonidine. PMID- 10524272 TI - Gender-related differences in polyamine oxidase activity in rat tissues. AB - Variations in level of polyamines and their related enzymes are frequently observed in response to some treatments which affect in a different way male and female. The possibility of a gender-related difference in the oxidation of polyamines was investigated in rats by measuring the activity of polyamine oxidase, a ubiquitous enzyme of vertebrate tissues, which transforms spermine into spermidine and spermidine into putrescine. The study was carried out on thymus, spleen, kidney and liver of young rats of both sexes, and female rats showed a lower polyamine oxidase activity than male rats in all the tissues. We also found higher values of spermidine acetylation in female than male rats in thymus and liver. Owing to these gender-related differences, a higher spermidine N-acetyltransferase/polyamine oxidase ratio was found in female than in male rats. A second gender-related difference was a higher spermidine/spermine ratio in female than in male, the only exception being the thymus. These basal differences possibly account for the gender-related differences of polyamine metabolic enzyme activities in response to some treatments, including drugs or hormones. PMID- 10524273 TI - Synthesis of biheterocyclic alpha-amino acids. AB - We report here the synthesis of biheterocyclic alpha-amino acids by 1,3 dipolar cycloaddition of acetylenic compounds on alpha-azido alpha-amino esters. PMID- 10524274 TI - Effect of some herbicides on the production of lysine by Azotobacter chroococcum. AB - Production of lysine by Azotobacter chroococcum strain H23 was studied in chemically-defined media amended with different concentrations of alachlor, metolachlor, 2,4-D, 2,4,5-T and 2,3,6-TBA. The presence of 5, 10, and 50 micrograms/ml of alachlor or 2,3,6-TBA significantly decreased quantitative production of lysine. However, the presence 2,4-D or 2,4,5-T at concentrations of 10 and 50 micrograms/ml enhanced the production of lysine. Quantitative production of lysine was not affected as consequence of the addition of metolachlor to the culture medium, showing that the release lysine to the culture media by A. chroococcum was not affected by that herbicide. PMID- 10524275 TI - Synthesis and enzymology of modified N-benzyloxycarbonyl-L-cysteinylglycyl-3,3 dimethylaminopropylamide++ + disulphides as alternative substrates for trypanothione reductase from Trypanosoma cruzi: Part 3. AB - Kinetic data for alternative substrates of recombinant trypanothione reductase from Trypanosoma cruzi were measured for a series of N-substituted-L cysteinylglycyl-3-dimethylaminopropylamides, in which the cysteine N-substituent was either a variant of the benzyloxycarbonyl group or was L-phenylalanine or L tryptophan. Replacing the benzylic ether oxygen atom by CH2 or NH had relatively minor effects on kcat, but raised the value of K(m) 4.5- and 10-fold, respectively. Similarly, relative to the carbobenzoxy group, an N-L-phenylalanyl or N-L-tryptophanyl replacement on the cysteine hardly altered kcat, but increased K(m) values by 16.6 and 7.4 fold, respectively. These observations were consistent with the K(m) values referring primarily to binding for this series of nonspecific substrates. PMID- 10524276 TI - Utilization of D-amino acids by Fusobacterium nucleatum and Fusobacterium varium. AB - The utilization of D- and L-amino acids with acidic, basic or polar side chains was demonstrated by HPLC. Two species of the anaerobe Fusobacterium utilized D lysine and the L isomers of glutamate, glutamine, histidine, lysine and serine. Only F. varium used L-arginine, D-glutamate and D-serine as substrates, whereas F. nucleatum specifically utilized D-histidine and D-glutamine. D-Glutamate accumulated in F. nucleatum cultures supplemented with D-glutamine, and ornithine was detected when either DL- or L-arginine was included in F. varium cultures. Based on literature precedents, D-glutamate and D-histidine are isomerized to their L isomers prior to degradation, but separate catabolic pathways are possible for each enantiomer of lysine and serine. PMID- 10524277 TI - Amino acids and osmolarity in honeybee drone haemolymph. AB - In the haemolymph of honeybee drones, concentrations of free amino acids were higher than in worker haemolymph, with different relative proportions of individual amino acids. The overall concentration of free amino acids reached its highest level at the 5th day after adult drone emergence, and after the 9th day only minor changes in the concentration and distribution of free amino acids were observed. This coincides with the age when drones reach sexual maturity and change their feeding behaviour. Levels of essential free amino acids were high during the first 3 days of life and thereafter decreased. Osmolarity was lowest at emergence (334 +/- 42 mOsm), increased until the age of 3 days (423 +/- 32 mOsm) and then stayed generally constant until the 16th day of life. Only 25-day old drones had significantly higher osmolarity (532 +/- 38 mOsm). The overall change in osmolarity during a drone's lifetime was about 40%. PMID- 10524278 TI - Evolutionary changes reflected by the cellular amino acid composition. AB - Comparison of the amino acid composition of cell-proteins using 17 amino acids has been used to investigate the biological evolution of organisms such as bacteria, blue-green alga, green alga, fungi, slime mold, protozoa and vertebrates. The degree of difference in the amino acid ratios between any two groups reflects the degree of divergency in biological evolution. The amino acid composition of the Gram-negative bacteria (Escherichia coli, Klebsiella, Proteus, and Vibrio alginolyticus) was identical. However, the amino acid composition of Staphylococcus aureus and Bacillus subtilis, which are Gram-positive bacteria, differed from each other and from the Gram-negative bacteria. The amino acid composition of the blue-green alga (Cyanobacterium, Chroococidiopsis) was quite similar to that of E. coli. A marked difference in the amino acid composition was observed between E. coli and green alga (Chlorella), and significant differences were observed between E. coli and other organisms, such as fungi, protozoa (Tetrahymena), slime mold (Dictyostelium discoideum) and vertebrates. In conclusion, the change in cellular amino acid composition reflects the divergence which has occurred during biological evolution, whereas a basic pattern of amino acid composition is maintained in spite of a long period of evolutional divergence among the various organisms. Thus, it is proposed that the primitive life forms established at the end of prebiotic evolution had a similar amino acid composition. PMID- 10524279 TI - Pathogenic amyloid beta-protein induces apoptosis in cultured human cerebrovascular smooth muscle cells. AB - The amyloid beta-protein (A beta) pathologically accumulates in cerebral vascular and senile plaque deposits in the brains of patients with Alzheimer's disease (AD) and related disorders including hereditary cerebral hemorrhage with amyloidosis Dutch type (HCHWA-D). The cerebrovascular deposits are accompanied by degeneration and eventual loss of smooth muscle cells in cerebral vessel wall. Similarly, we have shown that pathogenic forms of A beta cause cell death in cultured human cerebrovascular smooth muscle (HCSM) cells in vitro. Here we show that pathogenic A beta induces a number of structural changes in HCSM cells including shrinkage of cell bodies, retraction of processes, disruption of the intracellular actin network, and nuclear condensation and fragmentation. These changes were accompanied by a number of biochemical alterations in the cells shown by in situ end labeling of nuclear DNA, proteolytic breakdown of smooth muscle cell a actin, and proteolytic activation of the proteinase caspase 3. Together, these characteristics are consistent with an apoptotic mechanism of cell death in HCSM cells in response to pathogenic A beta. PMID- 10524280 TI - Tertiary structure of human lambda 6 light chains. AB - AL amyloidosis is a disease process characterized by the pathologic deposition of monoclonal light chains in tissue. To date, only limited information has been obtained on the molecular features that render such light chains amyloidogenic. Although protein products of the major human V kappa and V lambda gene families have been identified in AL deposits, one particular subgroup--lambda 6--has been found to be preferentially associated with this disease. Notably, the variable region of lambda 6 proteins (V lambda 6) has distinctive primary structural features including the presence in the third framework region (FR3) of two additional amino acid residues that distinguish members of this subgroup from other types of light chains. However, the structural consequences of these alterations have not been elucidated. To determine if lambda 6 proteins possess unique tertiary structural features, as compared to light chains of other V lambda subgroups, we have obtained x-ray diffraction data on crystals prepared from two recombinant V lambda 6 molecules. These components, isolated from a bacterial expression system, were generated from lambda 6-related cDNAs cloned from bone marrow-derived plasma cells from a patient (Wil) who had documented AL amyloidosis and another (Jto) with multiple myeloma and tubular cast nephropathy, but no evident fibrillar deposits. The x-ray crystallographic analyses revealed that the two-residue insertion located between positions 68 and 69 (not between 66 and 67 as previously surmised) extended an existing loop region that effectively increased the surface area adjacent to the first complementarity determining region (CDR1). Further, an unusual interaction between the Arg 25 and Phe 2 residues commonly found in lambda 6 molecules was noted. However, the structures of V lambda 6 Wil and Jto also differed from each other, as evidenced by the presence in the latter of certain ionic and hydrophobic interactions that we posit increased protein stability and thus prevented amyloid formation. PMID- 10524282 TI - Enhanced amyloidogenicity of sulfonated transthyretin in vitro, a hypothetical etiology of senile amyloidosis. AB - Genetic variants of transthyretin (TTR) cause systemic amyloidosis and wild-type TTR may also in some situations produce amyloid fibrils. We have analyzed wild type and variant TTRs by mass spectrometry and found that TTR preparations from all individuals demonstrated free TTR, TTR conjugated with thiol compounds and several minor components. We previously described a component which had a molecular mass 80 Da larger than free TTR and was proved to be TTR conjugated with sulfite. Here, the amyloid fibril formation of the TTR isoforms was monitored by the turbidity at 330 nm, and by a Congo red-binding assay as a function of pH, according to the method of Lai et al. The S-sulfonated TTR showed the highest level of amyloid fibril formation. In contrast, TTR reduced by dithiothreitol, which was free of the S-sulfonated component, showed neither elevation of the turbidity nor the Congo red binding. Commercially purchased TTR without further treatment containing free, S-sulfonated and other species of TTR molecules showed an intermediate elevation. These results suggested that the S sulfonated wild-type TTR is highly amyloidogenic. Although further experiments are needed to apply the observation to in vivo phenomenon, exogenous sulfite may be a cause of senile systemic amyloidosis. PMID- 10524281 TI - Cellular processing of the amyloidogenic cystatin C variant of hereditary cerebral hemorrhage with amyloidosis, Icelandic type. AB - An important gap in our understanding of the pathogenesis of the amyloidoses is the identification of the cellular events that lead from synthesis of an amyloid precursor protein to its conversion to the amyloid fiber subunit. We address this question by characterizing the effects of an amyloidogenic mutation on the intracellular processing of its protein product. The protein, a mutant of the cysteine protease inhibitor cystatin C, is the amyloid precursor protein in Hereditary Cerebral Hemorrhage with Amyloidosis--Icelandic type (HCHWA-I). The amyloid fibers are composed of mutant cystatin C (L68Q) that lacks the first 10 amino acids. We have previously shown that processing of wild-type cystatin C entails formation of a transient intracellular dimer that dissociates prior to secretion, such that extracellular cystatin C is monomeric. We report here that the cystatin C mutation engenders several alterations in its intracellular trafficking. It forms a stable intracellular dimer that is partially retained in the endoplasmic reticulum and degraded. The bulk of mutant cystatin C that is secreted does not dissociate and is secreted as an inactive dimer. Thus, formation of the stable mutant cystatin C dimer is an early event in the pathogenesis of this disease. PMID- 10524283 TI - Impact of age and amyloidosis on thiol conjugation of transthyretin in hereditary transthyretin amyloidosis. AB - Variant forms and post-translational modifications of transthyretin (TTR) can be identified by electrospray ionisation mass spectrometry (ESI-MS). The aim of the present study was to investigate thiol conjugation of transthyretin and it's relation to age and symptomatic amyloid disease in different populations of variant TTR carriers. Plasma samples from 70 individuals from Denmark, Argentina, Sweden and Japan, with 2 different TTR mutations were analysed. The percentage cysteine (Cys) conjugated wild and variant TTR were calculated from the corresponding peaks of the spectra, and multiple regression analysis was employed to disclose relationships between age, symptomatic amyloid disease and origin. Age, origin and presence of symptomatic disease, were found to be independent factors related to transthyretin conjugation. A higher percentage of conjugated to unconjugated TTR was disclosed in symptomatic, but not in asymptomatic carriers. In summary: Thiol conjugation of TTR is dependent on age and presence of symptomatic amyloid disease. Furthermore, it varies between different populations. Variant TTR is more susceptible to thiol conjugation than the wild type. Post-translational factors may be related to amyloid formation and/or toxicity. PMID- 10524284 TI - Endocrine cells in the upper gastrointestinal tract in relation to gastrointestinal dysfunction in patients with familial amyloidotic polyneuropathy. AB - Gastrointestinal (GI) dysfunction is a common complication of familial amyloidotic polyneuropathy (FAP). In previous reports, a decreased content of small and large intestinal endocrine cells has been found in patients with FAP and it has been suggested that this may contribute to the development of GI disturbances. The aim of the present study was to investigate the endocrine cell content in the stomach and duodenum of FAP patients, and to correlate the findings with gastric emptying. Fifteen patients with FAP were included in the study. Twenty-eight subjects with macroscopically and histologically normal mucosa were used as controls for endocrine cell contents and 14 healthy subjects for gastric scintigraphy. The endocrine cells were identified by immunohistochemistry and quantified with image analysis. Gastric emptying time was detected by scintigraphy and endoscopy. The number of chromogranin A immunoreactive (IR) cells was reduced in all investigated parts of the GI tract except bulbus duodeni. Gastrin/CCK cell content was reduced in duodenum, but tended to be increased in antrum of the stomach (P = 0.07). Otherwise, the content of all other endocrine cells types in the upper GI tract was reduced compared with controls. A correlation with malnutrition was found for gastric inhibitory polypeptide and secretin cell content in bulbus duodeni. Gastric scintigraphy disclosed delayed gastric emptying of solid food, but the finding was not correlated to the decreased content of neuroendocrine cells. The severity of endocrine cell depletion was not correlated to duration of GI disturbances. The present study showed that the endocrine cells of the stomach are affected in FAP patients and that the abnormalities in the upper GI endocrine cells occur early during the course of the disease. PMID- 10524285 TI - Serum amyloid A1 alleles and plasma concentrations of serum amyloid A. AB - Serum amyloid A1 (SAA1), the predominant isotype of acute phase SAA in plasma and the predominant precursor of fibrillar deposits in reactive amyloidosis, is encoded by a gene, for which six allelic variants have been described. Recent studies proposed that the allele SAA1.3 was positively correlated with the development of reactive amyloidosis in Japanese. This study examined whether the plasma concentration of total SAA is influenced by specific SAA1 alleles. Two hundred and eighty healthy Japanese subjects were examined to determine the allelic distribution of SAA1 and SAA2 genes by the PCR-RFLP method, and to measure the total plasma SAA concentrations. SAA concentrations were significantly higher (p < 0.001) in subjects with the allele SAA1.5 than those without it, suggesting that SAA1.5 may have a distinctive feature in the process of synthesis or catabolism. Subjects with the allele SAA1.3 had lower SAA concentrations, though not statistically significant, than those with SAA1.1. There was not significant correlation of SAA2 alleles with SAA concentrations. These results are discussed in terms of amyloidogenicity. PMID- 10524286 TI - Familial amyloidosis in cats: Siamese and Abyssinian AA proteins differ in primary sequence and pattern of deposition. AB - Familial AA amyloidosis is a hereditary trait in Abyssinian cats, with the kidney as the main target organ. The amino acid sequence of the amyloid A protein of the Abyssinian cat has been described earlier. Recently, familial amyloidosis has been found in Siamese cats, with the liver as the main target organ. In the present paper, we describe the complete amino amid sequence of the major constituent protein, of two Siamese cats. Siamese hepatic protein AA showed homology with, but was different from all feline SAA and AA sequences hitherto reported. Two substitutions (46Q-R and 52A-V) from the Abyssinian protein sequence were identified, one of which (46Q-R) is a non-homologous substitution not found in mammalian SAA, but is present in two bird AA amyloid proteins. This shows the presence of an unique amyloidogenic SAA isotype in Siamese cats. Both the Siamese and the Abyssinian sequence are amyloidogenic, thus making identification of amyloidogenic residues difficult. Apart from the apparent inherent amyloidogenicity of SAA, it can not be excluded that certain amino acid substitutions could enhance its amyloidogenicity but also could contribute to tissue predilection in amyloidosis. PMID- 10524287 TI - Conjunctival AL amyloidosis associated with a low-grade B-cell lymphoma. AB - A rare case of localized amyloidosis associated with a low-grade B-cell lymphoma involving the conjunctiva is described. Although infiltrating small lymphocytes and plasma cells showed little cytological atypia, molecular genetic examination revealed a prominent B-cell clonal immunoglobulin heavy chain (IgH) gene rearrangement in the tumor tissue. Immunoelectronmicroscopic examination showed immunoglobulin lambda light chain specificity in the amyloid deposit and Russell bodies in the surrounding plasma cells. We concluded that the immunoglobulin lambda light chain, produced by the tumor's differentiated plasma cells, is the precursor protein of the localized amyloidosis found in this case. PMID- 10524288 TI - Hereditary cerebral hemorrhage with amyloidosis--Dutch type (HCHWA-D): a review of the variety in phenotypic expression. PMID- 10524289 TI - International Workshop on Conformational Diseases, Dead Sea, Israel, November 8 12, 1998 (sponsored by the Center for the Study of Emerging Diseases (CSED)). PMID- 10524290 TI - Amount of metallic ions released from Ti-Ni alloy by abrasion in simulated bioliquids. AB - The current density of Ti-56mass%Ni (Ti-50at.%) alloy after abrasion in simulated bioliquids was measured using a potentiostat to estimate the amount of metallic ions released from the alloy during repassivation and maturation. The current density in saline, saline with and without N2 bubbling, and Hanks' solutions with and without proteins after abrasion was measured and the amount of released ion was calculated from the integrated current density with time, assuming that Ti4+ and Ni2+ are equivalently released. No difference in the amount of released ion was observed between saline with and without N2 bubbling. Also, no difference was observed between saline and pH 7.4 Hanks' solution. More Ti4+ and Ni2+ were released in bioliquids with proteins than in saline with and without N2 bubbling (p < 0.05). That is, dissolved oxygen and inorganic ions in Hanks' solution did not influence the amount of released ion, but proteins influenced it. The release of metallic ions from metals and alloys in biological systems can be estimated by the methodology employed in this study. PMID- 10524291 TI - Characterization and application of thin film pressure sensors. AB - Data regarding intra-joint loads during range of motion is essential to understanding normal joint mechanics and pathology. The investigators configured and characterized the response of a 440 N range, 0.076 mm thick commercial thin film sensor to monitor joint loads through a range of motion. Following preconditioning, static and dynamic tests were performed to evaluate the sensor response under varied environmental conditions. Both tests utilized a fixture to align the sensor and applied load. Under conditions that included dry, wet, folded and kinked configurations, a static load of 100 N was applied and the sensor output monitored up to thirty minutes from the time of loading. A load of 400 N at 50 N/s was applied to the sensor to determine dynamic characteristics and calibration curves under the conditions described for static tests in addition to curved, hard and soft contact surfaces and sensor overload. Static kinked data was significantly different from the dry, wet and folded conditions. Dynamic data showed that inter-package variability was not significant but that differences between sensor packages and sensor configuration conditions were significant. To investigate applicability of these sensors to the field of orthopaedics, a cadaveric knee was instrumented with sensors to examine the role of the meniscus in load transmission and distribution across the knee. The sensors were placed bilaterally below each meniscus on the anterior, posterior and center of the tibial plateau. Sensor data were obtained at these locations during manually flexed range of motion for the intact, re-attached and lateral menisectomized conditions of the knee specimen. PMID- 10524292 TI - Surface treatments of titanium in aqueous solutions containing calcium and phosphate ions. AB - Immersion treatment of titanium in aqueous solutions containing various kinds of ion concentrations of calcium and phosphate (pH 5.8, 7.0, and 8.0) were attempted to accelerate calcium phosphate precipitation on titanium in body fluid. The performance was confirmed using scanning electron microscopy, X-ray diffractometry, and Fourier transformed infrared absorption spectrometry with a reflection absorption spectrometer of the specimen immersed in Hanks' solution. Calcium phosphate precipitation on titanium in Hanks' solution is accelerated by the immersion treatment in aqueous solutions containing calcium and phosphate ions. The amount, composition, and shape of calcium phosphate precipitate vary according to the pH and ion concentrations of the solutions in which titanium is immersed. This method is effective for the surface treatment of inside pore narrow space of titanium materials. PMID- 10524294 TI - A comparison of the wear and debris generation of GUR 1120 (compression moulded) and GUR 4150HP (ram extruded) ultra high molecular weight polyethylene. AB - The wear debris generated from UHMWPE (ultra high molecular weight polyethylene) has been recognised as one of the major causes of failure in THR (total hip replacement). GUR 1120 (compression moulded) and GUR 4150HP (ram extruded) which are currently the most frequently used materials in THR were studied in pin-on plate wear test. The wear particles generated from this test were observed by scanning electron micrograph and analysed by image analysis. The results from this study showed that GUR 4150HP had superior wear resistance than GUR 1120 under relatively high wear factor conditions. These results also highlighted the importance of multidirectional motion and its effect on the wear rates of UHMWPE. The multidirectional motion tended to show a higher wear factor than previous studies using unidirectional motion conducted under otherwise similar conditions. The wear debris analysis conducted with the wear particles collected from unidirectional (relatively rough) pin-on-plate wear tests (GUR 1120 and GUR 4150HP) showed that the greatest number of particles had a size range of 0.1-0.5 micron followed by 0.5-1.0 micron, 1.0-5.0 microns and 5.0-10.0 microns, in both GUR 1120 and GUR 4150HP. However, comparing the masses of the wear particles, the bigger size range of greater than 10 microns, had the highest percent mass followed by 1.0-5.0 microns, 0.5-1.0 micron, 0.1-0.5 micron and 5.0-10.0 microns. PMID- 10524293 TI - Artificial SMA valve for treatment of urinary incontinence: upgrading of valve and introduction of transcutaneous transformer. AB - This paper is concerned with the development of an artificial urethral valve driven by shape memory alloy actuators, which is attached onto the urethra of a urinary incontinence sufferer for treating the involuntary micturition. Three types of compact cylindrical valves are assembled and their opening and closing functions are examined experimentally. The updated valve is heated and opened by using the transcutaneous energy transformer consisting of a pair of flexible spiral-formed copper wire coils. The experiment using the canine urinary canal verifies that the total system of the valve and the transformer works well as an artificial sphictor muscle and controls the urinary flow through the canal appropriately. PMID- 10524295 TI - Water-sorption kinetics of dental polymeric resin under tensile stressing conditions. AB - Water sorption tests were conducted on unfilled poly(methyl methacrylate) samples in distilled water at 5, 37, and 60 degrees C under three different tensile stress ratios (sigma appl/sigma ys = 0%, 5%, and 10%). Each sample was placed in a modified Hoffman open-side tubing clamp and subjected to four-point bending at pre-determined stress level for 1 day, 3 days, 1 week, 2 weeks, and 4 weeks. Water sorption was measured by weight change calculations, without accounting for any weight loss due to solubility of uncured monomer. A generalized diffusion equation can be used to express both stress-free and stress conditions; D = D0exp[-E (sigma)/kT]. It was found that the activation energy for water sorption diffusion was linearly related to applied stress ratios; i.e., E = 1.15 sigma appl/sigma ys + 10.76 (kJ/mol), with the correlation coefficient r = 0.97. Since the proportional pre-exponential constant, D0, is independent of temperature, it is speculated that the loading percentage of reinforcing filler elements in composite resin materials can be related to this constant. PMID- 10524296 TI - Manual DNA sequencing using fluorescent-labeled primers and a fluorescence scanner. PMID- 10524297 TI - Sequencing errors in reactions using labeled terminators. PMID- 10524298 TI - Increasing sample throughput on a standard 36-lane model 377 DNA sequencer to 48 or 64 samples per run. PMID- 10524299 TI - Inverse single-strand RACE: an adapter-independent method of 5' RACE. PMID- 10524300 TI - Modified inverse PCR method for cloning the flanking sequences from human cell pools. PMID- 10524301 TI - Detection of frame-shifts within homopolymeric DNA tracts using the amplification refractory mutation system (ARMS). PMID- 10524302 TI - Empirical midpoint dissociation temperature (Td) determination for oligonucleotide probes using a PCR thermal cycler. PMID- 10524303 TI - Synthesis of radioactive single-stranded DNA probes using asymmetrical PCR and oligonucleotide random priming. PMID- 10524304 TI - Simple identification of mutant clones. PMID- 10524305 TI - Cloning of transfected cells without cloning rings. PMID- 10524306 TI - Tetracycline-resistant gene cassette designed for construction of mutant libraries of a target gene. PMID- 10524307 TI - Estimation of viable cell count after fluorescein diacetate staining using phosphorimager analysis. PMID- 10524308 TI - Cloning and purification of bacteriophage K11 RNA polymerase. PMID- 10524309 TI - Direct fluorescence-based lipase activity assay. PMID- 10524310 TI - Convenient method to improve the graphical quality of phylogenetic trees computed by the MEGA program. PMID- 10524311 TI - Enhanced sensitivity of a modified protein truncation test for exons 1-14 of the adenomatous polyposis coli gene. PMID- 10524312 TI - Internet training for biologists on BioMOO. PMID- 10524313 TI - Micro-perfusion flow cell for imaging cultured cells. AB - We present a unique design for a flow cell with a small working volume that allows rapid displacement of media viewed under high power and short working distance objectives. The flow cell has a small internal depth (ca. 0.033 cm) and volume (ca. 0.05 mL) and is easy to handle. Made of Delrin, the flow cell is biologically inert. We have used the flow cell for fluorescence imaging of PC12 cells loaded with tetramethylrhodamine dextran (TMRD) and other dyes. PMID- 10524314 TI - Co-transfected SV40 origin of replication activates expression from SV40 promoterless constructs. AB - Co-transfection with expression plasmids is widely used to control DNA uptake efficiency in transient transfection experiments. However, a number of problems have been associated with their use. Here, we describe the activation of expression of constructs not containing the simian virus 40 (SV40) origin of replication (ori) by co-transfection in COS-7 cells with plasmids containing the SV40 ori. This effect has consequences for the use of such plasmids to control transfection efficiency. PMID- 10524315 TI - Site-directed mutagenesis using uracil-containing double-stranded DNA templates and DpnI digestion. AB - DpnI can cleave fully methylated parental DNA while leaving hemi-methylated DNA intact. Based on this observation, we developed a rapid site-directed mutagenesis method using uracil-containing, double-stranded (ds)DNA templates and DpnI digestion. A 38% mutation efficiency was achieved by DpnI treatment of the mutagenic strand-extension reaction, and it increased to 70%-91% when uracil containing dsDNA templates were used. This method compares favorably to the most efficient current methods, but is simpler and does not require the use of single stranded templates or phage vectors. PMID- 10524316 TI - Homogeneous noncompetitive immunoassay based on the energy transfer between fluorolabeled antibody variable domains (open sandwich fluoroimmunoassay). AB - The antigen-dependent stabilization of an anti-hen egg lysozyme (HEL) antibody HyHEL-10 variable region was monitored with fluorescence resonance energy transfer (FRET) between fluorolabeled heavy chain (VH) and light chain (VL) fragments. The VH and VL fragments labeled with succinimide esters of fluorescein and rhodamine-X, respectively, were mixed in a cooled cuvette, and the change in fluorescence spectra upon antigen addition was monitored. When excited at 490 nm, significant decrease in the fluorescence at 520 nm and its increase at 605 nm were observed when an increasing amount of HEL was added to the mixture in the concentration range of 1-100 micrograms/mL. The assay, named open sandwich fluoroimmunoassay (FIA), is noncompetitive and homogeneous and can be conducted with one clone of antibody. With the use of appropriate antibodies, it is thought to be a quick and inexpensive alternative to the conventional laborious and/or expensive immunoassays. PMID- 10524318 TI - Changing functionality of surfaces by directed self-assembly using oligonucleotides--the Oligo-Tag. AB - A method is presented to modify surfaces for biotechnological applications. Oligonucleotides have been coupled covalently to a pre-activated surface. Complementary oligonucleotides hybridize to the surface, which are conjugated with functionalities. The oligonucleotides serve as "Oligo-Tags" for these functionalities that now are linked specifically and reversibly. The approach might be used to change DNA-arrays into arrays of arbitrary ligands. We demonstrate the method with an optical wave guide grating coupler as a sensing surface using two different haptens as examples for a variety of functionalities. The haptens were 2,4-dichlorophenoxyacetic acid and atrazin and are recognized by specific antibodies. The surface created was completely regenerable by alkaline washing or temperature increase without any loss of binding capacity. Specificity was demonstrated by competitive binding of antibody in presence and absence of analyte; unspecific binding has not been observed. PMID- 10524317 TI - Quantification of 5-methylcytosine in DNA by the chloroacetaldehyde reaction. AB - The study of changes in genome-wide levels of DNA methylation has become a key focus for understanding the epigenetic regulation of gene expression. Many procedures exist to study DNA methylation, falling into two categories: gene specific and genome-wide. Genome-wide methylation analysis is best performed by DNA hydrolysis followed by HPLC; however, it requires access to an HPLC machine, which is not always available. Alternative procedures, such as the radioactive labeling of CpG sites using SssI DNA methyltransferase, have been developed to address this problem, but it can only monitor CpG methylation changes, and CpNpG methylation is not detected. Here, we present a method for the analysis of DNA methylation in any sequence context by fluorescent labeling. We present control analyses using synthetic oligonucleotides of known methylation levels and a comparison of genomic DNA from two transgenic tobacco lines known to differ in their methylation levels. The results indicate that hygromycin-induced hypermethylation acts equally on all classes of methylatable cytosine, perhaps indicating a common mechanism. PMID- 10524319 TI - Quantitative ratio of primer pairs and annealing temperature affecting PCR products in duplex amplification. AB - The quantity of PCR products that are simultaneously amplified from two different loci in a duplex amplification (DA) are significantly lower for one of the loci, as compared to identical PCR amplification in separate single-band amplifications (SBA). This difference in amplification probably occurs already after the second cycle of amplification. To further analyze this phenomenon, we tested different reaction conditions, including annealing times, a wide range of temperatures, various quantities of the template, several nucleotide concentrations, different amounts of TaqI DNA Polymerase, number of amplification cycles and various amounts of primers and primers ratio. Changing the ratio between the sets of primers in DA had the most significant effect on the relative levels of amplification of the loci with an optimal ratio of 4:1 in favor of the set of primers used to amplify the underrepresented fragment. The optimal annealing temperatures for the tested sets of primers were identical in SBA and different in DA. Possible reasons for this phenomenon are discussed. PMID- 10524320 TI - Budding yeast as a screening tool for discovery of nucleoside analogs for use in HSV-1 TK suicide-gene therapy. AB - We present a fast, convenient and inexpensive method that allows the automated, large-scale screening of chemical libraries for compounds that are converted by the herpes simplex virus type 1 (HSV-1) thymidine kinase (TK) into inhibitors of cell growth. The method is based on the use of budding yeast (Saccharomyces cerevisiae) transformed with the HSV-1 TK gene on a multicopy plasmid. Eight nucleoside analogs (acyclovir, ganciclovir, penciclovir, lobucavir, brivudin, sorivudine, IVDU and ara-T), for which the cytostatic action against mammalian cells expressing the HSV-1 TK gene has been well documented, were studied for their inhibitory effect on the growth of yeast expressing the viral TK. These nucleoside analogs had little or no inhibitory effect on the growth of yeasts transformed with the empty vector, but inhibited to a significant extent the growth of yeast expressing the viral TK. Use of HSV-1 TK-expressing yeast allows quick screening in multi-well plate format for compounds with potential use in HSV-1 TK suicide gene therapy. The method may also be used as a tool to selectively suppress or arrest the growth of one population of yeast out of mixed yeast cell cultures. PMID- 10524322 TI - Identification of ribonucleoprotein (RNP)-specific protein interactions using a yeast RNP interaction trap assay (RITA). AB - We describe an adaptation of the yeast three-hybrid system that allows the reconstitution in vivo of tripartite (protein-RNA-protein) ribonucleoproteins (RNPs). To build and try this system that we called RNP interaction trap assay (RITA), we used as a model the autoantigenic Ro RNPs. hY RNAs bear distinct binding sites for Ro60 and La proteins, and Ro RNPs are thus physiologically tripartite (Ro60/hY RNA/La). Using recombinant La (rLa) and Ro60 (rRo60) proteins and recombinant hY RNAs (rhY) co-expressed in yeast, we found that RNPs made of rRo60/rhY/rLa were readily reassembled. Reconstitution of tripartite RNPs was critically dependent on the presence of an appropriate Ro60 binding site on the recombinant RNA. The RITA assay was further used to detect (rRo60/rhY RNP) binding proteins from a HeLa cell cDNA library, allowing specific identification of La and of a novel Ro RNP-binding protein (RoBPI) in more than 70% of positive clones. RITA assay may complement already available two- and three-hybrid systems to characterize RNP-binding proteins by allowing the in vivo identification of interactions strictly dependent upon the simultaneous presence of a protein and of its cognate RNA. PMID- 10524321 TI - High-throughput microarray-based enzyme-linked immunosorbent assay (ELISA). AB - A new generation biochip is described as capable of supporting high-throughput (HT), multiplexed enzyme-linked immunosorbent assays (ELISAs). These biochips consist of an optically flat, glass plate containing 96 wells formed by an enclosing hydrophobic Teflon mask. The footprint dimensions of each well and the plate precisely match those of a standard microplate. Each well contains four identical 36-element arrays (144 elements per well) comprising 8 different antigens and a marker protein. Arrays are formed by a custom, continuous flow, capillary-based print head attached to a precise, high-speed, X-Y-Z robot. The array printing capacity of a single robot exceeds 20,000 arrays per day. Arrays are quantitatively imaged using a custom, high-resolution, scanning charge coupled device (CCD) detector with an imaging throughout of 96 arrays every 30 s. Using this new process, arrayed antigens were individually and collectively detected using standard ELISA techniques. Experiments demonstrate that specific multiplex detection of protein antigens arrayed on a glass substrate is feasible. Because of the open microarray architecture, the 96-well microarray format is compatible with automated robotic systems and supports a low-cost, highly parallel assay format. Future applications of this new high-throughput screening (HTS) format include direct cellular protein expression profiling, multiplexed assays for detection of infectious agents and cancer diagnostics. PMID- 10524323 TI - Automated detection of point mutations by electrophoresis in peptide-nucleic acid containing gels. AB - Polymerization of electronically, essentially neutral peptide nucleic acids (PNA) into polyacrylamide gels creates a medium in which the salt-independent properties of PNA/DNA interactions are used to achieve hybridization with target DNA during affinity electrophoresis. Such physical entrapment of PNA has been used to differentiate between a retarded, complementary DNA strand and a non retarded sequence differing by a single point mutation. Analysis of fluorescent PCR products--from both model mismatches and clinically relevant point mutations using a conventional automated DNA sequencer--allows one to follow this hybridization in real time and to distinguish homo- and heterozygous mutants visually. It has been shown that parameters affecting the resolution of these species include not only temperature and concentration (as a function of the [GC] content of the PNA), but also the position of the PNA binding sequence within the interacting DNA segment. Under conditions optimized in terms of temperature and PNA concentration, the maximum separation of retarded from non-retarded DNA single strands is obtained when the PNA binding sequence is close to either DNA terminus. Strategies of PNA and PCR primer design that permit a diagnostic application ranging over 85-451 DNA base pairs are proposed. PMID- 10524324 TI - Automated purification and quantification of oligonucleotides. AB - We have developed automated methods for the trityl-on purification and quantification of synthetic oligonucleotides. Oligonucleotide purification is by solid-phase extraction cartridges using Amberchrom CG-50 resin on an XYZ-axis robotic system. Quantification is by OD260nm using an online UV-visible spectrophotometer with sipper. The purification of 20 oligonucleotides requires 5 min of user set-up time, plus 20 min per sample of robot time. For a 15-25-mer at the 40 nmol scale of synthesis, the method gives a yield of 2.8 ODs from a load of 10.1 OD, i.e., a 28% average yield. Oligonucleotides purified by this method have proven to be successful for primers for automated DNA sequencing. PMID- 10524325 TI - Evaluation of methods for transient transfection of a murine macrophage cell line, RAW 264.7. AB - Monocyte/macrophage cell lines are fastidious cells commonly used in transient transfection experiments. In the course of a study of gene regulation by lipopolysaccharide (LPS), we have compared several methods for DNA-mediated cell transfection to determine which would be optimally applicable to the macrophage line, RAW 264.7. Both the response level (LPS inducibility) and the degree of inter-assay variation were evaluated for each transfection technique. The following methods were compared: Lipofectin, LipofectAMINE, LipofectAMINE PLUS, SuperFect, Ca3(PO4)2 DNA co-precipitation, DEAE dextran-mediated transfection and electroporation. The transfected plasmid DNA included a luciferase reporter construct containing the junB minimal promoter under the control of an LPS inducible 1300-bp regulatory fragment downstream of junB 5'-flanking sequence, as well as a beta-galactosidase reporter construct under the adenovirus promoter and enhancer used as an internal control. Electroporation, followed by a resting period of 16-24 h before stimulation with LPS, had the highest inducibility of all methods. DEAE dextran and Ca3(PO4)2 precipitation showed the least and the greatest inter-assay variation, respectively. For all other methods, inter-assay variability fell within this range. The results presented may serve as both a general reference and a guide for reporter gene studies in this or other macrophage cell lines. PMID- 10524326 TI - RecA-mediated affinity capture: a method for full-length cDNA cloning. AB - We describe an improved method for rapid cloning of full-length cDNA from cDNA libraries. This approach is based on the ability of Escherichia coli RecA protein to form a stable nucleoprotein complex with a linear single-stranded DNA probe and homologous sequences in circular double-stranded DNA. Hybridization of RecA coated biotinylated DNA probes to homologous plasmid DNA creates triple-stranded complexes, which are then captured on streptavidin-coated magnetic beads. Following magnetic separation of the hybrid molecules, the enriched plasmid population is recovered by alkaline treatment, precipitated, resuspended and used to transform bacteria. Typically, many clones can then be recovered by colony hybridization screening of a single plate of the enriched library. We have used this technology to clone full-length and alternatively spliced forms of the human bcl-xL cDNA from a human liver cDNA library. PMID- 10524327 TI - Detection of DNA damage and identification of UV-induced photoproducts using the CometAssay kit. AB - We introduce the first commercially available comet assay for the detection and quantification of DNA damage in individual eukaryotic cells. The major difficulty of the comet assay is the preparation of the slides needed to immobilize the samples throughout the lysis and electrophoretic procedures. The CometAssay kit uses a proprietary technology to precoat glass microscope slides to allow direct application of the agarose embedded sample without any additional slide treatment. In this report, we discuss the detection of DNA damage in individual cells exposed to ultraviolet irradiation using the new CometSlides and their cost compared to traditional slides. PMID- 10524328 TI - Bacterial interference. AB - Bacterial interactions, antagonistic and synergistic, help maintain the balance in the normal endogenous flora. The production of bacteriocins by microorganisms is one of the important mechanisms used for interference. The ability of various microorganisms to produce bacteriocins and exhibit interfering capability is detailed in the review. These organisms include Gram-positive and Gram-negative aerobic and anaerobic bacteria. The role of bacterial interference (BI) in clinical infections and the effect of this phenomenon on their eradication is detailed. The infections discussed include those of the upper respiratory (pharyngo-tonsillitis, otitis media), urogenital, and gastrointestinal tracts. The influence of antimicrobial agents on these organisms and their interactions with other bacteria are also described. PMID- 10524329 TI - Cuban allegations of biological warfare by the United States: assessing the evidence. PMID- 10524330 TI - Mycopesticide production by fermentation: potential and challenges. AB - The agriculture industry is in need of novel biopesticides and development of large-scale production of mycopesticide, either fungal cells themselves or cell free fungal components. The identification of a fungal strain with pesticide activity, and its improvement, is the primary step in developing infective propagules such as conidia, blastospores, chlamydospores, oospores, and zygospores as well as in preparing hydrolytic enzyme mixtures. This review discusses various parameters for submerged and solid state fermentation to produce fungal structures, particularly of mycoparasitic and entomopathogenic species that are prospective candidates for use as mycopesticides. The understanding of the molecular aspects of fungus-fungus and fungus-insect interactions, the role of hydrolytic enzymes especially chitinases in the killing process, and the possible use of chitin synthesis inhibitors are the prime areas of research aimed at making fungi more effective either singly or in combination as mycopesticides. PMID- 10524331 TI - An improved amplification system for the production of Endo F3. PMID- 10524332 TI - Two systems of giant axon terminals in the cat medial geniculate body: convergence of cortical and GABAergic inputs. AB - The thalamus plays a critical role in processing sensory information that involves interactions between extrinsic connections and intrinsic circuitry. Little is known regarding how these different systems might interact. We found an unexpected nuclear convergence of two types of giant axon terminals, each of which must have independent origins, in the dorsal division of the cat medial geniculate body. The first class of giant terminal was labeled after injections of biotinylated dextran amines (BDA) in seven auditory cortical areas. A second type was found in sections immunostained for gamma-aminobutyric acid (GABA); these endings had the same nuclear distribution, and they were numerous. The origin of this GABAergic terminal is unknown. The giant corticothalamic terminals were presumably those described in prior accounts using different tracers (Rouiller and de Ribaupierre [1990] Neurosci. Lett. 208:29-35; Ojima [1994] Cerebral Cortex 6:646-663), but with BDA they are labeled more fully. Clusters of such endings were often linked, and hundreds may occur in a single section. Their boutons formed a substantial proportion of the corticothalamic population. Other types of corticogeniculate axon terminals were also labeled, including two kinds that are much smaller and that match closely the classical descriptions of corticothalamic axons. The giant GABAergic endings were found in all dorsal division nuclei and in thalamic visual nuclei such as the lateral posterior nucleus. Like the giant cortical endings, the giant GABAergic terminals often encircled large, pale, immunonegative profiles that may be dendritic. This implies a close spatial, and perhaps a close functional, relationship between the populations of giant axon terminals. Insofar as physiological studies found that pharmacological inactivation of rat somatic sensory cortex suppresses peripheral information transmission through the posterior thalamus, corticofugal input may be essential for normal processing (Diamond et al. [1992] J. Comp. Neurol. 319:66 84). Our findings suggest that the giant corticothalamic endings could play an important role in descending control. Perhaps they are counterbalanced by a GABAergic system and affect thalamic oscillations implicated in shifts in vigilance and attention. PMID- 10524333 TI - Apoptosis during development of the human retina: relationship to foveal development and retinal synaptogenesis. AB - Apoptosis in the ganglion cell (GCL) and inner nuclear (INL) layers of human fetal retinae aged 14-35 weeks of gestation (WG) was investigated in relation to synaptogenesis and foveal depression formation. Terminal transferase dUTP-biotin nick end labeling (TUNEL) was used to identify apoptosis, and synapse development was demonstrated by synaptophysin immunoreactivity (-IR). The distribution of apoptotic cells and synaptophysin-IR was studied as a function of eccentricity. Between 14 and 23-24 WG in the GCL, rates of apoptosis were relatively low in central retina. A shallow fovea was detected at 23-24 WG. In the central GCL, the rate of apoptosis was 0.21% of viable cells compared with a higher incidence of 0.79-1.64% peripherally. Apoptosis in the INL was 2-8 times greater than that in the GCL. At 14-15 WG, peak death occurred at the incipient fovea; however, by 20 WG the distribution was bimodal, with peaks at more eccentric locations on either side of the incipient fovea with increasing age. Approximately 90% of INL apoptotic cells were in the middle and outer regions, suggesting that bipolar cells formed the majority of dying neurons. Synaptophysin-IR was present in cones, bipolar cells, and processes in the inner and outer plexiform layers at the incipient fovea at 14 WG and spread peripherally with increasing age. The peripheral margin of synaptophysin-IR coincided with areas of peak INL apoptosis. This pattern suggests that bipolar cell elimination is associated with the onset of synaptogenesis. Apoptosis in the GCL and INL is not a significant factor in foveal depression morphogenesis. PMID- 10524334 TI - Antigenic epitopes of the photoreceptor synaptic ribbon. AB - The purpose of this study was twofold: 1) to purify and identify a protein containing an epitope recognized by an anti-synaptic ribbon antibody B16 and 2) to identify and sequence the epitope. B16 recognizes several unrelated proteins in retina immunoblots. Purification and microsequencing of the strongest band (88 kDa) demonstrate 94% identity to aconitase over 111 amino acids. Polyclonal antibodies against aconitase recognize aconitase on Western blots, but not synaptic ribbons in sections. We conclude that although aconitase contains the epitope, aconitase is not the synaptic ribbon protein. The B16 epitope was identified to be 542DTYQHPPKDS551. A synthetic peptide to this sequence absorbs B16 activity in both Western blots and immunohistochemistry studies, whereas partial peptides fail to absorb activity. Additional antibodies against this peptide label synaptic ribbons. When mouse retina were double labeled with B16 and anti-alpha-actinin, B16 was found to label synaptic ribbons in the outer plexiform layer that partially enclosed the alpha-actinin label. We have determined the amino acid sequence of the B16 epitope and found that the B16 labeling colocalizes with alpha-actinin at the photoreceptor synapse. PMID- 10524335 TI - Subcellular distribution of 5-hydroxytryptamine2A and N-methyl-D-aspartate receptors within single neurons in rat motor and limbic striatum. AB - The dorsolateral caudate-putamen nucleus (CPN) and the nucleus accumbens (NAc) shell, respectively, are involved in many motor and limbic functions that are affected by activation of the 5-hydroxytryptamine2A receptor (5HT2AR) and the N methyl-D-aspartate subtype of glutamate receptor (NMDAR). We examined the functional sites for 5HT2AR activation and potential interactions involving the NMDAR subunit NR1 (NMDAR1) within these striatal regions. For this examination, sequence-specific antipeptide antisera against these receptors were localized by electron microscopic dual-labeling immunocytochemistry in the rat brain. In the dorsolateral CPN and the NAc shell, the 5HT2AR-labeled profiles were mainly dendrites, but somata and axons were also immunoreactive. The neuronal somata contained round unindented nuclei that are typical of spiny striatal neurons, although few dendritic spines were 5HT2AR immunolabeled. In all neuronal profiles, the 5HT2AR labeling was primarily associated with cytoplasmic organelles and more rarely was localized to synaptic or nonsynaptic plasma membranes. Colocalization of 5HT2AR and NMDAR1 was seen primarily in somata and dendrites. Significantly greter numbers of 5HT2AR- or 5HT2AR- and NMDAR1 containing dendrites were seen in the dorsolateral CPN than in the NAc shell. As compared with 5HT2AR, NMDAR1 labeling was more often observed in dendritic spines, and these were also more numerous in the CPN. These results indicate that 5HT2A and NMDA receptors are coexpressed but differentially targeted in single spiny striatal neurons and are likely to play a major role in control of motor functions involving the dorsolateral CPN. PMID- 10524336 TI - Ultrastructural localization of neuropeptide Y and expression of its mRNA in the glomus cells distributed in the wall of the common carotid artery of the chicken. AB - In the chicken, glomus cells are widely distributed in the carotid body and in the wall of the common carotid artery and around its branches. The cells located in the wall of the common carotid artery express intense immunoreactivity for neuropeptide Y (NPY). They contain abundant dense-cored vesicles ranging from 70 to 220 nm in diameter. In this study, we examined ultrastructural localization of NPY in the glomus cells by using the postembedding immunogold method. Gold particles representing immunoreactivity for NPY were specifically localized on the dense-cored vesicles of the glomus cells. In addition, the localization of NPY mRNA in the glomus cells was examined by in situ hybridization with digoxigenin-labeled chicken NPY cRNA probe. A strong hybridization signal for NPY mRNA was detected in the glomus cells located in the wall of the common carotid artery. Few glomus cells of the carotid body, however, displayed labeling for NPY mRNA. Northern blot analysis with the chicken NPY exon 4 probe demonstrated that a single band for NPY mRNA was present in the poly (A) + RNA isolated from the common carotid artery where the glomus cells were distributed. Furthermore, the expression of NPY mRNA in the common carotid artery was confirmed by the reverse transcription-polymerase chain reaction. These results indicate that the chicken glomus cells are able to produce NPY but that the level of its translation varies according to the location of the cells. PMID- 10524337 TI - Ultrastructural localization of the corticotropin-releasing factor-binding protein in rat brain and pituitary. AB - Preembedding immunoperoxidase staining methods were used to permit ultrastructural analyses of the distribution in rat brain and pituitary of the corticotropin-releasing factor-binding protein (CRF-BP), a moiety distinct from CRF receptors, but which is nonetheless capable of binding the peptide and reversibly neutralizing its biological actions. In anterior pituitary, CRF-BP immunoreactivity (ir) was detected in corticotropelike cells, with reaction product associated principally with secondary lysosomes and multivesicular bodies and not at all with secretory granules. In brain, marked regional differences in the subcellular pattern of CRF-BP staining were evident. In isocortex, where BP/peptide colocalization is rare, BP-ir was distributed in cells and processes in a manner similar to that of a prototypic neuropeptide, including in terminals commonly engaging in synaptic contacts with unlabeled dendritic profiles. In the bed nucleus of the stria terminalis, a site that contains overlapping accumulations of CRF-BP-ir projections and CRF-ir perikarya, BP staining was restricted to vesicle-laden varicosities that rarely engaged in synaptic contacts with somatic or dendritic elements but were frequently apposed to unlabeled axon varicosities and terminals. In the ventromedial medulla, a site of partial CRF/BP overlap, most cells displayed a subcellular localization CRF-BP-ir like that seen in cortex, whereas in others the distribution shared similarities with that observed in pituitary. The results suggest that the function of the CRF-BP may differ in different cellular contexts. In cellular targets of CRF or in neurons in which peptide and BP coexist, the CRF-BP may play a role in processing and degradation of CRF and/or ligand-receptor complexes. In other areas of the central nervous system, the BP seems positioned to serve as a transmitter/modulator at conventional synapses or as an autocrine or paracrine modulator of local CRF effects. PMID- 10524338 TI - Localization of GABA-like immunoreactivity in the central nervous system of Aplysia californica. AB - Gamma-aminobutyric acid (GABA) is present in the central nervous system of Aplysia californica (Gastropoda, Opisthobranchia) where its role as a neurotransmitter is supported by pharmacological, biochemical, and anatomical investigations. In this study, the distribution of GABA-immunoreactive (GABAi) neurons and fiber systems in Aplysia was examined by using wholemount immunohistochemistry and nerve backfill methods. GABAi neurons were located in the buccal, cerebral, and pedal ganglia. Major commissural fiber systems were present in each of these ganglia, whereas more limited fiber systems were observed in the ganglionic connectives. Some of the interganglionic fibers were found to originate from two unpaired GABAi neurons, one in the buccal ganglion and one in the right pedal ganglion, each of which exhibited bilateral projections. No GABAi fibers were found in the nerves that innervate peripheral sensory, motor, or visceral organs. Although GABAi cells were not observed in the pleural or abdominal ganglia, these ganglia did receive limited projections of GABAi fibers originating from neurons in the pedal ganglia. The distribution of GABAi neurons suggests that this transmitter system may be primarily involved in coordinating certain bilateral central pattern generator (CPG) systems related to feeding and locomotion. In addition, the presence of specific interganglionic GABAi projections also suggests a role in the regulation or coordination of circuits that produce components of complex behaviors. PMID- 10524339 TI - Ontogenetic expression of trk neurotrophin receptors in the chick auditory system. AB - Neurotrophins and their cognate receptors are critical to normal nervous system development. Trk receptors are high-affinity receptors for nerve-growth factor (trkA), brain-derived neurotrophic factor and neurotrophin-4/5 (trkB), and neurotrophin-3 (trkC). We examine the expression of these three neurotrophin tyrosine kinase receptors in the chick auditory system throughout most of development. Trks were localized in the auditory brainstem, the cochlear ganglion, and the basilar papilla of chicks from embryonic (E) day 5 to E21, by using antibodies and standard immunocytochemical methods. TrkB mRNA was localized in brainstem nuclei by in situ hybridization. TrkB and trkC are highly expressed in the embryonic auditory brainstem, and their patterns of expression are both spatially and temporally dynamic. During early brainstem development, trkB and trkC are localized in the neuronal cell bodies and in the surrounding neuropil of nucleus magnocellularis (NM) and nucleus laminaris (NL). During later development, trkC is expressed in the cell bodies of NM and NL, whereas trkB is expressed in the nerve calyces surrounding NM neurons and in the ventral, but not the dorsal, dendrites of NL. In the periphery, trkB and trkC are located in the cochlear ganglion neurons and in peripheral fibers innervating the basilar papilla and synapsing at the base of hair cells. The protracted expression of trks seen in our materials is consistent with the hypothesis that the neurotrophins/tyrosine kinase receptors play one or several roles in the development of auditory circuitry. In particular, the polarized expression of trkB in NL is coincident with refinement of NM terminal arborizations on NL. PMID- 10524340 TI - Glomerular formation in the developing rat olfactory bulb. AB - Using the confocal microscope together with markers for the cellular components of glomeruli, we examined the spatiotemporal cellular interactions that occur between the axons of olfactory receptor cells, their dendritic targets, and glial cells during the critical period of glomerular formation. We have employed markers of immature and mature olfactory receptor cell axons, mitral/tufted cell dendrites, and glial cells as well as a synapse-associated protein for double- and triple-label immunocytochemistry. Axons of olfactory receptor cells grew into a dense dendritic zone of the olfactory bulb (comprising the dendrites of both mitral and tufted cells) between E17 and E18. At E19, these axons coalesced into protoglomeruli, which continued to develop until birth, when the basic anatomical structure of adult glomeruli emerged. Neither mitral/tufted cell dendrites nor olfactory bulb astrocytes became specifically associated with these protoglomeruli until E21. Ensheathing cells remained restricted to the outer nerve fiber layer and did not appear to contribute to glomerular formation. Finally, the synaptophysin staining has shown that synaptic constituents are expressed as early as E17, prior to the appearance of mature olfactory receptor cell axons. Based on these data, we have established a time line detailing the temporal and spatial interactions that occur between cell types during late embryonic rat olfactory bulb development. We conclude that the initial event in the formation of glomeruli is the penetration of the mitral/tufted cell dendritic zone by olfactory receptor cell axons. The coalescence of dendritic and glial processes into glomerular structures appears secondary to the arrival of the olfactory receptor cell axons. PMID- 10524341 TI - The shapes and numbers of amacrine cells: matching of photofilled with Golgi stained cells in the rabbit retina and comparison with other mammalian species. AB - Amacrine cells of the rabbit retina were studied by "photofilling" a photochemical method in which a fluorescent product is created within an individual cell by focal irradiation of the nucleus; and by Golgi impregnation. The photofilling method is quantitative, allowing an estimate of the frequency of the cells. The Golgi method shows their morphology in better detail. The photofilled sample consisted of 261 cells that were imaged digitally in through focus series from a previous study (MacNeil and Masland [1998] Neuron 20:971 982). The Golgi material consisted of 49 retinas that were stained as wholemounts. Eleven of these subsequently were cut in vertical section. Of the many hundreds of cells stained, digital through-focus series were recorded for 208 of the Golgi-impregnated cells. The two methods were found to confirm one another: Most cells revealed by photofilling were recognized easily by Golgi staining, and vice versa. The greater resolution of the Golgi method allowed a more precise description of the cells and several types of amacrine cell were redefined. Two new types were identified. The two methods, taken together, provide an essentially complete accounting of the populations of amacrine cells present in the rabbit retina. Many of them correspond to amacrine cells that have been described in other mammalian species, and these homologies are reviewed. PMID- 10524342 TI - Anatomical evidence supporting the potential for modulation by multiple neurotrophins in the majority of adult lumbar sensory neurons. AB - Neurotrophins exert effects on sensory neurons through receptor tyrosine kinases (trks) and a common neurotrophin receptor (p75). Quantitative in situ hybridization studies were performed on serial sections to identify neurons expressing single or multiple neurotrophin trk receptor mRNA(s) in adult lumbar dorsal root ganglion (DRG) in order to examine the possibility of multi neurotrophin modulation of phenotype via different trk receptors or various trk isoforms. Expression of mRNA encoding trkA, trkB, trkC, or p75 is restricted to select subpopulations representing approximately 41%, 33%, 43%, and 79% of DRG neurons, respectively. Colocalization studies reveal that approximately 10% of DRG neurons coexpress trkA and trkB mRNA; 19% coexpress trkA and trkC mRNA; and 18% coexpress trkB and trkC mRNA. Trilocalization of all three trk mRNAs is rare, with approximately 3-4% of neurons in this category. Overall incidence of expression of more than one full length trk mRNA occurs in approximately 40% of DRG neurons, whereas expression of individual trk mRNA is found in approximately 34%. Full length trk receptor mRNA is rarely detected without p75, implicating the latter in neuronal response to neurotrophins. Examination of two full-length isoforms of trkA reveal that they are coexpressed with relative levels of expression positively correlated. TrkC mRNAs corresponding to 14- or 39-amino acid insert isoforms colocalize with the non-insert trkC isoform, but the converse is not necessarily true. The data suggest that substantial subpopulations of adult sensory neurons may be modulated through interactions with multiple neurotrophins, the consequences of which are largely unknown. PMID- 10524343 TI - Topographic organization of serotonergic dorsal raphe neurons projecting to the superior colliculus in the Mongolian gerbil (Meriones unguiculatus). AB - Recent evidence suggests that the dorsal raphe nucleus (DRN) of the brainstem is a collection of neuronal clusters having different neurochemical characteristics and efferent projection patterns. To gain further insight into the neuroanatomic organization of the DRN, neuronal populations projecting to the superior colliculus (SC) were mapped in a highly visual rodent, the Mongolian gerbil (Meriones unguiculatus). Retrograde tracers Fluoro-Gold (FG) or cholera toxin subunit-B (CTB) were injected into the superficial layers of the SC, and serotonin (5-hydroxytryptamine, 5-HT) -positive cells were identified by using immunocytochemistry in the FG-injected animals. Based on its projections to the SC, the DRN was divided into five rostrocaudal levels. In the rostral and middle levels of the DRN, virtually all FG-filled cells occurred in the lateral DRN, and 36-55% of 5-HT-immunoreactive (5-HT-ir) cells were also double-labeled with FG. Caudally, FG-filled cells occurred in the lateral, ventromedial, and interfascicular DRN; and 44, 12, and 31% of 5-HT-ir cells, respectively, were also FG-filled. The dorsomedial DRN contained only a small proportion of FG filled cells at its most caudal level and was completely devoid of FG-filled cells more rostrally. The CTB-injected animals showed a similar distribution of retrogradely labeled cells in the DRN. Topographically, the dorsal tegmental nucleus and the laterodorsal tegmental nucleus appeared to be closely associated with 5-HT-ir cells in the caudal DRN. These results suggest that the lateral DRN and the ventromedial/interfascicular DRN may be anatomically, morphologically, and neurochemically unique subdivisions of the gerbil DRN. PMID- 10524344 TI - Comparison of 26,27-hexafluoro-1 alpha,25-dihydroxyvitamin D3 and 1 alpha,25 dihydroxyvitamin D3 on the resorption of bone explants ex vivo. AB - 26,27-Hexafluoro-1 alpha,25-dihydroxyvitamin D3 [F6-1,25-(OH)2D3] is more potent than 1 alpha,25-dihydroxyvitamin D3 [1,25(OH)2D3] in stimulating bone resorption in vitro and in vivo. The reason why F6-1,25(OH)2D3 is more active remains unclear. To clarify the relationship between the bone-resorbing activity of each vitamin D3 analogue and the metabolism of each analogue, in the present study, we used an ex vivo method that was established by Reynolds et al (Calcif Tissue Res, 1974, 15, 333-339). The effect of F6-1,25(OH)2D3 or 1,25(OH)2D3 on 45Ca release from parietal bones, prepared at 3, 14 and 24 h after injection of 1.9, 3.8, 7.6 or 15.2 pmol vitamin D analog/g body weight, was examined. F6-1,25(OH)2D3 was more potent than 1,25(OH)2D3 during each in vivo time period. 1,25(OH)2D3 at 3 h after the injection was more active compared to the control (no injection of 1,25(OH)2D3) but not at 14 and 24 h. The radioactivity of the bones after the injection of [3H]-F6-1,25(OH)2D3 was retained even at 24 h. In the case of [3H] 1,25(OH)2D3, the radioactivity of bones decreased with an increase in the in vivo period. In a HPLC analysis of the lipid extract of bone homogenate, [3H]-F6 1,25(OH)2D3 alone was detected at 3 h after the injection and both [3H]-F6 1,25(OH)2D3 and [3H]-26,27-hexafluoro-1 alpha, 23S,25-trihydroxyvitamin D3 [F6 1,23,25(OH)3D3] were detected at 14 and 24 h after the injection. [3H] 1,25(OH)2D3 was highly detected at 3 h after the injection, but it decreased with an increase in the in vivo period. In the ex vivo test, the activity of F6 1,23,25(OH)3D3 was less than that of F6-1,25(OH)2D3 but similar to that of 1,25(OH)2D3. The present study indicates that F6-1,25(OH)2D3 is more active and more long-lasting than 1,25(OH)2D3 in the ex vivo method. A higher potency of F6 1,25(OH)2D3 is explained, at least partly, by the results that the amounts of both F6-1,25(OH)2D3 and its active metabolite, F6-1,23,25(OH)3D3, in the bones are higher than that of 1,25(OH)2D3, and that F6-1,25(OH)2D3 and its metabolite are retained in bones longer than 1,25(OH)2D3. PMID- 10524345 TI - Curdlan and gellan gum, bacterial gel-forming polysaccharides, exhibit different effects on lipid metabolism, cecal fermentation and fecal bile acid excretion in rats. AB - The effects of curdlan (CD) and gellan gum (GG), bacteria-producing polysaccharides, on lipid concentrations of serum and liver, fecal bile acid composition and intestinal fermentation products were studied in rats fed diets containing cellulose powder (CP), CD or GG at 5% for 4 wk. The cecal weight of the CD group increased significantly as compared to that of the other two groups and the pH of its contents was significantly low. The gastrointestinal transit time in the GG group was significantly shorter than that in the CP and CD groups. No significant inter-group differences were observed in the serum concentrations of total cholesterol and HDL-cholesterol, but a significant decrease was observed in the hepatic total cholesterol concentration of the CD group as compared to that of the CP and GG groups. No significant difference in the total bile acid excretion in feces was observed among the groups, but significantly low values were observed in the proportion of secondary bile acids in the CD group as compared to those of the CP and GG groups. Amounts of short-chain fatty acids (acetic, propionic and butyric acid) and lactic acid in the cecal contents were significantly higher in the CD group than in the other two groups. These results reveal that dietary CD is easily degraded and fermented by intestinal bacteria in the cecum and lowers cholesterol concentration in the liver, while dietary GG shortens the gastrointestinal transit time, suggesting the promotion of evacuation. PMID- 10524346 TI - Increase of the cellular growth of old human diploid fibroblasts by radical scavenger: methanolic extract of broad beans. AB - The methanolic extract from broad beans (MEBB), which is comprised of phenolic compounds, has free-radical scavenging activity. The effects of MEBB on cytosolic antioxidant enzymes and cell proliferation were examined in cultures of old (78 84% life-span completed) WI-38 human diploid fibroblasts. Because catechin is polyphenol and has radical scavenging activity, it was used as the control in experiments. We observed that MEBB increased cellular growth when added to the cell culture. In MEBB at 40 and 120 micrograms/mL, the cell proliferation increased by 14 and 27%, respectively, as compared to the control. In catechin, cell proliferation increased as well. Regarding cytosolic glutathione peroxidase (GSH-Px) activity, treatment of old cells with MEBB at 40 and 120 micrograms/mL resulted in decreases as compared to the control. In contrast, catechin showed no similarities to the modification of GSH-Px activity. Cytosolic SOD activity was increased by treatment with 40 micrograms/mL MEBB, and the activity showed a gradual decrease with increased MEBB concentrations. A similar trend occurred in the cells treated with catechin (4-20 microM). These results suggest that cytosolic antioxidant enzyme activities in old cells may be modulated by MEBB treatment. We conclude that there may be a relation between the optimum MEBB concentration for the increase of cellular growth and the MEBB concentration required to exhibit a decrease in GSH-Px activity. PMID- 10524348 TI - Dietary phospholipid-dependent reductions in gene expression and activity of liver enzymes in fatty acid synthesis in fasted-refed rats. AB - The effects of dietary soybean phospholipid, its hydrogenation product and safflower phospholipid on gene expression and the activity of hepatic enzymes in fatty acid biosynthesis were examined in fasted-refed rats. Phospholipid composition of soybean phospholipid and its hydrogenation product were the same, but the hydrogenation product contained negligible amounts of unsaturated fatty acids. Among phospholipid classes, lysophosphatidylcholine and phosphatidylinositol proportions were slightly higher in safflower phospholipid than in soybean phospholipid or its hydrogenation product. Rats were fasted for 2 d and refed a fat-free diet or a diet containing 4% fatty acids either as soybean oil or various phospholipid preparations for 3 d. Compared to the fat-free diet, the soybean oil diet only slightly decreased specific, but not total hepatic fatty acid synthetase and malic enzyme activity, and it was totally ineffective in modulating glucose 6-phosphate dehydrogenase and pyruvate kinase activity under our experimental conditions. The diets containing phospholipids, however, markedly decreased the activity of these enzymes. The extent of reduction was somewhat attenuated with hydrogenated soybean phospholipid as compared with soybean and safflower phospholipids. Dot and Northern blot hybridization using specific cDNA probes showed that, compared to a fat-free diet, diets containing phospholipids profoundly decreased the hepatic mRNA levels of enzymes in fatty acid synthesis. Soybean oil, however, only marginally affected these parameters. Hepatic mRNA levels for enzymes correlated well with enzyme activity. Dietary phospholipids therefore appear to have decreased enzyme activity in fatty acid synthesis primarily by suppressing the mRNA levels of these enzymes. Compared to soybean oil, hydrogenated soybean phospholipid is still effective in decreasing the activity and mRNA level of enzymes in fatty acid synthesis. Therefore, it is difficult to ascribe the potent physiological activity of phospholipid in reducing fatty acid synthesis entirely to polyunsaturated fatty acid moiety. PMID- 10524347 TI - Combined effects of ethanol and garlic on hepatic ethanol metabolism in mice. AB - The combined effects of ethanol and components in fresh garlic on ethanol metabolism were investigated in the livers of mice. Male, 11-wk-old C3H/HeNCrj mice were intragastrically administered 2 g ethanol/kg body weight after being administered fresh garlic juice for 8 d (garlic group), and changes in the concentrations of ethanol, acetaldehyde and acetate in the serum, and changes in the activity of hepatic enzymes related to ethanol metabolism in mice were examined. The increases in the concentrations of acetaldehyde and acetate in the serum after ethanol administration tended to be diminished following garlic administration. The microsomal ethanol-oxidizing system (MEOS) in the livers of the garlic groups was significantly lower than that of the control microsomes at 2 h after ethanol administration. It therefore seems that the decrease of MEOS in hepatic microsomes caused a smaller increase in the acetaldehyde concentration in the serum of the garlic groups because cytosolic alcohol dehydrogenase showed no significant difference between the control and garlic groups. After ethanol administration, the content of cytochrome P-450 in the hepatic microsomes of the control groups increased, while that of the garlic groups did not change although cytochrome P-450 (CYP) 2E1 and 1A2 in the hepatic microsomes of the garlic groups increased. These results indicate that the induction of isozymes of cytochrome P 450 other than CYP 2E1 and 1A2 was inhibited following garlic administration. Cytosolic high Km and total aldehyde dehydrogenase (AIDH) in the liver of the garlic groups tended to be lower than those activities of the control groups at 1 and 2 h after ethanol administration. It therefore seems that the decreases of AIDH in the hepatic cytosols diminished the increase of acetate in the serum of the garlic groups after ethanol administration. These results suggest that the ethanol metabolism in the mouse liver is controlled by components in fresh garlic juice. PMID- 10524349 TI - Hepatic branched-chain alpha-keto acid dehydrogenase complex in female rats: activation by exercise and starvation. AB - The effects of acute exercise and starvation on hepatic branched-chain alpha-keto acid dehydrogenase (BCKDH) complex activity were examined in female rats fed high (30%)- or low (8%)-protein diets. The total activity of the complex was significantly higher in the high protein-fed rats than in the low protein-fed rats but was not affected by acute exercise and starvation in either diet group. The proportion of the active form of BCKDH complex was less than 10% in both diet groups. Acute exercise and starvation markedly increased the active form of the complex in both diet groups. The activity of BCKDH kinase, which is responsible for inactivation of the BCKDH complex by phosphorylation, tended to be decreased by acute exercise and starvation in both diet groups. These results suggest that the activity of the BCKDH kinase is an important factor determining the proportion of the active form of BCKDH complex in exercise and starvation, and that the female rat is a useful model for studying the regulation of hepatic BCKDH complex activity. PMID- 10524350 TI - Bioavailability of milk micellar calcium phosphate-phosphopeptide complex in rats. AB - We evaluated the bioavailability of two types of calcium from milk in two experiments. One was a micellar calcium phosphate-phosphopeptide (MCP-PP) complex in which the chemical form was similar to the original form of milk, and the other was a commercial whey calcium in which the chemical form was different from that of milk. In experiment 1, the calcium absorption, bone mineral density, and bone strength were examined when growing female rats were fed either MCP-PP complex or whey calcium as the sole source of calcium for 46 d. In experiment 2, the calcium solubility in the small intestine was measured when female rats were meal-fed either MCP-PP complex or whey calcium. The apparent calcium absorption rate in both groups decreased time-dependently during the experimental period, but the time-dependent change in the apparent calcium absorption rate was statistically different. It decreased more slowly in rats fed the MCP-PP diet than in rats fed the whey calcium diet. The bone mineral density of the femur in rats fed the MCP-PP diet was significantly higher than that of the rats fed the whey calcium diet. The bone strength (breaking force and energy) of the femur in rats fed the MCP-PP diet was higher than in the rats fed the whey calcium diet. The amount of soluble calcium in the small intestinal contents in rats at 2.5 h after ingestion of the MCP-PP diet was approximately three times higher than that found in rats fed the whey calcium diet. These results indicate that the calcium bioavailability of MCP-PP complex is higher than that of whey calcium, and this difference is due in part to the solubility in the intestine. PMID- 10524351 TI - Inhibitory effect of Cladosiphon fucoidan on the adhesion of Helicobacter pylori to human gastric cells. AB - We studied the inhibitory effect of Cladosiphon fucoidan on the attachment of Helicobacter pylori (H. pylori), a gastroduodenal pathogen, to human gastric cell lines. The bacterial binding in these cell lines was inhibited more by Cladosiphon fucoidan (IC50 = 16-30 mg/mL), than by the fucoidan from Fucus (IC50 > 30 mg/mL). Dextran sulfate, another sulfated polysaccharide, did not inhibit the binding at all. Pre-incubating the bacterial suspension with fucoidans reinforced the inhibitory ability of these components, and reduced the IC50 value of Cladosiphon fucoidan to approximately 1 mg/mL. However, the binding was not inhibited by pre-treatment of gastric cells with these components. It was also shown that this fucoidan blocks both Leb- and sulfatide-mediated attachment of H. pylori to gastric cells. Furthermore, fucoidan-binding proteins were found on the H. pylori cell surface by Western blot analysis. Thus, the inhibitory effect exerted by Cladosiphon fucoidan on binding between H. pylori and gastric cells might result from the coating with this component of the bacterial surface. PMID- 10524352 TI - Effects of green tea catechin on phospholipase A2 activity and antithrombus in streptozotocin diabetic rats. AB - The purpose of this study was to investigate the effects of dietary green tea catechin on phospholipase A2 (PLA2) activity and the antithrombotic reaction of platelets in streptozotocin (STZ)-diabetic rats. Sprague-Dawley male rats weighing 100 +/- 10 g were randomly divided into one normal and three STZ diabetic groups, which were subdivided into catechin-free group (DM-0C), 0.5% catechin group (DM-0.5C) and 1% catechin group (DM-1C). The activity level of platelet phospholipase A2 was higher in the diabetic groups than in the normal group, while it was lower in DM-0.5C and DM-1C than in DM-0C. The activity of platelet cyclooxygenase in DM-0C was 1.1-fold as high as in the normal group, but was significantly reduced by catechin supplementation. The platelet thromboxane A2 (TXA2) formation became higher in DM-0C as compared to the normal group, but not in DM-0.5C and DM-1C. The synthesis of aortic prostacyclin (PGI2) was lower in DM-0C and DM-0.5C than in the normal group. The PGI2/TXA2 ratio was decreased to 55% in DM-0C, but was restored by catechin supplementation. These results indicate that STZ-diabetic rats are sensitive to platelet aggregation and thrombosis, and that the abnormality can be improved by dietary catechin. PMID- 10524353 TI - Percutaneous absorption of biotin in healthy subjects and in atopic dermatitis patients. AB - The study was designed to test the ability of sequential applications of biotin containing ointment to increase serum biotin levels. Twenty atopic dermatitis patients (mean age, 20.5 yr) and 11 healthy subjects (mean age, 25.5 yr) volunteered to participate in this study. The diagnosis of atopic dermatitis was established dermatologically. Seven grams per day of ointment containing 0.3% biotin and 1-4 g per day of steroid ointment were both applied sequentially. The healthy subjects applied only biotin ointment. The biotin concentration was determined microbiologically. Before biotin treatment, the average serum biotin level was significantly lower in atopic dermatitis patients than in healthy subjects. The percutaneous application of biotin-containing ointment caused a significant increase in the serum biotin concentration in both healthy subjects (from 41.5 +/- 10.0 to 50.2 +/- 9.2 nmol/L) and in atopic dermatitis patients (from 27.9 +/- 17.4 to 50.7 +/- 21.6 nmol/L), especially in patients whose initial level was low, and also could be effective in regulating the atopic allergic response involving eosinophils. In conclusion, biotin appears to be readily absorbed through both normal and dermatitis-affected human skin. PMID- 10524354 TI - Esterification makes retinol more labile to photolysis. AB - Because retinyl palmitate was reported to be more stable to oxidation than retinol, we wondered if retinyl palmitate was also more resistant to photolysis as compared to free alcohol. We investigated the resistance of ethanolic solutions of retinol, retinyl palmitate, or both to air oxidation and (or) photolysis using fluorescent light. The initial concentrations were all-trans retinol, 14 mumol/L, and all-trans-retinyl palmitate, 14 mumol/L. The concentrations of retinol and retinyl palmitate were determined by HPLC and are expressed as a percentage of their original concentrations. After 4 h of exposure to an 18 W fluorescent lamp at 15 cm from the solution, the means (SD) of the surviving analytes were 64% (3%) for retinol and 5% (2%) for retinyl palmitate in a solution containing both retinol and retinyl palmitate. Taking account of the cis isomer arising from retinyl palmitate, 29% (3%) of the retinyl palmitate survived after 4 h of photolysis. Degradation of retinyl palmitate might occur after the conversion of trans isomer to cis isomer during photolysis, however, trans isomer could be degraded with a lesser extent of isomerization. After 4 h of bubbling air through the solution in the dark, 49% (6%) of retinol and 69% (4%) of retinyl palmitate survived. Exposing retinol or retinyl palmitate separately to air oxidation, bubbling air through the solution, or photolysis, exposing them to light, we found that retinyl palmitate could retard the air oxidation of retinol (p < 0.001), but it had no effect on the light-induced degradation of retinol. We also studied the effect of the addition of approximately 1,560 mumol/L alpha-tocopherol, approximately 190 mumol/L beta carotene and approximately 2,000 mumol/L ascorbic acid as antioxidants. In the presence of 156 mumol/L alpha-tocopherol, 87% (1%) of the retinol and 91% (4%) of the retinyl palmitate remained after air oxidation. Although the photolysis of retinol and retinyl palmitate was also inhibited by 190 mumol/L beta-carotene, alpha-tocopherol and ascorbic acid did not exert inhibiting effects. We conclude that retinyl palmitate is physico-chemically more labile to photolysis but is more resistant to air oxidation than retinol. PMID- 10524355 TI - Differences in the effect of iron-deficient diet on tissue weight, hemoglobin concentration and serum triglycerides in Fischer-344, Sprague-Dawley and Wistar rats. AB - This study was designed to examine the differences in the effect of an iron deficient diet on iron metabolism in Fischer-344 (FC), Sprague-Dawley (SD) and Wistar (WT) rats based on hemoglobin (Hb), hematocrit (Hct), serum iron levels, growth rate and organ weight. Hb concentration was higher in FC rats (14 mg/100 mL) on the initial day than in SD (10) and WT (10) rats. Although the Hb level was significantly decreased in FC rats fed an iron-deficient (ID, 8 mg/kg) diet for 33 d compared to the FC rats fed an iron-adequate (IA, 50 mg/kg) diet, the relative concentration of Hb was high in FC rats fed the ID diet as compared to the SD and WT rats fed the same diet. A similar relationship was detected between Hct and serum iron concentrations. Although serum triglycerides (TG) were significantly increased in each rat strain fed the ID diet as compared to the IA diet, the percentage of the value for the IA diet was lowest in FC rats (119%) fed the ID diet as compared to the SD (328) and WT (394) rats fed the same diet. Retroperitoneal fat pad was decreased in FC, SD and WT rats fed the ID diet as compared to the IA diet. SD rats were particularly sensitive to the reduction of retroperitoneal fat pad. The results suggested that rat strains responded differently to dietary iron inadequacy, and that FC rats were less sensitive to an iron-deficient diet as compared to the SD and WT rats. PMID- 10524356 TI - Possible contribution of a decrease in serum albumin concentration to a low level of blood L-tryptophan in nephrotic rats. AB - In order to clarify whether or not a decrease in serum albumin concentration contributes to a low level of blood L-tryptophan (Trp) in nephrosis, blood Trp concentration at 30, 60, 90, and 120 min after oral administration of Trp (100 mumol/kg body weight) in the same rats injected once with puromycin aminonucleoside (PAN) (100 mg/kg body weight, i.p.), an inducer of nephrosis, was examined at different stages of nephrosis. The increase and decrease in blood Trp concentration after Trp administration were similar in the PAN-treated rats without nephrosis, the PAN-treated rats recovered from nephrosis, and untreated control rats. The maximum increase in blood Trp concentration at 30 min after Trp administration was lower in nephrotic rats than in control rats. In all rats treated with and without PAN, increased blood L-tryptophan concentrations at 30 min after L-tryptophan administration were positively correlated well with serum albumin concentrations (r = 0.88, p < 0.001). There was no difference in the intestinal absorption of the same dose of orally administered Trp between nephrotic and control rats. These results suggest that a decrease in serum albumin concentration may contribute to a low level of blood L-tryptophan in nephrosis. PMID- 10524357 TI - Effects of angiotensin I-converting enzyme inhibitor from Ashitaba (Angelica keiskei) on blood pressure of spontaneously hypertensive rats. AB - The inhibitory activity of angiotensin I-converting enzyme (ACE) was extracted with 80% ethanol from the leaves of Ashitaba (Angelica keiskei). The present ACE inhibitor was fractionated and separated with various chromatographies. The antihypertensive effects of the sample (G fraction) from Ashitaba on spontaneously hypertensive rats (SHR) were observed by long-term administration for 10 wk. Another sample (S fraction) from Ashitaba also had antihypertensive effects after a single intravenous administration to SHR. The sample was further purified by using several chromatographies. The ACE inhibitor fraction was characterized as follows: no significant absorbance, a zwitterion, a water soluble substance and a positive ninhydrin reaction. According to a mass spectrum analysis, the molecular weight of the ACE inhibitor was determined to be 303 and Na-salt ions of carboxyl groups were detected. The ACE inhibitor from Ashitaba contained in the anti-hypertensive fraction was speculated to be very similar to authentic nicotianamine based on a comparative study of inhibitory activity, mass spectrum analysis and thin-layer chromatographies. PMID- 10524358 TI - Effect of maitake (Grifola frondosa) water extract on inhibition of adipocyte conversion of C3H10T1/2B2C1 cells. AB - We investigated the effect of maitake (Grifola frondosa) water extract on inhibiting the conversion of C3H10T1/2B2C1 cells into adipocytes. Maitake water extract was fractionated by molecular sieve. Heat-labile compounds strongly inhibiting adipocyte conversion proved to occur in fractions of molecular weight of more than 10,000 on the basis of activity measurement of glycerol-3-phosphate dehydrogenase. PMID- 10524359 TI - [Pharmacological characteristics and clinical application of losartan, an orally active AT1 angiotensin II receptor antagonist]. AB - Losartan is the first orally active angiotensin II receptor type 1 antagonist for a new class of cardiovascular therapeutic agent. Losartan is converted to an active metabolite (E3174) after oral administration in humans and rats. Both losartan and E3174 contribute to the net angiotensin II receptor blockade and produce anti-hypertensive effect. Losartan not only blocks the vasoconstrictive effect of angiotensin II but also inhibits its mitogenic effect; thus losartan is expected to protect against end-organ-damage-related hypertension and chronic heart failure. Unlike angiotensin-coverting-enzyme inhibitor, losartan does not elicit adverse effects of cough and angioneurotic edema by its blockade of angiotensin II receptor. It is also expected to reduce proteinuria in nephropathy. In addition to its blockade of angiotensin II receptor, losartan blocks thromboxane A2 receptor and facilitates excretion of uric acid, although therapeutic importance of these effects are under investigation. In summary, losartan, an angiotensin II type 1 receptor antagonist is a new class of antihypertensive agent and its therapeutic potentials are not merely reduction of blood pressure but total protection from end-organ damage resulting from activation of both the systemic and local renin-angiotensin system. PMID- 10524360 TI - [Gramicidin perforated patch recording technique]. AB - Cl- is one of the major ionic constituents of cells and extracellular spaces. Intracellular Cl- plays an important role in regulating the cell volume and pH, in both salt secretion and reabsorption, in membrane excitability, and G-protein dependent intracellular signal transduction. GABA and glycine are the primary inhibitory neurotransmitters. Such agonist-stimulated responses are affected by the intracellular Cl- concentration ([Cl-]i). However, it was difficult to make an electrical recording of the physiological Cl- response from cells with native intracellular Cl- activity because of the limitations of present recording techniques using conventional glass-microelectrode, whole-cell patch and nystatin perforated patch recording modes. Recently, this difficulty was overcome by developing the gramicidin perforated patch recording mode in our laboratory. Gramicidin is a polypeptide antibiotic that forms pores in the cell membrane as well as nystatin but allows only monovalent cations to permeate the membrane, enabling both [Cl-]i and the second messenger system to remain undisturbed. Here, I would like to primarily focus on the GABA- and glycine-induced Cl- responses to in mammalian CNS neurons maintaining native cellular [Cl-]i under normal and pathological neuronal conditions by use of gramicidin perforated patch recording configuration. Age-related and developmental changes in neuronal [Cl-]i are also described in detail. PMID- 10524361 TI - [Detection of drug-drug interaction by ESR spectroscopy]. AB - Vitamin C (sodium ascorbate), gallic acid and dopamine induced apoptosis (characterized by internucleosomal DNA cleavage, nuclear fragmentation, chromatin condensation near nuclear membrane and loss of cell surface microvilli) in human promyelocytic leukemic HL-60 cells. ESR spectroscopy demonstrated the connection between the radical intensity and apoptosis-inducing activity of derivatives of these compounds. When vitamin C and gallic acid were mixed together, the radical intensity and cytotoxic activity of gallic acid was completely scavenged by one or two orders lower concentrations of vitamin C, suggesting the predominant action of vitamin C over gallic acid. When vitamin C and dopamine were mixed together, their radical intensity and apoptosis-inducing activity counteracted with each other. On the other hand, lignins significantly enhanced both the radical intensity and apoptosis-inducing activity of vitamin C. ESR spectroscopy might be applicable for the detection of drug-drug interaction. PMID- 10524362 TI - [Effects of local anesthetics on rat leukocyte functions]. AB - To determine whether local anesthetics affect functions in macrophages, I examined the effects of 5 local anesthetics, lidocaine HCl, mepivacaine HCl, propitocaine HCl, procaine HCl, and tetracaine HCl, on chemotaxis and production of superoxide anion in rat peripheral macrophages. Rats were intraperitoneally injected with 1% glycogen. Peritoneal exudate cells containing macrophages were obtained from the peritoneal cavity 4 days after the administration. Chemotaxis was evaluated using a 48-well microchemotaxis chamber with a polycarbonate membrane filter. Production of superoxide anion was measured spectrophotometrically by a superoxide dismutase-sensitive reduction of ferricytochrome c. All of the local anesthetics examined at a dose of 1 mg ml inhibited (P < 0.05) chemotaxis and production of superoxide anion in macrophages. Moreover, pretreatment of macrophage suspensions with mepivacaine HCl, propitocaine HCl, procaine HCl, or tetracaine HCl at a dose of 1 mg ml resulted in inhibition of the production of superoxide anion. In contrast, pretreatment with lidocaine HCl at this concentration did not significantly affect the production of superoxide anion. These results suggest that all of the local anesthetics examined at a therapeutic concentration inhibit chemotaxis and production of superoxide anion in rat macrophages. PMID- 10524363 TI - Current awareness in prenatal diagnosis. PMID- 10524364 TI - Managed care and the workers' compensation bargain. PMID- 10524365 TI - Evaluation of the Washington State Workers' Compensation Managed Care Pilot Project I: medical outcomes and patient satisfaction. AB - OBJECTIVES: This study examined the effect of managed care on medical outcomes and patient satisfaction as part of an evaluation of the Washington State Workers' Compensation Managed Care Pilot. METHODS: One hundred twenty firms (7,041 employees) agreed to have their injured workers treated in managed-care plans. Managed care introduced two changes from the fee-for-service (FFS) delivery system currently used by injured workers in Washington State: (1) experience-rated capitation, and (2) a primary occupational-medicine delivery model. The FFS control group included injured workers employed at 392 firms (12,000 employees). A total of 1,313 workers who experienced occupationally related injuries or illnesses between April 1995 and June 1996 were interviewed by telephone at 6 weeks after injury regarding their medical outcomes and satisfaction with care. Workers whose injuries resulted in four or more lost workdays (n = 372) were also interviewed at 6 months after injury on the same topics. The areas surveyed included functional outcomes and satisfaction with care, providers, and access to providers. RESULTS: The measures of functional outcome reflected no consistent differences between the managed care and the FFS conditions. The workers who attended the managed-care system reported lower levels of satisfaction with care, particularly with access to providers. For example, 58% of managed-care patients reported satisfaction with their attending physician as compared with 69% of FFS patients (P<0.01). CONCLUSIONS: Workers treated through managed-care arrangements were less satisfied with their care, but their medical outcomes were similar to those of workers who received traditional FFS care. The current workers' compensation system in Washington State affords injured workers great latitude in choosing providers. If provider choice is substantially restricted by managed care, worker satisfaction is likely to diminish. PMID- 10524366 TI - Evaluation of the Washington State Workers' Compensation Managed Care Pilot Project II: medical and disability costs. AB - OBJECTIVES: This study examined the effect of managed care on medical and disability costs as part of an evaluation of the Washington State Workers' Compensation Managed Care Pilot (MCP). METHODS: One hundred twenty firms (7,041 employees) agreed to have their injured workers treated in managed care plans. Managed care introduced two changes from the fee-for-service (FFS) delivery system currently used by injured workers in Washington State: experience- rated capitation and a primary occupational medicine delivery network. The FFS control group included injured workers employed at 392 firms (12,000 employees). Medical and disability costs were compared for 1,058 injuries in the managed care group and 1,159 injuries in the FFS group occurring between April 1995 and June 1996. Univariate and multivariate statistical methods were used to analyze the effects of managed care on medical and disability costs. RESULTS: The mean unadjusted medical cost per injury ($587) for the managed care group was 21.5% lower (P = 0.06) than for the FFS group ($748). Adjustment for differences in worker and firm-level characteristics through multivariate analysis had little effect on the unadjusted results, except that the difference in costs between managed care and FFS groups became statistically significant (P<0.01). The major cost differences were for outpatient surgery (cost per surgery) and ancillary services (pharmacy, x-ray, physical therapy, and all other costs). In addition, disability costs, particularly percent on time loss and time-loss cost per injury, were significantly lower (P<0.01) in the managed care group. CONCLUSIONS: The results from the MCP suggest that substantial savings in workers' compensation medical and disability costs may be realized using the type of managed care intervention designed for this study. Delivering occupational health services through managed care arrangements whose design is based on an integrated, occupational health centered delivery model may offer a viable approach for improving delivery systems, reducing costs and encouraging greater attention to disability prevention. PMID- 10524367 TI - Medical care expenditures for selected circulatory diseases: opportunities for reducing national health expenditures. AB - OBJECTIVES: Circulatory system diseases are a significant burden in terms of morbidity, mortality, and use of health care services. This article presents total, per capita, and per condition US medical care expenditures in 1995 for circulatory diseases according to sex, age, and type of health service. METHODS: Total personal health care expenditures estimated by the Health Care Financing Administration for each type of health care service are separated into components to estimate patient expenditures by age, sex, primary medical diagnosis, and health care service for all diseases of the circulatory system, heart disease, coronary heart disease, congestive heart failure, hypertensive disease, and cerebrovascular disease. RESULTS: Expenditures for circulatory diseases totaled $127.8 billion in 1995 (17% of all personal health care expenditures), $486 per capita, and $1,636 per condition. Approximately one half of expenditures was for hospital care and 20% was for nursing home care. Heart disease accounted for 60% of circulatory expenditures. Expenditures increased with age and reached 35% of expenditures among persons aged 85 years and older, which was almost $7,000 per capita. These relationships vary somewhat according to the specific circulatory disease, type of health care, and age. CONCLUSIONS: Expenditures increase with age and circulatory diseases can be expected to command an increasing share of national health expenditures as the number and proportion of the population that is elderly grows. The alteration of lifestyles and medical interventions provide many opportunities to prevent circulatory diseases and to reduce national health expenditures. PMID- 10524368 TI - Strategic hospital alliances: do the type and market structure of strategic hospital alliances matter? AB - BACKGROUND: Throughout the 1990s, hospitals formed local alliances to defend against increasingly powerful hospital rivals and to improve their market positions relative to aggressive and consolidating managed-care organizations. An important consequence of hospitals combining or aligning horizontally at the local level is a significant consolidation of hospital markets. OBJECTIVE: The aim of this study was to examine the relationship between the type of the local strategic hospital alliances (SHAs), market, environment, and operational factors with financial performance. METHODS: The study is a cross-sectional analysis of the financial performance across SHAs in all metropolitan statistical areas in 1995. RESULTS: SHAs with dominant or dominant for-profit (FP) hospitals are not more financially successful than other SHAs. SHAs in markets with high health maintenance organization (HMO) or SHA penetration have lower revenues per case mix adjusted discharge. The operational characteristics, proportion of teaching members in the SHA, and SHA bed size, result in higher revenues and expenses, whereas greater SHA technical efficiency results in lower costs. CONCLUSIONS: Health care organizations are centralizing their operations and governance. This study shows that this trend has not added financial value to hospital collectives, at least at this point in their development. PMID- 10524369 TI - Effects of a behavioral health carve-out on inpatient-related quality indicators for major depression treatment. AB - OBJECTIVES: To analyze the effects of the 1993 Massachusetts behavioral health carve-out for state employees on readmissions and follow-up treatment after hospitalization for major depressive disorder (MDD). METHODS: The sample consisted of 218 continuous enrollees in preferred provider organization and/or indemnity plans who had any MDD admissions during fiscal years 1992 to 1995. These users accounted for 310 MDD admissions. Eligibility files and behavioral health claims were used to analyze readmissions and follow-up treatment after discharge. Kaplan-Meier survival functions were obtained for pre/post (pre-carve out vs. post-carve-out) comparisons of the two indicators. Cox regression models were used to estimate carve-out effects on readmission and follow-up treatment while controlling for patient variables. Postdischarge contact categories were also compared. RESULTS: The risk of readmission did not change significantly after the carve-out, in either the Kaplan-Meier or Cox regression analyses. Follow-up treatment was significantly more likely after the carve-out, including in the early postdischarge period. There was a significant decrease in the proportion of discharged patients followed by readmission only, and a significant increase in patients receiving follow-up treatment prior to a readmission. CONCLUSIONS: Under this behavioral health carve-out, follow-up treatment was more likely, and estimated risk of readmission did not change significantly for a seriously ill subgroup of enrollees. This was true even when controlling for patient variables and using data for extended time "at risk" for each indicator. Future research on carveouts should move toward direct clinical quality measurement. PMID- 10524370 TI - Mental health service utilization by African Americans and Whites: the Baltimore Epidemiologic Catchment Area Follow-Up. AB - OBJECTIVE: To compare mental health service utilization and its associated factors between African Americans and whites in the 1980s and 1990s. DESIGN: Household-based longitudinal study with baseline interviews in 1981 and follow-up interviews from 1993 to 1996. SETTING: The Baltimore Epidemiologic Catchment Area (ECA) Follow-Up. SUBJECTS: Subjects included 1,662 adults (590 African Americans and 1,072 whites). MAIN OUTCOME VARIABLE: Use of mental health services, defined as talking to any health professional about emotional or nervous problems or alcohol or drug-related problems within the 6 months preceding each interview. RESULTS: In 1981, crude rates of mental health service use in general medical (GM) settings and specialty mental health settings were similar for African Americans and whites (11.7%). However, after adjustment for predisposing, need, and enabling factors, individuals receiving mental health services were less likely to be African American. Mental health service use increased by 6.5% over follow-up, and African Americans were no longer less likely to report receiving any mental health services in the 1990s. African Americans were more likely than whites to report discussing mental health problems in GM settings without having seen a mental health specialist. They were less likely than whites to report use of specialty mental health services, but this finding was not statistically significant, possibly because of low rates of specialty mental health use by both race groups. Psychiatric distress was the strongest predictor of mental health service use. Attitudes positively associated with use of mental health services were more prevalent among African Americans than whites. CONCLUSIONS: Mental health service use increased in the past decade, with the greatest increase among African Americans in GM settings. Although it is possible that the racial disparity in use of specialty mental health services remains, the GM setting may offer a safety net for some mental health concerns of African Americans. PMID- 10524371 TI - An evaluation of radical prostatectomy at Veterans Affairs Medical Centers: time trends and geographic variation in utilization and outcomes. AB - OBJECTIVE: To examine temporal trends and geographic variation in utilization of radical prostatectomy (RP) as well as 30-day mortality and complication rates. DESIGN: Administrative data-base study of radical prostatectomy (RP) using the Department of Veterans Affairs Patient Treatment File and Outpatient Clinic File between 1986 to 1996. Logistic regression was used to estimate temporal and geographic effects on the use of RP. SETTING: All Departments of Veterans Affairs Medical Centers (VAMC) in the contiguous United States. PATIENTS: Men aged 45 to 84 years who underwent RP at a VAMC (n = 13,398). MAIN OUTCOME MEASURES: Number and utilization of RP, rate of 30-day mortality, major cardiopulmonary or vascular complications, and colorectal injuries requiring surgical repair within 30 days of RP. RESULTS: From 1986 to 1996, the annual number of RP at VAMCs (range, 695-1,545 RP) more than doubled, and the rate of RP at VAMCs per male VA user increased by 40% (range, 48/100,000-66/100,000). After controlling for age and year, the utilization of RP in West North Central, Mountain, West South Central, and Pacific census divisions was 70%, 14%, 10%, and 8% higher, respectively, whereas the utilization of RP in New England, East North Central, and Mid-Atlantic divisions was 38%, 31%, and 25% lower, respectively, than the rest of the nation (P<0.001). Geographic variation in utilization decreased during the period between 1986 and 1996, but a twofold difference in RP utilization in 1996 remained between high- and low-utilization divisions. Major cardiopulmonary complications, vascular complications, and colorectal injuries occurred in 1.7%, 0.2%, and 1.8% of men, respectively. Thirty-day mortality was 0.73%, declined from 1986 to 1996, and was associated with a history of diabetes and congestive heart failure. CONCLUSION: Utilization of RP at VAMCs increased over time and varied across geographic areas. Thirty-day mortality was less than 1% and decreased with time. Differences in utilization may be caused by uncertainty regarding the effectiveness of early detection and treatment of prostate cancer. PMID- 10524372 TI - The care of older women with early-stage breast cancer: what is the role of surgeon gender? AB - BACKGROUND: Over the past decade and a half, a substantial literature has documented age-dependent variations in breast cancer care. Accumulating evidence suggests that these variations impact the health outcomes of older women with breast cancer. Surgeon gender may be an important source of age-dependent variations in care. OBJECTIVE: To examine the relationship between surgeon gender and primary tumor therapy and systemic adjuvant therapy among 303 older women with early-stage breast cancer cared for by 20 surgeons in Boston, Massachusetts. METHODS: The research design was a cross-sectional observational study. The subjects were women at least 55 years of age with newly diagnosed Stage I or II breast cancer. The main outcome measure was definitive primary tumor therapy and systemic adjuvant therapy. RESULTS: After adjustment for patient and tumor characteristics, patients of female surgeons were more likely to receive definitive treatment, with the strongest effect being observed for the receipt of both definitive primary tumor therapy and systemic adjuvant therapy (odds ratio 4.5; 95% confidence interval 2.7, 7.7). CONCLUSIONS: Women with early-stage breast cancer cared for by female surgeons are more likely to receive standard therapies. Surgeons provide the initial care, both diagnostic and therapeutic, for all women with breast cancer. Their role in breast cancer care is pivotal and has a substantial impact on the nature of breast cancer care received. PMID- 10524373 TI - Experience of primary care by racial and ethnic groups in the United States. AB - OBJECTIVES: The purpose of this study was to examine the experience of primary care by racial and ethnic groups and identify aspects of primary care where significant disparities in experience exist across racial and ethnic groups. METHODS: Data for this study came from the Household Component of the 1997-1998 Medical Expenditure Panel Survey (MEPS), a nationally representative survey of the civilian noninstitutionalized population of the United States. Measures were identified within MEPS that denote race, ethnicity, experience of primary care, and socioeconomic covariates associated with access to care. RESULTS: Racial and ethnic minorities experienced worse primary care, particularly in the first contact aspect, than did white Americans. Their usual sources of care were more likely to be hospital settings than private clinics. They faced greater barriers accessing their usual source of care (USC), finding it more difficult to get an appointment and waiting longer during an appointment. Many of the significant differences persist after adjustment for sociodemographic and health-status characteristics. CONCLUSIONS: Racial and ethnic disparity in primary care experience is not simply a reflection of sociodemographic and health-status differences across racial/ethnic groups. Efforts must be made to reduce nonfinancial as well as financial barriers to care and ensure that quality primary care is provided in all settings, public as well as private, and to individuals of all colors. PMID- 10524374 TI - Gender differences in fatigue: biopsychosocial factors relating to fatigue in men and women. AB - BACKGROUND: Fatigue is a common problem, which is found more frequently among women than men. To date, neither the etiology of fatigue nor the factors that explain the gender difference in its incidence are still fully understood. METHODS: In a sample of men (n = 4,681) and women (n = 4,698) (age range, 15-64 years) in the Dutch National Survey of Morbidity and Interventions in General Practice, the gender differences in the underlying biological, psychological, and social factors of fatigue were analyzed. RESULTS: Both general and gender specific factors were recognized. Men and women who experience complaints of fatigue appeared to be younger and more highly educated. They had more acute health complaints and more psychosocial problems and also showed a lower level of perceived health. Among women, only gender-specific biological complaints and psychosocial problems were related to fatigue. In addition, relevant sociodemographic variables included taking care of young children and being employed. Among men, fatigue was particularly related to having handicaps and severe chronic complaints. Taking care of young children did not make a difference in the male sample. CONCLUSIONS: Fatigue can only be adequately understood in a multicausal model with biomedical and psychosocial factors. Complaints of fatigue are too often ignored in general practice. By adopting a patient-centered style of communication, physicians can acquire a more complete picture of the patients' fatigue. PMID- 10524375 TI - The cost of efforts to improve quality. AB - BACKGROUND: Virtually all hospitals in the United States report that they engage in efforts to improve quality, such as continuous quality improvement (CQI). Little is known about the costs of these efforts and whether they are associated with improved outcomes or lower patient-care costs. OBJECTIVES: The principal objective of this study was to provide benchmark data on the costs of efforts to improve quality. The authors also attempted to determine if quality improvement expenditures are correlated with outcomes and/or condition-specific hospital costs. METHODS: Detailed information on the cost of quality improvement was obtained from hospitals participating in a broad study of CQI activities. These data were correlated with patient outcomes and condition-specific costs. The subjects were medium to large hospitals throughout the United States. Senior managers provided budgetary information on direct costs of quality improvement, and details about meetings associated with quality improvement. They also provided summary medical bills for all patients undergoing total hip replacement and coronary artery bypass graft surgery. The billing information was combined with data provided by the Health Care Finance Administration to estimate condition-specific costs. Patients were directly surveyed to obtain information about satisfaction and outcomes. RESULTS: There is a wide range of expenditures on quality improvement activities. Meeting costs are a substantial percentage of total costs. Neither total costs nor meeting costs are correlated with condition specific costs. DISCUSSION: Hospital managers can be expected to insist on evidence that quality improvement expenditures produce tangible benefits. This article provides benchmark estimates of those benefits and a methodology for further research. PMID- 10524376 TI - Creating good news and helping the public--it's in our hands. PMID- 10524377 TI - Meeting the challenges of recruiting and retaining participants in clinical trials. PMID- 10524378 TI - Drug and supplement sales on the Web: novel marketing method or potential time bomb? PMID- 10524379 TI - Do patient education materials for physicians help or hurt dietetics professionals? PMID- 10524380 TI - Do patient education materials for physicians help or hurt dietetics professionals? PMID- 10524381 TI - The Campaign for Coverage of Medical Nutrition Therapy--ADA's public policy activities reach beyond Medicare. PMID- 10524382 TI - Public confusion over food portions and servings. PMID- 10524383 TI - Factors associated with weight gain in women after diagnosis of breast cancer. Women's Healthy Eating and Living Study Group. AB - OBJECTIVE: To identify the factors associated with weight gain after diagnosis of breast cancer in a heterogeneous population of women. DESIGN: Descriptive cross sectional study. SUBJECTS: 1,116 patients who had been diagnosed with stage I, stage II, or stage IIIA primary, operable breast cancer within the previous 4 years. Patients were recruited during enrollment into a diet intervention trial to reduce risk for breast cancer recurrence. Analysis Demographic data, weight history, and physical activity information obtained by questionnaire and medical information obtained by chart review; dietary assessment based on four 24-hour dietary recalls collected by telephone. Associations between weight change after the diagnosis of breast cancer and prediction variables were examined using univariate and multiple linear regression analyses. RESULTS: Overall, 60% of the subjects reported weight gain, 26% reported weight loss, and 14% reported no change in weight after the diagnosis of breast cancer. The overall mean weight change was a gain of 2.7 kg (6 lb). Factors positively and independently associated with weight gain were time since diagnosis of breast cancer, adjuvant chemotherapy, African-American ethnicity, current energy intake, and postmenopausal status at time of study entry. Factors inversely and independently associated with weight gain were prediagnosis body mass index, age at diagnosis, education level, and exercise index score. APPLICATIONS: Higher energy intake and lower level of physical activity are independently associated with increased risk for weight gain after the diagnosis of breast cancer. Strategies to modify these behaviors are likely to influence the long-term pattern of weight change. PMID- 10524384 TI - Individualizing nutrition counseling for patients with cancer. PMID- 10524385 TI - Energy balance in women with breast cancer during adjuvant treatment. AB - OBJECTIVE: To compare weight, body composition, and major determinants of energy balance of women treated with adjuvant chemotherapy (n = 8) using Adriamycin and cyclophosphamide (AC), or radiation therapy (n = 10). DESIGN: The study used a nonrandomized prospective design. Pretreatment and posttreatment measurements, obtained at baseline and 12 weeks, respectively, included weight, body composition (determined using dual-energy x-ray absorptiometry), energy intake (determined using 3-day food records), resting energy expenditure (determined in indirect calorimetry), and physical activity (determined using 3-day physical activity records). Poststudy follow-up weights were obtained for 13 women. SUBJECTS/SETTING: Eighteen premenopausal women with breast cancer in the early stage, recruited from outpatient clinics, participated in and completed the study. STATISTICAL ANALYSES PERFORMED: Unpaired Student t tests or X2 tests were used to test for differences in baseline subject characteristics, and repeated measures analysis of variance was used to compare groups before and after treatment. RESULTS: Body weight was unchanged in both treatment groups during the study, although poststudy follow-up weights (n = 13) suggested a tendency for weight gain in both groups. Significant changes in body composition for both groups included a mean loss of 0.8 kg total lean body mass (LBM), a mean loss of 0.4 kg LBM in the leg region, and a mean 1.3% increase in percent body fat, from 40.0% to 41.3%. Overall, no between-group differences were observed in any factors associated with energy balance. APPLICATIONS: In this short-term study, AC chemotherapy using fewer antineoplastic agents and number of treatments than most chemotherapy protocols for breast cancer, did not result in weight gain during treatment. Regardless of weight gain, changes in body composition may occur in women with breast cancer during or after treatment. These potential changes have important implications for preventive nutrition counseling. PMID- 10524386 TI - Dietary changes favorably affect bone remodeling in older adults. AB - OBJECTIVE: To determine whether dietary counseling to increase milk intake could produce useful changes in the calcium economy and what, if any, other nutrition related changes might be produced. DESIGN: Randomized, open trial. SUBJECTS/SETTING: Two hundred four healthy men and women, aged 55 to 85 years, who habitually consumed fewer than 1.5 servings of dairy foods per day. Six academic health centers in the United States. INTERVENTION: Subjects were instructed to consume 3 servings per day of nonfat milk or 1% milk as a part of their daily diets, or to maintain their usual diets, for a 12-week intervention period, which followed 4 weeks of baseline observations. MAIN OUTCOME MEASURES: Energy and nutrient intake assessed from milk intake logs and 3-day food records; serum calciotrophic hormone levels at baseline and at 8 and 12 weeks; urinary excretion of calcium and N-telopeptide at 12 weeks. STATISTICAL ANALYSES: Repeated-measures analysis of variance. RESULTS: In the milk-supplemented group, calcium intake increased by 729 +/- 45 mg/day (mean +/- standard error), serum parathyroid hormone level decreased by approximately 9%, and urinary excretion of N-telopeptide, a bone resorption marker, decreased by 13%. Urine calcium excretion increased in milk-supplemented subjects by 21 +/- 7.6 mg/day (mean +/- standard error), less than half the amount predicted to be absorbed from the increment in calcium intake. All of these changes were significantly different from baseline values in the milk group and from the corresponding changes in the control group. Bone-specific alkaline phosphatase level (a bone formation marker) fell by approximately 9% in both groups. Serum level of insulin-like growth factor-1 (IGF-1) rose by 10% in the milk group (P < .001), and the level of insulin-like growth factor binding protein-4 (IGFBP-4) fell slightly (1.9%) in the milk group and rose significantly (7.9%) in the control group (P < .05). APPLICATIONS/CONCLUSIONS: The changes observed in the calcium economy through consumption of food sources of calcium are similar in kind and extent to those reported previously for calcium supplement tablets. The increase in IGF-1 level and the decrease in IBFBP-4 level are new observations that are beneficial for bone health. Important improvements in skeletal metabolism can feasibly occur in older adults by consumption of food sources of calcium. Dietitians can be confident that food works, and that desired calcium intakes can be achieved using food sources. PMID- 10524387 TI - Counseling older adults about calcium and bone health. PMID- 10524388 TI - Infant formula preparation, handling, and related practices in the United States. AB - OBJECTIVE: To describe practices related to infant formula feeding: diluting and concentrating formula, mixing formula with warm tap water, sterilizing, storing prepared formula, heating in a microwave oven, putting the baby to bed with a bottle, and adding cereal and sweeteners to formula; to analyze characteristics related to compliance with recommended practices; and to examine the relation between formula handling and infant diarrhea. SUBJECTS/DESIGN: Subjects were mothers who fed their infants formula (more than 1,000 subjects at each infant age). Data are from the US Food and Drug Administration's Infant Feeding Practices Study (IFPS), a national longitudinal survey with a nonprobability sample. Data were collected by mail, and formula practices were included at infant ages 2, 5, and 7 months. STATISTICAL ANALYSES PERFORMED: Logistic regression was conducted and percentages and odds ratios were calculated, adjusting for instruction in preparing formula from a health care professional, education, income, age, parity, work status, and breast-feeding practices. RESULTS: Failure to comply with recommendations was high for several practices with clear health implications; 33% of mothers mixed formula with warm tap water and up to 48% heated bottles in a microwave oven. Mothers of 2-month-old infants who received instruction from a health care professional and who breast-fed showed increased compliance, but few demographic characteristics, such as education, were related. Diarrhea increased with ambient holding of formula for older infants. APPLICATION: Advice from a health care professional can improve formula-handling behaviors. Dietitians and other health care professionals should provide information on proper preparation and handling of infant formula to all infant caregivers. PMID- 10524389 TI - Baseline fruit and vegetable intake among adults in seven 5 a day study centers located in diverse geographic areas. AB - OBJECTIVE: To examine baseline rates of fruit and vegetable consumption among adults in the 5 A Day research trials in order to identify any regional and sociodemographic differences associated with daily servings. DESIGN: The main outcome measure was the frequency of fruits and vegetables consumed within 1 month of the baseline survey as assessed by a 7-item food frequency questionnaire (FFQ). SUBJECTS/SETTING: Participants (N = 15,060) were from 7 study centers. Study centers included schools (N = 48), worksites (N = 60), churches (N = 50), or the Special Supplemental Nutrition Program for Women, Infants, and Children (WIC) clinics (N = 15) in interventions to increase fruit and vegetable consumption. STATISTICAL ANALYSES: Means and standard errors, adjusting for clusters, were calculated. A mixed linear model analyzed relationships between fruit and vegetable consumption and regional center, gender, age, race, education, income, marital status, food-shopping responsibility, and whether one lives with children. RESULTS: Results indicate an overall mean intake of 3.6 daily servings of fruits and vegetables. Significant differences in mean daily servings were found among the regional study centers (low of 3.0 to high of 4.1). There were significant differences in mean daily consumption by age (< 30 years = 3.7 servings per day; 30 to 49 years = 3.4; > or = 50 years = 3.7), education (> high school = 3.4 servings per day; high school graduate = 3.4; some college = 3.5; college graduate = 3.9), race (black = 3.7 servings per day; Hispanic = 3.0; white = 3.6; other = 3.7), marital status (married = 3.6 servings per day; single = 3.5), and food-shopping responsibilities (little = 3.2 servings per day; about half = 3.6; most = 3.8). Only 17% of respondents ate 5 or more servings of fruits and vegetables per day. CONCLUSIONS: The 7 regions showed significant variability in daily fruit and vegetable consumption, suggesting that a single national message to increase fruit and vegetable consumption may not reach the population segments most in need of changing. It is advisable to spend more time understanding the food consumption habits of the population under investigation to develop messages to foster behavior change. PMID- 10524390 TI - Chocolate: food or drug? AB - Although addictive behavior is generally associated with drug and alcohol abuse or compulsive sexual activity, chocolate may evoke similar psychopharmacologic and behavioral reactions in susceptible persons. A review of the literature on chocolate cravings indicates that the hedonic appeal of chocolate (fat, sugar, texture, and aroma) is likely to be a predominant factor in such cravings. Other characteristics of chocolate, however, may be equally as important contributors to the phenomena of chocolate cravings. Chocolate may be used by some as a form of self-medication for dietary deficiencies (eg, magnesium) or to balance low levels of neurotransmitters involved in the regulation of mood, food intake, and compulsive behaviors (eg, serotonin and dopamine). Chocolate cravings are often episodic and fluctuate with hormonal changes just before and during the menses, which suggests a hormonal link and confirms the assumed gender-specific nature of chocolate cravings. Chocolate contains several biologically active constituents (methylxanthines, biogenic amines, and cannabinoid-like fatty acids), all of which potentially cause abnormal behaviors and psychological sensations that parallel those of other addictive substances. Most likely, a combination of chocolate's sensory characteristics, nutrient composition, and psychoactive ingredients, compounded with monthly hormonal fluctuations and mood swings among women, will ultimately form the model of chocolate cravings. Dietetics professionals must be aware that chocolate cravings are real. The psychopharmacologic and chemosensory effects of chocolate must be considered when formulating recommendations for overall healthful eating and for treatment of nutritionally related health issues. PMID- 10524391 TI - A multidimensional weight-management program for women. AB - Real and imagined overweight and obesity, and resulting weight-reduction efforts, are associated with the development of a variety of health problems and eating disorders. For many years, research and practice in the field of weight management have been based largely on a unidimensional, simplistic, weight-loss paradigm. The long-term success rate for persons using this paradigm has been low. This article presents a multidimensional paradigm that focuses on all aspects of the prevention, treatment, and management of weight-related problems. The goal is to stimulate a shift from the unidimensional to a more multidimensional approach in weight-management interventions. The paradigm presents weight management as a continuum on which 5 prominent points are identified: formulation of reasonable weight goals, prevention of unnecessary weight; gain or loss, weight loss when necessary, prevention of relapse, and acceptance of an overweight/obese physique when necessary. The intrapersonal characteristics and skills associated with this continuum, namely, self-esteem, body image, self-efficacy, locus of control, motivation, stress management, problem solving and decision making, and assertiveness, as well as the role of stage of change and environmental influences on weight management, are examined. Issues concerning the different dimensions of the paradigm are discussed as are challenges and applications for researchers and practitioners in the field of weight management. PMID- 10524392 TI - Marketing can change consumers' perceptions of healthfulness of items served in a worksite cafeteria. PMID- 10524393 TI - Vitamin and mineral diet adequacy and supplement use by full-time employed women with preschool children. PMID- 10524394 TI - Breast-feeding practices of WIC participants from the former USSR. PMID- 10524395 TI - Impact of combined weight-control and smoking-cessation interventions on body weight: review of the literature. PMID- 10524396 TI - A comparison of breast and colon cancer incidence rates among native Asian Indians, US immigrant Asian Indians, and whites. PMID- 10524397 TI - Position of the American Dietetic Association: functional foods. AB - It is the position of The American Dietetic Association that functional foods, including whole foods and fortified, enriched, or enhanced foods, have a potentially beneficial effect on health when consumed as part of a varied diet on a regular basis, at effective levels. The Association supports research to further define the health benefits and risks of individual functional foods and their physiologically active components. Dietetics professionals will continue to work with the food industry, government, the scientific community, and the media to ensure that the public has accurate information regarding this emerging area of food and nutrition science. Knowledge of the role of physiologically active food components, both from phytochemicals and zoochemicals, has changed the role of diet in health. Functional foods have evolved as food and nutrition science has advanced beyond the treatment of deficiency syndromes to reduction of disease risk. This position reviews the definition of functional foods, their regulation, and the scientific evidence supporting this emerging area of food and nutrition. Foods can no longer be evaluated only in terms of macronurtrient and micronutrient intake. Analyzing the content, of other physiologically active components will be necessary. The availability of health-promoting functional foods in the US diet has the potential to help ensure a healthier population. However, each functional food should be evaluated on the basis of scientific evidence to ensure appropriate integration into a varied diet. PMID- 10524398 TI - Kaye Stanek wins Huddelson Award. PMID- 10524399 TI - Case problem: balancing nutrition advice with dental care in patients with anorexia and bulimia. PMID- 10524400 TI - Professional identity and the soul of radiation oncology. PMID- 10524401 TI - Physicians and non-physician practitioners: working together for improved patient care. PMID- 10524402 TI - Finding our sensitive patients. PMID- 10524403 TI - Esophageal brachytherapy: a phantom menace? PMID- 10524404 TI - Editorial comment on "Long-term adverse effects of radiation inhibition of restenosis". PMID- 10524405 TI - The importance of adequate follow-up in defining treatment success after external beam irradiation for prostate cancer. AB - PURPOSE: We reviewed our institution's experience treating patients with localized prostate cancer with external beam radiation therapy (RT) to determine how differences in the length of follow-up affect the determination of treatment outcome using the American Society for Therapeutic Radiology and Oncology (ASTRO) Consensus Panel Definition of biochemical failure (BF). METHODS AND MATERIALS: From January 1987 through December 1997, 1109 patients with localized prostate cancer were treated with definitive external beam RT at William Beaumont Hospital, Royal Oak, Michigan. All patients received external beam RT to a median total prostate dose of 66.6 Gy (range: 59.4-70.4 Gy). A total of 1096 patients (99%) had sufficient prostate-specific antigen (PSA) follow-up to determine their biochemical status. To test the impact of differences in follow-up on the calculation of BF, 389 patients with at least 5 years of PSA follow-up were selected as the reference group for the initial analysis. BF was then retrospectively determined using the Consensus Panel definition at yearly intervals, ignoring the remainder of each patient's follow-up. The median follow up for this group of patients was 6.6 years (range: 5.0-11.6 years). In a second analysis, patient cohorts were randomly selected with varying median PSA follow up intervals in order to more accurately represent a population whose follow-up is distributed continuously over a defined range. Seven cohorts were randomly selected with 200 patients in each cohort. Cohorts were individually identified such that half of the patients (100) had 2 years or less follow-up than the stated time point for analysis and half (100) had up to 2 years more follow-up than the time point chosen for analysis. For example, in the cohort with a median follow-up of 3 years, 100 patients with a PSA follow-up from 1 to 3 years were randomly selected, and 100 patients with a follow-up from 3 to 5 years were randomly selected, thus generating a median follow-up of 3 years for this cohort (range: 1 to 5 years). This process was repeated five times for five random samples of seven cohorts each. Biochemical failure was calculated according to the Consensus Panel definition. RESULTS: In the first analysis, significantly different rates of biochemical control (varying by 6-21%) were calculated for the same actuarial year chosen for analysis depending only upon the length of follow up used. For example, the 3-year actuarial rate of biochemical control (BC) varied from 71% when calculated with 3 years of follow-up versus 50.4% with 7 years (p < 0.01). These differences in actuarial rates of BC were observed in all subsets of patients analyzed (e.g., PSA < 10, Gleason < or = 6, n = 132,p < 0.001; PSA < 10, Gleason > or = 7, n = 33, p = 0.03; PSA > or = 10, Gleason < or = 6, n = 109, p < 0.001; and PSA > or = 10, Gleason > or = 7, n = 72, p = 0.002). The absolute magnitude of the difference in actuarial rates of BC was greatest during years 2 (range 18-30%), 3 (range 16-25%), and 4 (range 15-24%) after treatment. In the second analysis using median PSA follow-ups (as defined above), statistically significant differences in actuarial rates of BC were again observed. For example, the 3-year actuarial rate of BC varied from 74.8% with a median follow-up of 2 years versus 49.2% with a median follow-up of 6 years. These dramatic differences in BC were still observed beyond 5 years. CONCLUSION: When the ASTRO Consensus Panel definition of BF is used to calculate treatment success with external beam RT for prostate cancer, adequate follow-up is critical. Depending upon the length of time after treatment, significantly different rates of BC (varying by 15% to 30%) can be calculated for the same time interval chosen for analysis. These results suggest that data should only be reported if the length of follow-up extends at least beyond the time point at which actuarial results are examined for the majority of patients. PMID- 10524406 TI - Tamsulosin palliates radiation-induced urethritis in patients with prostate cancer: results of a pilot study. AB - PURPOSE: A pilot study was performed to determine the effectiveness of Flomax (tamsulosin HCl) in the management of acute radiation urethritis in prostate cancer patients undergoing conformal external beam radiation therapy (RT). Potential predictors of response to Flomax were evaluated. METHODS AND MATERIALS: From January 1998 to April 1998, 26 consecutive patients who developed symptoms of radiation urethritis while undergoing RT for prostate cancer were treated with Flomax, a superselective alpha1A-adrenergic antagonist. A genitourinary review of systems served as the instrument used to assess baseline urinary function and treatment response. RESULTS: The initial response rate to Flomax was 62% (16/26) at the 0.4 mg level and 60% (6/10) at the 0.8 mg level. Half of the 16 patients who initially responded to 0.4 mg subsequently progressed. Three-fourths of those patients who progressed, however, achieved a durable response with the 0.8 mg dose. Therefore urinary symptoms were ultimately controlled in 77% (20/26) of the patients. After correcting for the testing of multiple hypotheses (n = 5), the presence of benign prostatic hyperplasia (BPH) approached statistical significance for predicting the initial response to the 0.4 mg dose of Flomax (78% vs. 25%, p = 0.03). CONCLUSION: Flomax appears to be effective in relieving the symptoms of radiation urethritis. A Phase II trial is justified and in progress. PMID- 10524407 TI - Efficacy of selective alpha-1 blocker therapy in the treatment of acute urinary symptoms during radiotherapy for localized prostate cancer. AB - PURPOSE: To determine the efficacy of an alpha-1 adrenoreceptor blocking agent for acute urinary symptoms in patients treated with radiotherapy for localized prostate cancer. METHODS AND MATERIALS: Between 1987 and 1995, 743 patients with clinically localized prostate cancer were treated with 3D-CRT. A total of 275 (37%) patients developed Grade 2 acute urinary symptoms as defined by the RTOG morbidity scoring system. Terazosin hydrochloride (THC), a selective alpha-1 adrenoceptor blocking agent, was given to 119 (43%) patients for treatment of their urinary symptoms, whereas nonsteroidal anti-inflammatory medications (NSAID) were administered to 71 patients (26%). Thirty-one patients (11%) were treated with other medications, and 54 (20%) did not seek pharmacologic intervention for their urinary symptoms. Patients were monitored weekly to assess changes in urinary urgency, frequency, and nocturia. RESULTS: Treatment with THC resulted in a significant resolution of urinary symptoms in 79 of 119 patients (66%), while 26 (22%) had moderate improvement, and 14 (12%) had minimal to no response to this drug. In contrast, only 11 of 71 (16%) of the patients treated with NSAIDs experienced significant symptom relief, 20 (28%) had moderate improvement, and 40 (56%) had minimal to no response. The difference in the significant symptomatic improvement between THC and NSAID therapy (66% vs. 16%) was highly significant (p < 0.001). For patients treated with THC, a higher likelihood of significant symptom relief was observed in patients who did not receive neoadjuvant androgen ablation (p = 0.04) and in those who were younger than 65 years of age (p = 0.02). CONCLUSION: Alpha-1 selective adrenoceptor blocking agents are effective in ameliorating the acute urinary symptoms in patients receiving radiotherapy for localized prostate cancer. Although this was not a randomized prospective study, the data suggest that NSAIDs were less effective in relieving radiation-induced urinary symptoms. PMID- 10524408 TI - Effect of post-implant edema on the rectal dose in prostate brachytherapy. AB - PURPOSE: To characterize the effect of prostate edema on the determination of the dose delivered to the rectum following the implantation of 125I or 103Pd seeds into the prostate. METHODS AND MATERIALS: From 3 to 5 post-implant computed tomography (CT) scans were obtained on 9 patients who received either 125I or 103Pd seed implants. None of the patients received hormone therapy. The outer surface of the rectum was outlined on each axial CT image from the base to the apex of the prostate. The D10 rectal surface dose, defined as the dose which encompasses only 10% of the surface area of the rectum, was determined from each CT scan by compiling a dose-surface histogram (DSH) of the rectal surface. The magnitude and half-life of the post-implant edema in each of these implants is known from the results of a previously published study based on the analysis of the serial CT scans. RESULTS: As the prostate edema resolved, the distance between the most posterior implanted seeds and the anterior surface of the rectum decreased. As a result, the D10 rectal surface dose increased with each successive post-implant CT scan until the edema resolved. The dose increased exponentially at approximately the same rate the prostate volume decreased. The D10 rectal surface dose at 30 days post-implant ranged from 16% to 190% (mean 68 +/- 50%) greater than on day 0. The dose on day 30 was at least 50% greater in 6 of 9 cases. CONCLUSION: The rectal surface dose determined by analysis of a post implant CT scan of an 125I or 103Pd prostate seed implant depends upon the timing of the CT scan. The dose indicated by the CT scan on day 30 is typically at least 50% greater than that indicated by the CT scan on day 0. Because of this difference, it is important to keep the timing of the post-implant CT in mind when specifying dose thresholds for rectal morbidity. PMID- 10524410 TI - Accelerated radiotherapy with delayed concomitant boost in locally advanced squamous cell carcinoma of the head and neck. AB - PURPOSE: To determine the toxicity, maximum tolerated dose (MTD), and clinical effectiveness of a 5-week course of accelerated radiotherapy with delayed concomitant boost in locally advanced squamous cell carcinoma of the head and neck (SCCHN). METHODS AND MATERIALS: Thirty-five patients with untreated T3T4NM0 or TN2 (> 3 cm) N3M0 SCC of the oral cavity, oropharynx, hypopharynx, or larynx were entered in the study between January 1994 and October 1997. The initial target volume was treated with conventional daily fractions. A small field boost covering gross disease was added as a second daily fraction during the last 2 weeks of the 5-week schedule, using a minimum interfraction interval of 6 h. The study was initiated using 180-cGy fractions to deliver a total dose of 63 Gy over 33-35 days. A classical dose escalation strategy was planned to increase the delivered dose in steps using minimum cohorts of three patients, up to a maximum of 70 Gy in 200-cGy fractions. RESULTS: In the dose escalation study, 4 patients were entered at level 1 (63 Gy), 9 at level 2 (65 Gy), and 8 at level 3 (67 Gy). One patient was withdrawn at level 2 because of unstable angina, and 1 at level 3 because of uncontrolled diabetes. One patient at level 3 failed to complete treatment because of radiation toxicity. RTOG Grade 3 mucositis, dermatitis, or pharyngitis was documented in 1 (25%), 5 (63%), and 7 (100%) evaluable patients at levels 1, 2, and 3, respectively. Grade 4 reactions were documented in 1 patient at each level. One patient at level 3 died 5 weeks post-treatment of unknown causes. Two additional patients at level 3 died of progressive disease and RT toxicity. Sixty-five Gy (level 2) was chosen as the MTD. In the MTD study, 14 additional patients were entered at level 2, providing a total of 22 evaluable patients with a median follow-up of 21 months (range 12-41 months). Grade 3 mucositis, dermatitis, or pharyngitis were documented in 11 (50%), 8 (36%), and 6 (27%) patients, respectively. One patient developed Grade 4 mucositis. A complete response was recorded in 16 (77%). Three of 5 patients with uncontrolled disease and 3 of 3 patients with recurrent disease underwent salvage surgery with no postoperative complications. Radiotherapy controlled disease above the clavicles in 14 (68%). Ultimate locoregional control was achieved in 17 (77%). The disease free, overall, and cause-specific survival of all patients entered at level 2 was 56%, 76%, and 80%, respectively, at 2 years. Late complications have been limited to 3 patients (trismus, chronic mucosal ulcer, and soft tissue necrosis). CONCLUSION: A 5-week course of accelerated radiotherapy with delayed concomitant boost can deliver 65 Gy with acceptable toxicity, encouraging rates of complete response, and locoregional control, and no compromise of salvage surgery in patients with locally advanced SCCHN. The regimen is worthy of further study in a Phase III trial. PMID- 10524409 TI - Dose, volume, and function relationships in parotid salivary glands following conformal and intensity-modulated irradiation of head and neck cancer. AB - PURPOSE: To determine the relationships between the three-dimensional dose distributions in parotid glands and their saliva production, and to find the doses and irradiated volumes that permit preservation of the salivary flow following irradiation (RT). METHODS AND MATERIALS: Eighty-eight patients with head and neck cancer irradiated with parotid-sparing conformal and multisegmental intensity modulation techniques between March 1994 and August 1997 participated in the study. The mean dose and the partial volumes receiving specified doses were determined for each gland from dose-volume histograms (DVHs). Nonstimulated and stimulated saliva flow rates were selectively measured from each parotid gland before RT and at 1, 3, 6, and 12 months after the completion of RT. The data were fit using a generalized linear model and the normal tissue complication probability (NTCP) model of Lyman-Kutcher. In the latter model, a "severe complication" was defined as salivary flow rate reduced to < or =25% pre-RT flow at 12 months. RESULTS: Saliva flow rates data were available for 152 parotid glands. Glands receiving a mean dose below or equal to a threshold (24 Gy for the unstimulated and 26 Gy for the stimulated saliva) showed substantial preservation of the flow rates following RT and continued to improve over time (to median 76% and 114% of pre-RT for the unstimulated and stimulated flow rates, respectively, at 12 months). In contrast, most glands receiving a mean dose higher than the threshold produced little saliva with no recovery over time. The output was not found to decrease as mean dose increased, as long as the threshold dose was not reached. Similarly, partial volume thresholds were found: 67%, 45%, and 24% gland volumes receiving more than 15 Gy, 30 Gy, and 45 Gy, respectively. The partial volume thresholds correlated highly with the mean dose and did not add significantly to a model predicting the saliva flow rate from the mean dose and the time since RT. The NTCP model parameters were found to be TD50 (the tolerance dose for 50% complications rate for whole organ irradiated uniformly) = 28.4 Gy, n (volume dependence parameter) = 1, and m (the slope of the dose/response relationship) = 0.18. Clinical factors including age, gender, pre-RT surgery, chemotherapy, and certain medical conditions were not found to be significantly associated with the salivary flow rates. Medications (diuretics, antidepressants, and narcotics) were found to adversely affect the unstimulated but not the stimulated flow rates. CONCLUSIONS: Dose/volume/function relationships in the parotid glands are characterized by dose and volume thresholds, steep dose/response relationships when the thresholds are reached, and a maximal volume dependence parameter in the NTCP model. A parotid gland mean dose of < or =26 Gy should be a planning goal if substantial sparing of the gland function is desired. PMID- 10524411 TI - Concurrent chemoradiotherapy followed by adjuvant chemotherapy in Asian patients with nasopharyngeal carcinoma: toxicities and preliminary results. AB - PURPOSE: Nasopharyngeal carcinoma (NPC) is endemic in Singapore. Nearly 60% of the patients diagnosed with NPC will present with locally advanced disease. The North American Intergroup study 0099 reported improved survival outcome in patients with locally advanced NPC who received combined chemoradiotherapy when compared to radiotherapy alone. Hence we explored the feasibility and efficacy of a similar protocol in our patients. METHODS AND MATERIALS: Between June 1996 and December 1997, 57 patients were treated with the following schedule as described. Radical radiotherapy (RT) of 66-70 Gy to the primary and neck with cisplatin (CDDP) 25 mg/m2 on days 1-4 given by infusion over 6-8 hours daily on weeks 1, 4, and 7 of the RT. This is followed by a further 3 cycles of adjuvant chemotherapy starting from week 11 from the first dose of radiation (CDDP 20 mg/m2/d and 5 fluorouracil [5-FU] 1 gm/m2/d on days 1-4 every 28 days). RESULTS: The majority of patients (68%) had Stage IV disease. About 54% of patients received all the intended treatment; 75% received all 3 cycles of CDDP during the RT phase and 63% received all three cycles of adjuvant chemotherapy. The received dose intensity of CDDP and 5-FU of greater than 0.8 was achieved in 58% and 60% of the patients respectively. Two treatment-related deaths due to reactivation of hepatitis B and neutropenic sepsis respectively, were encountered. At median follow-up of 16 months, 14 patients had relapsed, 12 systemically and 2 loco-regionally. CONCLUSION: Due to the acceptable tolerability of such a protocol in our cohort of patients, we have embarked on a Phase III study to confirm the results of the 0099 Intergroup study in the Asian context. PMID- 10524412 TI - Deep inspiration breath-hold technique for lung tumors: the potential value of target immobilization and reduced lung density in dose escalation. AB - PURPOSE/OBJECTIVE: This study evaluates the dosimetric benefits and feasibility of a deep inspiration breath-hold (DIBH) technique in the treatment of lung tumors. The technique has two distinct features--deep inspiration, which reduces lung density, and breath-hold, which immobilizes lung tumors, thereby allowing for reduced margins. Both of these properties can potentially reduce the amount of normal lung tissue in the high-dose region, thus reducing morbidity and improving the possibility of dose escalation. METHODS AND MATERIALS: Five patients treated for non-small cell lung carcinoma (Stage IIA-IIIB) received computed tomography (CT) scans under 4 respiration conditions: free-breathing, DIBH, shallow inspiration breath-hold, and shallow expiration breath-hold. The free-breathing and DIBH scans were used to generate 3-dimensional conformal treatment plans for comparison, while the shallow inspiration and expiration scans determined the extent of tumor motion under free-breathing conditions. To acquire the breath-hold scans, the patients are brought to reproducible respiration levels using spirometry, and for DIBH, modified slow vital capacity maneuvers. Planning target volumes (PTVs) for free-breathing plans included a margin for setup error (0.75 cm) plus a margin equal to the extent of tumor motion due to respiration (1-2 cm). Planning target volumes for DIBH plans included the same margin for setup error, with a reduced margin for residual uncertainty in tumor position (0.2-0.5 cm) as determined from repeat fluoroscopic movies. To simulate the effects of respiration-gated treatments and estimate the role of target immobilization alone (i.e., without the benefit of reduced lung density), a third plan is generated from the free-breathing scan using a PTV with the same margins as for DIBH plans. RESULTS: The treatment plan comparison suggests that, on average, the DIBH technique can reduce the volume of lung receiving more than 25 Gy by 30% compared to free-breathing plans, while respiration gating can reduce the volume by 18%. The DIBH maneuver was found to be highly reproducible, with intra breath-hold reproducibility of 1.0 (+/- 0.9) mm and inter breath-hold reproducibility of 2.5 (+/- 1.6) mm, as determined from diaphragm position. Patients were able to perform 10-13 breath-holds in one session, with a comfortable breath-hold duration of 12-16 s. CONCLUSION: Patients tolerate DIBH maneuvers well and can perform them in a highly reproducible fashion. Compared to conventional free-breathing treatment, the DIBH technique benefits from reduced margins, as a result of the suppressed target motion, as well as a decreased lung density; both contribute to moving normal lung tissue out of the high-dose region. Because less normal lung tissue is irradiated to high dose, the possibility for dose escalation is significantly improved. PMID- 10524413 TI - Radiotherapy and concurrent continuous infusion of cisplatin with adjuvant surgery in nonresectable Stage III lung carcinoma: short- and long-term results of a Phase II study. AB - PURPOSE: Cisplatin-enhanced radiotherapy plus adjuvant surgery was evaluated in nonresectable non-small cell lung carcinoma (NSCLC). METHODS AND MATERIALS: Doses of 50 Gy (administered in standard fractionation in 5 weeks) were delivered with concurrent cisplatin in continuous infusion (daily dose: 4 mg/m2), to 32 Stage IIIa and 45 Stage IIIb patients enrolled in a Phase II study. Patients without progression underwent surgery. RESULTS: Esophagitis (64%), nausea/vomiting (34%), and pulmonary toxicity (14%) were the main side effects. Grade 3 toxicity occurred in 4 instances. A clinical locoregional major response was achieved by 55 patients (there were 10 complete responses). Forty patients underwent surgery, 7 with a nonradical procedure. Seven patients died due to surgery-related complications, which were significantly impacted by right pneumonectomy (71% vs. 6% of the other procedures, p < 0.0001). Eighteen of the 40 surgical patients were assessed to be without viable tumor and 11 with microresidual carcinoma. There were 13 disease-free, 5-year survivors. CONCLUSIONS: Toxicity was low but activity high with the chemoradiotherapy. Adjuvant surgery increased the rate of complete responses, but right pneumonectomy had an unacceptable mortality. The role of surgery needs further refinement. Integration of the chemoradiotherapy schedule with cisplatin-based induction chemotherapy is advisable. PMID- 10524414 TI - Multi-institutional randomized trial of external radiotherapy with and without intraluminal brachytherapy for esophageal cancer in Japan. Japanese Society of Therapeutic Radiology and Oncology (JASTRO) Study Group. AB - PURPOSE: With the aim of improving the results of treatment of esophageal cancer, we designed this multi-institutional, randomized trial to establish the optimal irradiation method in radical radiation therapy for esophageal cancer by clinically evaluating external irradiation alone and in combination with intraluminal brachytherapy. METHODS AND MATERIALS: The study population consisted of patients with squamous cell carcinoma who were expected to be successfully treated with radical radiation therapy. The patients who could be given intraluminal brachytherapy at the end of external irradiation of 60 Gy were stratified into 2 groups. Patients assigned to receive external irradiation alone received boost irradiation of 10 Gy/week on a schedule similar to the previous one, and with the same or smaller irradiation field. Intraluminal brachytherapy was performed, as a rule, with the reference dose point set at a depth of 5 mm of the esophageal submucosa, and a total of 10 Gy was irradiated at a daily dose of 5 Gy, on a once-weekly schedule with low-dose-rate or high-dose-rate brachytherapy equipment. RESULTS: A total of 103 patients were registered, 94 of whom were analyzable, with 8 ineligible, and 1 for whom complete information was unavailable. The overall cumulative survival rate was 20.3% at 5 years. The cause specific survival rate was 31.8% at 5 years. The cause-specific survival rate at 5 years was 27% in the external irradiation alone group and 38% in intraluminal brachytherapy combined group. There was no significant difference between the 2 groups (p = 0.385). However, in the patients with 5 cm or less tumor length, the cause-specific survival rate was 64% at 5 years in the intraluminal brachytherapy combined group, which showed a significant improvement over 31.5% in the external irradiation alone group (p = 0.025). In the patients with Stage T1 and T2 disease, cause-specific survival rates tended to be better in the intraluminal brachytherapy combined group than in the external irradiation alone group (p = 0.088). In the patients with more than 5 cm tumor length or Stage T3-4 disease, there were no significant differences between the two groups by treatment methods (p = 0.290). The incidence of early and late complications did not differ according to whether intraluminal brachytherapy was used. CONCLUSION: For the purpose of establishing the usefulness of intraluminal brachytherapy, further prospective randomized studies are necessary to evaluate the efficacy in tumors with short length and those with shallow invasion, or to assess the usefulness of intraluminal brachytherapy, as additional irradiation in large advanced tumors have been shown to have disappeared by diagnostic imaging after chemoradiotherapy with 60 Gy/6w external irradiation. PMID- 10524415 TI - Phase II study of radiochemotherapy with UFT and low-dose oral leucovorin in patients with unresectable rectal cancer. AB - PURPOSE: To determine the activity and evaluate the toxicity of uracil and tegafur in a 4:1 molar concentration (UFT) plus low-dose leucovorin administered concomitantly with pelvic irradiation in patients with unresectable or recurrent rectal cancer. METHODS AND MATERIALS: Thirty-five patients (22 with primary unresectable tumors and 13 with locally recurrent tumors) were enrolled in the trial. Thirty-five patients were evaluable for toxicity and 32 of these were evaluable for clinical response. Patients received 300 mg/m2/day UFT and 30 mg/day leucovorin on days 8-35 concomitantly with pelvic radiotherapy, to a total dose of 45 Gy. RESULTS: Eight of the 35 (23%) patients developed Grade 3 diarrhea and were treated with radiotherapy alone after this event. Of the 22 patients with unresectable primary tumors, 17 underwent surgery, and resection was feasible in 15 cases (88%). Of the 32 patients evaluable for clinical response, 4 (13%) had a complete clinical response (CR) and 22 (69%) a partial response (PR). A complete pathologic response was observed in 3 cases (18%) and, a PR in 11 cases (65%). CONCLUSION: The response rates achieved with this schedule seem comparable to those obtained with 5-FU and radiotherapy. These results warrant further evaluation of this combination in patients with unresectable or locally advanced tumors. PMID- 10524416 TI - Intraoperative radiotherapy for resectable extrahepatic bile duct cancer. AB - PURPOSE: Through a retrospective study of intraoperative radiation therapy (IORT) in bile duct cancer, we hope to help clarify its clinical usefulness. METHODS AND MATERIALS: Between 1976 and 1996, IORT was carried out in 35 patients with bile duct cancer at the Tokyo Metropolitan Komagome Hospital. Of the 35 patients, resection proved to be curative in 15. Intraoperative irradiation of 15-30 Gy (average 20.1 Gy) was delivered by electron beam in the 5- to 19-MeV energy ranges. Postoperative external-beam radiation therapy (EBRT) was also delivered in 16 patients. The EBRT was fractionated to 2 Gy/day, in principle, and was delivered at 8.8-54 Gy (average 40.4 Gy) by 10-MV X-rays. RESULTS: The median survival in our patients was 19 months. The 1-year, 2-year, and 5-year survival rates were 57%, 43%, and 19%, respectively. Statistical analysis identified the following prognostic factors: performance status, curative surgical resection, lymph node metastasis, IORT dosage, and treatment period. Only 1 patient (3%) died within 30 days after surgery, and the incidence of late-onset complications was 21%. CONCLUSION: The combination of IORT and EBRT is useful for patients with bile duct cancer who undergo noncurative resection or who have lymph node metastasis. PMID- 10524417 TI - Evaluation of the therapeutic effect of radiotherapy on cervical cancer using magnetic resonance imaging. AB - PURPOSE: This study was performed to evaluate magnetic resonance imaging (MRI) in determining the therapeutic effect of radiotherapy (RT) on cervical cancer. METHODS AND MATERIALS: Serial MRI studies were performed in 42 patients with predominantly advanced cervical cancer before, during, and after radiotherapy. Patients underwent external irradiation combined with high-dose-rate intracavitary (HDR) brachytherapy. T-2 weighted spin-echo pulse sequences with long repetition and echo times were used at a field strength of 1.5 T. Multiple punch biopsies of the cervix were obtained from the high-signal intensity area in all patients at the same time as the MRI. RESULT: In biopsies performed immediately after RT, no residual tumors were found in 36 patients (86%); in 6 patients, residual tumors were observed. The simultaneous MRI study demonstrated no high-signal intensity on T2-weighted images in 28 patients. A high-signal area was observed in 14 patients, and this disappeared 3 months after RT in 8 patients with a negative histological study. The sensitivity, specificity, and accuracy of MRI studies at 3 months after RT were 100%. When the relationship between reduction of tumor volume at 30 Gy and local tumor control was analyzed, every patient with a reduction under 30% gained local control. Also, patients with no residual tumors 3 months after RT gained local control. CONCLUSION: MRI studies performed at 30 Gy of external irradiation and 3 months after RT were predictive factors of local control. PMID- 10524418 TI - Lack of effect of tumor size on the prognosis of carcinoma of the uterine cervix Stage IB and IIA treated with preoperative irradiation and surgery. AB - PURPOSE: The purpose of this analysis was to evaluate the prognostic significance of cervical tumor size in patients with Stages Ib and IIa carcinoma of the cervix treated with preoperative irradiation and radical or conservative hysterectomy. METHODS AND MATERIALS: This study is a retrospective analysis of 177 patients. One hundred forty-one patients had Stage Ib and 36 patients had Stage IIa carcinoma of the cervix. All patients were treated with preoperative irradiation and surgery. Radiation therapy consisted of external pelvic irradiation and intracavitary brachytherapy; total doses ranged from 30 to 60 Gy to the pelvic sidewall and 60 to 70 Gy to point A. Surgery consisting of radical hysterectomy and lymph node dissection or a conservative hysterectomy and lymph node dissection was performed 4 to 6 weeks after completion of irradiation. RESULTS: The 5-year progression-free survivals were 80% for Stage Ib and 63% for Stage IIa (p = 0.03). The 5-year cumulative pelvic failure rates for Stage Ib were 16% for tumors <3 cm and 9% for tumors >3 cm (p = 0.90). The 5-year cumulative pelvic failure rates for Stage IIa were 22% for tumors <3 cm and 22% for tumors >3 cm (p = 0.75). The corresponding cumulative distant metastasis failure rates at 5 years for Stage Ib were 21% for tumors <3 cm and 21% for tumors >3 cm (p = 0.60). For patients with Stage IIa disease, the 5-year cumulative distant metastasis rates were 33% for tumors <3 cm and 36% for tumors >3 cm (p = 0.70). A multivariate analysis was performed to evaluate prognostic factors for the endpoint of progression-free survival. The variables that were analyzed were patient age, tumor histology, tumor size, clinical stage, point A and pelvic lymph node irradiation dose, and cervical tumor status and pelvic lymph node status at the time of hysterectomy. The variables that were found to be of independent significance for progression-free survival by multivariate analysis were pelvic lymph node irradiation dose (p <0.001), pelvic lymph node status at the time of hysterectomy (p = 0.01), and clinical stage (p = 0.02). Cervical tumor size at the time of diagnosis and the presence of tumor cells in the cervix in the hysterectomy specimen was not an independent prognostic factor by multivariate analysis. The overall severe complication rate was 11% for all patients. CONCLUSIONS: For this population of patients treated with preoperative irradiation and surgery, pelvic lymph node status at the time of hysterectomy and the preoperative irradiation dose to the pelvic lymph nodes are independent predictors of progression-free survival and the development of distant metastasis. The pretreatment cervical tumor size is of less importance for predicting progression-free survival and the development of distant metastasis but clinical stage is an important prognostic variable. These results are in contrast with those of surgery or irradiation alone, in which primary tumor size is a critical prognostic factor for all outcome parameters. PMID- 10524419 TI - Hyperfractionation in carcinoma of the cervix: tumor control and late bowel complications. AB - PURPOSE: Hyperfractionation has been advocated to improve local tumor control by increasing radiation dose without increasing late normal tissue complications. The aim of this study was to determine if hyperfractionation decreased late bowel complications. METHODS AND MATERIALS: Thirty patients with Stage II and III cervical cancer were randomized to receive either hyperfractionation or conventional fractionation. Patients were followed for 5 years and monitored for tumor control, recurrence, and bowel complications. The relative risks of tumor control and bowel complications were computed at 1 year and 5 years of follow-up. Kaplan-Meier survival curves were plotted to determine probabilities of being tumor-free and bowel complication-free. RESULTS: There were 15 patients in each group. At 1 year of follow-up, 2 patients in the hyperfractionation group (13%) and 7 patients in the conventional treatment group (45%) had tumor (relative risk [RR] 0.3; 95% confidence interval [CI] 0.1, 1.1; p = 0.054). Delayed bowel complications were seen in 8 patients in the hyperfractionation group and 1 patient in the conventional treatment group (RR 7.5; 95% CI 1.1, 52; p = 0.014). At 5 years, 2 patients in the hyperfractionation group and 8 patients in the conventional treatment group had tumor (RR 0.3; 95% CI 0.1, 1.1; p = 0.04). Delayed bowel complications (Grades 2 and 3) occurred in 9 women in the hyperfractionation group and 2 patients in the conventional group (RR 5.4; 95% CI 1.5, 19.5; p = 0.0006). Kaplan-Meier analysis showed that the hyperfractionation group had significantly more bowel complications over the 5 years of follow-up (p = 0.024). CONCLUSION: Hyperfractionation may result in better tumor control both at 1 year and at 5 years following treatment of cervical cancer. However, hyperfractionation could lead to increased late bowel complications and must be used judiciously in the treatment of cervical cancer. PMID- 10524420 TI - Treatment of early epithelial ovarian cancer with chemotherapy and abdominopelvic radiotherapy: results of a prospective treatment protocol. AB - PURPOSE: To test the hypothesis that the combination of adjuvant chemotherapy and abdominopelvic radiation (APRT) improves the outcome of patients with early ovarian cancer compared to treatment with APRT alone. METHODS AND MATERIALS: Between 1991 and 1994, 93 patients with Stage I to III, optimally cytoreduced, invasive, epithelial ovarian cancer were treated with sequential chemotherapy and APRT. Treatment was assigned using a prognostic classification that was derived from previous cohorts of patients. Low-risk patients (n = 9) received APRT alone, intermediate-risk patients (n = 66) received two courses of cisplatin followed by APRT, and high-risk patients (n = 18) received 6 courses of cisplatin and cyclophosphamide followed by APRT. RESULTS: Disease recurred in 22 patients, and was confined to the pelvis or abdomen in 15. Nine patients died and the remainder were alive with disease after receiving salvage chemotherapy. The 3-year disease free and overall survivals were 78% and 91%, respectively. The prognostic classification used to assign treatment was the only factor that predicted disease-free survival (83% and 59% at 3 years for low/intermediate- and high-risk patients, respectively; p = 0.03). There was no detectable difference in outcome between the present series and an historical control group treated with APRT alone. Treatment was well tolerated and only 2 patients (2.5%) developed serious complications. CONCLUSION: APRT is an effective adjuvant treatment for carefully selected patients with early ovarian cancer. The addition of chemotherapy as used in this study to APRT does not significantly improve outcome compared to APRT alone. PMID- 10524421 TI - Validation of the methods of cosmetic assessment after breast-conserving therapy in the EORTC "boost versus no boost" trial. EORTC Radiotherapy and Breast Cancer Cooperative Groups. European Organization for Research and Treatment of Cancer. AB - PURPOSE: To evaluate both qualitative and quantitative scoring methods for the cosmetic result after breast-conserving therapy (BCT), and to compare the usefulness and reliability of these methods. METHODS AND MATERIALS: In EORTC trial 22881/10882, stage I and II breast cancer patients were treated with tumorectomy and axillary dissection. A total of 5318 patients were randomized between no boost and a boost of 16 Gy following whole-breast irradiation of 50 Gy. The cosmetic result was assessed for 731 patients in two ways. A panel scored the qualitative appearance of the breast using photographs taken after surgery and 3 years later. Digitizer measurements of the displacement of the nipple were also made using these photographs in order to calculate the breast retraction assessment (BRA). The cosmetic results after 3-year follow-up were used to analyze the correlation between the panel evaluation and digitizer measurements. RESULTS: For the panel evaluation the intraobserver agreement for the global cosmetic score as measured by the simple Kappa statistic was 0.42, considered moderate agreement. The multiple Kappa statistic for interobserver agreement for the global cosmetic score was 0.28, considered fair agreement. The specific cosmetic items scored by the panel were all significantly related to the global cosmetic score; breast size and shape influenced the global score most. For the digitizer measurements, the standard deviation from the average value of 30.0 mm was 2.3 mm (7.7%) for the intraobserver variability and 2.6 mm (8.7%) for the interobserver variability. The two methods were significantly, though moderately, correlated; some items scored by the panel were only correlated to the digitizer measurements if the tumor was not located in the inferior quadrant of the breast. CONCLUSIONS: The intra- and interobserver variability of the digitizer evaluation of cosmesis was smaller than that of the panel evaluation. However, there are some treatment sequelae, such as disturbing scars and skin changes, that can not be evaluated by BRA measurements. Therefore, the methods of cosmetic evaluation used in a study must be chosen in a way that balances reliability and comprehensiveness. PMID- 10524422 TI - The influence of the boost in breast-conserving therapy on cosmetic outcome in the EORTC "boost versus no boost" trial. EORTC Radiotherapy and Breast Cancer Cooperative Groups. European Organization for Research and Treatment of Cancer. AB - PURPOSE: To evaluate the influence of a radiotherapy boost on the cosmetic outcome after 3 years of follow-up in patients treated with breast-conserving therapy (BCT). METHODS AND MATERIALS: In EORTC trial 22881/10882, 5569 Stage I and II breast cancer patients were treated with tumorectomy and axillary dissection, followed by tangential irradiation of the breast to a dose of 50 Gy in 5 weeks, at 2 Gy per fraction. Patients having a microscopically complete tumor excision were randomized between no boost and a boost of 16 Gy. The cosmetic outcome was evaluated by a panel, scoring photographs of 731 patients taken soon after surgery and 3 years later, and by digitizer measurements, measuring the displacement of the nipple of 3000 patients postoperatively and of 1141 patients 3 years later. RESULTS: There was no difference in the cosmetic outcome between the two treatment arms after surgery, before the start of radiotherapy. At 3-year follow-up, both the panel evaluation and the digitizer measurements showed that the boost had a significant adverse effect on the cosmetic result. The panel evaluation at 3 years showed that 86% of patients in the no-boost group had an excellent or good global result, compared to 71% of patients in the boost group (p = 0.0001). The digitizer measurements at 3 years showed a relative breast retraction assessment (pBRA) of 7.6 pBRA in the no-boost group, compared to 8.3 pBRA in the boost group, indicating a worse cosmetic result in the boost group at follow-up (p = 0.04). CONCLUSIONS: These results showed that a boost dose of 16 Gy had a negative, but limited, impact on the cosmetic outcome after 3 years. PMID- 10524423 TI - Use of the RTOG recursive partitioning analysis to validate the benefit of iodine 125 implants in the primary treatment of malignant gliomas. AB - PURPOSE: To date, numerous retrospective studies have suggested that the addition of brachytherapy to the conventional treatment of malignant gliomas (MG) (surgical resection followed by radiotherapy +/- chemotherapy) leads to improvements in survival. Two randomized trials have suggested either a positive or no survival benefit with implants. Critics of retrospective reports have suggested that the improvement in patient survival is due to selection bias. A recursive analysis by the RTOG of MG trials has stratified MG patients into 6 prognostically significant classes. We used the RTOG criteria to analyze the implant data at Wayne State University to determine the impact of selection bias. METHODS AND MATERIALS: Between July 1991 and January 1998, 75 patients were treated with a combination of surgery, radiotherapy, and stereotactic I-125 implant as primary MG management. Forty-one (54.7%) were male; 34 (45.3%) female. Median age was 52 years (range 4-79). Twenty-two (29.3%) had anaplastic astrocytoma (AA); 53 (70.7%), glioblastoma multiforme (GBM). Seventy-two patients had data making them eligible for stratification into the 6 RTOG prognostic classes (I-VI). Median Karnofsky performance status (KPS) was 90 (range 50-100). There were 14, 0, 14, 31, 12, and 1 patients in Classes I to VI, respectively. Median follow-up time for AA, GBM, and any surviving patient was 29, 12.5, and 35 months, respectively. RESULTS: At analysis, 29 (40.3%) patients were alive; 43 (59.7%), dead. For AA and GBM patients, 2-year and median survivals were: 58% and 40%; 38 and 17 months, respectively. For analysis purposes, Classes I and II, V and VI were merged. By class, the 2-year survival for implanted patients compared to the RTOG data base was: III--68% vs. I--76%; III--74% vs. 35%; IV--34% vs. 15%; V/VI--29% vs. V--6%. For implant patients, median survival by class was (in months): I/II--37; III--31; IV--16; V/VI--11. CONCLUSION: When applied to MG patients receiving permanent I-125 implant, the criteria of the RTOG recursive partitioning analysis are a valid tool to define prognostically distinct survival groups. As reflected in the RTOG study, a downward survival trend for the implant patients is seen from "best to worse" class patients. Compared to the RTOG database, median survival achieved by the addition of implant is improved most demonstrably for the poorer prognostic classes. This would suggest that selection bias alone does not account for the survival benefit seen with I-125 implant and would contradict the notion that the patients most eligible for implant are those gaining the most benefit from the treatment. In light of the contradictory results from two randomized studies and given the present results, further randomized studies with effective stratification are required since the evidence for a survival benefit with brachytherapy (as seen in retrospective studies) is substantial. PMID- 10524424 TI - The incidence of cerebrovascular accidents in patients with pituitary adenoma. AB - BACKGROUND AND PURPOSE: Patients with pituitary adenomas are effectively treated with a combination of surgery, radiotherapy, and medical therapy. Nevertheless, long-term studies suggest increased mortality that is independent of tumor control, with cerebrovascular accidents (CVA) as the major contributing cause. The purpose of this study was to define the frequency of CVAs in a cohort of patients with pituitary adenoma and identify potential predisposing factors. PATIENTS AND METHODS: A cohort of 331 United Kingdom (UK) residents with pituitary adenoma treated at the Royal Marsden Hospital (RMH) between 1962 and 1986 was studied. The frequency of CVA was assessed from RMH and referring hospital records and clinicians, by postal questionnaire of referring hospitals and general practitioners, and by examination of all death certificates. The data were analyzed by actuarial methods, and risk factors were assessed by multivariate analysis. The data were compared to the incidence of CVA in the general population using a published UK population cohort. RESULTS: Sixty-four of 331 patients developed CVA after primary treatment of pituitary adenoma. The actuarial incidence of CVA was 4% (95% CI: 2-7%) at 5 years, 11% (95% CI: 8-14%) at 10 years, and 21% (95% CI: 16-28%) at 20 years measured from the date of radiotherapy. The relative risk of CVA compared to the general population in the UK was 4.1. Age was an independent predictive factor for CVA. However, the relative risk in comparison to the general population was independent of age. The relative risk of developing CVA was higher in women compared to men, in patients undergoing debulking surgery compared to less radical procedures, and in patients diagnosed and treated in the 1980s compared to previous decades. The dose of radiotherapy was an additional independent prognostic factor on multivariate analysis. CONCLUSION: Patients with pituitary adenoma treated with surgery and radiotherapy have a significantly increased risk of CVA compared to the general population. The factors which may contribute to the increased risk include the presence of pituitary adenoma and consequent endocrine disturbances and the treatment, particularly the extent of surgery and the dose of radiotherapy. When assessing the value of treatment strategies, it is therefore important to include not only intermediate endpoints of tumor and hormonal control, but also late toxicity, including the incidence of CVA and overall survival as the primary endpoint. The potential predisposing factors for CVA need further elucidation to develop treatment strategies with lower risk and consequently, reduced mortality. PMID- 10524425 TI - Temporal lobe necrosis following radiation therapy for nasopharyngeal carcinoma: 1H MR spectroscopic findings. AB - PURPOSE: To observe the patterns of radiation-induced temporal lobe necrosis (TLN) following radiation therapy for nasopharyngeal carcinoma (NPC). METHODS AND MATERIALS: Twenty-five proton magnetic resonance spectroscopic (1H MRS) examinations were acquired from 13 healthy adult volunteers for comparison with data from the patient population. There were 18 patients (28 spectra) with radiologic evidence of TLN and all patients were confirmed cases of NPC treated with radiation therapy. Six patients (33%) had a single treatment while 12 (67%) patients had two treatments. All 1H MRS examinations were performed on a 2-T whole body system (Bruker) using the point-resolved spectroscopy (PRESS) method with TE = 135 ms, TR = 3000 ms, and data processed automatically using the LCModel software package for metabolite quantification. RESULTS: The N-acetyl aspartate (NAA) levels were reduced in all except one spectrum (96%). Choline (Cho) was increased in 3 (11%), normal in 4 (14%), and reduced in 21 (75%) spectra. The creatine (Cr) level was normal in 8 (29%) spectra and reduced in 20 (71%) spectra. In four patients with normal imaging findings 1H MRS was abnormal. CONCLUSION: 1H MRS can characterize radiation-induced TLN. Spectra with increased Cho can be mistaken for neoplasm. Spectroscopy can also identify metabolic derangement before imaging. PMID- 10524426 TI - Altered apoptotic profiles in irradiated patients with increased toxicity. AB - PURPOSE: A retrospective study of radiation-induced apoptosis in CD4 and CD8 T lymphocytes, from 12 cancer patients who displayed enhanced toxicity to radiation therapy and 9 ataxia telangiectasia patients, was performed to test for altered response compared to healthy blood-donors and normal cancer patients. METHODS AND MATERIALS: Three milliliters of heparinized blood from each donor was sent via express post to the Paul Scherrer Institute (PSI) for subsequent examination. The blood was diluted 1:10 in RPMI medium, irradiated with 0-, 2-, or 9-Gy X-rays, and incubated for 48 h. CD4 and CD8 T-lymphocytes were then labeled using FITC conjugated antibodies, erythrocytes were lysed, and the DNA stained with propidium iodide. Subsequently, cells were analyzed using a Becton Dickinson FACScan flow cytometer. Radiation-induced apoptosis was recognized in leukocytes as reduced DNA content attributed to apoptosis-associated changes in chromatin structure. Apoptosis was confirmed by light microscopy, electron microscopy, and by the use of commercially available apoptosis detection kits (in situ nick translation and Annexin V). Data from hypersensitive individuals were compared to a standard database of 105 healthy blood-donors, and a database of 48 cancer patient blood donors who displayed normal toxicity to radiation therapy. To integrate radiosensitivity results from CD4 and CD8 T-lymphocytes after 2 and 9 Gy, z-score analyses were performed. RESULTS: A cohort of 12 hypersensitive patients was evaluated; 8 showed enhanced early toxicity, 3 showed enhanced late toxicity, and 1 showed both. The cohort displayed less radiation-induced apoptosis (-1.8 sigma) than average age-matched donors. A cohort of 9 ataxia telangiectasia homozygotes displayed even less apoptosis (-3.6 sigma). CONCLUSION: The leukocyte apoptosis assay appears to be a useful predictor of individuals likely to display increased toxicity to radiation therapy; however, validation of this requires a prospective study. PMID- 10524427 TI - External beam radiotherapy for painful osseous metastases: pooled data dose response analysis. AB - PURPOSE: Although the effectiveness of external beam irradiation in palliation of pain from osseous metastases is well established, the optimal fractionation schedule has not been determined. Clinical studies to date have failed to demonstrate an advantage for higher doses. To further address this issue, we conducted a pooled dose response analysis using data from published Phase III clinical trials. METHODS AND MATERIALS: Complete response (CR) was used as an endpoint because it was felt to be least susceptible to inconsistencies in assessment.The biological effective dose (BED) was calculated for each schedule using the linear-quadratic model and an alpha/beta of 10. Using SAS version 6.12, the data were fitted using a weighted linear regression, a logistic model, and the spline technique. Finally, BED was categorized, and odds ratios for each level were calculated. RESULTS: CR was assessed early and late in 383 and 1,007 patients, respectively. Linear regression on the early-response data yielded a poor fit and a nonsignificant dose coefficient. With the late-response data, there was an excellent fit (R-square = 0.842) and a highly significant dose coefficient (p = 0.0002). Fitting early CR to a logistic model, we could not establish a significant dose response relationship. However, with the late response data there was an excellent fit and the dose coefficient was significantly different from zero (0.017 +/- 0.00524; p = 0.0012). Application of the spline technique or removal of an outlier resulted in an improved fit (p = 0.048 and p = 0.0001, respectively). Using BED of < 14.4 Gy as a reference level, the odds ratios for late CR were 2.29-3.32 (BED of 19.5-51.4 Gy, respectively). CONCLUSION: Our results demonstrate a clear dose-response for pain relief. Further testing of high intensity regiments is warranted. PMID- 10524428 TI - High-dose spatially-fractionated radiation (GRID): a new paradigm in the management of advanced cancers. AB - PURPOSE: With the advent of megavoltage radiation, the concept of spatially fractionated (SFR) radiation has been abandoned for the last several decades; yet, historically, it has been proven to be safe and effective in delivering large cumulative doses (> 100 Gy) of radiation in the treatment of cancer. SFR radiation has been adapted to megavoltage beams using a specially constructed grid. This study evaluates the toxicity and effectiveness of this approach in treatment of advanced and bulky cancers. METHODS AND MATERIALS: From January 1995 through March 1998, 71 patients with advanced bulky tumors (tumor sizes > 8 cm) were treated with SFR high-dose external beam megavoltage radiation using a GRID technique. Sixteen patients received GRID treatments to multiple sites and a total of 87 sites were irradiated. A 50:50 GRID (open to closed area) was utilized, and a single dose of 1,000-2,000 cGy (median 1,500 cGy) to Dmax was delivered utilizing 6 MV photons. Sixty-three patients received high-dose GRID therapy for palliation (pain, mass, bleeding, or dyspnea). In 8 patients, GRID therapy was given as part of a definitive treatment combined with conventionally fractionated external beam irradiation (dose range 5,000-7,000 cGy) followed by subsequent surgery. Forty-seven patients were treated with GRID radiation followed by additional fractionated external beam irradiation, and 14 patients were treated with GRID alone. Thirty-one treatments were delivered to the abdomen and pelvis, 30 to the head and neck region, 15 to the thorax, and 11 to the extremities. RESULTS: For palliative treatments, a 78% response rate was observed for pain, including a complete response (CR) of 19.5%, and a partial response (PR) of 58.5% in these large bulky tumors. A 72.5% response rate was observed for mass effect (CR 14.6%, PR 52.9%). The response rate observed for bleeding was 100% (50% CR, 50% PR) and for dyspnea, a 60% PR rate only. A relatively higher response rate (CR 23.3%, PR 60%) was observed in patients who received GRID treatment in the head and neck area. No grade 3 late skin, subcutaneous, mucosal, GI, or CNS complications were observed in any patient in spite of these high doses. In the 8 patients who received GRID treatment for definitive treatment, a clinical CR was observed in 5 patients (62.5%) and a pathological complete response was confirmed in the operative specimen in 4 patients (50%). CONCLUSION: The efficacy and safety of using a large fraction of SFR radiation was confirmed by this study and substantiates our earlier results. In selected patients with bulky tumors (> 8 cm), SFR radiation can be combined with fractionated external beam irradiation to yield improved local control of disease, both for palliation and selective definitive treatment, especially where conventional treatment alone has a limited chance of success. PMID- 10524429 TI - Ambulatory patient classifications and the regressive nature of Medicare reform: is the reduction in outpatient health care reimbursement worth the price? AB - PURPOSE: To evaluate the impact of the proposed Ambulatory Patient Classification (APC) system on reimbursement for hospital outpatient Medicare procedures at the Massachusetts General Hospital (MGH) Department of Radiation Oncology. METHODS AND MATERIALS: Treatment and cost data for the MGH Department of Radiation Oncology for the fiscal year 1997 were analyzed. This represented 66,981 technical procedures and 41 CPT-4 codes. The cost of each procedure was calculated by allocating departmental costs to the relative value units (RVUs) for each procedure according to accepted accounting principles. Net reimbursement for each CPT-4 procedure was then calculated by subtracting its cost from the allowed 1998 Boston area Medicare reimbursement or from the proposed Boston area APC reimbursement. The impact of the proposed APC reimbursement system on changes in reimbursement per procedure and on volume-adjusted changes in overall net reimbursements per procedure was determined. RESULTS: Although the overall effect of APCs on volume-adjusted net reimbursements for Medicare patients was projected to be budget-neutral, treatment planning revenues would have decreased by 514% and treatment delivery revenues would have increased by 151%. Net reimbursements for less complicated courses of treatment would have increased while those for treatment courses requiring more complicated or more frequent treatment planning would have decreased. Net reimbursements for a typical prostate interstitial implant and a three-treatment high-dose-rate intracavitary application would have decreased by 481% and 632%, respectively. CONCLUSION: The financial incentives designed into the proposed APC reimbursement structure could lead to compromises in currently accepted standards of care, and may make it increasingly difficult for academic institutions to continue to fulfill their missions of research and service to their communities. The ability of many smaller, low patient volume, high Medicare mix hospital-based radiation oncology departments to continue to deliver their current level of care could be compromised. APC reform may carry monetary and opportunity costs which far outweigh its apparent savings. As payment systems continue to place pressure on operating margins, it becomes even more critical that both academic and community radiation oncology practices know the cost of providing services. PMID- 10524430 TI - Ionizing radiation enhances immunogenicity of cells expressing a tumor-specific T cell epitope. AB - BACKGROUND: p53 point mutations represent potential tumor-specific cytolytic T lymphocyte (CTL) epitopes. Whether ionizing radiation (IR) alters the immunological properties of cells expressing mutant p53 in respect of the CTL epitope generated by a defined point mutation has not been evaluated. METHODS: Mutant p53-expressing syngeneic, nontumor forming BALB/c 3T3 fibroblasts, tumor forming ras-transfected BALB/c 3T3 sarcomas, and DBA/2-derived P815 mastocytoma cells, which differ at the level of minor histocompatibility antigens, were used as cellular vaccines. Cells were either injected with or without prior IR into naive BALB/c mice. Cellular cytotoxicity was assessed after secondary restimulation of effector spleen cells in vitro. RESULTS: Injection of P815 mastocytoma cells expressing the mutant p53 induced mutation-specific CTL in BALB/c mice irrespective of prior irradiation. However, syngeneic fibroblasts or fibrosarcomas endogenously expressing mutant p53 were able to induce significant mutation-specific CTL only when irradiated prior to injection into BALB/c mice. IR of fibroblasts did not detectably alter the expression of cell surface molecules involved in immune response induction, nor did it alter the short-term in vitro viability of the fibroblasts. Interestingly, radioactively-labeled fibroblasts injected into mice after irradiation showed altered organ distribution, suggesting that the in vivo fate of these cells may play a crucial role in their immunogenicity. CONCLUSIONS: These findings indicate that IR can alter the immunogenicity of syngeneic normal as well as tumor forming fibroblasts in vivo, and support the view that ionizing radiation enhances immunogenicity of cellular tumor vaccines. PMID- 10524431 TI - Repair of DNA double-strand breaks and radiosensitivity to killing in an isogenic group of p53 mutant cell lines. AB - PURPOSE: Accumulation of the p53 protein can result in G1 arrest that may facilitate DNA repair, or alternatively, it may lead to apoptosis. Mutations that alter p53's ability to mediate these responses are expected to alter cell radiosensitivity to killing. However, the relationship between p53 status and cell radiosensitivity has proven to be complex. Several studies have suggested that p53 mutations are associated with increased radioresistance to killing, while others have shown no such correlation. These differences may be derived from the fact that different mutations of p53 exert different effects on cell radiosensitivity. METHODS AND MATERIALS: To address this question, we examined a group of isogenic cell lines that express different "hot spot" mutant forms of p53. These cells were generated from human osteosarcoma (SAOS) cells, a p53 null cell line, by transfection with vectors expressing different p53 mutants. Vectors with the following p53 mutations were utilized: 143Ala, 175His, 248Try, 273His, and 281Gly. As controls, we used the original SAOS cells and cells transfected with the vector alone. Results were compared to those obtained with a cell line expressing wild-type p53 (wt p53). Radiosensitivity to killing was determined in the exponential phase of growth by measuring loss of colony-forming ability. Induction and repair of DNA double-strand breaks (dsb) was measured in irradiated cells using pulsed-field gel electrophoresis. Apoptosis was assessed using morphologic evaluation of DAPI-stained cells after treatment either with radiation or paclitaxel. RESULTS: Transfected SAOS-2 cell lines expressed a mutant form of p53 that could not be induced by radiation, and which was transcriptionally inactive. Among the 7 cell lines studied, we observed no difference in cellular radiosensitivity to killing (p = NS). When examining DNA repair, no difference in either the induction or repair of DNA dsb was noted in any of the cell lines studied (p = NS). Also, induction of apoptosis, either after exposure to radiation or paclitaxel, was low, and similar in all cell lines (p = NS). Non-isogenic cells expressing wt p53 were more radioresistant to killing by radiation, but showed similar kinetics of dsb rejoining. CONCLUSION: The results suggest that expression of different p53 mutants does not alter the yields of radiation-induced dsb, or the ability of cells to repair this type of lesion. In addition, the same p53 mutants do not affect cellular radiosensitivity to killing, or the induction of apoptosis after exposure to radiation or paclitaxel. PMID- 10524432 TI - Long-term adverse effects of radiation inhibition of restenosis: radiation injury to the aorta and branch arteries in a canine model. AB - PURPOSE: To determine the long-term effects of irradiation on large arteries in view of the possible use of radiation to prevent restenosis after angioplasty. METHODS AND MATERIALS: Groups of dogs received 10-55 Gy single-dose alone, or in combination with 50 Gy in 2-Gy fractions, or 50-80 Gy in 2-2.7-Gy fractions to an 8-cm length of aorta and branch arteries. Single doses were delivered intraoperatively. Two or 5 years after irradiation, aortas and branch arteries were evaluated histomorphometrically to determine areas of intima, media, and adventitia, and qualitatively to determine other adverse effects. RESULTS: Intimal area increased at single doses < 20 Gy and after all fractionated doses, but was normal at doses > 20 Gy 2 years after irradiation. Intimal area was greater at 5 years than at 2 years after irradiation. Adventitial area increased with increasing dose at 2 and 5 years after irradiation. Thrombosis of the aorta and branch arteries occurred at 4-5 years after irradiation with ED50s of 29.7 Gy and about 25 Gy, respectively, but did not occur after fractionated irradiation. CONCLUSION: Intimal proliferation is inhibited at single doses > 20 Gy, but may be stimulated at single doses of < 20 Gy or after fractionated irradiation. Adventitial fibrosis increases with increasing dose and could contribute to adverse late vascular remodeling. Severe adverse effects were not evident until 4 5 years after irradiation at does of > 20 Gy to an 8-cm vessel length. PMID- 10524433 TI - Boronated protoporphyrin (BOPP): localization in lysosomes of the human glioma cell line SF-767 with uptake modulated by lipoprotein levels. AB - PURPOSE: Boronated protoporphyrin (BOPP) is a candidate for use in both boron neutron capture therapy (BNCT) and photodynamic therapy (PDT) of glioblastoma multiforme (GBM). Our objectives are to identify factors that influence the uptake and retention of BOPP in vitro and to determine BOPP distribution in a human glioma cell line in vitro. This information will aid the development of compounds and treatment strategies that increase the effectiveness of BNCT therapy for GBM. METHODS AND MATERIALS: The amount, distribution pattern, and site of internalization of BOPP were assessed using fluorescence microscopy. Living human glioma (SF-767) cells were imaged after a 24-h exposure to BOPP (20 135.6 microg/ml, normal serum). Dose-dependent uptake of BOPP was determined using both fluorescence microscopy of individual living cells and inductively coupled plasma-atomic emission spectroscopy (ICP-AES) analysis of cell pellets. Lysosome- or mitochondria-specific fluorescent probes were used to identify the cellular compartment containing BOPP. Two human fibroblast cell lines, AG-1522 (LDL receptor-positive) and GM019-15C (LDL receptor-deficient), were used to investigate LDL receptor-dependent BOPP uptake. The dependence of BOPP uptake on lipoproteins in the media was determined by exposing each of the three cell types to BOPP in medium containing either normal (NS) or lipoprotein deficient serum (LPDS). RESULTS: BOPP accumulated in the lysosomes of human glioma cells in vitro, and not in the mitochondria, as reported for C6 rat glioma cells in vitro. BOPP uptake was concentration-dependent and was also dependent on the amount of lipoproteins in the medium. Over the range of incubation concentrations studied and at the single exposure duration time point investigated (24 h), all cells retained a similar amount of BOPP. At the lowest incubation concentration (20 microg/ml, NS), the amount of boron retained was near 10(9) atoms per cell (15 microg B/g cells). Lysosomes containing high concentrations of BOPP were randomly distributed throughout the cytoplasm; however, larger lysosomes containing BOPP were concentrated around the cell nucleus. Little or no BOPP accumulated in the cell nucleus. At incubation concentrations of 20 and 40 microg/ml (24-h time point), BOPP uptake in SF-767 cells was reduced in LPDS compared with NS (66% reduction). A similar result was observed for normal human fibroblasts (AG-1522 cells, 40 microg/ml, 24 h). At 40 microg/ml, in both NS and LPDS at 24 h, BOPP accumulation in LDL receptor-deficient human fibroblasts (GM019-15C cells) was reduced relative to AG-1522 cells. BOPP accumulation in GM019-15C cells (40 microg/ml, 24 h) was not affected by serum lipoprotein levels. CONCLUSION: In cell culture, BOPP is taken up by human glioma cells via the LDL pathway and is compartmentalized into cellular lysosomes. Knowledge of this mechanism of BOPP uptake and retention will be important in attempts to modify toxicity and efficacy of this drug. PMID- 10524434 TI - A radiographic and tomographic imaging system integrated into a medical linear accelerator for localization of bone and soft-tissue targets. AB - PURPOSE: Dose escalation in conformal radiation therapy requires accurate field placement. Electronic portal imaging devices are used to verify field placement but are limited by the low subject contrast of bony anatomy at megavoltage (MV) energies, the large imaging dose, and the small size of the radiation fields. In this article, we describe the in-house modification of a medical linear accelerator to provide radiographic and tomographic localization of bone and soft tissue targets in the reference frame of the accelerator. This system separates the verification of beam delivery (machine settings, field shaping) from patient and target localization. MATERIALS AND METHODS: A kilovoltage (kV) x-ray source is mounted on the drum assembly of an Elekta SL-20 medical linear accelerator, maintaining the same isocenter as the treatment beam with the central axis at 90 degrees to the treatment beam axis. The x-ray tube is powered by a high-frequency generator and can be retracted to the drum-face. Two CCD-based fluoroscopic imaging systems are mounted on the accelerator to collect MV and kV radiographic images. The system is also capable of cone-beam tomographic imaging at both MV and kV energies. The gain stages of the two imaging systems have been modeled to assess imaging performance. The contrast-resolution of the kV and MV systems was measured using a contrast-detail (C-D) phantom. The dosimetric advantage of using the kV imaging system over the MV system for the detection of bone-like objects is quantified for a specific imaging geometry using a C-D phantom. Accurate guidance of the treatment beam requires registration of the imaging and treatment coordinate systems. The mechanical characteristics of the treatment and imaging gantries are examined to determine a localizing precision assuming an unambiguous object. MV and kV radiographs of patients receiving radiation therapy are acquired to demonstrate the radiographic performance of the system. The tomographic performance is demonstrated on phantoms using both the MV and the kV imaging system, and the visibility of soft-tissue targets is assessed. RESULTS AND DISCUSSION: Characterization of the gains in the two systems demonstrates that the MV system is x-ray quantum noise-limited at very low spatial frequencies; this is not the case for the kV system. The estimates of gain used in the model are validated by measurements of the total gain in each system. Contrast-detail measurements demonstrate that the MV system is capable of detecting subject contrasts of less than 0.1% (at 6 and 18 MV). A comparison of the kV and MV contrast-detail performance indicates that equivalent bony object detection can be achieved with the kV system at significantly lower doses (factors of 40 and 90 lower than for 6 and 18 MV, respectively). The tomographic performance of the system is promising; soft-tissue visibility is demonstrated at relatively low imaging doses (3 cGy) using four laboratory rats. CONCLUSIONS: We have integrated a kV radiographic and tomographic imaging system with a medical linear accelerator to allow localization of bone and soft-tissue structures in the reference frame of the accelerator. Modeling and experiments have demonstrated the feasibility of acquiring high-quality radiographic and tomographic images at acceptable imaging doses. Full integration of the kV and MV imaging systems with the treatment machine will allow on-line radiographic and tomographic guidance of field placement. PMID- 10524435 TI - A comparison of field-only electronic portal imaging hard copies with double exposure port films in radiation therapy treatment setup confirmation to determine its clinical application in a radiotherapy center. AB - PURPOSE: To determine in which treatment sites field-only hard copy electronic portal images (EPI) captured during a treatment exposure could replace traditional double exposed port films in a busy radiation oncology department. METHODS AND MATERIALS: The three linear accelerators in the William Buckland Radiotherapy Centre (WBRC) at the Alfred Hospital in Melbourne are each equipped with an electronic portal imaging device (EPID). These devices can be used daily on all patients where the treatment fields are within the size constraint of the cassette, for example, less than 25 x 25 cm. Port films using radiographic film in hard cassettes were previously considered the standard method of field placement verification. After the radiation therapists were trained in all program aspects of capturing, enhancing, and producing hard copies of EPIs, a study was developed to evaluate the possibility of replacing port films with EPI hard copies within the established departmental procedures. Comparison of EPI hard copy with the simulator film and the port film of the same field was carried out by the radiation oncologist specialists. Seventy-eight comparison sets were generated and grouped into seven anatomical regions for evaluation by the radiation oncologist specialist responsible for each particular region. The outcome decision was the preferred imaging option. Where no preference was stated, EPI became the modality of choice, as it increased the efficiency of work practice. RESULTS: The results indicate that field-only EPI can be considered to be at least as clinically useful for treatment verification in the following sites: breast, chest, hip, spine, and large pelvic fields. Port films using a standard, double exposure technique were considered necessary for partial brain fields, small pelvis fields, extremities, and radical head and neck fields. CONCLUSION: The quality of field-only images captured using an EPID has been favorably assessed to be equivalent to, or an improvement on, the traditional double exposed port films for some treatment areas. Departmental policy has been altered to incorporate this new imaging modality as a practical alternative to port films, resulting in a direct benefit in terms of resource management and patient care. Continuing research is currently evaluating open area exposed EPI hard copies as a potential alternative to port films. PMID- 10524436 TI - Clinical application of digitally-reconstructed radiographs generated from magnetic resonance imaging for intracranial lesions. AB - PURPOSE: The purpose of this work is to demonstrate the clinical utility of magnetic resonance (MR) imaging-based digitally reconstructed radiographs (DRRs) for the setup and verification of patients with intracranial lesions. METHODS AND MATERIALS: MR images of 16 patients with various intracranial lesions were obtained for treatment planning and virtual simulation. Five-millimeter-thick contiguous T1-weighted postcontrast transverse slices were obtained using a standard head coil in a General Electric Signa 1.5T MR scanner. MR-DRRs were generated using the "pseudo density" technique on an existing treatment planning computer without any special modifications. Anterior and lateral verification films were taken for each patient for visual comparison with MR-based DRRs. RESULTS: Visual alignment with bony landmarks, including the orbits, frontal sinus, sphenoid sinus, auditory meatus, nasal bone, vomer bone, mastoid process, and the cranium were used by physicians, physicists, and therapists to verify patient positioning. Misalignments from 3 to 10 mm were visually identified and corrected using this technique. CONCLUSION: A method for visually utilizing MR based DRRs during simulation has been developed and clinically implemented. The quality of MR-DRRs generated using this technique is such that physicians, physicists, and therapists can easily and routinely compare MR-DRRs side-by-side with simulation films. PMID- 10524437 TI - Dose-volume analysis for quality assurance of interstitial brachytherapy for breast cancer. AB - PURPOSE/OBJECTIVE: The use of brachytherapy in the management of breast cancer has increased significantly over the past several years. Unfortunately, few techniques have been developed to compare dosimetric quality and target volume coverage concurrently. We present a new method of implant evaluation that incorporates computed tomography-based three-dimensional (3D) dose-volume analysis with traditional measures of brachytherapy quality. Analyses performed in this fashion will be needed to ultimately assist in determining the efficacy of breast implants. METHODS AND MATERIALS: Since March of 1993, brachytherapy has been used as the sole radiation modality after lumpectomy in selected protocol patients with early-stage breast cancer treated with breast-conserving therapy. Eight patients treated with high-dose-rate (HDR) brachytherapy who had surgical clips outlining the lumpectomy cavity and underwent computed tomography (CT) scanning after implant placement were selected for this study. For each patient, the postimplant CT dataset was transferred to a 3D treatment planning system. The lumpectomy cavity, target volume (lumpectomy cavity plus a 1-cm margin), and entire breast were outlined on each axial slice. Once all volumes were entered, the programmed HDR brachytherapy source positions and dwell times were imported into the 3D planning system. Using the tools provided by the 3D planning system, the implant dataset was then registered to the visible implant template in the CT dataset. The distribution of the implant dose was analyzed with respect to defined volumes via dose-volume histograms (DVH). Isodose surfaces, the dose homogeneity index, and dosimetric coverage of the defined volumes were calculated and contrasted. All patients received 32 Gy to the entire implanted volume in 8 fractions of 4 Gy over 4 days. RESULTS: Three-plane implants were used for 7 patients and a two-plane implant for 1 patient. The median number of needles per implant was 16.5 (range 11-18). Despite visual verification by the treating physician that surgical clips (with an appropriate margin) were within the boundaries of the implant needles, the median proportion of the lumpectomy cavity that received the prescribed dose was only 87% (range 73-98%). With respect to the target volume, a median of only 68% (range 56-81%) of this volume received 100% of the prescribed dose. On average, the minimum dose received by at least 90% of the target volume was 22 Gy (range 17.3-26.9), which corresponds to 69% of the prescribed dose. CONCLUSION: Preliminary results using our new technique to evaluate implant quality with CT-based 3D dose-volume analysis appear promising. Dosimetric quality and target volume coverage can be concurrently analyzed, allowing the possibility of evaluating implants prospectively. Considering that target volume coverage may be suboptimal even after radiographically verifying accurate implant placement, techniques similar to this need to be developed to ultimately determine the true efficacy of brachytherapy in the management of breast cancer. PMID- 10524438 TI - Volume-based dose optimization in brachytherapy. AB - PURPOSE: We address the question of how to optimize the dwell time distribution in brachytherapy with a stepping source if a minimal tumor dose is prescribed within the planning target volume (PTV). METHODS AND MATERIALS: For a given PTV, reference points inside and at the surface of the PTV are generated and dose constraints are prescribed. The dose at these reference points can be calculated if the positions of the sources are known. We determine a set of dwell times such that the dose constraints are fulfilled, and at the same time, the total irradiation time is minimized. The simplex algorithm allows us to find a solution (if any exists) for this problem. RESULTS: The performance of this method has been tested for a geometrically simple PTV. This method gives better results than conventionally used algorithms for dwell time optimization. CONCLUSION: The method described in this paper allows a volume-oriented optimization for brachytherapy dose distribution. The algorithm guarantees finding a dwell time distribution which fulfills the prescribed dose constraints, if any solution exists. PMID- 10524439 TI - Dosimetric comparison of three photon radiosurgery techniques for an elongated ellipsoid target. AB - PURPOSE: To examine the dosimetric differences among three radiosurgery techniques: gamma knife, linac multiple arcs, and conformally-shaped static fields. METHODS AND MATERIALS: A simulated target was taken to be a prolate ellipsoid, 25 mm in diameter, 35 mm in length, centrally located in a three dimensional (3D) model of a patient head taken from MR images. Single isocenter linac treatment plans were developed, 9 portals for the static shaped field technique, and a 7-arc plan for the multiple arc method. A total of 13 isocenters with 3 different collimators were used in the gamma knife plan. RESULTS: At dose levels from 25% to 50% of the reference dose, multiple arc and shaped-field plans treated a greater volume than the gamma knife plan. The linac plans, however, delivered the dose more homogeneously across the target volume as compared to the gamma knife plan. For the dose levels between 50-100%, the shaped fields and gamma knife plan have a similar dose distribution, and treated slightly less volume than the multiple arc plan. CONCLUSION: For a target of limited volume and essentially any shape, one can obtain closely conformal dosimetry with the gamma knife. For a regular-shaped target, the single isocenter multiple arc technique gives a more homogenous dose distribution within the target. Static shaped fields offer an alternative radiosurgery technique, with dosimetry similar to the multiple arc method, applicable to targets of any shape. PMID- 10524440 TI - Regarding Yamada et al. IJROBP 44:99-104;1999. PMID- 10524441 TI - Regarding Seong, Keum, Han, et al. IJROBP 43:393-397; 1999. PMID- 10524442 TI - The integral biologically effective dose (IBED) serves as an indicator of relative damage to an organ: plausible but incorrect. PMID- 10524443 TI - Predictors of subclinical nodal involvement in clinical stages I and II non-small cell lung cancer--why change the study objectives? Regarding Sawyer et al. IJROBP 43(5):965-970; 1995. PMID- 10524444 TI - Dyskeratosis congenita: recent advances and future directions. PMID- 10524445 TI - Therapeutic amenorrhea. PMID- 10524446 TI - Commentary on: ferrokinetics in the syndrome of familial hypoferremic microcytic anemia with iron malabsorption. PMID- 10524447 TI - Essential thrombocythemia in children. AB - PURPOSE: The objective of this study was to evaluate the clinical course, laboratory findings, and outcomes of children with essential thrombocythemia (ET). PATIENTS AND METHODS: The authors analyzed 36 children, ages 6 weeks to 18 years, by combining descriptions of 2 patients observed at their institution with 34 patients reported in the English medical literature. RESULTS: Fifteen patients (10 at diagnosis and 5 later on) had symptoms directly related to ET, including 9 who had severe thrombohemorrhagic phenomena. Common abnormalities included large platelets, increased marrow megakaryocytes with hyperlobulated forms, and abnormal platelet aggregation. Symptomatic patients had significantly higher platelet counts (2,419 versus 904 x 10(9)/L, P < 0.001); however, three patients with platelet counts that were only moderately elevated (600-800 x 10(9)/L) had thrombotic events. Eleven patients received various therapeutic agents. Interestingly, three patients who had one thrombotic event, and did not receive therapy, went on to have a benign clinical course. Leukemia developed in two treated patients, and they died; two others died of thrombotic complications; and myelofibrosis developed in one patient. Seventeen cases (47%) were familial. Patients with familial cases had significantly lower platelet counts, a lower incidence of hepatomegaly, and no thrombotic complications. CONCLUSIONS: This analysis of children with ET found that severe vascular complications developed in a substantial number. Platelet counts usually, but not always, correlate with the occurrence of complications. The indications for treatment and the best treatment of children with ET are currently not known, and guidelines for the management of children with ET are needed. Familial thrombocythemia is common among children with primary thrombocytosis and appears to be a different disease from ET, with a more benign course. PMID- 10524448 TI - Adjuvant chemotherapy with vincristine, doxorubicin, and cyclophosphamide in the treatment of postenucleation high risk retinoblastoma. AB - PURPOSE: To study risk factors and outcome of children with high risk retinoblastoma who receive postenucleation vincristine, doxorubicin, and cyclophosphamide. PATIENTS AND METHODS: Charts of all patients who received adjuvant chemotherapy for retinoblastoma were reviewed. Thirty-six patients were identified who received chemotherapy for high risk histopathologic features. Histopathology slides of these 36 patients were retrieved and reviewed, and the disease was staged according to the modified St. Jude staging system. The disease was unilateral in 23 patients (64%). There were 9 patients with stage I disease, 18 with stage II, and 9 with stage III. Twenty-four patients (67%) completed 12 of the 12 scheduled chemotherapy cycles, and 11 patients (30%) received 4 to 11 cycles because of relapse, disease progression, or family reasons. A life threatening complication developed in one patient after the first cycle, and this patient received no further chemotherapy. RESULTS: Five (3 with unilateral and 2 with bilateral disease) of the 36 patients developed distant metastasis and subsequently died. All had massive tumors; three had choroidal and up to surgical margin optic nerve invasion, and two had tumor extending posterior to lamina cribrosa. Six other patients had local relapse or progressive disease. All of these six patients had bilateral disease and failed in the intact eye during (three patients) or after (three patients) chemotherapy. Only two of the six patients were alive with no disease 50 and 102 months from diagnosis. With a median follow-up of 5.6 years, the 5-year and 10-year actuarial overall survival rates were 86% and 74%, respectively. The 5-year survival rates for patients with modified St. Jude stage I, II, and III disease were 100%, 91% (95% confidence interval, 57% to 100%), and 58% (95% confidence interval, 22% to 94%), respectively (P = 0.008). The survival rate was significantly different among patients with optic nerve involvement anterior to lamina cribrosa, posterior to lamina cribrosa, and surgical margin involvement (100%, 55%, and 41%, respectively; P = 0.003). Multivariate analysis showed that only the degree of optic nerve involvement (and therefore, modified St. Jude stage) was predictive of poor outcome. CONCLUSION: Patients with retinoblastoma involving the optic nerve beyond the lamina cribrosa have low survival rate despite local therapy and adjuvant chemotherapy with vincristine, doxorubicin, and cyclophosphamide. Progression of disease in the intact eye of three patients receiving chemotherapy is of concern. Alternative chemotherapeutic agents should be considered for patients with such high risk features. PMID- 10524450 TI - Regional cerebral blood flow and neuron-specific enolase in cerebrospinal fluid in children with acute lymphoblastic leukemia during induction treatment. AB - PURPOSE: To investigate possible side effects on the central nervous system from intrathecal methotrexate given during induction treatment for acute lymphoblastic leukemia in childhood. PATIENTS AND METHODS: Twenty-five children with acute lymphoblastic leukemia were examined by cerebral single photon emission computed tomography at the beginning of treatment (16 untreated, 9 during the first week) and after 4 weeks of treatment. Cerebrospinal fluid was sampled for analyses of neuron-specific enolase on four occasions in 54 patients. RESULTS: Regional cerebral blood flow became impaired during treatment in all patients. The single photon emission computed tomography score for nonhomogeneous perfusion increased from 6.4/50 to 16.6/50. Hypoperfusion was global without any clear preference for any lobe. The cerebellum was not affected. Neuron-specific enolase increased significantly during treatment, with a peak after 1 week, followed by a gradual decrease, but it was still significantly elevated after 4 weeks. CONCLUSIONS: Nonhomogeneous cerebral hypoperfusion was found in all patients during induction treatment, including repeated intrathecal administration of methotrexate, but before systemic high-dose methotrexate. Signs of neuronal injury, in the form of a moderate increase in neuron-specific enolase in the cerebrospinal fluid, were found early in the treatment. Follow-up is needed to evaluate the long-term impact of these findings. PMID- 10524449 TI - Brain metastases in pediatric Ewing sarcoma and rhabdomyosarcoma: the St. Jude Children's Research Hospital experience. AB - PURPOSE: Although brain metastases rarely occur in children with solid tumors, pediatric Ewing sarcoma (ES) and rhabdomyosarcoma (RMS) are among those most likely to metastasize to the brain. The authors review their institution's experience of brain metastases of ES and RMS. PATIENTS AND METHODS: The clinical characteristics, therapy, and outcome of all patients treated at St. Jude Children's Research Hospital over a 36-year period who had ES or RMS with brain metastases were reviewed. RESULTS: Of 419 patients with RMS, 10 (2.4%) had brain metastases. Of 335 patients with ES, 11 (3.3%) had brain metastases. The median age of the 21 patients was 10.4 years (range, 0.4-18.0 years) at the time of primary diagnosis. All had clinical signs of central nervous system (CNS) involvement. Outcome was dismal: The median duration of survival after diagnosis of brain metastasis was 2.7 months. The estimated survival 1 year after detection of brain involvement was 23.8%+/-8.5% (mean +/- standard error). One patient, who underwent chemotherapy, surgical resection, and radiotherapy, at the time of this writing is a long-term survivor. CONCLUSIONS: Brain metastases are rare in children with ES and RMS, but carry a grave prognosis. Because most brain metastases are accompanied by signs of neurologic involvement, routine imaging studies of asymptomatic children are not necessary. Combined-modality treatment offers the best chance of long-term survival. PMID- 10524451 TI - Serum soluble CD44 in pediatric patients with acute leukemia. AB - PURPOSE: CD44 is an adhesion molecule expressed on a variety of cells, and its level correlates with the metastatic potential of malignant tumors. Serum concentrations of soluble CD44 (sCD44) are elevated in various cancers. The purpose of this study was to measure the serum concentrations of CD44 in pediatric patients with acute leukemia. PATIENTS AND METHODS: Fourteen pediatric patients with acute leukemia were studied. The authors measured the serum concentration of sCD44 by enzyme-linked immunosorbent assay before and after therapy. The concentrations were compared with those of 15 control healthy children and 10 patients with bacterial infections. RESULTS: The mean serum concentration of sCD44 at presentation was significantly higher in patients than in control subjects, but decreased to a normal range in complete remission after chemotherapy. There was no difference in sCD44 concentrations between patients with acute lymphocytic leukemia and those with acute myeloid leukemia. Serum concentrations of sCD44 did not correlate with lactic dehydrogenase concentrations or bone marrow nucleated cell counts and only weakly with peripheral leukocyte count. sCD44 levels in patients with bacterial infections were similar to those of control subjects. CONCLUSION: Serum concentration of sCD44 may reflect disease status in pediatric patients with acute leukemia and might be a useful tumor marker in these patients. PMID- 10524452 TI - Costs, charges, and reimbursements for persons with sickle cell disease. AB - PURPOSE: The aims of this study were to describe health care costs and charges for patients with sickle cell disease (SCD) and identify predictors of high use. PATIENTS AND METHODS: Patients with SCD were identified by International Classification of Diseases, 9th revision, Clinical Modification (ICD-9-CM) codes from a university hospital's administrative databases from January 1, 1996, to September 30, 1997. Clinical and administrative data were gathered on each patient for all hospital admissions and ambulatory clinic visits. Logistic regression models were used to determine predictors of high health care use. RESULTS: A total of 947 patients with SCD were identified, 73% of whom resided within three South Carolina counties. On average, there were 0.9 admissions per patient per year and 8.0 outpatient visits per patient per year. Mean inpatient hospital charges, physician charges, and direct hospital costs per admission were $7290, $1589, and $5405, respectively, and the average length of stay was 4.5 days. Mean hospital charges, physician charges, and direct hospital costs per outpatient visit were $305, $169, and $688, respectively. Forty percent of the inpatient hospital charges were accounted for by only 4.2% of the patients. Residing in a distant county and being admitted with a diagnosis of painful respiration were found to be predictors of excessive charges and expenses beyond expected reimbursements. CONCLUSIONS: Patients with SCD are frequent users of health care services. Charges and costs are distributed disproportionately across these patients. Predictors of excessive hospital charges include living geographically distant from the hospital and being admitted with a diagnosis of painful respiration. PMID- 10524453 TI - Correlation of the C677T MTHFR genotype with homocysteine levels in children with sickle cell disease. AB - Recently, a mild to moderate elevation in the plasma homocysteine (Hcy) level has been found to be an important risk factor for stroke. Homozygosity for a common mutation (C677T) in the gene encoding for the enzyme methylenetetrahydrofolate reductase (MTHFR) involved in Hcy metabolism has been associated with increased levels of Hcy. To determine the role of hyperhomocysteinemia in the pathogenesis of stroke in children with sickle cell disease (SCD), Hcy levels and C677T MTHFR genotype were determined in 40 patients homozygous for hemoglobin SS and compared with 197 healthy children. Eleven of 40 patients with SCD had a history of stroke. The prevalence of homozygosity for the C677T MTHFR variant was 5% in the patients with SCD. The median Hcy level was 5.8 micromol/L in the patients versus 5.4 micromol/L in the controls (Fisher's, P > 0.05). There was no correlation of Hcy levels with the MTHFR genotype in patients with SCD. In patients with SCD and stroke, the median Hcy level was 4.8 micromol/L versus 6.0 micromol/L in those without stroke (P = 0.44, Mann-Whitney rank sum test). There was no difference in the proportion of patients with SCD with or without stroke who were homozygous for the C677T MTHFR mutation (0/11 versus 2/29; Fisher's, P = 1.000). In conclusion, this study failed to demonstrate an elevation in plasma Hcy levels in children with SCD compared with normal controls. Furthermore, hyperhomocysteinemia did not seem to be a significant factor in the pathogenesis of stroke in children with SCD. PMID- 10524454 TI - Splenic complications of the sickling syndromes and the role of splenectomy. AB - PURPOSE: To analyze the authors' experience with splenectomy for sickling disorders and evaluate the indications, complications, and outcome. PATIENTS AND METHODS: Over a period of 10 years (1987-1997), 113 patients with sickling disorders (100 with sickle cell disease and 13 with sickle-beta-thalassemia) had splenectomy at the authors' hospital as part of their management. The indications for splenectomy were hypersplenism (26 patients), major splenic sequestration crisis (MSSC) (23 patients), minor recurrent splenic sequestration crisis (MRSSC) (50 patients), splenic abscess (12 patients), and massive splenic infarction (2 patients). RESULTS: Splenectomy in patients with sickle cell disease (SCD) and sickle-beta-thalassemia (S-beta-Thal) was beneficial in reducing their transfusion requirements and its attendant risks, eliminating the discomfort from mechanical pressure of the enlarged spleen, and avoiding the risks of acute splenic sequestration crisis. It also was curative for patients with splenic abscess and massive splenic infarction. Twenty-four patients with SCD (24%) had splenectomy and cholecystectomy caused by concomitant gallstones. There was no mortality, and the postoperative morbidity was 7%. CONCLUSIONS: With careful perioperative management, splenectomy is both safe and beneficial in a select group of patients with SCD and S-beta-Thal. PMID- 10524455 TI - Physiologic decline in fetal hemoglobin parameters in infants with sickle cell disease: implications for pharmacological intervention. AB - PURPOSE: Fetal hemoglobin (HbF) is an important determinant in the clinical severity of patients with sickle cell disease. The physiologic decline in HbF parameters in a cohort of infants with sickle cell disease was investigated. PATIENTS AND METHODS: The percent HbF and F cells were quantitated, and the HbF per F cell was then calculated. One hundred thirty-eight blood samples from 44 infants with homozygous sickle cell anemia (HbSS) and 56 samples from 24 infants with sickle cell hemoglobin (HbSC) were studied. RESULTS: Infants with HbSS had a logarithmic decline in HbF parameters; at 24 months, the average HbF was 14.6%+/ 7.3% and the % F cells was 64.7%+/-16.9%. The amount of HbF in each F cell (HbF per F cell) was <15 pg/cell, a suggested threshold for intracellular sickle polymerization, by age 12 months. Infants with HbSC had a more rapid decline: at 12 months the average % HbF was 12.2%+/-9.3%, the % F cells was 60.5%+/-18.7%, and the HbF per F cell was <10 pg/cell. CONCLUSIONS: By age 2 years, HbF parameters including the % HbF, % F cells, and the HbF per F cell decrease to levels insufficient to inhibit sickling. Pharmacologic intervention designed to enhance HbF production and prevent chronic organ damage should be considered during infancy. PMID- 10524456 TI - Ferrokinetics in the syndrome of familial hypoferremic microcytic anemia with iron malabsorption. AB - PURPOSE: In 1981, Buchanan and Sheehan described a previously unreported syndrome in three siblings who had iron malabsorption, hypoferremia, and microcytic anemia that did not respond to oral iron and responded only partly to parenteral iron dextran. Ferrokinetic studies were not done in these or subsequently reported patients with this syndrome. It has been postulated that this syndrome of abnormal iron metabolism is analogous to that observed in the mk/mk mouse, which has similar hematologic findings but also has abnormal ferrokinetics. Ferrokinetic studies were performed in one patient to determine whether the abnormality of iron metabolism in the human syndrome is analogous to the mk/mk mouse. PATIENTS AND METHODS: Two sisters with severe microcytic anemia and iron malabsorption who have had only partial response to parenteral iron have been followed up for 15 years. Ferrokinetic studies with 59Fe were performed in one sister. RESULTS: Ferrokinetic studies with radio iron were characteristic of iron deficient erythropoiesis (rapid 59Fe T1/2; rapid, complete incorporation of 59Fe into erythrocyte hemoglobin). These ferrokinetics differ from those of the mk/mk mouse, which has a missense mutation in Nramp2, a putative iron transporter protein. In these children, once iron enters the plasma its subsequent metabolism (including binding to transferrin), transfer into erythroid bone marrow cells, and subsequent incorporation into erythrocyte hemoglobin are all normal. The defect in these patients appears to be an undefined, novel abnormality that governs mobilization of iron into the plasma from both the intestinal mucosal and reticuloendothelial cells. Despite lifelong severe hypoferremia, the growth, development and intellectual performance of these children, who are teen-agers, are normal. PMID- 10524457 TI - Refractory anemia with ringed sideroblasts in children: two diseases with a similar phenotype? AB - Three pediatric patients with refractory anemia with ringed sideroblasts (RARS) are presented. Bone marrow aspirates were examined using Romanowsky and Prussian blue iron stains in all three patients, and electron microscopic analysis was performed in one patient. All three patients had cytogenetic analysis of the bone marrow. Other studies included analysis of serum iron, total iron-binding capacity, ferritin, copper, vitamins B6 and B12, and folate levels. Antibody titers to Parvovirus, HIV, and other viruses were measured. The patients had contrasting clinical courses. Patients 1 and 2 had dysplastic hematopoietic features and cytogenetic findings (with either partial or one allele loss of chromosome 7), suggestive of myelodysplastic syndrome. Patient 1 experienced acute myeloid leukemia (AML) and had a good response to AML-directed therapy. Patient 2 had prolonged cytopenias and underwent bone marrow transplantation (BMT). Patient 3 had features suggestive of refractory anemia associated with mitochondrial cytopathy, including normal cytogenetics with pronounced vacuolization of marrow precursors. His anemia regressed spontaneously a few months after diagnosis. These patients represent two subgroups of pediatric RARS. Patients with the myelodysplastic syndrome (MDS) type may progress to cytopenias or leukemia and may require aggressive therapy; the type is characterized by clonal cytogenetic findings. The non-MDS type, which may relate to mitochondrial cytopathy, often shows spontaneous regression and requires only supportive treatment; it has normal cytogenetic findings. PMID- 10524458 TI - Rhabdomyosarcoma in a patient with cardio-facio-cutaneous syndrome. AB - A boy with characteristic facial features, pulmonary valvular stenosis, ectodermal abnormalities, growth failure, and mental retardation was admitted for intestinal occlusion at 20 months of age. Clinical findings were consistent with a diagnosis of cardio-facio-cutaneous syndrome (CFC-s), and a huge abdominal mass was evident on computed tomography scan. A biopsy was performed, and embryonal rhabdomyosarcoma was diagnosed. Molecular analysis was performed by reverse transcription (RT) polymerase chain reaction (PCR) on tumor RNA to seek the chimerical transcript of the most common soft tissue sarcoma translocations and analyze neurofibromatosis 1 (NF1) gene expression. Translocations involving 1;13, 2;13, and 11;22 were not found, and the specific transcripts of the NF1 gene were present. Chemotherapy was implemented, but the child died 7 months later of tumor progression. Few patients with CFC-s have been described, and their follow-up is not well known. The association of CFC-s with rhabdomyosarcoma has not been reported previously, but other neoplasms have been reported in patients with Noonan syndrome, a condition similar to CFC-s. More observations are needed, but this and other reports suggest there could be a higher risk of malignancy in patients with syndromes in the Noonan phenotype category. PMID- 10524459 TI - Congenital acute megakaryoblastic leukemia (M7) with chromosomal t(1;22)(p13;q13) translocation in a set of identical twins. AB - Chromosomal translocations at t(1;22)(p13;q13) have been reported to occur in a number of infants with acute megakaryoblastic leukemia. A set of female twins with acute megakaryoblastic leukemia are reported with this unique translocation of 1p13 to 22q13. The twins presented at 2 months of age with fever and poor feeding and subsequently developed progressive hepatosplenomegaly. One twin died before treatment could be started; the other became septicemic 5 days after initiation of chemotherapy and eventually died. PMID- 10524460 TI - Bacillus cereus causing fulminant sepsis and hemolysis in two patients with acute leukemia. AB - PURPOSE: Hemolysis is so rarely associated with Bacillus cereus sepsis that only two very well documented cases have been reported. This article reports two unusual cases of Bacillus cereus sepsis with massive intravascular hemolysis in patients who had acute lymphoblastic leukemia (ALL). PATIENTS AND METHODS: A 20 year-old woman who was 9 weeks pregnant experienced a relapse of ALL. A therapeutic abortion was performed. During week 4 of reinduction the patient had abdominal pain, nausea, and vomiting, with severe neutropenia but no fever. Her condition deteriorated rapidly with cardiovascular collapse, acute massive intravascular hemolysis, and death within hours of the onset of symptoms. Blood cultures were positive for Bacillus cereus. Postmortem histologic examination and cultures revealed Bacillus cereus and Candida albicans in multiple organs. The second patient, a 10-year-old girl, presented with relapsed T-cell ALL. In the second week of reinduction, she had abdominal pain followed by hypotension. Again, no fever was noted. Laboratory studies showed intravascular hemolysis 12 hours after admission. Aggressive support was promptly initiated. Despite disseminated intravascular coagulation; cardiovascular, hepatic, and renal failure; and multiple intracerebral hypodense lesions believed to be infarcts, the patient recovered fully and resumed reinduction therapy. CONCLUSIONS: Bacillus cereus infection can have a fulminant clinical course that may be complicated by massive intravascular hemolysis. This pathogen should be suspected in immunosuppressed patients who experience gastrointestinal symptoms and should not be precluded by the absence of fever, especially if steroids such as dexamethasone are being given. Exchange transfusion may be lifesaving in Bacillus cereus septicemia associated with massive hemolysis. PMID- 10524461 TI - Oral contraceptives: a cause of hyperbilirubinemia in stem cell transplant patients. AB - Conjugated hyperbilirubinemia in the clinical setting of hematopoietic stem cell transplantation can have multiple etiologies that may prompt various therapeutic interventions. Two patients who received short courses of a high-dose estrogen progesterone combination to treat breakthrough menstrual bleeding during transplant are reported. Conjugated hyperbilirubinemia developed in both patients within days of beginning therapy and resolved after the ethinyl estradiol and norgestrel (Ovral; Pharmacia and Upjohn, Kalamazoo, MI, U.S.A.) was discontinued. In one of the patients, this occurred on three separate occasions during the course of transplantation. Recognizing the cholestatic effect of estrogens during transplantation may prevent unnecessary alterations in therapy beyond the simple discontinuation of these medications. PMID- 10524462 TI - Partial splenectomy in a child with human immunodeficiency virus-related immune thrombocytopenia. AB - Immune thrombocytopenia (IT) is a frequently occurring disease in childhood and a well known complication of HIV infection. Splenectomy is a part of the treatment strategy for severe chronic IT. However, overwhelming infections after splenectomy have limited its use, especially in young children. A 7-year-old child with maternal-fetal HIV-1 infection and related thrombocytopenia underwent splenectomy after previous treatment failed to improve her platelet count. Approximately 75% of the spleen was removed. The postoperative period was uncomplicated, and the platelet count increased significantly to greater than 500,000/mm3. Ultrasonographic examination performed 3 months later showed a stable volume of the spleen stump (40 x 40 x 20 mm) with effective "vascularization." The platelet count 12 months after surgery showed a sustained increase greater than 150,000 cell/mm3. Subtotal splenectomy may be a safe and effective alternative for patients with HIV and immune thrombocytopenia. PMID- 10524463 TI - Aplastic anemia in a patient with Rothmund-Thomson syndrome. AB - This report is the first to describe constitutional aplastic anemia in a patient with Rothmund-Thomson syndrome (also called poikiloderma congenitale), a disease characterized by multiple cutaneous and extracutaneous findings. The findings suggest that although Rothmund-Thomson syndrome is a rare disease, vigilance for the development of associated hematologic abnormalities is warranted. PMID- 10524464 TI - Alpha1-antitrypsin deficiency with fatal intracranial hemorrhage in a newborn. AB - A 4-week-old boy had a fatal intracranial hemorrhage resulting from vitamin K deficiency. The infant had received no vitamin K prophylaxis and was exclusively breastfed. At autopsy, examination of the liver showed cholestasis and fibrosis. DNA was isolated from a blood spot on a Gutherie sample card obtained from the infant for routine metabolic screening. This DNA was used for alpha1-antitrypsin genotyping studies. Genotyping studies identified homozygosity for the point mutation 9989G-->A, confirming a diagnosis of alpha1-antitrypsin deficiency (ZZ phenotype), and resulted in appropriate screening of siblings born after this child's death. Alpha1-antitrypsin deficiency should be considered in the differential diagnosis of infants with late hemorrhagic disease of the newborn. Use of blood from the metabolic screening card as a source of DNA allowed confirmation of this diagnosis after the infant's death. PMID- 10524465 TI - Simultaneous detection of platelet-associated antigen and platelet peroxidase on buffy coat cells from bone marrow in two patients with pediatric acute megakaryocytic leukemia. PMID- 10524466 TI - Novel three-way translocation t(5;8;21) in acute myeloblastic leukemia (M2) with chloroma. PMID- 10524467 TI - Continuity and change. PMID- 10524468 TI - Intrathecal coadministration of bupivacaine diminishes morphine dose progression during long-term intrathecal infusion in cancer patients. AB - OBJECTIVE: To determine the difference in intrathecal morphine dose progression between a continuous intrathecal infusion of a morphine/bupivacaine mixture and morphine for pain relief in patients with cancer. DESIGN: Patients were treated with intrathecal drugs in a randomized study and followed prospectively until death. SETTING: Institute for Anesthesiology, Department of Pain Treatment, University Hospital Nijmegen, St Radboud, The Netherlands. PATIENTS: Twenty patients with cancer were selected for intrathecal treatment because of either side effects or inadequate relief during conventional pain treatment. INTERVENTIONS: Intrathecal drug infusion rates and medication were adjusted according to pain relief and side effects. OUTCOME MEASURES: Progression of intrathecal morphine dose during a phase of adequate analgesia in both groups following regression analysis and analysis of possible treatment related side effects. RESULTS: The combination of intrathecal morphine and bupivacaine resulted in a diminished progression of the intrathecal morphine dose (slope of regression line = 0.0003 vs. 0.005, p = 0.0001) during a phase of stable analgesia in comparison with the morphine group. No serious side effects presented. CONCLUSION: The diminished intrathecal morphine dose increase in the combination group is considered to be due to a synergistic effect of bupivacaine on the intrathecal morphine-induced antinociception. A dose increment during long term intrathecal infusion in cancer patients appears to be related to both disease progression and tolerance phenomena. PMID- 10524469 TI - Intravenous titration with morphine for severe cancer pain: report of 28 cases. AB - BACKGROUND: In a multicenter study, 28 patients with cancer pain and insufficient pain relief with analgesic treatment according to step II of the guidelines of the World Health Organization (WHO) were switched to oral slow-release morphine. METHODS: Patients received intravenous morphine through a patient-controlled pump (PCA) for the first 24 hours (bolus = 1 mg, lockout interval = 5 minutes, maximum dose = 12 mg/hour). From day 2 patients were treated with oral slow-release morphine. Daily doses were calculated from the requirements of the day before. Breakthrough pain was treated with PCA until stable doses were reached (<2 boluses/day) and then with oral immediate-release morphine solution. Pain intensity was reported in a diary four times a day, in addition to mood, activity, and quality of sleep once daily. RESULTS: Mean duration until adequate pain relief reported (<30 on a 101-step numerical scale; NRS) was 5 hours (range = 80-620 minutes). Mean pain intensity was reduced from 67 NRS to 22 NRS. Mean doses of oral morphine were 133 mg/day initially and then 154 mg/day on day 14. Serious adverse events such as respiratory depression were not observed. Two patients terminated the study due to progressive symptoms of gastrointestinal obstruction. Seventy-five percent of the patients evaluated the effectiveness of the analgesic regime as good. CONCLUSIONS: Dose finding with intravenous PCA may be appropriate for a small minority of patients with severe pain. Higher treatment costs and the risk of complications are drawbacks of this method compared with conventional oral titration. PMID- 10524470 TI - Efficacy and safety of controlled-release versus immediate-release oxycodone: randomized, double-blind evaluation in patients with chronic back pain. AB - OBJECTIVE: To compare the efficacy and safety of controlled-release oxycodone given every 12 hours with immediate-release oxycodone given four times daily in patients with persistent back pain. DESIGN: Randomized, double-blind, active controlled, two-period crossover trial. PATIENTS: Fifty-seven adult outpatients with stable, chronic, moderate-to-severe low back pain despite analgesic therapy were enrolled; 47 were randomized; 11 discontinued for side effects, most commonly nausea and vomiting. INTERVENTIONS: Controlled-release oxycodone tablets given every 12 hours; immediate-release oxycodone tablets given four times daily; dose titration with controlled-release or immediate-release for up to 10 days; double-blind treatment for 4-7 days each. OUTCOME MEASURES: Patients' pain scores (0 = none, 1 = slight, 2 = moderate, 3 = severe). RESULTS: Pain intensity decreased from moderate to severe at baseline to slight at the end of titration with both oxycodone formulations. The daily oxycodone dose was 40 mg or less in 68% of patients. During double-blind treatment, mean pain intensity was maintained at 1.2 (0.1 SE) with controlled-release and at 1.1 (0.1 SE) with immediate-release oxycodone. The most common adverse events were constipation, nausea, pruritus, somnolence, and dizziness. CONCLUSIONS: Controlled-release oxycodone given every 12 hours was comparable with immediate-release oxycodone given four times daily in efficacy and safety, and it provides convenient, twice daily, around-the-clock treatment for selected patients with persistent back pain that is inadequately controlled by nonopioids or as-needed opioid therapy. PMID- 10524471 TI - Validity of self-reported drug use in chronic pain patients. AB - OBJECTIVE: Previous researchers have reported that in psychiatric populations many patients provide incorrect self-report information on current drug use. Therefore, the purposes of the present study were to determine the percentage of chronic pain patients (CPPs) using illicit drugs (cannabis, cocaine), to determine the percentage of CPPs who provide incorrect self-report drug use information in the psychiatric examination, and to identify some variables that could help in identifying the CPP likely to provide an incorrect drug use history using drug urine toxicologies. DESIGN/SETTING/PARTICIPANTS/OUTCOME MEASURES: Two hundred seventy-four CPP consecutive admissions to a pain facility were psychiatrically examined according to criteria in the Diagnostic and statistical manual of mental disorders (3rd ed., rev; DSM-III-R), with special emphasis on all current drug use. Immediately after the psychiatric examination, all CPPs were asked to consent to urine toxicology. Urine was tested for benzodiazepines, opioids, tricyclics, propoxyphene, cannabinoids, barbiturates, amphetamines, methadone, methaqualone, phencyclidine, alcohol, and cocaine. CPPs were then segregated into three groups: negative toxicology, positive toxicology but concordant with self-report of current drug use, and positive toxicology discordant with self-report of current drug use. These groups were statistically compared with each other with regard to age, gender, race, workers' compensation status, and prevalence of individual DSM-III-R psychoactive substance use disorders. Sensitivities were also calculated for two conditions: accuracy of toxicology and accuracy of self-report. RESULTS: Toxicologies were obtained from 226 (82.5%) of the CPPs. Toxicologies were negative in 121 (53.5%) and positive in 105 (46.5%) of the CPPs. Of the 226 CPPs, 8.4% had illicit drugs in the urine (6.2% cannabis, 2.2% cocaine). Twenty (8.8%) of the CPPs provided incorrect self report information about current drug use, the incorrect information most frequently about illicit drugs. Drug urine toxicology sensitivity results indicated that a significant percentage of CPPs was claiming to be taking a drug but was not taking it or taking it incorrectly. The psychiatric examination drug self-report sensitivity results indicated that a significant percentage of CPPs was withholding or providing incorrect information on current drug use. Lowest self-report sensitivity results were in reference to illicit drugs. CPPs who were more likely to provide incorrect psychiatric examination self-report information about current drug use were more likely to be younger, to be a workers' compensation CPP, and to have been assigned a DSM-III-R diagnosis of polysubstance abuse in remission. CONCLUSIONS: A significant percentage of CPPs appears to provide incorrect information on current illicit drug use. Urine toxicology studies may have a place in the identification of drugs for which incorrect information may be provided by CPPs. There are many possible reasons, such as assay error, that could lead to apparent misinformation. In the clinical setting, these possibilities should be considered if urine toxicology results appear to be incongruent with psychiatric examination drug use self-report. PMID- 10524472 TI - Postoperative pain expression in preschool children: validation of the child facial coding system. AB - OBJECTIVE: The purposes of the study were threefold: (a) to determine whether a measurement system based on facial expression would be useful in the assessment of post-operative pain in young children; (b) to examine construct validity in terms of structure, consistency, and dynamics of the facial display; and (c) to evaluate concurrent validity in terms of associations with global judgments of the children's pain. PATIENTS: One hundred children between the ages of 13 and 74 months were video-taped for a maximum of 1 hour after arrival in the postanesthesia care unit (PACU) at British Columbia's Children's Hospital. OUTCOME MEASURES: Videotapes were edited into 20-second blocks, randomly selected from each 2-minute time period taped during the hour following surgery, and coded for the presence or absence of 13 facial actions in the Child Facial Coding System (CFCS). RESULTS: Facial expressions were characterized primarily by the following constellation of actions: open lips, lowered brows, a deepened nasolabial furrow, mouth stretched wide in both horizontal and vertical directions, eyes squeezed shut or squinted, and raised cheeks. A principal components analysis indicated that these actions comprised a single factor, accounting for 55% of the variance in CFCS actions. Facial action summary scores were correlated with a visual analog rating of global pain, confirming that the CFCS has convergent validity. Facial action summary scores, i.e., pain displays, were at their lowest immediately after admittance to the PACU and just before the child's release from the PACU. CONCLUSIONS: The present study demonstrated that the CFCS serves as a valid measurement tool for persistent pain in children. PMID- 10524473 TI - Biobehavioral responses to acute pain in adolescents with a significant neurologic impairment. AB - OBJECTIVE: To further understand acute pain response in children with a significant neurologic impairment (SNI), we undertook a descriptive hypothesis generating study of the response to a routine vaccine among adolescents with SNI. DESIGN: Within-subject crossover design. SETTING: Tertiary care facility for children and adolescents with SNI. PATIENTS: Eight adolescents (mean age = 15 years). INTERVENTIONS: Mock and real vaccine injections. OUTCOME MEASURES: Quantitative measures of heart rate, videotaped facial action, Child Facial Coding System (CFCS), and Facial Action Coding System (FACS); observer ratings visual analog scale (VAS) were obtained before, during, and after a mock injection and routine annual influenza vaccine injection presented in a counterbalanced order. RESULTS: VAS scores were significantly higher during the injection phase than during the other time periods; however, there were no significant differences across study time periods when using the other outcome measures. CONCLUSIONS: Although the dampened behavioral and physiologic reactions to an acute noxious stimulus were similar to those of previous work with developmentally delayed children and frail elderly, it remains unclear what underlies the apparent reduced pain response in this setting. These findings have potentially important implications for the daily care of individuals with significant neurologic impairment and illustrate the compelling need tor further study of the unique character of the pain experience in this setting. PMID- 10524474 TI - Relation between pain location and disc pathology: a study of pain drawings and CT/discography. AB - OBJECTIVE: The purpose of this study was to determine whether pain location indicated in pain drawings was related to the specific lumbar disc level(s) that was abnormal in appearance and painful upon discographic injection. DESIGN: Data were collected prospectively. SETTING: This study was conducted in a spine specialty clinic. PATIENTS: The study group consisted of 187 patients (118 men, 69 women; mean age = 37.2 years, range = 18-62 years) with low back pain with or without leg pain. All patients were undergoing computed tomography (CT)/discography at the three lowest lumbar levels for diagnostic purposes. INTERVENTIONS: Pain drawings were completed the day of but prior to undergoing discography. Discographic pain responses were recorded with respect to the similarity to the patient's clinical symptoms. Pain drawings were classified based on the presence or absence of pain in five areas: low back and/or buttocks, posterior thigh, posterior leg, anterior thigh, and anterior leg. The drawings were scored with the system described by Ransford et al. (1976, Spine 1: 127-34), and those likely to be indicative of psychological problems were analyzed separately (n = 43). OUTCOME MEASURES: Results were determined by analyzing the relation between the location of pain in the drawings and the specific lumbar disc level(s) found to be painful and disrupted by discography. RESULTS: There was a significant relation between pain location indicated in the drawing and the lumbar disc level(s) identified as clinically painful and disrupted by CT/discography (p < 0.05, chi-square). Pain limited to the low back and buttocks was frequently associated with the absence of disc pathology (58.3%). When pain in the posterior thigh or leg was present but there was no pain in the anterior drawing, patients frequently had a positive L5-S 1 disc (> or =75%). In patients with anterior thigh pain, with or without posterior thigh or leg pain, the L4-5 disc was frequently symptomatic (>63%). The pattern of no posterior thigh or leg pain but with pain radiating into the leg anteriorly was most commonly associated with the L3-4 disc (71.4%). CONCLUSIONS: The results of this study indicate that pain drawings may be helpful in identifying which specific discs are associated with pain complaints. As with any evaluation, the drawings should be considered in combination with findings from other assessments. PMID- 10524475 TI - Short- and long-term outcomes of children with complex regional pain syndrome type I treated with exercise therapy. AB - OBJECTIVE: To report the initial and long-term outcome after an intensive exercise therapy program for childhood complex regional pain syndrome, type I (CRPS). DESIGN: Prospective follow-up. SETTING: A children's hospital. SUBJECTS: We followed 103 children (87 girls; mean age = 13.0 years) with CRPS. Forty-nine subjects were followed for more than 2 years (mean = 5 years 3 months). INTERVENTIONS: An intensive exercise program (most received a daily program of 4 hours of aerobic, functionally directed exercises, 1-2 hours of hydrotherapy, and desensitization). No medications or modalities were used. All had a screening psychological evaluation, and 79 (77%) were referred for psychological counseling. MAIN OUTCOME MEASURES: Outcomes included pain, presence of physical dysfunction, or recurrent episodes of CRPS or other disproportional musculoskeletal pain. RESULTS: The mean duration of exercise therapy was 14 days, but over the past 2 years has decreased to 6 days. Ninety-five children (92%) initially became symptom free. Of those followed for more than 2 years, 43 (88%) were symptom free (15, or 31 %, of these patients had had a reoccurrence), 5 (10%) were fully functional but had some continued pain, and 1 (2%) had functional limitations. The median time to recurrence was 2 months; 79% of the recurrences were during the first 6 months after treatment. CONCLUSION: Intense exercise therapy is effective in initially treating childhood CRPS and is associated with low rate of long-term symptoms or dysfunction. PMID- 10524476 TI - Signs and symptoms in complex regional pain syndrome type I/reflex sympathetic dystrophy: judgment of the physician versus objective measurement. AB - OBJECTIVE: To assess the relation between the subjectively assessed and objectively measured diagnostic signs and symptoms in complex regional pain syndrome type I (CRPS I) and to quantify their severity. DESIGN: Diagnostic signs and symptoms were recorded in patients suffering from CRPS I of one upper extremity for less than 1 year. Independent assessors measured (a) pain by using four visual analog scales (VAS) and the McGill Questionnaire list of adjectives (MPQ), (b) edema with a hand volumeter, (c) skin temperature with an infrared thermometer, and (d) active range of motion (AROM) with goniometers. SETTING: Two university hospitals. PATIENTS: Ninety-five women and 40 men with CRPS I of one upper extremity. RESULTS: Four signs and symptoms were diagnosed in 50 patients, and five in the remaining 85 patients. The mean score for present pain intensity was 31.5 mm and that for pain resulting from exertion of the affected extremity was 71.9 mm. A median of 11.5 words was chosen from the MPQ, with the highest number from its evaluative part. The difference in volume between both hands was 30.4 ml. The mean difference in temperature between the two hands was 0.78 degrees C dorsally and 0.66 degrees C palmarly. The largest decrease in mobility was seen in the wrist and fingers; the thumb was relatively less affected and the little finger relatively more affected than the other fingers. CONCLUSIONS: Bedside evaluation of CRPS I with Veldman's criteria was in good accord with psychometric or laboratory testing of these criteria. PMID- 10524477 TI - Effects of gender and acute dental pain on thermal pain responses. AB - OBJECTIVE: Considerable research suggests that females exhibit greater sensitivity to laboratory pain procedures than do males; however, whether the presence of acute clinical pain influences this sex difference in pain sensitivity has not been investigated. The present experiment investigated the effects of sex and acute dental pain on laboratory pain responses. DESIGN: Thermal pain onset and tolerance were determined in 46 dental patients (15 male, 31 female) experiencing pain due to acute irreversible pulpitis and in 33 healthy controls (13 male, 20 female). In addition, measures of mood and coping were obtained in all participants. All subjects participated in two experimental sessions. The first session took place immediately before the patients underwent endodontic treatment for relief of pulpal pain. The second session took place approximately 1-2 weeks later, when pulpitis patients were pain free after treatment. During each session, thermal pain onset and tolerance were assessed with a 1-cm2 contact thermode applied to the right volar forearm using an ascending method of limits. RESULTS: During both sessions, thermal pain onset and tolerance were lower in control females than in control males; however, male and female pulpitis patients did not differ in their thermal pain responses during either session. Pulpitis patients also showed greater affective distress than controls. CONCLUSIONS: These data suggest that the sex difference in thermal pain sensitivity frequently reported in pain-free subjects appears to be absent in patients presenting with acute dental pain. However, this effect cannot be explained solely based on the presence of clinical pain because the effect on pain threshold and tolerance persisted into session 2, when pulpitis patients were pain free. Potential explanations for these results are discussed. PMID- 10524478 TI - Abacus VAS in burn pain assessment. PMID- 10524479 TI - Clinical criteria as a preliminary screen for cervical spine injury. PMID- 10524480 TI - Speech and language problems in international adoptees. PMID- 10524481 TI - Thinking about sexually transmitted diseases. PMID- 10524482 TI - Medical care in the home. PMID- 10524483 TI - Abnormal uterine bleeding. AB - The most probable etiology of abnormal uterine bleeding relates to the patient's reproductive age, as does the likelihood of serious endometrial pathology. The specific diagnostic approach depends on whether the patient is premenopausal, perimenopausal or postmenopausal. In premenopausal women with normal findings on physical examination, the most likely diagnosis is dysfunctional uterine bleeding (DUB) secondary to anovulation, and the diagnostic investigation is targeted at identifying the etiology of anovulation. In perimenopausal patients, endometrial biopsy and other methods of detecting endometrial hyperplasia or carcinoma must be considered early in the investigation. Uterine pathology, particularly endometrial carcinoma, is common in postmenopausal women with abnormal uterine bleeding. Thus, in this age group, endometrial biopsy or transvaginal ultrasonography is included in the initial investigation. Premenopausal women with DUB may respond to oral contraceptives, cyclic medroxyprogesterone therapy or cyclic clomiphene. Perimenopausal women may also be treated with low-dose oral contraceptives or medroxyprogesterone. Erratic bleeding during hormone replacement therapy in postmenopausal women with no demonstrable pathology may respond to manipulation of the hormone regimen. PMID- 10524484 TI - Drug treatment of common STDs: part I. Herpes, syphilis, urethritis, chlamydia and gonorrhea. AB - In 1998, the Centers for Disease Control and Prevention released guidelines for the treatment of sexually transmitted diseases. Several treatment advances have been made since the previous guidelines were published. Part I of this two-part article describes current recommendations for the treatment of genital ulcer diseases, urethritis and cervicitis. Treatment advances include effective single dose regimens for many sexually transmitted diseases and improved therapies for herpes infections. Two single-dose regimens, 1 g of oral azithromycin and 250 mg of intramuscular ceftriaxone, are effective for the treatment of chancroid. A three-day course of 500 mg of oral ciprofloxacin twice daily may be used to treat chancroid in patients who are not pregnant. Parenteral penicillin continues to be the drug of choice for treatment of all stages of syphilis. Three antiviral medications have been shown to provide clinical benefit in the treatment of genital herpes: acyclovir, valacyclovir and famciclovir. Valacyclovir and famciclovir are not yet recommended for use during pregnancy. Azithromycin in a single oral 1-g dose is now a recommended regimen for the treatment of nongonococcal urethritis. PMID- 10524485 TI - Pigmented villonodular synovitis of the hip and knee. AB - Pigmented villonodular synovitis is an uncommon disease that remains a diagnostic challenge. Presenting complaints commonly involve one joint, most often the knee or hip. Symptoms of pain and swelling characteristically have an insidious onset and are slowly progressive. The physical examination may be completely normal. Radiographs of the knee may appear normal or may show a periarticular soft tissue density, expansion of the suprapatellar pouch and local osseous changes confined to the patellofemoral articulation. Radiographs of the hip may show erosions in the head and neck of the femur and acetabulum. Magnetic resonance imaging usually demonstrates key diagnostic features, which include joint effusion, elevation of the joint capsule, hyperplastic synovium and low signal intensity resulting from hemosiderin deposition. The diagnosis of pigmented villonodular synovitis is confirmed by biopsy, and the treatment of choice is synovectomy. PMID- 10524486 TI - An approach to diagnosis and initial management of systemic vasculitis. AB - Systemic vasculitis occurs in a heterogeneous group of primary disorders or can be a manifestation of infection, an adverse drug reaction, malignancy or a connective tissue disease. A vasculitic process should be suspected in patients with unexplained ischemia or multiple organ involvement, especially when such features as polymyalgia rheumatica, inflammatory arthritis, palpable purpura, glomerulonephritis or multiple mononeuropathy are also present. The clinical features of systemic vasculitis depend on the organs involved and, in turn, organ involvement is largely influenced by the size of the affected blood vessels. The diagnostic work-up should be tailored to the clinical situation and geared toward a tissue or angiographic diagnosis, bearing in mind that the findings from these studies are not always pathognomonic. Emphasis should also be placed on exclusion of a secondary process. The diagnosis of the specific type of vasculitis may be made on the basis of the clinical features and the histopathologic or angiographic findings. Initial therapy for most types of systemic vasculitis consists of high-dose corticosteroids, with the addition of immunosuppressive therapy in certain patients. PMID- 10524487 TI - Chronic insomnia: a practical review. AB - Insomnia has numerous, often concurrent etiologies, including medical conditions, medications, psychiatric disorders and poor sleep hygiene. In the elderly, insomnia is complex and often difficult to relieve because the physiologic parameters of sleep normally change with age. In most cases, however, a practical management approach is to first consider depression, medications, or both, as potential causes. Sleep apnea also should be considered in the differential assessment. Regardless of the cause of insomnia, most patients benefit from behavioral approaches that focus on good sleep habits. Exposure to bright light at appropriate times can help realign the circadian rhythm in patients whose sleep-wake cycle has shifted to undesirable times. Periodic limb movements during sleep are very common in the elderly and may merit treatment if the movements cause frequent arousals from sleep. When medication is deemed necessary for relief of insomnia, a low-dose sedating antidepressant or a nonbenzodiazepine anxiolytic may offer advantages over traditional sedative-hypnotics. Longterm use of long-acting benzodiazepines should, in particular, be avoided. Melatonin may be helpful when insomnia is related to shift work and jet lag; however, its use remains controversial. PMID- 10524488 TI - Clinical utility of the erythrocyte sedimentation rate. AB - The erythrocyte sedimentation rate (ESR) determination is a commonly performed laboratory test with a time-honored role. However, the usefulness of this test has decreased as new methods of evaluating disease have been developed. The test remains helpful in the specific diagnosis of a few conditions, including temporal arteritis, polymyalgia rheumatica and, possibly, rheumatoid arthritis. It is useful in monitoring these conditions and may predict relapse in patients with Hodgkin's disease. Use of the ESR as a screening test to identify patients who have serious disease is not supported by the literature. Some studies suggest that the test may be useful as a "sickness index" in the elderly or as a screening tool for a few specific infections in certain settings. An extreme elevation of the ESR is strongly associated with serious underlying disease, most often infection, collagen vascular disease or metastatic malignancy. When an increased rate is encountered with no obvious clinical explanation, the physician should repeat the test after an appropriate interval rather than pursue an exhaustive search for occult disease. PMID- 10524489 TI - Parvovirus B19 infections. AB - Infections caused by human parvovirus B19 can result in a wide spectrum of manifestations, which are usually influenced by the patient's immunologic and hematologic status. In the normal host, parvovirus infection can be asymptomatic or can result in erythema infectiosum or arthropathy. Patients with underlying hematologic and immunologic disorders who become infected with this virus are at risk for aplastic anemia. Hydrops fetalis and fetal death are complications of intrauterine parvovirus B19 infection. PMID- 10524490 TI - Issues in newborn screening for phenylketonuria. AB - The blood sample for phenylketonuria (PKU) screening should be obtained at least 12 hours after the infant's birth. Newborn screening for PKU has largely eliminated mental retardation caused by this disease. If the first phenylalanine test demonstrates positive results, a repeat test should be performed. Treatment to prevent sequelae from this disorder is best carried out in cooperation with an experienced PKU center. Dietary care is expensive, and financial assistance may be necessary for many families. A phenylalanine-restricted diet should be started as soon as possible. Occasionally, cases of PKU are missed by newborn screening. Thus, a repeat PKU test should be performed in an infant who exhibits slow development. PMID- 10524491 TI - Management of the hyperosmolar hyperglycemic syndrome. AB - Hyperglycemic hyperosmolarity is part of a clinical spectrum of severe hyperglycemic disorders ranging from pure hyperglycemic hyperosmolarity without ketosis to diabetic ketoacidosis, with significant overlap in the middle. From 50 to 75 percent of hospitalizable patients who have uncontrolled diabetes present with significant hyperosmolarity. An altered state of consciousness attributable to uncontrolled diabetes is virtually always the result of severe hyperosmolar hyperglycemia. The linchpin of therapy is prompt, rapid administration of crystalloid solutions that have tonicity appropriate to the level of hyperosmolarity. A decrease in the plasma glucose concentration indicates the adequacy of therapy, especially rehydration; the goal is for the plasma glucose level to decline by at least 75 to 100 mg per dL (4.2 to 5.6 mmol per L) per hour. Patients with hyperosmolar hyperglycemic syndrome are often chronically ill, and they may have major total body deficits of potassium, phosphate and magnesium, as well as B-complex vitamins (especially thiamine). These deficits also require attention and correction during therapy. PMID- 10524492 TI - The home visit. AB - With the advent of effective home health programs, an increasing proportion of medical care is being delivered in patients' homes. Since the time before World War II, direct physician involvement in home health care has been minimal. However, patient preferences and key changes in the health care system are now creating an increased need for physician-conducted home visits. To conduct home visits effectively, physicians must acquire fundamental and well-defined attitudes, knowledge and skills in addition to an inexpensive set of portable equipment. "INHOMESSS" (standing for: immobility, nutrition, housing, others, medication, examination, safety, spirituality, services) is an easily remembered mnemonic that provides a framework for the evaluation of a patient's functional status and home environment. Expanded use of the telephone and telemedicine technology may allow busy physicians to conduct time-efficient "virtual" house calls that complement and sometimes replace in-person visits. PMID- 10524493 TI - Photo quiz. White patch on back. PMID- 10524494 TI - Approaching a terminally ill patient in denial. PMID- 10524495 TI - 'Be the doctor, always'. PMID- 10524496 TI - Escape from self-tolerance leads to neonatal insulin-dependent diabetes mellitus. AB - Double transgenic (dTg) mice expressing the hemagglutinin (HA) of influenza virus under the insulin promoter and the TCR specific for the immunodominant CD4 T cell epitope of HA (HA110-120) develop insulin-dependent diabetes mellitus (IDDM). In order to gain information on the breaking down of neonatal self-tolerance we studied the occurrence of IDDM after birth. Our results showed that newborn mice develop fulminant IDDM characterized by occurrence of insulitis as early as 3 days after birth, followed by hyperglycemia by 7 days, and significant hypoinsulinemia by 28 days. The neonatal breakdown of self-tolerance of T cells positively selected in the thymus is supported by the facts that: (i) peripheral HA110-120 specific T cells from neonates are fully functional and proliferated upon stimulation with the nominal peptide, and (ii) peptide-specific T cells were accumulated in the pancreas of dTg mice as early as 3 days after birth. Our results demonstrate that diabetes occurring in young dTg mice is due to early activation of self-reactive T cells immediately after birth. Accumulation of specific T cells in the target organ leads to destruction of pancreatic beta cells and IDDM. These mice may provide a useful model to evaluate new strategies for the prevention of diabetes. PMID- 10524497 TI - Reversal of immunodominance among autoantigenic T-cell epitopes. AB - Studies spanning several decades have revealed how the complex forces of antigen processing distinguish those epitopes of a protein that dominate the immune response from those that remain cryptic. Since foreign antigens and self-proteins are subjected to the same proteolytic pathways before presentation to the T-cell repertoire, it has long been assumed that they comply equally with the established rules of immunodominance. Nevertheless, the pathological determinants of some autoantigens appear ill-equipped for the dominant role they adopt, displaying features more befitting subdominant or cryptic epitopes, such as low affinity for their MHC restriction element. These findings may be reconciled by suggesting that, far from remaining sequestered during ontogeny, many classical autoantigens participate in the establishment of self-tolerance, the efficiency with which individual epitopes purge the T-cell repertoire being determined by the conventional rules of immunodominance: while those epitopes that are truly dominant induce profound non-responsiveness, those that are poorly presented may leave residual reactivity, manifest in the periphery as responses to epitopes that appear inappropriately dominant. Here we review recent evidence showing the process of self-tolerance to be uniquely responsible for the reversal of immunodominance which promotes such epitopes to an undeserved position of importance within the determinant hierarchy. PMID- 10524498 TI - In vitro stimulation by islet antigen facilitates the detectability of beta-cell reactive cells in diabetes-prone BB/OK rats. AB - It has been supposed that beta-cell destruction in man and animals is due to autoreactive T-cells. We used the [51Cr]-release assay to identify the presence of beta-cell reactive cells in the spleen of diabetes-prone BB/OK rats before and after diabetes manifestation as well as in long-term normoglycaemic rats with a reduced diabetes risk of 3%. Splenic mononuclear cells (MNCs) obtained from diabetes-resistant LEW.1W and the majority of long-term normoglycaemic BB/OK rats (86.4%) showed no reactivity to pancreatic islets in vitro. In contrast, beta cell reactive cells were identified in dependence on age in 30.4-65.0% of 75-120 days old normoglycaemic rats and in relation to diabetes duration (1 and 20 days) in 75.0% and 16.0% of diabetic BB/OK rats. Islet antigen-specific stimulation of splenic MNCs, that showed no spontaneous islet-directed reactivity, resulted in a concentration-dependent activation of cytolytically reactive cells in BB/OK but not in LEW.1W rats. Splenic MNCs derived from all diabetic, from 82.4% of young normoglycaemic and from 46.2% of long-term normoglycaemic BB/OK rats developed an islet-directed reactivity in vitro. Phenotyping of MNCs showed a significant increase of activated IL2R+ T-lymphocytes in diabetic BB/OK rats, but without any correlation to their cytolytic potential in the [51Cr]-release assay. Despite this fact, IL2R+ cells enriched from the pool of MNCs mediated an enhanced [51Cr] release from islets, indicating their relevance in the beta-cell destruction. These data suggest, that functional reactivity rather than phenotypic characterization of MNCs is useful to identify the existence of beta-cell reactive cells. Furthermore, for screening diabetes risk in young normoglycaemic BB/OK rats besides the detection of beta-cell reactive cells the occurrence of regulatory cells seems to be decisive. PMID- 10524499 TI - Production of IL-1beta, IL-1 receptor antagonist and IL-10 by mononuclear cells from patients with SLE. AB - Depositions of immune-complexes are responsible for many of the pathological features of systemic lupus erythematosus (SLE). For example, immune-complex induced tissue damage in glomerulonephritis has been shown to be mediated, at least in part, by interleukin (IL)-1. Inappropriate production or function of IL 1 may therefore contribute to disease manifestations in SLE. We investigated lipopolysaccharide (LPS)- and adherent IgG-stimulated release of IL-1beta, IL-1 receptor antagonist (IL-1ra) and IL-10, a potent modulator of IL-1, by blood mononuclear cells from patients with SLE. Mediator production was measured as ng cytokines/10(6) monocytes and compared with clinical parameters. Release of IL 1beta was only detectable in LPS-stimulated cultures and substantially reduced in patients with both active and inactive disease (P < 0.001). LPS-stimulated IL-1ra release was normal and the IL-1ra/IL-1beta ratio was therefore increased (P < 0.05) and correlated inversely to prednisolone dosage (P = 0.009). IgG-stimulated release of IL-1ra was reduced in patients with active disease compared to those with inactive disease and controls (P = 0.002). IL-10 release was similar in patients and controls. We conclude that monocytes from patients with active SLE are deficient in Fc gamma-R-mediated production of IL-1ra, whereas LPS-stimulated IL-1beta release by SLE monocytes is reduced regardless of disease activity. The former may contribute to immune-complex-mediated tissue damage in SLE. PMID- 10524500 TI - Intermittent feeding and fasting reduces diabetes incidence in BB rats. AB - Food intake may be one of several factors which influence the risk of development of insulin dependent diabetes mellitus, but the influence of the pattern of food supply has not been studied previously. The aim of the present study was to investigate the effect of intermittent feeding and fasting upon diabetes in BB rats. This study included three groups. Group 1 served as control and included 77 animals, 79% became diabetic. In groups 2 and 3, after weaning, food but not water was withdrawn from the animals: 24 h twice a week in group 2; 24 h every second day in group 3. Group 2 included 40 BB rats, 50% (p < 0.004) became diabetic. Group 3 included 44 BB rats, 52% (p < 0.01) became diabetic. No differences were seen between sexes. Degree of insulitis was not influenced by changed food supply. Regarding blood glucose, no influence was seen among diabetic animals, among non-diabetic animals changed food supply reduced blood glucose values obtained at the end of the study. Intermittent feeding and fasting tended to reduce mean age at the time of diagnosis of diabetes, significance was reached only in female animals from group 3 compared to group 1. Body weight was obtained weekly. Intermittent feeding and fasting caused a reduced weight gain in group 2 as well as in group 3 compared to control animals; however, most pronounced in group 3 and also more pronounced among males compared to females. For pre-diabetic and non-diabetic animals comparable influence on body weight was seen. The main conclusion in the study is that intermittent feeding and fasting reduced diabetes incidence in BB rats. PMID- 10524502 TI - What's new? PMID- 10524501 TI - The role of Fas-mediated apoptosis in thyroid autoimmune disease. AB - Apoptosis is a carefully regulated mechanism of cell death that differs from necrosis and plays an important role in normal tissue development and homeostasis, as well as disease processes. Apoptosis also plays an important role in autoimmunity. Defective apoptosis can cause systemic autoimmunity by allowing the survival of autoreactive lymphocytes. It may also be involved in the pathogenesis of organ-specific autoimmune diseases, such as Hashimoto's thyroiditis, through altered target organ susceptibility. Apoptosis signaling pathways can be initiated through activation of death receptors. One of these pathways employs the death receptor Fas and its ligand (FasL). Fas expression and death pathway signaling have been demonstrated in the thyroid, but there is controversy surrounding the expression of FasL and its role in thyroid autoimmunity. A number of proteins, including FAP-1, Bcl-2 and I-FLICE may regulate the Fas pathway in the thyroid and provide potential mechanisms for modifying the pathogenesis of autoimmune thyroid disease. PMID- 10524503 TI - A brief history of nuclear criticality accidents in Russia--1953-1997. AB - Fourteen nuclear criticality accidents that occurred in Russia between 1953 and 1997 are described. These accidents are significant because of the loss of control of special nuclear material and the resultant radiation doses to personnel, potential damage to equipment, and release of radioactive material to the workplace and the environment. A qualitative analysis of the causes and contributing factors to these accidents is presented along with a description of the radiation health effects to workers. The primary cause of most of these accidents was inadequate design that allowed the use of process equipment that did not preclude nuclear criticality on the basis of geometry. Personnel errors and violations of procedures were major contributing factors to these accidents. PMID- 10524504 TI - Elevated urine uranium excretion by soldiers with retained uranium shrapnel. AB - The use of depleted uranium in munitions has given rise to a new exposure route for this chemically and radioactively hazardous metal. A cohort of U.S. soldiers wounded while on or in vehicles struck by depleted uranium penetrators during the Persian Gulf War was identified. Thirty-three members of this cohort were clinically evaluated, with particular attention to renal abnormalities, approximately 3 y after their injury. The presence of retained shrapnel was identified by x ray, and urine uranium concentrations were measured on two occasions. The absorption of uranium from embedded shrapnel was strongly suggested by measurements of urine uranium excretion at two time intervals: one in 1993/1994 and one in 1995. Mean urine uranium excretion was significantly higher in soldiers with retained shrapnel compared to those without shrapnel at both time points (4.47 vs. 0.03 microg g(-1) creatinine in 1993/1994 and 6.40 vs. 0.01 microg g(-1) creatinine in 1995, respectively). Urine uranium concentrations measured in 1995 were consistent with those measured in 1994/1993, with a correlation coefficient of 0.9. Spot urine measurements of uranium excretion were also well correlated with 24-h urine collections (r = 0.95), indicating that spot urine samples can be reliably used to monitor depleted uranium excretion in the surveillance program for this cohort of soldiers. The presence of uranium in the urine can be used to determine the rate at which embedded depleted uranium fragments are releasing biologically active uranium ions. No evidence of a relationship between urine uranium excretion and renal function could be demonstrated. Evaluation of this cohort continues. PMID- 10524505 TI - 134Cs and 85Sr in fruit plants following wet aerial deposition. AB - The knowledge of processes concerning the radiocontamination of fruit after a spike release can improve the understanding of exposure through ingestion of food and better assess the public dose. The fate of 134Cs and 85Sr in the above ground part of different species of fruit plants after wet deposition on leaves or on fruits was compared. Grapevines, apple trees, and pear trees grown under field conditions were contaminated with 134Cs and 85Sr either via leaves or via fruits before ripening. Spiked and non-spiked fruits and leaves were picked 50 d later, at harvest time, and their 134Cs and 85Sr contents were measured by gamma spectrometry. The residual fraction in leaves was on average 7% of the initially applied 134Cs and 8% of 85Sr, while that in fruits was 60% of 134Cs and 28% of 85Sr. Rinsing of fruits before consumption causes a loss of 24% for 134Cs and 36% for 85Sr present in fruit at harvest. Leaf-to-fruit transfer factors are considerably higher for 134Cs, 4% of the applied activity, than for 85Sr, 0.04%. Leaf-to-leaf are also higher on average for 134Cs than for 85Sr. Transfer also occurs from spiked fruits to leaves; its extent is affected more by the kind of plant than by the radionuclide. 134Cs and 85Sr are transferred to fruits and leaves of non-contaminated branches to a lesser extent than within the contaminated branches. PMID- 10524506 TI - Finger doses received during 153Sm injections. AB - This study was undertaken to determine the dose received by the skin of the fingers of clinical and laboratory staff during injections of 153Sm. The use of 153Sm, chelated with ethylenediaminetetramethylene phosphonic acid (153Sm-EDTMP), is coming into more frequent use in radionuclide therapy since its approval by the U.S. Food and Drug Administration in March 1997. 153Sm emits a range of medium-energy therapeutically useful beta particles that have been found beneficial in the palliation of metastatic bone cancer pain. It also emits a range of gamma rays. Calculations have been undertaken to provide the beta particle and gamma-ray dose rates, at a depth within the skin corresponding roughly to the basal cell layer, when the finger is placed in direct contact with the external surface of a syringe containing 153Sm. The beta-particle dose rates were modeled using an empirically based Monte Carlo approach previously described by Beddoe and Kelly. The gamma-ray dose rates were modeled using a distributed point source approach previously reported by Pattison et al. In the calculations it is assumed that a typical administered activity is 2.6 GBq, with a finger syringe contact time of 30 s. The skin dose, due to both beta particles and gamma rays when the finger is centrally placed over an active volume of 0.3 mL in a 1 mL syringe, is calculated to be 77 mGy per injection. Similarly, for an active volume of 1.0 mL in a 2.5-mL syringe, the dose is calculated to be 10 mGy per injection. In view of the ICRP recommended weekly skin dose limit of 10 mGy, both of the above two doses are excessive. If, however, the fingers are placed at the rear end of the syringe barrel, where they are only exposed to the gamma rays, the above two doses are reduced to 0.069 and 0.139 mGy per injection, respectively. Both of these two doses are well within the recommended weekly dose limits for the skin. It is found that the weekly dose limit for the skin is readily exceeded if the fingers are in direct contact with the external surface of the syringe and located over the active volume. However, if handled at the rear end of the syringe barrel, a typical weekly workload can be managed without exceeding the recommended dose limits. PMID- 10524507 TI - Improved estimates of effective dose equivalent using two optimal anisotropic responding dosimeters. AB - Although the use of two dosimeters, one on the chest and the other on the back, successfully solved the underestimation problem for posterior incident photon beams, the two-dosimeter approach still has some problems-significant overestimations for lateral, overhead, and underfoot beam directions when isotropic-responding dosimeters are used for measurement. A solution to this problem is to intentionally construct the dosimeters to under-respond as the beam direction departs from normal incidence and approaches lateral, overhead, or underfoot beam directions. The objective of this study is to develop a dosimeter that does not significantly overestimate effective dose equivalent (H(E)) for lateral, overhead, and underfoot beam directions, while maintaining good performance for anterior and posterior beam directions. Several dosimeter geometries were investigated using Monte Carlo simulation to find the best geometry using aluminum oxide (Al2O3) as dosimeter attenuator material. Then, the developed Optimal Anisotropic Responding dosimeters were tested for 0.08, 0.30 and 1.00 MeV photon beams of various beam directions. The dosimeters did not overestimate H(E) by more than 80% considering all photon energies and beam directions, which is much less than the overestimation of isotropic-responding dosimeters (202%). The dosimeters also showed similar performance compared to isotropic-responding dosimeters for anterior and posterior beam directions. Finally, the dosimeters were applied to effective dose (E) and the results are compared with those of H(E). PMID- 10524508 TI - Validation of a geologically based radon risk map: are the indoor radon concentrations higher in high-risk areas? AB - Since geographically coded information is frequently used in studies of the relationships between environmental factors and illness at the population level and by authorities for promotion of mitigation, knowledge about the validity of proxy measures is essential. This study was an evaluation of a geologically based map describing the risk for high radon levels, which was used by the municipal authorities to determine the necessity of remedial actions. Annual mean radon gas concentrations for a random sample of one-family homes selected from high-risk areas (n = 252) were compared with those of a random sample of homes from normal and low-risk areas (n = 259). No difference in geometric mean radon concentration was found between the areas, 101 Bq m(-3) and 103 Bq m(-3), respectively. The proportion of homes in each area with radon gas concentrations above the current Swedish administrative limit value for mitigation (400 Bq m(-3)) was similar, approximately 10%. We conclude that the radon risk map was unsuitable for identifying areas of concern. The findings also indicate that geologically based and geographically coded information as a proxy for human exposures can be safely used for scientific and administrative purposes only following validation. PMID- 10524509 TI - An application of the NCRP screening techniques to atmospheric radon releases from the former feed materials production center near Fernald, Ohio. National Council on Radiation Protection and Measurements. AB - The National Council on Radiation Protection and Measurements has published a series of screening models for releases of radionuclides to the environment. These models have been used to prioritize radionuclides being considered in environmental dose reconstructions. The NCRP atmospheric models are also accepted by the U.S. Nuclear Regulatory Commission for demonstrating compliance with the constraint on releases of airborne radioactive materials to the environment from licensees other than power reactors. This study tested the NCRP atmospheric techniques by comparing annual average predicted air concentrations of radon with measured radon concentrations at 14 locations 43 m to 598 m downwind of the former U.S. Department of Energy Feed Materials Production Center (FMPC) near Fernald, Ohio, for the period 2 July 1985 to 2 July 1986. Predictions were made using five different sets of meteorological data as input: (1) NCRP default values; (2) composite FMPC site data; (3) data from the Greater Cincinnati Airport; (4) data from the Dayton, Ohio, airport; and (5) data collected at Miami University, located near Oxford, Ohio. Following are the respective medians and ranges of the ratio of the predicted to observed annual radon air concentrations for each of these sources of meteorological data: (1) 5.2, 0.9-54; (2) 1.4, 0.1 8.2; (3) 0.7, 0.1-7.2; (4) 0.7, 0.1-8.4; and (5) 0.6, 0.1-10. The stated goal of the NCRP models is to predict doses that do not underpredict actual doses by greater than a factor of 10. In this comparison, all of the meteorological data produced air concentration predictions that meet this criteria. However, to ensure that final doses meet this criterion, one would need to carefully evaluate all assumptions used to calculate dose from each of these air concentrations. PMID- 10524510 TI - Investigation of atmospheric, mechanical and other pressure effects influencing the levels of radon and radon progeny in buildings. AB - Real-time data measurement and analysis have identified a number of influences affecting the variability and accumulation of radon and its progeny in indoor air. Observed cycles in radon concentrations were shown to be related to the influence of air-conditioning and water-heated central heating systems. The cyclical pattern, related to operation of the air-condition system, showed radon levels almost four times lower during the period when the system was switched On than when it was Off. When the heating system was switched On the radon and radon progeny levels were 40% lower than when it was switched Off. Under both regimes, it was possible to establish the over-riding influence of meteorological factors by separating the recurring cyclical component from the relevant data set. The general or trend level of indoor radon was determined substantially by the prevailing atmospheric conditions. PMID- 10524511 TI - Mass attenuation coefficients of clear-Pb for photons from 125I, 103Pd, 99mTc, 192Ir, 137Cs and 60Co. AB - The mass attenuation coefficients, mu/rho, for Clear-Pb for photon energies ranging from 10 keV to 10 MeV were determined using Monte Carlo methods and simple equations used to manipulate elemental mass attenuation coefficients. It was determined that the effectiveness of Clear-Pb as a radiation shielding material was greater than plain acrylic for all photon energies, especially those less than 150 keV, and for deep penetration problems where the differences in mu/rho between Clear-Pb and acrylic became more significant. Finally, the usefulness of Clear-Pb as a shielding material when compared with acrylic was determined for the following commonly used radionuclides: 125I, 103Pd, 99mTc, 192Ir, 137Cs, and 60Co. PMID- 10524512 TI - MAXED, a computer code for maximum entropy deconvolution of multisphere neutron spectrometer data. AB - Reliable neutron dosimetry requires knowledge of the neutron spectrum. We discuss the problem of analyzing data from a multisphere neutron spectrometer to infer the energy spectrum of the incident neutrons and describe the code MAXED, a computer program developed to apply the maximum entropy principle to this problem. The code and documentation are available from the authors upon request. PMID- 10524513 TI - An instrument for measuring equilibrium-equivalent 222Rn and 220Rn concentrations with etched track detectors. AB - To simultaneously measure both 222Rn and 220Rn progeny concentrations, a new type of portable integrating monitor with allyl diglycol carbonate (CR-39) plastic detectors was developed. The monitor gives the average equilibrium-equivalent 222Rn and 220Rn concentrations (EEC(RN) and EEC(Tn)) during sampling intervals. The detection efficiencies of the alpha particles were calculated by Monte Carlo method. The lower limits of detection for EEC(Rn) and EEC(Tn) are estimated to be 0.57 Bq m(-3) and 0.07 Bq m(-3) for 24 h continuously sampling at a flow rate of 0.8 L min(-1). The measuring results with the new type monitors were confirmed through intercomparison experiments. In a small survey, a rather high 220Rn progeny concentration with an average of 1.73 Bq m(-3) was observed in traditional Japanese dwellings with soil/mud plastered walls. On the other hand, a very high 232Th concentration in soil was reported in China. They suggested that there is a possibility of high 220Rn progeny concentration in both Japan and China. PMID- 10524514 TI - Radiation safety system of the B-Factory at the Stanford Linear Accelerator Center. AB - The radiation safety system of the B-Factory accelerator facility at the Stanford Linear Accelerator Center is described. The radiation safety system, which is designed to protect people from prompt radiation exposure due to beam operation, consists of the access control system and the radiation containment system. The access control system prevents people from being exposed to the very high radiation levels inside a beamline shielding enclosure. The access control system consists of barriers, a standard entry module at every entrance, and beam stoppers. The radiation containment system prevents people from being exposed to the radiation outside a shielding enclosure due to either normal or abnormal operation. The radiation containment system consists of power limiting devices, shielding, dump and collimator, and an active radiation monitoring system. The inter-related elements for the access control system and radiation containment system, as well as the associated interlock network, are described. The policies and practices used in establishing the radiation safety system are also compared with the regulatory requirements. PMID- 10524515 TI - Characterization and minimization of extremity doses during 32P metabolic cell labeling. AB - A study was performed to characterize hand extremity doses received by a laboratory researcher performing a metabolic cell labeling procedure using 10-cm diameter cell culture dishes containing GBq amounts of 32P-orthophosphate. Specifically, the optimal location for placement of thermoluminescent dosimeters on the extremities was determined as was the phase (time frame) during the cell labeling procedure when the highest dose was received. The cell labeling procedure was divided into four phases. Thermoluminescent dosimeters were placed on three fingers of each hand at the start of each phase and after the completion of that phase, collected, and sent to the processor. It was determined that in the case of this right-handed worker, the left-hand index finger should be used for determining the maximum extremity dose. The time frame during which the highest extremity dose was received was the cell lysis phase. Acrylic shielding was fabricated, which significantly reduced worker extremity dose. PMID- 10524516 TI - Xenon spill distribution and room clearance. AB - The purpose of these studies was to investigate actual xenon gas clearance times under different exhaust conditions, to compare them with the calculated clearance times, to observe the distribution of the xenon gas while it was being exhausted from the room, and to determine the cause of a stationary xenon cloud that appeared on some clinical images. Clearance times with and without a flexible exhaust hose placed next to a simulated 133Xe gas spill were compared with clearance times measured in a room with all exhaust closed off. Two gamma cameras were used to observe the transport and exhaust of xenon following a simulated spill. Clearance times with the flexible exhaust hose were less than one minute because the xenon gas was removed before it had a chance to disperse into the room. Conventional room clearance calculations based on uniform mixing and measured exhaust rates yielded a clearance time of 22 min. The source of an artifactual stationary cloud image was discovered to be a small amount of xenon trapped between the collimator and camera face. A negative pressure and dedicated exhaust can be even more effective in exhausting spilled xenon from a room than air transfer calculations predict. The authors believe the flexible hose should always be used. PMID- 10524517 TI - Surgical pathology remains pivotal in the evaluation of 'sentinel' lymph nodes. PMID- 10524518 TI - Diagnosis of prostate cancer in needle biopsies after radiation therapy. AB - Interpretation of postirradiation needle biopsies is a major diagnostic challenge for the pathologist because of substantial radiation-induced changes in benign and malignant prostatic tissue. Reports that have systematically evaluated the histopathologic findings in postirradiation needle biopsies are limited. In this study, we evaluated 46 histologic features in 29 postirradiation needle biopsy specimens from 29 patients. All patients had recurrent cancer on needle biopsies after external beam radiation, and all subsequently underwent salvage radical prostatectomy and bilateral pelvic lymphadenectomy. Patient age ranged from 57 to 78 years (mean, 61 years). The interval from radiation therapy to biopsy ranged from 1.0 to 17 years (mean, 3.9 years). Histologic features that were helpful in the diagnosis of cancer after radiation therapy included infiltrative growth, perineural invasion, intraluminal crystalloids, blue mucin secretions, the absence of corpora amylacea, and the presence of coexistent high-grade prostatic intraepithelial neoplasia. Benign glands usually showed nuclear enlargement (86%) and prominent nucleoli (50%), and therefore, these cytologic features alone were not reliable for the diagnosis of cancer after irradiation. Postirradiation needle biopsies underestimated the prostatectomy Gleason grade in 35% of cases and overestimated it in 14% of cases; these results were similar to published reports from patients not receiving radiation therapy. There was a major discrepancy in degree of radiation effect between radical prostatectomy and biopsies. Moderate or severe radiation effect on cancer was present in 48% of needle biopsy specimens, whereas 94% of radical prostatectomy specimens had no or minimal radiation effect on cancer when the areas with the least amount of radiation effect were chosen for quantification. These findings indicate that quantification of radiation effect in needle biopsy specimens was inaccurate and potentially misleading. Conversely, Gleason grade in postirradiation needle biopsy specimens appeared to provide useful predictive information and should be reported. PMID- 10524519 TI - Nodal cytotoxic lymphoma spectrum: a clinicopathologic study of 66 patients. AB - The expression of cytotoxic granule-associated proteins has been reported in some T-cell or natural killer (NK)-cell lymphomas of mostly extranodal origin, but rarely of nodal origin except for anaplastic large cell lymphoma (ALCL) and Hodgkin's disease (HD). This study analyzed 66 nodal lymphomas expressing T-cell intracellular antigen-1 (TIA-1) and/or granzyme B to characterize the clinicopathologic spectrum of these neoplasms. Four main groups could be delineated. The first group consisted of p80/anaplastic lymphoma kinase (ALK) positive ALCL (n = 35). The patients were 2 to 62 years of age (median age, 16 years), and the lymphomas pursued a relatively indolent clinical course. The tumors were phenotypically of either T- or null-cell type with constant expression of CD30, epithelial membrane antigen (EMA), and p80/ALK, but not CD15 or BCL2. None harbored Epstein-Barr virus (EBV). The second group consisted of peripheral T/NK-cell lymphoma, the nodal high-grade cytotoxic type (n = 13). The patients were 29 to 72 years in age (median age, 55 years), and the tumors pursued an aggressive clinical course. The tumors often showed pleomorphic, anaplastic, or centroblastoid morphology, and were featured by either EBV association or CD56 expression. The third group consisted of peripheral T-cell lymphoma, of the nodal low-grade cytotoxic type (n = 8). The patients, three men and five women, were 31 to 75 years old (median age, 61 years). Notably, six of them exhibited lymphoepithelioid (Lennert's) lymphoma. The fourth group consisted of cytotoxic Hodgkin's-like ALCL/HD (n = 10), included seven cases of Hodgkin's like ALCL and three cases of HD, and was characterized by the presence of Reed Sternberg cells and often the CD15+ phenotype. The patients were all men except for one woman, and they ranged in age from 24 to 84 years (median age, 62 years). The link among these four groups was reinforced by the presence of a highly characteristic large cell with horseshoelike or reniform nuclei-the frequent expression of CD30 and EMA-and the often lack of T-cell receptor-alphabeta. In this series, the expression of p80/ALK and CD56 was also associated with favorable and poor prognoses respectively (p<0.001, log-rank test). PMID- 10524520 TI - Lipomatous hemangiopericytoma: a rare variant of hemangiopericytoma that may be confused with liposarcoma. AB - We describe the clinicopathologic features and biologic behavior of 16 cases of histologically benign hemangiopericytoma containing a variable amount of mature fat as an intrinsic part of the neoplasm. These so-called lipomatous hemangiopericytomas occurred primarily in men (12 men and 4 women) with a mean age of 54 years (range, 33-74 years). All occurred in deep soft tissue and had an average size of 10 cm when first detected. All were characterized by a relatively sharp border and typical histologic features of hemangiopericytomas, including oval to round cells surrounding a sinusoidal and staghorn vasculature often with perivascular hyalinization. Mature fat varied in amount but usually occupied approximately one quarter to three quarters of the area of tumor. Mitotic activity was low, with more than half the cases having no mitotic activity. Five cases showed moderate nuclear atypia. In four cases, the pericytic regions had sclerotic zones. In contrast to liposarcoma, neither lipoblasts nor isolated atypical hyperchromatic cells within mature fat, as are seen in well differentiated liposarcoma, were present. Immunohistochemistry performed in four cases showed factor XIIIa in tumor cells and an intricate pattern of immunoreactivity around cells for type IV collagen. CD34 and smooth-muscle actins were identified in two of four cases. Follow-up in seven cases showed no recurrences or metastases within the follow-up period of 1 to 7 years. Because these lesions are located in deep soft tissue and contain large amounts of mature fat, they could be mistaken for well-differentiated liposarcomas in limited biopsy material, although the distinction is easily made in examining the entire specimen. The lipomatous hemangiopericytoma represents yet another example of a bimodal mesenchymal tumor containing mature fat and raises the question of whether a common cytogenetic abnormality can explain the emergence of two clonal populations in this hybrid tumor. PMID- 10524521 TI - The Spectrum of Cutaneous Lymphomas in HIV infection: a study of 21 cases. AB - We studied 21 HIV-associated lymphomas with cutaneous presentation to determine whether they showed features of primary cutaneous lymphoma arising fortuitously or whether they represented the cutaneous involvement of AIDS systemic lymphoma. Besides rare mycosis fungoides (n = 3), which shared typical clinicopathologic lesions, nonepidermotropic large-cell lymphomas (n = 18) were predominant. They frequently presented as a solitary nodule or tumor. Seven of the eight large T cell lymphomas had a CD30-positive (CD30+) phenotype but did not express ALK protein. Overexpression of p53 protein was observed in six cases. Although EBV EBER transcripts were detected in two of them, LMP1 protein was absent. Except for their original prevalence, the features of these T-cell CD30+ cutaneous lymphomas were the same as in immunocompetent patients. The 10 B-cell cutaneous lymphoma were immunoblastic or centroblastic lymphomas, with a differential expression of BCL-6 and Syndecan. Four of them expressed CD30, EBER-EBV transcripts, and LMP1 and p53 proteins. This B-cell CD30+ EBV+ phenotype contrasts with cutaneous lymphoma in immunocompetent patients. Human herpesvirus 8 was not involved in lymphomagenesis since its sequences were detected in a single patient with Kaposi's sarcoma and Castleman's disease. These lymphomas occurred in severely immunocompromised patients with a low CD4 count. Death was due to immunodepression rather than to lymphoma spread, suggesting avoiding aggressive immunosuppressive treatment in such patients. PMID- 10524522 TI - Electron microscopic diagnosis of human flavivirus encephalitis: use of confocal microscopy as an aid. AB - The distinction between intracranial viral infections and inflammatory conditions requiring immunosuppression is important. Although specific laboratory reagents are readily available for some viruses, diagnosis of arbovirus infection is more difficult. Transmission electron microscopy (TEM) theoretically allows identification of viral particles independent of reagent availability, but it has limited sensitivity. We report two cases of human flavivirus encephalitis diagnosed by TEM. Laser scanning confocal microscopy (LSCM) was used in one case to survey unembedded tissue slices for focal abnormalities, from which fragments smaller than 1 mm2 were excised for epoxy embedding. This facilitated TEM identification of intracytoplasmic, budding, 35-40 nm spherical virus particles, confirmed by serology as St. Louis encephalitis. In contrast to mosquitoes and newborn mice, in which high viral loads are associated with minimal tissue responses, these biopsies showed florid angiodestructive inflammation and microgliosis, with rare virions in necrotic perivascular cells and astrocytes. To our knowledge, this represents the first ultrastructural study of St. Louis encephalitis in humans, indicating the potential value of LSCM-aided TEM. PMID- 10524523 TI - Hyalinizing spindle cell tumor with giant rosettes: a report of three cases with ultrastructural analysis. AB - We report the light microscopic, ultrastructural appearance and immunohistochemical staining profile of three distinctive soft-tissue tumors recently designated hyalinizing spindle cell tumor with giant rosettes. The tumors occurred in two men, 41 and 54 years old, and one woman, 62 years old. Two tumors arose in the lower extremities and one in the upper arm. Two tumors were resected and measured 3 and 13.2 cm in greatest diameter; a biopsy only was done of the third tumor. Grossly, the tumors had a tan, pink, or white cut surface. The largest tumor exhibited central cystic change. Microscopically, they all displayed similar features and were composed of fibromyxoid regions, with areas of hyalinization in two tumors and focal ossification in one tumor. Scattered throughout each of the tumors were rosette-like structures in which neoplastic cells were arranged around a central collagenous core. Ultrastructurally, the neoplastic cells demonstrated the features of fibroblasts. In all tumors, there was abundant extracellular collagen fibers and in one there were large aggregates of amorphous extracellular external lamina-like material. The center of the rosette-like structures was composed of banded collagen fibers and the cells at the periphery of the rosettes had ultrastructural features similar to the neoplastic spindle cells located elsewhere in the tumor. Immunohistochemically, the tumor cells stained for vimentin. There was focal staining of the widely distributed spindle cells and cells that formed the rosettes for Leu-7, S-100 protein, and CD34. In one tumor, there was faint diffuse staining of the spindle cells for neuron-specific enolase. One tumor (with the amorphous extracellular material) stained for type IV collagen. There was no staining for desmin, muscle actin, smooth muscle actin, keratin, or epithelial membrane antigen. These results demonstrate that hyalinizing spindle cell tumor with giant rosettes is composed of fibroblasts. We did not demonstrate any ultrastructural or immunohistochemical differences between the spindle cells that comprised the majority of the mass and those that surrounded the rosette-like structures. There was no ultrastructural evidence of neural differentiation to explain the focal S 100 protein and Leu-7 staining of the tumor cells. PMID- 10524524 TI - Peripheral T-cell lymphoma with Reed-Sternberg-like cells of B-cell phenotype and genotype associated with Epstein-Barr virus infection. AB - We report three cases of nodal peripheral T-cell lymphoma (PTCL) with Reed Sternberg-like (RS-like) cells of B-cell pheno- and/or genotype. Histologic analysis in all cases revealed diffuse nodal effacement by atypical lymphoid cells of variable size. Two of the three cases had features of angioimmunoblastic T-cell lymphoma (AILT). Large mononuclear and binucleated cells with prominent eosinophilic nucleoli and abundant cytoplasm resembling classic RS cells and mononuclear variants were scattered throughout all biopsies. The lymphoma cells in the three cases were of T-cell lineage (CD3+, CD43+, and CD45RO+). The RS-like cells from all cases were CD30 and CD15 positive. In contrast to the neoplastic T cells, the RS-like cells lacked all T-cell markers and in two cases were positive for CD20. Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) and EBER 1 (2/2) were detected in the RS-like cells in all cases. The neoplastic T cells were negative for EBV. Polymerase chain reaction (PCR) analysis demonstrated clonal rearrangements of the T-cell receptor gamma chain gene in the three cases. PCR analysis of microdissected RS-like cells for immunoglobulin heavy chain gene rearrangements in cases 1 and 3 showed an oligoclonal pattern. The presence of RS like cells in PTCL represents a diagnostic pitfall, because in one case this observation led to a misdiagnosis of Hodgkin's disease (HD). The oligoclonal expansion of EBV-infected cells may be related to underlying immunodeficiency associated with T-cell lymphomas and AILT in particular. This phenomenon may provide the basis for some cases of Hodgkin's disease after T-cell lymphomas and suggests that they are clonally unrelated neoplasms. The expression of LMP1 appears to be crucial for the immunophenotype and probably for the morphology of the RS and RS-like cells appearing in diverse lymphoid malignancies, including HD, chronic lymphocytic leukemia, and PTCL. PMID- 10524525 TI - Erosive injury to the upper gastrointestinal tract in patients receiving iron medication: an underrecognized entity. AB - Severe gastrointestinal necrosis and strictures after an iron overdose are well described. However, mucosal injury in patients receiving therapeutic iron has received only scant recognition despite its wide use. We studied the clinical and histologic features of 36 upper gastrointestinal tract biopsies from 33 patients (24 gastric, 9 esophageal, 1 gastroesophageal junction, and 2 duodenal) containing characteristic brown crystalline iron material, and evaluated the amount and tissue distribution of the iron. In addition, we investigated the prevalence of iron-associated mucosal injury in upper gastrointestinal endoscopic examinations. The majority of the biopsies (32 of 36, 89%) contained luminal crystalline iron adjacent to the surface epithelium or admixed with luminal fibrinoinflammatory exudate. Thirty biopsies (83%) showed crystalline iron deposition in the lamina propria, either covered by an intact epithelium, subjacent to small superficial erosions, or admixed with granulation tissue. Three biopsies (8%) demonstrated iron-containing thrombi in mucosal blood vessels. Erosive or ulcerative mucosal injury was present in 30 of 36 biopsies (83%). The amount of iron accumulation in cases with mucosal injury was greater than in cases without mucosal injury (mean grades, 2.4+ vs. 1.3+ on a 1+ to 3+ scale; p = 0.002). Iron medication was confirmed in 25 of 33 patients (76%) 22 patients were receiving ferrous sulfate. Approximately half of the patients (17 of 33, 51%) also had underlying infectious, mechanical, toxic, or systemic medical conditions that could have initiated or exacerbated tissue injury. Crystalline iron deposition was found in 0.9% of upper gastrointestinal endoscopic examinations (12 of 1,300), and iron medication-associated erosive mucosal injury was present in 0.7% (9 of 1,300). These results indicate that crystalline iron deposition in the upper gastrointestinal tract is not uncommon. It can induce or exacerbate a distinctive histologic pattern of erosive mucosal injury, especially in patients with associated upper gastrointestinal disorders. Recognition of this pattern by pathologists and its communication to clinicians may aid in optimizing therapy. PMID- 10524527 TI - Aberrant crypt foci in the human colon: frequency and histologic patterns in patients with colorectal cancer or diverticular disease. AB - Aberrant crypt foci are considered potential markers of colorectal cancer risk. The aim of this study was to analyze a large series of human aberrant crypt foci according to frequency, distribution, and histology. Aberrant crypt foci were identified in methylene blue-stained colonic mucosa from 103 patients undergoing surgery for colorectal cancer or diverticular disease. Foci were histologically classified into surface hyperplastic type, surface and glandular hyperplastic type, mixed hyperplastic and adenomatous type, and adenomatous type. The mean frequency of aberrant crypt foci (n = 720) was higher in the colorectal cancer group (0.20/cm2) than in the diverticular disease group (0.07/cm2), and in distal colonic segments than in proximal segments. Most of the histologically examined foci (n = 366) were hyperplastic (88.8%). Surface hyperplasia accounted for 30.6% and prevailed in small lesions. Surface and glandular hyperplasia accounted for 58.2% and prevailed in medium-sized to large foci. Partially or totally dysplastic foci accounted for 10.1% of examined lesions (10.8% and 2.8% in the colorectal cancer and diverticular disease groups, respectively). Most of them (94.6%) were composed of mixed hyperplastic and adenomatous crypts and prevailed in large lesions. The higher frequency of aberrant crypt foci in patients with colorectal cancer sustains their putative role as preneoplastic markers. The high rate of mixed hyperplastic and adenomatous lesions supports the possible adenomatous transformation of hyperplastic lesions. PMID- 10524526 TI - Immunohistochemistry for hMLH1 and hMSH2: a practical test for DNA mismatch repair-deficient tumors. AB - Inactivation of deoxyribonucleic acid (DNA) mismatch repair genes, most commonly human mutL homologue 1 (hMLH1) or human mutS homologue 2 (hMSH2), is a recently described alternate pathway in cancer development and progression. The resulting genetic instability is characterized by widespread somatic mutations in tumor DNA, and is termed high-frequency microsatellite instability (MSI-H). Although described in a variety of tumors, mismatch repair deficiency has been studied predominantly in colorectal carcinoma. Most MSI-H colorectal carcinomas are sporadic, but some occur in patients with hereditary nonpolyposis colorectal cancer (HNPCC), and are associated with germline mutations in mismatch repair genes. Until now, the identification of MSI-H cancers has required molecular testing. To evaluate the role of immunohistochemistry as a new screening tool for mismatch repair-deficient neoplasms, the authors studied the expression of hMLH1 and hMSH2, using commercially available monoclonal antibodies, in 72 formalin fixed, paraffin-embedded tumors that had been tested previously for microsatellite instability. They compared immunohistochemical patterns of 38 MSI H neoplasms, including 16 cases from HNPCC patients with known germline mutations in hMLH1 or hMSH2, with 34 neoplasms that did not show microsatellite instability. Thirty-seven of 38 MSI-H neoplasms were predicted to have a mismatch repair gene defect, as demonstrated by the absence of hMLH1 and/or hMSH2 expression. This included correspondence with all 16 cases with germline mutations. All 34 microsatellite-stable cancers had intact staining with both antibodies. These findings clearly demonstrate that immunohistochemistry can discriminate accurately between MSI-H and microsatellite-stable tumors, providing a practical new technique with important clinical and research applications. PMID- 10524528 TI - Benign mesothelial cells in mediastinal lymph nodes. AB - Inclusions of benign tissues in lymph nodes are most often aberrant glandular tissue, including endosalpingiosis, the thyroid, parotid, breast, and pancreas. Nonglandular inclusions are rare and include nevus cells and decidua. Mesothelial cells in lymph nodes are exceedingly rare; only eight cases have been reported in mediastinal lymph nodes and three cases in abdominal lymph nodes. The incidence of benign mesothelial cells in mediastinal lymph nodes in patients with a history of pericarditis or pleuritis is reported in this study. A retrospective search showed eight cases with removal of mediastinal lymph nodes in the absence of neoplasm. Hematoxylin and eosin-stained sections were examined in all cases. Immunohistochemical stains for CAM 5.2 were performed in all cases, and stains for AE1/AE3, Ber-EP4, carcinoembryonic antigen, Leu-M1, B72.3, and S-100 were performed in one case. CAM 5.2-positive cells with features of mesothelial cells were present in five of eight cases. In all cases, the cells were present in nodal sinuses and appeared as single cells or small clusters. The cells were missed on routine hematoxylin and eosin sections in all cases but one, in which they were numerous and mimicked metastatic carcinoma. Malignancy was not found in any of the cases preoperatively, at the time of surgery, or during the follow-up period. Benign mesothelial cells may embolize to regional lymph nodes in pleuritis or pericarditis. In most cases, these cells are few and undetectable on routine sections. Rarely, hyperplastic mesothelial cells may be present and must be distinguished from metastatic carcinoma, mesothelioma, and melanoma. PMID- 10524530 TI - Cysticercosis in Nepal: a histopathologic study of sixty-two cases. AB - Human cysticercosis, an infection caused by larvae of Taenia solium, is a major public health problem in many developing countries. Sixty-two of 23,402 biopsy cases have been detected as cysticercosis in the last 5 years in Patan Hospital. Most (82%) of the patients presented with solitary skin nodules, another 10% with nodules in the oral mucosa, and 8% in the breast. Forty cases were identified from the Kathmandu valley and the rest from outside Kathmandu. Most patients were younger than 30 years of age (mean, 21+/-11 years). Statistically, there was no difference between males (0.28%) and females (0.24%). The average size of cysticercosis was 19 mm in diameter, and the histology of cysticercosis showed fibrous walled cysts covered by several layered epithelioid cells with a few Langhans' giant cells and infiltration of eosinophils without caseous necrosis. These cysticercosis findings from an endemic area will be helpful for doctors who examine immigrant patients in nonendemic areas. PMID- 10524529 TI - Fibro-osseous lesions of the central nervous system: report of four cases and literature review. AB - Fibro-osseous lesions, also reported as calcifying pseudoneoplasms of the neural axis, are uncommon lesions of the CNS. We report four additional cases: two extraaxial and two intraaxial, in patients ages 33, 47, 49, and 59 years at presentation. Fibro-osseous lesions involving the CNS demonstrate variable proportions of fibrous stroma, bone, palisading spindle to epithelioid to multinucleated cells in association with a highly distinctive, perhaps pathognomonic, chondromyxoid-like matrix often distributed in a nodular pattern. This histopathologically distinctive lesion can be seen in many regions of the neuraxis, often with a dural association, and most commonly along the vertebral column. It appears to be a slow-growing lesion and, with wide excision, the prognosis is excellent. The etiology remains unclear, but the preponderance of data favors a reactive rather than neoplastic process. If this putative pseudotumor is not recognized histopathologically, a neoplastic or infectious differential might result in inappropriate investigations and potentially harmful therapies. PMID- 10524531 TI - Breast carcinoma diverging to aberrant melanocytic differentiation: a case report with histopathologic and loss of heterozygosity analyses. AB - A case of primary breast cancer showing differentiation to malignant melanoma is reported. To obtain insight into the clonal relationship between the two components of the tumor, polymerase chain reaction-based microsatellite analysis to detect loss of heterozygosity on chromosome arms 1p, 1q, 3q, 4q, 6q, 8p, 9p, 10q, 11q, 13q, 16q, 17p, 17q, and 18q with microdissected tissues of both components was performed in addition to histologic, histochemical, immunohistochemical, and ultrastructural techniques. The tumor consisted of a combination of carcinoma and melanoma with morphologic transition. Metastases in the lymph nodes and thoracic spinal bone marrow showed dual tissue structure. One of the metastatic lung tumors showed melanomatous tissue structure. The abundant pigment in the cells was positive for Fontana-Masson staining and bleached with potassium permanganate. The carcinoma component was positive for epithelial membrane antigen and CA19-9, but the melanoma component was negative. Conversely, the melanoma component was positive for HMB45 and vimentin, but the carcinoma component was negative. Electron microscopic analysis showed premelanosomes and melanosomes in the melanoma component. Microsatellite analysis showed the same genetic alterations with loss of heterozygosity on chromosome arms 1p, 3q, 4q, 6q, 9p, 10q, 11q, 13q, 16q, 17p, and 17q in in situ, invasive, and metastatic foci. We concluded that the carcinoma and melanoma components had arisen from the same clone and that this breast carcinoma might have diverged to aberrant malignant melanoma through multiple genetic alterations in the early period of ductal carcinoma in situ. PMID- 10524532 TI - Regional proliferation of HMB-45-positive clear cells of the lung with lymphangioleiomyomatosislike distribution, replacing the lobes with multiple cysts and a nodule. AB - The authors report a case of a localized lesion of the lung presenting as multiple cysts and as a tumor in the right upper and middle lobes, consisting of a diffuse proliferation of clear cells with intralysosomal glycogen granules and human melanin black (HMB)-45 immunoreactivity. A 33-year-old woman complained of dyspnea because of the enlargement of bullae in the right upper and middle lung fields without stigmata of tuberous sclerosis. Resection showed multiple, various size air-filled cysts and a tumor. The cysts in the resected lungs were reminiscent of lymphangioleiomyomatosis (LAM), accompanied by the diffuse proliferation of clear cells in the interstitium. The tumor, 1.8 cm in diameter, resembled a clear cell tumor of the lung (CCTL) and showed proliferation of clear cells with sinusoidlike vascular spaces. Both forms of proliferation were continuous spatially, and both constituent cells showed diffuse HMB-45 immunoreactivity. The cells that comprised a nodule revealed ultrastructurally abundant cytoplasmic glycogen, which was in the form of free and membrane-bound glycogen granules. This case may represent a particular pulmonary lesion consisting of CCTL-LAM hybrid cells, which share the cytologic features with CCTL cells on one hand, and the proliferative pattern and potential with LAM cells on the other. PMID- 10524533 TI - Mycobacterial spindle cell pseudotumor of the brain: a case report and review of the literature. AB - Spindle cell pseudotumors found in the skin, lymph nodes, bone marrow, spleen, lungs, and retroperitoneum have been reported recently in immunosuppressed patients, including those with acquired immunodeficiency syndrome. The authors report a similar lesion limited to the brain in a 38-year-old human immunodeficiency virus-negative man receiving steroid therapy for treatment of sarcoidosis. Histopathologically the lesions were composed of spindle and epithelioid histiocytes, small foci of necrosis, and numerous acid-fast bacilli. The acid-fast bacilli were determined by culture and polymerase chain reaction to be Mycobacterium avium intracellulare. Because of the uncommon histologic appearance of this lesion and the potential for treatment if recognized, mycobacterial spindle cell pseudotumors should be included in the differential diagnosis of spindle cell lesions in the brain in immunosuppressed patients. PMID- 10524534 TI - Unique cytological features and chromosome aberrations in chondroid lipoma: a case report based on fine-needle aspiration cytology, histopathology, electron microscopy, chromosome banding, and molecular cytogenetics. AB - Chondroid lipoma is a rare, benign tumor that may mimic soft-tissue sarcoma clinically. Its histopathologic features may resemble hibernoma, myxoid liposarcoma, myxoid chondrosarcoma, and other lipomatous or chondroid neoplasms. In this study, a chondroid lipoma was analyzed by fine-needle aspiration cytology, histopathology, electron microscopy, chromosome banding, and metaphase fluorescence in situ hybridization. The results demonstrate that chondroid lipoma exhibits a characteristic pattern by fine-needle aspiration cytology, including a mixture of benign adipose tissue with lipoblastlike cells, and chondroblastlike cells with a fibrochondroid matrix. Cytogenetically, a three-way rearrangement between chromosomes 1, 2, and 5 was found, together with an 11;16 translocation with a breakpoint in 11q13, approximately 1 Mb proximal to the MEN1 region shown to be rearranged frequently in hibernoma. The presence of a karyotype of low complexity, but without any of the genetic aberrations characteristic for other types of soft-tissue tumors, indicate that chondroid lipoma develops along a unique pathogenetic pathway. PMID- 10524535 TI - Peritumoral and nodal muciphages. PMID- 10524536 TI - Cervical adenoid basal epitheliomas. PMID- 10524537 TI - Recurrent wheezing in infants and young children: a perspective. PMID- 10524538 TI - The effect of corticosteroid therapy on blood eosinophils and eosinophilic cationic protein in patients with acute and chronic asthma. AB - There is evidence that eosinophils are involved in inflammation in asthma, a correlation having been observed between blood eosinophil (B-EOS) count and pulmonary function. It has been suggested that eosinophils, and its product, eosinophil cationic protein (ECP), can serve as markers of disease activity. This paper examines this hypothesis. B-EOS count, serum ECP level, and peak expiratory flow (PEF) were estimated in two groups of asthmatics and controls at three visits in 4 weeks. The mean B-EOS count in acute and stable asthmatic groups was higher than in controls at presentation; the difference was statistically significant (p<0.02). Similarly, mean ECP was higher in the two groups than in controls, but with no statistically significant difference. The B-EOS count and serum ECP level within the groups fell between week 0 and week 4 because of treatment. There was positive correlation between ECP and PEF and also between B EOS and ECP and PEF. The findings reveal that blood eosinophils reflect some degree of activity in asthmatic patients in the acute and chronic state. PMID- 10524539 TI - Sensitivity and specificity of asthma definitions and symptoms used in a survey of childhood asthma. AB - We compared the ability of definitions/symptoms of asthma to identify urban, elementary schoolchildren with physician-diagnosed asthma and bronchial hyperresponsiveness (BHR) post-exercise challenge. Definitions of asthma from the literature were compared, including American Thoracic Society (ATS) and British Medical Research Council (BMRC) definitions. Modified ATS had the highest sensitivity (77%), whereas BMRC had the highest specificity (99%). The most sensitive symptom was "wheeze with cold" (89%). The most specific symptoms were "medication required," and "breathing normal between attacks" (95%). Definitions and symptoms were poor predictors of BHR. Researchers can use these estimates in selecting and defining specific populations of children with asthma. PMID- 10524540 TI - Asthma hospitalization trends in California, 1983-1996. AB - Annual asthma hospitalization rates were calculated for California's ethnically diverse population from 1983 through 1996. Trends were examined for four race/ethnicity groups: Hispanics, African-Americans, non-Hispanic Caucasians, and Asians/Pacific Islanders. The overall rate decreased by 30% during the time period. African-Americans had the highest rate, more that three times greater than the rate for Caucasians. Among children, the rates for Caucasians decreased by one-third, while rates increased for Hispanics and Asians. The rate for African-American children remained generally constant and was four times higher than the rate for Caucasians. Data from 1996 were assessed for repeat admissions, age and sex differences in rates, costs, and progress toward national goals. PMID- 10524541 TI - Beta2-adrenergic receptor polymorphisms affect airway responsiveness to salbutamol in asthmatics. AB - We examined the beta2-adrenergic receptor (beta2AR) polymorphisms (Arg16-->Gly, Gln27-->Glu) and clinical status for 117 asthmatics. Airway responsiveness to methacholine and beta2-agonists was evaluated with Astograph. The atopic factors, pulmonary function test, and airway responsiveness to methacholine did not differ significantly among the different beta2AR genotypes. Asthmatics homozygous for Gly16 showed significantly lower airway responsiveness to inhaled salbutamol than those heterozygous for Arg/Gly16 or homozygous for Arg16. Asthmatics heterozygous for Gln/Glu27 had significantly later asthma onsets than those homozygous for Gln27. These results suggest that beta2AR polymorphisms play an important role in the airway responsiveness to inhaled beta2-agonist and the initial asthma onset. PMID- 10524542 TI - Asthma controller medications: what do patients want? AB - Our goal was to understand which features of asthma controller medications are important to patients. We used a cross-sectional survey of primary care patients (N = 394) with the diagnosis of asthma. Using conjoint or "trade-off analysis," we measured patient preferences for hypothetical asthma controller medications based upon their route and frequency of administration, and need for blood test monitoring. Patients were not willing to use medications that required blood test monitoring. Preference regarding blood test monitoring was the strongest of any medication attribute that we studied, accounting for 45% of the variation. Patients' decisions were also highly affected by the frequency of dosing (40% of the variation). Patients did not have strong preferences regarding the route of administration (15% of the variation). Understanding these patient preferences may lead to increased compliance with treatment plans and promote physician patient partnership. PMID- 10524543 TI - A tool to organize instructions at discharge after treatment of asthmatic children in an emergency department. AB - Asthma exacerbations continue to be a major cause of visits to emergency departments (ED). Comprehensive care in the outpatient setting, with planning for early intervention for exacerbations, can reduce emergency visits. Thus, a major goal of ED intervention is to establish a link between the patient and the provider of ongoing asthma care, where complete education can be achieved and reinforced over time. When designing the Asthma 1-2-3 Plan discharge teaching tool for the ED, consideration was given to educational format, readability, patient population, and setting in which education was to be delivered. To evaluate use of the plan, ED records of patients enrolled in a separate asthma study, the Neighborhood Asthma Coalition (NAC), were audited for two 8-month intervals, May-December 1993 (before initiation of the plan) and May December 1994 (starting 1 month after completion of pilot testing on the plan in the ED). To evaluate effectiveness of the plan, records of physicians who cared for children in the NAC were evaluated. The database was reviewed for the date of the first visit for planned review of asthma that occurred after the acute asthma ED visit. After introduction of the plan, the proportion of children told to return to the physician for follow-up increased from 54% to 81%. The proportion of children given advice to return to their physician within the recommended 3 days or less increased from 11% to 54%. Chi2 Analyses showed that these changes were both statistically significant (p<0.0001). The plan was not effective in achieving increased follow-up visits for regular asthma care, in that 7% returned for follow-up within 7 days after an ED visit before the plan and only 6% returned for such a visit after the Plan. Successful initiation of a focused discharge teaching tool into the routine of the ED increased appropriate advice given at time of discharge from the ED. Although unsuccessful in increasing appropriate follow-up, the present intervention uses the ED not as a base for asthma education, but as a point for contacting patients in need of regular care and education, and for promoting access to that regular care. PMID- 10524545 TI - Predictors of asthma severity in the elderly: results of a community survey in Northeast England. AB - A number of risk factors for the development and severity of asthma in childhood are known. Particularly, there is information on allergens, excessive use of beta2- agonists, and indoor environmental pollutants. Similar information on elderly patients is lacking. We examined the risk factors for current asthma and for the severity of asthma in 95 elderly subjects (>65 years old) compared to 274 elderly subjects with obstructive spirometry who did not have asthma as defined by the following criteria: symptoms of episodic wheeze, cough, or chest tightness and forced expiratory volume in 1 sec/vital capacity (FEV1/VC) <70% with >15% or 200 mL reversibility in FEV1 to 200 microg salbutamol given from a metered-dose inhaler. The severity of airflow limitation was graded on the basis of the FEV1/VC ratio as mild (60%-70%), moderate (40%-60%), and severe (<40%). Asthma history was collected using the Medical Research Council respiratory questionnaire and a follow-up postal questionnaire. Data were analyzed using multiple logistic regression and the overall goodness-of-fit of the model was checked using the Hosmer-Lemeshow (HL) statistic. History of allergy (to one or more of the following allergens: cat, house dust, or grass or tree pollen) (odds ratio [OR] 25; 95% confidence interval [CI] 13-51; p = 0.0001) and history of childhood wheeze (OR 8; 95% CI 4-9; p = 0.004) were strong predictors of current asthma. Duration of wheezing, smoking history, indoor heating, history of working in coal mines, and sex were not predictors (HL 6.75, degrees of freedom [df] = 8, p = 0.56). Use of >4 puffs of salbutamol/ day (OR 5.3; 95% CI 2-14; p = 0.005), more than 10 years of asthma symptoms (OR 4.2; 95% CI 4.1-36.2; p = 0.0001), and >500 mL reversibility in FEV1 (OR 4.2; 95% CI 1.2-14.3; p = 0.05) were independent predictors of moderate to severe asthma. History of atopy was the strongest predictor of asthma in the elderly population studied. Indoor heating, presence of pets at home, sex, smoking history, and history of working in coal mines were not predictors of asthma. The severity of asthma as assessed by measurement of airflow limitation was related to the frequency of use of beta2 agonists, duration of symptoms of asthma, and increased reversibility of FEV1 to beta2-agonist. PMID- 10524544 TI - Community study using a polymerase chain reaction panel to determine the prevalence of common respiratory viruses in asthmatic and nonasthmatic children. AB - We developed a sensitive polymerase chain reaction (PCR) panel, suitable for the detection of seven common respiratory viruses, to study the prevalence of viruses in nasal swabs obtained from clinically stable asthmatic children (n = 21), non physician diagnosed asthmatic children with exercise-induced bronchoconstriction (EIB) (n = 16), and nonasthmatic, non-EIB controls (n = 33). The PCR panel detected viruses in 43/70 (61.4%) specimens but there were no significant differences in prevalence of these viruses between the three groups of children. These results indicate that clinically stable asthmatic and nonasthmatic children frequently harbor viruses in the upper respiratory tract. PMID- 10524546 TI - Effect of beta-agonists on production of cytokines by activated T cells obtained from asthmatic patients and normal subjects. AB - Intracellular levels of cAMP were found to regulate T cell activity. We examined whether beta2-agonists altered cytokine production and cyclic adenosine monophosphate (cAMP) accumulation in concanavalin A (ConA)-activated peripheral T cells from asthmatic patients. Procaterol and isoproterenol weakly decreased the ConA-elicited interleukin (IL)-4 and IL-5 secretion; however, the inhibitory effect of procaterol on the ConA-induced IL-2 secretion was inferior to that of isoproterenol in normal controls and was little in asthmatics. The intracellular accumulation of cAMP by procaterol was not altered compared with that by isoproterenol. Results suggest that there is a qualitative difference between procaterol- and isoproterenol-induced cAMP accumulation in T cells. PMID- 10524547 TI - Evidence of the radiographic diagnosis of osteonecrosis disputed. PMID- 10524548 TI - Investigation of the alveolar macrophages and T lymphocytes in 15 patients with systemic sclerosis. AB - The cell distribution and function of alveolar macrophages and T lymphocytes were investigated in the bronchoalveolar lavage (BAL) of 15 patients with systemic sclerosis (SSc). In alveolar macrophages, both spontaneous and PMA-stimulated TNF alpha production were increased in SSc. PMA-induced IL-6 production was also elevated. Spontaneous IL-6 excretion of scleroderma alveolar macrophages was similar to the controls. Yeast and C3b-coated yeast (opsonised yeast) phagocytosis, chemotaxis and Fc receptor activity of alveolar macrophages were normal. The proportion of CD3, CD4 and CD8 T-lymphocyte subsets in the BAL was similar to the control values. The lymphocyte blast transformation index of the non-adherent cells deriving from the BAL fluid was markedly decreased. PMID- 10524549 TI - Influence of disease activity and chronicity on ankylosing spondylitis bone mass loss. AB - We investigated 30 consecutive Brazilian patients with definite ankylosing spondylitis (AS) fulfilling the New York and the European spondyloarthropathy study group classification criteria. The mean age at study was 37 years old and the mean disease duration was 17 years. Bone densitometry employed the dual energy X-ray absorptiometry (DEXA) technique, using a Hologic QDR-1000/W densitometer. Axial bone mineral density (BMD) was measured in the lumbar spine (L1-L4) and appendicular BMD was measured in the total proximal femur and sub regions (neck, greater trochanter, intertrochanter and Ward's triangle). Based on World Health Organisation criteria, the lumbar spine showed osteopenia or osteoporosis in 50% of the patients, while 86% had osteopenia or osteoporosis in the total proximal femur. When compared with the normal population, the patients showed a significant BMD decrease in the lumbar spine and total proximal femur with sub-regions, except for the femoral neck. A comparison of BMD between patients with active and inactive disease did not reveal a significant effect of clinical disease activity on the lumbar spine and total proximal femur with sub regions, except for Ward's triangle. Concerning disease chronicity, there were significant positive correlations between disease duration and lumbar spine, total proximal femur, greater trochanter and intertrochanteric regional BMD. This false increase in lumbar spine BMD found mostly in patients with long standing AS was due to the presence of paravertebral calcification and ossification. We conclude that the bone mass loss in AS is better evaluated in the proximal femur, because of the greater sensitivity of bone densitometry in this region, which is almost free of artefacts. PMID- 10524551 TI - The influence of a joint orthosis on the grip force of the rheumatoid hand. AB - Distribution of force was studied for the distal, medial and proximal digits of 60 patients with rheumatoid arthritis during a cylindrical grip before correction of the flattened transverse arch of the hand and after its correction using a felt pad placed under the capitulum of the third metacarpal bone, and also before and after the placing of a wrist band. The cylindrical grip involving 22 degrees of ease of movement is the best indicator of hand function and deteriorates most (in approximately 80-90%) in cases of disease. PMID- 10524552 TI - Urinary thrombomodulin in patients with rheumatoid arthritis: relationship to disease subset. AB - In 110 patients with rheumatoid arthritis (RA), the mean (+/- SD) urinary thrombomodulin (TM) concentration was 74.4+/-19.5 ng/mg creatinine (Cre), which was significantly higher than the mean in age-matched healthy controls (49.9+/ 10.8 ng/mg Cre; p<0.0001). The mean urinary TM concentration in the RA subset with least erosive disease (LES) was 65.2+/-12.4 ng/mg Cre (n = 41), with more erosive disease (MES) was 77.4+/-20.4 ng/mg Cre (n = 58) and with mutilating disease (MUD) was 92.6+/-20.2 ng/mg Cre (n = 11). TM in the MUD group was the highest of the three subsets (ANOVA, p<0.0001). By contrast, the erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) in the MES and MUD groups were not significantly different. Urinary TM levels may allow differentiation of RA subsets, unlike markers of inflammation such as ESR and CRP. PMID- 10524550 TI - Chloroquine therapy in patients with recent-onset rheumatoid arthritis: the clinical response can be predicted by the low level of acute-phase reaction at baseline. AB - If rheumatoid arthritis (RA) patients with a mild disease course could be identified early in the phase of the disease, therapy with less aggressive and probably less toxic antirheumatic drugs seems to be rational. The aim of this study was to investigate which factors at baseline could predict a clinical response (American College of Rheumatology preliminary response criteria) after treatment with chloroquine for 16 weeks. Two hundred and three early RA patients with active disease were treated with oral chloroquine sulphate (Nivaquine) at a daily dose of 300 mg during the first 4 weeks, 200 mg during the second 4 weeks and 100 mg thereafter. One hundred and eighty-three patients (90%) completed the study and 20 patients prematurely discontinued treatment. Of all the patients, 43 patients (21%) met the response criteria. A low level of C-reactive protein (CRP) was the only independent predictor for clinical response [relative risk: 0.97 (95% confidence interval: 0.95-0.98)]. It was concluded that a clinical response to chloroquine therapy in early RA patients can be predicted by a low CRP level at baseline. PMID- 10524553 TI - Calcium pyrophosphate dihydrate crystal deposition disease of the knee simulating spontaneous osteonecrosis. AB - A retrospective review of the frontal and lateral knee radiographs of 200 patients with calcium pyrophosphate dihydrate (CPPD) crystal deposition disease was performed. Of these 200 patients, nine patients (four male, five female, mean age 74 years, age range 63-87 years) had radiographic findings simulating osteonecrosis of the knee. One patient also had magnetic resonance imaging of the involved knee. A total of 10 knee radiographs in nine patients showed articular and periarticular calcification diagnostic of CPPD crystal deposition of the knee. In addition, all 10 radiographs showed flattening of one femoral condyle. Four of the 10 cases demonstrated an area of radiolucency in the subchondral bone surrounded by a halo of sclerosis. Eight of the 10 cases had narrowing of the involved joint compartment and osteophytosis. These findings mimic the radiographic signs of spontaneous osteonecrosis of the knee. In conclusion, flattening of the femoral condyles in CPPD crystal deposition disease simulates that of spontaneous osteonecrosis and probably relates to articular cartilage and meniscal damage that subsequently leads to stress fracture of subchondral bone with bone collapse. PMID- 10524555 TI - Validation study of a Hebrew version of WOMAC in patients with osteoarthritis of the knee. AB - The aim of the study was to validate a translated version of the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) in Hebrew speaking populations. The WOMAC was translated into Hebrew from its English version and its reliability and validity were studied. Before its use in patients, the Hebrew version was translated back into English by an independent translator and minor amendments were made to satisfy the original English versions designed by Bellamy et al. The Hebrew version of the WOMAC questionnaire was administered to 114 patients with osteoarthritis of the knee. All the subjects were asked about the presence and severity of pain during movement and handicap, using a visual analogue scale (VAS) of pain and handicap. Test-retest reliability was assessed using Pearson's and intraclass correlation coefficients. Internal consistency was evaluated by Cronbach's alpha coefficient of reliability. Construct validity was tested by correlating the WOMAC items with VAS of pain and handicap. The test retest reliability Pearson's correlation coefficients for the WOMAC items ranged from 0.55 to 0.78 (p<0.01), and the Cronbach's alpha was 0.97 at time 1 and 0.98 at time 2. Significant correlations (p<0.01) were obtained between the WOMAC items and VAS of pain and handicap. The Hebrew version of WOMAC is a reliable and valid instrument for evaluating the severity of osteoarthritis of the knee in Israeli patients. PMID- 10524554 TI - Decreased CD4+ lymphocyte activation and increased interleukin-4 production in peripheral blood of rheumatoid arthritis patients after acute starvation. AB - We investigated the effects of acute starvation on mitogen-induced T-cell activation and Th1/Th2 cytokine responses in rheumatoid arthritis (RA) patients. Ten RA patients with active disease underwent a 7-day fast followed by a 2-week refeeding period. Immunological, hormonal, laboratory and clinical evaluations were carried out on days 0, 7 and 21. Using flow cytometry, mitogen-stimulated T cell activation was assessed in fresh heparinised blood via analysis of CD69 expression. Production of Th1 (interferon-gamma) and Th2 (interleukin-4, IL-4) cytokines was also assessed by ELISA. The 7-day fast significantly decreased the erythrocyte sedimentation rate, C-reactive protein level, joint count, morning stiffness, body weight, CD4+ and CD8+ counts and CD69+ expression on mitogen stimulated CD4+ lymphocytes. A significant increase in mitogen-induced IL-4 production after fasting was found. The fast markedly reduced serum leptin and insulin-like growth factor-1 concentrations. No significant differences occurred in serum cortisol or prolactin before and after fasting. Decreases in CD4+ lymphocyte activation during fasting correlated with decreases in body weight. Our results suggest that the clinical and laboratory improvements in fasting RA patients may be attributed to decreased CD4+ T-cell activation and an increase in the number and/or function of IL-4-producing Th2 cells. Factors associated with loss of body weight during acute starvation appear to have an inhibitory effect on CD4+ lymphocyte activation. PMID- 10524556 TI - Candida glabrata arthritis: case report and review of the literature of Candida arthritis. AB - We report a case of arthritis due to Candida (Torulopsis) glabrata in two different joints at different times in the same patient. The first episode of arthritis was situated in the right ankle and lasted more than 1 year before the patient agreed to the proposed treatment. Therapy with intravenous amphotericin B and oral fluconazole failed. A cure was achieved with weekly intra-articular administration of amphotericin B, which was continued for more than 20 weeks and combined with oral itraconazole. Several weeks later the patient developed Candida glabrata arthritis of the left knee while still taking itraconazole. Immediately, intravenous amphotericin B therapy was started and was successful. Because there were no previous invasive point manipulations or trauma, the infections were considered to be haematogenously disseminated. Chronic corticosteroid and repeated antibiotic therapy for infectious exacerbations of chronic obstructive pulmonary disease and alcohol abuse are the presumed risk factors in this otherwise immunocompetent patient. PMID- 10524557 TI - Low-dose cyclosporin for multiple colonic ulcers associated with mixed connective tissue disease. AB - This report describes a patient with multiple colonic ulcers and mixed connective tissue disease. The histological findings of the colonic lesions showed vasculitis with T-cell infiltration, and the peripheral T cells were frequently in the activated phase of the cell cycle. In this patient, low-dose cyclosporin treatment (2.5 mg/kg/day) inhibited the T-cell activation in the peripheral lymphocytes and was very effective in the gastrointestinal disorder, which might be related to T-cell activation. This case suggests the possibility that even low dose cyclosporin can exert a great influence on peripheral T cells and directly inhibit T-cell activation, thereby improving symptoms related to T-cell activation. PMID- 10524558 TI - A complex case of hepatitis in a patient with systemic lupus erythematosus. AB - Liver involvement in patients with systemic lupus erythematosus (SLE) is considered rare. Previous treatment with potentially hepatotoxic drugs or viral hepatitis have usually been implicated as the main causes of liver disease in SLE patients. On the other hand, even after careful exclusion of these aetiologies, the problem remains whether to classify the patient as having a primary liver disease with associated autoimmune clinical and laboratory features resembling SLE, such as autoimmune hepatitis, or as having liver disease as a manifestation of SLE. We report the case of an elderly woman who presented with acute hepatitis, who had been diagnosed with SLE 14 years ago and who also had Sjogren's syndrome and anti-phospholipid's syndrome for several years. The histology depicted chronic active hepatitis and, after drug-induced hepatitis and viral hepatitis were excluded, the serological and clinical features were shown to be typical of liver damage caused by SLE. The patient was treated with azathioprine 100 mg/d and prednisone 30 mg/d. The clinical symptoms resolved in 10 days and the laboratory values were normal at the end of the first month of therapy. Prednisone was progressively reduced, during a period of 4 months, to 10 mg/d but azathioprine was kept to the same dose. One year after the diagnoses the patient is still in remission. Although uncommon, hepatic involvement is well recognised in SLE. The interest of this case lies in the differential diagnosis and recognition of this condition, which deserves an aggressive treatment. PMID- 10524559 TI - Spondylodiscitis caused by viridans streptococci: three cases and a review of the literature. AB - Three cases of spondylodiscitis caused by viridans streptococci were observed within the course of 1 month. Although streptococci have been reported as the third most frequent cause of spondylodiscitis after staphylococci and gram negative bacteria, alpha-haemolytic streptococci are rarely seen. The three patients presented with symptoms of low back pain; they felt well and did not have a fever or chills. Laboratory examinations revealed inflammation. Further examinations such as scintigraphy, computed tomography or magnetic resonance imaging were done. Bacteriological diagnosis was established by blood cultures in two cases and by needle biopsy of the disco-vertebral space in one. In one patient endocarditis was also documented. Because the prevalence of endocarditis was found to be higher in our cases of spondylodiscitis due to Streptococcus viridans than for other bacteria, the exclusion of this diagnosis must be pursued aggressively. These observations lead us to question if the spectrum of bacteria causing spondylodiscitis is undergoing a change. an aetiological agent could be isolated in 1168 patients (85.4%): in 48% a staphylococcus, in 28% a gram negative bacterium and in only 10% a streptococcus. There were two cases of viridans streptococci (0.2%). These two cases together with other single case reports [14-22] account for 15 cases of spondylodiscitis due to alpha-haemolytic streptococci. Differentiation of the organisms to the species level was accomplished in six cases: S. mitis (3), S. sanguis (2) and S. anginosus (1). Although a multitude of organisms, bacterial as well as fungal, causing spondylodiscitis has been reported in recent years, almost all were single cases [23-42]. The unusual observation of three cases of spondylodiscitis due to alpha haemolytic streptococci within 1 month prompted us to review the clinical and laboratory findings and to compare these cases with those caused by Staphylococcus aureus. PMID- 10524560 TI - Bleomycin-induced scleroderma: report of a case with a chronic course rather than the typical acute/subacute self-limiting form. AB - We report a case of bleomycin-induced scleroderma in a 35-year-old woman treated with chemotherapy for Hodgkin's disease. Approximately 6 months after the first chemotherapy cycle, the patient developed skin sclerosis in both arms. The lesion showed no signs of spontaneous clinical amelioration and treatment with steroids was unsuccessful. A partial remission of the skin sclerosis was instead obtained by the administration of D-penicillamine. A family history revealed other cases of autoimmune diseases and HLA typing showed the presence of antigens associated with scleroderma. The association between bleomycin therapy and scleroderma is discussed. PMID- 10524561 TI - Immunohistological study endothelin-1 and endothelin-A and B receptors in two patients with scleroderma renal crisis. AB - Scleroderma renal crisis (SRC) developed in two patients with systemic sclerosis (SSc) and they died from respiratory failure. Findings on autopsy revealed congestion and oedema in both lungs and intimal thickening of the small renal arteries in both patients. Immunohistological investigations showed positive staining of anti-human endothelin (ET)-1 in the media of the small renal arteries and ET-B receptor in the medial smooth muscle of the small renal arteries. This observation suggests an important pathophysiological role of ET-1 in the development of SRC in some patients with SSc. PMID- 10524562 TI - Comparison of recombinant human thyrotropin receptors versus porcine thyrotropin receptors in the thyrotropin binding inhibition assay for thyrotropin receptor autoantibodies. AB - Thyrotropin receptor autoantibodies (TRAb) are most commonly measured in a thyrotropin-binding inhibition (TBI) assay using solubilized porcine thyrotropin receptors (pTSHR). Recently, we reported modifications in recombinant human thyrotropin receptor (hTSHR) production and extraction that made substitution of this antigen for the pTSHR practical. We now report the first comparison of the behavior in a TBI assay of the recombinant, solubilized hTSHR with the pTSHR in a large series of clinically characterized patients with autoimmune thyroid disease. We studied 227 patients with Graves' disease (32 untreated patients, 156 patients receiving antithyroid medications, 24 patients in remission, 9 patients with recurrence of disease, and 6 thyroidectomized patients), as well as 32 patients with Hashimoto's thyroiditis and 28 normal individuals. In patients with untreated Graves' disease, 29 of 32 (90.6%) were TBI positive with either antigen, although two sera gave discrepant data in the two assay. Of the patients receiving antithyroid drugs, 94 of 156 (60.3%) were positive with the pTSHR and 106 of 156 (67.9%) were positive with the hTSHR TBI assay (p < 0.05%). In all other respects, however, there was no difference between the two TBI assays. Neither assay performed well in providing clinical guidance in the remission or relapse of disease. Of the 24 Graves' patients in remission, 75.0% and 79.2% were TBI negative with the hTSHR and pTSHR assays, respectively. The TBI assay at the time of relapse was even less informative; 6 of 9 (66.7%) being TBI negative in the pTSHR assay and 3/9 (33.3%) being negative in the hTSHR assay. In TBI assays with both species of TSHR, 3 of 32 hypothyroid patients with Hashimoto's thyroiditis were TBI positive. In summary, production of the recombinant hTSHR is now a practical reality and this antigen can clearly substitute at least as well for the pTSHR in the imperfect, although most commonly used, TBI assay. It is, therefore, likely that the hTSHR will supplant the pTSHR in this important assay. However, the use of the hTSHR rather than pTSHR does not appear to provide a major advantage, at least in terms of TBI assay sensitivity, specificity and predictive value. PMID- 10524563 TI - Hypothalamic-pituitary-testicular axis and seminal parameters in hyperthyroid males. AB - Information on the effect of abnormal thyroid function on male reproduction is less available than that for the female. To assess the effects of hyperthyroidism on hypothalamic-pituitary-testicular axis and on spermogram parameters, 25 male patients (19-47 years old) suffering from active Graves' disease were studied. Serum luteinizing hormone (LH), follicle stimulating hormone (FSH), and prolactin (PRL) were measured before and after administration of 100 microg GnRH plus 200 microg thyrotropin-releasing hormone (TRH). Testosterone (T), estradiol (E2), and 17-hydroxyprogesterone (17-OHP) were determined before and after 5000 IU human chorionic gonadotropin (HCG) administration. Serum sex hormone-binding globulin (SHBG), cortisol-binding globulin (CBG), androstenedione and bioavailable testosterone (bioT), and bioavailable estradiol (bioE2) were also measured. Spermograms according to World Health Organization (WHO) criteria were determined in 21 patients. Hormonal and seminal studies were repeated in six patients after 7 to 19 months of euthyroidism achieved after treatment for hyperthyroidism. As a control group, 10 normal men were evaluated. Impaired sexual function, gynecomastia, and low testicular volume were found in 12, 6, and 3 hyperthyroid patients. Mean basal LH was significantly higher than the control group (7.8 +/- 4.7 vs. 5.0 +/- 1.9 mIU/mL, respectively, p < 0.02), with hyperresponse to GnRH. The response of PRL to TRH was lower in patients versus control group (30 minutes: 3.9 +/- 3.4 and 12.0 +/- 2.8 ng/mL, p < 0.01). Basal levels of steroids and SHBG were significantly higher in patients than in normal men (T: 9.3 +/- 3.3 vs. 5.4 +/- 1.6 ng/mL, p < 0.005; E2: 62.2 +/- 25.2 vs. 32.1 +/- 11.0 pg/mL, p < 0.005; 17-OHP: 2.4 +/- 0.9 vs. 1.1 +/- 0.5 ng/mL, p < 0.001; SHBG: 102.3 +/- 37.3 vs. 19.0 +/- 5.0 nmol/L, p < 0.01). The maximal increment of T and 17-OHP after HCG was lower in hyperthyroid patients than in normal men (p < 0.019 and p < 0.001, respectively). Basal bioT was lower in patients than controls (1.7 +/- 0.8 and 3.1 +/- 1.9 ng/mL, p < 0.02). The following incidence of abnormal semen parameters was found: asthenospermia 85.7%, hypospermia 61.9%, oligospermia 42.9%, necrospermia 42.9% and teratospermia 19.0%. In euthyroidism, a normalization of 85% of seminal alterations was observed in the limited number of patients evaluated. Our results confirm that hyperthyroidism causes marked alterations of the gonadotropic and PRL axis and dramatically affects spermatic function. BioT measurement was useful to identify hypoandrogenism in these patients in spite of the high concentration of total testosterone. The restoration of most semen parameters when euthyroidism was achieved suggests that the alterations were induced by the Graves' disease. PMID- 10524564 TI - Calculation of the radioiodine dose for the treatment of Graves' hyperthyroidism: is more than seven-thousand rad target dose necessary? AB - Some authors recently suggested a significant increase in the target dose of radioiodine treatment in Graves' disease. The aim of the present study was to investigate the impact of thyroid gland mass on the success rate of radioiodine treatment. For this purpose, the thyroid function of 105 consecutive Graves' patients was assessed 6 and 12 months after a 131I treatment and correlated to the gland mass. The patients were categorized according to the gland mass into small (< or = 30 g; 19 patients), medium size (31-50 g; 40 patients), and large size (> 50 g; 46 patients) groups (S, M, L groups, respectively). None of the patients received more than a 10,000-rad (100-Gy) target dose. During the calculation of administered 131I activity, late uptake measurement has also been routinely used, in addition to the usual maximal uptake parameter. The established effective half-life of 131I was highly variable (5 +/- 1.2 days; range: 2-7.6 days) and could not be predicted based on other clinical data without measuring an extended radioiodine uptake curve of the given patient. However, the correlation between the administered activity calculated from the complete set of uptake values and that of only a single late one was excellent (r = 0.99). Six months after the 131I treatment, hyperthyroidism was cured in 81% of patients with small and medium size thyroid glands, with 62% euthyroid and 19% hypothyroid ratios respectively. In the early phase of study for large goiters, the same linear mass activity function was used during calculation as in smaller glands. In these 17 patients the nonhyperthyroid result was comparable to the results of treatment of the small and medium size gland groups only after 1 year (77%), but the 6-month success rate was significantly lower (53%; p < 0.05). After obtaining these results, the usual 7000-rad target dose was increased to 8000-10,000 rad (depending on the gland mass) in another group of 29 patients with large thyroid glands that result in an acceptable 6-month success rate of 72%. In conclusion, instead of the "mCi 131I/g gland mass/maximal uptake" dose calculation, we suggest a method in which (1) the late 131I uptake measurement is taken into account and (2) for large goiters there is an additional dose adjustment, ie, increase is needed over the usual linear, size driven calculation. No overall increase of target dose over 10,000 rad is necessary if no antithyroid medication is given shortly before 131I treatment. PMID- 10524565 TI - Special features of Graves' disease in early childhood. AB - Graves' disease (GD) is extremely rare in children younger than 4 years of age, but if not recognized and treated it can seriously interfere with growth and development. We report three unrelated children, all females, in whom GD occurred before the age of 3. These children presented with goiter, exophthalmos, tachycardia, and hyperactivity. Moreover, one showed a severe psychomotor delay, and had previously undergone surgery due to craniosynostosis; the other two manifested a language delay. All had high thyroid hormones and thyrotropin receptor antibody (TRAb) serum levels that clearly indicated autoimmune hyperthyroidism. In all of them, the disease presumably had developed during the first or second year of life. No maternal history of GD was present in two. The third child was born to a mother affected with GD during pregnancy, but it is likely that her GD began to develop after 6 months of life. These children are being treated with methimazole, and treatment is still necessary after 32 months. TRAb levels were persistently high at follow-up. Psychological evaluation including language development at follow-up was appropriate for age in two children; the third child improved, but severe mental retardation is still evident. GD assessment in early childhood also needs to focus on psychological evaluation. Pediatricians should be aware of the possibility of permanent brain damage and craniosynostosis due to hyperthyroidism in infancy. PMID- 10524566 TI - Selective binding of thyrotropin receptor autoantibodies to recombinant extracellular domain of thyrotropin/lutropin-chorionic gonadotropin receptor chimeric proteins. AB - The extracellular domain of the glycosylated human thyrotropin receptor (ET-gp) contains epitopes that can adsorb pathogenic antibodies from sera of patients with Graves' disease (GD). In an attempt to define the regions within the ETSHR with which autoantibodies interact, we expressed extracellular domains of eight thyrotropin receptor/chorionic gonadotropin receptor (TSHR/LH-CGR) chimeric proteins in insect cells. The levels of expression were high and chimeric proteins were glycosylated. Chimeric proteins designated as EMc2+4 and EMc2+3+4, in which amino acids (aa) 90-165 and 261-370, and aa 90-370, respectively, of TSHR were replaced with corresponding aa of LH-CGR, partially reversed the thyrotropin binding inhibitory immunoglobulin (TBII) activity of experimental anti-TSHR antisera (anti-ET-gp). The other six chimeras almost completely reversed the TBII activity of these anti-ET-GP antisera. Next, we tested the ability of these chimeric proteins to reverse the TBII activity of GD patients' sera. Similar to our earlier study, ET-gp protein reversed the TBII activity of all eight GD patients' sera tested. Chimera EMc2, in which aa 90-165 of TSHR has been replaced with corresponding aa of LH-CGR, and EMc2+4 partially reversed the TBII activity of only three of the eight GD patients' sera. However, the other six chimeric proteins failed to neutralize the TBII activity of any of GD patients' sera. These data showed the following: (1) There is considerable heterogeneity amongst autoantibodies in GD patients' sera, (2) The TBII activity of some, but not others, is dependent on aa 90-165 and 261-370, and (3) Most Graves' sera, with TBII activity, failed to react with chimeric proteins in which either N-terminal or C-terminal regions of the extra cellular domain of the TSHR were replaced with corresponding regions of LH-CGR. These results suggest that the TBII activity of GD patients' sera is dependent on conformational epitopes and replacement of certain regions of TSHR with homologous regions of LH-CGR results in sufficient alteration in the conformation of the protein leading to loss of reactivity. PMID- 10524567 TI - The hypothyroidism in an inbred kindred with congenital thyroid hormone and glucocorticoid deficiency is due to a mutation producing a truncated thyrotropin receptor. AB - Growth and function of the thyroid and adrenal glands are maintained and controlled by thyrotropin (TSH) and adrenocorticotrophic hormone (ACTH), respectively. The action of these trophic hormones requires the presence of functional TSH and ACTH receptors. We describe a large inbred Bedouin kindred in which profound congenital hypothyroidism and hypoadrenocortisolism occurred alone or together in eight family members belonging to four nuclear families. The high serum TSH and ACTH levels in the presence of normal or hypoplastic thyroid glands and low glucocorticoid, but not mineralocorticoid concentrations, are characteristic of resistance to TSH and ACTH. Linkage analysis, using specific polymorphic markers, excluded the involvement of the ACTH receptor but not thyrotropin receptor (TSHR). A novel point mutation was identified in exon 10 of the TSHR that replaces the normal cytosine in nucleotide 2024 with a thymidine. As a result the normal arginine in codon 609 (CGA) is replaced with a stop codon (TGA). This mutation produces a truncated TSHR lacking the third intracellular and extracellular loops, the sixth and seventh transmembrane segments, and the intracytoplasmic tail. The presence of hypothyroidism did not affect the timing, severity, and manner of clinical manifestation of hypoadrenocortisolism. PMID- 10524569 TI - Genomic organization of the 3' region of the human thyroglobulin gene. AB - The genomic organization of the 3' end of the human Thyroglobulin (Tg) gene has not previously been characterized. We isolated and characterized seventeen lambda phage clones from a human genomic library that included nucleotides 6263 to 8410 of the Tg mRNA, encompassing the last thirteen 3' exons of the Tg gene. The region contained exons ranging in size from 94 to 222 nucleotides, split by introns of 1 to 64 kb. We estimate a total of 48 exons in the Tg gene. All the intron-exon boundaries were sequenced. We found that the splicing sequences diverged considerably from the 3' and 5' consensus. However, the GT-AG rule was perfectly respected in all the exons. A total of 5788 intronic bases and most of the sequences contained in the 13 exons were analyzed (1846 bases). One sequence variation, TT to CC at positions 8377-8378, was found in the 3' untranslated segment. The three tyrosine residues involved in thyroid hormones synthesis (amino acids 2554, 2568, and 2747) at the carbosyl termini of Tg, are encoded by exons 44, 45, and 48. The knowledge of the precise organization of the Tg gene should help to direct studies of Tg gene mutations in families in which a defect in the synthesis of Tg occurs. PMID- 10524570 TI - Degradation of extracellular matrix by metastatic follicular thyroid carcinoma cell lines: role of the plasmin activation system. AB - The plasmin activation system plays a key role in extracellular matrix degradation in many malignant tumors. Because no data are available on the involvement of the plasmin activation system in matrix degradation by thyroid carcinoma, the present study was performed using follicular thyroid carcinoma cell lines obtained from a primary tumor (FTC-133) and metastases (FTC-236 and FTC-238) of one patient. Matrix degradation by these cell lines was studied assessing the release of radioactivity from S35-methionine labeled extracellular matrix coated onto plastic. The involvement of constituents of the plasmin activation system as well as matrix metalloproteinases (MMPs), another class of proteolytic enzymes, which can be activated by plasmin, were assessed by semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and zymography. In the matrix degradation experiment, S35 release by FTC-133 was significantly higher than FTC-236 and FTC-238. S35 degradation could be inhibited by the plasmin inhibitor aprotinin and by anti-human urokinase-type plasminogen activator (uPA) antibody, indicating the involvement of the plasmin activation system. Matrix degradation could also be inhibited by the MMP inhibitor marimastat, thus demonstrating the involvement of MMPs in matrix degradation by these cell lines. Zymographic assays revealed activity of uPA in all cell lines. However, in contrast with FTC-236 and FTC-238, no plasminogen activator inhibitor (PAI) or PAI1 mRNA were found in FTC-133. Therefore, the differences in PAI activity as observed between the cell lines may originate from differences in PAI1 gene transcription. Differences in PAI1 expression did not affect the attachment of these cell lines to vitronectin. We conclude that the plasmin activation system is involved in extracellular matrix degradation by these metastatic follicular thyroid carcinoma cell lines. Differences in extracellular matrix degradation between the cell lines correspond with differences in PAI1 gene expression, indicating the significance of PAI1 in extracellular matrix degradation by metastatic follicular thyroid carcinoma. PMID- 10524568 TI - Assessment of goiter in an area of endemic iodine deficiency. AB - Urinary iodone (UI) excretion and sonographically measured thyroid volume were investigated in 195 subjects living in 6 separate villages in the Casamance region of southeastern Senegal, West Africa. A comparison of goiter prevalence using thyroid palpation and volume measurement and of iodine excretion expressed as micrograms per gram (microg/g) creatinine or micrograms per deciliter (microg/dl) urine was undertaken, and possible pathogenetic factors were investigated. Ultrasound measured thyroid volumes were above the recommended upper limit of the reference range for an area replete in iodine in 83.1% or females, 52.3% of males, and 80.0% of children aged 13 years or younger. Overall sensitivity and specificity for palpation compared to sonographically demonstrated thyroid enlargement was 51.7% and 91.5%, respectively. Thyroid enlargement was not associated with ethnic origin, thiocyanate ingestion, HLA DR/DQ phenotype frequency, or thyroid growth-stimulating immunoglobulin (TGI) positivity. Median UI was 32 microg/g creatinine with 65.0% having values consistent with iodine deficiency (< 50 microg/g). When results were expressed as micrograms per deciliter, the percentage having values consistent with iodine deficiency (< 5.0 microg/dl) increased to 95.7%. The findings suggest a primary role for iodine deficiency in goitrogenesis in the study population. They demonstrate that classification of the severity of the endemia in this or other study populations in areas of iodine deficiency is dependent on the methods used to determine goiter prevalence (palpation or ultrasound measured thyroid enlargement), or dietary iodine status (iodine excretion expressed as micrograms per gram creatinine or micrograms per deciliter urine). PMID- 10524571 TI - Clinical evaluation of serum tissue polypeptide specific antigen in patients with thyroid carcinoma. AB - There is no specific marker for diagnosis of thyroid cancer with the exception of carcinoembryonic antigen (CEA) and calcitonin for medullary thyroid carcinoma. Tissue polypeptide antigen (TPA) is known to be a diagnostic marker of malignant neoplasms. TPA is also one of the tumor markers for thyroid cancer, but serum TPA shows poor specificity because of polyclonal antibody. Tissue polypeptide specific antigen (TPS) is detected by the monoclonal antibody M3, directed at 1 of the 35 epitopes in TPA. TPS appears in proliferative tumor cells. We examined 72 patients (32 thyroid carcinoma, 20 thyroid adenoma, 5 adenomatous goiter, and 15 diffuse goiter) and 24 healthy volunteers for serum TPS, TPA, and thyroglobulin (Tg). The TPS in thyroid carcinoma was significantly higher than in healthy controls and thyroid adenoma (p < 0.05). The mean value of TPS for all diseases except thyroid carcinoma was below the cutoff value (95 U/L). The TPS sensitivity, specificity, and accuracy for thyroid carcinoma was 37.5%, 100%, and 64.9% respectively. There were no significant differences in mean values of serum Tg and TPA between thyroid carcinomas and adenomas. Positive rates of tissue TPS expression in thyroid carcinomas and adenomas were 87.5% (7/8) and 14.3% (1/7), respectively. The positive rates for serum TPS and immunoreactive TPS in the tissue correlated significantly (p < 0.0001). It is concluded that TPS is a useful marker for diagnosis of thyroid carcinoma. PMID- 10524572 TI - Three-dimensional structure of the micro-blood vessels in thyroid tumors analyzed by immunohistochemistry coupled with image analysis. AB - The structure of micro-blood vessels, one of the most important factors influencing the tumor growth and tumor metastasis among histological types of thyroid malignancy, was analyzed immunochemically by staining tissues for platelet endothelial cell adhesion molecule-1 (PECAM-1). Human thyroid tumor tissue obtained at surgery, consisting of 18 cases of papillary carcinoma, 9 cases of follicular carcinoma, and 9 cases of anaplastic carcinoma were fixed in formalin solution, and paraffin sections were made. They were stained for PECAM-1 using the avidin-biotin complex (ABC) technique. The volume of the blood vessels and their three-dimensional (3D) structure were analyzed using an image analyzer. The volume ratios of blood vessels in thyroid tissues were: normal tissues, 1.10%; papillary carcinoma, 3.01%; follicular carcinoma, 8.13%; and anaplastic carcinoma, 0.91%. Ratios in malignant tumors were larger than in normal tissues, except for anaplastic carcinoma. The typical 3D structure of micro-blood vessels was histopathologically varied: branching tree-like blood vessels in papillary carcinomas; vessels of varied diameter surrounding follicle structure in follicular carcinomas; and simple and immature vessels in anaplastic carcinomas. The volume and 3D structure of micro-blood vessels in thyroid malignant tumors differed from those in normal tissues, and varied according to histological classification. PMID- 10524573 TI - Comparison of two thyroglobulin immunoradiometric assays on the basis of comprehensive imaging in differentiated thyroid carcinoma. AB - The aim was to compare two thyroglobulin-immunoradiometric assays (Tg-IRMA) in the follow-up of patients with differentiated thyroid carcinoma (DTC) in order to set up interassay correlation, correlation to clinical background, and to determine whether a lower functional sensitivity (kit A: 0.5 ng/mL, kit B: 0.3 ng/mL) would allow an earlier detection of recurrences. Three hundred eight samples from 181 patients with DTC were investigated. The clinical interpretation of the Tg-IRMA results was based on comprehensive imaging and the clinical history before and during the study period. Groups were formed against this background and against the thyrotropin (TSH) levels of the samples (LT4- on and LT4-off). During a follow-up period that lasted until September 1998, the clinical situation was reevaluated in order to determine any changes in the patients' clinical status. The two assays presented a good interassay correlation of 0.838. Both assays had a high and comparably good sensitivity in the detection of recurrence of malignancy or distant metastases. Patients in remission had, in most cases, nonmeasurable or Tg values below 1 ng/mL. Kit B presented slightly measurable Tg results in a larger number of patients in remission; however, during the follow-up most of these slightly measurable Tg results were not reproducible, thus being most likely artifacts. Consequently, the functional sensitivity of 0.3 ng/mL of kit B showed no advantages in terms of an earlier tumor detection and seems to be unacceptably low. Negative consequences may be an increase in the number of investigations during the follow-up, which may be disconcerting for both the clinicians and the patients. PMID- 10524574 TI - Basal calcitonin levels and the response to pentagastrin stimulation in patients after kidney transplantation or on chronic hemodialysis as indicators of medullary carcinoma. AB - Plasma concentrations of calcitonin (hCT) were determined in 150 patients with chronic renal failure on chronic hemodialysis therapy (CHD) and in 800 patients after successful kidney transplantation (KT). Basal hCT concentrations exceeded 10 pg/mL in 44 of 150 patients (29%) with CHD and in 48 of 800 (6%) in patients with KT. Among these patients with elevated basal hCT, pentagastrin-stimulated concentrations of hCT exceeded 100 pg/mL in 4 patients with CHD and in 7 with KT. Thyroidectomy was performed in 8 patients (5 with KT, 3 with CHD) revealing the presence of medullary thyroid carcinoma (MTC) (n = 2) or of C-cell hyperplasia (n = 6). Two patients with C-cell hyperplasia had the neoplastic form of this disorder. One patient with MTC and 1 with C-cell hyperplasia also presented a papillary microcarcinoma. Stimulated concentrations of hCT were only moderately elevated in the remaining 3 patients and follow-up rather than surgery was deemed appropriate due to their concomitant severe medical problems. In conclusion, basal concentrations of hCT higher than 10 pg/mL are more common in patients with CHD (29%) and after successful KT (6%) than previously described in patients with thyroid nodular disease (3%). In spite of various additional factors complicating the interpretation of elevated hCT in CHD, pentagastrin-stimulated values above 100 pg/mL must be considered to indicate the presence of C-cell hyperplasia and/or of medullary thyroid carcinoma. Although thyroidectomy would theoretically be the therapy of choice, the potential benefit of the operation must be seen in the context of the patient's general condition. PMID- 10524575 TI - Monomorphic teratoma of the ovary: a rare cause of triiodothyronine toxicosis. AB - A case of low thyroid radioactive iodine uptake (RAIU) thyrotoxicosis due to a large struma ovarii comprising pure thyroid tissue is presented, including a detailed diagnostic evaluation, histopathology, and demonstration of rapid recovery of native thyroid function after surgical excision. In addition, the first comprehensive analysis of thyroglobulin obtained from an ovarian struma is reported. PMID- 10524576 TI - Recurrent pregnancy-related upper airway obstruction caused by intratracheal ectopic thyroid tissue. AB - An unusual case of recurrent pregnancy-related thyroid growth stimulation is reported. A 27-year-old euthyroid woman had pulmonary symptoms, thought to be asthma during her first pregnancy, that improved postpartum. Bronchodilatators had no effect and symptoms recurred from gestational week 22 during her second pregnancy. Her 58-mL multinodular goiter (by ultrasound) was not thought to be responsible for her upper airway symptoms. Therefore, fiber laryngoscopy and computed tomographic (CT) scan were performed and revealed a 20 x 15 x 10 mm intratracheal tumor. After tracheostomy and microlaryngoscopy, benign goitrous thyroid tissue was removed through a tracheal fissure during gestational week 35. Postoperatively the patient had stopped medication and was without any pulmonary symptoms. The child was delivered by cesarean section in gestational week 39. Apgar score was normal and the child has developed normally. We believe that this case illustrates the recurrent effect of pregnancy-related thyroid tissue stimulation by a combination of increasing human chorionic gonadotropin (hCG) stimulation and iodine deficiency in a borderline iodine-deficient region. This is the first report on symptomatic intratracheal ectopic thyroid tissue diagnosed during pregnancy. PMID- 10524578 TI - Satellite communication and medical assistance for thyroid disease diagnosis from Nagasaki to Chernobyl. PMID- 10524577 TI - Responsiveness to thyroid hormone is enhanced in rat hepatocytes cultured as spheroids compared with that in monolayers: altered responsiveness to thyroid hormone possibly involves complex formed on thyroid hormone response elements. AB - We previously reported that the expression of type I iodothyronine 5'-deiodinase (5'DI) gene was increased by 3,3,',5-triiodothyronine (T3) in isolated rat hepatocytes when cultured as spherical aggregates (spheroids), whereas this effect was greatly attenuated in conventional monolayer cultures. In the current study, we examined whether the enhanced T3 responsiveness in spheroid cultures extends to other T3-responsive genes. As observed for 5'DI, T3 increased spot 14, malic enzyme and fibronectin messenger RNAs (mRNAs) by fourfold to fivefold in spheroid cultures, while the effect in monolayer cultures was blunted. This difference in T3 responsiveness was also observed when T3-responsive reporters consisting of the luciferase gene under the control of triiodothyronine response element (TRE) were introduced into hepatocytes using a replication-defective adenovirus vector. These results suggest that the factors required for T3 dependent transcriptional activation are preserved in spheroid cultures and that they must exert their effect by interacting with TRE. Maximal binding capacity of nuclear T3 receptor was not different between monolayer and spheroid cultures while the expression of retinoid X receptor-alpha (RXR alpha) mRNA was higher in spheroid cultures compared with that in monolayers. The difference in RXR alpha mRNA expression, together with enhanced proteolytic cleavage in monolayers that we demonstrated recently, may account for the difference in T3 responsiveness between the two hepatocyte culture systems. PMID- 10524579 TI - How much isoniazid is needed for prevention of tuberculosis among immunocompetent adults? AB - Recommendations regarding how long isoniazid should be taken to prevent tuberculosis have gradually shifted from 12 to 6 months of treatment. These recommendations and the relevant evidence from controlled trials on duration of treatment are reviewed. The conclusions for immunocompetent adults are: 1) 6 months of preventive treatment does not give optimal protection; 2) more than 12 months of preventive treatment is not necessary; 3) 9-10 months appears to be the optimal duration; and 4) total duration of preventive treatment may be more important than its continuity. PMID- 10524580 TI - Recurrent tuberculosis: definitions and treatment regimens. AB - Within the National Tuberculosis Control Programme of Malawi, misunderstandings sometimes occur about the diagnosis and management of recurrent tuberculosis (TB). Patients with smear-positive pulmonary tuberculosis (PTB) who have had a previous, treated episode of smear-negative TB may be registered as 'new cases' rather than relapse cases, and thus denied the benefits of a retreatment regimen. Patients with a recurrent episode of smear-negative PTB or extra-pulmonary TB (EPTB) may also be wrongly registered as 'new cases' rather than recurrent cases. International guidelines about the treatment of recurrent smear-negative PTB and EPTB are not explicit, resulting in confusion about how best to manage these cases. It is suggested that all such cases be considered for re-treatment regimen because of concerns about acquired drug resistance. WHO and IUATLD guidelines on the diagnosis and management of recurrent and relapse TB need to be improved, and operational research studies should be conducted to provide answers to some outstanding questions. PMID- 10524581 TI - Shifting the paradigm in tuberculosis control: illustrations from India. AB - Drawing on literature from India and key contributions from social science, this paper asks and attempts to answer the question 'who is to blame for treatment failures in TB'? Some key lessons emerge: effective tuberculosis control cannot be achieved so long as the disease is considered in isolation from the social processes that maintain it, create the conditions facilitating its spread and act as barriers to care. Insights into the economic and social burdens incurred with a diagnosis of TB are essential to understand why many patients, especially the most disadvantaged, are unable to comply with treatment regimens. TB and health care interventions need to be appropriate to the health service contexts in which they are applied, and sensitive to the competing demands, needs and priorities of people's lives. The paper argues for the need to reorient TB control programmes towards enabling patients to obtain care. The problem of access emerges as central to people's ability to obtain and maintain appropriate therapy. Examples and characteristics of successful non-governmental projects, from which policy makers, programmers and practitioners could learn, are outlined and contrasted with more rigid directly observed treatment approaches. We conclude that treatment failures are not patient failures, and that TB control programmes need to address the social dimensions of TB, and adhere to the principles of good TB care, with the same commitment that is devoted to ensuring patients follow treatment guidelines. We suggest a paradigm shift away from a focus on diseased patients towards enabling health in the community. PMID- 10524583 TI - Socio-economic impact of tuberculosis on patients and family in India. AB - OBJECTIVE: To quantify the socio-economic impact of tuberculosis on patients and their families from the costs incurred by patients in rural and urban areas. DESIGN: An interview schedule prepared from 17 focus group discussions was used to collect socio-economic demographic characteristics, employment, income particulars, expenditure on illness and effects on children from newly detected sputum-positive pulmonary tuberculosis patients. The direct and indirect costs included money spent on diagnosis, drugs, investigations, travel and loss of wages. Total costs were projected for the entire 6 months of treatment. RESULTS: The study population consisted of 304 patients (government health care 202, non governmental organisation 77, private practitioner 25), 120 of whom were females. Mean direct cost was Rs.2052/-, indirect Rs.3934/-, and total cost was Rs.5986/- ($171 US). The mean number of work days lost was 83 and mean debts totalled Rs.2079/-. Both rural and urban female patients faced rejection by their families (15%). Eleven per cent of schoolchildren discontinued their studies; an additional 8% took up employment to support their family. CONCLUSIONS: The total costs, and particularly indirect costs due to TB, were relatively high. The average period of loss of wages was 3 months. Care giving activities of female patients decreased significantly, and a fifth of schoolchildren discontinued their studies. PMID- 10524582 TI - Attitudes to compliance with tuberculosis treatment among women and men in Vietnam. AB - SETTING: A study carried out in 1996 in four districts representing south and north as well as urban and rural areas of Vietnam. OBJECTIVE: To explore gender differences in knowledge, beliefs and attitudes towards tuberculosis and its treatment, and how these factors influence patients' compliance with treatment. DESIGN: Sixteen focus group discussions were performed by a multi-disciplinary research team from Vietnam and Sweden. Analysis was performed using modified Grounded Theory technique, specifically evaluating gender differences. RESULTS: Women were believed to be more compliant than men. Insufficient knowledge and individual cost during treatment were reported as main obstacles to compliance among men (poor patient compliance), while sensitivity to interaction with health staff and stigma in society (poor health staff and system compliance) were reported as the main obstacles among women. CONCLUSIONS: It is time to adopt a more comprehensive and gender-sensitive approach to compliance, which incorporates patient compliance, doctor compliance and system compliance, in order to fully support individual patients in their efforts to comply with treatment. PMID- 10524584 TI - Developing nursing practice as part of the collaborative TB control programme, Tomsk, Siberia. AB - SETTING: Review of nursing practice, identification of training needs and implementation of training for nurses working in the Tomsk Oblast' Tuberculosis Services (TOTBS), Russia. OBJECTIVE: Preparation of TOTBS nurses for the implementation of a WHO-style TB control programme in January 1997. METHODS: Nursing services and training needs were assessed through observation visits to a number of institutions providing care for TB patients, semi-structured interviews, and discussions at staff meetings. Training sessions focused on the WHO DOTS strategy, patient education and default tracing. An evaluation visit focused on nurses' attitudes and levels of treatment completion. RESULTS: Out of a total of 165 TB cases notified in Tomsk Oblast' between January and March 1997, 53 were started on DOTS on an ambulatory basis. Five patients who defaulted returned to treatment within five days (range 2-5) and no patients were lost to follow up. Improved compliance was attributed to better patient education offered by nurses and a reliable supply of medication. Quarterly reports continue to show satisfactory levels of treatment completion. CONCLUSIONS: Obstacles to the development of nursing practice included resistance to change and low morale due to enormous workloads, no pay and staff shortages. Motivation improved through the setting of achievable targets. PMID- 10524585 TI - DOTS (Directly Observed Treatment Strategy) project in Mongolia, 1995. AB - OBJECTIVE: To establish a tuberculosis (TB) control programme consistent with recommendations made by the WHO and the International Union Against Tuberculosis and Lung Disease (IUATLD) in a country where, as in control programmes of the former USSR, TB management previously relied on active case-finding with radiology and long-term monitoring and treatment of patients. DESIGN AND METHODS: A pilot DOTS strategy (directly observed treatment, short course) project was implemented in Dornod Aimak, Eastern Mongolia During a 6-week period, individuals with chronic cough of > or =3 weeks were screened with sputum smear microscopy. Smear-positive patients received a supervised 6-month regimen (2SRHZ/4RH). Outcome was assessed with smear examination 2, 5, and 6 months after the initiation of treatment. RESULTS: Screening of 1241 symptomatic individuals identified 169 smear-positive TB cases (14%). Most of them (92%) were cured as demonstrated by documented sputum conversion. Five patients completed treatment, but were not available for follow-up smear examination, four patients died and four defaulted. CONCLUSION: The DOTS strategy was successfully introduced in a former socialist model country, paving the way to national DOTS implementation in Mongolia. It may serve as an example for countries with a health care tradition similar to that of the Commonwealth of Independent States. PMID- 10524586 TI - The effect of bacille Calmette-Guerin vaccination at birth on tuberculin skin test reactivity in Ugandan children. AB - SETTING: In Uganda, bacille-Calmette Guerin (BCG) vaccination coverage at birth is between 82 and 84%. OBJECTIVE: To evaluate the effect of neonatal BCG vaccination on tuberculin skin test positivity in Ugandan children exposed to infectious cases. DESIGN: As part of an ongoing prevalence study of household contacts of new tuberculosis cases, 365 children were evaluated to determine if BCG vaccination at birth had an impact on tuberculin skin testing. The children were classified as contacts (179) and non-contacts (186) depending on the presence of a sputum acid-fast bacilli (AFB) smear-positive adult tuberculosis case in the household. RESULTS: Regardless of prior BCG vaccination, children exposed to a smear-positive adult were more likely to have a positive skin test (purified protein derivative >5mm) (68% versus 36%, P < 0.01). BCG-vaccinated children below 1 year of age without a known household contact with active tuberculosis had a lower frequency of tuberculin skin reactions (29%) compared to their counterparts in the contact households (65%, P = 0.031). CONCLUSION: BCG vaccination at birth had no important effect on the interpretation of the tuberculin skin test reactivity in this group of Ugandan children. The tuberculin skin test remains a valuable tool for the evaluation of household contacts and suspected cases of tuberculosis in BCG-vaccinated children. PMID- 10524587 TI - The diagnosis of smear-negative pulmonary tuberculosis: the practice of sputum smear examination in Malawi. AB - SETTING: Forty hospitals in Malawi (3 central, 22 district and 15 mission) performing smear microscopy and registering tuberculosis patients. OBJECTIVE: To determine, in patients aged 15 years or above, 1) the proportion with smear negative pulmonary tuberculosis (PTB) who had sputum smears examined, 2) the number of sputum smears examined per patient, and 3) the proportion of patients registered with smear-positive and smear-negative PTB. DESIGN: Data collection during three 6-month periods, from January 1997 to June 1998, using tuberculosis registers, laboratory sputum registers and quarterly reports. RESULTS: Of 6301 smear-negative PTB patients, 84% had sputum smears examined, the rate increasing from 76% in January-June 1997, to 85% in July-December 1997, to 89% in January June 1998. Of patients who submitted sputum (where the number of smears was recorded), 99% had two or more smears examined and 93% had three smears examined. In district and mission hospitals performance improved over time, while in central hospitals results were more variable. During the same 18-month period 21 422 patients aged 15 years or more were registered with PTB: 59% with smear positive PTB and 41% with smear-negative PTB; this pattern was similar in each 6 month period. CONCLUSION: The study suggests that it is reasonable to aim for a target of 90% or more of smear-negative PTB patients having sputum smears examined. PMID- 10524588 TI - Tuberculosis infection and homelessness in Melbourne, Australia, 1995-1996. AB - OBJECTIVE: To describe tuberculosis infection among persons experiencing homelessness in inner Melbourne, Australia. DESIGN: Homeless people were surveyed during late 1995 and early 1996. In stage one of the study 284 homeless people from crisis and long-term accommodation sites were recruited by means of stratified, systematic, random sampling. In stage two a convenience sample of 100 homeless people from squats and the streets were recruited. Participants completed a questionnaire and Mantoux testing was performed. RESULTS: A past history of tuberculosis was reported by 3%. Thirty-seven per cent had a Mantoux > or =10 mm; 21% > or =15 mm; and 11% > or =20 mm. A Mantoux > or =15 mm was independently associated with being aged > or =40 years, coming from the accommodated sample, overseas birth, and a past history of tuberculosis. Using logistic regression modelling, a Mantoux > or =15 mm was predicted by being aged > or =40 years, overseas birth, and past history of tuberculosis. CONCLUSION: Mantoux test results suggest that this group of homeless people had a high prevalence of infection with the tubercle bacillus. Many aspects of the physical and social circumstances of homeless people predispose to reactivation and have the potential to enhance rapid spread should latent infection become active disease. PMID- 10524589 TI - Bone marrow cultures for the diagnosis of mycobacterial and fungal infections in patients infected with the human immunodeficiency virus. AB - SETTING: University medical center. OBJECTIVE: To determine the value of bone marrow cultures for mycobacteria and fungi in patients infected with the human immunodeficiency virus (HIV). DESIGN: Retrospective review of charts and laboratory records. RESULTS: From 1992-1996, 1225 bone marrow specimens were submitted for mycobacterial and fungal cultures. The number of specimens submitted,declined sharply from 435 in 1992 to 94 in 1996 (P = 0.002 for trend). The yield remained stable. Thirty-one of 1225 specimens grew mycobacteria or fungi; 26 isolates were from 24 HIV-infected patients. These 24 patients were infected with Mycobacterium avium complex (19), M. tuberculosis (one), M. chelonae (one), Histoplasma capsulatum (two), and Cryptococcus neoformans (one). All 24 HIV-infected patients had a culture submitted from at least one other site within 4 weeks of the positive bone marrow culture. The identical organism was grown from another site (usually blood) in 18 of these 24 patients. The bone marrow culture provided the only positive result in six patients and the first positive result in eight patients. CONCLUSIONS: Utilization of bone marrow cultures for mycobacteria and fungi declined at our institution. Bone marrow and blood cultures were highly concordant. However, the majority of positive bone marrow cultures provided useful information. PMID- 10524590 TI - Lung health in Alberta farmers. AB - SETTING: A study conducted in the rural areas of two counties in east-central Alberta, Canada. OBJECTIVE: To investigate the relationship between lung health and dust exposure in farmers. DESIGN: A cross-sectional study of 781 farmers growing grain crops and raising livestock. Measurements included a questionnaire on respiratory symptoms, smoking habits and occupation, skin prick tests using common aeroallergens, and spirometry. RESULTS: Immediate skin reactivity to common aeroallergens was less prevalent in farmers with higher reported intensity of dust exposure. Respiratory symptoms suggestive of bronchitis had a significant dose-response relationship with the reported intensity of dust exposure. Respiratory symptoms consistent with bronchial responsiveness were significantly positively associated with cumulative dust exposure. There was a significant positive association between a physician's diagnosis of bronchitis and intensity of dust exposure. FEV1 and FEV1/FVC were significantly negatively associated with cumulative dust exposure. Ten years of exposure to a moderate dust level was associated with a deficit of 43 ml in the FEV1 and a deficit of 0.44% in the FEV1/FVC. CONCLUSIONS: Despite evidence of worker selection related to dust exposure, these farmers experienced respiratory symptoms, respiratory conditions, and reduced lung function associated with reported occupational dust exposure. PMID- 10524591 TI - Health status, dyspnea, lung function and exercise capacity in patients with chronic obstructive pulmonary disease. AB - SETTING: A secondary hospital outside Oslo. OBJECTIVE: To assess relationships between health status and measures of dyspnea, lung function and exercise capacity in patients with chronic obstructive pulmonary disease (COPD), to identify dimensions where lung-specific instruments associate and discriminate better than general measures. DESIGN: We assessed health status in 59 out patients with COPD, using the following instruments: Short Form 36 (SF-36)-a general health status measure, Respiratory Quality of Life Questionnaire (RQLQ)-a lung-specific measure, the Karnofsky performance scale, and a rating scale. All patients rated their dyspnea and had spirometry and exercise capacity measured. RESULTS: Mean (SD) patient age was 57.3 (9.7) years, FEV1 47% (15%) of predicted, 6 minute walk distance 503 m (122 m). Dyspnea was the strongest predictor for health status. Both SF-36 and RQLQ had dimensions associating well with dyspnea and exercise capacity. The associations with FEV1 ranged from none to moderate. CONCLUSION: All RQLQ scales had a moderate to substantial association with indices of dyspnea and exercise capacity, while the SF-36 associated well only in dimensions related to physical health. The general measure has a broader scope and complements the lung-specific measure. These findings support the construct validity of both the SF-36 and the RQLQ, and justify using a general measure to supplement a lung-specific measure. PMID- 10524592 TI - Smoking in Mediterranean countries: Europe, North Africa and the Middle-East. Results from a co-operative study. AB - SETTING: Although smoking is considered a major public health problem, it remains an important component of social behaviour and economic activity. OBJECTIVE: To provide an initial evaluation, from available data, of the main characteristics of smoking in the Mediterranean region. DESIGN: A questionnaire was sent to a group of correspondents-clinicians or epidemiologists involved in tobacco prevention-in the different countries. RESULTS: The proportion of smokers was quite different in men and women. In the majority of the countries over 45% of men and under 15% of women were smokers. The mean age of initiation of smoking was about 15. In every country mainly manufactured cigarettes were smoked, and the younger population preferred American cigarettes. Doctors and medical students had smoking habits similar to those of the general population. All of the countries included in the study had antismoking legislation, but only some put restrictions on advertising or sponsoring from the tobacco industry. CONCLUSION: To deal with the situation, recommendations have been proposed by a group of IUATLD experts in the different regions. The first of these is the implementation of a co-operative study in order to collect reliable data on smoking. Other recommendations are to set up educational programmes for health professionals to aid them in their smoking prevention activities. PMID- 10524593 TI - Tuberculosis and health sector reform: experience of integrating tuberculosis services into the district health system in rural South Africa. AB - SETTING: Hlabisa health district, South Africa. OBJECTIVE: To describe the integration of a 'vertical' tuberculosis control programme into an emerging 'horizontal' district health system, within the context of health sector reform. DESIGN: Descriptive account of the process of integration of the programme into the health system. RESULTS: A highly 'vertical' system of delivering tuberculosis treatment (with poor programme outcomes) was converted into a 'horizontal' team, integrated within the district health system, that used available resources such as village clinics and community health workers, with improved programme outcomes. CONCLUSIONS: In some settings at least, integration of tuberculosis 'programmes' into the district health system as tuberculosis 'teams' is feasible, and may produce highly cost-effective outcomes. PMID- 10524594 TI - Immune mediated 'HAART' attack during treatment for tuberculosis. Highly active antiretroviral therapy. AB - Highly active antiretroviral therapy (HAART) suppresses viral replication and improves immune function. However the inflammatory component of immune restoration can have clinically deleterious effects on previously asymptomatic infections. We report the development of acute respiratory failure in a patient after the institution of HAART, following 2 months of appropriate therapy for pulmonary tuberculosis. Necrotizing granulomas with acid-fast bacilli were found on lung biopsy, but cultures were negative for Mycobacterium tuberculosis and no other pathogens were isolated. Polymerase chain reaction of lung biopsy tissue for all mycobacterial species was positive only for M. tuberculosis. Rapid clinical improvement followed corticosteroid therapy. After initiating HAART, clinicians should be aware of the possibility of an inflammatory response to a previously quiescent tuberculous infection, even while on antituberculosis therapy. PMID- 10524595 TI - Creation and use of a Talairach-compatible atlas for accurate, automated, nonlinear intersubject registration, and analysis of functional imaging data. AB - Spatial normalization in functional imaging can encompass various processes, including nonlinear warping to correct for intersubject differences, linear transformations to correct for identifiable head movements, and data detrending to remove residual motion correlated artifacts. We describe the use of AIR to create a custom, site-specific, normal averaged brain atlas that can be used to map T2 weighted echo-planar images and coplanar functional images directly into a Talairach-compatible space. We also discuss extraction of characteristic descriptors from sets of linear transformation matrices describing head movements in a functional imaging series. Scores for these descriptors, derived using principal components analysis with singular value decomposition, can be treated as confounds associated with each individual image in the series and systematically removed prior to voxel-by-voxel statistical analysis. PMID- 10524596 TI - Sources of distortion in functional MRI data. AB - Functional magnetic resonance image (fMRI) experiments rely on the ability to detect subtle signal changes in magnetic resonance image time series. Any areas of signal change that correlate with the neurological stimulus can then be identified and compared with a corresponding high-resolution anatomical scan. This report reviews some of the several artefacts that are frequently present in fMRI data, degrading their quality and hence their interpretation. In particular, the effects of magnetic field inhomogeneities are described, both on echo planar imaging (EPI) data and on spiral imaging data. The modulation of these distortions as the subject moves in the magnet is described. The effects of gradient coil nonlinearities and EPI ghost correction schemes are also discussed. PMID- 10524597 TI - Functional MR imaging of confounded hypofrontality. AB - Comparatively reduced blood flow to frontal brain regions in patients with schizophrenia (hypofrontality) has been frequently observed in the last 25 years. However, there is an inconstant quality to hypofrontality, suggesting either confounded observation of a static (trait-like) abnormality, or that it is a genuinely dynamic (state-like) phenomenon. Possible confounds in functional magnetic resonance imaging (fMRI) studies of hypofrontality are classified. Methods for assessment and correction of stimulus correlated motion (an extracerebral confound) are reviewed in the context of fMRI data acquired from five schizophrenic patients and five comparison subjects during performance of a verbal fluency task. Factorial analysis of these and other data, acquired from the same subjects during a semantic decision task, is used to exclude a number of possible intracerebral confounds. By analogy to the historical controversy concerning the appearance of the planet Saturn viewed through early telescopes, understanding the inconstancy of hypofrontality in schizophrenia is likely to progress more by theoretically driven experiments that exploit the repeatability of fMRI than by further technological development alone. PMID- 10524598 TI - Revealing interactions among brain systems with nonlinear PCA. AB - In this work, we present a nonlinear principal component analysis (PCA) that identifies underlying sources causing the expression of spatial modes or patterns of activity in neuroimaging time series where these sources can interact to produce second-order modes. This nonlinear PCA uses a neural network architecture that embodies a specific form for the mixing of sources that is based on a second order approximation to any general nonlinear mixing. The modes obtained have a unique rotation and scaling that does not depend on the biologically implausible constraints adopted by conventional PCA. Interactions among sources render the expression of any mode or brain system sensitive to the expression of others. The example considers interactions among functionally specialized brain systems (using a fMRI study of colour and motion processing). PMID- 10524599 TI - Detecting changes in nonisotropic images. AB - If the noise component of image data is nonisotropic, i.e., if it has nonconstant smoothness or effective point spread function, then theoretical results for the P value of local maxima and the size of suprathreshold clusters of a statistical parametric map (SPM) based on random field theory are not valid. This assumption is reasonable for PET or smoothed fMRI data, but not if these data are projected onto an unfolded, inflated, or flattened 2D cortical surface. Anatomical data such as structure masks, surface displacements, and deformation vectors are also highly nonisotropic. The solution offered here is to suppose that the image can be warped or flattened (in a statistical sense) into a space where the data are isotropic. The subsequent corrected P values do not depend on finding this warping; it is sufficient only to know that such a warping exists. PMID- 10524600 TI - Scanning patients with tasks they can perform. AB - We present an overview of the types of imaging experiments that can be performed on psychologically impaired patients. The critical observation from such studies is a differential pattern of activation in the patients and normals. Underactivity is interpretable only when the patients make normal responses. In this context, a failure to activate a component region of the normal system implies that this region was not necessary for task performance. Overactivity indicates either cognitive or neuronal reorganisation. Neuronal reorganisation is indicated only if the patient performs the task using the same set of cognitive operations as normal subjects. Cognitive reorganisation can be demonstrated if the same activation pattern is elicited by normals when they are co-erced into using the same cognitive implementation as the patient. We conclude that the interpretation of neuroimaging studies of psychologically impaired patients depends on intact task performance and a detailed task analysis. When these criteria are met, patient studies can be used to identify: (1) necessary and sufficient brain systems, (2) dysfunction at sites distant to damage, (3) peri damage activation, and (4) compensation either at a neuronal level when pre existing cognitive strategies are re-instantiated using duplicated neuronal systems (degeneracy), or at a cognitive level when alternative cognitive strategies (and their corresponding brain systems) are adopted. PMID- 10524601 TI - Optimal experimental design for event-related fMRI. AB - An important challenge in the design and analysis of event-related or single trial functional magnetic resonance imaging (fMRI) experiments is to optimize statistical efficiency, i.e., the accuracy with which the event-related hemodynamic response to different stimuli can be estimated for a given amount of imaging time. Several studies have suggested that using a fixed inter-stimulus interval (ISI) of at least 15 sec results in optimal statistical efficiency or power and that using shorter ISIs results in a severe loss of power. In contrast, recent studies have demonstrated the feasibility of using ISIs as short as 500 ms while still maintaining considerable efficiency or power. Here, we attempt to resolve this apparent contradiction by a quantitative analysis of the relative efficiency afforded by different event-related experimental designs. This analysis shows that statistical efficiency falls off dramatically as the ISI gets sufficiently short, if the ISI is kept fixed for all trials. However, if the ISI is properly jittered or randomized from trial to trial, the efficiency improves monotonically with decreasing mean ISI. Importantly, the efficiency afforded by such variable ISI designs can be more than 10 times greater than that which can be achieved by fixed ISI designs. These results further demonstrate the feasibility of using identical experimental designs with fMRI and electro /magnetoencephalography (EEG/MEG) without sacrificing statistical power or efficiency of either technique, thereby facilitating comparison and integration across imaging modalities. PMID- 10524602 TI - Direct and indirect integration of event-related potentials, functional magnetic resonance images, and single-unit recordings. AB - Cognitive neuroimaging techniques vary along three primary dimensions: invasiveness, temporal resolution, and spatial resolution. Several of the major techniques excel on two of these three dimensions, but none of them excels on all three. In principle, multiple techniques with different strengths and weaknesses could be combined to obtain high temporal and spatial resolution data about human neural activity, and this article compares two approaches to combining microelectrode, hemodynamic, and electromagnetic measures of neural activity. The first approach involves using structural magnetic resonance images to provide a common reference frame for the mathematical estimation of neural activity, and the second approach involves parallel experimental manipulations and converging evidence. At present, neither approach is entirely satisfactory, and the integration of different measures of neural activity, therefore, requires a combination of direct and indirect approaches. PMID- 10524603 TI - Hemodynamic and electrophysiological study of the role of the anterior cingulate in target-related processing and selection for action. AB - A number of experiments requiring attention or other complex cognitive functions have found substantial activation in the anterior cingulate cortex (ACC). Some of these studies have suggested that this area may be involved in "selection for action," such as for selecting to respond to a target stimulus. Here, positron emission tomography (PET) and event-related potentials (ERPs) were used to study the effects of target probability during a demanding visual spatial attention task, in which the target percentage was either low (2%, 1 per approximately 26 sec) or high (16%, 1 per approximately 3.5 sec). As expected, ERPs to detected targets evoked large, bilaterally distributed P300 waves. The PET showed strong activation of the ACC, particularly dorsally, during all the attend conditions relative to passive. However, these PET activations did not significantly differ between the few-target and many-target conditions, showing only a small trend to be larger in the many-target case. Such results indicate that the bulk of the ACC activation does not reflect selection for action per se, while also suggesting that the ACC is not a likely source of the P300 effect. The current data, however, do not argue against the ACC serving a role in maintaining a vigilant or anticipatory state in which one may need to select for action, or in continually or repeatedly (i.e., for each stimulus) needing to resolve whether to select to act or to not act. PMID- 10524604 TI - Computational modeling of high-level cognition and brain function. AB - This article describes a computational modeling architecture, 4CAPS, which is consistent with key properties of cortical function and makes good contact with functional neuroimaging results. Like earlier cognitive models such as SOAR, ACT R, 3CAPS, and EPIC, the proposed cognitive model is implemented in a computer simulation that predicts observable variables such as human response times and error patterns. In addition, the proposed 4CAPS model accounts for the functional decomposition of the cognitive system and predicts fMRI activation levels and their localization within specific cortical regions, by incorporating key properties of cortical function into the design of the modeling system. PMID- 10524605 TI - Predicting human functional maps with neural net modeling. AB - Formidable difficulties exist in interpreting positron emission tomography (PET) and functional magnetic resonance imaging (fMRI) hemodynamic signals in terms of the underlying neural activity. These include issues of spatial and temporal resolution and problems relating neuronal activity (i.e., action potentials) measured in nonhuman studies by single unit electrodes to hemodynamic measurements reflecting synaptic activity. Also, regional hemodynamic measurements correspond to a mixture of local and afferent synaptic activity. To surmount these difficulties, we propose using large-scale neurobiologically realistic models in which data at various spatial and temporal levels can be simulated and cross-validated by multiple disciplines, including functional neuroimaging. A delayed match-to-sample visual task is used to illustrate this approach. PMID- 10524606 TI - Functional volumes modeling: scaling for group size in averaged images. AB - Functional volumes modeling (FVM) is a statistical construct for metanalytic modeling of the locations of brain functional areas as spatial probability distributions. FV models have a variety of applications, in particular, to serve as spatially explicit predictions of the Talairach-space locations of functional activations, thereby allowing voxel-based analyses to be hypothesis testing rather than hypothesis generating. As image averaging is often applied in the analysis of functional images, an important feature of FVM is that a model can be scaled to accommodate any degree of intersubject image averaging in the data set to which the model is applied. In this report, the group-size scaling properties of FVM were tested. This was done by: (1) scaling a previously constructed FV model of the mouth representation of primary motor cortex (M1-mouth) to accommodate various degrees of averaging (number of subjects per image = n = 1, 2, 5, 10), and (2) comparing FVM-predicted spatial probability contours to location-distributions observed in averaged images of varying n composed from randomly sampling a 30-subject validation data set. PMID- 10524607 TI - Interregional connectivity to primary motor cortex revealed using MRI resting state images. AB - The topographic organization of cortical neurons is traditionally examined using histological procedures. Functional magnetic resonance imaging (fMRI) offers the potential noninvasively to detect interregional connectivity of human brain. In the brain, there is spontaneous firing of neurons even in the resting state. Such spontaneous firing will increase local blood flow, cause MRI signal fluctuations, and affect remotely located neurons through the efferent output. By calculating covariance of each voxel referenced to the time course of a selected brain region, it is possible to detect the neurons connected to the selected region. Using this covariance method, neural connectivity to primary motor cortex was assessed during a resting state in six healthy right-handed volunteers. This interregional connectivity is similar to connectivity established by other anatomical, histochemical, and physiological techniques. This method may offer in vivo noninvasive measurements of neural projections. PMID- 10524608 TI - Perils of inadequacies in safety regulation. PMID- 10524609 TI - Terms of access to cloned mice comes under researchers' fire. PMID- 10524610 TI - Company to use advertising to cover Pubmed Central costs. PMID- 10524611 TI - Virus treatment questioned after gene therapy death. PMID- 10524612 TI - Japan joins efforts to patent cDNA clones. PMID- 10524613 TI - US provides funding for sequencing rice genome. PMID- 10524614 TI - Beyond 'substantial equivalence'. PMID- 10524616 TI - Immunology. Wavering on commitment. PMID- 10524615 TI - Cell cycle. Fools rush in. PMID- 10524617 TI - Cell motility. Bare bones of the cytoskeleton. PMID- 10524618 TI - Plant defence. Long view from a high plateau. PMID- 10524619 TI - Arsenic poisoning in the Ganges delta. PMID- 10524620 TI - Arsenic poisoning in the Ganges delta. PMID- 10524621 TI - A histone-H3-like protein in C. elegans. PMID- 10524622 TI - Commitment to the B-lymphoid lineage depends on the transcription factor Pax5. AB - The Pax5 gene encoding the B-cell-specific activator protein (BSAP) is expressed within the haematopoietic system exclusively in the B-lymphoid lineage, where it is required in vivo for progression beyond the pro-B-cell stage. However, Pax5 is not essential for in vitro propagation of pro-B cells in the presence of interleukin-7 and stromal cells. Here we show that pro-B cells lacking Pax5 are also incapable of in vitro B-cell differentiation unless Pax5 expression is restored by retroviral transduction. Pax5-/- pro-B cells are not restricted in their lineage fate, as stimulation with appropriate cytokines induces them to differentiate into functional macrophages, osteoclasts, dendritic cells, granulocytes and natural killer cells. As expected for a clonogenic haematopoietic progenitor with lymphomyeloid developmental potential, the Pax5-/- pro-B cell expresses genes of different lineage-affiliated programmes, and restoration of Pax5 activity represses this lineage-promiscuous transcription. Pax5 therefore plays an essential role in B-lineage commitment by suppressing alternative lineage choices. PMID- 10524623 TI - Low-temperature crystallization of silicate dust in circumstellar disks. AB - Silicate dust in the interstellar medium is observed to be amorphous, yet silicate dust in comets and interplanetary dust particles is sometimes partially crystalline. The dust in disks that are thought to be forming planets around some young stars also appears to be partially crystalline. These observations suggest that as the dust goes from the precursor clouds to a planetary system, it must undergo some processing, but the nature and extent of this processing remain unknown. Here we report observations of highly crystalline silicate dust in the disks surrounding binary red-giant stars. The dust was created in amorphous form in the outer atmospheres of the red giants, and therefore must be processed in the disks to become crystalline. The temperatures in these disks are too low for the grains to anneal; therefore, some low-temperature process must be responsible. As the physical properties of the disks around young stars and red giants are similar, our results suggest that low-temperature crystallization of silicate grains also can occur in protoplanetary systems. PMID- 10524624 TI - fMRI evidence for objects as the units of attentional selection. AB - Contrasting theories of visual attention emphasize selection by spatial location, visual features (such as motion or colour) or whole objects. Here we used functional magnetic resonance imaging (fMRI) to test key predictions of the object-based theory, which proposes that pre-attentive mechanisms segment the visual array into discrete objects, groups, or surfaces, which serve as targets for visual attention. Subjects viewed stimuli consisting of a face transparently superimposed on a house, with one moving and the other stationary. In different conditions, subjects attended to the face, the house or the motion. The magnetic resonance signal from each subject's fusiform face area, parahippocampal place area and area MT/MST provided a measure of the processing of faces, houses and visual motion, respectively. Although all three attributes occupied the same location, attending to one attribute of an object (such as the motion of a moving face) enhanced the neural representation not only of that attribute but also of the other attribute of the same object (for example, the face), compared with attributes of the other object (for example, the house). These results cannot be explained by models in which attention selects locations or features, and provide physiological evidence that whole objects are selected even when only one visual attribute is relevant. PMID- 10524625 TI - Involvement of visual cortex in tactile discrimination of orientation. AB - The primary sense modalities (vision, touch and so on) are generally thought of as distinct. However, visual imagery is implicated in the normal tactile perception of some object properties, such as orientation, shape and size. Furthermore, certain tactile tasks, such as discrimination of grating orientation and object recognition, are associated with activity in areas of visual cortex. Here we show that disrupting function of the occipital cortex using focal transcranial magnetic stimulation (TMS) interferes with the tactile discrimination of grating orientation. The specificity of this effect is illustrated by its time course and spatial restriction over the scalp, and by the failure of occipital TMS to affect either detection of an electrical stimulus applied to the fingerpad or tactile discrimination of grating texture. In contrast, TMS over the somatosensory cortex blocked discrimination of grating texture as well as orientation. We also report that, during tactile discrimination of grating orientation, an evoked potential is recorded over posterior scalp regions with a latency corresponding to the peak of the TMS interference effect (about 180 ms). The findings indicate that visual cortex is closely involved in tactile discrimination of orientation. To our knowledge, this is the first demonstration that visual cortical processing is necessary for normal tactile perception. PMID- 10524626 TI - Primate spinal interneurons show pre-movement instructed delay activity. AB - Preparatory changes in neural activity before the execution of a movement have been documented in tasks that involve an instructed delay period (an interval between a transient instruction cue and a subsequently triggered movement). Such preparatory activity occurs in many motor centres in the brain, including the primary motor cortex, premotor cortex, supplementary motor area and basal ganglia. Activity during the instructed delay period reflects movement planning, as it correlates with parameters of the cue and the subsequent movement (such as direction and extent), although it occurs well before muscle activity. How such delay-period activity shapes the ensuing motor action remains unknown. Here we show that spinal interneurons also exhibit early pre-movement delay activity that often differs from their responses during the subsequent muscle activity. This delay activity resembles the set-related activity found in various supraspinal areas, indicating that movement preparation may occur simultaneously over widely distributed regions, including spinal levels. Our results also suggest that two processes occur in the spinal circuitry during this delay period: the motor network is primed with rate changes in the same direction as subsequent movement related activity; and a superimposed global inhibition suppresses the expression of this activity in muscles. PMID- 10524627 TI - Polyamine-dependent facilitation of postsynaptic AMPA receptors counteracts paired-pulse depression. AB - At many glutamatergic synapses in the brain, calcium-permeable alpha - amino - 3 hydro - 5 - methyl - 4 - isoxazolepropionate receptor (AMPAR) channels mediate fast excitatory transmission. These channels are blocked by endogenous intracellular polyamines, which are found in virtually every type of cell. In excised patches, use-dependent relief of polyamine block enhances glutamate evoked currents through recombinant and native calcium-permeable, polyamine sensitive AMPAR channels. The contribution of polyamine unblock to synaptic currents during high-frequency stimulation may be to facilitate currents and maintain current amplitudes in the face of a slow recovery from desensitization or presynaptic depression. Here we show, on pairs and triples of synaptically connected neurons in slices, that this mechanism contributes to short-term plasticity in local circuits formed by presynaptic pyramidal neurons and postsynaptic multipolar interneurons in layer 2/3 of rat neocortex. Activity dependent relief from polyamine block of postsynaptic calcium-permeable AMPARs in the interneurons either reduces the rate of paired-pulse depression in a frequency-dependent manner or, at a given stimulation frequency, induces facilitation of a synaptic response that would otherwise depress. This mechanism for the enhancement of synaptic gain appears to be entirely postsynaptic. PMID- 10524628 TI - Signal relay by BMP antagonism controls the SHH/FGF4 feedback loop in vertebrate limb buds. AB - Outgrowth and patterning of the vertebrate limb are controlled by reciprocal interactions between the posterior mesenchyme (polarizing region) and a specialized ectodermal structure, the apical ectodermal ridge (AER). Sonic hedgehog (SHH) signalling by the polarizing region modulates fibroblast growth factor (FGF)4 signalling by the posterior AER, which in turn maintains the polarizing region (SHH/FGF4 feedback loop). Here we report that the secreted bone morphogenetic-protein (BMP) antagonist Gremlin relays the SHH signal from the polarizing region to the AER. Mesenchymal Gremlin expression is lost in limb buds of mouse embryos homozygous for the limb deformity (Id) mutation, which disrupts establishment of the SHH/FGF4 feedback loop. Grafting Gremlin-expressing cells into ld mutant limb buds rescues Fgf4 expression and restores the SHH/FGF4 feedback loop. Analysis of Shh-null mutant embryos reveals that SHH signalling is required for maintenance of Gremlin and Formin (the gene disrupted by the ld mutations). In contrast, Formin, Gremlin and Fgf4 activation are independent of SHH signalling. This study uncovers the cascade by which the SHH signal is relayed from the posterior mesenchyme to the AER and establishes that Formin dependent activation of the BMP antagonist Gremlin is sufficient to induce Fgf4 and establish the SHH/FGF4 feedback loop. PMID- 10524629 TI - Long-term in vivo reconstitution of T-cell development by Pax5-deficient B-cell progenitors. AB - The mechanisms controlling the commitment of haematopoietic progenitors to the B lymphoid lineage are poorly understood. The observations that mice deficient in E2A and EBF lack B-lineage cells have implicated these two transcription factors in the commitment process. Moreover, the expression of genes encoding components of the rearrangement machinery (RAG1, RAG2, TdT) or pre-B-cell receptor (lambda5, VpreB, Igalpha, Igbeta) has been considered to indicate B-lineage commitment. All these genes including E2A and EBF are expressed in pro-B cells lacking the transcription factor Pax5. Here we show that cloned Pax5-deficient pro-B cells transferred into RAG2-deficient mice provide long-term reconstitution of the thymus and give rise to mature T cells expressing alpha/beta-T-cell receptors. The bone marrow of these mice contains a population of cells of Pax5-/- origin with the same phenotype as the donor pro-B cells. When transferred into secondary recipients, these pro-B cells again home to the bone marrow and reconstitute the thymus. Hence, B-lineage commitment is determined neither by immunoglobulin DJ rearrangement nor by the expression of E2A, EBF, lambda5, VpreB, Igalpha and Igbeta. Instead, our data implicate Pax5 in the control of B-lineage commitment. PMID- 10524630 TI - Dimerization inhibits the activity of receptor-like protein-tyrosine phosphatase alpha. AB - Protein-tyrosine phosphatases (PTPs) are vital for regulating tryosine phosphorylation in many processes, including growth and differentiation. The regulation of receptor-like PTP (RPTP) activity remains poorly understood, but based on the crystal structure of RPTPalpha domain 1 we have proposed that dimerization can negatively regulate activity, through the interaction of an inhibitory 'wedge' on one monomer with the catalytic cleft of domain 1 in the other monomer. Here we show that dimerization inhibits the activity of a full length RPTP in vivo. We generated stable disulphide-bonded full-length RPTPalpha homodimers by expressing mutants with single cysteines at different positions in the ectodomain juxtamembrane region. Expression of wild-type RPTPalpha and Phe135Cys and Thr141Cys mutants in RPTPalpha-null mouse embryo cells increased dephosphorylation and activity of Tyr 529 in the protein tyrosine kinase c-Src; in contrast, expression of a Pro137Cys mutant did not. Mutation of Pro 210/211 to leucine in the inhibitory wedge of the Pro137Cys mutant restored its ability to activate c-Src, indicating that dimerization may inhibit full-length RPTPalpha activity in a manner stereochemically consistent with RPTPalpha crystal structures. Our results suggest that RPTPalpha activity can in principle be negatively regulated by dimerization in vivo. PMID- 10524631 TI - Phytochrome signalling is mediated through nucleoside diphosphate kinase 2. AB - Because plants are sessile, they have developed intricate strategies to adapt to changing environmental variables, including light. Their growth and development, from germination to flowering, is critically influenced by light, particularly at red (660 nm) and far-red (730 nm) wavelengths. Higher plants perceive red and far red light by means of specific light sensors called phytochromes(A-E). However, very little is known about how light signals are transduced to elicit responses in plants. Here we report that nucleoside diphosphate kinase 2 (NDPK2) is an upstream component in the phytochrome signalling pathway in the plant Arabidopsis thaliana. In animal and human cells, NDPK acts as a tumour suppressor. We show that recombinant NDPK2 in Arabidopsis preferentially binds to the red-light activated form of phytochrome in vitro and that this interaction increases the activity of recombinant NDPK2. Furthermore, a mutant lacking NDPK2 showed a partial defect in responses to both red and farred light, including cotyledon opening and greening. These results indicate that NDPK2 is a positive signalling component of the phytochrome-mediated light-signal-transduction pathway in Arabidopsis. PMID- 10524633 TI - 14-3-3Sigma is required to prevent mitotic catastrophe after DNA damage. AB - 14-3-3Sigma is a member of a family of proteins that regulate cellular activity by binding and sequestering phosphorylated proteins. It has been suggested that 14-3-3sigma promotes pre-mitotic cell-cycle arrest following DNA damage, and that its expression can be controlled by the p53 tumour suppressor gene. Here we describe an improved approach to the generation of human somatic-cell knockouts, which we have used to generate human colorectal cancer cells in which both 14-3 3sigma alleles are inactivated. After DNA damage, these cells initially arrested in the G2 phase of the cell cycle, but, unlike cells containing 14-3-3sigma, the 14-3-3sigma-/- cells were unable to maintain cell-cycle arrest. The 14-3-3sigma-/ cells died ('mitotic catastrophe') as they entered mitosis. This process was associated with a failure of the 14-3-3sigma-deficient cells to sequester the proteins (cyclin B1 and cdc2) that initiate mitosis and prevent them from entering the nucleus. These results may indicate a mechanism for maintaining the G2 checkpoint and preventing mitotic death. PMID- 10524632 TI - Reconstitution of actin-based motility of Listeria and Shigella using pure proteins. AB - Actin polymerization is essential for cell locomotion and is thought to generate the force responsible for cellular protrusions. The Arp2/3 complex is required to stimulate actin assembly at the leading edge in response to signalling. The bacteria Listeria and Shigella bypass the signalling pathway and harness the Arp2/3 complex to induce actin assembly and to propel themselves in living cells. However, the Arp2/3 complex alone is insufficient to promote movement. Here we have used pure components of the actin cytoskeleton to reconstitute sustained movement in Listeria and Shigella in vitro. Actin-based propulsion is driven by the free energy released by ATP hydrolysis linked to actin polymerization, and does not require myosin. In addition to actin and activated Arp2/3 complex, actin depolymerizing factor (ADF, or cofilin) and capping protein are also required for motility as they maintain a high steady-state level of G-actin, which controls the rate of unidirectional growth of actin filaments at the surface of the bacterium. The movement is more effective when profilin, alpha-actinin and VASP (for Listeria) are also included. These results have implications for our understanding of the mechanism of actin-based motility in cells. PMID- 10524634 TI - Optimum dose of ursodeoxycholic acid in primary biliary cirrhosis. AB - BACKGROUND: Ursodeoxycholic acid (UDCA) improves liver function tests and prolongs survival in primary biliary cirrhosis (PBC). The dose of 10- 15 mg/kg/day used in the large trials has largely been based on that used for gallstone dissolution. The only dose-response study of UDCA in PBC suggested that a dose of 8 mg/kg/day was the most efficacious. However, disease stage of the patients was not known, higher doses of UDCA were not tried and there was no 'washout period' between the different doses. The aim of this study was to determine the optimum dose of UDCA in early-stage PBC (stage 1 and 2). METHODS: Twenty-four biopsy-proven early-stage PBC patients (one male, 23 female) received five doses of UDCA (0, 300, 600, 900, 1200 mg/day) each for 8 weeks with 4-week washout periods between doses. Symptoms (pruritus, fatigue, diarrhoea) were assessed on a four-point scale (none, mild, moderate, severe). Liver function tests (LFTs) were performed using conventional methods, and serum bile acids were measured using gas liquid chromatography. RESULTS: The dose of 900 mg/day produced the greatest enrichment of UDCA in serum bile acids; although there was no difference in the enrichment of UDCA between the different doses. There was a trend towards normalization of the abnormal LFTs in a dose-dependent manner (for y-glutamyl transferase (yGT), alkaline phosphatase (ALP), alanine transaminase (ALT) and IgM). Multi-factorial analysis showed that UDCA treatment, irrespective of dose, was significantly better than placebo for all the variables. The 900 and 1200 mg doses were better than both 300 and 600 mg using yGT and total bilirubin as variables, better than 300 mg using ALP and IgM as variables, and better than 600 mg using albumin as a variable. No variables showed a significant difference between 900 and 1200 mg. CONCLUSION: The optimum dose of UDCA is 900 mg/day (equivalent to 13.5 mg/kg/day). PMID- 10524635 TI - Combined ursodeoxycholic acid and glycyrrhizin therapy for chronic hepatitis C virus infection: a randomized controlled trial in 170 patients. AB - OBJECTIVE AND DESIGN: To assess the efficacy and safety of combination therapy using ursodeoxycholic acid with glycyrrhizin for chronic hepatitis C virus infection, we conducted a prospective randomized controlled trial of glycyrrhizin (group G) compared with glycyrrhizin plus ursodeoxycholic acid (group G+U) in 170 patients. METHODS: All patients had elevated serum aminotransferase levels over 6 months before entry into the trial. Glycyrrhizin was administered to both groups for 24 weeks, and in group G+U, ursodeoxycholic acid (600 mg/day) was administered orally as well. RESULTS: Serum aspartate transaminase and alanine transaminase concentrations significantly decreased during treatment in both groups, but serum gamma-glutamyl transpeptidase concentrations fell significantly only in group G+U. Concentrations of all three enzymes fell significantly more in group G+U than in group G, and had normalized in more cases when the trial ended at 24 weeks. However, levels of HCV viraemia did not change during the trial in either group. Multiple regression analysis linked only the treatment regimen, not HCV-related factors or liver histology, to the degree of serum enzyme reduction. No adverse effects were noted in either group. CONCLUSIONS: The combined therapy with ursodeoxycholic acid and glycyrrhizin is safe and effective in improving liver-specific enzyme abnormalities, and may be an alternative to interferon in chronic hepatitis C virus infection, especially for interferon-resistant or unstable patients. PMID- 10524637 TI - Abnormal clotting parameters before therapeutic ERCP: do they predict major bleeding? AB - A total of 399 consecutive patients undergoing 598 ERCPs (endoscopic retrograde cholangiopancreatographies), including 88 pre-cut papillotomies and 206 conventional papillotomies, are described in a retrospective study. Clotting parameters, haemoglobin levels, indications for pre-cut and/or conventional papillotomy and the use of drugs assumed to interfere with blood clotting (anticoagulants, platelet-aggregation inhibitors, low-molecular-weight heparin) were evaluated in order to detect risk factors for ERCP-associated bleeding. The overall incidence of ERCP-associated bleeding was 18/598 (3.0%). The incidence of bleeding in the group without papillotomy was 7/346 (2.0%). This group consisted of patients who underwent only a diagnostic ERCP, patients who had undergone papillotomy previously, patients in whom a renewed attempt was made to extract biliary stones, and patients in whom removal or change of a stent was necessary. The incidence of papillotomy-associated bleeding was 11/252 (4.4%). Pre-cut papillotomy did and conventional papillotomy did not significantly increase the incidence of bleeding: 15.2% (P < 0.001) and 1.9% (P= 1.00) respectively. The incidence of ERCP-associated bleeding in the group not using any drugs interfering with blood clotting was 2.5%. The use of low-molecular-weight heparin (10.3%) during ERCP significantly increased the risk of bleeding (P= 0.01). However, the use of platelet aggregation inhibitors (2.4%) did not (P= 1.00). As the incidence of bleeding in patients with normal clotting parameters, including the patients with abnormal parameters which were well corrected (4.3%), was higher than in patients with abnormal haemostatic screens (2.7%), abnormal coagulation tests did not predict ERCP-associated bleeding. PMID- 10524636 TI - Non-adrenergic, non-cholinergic regulation of stone-diseased and stone-free human gallbladders. AB - OBJECTIVE: The aim of this study was to determine the role of nitric oxide (NO), as the primary neurotransmitter of non-adrenergic, non-cholinergic (NANC) innervation, in stone-diseased and stone-free human gallbladders. METHODS: Human gallbladder muscle strips were mounted in modified Krebs-Henseleit solution with atropine (1 mM), guanethidine sulfate (5 mM) and aerated with Carbogen. Electric field stimulation (EFS) (70 V, 0.5 ms, 100 pulses) was used to activate NANC nerves. N-omega-nitro-L-arginine (L-NNA, 100 mM) and L-arginine (L-Arg, 120 mM) were used to manipulate the NO-synthase. Gallbladder slices were stained by using the alkaline phosphatase, anti-alkaline phosphatase (APAAP) method for histological examination. RESULTS: In the control group (basal tone, 8.94 +/- 1.17 mN) caused a frequency-dependent reduction of basal tone (1 Hz = 5.73 +/- 0.81 mN; 3 Hz = 5.18 +/- 0.65 mN; 10 Hz = 4.63 +/- 0.49 mN), inhibition of NO synthesis with L-NNA increased the tone (7.63 +/- 0.76 mN). Stone-diseased groups were divided into two groups (contractor and subcontractor), according to their ability to contract by CCK. Under the influence of EFS the contractor group (basal tone = 7.79 +/- 0.93 mN) reacted like the control group, but was frequency independent and, additionally, showed spontaneous phasic contractions. In the sub contractor group (basal tone 4.13 +/- 0.65 mN) EFS only decreased the frequency of spontaneous phasic contractions. L-NNA caused an increase in tone (5.97 +/- 0.84 mN) and frequency. L-arginine, the substrate for NO-synthase, significantly reversed this effect, indicating the dependence on NO. Histologically, the contractor group showed a wrinkled mucosal membrane and low-grade inflammation. Shallow mucosa, necrosis and high-grade inflammation were found in the sub contractor group. CONCLUSIONS: In vitro, NANC relaxation of human gallbladder is NO dependent. The motility of stone-diseased gallbladders is modulated by NO and seems to depend on the degree of scarification. PMID- 10524638 TI - The network: a strategy to describe the relationship between quality of life and disease activity. The case of inflammatory bowel disease. AB - OBJECTIVE: Health is a complex and multi-dimensional entity and is neither easily determined nor easily conveyed to others. Publications have often combined various variables of disease activity and health-related quality of life (HRQoL), used the variables interchangeably or utilized summation indices to compare health assessment. The aim of this study is to investigate the relationship between measurements of disease activity and HRQoL. STUDY: design Cross-sectional evaluation of disease activity and HRQoL. STUDY POPULATION: Two hundred and eleven consecutive patients with ulcerative colitis. SETTING: The catchment area of Linkoping University Hospital. MEASUREMENTS: HRQoL was measured using two questionnaires, the Sickness Impact Profile (SIP) and the Rating Form of IBD Patient Concerns (RFIPC). Patients were also asked if they were 'feeling fit and well', as a measurement of general health perception. Disease activity was measured by means of symptom cards, laboratory tests and sigmoidoscopy. RESULTS: The correlations (Spearman's r (r5)) between variables of disease activity and HRQoL were low. 'Feeling fit and well' was best correlated to worries and concerns (the RFIPC, rs 0.32, P < 0.05), while there was a decreasing association with subjective functional status (the SIP, rs 0.31, P < 0.05), symptoms (stools per day, rs 0.15, not significant) and biological variables (endoscopy score, rs 0.04, not significant). CONCLUSION: The correlations between traditional measurements of disease activity and various measures of HRQoL are low. We therefore propose a system whereby the process is conceptualized using a 'network strategy', ordering the measurements of disease activity and HRQoL into five dimensions: biological variables, symptoms, functional status, worries and concerns, and health perceptions. We feel that this method of interpretation more accurately reflects the overall health of a group of patients with IBD than more traditional summation indices. PMID- 10524639 TI - T-cell co-stimulatory molecules are upregulated on intestinal macrophages from inflammatory bowel disease mucosa. AB - BACKGROUND AND AIMS: Macrophages play an important role during mucosal inflammation in inflammatory bowel disease (IBD). As the co-stimulatory molecules B7-1 (CD80) and B7-2 (CD86) play an integral role in the activation of T cells by antigen-presenting cells (APC) we investigated the surface expression of B7-1 and B7-2 on colonic macrophages from normal and IBD mucosa. METHODS: Intestinal macrophages were isolated from biopsies of 13 control persons and 14 patients with IBD (seven with Crohn's disease (CD); and seven with ulcerative colitis (UC)). Cells were characterized by triple fluorescence flow cytometrical analysis using CD33 as macrophage marker. RESULTS: The expression of B7-1 (CD80) (9.2% +/- 4.2%) and B7-2 (CD86) (15.1% +/- 7.3%) was low on colonic macrophages from normal mucosa, indicating only a low antigen presenting potential. However, on macrophages from IBD colon there was a significant increase in the expression of co-stimulatory molecules (CD80, 33.8% +/- 8.9%, P = 0.00005 vs. control; CD86, 39.9% +/- 8.8%, P = 0.00002). There was no significant difference between CD and UC in the expression of CD80 (CD, 31.3% +/- 6.7%; UC, 34.4% +/- 13.3%) and CD86 (CD, 41.9% +/- 3.8%; UC, 35.6% +/- 13.8%). While in normal mucosa only 10.6% +/- 4.9% of the macrophages expressed CD14, more than 90% of the CD86/CD80 positive cells of the inflamed mucosa were positive for CD14. CONCLUSION: Colonic macrophages from normal mucosa rarely express the co-stimulatory molecules CD80 and CD86. In IBD a new macrophage population is found with high expression of co stimulatory molecules presumably responsible for the perpetuated immune response. PMID- 10524640 TI - Acute inflammatory intestinal vascular lesions and in situ abnormalities of the plasminogen activation system in Crohn's disease. AB - OBJECTIVES: The distribution of the intestinal vascular lesions and their relation with the fibrinolysis process are poorly known in Crohn's disease (CD). The mediators of the plasminogen activator system, namely urokinase-type plasminogen activator (u-PA), tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor type-1 (PAI-1), are a key complex involved in fibrinolysis. The aims of this study were: (1) to further define vascular lesions and their distribution in the intestine; and (2) to study concomitantly the qualitative in situ expression and the levels of u-PA, t-PA and PAI-1 in the ileum of patients with CD. PATIENTS AND METHODS: Histological, immunohistochemical and ultrastructural studies of vascular lesions in the resected ileum of 27 patients with CD were performed and compared with 36 control patients. Levels of u-PA, t-PA and PAI-1 measured by ELISA methods were compared in healthy and inflamed ileal tissues of 17 patients with CD. RESULTS: Acute vascular lesions involving mainly serosal venules and capillaries were present in 63% of patients with CD vs 3/36 controls and were associated with PAI-1 expression. They were prominent on the mesenteric border beneath macroscopically normal mucosa. In contrast, chronic vascular lesions were present in all layers beneath mucosal ulcerations, where a significant increase of PAI-1 levels was found. CONCLUSIONS: These results suggest that vascular involvement associated with abnormalities of PAI-1 expression is an early and widespread event in CD. Their prominence on the mesenteric border might explain the characteristic location of CD ulceration along the mesenteric margin. PMID- 10524641 TI - Epidemiological and clinical features of Spanish patients with Crohn's disease. Spanish Epidemiological and Economic Study Group on Crohn's disease. AB - OBJECTIVES: To study the epidemiological features, clinical profile and drug utilization patterns of patients with Crohn's disease (CD). DESIGN: A cross sectional study of 635 Spanish patients with CD included through a stratified and proportional random sampling. RESULTS: The mean age of the patients was 33 years (SD 11.9) (52% were women). Nine per cent of the patients had a family history of CD and 14% had a history of inflammatory bowel disease (IBD). Over half (54%) of the patients were smokers. The most frequent localization of CD was ileocolonic involvement (47%). Twelve per cent had peri-anal disease. Regarding symptom pattern, 23% of the patients had chronic active disease and 42% had experienced no relapses during the previous 12 months. Compared with the inflammatory pattern, fistulizing pattern was associated with a significantly higher proportion of patients with chronic active disease, a higher number of relapses per year, and a higher incidence of post-surgical relapses. Sixty-one per cent of the patients had complications and 35% were hospitalized. Acute relapse and bowel stricture were the most common complications. The need for hospitalization was higher in patients with fistulizing pattern. Regarding medication, 77% and 41 % of the patients were taking aminosalicylates and corticosteroids respectively, mainly on a long-term basis. Mean daily doses (MDD) were 2.2 g/day and 31.7 mg/day respectively. Twenty-one per cent had received immunosuppressors with a MDD of 1.6 mg/kg/day. Fifteen per cent of the patients had received metronidazole or ciprofloxacin while 5% were taking anti-diarrhoeal drugs. CONCLUSIONS: Spanish patients with CD are young, have frequent hospitalization requirements, complications, and a high consumption of drugs. PMID- 10524642 TI - Risk indicators of organic diseases in uninvestigated dyspepsia: a one-week survey in 246 Italian endoscopy units. AB - OBJECTIVE: To evaluate the predictive value of demographic and clinical features and of the results of an office-based test for Helicobacter pylori antibodies, in the presence of organic dyspepsia. DESIGN: Over a 1-week period, 2206 consecutive patients first referred for endoscopy in 246 Italian centres were included. METHODS: Demographic and clinical features, endoscopy findings, and histological diagnosis of H. pylori infection were recorded for all patients. IgG antibodies to H. pylori were determined in 2128 cases by a rapid, immunochromatographic method (Flex Sure HP, S.K.D., San Jose, CA). RESULTS: Endoscopic abnormalities were found in 939 patients (42.6%). Histologically assessed H. pylori infection was predictive for duodenal ulcer (odds ratio (OR), 6.79; 95% confidence interval (CI), 4.4-10.5). Being male (OR, 1.97; 95% CI, 1.7-2.3), older than 40 years (OR, 1.81; 95% CI, 1.5-2.2), a smoker (OR, 1.88; 95% CI, 1.6-2.3), and presenting nocturnal awakening (OR, 1.62; 95% CI, 1.3-2.0) were independently associated with secondary dyspepsia. Epigastric (OR, 1.50; 95% CI, 1.2-1.9) and retrosternal pain (OR, 1.39; 95% CI, 1.1 -1.8) severe enough to affect the usual activities were predictive of organic diseases. The results of the Flex Sure HP test correlated poorly with histological findings. CONCLUSIONS: Male gender, older age, cigarette smoking, a family history of peptic ulcer, symptoms severe enough to induce awakening, epigastric/retrosternal pain severe enough to influence the usual activities are all independently (although weakly) associated with organic dyspepsia. H. pylori infection is strongly associated with duodenal ulcer, but the rapid test we used was not sensitive enough to achieve clinical utility. PMID- 10524644 TI - A new, non-invasive method for detection of Helicobacter pylori: validity in the routine clinical setting. AB - BACKGROUND: Helicobacter pylori has been recognised as a major gastric pathogen. Many techniques to identify infection have been developed, including histology and culture as invasive tests and the urea breath test as non-invasive technology. Recently, another non-invasive test based on the detection of H. pylori antigens in stool specimens was introduced. AIM: To evaluate the sensitivity and specificity of this novel enzyme-linked immunosorbent assay for detection of H. pylori infection in comparison with histology. METHODS: Stool specimens of 72 consecutive patients who had gastroscopy with biopsy at our endoscopy unit were collected and frozen at -20 degrees C until further processing. H. pylori status was determined as part of the routine histological work-up. In a second step, histology slides were reviewed by a pathologist who specialized in gastrointestinal pathology. Stool samples were tested for the presence of H. pylori antigen using the PREMIER H. pylori Stool Antigen Test. Sensitivity and specificity of routine histopathology and the H. pylori Stool Antigen Test were determined using the diagnosis of the expert gastrointestinal histopathologist as gold standard. RESULTS: Routine histopathology resulted in a sensitivity of 90% and specificity of 92% in the diagnosis of H. pylori. These results compare well with the results of the H. pylori Stool Antigen Test that revealed a sensitivity of 80% and a specificity of 98%. CONCLUSION: The H. pylori Stool Antigen Test is a promising non-invasive test for the detection of H. pylori infection. PMID- 10524643 TI - The impact of short-term ranitidine use on the precision of the 13C-urea breath test in subjects infected with Helicobacter pylori. AB - BACKGROUND: The 13C-urea breath test (13C-UBT) is a very accurate method of Helicobacter pylori diagnosis with a false-negative rate of 1-3%. However, the accuracy of the 13C-UBT is affected by potent acid inhibition with proton-pump inhibitors, which may suppress H. pylori and cause false-negative results. It is not known whether this occurs with less potent acid inhibition by H2-antagonists and any effect may be important clinically. OBJECTIVE: To determine the kinetics of 13CO2 excretion in H. pylori infected subjects during and after short-term ranitidine use. METHODS: Volunteers underwent a baseline 13C-UBT (positive: delta13CO2 > or = 5.0; negative: < or = 3.5; indeterminate: > 3.5 to < 5.0). Infected subjects took ranitidine 300 mg each evening for up to 28 days. 13C-UBTs were performed at weekly intervals and then every other day after ranitidine was ceased. If the 13C-UBT remained positive after 14 days, ranitidine was continued for a further 14 days. RESULTS: Thirty-one subjects were studied (mean age 40.4 +/- 2.1 years; 23 female/8 male; mean baseline delta13CO2 27.3 +/- 2.5). In 28 subjects the 13C-UBT remained positive during ranitidine use. The mean delta13CO2 rose to 124% (P< 0.06) and 121% (P < 0.05) of baseline at 14 and 28 days respectively. In two subjects, the delta13CO2 became indeterminate at day 7 (delta13CO2 4.3 and 3.8). In one of these, return to a positive value (delta13CO2 13.6; 103% of baseline) occurred while still on ranitidine. The other subject became positive again by day 3 off ranitidine (17.8; 119% of baseline). One subject had a transiently negative test after 21 days and this became positive again while still taking ranitidine. CONCLUSIONS: Ranitidine has a minimal effect on the 13C-UBT. The rate of indeterminate or false-negative tests is no greater than in patients on no anti-secretory medication. PMID- 10524645 TI - Motility of Helicobacter pylori in a viscous environment. AB - BACKGROUND: Patients with gastroduodenal disease produce gastric mucus of higher viscosity, and mucins that are of a smaller size, than normal. We have modelled these changes to the mucus layer in solutions of methylcellulose, and measured bacterial motility in biopsied mucus, to assess how they might influence the movements of Helicobacter pylori. METHODS: Motilities of Helicobacter pylori were measured in solutions of methylcellulose with molecular mass of 14 and 41 kDa, and in biopsied mucus with a Hobson BacTracker. Four parameters of bacterial motility were quantified: curvilinear velocity (CLV), path length, track linearity and curvature rate. RESULTS: All H. pylori were motile in methylcellulose solutions, and had optimal motilities at a viscosity of 3 cp (CLV in methylcellulose of 41 kDa, for instance, was 33 +/- 1.4 microm/s (mean +/- SEM) and the path length in methylcellulose of 41 kDa was 22.4 +/- 2 microm). At higher viscosities, mean CLVs, path lengths and curvature rates decreased, and track linearities increased in direct proportion to the increase in methylcellulose viscosity. Bacteria become non-motile at a viscosity of 50 cp in methylcellulose of 14 kDa, and at 70 cp in methylcellulose of 41 kDa. Mean CLVs, path lengths and curvature rates (but not track linearities) were greater in methylcellulose of 41 kDa than in methylcellulose of 14 kDa at each viscosity tested. Motilities of H. pylori from patients with duodenal ulcer or non-ulcer dyspepsia in methylcellulose solutions were not significantly different. H. pylori had poor motility in biopsied mucus, but became highly motile when biopsied mucus was diluted with saline. CONCLUSIONS: The viscosity-motility profiles of H. pylori in methylcellulose and the motilities of H. pylori in biopsied mucus suggest (1) that H. pylori may have poor motility in mucus at the epithelial surface, but high motility at the luminal surface of the mucus layer, and (2) that the increased mucus viscosity and decreased mucin size in patients with gastroduodenal disease act in combination to decrease H. pylori motility in vivo. PMID- 10524646 TI - Gastric involvement in progressive systemic sclerosis: electrogastrographic and sonographic findings. AB - OBJECTIVE: To determine whether electrogastrography (EGG) can discern sonographically demonstrated motility disorders in patients with progressive systemic sclerosis (SSc) and to evaluate EGG as a possible diagnostic tool. DESIGN: Prospective study with control group and testing for reliability. SUBJECTS: 15 SSc patients [women aged 33-70 years (mean 53.3 years)] and 15 healthy volunteers. METHODS: Bipolar cutaneous EGG was recorded to obtain the following parameters: period dominant frequency (PDF), percentage of gastric dysrhythmia and normogastria (defined as 2-4/min), period dominant power (PDP) and its change after a standardized meal of 500 kcal (2093 kJ), and instability coefficients of dominant frequency and power (DFIC, DPIC). Simultaneously, real time sonography was performed in the aortomesenteric plane (3.5-MHz curved-array probe). In 10 patients and 13 control subjects, the distance from the anterior wall of the gastric antrum to the abdominal skin was measured. RESULTS: Three patients (20%) showed hypomotility of the gastric antrum sonographically. The percentage of bradygastria was significantly lower in these patients, but the PDF, DFIC and DPIC values were not significantly different. The distance between the cutaneous electrodes and the antrum bore a greater relationship to the PDP values than did the sonographically demonstrated number of gastric contractions. CONCLUSIONS: Although cutaneous EGG can be performed in SSc patients without apparent derangement in frequency and stability of the signal, it offers no advantage over sonography in diagnosis and follow-up. PMID- 10524647 TI - Long-term palliation in metastatic carcinoid tumours with various applications of meta-iodobenzylguanidin (MIBG): pharmacological MIBG, 131I-labelled MIBG and the combination. AB - Carcinoid tumours are rare, but well known for their characteristic presentation with diarrhoea and flushes due to overproduction of serotonin in the case of liver metastases. Treatment is mainly based on the reduction of vasoactive peptide hypersecretion and symptomatic improvement Octreotide and interferon are widely applied and effective treatment options to induce symptomatic improvement and, to a lesser extent, biochemical response. The main drawbacks, however, are the need for frequent injections and/or the occurrence of side effects. A rather new approach is the application of meta-iodobenzylguanidine (MIBG), which resembles noradrenalin and serotonin. In carcinoid patients, MIBG is taken up in the tumour cells and stored in the neurosecretory granules. When labelled with 131 iodine, radionuclide imaging is positive in up to 70% of the patients. In these patients, two cycles of a therapeutic dose of radioactive MIBG may induce long-lasting palliation (8 months) by internal irradiation. Also, the non radioactive MIBG compound may be effective in palliation, even in patients with a negative scan. The mode of action is based on specific tumour acidification as found in animal models, and/or based on its effect as a false neurotransmittor. Three case reports demonstrate different therapeutic possibilities of MIBG: 1) symptomatic relief with unlabelled MIBG, which is a safe and simple treatment; 2) the longterm palliation following radioactive treatment; and 3) an additional new aspect of predosing with unlabelled MIBG followed by radioactive MIBG led to improved tumour targeting and impressive clinical response. PMID- 10524648 TI - Combination therapy with mycophenolate mofetil and ursodeoxycholic acid for primary biliary cirrhosis. AB - Evidence of autoimmunity in primary biliary cirrhosis (PBC) provides a rationale for treatment with an immunosuppressant. Such a drug should be sufficiently free from serious toxicities to enable patients with asymptomatic, but progressive, disease to be treated longterm. Mycophenolate mofetil (MMF) mediates immunosuppression by selectively and reversibly inhibiting lymphocyte function, and has a more acceptable safety profile than other immunosuppressants that have been used in the treatment of this disease. Two patients, whose response to long term treatment with ursodeoxycholic acid (UDCA) had been inadequate, were treated with a combination of MMF 2 g daily and UDCA 1 g daily for 12 months. In both patients this regimen was associated with no clinically significant adverse events, a decrease in elevated serum alkaline phosphatase levels to values close to the upper limit of normal, and an almost complete disappearance of the chronic inflammatory cell infiltrate that had been present in pre-combination treatment liver biopsies. MMF may be an appropriate immunosuppressive drug for use in the long-term treatment of patients with PBC, including asymptomatic patients. PMID- 10524649 TI - Intra-hepatic false aneurysm: a rare complication of ERCP. AB - We report a rare case in which an intra-hepatic false aneurysm formed following endoscopic retrograde cholangiopancreatography and presented with life threatening gastrointestinal bleeding. The aetiology, investigation and management of intra-hepatic false aneurysm is discussed. PMID- 10524650 TI - Ileal aberrant pancreas induces intussusception and gastrointestinal bleeding in an adult woman--case report. AB - Aberrant pancreas is a congenital anomaly. In surgical series, its incidence varied from 0.2 to 0.8%. About 70% of aberrant pancreas occur in the gastrointestinal tract. Eighty percent of them locate in the stomach and duodenum, and only 0.2% in the ileum. We report on a 25-year-old woman with ileal aberrant pancreas who suffered from ileal intussusception and recurrent gastrointestinal bleeding. The diagnosis was confirmed by surgery and histology. She is symptom-free after surgery. PMID- 10524651 TI - Treatment of mesenteric desmoid tumours with the anti-oestrogenic agent toremifene: case histories and an overview of the literature. AB - Desmoid tumours are histologically benign but due to their infiltration and compression of surrounding structures potentially life-threatening fibromatous lesions of unknown aetiology. The annual incidence rate is 2-4 per million people. The mesenteric variant constitutes about 10% of all desmoid tumours, although in familial adenomatous polyposis (FAP) patients this may be up to 70%. Due to the small number of patients with mesenteric desmoids the therapy is mainly empirical. This report describes the rationale as well as the value of the short- and long-term treatment (up to 6 years) with the anti-oestrogenic agent toremifene in combination with sulindac in two patients suffering from such a mesenteric desmoid tumour. These patients did not respond to sulindac alone and previous treatment with tamoxifen together with this non-steroidal anti inflammatory drug had also failed. An overview of the literature on the management of these dismal tumours is presented. PMID- 10524652 TI - The histopathology of coeliac disease: time for a standardized report scheme for pathologists. AB - In this paper, we review the histological features of coeliac disease and propose a standardized report scheme based on the Marsh classification. Furthermore, terms used by pathologists are defined. The most important histological differential diagnoses are given, as well as a definition of the different clinical forms of coeliac disease such as symptomatic, silent, latent, potential, treated and refractory coeliac disease. PMID- 10524653 TI - Spreading depression and trigeminovascular sensory neurons. PMID- 10524654 TI - Efficacy of 5HT in migraine. PMID- 10524655 TI - Great pains: famous people with headaches. AB - Have headaches influenced the course of history? It is very difficult to prove, but there is no doubt that head pains have affected some of the most influential people in history. This review explores how headaches have affected some of the world's most famous people. PMID- 10524656 TI - Effect of cortical spreading depression on activity of trigeminovascular sensory neurons. AB - The effect of cortical spreading depression, a proposed initiating event for migraine pain, on cortical blood flow (laser Doppler method) and on the spontaneous firing rate and stimulus-evoked responses of trigemino-cervical neurons with craniovascular input was studied in 17 neurons in 8 cats anesthetized with chloralose. Cortical spreading depression, induced via cortical pinprick injury, produced an initial wave of cortical hyperemia (243+/-57% of control) and a later and smaller phase of oligemia (96+/-4% of control). Neither the basal discharge rate (6.7+/-1.7 sec(-1)) nor the evoked responses to electrical stimulation of the superior sagittal sinus (4.1+/-0.8 discharges per stimulus) of upper cervical spinal cord neurons was altered over periods of up to 2 h following one, two, or three waves of spreading cortical depression. We conclude that a small number of episodes of cortical spreading depression is not capable of activating C2 cervical spinal cord craniovascular sensory neurons in the cat. PMID- 10524657 TI - Serotonin infusions inhibit sensory input from the dural vasculature. AB - Intravenous infusions of serotonin (5-hydroxtryptamine creatinine sulphate, 5HT, 50-300 microg/kg/min) in cats reversibly inhibited the responses of cervical spinal cord neurons to electrical stimulation of the superior sagittal sinus. Inhibition developed over 20-30 min and resolved over the same time course, suggesting a dependence on accumulation of 5HT in the central nervous system. Inhibition was suppressed by prior intravenous injection of the 5HT antagonists methysergide (1 mg/kg) and methiothepin (1 mg/kg). Infusions of 5HT (50 microg/kg/min) caused a rise in whole blood levels of 5HT by a factor of 1.5 of control values. 5HT levels in platelet-free plasma rose by a factor of 50. Levels of 5HT and 5 hydroxyindole acetic acid released into the cerebrospinal fluid rose significantly. The results suggest that earlier clinical observations that 5HT infusions can ameliorate the pain of migraine may not have been due to cranial vasoconstriction alone, but could have involved a central action of 5HT. PMID- 10524658 TI - Human isolated coronary artery contraction to sumatriptan: a post hoc analysis. AB - A post hoc analysis was performed on concentration response curves to sumatriptan in 62 human isolated coronary arteries. We determined whether donor-related clinical characteristics (age, sex, cause of death) and properties of the coronary artery (functional endothelial integrity, muscle mass) were related to the potency and efficacy of sumatriptan in contracting the human isolated coronary artery. The efficacy of sumatriptan was inversely related to the functional integrity of the vessel endothelium. Thus, contrary to expectation, coronary artery constriction to sumatriptan seems to be more pronounced in patients with nondiseased coronary arteries where the endothelium is intact. Nevertheless, in view of the high coronary reserve in these patients, myocardial ischemia after the use of sumatriptan is unlikely to occur, whereas in patients with coronary artery disease even a small contraction may be deleterious. PMID- 10524659 TI - Resolution of MRI abnormalities of the oculomotor nerve in childhood ophthalmoplegic migraine. AB - Ophthalmoplegic migraine is an uncommon disorder, usually starting in older childhood. Its physiopathology remains obscure and diagnosis is reliant on clinical grounds and exclusion of other disorders. We report four cases of childhood ophthalmoplegic migraine, one of them starting in infancy. Association with other types of migraine is common. Two of the three patients studied by magnetic resonance imaging (MRI) showed enhancement and enlargement of the cisternal portion of the oculomotor nerve, which spontaneously resolved after 2 and 4 years, respectively. Persistence of clinical recurrences was associated with long-lasting presence of the MRI finding, and possibly with mild sequelae. These radiological abnormalities suggest a common physiopathological mechanism with other inflammatory diseases, except for a benign evolution which, added to its specific anatomic site, seems to be the only neuroradiological marker, besides normality, in ophthalmoplegic migraine. The very long potential duration of MRI changes and the scarcity of clinical episodes make feasible its incident discovery once the migraine attack has become a remote memory. PMID- 10524660 TI - Glyceryl trinitrate induces attacks of migraine without aura in sufferers of migraine with aura. AB - Migraine with aura and migraine without aura have the same pain phase, thus indicating that migraine with aura and migraine without aura share a common pathway of nociception. In recent years, increasing evidence has suggested that the messenger molecule nitric oxide (NO) is involved in pain mechanisms of migraine without aura. In order to clarify whether the same is true for migraine with aura, in the present study we examined the headache response to intravenous infusion of glyceryl trinitrate (GTN) (0.5 microg/kg/min for 20 min) in 12 sufferers of migraine with aura. The specific aim was to elucidate whether an aura and/or an attack of migraine without aura could be induced. Fourteen healthy subjects served as controls. Aura symptoms were not elicited in any subject. Headache was more severe in migraineurs than in the controls during and immediately after GTN infusion (p=0.037) as well as during the following 11 h (p = 0.008). In the controls, the GTN-induced headache gradually disappeared, whereas in migraineurs peak headache intensity occurred at a mean time of 240 min post-infusion. At this time the induced headache in 6 of 12 migraineurs fulfilled the diagnostic criteria for migraine without aura of the International Headache Society. The results therefore suggest that NO is involved in the pain mechanisms of migraine with aura. Since cortical spreading depression has been shown to liberate NO in animals, this finding may help our understanding of the coupling between cortical spreading depression and headache in migraine with aura. PMID- 10524661 TI - Migraine polypharmacy and the tolerability of sumatriptan: a large-scale, prospective study. AB - Polypharmacy (the prescription of more than one therapy for a single patient) and subcutaneous (s.c.) sumatriptan tolerability were prospectively studied in 12,339 migraineurs, each followed for up to 1 year. Inclusion/exclusion criteria were minimal and mirrored United States Imitrex labeling. Drug usage and compliance monitoring were automatically interfaced with prescription refill. Concomitant drugs were used by 79% of patients, with analgesics, antidepressants, and sedatives used most commonly. No adverse interactions between sumatriptan and neurological drugs were found, possibly reflecting relative inability of the former to cross the blood-brain barrier. No difference in cardiovascular adverse events was associated with oral contraceptive use, which was more common than expected. No other drug class influenced adverse event probability, although sample sizes for these comparisons was sometimes <400 patients. This study confirms the prevalence of polypharmacy in migraine, identifies the drugs used, and concludes that, on a population basis, the tolerability of s.c. sumatriptan, when used according to labeled instructions, is unaffected by these concomitant drugs. PMID- 10524662 TI - Open-labeled long-term study of the efficacy, safety, and tolerability of subcutaneous sumatriptan in acute migraine treatment. AB - In a multicenter study, a long-term analysis was made of the efficacy, safety, and tolerability of subcutaneous (s.c.) sumatriptan in the acute treatment of migraine attacks over a period of up to 18 months. A total of 2263 patients took part in the study, all able to perform their own acute treatment of migraine attacks at home by s.c. administration of 6 mg of sumatriptan. A headache diary was used by each patient to record the various migraine parameters before the injection and 1 h and 2 h after it. A total of 43,691 attacks were treated and analyzed during the study period from October 1991 to June 1993. Therapy was successful in 89.5% of attacks. Freedom from headache was achieved in 71.0% of cases. In 22.7% of the attacks a second injection was administered on recurrence of the headache; 82.9% of the patients achieved an intraindividual therapy success rate ranging from over 80% to 100%. In the course of treatment there was no change in either the therapy success rate or in the frequency of attacks. Some 4.9% of the patients withdrew from the study because of insufficient efficacy or adverse events. A total of 44.5% of patients reported adverse events, and these were rated serious in the case of 1.7%. S.c. administration of sumatriptan for acute migraine therapy is an effective treatment method, with reliable action, that can be used with good tolerability provided the contraindications are taken into account. PMID- 10524663 TI - Treatment of severe, disabling migraine attacks in an over-the-counter population of migraine sufferers: results from three randomized, placebo-controlled studies of the combination of acetaminophen, aspirin, and caffeine. AB - OBJECTIVE: To examine the benefits of acetaminophen, aspirin, and caffeine (AAC) in the treatment of severe, disabling migraine attacks, in a population of migraine sufferers for whom over-the-counter (OTC) medications are appropriate. BACKGROUND: Subjects (n = 1220) who met the International Headache Society criteria for migraine with or without aura were included in three independent clinical studies. DESIGN/METHODS: Post-hoc analysis of 172 subjects who met the criteria for severe, disabling migraine reported a history of migraine attacks characterized by at least severe pain and severe disability, and treated attacks with severe pain and at least severe disability. Subjects who usually vomited with 20% or more of their migraine attacks, and those with incapacitating disability (subjects who required bed rest for more than 50% of their attacks) were not eligible for enrollment. RESULTS: From 1 h and continuing through 6 h postdose, the proportion of responders was significantly greater (p< or =0.01) for AAC than placebo. The pain intensity difference from baseline was significantly greater (p< or =0.05) for AAC than placebo from 0.5 h through 6 h. The proportion of subjects reporting improvement in functional disability, photophobia, and phonophobia was significantly greater for AAC than placebo from 2 h through 6 h postdose. CONCLUSIONS: The nonprescription combination of AAC was well tolerated and effective. PMID- 10524664 TI - Can postoperative infection be prevented? PMID- 10524665 TI - Transvaginal ultrasound-guided aspiration of pelvic abscesses. AB - OBJECTIVE: To assess the utility of a less invasive approach to the care of women with a pelvic abscess, we retrospectively reviewed the outcome of women with pelvic abscesses managed by transvaginal ultrasound-guided aspiration. METHODS: A retrospective analysis of 27 pelvic abscesses in 22 consecutive women undergoing transvaginal drainage, including 13 tuboovarian abscesses (TOAs) and 14 postoperative abscesses (POAs). All patients received broad-spectrum intravenous antibiotics from the time infection was diagnosed to resolution of signs and symptoms. Chart review and examination of ultrasound files were utilized to extract demographic clinical, laboratory, and outcome data. RESULTS: The mean age for the study group was 30 years old. Mean duration from diagnosis to drainage was 5.6 days (TOA) and 2.0 days (POA), P < 0.01. The mean diameter of the abscesses was 86 mm. The volume of purulent material drained ranged from 70-750 mL. Perceived adequacy of drainage was correlated with lack of abscess septation. Cultures for aerobic and anaerobic pathogens were positive in 51% of cases (79% POA versus 23% TOA, P < 0.05) with 1.9 organisms/ positive culture. Transvaginal drainage was successful in 25 of 27 abscesses. No complications were reported. CONCLUSION: In skilled hands, transvaginal guided aspiration of pelvic abscess is a highly successful technique with minimal risk to the patient. Follow-up studies are needed to assess the long-term sequelae, such as frequency of infertility, ectopic pregnancy, and chronic pelvic pain. PMID- 10524666 TI - Germ tube formation changes surface hydrophobicity of Candida cells. AB - Hydrophobic interaction is generally considered to play an important role in the adherence of microorganisms to eukaryotic cells and also to certain inert surfaces. Using a microbe adhesion assay to hydrocarbons (n-hexadecane), 68 strains of Candida albicans and 30 non-albicans strains were studied. Influence of source of isolate, age of the culture, and percentage of germ tube formation on adhesion were studied. C. albicans blastoconidia were found to be hydrophilic; conversely, blastoconidia of non-albicans strains were slightly more hydrophobic. Germ tube formation was associated with a significant rise in cell surface hydrophobicity. PMID- 10524667 TI - Semiquantitative bacterial observations with group B streptococcal vulvovaginitis. AB - OBJECTIVE: Group B streptococcal (GBS) vulvovaginitis is a poorly-delineated clinical entity. The purpose of this study is to report semiquantitative data from four cases of GBS vulvovaginitis and to comment on their significance in terms of the in vitro inhibitory capabilities of GBS. METHODOLOGY: Four patients whose clinical presentations were consistent with GBS vulvovaginitis, from whom GBS was isolated and for whom semi-quantitative as well as qualitative microbiologic data existed, were identified. RESULTS: To produce vulvovaginitis, GBS must be at a high multiplicity (10(8) CFU/g of vaginal fluid). Single coisolates were identified in three of the four cases (two cases of Escherichia coli and one case of Staphylococcus aureus). Group B streptococcus does not inhibit either of these bacteria in vitro. CONCLUSION: When the growth requirements for the demonstration of in vitro inhibition for GBS or lack thereof are met in vivo, the in vivo observations are consistent with those projected from the in vitro data. PMID- 10524668 TI - Do antepartum herpes simplex virus cultures predict intrapartum shedding for pregnant women with recurrent disease? AB - OBJECTIVE: To examine antenatal screening as a predictor of intrapartum shedding of herpes simplex virus (HSV) and to determine its usefulness in guiding the appropriate route of delivery for patients with recurrent HSV in pregnancy. METHODS: A population of 198 pregnant women with a history of recurrent genital HSV were cultured in the last weeks of their pregnancy by specially-trained personnel and intrapartum by their delivering attendants. RESULTS: Of cultures from a total of 906 antenatal visits, 17% were culture positive, with an asymptomatic shedding rate of 3.4%. Asymptomatic shedding occurred in 12.6% of women. Over the 8-week antepartum period, viral culture-positivity rates for each visit ranged from 11% to 19.5%. This provided an expected delivery culture positivity rate of 15.3%. However, actual intrapartum viral culture positivity occurred in only three of 191 women (1.5%; P < 0.001). Because previous studies have suggested antepartum culture positivity fails to predict intrapartum viral shedding, evaluations, including cultures, as well as predictive values for subsequent culture positivities, were determined under the supervision of an infectious disease specialist. Under these conditions, positive predictive values were 59% when the interval between visits was 2 days, but only 19% when days between visits were >2 (P < 0.0001). No cases of neonatal herpes were seen in this population, although cesarean deliveries were performed in 31% of the patient population, with genital herpes as the indication for 56% of those. CONCLUSIONS: Antepartum serial screening by viral culture is not predictive of an infant's risk of intrapartum viral exposure when conducted at weekly intervals. However, more frequent assessments of patients can be predictive of an infant's exposure risk to HSV; for patients with frequent recurrent disease near term or primary infection in pregnancy, frequent late antepartum screening may be appropriate. PMID- 10524669 TI - Human papillomavirus cervical infection and associated risk factors in a region of Argentina with a high incidence of cervical carcinoma. AB - OBJECTIVE: To assess the prevalence and potential risk factors associated with human papillomavirus (HPV) cervical infection among women residing in a region of northeastern Argentina with a high incidence of cervical cancer. METHODS: A case control study of 330 women participating in a cervical cytological screening program conducted in Posadas city, Misiones, Argentina, from February 1997 to November 1998 was carried out. Standardized questionnaires were administered, and clinical examination including colposcopy was performed. Fresh endocervical specimens for HPV DNA detection by generic polymerase chain reaction were collected and the products typed by dot-blot hybridization. RESULTS: Human papillomavirus DNA was found in 61% of samples analyzed (185/301). Samples with normal cytology had a 43% infection rate (85/199), while those classified as low grade squamous intraepithelial lesion, high-grade squamous intraepithelial lesion, and invasive cervical carcinoma had an infection rate of 96% (53/55), 100% (29/29), and 100% (18/18), respectively. Human papillomavirus typing showed a 64% (118/185) prevalence of type 16 among all the infected population analyzed; type 16 was detected among 49% (42/85) of infected samples with normal cytology and in an average of 74% (74/100) with abnormal cytology. Sexual behavior, residence in southern Paraguay, and history of a previous sexually transmitted diseases were the main risk factors associated with high-grade cervical lesions. CONCLUSIONS: An elevated prevalence of HPV infection was detected in this population, which also has a high incidence of cervical cancer. The broad distribution of high-risk HPV type 16 in women with normal cytology and colposcopy suggests that viral infection is an important determinant of regional cancer incidence. PMID- 10524670 TI - Peritoneal tuberculosis: diagnostic options. AB - BACKGROUND: Extrapulmonary tuberculosis has vague symptoms and few signs. It is essential to recognize and diagnose this curable disease prior to performing definitive surgery. Newer tests such as DNA or RNA amplification allow for early diagnosis but have limitations. CASE: We report a case of peritoneal tuberculosis in an immigrant woman. She had vague symptoms of low-grade fever, mild abdominal pain, obstipation, and bloating. Diagnostic laparoscopy was performed to establish the diagnosis. Tuberculosis was confirmed by DNA extraction from the frozen section specimen with subsequent analysis using polymerase chain reaction. CONCLUSION: Peritoneal tuberculosis is a disease that often simulates malignancies. With the increasing prevalence of human immunodeficiency virus in developed countries, tuberculosis is also on the rise and should be considered in the differential diagnosis of a patient with an abdominal/pelvic mass and ascites. PMID- 10524671 TI - Pelvic inflammatory disease in the postmenopausal woman. AB - OBJECTIVE: Review available literature on pelvic inflammatory disease in postmenopausal women. DESIGN: MEDLINE literature review from 1966 to 1999. RESULTS: Pelvic inflammatory disease is uncommon in postmenopausal women. It is polymicrobial, often is concurrent with tuboovarian abscess formation, and is often associated with other diagnoses. CONCLUSION: Postmenopausal women with pelvic inflammatory disease are best treated with inpatient parenteral antimicrobials and appropriate imaging studies. Failure to respond to antibiotics should yield a low threshold for surgery, and consideration of alternative diagnoses should be entertained. PMID- 10524672 TI - Complement deficiency states, disease susceptibility, and infection risk in systemic lupus erythematosus. PMID- 10524673 TI - Neuroimaging in neuropsychiatric systemic lupus erythematosus. PMID- 10524674 TI - Low secretion of tumor necrosis factor alpha, but no other Th1 or Th2 cytokines, by peripheral blood mononuclear cells correlates with chronicity in reactive arthritis. AB - OBJECTIVE: To determine Th1 and Th2 cytokine production in patients with reactive arthritis (ReA) in relation to disease outcome and in comparison with rheumatoid arthritis (RA). METHODS: Secretion of tumor necrosis factor alpha (TNFalpha), interferon-gamma, interleukin-10 (IL-10), and IL-4 by peripheral blood mononuclear cells (PBMC) from 53 patients with early ReA (disease duration <8 weeks, 64% HLA-B27 positive) and 30 patients with early, untreated RA (disease duration <6 months) was determined by enzyme-linked immunosorbent assay (ELISA) after ex vivo stimulation. Intracellular cytokine staining with quantification of positive T cells by fluorescence-activated cell sorting (FACS) was performed in 12 ReA patients and 12 RA patients. In 27 ReA patients, cytokine secretion was measured again after 3 months. Patients were followed up for 1 year, and cytokine patterns were correlated with disease duration. RESULTS: TNFalpha secreted by whole PBMC and by T cells was significantly lower, by ELISA and by FACS, in ReA patients than in RA patients, while no significant differences were detected for the other cytokines. ReA patients with a disease duration of > or =6 months showed significantly lower TNFalpha secretion than patients with a disease duration of <6 months (mean +/- SD 385 +/- 207 pg/ml versus 684 +/- 277 pg/ml; P = 0.003). Furthermore, low TNFalpha secretion after 3 months also correlated significantly with a more chronic course of disease. HLA-B27 positive patients secreted less TNFalpha than did those who were B27 negative (338 +/- 214 pg/ml versus 512 +/- 207 pg/ml; P = 0.05), and patients with a more chronic course had a higher frequency of B27 positivity (47% versus 80%; P = 0.01). Among the 27 HLA B27 positive patients, TNFalpha secretion in those with a disease duration of > or = 6 months was lower than that in the 7 with a disease duration of <6 months (308 +/- 167 pg/ml versus 562 +/- 308 pg/ml; P = 0.04). CONCLUSION: Low TNFalpha secretion and HLA-B27 status correlate with longer disease duration in ReA patients, possibly with an additive effect. The diminished TNFalpha production might reflect a state of relative immunodeficiency contributing to bacterial persistence in ReA. PMID- 10524675 TI - Modulation of peripheral blood mononuclear cell activation status during Salmonella-triggered reactive arthritis. AB - OBJECTIVE: To determine the activation status of mononuclear cells in the peripheral circulation during the acute phase and the recovery phase of Salmonella-triggered reactive arthritis (ReA). METHODS: Peripheral blood mononuclear cells (PBMC) were obtained from 8 patients with Salmonella infection (4 with ReA and 4 without) and were studied by reverse transcription-polymerase chain reaction for messenger RNA (mRNA) of proinflammatory and antiinflammatory cytokines, by flow cytometry (FC) for cell surface activation and adhesion molecules, by immunofluorescence (IF) microscopy for bacterial antigens, and by FC, IF, and DNA fragmentation on gel for signs of apoptosis. RESULTS: During the acute phase of the infection, PBMC were activated in all patients, as characterized by high levels of expression of CD14, CD11b, and CD11c on monocytes. In the patients with ReA, PBMC also had the capacity to produce interleukin-1beta (IL-1beta), IL-6, IL-8, IL-10, and tumor necrosis factor alpha. During the amelioration of disease, monocyte activation was decreased in all patients. A complete down-regulation of CD14 was detected only in the patients with ReA, whereas the expression of CD14 in the patients without ReA was positive and was similar to that in healthy controls. In addition, cytokine mRNA levels decreased regardless of the presence of Salmonella antigens in blood cells in all 4 patients with ReA. CONCLUSION: High levels of expression of some activation and adhesion molecules and elevated levels of mRNA for certain cytokines that are predominantly produced by monocytes were found in PBMC from patients with acute Salmonella-triggered ReA, which suggests that these cells are activated. On the other hand, complete down-regulation of CD14 and a marked decrease in the cytokine production capacity during amelioration of the disease suggest that suppression of PBMC activity might be involved in recovery from ReA. PMID- 10524676 TI - V2 regions of 16S ribosomal RNA used as a molecular marker for the species identification of streptococci in peripheral blood and synovial fluid from patients with psoriatic arthritis. AB - OBJECTIVE: To detect the 16S ribosomal RNA (rRNA) of 3 streptococcal species in the peripheral blood and synovial fluid of patients with psoriatic arthritis (PsA). METHODS: Reverse transcription-polymerase chain reaction (RT-PCR) detection targets bacterial 16S rRNA, which is present in bacteria at high copy numbers. The 3 species-specific primers for group A streptococci (GAS; Streptococcus pyogenes), group B streptococci (GBS; Streptococcus agalactiae), and Streptococcus pneumoniae were designed from the fragments of highly variable V2 regions of 16S rRNA. Total RNA was prepared from whole peripheral blood and joint fluid obtained from patients with PsA and rheumatoid arthritis (RA). All positive PCR reactions were then sequenced with a Pharmacia ALF DNA sequencing system. RESULTS: Our data in 19 PsA patients showed that 7 peripheral blood samples were positive for GAS (P = 0.006 versus GAS-positive RA patients [n = 0], by Fisher's exact test), and 2 were also positive for GBS. One synovial fluid sample from a PsA patient was positive for GAS. S pneumoniae was absent from all specimens. Seventeen patients with RA were PCR negative for the 3 streptococcal species. Peripheral blood from a patient with inflammatory bowel disease was positive for GAS. CONCLUSION: The presence of GAS 16S rRNA in the peripheral blood and synovial fluid of patients with PsA supports the concept that PsA is a reactive arthritis to certain streptococci. PMID- 10524677 TI - A new spin on an old model: in vivo evaluation of disease progression by magnetic resonance imaging with respect to standard inflammatory parameters and histopathology in the adjuvant arthritic rat. AB - OBJECTIVE: To noninvasively examine the pathogenesis of rat adjuvant-induced arthritis (AIA) by magnetic resonance imaging (MRI), and to correlate MRI indices of disease progression with classic inflammatory parameters and histologic evaluation. METHODS: AIA was established in male Lewis rats following subcutaneous injection in the right hindpaw with 0.5 mg of heat-killed Mycobacterium butyricum suspended in light mineral oil. In vivo MRI evaluations of soft tissue and bony changes in AIA rats with matched histopathology were correlated with changes in left hindpaw volumes, circulating leukocytes, acute phase reactants, and urinary collagen crosslinks throughout the disease process. RESULTS: MRI of arthritic tibiotarsal joints of the uninjected left hindpaws from AIA rats demonstrated 2 distinct phases of disease activity. The first phase, apparent between days 10 and 18, was characterized by periarticular inflammation with marked synovitis, synovial fibroplasia, and distension of the joint capsule into the surrounding tissue. The secondary phase, occurring between days 18 and 30, was marked by continued soft tissue inflammation, periostitis with osteolysis, and periosteal new bone formation progressing to a state of near complete ankylosis by day 30. These 2 phases of disease activity observed by MRI paralleled biochemical, cellular, and histologic markers of disease progression. CONCLUSION: MRI can be used to noninvasively detect, monitor, and quantify the chronic synovitis and progressive destruction of soft tissue and bone in live AIA rats, thereby improving the ability to evaluate disease progression in this preclinical animal model of rheumatoid arthritis. PMID- 10524678 TI - Cleavage of aggrecan at the Asn341-Phe342 site coincides with the initiation of collagen damage in murine antigen-induced arthritis: a pivotal role for stromelysin 1 in matrix metalloproteinase activity. AB - OBJECTIVE: The destruction of articular cartilage during arthritis is due to proteolytic cleavage of the extracellular matrix components. This study investigates the kinetic involvement of metalloproteinases (MMPs) in the degradation of the 2 major cartilage components, aggrecan and type II collagen, during murine antigen-induced arthritis (AIA). In addition, the role of stromelysin 1 (SLN-1) induction of MMP-induced neoepitopes was studied. METHODS: VDIPEN neoepitopes in aggrecan and collagenase-induced COL2-3/4C neoepitopes in type II collagen were identified by immunolocalization. Stromelysin 1-deficient knockout (SLN1-KO) mice were used to study SLN-1 involvement. RESULTS: In AIA, the VDIPEN epitopes in aggrecan appeared after initial proteoglycan (PG) depletion. The collagenase-induced type II collagen neoepitopes colocalized with VDIPEN epitopes. Remarkably, cartilage from arthritic SLN1-KO mice showed neither the induction of VDIPEN nor collagen cleavage-site neoepitopes during AIA, suggesting that stromelysin is a pivotal mediator in this process. PG depletion, as measured by the loss of Safranin O staining, was similar in SLN1-KO mice and wild-type strains. Furthermore, in vitro induction of VDIPEN epitopes in aggrecan and COL2-3/4C epitopes in type II collagen, on exposure of cartilage to interleukin-1, could not be accomplished in SLN1-KO mice, whereas intense staining was achieved for both epitopes in cartilage of wild-type strains. CONCLUSION: This study emphasizes that SLN-1 is essential in the induction of MMP specific aggrecan and collagen cleavage sites during AIA. It suggests that SLN-1 is not a dominant enzyme in PG breakdown, but that it activates procollagenases and is crucial in the initiation of collagen damage. PMID- 10524679 TI - Enhanced T cell proliferative response to type II collagen and synthetic peptide CII (255-274) in patients with rheumatoid arthritis. AB - OBJECTIVE: To determine the presence of specific immune recognition of type II collagen (CII) and its immunodominant epitope CII (255-274) in patients with rheumatoid arthritis (RA). METHODS: T cell proliferative responses to bovine CII and a synthetic peptide encompassing CII (255-274) in peripheral blood mononuclear cells (PBMC) and synovial fluid mononuclear cells (SFMC) from RA patients, and in PBMC from osteoarthritis (OA) patients and healthy controls were assayed by mixed lymphocyte culture. RESULTS: The stimulation index (SI) and the number of positive (SI > or = 2) T cell responses to CII were higher in RA patients (n = 106) than in OA patients (n = 26) and healthy controls (n = 34). T cell responses to CII (255-274) were also enhanced in RA patients and correlated well with those to CII. In SFMC, positive responses to CII or CII (255-274) were detected in 61.9% of 42 RA patients. T cell responses to CII in SFMC were stronger and more prevalent than peripheral responses. The SI and positive responses to CII were higher in early RA than in late RA. Levels of IgG antibodies to CII in synovial fluid inversely correlated with T cell responses to CII. CONCLUSION: T cell responses to CII or CII (255-274) were enhanced in RA, especially in early disease. Synthetic peptide CII (255-274), as well as native CII, could be recognized as immunogenic antigens by T cells, particularly in the synovial fluid. These observations suggest that CII-reactive T cells play an important role in the pathogenesis of RA. Peripheral tolerance induction using CII (255-274) might be useful in the treatment of RA. PMID- 10524681 TI - Estrogen replacement therapy modulation of the insulin-like growth factor system in monkey knee joints. AB - OBJECTIVE: Epidemiologic studies have suggested that estrogen replacement therapy may lower the risk of osteoarthritis in women, but the mechanism of this effect is unknown. Since estrogen acts in other tissues in part through regulation of the insulin-like growth factor (IGF) system as well as cytokines including interleukin-6 (IL-6), we determined whether estrogen replacement regulates the levels of these factors in synovial fluid (SF). METHODS: Levels of IGF-1, IGF-2, IGF binding proteins (IGFBP) 1-3, and IL-6 were measured in SF samples obtained from 67 female adult cynomolgus monkeys that had been ovariectomized and treated for 30 months in 1 of 3 groups. Group 1 (n = 24) had no estrogen replacement (control), group 2 (n = 22) received estrogen (Premarin) at the human equivalent of 0.625 mg/day, and group 3 (n = 21) received estrogen at the same dose as group 2, plus progesterone (Provera) at the equivalent of 2.5 mg/day. RESULTS: Compared with controls, estrogen-treated monkeys had 2-fold higher SF levels of IGF-1 (P < 0.001), 1.7-fold higher IGF-2 (P < 0.006), 5.9-fold higher IGFBP-1 (P < 0.02), and 2.5-fold higher IGFBP-3 (P < 0.001). Estrogen plus progesterone-treated monkeys had SF levels of IGF-1, IGF-2, IGFBP-1, and IGFBP-3 that were intermediate between the levels in the control and estrogen groups, except that the level of IGFBP-3 was significantly greater than that in the control group (P < 0.001). SF levels of IGFBP-2 and IL-6 did not differ by treatment group. Treatment group did not affect the serum levels of IGF-1 and IL-6, but IGF-2 and IGFBP-3 were increased by 1.6- and 1.8-fold, respectively, in the estrogen group (P < 0.001). There was no correlation between changes in serum and SF levels of IGF components, except for a weak correlation for IGFBP-3 levels from control (r = 0.464, P = 0.04) and estrogen-treated (r = 0.577, P = 0.008) animals. CONCLUSION: This study demonstrates a significant effect of estrogen replacement on IGF system components in synovial fluid, of which at least some are distinct from any systemic changes observed. The results indicate a potential stimulatory effect of estrogen on joint tissues in vivo. PMID- 10524680 TI - Cartilage protection by nitric oxide synthase inhibitors after intraarticular injection of interleukin-1beta in rats. AB - OBJECTIVE: To evaluate the effect of nitric oxide synthase (NOS) inhibitors on proteoglycan synthesis following intraarticular administration of interleukin 1beta (IL-1beta) in rats. METHODS: Recombinant human IL-1beta and NOS inhibitors with different selectivity for inducible NOS (N-monomethyl-L-arginine [L-NMA], N iminoethyl-L-ornithine [L-NIO], and S-methylisothiourea [SMT]) were simultaneously administered in rats by a single intraarticular injection in each knee. L-NMA was also infused for 72 hours using an Alzet mini osmotic pump implanted into the peritoneal cavity 24 hours before IL-1beta challenge. NO production was determined as nitrate and nitrite, either in synovial fluid or ex vivo in supernatants of synovium and patellae. Proteoglycan synthesis was measured by ex vivo incorporation of 35SO4(2-) into patellar cartilage. RESULTS: IL-1beta induced a time-dependent increase in NO production in synovial fluid. Synovium and patellae released large amounts of nitrate and nitrite under ex vivo conditions, indicating that both tissues are effective sources of NO within the joint. This production of NO was accompanied by a delayed inhibition of proteoglycan synthesis. The intraarticular administration of L-NMA and L-NIO reduced NO release in synovial fluid and resulted in a partial recovery of proteoglycan synthesis. Under our experimental conditions, SMT failed to reduce NO synthesis and to restore proteoglycan synthesis. The protection of cartilage was improved by the systemic and sustained delivery of L-NMA. However, the complete inhibition of NO production in synovial fluid was not sufficient to fully restore cartilage anabolism. CONCLUSION: Our findings show that in rats: 1) NO may be an early mediator of the effect of IL-1beta on cartilage, 2) NO inhibition may have therapeutic relevance, although it is not sufficient to fully reverse the deleterious effects of IL-1beta, 3) among NOS inhibitors tested, only amino acid derivatives are effective, 4) protection can be achieved by local administration of NOS inhibitors, and 5) systemic and sustained delivery of the NOS inhibitor with the highest efficacy after intraarticular injection provides the most benefit. PMID- 10524682 TI - Abnormal regulation of urokinase plasminogen activator by insulin-like growth factor 1 in human osteoarthritic subchondral osteoblasts. AB - OBJECTIVE: Subchondral bone sclerosis is a common feature of osteoarthritis (OA), but the mechanisms responsible for this condition remain unresolved. We investigated the role of insulin-like growth factor 1 (IGF-1) and urokinase plasminogen activator (uPA) in human osteoblasts from subchondral bone obtained from the tibial plateaus of OA patients and normal individuals. METHODS: Primary in vitro osteoblasts were prepared from subchondral bone specimens obtained from OA patients at surgery and from normal individuals at autopsy. Levels of uPA and PA inhibitor 1 (PAI-1) levels were determined under basal conditions and after IGF-1 stimulation in conditioned media from osteoblasts by enzyme-linked immunosorbent assay. The activity of uPA was evaluated by specific substrate hydrolysis and zymography under basal conditions and after plasminogen stimulation, in the presence and absence of added IGF-1. Plasmin activity was also evaluated by specific substrate hydrolysis. RESULTS: Levels of uPA released by OA osteoblasts were significantly higher than normal. Addition of IGF-1 to osteoblasts significantly reduced uPA protein levels only in OA patients (P < 0.05). In contrast, the addition of uPA to osteoblasts did not modify IGF-1 levels in either normal or OA osteoblasts. Basal uPA activity was higher in OA than in normal osteoblasts. Interestingly, IGF-1 enhanced basal uPA activity in OA specimens in a dose-dependent manner. Addition of plasminogen promoted uPA activity in both normal and OA osteoblasts via a positive feedback loop due to plasmin generation, since this activity was inhibited by both PAI-1 and alpha2 antiplasmin. Unexpectedly, incubation with IGF-1 inhibited this positive feedback of plasminogen-dependent uPA activity in OA osteoblasts, but not in normal osteoblasts, in a dose-dependent manner. Hence, normal osteoblasts were relatively insensitive to IGF-1, whereas the same treatment reduced both uPA levels and plasminogen-dependent uPA activity in OA osteoblasts while it increased basal uPA activity in OA osteoblasts. This could not be explained by PAI-1 protein levels, which were similar in normal and OA osteoblasts in the presence and absence of IGF-1. IGF-1 also reduced plasmin activity in OA osteoblasts while it did not modify this activity in normal osteoblasts. CONCLUSION: These results suggest that in OA osteoblasts, the uPA/plasmin system functions normally, yet IGF-1 inhibits the positive feedback of plasmin on uPA activity. This inhibition may contribute to abnormal IGF-1- and uPA-dependent bone remodeling, ultimately leading to abnormal bone sclerosis in OA. PMID- 10524683 TI - Nitric oxide production and apoptosis in cells of the meniscus during experimental osteoarthritis. AB - OBJECTIVE: To examine the pathologic changes in meniscus tissue during experimental osteoarthritis (OA) and to determine the relationship between nitric oxide (NO) synthesis, apoptosis, and meniscus degradation. METHODS: OA was induced in rabbits by anterior cruciate ligament (ACL) transection. Knees were harvested after 9 weeks and assessed for OA severity. Menisci were subjected to histologic, immunohistochemical, and electron microscopic analyses for the presence of nitrotyrosine and apoptosis. Menisci were also cultured for analysis of NO production. RESULTS: All menisci from joints with ACL transection demonstrated degenerative changes. A high number of apoptotic cells was present in the medial part of menisci, which contains chondrocytic cells. Menisci from nonoperated contralateral knees contained only small numbers of cells in apoptosis. Conditioned media from meniscus cultures contained similarly elevated levels of nitrite as cartilage cultures from the same arthritic knees. Nitrotyrosine immunoreactivity, an indicator of in vivo NO production, was prominent in menisci from knees with ACL transection. In addition, menisci from normal knees produced high levels of NO in response to in vitro stimulation with interleukin-1beta or lipopolysaccharide. CONCLUSION: These observations suggest that pathologic changes in menisci are a regular feature of experimentally induced OA and are associated with NO production and meniscus cell apoptosis. PMID- 10524684 TI - An IgG antiprothrombin antibody enhances prothrombin binding to damaged endothelial cells and shortens plasma coagulation times. AB - OBJECTIVE: To test the hypothesis that some lupus anticoagulants are antiprothrombin antibodies, and that such antibodies enhance prothrombin binding to endothelial cells (EC) and thus promote clotting on the cell surface. METHODS: We generated a monoclonal antiprothrombin antibody (designated IS6) from a patient with primary antiphospholipid syndrome (APS). The antibody was analyzed for its binding properties, lupus anticoagulant activity, and pathophysiologic activity, using an EC-based plasma coagulation assay. RESULTS: IS6 is the first patient-derived monoclonal IgG antiprothrombin antibody. It bound to prothrombin with low affinity, reacted with 3 phospholipids (cardiolipin, phosphatidylethanolamine, and phosphatidylserine), and showed lupus anticoagulant activity. Moreover, IS6 enhanced the binding of prothrombin to damaged EC and shortened the EC-based plasma coagulation times. CONCLUSION: These findings suggest that IS6 may promote coagulation in areas of damaged EC in the host, and thus contribute to thrombosis in patients with APS. PMID- 10524685 TI - Exclusion of the gene for human cartilage intermediate layer protein in currently mapped calcium pyrophosphate dihydrate deposition syndromes. AB - OBJECTIVE: To map the gene for human cartilage intermediate layer protein (CILP) in order to assess its involvement in some familial forms of calcium pyrophosphate dihydrate (CPPD) deposition disease. METHODS: A radiation hybrid panel was analyzed for chromosomal assignment of the CILP gene within a 1-cM limit of resolution. The location of the gene for CILP was confirmed to reside at the observed radiation hybrid locus by fluorescence in situ hybridization. RESULTS: The human CILP gene resides at chromosome 15q21. CONCLUSION: This map location definitively excludes mutations in the CILP gene as the cause of certain familial forms of CPPD deposition disease that have been genetically mapped to chromosomes 8q and 5p. PMID- 10524686 TI - Mannose-binding lectin polymorphisms and susceptibility to infection in systemic lupus erythematosus. AB - OBJECTIVE: To determine whether variant alleles in the coding portion of the mannose-binding lectin (MBL) gene are associated with increased susceptibility to systemic lupus erythematosus (SLE) and concomitant infections. METHODS: MBL alleles and serum concentrations were determined by polymerase chain reaction and enzyme-linked immunosorbent assay, respectively, in 91 Danish patients with SLE and in 250 controls. RESULTS: Homozygosity for MBL variant alleles was observed in 7.7% of the SLE patients compared with 2.8% of the controls (P = 0.06), while no difference was seen for heterozygosity (33.0% versus 34.4%). Homozygotes had an increased risk of acquiring serious infections compared with patients who were heterozygous or homozygous for the normal allele (odds ratio 8.6, 95% confidence interval 1.5-47.6, P = 0.01). The time interval from the diagnosis of SLE to the first infectious event was shorter (P = 0.017), and the annual number of infectious events was 4 times higher, among homozygotes (P = 0.00002). They were especially prone to acquire pneumonia (P = 0.00004). CONCLUSION; Homozygosity for MBL variant alleles may explain much of the increased risk of complicating infections seen in SLE patients. Additionally, it is a minor risk factor for acquiring SLE. PMID- 10524687 TI - The Prosorba column for treatment of refractory rheumatoid arthritis: a randomized, double-blind, sham-controlled trial. AB - OBJECTIVE: To evaluate the efficacy and safety of the Prosorba column as a treatment for rheumatoid arthritis (RA) in patients with active and treatment resistant (refractory) disease. METHODS: A sham-controlled, randomized, double blind, multicenter trial of Prosorba versus sham apheresis was performed in patients with RA who had failed to respond to treatment with methotrexate or at least 2 other second-line drugs. Patients received 12 weekly treatments with Prosorba or sham apheresis, with efficacy evaluated 7-8 weeks after treatment ended. Patients were characterized as responders if they experienced improvement according to the American College of Rheumatology (ACR) response criteria at the efficacy time point. A data safety monitoring board (DSMB) evaluated interim analyses for the possibility of early completion of the trial. RESULTS: Patients in the trial had RA for an average of 15.5 years (range 1.7-50.6) and had failed an average of 4.2 second-line drug treatments prior to entry. After the completion of treatment of 91 randomized patients, the DSMB stopped the trial early due to successful outcomes. Of the 47 patients in the Prosorba arm, 31.9% experienced ACR-defined improvement versus 11.4% of the 44 patients in the sham treated arm (P = 0.019 after adjustment for interim analysis). When results from 8 additional patients, who had completed blinded treatments at the time of DSMB action, were added to the analysis (n = 99), results were unchanged. The most common adverse events were a short-term flare in joint pain and swelling following treatment, a side effect that occurred in most subjects at least once in both treatment arms. Other side effects, although common, occurred equally as frequently in both treatment groups. CONCLUSION: Apheresis with the Prosorba column is an efficacious treatment for RA in patients with active disease who have failed other treatments. PMID- 10524688 TI - Comparison of two schedules for administering oral low-dose methotrexate (weekly versus every-other-week) in patients with rheumatoid arthritis in remission: a twenty-four week, single blind, randomized study. AB - OBJECTIVE: To compare the efficacy of 2 low-dose oral methotrexate (MTX) schedules in maintaining remission in patients with rheumatoid arthritis (RA). METHODS: Patients with RA were included if they were receiving treatment with weekly MTX for at least 9 months and the RA was in remission (defined by American College of Rheumatology [ACR] criteria) for at least 6 months. Patients were stratified by treatment and randomly assigned to weekly or every-other-weekly (EOW; reducing their monthly dose by half) treatment with MTX. Patients were evaluated by a rheumatologist (blinded to the treatment schedule) at baseline and at 6, 12, and 24 weeks. The evaluations included joint counts, Ritchie Articular Index, Health Assessment Questionnaire Disability Index, physician's and patient's global health assessments, visual analog scale for pain, and incidence of adverse effects. Laboratory evaluations were done at baseline and at week 24. RESULTS: Fifty-one patients were included (26 taking weekly MTX, 25 taking EOW MTX). Baseline comparisons showed no differences between the groups. The mean duration of RA was <3 years in both groups, and they had been started on weekly MTX treatment early after diagnosis. After 24 weeks, >90% of the patients in both groups continued in remission. Evaluations of disease activity at 6 and 12 weeks showed no between-group differences. EOW MTX patients who experienced relapse were switched back to weekly MTX, and after a few weeks, their RA was again controlled. The incidence of adverse effects was slightly higher in the weekly MTX group, although the difference did not reach statistical significance. The observed laboratory values were very similar for both groups, except for the serum aspartate aminotransferase and alanine aminotransferase levels, which decreased in the EOW MTX group and were statistically significant at week 24 (P = 0.04 and P = 0.006, respectively). CONCLUSION: EOW MTX represents a valid therapeutic alternative for a specific subgroup of RA patients, as outlined by the ACR remission criteria. Patients with a short disease duration who were treated early after disease onset with weekly MTX and who achieve sustained remission have a higher probability of success with the EOW MTX schedule. PMID- 10524689 TI - Treatment with monoclonal anti-tumor necrosis factor alpha antibody results in an accumulation of Th1 CD4+ T cells in the peripheral blood of patients with rheumatoid arthritis. AB - OBJECTIVE: In rheumatoid arthritis (RA), treatment with tumor necrosis factor alpha (TNFalpha) binding agents has proven to be highly effective. Downregulation of the proinflammatory cytokine cascade and a reduced migration of leukocytes into the joints have been proposed as modes of action of TNFalpha blockade. We investigated whether alterations in the number of circulating pro- and antiinflammatory T cell subsets contribute to the therapeutic effect of monoclonal antibodies (mAb) against TNFalpha in RA patients. METHODS: Phenotypic analysis of peripheral blood T cell subsets was performed on blood from RA patients before and after treatment with an anti-TNFalpha mAb. RESULTS: An accumulation of primed CD45RA- T cells of both the CD4+ and the CD8+ T cell population was seen shortly after treatment. Most notably, within the CD4+,CD45RA T cell subset, the number of interferon-gamma-producing T cells was significantly increased after anti-TNFalpha mAb treatment, resulting in a significant rise in the Th1:Th2 ratio. In addition, an increase in the number of CD4+ T cells expressing the homing receptor CD49d in high density was observed after treatment, which correlated positively with the increase in the Th1:Th2 ratio. Conclusion. We show that the Th1:Th2 ratio in the peripheral blood is raised by anti-TNFalpha mAb treatment. PMID- 10524690 TI - The influence of HLA-DRB1 alleles and rheumatoid factor on disease outcome in an inception cohort of patients with early inflammatory arthritis. AB - OBJECTIVE: There are conflicting data concerning the role of HLA-DRB1 alleles in disease outcome in early rheumatoid arthritis. The exact role of these alleles in short-term outcome is determined in this large, prospective, population-based study. METHODS: We recruited 532 patients with inflammatory polyarthritis from the Norfolk Arthritis Register and typed their sera for HLA-DRB1 alleles using polymerase chain reaction-based methods. Disease outcome was assessed at 2 years in terms of persistent joint inflammation, functional disability, and radiologic erosions. Results are expressed as risk ratios (RR) with 95% confidence intervals (95% CI). RESULTS: There was no influence of HLA-DRB1 alleles, in any combination, on the likelihood of disease persistence, and only a modest effect on functional disability (Health Assessment Questionnaire score > or = 1). The most obvious effect was on the development of erosions (RR 1.9, 95% CI 1.4-2.6 for those who carried at least 1 DRB1 shared epitope [SE] allele), with slightly greater effects for those who were homozygous for SE-bearing alleles (RR 2.5, 95% CI 1.8-3.6). This effect of HLA-DRB1 was restricted to patients whose sera were negative for rheumatoid factor. Among patients with erosions, HLA-DRB1 had no influence on the severity of radiologic damage (defined as the number of eroded joints, or total Larsen score). CONCLUSION: These data do not support routine HLA DRB1 screening of patients with early arthritis to identify those at risk for subsequent severe disease. PMID- 10524691 TI - Persistence of mild, early inflammatory arthritis: the importance of disease duration, rheumatoid factor, and the shared epitope. AB - OBJECTIVE: To determine the factors that predict clinical outcome at 6 months for patients with mild, early inflammatory arthritis. METHODS: Sixty-three patients with mild, untreated, early arthritis were given a single dose of corticosteroids at presentation. Administration was intramuscular if disease was polyarticular (n = 53) or intraarticular if patients had <5 synovitic joints (n = 10). The primary outcome measure was clinical disease remission or persistence of arthritis at 6 months following injection. RESULTS: At 6 months following injection, 49 of the 63 patients (78%) had persistent inflammatory joint disease. The other 14 (22%) had clinical disease remission. Regression analysis showed that only disease duration was significantly associated with persistent arthritis (P < 0.05). The other significant factor (by chi-square test) was the presence of the shared epitope (SE). Of the patients fulfilling the American College of Rheumatology (ACR) criteria at presentation (51% of the total), 53% with disease duration of < or = 12 weeks at presentation had persistent disease 6 months later, compared with 94% of those who presented with disease duration of >12 weeks. CONCLUSION: The strongest predictor of persistent disease was a disease duration of >12 weeks. Rheumatoid factor and SE were also predictors to a lesser extent. Patients who both fulfilled the ACR classification criteria for rheumatoid arthritis (RA) and had a short disease duration included some with an excellent prognosis. Therefore, 12 weeks may be a more appropriate disease duration to use for the RA classification criteria. Administering a bolus of corticosteroids may be a useful diagnostic/therapeutic approach. PMID- 10524692 TI - Objective and subjective sleep disturbances in patients with systemic lupus erythematosus. AB - OBJECTIVE: To assess objective and subjective evidence of sleep disorders in patients with systemic lupus erythematosus (SLE) and to examine correlations between parameters of lupus activity, depression, and sleep disturbances. METHODS: Fourteen SLE patients and 11 normal control subjects of similar age underwent all-night polysomnography on 3 consecutive nights. The patients and controls were also evaluated for daytime sleepiness by the Multiple Sleep Latency Test and completed a sleep disorders questionnaire and the Beck Depression Inventory. RESULTS: The polysomnographic data showed that sleep in SLE patients was characterized by respiratory and movement disorders. These intrinsic primary sleep disorders are related to the symptom of restless, poor sleep at night. Lupus patients were more sleepy during the day, and their sleepiness was related to sleep fragmentation, with more arousals and stage transitions than the control group. Disease activity was associated with decreases in sleep efficiency and delta sleep and with increases in sleep fragmentation. Depression was not correlated with the activity of the disease. CONCLUSION: There is an enhanced presence of sleep disorders in patients with SLE. The most frequent primary sleep disorders are respiratory and movement disorders. PMID- 10524693 TI - Poly(ADP-ribose) polymerase alleles in French Caucasians are associated neither with lupus nor with primary antiphospholipid syndrome. GRAID Research Group. Group for Research on Auto-Immune Disorders. AB - OBJECTIVE: To investigate the putative involvement of poly(ADP-ribose) polymerase (PARP) alleles in systemic lupus erythematosus (SLE) and primary antiphospholipid syndrome (APS). METHODS: This study of French Caucasians included 171 unrelated patients with SLE, 88 unrelated patients with primary APS, and 193 ethnically matched healthy controls. The SLE group comprised 89 patients with sporadic SLE and 82 patients with familial SLE. Patients' and controls' DNA were genotyped for the various alleles of a polymorphic CA dinucleotide repeat located within the promoter region of PARP. RESULTS: No statistically significant difference was observed for the distribution of PARP alleles between the healthy control group and each patient group or the pooled SLE patient group. CONCLUSION: The study findings strongly suggest that these alleles do not influence susceptibility to SLE or primary APS in French Caucasians. PMID- 10524694 TI - Detection of bacterial DNA in serial synovial samples obtained during antibiotic treatment from patients with septic arthritis. AB - OBJECTIVE: The management of septic arthritis could benefit from sensitive tests that detect the persistence of microorganisms in the joint. The aim of this study was to determine the feasibility of monitoring the presence of bacterial DNA in synovial samples from septic arthritis patients during antibiotic treatment. METHODS: Synovial fluid (SF) and synovial tissue (ST) samples were collected serially from 6 patients with septic arthritis before and during antibiotic therapy. In addition, peripheral blood (PB) samples were available for polymerase chain reaction (PCR) analysis from 5 of the 6 patients before treatment. All samples were analyzed for the presence of bacterial DNA with the use of a PCR with universal 16S ribosomal RNA primers. Automated sequencing and comparative data analysis were performed to identify the species. These data were compared with Gram staining and culture results. RESULTS: The bacterial species cultured from the synovium could be identified in all 6 patients using PCR and subsequent sequence analysis of the amplicons. In virtually all cases, positive Gram stain and culture findings in the synovial samples became negative after 2-3 days of antibiotic treatment. Bacterial DNA persisted in the SF and/or ST after culture conversion; in 2 patients, bacterial DNA was still detected at day 10, in 1 patient, at day 20, and in another patient, at day 22 after the initiation of treatment. Synovial samples were available for PCR analysis from 2 patients at day 26. At this time point, bacterial DNA could not be detected anymore. All PB samples were negative by both culture and PCR analysis. CONCLUSION: PCR analysis can be used to monitor the presence of bacterial DNA in synovial samples from patients with septic arthritis during antibiotic treatment. The absence of bacterial DNA could help in the decision to discontinue antibiotic treatment. PMID- 10524695 TI - Extrahepatic manifestations of chronic hepatitis C. MULTIVIRC Group. Multidepartment Virus C. AB - OBJECTIVE: To assess the prevalence of clinical and biologic extrahepatic manifestations of hepatitis C virus (HCV) infection and to identify associations between clinical and biologic manifestations. METHODS: To analyze the natural history of extrahepatic manifestations of HCV infection, we reviewed only the data recorded prospectively during the first visit of 1,614 patients with chronic HCV infection, coming from a single monocenter cohort. Exclusion criteria were positivity for hepatitis B surface antigen or human immunodeficiency virus. The prevalence of dermatologic, rheumatologic, neurologic, and nephrologic manifestations; diabetes; arterial hypertension; autoantibodies; and cryoglobulins were assessed. Then, using multivariate analysis, we identified demographic, biochemical, immunologic, virologic, and liver histologic factors associated with the presence of extrahepatic manifestations. RESULTS: At least 1 clinical extrahepatic manifestation was observed in each of 1,202 patients (74%). Five manifestations had a prevalence >10%: arthralgia (23%), paresthesia (17%), myalgia (15%), pruritus (15%), and sicca syndrome (11%). Four biologic abnormalities had a prevalence >5%: cryoglobulins (40%), antinuclear antibodies (10%), low thyroxine level (10%), and anti-smooth muscle antibodies (7%). Only vasculitis, arterial hypertension, purpura, lichen planus, arthralgia, and low thyroxine level were associated with cryoglobulin positivity. By univariate and multivariate analyses, the most frequent risk factors for the presence of clinical and biologic extrahepatic manifestations were age, female sex, and extensive liver fibrosis. CONCLUSION: Extrahepatic clinical manifestations are frequently observed in HCV patients and involve primarily the joints, muscles, and skin. The most frequent immunologic abnormalities include mixed cryoglobulins, antinuclear antibodies, and anti-smooth muscle antibodies. The most frequent risk factors for the presence of clinical and biologic extrahepatic manifestations are advanced age, female sex, and extensive liver fibrosis. PMID- 10524696 TI - Development of validated disease activity and damage indices for the juvenile idiopathic inflammatory myopathies. II. The Childhood Myositis Assessment Scale (CMAS): a quantitative tool for the evaluation of muscle function. The Juvenile Dermatomyositis Disease Activity Collaborative Study Group. AB - OBJECTIVE: To develop, validate, and determine the measurement characteristics of a quantitative tool for assessing the severity of muscle involvement in children with idiopathic inflammatory myopathies. METHODS: The Childhood Myositis Assessment Scale (CMAS) was developed from 2 existing observational functional assessment tools to assess muscle function in the areas of strength and endurance across a wide range of ability and ages. The 14 ordinal items included were chosen to assess primarily axial and proximal muscle groups and are ranked with standard performance and scoring methods. Following the development of the CMAS, a training video and written instructions were developed and reviewed by the physicians participating in this study. Subsequently, utilizing a randomized block design, 12 physicians independently scored 10 children (9 with dermatomyositis, 1 with polymyositis; ages 4-15 years) twice in one day (morning and afternoon) on the CMAS. A pediatric physical therapist performed quantitative manual muscle strength testing (MMT) twice on each child (morning and afternoon), including the neck, trunk, and proximal and distal extremity muscle groups. RESULTS: The CMAS has a potential range of 0-51, with higher scores indicating greater muscle strength and endurance. The observed mean for the 10 patients was 36.4 (median 44, SD 14.1, observed range 5-51). The total score for the CMAS correlated with the physician's global assessment (by visual analog scale) of disease activity, the MMT score, serum creatine kinase level, and the Juvenile Arthritis Functional Assessment Report score. The score on the CMAS was not correlated with patient age. Interrater reliability (Kendall's coefficient of concordance) ranged from 0.77 to 1.0 for individual items (all P < 0.001), and overall, it was 0.95 (P < 0.001). Intrarater reliability for the individual physicians was measured by correlation of the CMAS scores for each patient on 2 separate evaluations and ranged from 0.97 to 0.99, with an overall correlation for all physicians of 0.98 (all P < 0.001). CONCLUSION: The CMAS demonstrated an acceptable range of observed scores, excellent convergent validity, and excellent inter- and intrarater reliability. The CMAS is validated to quantitatively assess muscle function in the areas of strength and endurance in children with idiopathic inflammatory myopathies. It can be used in routine clinical care as well as therapeutic trials. PMID- 10524697 TI - Toward a multidimensional Health Assessment Questionnaire (MDHAQ): assessment of advanced activities of daily living and psychological status in the patient friendly health assessment questionnaire format. AB - OBJECTIVE: To develop components of a multidimensional Health Assessment Questionnaire (MDHAQ) through the addition of new items in the "patient-friendly" HAQ format, including advanced activities of daily living (ADL), designed to overcome "floor effects" of the HAQ and modified HAQ (MHAQ) in which patients may report normal scores although they experience meaningful functional limitations, and psychological items, designed to screen efficiently for psychological distress in routine care. METHODS: The new MDHAQ items, as well as scales for pain, fatigue, helplessness, and global health status on a 2-page questionnaire, were completed by 688 consecutive patients with various rheumatic diseases, including 162 with rheumatoid arthritis (RA), 114 with fibromyalgia, 63 with osteoarthritis, 34 with systemic lupus erythematosus, 20 with vasculitis, 18 with psoriatic arthritis, 16 with scleroderma, and 261 with various other rheumatic diseases, over 2 years at a weekly academic rheumatology clinic. RESULTS: The new MDHAQ items have good test-retest reliability and face validity. MHAQ scores were highest in patients with RA, and scores for other scales were highest in patients with fibromyalgia. On the advanced ADL, 58% of patients reported difficulty with errands, 68% with climbing stairs, 79% with walking two miles, 87% with participating in sports and games, and 94% with running or jogging two miles. On the psychological items, 75% of patients reported difficulty with sleep, 63% with stress, 61% with anxiety, and 57% with depression. Normal MHAQ scores were reported by 23% of patients and normal HAQ scores by 16% of patients who completed these questionnaires, while fewer than 5% had normal scores on the MDHAQ. CONCLUSION: The MDHAQ items overcome in large part the "floor effects" seen on the HAQ and MHAQ, and are useful to screen for problems with sleep, stress, anxiety, and depression in the "patient-friendly" HAQ format. These data support the value of completion of a simple 2-page patient questionnaire by each patient at each visit to a rheumatologist. PMID- 10524698 TI - Sonography and magnetic resonance imaging equivalent for the assessment of full thickness rotator cuff tears. AB - OBJECTIVE: To investigate the diagnostic value of sonography (SG) and magnetic resonance imaging (MRI) in the assessment of full-thickness rotator cuff tears (RCTs). METHODS: Twenty-one consecutive, otherwise healthy patients with noninflammatory unilateral chronic (>3 months) shoulder complaints due to a possible full-thickness RCT were studied (9 women and 12 men, mean +/- SD age 56 +/- 12). According to standardized procedures, SG was performed by both a radiologist and a rheumatologist, and MRI was evaluated by 2 radiologists. All assessors were blinded to the patient's diagnosis. Within 3 weeks after SG and MRI, arthroscopy was performed. SG, MRI, and arthroscopy results were scored as negative or positive for the presence of a full-thickness RCT. The result of surgical inspection was used as the "gold standard." RESULTS: For full-thickness RCTs, the sensitivity was 0.81 for SG and 0.81 for MRI. The specificity was 0.94 for SG and 0.88 for MRI. The positive predictive value was 0.96 for SG and 0.91 for MRI. The negative predictive value was 0.77 for SG and 0.74 for MRI. Accuracy was 0.86 for SG and 0.83 for MRI. CONCLUSION: Full-thickness RCTs can be identified accurately by both SG and MRI. Because of its low cost and because it can be performed in the rheumatology unit, SG seems to be a promising diagnostic tool for use by the rheumatologist. PMID- 10524699 TI - Clinical and experimental evidence for persistent Yersinia infection in reactive arthritis. AB - The findings of bacterial antigens in the joint and persistent triggering infection elsewhere in the body are thought to be important in the pathogenesis of reactive arthritis (ReA). We describe a patient with clinical and laboratory features consistent with this. The initial presentation with erythema nodosum and periarthritis due to infection with Yersinia pseudotuberculosis IV was followed 13 months later by recurrent erythema nodosum with joint effusion. At that time, synovial fluid was shown to contain Yersinia antigens, and, surprisingly, Yersinia-specific 16S ribosomal RNA (rRNA) sequences were also identified by reverse transcriptase-polymerase chain reaction and sequencing. Since there was no serologic evidence of reinfection, we postulate that a silent persistent Yersinia infection was reactivated, leading to dissemination of organisms to the joint, with consequent induction of ReA. Although the finding of synovial Yersinia antigens years after the original infection in ReA has previously been reported, the presence of Yersinia 16S rRNA indicates that viable organisms were also able to reach the joint. PMID- 10524700 TI - Distinguishing primary angiitis of the central nervous system from cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy: the importance of family history. AB - Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is a genetically linked neurologic disease characterized by recurrent strokes and progressive or stepwise dementia, with or without migraine-like headaches, seizures, and pseudobulbar palsy. We describe a patient referred with a diagnosis of treatment-refractory primary angiitis of the central nervous system. Meningocortical and skin biopsies confirmed that the patient had CADASIL. Clinical and radiographic differences in these disorders may be subtle, but awareness of them is crucial if the patient is to avoid unnecessary exposure to potentially deleterious immunosuppressive therapy. PMID- 10524701 TI - Shrinking lung in primary Sjogren's syndrome. PMID- 10524702 TI - Antiphospholipid antibodies in patients with Wegener's granulomatosis and polyarteritis nodosa. PMID- 10524703 TI - Long-term followup of naproxen-induced pseudoporphyria in juvenile rheumatoid arthritis. PMID- 10524704 TI - Leukocytapheresis and rheumatoid arthritis: comment on the article by Hidaka et al. PMID- 10524705 TI - Association between the inflammatory response and the risk of developing irreversible cranial ischemic complications: comment on the article by Cid et al and the letter by Nesher and Sonnenblick. PMID- 10524706 TI - Giant cell arteritis disease patterns: comment on the article by Brack et al. PMID- 10524707 TI - Ovarian failure with thalidomide treatment in complex aphthosis: comment on the concise communication by Ordi et al. PMID- 10524708 TI - Mechanisms through which PDGF alters intracellular calcium levels in U-1242 MG human glioma cells. AB - PDGF-BB induces a rapid, sustained increase in intracellular calcium levels in U 1242 MG cells. We used several calcium channel blockers to identify the types of channels involved. L channel blockers (verapamil, nimodipine, nicardipine, nitrendipine and taicatoxin) had no effect on PDGF-BB induced alterations in intracellular calcium. Blockers of P, Q and N channels (omega-agatoxin-IVA, omega conotoxin MVIIC and omega-conotoxin GVIA) also had no effect. This indicates that these channels play an insignificant role in supplying the Ca2+ necessary for PDGF stimulated events in U-1242 MG cells. However, a T channel blocker (NDGA) and the non-specific (NS) calcium channel blockers (FFA and SK&F 9365) abolished PDGF-induced increases in intracellular calcium. This indicates that PDGF causes calcium influx through both non-specific cationic channels and T channels. To study the participation of intracellular calcium stores in this process, we used thapsigargin, caffeine and ryanodine, all of which cause depletion of intracellular calcium stores. The PDGF effect was abolished using both thapsigargin and caffeine but not ryanodine. Collectively, these data indicate that in these human glioma cells PDGF-BB induces release of intracellular calcium from caffeine- and thapsigargin-sensitive calcium stores which in turn lead to further calcium influx through both NS and T channels. PMID- 10524709 TI - Amino acid efflux and cell volume regulation in cerebrocortical minislices prepared from chronically hyponatraemic and hypernatraemic rats. AB - The rates of efflux of pre-loaded amino acids, and associated steady-state volumes, were measured in cells in cerebrocortical minislices prepared from chronically (4 day) hypo- and hypernatraemic rats. The findings were compared with those obtained when cells from normonatraemic rats were acutely exposed to comparable levels of anisosmotic stress. In the presence of 122 mmol/l Na+ cells from normal rats showed increases in the rates of efflux of D-aspartate and GABA, and significant swelling (both by comparison with levels in media containing 142 mmol/l Na+). Conversely there was no acceleration of efflux in cells from hyponatraemic rats (plasma Na+ = 119-126 mmol/l) and volumes were preserved at levels comparable with those in isomotically incubated cells from normal rats. In media containing 164 mmol/l Na+ amino acid efflux in cells from normal rats was retarded, and shrinkage occurred. In cells from chronically hypernatraemic rats (plasma Na+ = 160-166 mmol/l) the rates of efflux of D-aspartate and D-glutamate were accelerated by comparison with cells from normal rats, with volume preservation. However there was no increase in the rate of GABA or glycine efflux, and cell swelling was observed. It is concluded (i) that during chronic hyponatraemia the presence of D-aspartate or GABA is associated with cell volume preservation, (ii) during chronic hypernatraemia acidic, but not neutral, amino acids are also effective in this respect, and (iii) that the markedly differing patterns of efflux responses to acute and chronic anisosmotic stress are likely to reflect chronic volume-regulatory adaptations of the efflux mechanism(s). PMID- 10524710 TI - The in vivo modulation of dopamine synthesis by calcium ions: influences on the calcium independent release. AB - To investigate the contribution of the dopamine (DA) synthesis to both the calcium-dependent and the carrier-mediated, mechanisms of DA release in the striatum, anaesthetized rats were locally superfused in the striatum with a push pull cannula supplied with an artificial CSF containing tritiated tyrosine. DA, dihydroxyphenylacetic acid (DOPAC) and their respective specific activity were measured in effluent and used to evaluate changes in the DA synthesizing rate. Excluding calcium ions from the CSF only partially reduced spontaneous DA release (70%) still leaving a possible carrier-mediated DA release. This effect was not additive with a local superfusion with 0.1 mM a-methyl-p-tyrosine, a blocker of DA synthesis, suggesting that synthesis could already be reduced by calcium-free superfusion. Local superfusion with 100 microM cadmium in the presence or not of calcium ions, increased the DA release (220 and 350%, respectively), simultaneously reducing DA synthesis. Local application of 1 microM calcium ionophore (A23187) was without effect on the basal release of DA but enhanced DA synthesis and increased the amphetamine-evoked and carrier-mediated amine release. We conclude that DA synthesis can be a modulatory process of the firing independent and carrier-mediated amine release while it weakly affects the classical calcium-dependent release. PMID- 10524711 TI - Effects of ouabain on in situ cardiac sympathetic nerve endings. AB - Using dialysis technique, the effects of ouabain on in situ cardiac sympathetic nerve endings were examined in anesthetized cats. Dialysis probes were implanted in the left ventricular myocardium, and the concentration of dialysate norepinephrine (NE) was used as an indicator of NE output at the cardiac sympathetic nerve ending. Locally applied ouabain dose-dependently (1, 10, 100 microM) increased dialysate NE levels. This finding suggested that ouabain causes an increase in NE efflux without any requirement for prior mobilization of NE from vesicular stores. Transection of sympathetic nerves innervating the heart, was without effect on the ouabain (100 microM)-induced increase in NE efflux. Pretreatment with a Ca2+-channel blocker, omega-conotoxin GVIA (10 microg/kg i.v.) suppressed the ouabain-induced NE efflux. These data suggested that ouabain opened N-type calcium channels coupled to NE release without centrally mediated neural transmission. Furthermore, ouabain-induced NE efflux was suppressed by pretreatment with desipramine (neuronal NE uptake inhibitor, 100 microM). Our data suggest that the two mechanisms (exocytosis and carrier-mediated outward transport), to the same extent, contributed to the amount of NE efflux evoked by ouabain in in situ cardiac sympathetic nerve endings. PMID- 10524712 TI - Rapid and simple measurement of serotonin N-acetyltransferase activity by liquid biphasic diffusion assay. AB - We report here a rapid, simple, and accurate method to assay for serotonin N acetyltransferase (NAT) activity. This assay relies on the selective diffusion of radiolabeled acetyltryptamine into a water-immiscible scintillation fluid. Unlike organic solvent extraction, thin-layer chromatography, or high performance liquid chromatography, the separation of acetyltryptamine from acetyl CoA and tryptamine is not required in the method. Moreover, the limit of sensitivity is less than 4 pmol of N-acetyltryptamine formed per sample. Enhancement of NAT activity upon beta-adrenergic receptor stimulation in the rat pineal gland was clearly detected with this method. In addition, the NAT activity measurements obtained with this method agreed quantitatively in the pineal gland and other brain tissues with the conventional organic solvent extraction method. The results suggest that this liquid biphasic diffusion assay is applicable to the detection of NAT activity in tissues and cells. PMID- 10524713 TI - Sustained potentiation of AP1 DNA binding is not always associated with neuronal death following systemic administration of kainic acid in murine hippocampus. AB - Mice were intraperitoneally injected with kainic acid (KA), followed by dissection of frozen coronal sections and subsequent punching out of the pyramidal and granular cell layers in the hippocampus under a binocular microscope. Systemic administration of KA resulted in marked and sustained potentiation of binding of a radiolabeled double stranded oligonucleotide probe for the nuclear transcription factor activator protein-1 (AP1) in the pyramidal cell layers of the CA1 and CA3 subfields and the granule cell layers of the dentate gyrus 2-18 h later. Morphological evaluation using cresyl violet revealed marked losses of neuronal layers in the pyramidal CA1 and CA3 subfields, but not in the granular dentate gyrus, within 6 weeks after administration. Supershift analysis using antibodies against different Jun and Fos family members differentiated between AP1 DNA binding in hippocampal nuclear extracts obtained 2 and 18 h after the administration of KA. These results suggest that neuronal death may not always follow modulation of de novo synthesis of particular proteins through sustained potentiation of AP1 DNA binding which involves expression of different Jun and Fos family members in response to systemic administration of KA in murine hippocampus. PMID- 10524714 TI - Evidence that endogenous thymosin alpha-1 is present in the rat central nervous system. AB - We have previously reported that administration of Thymosin alpha 1 (T-alpha1) can enhance the level of the Nerve Growth Factor and the distribution of its receptor in the developing Central Nervous System (CNS) of rat. To further explore the role of T-alpha1 and verify its presence in cells of rat CNS, we carried out an immunohistochemical study using a polyclonal antibody against T alpha1. T-alpha1 immunoreactivity was found mainly in neurons of the hippocampus and spinal cord and in several small cells, resembling glial cells, of specific regions of the brain. Moreover, to study whether cerebral cells were receptive to T-alpha1, we injected iodinated T-alpha1 (125I-T-alpha1) i.c.v.. 125I-T-alpha1 labelled neurons were observed in the hypothalamus and septal nuclei. Our results indicate that specific neuronal populations in the rat CNS are able to express and respond to T-alpha1. PMID- 10524715 TI - Opposite modulation of capsaicin-evoked substance P release by glutamate receptors. AB - Substance P and glutamate are present in primary afferent C-fibers and play important roles in persistent inflammatory and neuropathic pain. In the present study, we have examined whether activation of different glutamate receptor subtypes modulates the release of substance P evoked by the C-fiber selective stimulant capsaicin (1 microM) from rat trigeminal nucleus slices. The selective NMDA glutamate receptor agonist L-CCG-IV (1-10 microM) enhanced capsaicin-evoked substance P release about 100%. This facilitatory effect was blocked by 0.3 microM MK-801, a selective NMDA receptor antagonist. The metabotropic glutamate receptor agonists L-AP4 (group III) and DHPG (group I) (30-100 microM) inhibited capsaicin-evoked substance P release by approximately 60%. These inhibitory effects were blocked by the selective metabotropic glutamate receptor antagonist (+/-)-MCPG (5 microM). On the other hand, AMPA and kainate (0.1-10 microM), did not significantly affect capsaicin-evoked substance P release. Thus, substance P release from non-myelinated primary afferents, and possibly nociception, may be under the functional antagonistic control of some metabotropic and ionotropic glutamate receptor subtypes. PMID- 10524716 TI - Developmental vs. social personality models of adult attachment and mental ill health. AB - Both the developmental and social personality approaches to the study of adult attachment are concerned with understanding those factors that describe an individual's quality of relational adaptation and risk for mental ill health. This paper examines the theoretical and methodological assumptions of these alternative models and how these assumptions have markedly different implications for addressing clinical issues. It is suggested that recent evidence necessarily leads to the conclusion that mental and relational difficulties, such as partner violence and victimization, borderline personality, dissociation, suicidal behaviour and other clinical symptomology thought to be related to experiences of severe relationship distress, are best explained in terms of attachment disorganization rather than as normative forms of attachment insecurity or fearful avoidant adult romantic attachment. PMID- 10524717 TI - Attachment style and adult love relationships and friendships: a study of a group of women at risk of experiencing relationship difficulties. AB - This study examines the relationship between attachment style and love relationships and friendships in a group of women (N = 40) known to be at risk of experiencing relationship problems. The association between attachment style and measures of self-esteem and depression were also investigated. Women with a secure attachment style had more positive ratings in the domain of adult love relationships than women with insecure attachment style (avoidant and ambivalent) and difficulties in adult love relationships were found to be particularly related to an avoidant attachment style. Insecure attachment style was also related to having cohabited with a deviant partner. Adult attachment style was not found to be related to ratings of current mood but was significantly related to self-esteem and to ratings of functioning in the domain of adult friendships. In particular, participants with an anxious-ambivalent attachment style had more negative self-esteem than secure participants. Secure participants had more positive ratings in the domain of adult friendships than insecure participants and a moderately significant association was also found between difficulties in the domain of adult friendships and an anxious-ambivalent attachment style. In addition, 20% (N = 8) of the women also reported attachment styles characterized by high levels of avoidance and ambivalence: this group was found to have more pervasive difficulties in their close relationships than women who endorsed a single dominant attachment style. PMID- 10524718 TI - An investigation of shame and guilt in a depressed sample. AB - A self-report measure of proneness to shame and guilt was administered to 86 patients with moderate to severe depression, with the prediction that there would be a positive correlation of shame with severity of depression. Contrary to other, non-clinical studies, we found that guilt but not shame was associated with levels of depression. Shame-proneness demonstrated a unique association with a stable attributional style for negative outcomes, global negative self evaluation, submissive behaviour and internalized anger. Contrary to prediction, no relationship was found between shame- or guilt-proneness and a reported history of childhood sexual abuse. PMID- 10524719 TI - Linking verbal and non-verbal representations: computer analysis of referential activity. AB - The objective of this study was to develop a computer assisted procedure to model the Referencial Activity scales as scored by raters. Referential Activity is defined as the function of connecting non-verbal experience with language. Using a large text corpus that had been rated by experienced and reliable judges, extreme samples from both ends of the Referential Activity Scales were selected. The Characteristic Vocabularies for each of these corpora, words that were significantly more frequent in each corpus as compared to the other, were then identified. A small set of 181 frequent words was derived that accounted for half of all words in the text corpora. These words were used as dictionaries for a Computerized Referential Activity measure based on computer assisted content analysis techniques. The new measure showed a correlation with judge-scored Referential Activity of around .50 across both the development and test corpora. PMID- 10524720 TI - A cross-case comparison of two independent analyses of intake workers' descriptions of the process of assessment for psychotherapy. AB - This study compares the analyses by two clinical psychologists of material gathered during research interviews held with 18 intake workers employed by four different institutes for psychotherapy. Each intake worker responded to questions regarding their experiences with two clients during intake interviews for psychotherapy. During the research interviews, the intake workers described their impressions regarding the clients as persons, the course of the encounter with these clients as well as motives for proposing a particular type of treatment. Comparison of the two psychologists' analyses of the original material revealed remarkably similar descriptions of the structure of the intake workers' experiences and of that of their motivation for proposing several types of treatment. In line with contemporary qualitative research practices, the current comparative study may be seen as a form of interpreter triangulation. PMID- 10524721 TI - Therapeutic commitment and role security in work with men with violence-related problems: an investigation and test of a model. AB - This study investigated whether therapists' therapeutic commitment and role security could be important therapist factors in therapy with those client groups that are perceived as being difficult to work with. Therapeutic commitment represents how committed therapists were to such work and role security measures to how secure they felt in their role. This study explored whether these factors could potentially explain some of the individual variation found between therapists in treatment outcomes. Following a pilot study, 209 therapists, consisting of 20 violence counsellors, 56 Relate counsellors, 58 forensic and prison psychologists and 75 clinical psychologists, completed self-report questionnaires by post about their work with violent men. Therapists' therapeutic commitment and role security were found to correlate significantly with expectations of clinical outcome on a case vignette, and with the level of violence that was usually worked with. Role security and therapeutic commitment also correlated significantly with therapists' level of experience, education about violence, role support and self-esteem. Differences existed between the professional groups on these latter factors and also in therapeutic commitment and role security. The results indicated the importance of role security in dealing with violence. PMID- 10524722 TI - Coping with the disfiguring effects of vitiligo: a preliminary investigation into the effects of cognitive-behavioural therapy. AB - Vitiligo is a progressive condition involving a loss of pigmentation in the skin; it can be disfiguring and no effective treatment or cure exists. Although vitiligo's medical effects have been studied extensively, little attention has been paid to its psychological impact or to the effects of psychological state on the illness itself. To address these issues, the present study examined the effect of cognitive behavioural therapy on coping with vitiligo and adaptation to the negative effects on body image, quality of life and self-esteem in adult patients. The study also examined whether any psychological gains acquired from psychological therapy would influence the progression of the condition itself. Two matched groups of vitiligo patients were compared, one of which received cognitive-behavioural therapy over a period of 8 weeks, while the other received no changes to their treatment status. All patients were assessed on self-esteem, body image and quality of life, prior to, immediately following and 5 months following the end of therapy. The progression of the condition was assessed by photographing patients prior to the start of counselling and 5 months following counselling. Results suggest that patients can benefit from cognitive behavioural therapy in terms of coping and living with vitiligo. There is also preliminary evidence to suggest that psychological therapy may have a positive effect on the progression of the condition itself. Implications for incorporating psychological counselling into patient care and management are discussed. PMID- 10524723 TI - The use of stop signals to reduce the pain and distress of patients undergoing a stressful medical procedure: an exploratory clinical study. AB - The present study investigates stop signals and their effects on the pain and distress of patients undergoing a stressful medical procedure. Thirty-six chronic pain patients (17 men, 19 women) attending an out-patient operating theatre for diagnostic nerve blocks/local anaesthetic injections were allocated to one of two conditions (experimental and control). All patients received a standard information leaflet concerning the forthcoming injections. Additional information was given to those in the experimental group on four occasions (three orally, one written) before the injections which stated that they could halt the procedure at any time by saying 'stop'. Subjective measures of anxiety, pain, distress, sense of control over the procedure as well as observer ratings of patient distress and pain behaviour were obtained before, during and after the injections. After initial differences in pre-injection pain were controlled for the experimental group, patients rated themselves as less distressed during the injections and recorded lower state anxiety following treatment. In view of various methodological limitations of the present study its findings may only be accorded 'pilot study' status. These limitations are explored in the discussion along with their implications for a more robust replication study. Nevertheless the present findings provide tentative support for the hypothesis that the use of stop signals can reduce the stressful nature of diagnostic nerve blocks. PMID- 10524724 TI - Interpreting the Inventory of Interpersonal Problems: subscales based on an interpersonal theory model. AB - There remains considerable debate about a theoretically interpretable subscale structure for the Inventory of Interpersonal Problems (IIP). In this paper items are extracted from the IIP to form a 40-item shortened version (the IIP-40) comprising eight subscales, each of five items, which conform to the eight octant positions within Birtchnell's interpersonal octagon: a version of interpersonal theory. The inter-item reliability of the subscale structure is found to be acceptable when tested against a sample of 150 pre-assessment for psychotherapy IIP completions within the Centre for the Study of Psychotherapy (CSP), University of Kent. The face validity is established through acceptable inter rater reliability scores in an experiment using blind raters. Subscale scores are shown for patients within CSP and are found to be significantly different between genders in the octant positions of Upper Close and Neutral Distant. High scores in the octant position of Upper Distant is found to be a significant predictor of therapeutic drop-out. The results for gender and therapeutic engagement are consistent with other published work in the interpersonal theory field. PMID- 10524725 TI - Confirmatory factor analysis of the Level of Expressed Emotion (LEE) scale. AB - A confirmatory analysis of a previously reported structure for the Level of Expressed Emotion (LEE) scale was conducted with data from 75 volunteers. Three principal components were extracted and rotated to maximum congruence with a target based on previous research. The fit was found to be highly significant and retest correlations for the subscales of the LEE were good. However, the revised LEE scale could probably be improved, especially by the addition of a subscale designed to measure perceived criticism. PMID- 10524726 TI - Improving survival: a multi-portal approach to improving cardiopulmonary resuscitation outcomes. PMID- 10524727 TI - Acute changes in arterial carbon dioxide tension and acid-base status and early neurologic characteristics in term infants following perinatal asphyxia. AB - BACKGROUND: Marked acute changes in arterial carbon dioxide tension (PaCO2) and acid-base status occur in the immediate postnatal period in infants delivered in the presence,of pathologic fetal acidemia (FA) in whom the risk for hypoxic ischemic cerebral injury is high. The cerebral vasculature is extremely sensitive to changes in PaCO2. However, the relationship between the acute changes in PaCO2 and subsequent neonatal neurologic characteristics remains unclear. OBJECTIVES: (1) To determine the extent of the acute changes in PaCO2 and acid-base status following birth in infants delivered in the presence of pathologic FA and (2) to determine the potential relationship of the initial changes in PaCO2 and neonatal neurologic characteristics. METHODS: PaCO2 and acid base status of cord umbilical arterial blood and initial postnatal arterial blood were studied in 73 term infants admitted to the Neonatal Intensive Care Unit. Infants were categorized in three groups: I, no FA, no respiratory support and normal neonatal neurologic examination (n = 49); II, pathologic FA (umbilical artery pH < or = 7.00, base deficit > or = 12 mEq/l), no respiratory support and normal neonatal neurologic examination (n = 17); III, FA, intubated and with evidence of hypoxic ischemic encephalopathy (HIE) including seizures (n = 7). RESULTS: Demographic characteristics were similar among the three groups, although 5-min Apgar score < or = 5 was more common in group II (47%) and group III (100%) than in group I (4%). Umbilical arterial pH was lower in group III (6.75 +/- 0.18) vs. group II (6.90 +/- 0.09) and in group II vs. group I (6.90 +/- 0.09 vs. 7.19 +/- 0.09) (P < 0.005) and the PaCO2 was higher in group III (141 +/- 37 mmHg) vs. group II (94 +/- 22 mmHg) and in group II vs. group I (94 +/- 22 vs. 60 +/- 13 mmHg) (P < 0.05). The mean base deficit was large but comparable between groups III and II, i.e. 18 +/- 6 vs. 18 +/- 5 mEq/l, respectively, and higher than in group I infants (6 +/- 4 mEq/l) (P < 0.00). At 1 h postnatal age, the mean arterial pH had increased in all groups, i.e. 7.06 +/- 0.15 (group III), 7.25 +/- 0.09 (group II), and 7.31 +/- 0.06 (group I); however, the differences amongst the groups remained significant (P < 0.005). The mean PaCO2 decreased from 94 +/- 22 mmHg (12.5 +/- 2.9 kPa) to 30 +/- 6 mmHg (4.0 +/- 0.8 kPa) for the spontaneously ventilating group II infants and from 141 +/- 37 mmHg (18.8 +/- 4.9 kPa) to 45 +/ 14 mmHg (6.0 +/- 1.9 kPa) in the intubated group III infants (P < 0.005). A repeat PaCO2 at 2 h of age in group III infants had decreased to 29 + 2 mmHg (3.9 +/- 0.3 kPa),which was not different from the PaCO2 at 2 h in group II infants (30 +/- 8 mmHg; 4.0 +/- 1.1 kPa). No significant differences were observed for pH or base deficit at this time. CONCLUSIONS: Marked and rapid changes in PaCO2 and pH were observed in term infants delivered in the presence of pathologic FA. Initial postnatal PaCO2 values varied significantly with the lowest values noted in those infants breathing spontaneously and who exhibited an uneventful neonatal course; higher initial postnatal values, despite mechanical ventilation, were noted in infants with HIE including seizures. Further investigation in this area is imperative in order to better define the optimal respiratory management of the neurologically at-risk infant. PMID- 10524728 TI - Sustained ventricular tachycardia in the emergency department. AB - The aim of the study was to evaluate the demographics, haemodynamics, ECG characteristics, underlying disease, tachycardia termination and outcome of patients with sustained ventricular tachycardia (VT). We registered 75 patients presenting with VT (51 male, median age 63) from December 1993 to August 1998 in our emergency department (ED). Seventeen of these patients were haemodynamically unstable (23%), and 58 patients were stable (77%); there was no difference in the tachycardia cycle length (median 320 ms) and QRS width (median 140ms) between the two groups; however, five of the seven patients with polymorphic VT pattern were in the unstable group. Ischaemic heart disease was the underlying disorder in 57 patients (76%). Acute myocardial infarction (AMI) was present in 12 of the 58 stable (21%) compared to 11 of the 17 unstable (65%) patients. In three patients (4%) VT terminated spontaneously, in 34 patients (45%) VT was terminated by first line intravenous drug therapy, and in 38 patients (51%) including all 17 unstable and 22 stable who failed to respond to the intravenous antiarrhythmic therapy challenge out of 55 patients, VT was terminated by electrical therapy. Within 2 days, 48 patients (64%) were transferred to an open ward, 13 (17%) still needed intensive care, nine (12%) were discharged to home and five (7%) died. Death occurred due to cardiac failure from AMI with extensive anterior wall infarction in three patients, and due to constrictive pericarditis and reocclusion of stented LAD each in one patient. At presentation in the emergency department, the majority of the patients with VT were haemodynamically stable, thus allowing first-line antiarrhythmic drug administration. However, in the course of the disease, half needed electrical therapy for definitive termination of the tachycardia. Therefore, direct current cardioversion must be available in the emergency department. Haemodynamic instability and death occurs significantly more often if VT occurs during the course of AMI. PMID- 10524729 TI - Leadership of resuscitation teams: "Lighthouse Leadership'. AB - AIM: The purpose of this study was to determine the relationship between leadership behaviour, team dynamics and task performance. METHODS: This was as an observational study, using video recordings of 20 resuscitation attempts. The Leadership Behaviour Description Questionnaire (LBDQ) was used to measure the level of structure built within the team. Interpersonal behaviour and the tasks of resuscitation were measured with a team dynamics and a task performance scale. The degree to which the leader actively participated, 'hands on', with the tasks of resuscitation, and their previous training in advanced life support (ALS), and experience of resuscitation attempts, were evaluated against the leadership rating. RESULTS: The degree to which the leader built a structure within the team was found to correlate significantly with the team dynamics (P = 0.000) and the task performance (P = 0.013). Where the leaders participated 'hands on' they were less likely to build a structured team (P = 0.005), the team were less dynamic (P = 0.028) and the tasks of resuscitation were performed less effectively (P = 0.099). Experience gained over a 1-year period did not enhance leadership performance, but leaders who had up to 3 years experience were more likely to be effective in this role (P = 0.072). Interestingly, ALS training did not enhance leadership performance per se. However those leaders who had had recent ALS training were more likely not to participate 'hands on' (P = 0.035). There were some notable shortcomings in the performance of the task and some interesting correlations relating to duration of resuscitation, survival rate estimations, the leaders' attitudes and the teams' level of experience. CONCLUSION: Leaders must build a structure within a resuscitation team in order for them to perform effectively. An emergency leadership training programme is essential to enhance the performance of leaders and their teams. PMID- 10524730 TI - Cardiopulmonary resuscitation: errors made by pre-hospital emergency medical personnel. AB - The purpose of the current study was to evaluate the CPR techniques of emergency healthcare professionals (emergency medical technicians, firemen, emergency first responders, CPR instructors). Skills were evaluated using a Laerdal Skillmeter Manikin, which provided a computerized printout of the quantifiable data during the CPR sequence. All of the 66 subjects in the study had completed a recertification course within the last 2 years (mean = 0.86 +/- 0.18, 95% CI). The sequence was videotaped for later viewing and for correlating the errors with the data. In addition, the participants were required to fill in a questionnaire. The most frequently occurring errors were observed in landmarking, overcompression, palpating a carotid pulse and insufficient ventilation. Although 98.5% of participants made an attempt to landmark their position for compression on the sternum, 35.9% of the total compressions performed by all subjects were incorrectly positioned on the patient's chest. Overcompression of the patient's chest accounted for 55.3% of incorrect compressions. Although 94% of participants attempted to verify a carotid pulse, only 45% were able to feel it and therefore stop performing cardiac massage. Of the total ventilations, 49% were below the American Heart Association (AHA) recommended minimum (800 ml). The results of this study showed a high rate of errors occurring in the CPR provided by emergency healthcare professionals. PMID- 10524731 TI - Intra-aortic administration of epinephrine above aortic occlusion does not alter outcome of experimental cardiopulmonary resuscitation. AB - Intra-aortic balloon occlusion during experimental cardiopulmonary resuscitation (CPR) improves coronary perfusion pressure and resuscitability and provides unique access to the central circulation. It has been hypothesized that administration of epinephrine into the aortic arch in combination with aortic occlusion would further improve haemodynamics during CPR, resuscitability and 24 h survival. In 16 anaesthetised dogs intravascular catheters were placed for hemodynamic and blood gas monitoring. An aortic balloon catheter was placed by femoral artery insertion with its tip just distal to the left subclavian artery. Ventricular fibrillation for 7.5 min without CPR, 2.5 min of Basic Life Support with chest compressions and ventilation with 100% oxygen were followed by 30 min of Advanced Cardiac Life Support (ACLS) with systemic canine drug dosages. The intra-aortic balloon was inflated when ACLS started and gradually deflated shortly after restoration of spontaneous circulation (ROSC). Epinephrine, in 100 microg/kg boluses every 5 min until the heart was restarted or 30 min had elapsed was administered through the intra-aortic catheter in the experimental group (n = 8) and via a central venous catheter in the control group (n = 8). Coronary perfusion pressure increased during the ACLS period in both groups (P < 0.05) with no difference between the groups and there was no difference in the frequency of ROSC (experimental group 5/8, control group 4/8). Furthermore with respect to 24 h survival, there was no difference between the experimental group (2/8) and the control group (3/8). Severe macroscopic haemorrhagic necrosis of the myocardium in the dogs with ROSC was found in 4/5 in the experimental group compared to 1/4 in the control group. In conclusion, intra-aortic administration of 100 microg/kg epinephrine doses combined with aortic occlusion during experimental CPR did not alter outcome. PMID- 10524732 TI - Meglumine antimoniate, amiodarone and torsades de pointes: a case report. AB - Pentavalent antimonial drugs used for the treatment of leishmaniasis have been associated with sudden deaths, probably due to the development of ventricular tachyarrhythmias. Prolongation of the QT interval and ventricular tachyarrhymias have been described in patients on amiodarone therapy. We report a case of recurrent torsades de pointes following treatment with pentavalent antimonial drugs and amiodarone. PMID- 10524733 TI - A case of progressive congestive heart failure secondary to severe anemia in a patient presenting with uterine hemorrhage. AB - In this report, we present a 42-year-old female patient who was transferred to our emergency department due to symptoms of congestive heart failure. She presented with severe anemia (hemoglobin was 1.3 g dl(-1), and hematocrit was 6.0%) due to continuous uterine hemorrhage and metabolic acidosis, otherwise she seemed to be free from illness. We diagnosed that she was suffered from chronic severe anemia due to uterine hemorrhage and congestive heart failure. Monitoring her hemodynamic status, treatment of congestive heart failure using diuretics and inotropes in combination with blood transfusion brought her good recovery. We discussed this case from the mechanisms of development of congestive heart failure in a chronic severe anemic condition, and pointed out that distributive effects of sodium and water may develop congestive heart failure without myocardial dysfunction in such a condition. PMID- 10524734 TI - Tracheal intubation with the aid of a magnet. PMID- 10524735 TI - Former SLEH-THI cardiology fellow is elected president of the American Heart Association. PMID- 10524736 TI - A tribute to C. Walton Lillehei, the "Father of open heart surgery". PMID- 10524737 TI - Evolution of the ventricles. AB - We studied the evolution of ventricles by macroscopic examination of the hearts of marine cartilaginous and bony fish, and by angiocardiography and gross examination of the hearts of air-breathing freshwater fish, frogs, turtles, snakes, and crocodiles. A right-sided, thin-walled ventricular lumen is seen in the fish, frog, turtle, and snake. In fish, there is external symmetry of the ventricle, internal asymmetry, and a thick-walled left ventricle with a small inlet chamber. In animals such as frogs, turtles, and snakes, the left ventricle exists as a small-cavitied contractile sponge. The high pressure generated by this spongy left ventricle, the direction of the jet, the ventriculoarterial orientation, and the bulbar spiral valve in the frog help to separate the systemic and pulmonary circulations. In the crocodile, the right aorta is connected to the left ventricle, and there is a complete interventricular septum and an improved left ventricular lumen when compared with turtles and snakes. The heart is housed in a rigid pericardial cavity in the shark, possibly to protect it from changing underwater pressure. The pericardial cavity in various species permits movements of the heart-which vary depending on the ventriculoarterial orientation and need for the ventricle to generate torque or spin on the ejected blood- that favor run-off into the appropriate arteries and their branches. In the lower species, it is not clear whether the spongy myocardium contributes to myocardial oxygenation. In human beings, spongy myocardium constitutes a rare form of congenital heart disease. PMID- 10524738 TI - Central venous injuries of the subclavian-jugular and innominate-caval confluences. AB - Injuries to the central venous system can result from penetrating trauma or iatrogenic causes. Injuries to major venous confluences can be particularly problematic, because the clavicle and sternum seriously limit exposure of the injury site. We report our institution's experience with central venous injuries of the subclavian-jugular and innominate-caval venous confluences. Significant injuries of the subclavian-jugular venous confluence frequently result from penetrating trauma, while injuries to the innominate-caval confluence are usually catheter-related. Median sternotomy provides adequate exposure of the innominate caval confluence, while exposure of the subclavian-jugular venous confluence requires extension of the median sternotomy incision into the neck and resection of the clavicle. The literature is reviewed. PMID- 10524739 TI - Diagnosis and treatment of concomitant aortic and coronary disease: a retrospective study and brief review. AB - Coronary arteriosclerosis seriously complicates the surgical treatment of aortic diseases. The aim of our retrospective study was to determine the incidence of coronary artery disease among our surgical patients in treatment for aortic dissection or aneurysm, and to determine whether coronary intervention before aortic surgery appears to affect outcomes. Between 1 January 1993 and 1 March 1998, our center treated 253 patients for aortic dissection or aneurysm. We examined these cases retrospectively for information on diagnostic and treatment methods, both for the aortic lesions and for concomitant coronary arteriosclerosis. Aortic dissection had been detected in 86 (33.9%) patients and aortic aneurysm in 167 (66.1%). Coronary angiography was performed to search for concomitant coronary artery disease in 29 (33.8%) patients with dissection and in 112 (67.1%) patients with aneurysm; of these, 11 (12. 7%) and 54 (32.3%), respectively, were found to have coronary disease. Among 43 patients with abdominal aortic aneurysm in whom coronary angiography was performed, concomitant coronary disease was detected in 36 (83.7%). Coronary artery bypass surgery was performed in 10 patients who had dissection and in 30 patients who had aneurysm; percutaneous transluminal coronary angioplasty was performed in 7 patients who had aneurysm. Perioperative mortality rates in the dissection and aneurysm groups, overall, were 23.2% and 13.8%, respectively Unfortunately, the prospective, random clinical study that would be necessary to prove the case for or against preoperative coronary angiography among subsets of patients in need of aortic repair would raise ethical questions, given the strength of the information already in our possession, gathered by less formal methods. Our study reinforces existing evidence that preoperative angiography can reduce mortality and morbidity in the elective repair of aortic aneurysm, especially thoracic or abdominal aneurysm. However, angiography should not be performed routinely in cases of aortic dissection and should be withheld in cases of type A dissection. PMID- 10524740 TI - Intraluminal milrinone for dilation of the radial artery graft. AB - There is renewed interest in the use of the radial artery as a conduit for coronary artery bypass surgery. The radial artery is, however, a very muscular artery, prone to vasospasm. Milrinone, a potent vasodilator, has demonstrated vasodilatory properties superior to those of papaverine. In this report, we describe our technique of radial artery harvesting and the adjunctive use of intraluminal milrinone as a vasodilator in the preparation of this conduit for coronary artery bypass grafting. We have used these techniques in 25 patients who have undergone coronary artery bypass grafting using the radial artery. No hand ischemic complications have been observed in this group. Intraluminal milrinone appears to dilate and relax the radial artery, rendering this large conduit spasm free and very easy to use. We recommend the skeletonization technique for radial artery harvesting and the use of intraluminal milrinone as a radial artery vasodilator in routine myocardial revascularization. PMID- 10524741 TI - Technical aspects of mitral valve replacement with an allograft for acute bacterial endocarditis. AB - Mitral valve replacement with a mitral valve allograft is receiving a resurgence of interest. We discuss the technical aspects of this procedure as it applies to cases of acute bacterial endocarditis infecting the mitral valve. PMID- 10524742 TI - The use of a stentless porcine bioprosthesis to repair an ascending aortic aneurysm in combination with aortic valve regurgitation. AB - Over the years, many surgical methods have evolved for the treatment of ascending aortic aneurysm in combination with aortic valve regurgitation; however, precise guidelines for optimal surgical techniques for varying presentations have not been defined. We describe the use of a stentless porcine bioprosthesis (Medtronic Freestyle) in a patient with an ascending aortic aneurysm and aortic regurgitation. We used the complete root replacement method, and anastomosed a Dacron graft (Hemashield) between the bioprosthetic valve and the native aorta to replace the distal part of the aneurysm. PMID- 10524743 TI - The history and development of cardiac transplantation. AB - The history of heart surgery, spanning only 100 years to date, has seen some of the most daring and persistent men and women in all of medical history. Many aspects of heart surgery, including such innovations as the heart-lung machine, aortic aneurysm surgery, and the correction of congenital heart defects, have provided future surgeons with an important lesson: diligent research can solve complex problems. The history and development of cardiac transplantation is particularly full of challenges that have been overcome, with the research phase alone spanning more than 90 years. During that time, essential contributions came from all over the world, including the United States, Russia, England, and South Africa. As is typical of medical advancement, individual contributions did not stand alone but added to the experience of those who had come before. Even so, the work of a few particular groups deserves special recognition. Most notable is the Stanford team, led by Dr. Norman Shumway, who continued to transplant human hearts when other institutions had abandoned hopes for the operation. Largely because of the commitment of that team, cardiac transplantation has become a standard option in the treatment of end-stage heart disease. Currently, only the availability of donor hearts limits the number of cardiac transplantations performed worldwide. PMID- 10524744 TI - Cor triatriatum in adults: three new cases and a brief review. AB - We report 3 cases of cor triatriatum that were diagnosed late, in the 4th and 5th decades of life. The presentations of these 2 men and 1 woman varied substantially, both in anatomic and symptomatic aspects. The woman had an associated complex congenital anomaly, which is not uncommon in cases of cor triatriatum. We present our cases, along with a review of this rare congenital cardiac anomaly. PMID- 10524745 TI - Differential pulmonary flow in hypoplastic left heart syndrome. AB - We report a case of hypoplastic left heart syndrome associated with restrictive interatrial communication and partial anomalous pulmonary venous connection via a right lower pulmonary vein draining to the inferior vena cava. We found unequal pulmonary artery pressure and different pulmonary artery structure, with the right pulmonary artery being lower in pressure and more tortuous and dilated in its peripheral branches than the left. This was attributed to the variant degrees of pulmonary venous obstruction. The left pulmonary venous return was severely obstructed by the restrictive interatrial communication, whereas the anomalous right lower pulmonary vein drained into the inferior vena cava, with less obstruction. To the best of our knowledge, there has never before been a report of differential pulmonary flow associated with a partial anomalous pulmonary vein, in a case of hypoplastic left heart syndrome. PMID- 10524746 TI - Asymptomatic rupture of an aortoiliac aneurysm. AB - The rupture of an abdominal aortic aneurysm is one of the most feared complications confronted by cardiovascular surgeons. Such ruptures are usually catastrophic, but in some instances the rupture is posterior and remains sealed. These chronic ruptures may manifest with any of a variety of clinical presentations. This report describes an uncommon presentation of a chronic rupture of an aortoiliac aneurysm in a patient with generalized aneurysmal disease. The rupture presented as an asymptomatic giant pulsatile mass in the patient's abdomen. The mass had developed over a period of several years. The literature is also reviewed. PMID- 10524747 TI - Autologous vein-coated stent for exclusion of a coronary artery aneurysm: case report with postimplantation intravascular ultrasound characteristics. AB - This report describes the successful use of an autologous cephalic vein-coated coronary stent to exclude an aneurysm of the distal right coronary artery. Post implantation angiography confirmed successful exclusion of the aneurysm with no evidence of leakage. Intravascular ultrasonography showed complete apposition of the stent to the arterial wall proximal and distal to the aneurysm. The vein could be seen clearly around the stent. Symmetrical stent expansion (minimal luminal diameter, 2.8 mm) was verified. Increased echogenicity in the excluded aneurysm indicated early thrombus formation. Evidently, this is the 1st report of the successful use of an autologous cephalic vein-coated coronary stent to exclude an aneurysm of the distal right coronary artery. PMID- 10524748 TI - Replacement of an immobile prosthetic mitral valve: a case report. AB - A mechanical prosthetic heart valve can become acutely obstructed despite anticoagulation therapy. This can be a life-threatening complication. We report the case of a 38-year-old woman who survived obstruction of her Sorin prosthetic mitral valve. She was admitted to the hospital because of severe pulmonary edema. On auscultation, mechanical valve sounds were absent. Transthoracic echocardiography showed an immobile mechanical valve. The patient suffered a cardiac arrest while being prepared for surgery, but she underwent successful mitral valve replacement after cardiopulmonary resuscitation. When patients with prosthetic mitral valves present with acute dyspnea, the possibility of an obstructed prosthetic valve must be considered in the differential diagnosis. PMID- 10524750 TI - Tuberculous pseudoaneurysm of the descending thoracic aorta: successful treatment by surgical excision and primary repair. AB - Tuberculous pseudoaneurysm of the aorta is a rare disease with a high mortality rate. We present the case of a 27-year-old woman who had a tuberculous pseudoaneurysm of the descending thoracic aorta. The patient underwent successful excision and primary repair of the lesion while under hypothermic circulatory arrest and partial femoral bypass. To the best of our knowledge, this is the youngest patient to be successfully treated with surgery for a tuberculous pseudoaneurysm of the descending thoracic aorta. The pathogenesis, diagnosis, and treatment of this disease are reviewed, and the need to include tuberculous pseudoaneurysm in the differential diagnosis of chest lesions is emphasized. PMID- 10524749 TI - The significance of incidental noncardiac findings in Tc-99m sestamibi myocardial perfusion imaging: illustrated by a case. AB - Technetium 99m sestamibi is widely used in the evaluation of myocardial perfusion imaging. Although the aim of such imaging is cardiac evaluation, numerous other organs are included in the imaging field. Failure to identify incidental abnormal findings in these organs delays diagnosis and treatment. In common with other radiopharmaceutical agents, technetium 99m sestamibi is distributed throughout the body and accumulates in multiple tissues. When interpreting studies that involve this radiotracer, the physician must be aware of its physiologic distribution, in order to recognize abnormal uptake. We present an illustrative case in which areas of decreased tracer activity were noted incidentally during the evaluation of unprocessed single photon emission computed tomography data. These findings were due to metastasis of colon cancer to the liver. PMID- 10524751 TI - Ectopic origin of the left anterior descending artery from the posterior descending branch. PMID- 10524752 TI - Primary bronchomalacia and patent ductus arteriosus: simultaneous surgical correction in an infant. AB - We report the clinical course of a 6-month-old girl with recurrent infection of the left lung, persistent wheezing, and a suspected congenital heart anomaly (patent ductus arteriosus. Chest radiography revealed hyperinflation and slight inflammation of the left lung. Tracheobronchoscopy and left-sided bronchography showed a collapsed segment of the left main bronchus, 3 cm long. Computed tomography confirmed hyperinflation of the left lung and atelectasis of the superior lobe. There were no signs of extramural compression. Color-flow Doppler echocardiography confirmed the suspicion of patent ductus arteriosus. To the best of our knowledge, there is no other report in the literature of a patient with this combination of anomalies. After receiving 2 weeks of antibiotic treatment, the patient underwent surgical repair The patent ductus arteriosus was closed by means of a triple-ligature procedure, and during the same operation a bronchopexy was performed, securing the left main bronchus to the closed ductus tissue by means of sutures. There have been no complications in the postoperative period. Clinical follow-up, as well as echocardiography and bronchoscopy, have yielded normal results 14 months after surgery. PMID- 10524753 TI - Evidence for a mosaic structure of the Tn5481 in Lactococcus lactis N8. AB - The sequences of the left end of the nisin-sucrose transposon Tn5481 in Lactococcus lactis subsp. lactis N8, the adjacent 1.5 kb chromosomal region upstream of the junction site as well as a 5.0 kb region downstream of the nisZBTCIPRKFEG genes within the transposon have been determined. In the upstream chromosomal region, an incomplete open reading frame encoding a protein with strong N-terminal homology to the low-affinity branched chain amino acid carriers was identified. Within the transposon, downstream of the nisin gene cluster, a 186 bp almost identical copy of the left hand sequence was located. Further downstream, four new open reading frames were found. The codon usage in these reading frames as well as the G+C content of the region are clearly different from those of the nisin genes, suggesting that the functionally unrelated areas of Tn5481 are gathered from different origins during the evolution of the transposon. PMID- 10524754 TI - Characterization of the nisFEG operon of the nisin Z producing Lactococcus lactis subsp. lactis N8 strain. AB - Biosynthesis of the food additive nisin, a posttranslationally modified peptide antibiotic existing as two natural variants (A and Z), requires eleven genes (nisA/ZBTCIPRKFEG) involved in modification, secretion, regulation and self immunity. The suggested self-immunity genes (nisFEG) of the nisin Z producer Lactococcus lactis subsp. lactis N8 were cloned and sequenced. Putative binding sites of the NisR transcription factor were recognized upstream of the nisF promoter. The hydrophilic NisF protein was expressed in Escherichia coli and shown to be associated with the membrane. Expression of the nisF gene from a plasmid in L. lactis MG1614, a strain lacking the nisin operons, did not increase the nisin resistance of the cells. This showed that NisF alone does not protect against nisin. Overexpression of the nisF gene in the N8 nisin producer did not affect the level of nisin immunity, indicating that the wild-type amount of NisF is not limiting the level of nisin immunity. Production of antisense-nisEG or antisense-nisG RNA in L. lactis N8 resulted in severe reduction in the level of nisFEG mRNA and a clearly reduced immunity showing that the nisFEG transcript is important for development of nisin self-immunity. PMID- 10524755 TI - Variability of P1 protein of zucchini yellow mosaic virus for strain differentiation and phylogenetic analysis with other potyviruses. AB - The complete nucleotide sequence of a Singapore isolate of zucchini yellow mosaic potyvirus (ZYMV-S) was determined from viral cDNA clones. The complete genome is 9603 nucleotides in length excluding the poly (A) tail. Computer analysis of the sequence revealed a single large open reading frame (ORF) that presumably encodes a polyprotein of 3082 amino acids with a calculated molecular weight of 350 kDa. Analysis of the helper component (HC) protein showed that the highly conserved motif K-I-T-C which is involved in aphid transmission appeared as K-L-S-C. There is also a change of D-A-G to G-A-G triplet near the N-terminal of the coat protein (CP). Amino acid sequence identity comparison of ZYMV-S gene products with the California and Reunion Island isolates of ZYMV revealed a minimum range of 65-75% to a maximum range of 95-98%. Comparison with other distinct potyviruses showed a low degree of identity from 19-74%. The 5' untranslated region (UTR) of ZYMV-S showed 67% and 72% identity when compared with the California and Reunion Island isolates, respectively. The sequence variability in the 5' UTR of ZYMV can be exploited for strain differentiation and phylogenetic analysis. ZYMV-S shared 94% and 82% identity in the 3' UTR as compared to the California and Reunion Island isolates, respectively. The P1 protein of ZYMV-S shared moderate sequence variability among ZYMV isolates but high sequence variability among all potyviruses. In addition, phylogenetic analysis using the P1 protein indicated that highly variable proteins in the viral genome could also be employed in the study of potyvirus taxonomy and used for strain differentiation. PMID- 10524758 TI - Sequence analysis of the Rhizobium etli ribose kinase gene rbsK and its phylogenetic position. AB - DNA sequence analysis of a 1878-bp DNA fragment located downstream from the Rhizobium etli ptsN gene revealed the presence of an open reading frame coding for a protein of 300 amino acids. This protein is homologous to members of the PfkB subfamily of carbohydrate and carbohydrate phosphate kinases. Since the highest homology is observed with the ribokinases of Escherichia coli, Haemophilus influenzae and Bacillus subtilis, the isolated gene was named the R. etli rbsK gene. The eubacterial ribokinases form a cluster distinct from the cluster of ribokinase proteins of the archaebacteria Methanobacterium thermoautotrophicum, Methanococcus jannaschii and Sulfolobus solfoataricus, which form a more divergent group of proteins. R. etli RbsK has a molecular mass of 30.6 kDa and a calculated isoelectric point of 4.5. No homologues of Escherichia coli ORF284 and ORF90 were found downstream from R. etli ptsN. PMID- 10524756 TI - Molecular cloning of Nedd4 from Xenopus laevis. AB - The tryptophan-bounded WW domains ofNedd4 bind to the proline-tyrosine (PY) motifs contained in the C-terminal cytoplasmic region of the beta and gamma subunits of the rat amiloride-sensitive sodium channel (ENaC). In patients with Liddle's syndrome, the PY motif is mutated and the channel remains constitutively activated leading to sodium retention and hypertension. Although the function ofNedd4 is unknown, it contains a highly conserved ubiquitin protein ligase domain that may attach ubiquitin to ENaC, targeting it for degradation or it may modulate ENaC activity through another undetermined pathway. Xenopus laevis derived cells, such as oocytes and the A6 kidney cell line, are important models currently used for the study of ENaC regulation. We describe the X. laevis homologue of Nedd4 (xNedd4). A partial clone, approximately 2.6 Kb, was isolated from an aldosterone-treated A6 cell cDNA library. Further 5' sequence, approximately 1.2 Kb, was obtained using a modified 5' rapid amplification of cDNA (RACE) protocol and cDNA from untreated A6 cells as the substrate. The identity and similarity of xNedd4 with human Nedd4 are approximately 63 and 71%, respectively. xNedd4 contains the C2, ubiquitin protein ligase, and 4 WW domains previously described for Nedd4 from other species. PMID- 10524759 TI - Cloning and characterisation of a prohibitin gene from infective larvae of the parasitic nematode Toxocara canis. AB - Infective larvae of the parasitic nematode Toxocara canis express an mRNA (Tc-pro 1) encoding a predicted protein that shares significant homology with prohibitin, a protein involved in inhibition of cell proliferation - The closest homologues of Tc-pro-1 include an expressed sequence tag (EST) from Caenorhabditis elegans and Drosophila L2Cc, a protein thought to be essential for larval development and moulting. Other homologues include prohibitin from rat and human and an EST from Saccharomyces cerevisiae. Parasite life cycles generally include periods of developmental arrest, which in the larvae of T. canis may persist for many years without loss of metabolic activity. This report of the first full-length gene encoding prohibitin from a parasitic nematode raises interesting suggestions about the potential role of prohibitin in diapause and in the regulation of moulting in development. PMID- 10524760 TI - Isolation and characterization of an auxin-inducible SAUR gene from radish seedlings. AB - A gene homologue to the auxin-inducible SAUR (Small Auxin Up RNAs) of soybean was isolated from a cDNA and a genomic library of radish. The cDNA clone was about 470 bp in length and contained a DST (Down STream) element that was involved in mRNA instability in the 3'-untranslated region. The genomic clone contained two short sequences that related to auxin-responsible core sequences (AGTCTC and TATCCCAC) in its promoter region. The radish SAUR transcript was accumulated by auxin and cycloheximide treatments in the same way as that in other plant species. However, the NDE and DUE elements that were reported in all of the other SAUR gene promoters were not conserved in this radish SAUR gene. The results suggest that the promoter activity of the SAUR gene will be regulated by only two short sequences. PMID- 10524757 TI - Sequencing analysis of forty-eight human image cDNA clones similar to Drosophila mutant protein. AB - We have sequenced 48 human IMAGE cDNA clones selected from the public EST database (dbEST) for their significant homology to Drosophila mutant genes. A dynamically updated analysis report was produced by BlastX and BlastN analysis searches in the latest databases available. This analysis led us to estimate the grade of similarity with homologous genes isolated in other species. Bottlenecks were detected in the sequencing process and here we have presented our problem solving approach. We think the value of this full-length sequencing project is an enrichment of the sequence database information that is currently available to the human genome community. PMID- 10524762 TI - Nucleotide sequence of a three gene cluster in Neisseria Gonorrhoeae encoding ribosomal proteins S6, S18, and L9. AB - A cluster of three genes, rpsF, rpsR, and rpII, encoding the ribosomal proteins S6, S18, and L9, respectively, were cloned and sequenced from Neisseria gonorrhoeae. The order of the genes within the cluster was established as rpsF rpsR-rpII. Within this cluster an additional open reading frame of unknown identity spanning 108 bp was found between rpsF and rpsR. The putative amino acid sequences deduced from all three genes show a high degree of homology to other bacterial ribosomal proteins. PMID- 10524761 TI - Molecular cloning of Arabidopsis photolyase gene (PHR1) and characterization of its promoter region. AB - Photolyase is an enzyme that repairs ultraviolet (UV)-damaged DNA by photoreactivation. In higher plants, accumulation of photolyase (PHR1) mRNA is induced by either UV or visible light. In order to know the molecular mechanism by which PHR1 gene expression is induced by light, we have determined the genomic structure and the 5'-flanking sequence of the Arabidopsis PHR1 gene. The PHR1 gene spans approximately 2.5 kb of genomic DNA and consists of 9 exons. In the promoter region of PHR1, there are two pairs of inverted repeats spanning more than sixty base pairs. The promoter also contains DNA motifs similar to the GT-1 box or G-box found in many light-inducible gene promoters. EMSA analysis showed that several proteins in Arabidopsis nuclear extract bound to the G-box-like motifs. These results raise the possibility that the Arabidopsis PHR1 gene is regulated by transcription factors which interact with these motifs. PMID- 10524765 TI - Identification and nucleotide sequence of the heat shock protein 60 (GroEL) gene of Bacteroides forsythus. AB - Bacteroides forsythus is a Gram negative anaerobe associated with human periodontal disease. Heat shock proteins are immunodominant antigens in bacteria that can elicit strong and protective immune responses. The gene specifying the GroEL protein (hsp60, heat shock protein 60) of B. forsythus was isolated by PCR amplification using consensus primers based upon published nucleotide sequences of the groEL genes of several bacterial species. Translation of the gene sequence predicts a protein of 544 amino acids in length with a molecular mass of 58 kDa. B. forsythus GroEL demonstrates identities of 50 to 81% with the predicted amino acid sequences of GroEL proteins of several bacterial species and the human mitochondrial P1 protein. PMID- 10524763 TI - cDNA and deduced amino acid sequences of dog catalase. AB - The nucleotide sequence of dog catalase was determined from cDNA obtained by reverse transcriptase-polymerase chain reaction (RT-PCR) and 5'- and 3'- rapid amplification of cDNA ends (RACE) using dog liver mRNA. The deduced amino acid sequence was compared with the sequences of other mammalian catalases and was found to show 93.0, 92.0, 93.3 and 91.7% similarity with mouse, rat, bovine and human catalases, respectively. PMID- 10524764 TI - The genomic and sequence analysis of rat histone H2B genes. AB - In addition to previously characterized testis-specific and somatic H2B histone genes, two somatic histone H2B genes, hereafter called sH2B-2 and sH2B-3, were isolated from a rat genomic library, genomic organization was determined, and their promoter was sequenced. Like many other H2B genes, sH2B-2 gene was closely linked to H2A gene whereas H3 gene was located upstream of sH2B-3 gene. Deletion and mutation analysis of 5' sequence fused to CAT reporter gene revealed that the interaction between CCAAT and octamer binding factors is important for S-phase specific activation of sH2B-3 gene. PMID- 10524766 TI - Cloning and sequencing of the rat cDNAs encoding class I beta-tubulin. AB - Two kinds of rat cDNA clones encoding class I beta-tubulin were isolated from the neonatal rat brain, and sequenced. They corresponded to mRNA species formed by the alternative usage of polyadenylation signals. The determined nucleotide sequence showed high (84.6%) identity to rat class II beta-tubulin, the only rat beta-tubulin sequence reported in the nucleotide database, in the coding region, but relatively low (47.3-50.7%) in the noncoding region. The Northern blot analysis using the 3'-noncoding fragment as a probe showed that both mRNA species of the class I beta-tubulin were down-regulated during brain development. PMID- 10524767 TI - Sequence comparison of the VP7 of serotype G2 rotaviruses from diverse geographical locations. AB - The sequences of the genes encoding the outer capsid glycoprotein, VP7, of twelve serotype G2 rotavirus isolates from Australia were determined. The deduced amino acid sequences were compared by phylogenetic analysis to published sequences from strains collected from various geographical locations worldwide between 1976 an 1993. This analysis showed that geographical clustering of strains was apparent and that most strains were distantly related to strain DS1 whose VP7 is included in the rhesus rotavirus tetravalent vaccine formulation. Comparisons of the neutralization epitope regions of VP7 also indicated that most strains might exhibit antigenic differences to DS1. These results have implications for the development of future vaccine strategies. PMID- 10524768 TI - Crystal structure of the complex formed between bovine beta-trypsin and MCTI-A, a trypsin inhibitor of squash family, at 1.8-A resolution. AB - The stoichiometric complex formed between bovine beta-trypsin and Momordica charantia, Linn. Cucurbitaceae trypsin inhibitor A (MCTI-A) was crystallized and its X-ray crystal structure was refined to a final R value of 0.179 using data of 7.0- to 1.8-A resolution. Combination with results on the complex of MCTI-A with porcine trypsin gives the sequence of MCTI-A definitely, of which 13 residues are conserved compared with other squash family trypsin inhibitors. Its spatial structure and the conformation of its primary binding segment from Cys3I (P3) to Glu7I (P3'), which contains a reactive scissile bond Arg5I C-Ile6I N, were found to be very similar to the other squash family proteinase inhibitors. PMID- 10524769 TI - Sequence pattern for the occurrence of N-glycosylation in proteins. AB - To further understand the occurrence of N-glycosylation, 21 nonhomologous proteins with Asn-x-Ser/Thr sequence were investigated. The results showed that some oligopeptides with Gly residues (G-x-y or y-x-G) are adjacent to the N glycosylated sequences. These oligopeptides are not only essential for the structure and function of the proteins, but they are also found to be often proteolytic processing sites. These properties suggest that these oligopeptides may be a "sequence pattern" for the occurrence of N-glycosylation. The implications of the findings for protein structure and function are discussed. PMID- 10524770 TI - Antibody immunodiversity: a study on the marked specificity difference between two anti-yeast iso-1 cytochrome c monoclonal antibodies whose epitopes are closely related. AB - Anti-yeast iso-1 cytochrome c (cyt. c) monoclonal antibodies 2-96-12 and 4-74-6 have closely related epitopes (antigenic determinants). However, while the specificity of 4-74-6 is stringent, 2-96-12 cross-reacts with many evolutionarily related cytochromes c. Such a marked difference in specificity of antibodies with overlapping epitopes may represent unique antibody immunodiversity. Thus, we constructed Fv fragment models consisting of the variable domains of the heavy and light chains of 2-96-12 and 4-74-6 and that of another anti-iso-1 cyt. c as a control to gain insight into the origin of this difference in specificity. Our models show that 4-74-6 and 2-96-12 contain five and two aromatic side chains, respectively, in or near the central area of the antigen-combining site. The side chains of Arg95H (heavy chain) in 2-96-12 and Arg91L (light chain) in 4-74-6 project toward the central area of the combining site in our model. Antigen docking to our Fv models, combined with previous immunological studies, suggests that iso-1 cyt. c Asp60 may interact with Arg95H in 2-96-12 and Arg91L in 4-74-6 and that both epitopes of 2-96-12 and 4-74-7 may include iso-1 cyt. c Leu58, Asp60, Asn62, and Asn63. The effect of the Arg95H to Lys mutation on the antigen binding is also in accord with our model. The difference in specificity may be partly explained by a greater degree of conformational flexibility in and around the central area of the combining site in 2-96-12 compared to 4-74-6 due to differences in aromatic side chain packing. PMID- 10524771 TI - Suppression of kinetic AMP cooperativity of fructose-1,6-bisphosphatase by carbamoylation of lysine 50. AB - Selective treatment of pig kidney fructose 1,6-bisphosphatase with cyanate leads to the formation of an active carbamoylated derivative that shows no cooperative interaction between the AMP-binding sites, but completely retains the sensitivity to the inhibitor. By an exhaustive carbamoylation of the enzyme a derivative is formed that has a complete loss of cooperativity and a decrease of sensitivity to AMP. It was proposed that the observed changes of allosteric properties were due to the chemical modification of two lysine residues per enzyme subunit [Slebe et al. (1983), J. Protein Chem. 2, 437-443]. Studies of the temperature dependence of AMP sensitivity and the interaction with Cibacron Blue Sepharose of carbamoylated fructose 1,6-bisphosphatase derivatives indicate that the lysine residue involved in AMP sensitivity is located at the allosteric AMP site, while the lysine residue involved in AMP cooperativity is at a distinct location. Using [14C]cyanate, we identified both lysine residues in the primary structure of the enzyme; Lys50 is essential for AMP cooperativity and Lys112 appears to be the reactive residue involved in the AMP sensitivity. According to the fructose 1,6 bisphosphatase crystal structure, Lys50 is strategically positioned at the C1-C2 interface, near the molecular center of the tetramer, and Lys112 is in the AMP binding site. The results reported here, combined with the structural data of the enzyme, strongly suggest that the C1-C2 interface is critical for the propagation of the allosteric signal among the AMP sites on different subunits. PMID- 10524772 TI - Methanol-induced unfolding and refolding of cytochrome b5 and its P40V mutant monitored by UV-visible, CD, and fluorescence spectra. AB - In order to illustrate the structural importance of proline-40 of cytochrome b5 (Cyt b5), the P40V mutant gene was constructed. Unfolding and refolding of Cyt b5 induced by methanol was investigated by means of the UV-visible spectrum, circular dichroism, and the fluorescence spectrum. Methanol denaturation of Cyt b5 is a cooperative process, that is, the heme group dissociates from the heme pocket accompanied by unfolding of the polypeptide chain both in the secondary and tertiary structures. Substitution of proline by valine reduces the stability of the mutant under methanol denaturation. The unfolding process is almost reversible by dilution. During refolding, the denatured polypeptide must be folded to a more ordered structure prior to the heme capture. Pro40 plays an important role in modulating the protein's stability. The role of tyrosine in the unfolding and refolding of Cyt b5 is evaluated for the first time. A mechanism of methanol denaturation is also proposed. PMID- 10524773 TI - Solvation energy and thermal stability of hydrophilization-modified alpha chymotrypsin. AB - As reported in the literature [Mozhaev et al. (1988), Eur. J. Biochem. 173, 147 154], when a series of modifiers, especially the cyclic anhydrides of pyromellitic and mellitic acids, are introduced into each lysine located in the alpha-chymotrypsin (CT) surface, a substantial hydrophilization of the enzyme surface can occur and remarkable stabilization effects of modified enzymes can be obtained. In this paper, four models are applied to calculate the solvation energy of native and the modified CT based on their tertiary structures, which can be built by the CVFF force field. Analyzing the relationship between the solvation energy and the thermal stability in detail, we find that the results of three solvation energy models (Ooi model, WE-1 model, and WE-2 model) can be used to illustrate the relative stability among these enzymes qualitatively. The present study should be of practical value as well as of some theoretical interest. PMID- 10524774 TI - Relationship between amino acid properties and protein stability: buried mutations. AB - In order to understand the mechanism of protein stability and to develop a simple method for predicting mutation-induced stability changes, we analyzed the relationship between stability changes caused by buried mutations and changes in 48 amino acid properties. As expected from the importance of hydrophobicity, properties reflecting hydrophobicity are strongly correlated with the stability of proteins. We found that subgroup classification based on secondary structure increased correlations significantly, and mutations within beta-strand segments correlated better than did those in alpha-helical segments, which may result from stronger hydrophobicity of the beta-strands. Multiple regression analyses incorporating combinations of three properties from among all possible combinations of the 48 properties increased the correlation coefficient to 0.88 and by an average of 13% for all data sets. Analyzing the stability of tryptophan synthase mutants with Glu49 replaced by all other residues except Arg revealed that combining buriedness, solvent-accessible surface area for denatured protein, and unfolding Gibbs free energy change increased the correlation to 0.95. Consideration of sequence and structural information (neighboring residues in sequence and in space) did not significantly strengthen the correlations in buried mutations, suggesting that nonspecific interactions dominate in the interior of proteins. PMID- 10524775 TI - What is the minimum number of residues to determine the secondary structural state? AB - The failure of protein secondary structural prediction is commonly attributed to the neglect of long-range interactions. The question is, what is the minimum length of subsequence required to determine the central secondary structural state, stabilized only by local interactions? In the present work, the 20 amino acids were classified into eight groups to analyze systematically the relationship between the length and secondary structural state of subsequences in the PDB database. It was found that the fraction of subsequences with a unique central secondary structural state increases with increasing length, and the minimum length of subsequence required to determine the central secondary structural state is about 14-17 residues. The low accuracy of secondary structure prediction does not result from the neglect of long-range interactions, but may result from the limitation of the available protein database size or prediction algorithm. PMID- 10524776 TI - The intermolecular disulfide bridge of human glial cell line-derived neurotrophic factor: its selective reduction and biological activity of the modified protein. AB - Recombinant human glial cell line-derived neurotrophic factor has been implicated to have therapeutic potential in the treatment of neurodegenerative diseases. The mature protein is a single polypeptide of 134 amino acid residues and functions as a disulfide-linked dimer. Reduction of the protein with dithiothreitol at pH 7.0 and in the absence of denaturant showed that the single intermolecular cystine bridge was reduced preferentially. Direct alkylation of the generated free sulfhydryl group using iodoacetamide or iodoacetate without denaturant was incomplete. Unfolding the protein in 6 M guanidine hydrochloride prior to the modification showed rapid disulfide scrambling. However, the sulfhydryl-modifying reagent N-ethylmaleimide was able to label quantitatively the free cysteinyl residue in the absence of any added chaotropic agent. By a combination of peptide mapping, Edman degradation, and mass spectrometric analysis, the labeled residue was identified to be Cys101, hence verifying the location of the intermolecular disulfide bond. The modified protein behaved as a noncovalent dimer when chromatographed through a Superdex 75 column under nondenaturing conditions and was comparable in biological activity to an unmodified control sample. The results therefore indicate that the intermolecular disulfide bridge of the protein is not essential for its biological function. PMID- 10524777 TI - Alternative nonallelic deletion is constitutive of ruminant alpha(s1)-casein. AB - Multiple forms of alpha(s1)-casein were identified in the four major ruminant species by structural characterization of the protein fraction. While alpha(s1) casein phenotypes were constituted by a mixture of at least seven molecular forms in ovine and caprine species, there were only two forms in bovine and water buffalo species. In ovine and caprine forms the main component corresponded to the 199-residue-long form, and the deleted proteins differed from the complete one by the absence of peptides 141-148, 110-117, or Gln78, or a combination of such deletions. The deleted segments corresponded to the sequence regions encoded by exons 13 and 16, and by the first triplet of exon 11 (CAG), suggesting that the occurrence of the short protein forms is due to alternative skipping, as previously demonstrated for some caprine and ovine phenotypes. The alternative deletion of Gln78 in alpha(s1)-casein, the only form common to the milk of all the species examined and located in a sequence region joining the polar phosphorylation cluster and the hydrophobic C-terminal domain of the protein, may play a functional role in the stabilization of the milk micelle structure. PMID- 10524778 TI - Kinetics of inhibition of green crab (Scylla serrata) alkaline phosphatase by L cysteine. AB - The inhibition of alkaline phosphatase from green crab (Scylla serrata) by L cysteine has been studied. The results show that L-cysteine gives a mixed-type inhibition. The progress-of-substrate-reaction method previously described by Tsou [(1988), Adv. Enzymol. Related Areas Mol. Biol. 61, 391-436] was used to study the inactivation kinetics of the enzyme by L-cysteine. The microscopic rate constants were determined for reaction of the inhibitor with the free enzyme and the enzyme-substrate complex (ES) The results show that inactivation of the enzyme by L-cysteine is a slow, reversible reaction. Comparison of the inactivation rate constants of free enzyme and ES suggests that the presence of the substrate offers marked protection of this enzyme against inactivation by L cysteine. PMID- 10524780 TI - Taking a bigger slice of the pie. PMID- 10524781 TI - Relationship between anisometropia, amblyopia, and binocularity. PMID- 10524779 TI - Three-dimensional structure of Selenocosmia huwena lectin-I (SHL-I) from the venom of the spider Selenocosmia huwena by 2D-NMR. AB - The three-dimensional structure of native SHL-I, a lectin from the venom of the Chinese bird spider Selenocosmia huwena, has been determined from two-dimensional 1H NMR spectroscopy recorded at 500 and 600 MHz. The best 10 structures have NOE violation <0.3 A, dihedral violation <2 deg, and average root-mean-square differences of 0.85 + 0.06 A over backbone atoms. The structure consists of a three-stranded antiparallel beta-sheet and three turns. The three disulfide bridges and three-stranded antiparallel beta-sheet form a inhibitor cystine knot motif which is adopted by several other small proteins, such as huwentoxin-I, omega-conotoxin, and gurmarin. The C-terminal fragment from Leu28 to Trp32 adopts two sets of conformations corresponding to the cis and trans conformations of Pro31. The structure of SHL-I also has high similarity with that of the N terminus of hevein, a lectin from rubber-tree latex. PMID- 10524782 TI - Fetal alcohol syndrome. AB - BACKGROUND: Fetal alcohol syndrome (FAS) describes the systemic and ocular anomalies resulting from the teratogenic effect of maternal alcohol abuse during pregnancy. It is a leading cause of preventable birth defects in the U.S. CASE REPORTS: Two case reports illustrate the characteristic findings in FAS. These include growth retardation, cognitive impairment, and facial dysmorphism. Ocular signs are prevalent, including small palpebral fissure, microcornea, strabismus, myopia, astigmatism, and optic nerve hypoplasia. DISCUSSION: Fetal alcohol exposure can lead to a wide spectrum of systemic defects and vision deficits. The increasing frequency of drinking among pregnant women in recent years should call more public attention to this detrimental yet preventable syndrome. CONCLUSION: The high frequency of ocular manifestations aids in making a diagnosis of FAS, which can be challenging. Eye-care professionals can play an important role in patient management and the educational process. PMID- 10524783 TI - Complications of neodymium:YAG cyclophotocoagulation in the treatment of open angle glaucoma. AB - A retrospective study was performed to determine the complications that occurred after cyclophotocoagulation. The condition of 33 eyes in 25 patients was observed from 1 month to 1 year after application of cyclophotocoagulation with a neodymium:YAG (Nd:YAG) laser for open-angle glaucoma. The number of treatment burns ranged from 16 to 50, and laser output energy ranged from 3 to 7 J. Complications included anterior uveitis in 14 eyes (42%), conjunctival injection in 12 eyes (36%), pain in 9 eyes (30%), and conjunctival hemorrhage in 5 eyes (15%). Corneal edema, intraocular pressure spikes, and corneal epithelial defects were each noted in three (9%) of the eyes treated, whereas cataracts developed in four (12%) of the eyes. Two eyes (6%) developed anterior segment ischemia with subsequent phthisis bulbi. Seven eyes (21 %) demonstrated no adverse reactions. PMID- 10524784 TI - Rigid contact lens fitting relationships in keratoconus. Collaborative Longitudinal Evaluation of Keratoconus (CLEK) Study Group. AB - PURPOSE: Although the influence of flat-fitting contact lenses on corneal scarring in keratoconus is frequently debated, the current standard of care with regard to the apical fitting relationship in keratoconus remains undocumented. METHODS: Patients were examined at baseline in the Collaborative Longitudinal Evaluation of Keratoconus (CLEK) Study (N = 1209). Patients wearing a rigid contact lens in one or both eyes (N = 808) had their habitual rigid contact lenses analyzed, and the fluorescein patterns and base curves were compared to the first definite apical clearance lens (FDACL). The FDACL is the flattest lens in the CLEK Study trial lens set that exhibits an apical clearance fluorescein pattern. For patients wearing a rigid contact lens in both eyes, one eye was selected randomly for analysis. RESULTS: Twelve percent of the rigid contact lens wearing eyes were wearing lenses fitted with apical clearance based upon the clinician's fluorescein pattern interpretation. The remainder (88%) was wearing lenses fitted with apical touch. For mild (steep keratometric reading <45 D) keratoconus corneas, the mean estimate of the base curve to cornea-fitting relationship was 1.18 D flat (SD +/- 1.84 D); moderate (steep keratometric reading: 45 to 52 D) corneas were fitted on average 2.38 D flat (SD +/- 2.56 D); and severe (steep keratometric reading > 52 D) corneas were fitted an average of 4.01 D flat (SD +/- 4.11 D). CONCLUSIONS: Despite the potential risk for corneal scarring imposed by flat-fitting rigid contact lenses, most CLEK Study patients wear flat-fitting lenses. Overall, rigid lenses were fitted an average of 2.86 D (SD +/- 3.31 D) flatter than the FDACL. PMID- 10524785 TI - Hydrogel lens dehydration and subjective comfort and dryness ratings in symptomatic and asymptomatic contact lens wearers. AB - PURPOSE: To determine whether lens dehydration correlates with discomfort, dryness, and noninvasive tear break-up time in symptomatic and asymptomatic contact lens wearers and whether dehydration of the two lens types varies. METHOD: Twenty hydrogel contact lens wearers with dryness-related symptoms and 20 asymptomatic wearers wore an Etafilcon A lens (Acuvue; Vistakon, Inc., Jacksonville, Florida) in one eye and an Omafilcon A lens (Proclear; Biocompatibles, Norfolk, Virginia) in the contralateral eye for 7 h in a randomized, double-masked study. Lens water content was measured before and after 7 h of lens wear and prelens noninvasive tear film break-up time (NIBUT) was measured immediately after insertion and after 5 h of lens wear. Subjective comfort and dryness were rated at 0, 1, 3, 5, and 7 h of lens wear. RESULTS: The symptomatic group had significantly reduced prelens NIBUT, decreased comfort, and increased dryness, but there was no difference between lenses for these variables. The Omafilcon A lenses dehydrated significantly less than the Etafilcon A lenses, but there was no significant difference in lens dehydration between two subject groups. CONCLUSION: No correlation was found between lens dehydration and subjective dryness and comfort. Symptomatic hydrogel contact lens wearers with decreased wearing time had measurably decreased comfort, increased dryness ratings, and reduced NIBUT. PMID- 10524786 TI - The frequency of ocular symptoms during spectacle and daily soft and rigid contact lens wear. AB - PURPOSE: Ocular discomfort is the primary reason for discontinuation of contact lens wear. The purpose of the study was to quantify and compare the frequency of ocular symptoms experienced by spectacle wearers and wearers of soft and rigid daily contact lenses. METHODS: We analyzed the results of an ocular symptom survey of prospective volunteers for contact lens clinical trials during the period 1989 to 1995. Questions pertaining to lens-wear experience and ocular symptoms were answered by 883 untrained individuals without active ocular disease. The sample included 664 spectacle wearers, 171 soft contact lens (SCL) wearers, and 48 rigid gas-permeable (RGP) lens wearers. The frequencies of 10 ocular symptoms were compared for each group. Spearman's Rank Correlation was used to test for correlations between symptoms. The chi2 test was used to determine differences between subject groups, adjusted for multiple comparisons. RESULTS: There were no significant differences in the frequency of ocular symptoms between the soft contact lens (SCL) and RGP wearers. The most common symptom was ocular tiredness (27%). None of the symptoms were highly correlated, indicating that they are somewhat different "sensations." Despite ocular discomfort being the primary reason for discontinuation of lens wear, contact lens wearers experienced the same type and severity of symptoms as spectacle wearers. Thus (in order of frequency of occurrence), tiredness, itchiness, watering, pain, aching, excessive blinking, and burning had similar rates of occurrence for all three groups. The two major distinguishing symptoms were dryness and redness, which were reported far more frequently and with greater severity in both contact lens groups (p < 0.001). Grittiness was also reported more with RGP wearers than with spectacle wearers (p < 0.001). CONCLUSIONS: Contact lenses disturb the ocular environment, as evidenced by responses of increased ocular dryness, redness, and grittiness. Despite fundamental differences in SCL's and RGP contact lenses, both groups of contact lens wearers surveyed experienced a similar type and frequency of ocular symptoms. PMID- 10524787 TI - Re-evaluation of the oxygen diffusion model for predicting minimum contact lens Dk/t values needed to avoid corneal anoxia. AB - PURPOSE: (1) To update Fatt's mathematical model of the distribution of oxygen tension (pO2) across the cornea and contact lens (CL) to include the recent finding that corneal oxygen consumption increases with the acidification that occurs with CL wear. (2) To estimate the minimum transmissibility (CL Dk/t) to avoid epithelial anoxia or to avoid stromal anoxia. METHODS: A five-layer static and one-dimensional mathematical model of oxygen diffusion through the cornea based on Fatt's models was used. The relationships between acidosis and increased QO2, and acidosis and CL Dk/t were used to estimate corneal QO2 for a given CL Dk/t. RESULTS: (1) Revised model predictions are in agreement with direct tear pO2 measurements beneath CLs in the rabbit. (2) For the human eye, the minimum CL Dk/t for oxygen delivery to the basal epithelial cells was determined to be 23 for the open eye and 89 for the closed eye. To prevent anoxia throughout the entire corneal thickness the Dk/t requirements are 35 for the open eye and 125 for the closed eye. CONCLUSIONS: (1) Model predictions of the oxygen distribution beneath contact lenses are significantly lower than previous models that did not include the effect of acidosis on corneal QO2. (2) Minimum Dk/t values that allow oxygen delivery to the basal epithelium are in agreement with the Dk/t needed to avoid corneal edema. PMID- 10524788 TI - A new schematic eye model incorporating accommodation. AB - PURPOSE: To create a schematic eye model with four refracting surfaces and with the new function describing the crystalline lens profile. METHODS: The new function describing the crystalline lens profile is a combination of hyperbolic cosine functions modulated by hyperbolic tangent functions. It ensures the continuity of the radius of curvature along the whole lens profile. Recent experimental results on cornea and lens shape measurements have been applied to the proposed model. The new function describing the lens profile has been used to model the eye during accommodation. RESULTS: Spherical aberration and modulation transfer function have been computed to test the eye model. Results of calculations are in agreement with experimental findings. CONCLUSIONS: The calculations show that spherical aberrations of the cornea and crystalline lens cancel each other. The new description of the crystalline lens offers a reduction of longitudinal spherical aberration. The image quality of the unaccommodated eye model is slightly better than that of the accommodated one. PMID- 10524789 TI - Human IgM antibody therapy for HIV-1 infection. PMID- 10524790 TI - Detection of Clostridium difficile toxin A by reversed passive latex agglutination. AB - A reversed passive latex agglutination (RPLA) assay for detecting Clostridium difficile toxin A is presented. Purified monoclonal antibody (mAb 37B5) was used for latex sensitization. The culture supernatants of 93 strains of C. difficile were tested by RPLA assay and the results compared with those of a commercially available latex agglutination test, PCR and cytotoxin assay with Vero cells. There was agreement between RPLA, cytotoxicity and PCR assays, but 29 strains were positive in the RPLA assay while 35 were positive in the cytotoxicity test and PCR using primer pair NK3-NK2 directed to the nonrepeating portion of the C. difficile toxin A gene. The 6 cytotoxic but RPLA-negative strains were demonstrated to be toxin A-negative/toxin B-positive strains in the PCR assay by using primer pair NK11-NK9 directed to the repeating portion of the C. difficile toxin A gene. There were no cross-reactions with culture supernatants of the other clostridial strains except for two strains of C. sordelli that produced hemorrhagic toxin (which is immunologically related to C. difficile toxin A). PMID- 10524791 TI - Characterization of the Coxiella burnetti sucB gene encoding an immunogenic dihydrolipoamide succinyltransferase. AB - The Coxiella burnetii sucB gene encoding the dihydrolipoamide succinyltransferase (E2o) enzyme was cloned by immunological screening of a lambda EMBL3 genomic library prepared from strain Nine Mile DNA and sequenced. The homology of the cloned gene product to the counterpart in Escherichia coli was 54.3%, but the homology of the N-terminal region was only 42%. The gene was expressed in E. coli as an independent unit from its own promoter, producing an immunoreactive protein of about 50 kDa on SDS-PAGE which reacted with antisera from laboratory animals and sera from human patients with acute Q fever. The study results suggest that the C. burnetii E2o enzyme may serve as a potential target antigen for diagnostic assays for Q fever. PMID- 10524792 TI - Apoptosis of endothelial cell line ECV304 persistently infected with Orientia tsutsugamushi. AB - Endothelial cells are major targets of Orientia tsutsugamushi. To examine the consequences of the infection of endothelial cells with O. tsutsugamushi, we used human endothelial cell line ECV304. Persistent infection was established and infected cultures could be maintained for over seven months without the addition of normal cells. The heavily infected cells became round and floated in the culture medium, harboring large numbers of organisms inside them. Some of the infected ECV304 cells showed features of apoptotic cells, as determined by the terminal deoxytransferase-mediated dUTP nick end-labeling reaction and DNA fragmentation. We also found that O. tsutsugamushi increased transcription of the mRNAs of proinflammatory cytokines such as IL-6 and IL-8. These results show the first evidence of in vitro-persistent infection by O. tsutsugamushi, which may be related to in vivo persistence reported previously. PMID- 10524793 TI - Crystallization of lipopolysaccharide from a Salmonella typhimurium semi-rough (SR) mutant. AB - Salmonella typhimurium SR-form lipopolysaccharide (LPS), consisting of a single repeating unit of the O-antigenic polysaccharide, linked to the R-core consisting of oligosaccharide that is, in turn, linked to lipid A, formed crystals whose shapes were hexagonal plates, discoids, and solid columns when precipitated by the addition of 2 volumes of 95% ethanol containing 375 mM MgCl2 and kept in 70% ethanol containing 250 mm MgCl2 at 4 C for 10 days. Among these crystals, the basic form is considered to be the hexagonal plates. Analyses of hexagonal plate crystals showed that they consist of hexagonal lattices with a lattice constant (a axis) of 4.62 A and longitudinal axis (c axis) of approximately 100 A. In X ray diffraction patterns in the low-angle region, crystals of S. typhimurium SR form LPS exhibited much less distinct reflections when compared with crystals of synthetic Escherichia coli-type lipid A. In contrast to the previous finding that S. minnesota S-form LPS possessing the O-antigenic polysaccharide does not crystallize under the same experimental conditions as used in the present study, the presence of a single repeating unit of the O-antigenic polysaccharide does not inhibit crystallization. PMID- 10524794 TI - Protective immune response to 16 kDa immunoreactive recombinant protein encoding the C-terminal VP1 portion of Foot and Mouth Disease Virus type Asia 1. AB - Recombinant protein of Foot and Mouth Disease Virus (FMDV) type Asia 1 corresponding to the C-terminal half of VP1 was expressed in Escherichia coli. As an alternative to the synthetic peptide, this selected C-terminal region was used as a protein vaccine in guinea pigs in order to study the immune response with various adjuvant formulations: immune stimulatory complexes (ISCOMs), Montanide ISA 206, Freund's incomplete adjuvant (FIA), lipopolysaccharide (LPS) and cytokine mixture. A primary dose of 40 microg/animal followed by a booster of the same dose was injected after a 21-day interval. The sera were collected at intervals of 21, 42 and 63 days after the booster. The humoral response to vaccine was monitored by sandwich enzyme-linked immunosorbent assay (ELISA) and a serum neutralization test (SNT). The guinea pig sera showed high titers both in ELISA and SNT, which could be protective. Further, irrespective of the adjuvant preparation used, the vaccine conferred protection against the challenge virus 105 days post-vaccination in 13 of 15 animals (86%). The results indicated that a combination of recombinant protein ISCOMs and Montanide ISA 206 would be a better choice for achieving early protective titers and longer lasting immunity and that the C-terminal half of the VP1 protein may be tried as a safe vaccine for secondary immunization. PMID- 10524795 TI - Cauliflower mosaic virus ORF III product forms a tetramer in planta: its implication in viral DNA folding during encapsidation. AB - Cauliflower mosaic virus (CaMV) open reading frame (ORF) III encodes a 15 kDa protein; the function of which is as yet unknown. This protein has non-sequence specific DNA binding activity and is associated with viral particles, suggesting that the ORF III product (P3) is involved in the folding of CaMV DNA during encapsidation. In this study, we demonstrated that P3 forms a tetramer in CaMV infected plants. A P3-related protein with an apparent molecular weight of 60 kDa was detected by Western blotting analysis using anti-P3 antiserum under non reducing conditions, while only 15 kDa P3 was detected under reducing conditions. Analysis of P3 using viable mutants with a 27-bp insertion in either ORF III or IV revealed that the 60 kDa protein was a tetramer of P3. The P3 tetramer co sedimented with viral coat protein in multiple fractions on sucrose gradient centrifugation, suggesting that P3 tetramer binds to mature and immature virions. These results strongly suggested that CaMV P3 forms a tetramer in planta and that disulfide bonds are involved in its formation and/or stabilization. The finding of P3 tetramer in planta suggested that viral DNA would be folded compactly by the interaction with multiple P3 molecules, which would form tetramers, while being packaged into the capsid shell. PMID- 10524796 TI - Prevalence of maternal cytomegalovirus (CMV) antibody and detection of CMV DNA in amniotic fluid. AB - The prevalence of cytomegalovirus (CMV) IgG antibody was determined in 573 pregnant women in the first trimester. The overall prevalence of CMV IgG antibody was 77.5%. The rate of seropositivity was 67.7% in women < 25 yr, and increased with age to 85.7% in women 40 yr. These results imply that young women in Japan are at increased risk for primary CMV infection during pregnancy and that congenital CMV infection rates might increase in the future. We conducted a prospective study of 75 pregnant women who underwent amniocentesis for various indications to determine if CMV DNA could be detected in the amniotic fluid. None had symptoms associated with CMV infection, CMV IgM antibody, or seroconversion to CMV IgG antibody during pregnancy. CMV DNA was not detected in the amniotic fluid using a polymerase chain reaction assay. The 65 fetuses, including 3 sets of twins, were followed through birth. CMV DNA was not detected in urine samples obtained within the first 2 weeks of life. In conclusion, CMV DNA was not detected in the amniotic fluid of women who did not have CMV infection. These results, however, suggest that the negative predictive value of prenatal amniotic fluid analysis is high and that the presence of CMV DNA in the amniotic fluid has clinical significance for the diagnosis of congenital CMV infection if detected in pregnant women. PMID- 10524797 TI - Production and partial characterization of antibody to cord factor (trehalose 6,6'-dimycolate) in mice. AB - Antibody production against the trehalose 6,6'-dimycolate (TDM, cord factor) of Rhodococcus ruber, a non-pathogenic species of the Actinomycetales group, was investigated in mice by repeated intraperitoneal injection of TDM in water-in-oil in-water micelles without carrier protein. The antigenic TDM was isolated and purified chromatographically from the chloroform-methanol extractable lipids of R. ruber. The hydrophobic moiety of this TDM was composed of two molecules of monoenoic or dienoic alpha-mycolic acids with a carbon chain length ranging from C44 to C48 centering at C46. To detect the antibody, an enzyme-linked immunosorbent assay (ELISA) system was employed using plastic plates coated with TDM. The antibody reacted against the TDM of R. ruber. The antibody was reactive in similar fashion against glycosyl monomycolates differing in the carbohydrate moiety, such as that of glucose mycolate (GM) and mannose mycolate (MM), obtained from R. ruber. Moreover, the antibody reacted against mycolic acid methyl ester itself when it was used as the antigen in ELISA, and trehalose did not absorb the antibody to TDM or inhibit the reaction. These results indicate that the epitope of TDM recognized by the antibody is mycolic acid, an extremely hydrophobic part of the molecule. Next, we prepared monoclonal anti-TDM antibody (moAb) in mice myeloma cells to examine its biological activities and the role of humoral immunity in mycobacterial infection. MoAb reacted against the TDM, glycosyl mycolate, and mycolic acid methyl ester in ELISA in the same manner as our polyclonal antibody did. The administration of moAb suppressed granuloma formation in the lungs, spleen, and liver induced by TDM and inhibited the production of interleukin-1 (IL-1) and chemotactic factor, which is reported to precede granuloma formation. PMID- 10524798 TI - Production of murine collagen-induced arthritis using Klebsiella pneumoniae O3 lipopolysaccharide as a potent immunological adjuvant. AB - Collagen-induced arthritis (CIA) was produced in mice with non H-2q and H-2r haplotypes by repeated immunization of porcine type-II collagen (CII) together with Klebsiella O3 lipopolysaccharide (KO3 LPS) as an immunological adjuvant. Histological changes that appeared in joints of repeatedly immunized mice were characterized by destruction of normal joint structure, synovial hyperplasia with proliferation of synovial cells, and infiltration of inflammatory cells. No such lesions were produced in mice receiving repeated injections of CII alone or KO3 LPS alone. Development of the humoral antibody and the delayed-type hypersensitivity to CII was exclusively found in mice immunized with the mixture of CII and KO3 LPS. It was therefore suggested that arthritis lesions induced by repeated immunization with the mixture of CII and KO3 LPS might be caused by an autoimmune mechanism, and that the experimental model might be useful for characterization of human rheumatoid arthritis (RA). PMID- 10524799 TI - Evaluation of AFLP, a high-resolution DNA fingerprinting method, as a tool for molecular subtyping of enterohemorrhagic Escherichia coli O157:H7 isolates. AB - The amplified fragment-length polymorphism (AFLPTM) technique is based on the selective PCR amplification of restriction fragments. We investigated the utility of AFLP in the molecular subtyping of enterohemorrhagic Escherichia coli serotype O157:H7 isolates. We analyzed a total of 46 isolates of E. coli O157:H7 along with other serotypes, O26:H11, 0114:H19 and 0119:NT. Isolates of E. coli O157:H7 derived from the same outbreak showed an identical AFLP-banding pattern and were subtyped into the same group, giving results almost consistent with those of a pulsed-field gel electrophoresis (PFGE) study, while other serotypes showed clearly different patterns from those of E. coli O157:H7. These results suggest that the AFLP technique has potential as an alternative tool for the molecular epidemiology of E. coli O157:H7. PMID- 10524800 TI - Limited stress response in Streptococcus pneumoniae. AB - In Streptococcus pneumoniae, heat shock induces the synthesis of 65-, 73-, and 84 kDa proteins, and ethanol shock induces a 104-kDa protein. In this study, the 65 , 84-, and 104-kDa proteins were identified as members of the GroEL, ClpL and alcohol dehydrogenase families, respectively, and the general properties of the stress response of S. pneumoniae to several other stresses were characterized. However, several stresses which are known to induce stress responses in Escherichia coli and Bacillus subtilis failed to induce any high molecular weight heat-shock proteins (HSPs) such as GroEL and DnaK homologues. A minor temperature shift from 30 to 37 C triggered induction of the homologues of DnaK and GroEL of E. coli. These features may provide a foundation for evaluating the role of heat shock proteins relative to the physiology and pathogenesis of pneumococcus. PMID- 10524801 TI - Chitin synthase 2 gene sequence of Malassezia species. AB - Nucleotide sequences of the chitin synthase 2 (CHS2) gene of seven species, Malassezia furfur, M. globosa, M. obtusa, M. pachydermatis, M. restricta, M. slooffiae and M. sympodialis, were analyzed for their phylogenetic relationship. About 620-bp genomic DNA fragments of the CHS2 gene were amplified from these Malassezia species by polymerase chain reaction (PCR) and sequenced. The CHS2 nucleotide sequences of these Malassezia species showed more than 95% similarity between the species. A phylogenetic analysis of the nucleotide sequences of CHS2 gene fragments of seven Malassezia species revealed that the species were genetically distinct from each other. PMID- 10524802 TI - Development of restriction fragment-length polymorphism method to differentiate five subtypes of feline immunodeficiency virus. AB - Feline immunodeficiency virus (FIV) isolates have been classified into five subtypes (A to E) based on the sequences of the env variable V3 to V5 region. In this study, we sequenced a partial gag region of 4 and 3 isolates belonging to subtypes C and E, respectively. Phylogenetic analysis revealed that the branching pattern based on the region was similar to that based on the env V3 to V5 region. Here, we propose a protocol to differentiate five subtypes by polymerase chain reaction amplifying 329 bp within the region followed by restriction fragment length polymorphism analysis using four restriction enzymes. PMID- 10524803 TI - Lung cancer epidemiology and genetics. AB - Lung cancer is the number one cause of cancer death for all citizens of the United States regardless of gender, race, or ethnic background. Surgery, the only curative treatment available, is feasible in only 35% of all patients with lung cancer. But even in this group of patients, most will die of recurrent disease. Treatment advances have been modest at best, and the 5-year survival rate of approximately 15% has not changed appreciably in the past two decades. Population projections indicate a continuation of the lung cancer epidemic among men and a rise among women, in whom rates of smoking have remained relatively high. It is estimated that 90% of all lung cancers are smoking related, and gender, race, and family history are important risk modifiers. Several promising markers of lung cancer susceptibility have recently been identified. However, the relative contribution of individual intrinsic factors (genetic markers, family history, race, gender) on lung cancer susceptibility in the context of complex extrinsic factors (behavior, environmental exposure, socioeconomic status) is unknown and requires further study for more informed screening and primary prevention. PMID- 10524804 TI - Radiologic staging of lung cancer. AB - Preoperative tumor staging in patients with known or suspected non-small cell lung cancer is generally performed using contrast enhanced chest computed tomography (CT) (including the adrenal glands). Abdominal CT is generally unnecessary, given the low frequency of isolated liver metastases. The role of MRI is limited, and it is used mainly as a problem solving tool in certain specific situations. A CT showing no mediastinal lymph node enlargement usually oviates preoperative mediastinal lymph node sampling, with certain exceptions. If enlarged mediastinal lymph nodes are demonstrated at CT, then CT may be used to direct preoperative lymph node sampling via transbronchoscopic Wang needle biopsy, mediastinoscopy, mediastinotomy, or video assisted thoracoscopy. PMID- 10524805 TI - Non-small cell lung cancer: FDG-PET imaging. AB - Positron emission tomography is a clinically useful imaging modality that complements conventional radiologic studies in the evaluation of focal pulmonary opacities, lung cancer staging, tumor recurrence, treatment response, and prognosis. PMID- 10524806 TI - Medical treatment of lung cancer. AB - Lung cancer is the leading cause of cancer deaths for both men and women in the United States. Most patients present with late stages of disease, rendering them incurable. Despite this dismal outcome, there is optimism; modern treatment for lung cancer has been shown to increase survival, improve quality of life, and be cost effective. This article reviews the current treatment strategies for non small cell lung cancer and small cell lung cancer as well as discusses future treatments, including the exciting biologic agents that are in clinical trials. PMID- 10524807 TI - Surgical management of lung cancer. AB - Surgery offers the best opportunity for long term survival for a patient with lung cancer. In this review, the principles guiding surgical intervention are discussed. PMID- 10524808 TI - A CT sign of chronic pulmonary arterial hypertension: the ratio of main pulmonary artery to aortic diameter. AB - The aim of this study was to determine whether the ratio of the diameters of the main pulmonary artery and of the ascending aorta (rPA), as assessed on computed tomography (CT), is predictive of pulmonary arterial hypertension (PAH). We undertook a retrospective review of 50 patients with a wide range of pulmonary and cardiovascular diseases, who had undergone both chest CT and pulmonary arterial pressure measurements at right heart catheterization. Two independent observers made measurements of the diameter of the main pulmonary artery and of the ascending aorta on a single defined CT section. Body surface area (BSA, n = 48), pulmonary arteriolar resistance (n = 39), total lung capacity (n = 40), and aortic pressures (n = 50) were also recorded. rPA and pulmonary arterial diameter (dPA) were positively related to mean pulmonary artery pressure (Rs = 0.74, p < 0.0005 for both analyses). For patients younger than 50 years of age, mean pulmonary artery pressure correlated more strongly with rPA than dPA (Rs = 0.77, p < 0.00005, compared with Rs = 0.59, p < 0.005); and vice versa for patients older than 50 years of age (Rs = 0.63, p < 0.005, compared with Rs = 0.75, p < 0.00005). Using a mean pulmonary artery pressure greater than 20 mm Hg as indicative of PAH and a value of rPA > 1, the sensitivity, specificity, and positive and negative predictive values for determining PAH were 70% (26/37), 92% (12/13), 96% (26/27), and 52% (12/23), respectively. On multivariate analysis, rPA was positively related to mean pulmonary artery pressure (p < 0.0005), and negatively related to age (p < 0.0005), but was not related to BSA. By contrast, dPA showed some dependency on BSA (p < 0.0005), as well as on mean pulmonary arterial pressure. In patients younger than 50 years of age, we have found a strong correlation between rPA and mean pulmonary artery pressure in a heterogeneous study population, and this relationship is independent of BSA and sex. The presence of the sign "rPA > 1" is simple in practical CT reading to determine; if this is identified, there is a very high probability of pulmonary arterial hypertension, and clinicians should be alerted to this possibility. PMID- 10524810 TI - Thoracic venous anatomy delineated by malpositioned central venous catheters on plain chest films. AB - The aim of this essay was to demonstrate the thoracic venous anatomy as delineated by malpositioned central venous catheters on plain chest radiographs. We therefore used the didactic advantage of clinically inadvertent catheter positions. This approach was chosen to illustrate venous anatomy with plain chest radiographs, and, thereby, to recognize malpositions promptly on the modality with which positions of central venous catheters is routinely performed. PMID- 10524809 TI - Visual quantitation and observer variation of signs of small airways disease at inspiratory and expiratory CT. AB - Areas of decreased pulmonary attenuation representing small airways disease can be identified on computed tomography (CT). The objective was to quantify differences between inspiratory and expiratory CT for the detection of signs of small airways disease by four observers. Observer variation and the superiority of a fine versus a coarse grading system were also evaluated. Inspiratory and expiratory CT scans of 106 patients with conditions characterized by small airways disease and 19 healthy individuals were assessed by four observers. The extent of decreased attenuation was scored on a fine scale to the nearest 5% and also semiquantitatively on a coarser 5-point scale. Decreased attenuation was more extensive on expiratory CT (median. 6.7%; 0-76.7%) than on inspiratory CT (median, 3.8%; 0-81.7%). The fine scoring system had unacceptable interobserver variation (coefficient of variation, 80% for inspiratory CT, 70% for expiratory CT). The semiquantitative system had acceptable interobserver agreement (inspiratory CT k(w) = 0.64; expiratory CT, k(w) = 0.69) and good intra-observer agreement (inspiratory CT, k(w) = 0.80; expiratory CT, k(w) = 0.64). The major CT sign of small airways disease is more confidently quantified on expiratory CT. A fine scoring system is associated with unacceptable observer variation, and a coarse semiquantitative system is more suitable for quantitative studies of small airways disease. PMID- 10524811 TI - Radiologic anatomy of ventricular assist devices. AB - Ventricular assist devices (VAD) allow for long-term circulatory support of patients with end-stage heart failure. With the increasing duration of circulatory support, diagnostic imaging plays an important role in the management of patients on a VAD. The aim of our review was to analyze the radiologic features of different VADs. From 1987 to 1996, 319 patients (mean age 42 years, range 3 to 74 years) were treated with a VAD. A Berlin Heart VAD was implanted in 263 of the patients, the univentricular Baxtor Novacor was implanted in three patients, and the univentricular CI Heartmate was implanted in 19 patients. All patients were studied by serial chest radiographs. In addition, 70 patients underwent computed tomography (CT), and five patients underwent electron beam CT. The Berlin Heart VAD was used as a biventricular support system in 218 patients. In all cases, the position of the wire-directed cannulae was identified on the chest radiographs, while the exact position of the cannula tip could be visualized by CT only. The plastic cannulae of both the Novacor and the Heartmate were not discernible on radiographs, but required CT for evaluation. Computed tomography also resolved the metal components of the pumps. The titanium-made pump housing of the Heartmate caused beam-hardening artefacts that might conceal fluid accumulations in the pump pocket. Computed tomography is the standard of reference for examinations of cannula position, pump position, and pump components of ventricular assist devices. PMID- 10524812 TI - Posttraumatic thoracic splenosis and chronic aortic pseudoaneurysm. AB - Thoracic splenosis is the autotransplantation of splenic tissue into the pleural cavity, usually following traumatic injury that simultaneously involves the spleen and diaphragm. Intrathoracic splenosis is an uncommonly reported phenomenon. We present a case of intrathoracic splenosis in a patient with a chronic aortic pseudoaneurysm, both detected, incidentally, three decades after the traumatic event. The diagnosis of thoracic splenosis can be confirmed noninvasively, using 99mTc sulfur colloid nuclear scintigraphy. PMID- 10524813 TI - Pyrofluid inhalation in "fire-eaters": sequential findings on CT. AB - We report the sequential computed tomography (CT) findings in two fire-eaters after accidental inhalation of pyrofluid. The initial chest radiographic findings were ambiguous and the interpretation of the radiographs was biased by clinical history unrelated to fire eating. On CT, pneumatoceles were the major findings in both patients. The pneumatoceles resolved rapidly, leaving only minimal scarring. Our cases illustrate the sequential evolution of pneumatoceles in fire-eaters after the inhalation of pyrofluid and documents the rapidity with which the lesions regress. The rare accidental inhalation of pyrofluid in fire-eaters may produce a puzzling clinical and radiographic picture and can be confused with other lung disorders. PMID- 10524814 TI - Mycobacterium xenopi infection of a 50-year-old oil plombage complicated by bronchopleural and pleurocutaneous fistulas. AB - We report the rare combination of simultaneous bronchopleural and pleurocutaneous fistulas 50 years after oil plombage, together with infection of both the plombage and the contralateral lung with Mycobacterium xenopi. Our case documents imaging patterns of complex fistula formation and subsequent infection resulting from oil plombage. Our case also emphasizes the infectious potential of Mycobacterium xenopi. PMID- 10524815 TI - False radiographic appearance of a right bronchial stump following extended right pneumonectomy. AB - The authors describe three cases in which postoperative frontal chest radiographs following extended right pneumonectomy showed a right hilar lucency producing the false appearance of a residual main bronchus that is shown by additional studies to represent a dilated esophagus. PMID- 10524816 TI - Giant liposarcoma of the esophagus: radiological findings. AB - This case of an esophageal liposarcoma illustrates a polypoid lesion within the esophagus that extended from the left pyriform sinus to the distal esophagus above the gastric cardia. Contrast-enhanced computed tomography (CT) and magnetic resonance imaging (MRI) showed an inhomogenously-enhancing intraluminal mass, while video-fluoroscopy revealed that the mass was adherent to the esophageal wall and was associated with esophageal dilatation and diminished peristalsis. This ninth reported case of esophageal liposarcoma is the first described where preoperative radiologic studies and endoscopy showed broad fixation of the tumor to the esophageal wall. PMID- 10524817 TI - Bioabsorbable suture anchor (co-polymer 85/15 D,L lactide/glycolide) implanted in bone: correlation of physical/mechanical properties, magnetic resonance imaging, and histological response. AB - A novel bioabsorbable suture anchor has been introduced for shoulder rotator cuff surgical repair made of the co-polymer 85/15 D,L lactide/glycolide. Previous clinical reports on the use of this material in anterior cruciate ligament reconstruction have described intraosseous edema at various time intervals following implantation. The purpose of this study was to analyze the implant's loss of physical properties and to correlate magnetic resonance imaging (MRI) finding with gross and histological observations at various time intervals after intraosseous implantation in the experimental animal. Six drill holes were made in the tibias of 11 dogs. The spherical implant was placed in 5 of the drill holes and the sixth was preserved as a sham control. The dogs were killed at 3, 4, 6, 9, 12, and 26 weeks for gross and microscopic inspection. Correlative MRIs were taken from the 4-, 12-, and 26-week specimens. Gross inspection showed that the overlying soft tissue healed to bone in 3 weeks. The implants were surrounded by new bone by 6 weeks. The implants maintained gross physical integrity for 6 to 12 weeks. Histologically, there was minimal inflammatory response to the degrading implant. The implant site had been completely replaced by bone at 12 weeks. Correlative MRI showed edema adjacent to the implant sites, but there was no correlative inflammation or cyst formation through the time necessary for complete absorption of the implant. Correlative MRI identified and differentiated the image of the intact and degrading implant. PMID- 10524818 TI - Serial magnetic resonance imaging evaluation of operative site after fixation of patellar tendon graft with bioabsorbable interference screws in anterior cruciate ligament reconstruction. AB - Magnetic resonance imaging (MRI) is accepted as the imaging procedure of choice for showing internal derangement of the knee. In contrast to metal implants, bioabsorbable interference screws do not produce an artifact and provide an opportunity to expand the evaluation of the postoperative anterior cruciate ligament (ACL) ligament repair. There is the potential to evaluate the implant, the graft, the adjacent tissue, and the surgically created bone tunnels. The purpose of this study was to evaluate with MRI the postoperative site of ACL patellar tendon autografts in which bioabsorbable screws were used for fixation. It was hypothesized that a time line of bone tissue changes resulting from this type of surgery could be developed based on the expanded evaluation of MRI. From January 1993 through October 1997, 270 patients underwent surgical repair of a disrupted native ACL. There were 173 men 97 women; the average age was 25.1 years, (range, 17 to 50 years). There were 155 right knees and 115 left knees. In addition to the conventional postoperative clinical assessment and plain film radiographs, opportunistic MRIs were obtained with the patient's permission. The examinations were performed at different postoperative intervals from the third postoperative day to 4 years postoperatively. A total 206 MRIs from various time intervals were available for study. The study protocol was designed to look for loss of integrity of the screws, adjacent fluid collection, tunnel widening, and tunnel healing or narrowing. The hypothesis was substantiated in this study. The use of MRI provided observations not available by other imaging methods. The absence of metal implants for fixation provided an opportunity to examine the adjacent tissue in detail and to form a time line of the tissue response in this type of surgery. PMID- 10524819 TI - Suture anchors--update 1999. AB - New suture anchors continue to become available. Our prior reports on the pullout strength of over 50 different anchors is supplemented by a similar test conducted on 25 additional new anchors. This anchor comparison, using an established protocol in fresh porcine femurs, recorded failure strength, failure mode (anchor pullout, suture eyelet cutout, or wire breakage), eyelet size, minor and major diameters, and drill hole sizes. These new anchors were tested in diaphyseal cortex, metaphyseal cortex, and a cancellous trough. Tensile stress parallel to the axis of insertion was applied at a rate of 12.5 mm/sec by an Instron 1321 until failure and mean anchor failure strengths were calculated. Anchors tested included DePuy 4.5 prototypes D1, D2 Catera 4.5, and D3; DePuy 3.5 prototypes D4- Catera 3.5, D5, and D6; Mainstay 2.7, 3.5, 4.5; ROC EZ 2.8, EZ 3.5, and XS 3.5; Ultrafix RC and Ultrafix MiniMite; 1.3 MicroMitek, Panalok 3.5, and Tacit 2.0; Umbrella Harpoon; PeBA 2.8, 4.0, 6.5; and Stryker 1.9, 2.7, 3.4, and 4.5 prototypes. Screw anchors still tend to have higher values, but for the newer nonscrew designs this distinction is less apparent. The new biodegradable anchors were all composed of poly L-lactic acid suggesting a trend away from other polymers, and these new biodegradable anchors showed load-to-failure strengths comparable to others in their class. All anchors were stronger than the suture for which they are designed to accommodate. PMID- 10524820 TI - Biomechanical test comparing the load to failure of the biodegradable meniscus arrow versus meniscal suture. AB - The biodegradable Meniscus Arrow (Bionx, Blue Bell, PA) is a newly introduced method for repair of meniscal injury. However, because of a lack of studies, there is still little understanding of its biomechanical behavior. Biomechanical test was carried out to evaluate the load to failure and failure modes of the Meniscus Arrow and to compare the results with those of three traditionally used methods. Thirty-five lateral menisci obtained from Yorkshire pigs were incised longitudinally, simulating a peripheral longitudinal tear, and were repaired by means of Meniscus Arrows (one-point and two-point fixation), as well as with the techniques of knot-end, horizontal, and vertical suture using No. I PDS II monofilament suture. Tensile strength testing was performed according to each repair method with the Instron tensometer (Model No. 5569; Instron, Canton, MA) and its results were analyzed using a computerized statistical program. The average maximal tensile strengths were 113.9 +/- 14.6 N in vertical suture, 75.1 +/- 18.4 N in horizontal suture, 53.9 +/- 6.4 N in knot-end suture, 38.3 +/- 4.3 N in one-point fixation Meniscus Arrow repair, and 56.5 +/- 3.5 N in two-point fixation Meniscus Arrow repair. The initial failure strength for two-point fixation Meniscus Arrow repair is comparable to that of a knot-end suture. Following the results of this study, meticulous caution should be taken when meniscal tears are repaired with Meniscus Arrows. PMID- 10524821 TI - Magnetic resonance imaging of tunnel placement in posterior cruciate ligament reconstruction. AB - The aim of this study was to define a reproducible method for evaluating posterior cruciate ligament (PCL) reconstructions using magnetic resonance imaging (MRI). A 2-fold investigation was performed. In part I, the "footprints" of an intact PCL were located on MRI and their coordinates were defined. Measurements were made on the images of 50 subjects using axial, coronal, and sagittal planes. Interobserver variability was calculated by averaging the measurements of the 2 reviewers and using the Kappa coefficient. Three points of reference were located: tibial attachment on the tibial axial plane, and two femoral attachments on the sagittal and coronal oblique planes. In part II, stability of 20 PCL reconstructions with a bone-patellar tendon-bone (BPTB) autograft were evaluated and scored using the IKDC evaluation form after a 2-year follow-up. Stability was evaluated clinically and instrumentally using a KT-2000 arthrometer at 89 N with the knee flexed at a neutral quadriceps knee angle of approximately 70 degrees . Seven cases were graded A (0 to 2 mm), 11 graded B (3 to 5 mm), and 2 graded C (6 to 10 mm). All patients had an MRI after an average of 16 months (range, 12 to 24 months, 2 SD). The previous measurements from part I of the study were used to make a correlation between achieved stability and tunnel location. A 1-factor analysis of variance (ANOVA), nonparametric ANOVA, and the Fisher Exact test were used to determine if clinical outcome of the 3 groups was influenced by graft placement. At MRI evaluation, excessive deep placement was observed in 4 cases and a correlation between improper femoral tunnel location and stability was statistically significant (P < .05). A correct placement of tibial tunnel was observed in all patients. In our analysis, proper location of the femoral tunnel seems to be more critical and difficult to achieve than tibial tunnel placement, probably because of the lack of specific anatomic landmarks during surgery. PMID- 10524822 TI - Quantitative analysis of human cruciate ligament insertions. AB - The objective of this study was to provide quantitative data on the insertion sites of the cruciate ligaments. In the first part of the study, we determined the shapes and sizes of the insertions of the anterior and posterior cruciate ligaments (ACL and PCL), and further compared these data with the midsubstance cross-sectional areas of the ligaments. The cross-sectional area of the ACL and PCL midsubstance of 5 human knees was measured using a laser micrometer system. The insertion sites of each ligament were then digitized and the 2-dimensional insertion site areas were determined. Relative to the ligament midsubstance, the PCL tibial and femoral insertions were approximately 3 times larger, whereas those of the ACL were over 3.5 times larger. In the second part of the study, the ACLs and PCLs of 10 knees were each divided into their 2 components and the areas of each insertion were determined. Each component was approximately 50% of the total ligament insertion area and no significant difference between the 2 could be shown. PMID- 10524823 TI - Femoral tunnel position in anterior cruciate ligament reconstruction using three techniques. A cadaver study. AB - The possibility of achieving correct deep femoral tunnel positioning during anterior cruciate ligament (ACL) reconstruction with the double incision technique (DI), the transtibial technique (TT), and the anteromedial technique (AM) was evaluated in 30 cadaver knees. A reference hole was made just deep to the insertion of the anteromedial bundle of the ACL through an anteromedial arthrotomy. In the DI technique, a Kirshner wire was inserted outside-in using a rear entry C guide. In the TT and AM techniques, the K-wire was inserted inside out through the tibial tunnel and through the arthrotomy, respectively. The reference hole could be achieved with each technique. Using lateral radiographs, the superficial aspect of the intra-articular exit of the femoral tunnel was found to be located on average at 36%, 36%, and 34% of the width of the condyles from the posterior margin (NS). None of the holes was more anterior than 40%. In conclusion, a deep femoral tunnel positioning could be achieved with each technique. The choice of technique must be based on the surgeon's preference and clinical results. PMID- 10524825 TI - Arthroscopic removal of bullet fragments from the subtalar joint. AB - A case of arthroscopic removal of a bullet fragment from the subtalar joint and the calcaneus is presented. The bullet fragments impinged on the fibula, limiting eversion and causing pain. The fragments were removed both arthroscopically and through open incision. The patient noted complete relief of pain and improved range of motion within 1 week, and complete recovery soon thereafter. PMID- 10524824 TI - The cyclops lesion: a cause of diminished knee extension after rupture of the anterior cruciate ligament. AB - Four patients presented with persistent diminution of knee motion after rupture of the anterior cruciate ligament with a novel lesion as the cause. Each had participated in an aggressive rehabilitation program for a minimum of 2 months with emphasis on regaining full range of knee motion. Because chronic impairment of knee extension can be disabling, in those who did not regain full range of motion, arthroscopy of the knee ensued. All had a lesion in the intercondylar notch near the tibial insertion of the anterior cruciate ligament that acted as a mechanical obstruction to full knee extension. Grossly and histologically, these were similar to the cyclops lesion that also has been shown to cause loss of knee extension after anterior cruciate ligament reconstruction. Arthroscopic debridement of the cyclops lesion and manual manipulation of the knee under anesthesia lead to restoration of full knee extension in all knees. In 1 other knee with chronic instability after anterior cruciate ligament rupture, the cyclops lesion was present but was very small and was not associated with diminished knee extension. When loss of full extension persists for 2 months after anterior cruciate ligament disruption despite aggressive rehabilitation, the presence of a cyclops lesion should be considered. PMID- 10524826 TI - Tibial tuberosity avulsion fracture combined with meniscal tear. AB - Avulsion fractures of the tibial tuberosity are uncommon injuries. They usually occur during athletic activities in adolescents. The classification of these injuries has been divided into three types. Only two cases of avulsion fractures of tibial tuberosity have previously been reported with associated damages to menisci. We report a type III fracture of the tibial tuberosity associated with tear of the medial meniscus. PMID- 10524827 TI - Inflammatory foreign-body reaction to an arthroscopic bioabsorbable meniscal arrow repair. AB - Various arthroscopic meniscal repair techniques have been developed in recent years to preserve meniscal function. We report the case of a patient with a failed arthroscopic meniscal repair demonstrating an inflammatory foreign-body reaction to bioabsorbable meniscal arrows. PMID- 10524828 TI - Common peroneal nerve palsy following knee arthroscopy. AB - We report the case of a 43-year-old woman who underwent knee arthroscopy. Postoperatively, she developed a lesion of the common peroneal nerve, which was confirmed by neurophysiological studies. Exploration showed the nerve to be in continuity and externally undamaged. At review 17 months later, there was incomplete recovery. We believe this lesion was caused by a traction injury related to patient positioning, which has not been reported previously. PMID- 10524829 TI - Intra-articular detachment of the Endobutton more than 18 months after anterior cruciate ligament reconstruction. AB - We report a case of detachment of an Endobutton (Acufex Microsurgical, Mansfield MA) used for femoral fixation of a reconstructed anterior cruciate ligament. The Endobutton, which was confirmed to be in place on the suprapatellar space of the femur by radiograph 18 months postoperatively, was found in the popliteal space by radiograph 25 months after surgery. This is a rare complication, but our case suggests that the Endobutton should not be fixed too distal close to the femoral groove. PMID- 10524830 TI - Wrist arthroscopy and dislocation of the radiocarpal joint without fracture. AB - The authors report a rare case of dorsal dislocation of the radiocarpal joint without any bony lesion associated. The traumatic cause was a high energy motorbike accident. Fractures of the other limbs were associated. The authors report the clinical, radiological, and arthroscopic features. Wrist arthroscopy showed a complete tear of all the extrinsic ligaments, a radial avulsion of the triangular fibrocartilage complex, and the integrity of the intracarpal ligaments, which guided the treatment. The dislocation was treated by closed reduction and radiocarpal pinning. The authors propose wrist arthroscopy in radiocarpal dislocation for diagnosis of soft tissue and cartilaginous lesions to guide the treatment (close or open). PMID- 10524831 TI - Complete transection of the median and radial nerves during arthroscopic release of post-traumatic elbow contracture. AB - Arthroscopic debridement and capsular release was performed in a 57-year-old woman because of post-traumatic stiffness in the dominant right elbow joint. During this procedure, the median and radial nerves were completely transected. A few recent reports of small series have described encouraging results after arthroscopic capsular release of post-traumatic elbow contracture, but the present case demonstrates the inherent risk of damage to neurovascular structures. PMID- 10524832 TI - Spur reformation after arthroscopic acromioplasty. AB - Rotator cuff pathology has been associated with a hooked acromial morphology. Impingement syndrome has traditionally been considered to be the result of bony encroachment into the subacromial space. This report of a spur recurrence after acromioplasty presents evidence that acromial morphology may be a reactive change attributable to primary rotator cuff insufficiency. PMID- 10524833 TI - Anterior cruciate ligament reconstruction in the young patient without violation of the epiphyseal plate. AB - A technique of arthroscopic anterior cruciate ligament reconstruction that does not disturb the epiphyseal plate in the young patient with open physis is presented. A cryopreserved bone-Achilles tendon allograft was incorporated by an interference screw fixation to the bone plug in the tibia and an over-the-top positioning of the tendon on the femoral side. For this procedure, the minimal patient age that the thickness of the epiphysis can accept an interference screw greater than 15 mm in length is 8 years. An intra-articular reconstruction of anterior cruciate ligament with the cryopreserved Achilles allograft using our technique is safe and recommendable for young patients with open physis. PMID- 10524834 TI - Overlap endoscopic SLAP lesion repair. AB - The fixation of the superior labrum using the punch-chop needle overlap technique (Aeratec Inc, Uniondale, NY) for reattaching torn labral tissue to bone allows ease of suture placement for type II SLAP lesion repair with fixation overlapping the superior rim of the glenoid and without the use of anchors. The technique presented includes preparation of the superior glenoid rim, drilling of the glenoid tunnels for the punch needles, and peripheral suturing of the labrum. PMID- 10524835 TI - Lamotrigine and the treatment of bipolar disorder. Introduction. PMID- 10524836 TI - Controlled trials in bipolar I depression: focus on switch rates and efficacy. AB - Until recently, the rate at which patients switch from bipolar depression to the manic or hypomanic phase of the disorder during treatment with antidepressant medications was poorly defined. The completion of three large-scale, double-blind controlled trials in bipolar I depression has improved understanding of this phenomenon. The low switching rates observed in these studies of lamotrigine, paroxetine and moclobemide may indicate a special application of these drugs in the management of patients prone to antidepressant-induced switching. These studies also confirm prior suggestions that tricyclic antidepressants present the highest risk of switching. At present there is no consensus over the optimal definition of switching. Standardising the definition may lead to improvements in the clinical management of bipolar disorder. PMID- 10524837 TI - Lamotrigine in the treatment of bipolar depression. AB - Several case reports and open studies have reported the efficacy of lamotrigine in bipolar depression. A randomised placebo-controlled 7-week study comparing two doses of lamotrigine with placebo in 195 patients with moderate to severe bipolar depression has now been completed. Lamotrigine was superior to placebo after 3 weeks as assessed by changes in the Montgomery-Asberg Depression Rating Scale (MADRS). A response, defined as more than 50% improvement on the MADRS occurred in 56 and 48% of the lamotrigine 200 and 50 mg/day groups, respectively, compared with 29% for placebo (P<0.05). There was no evidence that lamotrigine destabilised mood or precipitated mania. Tolerability was good and there were no cases of serious rashes. Preliminary results from an ongoing study also indicate that lamotrigine is more effective than gabapentin in bipolar depression. In conclusion, lamotrigine is effective in alleviating bipolar depression, without causing mood destabilisation. Slow dosage escalation yields good tolerability. PMID- 10524838 TI - Lamotrigine and the treatment of mania in bipolar disorder. AB - Anticonvulsants, including valproate and carbamazepine, have established efficacy in the treatment of mania. The anticonvulsant, lamotrigine. has been reported to have antimanic and antidepressant efficacy, and mood-stabilising effects in case reports and preliminary open trials. The efficacy and tolerability of lamotrigine has been compared with olanzapine and lithium in a randomised, prospective, controlled fashion over a period of 4 weeks treatment in a total of 45 hospitalised patients with DSM-IV-defined mania. Significant improvements of a similar magnitude were observed for all treatment groups and lamotrigine was well tolerated. Mechanisms of action proposed to explain the antimanic activity of lamotrigine include inhibition of voltage-sensitive and use-dependent sodium channels, inhibition of glutamate release and calcium channel blockade. Platelet studies have indicated supersensitivity of glutamate receptors and increased intracellular calcium concentrations in patients with mania. Further clinical and mechanistic studies of lamotrigine use in mania are warranted. PMID- 10524839 TI - Prophylaxis of bipolar disorder: how and who should we treat in the long term? AB - Bipolar disorder is a devastating and chronic mood disorder, which can require life-long treatment. The vast majority of patients will suffer relapse of symptoms in the absence of effective therapy. Of those patients receiving treatment, compliance to medication regimens is poor. Non-compliance, when associated with lithium treatment in particular, increases the risk of recurrence of illness. Problems associated with withdrawal serve as powerful stimuli to develop alternatives to lithium monotherapy. Conventional placebo-controlled studies of treatments are difficult in patients with bipolar disorder. Large scale, pragmatic and clinically relevant trials should be employed to assess existing and novel treatments for bipolar disorder. These can only develop out of genuine clinician and patient uncertainty and the creation of a trial culture in everyday practice. PMID- 10524840 TI - Planning nephron-sparing renal surgery using 3D helical CT angiography. PMID- 10524841 TI - CT evaluation of hepatic injury following proton beam irradiation: appearance, enhancement, and 3D size reduction pattern. AB - PURPOSE: The purpose of this study was to investigate the long-term imaging appearances of hepatic injury following proton beam irradiation. The time attenuation curves, time of appearance and recovery, and 3D size reduction pattern are described in patients of different ages and genders with different irradiation doses, irradiated portals, and Child groups. METHOD: Forty-six patients with hepatocellular carcinoma underwent 50 to 84 Gy proton beam irradiation in periods of 14-52 days. CT including noncontrast and dynamic study was performed every 3 months starting 3 weeks after the end of irradiation. The 3D volume measurement of areas of radiation-induced hepatic injury was performed through incremental dynamic CT images in every follow-up study. CT follow-up study of the patients was done for 12-76 months. RESULTS: Radiation-induced hepatic injury was observed as low attenuation areas on noncontrast CT and enhanced areas on dynamic study in the regions corresponding to the irradiation portals. Of our cases, 67.5% showed the appearance of radiation hepatitis in 3-4 weeks and 95.3% in 3-4 months after the end of irradiation. In both periods, there was a significant delay in the female patients. The time-attenuation curve showed an early and prolonged enhancement of the irradiated regions. The volume reduction pattern of the injured areas was found to be longstanding, exponential, and directed from periphery to the center. CONCLUSION: Early appearance of radiation-induced hepatic injury was found only to be gender dependent, with a tendency to occur with higher irradiated doses; no other parameters affected this phenomenon in our cases. Disappearance of the injured areas, if present, takes a long time (at least 42 months). PMID- 10524842 TI - Gadolinium-enhanced 3D MRA prior to isolated hepatic perfusion for metastases. AB - PURPOSE: Isolated hepatic perfusion (IHP) is a new treatment for patients with isolated unresectable liver metastases, which can result in a partial or complete response in approximately 75% of patients. Preoperative knowledge of hepatic arterial anatomy is important to adequately perfuse the liver. Digital subtraction angiography (DSA) is currently used to identify the hepatic arterial anatomy. The purpose of this study was to determine if MR angiography (MRA) could replace DSA prior to IHP. METHOD: Twenty-seven patients scheduled to undergo IHP underwent MRA with a contrast-enhanced 3D time-of-flight gradient echo sequence. Both maximal intensity projections (MIPs) and source coronal images were used to evaluate the images. The results of the MRA were interpreted by two readers who were blinded to the surgical results. The first 17 patients also underwent DSA, and a separate comparison was made with those results. Anatomy was characterized as either normal hepatic arteries (NHAs), normal vasculature with an accessory left hepatic artery (aLHA), or a replaced right hepatic artery (rRHA). RESULTS: MRA correctly detected all 22 patients with NHAs but also identified 6 aLHAs, of which only 2 were confirmed surgically. MRA correctly detected all five rRHAs. MIP images alone accurately depicted the hepatic arterial anatomy in only 9 of 27 (33%), usually because significant vessels were not visualized or their origin could not be determined. Source coronal images were required to accurately determine the anatomy in all patients. Among the 17 patients who underwent DSA, MRA detected 14 of 14 with NHA and 3 of 3 with rRHA. Six aLHAs were identified by MRA and five were confirmed by DSA. CONCLUSION: Enhanced 3D MRA is an accurate method of depicting the hepatic arterial supply. In comparison to surgery, MRA overestimates the number of aLHAs, but this may be because these small vessels are not detected at surgery. Based on the results of this study, DSA has been replaced by MRA in the planning of IHP at our institution. A better display of MRA images is needed as MIP images were usually insensitive for the small caliber arteries supplying the liver. PMID- 10524843 TI - Intrahepatic peripheral cholangiocarcinoma: comparison of dynamic CT and dynamic MRI. AB - PURPOSE: The purpose of this work was to compare dynamic MRI (D-MRI) with dynamic CT (D-CT) for the diagnosis of peripheral cholangiocarcinoma (PCC) of the liver. METHOD: Twenty patients with PCC underwent both D-CT and D-MRI during the early, middle, and delayed phase after contrast medium administration. The findings from D-MRI were compared with those from D-CT. RESULTS: D-CT and D-MRI exhibited a similar tumoral enhancement pattern, and this enhancement was more conspicuous on D-MRI. A wedge-like enhancement area peripheral to the tumor was observed in 9 (45%) patients on D-CT and 11 (55%) patients on D-MRI. Ductal dilatation was found in 13 (65%) patients on both techniques. Vascular involvement and extrahepatic invasion were seen in nine (45%) and two (10%) patients, respectively. The relationship of the tumor to the vessels and surrounding organs was more easily evaluated on D-CT. CONCLUSION: Both D-CT and D-MRI can provide important information for the diagnosis of PCC. D-CT is better than D-MRI for demonstrating vascular involvement and extrahepatic invasion. D-MRI gives more conspicuous enhancement. PMID- 10524844 TI - CT features of abdominal manifestations of primary antiphospholipid syndrome. AB - PURPOSE: The purpose of this work was to evaluate the CT features of the abdominal manifestations of primary antiphospholipid syndrome (PAPS). METHOD: Of the 32 patients who were confirmed to have PAPS among 751 patients with elevated antiphospholipid antibodies during a 2 year period, we retrospectively reviewed the 14 patients who underwent abdominal CT. The clinical indications for abdominal CT included abdominal pain, abdominal distension, or lower leg swelling. CT findings were analyzed with regard to the abdominal vascular system and abdominal organ involvement patterns as well as ancillary findings. RESULTS: Of the 14 patients with PAPS, 10 had involvement of the venous system (72%), 2 of the arterial system (14%), and 2 of both systems (14%). Of the 12 patients who had venous system involvement, 4 had thrombosis in the inferior vena cava (IVC), 2 in both the IVC and the hepatic vein, 1 in the IVC and splenic and portal veins, 1 in the IVC and hepatic and adrenal veins, 1 in the hepatic, portal, and renal veins, and 3 in the portal and superior mesenteric veins. Budd-Chiari syndrome developed in five of the nine patients who had thrombosis of the IVC or hepatic vein. Arterial thrombosis was noted in four patients, hepatic artery in two, aorta in one, renal artery in one, pancreatic arcade in one, and splenic artery in one, with infarct of multiple organs including the liver, jejunum, colon, kidney, and adrenal gland. Seven of the 14 patients (50%) manifested thrombosis or infarct of multiple extra-abdominal organs. CONCLUSION: PAPS should be included in the differential diagnosis when CT demonstrates infarcts in multiple organs or patients have recurrent episodes of venous or arterial thrombosis. PMID- 10524845 TI - MRI of intralesional hemolysis in focal nodular hyperplasia of the liver. AB - A case of Kasabach-Merritt syndrome caused by focal nodular hyperplasia of the liver is presented with atypical magnetic resonance findings due to intratumoral hemosiderin deposition. The high sensitivity of magnetic resonance imaging for iron served to identify the site of hemolysis in this patient with Kasabach Merritt syndrome. PMID- 10524846 TI - Perineal angiomyofibroblastoma: CT and MR findings with pathologic correlation. AB - We present a case of a perineal angiomyofibroblastoma (AMFB) arising in the right perirectal fossa in a middle-aged woman, documented with CT and MRI. Compounding the rarity of the entity, this case is unique as it is the first radiological report illustrating the CT and MR features of this recently clinicopathologically described neoplasm. PMID- 10524848 TI - Retroperitoneal paragonimiasis: a case of ectopic paragonimiasis presenting as periureteral masses. AB - We describe a case of retroperitoneal paragonimiasis presenting as periureteral masses. CT showed a conglomerate of enhancing nodules with subtle low attenuation in the center at the left iliac fossa and clustered, ring-like, enhancing lesions at the left renal hilum. When a retroperitoneal conglomerate of ring-like, enhancing lesions in association with pleuropulmonary disease suggestive of paragonimiasis can be found in endemic regions or in migrants from those regions, one may expect ectopic-retroperitoneal paragonimiasis. PMID- 10524849 TI - Peritoneal location of fascioliasis mimicking a peritoneal carcinomatosis. PMID- 10524847 TI - Radiologic features of intrahepatic bile duct adenoma: a look at the surface of the liver. AB - We report the radiological features of intrahepatic bile duct adenoma (BDA) in three patients. BDA was shown as a small mass located in the peripheral region of the liver with each imaging modality: a hypervascular mass on angiography and a mass appearing as early nodular enhancement found disproportionately evident compared with their small size and distinct delayed or prolonged enhancement on CT. BDA should be included in the diseases to be differentiated from hypervascular hepatic tumors. PMID- 10524850 TI - Isolated traumatic rupture of the cisterna chyli: CT diagnosis. PMID- 10524851 TI - Imaging of dermoid cysts with foci of immature tissue. PMID- 10524852 TI - Foot pain after a plantar fasciotomy: an MR analysis to determine potential causes. AB - PURPOSE: The purpose of this work was to determine potential causes of foot pain in patients who have had a surgical release of the plantar fascia for treatment of fasciitis. METHOD: We studied 17 patients (15 women, 2 men; age range 22-59 years, mean 40 years) with foot pain after undergoing a fasciotomy. Fourteen unilateral and three bilateral procedures accounted for the 20 ankles evaluated. Mean duration after surgery was 22 months (range 3-53 months). Each patient was instructed to localize the pain to a region of the foot; classify the pain as new onset, persistent, or recurrent; and characterize it as to the action that produced the greatest pain. T1-weighted sagittal and dual-echo T2-weighted images in the sagittal, coronal, and axial planes were obtained in a 1.5 T magnet. The MR studies were evaluated for abnormalities of the plantar fascia, perifascial soft tissues, tendons, and osseous structures. RESULTS: The plantar fascia appeared thick in all ankles (mean 8.0 mm, range 6-12 mm). A total of 25 symptomatic sites were assessed. An acute plantar fascia rupture explained plantar symptoms in two feet. In another 16 feet (12 with plantar heel pain and 4 with nonspecific heel pain), 6 had documentation of acute plantar fasciitis and 9 demonstrated perifascial edema. Of the latter nine feet, five demonstrated abnormalities of the posterior tibialis, peroneus longus, and peroneus brevis tendons. The pain localized to the medial arch in six feet; five feet had abnormalities of the posterior tibialis tendon and one foot demonstrated edema in the flexor digitorum brevis muscle. The pain localized to the lateral midfoot in one foot, which had a cuboid stress fracture. CONCLUSION: The cause of foot pain in patients who had a plantar fasciotomy appeared to be multifactorial. Three likely causes of pain were identified: persistent or recurrent acute plantar fasciitis, pathology related to arch instability, and structural failure from overload. PMID- 10524853 TI - MR evaluation of subscapularis tears. AB - PURPOSE: The purpose of this work was to describe the MR appearance of tears of the subscapularis tendon and compare the usefulness of different imaging planes as well as note the association of subscapularis tears with other rotator cuff tears and biceps tendon dislocations. METHOD: MR studies at 1.5 T over an 8 year interval were retrospectively assessed for the presence of a rotator cuff tear and/or tear of the subscapularis tendon. Images that showed a subscapularis tear were reviewed for the presence of a visible tear separately on the axial, coronal, and sagittal images. The MR studies were also evaluated for associated tears of the supraspinatus, infraspinatus, and teres minor muscles as well as biceps tendon dislocation and the "naked humerus sign" on coronal images. Last, clinical records and surgical reports were reviewed. RESULTS: Forty-five (2%) of 2,167 rotator cuff tears involved the subscapularis; 27% were partial and 73% were complete tears. Tears were best seen in the sagittal oblique plane. Almost all subscapularis tears were an extension of typical rotator cuff tears: supraspinatus in 35 patients (79%), extending into infraspinatus tears in 25 (56%) and into teres minor tears in 2 patients (4%). Bicipital dislocations were seen in 22 patients (49%), and three complete tears of the biceps (7%) were noted as well. The naked humerus sign was demonstrated in 31 patients (69%). Surgical reports that confirmed the MR findings were available for 15 patients. CONCLUSION: About 2% of rotator cuff tears involve the subscapularis tendon. Most subscapularis tears are extensions of supraspinatus tears and frequently involve the biceps tendon. PMID- 10524855 TI - Chondroblastoma: MR characteristics with pathologic correlation. AB - PURPOSE: The purpose of this study was to describe the MR findings of chondroblastoma with pathologic correlation. METHOD: In 22 patients with pathologically proven chondroblastoma, MR signal characteristics were correlated with pathological findings. RESULTS: On T2-weighted images, 12 (55%) lesions were hyperintense with hypointense areas in 9 lesions, whereas 10 (45%) were hypointense. Therefore, 19 of 22 (86%) lesions with pathologic correlation had hypointense areas entirely (n = 10) or partly (n = 9) on T2-weighted images. On gadolinium-enhanced images, 13 (59%) lesions showed lobular enhancement and 9 (41%) showed marginal and septal enhancement. Low signal intensity on T2-weighted MR images was most strongly associated with an abundance of immature chondroid matrix, hypercellularity of the chondroblasts, calcifications, and hemosiderin on histology. CONCLUSION: Chondroblastoma was found to show hypointense portions on T2-weighted images. Signal intensity on T1- and T2-weighted MR images in chondroblastoma was dependent on the amounts of histopathological components. PMID- 10524854 TI - Central pseudodefect of the talus: a potential ankle MR interpretation pitfall. AB - PURPOSE: The purpose of our study was to outline the MR features of the central pseudodefect of the talus (a normal finding that can simulate an osteochondral lesion on ankle MR studies), assess the prevalence of the central pseudodefect of the talus, and provide insight into the origin of this misleading MR appearance. METHOD: We retrospectively evaluated 31 ankle MR studies in 10 asymptomatic volunteers and 21 consecutive patients for the presence of the central pseudodefect of the talus. None of the patients had a history of trauma to the ankle. The signal, size, and shape of the pseudodefect were documented in each patient. The sagittal images were cross-referenced with the axial and coronal images in all patients in whom the central pseudodefect was identified. RESULTS: Six volunteers (60%) and 13 patients (62%) showed a curvilinear band in the middle third of the talus on far medial sagittal images, consistent with the central pseudodefect of the talus. The band measured 8-15 x 3-8 mm (mean 11 x 4 mm) and was hypointense on T1 and STIR pulse sequences. In two cases, the pseudodefect was subchondral; in the rest, it was found a few millimeters below the articular surface. On cross-referenced axial and coronal images, the band corresponded to the talar insertion site of the deep tibiotalar fibers of the deltoid ligament. CONCLUSION: The central pseudodefect of the talus is a common finding that is produced by the insertion of the tibiotalar fibers of the deltoid ligament into the talus. Familiarity with its appearance is necessary to avoid misinterpreting it as an osteochondral lesion of the talus. PMID- 10524856 TI - Intraosseous ganglion of the metatarsal bone. AB - We describe a rare case of intraosseous ganglion arising in the metatarsal bone. Radiographs revealed an osteolytic lesion with a fracture in the third metatarsal bone. A biopsied specimen exhibited hyaline fibrous tissue with marked myxoid change. Gadolinium-enhanced MRI, which revealed the network-like enhancement of the rim of the lesion and polycystic lesions adjacent to the joint, was helpful in making a diagnosis of intraosseous ganglion. PMID- 10524857 TI - Squalene-induced extrinsic lipoid pneumonia: serial radiologic findings in nine patients. AB - PURPOSE: The purpose of this work was to demonstrate the initial and follow-up radiologic findings of squalene-induced extrinsic lipoid pneumonia. METHOD: Follow-up chest radiographs (n = 9) and high-resolution CT scans (n = 3) as well as initial radiographs (n = 9) and CT scans (n = 8) were obtained in nine patients with squalene-induced extrinsic lipoid pneumonia. The serial radiologic findings were analyzed retrospectively by three chest radiologists, focusing on the pattern and distribution of parenchymal abnormalities. RESULTS: The most frequent pattern of parenchymal abnormalities on chest radiograph was areas of ground-glass opacity (n = 9, bilateral 6), followed by consolidation (n = 7, bilateral 3) and poorly defined small nodules (n = 4, bilateral 2). The abnormalities were distributed in the right lower lung (n = 9), left lower lung (n = 6), and right middle lung (n = 6) zones. Initial CT scans (n = 8) demonstrated bilateral areas of ground-glass attenuation (n = 8), poorly defined centrilobular nodules (n = 8), crazy paving (n = 6), and consolidation (n = 3). The abnormalities were distributed in the right middle lobe (n = 8) and in both lower lobes (n = 5). Follow-up chest radiograph (n = 9) showed complete disappearance (n = 2) and decrease (n = 7) in the extent of the parenchymal abnormalities. Follow-up CT scans (n = 3) demonstrated decrease (n = 2) and no change (n = 1) in the extent of the abnormalities. CONCLUSION: Squalene-induced extrinsic lipoid pneumonia most commonly appears as areas of ground-glass attenuation mixed with poorly defined centrilobular nodules and crazy paving on CT, being distributed mainly in the right middle and both lower lobes. The lesions are indolent and remain after cessation of squalene ingestion. PMID- 10524858 TI - CT findings of pulmonary tuberculosis presenting as segmental consolidation. AB - PURPOSE: The purpose of our study was to determine specific CT findings of tuberculous pneumonia presenting as segmental or lobar consolidation along with a pathologic review of specimens with similar radiographic patterns. METHOD: CT findings of 45 cases of proven tuberculous pneumonia and 21 proven nontuberculous pneumonia were compared. Pathologic findings of five surgically resected tuberculous pneumonia cases were also investigated. The presence of fluid bronchogram (linear, branching shadow of fluid attenuation) and inner low attenuation/cavitation in the area of consolidation, luminal dilatation, and wall thickening of proximal bronchi were the main points sought on CT scan. In addition, the presence of bronchogenic dissemination, lymph node enlargement, and pleural lesions was also checked for in the unaffected area of both lungs. RESULTS: The following bronchial changes were seen in the tuberculous pneumonia and nontuberculous pneumonia groups, respectively: fluid bronchogram in 68.9 and 23.8% (p < 0.05), bronchial luminal dilatation in 60.0 and 23.8% (p < 0.05), and bronchial wall thickening of the proximal airway leading to the area of consolidation in 52.8 and 7% (p < 0.05). Bronchogenic dissemination outside the consolidation appeared in 88.9 and 52.4% (p < 0.05), respectively. In the tuberculous pneumonia group, lymph node enlargement and pleural reaction were seen in 55.6 and 35.6%, respectively, but in 42.9 and 57.1% in the nontuberculous pneumonia group (p > 0.05). Histologically, tuberculous pneumonia showed either bronchioles containing inflammatory exudates and submucosal granuloma or alveoli containing aggregates of alveolar macrophages or cellular debris. CONCLUSION: Fluid bronchogram in the area of homogeneous consolidation, bronchial luminal dilatation, and bronchial wall thickening of the proximal airway were the bronchial changes more significantly prominent in the tuberculous pneumonia group. We suspect that these findings may represent tuberculous bronchitis in small airways. PMID- 10524859 TI - Estimation of gas and tissue lung volumes by MRI: functional approach of lung imaging. AB - PURPOSE: The purpose of this work was to assess the accuracy of MRI for the determination of lung gas and tissue volumes. METHOD: Fifteen healthy subjects underwent MRI of the thorax and pulmonary function tests [vital capacity (VC) and total lung capacity (TLC)] in the supine position. MR examinations were performed at inspiration and expiration. Lung volumes were measured by a previously validated technique on phantoms. Both individual and total lung volumes and capacities were calculated. MRI total vital capacity (VC(MRI)) was compared with spirometric vital capacity (VC(SP)). Capacities were correlated to lung volumes. Tissue volume (V(T)) was estimated as the difference between the total lung volume at full inspiration and the TLC. RESULTS: No significant difference was seen between VC(MRI) and VC(SP). Individual capacities were well correlated (r = 0.9) to static volume at full inspiration. The V(T) was estimated to be 836+/-393 ml. CONCLUSION: This preliminary study demonstrates that MRI can accurately estimate lung gas and tissue volumes. The proposed approach appears well suited for functional imaging of the lung. PMID- 10524860 TI - "Crazy paving appearance" on high resolution CT in various diseases. AB - PURPOSE: The purpose of this work was to demonstrate the variety of causes of crazy-paving appearance (CPA) on high resolution CT (HRCT). METHOD: To identify cases exhibiting CPA (ground-glass opacity with superimposed interlobular septal thickening and intralobular interstitial thickening) on HRCT, we prospectively searched for them over a period of 29 months. RESULTS: We identified 10 cases of CPA on HRCT, including 4 Pneumocystis carinii pneumonia, 1 alveolar proteinosis, 1 usual interstitial pneumonia, 1 pulmonary hemorrhage, 1 acute radiation pneumonitis, 1 adult respiratory distress syndrome, and 1 drug-induced pneumonitis. CONCLUSION: CPA can result from a variety of diseases. When we encounter CPA on HRCT, clinical information is necessary for differentiation of these entities. PMID- 10524862 TI - Mucous gland adenoma of the bronchus: CT findings in two patients. AB - Mucous gland adenoma of the bronchus is a truly benign, well defined, intraluminal mass that manifests on CT with air-meniscus sign or abutting the bronchus. We report the CT findings of mucous gland adenoma in two patients. PMID- 10524861 TI - Analysis of time-density curves of contrast media for improvement of chest dynamic incremental CT. AB - PURPOSE: The goal of this work was to analyze time-density curves (TDCs) of contrast media (CM) in the mediastinal vasculature to optimize chest dynamic incremental CT. METHOD: Forty-three patients were injected with nonionic CM into the forearm veins with injection rates (ml/s), durations (s), and total amounts (ml) of 2.0, 20, and 40 (protocol 1); 4.0, 20, and 80 (protocol 2); and 2.0, 40, and 80 (protocol 3). TDCs were obtained for the pulmonary trunk (PA) and ascending (AA) and descending (DA) aorta from dynamic scans. Areas under the curves (AUCs) of TDCs for imaginary 30 s scans were evaluated. RESULTS: AUC peaks were obtained after 10, 17, and 19 s (PA, AA and DA; protocol 1; 9, 16, and 18 s (protocol 2); and 18, 25, and 28 s (protocol 3) delay time. CONCLUSION: Better chest dynamic incremental CT would be expected with scan midpoints a little after the end of injection of CM. PMID- 10524863 TI - Lymphangitic carcinomatosis from prostate carcinoma. PMID- 10524864 TI - Liposarcoma of the breast arising within a phyllodes tumor. PMID- 10524865 TI - Rapid imaging of olfaction by functional MRI (fMRI): identification of presence and type of hyposmia. AB - PURPOSE: Our goal was to develop a rapid, simple, near-real-time method of functional MRI (fMRI) to measure brain activation in response to olfactory stimuli, to use it to identify patients with smell loss (hyposmia), and to differentiate their types of hyposmia. METHOD: fMRI was obtained in 16 patients with Type I hyposmia (who could detect but not recognize odors), 5 patients with Type II hyposmia (who could both detect and recognize odors, albeit with less than normal acuity), and 2 volunteers with normal olfactory acuity by use of a rapid echo planar imaging technique in which one coronal brain section from the anterior cortical region was studied and a single olfactory stimulus was used. Actual scanning time performed by a variation of methods previously published required 26 s. Three patients with Type I hyposmia were treated with theophylline 250-500 mg for 4-6 months and were studied before and after treatment. RESULTS: Brain activation in response to olfactory stimuli was demonstrated using a new, rapid, and simple fMRI technique. Patients with Type I hyposmia had less activation than patients with Type II hyposmia. Both patient groups had less activation than normal volunteers. Activation in patients with Type I hyposmia was essentially absent from regions of the middle frontal, orbitofrontal, and temporal cortex and was totally absent in regions of inferior frontal, insular, and cingulate cortex. Activation in patients with Type II hyposmia was greatest in the middle frontal cortex and the orbitofrontal cortex bilaterally and was present in regions of inferior frontal, temporal, and cingulate cortex. Each patient with Type I hyposmia treated with theophylline had improved smell function to Type II hyposmia and after treatment demonstrated activation in inferior frontal and cingulate cortex bilaterally, whereas before treatment, no activation in these regions was apparent. CONCLUSION: We describe a simple, rapid technique that can be used in a practical clinical setting to identify patients with hyposmia and to differentiate patients with different types of olfactory loss. These studies confirm the presence and classification of patients with Type I and Type II hyposmia. Results of this study suggest that regions of the frontal cortex may act to guide or direct olfactory signals to other brain areas such as temporal and cingulate regions. Although these latter regions are involved with olfactory recognition, their role in olfactory memory, olfactory meaning, and attention needs to be considered. PMID- 10524866 TI - Normal canals at the fundus of the internal auditory canal: CT evaluation. AB - PURPOSE: Knowledge of the normal anatomy of the four bony canals located at the fundus of the internal auditory canal (IAC) is necessary during evaluation of temporal bone trauma, congenital anomalies affecting the individual nerves, and some neuro-otologic surgeries. The purpose of this work was therefore to characterize the normal appearance of the four bony canals and to measure their dimensions. METHOD: A retrospective study was performed using CT studies of the temporal bones in 50 patients to identify and characterize the bony canals for the labyrinthine segment of the facial nerve (BCFN), superior vestibular nerve (BCSVN), cochlear nerve (BCNC), and the inferior vestibular nerve (singular canal; SC) located at the fundus of the IAC. All the patients underwent high resolution temporal bone CT for evaluation of uncomplicated inflammatory (n = 49) and neoplastic (n = 1) diseases involving the temporal bone. CT studies were done using 1-mm-thick contiguous sections in axial and coronal planes. Measurements of the canals were performed by one radiologist. No patient had a prior history of trauma, vertigo, and sensorineural hearing loss or facial nerve paralysis. RESULTS: The BCFN, BCSVN, and BCNC were identified in all studies, whereas the SC was seen in 93% of studies. The BCFN, BCSVN, and BCNC arise from the fundus of the IAC, whereas the SC arises medial to the fundus. Mean +/- SD measurements (in mm) of the length and width were as follows: BCFN = 2.92+/-0.48 and 0.91+/-0.28; BCSVN = 2.36+/-0.53 and 0.89+/-0.28; BCNC = 0.93+/-0.21 and 2.13+/-0.44; and SC = 3.22+/-0.73 and 0.50+/-0.14. CONCLUSION: These small canals are routinely visualized on thin section (1 mm) CT of the temporal bone and should not be confused with fractures. This study provides baseline measurements that may be used to evaluate congenital anomalies of these canals. These data may also be helpful in the presurgical evaluation of patients undergoing singular neurectomies for benign positional vertigo. PMID- 10524867 TI - Intracranial papillary endothelial hyperplasia: occurrence of a case after surgery and radiosurgery. AB - Papillary endothelial hyperplasia (PEH) is considered a form of endothelial proliferation rather than a true neoplasm and is usually located in the skin or subcutis. We report a case of intracranial PEH that occurred after surgery for glioma and subsequent radiosurgery. CT and MR revealed an enhancing extra-axial mass located left posterolateral to the brainstem. Intracranial PEH is rare; to our knowledge, development of an intracranial PEH after surgery and radiosurgery has not been previously reported. PMID- 10524868 TI - Computing the centerline of a colon: a robust and efficient method based on 3D skeletons. AB - We present a robust and efficient algorithm for calculating the centerline of a computer-generated colon model created from helical CT image data. The centerline is an essential aid for navigating through complex anatomy such as the colon. Our algorithm involves three steps. In the first step, we generate a 3D skeleton of the binary colon volume using a fast topological thinning algorithm. In the second step, we employ a graph search algorithm to remove extra loops and branches. These loops and branches are caused by holes in the object that are artifacts produced during image segmentation. In the final step, we compute a smooth representation of the centerline by approximating the skeleton with cubic B-splines. This final step is necessary because the skeleton contains many abrupt changes in direction due to the discrete nature of image data. The user supplies two endpoints for the centerline; otherwise, the algorithm is fully automated. Experimental results demonstrate that the algorithm is not only robust but also efficient. PMID- 10524869 TI - Xenon versus ceramics: a comparison of two CT X-ray detector systems. AB - PURPOSE: The purpose of this work was to compare image quality in phantom and patient CT scans acquired by xenon and ceramic CT detector systems. METHOD: High and low contrast resolution and image noise were determined with a standard CT phantom for both detector systems. In patient CT images, the effect on image noise was measured in anatomical regions of interest in the head, lumbar spine, chest, and abdomen. RESULTS: In phantom studies, image noise was significantly lower using ceramic versus xenon detectors. Also, in images of the head and lumbar spine, the signal-to-noise ratio was significantly higher with ceramic than with xenon detectors. In chest scans, ceramic significantly reduced beam hardening artifacts caused by the thoracic spine. However, in abdominal images, the signal-to-noise ratio was not significantly different between ceramic and xenon detector systems. CONCLUSION: For reduced image noise in CT images of the head, lumbar spine, and chest and high resolution CT, ceramic detector systems appear to be superior to xenon detector systems. PMID- 10524870 TI - Comparison and evaluation of rigid, affine, and nonrigid registration of breast MR images. AB - PURPOSE: A new nonrigid registration method, designed to reduce the effect of movement artifact in subtraction images from breast MR, is compared with existing rigid and affine registration methods. METHOD: Nonrigid registration was compared with rigid and affine registration methods and unregistered images using 54 gadolinium-enhanced 3D breast MR data sets. Twenty-seven data sets had been previously reported normal, and 27 contained a histologically proven carcinoma. The comparison was based on visual assessment and ranking by two radiologists. RESULTS: When analyzed by two radiologists independently, all three registration methods gave better-quality subtraction images than unregistered images (p < 0.01), but nonrigid registration gave significantly better results than the rigid and affine registration methods (p < 0.01). There was no significant difference between rigid and affine registration methods. CONCLUSION: Nonrigid registration significantly reduces the effects of movement artifact in subtracted contrast enhanced breast MRI. This may enable better visualization of small tumors and those within a glandular breast. PMID- 10524871 TI - Aunt Minnie's Corner. Well-differentiated retroperitoneal liposarcoma. PMID- 10524872 TI - von Recklinghausen's disease and pheochromocytomas. AB - PURPOSE: We review the literature and characterize the clinical findings of von Recklinghausen's associated pheochromocytoma. MATERIALS AND METHODS: A Grateful Med search for the years 1966 to 1999 was performed on the subjects, "von Recklinghausen" and "neurofibromatosis." Articles from the Grateful Med search were then reviewed to identify older publications. Of 325 articles 118 are included in this review. RESULTS: Pheochromocytomas have been clinically identified in 0.1 to 5.7% of patients with von Recklinghausen's disease. Mean patient age was 42 years (range 1.5 to 74) in 87 women and 61 men at presentation with pheochromocytoma. Of the 148 patients 84% had solitary adrenal tumors, 9.6% bilateral adrenal disease and 6.1% ectopic pheochromocytomas. Symptoms related to pheochromocytoma or hypertension were noted in 78% of the patients. Tumors secreted epinephrine and norepinephrine, and 87% demonstrated metaiodobenzylguanidine uptake. Of the 148 patients 6% died during pregnancy or a medical procedure, or due to hypertensive crisis without apparent provocation, 8.8% had gastrointestinal carcinoid tumors and 11.5% had metastases or local invasion from pheochromocytoma. CONCLUSIONS: Pheochromocytomas occur in a small but defined number of patients with von Recklinghausen's disease, and can be associated with significant morbidity and mortality if not detected. Screening of patients with von Recklinghausen's disease and hypertension or before provocative procedures or pregnancy seems to be indicated. PMID- 10524873 TI - The effect of high grade prostatic intraepithelial neoplasia on serum total and percentage of free prostate specific antigen levels. AB - PURPOSE: It is established that the percentage of free prostate specific antigen (PSA) in serum is low in patients with prostate cancer. An unanswered question is whether a low percentage of free PSA can be explained by high grade prostatic intraepithelial neoplasia alone. We compared the percentage of free PSA in men with high grade prostatic intraepithelial neoplasia alone, prostate cancer, benign prostatic hyperplasia (BPH) and a normal prostate (that is normal digital rectal examination and PSA less than or equal to 2.5 ng./ml.). MATERIALS AND METHODS: From October 1994 through December 1997, 48 men were diagnosed with high grade prostatic intraepithelial neoplasia without concomitant prostate cancer. Of these men 43 with a mean age plus or minus standard deviation of 67.4 +/- 7.8 years comprised our study group. To date none has been diagnosed with cancer during followup. Serum free and total PSA levels were measured, and the percentage of free PSA was calculated. The percentage of free PSA in the 43 men was compared to that in 50 with prostate cancer (mean age 65.4 +/- 7.8 years), 50 with biopsy proved BPH (67 +/- 7) and 43 with a normal prostate (61 +/- 8). RESULTS: There was no significant difference in mean total serum PSA in patients with high grade prostatic intraepithelial neoplasia, prostate cancer or BPH. The percentage of free PSA was significantly lower in patients with prostate cancer (14.9 +/- 6.5%) than those with high grade prostatic intraepithelial neoplasia (20.8 +/- 7.1%), BPH (20.1 +/- 7.3%) or a normal prostate (27.7 +/- 12.2%). There was also no significant difference in the percentage of free PSA between men with high grade prostatic intraepithelial neoplasia (20.8 +/- 7.1%) and those with BPH (20.1 +/- 7.3%). Additionally, men with a normal prostate had a higher percentage of free PSA (27.7%) than those with BPH (20.1%), high grade prostatic intraepithelial neoplasia (20.8%) or prostate cancer (14.9%). CONCLUSIONS: The percentages of free PSA in men with high grade prostatic intraepithelial neoplasia and BPH are similar, and significantly higher than those found in men with prostate cancer. PMID- 10524874 TI - Simultaneous versus staged bilateral extracorporeal shock wave lithotripsy: long term effect on renal function. AB - PURPOSE: We determine whether there is a clinically significant difference in the long-term effect on renal function following simultaneous versus staged bilateral extracorporeal shock wave lithotripsy (ESWL). MATERIALS AND METHODS: Between July 1986 and October 1995, 360 patients underwent treatment for bilateral renal calculi with ESWL using a Dornier HM-3 lithotriptor. Of the patients 319 had both kidneys treated simultaneously (simultaneous group) and the remaining 41 were treated in a planned, staged fashion (staged group) with the procedures separated by 3 to 20 weeks (mean 6.0). Followup data of at least 1 year were available for 49 men and 30 women (mean age 52.4) in the simultaneous group, and 8 and 12 (mean age 45.1), respectively, in the staged group. Mean stone burden was 0.70 cm.2 on the right and 0.87 on the left side in the simultaneous group, and 1.6 and 1.5, respectively, in the staged group. RESULTS: Patients with at least 1 calculus greater than 0.5 cm.2 treated simultaneously received a mean of 1,386 shock waves to the right and 1,637 to the left side, and the staged group received a mean of 1,802 and 2,094, respectively. Mean serum creatinine was 1.03 mg./dl. before and 1.04 after treatment (normal 0.7 to 1.4 mg./dl.) in the simultaneous group, representing a mean increase of 0.01 at followup of 1.0 to 10.5 years (mean 3.7), and 0.88 and 0.88, respectively, in the staged group, representing a decrease of 0.005 at followup of 1.0 to 7.2 years (mean 3.2). The effects of simultaneous versus staged ESWL on renal function as measured by serum creatinine were not statistically significant using a multiple regression model which controlled for the effects of stone burden, number of shock waves, patient age, pretreatment serum creatinine and length of followup (p = 0.19). CONCLUSIONS: There is no clinically apparent difference in the long-term effect on renal function for patients with bilateral renal calculi treated with ESWL in a simultaneous versus staged fashion. PMID- 10524875 TI - Complications of retrograde balloon cautery endopyelotomy. AB - PURPOSE: Adult ureteropelvic junction obstruction is increasingly managed with endoscopic techniques. Retrograde balloon cautery endopyelotomy is quick, requires minimal hospital stay and allows most patients a rapid return to work. The complication rate of retrograde balloon cautery endopyelotomy ranges from 13 to 34%, with vascular injury in 0 to 16% of patients. We report 5 uncommon complications, including 4 vascular injuries, that clinicians should be familiar with when using this technique. MATERIALS AND METHODS: We reviewed 52 retrograde endoscopic endopyelotomy procedures performed during a 5-year period. There were 5 uncommon complications. RESULTS: Accessory lower pole renal artery injuries occurred in 3 patients, 1 of whom presented 12 days after endopyelotomy. Embolization was successfully performed in all 3 cases and none had subsequent hypertension. In 1 case a right ovarian vein laceration was not evident on preoperative or postoperative angiography. Emergency post-embolization abdominal exploration revealed a 2 mm. injury to the right ovarian vein before entering the right renal vein close to the ureteropelvic junction incision. Nephrectomy and ovarian vein ligature were curative. In 1 case the electrocautery wire broke intracorporeally after firing, resulting in a bobby pin-like configuration. Successful removal was accomplished by twisting the catheter and wrapping the wire around the tip, enabling atraumatic removal. CONCLUSIONS: Retrograde balloon cautery endopyelotomy is an emerging technology with potential adverse outcomes. The complications we noted are complex and potentially life threatening. Awareness of these complications may help avoid poor outcomes and expedite appropriate treatment. PMID- 10524876 TI - Early nasogastric tube removal combined with metoclopramide after radical cystectomy and urinary diversion. AB - PURPOSE: Prolonged nasogastric decompression increases pulmonary complications by inhibiting clearance of respiratory secretions. The literature supports early nasogastric tube removal following bowel resection. Metoclopramide stimulates bowel activity, promoting return of function. We examined combining early nasogastric tube removal with metoclopramide after radical cystectomy. MATERIALS AND METHODS: From 1994 to 1996, 27 prospective cystectomy patients received intravenous metoclopramide (metoclopramide group) combined with early nasogastric tube removal (less than 24 hours). A total of 54 concurrent cystectomy controls received no metoclopramide and nasogastric tubes remained until return of normal bowel function. RESULTS: Preoperative and perioperative factors were comparable between the 2 groups. Nasogastric tubes were removed from 78% of the metoclopramide group in less than 24 hours, 11% on day 2 and 11% on day 3 compared to none on day 1, 50% on day 2 and 50% on day 3 or greater in controls. The metoclopramide group had a more rapid return of normal bowel sounds (2.9 versus 4.0 days, p = 0.0002) and earlier tolerance of solid food (6.7 versus 7.9 days, p = 0.04). Nasogastric tube replacement was required in 3 of 27 metoclopramide cases versus 5 of 54 controls. Atelectasis occurred more often in the control group (33 versus 15%). There were no bowel related complications in the metoclopramide group but partial small bowel obstruction in 2 controls was treated conservatively. CONCLUSIONS: This preliminary study suggests that combining intravenous metoclopramide with early nasogastric tube removal after cystectomy and urinary diversion may reduce postoperative atelectasis and speed return of bowel function while posing no danger to the small bowel anastomosis. This regimen may result in fewer complications and shorter hospitalizations, translating into lower costs without compromising quality of care. PMID- 10524877 TI - Upper tract recurrences following radical cystectomy: an analysis of prognostic factors, recurrence pattern and stage at presentation. AB - PURPOSE: We study the incidence and pattern of upper tract recurrences following radical cystectomy for bladder cancer, and analyze the prognostic factors. MATERIALS AND METHODS: A retrospective study was performed on 529 patients who underwent radical cystectomy and urinary diversion at Memorial Sloan-Kettering Cancer Center between July 1989 and June 1997. Data related to upper tract recurrence were analyzed. RESULTS: Of the 529 patients 16 (3%) had upper tract recurrence. Median followup was 16.9 months for the entire group and 49.1 months for patients with upper tract recurrence, with a median time to recurrence of 37.2 months. Of 12 upper tract recurrences 7 (58%) were locally advanced at surgery (p3a or greater with or without lymph node metastasis) and 5 of 16 patients with recurrence (31.3%) had bilateral tumors (2 synchronous and 3 metachronous). Overall survival from the time of diagnosis of upper tract recurrence after radical cystectomy was poor, with a median of 10 months (confidence interval 1 to 19). CONCLUSIONS: The incidence of upper tract recurrence following radical cystectomy is low (3%). However, the incidence of bilateral tumors (31.3%) and locally advanced stage at the time of operation (58%) is higher than expected for upper tract tumors in the general population. Survival of patients with upper tract recurrence is poor, with a median of 10 months. PMID- 10524878 TI - Tailored laminectomy: a new technique for neuromodulator implantation. AB - PURPOSE: Neuromodulation of sacral roots is an alternative mode of therapy for patients with urge incontinence or detrusor hypocontractility. We investigated the effects of sacral (S3) nerve stimulation in patients using a new surgical approach for sacral neuromodulator implantation. Modification of the implantation method with sacral laminectomy and bilateral electrode placement led to distinct improvement of stimulation, positioning and dislocation. We developed tailored laminectomy for bilateral neuromodulator electrode implantation to minimize surgical trauma. MATERIALS AND METHODS: Tailored laminectomy was performed in 6 patients with urge incontinence and 3 with a hypocontractile detrusor. After making a 10 cm. longitudinal skin incision we exposed the spinous processes of S2 and S3. Instead of complete 2-level laminectomy, only 2 oval laminectomy holes were made with a high speed ball drill. An electrode fixation hole was drilled at the edge of the laminectomy window and the wire was fixed with nonabsorbable suture material. RESULTS: In patients with idiopathic urge incontinence (followup 12.5 months, range 7 to 18) the number of leaks decreased from 7.2 to 0 daily and functional bladder capacity increased from 298 to 352 ml. In patients with a hypocontractile detrusor (followup 10.5 months, range 6 to 20) detrusor pressure increased during voiding from 12 to 34 cm. water and post-void residual decreased from 350 to 58 ml. Average surgery time was 2 hours 15 minutes. In 1 case a seroma developed near the impulse generator. CONCLUSIONS: Tailored laminectomy is a fast, minimally invasive and reliable technique for neuromodulator implantation. PMID- 10524879 TI - Is intraoperative electrostimulation of erectile nerves possible? AB - PURPOSE: We improved intraoperative conditions to achieve better corpora cavernosal response to stimulation of the erectile nerves. MATERIALS AND METHODS: A total of 18 men undergoing nerve sparing retropubic prostatectomy were evaluated with intraoperative stimulation for identification of the erectile nerves. Intracavernosal pressure was measured directly or via electromyography of the corpora cavernosa. Different kinds of anesthesia were used with or without urapidil. RESULTS: Intracavernosal pressure was recorded in all patients. Use of isoflurane based anesthesia blocked change, and total intravenous anesthesia with propofol resulted in a measurable change in intracavernosal pressure during electrostimulation. However, local urapidil, a potent alpha-blocking agent, doubled or tripled intracavernosal pressure. Electromyography of the corpora cavernosa demonstrated no measurable change. CONCLUSIONS: Intraoperative electrostimulation of erectile nerves requires special anesthesia as well as local blocking of alpha-receptors. The functional anatomy of the erectile nerves is variable. PMID- 10524880 TI - Sildenafil citrate after radical retropubic prostatectomy. AB - PURPOSE: Erectile dysfunction continues to be a significant problem for men after radical retropubic prostatectomy despite nerve sparing techniques. Sildenafil citrate (Viagra) has proved effective for erectile dysfunction in many men. We determine the efficacy of sildenafil in men with erectile dysfunction after radical retropubic prostatectomy and examine variables that may impact the response to treatment. MATERIALS AND METHODS: A total of 84 men were prescribed sildenafil after radical retropubic prostatectomy and asked to complete a series of questionnaires, including the International Index of Erectile Function (IIEF), on erectile function before and after sildenafil administration. The importance of factors, such as patient age, time since surgery, degree of cavernous nerve sparing, preoperative prostate specific antigen, Gleason score, clinical and pathological stage, and baseline postoperative erectile function, was examined. RESULTS: Of the 84 patients 45 (53%) had improved erections and 34 (40%) had improved ability for intercourse while taking sildenafil. Mean IIEF score for the erectile function domain increased from 9 to 14 (p <0.001). Orgasmic function (p = 0.004) and intercourse satisfaction (p = 0.009) also significantly improved. The degree of nerve sparing and baseline postoperative erectile dysfunction had a significant impact on the ability of sildenafil to improve erectile function (p = 0.010 and p <0.001, respectively) and total IIEF questionnaire responses (p = 0.031 and p <0.001, respectively). Age and pathological stage also appeared to have a significant effect. CONCLUSIONS: Sildenafil improved erectile function and the ability to have intercourse in more than half of men after radical retropubic prostatectomy. Baseline postoperative erectile function, which is dependent on the degree of nerve sparing technique, significantly impacts the likelihood that patients will respond to sildenafil. PMID- 10524881 TI - Is routine scrotal ultrasound advantageous in infertile men? AB - PURPOSE: We determine the value of routine scrotal ultrasonography in the evaluation of male infertility. MATERIALS AND METHODS: Scrotal color Doppler ultrasonography reports of 1,372 infertile men were reviewed to assess the prevalence of scrotal abnormalities and compared to clinical findings. RESULTS: The prevalence of scrotal abnormalities was 38%. Testicular tumor was found in 0.5%, varicocele in 29.7%, testicular cyst in 0.7%, testicular microlithiasis in 0.9%, epididymal cyst in 7.6% and hydrocele in 3.2% of the cases. Overall, 67% of sonography findings were not evident on palpation, and only 1 of 7 testicular tumors was suspected. Of the varicoceles 60% were not found on physical examination. The rate of testicular tumors (1/200) was higher than that reported for the general European population (1/20,000). CONCLUSIONS: Routine scrotal ultrasound provides valuable information in the diagnostic evaluation of infertile men and substantially more pathological conditions are detected compared to clinical palpation. The high prevalence of testicular malignancies underlines the clinical relevance of routine scrotal ultrasonography in infertile men. PMID- 10524882 TI - A comparative study of the no scalpel and standard incision approaches to vasectomy in 5 countries. The Male Sterilization Investigator Team. AB - PURPOSE: We compare the safety, ease of use and effectiveness of the no scalpel and standard incision approaches to vasectomy. MATERIALS AND METHODS: A multicenter, randomized, partially masked controlled trial was conducted at 8 sites in Brazil, Guatemala, Indonesia, Sri Lanka and Thailand. Semen samples were collected 10 weeks postoperatively and tested to ascertain sterility using verification of no living spermatozoa. RESULTS: The study included 1,429 men seeking vasectomy. The efficacy of the 2 approaches was virtually identical. In the no scalpel group operating time was significantly shorter, and complications and pain were less frequent than in the standard incision group. The no scalpel group resumed intercourse sooner, probably as a result of less pain following the procedure. CONCLUSIONS: The no scalpel approach is an important advance in the surgical approach to vasectomy, and offers fewer side effects and greater comfort compared to the standard incision technique, without compromising efficacy. PMID- 10524883 TI - Fertility outcome after repeat vasoepididymostomy. AB - PURPOSE: Historically, epididymal obstruction has been treated with surgical reconstruction. We determine whether it is worthwhile for patients to undergo repeat surgical reconstruction after failed vasoepididymostomy or whether they should be advised only to undergo sperm acquisition for assisted reproductive technique. MATERIALS AND METHODS: A total of 18 patients underwent repeat vasoepididymostomy performed by a single urologist (A. J. T.). Cases were divided based on the etiology of obstruction into groups 1--prior vasectomy (4), 2- congenital (7) and 3--inflammatory (7). Data were available regarding time of obstruction between initial and repeat vasoepididymostomy, quality of epididymal fluid, levels of anastomoses, semen analyses at least 12 months after surgery for all 18 men and pregnancy rates based on more than 18 months of followup in 12. RESULTS: Mean patient age at repeat vasoepididymostomy was 40.6 years (50.5, 36 and 39.4 years for groups 1, 2 and 3, respectively). Mean interval between vasectomy and initial vasoepididymostomy was 12.3 years (range 10 to 18). Mean interval between initial and repeat vasoepididymostomy was 19 months (range 12 to 41). Of the patients 10 underwent unilateral and 8 bilateral anastomoses, for a total of 26 repeat anastomoses. Overall patency rate was 66.7% (12 of 18) with sperm in the ejaculate in 75, 85 and 43% of patients in groups 1, 2 and 3, respectively. The patency rates according to the levels of the anastomosis were 66.7, 62.5 and 100% in the caput, corpus and cauda, respectively. Natural conception occurred in 3 of 12 couples (25%, 2 caput and 1 caudal anastomosis) during a mean followup of 23 months (range 13 to 34). All 3 cases had congenital obstruction. Pregnancy was achieved in 2 group 1 cases with cryopreserved sperm extracted at repeat vasoepididymostomy, and in 1 case each in groups 1 and 2 with microsurgical epididymal sperm aspiration and intracytoplasmic sperm injection. CONCLUSIONS: After repeat vasoepididymostomy two-thirds of men have sperm in the semen. Natural conception occurred in 25% of patients (3 of 12) followed for more than 18 months. Inability to establish pregnancy in the remaining 7 of 9 patients with sperm in the semen with a followup longer than 18 months may be due to epididymal dysfunction or partial obstruction and subsequent poor sperm quality. Aspiration of motile sperm and cryopreservation were possible in 11 of 18 cases at repeat vasoepididymostomy and should be recommended in case azoospermia remains or occurs after surgery. It appears worthwhile to offer patients repeat vasoepididymostomy after a failed initial procedure. PMID- 10524884 TI - A randomized double-blind study assessing 4 versus 8 mg. doxazosin for benign prostatic hyperplasia. AB - PURPOSE: We compare the efficacy of 4 versus 8 mg. doxazosin for benign prostatic hyperplasia. MATERIALS AND METHODS: A total of 82 patients with benign prostatic hyperplasia successfully treated with 4 mg. doxazosin were randomized in a double blind fashion to take 4 or 8 mg. Patients were evaluated with American Urological Association (AUA) symptom score, Boyarsky score, uroflowmetry and side effect profile before, and 1 and 3 months following randomization. RESULTS: Of the patients 42 and 40 were randomized to receive 4 and 8 mg. doxazosin, respectively. Both groups were similar with respect to patient age, baseline Boyarsky and AUA symptom scores, and baseline maximum urinary flow rate. At 3 months mean improvement from baseline plus or minus standard deviation in Boyarsky score was 0.6 +/- 6.5 and 4.9 +/- 6.6 in the 4 and 8 mg. groups (p <0.05), respectively, mean improvement in AUA symptom score was 1.6 +/- 5.3 and 5.3 +/- 8.0 (p <0.05), and mean maximum flow rate difference was -0.6 +/- 6.4 and +1.4 +/- 7.9 (p >0.05). Of the patients 7 and 8 in the 4 and 8 mg. groups dropped out of the study, and there were no statistical differences in side effects between dosages. CONCLUSIONS: A dose of 8 mg. doxazosin was more efficacious than 4 mg. and the side effects associated with both dosages appeared to be similar. The 8 mg. dose should be tried in patients who have not achieved an adequate therapeutic response to 4 mg. and are tolerating the medication. Consideration should be given to increasing the dosage to 8 mg. in patients who are clinically improved at lower dosages. PMID- 10524885 TI - A computer generated interactive transurethral prostatic resection simulator. AB - PURPOSE: We developed a computer generated model of the prostate gland and an interactive simulator for use in training urologists in transurethral resection of the prostate. MATERIALS AND METHODS: Software was developed on a standard personal computer which allowed images of the lumen of the prostatic urethra and resectoscope loop to be generated and interacted with using a magnetic position sensor input device attached to a dummy resectoscope. RESULTS: An anatomically accurate computer model of the prostate was generated at low cost which permitted user interaction and which simulated key elements of transurethral prostatic resection. CONCLUSIONS: Although not a substitute for learning transurethral prostatic resection on patients, the simulator enabled the user to become familiar with the technique of transurethral prostatic resection in the absence of time constraints and without risk to patients. The simulator may become an important tool in training and assessing surgeon competency, and may reduce the costs of training. Further development is needed to refine the transurethral prostatic resection simulator and expand its surgical range. PMID- 10524886 TI - A prospective study of transperineal prostatic block for transurethral needle ablation for benign prostatic hyperplasia: the Emory University Experience. AB - PURPOSE: We evaluate the feasibility, effectiveness and role of transperineal prostate block in providing anesthesia during minimally invasive radio frequency thermal therapy of the prostate. MATERIALS AND METHODS: A total of 38 consecutive patients undergoing transurethral needle ablation for symptomatic benign prostatic hyperplasia were entered in this prospective study. All patients received transperineal prostatic block as the main method of anesthesia. A mixture of equal volumes of 1% lidocaine and 0.25% bupivacaine, each with epinephrine (1:100,000 concentration ratio) was used. Pain control during the instillation of transperineal prostatic block and transurethral needle ablation was assessed using a 10-point linear analog pain scale and questionnaire. RESULTS: Median patient age was 65.5 years (range 47 to 79), with 21% of men in the eighth decade of life. Median American Urological Association symptom score was 25.0 (range 14 to 35), bother score was 20.0 (11 to 28), quality of life score was 4.0 (3 to 6) and peak urinary flow rate was 8.9 cc per second (3.5 to 15.7). Median sonographic prostate volume was 35.0 cc (range 17 to 129). Median volume of anesthetic agent used was 40.0 cc (range 30 to 60) per case (1.1 cc solution per 1 cc prostate tissue). No adverse events were encountered. Median pain score was 3.3 (range 1 to 6) during instillation of transperineal prostatic block and 1.0 (0 to 6) during transurethral needle ablation. Transperineal prostatic block proved highly effective and was a satisfactory method of anesthesia during transurethral needle ablation as judged by postoperative questionnaire. No sedation, narcotic or analgesia was required. All procedures were performed in the outpatient cystoscopy suite or office setting without support of an anesthesia team or conscious sedation monitoring. CONCLUSIONS: Transperineal prostatic block is a safe, convenient, effective and satisfactory method of minimally invasive anesthesia for transurethral needle ablation of the prostate in an outpatient office setting. Elderly patients and those at high surgical risk can be treated safely using this approach. Considerable cost saving is seen secondary to omission of charges related to anesthesia team support, recovery room facility and conscious sedation monitoring. PMID- 10524887 TI - Holmium laser versus transurethral resection of the prostate: a randomized prospective trial with 1-year followup. AB - PURPOSE: The high-powered holmium:YAG laser can be used for incision, ablation and resection of the prostate. The technique of holmium laser resection of the prostate is compared to transurethral prostatic resection for surgical management of benign prostatic hyperplasia in this prospective randomized study. MATERIALS AND METHODS: A total of 120 urodynamically obstructed cases were randomized to holmium laser or transurethral prostatic resection. All eligible patients were assessed preoperatively and at 3 weeks, and 3, 6 and 12 months postoperatively with an American Urological Association symptom score, peak urinary flow rate, and questionnaires concerning sexual function and continence. Preoperative pressure flow study, ultrasound prostate volume assessment and post-void residual volume measurement were repeated at the 6-month visit. All complications were noted. RESULTS: Holmium laser and transurethral resections resulted in significant improvements in symptom score, quality of life score, peak urinary flow rate and post-void residual urine measurements. Operating time was significantly longer in the holmium group but nursing contact time, catheter time and hospital stay were significantly less compared to the transurethral prostatic resection group. Urodynamic results were equivalent at 6 months. There were fewer side effects in the holmium group. Effects on continence, potency and symptoms were similar with 1-year followup. CONCLUSIONS: Holmium and transurethral resections of the prostate appear to be equivalent in surgical management of bladder outflow obstruction due to benign prostate hyperplasia. Perioperative morbidity was less in the holmium group. PMID- 10524888 TI - A randomized comparative study of the Bandloop versus the standard loop for transurethral resection of the prostate. AB - PURPOSE: The Bandloop is a new electroresection loop that is broader than the standard loop and thickens from front to back. We compared the safety and efficacy of the Bandloop to the conventional loop electrode for transurethral prostatectomy. MATERIALS AND METHODS: A randomized prospective study was performed on 53 patients with lower urinary tract symptoms and estimated prostatic volume greater than 30 ml. on transrectal ultrasonography. The Bandloop was used in 25 cases (Bandloop group) and standard loop was used in 28 (standard group). Surgical outcomes, including International Prostate Symptom Score, resected prostatic weight, operative time, uroflowmetry, post-void residual urine, postoperative catheterization period, time to disappearance of macrohematuria, and serial changes of hemoglobin and urinalysis, between the 2 groups were compared. RESULTS: A total of 23 Bandloop and 28 standard loop cases were evaluable. There was no difference in preoperative estimated prostatic volume (mean 44.7 versus 47.8 ml.), resected prostatic weight (36.5 versus 29.4 gm.) or operative time (61 versus 60 minutes) between the 2 groups. The ratio of resected weight-to-estimated prostatic volume was significantly greater in the standard group (81.4% versus 61.9%). There was no difference in postoperative catheterization period (3.3 versus 3.4 days) or time to disappearance of macroscopic hematuria (5.0 versus 5.0 days). Postoperative changes in symptom score (-15.1 versus -15.9), maximum flow rate (11.8 versus 16.3 ml. per second at 12 weeks) and residual volume (-31.6 versus -48.6 ml. at 12 weeks) demonstrated no significant difference between the 2 groups. There was no difference in intraoperative and postoperative bleeding estimated by serial changes in serum hemoglobin and urinalysis after surgery. No patient had major complications or required transfusion. CONCLUSIONS: Transurethral prostatectomy using the Bandloop is as safe and effective in achieving subjective and objective improvements as standard transurethral prostatectomy. However, our randomized study indicates that the Bandloop offers no advantage compared to the standard loop electrode for transurethral prostatectomy. PMID- 10524889 TI - Invasive therapy for benign prostatic hyperplasia--what's new. PMID- 10524890 TI - The significance of prior benign needle biopsies in men subsequently diagnosed with prostate cancer. AB - PURPOSE: We determine the relationship between a history of benign needle biopsies, and the volume and location of cancer. MATERIALS AND METHODS: We evaluated 395 men who underwent radical prostatectomy for stage T1c (nonpalpable) prostate cancer. RESULTS: Of the men 74 had 1 or more prior benign needle biopsies. Prior benign biopsy correlated with tumor in the anterior or lateral portion of the radical prostatectomy specimen (p = 0.044) and prostate weight (p = 0.002). The likelihood of prior benign biopsy was 32.5% for men with a 75 gm. or greater prostate compared to 15.2% for those with a less than 75 gm. prostate. Although prior benign biopsy correlated with "very limited" tumor in the prostate (less than 0.2 cc, no Gleason pattern 4 or 5 and organ confined disease) (p = 0.005), only 28.4% of patients with prior benign biopsy had "very limited" tumor. In a multivariate analysis prior benign biopsy correlated only with anterior or lateral distribution and enlarged prostate size. Of the prior benign biopsy cases 12% had positive margins, average tumor volume was 1.15 cc and 27% had nonorgan confined disease. These figures were not different from those in cases with cancer on the first biopsy. In prior benign biopsy cases although PSA velocity predicted tumor volume and "very limited" tumor, a specific clinically useful cutoff value was not present. Needle biopsy grade and number of positive cores were not predictive of tumor volume or "very limited" cancer. CONCLUSIONS: Prior benign biopsy in men subsequently diagnosed with prostate cancer does not indicate indolent tumor. Benign biopsies are more likely in larger prostate glands and when cancer is in the anterior and lateral regions of the gland, suggesting the need for different biopsy strategies to improve cancer detection. PMID- 10524891 TI - Total cryosurgery of the prostate versus standard cryosurgery versus radical prostatectomy: comparison of early results and the role of transurethral resection in cryosurgery. AB - PURPOSE: Results of standard cryosurgery of the prostate for prostate cancer in 49 patients were compared to those of destruction of the urethra during or after cryosurgery with subsequent transurethral resection or total freezing of the prostate (total cryosurgery) in 27. These results were compared to those of radical surgery in 83 patients with similar age, stage and grade of disease, and prostate specific antigen (PSA). MATERIALS AND METHODS: The 76 cryosurgery cases included all of those treated by 1 surgeon (R. S. G.) for localized prostate cancer after July 1, 1995. The 83 radical perineal prostatectomy cases consisted of all of those treated by another surgeon during the study period and by R. S. G. before cryosurgery use. Success was defined as a PSA of 0.2 or less 6 months after the procedure and a stricter standard, 0.0 PSA, was also assessed. RESULTS: The success rate was 96% for total cryosurgery, 48.9% for standard cryosurgery and 73.4% for radical surgery. Using 0.0 PSA as a criterion, 66.7% of total cryosurgery, 16.3% of standard cryosurgery and 48.2% of radical surgery cases were successfully treated. CONCLUSIONS: Total cryosurgical destruction of the prostate may offer new opportunities for cancer treatment heretofore unrecognized and should undergo more investigational analysis. PMID- 10524892 TI - Orchiectomy and orchiectomy plus mitomycin C for metastatic prostate cancer in patients with poor prognosis: the final results of a European Organization for Research in Cancer Therapy Genitourinary Group Trial. AB - PURPOSE: The outcome of patients with symptomatic metastatic prostate cancer is poor and improved treatment regimens are urgently needed. Theoretically, the combination of orchiectomy and chemotherapy could reduce androgen sensitive and insensitive cells in the prostate. This European Organization for Research in Cancer Therapy Genitourinary Group randomized, multicenter phase III trial demonstrates the outcome of orchiectomy alone versus orchiectomy followed by intravenous mitomycin C. MATERIALS AND METHODS: A total of 189 patients with metastatic prostate cancer and poor prognostic factors were randomized in this trial by 42 institutions. Of these patients 184 (97%) were eligible for study, including 90 treated with orchiectomy alone (orchiectomy only arm) and 94 treated with orchiectomy followed by 15 mg./m.2 mitomycin C in 1 week (combined treatment arm). Mitomycin C was administered every 6 weeks and treatment was continued as long as tolerance and patient compliance allowed, and no progression was observed. Objective and subjective criteria for progression were clearly defined in the protocol. RESULTS: Patient and tumor characteristics were well balanced between the 2 treatment arms. At a median followup of 4.2 years 144 patients had died, including 112 of prostate cancer. No significant differences for time to overall (p = 0.17), subjective (p = 0.25) and objective (p = 0.08) progression were found between the 2 treatment groups. For progression-free survival no difference was noted (p = 0.67) between the 2 treatment groups but a trend in favor of orchiectomy alone was observed for overall survival (p = 0.04). Mitomycin C induced considerable hematological, gastrointestinal, renal and pulmonary toxicity leading to discontinuation in 31% of patients with pulmonary toxicity and 7% with renal deterioration. In addition, the quality of life evaluation revealed significant reduction in the combined treatment arm. CONCLUSIONS: Based on the results of this randomized phase III study orchiectomy plus mitomycin C for metastatic prostate cancer in patients with poor prognostic factors cannot be recommended due to failure of improvement in survival and reduced quality of life parameters. PMID- 10524893 TI - Proteus mirabilis viability after lithotripsy of struvite calculi. AB - PURPOSE: We tested the hypotheses that Proteus mirabilis viability of struvite calculi differs after exposure to different lithotripsy modalities and that the photothermal mechanism of holmium:YAG lithotripsy is antibacterial. MATERIALS AND METHODS: Human calculi of known struvite composition (greater than 90% magnesium ammonium phosphate hexohydrate) were incubated with P. mirabilis. Calculi were randomly distributed and fragmented with no lithotripsy (controls), or shock wave, intracorporeal ultrasonic, electrohydraulic, pneumatic, holmium:YAG or pulsed dye laser lithotripsy. After lithotripsy fragments were sonicated and specimens were serially plated for 48 hours at 38C. Bacterial counts and the rate of bacterial sterilization were compared. RESULTS: Median bacterial counts (colony-forming units per ml.) were 8 x 10(6) in controls and 3 x 10(6) in shock wave, 3 x 10(7) in ultrasonic, 4 x 10(5) in electrohydraulic, 8 x 10(6) in pneumatic, 5 x 10(4) in holmium:YAG and 1 x 10(6) in pulsed dye laser lithotripsy cases (p <0.001). The rate of bacterial sterilization was 50% for holmium:YAG lithotripsy treated stones versus 0% for each of the other cohorts (p <0.01). CONCLUSIONS: P. mirabilis viability varies among lithotrites. The photothermal mechanism of holmium:YAG lithotripsy is antibacterial. PMID- 10524894 TI - Results of pubovaginal sling for stress incontinence: a prospective comparison of 4 instruments for outcome analysis. AB - PURPOSE: Presently to our knowledge there are no standardized techniques to assess outcomes after surgery for stress incontinence. We performed a prospective blinded study to assess the correlation among physician and patient assessments, and a validated 24-hour pad test and voiding diary. MATERIALS AND METHODS: A total of 84 women were evaluated before and after pubovaginal sling for stress incontinence with a voiding diary, pad test and symptom questionnaire (patient assessment) administered by a blinded third party. The operating surgeon evaluated the patient using history, physical examination, pad test and voiding diary but was blinded to results of the outcome questionnaire. Preoperative focused neurourological examination and video urodynamics confirmed stress incontinence. Patients were assessed at least 1 year postoperatively. We compared patient assessment (cured, improved, failure) to the outcome of the pad test, voiding diary and physician assessment. The physician and questioner were blinded to each other. We considered patients with a pad test of 0 to 2 ml. as cured, 50% or more volume reduction as improved and less than 50% volume reduction as failure. Postoperative assessment did not differentiate between stress and urge incontinence. The kappa coefficient was used for statistical comparison. RESULTS: Average patient age was 58 years and average followup for the entire group was 4 years. Agreement among the 4 instruments to assess outcome was excellent (k >0.9) with respect to cured/improved versus failure but only good for cured versus improved versus failure (k >0.5). CONCLUSIONS: Outcomes following incontinence surgery may vary depending on how the analysis was performed, patient selection, definition of success and so forth. Our results indicate that a pad test and voiding diary are reliable and should be part of the normal followup after pubovaginal sling for sphincteric incontinence. When these tests are used in conjunction with defined parameters of success, there is excellent agreement with patient feelings in regard to success or failure of surgery. Nevertheless, these instruments and methods are imperfect at best. PMID- 10524895 TI - A simple objective method of adjusting sling tension. AB - PURPOSE: Pubovaginal sling is gaining widespread acceptance as a primary form of treatment for types II and III stress urinary incontinence. However, a major drawback is postoperative obstructed voiding due to excessive force placed on the suspension suture. We describe a simple objective method for intraoperative adjustment of sling tension that can be performed by a single surgeon during pubovaginal sling surgery. MATERIALS AND METHODS: A cotton swab is inserted into the urethra and placed at the urethrovesical junction after the sling is fixed suburethrally and the vaginal mucosa is closed. The suspension sutures are tied down directly onto the rectus fascia with enough tension to keep the cotton swab angle between 0 and 10 degrees to the horizontal plane. A total of 29 patients with an average age of 62 years underwent pubovaginal sling surgery with rectus and cadaveric fascia using this technique for tension adjustment. Of the patients 21 were diagnosed with types II and III, 5 had type II only and 3 had type III only incontinence. Preoperative evaluation revealed detrusor instability in 5 patients. Mean postoperative indwelling catheterization period was 6.2 days. Average followup was 15.6 months. RESULTS: To date no permanent urinary retention has occurred. Of the patients 15 voided without difficulty after catheter removal, 13 had urinary difficulty requiring intermittent catheterization for 1 week or less and 1 had retention requiring intermittent catheterization for 10 weeks. Preoperative symptoms of detrusor instability resolved in all cases. De novo detrusor instability in 3 cases was controlled with anticholinergics. CONCLUSIONS: Overzealous sling tension adjustment has been recognized as a cause of treatment failure leading to urethral obstruction. Our technique is effective in preventing over adjustment of tension, is reproducible and can be performed by 1 surgeon. PMID- 10524896 TI - Retractile mesenteritis mimicking an adrenal tumor. PMID- 10524897 TI - Successful renal transplantation in 3 family members with type 1 renal tubular acidosis. PMID- 10524898 TI - Idiopathic retroperitoneal fibrosis presenting as duodenal obstruction. PMID- 10524899 TI - Retroperitoneal plasmacytoma associated with hyperamylasemia. PMID- 10524900 TI - Pouch conservation during Studer enterocystoplasty in ileal nondetubularized segmental stenosis. PMID- 10524901 TI - Carcinoid tumor in an ileal conduit diversion. PMID- 10524902 TI - Transitional cell carcinoma in a urachal cyst. PMID- 10524903 TI - Successful endoscopic closure of radiation induced vesicovaginal fistula with fibrin glue and bovine collagen. PMID- 10524904 TI - Spontaneous perforation of the gallbladder as a complication of radical cystoprostatectomy. PMID- 10524905 TI - Endoscopic retrieval of a proximal corpus cavernosum polytetrafluoroethylene sleeve during explantation of an infected penile prosthesis. PMID- 10524906 TI - Scrotal involvement with idiopathic retroperitoneal fibrosis. PMID- 10524907 TI - Verrucous scrotal carcinoma: a radioresponsive tumor. PMID- 10524908 TI - An unusual complication of transurethral resection: reflux into the vas deferens, seminal vesicles and epididymis. PMID- 10524909 TI - Bladder cancer clinical guidelines panel summary report on the management of nonmuscle invasive bladder cancer (stages Ta, T1 and TIS). The American Urological Association. AB - PURPOSE: The American Urological Association convened the Bladder Cancer Clinical Guidelines Panel to analyze the literature regarding available methods of treating nonmuscle invasive bladder cancer, and to make practice policy recommendations based primarily on treatment outcomes data. MATERIALS AND METHODS: The panel searched the MEDLINE database for all articles related to nonmuscle invasive bladder cancer published from 1966 to January 1998. Outcomes data were extracted from articles accepted after panel review and meta-analyzed to produce comparative probability estimates for alternative treatments. RESULTS: All of the intravesical agents (thiotepa, bacillus Calmette-Guerin, mitomycin C and doxorubicin) when used as adjuvant therapy after transurethral resection resulted in a lower probability of recurrence compared to resection alone. However, there is no evidence that intravesical therapy affects long-term progression. CONCLUSIONS: For patients with no prior intravesical therapy adjuvant intravesical chemotherapy or immunotherapy is a treatment option after endoscopic removal of low grade Ta bladder cancers. Intravesical instillation of bacillus Calmette-Guerin or mitomycin C is recommended for carcinoma in situ, and after endoscopic removal of T1 and high grade Ta tumors. PMID- 10524910 TI - 1998 American Urological Association Gallup Survey: changes in physician practice patterns, treatment of ureteral stones and impact of managed care. AB - PURPOSE: The American Urological Association first commissioned the Gallup Organization to conduct a study to assess urologist practice patterns in 1992. We present the results of the seventh consecutive Gallup Survey performed for the Association. MATERIALS AND METHODS: A random sample of 537 American urologists who completed urological residency and practiced at least 20 hours weekly in 1997 were interviewed by telephone in March 1998. Major topic areas included general demographics, practice patterns, treatment of ureteral stones and experience with managed care. RESULTS: Demographic trends indicated a significant decrease in average urologist age from 49.4 years in 1992 to 46.8 in 1998. Of the urologists 99% reported that they treat ureteral stones. Managed care had an increasingly larger role in most practices, particularly in the western United States, where 73% of urologists reported that they contract with a Medicare health maintenance organization. CONCLUSIONS: The average age of practicing urologists significantly decreased, which may be due to an increasing number of urologists retiring at an earlier age, although this finding is not clear. Nearly all urologists treated ureteral stones with considerable consistency. Finally, managed care appeared to have a major impact on most urologists throughout the United States. PMID- 10524911 TI - Re: Floating kidneys: a century of nephroptosis and nephropexy. PMID- 10524912 TI - Re: Vesicoureteral reflux in infants with prenatal hydronephrosis confirmed at birth: racial differences. PMID- 10524913 TI - Re: Intravesical interleukin-2 in T1 papillary bladder carcinoma: regression of marker lesion in 8 of 10 patients. PMID- 10524914 TI - Re: Strategies for reconstruction after unsuccessful or unsatisfactory primary treatment of patients with bladder exstrophy or incontinent epispadias. PMID- 10524915 TI - Re: Congenital dorsal penile curvature: a potential problem of the long phallus. PMID- 10524916 TI - Re: A comparison of noncontrast computerized tomography with excretory urography in the assessment of acute flank pain. PMID- 10524917 TI - Re: Frere Jacques Beaulieu: from rogue lithotomist to nursery rhyme character. PMID- 10524918 TI - Re: AUA code of ethics. American Urological Association. PMID- 10524919 TI - Percutaneous management of renal calculi: experience with percutaneous nephrolithotomy in 60 children. AB - PURPOSE: We report our experience with percutaneous nephrolithotomy in a pediatric population in which primary as well as recurrent stone episodes are frequent and the need for less invasive procedures is imperative. MATERIALS AND METHODS: Percutaneous nephrolithotomy was performed in 60 children 3 to 13 years old (average age 6), including 44 boys (73.3%) and 16 girls (26.7%). There was a single obstructing renal calculus in 43 patients, while 17 had multiple calculi. The procedure was performed in 1 stage in 49 patients, and it was staged with preliminary nephrostomy in 11 who presented with calculous anuria and elevated serum creatinine. Normal saline was used as an irrigant and perioperatively serum electrolytes were measured to monitor fluid absorption in 18 patients. Stones were extracted intact from 40 patients (66.6%) and ultrasonic lithotripsy was performed in 20 (33.3%). RESULTS: Of the 60 patients 50 (83.3%) were rendered stone-free at 1 session. Incomplete stone clearance at 1 session was due to intraoperative bleeding requiring blood transfusion, extravasation, multiple stones that were inaccessible via 1 tract, displacement of stone fragments into an inaccessible calix and insignificant residual fragments less than 3 mm. in 2 cases each. During followup of 3 months to 6 years (average 1 year) no late complications were noted. CONCLUSIONS: Percutaneous nephrolithotomy is a safe and relatively efficacious mode of managing pediatric renal calculi. Although higher success rates are achieved in adults, caution should be exercised in children, in whom diligent attempts at stone clearance in 1 session may be made at the expense of safety. PMID- 10524920 TI - Initial experience with endoscopic holmium laser lithotripsy for pediatric urolithiasis. AB - PURPOSE: Due to the unavailability of suitable pediatric instruments children have not benefited from advances in endoscopic lithotripsy. This limitation may be overcome by the holmium: YAG laser. We evaluated the indications for, and efficacy and complications of holmium:YAG laser lithotripsy. MATERIALS AND METHODS: We retrospectively reviewed all cases of laser lithotripsy. Access to the calculus was antegrade or retrograde. A solid state holmium:YAG laser was used. RESULTS: Eight patients 4 to 14 years old underwent laser lithotripsy during the study period. Average calculous surface area was 357.13 mm.2 (range 14 to 1,645). Five patients required 1 procedure to render them stone-free, while the remaining 3 required multiple procedures. No complications were associated with laser lithotripsy. CONCLUSIONS: The ability of the holmium:YAG laser to pulverize urinary calculi makes it an alternative choice for lithotripsy. In our series all patients are stone-free with stable renal function. The advantages of the holmium:YAG laser are that it may be precisely applied via small fibers, and it pulverizes calculi with minimal scattering of energy and retropulsion of the calculus, decreasing trauma to tissues at the perioperative site. There is also a lower risk of residual fragments, which is associated with a lower incidence of calculous regrowth. Holmium: YAG laser is safe and effective for treating pediatric urolithiasis. PMID- 10524921 TI - Holmium:YAG lithotripsy in children. AB - PURPOSE: We determined the safety and efficacy of holmium:YAG lithotripsy in children. MATERIALS AND METHODS: We retrospectively reviewed the records of all holmium:YAG lithotripsy done in patients 17 years old or younger. Demographic, preoperative, intraoperative and postoperative data were collected. RESULTS: A total of 9 boys and 10 girls (26 stones) with a mean age of 11 years (range 1 to 17) were treated with holmium:YAG lithotripsy, which was chosen as initial therapy in 10 (53%). Retrograde ureteroscopy was performed in 15 patients to treat 13 ureteral and 6 renal calculi, and percutaneous nephrolithotripsy was done in 4 to treat 3 ureteral and 4 renal calculi. A complete stone-free outcome after 1 procedure was achieved in 16 children (84%) and 3 patients were rendered stone-free after 2 procedures. No patient had an intraoperative injury. Followup ranged from 0.5 to 12 months (mean 3). Followup imaging has shown no evidence of stricture or hydronephrosis. CONCLUSIONS: Holmium:YAG lithotripsy is safe and effective in children. It is a reasonable option for failed shock wave lithotripsy, or in children with a known durile stone composition or contraindications to shock wave lithotripsy. PMID- 10524922 TI - Percutaneous nephrolithotomy in the pediatric population. AB - PURPOSE: Percutaneous nephrolithotomy is an established technique used in children with renal calculi. We review our experience with percutaneous nephrolithotomy for treating nephrolithiasis in childhood. MATERIALS AND METHODS: We retrospectively reviewed the records of children who underwent percutaneous nephrolithotomy procedures for renal calculi from 1985 to 1996. Antegrade percutaneous access was obtained in all patients and the tract was dilated to 24F. Grasper forceps, ultrasound and/or electrohydraulic lithotripsy was used to remove and disintegrate stones. In all patients a nephrostomy tube was placed intraoperatively, and a plain abdominal x-ray and nephrostogram were done postoperatively. The nephrostomy tube was removed after ensuring free drainage down the ureter and no untoward effects from clamping. Complete anatomical and metabolic evaluation was performed in all cases. Patients were followed 2 to 6 weeks, and 3 and 6 months postoperatively with a plain abdominal x-ray and excretory urography or renal ultrasound. RESULTS: In 5 boys and 3 girls (9 renal units) 4 to 11 years old (mean age 6.4) a total of 10 percutaneous nephrolithotomy procedures were performed. At presentation 6 children had flank and/or abdominal pain, 5 gross hematuria and 3 urinary tract infection. Three patients had associated metabolic abnormalities. One patient with a staghorn calculus had hydronephrosis and multiple infundibular stenoses. No underlying urological anatomical abnormalities were noted in the remaining cases. Four renal units that were obstructed at presentation required initial nephrostomy tube insertion. Average operative time was 131.8 minutes (range 58 to 240). An 87.5% stone-free rate was achieved using percutaneous nephrolithotomy monotherapy. Percutaneous nephrolithotomy was not successful for eradicating a staghorn stone in 1 patient. Hypothermia developed in 2 patients in whom operative time exceeded 150 minutes. No blood transfusions were required. CONCLUSIONS: Percutaneous nephrolithotomy is safe and effective in children, and should be considered a viable management option. However, staghorn calculi may require alternative management, particularly in the setting of underlying anatomical abnormalities. Children with renal calculi should undergo a complete anatomical and metabolic assessment with the institution of medical therapy, as appropriate. PMID- 10524923 TI - Pediatric retroperitoneoscopic nephrectomy using 2 mm. instrumentation. AB - PURPOSE: We describe several modifications of the retroperitoneoscopic approach to nephrectomy for benign renal disease, including the use of 2 mm. instrumentation and prone patient positioning. MATERIALS AND METHODS: A total of 14 children underwent retroperitoneoscopic nephrectomy in the prone position. An inflatable dissecting device was inserted into the retroperitoneum after a small muscle splitting incision was made at the lateral border of the sacrospinalis muscle approximately 1 cm. below the costovertebral angle. After inflation the dissecting device was replaced with a 5 mm. cannula and pneumoretroperitoneum was maintained with carbon dioxide insufflation. Two 2 mm. trocars were then placed under endoscopic guidance. Dissection was performed using 2 mm. instrumentation and the specimen was retrieved through the largest port site. RESULTS: Nephrectomy was performed in 9 girls and 5 boys 3 months to 9.8 years old. The preoperative diagnosis included chronic pyelonephritis with minimal renal function, reflux with a nonfunctioning kidney, multicystic dysplastic kidney, an upper pole dysplastic moiety with an associated ureterocele and a dysplastic kidney with a vaginal ectopic ureter. Mean operative time for retroperitoneoscopic nephrectomy was 142 minutes with an estimated blood loss of less than 15 ml. Contralateral ureteral reimplantation was performed after retroperitoneoscopic dissection in 5 patients. Overall average hospital stay was 2 days and there were no complications. CONCLUSIONS: Several modifications of the retroperitoneal approach, including the use of prone patient positioning and 2 mm. instrumentation for visualization and dissection, may improve the safety and efficacy of this technique in children. PMID- 10524924 TI - Pediatric endourology--coming into focus. PMID- 10524925 TI - Urodynamic pattern in asymptomatic infants: siblings of children with vesicoureteral reflux. AB - PURPOSE: We studied the urodynamic pattern in asymptomatic infants who are siblings of children with vesicoureteral reflux. MATERIALS AND METHODS: Cystometry and perineal electromyography were performed with voiding cystourethrography in 16 male and 21 female infant siblings screened for reflux at age 0.2 to 7.3 months (median 1.1). RESULTS: Vesicoureteral reflux was present in 25% of the male and 10% of the female infants. In those without vesicoureteral reflux unstable bladder contractions were noted in 8% of the male and 16% of the female subjects. In these infants median maximum voiding detrusor pressure was 127 (range 84 to 211) and 72 cm. water (range 42 to 240), respectively, and median bladder capacity was 20 ml. (range 10 to 49 and 10 to 120, respectively). Maximum voiding detrusor pressure was significantly higher in male than in female infants (p <0.01). Perineal electromyography was interpretable in 13 of the 16 male and 16 of the 21 female infants overall. All but 1 female subject had increased activity during voiding, which was also present intermittently in all subjects. CONCLUSIONS: Our study of asymptomatic siblings of children with vesicoureteral reflux has provided results that may be used as reference data for normal urodynamics in early infancy. Instability was rare. Bladder capacity was lower than expected with a predicted capacity at birth of approximately 20 ml. Maximum voiding pressure was high, especially in male subjects. The urodynamic voiding pattern suggests physiological dyscoordination, probably due to immature detrusor-sphincter function. PMID- 10524926 TI - Urinary cytokines as markers of reflux nephropathy. AB - PURPOSE: We established whether the urinary cytokines interleukin-6 (IL-6), tumor necrosis factor (TNF)-alpha and soluble TNF receptor-1 have a role as noninvasive markers of renal damage in children with vesicoureteral reflux. MATERIALS AND METHODS: We performed an observational study in a surgical and urological unit at a pediatric teaching hospital. Urine cytokine levels of IL-6, TNF-alpha and soluble TNF receptor-1 were measured using a standard enzyme-linked immunosorbent assay technique in children stratified into group 1--11 with vesicoureteral reflux and reflux nephropathy, group 2--6 with vesicoureteral reflux only and no associated nephropathy, and group 3--15 age and sex matched controls. RESULTS: Urinary levels of the cytokines IL-6 and soluble TNF receptor-1 were significantly elevated in group 1 versus group 3 (0.048 to 13.25 pg./micromol. creatinine, mean 3.658 versus 0.027 to 0.677, mean 0.247 and 102.89 to 4,502.9 pg./micromol. creatinine, mean 1,395.3 versus 13.06 to 569.6, mean 145.357, respectively). Neither cytokine in group 2 (0.074 to 10.96 pg./micromol. creatinine, mean 2.94 and 51.52 to 1,115.48, mean 413.137, respectively) was elevated compared to that in group 3. TNF-alpha was not elevated in group 1 or 2 compared to that in group 3 (0.104 to 2.518 pg./micromol. creatinine, mean 0.56, 0.094 to 1.278, mean 0.334 and 0.065 to 0.694, mean 0.241, respectively). CONCLUSIONS: Measuring the urinary levels of the cytokines IL-6 and soluble TNF receptor-1 may be useful as a noninvasive marker of reflux associated renal damage. Further studies with larger patient groups are necessary to locate the source of production of the elevated urinary cytokines measured in our study. PMID- 10524927 TI - A long continent ileovesicostomy using a single piece of bowel. AB - PURPOSE: In 1981 Mitrofanoff presented a procedure to create a continent urinary stoma for intermittent catheterization. Since then, several other methods have been introduced, including the Yang-Monti ileovesicostomy. The length of these ileovesicostomies is limited by the circumference of the bowel segment used, which is inadequate in some cases. We developed a procedure to double the length of the Yang-Monti ileovesicostomy using a single section of bowel. MATERIALS AND METHODS: A 3.5 cm. section of ileum is isolated on its mesentery. The bowel is divided into 2 segments for 80% of its circumference, leaving the bowel intact over the mesentery. Each ring of bowel is then divided adjacent to the mesentery but on opposite sides, allowing the bowel to be unfolded and reconfigured in a single long strip that may then be tubularized. The blood supply to the tube is excellent and it is in the center of the reconfigured ileum. The ends may be trimmed or widely spatulated as necessary. RESULTS: We have performed this procedure in 8 patients. The resulting ileovesicostomy created from a 3.5 cm. section ofileum is 10 to 14 cm. long and accepts a 12F catheter. A larger tube may be created from a longer piece of ileum. All patients are dry and they perform catheterization easily. CONCLUSIONS: This form of ileovesicostomy allows the creation of a long bowel tube that is easily catheterized. The longer length of the tube increases application of the continent stoma principle to more patients and enables reconstruction to be performed with optimal placement and without tension. PMID- 10524928 TI - Concomitant modified bladder neck closure and Mitrofanoff urinary diversion. AB - PURPOSE: We describe a modification of bladder neck closure for managing urinary incontinence in children. MATERIALS AND METHODS: In 11 patients with intractable urinary incontinence that persisted after multiple failed surgical procedures we performed modified bladder neck closure with construction of a catheterizable continent conduit. RESULTS: Mean followup is 3 years. All patients were continent after the procedure and none had a fistula or urinary leakage. One patient required stomal and conduit revision, and bladder stones in 3 necessitated endoscopic removal. CONCLUSIONS: We recommend this modified technique of bladder neck closure as an option for managing urinary incontinence in a complex group of children because it allows the achievement of continence with minimal morbidity. PMID- 10524929 TI - Appendicovesicostomy and newer alternatives for the Mitrofanoff procedure: results in the last 100 patients at Riley Children's Hospital. AB - PURPOSE: We present our experience using the various Mitrofanoff techniques to create a continent catheterizable stoma as an adjunct to continent urinary tract reconstruction in children and young adults. MATERIALS AND METHODS: Between 1990 and 1998 a Mitrofanoff procedure was performed at our institution in 55 male and 45 female patients with a mean age of 10.5 years. The etiology of incontinence was diverse but more than 90% of the patients had neurogenic bladder, the epispadias-exstrophy complex or a cloacal anomaly. Surgery included appendicovesicostomy in 57 cases, a Yang-Monti ileovesicostomy in 21, continent vesicostomy in 21 and formation of a tapered ileal segment as a catheterizable channel in 1. Simultaneously bladder augmentation was performed in 52 patients, bladder neck reconstruction was done in 48 and a Malone antegrade colonic enema stoma was constructed for fecal incontinence in 17. RESULTS: The abdominal stoma is continent in 98 of our 100 patients. Mean followup is 2 years (range 2 months to 8 years) with the longer followup in the appendicovesicostomy group. One patient with stomal incontinence who underwent revision is now dry. Postoperative complications requiring an additional procedure developed in 20 patients, including stomal stenosis in 12. Continent vesicostomy was most prone to stomal problems (6 of 21 patients, 29%). CONCLUSIONS: The Mitrofanoff procedure is a reliable technique for creating a continent catheterizable urinary stoma. Appendicovesicostomy continues to be our first option for this procedure, although we have also had good results with the Yang-Monti ileovesicostomy and continent vesicostomy. These newer options have allowed preservation of the appendix for the Malone antegrade colonic enema stoma procedure in patients with urinary and fecal incontinence. PMID- 10524930 TI - Enlarged penis due to a plexiform neurofibroma. PMID- 10524931 TI - Varicocele: a multidisciplinary approach in children and adolescents. AB - PURPOSE: To minimize varicocele treatment in children and adolescents a multidisciplinary approach that includes surgery and operative radiology has been used at our institution since 1991. We present our results during this 7-year period. MATERIALS AND METHODS: From January 1991 to December 1997 we examined 477 patients 4 years 5 months to 25 years 4 months old (mean age 13 years 3 months) with varicocele, of whom 367 (396 varicoceles) required treatment. Percutaneous sclero-embolization was suggested as the primary treatment of choice in all cases, while surgery was reserved for select cases. A total of 366 cases followed at least 6 months (mean 1 year) were entered into this study. RESULTS: Only 7.1% of the patients or families preferred surgery. In 47 patients sclero-embolization was not possible due to technical problems or vascular anomalies. Sclero embolization was successful in 79.4% of 277 patients, and retroperitoneal ligation was successful in 88.7% of 124. Since 1995 ligation of the whole spermatic bundle above the vas deferens has been preferred, and only 1 recurrence has been observed in 60 cases. CONCLUSIONS: Percutaneous sclero-embolization is a minimally invasive treatment of varicocele that is feasible in children and adolescents. Most patients prefer this therapy, although it is not as safe as surgery. When open surgery is required, complete ligation of the whole vascular pedicle above the vas deferens offers excellent success. PMID- 10524932 TI - Biocompatibility testing of a new bioabsorbable X-ray positive SR-PLA 96/4 urethral stent. AB - PURPOSE: Recently a first X-ray-positive bioabsorbable urethral stent was developed by our group. The stent is made from self-reinforced poly-L,D-lactic acid (SR-PLA 96/4) blended with barium sulfate. The aim of this study was to evaluate the biocompatibility properties of the new stent materials. MATERIALS AND METHODS: Rods made from pure SR-PLA 96/4 and SR-PLA 96/4 blended with barium sulfate were inserted into the dorsal muscles of a rabbit. Rods made from latex and silicone were used as positive and negative controls. To evaluate the long term effect of BASO4 after the bioabsorption of the polymer, fast degrading SR PGA (self-reinforced polyglycolic acid) and SR-PLA + BASO4 rods were used as controls. Urethral stents made from SR-PLA 96/4 and X-ray-positive SR-PLA 96/4 stents were inserted cystoscopically into the rabbit urethra. Metal stents were used as controls. The animals were sacrificed after 1 week, 1 month or 6 months. RESULTS: In the muscle implantation samples acute tissue reactions due to operative trauma were seen in all specimens at 1 week. After 6 months chronic inflammatory changes and foreign body reaction were seen only in the positive controls. The stent worked well in the rabbit urethra, its biocompatibility was good and there was less encrustation than in the metal stents. CONCLUSIONS: This first X-ray-positive bioabsorbable urethral stent showed no toxic tissue effects. PMID- 10524933 TI - Overactivity and structural changes in the chronically ischemic bladder. AB - PURPOSE: Our aim was to study the effect of chronic ischemia on bladder contraction and detrusor smooth muscle reactivity. The relationship between structural damage and functional changes in the chronically ischemic bladder was also investigated. MATERIAL AND METHODS: Male New Zealand White rabbits were divided into arterial injury (AI), hypercholesterolemia (Hch) and control groups. The AI group (n = 18) underwent balloon endothelial injury of the iliac arteries and received a 0.5% cholesterol diet. The Hch group (n = 8) received a 0.5% cholesterol diet alone. The control group (n = 8) received a regular diet. After 16 weeks, iliac artery and bladder wall blood flows were recorded. Cystometrograms and arteriography were obtained and bladder tissues were processed for isometric tension measurement in the organ bath and for histological evaluation. RESULTS: At 16 weeks, blood flow through the iliac arteries was significantly reduced in the AI group compared with the Hch and control groups. In the AI group, 8 animals developed severe bladder ischemia (SBI) defined as greater than 60% decrease in bladder blood flow, 7 animals developed moderate bladder ischemia (MBI) defined as 40 to 60% decrease in bladder blood flow, and 3 animals failed to develop significant bladder ischemia (<40% decrease in bladder blood flow). In the control animals, bladder blood flow increased prior to contraction, decreased during contraction and rebounded to baseline levels after contraction. In animals with MBI and SBI, the increase in bladder blood flow prior to contraction and the rebound of blood flow after contraction, both seen in control animals, were diminished. Detrusor overactivity (significant increase in the frequency of spontaneous bladder contractions) was observed in the MBI group and impaired bladder contraction in the SBI group. In the organ bath, bladder strips from the MBI group demonstrated increased contractile response to carbachol and electrical field stimulation (EFS) while bladder strips from the SBI group showed impaired contractility. Hch alone produced only short-lived ischemia during bladder contraction and caused significantly lesser functional changes compared with those seen in MBI. Histological examination showed atherosclerotic occlusion in the iliac arteries and bladder microcirculation and marked disruption of urothelium in the MBI and SBI groups. Severe fibrosis was seen in bladder tissue from the SBI group, moderate fibrosis in tissue from the MBI group and mild fibrosis in tissue from the Hch group. CONCLUSIONS: Our studies show that chronic MBI is associated with detrusor overactivity and increased smooth muscle contractility to carbachol and EFS while chronic SBI is associated with impaired detrusor contraction. The mechanism of chronic ischemia-induced bladder dysfunction is not known and may involve multiple physiologic and structural changes in the bladder nerves, receptors and contractile components. Our studies suggest that ischemia-induced structural damage in the urothelium and possible chronic exposure of the underlying tissue and nerves to the urine may also play a role in MBI-induced detrusor overactivity. SBI-induced impairment of bladder contraction may involve, in part, extensive fibrosis and loss of bladder smooth muscle. Histopathophysiologic changes in bladder tissue from our MBI model are similar to those seen in patients with detrusor instability, suggesting that chronic ischemia may play a role in the development of idiopathic detrusor instability. PMID- 10524934 TI - Characterization of cultured bladder smooth muscle cells: assessment of in vitro contractility. AB - PURPOSE: The contractile properties of in vitro cultured bladder smooth muscle cells (SMC) are unknown. This study characterized the in vitro contractile response of human and rat bladder SMC to several pharmacological agonists known to induce in vivo contraction of intact bladder muscle. MATERIALS AND METHODS: Human and rat bladder SMC were seeded separately within attached collagen lattices. Contractility of SMC was analyzed by measuring alterations in lattice diameter after exposure and release to the following contractile agonists: carbachol (10(-7)-10(-3) microM), calcium-ionophore (10 microM), lysophosphatidic acid (LPA) (1 microM), endothelin (0.1 microM), KCl (3.33 mmicroM) angiotensin II (10 microM), and serotonin (100 microM). Results were recorded as a mean reduction of the lattice diameter. In addition, immunohistochemical analysis for phenotypic markers of smooth muscle cell differentiation was performed on bladder SMC cultured within collagen lattices. Human palmar fascia fibroblasts, which have been previously well characterized by in vitro contractility and immunohistochemistry, were tested in parallel and used as controls for all the above experiments. RESULTS: Human SMC had significant contractile responses to calcium-ionophore (31% +/- 4 relative percent contraction, p <0.05), LPA (34% +/- 4, p <0.05), and endothelin (37 +/- 5%, p <05). There was no significant contraction in response to carbachol, angiotensin II, KCl, or serotonin. Rat bladder SMC had a similar contractile response but did not contract in response to endothelin. In contrast to human and rat bladder SMC, fibroblasts did not contract to calcium-ionophore. CONCLUSIONS: In vitro cultured bladder SMC demonstrate loss of contractile response to normal in vivo pharmacologic agonists. Both human and rat bladder SMC can be distinguished in vitro from fibroblasts based upon their lack of contractile response to calcium- ionophore. These results demonstrate the ability to further characterize cultured bladder SMC with in vitro contractility. Further characterization is essential if we are to advance our understanding of the clinical applicability of in vitro studies utilizing cultured bladder SMC. PMID- 10524935 TI - Electrical stimulation has no adverse effect on pregnant rats and fetuses. AB - PURPOSE: Electrical stimulation has been considered a contraindication in pregnant women with various voiding dysfunctions, because of the potential to cause teratogenicity or abortion. However, it is not known whether electrical stimulation can cause fetal malformation or abortion. The purpose of this study is to evaluate whether electrical stimulation has any adverse effect on pregnant rats and fetuses. MATERIALS AND METHODS: Twenty Sprague-Dawley pregnant rats were divided into two groups: electrical stimulation group (n = 10) and sham controls (n = 10). Rats in the stimulation group were stimulated 7 hours every day from Day 4 to Day 20 of gestation. All pregnant rats were sacrificed and fetuses were examined at near term (Day 20 of gestation). The number of fetuses, resorptions, fetal liability, body weight and gross appearance were recorded. Viscera and skeleton stained with Alizarin Red S were examined under stereoscope. RESULTS: All pregnant rats were healthy during the gestation period and no abortions were noted. Fetal body weight in the stimulation group (2.27 +/- 0.51 gm.) was not significantly different from sham group (2.13 +/- 0.51 gm.; p = 0.91). No significant difference was found in the number of resorptions between both groups. All fetuses were alive at the time of cesarean section. No fetal malformation was observed in gross appearance, viscera and skeleton of all rats. CONCLUSIONS: Electrical stimulation did not have any adverse effect on pregnant rats and their fetuses. Termination of pregnancy is not advised for prospective mothers when electrical stimulation has been performed inadvertently in early pregnancy. PMID- 10524936 TI - Association between outcome and telomere DNA content in prostate cancer. AB - PURPOSE: To perform an initial retrospective investigation of the relationship between outcome in patients with organ confined prostate adenocarcinoma and the tumor cells' content of telomere DNA. MATERIALS AND METHODS: The case-controlled study group was composed of eighteen men diagnosed with prostatic adenocarcinoma prior to 1993. The group was selected so that approximately one half died within ten years of diagnosis and one half survived ten years or longer. Archival, paraffin-embedded tumor tissue was recovered for each patient. DNA was extracted from newly cut sections, fixed to nylon membranes and hybridized with P32-labeled centromere- and telomere-specific probes. Telomere DNA contents were quantitated from the hybridized radioactivities. The relationships between telomere DNA content and survival, and telomere DNA content and disease recurrence in men receiving prostatectomies were determined. RESULTS: Death and disease recurrence were associated with reduced telomere DNA content (p <0.0001, p <0.0001, respectively). CONCLUSIONS: Telomere DNA content may differentiate high-risk patients with metastatic prostate cancer from men with indolent disease who can be spared the unnecessary side effects and expense of treatment by management with "watchful waiting." PMID- 10524937 TI - Characterization of alpha-adrenoceptor subtypes in the corpus cavernosum of patients undergoing sex change surgery. AB - PURPOSE: To characterize the subtypes of alpha1- and alpha2-adrenoceptors in the human corpus cavernosum from patients undergoing sex change surgery. MATERIALS AND METHODS: Saturation and competition radioligand binding studies were performed for characterization at the protein level. Alpha1-adrenoceptors were labeled with [3H]prazosin and [3H]tamsulosin, while alpha2-adrenoceptors were labeled with [3H]RX 821002. Alpha1-adrenoceptor subtype mRNA was additionally determined by reverse-transcriptase polymerase chain reaction and RNase protection assays. RESULTS: Human corpus cavernosum expressed approximately 32 and approximately 22 fmol./mg. protein alpha1- and alpha2-adrenoceptors, respectively. Competition studies with the alpha1A-selective antagonists 5 methylurapidil and (+)-niguldipine and the alpha1D-selective BMY 7378 revealed a mixed alpha1A/alpha1B-adrenoceptor population with no evidence for alpha1D adrenoceptor protein. In contrast alpha1D-adrenoceptors were readily detected at the mRNA level. Competition binding studies with the alpha2A-selective oxymetazoline and the alpha2B-selective prazosin and ARC 239 revealed a homogeneous population of alpha2A-adrenoceptors. CONCLUSIONS: We conclude that human corpus cavernosum expresses predominantly alpha1A-, alpha1B- and alpha2A adrenoceptor protein; additionally the alpha1D-adrenoceptor is present at the mRNA level. PMID- 10524938 TI - Transdifferentiation of prostate cancer cells to a neuroendocrine cell phenotype in vitro and in vivo. AB - PURPOSE: To better understand the source of neuroendocrine cells associated with human prostate cancer progression, we studied the ability of a cultured prostate cancer cell line, LNCaP, to transdifferentiate into neuroendocrine-like cells in vitro and in vivo. MATERIALS AND METHODS: Cyclic AMP concentrations were measured in extracts of LNCaP cells cultured in the presence of normal or hormone deficient medium (containing charcoal-stripped serum) with the use of an immunoassay. Quantitative RT-PCR procedures were used to determine whether hormone depletion affects TGF-beta2 mRNA expression. Western blotting procedures (for neuron specific enolase [NSE]) were used to determine whether TGF-beta2 supplementation or antibody neutralization might affect the ability of cultured LNCaP cells to transdifferentiate to neuroendocrine-like cells. Finally, tumors formed from LNCaP cells xenografted into male nude mice were evaluated for the presence of neuroendocrine cells (prior and subsequent to castration of the host mouse) using an immunohistochemical stain for chromogranin A. RESULTS: LNCaP cells cultured in a hormone-deficient medium have a mean 9-fold increase in cyclic AMP (p = 0.02) and a significant decline in the expression of TGF-beta2 mRNA when compared with cells grown in normal medium. Supplementation or depletion of TGF-beta2 did not affect the neuroendocrine conversion of LNCaP cells as assessed by NSE expression patterns. LNCaP tumors growing in castrated male nude mice were found to have significantly increased numbers of chromogranin A positive neuroendocrine cells (46/high powered field) when compared with tumors growing in intact male mice (3/high powered field) (p = 0.0038). CONCLUSIONS: Exposure of LNCaP cells to a hormone deficient medium drastically increased cyclic AMP production and this may identify the biochemical pathway through which hormone depletion induces a neuroendocrine conversion of prostate cancer cells. Hormone depletion also reduced TGF-beta2 mRNA expression and this finding was consistent with our inability to demonstrate any effect of TGF-beta2 on neuroendocrine conversion in vitro. Finally, our demonstration of increased neuroendocrine cells found in LNCaP tumors growing in castrated immunodeficient mice suggests that the neuroendocrine cells associated with advanced human prostate tumors in vivo, arise from prostate cancer cells through the transdifferentiation process. PMID- 10524939 TI - Immunotherapy of bladder cancer targeting P53. AB - PURPOSE: Superficial bladder cancer is often responsive to immunotherapy with bacillus Calmette-Guerin (BCG). However, some tumors progress despite BCG treatment, and most of these have mutations in the p53 tumor suppressor gene resulting in its over-expression. Overexpressed p53 is therefore a potential target for immunotherapy. The objective of this study was to demonstrate whether human bladder cancer xenografts in SCID mice could be eliminated by cytotoxic T cells (CTL) which recognize over-expressed p53. MATERIALS AND METHODS: Murine CTL which are specific for human p53 were previously generated in our laboratory by peptide immunization of HLA A2.1 transgenic mice. These CTL recognize and lyse human tumor cell lines which over-express p53 in the context of HLA A2.1. The p53 over-expressing HLA A2.1+ human bladder cancer cell line J82 was used to establish subcutaneous or intravesicular tumors in SCID mice. The mice were then administered tail vein injections of 5 x 10(7) p53-specific CTL, control CTL, or phosphate buffered saline (PBS). RESULTS: The subcutaneous tumor mean volume at 5 weeks for the p53-specific CTL treatment group was significantly lower than for both the control CTL or the PBS group (32 mm.3 versus 185 mm.3, p = 0.04 and 32 mm.3 versus 418 mm.3, p = 0.0001). In the mice with intravesicular tumors, a reduction to nonpalpable tumor size in vivo was seen with specific CTL therapy (14% palpable) versus control CTL treatment (86% palpable), the final tumor volume at necropsy was 127 mm.3 versus 246 mm.3 (N.S.). CONCLUSION: The overall response of the human bladder tumors in the SCID mouse model suggests the possibility of targeting p53 in patients with bladder cancer. PMID- 10524940 TI - Immunoregulatory potential of urothelium: characterization of NF-kappaB signal transduction. AB - PURPOSE: To characterize the basal and activated form of nuclear factor-kappaB (NF-kappaB) complex in normal urothelial cell cultures after stimulation with tumor necrosis factor-alpha (TNF-alpha), lipopolysaccharide (LPS), and double stranded ribonucleic acid (dsRNA). MATERIALS AND METHODS: Human urothelial cells cultured from normal bladder specimens underwent immunohistochemical staining and cellular extracts were prepared for Electrophoretic Mobility Shift Assays (EMSA), Western blot analyses, RNA isolation and Northern Blot analyses before and after stimulation with TNF-alpha, LPS, and dsRNA. RESULTS: In normal human urothelial cells, activation of the NF-kappaB complex in response to stimulation with TNF alpha, LPS, and dsRNA was detected by immunohistochemical methods and EMSA. Depending on the stimulus, a specific NF-kappaB complex was activated as seen by supershift experiments in EMSA. By Western blot, the inhibitor of NF-kappaB complex, IkappaB-alpha, degraded in response to stimulation. Northern blot analysis from total RNA revealed subsequent inducible interleukin-8 (IL-8) mRNA expression of normal urothelial cells when treated with TNF-alpha, LPS, and dsRNA. CONCLUSIONS: Normal human urothelial cells contain basal NF-kappaB complexes in an inactivated state. When these cells are challenged by different agents such as TNF-alpha, LPS, and dsRNA, the cells respond by activation of the NF-kappaB signal transduction pathway, degradation of its inhibitor, IkappaB alpha, and translocation of this primary factor into the nucleus to induce specific genetic responses such as IL-8 expression. PMID- 10524941 TI - Intraoperative use of the absorbable fibrin adhesive bandage: long term effects. AB - PURPOSE: The absorbable fibrin adhesive bandage (AFAB) reduces acute blood loss in experimental trauma models, but the effects on wound healing and subsequent function have heretofore not been investigated. Retropubic prostatectomy was selected to evaluate short and long term effects of using the AFAB intraoperatively. MATERIALS AND METHODS: Dogs undergoing prostatectomy were randomly assigned to one of four treatments: CONTROL- sponges and manual pressure were applied after transecting the prostatic pedicles. Sponges were removed when the prostate was delivered. Vessels were isolated and ligated if bleeding continued after removal. AFAB- hemostatically active bandages were applied to the prostatic bed prior to sponges and pressure. Additional bandages were applied at the urethrovesical junction after completing the anastomosis. PLACEBO- visually identical (hemostatically inert) bandages were applied in an identical fashion. LIQUID SEALANT- concentrated thrombin and fibrinogen solution was applied to the vessels prior to sponges and pressure. Additional sealant solution was applied around the anastomosis. RESULTS: Blood loss and time to achieve hemostasis were significantly less in the AFAB group compared with the other treatments. There were no differences in days to anastomotic integrity, continence, or intra abdominal adhesions at necropsy six weeks later. CONCLUSIONS: The AFAB can reduce surgery time and blood loss, with no decrement in wound healing or subsequent function. PMID- 10524942 TI - Differential effects of sex hormones and phytoestrogens on peak and steady state contractions in isolated rabbit detrusor. AB - PURPOSE: Recent evidence suggests that sex steroids may produce rapid inhibition of voltage operated Ca2+ channels (VOCCs). Detrusor smooth muscle is highly dependent upon Ca2+ influx for receptor-activated contractions. Thus, we examined the relative effectiveness of a select group of sex steroids and dietary phytoestrogens to relax detrusor contracted with the muscarinic receptor agonist, bethanechol (BE) and the purinergic P2X receptor agonist, alpha,beta-methylene ATP (alpha,beta-MeATP). MATERIALS AND METHODS: Isolated strips of rabbit detrusor were secured to isometric force transducers in a tissue bath and length-adjusted until maximum contractions were achieved. Peak (P) contractile responses were recorded for alpha,beta-MeATP (P(ATP)) and BE (P(BE)) and steady-state (SS) responses were recorded for BE (SS(BE)) in the presence and absence of selected sex steroids and phytoestrogens (10 microM, unless indicated). RESULTS: The L type VOCC inhibitor, nifedipine (1 to 10 microM), completely inhibited P(ATP) but reduced SS(BE) by approximately 50%, whereas the VOCC and non-VOCC inhibitor, SKF 96365, inhibited SS(BE) by approximately 95%, suggesting that P(ATP) was entirely dependent on L-type VOCCs, but (BE)-induced contractions depended also on activation of non-VOCCs. 17Beta-estradiol (estradiol) and progesterone inhibited P(ATP) by approximately 60% and 20%, respectively, and 32 microM estradiol and ethinyl estradiol inhibited SS(BE) by approximately 80 and 95%, respectively. Inhibition by estradiol was potentiated, rather than blocked, by the nuclear estrogen receptor antagonist, tamoxifen. Moreover, tamoxifen alone nearly completely relaxed SS(BE). The inactive metabolite of estradiol, 17alpha estradiol, inhibited both P(ATP) and P(BE) by approximately 40%. Testosterone had no effect on P(ATP) and P(BE). The phytoestrogen and tyrosine kinase inhibitor, genistein, inhibited SS(BE) by 44%, whereas daidzein, a phytoestrogen without tyrosine kinase inhibitory activity, produced only a 7% inhibition. None of the phytoestrogens examined inhibited P(BE), whereas all inhibited P(ATP) by approximately 20 to 35%. A comparison of inhibition of (BE) and alpha,beta-MeATP induced contractions by selected estrogen isomers showed some distinct differences. For example, estrone did not inhibit P(BE) or SS(BE), but inhibited P(ATP) by approximately 20%, whereas DES inhibited SS(BE) by nearly 90%, but P(ATP) by a lesser degree (approximately 70%). CONCLUSIONS: Our data support the hypothesis that 17beta-estradiol, ethinyl estradiol, DES, tamoxifen and genistein may relax detrusor contractions by inhibition of both VOCCs and non-VOCCs. Moreover, our data show that genistein, a dietary phytoestrogen with tyrosine kinase inhibitory activity, selectively reduced alpha,beta-MeATP-induced peak and BE-induced steady-state contractions, sparing the maximum response to BE. Lastly, the inactive isomer, 17alpha-estradiol, inhibited both BE- and alpha,beta-MeATP induced contractions. These data suggest that certain dietary phytoestrogens (for example, genistein) or sex steroids, especially those with weak activity at the nuclear steroid site (for example, 17alpha-estradiol), or tamoxifen may prove therapeutically useful in treating overactive bladder caused by elevated muscarinic and purinergic receptor activation. PMID- 10524943 TI - Evidence for predominant mediation of alpha1-adrenoceptor in the tonus of entire urethra of women. AB - PURPOSE: We separated the entire length of the isolated human female urethra into seven parts from external urethral meatus to bladder neck and examined regional differences in contractile responses to noradrenaline, clonidine, acetylcholine and KCl. MATERIALS AND METHODS: The entire urethra was obtained from 9 female patients with a mean age of 72.2 +/- 1.8 years. The entire urethra (35 to 42 mm. in length) was transversely cut into seven parts, and the contractile responses to noradrenaline, clonidine, acetylcholine and KCl of these parts were examined. RESULTS: Noradrenaline but not clonidine produced concentration-dependent contraction in all parts, with a peak amplitude in middle to proximal urethra. In contrast, acetylcholine produced contraction only in proximal urethra and bladder neck. The amplitudes of noradrenaline-induced contraction were normalized against those induced by KCl, revealing similarity in patterns between noradrenaline induced contractions and urethral pressure profile in human female urethra. These contractions to noradrenaline and acetylcholine were competitively inhibited by prazosin (pK(B): 8.38 +/- 0.10) and atropine (pK(B): 8.52 +/- 0.43), respectively. CONCLUSION: These findings suggest that sympathetic innervation helps maintain resting urethral tonus, mainly through alpha1-adrenoceptors. PMID- 10524944 TI - A quantitative study of atropine-resistant contractile responses in human detrusor smooth muscle, from stable, unstable and obstructed bladders. AB - PURPOSE: The objective of the study was to quantify in vitro the magnitude of atropine-resistant contractions using human detrusor samples and to determine the cellular processes underlying these contractions. MATERIALS AND METHODS: Isometric contractile responses were measured in isolated strips of human detrusor muscle obtained from patients with i) stable, ii) unstable or iii) obstructed bladders. Preparations were electrically stimulated or exposed to carbachol and ATP in the superfusate. RESULTS: Force-frequency curves were shifted to the right in samples from unstable and obstructed bladders. These same tissue groups also showed significant atropine-resistant contractions which were abolished by the neurotoxin TTX, or the non-hydrolysable ATP analog, alpha,beta methylene ATP, suggesting that these contractions were mediated by neurally released ATP. Sub-division of the patient group with unstable bladders demonstrated that those with neuropathic instability did not show atropine resistance, whereas those with idiopathic instability or secondary instability after obstruction did show atropine-resistant contractions. The potency of carbachol in generating a contracture was significantly greater than ATP (mean EC50 0.65 microM and 151 microM respectively) however, for each agonist there was no difference in potency between the three patient groups. Direct muscle excitability was similar in all three patient groups. CONCLUSIONS: It is concluded that purinergic, atropine-resistant contractions are present in some types of dysfunctional bladder, and these are not caused by a differential sensitivity of the muscle to ATP and cholinergic agonists. PMID- 10524945 TI - A comparison of the mode of action of ATP and carbachol on isolated human detrusor smooth muscle. AB - PURPOSE: The objectives of the study were: i) to examine the ability of carbachol and ATP to raise intracellular [Ca2+] in isolated detrusor myocytes; ii) to determine the origin of the intracellular Ca2+ and iii) to address the question of whether the appearance of purinergic contractions in detrusor from unstable and obstructed human bladders is reflected in the sensitivity of the cell to the two agonists. MATERIALS AND METHODS: Intracellular Ca2+ transients generated by extracellular ATP and carbachol were recorded from isolated human detrusor myocytes. Cells were dissociated by collagenase disruption of the biopsy. Intracellular Ca2+ was measured by epifluorescence microscopy using Fura-2 and electrophysiological recordings were made with patch electrodes. RESULTS: In cells from stable bladder biopsies the half-maximal concentrations (EC50) for ATP and carbachol to generate Ca2+ transients were 0.10 and 0.25 microM respectively. With cells from unstable bladders the EC50 values for both agonists and the magnitude of the Ca2+ transients were not significantly different from those obtained in cells from normal bladders. The transient in ATP was preceded by a transient depolarisation generated by a large inward current. The carbachol-Ca2+ transient was independent of changes to membrane potential, except in a subset of cells where complex membrane potential changes followed the rise of intracellular Ca2+. The ATP-Ca2+ transient was partially abolished by nicardipine and completely abolished by zero-Ca solutions, the carbachol-Ca2+ transient was unaffected by nicardipine and less completely attenuated by zero-Ca solutions. Prior exposure to caffeine suggested that the carbachol-Ca2+ transient, but not the ATP-Ca2+ transient, originated from intracellular stores. CONCLUSIONS: It is concluded that both agonists are equipotent in increasing intracellular Ca2+, but by different routes. The generation of purinergic contractions in detrusor from unstable bladder is not due to altered sensitivities of the detrusor myocyte to ATP or cholinergic agonists. PMID- 10524946 TI - Potentiation of erectile response and cAMP accumulation by combination of prostaglandin E1 and rolipram, a selective inhibitor of the type 4 phosphodiesterase (PDE 4). AB - PURPOSE: Phosphodiesterases (PDEs) are an important component of the signal transduction pathway during the erectile response. To determine the PDE isoforms in the corpora cavernosa in the cat and to establish the functional presence of PDE 4 in human cavernosal tissue, the erectile response to intracavernosal phosphodiesterase (PDE) inhibitors alone and the combination of PDE inhibitors and prostaglandin E1 (PGE1) was evaluated in the anesthetized cat. The in vitro formation of cAMP and cGMP in human cavernosal smooth muscle cells (HCSMCs) treated with PGE1 and rolipram in primary culture was also measured. MATERIALS AND METHODS: In pentobarbital-anesthetized cats, increases in intracavernosal pressure, penile length, and duration of erectile response were determined after intracavernosal injections of (i) the type 3 cAMP-specific, cGMP-inhibitable PDE inhibitor, milrinone, (ii) the type 4 cAMP-specific PDE inhibitor, rolipram, (iii) the type 5 cGMP-specific PDE inhibitor, zaprinast, and (iv) the combination of rolipram and PGE1. Systemic arterial pressure was concurrently assessed in these experiments. All responses to PDE inhibitors were compared with a control triple-drug combination comprised of papaverine (1.65 mg.), PGE1 (0.5 microg.), and phentolamine (25 microg.). HCSMCs were incubated with PGE1 (3 microM) and rolipram (10 microM) individually or in combination up to 2 hours at 37C. The intracellular cAMP and cGMP was extracted by cold absolute ethanol and measured (pmol./10(6) cells) by a commercially available EIA kit. RESULTS: Milrinone (3 to 100 microg.), rolipram (3 to 100 microg.), and zaprinast (3 to 100 microg.) induced dose-dependent increases in intracavernosal pressure and penile length (p <0.05) when administered intracavernosally. The maximum increase in cavernosal pressure in response to zaprinast was associated with no significant change in systemic arterial pressure. When rolipram was combined with PGE1 (0.1 microg.), the increases in intracavernosal pressure and the duration of erectile response were significantly higher (p <0.05) and longer (p <0.05) than those observed when rolipram alone was injected intracavernosally. PGE1 (3 microM) and rolipram (10 microM) produced significant increases (p <0.05) in the accumulation of intracellular cAMP levels in HCSMCs in primary culture above those of the baseline values while intracellular levels of cGMP did not change. CONCLUSIONS: PDE inhibitors administered intracavernosally caused dose-dependent increases in cavernosal pressure in the cat. When a specific cAMP PDE inhibitor was combined with PGE1, the erectile response was enhanced and intracellular levels of cAMP were increased in HCSMCs in primary culture. These data suggest further exploration of the combination of various PDE inhibitors and PGE1 in the pharmacologic treatment of erectile dysfunction and provide functional evidence for the presence of PDE 4 isoenzyme in human penile cavernosal cells. PMID- 10524948 TI - Virtual endoscopy for planning and simulation of minimally invasive neurosurgery. PMID- 10524947 TI - In vivo pressure measurements of lithotripsy shock waves in pigs. PMID- 10524949 TI - Outpatient treatment of middle and lower ureteric stones: extracorporeal shock wave lithotripsy versus ureteroscopic laser lithotripsy. PMID- 10524950 TI - Diagnosis and management of amebiasis. PMID- 10524951 TI - Photo quiz. Diagnosis: cutaneous miliary tuberculosis. PMID- 10524952 TI - Involvement of Panton-Valentine leukocidin-producing Staphylococcus aureus in primary skin infections and pneumonia. AB - Panton-Valentine leukocidin (PVL) is a cytotoxin that causes leukocyte destruction and tissue necrosis. It is produced by fewer than 5% of Staphylococcus aureus strains. A collection of 172 S. aureus strains were screened for PVL genes by polymerase chain reaction amplification. PVL genes were detected in 93% of strains associated with furunculosis and in 85% of those associated with severe necrotic hemorrhagic pneumonia (all community-acquired). They were detected in 55% of cellulitis strains, 50% of cutaneous abscess strains, 23% of osteomyelitis strains, and 13% of finger-pulp-infection strains. PVL genes were not detected in strains responsible for other infections, such as infective endocarditis, mediastinitis, hospital-acquired pneumonia, urinary tract infection, and enterocolitis, or in those associated with toxic-shock syndrome. It thus appears that PVL is mainly associated with necrotic lesions involving the skin or mucosa. PMID- 10524953 TI - Seasonal variation of Acinetobacter infections: 1987-1996. Nosocomial Infections Surveillance System. AB - To determine whether nosocomial infections due to Acinetobacter species have increased over the past 10 years and whether infections continue to have a pronounced seasonal variation, we analyzed infections reported by hospitals in the National Nosocomial Infections Surveillance System that performed adult and pediatric intensive care unit surveillance from 1987 through 1996. Overall, 3447 nosocomial acinetobacter infections were reported during 5,596, 156 patient-days. There was a yearly median of 7.2 infections (range, 5.0-10.5) per 10,000 patient days and a downward trend in the rate of acinetobacter infections overall (P<.05) and of 2 major types of infection (P<.05): bloodstream infections (yearly median, 1.6 per 10, 000 central venous catheter-days; range, 1.3-2.9) and pneumonia (yearly median, 7.6 per 10,000 ventilator-days; range, 6.5-12.0). Throughout this period, average rates were significantly higher during July-October than during November-June for acinetobacter infections overall (8.0 vs. 5.2; P<.01) and for bloodstream infections (2.0 vs. 1.2; P<.01) and pneumonia (9.7 vs. 6.6; P<.01). PMID- 10524954 TI - Tuberculosis in the inner city: impact of a continuing epidemic in the 1990s. AB - Tuberculosis cases have recently declined in the United States, renewing interest in disease elimination. We examined the epidemiology of tuberculosis from 1991 through 1997 at an inner-city public hospital and assessed population-based tuberculosis rates by ZIP code in the 8 metropolitan Atlanta counties. During the 7 years, 1378 new patients had tuberculosis diagnosed at our hospital (mean, 197 patients/year), accounting for 25% of tuberculosis cases in Georgia. Coinfection with human immunodeficiency virus (HIV) was common, but a significant decrease in the proportion of HIV-infected patients with tuberculosis was noted over time. Most patients were members of a minority group (93%) and were born in the United States (96%). Two inner-city ZIP code areas had annual tuberculosis rates >120 cases per 100,000 persons, and 8 ZIP code areas had annual rates of 47-88 cases per 100,000 persons between 1993 and 1997, compared with the annual national average of 8.7 cases per 100,000 persons. Our hospital continues to care for large numbers of tuberculosis patients, and rates of tuberculosis remain high in the inner city. These data mandate a concentration of efforts and resources in urban locations if tuberculosis control and elimination is to be achieved in the United States. PMID- 10524956 TI - Clinical and prognostic categorization of extraintestinal nontyphoidal Salmonella infections in infants and children. AB - The study included 172 patients, aged 0-15 years, for whom at least 1 nonfecal, nonurinary specimen was culture-positive for nontyphoidal Salmonella. Ninety-five percent had positive blood cultures. Immunocompromising diseases were found in 19% of 74 infants and 77% of 98 children. Associations between the study factors and outcomes, as localized infection or death, were assessed by logistic regression analysis. Thirty-three patients had localized infections. An adjusted risk factor for development of localized infections was an age of <12 months (P=.003). There were 17 deaths. The case-fatality rates were 43% and 10% for immunocompromised and 5% and 0% for nonimmunocompromised infants and children, respectively. Adjusted risk factors for death were age of <12 months (P=.006), inappropriate antimicrobial therapy (P=.014), meningitis or culture-proven pneumonia due to nontyphoidal Salmonella (P=.004), and immunocompromised status (P<.001). The clinical courses and prognoses for infants and children with extraintestinal infection due to nontyphoidal Salmonella can be categorized into 4 groups according to the characteristics of age (infants vs. children) and host status (immunocompromised vs. nonimmunocompromised). PMID- 10524955 TI - Prevalence of lower genital tract infections among human immunodeficiency virus (HIV)-seropositive and high-risk HIV-seronegative women. HIV Epidemiology Research Study Group. AB - This study was undertaken to assess whether the prevalence of lower genital tract infections among human immunodeficiency virus (HIV)-seropositive women was higher than among high-risk HIV-seronegative women at their baseline visit for the HIV Epidemiology Research Study. Results were available for 851 HIV-seropositive and 434 HIV-seronegative women. Human papilloma virus (HPV) infection was more prevalent among HIV-seropositive women (64% vs. 28%). Bacterial vaginosis was common (35% vs. 33%), followed by trichomoniasis (12% vs. 10%), syphilis (8% vs. 6%), Chlamydia trachomatis infection (4% vs. 5%), candidal vaginitis (3% vs. 2%), and Neisseria gonorrhoeae infection (0.8% vs. 0.3%). Alcohol use (odds ratio [OR], 1.8; 95% confidence interval [CI], 1. 3-2.4) and smoking (OR, 1.8; 95% CI, 1.3-2.5) were associated with bacterial vaginosis. Bacterial vaginosis (OR, 2.3; 95% CI, 1.5-3.4), trichomoniasis (OR, 2.3; 95% CI, 1.1-4.7), and syphilis (OR, 3.1; 95% CI, 1.3-7.4) were found to be more prevalent among black women. Our study showed no statistically significant difference in the prevalence of lower genital tract infections except for HPV between HIV-infected and demographically and behaviorally similar HIV-uninfected high-risk women. PMID- 10524957 TI - Pericystic metabolic activity in alveolar echinococcosis: assessment and follow up by positron emission tomography. AB - Information on parasite viability in alveolar echinococcosis (AE) cannot be obtained by conventional imaging techniques. We evaluated the glucose metabolism of AE lesions by use of [18F]fluorodeoxyglucose positron emission tomography (FDG PET) in 12 inoperable patients. Eight patients showed either perilesional or focal enhancement ("hot spots"), whereas 4 patients had nonenhancing (metabolically inactive) lesions. With PET, necrotic parasitic lesions and areas of enhanced metabolic activity could be clearly discriminated. Most notably, 3 of 8 patients with metabolically active lesions who were reexamined after chemotherapy treatment clearly showed improvement: the initial surrounding hot spots had disappeared in 2 of them, and had significantly decreased in 1. PET may prove valuable in assessing the efficacy of chemotherapy by showing the disappearance of metabolic activity and may also be useful for timely detection of relapses and metastases. Although costly and not readily available, FDG-PET is a promising tool toward improved management of AE and may thus help lower costs of long-term chemotherapy. PMID- 10524958 TI - The epidemiology of candidemia in two United States cities: results of a population-based active surveillance. AB - We conducted prospective, active population-based surveillance for candidemia (defined as any Candida species isolated from blood) in Atlanta and San Francisco (total population, 5.34 million) during 1992-1993. The average annual incidence of candidemia at both sites was 8 per 100,000 population. The highest incidence (75 per 100,000) occurred among infants 128 microg/mL, respectively). The efflux mechanism is the predominant form of macrolide resistance in the United States. PMID- 10524962 TI - Parenteral antibiotic use in acute-care hospitals: A standardized analysis of fourteen institutions. AB - Despite increasing concerns regarding the need to optimize appropriate antibiotic use in hospitals, a standardized method for evaluating interinstitutional antibiotic use has not been developed. To address this issue, antibiotic use was analyzed by means of a uniform methodology among 14 acute-care hospitals. Data were standardized by use of a defined daily dose for each antibiotic while adjusting for patient volume by calculating use per 1000 patient-days. Within the group, there was a 68% range in total parenteral antibiotic expenditures and wide variability in the use of individual agents. Analysis of these differences indicated that only the use of active antibiotic-management programs clearly correlated with antibiotic cost per 1000 patient-days (P<.001). Given these results, we believe that wider comparative analysis of antibiotic use with a standardized methodology in conjunction with standardized analysis of nosocomial infection rates and antibiotic resistance data may enhance the stewardship of antibiotics in acute-care hospitals. PMID- 10524963 TI - Diagnosis of venous access port-related infections. AB - The accumulation of infected clots under the silicone septum of the reservoir of venous access ports (VAPs) has been reported. We analyzed the relationship between these deposits and the occurrence of VAP-related bloodstream infections (VAP-BSIs) by (1) evaluating the accuracy of paired quantitative blood cultures for diagnosing VAP-BSI before the removal of the device and (2) assessing the accuracy of cultures of the tip and septum (i.e., the internal lumen of the VAP) for diagnosing VAP-BSI after removal of the device. Over a 16-month period, all VAPs removed were prospectively investigated. Before VAP removal, paired quantitative blood cultures were 77% sensitive and 100% specific and had a positive predictive value of 100% and a negative predictive value of 98% for diagnosing VAP-BSI. After VAP removal, tip culture was only 46% sensitive, whereas septum culture was 93.3% sensitive for confirming the diagnosis of VAP BSI. Thus infected deposits that accumulate under the VAP septum are the source of VAP-BSI. PMID- 10524966 TI - Molecular epidemiology of the global and temporal diversity of Candida albicans. AB - The epidemiology of Candida albicans has changed with the rise in immunocompromised patients and the pressures of antifungal treatment and prophylaxis. We assessed the genotype distribution of recently obtained, globally diverse isolates in comparison with isolates recovered in the United States and United Kingdom before 1985, in order to determine temporal and geographic differences. We used EcoRI digestion of cellular DNA to generate restriction fragment length polymorphisms, dividing the isolates into 4 groups. From 15 diverse geographic areas, 439 isolates obtained over 20 years were divided into 121 genotypes within groups A (289 isolates), B (85), C (56), and D (9). Differences in genotype distribution existed among the localities (P<.0001) and between isolates obtained before 1990 versus those recovered since then (P=.009). Comparison of pre-1985 United States/United Kingdom isolates with post-1994 United States isolates revealed a trend toward a changing genotype distribution (P=.057). Global post-1985 isolates were different in genotype distribution from United States/United Kingdom isolates (P<.0001). The distribution of isolates from Israel was unique (P<.0001). These differences could be due in part to the increasing prevalence of group C strains worldwide. PMID- 10524964 TI - Magnitude of the disease burden from neurocysticercosis in a developing country. AB - Cysticercosis contributes to higher epilepsy rates in developing countries than in industrialized ones, yet no estimate exists for the associated burden of disease. We used epidemiological data on neurocysticercosis in Peru to calculate the burden of disease and applied our model to the other countries of Latin America where neurocysticercosis is endemic to determine a regional estimate. Analysis of 12 population-based community studies demonstrated that neurocysticercosis was endemic in highland areas and high jungles, with seroprevalences from 6% to 24%. In one community, the adult seizure disorder rate was 9.1% among seropositive persons versus 4. 6% among seronegative persons; we used this difference for estimates. On the basis of average prevalence rates in areas of endemicity of 6%-10%, we estimated that there are 23,512-39,186 symptomatic neurocysticercosis cases in Peru. In Latin America, an estimated 75 million persons live in areas where cysticercosis is endemic, and approximately 400,000 have symptomatic disease. Cysticercosis contributes substantially to neurological disease in Peru and in all of Latin America. PMID- 10524965 TI - Aspergillosis in children with cancer: A 34-year experience. AB - A retrospective review of medical records, microbiology and pathology laboratory records, and nosocomial infection surveillance data was undertaken to describe the experience with culture-documented aspergillus infection in pediatric cancer patients at our facility. Sixty-six patients were identified from a 34-year period. The most common underlying diagnosis was leukemia. Risk factors included neutropenia, immunosuppression, and prior antibiotic therapy. On the basis of clinical presentation, 23 patients were believed to have disseminated disease and 43 to have localized disease. The lung was the most frequently affected organ. Despite aggressive medical and surgical management, overall mortality was 85% within the first year after diagnosis. Patients who presented with disease in sites other than the lungs fared better than patients with initial pulmonary involvement (P=.0014). Aspergillosis continues to be associated with poor outcome. Development of improved medical and adjuvant therapies, including surgery, is warranted. PMID- 10524967 TI - Crusted scabies: A molecular analysis of Sarcoptes scabiei variety hominis populations from patients with repeated infestations. AB - Crusted scabies is a severe debilitating disease due to hyperinfestation with the ectoparasite Sarcoptes scabiei. Treatment protocols include oral ivermectin and topical scabicides. After single-dose ivermectin, there may be early recrudescence, whereas after 3 doses at 14-day intervals, there is an apparent cure. However, such patients often present again after 6-12 months. To clarify the biology of recurrence, we studied genetic markers in sequential populations of S. scabiei mites from treated patients with multiple episodes of crusted scabies. Individual mites were genotyped at hypervariable microsatellite loci by a fluorescence-based polymerase chain reaction. Results indicated that sequential populations of mites were genetically more similar to each other than to mites from other patients. Although the majority of recurrent scabies is probably due to reinfestation from inadequately treated contacts, there was evidence that in very severe crusted scabies, treatment with even 3 doses of ivermectin 14 days apart may be inadequate and relapse may occur. PMID- 10524968 TI - Methicillin-resistant Staphylococcus aureus as a causative agent of postoperative intra-abdominal infection: relation to nasal colonization. AB - In the surgical intensive care unit of a university hospital, we investigated the frequency of and the risk factors for acquisition of methicillin-resistant Staphylococcus aureus (MRSA) during postoperative intra-abdominal infection (pIAI). We conducted a prospective MRSA nasal screening and case evaluation for 17 months among 73 consecutive patients with having pIAI. MRSA pIAI was diagnosed when MRSA was obtained from culture of intraperitoneal fluids. The identity of nasal and peritoneal MRSA strains was assessed by genomic analysis. Twelve patients had MRSA pIAI, representing 21% of all MRSA infections acquired by the 73 patients. An organ system failure score of >/=1 and MRSA nasal carriage prior to pIAI were the independent risk factors for acquisition of MRSA pIAI. Patients with MRSA pIAI had a longer intensive care unit stay and more reoperations than did those free of MRSA pIAI. We conclude that MRSA may be a causative pathogen in pIAI and may be related to nasal colonization. PMID- 10524969 TI - Bordetella pertussis and chronic cough in adults. AB - To evaluate Bordetella pertussis as a cause of persistent cough in adults, we examined 201 patients who had a cough for 2-12 weeks and no pulmonary disease. We obtained the following at presentation: medical history, chest radiograph, respiratory function measurement, nasopharyngeal aspirate for polymerase chain reaction (PCR), nasopharyngeal swab specimen for culture, and a blood sample (acute serum). Four weeks later a second blood sample (convalescent serum) was obtained. Control sera were obtained from 164 age-matched healthy blood donors with no history of cough during the previous 12 weeks. Four patients were B. pertussis culture-positive; 11 (including the culture-positive patients) were B. pertussis PCR-positive; and 33, including 10 of the 11 PCR-positive patients, had serological evidence of recent B. pertussis infection. Pertussis-positive and negative patients could not be discriminated by a history of cough. We conclude that B. pertussis infection is a common cause of persistent cough in adults. This is of concern, because these patients may be B. pertussis reservoirs from which transmission may occur to infants, in whom the disease can be devastating. PMID- 10524970 TI - O'nyong-nyong fever in south-central Uganda, 1996-1997: clinical features and validation of a clinical case definition for surveillance purposes. AB - O'nyong-nyong (ONN) fever, caused by infection with a mosquito-borne central African alphavirus, is an acute, nonfatal illness characterized by polyarthralgia. During 1996-1997, south-central Uganda experienced the second ONN fever epidemic ever recognized. Among 391 persons interviewed and sampled, 40 cases of confirmed and 21 of presumptive, well-characterized acute, recent, or previous ONN fever were identified through active case-finding efforts or during a household serosurvey and by the application of clinical and laboratory criteria. Among confirmed cases, the knees and ankles were the joints most commonly affected. The median duration of arthralgia was 6 days (range, 2-21 days) and of immobilization was 4 days (range, 1-14 days). In the majority, generalized skin rash was reported, and nearly half had lymphadenopathy, mainly of the cervical region. Viremia was documented in 16 cases, primarily during the first 3 days of illness, and in some of these, body temperature was normal. During this epidemic, the combination of fever, arthralgia, and lymphadenopathy had a specificity of 83% and a sensitivity of 61% in the identification of cases of ONN fever and thus could be useful for surveillance purposes. PMID- 10524971 TI - Questionable history of immediate-type hypersensitivity to penicillin in Staphylococcal endocarditis: treatment based on skin-test results vers-us empirical alternative treatment--A decision analysis. AB - Approximately 10% of the population claim to be allergic to penicillins, but only approximately 10%-30% of these have IgE-mediated reactions to penicillin skin tests. Alternatives to penicillins may be less effective, more toxic, and more expensive. Therefore, we used decision analysis to calculate maximum expected utility and minimum cost for skin-testing or not skin-testing patients who have endocarditis due to Staphylococcus aureus that is susceptible to cloxacillin and who have a questionable history of immediate-type hypersensitivity to penicillin. We used known probabilities of intermediate outcomes, actual costs, and measured utilities and included one-way sensitivity analysis. Whether utility, cost, or average cost-utility was the outcome of interest, skin-testing was preferred to no skin-testing in most conditions. Patients who have endocarditis due to S. aureus that is susceptible to cloxacillin and who also have a questionable history of immediate-type hypersensitivity to penicillin should be skin-tested before starting antibiotic therapy. PMID- 10524972 TI - Carriage of multidrug-resistant Streptococcus pneumoniae and impact of chemoprophylaxis during an outbreak of meningitis at a day care center. AB - Three cases of meningitis due to multidrug-resistant serotype 14 Streptococcus pneumoniae occurred at a day care center (DCC) over 5 days. Cultures of nasopharyngeal samples were done at the index DCC, 2 comparison DCCs, and a pediatrics practice. Isolates were serotyped and subtyped by pulsed-field gel electrophoresis (PFGE) with SmaI. Pneumococcal carriage rates ranged from 44%-65% at the 3 DCCs and 29% in the pediatrics practice. Carriage of multidrug-resistant serotype 14 S. pneumoniae was noted in 13%-19% of children at the 3 DCCs. An outbreak strain was identified by PFGE at the index DCC and 1 other DCC; a closely related strain was found in the third DCC. Carriage of the outbreak strain was associated with being age 0-24 months, antibiotic use, upper respiratory tract infections, and otitis media. DCC contacts of the ill children were offered chemoprophylaxis with rifampin and clindamycin, which produced a profound but transient decrease in carriage. No additional cases occurred. PMID- 10524973 TI - Multidrug-resistant Streptococcus pneumoniae: An opportunity to further understand pneumococcal ecology and to better predict intervention outcomes. PMID- 10524974 TI - An outbreak of vancomycin-dependent Enterococcus faecium in a bone marrow transplant unit. AB - Outbreaks of vancomycin-resistant enterococci (VRE) are well described. The presence of mutants of VRE, such as vancomycin-dependent enterococci (VDE), in individual patients has been documented, but their potential to spread nosocomially has not been known. We present the first cluster of patients who acquired VDE nosocomially. Five bone marrow transplantation patients were infected or colonized by a genotypically indistinguishable multiantibiotic resistant strain of Enterococcus faecium. Vancomycin dependence in 3 of the 5 isolates was demonstrated. All cluster patients had received protracted prophylactic treatment with vancomycin (mean, 22.6 days), and specimens from >/=2 body sites were repeatedly culture-positive for the outbreak strain. The outbreak was controlled with aggressive infection control strategies, and prophylactic antibiotic policies were revised. Awareness of the potential for nosocomial spread of multiantibiotic-resistant VDE is vital for the care of immunocompromised patients, especially those receiving prophylactic antibiotics. PMID- 10524975 TI - Marked differences in pneumococcal carriage and resistance patterns between day care centers located within a small area. AB - Carriage rates of Streptococcus pneumoniae and their antibiotic resistance, capsular types, and genetic patterns were studied among 264 children aged 12-35 months attending 8 day care centers located within a 2.5-mile radius in the same city. Nasopharyngeal cultures were obtained within a 2-month interval from all 264 children. Significant differences in each of the studied characteristics were found between day care centers, and each day care center had a unique pattern of the carried pneumococci. Our findings show that day care centers are independent microenvironments and emphasize their role in the transmission and augmentation of antibiotic-resistant S. pneumoniae in the community. PMID- 10524976 TI - A multistate nosocomial outbreak of Ralstonia pickettii colonization associated with an intrinsically contaminated respiratory care solution. AB - From 1 February through 30 April 1998, 4 hospitals reported a total of 34 patients colonized with Ralstonia pickettii. All but 1 had been exposed to 0.9% saline solution manufactured by 1 company (Modudose; Kendall, Mainsfield, MA), which was used during endotracheal suctioning. Culture of saline solution from previously unopened vials yielded R. pickettii. All available product and patient isolates were genotypically related by pulsed-field gel electrophoresis (PFGE) analysis. The contaminated saline solution was manufactured at the same plant that had been associated with a similar outbreak in 1983. The 1983 and 1998 R. pickettii isolates were unrelated, as determined by PFGE. In both 1983 and 1998, a 0. 2-microm cartridge filter was used for terminal sterilization. The detection of R. pickettii should alert hospital personnel to the possibility of product contamination. In this outbreak, prompt notification of public health agencies resulted in rapid notification of other health care providers, which likely prevented additional outbreaks. PMID- 10524977 TI - Skin hygiene and infection prevention: more of the same or different approaches? AB - The purpose of this article is to review research indicating a link between hand hygiene and nosocomial infections and the effects of hand care practices on skin integrity and to make recommendations for potential changes in clinical practice and for further research regarding hand hygiene practices. Despite some methodological flaws and data gaps, evidence for a causal relationship between hand hygiene and reduced transmission of infections is convincing, but frequent handwashing causes skin damage, with resultant changes in microbial flora, increased skin shedding, and risk of transmission of microorganisms, suggesting that some traditional hand hygiene practices warrant reexamination. Some recommended changes in practice include use of waterless alcohol-based products rather than detergent-based antiseptics, modifications in lengthy surgical scrub protocols, and incorporation of moisturizers into skin care regimens of health care professionals. PMID- 10524978 TI - Toward the development of antibacterial vaccines: report of a symposium and workshop. Organizing Committee. AB - On 26 and 27 October 1998, the Department of Medicine at the University of California, San Francisco (UCSF), hosted a symposium and workshop on bacterial vaccines. The symposium featured invited speakers who are internationally recognized authorities in their fields and who discussed selected topics related to specific pathogens or specific principles of bacterial vaccine development. The workshop, held on the day following the symposium, brought together the invited speakers and members of the organizing committee, who came from UCSF and the University of California, Berkeley, to discuss 4 specific topics and to define priorities for future vaccine development. Considerable knowledge has been gained from successful and unsuccessful vaccine development efforts, and large gains in knowledge relevant to vaccine development have resulted from studies of basic immunology and microbial pathogenesis. This report summarizes the presentations at the symposium and the discussions of the workshop sessions. PMID- 10524980 TI - Answer to photo quiz (see pages 1126-7) PMID- 10524979 TI - Dr. Kiyoshi Shiga: discoverer of the dysentery bacillus. AB - The clinical manifestations of dysentery have been described for centuries, and the prototypic bacterial agent, Shigella dysenteriae, was identified 100 years ago. In the English language there has been remarkably little written about Dr. Kiyoshi Shiga, discoverer of the dysentery bacillus. We submit a brief biography of Dr. Shiga and the circumstances leading to his discovery, which proved the bacterial etiology of nonamebic dysentery. PMID- 10524981 TI - Bacteroides fragilis bacteremia and infected aortic aneurysm presenting as fever of unknown origin: diagnostic delay without routine anaerobic blood cultures. AB - We report the case of a 71-year-old male with Bacteroides fragilis bactermia and infected aortic aneurysm that went undiagnosed, in part, because routine anaerobic blood cultures were not obtained. Bacteremia caused by anaerobes has been reported to be declining, and recommendations to discontinue routine anaerobic blood cultures have been implemented in some hospitals. To our knowledge, this is the first report of an anaerobic bacteremia and infection that had a delay in diagnosis due to this change in blood-culturing protocol. The potential impact of deleting anaerobic blood cultures from routine protocols is discussed. PMID- 10524982 TI - Hepatitis E virus infection in travelers. AB - Hepatitis E virus (HEV) is a major cause of clinical hepatitis in regions of endemicity, affecting primarily young adults and travelers to these areas. We present 5 cases of acute HEV infection in travelers and review 143 cases of HEV infection found by a literature search that were contracted in areas of endemicity. Fulminant hepatitis occurred in 2.7% of the reported cases; 2 of these were fatal. The destination of most of the travelers with acute HEV infection was the Indian subcontinent. The overall risk of contracting HEV infection for travelers appears to be lower than the risk for hepatitis A virus infection. Pregnant women and individuals with underlying liver disease may be a risk for severe infection. PMID- 10524983 TI - Entamoeba histolytica and Entamoeba dispar: epidemiology and comparison of diagnostic methods in a setting of nonendemicity. AB - Recent studies suggest that stool antigen assays are more sensitive and specific than microscopy for the diagnosis of Entamoeba histolytica infection. One hundred twelve patients presenting at 3 centers with symptoms or risk factors of E. histolytica infection were prospectively enrolled in this study to evaluate new diagnostic tests for infections with E. histolytica and Entamoeba dispar. Four ELISA-based stool antigen kits for detecting E. histolytica or E. dispar were blindly compared with stool microscopy. Amebic serology was assessed by indirect hemagglutination. When antigen assays were used as the reference standard, microscopy performed at referral centers was more specific (68.4% vs. 9.5%) but less sensitive (70.4% vs. 92.1%) than microscopy performed in community laboratories. Diagnosis with the E. histolytica test and Merlin Optimun S ELISA indicated that only 3 (4.2%) of 72 coproantigen-positive stools were positive for E. histolytica. Indirect hemagglutination was a good predictor of E. histolytica infection when titers of antibody to ameba were >/=1:512. PMID- 10524984 TI - Anal colonization of group G beta-hemolytic streptococci in relapsing erysipelas of the lower extremity. AB - Four patients who had frequent relapses of erysipelas but no obvious portal of entry and no beta-hemolytic streptococci in specimens from conventional culture sites all had group G streptococci in cultures of specimens from the anal canal. It is suggested that anal colonization with group G streptococci, and possibly group A and other beta-hemolytic streptococci, may constitute a reservoir for streptococci in such cases. PMID- 10524985 TI - Epidemiological characteristics of spotted fever in Israel over 26 years. AB - During the summer of 1997, 2 confirmed and several suspected fatal cases of spotted fever (SF) occurred in previously healthy young adults in Israel. This unusual cluster of events stimulated the current study. The incidence of SF in Israel from 1971-98 was analyzed. Incidence increased until 1980, declined until 1994, and increased slightly from 1994-97. Incidence was higher during the summer, among children aged 0-9 years, and in rural settlements in central Israel. From 1971-1997, 31 deaths were reported, mostly in the elderly. The deaths that occurred in 1997 are a reminder that, despite the fact that morbidity due to SF is described mainly in children, SF can have a rapidly fatal outcome in healthy young adults. Thus, even during periods of low incidence, careful monitoring and high awareness for prompt diagnosis and treatment are needed. PMID- 10524986 TI - Melioidosis in Southern Vietnam: clinical surveillance and environmental sampling. AB - From 1992-1998, Burkholderia pseudomallei was isolated from only 9 (0.25%) of 3653 cultures of blood from febrile patients admitted to the Centre for Tropical Diseases in Ho Chi Minh City, an infectious disease referral center for southern Vietnam. Soil was sampled from 407 sites in 147 rice fields along the 5 major roads radiating from Ho Chi Minh City. B. pseudomallei was isolated from 73 sites (18%) in 39 rice fields (27%), but only 15 (21%) of the 71 isolates from 9 (6%) of 147 fields were the virulent l-arabinose (ara)-negative biotype. All except 1 of the fields with the ara-negative biotype were close to the homes of the patients with melioidosis. The low incidence of melioidosis in the provinces around Ho Chi Minh City may be explained by the restricted distribution of ara negative B. pseudomallei in the soil in this area. PMID- 10524987 TI - Low risk of vertical transmission of hepatitis C virus by breast milk. AB - To evaluate the risk of hepatitis C virus (HCV) transmission via breast milk, we collected 76 samples of breast milk from 73 chronically HCV-infected women and serum samples from their 76 perinatally HCV-exposed children. Enzyme immunoassay and strip immunoblot assay were used for detection of antibodies to HCV, and reverse transcriptase-polymerase chain reaction analysis was used for detection of HCV RNA. None of the 76 samples of breast milk contained HCV RNA, whereas 37 (59.7%) of 62 mothers tested for HCV RNA had HCV viremia. Only 1 of the 76 breast fed infants had evidence of HCV infection. Because HCV infection in this child was detected 1 month after birth, it seems unlikely that it was transmitted by breast-feeding. These results indicate that HCV infection in pregnant women should not be a contra-indication for breast-feeding. PMID- 10524988 TI - Hepatitis B and pupil-sparing oculomotor nerve paresis. PMID- 10524989 TI - Pulmonary colonization with Pneumocystis carinii in human immunodeficiency virus negative patients: assessing risk with blood CD4+ T cell counts. PMID- 10524990 TI - Clearance of hepatitis G viremia in a human immunodeficiency virus-positive patient by high-activity antiretroviral therapy. PMID- 10524991 TI - St. Louis encephalitis with focal neurological signs. PMID- 10524992 TI - Spontaneously resolving pulmonary mucormycosis. PMID- 10524993 TI - Treatment of severe pulmonary blastomycosis with oral itraconazole: case report. PMID- 10524994 TI - Ceftriaxone therapy for syphilis: report from the emerging infections network. PMID- 10524995 TI - Bacteremia due to Dietzia maris in an immunocompromised patient. PMID- 10524996 TI - Development of listerial meningitis during ciprofloxacin treatment. PMID- 10524997 TI - Methicillin-resistant Staphylococcus aureus infection in a renal allograft recipient treated successfully with a novel new antimicrobial agent (linezolid): new treatment options for infections due to resistant organisms. PMID- 10524998 TI - Septic arthritis in the knee due to Mycobacterium xenopi in a patient undergoing hemodialysis. PMID- 10524999 TI - Chronic otitis media due to EF-4 bacteria. PMID- 10525000 TI - Lung abscess due to beta-lactamase-producing Pasteurella multocida. PMID- 10525001 TI - Lethal infection due to Armillifer armillatus (Porocephalida): A snake-related parasitic disease. PMID- 10525002 TI - Multidrug-resistant Salmonella associated with AmpC hyperproduction. PMID- 10525003 TI - Interstitial nephritis, thrombocytopenia, hepatitis, and elevated serum amylase levels in a patient receiving clarithromycin therapy. PMID- 10525004 TI - Valvular and myocardial abscesses due to Erysipelothrix rhusiopathiae. PMID- 10525005 TI - Subacute thyroiditis presenting as pyrexia of unknown origin in a patient with human immunodeficiency virus infection. PMID- 10525006 TI - Clostridium cadaveris bacteremia in an immunocompetent host. PMID- 10525007 TI - Reply PMID- 10525008 TI - Risk of Candida infection from contaminated aortic valve allografts. PMID- 10525010 TI - Reply PMID- 10525009 TI - Reply PMID- 10525011 TI - Necrotizing fasciitis associated with Klebsiella pneumoniae liver abscess. PMID- 10525012 TI - Orbital cellulitis due to Streptococcus pneumoniae in a previously healthy adult. PMID- 10525014 TI - Evidence in perinatal medicine: enough of trial and error? PMID- 10525015 TI - Role of ureaplasma urealyticum in lung disease of prematurity. AB - AIM: To examine the role of Ureaplasma urealyticum colonisation or infection in neonatal lung disease. METHODS: Endotracheal aspirates from ventilated infants less than 28 weeks of gestation were cultured for U urealyticum and outcomes compared in infants with positive and negative cultures. RESULTS: U urealyticum was isolated from aspirates of 39 of 143 (27%) infants. Respiratory distress syndrome (RDS) occurred significantly less often in colonised, than in non colonised infants (p=0.002). Multivariate logistic regression analysis showed that in singleton infants, ureaplasma colonisation was the only independent (negative) predictor of RDS (OR 0.36; p=0. 02). Both gestational age (OR 0.46; p=0.006) and isolation of U urealyticum (OR 3.0; p=0.05) were independent predictors of chronic lung disease (CLD), as defined by requirement for supplemental oxygen at 36 weeks of gestational age. Multiple gestation was also a major independent predictor of RDS and CLD. CONCLUSIONS: Colonisation or infection with ureaplasma apparently protects premature infants against the development of RDS (suggesting intrauterine infection). However, in singleton infants, it predisposes to development of CLD, independently of gestational age. Treatment of affected infants after birth is unlikely to significantly improve the outcome and methods are required to identify and treat the women with intrauterine ureaplasmal infection, before preterm delivery occurs. PMID- 10525016 TI - Blood pressure standards for very low birthweight infants during the first day of life. AB - Blood pressures during the first day of life were measured prospectively in 61 very low birthweight infants using umbilical or peripheral arterial lines. Video recordings of real time waveforms were reviewed. Blood pressure correlated linearly with birthweight and gestation. Comparison with available standards showed that infants weighing under 800 g had lower acceptable mean arterial pressure (MAP). The lower limits of MAP for infants between 26 to 32 weeks of gestation were numerically similar to the gestational ages. PMID- 10525017 TI - Reference range for serum cortisol in well preterm infants. AB - AIM: To establish a reference range for serum cortisol concentrations in preterm infants with a gestational age of less than 30 weeks during the first two weeks of life. METHODS: Infants were prospectively classified by the following exclusion criteria: surfactant administration, arterial hypotension, acute or uncontrolled infection, ventricular haemorrhage II degrees or above, serum glucose < 2.2 mmol/l, exchange transfusion, stress as a result of any kind of examination or nursing for at least 4 hours before blood sampling. The cortisol value was measured once using radioimmunoassay in each infant. RESULTS: In appropriate for gestational age (AGA) infants (n = 37, median gestational age 27.7 weeks, median birthweight 1030 g) the distribution of the cortisol concentrations was non-Gaussian. These had a nearly normal distribution, when log(10) values of the data were used. The points determined by mean (2 SD) on the logarithmic scale were transformed back to the original units to provide a reference range: 73-562 nmol/l. Gestational age was significantly (p = 0.033) associated with cortisol values (log(10)) with a regression coefficient (standard error) of -0.045 (0.020). Small for gestational age (SGA) infants (n = 8) had significantly higher cortisol values (median 357 nmol/l) than AGA infants (median 199 nmol/l) (p=0.028). CONCLUSIONS: There is a strictly defined reference range of serum cortisol concentrations in AGA preterm infants. PMID- 10525018 TI - Longitudinal measurements of 17alpha-hydroxyprogesterone in premature infants during the first three months of life. AB - AIMS: To determine normal concentrations of 17alpha-hydroxyprogesterone (17OHP) for premature infants. METHODS: 17OHP was measured in 66 consecutive premature infants once a week during the first month, and once every two weeks thereafter, until the age of 3 months. The 17OHP values in 100 full term healthy neonates on the third day of life served as controls. Blood was sampled on filter paper using a neonatal radioimmunoassay kit. Findings were correlated with gestational age, birthweight, mode of delivery, Apgar scores, presence of respiratory distress syndrome and intake of maternal steroids. RESULTS: Mean 17OHP was raised at 7 days of age (138.9, 46.3, 53.3, 29.9 nmol/l, respectively, for infants whose gestational age was under 29 weeks, 29 to 30 weeks, 31 to 32 weeks, and 33 weeks and above). It fell sharply in the first two weeks after which it gradually decreased further, reaching 32.7, 23.6, 16.9, and 13.0 nmol/l, respectively, by the age of 90 days. The mean (SEM) 17OHP concentration in full term infants on day 3 of life was 17.8 (8.9) nmol/l. These values were independent of the presence and severity of respiratory distress syndrome and of prenatal maternal steroids. CONCLUSIONS: The increased 17OHP concentrations found at birth fell to those found in term infants during the first three months of life in infants over 31 weeks of gestation. Postconceptional age is the most important factor determining 17OHP concentration. PMID- 10525019 TI - Influence of spironolactone on neonatal screening for congenital adrenal hyperplasia. AB - AIM: To determine if the diuretic spironolactone cross reacts with 17alpha hydroxyprogesterone (17OHP) in an enzyme linked immunosorbent assay (ELISA) kit used for the mass screening of congenital adrenal hyperplasia. METHODS: Concentrations of 17OHP on a blood filter paper disc were measured using an ELISA kit (kit C-7: ENZAPLATE N-17alpha -OHP-7; Chiron, Tokyo, Japan). The cross reactivity of spironolactone and its metabolites with 17OHP was determined. The concentrations of spironolactone and its metabolites in blood were measured using HPLC (high performance liquid chromatography). RESULTS: Spironolactone cross reacted with 17OHP using kit C-7 (0.01%), by increasing 17OHP concentration in a dose dependent manner. The blood concentration of spironolactone and its metabolites was nearly 900 ng/ml, high enough to show an additive effect on the 17OHP concentration. About 12% of the false positive cases screened using the kit were due to the administration of spironolactone. CONCLUSIONS: Spironolactone interferes with 17OHP concentrations, leading to false positive test results for CAH. PMID- 10525020 TI - Sodium potassium adenosine triphosphatase activity in preterm and term infants and its possible role in sodium homeostasis during maturation. AB - AIM: To investigate sodium (NA(+)) potassium (K(+)) adenosine triphosphatase (ATPase) activity in newborn infants at different gestational ages, to elucidate the mechanism underlying poor renal sodium conservation in preterm infants. METHODS: Fifty three healthy newborn infants, gestational age 30-42 weeks, were studied. Umbilical cord red blood cell Na(+) K(+)ATPase activity, plasma renin activity, and plasma aldosterone activities were measured in all of them. Red blood cell Na(+) K(+)ATPase activity was re-examined in eight preterm infants, one and two weeks after birth. Total and ouabain sensitive ATPase activity was measured spectrophotometrically using a method that couples ATP hydrolysis with NADH oxidation. RESULTS: Red blood cell Na(+) K(+)ATPase activity was significantly lower (p<0.01) in preterm babies with a gestational age below 35 weeks, compared with those with aged 35 weeks and above: 2.3 (0.8) and 6.7 (1.3) nmol NADH/minute/mg protein, respectively. There was no correlation between gestational age, Na(+) K(+)ATPase, plasma renin activity and aldosterone values either in the preterm or term babies. Two weeks after birth, irrespective of gestational age, the enzyme activity of the preterm babies increased to values similar to those observed in the term neonates at birth. CONCLUSION: The differences in sodium homeostasis between term and preterm babies are modulated via changes in Na(+) K(+)ATPase activity. PMID- 10525022 TI - Individualised pulse oximetry limits in neonatal intensive care. AB - AIM: To determine whether individualised limits for arterial oxyhaemaglobin saturation by pulse oximetry (SpO(2)) are more effective for detecting hypoxia and hyperoxia in sick newborn infants than setting fixed limits. METHODS: Six hundred and ninety two simultaneous measurements of SpO(2) and partial pressure of oxygen in arterial blood (PaO(2)) were made in 95 infants. The sensitivity and specificity for predicting hypoxia and hyperoxia using various fixed SpO(2) limits and also individualised SpO(2) limits, calculated using a standard equation, were determined and compared. RESULTS: None of the fixed limits for SpO(2) was both sensitive and specific for predicting hypoxia and/or hyperoxia. There was no difference between these and individualised limits. CONCLUSION: Individualised SpO(2) limits are no more effective than fixed SpO(2) limits for predicting hypoxia and/or hyperoxia in sick newborn infants. SpO(2) monitoring is not an ideal method for assessing PaO(2). PMID- 10525021 TI - Peripheral blood lymphocyte subpopulations in schoolchildren born very preterm. AB - AIM: To investigate whether lymphocytes or serum inflammatory markers are associated with obstructive lung disease and bronchial lability in schoolchildren born very preterm. METHOD: Lymphocyte subsets were studied in the peripheral venous blood of 29 such children (median age 8.8 years). Serum eosinophil cationic protein (ECP) and myeloperoxidase (MPO) concentrations and the association between them, lymphocyte subsets, and lung function were studied. Fourteen healthy children born at term, median age 9.1 years, served as controls. T lymphocytes (CD3), T lymphocyte subpopulations (CD4 and CD8), B lymphocytes (CD19), natural killer cells (CD16+56) and activation markers of T and B lymphocytes (CD23 and CD25) were determined using flow cytometry. Lung function was measured in all children both in the clinic and at home (Vitalograph Data Storage Spirometer). RESULTS: Compared with the controls, schoolchildren born very preterm had significantly lower CD4(+) T cell percentages and CD4:CD8 ratios (p < 0.05 for both), whereas natural killer cell percentages and serum ECP values were significantly higher (p < 0. 05). The very preterm schoolchildren had significantly lower spirometric values than the control group (p < 0.05)-except forced vital capacity. When all the subjects were considered together, a weak, but significant, negative association was observed between the bronchial responsiveness in peak expiratory flow, after a beta(2) agonist during home monitoring, and the CD4(+) T cell percentage (r = -0.45; p = 0.008) and the CD4:CD8 ratio (r = -0.50; p = 0.003), indicating a relation between bronchial lability and imbalance of T cell subpopulations. CONCLUSIONS: These results suggest that there is an inflammatory basis for lung function abnormalities in schoolchildren born very preterm. PMID- 10525023 TI - Echocardiographic flow pattern of patent ductus arteriosus: a guide to indomethacin treatment in premature infants. AB - AIM: To compare the efficacy and safety of an indomethacin treatment strategy based on serial echocardiographic measurement of patent ductus arteriosus (PDA) flow pattern with a standard protocol. METHODS: Neonates weighing less than 1500 g at birth, who required respiratory support, and who had developed symptomatic PDA, were studied. PDA was confirmed in all infants using colour Doppler echocardiography, and serial observations of the ductal flow pattern were made. Infants randomly assigned to receive conventional indomethacin treatment (protocol group) were given an initial dose of 0.2 mg/kg, followed by 0.1 or 0.2 mg/kg, depending on age, 12 hourly for two further doses, and were eligible for a second course. Those randomly assigned to the ductal flow pattern assessment (ECHO group) received further doses of indomethacin after 24 hours, only if their flow pattern was "pulsatile" or "growing." RESULTS: There was no significant difference in the primary outcome measures between the two groups. The closure rate was 89.1% and 87.2%, respectively, in the protocol and ECHO groups. The mean (SD) doses of indomethacin were significantly higher in the protocol group: 3.2 (1.4) doses compared with 1.6 (0.9) doses. There was a significantly higher incidence of hypoglycaemia, impaired urine output, and gastrointestinal bleeding in the protocol group. CONCLUSIONS: An indomethacin treatment strategy for PDA based on measurement of the ductal flow pattern is associated with a reduction in the total doses of indomethacin administered, and a reduced rate of complications, compared with a conventional protocol. There is no difference in closure rate. PMID- 10525024 TI - Prone and left lateral positioning reduce gastro-oesophageal reflux in preterm infants. AB - AIM: To examine the effect of body position on clinically significant gastro oesophageal reflux (GOR) in preterm infants. METHODS: Eighteen preterm infants with clinically significant GOR were studied prospectively using 24 hour lower oesophageal pH monitoring. Infants were nursed in three positions (prone, left, and right lateral) for 8 hours in each position, with the order randomly assigned. Data were analysed using analysis of covariance. RESULTS: The median (range) reflux index (RI) for the group was 13.8% (5.8-40. 4). There was no significant difference in the mean time spent in each position. RI (mean % (SEM)) was significantly less in prone (6. 3 (1.7)) and left lateral positions (11.0 (2.2)), when compared with the right lateral position (29.4 (3.2)); p<0.001. The mean (SEM) longest episodes (mins) of GOR were reduced by prone and left positions (8.6 (2.2) and 10.0 (2.4), respectively) compared with the right position (26.0 (3.9)); p<0.001. The mean (SE) number of episodes was reduced by prone (15.4 (2.8)) and left (24.6 (3.5)) positions when compared with right (41.6 (4.6)) (p<0.001). CONCLUSIONS: Prone and left lateral positions significantly reduce the severity of GOR, by reducing the number of episodes and the duration of the longest episodes. Such positioning offers a useful adjunct to the treatment in hospital of preterm infants with gastro-oesophageal reflux. PMID- 10525025 TI - Unexplained fever in neonates may be associated with hepatitis B vaccine. AB - AIM: To investigate whether hepatitis B vaccination has increased the number of cases of unexplained neonatal fever. METHOD: The files of all infants born from 1 January 1991 to 31 December 1992, in whom a diagnosis of "injected antibiotic" or "disease of temperature regulation" was recorded, were reviewed. Those who had unexplained fever of 38 degrees C or higher during the first three days of life were divided into two groups: infants who did not receive the hepatitis B vaccine (1991) and infants who did (1992). RESULTS: In 1992 the incidence of unexplained fever in hepatitis B vaccinated neonates was significantly higher than in the 1991 group of pre-vaccination neonates (35 out of 5819 (0.6%) vs 14 out of 5010 neonates (0.28%) respectively, p=0.013). CONCLUSIONS: The increase in the number of cases of unexplained neonatal fever seems to be associated with the introduction of routine hepatitis B vaccination on the first day of life. The possibility that an excess number of neonates will undergo unnecessary procedures and treatment to diagnose unexplained fever justifies planning a controlled study to determine whether these preliminary findings point to a significant problem. PMID- 10525026 TI - Effect of maternal anticonvulsant treatment on neonatal blood coagulation. AB - AIMS: To investigate the impact of maternal anticonvulsant use on the ability of cord blood to coagulate. METHODS: Cord blood prothrombin times were measured, over 15 years in a consecutive series of 137 term babies born to women taking phenobarbitone, phenytoin, and/or carbamazepine while pregnant. The response to parenteral vitamin K was measured in 83 neonates. RESULTS: Only 14 of the 105 babies born to the mothers who had therapeutic anticonvulsant blood concentrations at birth had a prolonged prothrombin time (outside the 95% reference range). None had an overt bleeding tendency. The abnormality was corrected within 2 hours by 1 mg of parenteral vitamin K, but rapid intravenous prophylaxis produced complications in three infants. CONCLUSIONS: A policy of giving vitamin K throughout the last third of pregnancy to all women being treated with anticonvulsants, as recently recommended, is not justified by the available evidence. The belief that there is a distinct, early form of neonatal vitamin K deficiency that is different from, and more dangerous than, the classic form of the disease, is not supported by a review of the published evidence. PMID- 10525027 TI - Neonatal focal temporal lobe or atrial wall haemorrhagic infarction. AB - AIMS: To describe two variants of infarction within the temporal lobe, associated with local matrix bleeding and mild to moderate intraventricular haemorrhage. METHODS: The files of 10 neonates, extracted from a sonographic study of 560 very low birthweight infants conducted between 1993 and 1997, were retrospectively examined. RESULTS: Seven lesions were located in the middle to posterior area of the temporal lobe, three others faced the atrium. All except two of those with a temporal site were VLBW infants with hyaline membrane disease. Except for one fatal case, intraventricular bleeding was mild to moderate. Computed tomograms or magnetic resonance imaging were used to illustrate the haemorrhagic nature of three lesions. Survivors of this so far undescribed entity who were followed up for more than 18 months did not have a uniform type of cerebral palsy but some scored in the low normal range on the Bayley Mental Development Index. One girl developed temporal lobe epilepsy. CONCLUSIONS: This pattern of injury seems to be one of venous infarction associated with temporal or para-atrial matrix haemorrhage. The temporal site fits the picture of venous infarction within the area drained by the inferior ventricular vein. A less constant lateral atrial vein, either draining into the basal or internal cerebral vein, is probably involved in the para-atrial lesion. Sonography may be the only practical tool currently available for detection in life. PMID- 10525028 TI - Concentration of nitric oxide products in bronchoalveolar fluid obtained from infants who develop chronic lung disease of prematurity. AB - AIMS: To determine if nitric oxide (NO) products (nitrate and nitrite) are increased in bronchoalveolar lavage (BAL) fluid obtained from infants who develop chronic lung disease of prematurity (CLD). METHODS: One hundred and thirty six serial bronchoalveolar lavages were performed on 37 ventilated infants (12 with CLD, 18 with respiratory distress syndrome (RDS), and seven control infants) who did not receive inhaled NO. RESULTS: During the first week of life nitrate concentration was between 25-31 micromol/l in all three groups. Thereafter, the concentration of BAL fluid nitrate decreased to 14 micromol/l and 5.5 micromol/l, respectively in the RDS and control groups by 14 days of age. In contrast, nitrate in the CLD infants remained constant until 28 days of age (31.3 micromol/l at day 14; p<0.05). In all BAL fluid samples the mean concentration of nitrite was <1.2 micromol/l throughout the first 28 days with no significant differences noted among the three groups. CONCLUSION: The similar concentration of BAL fluid nitrate in all groups during the first week of life suggest that NO may be important in the adaptation of the pulmonary circulation after birth. However, persistence of nitrate in the BAL fluid of infants with CLD during the second week may reflect pulmonary maladaptation, or, more likely, persisting pulmonary inflammation. PMID- 10525029 TI - Pharmacokinetics and dose requirements of vancomycin in neonates. AB - AIMS: To design and evaluate dosing guidelines for vancomycin based on data collected during routine use of the drug. METHODS: Following the observation that 66% of neonatal vancomycin trough concentrations were outside the target range, new dose guidelines were developed using a population pharmacokinetic approach. NONMEM (non-linear mixed effects model) was used to analyse dose histories and 347 concentration measurements collected during routine therapeutic drug monitoring in 59 neonates. RESULTS: Postconceptual ages in the patient group ranged from 26-45 weeks, weights from 0. 57-4.23 kg, and creatinine concentrations from 18-172 micromol/l. The population estimate of vancomycin clearance (l/h/kg) was 3. 56/creatinine concentration (micromol/l) with an interpatient coefficient of variation (CV) of 22% and volume of distribution 0.67 l/kg with a CV of 18%. Residual error was 4.5 mg/l. When the new recommendations on dosing were used prospectively in a separate group of neonates the proportion of acceptable troughs increased from 33% to 72%. CONCLUSIONS: The pharmacokinetics of vancomycin in neonates and young infants depend on weight and serum creatinine. Preliminary results from the new guidelines indicate an improvement on previous practice, but also an ongoing need to monitor concentrations. PMID- 10525030 TI - Children of the 90s II: challenges for the ethics and law committee. PMID- 10525031 TI - The Chamberlen family (1560-1728) and obstetric forceps. PMID- 10525033 TI - Chemokines and asthma: redundancy of function or a coordinated effort? PMID- 10525032 TI - Th2 cells and GATA-3 in asthma: new insights into the regulation of airway inflammation. PMID- 10525034 TI - Airway remodeling in asthma. PMID- 10525035 TI - The macula densa is worth its salt. PMID- 10525036 TI - Igalpha: B all that you can B. PMID- 10525037 TI - The interaction of nitric oxide, bradykinin, and the angiotensin II type 2 receptor: lessons learned from transgenic mice. PMID- 10525038 TI - ABC1: connecting yellow tonsils, neuropathy, and very low HDL. PMID- 10525039 TI - Two of the usual suspects, platelet-activating factor and its receptor, implicated in acute lung injury. PMID- 10525040 TI - Cooperation between Th1 and Th2 cells in a murine model of eosinophilic airway inflammation. AB - We have studied the actions of helper T lymphocyte-1 and -2 (Th1 and Th2) cells in an acute model of eosinophilic airway inflammation by infusing chicken ovalbumin-specific (OVA-specific) Th1 cells, Th2 cells, or both into unsensitized mice and challenging the mice with an OVA aerosol. OVA challenge after infusion of Th1 cells alone resulted in airway inflammation with lymphocytes and monocytes. Challenge after the infusion of Th2 cells alone resulted in minimal inflammation. In contrast, when Th1 and Th2 cells were transferred together, they cooperated to promote a robust eosinophil-predominant inflammatory response. Th1 cells alone were readily recruited to the airways after challenge, but in the absence of Th1 cells, Th2 cells did not accumulate in the airways. When transferred together, both Th1 and Th2 cells, as well as endogenous eosinophils, were effectively recruited. This recruitment was correlated with increased VCAM-1 expression in the medium- and large-sized vessels of the lung and could be inhibited by treating the mice with neutralizing antibodies to TNF-alpha or VCAM 1. These data indicate that Th2 cells require signals in addition to antigen for their effective recruitment to the airways. Th1 cells can provide these signals. PMID- 10525041 TI - Analysis of the human thymic perivascular space during aging. AB - The perivascular space (PVS) of human thymus increases in volume during aging as thymopoiesis declines. Understanding the composition of the PVS is therefore vital to understanding mechanisms of thymic atrophy. We have analyzed 87 normal and 31 myasthenia gravis (MG) thymus tissues from patients ranging in age from newborn to 78 years, using immunohistologic and molecular assays. We confirmed that although thymic epithelial space (TES) volume decreases progressively with age, thymopoiesis with active T-cell receptor gene rearrangement continued normally within the TES into late life. Hematopoietic cells present in the adult PVS include T cells, B cells, and monocytes. Eosinophils are prominent in PVS of infants 2 years of age or younger. In the normal adult and the MG thymus, the PVS includes mature single-positive (CD1a(-) and CD4(+) or CD8(+)) T lymphocytes that express CD45RO, and contains clusters of T cells expressing the TIA-1 cytotoxic granule antigen, suggesting a peripheral origin. PBMCs bind in vitro to MECA 79(+) high endothelial venules present in the PVS, suggesting a mechanism for the recruitment of peripheral cells to thymic PVS. Therefore, in both normal subjects and MG patients, thymic PVS may be a compartment of the peripheral immune system that is not directly involved in thymopoiesis. PMID- 10525042 TI - Differential expression of three T lymphocyte-activating CXC chemokines by human atheroma-associated cells. AB - Activated T lymphocytes accumulate early in atheroma formation and persist at sites of lesion growth and rupture, suggesting that they may play an important role in the pathogenesis of atherosclerosis. Moreover, atherosclerotic lesions contain the Th1-type cytokine IFN-gamma, a potentiator of atherosclerosis. The present study demonstrates the differential expression of the 3 IFN-gamma inducible CXC chemokines--IFN-inducible protein 10 (IP-10), monokine induced by IFN-gamma (Mig), and IFN-inducible T-cell alpha chemoattractant (I-TAC)--by atheroma-associated cells, as well as the expression of their receptor, CXCR3, by all T lymphocytes within human atherosclerotic lesions in situ. Atheroma associated endothelial cells (ECs), smooth muscle cells (SMCs), and macrophages (MO) all expressed IP-10, whereas Mig and I-TAC were mainly expressed in ECs and MO, as detected by double immunofluorescence staining. ECs of microvessels within lesions also expressed abundant I-TAC. In vitro experiments supported these results and showed that IL-1beta, TNF-alpha, and CD40 ligand potentiated IP-10 expression from IFN-gamma-stimulated ECs. In addition, nitric oxide (NO) treatment decreased IFN-gamma induction of IP-10. Our findings suggest that the differential expression of IP-10, Mig, and I-TAC by atheroma-associated cells plays a role in the recruitment and retention of activated T lymphocytes observed within vascular wall lesions during atherogenesis. PMID- 10525043 TI - Leptin protects mice from starvation-induced lymphoid atrophy and increases thymic cellularity in ob/ob mice. AB - Thymic atrophy is a prominent feature of malnutrition. Forty-eight hours' starvation of normal mice reduced the total thymocyte count to 13% of that observed in freely fed controls, predominantly because of a diminution in the cortical CD4(+)CD8(+) thymocyte subpopulation. Prevention of the fasting-induced fall in the level of the adipocyte-derived hormone leptin by administering exogenous recombinant leptin protected mice from these starvation-induced thymic changes. The ob/ob mouse, which is unable to produce functional leptin because of a mutation in the obese gene, has impaired cellular immunity together with a marked reduction in the size and cellularity of the thymus. We found that ob/ob mice had a high level of thymocyte apoptosis resulting in a ratio of CD4(+)CD8(+) (cortical) to CD4(-)CD8(-) (precursor) thymocytes that was 4-fold lower than that observed in wild-type mice. Peripheral administration of recombinant leptin to ob/ob mice reduced thymocyte apoptosis and substantially increased both thymic cellularity and the CD4(+)CD8(+)/CD4(-)CD8(-) ratio. In contrast, a comparable weight loss in pair-fed PBS-treated ob/ob mice had no impact on thymocyte number. In vitro, leptin protected thymocytes from dexamethasone-induced apoptosis. These data indicate that reduced circulating leptin concentrations are pivotal in the pathogenesis of starvation-induced lymphoid atrophy. PMID- 10525044 TI - Expression of functional CXCR4 chemokine receptors on human colonic epithelial cells. AB - In addition to their role as regulators of leukocyte migration and activation, chemokines and their receptors also function in angiogenesis, growth regulation, and HIV-1 pathogenesis--effects that involve the action of chemokines on nonhematopoietic cells. To determine whether chemokine receptors are expressed in human colonic epithelium, HT-29 cells were examined by RT-PCR for the expression of the chemokine receptors for lymphotactin, fractalkine, CCR1-10, and CXCR1-5. The only receptor consistently detected was CXCR4 (fusin/LESTR), although HT-29 cells did not express mRNA for its ligand, stromal cell-derived factor (SDF 1alpha). Flow cytometric analysis with anti-CXCR4 antibody indicated that the CXCR4 protein was expressed on the surface of roughly half of HT-29 cells. CXCR4 was also expressed in colonic epithelial cells in vivo as shown by immunohistochemistry on biopsies from normal and inflamed human colonic mucosa. The mRNA for SDF-1alpha and other CC and CXC chemokines was present in normal colonic biopsies. The CXCR4 receptor in HT-29 cells was functionally coupled, as demonstrated by the elevation in [Ca2+]i, which occurred in response to 25 nM SDF 1alpha and by the SDF-1alpha-induced upregulation of ICAM-1 mRNA. Sodium butyrate downregulated CXCR4 expression and induced differentiation of HT-29 cells, suggesting a role for CXCR4 in maintenance and renewal of the colonic epithelium. This receptor, which also serves as a coreceptor for HIV, may mediate viral infection of colonic epithelial cells. PMID- 10525045 TI - Platelet-activating factor mediates acid-induced lung injury in genetically engineered mice. AB - Adult respiratory distress syndrome (ARDS) is an acute lung injury of high mortality rate, and the molecular mechanisms underlying it are poorly understood. Acid aspiration-induced lung injury is one of the most common causes of ARDS, characterized by an increase in lung permeability, enhanced polymorphonuclear neutrophil (PMN) sequestration, and respiratory failure. Here, we investigated the role of platelet-activating factor (PAF) and the PAF receptor (PAFR) gene in a murine model of acid aspiration-induced lung injury. Overexpression of the PAFR gene in transgenic mice enhanced lung injury, pulmonary edema, and deterioration of gas exchange caused by HCl aspiration. Conversely, mice carrying a targeted disruption of the PAFR gene experienced significantly less acid-induced injury, edema, and respiratory failure. Nevertheless, the efficiency of PMN sequestration in response to acid aspiration was unaffected by differences in PAFR expression level. The current observations suggest that PAF is involved in the pathogenesis of acute lung injury caused by acid aspiration. Thus, inhibition of this pathway might provide a novel therapeutic approach to acute lung injury, for which no specific pharmaceutical agents are currently available. PMID- 10525046 TI - Adenoviral cardiotrophin-1 gene transfer protects pmn mice from progressive motor neuronopathy. AB - Cardiotrophin-1 (CT-1), an IL-6-related cytokine, causes hypertrophy of cardiac myocytes and has pleiotropic effects on various other cell types, including motoneurons. Here, we analyzed systemic CT-1 effects in progressive motor neuronopathy (pmn) mice that suffer from progressive motoneuronal degeneration, muscle paralysis, and premature death. Administration of an adenoviral CT-1 vector to newborn pmn mice leads to sustained CT-1 expression in the injected muscles and bloodstream, prolonged survival of animals, and improved motor functions. CT-1-treated pmn mice showed a significantly reduced degeneration of facial motoneuron cytons and phrenic nerve myelinated axons. The terminal innervation of skeletal muscle, grossly disturbed in untreated pmn mice, was almost completely preserved in CT-1-treated pmn mice. The remarkable neuroprotection conferred by CT-1 might become clinically relevant if CT-1 side effects, including cardiotoxicity, could be circumvented by a more targeted delivery of this cytokine to the nervous system. PMID- 10525047 TI - Effects of a low-fat, high-carbohydrate diet on VLDL-triglyceride assembly, production, and clearance. AB - Low-fat, high-carbohydrate (LF/HC) diets commonly elevate plasma triglyceride (TG) concentrations, but the kinetic mechanisms responsible for this effect remain uncertain. Subjects with low TG (normolipidemic [NL]) and those with moderately elevated TG (hypertriglyceridemic [HTG]) were studied on both a control and an LF/HC diet. We measured VLDL particle and TG transport rates, plasma nonesterified fatty acid (NEFA) flux, and sources of fatty acids used for the assembly of VLDL-TG. The LF/HC diet resulted in a 60% elevation in TG, a 37% reduction in VLDL-TG clearance, and an 18% reduction in whole-body fat oxidation, but no significant change in VLDL-apo B or VLDL-TG secretion rates. Significant elevations in fasting apo B-48 concentrations were observed on the LF/HC in HTG subjects. In both groups, fasting de novo lipogenesis was low regardless of diet. The NEFA pool contributed the great majority of fatty acids to VLDL-TG in NL subjects on both diets, whereas in HTG subjects, the contribution of NEFA was somewhat lower overall and was reduced further in individuals on the LF/HC diet. Between 13% and 29% of VLDL-TG fatty acids remained unaccounted for by the sum of de novo lipogenesis and plasma NEFA input in HTG subjects. We conclude that (a) whole-food LF/HC diets reduce VLDL-TG clearance and do not increase VLDL-TG secretion or de novo lipogenesis; (b) sources of fatty acids for assembly of VLDL TG differ between HTG and NL subjects and are further affected by diet composition; (c) the presence of chylomicron remnants in the fasting state on LF/HC diets may contribute to elevated TG levels by competing for VLDL-TG lipolysis and by providing a source of fatty acids for hepatic VLDL-TG synthesis; and (d) the assembly, production, and clearance of elevated plasma VLDL-TG in response to LF/HC diets therefore differ from those for elevated TG on higher-fat diets. PMID- 10525048 TI - Overproduction of Th2-specific chemokines in NC/Nga mice exhibiting atopic dermatitis-like lesions. AB - We have examined the expression of chemokines and their receptors in the atopic dermatitis-like (AD-like) lesions of NC/Nga mice. Such lesions develop when the mice are kept in conventional conditions, but not when they are kept isolated from specific pathogens. The thymus- and activation-regulated chemokine TARC is unexpectedly highly expressed in the basal epidermis of 14-week-old mice with lesions, whereas it is not expressed in the skin without lesions. Production of TARC by keratinocytes was confirmed by culturing murine keratinocytic cell line cells (PAM212) with TNF-alpha, IFN-gamma, or IL-1beta. Expression of another Th2 chemokine, macrophage-derived chemokine (MDC), was observed in the skin from mice kept in both conventional and pathogen-free conditions, but expression of MDC was increased severalfold in the skin with lesions. The cellular origin of MDC was identified to be dermal dendritic cells. Infiltration of the skin by IL-4 producing T cells and mast cells, and the increase of CCR4 mRNA in the skin, coincided with the development of AD lesions. These observations indicate that TARC and MDC actively participate in the pathogenesis of AD-like lesions in NC/Nga mice and that these Th2 chemokines could be novel targets for intervention therapy of AD in humans. PMID- 10525049 TI - Detoxification of hydrogen sulfide and methanethiol in the cecal mucosa. AB - Colonic bacteria liberate large quantities of the highly toxic gases hydrogen sulfide (H(2)S) and methanethiol (CH(3)SH). The colonic mucosa presumably has an efficient means of detoxifying these compounds, which is thought to occur through methylation of H(2)S to CH(3)SH and CH(3)SH to dimethylsulfide (CH(3)SCH(3)). We investigated this detoxification pathway by incubating rat cecal mucosal homogenates with gas containing H(2)S, CH(3)SH, or CH(3)SCH(3). Neither CH(3)SH nor CH(3)SCH(3) was produced during H(2)S catabolism, whereas catabolism of CH(3)SH liberated H(2)S but not CH(3)SCH(3). Thus, H(2)S and CH(3)SH are not detoxified by methylation to CH(3)SCH(3). Rather, CH(3)SH is demethylated to H(2)S, and H(2)S is converted to nonvolatile metabolites. HPLC analysis of the homogenate showed the metabolite to be primarily thiosulfate. Analysis of cecal venous blood obtained after intracecal instillation of H(2)(35)S revealed that virtually all absorbed H(2)S had been oxidized to thiosulfate. The oxidation rate of H(2)S by colonic mucosa was 10,000 times greater than the reported methylation rate. Conversion to thiosulfate appears to be the mechanism whereby the cecal mucosa protects itself from the injurious effects of H(2)S and CH(3)SH, and defects in this detoxification possibly could play a role in colonic diseases such as ulcerative colitis. PMID- 10525050 TI - Mutations in Igalpha (CD79a) result in a complete block in B-cell development. AB - Mutations in Btk, mu heavy chain, or the surrogate light chain account for 85-90% of patients with early onset hypogammaglobulinemia and absent B cells. The nature of the defect in the remaining patients is unknown. We screened 25 such patients for mutations in genes encoding components of the pre-B-cell receptor (pre-BCR) complex. A 2-year-old girl was found to have a homozygous splice defect in Igalpha, a transmembrane protein that forms part of the Igalpha/Igbeta signal transduction module of the pre-BCR. Studies in mice suggest that the Igbeta component of the pre-BCR influences V-DJ rearrangement before cell-surface expression of mu heavy chain. To determine whether Igalpha plays a similar role, we compared B-cell development in an Igalpha-deficient patient with that seen in a mu heavy chain-deficient patient. By immunofluorescence, both patients had a complete block in B-cell development at the pro-B to pre-B transition; both patients also had an equivalent number and diversity of rearranged V-DJ sequences. These results indicate that mutations in Igalpha can be a cause of agammaglobulinemia. Furthermore, they suggest that Igalpha does not play a critical role in B-cell development until it is expressed, along with mu heavy chain, as part of the pre-BCR. PMID- 10525051 TI - CREB-independent regulation by CBP is a novel mechanism of human growth hormone gene expression. AB - Hypothalamic growth hormone-releasing hormone (GHRH) stimulates growth hormone (GH) gene expression in anterior pituitary somatotrophs by binding to the GHRH receptor, a G-protein-coupled transmembrane receptor, and by mediating a cAMP mediated protein kinase A (PKA) signal-transduction pathway. Two nonclassical cAMP-response element motifs (CGTCA) are located at nucleotides -187/-183 (distal cAMP-response element; dCRE) and -99/-95 (proximal cAMP-response element; pCRE) of the human GH promoter and are required for cAMP responsiveness, along with the pituitary-specific transcription factor Pit-1 (official nomenclature, POU1F1). Although a role for cAMP-response element binding protein (CREB) in GH stimulation by PKA has been suggested, it is unclear how the effect may be mediated. CREB binding protein (CBP) is a nuclear cofactor named for its ability to bind CREB. However, CBP also binds other nuclear proteins. We determined that CBP interacts with Pit-1 and is a cofactor for Pit-1-dependent activation of the human GH promoter. This pathway appears to be independent of CREB, with CPB being the likely target of phosphorylation by PKA. PMID- 10525052 TI - Renal cytochrome P450 omega-hydroxylase and epoxygenase activity are differentially modified by nitric oxide and sodium chloride. AB - Renal function is perturbed by inhibition of nitric oxide synthase (NOS). To probe the basis of this effect, we characterized the effects of nitric oxide (NO), a known suppressor of cytochrome P450 (CYP) enzymes, on metabolism of arachidonic acid (AA), the expression of omega-hydroxylase, and the efflux of 20 hydroxyeicosatetraenoic acid (20-HETE) from the isolated kidney. The capacity to convert [(14)C]AA to HETEs and epoxides (EETs) was greater in cortical microsomes than in medullary microsomes. Sodium nitroprusside (10-100 microM), an NO donor, inhibited renal microsomal conversion of [(14)C]AA to HETEs and EETs in a dose dependent manner. 8-bromo cGMP (100 microM), the cell-permeable analogue of cGMP, did not affect conversion of [(14)C]AA. Inhibition of NOS with N(omega)-nitro-L arginine-methyl ester (L-NAME) significantly increased conversion of [(14)C]AA to HETE and greatly increased the expression of omega-hydroxylase protein, but this treatment had only a modest effect on epoxygenase activity. L-NAME induced a 4 fold increase in renal efflux of 20-HETE, as did L-nitroarginine. Oral treatment with 2% sodium chloride (NaCl) for 7 days increased renal epoxygenase activity, both in the cortex and the medulla. In contrast, cortical omega-hydroxylase activity was reduced by treatment with 2% NaCl. Coadministration of L-NAME and 2% NaCl decreased conversion of [(14)C]AA to HETEs without affecting epoxygenase activity. Thus, inhibition of NOS increased omega-hydroxylase activity, CYP4A expression, and renal efflux of 20-HETE, whereas 2% NaCl stimulated epoxygenase activity. PMID- 10525053 TI - Killing of Streptococcus pneumoniae by capsular polysaccharide-specific polymeric IgA, complement, and phagocytes. AB - The role of IgA in the control of invasive mucosal pathogens such as Streptococcus pneumoniae is poorly understood. We demonstrate that human pneumococcal capsular polysaccharide-specific IgA initiated dose-dependent killing of S. pneumoniae with complement and phagocytes. The majority of specific IgA in serum was of the polymeric form (pIgA), and the efficiency of pIgA initiated killing exceeded that of monomeric IgA-initiated killing. In the absence of complement, specific IgA induced minimal bacterial adherence, uptake, and killing. Killing of S. pneumoniae by resting phagocytes with immune IgA required complement, predominantly via the C2-independent alternative pathway, which requires factor B, but not calcium. Both S. pneumoniae-bound IgA and complement were involved, as demonstrated by a 50% decrease in killing with blocking of Fcalpha receptor (CD89) and CR1/CR3 (CD35/CD11b). However, IgA mediated killing by phagocytes could be reproduced in the absence of opsonic complement by pre-activating phagocytes with the inflammatory products C5a and TNF-alpha. Thus, S. pneumoniae capsule-specific IgA may show distinct roles in effecting clearance of S. pneumoniae in the presence or absence of inflammation. These data suggest mechanisms whereby pIgA may serve to control pneumococcal infections locally and upon the pathogen's entry into the bloodstream. PMID- 10525054 TI - Temporal adjustment of the juxtaglomerular apparatus during sustained inhibition of proximal reabsorption. AB - Tubuloglomerular feedback (TGF) stabilizes nephron function by causing changes in single-nephron GFR (SNGFR) to compensate for changes in late proximal flow (VLP). TGF responds within seconds and reacts over a narrow range of VLP that surrounds normal VLP. To accommodate sustained increases in VLP, TGF must reset around the new flow. We studied TGF resetting by inhibiting proximal reabsorption with benzolamide (BNZ; administered repeatedly over a 24-hour period) in Wistar Froemter rats. BNZ acutely activates TGF, thereby reducing SNGFR. Micropuncture was performed 6-10 hours after the fourth BNZ dose, when diuresis had subsided. BNZ caused glomerular hyperfiltration, which was prevented with inhibitors of macula densa nitric oxide synthase (NOS). Because of hyperfiltration, BNZ increased VLP and distal flow, but did not affect the basal TGF stimulus (early distal salt concentration). BNZ slightly blunted normalized maximum TGF response and the basal state of TGF activation. BNZ sensitized SNGFR to reduction by S methyl-thiocitrulline (SMTC) and caused the maximum TGF response to be strengthened by SMTC. Sensitization to type I NOS (NOS-I) blockers correlated with increased macula densa NOS-I immunoreactivity. Tubular transport measurements confirmed that BNZ affected TGF within the juxtaglomerular apparatus. During reduced proximal reabsorption, TGF resets to accommodate increased flow and SNGFR through a mechanism involving macula densa NOS. PMID- 10525055 TI - The Tangier disease gene product ABC1 controls the cellular apolipoprotein mediated lipid removal pathway. AB - The ABC1 transporter was identified as the defect in Tangier disease by a combined strategy of gene expression microarray analysis, genetic mapping, and biochemical studies. Patients with Tangier disease have a defect in cellular cholesterol removal, which results in near zero plasma levels of HDL and in massive tissue deposition of cholesteryl esters. Blocking the expression or activity of ABC1 reduces apolipoprotein-mediated lipid efflux from cultured cells, and increasing expression of ABC1 enhances it. ABC1 expression is induced by cholesterol loading and cAMP treatment and is reduced upon subsequent cholesterol removal by apolipoproteins. The protein is incorporated into the plasma membrane in proportion to its level of expression. Different mutations were detected in the ABC1 gene of 3 unrelated patients. Thus, ABC1 has the properties of a key protein in the cellular lipid removal pathway, as emphasized by the consequences of its defect in patients with Tangier disease. PMID- 10525056 TI - Effect of molecular charge on intestinal epithelial drug transport: pH-dependent transport of cationic drugs. AB - The aim of this study was to investigate the effect of ionization on drug transport across the intestinal epithelium in order to include this effect in structure-absorption relationships. The pH-dependent permeation of one rapidly (alfentanil) and one slowly (cimetidine) transported basic model drug across Caco 2 cell monolayers was investigated. Both drugs had pK(a)values in the physiological pH range. The permeability coefficients (P(c)) of the model drugs were obtained at varying apical buffer pHs, thus varying the degree of drug ionization (from 5 to 95%). The relationship between P(c) and the fraction of the drug in un-ionized form (f(u)) was analyzed to delineate the permeability coefficients of the un-ionized (P(c,u)) and ionized (P(c,i)) forms of the drugs. Theoretical estimates of the pK(a) values were also calculated from ionization energies for each model compound. For both drugs, a linear increase in P(c) was observed with increasing f(u). Transport of the un-ionized form was 150- and 30 fold more rapid than transport of the ionized form for alfentanil and cimetidine, respectively. However, when f(u) <0.1, the contribution of the ionized form was significant. Because f(u) is <0.1 over the entire physiological pH range for a large number of drugs, these results will have implications on predictions of in vivo intestinal drug absorption both from in vitro studies in cell cultures and from computed structural properties of drug molecules. PMID- 10525057 TI - Pharmacogenetic evidence for the involvement of 5-hydroxytryptamine (Serotonin) 1B receptors in the mediation of morphine antinociceptive sensitivity. AB - Morphine antinociception has been shown to be influenced significantly by genetic factors, now beginning to be identified in mice. A recent quantitative trait locus analysis revealed a significant statistical association between morphine antinociceptive magnitude and a region of mouse chromosome 9. This region contains the Htr1b gene, which encodes the 5-hydroxytryptamine (serotonin)-1B (5 HT(1B)) receptor subtype. To investigate the possibility that Htr1b represents the quantitative trait locus, C57BL/6 and DBA/2 inbred strains, the progenitors of the original quantitative trait locus mapping populations, were administered a novel 5-HT(1B) receptor antagonist (GR127935) concomitant with morphine. These mice are known to differ in morphine antinociceptive sensitivity on thermal pain assays (DBA/2 high; C57BL/6 low). GR127935 caused a dose-dependent antagonism (both reversal and prevention) of morphine antinociception in DBA/2 mice but had no effect in C57BL/6 mice. However, a 5-hydroxytryptamine-1A subtype (5-HT(1A)) receptor agonist, 8-hydroxydipropylaminotetralin, reversed morphine antinociception equally in the two strains. DBA/2 mice also exhibited significantly greater antinociception than did C57BL/6 mice from the administration of a 5-HT(1B) agonist, CGS12066. These data collectively support a role for 5-HT(1B) receptors in the mediation of morphine antinociception and support the contention that polymorphisms in the Htr1b gene may underlie individual differences in morphine sensitivity. PMID- 10525058 TI - Biological effects of 1alpha-hydroxy- and 1beta-(hydroxymethyl)-vitamin D compounds relevant for potential colorectal cancer therapy. AB - 1alpha,25-dihydroxyvitamin D(3) and two synthetic analogs, 1alpha, 25-dihydroxy 16-ene-23-yne-vitamin D(3) (Ro 23-7553) and 1alpha, 25-dihydroxy-16-ene-24-oxo vitamin D(3) (JK-1624-3), were tested for their ability to specifically inhibit growth and promote differentiation of human colon cancer cells in comparison with a series of 1beta-(hydroxymethyl) congeners of the natural hormone, such as 1beta (hydroxymethyl)-3alpha,25(OH)(2)-16-ene,24-oxo-vitamin D(3) (JK-1624-2), 1beta (hydroxymethyl)-3alpha, 25-dihydroxy-16-ene-26,27-dihomo vitamin D(3) (JK-1626 2), and 1beta-(hydroxymethyl)-3alpha,25-dihydroxy-22,24-diene-26,27- dihomo vitamin D(3) (MCW-EE). Western blot analysis revealed that reduction of cyclin D1 levels is a key mechanism by which the vitamin D compounds under investigation inhibit Caco-2 tumor cell growth. Both the 1alpha-hydroxy- as well as the 1beta hydroxymethyl-type vitamin D compounds, which exhibit only low affinity for the vitamin D receptor, significantly reduced [(3)H]thymidine DNA labeling in confluent Caco-2 cell cultures. This suggests that high-affinity binding to the vitamin D receptor is not an absolute prerequisite for genomic action on tumor cell growth. Hybrid analogs JK-1624-2 and MCW-EE, although antimitotically active, were rather ineffective in promoting phenotypic differentiation of human colon cancer cells. However, because both compounds also do not promote osteoclast differentiation from hematopoetic bone marrow cells, they still could be used as antimitotic agents in cancer therapy, even at dose levels that, with other analogs, could cause hypercalcemia. PMID- 10525059 TI - LLC-PK(1) cells stably expressing the human norepinephrine transporter: A functional model of carrier-mediated norepinephrine release in protracted myocardial ischemia. AB - In myocardial ischemia, adrenergic terminals undergo ATP depletion, hypoxia, and intracellular pH reduction, causing the accumulation of axoplasmic norepinephrine (NE) and intracellular Na(+) [via the Na(+)-H(+) exchanger (NHE)]. This forces the reversal of the Na(+)- and Cl(-)-dependent NE transporter (NET), triggering massive carrier-mediated NE release and, thus, arrhythmias. We have now developed a cellular model of carrier-mediated NE release using an LLC-PK(1) cell line stably transfected with human NET cDNA (LLC-NET). LLC-NET cells transported [(3)H]NE and [(3)H]N-methyl-4-phenylpyridinium ([(3)H]MPP(+)) in an inward direction. This uptake was abolished by the NET inhibitors desipramine (100 nM) and mazindol (300 nM) and by extracellular Na(+) removal. Na(+)-gradient reversal induced an efflux of (3)H-substrate from preloaded LLC-NET cells. Desipramine and mazindol blocked this efflux. Because of its greater intracellular stability and higher sensitivity to Na(+)-gradient reversal, [(3)H]MPP(+) proved preferable to [(3)H]NE as an NET substrate; therefore, only [(3)H]MPP(+) was used for subsequent studies. The K(+)/H(+) ionophore nigericin (10 microM) evoked a large efflux of [(3)H]MPP(+). This efflux was potentiated by the Na(+),K(+)-ATPase inhibitor ouabain (100 microM), was sensitive to desipramine, and was blocked by the NHE inhibitor 5-(N-ethyl-N-isopropyl)-amiloride (EIPA; 10 microM). In contrast, EIPA failed to inhibit the [(3)H]MPP(+) efflux elicited by the Na(+) ionophore gramicidin (10 microM). Furthermore, [(3)H]MPP(+) efflux induced by the NHE-stimulant proprionate (25 mM) was negatively modulated by imidazoline receptor activation. Our findings suggest that LLC-NET cells are a sensitive model for studying transductional processes of carrier-mediated NE release associated with myocardial ischemia. PMID- 10525060 TI - Selectivities of dihydropyridine derivatives in blocking Ca(2+) channel subtypes expressed in Xenopus oocytes. AB - Some dihydropyridines (DHPs), such as amlodipine and cilnidipine, have been shown to block not only L-type but also N-type Ca(2+) channels; therefore, DHPs are no longer considered as L-type-specific Ca(2+) channel blockers. However, selectivity of DHPs for Ca(2+) channel subtypes including N-, P/Q-, and R-types are poorly understood. To address this issue at the molecular level, blocking effects of 10 DHPs (nifedipine, nilvadipine, barnidipine, nimodipine, nitrendipine, amlodipine, nicardipine, benidipine, felodipine, and cilnidipine) on four subtypes of Ca(2+) channels (L-, N-, P/Q-, and R-types) were investigated in the Xenopus oocyte expression system with the use of the two-microelectrode voltage-clamp technique. L-type Ca(2+) channels expressed as alpha(1C)alpha(2)beta(1a) combination were profoundly blocked by all DHPs examined, whereas blocking actions of these DHPs on R-type (alpha(1E)alpha(2)beta(1a)) channels were equally weak. In contrast, 5 of the 10 DHPs (amlodipine, benidipine, cilnidipine, nicardipine, and barnidipine) significantly blocked N-type (alpha(1B)alpha(2)beta(1a)) and P/Q-type (alpha(1A)alpha(2)beta(1a)) Ca(2+) channels. These selectivities of DHPs in blocking Ca(2+) channel subtypes would provide useful pharmacological and clinical information on the mode of action of the drugs including side effects and adverse effects. PMID- 10525061 TI - ATP-sensitive potassium channel blocker HMR 1883 reduces mortality and ischemia associated electrocardiographic changes in pigs with coronary occlusion. AB - ATP-sensitive potassium (K(ATP)) channels are activated during myocardial ischemia. The ensuing potassium efflux leads to a shortening of the action potential duration and depolarization of the membrane by accumulation of extracellular potassium favoring the development of reentrant arrhythmias, including ventricular fibrillation. The sulfonylthiourea HMR 1883 was designed as a cardioselective blocker of myocardial K(ATP) channels for the prevention of arrhythmic sudden death in patients with ischemic heart disease. We investigated the effect of HMR 1883 on sudden cardiac arrhythmic death and electrocardiography (ECG) changes induced by 20 min of left anterior descending coronary artery occlusion in pentobarbital-anesthetized pigs. HMR 1883 (3 mg/kg i.v.) protected pigs from arrhythmic death (91% survival rate versus 33% in control animals; n = 12; p<.05). Ischemic areas were of a similar size. The compound had no effect on hemodynamics and ECG, including Q-T interval, under baseline conditions and no effect on hemodynamics during occlusion. In control animals, left anterior descending coronary artery occlusion lead to a prompt and significant depression of the S-T segment (-0.35 mV) and a prolongation of the Q-J time (+46 ms), the former reflecting heterogeneity in the plateau phase of the action potentials and the latter reflecting irregular impulse propagation and delayed ventricular activation. Both ischemic ECG changes were significantly attenuated by HMR 1883 (S-T segment, -0.14 mV; Q-J time, +15 ms), indicating the importance of K(ATP) channels in the genesis of these changes. In conclusion, the K(ATP) channel blocker HMR 1883, which had no effect on hemodynamics and ECG under baseline conditions, reduced the extent of ischemic ECG changes and sudden death due to ventricular fibrillation during coronary occlusion. PMID- 10525062 TI - Effects of regulators of G protein-signaling proteins on the functional response of the mu-opioid receptor in a melanophore-based assay. AB - The goal of the present study was to investigate a possible role for regulators of G protein-signaling (RGS) proteins in opioid receptor (OR) desensitization using cultured Xenopus laevis dermal melanophores. Morphine-induced pigment aggregation in a melanophore cell line stably expressing the murine mu OR (muOR) was quantified over time. Responses of the muOR (a G(i)-linked receptor) exhibited a time-dependent desensitization, which varied with the concentration of morphine used. In contrast, much less desensitization was observed in response to melatonin, effects mediated through the cells' endogenous melatonin receptor (which is also G(i)-linked). To further study OR desensitization, melanophores lacking a muOR were transiently transfected with plasmids encoding the muOR alone or in combination with plasmids encoding one of several RGS subtypes (RGS1, RGS2, RGS3, or RGS4). Overexpression of RGS2, but not the other RGS subtypes, produced a rightward shift in the morphine concentration-response curve. RGS protein overexpression also decreased the magnitude of morphine-induced responses. Finally, the effect of a mutant form of Galpha(i1), which is insensitive to RGS action, was investigated with respect to its ability to alter the response of the muOR to morphine. Expression of the mutant Galpha(i1) prolonged morphine-induced pigment aggregation and produced leftward shifts in concentration-response curves, compared with expression of wild-type Galpha(i1). These results demonstrate that specific RGS proteins can dampen signals initiated by agonist activation of the muOR, and support a possible role for RGS proteins in OR desensitization. PMID- 10525063 TI - Influence of exogenous thiols on inorganic mercury-induced injury in renal proximal and distal tubular cells from normal and uninephrectomized rats. AB - Inorganic mercury (Hg(2+)) induced time- and concentration-dependent cellular injury in freshly isolated proximal tubular (PT) and distal tubular (DT) cells from normal (control) rats or uninephrectomized (NPX) rats. PT cells from NPX rats were more susceptible than PT cells from control rats, and DT cells were slightly more susceptible than PT cells to cellular injury induced by Hg(2+) (not bound to a thiol). Preloading cells with glutathione increased Hg(2+)-induced cellular injury in PT cells from control rats. However, coincubation of PT or DT cells from control or NPX rats with Hg(2+) and glutathione (1:4) provided significant protection relative to incubations with Hg(2+) alone. No support was obtained for a role for gamma-glutamyltransferase in glutathione-dependent protection. However, the organic anion carrier does appear to play a role in accumulation and toxicity of mercuric conjugates of cysteine in PT cells from control, but not NPX, rats. Coincubation with Hg(2+) and cysteine (1:4) had little effect on, or slightly enhanced, Hg(2+)-induced cellular injury at low concentrations of Hg(2+) in all cells studied. Coincubation with Hg(2+) and albumin (1:4) markedly protected PT and DT cells from control and NPX rats at all concentrations except the highest concentration of Hg(2+) in DT cells from NPX rats. 2,3-Dimercapto-1-propanesulfonic acid protected cells both when preloaded or added simultaneously with Hg(2+). Thus, renal cells from NPX rats are more susceptible to Hg(2+)-induced injury, PT and DT cells respond differently to exposure to Hg(2+), and thiols can significantly modulate the toxic response to Hg(2+). PMID- 10525064 TI - Agonist-induced sensitization of beta-adrenoceptor signaling in neonatal rat heart: expression and catalytic activity of adenylyl cyclase. AB - Agonist stimulation of neonatal cardiac beta-adrenoceptors produces heterologous sensitization of adenylyl cyclase (AC) signaling, rather than desensitization, as seen in adults. We examined the ontogenetic patterns of AC expression and activity, and evaluated isoproterenol effects on this pattern. [(3)H]Forskolin binding showed an increase in AC concentration across the period (birth to 25 days of age) in which agonist-induced sensitization is replaced by desensitization; binding affinity also increased, suggesting a shift in conformation and/or isoform. Indeed, catalytic properties of AC changed substantially with development, as evaluated by AC responses to forskolin versus Mn(2+). In contrast, there were only minor changes in the levels of mRNAs encoding the two major isoforms. Neonates given repeated isoproterenol treatment showed an enhancement of [(3)H]forskolin binding B(max) and a precocious shift to the mature affinity state and corresponding catalytic properties. Although isoproterenol caused significant increases in AC mRNAs, the effects were small and showed no isoform preference. Thus, a primary mode for ontogenetic increases in cardiac cellular responsiveness to adrenergic stimulation is the increase in AC activity attendant upon an absolute increase in the membrane concentration of AC molecules, along with changes in the catalytic properties of AC. The lack of correlation between mRNA and AC protein suggests that the primary regulatory events are post-transcriptional. The induction of AC by beta-adrenoceptor stimulation in the fetus and neonate accounts for heterologous, agonist-induced sensitization, a phenomenon that preserves cellular responses during the period of the perinatal transition. PMID- 10525065 TI - 5-Oxo-6,8,11,14-eicosatetraenoic acid stimulates isotonic volume reduction of guinea pig jejunal crypt epithelial cells. AB - 5-Oxo-6,8,11,14-eicosatetraenoic acid (5-oxo-ETE) is a recently discovered arachidonate metabolite that is a potent activator of eosinophils and neutrophils and may be an important mediator of inflammation. The objective of the present investigation was to determine whether 5-oxo-ETE affects the isotonic volume of Cl(-) secretory intestinal crypt epithelial cells. 5-Oxo-ETE caused rapid shrinkage of guinea pig jejunal crypt epithelial cells to a reduced but stable volume, which was measured electronically. This effect was prevented by Cl(-) and K(+) channel blockers and inhibitors of protein kinase C. 5-Oxo-ETE (EC(50) = 20 pM) was more potent than any of the other agonists tested, including its precursor, 5-hydroxy-6,8,11,14-eicosatetraenoic acid (EC(50) = 5 nM); leukotriene D(4) (EC(50) = 1 nM); vasoactive intestinal peptide (EC(50) = 200 pM); and bradykinin (EC(50) = 50 nM). Leukotriene B(4) had no effect on crypt cell volume. In contrast to its effects on crypt cells, 5-oxo-ETE had no effect on the volume of jejunal villus cells. These results indicate that 5-oxo-ETE induces an isotonic volume reduction in intestinal crypt epithelial cells that appears to be dependent on Cl(-) secretion and activation of protein kinase C. PMID- 10525066 TI - Inhibitors of chymase as mast cell-stabilizing agents: contribution of chymase in the activation of human mast cells. AB - There has long been evidence that inhibitors of chymotryptic proteinases can inhibit the degranulation of rodent mast cells, but their actions on human mast cells and the contribution of mast cell chymase itself have received little attention. We investigated the ability of the selective chymase inhibitor Z-Ile Glu-Pro-Phe-CO(2)Me and other proteinase inhibitors to inhibit chymase and cathepsin G activity, and we examined their potential to modulate the responsiveness of mast cells dispersed from human skin, lung, and tonsil tissues. IgE-dependent histamine release from skin mast cells was inhibited by up to about 80% after preincubation with Z-Ile-Glu-Pro-Phe- CO(2)Me (up to 0.1 microM), 70% with chymostatin (17 microM), and 60% with soybean trypsin inhibitor (0.5 microM). The mast cell-stabilizing properties of chymase inhibitors appeared to be greater for skin mast cells than for those from lung, whereas tonsil mast cells were relatively unresponsive. There were marked differences in the time course of responses to inhibitors, and the effect was dependent on the stimulus, with calcium ionophore-induced histamine release being unaffected. Incubation of dispersed skin, lung, or tonsil cells for up to 45 min with purified chymase failed to induce histamine release, although preincubation of cells with chymase was able to suppress IgE-dependent activation. Chymase could thus contribute to mast cell degranulation and after secretion could provide a feedback mechanism to limit this process. Nevertheless, inhibitors of chymase can be potent mast cell stabilizers, particularly in the skin. PMID- 10525067 TI - Estrogen alters relative contributions of nitric oxide and cyclooxygenase products to endothelium-dependent vasodilation. AB - The purpose of this study was to determine the effects of in vivo estrogen manipulations on mechanisms of endothelium-dependent vasodilation. Ovary-intact, ovariectomized (OVX), or OVX with estrogen replacement (OVX + E(2)) female Sprague-Dawley rats were studied (n = 8). Mesenteric arteries (approximately 300 microm) were isolated, cannulated, and pressurized to 60 mm Hg in an arteriograph containing bicarbonate buffer and vessel diameter was monitored. Concentration response curves to the endothelium-dependent histamine H(1) agonist 2 thiazolylethylamine (2-TEA; 1 nM-100 microM) and to acetylcholine (1 nM-10 microM) were performed in preconstricted arteries. The effect of Nomega-nitro-L arginine (LNA; 100 microM) or LNA + indomethacin (INDO) (10 microM) on agonist induced vasodilation was determined. There was no difference between treatment groups in the sensitivity of mesenteric arteries to 2-TEA or acetylcholine. LNA produced a significant decrease in sensitivity to 2-TEA in arteries from ovary intact and OVX + E(2) rats but not in those from OVX rats. The addition of INDO produced a small additional decrease in sensitivity to 2-TEA in arteries from ovary-intact rats, a significant decrease in OVX, and no shift in OVX + E(2). LNA + INDO produced a similar degree of inhibition of the 2-TEA response in the three treatment groups. In contrast, when acetylcholine was used, the decrease in sensitivity produced by LNA or LNA + INDO was similar in the three rat groups. We conclude that estrogen increases the nitric oxide component of endothelium dependent dilation and decreases the cyclooxygenase component. These effects of estrogen appear to be agonist-specific. Our findings suggest that estrogen modulates cross talk between the nitric oxide synthase and cyclooxygenase pathways of vasodilation. PMID- 10525068 TI - Role of adenosine and N-methyl-D-aspartate receptors in mediating haloperidol induced gene expression and catalepsy. AB - Acute blockade of dopamine D(2) receptors by the typical antipsychotic drug haloperidol leads to alterations in neuronal gene expression and behavior. In the dorsolateral striatum, the levels of mRNA for the immediate-early gene c-fos and the neuropeptide gene neurotensin/neuromedin N (NT/N) are significantly increased by haloperidol. An acute behavioral response to haloperidol is catalepsy, considered to be a rodent correlate of some of the immediate extrapyramidal motor side effects seen in humans. Several lines of evidence suggest a link between neurotensin induction in the dorsolateral striatum and catalepsy. We hypothesize that both striatal gene induction and catalepsy elicited by haloperidol arise from the combined effect of excitatory adenosinergic and glutamatergic inputs acting at adenosine A(2A) and N-methyl-D-aspartate (NMDA) receptors, respectively. In agreement with our previous reports, adenosine antagonists reduced haloperidol-induced c-fos and neurotensin gene expression as well as catalepsy. In agreement with other reports, the noncompetitive NMDA receptor antagonist MK-801 also reduced gene expression and catalepsy in response to haloperidol. The competitive NMDA receptor antagonist LY235959 decreased haloperidol-induced catalepsy. We show here that blocking both A(2A) and NMDA receptors simultaneously in conjunction with haloperidol resulted in a combined effect on gene expression and behavior that was greater than that for block of either receptor alone. Both c-fos and NT/N mRNA levels were reduced, and catalepsy was completely abolished. These results indicate that the haloperidol induced increases in c-fos and NT gene expression in the dorsolateral striatum and catalepsy are driven largely by adenosine and glutamatergic inputs acting at A(2A) and NMDA receptors. PMID- 10525069 TI - Effects of Delphinium alkaloids on neuromuscular transmission. AB - The Delphinium alkaloids methyllycaconitine (MLA), nudicauline, 14 deacetylnudicauline (14-DN), barbinine, and deltaline were investigated for their effects on neuromuscular transmission in lizards. The substituent at C14 provides the only structural difference among the alkaloids MLA, nudicauline, 14-DN, and barbinine. Deltaline lacks the N-(methylsuccinyl)anthranilic acid at C18 common to the other four alkaloids. Each alkaloid reversibly reduced extracellularly recorded compound muscle action potential (CMAP) amplitudes in a concentration dependent manner. The IC(50) values for CMAP blockade were between 0.32 and 13.2 microM for the N-(methylsuccinimido)anthranoyllycacotonine-type alkaloids and varied with the C14 moiety; the IC(50) value for deltaline was 156 microM. The slopes of the concentration-response curves for CMAP blockade were similar for each alkaloid except barbinine, whose shallower curve suggested alternative or additional mechanisms of action. Each alkaloid reversibly reduced intracellularly recorded spontaneous, miniature end-plate potential (MEPP) amplitudes. Alkaloid concentrations producing similar reductions in MEPP amplitude were 0.05 microM for 14-DN, 0.10 microM for MLA, 0.50 microM for barbinine, and 20 microM for deltaline. Only barbinine altered the time constant for MEPP decay, further suggesting additional or alternative effects for this alkaloid. MLA and 14-DN blocked muscle contractions induced by exogenously added acetylcholine. All five alkaloids are likely nicotinic receptor antagonists that reduce synaptic efficacy and block neuromuscular transmission. The substituent at C14 determines the potency and possibly the mechanism of nicotinic acetylcholine receptor blockade for MLA, nudicauline, 14-DN, and barbinine at neuromuscular synapses. The lower potency of deltaline indicates that the N-(methylsuccinyl)anthranilic acid at C18 affects alkaloid interactions with nicotinic acetylcholine receptors at neuromuscular junctions. PMID- 10525071 TI - Chlorotrifluoroethylcysteine interaction with rabbit proximal tubule cell basolateral membrane organic anion transport and apical membrane amino acid transport. AB - The interaction of the cysteine conjugate S-(1-chloro-1,2,2, -trifluoroethyl)-L cysteine (CTFC) with organic anion and amino acid transport in the basolateral and apical membranes was examined with rabbit renal proximal tubule suspensions and primary cultures of rabbit renal proximal tubule cells. The apparent K(i) for CTFC inhibition of the 1-min uptake of [(3)H]p-aminohippurate in tubule suspensions was 105+/-3 microM and suggests that CTFC interacts with basolateral organic anion transport. Also, the addition of 1 mM CTFC decreased the secretion and intracellular accumulation of fluorescein by approximately 70 to 75%. The addition of 1 mM CTFC to the apical compartment decreased the reabsorption and intracellular accumulation of the amino acid [(3)H]phenylalanine by approximately 60 to 70%. Similar to CTFC, saturating concentrations of the organic anion [(3)H]p-aminohippurate and the amino acid phenylalanine reduced by approximately 75% fluorescein secretion and [(3)H]phenylalanine reabsorption, respectively, by approximately 60 to 70%. Thus, the cysteine conjugate CTFC appears to be a potent inhibitor of basolateral organic anion and apical amino acid transepithelial transport. In contrast to its effects on apical phenylalanine uptake, CTFC had no effect on the basal uptake of [(3)H]phenylalanine by primary cultures. The presence of CTFC in the external bath did trans-stimulate the efflux of fluorescein and [(3)H]phenylalanine across the basal and apical membrane in tubule suspensions or primary cultures, respectively, grown on plastic. Collectively, these data demonstrate that CTFC interacts with, and is transported by, two anatomically and functionally distinct transporters, the basolateral organic anion and apical neutral amino acid pathways, in the rabbit renal proximal tubule cell. PMID- 10525070 TI - Pharmacokinetic-pharmacodynamic modeling of tolerance to the prolactin-secreting effect of chlorprothixene after different modes of drug administration. AB - The objective of this study was the construction of a pharmacokinetic pharmacodynamic model to describe the effects of chlorprothixene on prolactin secretion and the time-dependent alterations in the concentration-effect relationship due to tolerance development. Prolactin and chlorprothixene serum concentrations were determined in eight healthy men for up to 72 h after the intravenous and oral administration of chlorprothixene. An integrated pharmacokinetic model and a physiological indirect pharmacodynamic/tolerance model were applied to describe the prolactin-secreting effect of chlorprothixene. A three-compartment model served as pharmacokinetic model. The pharmacodynamic and tolerance model accounted for the baseline effect, the effect induced by the drug, and the regulatory mechanism that opposes the effect of the drug. This model adequately characterized the prolactin response after intravenous and oral drug administration of each individual by the sensitivity (dissociation constant), the efficacy (maximal prolactin secretion rate), the extent, and the rate of tolerance development. We speculate that this approach improves the quality of neuroendocrine challenge tests to determine the subject's sensitivity to drugs and the time course of adaptation. PMID- 10525072 TI - Bepridil blunts the shortening of action potential duration caused by metabolic inhibition via blockade of ATP-sensitive K(+) channels and Na(+)-activated K(+) channels. AB - The effects of bepridil, a potent antiarrhythmic drug, on the activity of ATP sensitive K(+) (K(ATP)) channels and Na(+)-activated K(+) (K(Na)) channels were examined in isolated patches from guinea pig ventricular myocytes. In inside-out membrane patches, K(ATP) channel currents were recorded with 140 mM [K(+)](i) and 140 mM [K(+)](o) solutions, and K(Na) channel currents were recorded by increasing [Na(+)](i) to 100 mM with 40 mM [K(+)](i), respectively. Bepridil (1 100 microM) inhibited the K(ATP) channel current in a concentration-dependent manner. The IC(50) value of bepridil was estimated to be 10.5 microM for outward K(ATP) channel currents (holding potential, +60 mV) and 6.6 microM for inward K(ATP) channel currents (holding potential, -60 mV). Bepridil (0.1-30 microM) also inhibited K(Na) channel currents measured at the holding potential of -60 mV, in a concentration-dependent manner with an IC(50) value of 2.2 microM. In coronary-perfused guinea pig right ventricular preparations, the metabolic inhibition (MI) achieved with the application of 0.1 microM carbonyl cyanide p (trifluoromethoxy)phenylhydrazone shortened the action potential duration (APD) in a time-dependent manner. When bepridil (10 microM) was applied 5 min after the introduction of MI, the APD shortening was significantly blunted. The concomitant application of a K(ATP) channel antagonist (glibenclamide, 1 microM) and a K(Na) channel antagonist (R56865, 10 microM) could mimic the effect of bepridil and attenuated the shortening otherwise produced by MI. These results suggest that bepridil inhibits both K(ATP) channels and K(Na) channels and blunts the shortening of APD during MI. These effects of bepridil may partly account for the alleged antiarrhythmic action of this drug during ischemia. PMID- 10525073 TI - Effect of poststroke captopril treatment on mortality associated with hemorrhagic stroke in stroke-prone rats. AB - We tested the ability of captopril treatment (50 mg/kg/day p.o.), initiated 2 weeks before stroke or up to 5 days after stroke, to alter the onset of stroke and death after stroke in Kyoto Wistar stroke-prone spontaneously hypertensive rats (SHRsp). The benefits of blood pressure and aldosterone suppression during captopril treatment were assessed. SHRsp developed a 100% mortality rate with intracerebral hemorrhage by 16 weeks of age. Captopril treatment, started 2 weeks before or at the initiation of stroke, suppressed plasma aldosterone and equally prevented mortality to a mean age of >27 weeks. Treatment started 5 days after stroke extended the mean lifespan to >23 weeks. The re-elevation of plasma aldosterone (via osmotic pumps to levels in untreated SHRsp) during captopril treatment, before stroke, allowed stroke to develop. The initiation of the latter manipulation in pre- or poststroke captopril-treated SHRsp at a latter age (23 weeks) didn't alter the lifespan of SHRsp (death occurred at about 28 weeks). The antistroke effects of captopril treatment occurred without an antihypertensive effect, weren't altered by enhancing hypertension during treatment (with dexamethasone), and couldn't be duplicated by antihypertensive treatment with hydralazine. Spironolactone treatment didn't duplicate the effects of captopril. The suppression of plasma aldosterone may retard the onset of stroke in SHRsp during captopril treatment but likely other factors prolong life in pre- and poststroke SHRsp receiving long-term captopril treatment. The observation that spironolactone treatment couldn't duplicate the effects of captopril suggests that aldosterone may facilitate stroke through nongenomic receptor mechanisms. PMID- 10525074 TI - Sex differences in the pentylenetetrazol-like stimulus induced by ethanol withdrawal. AB - This study investigated sex differences in responding to the pentylenetetrazol (PTZ, a gamma-aminobutyric acid A antagonist) discriminative stimulus and to substitution to PTZ during ethanol withdrawal. The PTZ stimulus has served as an anxiogenic stimulus in numerous studies. Adult male and female rats were trained to discriminate PTZ (16 mg/kg i.p.) from saline in a two-lever food-reinforced task. They were then gonadectomized or sham-operated. Ovariectomized (OVX) rats were also tested during 17beta-estradiol (2.5 mg, 21 days release, s.c.) replacement. The PTZ dose response (0-16 mg/kg i.p.) was tested in all groups. In general, fewer females than males responded to PTZ. Diazepam (DZP; 0-10 mg/kg i.p.) injected before PTZ (16 mg/kg) decreased the number of rats selecting the PTZ lever. This effect was greater in sham female and estradiol-replaced-OVX rats than in male or OVX rats. Rats then received chronic ethanol diet (6.5%) for 10 days. During ethanol withdrawal (12 h after termination of the ethanol diet), they were tested for PTZ lever selection. PTZ lever selection differed between groups: sham or castrated male rats > OVX > sham female or estradiol-replaced-OVX rats. In sham female rats, estradiol concentrations showed a cyclic pattern with an estradiol surge that did not influence their PTZ discrimination performance. After i.p. injection of ethanol (2 g/kg), blood ethanol concentrations were not different in male and female rats. These findings suggest that 1) female rats are less sensitive to the anxiogenic effects of PTZ; 2) female rats are less sensitive to the anxiogenic effects of ethanol withdrawal; and 3) estrogen plays some role in mediation of these sex differences. PMID- 10525075 TI - Functional evidence for an angiotensin IV receptor in rat resistance arteries. AB - To distinguish between the different effects of angiotensin IV (Ang IV) on resistance artery vasoreactivity, freshly isolated rat mesenteric arteries were perfused and the changes in their diameter were recorded under various conditions. Ang IV exerted vasoconstrictor effects on both normal vessels and vessels that had been precontracted with phenylephrine or serotonin. This effect was abolished by losartan or candesartan cilexetil, two type 1 angiotensin receptor antagonists, but not by PD 123319, a type 2 angiotensin receptor antagonist. No tachyphylaxis was observed for the vasoconstrictor effect of Ang IV. N(G)-nitro-L-arginine methyl ester, a nitric oxide synthase inhibitor, had no effect on Ang IV-induced vasoconstriction, whereas indomethacin, a cyclooxygenase inhibitor that was inactive by itself, influenced Ang IV-induced vasoconstriction, suggesting that Ang IV could stimulate the release of prostaglandins. Treatment of preconstricted vessels by candesartan cilexetil unraveled a vasodilator effect of Ang IV that was abolished by PD 123319, a type 2 angiotensin receptor antagonist. Unexpectedly, Ang IV still produced a vasoconstrictor effect on normal or preconstricted vessels after blockade of both type 1 and type 2 angiotensin receptors. Taken together, these results show that Ang IV influences resistance artery vasoreactivity via different mechanisms, one of which implicates a functionally active type 4 angiotensin receptor. PMID- 10525076 TI - Effects of atrial natriuretic peptide on left ventricular performance in conscious dogs before and after pacing-induced heart failure. AB - Atrial natriuretic peptide (ANP) has potent vasodilatory and natriuretic actions and may have therapeutic benefit in congestive heart failure (CHF). These benefits may be offset by a negative inotropic effect of ANP seen in isolated preparations. However, ANP's integrated effect on left ventricular (LV) contraction and relaxation, independent of loading conditions, both under normal conditions and after CHF, is not known. We studied six conscious dogs, instrumented to measure LV and left atrial pressures and to determine LV volume from three dimensions. ANP produced significant (P<.05) decreases in LV end systolic pressure (101.2+/-11.8 versus 91.7+/-11.2 mm Hg, P<.05) in normal dogs and in dogs with CHF (93.1+/-6.4 versus 87.1+/- 4.4 mm Hg, P<.05). ANP also caused significant reductions of the slope of end-systolic pressure-end-systolic volume relation both before (7.0 +/-1.5 versus 6.3+/-1.5 mm Hg/ml) and after CHF (4.8+/-1.3 versus 4.4+/-1.2 mm Hg/ml, P<.05). Both before and after CHF, ANP slowed LV relaxation at matched end-systolic pressure. Before CHF, steady-state stroke volume and peak LV filling rate (dV/dt(max)) were reduced. However, after CHF, the fall in end-systolic pressure more than offset the load-independent LV depression, as stroke volume, the rate LV relaxation, and dV/dt(max) were increased and minimum LV pressure reduced. ANP has negative effects on LV contractility and relaxation both before and after CHF. However, after CHF, afterload reduction with ANP overcomes its negative effects, resulting in net improvement of LV ejection and relaxation. Thus, the direct cardiodepressant effects of ANP should not limit its usefulness in CHF. PMID- 10525077 TI - Molecular cloning and functional characterization of a polyspecific organic anion transporter from Caenorhabditis elegans. AB - We have cloned a polyspecific organic anion transporter from Caenorhabditis elegans and elucidated its functional characteristics. The C. elegans anion transporter (CeOAT1) codes for a protein of 526 amino acids containing 12 putative transmembrane domains. It exhibits significant homology at the level of amino acid sequence to the C. elegans organic cation transporter and to the mammalian organic cation and anion transporters. The function of CeOAT1 was investigated by expressing the transporter heterologously in mammalian cells. CeOAT1 transports p-aminohippurate (PAH) in a Na(+)-independent manner. The transport mechanism appears to involve anion exchange because CeOAT1-mediated PAH transport is stimulated by a cell-to-medium concentration gradient of alpha ketoglutarate or fumarate generated by coexpression in the cells of a mammalian Na(+)-coupled dicarboxylate transporter. CeOAT1 exhibits broad specificity, accepting anions such as folate, indomethacin, furosemide, probenecid, and benzylpenicillin as substrates. The Michaelis-Menten constant for the prototypical organic anion PAH is 0.43+/-0.07 mM. This constitutes the first report of the molecular and functional identification of a polyspecific organic anion transporter in C. elegans. PMID- 10525078 TI - Mechanisms of action of OPC-28326, a selective hindlimb vasodilator. AB - The unique cardiovascular profile of OPC-28326 [4-(N-methyl-2-phenylethylamino)-1 (3, 5-dimethyl-4-propionylaminobenzoyl)piperidine hydrochloride monohydrate] provides insight into basic mechanisms of this new drug as determined by experiments in dogs and rats. In anesthetized open-chest dogs, an i.v. administration of a low dose (0.3 and 1.0 microg/kg) of OPC-28326 selectively increased femoral artery blood flow with only minimal action on systemic blood pressure, heart rate and coronary, carotid, vertebral, renal, and mesenteric blood flows. Biochemical study suggests that OPC-28326 had no effect on phosphodiesterase-3 and -5. OPC-28326 dose-dependently inhibited phenylephrine induced increases in blood pressure in spinally anesthetized dogs. The potency of OPC-28326 was, however, about 180 times lower than that of prazosin. Although binding studies have revealed an affinity of OPC-28326 to serotonin 5-HT(2) receptors, the drug is without effect, except at very high concentrations, on serotonin-induced contraction in an isolated canine femoral artery preparation. The potency of OPC-28326 on the increase in femoral artery blood flow was about 14 times higher than that of prazosin but was at about the same level as that obtained with yohimbine in canine autoperfused femoral artery preparations. In perfused rat hindlimb preparations, OPC-28326 inhibited the decrease in perfusion flow induced by brimonidine, a selective alpha(2)-adrenoceptor agonist. The potency of OPC-28326 was at least 10 times less than that of yohimbine. Taken together, the results show that at low doses, OPC-28326 selectively exerts a potent vasodilating effect on the femoral arterial bed, in part due to an alpha(2)-adrenoceptor-blocking activity. PMID- 10525080 TI - Cannabinoid receptors can activate and inhibit G protein-coupled inwardly rectifying potassium channels in a xenopus oocyte expression system. AB - In this study, we focused on the pharmacological characterization of cannabinoid receptor coupling to G protein-gated inwardly rectifying potassium (GIRK) channels. Cannabinoids were tested on Xenopus laevis oocytes coexpressing the CB(1) receptor and GIRK1 and GIRK4 channels (CB(1)/GIRK1/4) or the CB(2) receptor and GIRK1/4 channels (CB(2)/GIRK1/4). WIN 55,212-2 enhanced currents carried by GIRK channels in the CB(1)/GIRK1/4 and CB(2)/GIRK1/4 system; however, the CB(2) receptor did not couple efficiently to GIRK1/4 channels. In the CB(1)/GIRK1/4 system, WIN 55,212-2 was the most efficacious compound tested. CP 55,940 and anandamide acted as partial agonists. The rank order of potency was CP 55,940 > WIN 55,212-2 = anandamide. The CB(1)-selective antagonist SR141716A alone acted as a inverse agonist by inhibiting GIRK currents in oocytes expressing CB(1)/GIRK1/4, suggesting the CB(1) receptor is constitutively activated. A conserved aspartate residue, which was previously shown to be critical for G protein coupling in cannabinoid receptors, was mutated (to asparagine, D163N) and analyzed. Oocytes coexpressing CB(1)/GIRK1/4 or D163N/GIRK1/4 were compared. The potency of WIN 55, 212-2 at the mutant receptor was similar to wild type, but its efficacy was substantially reduced. CP 55,940 did not elicit currents in oocytes expressing D163N/GIRK1/4. In summary, it appears the CB(1) and CB(2) receptors couple differently to GIRK1/4 channels. In the CB(1)/GIRK1/4 system, cannabinoids evaluated demonstrated the ability to enhance or inhibit GIRK currents. Furthermore, a conserved aspartate residue in the CB(1) receptor is required for normal communication with GIRK channels in oocytes demonstrating the interaction between receptor and channels is G protein dependent. PMID- 10525079 TI - Design and characterization of orally active Arg-Gly-Asp peptidomimetic vitronectin receptor antagonist SB 265123 for prevention of bone loss in osteoporosis. AB - The Arg-Gly-Asp (RGD)-binding integrin alpha(V)beta(3) is highly expressed on osteoclasts and has been proposed to mediate cell-matrix adhesion required for osteoclast-mediated bone resorption. Antagonism of this receptor should prevent stable osteoclast adhesion and thereby inhibit bone resorption. We have generated an orally bioavailable, nonpeptide RGD mimetic alpha(v)beta(3) antagonist, SB 265123, which prevents bone loss in vivo when dosed by oral administration. SB 265123 binds alpha(v)beta(3) and the closely related integrin alpha(v)beta(5) with high affinity (K(i) = 3.5 and 1.3 nM, respectively), but binds only weakly to the related RGD-binding integrins alpha(IIb)beta(3) (K(i) >1 microM) and alpha(5)beta(1) (K(i) >1 microM). The compound inhibits alpha(v)beta(3)-mediated cell adhesion with an IC(50) = 60 nM and more importantly, inhibits human osteoclast-mediated bone resorption in vitro with an IC(50) = 48 nM. In vivo, SB 265123 completely blocks bone resorption in a thyroparathyroidectomized rat model of acute bone resorption when dosed at 2.5 mg/kg/h by continuous i.v. infusion. When dosed orally with 3 to 30 mg/kg b.i.d. , in the ovariectomy-induced rat model of osteoporosis, SB 265123 prevents bone resorption in a dose-dependent fashion. This is the first report of an orally active alpha(v)beta(3) antagonist that is effective at inhibiting bone resorption when dosed in a pharmaceutically acceptable fashion. Such a molecule may provide a novel therapeutic agent for the treatment of postmenopausal osteoporosis. PMID- 10525081 TI - Developmental regulation of endothelial nitric oxide synthase in cerebral vessels of newborn pig by prostaglandin E(2). AB - We investigated whether prostaglandins regulate endothelial nitric oxide synthase (eNOS) in the pig cerebral vasculature during the neonatal period. Prostaglandins, eNOS mRNA, eNOS protein, and NO production were higher in cerebral microvessels of newborn (1 day old) than in those of adult (6- to 8 month-old) pigs. The treatment of isolated cerebral microvessels of newborn animals with ibuprofen for 24 h reduced eNOS mRNA and nitrite production to levels in the adult; this effect of ibuprofen was prevented by concurrent treatment with prostaglandin (PG)E(2) analog 16,16-dimethyl-PGE(2), nonselective PGE(2) receptor analog 11-deoxy PGE(1), and prostaglandin EP(3) receptor agonists sulprostone and M&B 28,767 but was not modified by PGI(2) analog carbaprostacyclin, PGD(2), and EP(1) receptor agonist 17-phenyl trinor PGE(2). Correspondingly, 16, 16-dimethyl-PGE(2) and M&B 28,767 increased eNOS mRNA expression of adult microvessels to values in the newborn. Data similar to those with isolated cerebral vessels were obtained through histochemical analysis (NADPH-diaphorase positivity) of brain from newborn animals treated in vivo with ibuprofen in combination or not with sulprostone. Furthermore, substance P induced NO-mediated cerebral vasorelaxation was decreased to adult values through the treatment of newborn pigs with ibuprofen; this effect was prevented by concomitant treatment with sulprostone. It is concluded that PGE(2) regulates eNOS in newborn pig cerebral microvessels via EP(3) receptors; this may be physiologically required during normal neurovascular development. PMID- 10525082 TI - Regulation by endogenous interleukin-1 of mRNA expression of healing-related factors in gastric ulcers in rats. AB - We investigated the role of endogenous interleukin (IL)-1 in the mRNA expression of cyclooxygenase (COX)-1, COX-2, inducible nitric oxide synthase (iNOS), cytokine-induced neutrophil chemoattractant (CINC)-1, epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), hepatocyte growth factor (HGF), and transforming growth factor (TGF)-beta1 in acetic acid-induced gastric ulcers in rats. IL-1beta mRNA was not detected in the normal or intact mucosa of ulcerated stomachs, but its expression was induced in the ulcerated tissue. IL-1beta immunoreactivity was observed in macrophages/monocytes and fibroblasts in the ulcer base. COX-2, iNOS, and CINC-1 mRNAs were expressed by ulceration. EGF, bFGF, HGF, and TGF-beta1 mRNA expression was detected in the normal mucosa, and their levels were significantly elevated by ulceration. In contrast, COX-1 mRNA level did not differ between the normal and ulcerated tissues. In a culture of isolated ulcer bases, block of IL-1 with IL-1 receptor antagonist (IL-1RA) dose dependently and significantly reduced the mRNA levels of COX-2, iNOS, CINC-1, HGF, and bFGF. In contrast, COX-1, EGF, and TGF-beta1 mRNA expression was not affected by IL-1RA. IL-1RA dose-dependently reduced prostaglandin E(2) production, total and iNOS activities, neutrophil chemotactic activity, and growth-promoting activity toward gastric epithelial cells in the ulcer base. Finally, the administration of IL-1RA caused a significant impairment of ulcer healing. These results indicate that IL-1, expressed in macrophages/monocytes and fibroblasts in the ulcer base, might up-regulate the mRNA expression of COX-2, iNOS, CINC-1, HGF, and bFGF, thereby contributing to gastric ulcer healing in rats. PMID- 10525083 TI - Beneficial effects of ropivacaine in rat experimental colitis. AB - Ropivacaine, a new, long-acting local anesthetic agent, has been shown to have beneficial effects in the treatment of ulcerative colitis. Treatment with this drug results in prompt symptomatic relief. The aim of this study was to examine the effects of ropivacaine on mucosal healing and to investigate whether ropivacaine can restore the decreased colonic contractility seen in the diseased state. Colitis was induced in rats by a single intrarectal administration of trinitrobenzene sulfonic acid. Mucosal healing was assessed after 1 week of therapy. The effects on colonic contractility were examined either after 1 week of treatment or by application of the drugs to untreated, inflamed rat colon segments placed in organ baths. After the induction of colitis, daily intracolonic treatment with ropivacaine for 1 week reduced morphological damage and myeloperoxidase activity. One week of treatment also restored the contractile response to acetylcholine. By adding ropivacaine directly to untreated inflamed colonic segments in organ baths, the contractile response to acetylcholine was increased compared with controls. For comparison, the effects of budesonide and 5 aminosalicylic acid were also examined. Ropivacaine improved mucosal healing and restored colonic motor activity in experimental colitis, similar to budesonide but superior to 5-aminosalicylic acid. PMID- 10525084 TI - Pharmacological characterization of (E)-N-(3-iodoprop-2-enyl)-2beta-carbomethoxy 3beta-(4'-methylphenyl)n ortropane as a selective and potent inhibitor of the neuronal dopamine transporter. AB - The pharmacological properties of the iodinated derivative of cocaine (E)-N-(3 iodoprop-2-enyl)-2beta-carbomethoxy-3beta-(4'-me thylphenyl)nortropane (PE2I) were evaluated in vitro in the rat. Binding experiments on rat striatal membranes showed that PE2I selectively recognized the dopamine transporter (DAT) according to a single binding site model with high affinity (K(d) = 4 nM, B(max) = 12 pmol/mg protein). In the cortical membranes, the binding of PE2I was also selectively associated with the DAT (IC(50) for GBR 12909 = 6 nM versus more than 1000 nM for paroxetine), with similar affinity to that of the striatum. Autoradiographic experiments on rat brain sections with [(125)I]PE2I were in agreement with the localization of the DAT. In addition, PE2I was shown to be a potent inhibitor of dopamine uptake, with IC(50) values similar to those for GBR 12909 and 2beta-carbomethoxy-3beta-(4'-iodophenyl)-tropane (beta-CIT) (2-6 nM). All of these findings, combined with previously published data, support the use of PE2I as a selective and potent tool to study the DAT both in vivo and in vitro. PMID- 10525085 TI - Adenosine A(2A) receptors mediate coronary microvascular dilation to adenosine: role of nitric oxide and ATP-sensitive potassium channels. AB - Adenosine is a potent vasodilator that plays an important role in the regulation of coronary microvascular diameter. Although multiple adenosine receptor subtypes have been recently cloned, the specific adenosine receptor subtypes and the underlying mechanisms responsible for the vasodilation to adenosine in the coronary microcirculation remain unknown. Therefore, in the present study we determined the receptor subtypes for coronary arteriolar dilation to adenosine and investigated the role of nitric oxide (NO) and ATP-sensitive potassium (K(ATP)) channels in this vasodilatory response. Pig coronary arterioles (50-100 microm in situ) were isolated, cannulated, and pressurized without flow for in vitro study. Arterioles developed basal tone and dilated in a concentration dependent manner to adenosine and to adenosine receptor agonists (2S)-N(6)-[2 endo-norbornyl]adenosine (A(1)), 2-[p-(2-carboxyethyl)]phenylethyl-amino-5'-N ethylcarboxamidoadenosin e (CGS21680; A(2A)), N(6)-(3-iodobenzyl)adenosine-5'-N methyluronamide (A(3)), and N-ethylcarboxamidoadenosine (nonselective adenosine receptor activation). The selective A(2A) receptor antagonist 4-(2-[7-amino-2-(2 furyl)[1,2,4]-triazolo[2,3-a][1,3,5]triazin-5-y l amino]ethyl)phenol attenuated vasodilation to adenosine and to all adenosine receptor agonists tested, suggesting that the vasodilatory responses were primarily mediated by A(2A) receptors. Adenosine- and CGS21680-induced dilations were attenuated in a similar manner by endothelial removal and by the NO synthase inhibitor N(G)-nitro-L arginine methyl ester. In denuded vessels, both adenosine- and CGS21680-induced dilations were nearly abolished by the K(ATP) channel inhibitor glibenclamide. The selective A(2A) agonist CGS21680 mechanistically mimics the vasodilation in response to adenosine. Collectively, our results suggest that the dilation of coronary arterioles to adenosine is mediated predominantly by A(2A) receptors. Activation of this receptor subtype elicits vasodilation by endothelial release of NO and by the smooth muscle opening of K(ATP) channels. PMID- 10525086 TI - Characterization of interleukin-1alpha binding to mouse brain endothelial cells. AB - In vivo studies have shown that interleukin (IL)-1alpha binds to and is transported across brain endothelial cells, whereas in vitro studies have shown that brain endothelial cells respond to IL and contain mRNA for the IL-type 1 receptor. However, these binding sites have yet to be characterized. Herein, we used murine brain microvessels to characterize the binding of IL labeled with (125)I. Binding was temperature- and time-dependent with maximal binding after 4 h of incubation at 37 degrees C. The amount of radioactivity determined by HPLC to represent intact (125)I-labeled murine IL-1alpha at 4 h was approximately 100% in the incubation fluid and 80 to 90% for radioactive material recovered from the incubated cells. B(max) was 0.955 fmol and the K(d) was 292 pM for human (125)I IL and binding was displaced by interleukin-1beta and interleukin-1 receptor antagonist but not by tumor necrosis factor alpha. Binding was dependent on magnesium and glucose. Incubation with antibodies showed that the binding site was not identical with the IL-type 1 receptor but closely resembled the blood brain barrier transporter. These results show that murine brain endothelial cells have specific binding sites for IL and that these sites more closely resemble the transporter than the type 1 receptor. PMID- 10525087 TI - Alpha-adrenoceptors in canine mesenteric artery are predominantly 1A subtype: pharmacological and immunochemical evidence. AB - We wanted to determine which alpha-adrenoceptor subtypes mediate phenylephrine (PE) contraction of dog mesenteric artery in vitro. We studied antagonisms in response to prazosin, 2-(2, 6-dimethoxyphenoxyethyl)-aminomethyl-1,4 benzodioxane, 5-methylurapidil, N-[2-(2-cyclopropyl methoxy phenoxy)ethyl]5 chloro-alpha,alpha-dimethyl-1H-indole-3-ethanamine HCl (RS 17053), 8-3-[4-(2 methoxyphenyl)-1-piperazinyl]propylcarbamoyl)-3-methyl-4 -oxo-22-phenyl-4H-1 benzopyran 2HCl [SB216469 (Rec 15/2739)], BMY 7378, 8-[2-(1,4-benzodioxan-2 ylmethylamino)ethyl]8-azaspirol++ + [4,5]decane-7,9-dione HCl, MDL 72832, and 7 chloro-2-bromo-3,4,5, 6-tetrahydro-4-methylfurol[4,3,2-ef]3-benzapine. pK(B) values for prazosin, 5-methylurapidil, MDL 72832, and RS-17053 were consistent with action on alpha(1A)-adrenoceptors but decreased with concentration. pK(B) values (9.6) for Rec 15/2739 (alpha(1L/1A)-adrenoceptor selective) were constant. Antagonism by BMY 7378, 7-chloro-2-bromo-3,4,5,6-tetrahydro-4-methylfurol[4,3, 2 ef]3-benzapine, and 8-[2-(1, 4-benzodioxan-2-ylmethylamino)ethyl]8 azaspirol[4,5]de cane-7,9-dione HCl gave pK(B) values between those expected for alpha(1A)- and alpha(1D)-adrenoceptors. Chloroethylclonidine (100 microM) shifted EC(50) values for PE rightward and decreased E(max) values but left large residual responses. After 100 microM chloroethylclonidine, either BMY 7378 (100 nM) or RS-17053 (300 nM) increased EC(50) values for PE contractions with pK(B) values like those of controls. At 6 nM, phenoxybenzamine increased the EC(50) values and reduced E(max) values; prior Rec 15/2739, but not prior BMY 7378, protected receptors against inactivation. An antibody against the alpha(1B) adrenoceptors immunostained muscle of aorta but not mesenteric artery. We conclude that dog mesenteric artery contains alpha(1A)-adrenoceptors. Discrepancies among responses expected if only these receptors are present may result from pleiotropic functional effects at this receptor and the presence of alpha(1L)-adrenoceptors. PMID- 10525088 TI - RWJ 67657, a potent, orally active inhibitor of p38 mitogen-activated protein kinase. AB - Tumor necrosis factor-alpha (TNF-alpha), a cytokine secreted by activated monocytes/macrophages and T lymphocytes, has been implicated in several disease states, including rheumatoid arthritis, inflammatory bowel disease, septic shock, and osteoporosis. Monocyte/macrophage production of TNF-alpha is dependent on the mitogen-activated protein kinase p38. RWJ 67657 (4-[4-(4-fluorophenyl)-1-(3 phenylpropyl)-5-(4-pyridinyl)-1H-imidazol -2-yl]-3-butyn-1-ol) inhibited the release of TNF-alpha by lipopolysaccharide (a monocyte stimulus)-treated human peripheral blood mononuclear cells with an IC(50) of 3 nM, as well as the release of TNF-alpha from peripheral blood mononuclear cells treated with the superantigen staphylococcal enterotoxin B (a T cell stimulus), with an IC(50) value of 13 nM. This compound was approximately 10-fold more potent than the literature standard p38 kinase inhibitor SB 203580 in all p38 dependent in vitro systems tested. RWJ 67657 inhibited the enzymatic activity of recombinant p38alpha and beta, but not gamma or delta, in vitro and had no significant activity against a variety of other enzymes. In contrast, SB 203580 significantly inhibited the tyrosine kinases p56 lck and c-src (IC(50) = 5 microM). RWJ 67657 did not inhibit T cell production of interleukin-2 or interferon-gamma and did not inhibit T cell proliferation in response to mitogens. RWJ 67657 inhibited TNF alpha production in lipopolysaccharide-injected mice (87% inhibition at 50 mg/kg) and in rats (91% inhibition at 25 mg/kg) after oral administration. Based on these favorable biological properties, RWJ 67657 may have use as a treatment for inflammatory diseases. PMID- 10525089 TI - Effects of treatment with haloperidol, chlorpromazine, and clozapine on protein kinase C (PKC) and phosphoinositide-specific phospholipase C (PI-PLC) activity and on mRNA and protein expression of PKC and PLC isozymes in rat brain. AB - The effects of acute (single) and chronic (21-day) administration of haloperidol (HAL), chlorpromazine (CPZ), or clozapine (CLOZ) on components of the phosphoinositide (PI)-signaling pathway were studied in rat brain. Chronic administration of HAL decreased protein kinase C (PKC) activity and mRNA and protein levels of PKC alpha and epsilon isozymes in both membrane and cytosol fractions of cortex, hippocampus, and striatum. Chronic administration of CPZ, however, decreased PKC activity only in the membrane fraction of cortex, hippocampus, and striatum, and had no effect on the levels of any PKC isozymes. On the other hand, chronic administration of CLOZ decreased PKC activity and mRNA and protein levels of PKC alpha, gamma, and epsilon isozymes in membrane and cytosol fractions of cortex, hippocampus, and cerebellum. Studies of the effects on phospholipase C (PLC) revealed that only chronic administration of CPZ significantly decreased PI-PLC activity and mRNA and protein levels of the specific PLC beta(1) isozyme in membrane and cytosol fractions of cortex, hippocampus, cerebellum, and striatum. Acute-treatment data suggest that CPZ or CLOZ had no significant effects on PI-PLC or PKC; however, HAL translocated PKC, as evidenced from increased PKC activity and protein levels of PKC alpha and epsilon isozymes in the membrane fraction and the decrease in these parameters in the cytosol fraction of cortex, hippocampus, and striatum. Our results thus suggest that the interaction of antipsychotic drugs with PKC and PLC may be associated with their mechanisms of action. PMID- 10525090 TI - Recognition of L-amino acid ester compounds by rat peptide transporters PEPT1 and PEPT2. AB - Peptide transporters (PEPT1 and PEPT2) in epithelia play an important role in the absorption of small peptides and peptide-like drugs. Recently, it was demonstrated that various nonpeptidic compounds can be transported by these transporters. In the present study, we focused on the L-amino acid ester compounds and examined the mechanisms of their interaction with rat PEPTs (rPEPTs) using stable transfectants. Valacyclovir, the L-valyl ester prodrug of the antiherpetic agent acyclovir, competitively inhibited [(14)C]glycylsarcosine uptake in the rPEPT1- or rPEPT2-expressing cells. Dixon plot analyses showed that the inhibition constant (K(i)) values of valacyclovir were 2.7 and 0.22 mM for rPEPT1 and rPEPT2, respectively, suggesting that rPEPT2 had higher affinity for this agent. Various L-valine alkyl esters significantly inhibited [(14)C]glycylsarcosine uptake. L-Valine methyl ester (Val-OMe) competitively inhibited [(14)C]glycylsarcosine uptake with K(i) values of 3.6 and 0.83 mM for rPEPT1 and rPEPT2, respectively, indicating that Val-OMe is also a high-affinity substrate for rPEPT2. Val-OMe had a trans-stimulation effect on [(14)C]glycylsarcosine efflux from both transfectants, suggesting the translocation of L-valine methyl ester via rPEPTs. Val-OMe showed the most potent inhibitory effect among the several L-amino acid methyl esters examined. We conclude that Val-OMe, as well as valacyclovir, could be recognized and transported by rPEPT1 and rPEPT2 and that these L-valyl esters showed higher affinity for rPEPT2 as do most substrates of these transporters. Our results suggest that L-valine is a desirable L-amino acid for the esterification of poorly permeable drugs to enhance their oral bioavailability targeting intestinal PEPT1. PMID- 10525091 TI - Role of an ATP-sensitive potassium channel opener, YM934, in mitochondrial energy production in ischemic/reperfused heart. AB - We examined a possible mechanism of action of an ATP-sensitive potassium (K(ATP)) channel opener, YM934, for the improvement of energy metabolism in hearts subjected to 35-min ischemia and 60-min reperfusion. The treatment with 30 nM YM934 for the final 15 min of preischemia enhanced postischemic recovery of left ventricular developed pressure, attenuated the postischemic rise in left ventricular end-diastolic pressure, and suppressed the release of creatine kinase and ATP metabolites during reperfusion. The treatment also restored myocardial ATP and creatine phosphate contents and attenuated the decrease in mitochondrial oxygen consumption rate during reperfusion. The higher mitochondrial function was also seen in YM934-treated hearts at the end of ischemia. In another set of experiments, myocardial skinned bundles were incubated for 30 min under hypoxic conditions in the presence and absence of YM934, and then mitochondrial oxygen consumption rate was determined. Hypoxia decreased the mitochondrial oxygen consumption rate of skinned bundles to approximately 40% of the prehypoxic value. In contrast, the treatment of skinned bundles with 30 nM YM934 preserved the mitochondrial oxygen consumption rate during hypoxia. The effect of YM934 on the hypoxic skinned bundles was abolished by combined treatment with either the K(ATP) channel blocker glyburide or the mitochondrial K(ATP) channel blocker 5 hydroxydecanoate in a concentration-dependent manner. The results suggest that YM934 is capable of attenuating ischemia/reperfusion injury of isolated perfused hearts due to preservation of mitochondrial function during ischemia, probably through opening of mitochondrial K(ATP) channels. PMID- 10525092 TI - K(+)-induced neurogenic relaxation of rat distal colon. AB - Relaxations of segments of rat distal colon were elicited by hypertonic solutions of potassium (K(+); final concentration, 20.8 or 50.8 mM). The initial part of the response to K(+) was antagonized by the nerve blocker tetrodotoxin. This effect could, moreover, be significantly antagonized by apamin (a blocker of K(+) channels), reactive blue 2 (a P(2y)-purinoceptor antagonist), N(G)-nitro-L arginine (an inhibitor of NO synthase), 1H-[1,2,4]- oxadiazolo[4,3-a]quinoxaline 1-one (ODQ; an inhibitor of soluble guanylyl cyclase), or N-[2-(p bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide (H-89; an inhibitor of cAMP dependent protein kinase). Sodium nitroprusside (a donor of NO) and vasoactive intestinal peptide (VIP) both relaxed the tissues. The response to sodium nitroprusside was abolished by ODQ and unaffected by H-89, and that to VIP was partially inhibited by VIP(10-28) (a VIP receptor antagonist), ODQ, or H-89. When combining reactive blue 2 and N(G)-nitro-L-arginine, the response to 50.8 mM K(+) was reduced by approximately 70% and was abolished by the concomitant administration of these antagonists and VIP(10-28). ATP, NO, and VIP may, thus, be inhibitory neurotransmitters in rat distal colon. PMID- 10525093 TI - Canine cardiac muscarinic receptors, G proteins, and adenylate cyclase after long term morphine. AB - Short-term morphine stimulates vagal bradycardia. This led us to propose the hypothesis that chronically administered morphine would down-regulate myocardial muscarinic receptor systems. Dogs received morphine continuously for 2 weeks through an s.c. catheter, and cellular aspects of parasympathetic control of the heart were examined. Contrary to expectations, morphine increased muscarinic receptor density in the right atrium and left ventricle by 17 and 34%, respectively, with no change in the apparent affinity of the receptor (K(D)). Morphine also increased the expression of the G protein G(ialpha) by 115 and 233%, respectively, in right atrial and left ventricular sarcolemmal membranes. Morphine increased ventricular and atrial G(salpha) to a much lesser degree (49 and 25%). Morphine failed to alter basal or maximally stimulated (forskolin plus MnCl(2)) adenylate cyclase activity. The maximum cyclase activation by isoproterenol and the maximum inhibition by carbachol were similarly unaltered by morphine. Morphine reduced the ventricular but not atrial norepinephrine. Both long- and short-term morphine lowered tissue epinephrine content, suggesting that short-term morphine reduces extraneuronal uptake. Potential systemic and cellular models for myocardial adaptation to morphine are proposed, including sequential sympathetic and parasympathetic compensations. PMID- 10525094 TI - Hypoxia-reoxygenation-induced apoptosis in cultured adult rat myocytes and the protective effect of platelets and transforming growth factor-beta(1). AB - The outcome of myocardial ischemia-reperfusion has been partially attributed to the degree of apoptosis in cardiomyocytes. Aggregating platelets by release of transforming growth factor-beta(1) (TGF-beta(1)) protect the isolated heart against ischemia-reperfusion injury and preserve myocardial TGF-beta(1) content. To gain more insight into the modulation of hypoxia-reoxygenation-induced injury (apoptosis and necrosis) to myocytes by TGF-beta(1) and aggregating platelets, cultured adult rat myocytes were exposed for 48 or 72 h to hypoxia alone, or to hypoxia followed by 3 h of reoxygenation. Apoptosis in the cells was determined by in situ terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling staining and DNA fragmentation on gel electrophoresis. Hypoxia alone caused a time-dependent increase in myocyte apoptosis (number of apoptotic cells: 19+/-3% at 48 h and 39+/-5% at 72 h compared with 5+/-1% in control cells, based on a 500-cell count). Three hours of reoxygenation after 48 h of hypoxia further increased the number of apoptotic cells (34+/-8 versus 19+/-3% in hypoxia for 48 h), but reoxygenation after 72 h of hypoxia did not additionally increase the number of apoptotic cells, perhaps because of extensive cell necrosis on prolonged hypoxia. Forty-eight hours of hypoxia followed by 3 h of reoxygenation also resulted in a decrease in Bcl-2 and an increase in Fas protein level. Incubation of myocytes with either recombinant TGF-beta(1) (0.5-5 ng/ml) or aggregated platelet supernatant (from 2-3 x10(7) platelets/ml, containing approximately 0.5 ng/ml of TGF-beta(1)) markedly (P<.01) decreased the number of apoptotic cells after hypoxia-reoxygenation. Incubation with TGF-beta(1) also reduced myocyte necrosis as evident from lactate dehydrogenase release and trypan blue dye exclusion. These data demonstrate that hypoxia-reoxygenation results in apoptosis and necrosis in cultured adult rat myocytes; this can be attenuated by TGF-beta(1). Similarity of data with TGF-beta(1) and aggregated platelet supernatant suggests that platelet-mediated cardioprotection during hypoxia reoxygenation may relate in part to the release of TGF-beta(1). PMID- 10525095 TI - Inhibition of epidermal growth factor receptor-associated tyrosine phosphorylation in human carcinomas with CP-358,774: dynamics of receptor inhibition in situ and antitumor effects in athymic mice. AB - Phosphorylation of tyrosine residues on the epidermal growth factor (EGF) receptor (EGFr) is an important early event in signal transduction, leading to cell replication for major human carcinomas. CP-358,774 is a potent and selective inhibitor of the EGFr tyrosine kinase and produces selective inhibition of EGF mediated tumor cell mitogenesis. To assess the pharmacodynamic aspects of EGFr inhibition, we devised an ex vivo enzyme-linked immunosorbent assay for quantification of EGFr-specific tyrosine phosphorylation in human tumor tissue specimens obtained from xenografts growing s.c. in athymic mice. When coupled with pharmacokinetic analyses, this measurement can be used to describe the extent and duration of kinase inhibition in vivo. CP-358,774 is an effective, orally active inhibitor of EGFr-specific tyrosine phosphorylation (ED(50) = 10 mg/kg, single dose). It has a significant duration of action, producing, on average, a 70% reduction in EGFr-associated phosphotyrosine over a 24-h period after a single 100 mg/kg dose. Inhibition of EGFr phosphotyrosine in an ex vivo assay format effectively estimates the potency and degree of inhibition of EGFr dependent human LICR-LON-HN5 head and neck carcinoma tumor growth. Substantial growth inhibition of human tumor xenografts was achieved with p.o. doses of the compound (ED(50) = 10 mg/kg q.d. for 20 days). Combination chemotherapy with cisplatin produced a significant response above that of cisplatin alone with no detectable effects on body weight or lethal toxicity. Taken together, these observations suggest that CP-358,774 may be useful for the treatment of EGFr driven human carcinomas. PMID- 10525096 TI - Role of a potent inhibitory monoclonal antibody to cytochrome P-450 3A4 in assessment of human drug metabolism. AB - Cytochrome P-450 (CYP) 3A4 is an inordinately important CYP enzyme that catalyzes the metabolism of a vast array of clinically used drugs. Microsomal proteins of Spodoptera frugiperda (Sf21) insect cells infected with recombinant baculoviruses encoding CYP3A4 cDNA were used to immunize mice and to develop a monoclonal antibody (mAb(3A4a)) specific to CYP3A4 through the use of hybridoma technology. The mAb is both a potent inhibitor and a strong binder of CYP3A4. One and 5 microl (0.5 and 2.5 microM IgG(2a)) of the mAb mouse ascites in 1-ml incubation containing 20 pmol of CYP3A4 strongly inhibited the testosterone 6beta hydroxylation by 95 and 99%, respectively, and, to a lesser extent, cross inhibited CYP3A5 and CYP3A7 activity. mAb(3A4a) exhibited no cross-reactivity with any of the other recombinant human CYP isoforms (CYP1A1, CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, and CYP2E1) in the course of CYP reaction phenotyping and Western immunoblot analyses. The potency of mAb-induced inhibition is insensitive to substrate concentration in human liver microsomes. Therefore, mAb(3A4a) was used to assess the quantitative role of CYP3A4/5 to the metabolism of testosterone and diazepam in five human liver microsomes. The results showed that CYP3A4 and CYP3A5 contribute >95% to both testosterone 6beta hydroxylation and diazepam 3-hydroxylation and 52 to 73% to diazepam N demethylation, respectively. In addition, mAb(3A4a) significantly inhibited testosterone 6beta-hydroxylase activity in rhesus monkey liver microsomes to a degree equal to that observed with CYP3A4 in human liver microsomes. By comparison, no inhibition of testosterone 6beta-hydroxylase activity was observed in the presence of dog, rat, and mouse liver microsomes. The selectivity of ketoconazole, a chemical inhibitor of CYP3A4, was probed with mAb(3A4a) and was shown to be highly concentration dependent in the diazepam N-demethylation by human liver microsomes. The results demonstrate that inhibitory and immunoblotting mAb(3A4a) can offer a precise and useful tool for quantitative identification of CYP3A4/5 in the metabolism of drugs in clinical use and drugs in development. PMID- 10525097 TI - High-affinity interaction of (des-Tyrosyl)dynorphin A(2-17) with NMDA receptors. AB - The opioid peptide dynorphin A elicits non-opioid receptor-mediated, neurotoxic response in vivo, which is blocked by pretreatment with MK-801, a noncompetitive N-methyl-D-aspartate receptor (NMDAR) antagonist. In the present study, we examined the possible direct interaction of dynorphin A on the NMDAR. A nonopioid dynorphin A analog, (125)I-(des-tyrosyl) dynorphin A(2-17), was used in radioligand binding analysis on rat cortical brain membranes. This radioligand exhibited a saturable, specific binding at high affinity with a K(d) value of 9.4+/-1.6 nM and maximal binding of 2.4+/-0.6 pmol/mg protein. This binding site was associated with the NMDAR complex because it was modulated by a number of NMDAR ligands. Transient expression of the rat NR1a/NR2A complex in human embryonic kidney 293 cells confirmed a coexpression of (125)I-(des-tyrosyl) dynorphin A(2-17), [(3)H]CGP39,653, and [(3)H]MK-801 binding. These data provide direct evidence of the presence of a high-affinity binding site for dynorphin A on the NMDAR. The modulatory effect of the various NMDAR-selective ligands on dynorphin A binding suggests that dynorphin A may bind preferentially to the closed/desensitized state of the NMDAR. The physiological role of dynorphin A binding to the NMDAR remains to be established. PMID- 10525098 TI - A transgenic model of acetaldehyde overproduction accelerates alcohol cardiomyopathy. AB - Chronic alcohol consumption produces alcoholic heart muscle disease (AHMD), a prevalent form of congestive heart failure. Several hypotheses have been proposed to explain the damaging effects of alcohol on the heart, but neither the mechanism nor the ultimate toxin has been established. In this study, we use transgenic overexpression of alcohol dehydrogenase to elevate cardiac exposure to acetaldehyde, the major and most reactive metabolite of alcohol. Overexpression of alcohol dehydrogenase by 40-fold produced no detectable deleterious effects to the heart in the absence of alcohol. In the presence of alcohol, transgenic hearts contained 4-fold higher acetaldehyde than control hearts. Chronic alcohol exposure produced many changes similar to AHMD in transgenic hearts. Compared with control hearts, these pathological changes occurred more rapidly and to a greater extent: alcohol-exposed transgenic hearts were almost twice as large as control hearts. They demonstrated ultrastructural damage consistent with AHMD and had much lower contractility than alcohol-exposed control hearts. In addition, the transgenic hearts showed greater changes in mRNA expression for alpha skeletal actin and atrial natriuretic factor than alcohol-exposed control hearts. Alterations in NAD(+)/NADH levels were insufficient to account for such severe damage in cardiomyopathic hearts. The increased damage produced in transgenic hearts suggests an important role for acetaldehyde in AHMD. PMID- 10525099 TI - Combined effects of buffer and adrenergic agents on postresuscitation myocardial function. AB - Although buffer agents alone have failed to improve the success of resuscitation, we now examine the widely held concept that it is the combined effect of alkaline buffer and adrenergic agents that improves outcomes of cardiopulmonary resuscitation. In the present report, the effects of both CO(2)-consuming and CO(2)-generating buffer agents in combination with adrenergic vasopressor drugs were investigated. Ventricular fibrillation was electrically induced in Sprague Dawley rats weighing between 450 and 550 g. Precordial compression and mechanical ventilation were initiated after 8 min of untreated ventricular fibrillation. Animals were then randomized to receive bolus injections of either inorganic sodium bicarbonate buffer, organic tromethamine buffer, or saline placebo. The beta(1) adrenergic effects of epinephrine were blocked with esmolol. The vasopressor amine was injected 2 min after injection of the buffer agent. Electrical defibrillation was attempted at the end of 8 min of precordial compression. In 15 additional animals, the sequence of administration of the adrenergic vasopressor and buffer agents was reversed such that the adrenergic vasopressor was injected before the buffer agents. All animals were restored to spontaneous circulation. Both bicarbonate and tromethamine significantly decreased coronary perfusion pressure from 26 to 15 mm Hg and reduced the magnitude of the vasopressor effect of the adrenergic vasopressor. When the vasopressor preceded the buffer, declines in coronary perfusion pressure after administration of buffer agents were prevented. In each instance, however, greater impairment of postresuscitation myocardial function and decreased postresuscitation survival were observed after treatment with buffer agents. PMID- 10525100 TI - Na(+)-dependent carnitine transport by organic cation transporter (OCTN2): its pharmacological and toxicological relevance. AB - Carnitine deficiency, either primary or drug-induced, causes critical symptoms and is thought to involve alteration of active transport of carnitine across the plasma membrane of tissues as the underlying mechanism. Recently, we showed that human organic cation transporter, hOCTN2, cloned as a member of the organic cation transporter family, is a physiologically important Na(+)-dependent high affinity carnitine transporter in humans. In this study, we further characterized the functional properties of hOCTN2 and examined the interaction between hOCTN2 mediated carnitine transport and clinically used drugs to assess possible toxicological effects. When expressed in human embryonic kidney (HEK)293 cells, hOCTN2 showed low but significant stereospecific transport activity: D-carnitine was transported with lower affinity (K(m) = 10.9 microM) than the L-isomer (K(m) = 4.3 microM). One Na(+) appeared to be associated with the transport of one carnitine molecule. hOCTN2-mediated transport of acetyl-L-carnitine was also Na(+)-dependent and of high affinity, with a K(m) value of 8.5 microM. To examine the transport activity for organic cations other than carnitine and the possible relationship of drug-induced carnitine deficiency with hOCTN2, the inhibitory effect of several drugs on hOCTN2-mediated L-carnitine transport was examined. Many zwitterionic drugs, such as cephaloridine, and many cationic drugs, such as quinidine and verapamil, exhibited significant inhibitory effects. Among these inhibitors, tetraethylammonium, pyrilamine, quinidine, verapamil, and valproate were found to be transported by hOCTN2. The results suggest that the carnitine deficiency-related toxicological effects by long-term treatment with such drugs might be ascribed to a functional alteration of hOCTN2-mediated carnitine transport. PMID- 10525101 TI - A critical role of the N-methyl-D-aspartate (NMDA) receptor subunit (NR) 2A in the expression of redox sensitivity of NR1/NR2A recombinant NMDA receptors. AB - In recombinant N-methyl-D-aspartate (NMDA) receptors, two redox modulatory sites are thought to exist, one formed by Cys744 and Cys798 on NMDA receptor subunit (NR) 1, and a second one, not yet localized, on NR2A. Reductants increase the open dwell-time and opening frequency of NR1/NR2A channels. In contrast, NR1/NR2B and NR1/NR2C channels exhibit changes only in opening frequency after redox treatments. Here, we evaluated whether the two redox sites act independently of each other, with the NR1 site affecting the opening frequency and the NR2A site altering open dwell-time. Unitary and whole-cell currents mediated by NMDA receptors composed of a cysteine-mutated NR1 subunit, NR1(C744A, C798A) were thus investigated. Dithiothreitol increased the open dwell-time and opening frequency of NR1(C744A, C798A)/NR2A receptors in a manner indistinguishable from that previously seen in wild-type channels. Marginal redox-induced changes in opening frequency of NR1(C744A, C798A)/NR2B receptors were noted. Redox modulation was completely abolished in NR1(C744A, C798A)/NR2C channels. Whole-cell recordings confirmed the single-channel results. Sulfhydryl reagents modulated NR1(C744A, C798A)/NR2A receptors identically to wild-type NR1/NR2A channels, whereas NR1(C744A, C798A)/NR2C receptors were insensitive to redox modulation. The oxidant 5,5'-dithio-bis-(2-nitrobenzoate) attenuated NR1(C744A, C798A)/NR2B receptor-mediated responses in a dithiothreitol-reversible manner. We conclude that cysteines 744 and 798 on the NR1 subunit are not involved in the redox modulation of NR1/NR2A receptors, but are crucial for the modulation of NR1/NR2C containing receptors. This suggests that the NR2A subunit is necessary and sufficient for the expression of redox sensitivity in NR1/NR2A channels. The slight, but measurable residual redox sensitivity of the mutant NR1(C744A, C798A)/NR2B receptors suggests the existence of an additional redox-sensitive site on NR2B. PMID- 10525102 TI - Variability in phenylephrine response and essential hypertension: a search for human alpha(1B)-adrenergic receptor polymorphisms. AB - Genetic polymorphisms in drug receptors, in particular adrenergic receptors, may contribute to intersubject differences in pharmacologic response. We tested patients and first-degree normotensive and hypertensive relatives of patients with essential hypertension and found substantial intersubject variability in blood pressure response to infusion of the alpha(1)-adrenergic agonist phenylephrine. Because response to phenylephrine depends upon interaction with alpha(1B)-adrenergic receptors, we tested whether polymorphisms in this receptor contribute to the variable responses. Accordingly, we developed a polymerase chain reaction-based method, generating four exon-spanning fragments, to identify polymorphisms in the coding sequence of the two exons of the human alpha(1B) adrenergic receptor. We sequenced the entire coding sequence of exon 1 from 51 subjects and exon 2 from 16 of these 51 subjects. Compared with the published sequence for the alpha(1B)-adrenergic receptor, we found one amino acid addition in exon 2 at position 368 (Arg) and one substitution (Arg371Gly) in all subjects. We thus suggest we have defined the correct coding sequence of the human alpha(1B) receptor. We found two "silent" polymorphisms in exon 1, one of which occurred in 3 of 51 subjects. These polymorphisms were unrelated to blood pressure status or response to phenylephrine. The 95% confidence intervals for expression of polymorphisms in exons 1 and 2 were 0 to 11%. Our data reveal that although phenylephrine response varies in humans, frequent polymorphisms in the coding sequence of the human alpha(1B)-adrenergic receptor appear not to account for this variation or for the increased blood pressure in patients with essential hypertension. PMID- 10525103 TI - Angiotensin-converting enzyme and matrix metalloproteinase inhibition with developing heart failure: comparative effects on left ventricular function and geometry. AB - The progression of congestive heart failure (CHF) is left ventricular (LV) myocardial remodeling. The matrix metalloproteinases (MMPs) contribute to tissue remodeling and therefore MMP inhibition may serve as a useful therapeutic target in CHF. Angiotensin converting enzyme (ACE) inhibition favorably affects LV myocardial remodeling in CHF. This study examined the effects of specific MMP inhibition, ACE inhibition, and combined treatment on LV systolic and diastolic function in a model of CHF. Pigs were randomly assigned to five groups: 1) rapid atrial pacing (240 beats/min) for 3 weeks (n = 8); 2) ACE inhibition (fosinopril, 2.5 mg/kg b.i.d. orally) and rapid pacing (n = 8); 3) MMP inhibition (PD166793 2 mg/kg/day p.o.) and rapid pacing (n = 8); 4) combined ACE and MMP inhibition (2.5 mg/kg b.i.d. and 2 mg/kg/day, respectively) and rapid pacing (n = 8); and 5) controls (n = 9). LV peak wall stress increased by 2-fold with rapid pacing and was reduced in all treatment groups. LV fractional shortening fell by nearly 2 fold with rapid pacing and increased in all treatment groups. The circumferential fiber shortening-systolic stress relation was reduced with rapid pacing and increased in the ACE inhibition and combination groups. LV myocardial stiffness constant was unchanged in the rapid pacing group, increased nearly 2-fold in the MMP inhibition group, and was normalized in the ACE inhibition and combination treatment groups. Increased MMP activation contributes to the LV dilation and increased wall stress with pacing CHF and a contributory downstream mechanism of ACE inhibition is an effect on MMP activity. PMID- 10525104 TI - Milameline (CI-979/RU35926): a muscarinic receptor agonist with cognition activating properties: biochemical and in vivo characterization. AB - Milameline (E-1,2,5,6-tetrahydro-1-methyl-3-pyridinecarboxaldehyde, O-methyloxime monohydrochloride, CI-979, PD129409, RU35926) was characterized in vitro and evaluated for effects on central and peripheral cholinergic activity in rats and rhesus monkeys. In muscarinic binding studies, milameline displayed nanomolar affinity with an agonist ligand and micromolar affinity with antagonist ligands, with approximately equal affinities determined at the five subtypes of human muscarinic receptors (hM(1)-hM(5)) with whole cells or membranes from stably transfected Chinese hamster ovary (CHO) cells. On binding, milameline stimulated phosphatidylinositol hydrolysis in hM(1) and hM(3) CHO cells and inhibited forskolin-activated cAMP accumulation in hM(2) and hM(4) CHO cells. Additionally, it decreased K(+)-stimulated release of [(3)H]acetylcholine from rat cortical slices. Responses were not caused by the inhibition of acetylcholinesterase, and there was no significant binding to approximately 30 other neurotransmitter binding sites. In rats, milameline decreased spontaneous and scopolamine-induced swimming activity, improved water-maze performance of animals impaired by basal forebrain lesions, increased cortical blood flow, decreased core body temperature, and increased gastrointestinal motility. Electroencephalogram activity in both rats and monkeys was characterized by a predominance of low voltage desynchronized activity consistent with an increase in arousal. Milameline also reversed a scopolamine-induced impairment of attention on a continuous-performance task in monkeys. Thus, milameline possesses a pharmacological profile consistent with that of a partial muscarinic agonist, with central cholinergic actions being produced in rats and monkeys at doses slightly lower than those stimulating peripheral cholinergic receptors. PMID- 10525105 TI - An unusual Ca(2+) entry pathway activated by protein kinase C in dog splenic artery. AB - The characteristics of the Ca(2+) entry pathways that are activated by protein kinase C (PKC) in canine splenic artery were investigated. Phorbol 12, 13 dibutyrate (PDB) contracted tissues and increased Ca(2+) influx. PDB-induced contraction was reduced by preincubation of tissues in Ca(2+)-free Krebs' solution (1 mM EGTA) but was unaffected when Ca(2+)-free solution was applied after contraction was initiated with PDB. In contrast, (45)Ca influx and contraction induced by PDB were resistant to nifedipine, Cd(2+), Gd(3+), La(3+), or Ni(2+) whether added before or during exposure to PDB. Indeed, Cd(2+) reduced (45)Ca(2+) efflux and potentiated Ca(2+) influx, but not PDB-induced contraction. Norepinephrine (NE)-induced contractions were inhibited by preincubation in Ca(2+)-free Krebs' solution (1 mM EGTA). Nifedipine (10 microM) led to a small reduction in the NE-induced contraction but was without effect on (45)Ca(2+) influx. Pretreatment for 16 min with Cd(2+), Gd(3+), or La(3+) (each 1 mM) reduced or abolished NE-induced contraction and Ca(2+) influx. Application of these cations after exposure to NE did not affect (45)Ca(2+) influx but reduced tension. The Q(10) for the increase in (45)Ca(2+) influx was approximately 2 for high K(+) and NE, but 4 for PDB. The results suggest that stimulation of PKC in dog splenic artery activates a Ca(2+) entry pathway that is resistant to di- and trivalent cations. The inhibition of Ca(2+) influx by preincubating with cations during short-term exposure to NE may represent an action on Ca(2+) turnover that precedes activation of PKC. PMID- 10525106 TI - Alpha(1)-adrenoceptor subtypes mediating inotropic responses in rat heart. AB - We studied the distribution of alpha(1)-adrenoceptor subtypes by radioligand binding assays using (125)I-labeled 2-beta(4-hydroxyphenyl)-ethylaminomethyl) tetralone (BE2254) and RNase protection assays, and determined the role of each subtype in mediating the inotropic response in rat heart. Chlorethylclonidine preincubation causes a approximately 72% decrease in the maximal binding capacity (B(max)). On the other hand, protection from phenoxybenzamine alkylation by 5 methyl-urapidil or BMY7378 decreased B(max) by 59 and 70%. By competitive inhibition, we have identified 19 to 28% and 30% high-affinity binding sites for the alpha(1A)- and alpha(1D)-selective antagonists in rat ventricles, with the alpha(1B)-adrenoceptor estimated as 45%. Consistent with the receptor-binding result, a similar distribution of mRNAs encoding alpha(1A), alpha(1B,) and alpha(1D) (22, 39, and 39%), based on RNase protection assays, was observed. In addition, we demonstrated that the noradrenaline response through alpha(1) adrenoceptor was antagonisted by 5-methyl-urapidil, RS-17053, BMY7378, and WB4101 in contraction functional experiments. K(I) values for the above compounds were defined for all three alpha(1)-adrenoceptor subtypes expressed in the human embryonic kidney 293 cell stably, and were further compared with the corresponding pA(2) values. Interestingly, the correlation was significantly higher for alpha(1A) (r(2) = 0.73) and alpha(1B) (r(2) = 0.66) than alpha(1D) (r(2) = 0.35) in these experiments. Because the potential of alpha(1D) measured to be 21% based on protection from phenoxybenzamine-caused inhibition by BMY7378, the combined potential of alpha(1A) and alpha(1B) can be estimated as approximately 80%. Taken together, these results suggest that the three alpha(1) adrenoceptor subtypes coexist in rat heart, with alpha(1A) and alpha(1B) playing a more prominent role in the positive inotropic response to noradrenaline. PMID- 10525107 TI - The third transmembrane helix of the cannabinoid receptor plays a role in the selectivity of aminoalkylindoles for CB2, peripheral cannabinoid receptor. AB - Two subtypes of the human cannabinoid receptor have been identified. The CB1 receptor is primarily distributed in the central nervous system, whereas the CB2 receptor is associated with peripheral tissue, including the spleen. These two subtypes are also distinguished by their ligand-binding profiles. The goal of this study was to identify critical residues in transmembrane region III (TM3) of the receptors that contribute to subtype specificity in ligand binding. For this purpose, a chimeric cannabinoid receptor [CB1/2(TM3)] was generated in which the TM3 of CB1 was replaced with the corresponding region of CB2. These receptors were stably expressed in Chinese hamster ovary cells for evaluation. The binding affinities of CB1/2(TM3) and the wild-type CB1 receptor to several prototype ligands were similar with one notable exception: the chimeric receptor exhibited a 4-fold enhancement in binding affinity to WIN 55,212-2 (K(d) = 4.8 nM) relative to that observed with CB1 (K(d) = 21.7 nM). Two additional aminoalkylindoles, JWH 015 and JWH 018, also bound the chimeric receptor (K(i) = 1.0 microM and 1.4 nM, respectively) with higher affinity compared with the wild-type CB1 (K(i) = 5.2 microM and 9.8 nM, respectively). Furthermore, the increase in binding affinities of the aminoalkylindoles were reflected in the EC(50) values for the ligand induced inhibition of intracellular cAMP levels mediated by the chimeric receptor. This pattern mirrors the selectivity of WIN 55,212-2 binding to CB2 compared with CB1. Site-specific mutagenesis of the most notable amino acid changes in the chimeric receptor, Gly195 to Ser and Ala198 to Met, revealed that the enhancement in WIN 55,212-2 binding is contributed to by the Ser but not by the Met residue. The data indicate that the amino acid differences in TM3 between CB1 and CB2 play a critical role in subtype selectivity for this class of compounds. PMID- 10525108 TI - Modification of cardiac Na(+) current by RWJ 24517 and its enantiomers in guinea pig ventricular myocytes. AB - We examined the effects of the cardiotonic agent RWJ 24517 (Carsatrin, racemate) and its (S)- and (R)-enantiomers on action potential duration, Na(+) current (I(Na)), and delayed rectifier K(+) current (I(K)) of guinea pig ventricular myocytes. RWJ 24517 (0. 1 and 1 microM) prolongation of action potential duration could not be accounted for by suppression of either the rapid (I(Kr)) or slow (I(Ks),) component of I(K), although RWJ 24517 did reduce I(Kr) at concentrations of 1 microM. A more dramatic effect of RWJ 24517 (0.1-1 microM) and the (S) enantiomer of RWJ 24517 (0.1-3 microM) was an increase in peak I(Na) and slowing of the rate of I(Na) decay, eliciting a large steady-state current. Neither RWJ 24517 nor the (S)-enantiomer affected the fast time constant for I(Na) decay, but both significantly increased the slow time constant, in addition to increasing the proportion of I(Na) decaying at the slow rate. Both agents elicited a use dependent decrease of peak I(Na) (3-10 microM), which probably resulted from a slowing of both fast and slow rates of recovery from inactivation. In contrast, the (R)-enantiomer of RWJ 24517 did not induce a steady-state component I(Na) or increase peak I(Na) up to 10 microM, but it decreased peak I(Na) at 30 microM. The (R)-enantiomer displayed little use-dependent reduction of I(Na) during trains of repetitive pulses and had no effect on rates of inactivation or recovery from inactivation. These actions of the racemate and the (S) stereoisomer to slow inactivation and to prolong both Na(+) influx and action potential duration may contribute to the positive inotropic actions of these agents because the resulting accumulation of intracellular Na(+) would increase intracellular Ca(2+) via Na(+)/Ca(2+) exchange. PMID- 10525109 TI - Species-dependent differences in monoamine oxidase A and B-catalyzed oxidation of various C4 substituted 1-methyl-4-phenyl-1,2,3, 6-tetrahydropyridinyl derivatives. AB - In an attempt to provide a better understanding of the scope and limitations of animal models used in some drug development programs and to further our understanding of potential metabolic bioactivation reactions, we have undertaken studies to profile the monoamine oxidase A and B (MAO-A and -B, respectively) activities in liver and brain mitochondrial preparations obtained from a variety of species using a series of 1-methyl-4-aryl-1,2,3, 6-tetrahydropyridinyl substrates. Mitochondrial preparations were incubated with substrates at 37 degrees C in the presence or absence of clorgyline, (R)-deprenyl, or a mixture of these two propargylamines to inhibit MAO-A, MAO-B, or both enzymes. The rates of formation of the corresponding dihydropyridinium metabolites were estimated spectrophotometrically. MAO-B was found to be the principal enzyme present in all tissues. Human liver displayed more MAO-A activity than the liver of any other species studied; subhuman primates displayed little or no detectable MAO-A activity. The properties of the preparations from rat liver were most similar to those from human liver with respect to the MAO-A/MAO-B ratios and the kinetic parameters of the four substrates used to profile enzymatic activity. The kinetic properties of mitochondrial preparations from bovine liver, a commonly used source of purified MAO-B preparations, were consistently different from all of the other species studied. The mitochondrial preparations from rabbit brain and liver also were unusual in that they displayed relatively low MAO activities. Additionally, these enzyme activities were considerably less susceptible to inhibition by clorgyline and (R)-deprenyl. Finally, an exceptionally low MAO-B liver/brain V(max)/K(m) ratio was observed with the mitochondria obtained from the C57BL/6 mouse, an effect that may contribute to the susceptibility of this strain to the toxic effects of the parkinsonian-inducing neurotoxin 1-methyl-4 phenyl-1,2,3, 6-tetrahydropyridine. PMID- 10525110 TI - Safety, pharmacokinetics, and tissue distribution of liposomal P-ethoxy antisense oligonucleotides targeted to Bcl-2. AB - Antisense oligonucleotides (oligos) have the ability to selectively block disease causing genes, thereby inhibiting production of disease-associated proteins. However, their effectiveness has been limited by their low intracellular delivery. We had previously demonstrated that liposomes could increase the intracellular uptake of P-ethoxy oligos, hydrophobic analogs of phosphodiesters, and that liposomal Bcl-2 P-ethoxy antisense oligos (L-Bcl-2) could selectively inhibit Bcl-2 protein production, thereby inducing growth inhibition in Follicular Lymphoma cell lines. To understand the in vivo behavior of L-Bcl-2, we conducted a series of studies to evaluate the safety, pharmacokinetics, and tissue distribution of i.v. injections of L-Bcl-2 in normal rodents. Daily administration of 20 mg of L-Bcl-2/kg of body weight in 5 consecutive days had no adverse effects on renal or hepatic functions, nor on hematological parameters. Histopathology also did not reveal any significant changes in the morphology of the organs studied. In rats, the area under the curve of L-Bcl-2 reflects a two compartment model of distribution with a biphasic plasma clearance. The T(1/2alpha) and T(1/2beta) were approximately 8 min and 4.2 h, respectively, and the V(d) was 79 ml, indicating a broad body distribution. The highest concentrations of L-Bcl-2 were found in spleen > liver > kidneys. These studies showed that in the schedules studied no significant toxicity associated with L Bcl-2 was observed over 6 weeks, and that L-Bcl-2 could be widely distributed in the body. PMID- 10525111 TI - Thioguanine administered as a continuous intravenous infusion to pediatric patients is metabolized to the novel metabolite 8-hydroxy-thioguanine. AB - Thiopurine antimetabolites have been in clinical use for more than 40 years, yet the metabolism of thiopurines remains only partially understood. Data from our previous pediatric phase 1 trial of continuous i.v. infusion of thioguanine (CIVI TG) suggested that TG was eliminated by saturable mechanism, with conversion of the drug to an unknown metabolite. In this study we have identified this metabolite as 8-hydroxy-thioguanine (8-OH-TG). The metabolite coeluted with the 8 OH-TG standard on HPLC and had an identical UV spectrum, with a lambda(max) of 350 nm. On mass spectroscopy, the positive ion, single quad scan of 8-OH-TG yielded a protonated molecular ion at 184 Da and contained diagnostic ions at m/z 167, 156, 142, and 125 Da. Incubation of TG in vitro with partially purified aldehyde oxidase resulted in 8-OH-TG formation. 8-OH-TG is the predominant circulating metabolite found in patients receiving CIVI-TG and is likely generated by the action of aldehyde oxidase. PMID- 10525112 TI - Beta-adrenoceptor subtype activities of trimetoquinol derivatives: biochemical studies on human beta-adrenoceptors expressed in chinese hamster ovary cells. AB - The beta-adrenoceptor activities of trimetoquinol (TMQ) isomers and selected derivatives were evaluated on human beta-adrenoceptor subtypes expressed in Chinese hamster ovary cells. In cAMP accumulation assays, (-)-TMQ was 214-, 281-, and 776-fold more potent than (+)-TMQ at stimulating beta(1)-, beta(2)-, and beta(3)-adrenoceptor subtypes, respectively. In radioligand binding assays, (-) TMQ exhibited 123-, 331-, and 5-fold greater affinity than (+)-TMQ for beta(1)-, beta(2)-, and beta(3)-adrenoceptor subtypes, respectively. (-)-TMQ and (+/-)-TMQ activated the human beta(3)-adrenoceptor with an 8.2- and 3.4-fold greater efficacy, respectively, than the reference beta-adrenoceptor agonist (-) isoproterenol (efficacy = 1). The 3',5'-diiodo analogs of TMQ were partial agonists of the beta(2)-adrenoceptor relative to (-)-isoproterenol, and their potencies were 5- to 10-fold higher at the beta(3)-adrenoceptor as compared with beta(1)-adrenoceptors. Modification of the catechol (6,7-dihydroxy) nucleus, such as replacement of the 7-hydroxy group with a chloro group (7-chloroTMQ), ring fluorination (8-fluoro and 5,8-difluoro analogs), or preparation of bioisosteric tetrahydrothiazolopyridine (THP) derivatives of TMQ yielded compounds that displayed partial agonist activity (relative to (-)-isoproterenol) or were inactive at the beta(2)-adrenoceptor and exhibited beta(3)-adrenoceptor-selective stimulation compared with the beta(1)-adrenoceptor. Furthermore, the 3',5'-diiodo 4'-methoxybenzylTHP derivative of TMQ was 65-fold more potent than the corresponding 3',4',5'-trimethoxybenzylTHP at the human beta(3)-adrenoceptor. Our results indicate that 6, 7-dihydroxy-catechol-modified and 1-benzyl halogen substituted derivatives of TMQ represent promising leads for the development of beta(3)-adrenoceptor-selective agonists. PMID- 10525113 TI - Nicotine infusion modulates immobilization stress-triggered induction of gene expression of rat catecholamine biosynthetic enzymes. AB - The relationship between nicotine and stress is complex and paradoxical. Although people claim they smoke because it relaxes them, nicotine can trigger some of the effects observed with stress, including the release and synthesis of the catecholamines and their biosynthetic enzymes. This study examined one aspect of this confusing relationship between nicotine and stress. Multiple injections of nicotine bitartrate (5 mg/kg) elevated mRNA levels for the catecholamine biosynthetic enzymes, tyrosine hydroxylase (TH), dopamine beta-hydroxylase (DBH), and phenylethanolamine N-methyltransferase, and of preproneuropeptide Y in rat adrenal medulla more than did 1 mg/kg of nicotine bitartrate. In the locus ceruleus, substantia nigra, and ventral tegmental area both doses equally induced TH mRNA levels. Nicotine infusion (15 mg/kg/day) did not affect adrenal mRNA levels for any of the genes of interest and did not increase plasma corticosterone levels. However, in rats pre-exposed to nicotinic infusions, the response to a single immobilization (IMO) stress was markedly attenuated with respect to changes in adrenomedullary TH, DBH, and phenylethanolamine N methyltransferase mRNA levels and in c-Fos protein levels. In the central nervous system, the chronic infusion of nicotine prevented the induction of TH mRNA by repeated IMO stress in the ventral tegmental area (but not in substantia nigra) and of DBH mRNA by single IMO in the locus ceruleus. These findings may explain some of the complex interactions between stress and exposure to nicotine. PMID- 10525114 TI - Cannabinoid receptors differentially modulate potassium A and D currents in hippocampal neurons in culture. AB - Cannabinoid (CB(1)) receptor activation produced differential effects on voltage gated outward potassium currents in whole-cell recordings from cultured (7-15 days) rat hippocampal neurons. Voltage-dependent potassium currents A (I(A)) and D (I(D)) were isolated from a composite tetraethylammonium-insensitive current (I(comp)) by blockade with either 4-aminopyridine (500 microM) or dendrotoxin (2 microM) and subtraction of the residual I(A) from I(comp) to reveal I(D). The time constants of inactivation (tau) of I(A) and I(D) as determined in this manner were found to be quite different. The CB(1) agonist WIN 55,212-2 produced a 15- to 20-mV positive shift in voltage-dependent inactivation of I(A) and a simultaneous voltage-independent reduction in the amplitude of I(D) in the same neurons. The EC(50) value for the effect of WIN 55,212-2 on I(D) amplitude (13.9 nM) was slightly lower than the EC(50) value for its effect on I(A) voltage dependence (20.6 nM). Pretreatment with either the CB(1) antagonist SR141716A or pertussis toxin completely blocked the differential effects of WIN 55,212-2 on I(A) and I(D), whereas cellular dialysis with guanosine-5'-O-(3-thio)triphosphate mimicked the action of cannabinoids but blocked the action of simultaneously administered cannabinoid receptor ligands. Finally, the differential effects of cannabinoids on I(A) and I(D) were both shown to be mediated via the well documented cannabinoid receptor inhibition of adenylyl cyclase and subsequent modulation of cAMP and protein kinase. These actions are considered in terms of cAMP-mediated phosphorylation of separate I(A) and I(D) channels and the contribution of each to composite voltage-gated potassium currents in these cells. PMID- 10525115 TI - Repeated measurement of intestinal permeability as an assessment of colitis severity in HLA-B27 transgenic rats. AB - We report on the development of a method for repeated monitoring of mucosal permeability that allows assessment of the severity of colitis and evaluation of treatment efficacy in HLA-B27 transgenic rats. We determined the extent to which intestinal permeability related to stool condition, colon weight, and histological pathology in precolitic and diseased rats up to 29 weeks old. Intestinal permeability was measured by the urinary excretion of iodixanol at 24 h after oral administration. Mean permeability values increased significantly with age in HLA-B27 rats but remained decreased in the background strain Fischer 344 (F-344) control animals. Macroscopic evaluation of HLA-B27 rat colons between 20 and 24 weeks old showed colonic thickening with colonic wet weights increased from 3.4+/-0.13 mg/kg b.wt. in F-344 rats to 6.79+/-0.73 mg/kg b.wt. (p<.05) in HLA-B27 rats. Histological examination of HLA-B27 rat colons confirmed the colonic inflammation as a chronic active mononuclear cell infiltrate. The increase in colon weight was associated with an increase in permeability: 1.16+/ 0.17 mg iodixanol versus 5.37+/-1.3 mg of iodixanol in F-344 and HLA-B27 rats, respectively. Three weeks treatment of HLA-B27 rats with cyclosporin A, but not sulfasalazine, showed a dose-dependent decrease in mucosal permeability and colon weight. Neither treatment improved stool condition. We conclude that the measurement of intestinal permeability by iodixanol excretion is a useful biochemical marker that is associated with increases in colonic weight and histological evaluation of inflammation. These data indicate that this technique may be valuable for diagnostic and evaluation purposes in preclinical models of inflammatory bowel disease. PMID- 10525116 TI - Spin trap (N-t-butyl-alpha-phenylnitrone)-mediated suprainduction of heme oxygenase-1 in kidney ischemia/reperfusion model: role of the oxygenase in protection against oxidative injury. AB - In mammals the rate-limiting step in heme catabolism is the heme oxygenase (HO) system. Two isozymes, HO-1 and HO-2, oxidatively cleave the substrate to form biliverdin, and the potential cellular messenger, CO; the chelated iron is released as the result of the tetrapyrrole ring opening. Biliverdin is subsequently reduced to bilirubin, an antioxidant, by biliverdin reductase. The aim of the present study was to investigate the involvement of HO-1, a heat shock/stress protein, in protection offered by the spin trap agent, N-tert-butyl alpha-phenyl-nitrone (PBN), against kidney ischemia/reperfusion injury. For this, HO-1 expression and assessment of the parameters associated with tissue-oxidative injury were compared in the presence or absence of PBN pretreatment of rats (100 mg/kg i.p., 30 min) before the onset of 30-min ischemia. Twenty-four hours after reperfusion, Northern blot analysis showed an unprecedented approximately 37-fold increase in 1.8-kb HO-1 mRNA in PBN pretreated rat kidney; HO-2 mRNA levels did not increase. At 48 h, the levels of HO-1 mRNA remained nearly 14-fold higher than the control value. In the absence of PBN, the levels measured approximately 5- and 2-fold higher than control values at the 24- and 48-h intervals, respectively. PBN pretreatment also resulted in a most impressive increase in the levels of HO-1 protein as judged by Western blot analysis and measurement of enzyme activity at the 24-h time point. As detected by immunohistochemical analysis, PBN pretreatment caused an increase in HO-1 and biliverdin reductase immunoreactive proteins in the cortex and in the outer stripe of the outer medulla. In the absence of PBN pretreatment, there was an intense immunostaining for HO-1 in the medullary rays, which corresponded with iron and lipid peroxidation staining of the region; these observations were not made with PBN pretreated kidneys. Collectively, the findings are consistent with the likelihood that suprainduction of HO-1 gene expression protects the kidney from free radical mediated injury by increasing the capacity to produce the potent cellular antioxidant bilirubin. We also suggest spin trap-mediated protection against ischemia/reperfusion injury is likely due to a sustained elevation of HO-1 gene expression by formation of stable radicals. PMID- 10525118 TI - F(2)-isoprostanes: sensitive and specific non-invasive indices of lipid peroxidation in vivo. AB - Isoprostanes are members of a complex family of lipids, isomers of the conventional enzymatically derived prostaglandins (PG), which are produced in vivo primarily, if not exclusively, by a free radical-catalyzed peroxidation of polyunsaturated fatty acids. Most of the work has been focused upon a group of isomers of the enzyme-derived PGF(2alpha), called F(2)-isoprostanes (F(2)-iPs). Because of their mechanism of formation, chemical stability and the rapid development of sensitive methods for their measurement, they have the attraction as non-invasive indices of oxidant stress in vivo. Altered generation of F(2)-iPs has been reported in a variety of clinical settings putatively associated with oxidant stress. These include atherosclerosis, chronic obstructive pulmonary disease and Alzheimer's disease. Furthermore, the measurement of specific F(2) iPs may provide a sensitive biochemical basis for rational dose-selection of natural and synthetic inhibitor of lipid peroxidation. Although F(2)-iPs possess biological activities in vitro and in vivo, much remains to be learned about their role and as mediators of the cellular effects of lipid peroxidation and oxidant stress. PMID- 10525117 TI - Regulation of aquaporin-2 expression by the alpha(2)-adrenoceptor agonist clonidine in the rat. AB - Aquaporin-2 (AQP-2), the major water channel responsible for water balance, has been shown to be regulated by the binding of vasopressin to V(2) vasopressin receptors in the medullary collecting duct. alpha(2)-Adrenoceptor agonists such as clonidine have been associated with an increase in free water clearance that was secondary to an inhibition of the ability of vasopressin to increase cAMP levels in the collecting ducts. This investigation focused on the possibility that this increase in free water clearance following administration of an alpha(2)-adrenoceptor agonist was associated with a reduction in medullary AQP-2 expression. In the anesthetized rat, clonidine increased urine flow rate (32+/-5 versus 137+/-16 microl/min, p<.05) and free water clearance (-58+/-6 versus 3+/-8 microl/min, p<.05) compared with the group receiving the saline vehicle infusion. The increase in free water clearance with clonidine administration was associated with a reduction in whole kidney AQP-2 mRNA levels (282+/-25 versus 216+/-11 A units, p<.05). This decrease in water reabsorption was associated with a redistribution of AQP-2 away from the luminal membrane of the medullary collecting duct to the cytosol. These effects were not secondary to changes in serum vasopressin levels, as these were similar in the vehicle control and clonidine groups (59+/-5 pg/ml versus 64+/-7 pg/ml, p = NS). The rapid redistribution of AQP-2 and the reduction in AQP-2 mRNA following clonidine administration are consistent with the hypothesis that the alpha(2) adrenoceptor regulates water excretion at least in part by effects on AQP-2. PMID- 10525119 TI - Impaired postprandial tissue regulation of blood flow in insulin resistance: a determinant of cardiovascular risk? AB - The insulin resistant state is a major risk factor for coronary artery disease. This increased risk is likely to be due to associated lipid and coagulation abnormalities rather than just abnormalities in glucose metabolism or hyperinsulinaemia alone. Exaggerated postprandial lipaemia is a well-recognised associate of insulin resistance and postprandial hypertriglyceridaemia is particularly important in the development of coronary atheroma. It seems likely that insulin is one of the hormonal regulators of adipose tissue and skeletal muscle blood flow. The reduced blood flow and blunting of the postprandial rise of peripheral blood flow in insulin resistance may decrease chylomicron triglyceride delivery to muscle in subjects with insulin resistance. This, in turn, will lead to increased production of atherogenic particles. We propose that impaired postprandial vasodilation, already recognised as a key feature of glucose intolerance, is also the cause of impaired lipid metabolism in insulin resistant subjects and predisposes them to cardiovascular disease. PMID- 10525120 TI - Contribution of apolipoprotein(a) size, pentanucleotide TTTTA repeat and C/T(+93) polymorphisms of the apo(a) gene to regulation of lipoprotein(a) plasma levels in a population of young European Caucasians. AB - Several studies indicate that the inter-individual variation in plasma concentrations of lipoprotein(a) (Lp(a)) is mainly under genetic control. To define the effect of three DNA polymorphisms on apolipoprotein(a) (apo(a)) expression, we have determined plasma Lp(a) concentrations, apo(a) isoform size, KpnI allele size, the TTTTA pentanucleotide repeat number in the 5' control region of the apo(a) gene and the +93 C/T polymorphism in a European Caucasian population. The simultaneous determination of the kringle 4 (K4) number by genotyping and by phenotyping revealed that the size distribution of non expressed apo(a) alleles was markedly skewed towards alleles with greater than 25 K4 repeats. This is consistent with the inverse relationship frequently described between the kringle 4 number and the plasma Lp(a) level. Apportioning the Lp(a) concentration from the surface of the peaks on apo(a) phenotyping blots, we have observed that the Lp(a) plasma concentration associated with alleles having more than 25 K4 units does not exceed 400 mg/l, whereas the range of Lp(a) concentrations associated with smaller alleles was broad, from 0 to more than 1000 mg/l. It can thus be concluded that the number of K4 repeats is the main determinant of Lp(a) concentration when this number is more than 25, whereas other polymorphisms may be involved in the alleles with fewer than 26 K4. Analyses of the TTTTA repeat number and of the +93 C/T polymorphism were performed in subjects with KpnI alleles of the same length: low Lp(a) concentrations were shown to be preferentially associated with the presence of apo(a) alleles with more than eight pentanucleotide repeats while no association was revealed between Lp(a) plasma levels and the C/T polymorphism. These results demonstrate that the (TTTTA)(n) polymorphism affects the Lp(a) expression independently of apo(a) size polymorphism. PMID- 10525121 TI - Endothelial cells regulate the proliferation of monocytes in vitro. AB - Monocytes (MPhis) are among the first cells to accumulate in early atherosclerotic lesions and generally are believed to be incapable of proliferation. However, recent studies indicate that the number of MPhis in atherosclerotic lesion may increase due to induction of local proliferation. Since proliferation of hematopoietic lineage cells is strongly influenced by interaction with neighboring cell types, we examined the ability of vascular endothelial cells (EC), smooth muscle cells or fibroblasts to stimulate MPhi proliferation. In this study, we show that only when seeded at high densities MPhis could proliferate in culture. However, when contact co-cultured with EC, MPhis proliferated at a higher rate (260% on day 6) than those cultured alone or co-cultured with smooth muscle cells or fibroblasts. Endothelial cells could stimulate the proliferation of MPhis even at non-proliferating densities. Only EC that were growth arrested or in lag phase could induce MPhi proliferation, whereas those in the exponential proliferating phase were non-stimulatory. Conditioned medium prepared from EC in growth arrested or lag phase failed to stimulate MPhi proliferation. Similarly physical separation of MPhis from EC also resulted in no proliferation. These results suggest that EC induced MPhi proliferation is contact dependent and no soluble factors are involved in this induction. This EC induced MPhi proliferation may have a profound effect on the rate of progression of atherosclerosis. PMID- 10525122 TI - Macrophage colony-stimulating factor reduces tert-butyl hydroperoxide induced oxidative injury to monocytes/macrophages. AB - The transformation of macrophages into foam cells is an important event in the development of atherosclerosis, and the oxidative injury caused by oxidized low density lipoprotein (Ox-LDL) plays an essential role in that process. It has been proved that macrophage colony-stimulating factor (M-CSF) could prevent the progression of atherosclerosis in Watanabe heritable hypercholesterolemic (WHHL) rabbits. We proposed that the anti-atherogenic effect of M-CSF was partly associated with its protective effect on monocyte-derived macrophages from Ox-LDL induced oxidative injury. In order to prove this, we investigated the effect of M CSF on the oxidative injury caused by tert-butyl hydroperoxide (tbOOH) to mouse peritoneal macrophages and U937/J774 cell lines. The results showed that M-CSF could protect mouse peritoneal macrophages from oxidative injury (presented by cell morphology and cell survival rate); L929 cell-conditioned medium (L929-CM) had the same effect as M-CSF; and anti-M-CSF monoclonal antibody could mostly block the protective effect of L929-CM on macrophages. L929-CM was proved to be also able to decrease the impact of plasma membrane fluidity in U937 and J774 cells treated with tbOOH. Incubation with tbOOH caused DNA fragmentation in U937 cells. The presence of L929-CM greatly reduced the number of apoptotic U937 cells characterized by DNA fragmentation. From these results, we concluded that M-CSF could protect monocytes/macrophages from oxidative injury. It may be one of the mechanisms which explain the anti-atherogenic effect of exogenous M-CSF in WHHL rabbits. PMID- 10525123 TI - Evidence for a cholesteryl ester donor activity of LDL particles during alimentary lipemia in normolipidemic subjects. AB - Postprandial hypertriglyceridemia represents an independent risk factor for coronary artery disease. In the postprandial state, elevated levels of triglyceride-rich lipoproteins (TRL) are minor acceptors of HDL-cholesteryl ester (CE) transferred by CETP in normolipidemic subjects: indeed, LDL particles represent the major CE acceptors. In order to evaluate further the potential atherogenicity of lipoprotein particles characteristic of the postprandial phase in normolipidemic subjects, we determined the quantitative and qualitative features of apoB- and apoAI-containing lipoproteins over an 8-h period following consumption of a mixed meal. During postprandial lipemia, we observed a significant decrease (-12%) in plasma AI concentration (138+/-4 and 156+/-4 mg/dl, at 3 h and baseline, respectively, P<0.005). Concomitantly, a progressive increase (+13%) was detected in HDL2 concentrations (138+/-7 mg/dl at 4 h vs. 122+/-12 mg/dl at baseline, P<0.005), as well as a significant reduction (-9%) in HDL3 levels (137+/-6 mg/dl at 3 h vs. 150+/-4 mg/dl at baseline; P<0.05). Additionally, plasma LDL was reduced by 5% (247+/-12 mg/dl at 3 h vs. 260+/-15 mg/dl at baseline; P<0.05) 3 h following meal intake. Moreover, a significant reduction (-10%) occurred in the CE/TG ratio in LDL at 2 h postprandially (8+/-2 at 2 h vs. 9+/-3 at baseline; P<0.005). These changes reflected an increment (17+/-3 mg/dl at 3 h vs. 15+/-4 mg/dl at baseline; P<0.05) in LDL triglyceride concentrations. Despite the high CE acceptor capacity of LDL particles, no measurable increase in their CE content was detected during the postprandial phase. We demonstrated that CE accepted by LDL particles from HDL are secondarily transferred to chylomicrons by CETP. As chylomicrons displayed a 260-fold lower CE/TG ratio than LDL (0.03:1 and 7.8:1 in chylomicrons and LDL, respectively), CE rich LDL may act to donate CE to chylomicrons. In conclusion, our data indicate that the presence of elevated levels of chylomicrons induces LDL to act as a secondary donor of CE during the postprandial phase. PMID- 10525124 TI - C-reactive protein (CRP) in cerebro-vascular events. AB - BACKGROUND AND PURPOSE: C-reactive protein (CRP) is a useful prognostic factor in coronary heart disease. It has not been previously studied in acute cerebro vascular events, which was the topic of the present study. METHODS: Patients admitted to the hospital for an acute cerebro-vascular event were prospectively investigated. C-reactive protein was determined nephelometrically. Infection or inflammation were excluded clinically and with an erythrocyte sedimentation rate <30 mm/h. Computed tomography or nuclear magnetic resonance imaging of the brain was performed. RESULTS: According to initial brain imaging and the clinical course the 138 patients were divided into five groups: 20 with transient ischemic attack, 20 with reversible neurological deficit lasting less than 2 weeks, 61 with completed stroke and restitution, 16 with stroke without restitution and 21 with cerebral hemorrhage. Median CRP values (range) were 3.2 (2.4-13.5), 3.3 (2.4 39.4), 4.2 (2.4-73. 4), 3.4 (3.2-44.0) and 3.5 (2.4-104.0 mg/l), respectively with no significant differences between groups in a non-parametric test (Kruskal Wallis). Risk factors for vascular disease in general and stroke in particular had no visible influence on CRP levels. No relationship was found between time interval since onset of symptoms and CRP measurement, suggesting that an acute cerebro-vascular event has little influence on CRP values. CONCLUSION: CRP is not a useful marker to predict the outcome of an acute cerebro-vascular event on hospital admission. This is in contrast to acute coronary events. PMID- 10525125 TI - The GPIIIa Pl(A) polymorphism in the progression of abdominal aortic atherosclerosis. AB - Glycoprotein IIIa is expressed in platelets as part of the fibrinogen receptor and also in vascular endothelium where it mediates smooth muscle cell proliferation. The association between the glycoprotein GPIIIa Pl(A) polymorphism and the stage of atherosclerosis in the abdominal aorta was studied in a prospective autopsy study series of 300 middle-aged men (33-69 years). The Pl(A) genotype was determined by RFLP-PCR. The stage of atherosclerosis in the abdominal aorta was determined by computer-assisted morphometry. Elevated, fibrous lesions were more frequently (P=0.05) found in the abdominal aortas of men with the Pl(A1) homozygous genotype compared to men with the A2 allele (OR 2.3; 95% CI 0.99-5.2). The area of complicated lesions was significantly greater in men with Pl(A2)-positive genotypes compared to A1 homozygotes. The association with complicated lesions was especially strong in men over 60 (P=0.002). These results suggest that Pl(A) polymorphism is involved in the progression of atherosclerosis in the abdominal aorta. The association of men possessing the Pl(A2) allele with slower development of fibrous lesions and with greater area of complicated lesions in the abdominal aorta may result from genotypic differences in the smooth muscle cell proliferation after slight injuries to the endothelium mediated by glycoprotein IIIa or from genotypic differences in platelet fibrinogen binding or both. PMID- 10525126 TI - Reduced progression of atherosclerosis in apolipoprotein E-deficient mice with phenylhydrazine-induced anemia. AB - Epidemiological and experimental studies suggest that circulating erythrocytes play a role in the incidence of coronary heart disease. We investigated the influence of phenylhydrazine (PHZ)-induced anemia on the formation of atherosclerotic lesions in apo E-deficient mice on regular chow and on a high fat, high-cholesterol diet during 10 weeks. The repeated doses of PHZ caused sustained anemia throughout the study, changes in the physical characteristics of erythrocytes and increased reticulocyte count. The lesions of the anemic animals were smaller than in the controls and this was even more evident in mice fed with the atherogenic diet. A positive correlation was found between circulating red blood cells at the end of the experiment and the area of aortic lesion. There was also a negative association between the lesion and the reticulocyte count. This reduced progression of atherosclerotic lesions is independent of nutritional status or the lipoprotein cholesterol distribution. The results suggest that mechanisms related to the number of circulating red blood cells may have a significant influence on the development of atherosclerosis. PMID- 10525127 TI - Lipoxygenase inhibition decreases neointimal formation following vascular injury. AB - Our aim was to assess the potential role of lipoxygenase (LO) products in neointimal formation following vascular injury. We investigated the effect of LO pathway inhibition, by phenidone, on the concentration of 12- and 5 hydroxyeicosatetraenoic acid (12- and 5-HETE) in rat whole blood and in aortic tissue. We also examined the effect of phenidone on myoneointimal formation in balloon-injured rat carotid arteries. Phenidone significantly decreases the concentration of HETEs in aortic tissue, and decreases neointimal size even though there is no difference in the BrdU index. These data indicate that the LO product participates in developing neointima following balloon-induced vascular injury, and that the LO blocker phenidone decreases neointimal size possibly by suppressing migration of smooth muscle cells. PMID- 10525128 TI - Low density lipoprotein (LDL) binding affinity for the LDL receptor in hyperlipoproteinemia. AB - We measured the binding affinity of low density lipoprotein (LDL) for the LDL receptor in patients with various types of hyperlipoproteinemia and investigated the effects of LDL lipid composition and particle size on receptor affinity. LDL (1.019 < d < 1.063) was isolated by sequential ultracentrifugation from the serum of normolipidemic controls and patients with hyperlipoproteinemia. Patients with type IIa hyperlipoproteinemia had LDL with a similar receptor affinity to that of normal LDL. However, patients with hypertriglyceridemia (type IIb and type IV hyperlipoproteinemia) had LDL with a low receptor affinity, and the degree of the reduction in affinity paralleled the severity of the hypertriglyceridemia. The LDL of hypertriglyceridemic patients was rich in protein and triglycerides, had a low content of cholesterol and phospholipids, and was smaller than normal, thus resembling the atherogenic lipoprotein known as small, dense LDL. These abnormalities were observed even in patients with mild hypertriglyceridemia regardless of their serum cholesterol levels. The degree of alteration in LDL lipid composition and particle size was strongly associated with the reduction of LDL receptor affinity. We also examined the effects of two lipid-lowering agents (bezafibrate and probucol) on the characteristics of LDL. LDL receptor affinity was only improved when the lipid composition and particle size were normalized by drug therapy. Although it has been reported that decreased cholesteryl ester transfer protein (CETP) activity results in the formation of small LDL, plasma CETP activity was normal in the hyperlipoproteinemic patients and the normalization of LDL characteristics by drug therapy was not accompanied by an increase of CETP activity. Our results suggested that an abnormal lipid composition and/or small particle size might cause a decrease in the receptor affinity of LDL. These structural and functional abnormalities were reversed by drug therapy, underlining the importance of treating hypertriglyceridemia for the prevention of atherosclerosis. PMID- 10525129 TI - Dietary corn oil versus olive oil enhances HDL protein turnover and lowers HDL cholesterol levels in hamsters. AB - We studied the effect of dietary olive and corn oil on high-density lipoprotein (HDL) metabolism in golden Syrian hamsters. The animals were fed a semipurified diet containing 0.1% cholesterol and 40 energy % in the form of either olive or corn oil for a period of nine weeks. Hamsters fed corn oil had significantly lower very-low density and low-density lipoprotein (VLDL+LDL) cholesterol concentrations than those fed olive oil (0.98+/-0.24 vs. 1.40+/-0.34 mmol/l, means+/-S.D., n = 12), as well as significantly lower HDL cholesterol concentrations (3.31+/-0.50 vs. 3.91+/-0.12 mmol/l). The binding capacity of 125I labelled HDL to liver membranes was 33% higher in the hamsters fed corn oil instead of olive oil (571+/-29 vs. 429+/-24 ng HDL protein/mg membrane protein, P<0.05, n = 4). HDL protein kinetics were studied with 125I-HDL using a constant infusion technique. Both HDL fractional catabolic rate (0.255+/-0. 058 vs. 0.121+/-0.023 /h, P<0.01, n = 5) and transport rate (2.386+/-0. 753 vs. 1.218+/ 0.101 mg/h, P<0.01, n = 5) were about 2-fold higher in the hamsters fed corn oil. The rate of plasma cholesterol esterification by lecithin: cholesterol acyltransferase (LCAT) was essentially the same for the two diets. It is concluded that the low HDL level in the hamsters fed corn oil diets is linked with increased HDL binding and degradation in the liver and possibly other tissues. Due to increased HDL protein turnover, the capacity for reverse cholesterol transport is increased in hamsters fed corn oil despite the relative low HDL concentrations PMID- 10525130 TI - Oxidized-LDL induce apoptosis in HUVEC but not in the endothelial cell line EA.hy 926. AB - We studied the cytotoxic effect of copper-oxidized LDL in human primary human umbilical vein endothelial cells (HUVEC) and the immortalized EA.hy 926 cell line. Copper oxidized LDL (50-200 microg apoB/ml) induced concentration-dependent apoptotic cell death in HUVEC but did not induce apoptosis in EA.hy 926 cells. Only necrotic EA.hy 926 cells were evidenced at all copper oxidized LDL concentrations (25-200 microg apoB/ml), oxidation states (lightly, moderately and extensively copper-oxidized LDL) and incubation periods (4, 8 and 20 h). The different mechanisms of cell death induced by copper-oxidized LDL in EA.hy 926 cells and HUVEC may be related to various factors such as cytokines. In this study, we investigated whether interleukin-8 may be implicated in this process. The interleukin-8 production was increased in EA.hy 926 cells but not in HUVEC incubated with oxidized LDL. This increase in EA.hy 926 cells was associated with necrosis but not apoptosis. Nevertheless, the addition of interleukin-8 to HUVEC did not inhibit apoptosis induced by oxidized LDL. As the lower antioxidant capacity of EA.hy 926 cells results in higher sensitivity to oxidized LDL cytotoxicity (as we previously described), the redox status of cells may also control the form of endothelial cell death. In atherosclerotic lesions, the formation of apoptotic endothelial cells may result in part from the induction by oxidized LDL. PMID- 10525131 TI - The rebound of lipoproteins after LDL-apheresis. Effects on chemical composition and LDL-oxidizability. AB - The changes in low density lipoprotein (LDL) composition and oxidizability after LDL-apheresis (LA) using dextran sulfate cellulose columns were evaluated in 12 hypercholesterolemic men (mean+/-S.D. total cholesterol (TC) 9.7+/-1.8 mmol/l). After 10-20 months on biweekly LA combined with simvastatin 40 mg per day immediate pre-apheresis levels of TC, LDL-cholesterol, and apolipoprotein B were decreased to 5.3+/-1.3 mmol/l, 3.3+/-1.2 mmol/l, and 1.6+/-0.4 g/l, respectively, whereas apheresis induced mean acute reductions of 61, 78, and 76%, respectively. Measurements of copper-induced LDL-oxidizability in vitro showed an increased resistance against oxidation after LA until day 3 post-treatment: lag time (min) (day 0 (before LA) versus day 1 (post-LA)) 112+/-27 versus 130+/-26 (P=0.001), maximal rate of diene production (nmol/min per mg LDL) 11.1+/-2.7 versus 9.1+/ 2.1 (P=0.001), and time to maximal diene production (min) 186+/-39 versus 209+/ 35 (P=0. 001). Analysis of the chemical composition of LDL revealed a 25% (P<0.001) reduced content of cholesteryl esters and a decrease of the cholesterol to protein ratio of 1.20+/-0.25 to 0.70+/-0.22 (P<0. 001) through the 3rd day post-LA. Linoleic acid and arachidonic acid content of LDL decreased 11 and 18%, respectively, at the expense of palmitic acid. Vitamin E levels (mg/l) were significantly lowered due to reduction of the lipoprotein pool by apheresis; however, vitamin E content of LDL did not change in the days after apheresis when expressed per g protein or per micromol linoleic acid. The changes in fatty acid pattern were strongly associated with changes in LDL-oxidizability indices (P/=200 microM, decreased heart rates at >/=10 microM, and decreased somite and protein values at 500 microM. Defects involved cranial neural tube, optic vesicle, heart, and somites. A malformation rate of 59% in embryos exposed to 110 microg/ml tolbutamide was reduced to 13% by adding 1 microM cromakalim to the culture medium. Heart rate, somite number, and protein values were also improved by combined exposure to cromakalim and tolbutamide compared with exposure to tolbutamide alone. These results support previous findings with diazoxide (K(+) channel opener) and chlorpropamide (sulfonylurea) and further suggest a potential role for K(ATP) channel effects in sulfonylurea-induced dysmorphogenesis. PMID- 10525204 TI - Laterality patterns in infants with external birth defects. AB - The lateral distribution of external birth defects has not been reported in a comprehensive way, and patterns in this distribution have not been examined. This study presents the lateral distribution of 6,390 unilateral defects from among 102 defect categories in data collected by the Metropolitan Atlanta Congenital Defects Program. Among all defects, 49% (95% CI 48-51%) were right-sided. Among males and females, 51% (95% CI 50-53%) and 47% (95% CI 46-49%) of the defects, respectively, were right-sided. Of the 102 defect types, 57 had an excess of defects on the right side of the body; 39 had an excess of defects on the left side; and 6 were equally distributed. The excess on the right side was statistically significant for inguinal hernia, incarcerated inguinal hernia, microtia, preauricular sinus, talipes calcaneovalgus, and lambdoidal craniosynostosis. For the left side, the excess was statistically significant for preauricular tags, cleft lip, fused lip and cleft gum, cleft lip with cleft palate, congenital hip dysplasia, unstable hip, absent forearm or hand, anomaly of the knee, and skin tags. The percentage of right-sided defects among case subjects with unilateral defects was correlated with the percentage of males among all case subjects (r = 0.24, P < 0.05). Among male case subjects with unilateral defects, the correlation coefficient was 0.31 (P < 0. 01), and among females with unilateral defects, it was 0.11 (P > 0. 10). Differences in the lateral distribution of specific birth defects may be due to subtle differences in morphogenesis on the left and right sides of the embryo brought about by establishment of left-right asymmetry prior to organogenesis. The fact that more defect categories were right-sided than left-sided may be related to the observation that mitochondrial maturation in rat embryos is delayed on the right side. The right side, therefore, may be more susceptible than the left to defects caused by prenatal hypoxia. The significant correlation between the percentage right-sided and percentage male may then also be related to the observation that male sex hormones lower the mitochondrial respiration rate in rats and increase rat sensitivity to chemical hypoxia. Investigators should consider reporting the laterality of specific defects in both laboratory and epidemiological studies of birth defects. Right- and left-sided defects should perhaps be considered separately in etiologic studies of birth defects. Teratology 60:265-271, 1999. Published 1999 Wiley-Liss, Inc. PMID- 10525205 TI - Anatomy of a duplicated human foot from a limb with fibular dimelia. AB - At birth, a patient presented with a right lower limb featuring preaxial polydactyly and fibular dimelia with a complete absence of the tibia. Radiographic studies of the patient's foot revealed a duplicated tarsus with eight metatarsals and toes. The three preaxial toes were surgically removed at 1 year of age. A hallux and four normal-appearing postaxial toes remained. The foot was amputated when the patient was 3 years old. Dissection of the amputated foot revealed that the muscles of the dorsum were normal, except that the tendon of the extensor hallucis brevis muscle inserted into both the hallux and toe 2, rather than only into the hallux. The few abnormalities observed among the muscles on the plantar surface of the foot included absence of the insertions of the tibialis posterior and the abductor hallucis muscles. In addition, the two heads of the adductor hallucis muscle inserted abnormally into the medial (tibial) side of metatarsal 1, rather than into the lateral side. These various muscular anomalies, in addition to the mirror duplication of the foot with the presence of only a single metatarsal 1, leads us to propose that this metatarsal probably represents two lateral (fibular) halves that form a laterally duplicated bone. Although the dorsalis pedis artery was present on the dorsal surface of the foot, most of its derivatives were absent. This artery did give rise to a supernumerary medial branch that ended abruptly in the connective tissue (presumably postsurgical scar) at the medial border of the foot. This branch may have represented a duplicated dorsalis pedis artery associated with the duplicated preaxial portion of the foot. The arteries on the plantar surface of the foot were normal. Even though some anomalies in the pattern of the cutaneous innervation were observed, the nerves of the foot were largely normal. The gross and radiographic anatomy of this specimen and the radiographic anatomy of the leg suggest that some teratogenic event occurred when developmental specification reached the level of the future knee. The teratogenic event, which probably occurred early in the fifth week of development, may have caused damage that led to a lateral duplication of both the leg and the foot with the absence of some of the most medial structures. Teratology 60:272-282, 1999. PMID- 10525206 TI - Comparative effects of single intraperitoneal or oral doses of sodium arsenate or arsenic trioxide during in utero development. AB - Numerous studies have suggested that single-day intraperitoneal (IP) injection of inorganic arsenic results in failure of neural tube closure and other malformations in rats, hamsters, and mice. Most of these studies involved treatment of limited numbers of animals with maternally toxic doses of arsenic (generally As(V)), without defining a dose-response relationship. In the present Good Laboratory Practice-compliant study, sodium arsenate (As(V)) was administered IP and arsenic trioxide (As(III)) was administered either IP or orally (by gavage) on gestational day 9 to groups of 25 mated Crl:CD(R)(SD)BR rats. Only at dose levels that caused severe maternal toxicity, including lethality, did IP injection of arsenic trioxide produce neural tube and ocular defects; oral administration of higher doses of arsenic trioxide caused some maternal deaths but no treatment-related fetal malformations. In contrast, IP injection of similar amounts of sodium arsenate (based on the molar amount of arsenic) caused mild maternal toxicity but a large increase in malformations, including neural tube, eye, and jaw defects. In summary, neural tube and craniofacial defects were observed after IP injection of both As(V) and As(III); however, no increase in malformations was seen following oral administration of As(III), even at maternally lethal doses. These results demonstrate that the frequently cited association between prenatal exposure to inorganic arsenic and malformations in laboratory animals is dependent on a route of administration that is not appropriate for human risk assessment. PMID- 10525207 TI - Mini-review: toward understanding mechanisms of genetic neural tube defects in mice. AB - We review the data from studies of mouse mutants that lend insight to the mechanisms that lead to neural tube defects (NTDs). Most of the 50 single-gene mutations that cause neural tube defects (NTDs) in mice also cause severe embryonic-lethal syndromes, in which exencephaly is a nonspecific feature. In a few mutants (e.g., Trp53, Macs, Mlp or Sp), other defects may be present, but affected fetuses can survive to birth. Multifactorial genetic causes, as are present in the curly tail stock (15-20% spina bifida), or the SELH/Bc strain (15 20% exencephaly), lead to nonsyndromic NTDs. The mutations indicate that "spina bifida occulta," a dorsal gap in the vertebral arches over an intact neural tube, is usually genetically and developmentally unrelated to exencephaly or "spina bifida" (aperta). Almost all exencephaly or spina bifida aperta of genetic origin is caused by failure of neural fold elevation. The developmental mechanisms in genetic NTDs are considered in terms of distinct rostro-caudal zones along the neural folds that likely differ in mechanism of elevation. Failure of elevation leads to: split face (zone A), exencephaly (zone B), rachischisis (all of zone D), or spina bifida (caudal zone D). The developmental mechanisms leading to these genetic NTDs are heterogeneous, even within one zone. At the tissue level, the mutants show that the mechanism of failure of elevation can involve, e.g., (1) slow growth of adjacent tethered tissue (curly tail), (2) defective forebrain mesenchyme (Cart1 or twist), (3) defective basal lamina in surface ectoderm (Lama5), (4) excessive breadth of floorplate and notochord (Lp), (5) abnormal neuroepithelium (Apob, Sp, Tcfap2a), (6) morphological deformation of neural folds (jmj), (7) abnormal neuroepithelial and neural crest cell gap-junction communication (Gja1), or (8) incomplete compensation for a defective step in the elevation sequence (SELH/Bc). At the biochemical level, mutants suggest involvement of: (1) faulty regulation of apoptosis (Trp53 or p300), (2) premature differentiation (Hes1), (3) disruption of actin function (Macs or Mlp), (4) abnormal telomerase complex (Terc), or (5) faulty pyrimidine synthesis (Sp). The NTD preventative effect of maternal dietary supplementation is also heterogeneous, as demonstrated by: (1) methionine (Axd), (2) folic acid or thymidine (Sp), or (3) inositol (curly tail). The heterogeneity of mechanism of mouse NTDs suggests that human NTDs, including the common nonsyndromic anencephaly or spina bifida, may also reflect a variety of genetically caused defects in developmental mechanisms normally responsible for elevation of the neural folds. PMID- 10525208 TI - Teratogen update: thalidomide: a review, with a focus on ocular findings and new potential uses. PMID- 10525209 TI - Assessment of breast cancer size: sonographic and pathologic correlation. AB - PURPOSE: Accurate presurgical assessment of tumor size in breast cancer is important for choosing appropriate treatment. We retrospectively compared presurgical sonographic measurements of tumor size with postsurgical measurements of size and other variables. METHODS: In 174 cases, tumor size was measured by sonography before surgery, and those measurements were compared with values obtained by histopathologic examination of the specimens. The histologic type and grade, the number of lesions, and the presence of an extensive intraductal component also were considered in the intramodal correlations of tumor size. RESULTS: Sonographic measurements of tumor size correlated well with size measured after surgery (r = 0.72; 95% confidence interval, 0.64-0.78). The correlation was higher for lesions of 20 mm or less in their longest diameter than for larger lesions. The intramodal size correlation was lower for tumors with an extensive intraductal component than for tumors without an extensive intraductal component. The sonographic versus pathologic correlation of tumor size was less accurate when several lesions were present. CONCLUSIONS: Sonography is useful for presurgical assessment of tumor size in patients with breast cancer, especially for single lesions of 20 mm or less and without an extensive intraductal component. PMID- 10525210 TI - Detection and diagnosis of parathyroid incidentalomas during thyroid sonography. AB - PURPOSE: The aim of our study was to evaluate the incidence of incidentally found parathyroid adenomas (incidentalomas) in patients undergoing sonography of the neck for thyroid disease. METHODS: A total of 1,686 patients (305 men and 1,381 women) underwent sonography of the neck; the mean age was 49.6 +/- 21.7 years. In 38 patients (2.3%; 7 men and 31 women) with a mean age of 48.7 +/- 14.7 years, hypoechoic, homogeneous, oval nodules (mean volume, 1.0 +/- 0. 9 cm(3)) adjacent to the thyroid parenchyma were observed. All these lesions, compatible with the shape of an enlarged parathyroid gland, underwent ultrasound-guided fine-needle aspiration biopsy (FNAB), with measurement of parathyroid hormone (PTH) and thyroglobulin (Tg) levels in the needle washings (FNAB-PTH and FNAB-Tg). Biochemical screening for hyperparathyroidism was also performed. RESULTS: Cytologic examination plus FNAB-PTH/FNAB-Tg measurements revealed the presence of cellular material consistent with parathyroid tissue in 9 patients (24%), thyroid tissue in 22 patients (58%), and lymphoid tissue in 4 patients (11%). A tissue diagnosis was not established in 3 patients (8%). Five of 9 patients with parathyroid enlargement had high serum PTH and calcium levels. CONCLUSIONS: Enlarged parathyroid glands may be incidentally discovered during sonography of the thyroid. In patients with thyroid disease, the positive-predictive value of sonography in the identification of parathyroid tissue was low. Ultrasound-guided FNAB-PTH determination should be carried out when parathyroid adenoma is suspected. The incidental finding of an enlarged parathyroid may or may not be associated with yet undiagnosed hyperparathyroidism. PMID- 10525211 TI - Hydrogastric sonography in the preoperative staging of gastric cancer. AB - PURPOSE: Depth of wall invasion is the main prognostic factor in gastric cancer. We studied the utility of hydrogastric sonography in the evaluation of transmural infiltration by gastric cancer. METHODS: Thirty-seven patients with gastric adenocarcinoma were examined before surgery with a 5-MHz probe after the ingestion of 100-400 ml of water (mean, 330 ml). Sonographic results were compared with pathologic classifications obtained after surgery. RESULTS: Of the 37 tumors, 15 were found at surgery to be in the antrum, 10 were in the gastric body, 5 were proximal, and 7 were diffuse. After surgery, tumors were classified as follows: 2 (5%) T1, 4 (11%) T2, 15 (41%) T3, and 16 (43%) T4. Hydrogastric sonography correctly classified 30 (81%) of the 37 tumors. Sonography was correct for 2 (100%) of the 2 T1 tumors, 2 (50%) of the 4 T2 tumors, 13 (87%) of the 15 T3 tumors, and 13 (81%) of the 16 T4 tumors. Five sonographic errors were due to understaging and 4 to overstaging. With regard to tumor site, sonographic results were correct for 4 (57%) of the 7 diffuse tumors, 3 (60%) of the 5 proximal tumors, 9 (90%) of the 10 gastric body tumors, and 14 (93%) of the 15 antral tumors. CONCLUSIONS: Hydrogastric sonography is useful for preoperative evaluation of transmural infiltration by gastric cancers, particularly tumors in the antrum or gastric body. PMID- 10525212 TI - Transabdominal sonography of gastroesophageal junctions. AB - PURPOSE: We compared transabdominal sonography with upper gastrointestinal tract x-ray series (barium study) for evaluating gastroesophageal junction disease. METHODS: Fifty-five patients underwent barium study and sonography, which were performed independently by 2 radiologists. The results were compared. Normal findings were verified by esophagoscopy or by clinical follow-up; all abnormal findings were verified by biopsy, surgery, or manometry. RESULTS: Findings from barium study and sonography agreed in all 30 of the normal cases. On sonography, normal gastroesophageal junctions had multiple layers of different echogenicities (mean wall thickness, 4.9 mm); the 20 cancer cases all appeared as a mass-like thickening (mean, 14.9 mm) on sonography. Barium study findings were misinterpreted as achalasia in 2 cancer cases. One benign stricture was misinterpreted as cancer by both sonography and barium study. Of the 4 cases of achalasia, 3 were revealed by sonography as normal gastroesophageal junctions with proximal dilatation. CONCLUSIONS: Transabdominal sonography is useful for revealing the extramucosal component of gastroesophageal junction disease. The modality is especially useful for distinguishing between achalasia and infiltrative cancer when barium study shows smooth circumferential narrowing. PMID- 10525213 TI - Sonographic investigation of flow patterns in the perfused human placenta and their modulation by vasoactive agents with enhanced visualization by the ultrasound contrast agent Albunex. AB - PURPOSE: Our objective was to demonstrate sonographically the flow distribution in the circulation of human placentae as well as the sensitivity of the human fetal capillary bed to vasoconstriction and dilatation. METHODS: Five human full term placental lobules were maintained in vitro with fetal and maternal flow. Commercial ultrasound scanners were used for imaging. Albunex (1 ml bolus) was administered to the fetal "artery" to monitor patterns of flow. U46619 (1 ml, 10( 6) M; a thromboxane agonist and potent vasoconstrictor) and/or nitroglycerin (a potent vasodilator) were added to the fetal artery. RESULTS: Following the addition of U46619, mean "fetal pressures" rapidly rose from 23.2 +/- 0.8 to 118 +/- 2. 9 mm Hg (mean +/- standard error of mean; p < 0.001); venous flow rates decreased. As demonstrated by color Doppler imaging, flow markedly changed from a pattern of general distribution throughout the lobule to flow only near the chorionic plate. Color persistence was 94.4 +/- 6.5 seconds with Albunex after nitroglycerin and 39.8 +/- 3.4 seconds with Albunex after injection of U46619 (p < 0.001). Nitroglycerin had no effect when injected by itself but returned "constricted" flow to a "normal" pattern when injected after U46619. CONCLUSIONS: The contrast medium Albunex improved visualization of the fetal circulation throughout the lobule. Flow in the human placental capillary bed can be regionally manipulated throughout the placental lobule by vasomodulators and monitored by Albunex-enhanced sonographic examination. PMID- 10525214 TI - Sonographic spectrum of pelvic vascular malformations in women. PMID- 10525215 TI - Sonography in the diagnosis of rhabdomyolysis. AB - This case report describes the use of musculoskeletal sonography in the diagnosis of rhabdomyolysis. The case involved an episode of severe muscle lysis following a heroin overdose in an addict who lay comatose for an uncertain period. Sonography revealed multiple hyperechoic areas within the muscles examined, consistent with a recent injury. The clinical diagnosis of rhabdomyolysis may be difficult but is important in view of the attendant danger of acute renal failure, and sonography was instrumental in the diagnosis in this case. PMID- 10525216 TI - Symptomatic wandering accessory spleen in the pelvis: sonographic findings. AB - We describe the case of a mobile left lower quadrant mass associated with recurrent abdominal pain; at surgical exploration, the mass was found to be an accessory pelvic spleen. Although accessory spleens are present in 10-30% of individuals, a wandering accessory spleen located in the pelvis is not frequently seen. On sonography, the mass in our patient appeared well-defined and homogeneous. Spectral analysis and color Doppler imaging demonstrated a normal vascular branching pattern and high diastolic flow due to low resistance in the vascular bed. The parenchymal resistance index of the mass was similar to that of the native spleen. PMID- 10525217 TI - Sonographic findings in abdominal hereditary angioedema. AB - Patients with hereditary angioedema (HAE) may suffer from abdominal pain severe enough to prompt unnecessary surgical intervention. The diagnostic approach to abdominal pain during HAE attacks is not established. We describe abdominal sonographic findings during severe colic in 2 patients with known HAE. Sonography demonstrated marked mucosal thickening and edema of the bowel wall with a variable amount of free peritoneal fluid. These findings are not specific but are consistent with the hypothesized mechanism of attack and resolve after therapy. Abdominal sonography is useful for evaluating acute abdominal pain in patients with known HAE to prevent unnecessary surgery. Conversely, if the described sonographic findings appear in a case of abdominal colic of unknown origin, HAE should be included in the differential diagnosis. PMID- 10525218 TI - Endometrial carcinoma presenting as hematometra mimicking a large pelvic cyst. AB - Large pelvic cysts are commonly seen in gynecologic practice; their heterogeneous origin is reflected in their pleomorphic clinical features. We report the case of a 64-year-old multiparous postmenopausal woman with an unusual manifestation of endometrial adenocarcinoma that presented as hematometra mimicking a large pelvic cyst. In this case, hematometra was well demonstrated by transabdominal sonography, but transvaginal sonography allowed better visualization of the endometrial lining and suggested the correct diagnosis of endometrial cancer. Abnormal vaginal bleeding or hematometra in postmenopausal women should lead to assessment of the endometrial mucosa. Transvaginal sonography can be used to visualize neoplastic lesions in the endometrium when hematometra is detected through transabdominal sonography. PMID- 10525219 TI - Gallstones in sickle cell anemia. PMID- 10525221 TI - Sharp minds and blurred images. PMID- 10525220 TI - IVUS analysis of the acute and long-term stent result using motorized pullback: intraobserver and interobserver variability. AB - Intravascular ultrasound imaging has become an established method for analysis of intra-coronary stents. We analyzed the reproducibility of morphometric measurements immediately and late after stent implantation and the variability in the selection of predefined sites during motorized catheter pullback. Fifty consecutive patients were investigated immediately and 6 months after Palmaz Schatz stent implantation (motorized catheter pullback 0.5 mm/sec; 2.9 Fr; 30-MHz transducer). Two experienced investigators independently identified the proximal and distal reference, stent inlet, stent outlet, and the minimal in-stent area in each imaging run. The longitudinal distance between corresponding measurement sites was calculated. Lumen, stent, and vessel area were assessed by planimetry, mean difference was calculated. Long-term reproducibility was analyzed by comparison of measurements made at predefined sites within the stent, immediately and late after implantation. Observer agreement in identification of predefined measurement sites was high. Longitudinal distance between corresponding measurement sites was low and pronounced for the minimal in-stent lumen area. Variabilities for the intra- and interobserver comparison were similar. Values for interobserver comparison were given in brackets. Acute after stent implantation, the variability for the reference proximal was 4.9% (0.4%), distal 1.0% (-4.2%), minimal in-stent lumen -0.5% (1.3%). At follow-up, variability for the reference proximal was -11.0% (-2.2%), distal -1.0% (-2.3%), minimal in-stent lumen 1.9% (6.1%). Long-term reproducibility for the proximal stent inlet was 2.7% (observer 1) and -0.4% (observer 2), for the distal stent outlet 1.3% (observer 1), -3.0% (observer 2), respectively. IVUS investigations with motorized IVUS pullback in stented coronary segments show a low intra- and interobserver variability, both immediately and late after stent implantation. Absolute and relative area differences are low. Long-term reproducibility of measurements within predefined stent sites was high. Motorized catheter pullback guarantees high reliability of IVUS measurements and should be routinely used for clinical IVUS studies. PMID- 10525222 TI - Risk predictors in patients scheduled for percutaneous coronary revascularization. AB - Traditionally, procedural risks associated with conventional balloon coronary angioplasty have been largely attributed to unfavorable lesion morphology. However, factors predicting adverse events in the current practice of percutaneous coronary revascularization are unclear. The present study was undertaken to determine factors predicting major adverse events (death or Q-wave myocardial infarction or emergency bypass surgery) in 3,335 consecutive patients undergoing percutaneous coronary revascularization in the current practice of percutaneous coronary revascularization. During the period of observation, the rate of lesions treated successfully increased from 91% to 95% (P < 0.0001), whereas the rate of major adverse events (MACE) decreased from 3.6% to 1.6% (odds ratio [OR], 0.70 per year). Using multiple stepwise logistic regression analysis, cardiogenic shock (OR, 8.59; confidence interval [CI], 4.27-17.27), renal disease (OR, 3.33; CI, 1.95-5.69), evolving myocardial infarction (OR, 2.80; CI, 1.47 5.31), congestive heart failure (OR, 2.18; CI, 1.23-3.86), total number of lesions treated (OR, 1.28; CI, 1.03-1.59), age (OR, 1.03; CI, 1.01-1.06), and history of prior coronary intervention (OR, 0.51; CI 0.26-0.99) were identified as independent predictors of MACE. In addition, vascular disease (OR, 2. 48; CI 1.37-4.50) and unstable angina pectoris (OR, 0.44; CI 0.25-0. 79) were related to adverse events when patients in cardiogenic shock were excluded from the model. With the exception of most unfavorable lesion morphology (AHA/ACC lesion type C; OR, 2.05; CI, 1.19-3.52), anatomic parameters added no further information. In the present era of device technology, success rates of percutaneous coronary revascularization procedures have increased and remain to be determined by lesion morphology. In contrast, the rate of MACE is declining and best predicted by easily identified patient characteristics. PMID- 10525223 TI - Doing it--where's the risk? PMID- 10525224 TI - Primary stent implantation is superior to balloon angioplasty in acute myocardial infarction: final results of the primary angioplasty versus stent implantation in acute myocardial infarction (PASTA) trial. PASTA Trial Investigators. AB - Several studies have shown that stent implantations in acute myocardial infarction (AMI) result in better short- and long-term outcomes than primary balloon angioplasty. These results, however, have not been ascertained in randomized trials. We randomized 136 patients out of 208 patients with AMI within 12 hr from onset into two groups: 69 patients with primary balloon angioplasty (POBA group) and 67 patients with primary stent implantation (STENT group). We compared the incidences of major cardiac events (repeat MI, target lesion revascularization, and cardiac death) and angiographic parameters during hospitalization and follow-up periods up to 12 months in these two groups. There was no significant difference in the reperfusion success rates. The incidences of major cardiac events were lower in the STENT group than in the POBA group during hospitalization, the first 6 months and 12 months (6% vs. 19%, P = 0.023; 21% vs. 46%, P < 0.0001; 22% vs. 49%, P = 0.0011). Minimum lumen diameters were significantly bigger in the STENT group than the POBA group at predischarge angiogram and 6-month follow-up (2.85 +/- 0.62 vs. 2.08 +/- 0.82 mm, P < 0.0001; 2.24 +/- 0.64 vs. 1.72 +/- 0.76, P = 0.002). Restenosis rates at 6-month follow up were significantly lower in the STENT group than in the POBA group (17% vs. 37.5%, P = 0.02). In selected patients with AMI, primary stent implantation results in a lower incidence of major cardiac events during the first 12 months, postprocedure, and less frequent 6-month restenosis than primary balloon angioplasty. PMID- 10525225 TI - Pasta without sauce? PMID- 10525226 TI - Transradial cardiac catheterization in patients with prior brachial artery cutdown. AB - The safety and efficacy of transradial cardiac catheterization in patients with prior ipsilateral brachial cutdown is not known. Using standard techniques we performed transradial catheterization in 278 consecutive patients, of which 63 had prior brachial cutdown. All patients had a strongly palpable radial pulse and a negative Allen's test. Although patients with prior cutdown were older and had a higher incidence of hypertension and prior coronary artery bypass surgery, there was no significant difference in success rates for transradial catheterization (93.6% vs. 95.3%; P = NS). There were no periprocedural complications. Brachial artery occlusion was responsible for only two unsuccessful catheterization attempts. We conclude that, with careful preprocedural screening, ipsilateral transradial cardiac catheterization can be successfully performed in a majority of patients with prior brachial cutdown. PMID- 10525228 TI - Successful stent delivery with deep seating of 6 French guiding catheters in difficult coronary anatomy. AB - Despite improvements in coronary stent design, delivery difficulties may still be encountered. Between April 1996 and September 1998, 945 patients underwent coronary stenting in our Institute. New 6 Fr Long Brite Tip (LBT) guiding catheters, allowing deep coronary artery intubation and increased backup support, were used in 25 (2.6%) of these patients presenting complex coronary anatomy and poor stent accessibility, electively in 3 (12%) and after stent delivery failure with multiple (2.1 +/- 1.2) standard guiding catheters in 22 (88%). Deep coronary artery intubation (>/= 20 mm) was successfully performed in 22 (88%) patients and was associated with adequate pressure recording and contrast opacification without blood flow compromise. Ten (22.7%) Palmaz-Schatz stents and 34 (77.3%) second-generation stents of various lengths were successfully delivered to different coronary vessels (RCA = 15, LAD = 9, LCx = 1) in all patients in whom deep coronary intubation was obtained. These data demonstrate that deep coronary artery cannulation with LBT catheters is feasible and safe and may markedly increase the rate of stent delivery success in very complex coronary anatomy and when standard guiding catheters have failed. PMID- 10525227 TI - Intracoronary adenosine administered during rotational atherectomy of complex lesions in native coronary arteries reduces the incidence of no-reflow phenomenon. AB - Rotational atherectomy (RA) of complex, calcified lesions has been associated with a high incidence of no reflow ranging from 6%-15% and concomitant myocardial necrosis with adverse prognostic implications. There are no uniform strategies for preventing this complication. The role of intracoronary adenosine for the prevention of this phenomenon during RA has not been fully evaluated. We studied the procedural outcome of 122 patients who underwent RA of complex native coronary artery lesions. Fifty-two patients received no adenosine but a variety of other agents. Seventy patients received intracoronary adenosine boluses (24 to 48 microgram prior to and after each RA run). There was no difference in the type of lesion studied, run time, or Burr to artery ratio (0.6-0.7) between the two groups. Six patients without adenosine experienced no reflow (11.6%), with resultant infarction in the target artery territory, while only 1 of 70 patients (1.4%, P - 0.023) in the adenosine group experienced no reflow. No untoward complications were observed during adenosine infusion. Intracoronary adenosine bolus administered during rotational atherectomy is easy, safe, and may significantly reduce the incidence of no reflow, which may improve the 30-day outcome of this procedure. PMID- 10525229 TI - Tricks for overcoming difficult stent delivery. PMID- 10525230 TI - Angiographic and clinical restenosis following the use of long coronary Wallstents. AB - This study assessed clinical and angiographic restenosis following the deployment of the long coronary Wallstent. Between May 1995 and June 1997, 182 Wallstents were deployed in 162 vessels in this unit. Forty-eight percent had an unstable coronary syndrome and 94% had AHA grade B or C lesions. The mean lesion length was 37 +/- 20 mm and the mean stent length was 48 +/- 20 mm. The procedural success rate was 99% and the primary success rate was 93%. Six in-patients suffered subacute stent thrombosis, the majority being in the era of anticoagulation rather than antiplatelet regimes. Seventy-three percent remained free of major adverse clinical events in the follow-up period, but 41% had angiographic restenosis. The Wallstent can be deployed in complex lesions with a high primary success rate and an acceptably low restenosis rate. The optimal management of in-stent restenosis remains to be defined. PMID- 10525231 TI - Can wallstent be the great wall against neointimal invasion? PMID- 10525232 TI - Subclinical aortic perforation with the infant double-button patent ductus arteriosus occluder. AB - Modification of the double-button (Sideris) patent ductus arteriosus (PDA) occluder has resulted in a single-strut aortic component rather than the conventional cross-strut design. We report the use of this infant PDA occluder for transcatheter closure in three patients with PDA measuring 2 mm, 3.7 mm, and 4 mm. Subclinical aortic perforation with a small aortic aneurysm developed in two patients 1 year after occluder implantation. The third patient had developed a small aortic aneurysm without perforation at 3-month follow-up. All three patients had a residual shunt and underwent successful PDA surgical closure with aortic aneurysmal repair. Single-strut umbrella designs are not recommended for PDA transcatheter closure. PMID- 10525233 TI - Cardiac perforation after device implantation for congenital heart disease: should we worry? PMID- 10525234 TI - Infectious complications related to the use of the angio-seal hemostatic puncture closure device. AB - One hundred and eight coronary angiography procedures in which the Angio-Seal device was utilized were complicated by eight (7.4%) hematomas, of which two (1.9%) subsequently developed infection (Staphylococcus aureus endarteritis and S. aureus septic hematoma). The Angio-Seal device may be a risk factor for infection for two reasons: excessive hematoma formation (a known risk factor for endarteritis), and foreign material remaining within the arterial lumen and wall, thereby creating a nidus for infection. PMID- 10525235 TI - Repair of left anterior descending coronary artery perforation by Magic Wallstent implantation. AB - Coronary rupture is a rare complication of percutaneous coronary intervention. However, it may be associated with serious hemodynamic consequences often leading to tamponade, myocardial infarction, emergency surgical intervention, or death. We report a successful percutaneous repair of a brisk left anterior descending coronary artery perforation by the implantation of a Magic Wallstent. PMID- 10525236 TI - Expanding subintimal coronary dissection under a stent-covered arterial segment: serial intravascular ultrasound observations. AB - A patient with an angiographically unrecognized minor coronary dissection in a stent-covered coronary segment in which a type D spiral dissection extended submedially to the distal artery is described. This complication occurred 6 months after stent implantation and was ascribed to injury of the stented vessel wall during an intravascular ultrasound study. PMID- 10525237 TI - Intra-arterial thrombolysis in a patient presenting with an ischemic stroke due to spontaneous internal carotid artery dissection. AB - We describe a case of a 38-year-old male who presented with acute onset of right sided hemiplegia and aphasia, who was transferred for emergent percutaneous intervention. Angiography revealed a dissection with total occlusion of the left internal carotid artery (ICA) with propagation of thrombus in the distribution of the middle cerebral artery (MCA). Therapy was directed at the MCA and not the ICA. Intra-arterial thrombolysis was performed on the M1 and M2 branches of the left middle cerebral artery, resulting in almost complete resolution of symptoms during the angiography procedure. Heparin was continued postprocedure, and the patient was discharged home on warfarin and aspirin with minimal residual symptoms. PMID- 10525238 TI - Short- and long-term histopathologic evaluation of stenting using a self expanding nitinol stent in pig carotid and iliac arteries. AB - Stenting is increasingly being used to treat carotid artery disease. However, complications including distal embolization, stent thrombosis, stent collapse from external compression, the need for high-pressure inflation with increased neointimal response, or balloon rupture during stent expansion and stent loss are all potential problems and of concern. To address each of these specific concerns, a new stent was designed, which is self-expandable, made of nitinol, with temperature-dependent superelastic properties, and with high vessel wall surface coverage. Since this device has a number of novel characteristics, we aimed to assess the short- and long-term histopathologic response in pig carotid and iliac arteries. Single stents were deployed in pig carotid and iliac arteries after overstretch balloon injury. Angiograms were performed pre- and poststenting and prior to sacrifice. Intravascular ultrasound was used before implantation to determine vessel size. Vessels were examined histologically at 1 month (n = 6) and 6 months (n = 6) for morphometric analysis, hemorrhage and thrombus, endothelialization, and inflammatory and fibrotic responses. There was a 100% angiographic success rate at implantation. In one case, it was determined histologically that a single stent was implanted in a dissection plane of a pig's left iliac artery and was occluded by organized thrombus, with the true lumen being patent. At 6-month follow-up, this was the only evidence of a single stent occlusion, with flow adjacent to the stent in the true lumen. In the other vessels, the stents showed good vessel wall-stent apposition and the lumens were patent with a concentric and thin neointima. Inflammatory cells were rare and there were no mural thrombi. Coverage of the vessel wall by endothelial-like cells was complete at 1 month. The novel nitinol EndoStent appears to have favorable biocompatibility with minimal thrombus deposition or inflammatory response, and its use is feasible for clinical application in carotid and iliac arteries. PMID- 10525239 TI - Preventive effects of the heparin-coated stent on restenosis in the porcine model. AB - The coronary stent reduces acute coronary arterial occlusion and late restenosis during and after coronary intervention. However, stent thrombosis and restenosis are still major limitations in the widespread use of the coronary stent. Local drug delivery using the heparin-coated stent may be a new approach, which reduces the incidence of stent thrombosis and restenosis. In order to evaluate the effects of the heparin-coated stent on stent restenosis, heparin-coated stents were compared with control stents in a porcine coronary stent restenosis model. Stent overdilation injury (stent:artery = 1.3:1.0) was performed with bare Wiktor stents (group I, n = 10) and heparin-coated Wiktor stents (group II, n = 20; HEPAMED, Medtronics) in porcine coronary arteries. Follow-up quantitative coronary angiography (QCA) was performed at 4 weeks after stenting, and histo pathologic assessments of stented porcine coronary arteries were compared in both groups. On QCA, percent diameter stenosis was significantly higher in group I than in group II (16.3% +/- 6.62% vs. 9.6% +/- 5.06%, P < 0.05). The injury score of stented porcine coronary arteries was the same in both groups (1. 26 +/- 0.23 vs. 1.20 +/- 0.22). The area of pathologic stenosis of the stented arteries was higher in group I than in group II (41.6% +/- 12.5% vs. 27.1% +/- 9.9%, P < 0.005). The neointimal area was higher in group I than in group II (4.58 +/- 1.41 mm(2) vs. 2.57 +/- 1.07 mm(2), P < 0.05). By immunohistochemistry, the proliferating cell nuclear antigen (PCNA) index was higher in group I compared with group II (11.2% +/- 6.75% vs. 6.3% +/- 4.14%, P < 0.05). The heparin-coated stent is effective in the prevention of late coronary stent restenosis in a porcine coronary stent restenosis model. This may be related to the inhibition of neointimal cell proliferation. PMID- 10525240 TI - String-plucking as a mechanism of chordal rupture during balloon mitral valvuloplasty using inoue balloon catheter. PMID- 10525241 TI - Reply to letter to the editor by francis Y.K. Lau PMID- 10525242 TI - Archives of pacing and electrophysiology: In search of a permanent home... PMID- 10525243 TI - Induction of atrial fibrillation and flutter in dogs using methacholine. AB - Systemic infusion of methacholine has been used to facilitate induction of atrial fibrillation. However, the dose-response relationship, reproducibility and effect of anesthetic agents on induction are not well understood. The use of methacholine to facilitate electrical induction of sustained (>10 minutes duration) atrial fibrillation or flutter was examined. In 25 dogs induction of atrial arrhythmias was attempted using a series of ten 50 Hz trains of 10 seconds duration delivered via an endocardial catheter in the baseline anaesthetized state and subsequently in the presence of graded doses of intravenous methacholine (maximum 5 microg/kg/min). Studies were repeated in 13 dogs to assess reproducibility. Twelve dogs (48%) had inducible sustained atrial flutter or fibrillation lasting greater than 10 minutes in the baseline state. During infusion of methacholine the remaining 13 (52%) dogs also had inducible sustained atrial flutter or fibrillation (mean infusion rate 1.6 +/- 1.9 microg/kg/min). Induction of sustained atrial flutter or fibrillation was reproducible in all but one dog. The type of anesthetic did not significantly affect inducibility. Induction of prolonged atrial fibrillation or flutter is possible in the baseline anaesthetized state in approximately half of dogs using high frequency programmed electrical stimulation. The yield of inducible sustained atrial fibrillation or flutter with programmed stimulation during intravenous infusion of methacholine was increased to 100%. Induction of sustained atrial fibrillation or flutter was highly reproducible. PMID- 10525244 TI - Effect of atrial pressure increase on effective refractory period and vulnerability to atrial fibrillation in patients with lone atrial fibrillation. AB - BACKGROUND: There is evidence suggesting that atrial fibrillation (AF) may be induced by acute increase of atrial pressure. The aim of the present study was to investigate the effect of alterations in atrial pressure, induced by varying the atrioventricular (AV) interval, on atrial refractoriness, and on the frequency of induction of (AF), in patients with a history of lone atrial fibrillation (LAF). METHODS AND RESULTS: Twenty-five patients were included in this study. The patients were divided in two groups: the LAF group, and the control group. None of the patients in either group had organic heart disease. Effective refractory period (ERP) and duration of atrial extrastimulus electrogram (A(2)) were measured at two right atrial sites (high lateral wall, atrial appendage) during AV pacing (cycle length: 500 msec) with different AV intervals. Peak, minimal and mean atrial pressure increased from 8.57 +/- 2.37 to 18.14 +/- 4.74 mm Hg, 2 +/- 2.23 to 5.14 +/- 2.60 mm Hg (p = 0.0001) and from 4.28 +/- 1.6 mm Hg to 9.77 +/- 2.9 mm Hg (p = 0.001), respectively during AV interval modification. During lateral and atrial appendage pacing, with a progressive decrease of AV interval to 160, 100, 80, 40, 0 msec, the ERP, the dispersion of ERP, functional refractory period (FRP), A2 and latency period (LP) did not change significantly, in both groups. The frequency of induction of AF was not statistically different in both lateral atrial wall and appendage, during pacing in different AV intervals. CONCLUSIONS: This study demonstrates that alterations in the intraatrial pressure does not have important effects on atrial refractoriness and does not increase vulnerability to AF in patients with a history of LAF. PMID- 10525246 TI - DDD-pacing-induced cardiomyopathy following AV node ablation for persistent atrial tachycardia. AB - Ventricular rate control by catheter ablation of the AV node and pacing in patients with persistent atrial tachycardia has been reported to improve left ventricular function. However, this approach requires careful selection of the pacing mode. We report a patient who underwent AV node ablation for persistent multiple atrial tachycardias, and who then had a non-mode-switching pacemaker implanted. Because of an inappropriately programmed relatively high upper rate limit, the patient developed left ventricular dysfunction after 6 years. This resolved after programming the pacemaker to VVI at 70 bpm. PMID- 10525245 TI - The spatial dispersion of atrial refractoriness and atrial fibrillation vulnerability. AB - The local dispersion of conduction and refractoriness has been considered essential for induction of atrial arrhythmias. This study sought to determine whether a difference of refractoriness and vulnerability for induction of atrial fibrillation between trabeculated and smooth as well as high and low right atrium may contribute to initiation of atrial fibrillation in dogs. In 14 healthy mongrel dogs weighing 22.4 +/- 1 kg, closed-chest endocardial programmed stimulation was performed from four distinct right atrial sites. Atrial refractory periods and vulnerability for induction of atrial fibrillation or premature atrial complexes were determined during a basic cycle length of 400 and 300 ms and an increasing pacing current strength. For a pacing cycle length of 300 ms, atrial refractory periods were longer on the smooth, as compared to the trabeculated right atrium (102 +/- 25 vs. 97 +/- 17 ms, p < 0.05), whereas for a pacing cycle length of 400 ms, there was no significant difference. The duration of the vulnerability zone for induction of atrial fibrillation was longer on the smooth right atrium, for a cycle length of both 400 ms (40 +/- 30 vs. 31 +/- 22 ms; p < 0.05) and 300 ms (33 +/- 25 vs. 23 +/- 21 ms; p < 0. 01). When comparing high and low right atrium, refractory periods were longer on the the low right atrium, for a cycle length of both 400 ms (111 +/- 23 vs. 94 +/- 24 ms; p < 0.01) and 300 ms (104 +/- 20 vs. 96 +/- 23 ms; p < 0.01). For a pacing cycle length of 300 ms, the duration of the atrial fibrillation vulnerability zone was longer for the high, as compared to the low right atrium (34 +/- 22 vs. 22 +/- 22, p < 0.01). Seven dogs with easily inducible episodes of atrial fibrillation demonstrated significantly shorter refractory periods as compared to 7 non vulnerable dogs, regardless of pacing site and current strength. In conclusion, significant differences in refractoriness and vulnerability for induction of atrial fibrillation can be observed in the area of the crista terminalis in healthy dogs. Thus, local anatomic factors may play a role in the initiation of atrial fibrillation. PMID- 10525247 TI - Effect of general anesthesia on the defibrillation energy requirement in patients undergoing defibrillator implantation. AB - BACKGROUND: The effect of general anesthesia on defibrillation efficacy in humans is not known. The purpose of this study was to determine the effect of general anesthesia on the defibrillation energy requirements in patients undergoing implantation of a pectoral defibrillator. METHODS AND RESULTS: Nineteen consecutive patients who underwent defibrillator implantation under general anesthesia were prospectively compared to 16 consecutive patients who underwent defibrillator implantation by the same physicians, using similar devices, at another hospital under conscious sedation. Pre-discharge testing was performed 1.4 +/- 1.0 days after implant using sedation in both groups. The defibrillation energy requirement was determined using the same predefined step-down protocol (15, 10, 8, 5, 3, 1 J) at the time of implantation and during pre-discharge testing. The clinical characteristics of the patients were similar between groups. There was no significant difference in the mean implant defibrillation energy requirement compared to the mean pre-discharge defibrillation energy requirement in either the general anesthesia group (8.5 +/- 4.7 vs. 8.4 +/- 3.4 J; p = 0.9) or in the conscious sedation group (9.4 +/- 3.9 vs. 9.0 +/- 3.8 J; p = 0.7). CONCLUSIONS: When compared to conscious sedation, general anesthesia with mechanical ventilation has no significant effect on defibrillation efficacy in patients undergoing defibrillator implantation. PMID- 10525248 TI - Comparison of therapy detection times between implantable cardioverter defibrillators with standard dual- and single-chamber pacing. AB - Previous implantable cardioverter defibrillators (ICDs) required patients in need of dual-chamber (DDD) pacing for improved hemodynamic status to undergo implantation of separate devices to treat bradycardia and/or ventricular arrhythmias. An investigation was conducted to verify the performance of a new ICD that combines both therapies.Sixty-nine patients at 17 European and Canadian centers were implanted with VENTAK AV models 1810/1815, ICD's that includes DDD pacing and algorithms designed to differentiate between atrial and ventricular arrhythmias. 36 of the cohort were compared to 32 patients tested at six centers with an external test device (VENTAK MINI). In both cohorts detection times were calculated for ventricular fibrillation (VF) induced at implant. The mean detection times (DT) from the VENTAK AV device were compared to the DT from the VENTAK MINI device. Patient characteristics of the VENTAK AV and the VENTAK MINI control groups were similar. Mean VF detection time (+/-SD) with the VENTAK AV device was 2.21 +/- 0.54 seconds, as compared with 1.87 +/- 0.62 seconds with the VENTAK MINI (p < 0.01), indicating that the difference in means did not exceed one second. The VENTAK AV system function did not demonstrate interaction with the pacemaker function, as indicated by the clinical significance with the detection times of the study device. The difference in detection times between cohorts did not statistically exceed one second. Appropriate detection of the new ICD was not compromised by the addition of the dual-chamber pacing therapy. PMID- 10525249 TI - Induction of ventricular fibrillation by T wave shocks: observations from monophasic action potential recordings. AB - INTRODUCTION: Shocks given during the vulnerable period of cardiac repolarization may induce ventricular fibrillation (VF). However, the relationship of the vulnerable period and the monophasic action potential (MAP) has not yet been reported in humans. The purpose of this study was, therefore, to determine how the monophasic action potential recorded from the right ventricle correlates with inducibility of VF using T wave shocks during ventricular pacing. METHODS: Eleven patients undergoing implantable cardioverter defibrillator (ICD) implantation had a MAP catheter positioned in the right ventricle (RV). The local monophasic action potential duration at 90% repolarization (MAP90) duration was measured during pacing at 400 ms. VF induction was attempted by pacing at 400 ms for 10 cycles and then giving a 1.0 joule monophasic T wave shock at varying coupling intervals (CI) to the last paced stimulus. The maximum and minimum CI that induced VF were determined and mapped in relation to the MAP90 recording. RESULTS: The average paced MAP duration was 275 +/- 20 ms. The minimum and maximum CI to induce VF were 255 +/- 24 ms and 325 +/- 36 ms respectively. This ranged from 93% to 118% of the MAP90 duration but because of delay in conduction time to the MAP catheter, shocks that induced ventricular fibrillation occurred between 74% and 99% of local repolarization time. CONCLUSION: VF is inducible with low energy T wave shocks falling during the last 25% of the right ventricular MAP90 recording. This corresponds with VF initiation during phase III repolarization. PMID- 10525250 TI - A shocking case of pseudoephedrine use. PMID- 10525253 TI - The World of Pacing and Electrophysiology. PMID- 10525252 TI - 95 years of electrocardiography. PMID- 10525254 TI - Cardiac Pacing and Electrophysiology International Calendar of Events. PMID- 10525251 TI - Temperature-controlled radiofrequency catheter ablation with a 10-mm tip electrode creates larger lesions without charring in the porcine heart. AB - BACKGROUND: Radiofrequency catheter ablation of atrial flutter, atrial fibrillation or ventricular tachycardia may be favoured by large lesions. We compared lesions created in unipolar mode using 10-mm/8 F electrodes with those of 4-mm/7 F catheters. METHODS: Ablations were first performed in porcine hearts in vitro (70 degrees C, 60 s, tangential catheter tip-tissue orientation). Anaesthetized pigs were thereafter ablated with 10- or 4-mm catheters in the right atrial free wall (RAFW), inferior vena cava-tricuspid valve (IVC-TV) isthmus and left ventricle (LV). RESULTS: In vitro, lesion length doubled and lesion volume tripled using the 10-mm catheter. Average power supply was 69 (SD12) (10-mm tip) versus 26 (SD7) W (4-mm tip). In vivo, lesion length increased by 50% and lesion volume fivefold. Charring at the lesion surface or sudden impedance rises were not observed in vivo. Histologically, coagulation necrosis and minor haemorrhages were found. One RAFW lesion (10-mm) showed a dissection approaching the epicardium. Fibrinous platelet clots or overt thromboses covered the endocardial surface in half of all lesions. Three 10-mm electrode isthmus lesions extended to the right descending posterior artery and one LV lesion to the left anterior descending artery, but there was no damage to the arterial walls. Following six ablations with the 10-mm electrode and two with the 4-mm tip, injury to the adjacent lung tissue of 0.5 to 6.0 mm depth was found (p = 0.22). CONCLUSION: RF ablation using 10-mm/8 F electrodes created significantly larger lesions. 10-mm electrodes appeared safe in the porcine IVC-TV isthmus and LV, but not in the RAFW. PMID- 10525255 TI - Polymerase chain reaction analysis for diagnosis of Tropheryma whippelii infective endocarditis in two patients with no previous evidence of Whipple's disease. PMID- 10525256 TI - Is human T-cell lymphotropic virus type I more clever than human immunodeficiency virus type 1? PMID- 10525257 TI - Computer keyboards as reservoirs for Acinetobacter baumannii in a burn hospital. PMID- 10525258 TI - Reply PMID- 10525259 TI - The author replies PMID- 10525260 TI - Adducts of acrylonitrile with hemoglobin in nonsmokers and in participants in a smoking cessation program. AB - Hemoglobin adducts have been used to assess exposure to carcinogenic compounds in tobacco smoke. However, because of background levels in nonsmokers, most adducts that have been studied are not useful for monitoring low-level exposure. Bergmark [(1997) Chem. Res. Toxicol. 10, 78-84] showed that the level of adducts of acrylonitrile (AN) with N-terminal valine (ANVal) increases with increasing cigarette consumption, and the increment from 1 cigarette/day was estimated to be 8 pmol/g of globin. The background level of ANVal in nonsmokers was not quantified (<2 pmol/g of globin). The objective of this study was to determine the background level of ANVal in hemoglobin and to study the stability of this adduct in vivo. Globin samples previously analyzed by Bergmark from 17 nonsmokers and 2 smokers were reanalyzed in the study presented here. Globin samples from 7 additional nonsmokers and from 10 participants in a smoking cessation program were also analyzed. Smoking habits and exposure to environmental tobacco smoke (ETS) were assessed by interview. Only two of the participants completed the program. The levels of ANVal in these 2 subjects decreased after quitting and were at background level by 126 days. The time course of the decrease was compatible with removal of stable adducts. The levels of ANVal in the nonsmokers were 0.76 +/- 0.36 (mean +/- SD) (n = 18; reporting no exposure ETS), 1.1 +/- 0.6 (n = 3; reporting exposure to ETS), and 1. 2 +/- 0.5 pmol/g of globin (n = 3; snuff users). Thus, the adduct level in nonsmokers corresponds to the adduct increment from about 0. 1 cigarette/day. Measurements of the level of ANVal could be used to distinguish between nonsmokers and low-level smokers on an individual level, but larger groups of individuals would be required to detect a possible contribution to the background from passive smoking. PMID- 10525261 TI - Tributyltin-induced apoptosis requires glycolytic adenosine trisphosphate production. AB - The toxicity of tributyltin chloride (TBT) involves Ca(2+) overload, cytoskeletal damage, and mitochondrial failure leading to cell death by apoptosis or necrosis. Here, we examined whether the intracellular ATP level modulates the mode of cell death after exposure to TBT. When Jurkat cells were energized by the mitochondrial substrate, pyruvate, low concentrations of TBT (1-2 microM) triggered an immediate depletion of intracellular ATP followed by necrotic death. When ATP levels were maintained by the addition of glucose, the mode of cell death was typically apoptotic. Glycolytic ATP production was required for apoptosis at two distinct steps. First, maintenance of adequate ATP levels accelerated the decrease of mitochondrial membrane potential, and the release of the intermembrane proteins adenylate kinase and cytochrome c from mitochondria. A possible role of the adenine nucleotide exchanger in this first ATP-dependent step is suggested by experiments performed with the specific inhibitor, bongkrekic acid. This substance delayed cytochrome c release in a manner similar to that caused by ATP depletion. Second, caspase activation following cytochrome c release was only observed in ATP-containing cells. Bcl-2 had only a minor effect on TBT-triggered caspase activation or cell death. We conclude that intracellular ATP concentrations control the mode of cell death in TBT-treated Jurkat cells at both the mitochondrial and caspase activation levels. PMID- 10525262 TI - Formation of deaminated products in styrene oxide reactions with deoxycytidine. AB - The reaction of racemic styrene oxide with deoxycytidine under aqueous conditions was studied. The four principal products isolated were a pair of diastereomeric N(4)-(2-hydroxy-1-phenylethyl)deoxycytidines ( approximately 20% of the products) and a pair of diastereomeric 3-(2-hydroxy-2-phenylethyl)deoxyuridines ( approximately 80% of the products). Reactions with optically active styrene oxides allowed the configurations of the 3-(2-hydroxy-2-phenylethyl)deoxyuridines to be assigned, and these structures were confirmed by an independent synthesis from deoxyuridine. Also, it was possible to tentatively assign the configurations of the N(4)-(2-hydroxy-1-phenylethyl)deoxycytidines that had undergone some racemization during the reaction (the ratio of the retained to inverted configuration of the products was approximately 1:7). PMID- 10525263 TI - Analysis in the rat of 4-hydroxynonenal metabolites excreted in bile: evidence of enterohepatic circulation of these byproducts of lipid peroxidation. AB - 4-Hydroxynonenal (HNE) is a cytotoxic product resulting from the lipid peroxidation of membrane polyunsaturated fatty acids. In vitro, metabolism mainly leads to the corresponding alcohol (DHN), carboxylic acid (HNA), and the glutathione conjugate, whereas in vivo, mercapturic acid conjugates of HNE, DHN, HNA, and HNA-lactone and, more recently, dicarboxylic acids and related mercapturate conjugates were identified in urine of rats. In the study presented here, the identity of the HNE biotransformation products in the bile of rats following a single iv administration of [4-(3)H]HNE and the potential for enterohepatic recycling of HNE metabolites were investigated. The identity of metabolites was assessed by comparison of their HPLC retention times with those of the corresponding synthesized standards and by mass spectrometry analysis. Five metabolites were present in the bile; two of them corresponded to HNE- and DHN-glutathione conjugates. Two others metabolites were identified as DHN- and HNA-lactone mercapturic acid conjugates. The fifth metabolite was isolated but remained unidentified. As previously observed for urinary elimination, the kinetic excretion of biliary metabolites exhibited a rapid metabolism of HNE in rats. Within 4 h of injection, the bile accounted for 19.5% (+/-2.8%) of the injected radioactivity, whereas only 3% was found in the feces within 48 h [Alary, J., et al. (1995) Chem. Res. Toxicol. 8, 34-39]. The extent of HNE enterohepatic recycling was estimated utilizing a modified version of the linked rat model in three animals. All rat recipients were found to have measurable levels of HNE metabolites in bile, confirming that HNE is likely to undergo enterohepatic recirculation in the rat. The extent of recycling was approximatly 7. 7% of the total dose in this model. Two unknown metabolites were present in the bile of recipient rats and not found in the bile of donors rats, suggesting that intestinal microflora and/or intestinal mucosa could biotransform HNE related compounds before or during the reabsorption process. PMID- 10525264 TI - Conformational analysis of the major DNA adduct derived from the food mutagen 2 amino-3-methylimidazo[4,5-f]quinoline. AB - The heterocyclic amine 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) is one of a number of carcinogens found in barbecued meat and fish. It is mutagenic in bacterial and mammalian assays and induces tumors in mammals. IQ is biochemically activated to a derivative which reacts with DNA to form a major covalent adduct at carbon 8 of guanine. This adduct may deform the DNA and consequently cause a mutation, which may be responsible for initiating IQ's carcinogenicity. Atomic resolution structures of the IQ-damaged DNA are not yet available experimentally. We have carried out an extensive molecular mechanics energy minimization search to locate feasible structures for the major IQ-DNA adduct in the representative sequence d(5'-G1-G2-C3-G4-C5-C6-A7-3'). d(5'-T8-G9-G10-C11-G12-C13-C14-3') with IQ modification at G4; this contains the GGCGCC mutational hotspot sequence known as NarI. The molecular mechanics program AMBER 5.0 with the force field of Cornell et al. [(1995) J. Am. Chem. Soc. 117, 5179-5197] was employed, including explicit Na(+) counterions and an implicit treatment for solvation. However, key parameters, the partial charges, bond lengths, bond angles, and dihedral parameters of the modified residue, are not available in the AMBER database. We carefully parametrized the force field, created 800 starting conformations which uniformly sampled at 18 degrees intervals each of the three flexible torsion angles that govern the IQ-DNA orientation, and minimized their energy. A conformational mix of structural types, including major groove, minor groove, and base-displaced intercalated carcinogen positions, was generated. This mixture may be related to the diversity of mutational outcomes induced by IQ. PMID- 10525265 TI - Formation and reactions of N(7)-aminoguanosine and derivatives. AB - Arylamines are mutagens and carcinogens and are thought to initiate tumors by forming adducts with DNA. The major adducts are C(8)-guanyl, and we have previously suggested a role for guanyl-N(7) intermediates in the formation process. N(7)-Aminoguanosine (Guo) was synthesized and characterized, with the position of the NH(2) at N7 established by two-dimensional rotating frame Overhauser enhancement NMR spectroscopy. In DMF, N(7)-NH(2)Guo formed C(8) NH(2)Guo and the cyclic product C(8):5'-O-cycloGuo. In aqueous media, these products were formed along with 8-oxo-7,8-dihydroGuo, N(7)-NH(2)guanine, and a product characterized as a purine 8, 9-ring-opened derivative (N aminoformamidopyrimidine). The rate of aqueous decomposition of N(7)-NH(2)Guo increased with pH, with a t(1/2) of 10 h at pH 7 and a t(1/2) of 2 h at pH 9. The rate of migration of NH(2) from N7 to C8 is fast enough to explain the formation of C(8)-NH(2)Guo from the reaction of 2, 4-dinitrophenoxyamine with Guo but not the formation of C(8)-(arylamino)Guo in the reaction of Guo with aryl hydroxylamine esters; however, the fluorenyl moiety may facilitate the proposed rearrangement by stabilizing an incipient negative charge in the transfer. In the reaction of Guo with N-hydroxy-2-aminofluorene and acetylsalicylic acid, a peak with the mass spectrum expected for N(7)-(2-aminofluorenyl)Guo was detected early in the reaction and was distinguished from C(8)-(2-aminofluorenyl)Guo. NMR experiments with [8-(13)C]Guo also provided some additional support for transient formation of N(7)-(2-aminofluorenyl)Guo. We conclude that a guanyl-N(7) intermediate is reasonable in the reaction of activated arylamines with nucleic acids, although an exact rate of transfer of an N(7)-arylamine group to the C8 position has not yet been quantified. The results provide an explanation for the numerous products associated with modification of DNA by activated arylamines. However, the contribution of "direct" reaction at the guanine C8 atom cannot be excluded. PMID- 10525266 TI - Sequence-dependent repair of synthetic AP sites in 15-mer and 35-mer oligonucleotides: role of thermodynamic stability imposed by neighbor bases. AB - We previously reported that 15-mer oligonucleotides with a central 1, N(6) epsilonA were cleaved by alkylpurine-DNA N-glycosylase as a function of T(m), modulated by neighbor bases [Hang, B., Sagi, J., and Singer, B. (1998) J. Biol. Chem. 273, 33406-33413]. This type of investigation has now been extended to cleavage by Escherichia coli endonuclease IV of a centrally placed synthetic AP site using both 15-mer and 35-mer duplexes. In 15-mers, the triplet sequences adjunct to the central AP site greatly affected the thermodynamic stability. The repair rate paralleled the thermal stability since endonuclease IV requires a double-stranded substrate. When the AP site-containing duplexes were 35-mers, there was also a general correlation between the thermostability and cleavage efficiency. However, the difference in the cleavage rates between different sequences was much less than with the 15-mers. Since the 35-mers were more than 96% annealed, this difference presumably results from local stability and structure adjacent to the AP site. These results suggest that under enzyme limiting conditions or overproduction of AP sites, sequence-dependent differential repair could occur in vivo. PMID- 10525267 TI - Methylarsenicals and arsinothiols are potent inhibitors of mouse liver thioredoxin reductase. AB - Thioredoxin reductase (TR, EC 1.6.4.5) was purified 5800-fold from the livers of adult male B6C3F1 mice. The estimated molecular mass of the purified protein was about 57 kDa. The activity of the purified enzyme was monitored by the NADPH dependent reduction of 5, 5'-dithiobis(2-nitrobenzoic acid) (DTNB); this activity was fully inhibited by 1 microM aurothioglucose. Arsenicals and arsinothiols, complexes of As(III)-containing compounds with L-cysteine or glutathione, were tested as inhibitors of the DTNB reductase activity of the purified enzyme. Pentavalent arsenicals were much less potent inhibitors than trivalent arsenicals. Among all the arsenicals, CH(3)As(III) was the most potent inhibitor of TR. CH(3)As(III) was found to be a competitive inhibitor of the reduction of DTNB (K(i) approximately 100 nM) and a noncompetitive inhibitor of the oxidation of NADPH. The inhibition of TR by CH(3)As(III) was time-dependent and could not be reversed by the addition of a dithiol-containing molecule, 2,3 dimercaptosuccinic acid, to the reaction mixture. The inhibition of TR by CH(3)As(III) required the simultaneous presence of NADPH in the reaction mixture. However, unlike other pyridine nucleotide disulfide oxidoreductases, there was no evidence that mouse liver TR was inactivated by exposure to NADPH. Treatment with CH(3)As(III) did not increase the NADPH oxidase activity of the purified enzyme. Thus, CH(3)As(III), a putative intermediate in the pathway for the biomethylation of As, is a potent and irreversible inhibitor of an enzyme involved in the response of the cell to oxidative stress. PMID- 10525268 TI - Intracellular S-glutathionyl adducts in murine lung and human bronchoepithelial cells after exposure to diisocyanatotoluene. AB - Diisocyanatotoluene (toluene diisocyanate, TDI), a 4:1 mixture of 2, 4- and 2,6 isomers used in the preparation of polyurethanes, causes occupational asthma by an as yet unknown mechanism. We previously showed that it forms adducts with the apical surface of the bronchoepithelium in vivo, and with ciliary microtubules in cultured human bronchoepithelial (HBE) cells. These results suggested that TDI may not enter HBE cells. In vitro studies, however, showed that TDI avidly forms bis adducts with glutathione (GSH) and that these adducts transfer monoisocyanato monoglutathionyl-TDI to a sulfhydryl-containing peptide. This study sought to elucidate intracellular reactions of TDI. Using an electron paramagnetic resonance spectrometric (EPR) method, we established that the level of thiol dependent quenching of phenoxyl radicals of etoposide was decreased >40% in pulmonary tissue of mice that received TDI intrabronchially. Similarly, HBE cells exposed to 100 ppb TDI vapor experienced a >30% reduction in thiol levels as determined with a thiol-specific fluorescent probe (ThioGlo 1). HPLC/UV analysis of lysates from HBE cells exposed to 200 and 500 ppb TDI vapor suggested a dose related formation of S-glutathionyl adducts. Data from the 500 ppb TDI-treated HBE cells verified the identity of the 2-monoglutathionyl-4-monoisocyanato adduct. The results provide firm evidence that TDI enters pulmonary cells and reacts with GSH. This rapid reaction leading to formation of S-glutathionyl adducts of TDI suggests the importance of cellular thiols in TDI-induced pulmonary disease. PMID- 10525269 TI - Acylase-catalyzed deacetylation of haloalkene-derived mercapturates. AB - Mercapturates (S-substituted N-acetyl-L-cysteines) are terminal metabolites formed by the glutathione-dependent metabolism of electrophilic xenobiotics, including haloalkenes. Acylases catalyze the hydrolysis of N-acyl-L-amino acids, including many xenobiotic-derived mercapturates, to give fatty acids and amino acids as products. Although several acylases have been identified, the acylases that catalyze the deacetylation of the haloalkene-derived mercapturates have not been identified and characterized. Acylase I catalyzes the deacetylation of some haloalkene-derived mercapturates, including S-(1,1,2, 2-tetrafluoroethyl)-N acetyl-L-cysteine, S-(2-chloro-1,1, 2-trifluoroethyl)-N-acetyl-L-cysteine, and S (2-bromo-1,1, 2-trifluoroethyl)-N-acetyl-L-cysteine [Uttamsingh, V., et al. (1998) Chem. Res. Toxicol. 11, 800-809]. In the studies presented here, we identified a rat kidney acylase that catalyzed the hydrolysis of the haloalkene derived mercapturates S-(1, 2-dichlorovinyl)-N-acetyl-L-cysteine, S-(1,2,3,4,4 pentachloro-1, 3-butadienyl)-N-acetyl-L-cysteine, and S-(2,2-dibromo-1, 1 difluoroethyl)-N-acetyl-L-cysteine. The substrate selectivity and amino acid sequence of the purified rat kidney acylase were studied. Although the sequence of the purified rat kidney acylase was somewhat identical with that of aspartoacylase, it did not catalyze the hydrolysis of N-acetyl-L-aspartate. PMID- 10525270 TI - Distinct endoplasmic reticulum signaling pathways regulate apoptotic and necrotic cell death following iodoacetamide treatment. AB - Environmental stress induces the synthesis of glucose-regulated proteins (Grps) in the endoplasmic reticulum (ER) and heat shock proteins (Hsps) in the cytoplasm. Iodoacetamide (IDAM), a prototypical alkyating agent, induces both Grp and Hsp synthesis in renal epithelial cells and causes necrosis which is prevented by prior activation of the ER stress response (pre-ER stress) [Liu, H., et al. (1997) J. Biol. Chem. 272, 21751-21759]. In this study, we examined the biochemical pathways leading to IDAM-induced apoptosis and investigated the role of the ER stress response in apoptotic cell death. The antioxidant N,N'-diphenyl p-phenylenediamine (DPPD) prevented necrosis after IDAM treatment, but the cells went on to die with hallmarks of apoptosis, i.e., cell detachment, caspase-3 activation, cleavage of poly(ADP-ribose)polymerase (PARP), and DNA-ladder formation, all of which were blocked by the general caspase inhibitor zVAD. As with IDAM-induced necrosis, dithiothreitol protected against apoptosis, but cell permeable calcium chelators did not, suggesting that distinct biochemical pathways mediate these two forms of cell death. Pre-ER stress, but not heat shock, prevented IDAM-induced apoptosis. pkASgrp78 cells are deficient in Grp78 induction due to expression of a grp78 antisense RNA and are more sensitive to necrosis. However, these cells were resistant to IDAM-induced apoptosis and had increased basal levels of Grp94 and a KDEL-containing protein of about 50 kDa. Thus, the expression of grp78 antisense perturbs ER functions and activates expression of other ER stress genes accounting for the resistance to apoptosis. Taken together, the data describe functionally distinct signaling pathways through which the ER regulates apoptosis and necrosis caused by chemical toxicants. PMID- 10525271 TI - In vivo production of nitric oxide after administration of cyclohexanone oxime. AB - Cyclohexanone oxime (CHOX), an intermediate used in the synthesis of polycaprolactam (Nylon-6), has been reported to be hematotoxic in Fischer rats. The in vivo metabolism of CHOX was found to release nitric oxide, which was detected in venous blood by electron paramagnetic resonance spectroscopy as the nitrosylhemoglobin complex. In vitro incubation of CHOX with venous blood resulted in the formation of the characteristic nitrosylhemoglobin complex, suggesting that the blood was a possible site for metabolism. Excessive nitric oxide production may, in part, contribute to the observed toxicity of CHOX. PMID- 10525272 TI - Biotransformation of [(12)C]- and [(13)C]-tert-amyl methyl ether and tert-amyl alcohol. AB - tert-Amyl methyl ether (TAME) is intended for use as a gasoline additive to increase oxygen content. Increased oxygen content in gasoline reduces tailpipe emissions of hydrocarbons and carbon monoxide from cars. Due to possible widespread use of TAME, the toxicity of TAME is under investigation. We studied the biotransformation of TAME in rats and one human volunteer after inhalation of (12)C- or (13)C-labeled TAME. In addition, the biotransformation of [(13)C]-tert amyl alcohol was studied in rats after gavage. Urinary metabolites were identified by GC/MS and (13)C NMR. Rats (two males and two females) were individually exposed to 2000 ppm [(12)C]- or [(13)C]TAME for 6 h, and urine was collected for 48 h. Free and glucuronidated 2-methyl-2,3-butanediol and a glucuronide of tert-amyl alcohol were identified by (13)C NMR, GC/MS, and LC/MS/MS as major urinary metabolites on the basis of the relative intensities of the (13)C NMR signals. The presence of several minor metabolites was also indicated by (13)C NMR; they were identified as tert-amyl alcohol, 2-hydroxy-2 methylbutyric acid, and 3-hydroxy-3-methylbutyric acid. One human volunteer was exposed to an initial concentration of 27 000 ppm [(13)C]TAME by inhalation for 4 min from a 2 L gas sampling bag, and metabolites of TAME excreted in urine were analyzed by (13)C NMR. All TAME metabolites identified in rats were also present in the human urine samples. To study tert-amyl alcohol biotransformation, male rats (n = 3) were treated with 250 mg/kg [(13)C]-tert-amyl alcohol dissolved in corn oil by gavage, and urine was collected for 48 h. (13)C NMR of the urine samples showed the presence of metabolites identical to those in the urine of [(13)C]TAME-treated rats. Our results suggest that TAME is extensively metabolized by rats and humans to tert-amyl alcohol which may be further oxidized to diols and carboxylic acids. These reactions are likely mediated by cytochrome P450-dependent oxidations. PMID- 10525273 TI - Effect of cations on the formation of DNA alkylation products in DNA reacted with 1-(2-Chloroethyl)-1-nitrosourea. AB - The purpose of this study was to examine the influence of cations on the formation of the individual DNA alkylation products derived from 1-(2 chloroethyl)-1-nitrosourea (CNU). Reaction of calf-thymus DNA with [(3)H]CNU in 10 mM triethanolamine buffer produced 13 DNA adducts. Seven of these adducts were identified as N7-(2-hydroxyethyl)guanine, N7-(2-chloroethyl)guanine, 1, 2-(diguan 7-yl)ethane, N1-(2-hydroxyethyl)-2-deoxyguanosine, 1-(N1-2-deoxyguanosinyl)-2-(N3 2-deoxycytidyl)ethane, O(6)-(2-hydroxyethyl)-2-deoxyguanosine, and phosphotriesters. The ratios of the individual products indicated that the chloroethyl and hydroxyethyl adducts are derived from different alkylating intermediates. The influence of cations on the formation of these DNA alkylation products was investigated by the addition of either NaCl, MgCl(2), or spermine. The results demonstrated that (1) the levels of DNA alkylation were inversely proportional to ionic strength, (2) the extent of inhibition was dependent on the alkylation product, and (3) the order of relative effectiveness of inhibition of DNA alkylation by these cations was as follows: spermine > Mg > Na. These results support a model whereby reactions which proceed via an S(N)2 mechanism are more sensitive to the effects of ionic strength than reactions which proceed via an S(N)1 mechanism. In 9L cells treated with CNU, the same alkylation products were formed as in purified DNA; however, the product distribution was different. We interpret this to indicate that within cells, cations modify the reaction of intermediates derived from CNU with DNA. PMID- 10525274 TI - Phototoxicity of naphazoline. Evidence that hydrated electrons, nitrogen-centered radicals, and OH radicals trigger DNA damage: a combined photocleavage and laser flash photolysis study. AB - The potential phototoxic activity of naphazoline (NP), 2-(1 naphthylmethyl)imidazoline, was investigated by studying its photoreactivity toward DNA. Photocleavage studies combined with laser flash photolysis experiments provide clear evidence that the transient species produced under NP photolysis react with DNA, thereby promoting its breakage under both aerobic and anaerobic conditions. Hydrated electrons and nitrogen-centered radicals are involved in the photodamage under anaerobic conditions. Hydroxyl radicals generated by Haber-Weiss reaction seem to initiate the photocleavage observed under aerobic conditions. A photodynamic mechanism involving the participation of singlet oxygen does not seem to play a crucial role in the photoinduced DNA breakage. The interaction between the NP and the biopolymer is also investigated by using both steady state and time-resolved spectroscopy. PMID- 10525275 TI - Comparison of (32)P-postlabeling and high-resolution GC/MS in quantifying N7-(2 Hydroxyethyl)guanine adducts. AB - This study compares (32)P-postlabeling and high-resolution gas chromatography/mass spectrometry (GC/MS) in the quantification of N7-(2 hydroxyethyl)guanine adducts (7-HEG) in DNA obtained from the same tissue samples of control rats and rats exposed to ethene. The samples were obtained from two independent studies. In one study, male Sprague-Dawley rats were exposed to 300 ppm ethene for 12 h/day for 3 days ("Euro samples"). In the other study, male F 344 rats were exposed to 3000 ppm ethene for 6 h/day for 5 days ("U.S. samples"). DNA from liver and kidney from the European study was isolated in the European laboratory, and DNA from liver and spleen from the U.S. study was isolated in the U.S. laboratory. The DNA samples were coded, divided into two portions, and exchanged between the two laboratories. All DNA samples from both laboratories were analyzed with respect to 7-HEG adducts by (32)P-postlabeling and high resolution GC/MS in the European and U.S. laboratories, respectively. However, the U.S. samples were repurified in the European laboratory before the postlabeling analysis. The data from the Euro and the U.S. samples were therefore treated separately in the regression analysis of the (32)P-postlabeling versus GC/MS data. The slope of the regression line for the Euro samples was 1.19 (r = 0.97), implying that the GC/MS data were slightly lower than the postlabeling data (one possible outlier was excluded). The slope of the regression line for the U.S. samples was 0.61 (r = 0.94), implying that the GC/MS data were somewhat higher than the postlabeling data. The main conclusion from this study is that there is very good agreement between the (32)P-postlabeling and high-resolution GC/MS methods in quantifying 7-HEG adducts to DNA, particularly when identical DNA samples are analyzed and the RNA content is <2%. The paper also discusses the background levels of adducts, the interorgan distribution, comparison between different strains, and exposure conditions. PMID- 10525276 TI - 5,6-Dihydroxyindoles in the fenton reaction: a model study of the role of melanin precursors in oxidative stress and hyperpigmentary processes. AB - Increasing evidence supports the view that diffusible melanin-related metabolites do not serve merely as pigment precursors, but may also act as modulators of the responses of the pigmentary cell melanocyte to external stimuli, especially to inflammation. In this study, the effect of melanin precursors 5, 6 dihydroxyindole (DHI) and 5,6-dihydroxyindole-2-carboxylic acid (DHICA) on the Fenton-induced oxidation of deoxyribose was investigated as a model of the oxidative stress processes triggered by the release of iron during inflammation. DHICA caused a powerful inhibition of the H(2)O(2)-Fe(II)/EDTA oxidation under both aerobic and anaerobic conditions, proving to be more efficient than typical hydroxyl radical (HO(*)) scavengers even at low concentrations with respect to deoxyribose. Conversely, DHI in air was a prooxidant at low indole:Fe(II) ratios, but shifted to an antioxidant at higher ratios (>6). The magnitude of the prooxidant effect increased by lowering the pH of the medium or by replacing Fe(II) with Fe(III), but was suppressed by exclusion of oxygen. Both the indoles retained their effects on the Fenton reaction in the absence of EDTA, as a result of their ability to chelate iron ions as evidenced by spectrophotometric experiments. Investigation of the reaction of DHI and DHICA with the Fenton reagent led to the conclusion that the indoles interact efficiently with HO(*), yielding indolesemiquinone species which are then converted to melanin pigments by self-coupling or disproportionation. At low DHI:iron molar ratios, the ability of semiquinones, generated by autoxidation of indoles, to recycle Fe(II) ions prevails, accounting for the observed prooxidant effect. Collectively, the results of this study provide new evidence for melanogenic 5,6-dihydroxyindoles as a novel class of biological antioxidants and point to these compounds as the key to interpreting the response of melanocytes to oxidative injuries. Moreover, the rapid formation of melanin following the exposure of 5, 6-dihydroxyindoles to the Fenton oxidation suggests new mechanisms of skin hyperpigmentation associated with inflammation. PMID- 10525278 TI - Effects of mannitol or catalase on the generation of reactive oxygen species leading to DNA damage by Chromium(VI) reduction with ascorbate. AB - Interaction of Cr(VI) and ascorbate in vitro generates Cr(V), Cr(IV), Cr(III), carbon-based alkyl radicals, COO(*)(-), (*)OH, and ascorbate radicals and induces DNA interstrand cross-links at guanines. To determine which specific Cr species and free radicals cause DNA damage, we investigated the effects of mannitol and catalase on the formation of reactive intermediates, Cr-DNA associations, DNA polymerase-stop sites, and 8-hydroxydeoxyguanosine (8-OHdG) adducts induced by Cr(VI)/ascorbate in a Hepes buffer. EPR spectra showed that mannitol trapped reactive Cr(V), forming a stable Cr(V)-diol complex, and inhibited the radicals induced by Cr(VI)/ascorbate, whereas catalase or heat-denatured catalase enhanced the levels of Cr(V) without altering the radical signals. Mannitol markedly inhibited the retarded gel electrophoretic mobility of supercoiled plasmids and the formation of DNA polymerase-stop sites induced by Cr(VI)/ascorbate, but catalase did not. On the other hand, mannitol reduced only 32% of the Cr-DNA adducts induced by Cr(VI)/ascorbate, suggesting that Cr monoadducts (possibly DNA Cr-mannitol adducts) are the major lesions generated in the Cr(VI)/ascorbate/mannitol/DNA solution. Native catalase but not heat-denatured catalase protected approximately 25% of the Cr-DNA adducts induced by Cr(VI)/ascorbate, suggesting that hydrogen peroxide may be involved. Mannitol could not completely inhibit the formation of 8-OHdG adducts induced by Cr(VI)/ascorbate, indicating that this DNA damage may be generated before the action of mannitol to trap Cr(V) and reactive oxygen species. Alternatively, Cr peroxide intermediates may also lead to 8-OHdG formation to account for the incomplete prevention by mannitol. Catalase or heat-denatured catalase partially protected the formation of 8-OHdG adducts induced by Cr(VI)/ascorbate, suggesting an effect of proteins. Together, the results from this study suggest that the primary species generated during the reduction of Cr(VI) by ascorbate are hydroxyl radicals and Cr(V) species, responsible for the formation of 8-OHdG and DNA cross-links, respectively. PMID- 10525277 TI - Maitotoxin-induced calcium influx in erythrocyte ghosts and rat glioma C6 cells, and blockade by gangliosides and other membrane lipids. AB - Maitotoxin (MTX) at 0.3 nM elicited a 10-20-fold increase in the level of Ca(2+) influx in rat glioma C6 cells. At higher doses (3-30 nM), MTX induced marked Ca(2+) influx in human erythrocyte ghosts when monitored with the fluorescent dye Fura-2. Although the ghosts were not as susceptible to MTX as intact erythrocytes or other cell lines, Fura-2 experiments under various conditions suggested that the MTX-induced entry of ions into the ghosts was mediated by a mechanism similar to that reported for cells or tissues. These ghosts are the simplest system known to be sensitive to MTX and thus may be suitable for research on the direct action of MTX. Gangliosides GM1 and GM3, glycosphingolipids which have a sialic acid residue, strongly inhibited MTX-induced Ca(2+) influx in C6 cells, while the inhibitory action by asialo-GM1, which lacks a sialic acid residue, was somewhat weaker. Their inhibitory potencies were in the following order: GM1 (IC(50) approximately 2 microM) > GM3 (IC(50) approximately 5 microM) > asialo-GM1 (IC(50) approximately 20 microM). GM1 (3 microM) completely blocked MTX (30 nM) induced Ca(2+) influx in human erythrocyte ghosts. When C6 cells were pretreated with tunicamycin, an antibiotic which inhibits N-linked glycosylation, or concanavalin A, a lectin which exhibits a high affinity for cell-surface oligosaccharides, MTX-induced Ca(2+) influx was significantly potentiated. This suggests that removal of oligosaccharides from the cell surface by tunicamycin or capping of sugar chains on plasma membranes by concanavalin A can potentiate the action of MTX. PMID- 10525279 TI - Oxidation of acetaldehyde by peroxynitrite and hydrogen Peroxide/Iron(II). Production Of acetate, formate, and methyl radicals. AB - Production of free radicals from acetaldehyde oxidation by enzymes and cellular fractions is a well-known process. The toxic effects of acetaldehyde, however, are usually attributed to its reactions with biomolecules to produce adducts. Here, we demonstrate that hypothetical adducts produced from attack of acetaldehyde by two important biological oxidants, peroxynitrite and hydrogen peroxide, decompose to produce acetate, formate, and methyl radicals. Acetate, formate, nitrate, and nitrite were characterized and quantified by capillary electrophoresis. Radicals were detected and quantified by the EPR spectra produced in the presence of spin traps 3, 5-dibromo-4-nitrosobenzenesulfonic acid and 5,5-dimethyl-1-pyrroline N-oxide. Kinetic studies and product analysis were performed at different pHs. The results demonstrate that production of methyl radicals during oxidation of acetaldehyde by hydrogen peroxide was strictly dependent on the presence of iron(II) and occurred via two routes. One involved acetaldehyde attack by the hydroxyl radical to produce the acetyl radical that decomposes to methyl radical and carbon monoxide. The other route involved acetaldehyde attack by deprotonated hydrogen peroxide to produce a hypothetical intermediate that reductively cleaves via the action of present iron(II) to produce radicals. The latter mechanism predominates in the case of peroxynitrite, but radical formation does not require metal ions. Most of the hypothetical adduct produced from acetaldehyde and peroxynitrite (k = 680 M(-)(1) s(-)(1) at pH 7.4 and 37 degrees C) decays to nitrate and regenerates the aldehyde [Uppu, R. M., et al. (1997) Chem. Res. Toxicol. 10, 1331], but about 30% of it produces acetate, formate, and methyl radicals. Part of these oxidized products result from beta-scission and 1,2-shift reactions of the 1-hydroxyethoxyl radical which, together with nitrogen dioxide, freely diffuses from the adduct (20% yields). The results provide yet another example of the metal-independent free radical reactivity of peroxynitrite and may be relevant to the toxic effects associated with heavy drinking and diabetes. PMID- 10525280 TI - Determination of in vitro- and in vivo-formed DNA adducts of 2-amino-3 methylimidazo[4,5-f]quinoline by capillary liquid chromatography/microelectrospray mass spectrometry. AB - Capillary liquid chromatography/microelectrospray mass spectrometry has been applied to the detection of deoxyribonucleoside adducts of the food-derived mutagen 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) from in vitro and in vivo sources. Constant neutral loss (CNL) and selective reaction monitoring (SRM) techniques with a triple-quadrupole mass spectrometer enabled sensitive and specific detection of IQ adducts in vitro and in animals. Detection of 1 adduct in 10(4) unmodified bases is achieved using CNL scanning detection, while the lower detection limits using SRM approach 1 adduct in 10(7) unmodified bases using 300 microg of DNA. The DNA adducts N-(deoxyguanosin-8-yl)-2-amino-3 methylimidazo[4, 5-f]quinoline (dG-C8-IQ) and 5-(deoxyguanosin-N(2)-yl)-2-amino-3 methylimidazo[4,5-f]quinoline (dG-N(2)-IQ) were detected in kidney tissues of chronically treated cynomolgus monkeys at levels and in proportions consistent with previously published (32)P-postlabeling data [Turesky, R. J., et al. (1996) Chem. Res. Toxicol. 9, 403-408]. Thus, capillary tandem LC/MS is a highly sensitive technique, which can be used to screen for DNA adducts in vivo. PMID- 10525281 TI - Mechanism-based inactivation of cytochrome P450s 1A2 and 3A4 by dihydralazine in human liver microsomes. AB - Dihydralazine is known to induce immunoallergic hepatitis. Since anti-liver microsome (anti-LM) autoantibodies found in the serum of the patients react with P450 1A2, it is suggested that dihydralazine is biotransformed into a reactive metabolite, which covalently binds to cytochrome P450 1A2 and triggers an immunological response as a neoantigen. We investigated inactivation of P450 enzymes, including P450 1A2, during the metabolism of dihydralazine to evaluate the selectivity of P450 1A2 as a catalyst and a target of dihydralazine. Human liver microsomes or microsomes from lymphoblastoid cells expressing P450 enzymes were preincubated with dihydralazine in the presence of NADPH, followed by an assay of several monooxygenase activities. Preincubation of human liver microsomes with dihydralazine in the presence of NADPH resulted in decreases in phenacetin O-deethylase activity (an indicator of P450 1A2 activity) and testosterone 6beta-hydroxylase activity (P450 3A4), but not in diclofenac 4' hydroxylase activity (P450 2C9), an indication of inactivation of P450s 1A2 and 3A4 during the dihydralazine metabolism. The inactivation of both of the P450s followed pseudo-first-order kinetics and was saturable with increasing dihydralazine concentrations. Similar time-dependent decreases in the activities were obtained in the case for use in microsomes expressing P450 1A2 and P450 3A4 instead of the human liver microsomes. The data presented here demonstrated that dihydralazine was metabolically activated not only by P450 1A2 but also by P450 3A4, and the chemically reactive metabolite bound to and inactivated the enzyme themselves, suggesting that dihydralazine is a mechanism-based inactivator of P450s 1A2 and 3A4. The data support the postulated covalent binding of a reactive metabolite of dihydralazine to P450 1A2 as a step in the formation of anti-LM antibodies in dihydralazine hepatitis, but it is not the unique factor for determining the specificity of the autoantibodies. PMID- 10525282 TI - AFM study of membrane proteins, cytochrome P450 2B4, and NADPH-cytochrome P450 reductase and their complex formation. AB - The application of the AFM technique for visualization of membrane proteins and for measuring their dimensions was demonstrated. The AFM images of the microsomal monooxygenase system components-cytochrome P450 2B4 and NADPH-cytochrome P450 reductase-were obtained by using two types of supports-hydrophobic, highly oriented pyrolytic graphite (HOPG) and hydrophilic mica. It was shown that hemo- and flavoprotein monomers and oligomers can be adsorbed to and visualized on HOPG. On the negatively charged mica matrix, flavoprotein oligomers dissociated to monomers while hemoprotein oligomers dissociated into less aggregated particles. The images of cytochrome P450 2B4 and NADPH-cytochrome P450 reductase monomers were about 3 and 5 nm high, respectively, while the images of oligomeric forms of these proteins were about 10 and 8 nm high, respectively. We were able to observe the binary complexes composed of monomeric proteins, cytochrome P450 2B4 and its reductase and to measure the heights of these complexes (7 nm). The method is applicable for visualization of not only individual proteins but also their complexes. PMID- 10525283 TI - Effect of sodium arsenite on heme metabolism in cultured chick embryo hepatocytes. AB - We had previously reported that low concentrations of sodium arsenite (1-5 microM) decreased the induction of cytochrome P450 CYP1A and CYP2H in cultured chick embryo hepatocytes in parallel with increases in heme oxygenase. However, in those studies exogenous heme did not prevent the decrease in CYPs. In this study, we investigated the effect of arsenite on the synthesis and degradation of heme. Arsenite had no effect on induction of 5-aminolevulinic acid synthase mRNA or activity. Arsenite, at concentrations from 1 to 25 microM, had no effect on protoporphyrin synthesis from 5-aminolevulinic acid and did not increase the accumulation of other porphyrins, indicating that the enzymes in the pathway between 5-aminolevulinic acid synthase and ferrochelatase were unaffected by arsenite. Synthesis of heme from radioactive 5-aminolevulinic acid was slightly decreased (less than 20%) by 2.5 microM arsenite, a concentration that decreased induction of CYP1A and CYP2H by greater than 50%. Rates of biliverdin formation and degradation of exogenous heme were not different in cultures treated simultaneously with arsenite and heme or with heme alone. However, arsenite treatment increased biliverdin formation from heme synthesized from added 5 aminolevulinic acid by 60% and decreased the endogenous heme content of the cells by 30%. Our results suggest that although 2.5 microM arsenite induced heme oxygenase four- to sixfold, this had no effect on degradation of exogenous heme. Degradation of heme synthesized from 5-aminolevulinic acid was increased but this did not affect the regulatory heme pool. PMID- 10525284 TI - Mutations in the charged residues of the amino terminus of rat liver fructose 6 phosphate,2-kinase:Fructose 2,6-bisphosphatase: effects on regulation. AB - Amino and carboxyl termini of the bifunctional enzyme Fru 6-P, 2-kinase:Fru 2,6 bisphosphatase regulate the relative activities of the kinase/phosphatase. The N terminus of the rat liver bifunctional enzyme is highly basic, containing a protein kinase A phosphorylation site that regulates these enzyme activities in a reciprocal manner. To determine the role of charged residues in the N-terminal peptide, mutant enzymes were constructed in which these residues were altered to residues carrying opposite charges, and the effect on the catalytic properties, thermal lability, and susceptibility to trypsin digestion and phosphorylation by protein kinase A was determined. Most of these mutations decreased k(cat)/K(ATP) and/or k(cat)/K(Fru) (6-P) of the kinase and increased k(cat)/K(Fru 2,6-P2) of the phosphatase. These mutant enzymes were more susceptible to trypsin digestion, phosphorylation by protein kinase A, and thermal inactivation. In general, the effect was greater with amino acid residues located more distant from the N terminus. The resulting changes were not as large as observed with the phosphorylated enzyme. Mutation of Ser22 to Pro produced large changes in the kinetic properties comparable to those of phosphorylation, suggesting that the flexible region of the N-terminus containing five serines (Ser20 to S24) is essential for the enzyme activities. These results indicated that the charged residues as well as Ser20-Ser24 in the N-terminus of the liver Fru 6-P,2 kinase:Fru 2,6-Pase are essential in the allosteric regulation and probably involved in interactions with the catalytic domains that induce a conformation that has high Fru 6-P,2-kinase and low Fru 2,6-Pase activities. Any disruption of this N-terminal interaction results in inhibition of the kinase and activation of the phosphatase. PMID- 10525285 TI - Molecular cloning and sequence analysis of cDNAs coding for 3-methylcholanthrene inducible cytochromes P450 in Xenopus laevis liver. AB - Liver microsomes of Xenopus laevis were investigated for specific cytochrome P450s (CYPs) that would be inducible in response to the administration of either 3-methylcholanthrene (3MC) or beta-naphthoflavone (BNF), potent inducers for mammalian CYP1A. When probed with antibodies raised against rat CYP1A1, a 54-kDa protein was detected after administration of polycyclic aromatic hydrocarbons. However, there was no immunoreactive protein in microsomes from untreated frogs. In order to obtain structural information about this CYP1A-like protein, a liver cDNA library of 3MC-treated frog was constructed and screened using a fragment of rat CYP1A2 cDNA under low stringency conditions. We have isolated two cDNA clones (MC1 and MC2) with inherent features of the CYP1A subfamily. The sequence determination revealed that both of them coded for polypeptides composed of 526 amino acid residues, which differed from each other by 30 amino acids. A comparison with other mammalian CYP enzymes demonstrated that both of the sequences share 55 to 63% identity with the sequences of CYP1A family members. Northern blot analysis and RT-PCR results further demonstrated that two discrete transcripts corresponding to clones MC1 and MC2 are indeed inducible in the frog liver by treatment with 3MC or BNF. The names CYP1A6 and CYP1A7 were given to clones MC1 and MC2, respectively. PMID- 10525286 TI - Failure of selenomethionine residues in albumin and immunoglobulin G to protect against peroxynitrite. AB - Selenomethionine has been suggested to protect against peroxynitrite by quenching it in vivo. Selenomethionine is distributed randomly in the methionine pool. Albumin and IgG were purified from plasma of a human being before and after 28 days of supplementation with 400 microg selenium/day as selenomethionine. The albumin contained 1 selenium atom, presumably as selenomethionine, per 8000 methionine residues before supplementation and 1 per 2800 after supplementation. Although this ratio suggested that selenomethionine would not have as great an effect in quenching peroxynitrite as would methionine, direct testing of the albumin and IgG fractions was carried out to assess the ability of these proteins to prevent peroxynitrite oxidation of dihydrorhodamine 123 to rhodamine 123. The ability of the albumin preparations to resist nitration of tyrosine residues was also assessed. The high-selenomethionine preparations of the proteins had no greater effect in quenching the peroxynitrite than did the normal selenomethionine preparations. These results do not support the proposal that selenomethionine in proteins contributes to in vivo protection against peroxynitrite. PMID- 10525287 TI - Oligomerization of the EK18 mutant of the trp repressor of Escherichia coli as observed by NMR spectroscopy. AB - The regulation of the trp repressor system of Escherichia coli is frequently modeled by a single equilibrium, that between the aporepressor (TR) and the corepressor, l-tryptophan (Trp), at their intracellular concentrations. The actual mechanism, which is much more complex and more finely tuned, involves multiple equilibria: TR and Trp association, TR oligomerization, specific and nonspecific binding of various states of TR to DNA, and interactions between these various species and ions. TR in isolation exists primarily as a homodimer, but the state of oligomerization increases as the TR concentration goes up and/or the salt concentration goes down, leading to species with lower affinity for DNA. We have used multinuclear, multidimensional NMR spectroscopy to investigate structural changes that accompany the oligomerization of TR. For these investigations, the superrepressor mutant EK18 (TR with Glu 18 replaced by Lys) was chosen because it exhibits less severe oligomerization at higher protein concentration than other known variants; this made it possible to study the dimer to tetramer oligomerization step by NMR. The NMR results suggest that the interaction between TR dimers is structurally linked to folding of the DNA binding domain and that it likely involves direct contacts between the C-terminal residues of the C-helix of one dimer with the next dimer. This implies that oligomerization can compete with DNA binding and thus serves as a factor in the fine-tuning of gene expression. PMID- 10525288 TI - Manganese and iron porphyrins catalyze peroxynitrite decomposition and simultaneously increase nitration and oxidant yield: implications for their use as peroxynitrite scavengers in vivo. AB - Twelve substituted metalloporphyrins (MPs), some of which have been previously characterized with respect to superoxide dismutase and peroxynitrite decomposing activities, were evaluated for their ability to scavenge peroxynitrite in vitro at 37 degrees C. Because the overall effectiveness of MPs as catalytic peroxynitrite scavengers is a function of (1) how fast they react with peroxynitrite, (2) how fast they cycle back to the starting compound, and (3) how well they contain or quench the reactive intermediates generated, all of these properties were evaluated and compared directly under the same conditions. Of the various MPs tested, only the iron and manganese porphyrins showed significant reactivity with peroxynitrite. The Mn(IV) intermediates resulting from oxidation by peroxynitrite were relatively stable and rereduction to the Mn(III) forms was rate-limiting to catalytic decomposition of peroxynitrite. However, in the presence of oxidizeable substrates like phenolics, rereduction of Mn(IV) forms occurred very rapidly and both the Mn- and Fe-porphyrins catalyzed nitration and oxidation by peroxynitrite. Mn- and Fe-porphyrins enhanced the yield of nitrated phenolics by peroxynitrite as much as 5-fold at pH 7.4 and up to 12-fold at pH 9. 1, while total oxidative yield was more than doubled. Nitration enhancement by MPs was effectively inhibited by ascorbate, glutathione, or serum, although much higher concentrations of ascorbate were required to inhibit nitration catalyzed by either Mn or Fe tetramethylpyridyl porphyrin. Catalysis of peroxynitrite nitration by MPs appears to proceed via a radical-mediated reaction mechanism whereby the phenolic substrate rapidly reduces Mn(IV) = O or Fe[IV] = O to the +3 state to yield phenoxyl radical which then combines with the other primary product, nitrogen dioxide. Based on the rate constants and the proposed reaction mechanism, it is reasonable to suggest that Mn and Fe porphyrins could detoxify peroxynitrite in vivo by efficiently trapping the relatively unreactive peroxynitrite anion and, in effect, channeling it into a single reaction pathway which could then be more effectively scavenged by cellular reductants like ascorbate. PMID- 10525289 TI - Changes in fructose-induced production of glucose in the rat liver following partial hepatectomy. AB - The fructose-induced production of glucose in the liver after partial hepatectomy (PH) was evaluated by using the liver-perfusion system. There was no significant difference in plasma glucose level between hepatectomized (HX) and sham-operated (SO) rats at 24 h after surgery, and, thereafter, almost similar levels were obtained in both groups. However, the level of serum free fatty acids (FFA) was significantly higher in HX rats than that in SO rats at 24 and 48 h after surgery. When both groups of rats were given fructose by gavage, the increment of plasma glucose was significantly larger in HX rats than in SO rats. Lactate infusion failed to increase the rate of glucose production in perfused livers of both HX and SO rats and there was no significant difference in the activity of hepatic phosphoenolpyruvate carboxykinase. By contrast, fructose infusion elicited a large increase in glucose production in the perfused livers of HX rats at 24 and 48 h after PH. The increase was closely associated with not the change in fructose 2,6-bisphosphate levels but the increment of the intracellular levels of citrate. Treatment of octanoate or oleate, which supplies acetyl-CoA via fatty acid oxidation, mimicked the fructose-induced increase in glucose production in SO rats with a concomitant increase in hepatic levels of citrate. These results suggest that the oxidation of FFA may play an important role in glucose production induced by fructose administration during the early phase of liver regeneration. PMID- 10525290 TI - Fluorescence properties of melanins from opioid peptides. AB - Recently our group synthesized a new class of melanins obtained by the tyrosinase catalyzed oxidation of opioid peptides (opiomelanins). Owing to the presence of the peptide moiety such pigments exhibit high solubility in hydrophilic solvents, which allows spectroscopic investigations. In particular, the absence of solid state quenching effects enables the study of melanin fluorescence properties, till now poorly investigated due to the complete insolubility of melanins produced from tyrosine or Dopa. Opiomelanins dissolved in aqueous medium show a characteristic emission peaked at 440 and 520 nm when excited around 330 nm, where a maximum is observed in the absorption spectrum. Kinetic measurements performed on the tyrosinase-catalyzed oxidation of opioid peptides show that the 440-nm fluorescence band arises in the early stages of peptide oxidation, whereas the 520-nm band appears in later stages of oxidation, i.e., during the polymerization of indole-quinone units. Moreover, molecular sieve fractionation shows that in the opiomelanin fraction with a molecular weight lower than 10 kDa the 440-nm band is dominant in the fluorescence spectrum. The breakdown of the polymer induced by hydrogen peroxide and light (i.e., the photobleaching of melanin pigments) produces a marked enhancement of the 440-nm fluorescence band while the 520-nm band disappears. Hence, our findings suggest that the observed fluorescence contains contributions from both oligomeric units (440-nm band) and high-molecular-weight polymers (520-nm band). PMID- 10525291 TI - Soybean nodule sucrose synthase (nodulin-100): further analysis of its phosphorylation using recombinant and authentic root-nodule enzymes. AB - Sucrose synthase (SS) is a known phosphoserine-containing enzyme in legume root nodules and various other plant "sink" tissues. In order to begin to investigate the possible physiological significance of this posttranslational modification, we have cloned a full-length soybean nodule SS (nodulin-100) cDNA and overexpressed it in Escherichia coli. Authentic nodule SS and recombinant wild type and mutant forms of the enzyme were purified and characterized. We document that a conserved serine near the N-terminus (Ser(11)) is the primary phosphorylation site for a nodule Ca(2+)-dependent protein kinase (CDPK) in vitro. Related tryptic digestion and mass spectral analyses indicated that this target residue was also phosphorylated in planta in authentic nodulin-100. In addition, a secondary phosphorylation site(s) in recombinant nodule SS was implicated given that all active mutant enzyme forms (S11A, S11D, S11C, and N terminal truncation between Ala(2) and Arg(13)) were phosphorylated, albeit weakly, by the CDPK. This secondary site(s) likely resides between Glu(14) and Met(193) as evidenced by CNBr cleavage and phosphopeptide mapping. Phosphorylation of the recombinant and authentic nodule Ser(11) enzymes in vitro by the nodule CDPK had no major effect on the sucrose-cleavage activity and/or kinetic properties. However, phosphorylation decreased the apparent surface hydrophobicity of the recombinant wild-type enzyme, suggesting that this covalent modification could potentially play some role in the documented partitioning of nodulin-100 between the nodule symbiosome/plasma membranes and cytosol in planta. PMID- 10525292 TI - Chemical synthesis of biotinylated histones and analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis/streptavidin-peroxidase. AB - Recently, Hymes and co-workers demonstrated that human biotinidase (EC 3.5.1.12) specifically biotinylates histones, suggesting that biotin may have a specific role in transcription and replication of DNA. In the present study, we sought to biotinylate histones in vitro for later use as standards in the quantitation of histones biotinylated in vivo. We also sought to develop a procedure for electrophoretic separation and streptavidin-peroxidase detection of the various classes of biotinylated histones. Histones H1, H2a, H2b, H3, and H4 from calf thymus were biotinylated using sulfosuccinimidobiotin at pH 7.5. Stoichiometries of biotin/histone were determined either by 4'-hydroxyazobenzene-2-carboxylic acid/avidin assay or by avidin-binding assay. The stoichiometries of biotinylation (mol biotin/mol histone) were as follows: H1, 3.9 +/- 0.17; H2a, 1.7 +/- 0.11; H2b, 1.8 +/- 0.11; H3, 0.029 +/- 0.0012; H4, 0.006 +/- 0.0002. When two synthetic polypeptides were used as substrates for biotinylation, the stoichiometry of poly-l-lysine was 2.8 +/- 0.14 mol biotin/mol; in contrast, the stoichiometry of poly-l-arginine was less than 0.3 x 10(-3) mol biotin/mol. These data suggest that primary amino groups of histones biotinylated by sulfosuccinimidobiotin were lysine rather than arginine. Detection and identification of biotinylated histones were accomplished by electrophoretic separation on 16% polyacrylamide gels; the separated histones on nitrocellulose transblots of the gels were detected using streptavidin-peroxidase with 4-chloro 1-naphthol as the substrate. We conclude that sulfosuccinimidobiotin does biotinylate each of the five classes of histones and that the stoichiometry of biotinylation is sufficient for detection on nitrocellulose transblots by streptavidin-peroxidase. PMID- 10525293 TI - 5-Doxylstearate-induced displacement of phencyclidine from its low-affinity binding sites on the nicotinic acetylcholine receptor. AB - Fatty acids as well as phencyclidine (PCP) inhibit the ion channel activity of the nicotinic acetylcholine receptor (AChR) by a noncompetitive mechanism. However, the exact localization of the fatty acid binding sites is unknown and, thus, the noncompetitive inhibitory mechanism for these endogenous modulators remains to be elucidated. In an attempt to determine the location of the fatty acid binding sites, we study the mutually exclusive action between 5 doxylstearate (5-SASL), a derivative of the endogenous noncompetitive antagonist (NCA) stearic acid, and other exogenous NCAs. For this purpose, both equilibrium and competitive binding assays using fluorescent and radiolabeled ligands were performed on desensitized AChRs. More specifically, we determined: (i) the effect of 5-SASL on the binding of the exogenous NCA [(3)H]PCP; (ii) the effect of 5 SASL on the binding of either quinacrine or ethidium, two fluorescent NCAs from exogenous origin; and (iii) the PCP-induced displacement of quinacrine and ethidium from their respective high-affinity binding sites. Our first target (i) is carried out by measuring the [(3)H]PCP binding in the absence or in the presence of increasing concentrations of 5-SASL. We found that 5-SASL displaces PCP from its low-affinity binding sites. The low-affinity PCP binding sites were pharmacologically characterized by an apparent dissociation constant (K(d)) of 6.1 +/- 5.0 microM and a stoichiometry of 3.7 +/- 1.5 sites per AChR. The fact that 5-SASL increased the apparent K(d) without changing the number of sites per AChR is indicative of a mutually exclusive action. From these results, an apparent inhibition constant (K(i)) of 75 +/- 31 microM for 5-SASL was calculated. In addition, 5-SASL affected neither the apparent K(d) (0.46 +/- 0.37 microM) nor the stoichiometry (1.07 +/- 0.57 sites per AChR) of the high-affinity PCP binding site. The second objective (ii) is achieved by titrating either quinacrine or ethidium into AChR native membranes in the absence or in the presence of increasing concentrations of 5-SASL. These experiments showed that 5 SASL efficiently increased the apparent K(d) of quinacrine without perturbing the interaction of ethidium with its high-affinity locus. Considering that (a) 5-SASL effectively quenched the AChR-bound quinacrine fluorescence (H. R. Arias, Biochim. Biophys. Acta 1347, 9-22, 1997) and (b) fluorescence-quenching is a short-range process, it is possible to suggest that 5-SASL displaces quinacrine from its high-affinity binding site by a steric mechanism. In this regard, a K(i) of 38 +/- 5 microM for 5-SASL was calculated. Concerning the last objective (iii), AChR-bound quinacrine or ethidium was back titrated with PCP. Two PCP K(i) values were obtained by fitting the displacement plots by nonlinear regression with two components. The lowest K(i) values obtained for either quinacrine (0.86 +/- 0.37 microM) or ethidium (0. 29 +/- 0.23 microM) displacement from their respective high-affinity binding sites coincide with the previously determined high-affinity [(3)H]PCP K(d). In addition, the highest K(i) values obtained for either NCA displacement are in the same concentration range as the observed low affinity [(3)H]PCP K(d). Taking into account all experimental data, we reached the following conclusions: (i) fatty acid molecules, or at least 5-SASL, sterically interact with both the PCP low-affinity and the quinacrine high affinity binding sites; (ii) the low-affinity PCP binding sites, as well as the high-affinity quinacrine locus, are located at the nonannular lipid domain of the AChR; and, finally, (iii) fatty acid molecules are not accessible to the lumen of the ion channel, indicating an allosteric mode of action for fatty acids to inhibit ion flux. Thus, the 5-SASL, the quinacrine high-affinity, and the PCP low affinity binding sites are all located at overlapping nonannular loci on the muscle-type AChR. PMID- 10525294 TI - Formation of a new quinone methide intermediate during the oxidative transformation of 3,4-dihydroxyphenylacetic acids: implication for eumelanin biosynthesis. AB - Oxidation of dopa and dopamine requires a net removal six electrons to produce indolequinones, the monomeric precursors of eumelanin pigment. On the other hand, their 6-fluoroderivatives suffer only four-electron oxidation to yield the same products (M. E. Rice, B. Mogaddam, C. R. Creveling, and K. L. Kirk, Anal. Chem. 59, 1534-1536, 1987). Taking advantage of this novel fluorochemistry, we reexamined the oxidative mechanism of 3,4-dihydroxyphenylacetic acid and 6-fluoro 3,4-dihydroxyphenylacetic acid to throw more light on the nature of reactive intermediates formed during the reaction. Enzymatic or chemical oxidation of 3,4 dihydroxyphenylacetic acid generated the transient o-quinone which exhibited rapid intramolecular cyclization and side chain modification to produce 2, 5,6 trihydrobenzofuran and 3,4-dihydroxymandelic acid, respectively. However, when 6 fluoro-3,4-dihydroxyphenylacetic acid was oxidized either by tyrosinase or by sodium periodate, the resultant quinone uniquely exhibited only cyclization coupled with loss of fluoride ion. This clean reaction allowed us to establish the structures of the transient reactive intermediates. Two interconvertable isomeric forms of the product were isolated and characterized from the reaction mixture. If the oxidation was carried out in water, a yellow quinolactone accumulated in the reaction mixture. This compound was instantaneously converted to a purple quinone methide upon addition of a trace amount of sodium phosphate. Passage through a C(18) HPLC column caused the reverse transformation. The structures of these products were established by semiempirical molecular orbital calculations and NMR spectrometry. Comparison of the oxidation mechanisms of melanin precursors, dopa and dopamine, with that of 3,4-dihydroxyphenylacetic acids reveals that a similar quinone methide intermediate is likely to be formed during eumelanin biosynthesis. PMID- 10525295 TI - Consecutive halogen transfer between various functional groups induced by reaction of hypohalous acids: NADH oxidation by halogenated amide groups. AB - Cyclic dipeptides (c-Gly(2), c-Ser(2), c-Gly-Phe, etc.) were used as simple protein models to investigate the HOCl-induced generation and reactivity of chlorinated amide groups. The pH dependence of the kinetics of amide chlorination reveals that ClO(-) (not HOCl) is the reactive agent. N-Chlorinated cyclopeptides are stable up to 30 min, they exhibit narrow absorption bands around 215 nm, and they are capable of oxidizing certain biological substrates, the reactivity decreasing in the order GSH > ascorbate > methionine > NADH >> GSSG. The chloroamide is less reactive, but much more selective in its reactions, than HOCl or ClO(-); thus, with formation of the chloroamide prolonged oxidative effects, directed toward specific target molecules, can be expected. Chlorination of NADH, yielding a catalytically inactive species (NAD/Cl), was investigated in most detail because it is likely to be an important and highly lethal process. The chloroamide group is far more reactive toward NADH than chloroamines derived from primary amines. Chloronucleotides formed by reaction of ClO(-) with inosine, GMP, TMP, or UMP are capable of quantitative chlorine transfer to cyclopeptides; however, no chlorine transfer between the amide nitrogen and primary amines is detectable, in either direction. The results presented enable prediction of chlorine transfer cascades induced by HOCl/ClO(-), involving nucleotides, peptide amide groups, and final target molecules. Chlorinated NAD(P)H, as a stable terminal product of consecutive chlorine transfer reactions, might be a useful biological marker for assessing the role of HOCl in inflammatory events. Bromination by BrO(-) of cyclopeptides is more than two orders of magnitude faster than chlorination by ClO(-), and the reactivity of bromoamide with NADH exceeds that of chloroamide by more than four orders of magnitude. PMID- 10525296 TI - Tetrameric N(5)-(L-1-carboxyethyl)-L-ornithine synthase: guanidine. HCl-induced unfolding and a low temperature requirement for refolding. AB - Guanidine x HCl (GdnHCl)-induced unfolding of tetrameric N(5)-(L-1-carboxyethyl) L-ornithine synthase (CEOS; 141,300 M(r)) from Lactococcus lactis at pH 7.2 and 25 degrees C occurred in several phases. The enzyme was inactivated at approximately 1 M GdnHCl. A time-, temperature-, and concentration-dependent formation of soluble protein aggregates occurred at 0.5-1.5 M GdnHCl due to an increased exposure of apolar surfaces. A transition from tetramer to unfolded monomer was observed between 2 and 3.5 M GdnHCl (without observable dimer or trimer intermediates), as evidenced by tyrosyl and tryptophanyl fluorescence changes, sulfhydryl group exposure, loss of secondary structure, size-exclusion chromatography, and sedimentation equilibrium data. GdnHCl-induced dissociation and unfolding of tetrameric CEOS was concerted, and yields of reactivated CEOS by dilution from 5 M GdnHCl were improved when unfolding took place on ice rather than at 25 degrees C. Refolding and reconstitution of the enzyme were optimal at NOD-E and NOD-E-->NOD chimeras have elevated levels of IL-4 compared to NOD-->NOD and NOD-->NOD-E chimeras in the thymus. However, in the periphery the protected NOD-E-->NOD-E show much higher IL-4 levels than any of the other chimeras. This drop in peripheral IL-4 production seen in NOD-E- >NOD, NOD-->NOD-E and NOD-->NOD chimeras correlates with the increased insulitis seen in these mice compared to NOD-E-->NOD-E. In contrast, there were no differences in IFN-gamma production between the chimeras. We suggest that the precommitted, regulatory T cells, selected in an E-expressing thymic environment, need continuous interaction with E-expressing primary antigen presenting cells in the periphery for optimal IL-4 production. Decrease in IL-4 production correlates with increased insulitis. PMID- 10525317 TI - Flow cytometric detection of type 1 (IL-2, IFN-gamma) and type 2 (IL-4, IL-5) cytokines in T-helper and T-suppressor/cytotoxic cells in rheumatoid arthritis, allergic asthma and atopic dermatitis. AB - Type 1 cytokines (a.o. IL-2 and IFN-gamma) play an important role in the pathogenesis of rheumatoid arthritis. On the other hand, IgE-mediated diseases such as allergic asthma and atopic dermatitis show a type 2 cytokine (amongst others IL-4 and IL-5) profile. This study examined simultaneously the intracellular production of IL-2, IFN-gamma, IL-4 and IL-5 in T-lymphocytes of patients with rheumatoid arthritis during treatment with methotrexate or salazopyrin, patients with allergic asthma or atopic dermatitis under stable treatment, compared to healthy controls.A three-colour flow cytometric analysis was used for cytokine detection in T-helper cells and T-suppressor/cytotoxic cells. Compared to controls, patients with symptomatic atopic dermatitis showed an increased number of IL-4-producing T-helper lymphocytes in basal circumstances (P=0.001), in contrast to asymptomatic allergic asthma patients. Compared to controls, rheumatoid arthritis patients, treated with salazopyrin, showed an increased number of IL-2-producing T-helper and T-suppressor/cytotoxic lymphocytes after in vitro stimulation with PMA and ionomycin (P=0.01). In contrast, rheumatoid arthritis patients, treated with methotrexate, a more potent disease modifying drug, did not show this type 1 cytokine profile. A positive correlation was found between the number of IFN-gamma producing T-helper cells and disease activity (Ritchie Index and number of swollen joints) in both rheumatoid arthritis patient groups. Active atopic dermatitis patients showed a type 2 cytokine profile, whereas stable asthma patients with lower disease activity did not show a predominance of type 2 cytokines. Rheumatoid arthritis patients under treatment with salazopyrin had a type 1 cytokine profile, which could not be demonstrated in patients treated with methotrexate. This imbalance between type 1 and type 2 cytokines in different immune mediated disorders can be related with treatment and the grade of disease activity. These results stress the need for further investigation of the influence of therapy on cytokine profiles. PMID- 10525318 TI - Induced heteroduplex genotyping of TNF-alpha, IL-1beta, IL-6 and IL-10 polymorphisms associated with transcriptional regulation. AB - We describe the construction and use of 7 induced heteroduplex generators, reagents for the rapid and unequivocal genotyping of nucleotide sequence polymorphism in TNF-alpha, IL-1beta, IL-6 and IL-10. Polymorphisms detected are those previously associated with regulation of gene transcription: TNF-alpha positions -308 and -238; IL-1beta position +3953; IL-6 position -174; and IL-10 positions -1082, -819 and -592. The reagents were used for analysis of allele and haplotype frequencies in a population of healthy Caucasian volunteer blood donors. PMID- 10525319 TI - Endotoxin tolerance in rats: expression of TNF-alpha, IL-6, IL-10, VCAM-1 AND HSP 70 in lung and liver during endotoxin shock. AB - Endotoxin can induce a state of tolerance against its own pathological effects, commonly referred to as endotoxin tolerance. This phenomenon has been found to be associated with reduced serum levels of cytokines such as TNF-alpha, IL-1, IL-6 and IL-10. In the present study the expression of TNF-alpha, IL-6, IL-10, the adhesion molecule VCAM-1 and the heat shock protein 70 was determined in vivo in lung and liver of LPS-tolerant and naive rats by means of semiquantitative RT-PCR after i.v. LPS injection. TNFalpha, IL-6, IL-10, HSP 70 and VCAM-1 were induced in lung and liver after LPS injection. In liver and lung of endotoxin-tolerant rats TNF-alpha and IL-6 were induced to a lower degree after LPS treatment when compared to non-tolerant controls. The LPS-induced IL-10 expression was also slightly attenuated in the lung of tolerant rats, but in the liver no differences between tolerant and non-tolerant animals were observed. HSP 70 and VCAM-1 were expressed after systemic LPS treatment in liver and lung. The degree of induction, however, was the same in tolerant and untreated controls. The presented data show that endotoxin tolerance is reflected by a reduced cytokine expression in lung and liver in vivo. On the other hand, levels of expression of the adhesion molecule VCAM-1 and the stress protein HSP 70 do not appear to be changed by endotoxin tolerance. PMID- 10525320 TI - Impaired production of IL-12 in systemic lupus erythematosus. III: deficient IL 12 p40 gene expression and cross-regulation of IL-12, IL-10 and IFN-gamma gene expression. AB - Interleukin 12 (IL-12) is a heterodimer comprising p35 and p40 subunits which are encoded and regulated separately. The authors previously demonstrated deficient IL-12 production in SLE which correlates negatively with disease activity. The present study was designed to determine whether deficiency of IL-12 and excess production of IL-10 and IL-6 in systemic lupus erythematosus (SLE) are due to aberrant regulation at the gene level. Using semiquantitative RT-PCR assay, it was shown that constitutive expression of IL-12 p35 gene is somewhat impaired in SLE compared with controls and that IL-12 p40 mRNA, which was present at low levels in controls, was undetectable in unstimulated SLE peripheral blood mononuclear cells (PBMC). Gene expression of IL-12 p35 and p40 was significantly increased in response to SAC, with significantly lower SAC-induced expression of p40 in SLE patients than controls. SAC-stimulated IL-12 p35 and p40 mRNAs were significantly augmented by interferon gamma (IFN-gamma). Exogenous IL-12 or IFN gamma significantly inhibited IL-10 gene expression, without affecting IL-6 mRNA or other proinflammatory cytokine mRNA levels. These observations were further confirmed by studies of protein production at the single cell level using ELISPOT assay. Downregulation of IL-12 p40 expression appears to be the cause of IL12 p70 deficiency in SLE. If this defect could be repaired, normalization of IL-12 and IFN-gamma production should reduce excessive IL-10 and prevent pathology. PMID- 10525332 TI - Intracellular fate mapping in a basal metazoan, the ctenophore Mnemiopsis leidyi, reveals the origins of mesoderm and the existence of indeterminate cell lineages. AB - Ctenophores are marine invertebrates that develop rapidly and directly into juvenile adults. They are likely to be the simplest metazoans possessing definitive muscle cells and are possibly the sister group to the Bilateria. All ctenophore embryos display a highly stereotyped, phylum-specific pattern of development in which every cell can be identified by its lineage history. We generated a cell lineage fate map for Mnemiopsis leidyi by injecting fluorescent lineage tracers into individual blastomeres up through the 60-cell stage. The adult ctenophore body plan is composed of four nearly identical quadrants organized along the oral-aboral axis. Each of the four quadrants is derived largely from one cell of the four-cell-stage embryo. At the eight-cell stage each quadrant contains a single E ("end") and M ("middle") blastomere. Subsequently, micromeres are formed first at the aboral pole and later at the oral pole. The ctene rows, apical organ, and tentacle apparatus are complex structures that are generated by both E and M blastomere lineages from all four quadrants. All muscle cells are derived from micromeres born at the oral pole of endomesodermal precursors (2M and 3E macromeres). While the development of the four quadrants is similar, diagonally opposed quadrants share more similarities than adjacent quadrants. Adult ctenophores possess two diagonally opposed endodermal anal canals that open at the base of the apical organ. These two structures are derived from the two diagonally opposed 2M/ macromeres. The two opposing 2M/ macromeres generated a unique set of circumpharyngeal muscle cells, but do not contribute to the anal canals. No other lineages displayed such diagonal asymmetries. Clones from each blastomere yielded regular, but not completely invariant patterns of descendents. Ectodermal descendents normally, but not always, remained within their corresponding quadrants. On the other hand, endodermal and mesodermal progeny dispersed throughout the body. The variability in the exact complements of adult structures, along with previously published cell deletion experiments, demonstrates that cell interactions are required for normal cell fate determination. Ctenophore embryos, like those of many bilaterian phyla (e.g., spiralians, nematodes, and echinoids), display a highly stereotyped cleavage program in which some, but not all, blastomeres are determined at the time of their birth. The results suggest that mesodermal tissues originally evolved from endoderm tissue. PMID- 10525333 TI - Sequential steps in synaptic targeting of sensory afferents are mediated by constitutive and developmentally regulated glycosylations of CAMs. AB - Sensory afferents in the leech are labeled with both constitutive and developmentally regulated glycosylations (markers) of their cell adhesion molecules (CAMs). Their constitutive mannose marker, recognized by Lan3-2 monoclonal antibody (mAb), mediates the formation of their diffuse central arbors. We show that, at the ultrastructural level, these arbors consist of large, loosely organized axons rich with filopodia and synaptic vesicles. Perturbing the mannose-specific adhesion of this first targeting step leads to a gain in cell-cell contact but a loss of filopodia and synaptic vesicles. During the second targeting step, galactose markers divide afferents into different subsets. We focus on the subset labeled by the marker recognized by Laz2-369 mAb. Initially, the galactose marker appears where afferents contact central neurons. Subsequently it spreads proximally and distally, covering the entire afferent surface. Afferents now gain cell-cell contact, with central neurons and self similar afferents, but lose filopodia and synaptic vesicles. Extant synaptic vesicles prevail where afferents are apposed to central neurons. These neurons develop postsynaptic densities and en passant synapses are forming. Perturbing the galactose-specific adhesion of this second targeting step causes a loss of cell-cell contact but a gain in filopodia and synaptic vesicles, essentially returning afferents to the first targeting step. The transformation of afferent growth, progressing from mannose- to galactose-specific adhesion, is consistent with a change from cell-matrix to cell-cell adhesion. By performing opposing functions in a temporal sequence, constitutive and developmentally regulated glycosylations of CAMs collaborate in the synaptogenesis of afferents and the consolidation of self-similar afferents. PMID- 10525334 TI - In vivo regulation of somite differentiation and proliferation by Sonic Hedgehog. AB - In vertebrates, somite differentiation is mediated in part by Sonic Hedgehog (Shh), secreted by the notochord and the floor plate. However, Shh-null mice display close to normal expression of molecular markers for dermomytome, myotome, and sclerotome, indicating that Shh might not be required for their initial induction. In this paper, we have addressed the capacity of Shh to regulate in vivo the expression of the somite differentiation markers Pax-1, MyoD, and Pax-3 after separation of paraxial mesoderm from axial structures. We show that Pax-1, which is lost under these experimental conditions, is rescued by Shh. In contrast, Shh maintains, but cannot induce MyoD expression, while Pax-3 expression is independent of the presence of axial structures or Shh. Finally, we demonstrate that Shh is a potent mitogen for somitic cells, supporting the idea that it may serve to expand subpopulations of cells within the somite. PMID- 10525335 TI - Spatially regulated translation in embryos: asymmetric expression of maternal Wnt 11 along the dorsal-ventral axis in Xenopus. AB - Transition from symmetry to asymmetry is a central theme in cell and developmental biology. In Xenopus embryos, dorsal-ventral asymmetry is initiated by a microtubule-dependent cytoplasmic rotation during the first cell cycle after fertilization. Here we show that the cytoplasmic rotation initiates differential cytoplasmic polyadenylation of maternal Xwnt-11 RNA, encoding a member of the Wnt family of cell-cell signaling factors. Translational regulation of Xwnt-11 mRNA along the dorsal-ventral axis results in asymmetric accumulation of Xwnt-11 protein. These results demonstrate spatially regulated translation of a maternal cell-signaling factor along the vertebrate dorsal-ventral axis and represent a novel mechanism for Wnt gene regulation. Spatial regulation of maternal RNA translation, which has been established in invertebrates, appears to be an evolutionarily conserved mechanism in the generation of intracellular asymmetry and the consequential formation of the multicellular body pattern. PMID- 10525336 TI - Flik, a chick follistatin-related gene, functions in gastrular dorsalisation/neural induction and in subsequent maintenance of midline Sonic hedgehog signalling. AB - We have targetted the chick gene Flik with antisense oligodeoxynucleotide treatment at gastrular stages, when it is expressed in organiser-derived structures of the midline (K. Patel et al., 1996, Dev. Biol. 178, 327-342). A specific syndrome of deficient axial patterning and holoprosencephaly is produced. Most aspects of this syndrome can be understood as due to attenuation of dorsalising and neural-inducing signals during gastrulation, followed by failure to maintain the later signals from chordamesoderm/neural midline that pattern the mesodermal and neural cross sections during subsequent stages. Anatomical effects are first apparent at early neurula stages and correspond with what might be expected from a reduced counteraction of the ventralising Bone morphogenetic protein (BMP) pathway at the earlier stages, coupled with inadequate Sonic hedgehog (Shh) signalling subsequently. Delay in the clearing of BMP-4 RNA expression from the presumptive neural region at gastrulation is indeed seen, though chordin RNA expression within organiser derivatives remains normal. Subsequently, specific attenuation of chordamesoderm and neural midline Shh expression is observed. Brief preincubation of stage 4 chick blastoderms in supernatant from Xenopus oocytes that have been injected with Flik RNA prolongs and enhances the competence of their peripheral epiblast to respond to neural inductive signals from grafted Hensen's nodes. This effect specifically mimics that recently observed using microg/ml solutions of recombinant Follistatin (D. J. Connolly et al., 1999, Int. J. Dev. Biol., in press), further suggesting that Flik protein might act in vivo by somehow modulating activity of signalling pathways through BMP or other TGFbeta-related ligands. We discuss the significance of the observations in relation to recent ideas about neural induction, about possible redundancy in gene action, and about subsequent patterning of the axial cross section, suggesting that a Flik function in autocrine/paracrine maintenance of later midline Shh signalling represents a role of the gene separate from that in primary dorsalisation/neural induction. PMID- 10525337 TI - The early expression control of Xepsin by nonaxial and planar posteriorizing signals in Xenopus epidermis. AB - The control mechanism of the anteroposterior axis specification in Xenopus epidermis was studied by comparing the expression of a novel anterior marker, Xepsin, with that of a panepidermal marker, type I keratin. Xepsin mRNA, which encodes a novel Xenopus serine protease, is transcribed zygotically with the expression peak in neurula stages. In normal development, its expression is limited to the anterior and anterior-dorsal portions within epidermis during neurula and tailbud stages, respectively. In UV-irradiated ventralized embryos (dorsoanterior index, DAI 0 and 1), an expression boundary for Xepsin is apparently formed within the epidermis. In contrast, Xepsin expression was observed throughout the epidermis in LiCl-treated dorsalized embryos (DAI 10), as seen from an expression pattern indistinguishable from that of type I keratin. These data suggest that posteriorizing signals which suppress the transcription of Xepsin are present in nonaxial regions and absent in the anterior dorsal mesoderm. That posteriorizing signals were present in nonaxial regions was also supported by a conjugation experiment in which Xepsin expression was suppressed in ectodermal explants conjugated with lateral or ventral marginal zone. Moreover, the partly suppressed expression of Xepsin in the epidermal region of exogastrulae indicates that the signals may travel horizontally within the plane of the epidermis. We also present data showing that both treatment with retinoic acid and the overexpression of a constitutively active form of a retinoic acid receptor caused the suppression of Xepsin mRNA transcription, suggesting that anterior-posterior patterning in the central nervous system and in the epidermis may share common endogenous factors, i.e. , retinoids, in the Xenopus embryo. PMID- 10525338 TI - Role of myosin VI in the differentiation of cochlear hair cells. AB - The mouse mutant Snell's waltzer (sv) has an intragenic deletion of the Myo6 gene, which encodes the unconventional myosin molecule myosin VI (K. B. Avraham et al., 1995, Nat. Genet. 11, 369-375). Snell's waltzer mutants exhibit behavioural abnormalities suggestive of an inner ear defect, including lack of responsiveness to sound, hyperactivity, head tossing, and circling. We have investigated the effects of a lack of myosin VI on the development of the sensory hair cells of the cochlea in these mutants. In normal mice, the hair cells sprout microvilli on their upper surface, and some of these grow to form a crescent or V shaped array of modified microvilli, the stereocilia. In the mutants, early stages of stereocilia development appear to proceed normally because at birth many stereocilia bundles have a normal appearance, but in places there are signs of disorganisation of the bundles. Over the next few days, the stereocilia become progressively more disorganised and fuse together. Practically all hair cells show fused stereocilia by 3 days after birth, and there is extensive stereocilia fusion by 7 days. By 20 days, giant stereocilia are observed on top of the hair cells. At 1 and 3 days after birth, hair cells of mutants and controls take up the membrane dye FM1-43, suggesting that endocytosis occurs in mutant hair cells. One possible model for the fusion is that myosin VI may be involved in anchoring the apical hair cell membrane to the underlying actin-rich cuticular plate, and in the absence of normal myosin VI this apical membrane will tend to pull up between stereocilia, leading to fusion. PMID- 10525339 TI - Oocyte regulation of kit ligand expression in mouse ovarian follicles. AB - Kit ligand (KL), a product of granulosa cells in ovarian follicles, is a putative regulator of oocyte development. However, the factors that regulate KL mRNA levels in granulosa cells remain unclear. This study tested the hypothesis that oocytes regulate granulosa cell steady-state KL mRNA expression levels and that the characteristics of this regulation are dependent on the stage of growth and development of both oocytes and follicles. Levels of mRNA for the KL splice variants (KL-1 and KL-2) were shown to be high in granulosa cells from preantral follicles and then decline after follicular antrum formation. Preovulatory follicular development was associated with a dramatic increase in steady-state levels of KL-1 mRNA in mural granulosa but not cumulus cells. Regulation of these changes was examined in vitro using partly grown oocytes isolated from preantral follicles and fully grown oocytes isolated from preovulatory follicles. FSH increased the steady-state KL mRNA levels in preantral granulosa cells in vitro. Partly grown oocytes either increased or decreased KL mRNA levels in preantral granulosa cells depending on the absence or presence of FSH stimulation, respectively. Fully grown oocytes reduced the KL mRNA level in preantral granulosa cells and increased the ratio of KL-1 to KL-2 mRNA. In mural granulosa cell culture, FSH augmented testosterone-dependent elevation of the steady-state KL mRNA level, but had no effect alone. Fully grown oocytes reduced KL-2 but not KL-1 mRNA levels in mural granulosa cells treated with testosterone plus FSH, whereas fully grown oocytes reduced levels of both KL transcripts in cumulus cell culture. These effects of oocytes on steady-state KL mRNA expression levels in vitro explain the changes in granulosa cell KL mRNA levels observed during follicle development in vivo. The results therefore support the hypothesis that oocytes regulate granulosa cell kit ligand mRNA levels in a way that is characteristic of the stage of growth and development of the oocyte. Moreover, the results suggest that oocytes play a major role in promoting dynamic changes in gene expression by granulosa cells appropriate to the stage of follicular development. PMID- 10525340 TI - Xenopus Smad4beta is the co-Smad component of developmentally regulated transcription factor complexes responsible for induction of early mesodermal genes. AB - Smad4 is defined as the common-mediator Smad (co-Smad) required for transducing signals for all TGF-beta superfamily members. This paper describes two Smad4s in Xenopus: XSmad4alpha, which is probably the Xenopus orthologue of human Smad4, and a distinct family member, XSmad4beta, which differs primarily at the extreme N-terminus and in the linker region. Both XSmad4s act as co-Smads, forming ligand dependent complexes with receptor-regulated Smads 1 and 2 and synergizing with them to activate transcription of mesodermal genes in Xenopus embryos. The two XSmad4 genes have reciprocal temporal expression patterns in Xenopus embryos and are expressed in varying ratios in adult tissues, suggesting distinct functional roles in vivo. XSmad4beta is the predominant maternal co-Smad and we go on to demonstrate its role in the transcriptional regulation of early mesodermal genes. We have identified two distinct nuclear complexes that bind the activin responsive element of the Xenopus Mix.2 promoter: one formed in response to high levels of activin signaling and the other activated by endogenous signaling pathways. Using specific antisera we demonstrate the presence of endogenous XSmad4beta and also XSmad2 in both of these complexes, and our data indicate that the DNA-binding components of the complexes are different. Furthermore, we show that the presence of these complexes in the nucleus perfectly correlates with the transcriptional activity of the target gene, Mix.2, and we show that one of the XSmad4beta-containing transcription factor complexes undergoes a developmentally regulated nuclear translocation. PMID- 10525341 TI - Production and design of more effective avian replication-incompetent retroviral vectors. AB - Retroviral vectors have been invaluable tools for studies of development in vertebrates. Their use has been somewhat constrained, however, by the low viral titers typically obtained with replication-incompetent vectors, particularly of the avian type. We have addressed this problem in several ways. We optimized the transient production of avian replication-incompetent viruses in a series of cell lines. One of the optimal cell lines was the mammalian line 293T, which was surprising in light of previous reports that avian viral replication was not supported by mammalian cells. We also greatly increased the efficiency of viral infection. Pseudotyping with the vesicular stomatitus virus G (VSV-G) protein led to an over 350-fold increase in the efficiency of infection in ovo relative to infection with virus particles bearing an avian retroviral envelope protein. To further increase the utility of the system, we developed new Rous sarcoma virus (RSV)-based replication-incompetent vectors, designed to express a histochemical marker gene, human placental alkaline phosphatase, as well as an additional gene. These modified retroviral vectors and the VSV-G pseudotyping technique constitute significant improvements that allow for expanded use of avian replication incompetent viral vectors in ovo. PMID- 10525342 TI - Autoregulation of the Drosophila disconnected gene in the developing visual system. AB - The Drosophila disconnected (disco) gene is required for the formation of appropriate connections between the larval optic nerve and its target cells in the brain. The disco gene encodes a nuclear protein with two zinc fingers, which suggests that the gene product is a transcription factor. Here, we present data supporting this notion. We find that disco expression in the optic lobe primordium, a group of cells contacted by the developing optic nerve, depends on an autoregulatory feedback loop. We show that wild-type disco function is required for maintenance of disco mRNA and protein expression in the developing optic lobe. In addition, we demonstrate that ubiquitous Disco activity supplied by a heat-inducible gene construct activates expression from the endogenous disco gene specifically in the optic lobe primordium. Consistent with a role of Disco as a transcriptional regulatory protein, we show that portions of the Disco protein are capable of activating the transcription of reporter constructs in a heterologous system. Moreover, we find that the zinc finger portion of Disco binds in vitro to sequences located near the disco transcription unit, suggesting that Disco autoregulates its transcription in the optic lobe primordium by direct binding to a regulatory element in its own promoter. PMID- 10525343 TI - Calcium release at fertilization of Xenopus eggs requires type I IP(3) receptors, but not SH2 domain-mediated activation of PLCgamma or G(q)-mediated activation of PLCbeta. AB - Elevation of intracellular Ca2+ at fertilization is essential for the initiation of development in the Xenopus egg, but the pathway between sperm-egg interaction and Ca2+ release from the egg's endoplasmic reticulum is not well understood. Here we show that injection of an inhibitory antibody against the type I IP(3) receptor reduces Ca2+ release at fertilization, indicating that the Ca2+ release requires IP(3). We then examine how IP(3) production is initiated. Xenopus eggs were injected with specific inhibitors of the activation of two phospholipase C isoforms, PLCgamma and PLCbeta. The Src-homology 2 (SH2) domains of PLCgamma were used to inhibit SH2-mediated activation of PLCgamma, and an antibody against G(q) family G-proteins was used to inhibit G(q)-mediated activation of PLCbeta. Though the PLCgamma SH2 domains inhibited platelet-derived growth factor (PDGF)-induced Ca2+ release in eggs with exogenously expressed PDGF receptors, they did not inhibit the Ca2+ rise at fertilization. Similarly, the G(q) family antibody blocked serotonin-induced Ca2+ release in eggs with exogenously expressed serotonin 2C receptors, but not the Ca2+ rise at fertilization. A mixture of PLCgamma SH2 domains and the G(q) antibody also did not inhibit the Ca2+ rise at fertilization. These results indicate that Ca2+ release at fertilization of Xenopus eggs requires type I IP(3)-gated Ca2+ channels, but not SH2 domain mediated activation of PLCgamma or G(q)-mediated activation of PLCbeta. PMID- 10525344 TI - Synapse formation and agrin expression in stratospheroid cultures from embryonic chick retina. AB - Stratospheroids are three-dimensional cellular spheres which develop in vitro through the proliferation and differentiation of retinal neuroepithelial precursor cells. We investigated synapse formation in stratospheroids by analyzing the development of aggregates of synapse-associated molecules and of electron microscopically identifiable synaptic specializations. Our results show that the first aggregates of the GABA(A) receptor, the glycine receptor, and gephyrin appear in the inner plexiform layer after 8 days in culture simultaneously with the development of the first active zones and postsynaptic densities. In contrast, presynaptic molecules including synaptophysin could be detected in the inner plexiform layer before synaptogenesis, suggesting functions for these molecules in addition to neurotransmitter exocytosis at mature synapses. Similar to the retina in vivo, synapses were not found in the nuclear layers of stratospheroids. We also analyzed the isoform pattern, expression, and distribution of the extracellular matrix molecule agrin, a key regulator during formation, maintenance, and regeneration of the neuromuscular junction. In stratospheroids, several agrin isoforms were expressed as highly glycosylated proteins with an apparent molecular weight of approximately 400 kDa, similar to the molecular weight of agrin in the retina in vivo. The expression specifically of the neuronal isoforms of agrin was concurrent with the onset of synaptogenesis. Moreover, the neuronal agrin isoforms were exclusively found in the synapse-containing inner plexiform layer, whereas other agrin isoforms were associated also with the inner limiting membrane and with Muller glial cells. These results show that synapse formation is very similar in stratospheroids and in the retina in vivo, and they suggest an important role for agrin during CNS development. PMID- 10525345 TI - Cholesterol efflux-mediated signal transduction in mammalian sperm: cholesterol release signals an increase in protein tyrosine phosphorylation during mouse sperm capacitation. AB - We previously demonstrated that mouse sperm capacitation is accompanied by a time dependent increase in protein tyrosine phosphorylation that is dependent on the presence of BSA, Ca2+, and NaHCO(3), all three of which are also required for this maturational event. We also demonstrated that activation of protein kinase A (PK-A) is upstream of this capacitation-associated increase in protein tyrosine phosphorylation. BSA is hypothesized to modulate capacitation through the removal of cholesterol from the sperm plasma membrane. In this report, we demonstrate that incubation of mouse sperm medium containing BSA results in a release of cholesterol from the sperm plasma membrane to the medium; release of this sterol does not occur in medium devoid of BSA. We next determined whether cholesterol release leads to changes in protein tyrosine phosphorylation. Blocking the action of BSA by adding exogenous cholesterol-SO-(4) to the BSA-containing medium inhibits the increase in protein tyrosine phosphorylation as well as capacitation. This inhibitory effect is overcome by (1) the addition of increasing concentrations of BSA at a given concentration of cholesterol-SO-(4) and (2) the addition of dibutyryl cAMP plus IBMX. High-density lipoprotein (HDL), another cholesterol binding protein, also supports the capacitation-associated increase in protein tyrosine phosphorylation through a cAMP-dependent pathway, whereas proteins that do not interact with cholesterol have no effect. HDL also supports sperm capacitation, as assessed by fertilization in vitro. Finally, we previously demonstrated that HCO-(3) is necessary for the capacitation-associated increase in protein tyrosine phosphorylation and demonstrate here, by examining the effectiveness of HCO-(3) or BSA addition to sperm on protein tyrosine phosphorylation, that the HCO-(3) effect is downstream of the site of BSA action. Taken together, these data demonstrate that cholesterol release is associated with the activation of a transmembrane signal transduction pathway involving PK-A and protein tyrosine phosphorylation, leading to functional maturation of the sperm. PMID- 10525346 TI - Regulative development in a nematode embryo: a hierarchy of cell fate transformations. AB - Cell specification during embryogenesis of the model system Caenorhabditis elegans involves a combination of inductive and autonomous mechanisms. We have begun to study the development of other nematodes to investigate how well cell specification mechanisms are preserved among closely related species. Here we report that the embryo of the soil nematode Acrobeloides nanus expresses a so far undescribed regulative potential. When, for instance, the first somatic founder cell AB is eliminated it is replaced by its posterior neighbor EMS, which in turn is replaced by the C cell. This allows-different from C. elegans-the development of partial embryos up to hatching and sometimes to fertile adults. Thus, early somatic blastomeres in A. nanus are multipotent, each being capable of giving rise to more than one somatic founder cell. Lost germ-line cells, however, are not replaced. A model is presented, according to which in A. nanus cellular identities are assigned by specific reciprocal inhibitory cell-cell interactions absent in C. elegans. Differences and similarities in cell specification between the two species are discussed and related to different developmental strategies. PMID- 10525347 TI - Laminin and beta1 integrins are crucial for normal mammary gland development in the mouse. AB - We have examined the role of integrin-extracellular matrix interactions in the morphogenesis of ductal structures in vivo using the developing mouse mammary gland as a model. At puberty, ductal growth from terminal end buds results in an arborescent network that eventually fills the gland, whereupon the buds shrink in size and become mitotically inactive. End buds are surrounded by a basement membrane, which we show contains laminin-1 and collagen IV. To address the role of cell-matrix interactions in gland development, pellets containing function perturbing anti-beta1 integrin, anti-alpha6 integrin, and anti-laminin antibodies respectively were implanted into mammary glands at puberty. Blocking beta1 integrins dramatically reduced both the number of end buds per gland and the extent of the mammary ductal network, compared with controls. These effects were specific to the end buds since the rest of the gland architecture remained intact. Reduced development was still apparent after 6 days, but end buds subsequently reappeared, indicating that the inhibition of beta1 integrins was reversible. Similar results were obtained with anti-laminin antibodies. In contrast, no effect on morphogenesis in vivo was seen with anti-alpha6 integrin antibody, suggesting that alpha6 is not the important partner for beta1 in this system. The studies with beta1 integrin were confirmed in a culture model of ductal morphogenesis, where we show that hepatocyte growth factor (HGF)-induced tubulogenesis is dependent on functional beta1 integrins. Thus integrins and HGF cooperate to regulate ductal morphogenesis. We propose that both laminin and beta1 integrins are required to permit cellular traction through the stromal matrix and are therefore essential for maintaining end bud structure and function in normal pubertal mammary gland development. PMID- 10525348 TI - Identification of regulatory regions driving the expression of the Drosophila spalt complex at different developmental stages. AB - The zinc finger transcription factors Spalt and Spalt-related have been implicated in multiple developmental processes. In the wing they are regulated by the secreted protein Decapentaplegic and participate in the positioning of the wing veins. The function of Spalt has been also analyzed during tracheal development and embryonic segmentation. Here, we present the isolation and characterization of novel spalt/spalt-related alleles, which analysis indicates that these genes cannot substitute for each other in the developmental processes studied. The mutants present embryonic or pupal lethality, with phenotypes consistent with the loss of spalt function. We also present a detailed functional analysis of the DNA regions implicated in the regulation of these genes. This regulation is complex, integrating the information from both negative and positive regulators, and it is modular, with discrete fragments of DNA directing expression to discrete regions in embryonic and larval tissues. PMID- 10525349 TI - Evidence for a role of Smad6 in chick cardiac development. AB - Bone morphogenetic proteins (BMPs), members of the transforming growth factor beta (TGF-beta) superfamily, are obligatory growth factors for early embryogenesis and heart formation. SMAD proteins transduce signals of the TGF beta superfamily. We isolated chicken Smad6 (cSmad6), a member of inhibitory SMADs, and found its expression to be remarkably restricted to the developing heart, eyes, and limbs. cSmad6 expression was detected in the cardiogenic region of stage 5 embryos and overlapped Nkx2-5 and bmp-2, -4, and -7 expression. Throughout development, cSmad6 was expressed strongly in the heart, primarily in the myocardium, endocardium, and endocardial cushion tissue. Myocardial expression of cSmad6 was stronger in the forming septum, where highly localized expression of bmp-2 and -4 was also observed. Ectopically applied BMP-2 protein induced the expression of cSmad6, a putative negative regulator of BMP-signaling pathway, in anterior medial mesoendoderm of stage 4-5 embryos. In addition, blocking of BMP signaling using Noggin downregulated cSmad6 in cardiogenic tissue. cSmad1, one of the positive mediators of BMP signaling, was also expressed in cardiogenic region, but was not BMP-2 inducible. Our data suggest that cSmad6 has a role in orchestrating BMP-mediated cardiac development. We propose the possible mechanism of action of cSmad6 as modulating BMP signal by keeping a balance between constitutively expressed pathway-specific cSmad1 and ligand-induced inhibitory cSmad6 in the developing heart. PMID- 10525350 TI - Expression of HAND gene products may be sufficient for the differentiation of avian neural crest-derived cells into catecholaminergic neurons in culture. AB - Members of the basic helix-loop-helix family of DNA binding proteins have important roles in the development of subpopulations of neural crest-derived neurons. We have cloned the chicken homologues of dHAND (HAND2) and eHAND (HAND1), basic helix-loop-helix DNA binding proteins whose neuronal expression is restricted to sympathetic and enteric neural crest-derived ganglia. Transcripts encoding dHAND and eHAND are expressed in sympathetic ganglia beginning at Hamburger and Hamilton stage 17-18. Antisense blockade of transcripts encoding HAND genes in neural crest-derived cells in vitro results in a significant reduction in neurogenesis. Differentiation of catecholaminergic neurons is also reduced by 52% if the expression of transcripts encoding dHAND and eHAND is reduced using antisense oligonucleotide blockade. The effect on neurogenesis and phenotypic expression of neural crest-derived neurons is specific; blockade of HAND gene expression has no apparent influence on the differentiation in vitro of neural tube-derived neurons. Use of a replication-competent avian retrovirus to constitutively express HAND genes in neural crest-derived cells in vitro, under nonpermissive growth conditions in medium supplemented with 2% chick embryo extract (CEE), induced precocious catecholaminergic differentiation. Constitutive expression of HAND gene products resulted in a significant increase in catecholaminergic differentiation of cells grown in medium supplemented with 10% CEE, a permissive growth condition for catecholaminergic development. These results suggest that the expression by neural crest cells of dHAND and eHAND may be both sufficient and necessary for catecholaminergic phenotypic expression. PMID- 10525351 TI - Deficiency of Trp53 rescues the male fertility defects of Kit(W-v) mice but has no effect on the survival of melanocytes and mast cells. AB - Mutations of the receptor tyrosine kinase, Kit, or its ligand, mast growth factor (Mgf), affect three unrelated cell populations: melanocytes, germ cells, and mast cells. Kit signaling is required initially to prevent cell death in these lineages both in vitro and in vivo. Mgf appears to play a role in the survival of some hematopoietic cells in vitro by modulating the activity of p53. Signaling by Mgf inhibits p53-induced apoptosis of erythroleukemia cell lines and suppresses p53-dependent radiation-induced apoptosis of bone marrow cells. We tested the hypothesis that cell survival in Kit mutant mice would be enhanced by p53 deficiency in vivo. Double-mutant mice, which have greatly reduced Kit receptor tyrosine kinase activity and also lack Trp53, were generated and the affected cell lineages examined. Mast cell, melanoblast, and melanocyte survival in the double Kit(W-v/W-v):Trp53(-/-) mutants was not increased compared to the single Kit(W-v/W-v):Trp53(+/+) mutants. However, double-mutant males showed an increase in sperm viability and could father litters, in contrast to their homozygous Kit mutant, wild-type p53 littermates. This germ cell rescue appears to be male specific, as female ovaries were similar in mice homozygous for the Kit mutant allele with or without p53. We conclude that defective Kit signaling in vivo results in apoptosis by a p53-independent pathway in melanocyte and mast cell lineages but that in male germ cells apoptosis in the absence of Kit is p53 dependent. PMID- 10525352 TI - The Drosophila CPEB homolog, orb, is required for oskar protein expression in oocytes. AB - The establishment of polarity axes in the Drosophila egg and embryo depends upon the localization and on-site expression of maternal mRNAs. The critical step in the targeting of posterior determinants is the localization of oskar (osk) mRNA to the pole and its on-site translation. Osk protein then recruits other posterior group gene products involved in the formation of pole plasm and in the localization and regulation of the posterior determinant, nanos. Here we have investigated the role of the Drosophila CPEB homolog, the orb gene, in the osk mRNA localization pathway. We demonstrate that the expression of Osk protein is dependent upon the orb gene. In strong orb mutants, Osk protein expression is undetectable, while in the hypomorphic mutant, orb(mel), little or no on-site expression of Osk protein at the posterior pole is observed. The defects in Osk protein accumulation in orb mutant ovaries are correlated with a reduction in the length of the osk poly(A) tails. We show that osk mRNA is in immunoprecipitable complexes with Orb protein in ovaries and that the osk 3' UTR can be UV cross linked to Orb protein in ovarian extracts. These data suggest that Orb is required to activate the translation of osk mRNA and at that this may be accomplished by a mechanism similar to that used by the Xenopus CPEB protein to control translation of "masked" mRNAs. PMID- 10525353 TI - Functional analysis of placental 57-kDa Ca(2+)-binding protein: overexpression and downregulation in a trophoblastic cell line. AB - The placental trophoblastic epithelium functions to transport nutrients needed by the fetus, including calcium, which is required in the greatest amounts during the last third of pregnancy when the majority of fetal skeletal mineralization occurs. The mechanism of placental calcium transport and the developmental changes in the trophoblast that facilitate this process are currently incompletely understood. We have previously identified a 57-kDa, Ca(2+)-binding protein (CaBP) functionally implicated in placental calcium transport and trophoblast differentiation. In this study we have directly examined the role of CaBP in these processes by (1) recombinantly overexpressing CaBP in an inducible manner and (2) downregulating CaBP expression using antisense technology, using the rat choriocarcinoma cell line Rcho-1 as a trophoblastic cell model system. Our results show that overexpression of CaBP stimulates both cellular calcium uptake and vectorial calcium transport activities in Rcho-1 cells. Those cells stably expressing CaBP also exhibit higher levels of steady-state intracellular calcium and enhanced calcium-buffering ability. In addition, prolonged overexpression of CaBP in Rcho-1 cultures promotes trophoblast differentiation. Conversely, downregulation of CaBP expression had a negative effect on calcium uptake, calcium transport, and trophoblast differentiation in Rcho-1 cells. These data indicate that CaBP plays a direct role in placental calcium transport, functioning both as an intracellular calcium buffer and as a shuttle. These results also support a more direct role for CaBP in the trophoblast differentiation pathway. PMID- 10525355 TI - Different levels, but not different isoforms, of the Drosophila transcription factor DMEF2 affect distinct aspects of muscle differentiation. AB - mef2 genes encode alternatively spliced transcription factor isoforms that function in muscle differentiation in both Drosophila and vertebrates. Drosophila mef2 (Dmef2) has been shown to be required for the differentiation of a variety of distinct muscle types. However, many possible aspects of its function in muscle remain unexplored. There has also been no analysis in vivo of the activity of different MEF2 isoforms in any species. Our investigation centred on the role of different levels of DMEF2 in the Drosophila embryo in regulating diverse events of muscle differentiation and on the functional significance of Dmef2 alternative splicing. We used the GAL4/UAS system to both misexpress and overexpress individual DMEF2 isoforms and to rescue the different aspects of the Dmef2 mutant phenotype. Ectopic ectodermal expression of DMEF2 activated muscle gene expression and inhibited epidermal differentiation. Overexpression of DMEF2 in the mesoderm disrupted differentiation of the somatic and visceral muscle and the heart. The use of different DMEF2 levels in the rescue experiments revealed an activity range compatible with differentiation of the different muscle types: the consequence of too little or too much DMEF2 activity was disrupted differentiation. These rescue experiments also revealed that distinct DMEF2 thresholds are required for different properties within a cell and also for different cells within a muscle type and for different muscle types. Finally, each isoform functioned equivalently in these experiments, including in the stringent test of rescue of the Dmef2 mutant phenotype. PMID- 10525354 TI - Sonic hedgehog and BMP2 exert opposing actions on proliferation and differentiation of embryonic neural progenitor cells. AB - Although Sonic Hedgehog (Shh) plays a critical role in brain development, its actions on neural progenitor cell proliferation and differentiation have not been clearly defined. Transcripts for the putative Shh-receptor genes patched (Ptc) and smoothened (Smo) are expressed by embryonic, postnatal, and adult progenitor cells, suggesting that Shh can act directly on these cells. The recombinant human amino-terminal fragment of Shh protein (Shh-N) alone did not support the survival of cultured progenitor cells, but treatment with Shh-N in the presence of bFGF increased progenitor cell proliferation. Furthermore, treatment of embryonic rat progenitor cells propagated either in primary culture or after mitogen expansion significantly increased the proportions of both beta-tubulin- (neuronal marker) and O4- (oligodendroglial marker) immunoreactive cells and reduced the proportion of nestin- (uncommitted neural progenitor cell marker) immunoreactive cells. By contrast Shh-N had no effect on the elaboration of GFAP- (astroglial marker) immunoreactive cells. Cotreatment with Shh-N and bone morphogenetic protein-2 (BMP2) inhibited the anti-proliferative, astroglial-inductive, and oligodendroglial-suppressive effects of BMP2. Our observations suggest that Shh-N selectively promotes the elaboration of both neuronal and oligodendroglial lineage species and inhibits the effects of BMP2 on progenitor cell proliferation and astroglial differentiation. PMID- 10525356 TI - Purification and characterization of insulin from the Australian lungfish, Neoceratodus forsteri (Dipnoi). AB - The Australian lungfish Neoceratodus forsteri, a facultative air breather, is considered to be the most primitive of the extant Dipnoi and so occupies a uniquely important evolutionary position in the transition from fish to tetrapods. Insulin was isolated from an extract of the pancreas of N. forsteri and its primary structure established as: A-Chain, Gly-Ile-Val-Glu-Gln-Cys-Cys His-Thr-Pro(10)-Cys-Ser-Leu-Tyr-Gln-Leu-G lu-Asn-Tyr-Cys(20)-Asn-Glu-Thr-Glu; B Chain, Ala-Ala-Val-Asn-Gln-His-Leu-Cys-Gly-Ser(10)-His-Leu-Val-Glu-Ala-Leu- Tyr Phe-Val-Cys(20)-Gly-Glu-Arg-Gly-Phe-Phe-Tyr-Leu-Pro- Lys(30)-Gly. This amino acid sequence is more similar to that of human insulin than to insulins from present day amphibians. All the residues in human insulin that are considered to be important in receptor binding, dimerization, and hexamerization are conserved in lungfish insulin except for the substitution (Leu --> Phe) at the position corresponding to B17 in human insulin. Consistent with the assertion that the Dipnoi is a monophyletic group, insulins from N. forsteri and from the African lungfish Protopterus annectens contain extensions to the C-terminus of the A chain and to the N-terminus of the B-chain that have not been found in other sarcopterygian species. However, the unusual amino acid substitutions found in insulin from P. annectens (e.g., GlyB21 --> Ala, GluB22 --> Asp, and ArgB23 --> Asn) are not present in N. forsteri insulin, suggesting that they occurred in the Protopterus lineage after divergence of the genera. PMID- 10525358 TI - Tachykinins (substance P and neuropeptide gamma) from the brains of the pallid sturgeon, Scaphirhynchus albus and the paddlefish, Polyodon spathula (Acipenseriformes). AB - A peptide with substance P-like immunoreactivity was isolated from extracts of the brains of the pallid sturgeon, Scaphirhynchus albus and the North American paddlefish, Polyodon spathula. The primary structure of the peptide (Lys-Pro-Lys Pro-His-Gln-Phe-Phe-Gly-Leu-Met.NH(2)) is the same in both species and contains 2 amino acid substitutions (Arg(1) --> Lys and Gln(5) --> His) compared with human substance P and 1 substitution (Arg(3) --> Lys) compared with substance P from the trout (Teleostei). Scyliorhinin I, a tachykinin previously isolated from an extract of sturgeon intestine, was not detected in either brain extract. A peptide with neurokinin A-like immunoreactivity (Ser-Ser-Ala-Asn-Arg-Gln-Ile-Thr Gly-Lys(10)Arg-Gln-Lys-Ile-Asn-Ser-P he-Val-Gly-Leu(20)Met.NH(2)) was isolated from sturgeon brain and contains 10 amino acid substitutions compared with human neuropeptide gamma (a specific product of the posttranslational processing of gamma-preprotachykinin A) but only 4 substitutions compared with trout neuropeptide gamma. It was not possible to obtain the paddlefish neurokinin A related peptide in pure form. The structural similarity between the sturgeon and the trout tachykinins supports the hypothesis that the Acipenseriformes (sturgeons and paddlefish) represent the sister group of the Neopterygii (gars, bowfin, and teleosts). PMID- 10525357 TI - Studies on the origin of ovarian interstitial tissue and the incidence of endometrial hyperplasia in domestic and feral cats. AB - Ovarian interstitial cells (OICs) are a common feature of mammalian gonads but little is understood concerning their origin or functional significance. This study investigated the development and steroidogenic potential of OIC in feral and colony-reared feline queens. Reproductive tracts, collected from a total of 50 female colony and feral cats, were fixed and analyzed by morphometry. Ovarian sections were also immuno-stained for the expression of the steroidogenic enzymes 17alpha-hydroxylase/17,20 lyase cytochrome P450 (P450c17), 3beta-hydroxysteroid dehydrogenase/Delta5-Delta4 isomerase (3beta-HSD), and aromatase. These findings were related to serum estradiol and testosterone concentrations and to the degree of existing cystic endometrial hyperplasia (CEH). Feral cats had three times as many OICs as colony-reared queens (2713 +/- 855 vs 744 +/- 494 cells/mm(2), P < 0.01). These cells were lipid laden and expressed both P450c17 and 3beta-HSD at levels that were higher than those seen in the theca interna of adjacent follicles. Aromatase expression was undetectable. The pattern of enzyme expression was consistent with development of interstitial tissue from atretic follicles and the potential for continued steroid secretion during the anestrum. The incidence of CEH was higher in older (>5 years old; 88.2%) than in younger (2 4 years; 30%) colony queens (P < 0. 01), whereas no such disease was evident in any of the feral cats. Estradiol levels were higher in colony-reared than in feral cats, but testosterone levels were not different. These data are consistent with the transformation of the theca interna of atretic follicles in cats into OICs that retain a similar, or even enhanced, steroidogenic phenotype. Colony reared cats exhibit a predisposition to CEH compared with feral queens that is associated with elevated serum estradiol concentrations. Whether or not OICs somehow prevent the development of uterine disease or otherwise reflect a gonadal response to reduced negative feedback on the hypothalamic-pituitary axis remains to be determined. PMID- 10525359 TI - Seasonal changes in expression of neurohypophysial hormone genes in the preoptic nucleus of immature female masu salmon. AB - In relevance to osmoregulatory and reproductive functions, activity of the hypothalamic magnocellular neurosecretory system may vary seasonally in teleosts. The changes in the expression of vasotocin (VT) and isotocin (IT) genes were thus studied by an in situ hybridization technique and an immunohistochemical avidin biotin complex method in immature female masu salmon (Oncorhynchus masou). The plasma levels of testosterone and estradiol were also measured by enzyme immunoassay. Fish were sampled in March, May, August, and November 1994 and January 1995. The intensity of autoradiographic hybridization signals and immunoreactivity were determined in individual neurosecretory cells (NSC) in the rostroventral, middle, and dorsocaudal regions of the magnocellular part of the preoptic nucleus (PM). The VT hybridization signals and immunoreactivity were high in November, along with the elevation of plasma levels of testosterone and estradiol. These results suggest that sex steroid hormones are involved in seasonal regulation of VT gene expression. The hybridization signals for IT mRNA were increased in May and decreased in November, whereas IT immunoreactivity was low in March and high in November. NSCs thus showed seasonal variations in the intensity of hybridization signals for VT and IT mRNAs and immunoreactivity of VT and IT, although the patterns of changes were different between VT and IT. VT and IT genes may be seasonally expressed under different regulatory mechanisms. PMID- 10525360 TI - Differences in seasonal expression of neurohypophysial hormone genes in ordinary and precocious male masu salmon. AB - Our previous study showed the seasonal variations in expression of vasotocin (VT) and isotocin (IT) genes in preoptic magnocellular neurons of female masu salmon (Oncorhynchus masou). The changes in the level of VT mRNA were coincident with those in plasma testosterone and estradiol levels. In the present study, generality of this phenomenon in salmonid was verified in males. We examined changes in expression of VT and IT genes by an in situ hybridization technique and an immunohistochemical avidin-biotin complex method in the preoptic nuclei of ordinary and precocious male masu salmon. Plasma levels of testosterone and estradiol were measured by enzyme immunoassay. Fish were sampled in March, May, August, and November 1994 and January 1995. The intensities of hybridization signals for VT and IT mRNAs, as well as immunoreactivity of VT and IT, showed seasonal variations, although the profiles were different between the ordinary and precocious males. In the ordinary males, the intensities of hybridization signals for VT and IT mRNAs were high in January. These strong hybridization signals, representing elevation of VT and IT gene expression, were accompanied by increases in plasma levels of testosterone and estradiol. However, in precocious males, changes in VT and IT mRNA levels were not coincident with variation of plasma levels of sex steroid hormones. The sensitivity to sex steroid hormones of VT and IT gene expression may be different between the ordinary and precocious male masu salmon. PMID- 10525361 TI - Changes in patterns of corticosterone secretion concurrent with migratory fattening in a neotropical migratory bird. AB - Several studies on free-living birds have shown a change in corticosterone secretion (elevated baseline levels and a reduced corticosterone response to stress) during migration. It was not known, however, if this change was concurrent with the development of migratory condition or if it was an independent response to unknown environmental stressors experienced by the birds prior to capture. In this study, a Neotropical annual migrant, the yellow-rumped warbler (Dendroica coronata), held under controlled laboratory conditions, was used to test the Migration Modulation Hypothesis (MMH): during the migratory period migrants exhibit (1) elevated baseline corticosterone to facilitate migratory fattening and (2) a reduced corticosterone stress response, a means by which skeletal muscle needed for migration can be protected against catabolism by high levels of corticosterone. Fifteen hatching-year warblers were maintained on insect larvae and water ad libitum for 43 weeks, experiencing two transitions from a short- to long-day photoperiod to bring them into spring migratory condition. Corticosterone profiles comprising three blood samples from each individual (baseline at the time of initial disturbance and 30 and 60 min later), body mass, fat reserves, molt, and state of cloacal protuberance (males only) were measured at key intervals throughout the study. Over the entire study, mean baseline corticosterone levels were positively correlated with mean body mass, which increased predictably in response to long days. Individual baseline corticosterone was not correlated with individual body mass at any time. During periods when the birds were lean and held on short days, the corticosterone stress profiles were characterized by low initial hormone concentration followed by a significant increase in corticosterone with handling time. In response to long days, the warblers showed a significant increase in body mass and fat reserves concurrent with corticosterone stress profiles characterized by significantly elevated baseline levels and no further increase in corticosterone with handling time. These results support both components of the MMH illustrating changes in corticosterone secretion concurrent with migratory fattening but the exact nature of this change is unknown. PMID- 10525362 TI - The effect of estrogen on the gonads and on in vitro conversion of androstenedione to testosterone, 11-ketotestosterone, and estradiol-17beta in Sparus aurata (Teleostei, Sparidae). AB - The effects of estrogen on gonad morphology and steroidogenesis of sea bream, Sparus aurata, a protandrous hermaphrodite teleost, were investigated. Fish were treated in winter/spring for different periods with 17alpha-ethynylestradiol (ethE(2); experiment 1) and in summer with two doses of estradiol-17beta (E(2); experiment 2). Estrogen was more effective in summer. Its main effect on the gonad was inhibition of testicular growth and of male germ cell development beyond the spermatogonia stage, including mitosis. The effect of estrogen on ovarian development was slight and only apparent at the end of experiment 2 in the higher dose group. Gonadal fragments were incubated at different times during treatment with androstenedione and the output of testosterone (T), estradiol (E(2)), and 11-ketotestosterone (11KT) was measured by radioimmunoassay. T and E(2) production was inversely correlated with the proportion of testicular tissue (and positively with ovarian tissue) in the gonad in experiment 2. However, the production of 11KT was not correlated with any type of tissue, possibly because of further metabolism. Inhibition of testicular development by estrogen was also associated with higher output of steroid conjugates. PMID- 10525363 TI - Plasma gonadotropin concentrations in the cyclic female brushtail possum (Trichosurus vulpecula). AB - Changes in plasma concentrations of follicle-stimulating hormone (FSH) and luteinizing hormone (LH), and their relationship to antral follicle development and ovulation, were determined in female brushtail possums (Trichosurus vulpecula) in experiments in which pouch young were removed (RPY) from lactating females to promote ovarian activity. In Experiment 1 (n = 8), the development of preovulatory follicles and ovulation was monitored by laparoscopy. In Experiment 2 (n = 15) estrus and mating were monitored by cytology of urine. Ovulation occurred in 4/8 (Experiment 1) and 9/16 (Experiment 2) possums, and in these animals, plasma FSH concentrations fell progressively over the period of preovulatory follicle development and returned to pretreatment levels after ovulation. With the exception of samples taken at the time of the preovulatory gonadotropin surge, mean plasma LH levels remained basal. In those possums that failed to ovulate, plasma FSH concentrations were elevated while plasma LH concentrations were low; these patterns remained unchanged throughout the sampling period. It was not possible to distinguish between animals that would ovulate and those that would not ovulate after RPY on the basis of gonadotropin profiles at the time of RPY. A further group of possums (Experiment 3, n = 10) were blood-sampled at hourly intervals for 48 h to characterize preovulatory gonadotropin surges, using laparoscopy to monitor preovulatory follicular development and predict ovulation. A preovulatory LH surge (max. conc. 10.2-43.5 ng/ml, duration 7-9 h) was recorded in 4 animals, with a coincident preovulatory FSH surge (max. conc. 1.4-21.4 ng/ml, duration 3-11 h) observed in 3 of these possums. The patterns of gonadotropin secretion in the cycling brushtail possum conform to those reported for eutherians that ovulate spontaneously and appear to be regulated by similar mechanisms. PMID- 10525364 TI - Recombinant goldfish activin B stimulates gonadotropin-Ibeta but inhibits gonadotropin-IIbeta expression in the goldfish, Carassius auratus. AB - It is well documented that the pituitary in teleosts produces two gonadotropins, namely gonadotropin-I (GTH-I) and gonadotropin-II (GTH-II), which may regulate different phases of the reproductive cycle. However, unlike in mammals, very little is known about the differential regulation of the two GTHs in fish. Using goldfish as a model, the present study demonstrates, for the first time, that activin, a protein factor that plays a critical role in the differential regulation of mammalian FSH and LH, has opposite effects on GTH-Ibeta and GTH IIbeta mRNA expression. Recombinant goldfish activin B stimulates GTH-Ibeta but significantly suppresses GTH-IIbeta mRNA levels in a dose-dependent manner in cultured goldfish pituitary cells. Administration of recombinant human follistatin completely abolished the effects of activin, thus demonstrating the specificity of the activin activities. The novel opposite effects of activin on the two goldfish GTHs make goldfish a very unique vertebrate model for activin studies. The present study not only contributes to our understanding of the mechanisms that control the temporal expression patterns of the two GTHs during the fish reproductive cycle, but also provides important information on the evolution of gonadotropin regulation in vertebrates. PMID- 10525365 TI - Gonadotropin regulation of inhibin alpha-subunit mRNA and immunoreactive protein in cultured chicken granulosa cells. AB - Gonadotropin regulation of the inhibin alpha-subunit was investigated in chicken granulosa cell cultures. Granulosa layers were isolated from the F(1) and F(3) + F(4) follicles from three to four hens, pooled according to size, dispersed, and cultured (n = 3 replications for each experiment). In Experiments 1 and 2 either ovine LH or FSH was added to the cultures at doses of 0, 5, and 25 ng/ml. The cultures were terminated at 4, 24, and 48 h after plating. For both follicle sizes the expression of mRNA for the inhibin alpha-subunit was less (P < 0.05) at 24 and 48 h in untreated cells than in those treated with both doses of LH. Expression of the mRNA for the inhibin alpha-subunit was significantly increased only by the 25 ng/ml dose of FSH and only in the F(1) follicle at 24 and 48 h compared to the untreated cells. After 48 h of culture, immunoreactive alpha subunit protein accumulation was greater for both follicle types in the media of cells treated with the highest dose of LH and FSH than in the media from untreated cells. In Experiment 3, doses of 0, 5, 25, or 50 ng/ml of either LH or FSH were added to F(1) and F(3) + F(4) granulosa cells. All cultures were terminated at 48 h. LH and FSH increased the expression of the mRNA and immunoreactive protein for the inhibin alpha-subunit equally in a time-dependent manner. These experiments indicate that gonadotropins enhance the expression of both the mRNA and the protein for the inhibin alpha-subunit in chicken granulosa cells. PMID- 10525366 TI - Plasma glucose and insulin levels in genetically lean and fat sheep. AB - This study investigated whether genetically lean and fat sheep displayed differences in insulin and glucose statuses. Lean genotype sheep had significantly (P < 0.05) greater basal glucose concentrations than fat genotype sheep (4.78 versus 4.52, SED = 0. 104 mmol/l), although basal plasma insulin was not significantly different (mean 304, SEM = 37.3 pmol/l) between the genotypes. During glucose tolerance tests (GTT), carried out at 4 levels of injection: 0, 0.28, 1.39 or 2.78 mmol glucose/kg liveweight, the area under the plasma insulin curve was significantly (P < 0.05) greater for fat than lean genotype sheep, although there were no differences in any glucose parameters. There were no significant differences between genotypes in insulin or glucose concentrations during or following glucose infusion (GINF) experiments at 0, 0.09, 0.46 or 0.93 mmol glucose/kg live-weight/h over 3 hours. Elevated plasma insulin concentrations after a glucose tolerance test are concluded to be associated with increased fatness in this genetically selected line of sheep. However, the differences in insulin and glucose levels between the lean and fat genotype sheep are minor, relative to the differences in carcass composition. PMID- 10525367 TI - Ontogenetic profile of FSH and LH in Rana esculenta. AB - Circulating levels and pituitary content of FSH and LH were determined by specific radioimmunoassays in Rana esculenta starting a few days after hatching until the completion of metamorphosis. Both gonadotropins were found in the pituitary as well as in the blood plasma at all stages of development examined here. The plasma concentrations of FSH and LH were more or less uniform during pre- and prometamorphosis, but increased significantly at the onset of metamorphic climax. The plasma levels of FSH and LH remained high at the completion of metamorphosis. The pituitary content of FSH and LH was low in early premetamorphosis. It increased slightly through prometamorphosis and metamorphic climax, following which a highly significant increase occurred. Whereas plasma concentrations of FSH and LH were essentially similar within a single stage of development, the pituitary FSH content was severalfold higher than pituitary LH. The significance of these results is discussed in relation to the functional maturation of the brain-pituitary-gonadal axis in the frog. PMID- 10525368 TI - Modification of the plasma cortisol response to stress in rainbow trout by selective breeding. AB - Male and female rainbow trout were segregated into high- and low-responding individuals (HR, LR) on the basis of their plasma cortisol response to a 3-h period of confinement imposed at monthly intervals for 5 months. Consistent divergence was obtained in the responsiveness of the two groups, although the difference between LR and HR groups was greater in female fish (56 c.f. 116 ng ml(-1)) than in males (45 c.f. 69 ng ml(-1)). Progeny groups (full-sib families) were obtained from the pairing of HR males and females and LR males and females. A third progeny group (US) was obtained by random pairing of parents which were not selected as HR or LR. Poststress plasma cortisol levels in the progeny were first tested at 6 months after hatch and were significantly correlated with the response of the corresponding parental groups, HR > US > LR (178, 126, 81 ng ml( 1), respectively). The difference in responsiveness between LR and HR groups was demonstrated in all four subsequent tests over a 12-month period. There were no significant differences in baseline plasma cortisol levels in LR and HR groups prior to confinement. During a 4-h period of confinement, the differences in plasma cortisol levels between LR and HR fish were sustained throughout, indicating that the trait upon which the fish were selected was related to absolute maximum levels of circulating cortisol and not to the rate of change of cortisol levels during exposure to a stressor. A moderately high heritability (h(2)) for confinement-induced plasma cortisol of 0.41 was obtained by a parent progeny regression. Manipulation of stress responsiveness in fish by selective breeding offers scope for optimizing performance under intensive rearing conditions but also provides a useful research tool for investigating the operation of the endocrine stress response. PMID- 10525369 TI - Colocalization of GnRH binding sites with gonadotropin-, somatotropin-, somatolactin-, and prolactin-expressing pituitary cells of the pejerrey, Odontesthes bonariensis, in vitro. AB - Previous studies in the pejerrey, Odontesthes bonariensis, have demonstrated that fibers with immunoreactivity to gonadotropin-releasing hormone (ir-GnRH) reach all areas of the pituitary gland, the rostral pars distalis (RPD), the proximal pars distalis (PPD), and the pars intemedia (PI). A close association was shown between ir-GnRH fibers and gonadotropin (GtH)-, growth hormone (GH)-, somatolactin (SL)-, and prolactin (PRL)-expressing cells. The presence of only one GnRH variant, suspected to be a novel form, has been shown in pituitary extracts of this fish. In addition, GnRH may stimulate GtHs, GH, SL, and PRL levels in different fish species. The objective of the present study was to seek GnRH receptors and therefore colocalization with GtHs, GH, SL, and PRL cells in O. bonariensis using a pituitary primary cell culture system. GnRH binding sites were revealed by autoradiography of an iodinated superactive GnRH agonist ([(125)I]GnRH-A) and pituitary cells were identified by immunocytochemistry using piscine antisera. Following autoradiography, silver grains representing specific [(125)I]GnRH-A binding were associated with anti GtH, GH, SL, and PRL positive cells. These results demonstrate the presence of GnRH binding sites on these cells. It is suggested that GnRH may play a wide role in the neuroendocrine control of different pituitary hormones in addition to the GtHs. PMID- 10525370 TI - A multicenter phase II study with triptorelin (sustained-release LHRH agonist) in advanced or recurrent endometrial carcinoma: a French anticancer federation study. AB - The objective of this phase II multicenter study was to assess the efficacy and tolerance of triptorelin (a sustained-release LHRH agonist) in advanced or recurrent endometrial cancer. A total of 101 monthly intramuscular injections were administered to 24 eligible patients (median number/patient = 3; range 1 12). Mainly due to progression, only 16 patients received 3 or more injections. Among the 23 evaluable patients, 1 complete and 1 partial response (response rate of 8.7%) and 5 disease stabilizations were observed, often of long duration, but never in an irradiated area or after progestogens treatment failure. Median survival for eligible patients was 7.2 months (range: 1-36 months). Only grade 1 toxicities possibly related to the treatment were observed in 4 patients. In conclusion, triptorelin was safe, well tolerated, and easily manageable, and the very low toxicity did not impair the quality of life in these patients with a very poor prognosis. Although the response rate was disappointing, several patients showed early evidence of efficacy which may be of long duration. Response rates range between 0 and 45% in different published studies. Additional studies with stricter inclusion criteria and a larger sample size are necessary to better evaluate the role of LHRH agonists in endometrial adenocarcinomas. PMID- 10525371 TI - The effect of amifostine on the in vitro cytotoxicity of chemotherapeutic agents in three epithelial ovarian carcinoma cell lines. AB - OBJECTIVES: Amifostine protects against a spectrum of toxicities induced by chemotherapy without affecting tumor cell kill. This is supported by clinical data and in vivo animal studies. However, there is a paucity of data on its effect on the tumor cytotoxicity of several chemotherapeutic agents used in recurrent epithelial ovarian cancer. This study compares in vitro cytotoxicity before and after addition of amifostine. METHODS: Three epithelial ovarian carcinoma cell lines (SKOV3, 420, 429) were exposed to cis-platinum, paclitaxel, doxorubicin, etoposide, 5-fluorouracil, bleomycin, 4-epidoxorubicin, 4-HC (activated cyclophosphamide), vincristine, actinomycin D, mitomycin C, and topotecan. Cells were pretreated with either 0 or 1.2 mM amifostine. Tumor cell kill after 6 days of incubation was measured using the ATP cell viability assay. Paired samples Student's t statistic was used to test the difference in mean ATP levels between drug-treated samples with and without pretreatment with amifostine. RESULTS: SKOV3 was sensitive to paclitaxel, actinomycin D, 4 epidoxorubicin, and vincristine. Cell line 420 was sensitive to paclitaxel, etoposide, and 5-fluorouracil. Cell line 429 was sensitive to paclitaxel and 5 fluorouracil. There was no significant difference in the mean ATP levels between drug-treated samples with and without pretreatment with amifostine for each of the sensitive drugs in all three cell lines. Similarly, there was no significant difference in the mean ATP levels in cis-platinum and 4-HC treated samples with and without pretreatment with amifostine. CONCLUSIONS: These results show that at the cellular level amifostine did not protect epithelial ovarian carcinoma cells against tumor cell kill. PMID- 10525372 TI - IL-2 enhances standard IFNgamma/LPS activation of macrophage cytotoxicity to human ovarian carcinoma in vitro: a potential for adoptive cellular immunotherapy. AB - OBJECTIVE: The objective was to evaluate the enhancement of human peritoneal macrophage cytotoxic in vitro activity by the addition of interleukin-2 (IL-2) to the standard interferon gama (IFNgamma) and lipopolysaccharide (LPS) activation procedure used for cellular adoptive immunotherapy in a human ovarian cancer system. This cytotoxic effect of these activated macrophages was tested on cells from ovarian cancers of various stages, histology type, and grade, both prior to chemotherapy and at recurrence, in ovarian carcinoma cells lines and normal cells. Increased activation of the macrophage may make it a better candidate for intraperitoneal cellular adoptive immunotherapy as a component of ovarian cancer therapy. This was not a study of the mechanism of macrophage killing. METHODS: Ascites specimens were collected from 24 ovarian cancer patients at the time of surgery or by paracentesis. The mononuclear cell fraction was isolated by discontinuous density gradient centrifugation and used as a cellular source of peritoneal macrophages (PMs) and primary cultured ovarian cancer cells. PMs were separated by 1-h adhesion followed by intensive washing to remove floating cells. The floating cells were cultured for 24 h which left the cancer cells attached after unattached cells were removed by washing. These cells formed a monolayer of cancer cells, which could be subcultured in 22 patients. The cells from the third to fifth passages were used as target cells without coculture with other cells. PMs were identified by latex ingestion, and their purity after isolation by adhesion culture was tested by flow cytometry and immunofluorescence. PMs were activated by culturing in the presence of IFNgamma, with or without IL-2, for 18 h followed by the addition of LPS 6 h prior to use as effector cells in cytotoxicity assays. Ovarian cancer cells of both established cell lines and primary cultures were labeled with (51)Cr and utilized as target cells to quantitatively measure PM-mediated cytotoxicity. Ovarian cancer cells were also cocultured with PMs for morphologic observations to provide supporting evidence to the cytotoxicity assays. RESULTS: IL-2 enhances the cytotoxicity of the standard IFNgamma/LPS macrophage activation in this system. Peritoneal macrophages so activated are cytotoxic to autologous and allogenic primary cultured ovarian tumors and to ovarian carcinoma cell lines. The macrophages are cytotoxic to cells both prior to treatment and at recurrence, but the data from the few recurrent patients did reach statistical significance. This cytotoxicity is not MHC associated. Normal cells are minimally affected. CONCLUSIONS: IL-2 augmented the standard IFNgamma/LPS method of activating peritoneal macrophage cell killing of human ovarian cancer cells in this in vitro system. The cell killing occurred with autologous and allogenic tumor cells from patients with primary and possibly recurrent tumors. Activated PMs minimally affected the normal cells tested. This enhanced activation may improve the disappointing results of previous adoptive cellular immunotherapy human trials and should be considered for ovarian cancer clinical trials. PMID- 10525373 TI - FIGO stage IIIC endometrial carcinoma with metastases confined to pelvic lymph nodes: analysis of treatment outcomes, prognostic variables, and failure patterns following adjuvant radiation therapy. AB - OBJECTIVES: This study was undertaken to evaluate the prognostic significance of isolated positive pelvic lymph nodes on survival and to analyze other prognostic variables, overall survival, and failure patterns in surgically staged endometrial carcinoma patients with positive pelvic lymph nodes and negative para aortic lymph nodes following radiation therapy (RT). METHODS: Between January 1, 1987, and December 31, 1997, 782 women underwent primary treatment for uterine cancer at Indiana University Medical Center. Through a review of the medical records, we identified 58 patients with pathologic stage IIIA, 27 patients with pathologic stage IIIB, and 77 patients with pathologic stage IIIC endometrial carcinoma. Patients with pathologically positive or unsampled para-aortic lymph nodes and patients who received preoperative radiation therapy were excluded, leaving a study group of 17 patients with nodal metastases confined to pelvic lymph nodes. Thirteen patients received adjuvant pelvic RT using AP-PA or four field technique. A median dose of 5040 cGy was delivered. Four patients received whole abdominal irradiation (WAI) delivering a median dose of 3000 cGy. Two patients received vaginal cuff boosts of 1000 and 3560 cGy to 0.5 cm from the vaginal surface mucosa via Cs-137 brachytherapy. Two patients also received adjuvant chemotherapy (cis-platinum and doxorubicin) and/or hormonal therapy (megestrol acetate). Disease-free and overall survivals were estimated using the Kaplan-Meier method of statistical analysis and prognostic variables were analyzed using the log-rank test. RESULTS: With a median follow-up of 51 months the actuarial 5-year disease-free survival was 81% and the actuarial 2-year and 5 year overall survival rates were 81 and 72%, respectively. Univariate analysis revealed that positive peritoneal cytology in conjunction with positive pelvic lymph nodes imparts a greater risk of recurrence and decreased overall survival. There were no pelvic and/or upper abdominal failures, but there were recurrences in the para-aortic lymph nodes (two patients) and distantly (two patients). CONCLUSION: Surgery followed by postoperative pelvic RT is a viable treatment option for pathologically staged stage IIIC endometrial carcinoma with disease confined to the pelvic lymph nodes. Failures in the para-aortic region suggest a possible role for extended-field RT. Patients with positive peritoneal cytology in conjunction with nodal metastasis fared poorly with pelvic RT. Studies evaluating the efficacy of WAI are ongoing. Finally, substages within FIGO stage IIIC are recommended in an effort to better understand and define treatment strategies which might be appropriate for these patients. PMID- 10525374 TI - Effect of all-trans-retinoic acid on integrin receptors of human cervical cancer (SiHa) cells. AB - Cell surface receptors have been the subject of intensive investigations over the past few decades. One very important group of receptors on the cell surface is the "integrin" receptors which bind to extracellular matrix (ECM) proteins. Because of integrin's importance in cellular growth, development, and morphology the role of integrin receptors in cellular transformation, malignant growth, and metastasis has received wide attention. In this article we report on the effect of all-trans-retinoic acid (ATRA) on (a) the integrin family of cell surface receptors, (b) collagenase enzyme activity, and (c) invasive potential in human cervical cancer (SiHa) cells. A comparative cell adhesion assay clearly showed that ATRA affects the cell surface integrin receptors against different ECM proteins in a dose- and time-dependent manner. The binding of SiHa cells to ECM proteins (fibronectin, vitronectin, laminin, collagen IV) was drastically reduced when cells were treated with ATRA at 10 microM for 96 h in culture. Interestingly, when ATRA-treated (10 microM, 96 h) SiHa cells were allowed to grow for 15 days in ATRA-free complete medium the binding of SiHa cells to fibronectin and vitronectin was inhibited, even after 15 days of drug withdrawal, whereas cell adhesion to laminin and collagen IV returned to normal within 3-7 days. The comparative immunoprecipitation of two cell surface integrin receptors (alpha5beta1 and alphavbeta3) shows the effect of ATRA on the expression of alpha5, alphav, and beta1 subunits. In ATRA-treated SiHa cells the cell surface expression of the alphav subunit (in alphavbeta3 receptor) is much less than in untreated SiHa cells. In the case of the alpha5beta1 integrin receptor ATRA treatment caused a significant reduction in the expression of both alpha5 and beta1 subunits on the cell surface. Comparative zymography clearly demonstrated the inhibitory effect of ATRA on collagenase enzyme activity. Interestingly, the effect was irreversible, even after 15 days of culture in ATRA-free medium. The assay of the invasive potential of ATRA-treated and untreated SiHa cells in Boyden's invasion chamber demonstrated that ATRA treatment (10 microM, 96 h) inhibits the invasive potential of SiHa cells. The effect was not reversible even after 15 days of culture in ATRA-free medium. In conclusion, our observations indicate that ATRA has an inhibitory effect on the expression of SiHa cell surface integrin receptors and collagenase enzyme activity. The effect of ATRA on cell surface integrin receptors and collagenase enzyme activity may affect the invasive potential of SiHa cells. PMID- 10525375 TI - Surgical experiences and training of residents: perspective of experienced gynecologic oncologists. AB - OBJECTIVE: The aim of this study was to report the opinions of experienced gynecologic oncologists concerning the surgical education and experiences of residents. METHODS: The 1997 membership directory of the Society of Gynecologic Oncologists was used to identify individuals who were members for at least 5 years and on the faculty of residency training programs. One hundred seventy members were identified and a nine-question survey was mailed to them. RESULTS: One hundred nineteen (70%) surveys were returned. One hundred seventeen individuals were on the faculty of residency training programs and involved in the surgical training of residents. Ninety-six percent reported that gynecologic oncologists were a major resource for surgical education at their institution. Eight-nine (76%) reported a change in the volume of major abdominal and vaginal surgical procedures performed for noninvasive disease over the past 5 years. Of these 89, 16 (18%) reported that surgical volume at their institution decreased by 10%, 38 (43%) reported that surgical volume decreased by 10-25%, and 17 (19%) reported that surgical volume decreased by more than 25%. Sixty-three percent of all respondents reported that residents were not as well versed in pre- and postoperative care when compared to those of 5 years ago. Sixty-five percent of all respondents reported that graduating residents were less prepared in surgical techniques when compared to those of 5 years ago. Seventy-five percent of respondents reported that the primary care requirements of the RRC have decreased the amount of surgical experience, ICU rotations, and anesthesia rotations. Sixty percent of all respondents operated with other attending surgeons on more difficult cases, and 29% had changed to this practice within the past 5 years. Eighty-two percent believed that more time during residency training needs to be devoted gynecologic surgical experience. CONCLUSIONS: Experienced gynecologic oncologists on the faculty of residency training programs report a decrease in surgical skills and surgical experiences when compared to residents trained 5 years ago. PMID- 10525376 TI - Repetitive hydatidiform mole with different male partners. AB - OBJECTIVE: The aim of this study was to determine the role of parental factors that may relate to the pathogenesis of molar pregnancy. METHODS: A retrospective review of six patients who had a molar pregnancy with at least two different partners at New England Trophoblastic Disease Center between 1965 and March 1999 was performed. RESULTS: A total of 34 pregnancies with 20 different partners were observed in 6 patients. These pregnancies resulted in 15 (44.1%) hydatidiform moles, 8 (23.5%) term live births, 7 (20.6%) therapeutic abortions, 3 (8.8%) spontaneous abortions, and 1 preterm delivery. While 5 patients had a molar pregnancy with 2 different partners, 1 patient had a molar pregnancy with 3 different partners. Two patients developed persistent postmolar gestational trophoblastic tumor in 3 (20.0%) of the 15 episodes of molar pregnancy. Three of the male partners reported a total of 7 healthy children from prior relationships. CONCLUSION: The experience in these six patients suggests that a primary oocyte problem may contribute to the development of molar pregnancy. PMID- 10525377 TI - Benign and malignant serous and endometrioid epithelium in the omentum. AB - OBJECTIVES: Benign and malignant serous and endometrioid epithelial proliferations are found in the omentum, where their presence may be interpreted either as metastases from Mullerian tumors elsewhere or as primary peritoneal tumors. The present study was undertaken in an attempt to gather data that might help resolve the issue. METHODS: The ratios of serous epithelium to endometrioid epithelium in the omentum, in both the benign and malignant states, were determined for cases from January 1985 to July 1997 and January 1991 to December 1997, respectively. RESULTS: In ovarian carcinoma, the ratio of malignant serous epithelium to endometrioid epithelium involving the omentum is 15:1. This is comparable to the ratio of benign serous epithelium to endometrioid epithelium in the omentum, which is 10:1. The ratio of primary peritoneal serous carcinoma to endometrioid carcinoma is 10.5:1. CONCLUSION: It seems not reasonable that endometrioid carcinoma of the ovary is 15 times less likely to metastasize to the omentum than its serous counterpart. The ratio, however, is not unreasonable if endometrioid and serous carcinomas arise from preexisting endometrioid or serous epithelium. We conclude that serous and endometrioid carcinomas may arise primarily in the omentum and, in at least some cases, may derive from their benign counterparts. PMID- 10525378 TI - A limited role for adjuvant radiotherapy after the Wertheim/Okabayashi radical hysterectomy for cervical cancer confined to the cervix. AB - BACKGROUND: The indications for radiotherapy after radical hysterectomy for early stage cervical cancer are changing. In the past only tumor outside the cervix was considered an indication for radiotherapy. Today adjuvant radiotherapy is also considered for an "intermediate-risk" group with tumor confined to the cervix but poor prognostic primary tumor parameters such as large tumor diameter, vascular space invasion, and deep stromal penetration. OBJECTIVE: The aims of this study were to determine the risk of isolated pelvic recurrences in an intermediate-risk group (GOG Study No. 92) and to analyze whether this group will theoretically benefit from adjuvant pelvic radiotherapy. PATIENTS AND METHODS: A retrospective analysis was performed on 271 patients with early cervical cancer treated by a radical hysterectomy in a uniform fashion in one institute. Radiotherapy was administered only when tumor was found outside the cervix. Tumor diameter, capillary lymphatic space invasion, and depth of stromal penetration were assessed in all patients. Recurrence pattern, disease-specific survival, and recurrence-free interval were determined in the intermediate-risk group and compared with the remaining patients of the group with tumor confined to the cervix. RESULTS: A significant difference in disease-specific survival (89% versus 97%, P < 0.03) and 5-year recurrence-free interval (86% versus 95%, P < 0.02) was noted in the intermediate-risk group (n = 56) compared with the total group with tumor confined to the cervix. Three patients in the intermediate-risk group died of disease with a pelvic recurrence. Two of these patients had a combined pelvic and distant recurrence. CONCLUSION: Our retrospective results fail to support a survival benefit of extending indications for adjuvant radiotherapy other than postive nodes, parametrial extension, and positive margins. PMID- 10525379 TI - Pretreatment scalene node biopsy in gynecologic malignancy: prudent or passe? AB - OBJECTIVES: Surgicopathologic evaluation of the scalene fat pad is considered a critical step in the pretreatment evaluation of patients at our institution with cervical or corpus carcinoma when the periaortic lymph nodes (PAN) are involved. However, enthusiasm for this procedure at other centers has waned, largely due to a wide discrepancy in the reported rates of occult scalene node involvement. In an attempt to clarify the benefit of pretreatment scalene node sampling in gynecologic malignancies, we present our experience over the past 18 years. MATERIALS AND METHODS: We identified 57 patients who underwent scalene node sampling between 1980 and 1998. In 39 of 49 (80%), the decision to proceed with scalene node sampling was based entirely on histologically documented PAN metastases. In the remainder, scalene node sampling was prompted by the presence of suspicious clinical findings. RESULTS: Of the 49 patients included in the study, 33 had carcinoma of the cervix, while 16 had corpus carcinoma. Ninety percent of scalene node sampling was performed at the time of primary diagnosis. Overall, 9 patients (18%) had scalene node metastases. Notably, not a single patient with corpus cancer was found to have scalene node metastases in the absence of clinically evident scalene node enlargement independent of PAN status. In cervix cancer cases, the presence of grossly involved PAN was predictive of a high likelihood of scalene node metastases (44%), while no patient with occult PAN metastases had involvement of the scalene node. Only 1 minor complication was encountered following scalene node sampling. The 40 scalene node-negative patients were treated with either extended field radiation or whole abdominal radiation therapy, and 20% developed a major, RTOG grade >/=3 complication such as fistula formation, bowel obstruction, or ureteral stenosis. Only 1 case of mild radiation enteritis and cellulitis occurred during palliative radiation in the group of patients with scalene node metastases. CONCLUSIONS: Scalene node sampling may be of benefit in the pretreatment evaluation of patients with cervical carcinoma when PAN are grossly involved. Given that scalene node involvement satisfies the criteria for distant metastases, identification of such allows the clinician to avoid the morbidity of extended field radiotherapy in a setting without the chance for cure. PMID- 10525380 TI - Flow cytometric DNA analysis of early stage adenocarcinoma of the cervix. AB - OBJECTIVE: The aim of this study was to determine the utility of DNA flow cytometry as a prognostic indicator for risk of recurrence and overall survival in patients with early stage adenocarcinomas of the uterine cervix. METHODS: DNA flow cytometry was performed to determine ploidy, DNA index, and proliferative index in 66 women with stage IB and IIA pure mucinous adenocarcinomas of the cervix treated by primary surgical therapy with radical hysterectomy and pelvic lymphadenectomy. Fifty-seven of 66 (86.3%) tissue samples were analyzable. Three sections were obtained from paraffin-embedded tissue blocks containing primary tumor. Flow cytometric results, along with other known prognostic variables for risk for recurrent disease and survival, were analyzed using the Cox regression proportional hazards model and survival curves generated by the Kaplan-Meier method. RESULTS: Of 57 interpretable samples, DNA ploidy patterns were 18 (27%) diploid, 8 (12%) tetraploid, and 31 (47%) aneuploid. Thirteen of 66 patients (20%) experienced recurrence with a median time to recurrence of 1.6 years. No significant correlation was noted between DNA ploidy and risk of recurrence (P = 0.429). Multivariate analysis confirmed that positive metastatic lymph nodes were associated with risk of recurrence (P < 0.001). In node-negative patients, a high proliferative index (S% + G(2)M% > 20%), measured as a continuous variable, was the only significant factor for tumor recurrence (P = 0.002). CONCLUSION: DNA ploidy does not predict a patient's risk for tumor recurrence; however, a high proliferative index value warrants further investigation as a potential prognostic indicator for risk of recurrent disease in patients with adenocarcinoma of the uterine cervix. PMID- 10525382 TI - alphavbeta3 and vitronectin expression by normal ovarian surface epithelial cells: role in cell adhesion and cell proliferation. AB - The alphavbeta3 integrin and its ligand vitronectin are expressed by differentiated epithelial ovarian carcinomas and carcinoma cell lines in culture. Moreover, alphavbeta3/vitronectin interaction influences adhesion and migration of ovarian carcinoma cells in culture. For a better understanding of the behavior of these carcinomas, it appeared necessary to study the characteristics of their normal counterpart, the ovarian surface epithelium (OSE). The present study showed that normal cultured human OSE cells, like the carcinoma cells, have the ability to synthesize vitronectin. The vitronectin receptor, alphavbeta3 integrin, is also expressed by OSE cells and is localized in focal contacts close to paxillin, a focal contact-specific protein, and p125(FAK), a cytoskeletal and signaling molecule. This localization suggested an active participation of the integrin in the adhesion and/or proliferation of OSE cells. Indeed, the use of a blocking antibody demonstrated that alphav integrins promote OSE cell adhesion on vitronectin but not on fibronectin and that these integrins are required for maximal proliferative activity. The results suggest a role of the alphavbeta3/vitronectin system in normal OSE physiology and demonstrate that the expression of this system by well-differentiated ovarian carcinomas reflects the retention of normal cell properties. PMID- 10525381 TI - Matrix metalloproteinases and their inhibitors in gestational trophoblastic diseases and normal placenta. AB - OBJECTIVE: Our purpose was to investigate the expression of matrix metalloproteinases (MMPs) in gestational trophoblastic diseases and normal first trimester placenta. METHODS: Paraffin sections of 16 partial moles, 25 complete moles, 10 gestational choriocarcinomas, and 11 normal first-trimester placentas were studied immunohistochemically for expression of MMP-1, MMP-2, MMP-3, MMP-9, MMP-13, and tissue inhibitor of metalloproteinase-1 (TIMP-1). RESULTS: Nine (90.0%) of the choriocarcinoma cases showed strong intensity of staining for MMP 1. Choriocarcinoma exhibited significantly stronger staining for MMP-1 than syncytiotrophoblast in normal placenta (P < 0.01), partial mole (P < 0.01), and complete mole (P < 0.01). Choriocarcinoma also showed significantly stronger staining for MMP-1 than the extravillous trophoblast in placenta (P < 0.05). MMP 2 was expressed only in syncytio- and extravillous trophoblasts in normal placenta, partial mole, and complete mole. Choriocarcinoma and the extravillous trophoblast in partial mole and complete mole had significantly stronger staining for MMP-2 than the extravillous trophoblast in placenta (P < 0.05, P < 0.01, P < 0.01, respectively). Choriocarcinoma also exhibited significantly stronger staining for MMP-2 than syncytiotrophoblasts in placenta (P < 0.01), partial mole (P = 0.05), and complete mole (P < 0.01). The expression of MMP-3, MMP-9, and MMP 13 was similar in all four tissues with the predominance of syncytiotrophoblast for MMP-3 and MMP-13 and cytotrophoblast for MMP-9. While 8 (73.0%) placentas, 14 (87.5%) partial moles, and 19 (76.0%) complete moles showed strong immunoreactivity for TIMP-1 in syncytiotrophoblasts, no strong staining was found in choriocarcinomas (P < 0.01, P < 0.01, P < 0.01, respectively). CONCLUSION: The extravillous trophoblast of first-trimester placenta has significantly less expression of MMP-1 than choriocarcinoma and significantly less expression of MMP 2 than choriocarcinoma and extravillous trophoblast of partial and complete mole. The expression of TIMP-1 was significantly less in choriocarcinoma than the syncytiotrophoblast of normal placenta, partial mole, and complete mole. MMPs and their inhibitors may play a role in the pathogenesis of gestational trophoblastic diseases. PMID- 10525383 TI - Adenovirus-mediated p53 growth inhibition of ovarian cancer cells is independent of endogenous p53 status. AB - OBJECTIVE: The aim of this study was to determine the effect of transfection of adenovirus-mediated wild-type p53 into ovarian cancer cells with both wild-type and mutant endogenous p53. STUDY DESIGN: Eight human ovarian cancer cell lines were used: three with p53 mutations, one that is p53 null, and four with wild type p53. The recombinant p53 adenovirus (Adp53) contains the cytomegalovirus promoter, wild-type p53 cDNA, and SV40 polyadenylation signal in a minigene cassette inserted into the E1-deleted region of modified Ad5. The transduction efficiency of cells was assessed using a beta-gal-containing adenovirus. Cell counting assays were used to evaluate the effect of transfection with Adp53 on the growth of cells. P53 expression was evaluated using Western blot. Cell cycle analysis and apoptosis studies were done using a tunnel-based assay and fluorescent activated cell sorting. RESULTS: Transduction efficiencies varied between cell lines. More than 90% growth inhibition occurred in seven of eight cell lines after infection with adenovirus-mediated p53 if a viral dose leading to at least 50% of cells infected was used. Regardless of endogenous p53 status, apoptosis occurred in cells infected with p53. CONCLUSIONS: Ovarian cancer cells are growth inhibited by transfection with adenovirus-mediated p53 regardless of their endogenous p53 status. Growth inhibition is related to transduction efficiency. PMID- 10525384 TI - Pathology slide review in gynecologic oncology: routine or selective? AB - OBJECTIVE: The aims of this study were to assess the cost/benefit ratio for interinstitution pathology consultation (IPC) and to identify the types of specimens with little or no risk of diagnostic error in order to reduce the cost. METHODS: All gynecologic oncology referrals having IPC from 1993 to 1998 were reviewed. Each case was evaluated by comparing both the original and the consulted pathology reports. A discrepancy was major if it led to treatment alteration. A minor discrepancy was defined as differences without clinical consequences. Consultation error was determined by comparison with the final diagnosis and clinical data obtained from the records. The cost per review was adjusted to 1998 dollars for all cases over the 5-year study period. Statistical data were obtained by Fisher's exact test and Pearson's correlation test. RESULTS: Five hundred sixty-nine pathology specimens from 498 patients were analyzed in this study. The major discrepancy rate was 6.5% and the minor discrepancy rate was 12.5%. Cytological specimens accounted for no major discrepancy and 13 minor discrepancies compared to 37 major and 58 minor discrepancies in histological specimens. The difference was statistically significant (P = 0.003). Consultation errors occurred in 5 cases with no alteration of clinical care. By excluding cervical and vaginal smears and cervical biopsy specimens in cases with clinically gross tumors, the cost can be reduced by 25% with no detriment to the clinical management. CONCLUSIONS: The types of specimens that do not need consultative pathology review include (1) cervical biopsy specimens in those patients with gross tumors and (2) cervical and vaginal smears. PMID- 10525386 TI - Evaluation of CO(2) laser excision or vaporization for the treatment of vulvar intraepithelial neoplasia. AB - OBJECTIVE: Our objective was to evaluate the results of laser surgery in patients with vulvar intraepithelial neoplasia (VIN). METHODS: From January 1990 to December 1996, 52 patients with histologically proven VIN were treated with CO(2) laser vaporization or laser excision. The analysis included anamnestic characteristics, clinical aspects, types of treatment, correlation of the preoperative biopsy with the excised pathologic specimen, and follow-up results. RESULTS: Fourteen women underwent laser vaporization, and 38, laser excision. Of the patients submitted to vaporization, 11 were cured in one session (75%), 1 required two procedures, and 2 other patients, who underwent more treatments, eventually developed invasive squamous cell cancer 5 and 7 years from the initial treatment. The cure rate for laser excision was better, as a single session of treatment was curative in 33 of 38 patients (87%). In 3 cases the pathology report on the excised specimen showed an unrecognized invasive lesion (12%) and the women underwent radical surgery. The 2 remaining patients, both affected by multifocal disease, experienced recurrences and were treated with laser excision 2 and 3 years after the primary treatment, respectively. Symptom relief was obtained in all patients studied with both laser vaporization and excision. CONCLUSIONS: Excisional laser surgery is an effective treatment for patients with VIN. In addition, CO(2) laser excision allows evaluation of the operative specimen and detection of occult early invasion with good preservation of vulvar morphology; laser vaporization, while retaining good cosmetic results, is less effective in VIN treatment and does not allow evaluation of the surgical specimen. PMID- 10525385 TI - Primary uterine angiosarcoma. AB - OBJECTIVE: The aim of this study was to report the first case of primary uterine angiosarcoma described in a Hispanic American woman and to review the literature on uterine angiosarcomas. We review characteristic presenting symptoms, gross and microscopic pathologic findings, and treatment outcomes where available. METHODS: A case report is presented with a review of the English language literature via a Medline search. The key phrases used in the search were uterine angiosarcoma, hemangiosarcoma, hemangioendothelioma, and primary uterine neoplasm. RESULTS: Since the first report in 1902, there have been 19 reported cases of primary uterine angiosarcoma considered valid. Many early cases are questioned due to the lack of ultrastructural and immunohistochemical evidence available in later cases. Seventy-four percent (14 of 19) of these patients are perimenopausal with a mean age of 55 years (range 17-76 years). The common presenting findings are a pelvic mass, menorrhagia, and weight loss. Varying regimens of surgery, chemotherapy, and radiation have been utilized with limited success. CONCLUSIONS: Primary uterine angiosarcomas tend to exhibit a highly malignant behavior. The predominant prognostic factor seems to be the size of the tumor at diagnosis and the presence of extrapelvic disease. Recurrence occurs on average at 8.2 months. Of evaluable patients (n = 14), at 12 months the survival was only 43%. Although radiation and chemotherapy are options being utilized, no consensus exists for optimal therapy given the few cases from which to draw conclusions. Regardless of treatment, outcome is usually poor. PMID- 10525387 TI - Recurrent squamous cell carcinoma of the Bartholin's duct treated with en bloc resection. AB - Bartholin's gland carcinomas are a rare entity. A case of a recurrent Bartholin's gland carcinoma is described. These neoplasms have a myriad of treatment options for primary therapy but there is a paucity of information regarding treatment for a lethal recurrence. The patient's primary therapy consisted of an initial wide local excision followed by radiation therapy with chemosensitization. She was disease-free for 2 years before her recurrence. A novel treatment approach incorporating a mulitdisciplinary en bloc radical surgery is described. The patient is alive and well without evidence of disease at 22 months. PMID- 10525388 TI - Vulval squamous cell carcinoma arising in chronic hidradenitis suppurativa. AB - Hidradenitis suppurativa is a chronic inflammatory disease of the sweat glands and hair follicles which is rarely associated with squamous cell carcinoma (SCC). A case of vulval SCC complicating hidradenitis suppurativa is presented. In addition to being the first case to report the association of vulval SCC and hidradenitis suppurativa in the English language literature, it also illustrates the difficulty inherent in distinguishing between invasive SCC and pseudoepitheliomatous hyperplasia on histological examination. The success of wide local excision of the vulval lesion and primary closure without recourse to skin grafts, rotational flaps, or healing by secondary intention is demonstrated. PMID- 10525389 TI - Hemiballismus and brain metastases from squamous cell carcinoma of the cervix. AB - BACKGROUND: Brain metastases from cervical carcinoma are rare. Accompanying symptoms depend on the location of the metastatic lesions. Hemiballismus refers to a rare movement disorder characterized by involuntary, large amplitude movements of the limbs of one side of the body. The area of the brain controlling the limb movement is in the subthalamic nucleus of the contralateral side. In contrast, the usual location of brain metastases from cervical cancer is in the frontal and parietal parenchyma. There have been reported cases of hemiballismus secondary to metastatic carcinoma of the breast, lung, and gall bladder. This is the first reported case of putative cervical cancer metastases associated with hemiballismus. CASE: A 38-year-old Caucasian female was diagnosed with FIGO stage II-B poorly differentiated squamous cell carcinoma of the cervix. Para-aortic lymph nodes were positive for metastatic disease. The patient was treated by radiation with hydroxyurea chemosensitization. Four months after the initial diagnosis she presented with acute onset of hemiballismus. Magnetic resonance imaging of the head revealed a solitary lesion in the left cerebral peduncle extending into the inferior aspect of the left basal ganglia complex. The lesion was inaccessible to biopsy or excision. Palliative radiation therapy to the brain was unsuccessful and the patient expired 1 year following primary presentation. CONCLUSION: Treatment of hemiballismus is directed to its underlying causes. Some brain metastases from cervical cancer may be palliated or even cured by surgical resection and radiation therapy. Although not conclusive, it appears that hemiballismus in a setting of metastatic cervical cancer has a poor prognosis and little benefit from irradiation. PMID- 10525390 TI - Ovarian steroid cell tumors, not otherwise specified: a case report and literature review. AB - Steroid cell tumors, not otherwise specified, are rare ovarian sex cord-stromal tumors with malignant potential. The majority of these tumors produce steroids with testosterone being the most common. A case of a 46-year-old woman who presented with sudden onset of virilization and a pelvic mass is reported. Various aspects of the presentation, diagnosis, and treatment of these tumors are discussed. PMID- 10525391 TI - Endometrial adenocarcinoma in pregnancy. AB - OBJECTIVE: The coexistence of endometrial adenocarcinoma and pregnancy is rare. Most cases are discovered in the first trimester due to irregular bleeding or spontaneous abortion. CASE: A 44-year-old woman, gravida 3, para 2, was admitted due to abnormal vaginal bleeding. After complete history, physical examination, and laboratory evaluation, she was diagnosed with spontaneous abortion and underwent a suction curettage. Pathological examination of the tissue included chorionic villi and an area of atypical hyperplasia and endometrial cancer. CONCLUSION: Recent association between first-trimester spontaneous abortions and subsequent endometrial cancer makes these rare cases of concurrent endometrial cancer and first trimester of pregnancy attractive in that they may disclose insights into the pathophysiology of hormone-dependent cancers. PMID- 10525392 TI - Is uterine serous papillary carcinoma a BRCA1-related disease? Case report and review of the literature. AB - OBJECTIVES: Type II endometrial carcinomas are estrogen-independent and have adverse histologic features and a substantially poorer prognosis. No risk factors have been identified. Interestingly, there is a striking clinical and histopathological similarity between serous papillary carcinomas of the ovary (OSPC), endometrium, and peritoneal cavity, suggesting a common oncogenic mechanism. Several common molecular alterations were found using molecular comparative analysis of OSPC and uterine serous papillary carcinoma (USPC). Germline mutations in the BRCA1 tumor suppressor gene predispose to breast and ovarian cancer but no association with sporadic endometrial cancer has been found. A family of Ashkenazi Jewish origin, in which one sister was first diagnosed with USPC and the second diagnosed with OSPC, led to the hypothesis that a BRCA mutation may contribute to USPC. METHODS: Genomic DNA from both patients as well as two unaffected siblings was analyzed for the three mutations common in Ashkenazi Jews. Loss of heterozygosity (LOH) analysis was performed on DNA extracted from USPC tumor tissue. RESULTS: Both affected sisters tested positive for BRCA1 5382insC germline mutation. LOH analysis confirmed the results. CONCLUSIONS: We present a breast-ovarian cancer family including two sisters with advanced serous papillary carcinomas of endometrial and ovarian origins, carrying the same BRCA1 mutation (5382insC). LOH analysis on USPC tumor DNA showed loss of the wild-type allele, suggesting a causal relationship between the germline BRCA1 mutation and USPC. We believe a study examining BRCA1 mutations in a large cohort of women with this high-risk endometrial carcinoma is warranted. A positive finding may have implications for surveillance and prophylactic surgery in carriers of BRCA1 mutations. PMID- 10525393 TI - Multimodality therapy for carcinoma of the Bartholin gland. AB - OBJECTIVE: The aim of this study was to report the value of chemoradiation in the management of cancers of the Bartholin gland. METHODS: Primary treatment consisting of 45-46 Gy teletherapy radiation to the vulva, pelvis, and groins in combination with 50 mg/m(2) of cisplatin and 1000 mg/m(2)/day of 5-fluorouracil for 5 days during the first and fifth weeks of irradiation was delivered, followed by interstitial implant or excision. RESULTS: Two patients were free of disease at 30 and 59 months following therapy. Both patients required myocutaneous flap closure, one after excision of tumor after radiation and one after radionecrosis at the implant site. CONCLUSIONS: Primary chemoradiation may allow continence-sparing therapy for women with primary carcinoma of the Bartholin gland. PMID- 10525395 TI - Kin competition, the cost of inbreeding and the evolution of dispersal AB - Dispersal is often presented as a mechanism to avoid competition among relatives and inbreeding depression. However, the formal analysis of the effects of both these factors on the evolution of dispersal has only been conducted in few studies with strong restrictive assumptions. In this paper, I first derive the evolutionary stable dispersal rate as a function of three parameters: (1) the cost of dispersal, c, (2) the coefficient of relatedness among randomly chosen offspring, R, and (3) the cost of inbreeding, delta. In a second step, relatedness is used as a dynamical variable for the derivation of the evolutionarily stable dispersal rate. Finally, in a third step, relatedness and the cost of inbreeding are assumed to be dynamical variables. This allows to analyse the more realistic situation where dispersal, relatedness and the cost of inbreeding are coevolving simultaneously. Several subcases are considered depending on the genetic determinism (haploid or diploid), the control of the dispersal strategy (parent or offspring control of dispersal) and the plasticity of dispersal with sexes (with or without sex-specific dispersal rates). This analysis clarifies the role of the cost of inbreeding and kin competition on the evolution of dispersal (in particular on the evolution of sex-biased dispersal rates) and leads to quantitative and testable predictions. Copyright 1999 Academic Press. PMID- 10525394 TI - Radiopharmaceutical-guided surgery in primary malignant melanoma of the vagina. AB - The sentinel lymph node located in the right iliac basin was successfully pre- and intraoperatively identified by radiopharmaceutical-directed mapping in a case of primary malignant melanoma of the vagina. PMID- 10525396 TI - Amplification and spread of viruses in a growing plaque. AB - The two-dimensional propagation of viruses through a "lawn" of receptive hosts, commonly called plaque growth, reflects the dynamics of interactions between viruses and host cells. Here we treat the amplification of viruses during plaque growth as a reaction-diffusion system, where interactions among the virus, uninfected host cells, and virus-producing host-virus complexes are accounted for using rates of viral adsorption to and desorption from the host-cell surface, rates of reproduction and release of progeny viruses by lysis of the host, and by the coupling of these reactions with diffusion of free virus within the agar support. Numerical solution of the system shows the development of a traveling wave of reproducing viruses, where the velocity of the wave is governed by the kinetic and diffusion parameters. The model has been applied to predict the propagation velocity of a bacteriophage plaque. Different mechanisms may account for the dependence of this velocity on the host density during early stages of a growing plaque. The model provides a means to explore how changes in the virus host interactions may be manifest in a growing plaque. PMID- 10525397 TI - Simplified dynamics of human and mammalian neocortical neurons. AB - The behavior of human and mammalian neocortical neurons is governed by the interplay of approximately a dozen ion currents. Due to the complexity of the resulting dynamics, it is sometimes advantageous to resort to simpler systems in order to gain insight into the relationship between underlying dynamical principles and biophysics. This paper presents a new and extremely simple approximation to the dynamics of neocortical neurons based on just four simulated ion currents: I(Na), I(K), I(T), and I(AHP). The formulation incorporates Ohm's law plus explicit representation of Na(+), K(+), and Ca(2+)equilibrium potentials, yet mathematical simplicity is retained by restriction of the dynamics to cubic nonlinearities. The resulting equations produce a good approximation to spike shapes, firing rates, and bursting behavior throughout the physiological range. Analysis of the equations suggests that four unique dynamical regimes form the basis for different categories of neocortical neurons. Synaptic coupling between these model neurons demonstrates their potential utility in simulations by producing network models of bursting and of short-term memory function. PMID- 10525398 TI - A quantitative theory of affinity-driven T cell repertoire selection. AB - Binding of the T cell antigen receptor (TCR) to peptides presented on molecules encoded by major histocompatibility complex (MHC) genes is the key event driving T cell development and activation. Selection of the T cell repertoire in the thymus involves two steps. First, positive selection promotes the survival of cells binding thymic self-MHC-peptide complexes with sufficient affinity. The resulting repertoire is self-MHC restricted: it recognizes foreign peptides presented on self, but not foreign MHC. Second, negative selection deletes cells which may be potentially harmful because their receptors interact with self-MHC peptide complexes with too high an affinity. The mature repertoire is also highly alloreactive: a large fraction of T cells respond to tissues harboring foreign MHC. We derive mathematical expressions giving the frequency of alloreactivity, the level of self-MHC restriction, and the fraction of the repertoire activated by a foreign peptide, as a function of the parameters driving the generation and selection of the repertoire: self-MHC and self-peptide diversity, the stringencies of positive and negative selection, and the number of peptide and MHC polymorphic residues that contribute to T cell receptor binding. Although the model is based on a simplified digit string representation of receptors, all the parameters but one relate directly to experimentally determined quantities. The only parameter without a biological counterpart has no effect on the model's behavior besides a trivial and easily preventable discretization effect. We further analyse the role of the MHC and peptide contribution to TCR binding, and find that their relative, rather than absolute value, is important in shaping the mature repertoire. This result makes it possible to adopt different physical interpretations for the digit string formalism. We also find that the alloreactivity level can be inferred directly from data on the stringency of selection, and that, in agreement with recent experiments, it is not affected by thymic selection. PMID- 10525399 TI - Evolution of cooperation in spatially structured populations AB - Using a spatial lattice model of the Iterated Prisoner's Dilemma we studied the evolution of cooperation within the strategy space of all stochastic strategies with a memory of one round. Comparing the spatial model with a randomly mixed model showed that (1) there is more cooperative behaviour in a spatially structured population, (2) PAVLOV and generous variants of it are very successful strategies in the spatial context and (3) in spatially structured populations evolution is much less chaotic than in unstructured populations. In spatially structured populations, generous variants of PAVLOV are found to be very successful strategies in playing the Iterated Prisoner's Dilemma. The main weakness of PAVLOV is that it is exploitable by defective strategies. In a spatial context this disadvantage is much less important than the good error correction of PAVLOV, and especially of generous PAVLOV, because in a spatially structured population successful strategies always build clusters. Copyright 1999 Academic Press. PMID- 10525400 TI - The evolution of quantitatively responsive cooperative trade. AB - The iterated Prisoner's Dilemma reflects the essence of repeated cooperative interactions with selfish incentives. However, the classical form of this game assumes that individuals either cooperate or defect, whereas in practice different degrees of cooperation are usually possible. To overcome this limitation, we present a model of alternating cooperative trade in which individuals controlled the costs they incurred in benefiting their partners. Since the range of possible strategies is enormous, competitively successful solutions were identified using a genetic algorithm, a powerful search technique in which good performers are iteratively selected and recombined from an initial "strategy soup". Beginning with a population of asocial individuals, altruistic behaviour readily emerged. Like the pre-defined strategy of "Raise-the-Stakes", the emerging strategies evolved protection from cheats by investing relatively little in strangers and subsequently responding quantitatively to a partner's altruism. Unlike "Raise-the-Stakes", they began trading relations at intermediate levels and, when the benefit-to-cost ratio of cooperation was relatively low, mean investment was considerably below the maximum level. Our approach is novel in allowing us to predict not just whether cooperation will occur, but how cooperative individuals will be, in relation to factors such as the number of rounds and the cost effectiveness of cooperative trade. PMID- 10525401 TI - Kinetic analysis of parallel reactions, the initial reactant of which presents with two interconvertible isomeric forms (hydrolysis and hydroxylaminolysis of 6 phosphogluconolactone) AB - A mathematical treatment is presented of two parallel reactions each one of which can be described by the sum of two exponential functions of time. The two simplest reaction models consistent with this requirement are presented, and are shown to be described by a second-order non-homogeneous differential equation with constant coefficients. Solutions of this equation in closed form are given, and it is shown that, as is the case with pairs of parallel first-order reactions, description of the time course of one of the two reactions yields a complete description of the time course of the other reaction. Hypothetical cases, consistent with the reaction models presented, are given and are compared with experimental results, obtained on the parallel reactions of 6 phosphogluconolactone hydrolysis and hydroxylaminolysis. Results are compatible with the existence of enzymically generated 6-phosphogluconolactone in two interconvertible isomeric forms. Copyright 1999 Academic Press. PMID- 10525402 TI - Crystal structure of the C-terminal SH2 domain of the p85alpha regulatory subunit of phosphoinositide 3-kinase: an SH2 domain mimicking its own substrate. AB - The binding properties of Src homology-2 (SH2) domains to phosphotyrosine (pY) containing peptides have been studied in recent years with the elucidation of a large number of crystal and solution structures. Taken together, these structures suggest a general mode of binding of pY-containing peptides, explain the specificities of different SH2 domains, and may be used to design inhibitors of pY binding by SH2 domain-containing proteins. We now report the crystal structure to 1.8 A resolution of the C-terminal SH2 domain (C-SH2) of the P85alpha regulatory subunit of phosphoinositide 3-kinase (PI3 K). Surprisingly, the carboxylate group of Asp2 from a neighbouring molecule occupies the phosphotyrosine binding site and interacts with Arg18 (alphaA2) and Arg36 (betaB5), in a similar manner to the phosphotyrosine-protein interactions seen in structures of other SH2 domains complexed with pY peptides. It is the first example of a non-phosphate-containing, non-aromatic mimetic of phosphotyrosine binding to SH2 domains, and this could have implications for the design of substrate analogues and inhibitors. Overall, the crystal structure closely resembles the solution structure, but a number of loops which demonstrate mobility in solution are well defined by the crystal packing. C-SH2 has adopted a binding conformation reminiscent of the ligand bound N-terminal SH2 domain of PI3K, apparently induced by the substrate mimicking of a neighbouring molecule in the crystal. PMID- 10525403 TI - Interaction of the Escherichia coli DEAD box protein DbpA with 23 S ribosomal RNA. AB - The Escherichia coli DEAD box protein DbpA is unique among the DEAD box family in that its ATPase activity is specifically stimulated by bacterial 23 S ribosomal RNA. We have analysed the interaction between DbpA and a specific region within 23 S rRNA (namely nucleotides 2508-2580) which stimulates full ATPase activity. Using electrophoretic mobility shift assays we show that DbpA binds to this "specific" region with greater efficiency than to other regions of 23 S rRNA, and is not competed off by a non-specific RNA or a mutant RNA in which one of the stem-loops has been disrupted. These data suggest that the secondary structure within this region of 23 S rRNA is important for its recognition and binding by DbpA. We have also examined the ability of DbpA to unwind RNA and show that the purified protein does not behave as an RNA helicase in vitro with the substrates tested. PMID- 10525404 TI - Site-specific chromosomal integration in mammalian cells: highly efficient CRE recombinase-mediated cassette exchange. AB - Expression of experimental constructs in mammalian cells or transgenic animals is difficult to control because it is markedly influenced by position effects. This has limited both the analysis of cis -DNA regulatory elements for transcription and replication, and the physiological analysis of proteins expressed from transgenes. We report here two new methods based on the concept of recombinase mediated cassette exchange (RMCE) to perform site-specific chromosomal integration. The first method permits the exchange of a negative selectable marker pre-localized on the chromosome with a transgene via a CRE-mediated double recombination between inverted Lox sites. Integration efficiency is close to 100 % of negatively selected mouse erythroleukemia cells and ranges from 10 to 50 % in embryonic stem cells. The second method allows RMCE with no selection at all except for cells that have taken up plasmid transiently. While less efficient, this technique permits novel experimental approaches. We find that integration of a transgene at a given genomic site leads to reproducible expression. RMCE should be useful to develop artificial genetic loci that impart specific and reproducible regulation of transgenes in higher eukaryotes. This should facilitate the analysis of cis -regulatory DNA elements governing expression and position effects, improve our control over the physiological effects of transgenes, and accelerate the development of animal models for complex human diseases. PMID- 10525405 TI - The DNA translocation and ATPase activities of restriction-deficient mutants of Eco KI. AB - Eco KI, a type I restriction enzyme, specifies DNA methyltransferase, ATPase, endonuclease and DNA translocation activities. One subunit (HsdR) of the oligomeric enzyme contributes to those activities essential for restriction. These activities involve ATP-dependent DNA translocation and DNA cleavage. Mutations that change amino acids within recognisable motifs in HsdR impair restriction. We have used an in vivo assay to monitor the effect of these mutations on DNA translocation. The assay follows the Eco KI-dependent entry of phage T7 DNA from the phage particle into the host cell. Earlier experiments have shown that mutations within the seven motifs characteristic of the DEAD-box family of proteins that comprise known or putative helicases severely impair the ATPase activity of purified enzymes. We find that the mutations abolish DNA translocation in vivo. This provides evidence that these motifs are relevant to the coupling of ATP hydrolysis to DNA translocation. Mutations that identify an endonuclease motif similar to that found at the active site of type II restriction enzymes and other nucleases have been shown to abolish DNA nicking activity. When conservative changes are made at these residues, the enzymes lack nuclease activity but retain the ability to hydrolyse ATP and to translocate DNA at wild-type levels. It has been speculated that nicking may be necessary to resolve the topological problems associated with DNA translocation by type I restriction and modification systems. Our experiments show that loss of the nicking activity associated with the endonuclease motif of Eco KI has no effect on ATPase activity in vitro or DNA translocation of the T7 genome in vivo. PMID- 10525406 TI - Ataxia in prion protein (PrP)-deficient mice is associated with upregulation of the novel PrP-like protein doppel. AB - The novel locus Prnd is 16 kb downstream of the mouse prion protein (PrP) gene Prnp and encodes a 179 residue PrP-like protein designated doppel (Dpl). Prnd generates major transcripts of 1.7 and 2.7 kb as well as some unusual chimeric transcripts generated by intergenic splicing with Prnp. Like PrP, Dpl mRNA is expressed during embryogenesis but, in contrast to PrP, it is expressed minimally in the CNS. Unexpectedly, Dpl is upregulated in the CNS of two PrP-deficient (Prnp(0/0)) lines of mice, both of which develop late-onset ataxia, suggesting that Dpl may provoke neurodegeneration. Dpl is the first PrP-like protein to be described in mammals, and since Dpl seems to cause neurodegeneration similar to PrP, the linked expression of the Prnp and Prnd genes may play a previously unrecognized role in the pathogenesis of prion diseases or other illnesses. PMID- 10525407 TI - Mitochondrial localization and oligomeric structure of HClpP, the human homologue of E. coli ClpP. AB - A bacterially expressed recombinant HClpP protein, the human homologue of Escherichia coli ClpP protease, was used to obtain specific polyclonal antibodies. Those antibodies identify a 26 kDa polypeptide in mitochondrial subcellular fractions of rat and human liver. Immunofluorescence and electron microscopic studies demonstrate that the mammalian homologue of ClpP is located in the mitochondrial matrix with a tendency to be found in association with the inner mitochondrial membrane. An HClpP recombinant protein with a truncated NH2terminus (missing the first 58 amino acid residues) shows a molecular mass of 26 kDa under denaturing conditions. This N-truncated HClpP recombinant protein shows a native molecular mass of 340 kDa that is identical with the native molecular mass of the partially purified protein from rat liver mitochondria. Electron microscopy shows that the N-truncated recombinant HClpP has a ring shape with seven identical morphological units in the periphery, exhibiting a 7-fold symmetry. The native molecular mass and the electron microscopic studies suggest that mitochondrial ClpP is composed of two heptameric rings with 7-fold symmetry, similar to E. coli ClpP. PMID- 10525408 TI - Molecular modeling of the three-dimensional architecture of the RNA component of yeast RNase MRP. AB - RNase mitochondrial RNA processing (MRP) is a ribonucleoprotein endoribonuclease that is involved in RNA processing events in both the nucleus and the mitochondria. The MRP RNA is both structurally and evolutionarily related to RNase P, the ribonucleoprotein endoribonuclease that processes the 5'-end of tRNAs. Previous analysis of the RNase MRP RNA by phylogenetic analysis and chemical modification has revealed strikingly conserved secondary structural elements in all characterized RNase MRP RNAs. Utilizing successive constraint modeling and energy minimization I derived a three-dimensional model of the yeast RNase MRP RNA. The final model predicts several notable features. First, the enzyme appears to contain two separate structural domains, one that is highly conserved among all MRP and P RNAs and a second that is only conserved in MRP RNAs. Second, nearly all of the highly conserved nucleotides cluster in the first domain around a long-range interaction (LRI-I). This LRI-I is characterized by a ubiquitous uridine base, which points into a cleft between these two structural domains generating a potential active site for RNA cleavage. Third, helices III and IV (the yeast equivalent of the To-binding site) model as a long extended helix. This region is believed to be the binding site of shared proteins between RNase P and RNase MRP and would provide a necessary platform for binding these seven proteins. Indeed, several residues conserved between the yeast MRP and P RNAs cluster in the central region of these helixes. Lastly, characterized mutations in the MRP RNA localize in the model based on their severity. Those mutations with little or no effect on the activity of the enzyme localize to the periphery of the model, while the most severe mutations localize to the central portion of the molecule where they would be predicted to cause large structural defects. Press. PMID- 10525409 TI - Structure of recombinant mouse collagenase-3 (MMP-13). AB - The matrix metalloproteinases are crucial in the physiological and pathological degradation of the mammalian extracellular matrix, including breast tumours, and osteoarthritic cartilage. These enzymes are classified according to their matrix substrate specificity. Collagenase-3 (MMP-13) is a member of this family and preferentially cleaves type II collagen, cartilage, fibronectin and aggrecan. Collagenase-3 is normally expressed in hypertrophic chondrocytes, periosteal cells, and osteoblasts during bone development. The structure of the catalytic domain of recombinant mouse collagenase-3, complexed to the hydroxamate inhibitor (RS-113456), is reported at 2.0 A resolution. Molecular replacement and weak phasing information from a single derivative determined the structure. Neither molecular replacement nor derivative methods had a sufficient radius of convergence to yield a refinable structure. The structure illuminates the atomic zinc ion interactions with functional groups in the active site, emphasizing zinc ligation and the very voluminous hydrophobic P1' group for the inhibitor potency. The structure provides insight into the specificity of this enzyme, facilitating design of specific inhibitors to target various diseases. PMID- 10525410 TI - Protein titration in the crystal state. AB - Proteins are complex structures whose overall stability critically depends on a delicate balance of numerous interactions of similar strength, which are markedly influenced by their environment. Here, we present an analysis of the effect of pH on a protein structure in the crystalline state using RNase A as a model system. By altering only one physico-chemical parameter in a controlled manner, we are able to quantify the structural changes induced in the protein. Atomic resolution X-ray diffraction data were collected for crystals at six pH* values ranging from 5.2 to 8.8, and the six independently refined structures reveal subtle, albeit well-defined variations directly related to the pH titration of the protein. The deprotonation of the catalytic His12 residue is clearly evident in the electron density maps, confirming the reaction mechanism proposed by earlier enzymatic and structural studies. The concerted structural changes observed in the regions remote from the active-site point to an adaptation of the protein structure to the changes in the physico-chemical environment. Analysis of the stereochemistry of the six structures provided accurate estimates of p Kavalues of most of the histidine residues. This study gives further evidence for the advantage of atomic resolution X-ray crystallographic analyses for revealing small but significant structural changes which provide clues to the function of a biological macromolecule. PMID- 10525411 TI - Insight into odorant perception: the crystal structure and binding characteristics of antibody fragments directed against the musk odorant traseolide. AB - Monoclonal antibodies were elicited against the small hydrophobic hapten traseolide, a commercially available musk fragrance. Antibody variable region sequences were found to belong to different sequence groups, and the binding characteristics of the corresponding antibody fragments were investigated. The antibodies M02/01/01 and M02/05/01 are highly homologous and differ in the binding pocket only at position H93. M02/05/01 (H93 Val) binds the hapten traseolide about 75-fold better than M02/01/01 (H93 Ala). A traseolide analog, missing only one methyl group, does not have the characteristic musk odorant fragrance. The antibody M02/05/01 binds this hapten analog about tenfold less tightly than the original traseolide hapten, and mimics the odorant receptor in this respect, while the antibody M02/01/01 does not distinguish between the analog and traseolide. To elucidate the structural basis for the fine specificity of binding, we determined the crystal structure of the Fab fragment of M02/05/01 complexed with the hapten at 2.6 A resolution. The crystal structure showed that only van der Waals interactions are involved in binding. The somatic Ala H93 Val mutation in M02/05/01 fills up an empty cavity in the binding pocket. This leads to an increase in binding energy and to the ability to discriminate between the hapten traseolide and its derivatives. The structural understanding of odorant specificity in an antibody gives insight in the physical principles on how specificity for such hydrophobic molecules may be achieved. PMID- 10525412 TI - New insights into structure-function relationships in nitrogenase: A 1.6 A resolution X-ray crystallographic study of Klebsiella pneumoniae MoFe-protein. AB - The X-ray crystal structure of Klebsiella pneumoniae nitrogenase component 1 (Kp1) has been determined and refined to a resolution of 1.6 A, the highest resolution reported for any nitrogenase structure. Models derived from three 1.6 A resolution X-ray data sets are described; two represent distinct oxidation states, whilst the third appears to be a mixture of both oxidized and reduced states (or perhaps an intermediate state). The structures of the protein and the iron-molybdenum cofactor (FeMoco) appear to be largely unaffected by the redox status, although the movement of Ser beta90 and a surface helix in the beta subunit may be of functional significance. By contrast, the 8Fe-7S P-cluster undergoes discrete conformational changes involving the movement of two iron atoms. Comparisons with known component 1 structures reveal subtle differences in the FeMoco environment, which could account for the lower midpoint potential of this cluster in Kp1. Furthermore, a non-proline- cis peptide bond has been identified in the alpha subunit that may have a functional role. It is within 10 A of the FeMoco and may have been overlooked in other component 1 models. Finally, metal-metal and metal-sulphur distances within the metal clusters agree well with values derived from EXAFS studies, although they are generally longer than the values reported for the closely related protein from Azotobacter vinelandii. A number of bonds between the clusters and their ligands are distinctly longer than the EXAFS values, in particular, those involving the molybdenum atom of the FeMoco. PMID- 10525413 TI - Mutational analysis and NMR spectroscopy of quail cysteine and glycine-rich protein CRP2 reveal an intrinsic segmental flexibility of LIM domains. AB - The LIM domain is a conserved cysteine and histidine-containing structural module of two tandemly arranged zinc fingers. It has been identified in single or multiple copies in a variety of regulatory proteins, either in combination with defined functional domains, like homeodomains, or alone, like in the CRP family of LIM proteins. Structural studies of CRP proteins have allowed a detailed evaluation of interactions in LIM-domains at the molecular level. The packing interactions in the hydrophobic core have been identified as a significant contribution to the LIM domain fold, whereas hydrogen bonding within each single zinc binding site stabilizes zinc finger geometry in a so-called "outer" or "indirect" coordination sphere. Here we report the solution structure of a point mutant of the carboxyl-terminal LIM domain of quail cysteine and glycine-rich protein CRP2, CRP2(LIM2)R122A, and discuss the structural consequences of the disruption of the hydrogen bond formed between the guanidinium side-chain of Arg122 and the zinc-coordinating cysteine thiolate group in the CCHC rubredoxin knuckle. The structural analysis revealed that the three-dimensional structure of the CCHC zinc binding site in CRP2(LIM2)R122A is adapted as a consequence of the modified hydrogen bonding pattern. Additionally, as a result of the conformational rearrangement of the zinc binding site, the packing interactions in the hydrophobic core region are altered, leading to a change in the relative orientation of the two zinc fingers with a concomitant change in the solvent accessibilities of hydrophobic residues located at the interface of the two modules. The backbone dynamics of residues located in the folded part of CRP2(LIM2)R122A have been characterized by proton-detected(15)N NMR spectroscopy. Analysis of the R2/R1ratios revealed a rotational correlation time of approximately 6.2 ns and tumbling with an axially symmetric diffusion tensor (D parallel/D perpendicular=1.43). The relaxation data were also analyzed using a reduced spectral density mapping approach. As in wild-type CRP2(LIM2), significant mobility on a picosecond/nanosecond time-scale was detected, and conformational exchange on a microsecond time-scale was identified for residues located in loop regions between secondary structure elements. In summary, the relative orientation of the two zinc binding sites and the accessibility of hydrophobic residues is not only determined by hydrophobic interactions, but can also be modified by the formation and/or breakage of hydrogen bonds. This may be important for the molecular interactions of an adaptor-type LIM domain protein in macromolecular complexes, particularly for the modulation of protein-protein interactions. PMID- 10525414 TI - Shape and energetics of a cavity in c-Myb probed by natural and non-natural amino acid mutations. AB - The shape and the energetics of a functional cavity in the R2 subdomain (90-141) of the c-Myb DNA-binding domain were investigated by spectroscopy and thermodynamic analysis. We focused on the valine 103 residue located in front of the cavity. Nine mutants, in which valine 103 was substituted with alanine, 2 aminobutyric acid, norvaline, norleucine, leucine, isoleucine, allo -isoleucine, cyclohexylglycine, and cyclohexylalanine, were chemically synthesized and analyzed. These mutants provided a wide distribution of sizes which ranged from forming additional cavity space to filling and overflowing the cavity space. Temperature-scanning circular dichroism measurements and differential scanning calorimetry revealed a linear relationship between the van't Hoff enthalpy and the thermal transition temperature for the cavity-filling mutations. On the other hand, the mutants with side-chains larger than the side-chain of leucine resulted in a relatively low transition enthalpy and temperature, most likely due to the exposure of the side-chain to solvent and the increase in the entropy of the folded states. Branching at the beta-carbon atom reduced the unfolding free energy due to the steric constraint in the cavity. In particular, the mutational elongation of the side-chain from beta-carbon to the trans -to-CO direction proved to be more hindered than that from beta-carbon to the trans -to-NH. The unfolding free energy versus side-chain volume formed a bell-shaped plot with a maximum free energy for the leucine mutant. The difference in the transition free energy for cavity-filling mutants with beta-unbranched side-chains were two to four times larger than the difference in the transfer energy from organic solvent to water. Therefore, the increase in unfolding free energy would most likely be attributed to van der Waals interactions in the cavity wall, which would be a origin of stabilization by the sliding of tryptophan 95 into the cavity upon DNA binding. PMID- 10525416 TI - Acquisition of novel catalytic activity by the M1 RNA ribozyme: the cost of molecular adaptation. AB - The ribonucleoprotein RNase P is a critical component of metabolism in all known organisms. In Escherichia coli, RNase P processes a vast array of substrates, including precursor-tRNAs and precursor 4. 5S RNA. In order to understand how such catalytic versatility is achieved and how novel catalytic activity can be acquired, we evolve the M1 RNA ribozyme (the catalytic component of E. coli RNase P) in vitro for cleavage of a DNA substrate. In so doing, we probe the consequences of enhancing catalytic activity on a novel substrate and investigate the cost this versatile enzyme pays for molecular adaptation. A total of 25 generations of in vitro evolution yield a population showing more than a 1000 fold increase in DNA substrate cleavage efficiency (kcat/KM) relative to wild type M1 RNA. This enhancement is accompanied by a significant reduction in the ability of evolved ribozymes to process the ptRNA class of substrates but also a contrasting increase in activity on the p4.5S RNA class of substrates. This change in the catalytic versatility of the evolved ribozymes suggests that the acquired activity comes at the cost of substrate versatility, and indicates that E. coli RNase P catalytic flexibility is maintained in vivo by selection for the processing of multiple substrates. M1 RNA derivatives enhance cleavage of the DNA substrate by accelerating the catalytic step (kcat) of DNA cleavage, although overall processing efficiency is offset by reduced substrate binding. The enhanced ability to cleave a DNA substrate cannot be readily traced to any of the predominant mutations found in the evolved population, and must instead be due to multiple sequence changes dispersed throughout the molecule. This conclusion underscores the difficulty of correlating observed mutations with changes in catalytic behavior, even in simple biological catalysts for which three dimensional models are available. PMID- 10525415 TI - In vitro folding and thermodynamic stability of an antibody fragment selected in vivo for high expression levels in Escherichia coli cytoplasm. AB - We recently isolated a mutant of a human anti-beta-galactosidase single chain antibody fragment (scFv) able to fold at high levels in Escherichia coli cytoplasm. When targeted to the periplasm, this mutant and the wild-type scFv are both expressed at comparable levels in a soluble, active and oxidized form. If a reducing agent is added to the growth medium, only the mutant scFv is still able to fold, showing that in vivo aggregation is a direct consequence of the lack of disulphide bond formation and not of the cellular localization. In vitro denaturation/renaturation experiments show that the mutant protein is more stable than the wild-type scFv. Furthermore, refolding kinetics under reducing conditions show that the mutant folds faster than the wild-type protein. Aggregation does not proceed from the native or unfolded conformation of the protein, but from a species only present during the unfolding/refolding transition. In conclusion, the in vivo properties of the mutant scFv can be explained by, first, an increase in the stability of the protein in order to tolerate the removal of the two disulphide bonds and, second, a modification of its folding properties that reduces the kinetic competition between folding and aggregation of a reduced folding intermediate. PMID- 10525417 TI - Frequency-dependent increase in cardiac Ca2+ current is due to reduced Ca2+ release by the sarcoplasmic reticulum. AB - "Ca(2+)-current facilitation" describes several features of increase in current amplitude often associated with a reduction in inactivation rate. The aim of this study was to investigate the mechanism of frequency-dependent increase in L-type Ca2+ current, I(Ca) taking advantage of recent knowledge on the control of Ca2+ current inactivation in cardiac cells. The frequency-dependent increase in I(Ca) was studied in adult rat ventricular myocytes using the whole-cell patch-clamp technique. I(Ca) was elicited by a train of 200-ms depolarizing pulses to +20 mV applied at various frequencies (0.2 up to 1.3 Hz). The increase in frequency induced a rate-dependent enhancement of I(Ca), or facilitation phenomena. In most cells, that showed two inactivation phases of I(Ca), facilitation was mainly related to slowing of the fast I(Ca) inactivation phase that occurred besides increase in peak I(Ca) amplitude. Both the decrease and slowing of the fast component of inactivation phase were attenuated on beta -adrenergic-stimulated current. Frequency-dependent I(Ca) facilitation paralleled a reduction in Ca2+ transient measured with fluo-3. After blocking sarcoplasmic reticulum-Ca2+ release by thapsigargin, the fast I(Ca) inactivation phase was reduced and facilitation was eliminated. Facilitation could not then be restored by 1 microM isoprenaline. Thus in rat ventricular myocytes, frequency-dependent facilitation of I(Ca)reflects a reduced Ca(2+)-dependent inactivation consecutive, in most part, to reduced Ca2+ load and Ca2+ release by the sarcoplasmic reticulum rather than being an intrinsic characteristic of the L-type Ca2+ channel. PMID- 10525418 TI - Efficient limitation of intracellular edema and sodium accumulation by cardioplegia is dissociated from recovery of rat hearts from cold ischemic storage. AB - Energy deficiency and disturbances of sodium and water homeostasis are considered as mechanisms of injury during hypothermic preservation of cardiac muscle. The present study attempts to characterize the effect of potassium (K+) and magnesium (Mg2+) cardioplegia on these mechanisms. Cellular parameters were measured by multinuclear NMR spectroscopy in isolated rat hearts during 12 h of ischemia at 4 degrees C and 2 h of normothermic reperfusion with an isoosmotic Krebs-Henseleit (KH) solution. Potassium and magnesium cardioplegia (a) reduced the rate of ATP hydrolysis and cellular acidification during early stages of ischemia; (b) caused an early cessation of the phase of fast sodium influx after 40 min (P<0.001 vs 120 min with KH); (c) reduced intracellular sodium accumulation to 148-165 micromol/gdw after 12 h (P<0.01 vs 268+/-15 micromol/gdw with KH); (d) decreased ischemic volumes to 2.7+/-0.1 and 2.8+/-0.1 ml/gdw after 8 and 12 h of storage, respectively (P<0.005 v 3.0 and 3.3 ml/gdw with KH). Quantitative analysis of these parameters showed that both hypothermia and cardioplegia increased the relative contribution of sodium to intracellular water accumulation by a factor of 2-2.5. In view of the marked reduction in absolute sodium and water contents, the data indicate that cold cardioplegia limits the increase in intracellular osmolarity. Myocardial mechanical and metabolic recoveries, and cellular viability deteriorated during prolongation of the ischemic period from 8 to 12 h in all experimental groups (P<0.005). Reperfusion was efficient in reversing intracellular sodium and water accumulation in hearts stored with cardioplegia, in contrast to hearts stored in KH. Magnesium, but not potassium cardioplegia, lowered interstitial water contents (P<0.01 v KH), increased intracellular magnesium concentrations (P<0.001), improved mechanical and metabolic recoveries (P<0.01) and cellular viability (P<0.001). These results indicate (a) cardioplegia reduces intracellular sodium (by approximately 46%) and water accumulation (by 66%) during cold ischemia; (b) both hypothermia and cardioplegia limit the rise in intracellular osmolarity and increase the contribution of sodium to cellular swelling; (c) intracellular sodium and water contents were dissociated from myocardial viability and recovery from cold ischemia in potassium and magnesium cardioplegic solutions. It is concluded that intracellular sodium and water accumulation are not dominant factors in determination of cardiac outcome from ischemia. PMID- 10525419 TI - Anti-arrhythmic effect of kappa-opioid receptor stimulation in the perfused rat heart: involvement of a cAMP-dependent pathway. AB - During myocardial ischaemia the beta-adrenoceptor is activated, which contributes, at least partly, to cardiac arrhythmias via inducing [Ca2+]i oscillations. Since beta-adrenoceptor is negatively modulated by the kappa-opioid receptor in the heart, the present study attempted to determine if kappa-opioid receptor stimulation modulates the arrhythmogenic action of beta-adrenoceptor stimulation and to delineate the underlying mechanism. The effect of U50,488H, a selective kappa-opioid agonist, on arrhythmias in the isolated perfused rat heart subjected to low flow and 10(-6)mol/l norepinephrine (NE) were determined. Low flow induced arrhythmias, which were potentiated by NE, but not by 10(-6)mol/l U50,488H. The arrhythmia-potentiating effect of NE was antagonized by 10(-6)mol/l propranolol, a beta-adrenoceptor antagonist. U50,488H at 10(-6)mol/l also abolished the potentiation in arrhythmias by NE without affecting the arrhythmias induced by low flow. The anti-arrhythmic action of the kappa-opioid receptor agonist was abolished by 10(-6)mol/l nor-binaltorphimine, a selective kappa opioid receptor antagonist, but not by 10(-7)mol/l calphostin C, an inhibitor of protein kinase C. Similarly, kappa-opioid receptor stimulation with U50,488H also abolished the NE-induced [Ca2+]i oscillations which are believed to cause cardiac arrhythmias, in ventricular myocytes. To determine whether the inhibitory actions of U50,488H against the effects of beta-adrenoceptor stimulation was via a cAMP dependent or a cAMP-independent pathway, we determined the effects of U50,488H on NE-enhanced cAMP production and [Ca2+]i oscillations induced by either forskolin, an activator of adenylate cyclase, or Bay K-8644, a selective L-type Ca2+ channel agonist, in the ventricular myocytes. We found that U50,488H abolished the effect of forskolin, but did not alter the effect of Bay K-8644, on [Ca2+]i oscillations in the ventricular myocyte. In addition, U50, 488H also attenuated significantly the NE-induced elevation in cAMP in the heart. The observations suggest that kappa-opioid receptor stimulation abolishes the effect of beta-adrenoceptor stimulation on arrhythmias and [Ca2+]i oscillation via a cAMP-dependent pathway. The finding may be useful for the prevention and treatment of ischaemic heart diseases. PMID- 10525420 TI - Adaptation to high altitude hypoxia protects the rat heart against ischemia induced arrhythmias. Involvement of mitochondrial K(ATP) channel. AB - The aim was to determine whether adaptation to chronic hypoxia protects the heart against ischemic arrhythmias and whether ATP-dependent potassium channels (K(ATP)) play a role in the antiarrhythmic mechanism. Adult male rats were adapted to intermittent high altitude hypoxia (5000 m, 4 h/day) and susceptibility to ischemia-induced ventricular arrhythmias was evaluated in the Langendorff-perfused hearts subjected to either an occlusion of the coronary artery for 30 min or pre-conditioning by brief occlusion of the same artery prior to 30-min reocclusion. In separate groups, either a K(ATP) blocker, glibenclamide (10 micromol/l), or a mitochondrial K(ATP) opener, diazoxide (50 micromol/l), were added to a perfusion medium 20 min before the occlusion. Adaptation to hypoxia reduced the total number of ventricular arrhythmias by 64% as compared with normoxic controls. Preconditioning by a single 3-min coronary artery occlusion was antiarrhythmic only in the normoxic group, while two occlusion periods of 5 min each were needed to pre-condition the hypoxic hearts. Glibenclamide increased the number of arrhythmias in the normoxic hearts from 1316+/-215 to 2091+/-187 (by 59%) and in the hypoxic group from 636+/-103 to 1777+/-186 (by 179%). In contrast, diazoxide decreased the number of arrhythmias only in the normoxic group from 1374+/-96 to 582+/-149 (by 58%), while its effect in the hypoxic group was not significant. It is concluded that long-term adaptation of rats to high altitude hypoxia decreases the susceptibility of their hearts to ischemic arrhythmias and increases an antiarrhythmic threshold of pre conditioning. The mitochondrial K(ATP) channel, rather than the sarcolemmal K(ATP) channel, appears to be involved in the protective mechanism afforded by adaptation. PMID- 10525421 TI - Differential responses of adult cardiac fibroblasts to in vitro biaxial strain patterns. AB - Different patterns of extracellular matrix (ECM) remodeling in the heart are thought to be dependent on altered mechanical and chemical conditions and can contribute to cardiac dysfunction. Cardiac fibroblasts are the primary regulators of the ECM and may respond to mechanical factors in vitro. We hypothesized that different types of in vitro strains, e.g. tensile or compressive, can stimulate different functional responses in cultured adult rat cardiac fibroblasts. In this study, we first showed that a single step in strain applied by a uniaxial stretch system stimulated collagen III and fibronectin mRNA levels and transforming growth factor-beta(1) (TGF-beta(1)) activity in the adult phenotype of rat cardiac fibroblasts. Two-dimensional deformations were measured by tracking fluorescent microspheres attached to the substrate and cultured cells. For 10% uniaxial strain, mean principal strains were 0. 104 +/- 0.018 in the direction of stretch and -0.042 +/- 0.013 in the perpendicular direction, verifying that the fibroblasts were simultaneously subjected to tensile (positive) and compressive (negative) strains. Furthermore, these cells were also subjected to area change and to shear. In order to examine the distinct effects of different types of deformation on cardiac fibroblasts, an equibiaxial stretch system was used to apply either pure tensile or compressive area strains, in the absence of shear. Magnitudes of equibiaxial strain were selected to apply local cell area changes identical to those applied in the uniaxial system. Results showed that pure tensile and compressive area strains induced divergent responses in ECM mRNA levels. TGF-beta(1) activity was dependent on the magnitude of applied area strain regardless of the mode of deformation. These findings demonstrate that adult cardiac fibroblasts may respond differently to varied types of mechanical loading, suggesting that ECM remodeling may be locally regulated by specific mechanical stimuli in the heart. PMID- 10525422 TI - Functional and energetic consequences of chronic myocardial creatine depletion by beta-guanidinopropionate in perfused hearts and in intact rats. AB - Oral feeding with the creatine analogue beta-guanidinopropionate (beta-GP) reduces myocardial phosphocreatine and creatine concentrations by about 80%in vitro, this is accompanied by reduced contractile performance. We hypothesized, thus, that beta-GP feeding leads to hemodynamic changes in vivo characteristic of heart failure. beta-GP was fed to Wistar rats for up to 8 weeks. In isolated hearts, function was measured isovolumically, myocardial energetics were followed with (31)P-NMR spectroscopy. In vivo hemodynamics were measured with Millar-Tip catheters and an electromagnetic flow probe. Beta-GP feeding did not alter heart weight. In vitro, diastolic pressure-volume curves indicated structural left ventricular dilatation, and a 36% reduction of left ventricular developed pressure was found; phosphocreatine was reduced by approximately 80%, ATP unchanged and creatine kinase reaction velocity ((31)P-MR saturation transfer) decreased by approximately 90%. The total creatine pool (high-pressure liquid chromatography) was reduced by up to approximately 70%. In contrast to in vitro findings, in vivo cardiac hemodynamics (including left ventricular developed pressure, d P/d t(max), cardiac output and peripheral vascular resistance) at rest and during acute volume loading showed no alterations after beta-GP feeding. The only functional impairment observed in vivo was a 14% reduction of maximum left ventricular developed pressure during brief aortic occlusion. In the intact rat, cardiac and/or humoral compensatory mechanisms are sufficient to maintain normal hemodynamics in spite of a 90% reduction of creatine kinase reaction velocity. However, chronic beta-GP feeding leads to structural left ventricular dilatation. PMID- 10525423 TI - Decreased myocardial nNOS, increased iNOS and abnormal ECGs in mouse models of Duchenne muscular dystrophy. AB - Duchenne muscular dystrophy is a devastating neuromuscular disease caused by lack of the protein, dystrophin, in skeletal muscle and heart, although the biochemical mechanism by which dystrophin loss causes muscle dysfunction is unknown. Here we show that the dystrophin-deficient mdx mouse and a mouse lacking both dystrophin and the dystrophin-related protein, utrophin (dko), have abnormal electrocardiograms (ECGs). In skeletal muscle, dystrophin is normally associated with neuronal nitric oxide synthase (nNOS) at the sarcolemma. Consequently, we have measured NOS isoform activities in hearts from control, mdx and dko mice. In control mouse hearts, eNOS and nNOS activities increased by 120% and 47%, respectively, between 2 and 6 months of age. In mdx mice, myocardial nNOS activity was decreased by 60%, 84% and 80% at 2, 6 and 12 months of age, respectively. Similarly, hearts from dko mice showed a 65% decrease in nNOS activity compared to controls at 2 months of age. Endothelial NOS (eNOS) activity was not affected by dystrophin loss, but inducible NOS (iNOS) activity was seven fold higher than control in the mdx mouse heart by 12 months of age. We conclude that lack of dystrophin in the mdx mouse results in abnormal ECGs that are associated with decreased myocardial nNOS and increased iNOS activities. PMID- 10525424 TI - Antioxidant properties of pyruvate mediate its potentiation of beta-adrenergic inotropism in stunned myocardium. AB - This study tested the hypothesis that pyruvate's antioxidant actions, particularly its enhancement of the endogenous glutathione system, mediate its potentiation of beta-adrenergic inotropism in stunned myocardium. Isolated working guinea pig hearts, metabolizing 10 m M glucose and stunned by 45 min of low flow ischemia, were treated with 5 m M pyruvate, 5 m M N-acetylcysteine (NAC) and/or 2 n M isoproterenol beginning 15 min after reperfusion. The antioxidant NAC alone did not increase cardiac power (mJ/min/g wet: 11 +/- 1 in untreated and 15 +/- 2 in NAC treated stunned hearts), but NAC potentiated the increase in power produced by 2 n M isoproterenol (isoproterenol alone: 50+/-10; NAC plus isoproterenol: 133 +/- 24). Addition of NAC doubled cyclic AMP content but lowered cytosolic phosphorylation potential by 32% in isoproterenol-stimulated hearts. Stunning decreased the glutathione antioxidant ratio (GSH/GSSG) by 68%. The antioxidant ratio was completely restored by pyruvate alone or in combination with isoproterenol, but only partially restored by isoproterenol alone. Combining isoproterenol and NAC increased the GSH/GSSG ratio by an additional 36%. The combined treatment of pyruvate and isoproterenol increased the NADPH/NADP(+) ratio almost three-fold, and produced the greatest accumulation of glucose-6 phosphate of any treatment. CONCLUSIONS: like pyruvate, the antioxidant NAC potentiated beta-adrenergic inotropism of stunned myocardium. Unlike pyruvate, NAC did not increase cellular energy reserves, thus effectively limiting its potentiation of beta-adrenergic stimulation. Thus, pyruvate's potentiation of beta-adrenergic stimulation in stunned myocardium is most likely the result of the combined effects of its antioxidant and energetic properties. PMID- 10525425 TI - Chronic administration of nifedipine induces up-regulation of functional calcium channels in rat myocardium. AB - Previous studies from our laboratory demonstrated the up-regulation of cardiac dihydropyridine (DHP) receptors in rabbits chronically treated with nifedipine (NIFE). The goal of the present study was to further examine the functionality of this increased number of receptors by analysing different steps of excitation contraction coupling mechanism in adult rats chronically treated with NIFE (a single 10-mg oral dose/kg/day for 28 days). Ca2+ channel density was assessed by specific binding at the DHP receptors with [methyl-(3)H]PN 200-110 in rat ventricular membranes. Chronic NIFE treatment produced up-regulation of Ca2+ channels, being the maximal binding capacities 222+/-19 fmol/mg protein (n=14) and 310+/-21 fmol/mg protein (n=11) in untreated and treated animals, respectively (P<0.05). The functional consequences of this up-regulation of Ca2+ channels were determined in isolated ventricular myocytes by measuring L-type Ca2+ currents (I(Ca)) with the whole-cell configuration of patch-clamp technique and by intracellular Ca2+ (Ca2+(i)) transients estimated by the Indo-1/AM fluorescence ratio (410/482) simultaneously monitored with cell shortening. Peak I(Ca) density recorded at 0 mV was 32% greater in myocytes isolated from the treated group than in those obtained from the untreated group (-10.43+/-0.73 pA/pF (n=13) vs-7.10+/-0.59 pA/pF (n=12) P<0.05). Ca2+(i) transient amplitude and cell shortening, explored at 1 and 2 mM extracellular calcium ([Ca]0) were significantly higher in ventricular myocytes obtained fom NIFE-treated rats than in myocytes isolated from untreated animals. At 2 mM [Ca]0, the values of Ca2+(i) transient and shortening were 460+/-61 nM and 11+/-1 % of resting length (L(0)) in myocytes from treated rats (n=9) and 212+/-22 nM and 5.3+/-0.5% of L(0) in myocytes from control rats (n=6, P<0.05). The results demonstrate an up regulation of functionally-active cardiac Ca2+ channels after NIFE treatment, and offer a possible explanation for a "withdrawal effect" at myocardial level after the suppression of the treatment with this drug. PMID- 10525426 TI - Extract from Scutellaria baicalensis Georgi attenuates oxidant stress in cardiomyocytes. AB - Extract from Scutellaria baicalensis Georgi Attenuates Oxidant Stress in Cardiomyocytes. Journal of Molecular and Cellular Cardiology (1999) 31, 1885 1895. Scutellaria baicalensis Georgi is a Chinese herbal medicine used to treat allergic and inflammatory diseases. The medicinal effects of S. baicalensis root may result, in part, from its constituent flavones reported to have antioxidant properties. Since oxidants play multiple roles in cells, we tested whether S. baicalensis could confer protection in a cardiomyocyte model of ischemia and reperfusion. The intracellular fluorescent probes 2',7'-dichlorofluorescin diacetate (DCFH-DA, sensitive to H(2)O(2) and hydroxyl radicals) and dihydroethidium (DHE, sensitive to superoxide) were used to assess intracellular reactive oxygen species (ROS), and propidium iodide (PI) was used to assess viability in cultured embryonic cardiomyocytes. S. baicalensis extract (SbE) quickly attenuated levels of oxidants generated during transient hypoxia and during exposure to the mitochondrial site III inhibitor antimycin A, as measured by DCFH oxidation or by DHE oxidation. These attenuated oxidant levels were associated with improved survival and function. Cell death after ischemia/reperfusion decreased from 47+/-3 % in untreated to 26+/-2 % in S. baicalensis treated cells (P<0.001). After antimycin A exposure, S. baicalensis decreased cell death from 49+/-6 % in untreated to 23+/-4 % in treated cells. Return of contraction occurred in S. baicalensis-treated cells but was not observed in control cells. Other in vitro studies revealed that baicalein, a major flavone component of SbE can directly scavenge superoxide, hydrogen peroxide, and hydroxyl radicals. Collectively, these findings indicate that SbE and its constituent flavones such as baicalein can attenuate oxidant stress and protect cells from lethal oxidant damage in an ischemia-reperfusion model. PMID- 10525427 TI - Elevated levels of endogenous adenosine alter metabolism and enhance reduction in contractile function during low-flow ischemia: associated changes in expression of Ca(2+)-ATPase and phospholamban. AB - Adenosine has several potentially cardioprotective effects including vasodilatation, reduction in heart rate and alterations in metabolism. Adenosine inhibits catecholamine-induced increase in contractile function mainly through inhibition of phosphorylation of phospholamban (PLB), the main regulatory protein of Ca(2+)-ATPase in sarcoplasmic reticulum (SR), and during ischemia it reduces calcium (Ca2+) overload. In this study we examined the effects of endogenous adenosine on contractile function and metabolism during low-flow ischemia (LFI) and investigated whether endogenous adenosine can alter expression of the Ca(2+) ATPase/PLB-system and other Ca(2+)-regulatory proteins. Isolated blood-perfused piglet hearts underwent 120 min 10% flow. Hearts were treated with either saline, the adenosine receptor blocker (8)-sulfophenyl theophylline (8SPT, 300 micromol/l) or the nucleoside transport inhibitor draflazine (1 micromol/l). During LFI, 8SPT did not substantially influence metabolic or functional responses. However, draflazine enhanced the reduction in heart rate, contractile force and MVO(2), with less release of H+ and CO2. Before LFI there were no significant differences between groups for any of the proteins (Ca(2+)-ATPase, ryanodine-receptor, Na+/K(+)-ATPase) or mRNAs (Ca(2+)-ATPase, PLB, calsequestrin, Na+/Ca(2+)-exchanger) measured. At end of LFI mRNA-level of PLB was higher in draflazine-treated hearts compared to both other groups (P<0.01 vs both). Also, at end of LFI protein-level of Ca(2+)-ATPase was lower in draflazine-treated hearts (P<0.05 vs both), and a parallel trend towards a lower mRNA-level was seen (P=0.11 vs saline and P=0.43 vs 8SPT). During LFI tissue Ca2+ tended to rise in saline- and 8SPT-treated hearts but not in draflazine-treated hearts (at end of LFI, P=0.01 vs 8SPT). We conclude that the amount of adenosine normally produced during LFI does not substantially influence function and metabolism. However, increased endogenous levels by draflazine enhance downregulation of function and reduce signs of anaerobic metabolism. At end of LFI associated changes in expression of PLB and Ca(2+)-ATPase were seen. The functional significance was not determined in the present study. However, altered protein-levels might influence Ca(2+)-handling in sarcoplasmic reticulum and thus affect contractile force and tolerance to ischemia. PMID- 10525428 TI - The time course of haemodynamic, autonomic and skeletal muscle metabolic abnormalities following first extensive myocardial infarction in man. AB - We investigated the time course of genesis of skeletal muscle dysfunction and sympatho-vagal imbalance after myocardial infarction. We studied 22 normal controls, 22 patients with >6 months stable chronic heart failure and 10 patients after a first massive myocardial infarction at 1-3 weeks (the "early" period), 6 8 weeks ("mid") and 6-9 months ("late") following their infarct. Four patients developed overt heart failure. Forearm muscle metabolism was studied using (31)P magnetic resonance spectroscopy (MRS). Sympatho-vagal balance was assessed by heart rate variability and radiolabelled norepinephrine kinetics. Increased norepinephrine spillover (0.55+/-0.02 v 0.27+/-0.04 mg/min/m(2); P<0.01) and decreased heart rate variability were confined to those post-myocardial infarction patients who subsequently developed heart failure. Resting cardiac output was normal in all the post-myocardial infarction patients, although the response of cardiac output to supine bicycle exercise at the "mid" study point was less in the group who subsequently developed heart failure (9+/-1 v 41+/-8 %; P<0.005). In the MRS studies, there were no detectable differences between those who did or did not develop heart failure. The initial rate of ATP turnover, calculated from initial-exercise changes in pH and phosphocreatine (PCr), was increased in established chronic heart failure, but in the post-myocardial infarction patients a numerically similar increase reached statistical significance only in the early group (19+/-3 v 11+/-1 mM/min; P<0.005). The apparent maximum rate of oxidative ATP synthesis, calculated from post-exercise PCr recovery kinetics, was lower than control in the late post-myocardial infarction and established chronic heart failure groups 34+/-5 v 55+/-4 mM/min; P<0.03 and 38+/-3 v 55+/-4 mM/min; P<0.003, respectively). Skeletal muscle metabolism and autonomic function become abnormal after an extensive myocardial infarction. While skeletal muscle abnormalities are relatively slow to develop and unrelated to the degree of failure, excessive neurohormonal activation and impaired cardiac output response to exercise seem from an early stage to characterize patients who subsequently develop chronic heart failure. PMID- 10525429 TI - Differential expression of natriuretic peptides and their receptors in volume overload cardiac hypertrophy in the rat. AB - Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) upregulation are genetic markers for the chronic hypertrophic phenotype but also have important acute physiologic effects on salt and water balance and blood pressure control. The presence of a dual NP-system led us to hypothesize a differential expression of ANP and BNP in response to an acute hemodynamic stress of volume overload in the left ventricle (LV) and right ventricle (RV). Accordingly, we examined the temporal relationship between the RV and LV expression of ANP and BNP mRNA and NP receptor mRNA levels on days 1, 2, 3, and 7 after induction of aortocaval fistula in the rat. LV end-diastolic pressure was increased 1.5-fold by day 3 and 2.0-fold by day 7 compared to control (P<0.05). LV weight increased by day 7 compared to control (2.34+/-0.04 vs 3.07+/-0.10 mg/g, P<0.05) while RV weight did not change over the 7 days. There was a 7-fold increase of ANP mRNA in LV at day 1, which was sustained through day 7, while LV BNP mRNA levels did not differ from controls over the 7 days. In contrast, RV mRNA transcript levels for ANP and BNP were increased >2-fold by day 2 and this increase was sustained throughout 7 days. NP clearance receptor was decreased by 75% by day 7 in the LV but did not change in the RV. Thus, LV ANP mRNA levels increased before the onset of LV hypertrophy and RV BNP mRNA levels increased in the absence of RV hypertrophy. The disparate response of BNP and the NP clearance receptor transcript levels in the LV and RV may be related to differences in load and/or differential expression of the NP system in the LV and RV in response to acute haemodynamic stress. PMID- 10525431 TI - Editorial PMID- 10525430 TI - Protein kinase C-epsilon is responsible for the protection of preconditioning in rabbit cardiomyocytes. AB - The role of protein kinase C (PKC) in the protection of ischemic preconditioning (PC) is still controversial, partly because of the multiple isozymes of PKC and the inability to directly measure PKC activity in vivo. In this study we have used novel peptide inhibitors which correspond to part of the amino acid sequence from the isozyme-specific RACK-binding site on the PKC molecule. The peptides prevent binding of a specific activated PKC isozyme to its RACK, thus halting isozyme translocation and function. The inhibitor peptides are cross-linked to the membrane-translocating antennapedia homeodomain peptide that allows their entry into cells. The effect of inhibitors of PKC-beta, -delta, -epsilon and -eta were evaluated. Rabbit adult ventricular myocytes were obtained by enzymatic dissociation. Ischemia was simulated by centrifuging the myocytes into an oxygen free pellet for 180 min. PC was induced by 10 min of pelleting followed by resuspension in oxygenated medium for 15 min. During simulated ischemia cells undergo a predictable increase in osmotic fragility as judged by determination of the number of stained cells following their incubation in hypotonic (85 mOsm) trypan blue. The percentage of cells experiencing membrane rupture, and thus cell staining, was considered to be an index of ischemic injury. PC significantly delayed the progression of osmotic fragility during simulated ischemia (P<0.01). The protection of PC was abolished by the peptide inhibitor of PKC-epsilon but not by the peptide inhibitors selective for PKC-beta, PKC-delta, or PKC-eta; each was applied at 100 n N. Protection could also be induced by the PKC activator oleoylacetyl glycerol, and that protection was aborted by the inhibitor selective for PKC-epsilon, but not by the inhibitor for PKC-delta. None of the above peptide treatments affected the osmotic fragility in non-PC cells during simulated ischemia. Our studies further support PKC as a critical part of the signal transduction pathway in PC and indicate that PKC-epsilon alone is responsible for the early phase of PC's protection in rabbit cardiomyocytes. PMID- 10525432 TI - Assessment of the functional integrity of the humoral immune response: the plaque forming cell assay and the enzyme-linked immunosorbent assay. AB - The plaque-forming cell (PFC) assay and enzyme-linked immunosorbent assay (ELISA) appear to have comparable sensitivity and reproducibility for measuring IgM antibody production in mice and rats immunized with sheep red blood cells (sRBCs). Both assays can be manipulated, with respect to the immunizing antigen (e.g., T-dependent vs T-independent antigen), to provide evidence as to which cell type(s) may be adversely affected by a given compound. However, the PFC assay has more utility in dissecting out the target cell(s) involved. Since both the PFC assay and the ELISA may be readily conducted in the rat, it is feasible to incorporate either of these assays into standard acute and repeat dose toxicology studies. This may be accomplished by inclusion of satellite groups in the study. However, it has been suggested that the primary antibody response to sRBCs, as measured by an ELISA, may also be evaluated in the main group of animals in a toxicology study without compromise to the integrity of other toxicological endpoints (e.g., hematology, clinical chemistry, histopathology). Both approaches will provide a more extensive delineation of the safety profile of a drug or chemical. The latter approach will also reduce the number of animals needed and the cost of the study. PMID- 10525433 TI - Effects of xenobiotics on macrophage function: evaluation in vitro. AB - Macrophages participate in a variety of inflammatory and immunologic functions (e.g., phagocytosis, cytokine production, killing of microbes and tumor cells, and processing and presentation of antigen to T lymphocytes). Because these cells are widely distributed in the circulation and throughout tissue, the effects of xenobiotics on macrophage function can be significant. Measures in vitro of altered function elicited by xenobiotic exposure can include changes in expression of cell surface proteins, in production of oxygen and nitrogen free radicals, in production of cytokines [interleukin (IL)-1, IL-6, IL-12, tumor necrosis factor alpha (TNFalpha)], in expression of adhesion molecules (ICAM-1), in phagocytosis and intracellular killing, and in antigen presentation. PMID- 10525434 TI - Theory and practice of cytokine assessment in immunotoxicology. AB - The tools and concepts of immunotoxicology are increasingly being used in novel ways, such as using toxic reagents to understand immune system function. One of the most potentially useful of these new tools is the assessment of cytokines, the molecules responsible for regulating a variety of processes including immunity, inflammation, apoptosis, and hematopoiesis. Cytokine production or bioactivity may be affected by a variety of toxic mechanisms including direct toxicity to cytokine-producing cells, inhibition of cytokine production, inhibition of cytokine release, induction of immunosuppressive factors, alterations in cellular homeostasis, alterations in cellular activational or transcriptional mechanisms, and miscellaneous or undefined mechanisms. Moreover, alterations in the profile of cytokine production may provide important information regarding the nature of an immunotoxic insult (i.e., TH1 vs TH2 response). Proper evaluation of the role of cytokine modulation in immunotoxicology requires attention to myriad details. Some of the details discussed in this review include the source of the sample to be tested (circulating, local, or ex vivo isolated cells); the potential effects of collection, processing, and storage on the results of the assays; potential variables associated with the source material (matrix effects, relevance, inhibitory substances); and factors influencing the choice of assay used (bioassay, immunoassay, molecular biology technique, flow cytometry, hybrid assays). Other often-overlooked issues are discussed, including species considerations and quality control issues such as the use of reference standards and the expression of results. PMID- 10525435 TI - Uses and future applications of flow cytometry in immunotoxicity testing. AB - Flow cytometry is an emerging technology that has numerous applications to immunotoxicity testing. The use and development of high-speed single-cell laser based assays capable of quantitation of fluorescence, light scatter, and electrical impedance measurements can provide important information on xenobiotic induced toxicity in defined target cell populations. The purpose of this article is to briefly review established and emerging immunotoxicology assays that use flow cytometry. In the coming years it is likely that many new flow cytometry assays will be developed and validated that will improve the sensitivity and perhaps specificity of immunotoxicity testing. Since flow cytometry is readily adaptable to high-throughput screening, it is also likely that this technology will increasingly find its place in the preclinical testing of drugs and chemicals in the pharmaceutical and chemical industries. PMID- 10525436 TI - Assessment of apoptosis in xenobiotic-induced immunotoxicity. AB - Generation of immunity is a highly complex process in which proliferation and differentiation of immune-competent cells regulated by cytokines and cell-cell interactions play a major role. Reducing the number of immune-competent cells or altering the function, selection, and differentiation of lymphocytes after xenobiotic treatment may lead to serious adverse effects. Programmed cell death, or apoptosis, is a highly regulated process by which an organism eliminates unwanted cells without eliciting an inflammatory response. However, xenobiotics are also able to trigger unwanted apoptosis or to alter the regulation of programmed cell death. Cytological characteristics of apoptosis are generally different from those seen in acute pathological cell death resulting from cell injury. The morphological characteristics of apoptosis are unique including cell shrinkage, membrane blebbing, chromatin condensation, DNA fragmentation, disruption of the nuclear lamina, nuclear fragmentation, and emergence of apoptotic bodies. It is now established that apoptosis plays a critical role in both development and homeostasis of the immune system: thymic selection, cytotoxicity, deletion of autoreactive cells, and regulation of the size of the lymphoid compartment. Assessment of apoptosis relies on the morphological and biochemical modifications of the dying cells. As a rule, and because an apoptotic cell rarely displays all of the characteristic apoptotic features, several criteria should be monitored in parallel including morphological examination. The techniques described in this paper have been divided into five categories: analysis of cell morphology by microscopy, identification of DNA fragmentation, determination of mitochondrial membrane potential, detection of plasma membrane changes, analysis of caspase activation. PMID- 10525437 TI - Local lymph node assay: differentiating allergic and irritant responses using flow cytometry. AB - The murine local lymph node assay (LLNA) is a method for assessing the contact sensitization potential of chemicals. Based on events that occur during the induction phase of a contact sensitization response, the LLNA measures the in vivo proliferation of cells in the draining lymph nodes (DLNs) of mice following topical exposure to chemicals. In terms of predictive identification of important skin sensitizers, the LLNA has been shown to be at least as sensitive as, and much more reliable than, current guinea pig tests. However, proliferation has also been observed following treatment with some irritants. In an attempt to distinguish allergic from irritant-induced proliferation, flow cytometric techniques have been used to examine the phenotype of lymphocyte subsets in the DLNs as well as markers of T-lymphocyte activation and memory. Mice were treated on the ears for 3 consecutive days with allergens or irritants. The DLNs were harvested 72 h after the final treatment. Single-cell suspensions were prepared, counted, and stained for analysis of the percentages of T cells and B cells and T cell expression of two adhesion molecules that have been associated with differentiating naive and activated/memory T cells, CD62L (L-selectin) and CD44 (H-cam). Increases in lymph node cellularity were observed in both allergen- and irritant-treated mice relative to naive and vehicle-treated animals. Mice treated with allergens showed a preferential increase in the percentage of B220(+) B cells compared with irritant-treated mice. Treatment with allergens, but not irritants, resulted in a selective increase in the percentages of CD4(+) and CD8(+) cells expressing the T-cell activation/memory phenotype CD62L(lo)CD44(hi). Taken together, flow cytometric analysis of cell phenotype and expression of T cell activation/memory markers may provide important information for differentiating allergen- and irritant-induced proliferative responses in the DLNs of chemically treated mice. PMID- 10525438 TI - Cytokine fingerprinting: characterization of chemical allergens. AB - Chemical allergy is a common and important occupational health issue. Allergic sensitization induced by chemicals may take a variety of forms, including allergic contact dermatitis (skin sensitization) and allergic asthma and rhinitis (sensitization of the respiratory tract). There is a need to identify and characterize chemicals that have the potential to cause such sensitization reactions. Although a number of methods are available for the prospective analysis of skin sensitizing activity, there are currently no widely accepted tests for the identification of chemical respiratory allergens. We here describe a novel approach, cytokine fingerprinting, that has the potential to distinguish between chemical contact and respiratory allergens. The pattern of cytokine production by draining lymph node cells (LNCs) is evaluated following repeated topical exposure of mice to test chemicals. Experience to date reveals that contact allergens stimulate the selective development of type 1 immune responses associated with the secretion by draining LNCs of interferon gamma (IFN-gamma), but little interleukin-4 (IL-4) or interleukin-10 (IL-10). In contrast, chemical respiratory allergens are found to induce the appearance of preferential type 2 immune responses characterized by IL-4 and IL-10 production, but comparatively low levels of IFN-gamma. It is proposed that cytokine fingerprinting may permit the simultaneous identification and characterization of those chemicals that have the potential to cause allergic sensitization. PMID- 10525439 TI - Quantitation of cytokine mRNA expression as an endpoint for prediction and diagnosis of xenobiotic-induced hypersensitivity reactions. AB - Xenobiotic-induced hypersensitivity reactions are immune-mediated effects that involve specific antibodies and/or effector and regulatory T lymphocytes. Cytokines are key mediators of such responses and must be considered as possible endpoints for predicting sensitizing potency of drugs and chemicals, as well as for helping diagnosis of allergy. Detecting cytokine production at the protein level has been shown to not be always sensitive enough. This paper describes three examples of the utilization of semiquantitative or competitive reverse transcription polymerase chain reaction analysis of interleukin-4, interferon gamma, and interleukin-1beta mRNAs as endpoints for assessing T-cell or dendritic cell responses to sensitizing drugs (beta-lactam antibiotics) or chemicals (dinitrochlorobenzene). PMID- 10525440 TI - Assessment of autoimmunogenic potential of xenobiotics using the popliteal lymph node assay. AB - This article reviews the ability of the simple popliteal lymph node assay (PLNA) and variations thereof to assess the immunostimulating potential of low-molecular weight xenobiotics, including pharmaceuticals. In essence, all variations of the PLNA detect the immune reaction in the popliteal lymph node to subcutaneous injection of a chemical into the footpad. The primary PLNA, in which the enlargement of popliteal lymph node is measured on injection of the chemical as such, can be regarded as a fast, simple, and reliable assay to detect and grade the immunostimulating potential of chemicals in a preclinical production phase. To prove T-cell sensitization, i.e., the involvement of T cells and/or induction of T-cell memory, secondary PLNAs or the so-called modified PLNA can be used. Secondary PLNAs can be performed in previously sensitized animals or by using adoptive T-cell transfer techniques. In the modified PLNA the well-defined reporter antigens TNP-ovalbumin and TNP-Ficoll are injected together with the chemicals and the number and isotype of the antibody-forming cells in the draining lymph node are analyzed. This modification of the PLNA enables definition of the involvement of T cells as well as type of immune response (T cell sensitization vs mere inflammation as well as Th1 vs Th2) elicited by the chemical in an easy manner. To date, more than 100 chemicals have been tested in the PLNA and results indicate that all chemicals with documented adverse autoimmune or allergic effects in humans induce a positive PLN response. No false negatives have been found if metabolism is taken into consideration. It is important to realize that immunostimulation measured in the PLNA is only a first indication that a chemical can induce or exacerbate autoimmune(-like) disease. PMID- 10525441 TI - An oral sensitization model in Brown Norway rats to screen for potential allergenicity of food proteins. AB - We developed an oral sensitization protocol for food proteins for the rat. Young Brown Norway (BN) rats were exposed to 1 mg ovalbumin (OVA) by daily gavage dosing for 42 days without the use of an adjuvant. OVA-specific IgE and IgG responses were determined by ELISA. On an oral challenge with OVA some clinical symptoms of food allergy-like effects on the respiratory system, blood pressure, and permeability of the gastrointestinal barrier were studied. In addition, BN rats were orally exposed to a total hen egg white protein (HEW) extract and cow's milk (CM) and the specificities of induced antibody responses were compared with the specificities of antibodies in sera from egg- and milk-allergic patients using immunoblotting. Animals orally exposed to the allergens developed specific IgE and IgG antibodies which recognized the same proteins compared with antibodies from egg- or CM-allergic patients. Among the various clinical symptoms of food allergy, gut permeability was increased after an oral challenge. In addition, some animals demonstrated a temporary decrease in breathing frequency or systolic blood pressure. The results obtained show that the Brown Norway rat is a suitable animal model for inducing specific IgG and IgE responses on daily intragastric dosing of OVA without the use of an adjuvant. Moreover, local immune mediated effects on oral challenge are observed. The observation that enterally exposed BN rats and food-allergic patients demonstrate antibody responses to a comparable selection of proteins on exposure to different protein mixtures (HEW and CM) further supports the suitability of the BN rat as an animal model for food allergy research and for the study of the allergenicity of (novel) food proteins. PMID- 10525442 TI - Editorial PMID- 10525443 TI - Squalene and squalane emulsions as adjuvants. AB - Microfluidized squalene or squalane emulsions are efficient adjuvants, eliciting both humoral and cellular immune responses. Microfluidization stabilizes the emulsions and allows sterilization by terminal filtration. The emulsions are stable for years at ambient temperature and can be frozen. Antigens are added after emulsification so that conformational epitopes are not lost by denaturation and to facilitate manufacture. A Pluronic block copolymer can be added to the squalane or squalene emulsion. Soluble antigens administered in such emulsions generate cytotoxic T lymphocytes able to lyse target cells expressing the antigen in a genetically restricted fashion. Optionally a relatively nontoxic analog of muramyl dipeptide (MDP) or another immunomodulator can be added; however, the dose of MDP must be restricted to avoid systemic side effects in humans. Squalene or squalane emulsions without copolymers or MDP have very little toxicity and elicit potent antibody responses to several antigens in nonhuman primates. They could be used to improve a wide range of vaccines. Squalene or squalane emulsions have been administered in human cancer vaccines, with mild side effects and evidence of efficacy, in terms of both immune responses and antitumor activity. PMID- 10525444 TI - Immunomodulation by iscoms, immune stimulating complexes. AB - The iscom is a uniform stable complex consisting of cholesterol, phospholipid, adjuvant-active saponin, and antigen. The iscom matrix is a particulate complex with identical composition, shape, and morphology, but lacking the incorporated antigen. The assembly of the complex is based on hydrophobic interactions, but antigens that are not hydrophobic can be conjugated with a hydrophobic tail or hidden hydrophobic regions can be exposed, e.g., by acid treatment, to facilitate the incorporation into iscoms. The functional aspects of iscoms are described emphasizing immunomodulation in mouse models. Iscoms prominently enhance the antigen targeting, uptake, and activity of antigen presenting cells including dendritic and B cells and macrophages resulting in the production of proinflammatory cytokines, above all interleukin (IL)-1, IL-6, and IL-12. The expression of costimulatory molecules major histocompatibility complex (MHC) class II, B7.1 and B7.2, is also enhanced. The latter partly explains why the iscom is an efficient adjuvant for elderly mice. Iscoms enhance the Th1 type of response with increased production of IL-2 and interferon gamma. However, with some antigens and particularly in monkeys immunized with HIV iscoms, the production of IL-4 was enhanced. IL-4, IL-2, and interferon gamma (IFNgamma) together with the beta chemokines MIP-1alpha and MIP-1beta correlated with protection against challenge infection with a chimeric virus (simian immunodeficiency virus-human immunodeficiency virus). Iscoms were also shown to induce a potent immune response in the newborn and to be an efficient delivery system for mucosal administration. Technical information is given about formulation of iscoms and about handling of antigens to optimize their incorporation into iscoms. PMID- 10525445 TI - Monophosphoryl lipid A (MPL) formulations for the next generation of vaccines. AB - Many of the latest trends in vaccine development are dependent on immunological adjuvants that mediate and promote a wide variety of immune responses. One promising adjuvant candidate, monophosphoryl lipid A (MPL) immunostimulant, is being investigated with many of these new vaccine approaches in either preclinical or clinical trials. This is possible because different vehicle formulations can significantly influence the type of immunological response MPL promotes. Procedures are provided for formulating MPL in an aqueous vehicle or an oil-in-water emulsion. These two MPL formulations can be beneficial for most vaccine approaches being investigated today. PMID- 10525446 TI - Interleukin-1 and interleukin-1 fragments as vaccine adjuvants. AB - The human interleukin-1beta (IL-1beta) domain in position 163-171, comprising the amino acids VQGEESNDK, has been synthesized as a nine-amino-acid-long peptide and used in vivo as a nontoxic HCl salt. The IL-1beta nonapeptide reproduces the immunostimulatory and adjuvant effects of the whole mature IL-1beta, but does not possess any of the IL-1beta inflammatory, vasoactive, tumor-promoting, and systemically toxic effects, nor it can synergize with tumor necrosis factor alpha or other molecules in inducing toxicity and shock. The IL-1beta fragment is active as adjuvant either when administered together with the antigen or if inoculated separately; it can be physically linked to the antigen or used as a discrete peptide. Moreover, the DNA sequence encoding the IL-1beta domain has been included in an experimental DNA vaccine with positive results. Thus, immunostimulatory sequences can be identified within a pleiotropic cytokine like IL-1 and used in the rational design of novel vaccination strategies. PMID- 10525447 TI - Interleukin-12 as an adjuvant for cancer immunotherapy. AB - Interleukin-12 (IL-12) is a cytokine whose main effect is to drive Th-cell differentiation throughout a T helper type 1 cell type of response, thus inducing interferon gamma (IFNgamma) and favoring a switch from Ig to IgG2a. These properties make IL-12 a candidate adjuvant for vaccination against cancer and infection disease. Enthusiasm was generated in many animal studies where IL-12 was given either systemically or locally. The experience of some toxicity in humans has hampered its further development into clinical applications, which, however, are still possible if restricted to local administration. Gene transfer seems to be the preferred approach to obtain this local release of cytokine. Here we review the applications of IL-12 as adjuvant. PMID- 10525448 TI - Interleukin-18. AB - Interleukin (IL)-18 is a newly discovered cytokine, structurally similar to IL-1, with profound effects on T-cell activation. This short review summarizes the present knowledge on IL-18, to give an insight into the future perspectives for its possible use as vaccine adjuvant. Formerly called interferon (IFN) gamma inducing factor (IGIF), IL-18 is the new name of a novel cytokine that plays an important role in the T-cell-helper type 1 (Th1) response, primarily by its ability to induce IFNgamma production in T cells and natural killer (NK) cells. Mice deficient in IL-18 have suppressed IFNgamma production despite the presence of IL-12 IL-18 is related to the IL-1 family in terms of structure, receptor family, and function. In terms of structure, IL-18 and IL-1beta share primary amino acid sequences of the so-called "signature sequence" motif and are similarly folded as all-beta pleated sheet molecules. Also similar to IL-1beta, IL-18 is synthesized as a biologically inactive precursor molecule lacking a signal peptide which requires cleavage into an active, mature molecule by the intracellular cysteine protease called IL-1beta-converting enzyme (ICE, also called caspase-1). The activity of mature IL-18 is closely related to that of IL 1. IL-18 induces gene expression and synthesis of tumor necrosis factor (TNF), IL 1, Fas ligand, and several chemokines. The activity of IL-18 is via an IL-18 receptor (IL-18R) complex. This IL-18R complex is made up of a binding chain termed IL-18Ralpha, a member of the IL-1 receptor family previously identified as the IL-1 receptor-related protein (IL-1Rrp), and a signaling chain, also a member of the IL-1R family. The IL-18R complex recruits the IL-1R-activating kinase (IRAK) and TNFR-associated factor-6 (TRAF-6) which phosphorylates nuclear factor kappaB (NFkappaB)-inducing kinase (NIK) with subsequent activation of NFkappaB. Thus on the basis of primary structure, three-dimensional structure, receptor family, signal transduction pathways and biological effects, IL-18 appears to be a new member of the IL-1 family. Similar to IL-1, IL-18 participates in both innate and acquired immunity. PMID- 10525449 TI - Peptide carriers: A helicoid-type sequential oligopeptide carrier (SOC(n)) for multiple anchoring of antigenic/immunogenic peptides. AB - A new peptide carrier with three-dimensional predetermined structural motif has been constructed by the repetitive Lys-Aib-Gly moiety. The sequential oligopeptide carrier (SOC(n)), (Lys-Aib-Gly)(n), adopts a distorted 3(10)-helical conformation and the Lys-N(epsilon)H(2) anchoring groups exhibit defined spatial orientations. Conformational analysis of the SOC(n) conjugates showed that the coupled peptides retain their initial "active" structure, while prevalence of one conformer was also observed. It is concluded that the beneficial structural elements of SOC(n) induce a favorable arrangement of the conjugated peptides, so that potent antigens and immunogens are generated. PMID- 10525450 TI - Dendritic cells as natural adjuvants. AB - Dendritic cells (DCs) are professional antigen presenting cells that hold the key to the induction of T-cell responses. Therefore, the use of DCs for immunotherapy to stimulate immune responses has recently raised a great deal of interest. Many clinical trials using DCs have been initiated to stimulate immune responses against tumors or infectious agents. Several issues need to be considered before DCs can be used successfully as natural adjuvants: DCs have to be generated in sufficient numbers; they should display morphological, phenotypical, and functional properties of DCs; and they should be able to present antigens. In the present review we focus on methods for the purification of DCs from human bone marrow and peripheral blood and for the optimization of in vitro cell culture systems. Methods to generate growth factor-dependent mouse DC lines are also described. PMID- 10525451 TI - Mucosal delivery of vaccines. AB - Oral delivery represents one of the most pursued approaches for large-scale human vaccination. Due to the different characteristics of mucosal immune response, as compared with systemic response, oral immunization requires particular methods of antigen preparation and selective strategies of adjuvanticity. In this paper, we describe the preparation and use of genetically detoxified bacterial toxins as mucosal adjuvants and envisage the possibility of their future exploitation for human oral vaccines. PMID- 10525452 TI - Vaccine entrapment in liposomes. AB - The use of liposomes as carriers of peptide, protein, and DNA vaccines requires simple, easy-to-scale-up technology capable of high-yield vaccine entrapment. Work from this laboratory has led to the development of techniques that can generate liposomes of various sizes, containing soluble antigens such as proteins and particulate antigens (e.g., killed or attenuated bacteria or viruses), as well as antigen-encoding DNA vaccines. Entrapment of vaccines is carried out by the dehydration-rehydration procedure which entails freeze-drying of a mixture of "empty" small unilamellar vesicles and free vaccines. On rehydration, the large multilamellar vesicles formed incorporate up to 90% or more of the vaccine used. When such liposomes are microfluidized in the presence of nonentrapped material, their size is reduced to about 100 nm in diameter, with much of the originally entrapped vaccine still associated with the vesicles. A similar technique applied for the entrapment of particulate antigens (e.g., Bacillus subtilis spores) consists of freeze-drying giant vesicles (4-5 microm in diameter) in the presence of spores. On rehydration and sucrose gradient fractionation of the suspension, up to 30% or more of the spores used are associated with generated giant liposomes of similar mean size. PMID- 10525453 TI - Engineering the gram-positive cell surface for construction of bacterial vaccine vectors. AB - A genetic system for surface display of heterologous proteins has been developed in Streptococcus gordonii, a gram-positive human oral commensal that is naturally competent for genetic transformation. Our approach is based on chromosomal integration downstream from a resident promoter and translational fusion to an M6 protein. Using this strategy a variety of proteins, of different origin and size, were displayed on the cell surface and were shown to be stably expressed both in vitro and in vivo. Animal models of mucosal colonization (oral and vaginal) and intragastric immunization with recombinant S. gordonii were developed and the local and systemic immune responses were studied. Here we report the techniques for the construction of recombinant bacteria, use of animal models, and analysis of the immune response. PMID- 10525454 TI - Parvovirus-like particles as vaccine vectors. AB - A wide array of systems have been developed to improve "classic" vaccines. The use of small polypeptides able to elicit potent antibody and cytotoxic responses seems to have enormous potential in the design of safer vaccines. While peptide coupling to large soluble proteins such as keyhole limpet hemocyanin is the current method of choice for eliciting antibody responses and insertion in live viruses for cytotoxic T-lymphocyte responses, alternative cheaper and/or safer methods will clearly be required in the future. Virus-like particles constitute very immunogenic molecules that allow for covalent coupling of the epitopes of interest in a simple way. In this article, we detail the methodology employed for the preparation of efficient virus vectors as delivery systems. We used parvovirus as the model for the design of new vaccine vectors. Recently parvovirus-like particles have been engineered to express foreign polypeptides in certain positions, resulting in the production of large quantities of highly immunogenic peptides, and to induce strong antibody, helper-T-cell, and cytotoxic T-lymphocyte responses. We discuss the different alternatives and the necessary steps to carry out this process, placing special emphasis on the flow of decisions that need to be made during the project. PMID- 10525455 TI - DNA-based vaccination against tumors expressing the P1A antigen. AB - Based on experience acquired in the last few years, we describe some technical steps and provide suggestions on how to induce an immune response against tumors expressing the weakly immunogenic antigen P1A by means of a DNA-based vaccination approach. P1A is the product of a normal mouse gene, which shares many characteristics with already identified human tumor-associated antigens, and therefore represents a useful experimental model to evaluate the efficacy of new vaccination strategies potentially applicable to the field of human tumors. Information gained with this model has been applied with success in other experimental settings, and thus we think that the procedure described herein may constitute a valid platform that can be implemented and further refined. PMID- 10525456 TI - X-Rays from the Highly Polarized Broad Absorption Line QSO CSO 755. AB - We present results from a BeppoSAX observation of the broad absorption line (BAL) QSO CSO 755, which was observed as part of our program to investigate the X-ray properties of highly polarized BAL QSOs. CSO 755 is clearly detected by the BeppoSAX Medium-Energy Concentrator Spectrometers, making it the highest redshift (z=2.88) and most optically luminous (MV=-27.4) BAL QSO seen in X-rays. It is detected in several energy bands including the rest-frame 21-39 keV band, but we are able to place only loose constraints upon its X-ray spectral shape. Our X-ray detection is consistent with the hypothesis that the BAL QSOs with high optical continuum polarization tend to be the X-ray brighter members of the class. We examine a scattering interpretation of a polarization/X-ray flux connection, and we discuss the data needed to prove or refute such a connection. We also discuss a probable ROSAT detection of CSO 755. The observed-frame 2-10 keV flux from BeppoSAX (1.3x10-13 ergs cm-2 s-1) is high enough to allow X-Ray Multimirror Mission spectroscopy, and studies of iron K-line emission should prove of particular interest if a large amount of scattered X-ray flux is present. PMID- 10525458 TI - On the Cusp around Central Black Holes in Luminous Elliptical Galaxies. AB - In this Letter, we show that a massive black hole (MBH) that falls into the center of a galaxy on the dynamical timescale leaves a weak cusp (rho~r 1&solm0;2) around it, which is in good agreement with the recent observations of luminous elliptical galaxies by the Hubble Space Telescope. Such an event is a natural outcome of the merging of two galaxies that have central MBHs. This is the only known mechanism for forming weak cusps in luminous elliptical galaxies. Therefore, the existence of the weak cusps indicates that the central black holes of luminous elliptical galaxies have fallen to the center from outside, most likely during a major merger event. PMID- 10525457 TI - The BeppoSAX View of the Hot Cluster Abell 2319. AB - We present results from a BeppoSAX observation of the rich cluster Abell 2319. The broadband spectrum (2-50 keV) of the cluster can be adequately represented by an optically thin thermal emission model with a temperature of 9.6+/-0.3 keV and a metal abundance of 0.25+/-0.03 in solar units and with no evidence of a hard X ray excess in the PDS spectrum. From the upper limit to the hard-tail component, we derive a lower limit of approximately 0.04 uG for the volume-averaged intracluster magnetic field. By performing spatially resolved spectroscopy in the medium energy band (2-10 keV), we find that the projected radial temperature and metal abundance profiles are constant out to a radius of 16&arcmin; (1.4 Mpc). A reduction of the temperature of one-third, when going from the cluster core out to 16&arcmin;, can be excluded in the present data at the 99% confidence level. From the analysis of the temperature and abundance maps, we find evidence of a temperature enhancement and of an abundance decrement in a region localized 6&arcmin;-8&arcmin; northeast of the core, where a merger event may be taking place. Finally, the temperature map indicates that the subcluster located northwest of the main cluster may be somewhat cooler than the rest of the cluster. PMID- 10525459 TI - The Radio Afterglow and the Host Galaxy of the X-Ray-rich GRB 981226. AB - We report the discovery of a radio transient VLA 232937.2-235553, coincident with the proposed X-ray afterglow for the gamma-ray burst GRB 981226. This gamma-ray burst (GRB) has the highest ratio of X-ray to gamma-ray fluence of all the GRBs detected by BeppoSAX so far, and yet no corresponding optical transient was detected. The radio light curve of VLA 232937.2-235553 is qualitatively similar to that of several other radio afterglows. At the subarcsecond position provided by the radio detection, optical imaging reveals an extended R=24.9 mag object, which we identify as the host galaxy of GRB 981226. Afterglow models that invoke a jetlike geometry for the outflow or that require an ambient medium with a radial density dependence, such as that produced by a wind from a massive star, are both consistent with the radio data. Furthermore, we show that the observed properties of the radio afterglow can explain the absence of an optical transient without the need for large extinction local to the GRB. PMID- 10525460 TI - Near-Infrared Spectra of Ultraluminous Infrared Galaxies. AB - Near-infrared spectra with a resolution of lambda&solm0;Deltalambda approximately 1100 in the rest-wavelength range of 1.8-2.2 um have been obtained for a complete sample of 33 ultraluminous infrared galaxies (ULIRGs). Of the 33 objects observed, two show evidence of a central active galactic nucleus (AGN) through either a broad Paschen-alpha line or emission in the 1.963 um fine structure of [Si vi]. In the median spectrum of the remaining 31 objects, the lines present are recombination lines of hydrogen and neutral helium, vibration-rotation lines of H2, and [Fe ii]. There is no indication of AGN activity in the median spectrum, either through broad atomic recombination lines or through high ionization lines. No trends in luminosity are apparent when subsets of the 31 non AGN ULIRGs are binned by luminosity and median-combined. When secondary nuclei exist in ULIRGs, they typically have spectra very much like those seen in the primary nuclei. PMID- 10525461 TI - Possible Evidence for Truncated Thin Disks in the Low-Luminosity Active Galactic Nuclei M81 and NGC 4579. AB - M81 and NGC 4579 are two of the few low-luminosity active galactic nuclei that have an estimated mass for the central black hole, detected hard X-ray emission, and detected optical/UV emission. In contrast to the canonical "big blue bump," both have optical/UV spectra that decrease with increasing frequency in a nuLnu plot. Barring significant reddening by dust and/or large errors in the black hole mass estimates, the optical/UV spectra of these systems require that the inner edge of a geometrically thin, optically thick accretion disk lies at approximately 100 Schwarzschild radii. The observed X-ray radiation can be explained by an optically thin, two-temperature, advection-dominated accretion flow at smaller radii. PMID- 10525463 TI - Pinwheel Nebula around WR 98a. AB - We present the first near-infrared images of the dusty Wolf-Rayet star WR 98a. Aperture-masking interferometry has been utilized to recover images at the diffraction limit of the Keck I telescope, less, similar50 mas at 2.2 um. Multiepoch observations spanning about 1 yr have resolved the dust shell into a "pinwheel" nebula, the second example of a new class of dust shell first discovered around WR 104 by Tuthill, Monnier, & Danchi. Interpreting the collimated dust outflow in terms of an interacting winds model, the binary orbital parameters and apparent wind speed are derived: a period of 565+/-50 days, a viewing angle of 35&j0;+/-6 degrees from the pole, and a wind speed of 99+/-23 mas yr-1. This period is consistent with a possible approximately 588 day periodicity in the infrared light curve, linking the photometric variation to the binary orbit. Important implications for binary stellar evolution are discussed by identifying WR 104 and WR 98a as members of a class of massive, short-period binaries whose orbits were circularized during a previous red supergiant phase. The current component separation in each system is similar to the diameter of a red supergiant, which indicates that the supergiant phase was likely terminated by Roche lobe overflow, leading to the present Wolf-Rayet stage. PMID- 10525462 TI - Observation of Multi-TeV Gamma Rays from the Crab Nebula using the Tibet Air Shower Array. AB - The Tibet experiment, operating at Yangbajing (4300 m above sea level), is the lowest energy air shower array, and the new high-density array constructed in 1996 is sensitive to gamma-ray air showers at energies as low as 3 TeV. With this new array, the Crab Nebula was observed in multi-TeV gamma-rays and a signal was detected at the 5.5 sigma level. We also obtained the energy spectrum of gamma rays in the energy region above 3 TeV which partially overlaps those observed with imaging atmospheric Cerenkov telescopes. The Crab spectrum observed in this energy region can be represented by the power-law fit dJ&parl0;E&parr0;&solm0;dE=&parl0;4.61+/-0.90&parr0;x10-12&parl0;E&solm0;3 TeV&parr0;-2.62+/-0.17 cm-2 s-1 TeV-1. This is the first observation of gamma-ray signals from point sources with a conventional air shower array using scintillation detectors. PMID- 10525464 TI - Enhanced OH in C-Type Shock Waves in Molecular Clouds. AB - Cosmic-ray and X-ray ionizations in molecular gas produce a weak far-ultraviolet flux through the radiative decay of H2 molecules that have been excited by collisions with energetic electrons (the Prasad-Tarafdar mechanism). I consider the effect of this dissociating flux on the oxygen chemistry in C-type shocks. Typically, a few percent of the water molecules produced within the shock front are dissociated before the gas has cooled to 50 K. The resulting column density of warm OH rises from 1015 to 1016 cm-2 as the ionization rate is increased from 10-17 s-1 (typical of dark clouds) to 10-15 s-1 (adjacent to supernova remnants). These column densities produce substantial emission in the far-infrared rotational transitions of OH and are consistent with the OH/H2O ratios inferred from Infrared Space Observatory observations of emission from molecular shocks. For high ionization rates, the column of warm OH is sufficient to explain the OH(1720 MHz) masers that occur where molecular clouds are being shocked by supernova remnants. The predicted abundance of OH throughout the shock front will enable C-type shocks to be examined with high spectral resolution through radio observations of the four hyperfine ground-state transitions of OH at 18 cm and heterodyne measurements of emission in the far-infrared (e.g., from the Stratospheric Observatory for Infrared Astronomy). PMID- 10525465 TI - The Ortho-to-Para Ratio of Ammonia in the L1157 Outflow. AB - We have measured the ortho-to-para ratio of ammonia in the blueshifted gas of the L1157 outflow by observing the six metastable inversion lines from &parl0;J,K&parr0;=&parl0;1,1&parr0; to (6, 6). The highly excited (5, 5) and (6, 6) lines were first detected in the low-mass star-forming regions. The rotational temperature derived from the ratio of four transition lines from (3, 3) to (6, 6) is 130-140 K, suggesting that the blueshifted gas is heated by a factor of approximately 10 as compared to the quiescent gas. The ortho-to-para ratio of the NH3 molecules in the blueshifted gas is estimated to be 1.3-1.7, which is higher than the statistical equilibrium value. This ratio provides us with evidence that the NH3 molecules have been evaporated from dust grains with the formation temperature between 18 and 25 K. It is most likely that the NH3 molecules on dust grains have been released into the gas phase through the passage of strong shock waves produced by the outflow. Such a scenario is supported by the fact that the ammonia abundance in the blueshifted gas is enhanced by a factor of approximately 5 with respect to the dense quiescent gas. PMID- 10525466 TI - Detection of Polarized CO Emission from the Molecular Outflow in NGC 1333 IRAS 4A. AB - We report the first interferometric detection and mapping of linearly polarized spectral line emission due to the Goldreich-Kylafis effect. Our polarization maps of the CO J=2-->1 line in the molecular outflow powered by the very young stellar system NGC 1333 IRAS 4A make it possible to define the direction of the magnetic field in the outflow. Comparison with theoretical predictions implies that the magnetic field is parallel to the polarization. Our data suggest that the deflection of the outflow may be the result of the interaction between the outflow and the magnetic field. We also detect and map the linearly polarized dust continuum emission at 1.3 mm. The polarization map of the dust continuum is roughly consistent with an hourglass magnetic field morphology; i.e., it is in agreement with theoretical models of interstellar cloud contraction with a frozen in magnetic field. The two techniques for mapping magnetic field morphologies agree. In general, the two techniques sample different column densities, and together they allow the study of magnetic field morphology over wider areas than either technique by itself would permit. PMID- 10525467 TI - The Detached Dust Shell around the Massive Star HD 179821. AB - We have used the Keck I telescope to resolve at three mid-IR wavelengths the emission from HD 179821 (= RAFGL 2343), a G-type supergiant of perhaps 30 M middle dot in circle with a detached dust shell. The shell is very approximately circular in shape with an inner diameter of approximately 3&farcs;3, corresponding to 3.0x1017 cm. We estimate that the star was losing approximately 4x10-4 M middle dot in circle yr-1 until about 1800 yr ago, when the mass loss slowed dramatically. During the past approximately 104 yr, the star has lost approximately 10% of its initial mass. The star lies about 0&farcs;35 off center and is closer to the brighter, northern hemisphere of the nebula, which can be explained if the outflow velocity Vinfinity deviates by +20% from the average in the southern hemisphere and -20% from the average in the northern hemisphere. The mass-loss rate M&d2;(straight theta) may have been inversely correlated with the outflow velocity so that the momentum outflow p&d2; was isotropic during the mass loss phase. It also seems that M&d2;totalVinfinity was within a factor of 2 of L*/c, where L* is the current luminosity of the star; the mass loss may have been driven by radiation pressure. These results may help characterize the asymmetric circumstellar winds into which supernova explosions propagate. PMID- 10525468 TI - Meson Synchrotron Emission from Central Engines of Gamma-Ray Bursts with Strong Magnetic Fields. AB - Gamma-ray bursts (GRBs) are presumed to be powered by the still unknown central engines with timescales in the range from 1 ms to approximately a few seconds. We propose that the GRB central engines would be a viable site for strong meson synchrotron emission if they were compact astrophysical objects, such as neutron stars or rotating black holes with extremely strong magnetic fields (H approximately 1012-1017 G), and if protons or heavy nuclei were accelerated to ultrarelativistic energies on the order of approximately 1012-1022 eV. We show that the charged scalar mesons like pi+/- and heavy vector mesons like rho, which have several decay modes onto pi+/-, could be emitted, with a high intensity that is a thousand times larger than photons, through strong couplings to ultrarelativistic nucleons. These meson synchrotron emission processes eventually produce a burst of very high energy cosmic neutrinos with 1012 eV50%) of the in ecliptic interplanetary medium during our survey. We suggest that these suprathermal ions may therefore be a source population that is available for further acceleration by interplanetary shocks that accompany large SEP events, thereby leading to the 3He enhancements in a significant fraction of large SEP events. This impulsive SEP event material might also account for recent observations of large solar particle events with energetic particle ionization states that have a wide range of ionization states that encompass values expected for both gradual and impulsive solar SEP events. PMID- 10525473 TI - HIV-1 nuclear import: in search of a leader. AB - The ability of HIV-1 to use host cell nuclear import machinery to translocate the viral pre-integration complex into the cell nucleus is the critical determinant in the replication of the virus in non-dividing cells, such as macrophages. In this review, we describe the viral and cellular factors involved in this process. The available data suggest that the process of HIV-1 nuclear import is driven by interaction between nuclear localization signals (NLSs) present on viral proteins matrix and integrase and the cellular NLS receptor, karyopherin alpha. However, this interaction by itself is weak and insufficient to insure effective import of the pre-integration complex. Viral protein R (Vpr) functions to increase the affinity of interaction between viral NLSs and karyopherin alpha, thus substantially enhancing the karyophilic potential of the pre-integration complex. Interestingly, some cells, in particular HeLa, seem to contain a factor that can substitute for the Vpr's activity, making HIV-1 replication in such cells Vpr independent. We also describe a class of novel anti-HIV compounds that target the NLSs of HIV-1 and effectively block viral replication in T cells and macrophages. PMID- 10525474 TI - Metabolic disturbances and synovial joint responses in osteoarthritis. AB - Previously held views that the pathogenesis of idiopathic osteoarthritis (OA) originated in the synovial joint and was not influenced by systemic metabolic disturbances in the patient is inconsistent with recent data demonstrate skewing of the growth hormone/insulin-like growth factor-1 axis in the symptomatic OA patient. In light of this novel information, the role of growth hormone and insulin-like growth factor-1 in the pathogenesis and progression of OA requires further definition. In male patients with OA, the red blood cell sequesters more growth hormone than an aged-matched control group. Thus, this growth hormone "depot" may provide a mechanism for removal of "toxic" levels of growth hormone from the circulation. Storage of "excess" growth hormone in red cells may reduce the inflammatory or otherwise undesirable "toxic" actions of GH. In some patients, serum growth hormones levels may exceed three-times the average value considered normal. These "episodic" variations in growth hormone levels may play a significant role in the elevated levels of serum growth hormone seen in the OA patient. The connection between elevated growth hormone and decreased insulin like growth factor-1 levels and the defined cartilage anabolic and catabolic pathways defined in in vitro assays of articular cartilage derived from the OA patients remain to be more precisely defined. However, the dampened insulin-like growth factor-1 response in OA coupled with elevated cartilage extracellular matrix degradation (mediated by metalloproteinases) and depressed compensatory biosynthesis (induced and perpetuated by the presence of cytokines such as interleukin-1 and tumor necrosis factor-alpha) may, in fact, act synergistically to suppress normal cartilage repair mechanisms thus resulting in progressive destructive lesions of the cartilage and bone. PMID- 10525475 TI - Degradation and repair of articular cartilage. AB - Approximately 95,000 total knee replacements and 41,000 other surgical procedures to repair cartilaginous defects of the knee are performed annually in the United States (1). The response of normal articular cartilage to injury or arthritic degeneration is often a sub-optimal repair; the biochemical and mechanical properties of the new tissue differ from the native cartilage, resulting in inadequate or altered function. It is believed that the chondrocytes from the surrounding areas, although perhaps capable of some limited migration at the damaged site, are not able to proliferate and produce the macromolecules necessary to create an organized matrix characteristic of normal articular cartilage (2,3). Current therapeutic options for articular cartilage injuries and degeneration have resulted in repair tissue which may be hyaline-like, but does not approximate the durability and function of the normal articular surface. Numerous studies have been performed to increase our understanding of the normal repair process of articular cartilage and its limitations, and to devise methods and materials to regenerate the joint surface. PMID- 10525477 TI - Potential regulation of cartilage metabolism in osteoarthritis by fibronectin fragments. AB - There are few candidates for biochemical pathways that either initiate or amplify catabolic processes involved in osteoarthritis (OA). Perhaps, one of the most likely sources for such pathways may be within the extracellular matrix itself. This review focuses on an example of how specific degradation products of the extracellular matrix of cartilage, produced during proteolytic damage, have the potential to enhance OA-like processes. In this example, these products can induce or activate other factors, such as catabolic cytokines, that amplify the damage. The damage, in turn, enhances levels of the degradation products themselves, as in a positive feedback loop. Since these products are derived from the cartilage matrix, they could be considered barometers of the health of the cartilage that signal to the chondrocyte, through outside to inside signaling, the health or status of the surrounding matrix. The best example and most characterized system is that of fragments of the matrix protein, fibronectin (Fn), although as discussed later, other recently discovered fragment systems may also have the potential to regulate cartilage metabolism. In the case of Fn fragments (Fn-fs), the Fn-fs enhance levels of catabolic cytokines as in OA and, thus, are potentially earlier damage mediators than catabolic cytokines. The Fn fs up-regulate matrix metalloproteinase (MMP) expression, significantly enhance degradation and loss of proteoglycan (PG) from cartilage and temporarily suppress PG synthesis, all events observed in OA. However, this Fn-f system may be involved in normal cartilage homeostasis as well. For example, low concentrations of Fn-fs enhance anabolic activities and could play a role in normal homeostasis. This system may also be involved in not only amplifying damage but also coupling damage to repair. For example, high concentrations of Fn-fs that might arise in OA temporarily offset the anabolic response of lower Fn-f concentrations and cause short-term enhanced catabolic events that are followed by slowly increasing anabolic responses. Such effects would be expected for mediators with roles in regulation of metabolism in both normal or diseased cartilage. Other products of matrix degradation have also been shown to regulate cartilage metabolism. A common mechanistic theme to these systems may be that they perturb the cartilage matrix and directly or indirectly alter function of specific receptors involved in metabolism. These concepts illustrate the potential of the cartilage matrix to regulate its composition in both health and disease. PMID- 10525476 TI - Regulation of chondrocyte gene expression. AB - Extracellular influences known to affect the regulation of chondrocyte biosynthetic and catabolic activity have been shown to include soluble factors, extracellular matrix and mechanical stress. A balance of these numerous extracellular influences is required for normal function of articular cartilage. It is likely that OA is the result of an imbalance of regulatory influences, ultimately resulting in deleterious changes in gene expression, altered extracellular matrix (ECM) and tissue degeneration. Molecular signaling via soluble mediators has been shown to be crucial to cartilage homeostasis. A number of vitamins, hormones, growth/differentiation factors and cytokines have been implicated in chondrocyte differentiation and cartilage metabolism. During normal maintenance, as well as in aging and pathology, these soluble factors can significantly influence the physical properties and the function of cartilage. Chondrocytes, like cells in other tissues, exist within an information-rich extracellular environment consisting of ECM molecules, a milieu which interacts with and modulates the activity of growth factors, hormones and ECM remodeling enzymes. Cell surface matrix receptors, including a family of proteins known as integrins, connect structural information in the ECM to a complex cellular response mechanism in the cell's interior. Integrins on cell surfaces detect and transduce signals in a cooperative manner with other adhesion receptor classes and/or growth factor receptors. The effects of mechanical stress upon a number of chondrocyte biological parameters has been examined in several laboratories. Other investigations have addressed the mechanism by which mechanical force affects biochemical and biosynthetic processes in chondrocytes, in particular synthesis of aggrecan, a major component of the cartilage ECM. Each of these extracellular influences upon chondrocyte metabolism may affect regulation of chondrocyte ECM biosynthesis at many levels, including mRNA transcription, RNA splicing, nuclear transport, protein translation, post-translational modification, intracellular vesicular transport, and protein secretion. Transcriptional regulation of some of the major protein and proteoglycan components of the cartilage ECM has been examined in a number of species, and promoters have been characterized for aggrecan, link protein and collagen type II genes. There is evidence that gene expression may be altered in OA cartilage, providing clues as to which subsets of genes expressed in chondrocytes may be considered relevant to OA pathophysiology. PMID- 10525478 TI - Fundamental pathways in osteoarthritis: an overview. AB - Osteoarthritis (OA) is a significant world-wide health problem owing to the progressive and debilitating nature of the condition which results in high morbidity and a marked decrease in the quality of life. Significant advances in the medical and surgical management of OA have resulted from an understanding of the fundamental pathways governing the health and disease of synovial joint tissues. Continuing investigations into the nature of synovial joint pathophysiology at both the molecular and biochemical level should pave the way for the development of novel therapeutic strategies, including gene therapy and tissue engineering, in the treatment of the OA patient. PMID- 10525479 TI - Future directions for research and treatment of osteoarthritis. AB - Future directions in the research and treatment of osteoarthritis (OA) will be based on the emerging picture of pathophysiological events that govern the initiation and progression of OA. The fundamental event resulting in the destruction of articular cartilage in OA arises from an imbalance between anabolic and catabolic pathways. The extracellular matrix (ECM) of cartilage is degraded by matrix metalloproteinases (MMPs) induced by cytokines. Cytokines also blunt chondrocyte compensatory synthesis pathways required to restore the integrity of the degraded ECM. Inhibition of the MMPs, their activators, and cytokines that induce MMP gene up-regulation would appear to be fertile targets for drug development in the treatment of OA. Restoration of damaged articular surfaces via tissue engineering strategies which could employ chondroprogenitor cells in biomatrices appropriate for transplantation to cartilage surfaces appears feasible. A reduction in cytokine-mediated up-regulation of MMP gene expression as well as augmentation of cartilage ECM biosynthesis may also be possible by employing the principles of gene transfer using suitable vectors that establish long-term stable expression of genes which suppress MMPs while at the same time supporting cartilage ECM biosynthesis. PMID- 10525480 TI - Cytokines and their role in the pathophysiology of osteoarthritis. AB - The specific causative agent of the pathological process of osteoarthritis (OA) has not yet been identified, however, episodic inflammation at the clinical stage is now a well documented phenomenon and believed to be involved in the disease progression. Interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF alpha) are the predominant proinflammatory cytokines synthesized during the OA process. Other cytokines having proinflammatory properties or catabolic factors could also contribute to this pathological condition, and those having antiinflammatory properties may be able to counteract the negative effects of the former on the disease process. In this chapter, we will review cytokine interactions and their modulatory effects on joint articular tissue metabolism, including their stimulatory and/or inhibitory actions, and their potential relevance to OA. We will also briefly survey the major biological factors, in relation to cytokines, that look promising for future therapeutic approaches. PMID- 10525481 TI - An introduction to the pathophysiology of osteoarthritis. AB - Osteoarthritis involves the degeneration of articular cartilage together with changes in subchondral bone and limited intra-articular inflammation. In this chapter these changes are reviewed at the tissue, cell and molecular levels to reveal the complexity of a process which involves multiple changes in joint structure and turnover. PMID- 10525482 TI - Developmental patterns of cartilage. AB - The current state of knowledge of cartilage differentiation leaves many questions unanswered. This review provides an up-to-date examination of current thinking on the subject of developmental patterns in cartilage formation. We will discuss the current model of limb elongation as well as the molecular aspects of chondrogenesis and growth plate formation. This will then be compared with the limited information currently known about the molecular aspects of osteoarthritis. PMID- 10525483 TI - Degradative enzymes in osteoarthritis. AB - A central feature of the osteoarthritic disease process involves erosive destruction of the articular cartilage extracellular matrix (ECM) on the surfaces of diarthrotic joints. The resultant loss of joint function makes studies on mechanisms underlying ECM degradation critical for treatment of the disease and prevention of disability. Candidate pathways to account for the loss of cartilage involve expression of a combination of proteases that degrade the major cartilage matrix macromolecules, aggrecan and type II collagen. The specific types of enzymatic activities associated with the progressive removal of ECM and severity of joint disease include the matrix metalloproteinases, collagenase, gelatinase and aggrecanase(s). The degradative enzymes originate in synovial cells, cartilage cells, the chondrocytes, distributed within the ECM and leukocytes that actively invade the joint space. Specific enzymes arising from each of these tissues exhibit selective ECM degrading properties; the different categories of these tissue-derived enzymes will be discussed in this chapter. A perspective on the efficacy of existing agents and the potential for development of novel therapeutic agents is also included. While the degradative enzymes serve as a focal point for therapeutic intervention, a fundamental understanding of the mechanisms underlying degradative enzyme expression in osteoarthritis remains an important goal for prevention of disease. PMID- 10525484 TI - The role of cardiac MRI stress testing : "Make a better mouse trap...". PMID- 10525485 TI - Vasoconstriction to endogenous endothelin-1 is increased in the peripheral circulation of patients with essential hypertension. AB - BACKGROUND: In humans, endothelin (ET)-1 could be implicated in the pathophysiology of several cardiovascular diseases, including essential hypertension. We therefore evaluated the role of ET-1 in control of vascular tone in essential hypertension. METHODS AND RESULTS: We used strain-gauge venous plethysmography to test changes in forearm blood flow induced by intrabrachial infusion of TAK-044 (10, 30, and 100 microgram. 100 mL(-1). min(-1)), an ET(A)/ET(B) receptor antagonist, or sodium nitroprusside (1 and 2 microgram. 100 mL(-1). min(-1)), a vasodilator that acts on smooth muscle cells, in hypertensive patients and healthy controls (n=10 in each group). The NO pathway was also evaluated by infusion of N(G)-monomethyl-L-arginine, (L-NMMA; 10, 30, and 100 microgram. 100 mL(-1). min(-1)), an NO synthase inhibitor, and norepinephrine (3, 9, and 30 ng. 100 mL(-1). min(-1)) as control. Immunoreactive plasma ET-1 was measured by radioimmunoassay. In hypertensive patients, TAK-044 caused a vasodilation that was significantly (P<0.01) increased compared with normotensive subjects. Moreover, vasoconstriction to L-NMMA was significantly (P<0.01) decreased in hypertensive patients compared with controls. In contrast, the vascular responses to sodium nitroprusside and norepinephrine, as well as levels of immunoreactive plasma ET-1, were similar in hypertensive patients and controls. In the study population, vasodilation to TAK-044 and vasoconstriction to L-NMMA showed an inverse correlation (r=-0.56, P<0.05). CONCLUSIONS: These results indicate that TAK-044 caused a greater degree of vasodilation in the forearm vessels of essential hypertensive patients compared with normotensive subjects, an alteration associated with decreased tonic NO release. PMID- 10525486 TI - beta-Particle-emitting radioactive stent implantation. A safety and feasibility study. AB - BACKGROUND: This study represents the Heart Center Rotterdam's contribution to the Isostents for Restenosis Intervention Study, a nonrandomized multicenter trial evaluating the safety and feasibility of the radioactive Isostent in patients with single coronary artery disease. Restenosis after stent implantation is primarily caused by neointimal hyperplasia. In animal studies, beta-particle emitting radioactive stents decrease neointimal hyperplasia by inhibiting smooth muscle cell proliferation. METHODS AND RESULTS: The radioisotope (32)P, a beta particle emitter with a half-life of 14.3 days, was directly embedded into the Isostent. The calculated range of radioactivity was 0.75 to 1.5 microCi. Quantitative coronary angiography measurements were performed before and after the procedure and at 6-month follow-up. A total of 31 radioactive stents were used in 26 patients; 30 (97%) were successfully implanted, and 1 was embolized. Treated lesions were in the left anterior descending coronary artery (n=12), the right coronary artery (n=8), or the left circumflex coronary artery (n=6). Five patients received additional, nonradioactive stents. Treated lesion lengths were 13+/-4 mm, with a reference diameter of 2.93+/-0. 47 mm. Minimum lumen diameter increased from 0.87+/-0.28 mm preprocedure to 2.84+/-0.35 mm postprocedure. No in hospital adverse cardiac events occurred. All patients received aspirin indefinitely and ticlopidine for 4 weeks. Twenty-three patients (88%) returned for 6-month angiographic follow-up; 17% of them had in-stent restenosis, and 13% had repeat revascularization. No restenosis was observed at the stent edges. Minimum lumen diameter at follow-up averaged 1.85+/-0.69 mm, which resulted in a late loss of 0.99+/-0. 59 mm and a late loss index of 0.53+/-0.35. No other major cardiac events occurred during the 6-month follow-up. CONCLUSIONS: The use of radioactive stents with an activity of 0.75 to 1.5 microCi is safe and feasible. PMID- 10525487 TI - Effects of testosterone on coronary vasomotor regulation in men with coronary heart disease. AB - BACKGROUND: The increased incidence of coronary artery disease in men compared with premenopausal women suggests a detrimental role of male hormones on the cardiovascular system. However, testosterone has direct relaxing effects on coronary arteries in animals, as shown both in vitro and in vivo. The effect of testosterone on the human coronary circulation remains unknown. METHODS AND RESULTS: We studied 13 men (aged 61+/-11 years) with coronary artery disease. They underwent measurement of coronary artery diameter and blood flow after a 3 minute intracoronary infusion of vehicle control (ethanol) followed by 2-minute intracoronary infusions of acetylcholine (10(-7) to 10(-5) mol/L) until peak velocity response. A dose-response curve to 3-minute infusions of testosterone (10(-10) to 10(-7) mol/L) was then determined, and the acetylcholine infusions were repeated. Finally, an intracoronary bolus of isosorbide dinitrate (1000 microgram) was given. Coronary blood flow was calculated from measurements of blood flow velocity using intracoronary Doppler and coronary artery diameter using quantitative coronary angiography. Testosterone significantly increased coronary artery diameter compared with baseline (2.78+/-0. 74 mm versus 2.86+/ 0.72 mm [P=0.05], 2.87+/-0.71 mm [P=0.038], and 2.90+/-0.75 mm [P=0.005] for baseline versus testosterone 10(-9) to 10(-7) mol/L, respectively). A significant increase in coronary blood flow occurred at all concentrations of testosterone compared with baseline (geometric mean [95% CI]: 32 [25, 42] versus 36.3 [27, 48] (P=0.006), 35.3 [26, 47] (P=0.029), 36.8 [28, 49] (P=0.002), and 37 [28, 48] (P=0.002), mL/min for baseline versus testosterone 10(-10) to 10(-7) mol/L, respectively). No differences existed in coronary diameter or blood flow responses to acetylcholine before versus after testosterone. CONCLUSIONS: Short term intracoronary administration of testosterone, at physiological concentrations, induces coronary artery dilatation and increases coronary blood flow in men with established coronary artery disease. PMID- 10525488 TI - Utility of fast cine magnetic resonance imaging and display for the detection of myocardial ischemia in patients not well suited for second harmonic stress echocardiography. AB - BACKGROUND: Some patients referred for pharmacological stress testing with transthoracic echocardiography (TTE) are unable to undergo testing owing to poor acoustic windows. Fast cine MRI can be used to assess left ventricular contraction, but its utility for detection of myocardial ischemia in patients poorly suited for echocardiography is unknown. METHODS AND RESULTS: One hundred fifty-three patients (86 men and 67 women aged 30 to 88 years) with poor acoustic windows that prevented adequate second harmonic TTE imaging were consecutively referred for MRI to diagnose inducible myocardial ischemia during intravenous dobutamine and atropine. Diagnostic studies were completed in an average of 53 minutes. No patients experienced myocardial infarction, ventricular fibrillation, exacerbation of congestive heart failure, or death. In patients who underwent computer-assisted quantitative coronary angiography, the sensitivity and specificity for detecting a >50% luminal diameter narrowing were 83% and 83%, respectively. In the 103 patients with a negative MRI examination, the cardiovascular occurrence-free survival rate was 97%. CONCLUSIONS: Fast cine cardiac MRI provides a mechanism to assess left ventricular contraction and diagnose inducible myocardial ischemia in patients not well suited for stress echocardiography. PMID- 10525489 TI - Comparison of naive sixth-grade children with trained professionals in the use of an automated external defibrillator. AB - BACKGROUND: Survival after out-of-hospital cardiac arrest (OHCA) is strongly influenced by time to defibrillation. Wider availability of automated external defibrillators (AEDs) may decrease response times but only with increased lay use. Consequently, this study endeavored to improve our understanding of AED use in naive users by measuring times to shock and appropriateness of pad location. We chose sixth-grade students to simulate an extreme circumstance of unfamiliarity with the problem of OHCA and defibrillation. The children's AED use was then compared with that of professionals. METHODS AND RESULTS: With the use of a mock cardiac arrest scenario, AED use by 15 children was compared with that of 22 emergency medical technicians (EMTs) or paramedics. The primary end point was time from entry onto the cardiac arrest scene to delivery of the shock into simulated ventricular fibrillation. The secondary end point was appropriateness of pad placement. All subject performances were videotaped to assess safety of use and compliance with AED prompts to remain clear of the mannequin during shock delivery. Mean time to defibrillation was 90+/-14 seconds (range, 69 to 111 seconds) for the children and 67+/-10 seconds (range, 50 to 87 seconds) for the EMTs/paramedics (P<0.0001). Electrode pad placement was appropriate for all subjects. All remained clear of the "patient" during shock delivery. CONCLUSIONS: During mock cardiac arrest, the speed of AED use by untrained children is only modestly slower than that of professionals. The difference between the groups is surprisingly small, considering the naivete of the children as untutored first time users. These findings suggest that widespread use of AEDs will require only modest training. PMID- 10525490 TI - Firing properties of single muscle vasoconstrictor neurons in the sympathoexcitation associated with congestive heart failure. AB - BACKGROUND: Congestive heart failure (CHF) in humans is associated with a marked sympathoexcitation, including an augmented muscle sympathetic nerve activity (MSNA) in intraneural multiunit recordings. In the present study, single-unit recording was used to evaluate whether the firing properties of individual muscle vasoconstrictor neurons can reveal underlying mechanisms for this increase in MSNA. METHODS AND RESULTS: Eight patients with CHF (NYHA class II to IV; left ventricular ejection fraction, 29+/-5%, mean+/-SEM) were studied. In standard multiunit recordings, MSNA burst incidence (bursts/100 heartbeats) ranged from 65% to 100% (88+/-5%). Using selective tungsten microelectrodes, we made recordings from 16 single muscle vasoconstrictor axons. Mean unit firing probability (ie, the percentage of cardiac intervals in which a single axon fired) was 54.5+/-5.2% (range, 21 to 89%), and mean firing frequency was 0.98+/ 0.22 Hz (0.14 to 3.86 Hz), both of which were higher than seen previously in healthy subjects (P<0.001). Although single neurons occasionally generated multiple spikes per sympathetic burst, such multiple firing was rare and was not different from that seen in healthy subjects. CONCLUSIONS: An increased firing frequency of individual vasoconstrictor neurons is one mechanism for the increased number of multiunit MSNA bursts at rest in CHF. The neurons discharge in more diastoles than in healthy subjects (ie, firing probability is increased), but the likelihood of discharging >1 impulse per sympathetic burst is not increased. Despite the intense multiunit activity at rest, the firing characteristics of individual vasoconstrictor axons indicate a remaining capacity for transient increases of MSNA in CHF. PMID- 10525491 TI - Mechanical remodeling of the left atrium after loss of atrioventricular synchrony. A long-term study in humans. AB - BACKGROUND: Tachycardia-mediated mechanical remodeling of the atrium is considered central to the pathogenesis of thromboembolism associated with chronic atrial fibrillation. Whether atrial mechanical remodeling also occurs in response to atrial stretch induced by chronic asynchronous ventricular pacing in patients with permanent pacemakers is unknown. METHODS AND RESULTS: The study design was a prospective randomized comparison between 21 patients paced chronically in the VVI mode and 11 patients paced chronically in the DDD mode for 3 months. Left atrial appendage (LAA) function and the presence of spontaneous echo contrast (SEC) were determined with transesophageal echocardiography (TEE) within 24 hours of pacemaker implantation and after 3 months. The VVI patients were then programmed to DDD and underwent a third TEE after DDD pacing for an additional 3 months. After chronic VVI pacing, LAA velocity decreased from 82.4+/-29.0 to 42.1+/-25.4 cm/s (P<0.01), LAA fractional area change decreased from 74.9+/-17.2% to 49.8+/-22.0% (P<0.01), and 4 patients (19%) developed left atrial SEC (P<0.05). With the reestablishment of chronic AV synchrony, LAA velocity increased to 61.6+/-18.5 cm/s (P<0.01), LAA fractional area change increased to 76.4+/-18.1% (P<0.01), and SEC resolved. In the 11 patients undergoing chronic DDD pacing, no significant changes in LAA velocity (baseline, 86.0+/-28.8 cm/s versus 3 months, 79.6+/-14. 9 cm/s) or LAA fractional area change (baseline, 76.2+/-19.4% versus 72.5+/-15.7%) were demonstrated, and SEC did not develop. CONCLUSIONS: Chronic loss of AV synchrony induced by VVI pacing is associated with mechanical remodeling of the left atrium, which may reverse after the reestablishment of AV synchrony with DDD pacing. This process may be partly responsible for the higher incidence of thromboembolism observed in patients undergoing VVI pacing compared with AV sequential pacing. PMID- 10525492 TI - The gene encoding atrial natriuretic peptide and the risk of human stroke. AB - BACKGROUND: Recent evidence from an animal model of stroke, the stroke-prone spontaneously hypertensive rat, implicated the gene encoding atrial natriuretic peptide (ANP) as a possible candidate contributing to the likelihood of experiencing a stroke. The purpose of the present study was to investigate the role of ANP in the pathogenesis of cerebrovascular accidents in humans. METHODS AND RESULTS: We investigated 2 previously known markers at ANP, G1837A and T2238C, for their possible association with the occurrence of stroke. This was the largest matched case-controlled sample studied thus far; the sample was drawn from a large prospective study (the Physician's Health Study). When assuming a dominant mode of inheritance, a statistically significant positive association was observed for the 1837A allele, indicating an odds ratio of 1.64 (95% confidence interval, 1.01 to 2.65) for stroke. This observation led to the discovery of a new molecular variant in exon 1, G664A, which was responsible for a valine-to-methionine substitution in the proANP peptide. This mutation, which was in linkage disequilibrium with the G1837A marker, was associated with the occurrence of stroke (odds ratio, 2.0; 95% confidence interval, 1.17 to 3.19; P=0.01). CONCLUSIONS: Our findings suggest that molecular variants of the ANP gene may represent an independent risk factor for cerebrovascular accidents in humans. The strong parallelism to the experimental data obtained in the stroke prone animal model provides assurance for the relevance of our observation. PMID- 10525493 TI - The direct antiatherogenic effect of estrogen is present, absent, or reversed, depending on the state of the arterial endothelium. A time course study in cholesterol-clamped rabbits. AB - BACKGROUND: This study further investigated the relationship between estrogen, arterial endothelium, and nitric oxide (NO) in cholesterol-clamped rabbits. METHODS AND RESULTS: Rabbits were ovariectomized, balloon-injured in the thoracic aorta, and grouped to receive cholesterol-enriched chow together with either 17beta-estradiol or vehicle for 1, 2, 4, or 8 weeks. In the undamaged aorta, cholesterol accumulation of the placebo rabbits was significantly increased from week 4 to 8 (P<0.001). This increase was almost completely inhibited by estrogen (P<0.001). In the balloon-injured aorta, the estrogen and placebo rabbits accumulated similar amounts of cholesterol in the reendothelialized areas. In the deendothelialized areas, the estrogen group surprisingly accumulated significantly more cholesterol than the placebo group. This difference was apparent from week 2 and became significant at week 8 (P<0.01). Circulating nitrite/nitrate were significantly increased by estrogen at weeks 1, 2, and 4 but not at week 8. Similarly, in additional experiments, basal NO release was significantly higher in estrogen-treated than in placebo-treated rabbits after 4 (P<0.05) but not after 8 weeks. Stimulated NO release and endothelial NO synthase activity did not differ between groups. Mononuclear-endothelial cell binding was reduced by 50% by estrogen after 4 weeks (P<0.05). This difference, however, was abolished by coadministration of N(G)-nitro-L-arginine methyl ester, an inhibitor of NO production. CONCLUSIONS: The direct antiatherogenic effect of estrogen was present, absent, or reversed, depending on the state of the arterial endothelium, and preceded by a transient increase in NO production followed by a reduced mononuclear-endothelial cell binding. PMID- 10525494 TI - Progressive cardiac dysfunction and fibrosis in the cardiomyopathic hamster and effects of growth hormone and angiotensin-converting enzyme inhibition. AB - BACKGROUND: Growth hormone (GH) improves cardiac function in the rat with myocardial infarction, but its effects in a model of primary dilated cardiomyopathy have not been reported. GH effects were examined at early (4 months) and late (10 months) phases of disease in the cardiomyopathic (CM) hamster, and the combination of GH with chronic ACE inhibition was assessed in late-phase heart failure. METHODS AND RESULTS: CM hamsters (CHF 147 line) at 4 months showed severe systolic left ventricular (LV) dysfunction with normal LV filling pressure, and at 10 months there was more severe systolic as well as diastolic dysfunction with increasing myocardial fibrosis. Recombinant human GH alone for 3 weeks at age 4 months increased LV wall thickness and reduced systolic wall stress without altering diastolic wall stress, whereas at 10 months, wall stress and fractional shortening did not improve. The LV dP/dt(max) was enhanced at both ages by GH, which at 4 months reflected increased contractility, but at 10 months was most likely caused by elevation of the LV filling pressure. The increasing degree of fibrosis correlated inversely with LV function but was unaffected by GH. In other CM hamsters, high-dose ACE inhibition alone (quinapril), started at 8 months and continued for 11 weeks, improved LV function and inhibited unfavorable remodeling, but the addition of GH for 3 weeks at age 10 months produced increased wall thickness with little additional functional benefit and increased the LV filling pressure and diastolic wall stress. CONCLUSIONS: GH treatment alone improved LV dysfunction at 4 months of age in CM hamsters by increasing contractility and reducing wall stress but had few beneficial effects at 10 months in severe LV failure. After chronic ACE inhibition, addition of GH at 10 months had no additional beneficial effects and further increased LV diastolic pressure. These differing effects of GH may relate to the progressive increase of LV fibrosis in the CM hamster. PMID- 10525495 TI - Electroanatomic left ventricular mapping in the porcine model of healed anterior myocardial infarction. Correlation with intracardiac echocardiography and pathological analysis. AB - BACKGROUND: Catheter ablation for ventricular tachycardia in healed infarction is limited to patients with inducible, tolerated arrhythmias. Strategies that would allow mapping during sinus rhythm might obviate this limitation. METHODS AND RESULTS: Two sets of experiments were performed in adult pigs to refine a new technique for left ventricular mapping. First, detailed endocardial maps were done in 5 normal pigs and 7 pigs 6 to 10 weeks after left anterior descending coronary artery infarction to characterize electrograms in normal and infarcted tissue by electroanatomic mapping (CARTO, Biosense). Electrogram recording sites were verified by intracardiac echo (ICE, 9 MHz) and grouped by location: infarct (area of akinesis by ICE), border (0.5-cm perimeter of akinetic area), and remote. Compared with remote sites, electrograms from infarct sites had smaller amplitudes (1.2+/-0.5 versus 5.1+/-2.1 mV, P<0.001), longer durations (74.2+/ 26.3 versus 36.3+/-6.4 ms, P<0.001), and more frequent notched or late components. Border zone electrograms were intermediate in amplitude and duration. Second, infarct characterization by electroanatomic mapping was compared with pathological (exclusion of triphenyltetrazolium chloride staining) and ICE measurements. Infarct size by pathology correlated with the area defined by contiguous electrograms with amplitude /= 40 cigarettes per day. There was no interaction of smoking with use of oral contraceptives, but there were additive risks with other clinical risk factors such as hypertension and diabetes. It is estimated that if all women aged 16-44 years were able to stop smoking, 400 cases of myocardial infarction per annum (of whom 112 would die) would be prevented. CONCLUSIONS: In young women the risk of myocardial infarction from smoking was considerable, and heavy smokers with other risk factors were especially at risk. PMID- 10525514 TI - Non-adherence with ACE inhibitor treatment is common in heart failure and can be detected by routine serum ACE activity assays. AB - OBJECTIVE: To assess whether serum angiotensin converting enzyme (ACE) activity during routine clinical practice accurately reflects patient adherence to ACE inhibitor treatment for chronic heart failure (CHF). DESIGN: Retrospective assessment of ACE inhibitor adherence and serum ACE activity measurements. SETTING: Teaching hospital outpatient department PATIENTS AND INTERVENTIONS: During 1994-95, serum ACE was measured in 73 CHF patients who were routinely attending the heart failure clinic at Ninewells Hospital. At the same time, the medicines monitoring unit collected data on whether and when prescriptions for ACE inhibitors were redeemed at community pharmacies, which enabled each patient's adherence over a prolonged period to be assessed. MAIN OUTCOME MEASURES: Routine collected serum ACE measurements were correlated with measured adherence with ACE inhibitor treatment. RESULTS: In total, 18% of CHF patients appeared to exhibit < 70% adherence with their ACE inhibitor treatment with 34% exhibiting less than 85% adherence and 58% exhibiting < 100% adherence. A serum ACE activity of > 12 u/l gave 91% positive predictive accuracy that the patient was < 100% adherent with their ACE inhibitor treatment. At the other extreme, a serum ACE < 6.5 u/l gave 81% positive predictive accuracy that the patient was > 85% adherent with ACE inhibitor treatment. CONCLUSIONS: Non-adherence with ACE inhibitor treatment was found to be common in patients with CHF. The simple, inexpensive test of serum ACE activity can be used in CHF patients to identify many, although not all, non-adherent patients so that adherence enhancing strategies can be targeted towards them. Further work is clearly required to explore the precise clinical use of this promising test. PMID- 10525515 TI - Impact of concurrent amiodarone treatment on the tolerability and efficacy of carvedilol in patients with chronic heart failure. AB - OBJECTIVE: To assess the safety and efficacy of carvedilol when administered to heart failure patients already receiving amiodarone. DESIGN: Retrospective analysis of the clinical outcome of 230 patients treated with carvedilol for chronic heart failure, stratified according to whether they were already receiving amiodarone (amiodarone group, 80 patients) or not (non-amiodarone group, 130 patients) at baseline. SETTING: Heart failure clinic at a university affiliated public teaching hospital. MAIN OUTCOME MEASURES: Incidence of adverse events; changes in functional status and echocardiographic dimensions at three months. RESULTS: Adverse reactions to carvedilol occurred in 33 (41%) of the amiodarone group and 43 (29%) of the non-amiodarone group (p = 0.049). Carvedilol was discontinued in 21 (26%) of the amiodarone group and 37 (25%) of the non amiodarone group (NS). The clinical outcome at three months did not differ significantly between the two groups; 31 (39%) of the amiodarone group improved their New York Heart Association status, 28 (35%) were unchanged, and 21 (26%) deteriorated compared with 67 (45%), 51 (34%), and 32 (21%), respectively, for the non-amiodarone group (NS). Both groups had highly significant decreases in heart rate and left ventricular end systolic dimension, and a significant increase in left ventricular ejection fraction after three months of carvedilol treatment, with no significant differences between the groups. CONCLUSIONS: The beneficial effects of carvedilol on left ventricular remodelling, systolic function, and symptomatic status are not affected by concurrent treatment with amiodarone. Adverse reactions necessitating cessation of carvedilol are no more frequent in patients receiving amiodarone. PMID- 10525516 TI - Evolution and long term outcome in cases with fetal diagnosis of congenital heart disease: Italian multicentre study. Fetal Cardiology Study Group of the Italian Society of Pediatric Cardiology. AB - OBJECTIVES: To analyse the evolution and outcome in utero and after birth of infants with a fetal diagnosis of congenital heart disease. DESIGN: Inclusion criteria were the fetal diagnosis of congenital heart disease, confirmed postnatally or postmortem, and a complete follow up in utero and after birth. SETTING: 20 centres operating prenatal echocardiographic screening. PATIENTS: 847 cases were included in the study. Gestational age at diagnosis ranged from 15-39 weeks; in 370 cases (43.7%) the diagnosis was made before 24 weeks' gestation. RESULTS: 245/847 cases (28.9%) were terminated during pregnancy, 227 following early diagnosis; 128/245 cases (52. 2%) had associated anomalies and 117/245 (47.8%) had serious congenital heart disease. Of the remaining 602 cases that continued the pregnancy, 72 (11.9%) died in utero, 259 (43%) died postnatally (83 after surgery or invasive procedures), and 271 infants (45%) survived and presently range in age from 18 months to 13 years old. The mortality rate was higher in cases with associated extracardiac or chromosomal anomalies (68% and 74% of cases continuing pregnancy, respectively), and in cases with heart failure and complex cardiac defects. CONCLUSIONS: The data confirm a relevant fetal and postnatal loss in cases with complex congenital heart disease, and major clinical use of prenatal diagnosis in the management of ductus dependent anomalies. Negative prognostic factors for the outcome were associated anomalies and heart failure. PMID- 10525518 TI - Influence of left ventricular relaxation on the pressure half time of aortic regurgitation. AB - BACKGROUND: The severity of aortic regurgitation can be estimated using pressure half time (PHT) of the aortic regurgitation flow velocity, but the correlation between regurgitant fraction and PHT is weak. AIM: To test the hypothesis that the association between PHT and regurgitant fraction is substantially influenced by left ventricular relaxation. METHODS: In 63 patients with aortic regurgitation, subdivided into a group without (n = 22) and a group with (n = 41) left ventricular hypertrophy, regurgitant fraction was calculated using the difference between right and left ventricular cardiac outputs. Left ventricular relaxation was assessed using the early to late diastolic Doppler tissue velocity ratio of the mitral annulus (E/ADTI), the E/A ratio of mitral inflow (E/AM), and the E deceleration time (E-DT). Left ventricular hypertrophy was assessed using the M mode derived left ventricular mass index. RESULTS: The overall correlation between regurgitant fraction and PHT was weak (r = 0.36, p < 0.005). In patients without left ventricular hypertrophy, there was a significant correlation between regurgitant fraction and PHT (r = 0.62, p < 0.005), but not in patients with left ventricular hypertrophy. In patients with a left ventricular relaxation abnormality (defined as E/ADTI< 1, E/AM< age corrected lower limit, E-DT >/= 220 ms), no associations between regurgitant fraction and PHT were found, whereas in patients without left ventricular relaxation abnormalities, the regurgitant fraction to PHT relations were significant (normal E/AM: r = 0.57, p = 0.02; E DT< 220 ms: r = 0.50, p < 0.001; E/ADTI < 1: r = 0.57, p = 0.02). CONCLUSIONS: Only normal left ventricular relaxation allows a significant decay of PHT with increasing aortic regurgitation severity. In abnormal relaxation, which is usually present in left ventricular hypertrophy, wide variation in prolonged backward left ventricular filling may cause dissociation between the regurgitant fraction and PHT. Thus the PHT method should only be used in the absence of left ventricular relaxation abnormalities. PMID- 10525519 TI - Images in cardiology. Pitfall in the diagnosis of pericardial effusion by echocardiography. PMID- 10525517 TI - Stent implantation for aortic coarctation and recoarctation. AB - OBJECTIVE: To determine the early results of balloon expandable stent implantation for aortic coarctation or recoarctation. DESIGN: Prospective observational study. SETTING: Two paediatric cardiology tertiary referral centres. PATIENTS: 17 patients, median age 17 years (range 4.4 to 45) and median weight 61 kg (17 to 92). Six had native aortic coarctation and 11 had aortic recoarctation; 14 had upper limb systolic hypertension. Of those with recoarctation, eight had had at least one previous balloon dilatation attempt and two of these patients also had further surgical interventions. INTERVENTION: Balloon expandable Palmaz iliac stent implantation. MAIN OUTCOME MEASURES: Systolic pressures gradients, minimum aortic diameter, upper limb blood pressures, and incidence of aneurysm formation. RESULTS: 18 stents were implanted during 18 procedures in the 17 patients. Mean peak systolic pressure gradient fell from 26 mm Hg (95% confidence interval (CI), 21 to 31 mm Hg) before to 5 mm Hg (2 to 8 mm Hg) after stent implantation (p < 0.001), and mean minimum aortic diameter increased from 7 mm (95% CI, 6 to 8 mm) before to 11.3 mm (10 to 12.6 mm) after implantation (p < 0.001). Complications occurred in five patients (bleeding in two, stent migration in two, and aneurysm formation in one). Two patients remained borderline hypertensive and eight were receiving antihypertensive treatment at most recent assessment. CONCLUSIONS: Stent implantation for aortic recoarctation and native coarctation gives good immediate results. Careful follow up is necessary to evaluate complications and the long term effect on blood pressure. PMID- 10525520 TI - Utility of cardiac troponin I, creatine kinase-MB(mass), myosin light chain 1, and myoglobin in the early in-hospital triage of "high risk" patients with chest pain. AB - OBJECTIVE: To evaluate the use of cardiac troponin I (cTnI), creatine kinase MB(mass) (CK-MB(mass)), myosin light chain 1 (MLC 1), and myoglobin in identifying "high risk" patients with chest pain who will experience serious cardiac events (SCEs) in hospital. DESIGN: Prospective study. SETTING: University affiliated medical centre in Philadelphia, USA. PATIENTS: 208 patients with chest pain, at > 7% risk of acute myocardial infarction (MI), but without new ST segment elevation on their presenting ECG. INTERVENTIONS: cTnI, CK-MB(mass), MLC 1, and myoglobin concentrations were obtained on admission (0 hour) and at 4, 8, 16, and 24 hours. MAIN OUTCOME MEASURES: The sensitivity, specificity, positive and negative predictive value, and pre- and post-test probabilities of patients suffering an SCE in hospital were determined. SCEs included cardiac death, acute MI, cardiac arrest, life threatening cardiac arrhythmia, cardiogenic shock, and urgent coronary revascularisation. RESULTS: Admission concentrations of all markers were poor predictors of SCEs in hospital but improved substantially at subsequent timepoints. cTnI and CK-MB(mass) were consistently the most useful prognostic indicators. If both were negative at 0, 4, and 8 hours, then 99% (95% confidence interval 96% to 100%) of patients remained free from SCEs. The only SCEs not thus predicted were revascularisation procedures and associated complications. Additional tests after 8 hours, or the inclusion of additional markers, did not improve predictive accuracy further. CONCLUSIONS: Patients with high risk clinical features on admission who have negative cTnI and CK-MB(mass) concentrations at 0, 4, and 8 hours later have a favourable in-hospital prognosis and could be considered for early triage out of coronary care units. PMID- 10525521 TI - A new mutation of the cardiac troponin T gene causing familial hypertrophic cardiomyopathy without left ventricular hypertrophy. AB - AIM: To screen for a mutation of the cardiac troponin T gene in two families where there had been sudden deaths without an increase in left ventricular mass but with myocardial disarray suggesting hypertrophic cardiomyopathy. METHODS: DNA from affected individuals from both families was used to screen the cardiac troponin T gene on an exon by exon basis. Mutation screening was achieved by polymerase chain reaction and direct sequencing. Where appropriate, a mutation was confirmed by restriction digest. RESULTS: A novel missense mutation of exon 9 was found in the affected individuals of one of the families. This mutation at amino acid 94 resulted in the substitution of arginine for leucine and was not found in 100 normal control samples. A mutation of the cardiac troponin T gene was excluded in the second family. CONCLUSIONS: A mutation of the gene for the sarcomeric protein cardiac troponin T can cause familial hypertrophic cardiomyopathy with marked myocyte disarray and frequent premature sudden death in the absence of myocardial hypertrophy at clinical or macroscopic level. PMID- 10525522 TI - Malalignment of the sarcomeric filaments in hypertrophic cardiomyopathy with cardiac myosin heavy chain gene mutation. AB - OBJECTIVE: To investigate changes in the alignment of the sarcomeric filaments in hypertrophic cardiomyopathy and the effects of cardiac beta myosin heavy chain (beta-MHC) mutation on the sarcomeric ultrastructure. DESIGN: A retrospective analysis. PATIENTS: Endomyocardial biopsy samples were examined by transmission electron microscopy in seven patients with hypertrophic cardiomyopathy and beta MHC mutation, six with hypertrophic cardiomyopathy but without the mutation, and five controls (with chest pain syndromes). MAIN OUTCOME MEASURE: Alignment of the sarcomeric filaments and the distance between neighbouring thick myosin filaments. RESULTS: In controls, cross sections of the sarcomere at the A band showed a highly organised orthohexagonal array with 6 thin actin filaments surrounding one thick myosin filament, whereas in hypertrophic cardiomyopathy the alignment of the sarcomeric filaments was sparse and disrupted. In hypertrophic cardiomyopathy with a mutation, the distance between neighbouring thick myosin filaments was greater than in controls (mean (SD) 45.3 (4.7) v 38.5 (3.5) nm, p < 0.05), and the variance of the distance was greater than in controls (8.0 (0.7) v 4.8 (1.0) nm, p < 0.001) or in patients with hypertrophic cardiomyopathy without a mutation (6.7 (0.6) nm, p < 0.05). In the latter, the variance of the distance was also greater than in the controls (p < 0.01). A significant correlation was found between the grade of the myocyte hypertrophy and the variance of the distance (r = 0.654; p < 0.01). CONCLUSIONS: The alignment of the sarcomeric filaments is disrupted in hypertrophic cardiomyopathy, particularly when there is beta-MHC mutation. PMID- 10525523 TI - N-terminal proatrial natriuretic peptide correlates with systolic dysfunction and left ventricular filling pattern in patients with idiopathic dilated cardiomyopathy. AB - OBJECTIVE: To investigate the diastolic Doppler filling pattern in patients with idiopathic dilated cardiomyopathy and its relation to N-terminal pro-atrial natriuretic peptide (NT-pro-ANP). METHODS: 32 patients (26 male, six female) with idiopathic dilated cardiomyopathy were investigated. All were in sinus rhythm. Conventional M mode echocardiography and Doppler echocardiography was done in each patient. Pulsed wave Doppler inflow signals were obtained and the following variables were measured: maximum E wave, maximum A wave, E/A ratio, E wave deceleration time, A wave deceleration time. NT-pro-ANP was measured using radioimmunoassay. RESULTS: Mean (SD) left ventricular ejection fraction was 34 (7)% and mean left ventricular end diastolic diameter on M mode echocardiography was 69 (7) mm. Left ventricular filling indices were as follows: maximum E wave velocity, 0.86 (0.22) m/s; maximum A wave velocity, 0.71 (0.24) m/s; E/A ratio, 1.41 (0.65). Mean E wave deceleration time was 140 (50) ms; mean A wave deceleration time was 100 (20) ms. In a stepwise forward regression model, NT-pro ANP correlated significantly with left atrial diameter (r = 0.603; p < 0. 001), left ventricular ejection fraction (r = -0.758; p < 0.001), and Doppler derived E/A ratio (r = 0.740; p < 0.001). CONCLUSIONS: In patients with idiopathic dilated cardiomyopathy there is a relation between NT-pro-ANP and both systolic and diastolic variables. In a multivariate model NT-pro-ANP correlated with left atrial diameter, left ventricular ejection fraction, and Doppler derived E/A ratio on transmitral inflow. PMID- 10525525 TI - Mobile intracardiac calcinosis: a new risk of thromboembolism in patients with haemodialysed end stage renal disease. AB - Cardiac calcinosis is a common complication of end stage renal disease. A newly observed risk of thromboembolism is reported in four patients with mobile cardiac calcinosis, treated with long term dialysis. Rapidly growing mobile calcification was confirmed by echocardiography. Each patient had an imbalance in serum calcium x inorganic phosphate (Ca x P product >/= 50); this imbalance could not be treated due to the sudden death of the patient or the need for surgical resection to prevent recurrent cerebral thromboembolism. Histological examination revealed intracardiac calcinosis in three cases, and each case showed haemodialysis hypoparathyroidism (intact PTH < 160 pg/ml). Thromboembolism in such cases is rare, however it indicates a need for cautious echocardiographic monitoring in end stage renal disease in patients with an uncontrolled Ca x P product. PMID- 10525524 TI - Is it possible to identify infrahissian cardiac conduction abnormalities in myotonic dystrophy by non-invasive methods? AB - OBJECTIVE: To identify intracardiac conduction abnormalities in patients with myotonic dystrophy from their clinical, ECG, and genetic features. METHODS: 39 consecutive patients (mean (SD) age 42. 9 (12.1) years; 16 female, 23 male) underwent clinical examination, genetic studies, resting and 24 hour ambulatory ECG, signal averaged ECG, and electrophysiological studies. RESULTS: 23 patients suffered from cardiac symptoms, 23 had one or more cardiac conduction abnormality on resting ECG, one had sinus deficiency, and 21 (53.8%) had prolonged HV intervals. No correlation was found between the severity of the neurological symptoms, onset of disease, cardiac conduction abnormalities on ECG, and the intracardiac conduction abnormalities on electrophysiological study. The size of the DNA mutation was longer in the abnormal HV interval group than in the normal HV interval group (3.5 (1.8) v 2.2 (1.0) kb, p < 0.02). Signal averaged ECG parameters (total QRS duration (QRSD) and duration of low amplitude signals /= 100 ms with LAS 40 >/= 36 ms identified patients with an abnormal HV interval with good sensitivity (80%) and specificity (83. 3%). CONCLUSIONS: Infrahissian conduction abnormalities are common in myotonic dystrophy and can be identified using signal averaged electrocardiography. PMID- 10525526 TI - Carcinoid constrictive pericarditis. AB - A 78 year old man presented with diarrhoea, anorexia, and progressive lower limb oedema. He was in atrial fibrillation and had a right pleural effusion and ascites. Ultrasound of the abdomen and 24 hour urinary hydroxyindoleacetic acid output indicated metastatic carcinoid syndrome. Cardiac catheterisation revealed pericardial constriction, and pericardial exploration showed a greatly thickened pericardium with no evidence of tumour invasion. The patient died within 24 hours of surgery. Necropsy findings were consistent with a diagnosis of constrictive pericarditis secondary to metastatic carcinoid syndrome. PMID- 10525527 TI - Unusual congenital coronary anomaly and myocardial ischaemia. AB - Angiography was used to diagnose a rare congenital coronary anomaly with myocardial ischaemia in a woman with typical angina. All three coronary arteries arose from a solitary coronary ostium in the right aortic sinus; the left anterior descending coronary artery followed a septal course, the circumflex coronary artery ran behind the ascending aorta, and the right coronary artery followed a normal course. No significant coronary lumen narrowing was found. Transoesophageal echocardiography confirmed the anomalous origin and course of the aberrant coronary arteries. An exercise test reproduced angina, and ECG changes and myocardial perfusion study showed an anterior reversible defect. In contrast to previous reports, myocardial ischaemia was associated with the septal (intramuscular) course of the left anterior descending coronary artery; there was no other significant coronary artery disease. PMID- 10525528 TI - Nicorandil abolished repolarisation alternans in a patient with idiopathic long QT syndrome. AB - A 23 year old woman with idiopathic long QT syndrome had repeated syncopal attacks associated with torsades de pointes. T wave alternans (TWA) was recorded and the QT interval was abnormally prolonged during treadmill exercise test. Monophasic action potential (MAP) alternans also appeared after an abrupt shortening of the cycle length in electrophysiological study. After intravenous administration of nicorandil 6 mg, both TWA and MAP alternans disappeared. PMID- 10525529 TI - RAD51 and DMC1 form mixed complexes associated with mouse meiotic chromosome cores and synaptonemal complexes. AB - The eukaryotic RecA homologues RAD51 and DMC1 function in homology recognition and formation of joint-molecule recombination intermediates during yeast meiosis. The precise immunolocalization of these two proteins on the meiotic chromosomes of plants and animals has been complicated by their high degree of identity at the amino acid level. With antibodies that have been immunodepleted of cross reactive epitopes, we demonstrate that RAD51 and DMC1 have identical distribution patterns in extracts of mouse spermatocytes in successive prophase I stages, suggesting coordinate functionality. Immunofluorescence and immunoelectron microscopy with these antibodies demonstrate colocalization of the two proteins on the meiotic chromosome cores at early prophase I. We also show that mouse RAD51 and DMC1 establish protein-protein interactions with each other and with the chromosome core component COR1(SCP3) in a two-hybrid system and in vitro binding analyses. These results suggest that the formation of a multiprotein recombination complex associated with the meiotic chromosome cores is essential for the development and fulfillment of the meiotic recombination process. PMID- 10525530 TI - PML is critical for ND10 formation and recruits the PML-interacting protein daxx to this nuclear structure when modified by SUMO-1. AB - Nuclear domain 10 (ND10), also referred to as nuclear bodies, are discrete interchromosomal accumulations of several proteins including promyelocytic leukemia protein (PML) and Sp100. In this study, we investigated the mechanism of ND10 assembly by identifying proteins that are essential for this process using cells lines that lack individual ND10-associated proteins. We identified the adapter protein Daxx and BML, the RecQ helicase missing in Bloom syndrome, as new ND10-associated proteins. PML, but not BLM or Sp100, was found to be responsible for the proper localization of all other ND10-associated proteins since they are dispersed in PML-/- cells. Introducing PML into this cell line by transient expression or fusion with PML-producing cells recruited ND10-associated proteins into de novo formed ND10 attesting to PMLs essential nature in ND10 formation. In the absence of PML, Daxx is highly enriched in condensed chromatin. Its recruitment to ND10 from condensed chromatin requires a small ubiquitin-related modifier (SUMO-1) modification of PML and reflects the interaction between the COOH-terminal domain of Daxx and PML. The segregation of Daxx from condensed chromatin in the absence of PML to ND10 by increased accumulation of SUMO-1 modified PML suggests the presence of a variable equilibrium between these two nuclear sites. Our findings identify the basic requirements for ND10 formation and suggest a dynamic mechanism for protein recruitment to these nuclear domains controlled by the SUMO-1 modification state of PML. PMID- 10525531 TI - The karyopherin Kap122p/Pdr6p imports both subunits of the transcription factor IIA into the nucleus. AB - We discovered a nuclear import pathway mediated by the product of the previously identified Saccharomyces cerevisiae gene PDR6 (pleiotropic drug resistance). This gene product functions as a karyopherin (Kap) for nuclear import. Consistent with previously proposed nomenclature, we have renamed this gene KAP122. Kap122p was localized both to the cytoplasm and the nucleus. As a prominent import substrate of Kap122p, we identified the complex of the large and small subunit (Toa1p and Toa2p, respectively) of the general transcription factor IIA (TFIIA). Recombinant GST-Kap122p formed a complex with recombinant His(6)-Toa1p/Toa2p. In wild-type cells, Toa1p and Toa2p were localized to the nucleus. Consistent with Kap122p being the principal Kap for import of the Toa1p-Toa2p complex, we found that deletion of KAP122 results in increased cytoplasmic localization of both Toa1p and Toa2p. Deletion of KAP122 is not lethal, although deletion of TOA1 and TOA2 is. Together these data suggest that Kap122p is the major Kap for the import of Toa1p-Toa2p into the nucleus. Like other substrate-Kap complexes, the Toa1p/Toa2p/Kap122p complex isolated from yeast cytosol or reconstituted from recombinant proteins, was dissociated by RanGTP but not RanGDP. Kap122p bound to nucleoporins, specifically, to the peptide repeat-containing fragments of Nup1p and Nup2p. PMID- 10525532 TI - The cis-acting RNA trafficking signal from myelin basic protein mRNA and its cognate trans-acting ligand hnRNP A2 enhance cap-dependent translation. AB - The 21 nucleotide RNA trafficking signal (RTS), originally identified in myelin basic protein mRNA, but also found in a variety of other localized RNAs, is necessary and sufficient for transport of RNA along microtubules in oligodendrocytes. The RTS binds specifically to the RNA binding protein, hnRNP A2. Together, the RTS and hnRNP A2 comprise cis/trans determinants for several steps in the RNA trafficking pathway. Here we show that insertion of the RTS into green fluorescent protein (GFP) RNA enhances translation without affecting stability of microinjected RNA. In dicistronic RNA, the RTS enhances cap dependent translation without affecting internal ribosome entry site (IRES) dependent translation. The translation enhancer function of the RTS is position, copy number, and cell type independent, hnRNP A2 dependent, and saturable with increasing amounts of injected RNA. This represents one of the first specific translation enhancer elements identified in a mammalian system. PMID- 10525533 TI - Glycosylation can influence topogenesis of membrane proteins and reveals dynamic reorientation of nascent polypeptides within the translocon. AB - The topology of multispanning membrane proteins in the mammalian endoplasmic reticulum is thought to be dictated primarily by the first hydrophobic sequence. We analyzed the in vivo insertion of a series of chimeric model proteins containing two conflicting signal sequences, i.e., an NH(2)-terminal and an internal signal, each of which normally directs translocation of its COOH terminal end. When the signals were separated by more than 60 residues, linear insertion with the second signal acting as a stop-transfer sequence was observed. With shorter spacers, an increasing fraction of proteins inserted with a translocated COOH terminus as dictated by the second signal. Whether this resulted from membrane targeting via the second signal was tested by measuring the targeting efficiency of NH(2)-terminal signals followed by polypeptides of different lengths. The results show that targeting is mediated predominantly by the first signal in a protein. Most importantly, we discovered that glycosylation within the spacer sequence affects protein orientation. This indicates that the nascent polypeptide can reorient within the translocation machinery, a process that is blocked by glycosylation. Thus, topogenesis of membrane proteins is a dynamic process in which topogenic information of closely spaced signal and transmembrane sequences is integrated. PMID- 10525534 TI - The maize tha4 gene functions in sec-independent protein transport in chloroplasts and is related to hcf106, tatA, and tatB. AB - Proteins are translocated across the chloroplast thylakoid membrane by a variety of mechanisms. Some proteins engage a translocation machinery that is derived from the bacterial Sec export system and require an interaction with a chloroplast-localized SecA homologue. Other proteins engage a machinery that is SecA-independent, but requires a transmembrane pH gradient. Recently, a counterpart to this Delta pH mechanism was discovered in bacteria. Genetic studies revealed that one maize protein involved in this mechanism, HCF106, is related in both structure and function to the bacterial tatA and tatB gene products. We describe here the mutant phenotype and molecular cloning of a second maize gene that functions in the Delta pH mechanism. This gene, thylakoid assembly 4 (tha4), is required specifically for the translocation of proteins that engage the Delta pH pathway. The sequence of the tha4 gene product resembles those of the maize hcf106 gene and the bacterial tatA and tatB genes. Sequence comparisons suggest that tha4 more closely resembles tatA, and hcf106 more closely resembles tatB. These findings support the notion that this sec independent translocation mechanism has been highly conserved during the evolution of eucaryotic organelles from bacterial endosymbionts. PMID- 10525535 TI - Presenilin 1 controls gamma-secretase processing of amyloid precursor protein in pre-golgi compartments of hippocampal neurons. AB - Mutations of presenilin 1 (PS1) causing Alzheimer's disease selectively increase the secretion of the amyloidogenic betaA4(1-42), whereas knocking out the gene results in decreased production of both betaA4(1-40) and (1-42) amyloid peptides (De Strooper et al. 1998). Therefore, PS1 function is closely linked to the gamma secretase processing of the amyloid precursor protein (APP). Given the ongoing controversy on the subcellular localization of PS1, it remains unclear at what level of the secretory and endocytic pathways PS1 exerts its activity on APP and on the APP carboxy-terminal fragments that are the direct substrates for gamma secretase. Therefore, we have reinvestigated the subcellular localization of endogenously expressed PS1 in neurons in vitro and in vivo using confocal microscopy and fine-tuned subcellular fractionation. We show that uncleaved PS1 holoprotein is recovered in the nuclear envelope fraction, whereas the cleaved PS fragments are found mainly in post-ER membranes including the intermediate compartment (IC). PS1 is concentrated in discrete sec23p- and p58/ERGIC-53 positive patches, suggesting its localization in subdomains involved in ER export. PS1 is not found to significant amounts beyond the cis-Golgi. Surprisingly, we found that APP carboxy-terminal fragments also coenrich in the pre-Golgi membrane fractions, consistent with the idea that these fragments are the real substrates for gamma-secretase. Functional evidence that PS1 exerts its effects on gamma-secretase processing of APP in the ER/IC was obtained using a series of APP trafficking mutants. These mutants were investigated in hippocampal neurons derived from transgenic mice expressing PS1wt or PS1 containing clinical mutations (PS1(M146L) and PS1(L286V)) at physiologically relevant levels. We demonstrate that the APP-London and PS1 mutations have additive effects on the increased secretion of betaA4(1-42) relative to betaA4(1-40), indicating that both mutations operate independently. Overall, our data clearly establish that PS1 controls gamma(42)-secretase activity in pre-Golgi compartments. We discuss models that reconcile this conclusion with the effects of PS1 deficiency on the generation of betaA4(1-40) peptide in the late biosynthetic and endocytic pathways. PMID- 10525536 TI - Human cyclin A is required for mitosis until mid prophase. AB - We have used microinjection and time-lapse video microscopy to study the role of cyclin A in mitosis. We have injected purified, active cyclin A/cyclin-dependent kinase 2 (CDK2) into synchronized cells at specific points in the cell cycle and assayed its effect on cell division. We find that cyclin A/CDK2 will drive G2 phase cells into mitosis within 30 min of microinjection, up to 4 h before control cells enter mitosis. Often this premature mitosis is abnormal; the chromosomes do not completely condense and daughter cells fuse. Remarkably, microinjecting cyclin A/CDK2 into S phase cells has no effect on progress through the following G2 phase or mitosis. In complementary experiments we have microinjected the amino terminus of p21(Cip1/Waf1/Sdi1) (p21N) into cells to inhibit cyclin A/CDK2 activity. We find that p21N will prevent S phase or G2 phase cells from entering mitosis, and will cause early prophase cells to return to interphase. These results suggest that cyclin A/CDK2 is a rate-limiting component required for entry into mitosis, and for progress through mitosis until late prophase. They also suggest that cyclin A/CDK2 may be the target of the recently described prophase checkpoint. PMID- 10525538 TI - Dynactin is required for microtubule anchoring at centrosomes. AB - The multiprotein complex, dynactin, is an integral part of the cytoplasmic dynein motor and is required for dynein-based motility in vitro and in vivo. In living cells, perturbation of the dynein-dynactin interaction profoundly blocks mitotic spindle assembly, and inhibition or depletion of dynein or dynactin from meiotic or mitotic cell extracts prevents microtubules from focusing into spindles. In interphase cells, perturbation of the dynein-dynactin complex is correlated with an inhibition of ER-to-Golgi movement and reorganization of the Golgi apparatus and the endosome-lysosome system, but the effects on microtubule organization have not previously been defined. To explore this question, we overexpressed a variety of dynactin subunits in cultured fibroblasts. Subunits implicated in dynein binding have effects on both microtubule organization and centrosome integrity. Microtubules are reorganized into unfocused arrays. The pericentriolar components, gamma tubulin and dynactin, are lost from centrosomes, but pericentrin localization persists. Microtubule nucleation from centrosomes proceeds relatively normally, but microtubules become disorganized soon thereafter. Overexpression of some, but not all, dynactin subunits also affects endomembrane localization. These data indicate that dynein and dynactin play important roles in microtubule organization at centrosomes in fibroblastic cells and provide new insights into dynactin-cargo interactions. PMID- 10525537 TI - Analysis of dynactin subcomplexes reveals a novel actin-related protein associated with the arp1 minifilament pointed end. AB - The multisubunit protein, dynactin, is a critical component of the cytoplasmic dynein motor machinery. Dynactin contains two distinct structural domains: a projecting sidearm that interacts with dynein and an actin-like minifilament backbone that is thought to bind cargo. Here, we use biochemical, ultrastructural, and molecular cloning techniques to obtain a comprehensive picture of dynactin composition and structure. Treatment of purified dynactin with recombinant dynamitin yields two assemblies: the actin-related protein, Arp1, minifilament and the p150(Glued) sidearm. Both contain dynamitin. Treatment of dynactin with the chaotropic salt, potassium iodide, completely depolymerizes the Arp1 minifilament to reveal multiple protein complexes that contain the remaining dynactin subunits. The shoulder/sidearm complex contains p150(Glued), dynamitin, and p24 subunits and is ultrastructurally similar to dynactin's flexible projecting sidearm. The dynactin shoulder complex, which contains dynamitin and p24, is an elongated, flexible assembly that may link the shoulder/sidearm complex to the Arp1 minifilament. Pointed-end complex contains p62, p27, and p25 subunits, plus a novel actin-related protein, Arp11. p62, p27, and p25 contain predicted cargo-binding motifs, while the Arp11 sequence suggests a pointed-end capping activity. These isolated dynactin subdomains will be useful tools for further analysis of dynactin assembly and function. PMID- 10525539 TI - Novel roles for saccharomyces cerevisiae mitotic spindle motors. AB - The single cytoplasmic dynein and five of the six kinesin-related proteins encoded by Saccharomyces cerevisiae participate in mitotic spindle function. Some of the motors operate within the nucleus to assemble and elongate the bipolar spindle. Others operate on the cytoplasmic microtubules to effect spindle and nuclear positioning within the cell. This study reveals that kinesin-related Kar3p and Kip3p are unique in that they perform roles both inside and outside the nucleus. Kar3p, like Kip3p, was found to be required for spindle positioning in the absence of dynein. The spindle positioning role of Kar3p is performed in concert with the Cik1p accessory factor, but not the homologous Vik1p. Kar3p and Kip3p were also found to overlap for a function essential for the structural integrity of the bipolar spindle. The cytoplasmic and nuclear roles of both these motors could be partially substituted for by the microtubule-destabilizing agent benomyl, suggesting that these motors perform an essential microtubule destabilizing function. In addition, we found that yeast cell viability could be supported by as few as two microtubule-based motors: the BimC-type kinesin Cin8p, required for spindle structure, paired with either Kar3p or Kip3p, required for both spindle structure and positioning. PMID- 10525541 TI - A cell-free assay system for beta-catenin signaling that recapitulates direct inductive events in the early xenopus laevis embryo. AB - In vertebrate embryos, signaling via the beta-catenin protein is known to play an essential role in the induction of the dorsal axis. In its signaling capacity, beta-catenin acts directly to affect target gene transcription, in concert with transcription factors of the TCF/LEF family. We have developed a cell-free in vitro assay for beta-catenin signaling activity that utilizes transcriptionally active nuclei and cytoplasm from cleavage-blocked Xenopus laevis embryos. Under these assay conditions, we demonstrate that either addition of beta-catenin protein or upstream activation of the beta-catenin signaling pathway can induce the expression of developmentally relevant target genes. Addition of exogenous beta-catenin protein induced expression of Siamois, XTwin, Xnr3, and Cerberus mRNAs in a protein synthesis independent manner, whereas a panel of other Spemann organizer-specific genes did not respond to beta-catenin. Lithium induction of the beta-catenin signaling pathway, which is thought to cause beta-catenin accumulation by inhibiting its proteasome-dependent degradation, caused increased expression of Siamois in a protein synthesis independent fashion. This result suggests that beta-catenin derived from a preexisting pool can be activated to signal, and that accumulation of this activated form does not require ongoing synthesis. Furthermore, activation of the signaling pathway with lithium did not detectably alter cytoplasmic beta-catenin levels and was insensitive to inhibition of the proteasome- dependent degradation pathway. Taken together, these results suggest that activation of beta-catenin signaling by lithium in this system may occur through a distinct activation mechanism that does not require modulation of levels through regulation of proteasomal degradation. PMID- 10525540 TI - The kinesin-related protein, HSET, opposes the activity of Eg5 and cross-links microtubules in the mammalian mitotic spindle. AB - We have prepared antibodies specific for HSET, the human homologue of the KAR3 family of minus end-directed motors. Immuno-EM with these antibodies indicates that HSET frequently localizes between microtubules within the mammalian metaphase spindle consistent with a microtubule cross-linking function. Microinjection experiments show that HSET activity is essential for meiotic spindle organization in murine oocytes and taxol-induced aster assembly in cultured cells. However, inhibition of HSET did not affect mitotic spindle architecture or function in cultured cells, indicating that centrosomes mask the role of HSET during mitosis. We also show that (acentrosomal) microtubule asters fail to assemble in vitro without HSET activity, but simultaneous inhibition of HSET and Eg5, a plus end-directed motor, redresses the balance of forces acting on microtubules and restores aster organization. In vivo, centrosomes fail to separate and monopolar spindles assemble without Eg5 activity. Simultaneous inhibition of HSET and Eg5 restores centrosome separation and, in some cases, bipolar spindle formation. Thus, through microtubule cross-linking and oppositely oriented motor activity, HSET and Eg5 participate in spindle assembly and promote spindle bipolarity, although the activity of HSET is not essential for spindle assembly and function in cultured cells because of centrosomes. PMID- 10525542 TI - The receptor tyrosine phosphatase CRYPalpha promotes intraretinal axon growth. AB - Retinal ganglion cell axons grow towards the optic fissure in close contact with the basal membrane, an excellent growth substratum. One of the ligands of receptor tyrosine phosphatase CRYPalpha is located on the retinal and tectal basal membranes. To analyze the role of this RPTP and its ligand in intraretinal growth and guidance of ganglion cell axons, we disrupted ligand- receptor interactions on the retinal basal membrane in culture. Antibodies against CRYPalpha strongly reduced retinal axon growth on the basal membrane, and induced a dramatic change in morphology of retinal growth cones, reducing the size of growth cone lamellipodia. A similar effect was observed by blocking the ligand with a CRYPalpha ectodomain fusion protein. These effects did not occur, or were much reduced, when axons were grown either on laminin-1, on matrigel or on basal membranes with glial endfeet removed. This indicates that a ligand for CRYPalpha is located on glial endfeet. These results show for the first time in vertebrates that the interaction of a receptor tyrosine phosphatase with its ligand is crucial not only for promotion of retinal axon growth but also for maintenance of retinal growth cone lamellipodia on basal membranes. PMID- 10525543 TI - Integrin-associated protein stimulates alpha2beta1-dependent chemotaxis via Gi mediated inhibition of adenylate cyclase and extracellular-regulated kinases. AB - Integrin-associated protein (IAP/CD47) augments the function of alpha2beta1 integrin in smooth muscle cells (SMC), resulting in enhanced chemotaxis toward soluble collagen (Wang, X-Q., and W.A. Frazier. 1998. Mol. Biol. Cell. 9:865). IAP-deficient SMC derived from IAP(-/-) animals did not migrate in response to 4N1K (KRFYVVMWKK), a peptide agonist of IAP derived from the COOH-terminal domain of thrombospondin-1 (TSP1). When normal SMC were preincubated with 4N1K or an anti-alpha2beta1 function-stimulating antibody, cell migration to soluble collagen was significantly enhanced. 4N1K-induced chemotaxis was blocked by treatment of SMC with pertussis toxin indicating that IAP acts through Gi. In agreement with this, 4N1K evoked a rapid decrease in cAMP levels which was intensified in the presence of collagen, and forskolin and 8-Br-cAMP both inhibited SMC migration stimulated via IAP. 4N1K strongly inhibited extracellular regulated kinase (ERK) activation in SMC attaching to collagen and reduced basal ERK activity in suspended SMC. Pertussis toxin treatment of SMC significantly activated ERK, suggesting that an inhibitory input was alleviated. Inhibition of ERK activity by (a) the MAP kinase kinase (MEK) inhibitor, PD98059, (b) antisense oligonucleotide depletion of ERK, and (c) expression of mitogen-activated protein (MAP) kinase phosphatase-1 in SMC all led to increased migration to collagen, 4N1K, or 4N1K plus collagen. Thus, IAP stimulates alpha2beta1 integrin-mediated SMC migration via Gi-mediated inhibition of ERK activity and suppression of cyclic AMP levels. Both of these signaling pathways could directly modulate the state of the integrin as well as impact downstream components of the cell motility apparatus. PMID- 10525544 TI - Integrin alpha2beta1 mediates isoform-specific activation of p38 and upregulation of collagen gene transcription by a mechanism involving the alpha2 cytoplasmic tail. AB - Two collagen receptors, integrins alpha1beta1 and alpha2beta1, can regulate distinct functions in cells. Ligation of alpha1beta1, unlike alpha2beta1, has been shown to result in recruitment of Shc and activation of the Ras/ERK pathway. To identify the downstream signaling molecules activated by alpha2beta1 integrin, we have overexpressed wild-type alpha2, or chimeric alpha2 subunit with alpha1 integrin cytoplasmic domain in human osteosarcoma cells (Saos-2) lacking endogenous alpha2beta1. The chimeric alpha2/alpha1 chain formed a functional heterodimer with beta1. In contrast to alpha2/alpha1 chimera, forced expression of alpha2 integrin resulted in upregulation of alpha1 (I) collagen gene transcription in response to three-dimensional collagen, indicating that the cytoplasmic domain of alpha2 integrin was required for signaling. Furthermore, signals mediated by alpha2beta1 integrin specifically activated the p38alpha isoform, and selective p38 inhibitors blocked upregulation of collagen gene transcription. Dominant negative mutants of Cdc42, MKK3, and MKK4 prevented alpha2beta1 integrin-mediated activation of p38alpha. RhoA had also some inhibitory effect, whereas dominant negative Rac was not effective. Our findings show the isoform-specific activation of p38 by alpha2beta1 integrin ligation and identify Cdc42, MKK3, and MKK4 as possible downstream effectors. These observations reveal a novel signaling mechanism of alpha2beta1 integrin that is distinct from ones previously described for other integrins. PMID- 10525545 TI - Binding of integrin alpha6beta4 to plectin prevents plectin association with F actin but does not interfere with intermediate filament binding. AB - Hemidesmosomes are stable adhesion complexes in basal epithelial cells that provide a link between the intermediate filament network and the extracellular matrix. We have investigated the recruitment of plectin into hemidesmosomes by the alpha6beta4 integrin and have shown that the cytoplasmic domain of the beta4 subunit associates with an NH(2)-terminal fragment of plectin that contains the actin-binding domain (ABD). When expressed in immortalized plectin-deficient keratinocytes from human patients with epidermol- ysis bullosa (EB) simplex with muscular dystrophy (MD-EBS), this fragment is colocalized with alpha6beta4 in basal hemidesmosome-like clusters or associated with F-actin in stress fibers or focal contacts. We used a yeast two-hybrid binding assay in combination with an in vitro dot blot overlay assay to demonstrate that beta4 interacts directly with plectin, and identified a major plectin-binding site on the second fibronectin type III repeat of the beta4 cytoplasmic domain. Mapping of the beta4 and actin binding sites on plectin showed that the binding sites overlap and are both located in the plectin ABD. Using an in vitro competition assay, we could show that beta4 can compete out the plectin ABD fragment from its association with F actin. The ability of beta4 to prevent binding of F-actin to plectin explains why F-actin has never been found in association with hemidesmosomes, and provides a molecular mechanism for a switch in plectin localization from actin filaments to basal intermediate filament-anchoring hemidesmosomes when beta4 is expressed. Finally, by mapping of the COOH-terminally located binding site for several different intermediate filament proteins on plectin using yeast two-hybrid assays and cell transfection experiments with MD-EBS keratinocytes, we confirm that plectin interacts with different cytoskeletal networks. PMID- 10525546 TI - Formation process of autophagosome is traced with Apg8/Aut7p in yeast. AB - We characterized Apg8/Aut7p essential for autophagy in yeast. Apg8p was transcriptionally upregulated in response to starvation and mostly existed as a protein bound to membrane under both growing and starvation conditions. Immunofluorescence microscopy revealed that the intracellular localization of Apg8p changed drastically after shift to starvation. Apg8p resided on unidentified tiny dot structures dispersed in the cytoplasm at growing phase. During starvation, it was localized on large punctate structures, some of which were confirmed to be autophagosomes and autophagic bodies by immuno-EM. Besides these structures, we found that Apg8p was enriched on isolation membranes and in electron less-dense regions, which should contain Apg8p-localized membrane- or lipid-containing structures. These structures would represent intermediate structures during autophagosome formation. Here, we also showed that microtubule does not play an essential role in the autophagy in yeast. The result does not match with the previously proposed role of Apg8/Aut7p, delivery of autophagosome to the vacuole along microtubule. Moreover, it is revealed that autophagosome formation is severely impaired in the apg8 null mutant. Apg8p would play an important role in the autophagosome formation. PMID- 10525548 TI - Bone mineral density in adults with cystic fibrosis. PMID- 10525547 TI - Aggregation of lipid rafts accompanies signaling via the T cell antigen receptor. AB - The role of lipid rafts in T cell antigen receptor (TCR) signaling was investigated using fluorescence microscopy. Lipid rafts labeled with cholera toxin B subunit (CT-B) and cross-linked into patches displayed characteristics of rafts isolated biochemically, including detergent resistance and colocalization with raft-associated proteins. LCK, LAT, and the TCR all colocalized with lipid patches, although TCR association was sensitive to nonionic detergent. Aggregation of the TCR by anti-CD3 mAb cross-linking also caused coaggregation of raft-associated proteins. However, the protein tyrosine phosphatase CD45 did not colocalize to either CT-B or CD3 patches. Cross-linking of either CD3 or CT-B strongly induced tyrosine phosphorylation and recruitment of a ZAP-70(SH2)(2) green fluorescent protein (GFP) fusion protein to the lipid patches. Also, CT-B patching induced signaling events analagous to TCR stimulation, with the same dependence on expression of key TCR signaling molecules. Targeting of LCK to rafts was necessary for these events, as a nonraft- associated transmembrane LCK chimera, which did not colocalize with TCR patches, could not reconstitute CT-B induced signaling. Thus, our results indicate a mechanism whereby TCR engagement promotes aggregation of lipid rafts, which facilitates colocalization of LCK, LAT, and the TCR whilst excluding CD45, thereby triggering protein tyrosine phosphorylation. PMID- 10525549 TI - Sleep on the cheap: the role of overnight oximetry in the diagnosis of sleep apnoea hypopnoea syndrome. PMID- 10525550 TI - Inducible nitric oxide and pulmonary infection. PMID- 10525551 TI - Impact factors for 1998 PMID- 10525552 TI - Low bone mineral density in adults with cystic fibrosis. AB - BACKGROUND: Patients with cystic fibrosis have several risk factors for the development of low bone mineral density (BMD). To identify the prevalence and clinical correlates of low BMD in adult patients with cystic fibrosis, densitometry was performed in 151 patients (83 men) aged 15-52 years. METHODS: BMD was measured in the lumbar spine (L1-4) using dual energy x ray absorptiometry (DXA) and quantitative computed tomography (QCT). It was also measured in the proximal femur (total hip and femoral neck) using DXA, and in the distal and ultra distal forearm using single energy x ray absorptiometry (SXA). Biochemical markers of bone turnover, vitamin D levels, parathyroid hormone levels, and a variety of anthropometric variables were also assessed. RESULTS: The mean (SD) BMD Z score was -0.73 (0.85) in the distal forearm, -0.31 (0.92) in the ultra distal forearm, -1.21 (1. 18) in the lumbar spine using DXA, -0.56 (1.36) in the lumbar spine using QCT, -1.25 (1.30) in the femoral neck, and -1.01 (1.14) in the total hip. 34% of patients had a BMD Z score of -2 or less at one or more skeletal sites. Body mass index (0.527, p = 0.01), percentage predicted forced expiratory volume in one second (0.388, p = 0.01), and physical activity (0.249, p = 0.05) were positively related to the mean BMD Z score. Levels of C reactive protein (-0.328, p = 0. 01), parathyroid hormone (-0.311, p = 0.01) and biochemical markers of bone turnover (osteocalcin -0.261 and bone specific alkaline phosphatase -0.249, p = 0.05) were negatively related to the mean BMD Z score. Vitamin D insufficiency (25-hydroxyvitamin D <15 ng/ml) was prevalent (53/139 patients, 38%) despite supplementation with 900 IU vitamin D per day. CONCLUSIONS: Low bone density is prevalent in adult patients with cystic fibrosis. Current levels of vitamin D supplementation appear to be inadequate. PMID- 10525553 TI - Nocturnal oximetry for the diagnosis of the sleep apnoea hypopnoea syndrome: a method to reduce the number of polysomnographies? AB - BACKGROUND: Polysomnography (PSG) is currently the "gold standard" for the diagnosis of the sleep apnoea hypopnoea syndrome (SAHS). Nocturnal oximetry (NO) has been used with contradictory results. A prospective study was performed to determine the accuracy of NO as a diagnostic tool and to evaluate the reduction in the number of PSGs if the diagnosis of SAHS had been established by this method. METHODS: Two hundred and seventy five patients with a clinical suspicion of SAHS were admitted to undergo, in the same night, full PSG and NO. Desaturation was defined as a fall in the haemoglobin saturation level (SaO(2)) to lower than 4% from the baseline level and an oxygen desaturation index per hour (ODI) was obtained in each patient with three cut off points: >/= 5 (ODI-5), >/= 10 (ODI-10), and >/= 15 (ODI-15). RESULTS: SAHS was diagnosed in 216 patients (194 men). After withdrawing patients with abnormal lung function (forced expiratory volume in one second (FEV(1)) lower than 80% predicted), sensitivity (SE), specificity (SP), positive and negative predictive values (PPV and NPV) of NO were: ODI-5 (80%, 89%, 97%, 48%); ODI-10 (71%, 93%, 97%, 42%); ODI-15 (63%, 96%, 99%, 38%). The accuracy for each ODI was 0.81, 0.75, and 0.70, respectively. If NO had been considered as a diagnostic tool and PSG had been performed only in patients with a negative NO (false negative and true negative) and those with a positive NO and abnormal pulmonary function tests, 135/275 (ODI-5), 156/275 (ODI 10), and 170/275 (ODI-15) PSGs would have been performed, a reduction of 140, 119, and 105, respectively. CONCLUSION: Nocturnal oximetry in patients with suspected SAHS and normal spirometric values permits the institution of therapeutic measures in most patients. PMID- 10525554 TI - Mandibular advancement oral appliance therapy for obstructive sleep apnoea: effect on awake calibre of the velopharynx. AB - BACKGROUND: The mechanisms of action of oral appliance therapy in obstructive sleep apnoea are poorly understood. Videoendoscopy of the upper airway was used during wakefulness to examine whether the changes in pharyngeal dimensions produced by a mandibular advancement oral appliance are related to the improvement in the severity of obstructive sleep apnoea. METHODS: Fifteen patients with mild to moderate obstructive sleep apnoea (median (range) apnoea index (AI) 4(0-38)/h, apnoea-hypopnoea index (AHI) 28(9-45)/h) underwent overnight polysomnography and imaging of the upper airway before and after insertion of the oral appliance. Images were obtained in the hypopharynx, oropharynx, and velopharynx at end tidal expiration during quiet nasal breathing in the supine position. The cross sectional area and diameters of the upper airway were measured using image processing software with an intraluminal catheter as a linear calibration. RESULTS: AI decreased to a median (range) value of 0 (0-6)/h (p<0.01) and AHI to 8 (1-28)/h (p<0.001) following insertion of the oral appliance. The median (95% confidence interval) cross sectional area of the upper airway increased by 18% (3 to 35) (p<0.02) in the hypopharynx and by 25% (11 to 69) (p<0.005) in the velopharynx, but not significantly in the oropharynx. Although in general the shape of the pharynx did not change following insertion of the oral appliance, the lateral diameter of the velopharynx increased to a greater extent than the anteroposterior diameter. Following insertion of the oral appliance the reduction in AHI was related to the increase in cross sectional area of the velopharynx (p = 0.01). CONCLUSIONS: A mandibular advancement oral appliance increases the cross sectional area of the upper airway during wakefulness, particularly in the velopharynx. Assuming this effect on upper airway calibre is not eliminated by sleep, mandibular advancement oral appliances may reduce the severity of obstructive sleep apnoea by maintaining patency of the velopharynx, particularly in its lateral dimension. PMID- 10525555 TI - Scottish Confidential Inquiry into Asthma Deaths (SCIAD), 1994-6. AB - BACKGROUND: There have been important changes in the organisation of care for patients with asthma since asthma deaths were studied in the 1980s by the British Thoracic Association (BTA), with greater emphasis on long term control of symptoms and the use of preventive therapy. Recent trends in routine statistics show a decline in population death rates. METHODS: A confidential review was undertaken of general practice and hospital records and interviews with general practitioners of patients dying in mainland Scotland between January 1994 and December 1996 with a principal diagnosis of asthma recorded by the Registrar General's Office. Panel assessment of the cause of death was carried out and a number of possible adverse factors were identified. The data from the 15-64 year age group were compared with similar data from the earlier study by the BTA. RESULTS: Over the three year period 95 deaths of 235 studied (40%) were confirmed as being due to asthma. Taking account of different methods of case ascertainment used in the BTA and this study, a fall in the calculated rate of "deaths assessed as due to asthma" was found from 2.51 (95% CI 2.34 to 2.68) per 100,000 population in 1979 to 1.26 (95% CI 1.19 to 1.33) per 100,000 population in 1994 6. Fewer individual adverse factors were identified in clinical management, with appropriate routine management in 59% and management of the final attack satisfactory in 71%. Patient factors such as poor compliance, lack of peak expiratory flow (PEF) measurements, and overuse of reliever medication without inhaled corticosteroids, and psychosocial problems, notably depression, were confirmed as important contributing factors. Four of five patients under 16 years of age who died were found to have problems with routine management. CONCLUSIONS: This population based study documents important improvements in the standard of asthma care as well as a significant decline in the rate of deaths due to asthma over a period during which the organisation of care has changed and the chronic nature of the disease has been acknowledged. Strategies which might have a further impact include the greater use of PEF recordings, particularly during acute attacks, to document recovery, prescription monitoring of the underuse of inhaled corticosteroids, consideration of the use of combined preparations where persistent overuse of bronchodilators is occurring, and increased input for young patients whose routine management is proving difficult. PMID- 10525556 TI - A confidential inquiry into asthma deaths in Wales. AB - BACKGROUND: Death from asthma is regarded as preventable in principle, especially under the age of 65 years. METHODS: In 1994 a confidential inquiry was set up to investigate deaths attributable to asthma in residents of Wales under the age of 65 years. During the period of the inquiry 92 cases were notified as being ascribed to asthma, or (in 1996) having a mention of asthma on the death certificate. Of these, 80 were investigated further with the help of general practitioners, hospital notes, and relatives. The details were then considered by a small panel of doctors who endeavoured to identify factors that may have contributed to the deaths. RESULTS: Asthma was considered to be the underlying cause of 52 deaths. Although disease severity was usually a major factor, some aspect of the patient's behaviour or circumstances seemed to have contributed to 31 deaths, while in 15 cases there was probably a deficiency in medical care. CONCLUSIONS: Some preventable asthma deaths still occur, particularly in relation to inadequate treatment. Factors associated with patients' behaviour and circumstances are more difficult to tackle but, if doctors are aware of high risk patients, increased vigilance may prevent some deaths. PMID- 10525557 TI - Temporal trends and ethnic variations in asthma mortality in Singapore, 1976 1995. AB - BACKGROUND: A study was undertaken to examine temporal trends and ethnic differences in the asthma mortality rate in Singapore. METHODS: Asthma mortality rates in Singapore were estimated from vital data for the years from 1976 to 1995. Trends in sex and age specific (5-14, 15-34, 35-59, 60+ years) rates were obtained for four periods (1976-80, 1981-85, 1986-90, 1991-95) and for Chinese, Malay, and Indian subjects for the years when these data were available (1989 95). RESULTS: An increase in asthma mortality was observed in children aged 5-14 years from 0.21 per 100,000 person years in 1976-80 to 0.72 per 100,000 person years in 1991-95. No increases were noted in the other age groups but a small decrease was observed in the 1991-95 period for the 35-59 year age group. Marked ethnic differences in mortality rates were observed. In the group aged 5-34 years the asthma mortality rates were 0.5 per 100,000 in Chinese subjects, 1.3 per 100,000 in Indians, and 2.5 per 100,000 in Malay subjects. Similar 2-4 fold differences were observed in all other age groups. CONCLUSIONS: Apart from genetic factors, environmental exposures and medical care factors which influence asthma prevalence and severity are most likely to be the causes of the observed temporal trends and ethnic differences in the asthma mortality rate in Singapore, but further studies are needed to elucidate these more fully. PMID- 10525558 TI - An empirical comparison of the St George's Respiratory Questionnaire (SGRQ) and the Chronic Respiratory Disease Questionnaire (CRQ) in a clinical trial setting. AB - BACKGROUND: The Chronic Respiratory Questionnaire (CRQ) and the St George's Respiratory Questionnaire (SGRQ) are the two most widely used quality of life questionnaires in chronic obstructive pulmonary disease (COPD). A study was undertaken to compare directly the self-administered version of the CRQ and the SGRQ with respect to feasibility, internal consistency, validity, and sensitivity to changes resulting from bronchodilator therapy. METHODS: One hundred and forty four patients with moderate or severe COPD were randomly assigned to receive three months of treatment with either salmeterol, salmeterol + ipratropium bromide, or placebo. Quality of life was measured at baseline and after 12 weeks of treatment. RESULTS: The proportions of missing values per patient were low for both questionnaires (0.54% for the CRQ and 2% for the SGRQ). The internal consistency was good for both questionnaires (Cronbach's alpha coefficients >/= 0.84 for the CRQ and >/= 0.76 for the SGRQ). Factor analysis confirmed the original domain structure of the CRQ but not of the SGRQ. Correlations with forced expiratory volume in one second (FEV(1)) % predicted and peak expiratory flow rate (PEFR) were low for both questionnaires but better for the SGRQ than for the CRQ. The ability to discriminate between subjects with different levels of FEV(1) was somewhat better for the SGRQ. The correlations with symptom scores were comparable for both questionnaires. Cross sectionally, the scores of the two questionnaires were moderately to highly correlated (coefficients ranged from 0.35 to 0.72). Longitudinally, these correlations were lower (coefficients ranged from 0.17 to 0.54) but were still significant. The CRQ total and emotions score and the SGRQ symptoms score were the most responsive to change. The SGRQ symptoms domain was the only domain where the improvement in patients receiving combination treatment crossed the threshold for clinical relevance. CONCLUSIONS: Since this analysis of reliability, validity, and responsiveness to change did not clearly favour one instrument above the other, the choice between the CRQ and the SGRQ can be based on other considerations such as the required sample size or the availability of reference values. PMID- 10525559 TI - Domestic biomass fuel combustion and chronic bronchitis in two rural Bolivian villages. AB - BACKGROUND: Chronic bronchitis is an important public health problem worldwide. A study was undertaken to examine the association between exposure to air pollution from domestic biomass fuel combustion and chronic bronchitis in two rural Bolivian highland villages: a village in which cooking is done exclusively indoors and a village in which cooking is done primarily outdoors. Apart from this difference, the villages were virtually identical in terms of socioeconomic status, climate, altitude, access to health care, and other potential confounders. METHODS: Pollution exposure was assessed by combining information on concentrations of particulate matter of <10 microm diameter (PM(10)) in 12 randomly selected households in each village in all potential microenvironments of exposure with time allocation information. The prevalence of chronic bronchitis was assessed using the British Medical Research Council's questionnaire on individuals >20 years of age in both villages (n = 241). RESULTS: Daily pollution exposure was significantly higher in the indoor cooking village (range for adults: 9840-15 120 microg-h/m(3)) than in the outdoor cooking village (range for adults: 5520-6240 microg-h/m(3)) for both seasons and for men and women. The overall prevalence of chronic bronchitis was 22% and 13% for the indoor and outdoor cooking villages, respectively. Logistic regression analysis, which excluded the few smokers present in the population, showed a 60% reduced risk of chronic bronchitis in the outdoor cooking village compared with the indoor cooking village (OR 0.4; 95% CI 0.2 to 0.8; p = 0.0102) after adjusting for age and sex. Individuals aged >40 years were 4.3 times more likely to have chronic bronchitis than the younger age group (OR = 4.3; 95% CI 2.0 to 9.3; p = 0.0002). There was no significant difference in the prevalence of chronic bronchitis in men and women. CONCLUSIONS: The results of this study suggest an association between chronic bronchitis and exposure to domestic biomass fuel combustion, but further large scale studies from other areas of the developing world are needed to confirm the association. Results from this and other studies will assist the development of culturally acceptable and feasible alternatives to the high exposure cooking stoves currently being used by most people worldwide. PMID- 10525560 TI - Clinical significance of respiratory bronchiolitis on open lung biopsy and its relationship to smoking related interstitial lung disease. AB - BACKGROUND: Respiratory bronchiolitis-associated interstitial lung disease (RBILD) is a rare form of interstitial lung disease which may present in similar fashion to other types of chronic interstitial pneumonia. The purpose of this study was to undertake a clinicopathological review of 10 patients with RBILD and to examine the clinical and imaging data related to its histopathological pattern, in particular the relationship of RBILD to smoking. METHODS: Thirteen out of 168 retrospectively reviewed patients, from whom biopsy specimens were taken for suspected diffuse lung disease, were identified with a histopathological pattern of RBILD. Three cases were rejected as follow up data were unavailable. The 10 remaining cases constituted the study group and both clinical and imaging data were collected from patients' notes and referring physicians. RESULTS: Histopathologically, four cases of RBILD overlapped with the pattern of desquamative interstitial pneumonitis (DIP) and nine also had microscopic evidence of centrilobular emphysema. Nine patients were smokers, ranging from 3 to 80 pack years. The one non-smoker had an occupational exposure to the fumes of solder flux. The sex distribution was equal with an age range of 32-65 years. Two patients were clubbed. Lung function tests showed both restrictive and obstructive patterns together with severe reductions in carbon monoxide transfer factor in seven patients. Chest radiographs showed reticular or reticulonodular infiltrates in five patients and a ground glass pattern in two. CT scans were consistent with either DIP or RBILD in six of eight patients. Although seven patients remained stable or improved, either with or without treatment, three patients deteriorated. CONCLUSIONS: This study adds weight to the hypothesis that smoking can cause clinically significant interstitial lung disease, with deterioration in pulmonary function despite treatment. Given the overlapping histopathological patterns of RBILD and DIP and their strong association with smoking, the term "smoking related interstitial lung disease" is suggested for those patients who are smokers. PMID- 10525561 TI - Granulocyte-colony stimulating factor levels in bronchoalveolar lavage fluid from patients with idiopathic pulmonary fibrosis. AB - BACKGROUND: Granulocyte-colony stimulating factor (G-CSF) is known as a potent neutrophil chemotactic glycoprotein in vitro but its contribution to chemotactic activity in neutrophil mediated lung diseases is not yet known. The aims of this study were to determine whether G-CSF is present in high concentrations in bronchoalveolar lavage (BAL) fluid of patients with idiopathic pulmonary fibrosis (IPF, also called cryptogenic fibrosing alveolitis), a neutrophil mediated lung disease, and to what extent G-CSF in BAL fluid contributes to neutrophil accumulation in the lung of patients with IPF. METHODS: G-CSF concentrations in BAL fluid samples from 16 healthy volunteers, 24 patients with IPF, and 73 patients with non-IPF lung disease were measured by enzyme linked immunosorbent assay. The relationship between G-CSF concentrations and neutrophil count in BAL fluid was also examined. Neutrophil chemotactic activity (NCA) was measured in BAL fluid in healthy volunteers and patients with IPF. The contribution of G-CSF to overall NCA in lungs with IPF was assessed by repeating the measurement of NCA after a complete neutralisation of G-CSF bioactivity by anti-human G-CSF antiserum. RESULTS: Detectable levels of G-CSF were found in BAL fluid of 83% of patients with IPF while the levels in all healthy volunteers were below the detection limit. In patients with IPF a significant correlation was observed between the BAL fluid neutrophil count and the concentration of G-CSF in the BAL fluid. The neutrophil count also correlated significantly with percentage forced vital capacity. In BAL fluid samples from patients with IPF the mean NCA value was reduced by 35% after neutralisation with an anti-human G-CSF antiserum. CONCLUSIONS: G-CSF may be involved in enhancing neutrophil accumulation in the lungs of patients with IPF. PMID- 10525562 TI - Dietary factors and pulmonary function: a cross sectional study in middle aged men from three European countries. AB - BACKGROUND: Results of epidemiological studies relating individual dietary factors to chronic obstructive pulmonary disease (COPD) are inconsistent. To evaluate the cross sectional association of dietary factors with pulmonary function, data were collected from middle aged men in three European countries. METHODS: The data were collected in the 1960s in Finland (n = 1248), Italy (n = 1386), and the Netherlands (n = 691). Dietary intake was estimated using the cross-check dietary history method. Forced expiratory volume (FEV(0.75) or FEV(1), here called FEV) was measured by spirometry. Associations were adjusted for age, height, smoking, body mass index (BMI), alcohol consumption, and energy intake. RESULTS: FEV was positively associated with intake of vitamin E in Finland, with intake of fruit in Italy, and with intake of beta-carotene in the Netherlands. In all three countries men with intakes of both fruit and vegetables above the median had a higher FEV than those with a low intake of both foods. The difference in FEV ranged from 110 to 169 ml before and from 53 to 118 ml after energy adjustment. Differences in FEV for intake of three antioxidants (vitamins C and E and beta-carotene) above versus below the median ranged from 61 to 181 ml before and from -35 to 58 ml after energy adjustment. Intake of fish was not associated with FEV. CONCLUSIONS: In three European countries a high intake of fruit and vegetables was positively associated with pulmonary function. A high intake of all three antioxidants tended to be positively associated with pulmonary function before, but not after, adjustment for energy intake. Associations of individual antioxidants with pulmonary function were not consistent across countries. PMID- 10525563 TI - Particulate air pollution and the blood. AB - BACKGROUND: Particulate air pollution has been associated with excess deaths from, and increases in hospital admissions for, cardiovascular disease among older people. A study was undertaken to determine whether this may be a consequence of alterations in the blood, secondary to pulmonary inflammation caused by the action of fine particles on alveolar cells, by repeatedly measuring haematological factors in older people and relating them to measurements of exposure to airborne particles. METHODS: One hundred and twelve individuals aged 60+ years in two UK cities provided repeated blood samples over 18 months, 108 providing the maximum of 12 samples. Estimates of individual exposure to particles of less than 10 microm diameter (PM(10)), derived from a mathematical model based on activity diaries and comparative measurements of PM(10) at multiple sites and during a variety of activities, were made for each three day period prior to blood sampling. The relationships between blood values and estimates of both personal exposure and city centre measurements of PM(10) were investigated by analysis of covariance, adjusting for city, season, temperature, and repeated individual measurements. RESULTS: Estimated personal exposure to PM(10) over the previous three days showed negative correlations with haemoglobin concentration, packed cell volume (PCV), and red blood cell count (p<0.001), and with platelets and factor VII levels (p<0.05). The changes in red cell indices persisted after adjustment for plasma albumin in a sample of 60 of the subjects. City centre PM(10) measurements over three days also showed negative correlations with haemoglobin and red cell count (p<0.001) and with PCV and fibrinogen (p<0.05), the relationship with haemoglobin persisting after adjustment for albumin. C reactive protein levels showed a positive association with city centre measurements of PM(10) (p<0.01). Based on a linear relationship, the estimated change in haemoglobin associated with an alteration in particle concentration of 100 microg/m(3) is estimated to have been 0.44 g/dl (95% CI 0.62 to 0.26) for personal PM(10) and 0.73 g/dl (95% CI 1.11 to 0.36) for city centre PM(10) measurements. CONCLUSIONS: This investigation is the first to estimate personal exposures to PM(10) and to demonstrate associations between haematological indices and air pollution. The changes in haemoglobin adjusted for albumin suggest that inhalation of some component of PM(10) may cause sequestration of red cells in the circulation. We propose that an action of such particles either on lung endothelial cells or on erythrocytes themselves may be responsible for changing red cell adhesiveness. Peripheral sequestration of red cells offers an explanation for the observed cardiovascular effects of particulate air pollution. PMID- 10525564 TI - Effect of oral L-arginine on airway hyperresponsiveness to histamine in asthma. AB - BACKGROUND: Nitric oxide (NO) may exert protective properties within the airways of asthmatic patients. It was postulated that airways obstruction in asthma may be associated with endogenous NO deficiency caused by limited availability of NO synthase substrate. METHODS: In a double blind, crossover study 14 asthmatic patients received pretreatment with oral L-arginine (50 mg/kg body weight) or placebo prior to histamine challenge. Histamine challenge was performed until a 50% fall in forced expiratory volume in one second (FEV(1)) occurred and the response was expressed as the provocative concentration causing a 20% fall in FEV(1) (PC(20)) and as the dose-response slope (maximal % fall in FEV(1)/cumulative dose (micromol)). RESULTS: Pretreatment with L-arginine did not affect PC(20) histamine (mean change in doubling dose 0.18 (95% confidence interval (CI) -0.36 to 0.71), p = 0.5) but the dose-response slope to histamine was slightly reduced (mean change: 0.7 (95% CI 0.6 to 0. 9), p = 0.016). CONCLUSIONS: Oral L-arginine does not influence airway hyperresponsiveness to histamine as reflected by PC(20), although the dose-response slope is slightly reduced in patients with asthma. This indicates only marginal, clinically unimportant limitation of NO synthase substrate in asthma. PMID- 10525565 TI - Acute respiratory distress syndrome and nosocomial pneumonia. PMID- 10525567 TI - Successful treatment of post-influenza pseudomembranous necrotising bronchial aspergillosis with liposomal amphotericin, inhaled amphotericin B, gamma interferon and GM-CSF. AB - A case of aspergillus tracheobronchitis following influenza A infection in an immunocompetent 35 year old woman is described that required prolonged mechanical ventilation for airways obstruction. Treatment included liposomal amphotericin, inhaled amphotericin, gamma interferon and GM-CSF. Liposomal amphotericin therapy was associated with reversible hepatosplenomegaly. Inhaled corticosteroids with continued antifungal therapy were used for the management of severe recurrent airway obstruction. After a prolonged course of treatment she survived with fixed airways obstruction unresponsive to corticosteroids. PMID- 10525568 TI - Key topics in respiratory medicine PMID- 10525566 TI - Athletes and doping: effects of drugs on the respiratory system. PMID- 10525590 TI - [The patient's wishes--the first principle]. PMID- 10525591 TI - [Treatment of patients incapable of giving their consent. Legal requirements in anesthesia and intensive care medicine]. AB - A treatment procedure requires the consent of the patient, but this is legally effective only if he is capable of giving his consent and can be informed accordingly. Because of demographic development and the progress of medicine, the number of patients who are not able to give their consent is increasing. In practice, we make do with the presumed consent of the patient or, for procedures that can wait, with the consent of legitimate family members. An initiative action is suggested by physicians and hospitals that should reduce this gray zone and the forensic risks drastically. PMID- 10525592 TI - [Cisatricurium in the orbicularis oculi muscle. Comparisn of the neuromuscular action of cisatracurium and atracurium in the orbicularis oculi muscle and the adductor pollicis muscle]. AB - OBJECTIVES: Muscle relaxants have different pharmacodynamic profiles in various muscles. Therefore, results obtained for one muscle cannot be extrapolated to other muscles. In the adductor pollicis muscle cisatracurium exerts a pharmacodynamic profile comparable to atracurium, despite the known difference in onset time. However, studies evaluating the neuromuscular effect of cisatracurium in different muscles are lacking. Accordingly, this study compares the pharmacodynamic profile of cisatracurium and atracurium in the orbicularis oculi muscle (OO) - which shows a neuromuscular course similar to the diaphragm and the laryngeal muscles - and the adductor pollicis muscle (AP). METHODS: Forty-five patients (ASA I-II), scheduled for elective spinal surgery were anaesthetized with propofol and fentanyl. Endotracheal intubation was performed without using a muscle relaxant. Neuromuscular transmission was monitored using acceleromyography in both muscles. Patients received 0.1 mg/kg (2x ED(95)) or 0.15 mg/kg (3x ED(95)) cisatracurium, or 0.5 mg/kg atracurium (2x ED(95)) at random. Onset and recovery times were measured according to the recommendation of the Copenhagen Consensus Conference. RESULTS: Onset time was significantly shorter in the OO than in the AP following 0.15 mg/kg cisatracurium and 0.5 mg/kg atracurium (P<0.05). No differences in onset time between the two muscles were found after 0.1 mg/kg cisatracurium. The recovery of T(1) to 10% of its control was completed sooner in the OO than in the AP in all three groups (P<0.05). CONCLUSIONS: Cisatracurium shows a dose-dependent shorter onset time in the OO than in the AP. This is consistent with the current view that the onset of non-depolarizing neuromuscular blockers is more rapid in the OO than in the AP. However, at least a dose of 3x ED(95) of cisatracurium was necessary to show a difference in onset time between both muscles. In contrast, atracurium is reported to lead to a significantly shorter onset of neuromuscular block in the OO following 2x the ED(95). The more rapid recovery of T(1) to 10% of its control in all three groups in the OO is due to the relative resistance of this muscle to muscle relaxants. PMID- 10525593 TI - [Validation of a risk score for prediction of vomiting in the postoperative period]. AB - BACKGROUND: A risk score to predict postoperative vomiting was presented in a recent issue of this journal. In the present study this score was evaluated at another hospital under different surgical and anaesthetic conditions. Furthermore, we examined whether the score, which was originally designed to predict the occurrence of postoperative vomiting (POV) only, is also useful for prediction of postoperative nausea and vomiting (PONV). METHODS: The risk score was applied to 226 patients undergoing inpatient orthopaedic surgery under standardised general anaesthesia (propofol, desflurane in N(2)O/O(2), fentanyl, vecuronium, postoperative opioid analgesia). For 24 hours postoperatively, the patients were followed up for the occurrence of nausea, retching, and vomiting. Perioperatively, risk factors for POV were recorded (gender, age, smoking habits, history of previous PONV or motion sickness, duration of anaesthesia). Using these risk factors the individual risk for suffering POV was calculated for each patient. With these data two ROC-curves (for prediction of POV and PONV respectively) were constructed and the area under the ROC-curve (AUC) as a means of the prediction probabilities of the score was calculated. RESULTS: The incidence of POV as predicted by the score (22,8%) fits well to the actual incidence of this event (19,5%). The score predicts the occurrence of POV significantly better than can be expected by a random estimation. In spite of different surgical and anaesthetic conditions, the accuracy of the prediction in the present dataset was not significantly different from that reported by the authors of the scores in their validation set. Furthermore, the prediction properties for POV (AUC: 0,73) were not different from the prediction of PONV (AUC: 0,72). CONCLUSION: The present risk score provides valid prognostic results even under modified surgical and anaesthetic conditions, and, thus, may obviously be applied to other institutions. Furthermore our results support the hypothesis, that individual risk factors rather than the type of surgery or anaesthetic management have a major impact on the occurrence of POV and PONV. PMID- 10525594 TI - [Effects of long-term opioid therapy on psychomotor function in patients with cancer pain or non-malignant pain]. AB - OBJECTIVES: Despite increasing use of oral opioids in cancer and non cancer pain, little is known about the effects of long-term opioid therapy on psychomotor performance. This study was designed to investigate the effects of long-term opioid analgesia on attention and reaction time in cancer pain and in non malignant pain. METHODS: Three groups of patients (n=128) were studied: 48 patients on long-term opioid therapy (group O; including 33 patients with cancer pain and 15 patients with chronic non-malignant pain), 30 patients receiving non opioid analgesic therapy for chronic non-malignant pain (group NO) and a control group (group K) of 50 patients without pain and analgesic therapy. Attention was determined by Brickenkamp's d2-test, continuous reaction time by Schuhfried's method (Wiener Determinationsgerat, Modling, Austria). In addition, a modified questionnaire developed by Zerssen was used to determine the patient's current mood. Pain, fatigue and anxiety levels were estimated by visual analogue scales. RESULTS: Although no significant difference in attention/concentration could be demonstrated between the three groups, patients taking opioids performed Brickenkamp's test a little worse and also demonstrated a significant decline in this parameter with advancing age. Also, in cancer patients attention/ concentration was more impaired than in non-cancer opioid patients. Auditory and optical reaction times were significantly slower in patients on opioids than in the non-opioid analgesic group and highly significant slower than in the control group, while in the more complex combinations test no such difference could be demonstrated. In addition, a highly significant deterioration in reaction times with increasing age could be demonstrated for opioid patients compared to the other groups, while only a non significant prolongation was found between cancer and non-cancer patients on opioid therapy. CONCLUSIONS: Long-term opioid therapy produces a slight (non significant) impairment of psychomotor performance in patients with cancer pain or non-malignant chronic pain. These effects become significantly more pronounced with increasing age and in patients with cancer pain, indicating a higher susceptibility of the elderly towards opioids. These results indicate that, particularly in older patients receiving long-term opioid for cancer oder non-cancer pain, careful evaluation of their effects on psychomotor function is necessary in order to estimate patient's ability to perform his daily activities. However, since opioid effects were only minimal in the non-elderly other factors like basic disease, opioid dose, physical condition and age seem to be of greater importance than the effects of opioids per se. PMID- 10525595 TI - [The use of remifentanil in critically ill patients. Clinical findings and early experience]. AB - OBJECTIVES: It was the aim of this investigation to report our initial clinical experience on the use of remifentanil in critically ill patients undergoing mechanical ventilation. Additionally, we hypothesized that even under intensive care conditions remifentanil might facilitate a temporally predictable and "programmed" tracheal extubation. METHODS: Remifentanil was used for analgesia and sedation of mechanically ventilated patients who were admitted to the ICU following major noncardiac surgery or who had to be ventilated due to respiratory failure. The infusion was started with 0.15 microg/kg/min and then adapted in steps of 0.05 microg/kg/min according to clinical needs. After admission to the ICU the depth of sedation was adjusted to a Ramsay score level of 4 (sleeping patient, immediately arousable) and then targeted at a level of 2-3 (patient awake, co-operative and tranquil or responding to command only). In case of sufficient pain relief but inadequate sedation patients could receive bolus doses of midazolam (1-3 mg) or an infusion of clonidine (0.5 microg/kg/h), the latter especially in case of shivering or hypertension. Prior to extubation bolus doses of piritramide (3-5 mg) and a non-opioid analgesic (metamizol or propacetamol) could be used for postoperative pain relief. Data are presented as mean+/-SD. RESULTS: A total of 46 patients were studied, aged 62.8+/-15.4 yr with a mean APACHE II score of 19.2 points. The duration of remifentanil infusion ranged up to 78 h with a mean of 9. 8 h. The mean infusion rate was 0.14+/-0.08 microg/kg/min during ongoing analgesia and sedation and 0.10+/-0.08 microg/kg/min immediately before its discontinuance. Additional sedatives were necessary in 63% of all patients. Emergence was rapid in the majority of cases: 67% of all patients could safely be extubated within 15 min after termination of remifentanil, and a total of 87% were extubated within 45 min. A development of tolerance was not observed during the study period. CONCLUSIONS: Remifentanil appeared to be suitable for analgesia and sedation of critically ill patients undergoing mechanical ventilation: Even under intensive care conditions recovery was rapid in the majority of cases, and in two thirds of all patients tracheal extubation was temporally predictable and could be timed within 15 min. These results are best explained by the metabolism and offset of action of remifentanil obviously unaffected in the ICU area. However, for fast emergence the cautious use of additional sedatives is crucial. PMID- 10525596 TI - [Measurement technique. Systems and methods for intracranial pressure monitoring]. AB - GOAL: Intracranial pressure (ICP) monitoring has a key role within the neuromonitoring, although ICP does not monitor processes of the central neuron directly and only with delay. One of the important factors in ICP monitoring is measurement accuracy. For a better understanding of ICP probes and their differences, the function and principles of intracranial pressure transducers should be evaluated from a technical point of view. METHOD: The principles of ICP measurement were analyzed and compared. Practical applications of these principles were examined and examples of different ICP probes were discussed regarding their mode of pressure transformation. The technical advances of ICP monitoring were analyzed. RESULTS: Since LUNDBERG, a variety of different types of transducers has been developed. Ventricular ICP monitoring has been supplemented by extradural and intraparenchymatous probes. An increasing miniaturization of the transducers has emerged. Additionally, fiberoptic systems have been developed. Latest developments include multifunctional ICP probes. So far, the main problem of most types of transducers consists in the inability to assess measurement accuracy of a probe during the period of patient monitoring. CONCLUSION: ICP probes should be tested better for correct function by the manufacturer prior to sale. External controls of the measurement accuracy should be performed more frequently to ensure constant quality. Future ICP transducers have still to be more cost-effective. PMID- 10525597 TI - [Persisting respiratory depression following intrathecal administration of morphine and simultaneous sedation with midazolam]. AB - A 72-year-old patient received 0.1 mg morphine by the intrathecal route and 2 x 1.5 mg midazolam as adjuvant therapy. Severe respiratory depression and somnolence supervened 3.5 h later, which lasted over the next 24 h and necessitated intubation and mechanical ventilation. Continuous administration of >6 mg naloxone to antagonize the supposed effect of the morphine had no effect. The patient's condition was not normalized until a single dose of 0.3 mg flumazenil was administered. For the time being, especially in the case of elderly patients, we recommend that strict indications are adhered to when intrathecal administration of morphine is considered and that less than 0.1 mg morphine is given. Diazepines should be avoided. Respiration should be monitored for quite some time. PMID- 10525598 TI - [History of the development of intensive care medicine. Part 9: Architectural development of intensive treatment wards]. PMID- 10525599 TI - [Anesthesia and hyperbilirubinemia]. PMID- 10525600 TI - [ECG monitoring in electrical injury]. PMID- 10525601 TI - [ [In Process Citation] PMID- 10525602 TI - [46th German anesthesia congress. Wiesbaden, May 5-8, 1999]. PMID- 10525603 TI - [Jet ventilation and anaesthesia for diagnostic and therapeutic interventions of the airway]. PMID- 10525604 TI - [Reports from the Aachen annual meeting of the German Society for ENT Medicine, Head and Neck Surgery]. PMID- 10525605 TI - [Reports from the Aachen annual meeting of the German Society for ENT Medicine, Head and Neck Surgery. Alcohol drinking and risk of cancer in the area of the upper digestive tract]. PMID- 10525606 TI - [Prognostic significance of cell-cycle regulatory proteins for outcome after primary radiochemotherapy in patients with advanced head and neck cancer]. AB - BACKGROUND: Primary radiochemotherapy is gaining increasing importance for the treatment of advanced head and neck squamous cell carcinomas. However, there is a lack of clinical factors concerning prognostic information in relation to treatment. In this pilot study, we examined whether molecular factors can provide such information. PATIENTS AND METHODS: The expression patterns and their possible prognostic relevance of the cell cycle regulatory proteins p53, p21(WAF/CIP1), Rb, p16(INK4A), CDK4 and Cyclin D1, MIB1 (Ki-67) and BCL-2 were analysed in pretreatment tumor biopsies from 43 patients with advanced carcinomas of the oropharynx (n = 26), hypopharynx (n = 10) and larynx (n = 7) by applying immunohistochemistry to paraffin sections of tumor specimens. All patients were treated by the same method of an accelerated "concomitant boost" radiochemotherapy with carboplatin in a phase II study. Median followup time was 56 months. RESULTS: No correlation was found between any of the markers and the remission rate, T-stage, N-stage, rate of loco-regional recurrences and distant metastases. However, independent of the tumor stage, patients with CDK4/cyclin-D1 overexpressing tumors had a significantly shortened overall survival (P = 0.014 and 0.026, respectively). CONCLUSION: The results of this pilot study indicate that CDK4 and cyclin D1 over-expression provide useful prognostic information about clinical outcome after primary radiochemotherapy of head and neck cancer patients. PMID- 10525607 TI - [Category loudness scaling to evaluate sound perception in cochlear and retro cochlear lesions]. AB - Category loudness scaling was used to investigate the loudness perception of 31 patients with a cochlear hearing loss (Group 1) by comparing the results with those found in 15 patients with retro-cochlear hearing loss caused by an acoustic neuroma (Group 2). Narrow-band noise signals at four different frequencies (0.5 to 4.0 kHz) were used. In the cochlear hearing-impaired subjects the slopes of the level-loudness functions tended to increase with increasing hearing loss, indicating positive recruitment, whereas the much shallower slopes associated with retro-cochlear lesions were presumed to reflect negative recruitment. The graphic representation of the iso-loudness functions revealed a different dynamic range between Group 1 and 2 with the ability to discriminate small differences of stimulus levels reduced in the presence of an acoustic neuroma. Category loudness scaling has been shown to be a valuable tool describing the individual perception of sound in a qualitative and quantitative manner. Furthermore, the method can be employed as an indicator of recruitment without any restrictive preconditions. For this reason the categorical loudness scaling can be a desirable method for supplementing the audiological diagnosis of a retro-cochlear hearing impairment through the frequency-specific description of a usable hearing-field and its dynamic range. PMID- 10525608 TI - [The role of oncogenic human papillomaviruses in tonsillar squamous cell carcinomas with functional inactivation of the retinoblastoma protein]. AB - In order to identify squamous cell carcinomas of the head and neck (HNSCC) with common biological and clinical features, we investigated the incidence and properties of carcinomas lacking retinoblastoma protein (pR6) cell cycle control. Of 208 HNSCC investigated, 23 (11%) showed a lack of pRb expression. The majority of these tumors (65%) were tonsillar carcinomas. The pRb-negative tonsillar tumors were all stage IV, had metastasized to lymph nodes at the time of diagnosis and were in general poorly differentiated or undifferentiated. Very significantly, the pRb-negative phenotype was strongly associated with the presence of oncogenic human papilloma viruses, implying a viral etiology and functional inactivation of pRb by the viral E7 oncoprotein. Despite the very adverse histopathological factors, patients with pRb-negative tonsillar carcinomas had a better clinical outcome, which was consistent with a uniform favorable responsiveness of these tumors to postoperative radiation therapy. PMID- 10525610 TI - [Vocal capabilities of nonprofessional singers evaluated by measurement and superimposition of their speaking, shouting and singing voice range profiles]. AB - Voice range profile(VRP) measurement (Phonetography) was used for the evaluation of the vocal capabilities of 41 female (F) and 50 male (M) members of a nonprofessional choir. By means of an automatic VPR measurement system F0 and SPL dB(A) were determined and displayed real time, two-dimensionally. The speaking voice (reading a standard passage as well as counting from the softest to the loudest intensity), the shouting voice (3-4 times shouting a standard sentence) and the singing voice (sustained phonation / la:/ at minimum and maximum intensity level) were measured. The VRPs of these voice modalities were superimposed on the screen and the plot. The averaged values for the speaking VRP: intensity range (F): 48 dB (range 46 soft to 94 dB loud phonation), (M): 52 dB (range 46-98). Pitch range (F): 15 semitones (ST) (Cis3, 138-E4, 329 Hz), (M): 19 ST (E2, 82 Hz-H3, 246 Hz). The average slope for the speaking voice (F): 0,31 ST/dB, (M): 0,36 ST/dB. Shouting VRP highest intensity (F): 106,5 dB, (M): 108,5 dB, highest pitch (F): between Ais4, 466 and H4, 493 Hz. (M): E4, 329 Hz. Average slope for speaking and shouting voice (F): 0,36 ST/dB, (M): 0,39 ST/dB. Singing VRP pitch range (F): 34,6 ST, (M): 37 ST, intensity range (F): 60 dB, (M): 58 dB. The pitch extension of the speaking VRP ranges from 2,9 to 46,2%, speaking and shouting VRPs together with 2,9 to 65% of the pitch range of the singing VRP (F), (M) 2,7-54% and 2,7-67,5% accordingly. The average values for nonprofessional singers reflect an effective but not special use of the phonatory system for the speaking, shouting and singing voice functions with respect to pitch and intensity. PMID- 10525609 TI - [Objective auditory brainstem response threshold deficits in patients with cerebellopontine angle tumors]. AB - The established criteria of auditory brainstem responses (ABR) such as JV latency, JI-V interpeak latency and interaural differences of latency or amplitude have been found to be sensitive for detecting tumors of the cerebellopontine angle if a response is present. However, the ABR can be absent in cases of acoustic neuromas because of desynchronization, even though pure-one audiometry indicates that responses should be present. This retrospective study compared the ABR and pure-tone thresholds in 234 cases with cerebellopontine angle tumors and a control group of 181 cases with sensory hearing losses in order to quantify threshold discrepancies. The average deficit of the objective ABR threshold (DOABRT) to the subjective pure-tone threshold for those frequencies between 1-6 kHz was 3.6 dB for the control group (ABR and pure tone thresholds very close) and 31.2 dB for the tumor group (ABR threshold much higher than the pure-tone threshold). ABR thresholds 30 dB higher than the high frequency pure tone thresholds were found in 40.6% of the tumor group and in none of the control group. Thus, deficits of the ABR threshold >30 dB can be considered to be an additional criterion for detecting retrocochlear disease and increases ABR sensitivity for tumor detection even if responses are absent. PMID- 10525611 TI - [Management of large bilateral glomus jugulare tumors]. AB - Bilateral glomus jugulare tumors are rare. However, their treatment should preserve not only the function of the facial nerve but also the caudal cranial nerves and the middle ears in order to avoid bilateral hearing losses. Further, venous cerebral drainage has to be ensured in order to avoid cerebral hypertension and hemorrhagic infarction after bilateral jugular ligations. In the case presented bilateral glomus jugulare tumors required super-selective angiography and embolization. Complete tumor removal on both sides was then possible by a transmastoid-transcervical approach without any further functional deteriorations. Middle ear function was preserved on both sides by temporary ventral translocation of the posterior wall of the auditory meatus. As the sigmoid sinus and internal jugular vein had been ligated during initial previous surgery, venous drainage was tested one year later by angiography and compression of the remaining internal jugular vein. A sufficient collateral circulation was found and permitted surgery on the second side. PMID- 10525612 TI - [Cervical thymic cysts in the differential diagnosis of lateral neck tumors. Case report]. AB - Cervical thymic cysts belong to the rare causes of neck masses and therefore are frequently not included in a preoperative differential diagnosis. Here we report our experience in managing a 7-year-old boy who presented with a three-month history of a lateral neck mass causing stridor during sleep. Clinical findings and macroscopic and histopathological features are described and reviewed with respect to the available literature. The inclusion of a cervical thymic cyst in a preoperative differential diagnosis is important for determining the extent of the neck mass and planning any surgical procedure. PMID- 10525613 TI - [Medication-induced vocal cord paralysis as an uncommon cause of cord paralysis]. AB - A case is reported of a 57-year-old man who was found to have a right vocal cord paralysis that most likely followed prolonged treatment with the anti-arrythmic medication Amiodaron-HCl (Cordarex). Phoniatric treatment was given for 5 1/2 months, during which time microlaryngoscopy and stroboscopy were performed. With the help of speech therapy, mobility of the paralyzed cord was seen to begin to return 3 1/2 months after discontinuing the Amiodaron-HCl. Full cord mobility has not returned to date. PMID- 10525615 TI - [Unidentified swelling in the lateral neck area. Fusiform extracranial aneurysm of the internal carotid artery]. PMID- 10525614 TI - [Parapharyngeal tumor in an infant. Cervical neuroblastoma, stage IV]. PMID- 10525617 TI - [Injuries of the posterior cruciate ligament]. PMID- 10525616 TI - [p53 mutations/p53 protein overexpression. Differential significance for the progression of head-neck carcinomas]. PMID- 10525618 TI - [Diagnostic and incidence of the rupture of the posterior cruciate ligament]. AB - The purpose of this article is to evaluate the incidence and to give a general review of the examination of the posterior ligament complex. At least ca. 8-10 % of all severe ligament injuries concern the posterior cruciate ligament, which means, that an estimated 4,000-5,000 Germans suffer a PCL rupture every year. Motor-vehicle accidents are the most common cause of the injury, but sports related traumas (football, skiing) have increased in recent years. The high number of high-energy mechanisms involved (up to 90 %), cause ligament ruptures often to be associated with other injuries, especially fractures of the femur and tibia head. In polytrauma patients PCL ruptures are frequently recognized very late, because the possibility of this kind of injury is often not considered during the clinical examination. The same holds for the diagnosis of monotrauma patients. The initial step in the evaluation is to obtain a thorough history (including the mechanism of injury) and to perform a physical examination. The instability after a PCL rupture may present as an ACL rupture, because the anterior drawer test seems to be positive. The anterior/posterior drawer test must be assessed with other evaluation procedures to distinguish between anterior und posterior instabilities. The posterior sag sign, the quadriceps active test or the reversed pivot-shift may indicate a PCL rupture. A correct roentgenogram can reveal an avulsion of the tibia and can prove posterior instability due to a posterior translation of the tibia. A quantitative examination (clinical or X ray) of the instability and the indication of combined injury of the posterior cruciate ligament and the posterolateral complex are necessary for the therapeutic decision (operative/conservative). A rupture of the PCL may occur occasionally as a result of a luxation of the knee (reduced spontaneously) before the medical evaluation. A thorough neurovascular examination is essential. Magnetic resonance imaging can be important to the diagnosis of an acute injury, but it is not essential for the choice between operative and non-operative treatment. Arthroscopy has been found to have a high degree of accuracy in the diagnosis of ligament ruptures of the knee, but it is still an operative treatment, so that it can only be used if an operation of repair or reconstruction is planned anyway. Before operative treatment of chronic complex instability, potential osseous abnormalities (varus morphotype) must be revealed; in case of uncertainty, an X-ray control is necessary. PMID- 10525619 TI - [Augmented repair as therapy of fresh injury of the posterior cruciate ligament]. AB - The discussion about the therapy of the posterior cruciate ligament persists. Conservative treatment, augmented repair, and reconstruction with autografts are discussed. From 1993 to 1997, 49 patients with posterior cruciate ligament rupture had repair Trevira ligament augmentation of 3 mm. There were 21 isolated and 28 combined ruptures. In 5 cases bony avulsions were refixed by screw or additional hook plate. Investigation of 36 patients, in 15 cases with isolated ligamentous ruptures was made. Osseous avulsion had good results in all cases. Isolated posterior cruciate ligament rupture showed good stability in 7 of 15 cases and instability of 2 + in 8 cases. The medial range Lysholm score was 76.8 (+/- 21.6), the OAK score showed 2 very good and 5 good results, 3 fair and 5 bad results. Using the IKDC score led to 3 very good and 4 good results, 2 fair and 6 bad results. Using subjective criteria, 10 patients described results as very good or good, 2 fair and 3 bad. Posterior cruciate ligament rupture with additional knee injury or fracture of the leg showed bad results in 60 % of cases, and good or fair results in only 40 %. We think augmented repair of fresh injury of the posterior cruciate ligament can be used as an alternative therapy to reconstruction with autograft. PMID- 10525620 TI - [Treatment of the rupture of the distal tibiofibular syndesmosis with "Engelbrecht's syndesmosis hook"]. AB - After completing the osteosynthesis of the lateral malleolus, the hook is positioned 3-5 cm cranial of the syndesmosis and fixed to the tibia with a screw. It is expected to allow undisturbed ligament healing of the syndesmosis with early formation or fibres under limited motion. No immobilisation is necessary and no early removing of the implant. RESULTS: Of 62 patients questioned, 53 were examined, after an average of 50 months (2-6 years p. o.). Using the Weber score, 51 had excellent or good results. Twelve patients reported discomfort while jumping, in five cases the ROM concerned dorsal extension, and in three cases plantar flexion was reduced. Four patients showed distinct radiologic signs of osteoarthrosis, and two patients slight signs. PMID- 10525621 TI - [Late results after fracture of the femoral head]. AB - The dislocation fracture of the femoral head is the result of high speed trauma. Most of the patients have additional injuries. The prognosis of this kind of fracture of the femoral head depends on the type of fracture, the additional injuries and the age of the patients. The diagnosis and the specific treatment are most important, since most of the patients with this injury are of a younger age. The reposition of the fracture has to be performed within 6 hours. In our opinion, this should be done by surgery if possible. For the operation some routine pelvic X-rays and a CT of the pelvis should be prepared. The therapy depends on the type of fracture. In patients with Type I and II fractures the broken head fragments should be refixed by only taking out small parts of bone which are not elementary for the pressure zone of the femoral head. Younger patients with Type III fractures should always receive the possibility of a screw fixation of the neck of femur, whereas total hip replacement should generally be achieved in the older patient. An exact reconstruction of the dorsal acetabulum must be performed in Pipkin Type IV fractures. The usual approach for Type I-III fractures is the ventrolateral Smith-Peterson and lateral Watson-Jones, for Type IV fractures, the dorsal Kocher-Langenbeck approach. We suggest indometacine as a prophylaxis for ossifications due to high tissue damage. Several scores for the evaluation and documentation of the outcome of this kind of fracture are useful: the clinical results according to Merle d'Aubigne, social status scored by the Karnofsky Index and X-ray results using Brooker and Helfet to classify the heterotopic ossification and post traumatic joint changes. PMID- 10525622 TI - [Pediatric forearm fractures: indications, technique, and limits of conservative management]. AB - Although several "minimal invasive" techniques for the operative management of pediatric forearm fractures have been developed recently, conservative treatment still remains the option with the lowest risk for small patients. We present the results of our clinical and radiological follow-up after an average of 52.4 months (4-112) in 102 pediatric patients. All fractures were treated conservatively. There were 68 fractures (66.7 %) of the distal third of the forearm, 30 fractures (29.4 %) of the midshaft area, and four fractures (3.9 %) in the proximal third of the shaft. Greenstick fractures were seen in 58 cases (56.8 %), complete fractures with displacement of both corticalices in 26 patients (25.5 %), and folding fractures in 18 cases (17.7 %). With the exception of one fracture with the necessity of remanipulation after redisplacement in the cast, all fractures healed uneventfully without any further intervention. Functional results were excellent with a free range of motion of the wrist and elbow and without any signs of muscular atrophy in 96 children (94.1 %) at the time of follow-up. Six patients, however, showed a significant loss of forearm rotation of an average of 25 degrees (15 degrees -50 degrees ). In four of these six patients, the fracture had been situated in the proximal and midshaft area. Thus, two out of four fractures of the proximal forearm (50.0 %) showed a poor functional outcome. On the basis of our data we recommend conservative management for (closed) pediatric fractures of the distal and midshaft area. Operative treatment is indicated in forearm fractures close to the elbow. PMID- 10525623 TI - [Experimental stability of a new implant-free fixation technique in ACL replacement]. AB - The purpose of the study is to establish data on the stability of an ACL replacement. In 40 human cadaver knees, either a mid patellar ligament third with a trapezoid bone block on one side was fixed on the femoral side in a 2-diameter drill hole, or a conventional interference screw fixation was applied. Average primary stability amounted to 570 N (+/- 100 N) for the bone-blocking technique, and to 402 N ( +/- 79 N) for the interference screw fixation. When statistically tested by variance analysis, stability was significantly higher for the bone blocking technique than with interference screw fixation (p < 0.05). Thus, when using a mid-patellar ligament third for ACL reconstruction, the new implant-free technique described here appears to be a practical and reliable method for femoral graft fixation. PMID- 10525624 TI - [The complicated wound]. PMID- 10525625 TI - [Marburg modular prone positioning system (MBS) for positioning therapy]. AB - Despite the many benefits of prone positioning in critically ill patients with respiratory failure and ARDS in the ICU, its technical problems have not yet been adequately resolved. Different approaches with special beds for prone positioning do exist, but these devices are difficult to handle, often not available and involve high costs. With this in mind, we developed an easy handling prone positioning system (MBS) that requires no special beds and runs at low cost. The MBS is a cost-effective device, yielding many benefits for prone positioning in critically ill patients with severe athelectasis and ARDS. PMID- 10525626 TI - [Posttraumatic aneurysm of the brachiocephalic trunk: a rare injury]. AB - We report on a posttraumatic aneurysm of the brachiocephalic trunk, something which is rather rare. The injury was caused by blunt chest trauma following a car accident. Computed tomography could not find the brachiocephalic lesion. In patients with fractures of the upper ribs after blunt chest trauma, angiography should be done to exclude severe injuries of the aorta and brachiocephalic vessels. PMID- 10525627 TI - [Treatment of femoral fracture after total knee arthroplasty with the LIS system: a new method]. AB - The different treatments for femoral fracture after total knee arthroplasty are discussed. A new method with the LIS system (Synthes) is described and first clinical results presented. PMID- 10525628 TI - [Conflicting approaches to wound treatment dependent upon the specialty. How do we achieve a better working relationship between doctors and nurses?]. PMID- 10525629 TI - [Considerations on the differential indication of anterior cruciate ligament replacement]. PMID- 10525630 TI - [Tumescence-local anesthesia. Conversation with Prof. Werner Mang on the state of development of the new local anesthesia procedures (intervew by Werner Rossling/Hinrich Kuster)]. PMID- 10525631 TI - [MR imaging-guided interventions]. AB - The spectrum of MR imaging-guided interventions includes biopsies, thermal ablation modalities, vascular applications, MR endoscopy and intraoperative MR imaging. The concept of MR guidance is based on the excellent morphologic and functional imaging achieved by MR. The most important recently published experimental and clinical results are discussed. In the future, MR imaging will play an important role in interventional radiology, minimal invasive therapy and guidance of surgical procedures. PMID- 10525632 TI - [Image-guided interventions in liver tumors]. AB - Despite remarkable progress of diagnostic imaging and operative procedures radiological interventions play a major role in diagnostic and therapeutic liver tumor interventions. Percutaneous biopsies should be taken by 16-20 g needles. CT control is indicated in cases when sonographically guidance is impossible or of risk. MR guidance is still seldom. Accuracy rates of percutaneous biopsies are high (>90%), and safe with complications (e.g. bleeding) of less than 1%. Palliative percutaneous therapeutic interventions of primary or secondary liver malignancies are thermoablative procedures of laser (LITT), cryoablation or radio frequency, percutaneous ethanol injection (PEI) and intraarterial chemotherapy via port system or repetitive catheterisation with perfusion or embolization (TACE). For metastatic disease with less than five tumors of less than 4 cm LITT and PEI are recommended, more advanced cases should be treated by intra-arterial port system chemotherapy. For HCC best results are shown for PEI, in cases of UICC stage IIIB and IV only TACE is adequate. PMID- 10525633 TI - [MRI-controlled regional hyperthermia]. AB - PURPOSE: Regional hyperthermia in combination with chemotherapy or/and radiotherapy is a promising treatment concept for locally advanced, deep-seated tumors. The purpose of the project is the optimization of the therapy using non invasive, three-dimensional imaging of tissue changes or of the temperature distribution during regional hyperthermia. METHODS: MRI offers methods suitable in principle for tissue characterization and MR thermometry. A new MRI hyperthermia hybrid system has been developed based on an innovative hyperthermia applicator and an open MRI system. RESULTS: After successful testing of the new MRI-hyperthermia hybrid system simultaneous MRI and regional hyperthermia in patients could be accomplished for the first time. At present the T1 relaxation time seems to be a promising parameter for MR thermometry. CONCLUSION: The first clinical application of the MRI-hyperthermia hybrid system can be regarded as an important step towards the development of regional hyperthermia. This new hybrid system and the MR thermometry methods have to be investigated prospectively in clinical studies. PMID- 10525634 TI - [Interventional MR-guided laser induced thermotherapy in oncologic indications. Status and prospects]. AB - MR-guided LITT (laser-induced thermotherapy) is currently being evaluated for its effectiveness in clinical oncology. MR-guided LITT is defined as a minimally invasive technology based on the effects of the applied Nd-YAG laser on tumorous tissue. Due to specific characteristics of the laser-induced coagulative effect, online monitoring via MR thermometry is possible and extremely precise. In a period of 6 years 335 patients suffering from malignant soft tissue tumors were prospectively treated via MR-guided LITT. We evaluated the local tumor control rate, the rate of complications and the survival data from the clinical and MRI follow-up. Our results prove that MR-guided LITT results in a extremely low rate of side effects and an effective tumor control rate higher than 95%, depending on the size of the lesion. It is concluded that this therapeutic concept is of clinical value for patients with primary and secondary liver cancer, malignant lymph node involvement, abdominal recurrent tumors and tumors of the head and neck. PMID- 10525635 TI - [Percutaneous implantation of port-catheter systems for intraarterial chemotherapy of the liver]. AB - PURPOSE: The objective of this study was to determine the usefulness, safety and acceptance of a new technique of percutaneous implantation of port-catheter systems (PIPS) for locoregional intraarterial chemotherapy of the liver. MATERIAL AND METHODS: In 50 patients with malignant hepatic disease, 52 percutaneously implantable port-catheter systems were implanted for intraarterial chemotherapy of the liver as an interventional radiological technique. A commercially available angiographic catheter was placed in the hepatic artery under fluoroscopic control via a transfemoral approach and connected to a Port-A-Cath situated in the groin. This procedure was done on an outpatient basis; no medical treatment was administered. RESULTS: Percutaneous placement of the port-catheter system was successful in all cases, also in those with a hepatomesenteric trunk. No peri- and post-interventional complications occurred. The median patency was 312 days (13-547 days). The catheter-related complication rate was 12%. The function could be restored by replacement or an interventional procedure in all but one case (2%). Infection and leakage did not occur. The system had been withdrawn without complications in 7/52 cases for a variety of reasons (e.g. hemihepatectomy, desire of the patient or clinician, dissection after intervention, replacement). CONCLUSION: Percutaneous placement of a port-catheter system is a safe and easy alternative to the surgical placement of port systems for hepatic intraarterial chemotherapy. Long-term complication rates are comparable. The option of easy withdrawal and interventional correction of dysfunction as well as lower costs are additional advantages. PMID- 10525636 TI - [Percutaneous interventional radiologic implantation of intravenous port-catheter systems]. AB - PURPOSE: Percutaneous interventional radiologic and surgical techniques of port catheter implantation are described and compared with regard to the technical procedure and results. MATERIALS AND METHODS: In 53 patients with various malignancies interventional radiologic implantation of port-catheter systems into the subclavian vein was performed to provide long-term intravenous access for chemotherapy. The technical procedure, operation time, complication rates and long-term patency were compared with those of surgically implanted systems. RESULTS: Implantation was successful in all cases. Mean operation time was 36 min (range 20-55 min). Mean function time was 189 days (range 7-518). Primary patency rate was 92.5% with a total complication rate of 15% (8/53). In three patients (5.7%) pneumothorax was observed but did not require further treatment. In two cases (3.8%) local infection occurred, and in one patient (1.8%) a non complicated wound dehiscence. In 12/53 patients (22.6%) the system was withdrawn. Among these, withdrawal was due to complications in 4/53 (7.6%) cases. CONCLUSIONS: Interventional radiologic implantation of long-term intravenous port catheter systems is comparable to surgical placement with regard to both complication rate and long-term patency. PMID- 10525637 TI - [Renal tumor embolization]. AB - Today the relevance of renal tumor embolization is not determined only by the technical and clinical success of the method. Progress in diagnosis of early stages of renal carcinomas as well as the improvement of both surgical techniques and anesthetic procedures have lead to a change in the selection of patients for embolization. Preoperative embolization of advanced renal cell carcinomas with tumor thrombus into the vena cava or of T4 tumors is now an established clinical procedure. The complete occlusion of the vascular bed of the tumors leads to a considerable reduction in intraoperative blood loss and to simplification of the surgical preparation. By using Ethibloc for embolization, palliation of a hemorrhage or of tumor-related pain in inoperable patients is usually successful. Although local control of the tumor disease, including complete tumor ablation, is achieved by embolization, the median survival rate of our palliatively embolized patients is only 3.5 months. This short life expectancy in the group of inoperable patients has to be acknowledged individually in patients considered for palliative embolization who are free of symptoms related the tumor. PMID- 10525638 TI - [Intraarterial chemotherapy in cases of breast cancer]. AB - A general review of the results and technique of intraarterial chemotherapy in cases of breast cancer is given. The remission rate for untreated primary tumors is nearly 100%, for completely treated local relapses approximately 70%. The complication rate is quite low using an intraarterial well-tolerated cytostatic agent such as mitoxantrone. However, thromboembolism of the vertebral artery facing the internal mammary artery may occur. The indication for intraarterial chemotherapy should always be set by an interdisciplinary board. PMID- 10525639 TI - [Interventional treatment of hemorrhages in advanced cervical carcinoma]. AB - PURPOSE: Retrospective evaluation of percutaneous interventional treatment of locally advanced cervical carcinoma. MATERIALS AND METHODS: Since 1991, 13 patients with advanced tumor disease have been referred to our department for diagnosis and therapy of an acute blood loss. In all patients (age 40-88 years, mean 61 years) hemorrhage was detected by decrease in red blood cell count. In all cases patients suffered from locally advanced or recurrent disease after surgery and/or additional radio- or chemotherapy. Embolization was performed by transfemoral access using minicoils in most cases, liquid agents less often and a covered vascular stent in one patient. RESULTS: The site of the hemorrhage or the blood pooling of the tumor could be seen in all cases angiographically. Twenty seven treatment cycles (2.1 per patient) were performed at intervals of 3 days to 6 months. The maximum time of follow-up and additional treatments if necessary was 1 year. In 9 of 13 patients (69%) the bleeding could be stopped immediately with a single treatment or initial treatment via both iliac arteries. One patient (7,7%) died during therapy because of an uncontrollable bleeding and consecutive decrease in red blood cells count. The remaining three patients (23%) showed slight persistent or recurrent bleeding, which could be managed interventionally until the following episode. There were two complications (15%) during therapy, representing a coil misplacement and a coil wash-out, which both could be managed interventionally. CONCLUSION: Hemorrhage following locally advanced or recurrent cervical carcinoma can be stopped interventionally in about 70% of cases. Even in partial success it is possible to manage the acute life-threatening situation. Follow-up examinations of up to 1 year justify this therapeutic concept. PMID- 10525640 TI - [Patient with suspected struma. Bronchogenic cyst of the upper thoracic aperture]. PMID- 10525641 TI - [Distal choledochal cyst in hydro-spiral CT]. AB - Bile duct cysts are rare abnormalities of the biliary tract. Surgical therapy has been recommended because of possible complications such as cholestasis with jaundice and the risk of bile duct carcinoma. Accurate preoperative radiological imaging is available for surgical planning. In addition to direct imaging of the biliary system in ERCP, high-resolution axial computerized imaging techniques are necessary. The use of MRCP is becoming more frequent in diagnostic imaging of the biliary tract. Similar to the diagnosis of pancreatic tumors with hydro-spiral CT technique, we demonstrate the benefits of hydro-CT in imaging of a distal choledochal cyst, the problems of differential diagnosis and the classification in the generally accepted Todani system in a case in which accurate clarification with MRCP and ERCP was not possible. PMID- 10525643 TI - [ [In Process Citation] PMID- 10525644 TI - [Academy information. German Physician Group decides on introduction of voluntary proof of continuing education]. PMID- 10525645 TI - [ [In Process Citation] PMID- 10525642 TI - [Basics and techniques of spiral CT]. PMID- 10525647 TI - [ [In Process Citation] PMID- 10525646 TI - ["Do not trust a high-gloss brochure". On systematic search for suitable general practice software. 1: Basic principles, system configuration]. PMID- 10525648 TI - [Health reform 2000: hospitals must tighten their belts even more]. PMID- 10525649 TI - [What is ailing health reform proposal. Position of Bishop Voss on health care reform 2000]. PMID- 10525650 TI - [Unsecured e-mail communication between physicians is critically evaluated]. PMID- 10525651 TI - [ [In Process Citation] PMID- 10525652 TI - [ [In Process Citation] PMID- 10525653 TI - [Instead of ethics, monetics--or: the honest person is the fool. Comment on the reimbursement status]. PMID- 10525654 TI - Insulin analogues and their potential in the management of diabetes mellitus. PMID- 10525655 TI - Reduced prevalence of diabetes according to 1997 American Diabetes Association criteria. AB - AIMS/HYPOTHESIS: We examined the prevalence of diabetes and investigated the characteristics of subjects diagnosed by the American Diabetes Association and the World Health Organization criteria. METHODS: A total of 1235 Japanese Americans living in Hawaii and Los Angeles was studied. Of the subjects 114 were classified as previously diagnosed as having diabetes. A 75-g glucose tolerance test was given to the rest of the subjects. RESULTS: When abnormal glucose tolerance was diagnosed by the American Diabetes Association criteria, it was possible to identify only 40 % of diabetic subjects who had not been previously diagnosed compared with the current World Health Organization criteria based on a glucose tolerance test. In addition, the subjects identified by the American Diabetes Association criteria had higher glucose concentrations and had less insulin secretory capacity and they were in need of intensive treatment for diabetes. On the other hand, the subjects not diagnosed by the American Diabetes Association criteria alone were those whose glucose tolerance would be more likely to improve with lifestyle modification. CONCLUSION/INTERPRETATION: It might be better to use the fasting plasma glucose criterion advocated by the American Diabetes Association in combination with a glucose tolerance test after taking a detailed medical history. To reduce the number of subjects requiring the glucose tolerance test, priority should be given to subjects with impaired fasting glucose (6.1 His (CAG- >CAT), Exon 14: Val(648) -->Met (GTG-->ATG) and Arg(674)-->Cys (CGC--> TGC), and Exon 15: Ile(753)-->Leu (ATC-->CTC)]. The patients with Gln(108)-->His, Val(648)- >Met and Arg(674)-->Cys mutations, which may affect the E1P-E2P transition of SERCA3 during its enzyme cycle, had normal body weight with marked hyperglycaemia and beta-cell dysfunction. That is an unusual phenotype only found in 6 % of the Type II diabetic patients recruited for the UK Prospective Diabetes Study. In addition, five silent polymorphisms, six intron variants and two polymorphisms in the 3' untranslated region of exon 22 were found with similar frequency in diabetic and control subjects. CONCLUSION/INTERPRETATION: Our result suggests that in white Caucasians, the SERCA3 locus possibly contributes to the genetic susceptibility to Type II diabetes [Diabetologia (1999) 42: 1240-1243]. PMID- 10525667 TI - Search for variants of the gene-promoter and the potential phosphotyrosine encoding sequence of the insulin receptor substrate-2 gene: evaluation of their relation with alterations in insulin secretion and insulin sensitivity. AB - AIMS/HYPOTHESIS: The aim of this study was to screen part of the putative promoter sequence in addition to 14 potential phosphotyrosine residues of human IRS-2 for genetic variability which might cause changes in protein expression or function. Furthermore, the potential impact on insulin secretion and sensitivity of a previously identified IRS-2 variant (Gly1057Asp) was analysed. METHODS: The screenings were carried out by the SSCP-heteroduplex technique on DNA from Type II (non-insulin-dependent) diabetic patients. The impact of the Gly1057Asp variant was analysed in four glucose-tolerant Scandinavian study groups. RESULTS: The results showed no nucleotide substitutions in the promoter sequence, however, a novel heterozygous amino acid variant was identified (Leu647Val). In an association study, the new variant was found in 3 of 413 diabetic patients and in none of 280 glucose tolerant subjects. The variant did not affect the binding of IRS-2 to the insulin receptor or p85alpha of phosphatidylinositol 3-kinase when measured in the yeast two-hybrid system. Examination of the common Gly1057Asp variant in 363 young healthy subjects and in 228 glucose tolerant offspring of one diabetic parent showed no differences in insulin secretion or insulin sensitivity after an intravenous glucose tolerance test. Glucose tolerant middle aged subjects homozygous for the polymorphism (n = 31), however, had on average a 25 % decrease in fasting serum insulin concentrations (p = 0.009) and 28 % (p = 0.01) and 34 % (p = 0.003) reductions in serum insulin concentrations at 30 and 60 min, respectively, during an OGTT compared with wildtype carriers (n = 107). In a cohort of 639 elderly Swedish men the amino acid variant did not have any detectable impact on insulin secretion after an OGTT. CONCLUSION/INTERPRETATION: No genetic variability was found in the IRS-2 promoter. A rare IRS-2 variant at codon 647 has been identified in Type II diabetic patients. The prevalent codon 1057 polymorphism had no consistent effect on insulin secretion or insulin sensitivity. [Diabetologia (1999) 42: 1244-1249] PMID- 10525668 TI - Beta-fibrinogen gene G/A-455 polymorphism in relation to fibrinogen concentrations and ischaemic heart disease in Chinese patients with type II diabetes. AB - AIMS/HYPOTHESIS: We investigated the relation between the G/A-455 (Hae III) beta fibrinogen gene polymorphism and plasma fibrinogen concentration and its role in ischaemic heart disease in 264 Chinese patients with Type II (non-insulin dependent) diabetes mellitus and 182 non-diabetic control subjects. METHODS: The G/A-455 polymorphism was determined in genomic DNA using polymerase chain reaction and Hae III restriction enzyme digestion. Fibrinogen was measured with the Claus method. RESULTS: Fibrinogen concentrations were higher in diabetic patients (3.3 +/- 0.5 vs 2.5 +/- 0.9 g/l in controls, p < 0. 0001) and in women (p < 0.03 vs men). Allele frequency of the variant A allele was 27 % in both diabetic patients and control subjects' similar to findings in Caucasians. In control subjects, the AA genotype was associated with higher fibrinogen concentrations (2.8 +/- 0.38 g/l vs 2.5 +/- 0.5 in GG or GA, p < 0.03), contributing to 4 % of the variance in plasma fibrinogen. The genotype effect was smaller and not significant among non-smokers, women and diabetic patients. Higher fibrinogen concentrations and AA genotype frequency were found in diabetic patients with ischaemic heart disease (p < 0.05 and p < 0.005, respectively vs unaffected patients). In a multiple logistic regression model, AA genotype, age and mean arterial pressure were associated with ischaemic heart disease, with odds ratios of 4.19 (p < 0.01), 1.05 (p < 0.0001) and 1.03 (p < 0.03), respectively. CONCLUSION/INTERPRETATION: The G/A-455 polymorphism is a genetic determinant of fibrinogen concentrations and ischaemic heart disease in this Chinese cohort. It also interacts with environmental influences associated with smoking, the female sex and Type II diabetes in determining plasma fibrinogen concentrations. [Diabetologia (1999) 42: 1250-1253] PMID- 10525670 TI - Insulin resistance facilitates the development of coronary artery disease in Japanese type II diabetic patients: a single hospital-based follow-up study. PMID- 10525669 TI - Mechanism of protracted metabolic effects of fatty acid acylated insulin, NN304, in dogs: retention of NN304 by albumin. AB - AIMS/HYPOTHESIS: The provision of stable, reproducible basal insulin is crucial to diabetes management. This study in dogs examined the metabolic effects and interstitial fluid (ISF) profiles of fatty acid acylated insulin, Lys(B29) tetradecanoyl, des-(B30) human insulin (NN304). METHODS: Euglycaemic clamps were carried out under inhalant anaesthesia during equimolar intravenous infusions (3.6 pmol. min(-1) x kg(-1) for 480 min) of human insulin or NN304 (n = 8 per group). RESULTS: Steady-state total NN304 (albumin-bound and unbound) was considerably higher in plasma compared with human insulin (1895 +/- 127 vs 181 +/ 10 pmol/l, p < 0.001) and increased in interstitial fluid (163 +/- 14 vs 106 +/- 9 pmol/l, p < 0.01). The halftime for appearance of NN304 in interstitial fluid was slower than human insulin (92 vs 29 min, p < 0.001). Yet, equivalency of action was shown for glucose turnover; steady-state glucose uptake (Rd) of 7.28 +/- 0.55 and 6.76 +/- 0.24 mg. min(-1). kg(-1) and endogenous glucose production of 0.11 +/- 0.12 and 0.22 +/- 0.03 mg x min(-1) x kg(-1) (p > 0.40; NN304 and human insulin, respectively). Similar to interstitial fluid, half times for Rd and endogenous glucose production were delayed during NN304 infusion (162 vs 46 min and 80 vs 31 min, respectively; p < 0.01 vs human insulin). CONCLUSION/INTERPRETATION: Firstly equivalency of steady-state action is found at equimolar physiologic infusions of human insulin and NN304. Secondly NN304 binding to plasma albumin results in slower NN304 appearance in the interstitial compartment compared with human insulin. Thirdly the delay in appearance of NN304 in interstitial fluid may not in itself be a source of the protracted action of this insulin analogue. The protracted effect is due primarily to albumin binding of the insulin analogue NN304. [Diabetologia (1999) 42: 1254-1263] PMID- 10525671 TI - Absence of glutamic acid decarboxylase antibodies in Pima Indian children with diabetes mellitus. PMID- 10525672 TI - Characteristic neuroimaging findings in patients with diabetes and the 8296 mitochondrial tRNA(Lys) PMID- 10525673 TI - Cardiovascular risk factors in newly diagnosed abnormal glucose tolerance: comparison of 1997 ADA and 1985 WHO criteria. PMID- 10525675 TI - Sequence Characterization of a Unique Intergenic Spacer in Gadiformes Mitochondrial DNA. AB - : The nucleotide sequences of intergenic spacers located between the tRNA(Thr) and tRNA(Pro) genes in mitochondrial DNA of cod fishes (order Godiformes) were determined. Spacers from eight species representing two families of cod fishes were analyzed and found to vary in size from 25 to 99 bp. Each spacer sequence contains one or two copies of a conserved 17-bp motif. Four to five central nucleotides of this motif constitute a substitutional hot spot as observed from interspecific and intraspecific comparisons. The substitution rate of the spacer is approximately twice that of the variable part I of the mitochondrial DNA control region, making this sequence region interesting as a molecular marker in population studies or stock assessments of cod fishes. We propose that the spacer originated in a duplication event and evolved into a functional domain, perhaps by binding regulatory proteins. PMID- 10525676 TI - Organization of the Mitochondrial Genome of a Deep-Sea Fish, Gonostoma gracile (Teleostei: Stomiiformes): First Example of Transfer RNA Gene Rearrangements in Bony Fishes. AB - : We determined the complete nucleotide sequence of the mitochondrial genome (except for a portion of the putative control region) for a deep-sea fish, Gonostoma gracile. The entire mitochondrial genome was purified by gene amplification using long polymerase chain reaction (long PCR), and the products were subsequently used as templates for PCR with 30 sets of newly designed, fish universal primers that amplify contiguous, overlapping segments of the entire genome. Direct sequencing of the PCR products showed that the genome contained the same 37 mitochondrial structural genes as found in other vertebrates (two ribosomal RNA, 22 transfer RNA, and 13 protein-coding genes), with the order of all rRNA and protein-coding genes, and 19 tRNA genes being identical to that in typical vertebrates. The gene order of the three tRNAs (tRNA(Glu), tRNA(Thr), and tRNA(Pro)) relative to cytochrome b, however, differed from that determined in other vertebrates. Two steps of tandem duplication of gene regions, each followed by deletions of genes, can be invoked as mechanisms generating such rearrangements of tRNAs. This is the first example of tRNA gene rearrangements in a bony fish mitochondrial genome. PMID- 10525677 TI - Antisettlement and Narcotic Action of Analogues of Diterpene Marine Natural Product Antifoulants from Octocorals. AB - : Prevoius studies have determined that the octocorals Renilla reniformis and Leptogorgia virgulata contain diterpenes that are potent inhibitors of barnacle settlement. These antifoulants-the renillafoulins and pukalide-are, however, comparatively complex and thus are not amenable to commercial exploitation. The present study examined 19 analogues, based on the functional groups of lactone and furan rings in the parent molecules, for antisettlement activity and toxicity. The latter parameters are presented as EC(50) values for inhibition of cypris settlement and naupliar swimming, respectively. Assays of a subset of the analogues indicated that they were active in solution rather than when bound to a surface and that at relatively high concentrations they had a narcotic action. The mechanism whereby some of the analogues were able to inhibit settlement at nontoxic concentrations has yet to be explained but suggests that there is merit in the present approach to antifoulant development. PMID- 10525678 TI - Development of Polymorphic EST Markers Suitable for Genetic Linkage Mapping of Catfish. AB - : Expressed sequence tag (EST) markers are important for gene mapping and for marker-assisted selection (MAS). To develop EST markers for use in catfish gene mapping, 100 randomly picked complementary DNAs from the channel catfish (Ictalurus punctatus) pituitary library were sequenced. The EST sequences were used to design primers to amplify channel catfish and blue catfish (I. furcatus) genomic DNAs. Polymerase chain reaction products of the ESTs were analyzed to determine length polymorphism between the channel catfish and blue catfish. Eleven polymorphic EST markers were identified. Five of the 11 EST markers were from known genes and the other six were from unidentified ESTs. Seven ESTs were found to be associated with microsatellite sequences. Analysis of channel catfish gene sequences indicated highly biased codon usage, with 16 codons being preferably used. These codons were more preferably used in highly expressed ribosomal protein genes and in highly expressed pituitary hormone genes. G/C-rich codons are less used in channel catfish than those in other vertebrates suggesting AT-richness of the channel catfish genome. PMID- 10525679 TI - Green Fluorescent Protein As a Cell-Labeling Tool and a Reporter of Gene Expression in Transgenic Rainbow Trout. AB - : Green fluorescent protein (GFP) has been used as an indicator of transgene expression in living cells and organisms. For testing the utility of GFP in rainbow trout, we microinjected fertilized eggs with four types of supercoiled constructs containing two variants of GFP complementary DNA (S65T and EGFP), driven by two ubiquitous regulatory elements, human cytomegalovirus immediate early enhancer-promoter (CMV) and Xenopus laevis elongation factor 1alpha enhancer-promoter (EF1). Green fluorescence was first observed at 3 days postfertilization, when the embryo was in the mid-blastula stage. Fluorescence could be detected mosaically in various types of embryonic cells and tissues of swim-up fry. Both the percentage of fluorescent cells and the fluorescence intensity of GFP-expressing cells on blastoderms, measured with a microscopic photometry system, were highest in CMV-EGFP-microinjected embryos. We conclude that GFP is capable of producing detectable fluorescence in rainbow trout, and can be a powerful tool as a cell marker and reporter gene for cold-water fish, and that analysis of GFP expression in living cells is useful for characterizing the activity of cis-elements in vivo. PMID- 10525680 TI - Winter Flounder Expressed Sequence Tags: Establishment of an EST Database and Identification of Novel Fish Genes. AB - : An EST database of more than 900 sequences has been constructed from complementary DNAs from six different tissues (stomach, intestine, pyloric cecum, liver, spleen, and ovary) of the winter flounder Pleuronectes americanus. Template preparation and automated sequencing were optimized to generate high quality information in an economic fashion. Using computer scripts developed in our laboratory, the sequences were automatically compared with sequences in the databases via a Web-browser interface, and significant returns were recorded and organized on user-friendly HTML pages. Half (453) of the ESTs had significant matches to database sequences of known function, 33 matched ESTs from other organisms, 34 matched ribosomal RNAs, and 24 matched hypothetical open reading frames of unknown function. Forty-one percent (374) of the ESTs had no matches to sequences in the database and presumably represent previously unidentified cDNAs. Several sequences are the first isolated from teleost fish, and should be of interest for gene mapping and studies of developmental biology. PMID- 10525681 TI - Tissue-Specific Expressed Sequence Tags from the Black Tiger Shrimp Penaeus monodon. AB - : Expressed sequence tag data were generated from complementary DNA libraries created from cephalothorax, eyestalk, and pleopod tissue of the black tiger shrimp (Penaeus monodon). Significant database matches were found for 48 of 83 nuclear genes sequenced from the cephalothorax library, 22 of 55 nuclear genes from the eyestalk library, and 6 of 13 nuclear genes from the pleopod library. The putative identities of these genes reflected the expected tissue specificity. For example, genes for digestive enzymes were identified from the cephalothorax library and genes involved in the visual and neuroendocrine system from the eyestalk library. A few sequences matched anonymous EST or genomic sequences, and others contained mini-satellite or microsatellite repeat sequences. The remainder, 31 from the cephalothorax library, 25 from the eyestalk library, and 5 from the pleopod library, were sequences of high nucleotide complexity with no matches in any database searched and thus may represent novel genes. PMID- 10525682 TI - Sequences of 596 cDNA Clones (565,977 bp) of Japanese Flounder (Paralichthys olivaceus) Leukocytes Infected with Hirame Rhabdovirus. AB - : We have partially sequenced 785 sequences of 596 independent complementary DNA clones isolated from a cDNA library of Japanese flounder leukocytes infected with hirame rhabdovirus. These sequences consist of a total of 565,977 base pairs. The average size of the sequenced lengths was 721 bp. Of 596 clones, 386 (64.8%) were identified as previously reported genes by the BLASTN and BLASTX programs. About 30% of the identified clones could be recognized by only the BLASTX program. A total of 251 distinct genes were identified, and 181 of these genes are the first such genes reported from the teleostei. Approximately 27% of the identified Japanese flounder genes appear to be associated with cell division, cell structure or motility, and basic energy metabolism, 29% with gene or protein expression, and 17% with cell signaling, cell communication, and cell or organism defense. The most frequently identified expressed sequence tags of leukocytes of Japanese flounder were gelatinase b and ribosomal protein L23, which both had 1.34% prevalence. PMID- 10525683 TI - Secretagogues and Growth Factors in Fish and Crustacean Protein Hydrolysates. AB - : The search for new molecules in fish protein hydrolysates is of great interest in animal feeding as it is in aquaculture, fertilizer, cosmetic, and pharmacologic domains. Different sources of hydrolysates such as shrimp waste (Pandalus borealis), cod (Gadus morhua) head, and head and viscera of sardine (Sardina pilchardus), obtained after hydrolysis or autolysis, were tested on fibroblast cell cultures and by gastrin radioimmunoassay. The level of hydrolysis seems to play an important role in the presence of biological peptides. Elution profile on a gel filtration Sephadex G-50 column was used to estimate the degree of hydrolysis of the fractions studied. Growth-factor-like activities were found in less-hydrolyzed fractions. Conversely, the most-hydrolyzed fractions showed gastrin and cholecystokinin immunoreactivity. PMID- 10525684 TI - Bioprocess Intensification for Production of Novel Marine Bacterial Antibiotics Through Bioreactor Operation and Design. AB - : There is a lack of research into bioreactor engineering and fermentation protocol design in the field of marine bacterial antibiotic production. Most production strategies are carried out at the shake-flask level and lack a mechanistic understanding of the antibiotic production process, offering poor prospects for successful scale-up. This review shows that data need to be collated on media and physical optima differences between the trophophase and idiophase, along with investigations into the control mechanisms for biosynthesis, to allow implementation of novel fermentation protocols. Immobilization may play a part in bioprocess intensification of marine bacterial antibiotic production, through again this area is understudied. Similarly, mass transfer and shear stress data of fermentations are needed to provide the bioreactor design requirements to intensify antibiotic biosynthesis, with process scale-up in mind. The application of bioprocess intensification methods to the production of antibiotics (and other metabolites) from marine microbes will become an important strategy for improving supply of natural products, in order to assess their suitability as chemotherapeutic drugs. PMID- 10525685 TI - [Osteotomy of the hind-foot in children and adolescents]. AB - The position of the talus and the os calcaneum has consequences for the architecture of the forefoot. Medial tilting of the talus with vertical position provokes flattening of the medial arch and abduction of the forefoot. Varus position of the calcis and lateral direction of the talus (as in residual clubfoot), however, causes supination and adduction of the forefoot. Repair of deformities in the hindfoot therefore also influences the forefoot. A large number of various osteotomies have been proposed at both bones. The most popular ones are Dwyer's osteotomy at the corpus of the calcis and its modification described by Mitchell at the same location for correction of the cavus foot and the residual clubfoot, and the lengthening osteotomy at the neck of the calcaneum according to Evans. Indication and operative technique of these three procedures are described in detail. The closing wedge osteotomy according to Dwyer is indicated in cases of cavus feet. In clubfeet the os calcis is usually too short and the medial opening wedge osteotomy provokes skin problems. In these cases an osteotomy according to Mitchell with lateral displacement of the tuber calcanei is more suitable. In flat- and skew-feet a lengthening osteotomy at the neck of the calcaneum according to Evans can be indicated, especially when the load of the foot is bigger medially under the talus rather than at the lateral margin of the foot. If these procedures are carried out carefully, they have low complications rates. PMID- 10525686 TI - [Lateral column lengthening by calcaneal osteotomy combined with soft tissue reconstruction for treatment of severe posterior tibial tendon dysfunction. Methods and preliminary results]. AB - The purpose of this paper is to present principle and technique of proximal lateral column lengthening by calcaneal osteotomy and to critically analyze our preliminary results. 16 patients (7 female, 9 male; average age 52.3 years [24-72 years]) were treated for stage II to III posterior tibial tendon insufficiency by calcaneal osteotomy and medial soft tissue reconstruction (tendon reconstruction, 15; tendon transfer, 8; deltoid ligament repair, 10). When the AOFAS Ankle Hindfoot Rating Scale was applied, these patients were shown to have significantly increased their scores from an average preoperative value of 49.1 to a mean postoperative value of 91.1 after a mean follow-up of 24.6 months. In all but one case no loss of achieved foot correction was noted. In one case, a fusion of the calcaneocuboid joint had to be performed after 5 months due to painful degenerative joint disease. At follow-up, all patients had satisfactory restoration of their medial longitudinal arch, reduction of forefoot abduction, and restored arch height. All patients were able to fully weight-bear the operated foot, and all patients were satisfied with the achieved result. In the pes planovalgus deformity occurring in stage II to III (as significant degenerative joint disease has not already occurred), osteotomies appear to have a significant role in the operative management and to function by restoring more normal biomechanics, thus allowing tendon reconstruction and tendon transfers to return to successful function. PMID- 10525687 TI - [Arthrodesis of the talocalcaneal joint in adults. Indications, procedure, outcome]. AB - Affections of the rear-foot-complex may occur because of various etiologies and show differing pathogenetic patterns. Besides posttraumatic changes, the rheumatoid arthritis, primary disorders and neurologic complications are registered. Biomechanically the hindfoot-complex is characterised by the joint play of ligamentous, bony and tendineal structures. Referring to diagnostics the clinical and radiologic examination are predominant. In the literature there is no uniform opinion concerning the therapeutic algorithm. The arthrodesis as a surgical procedure is commonly used, nevertheless important differences may be stated with regard to the choice of the osteosynthesis or the number of the joints to be fused. We present biomechanical models, diagnostic examinations and operative procedures in this context. Our own results of patients who underwent surgical interventions involving joints of the lower rear-foot using the Kitaoka hindfoot-score are demonstrated. PMID- 10525688 TI - [Anatomic reconstruction of the lateral ligaments of the ankle using a plantaris tendon graft in the treatment of chronic ankle joint instability]. AB - The purpose of this work was to present our technique of anatomic reconstruction of the lateral ankle ligaments using a free plantaris tendon graft. Between 1988 and 1997, 52 ankles (48 patients) were treated for chronic ankle instability resisting to a training program of minimally 3 months. The average age was 28.6 years (16 to 46 years) at the time of surgery. There were 30 ankles in men and 22 ankles in females. 4 patients were treated on both ankles. 50 ankles were available for a follow-up investigation after a mean of 3.5 years (1 to 10 years). Based on the AOFAS-Hindfoot-Scale an average score of 97.9 points (90 to 100 points) was found. The functional result was found to be excellent in 39 ankles (78 %), good in 9 ankles (18 %), fair in 2 ankles (4 %), and poor in 0 ankle (0 %). Dorsi-/plantarflexion was not restricted in any ankle. Supination was slightly restricted in 2 ankles, but not increased in any ankle. High patient's satisfaction with respect to the achieved stability was found in all but one ankle. No deterioration with time was observed in any case. The overall good and excellent results with this method may be explained by the strictly anatomic reconstruction that did not alter the kinematics nor the mechanics of the ankle joint complex. In addition the peroneal tendons were not weakened. We feel that this procedure is a better alternative to other more complex ligament reconstructions, especially tenodesis operations by using the peroneal tendon. PMID- 10525689 TI - [Treatment of the ankle joint in rheumatoid arthritis with surgical and radiation synovectomy]. AB - Results of surgical synovectomy and radiation synovectomy (radiosynoviorthesis) of the tibiotalar joint in rheumatoid patients are reported. The staged concept for management of the rheumatoid ankle joint is presented which is based on the radiographic appearance of disease progression. Results of 16 rheumatoid patients with disease to the ankle joint suggest that pain and walking capability is positively influenced by synovectomy and radiosynoviorthesis. Follow-up of 30 months revealed no deterioration of postoperative clinical improvement. In the absence of contraindications to radiosynoviorthesis it is suggested to combine arthroscopic synovectomy with radiosynoviorthesis for the treatment of early stages of rheumatoid disease of the ankle joint. Open synovectomy is preferred to arthroscopic synovectomy, if tenosynovectomy is simultaneously required. PMID- 10525690 TI - [Short- and mid-term results with the STAR total ankle prosthesis]. AB - We evaluated the short- to mid-term results of an unconstrained total ankle prosthesis (S. T. A. R.) with uncemented fixation. Fifty consecutive ankle replacements were performed in 48 patients between 1996 and 1999. The initial diagnosis was posttraumatic osteoarthrosis in 31 cases (62 %), primary osteoarthrosis in 8 cases (16 %), and systemic joint affection in 11 cases (22 %), e. g. rheumatoid arthritis (6 cases), hemochromatosis (2 cases), psoriasis arthritis (1 case), lupus erythematodes (1 case), and sclerodermia (1 case). There were two perioperative complications: one superficial wound dehiscence that healed uneventfully, and one injury to the dorsal foot artery that necessitated primary reconstruction. Seven revisions, all in cases of posttraumatic arthrosis, were necessary: local revision of the fibula for painful lateral impingement (3 cases), posteromedial soft tissue revision for painful restriction of dorsiflexion (2 cases), percutaneous lengthening of the Achilles tendon (1 case), and osteotomy and callus distraction for angular correction after stress fracture of the distal tibia (1 case). At the last follow-up control, 21 patients (62 %) were very satisfied, 10 patients (29 %) were satisfied, and 3 patients (9 %) were satisfied with reservations. The obtained range of motion was 30 degrees (range, 15 to 55 degrees ), with a maximal plantarflexion of 25 degrees (range, 15 to 45 degrees ) and dorsiflexion of 5 degrees (-3 to 20 degrees ). When the AOFAS Hindfoot-Score was applied, the 34 patients scored 84.1 points (range, 44 to 100 points). After settling of the implants within 6 weeks, no migration was noted in any case, and all implants were considered to be stable. The favorable results were considered to be a result of the mechanical properties of the S. T. A. R. total ankle prosthesis that allows for unconstrained motion of the polyethylene inlay on the tibial component, e. g. anteroposterior translation, mediolateral translation and axial rotation. The success of implantation may depend on exact technique, correct hindfoot alignment, sufficient capsuloligamentous stability of the ankle, and a solid bone stock. Although our first results are very encouraging, a longer follow-up is mandatory to answer the question whether ankle replacement is a viable alternative to ankle arthrodesis. PMID- 10525691 TI - [The evolution of ankle arthroplasty]. AB - The results of ankle arthroplasty have generally been disappointing compared to other forms of arthroplasty. The present survey of results of ankle arthroplasty deals with the anatomical, the kinemasiological the biomechanical and the biological features of the ankle joint. These features all seem important prerequisites for a successful total ankle prosthesis design. From the study of the results displayed in the literature and those of my own series it can be stated that a modern ankle arthroplasty should respect the normal anatomy and kinemasiology. Biomechanically it should work as a normal ankle joint i. e. cylindrical mobility, congruency and a possibility for normal torque within the ankle mortise. The prosthesis components should only be fixed in solid subchondral bone, and preferably without cement. Only compressible forces should act at the bone-prosthesis interface. The axis of the ankle joint as well as of that of the hindfoot should be aligned to normal. Meticulous surgery and special guide instruments are absolute necessities. Restoration of muscle power and gait postoperatively are essential for a good and lasting result. The indication for ankle arthroplasty is mainly cases of osteoarthritis (primary or traumatic) and rheumatoid arthritis. Contraindications are talus necrosis, Charcot joints, extreme osteoporosis, severe arteriosclerosis, and very aggressive arthritis. Mental disorders and neurological disorders may also be contraindications. Furthermore, the patients should agree not to perform sports involving jumping and running or other ankle demanding activities. The average annual failure rate should not exceed 1 % if these recommendations are followed. PMID- 10525692 TI - [Arthroscopy for diagnosis and therapy of early osteoarthritis of the hip]. AB - Failure to conservative treatment in patients with less advanced radiographic signs of osteoarthritis of the hip (Danielsson grade 2-5) confronts with the decision of further treatment. Since radiographic imaging has not been proved very useful in demonstrating intraarticular structures and results of hip arthroscopies have been promising, arthroscopies have been performed in 17 hips from November 1997 to September 1998. Arthroscopic findings were exceeding preoperative imaging. In addition to cartilage degeneration, concomitant loose bodies, impinging osteophytes, degeneration of the labrum and synovial disease were found. Removal of loose bodies and osteophytes, partial resection of labral tears and partial synovectomy were performed. 1 month after arthroscopy (n = 15), mean Harris-Hip-Score was increased by 13 points und pain reduced by 39 % on average. 6 months after arthroscopy (n = 9), mean Harris-Hip-Score was increased by 14 points and pain reduced by 32 % on average. In addition to its therapeutic benefit, arthroscopy offers direct visualisation of the hip providing important information for the decision of further treatment. PMID- 10525693 TI - Meniscus repair. PMID- 10525694 TI - The arrow versus horizontal suture in arthroscopic meniscus repair. A prospective randomized study with arthroscopic evaluation. AB - In a prospectively randomized study including 68 patients, the results of inside out horizontal meniscus suturing were compared to meniscus repair using the meniscus arrow. 96% of the patients underwent re-arthroscopy after 3-4 months. Only lesions in the red/red or red/white areas were included. Patients were treated with a hinged brace for 9 weeks. 30 patients had an isolated bucket handle lesion. In 19 cases the repair was done in conjunction with an ACL reconstruction and in 19 cases the repair was performed in an ACL-insufficient knee. The two groups were comparable. Operating time in the arrow group was one half that of the suture group. Of 65 re-arthroscopies, 91% of the patients had healed or partially healed in the arrow group compared to 75% in the suture group (P = 0.11). In only 50% of the non-healed cases was this clinically suspected prior to control arthroscopy. The difference between healing in ACL-reconstructed and ACL-insufficient knees was not significant. Two patients in the suture group had a deep infection. There were no serious neurovascular injuries. Five patients in the suture group and two patients in the arrow group had symptoms in the saphenous nerve area. All patients had some synovial irritation at control arthroscopy but no severe reactions to suture or arrows were seen. Short-term results with meniscus arrows, based on healing and evaluated by second-look arthroscopy, seem promising. PMID- 10525695 TI - Magnetic resonance imaging of meniscal degeneration in torn menisci: a comparison between anterior cruciate ligament deficient knees and stable knees. AB - Signal anomalies observed in magnetic resonance imaging of the intrameniscal tissue adjacent to the tear were compared between stable knees (group 1, 54 menisci) and anterior cruciate ligament (ACL) deficient knees (group 2, 98 menisci). The histological significance of these signal anomalies was also studied (n = 25). The frequency of intrameniscal signal anomalies adjacent to the tear was significantly lower in ACL-deficient knees than in ACL-stable knees (P = 0.0022). There was a close correlation between the imaging anomalies and the presence of histological lesions (fissures, degeneration) within meniscal tissues adjacent to the tear (sensitivity: 0.95, specificity: 0.60). Our results suggest that the severity of intrameniscal degenerative changes adjacent to the tear are lower in ACL-deficient knees than in ACL-stable knees. PMID- 10525696 TI - The role of magnetic resonance imaging in routine decision making for meniscal surgery. AB - This clinical study evaluated factors affecting the decision for meniscal surgery in a patient population seen routinely at a trauma clinic. The study hypothesis was that patients who sustain a traumatic injury to the knee or have a long history of clinical symptoms are likelier to be operated on. We investigated 149 patients clinically and by magnetic resonance imaging (MRI). Group A (n = 62) underwent arthroscopic surgery and group B (n = 87) were treated conservatively. Multiple logistic regression analysis was used to examine correlations with regard to age, gender, injury pattern, period between the injury and first clinical examination, and MRI results. We found no significant difference between the two groups with regard to gender (P = 0.1), injury pattern (P = 0.44), or period between injury and first clinical examination (P = 0.5). Patients in group A were significantly older than those in group B (P = 0.044), and, as expected, MRI signal alterations were significantly higher in group A than in group B (P = 0.001). In acutely injured patients MRI helps to establish an accurate diagnosis, and in cases of positive MRI findings in a symptomatic patient, the surgeon should not wait 4-6 weeks but should immediately recommend surgery. PMID- 10525697 TI - Cyclops and cyclopoid formation after anterior cruciate ligament reconstruction: clinical and histomorphological differences. AB - Prospectively, 119 patients were pursued clinically and by follow-up-arthroscopy for the occurrence of a "cyclops syndrome" after ACL reconstruction with a patellar tendon autograft, augmented by LAD. Twenty-one patients showed nodular formations. Ten of these (group 1) developed early clinical evidence of a "cyclops syndrome" with a mean extension deficit of 19 degrees before follow-up arthroscopy, on average 5.9 months after the index operation. The nodular formations found and excised during debridement had a hard consistency. Histomorphological undecalcified microtome section evaluation of six specimens revealed fibrocartilagineous tissue with active bone formation in the center. The other 11 patients showed no clinical symptoms (group 2). A similar but soft nodulous scar formation was detected at follow-up-arthroscopy, on average 9.5 months after the index operation. Histomorphologically these so-called "cyclopoid" formations were only built-up fibrocartilagineous islands surrounded by granulation tissue. Neither remnants of tendon graft fibers nor old bone particles were found in specimens of either group. It can be concluded that both the hard cyclops and the soft "cyclopoid" are de novo scar formations. PMID- 10525698 TI - Bone tunnel enlargement after anterior cruciate ligament reconstruction with the hamstring autograft and endobutton fixation technique. A clinical, radiographic and magnetic resonance imaging study with 2 years follow-up. AB - The aim of this study was to describe the contrast-enhanced magnetic resonance imaging (MRI) appearance of bone tunnel enlargement detected on radiography after anterior cruciate ligament (ACL) reconstruction with semitendinosus and gracilis tendon endobutton (STG-endobutton) fixation technique. Fourteen patients with a STG-endobutton ACL reconstruction were examined 3 months (n = 1), 1 year (n = 1) and 2 years (n = 12) postoperatively. An age- and sex-matched group with a bone patellar tendon-bone (BTB) autograft ACL reconstruction with similar follow-up was taken as control. Data on clinical examination, laxity and isokinetic muscle torque measurements, anteroposterior and lateral view radiography were obtained, and knee scores (Lysholm and Tegner) were collected. Contrast-enhanced MRI was performed in the STG-endobutton group with a 1.5-T imager. There were no statistical differences between the groups with respect to clinical findings, stability tests, or knee scores. In the STG-endobutton group the average femoral and tibial bone tunnel diameter detected on anteroposterior view radiography had increased at 2-year follow-up by 33% and 23%, respectively. On MRI the ligamentous graft itself was not enhanced by the contrast medium whereas periligamentous tissue within and around the STG graft bundles showed mild contrast enhancement. In conclusion, the MRI results suggest that enhancing periligamentous tissue accumulated in and around the STG graft associated with the tunnel expansion. In spite of the significant bone tunnel enlargement observed on the follow-up radiography the STG-endobutton knees were stable and the patients satisfied. PMID- 10525699 TI - Comparison of polylactide screw and expansion bolt in bioabsorbable fixation with patellar tendon bone graft for anterior cruciate ligament rupture of the knee. A preliminary study. AB - In a preliminary study, 24 patients with rupture of the anterior cruciate ligament (ACL) were operated on using implants made of self-reinforced poly-l lactide (SR-PLLA). The operation method was outside-in bone-tendon-bone reconstruction. In 10 patients the fixation was made with an SR-PLLA screw with a diameter of 6.3 mm, in 12 with an SR-PLLA expansion plug with a diameter of 6.0 mm, and in two cases both implants were used, but these cases were excluded from comparison. The purpose of the study was to evaluate and compare the use and fixation results of these two implants. The follow-up time averaged 3.2 years. Twenty patients attended follow-up. On subjective evaluations, seven of the eight patients following SR-PLLA screw fixation and six of the ten after expansion plug fixation regarded their knee as normal or nearly normal. Arthrometric testing showed the side-to-side difference to average 2. 9 mm following SR-PLLA screw fixation and 2.6 mm after expansion plug fixation (NS). Six of the patients had giving-way symptoms (two after screw fixation and four after plug fixation). The pivot shift test was slightly positive in two patients and positive in one patient after SR-PLLA screw fixation, and in three knees slightly positive and in another three knees positive following expansion plug fixation. Radiography showed variation in the location and orientation of the bone channels. Magnetic resonance imaging was performed in seven cases, and in two cases an edema was found in the tendon of the anterior cruciate ligament graft and in six cases the implants were visible. No statistical difference in results between the SR-PLLA screw and SR-PLLA expansion bolt was noted. Fixation with expansion plug seems technically more challenging, with a tendency to inferior results compared to screw fixation. In the absorbable fixation of a bone-tendon-bone graft there are no metallic artifacts on magnetic resonance imaging and no need to remove the fixation material regarding the revision surgery. PMID- 10525700 TI - Proprioception after anterior cruciate ligament reconstruction with and without bracing. AB - In this investigation we evaluated the effect of ACL reconstruction and functional knee bracing on knee proprioception. Twenty subjects who experienced acute ACL disruption and underwent reconstruction with a bone-patellar tendon bone graft participated in a controlled rehabilitation program and were studied at a mean follow-up of 2 years. A control group of ten subjects were also studied. In both groups proprioception was evaluated by measuring the threshold to detection of passive motion (TDPM) with the knee at 15 degrees of flexion with and without a functional knee brace applied. The Knee Osteoarthritis Outcome Score, Cincinnati knee score, and two functional knee tests were also used as outcome measurements. Anterior-posterior displacement of the tibia relative to the femur was evaluated with the KT-1000 arthrometer. There were no significant differences in TDPM between the ACL-reconstructed and contralateral knees, or between the ACL reconstructed group and the healthy control group. Bracing did not produce a significant change in the TDPM for the ACL-reconstructed group or for the control group. There were low to moderate correlations between TDPM and the other outcome measurements. This study indicates that there is no significant differences in proprioception between the ACL-reconstructed knee and the contralateral uninvolved knee 1 year or more after surgery. Functional knee bracing does not seem to improve proprioception in patients who have undergone ACL reconstruction and been followed up on average 2 years after surgery. PMID- 10525701 TI - Proprioception in the posterior cruciate ligament deficient knee. AB - This study was undertaken to evaluate knee proprioception in patients with isolated unilateral posterior cruciate ligament (PCL) injuries. Eighteen subjects with isolated PCL tears were studied 1-234 months after injury. The threshold to detect passive motion (TTDPM) was used to evaluate kinesthesia and the ability to passively reproduce passive positioning (RPP) to test joint position sense. Two starting positions were tested in all knees: 45 degrees (middle range) and 110 degrees (end range) to evaluate knee proprioception when the PCL is under different amounts of tension. TTDPM and RPP were tested as the knee moved into flexion and extension from both starting positions. A statistically significant reduction in TTDPM was identified in PCL-injured knees tested from the 45 degrees starting position, moving into flexion and extension. RPP was statistically better in the PCL-deficient knee as tested from 110 degrees moving into flexion and extension. No difference was identified in the TTDPM starting at 110 degrees or in RPP with the presented angle at 45 degrees moving into flexion or extension. These subtle but statistically significant findings suggest that proprioceptive mechanoreceptors may play a clinical role in PCL-intact and PCL deficient patients. Further, it appears that kinesthesia and joint position sense may function through different mechanisms. PMID- 10525702 TI - Long-term results of tendon allografts for anterior cruciate ligament replacement in revision surgery and in cases of combined complex injuries. AB - We assessed the long-term validity of anterior cruciate ligament (ACL) reconstruction using tendon allografts. Nineteen patients were followed up for 8 years (mean 94 months) after tendon allograft replacement for ACL rupture. The evaluation used the International Knee Documentation Committee (IKDC) grades, the Lysholm score, and the Tegner scale. Two patients sustained a rerupture after a serious injury. Two others scored poorly because of associated proximal ipsilateral tibial and other fractures (IKDC grade D). Nine patients scored nearly normal (grade B) and six abnormal (grade C). The Lysholm score showed nine excellent (average 98), five good (average 87), and one fair result (70). Two of the excellent-rated patients were IKDC grade C solely because the X-rays showed a slight (1-mm) narrowing of the medial cartilage. One patient had 0.5-mm narrowing. The X-ray findings may indeed indicate future problems. On the Tegner scale the sports level decreased by an average of 2.1 points (from 6.87 to 4.73), and by 0.8 point compared to the level at which the patient had wished to perform. ACL repair using tendon allografts appears to provide satisfactory results on the Lysholm and Tegner scales. The IKDC scoring suggests future cartilage degeneration. Its value for knees with multiple ligament lesions and for revision cases is demonstrated. Only two reruptures were noted, suggesting good reliability of allografts on the long term. PMID- 10525704 TI - Midterm results of arthroscopic treatment of scapholunate ligament lesions associated with intra-articular distal radius fractures. AB - Since 1993, we have treated 30 patients with acute intra-articular distal radius fractures using arthroscopic assistance. Concomitant lesions of the intrinsic scapholunate (SL) ligaments were diagnosed in 12 patients (40%). Using the grading system of Geissler et al. [13], the identified lesions included a single grade I tear, three grade II, six grade III, and two grade IV. The grade III and IV lesions were accompanied by intraoperative findings of marked instability. Therefore operative stabilization was performed by temporary scapholunate and scaphocapitate arthrodesis. Seven patients in this group (87.5%) were followed up clinically and radiologically for an average of 3 years postoperatively. Clinical examination included range of motion and a subjective questionnaire concerning pain and ability to work. Objective grip strength was measured using a Jamar tester and compared to the contralateral wrist. Radiological evaluation consisted of posteroanterior and lateral views and of stress views in radial and ulnar deviation. Data were evaluated by the scoring systems of Jakim et al. [21], Cooney et al. [5] and by the demerit point system of Gartland and Werley [12], as modified by Sarmiento et al. [36]. An excellent result was present in 100% of our patients by the Gartland and Werley system, in 86% by that of Jakim et al., and in 60% by that of Cooney et al. Based on a subjective questionnaire, all of the patients had an excellent or good result. PMID- 10525703 TI - Knee arthroscopy in local versus general anaesthesia. The incidence of rearthroscopy. AB - The choice of anaesthesia in routine knee arthroscopy varies considerably. Concerns about local anaesthesia include the fear that it will take longer to perform surgery and that the anaesthesia will be inadequate, leading to patient discomfort. In this study, data from all patients (n = 6519) who had undergone a knee arthroscopy at St Goran Hospital Artro Clinic, in Stockholm, Sweden, during a 3.5 year period, between January 1993 and July 1996, were reviewed. Of these 6519 primary arthroscopies, 4101 were performed under local anaesthesia and 2418 under general anaesthesia. The purpose of the study was first to identify those arthroscopies that could not be successfully performed because the local anaesthesia was inadequate, and second, to investigate if arthroscopy under local anesthesia was associated with an increased number of rearthroscopies compared to general anaesthesia. The total number of rearthroscopies, performed within 180 days from the primary arthroscopy, was 214. Of these 214 rearthroscopies, 146 were due to a new indication for surgery and 30 were due to persisting clinical symptoms (true rearthroscopies). The remaining 38 rearthroscopies were due to an incomplete examination (because of patient discomfort) in a primary procedure where local anaesthesia was used. Of the 30 true rearthroscopies, 19 originated from the 4101 primary arthroscopies performed under local anaesthesia (0.46%) and 11 originated from the 2418 primary arthroscopies performed under general anaesthesia (0.45%). It is concluded that 0.9% of the primary arthroscopies performed under local anaesthesia could not be performed safely due to patient discomfort. There was no difference in the frequency of rearthroscopy between the arthroscopies performed under local anaesthesia compared to those performed under general anaesthesia. PMID- 10525705 TI - A technical solution for secondary arthritis due to chronic proximal tibiofibular joint instability. AB - Chronic instability of the proximal tibiofibular joint is an uncommon diagnosis and not frequently reported in the literature. The management options of this joint instability, complicated with secondary arthritis, have rarely been discussed and consist mainly of fibular head resection or arthrodesis of this joint. We describe a new technical procedure for addressing both the instability and the joint secondary arthritis. Stability of the joint is achieved by ligament reconstruction using a biceps femoris split passed through the tibial metaphysis and fixated back to the fibular head using bone anchors. The arthritic changes are addressed by interposition of a vascularized fascia lata strip. The described procedure offers a firm stabilization with no need for postoperative restrictions and an alternative to the inadvisable joint arthrodesis or resection. PMID- 10525706 TI - Comment on W. Petersen and B. Tillmann, "Blood and lymph supply of the posterior cruciate ligament". PMID- 10525707 TI - Rejoinder to R. scapinelli PMID- 10525708 TI - Vitamin D receptor gene polymorphisms, bone mass, bone loss and prevalence of vertebral fracture: differences in postmenopausal women and men. AB - Bone mineral density (BMD), the major determinant of fracture risk, is under strong genetic control. Although polymorphisms of the vitamin D receptor (VDR) gene have been suggested to account for some of the genetic variation in bone mass, the influence of VDR genotypes on osteoporosis remains controversial. Previous published studies have focused mainly on women, but the pattern of response in men has not been determined. Using the BsmI restriction enzyme, we studied the influence of the different VDR genotypes on bone mass, bone loss and the prevalence of vertebral fractures in a population-based sample of both sexes (n = 326). BMD was measured at the lumbar spine and femoral neck, with a 4-year interval, using dual-energy X-ray absorptiometry. Vertebral fractures were assessed by two lateral radiographs at the beginning and end of the study. The prevalence of the three possible VDR genotypes was similar to those in other Caucasian populations and no differences were found between men and women. Women with the favorable bb genotype showed significantly higher BMD values at the lumbar spine and femoral neck, and a positive rate of BMD change at the femoral neck compared with women with the BB and Bb genotypes. Moreover, women with the bb genotype showed a trend toward a lower prevalence and incidence of vertebral fractures (p = 0.07). We have not found any differences between VDR genotypes in men. In conclusion, VDR gene polymorphisms are related to bone mass and bone loss in women; also a trend in the prevalence of vertebral fractures was observed in postmenopausal women but not in men. PMID- 10525709 TI - A double-masked multicenter comparative study between alendronate and alfacalcidol in Japanese patients with osteoporosis. The Alendronate Phase III Osteoporosis Treatment Research Group. AB - To evaluate the efficacy and safety of alendronate, a double-masked, active (alfacalcidol) controlled comparative study for 48 weeks was carried out in a total of 210 Japanese patients with osteoporosis. The doses of alendronate and alfacalcidol were 5 mg/day and 1 microgram/day, respectively. The lumbar bone mineral density (LBMD) values observed at 12, 24, 36 and 48 weeks after the initiation of alendronate treatment were 3.53 +/- 0.53%, 5.37 +/- 0.62%, 5.87 +/- 0.74% and 6.21 +/- 0.59% (mean +/- SE), respectively, higher than the baseline value. Corresponding values in the alfacalcidol group were 1.50 +/- 0.43%, 0.69 +/- 0.63%, 1.12 +/- 0.60% and 1.36 +/- 0. 63%, respectively. There was a significant difference between the two groups at each time point (p<0.05 or p<0.001). The bone turnover markers were depressed during treatment in the alendronate group: -32.2% for alkaline phosphatase, -53.7% for N-terminal osteocalcin and -45.0% for urinary deoxypyridinoline compared with the corresponding baseline values. On the contrary, no notable changes in these parameters were observed in the alfacalcidol group. Treatment with alendronate caused a transient decrease in serum calcium concentrations associated with an increase in the serum level of intact parathyroid hormone. In contrast, treatment with alfacalcidol resulted in a tendency of these parameters to change in the opposite direction. No difference in fracture incidence between the two groups was observed. The overall safety of alendronate was comparable to that of alfacalcidol. In conclusion, although it was a relatively short-term study of 48 weeks, the results of the present study indicate that alendronate at the daily dose of 5 mg was effective in increasing LBMD and that no serious drug-related adverse events were observed in the alendronate-treated patients. Alendronate is more efficacious than alfacalcidol in increasing bone mineral density, although the mechanisms of the actions of the two drugs are apparently different. PMID- 10525710 TI - Effect and offset of effect of treatments for hip fracture on health outcomes. AB - We investigated the cost-effectiveness of treatments that reduce the risk of hip fracture using a computer simulation model. Cost-effectiveness was measured as cost per quality-adjusted life-year (QALY) gained using a threshold value for cost-effectiveness of $30,000/QALY gained. The baseline simulations assumed a 5 year intervention that reduced the risk of hip fracture by 50% during the intervention period, and an effect which reversed to the pretreatment risk during the next 5 years. Sensitivity analyses included the effects of age, different fracture risks, and different treatment costs and duration of therapeutic effect once treatment was stopped. Cost-effectiveness was critically dependent upon absolute risk determined by the age and the relative risk of hip fracture at any given age. Reasonable cost-effectiveness was shown even with relatively high intervention costs for women with a risk about twice the average at the age of 70 or more years. Cost-effectiveness was critically dependent upon the assumptions made concerning offset of effect of intervention after the end of treatment. Where no residual effect was assumed, it was difficult to show cost-effectiveness from any intervention except for the most effective and least expensive. Conversely, cost-effectiveness improved considerably where effectiveness persisted for a longer time. These studies support the view that intervention in the elderly with agents affecting skeletal metabolism alone may be preferred to such interventions at the time of the menopause, and that offset time, hitherto poorly characterized, is a critical component of cost-effectiveness, particularly in younger women. PMID- 10525711 TI - Bone mineral measurements: a comparison of delayed gamma neutron activation, dual energy X-ray absorptiometry and direct chemical analysis. AB - A system in vitro consisting of a femur from a cadaver and soft-tissue equivalent material was used to test the agreement between several techniques for measuring bone mineral. Calcium values measured by delayed gamma neutron activation (DGNA) and bone mineral content (BMC) by Lunar, Hologic and Norland dual-energy X-ray absorptiometers (DXA) were compared with calcium and ash content determined by direct chemical analysis. To assess the effect of soft-tissue thickness on measurements of bone mineral, we had three phantom configurations ranging from 15.0 to 26.0 cm in thickness, achieved by using soft-tissue equivalent overlays. Chemical analysis of the femur gave calcium and ash content values of 61.83 g +/- 0.51 g and 154.120 +/- 0.004 g, respectively. Calcium measured by DGNA did not differ from the ashed amount of calcium at any of the phantom configurations. The BMC measured by DXA was significantly higher, by 3-5%, than the amount determined by chemical analysis for the Lunar densitometer and significantly lower, by 3-6%, for the Norland densitometer (p<0.001-0.024), but only 1% lower (not significant) for the Hologic densitometer. DXA instruments showed a decreasing trend in BMC as the thickness increased from 20.5 to 26.0 cm (p<0.05). However, within the entire thickness range (15.0-26.0 cm), the overall influence of thickness on BMC by DXA was very small. These findings offer insight into the differences in these currently available methods for bone mineral measurement and challenge the comparability of different methods. PMID- 10525712 TI - Development and validation of the mini-osteoporosis quality of life questionnaire (OQLQ) in osteoporotic women with back pain due to vertebral fractures. Osteoporosis Quality of Life Study Group. AB - The objective of the study was to evaluate a shortened osteoporosis quality of life questionnaire (OQLQ) in osteoporotic women with back pain due to vertebral fractures. From the longer 30-item OQLQ (four to nine items per domain) we created the mini-OQLQ by choosing the two items with the highest impact in each of five domains (symptoms, physical function, activities of daily living, emotional function, leisure). We administered the OQLQ, the Sickness Impact Profile, the SF-36 and the Brief Pain Index to patients at baseline, after 2 weeks and after 6 months. The intraclass correlations between baseline and the 2 week follow-up for the five mini-OQLQ domains ranged from 0.72 to 0.86. Cross sectional correlations between the domains of the mini-OQLQ and other health instruments were moderate to large (0.35-0.80) and greater than predicted. The mini-OQLQ items showed moderate to large correlations with items omitted from the shortened questionnaire (0. 44-0.88). Correlations between the OQLQ domains and the other three instruments were greater than those of the mini-OQLQ, and partial correlations between OQLQ items omitted from the mini-OQLQ and the other three instruments after considering mini-OQLQ items were substantial (0.19-0.71) and statistically significant. Sample sizes of less than 200 per group should be required to detect minimally important differences in parallel-group clinical trials. Longitudinal correlations between the mini-OQLQ and the other measures were often significant but generally lower than predicted (0.10-0.49). The partial correlations revealed that the omitted items explained a significant portion of the longitudinal variance in each domain. We conclude that in a selected group of patients with back pain caused by vertebral fractures, the mini OQLQ demonstrated good discriminative and adequate evaluative properties. The mini-questionnaire should be useful in clinical settings. PMID- 10525713 TI - Vertebral fractures predict subsequent fractures. AB - This population-based study documents an increase in most types of fractures following the occurrence of a clinically recognized vertebral fracture among 820 Rochester, Minnesota, residents. During 4349 person-years of follow-up, 896 new fractures were observed. Relative to incidence rates in the community, there was a 2.8-fold increase in the risk of any fracture, which was greater in men (standardized incidence ratio (SIR), 4.2; 95% CI, 3.2-5.3) than women (SIR, 2.7; 95% CI, 2.4-3.0). The estimated cumulative incidence of any fracture after 10 years was 70%. The greatest increase in risk was for subsequent fractures of the axial skeleton, in particular a 12.6-fold increase (95% CI, 11-14) in additional vertebral fractures. There was a lesser increase in most limb fractures, including a 2.3-fold increase (95% CI, 1.8-2.9) in hip fractures and a 1.6-fold increase (95% CI, 1.01-2.4) in distal forearm fractures. There was a slightly greater association with distal forearm fractures among those whose first vertebral fracture occurred before age 70 years but a similar relationship with hip fractures, including cervical and intertrochanteric hip fractures separately, regardless of age at the initial vertebral fracture. There was also an equivalent increase in subsequent fracture risk whether the initial vertebral fracture was attributed to severe or moderate trauma. These data show that vertebral fractures represent an important risk factor for fractures in general, not just those of the spine and hip. PMID- 10525714 TI - Vitamin D status during puberty in French healthy male adolescents. AB - The vitamin D status was determined on one to four occasions either after summer (September-October) or after winter (March-April) in 175 male adolescents (13-17 years), resulting in 394 measurements of serum 25-hydroxyvitamin D (25(OH)D) and intact parathyroid hormone (iPTH). The subjects lived in a rural area to the north of Paris (49 degrees N). After summer the 25(OH)D concentration was 58.5 +/ 18.0 nmol/l (mean +/- SD), while after winter it had fallen to 20.6 +/-6.0 nmol/l (p = 0.0001). Meanwhile the iPTH concentration was 2.76 +/- 0.97 pmol/l (mean +/- SD) after summer and increased to 4.20 +/- 1.21 pmol/l after winter (p = 0. 0001). All the results were pooled and a nonlinear population model with random parameters was used to describe the relationship between serum iPTH and 25(OH)D. When the concentration of 25(OH)D was higher than 83 nmol/l, an iPTH mean 'plateau' level at 2.48 pmol/l was reached. When 25(OH)D concentrations fell below 83 nmol/l, the increase in iPTH concentration accelerates, and when the mean 25(OH)D concentration was equal to or lower than 10 nmol/l the mean iPTH level (4.97 pmol/l) was twice as high as the 'plateau' value. PMID- 10525715 TI - Vitamin D receptor gene polymorphisms and bone mineral density in elderly Chinese men and women in Hong Kong. AB - Although genetic factors have been strongly implicated in determining bone mineral density (BMD), the role of the vitamin D receptor (VDR) polymorphism remains controversial. An overall consensus is difficult, as the populations studied have been heterogeneous with respect to menopausal status and ethnicity. Moreover, some studies have examined only small populations, and relatively few studies have been conducted in Asian populations. There is mounting evidence that calcium homeostasis in Asian populations differs from that in Caucasians. This difference may be mediated, in part, through VDR effects. In a cross-sectional study we have examined the relationship between the VDR polymorphism and BMD in 272 women (mean age 75 years) and 237 men (mean age 73 years) of Chinese origin from Hong Kong. Consistent with other studies in Asian populations we found higher frequencies of the T, b and a alleles compared with those reported in Caucasian populations. Moreover, no significant difference in BMD was observed when subjects were grouped by a combination of the genotypes (bbAATT, bbAaTT, bbaaTT, BbAaTt, BbAATt). These results suggest that VDR polymorphism is not associated with BMD in elderly Hong Kong Chinese men and women. PMID- 10525716 TI - Trabecular bone architecture in the distal radius using magnetic resonance imaging in subjects with fractures of the proximal femur. Magnetic Resonance Science Center and Osteoporosis and Arthritis Research Group. AB - To determine whether magnetic resonance (MR)-derived measures of trabecular bone architecture in the distal radius are predictive for prevalent hip fractures, 20 subjects with hip fractures and 19 age-matched postmenopausal controls were studied. Bone mineral density (BMD) measures at the hip (dual-energy X-ray absorptiometry, DXA) and the distal radius (peripheral quantitative computed tomography, pQCT) were also obtained. We compared the MR-based structural measures derived in the radius with those in the calcaneus of the same patients. In the radius, images were acquired at an in-plane resolution of 156 microm and a slice thickness of 0.5 mm. Stereologic measures such as the apparent trabecular thickness (app. Tb.Th), fractional trabecular bone volume (app. BV/TV), trabecular spacing (app. Tb.Sp) and trabecular number (app. Tb.N) were derived from the images. Measures of app. Tb.Sp and app. Tb.N in the distal radius showed significant (p<0.05) differences between the two groups, as did hip BMD measures. However, radial trabecular BMD measures showed only a marginal difference (p = 0.05). Receiver operating curve analysis was used to determine the diagnostic efficacy of BMD, structural measures and a combination of the two. The area under the curve (AUC) for total hip BMD was 0.73, and for radial trabecular BMD was 0.69. AUC for most of the measures of trabecular bone structure at the distal radius was lower than for hip BMD measures; however, AUC for app. Tb.N at the radius was 0.69, comparable to trabecular BMD using pQCT. The AUC for combined BMD (hip) and structure measures was higher (0.87) when radius and calcaneus structure was included. Measures of trabecular architecture derived from MR images combined with BMD measures improve the discrimination between subjects with hip fractures and normal age-matched controls. PMID- 10525717 TI - Age- and gender-specific rate of fractures in Australia: a population-based study. AB - There is little population-based data concerning fracture rates in Australia. We ascertained all fractures occurring during 2 years in adults aged 35 years and over residing within a defined region (population 218 000), representative of the Australian population. The major strength of this study is the comprehensive ascertainment of fractures, which was ensured by regular searches of the only two radiologic providers in the Geelong Osteoporosis Study region. Nevertheless, vertebral fractures are likely to be underestimated since our ascertainment relied on a clinical indication for a medical imaging procedure. Among those aged 35-55 years, the fracture rate (persons per 10,000/year) in men was about double the rate in women (65 vs 35). The fracture rate was almost 7 times higher in women over 60 years versus women less than 55 years of age. In contrast, the fracture rate in men over 60 years was only 50% higher than in men less than 55 years of age (72 vs 104). Fracture rates in women and men were highest at the hip (28 and 10 respectively), spine (21 and 7), distal forearm (Colles') (18 and 4) and humerus (11 and 3), and were 3-4 times higher in women than men. These fractures accounted for 63% of all fractures in women and 32% in men. By contrast, the rate of lower leg and ankle fractures was less than 10 per 10,000 in both women and men and did not increase to the same extent with age. Hip fracture rates appear high, particularly among the older age strata, compared with retrospective ascertainment in other populations. In Australia, as in many other countries, there is an increasing longevity of the population. The number of women aged 90 years and over increased by 32% and the number of men of this age increased by 48% in the 5 years between the Australian national census of 1991 and 1996. Given stable fracture rates, the substantial health burden imposed by age-related fractures, particularly hip fractures, will continue to escalate in both women and men. PMID- 10525719 TI - Epidemiology of osteoporosis. PMID- 10525718 TI - Intracapsular hip fracture: increased cortical remodeling in the thinned and porous anterior region of the femoral neck. AB - It has been shown previously that the antero-inferior cortex is subjected to maximal tensile stress during a fall onto the greater trochanter. We have recently shown that in cases of femoral neck fracture, cortical thinning and porosity is greatest in the anterior and antero-inferior region of the femoral neck. To investigate whether this is due to increased remodeling, we have quantified surface-based parameters associated with Haversian remodeling in femoral neck biopsies from women with intracapsular hip fracture and post-mortem controls. Cryostat sections of chilled biopsies were reacted for either tartrate resistant acid phosphatase (TRAP) or alkaline phosphatase (ALP) activity. Proportions of active canals were determined in each quadrant (inferior, anterior, superior, posterior) of the femoral neck. The biopsies were then embedded in methacrylate to permit histomorphometry using Goldner's and Solochrome sections. In the cases there was no significant increase in the proportion of canals undergoing remodeling in the cortex as a whole (p = 0.846), but the regional distribution of remodeling was markedly different from that in the controls. In the anterior cortex, the proportion of canals undergoing remodeling was increased by 56% (p = 0.0087); in contrast there was a relative decrease of 35% in the superior region (p = 0.0047). In the anterior cortex of cases there were 76% and 42% increases in the proportions of eroded (p = 0.019) and osteoid-bearing (p = 0.041) canals, respectively. In the superior region, the decrease in the proportion of remodeling sites was due to a marked decrease in canals with an osteoid surface (51%; p = 0.0031). Covariance analysis with cortical porosity as the dependent variable showed that porosity was significantly dependent on the regional distribution of eroded (p = 0.033) but not on the distribution of forming (p = 0.153) canals (R(2)adj = 0.51). Cellular levels of TRAP and ALP were significantly elevated in the anterior region of cases compared with the controls (TRAP 55%, p = 0.006; ALP 36%, p = 0.003). For the posterior and inferior regions there were no marked differences in cellular TRAP and ALP levels compared with control values. These data show that the increased cortical thinning and increased porosity we have previously observed in the anterior cortex in cases of hip fracture are associated with increased indices of Haversian remodeling. These findings are consistent with the hypothesis that, in cases of hip fracture, remodeling imbalance in the anterior cortex is a continuing process up to the time of fracture and is due to increased osteoclastic cellular activity associated with an osteoblastic response that is inadequate to prevent bone loss. PMID- 10525720 TI - Risk factors for hip fracture not related to bone mass and their therapeutic implications. PMID- 10525721 TI - Determinants of peak bone mass and mechanisms of bone loss. PMID- 10525722 TI - Clinical assessment of bone mass, quality and architecture. PMID- 10525723 TI - Clinicial utility of biochemical markers. PMID- 10525724 TI - How should the risk of fracture in postmenopausal women be assessed? PMID- 10525725 TI - What can we learn from bone biology for the treatment for osteoporosis? PMID- 10525726 TI - Calcium, vitamin D and vitamin K in the prevention of fractures due to osteoporosis. PMID- 10525727 TI - An assessment of hormone replacement therapy to prevent postmenopausal osteoporosis. PMID- 10525728 TI - Sex steroid analogs. PMID- 10525729 TI - Bisphosphonates: pharmacology, mechanisms of action and clinical uses. PMID- 10525730 TI - Calcitonin, bone-active isoflavones and vitamin D metabolites. PMID- 10525731 TI - Treatment of osteoporosis: role of bone-forming agents. PMID- 10525732 TI - Osteoporosis in men. PMID- 10525733 TI - Cost-effective treatment strategies for osteoporosis. PMID- 10525734 TI - Ultraviolet and osmotic stresses induce and regulate the synthesis of mycosporines in the cyanobacterium chlorogloeopsis PCC 6912 AB - The cyanobacterium Chlorogloeopsis PCC 6912 was found to synthesize and accumulate two putative UV sunscreen compounds of the mycosporine (mycosporine like amino acid; MAA) type: mycosporine-glycine and shinorine. These MAAs were not constitutively present in the cells; their synthesis could be induced specifically either by exposure to UVB radiation (280-320 nm) or by osmotic stress, but not by other stress factors such as heat or cold shock, nutrient limitation, or photooxidative stress. A significant synergistic enhancement of MAA synthesis was observed when both stress factors were applied in combination. Although osmotic stress could induce MAA synthesis, comparison of the intracellular contents of MAAs with those of sugar osmolytes (glucose and trehalose) indicated that MAAs play no significant role in attaining osmotic homeostasis. UVB strongly enhanced the accumulation of shinorine, whereas osmotic stress had a more pronounced effect on mycosporine-glycine. This differential effect on the steady-state contents of each MAA could be explained either by differential regulation of biosynthesis or by differential loss rates of MAAs (leakage) under each condition. A preferential leakage of mycosporine-glycine from the cells after a hypoosmotic shock was detected. The results are interpreted in terms of an adaptive necessity for a combined regulatory control responding to both UV and external osmotic conditions in organisms that accumulate water-soluble sunscreens intracellularly. PMID- 10525735 TI - Physiology, phylogenetic relationships, and ecology of filamentous sulfate reducing bacteria (genus desulfonema) AB - Microscopy of organic-rich, sulfidic sediment samples of marine and freshwater origin revealed filamentous, multicellular microorganisms with gliding motility. Many of these neither contained sulfur droplets such as the Beggiatoa species nor exhibited the autofluorescence of the chlorophyll-containing cyanobacteria. A frequently observed morphological type of filamentous microorganism was enriched under anoxic conditions in the dark with isobutyrate plus sulfate. Two strains of filamentous, gliding sulfate-reducing bacteria, Tokyo 01 and Jade 02, were isolated in pure cultures. Both isolates oxidized acetate and other aliphatic acids. Enzyme assays indicated that the terminal oxidation occurs via the anaerobic C(1) pathway (carbon monoxide dehydrogenase pathway). The 16S rRNA genes of the new isolates and of the two formerly described filamentous species of sulfate-reducing bacteria, Desulfonema limicola and Desulfonema magnum, were analyzed. All four strains were closely related to each other and affiliated with the delta-subclass of Proteobacteria. Another close relative was the unicellular Desulfococcus multivorans. Based on phylogenetic relationships and physiological properties, Strains Tokyo 01 and Jade 02 are assigned to a new species, Desulfonema ishimotoi. A new, fluorescently labeled oligonucleotide probe targeted against 16S rRNA was designed so that that it hybridized specifically with whole cells of Desulfonema species. Filamentous bacteria that hybridized with the same probe were detected in sediment samples and in association with the filamentous sulfur-oxidizing bacterium Thioploca in its natural habitat. We conclude that Desulfonema species constitute an ecologically significant fraction of the sulfate-reducing bacteria in organic-rich sediments and microbial mats. PMID- 10525736 TI - Phototrophic utilization of toluene under anoxic conditions by a new strain of blastochloris sulfoviridis AB - The capacity of anoxygenic phototrophic bacteria to utilize aromatic hydrocarbons was investigated in enrichment cultures with toluene. When mineral medium with toluene (provided in an inert carrier phase) was inoculated with activated sludge and incubated under infrared illumination (> 750 nm), a red-to-brownish culture developed. Agar dilution series indicated the dominance of two types of phototrophic bacteria. One type formed red colonies, had rod-shaped cells with budding division, and grew on benzoate but not on toluene. The other type formed yellow-to-brown colonies, had oval cells, and utilized toluene and benzoate. One strain of the latter type, ToP1, was studied in detail. Sequence analysis of the 16S rRNA gene and DNA-DNA hybridization indicated an affiliation of strain ToP1 with the species Blastochloris sulfoviridis, a member of the alpha-subclass of Proteobacteria. However, the type strain (DSM 729) of Blc. sulfoviridis grew neither on toluene nor on benzoate. Light-dependent consumption of toluene in the presence of carbon dioxide and formation of cell mass by strain ToP1 were demonstrated in quantitative growth experiments. Strain ToP1 is the first phototrophic bacterium shown to utilize an aromatic hydrocarbon. In the supernatant of toluene-grown cultures and in cell-free extracts incubated with toluene and fumarate, the formation of benzylsuccinate was detected. These findings indicate that the phototrophic bacterium activates toluene anaerobically by the same mechanism that has been reported for denitrifying and sulfate reducing bacteria. The natural abundance of phototrophic bacteria with the capacity for toluene utilization was examined in freshwater habitats. Counting series revealed that up to around 1% (1.8 x 10(5) cells per gram dry mass of sample) of the photoheterotrophic population cultivable with acetate grew on toluene. PMID- 10525737 TI - Novel 16S rRNA gene sequences retrieved from highly saline brine sediments of kebrit deep, red Sea AB - In this study, we report on first 16S rRNA gene sequences from highly saline brine sediments taken at a depth of 1,515 m in the Kebrit Deep, northern Red Sea. Microbial DNA extracted directly from the sediments was subjected to PCR amplification with primers specific for bacterial and archaeal 16S rRNA gene sequences. The PCR products were cloned, and a total of 11 (6 bacterial and 5 archaeal) clone types were determined by restriction endonuclease digestion. Phylogenetic analysis revealed that most of the cloned sequences were unique, showing no close association with sequences of cultivated organisms or sequences derived from environmental samples. The bacterial clone sequences form a novel phylogenetic lineage (KB1 group) that branches between the Aquificales and the Thermotogales. The archaeal clone sequences group within the Euryarchaeota. Some of the sequences cluster with the group II and group III uncultivated archaea sequence clones, while two clone groups form separate branches. Our results suggest that hitherto unknown archaea and bacteria may thrive in highly saline brines of the Red Sea under extreme environmental conditions. PMID- 10525738 TI - The dcuD (former yhcL) gene product of escherichia coli as a member of the DcuC family of C4-dicarboxylate carriers: lack of evident expression AB - The dcuD gene (formerly yhcL) of Escherichia coli shows significant sequence similarity only to the dcuC gene of E. coli, which encodes a C4-dicarboxylate carrier (DcuC) that functions during anaerobic growth. Inactivation of dcuD had no effect on the growth of E. coli under a large number of conditions and led to no detectable changes in phenotype. Translational dcuD'-'lacZ gene fusions were not significantly expressed in the presence of dicarboxylates or monocarboxylates under oxic or anoxic conditions. Other potential substrates such as amino sugar derivatives, amino acids, and alpha-aspartyl dipeptides also did not lead to expression of dcuD. Changes in medium composition, pH, ionic strength, and temperature had no significant effects on dcuD expression. A dcuD gene amplified from a natural isolate of E. coli was not expressed in wild-type and E. coli K-12 backgrounds. Cloning of dcuD behind an inducible promoter resulted in the synthesis of a protein of the expected size (49 kDa), which, however, did not complement for the loss of DcuC or other C4-dicarboxylate carriers. It is suggested that dcuD encodes a protein of the DcuC family of anaerobic C4 dicarboxylate carriers and that dcuD is not significantly expressed or is expressed only under conditions not related to carboxylate metabolism. When two adjacent open reading frames (y0585 and y0586) from Haemophilus influenzae are fused, the resulting hypothetical protein has sequence similarity to DcuC and DcuD. PMID- 10525739 TI - The role of the twin-arginine motif in the signal peptide encoded by the hydA gene of the hydrogenase from wolinella succinogenes AB - The hydABC operon of Wolinella succinogenes encodes the three subunits of the membrane-integrated Ni-hydrogenase. The catalytic subunit, HydB, is on the periplasmic side of the membrane. Residues R41 and R42 of the twin-arginine motif within the signal peptide of the precursor of the iron-sulfur subunit, HydA, were replaced by two glutamine residues. The corresponding mutant did not grow with H(2) as the electron donor of anaerobic respiration. Mature HydB and the precursor protein of HydA were located exclusively in the cytoplasmic cell fraction of the mutant, which catalyzed the reduction of benzyl viologen by H(2), suggesting that HydB contained Ni. The HydC protein was located in the membrane fraction of the mutant in wild-type amounts. HydC was purified and was shown to contain heme. The results suggest that HydA and HydB are translocated across the membrane by the Tat (twin-arginine translocation) system. The translocation of HydA and HydB as well as the maturation of the precursor protein of HydA appear to depend on the presence of the twin-arginine motif. In contrast, maturation of HydB, the insertion of HydC into the membrane, and heme attachment to HydC are apparently independent of the twin-arginine motif and do not require translocation of the two other hydrogenase subunits. PMID- 10525740 TI - The correlation of the gene csoS2 of the carboxysome operon with two polypeptides of the carboxysome in thiobacillus neapolitanus AB - The carboxysomal polypeptides of Thiobacillus neapolitanus with apparent molecular masses of 85 and 130 kDa were isolated and subjected to N-terminal sequencing. The first 17 amino acids of the two peptides were identical. The sequence perfectly matched the deduced amino acid sequence of an open reading frame in the carboxysome operon. The gene was subsequently named csoS2. Expression of the gene in Escherichia coli resulted in the production of two peptides with apparent molecular masses of 85 and 130 kDa. Immunospecific antibodies generated against the smaller peptide recognized both peptides; the peptides were named CsoS2A and CsoS2B, respectively. A digoxigenin-hydrazide glycosylation assay revealed that both CsoS2A and CsoS2B are post-translationally modified by glycosylation. CsoS2 was localized to the edges of purified carboxysomes by immunogold electron microscopy using the monospecific CsoS2A antibodies. The molecular mass of CsoS2A calculated from the nucleotide sequence was 92.3 kDa. PMID- 10525741 TI - Ethene as an auxiliary substrate for the cooxidation of cis-1, 2-dichloroethene and vinyl chloride AB - Cultures able to dechlorinate cis-1,2-dichloroethene (cDCE) were selected with ethene (3-20%, v/v) as the sole source of carbon and energy. One mixed culture (K20) could degrade cDCE (400 &mgr;mol l(-1)) or vinyl chloride (100 &mgr;mol l( 1)) in the presence of ethene (/=4 weeks of sustained cocaine abstinence, whereas only one patient in the low and none in the zero magnitude condition achieved more than 2 weeks. Reinforcement magnitude was a critical determinant of the effectiveness of this abstinence reinforcement intervention. PMID- 10525749 TI - Strain-dependent effects of MK-801 on passive avoidance behaviour in mice: interactions with morphine and immobilization stress. AB - RATIONALE: Post-training treatments (drugs, stress, ECS) influence retention performance of laboratory animals, sometimes in a strain-dependent way. In a previous study, an interaction between the effects of morphine and of the non competitive N-methyl-d-aspartate (NMDA) receptor antagonist MK-801 on retention performance was observed, in random bred mice. OBJECTIVE: In the present research, we have investigated the effects on retention of C57BL/6 (C57) and DBA/2 (DBA) mice exerted by a) morphine, b) MK-801 and c) naltrexone. Further, we have studied in both strains the effects exerted on retention by combinations of morphine and MK-801, and of MK-801 and immobilization stress. Finally, the naltrexone-reversibility of the interaction between immobilization stress and MK 801 was also assessed. METHODS: All treatments were administered immediately after training in mice tested in a passive avoidance task. Drugs were injected IP. RESULTS: The results of our experiments showed that morphine and the non competitive NMDA receptor antagonist MK-801 exerted dose- and time-dependent facilitatory effects on retention performance in C57 mice, and dose- and time dependent impairments in DBA mice. Further, dose- and time-dependent deleterious effects on retention performance, in the C57 strain, and dose- and time-dependent enhancing effects, in the DBA strain, were observed following post-training IP naltrexone administration. MK-801 enhanced, in both strains, the effects of morphine. Finally, immobilization stress enhanced in both strains the effects of MK-801 and these effects were naltrexone-reversible. CONCLUSIONS: In conclusion, the results of this study show that the genetic make-up of the mice played an important role in all the effects observed, and, in particular, in the interaction between the opioid and the glutamatergic systems. Further, the naltrexone reversibility of the interaction between MK-801 and immobilization stress suggests that opioid mechanisms were involved. PMID- 10525750 TI - Interactions between social stress and morphine in the periaqueductal gray: effects on affective vocal and reflexive pain responses in rats. AB - RATIONALE: Endogenous opioid systems within the mesencephalic periaqueductal gray matter (PAG) appear to be intricately involved in many affective, defensive, submissive, and reflexive responses, and these systems are activated by aversive stimuli. OBJECTIVES: The present experiments evaluated the influence of opioid receptors within the PAG on affective vocal and reflexive responses to aversive stimuli in socially inexperienced, as well as defensive and submissive responses in defeated, adult male Long-Evans rats. METHODS: Defeat stress consisted of: (1) an aggressive confrontation with a "resident" stimulus rat in which the experimental "intruder" rat exhibited escape, defensive and submissive behaviors [i.e. upright, supine postures and ultrasonic vocalizations (USV)], and subsequently, (2) protection from the resident rat with a wire mesh screen for ca. 25 min. Defeat stress was immediately followed by an experimental session with thermal antinociceptive and tactile startle stimuli (20 psi airpuffs). RESULTS: The mu opioid receptor agonist morphine (0.3, 1, 3 microgram IC) attenuated startle-induced USV and the tail-flick reflex in socially inexperienced and defeated rats, with both groups of rats demonstrating equal sensitivity to morphine. Morphine decreased defeat-induced USV and increased the display of the crouch posture in defeated rats; these morphine effects in socially inexperienced and defeated rats were re- versed with the opioid receptor antagonist naltrexone (0.1 mg/kg IP). CONCLUSIONS: These results reveal that the ventrolateral PAG is an important site in which mu opioid receptor agonists such as morphine mediate affective vocal and submissive responses, yet this structure is not critical in the display of defeat stress-augmented effects of morphine. Endogenous opioid mechanisms appear to participate in the organization of defensive behavior, namely, to facilitate a shift from active to passive forms of coping. PMID- 10525752 TI - The effects of subchronic haloperidol on intact and dizocilpine-disrupted sensorimotor gating. AB - RATIONALE: Reversal of deficits in prepulse inhibition (PPI) of the startle reflex in rats is considered a preclinical screen for potential antipsychotics. Whereas acutely administered antipsychotics consistently reverse apomorphine induced deficits in PPI, some antipsychotics, including haloperidol, are unable to reverse deficits in PPI produced by non-competitive NMDA antagonists such as phencyclidine or dizocilpine (MK-801). Acute administration of antipsychotics tends to facilitate baseline PPI. However, the effect is generally not large enough in magnitude nor reliable enough to be considered a useful preclinical screen for antipsychotic activity. OBJECTIVE: Because the clinical effects of antipsychotics typically require subchronic administration, this study tested the hypothesis that reversal of NMDA antagonist-induced deficits in PPI by antipsychotics require subchronic administration. A second aim of this study was to determine if subchronic administration of an antipsychotic produces a more potent facilitation of baseline PPI than acute administration. METHODS: Rats received a subcutaneous injection of 0, 0.025, 0.1 or 0.5 mg/kg haloperidol for 16 consecutive days. On day 16, half the rats in each haloperidol dose group received a second subcutaneous injection consisting of either dizocilpine (0.1 mg/kg) or saline. RESULTS: None of the haloperidol doses tested had a significant effect on baseline PPI. The 0.1 mg/kg dose of haloperidol diminished but did not completely reverse dizocilpine-induced disruption of PPI. The other doses had no significant effect. CONCLUSIONS: These results suggest that time course factors may partially modify the effects of haloperidol on dizocilpine-induced disruption of PPI but not its effect on baseline PPI. PMID- 10525751 TI - Systemic sulpiride in young adult volunteers simulates the profile of cognitive deficits in Parkinson's disease. AB - RATIONALE: The mesotelencephalic dopamine system has been implicated in cognitive processes dependent on an intact prefrontal cortex. Most previous research in humans has focused on dopaminergic agonists and their effects on tasks of working memory. OBJECTIVES: The present study was designed to investigate the cognitive and subjective effects of two doses (200 mg and 400 mg) of the dopaminergic D(2) receptor antagonist, sulpiride on a broad range of well-validated neuropsychological tasks in a group of 34 young healthy male volunteers. METHODS: Cognitive tasks were administered to subjects after ingestion of either drug or placebo within a double-blind, placebo-controlled, cross-over design. The cognitive tests included tasks from the Cambridge Neuropsychological Test Automated Battery (CANTAB) and were designed to assess visuospatial recognition memory, planning ability, working memory, strategy learning, sustained attention and attentional set-shifting. In addition, the National Adult Reading Test (NART) was used to assess verbal IQ, and visual analogue scales to assess subjective effects of the drug. RESULTS: Subjects on sulpiride were impaired on the tasks of spatial recognition, spatial working memory (sequence generation), planning (one touch Tower of London) and attentional set-shifting. Only the spatial working memory task demonstrated a dose dependent effect. The impairments were not due to generalised sedative or motoric influences of sulpiride. CONCLUSIONS: All of the tasks impaired following sulpiride are known to be sensitive to frontal lobe damage and the precise pattern of deficits seen is consistent with the anatomical distribution of central dopamine receptors. The results are discussed with particular reference to their close simulation of the impairments seen in idiopathic Parkinson's disease. PMID- 10525753 TI - Assessment of GABA(A)benzodiazepine receptor (GBzR) sensitivity in patients on benzodiazepines. AB - OBJECTIVES: To measure GABA(A) benzodiazepine receptor sensitivity in patients taking benzodiazepines and compare with matched controls. METHODS: Seven patients who were on prescribed benzodiazepines for an anxiety disorder or insomnia were recruited from general practice and an adult mental health service outpatient clinic. They were matched with seven volunteers. All subjects received an intravenous injection of midazolam 50 microgram/kg in 10 ml normal saline over 10 min. Objective responses to midazolam were assessed using saccadic eye movement velocity slowing and subjective assessments using visual analogue scales. Measurements were recorded for 120 min and plasma midazolam concentrations obtained at 15-min intervals post-infusion to 120 min. Ratios of pharmacodynamic/pharmacokinetic effects were obtained for each individual to estimate GABA(A) benzodiazepine receptor sensitivity. RESULTS: Patients had an attenuated response to midazolam on both subjective and objective measures. GABA(A) benzodiazepine receptor sensitivity was significantly reduced in the patient group. CONCLUSIONS: Chronic treatment with benzodiazepines was associated with reduced effects of midazolam. Saccadic eye movement velocity was especially sensitive as a measure of attenuated response. PMID- 10525754 TI - Depletion of brain serotonin following intra-raphe injections of 5,7 dihydroxytryptamine does not alter d-amphetamine self-administration across different schedule and access conditions. AB - OBJECTIVES: These experiments investigated the effects of selective serotonin (5 HT) depletion on intravenous self-administration of d-amphetamine. METHODS: Depletion of brain 5-HT levels was induced by injecting the serotonergic neurotoxin 5,7-dihydroxytryptamine (5, 7-DHT) into the dorsal and median raphe nuclei. Rats were then trained to self-administer d-amphetamine according to various schedule and access conditions via chronically indwelling intravenous catheters. RESULTS: Large reductions of brain 5-HT did not alter responding for a training dose of 120 microgram/kg d-amphetamine delivered according to a fixed ratio 1 schedule during 3-h sessions. When the dose of d-amphetamine was altered (0, 3.75, 7. 5, 15, 30, 60 microgram/kg per infusion) a characteristic inverted U shaped dose response function was obtained. The 5-HT depleted rats showed increased responding for the lower doses of d-amphetamine, with a large significant increase in responding for the 7.5 microgram/kg dose. In these same rats, the suppressive effect of 10 mg/kg fluoxetine on d-amphetamine (60 microgram/kg) self-administration was prevented. The 5,7-DHT lesion also did not alter responding for d-amphetamine (120 microgram/kg) in longer (8 h) daily access sessions. Responding for d-amphetamine delivered on a progressive ratio schedule, in which response requirements increased for each successive infusion of d-amphetamine, was also determined in 5-HT depleted rats. The number of d amphetamine infusions was not different from the number of infusions earned by sham-lesioned rats across a range of doses of d-amphetamine (7.5-60 microgram/kg). In a final experiment, spontaneous acquisition of self administration of low doses of d-amphetamine (10 and 30 microgram/kg) was measured in 5-HT depleted and control rats. Again, self-administration behaviour in the 5-HT depleted rats did not differ from controls. CONCLUSIONS: These results provide no evidence that reducing 5-HT function alters the primary reinforcing effects of self-administered amphetamine. The increase in self administration of a low dose of amphetamine observed in experiment 1 probably involves some other process such as increased resistance to extinction. PMID- 10525755 TI - Acute effects of venlafaxine and paroxetine on serotonergic transmission in human volunteers. AB - RATIONALE: Antidepressant drugs are thought to enhance serotonergic neurotransmission through postsynaptic 5-HT(1A) receptors. This effect is delayed in animals and may be paralleled by a delay in the onset of a clinical response in humans. In humans, the growth hormone (GH) response to intravenous l tryptophan (IV l-TRP) is blocked by the 5-HT(1A) antagonist pindolol and the prolactin response is blunted. Both are therefore thought to be a useful measure of 5-HT(1A) receptor function. Clomipramine has previously been found to enhance the GH and prolactin responses to IV l-TRP after only 2 h. OBJECTIVE: The purpose of this study was to use this method to investigate the effects of newer antidepressants on 5-HT(1A) receptor-mediated function. METHODS: Twelve healthy male volunteers took part in a random order, double blind study, in which 18.75 mg venlafaxine, 5 mg paroxetine or placebo was administered 3 h before infusion of l-TRP. RESULTS: Pretreatment with venlafaxine significantly enhanced the growth hormone (GH) response to the infusion compared with pretreatment with placebo. There was no significant difference between the GH response following paroxetine compared with placebo or with venlafaxine. CONCLUSIONS: The data suggest enhancement of transmission through postsynaptic 5-HT(1A) receptors by venlafaxine but not paroxetine, after only 3 h. PMID- 10525756 TI - Selective effects of nicotine on attentional processes. AB - RATIONALE: It is now well established from electrophysiological and behavioural evidence that nicotine has effects on information processing. The results are usually explained either by a primary effect of nicotine or by a reversal effect of a nicotine-induced, abstinence deficit. In addition, there is dispute about the cognitive processes underlying the changes in performance. METHODS: This study has approached the first question by using the nicotine patch, in order to administer nicotine chronically. In addition, we examined the effects of nicotine on attention with a selection of tests which assessed the intensity and selectivity features of attention, using the Random Letter Generation test, the Flexibility of Attention test and the Stroop test. RESULTS: Nicotine enhanced the speed of number generation and the speed of processing in both the control and interference conditions of the Stroop test. There were no effects on attentional switching of the Flexibility of Attention test. CONCLUSION: The results are consistent with the hypothesis that nicotine mainly improves the intensity feature of attention, rather than the selectivity feature. PMID- 10525757 TI - New evidence that the pharmacological effects of benzodiazepine receptor ligands can be associated with activities at different BZ (omega) receptor subtypes. AB - RATIONALE: It has been suggested that different BZ (omega) receptor subtypes may mediate distinct behavioural effects of BZ receptor ligands. OBJECTIVE: The present study examined this hypothesis further. METHODS: The antagonism exerted by the selective BZ(1) (omega(1)) receptor antagonist beta-CCT on the pharmacological effects of the selective BZ(1) (omega(1)) receptor agonist zolpidem and the non-selective BZ (omega) receptor agonist diazepam in behavioural, biochemical and electrophysiological experiments was assessed. RESULTS: beta-CCT which was devoid of activity per se, antagonized the effects of the non-selective BZ (omega) receptor full agonist diazepam and the selective BZ(1) (omega(1)) receptor full agonist zolpidem against seizures produced by isoniazid, but beta-CCT failed to affect their action on seizures produced by pentylenetetrazole (PTZ), suggesting that BZ(2) (omega(2)) receptors may be primarily involved in the convulsant action of PTZ. In the light/dark test, beta CCT abolished the anxiolytic-like action of diazepam. In tests designed to investigate the central depressant activity of drugs, beta-CCT antagonized the sedative effects of diazepam and zolpidem, but failed to modify clearly the myorelaxant effects of diazepam. These differences may be related to the selectivity of beta-CCT for BZ(1) (omega(1)) sites as indicated by the preferential displacement of [(3)H]flumazenil in BZ(1) (omega(1))-enriched structures as compared to BZ(2) (omega(2))-enriched structures in the mouse. In in vitro experiments, beta-CCT antagonized the potentiation of the GABA-induced Cl(-) current produced by zolpidem in HEK cells expressing the alpha(1)beta(2)gamma(2) receptor or in cerebellar Purkinje neurones, while it failed to modify the diazepam potentiation at either alpha(3)beta(2)gamma(2) or alpha(5)beta(3)gamma(2) receptor subtypes. CONCLUSION: These results are consistent with the hypothesis that BZ(1) (omega(1)) receptors play an important role in the anxiolytic and sedative/hypnotic effects of BZ (omega) receptor ligands, whereas activity at BZ(2) (omega(2)) sites might be associated primarily with muscle relaxation. PMID- 10525758 TI - Adenosine and memory storage: effect of A(1) and A(2) receptor antagonists. AB - RATIONALE: Caffeine is a non-selective A(1)/A(2 )adenosine receptor antagonist which is known to improve cognitive performance in humans. This effect of caffeine has been attributed to its antagonism of adenosine receptors. OBJECTIVE: The present study was devised to identify the role of A(1 )and A(2A) adenosine receptors in the facilitation of memory consolidation in mice performing a passive avoidance task. METHODS: Adult albino Swiss male mice were used. The mice were trained in a step-through inhibitory avoidance task in which they were punished by a foot-shock (0.4 mA, 5 Hz, for 3 s) delivered through the grid floor. Caffeine (0.1, 0.3, 1.0 and 3.0 mg/kg), SCH 58261 (0.1, 0.3, 1.0 and 3.0 mg/kg) and DPCPX (0.1, 0.3, 1.0 and 3.0 mg/kg) were injected IP immediately or 180 min after training. The retention test was performed 24 h after training. RESULTS: Caffeine and the selective A(2A) adenosine receptor antagonist SCH 58261 facilitated retention when administered immediately after training, but not when administered 180 min later. The dose response was a bell-shaped curve. Conversely, post-training administration of the selective A(1) adenosine receptor antagonist DPCPX did not affect retention. Caffeine and SCH 58261 had no effect in mice not given the foot-shock on the training trial, a finding indicating that the drug's effect on retention was specific. CONCLUSIONS: These results suggest that A(2A) but not A(1) adenosine receptors are involved in memory retention and consolidation. PMID- 10525759 TI - A PET study of D(1)-like dopamine receptor ligand binding during altered endogenous dopamine levels in the primate brain. AB - RATIONALE: Several positron emission tomography (PET) studies have shown that radioligand binding to D(2)-like dopamine receptors competes with endogenous dopamine. OBJECTIVE: The purpose of this PET study was to examine the effect of amphetamine and reserpine on D(1)-like dopamine receptor binding. METHODS: Three Cynomolgus monkeys were examined with the radioligands [(11)C]SCH 23390 or [(11)C]NNC 112 at baseline condition and after pretreatment with amphetamine (2 mg/kg IV). The B/F values (binding potential) in the striatum and the neocortex were calculated at transient equilibrium. In two monkeys, the effect of the long lasting dopamine depletion after reserpine (1 mg/kg IV) was followed by a repeated Scatchard procedure in up to 77 days after drug administration. The Scatchard analysis was based on two PET measurements with high and low specific radioactivity and allowed the calculation of D(1)-like dopamine receptor density (B(max)) and apparent affinity (K(D)(app)). RESULTS: The effect of amphetamine on the B/F values was between -14 and 6%. These changes can be considered as within the range of the test-retest reliability. Thus, there was no evident effect of amphetamine-induced dopamine release on D(1)-like dopamine receptor binding. Five hours after reserpine administration, there was no change in B(max) or K(D)(app). At 3, 23, and 28 days after reserpine administration, the Scatchard analyses indicated a 13-20% reduction in B(max) without any evident change in K(D)(app) in both the striatum and the neocortex. CONCLUSIONS: The lack of evident effects of amphetamine and reserpine on D(1)-like dopamine receptor binding is markedly different from the 20% amphetamine-induced decrease and 50% reserpine-induced increase that has been consistently reported for D(2)-like dopamine receptor binding. The data indicated that D(1)-like dopamine receptor occupancy of endogenous dopamine is low at physiological condition. It is thus unlikely that D(1)-like dopamine receptor radioligands can be used to measure changes in the concentration of endogenous dopamine. PMID- 10525761 TI - Spinal tumours in neurofibromatosis type 1: an MRI study of frequency, multiplicity and variety. AB - In neurofibromatosis type 1 (NF1) spinal tumours cause neurological symptoms in about 2 % of patients. Among over 1400 patients with NF1 we saw symptomatic spinal tumours in 23 (1.6 %). MRI of the entire spinal canal was obtained in 54 patients aged 5-56 years with NF1. The number, site, morphology and signal characteristics of the spinal tumours were recorded and analysed. There were 24 patients with symptoms such as sensory impairment or paralysis; 30 patients had no neurological deficits. Of the 24 symptomatic patients, 23 (96 %) had spinal tumours, while we saw spinal tumours in 12 (40 %) of the 30 patients without neurological deficits. No spinal segment was preferred in symptomatic or asymptomatic patients. Most intraspinal extramedullary tumours were primarily extradural and intraforaminal. MRI showed intramedullary tumours in 3 patients (6 %), intraspinal extramedullary tumours in 18 (33 %) and intraforaminal tumours in 31 (57 %). Only neurological deficits in patients with NF1 should prompt further diagnostic clarification. In patients with neurological symptoms there may be a multiplicity of masses in the spinal canal, which can lead to difficulties in attaching symptoms to a certain tumour. In patients who do not satisfy the NIH criteria, it can be a helpful observation that spinal tumours in NF1 are primarily intraforaminal, extending into the spinal canal, while in NF2 they are mostly intraspinal intradural tumours. PMID- 10525760 TI - Sex differences in sensorimotor gating of the human startle reflex: all smoke? AB - RATIONALE: A recent report described sex differences in the effects of nicotine use and withdrawal on prepulse inhibition of acoustic startle (PPI), but no sex differences in PPI in non-smokers. OBJECTIVE: To determine whether previously reported male>female acoustic PPI reflect sex differences in smoking effects on PPI, rather than simple sex differences in the regulation of PPI. A retrospective analyses of >600 carefully screened normals tested over the past 12 years was completed. RESULTS: Male>female acoustic PPI was detected in analyses that included: 1) all subjects; or 2) self-declared non-smokers. CONCLUSIONS: Sex differences in PPI cannot be accounted for by smoking history, because they are present across a large sample of non-smoking normal controls. PMID- 10525762 TI - Posterior extradural migration of extruded thoracic and lumbar disc fragments: role of MRI. AB - We report three patients with a sequestrated disc fragment posterior to the thecal sac. The affected disc was lumbar in two cases and thoracic in the third. Disc fragment migration is usually limited to the anterior extra dural space. Migration of a disc fragment behind the dural sac is seldom encountered. MRI appears to be the method of choice to make this diagnosis. The disc fragments gave low signal on T1- and slightly high signal on T2-weighted images and showed rim contrast enhancement. The differential diagnosis includes abscess, metastatic tumour and haematoma. PMID- 10525763 TI - Accuracy of MRI-guided stereotactic thalamic functional neurosurgery. AB - Our goal was to evaluate the accuracy of stereotactic technique using MRI in thalamic functional neurosurgery. A phantom study was designed to estimate errors due to MRI distortion. Stereotactic mechanical accuracy was assessed with the Suetens-Gybels-Vandermeulen (SGV) angiographic localiser. Three-dimensional MRI reconstructions of 86 therapeutic lesions were performed. Their co-ordinates were corrected from adjustments based on peroperative electrophysiological data and compared to those planned. MR image distortion (maximum: 1 mm) and chemical shift of petroleum oil-filled localiser rods (2.2 mm) induced an anterior target displacement of 2.6 mm (at a field strength of 1.5 T, frequency encoding bandwidth of 187.7 kHz, on T1-weighted images). The average absolute error of the stereotactic material was 0.7 mm for anteroposterior (AP), 0.5 mm for mediolateral (ML) and 0.8 mm for dorsoventral (DV) co-ordinates (maximal absolute errors: 1.6 mm, 2.2 mm and 1.7 mm, respectively; mean euclidean error: 1 mm). Three-dimensional MRI reconstructions showed an average absolute error of 0.8 mm, 0.9 mm and 1.9 mm in AP, ML and DV co-ordinates, respectively (maximal absolute errors: 2.4 mm, 2.7 mm and 5.7 mm, respectively; mean euclidean error: 2.3 mm). MRI distortion and chemical-shift errors must be determined by a phantom study and then compensated for. The most likely explanation for an average absolute error of 1.9 mm in the DV plane is displacement of the brain under the pressure of the penetrating electrode. When this displacement is corrected for by microelectrode recordings and stimulation data, MRI offers a high degree of accuracy and reliability for thalamic stereotaxy. PMID- 10525764 TI - Thallium-201 SPECT of adjacent intracranial tumours: a contrast in thallium kinetics. AB - We report a case of adjacent intracranial tumours: malignant fibrous histiocytoma (MFH) and meningioma. Thallium-201 single-photon emission computed tomography demonstrated different thallium kinetics between the tumours (slow washout from the MFH and rapid clearance in the meningioma) and could be said to have been useful for preoperative histological estimation. PMID- 10525765 TI - Oligodendroglial gliomatosis cerebri: (1)H-MRS suggests elevated glycine/inositol levels. AB - Oligodendroglial gliomatosis cerebri is very rare. We describe 42-year-old woman who had low-grade oligodendroglial gliomatosis cerebri confirmed on stereotactic biopsy. The diffuse nature of the tumour was apparent clinically, neurophysiologically, on MRI and on proton magnetic resonance spectroscopy (MRS). She also had an isolated, false-localising partial seventh nerve palsy. MRS, of which there are no previous reports, suggested elevated glycine/inositol levels. This might be explained by the cell lineage from which the tumour arose. PMID- 10525766 TI - Choroid plexus papilloma of foramen of Luschka with multiple recurrences and cystic features. AB - We present a rare cerebellopontine angle choroid plexus papilloma arising at the foramen of Luschka, without an associated intraventricular component. Distinct features of the tumour on MRI, of multiple recurrences with cystic features, are described, with a review of the literature. PMID- 10525767 TI - The prevalence and distribution of white-matter changes on different MRI pulse sequences in a post-stroke cohort. AB - No uniform criteria currently exist for rating white-matter (WM) high-signal foci on MRI. Ratings are based on descriptive terms, different pulse sequences and different WM areas. Reports on the prevalence and clinical correlates of high signal foci have been contradictory. We wanted to examine the contribution of the pulse sequence and WM area on rating WM changes. We analysed WM changes separately on T2-, protondensity (PD)- and T1-weighted images in periventricular, subcortical, watershed area and deep WM. The difference between T2- and PD weighted images was significant for frontal caps, counting small foci or analysing subcortical changes. T1-weighted images showed significantly less change, but the number of foci detected was greater than previously thought. The prevalence of WM high-signal foci was greatest in the watershed zone and smallest in the subcortical area. There was a significant correlation between foci in different areas. PMID- 10525768 TI - One-and-a-half syndrome in pontine infarcts: MRI correlates. AB - The one-and-a-half syndrome is characterised by a lateral gaze palsy in one direction and internuclear ophthalmoplegia in the other. It is due to a unilateral lesion of the dorsal pontine tegmentum, involving the ipsilateral paramedian pontine reticular formation, internuclear fibres of the ipsilateral medical longitudinal fasciculus and, usually, the abducens nucleus. The main causes of this rare syndrome are stroke and multiple sclerosis. Few cases have been reported since the introduction of MRI. Our aim was to examine clinicoradiological correlations in six patients with a one-and-a-half syndrome due to a stroke. Ophthalmological symptoms were diplopia, oscillopsia or blurred vision. Four patients had an associated facial nerve palsy, three a hemiparesis and one a unilateral hemihypoaesthesia. MRI revealed an infarct in the pons in all patients. The cause of the infarct was a basilar artery dissection in one patient, bilateral vertebral artery dissection in a second and unknown in the other four. All patients recovered within 2 days to 8 weeks. This study showed a good correlation between the site of the lesion (superior, inferior or extensive pontine ischaemia) and clinical deficits. PMID- 10525769 TI - Bilateral basal ganglion haemorrhage in diabetic ketoacidotic coma: case report. AB - We report bilateral oedema and haemorrhagic transformation in the basal ganglia of a 59-year old woman with severe diabetic ketoacidosis. Lack of cerebral vascular autoregulation, followed by blood-brain barrier disruption due to the so called breakthrough mechanism is presumed to be the cause. PMID- 10525770 TI - "De novo" formation of intracranial aneurysms: who is at risk? AB - Although aneurysms are widely considered to be of congenital origin there is still debate as to whether some at least might be formed de novo during life. A review of all 49 reported cases plus one previously unpublished case reveals common clinical features and might aid in the management of this group of patients. Statistical analysis of all 50 cases of de novo aneurysms discloses a more frequent history of smoking (P = 0.0007) and arterial hypertension (P = 0.0026) than in a control cohort. Patients with de novo aneurysms are younger (P < 0.0001); the proportion with multiple aneurysms was 28 %. Of de novo aneurysms 44 % became symptomatic 3-6 years after the first subarachnoid haemorrhage (SAH), and the interval was significantly shorter in hypertensive patients. We suggest that young patients with a history of SAH and arterial hypertension and nicotine abuse should therefore be considered for conventional angiography after a 5-year interval. MRA might not be useful due to clip artefacts from even nonferromagnetic clips. Close control of blood pressure is essential in these patients. PMID- 10525771 TI - Artefact on MRA following aneurysm clipping: an in vitro study and prospective comparison with conventional angiography. AB - Using both an experimental model and clinical cases, we looked at the artefact produced by Aesculap titanium-alloy aneurysm clips on MRA. Experimentally, the volume affected by artefact was 50 % less when the clip was imaged lying parallel to the main ferromagnetic field than when lying perpendicular to it. Clinically, MRA was prospectively compared with digital subtraction angiography (DSA) in nine patients who had undergone aneurysm clipping. One patient with a non-diagnostic MRA due to movement artefact was excluded. In all other cases there was an area of signal loss surrounding the clips, obscuring the immediately adjacent vessel segments. There was good demonstration of the adjacent bifurcations in five cases and the contralateral circulation was seen well in all patients. In three cases in which the adjacent bifurcations were not seen, considerable vasospasm was suggested by MRA and confirmed with DSA. In one patient an unclipped contralateral ophthalmic artery aneurysm was identified using both modalities. In this series there were no adverse events relating to clips in either static or time-varying magnetic fields. PMID- 10525772 TI - MRI of the brain in HIV-positive patients: what is the value of routine intravenous contrast medium? AB - Our purpose was to assess the value of routine administration of intravenous gadolinium-DTPA (Gd-DTPA) for cranial MR in a series of human immunodeficiency virus (HIV)-positive patients. Two radiologists retrospectively reviewed 150 consecutive examinations of 104 patients. All patients underwent unenhanced and contrast-enhanced images. Each radiologist independently assessed first the unenhanced images alone and then the pre- and postinjection images together. Then both reviewed the complete study and produced a consensus report. The history, investigations and management were collated separately and were unknown to the radiologists. Contrast-enhanced T1-weighted images showed new focal abnormalities, not seen on the T2-weighted or unenhanced images in 15 (14 %) patients, but almost always in the context of abnormal unenhanced images. In only 2 patients (2 %) did contrast medium reveal abnormalities when the unenhanced study had been considered normal. In only 1 of these (1 %) was the new finding, cytomegalovirus diffuse ependymal enhancement, of clinical importance, although the diagnosis of encephalitis was made on routine examination of cerebrospinal fluid. The other revealed a toxoplasma lesion in a patient known to have resolving disease. Meningeal disease not suspected on the unenhanced images was shown in 2 patients (2 %). In these case the unenhanced images were abnormal in other respects. Intravenous Gd-DTPA was helpful to the radiologist in making a radiological diagnosis in 11 patients (11 %), usually by improving characterisation of a lesion seen on the unenhanced images. The contribution of intravenous Gd-DTPA in this series does not warrant recommending its use in every case. PMID- 10525773 TI - Meningeal leiomyoma in an adult with AIDS: CT and MRI with pathological correlation. AB - We describe the imaging features of a meningeal leiomyoma with pathological correlation in an adult with AIDS. On CT the tumour showed a central low attenuation area and an enhancing peripheral ring. It gave low signal on T1 weighting and on T2-weighted images a central high-signal area was surrounded by a markedly low-signal band, which showed contrast enhancement. As far as we know, this is the first report of this condition in the radiological literature. PMID- 10525774 TI - Severe otitis and mastoiditis due to Rhodococcus equi in a patient with AIDS. Case report. AB - We report a case of otitis media associated with pneumonia due to Rhodococcus equi. A 31-year-old patient with AIDS presented with cough and right facial palsy. Imaging revealed right otitis media and severe temporal bone destruction, associated with pneumonia. R. equi was isolated from ear secretions, blood, and sputum. The radiologic findings are described. This unusual pathogen should be included in the differential diagnosis of the immunocompromised patient with aggressive otitis. PMID- 10525775 TI - Announcements PMID- 10525776 TI - 2nd european course on pediatric neuroradiology PMID- 10525777 TI - Studies of iodixanol in the rabbit lung and peritoneum. AB - OBJECTIVE: To evaluate a new water-soluble contrast agent, iodixanol. The study evaluates absorption from the peritoneal cavity and toxicity in the lung. MATERIALS AND METHODS: Thirty New Zealand white rabbits were given the study agent and comparative agents into an endotracheal tube. Serial chest radiographs were evaluated for development of pulmonary edema. All lungs were evaluated histopathologically for toxic inflammatory response. Fifteen different rabbits were given intraperitoneal injection of the study and comparative contrast agents. Serial abdomen radiographs, taken up to 24 h after injection, were evaluated for contrast absorption from the peritoneal cavity. RESULTS: Evaluation of indicators of pulmonary edema demonstrated that iodixanol caused the same or less pulmonary edema than comparative agents. Histopathologic analysis showed that iodixanol caused less macrophage response than saline (P = 0.010), the same lymphocyte infiltration as saline (P = 0.472), the same neutrophil response as saline (P = 0.297), and the same vasculitic reaction as saline (P = 0.128). Compared to iohexol 270, iodixanol caused the same macrophage infiltrate (P = 0.924), the same lymphocyte infiltration (P = 0.523), more neutrophil reaction (P = 0.007), and less vasculitic reaction (P = 0.042). Iodixanol was rapidly absorbed from the peritoneal cavity. CONCLUSION: Iodixanol is a new contrast agent that is isotonic at all clinically useful iodine concentrations. It appears safe in the lung and is absorbed from the peritoneal cavity. PMID- 10525778 TI - High-resolution computed tomography of the chest in children with cystic fibrosis: support for use as an outcome surrogate. AB - BACKGROUND: Outcome surrogates are indicators that reflect, rather than directly measure, patient benefit. In order to provide useful results, however, outcome surrogates must be carefully chosen and must meet specific criteria. OBJECTIVE: To support development of high-resolution computed tomography (HRCT) as an outcome surrogate in cystic fibrosis (CF) by demonstrating the ability of HRCT to show short-term improvement in the appearance of the lungs in children with CF. MATERIALS AND METHODS: HRCT was performed at admission and after discharge on 8 children during 15 admissions for acute pulmonary exacerbation of CF. Three radiologists scored each study separately, then compared admission and discharge pairs. RESULTS: HRCT scores improved in 13/15 admissions. Mean score decreased from 25 to 22. The decrease was significant (P = 0.014). Comparison of admission and discharge scans showed improvement in peribronchial thickening (P = 0.007), mucous plugging (P = 0.002), and overall appearance (P = 0.025). CONCLUSION: HRCT has the potential to be a useful outcome surrogate in CF. A necessary attribute of an outcome surrogate is that it improves rapidly with effective therapy. Despite widespread belief among radiologists and pulmonologists that HRCT meets this criterion, no previous report has demonstrated this ability in children. These findings support further development of HRCT as an outcome surrogate in children with CF. PMID- 10525779 TI - Significance of thickening of the wall of the renal collecting system in children: an ultrasound study. AB - OBJECTIVE: To evaluate the significance of thickening of the wall of the renal collecting system by US. MATERIALS AND METHODS: Wall thickening of the renal collecting system was seen during US in 62 collecting systems of 51 patients over a period of 2 years. The medical and radiological records of these patients were reviewed with special attention to the definitive diagnosis and other clinical and radiological parameters. Moreover, a control group consisting of 48 renal collecting systems was examined to establish normal values for the thickness of the wall of the collecting system. RESULTS: Of the 62 collecting systems (mean wall thickness 1.6 mm, range 0.8-3.1 mm), vesicoureteric reflux (VUR) was present in 18 cases, urinary tract infection (UTI) in 11, and both VUR and UTI in 9 cases. In 10 cases, intermittent dilatation was present caused by primary obstructive megaureter (n = 2), pelvi-ureteric junction stenosis (n = 4), high pressure bladder (n = 3), or of unknown cause (n = 1). In 11 cases, transient dilatation had been present in the recent past (usually prenatally detected hydronephrosis), but had disappeared at the time of the US examination. In 3 patients, no definite cause for the wall thickening could be established. In the control group, wall thickness ranged from 0.1 to 0.8 mm. CONCLUSIONS: The upper limit for wall thickness of the normal collecting system in children is 0.8 mm. Thickening of the wall of more than 0.8 mm should be considered as pathological and is caused by urinary tract infection, intermittent dilatation (e. g., VUR), and dilatation in the recent past. PMID- 10525780 TI - Bilateral ovarian involvement at presentation in metastatic (stage 4) neuroblastoma. PMID- 10525782 TI - The development of hypertrophic pyloric stenosis in a patient with prostaglandin induced foveolar hyperplasia. AB - BACKGROUND: Hypertrophic pyloric stenosis (HPS) has been described in association with several obstructive antropyloric lesions including idiopathic foveolar hyperplasia (gastric mucosal hypertrophy), feeding tubes, eosinophilic gastroenteritis, and hypertrophic antral polyps. Non obstructive antral webs have also been described with HPS. PATIENT AND METHODS: We present a case of gastric outlet obstruction in association with HPS, namely, prostaglandin-induced foveolar hyperplasia. This entity has been previously described, but rarely in association with HPS. We report a female infant requiring prostaglandin therapy for pulmonary atresia who developed dose-related prostaglandin-induced foveolar hyperplasia and symptoms of progressive non-bilious vomiting. RESULTS: Initially, ultrasonography demonstrated evidence of antral mucosal hypertrophy as the cause for gastric-outlet obstruction. The patient subsequently developed progressive thickening of the antropyloric muscle, resulting in sonographic appearances of hypertrophic pyloric stenosis. Pyloromyotomy was eventually required for treatment of HPS. CONCLUSION: A common denominator of most of the above-described entities is thickening and/or hypertrophy of the antral mucosa. We suggest that the antropyloric musculature may hypertrophy in an effort to overcome the gastric outlet obstruction caused by the adjacent thickened antral mucosa. In other words, these entities may represent examples of "secondary" hypertrophic pyloric stenosis. PMID- 10525781 TI - Shortcomings of diuresis scintigraphy in evaluating urinary obstruction: comparison with pressure flow studies. AB - BACKGROUND: In at least 15 % of dilated urinary tracts, diuresis renography fails to assess the presence or absence of urinary obstruction. OBJECTIVE: To determine the shortcomings of (99 m)Tc-DTPA frusemide diuresis renography by reference to pressure flow studies. MATERIALS AND METHODS: Thirty-four patients, aged 1 month to 20 years, with questionable obstruction were evaluated by diuresis renography and pressure flow studies (the Whitaker test) as the reference method during the same short period of time. Discrepancies were analysed. RESULTS: In patients with type I or IIIa renographic response, pressure flow studies never led to any change in management. Poor function, major dilatation and prior surgery were found to be risk factors of inaccurately obstructive pattern (type II) on renography (n = 6). In patients with type IIIb response, pressure flow studies could show low-grade (n = 3) or intermittent obstruction (n = 2). Intermittent obstruction was also demonstrated in two patients with type II response. CONCLUSION: In patients with risk factors, type II response was sometimes inaccurate, and urodynamic evaluation showed absence of obstruction and led to conservative management. Type IIIb response should be considered equivocal rather than partially obstructive, and pressure flow studies could be considered in such patients. PMID- 10525783 TI - Intestinal intussusception survey about diagnostic and nonsurgical therapeutic procedures. AB - OBJECTIVE: To provide an overview of the diagnostic and therapeutic procedures performed by European paediatric radiologists in the management of intussusception. MATERIALS AND METHODS: A postal survey was sent to the European members of ESPR. Items surveyed included diagnostic imaging procedures (plain films, US, contrast enema [CE]), contrast medium used (barium, iodine, air, saline solution), and imaging technique used for monitoring during reduction (films, fluoroscopy, US). Multiple answers were possible. Other data, including contraindications, maximum pressure, pressure and irradiation monitoring, presence of a surgeon, sedation, number and duration of attempts, and hospitalisation were also obtained and analysed. RESULTS: There were 204 respondents (60.2 %). Regarding diagnosis, 72.5 % of respondents used plain radiographs, 93 % US, and 34 % CE. Reduction was performed using air (55 %), a barium suspension (32 %), iodinated contrast medium (24 %), or a saline solution (10 %). Reduction was monitored using fluoroscopy alone (46 %), fluoroscopy and radiographs (49.5 %), US alone (9.5 %), or a combination of radiology and US (18 %). Pressure was monitored by 81 % of respondents. Most respondents (82.4 %) used a maximum pressure between 100 and 120 mm Hg. CONCLUSIONS: US is widely used for diagnosing intussusception. For treatment, contrast medium and air reduction are used almost equally. A large number of radiologists are now performing intussusception reduction using US monitoring. PMID- 10525784 TI - Laryngeal penetration: a predictor of aspiration in infants? AB - BACKGROUND: We routinely observed "isolated" laryngeal penetration (ILP) on upper GI studies in infants with no risk for aspiration. OBJECTIVE: To determine whether "isolated" LP (LP during the pharyngeal phase of swallowing) is a benign process in infants and therefore not a predictor of aspiration as it is known to be in adults. PATIENTS AND METHODS: Two radiologists retrospectively reviewed videotaped upper GI studies done over a 2-year period on patients less than 2 years of age. A total of 110 studies was reviewed, which produced a study group of 34 patients who had no history or clinical suspicion for swallowing dysfunction. RESULTS: Of the 34 patients in the group, 33 demonstrated ILP without radiographic or clinical evidence of aspiration. CONCLUSION: ILP is a benign, normal process in infants likely due to immaturity of the swallowing mechanism. It is not a reliable predictor of aspiration as it is in adults. PMID- 10525785 TI - MRI of three siblings with Sjogren-Larsson syndrome. AB - Sjogren-Larsson syndrome (SLS) is a rare disorder with autosomal recessive inheritance. Its clinical, pathological, genetic, and biochemical manifestations have been thoroughly evaluated, but there is little imaging data, especially regarding MRI. We present brain MRI of three siblings with SLS and discuss our findings. PMID- 10525786 TI - Radiation dose reduction in paediatric cranial CT. AB - BACKGROUND: There is no consensus about the optimal milliamperage-second (mAs) settings for computed tomography (CT). Most operators follow the recommended settings of the manufacturers, but these may not be the most appropriate settings. OBJECTIVE: To determine whether a lower radiation dose technique could be used in CT of the paediatric brain without jeopardising the diagnostic accuracy of the images. MATERIALS AND METHODS: A randomised prospective trial. A group of 53 children underwent CT using manufacturer's default levels of 200 or 250 mAs; 47 underwent scanning at 125 or 150 mAs. Anatomical details and the confidence level in reaching a diagnosis were evaluated by two radiologists in a double-blinded manner using a 4-point scoring system. RESULTS: For both readers there was no statistically significant difference in the confidence level for reaching a diagnosis between the two groups. The 95 % confidence intervals and P values were -0.9-1.1 and 0.13 (reader 1) and -1.29-1.37 and 0.70 (reader 2), respectively. Reliability tests showed the results were consistent. CONCLUSIONS: The recommended level may not be the optimum setting. Dose reduction of 40 % is possible on our system in paediatric brain CT without affecting the diagnostic quality of the images. PMID- 10525787 TI - Congenital subependymal giant-cell astrocytoma: case report with prenatal ultrasonogram. AB - BACKGROUND: A prenatal sonogram at 27 weeks of gestation revealed a brain mass along the frontal horn and body of the lateral ventricle near the foramen of Monro in the fetus. MATERIALS AND METHODS: A huge subependymal giant-cell astrocytoma was nearly totally resected at 11 days of age. RESULTS: There was no syndromic family history, but features substantiating the diagnosis of tuberous sclerosis were recognized at 4 years of age. CONCLUSION: The sonographic finding of a tumor in the region of the foramen of Monro should raise the suspicion of a subependymal giant-cell astrocytoma, a tumor characteristically associated with tuberous sclerosis. PMID- 10525788 TI - Anomalies of ossification in the posterolateral femoral condyle: assessment by MRI. AB - BACKGROUND: Anomalies of ossification in the lower femoral epiphysis are often radiographically indistinguishable from juvenile osteochondritis dissecans. OBJECTIVE: To clarify the MRI characteristics of the anomalies of ossification in the posterolateral femoral condyle that distinguish it from juvenile osteochondritis dissecans. MATERIALS AND METHODS: We retrospectively examined the medical records, plain radiographs (n = 4), MRI (n = 4) and follow-up MRI (n = 2) of four boys (age 8-11 years) with anomalies of ossification in the posterolateral femoral condyle. RESULTS: Plain radiography showed symmetrical marginal irregularity of the posterolateral femoral condyles of both knees. These lesions were asymptomatic, and the areas of irregular radiographic appearances reduced in size or disappeared without treatment within a mean observation period of 3.5 months. MRI showed a clearly demarcated low-intensity islet with the same signal intensity as subchondral bone (which was considered to be an accessory ossification nucleus) in a high-signal area in which the signal intensity was equal to that of normal articular cartilage. The areas observed as radiolucent zones on plain radiography were visualised at the same signal intensity as articular cartilage, and were continuous with articular cartilage on MRI; thus they were regarded as uncalcified cartilage. These MR findings are different from MR images of osteochondritis dissecans. CONCLUSIONS: MRI is considered to be the most effective non-invasive diagnostic method for these two conditions. PMID- 10525790 TI - Subcutaneous fat necrosis of the newborn. AB - We present the MRI findings in a case of subcutaneous fat necrosis of the newborn. To our knowledge, the MRI findings of this entity have not been reported. Subcutaneous fat necrosis of the newborn is an uncommon, benign process in full-term infants. Hypercalcemia may be a potentially life-threatening complication of this otherwise self-limiting process. PMID- 10525789 TI - Imaging following allograft reconstruction in children with malignant bone tumours. AB - PURPOSE: To determine the nature of the imaging findings following reconstructive surgery using massive allografts in children with malignant bone tumours. MATERIALS AND METHODS: A retrospective review of the imaging studies and medical charts of 25 consecutive children who received an allograft as part of the management of a malignant bone tumour. RESULTS: Uncomplicated allografts were sclerotic relative to native bone on radiographs and showed a typical 'tramline' appearance on bone scintigraphy. On MR, the medullary canal of the allograft showed low signal, similar to or greater than skeletal muscle, but less than subcutaneous fat, on 91 % of T1-weighted images. On short-tau inversion recovery images, the medullary canal was inhomogeneous and hyperintense to subcutaneous fat in 70 % and hyperintense to muscle in the remainder. Complications occurred in 68 % of patients and included allograft fractures (36 %), recurrent tumour (20 %), infection (8 %), and non-union or delayed union (8 %). The radiographic findings alone permitted accurate diagnosis of most serious complications. Infection and rejection were difficult to distinguish with any technique. All complications were suspected on clinical and/or radiological grounds before being shown by MR or scintigraphy. CONCLUSIONS: Allografts, whether normal or complicated, have characteristic imaging findings, except that infection and bone resorption related to rejection and revascularisation are difficult to distinguish. Routine MR and bone scintigraphy appear to contribute little to the management of these patients. PMID- 10525791 TI - Periosteal reaction of the long bones associated with extracorporeal membrane oxygenation: cause and effect? PMID- 10525792 TI - The pathology of total joint arthroplasty.II. Mechanisms of implant failure. AB - Although the clinical results of total joint arthroplasty are usually excellent, some implants develop loosening and require revision. Implants usually fail by a combination of mechanisms, but different basic designs tend to show different dominant mechanisms of failure. Infection causes failure of about 1-5% of cases of primary arthroplasty. Clues to the presence of infection include clinical signs, a periosteal reaction, a positive culture of aspirated joint fluid, and acute inflammation identified in tissue around the implant. There are several different mechanisms and modes of implant wear, and perhaps the most important cause of aseptic loosening is an inflammatory reaction to particles of wear debris. Abrasive, adhesive, and fatigue wear of polyethylene, metal and bone cement produces debris particles that induce bone resorption and implant loosening. Particles can cause linear, geographic, or erosive patterns of bone resorption (osteolysis), the distributions of which are influenced by the implant design. Micromotion of implants that did not achieve adequate initial fixation is another important mechanism of loosening. Fatigue failure at the bone/cement and bone/implant interface may cause aseptic loosening, and may be especially important for implants with relatively smooth surfaces. Stress shielding can influence local bone density, but is rarely an isolated cause of implant loosening. Elevated hydrodynamic pressure has been associated with bone resorption in the absence of implants, and may also play a role in implant loosening. PMID- 10525793 TI - Masses and pseudomasses of the hand and wrist: MR findings in 134 cases. AB - OBJECTIVE: To assess the utility of magnetic resonance imaging (MRI) in the investigation of palpable masses in the hand or wrist. DESIGN AND PATIENTS. We retrospectively reviewed the MRI examinations and case records of 134 patients referred because of a palpable mass in the hand or wrist. MRI was performed on a 1.0 T magnet using an extremity coil. Intravenous gadolinium-DTPA was injected when considered appropriate. RESULTS AND CONCLUSIONS: MRI demonstrated the cause of the palpable mass in 126 cases (94.02%). Soft tissue neoplasms were found in 34 cases (25.37%). The majority were benign and included giant cell tumours of tendon sheath, lipomas and hemangiomas and had a characteristic appearance. There were three malignant tumours (myxoid liposarcoma, malignant fibroushistiocytoma and rhabdomyosarcoma). Ganglia were found in 36 cases (26.86%) and non-tumour tendon pathology in 31 cases (23.13%). Less common causes included articular diseases (5.97%) and anatomical variants (4.47%). No focal lesion was present in 8 cases (5.97%). In conclusion, MRI is an accurate diagnostic technique in patients who present with a palpable mass of the hand and wrist. PMID- 10525794 TI - Evaluation of the postoperative meniscus of the knee: a study comparing conventional arthrography, conventional MR imaging, MR arthrography with iodinated contrast material, and MR arthrography with gadolinium-based contrast material. AB - OBJECTIVE: To compare four imaging methods in the evaluation of the postoperative meniscus: conventional arthrography, conventional MR imaging, MR arthrography with iodinated contrast material, and MR arthrography with gadolinium-based contrast material. DESIGN AND PATIENTS: Thirty-three patients referred for knee MR examinations with a history of meniscal surgery were studied prospectively. At the first patient visit, conventional MR examination was followed by an MR arthrogram with gadolinium-based contrast material. At the second visit, a conventional arthrogram with iodinated contrast material was followed immediately by an MR examination. Imaging examinations were interpreted by a masked reader, and then compared with the results of repeat arthroscopic surgery in 12 patients. RESULTS: The correct evaluation of the status of postoperative menisci was allowed in 12 of 13 patients (92%) by MR arthrography using gadolinium-based contrast agent, 10 of 13 patients (77%) by conventional MR examination, 9 of 12 patients (75%) by MR arthrography, and 7 of 12 patients (58%) by conventional arthrography. CONCLUSION: Intra-articular fluid is advantageous in the evaluation of patients with a suspected meniscal retear. MR arthrography with gadolinium based contrast material is the most accurate imaging method for the diagnosis of meniscal retears. PMID- 10525796 TI - Osteosarcoma in a patient with McCune-Albright syndrome and Mazabraud's syndrome. AB - Sarcomas infrequently develop in osseous sites of fibrous dysplasia. We report a patient with Mazabraud's syndrome (polyostotic fibrous dysplasia and soft tissue myxomas) complicated by the development of osteogenic sarcoma in a bone affected by fibrous dysplasia. This is the third case of osteosarcoma within the small population of reported patients with Mazabraud's syndrome. There may be an increased incidence of malignant transformation in these individuals' dysplastic bones above that associated with patients suffering from fibrous dysplasia alone. PMID- 10525795 TI - Intramuscular vascular malformations of an extremity: findings on MR imaging and pathologic correlation. AB - OBJECTIVE: To analyze the findings of intramuscular vascular malformations of an extremity on MR imaging and to correlate these findings with histopathologic examination. DESIGN AND PATIENTS: The findings on MR imaging and the medical records of 14 patients with an intramuscular vascular malformation of the extremity were retrospectively studied. All patients underwent surgical excision. Diagnoses were based on the results of pathologic examination. Findings on MR imaging were noted and correlated with the histopathologic findings. RESULTS: Intramuscular vascular malformations of an extremity showed multi-septate, honeycomb, or mixed appearance on MR imaging. Multi-septate areas correlated with dilated and communicating vascular spaces with flattened endothelium. Honeycomb areas corresponded to vascular spaces with inconspicuous small lumina and thickened vascular walls. Areas of increased signal intensity on T2-weighted images were found in all intramuscular vascular malformations. Infiltrative margins were more commonly seen in intramuscular lymphaticovenous malformations. Adherence to neurovascular structures and orientation of the lesion along the long axis of the affected muscle were more commonly seen in intramuscular venous malformations. CONCLUSIONS: Intramuscular vascular malformations showed either a multi-septate, honeycomb, or mixed appearance, reflecting the size of the vascular spaces and the thickness of the smooth muscles of the vessel walls. Prediction of the subtype of an intramuscular vascular malformation of an extremity on MR imaging seems to be difficult, although there are associated findings that may be helpful in the differential diagnosis of each subtype. PMID- 10525797 TI - Dedifferentiated parosteal osteosarcoma with rhabdomyosarcomatous differentiation. AB - Dedifferentiated parosteal osteosarcomas are characterized histologically by the combination of low-grade fibroblastic osteosarcoma admixed with a high-grade component that typically has the appearance of malignant fibrous histiocytoma or osteosarcoma. Herein we report a case of dedifferentiated parosteal osteosarcoma of the distal femur, in which the high-grade component consisted of rhabdomyosarcoma. To our knowledge, a rhabdomyosarcomatous component has not been described previously in a dedifferentiated parosteal osteosarcoma. The clinical, radiologic, and pathologic features of this rare type of surface osteosarcoma are described. PMID- 10525799 TI - Neurilemmoma of the mandible. AB - Intraosseous neurilemmoma is rare, representing less than 1% of benign primary bone tumors. The site most commonly involved is the mandible. We report on a neurilemmoma of the mandible in a 30-year-old woman. A panoramic radiograph of the mandible showed a well-defined unilocular osteolytic lesion with a thin uniform sclerotic margin located in the body of the mandible. The CT scan confirmed a well-defined osteolytic lesion with thinning of the cortex of the body of the left side of the mandible. Histologically, the lesion was a moderately cellular neoplasm with distinct palisading and numerous Verocay bodies. Ultrastructurally, the cytoplasmic membranes were distinct and coated by amorphous bands of basal lamina. Complete excision was achieved by removing the tumor from the inferior alveolar nerve. PMID- 10525798 TI - Popliteal vascular malformation simulating a soft tissue sarcoma. AB - Differentiation of vascular abnormalities from soft tissue sarcomas may be difficult on clinical grounds, but is usually possible on imaging criteria. We report the MRI and digital subtraction angiography (DSA) findings in a patient presenting with a mass behind the knee. We discuss differentiating features and review the literature of similar cases. PMID- 10525800 TI - Malignant degeneration of an osteochondroma with unusual intra-bursal invasion. AB - Multiple hereditary osteochondromatosis is an uncommon autosomal dominant condition in which patients are predisposed to the development of chondrosarcoma. We report a case of a patient who developed a secondary low-grade chondrosarcoma in this setting. The tumor was associated with an unusual multinodular invasive growth pattern into a pre-existing bursa that was present overlying the osteochondroma. PMID- 10525801 TI - Laparoscopic adjustable silicone gastric banding: radiological appearances of a new surgical treatment for morbid obesity. AB - BACKGROUND: The purpose of this report is to describe the radiologic appearances of laparoscopic adjustable silicone gastric banding (LASGB), a new surgical treatment for morbid obesity. In this procedure, a silicone band is fastened around the fundus, delimitating a small proximal gastric pouch and stoma. The inner surface of the band is inflatable and connected by a thin silicone tube to an access port. This allows postoperative stoma size adjustment by puncturing the port and injecting or withdrawing saline solution. METHODS: One hundred eighty patients underwent LASGB. A radiologic study protocol was established and performed in all patients, including preoperative double-contrast upper gastrointestinal (GI) series and single-contrast upper GI series on the first postoperative day and 1 month after surgery. Radiologic evaluation was also performed at each band adjustment and in case of persistent vomiting or inadequate weight loss. RESULTS: Postoperative stoma adjustment was performed in all patients. The optimal volume of saline was 1-4.5 mL. Percutaneous puncture of the port was impossible in three patients because of an inverted port. We observed 15 cases of pouch dilatation with stomal obstruction requiring reoperation. There were also nine cases of spontaneous band deflation caused by leaking reservoir in five cases and by disconnection between the connecting tube and the port in the other four cases. CONCLUSIONS: Because radiologic evaluation is necessary after surgery and for band adjustments, radiologists are involved in the postoperative follow-up and may be asked to perform those adjustments themselves. PMID- 10525802 TI - Recurrent Crohn's disease in the duodenum and jejunum following extensive small bowel resection and jejunocolonic anastamosis: radiologic findings in twenty-five patients. AB - BACKGROUND: To evaluate the radiologic features of recurrent Crohn's disease after extensive enteric resection and jejunocolostomy. METHODS: We reviewed the small bowel studies of 25 patients with recurrent enteritis and less than 125 cm of jejunum following enteric resection and jejunocolostomy and the studies of 27 patients with jejunitis in an intact jejunum. RESULTS: Twenty-three patients with recurrences had neoterminal jejunitis, six under 10 cm, 10 over 10 cm and continuous, and seven with skip lesions (six jejunal, one duodenal). Two had isolated jejunitis or duodenitis. Three with continuous disease had lengthy recurrences. Enteritis showed only one or two abnormalities in 12 of 25 patients with recurrences and in two of 27 with disease in the intact jejunum. Recurrent jejunitis and jejunitis in the intact jejunum showed similar frequencies of mucosal thickening, strictures, ulceration and its complications, skip lesions, sacculation, obstructive dilatation, featureless mucosa, and polyps, and significantly different frequencies only of mesenteric masses. Recurrent jejunitis and terminal ileitis showed significantly different frequencies of mucosal thickening, strictures, ulceration and its complications, skip lesions, sacculation, obstructive dilatation, and mesenteric masses, and similar frequencies only of a featureless mucosa. CONCLUSIONS: The neoterminal jejunum is the most common site of recurrence and the only site in almost 25%. Jejunitis remote from the fecal stream is also frequent, but duodenitis is not. Recurrences are seldom extensive and often show only one or two radiographic findings. The frequencies of most lesions in recurrent jejunitis do not differ significantly from those in jejunitis in the intact jejunum but do differ from those in terminal ileitis. PMID- 10525803 TI - Spiral CT of the abdomen after distention of small bowel loops with transparent enema in patients with Crohn's disease. AB - BACKGROUND: [corrected] To evaluate the capability of a computed tomographic (CT) technique that combines distention of the small bowel loops with a transparent enema with contrast-enhanced spiral CT of the abdomen in patients with Crohn's disease. METHODS: We evaluated the abdomen with spiral CT after distention of the small bowel loops with a transparent enema of methylcellulose in 40 patients consecutively referred for radiologic evaluation of Crohn's disease of the small bowel. Fluid was infused through a nasojejunal catheter with a peristaltic pump. Ultrasonography was used to prevent bowel overdistention and detect arrival of methylcellulose to the cecum. Contrast-enhanced spiral CT of the abdomen was then performed, and the degree of contrast enhancement and the thickness of the walls of the involved loops were evaluated. A series of 10 patients with retrograde distention of the last ileal loop from large bowel water enema was used as a control. The results of the CT were compared with those of conventional radiographic small bowel studies. RESULTS: The normal small bowel wall was 1.9 2.5 mm thick (mean = 2. 1 mm); density values of the normal enhanced wall varied between 25 and 60 HU (mean = 32 HU) and presented a homogeneous structure. Bowel segments involved by the disease were 4-12.5 mm thick (mean = 9.2 mm), had density values of 75-150 HU (mean = 105 HU), and showed a multilayered appearance. Compared with conventional radiography, CT detected longer lesions or additional segments involved by the disease process in 14 cases, 11 additional fistulas, two abscesses, and mesenteric changes in 21 cases. CONCLUSIONS: The small bowel CT enema technique provides good results in the study of patients with Crohn's disease and can be used to evaluate patients with advanced lesions. PMID- 10525804 TI - Intestinal malrotation as an incidental finding on CT in adults. AB - BACKGROUND: Intestinal malrotation in adults is usually an incidental finding on computed tomography (CT). We present the CT findings of 18 adult patients with malrotation and discuss the clinical implications. METHODS: Abdominal scans of 18 patients (12 women, six men; age range = 15-79 years) with intestinal malrotation were reviewed. Special attention was directed to the location of the superior mesenteric vessels, the location of the small and large bowels, the size of the uncinate process, the situs definition, and additional anomalies. RESULTS: The malrotation was an incidental finding in all but one patient. The malrotation was type Ia in 17 patients and IIc in the one symptomatic patient. The superior mesenteric vessels were vertically oriented in 10, inverted in two, normally positioned in four, and mirror imaged in two cases with situs ambiguus. All patients had aplasia of the pancreatic uncinate process, five had a short pancreas, and two had a preduodenal portal vein. Fourteen patients had a normal situs and four had heterotaxia. Seven patients had polysplenia, six of which with associated inferior vena cava anomalies. CONCLUSIONS: Intestinal malrotation can be diagnosed on CT by the anatomic location of a right-sided small bowel, left sided colon, an abnormal relationship of the superior mesenteric vessels, and aplasia of the uncinate process. Awareness of these abnormalities is necessary to diagnose this anomaly. It should be sought in patients with a situs problem, inferior vena cava anomalies, polysplenia, or preduodenal portal vein. Although usually an incidental finding, it is important to diagnose such a malrotation because it may cause abdominal symptoms. Also, knowledge of associated vascular anomalies is important when abdominal surgery is planned. PMID- 10525805 TI - Middle mesenteric artery visualized by computed tomographic angiography. AB - The middle mesenteric artery, a third mesenteric artery arising from the aorta that principally feeds the transverse colon, is an extremely rare anomaly. We identified a middle mesenteric artery branching into the ileocolic artery and into the right, middle, and accessory middle colic arteries. It supplied the cecum and the entire ascending and transverse colon. This anomaly was detected with computed tomographic angiography. PMID- 10525806 TI - Sonography of pneumatosis cystoides intestinalis. AB - Pneumatosis cystoides intestinalis (PCI) is a relatively rare benign condition, and its sonographic findings have rarely been reported. We report on four cases of PCI in which sonography showed multiple immobile linear or spotty high echoes in the thickened colonic wall. These sonographic findings were more clearly visualized by using high-frequency probes and helped in establishing the diagnosis. In addition, color Doppler sonography confirmed the absence of portal gas and helped rule out fulminant PCI. When encountering patients with abundant abdominal gas, the possibility of PCI should be considered and the colonic wall and the portal vein should be meticulously observed by high-frequency probe and color Doppler sonography to prevent a delay in the diagnosis and to improve patient management. PMID- 10525807 TI - Vaginal opacification during defecography: utility of placing a folded gauze square at the introitus. AB - We evaluated the value of placement of a folded gauze square into the urogenital introitus to improve vaginal opacification in 90 patients who underwent defecography. Of the 50 patients who retained the gauze in the introitus, 96% demonstrated excellent or good vaginal opacification. By contrast, only 75% of the 40 patients who lost the gauze during the study were able to achieve the same level of opacification. This difference was shown to be statistically significant (p < 0.002), suggesting that placement of a folded gauze square in the introitus limits loss of contrast from the vagina, which improves vaginal opacification. PMID- 10525808 TI - Vaginal opacification during defecography: direction of vaginal migration aids in diagnosis of pelvic floor pathology. AB - BACKGROUND: To determine whether direction of vaginal displacement during defecography aids in diagnosing pelvic floor pathology. METHODS: Ninety patients underwent defecography over a 2-year period. Each study was retrospectively reviewed by three radiologists who recorded whether the vagina was displaced cephalad, caudad, or nondisplaced in relation to the urogenital hiatus. This information was then correlated with radiologic diagnosis rendered for the study. RESULTS: Of the 26 patients with normal defecograms, 19 (73%; p < 0. 001) demonstrated no vaginal displacement during the procedure. Comparatively, 10 (83%; p < 0.001) of the 12 patients with cystoceles showed caudad vaginal displacement, and no patients with cystoceles showed cephalad displacement of the vagina. Of the 17 patients with rectoceles, 10 (58%) showed cephalad displacement, one (6%) showed caudad displacement, and six (35%) patients showed no vaginal displacement. Thirteen (46%) of 28 patients with enteroceles showed cephalad vaginal displacement, nine (32%) showed no vaginal displacement, and six (21%) demonstrated caudad displacement. CONCLUSIONS: Caudad displacement of the opacified vagina suggests the presence of a cystocele. Cephalad vaginal displacement is suggestive of the presence of an enterocele or rectocele. PMID- 10525809 TI - Anal endosonography after sphincter repair: specific patterns related to clinical outcome. AB - BACKGROUND: This study evaluates the endosonographic patterns of the anal sphincter after sphincteroplasty to define specific postoperative findings and to identify factors related to clinical outcome after sphincter repair. METHODS: Thirty-one incontinent patients (29 women, two men; mean age = 57 years) who underwent surgical repair for an external sphincter defect were studied postoperatively by endosonography. Twenty patients were found to improve after surgery. RESULTS: Postoperative endosonograms showed specific images: direct visualization of the surgical process was represented by the "overlapping sign" in 17 cases and the "end-to-end suture" in four cases. These echographically favorable cases were associated with improvement after surgery in 18 of 21 patients (p < 0.005). Persistent defects were reduced in five patients and unchanged in five other patients and were associated with poor outcome in eight of 10 patients (p < 0.005). CONCLUSIONS: Postoperative endosonography of the external anal sphincter presented some specific endosonographic aspects. The association between anal endosonographic findings and clinical outcome suggests the use of this procedure to assess patients following sphincteroplasty. PMID- 10525810 TI - Anal endosonography after sphincter repair. PMID- 10525811 TI - High-resolution magnetic resonance imaging of the anorectal region without an endocoil. AB - BACKGROUND: To evaluate the feasibility of a high-resolution magnetic resonance imaging (MRI) technique in detailed imaging of the anal sphincter and lower pelvic region without the use of an endoluminal coil. METHODS: MRIs with an external phased array coil (T1- and T2-weighted turbo spin echo) were performed in 22 volunteers and 12 patients with an anal fistula, an anal sphincter defect, or a rectal tumor. The normal scans were evaluated by three independent observers. The scans of the patients were compared with surgical and/or histologic findings. RESULTS: The anal sphincter was visualized with detail. In the anal canal, hemorrhoidal tissue and the submucosae ani muscle could be seen. The MRI technique also allowed detailed imaging of anatomical structures above the pelvic floor. The MR findings in the 12 patients showed exact correlation with surgery and/or histology. CONCLUSIONS: High-resolution MRI of the anorectal region without an endoluminal coil is feasible. The MR technique with an external phased array coil allows detailed imaging of the anal sphincter at rest, the rectum, and the surrounding pelvic structures with one single investigation. The results are promising and suggest useful applications in the management of anorectal diseases. PMID- 10525813 TI - Portal tumor thrombus due to gastrointestinal cancer. AB - METHODS: We studied the clinical data of seven patients with portal tumor thrombus (PTT) due to gastrointestinal (GI) cancer to determine the radiologic patterns and clinical implications of this rare complication. RESULTS: (a) PTT was located along the entire splenic vein in three cases, at the splenomesenteric confluence in one case, and in the superior mesenteric vein in one case. Intrahepatic PTT occurred in two of four cases with liver metastasis. (b) One cirrhotic case was complicated by the occurrence of colon cancer associated with PTT in the splenic vein; the esophageal varices became rapidly enlarged and poorly controlled, and the patient died due to repeated variceal rupture. (c) In all patients, abdominal sonography (US) detected PTT and color Doppler sonography confirmed the US findings. CONCLUSIONS: The splenic vein should be meticulously observed by color Doppler sonography to check for PTT in patients with GI cancer to improve patient care. PMID- 10525814 TI - Right hepatic arterial supply to the posterior aspect of segment IV of the liver: analysis by CT during hepatic arteriography. AB - OBJECTIVE: To examine the frequency of the right hepatic arterial supply to the posterior aspect of segment IV (PASIV) of the liver shown on computed tomography (CT) during hepatic arteriography (CTHA). MATERIALS AND METHODS: Seventy-four patients who underwent CTHA from the right and/or left hepatic artery were studied. The right arterial supply to the PASIV was determined when the PASIV was stained on CT during right hepatic arteriography without any opacified arteries originating from the right hepatic artery and distributing to segment IV through the left hepatic hilum or when no staining was seen in the PASIV on CT during left hepatic arteriography. The frequency of the right hepatic arterial supply to the PASIV demonstrated on CTHA was analyzed. RESULTS: In six of 74 patients (8%), the PASIV was supplied from the right hepatic artery. CONCLUSION: This PASIV was supplied by the right hepatic artery in a significant proportion of cases. PMID- 10525815 TI - Hepatic infarction in preeclampsia as part of the HELLP syndrome: CT appearance. AB - We describe the computed tomographic (CT) findings of hepatic infarctions in two preeclamptic pregnant women. These infarcts were part of the HELLP syndrome (hemolysis, elevated liver function tests, and low platelets count). In both cases, CT disclosed features characteristic of multiple nonenhancing, low attenuation, peripheral lesions with vessels coursing through and a mottled appearance. The recognition of such CT findings in liver disease associated with preeclampsia can establish the correct diagnosis. PMID- 10525816 TI - Hepatic CT enhancement: effect of the rate and volume of contrast medium injection in an animal model. AB - BACKGROUND: To evaluate the relative effect of rate of injection and volume of contrast medium on aortic, portal, and hepatic enhancement during computed tomography (CT). METHODS: Thirty-eight nonincremental CT examinations were performed in three mini-pigs by using a combination of three different volumes (1.5, 2, and 3 mL/kg) and five different rates (1.5, 3, 4.5, 6, and 7.5 mL/s) of contrast material injection. Time-density enhancement curves of the aorta, portal vein, and liver were plotted over time for each rate of injection, each volume of contrast, and each volume-rate combination. In addition, aortic, portal, and liver peak enhancements, time-to-peak enhancements, optimal scanning intervals, and contrast enhancement indices were calculated for each volume-rate combination. RESULTS: Higher rates of injection increased peak aortic enhancement but had no effect on peak portal or hepatic enhancement. This result may be explained by the dilution of the bolus of contrast medium in the splanchnic circulation. When the results of a 6-mL/s injection of 1.5 mL/kg of contrast material were compared with a 3-mL/s injection of 2 mL/kg, maximum aortic enhancement increased by 32%, whereas maximum liver enhancement decreased by 35%. CONCLUSION: An increase in the rate of contrast injection results in an increase of peak aortic enhancement even when the total iodine load is decreased. However, an increase of the rate of contrast injection does not increase maximum liver enhancement, which is related to the total iodine dose injected. Therefore, one cannot compensate a decrease in the iodine load by an increase in injection rate in contrast-enhanced CT of the liver. PMID- 10525817 TI - Dynamic enhancement of upper abdominal organs in normal volunteers with MRI and effects of contrast dose reduction. AB - BACKGROUND: To quantify enhancement parameters of the upper abdominal organs over time during magnetic resonance (MR) examinations and to evaluate the effect of a dose reduction of contrast medium on these parameters. METHODS: Ten volunteers underwent two separate dynamic enhanced MR examinations with 0.1 and 0.075 mmol/kg of contrast medium, respectively. Breath-hold gradient-echo T1-weighted images were acquired every second for 118 s followed by delayed images. The percentages of enhancement, the time to maximum enhancement, and the area under the time-versus-enhancement curve were calculated for each organ. RESULTS: The mean times to maximum percentage of enhancement were less than 25 s for the pancreas, kidneys, and spleen and 50 s for the liver. The mean values of maximum percentage of enhancement for the standard/reduced doses were 72%/62% (pancreas), 165%/155% (kidneys), 114%/87% (spleen), and 67%/53% (liver). This difference was significant when liver enhancement was considered (p = 0.02). In addition, when the areas under the time-versus-enhancement curves were compared, the difference between the standard dose and reduced dose was significant for all organs tested (p < 0.05). CONCLUSIONS: Dynamic scanning of the upper abdomen should start early after contrast injection. Injection parameters should be standardized to capture arterial and venous enhancements in liver examinations. A 25% dose reduction did not significantly affect peak enhancement (except for the liver) but did significantly reduce overall enhancement. PMID- 10525818 TI - Ductal adenocarcinoma of the pancreas with intratumoral calcification. AB - We present two cases of ductal adenocarcinoma of the pancreas with intratumoral calcification. The two cases indicate two different etiologies for intratumoral calcification in ductal adenocarcinoma. Thus, the possibility of adenocarcinoma should be considered when a tumor with intratumoral calcification is found, although the incidence of intratumoral calcification in the ductal adenocarcinoma of the pancreas remains rare. PMID- 10525819 TI - Right anterior subphrenic space: an important site for the early detection of intraperitoneal fluid on abdominal CT. AB - BACKGROUND: Small rinds of free fluid are frequently seen in the right anterior subphrenic space. Therefore, we investigated patients undergoing abdominal computed tomography (CT) to see whether early fluid collections are seen preferentially at this or any other site. METHODS: CT examinations of 59 randomly selected patients with minor or moderate quantities of free intraperitoneal fluid were analyzed. The location of the fluid was determined [subphrenic spaces, the right hepatorenal fossa (Morison's pouch), paracolic gutters, pelvis (pouch of Douglas)]. The amount of fluid in each location was subjectively quantified (none, trace, small, or moderate). RESULTS: Free fluid was seen most frequently in the right anterior subphrenic space (44/59 patients, 75%) and pelvis (43/59 patients, 73%). Eight patients had intraperitoneal fluid at an isolated location (three in the pelvis alone, two in the left subphrenic space, one in the right subphrenic space, and one in each of the right and left paracolic spaces). CONCLUSIONS: Contrary to popular belief, intraperitoneal fluid is not always seen first in the pelvis when the patient is in the supine position. Although fluid is not consistently seen first in any specific location, the subphrenic space (especially the right anterior) is a common site where small quantities of fluid may be identified. PMID- 10525820 TI - Monitoring tumor response. PMID- 10525825 TI - Treatment of essential thrombocythemia with special emphasis on leukemogenic risk. AB - Acute leukemia and myelodysplastic syndromes are rare, but almost invariably fatal, evolutions of essential thrombocythemia (ET). Three major factors are associated with blastic transformation: cytogenetic abnormalities, myelofibrotic features, and the use of cytotoxic agents. Hematological malignancies have been reported in ET patients after treatment with alkylating agents, such as busulphan, as well as other cytoreductive drugs, such as hydroxyurea. Concerns about leukemogenicity have led some to suggest limiting the indications of these drugs to patients at higher risk of bleeding and thrombosis. Major risk factors for thrombosis are age above 60 years and a previous thrombotic event, whereas an increased bleeding tendency has been reported with platelet counts in excess of 1000-1500x10(9)/l. No myelosuppressive therapy is recommended for younger patients if they are asymptomatic or their platelet counts are below 1500x10(9)/l. The threshold of 1500x10(9)/l is controversial, however, and cytoreduction can be considered when platelets are above 1000x10(9)/l or in the presence of risk factors for cardiovascular disease. In the presence of thrombotic events or extreme thrombocytosis, young ET patients can be managed with cytoreductive agents theoretically devoid of leukemogenic risk, such as a interferon or anagrelide. Nevertheless, the mutagenic risk of anagrelide has not been investigated in long-term follow-up studies, and the ultimate place of these 'new' drugs in the management of ET patients remains to be established in prospective and controlled clinical trials. PMID- 10525826 TI - Iron status in Danes updated 1994. I: prevalence of iron deficiency and iron overload in 1332 men aged 40-70 years. Influence Of blood donation, alcohol intake, and iron supplementation. AB - Iron status, S-ferritin, and hemoglobin (Hb) were assessed in a population survey in 1994 (DAN-MONICA 10) comprising 1332 Caucasian Danish men equally distributed in age cohorts of 40, 50, 60 and 70 years. Blood donors (n=186) had lower S ferritin, median 76 microg/l, than nondonors, median 169 microg/l (p<0.0001). S ferritin in donors was inversely correlated with the number of phlebotomies (r(s)=-0.57, p<0.0001). S-ferritin in nondonors (n=1146) was similar in men 40-60 years of age, median 176 microg/l, and subsequently decreased at 70 years of age to a median of 146 microg/l (p=0.01). In the entire series, the prevalence of small iron stores (S-ferritin 16-32 microg/l) was 2.7%, that of depleted iron stores (S-ferritin <16 microg/l) 0.45%, and that of iron deficiency anemia (S ferritin <13 microg/l and Hb <129 g/l) 0.15%. Among nondonors, the prevalence of iron overload (S-ferritin >300 microg/l) was 20%. S-ferritin in nondonors correlated with body mass index (r(s)=0.19, p=0.0001) and with alcohol intake (r(s)=0.26, p=0.0001). In the entire series, 28% of the subjects took supplemental iron (median 14 mg ferrous iron daily). Iron supplements had no influence on iron status. Nondonors (n=170) treated with acetylsalicylic acid had lower S-ferritin, median 136 microg/l, than nontreated, median 169 microg/l (p<0.001) and those treated with H(2)-receptor antagonists (n=30) had lower S ferritin, median 142 microg/l, than nontreated, median 171 microg/l (p<0.04). Compared with the DAN-MONICA 1 iron status survey of Danish men in 1984, the prevalences of iron depletion and iron deficiency anemia are unchanged whereas the prevalence of iron overload has increased significantly. In Denmark, iron fortification of flour was abolished in 1987. This apparently had no negative effect on iron status in men. PMID- 10525827 TI - Serum levels of thrombopoietin, IL-11, and IL-6 in pediatric thrombocytopenias. AB - We measured serum levels of thrombopoietin (TPO), interleukin (IL)-11, and IL-6 in 90 different samples from 67 pediatric patients with thrombocytopenia (TP). The cytokine levels were determined by enzyme-linked immunosorbent assays (ELISA), and the biological activity of TPO was measured using a cell line transfected with human c-mpl. In patients with impaired megakaryocytopoiesis, as found in diseases such as aplastic anemia, amegakaryocytic TP, or TP with absent radii, we found TPO levels which were highly elevated compared with normal values (mean=261 AU/ml, n=52, vs. 22 AU/ml in healthy controls). In contrast, patients suffering from idiopathic thrombocytopenic purpura (mean=16 AU/ml, n=31) or platelet function defects (mean=23 AU/ml, n=7) demonstrated normal TPO levels. The biological activity tested in the bioassay correlated well with the ELISA data. However, sera of some patients with amegakaryocytic TP demonstrated a remarkably higher biological activity of TPO than expected from the ELISA data. Within the different groups there was no correlation between platelet counts and TPO levels. Only 27% of all samples had elevated levels of IL-11 (mean=450 pg/ml, n=20). Elevated IL-6 serum levels were detected in only 13% of all samples analyzed (mean=42 pg/ml, n=12). We conclude that megakaryocytopoiesis is regulated mainly by TPO, that it is dependent on the platelet and the megakaryocytic mass, and that IL-11 plays an additional role in supporting the platelet production. IL-6 does not appear to be up-regulated in children with thrombocytopenia. PMID- 10525828 TI - A hypothesis concerning deficiency of sunlight, cold temperature, and influenza epidemics associated with the onset of acute lymphoblastic leukemia in northern Finland. AB - Research to detect new factors contributing to the etiology of acute leukemia (AL) is urgently needed. Located between latitudes 65 degrees and 70 degrees north, the population in northern Finland is exposed to extreme seasonal alterations of ultraviolet-B light and temperature. There is also a seasonal variation of both the 25(OH)- and 1,25(OH)2-D3 vitamin serum concentrations. In the present work, the frequencies of different types and age-groups at diagnosis of AL were compared during the dark and light months of the year, to uncover seasonality. Between January 1972 and December 1986, 300 consecutive patients aged >/=16 years and diagnosed as having AL were enrolled. The observed mean monthly global solar radiation, temperature measurements, and influenza epidemics were compared with the monthly occurrence of AL. Both acute lymphoblastic leukemia (ALL) (p=0.006) and total AL (p=0.015) were diagnosed excessively in the dark and cold compared with light and warm period of the year. There was a tendency for de novo leukemia to increase also in the dark and cold, but for acute myeloid leukemia (AML) patients the excess was not significant. Age >/=65 was strongly associated with the dark and cold season (p=0.003). Significantly more ALL (p=0.005) and de novo leukemias (p=0.029) were observed during influenza epidemics than during nonepidemic periods. However, a seasonality, i. e., the fluctuation of numbers of AL cases, was not determined, either monthly or during different photo- and temperature periods or influenza epidemics; this might be due to the small numbers of patients studied. Nevertheless, it is hypothesized that sunlight deprivation in the arctic winter can lead to a deficiency of the 1, 25(OH)2D3 vitamin, which might stimulate leukemic cell proliferation and block cell differentiation through dysregulation of growth factors in the bone marrow stromal cells, causing one mutation and an overt ALL in progenitor cells damaged during the current or the previous winter by influenza virus, the other mutation. PMID- 10525829 TI - Serum levels of parathyroid hormone-related protein are not elevated in patients with T-cell prolymphocytic leukemia. AB - T-cell prolymphocytic leukemia (T-PLL) is a rare post-thymic T-cell neoplasm which shares most clinical features with adult T-cell leukemia (ATL). We measured serum level of C-terminal parathyroid hormone-related protein (C-PTHrP) in patients with T-PLL and ATL. Serum C-PTHrP levels of eight patients with T-PLL (median 36.8 pmol/l; range 27.0-50.2 pmol/l) did not differ from those of 30 human T-lymphotropic virus type I (HTLV-I)-seronegative blood donors (median 37.0 pmol/l; range 22.6-54.0 pmol/l). The C-PTHrP levels in ten ATL patients (median 69.6 pmol/l; range 42.5-899.4 pmol/l) were significantly higher than those in healthy controls (p<0.0001) or T-PLL patients (p=0.001). We suggest that the serum level of PTHrP can provide useful information for differentiating between T PLL and ATL. PMID- 10525831 TI - The platelet function analyzer (PFA-100) may not be suitable for monitoring the therapeutic efficiency of von willebrand concentrate in type III von willebrand disease. AB - We describe a type-III von Willebrand patient who was admitted to the hospital with severe deformity and functional deficit of the left knee joint due to recurrent hemarthrosis. Orthopedic intervention was necessary. To prevent bleeding episodes, von Willebrand factor (vWF) replacement therapy was given during and after surgery. APTT, plasma FVIII activity (FVIIIc), vWF antigen (vWF Ag), and vWF ristocetin cofactor (vWF Rco) were measured. Primary hemostasis was monitored using the PFA-100. This "Platelet Function Analyzer" is designed to measure platelet adhesion and aggregation capacities. Whole blood is aspirated through a capillary and is forced to flow through the central hole of a membrane coated with collagen and epinephrine (COL/EPI) or ADP (COL/ADP) as platelet activators. Irreversible platelet aggregation results in the formation of a stable platelet plug, closing the central hole. The result is expressed as "closure time" (CT), i.e., time necessary to stop the blood flow, and is a measure of platelet hemostasis capacity. Laboratory investigations during substitution therapy revealed no shortening of closure times with both COL/EPI and COL/ADP cartridges despite normalization of plasma vWF Ag, vWF Rco, and FVIIIc levels. These observations suggest that intraplatelet vWF, which is totally absent in type-III von Willebrand disease, plays an important function in the adhesion of platelets to the collagen-coated membrane of the PFA-100 system, simulating an injured vessel wall. Consequently, we conclude that the PFA-100 may not be suitable for monitoring the therapeutic efficacy of von Willebrand concentrate in type-III von Willebrand patients during substitution therapy. PMID- 10525830 TI - Phase-II trial of idarubicin, fludarabine, cytosine arabinoside, and filgrastim (Ida-FLAG) for treatment of refractory, relapsed, and secondary AML. AB - The current phase-II trial was initiated to assess the efficacy and toxicity of the Ida-FLAG regimen in patients with poor-risk acute myeloid leukemia (AML). Three subgroups of patients with AML were eligible for the study: (a) refractory, (b) first relapse, or (c) secondary AML (i.e., signs of trilineage myelodysplasia at diagnosis or the history of a myelodysplasia or myeloproliferative disorder). Fifty-seven fully evaluable patients were included in the study. Twenty patients received a second course of Ida-FLAG. Complete remission was achieved by 1/14 patients with refractory AML, 12/15 patients with relapsed AML, and 17/28 patients with secondary AML. The median duration of ANC <1000/microl was 17 days (10-36); of platelets <30,000/microl 23 days (9-65); of days with fever >38.0 degrees C 6 days (1-33). Thirteen patients (22.8%) died within 42 days of severe infection or hemorrhage. Overall survival at 20 weeks in the subgroups was 24% for patients with refractory, 78% for patients with relapsed, and 55% for patients with secondary AML. The toxicity of the first cycle of Ida-FLAG is moderate. The feasibility and subjective tolerance of the Ida-FLAG regimen are acceptable. There is no evidence for an increase of atypical infections. The efficacy for patients with secondary AML and especially those with first relapse of AML is good, with a high rate of complete remissions. Remission duration seems to be short. Therefore, an intensified post-remission therapy seems necessary. PMID- 10525832 TI - A dispermic chimerism in a 2-year-old Caucasian boy. AB - Detection of two different cell populations in a child is a rare event. The following case of a dispermic chimera was diagnosed before surgery due to problems in blood group determination. A 2-year-old phenotypically male child was admitted for correction of a penoscrotal hypospadia and unilateral cryptorchism. During presurgical laboratory investigation, difficulties in blood group determination occurred. Blood group typing was performed by the DiaMed-ID Micro Typing System and by FACS. Additionally, cytogenetic analysis of lymphocytes and analysis of DNA polymorphisms in different tissues were performed. Two populations of red blood cells were detected, O cells accounting for 75% and B cells for 25%. Analysis of DNA-PCR polymorphisms in lymphocytes, nails, and in cells of the oral mucous membrane demonstrated a chimerism, with two alleles inherited from the father and one from the mother. A cytogenetic analysis of cultured lymphocytes showed a mosaic 46, XY/46,XX. Surgery revealed a prostatic utricle grade III, also called pseudovagina; genitography confirmed a vagina. Bilateral gonad biopsy showed a testis on one side and an ovary on the other. This case of chimerism represents a true hermaphroditism that most probably developed by double fertilization of one or more egg nuclei by two sperms. PMID- 10525833 TI - Delayed and long-lasting complete response to fludarabine in two patients with B cell chronic lymphocytic leukemia. PMID- 10525834 TI - Bioconversion of progesterone by the activated immobilized conidia of Aspergillus ochraceus TS. AB - Progesterone was transformed to its 11alpha-hydroxy derivative (100% e.e) by the activated immobilized conidia of Aspergillus ochraceus TS. The immobilized preparation retained 79% of free conidial activity. The immobilized conidia, activated by nutrients, exhibited an increase in 11alpha-hydroxylation, and it was free of the side product 6beta, 11alpha-dihydroxy progesterone. The half life and turnover of immobilized and activated immobilized conidia were 14 and 12 days and 187 and 416 micromoles of the product/g of conidia respectively. The pH and temperature profiles of the free conidia remained unaltered after immobilization and activation. Some germination of conidia inside the matrix owing to incubation with nutrients was detected by scanning electron microscopy. PMID- 10525836 TI - Isolation and characterization of promoter regions from Streptococcus gordonii CH1. AB - We aimed to identify transcription signal sequences from Streptococcus gordonii strain CH1 by random chromosomal cloning. Five genomic fragments from a Sau3A digest, which constitutively activated transcription of a promoterless spectinomycin resistance gene in this strain, were isolated and characterized. Additionally, one promoter fragment was isolated that was specifically activated under iron-limiting conditions. A sequence motif with similarity to the consensus for Fur-binding regulatory DNA sequences (Fur box) in Escherichia coli was detected within the putative promoter region. The open reading frame downstream of this region possibly encodes a transmembrane protein involved in iron uptake. PMID- 10525835 TI - Inhibition of adherence of Actinobacillus pleuropneumoniae to porcine respiratory tract cells by monoclonal antibodies directed against LPS and partial characterization of the LPS receptors. AB - Actinobacillus pleuropneumoniae is the causative agent of porcine fibrinohemorrhagic necrotizing pleuropneumonia. We have previously identified the lipopolysaccharides (LPS) as the major adhesin of A. pleuropneumoniae involved in adherence to porcine respiratory tract cells. In the present study, adherence of A. pleuropneumoniae to porcine tracheal frozen sections was inhibited by homologous monovalent Fab fragments produced from monoclonal antibodies 5.1 G8F10 and 102-G02 directed, respectively, against the A. pleuropneumoniae serotype 1 or serotype 2 O-antigens. These results confirm the important role played by LPS in adherence of A. pleuropneumoniae and suggest that these adhesins might represent good vaccine candidates. We also investigated the presence of A. pleuropneumoniae receptors in tracheal cell preparations from piglets of four different breeds. Using Far-Western binding assays, we identified proteins recognized by whole cells of A. pleuropneumoniae reference strains for serotype 1 and 2, and local isolates belonging to the same serotypes, and also recognized by extracted LPS from both reference strains. We confirmed the proteinaceous nature of these LPS binding molecules by their staining with Coomassie brilliant blue, sensitivity to proteinase K digestion, resistance to sodium m-periodate oxidation, and their inability to stain with glycoprotein-specific reagents. Four low-molecular-mass bands (14-17 kDa) seemed to correspond to histones. We also identified proteins at Mr 38,500 that could represent putative receptors for A. pleuropneumoniae LPS in swine respiratory tract cells. PMID- 10525837 TI - Binding of selected extracellular matrix proteins to enterococci and Streptococcus bovis of animal origin. AB - Thirty-three enterococcal strains and 10 Streptococcus bovis strains were investigated for their protein-binding cell surface components. Seven extracellular matrix (ECM) proteins were immobilized on Difco latex beads to detect these components on the surface of all enterococcal strains and eight non autoaggregating S. bovis strains by a particle agglutination assay (PAA). Twenty three selected strains were also examined in microtiter plate assays. According to the absorbance readings (A(570nm)), 11 strains were classified as nonadherent (A(570nm) < 0.1), 10 strains as weakly adherent (0.1 < A(570nm) > 0.3), and 2 strains as strongly adherent (A(570nm) > 0.3) in these assays. A direct correlation was found between the values obtained in PAA and A(570nm) readings of microtiter plate assays. Binding of (125)I-labeled bovine lactoferrin to enterococci and streptococci was in the range of 6%-30% and of (125)I-labeled human vitronectin in the range of 9%-33% to streptococci. The binding of(125)I labeled ECM proteins to selected strains was much more effectively inhibited by sulfated carbohydrates than by non-sulfated hyaluronic acid, indicating the importance of the sulfate groups of these inhibitors. An inhibition effect of heparin on bLf binding to four selected strains was higher in comparison with fucoidan in the microtiter plates. Thirty-five out of 44 strains had agglutinated rabbit erythrocytes. However, these strains showed no ability to agglutinate bovine or sheep erythrocytes. PMID- 10525838 TI - Regulation of expression of the nonhemolytic phospholipase C of Burkholderia cepacia. AB - Burkholderia cepacia is an opportunistic pathogen that causes serious pulmonary infections in cystic fibrosis patients. We have purified and partially characterized one potential virulence factor for the organism-a nonhemolytic phospholipase C-and we studied the effect of iron restriction and choline and phosphate concentrations on the expression of phospholipase C. Iron limitation did not affect expression, the effect of choline was variable, and high phosphate concentrations repressed expression. Experiments with heat-treated spent culture supernatants suggested that autoinducers affected the expression of the phospholipase and two other potential virulence factors, a protease and a lipase. We screened 26 B. cepacia isolates for autoinducer activity: 11 induced violacein production in the autoinducer-deficient mutant Chromobacterium violaceum CV026. Spent supernatants from two strains, one that was positive in the C. violaceum assay and one that was negative, were tested for inducing early expression of phospholipase C, protease, and lipase in homologous and heterologous cultures. Expression of all three enzymes was increased or induced at an earlier stage in the growth curve in every case, suggesting not only that autoinducers were involved in the regulation of the expression of these enzymes, but also that the autoinducers were of two different classes. PMID- 10525839 TI - Expression of a new cold shock protein of 21.5 kDa and of the major cold shock protein by Streptococcus thermophilus after cold shock. AB - Streptococcus thermophilus is widely used in food fermentations; it commonly suffers diverse stress challenges during manufacturing. This study investigated the cold shock response of S. thermophilus when the cell culture temperature shifted from 42 degrees C to 15 degrees C or 20 degrees C. The growth of cells was affected more drastically after cold shock at 15 degrees C than at 20 degrees C. The generation time was increased by a factor of 19 when the temperature was lowered from 42 degrees to 20 degrees C, and by a factor of 72 after a cold shock at 15 degrees C. The two-dimensional electrophoretic protein patterns of S. thermophilus under cold shock conditions were compared with the reference protein pattern when cells were grown at optimal temperature. Two proteins of 21.5 and 7.5 kDa synthesized in response to cold shock were characterized. N-terminal sequencing and sequence homology searches have shown that the 7.5-kDa protein belonged to the family of the major cold shock proteins, while no homology was found for the new cold shock protein of 21.5 kDa. PMID- 10525840 TI - Cloning of Schizosaccharomyces pombe bio2 by heterologous complementation of a Saccharomyces cerevisiae mutant. AB - A Saccharomyces cerevisiae mutant affected in the last step of the biotin biosynthesis pathway was isolated by using a transposon mutagenesis method. The gene BIO2, encoding a biotin synthase, is shown to be interrupted in this mutant. Heterologous complementation experiment allowed the cloning and the characterization of a novel bio gene: bio2, encoding biotin synthase from Schizosaccharomyces pombe. PMID- 10525841 TI - Sequencing and expression of a beta-mannanase gene from the extreme thermophile Dictyoglomus thermophilum Rt46B.1, and characteristics of the recombinant enzyme. AB - A beta-mannanase gene (manA) was isolated from the extremely thermophilic bacterium Dictyoglomus thermophilum Rt46B.1. ManA is a single-domain enzyme related to one group of beta-mannanases (glycosyl hydrolase family 26). The manA gene was expressed in the heat-inducible vector pJLA602 and the expression product, ManA, purified to homogeneity. The recombinant ManA is a monomeric enzyme with a molecular mass of 40 kDa and an optimal temperature and pH for activity of 80 degrees C and 5.0. In the absence of substrate, the enzyme showed no loss of activity at 80 degrees C over 16 h, while at 90 degrees C the enzyme had a half-life of 5.4 min. Hydrolysis of the galactomannan locust bean gum (LBG) by purified ManA released mainly mannose, mannobiose, and mannotriose, confirming that ManA is an endo-acting beta-mannanase. Sequence comparisons with related beta-mannanases has allowed the design of consensus PCR primers for the identification and isolation of related genes. PMID- 10525842 TI - Characterization of Astragalus sinicus rhizobia by restriction fragment length polymorphism analysis of chromosomal and nodulation genes regions. AB - Two hundred and four isolates of rhizobia were sampled from root nodules of Astragalus sinicus grown in rice fields of six southern provinces of China. Genotypic diversity was determined by Southern hybridization using nodDBC genes as a probe, restriction fragment length polymorphism (RFLP) analysis of PCR amplified 16S-23S rDNA intergenic spacers (IGS), and plasmid profile. Our results show that rhizobia associated with A. sinicus were very diverse, and 10 genotypes were resolved within the previously identified dominant 16S rDNA type. Diversity levels varied greatly between different geographical locations. The same nod gene genotypes were harbored by distinct chromosomal types, suggesting that lateral plasmid transfer occurred during the evolution process. PMID- 10525843 TI - News & notes: diversity of superoxide-dismutases among clinical and soil isolates of Streptomyces species. AB - Comparison of the nature, activity, and cellular localization of superoxide dismutases (SOD) from soil and clinical isolates of Streptomyces species was investigated to identify possible factors that could account for the pathological role of the strains isolated from human lesions. Results showed that all of the studied strains possessed a cytoplasmic Ni-SOD. This particular SOD, found in isolates from patients, could be a new taxonomic criterion to identify Streptomyces species with greater precision. A second minor SOD, assimilated to an Fe/Zn-SOD, was detected in some strains, but no relationship was established between the presence of this enzyme and the clinical origin of the strains. PMID- 10525844 TI - Transcriptional and structural study of a region of two convergent overlapping yeast genes. AB - The exceptionally close packing of many yeast genes and other chromosomal elements raises the question of how those elements are functionally insulated. All published work shows that natural insulators are very effective, but transcriptional interference (TI) occurs if they are mutated or if their natural context is altered. Mechanisms to avoid TI are poorly understood, but are thought to involve an interplay of cis sequences and trans factors in a chromatin context. We have studied the case of two convergent closely packed ORFs (56 bp of separation) in chromosome IX of Saccharomyces cerevisiae. mRNAs from POT1 and YIL161w overlap by up to 115 nt. Convergent transcription causes a small but noticeable negative effect on the level of POT1 mRNA and nucleosome displacement in the intergenic region. This suggests for the first time that some TI could occur in convergently transcribed yeast genes, even in a natural chromosomal context. PMID- 10525845 TI - Syndrome of the trephined: theory and facts. AB - The pathophysiology behind "the syndrome of the trephined" has been under investigation over the past 50 years. Research related to barometric pressure, cellular metabolism, cerebrospinal fluid (CSF) dynamics, and the vasculature have attempted to decipher the mechanism of disease. These subjects are discussed in five papers along with specific topics related to the syndrome. The symptoms experienced after craniectomy, the resolution of symptoms with cranioplasty as well as CSF, cerebral blood flow, and metabolic studies are presented, respectively, with a review of the theories. PMID- 10525846 TI - Intraoperative use of magnetic resonance imaging: neurosurgical applications and technical limitations. AB - In this manuscript, the authors review current developments in intraoperative magnetic resonance imaging (MRI) and analyze the feasibility of obtaining intraoperative MRI images, current available instrumentation, and the advantages and disadvantages of performing a neurosurgical procedure using interactive or real time scans and neuronavigation. PMID- 10525847 TI - Perforating and leptomeningeal branches of the anterior communicating artery: an anatomical review. AB - The morphological variability, diameter, and length of the anterior communicating artery (ACoA) are important factors in clinical and surgical decisions. This artery presents branches that supply the optic nerves and chiasm, the lamina terminalis, the hypothalamus, and the subcallosal region. The ACoA has the most frequent incidence of saccular aneurysms in the anterior portion of the circle of Willis. Lesions to the ACoA's branches may be related to neuropsychological sequelae such as amnesia, confabulation and personality changes, besides other basal ganglia syndromes. In this paper, anatomical studies of the ACoA and its branches are reviewed and the results of an anatomical study carried out in our laboratory presented. PMID- 10525848 TI - High-dose chemotherapy with autologous hematopoietic stem-cell rescue for patients with malignant brain tumors. AB - This article reviews recent reports on high-dose chemotherapy combined with autologous stem-cell rescue for patients with newly diagnosed or recurrent malignant brain tumors. Children with newly diagnosed malignant central nervous system (CNS) tumors are included - in these patients it was desirable to avoid radiotherapy. The drugs used were thiotepa, etoposide, cyclophosphamide, busulfan, melphalan, and carboplatin. At a toxic death rate of 8-13%, their toxicity was considered acceptable. However, in patients receiving high-dose chemotherapy, neurologic complications were not uncommon. Therefore, the effects of this drug therapy on the post-treatment cognitive function of long-term survivors must be examined prospectively. Regarding the types of tumor that respond to drug therapy, medulloblastoma and cerebral primitive neuroectodermal tumor (PNET) appear to be good candidates, as are malignant astrocytic tumors in the cerebral hemisphere of children younger than 3 years. On the other hand, brain stem gliomas do not appear to respond well to high-dose chemotherapy. Radiotherapy can be avoided in a significant proportion of young children with malignant brain tumors. Larger patient series need to be studied to determine which patients are most likely to benefit from high-dose chemotherapy and to pinpoint the optimal time for treatment intervention. PMID- 10525849 TI - Oxyhemoglobin as the principal cause of cerebral vasospasm: a holistic view of its actions. AB - While oxyhemoglobin (oxyHb) is deemed to be the principal cause of cerebral vasospasm following subarachnoid hemorrhage, the mechanism(s) whereby it leads to vasospasm is by no means clear. Of importance is the fact that prolonged contraction of arterial smooth muscle is not the sole feature of cerebral vasospasm, particularly in humans. Vasospasm is also associated with the occurrence of organic changes in the arterial wall as well as the derangement of cerebral microcirculation. These additional features may play a pivotal role when vasospasm in the proximal arteries incurs delayed ischemic neurological deficits and cerebral infarction. The question then arises as to whether or not all the features of vasospasm are attributable to the actions of oxyHb. In this regard, owing to the recent advances in vascular physiology, it has become clear that the cerebral vasculature should be regarded as an organ, not a mere conduit, in which all intracellular mechanisms are functionally integrated for the maintenance and regulation of cerebral blood flow (CBF). In the sense that the arterial function is not simply a sum of the individual cellular functions, it may be described as "holistic". According to extant literature, oxyHb has multifarious actions that can be divided into the following three categories: (1) scavenging of nitric oxide (NO), (2) generation of reactive oxygen species (ROS), (3) activation of the tyrosine kinase/mitogen-activated kinase (TK/MAPK) pathway. Based on such knowledge, the present review aims at a speculative synthesis in terms of how oxyHb pertains to the occurrence of vasospasm, in which the highly integrated, holistic mechanisms within the cerebral artery are perturbed for a prolonged period. PMID- 10525850 TI - Solitary fibrous tumor in neurosurgical practice. AB - Solitary fibrous tumors most often affect the pleura, but examples are increasingly being reported in a wide variety of sites including the central nervous system. This tumor shows characteristic expression of CD34, which facilitates histopathologic differentiation of this lesion from other more common and better recognized spindle-cell tumors such as fibrous meningioma, hemangiopericytoma, or nerve sheath tumors. In this paper, we review current information on cranial and paracranial solitary fibrous tumors and emphasize the need for clinical recognition of this lesion as a distinct entity. PMID- 10525853 TI - Publications scanned for pertinent articles. PMID- 10525852 TI - Papers reviewed in this issue. PMID- 10525854 TI - Interventional MR imaging: percutaneous abdominal and skeletal biopsies and drainages of the abdomen. AB - Since the introduction of open magnets and short-bore closed magnets, and the availability of fast imaging sequences, MR imaging has become a tool for guidance and control of percutaneous nonvascular and vascular procedures. In general, percutaneous biopsies or drainages require no specific hardware or software modifications. For lesion localization and control of the needle track, simple fast imaging sequences such as fast T1-weighted gradient-echo techniques or fast single-shot T2-weighted spin-echo sequences are sufficient and can be applied depending on the best tissue-to-lesion contrast. Typical scan times range from 1 to 3 s. Different types of biopsy needles are commercially available, allowing sampling of sufficient amounts of tissue. For drainage procedures non ferromagnetic materials, such as nitinol wires, should be preferred to minimize image distortion by metallic artifacts. Indications for percutaneous biopsies include subdiaphragmatic liver lesions, lesions poorly visible on ultrasound or contrast-enhanced computed tomography, and lesions of the bone marrow characterized by an unspecific bone marrow edema. For percutaneous drainages, subdiaphragmatic lesions appear to be a good indication. With some experience the procedure time is not longer than that under CT or US guidance. PMID- 10525855 TI - Interventional MR: interstitial therapy. AB - The rationale and results for interstitial therapies via interventional MRI in the treatment of tumors in various regions are presented. Different interstitial treatment techniques are presented based on varying technologies both for tumor ablation and treatment monitoring. Data are presented based on 335 patients, 29 84 years of age (mean age 59 years, 196 men and 139 women) with a total of 932 liver tumors, 16 head and neck tumors and 14 abdominal recurrent pelvic and lymphatic tumors. All lesions had been treated with MR-guided laser-induced interstitial thermotherapy (LITT) via 2516 laser applications and 1856 cannulations. Data in the literature are extremely varying depending on author experience, treatment technique, and the included patient material. In our patient material we were able to achieve a local tumor control of 96.7% depending on the size of the tumorous lesion, the topographical relationship, and the applied laser parameters. The overall cumulative survival rate of patients with liver metastases was 45.74 months (median 40.97 months, 95 % confidence interval 31.42-50.52). The cumulative survival rate of the patient group with hepatic metastases of colorectal carcinoma was 42.71 months (median 39.33 months, 95% confidence interval 33.26-45.37). In patients with head and neck tumors a relevant reduction in clinically relevant symptoms such as pain, swallowing disorders, or nervous compression was achieved in 11 of 15 patients treated with LITT. In 14 soft tissue tumors, such as pelvic tumor recurrence and lymph node metastases, a local tumor control was obtained in 68% of lesions. Interstitial therapies under interventional MRI guidance, such as LITT, results in a high local tumor control with an improved survival rate. PMID- 10525856 TI - Interventional MR: vascular applications. AB - Three strategies for visualisation of MR-dedicated guidewires and catheters have been proposed, namely active tracking, the technique of locally induced field inhomogeneity and passive susceptibility-based tracking. In this article the pros and cons of these techniques are discussed, including the development of MR dedicated guidewires and catheters, scan techniques, post-processing tools, and display facilities for MR tracking. Finally, some of the results obtained with MR tracking are discussed. PMID- 10525857 TI - Abdominal MR: liver and pancreas. AB - Following the introduction of rapid, high-quality scan techniques and the development of new, tissue-specific contrast agents, the applications of MRI for abdominal imaging are experiencing unprecedented growth. This article examines the current status of liver and pancreatic MRI, highlighting technical and methodological approach, use of contrast agents, and main clinical applications. The MRI technique appears to be the ideal diagnostic tool for detection and characterization of benign and malignant liver neoplasms, and for evaluating tumor response after nonsurgical treatments. Dynamic imaging after bolus injection of a gadolinium chelate is currently a fundamental component of an MRI examination of the liver in many instances. Optimal dynamic scanning depends on the use of a multisection spoiled gradient-echo technique that allows one to image the entire region of interest during a single suspended respiration. Images are obtained during four phases relative to the injection of the contrast agent: precontrast, arterial (pre-sinusoidal), portal (sinusoidal), and delayed (extracellular) phase. Liver-specific contrast agents, including hepatobiliary agents and reticuloendothelial system-targeted iron oxide particles, however, may offer advantages over gadolinium chelates in some clinical settings. Computed tomography is still preferred to MRI for imaging the pancreas. However, state-of the-art MRI may currently be at least as accurate as spiral CT for depiction of inflammatory and neoplastic pancreatic diseases. Moreover, MRI has the advantage of allowing simultaneous investigation of the biliary tree, owing to cholangiopancreatography techniques. Hence, a comprehensive assessment of most pancreatic diseases can be achieved with a single examination. PMID- 10525858 TI - MRI of the biliary and pancreatic ducts. AB - Magnetic resonance Cholangiopancreatography (MRCP) is a non-invasive imaging technique able to provide projectional images of the bile ducts. Different sequences, using both breath-hold and non-breath-hold acquisition techniques, have been employed in order to obtain MRCP images. The authors discuss technical aspects, considering both three-dimensional non-breath-hold techniques and two dimensional breath-hold, multi-slice and thick slab sequences. Clinical applications of MRCP are evaluated, presenting data from both the literature and personal experience. The main indication for MRCP study is represented by the evaluation of common bile duct obstruction, with the aim of assessing the presence of the obstruction (accuracy 85-100%) and, subsequently, its level (accuracy 91-100%) and its cause. The utility of associating conventional MR images to MRCP in malignant strictures, in order to characterize and stage the malignant lesion, is also discussed. Finally, data are presented regarding the indications and the utility of MR-pancreatography in the evaluation of patients with pancreatic duct anomalies and chronic pancreatitis. PMID- 10525859 TI - MRI of the small and large bowel. AB - Driven by the improvements in gradient technology, breathhold T1- and T2-weighted imaging of the abdominal structures has become possible. These techniques allow exploitation of the advantages inherent to the MR imaging experiment: unsurpassed soft tissue contrast and multiplanar imaging capabilities. Magnetic resonance imaging of the small and large bowel has thus moved from a hypothetical possibility to a practical reality. This manuscript describes some of the underlying fast imaging techniques for display of the small and large bowel. Furthermore, it discusses the plethora of available oral and rectal contrast agents. Finally, clinical indications for MR of the small and large bowel as well as the rectum are described in light of the available literature. Advantages and disadvantages relative to computed tomography and other imaging techniques are discussed. PMID- 10525860 TI - Focal nodular hyperplasia of the liver on ferumoxides-enhanced MR imaging: features on conventional spin-echo, fast spin-echo and gradient-echo pulse sequences. AB - The aim of this study was to assess the efficacy of a superparamagnetic iron oxide, ferumoxides, in the detection and characterization of focal nodular hyperplasia (FNH) on MR conventional spin-echo (SE), fast spin-echo (FSE) and gradient-echo (GRE) images. Fourteen adults with 27 FNHs were evaluated at 1.5 T before and after injection of ferumoxides. T1-weighted and T2-weighted SE, T2 weighted FSE and T2(*)-weighted GRE sequences were used and analysed qualitatively and quantitatively. One hundred percent of FNHs showed a significant postcontrast decrease in signal intensity on T2- and T2*-weighted images. Heavily T2-weighted SE images showed the maximum decrease in FNH signal to-noise ratio (S/N). Postcontrast GRE T2(*)-weighted images improved the detection of the central scar and the delineation of FNHs and demonstrated the best lesion-to-liver contrast-to-noise ratio (C/N). Postcontrast T1-weighted SE images showed the least lesion-to-liver C/N. Ferumoxides-enhanced MR imaging can help detect and characterize FNH. Conventional pre- and postcontrast T2-weighted SE images and postcontrast GRE T2*-weighted images should be used preferentially. PMID- 10525861 TI - Is capsular retraction a specific CT sign of malignant liver tumor? AB - The aim of this study was to assess if a liver capsular retraction is a specific CT sign in malignant hepatic tumors. The authors reviewed retrospectively 320 hepatic CT scans obtained in 300 patients during a 3-year period. These patients presented with benign (n = 64) or malignant (n = 236) hepatic tumors. In 7 patients we found retraction of the capsule surrounding the tumor. All these tumors were histologically proven as malignant lesions: 4 metastases (none being chemically treated), 2 peripheral cholangiocarcinomas, and 1 epithelioid hemangioendothelioma. The prevalence of this sign was 2.18% (7 of 320) in this series. This capsular retraction pattern has never been found in hepato-cellular carcinomas (no fibrolamellar in this series) and benign lesions. Liver capsular retraction is an uncommon but specific (100%) sign in malignant hepatic tumors; however, a larger and prospective series is needed. PMID- 10525862 TI - Esophageal varices before and after endoscopic variceal ligation: evaluation using helical CT. AB - The purpose of this study was to demonstrate the utility of helical CT in assessing the therapeutic effects of endoscopic variceal ligation (EVL). Twenty four patients with esophageal varices were examined. Helical scanning was initiated 60 s after intravenous injection (Iopamidol 300 mgI/ml, total 120 ml, 3 ml/s) was started. Esophageal varices were clearly depicted as high-density areas. Multiplanar reformation and 3D images demonstrated collateral circulation three-dimensionally. After EVL, mucosal high-density areas had diminished markedly, but collateral veins around the esophagus, and gastro- and/or spleno renal shunts, were unchanged in all patients. Of 21 patients with collateral circulation, esophageal varices recurred endoscopically in 6 patients within 12 months. In 3 patients without collateral circulation, esophageal varices did not recur within 12 months. From these findings, we conclude that helical CT is a useful method for assessing the therapeutic effects of EVL. PMID- 10525863 TI - Clear cell sarcoma of the abdominal wall with peritoneal sarcomatosis: CT features. AB - Clear cell sarcoma, also called malignant melanoma of soft parts, is an uncommon neoplasm that involves tendons or aponeuroses of the lower extremity. The CT features of a clear cell sarcoma arising from the abdominal wall with later peritoneal dissemination are described. Peritoneal sarcomatosis from soft tissue sarcomas is a very rare condition previously unreported in the radiologic literature. Metastases to peritoneal surfaces must therefore be considered a possible site for systemic dissemination of soft tissue sarcomas. PMID- 10525864 TI - Diagnosis and treatment of expanding haematoma of the lateral abdominal wall after blunt abdominal trauma. AB - We report a rare case of an expanding post-traumatic lateral abdominal wall haematoma. A superselective arteriogram of the deep circumflex iliac artery showed extravasation from the ascending branch, urging emergency therapy. Microcoil and Gelfoam embolisation was successfully performed. Haematomas of the abdominal wall can be divided in the common rectus sheath haematomas and the rare haematomas of the lateral abdominal wall. Differentiating both entities is essential, since there is a strong difference in their vascular supply. The typical vascular supply of the lateral abdominal wall is discussed, with emphasis on the ascending branch of the deep circumflex iliac artery. PMID- 10525865 TI - Computed tomography of the thorax in HIV disease. AB - A wide variety of thoracic disorders can arise in patients infected with the human immunodeficiency virus (HIV), although recent developments in the therapeutic management of AIDS patients has resulted in a changing pattern of chest disease. The use of CT in the diagnosis and management of these thoracic manifestations is discussed along with the CT appearances of the various infectious and non-infectious complications of the acquired immune deficiency syndrome (AIDS) which are commonly encountered in clinical practice. PMID- 10525866 TI - Anomalous origin of the left coronary artery arising from the pulmonary trunk: report of an adult case with long-term follow-up after surgery. AB - An anomalous origin of the left coronary artery arising from the pulmonary artery is a congenital malformation rarely described in adults. We report the case of a 65-year-old patient with this anomaly. Clinical presentation, imaging identification (coronary angiogram, MRI and electron-beam CT), surgical treatment and angiographic long-term follow-up are described. PMID- 10525867 TI - Evolution of pulmonary perfusion defects demonstrated with contrast-enhanced dynamic MR perfusion imaging. AB - Pulmonary perfusion defects can be demonstrated with contrast-enhanced dynamic MR perfusion imaging. We present the case of a patient with a pulmonary artery sarcoma who presented with a post-operative pulmonary embolus and was followed in the post-operative period with dynamic contrast-enhanced MR perfusion imaging. This technique allows rapid imaging of the first passage of contrast material through the lung after bolus injection in a peripheral vein. To our knowledge, this case report is the first to describe the use of this MR technique in showing the evolution of peripheral pulmonary perfusion defects associated with pulmonary emboli. PMID- 10525868 TI - Altered distribution of Pneumocystis carinii pneumonia during radiation therapy. AB - The radiographic findings of Pneumocystis carinii pneumonia (PCP) are various. The typical findings are diffuse, bilateral, symmetric, finely granular, or reticular infiltrates. In patients taking aerosol pentamidine, atypical findings may be the first manifestation. One interesting radiologic finding of PCP is that the pneumonia may spare the irradiated lung. We report PCP developed in a patient undergoing irradiation for lung cancer. High-resolution CT revealed diffuse, bilateral, and symmetric ground-glass opacities with septal thickening in both lungs; however, the radiation port was spared and appeared as the "photographic negative of post-radiation pneumonia." The distribution of the pneumonic infiltrates was altered by radiotherapy. PMID- 10525869 TI - Opacification of the urinary tract in portal venous spiral CT without delayed scans. AB - In portal venous spiral CT there is no visible renal contrast excretion within the usual period of scanning. To opacify collecting systems additional delayed scanning is required. We administered an extra pre-dose of contrast medium before the main portal venous bolus in order to opacify the urinary tract and studied its effects on liver attenuation. In 32 patients examined first by non-contrast spiral CT 20 ml of a non-ionic IV CM were injected. Five minutes later, orientating cuts in the liver and along the urinary tract were obtained. Immediately thereafter, a 120-ml bolus was administered at 3 ml/s for portal venous phase helical CT (60-s delay craniocaudad). The quality of renal excretion was graded visually (excellent, fair, poor, none). Hepatic attenuation measurements were performed at comparable regions of interest. In all patients 20 ml CM opacified the renal pelvis after 5 min. Depiction of the ureters was excellent in 14, fair in 11 and poor or none in 7 cases. There was little effect on mean hepatic attenuation by the 20-ml pre-bolus after 5 min: mean enhancement 2.3 HU (range -0.6 to 7.8 HU). Mean hepatic enhancement after the 120-ml portal venous bolus ranged between 23.6 and 74.1 HU (mean 51.5 HU). When opacification of the urinary tract is necessary, pre-administration of a 20-ml bolus 5 min before portal venous scanning may save an extra delayed spiral. The effects on hepatic enhancement are negligible. PMID- 10525870 TI - Utility of coaxial technique for renal angioplasty in patients with a difficult to-cross stenosis. AB - The aim of this study was to evaluate the feasibility of coaxial approach in difficult-to-cross lesions in patients with failed percutaneous transluminal renal angioplasty by conventional over-the-wire exchange technique. Twelve stenoses in 10 patients (six women and four men; age range 19 +/- 7 years) with uncontrolled hypertension were treated by this method. The stenosis was caused by nonspecific aortoarteritis in 8 patients and fibromuscular dysplasia in 2 patients. It was ostial in seven and post-ostial in five vessels. Conventional exchange technique was unsuccessful in all of them. All procedures were done by femoral route. Technical success was seen in 11 (92%), without complication. The stenosis improved from 90 +/- 2.1% (range 80-100%) to 6 +/- 7% (range 0-20%), blood pressure decreased from 198 +/- 12.3 mm Hg (range 180-220 mm Hg)/130 +/- 6.7 mm Hg (range 120-140 mm Hg) to 119 +/- 5.7 mm Hg (range 110-130 mm Hg)/83 +/- 3.9 mm Hg (range 80-90 mm Hg), and number of drug treatments for hypertension fell from 3.6 +/- 0.52 (range 3-4) to 1 +/- 0.94 (range 0-3; p < 0.01). Percutaneous transluminal renal angioplasty resulted in "cure" in 3 patients and "improvement" in 7 patients. Follow-up period was 3-21 months (mean 6.4 months). No restenosis was detected. Coaxial approach is safe and effective in treating difficult-to-cross lesions in which renal angioplasty by conventional exchange technique is unsuccessful. PMID- 10525871 TI - Foreign-body granuloma of the kidney: CT, MR and pathologic correlation. AB - The differential diagnosis of renal masses containing fatty foci is limited to a small number of well-defined tumors, angiomyolipoma being the most frequent. In recent years clear cell carcinomas with intratumoral fatty foci have been reported, due to either entrapment of local fat or to regressive adipose metaplasia. Demonstration of focal calcifications is a valuable sign, being relatively common in carcinomas while rare in more benign lesions. We report a case of a foreign-body granuloma of the kidney, containing both calcifications and foci of fat. The value of this case, in our opinion, is that it demonstrates that detection of the previously mentioned features in a renal mass does not necessarily imply a presumptive diagnosis of renal cell carcinoma. PMID- 10525872 TI - Penile epithelioid sarcoma: MR imaging findings. AB - Magnetic resonance imaging findings of a 38-year-old man with epithelioid sarcoma of the penis is presented. It started as a firm, painless and slowly growing nodule at the base of his penis 6 months previously which caused pain radiating to the testis during coitus. It has been well known that sarcomas may well mimic reactive processes. Initial presentation of epithelioid sarcoma may provoke considerable diagnostic difficulty, and its differentiation from benign lesions, such as Peyronie's disease and chronic inflammation, may be a clinical problem. In our present report the MR findings are compared with those of the epithelioid sarcomas of various locations reported in the literature and differential diagnosis of the entity is discussed. To our knowledge, this is the first report regarding the MR findings of the epithelioid sarcoma of penis. PMID- 10525873 TI - Isolated penile metastasis from bladder carcinoma. AB - Metastases of the penis are uncommon, with only approximately 300 cases reported since 1870. In up to 70% of patients, the primary tumour is located in the urogenital tract. Furthermore, isolated metastases of the penis are exceptionally rare. We report a case of solitary squamous cell metastasis of the penis presenting with painful swelling initially thought to be inflammatory in origin. The CT and MR imaging findings are presented with a short review of the literature. PMID- 10525874 TI - Renal replacement lipomatosis: ultrasonography and computed tomography findings. AB - Replacement lipomatosis of the kidney is the result of severe atrophy or destruction of the renal parenchyma often caused by calculous disease with secondary marked proliferation of renal sinus, renal hilus, and perirenal fatty tissue. The diagnosis is difficult to establish with conventional radiographic methods. Although ultrasonography may show highly suggestive findings, computed tomography seems to be the most accurate method for demonstrating the distinctive features of replacement lipomatosis. Ultrasonographic and computed tomographic features in three cases of replacement lipomatosis of the kidney are reported. PMID- 10525875 TI - Contrast-media-induced nephrotoxicity: a consensus report. Contrast Media Safety Committee, European Society of Urogenital Radiology (ESUR). AB - The purpose of this study was, using consensus methodology, to document current understanding of contrast media nephrotoxicity (CMN) and to identify areas where there is disagreement or confusion. To draw up guidelines for avoiding CMN based on the current understanding of the condition established by the survey. One hundred sixty-four statements were mailed to 148 members of the European Society of Urogenital Radiology (ESUR) and to 48 experts in the field of CMN. They were asked about the definition, clinical features, predisposing factors and pathophysiology of CMN and about prophylactic measures. The importance of the statements was rated on a scale from 1 to 10 (1 least important, 10 most important). Fifty-three members (38%) and 23 experts (48%) responded. Both groups considered that an increase in serum creatinine that peaks within 3-4 days and a decrease in creatinine clearance are the most important (rating > 7) features of CMN. Enzymuria was not considered important (rating < 6). Pre-existing renal insufficiency, diabetic nephropathy, dehydration, congestive heart failure, concurrent administration of nephrotoxic drugs and the dose and type of contrast media were considered to be risk factors. Reduction in renal perfusion and damage to tubular cells were considered the main factors in the pathophysiology of CMN (rating > 6). Hydration and the use of low osmolar contrast media were thought to minimize the incidence of CMN (rating > 6). The majority of the responders (84.6% of members and 95.5% of experts) believe that the incidence of CMN in patients with normal renal function is less than 5%. Of the members, 62.5%, and 35.3% of experts, believe that the incidence of CMN is 20-30% in the presence of risk factors. There was disagreement about the definition of CMN, the threshold dose of contrast media above which renal complications may develop, the safe period between repeat injections, the relevance of contrast media renal retention shown on CT and whether contrast media have long-term effects on renal function. The survey showed good understanding of CMN among those who answered the questionnaires, although areas of disagreement remain which require further research. Simple guidelines are proposed. PMID- 10525876 TI - Use of single-slice thick slab phase-contrast angiography for the diagnosis of dural venous sinus thrombosis. AB - The aim of this study was to examine the reliability of single-slice phase contrast angiography (SSPCA) as a rapid technique for the investigation of suspected dural venous sinus occlusion. Images were obtained on 25 normal volunteers to document the accuracy of SSPCA in the demonstration of slow flow states. Normal volunteers were imaged using sagittal and coronal SSPCA (slice thickness 13 cm, matrix 256 x 256, TR 14 ms, TE 7 ms, flip angle 20 degrees, peak velocity encoding rate 30 cm/s). Sinus patency and flow rate were confirmed by measurement of flow in the superior sagittal and transverse sinuses using quantified single-slice phase difference images. Imaging was performed in 50 patients undergoing routine brain scans in order to determine the optimal slice orientation for clinical use. Twenty-one patients with suspected dural venous sinus thrombosis were also investigated with SSPCA and the diagnosis confirmed by one or more alternative imaging techniques. Imaging time was 29 s per acquisition and image quality was good in all cases. Variations in dural sinus patency and flow in normal volunteers were accurately predicted by SSPCA (kappa = 0.92). Use of a single angulated slice (130 mm thick, para-sagittal image angled 30 degrees towards coronal and 30 degrees towards transverse) provided sufficient separation of right- and left-sided venous structures to allow use of a single projection. The presence and extent of sinus occlusions in 14 patients and the absence of thrombosis in 7 were accurately identified by SSPCA. Sensitivity and specificity in this limited study were both 100%. The SSPCA technique takes less than 30 s and provides a reliable and rapid technique for the diagnosis of dural venous sinus thrombosis. PMID- 10525877 TI - Wernicke encephalopathy: MR findings in two patients. AB - Wernicke encephalopathy is a serious neurologic disorder caused by vitamin-B1 or thiamine deficiency. In the literature the characteristic symmetric paraventricular lesions of Wernicke encephalopathy are hyperintense on T2 weighted sequences spin-echo (SE) and enhance on T1-weighted SE sequences after intravenous gadolinium administration in the acute phase. We present two patients in the acute phase of Wernicke encephalopathy with special reference to the MR imaging. One of our reported cases is special because of the MR demonstration of a hemorrhagic focus in the caput of the right nucleus caudatus. The other case demonstrates no enhancement on SE T1-weighted sequences after intravenous gadolinium administration. PMID- 10525878 TI - Unusual chondrosarcoma of the larynx: CT findings. AB - A case of a large low-grade mixed clear-cell and conventional chondrosarcoma of the larynx is reported involving the paraglottic space, the cricoid and thyroid cartilage and characterized by an unusual long clinical course over 22 years, although multiple recurrences occurred without developing metastases. Computed tomography suggests diagnosis by detecting calcifications and adequately demonstrates the extension of the tumor. Innersurface and surface-rendering images document the airway stenosis in all directions. The unusual feature of this case consists in the peculiar histopathological differentiation of the observed chondrosarcoma showing a large clear-cell component. PMID- 10525879 TI - Radiological findings in acute adult epiglottitis. AB - Acute epiglottitis is a rare but life-threatening disease that commonly occurs in children, and also rarely in adults. The symptoms may be mild and non-specific before a rapid onset of airway obstruction occurs. Early diagnosis is essential, as delayed treatment is associated with a high rate of complications including death. We present the clinical and radiological findings of this unusual condition in an adult. PMID- 10525880 TI - Phalangeal US velocity discriminates between normal and vertebrally fractured subjects. AB - The purpose of this study was to evaluate the diagnostic sensitivity of phalangeal bone ultrasound velocity of the hand in the diagnosis of osteoporosis and to compare this technique to bone mineral density (BMD) measurement at the lumbar spine assessed by dual X-ray absorptiometry (DXA) and quantitative computed tomography (QCT). We investigated US velocity at the distal metaphysis of the proximal phalanx and spinal BMD in 101 women. Fifty-nine were healthy (mean age 50 +/- 11.6 years) and 42 were osteoporotic (mean age 65 +/- 6.6 years) with documented vertebral fractures. In the healthy population the relation with age was, respectively, r = -0.73 (p < 0. 0001) for quantitative US (QUS), r = 0.74 (p < 0.0001) for QCT and r = -0.48 (p < 0.01) for DXA. Both US and DXA were correlated with QCT: r = 0.74 and r = 0.77 (p < 0.0001), respectively. Correlation of QUS and DXA was r = 0.56 (p < 0.0001). Phalangeal US velocity and spinal BMD (QCT and DXA) values discriminate healthy from osteoporotic women. Age adjusted logistic regression analysis of the data showed standardized odds ratios (OR) for vertebral fracture to be similar for US and DXA (OR = 1.8 and 1.5, respectively) and stronger for QCT (OR = 2.9). Phalangeal US velocity reflects age-related bone loss and differentiates between healthy and osteoporotic subjects. PMID- 10525881 TI - Utility of CT scan for the diagnosis of chest wall tuberculosis. AB - The objective of this study was to determine the utility of CT scan findings for the diagnosis of chest wall tuberculosis, excluding the spine. We reviewed 15 patients (13 Africans and 2 Indians) with chest wall tuberculosis, retrospectively. The radiologic examination consisted of a plain X-ray and a CT scan of the chest for each patient. The site of disease was the rib in 13 patients or the body of the sternum in 2 patients. One rib was involved in 11 patients, 2 contiguous ribs (one site) in 2 patients, and bilateral disease (two sites) was observed in the remaining patient. The 14 rib sites involved the posterior arc or costovertebral joint in 11 cases, the anterior arc in 2 cases, and the anterior and middle arc in 1 case. The CT scan findings were an abscess (n = 14) or a soft tissue mass (n = 2), osteolytic lesions (n = 13), periosteal reaction (n = 10), and sequestrum (n = 14). Bone sclerosis was observed only in 3 cases of rib involvement. The association of a soft tissue abscess, an osteolytic lesion, and sequestrum, especially in immigrants to France, suggests chest wall tuberculosis on CT scan. PMID- 10525882 TI - Symptomatic fibrous lunato-triquetral coalition. AB - In general, carpal coalitions are considered to be asymptomatic. Incomplete separated joints and associated changes similar to osteoarthritis and pseudoarthrosis are known as possible causes of wrist pain. We present the clinical history, plain-film, and MR imaging findings of two patients with symptomatic fibrous lunato-triquetral coalition. Conventional films disclosed a narrowed space between the lunate and triquetral bone with cysts and sclerosis similar to pseudoarthrosis. Magnetic resonance imaging showed bone marrow edema adjacent to the incomplete separated lunato-triquetral joint and Gd-DTPA enhancing fibrovascular tissue in the synovium and subarticular cysts, explaining the pain over the ulnar-sided wrist. Patients with congenital lunato-triquetral coalition may poorly tolerate stress loading or trauma, resulting in a symptomatic state similar to degenerative arthritis or pseudoarthrosis, which is demonstrated by enhanced MR imaging. PMID- 10525883 TI - Acetabular pneumatocyst containing air-fluid level. AB - The presence of intraosseous gas most commonly occurs in osteomyelitis, vacuum phenomenon, and postsurgery or posttraumatic states. Several cases of subchondral gas-filled lesions, called pneumatocysts, have also been described in the sacroiliac joint and clavicle, none of them with intralesional air-fluid level. These pneumatocysts are innocuous lesions of uncertain origin. We describe one case of acetabular pneumatocyst containing air-fluid level in a 62-year-old man with long-standing ankylosing spondylitis involving hip joint. To our knowledge, this is the first reported case of a pneumatocyst in an acetabular location containing air-fluid level. PMID- 10525884 TI - Cranial fasciitis in an adult: CT and MR imaging findings. AB - Cranial fasciitis is a rare bone lesion in childhood. We report the first case in an adult, with CT and MR imaging, and suggest some diagnostic keys. PMID- 10525885 TI - Plexiform schwannoma of the foot. AB - The present report describes a plexiform schwannoma involving the subcutis of the foot in an 8-year-old boy. Gross findings revealed thin fibrous septa in a multilobulated tumor that was partly separated into free body-like nodules in the subcutis. Preoperative CT and MRI failed to delineate this multinodular architecture or free bodies. This is a case presentation including the CT and MR findings associated with plexiform schwannoma. PMID- 10525886 TI - MR-guided percutaneous excisional and incisional biopsy of breast lesions. AB - The aim of this study was the realisation and clinical application of MR-guided vacuum biopsy for percutaneous excisional and incisional biopsy of enhancing breast lesions. A breast biopsy system and procedure have been developed which allow precise and safe access to breast lesions in any location and use of vacuum biopsy (VB) under MR guidance. Fifty-one patients with 55 MR-detected lesions were examined. Verification of these diagnoses included re-excision histology of all 14 malignancies and for benign lesions retrospective correlation of histology and imaging, assessment of complete or partial removal of the enhancing area directly after VB (40 of 40 lesions) and follow-up MRI (33 of 40 lesions), which in contrast to conventional needle biopsy can be used as proof of representative removal. Fifty-four of 55 procedures (including 15 lesions /=50% diameter stenosis. With DSMR, significantly more patients yielded very good (69%) or good (13%) image quality in comparison with dobutamine stress echocardiography (20% and 31%, p<0. 05). Moderate image quality occurred in 16% with MR and 41% with dobutamine stress echocardiography (p<0.05), 2% and 8% were non-diagnostic. With each technique 18 patients could not be examined (DSE: emphysema: 10, adipositas: 8, DSMR: claustrophobia: 11, adipositas: 6, contraindication: 1). Four patients did not reach target heart rate. In 107 patients, significant coronary artery disease was found. With DSMR sensitivity was 88.7% (dobutamine stress echocardiography: 74.3%; p<0.05) and specificity 85.7% (dobutamine stress echocardiography: 69.8%; p <0.05). This difference was most pronounced in the group with moderate echocardiographic image quality. High dose DSMR is superior to dobutamine stress echocardiography and can replace this technique especially in patients with moderate echocardiographic image quality. PMID- 10525924 TI - Aortic regurgitant flow by color Doppler measurement of the local velocity 7 mm above the leak orifice--Part 1: in vitro measurements. AB - AIMS: The flow convergence method enables the determination of flow across restrictive orifices. It was validated for planar orifice plates and atrioventricular valves. However, its quantitative application to aortic regurgitation is complicated due to the complex valve anatomy which distorts the converging flow field. An open angle formed by the leaflets causes relatively lower velocities in the region near the orifice, resulting in underestimation of flow. Confinement of the flow convergence region by the ascending aorta relatively increases the velocities at greater distance to the orifice and can cause overestimation of flow. We hypothesized that there is a region at intermediate distance to the orifice, where both these effects on the flow field are minimal, so that the local velocity there is only a function of flow. METHODS AND RESULTS: In a flow model, aortic regurgitation was simulated. The flow convergence was imaged by color Doppler. Different scanning directions were used (analogue to apical and parasternal approach). Velocity profiles across the flow convergence were read along the line from the scanhead to the orifice. At a distance of 7 mm to the orifice, a uniform value was found for the ratio of local velocity v(7 mm)/flow (=0.28 cm(-2)). Variations in size (3.5 to 7 mm), site (central versus lateral) and leaflet angle (planar versus inverted funnel) of the orifice and in the scanning approach had only a minimal effect on this value. CONCLUSION: The aortic regurgitant flow convergence is characterized by a relatively uniform V (7 mm)/Q. Independent of variations in the anatomy and the scanning approach, this value directly reflects flow. PMID- 10525925 TI - [Clinical utility of contrast echocardiography for quantification of left ventricular volumes and function in patients with suboptimal echocardiograms]. AB - In the present study, we investigated whether the intravenous injection of air filled albumin microspheres (Infoson) as a contrast medium improves the echocardiographic quantification of left ventricular enddiastolic and endsystolic volumes, stroke volume, ejection fraction, and regional wall motion in patients with suboptimal endocardial border definition on echocardiography. In 30 adult patients, apical two and four chamber views were performed. In comparison to biplane cineventriculography enddiastolic and endsystolic volumes, stroke volume, ejection fraction, and regional wall function were assessed for heart cycles with and without left ventricular contrast. In comparison to biplane cineventriculography echocardiography underestimates enddiastolic (167+/-64 ml, 111+/-43; p<0.0001) and endsystolic volumes (77+/-63 ml, 54+/-40 ml; p<0.0002), stroke volume (90+/-25 ml, 57+/-17 ml; p<0.0001), and ejection fraction (58+/ 16%, 55+/-14%; p<0.03). By contrast echocardiography ejection fraction (58+/-16%) agreed with the angiocardiographically measured ejection fraction. Furthermore, after contrast injection correlations improved between cineventriculography and echocardiography for the assessment of left ventricular enddiastolic volumes (without contrast r = 0.90, SEE = 19 ml; with contrast r = 0.93, SEE = 19 ml), endsystolic volumes (without contrast r = 0.94, SEE = 14 ml; with contrast r = 0.95, SEE = 15 ml), stroke volume (without contrast r = 0.63, SEE = 14 ml; with contrast r = 0.67, SEE = 14 ml), ejection fraction (without contrast r = 0.84, SEE = 8%; with contrast r = 0.88, SEE = 7%), regional wall motion (p<0.01) and its reproducibility (p<0.02). In adult patients with suboptimal endocardial border delineation intravenous contrast echocardiography improves the assessment of left ventricular ejection fraction, regional wall motion, and its reproducibility without severe side effects. PMID- 10525926 TI - [Efficacy of metoprolol in prevention of supraventricular arrhythmias after coronary artery bypass grafting]. AB - Atrial fibrillation is in 20-50% the most frequent dysrhythmia after coronary artery bypass grafting (CABG) and a possible cause for hemodynamical complications and prolongation off the medical treatment in patients. Therefore, the effect of beta-blocking with metoprolol for prevention of supraventricular arrhythmias (SVA) was investigated in a prospective and randomized trial. 200 patients after CABG were randomized in a drug and control group (average age 63.2 years, 154 male, 46 female). Patients of the drug group (n=100) were treated with metoprolol (1mg/kg/BW) beginning on day one after operation, whereas patients of the control group (n=100) received therapy only in case of occurrence of atrial fibrillation. ECG, blood pressure, and electrolyte concentrations were measured regularly until the tenth day after surgery. Reasons for exclusion were an ejection fraction (< 30%, SA- and AV-block or simultaneous application of epinephrine and metoprolol. There were no significant differences between the patients of drug and control group with respect to age, sex ejection fraction, previous medication, number and type of bypass grafts, cardiopulmonary bypass time, and perioperative ischemic events. However, a statistically significant difference was seen in the occurrence of supraventricular arrhythmias in both groups, 4 patients of the therapy group (4%) in contrast to 37 patients of the control (37%) developed supraventricular arrhythmias during the postoperative observation period (p<0.0001). Both groups differed in total time of hospital stay by 1.5 days (control group: 9.83+/-2.88 days; drug group: 8.42+/-2.81 days), which was statistically significant (p<0.05). All patients of the drug group could be discharged with a stable sinus rhythm, whereas 7 patients of the control group were discharged with persistent atrial fibrillation. The difference was statistically significant as well (p<0.01). Neither typical side effects of metoprolol, nor AV-blocks, bradycardia (f<60/min) or symptoms of low blood pressure could be observed. The conclusion of this trial is a recommendation for a preventive application of 50mg metoprolol/day after coronary artery bypass surgery, which can reduce the incidence of SVA as well as the hospital stay statistically significant. PMID- 10525927 TI - [Prospective evaluation of effect of carvedilol therapy on heart rate variability in patients with dilated cardiomyopathy]. AB - The aim of the present study was to assess the effects of carvedilol therapy in addition to conventional heart failure therapy on heart rate variability (HRV) and on left ventricular function in 14 patients with mild to moderate heart failure due to idiopathic dilated cardiomyopathy (IDC). After a 3- to 4-week titration period, carvedilol was titrated up to 50mg daily, or the highest dose tolerated (at least 25mg daily). Maintenance treatment was then continued for 8 weeks. Digital 24-hour Holter recordings were obtained at baseline and after 8 weeks of carvedilol therapy. HRV for the entire 24-hour period was computed in the time domain using the Oxford Medilog Excel 2 analysis system. Measures of HRV included the mean of all coupling intervals between normal beats (RRm), the standard deviation of all normal RR intervals (SDNN), the square root of the mean of the squared differences between adjacent normal RR intervals (rMSSD), and the proportion of adjacent normal RR intervals differing >50 ms (pNN50). Additional treatment with carvedilol induced a significant increase in HRV: SDNN increased from 77+/-21 ms to 110+/-22 ms (p=0.001), rMSSD from 19+/-7 ms to 26+/-7 ms (p=0.02), and mean pNN50-value increased from 1.7+/-1.3% to 5.5+/-4.5% (p<0.01) under therapy with carvedilol. Mean heart rate on carvedilol calculated over 24 hours was 13 beats less than at baseline (75 bpm versus 88 bpm, p<0.01). After 2 months of additional treatment with carvedilol, both hemodynamic and clinical parameters improved: left ventricular ejection fraction increased from 24+/-7% to 30+/-10% (p<0.05), and New York Heart Association class decreased from 2.5+/-0.8 to 1.8+/-0.7 (p<0.05). In summary, eight weeks of additional carvedilol therapy induced a significant increase in HRV parameters related to parasympathetic activity in patients with IDC. Whether increased vagal tone may contribute to the protective effect of carvedilol has to be evaluated by further studies. PMID- 10525928 TI - [Carotid stenting: acute results and complications]. AB - The treatment of carotid stenoses with balloon angioplasty and stenting is a new and not generally established method. During a one year period 65 patients (22 female, 43 male, mean age: 73 years, 47 with neurologic symptoms, 8 with contralateral carotid occlusion) with significant (>70%) carotid stenosis were treated with balloon angioplasty and balloon expandable stents. The primary technical success rate was 98% (65/66 patients) respectively 99% (69/70 stenoses). A combined procedure was performed in 11 /17%) patients with stenting in both carotid arteries in 4 patients with additional coronary interventions in 6 patients and stenting of the origin of the common carotid artery in one patient.Severe neurologic complications occurred in 4 (6.2%) patients (1 death, 1 major stroke, 2 minor strokes) and short lasting neurologic deficits in additional 4 (6.2%) patients. Cardiovascular complications were not observed. Local (inguinal) problems occurred in 3 (4.5%) patients (2 aneurysma spuria, 1 transfusion for hematoma). Frequently, balloon insufflation was associated with bradycardia (40%) and additional hypotension (11%). In summary, carotid stenting can be performed with technically high success rates (99%), but it is adversely influenced by not infrequent thromboembolic cerebral events (12.4%). The possibility to perform combined procedures with interventions in other vessels (both carotid arteries, coronary arteries, aortic arch arteries) is advantageous. PMID- 10525929 TI - [Rupture of the pulmonary artery - fatal complication by pulmonary artery balloon tipped catheter]. AB - The flow directed balloon-tipped pulmonary artery catheter introduced by Swann and Ganz 1970 has made possible the measurement of filling pressures in the heart and is extensively used in operating rooms, in catheterization laboratories and in intensive care units. The rupture of the pulmonary artery is an uncommon complication associated with a high mortality rate exceeding 50 percent. Main symptoms are cough, hemoptysis, dyspnea and cardiac shock. Complications occur if guidelines for the safe use of the balloon-tipped catheters are not strictly followed: Excessive catheter manipulation, advancing the catheter tip too far peripherally and leaving the inflated balloon in the wedge position for long periods should be avoided. Patients with pulmonary hypertension as well as elderly and anticoagulated patients are at greater risk. PMID- 10525931 TI - [Cross your heart]. PMID- 10525930 TI - [Non-surgical management of a perforated left anterior descending coronary artery following cardiopulmonary resuscitation]. AB - A 49 year old male developed cardiocirculatory arrest following laparoscopic surgery of an inguinal hernia. Cardiopulmonary resuscitation (CPR) was started. The primary ECG showed ventricular fibrillation, after defibrillation a tachyarrhythmia and a newly developed right bundle branch block were documented. In addition, ST-elevations were seen in the left precordial leads. After 20 min of CPR the circulation was reestablished with high doses of catecholamines. Transthoracic echocardiography demonstrated a 7 mm pericardial effusion with mild impression of the right ventricular free wall. The patient underwent urgent heart catheterization for suspected pulmonary embolism (differential diagnosis: acute myocardial infarction). Pulmonary angiography demonstrated floating thrombi in the left main pulmonary artery, which could be fragmented using a pigtail catheter. Pulmonary angiography was followed by coronary angiography, which demonstrated a sharply interrupted left anterior descending artery (LAD), while the coronary arteries in general were found to be regular. The history, the morphology of the LAD-interruption, the concomitant pericardial effusion and the sternal and rib fractures were consistent with a type III coronary perforation in the classification of Sutton and Ellis. As contrast medium penetration into the pericardial space persisted after recanalization of the LAD, a 19 mm stent graft (Jostent) was used for closure. These grafts are constructed using a sandwich technique with an ultrathin layer of expandable polytetrafluorethylene being placed between two stents. After implantation of the stent graft no more contrast medium penetration was documented. CONCLUSION: Coronary perforations following blunt chest trauma is a rare complication, which has been described only once following CPR (2). Stent grafts can be used safely for acute closure of such perforations. PMID- 10525932 TI - [Cardiac chemoreflex sensitivity in relation to various forms of breathing]. PMID- 10525933 TI - [Reports by the German Society of Cardiology--Heart- and Cardiovascular Research. Clinical Epidemiology Working Group]. PMID- 10525934 TI - [Reports by the Working Group of Executive Cardiologic Hospital Physicians e. V]. PMID- 10525935 TI - [Reports by the German Society of Pediatric Cardiology. Quality Standards for echocardiography in children and adolescents. Recommendations by the German Society of Pediatric Cardiology for echocardiography studies in childhood and adolescence]. PMID- 10525936 TI - [31st Annual Meeting of the German Society of Pediatric Cardiology. Wuppertal, 2 to 5 October 1999]. PMID- 10525937 TI - Indications and results of fronto-lateral laryngectomy using a combined endolaryngeal and external approach. AB - Patients with laryngeal anterior commissure, cord-commissure, bilateral anterior cord-commissure carcinomas (T1 and T2 N0 M0) were subjected to a new method of frontolateral laryngectomy using a combined endolaryngeal and external approach. The proposed surgical procedure allowed the removal of the anterior commissure and part of one or both vocal cords in a single unit, together with the cartilaginous framework, respecting the integrity of the superior portion of the thyroid cartilage. The internal procedure permitted an accurate delimitation of the posterior part of the laryngeal neoplasm. In particular, this was performed during suspension microlaryngoscopy using the CO(2) laser or traditional cutting tools for section of the laryngeal visceral structures to the internal surface of the thyroid cartilage. Following this, the external approach included incision of the external perichondrium along the superior edge of the thyroid cartilage and along the median line, from the incisura to the inferior edge of the thyroid cartilage. The superior opening of the larynx is made side-to-side and the epiglottis separated at the level of the superior edge of the thyroid cartilage. The ends of the section are joined together with the superior parts of the section created during the laryngeal approach. Then progressive craniocaudal detachment of the internal perichondrium is performed backwards until the endolaryngeal sections are reached and downwards to the insertion of the cord ligaments. The inferior opening of the larynx is made by a horizontal section of the cricothyroid membrane at the level of the superior edge of the cricoid cartilage. Section of the thyroid cartilage is therefore performed in a trapezoidal shape. This section involves the inferior part of the protruding corner of the thyroid cartilage. After joining the ends of the cricothyroid section with the inferior extremities of the endolaryngeal sections, the surgical specimen is removed as a single unit. The method must be used only after accurate clinical evaluation. It is mostly recommended in subjects with cord-commissure carcinomas previously treated with radiotherapy. The results obtained were extremely satisfactory both as regards survival and functional results. In all, 27/28 patients (96.4%) were free from disease. The quality of voice was satisfactory but hoarse in 30% and breathy in 70% of the patients. PMID- 10525938 TI - Clinical application of proliferating cell nuclear antigen, oncoprotein p53 and tumor front grading analysis in patients operated on for laryngeal cancer. AB - The authors assessed proliferating cell nuclear antigen (PCNA), p-53 oncoprotein and morphologic tumor front grading (TFG) in patients with advanced squamous cell carcinoma (SCC), of the larynx and a poor prognosis and tried to find a correlation with tumor stage, the Broders grading system, local and neck lymph node metastases, as well as nodal and local recurrences. In addition, utility of the parameters investigated was evaluated in developing a prognostic factor model, using uni- and multivariate Cox regression analysis. Included in this study were 54 patients (mean age 57 years +/- 8.6). PCNA-positive staining was found in all but one patient with advanced disease, while p-53 stained positively in only 24 subjects (44.4%). The PCNA index ranged from 4.6 to 59.0% (mean, 23.4 +/- 11. 0) and the p-53 index varied from 4.0 to 42.0% (mean, 17.2 +/- 8.6). The TFG score ranged from 9 to 23 points (mean, 15.1 +/- 3.2). PCNA, p-53 and TFG were found to be the markers that provided significant additional information about the biological behavior of tumor cells. The high variability of the results (PCNA, p-53) and high percentage of negatively stained cells (p-53) reduced their application in clinical use. PCNA correlated with tumor grade, G (r = 0.38; P < 0. 01), but negatively with nodal (N) disease(r = -0.37; P < 0.01). The mean values of PCNA and p-53 index were higher in the subgroup with local recurrences. Our present attempt to develop a useful prognostic factor model failed. PMID- 10525939 TI - Immunohistochemical analysis of lymphocytic infiltration in the tumor microenvironment in patients operated on for laryngeal cancer. AB - The aim of this study was to evaluate semiquantitative and qualitative analysis of lymphocytic infiltrations in a neoplasm microenvironment in patients with laryngeal cancers and the correlation analysis between the intensitivity degree and composition of lymphocytic infiltration in foreseeing a survival time and probability of the appearance of lymph node metastases. Postoperative specimens from 43 patients (Upper Silesia region) operated on for laryngeal cancer in the 2nd ENT Department, Silesian Medical University in Zabrze between 1985 and 1995 all had unfavorable courses due to tumor recurrences. The patients' ages ranged from 39 to 79 years (mean 57 years). Tissue specimens were subjected to routine processing. The degree of pathological changes was ascertained and immunohistochemical preparations of laryngeal tissue were prepared according to generally accepted methods. The following primary monoclonal antibodies were used: CD 3, CD 20, CD 43, CD 45 RO, CD 56. The distribution analysis of the intensity of the phenotype CD 43 evaluated the lymphocytic infiltration in relation to differentiation of the whole study group. The intensity of CD 43 cell infiltration increased in the group of patients with lymph node metastases. In patients with stage IV disease, a relationship was found between survival time and intensity of cell infiltrations with CD 43 and CD 45 RO lymphocytes. The influence of these two lymphocyte phenotypes in the patient subgroups - one after total laryngectomy with confirmed lymph node metastases and the other group without lymph node metastases - showed their prognostic value. Our analysis of lymphocytic infiltration, mostly of CD 43 cells, in the neoplasm microenvironment indicated a prognostic value for determining a shorter survival time and the possibility of lymph node metastases in patients with recurrences of cancer. PMID- 10525940 TI - Laser arytenoidectomy in the treatment of bilateral vocal cord paralysis. AB - The introduction of the CO2 surgical laser into laryngeal microsurgery has made resection of the posterior vocal cord together with the arytenoid cartilage possible. Since November 1990, 30 arytenoidectomies, 17 partial cordectomies and 18 bilateral cordectomies as described by Kashima were performed by means of a CO(2) laser in patients with bilateral paralyses of the vocal cords. In this group there were 58 women and 7 men. The patients' ages ranged from 28 to 71 years (mean, 46.7 years). In one case the operation was performed twice: the right arytenoid cartilage was excised initially and the left arytenoid cartilage was removed in the second procedure. Three patients required tracheotomy before being transformed to the ENT Clinic, Poznan. The etiologies of the vocal cord paralyses were complications arising from thyroid gland surgery (n = 62), trauma (n = 2) and excision of a bilateral glomus caroticum tumor. In all patients except one postoperative recovery was correct and no breathing difficulties were observed after extubation. In the one failure after operation endolaryngeal scar tissue resulted in glottic stenosis. PMID- 10525941 TI - Extended fronto-lateral laryngectomy with simultaneous reconstruction by means of a mucochondral nasal septum flap. AB - As treatment for laryngeal cancers the extended fronto-lateral laryngectomy allows tumor to be removed that involves the glottis with the vestibule of the larynx. Since 1979 we have performed 417 extended fronto-lateral laryngectomies with simultaneous reconstruction of the anterior part of the larynx using a mucoperichondral flap from the anterior septum. This operation was indicated for stage T1b cancer (175 cases), stage T2 tumor (231 tumors) and T3 extended operations with epiglottectomy (11 cases). For the total number of 417 patients operated on, 19 cases during the 5-year period of following had local recurrences that required total laryngectomy. In 11 cases, metastases to the cervical lymph nodes required surgical neck dissection. In the remaining 387 cases, a normal air passage and a large laryngeal lumen were found. In 15 cases graft rotation into the larynx made respiration and phonation difficult but without dyspnea occurring. Follow-up to a maximum of 18 years showed that the oncological and functional results after this surgical procedures are encouraging. PMID- 10525942 TI - Endoscopic CO(2) laser therapy of selected cases of supraglottic marginal tumors. AB - Endoscopic CO(2) laser intervention can be used as conservation surgery for supraglottic laryngeal carcinomas in carefully selected patients. We analyzed retrospectively our experience in managing patients with early supraglottic carcinomas operated on at the Clinic of Otorhinolaryngology, Szeged, Hungary, during the 10-year period between 1987 and 1997. Conservation surgery was the treatment of choice in 187 patients, but only 23 (12%) were selected for endoscopic CO(2) laser surgery. Laser surgery was indicated predominantly for T1 cancer of the epiglottis (n = 15), but was also performed for T2 cancers (n = 8). Of the 23 supraglottic tumors treated, 16 had no signs of recurrence to date (1.5 to 9 years after surgery) a local control rate of 70%. Six patients with recurrences underwent salvage therapies that included repeated laser excisions (n = 3), radiotherapy (to 60 Gy), horizontal supraglottic laryngectomy and total laryngectomy. One patient was not resectable because of multiple metastases. Our experience with endolaryngeal CO(2) laser excision indicates that it is a reasonable method in selected cases of supraglottic tumors, but one-third of the patients required salvage treatment. PMID- 10525943 TI - The value of CT scans in improving laryngoscopy in patients with laryngeal cancer. AB - This retrospective study assessed the value of computed tomography (CT) scan with contrast in improving the staging accuracy of indirect and direct laryngoscopy. We compared the preoperative staging obtained by the two latter procedures with postoperative histopathological findings in 187 patients operated on for laryngeal cancer. Of these cancers, 98 were supraglottic, 82 glottic and 7 subglottic in origin. The staging accuracy of laryngoscopy was 51.3% and CT was 70.1%. When the two procedures were combined, the staging accuracy was 80.2%. The accuracy of the CT increased from glottic to supraglottic to subglottic tumors, although the accuracy of laryngoscopy decreased in the same direction. Laryngoscopy alone tended to understage larger tumors (pT3 and pT4), whereas CT underestimated the smaller ones (pT1 and pT2). Our data suggest that in order to plan the best treatment both laryngoscopy and CT should be used in making the diagnosis. PMID- 10525944 TI - Indication and surgical technique for extended hemilaryngectomy. AB - Previous experience has shown that conservation of the healthy hemilarynx is possible for the treatment of extended lateralized laryngeal and hypopharyngeal cancers. One major indication for supracricoid hemilaryngectomy is glottic cancer involving either the arytenoid or Morgagni's ventricle. Hemilaryngopharyngectomy is also indicated when tumor of the pyriform sinus involves its anterior part, lateral wall, medial wall and ary- and pharyngoepiglottic folds. Supracricoid hemilaryngopharyngectomy was performed in seven cases. The resection consisted of removal of the surpacricoid hemilarynx and ipsilateral pyriform sinus. The reconstruction was performed by elevating the posterior pharyngeal wall and suturing the mucosa to the midline. This surgical technique provided good functional results. Patients had no airway impairments, but the recovery of satisfactory deglutition may require 14-32 days. Phonatory rehabilitation was successful, and each patient had a satisfactory vocal quality. Postoperative irradiation did not affect the functional results. PMID- 10525945 TI - CO(2) laser posterior ventriculocordectomy for the treatment of bilateral vocal cord paralysis. AB - We reviewed our clinical experience between 1991 and 1997 concerning use of the CO(2) laser for posterior ventriculocordectomy (PVC) for the treatment of bilateral vocal cord paralysis. Pre- and postoperative functional evaluation was assessed in a prospective setting. In all, 41 patients (33 females and 8 males) underwent an endoscopic CO(2) laser PVC. Pre- and postoperative pulmonary function tests documented a significant statistical improvement in the parameters considered. Sixteen of 21 previously tracheostomized patients were decannulated within 15 months of operation. In no case was a postoperative tracheostomy required. We found no evidence of subclinical aspiration among our cases. Evaluation of vocal parameters by spectrographic analysis was assessed in 20 patients and revealed a postoperative reduction in voice quality. Laser CO(2) PVC seems to be an effective and reliable surgical procedure that allows for rapid decannulation and gives stable results with a low incidence of revision surgery and functional failures. PMID- 10525946 TI - Reversible immediate and definitive lateralization of paralyzed vocal cords. AB - The author reports on glottis dilation operations based on the endoextralaryngeal suture technique he has developed. In all, 101 patients were operated on for bilateral recurrent nerve paralysis using different variations of the above method, of which 73 have had more than 1 year of follow-up. Dilation was performed in 52 patients following tracheostomy, whereas no tracheostomy was performed in 21 patients. In 9 cases irreversible laterofixation without tracheostomy was performed with good results. In 12 patients a reversible glottis dilating operation was carried out without tracheostomy not long after the development of bilateral paramedian position of the vocal cords. Tracheostomy was necessary in 1 of 12 patients, who underwent reversible glottis dilating operations. In this case later reoperation, using a definitive endoscopic glottis dilating operation, was performed with success. Three patients required reoperation using open surgical procedures after irreversible endoscopic laterofixation methods. PMID- 10525947 TI - Endoscopic microsurgical management of scars in the posterior commissure and interarytenoid region resulting in vocal cord pseudoparalysis. AB - The author reports his treatment of scars in the interarytenoid region of the larynx and the surgical management of scars in the posterior commissure based on the endoextralaryngeal suture technique and the needle carrier he developed. After the examination by electromyography, the scars are separated endoscopically. To prevent recurrent scarring and adhesions, two procedures are applied to the posterior commissure. When the size of the scars does not exceed 5 mm, both vocal cords are lateralized temporarily once the scars have been separated. In this way, the surfaces of the scars do not touch, thus preventing adhesion. When the scars are larger than 5 mm but less than 10 mm, a soft silicon stent replicating the shape of the lumen is fixed between the scars in the lumen of the larynx using the author's technique and suture device. The laterofixing sutures and silicon stent are removed in the fourth postoperative week. As a result of the operation, the lumen of the larynx has been found to be of adequate width and suitable for normal breathing and sound formation. These procedures have been applied successfully in 12 out of 13 patients. PMID- 10525948 TI - The value of ultrasound examination in preoperative neck assessment and in early diagnosis of nodal recurrences in the follow-up of patients operated for laryngeal cancer. AB - The purpose of this study was to prove the superiority of ultrasound (US) examination of the neck in comparison to palpation, to reveal unpalpable nodes (pN0) before surgery, and to allow for the early detection of nodal recurrences in patients with laryngeal cancers. In all, 1,120 patients with laryngeal cancers were operated on between 1990 and 1997. All underwent palpation and neck US before surgery. In the pN0 group US revealed enlarged lymph nodes in 261/505 cases, of which 63 (24.14%) were confirmed as metastatic by histology. All 1,120 patients underwent regular postoperative US follow-up. Nearly 5,000 US examinations were performed; 136 patients who developed nodal recurrences had surgical salvage procedures. In this group 61 patients had small, nonpalpable lesions, and 46 patients discrete and slight changes in scarred necks. In this latter group of 117 patients with nonpalpable lesions, 105 cases were histologically confirmed as metastatic. In postoperative check-ups, metastases were suspected in sonographically detected subclinical nodes, but the US scans obtained were difficult to interpret. In these cases, because of the dynamics of lesion changes, US was repeated two to three times at 10- to 14-day intervals. This reappraisal enabled us to exclude malignancy in regressing nodes, as well as to obtain the stable picture of scar, and support a diagnosis while the lesion grew larger or a central area of low attenuation or hypoechogenic echos appeared in the nodal capsule. Successful radical reoperation for tumor was done on110 patients; 78 patients underwent successful salvage surgery after an early US diagnosis. The sonographic-surgical correlation was nearly 95% and the sonographic-histological correlation was 90%. The follow-up period was 1-49 months. In all preoperative assessments US was found to be a valuable tool in the staging of laryngeal cancer and planning the extent of surgery. Close follow-up with US after radiotherapy and initial operation was essential for the early detection of tumor recurrences, making surgical salvage still feasible. PMID- 10525949 TI - Voice function in patients after extended fronto-lateral laryngectomy. AB - The main purpose of this study was subjective and objective evaluation of voice function before and after extended frontolateral laryngectomy, followed by reconstruction with a mucochondrial graft from the nose. The next aim of this study was analysis of phoniatric and acoustic results depending on: (1) the extent and localization of the neoplastic process; (2) the wound-healing process (including the time of decannulation and effective deglutition); (3) the substitute mechanism of phonation. In all, 40 patients (37 men and 3 women) having T1B (67.5%) and T2 (32.5%) glottic cancers were examined before and after the operation. The ENT investigation included each patient's history from the beginning of illness, a subjective evaluation of the voice by patients before and after the operation, determination of laryngeal mobility of the larynx and indirect laryngoscopy during phonation. Phoniatric examination included voice recordings, average voice pitch, voice range, maximum phonation time, microlaryngoscopy and microstroboscopy. Acoustic analysis covered the harmonic structure of the voice, jitter, shimmer, noise components and basic frequency. Results showed that the extent of the neoplastic process before operation did not affect essential voice quality after operation in stage T1B and T2 disease. The maximum phonation time was shortened in T1B patients from 16.5 s to 9.28 s and in T2 from 16.7 s to 9.8 s. The average voice pitch was decreased in T1B from 177.3 Hz to 111.74 Hz and in T2 from 163.8 to 99 Hz. A correlation between acoustic analysis and phoniatric examinations was found. The value of the first formant decreased by 15.1 Hz, jitter increased 2%, shimmer increased 0.9 dB and basic frequency decreased. The voice quality after operation showed a statistically significant dependence on the mechanism of phonation. The best results were found in patients in whom phonation was the result of removing the vocal fold and postoperative scar vibration. The worst results were found in those patients with a sphincter mechanism of phonation. PMID- 10525950 TI - Indications for frontolateral laryngectomy and prognostic factors of failure. AB - The aim of this study was a retrospective analysis of the oncological results in a group of patients treated by frontolateral laryngectomy using clinical and histopathological correlations in order to review the indications for surgery. In all, 150 patients underwent frontolateral laryngectomy as described by Leroux Robert. All were staged according to the 1992 UICC TNM classification. Factors examined were clinical T, histopathological T, tumor infiltration of the anterior commissure and the vocal cord muscle, survival without disease and the percentage of local relapses. Twenty-one patients had local relapses (14%), while four patients developed second primary tumors (2.7%). Among the different correlations examined, microscopic infiltration of the anterior commissure was related to a greater number of local relapses (25.5% vs 5%) and a 55% survival with with no evidence of disease (NED). The crude 5-year NED survival was 66% and was influenced by second primary tumors and metastases (7.4%) and non-oncological diseases (14.6%). These data show the need for a re-evaluation of the indications for frontolateral laryngectomy because subtotal reconstructive laryngectomy could be performed more safely in the more advanced cases. In contrast, cases with more limited tumors might be better treated by laser for a more functional and cost beneficial result. PMID- 10525951 TI - Clinical experience with Ciaglia's percutaneous tracheostomy. AB - During a 6-year period the authors performed percutaneous dilational tracheostomy (PDT) in 304 cases using the technique introduced by Ciaglia in 1985. A following study on 40 patients evaluated late complications of PDT; none of the patients developed laryngotracheal stenosis. In our experience PDT has been a very sure technique of securing the airway. PMID- 10525952 TI - Basic aspects of photon transport through matter with respect to track structure formation. AB - After a short summary of the most important physical aspects of photon interaction with matter and of the main elements of photon transport simulation, some basic features of photon tracks in liquid water are discussed. These include the statistical distribution of the number of photon interactions caused during the complete photon slow-down, the corresponding mean value, the distance distribution of photon interactions, and the spectral distribution of secondary electrons or, which is the same, the spectral distribution of the start energy of secondary electron tracks. The latter distribution can easily be interpreted in terms of the track entity concept of Mozumder and Magee, which, therefore, proves to be the most natural concept of track structure analysis in the case of photon irradiation. PMID- 10525953 TI - Condensed-history Monte-Carlo simulation for charged particles: what can it do for us? AB - Condensed-history (CH) Monte-Carlo (MC) groups together the vast number of individual charged-particle collisions using multiple scattering theory for elastic angular changes and stopping power for energy losses. CH codes such as EGS4 have been enormously successful in simulating the transport of electrons, for example, in radiotherapy. MC-derived values of the water-to-air stopping power ratio, s(w/air), are used in all modern codes of practice for absolute dose determination in radiotherapy clinics. MC can also directly yield the dose ratio Dmed/Ddet for a dosimeter in a medium, and Correlated Sampling has been exploited to increase the efficiency, e. g., the central electrode in an ion chamber (aluminium vs. graphite). The extremely low density of the gas in an ion chamber poses problems for CH codes. However, multiple scattering can now be combined with single scattering and is expected to finally resolve important chamber perturbation effects. An exciting application of CH MC in radiotherapy is the computation of dose distributions in patients. Currently one can achieve an uncertainty around 1% (1 SD) in mm-sized voxels in several minutes for an electron beam and in around an hour for a photon treatment plan on hardware costing less than $20,000, and thus avoid all the various approximations conventionally used to account for inhomogeneities. In the microdosimetry/track structure field, CH codes have shown that the fluence (dPhi/dE) per unit dose at low electron energies is virtually independent of incident particle energy or depth, which simply explains the negligible RBE variation. PMID- 10525954 TI - The track structures of ionizing particles and their application to radiation biophysics. I. A new analytical method for investigating two biophysical models. AB - A new approach to the interpretation of the effects of radiation on cells is described, in which sample particle tracks are constructed using a Monte Carlo computer program and the exposure of cellular targets to these tracks is simulated using a second program known as BIOPHYS. Data on the shapes and DNA contents of the cell nuclei are obtained from the literature. It is assumed that the sensitive material is DNA, and that the target is divided into cubes of approximately 2 nm (the diameter of the DNA helix) per side; the numbers of these cubes containing different numbers of ionizations are derived. Two different methods of analysing the output of BIOPHYS are described. In the first, it is assumed that lethality is caused by the occurrence of a number of ionizations equal to or greater than a certain threshold in one cube; in the second method, it is assumed that only two ionizations are required, in different parts of the cube, but that only some fraction of the cube is sensitive. These models have been applied to the interpretation of the variation of radiosensitivity with a linear energy transfer (LET) of spores of Bacillus subtilis exposed wet and dry, and good fits to the published experimental data were obtained using both models. Fits to experimental data for a range of other cell lines will be presented in a second paper. PMID- 10525955 TI - Response of pig lung to irradiation with accelerated 12C-ions. AB - The response of pig lungs to irradiation with 12C-ions was assessed in two experiments to validate the procedures for heavy ion therapy planning at the Gesellschaft fur Schwerionenforschung (GSI) and to explore their range of applicability. In both experiments, the target volume (spread-out Bragg peak, SOBP) was planned to be a 4 cm long cylinder with a diameter of 4 cm. Doses in the SOBP were prescribed to be equivalent to 5x4 Gy, 5x5.5 Gy and 5x7 Gy of x rays in the first experiment, and to 5 fractions of 7 Gy and 9 Gy in the second experiment. The lung response in the first experiment was less than expected on the basis of earlier experiments with photons. Pneumonitis reaction and chronic fibrotic changes were observed outside the prescribed high-dose region. In the second experiment, the effects were more pronounced than had been expected on the basis of the first experiment. Changes were most intense in the high-dose region, but were also seen throughout the lung along the beam channel. Moreover, significant skin reactions were observed at the beam entrance site in all animals and - less pronounced - at the beam exit site in 3 of the 6 animals. In conclusion, the complex irradiation geometry of the pig lung, the changes of body weight between the two experiments, and insufficient accounting for a change in the relative biological effectiveness (RBE) computation led to substantial deviations of the observed reactions from expectations, the reasons for which could be identified in a subsequent analysis. The less pronounced lung reaction in the first experiment was due to an overestimation of RBE in a preliminary version of the algorithm for its determination. The extension of the fibrotic reaction resulted from the smear-out of the high-dose region due to density variations in tissue structures, respiratory movement, and limited positioning accuracy. The skin reactions at the entrance port reflect the different treatment geometry in the two experiments. The one unexplained observation is the mild skin reaction that was observed in the second experiment at the beam exit site. PMID- 10525956 TI - Induction of cAMP-dependent protein kinase A activity in human skin fibroblasts and rat osteoblasts by extremely low-frequency electromagnetic fields. AB - Sinusoidal extremely low-frequency electromagnetic fields (ELF-EMF; 7-8 mT, 20 Hz) have already been shown to inhibit proliferation and to accelerate terminal differentiation of human skin fibroblasts in vitro. In order to elucidate the underlying processes of signal transduction, we analysed the activity of cAMP dependent protein kinase (PKA). EMF exposure for 60 min resulted in an increased PKA activity in human skin fibroblasts (2-fold) and rat embryonic osteoblasts (1.7-fold). Long-term exposure for up to 7 days with a constant 1 h-on/1 h-off EMF exposure rhythm indicated a transient stimulation of PKA activity during the first two exposure rhythms followed by a decrease to the baseline levels of sham exposed controls. Based on these results, we postulate that a modulation of proliferation and differentiation processes in cells of mesenchymal origin is triggered by an immediate and transient EMF-induced increase in PKA activity. PMID- 10525957 TI - Estimation of the target stem-cell population size in chronic myeloid leukemogenesis. AB - Estimation of the number of hematopoietic stem cells capable of causing chronic myeloid leukemia (CML) is relevant to the development of biologically based risk models of radiation-induced CML. Through a comparison of the age structure of CML incidence data from the Surveillance, Epidemiology, and End Results (SEER) Program and the age structure of chromosomal translocations found in healthy subjects, the number of CML target stem cells is estimated for individuals above 20 years of age. The estimation involves three steps. First, CML incidence among adults is fit to an exponentially increasing function of age. Next, assuming a relatively short waiting time distribution between BCR-ABL induction and the appearance of CML, an exponential age function with rate constants fixed to the values found for CML is fitted to the translocation data. Finally, assuming that translocations are equally likely to occur between any two points in the genome, the parameter estimates found in the first two steps are used to estimate the number of target stem cells for CML. The population-averaged estimates of this number are found to be 1.86x10(8) for men and 1.21x10(8) for women; the 95% confidence intervals of these estimates are (1.34x10(8), 2. 50x10(8)) and (0.84x10(8), 1.83x10(8)), respectively. PMID- 10525958 TI - A registry for exposure and population health in the Altai region affected by fallout from the Semipalatinsk nuclear test site. AB - A registry of the rural population in the Altai region exposed to fallout from nuclear tests at the Semipalatinsk test site (STS) was established more than four decades after the first Soviet nuclear explosion on August 29, 1949. Information about individuals living in an exposed and a control area was collected using all available local sources, such as kolkhoz documentation, school registries, medical treatment records and interviews with residents. As a result, a database comprising an exposed group of 39 179 individuals from 53 Altai region villages, 6769 external and 3303 internal controls was compiled. For several settlements, effective dose estimates reached the level of 1.5 Sv, while the average effective dose estimate in the exposed group was 340 mSv. Dosimetric data, vital status information and health records gathered at rayon and village medical facilities are held in the registry. Cause-of-death information for deceased residents is obtained from death registration forms archived at the Altai region vital statistics office. At present, a follow-up of approximately 40% of the population exposed in 1949 has been done. More will be added by searching for migrants to the larger towns of the Altai region, i.e. Barnaul, Rubtsovsk and Biisk. In order to assess the influence of radiation exposure, analytical studies with a case control design for stomach and lung cancer are currently being prepared. The number of known cases is sufficient to detect an odds ratio of 1.5 at the 95% confidence level. Epidemiological studies in populations affected by fallout from STS may be equally important to the atomic bomb survivors' study for the direct quantification of radiation effects. The range of exposure rates experienced will extend the acute high-dose-rate findings from Hiroshima/Nagasaki towards acute and protracted lower exposures, which are more relevant for radiation protection issues. PMID- 10525962 TI - Announcement PMID- 10525959 TI - Childhood cancer and residential radon exposure - results of a population-based case-control study in Lower Saxony (Germany). AB - A population-based case-control study on risk factors for childhood malignancies was used to investigate a previously reported association between elevated indoor radon concentrations and childhood cancer, with special regard to leukaemia. The patients were all children suffering from leukaemia and common solid tumours (nephroblastoma, neuroblastoma, rhabdomyosarcoma, central nervous system (CNS) tumours) diagnosed between July 1988 and June 1993 in Lower Saxony (Germany) and aged less than 15 years. Two population-based control groups were matched by age and gender to the leukaemia patients. Long-term (1 year) radon measurements were performed in those homes where the children had been living for at least 1 year, with particular attention being paid to those rooms where they had stayed most of the time. Due to the sequential study design, radon measurements in these rooms could only be done for 36% (82 leukaemias, 82 solid tumours and 209 controls) of the 1038 families initially contacted. Overall mean indoor radon concentrations (27 Bq m(-3)) were low compared with the measured levels in other studies. Using a prespecified cutpoint of 70 Bq m(-3), no association with indoor radon concentrations was seen for the leukaemias (odds ratio (OR): 1.30; 95% confidence interval (95% CI): 0.32-5.33); however, the risk estimates were elevated for the solid tumours (OR: 2.61; 95% CI: 0.96-7.13), mainly based on 6 CNS tumours. We did not find any evidence for an association between indoor radon and childhood leukaemia, which is in line with a recently published American case-control study. There is little support for an association with CNS tumours in the literature. PMID- 10525963 TI - Variable stability of chromosomes containing amplified alpha-satellite sequences in human mesenchymal tumours. AB - Alpha-satellite sequences are found in the centromeric region of all human chromosomes and have been implicated in centromeric function. We describe the structure and behaviour of chromosomes containing amplified human alphoid DNA from chromosome 12, in an osteosarcoma cell line (OSA) and an atypical lipomatous tumour (ALT). In OSA, the amplified material was detected in one large marker chromosome, whereas in ALT amplified sequences were observed in chromosomes of variable number and appearance. The marker in OSA was mitotically stable, but those in ALT exhibited a high degree of mitotic instability, forming bridges at anaphase and chromatin strings between interphase nuclei. The amplified alpha satellite arrays reacted positively with human anti-centromeric antiserum and anti-centromere protein B antibodies in both tumours. Centromere protein C, previously shown to be present only in functional kinetochores, was invariably detected at the constriction of the marker in OSA, while one-fifth of markers in ALT appeared to exhibit additional centromere protein C-positive regions outside the primary constriction, indicating that the observed chromosomal instability in ALT might, at least in part, be a consequence of the occasional formation of more than one functional kinetochore. In OSA the alphoid DNA was coamplified with unique sequences from central 12q and the amplified material was C-band negative but in ALT amplified material from central 12q as well as sequences from proximal 12p were detected, resulting in C-band-positive areas. A propensity for additional kinetochore formation might thus be associated with the coamplification of alphoid DNA and pericentromeric sequences from chromosome 12. PMID- 10525964 TI - Tel2p, a regulator of yeast telomeric length in vivo, binds to single-stranded telomeric DNA in vitro. AB - The telomeres of the yeast Saccharomyces cerevisiae consist of a duplex region of TG(1-3) repeats that acquire a single-stranded 3' extension of the TG(1-3) strand at the end of S-phase. The length of these repeats is kept within a defined range by regulators such as the TEL2-encoded protein (Tel2p). Here we show that Tel2p can specifically bind to single-stranded TG(1-3). Tel2p binding produced several shifted bands; however, only the slowest migrating band contained Tel2p. Methylation protection and interference experiments as well as gel shift experiments using inosine-containing probes indicated that the faster migrating bands resulted from Tel2p-mediated formation of DNA secondary structures held together by G-G interactions. Tel2p bound to single-stranded substrates that were at least 19 bases in length and contained 14 bases of TG(1-3), and also to double stranded/single-stranded hybrid substrates with a 3' TG(1-3) overhang. Tel2p binding to a hybrid substrate with a 24 base single-stranded TG(1-3) extension also produced a band characteristic of G-G-mediated secondary structures. These data suggest that Tel2p could regulate telomeric length by binding to the 3' single-stranded TG(1-3) extension present at yeast telomeres. PMID- 10525965 TI - DNA replication during amplification of the C3 puff of Rhynchosciara americana initiates at multiple sites in a 6 kb region. AB - Two independent two-dimensional agarose gel electrophoresis methods have been used to map the origin of replication that directs amplification of the C3 DNA puff of Rhynchosciara americana. The results of neutral/neutral two-dimensional gel electrophoresis show that DNA replication initiates at multiple sites in a zone of at least 6 kb situated immediately upstream from the promoter of the main transcription unit of this puff. The complementary neutral/alkaline two dimensional gel electrophoresis technique shows that, within the initiation zone, forks move in both directions. In contrast, unidirectional fork movement away from the initiation zone is observed at the ends of the region, implying that it is the only place in the amplified region of the C3 puff where initiations occur. Since the initiation zone coincides with the region that is most highly amplified, amplification of the C3 puff probably occurs by an onion skin-type mechanism. PMID- 10525966 TI - Histochemical localization of reverse transcriptase in polytene chromosomes of chironomids. AB - The localization of a reverse transcriptase-related protein in salivary gland polytene chromosomes was investigated by immunohistochemistry in two species of Chironomus. The antibodies used were raised against a recombinant protein containing phylogenetically conserved motifs of reverse transcriptases and derived from an abundant non-LTR element previously identified in Chironomus. Immunoreactive protein was found in some telomeres, in a centromeric region, in a few interstitial bands and in Balbiani ring 3. The telomeric signal was probably dependent on transcription and increased dramatically when telomeric heat shock puffs were induced. A correlation with transcription was also seen in Balbiani ring 3, the immunobinding of which disappeared after inhibition of transcription with actinomycin D. PMID- 10525967 TI - Nucleo-cytoplasmic translocation of histone H1 during the HeLa cell cycle. AB - The distribution of unphosphorylated and phosphorylated isoforms of linker histone H1 protein was examined during the cell cycle of HeLa cells by quantitative light and electron microscopic immunocytochemistry. Immunolabeling with a monoclonal antibody directed against the globular domain of H1 (anti-H1), which recognized predominantly unphosphorylated H1, and a polyclonal antibody directed against hyperphosphorylated H1 (anti-H1P) revealed that: (1) H1 immunolabeling was lowest at the start of S phase (S(S)), and then increased progressively during the middle (S(i)) to end of S phase (S(e)), mitosis (M) and telophase (T) to reach the highest level in G1 phase, at which time there was a sudden reduction in H1 immunolabeling before the start of S phase; (2) H1P immunolabeling paralleled this progressive increase, but only until M phase, after which it abruptly disappeared and was virtually absent in G1; (3) H1P immunolabeling in S and M phase was found on both nuclear chromatin or chromosomes and in the cytoplasm, while H1 immunolabeling was found only on nuclear chromatin or chromosomes where it was predominantly localized on condensed chromatin. Our study indicates that H1 dissociates from the DNA to a large extent during replication and chromosome condensation, but not in interphase when cells are transcriptionally active. PMID- 10525968 TI - A family of dispersed repeats in the genome of Vicia faba: structure, chromosomal organization, redundancy modulation, and evolution. AB - A family of repeated DNA sequences of about 1200 bp in length and bordered by well-conserved, 18 bp inverted repeats (VfB family) was found in the nuclear genome of Vicia faba. The structure, chromosomal organization, redundancy modulation and evolution of these sequences were investigated. They are enriched in A+T base pairs (about 40% G+C) and lack any obvious internally repeated motif. A 64%-73% nucleotide sequence identity was found when pairwise comparisons between VfB sequences were carried out (average 69%). Direct repeats were not found to flank the inverted repeats that border these DNA sequences. The results obtained by hybridizing VfB repeats to Southern blots of V. faba genomic DNA digested with EcoRI indicated that these DNA elements are interspersed in the genome. The appearance of bands in these Southern blots and comparison of the structure of the sequences that flank different VfB elements showed that these repeats might be part of other, longer repeated DNA sequences. A high degree of dispersion throughout the genome was confirmed by cytological hybridization, which showed VfB sequences to be scattered along the length of all chromosomes and to be absent or rare only at heterochromatic chromosomal regions. These sequences contribute to intraspecific alterations of genomic size. Indeed, dot blot hybridizations proved that their redundancy, which is positively correlated with the overall amount of nuclear DNA in each accession, varies between V. faba land races (27x10(3)-230x10(3) copies per 1C DNA). Southern blot hybridization of VfB repeats to restriction endonuclease-digested genomic DNAs of V. faba, V. narbonensis, V. sativa, Phaseolus coccineus, Populus deltoides, and Triticum durum revealed nucleotide sequence homology of these DNA elements, whatever the stringency conditions, only to the DNAs of Vicia species, and to a reduced extent to the DNAs of V. narbonensis and V. sativa compared with that of V. faba. It is concluded that VfB repeats might be descended from mobile DNA elements and contribute to change genomic size and organization during evolution. PMID- 10525970 TI - Stroke prevention: which drugs to use and when? AB - This review summarizes recent findings from clinical trials regarding the prevention of stroke and translates these into therapeutic guidelines. A distinction is made between patients with previous cerebrovascular disease and those without, and between patients with and those without atrial fibrillation. Although the efficacy of aspirin is disappointingly small, the effects are consistent in all subgroups of patients with confirmed vascular disease, and this treatment remains superior as first choice except in patients with both atrial fibrillation and vascular risk factors, for whom oral anticoagulants are the optimal treatment. PMID- 10525969 TI - Nuclear organization studied with the help of a hypotonic shift: its use permits hydrophilic molecules to enter into living cells. AB - A new procedure for introduction of hydrophilic molecules into living cells based on efficient uptake of these molecules into the cells during hypotonic treatment is presented and its use is demonstrated by a variety of applications. Experiments with cultured vertebrate and Drosophila cells and various animal tissues demonstrated that the increase in cell membrane permeability under hypotonic conditions is a general phenomenon in all animal cells tested. The efficiency of the method depends on the composition and temperature of the hypotonic buffer, the duration of the hypotonic treatment and the molecular weight of the molecules introduced into living cells. The versatility of this approach is demonstrated with various types of molecules such as modified nucleotides, nucleotides with conjugated fluorochrome, peptides, phosphatase substrates and fluorescent dyes. The method opens new possibilities for the direct investigation of a variety of biological problems as documented here with data on the functional organization of the cell nucleus. PMID- 10525971 TI - Teaching course: ion channelopathies in neurology. PMID- 10525972 TI - A follow-up study of cognitive impairment due to inferior capsular genu infarction. AB - Abulia, memory loss, other cognitive deficits, and behavioral changes consistent with dementia can follow an inferior capsular genu infarction, but only little is known about the time course of these disturbances. The present study describes the long-term outcome of cognitive defects in four patients with inferior capsular genu infarction who underwent a neuropsychological examination within 3 and 12 months of onset. Three patients had infarcts in the inferior genu of the left internal capsule and had similar symptoms in the acute phase: disorientation, memory loss, language impairment, and behavioral changes. The patient with right-side infarct showed memory impairment and behavioral changes. Three patients had deficits in one or more cognitive domains on the first assessment, but none was demented. By the second evaluation all subjects had improved. In two patients there were a moderate memory defect persisted and a language disturbance. Improvement in these disturbances during long-time follow up demonstrates that there are alternative pathways that reestablish the functional connections damaged by the strategically located capsular genu infarct. Inferior capsular genu infarction is not a cause of persisting "strategic infarct dementia." PMID- 10525973 TI - Long-term follow-up of isolated optic neuritis: the risk of developing multiple sclerosis, its outcome, and the prognostic role of paraclinical tests. AB - We evaluated the risk of developing clinically definite multiple sclerosis (CDMS) after an acute attack of isolated optic neuritis (ON) in 112 patients, in relation to demographic and paraclinical findings. Patients were examined by brain MRI, CSF analysis, and multiple evoked potentials (EPs); 10 were lost to follow-up, and the other 102 were enrolled in a prospective study (follow-up duration 6. 3 +/- 2.2 years). Of these, 37 (36.3%) developed CDMS after a mean interval of 2.3 +/- 1.6 years. The risk of developing CDMS was 13% after 2 years, 30% after 4, 37% after 6, and 42% after 8 and 10 years. Gender, age, and season of ON onset did not affect the risk. MS occurred in 37 of 71 patients (52.1%) with one MRI lesion or more; no patient with a normal MRI developed the disease. MS developed more frequently in patients with intrathecal IgG synthesis than in those without (43% vs. 28%), but the difference was not statistically significant. Multiple EPs showed a slight predictive value only including somatosensory EPs of the lower limb. Multiple sclerosis was mild in most cases (EDSS 2.2 +/- 1.9). The EDSS was less than 4 in 32 cases (86%), between 4 and 6 in 2 (5%), higher than 6.5 in 3 (8%). PMID- 10525974 TI - Long-term recovery and fellow eye deterioration after optic neuritis, determined by serial visual evoked potentials. AB - Twelve optic neuritis patients (part of a larger group in whom the effects of intravenous methylprednisolone treatment were previously reported), were followed up 3 years from the onset of symptoms with visual evoked potentials (VEPs), contrast sensitivity and visual field examination. Findings from the previously "unaffected" eyes, none of which had had symptomatic optic neuritis, were also assessed. Between 6 months and 3 years after the onset of symptoms the VEPs of the affected eyes showed a significant shortening of mean latency (whole field, 131-123 ms; central field, 136-125 ms). Conversely, the responses of the previously unaffected eyes showed a contemporaneous latency prolongation (significant for the whole field, 110-113 ms) which exceeded the expected effect of aging. Contrast sensitivity tests showed no significant change in the affected eyes but a mild deterioration in the unaffected eyes, while the visual fields showed no overall pattern of improvement or deterioration. If the strong tendency for VEP latencies to shorten is due to ongoing remyelination, the lack of significant improvement in visual function may be because the visual deficit at 6 months is due to irreversible axonal loss rather than demyelination. The absence of functional deterioration in the affected eye, while VEPs and contrast sensitivity deteriorated in the unaffected eye, suggests that long-term remyelination may for a while counteract the effects of insidious demyelination and axonal degeneration which affect the visual pathway during clinical remission. PMID- 10525975 TI - Impairment of the supervisory attentional system in early untreated patients with Parkinson's disease. AB - The aim of this study was to specify the frontal type dysfunction widely reported in Parkinson's disease (PD) early in the course of the disease and before dopaminergic therapy. Seventeen "de novo" PD patients and 17 healthy control subjects performed modified versions of the Stroop word-color test and the Brown Peterson paradigm. A dissociation between results on the two tasks was observed in PD patients. They had difficulties in inhibiting a strong habitual response and establishing a new, better adapted pattern of response; but they performed as well as controls in a dual-task paradigm requiring correct allocation of the processing resources of working memory. Early in the course of the disease, untreated PD patients suffer from dysfunction of the supervisory attentional system. However, the present findings suggest that this system is not a single unit but rather could be composed of multiple subsystems whose sensitivity depends on the origin of frontal dysfunction. Indeed, only a few of these subsystems seemed to be impaired in de novo PD patients. It can be hypothesized that those involved in the phenomena of adaptation and consolidation of currently appropriate responses depend on the dorsolateral prefrontal loop, which is affected by the dopaminergic innervation of the caudate nucleus. PMID- 10525976 TI - Autosomal dominant cerebellar ataxia type I: oculomotor abnormalities in families with SCA1, SCA2, and SCA3. AB - Forty-six patients suffering from autosomal dominant cerebellar ataxia type I (ADCA I) underwent to a genotype-phenotype correlation analysis by molecular genetic assignment to the spinocerebellar ataxia type 1, 2, or 3 (SCA1, SCA2, SCA3) genetic locus and electro-oculography. Oculomotor deficits that are attributed to dysfunction of cerebellar structures occurred in all three mutations without major differences between the groups. Gaze-evoked nystagmus, however, was not found to be associated with SCA2. Square wave jerks were exclusively observed in SCA3. The gain in vestibulo-ocular reflex was significantly impaired in SCA3 and SCA1. In SCA3 the severity of vestibular impairment increased with CAG repeat length. Severe saccade slowing was a highly characteristic feature of SCA2. In SCA3 saccade velocity was normal to mildly reduced while SCA1 fell into an intermediate range. The present data show that each mutation is associated with a distinct syndrome of oculomotor deficits. Reduced saccade velocity and the absence of both square-wave jerks and gaze evoked nystagmus allow one SCA2 to be distinguished from SCA3 patients in almost all cases. The eye movement disorder of SCA1 patients, however, overlaps with both SCA2 and SCA3. PMID- 10525977 TI - Ipsilateral facial weakness in upper medullary infarction-supranuclear or infranuclear origin? AB - We describe two patients with upper medullary infarctions showing ipsilateral facial weakness and relative sparing of the upper facial muscles. Electrophysiological follow-up using transcranial magnetic stimulation of the motor cortex in combination with stimulation of the peripheral facial nerve disclosed a supranuclear (corticofacial) tract lesion in one patient and a partial nuclear/infranuclear intra-axial facial nerve lesion in another. PMID- 10525978 TI - A retrospective long-term analysis of the epidemiology and features of drug induced headache. AB - Drug-induced headache is well known to result from the abuse of compounds taken for the treatment of primary headache. The features of drug-induced headache depend on various features including the availability of drugs, the regional health system, and psychogenic factors of the patients. We performed a retrospective study on a series of 257 consecutive German patients presenting with drug-induced headache during the period 1983-1996. Our aim study was to evaluate the demographic features, the frequency of various drugs used, in particular of ergotamine derivates, and changes in these features during the study period. The frequency of drug-induced headache among all headache patients was 8%, with a female preponderance of 81%. Drug-induced headache occurred in all age groups, predominantly in migraine patients (35%). The mean number of substances used was 2.7, mainly, acetaminophen (47.9%), ergotamine tartrate (45%), and combined analgesics (56%). We did not find a significant difference between the associations with ergotamine tartrate and dihydroergotamine, although the latter was taken less frequently. Comparing the early and late years of our study period, there were no changes in the frequency of drug-induced headache (8% versus 7%), although changes in the frequency of some drugs changed (barbiturates, ergotamine tartrate, and codeine intake decreased whereas nonsteroidal anti-inflationary drugs, combined analgesics, and sumatriptan intake increased). Our data suggest that changes in drug availability and the introduction of classification criteria and treatment recommendations did not have a major impact on the frequency of drug-induced headache. PMID- 10525979 TI - The diagnostic accuracy of magnetic resonance imaging and cerebrospinal fluid cytology in leptomeningeal metastasis. AB - Diagnostic decision making in the case of patients suspected of having leptomeningeal metastasis (LM) can be very difficult. The results of cerebrospinal fluid (CSF) cytology can be repeatedly negative, and the predictive value of gadolinium-enhanced magnetic resonance imaging (MRI) is not well known. We report the results of CSF cytology and Gd MRI in 61 patients with known cancer, suspected of having LM. We combined our data with those from a similar study and calculated the sensitivity and specificity of CSF and Gd MRI, in the absence of a "gold standard diagnosis." CSF cytology was positive for LM in 35 patients and MRI in 38. With CSF cytology sensitivity 75% and specificity 100%, with Gd MRI sensitivity was 76% but specificity only 77%. We conclude that Gd MRI provides strong support in the diagnosis of LM in patients with cancer who have negative results on CSF cytology. PMID- 10525980 TI - Spinal epidural abscess complicating chronic epidural analgesia in 11 cancer patients: clinical findings and magnetic resonance imaging. AB - We reviewed the records of all patients who had received an epidural catheter for management of chronic cancer pain in a 3-year period (1993-1996). Patients with nervous system infections were identified, and pertinent clinical, radiological (magnetic resonance imaging), and bacteriological data were analyzed. We identified 11 patients who developed spinal epidural abscess (SEA). All of these had back pain; radicular signs occurred in seven patients and spinal cord compression in two patients. Magnetic resonance imaging revealed SEA in all 11 patients. SEA was iso- to hypointense on T1-weighted images and hyperintense on T2-weighted images relative to spinal cord. After gadolinium administration seven lesions showed characteristic rim enhancement while three showed minimal enhancement. No signs of diskitis or osteomyelitis were present, and the abscess was always localized to the posterior epidural space. Cultures were positive in all cases and revealed Staphylococcus epidermidis in eight and S. aureus in three. All patients were treated with intravenous antibiotics, and four had an additional decompressive laminectomy. Two patients died within 1 week of diagnosis from overwhelming septicemia despite apparently adequate antibiotic treatment. Within 4 weeks after diagnosis of SEA two patients died from widely metastatic disease, although infection may have contributed. One patient developed septicemia while receiving appropriate antibiotics and underwent emergency laminectomy. The neurological deficits recovered in all patients who survived the acute infectious episode. We conclude that patients with chronic epidural catheters for cancer pain require prompt neurological evaluation and magnetic resonance imaging when SEA is suspected. Early evaluation and treatment may lead to full recovery. PMID- 10525982 TI - Novel mutations in the muscle chloride channel CLCN1 gene causing myotonia congenita in Spanish families. AB - Mutations in the muscular voltage-dependent chloride channel gene (CLCN1), located at 7q35, lead to recessive and dominant myotonia congenita. We report four novel mutations identified in this gene, after clinical, electromyographic, and genetic studies performed on 13 unrelated families. Two of the four mutations (2512insCTCA and A218T) were identified in families with Thomsen's disease, one (Q658X) in a family with Becker's disease, and the fourth (R669C) in a presumably sporadic patient with the Becker phenotype. Although identification of the mutations allows us to establish some genotype/phenotype correlations, this does not wholly account for the clinical heterogeneity and the inheritance patterns of the disease. PMID- 10525981 TI - Promoter polymorphism (-491A/T) in the APOE gene of Finnish Alzheimer's disease patients and control individuals. AB - Apolipoprotein E (APOE) epsilon4 allele is a major risk factor for the development of Alzheimer's disease (AD). It has been suggested that the quantitative expression of APOE alleles results from mutations in the promoter region of this gene. We studied the -491A/T promoter polymorphism and whether it is dependent on the APOE epsilon4 allele in clinic-based AD (n = 106) and community-based control (n = 123) samples. The -491A/T and APOE polymorphisms were analyzed using the polymerase chain reaction method and restriction fragment length polymorphism analysis. The APOE epsilon4 allele was strongly associated with AD when compared with controls, P < 0.001 (odds ratio 5.85, 95% CI 3.29- 10. 41). The genotype distribution of the -491A/T polymorphism did not significantly differ between the study groups (P = 0.063), and the -491A allele was not associated with any significant risk in the AD group when compared to controls (odds ratio 1.82, 95% CI 0.95-3.49). However, haplotype estimation analysis indicated linkage disequilibrium between APOE -491A/T polymorphism and the APOE epsilon4 allele. Our findings confirm APOE polymorphism still to be the most efficient predictor of risk in AD. PMID- 10525983 TI - Cerebral hemorrhage and apoE. AB - The epsilon4 allele of apolipoprotein E (apoE) is found more commonly among patients with Alzheimer's disease (AD) than in the normal population. ApoE is associated with brain amyloid, a component of cerebral amyloid angiopathy (CAA), which is both a pathological feature of AD and a frequent cause of lobar intracerebral hemorrhage (ICH). We hypothesized that the frequency of epsilon4 allele is higher in patients with CAA-related ICH than in hypertensive ICH and in the normal population. To test this hypothesis we compared the frequency of apoE alleles in four populations: 24 patients with lobar ICH, 24 matched patients with hypertensive ICH, 24 matched normal controls, and 173 population controls. Although there was a tendency to a higher frequency of apoE epsilon4 in lobar ICH patients, we found no significant differences in the frequency of this allele between the four studied populations. In addition we did not confirm the finding of some authors of a higher frequency of apoE epsilon2 in patients with lobar ICH than in the normal population. Previous studies on the subject are discussed. The relationship between apoE polymorphism and lobar CAA-related ICH remains to be clearly defined. PMID- 10525984 TI - Mosaicism of unstable CAG repeats in the brain of spinocerebellar ataxia type 2. AB - Spinocerebellar ataxia type 2 (SCA2) is caused by expansion of unstable CAG repeats within the coding region of the novel gene, ataxin-2, on chromosome 12q24.1. We analyzed CAG repeat size of the SCA2 allele in two deceased patients (father and daughter) to investigate the repeat mosaicism in CNS regions. The CAG repeat size was examined using lymphoblastoid cell lines, frozen brain tissues, and paraffin-embedded tissues. In each patient the major repeat size of the expanded allele varied within the brain or spinal cord (father, 39-42; daughter, 39-47 repeats), and was smaller by three to eight repeats in the cerebellum than in other CNS regions. Our results are in agreement with the findings in other polyglutamine disorders showing somatic mosaicism. PMID- 10525985 TI - Machado-Joseph disease presenting as severe generalised dystonia in a German patient. PMID- 10525986 TI - The epileptogenic effect of tranexamic acid. PMID- 10525987 TI - A fourth ventricular papilloma presenting with transient autonomic failure. PMID- 10525988 TI - Visual evoked potentials and electroretinograms in an early stage of Leber's hereditary optic neuropathy. PMID- 10525990 TI - Nonhemorrhagic venous infarction of the spinal cord without spinal vascular malformation. PMID- 10525989 TI - Toxic leukoencephalopathy after intravenous consumption of heroin and cocaine with unexpected clinical recovery. PMID- 10525991 TI - Systemic lymphoma in patients with the initial diagnosis of primary central nervous system lymphoma: a report of two cases. PMID- 10525992 TI - Stiff-man syndrome with vacuolar degeneration of anterior horn motor neurons. PMID- 10525993 TI - Journal club PMID- 10525994 TI - Neuroscience news PMID- 10525995 TI - ENS news PMID- 10525996 TI - Clinical features differentiating patients with postmortem confirmed progressive supranuclear palsy and corticobasal degeneration. AB - Progressive supranuclear palsy (PSP) and cortocobasal degeneration (CBD) are often clinically confused with each other because they share a rapid disease progression, parkinsonism that responds poorly or transiently to levodopa therapy, and associated signs (e.g., ocular abnormalities, pyramidal signs and cognitive involvement). To improve the accuracy in diagnosing these disorders, this study examined the clinical features of 51 patients pathologically diagnosed with PSP and CBD. Logistic regression analysis identified two sets of predictors (models) for CBD patients, one consisting of asymmetric parkinsonism, cognitive disturbances at onset and instability and falls at first clinic visit, and the other one of asymmetric parkinsonism, cognitive disturbances at symptom onset and speech disturbances. While PSP patients often had severe postural instability at onset, symmetric parkinsonism, vertical supranuclear gaze palsy, speech and frontal lobe-type features, CBD patients presented with lateralized motor (e.g., parkinsonism, dystonia or myoclonus) and cognitive signs (e.g., ideomotor apraxia, aphasia or alien limb). On the other hand, CBD patients presenting with an alternate phenotype characterized by early severe frontal dementia and bilateral parkinsonism were generally misdiagnosed. PSP patients without vertical supranuclear gaze palsy were misdiagnosed. Recognizing the features which differentiate these disorders and the less obvious disease presentations as well as developing an increased index of suspicion will improve the diagnostic accuracy of these disorders. PMID- 10525998 TI - Parkinsonism-dementia complex on Guam - overview of clinical aspects. AB - Parkinsonism-dementia complex (PDC) is the second most common neurodegenerative disorder in Guam, after amyotrophic lateral sclerosis (ALS). PDC was first described by Hirano 1961. A familial appearance is seen among some PDC cases, which may also include ALS, and vice versa, but subsequent research including pedigree analysis, prospective case control registries, and the search for specific gene markers has failed to yield a satisfactory genetic explanation. Important diagnostic indicators of the illness include rigido-akinetic type Parkinsonism and severe dementia.In PDC, rigidity is so marked that postural deformities such as a generally flexed posture become rather prominent. Gait disturbances are a common initial symptom. Hyperreflexia and spinal muscular atrophy, developing mainly in the distal extremities, are frequently observed. These mixed-syndrome patients can be seen as clear support for the view that Guam ALS and PDC constitute a single mixed disease entity with a spectrum of clinical expression. The present paper offers an overview and description of the clinical features of PDC. PMID- 10525997 TI - Neuropathologic differentiation of progressive supranuclear palsy and corticobasal degeneration. AB - Progressive supranuclear palsy (PSP) and corticobasal degeneration (CBD) are usually sporadic multi-system degenerations associated with filamentous tau inclusions in neurons and glia. As such they can be considered sporadic tauopathies in contrast to familial tauopathies linked to mutations in the tau gene. Mutations have not been found in the tau gene in either PSP or CBD. The clinical syndromes and neuroimaging of typical cases of PSP and CBD are distinct; however, atypical cases are described that have overlapping clinical and pathologic features. Both PSP and CBD have similar biochemical alterations in the tau protein, with the abnormal tau protein containing predominantly four-repeat tau. While there is overlap in the pathology in PSP and CBD, there are sufficient differences to continue the present day trend to consider these separate disorders. Several important pathologic features differentiate PSP from CBD. Ballooned neurons are frequent and nearly a sine qua non for CBD, but they are found in PSP at a frequency similar to that of other neurodegenerative diseases, such as Alzheimer's disease. Astrocytic lesions are different, with tufted astrocytes found in motor cortex and striatum in PSP and astrocytic plaques in focal atrophic cortices in CBD. The most characteristic neuronal tau pathology in CBD is wispy, fine filamentous inclusions within neuronal cell bodies, while affected neurons in PSP have compact, dense filamentous aggregates characteristic of globose neurofibrillary tangles. Thread-like processes in gray and white matter are much more numerous and widespread in CBD than in PSP. The brunt of the pathology in CBD is in the cerebrum, while the basal ganglia, diencephalon and brainstem are the targets of PSP. Further clinicopathologic studies will refine our understanding of these disorders and open the possibility that common etiologic factors may be identified for these unusual sporadic tauopathies. PMID- 10525999 TI - Neuropathology of parkinsonism-dementia complex and amyotrophic lateral sclerosis of Guam: an update. AB - A comparative study was performed to investigate the differences and similarities of the neuropathological findings in the parkinsonism-dementia complex (PDC) and amyotrophic lateral sclerosis (ALS) of Guam, progressive supranuclear palsy and classic ALS. Based on the findings, it is proposed that (a) PDC is a discrete disease entity, (b) NFTs in Chamorro ALS are merely a background feature widely distributed in this population, (c) Chamorro ALS is a disease combined with classic ALS and neurofibrillary degeneration, (d) thus a subtype of "Guam ALS" is not present, and (e) PDC and ALS of Guam are different diseases. PMID- 10526000 TI - Posteroventral medial pallidotomy in Parkinson's disease. AB - There has been a resurgence in the use of functional neurosurgery for Parkinson's disease. An important factor that has played a role in this development is the recent understanding of the functional anatomy of the basal ganglia including a knowledge of the changes in the activities of neurons in the internal segment of the globus pallidus (Gpi) and the subthalamic nucleus (STN) in Parkinson's disease as well as the knowledge of the presence of segregated functional loops within the basal ganglia which include a sensory-motor loop that involves the posteromedial globus pallidus rather than the anterior GPi where earlier pallidotomy lesions had been made. Laitinen reintroduced the modern posteroventral medial pallidotomy (PVMP) in 1992. Since then it has become clear that this treatment has major effects on levodopa-induced dyskinesias and, unlike Vim thalamotomy, improves bradykinesia and rigidity as well as tremor. In this report, we review a number of topics related to PVMP including the clinical results of pallidotomy available in the literature as well as an update of our own 2 year follow-up data, studies evaluating factors that might predict the subsequent response to pallidotomy, the neuropsychological effects of the procedure, results of imaging studies including the correlation of clinical effects with lesion location, the question of bilateral pallidotomy and pallidotomy combined with deep brain stimulation and finally whether PVMP is effective in other parkinsonian disorders. PMID- 10526001 TI - Thalamic, subthalamic nucleus and internal pallidum stimulation in Parkinson's disease. AB - The limits of drug therapy in severe forms of Parkinson's disease have lead to a renewal of functional neurosurgery of the basal ganglia and the thalamus. Deep brain stimulation (DBS) of these structures was developed with the aims of reducing the morbidity of surgery and of offering an adaptative treatment. DBS was first applied to the thalamus in patients with severe tremor. Tremor of the hemibody is greatly reduced by stimulation of the contralateral electrode in 85% of the cases. There is little change in other symptoms. However, motor fluctuations and dyskinesias are a more frequent problem than severe tremor; in attempt to treat these symptoms, DBS has recently been applied to the subthalamic nucleus (STN) and the internal pallidum (GPi). STN stimulation greatly decreases off motor symptoms and motor fluctuations, which allows a reduction of drug dosage and consequently of dyskinesias. GPi stimulation decreases dyskinesias in most patients, but the effect on off motor symptoms is more variable from one series to another, from very good to nil. The severe morbidity of DBS applied to these 3 targets is low. Comparative studies of the cost and the efficacy of DBS and lesions applied to these different targets are now required. PMID- 10526002 TI - The nuclear matrix in pathology. AB - For a long time the molecular basis of nuclear structure has been a matter of debate rather than an established fact. In the last decade the concept of the nuclear matrix has emerged, and the molecular basis of this nuclear infrastructure, although still incomplete, is gradually being unravelled. In early studies concerning the nuclear structure, autoantibodies derived from patients with collagen disease had a significant role. This matrix, the structure remaining after extraction of membranes, nucleic acids and histones, consists of the nuclear lamina, the nucleolus and a fibrillogranular network. The nuclear lamina is composed of the lamins. The nucleolar matrix contains the proteins involved in rRNA processing. The fibrillogranular network is composed of nuclear matrix proteins, a wide variety of which has been discovered. It has become clear that the nuclear matrix not only provides a structural basis for nuclear architecture but also plays a part in regulating nuclear function. Lamins provide mechanical continuity between cytoskeleton and nuclear interior. Aberrant patterns of lamin expression have been described in cancer; these are not sufficiently specific to be used in histopathological diagnosis, however. Nucleolar size and expression levels of nucleolar proteins have been shown to correlate well with proliferative activity, which may revitalize interest in nucleolar organizing regions as a tool in histological diagnosis of cancer. The fibrillogranular network is involved in functional compartmentalization of replication and transcription. A variety of nuclear matrix proteins has been described, which appear to be specifically expressed in cancer cells. Analysis of expression of these proteins might play a significant role in cancer diagnosis. PMID- 10526003 TI - Primary natural killer/T-cell lymphomas of the oral cavity are aggressive neoplasms. AB - Thirty-four cases of primary non-Hodgkin's lymphoma of the oral cavity were investigated for their clinical findings, histopathological features, immunophenotypes and association with Epstein-Barr virus (EBV). Four cases (12%) were natural killer/T-cell lymphomas, 3 (9%) were T-cell lymphomas and 27 (79%) were B-cell lymphomas. Compared with T- and B-cell lymphomas, NK/T-cell lymphomas had a male predominance (M:F 4:0), and most presented as ulceration of the palate and/or maxillary gingiva. Histologically, the lesions showed diffuse infiltration of medium-sized or large lymphoid tumour cells. Angiocentricity and/or angioinvasion were found in all 4 cases. The immunophenotypes of the NK/T-cell lymphomas were CD3+, CD43+, CD45RO+, CD56+ and TIA-1+. EBV was detected in 2 NK/T cell lymphomas by in situ hybridization (ISH) and polymerase chain reaction (PCR) methods, and was not detected in T- and B-cell lymphomas. The survival rate of patients with NK/T-cell lymphoma was zero, but the survival rates for patients with T-cell and B-cell lymphomas were 67% and 38%, respectively. It appears that NK/T-cell lymphomas of the oral cavity have a predilection for originating in the palate and maxillary gingiva and are aggressive neoplasms. EBV positivity might be associated with more aggressive behaviour. PMID- 10526004 TI - Prognostic significance of p53 gene mutations and p53 protein expression in synovial sarcomas. AB - Alterations to p53 seem to be of prognostic significance in soft tissue sarcomas, but their significance for synovial sarcomas has not been studied. We analysed 34 synovial sarcomas in 19 patients for p53 alterations (p53 gene mutations + p53 immunopositivity) and examined this factor for its prognostic value in a group of 15 primary tumours. DNA was prepared from paraffin-embedded tumour material by a modified proteinase K/phenol/chloroform extraction. p53 gene mutations of exons 5 8 were analysed by the PCR-SSCP-sequencing method. p53 protein expression was evaluated by immunohistochemistry using the murine monoclonal antibody DO1. We found two missense mutations (5.9%) and ten p53 immunopositive cases (29.4%). Both tumours with p53 mutations showed p53 protein expression. There was no significant correlation between p53 alteration and histological subtype, age, sex, or tumour size. The 5-year survival rate was 24.1%. Overall survival was significantly reduced in patients having synovial sarcomas with p53 alterations (P<0.001). In the multivariate Cox's analysis, only p53 alterations (P=0.032) and tumour size (P=0.023) emerged as independent prognostic factors. We suggest that p53 alterations may be a useful prognostic indicator in synovial sarcomas, allowing rational clinical treatment and follow-up. PMID- 10526005 TI - Atypical cystic lobules: an early stage in the formation of low-grade ductal carcinoma in situ. AB - Evidence from many studies has established the neoplastic potential of ductal carcinoma in situ, but the origin and the morphological characteristics of the early stages of this proliferation remain unidentified. Workers writing in the early twentieth century observed a cystic transformation of lobules and proposed that it represented one such early stage, and contemporary European and Japanese pathologists have reached the same conclusion. We describe the characteristics of this cystic transformation, which we call us "atypical cystic lobules," and present evidence to support the proposal that the alteration is a step in the formation of low-grade ductal carcinoma in situ. Atypical cystic lobules are a proliferation of luminal cells showing low-grade cytological atypia without architectural atypia. The study group comprised 21 cases of atypical cystic lobules from specimens also showing conventional low-grade ductal carcinoma in situ or lobular neoplasia. Immunohistochemical staining for hormone receptors, keratin 19, and cyclin D1 revealed that atypical cystic lobules demonstrated a consistent immunophenotype, which differs from the pattern shown by normal lobules and benign lesions and matches that of low-grade ductal carcinoma in situ. In about 40% of the cases, atypical cystic lobules merged with fully established micropapillary/cribriform ductal carcinoma in situ. The similarities in the cytological and immunohistochemical features and the proximity of the two types of proliferation suggest that atypical cystic lobules represent an early stage in the formation of certain types of low-grade ductal carcinoma in situ. PMID- 10526006 TI - Glomerular expression of cell-cycle-regulatory proteins in human crescentic glomerulonephritis. AB - To elucidate the mechanism underlying crescentic formation, we assessed the phenotypic characterization and cell-cycle protein expression in human crescentic glomerulonephritis (CRGN). Kidney tissue specimens taken from CRGN patients (10 patients with pauci-immune type rapidly progressive glomerulonephritis (RPGN), 2 patients with Henoch-Schonlein purpura nephritis, and 1 patient with IgA nephropathy) were examined immunohistochemically. Most of the cellular components of the crescents expressed cytokeratin, whereas few cells expressed PHM-5. CD68 positive cells were minor components of cellular crescents, indicating that the major principal cellular component of the crescents is made up of cells with the parietal glomerular epithelial cell (PEC) phenotype. Additionally, serial section analysis revealed that Ki-67-positive cells in the crescents were frequently cyclin-A positive and Bcl-2 positive, but seldom cyclin-B(1) positive. Moreover, the expression of cyclin-dependent kinase inhibitor p27(Kip1) was low in the cellular crescents, despite being exclusively positive in podocytes within the same section. We concluded that the major component of the cellular crescents is made up of PECs and that apparent expression of cyclins and Bcl-2 and restrained expression of p27(Kip1) may be synergistically associated with the development of cellular crescents in human CRGN. PMID- 10526008 TI - Enhanced expression of EGF receptor and low frequency of ras mutations in X-ray induced rat thyroid tumours. AB - Radiation is recognized as a carcinogenic factor for the thyroid gland. In this experimental study, oncogene expression was investigated in radiation-induced rat thyroid tumours. Forty 3-month-old Wistar rats received X-ray-irradiation to the neck region; 40 animals were untreated controls. After 14 months, thyroid tumours had developed in 25 of the 29 irradiated animals still alive; 76% of these tumours were considered malignant. No tumours developed in controls. Mutations of codons 12-13 and 59-63 of H-, K- and N-ras were analysed by PCR-SSCP (single strand conformation polymorphism analysis) and sequencing of DNA from thyroid tissue. SSCP indicated a ras mutation frequency of 8%, but only one K-ras codon 12 (Gly-Cys) mutation was confirmed by sequencing. Protooncogene expression was analysed by mRNA slot blot hybridization analysis and immunohistochemistry. K-ras mRNA expression and EGF receptor mRNA and protein expression were significantly increased in the irradiated animals compared with controls, and in tumours versus nontumour tissue. This study of radiation-induced rat thyroid tumours demonstrates that ras expression may be subject to changes apart from activating mutations. Increased expression of EGF receptor in the tumours parallels the situation in human thyroid cancer. PMID- 10526007 TI - Immunohistochemical demonstration of oncocytes in nongonadotrophic pituitary adenomas. AB - An immunohistochemical study to demonstrate oncocytes in nongonadotrophic pituitary adenomas was performed. The adenomas were 10 prolactinomas, 2 ACTH producing adenomas (ACTHomas), and 28 growth hormone-producing adenomas (GHomas); we also studied 5 pituitary oncocytomas. GHomas were divided into two groups: GHomas with (GHomas-1) and without (GHomas-2) fibrous bodies. A small number of solitary large cells showed intense cytoplasmic granular reactivity for mitochondrial protein and cytochrome oxidase, resembling oncocytes in oncocytomas. The proportions of the mitochondrial protein-positive cells ranged from zero to 2.1% (0. 3+/-0.4%). They were more frequent in GHomas, GHomas-1 in particular, than other types of adenomas (P<0.01), and were mostly negative in prolactinomas and ACTHomas. In multivariate analysis, the proportions showed positive correlation with age (P<0.01) and the Ki-67 (MIB-1) labeling index (P<0.01) and tended to increase in number with recurrence (P<0.05). In GHomas, these cells were more common in cases with low basal GH level (P<0.01) and large tumor volume (P<0.01). We consider that these cells represent oncocytes existing in varying numbers in adenomas. We suggest that oncocytic change in nongonadotrophic adenomas indicates poor differentiation and/or some aggressiveness, which lead to a decrease in the endocrine activity of the tumor. PMID- 10526009 TI - "Composite" lymphoma, lymphoplasmacytoid and diffuse large B-cell lymphoma of the spleen: molecular-genetic evidence of a common clonal origin. AB - We describe here the first well-characterized case of "composite" lymphoma of the spleen in which the two components were a low-grade and a high-grade B-cell non Hodgkin's lymphomas. The patient was an elderly man with prominent splenomegaly and multiple hypoechogenic lesions of the spleen. A splenectomy was performed, and the macroscopic and histological findings showed the simultaneous presence of a "low-grade" B-cell lymphoma, lymphoplasmacytoid (immunocytoma) and a "high grade" B-cell lymphoma (immunoblastic), which were spatially separated. The two lesions expressed the same immunoglobulin light chain (lambda), but the Southern blot analysis showed different patterns of immunoglobulin heavy chain (IgH) clonal rearrangement. PCR analysis followed by direct sequencing of the IgH amplified rearrangement products provided molecular-genetic evidence that the two components of the composite lymphoma had the same clonal origin. Since both EBV LMP-1 and p53 were negative by immunohistochemistry, it is unlikely that EBV and p53 were involved in the neoplastic progression in this case. PCR analysis and direct sequencing of IgH-amplified rearrangement products are useful tools to investigate clonality in cases in which Southern blot analysis cannot be performed or does not provide conclusive findings. PMID- 10526010 TI - Granulocytic sarcoma of the thymus in a nonleukaemic patient. AB - We report a case of granulocytic sarcoma arising from the thymus in a 17-year-old nonleukaemic patient. The patient presented with an anterior mediastinal tumour and underwent surgical resection. Histological examination showed a diffuse infiltrate of immature round cells in the thymus. Tumour cells were diffusely peroxidase positive, but naphthol AS-D chloroacetate esterase negative. Immunohistochemical staining revealed expression of CD34 and terminal deoxynucleotidyl transferase (TdT), but not of CD13 and CD33. Ultrastructurally, electron-dense or medium-density granules were present in the cytoplasm. Four months after successful autogenic bone marrow transplantation, pleural and pericardial fluid contained tumour cells with azurophilic granules, which expressed CD13 and CD33, but not CD34 and TdT. The patient died of the disease 18 months after clinical manifestation, but still without developing leukaemia. The granulocytic sarcoma in the present case may have originated from myeloid precursors in the thymus and remained within the extramedullary site despite the differentiation into a more committed myeloid lineage at the relapse. PMID- 10526011 TI - 'Pyloric gland-type adenoma' arising in heterotopic gastric mucosa of the duodenum, with dysplastic progression of the gastric type. AB - 'Pyloric gland-type adenoma' is a recently described and very rare entity. We report a case of a pedunculated polyp of the duodenal bulb showing the features of pyloric gland-type adenoma. Heterotopic gastric mucosa was found adjacent to the tumour. Immunohistochemically, the tumour cells at the surface of the polyp showed foveolar-type mucin (M1) while most other tumour cells showed deep gastric mucin (M2), displaying a pattern of differentiation similar to the normal gastric mucosa. The polyp also showed villous or papillary structures with disorganization of gastric differentiation and marked increase of proliferating in foci cells. This is the first case of pyloric gland-type adenoma found to arise in heterotopic gastric mucosa of the duodenum, showing dysplastic progression of the gastric type. PMID- 10526013 TI - Reply PMID- 10526012 TI - Simple mucin-type carbohydrate antigens in Helicobacter pylori-positive chronic active gastritis. PMID- 10526014 TI - Sodium transport-related proteins in the mammalian distal nephron - distribution, ontogeny and functional aspects. AB - The mammalian distal nephron plays a pivotal role in adjusting urinary sodium excretion. Successive portions of the renal tubule are formed to adapt to this function, and an axial heterogeneity of the distal segments has been defined. The specific transport properties of these epithelia are accomplished by the expression of proteins (cotransporters, exchangers, channels) governing the movement of ions on either cell side. Molecular cloning of these proteins has had a marked impact on the study of their localization and function in the healthy and diseased kidney. Electroneutral cation-chloride cotransporters [Na(K)CC] have been localized to the thick ascending limb and the distal convoluted tubule using specific probes. Proteins implicated in the function of aldosterone target cells, such as the epithelial Na(+) channel (ENaC), the mineralocorticoid receptor (MR) and 11beta-hydroxysteroid dehydrogenase type 2 (11HSD2), an enzyme that confers mineralocorticoid specificity, have been found in the terminal portion of the nephron and the collecting duct. A mineralocorticoid-sensitive component of thiazide-sensitive NaCl transport has been identified in the distal convoluted tubule. Analysis of the ontogeny of these proteins in the maturing kidney has provided a detailed picture of epithelial differentiation and morphological specialization of the renal tubule. The study of mutations of the proteins related with NaCl transport has led to the identification of the molecular causes of inherited human diseases associated with hypo- or hypertension, and the respective sites of an impaired ion transport could be mapped to the renal tubule. PMID- 10526015 TI - Chronological study of the appearance of adenohypophysial cells in the ayu (Plecoglossus altivelis). AB - We previously reported the chronological appearance of adenohypophysial cells in freshwater teleosts using an immunocytochemical technique. The present study investigated the chronological appearance of adenohypophysial cells in the ayu, which is spawned and has its early development in brackish water, and the results were compared with those obtained in freshwater and seawater teleosts, as well as in other vertebrates. In the adult teleostean adenohypophysis, seven or eight types of secretory cells have been distinguished, each of which produce different hormones: prolactin (PRL), growth hormone (GH), thyroid stimulating hormone (TSH), gonadotropic hormones (GTH I and GTH II), adrenocorticotropic hormone (ACTH), melanophore stimulating hormone (MSH) and somatolactin (SL). In the pituitary of adult ayu, seven distinct types of glandular cells (PRL, GH, TSH, GTH, ACTH, MSH and SL cells) were identified. Chronologically, a few immunoreactive (ir)-PRL and ir-GH cells appeared in the ventral side of the pituitary one day before hatching. Then, just after hatching, ir-GTH cells were observed in the central to dorsal portion; ir-ACTH cells were found distributed in the anterior portion and some ir-MSH and a few ir-SL cells were seen in the posterior portion of the pituitary. Finally, a small number of ir-TSH cells were identified 50 days after hatching. These results differed from those obtained in other fishes previously reported with regard to the times of appearance of the PRL and GH cells. PRL cells appeared first, followed by GH cells in the freshwater teleosts, PRL and GH cells appeared at the same time in the brackishwater teleosts, while GH cells appeared first and PRL cells appeared last in the seawater teleosts. These results reflect the fact that PRL plays a major role in osmoregulation among freshwater teleosts, as compared with GH, which plays a similar role in seawater teleosts. It seems that both PRL and GH may play important roles in osmoregulation in brackishwater fish. PMID- 10526016 TI - Fine structure of dental epithelial cells and the enameloid during the enameloid formation stages in an elasmobranch, Heterodontus japonicus. AB - The structural features of the dental epithelial cells and the enameloid in tooth germs of the Japanese Port Jackson shark, Heterodontus japonicus, in the stages of enameloid formation, were investigated by light and transmission electron microscopy. At the enameloid matrix-formation stage, tall columnar inner dental epithelial cells contained large numbers of glycogen particles. At the enameloid mineralization stage, when many sharply outlined crystals appeared throughout the enameloid, the inner dental epithelial cells exhibited well-developed Golgi apparatuses and many mitochondria in the proximal cytoplasm, and abundant vesicles and vacuoles in the distal cytoplasm. Marked interdigitations of the lateral membrane were visible in the inner dental epithelial cells. The outer dental epithelial cells contained many mitochondria, lysosomal bodies, vesicles and microtubules, and the capillaries usually approached the outer dental epithelial cells. At the enameloid maturation stage, large numbers of crystals occupied the enameloid, and most of the organic matrix had disappeared from the enameloid area after demineralization. The organelles in the inner and outer dental epithelial cells decreased in number, but there were still widely distributed Golgi apparatuses, abundant intermediate filaments and granules containing an electron-dense substance in the inner dental epithelial cells. It is probable that the dental epithelial cells are involved in the removal of organic matrix from the enameloid and in the process of mineralization at the later stages of enameloid formation, i.e., the mineralization and the maturation stages. PMID- 10526017 TI - Neuronal and glial cell types revealed by NADPH-diaphorase histochemistry in the retina of a teleost fish, the grass goby (Zosterisessor ophiocephalus, Perciformes, Gobiidae). AB - The grass goby is a mud-burrowing fish with a rich retinal vasculature appropriate to its hypoxic habitat. NADPH-diaphorase histochemistry was performed on retinal sections and wholemounts to reveal cells that contain nitric oxide synthase and so may be presumed to synthesise nitric oxide, a gaseous intercellular messenger with many roles including vasodilation. Structures that were consistently stained by this method included cone ellipsoids, horizontal cells, Muller cells and their processes, large displaced ganglion cells in the inner nuclear layer (identified by their axons), large interstitial ganglion cells in the inner plexiform layer, and capillary endothelial cells. In wholemounts, horizontal cells were seen to form a regular pattern, contacting each other at their dendritic terminals. Some cells in the ganglion cell layer were weakly stained, but stained bipolar and amacrine cells were not seen. The diaphorase-positive large ganglion cells all formed large, sparsely branched dendritic trees, arborizing near the scleral border of the inner plexiform layer. The displaced and interstitial cells seemed to belong to distinct morphological types, the interstitial cells having smaller somata and trees. Analysis of their spatial distributions in one representative retina confirmed this: the displaced cells formed a highly regular mosaic with a mean spacing (nearest-neighbour distance) of 303 microm, whereas the interstitial cells formed a separate mosaic, almost as regular but with a smaller mean spacing of 193 microm, rising to 217 microm in a sample that excluded the area retinae temporalis. Spatial correlogram analysis showed that these two mosaics were spatially independent. Nitric oxide probably has many roles in the retina. The presence of its synthetic enzyme in Muller cells, which communicate with retinal blood vessels, is consistent with a role in the control of retinal blood flow. Its function in large, mosaic-forming retinal ganglion cells is unknown. PMID- 10526018 TI - Immunocytochemical mapping of NPY and VIP neuronal elements in the cat subcortical visual nuclei, with special reference to the pretectum and accessory optic system. AB - The aim of this study was to describe the distribution patterns of neuropeptide Y (NPY) and vasoactive intestinal polypeptide (VIP)-immunoreactive (ir) neuronal elements in subcortical visual centers of the cat. Numerous NPY-ir neurons were present in the feline nucleus of the optic tract and in the anterior pretectal nucleus. Only a few NPY-ir neurons were found in the posterior, medial and olivary pretectal nuclei and in the accessory optic nuclei. Diffuse and heavily beaded NPY-ir fiber plexuses were observed throughout the superior colliculus, pretectum, and accessory optic system. Extensively arborising NPY-ir fibers were present in the mesencephalon and ventral lateral geniculate nucleus, while the dorsal visual thalamic nuclei contained only a few NPY-ir fibers. VIP-ir cells were present mainly in the accessory optic nuclei, and they were absent in the dorsal visual thalamus. Both NPY- and VIP-ir neurons were multipolar and fusiform in shape in the regions studied. Enucleation did not alter the appearance of NPY- and VIP-containing neuronal elements in the superior colliculus and pretectum while in the thalamus a subset of NPY-ir fiber population disappeared, indicating their retinal origin. Although there is a partial overlap in the topographical localization of the NPY- and VIP-ergic neurons in the pretectum, the colocalization of the two peptides could not be demonstrated. The present observations demonstrate the existence of two different and separate peptidergic (NPY and VIP) neuronal populations in the pretectum. PMID- 10526019 TI - Programmed cell death and the morphogenesis of the hindbrain roof plate in the chick embryo. AB - The spatial and temporal distribution of apoptosis in the dorsal midline of the developing chick hindbrain was examined in relation to the development of the neuroepithelium and neural crest using scanning and transmission electron microscopy, immunocytochemistry and in situ hybridization. The pattern of TUNEL labeling and Slug expression in the dorsal midline at stages 10 and 11 differed from that at stages 12-15. At stages 10 and 11, TUNEL labeling and Slug expression were observed in the dorsal part of location II of rhombomere 1/2 (i.e., between the surface ectoderm and the neuroepithelium), but from stage 12 onward, they were observed in both the dorsal and ventral parts of location II. The implication is that whereas apoptosis may be restricted to a subpopulation of the early migrating neural crest at stages 10 and 11, it presumably occurs in subpopulations of both neural crest and neuroepithelial cells from stage 12 onward. Furthermore, as judged by the pattern of TUNEL labeling and Slug expression in r3 and r5, apoptosis in these two rhombomeres likely occurs in subpopulations of both neural crest and neuroepithelial cells. The eminence present in location I of r1/r2 between stages 10 and 12 consisted of both neural crest and neuroepithelial cells. These cells gradually underwent apoptosis until stage 12, when the eminence disappeared in most embryos. The formation of the inner (neuroepithelial) aspect of the hindbrain roof plate involved both cell migration from adjacent neuroepithelium and an alteration in the shapes of the cells, such that cells with flattened surfaces eventually lined the roof plate. During these processes, some of the neuroepithelial cells underwent apoptosis (i.e., in location IV). The results of this study thus demonstrate that subpopulations of both neuroepithelial and neural crest cells may be involved in programmed cell death in the hindbrain. Additionally, apoptosis in the hindbrain contributes significantly to morphogenetic thinning during roof plate formation. PMID- 10526020 TI - Does the perireticular thalamic nucleus project to the neocortex? AB - This study defines several features of the early connections of the developmentally transient perireticular thalamic nucleus of rats. The neocortex of developing rats was injected with either DiI, biotinylated dextran, WGA-HRP (wheatgerm agglutinin conjugated-horseradish peroxidase), fluorescent latex beads or cholera toxin subunit B (CTB) and their brains were processed for tracer detection with standard methods. In general, tracer injections into various regions of the developing neocortex revealed no labelled neurones within the perireticular nucleus, although some of these tracers (WGA-HRP, dextran) labelled many of the amoeboid microglial cells that are found within this nucleus. There were, however, many retrogradely labelled neurones in a region adjacent to the perireticular nucleus, within the nucleus basalis of the basal forebrain (medial edge of globus pallidus). Their identity was confirmed as neurones of the nucleus basalis since they were all were similar in morphology and somal size to neurones that were immunoreactive to NGFr (nerve growth factor receptor), an antigen found only among neurones of the nucleus basalis and basal forebrain. Moreover, double labelling experiments revealed that most, if not all, of the cortically labelled neurones were NGFr-immunoreactive also. Thus, in conclusion, our results suggest that the perireticular nucleus does not project to the neocortex; the only neurones in the general vicinity of the perireticular nucleus that have a cortical projection form part of the nucleus basalis. PMID- 10526021 TI - Prenatal development of coronary arteries in the rat: morphologic patterns. AB - The aim of this work was to address spatiotemporal and morphologic patterns of coronary artery development in rats, based on immunohistochemical and ultrastructural studies of hearts at different stages of prenatal development. Griffonia simplicifolia I lectin and alpha-smooth muscle antibody were used to demonstrate endothelial cells and/or their precursors and smooth muscle cells, respectively. Ultrastructural examination was performed on ED14-16 hearts to study the morphology of the developing coronary arteries in different regions of the truncus arteriosus and adjacent myocardium. On ED14 endothelial-like cells present within the mesenchyme surrounding the outflow tract penetrated the aortic wall and the truncoconal proximal myocardium. On ED15 these penetrating cells formed vascular clusters, which were the first signs of presumptive vascular channels. Development of the coronary artery proceeded by coalescence of discontinous vascular clusters, formation of the lumen (vascular channels) and establishing a connection of the proximal part with the aorta. The second layer of cells around vascular channels (embryonic media) consisted of mesenchymal cells that were attracted to the immature vessel and were first seen on ED15. At this time no lumenized connection of the coronary artery with the aorta has been seen. After the lumenized connection of the coronary artery with the aorta had been established perivascular cells of the media started to differentiate into vascular smooth muscle, as was shown by alpha-smooth muscle actin-staining. Further development and differentiation of the media and adventitia proceeded distally (towards the apex). PMID- 10526022 TI - Lectin histochemistry of the esophagus in several mammalian species. AB - The mucosa of the esophagus consists of stratified squamous epithelium that has a considerable resistance to injury. Intercellular glycoconjugates appear to constitute a major permeability barrier in the superficial portion of the esophageal mucosa. In the present study, we used a panel of lectins to investigate the differences in glycoconjugate production among different mammalian species. A battery of 12 lectins was used to study binding in sections from the esophagus of 6 mammalian species, including man. In general, the strongest staining was obtained in the stratum superficiale and the weakest staining in the stratum germinativum. In rabbit esophagus, exposure to pepsin/HCl produced a superficial damage to the epithelium, a considerable decrease in electrical resistance and a decreased staining of the esophageal epithelium with selected lectins. Pretreatment of the esophageal mucosa with sucrose octasulfate, a compound with protective properties, prevented, to some extent, the decrease in resistance and lectin staining. PMID- 10526024 TI - Editors' announcement PMID- 10526023 TI - Subretinal macrophages in the developing eye of eutherian mammals and marsupials. AB - Blood-borne mononuclear cells invade the developing retina via the hyaloid vasculature at the optic nerve head. Following removal of apoptotic cell debris they give rise to the network of resident microglia. The population of cells recently described in the peripheral subretinal space of developing human eyes may represent a further population of macrophages destined to become microglia. The aim of the present study was to confirm the presence of subretinal macrophages in the developing eye in other mammalian species and perform preliminary immunophenotypic analysis in rat tissues. The range of species chosen included eutherian mammals (rat and rabbit) and marsupials (wallaby and opossum). Ocular tissues from a range of developmental stages were studied by scanning electron microscopy and transmission electron microscopy. Distinctive networks of dendriform and pleomorphic macrophages were observed by scanning electron microscopy in the peripheral subretinal space of D2 rabbits, newborn and D2 rats and D75 wallaby. Transmission electron microscopic studies of D2 rabbit, newborn and D2 rat and all ages of North American opossum revealed cells with the ultrastructural features of macrophages in the peripheral subretinal space, cilio retinal junction and between ciliary epithelial cells. Preliminary immunoperoxidase studies using a panel of anti-leukocyte monoclonal antibodies on frozen sections of rat ocular tissues (newborn, D2 and D4) revealed ED1(+) Ox42(+) ED2(+) but Ox6(-) cells in the peripheral subretinal space, peripheral retina and ciliary body epithelia. The data confirms that subretinal macrophages are a feature of the developing eye in a broad range of mammalian species and immunophenotypic evidence leads the author to postulate that these cells arise from the ciliary body vasculature and may migrate into peripheral neural retina and mature into resident microglia. PMID- 10526025 TI - E.A.E.S. multicenter prospective randomized trial comparing two-stage vs single stage management of patients with gallstone disease and ductal calculi. AB - BACKGROUND: The current management of patients with gallstone disease and ductal calculi consists of endoscopic stone extraction (ESE) followed by laparoscopic cholecystectomy (LC). Following the advent of techniques of laparoscopic ductal stone clearance, an alternative single-stage laparoscopic treatment was introduced for these patients. The European Association of Endoscopic Surgery (E.A. E.S.) set up a ductal stone trial to compare the relative efficacy and outcome of these two management options. METHODS: A prospective randomized controlled clinical trial compared two management options. Group A (n = 150) received preoperative endoscopic retrograde cholangiography (ERC) with ESE followed by LC during the same hospital admission, and group B (n = 150) received single-stage laparoscopic management. RESULTS: There were no significant differences between the two groups in the clinical demographic details and the pretreatment biochemical findings. In group A, 14 of 150 patients received single stage treatment; in group B, 17 of 150 were managed by the two-stage approach (protocol violation = 31/300, 10%). In group A patients managed in accordance with randomization, ERC was successful in 129/136 (95%) and preoperative ESE succeeded in 82/98 (84%) with ductal calculi detected by the ERC. Two patients had malignancies and one refused surgery. Thus, 133 patients underwent surgery. Of this group, 116 had LC only and 17 had LC and attempted laparoscopic duct exploration. There were eight conversions to open surgery (6%), 17 complications for both stages (12.8%), and two postoperative deaths (1.5%). In group B patients managed in accordance with randomization, intraoperative cholangiography was successful in 132/133 (99%). Twenty-one (16%) had normal findings, ductal calculi were found in 109, and other pathology was noted in two (periampullary cancer, severe pancreatitis). These two patients and one other (who had gross adhesion in the triangle of Calot) were converted at the start of the procedure. Transcystic ductal stone clearance was successful in 45 of 56 patients (80%), and laparoscopic direct common duct (CBD) exploration was successful in 47 of 55 patients (85%). This group includes 53 patients who underwent primary direct exploration and two failed attempts at transcystic extraction. The conversion rate was 13%. Postoperative complications were encountered in 21 patients (15.8%), and one patient died of a major myocardial infarction (0. 75%). The one postoperative death and the 10/11 biliary complications occurred in the laparoscopic supraduodenal CBD exploration subgroup. The conversion rate was higher in group B (17 vs eight; p = 0.08). Laparotomy in the postoperative period was required in three patients in group A and four patients in group B. The group B patients were in hospital for 3 days less than patients who had two-stage management (median, 6.0, IQR = 4.25-12 vs median, 9.0, IQR = 5.5-14; p < 0.05). CONCLUSIONS: The results demonstrate equivalent success rates and patient morbidity for the two management options but a significantly shorter hospital stay with the single-stage laparoscopic treatment. The findings indicate that in fit patients (ASA I and II), single-stage laparoscopic treatment is the better option, and preoperative ESE should be confined to poor-risk patients-i.e., those with cholangitis or severe pancreatitis. PMID- 10526026 TI - Bursting strength evaluation after different types of mesh fixation in laparoscopic herniorrhaphy. AB - BACKGROUND: In laparoscopic inguinal herniorrhaphy, meshes commonly have been fixed with a stapler. Recently, a new mode of fixation using a helical fastener has been introduced. The purpose of this experimental study was to compare the stability achieved by various types of mesh fixation. METHODS: In 20 human cadavers, polypropylene meshes 10 x 15 cm in size were fixed in both groins by using either a helical fastener or a hernia stapler (4.8 mm). The mesh was fixed with 2, 4, and 8 elements and stressed with a dynamometer until the prosthesis ruptured. A paired and two-sided Student's t-test was used for statistical evaluation. RESULTS: With the helical fastener, the mesh could be fixed always at the desired site. However, with the stapler, it was not possible to fix the mesh in the pubic bone or, at times, in the Cooper's ligament. When two fixation elements were used, the mesh fixed by the helical fastener was able to withstand a median load of 34 N (range 23-53 N), and that fixed by the stapler 7.5 N (range 3-12 N; p < 0.001). When four fixation elements were used, the mesh fixed by the helical fastener was able to withstand 70.5 N (range 53-80 N) and that fixed by the stapler 17. 5 N (range 4-25 N; p < 0.001). With the use of eight elements, the mesh fixed by the helical fastener withstood 127 N (range 84-156 N) and that fixed by the stapler 32.5 N (range 15-59 N; p < 0.001). Thus, in all cases the helical fastener was significantly more stress resistant. The main reason for detachment of the mesh was tissue disruption or deformation of the fixation elements. Only when a stress of more than 130 N was applied did the mesh tear in two cases. CONCLUSIONS: The stress-bearing capacity (shear force resistance) of a mesh fixed by a helical fastener is up to four times that of a mesh fixed by a stapler. Therefore, the helical fastener provides significantly more stable fixation and will be able to protect the patient better from recurrent hernias caused by mesh migration. PMID- 10526027 TI - Treatment of endoscopic esophageal perforation. AB - BACKGROUND: The increasing usage of flexible endoscopy leads to a higher incidence of esophageal perforations, whose treatment strategies (conservative or operative) still are discussed controversially. We present our experiences and therapy concepts in relation to 75 iatrogenic esophageal perforations. PATIENTS: Between 1983 and 1997, 75 patients were treated for endoscopic perforation of the esophagus. The gender distribution was 31 females (41.3%) and 44 males (58.7%), with a mean age of 64.4 years (range 2-90 years). RESULTS: Therapeutic endoscopy was the most common cause of perforation (73 of 75 patients; 97.3%). Diagnostic endoscopy caused perforation in 2 patients (2.7%). The perforation was located in the cervical part of the esophagus in 7 patients (9.3%), the intrathoracic part in 25 patients (33.3%), and the abdominal part in 43 patients (57.3%). In this study population, 25 patients (33.3%) were treated surgically, and 50 patients (66.7%) conservatively. The overall in-hospital mortality rate was 14 of 75 patients (18.7%). In the surgically treated group the rate was 6 of 25 patients (24%) and in the conservative group 8 of 50 patients (16%). CONCLUSIONS: The decision of a treatment strategy depends on different factors such as the location and extent of the injury, the time interval between perforation and treatment onset, the preexisting diseases, and the patient's general condition. In view of these factors, an individual therapy concept should be determined for every patient. PMID- 10526028 TI - TNM staging and assessment of resectability of pancreatic cancer by laparoscopic ultrasonography. AB - BACKGROUND: Laparoscopic ultrasonography (LUS) is an imaging modality that combines laparoscopy and ultrasonography. The purpose of this prospective blinded study was to evaluate the TNM stage and assessment of resectability by LUS in patients with pancreatic cancer. METHODS: Of the 71 consecutive patients admitted to our department, 36 were excluded from the study, mainly due to evident signs of metastatic disease or another condition that would preclude surgery. Thus, a total of 35 patients were enrolled in the study. All patients underwent abdominal CT scan, ultrasonography, endoscopic ultrasonography (EUS), diagnostic laparoscopy, and LUS. Histopathologic examination was considered to be the final evaluation for LUS in all but three patients, where EUS was used as the reference. RESULTS: The accuracy of LUS in T staging was 29/33 (80%); in N staging it was 22/34 (76%); in M staging, it was 23/34 (68%); and in overall TNM staging, it was 23/34 (68%). In assessment of nonresectability, distant metastases, and lymph node metastases, the sensitivity was 0.86, 0.43 and 0.67, respectively, for LUS alone. Combining the information gleaned from laparoscopy and LUS, the accuracy in finding nonresectable tumors was 89%. CONCLUSIONS: Diagnostic laparoscopy with LUS is highly accurate in TNM staging and assessment of resectability of pancreatic cancer and should be considered an important modality in the assessment algorithm. PMID- 10526029 TI - Laparoscopic Duhamel procedure. Management of 30 cases. AB - BACKGROUND: Between February 1995 and June 1998, 30 laparoscopic Duhamel pull through procedures were performed in our department. METHODS: Our main aim was to prove the feasibility of the laparoscopic abdominal Duhamel procedure for different localizations of Hirschsprung disease. We used one camera port and three working ports. The sigmoid colon and posterior rectum were mobilized laparoscopically. A standard posterior colo-anal anastomosis was fashioned and a stapler was used for the anterior anastomosis. The top of the rectum was then closed by endo stapler under laparoscopic vision. RESULTS: Thirty patients underwent laparoscopic surgery for this procedure. Three laparoscopic procedures were converted because of technical difficulties. The operative time was 100-330 mn. Oral feeding was started at a mean postoperative time of 2.5 days. Mean postoperative hospitalization was 9 days. Early postoperative complications included 1 anastomotic leak, 1 retrorectal abscess, 2 urinary infections, and 1 evisceration (after conversion). No enterocolitis or enterocolitis-like symptoms were noted. All patients now have daily spontaneous bowel movements. CONCLUSION: The laparoscopic Duhamel procedure can be performed safely, giving good results. PMID- 10526030 TI - The use of multiplane transesophageal echocardiography to evaluate residual patent ductus arteriosus during video-assisted thoracoscopy in adults. AB - BACKGROUND: Video-assisted thoracoscopic surgery (VATS) has emerged as an innovative and popular procedure for interruption of patent ductus arteriosus (PDA), while intraoperative transesophageal echocardiography (TEE) has proven to be an effective monitor in the evaluation of residual patency. Previous reports on the adequacy of surgical interruption of PDA under VATS and TEE are available for pediatric patients, but only limited information is available for adults with PDA. MATEIALS AND METHODS: Between August 1995 and October 1997, we monitored 35 adult patients undergoing PDA interruption via VATS with Hewlett-Packard color Doppler multiplane TEE throughout the procedure. The average PDA diameter was 10.2 +/- 1.8 mm. All the PDA were completely ligated. RESULTS: Thirty-two patients showed no ductal flow after double ligation. In the other three patients, residual flow was detected intraoperatively after double ligation, but it was quickly abolished by the third ligation. One patient showed faint ductal flow by transthoracic echocardiography at postoperative follow-up, but no reintervention was needed. CONCLUSIONS: Our study showed that, with the refinement of adult PDA interruption via VATS, intraoperative multiplane TEE provides higher resolution for direct evaluation of the entire course of PDA ligation without interrupting the surgical procedure and minimizes the incidence of complications. PMID- 10526031 TI - Accuracy of endorectal ultrasound after preoperative radiochemotherapy in locally advanced rectal cancer. AB - OBJECTIVES: Factors limiting the accuracy of endorectal ultrasound in staging, locally advanced primary rectal cancer after preoperative neoadjuvant radiochemotherapy (RCT) were evaluated. METHODS: Patients (n = 84) with initial locally advanced rectal cancer (uT3/uT4) undergoing R0 resection were investigated after preoperative treatment that combined radiotherapy up to 45 Gy with two cycles of chemotherapy (5-FU and leucovorin on d 1-5 and 22-28). At 4 to 6 weeks after completion of RCT and before tumor resection, preoperative endoluminal ultrasound was performed. RESULTS: The accuracy to predict the depth of tumor infiltration (T-category) was found to correlate with downstaging. The T category was correctly staged before surgery in 15 of the 51 responders (29%) and in 27 of 33 nonresponders (82%), whereas misinterpretation occurred in 36 of the responders (71%) and in 6 of the nonresponders (18%) (p < 0.001). Neither tumor distance from anal verge nor tumor location correlated with the staging accuracy. Lymph node involvement was correctly assessed in 48 patients (57%). Wall invasion was correctly ascertained in 42 patients (50%), with under estimation in 11 patients (13%) and overestimation in 31 patients (37%). CONCLUSIONS: After radiochemotherapy, endosonography does not provide a satisfactory accuracy for preoperative staging of rectal cancer. New interpretation and diagnostic criteria are needed for the prediction of treatment response. PMID- 10526032 TI - Laparoscopic treatment of lymphocele after kidney transplantation. AB - BACKGROUND: Laparoscopic treatment of pelvic lymphocele secondary to kidney transplant has gained popularity in the last few years, although lesions of the urinary tract (ureter, renal pelvis, and bladder) have been reported frequently. To evaluate the result of this treatment and the associated risk of urinary tract lesions, we reviewed our experience and reports in the medical literature on open and laparoscopic surgery. METHODS: From 1991 to 1999, we laparoscopically treated 12 patients (7 men and 5 women; median age, 43 years; range, 17-59 years) with symptomatic pelvic lymphocele causing a deterioration of renal function because of compression on the ureter in 10 of the 12 patients and lymphocele compression of the iliac vein in the other 2 patients. In nine patients, the lymphocele wall was opened and sutured to the peritoneum to keep the window open. In two patients, an omentoplasty was performed, and in the remaining patient, both techniques were used. All patients were followed up clinically with ultrasound and biochemistry for a median period of 33 months (range, 1-96 months). Using Medline, we reviewed the medical literature from 1980 to 1998 and collected 252 cases in which operations had been performed to drain an internal lymphocele secondary to kidney transplantation. RESULTS: Laparoscopic treatment was successful in 11 of the 12 patients. One patient was converted to open surgery because of a lesion in the transplanted ureter. One patient needed repeat laparoscopy 24 hours after the operation because of bleeding from the peritoneal window. The median duration of the operation was 120 min (range, 70-200 min), and the median postoperative hospital stay was 5 days (range, 2-12 days). None of the patients needed to discontinue oral cyclosporine assumption. The serum creatinine level dropped significantly after surgery (p < 0. 05). No symptomatic recurrences were observed. Of the 252 patients found in the medical literature, in 129 the procedure was performed with open surgery and in 123 laparoscopically (our 12 patients included). The prevalence of iatrogenic lesions to the urinary tract increased threefold with the use of laparoscopic surgery (from 1.6% in open surgery to 7% in laparoscopy). The recurrence rate of symptomatic lymphocele, however, decreased from 15% to 4%. CONCLUSIONS: Laparoscopic drainage of posttransplantation lymphocele is a relatively simple method for treating this complication, although it bears the burden of an increased incidence of urinary tract lesions, as confirmed by a review of the literature. The major advantage of the laparoscopic approach is the absence of postoperative ileus with the opportunity to continue the enteral immunosuppressive regimen and a lower recurrence rate. These data suggest that laparoscopic lymphocele treatment might be considered the therapy of choice, provided the iatrogenic lesions of the urinary tract diminish as more experience with this technique is gained. PMID- 10526033 TI - Gastrostomy for enteral access. A comparison among placement by laparotomy, laparoscopy, and endoscopy. AB - BACKGROUND: Access to the stomach for long-term enteral feeding can be achieved via laparotomy (open GT), laparoscopy (lap GT) or endoscopy (PEG). We compared the three methods of gastrostomy to determine whether any one has an advantage over the others. METHODS: A retrospective analysis was done of 356 gastrostomies performed between January 1990 and June 1995. RESULTS: Of these 356 gastrostomies, 214 were open GT, 60 were lap GT, and 82 were PEG. The completion rate was high, 98.1% to 100%. The perioperative mortality rates were low and similar among the 3 methods; 4.2% for open GT, 5.3% for lap GT, and 4.9% for PEG (p = 0.87, Chi square test). Cardiac arrest was the predominant immediate cause of all perioperative deaths (68.8%). Overall, none of the deaths was directly related to the gastrostomy procedure. Major complications occurred in 24.9% of patients receiving open GT, in 18.3% of patients with lap GT, and in 17.1% of patients with PEG. Long-term complications developed in 25.9% of open GT, 25.6% of lap GT, and 30. 4% of PEG. The revision rates were similar for all 3 methods, 6.7% for open GT, 10% for lap GT, and 6.1% for PEG. CONCLUSIONS: Gastrostomy can be performed safely by all three techniques, with similar outcomes. PEG is our method of choice. Lap GT is preferred in patients with head and neck carcinoma, patients with obstructing esophageal carcinoma, and patients who have problems with overlying liver or colon. Open GT is reserved for cases with extensive intraabdominal adhesions or those where the procedure is done during an ongoing laparotomy. PMID- 10526034 TI - Primary laparoscopic placement of gastrostomy buttons for feeding tubes. A safer and simpler technique. AB - BACKGROUND: During a 4-year period, 240 gastrostomy buttons were placed in children, as the initial surgical feeding tube, using laparoscopic techniques. MATERIALS AND METHODS: The technique requires the use of a minilaparoscope (1.6 mm) and a single 5-mm trocar placed at the exit site for the gastrostomy button. It can also be performed in addition to a laparoscopic fundoplication using the same trocar sites. The technique requires no special instrumentation or kits. When performed alone, operative times average 15 min. When performed with fundoplication, it adds approximately 5-10 min to the time for the procedure. RESULTS: There were no intraoperative complications and five (2.1%) postoperative complications. CONCLUSIONS: This technique has proven to be simple and effective. It allows primary placement of a gastrostomy button that is cosmetically and functionally superior to a gastrostomy tube. PMID- 10526035 TI - The effect of CO2 pneumoperitoneum on the growth of a solid colon carcinoma in rats. AB - BACKGROUND: In order to investigate the effect of carbon dioxide (CO(2)) pneumoperitoneum on solid colon carcinomas, we used a colon anastomosis tumor model in 30 male syngeneic WAG rats, which were divided, at random into three groups. METHODS: In all rats, 10(6) CC531 S colon carcinoma cells were injected as an enema into the colon. Subsequently, a transection and a reanastomosis of the colon descendens was performed via laparotomy. After 2 weeks, group 1 (n = 10) was anesthetized as an anesthesia control group. Group 2 (n = 10) had a laparotomy that was closed after 20 min. In group 3 (n = 10), a CO(2) pneumoperitoneum of 22 (scores = 61.8, 29.4), while only one patient had a pathologic total time of reflux (percent time of reflux, 8%). The mean percent time of reflux in the other 13 patients was 1.9 +/- 0.6% (range, 0.1 4%), and the mean DeMeester score was 11.7 +/- 4.6 (range, 0.48-19.7). CONCLUSIONS: Laparoscopic Heller myotomy is effective for the relief of dysphagia in achalasia if the myotomy lowers the LES pressure to <17 mmHg. If performed without dissection of the entire esophagus, the laparoscopic Heller myotomy does not create significant GER in the postoperative period. Clearance of acid refluxate from the aperistaltic esophagus is an important component of the pathologic gastroesophageal reflux disease (GERD) seen after Heller myotomy for achalasia. Furthermore, GERD symptoms do not correlate with objective measurement of GE reflux in patients with achalasia. Objective measurement of GERD with 24 h pH probes may be indicated to identify those patients with pathologic acid reflux who need additional medical treatment. PMID- 10526039 TI - Operative manometry and endoscopy during laparoscopic Heller myotomy. An initial experience. AB - BACKGROUND: We report our initial experience using operative esophageal manometry as an adjunct to endoscopy to determine the completeness of esophagogastric high pressure zone (HPZ) obliteration during laparoscopic Heller myotomy. METHODS: Between July 1997 and October 1998, we performed laparoscopic Heller myotomies in 20 patients (eight male, 12 female; median age, 41 years). Mean duration of symptoms was 3.2 +/- 2.6 years (r = 0.5-11), and 45% of the patients had received prior dilation or toxin injection. A 16-channel esophageal manometry catheter was placed prior to anesthesia, with sites crossing the lower esophageal sphincter (LES). An endoscope was passed intraoperatively to localize the squamocolumnar junction, and the myotomy was performed. While the translucency was imaged in the area of the incision, we determined the adequacy of myotomy by visual assessment of LES and gastric cardia opening in response to endoscopic air insufflation. Manometry was then performed to detect any potential residual high pressure at the myotomized esophagogastric junction (EGJ). If it was found, the locus of persistent pressure was identified by probing along the myotomy, and residual muscle fibers were cut to yield a minimum pressure at the EGJ. RESULTS: A persistent HPZ was identified after the initial myotomy in 10 of 20 patients (50%). A Dor fundoplasty completed the operation. The mean operating time was 2.6 +/- 0.5 h (median, 2.5; r = 2-3.5 h), and the mean hospital stay was 1.6 +/- 1 days (median, 1, r = 1-5 days). The mean LES pressure was 2 +/- 3 mmHg immediately postmyotomy (p < 0.001 compared with preoperative value). Of 20 patients, only two have reported recurrence of dysphagia (10%). One had a recurrent HPZ on manometry, and one developed esophagitis, which resolved with omeprazole. CONCLUSIONS: Our initial experience suggests that operative esophageal manometry is a useful adjunct to upper endoscopy during laparoscopic Heller myotomy, quantitatively assuring obliteration of the nonrelaxing LES and HPZ. PMID- 10526040 TI - Influence of different gases and intraperitoneal instillation of antiadherent or cytotoxic agents on peritoneal tumor cell growth and implantation with laparoscopic surgery in a rat model. AB - BACKGROUND: A generally accepted approach to prevent tumor implantation with laparoscopic surgery does not exist. Alternative gases in combination with intraperitoneal instillation of different antiadherent or cytotoxic agents have not been evaluated. METHODS: The effect of taurolidine, heparin, and povidone iodine on the growth of colon adenocarcinoma DHD/K12/TRb was measured in rats undergoing laparoscopy with carbon dioxide (n = 40), helium (n = 40), or xenon (n = 40). In the procedure, 10(4) tumor cells were administered intraperitoneally, and pneumoperitoneum was established over 30 min at 8 mmHg with the different gases. The rats additionally received intraperitoneal instillation with one of the following: 1 ml of Ringer's solution, 1 ml of 0.5% taurolidine, 1 ml 0.5% taurolidine with heparin (10 U/ml), or 1 ml 0.25% of povidone-iodine. Tumor growth was measured after 4 weeks. RESULTS: Median intraperitoneal tumor weight was lower in rats receiving taurolidine (CO(2): 10 mg; helium: 50 mg; xenon: 39.5 mg) or taurolidine with heparin (CO(2): 4 mg; helium: 4.5 mg; xenon: 46.5 mg) in all gas groups than in the control groups (CO(2): 427 mg; helium: 268 mg; xenon: 345 mg) (p < 0.001). Whereas povidone-iodine caused significantly lower tumor growth in the CO(2) group (56.5 mg) (p < 0.01), the combination of helium (145 mg) and xenon (457 mg) with povidone-iodine produced no reduction of tumor growth as compared with the control groups (helium: 268 mg; xenon: 345 mg). CONCLUSIONS: Taurolidine and taurolidine with heparin significantly inhibit intraperitoneal tumor growth, with different gases used for pneumoperitoneum. Only povidone iodine caused significant decrease of tumor growth in combination with CO(2). The combination of xenon and povidone-iodine should not be used in patients with cancer because of increased tumor growth. PMID- 10526042 TI - Overview of randomized trials of inguinal hernia repair-a European Union concerted action. Report of first collaborators' meeting, Nieuwegein, the Netherlands, October 16-17, 1998. EU Hernia Trialists Collaboration. PMID- 10526041 TI - Laparoscopic extraperitoneal inguinal hernia repair with spinal anesthesia and nitrous oxide insufflation. AB - BACKGROUND: Laparoscopic repair of inguinal hernia is traditionally performed under general anesthesia mainly because of the adverse effects that carbon dioxide pneumoperitoneum has on awake patients. Since a mandatory use of general anesthesia for all hernia repairs is questionable, the feasibility of laparoscopic extraperitoneal herniorraphy using spinal anesthesia combined with nitrous oxide insufflation was investigated. METHODS: Over a 4-month period, February to May 1998, we performed 35 consecutive total extraperitoneal inguinal hernia procedures (24 unilateral, 11 bilateral) using spinal anesthesia and nitrous oxide extraperitoneal gas. Data on operative findings, self-reported operative and postoperative pain and discomfort (visual analog pain scale), procedure-related hemodynamics, and complications were collected prospectively. RESULTS: All 35 procedures were completed laparoscopically without the need to convert to general anesthesia. Mean operative time was 39 +/- 7 min for unilateral hernia and 65 +/- 10 min for bilateral hernia. Incidental peritoneal tears occurred in 22 patients (63%) resulting in nitrous oxide pneumoperitoneum, which was well tolerated. The patients remained hemodynamically stable throughout the procedure, and operative conditions and visibility were excellent. Complications at a mean of 4 months after the procedure included seven uninfected seromas (20%), three patients with transient testicular pain, and one (3%) recurrence. CONCLUSIONS: Laparoscopic total extraperitoneal hernia repair can be safely and comfortably performed using spinal anesthesia with extraperitoneal nitrous oxide insufflation gas. This method provides a good alternative to general anesthesia. PMID- 10526043 TI - Delayed-type hypersensitivity response is better preserved in mice following insufflation than after laparotomy. AB - BACKGROUND: Our laboratory has previously used pig and rat models to demonstrate that delayed-type hypersensitivity (DTH) response to an antigen challenge is suppressed following laparotomy compared to insufflation. The purpose of this study was to develop a practical and reliable mouse DTH model that could be used in future studies to test immunomodulating drugs and therapies. METHODS: Female C3H/HeN mice (n = 100) were given three serial DTH challenges of 25 microl of 4 mg/ml phytohemagglutinin (PHA) 12 days before the test procedure, immediately following the test procedure, and on the 2nd postoperative day. All challenges were administered via subcutaneous injection in alternating footpads. The thickness of the footpad was determined with electronic calipers immediately prior to injection and 24 h following injection in a blinded fashion. The difference in thickness represents the response. On the day of the procedure, mice were randomized into the following three groups: (a) control (AC), (b) insufflation (INS), and (c) open (OPEN). AC mice underwent no procedure. INS mice underwent CO(2) insufflation at 2-4 mmHg for 20 min. OPEN mice underwent a midline incision from xiphoid to pubis that was closed after 20 min. Data were analyzed using ANOVA and Tukey-Kramer tests to determine differences between groups. RESULTS: Preoperatively, there were no significant differences among the three groups. On POD1, the OPEN group had significantly less response than both the AC and INS groups. On POD3, there were significant differences between the OPEN group and both the INS and AC groups. There was no significant difference between the AC and INS group at any time. CONCLUSIONS: In conclusion, a DTH mouse model has been established that allows serial assessment of cell-mediated immune function. This model can be used to study immune function after open and minimal access procedures in a simple and cost-effective manner. PMID- 10526044 TI - Videoscopic surgery under local and regional anesthesia with helium abdominal insufflation. AB - BACKGROUND: High-risk patients may not be good candidates for laparoscopic surgery due to the metabolic consequences of transperitoneal absorption of insufflated CO(2) gas and the necessity of general anesthesia because CO(2) insufflation produces pain. Helium gas is metabolically inert and does not produce pain. Thus it permits an alternative approach to performing laparoscopic surgery in high-risk patients. METHODS: Laparoscopic cholecystectomy, appendectomy, hernia repair, and peritoneal dialysis catheter procedures were performed under local or regional anesthesia in high-risk patients utilizing helium gas as the insufflation agent. RESULTS: Twenty-one patients underwent laparoscopic procedures under local or regional anesthesia. None of the procedures initiated under local-regional anesthesia required abandonment of the laparoscopic approach or conversion to general anesthesia. There were no operative or perioperative mortalities. Two incidences of pneumothorax occurred with extraperitoneal hernia repair; one required a tube thoracostomy. CONCLUSIONS: Helium gas should be considered the agent of choice for intraperitoneal insufflation in high-risk patients not only because helium avoids the metabolic consequences of CO(2) insufflation but also because it permits selected procedures to be performed under local-regional anesthesia. Helium may be contraindicated for laparoscopic procedures involving extraperitoneal insufflation due to the increased risk for pneumothoraces. PMID- 10526045 TI - Recurrent pancreatitis in a child with pancreas divisum. Endoscopic therapy of a Santorinicele. AB - Pancreas divisum is a rare congenital anomaly of the pancreatic ducts that has been implicated in pancreatitis. In addition, the finding of a Santorinicele, which is a cystic dilatation of the dorsal duct, suggests that there is an obstruction associated with a congenital or acquired weakness of the mucosa. We used an endoscopic technique to treat a child with recurrent pancreatitis who was found to have pancreas divisum and a large Santorinicele. PMID- 10526046 TI - Esophageal perforation and mediastinal abscess following placement of a covered self-expanding metallic stent and radiation therapy in a cancer patient. AB - Patients with advanced esophageal cancer may require intubation with a stent to relieve debilitating dysphagia. However, if these patients also undergo radiation therapy, they may incur esophageal injury, thus increasing the risk of perforation after placement of the stent. Herein we report the case of a 71-year old man who received such combination therapy and died of severe sepsis 65 days after the stent was inserted. An autopsy revealed that the stent had perforated into the mediastinal pleura and that an abscess had developed around the perforation. We conclude that caution should be taken before combining radiation therapy with the use of a stent. PMID- 10526047 TI - The role of laparoscopy in symptomatic Meckel's diverticulum. AB - We report two cases of symptomatic Meckel's diverticulum in adults with recurrent abdominal pain and episodes of minor lower gastrointestinal bleeding. In case 1, the diagnosis was suggested by (99m)Tc pertechnetate scan and confirmed by laparoscopy; whereas in case 2, only diagnostic laparoscopy was performed because of suspected appendicitis. A segmental small bowel resection with attached diverticulum was performed extracorporeally after exteriorization through the umbilical port site in both cases. PMID- 10526048 TI - Laparoscopic resection of large leiomyomas of the gastric fundus. AB - Two patients with a large leiomyoma arising from the gastric fundus underwent laparoscopic resection. In case 1, the tumor was located in the anterior wall of the gastric fundus. To prevent stenosis and preserve the volume of the residual stomach, intragastric resection was adopted. The tumor was markedly and resected with laparosonic coagulating shears with a 1-cm safety margin. In case 2, a large tumor was detected in the duodenal bulb. Serious hemorrhage mandated emergency resection. The tumor originated from the posterior wall of the fundus. Attempts at reduction with the forceps failed. Reduction by digital manipulation via laparoscopic port sites was successful. An endostapler was used to resect the tumor and close the anterior wall. Both patients recovered uneventfully. PMID- 10526049 TI - A reliable and efficient technique for laparoscopic needle positioning. AB - Needle positioning can be a difficult, frustrating, and time-consuming step during laparoscopic suturing. Utilizing the reliable and efficient technique described in this article, needle positioning is expedited. This technique is applicable for any type of needledriver or suture. PMID- 10526050 TI - Laparoscopic splenectomy using a wall-lifting procedure. AB - A laparoscopic splenectomy using a hanger wall-lifting procedure is herein described. The patient is placed in the right lateral position. The left lower chest and left abdominal wall are then lifted by three wires in two directions, left laterally and vertical to the abdominal wall. The view of the operative field thus obtained is excellent. The lifting wires and bars do not hinder the movement of the forceps, since the angles of the instruments to approach the spleen are different from those of the wires. A laparoscopic splenectomy using this wall-lifting procedure avoids the usual complications associated with pneumoperitoneum while still being technically comparable to a procedure with pneumoperitoneum. PMID- 10526051 TI - A two-port technique of laparoscopic placement of Tenchkoff catheter with a means to prevent catheter migration. PMID- 10526053 TI - Bile duct cancer developed after cyst excision for choledochal cyst. AB - Oncogenesis after cyst excision for choledochal cyst and suitable surgical procedures for this operation are discussed. The clinical data of 23 patients with cancer of the biliary tree after excision of choledochal cyst reported in the English-language and Japanese literature were reviewed, and data for 1353 Japanese patients with choledochal cyst and/or pancreaticobiliary malunion were analyzed. In the 23 patients reported in the literature, age at cyst excision ranged from 1 to 55 years (average, 23.0 +/- 13.7 years), and cancers were detected at age 18-60 years (average, 32.1 +/- 12.2 years), with intervals between cyst excision and cancer detection of 1-19 years (average, 9.0 +/- 5.5 years). Sites of cancer development were: intrahepatic, six; anastomotic, eight; hepatic side residual cyst, three; and the intrapancreatic duct, six. In the Japanese patients with choledochal cyst and/or pancreaticobiliary malunion, the incidence of cancer associated with primary choledochal cyst and/or pancreaticobiliary malunion was 16.2% (219/1353). The incidence of cancer development after cyst excision in this population, of whom 1291/1353 underwent surgery, was assumed to be 0. 7%. Nearly half of the 23 patients in the literature had undergone inadequate cyst excision. Oncogenesis of cancers after cyst excision is possibly different from that of choledochal cysts. PMID- 10526054 TI - Mucosal cell proliferation activity of the gallbladder in children with anomalous arrangement of the pancreaticobiliary duct. AB - Anomalous arrangement of the pancreaticobiliary duct (AAPBD) is an anatomical maljunction of the bile duct and the pancreatic duct that is frequently associated with gallbladder carcinoma. In patients with AAPBD, it has been postulated that pancreatic juice regurgitates into the biliary tree, and the mixture of refluxed pancreatic juice and stagnant bile juice acts as an irritant factor to the biliary tract epithelium, leading to chronic inflammation and metaplasia. Eventually these mucosal changes may progress to invasive carcinoma. We reviewed clinicopathologic studies on epithelial changes of the gallbladder in patients with AAPBD to clarify the implications relevant to carcinogenesis. Conventional histological studies have shown that the most characteristic change observed in the gallbladder of children with this anomaly was epithelial hyperplasia. Furthermore, the incidence of mucosal hyperplasia was significantly increased in the gallbladder of children in whom the pancreatic duct joined the common bile duct (P-C type) compared with the incidence in children in whom the common bile duct joined the pancreatic duct (C-P type). In addition, cell kinetic studies have demonstrated increased cellular proliferative activity of the gallbladder in children with AAPBD. Cell proliferative activity was significantly elevated in children with the P-C type of AAPBD compared with that in children with the C-P type of anomaly. In conclusion, AAPBD may yield increased cell proliferation in the gallbladder of patients with this anomaly in early childhood, resulting in epithelial hyperplasia. Although it remains unknown which agents are responsible for promoting the activation of cellular function, it seems that bile acids and refluxed pancreatic proteases are likely play a role in such promotion. Further investigations are needed to elucidate the mechanism of increased cellular function. PMID- 10526055 TI - Carcinogenesis in the biliary system associated with APDJ. AB - Anomalous pancreaticobiliary ductal junction (APDJ) is a rare congenital anomaly which is considered to be an etiological factor in the development of carcinoma of the biliary tract. It is generally accepted that pancreatic juice reflux into the biliary tract due to APDJ is one of the etiologies of biliary tract cancers. Refluxing pancreatic juice results in changes of bile and induces chronic inflammation and increased cellular proliferation, leading to epithelial hyperplasia, metaplasia, and carcinoma of the biliary tract. K-ras mutations are more prevalent in the carcinomas of biliary tract associated with APDJ compared with those without APDJ. There is no difference in the overexpression of p53 between biliary tract carcinomas associated with APDJ and those unassociated with APDJ. Further studies are needed to evaluate the role of cytokines and growth factors in carcinogenesis of the biliary system associated with APDJ. PMID- 10526056 TI - Cellular kinetics and gene mutations in gallbladder mucosa with an anomalous junction of pancreaticobiliary duct. AB - Anomalous junction of the pancreaticobiliary duct (AJPBD) is thought to be an important risk factor for gallbladder carcinoma in Japan. We have reported the characteristic pathology, cellular kinetics, and gene mutations to clarify the mechanism of carcinogenesis in gallbladder mucosa with AJPBD. A comprehensive review of the literature was undertaken, with referencing of major articles on the subject. A sequence of hyperplastic changes, with a corresponding increase in cellular kinetics with progression through dysplasia to carcinoma is important in carcinogenesis of gallbladder mucosa with AJPBD. p53 mutations may contribute to the transition from premalignancy to malignancy in the early stage of carcinogenesis of the gallbladder mucosa, regardless of the presence of AJPBD. The specific mutation of GGT-to-GAT in codon 12 of K-ras may play an important role in carcinogenesis of gallbladder mucosa with AJPBD. PMID- 10526057 TI - Epithelial cell proliferation and gene mutation in the mucosa of gallbladder with pancreaticobiliary malunion and cancer. AB - The significant association between pancreaticobiliary malunion (PBM), especially undilated-type PBM, and a high risk of gallbladder cancer is known. Reflux and stasis of pancreatic juice induce various epithelial changes in the gallbladder. Recently, epithelial hyperplasia of the gallbladder was shown to be significantly and frequently associated with undilated-type PBM, and it is suggested that the majority of epithelial hyperplasia may exist at birth or be acquired in early childhood, and thereafter present throughout the lives of PBM patients. Cell kinetic studies demonstrated a significant stepwise increase in cellular proliferative activity from normal gallbladder mucosa, through epithelial hyperplasia to cancer. Epithelial hyperplasia with increased proliferative activity may predispose the mucosa to mutational events, thereby increasing cancer risk in PBM patients. K-ras mutations were frequently detected in gallbladder cancer in PBM patients and in epithelial hyperplasia as well. Epithelial hyperplasia is demonstrated to be an important premalignant lesion of gallbladder cancer. A multistep process of carcinogenesis as a consequence of multiple genetic alterations of oncogenes and tumor suppressor genes has been demonstrated in various organs; however, there is limited information on the molecular mechanism in gallbladder carcinogenesis with PBM. Recent findings support the idea that epithelial hyperplasia plays an important role in gallbladder carcinogenesis with PBM and also support the concept that neoplastic development in gallbladder with PBM also evolves through a multistep process associated with hyperproliferation and genetic alterations. PMID- 10526058 TI - Genetic abnormalities involved in the pathogenesis of gallbladder carcinoma. AB - While considerable progress has been made in the understanding of the genetic changes involved in the pathogenesis of several human neoplasms, there is limited information about the genetic changes involved in the development of gallbladder carcinoma. Several studies indicate that TP53 (17p13) and p16(Ink4)/CDKN2 (9p21 22) gene loci abnormalities are frequent and early events in the pathogenesis of this neoplasm, in some cases preceding the onset of histological changes of invasion. Preliminary data also suggest that deletions at other chromosomal regions (8p21 and DCC at 18q21 loci) may play an important role in the development of gallbladder carcinoma; however, they need to be further analyzed. K-ras gene mutations appear to be an infrequent event in this neoplasm, except in gallbladder carcinomas associated with congenital abnormalities of the biliary tract. Genetic studies confirm that the sequence dysplasia-carcinoma in situ (CIS) is the usual route for the development of gallbladder carcinoma, and our recent data strongly suggest that adenomas are not precursors of this neoplasm. PMID- 10526059 TI - Technical advances in living-related liver transplantation. AB - Since it was first reported in 1989, living-related liver transplantation (LRLT) has developed, and up to April, 1998, over 800 LRLTs had been performed worldwide. The past few years have seen considerable technical advances in LRLT, including selective vascular occlusion techniques for donor hepatectomy, arterial reconstruction using microscopy, and the introduction of intraoperative ultrasound and graft volume estimation, using the concept of standard liver volume, which have enabled LRLT to be extended to adult recipients. Successful results have led to LRLT becoming an indispensable modality to overcome the shortage of cadaveric liver grafts in Western countries. In places where transplantation of organs from brain-dead donors is rarely practiced, such as in most Asian countries, LRLT is the only means of saving patients with end-stage liver disease who otherwise would have no chance of survival. LRLT is now globally accepted as an effective liver transplantation modality. PMID- 10526060 TI - The Reg gene family and Reg proteins: with special attention to the regeneration of pancreatic beta-cells. AB - Pancreatic beta-cells of the islets of Langerhans are the only cells that produce insulin in humans, as well as in most animals, but they have been thought to have a limited capacity for regeneration, which is a predisposing factor for the development of diabetes mellitus. Strategies for influencing the replication and growth of the beta-cell mass are therefore important for the prevention and/or treatment of diabetes. We have established a model for islet regeneration in 90% depancreatized rats by the administration of poly(ADP-ribose) synthetase inhibitors such as nicotinamide. The regenerating islets in the remaining pancreas of poly(ADP-ribose) synthetase inhibitor-treated rats were markedly enlarged and consisted largely of insulin-producing beta-cells, preventing the development of diabetes mellitus that would otherwise be caused by the 90% pancreatectomy. In screening the regenerating islet-derived cDNA library, we found a novel gene and named it Reg (i.e., regenerating gene). The rate Reg cDNA had a single open reading frame that encoded a 165-amino acid protein with a 21 amino acid signal peptide. We also isolated the human REG cDNA which encoded a 166-amino acid protein with a 22-amino acid signal peptide. The amino acid sequence of human REG protein has 68% homology to that of rat Reg protein. Recombinant rat Reg protein without the signal peptide, produced in yeast, stimulated beta-cell replication and increased the beta-cell mass in the residual pancreas of 90% depancreatized rats, ameliorating the surgical diabetes. Recombinant human REG protein also induced an expansion of the beta-cell mass in non-obese diabetic (NOD) mice, resulting in the amelioration of diabetes. These results, as well as several other lines of evidence, indicate that Reg protein is a growth factor for pancreatic beta-cells and also suggest that the administration of Reg protein and/or activation of the Reg gene can be used as a potential therapeutic approach for diabetes mellitus. We have further isolated several Reg and Reg-related genes from human, rat and mouse, and revealed that they constitute a multigene family, the Reg gene family. Based on the primary structures of proteins encoded by the Reg gene family, we have grouped the members of the family into three subclasses, type I, II, and III. Type I Reg (Reg I) encodes a beta-cell growth factor, Reg I protein, as mentioned above. Some of the type III Reg (Reg III) have recently been suggested to play roles in the regeneration of cells other than pancreatic beta-cells, such as neuronal cells and epithelial cells in the alimentary tract. PMID- 10526061 TI - The role of percutaneous transhepatic abscess drainage for liver abscess. AB - To evaluate the efficacy of percutaneous transhepatic abscess drainage (PTAD) as an initial choice of treatment for liver abscess, the medical records of 28 patients with liver abscess were retrospectively analyzed. The patients were predominantly men (23 of 28) with a mean age of 59 years (range, 19-86 years). Their chief complaints were fever (86%), right hypochondralgia (32%), and jaundice (11%). Fifteen of the 28 patients (54%) had hepatobiliary and pancreatic carcinoma, and 31% had postoperative liver abscess. PTAD was performed in 23 patients and surgical drainage in 5. The overall success rate for PTAD was 83%. The success rate for PTAD for patients with multiple abscesses was 83% (5 of 6), compared with a success rate of 82% (14 of 17) for patients with solitary abscess. The prognostic factors for survival were cancer and sepsis and the mortality rate for patients with cancer was 40% (6 of 15) while the mortality rate for patients with sepsis was 56% (5 of 9). As a complication of drainage, 1 patient (4%) in the PTAD group had pleural abscess due to the transpleural puncture. Our findings support the use of PTAD as the primary treatment for liver abscess, as it is safe and effective irrespective of the number of abscesses and the patient's condition. PMID- 10526062 TI - Amelioration of tumor necrosis factor release by cyclosporine in warm ischemia/reperfusion injury of the rat liver: with special reference to hepatic ultrastructure. AB - Mechanisms by which an immunosuppressant (cyclosporine, CsA) ameliorates warm ischemic injury of the liver were studied. Female Sprague-Dawley rats were subjected to 60-min normothermic liver ischemia. Animals were assigned to one of two groups: group I, controls with vehicle treatment; group II, treatment with CsA (10 mg/kg). CsA was given orally for 4 consecutive days prior to the induction of hepatic ischemia. In addition to a survival study, plasma levels of endotoxin, serum activity of tumor necrosis factor-alpha (TNF), and serum levels of aminotransferases were measured in blood samples collected from the suprahepatic vena cava, and hepatic ultrastructural alterations were examined under an electron microscope. The 7-day survival rate was significantly higher in the CsA-treated animals. In the control group, serum TNF levels were elevated following reperfusion and peaked at 3 h. When the values at 3 h post reflow were compared, the animals given CsA had significantly lower levels of TNF (170.0 +/- 30.5 pg/ml for group I, 67.6 +/- 13.7 for group II, mean +/- SEM; P < 0.05). The sinusoidal lining cells and hepatocytes were drastically destroyed at 6 h post reflow in the control group, although the degree of injury at 1-3 h was less severe. On the other hand, the endothelium and parenchymal liver cells in the CsA treated group were well preserved at 6 h in comparison with those in the control group. Our data suggest that modulation of TNF production is one of the mechanisms through which CsA prevents the exacerbation of ischemia/reperfusion injury of the liver. PMID- 10526063 TI - Immediate increase of portal pressure, reflecting sinusoidal shear stress, induced liver regeneration after partial hepatectomy. AB - The mechanisms whereby hepatocytes in the normal liver can be primed for replication following partial hepatectomy (PHx) are poorly understood. To determine whether "shear stress," which is induced by acute portal hypertension after PHx, is involved in liver regeneration, we studied liver regeneration in rats with splenic transposition (SPT) in which we can minimize the postoperative elevation of portal pressure. Rats underwent 70% PHx following splenic transposition or sham surgery and were killed at various time points to measure portal pressure and other factors. In the control groups, the portal pressure was significantly increased immediately after surgery, peaking at 48 h, and returning to near the preoperative levels by 168 h after PHx. In the SPT group, although portal pressure increased immediately, it decreased to the control levels 6 h after PHx and thereafter repeatedly increased. Tumor necrosis factor-alpha (TNF alpha) and interleukin-6 (IL-6) levels peaked at 24 and 6 h after PHx, respectively. Proliferative cell analysis was done using MIB-5 antibody, and there were no significant differences between the two groups. Furthermore, liver weight was restored in the same way in both groups. Taken together, the results suggest that an immediate increase in portal pressure is necessary for the initiation of liver regeneration. PMID- 10526064 TI - Mucin-producing tumors of the pancreas: clinicopathological features, surgical treatment, and outcome. AB - Mucin-producing tumors (MPTs) of the pancreas are increasingly being recognized. To evaluate the appropriate surgical treatment and predict the prognosis of MPTs, we performed a retrospective clinicopathological study in 51 patients, 27 with benign tumors and 24 with borderline/malignant tumors. Three of the malignant tumors showed stromal invasion and lymph node metastasis on histological examination. Of the 24 patients with borderline/malignant tumors, 2 died of MPTs and 4 died of other diseases. At the last follow-up, 35 patients were alive and well. The 5-year postoperative survival rate was 90% for patients with benign tumors, and 78% of these with borderline/malignant tumors. Five of the patients with borderline/malignant tumors had multicentric tumors. Three of these patients underwent resection of the rest of the pancreas, 5, 6, and 8 years, respectively, after the first operation. Extended radical resection is required for malignant MPT with invasion of the pancreatic stroma. We prefer to perform pancreatogastrostomy or Imanaga's procedure to allow examination of the body and tail of the pancreas by endoscopic retrograde pancreatography after resection of the pancreatic head. Careful follow-up for a long period may be the most prudent approach for detecting multiple MPTs in the residual pancreas after surgical treatment. PMID- 10526065 TI - p53 gene mutations and overexpression of p53 product in cancerous and noncancerous biliary epithelium in patients with pancreaticobiliary maljunction. AB - To investigate the molecular mechanisms of the high incidence of carcinogenesis in the biliary epithelium of patients with pancreaticobiliary maljunction, we examined p53 gene mutations, loss of heterozygosity of p53, and overexpression of p53 gene product in the cancerous and noncancerous biliary epithelium of 27 patients with pancreaticobiliary maljunction. Mutations of the p53 gene were examined by polymerase chain reaction-single strand conformation polymorphism and a direct sequencing method. Loss of heterozygosity of the p53 gene was determined using a double-targeted fluorescence in situ hybridization method. Expression of p53 gene product was examined using immunohistochemical staining. Mutations of the p53 gene were found in 4 of 5 biliary carcinomas (80%) and in 10 of 26 noncancerous biliary lesions (38.5%). Point mutations of the p53 gene were detected at codons 207, 212, and 217 on exons 5 through 8. The incidence of p53 gene mutations on exons 5, 6, 7, and 8 was 12. 9%, 36.4%, 0.0%, and 13.8%, respectively. Loss of heterozygosity of p53 was shown in 72% of the cells obtained from the cancerous lesion, and in an average of 14% obtained from the noncancerous lesions. Overexpression of p53 protein was found in 57.1% of carcinoma, and in 31.3% of the noncancerous lesions. These results suggest that p53 gene mutations are involved in the carcinogenesis of biliary epithelium in patients with pancreaticobiliary maljunction. PMID- 10526066 TI - Epithelial cell proliferation activity of the biliary ductal system with congenital biliary malformations. AB - Congenital biliary malformations such as anomalous arrangement of the pancreaticobiliary ductal system (AAPB), congenital cystic dilatation of the common bile duct (CCDB), and congenital biliary strictures at the hepatic hilum (CBSH) are newly designated disease entities and are frequently found in adult patients with biliary malignancy such as gallbladder carcinoma, common bile duct carcinoma, and intrahepatic bile duct carcinoma. In the present study, the relationship of these malformations and biliary malignancy was investigated. We studied 61 gallbladders of patients with AAPB and 56 gallbladders of patients without AAPB; 16 common bile ducts of patients with CCDB (12 with AAPB and 4 without AAPB) and 11 gallbladders of patients without CCDB; and 17 intrahepatic bile ducts of patients with CBSH and 6 intrahepatic bile ducts of patients without CBSH. Tissue sections from the mucosa of the gallbladder, common bile duct, and intrahepatic bile duct were stained for proliferating cell nuclear antigen (PCNA). The PCNA labeling indexes of patients with these malformations were significantly higher than those of patients without these malformations (P < 0.05). Cell proliferation of the epithelia in the biliary ductal system in patients with these congenital biliary malformations was accelerated. Consequently, these congenital malformations appear to be an important risk factor for the occurrence of biliary malignancy. PMID- 10526067 TI - Early and late complications of pylorus-preserving pancreatoduodenectomy in Japan 1998. AB - Early (within 1 month after operation) and late (more than 1 month after surgery) complications after pylorus-preserving pancreatoduodenectomy (PpPD) were analyzed in 1066 Japanese patients collected from 74 authentic institutions in Japan. As early postoperative complications after PpPD, delayed gastric emptying was evident in 46% of patients, pancreatoenterostomy leakage in 16%, intra-abdominal infection in 14%, cholangitis in 8.9%, hepaticojejunostomy leakage in 4.7%, intra abdominal hemorrhage in 3. 5%, upper gastrointestinal hemorrhage in 3.2%, and duodenojejunostomy leakage in 2.0%. Delayed gastric emptying resolved 1-24 months after PpPD (mean, 3.1 months). The direct operative mortality (death within 1 month after the operation) was 2. 4%. Univariate and multivariate analysis of pancreatoenterostomy leakage showed that male sex (P = 0.0151) and soft consistency of the pancreas (P < 0.0001) were independent significant factors. Univariate analysis of delayed gastric emptying showed that establishment of gastrostomy (P < 0.0001), length of the preserved duodenum (P = 0.0406), gastric juice output (P = 0.0001), length of gastric tube placement (P < 0.0001), and administration of cisapride (P = 0.0059) were significant variants. As late complications, stomal ulcer was evident in 3.6% of patients, cholangitis in 6.7%, and liver abscess in 1.2%. Glucose intolerance appeared in 61 patients, resolved in 15, showed no change in 170, was absent in 695, and was ameliorated in 17. As a result, the dosage of hypoglycemic agents or insulin showed no change in 187 patients, decreased in 16, and increased in 52. Diabetes appeared 0-42 months after PpPD (mean, 102 months). When present, diabetes deteriorated 0-36 months postoperatively (mean, 6.3 months). Univariate analysis of the appearance or deterioration of diabetes showed that diabetes occurred more frequently in the following patients; those with Billroth I reconstruction compared with those with Billroth II (P = 0.0041), those with pancreatogastrostomy vs those with pancreatojejunostomy (P = 0.0229), those with pancreatogastrostomy vs those with end-to-side pancreatojejunostomy (P = 0.0165), and those with total tube drainage vs those with pancreatico-whole thickness anastomosis (P = 0.0392); a high American Society of Anesthesiologist (ASA) score (P = 0.0211) and pancreatoenterostomy leakage (P = 0.0361) were also significant factors. Postoperative body weight loss (>3 kg) was evident in 62% of patients. Body weight loss reached a maximum 4.2 +/- 5.8 months after PpPD (mean, 6.0 kg) and returned to the preoperative level 4.8 months thereafter. These results suggest that PpPD has been performed safely in Japan, the operative mortality being 2.4%. However, delayed gastric emptying was evident in 46% of the patients and pancreatoenterostomy leakage in 16%. Impairment of glucose tolerance occurred in about 10% of patients more than 1 month after PpPD. Therefore, during the early postoperative period, patients should be closely monitored for pancreatoenterostomy leakage and delayed gastric emptying and in the late postoperative period, glucose tolerance should be carefully followed-up. PMID- 10526068 TI - Photodynamic therapy using mono-L-aspartyl chlorin e6 for rabbit experimental hepatoma. AB - Photodynamic therapy and photodynamic diagnosis using photosensitizers have yet to be clinically employed for hepatoma. Mono-L-aspartyl chlorin e6, a chlorin derivative with high tumor affinity developed as a second-generation photosensitizer, enables rapid tumor detection after administration, without light-shielding. This study examined the potential of photodynamic therapy, using this photosensitizer, for hepatoma in rabbits. VX2 tumor cells were transplanted into the liver of Japanese white rabbits, and the animals were administered 2.5 mg/kg of mono-L-aspartyl chlorin 36 1 week later. Accumulation of mono-L-aspartyl chlorin e6 in hepatoma was observed over time with an epifluorescence stereoscope, and the efficacy of continuous photodynamic therapy following photodynamic diagnosis was examined. A diode laser system was used for treatment, and efficacy was examined in a control group and four other groups that were irradiated at different times following administration. Efficacy in suppressing tumor growth, tumor necrosis rates, and efficacy in suppressing pulmonary metastasis were studied. For all these aspects, treatment was significantly more effective in the group irradiated 5 minutes after administration than in groups irradiated at later times. Liver function testing in all groups revealed no distinct disorders. Photodynamic therapy with mono-L-aspartyl chlorin e6 may be suitable for clinical use in therapy for hepatoma. PMID- 10526069 TI - Successful resection of cecal hepatic metastasis extending into the right side of the heart under cardiopulmonary bypass. AB - Resection is the best hope for the cure of colorectal metastasis to the liver. However, surgery is indicated for only a few patients, especially those who have major vascular involvement. We report a 55-year-old woman with a liver metastasis from the cecum that showed a tumor thrombus in the right side of the heart. She had undergone laparoscopic right hemicolectomy for cecal cancer 6 months before, and presented with a palpable mass in the epigastrium. Abdominal ultrasonography, computed tomography, hepatic angiogram, and echocardiography showed a huge mass on the left lobe of the liver, with a tumor thrombus which extended to the right ventricle through the left hepatic vein and inferior vena cava. Tumor thrombectomy, through a right atriotomy, was success-fully performed under cardiopulmonary bypass, followed by left hepatic lobectomy. The patient's postoperative course was uneventful. PMID- 10526070 TI - Hepatobiliary cystadenoma presenting with intermittent inferior vena caval obstruction. AB - Hepatobiliary cystadenomas are rare benign tumours. They form as multilocular cysts in and around the liver. Their presentation is usually at a late stage when their size causes symptoms. We report a case of a 44-year-old woman presenting with abdominal pain, which was attributed to a multiloculate liver cyst. After intially refusing surgery she relented once intermittent inferior vena caval obstruction had developed. The cyst was found to be a hepatic cystadenoma. PMID- 10526071 TI - Ventral transdural herniation of the thoracic spinal cord: surgical treatment in four cases and review of literature. AB - BACKGROUND: A specific cause of progressive Brown-Sequard syndrome has been identified: a ventral herniation of the thoracic spinal cord through the dural sleeve on one side. METHOD: Four female patients who were affected by a progressive Brown Sequard syndrome related to a transdural spinal cord herniation have been investigated and were submitted to surgery and postoperative evaluation. FINDINGS: The MRI scan showed atrophy and forward displacement of the spinal cord on one side and adhesion of the spinal cord to the dura mater. CT myelography demonstrated the disappearance of the premedullar rim at the level of the herniation and the shadow of the extradural herniation. Surgical treatment consisted in the excision of the arachnoid cyst when there was one, section of the dentate ligament, release of the adhesions, detachment of the spinal cord from the hernial orifice, and lastly suture of the dural tear or placement by a patch. Follow-up examination showed motor improvement with persistent sensory deficit in two cases and stabilisation in two cases. INTERPRETATION: The cause of the dural tear, either traumatic or congenital could not be confirmed in the four cases. Symptoms probably occur when herniation fills the orifice and strangulation happens which explains the late appearance and progressive evolution of this myelopathy. Mobilisation of the herniated spinal cord back into the intradural space can be achieved by surgery and may stop the evolution of the symptoms and signs. PMID- 10526072 TI - Instrumented facet fusion for the degenerative lumbar disorders. AB - A new, simple, minimally morbid procedure used to treat degenerative lumbar spinal disorders is described. The authors based their treatment on the McBride technique of facet fusion, which was modified and supplemented with pedicle screw fixation. The first 32 consecutive patients with degenerative spondylolisthesis and failed back surgery syndrome were treated, and followed for more than a year. Surgically, 26 patients had a single level fusion, and 6 patients had a two level fusion. Postoperative radiographs and computed tomography (CT) scans were evaluated to determine the fusion status. There were no specific complications related with the facet fusion. Thirty of 32 (93.8%) had solid fusions according to the computed tomographic criteria. Two patients showed a questionable union on CT scan but motion was less than 5 degrees on dynamic films. While 96.2% (25/26) of patients with a single level procedure had solid fusion, rate of union in the patients with two level procedures was 83.3% (5/6). This preliminary study of 32 patients shows that instrumented facet fusion appears to be a safe and effective procedure for lumbar spine fusions, demonstrating a high fusion rate with rarely serious complications. PMID- 10526073 TI - Risk factors predicting recurrence in patients operated on for intracranial meningioma. A multivariate analysis. AB - The authors undertook a follow-up study of 286 patients who underwent surgical treatment for intracranial meningioma between 1973 and 1994, in order to analyse clinical, radiological, topographic, histopathological and therapeutic factors significantly influencing tumour recurrence. All patients were followed by using either computed tomography (CT) or magnetic resonance from 3 months to 17 years since first surgery (mean follow-up: 4.1 years). Forty-four (15.4%) recurrences were detected during this time period. Overall recurrence rates were 14%, 37% and 61% at 5, 10 and 15 years, respectively. Factors significantly associated with tumour relapse in bivariate analysis were: tumour location at petroclival and parasagittal (middle third) regions, incomplete surgical resection (assessed by Simpson's classification), atypical and malignant histological types (WHO classification), presence of nucleolar prominence, presence of more than 2 mitosis per 10 high-power fields, and heterogeneous tumour contrast enhancement on the CT scan. The multivariate analysis using the Cox's proportional hazards model identified the following risk factors for recurrence: incomplete surgical resection (Relative risk: 2.2; 95% Confidence interval: 1.33-3.64), non conventional histological type (RR: 2.13; 95%CI: 1-4.53), heterogeneous contrast enhancement on the CT scan (RR: 2.25; 95%CI: 1.1-4.72) and presence of more than 2 mitosis per 10 high-power fields (RR: 2.28; 95%CI: 0.99-5.27). Patients without any of these features showed low recurrence rates (4% and 18% at 5 and 10 years), and thus, they need less clinical and radiological controls through the follow-up than patients with some of these risk factors. PMID- 10526074 TI - Stereotactic irradiation of skull base meningiomas with high energy protons. AB - Nineteen patients with inextirpable skull base meningioma with involvement of neurovascular structures were given irradiation with a 180 MeV proton beam at the The Svedberg Laboratory, Uppsala, Sweden. The patients were treated seated in a fixed position with a stereotactic approach. Titanium-markers to the outer table served for identification and verification of the target positioning for dose planning and irradiation. The patients were given a total dose of 24 Gy in four consecutive daily 6 Gy fractions. All patients have been followed for at least 36 months. So far no meningiomas have progressed after treatment. Two patients have developed corticosteroid responsive oedema in the target area 6 moths after treatment. Late, but not serious, symptoms of side effects have been observed in one patient. PMID- 10526075 TI - Diagnostic in normal pressure hydrocephalus: A mathematical model for determination of the ICP-dependent resistance and compliance. AB - The internationally accepted calculation methods concerning cerebrospinal fluid dynamics proceed from a pressure independent resistance to cerebrospinal fluid outflow. In a new model we focus our attention on the pressure dependency of resistance. In our calculation model we are monitoring the complete pressure course p(t) over the time during and after the infusion. The comparison of the pressure rise On(p) during the infusion and the descent Off(p) after the infusion at the same pressure level allows one to construct all formulas for the compliance C(p) and resistance R(p). The computerized analysis of the results of the intrathecal infusion test using our mathematical computation leads to a simplification of this investigation. The simultaneous measurement of the resistance and compliance during a single investigation allows one to minimize the patient's discomfort. In contrast to the classical methods it is not necessary that the ICP reaches a plateau. Our mathematical method diverges with the description of a pressure dependent slope of the function for the resistance from the static examination models. For that we are able to take the non linearity of the cerebrospinal fluid resorption into consideration. PMID- 10526076 TI - Expansive suboccipital cranioplasty for the treatment of syringomyelia associated with Chiari malformation. AB - In order to treat syringomyelia associated with adult type Chiari malformation, the authors developed a method of expansive suboccipital cranioplasty (ESC) that involves enlarging the small posterior fossa to obtain a sufficient flow of cerebrospinal fluid (CSF). The relative effectiveness of ESC with the obex plugged and not plugged was also examined, as well as other factors influencing the operative results. Twenty patients without arachnoid adhesion at the major cistern underwent ESC without opening the arachnoid membrane at the major cistern. After surgery, all improved with no recurrence and CSF flow study using magnetic resonance (MR) imaging showed significant improvement of the flow at the major cistern. Another 20 patients without arachnoid adhesion also underwent ESC but with obex plugging. Sixteen improved and one displayed only temporary improvement with recurrent syringomyelia due to postoperative arachnoid adhesions. The remaining three showed no change in spite of shrinkage of the syrinx on postoperative MR imaging. These three patients had displayed pre operative symptoms over an approximately 10-year period involving almost the entire axial plain of the spinal cord, and presented a large syrinx before surgery. In 4 patients with arachnoid adhesions, all required intra-arachnoid procedures in addition to ESC. Intra-arachnoid procedures are not necessary to facilitate restoration of CSF flow in patients without arachnoid adhesions, because ESC can release the CSF flow blockage in the major cistern even without plugging of the obex. An associated arachnoid adhesion at the major cistern or a long-standing syringomyelia with irreversible damage of the spinal cord results in a poor operative prognosis. When posterior fossa surgery fails, insufficient decompression or postoperative arachnoid adhesions at the major cistern as the cause of treatment's failure should be evaluated by CSF flow studies using phase contrast MR imaging. PMID- 10526077 TI - Surgical treatment of Chiari I malformation with or without syringomyelia. AB - 102 patients with Chiari I malformation with or without syringomyelia underwent primary reconstructive operation at the craniocervical junction (CCJ). We present here the indications, methods of surgical management with clinical, radiological and operative findings. Here we present the results of a retrospective review. The authors discuss the possible ways of improvement in the results of "hindbrain related" syrngomyelia treatment. PMID- 10526078 TI - Syringobulbia caused by delayed postoperative tethering of the cervical spinal cord - delayed complication of foramen magnum decompression for Chiari malformation. AB - Postoperative tethering of the high cervical spinal cord is a rare cause of neurological deterioration after foramen magnum decompression (FMD) with duraplasty for Chiari type I malformation. A review of the literature revealed that only 5 cases have been reported. This entity is not widely known to occur as a complication of the common surgical procedure for Chiari type I malformation. A 17-year-old boy experienced rapidly progressive neurological deterioration over a 3-month period. FMD and duraplasty with lyophilized cadaver dura had been performed 8 years previously. Follow-up MR images showed that the cerebrospinal fluid (CSF) space dorsal to the cord was gradually disappearing and that syringobulbia had developed. Opening the dura mater of the posterior fossa revealed dense fibrous scarring, arachnoid thickening over the cervicomedullary area, and tethering the cord to the dura from the medulla to C2. The adhesions were dissected free, and the tethering was released. A syringosubarachnoid (SS) shunt was inserted and duraplasty was performed with an expended polytetrafluoroethylene sheet (Gore-Tex). Postoperative MR images demonstrated that the syringobulbia had completely collapsed and that a dorsal CSF space was present. Follow-up MR images provided significant information on the cervical spinal cord tethering after FMD with duraplasty for Chiari malformation. We encourage sharp surgical detethering and duraplasty with Gore-Tex to avoid retethering. Early recognition and treatment of this unusual but important complication are emphasized. PMID- 10526079 TI - Cerebral germinoma with syncytiotrophoblastic giant cells: feasibility of predicting prognosis using the serum hCG level. AB - As the biological behaviour of germinoma with syncytiotrophoblastic giant cells (STGC) is not well established, the present study was undertaken to ascertain the prognostic significance of serum hCG level in affected patients. Of a total of 23 cases studied, 12 patients were regarded as pure germinomas and 11 were germinomas with STGC. All but one of the former demonstrated an excellent outcome. The exception developed subarachnoid metastases, but the tumour disappeared on radiation therapy and the patient is enjoying a normal social life 13 years after the initial treatment. With the germinoma complicated by STGC, 3 cases showed local recurrence which were followed by a poor outcome. Their pretreatment hCG levels were 15.0, 26.0 and 29.6 mIU/ml respectively. The study showed a tendency, in germinomas with STGC, for a positive association between serum hCG, and the likelihood of a poor outcome. Germinomas with STGC and serum hCG levels higher than 15 mIU/ml thus have a high recurrence rate, and more aggressive treatment is indicated for the affected patients. PMID- 10526080 TI - The effect of mild hypothermia, mannitol and insulin-induced hypoglycaemia on ischaemic infarct volume in the early period after permanent middle cerebral artery occlusion in the rat. AB - We investigated the effect of mild hypothermia (32-34 degrees C), mannitol and insulin - induced hypoglycaemia on the ischaemic infarct volume on permanent middle cerebral artery occlusion with bilateral carotid artery ligation in rats. Temporalis muscle temperature as an indicator of brain temperature was monitored throughout the experiment in all rats, which were randomly divided into seven groups. During ischaemia, control rats received intravenous saline in a normothermic condition; treated rats had hypothermia and intravenous saline, hypothermia and mannitol, normothermia and mannitol, normothermia and insulin, normothermia, insulin and glucose, and hypothermia and insulin applied. After the experiment, the animals were killed, and brain sections were stained with haematoxylin and eosin. Images of infarct areas were determined using a camera attached to the microscope, and analysed by image analysis software. The total volume of infarcted tissue, right hemispheric volume, and the percentage of infarction were determined at the end of the image analysis investigation. The infarct volume on the control group was found to be 128.16+/-6.67 mm(3). Infarct volumes in hypothermic groups were significantly smaller than those of the control group (p<0.05). There were no significant differences between infarct volumes in the hypothermic groups. However, we found that hypothermia plus mannitol have the greatest neuro-protective effect. In normothermic rats, the infarct volume decreased proportionally but not statistically (p>0.05) whether mannitol or insulin was given. Our results also demonstrate that pre-, and post ischaemic serum glucose concentrations influence the volume of infarction. Rats that had had pre-ischaemic high serum glucose concentrations had a higher volume of infarct than the hypothermic rats (p<0.05), while rats with post-ischaemic low serum glucose concentrations had a lower volume of infarct than the control rats. PMID- 10526081 TI - Effect of acrylamide on neurological recovery following spinal cord injury in rats. AB - Acrylamide (ACR) is a cumulative neurotoxin which causes axonal degeneration in animals and man. Industrial workers exposed to ACR have been reported to suffer from a variety of central and peripheral neuropathological symptoms including numbness of hands and feet, skin peeling and muscular weakness of legs. These reports suggest that the body burden of ACR may be a risk factor in recovery patterns following neurotrauma. The present study was designed to assess the effect of ACR on neurological recovery following spinal cord injury (SCI) in rats. Male Sprague-Dawley rats weighing 200-230 g were anaesthetised with chloral hydrate and laminectomy was performed at T 7-8 level leaving the dura intact. A compression plate (2.2 x 5.0 mm) loaded with a weight of 35 g was placed on the exposed cord for 5 minutes. Animals were divided into seven groups of eight rats each. The animals in Group 1 served as control whereas rats in Group 2 underwent laminectomy alone (sham). The rats in Group 3 to 6 were subjected to SCI as mentioned above. Animals in Groups 4, 5 and 6 also received ACR in the doses of 10 mg/kg, 20 mg/kg and 40 mg/kg, i.p., respectively in addition to SCI, whereas the rats in Group 7 received ACR alone at a dose of 40 mg/kg body weight. The first dose of ACR was given 30 minutes before SCI, followed by daily administration of drug for 7 days. Post traumatic neurological recovery was recorded daily for 10 days using a modified Tarlov score, inclined plane test and sensory and vocal score. Electrophysiological changes were assessed using somatosensory and corticomotor evoked potentials. The animals were sacrificed at different time intervals and the injured site of the spinal cord was analysed for lipid hydroperoxides (LPH), conjugated dienes (CD) and glutathione (GSH). Neuropathological changes in the spinal cord were assessed using light microscopy. The rats exposed to compression injury alone showed a maximum neurological deficit at 24 hr and then a gradual recovery was observed over a period of 10 days. The rats treated with ACR along with SCI showed poor or no recovery over a period of 10 days. Our electrophysiological and histopathological studies also confirmed that concomitant exposure to ACR produces a significant deleterious effect on the recovery from SCI. SCI induced increase in oxidative stress (increase in LPH and CD and decrease in GSH) is also exacerbated by ACR suggesting a role of free radicals. The results of this study suggest that increased body burden of ACR may retard the recovery from neurotrauma or even lead to permanent disability. PMID- 10526082 TI - Ossifying pituitary gonadotroph adenoma: A case report. AB - BACKGROUND: Bone formation in pituitary adenomas is a rare finding. Only two previous cases were published, occurring in a prolactin- producing and a growth hormone- producing pituitary adenoma respectively, both in pre-menopausal women. CLINICAL MATERIAL: This is the third report of an ossified pituitary adenoma and the first report of a pituitary gonadotroph adenoma with bone formation occurring in an elderly man. MRI imaging revealed an unusual eggshell cap-like calcified structure surrounding the tumor. Histologically, the adenoma contained irregularly anastomosing trabecules with well formed lacunae and osteoblasts along the margins. CONCLUSIONS: Insufficient tumor blood supply may trigger proliferation of connective tissue that subsequently undergoes osteoid metaplasia. Pituitary adenoma with osteoid metaplasia should be included in the differential diagnosis of calcifying tumors in the sella region. PMID- 10526083 TI - Dissection of the middle cerebral artery caused by invasion of malignant glioma presenting as acute onset of hemiplegia. AB - A 57-year-old, previously healthy man who developed acute onset of hemiplegia is presented. Neuro-imaging studies on admission suggested dissection of the middle cerebral artery producing infarction in the frontotemporal region. In contrast to his stable clinical course, serial neuro-imaging studies disclosed rapid growth of malignant glioma, which was confirmed at surgery. Microscopic examination of the surgical specimen demonstrated invasion of glioma cells into the arterial wall associated with intramural haematoma formation of the middle cerebral artery. This case is the first to document dissection of an intracranial artery caused by invasion of tumour cells. PMID- 10526084 TI - Subarachnoid haemorrhage and vasospasm due to pituitary apoplexy after pituitary function tests. PMID- 10526085 TI - Metastasis along the stereotactic biopsy trajectory in glioblastoma multiforme. PMID- 10526086 TI - C5a receptor antagonists. AB - The anaphylatoxin C5a is an extremely potent proinflammatory peptide produced during activation of the complement system. The structure of C5a includes a core region (N-terminal residues 1-63) consisting of four, antiparallel alpha-helices held together by three disulfide linkages and a structured C-terminal tail (residues 64-74). The C5a receptor belongs to the large class of seven transmembrane, G-protein-linked receptors. C5a appears to interact with its receptor at two sites: the C5a core binds to the receptor s N-terminal extracellular domain while C5a s tail binds the receptor near Arg206, near the membrane surface of transmembrane helix V. C5a receptors are concentrated on blood granulocytes (neutrophils, eosinophils, and basophils) and tissue inflammatory cells (macrophages, mast cells, microglia); thus the main effects of C5a are manifest as inflammation. Additionally, C5a receptors are also present, albeit in lower concentrations, on non-myeloid cells, e.g. endothelial and smooth muscle cells where they may further influence inflammatory reactions such as blood cell emigration and tissue edema. C5a has been implicated in myriad disorders, both acute and chronic; therefore a C5a receptor antagonist is predicted to have utility as a therapeutic agent. Unfortunately, few specific C5a receptor antagonists have been reported, and only two have demonstrated activity in vivo. Furthermore, those reported are peptidic and hence have limited application therapeutically. The current state of C5a receptor antagonists is discussed as well as the potential for their use against various human disorders. A model of C5a receptor dimerization is presented to account for the high potency of the disulfide antagonist C5aRAD. PMID- 10526087 TI - Interferons and their role In inflammation. AB - Cytokines are pleiotropic molecules showing a wide variety of biologic functions on various cells and tissues, and several different cytokines exert similar and overlapping functions on certain cells. Interferons (IFNs), among the first cytokines identified, play a crucial role in human disease. The IFN cytokine family consists of type I IFNs (IFN-a and IFN-b) and type II IFN (IFN-g). In the first decades of IFN research, type I IFNs were considered primarily as viral inhibitors, whereas type II IFN, also termed "immune IFN", was generally considered to be uniquely involved in immune reactions. This view has changed considerably in the last years. The importance of type I IFNs in inflammation, immunoregulation and T-cell responses has been identified and has changed dramatically our interpretation of the biological relevance of type I and II IFNs. Recent data suggest that IFN-a is a multifunctional immunomodulatory cytokine with profound effects on the cytokine cascade including several anti inflammatory properties, whereas IFN-g remains a classical proinflammatory cytokine. These different effects on critical mediators of inflammation may also explain why type I and II IFNs are clinically successful in different diseases. These newly identified immunoregulatory and anti-inflammatory functions of type I IFNs may be of importance in the treatment of diseases such as chronic viral hepatitis or multiple sclerosis and help to explain some of the mechanisms of IFNs. PMID- 10526088 TI - Brain inflammation in Alzheimer disease and the therapeutic implications. AB - Immunohistochemical studies suggested the existence of a chronic inflammatory condition in affected regions of the brain in Alzheimer disease (AD). Since inflammation can be damaging to host tissue, it was hypothesized that antiinflammatory drugs might inhibit both the onset and the progression of AD. This hypothesis is supported by a number of epidemiological studies suggesting that the prevalence of AD in persons is reduced by 40 - 50% in persons using antiinflammatory drugs. In one small pilot trial in early AD, the nonsteroidal antiinflammatory drug indomethacin appeared to halt the progressive memory loss. Immunohistochemical and molecular biological studies on immune system components in AD brain are revealing the complexities of the innate immune reaction. This very complexity may offer points of therapeutic intervention for new types of antiinflammatory agents. The complement system, microglia and cytokines are key components. This review summarizes the present state of knowledge on the immune system elements found in AD brain. PMID- 10526089 TI - Pharmacological characterization of morphine-6-sulfate and codeine-6-sulfate. AB - Morphine-6-sulfate (M6S) and codeine-6-sulfate (C6S) are mu-selective opiates which have been isolated from brain. M6S is an effective analgesic, with a 30 fold greater potency than morphine in the mouse radiant heat tailflick assay and similar to the active morphine metabolite morphine-6beta-glucuronide (M6G). M6S analgesia is reversed by 3-methoxynaltrexone at low antagonist doses which are inactive against morphine, suggesting that M6S may be acting through the same mechanisms as M6G. Consistent with this possibility, antisense mapping of the MOR 1 clone revealed that M6S analgesia was lowered by probes targeting exon 2 and not by targeting exon 1, a sensitivity profile similar to that of M6G and not morphine. C6S also has analgesic activity at doses approximately 10-fold greater than M6S. However, its characterization was impeded by the appearance of seizures at doses below full analgesic activity. Thus, M6S is a potent analgesic with pharmacological properties similar to M6G. C6S has limited utility due to its high level of toxicity. PMID- 10526090 TI - Alterations in monoamines and GABA in the ventromedial and paraventricular nuclei of the hypothalamus following cold exposure: a reduction in noradrenaline induces hyperphagia. AB - A microdialysis technique for the in vivo assessment of the monoaminergic and GABAergic levels in the ventromedial (VMN) and paraventricular nuclei (PVN) of the hypothalamus was used in order to examine the activities of neurons that project to the hypothalamic regions and are implicated in the regulation of ingestive behavior and energy balance. Cold exposure increased food intake, as well as the circulating levels of glucose, noradrenaline (NA), 3,5,3' triiodothyronine, and corticosterone. The dialysate concentrations of NA, dopamine (DA), serotonin (5-HT), and gamma-aminobutyric acid (GABA) in the VMN all decreased after exposure to cold. The changes in extracellular NA, DA, and GABA in the PVN under cold conditions were similar to those in the VMN. NA release in the VMN or PVN was decreased after local electrolytic lesions, which significantly increased food intake. Thus, low activities of noradrenergic axons of neurons terminating in the VMN and PVN may be a good mechanism to induce feeding behavior. Extracellular 5-HT in the PVN was significantly increased, along with a significant decrease in 5-hydroxyindoleacetic acid, in cold-exposed rats, suggesting that serotonergic fibers terminating in the PVM are more closely related to the increases in adrenocortical and thyroid hormone secretion than to food intake. The neuronal activities, indicating that a sympathetic tone is activated on stimulation of the VMN and/or PVN, may be changes in GABAergic and/or serotonergic neurons. PMID- 10526091 TI - Cortical neurons immunoreactive for the potassium channel Kv3.1b subunit are predominantly surrounded by perineuronal nets presumed as a buffering system for cations. AB - Perineuronal nets (PNs) are known as chondroitin sulphate-rich, lattice-like coatings of the extracellular matrix. In the cortex of mammalian species investigated so far, they were mainly found around GABAergic neurons, but to a lesser degree also around pyramidal cells. Previous investigations in the rat revealed similar distribution patterns of fast-firing neurons expressing both the Kv3.1b subunit of voltage-gated potassium channels and the calcium-binding protein parvalbumin. In the present study, triple fluorescence labelling was applied for the simultaneous demonstration of PNs with the N-acetylgalactosamine specific Wisteria floribunda agglutinin (WFA), parvalbumin-immunoreactivity (ir) with a monoclonal antibody and of Kv3.1b-ir with several rabbit antibodies. Subsets of non-pyramidal neurons - enwrapped by PNs and expressing parvalbumin and Kv3.1b - were detected in the rat and monkey neocortex and hippocampus. In the rat, faintly stained PNs were additionally found around several layer II/III and V pyramidal cells immunonegative for Kv3.1b, but contacted by Kv3.1b containing boutons. In the monkey, more intensely labelled PNs frequently occurred around pyramidal cells which themselves appeared to be Kv3. 1b immunopositive. We also observed minor Kv3.1b-ir and parvalbumin-ir cortical cell populations which were devoid of PNs; occasionally, nets were detected around neurons lacking both immunoreactivities. By confocal laser scanning microscopy, Kv3.1b-ir and WFA-binding sites were found adjoining at the soma and proximal dendritic surface, while lectin-binding sites usually extended on more distal dendritic segments and the axon initial segments which failed to express detectable Kv3.1b-ir. This spatial relationship of both markers was also confirmed by combined WFA-gold labelling and Kv3.1b-immunoperoxidase staining at the electron microscopic level. The data are used for a critical examination of current hypotheses concerning the functional role of PNs. We conclude that PNs may serve as rapid local buffers of excess cation changes in the extracellular space. Somatic membranes of fast-spiking neurons seem to be a main, but not the only source of such changes. PMID- 10526092 TI - Cross-modal selective attention to visual and auditory stimuli modulates endogenous ERP components. AB - In two experiments event-related potentials (ERPs) to visual and auditory stimuli were measured in 12 healthy subjects. A cross-modal and delayed response paradigm was used that allows ERPs to be obtained separately to attended and unattended stimuli under conditions in which unattended stimuli are less likely to be covertly or randomly attended. The results showed: (1) N1 enhancement with attention for standard stimuli in auditory and visual modalities and for deviant stimuli in the visual modality; (2) The onset time and scalp distribution of both the N1 for attend condition and Nd1 were similar regardless of standard or deviant stimuli in the auditory and visual modality; the onset time of Nd1 elicited by auditory and visual deviant stimuli was earlier than that of the unattended N1, and their scalp distributions were different; and (3) The Nd1 components elicited by auditory and visual deviant stimuli were distributed over the respective primary sensory areas, but Nd1 components evoked by auditory and visual standard stimuli were distributed over the frontal scalp. These results suggest that the attended N1 enhancement is primarily caused by a component with endogenous origins and that the early attention effect occurs before the exogenous components. The results support the view that the cross-modal attention to deviant stimuli modulates modality-specific processing in the brain, whereas attention to standard stimuli affects modality-nonspecific or supramodal brain systems. PMID- 10526093 TI - Intracellular ATP depletion inhibits swelling-induced D-[3H]aspartate release from primary astrocyte cultures. AB - Volume expansion-sensing outward rectifier (VSOR) anion channel, also referred to as volume-sensitive organic osmolyte-anion channel (VSOAC), appears to be responsible for cell swelling-induced amino acid release in a variety of cells. One prominent feature of the VSOR/VSOAC is that non-hydrolyzed intracellular ATP binding to the channel or an accessory protein is required for its activation. In this study, the effect of intracellular ATP depletion on the swelling-induced release of D-[3H]aspartate from rat primary astrocyte cultures due to exposure to either high K(+) or hypotonic media was studied. When the cells were pretreated for 10 min with a combination of the metabolic inhibitors 2-deoxyglucose and rotenone, 100 mM K(+) media- or hypotonic media-induced D-[3H]aspartate release was completely suppressed. Added separately, each inhibitor showed only partial or no inhibition of D-[3H]aspartate release, which correlated with its relative effectiveness in decreasing intracellular ATP levels. These data are consistent with the view that during high [K(+)](o) or hypotonic media-induced swelling of primary astrocyte cultures an ATP-dependent swelling-activated VSOAC channel is responsible for D-[3H]aspartate release and close to normal ATP is required for full channel activation. PMID- 10526094 TI - Glial-derived proteins activate cultured astrocytes and enhance beta amyloid induced glial activation. AB - A prominent feature of Alzheimer's disease (AD) pathology is an abundance of activated glia (astrocytes and microglia) in close proximity to the amyloid plaques. These activated glia overexpress a number of proteins that may participate in the progression of the disease, possibly by propagation of inflammatory and oxidative stress responses. The beta-amyloid peptide 1-42 (Abeta), a major constituent of neuritic plaques, can itself induce glial activation. However, little is known about whether other plaque components, especially the upregulated glial proteins, can induce glial activation or modulate the effects of Abeta on glia. In this study, we focused on four glial proteins that are abundant in amyloid plaques and/or that are known to interact with Abeta: alpha1-antichymotrypsin (ACT), interleukin-1beta (IL-1beta), S100beta, and butyrylcholinesterase (BChE). We examined the ability of these proteins to activate rat cortical astrocyte cultures and to influence the ability of Abeta to activate astrocytes. Treatment of astrocytes with ACT, IL-1beta, or S100beta resulted in glial activation, as assessed by reactive morphology, upregulation of IL-1beta, and production of inducible nitric oxide synthase and nitric oxide. The ability of Abeta to induce astrocyte activation was also enhanced in the presence of each of these three proteins. In contrast, BChE alone did not activate astrocytes and had no effect on Abeta-induced activation. These results suggest that certain proteins produced by activated glia may contribute to the chronic glial activation seen in AD through their ability to stimulate astrocytes directly or through their ability to modulate Abeta-induced activation. PMID- 10526095 TI - Interactions of the systemic and brain renin-angiotensin systems in the control of drinking and the central mediation of pressor responses. AB - Most of the biological actions of the circulating (a.k.a., the systemic or blood borne) renin-angiotensin system require the generation of the octapeptide angiotensin (ANG) II from the decapeptide ANG I. In the case of circulating ANG I, the lungs are generally considered the major site for this conversion. The present experiments explored the possibility that under conditions of marked elevations of blood-borne ANG I, the generation of ANG II takes place within brain-associated target tissues, most notably circumventricular organs (CVOs) that lack a blood-brain barrier. The first important result of these experiments demonstrates that intracerebroventricular (i.c.v.) infusion of the converting enzyme inhibitor, captopril, completely blocks the drinking response and significantly attenuates the pressor response produced by systemically infused ANG I. This result indicates that under physiological/pathophysiological conditions associated with large elevations of circulating ANG I, an important part of the biological responses derived from blood-borne ANG may result from local conversion of ANG I to ANG II within specific brain target tissues which have high concentrations of converting enzyme. This local conversion process provides an important mechanism that would act to reinforce the "classic" conversion process which takes place in the lungs thereby delivering more ANG II immediately to central target receptors. The second important finding from these studies showed that drinking produced by systemically infused ANG II was not attenuated by an i.c.v. dose of captopril which was effective in blocking a comparable dipsogenic response induced by i.v. ANG I. This observation suggests that drinking induced by systemic ANG II does not require an intact metabolic cascade within the brain for the formation of ANG II (or ANG II-like effector peptide) from ANG I. PMID- 10526096 TI - A vulnerable period of colchicine toxicity during goldfish optic nerve regeneration. AB - The effects of intraocular (i.o.) administration of the alkaloid colchicine on visual recovery following axotomy of the goldfish optic nerve were investigated. Under the experimental conditions used, control goldfish recovered vision, measured behaviorally, within 5-7 weeks of retro-orbital optic nerve crush. Fish injected i. o. with 0.1 microg of colchicine within 3 days of optic nerve crush (post-crush; PC) recovered vision after some delay relative to control fish, while injection with colchicine between 7 and 14 days PC produced a much more profound inhibition of recovery of vision, in most cases a complete block for the duration of the study (98 days). Further evidence for a delayed susceptibility of the regenerating optic nerve to colchicine following crush was reflected in a suppression of neurite outgrowth normally seen in explanted retinal tissue taken from PC goldfish. In addition, retrograde transport of the fluorescent dye 4-(4 didecylaminostyryl)-N-methylpyridinium iodide from the optic tectum to the retina as a measure of axonal continuity revealed substantially less labeling following i.o. administration of colchicine 1 week PC when compared to retinas from fish receiving colchicine at the time of optic nerve crush. Histological sections of the retina showed no evidence of residual retinal damage resulting from the colchicine injections or from interactions of axotomy and the drug administration. These results indicate a period of increased vulnerability of the regenerating visual system to the toxic effects of i.o. administered colchicine, beginning 3-5 days PC, and remaining until regenerating optic nerve fibers have begun to reach the tectum. While colchicine has many known effects on nerve function, it is proposed that the delayed susceptibility to disruption of regeneration observed in these experiments is largely, if not entirely, attributable to a colchicine-induced accumulation of tubulin heterodimers, which are known to block microtubule assembly and to participate in a feedback inhibition of tubulin synthesis. Thus, it is during the maximal induction of tubulin synthesis and of microtubule formation which normally occurs several days following axotomy that colchicine has its greatest effect. The results suggest that colchicine may be especially neurotoxic during neural development and regeneration. PMID- 10526097 TI - Neonatal monosodium glutamate alters circadian organization of feeding, food anticipatory activity and photic masking in the rat. AB - In rodents, parenteral administration of monosodium glutamate (MSG) induces marked degeneration of the retina and arcuate nucleus (AN) and disrupts daily rhythms of food intake. We quantified the effects of neonatal MSG (2 mg/g SC, postnatal days 1, 3, 5, 7, 9) on the expression of feeding and activity rhythms in adult rats under schedules of light-dark (LD), constant dark (DD), restricted daily feeding and total food deprivation. AN lesions were confirmed by neuropeptide Y (NPY) immunocytochemistry and Nissl stain. Compared to age-matched control rats, the amplitude (quantified as LD ratios) of daily food intake and food-bin activity rhythms was significantly attenuated in MSG rats in LD 12:12 and on the first day of DD. Control rats, but not MSG rats, showed lower amplitude rhythms in DD compared to LD. The phase angle of feeding and activity rhythms did not differ between groups in either condition. In a short LD cycle (2:2), control rats, but not MSG rats, showed significant inhibition (masking) of activity during the 2 h light periods. When food access was restricted to a 4 h daily meal, MSG rats showed enhanced expression and persistence of food-entrained anticipatory activity rhythms by comparison with control rats. These results indicate that attenuation of daily feeding rhythms in MSG rats is due in part to loss of direct inhibitory effects of light on behavior, and that the AN likely modulates, but does not mediate entrainment of feeding-related rhythms to daily cycles of LD or food access. PMID- 10526098 TI - The effect of streptozotocin-induced diabetes mellitus on substance P and calcitonin gene-related peptide expression in the rat trigeminal ganglion. AB - Substance P (SP) and calcitonin gene-related peptide (CGRP) constitute the main sensory peptides in the trigeminal ganglion (TG). The objective of this study was to characterize peptidergic changes in the streptozotocin-induced diabetes mellitus rat model both quantitatively and qualitatively. Diabetes mellitus was induced by a single intraperitoneal injection of streptozotocin (65 mg/kg) and the levels of SP and CGRP were measured by means of radioimmunoassay (RIA) in a time-dependent manner. Peptide immunoreactivities were characterized by high pressure liquid chromatography (HPLC). The expression of both neuropeptides was examined 5 weeks after streptozotocin injection using in situ hybridization with 35S-labelled oligonucleotides. Saline-injected rats served as controls. SP was significantly decreased in the diabetic rat TG, i.e. , a 44.6% (+/-10.9) decrease after 1 week, 40.2% (+/-11.8) after 3 weeks and 72.3% (+/-14.6) after 5 weeks. CGRP was decreased only after 5 weeks (19.6% decrease +/-3.9), whereas at later stages, both peptide levels returned to normal values. HPLC revealed one major peak coeluting with the synthetic peptides. By using in situ hybridization, a significantly increased signal of both peptide-encoding mRNAs was found (43.8%), which seems to act to restore a diabetes-associated depletion of neuropeptides in the diabetic rat TG. The decreased SP- and CGRP levels in the diabetic rat TG reflect a diabetes-associated deficit which may be clinically relevant. Diabetes mellitus is associated with a variety of ocular complications, even corneal complications, including decreased corneal sensitivity, which in many ways resemble those after interruption of the normal trophic innervation of the eye. Our results point to reduced availability of neuropeptides for corneal innervation and may thus support the idea of a partial loss of trophic influences from the trigeminal nerve in diabetics. PMID- 10526099 TI - Early appearance of activated matrix metalloproteinase-9 and blood-brain barrier disruption in mice after focal cerebral ischemia and reperfusion. AB - Blood-brain barrier (BBB) disruption is thought to play a critical role in the pathophysiology of ischemia/reperfusion. Matrix metalloproteinases (MMPs) are a family of proteolytic enzymes that can degrade all the components of the extracellular matrix when they are activated. Gelatinase A (MMP-2) and gelatinase B (MMP-9) are able to digest the endothelial basal lamina, which plays a major role in maintaining BBB impermeability. The present study examined the expression and activation of gelatinases before and after transient focal cerebral ischemia (FCI) in mice. Adult male CD1 mice were subjected to 60 min FCI and reperfusion. Zymography was performed from 1 to 23 h after reperfusion using the protein extraction method with detergent extraction and affinity-support purification. MMP-9 expression was also examined by both immunohistochemistry and Western blot analysis, and tissue inhibitors to metalloproteinase-1 was measured by reverse zymography. The BBB opening was evaluated by the Evans blue extravasation method. The 88-kDa activated MMP-9 was absent from the control specimens, while it appeared 3 h after transient ischemia by zymography. At this time point, the BBB permeability alteration was detected in the ischemic brain. Both pro-MMP-9 (96 kDa) and pro-MMP-2 (72 kDa) were seen in the control specimens, and were markedly increased after FCI. A significant induction of MMP-9 was confirmed by both immunohistochemistry and Western blot analysis. The early appearance of activated MMP-9, associated with evidence of BBB permeability alteration, suggests that activation of MMP-9 contributes to the early formation of vasogenic edema after transient FCI. PMID- 10526100 TI - Synaptic excitability of the burst firing neurons in cat sensorimotor cortex in vitro. AB - We have recently reported that the burst firing neurons are found in layer III as well as in layer V of cat sensorimotor cortex in vitro. In the present study, we examined the synaptic excitability of layer III neurons by white matter stimulation and compared with their firing patterns against the current injections through the recording microelectrodes. The firing patterns of layer III neurons were classified into three main classes as in our previous study, i.e., (1) regular spiking (RS), i.e., the tonic firing that often exhibited spike frequency adaptation, (2) burst-and-single spiking (BS), i.e., the initial bursting followed by tonic firing, (3) repetitive-bursting (RB), the burst firing that recurred at fast frequency. In RS cells, single action potential was superimposed on the largest EPSPs among all cell types analyzed. BS cells also fired single action potential and never exhibited burst firing synaptically. Only in a part of RB cells, synaptic bursting instead of single action potential was evoked on smaller EPSPs. IPSPs could be observed in about 60% of all the recorded RS and BS cells, however, they were observed in only 10% of the RB cells. PMID- 10526101 TI - Systemic treatment with GPI 1046 improves spatial memory and reverses cholinergic neuron atrophy in the medial septal nucleus of aged mice. AB - Systemic treatment with GPI 1046, a non-immunosuppressive ligand of the immunophilin FKBP12 (FK-506-binding protein 12 kDa), has previously been shown to promote morphological recovery of the nigrostriatal dopaminergic projection after MPTP lesion in mice, and of lesioned sciatic nerve fibres after nerve crush in rats. In the present study, we investigated whether chronic systemic treatment with GPI 1046 could affect the decline of spatial learning and memory, and the atrophy of medial septal cholinergic neurons, associated with late senescence in C57 black mice. Three-month old (young) and 18-19-month old (aged) male C57BL/6N Nia mice were first trained in a place learning task in the Morris water maze. Based on their performance relative to young controls, aged animals were then allocated to treatment groups (10 mg/kg GPI 1046, or vehicle). Retention of the spatial platform location was assessed after 3 weeks of dosing. We found that aged animals that had been dosed with GPI 1046 now performed at a significantly better level than their vehicle control group. Aged animals that had shown the greatest degree of impairment during training in the place learning task showed the greatest relative degree of improvement under treatment and were statistically indistinguishable from young, or aged unimpaired control animals. Cell volumes of cholinergic cells in the medial septal nucleus were assessed after an additional 10 months of dosing at 30 months of age, using stereological methods. We found that aged animals displayed a significant 34% decrease in volume of these cells relative to young controls. This atrophy was significantly reversed in aged GPI 1046-treated animals (13% shrinkage). We conclude that chronic systemic treatment with GPI 1046 positively affects memory mechanisms in the aged mouse, possibly by acting on the septohippocampal cholinergic system. PMID- 10526102 TI - Recovery after nigral grafting in 6-hydroxydopamine lesioned rats is due to graft function and not significantly influenced by the remaining ipsilateral or contralateral host dopaminergic system. AB - The aim of this study was to evaluate whether the recovery observed after grafting of fetal nigral cells in 6-hydroxydopamine lesioned rats is due to the graft itself, and whether the participation of the remaining host dopaminergic system is necessary. The effects of unilateral 6-hydroxydopamine lesion on rotational behavior were not significantly affected by sham grafting or by sham grafting plus repeat ipsilateral lesion, but were suppressed by nigral grafting, and by contralateral lesion. Immunohistochemical and in situ hybridization study of right striata of rats subjected to right-side lesion then right-side sham grafting, and of right and left striata from rats subjected to right-side lesion then right-side sham-grafting then repeat right-side lesion then left-side lesion, revealed (a) no significant amphetamine-induced Fos activation, (b) marked increases in preproenkephalin mRNA levels, and (c) decreases in preprotachykinin levels, with no significant differences in any of these variables among these three types of striata. After nigral grafting, however, intense Fos expression was observed in the striatum, and preproenkephalin and preproenkephalin mRNA levels returned to normal. This recovery was maintained after subsequent repeat ipsilateral 6-hydroxydopamine lesion followed by contralateral lesion. The results demonstrate that, after dopaminergic denervation, the nigral graft itself is able to induce recovery in the assessed parameters, and that these effects of grafting into striata with maximal unilateral 6-hydroxydopamine lesion are due to graft function, and are not significantly influenced by the remaining ipsilateral or contralateral host dopaminergic system. Additionally, it is interesting to note that bilateral denervation led to changes in striatal preproenkephalin and preproenkephalin mRNA levels similar to those observed after unilateral lesion. PMID- 10526103 TI - Region-specific modulation of limbic seizure susceptibility by ovarian steroids. AB - Gonadal steroid hormones can markedly affect seizure susceptibility. Ovariohysterectomized female rats given ovarian steroid hormone supplements were used to evaluate the effects of ovarian steroids on epileptiform activity in hippocampal slices in vitro and on flurothyl-induced seizures in vivo. Seizure susceptibility was compared in the entorhinal cortex (EC) and CA1 regions of the hippocampus perfused with Mg(2+)-free medium, which leads to epileptiform discharges caused by a relief of voltage-dependent NMDA receptor block. After in vivo treatment with 500 microg of progesterone for 2 h prior to slice preparation, the latency to onset of low Mg(2+)-induced epileptiform activity of slices was significantly prolonged compared to slices from controls. In contrast, progesterone replacement accelerated the development of epileptiform activity in the CA1 region. Neither estrogen alone (2 x 2 microg of estradiol benzoate, 48 and 24 h prior to the experiment), nor a combined treatment with estrogen plus progesterone, significantly affected seizure susceptibility in either CA1 or the EC. There were no consistent effects of estrogen or progesterone, alone or in combination, on flurothyl-induced seizures in vivo. The data suggest that in vitro, progesterone alters seizure susceptibility in a site- and seizure model specific fashion. The differential effects of progesterone may be due to differential expression of progesterone receptor isoforms or metabolites in specific brain areas suggesting that selective modulation of NMDA receptor dependent epileptiform activity may play a role in hormonal effects on epileptogenesis. PMID- 10526104 TI - Lipopolysaccharide evokes the modulation of brain cytochrome P4501A in the rat. AB - The cytochrome P450 enzyme system is a multigene family of enzymes that is modulated in the liver during systemic inflammatory responses or during infection Several forms of the enzyme are expressed in discrete areas of the brain and likely play a critical role in the metabolism of drugs and endogenous chemicals in the central nervous system (CNS). Even though the brain responds to inflammation in a manner different from most tissues, we examined the possible modification of a major cytochrome P450 form (CYP1A) in the brain during inflammation confined to that organ. Total brain CYP1A activity, as measured by ethoxyresorufin dealkylase (EROD), was downregulated 24 and 48 h following the administration of a single dose of lipopolysaccharide (LPS). Regionally, a similar effect was determined in the cortex, hippocampus and the mid-brain but the activity in the cerebellum was unaffected. The examination of coronal brain sections using an antibody directed against CYP1A indicated that the enzyme was distributed in discrete cells of the hippocampus, thalamus and cortex and in the tanycytes surrounding the third ventricle. In each of these areas, the immunoreactivity was diminished in animals receiving LPS as compared to saline treated animals. LPS also evoked the expression of the small molecular weight heat shock protein hsp27 throughout the brain indicating the development of an inflammatory response. These studies indicate that inflammation localized to the CNS causes an alteration in the levels and activity of a major cytochrome P450 form in the brain. This could have implications to the metabolism or activation of drugs and endogenous chemicals in the CNS during a disease state that features an inflammatory component. PMID- 10526105 TI - The nature of penumbral depolarizations following focal cerebral ischemia in the rat. AB - It has been previously suggested that the transient ischemic depolarizations (IDs), thought involved in the gradual expansion of ischemic injury in the first hours following middle cerebral artery occlusion (MCAo), are akin to spreading depression (SD). However, previous studies indicate that the characteristics of these events are heterogeneous (unlike those of SDs). We therefore sought to determine whether different types of IDs exist or not. Using four cortical microelectrodes, we compared the spatial and the temporal characteristics of IDs that occur following intraluminal MCAo in halothane-anesthetized rats to those of electrically induced SDs. An average 4.6+/-3.2 series of events, sequentially affecting the four electrodes, were recorded in 5 h following the induction of ischemia. The distribution of ID duration disclosed two types: short IDs (<7 min, 53% of all events) and long IDs (>7 min; 9% of all events). Most long IDs occurred within the first 30 min and as the initial electrophysiological event. Later on and often restricted to a single or reduced number of recording sites, intermittent IDs were of reduced amplitude or even replaced entirely by suppressed electrocorticographic activity (38% of all events). While the amplitude, duration and spreading characteristics were similar between short IDs and SDs provoked in the cortex of non-ischemic rats, those of long IDs were markedly different. Our results indicate that two types of IDs exist and confirm that most IDs (short ones) are similar in nature to SDs. Long IDs may represent a penumbral anoxic depolarization (AD), reversed by an improvement of perfusion, in the early stages of ischemia. Furthermore, we show that intermittent blockade of depolarization waves occurs and that its incidence increases with time. This blockade may reflect adaptive mechanisms which take place to prevent further depolarizations, the nature of which remains to be determined. The present description of electrophysiological abnormalities might have implications for anti-depolarization therapy in focal cerebral ischemia and to interpret the results of non-invasive techniques which enable the imaging of depolarized areas following stroke. PMID- 10526107 TI - Evidence for subnucleus interpolaris in craniofacial muscle pain mechanisms demonstrated by intramuscular injections with hypertonic saline. AB - The subnucleus interpolaris (Vi) has been identified as a major recipient for trigeminal ganglionic input from jaw muscles, and contains neurons with nociceptive properties similar to those in the subnucleus caudalis (Vc). Therefore, Vi may be another important site for processing craniofacial muscle nociception. The aims of present study were to define functional properties of Vi neurons that receive input from masseter muscle afferents by characterizing their responses to electrical, mechanical, and to chemical stimulation of the muscle. Ninety cells were identified as masseter muscle units in 11 adult cats. Most of these units (79%) received additional inputs from orofacial skin. Following the intramuscular injection of 5% hypertonic saline, 49% of the cells showed a significant modulation of either the resting discharge and/or responses to innocuous mechanical stimulation on their cutaneous receptive fields (RFs). The most common response to saline injection was an induction or facilitation of resting discharge which declined as an exponential decay function, returning to pre-injection level within 3-4 min. Forty-five percent of the muscle units that were tested with mechanical stimulation (13/29) showed a prolonged inhibition of mechanically-evoked responses. In most cases (8/13), the inhibitory response was accompanied by initial facilitation. The observations that Vi contained a population of neurons that receive small diameter muscle afferent inputs, responded to noxious mechanical stimulation on the muscle and to a chemical irritant that is known to produce pain in humans provide compelling evidence for the involvement of Vi in craniofacial muscle pain mechanisms. PMID- 10526106 TI - Platelet-activating factor induced Ca(2+) signaling in human microglia. AB - Increases in intracellular Ca(2+) concentration in human microglial cells in response to platelet-activating factor (PAF) were studied using Ca(2+)-sensitive fluorescence microscopy. In normal physiological solution (PSS), PAF-induced transient increases in [Ca2+](i) which recovered to baseline values within 200 s. Application of PAF in zero-Ca(2+) solution caused the peak response to be decreased to a value near 20% of that recorded in PSS suggesting a primary contribution of Ca(2+) influx for the [Ca2+](i) increase in PSS. To investigate PAF-induced Ca(2+) influx, the contents of intracellular stores were modulated using the SERCA blocker cyclopiazonic acid (CPA). The Ca(2+) signal induced by CPA (10 microM) in zero-Ca(2+) solution showed a peak response about 20% of the amplitude in the presence of external Ca(2+), suggesting the latter response included significant contributions from store-operated Ca(2+) entry. The influx of divalent cations with PAF or CPA was directly measured using Mn(2+) quenching of the fluorescence signal. Although both PAF and CPA induced a similar degree of Mn(2+) influx over time, the PAF effect was very rapid, whereas the CPA action was delayed and only evident about 200 s after application. Overall, the results show that the primary source of the PAF-induced increase of [Ca2+](i) in human microglia was the influx of Ca(2+) from the extracellular space and intracellular Ca(2+)-release contributed only a small part of the total Ca(2+) signal. Nevertheless, Ca(2+)-release induced by PAF (or CPA) serves as an important factor in controlling Ca(2+) entry presumably mediated by activation of store operated-Ca(2+) channels. PMID- 10526108 TI - Postmortem diffusion of autoradiographic blood flow tracers. AB - The heterogeneity of blood flow in the brain under normo- and pathophysiological conditions, as well as during functional activation, has stimulated an interest in the use of autoradiography as a technique for the measurement of local cerebral blood flow. [14C]iodoantipyrine is the most prevalent tracer for the autoradiographic measurement of local cerebral blood flow since it is inert, nonvolatile, and is readily diffusible through the blood-brain barrier. The ability to diffuse freely in cerebral tissue, however, can lead to significant errors if the time duration between when the animal is sacrificed and when the tissue is frozen becomes appreciable, leading to significant postmortem diffusion of the tracer. Using an in vitro technique, the bulk diffusion coefficient for [14C]iodoantipyrine was measured in brain tissue (2.1 x 10(-6) cm(2)/s). Cerebral blood flow was measured with [14C]iodoantipyrine in anesthetized rats. At the end of the radiotracer infusion, the brain was freeze-captured using a device consisting of two rapidly spinning stainless steel blades that were pneumatically driven through the head, freezing the tissue several hundred milliseconds following sacrifice. Autoradiograms from these brains exhibit considerable heterogeneity in blood flow. Computer simulations of the effect of tracer diffusion on these autoradiograms show significant degradation of the images highlighting the importance of very rapid postmortem freezing. PMID- 10526109 TI - Short-term plasticity of the human auditory cortex. AB - Magnetoencephalographic measurements (MEG) were used to examine the effect on the human auditory cortex of removing specific frequencies from the acoustic environment. Subjects listened for 3 h on three consecutive days to music "notched" by removal of a narrow frequency band centered on 1 kHz. Immediately after listening to the notched music, the neural representation for a 1-kHz test stimulus centered on the notch was found to be significantly diminished compared to the neural representation for a 0.5-kHz control stimulus centered one octave below the region of notching. The diminished neural representation for 1 kHz reversed to baseline between the successive listening sessions. These results suggest that rapid changes can occur in the tuning of neurons in the adult human auditory cortex following manipulation of the acoustic environment. A dynamic form of neural plasticity may underlie the phenomenon observed here. PMID- 10526110 TI - Strain and sex differences in the locomotor response and behavioral sensitization to cocaine in hyperactive rats. AB - Individual variability in the behavioral responsiveness to psychostimulant drugs is due, in part, to genetic factors. The present study investigated the effects of acute and repeated administrations of cocaine (0, 5, 10 or 20 mg/kg, i.p.) on locomotor activity in male and female rats from genetically distinct strains often used as a model of human childhood hyperactivity/attentional deficit disorder: Wistar Kyoto Hyperactive (WKHA) rats, Spontaneous Hypertensive rats (SHR) and their control Wistar Kyoto (WKY). The results, expressed as percent change in locomotor activity relative to respective control groups, showed that cocaine elicits a dose-dependent hyperactivity in all strains and revealed neither strain nor sex differences in acute sensitivity to moderate doses of the drug. Nevertheless, across repeated administrations, strain and sex differences appeared: WKHA rats displayed a moderate extent of sensitization to psychomotor stimulant effects of cocaine and female rats showed more robust sensitization than males, whatever the strain. These findings support the genotype-dependence in the development of behavioral sensitization to cocaine and confirm the robustness of the sexual dimorphism across different inbred rat strains. Interestingly, the present results demonstrate that sensitization to psychostimulant drugs occurs in genetically hyperactive strains as well as in their normoactive control strain. PMID- 10526111 TI - Previously reported nerve growth factor levels are underestimated due to an incomplete release from receptors and interaction with standard curve media. AB - The 1996 research report by Hoener et al. [M.C. Hoener, E. Hewitt, J. M. Conner, J.W. Costello, S. Varon, Nerve growth factor (NGF) content in adult rat brain tissue is several-fold higher than generally reported and is largely associated with sedimentable fractions, Brain Res. 728 (1996) 47-56.] compares levels of nerve growth factor (NGF) found in rat brain by assaying both supernatant and pellet to previously reported data. However, Hoener et al. miscalculated when converting values previously reported in the literature to units of picogram per milliliter. Regardless of this mistake, the method of tissue extraction does affect the extent of release of NGF, which must be maximized in order to accurately determine NGF levels in the central nervous system. We now report that accurate measurement of NGF levels is not only affected by the incomplete release of NGF from receptors, but also the medium in which the standard curve is run. It is the combination of these two variables that has led to the underestimation of NGF levels in previous research. PMID- 10526112 TI - Topical application of neurotrophin-3 attenuates ischemic brain injury after transient middle cerebral artery occlusion in rats. AB - In order to examine the effect of neurotrophin-3 (NT-3) on ischemic brain injury, NT-3 was topically applied to brain surface just after 90 min of middle cerebral artery occlusion (MCAO) in rats. NT-3 significantly reduced the infarct size at 24 h of reperfusion. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin in situ nick labeling (TUNEL) staining and immunohistochemical study for caspase 3 and heat shock protein 72 (HSP72) showed that NT-3 treatment decreased the number of cells with DNA fragmentation and caspase-3 and HSP72 expressions. These data suggest that NT-3 protects neuronal cells from ischemic injury, and it is possibly associated with inhibition of DNA fragmentation. PMID- 10526113 TI - Inhibitors of mitogen-activated protein kinases protect axotomized developing neurons. AB - Axotomy kills developing neurons by mechanisms dependent on protein synthesis and influenced by the redox status. Amongst the redox-regulated transduction systems regulating gene expression are the mitogen-activated protein kinases (MAPKs). In the chick embryo, inhibitors of two different MAPK pathways, including notably the p38 kinase pathway, reduce the number of dying axotomized retinal ganglion cells. The regulation of the genetic events associated to axotomy-induced death thus seems to involve MAPKs. PMID- 10526114 TI - Role of the dorsal raphe nucleus in morphine-induced immediate early gene expression in the rat striatum. AB - Serotonin (5-HT) is thought to be involved in morphine action in the brain. To determine if the periaqueductal gray (PAG) and the dorsal raphe nucleus (DRN) are involved in morphine-induced c-Fos and JunB expression in the caudate-putamen (CPu), the mu receptor antagonist, beta-funaltrexamine (beta-FNA), was unilaterally infused into the PAG adjacent to DRN prior to morphine. Behaviorally, beta-FNA prevented morphine-induced loss of righting and Straub tail. In the CPu of beta-FNA treated rats, morphine-induced c-Fos and JunB were attenuated compared to vehicle-infused rats. These results suggest that morphine acts within the PAG-DRN to exert rapid behavioral effects and to induce c-Fos and JunB in the striatum. PMID- 10526115 TI - Dietary restriction protects hippocampal neurons against the death-promoting action of a presenilin-1 mutation. AB - Alzheimer's disease (AD) is an age-related disorder that involves degeneration of synapses and neurons in brain regions involved in learning and memory processes. Some cases of AD are caused by mutations in presenilin-1 (PS1), an integral membrane protein located in the endoplasmic reticulum. Previous studies have shown that PS1 mutations increase neuronal vulnerability to excitotoxicity and apoptosis. Although dietary restriction (DR) can increase lifespan and reduce the incidence of several age-related diseases in rodents, the possibility that DR can modify the pathogenic actions of mutations that cause AD has not been examined. The vulnerability of hippocampal neurons to excitotoxic injury was increased in PS1 mutant knockin mice. PS1 mutant knockin mice and wild-type mice maintained on a DR regimen for 3 months exhibited reduced excitotoxic damage to hippocampal CA1 and CA3 neurons compared to mice fed ad libitum; the DR regimen completely counteracted the endangering effect of the PS1 mutation. The magnitude of increase in levels of the lipid peroxidation product 4-hydroxynonenal following the excitotoxic insult was lower in DR mice compared to mice fed ad libitum, suggesting that suppression of oxidative stress may be one mechanism underlying the neuroprotective effect of DR. These findings indicate that the neurodegeneration-promoting effect of an AD-linked mutation is subject to modification by diet. PMID- 10526116 TI - Effect of upregulation of AMPA glutamate receptors on cerebral O(2) consumption and blood flow in rat. AB - AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate) receptors, in cerebral cortex, underwent upregulation (35% increase) following chronic blockade with a non-competitive AMPA receptor antagonist, GYKI 52466 (1-(aminophenyl)-4 methyl-7, 8-methylenedioxy-5H-2,3-benzodiazepine). Such upregulation did not alter basal cerebrocortical blood flow or O(2) consumption. There was a much higher increase in blood flow and O(2) consumption in the upregulated, agonist (AMPA) stimulated cortices of anesthetized rats. PMID- 10526117 TI - Effects of shear stress on nitric oxide levels of human cerebral endothelial cells cultured in an artificial capillary system. AB - Nitric oxide (NO) is an important vasodilator with various activities in the cerebral vasculature. Although the response of NO levels to shear stress has been investigated in various models using systemic endothelium, no study has evaluated human cerebral endothelial cells (HCE). We determined the NO levels of HCE cultured in an artificial capillary system in response to changes in shear stress. With direct measurement by a porphyrinic microsensor, we found that NO levels increased immediately with a peak at 7 h after changes in shear stress, and by 24 h dropped to a constant elevated baseline. Shear stress-mediated increases in NO levels were confirmed by the measurement of citrulline, an indirect measure of NO. Furthermore, NO levels by HCE were shown to decrease with decreasing shear stress levels. This study presents a novel system to study NO production by microvascular HCE, and indicates a linear relationship between shear stress and NO levels. As cerebral vessels age and lose transmural compliance, shear stress-mediated production of NO may play a greater role in cerebrovascular function and dysfunction. PMID- 10526118 TI - Regulation of norepinephrine transporter and tyrosine hydroxylase mRNAs after kainic acid-induced seizures. AB - Noradrenergic locus coeruleus (LC) efferents to the forebrain suppress seizures in several models of epilepsy. Using in situ hybridization, we demonstrate that tyrosine hydroxylase (TH) and norepinephrine transporter (NET) but not vesicular monoamine transporter 2 (VMAT2) mRNA levels are transiently elevated in LC neurons following kainic acid-induced status epilepticus. These increases of TH and NET mRNAs and presumably of the proteins themselves might enhance synthesis and reuptake of NE postictally. PMID- 10526119 TI - Lubeluzole shows neuroprotective effects in an "in-vitro"-model for neuronal lesions in the chicken retina. AB - In this study, the isolated chicken retina was used as an "in-vitro"-model for investigation of neuronal lesions to show the neuroprotective effects of lubeluzole. Lubeluzole is a neuroprotective compound that has been shown to stereoselectively rescue sensorimotor function and reduce infarct size in photochemical stroke models in rats. In the retina, the typical cell swelling of a developing lesion is accompanied by a very strong intrinsic optical signal (IOS), occurring simultaneous with the electrical signal which is based on changes in light scattering. In the presented model, lesions were elicited electrically with a tungsten microelectrode (0.1 MOmega). The degree of damage was evaluated with optical methods by measuring area and brightness of the affected tissue. Lubeluzole was much more effective in reducing the growth of the lesions than its R-isomer. However, both compounds enhanced the possibility of the neuronal tissue to recover after excitotoxic stimuli. PMID- 10526120 TI - N-acetylcysteine elicited increase in cytochrome c oxidase activity in mice synaptic mitochondria. AB - It has been suggested that thiolic groups are essential for cytochrome c oxidase (COX) activity and other respiratory mitochondrial enzymes. Recent experiments showed that the thiolic antioxidant N-acetylcysteine (NAC) can protect against age-related impairment in COX activity in mice hepatic mitochondria. The present paper shows that NAC enhances COX activity in vitro in synaptic mitochondria isolated from young and old mice. The optimum NAC concentration for maximum COX activity was 5 mM in young and 10 mM in old synaptic preparations. Our data suggest that mitochondrial thiolic groups, which are essentials to oxidative phosphorylation, are impaired by aging. PMID- 10526121 TI - Endothelin-1 contributes to normocapnic hyperoxic pial artery vasoconstriction. AB - The present study was designed to determine if hyperoxia elicits pial artery vasoconstriction and to characterize the contribution of endothelin-1 (ET-1) to that vascular response in newborn pigs equipped with a closed cranial window. Hyperoxic conditions were established by ventilating the piglets with 100% O(2) during normocapnia and concomitantly topically applying artificial CSF that had been bubbled with 100% O(2). Hyperoxia elevated CSF ET-1 from 23+/-1 to 45+/-4 pg/ml. Hyperoxia also elicited pial artery vasoconstriction that was attenuated by BQ123 (10(-6) M), an ET-1 antagonist (-15+/-1 vs. -5+/-1%). These data indicate that ET-1 contributes to hyperoxic pial artery vasoconstriction. PMID- 10526122 TI - Orexins/hypocretins regulate drinking behaviour. AB - Orexin/hypocretins are recently identified neuropeptides which regulate feeding behaviour. We found orexins increased water intake when administrated intracerebroventricularly to rats. The effect of orexin-A was more potent as compared with orexin-B, suggesting the possible involvement of OX(1) receptor. The efficacy of orexin-A was almost comparable with that of angiotensin II, and the effect lasted more than 3 h. Prepro-orexin mRNA level was up-regulated when rats were deprived of water. Orexin-immunoreactive varicose axons were observed in the subfornical organ and area postrema, regions implicated in drinking behaviour. These observations suggest a physiological role for orexin as mediators that regulate drinking behaviour. PMID- 10526123 TI - Distribution of rate-intensity function types in chick cochlear nerve after exposure to intense sound. AB - Intense sound exposure to the chick ear produces cochlear damage and losses in auditory function. At twelve days post exposure there is considerable structural repair, although a defect on the sensory epithelium remains in the form of an incompletely healed 'patch' lesion. Auditory function significantly recovers 12 days after the exposure, but it, too, is incomplete. In this paper we describe the relationship between stimulus intensity and cochlear nerve discharge rate (the rate-intensity function) in two groups of chicks. One is exposed to damaging sound levels but allowed 12 days to recover, while the other is a group of non exposed and age-matched control animals. Three different types of rate-intensity functions were identified; saturating, sloping, and straight. The percentage of saturating and sloping functions was compared across all characteristic frequencies in both groups of animals. A significant change was observed in the distribution of these types for recovered units with characteristic frequencies within the region of the patch lesion. In addition, the rate-intensity functions of these units exhibited a steeper slope and a higher maximum response. The distribution of rate-intensity function types and their slope and maximum responses, for units with characteristic frequencies outside of the patch lesion, was similar to those found in control ears. The changes in the cochlear nerve response in exposed chicks may be due to alterations in cochlear mechanics, hair cell or synaptic membrane properties, hair cell innervation, or the loss of a tonic suppression of afferent activity exerted by the damaged short hair cells. PMID- 10526124 TI - Protein expression of brain endothelial cell E-cadherin after hypoxia/aglycemia: influence of astrocyte contact. AB - The blood-brain barrier (BBB) plays a crucial role in protecting the central nervous system (CNS) from any changes in homeostasis brought about by pathological conditions. Cerebrovascular permeability is an important factor in the development of cerebral edema following stroke [M. Plateel, E. Teissier, R. Cecchelli, Hypoxia, dramatically increases the nonspecific transport of blood borne proteins to the brain. J. Neurochem. 68 (1997) 874-877] and any changes in its function can have detrimental neurological consequences. Recently, research has shown that an in vitro model of the BBB is sensitive to short exposures of hypoxia/aglycemia and that changes in endothelial cell calcium flux may be responsible for structural and functional variations in the BBB during ischemic stress [T.J. Abbruscato, T.P. Davis, Combination of hypoxia/aglycemia compromises in vitro BBB. J. Pharmacol. Exp. Ther. 289 (1999) 668-675]. Present experiments investigated bovine brain microvessel endothelial cell (BBMEC) expression of a Ca(2+)-dependent cell-cell adhesion molecule, E-cadherin, which has been shown to be important for blood-brain barrier function [D. Pal, K.L. Audus, T.J. Siahaan, Modulation of cellular adhesion in bovine brain microvessel endothelial cells by a decapeptide. Brain Research 747 (1997) 103-113]. Since it is believed that astrocyte-endothelial cell interaction is crucial for maintenance of in vivo BBB characteristics, we have attempted to optimize our isolation and culturing techniques to produce a reliable, in vitro model of the BBB that is suitable to study pathological conditions. Immunofluoresence experiments showed positive staining for E-cadherin, yet failed to show any change in cellular distribution of E-cadherin upon hypoxic/aglycemic exposure. In addition, culturing BBMECs with C6 conditioned medium (CM) had no effect on the localization of E-cadherin. Western blotting experiments showed that BBMECs express E-cadherin and this protein is decreased in a time dependent manner after various hypoxic/aglycemic exposures when endothelial cells are cultured alone or with C6 astrogliomas grown on a separate culture surface. When C6 astrocytes are grown directly opposed to endothelial cells, with a porous membrane between, we observed a slight attenuation in the decreased BBMEC expression of E-Cadherin after hypoxia/aglycemia exposure. This work has shown that the mammalian brain endothelial/astrocyte co-culture system is a useful model for studies of pathological conditions where BBB characteristics are maintained. PMID- 10526125 TI - Effect of age on calcium-dependent proteins in hippocampus of senescence accelerated mice. AB - The senescence-accelerated P8 mouse (SAMP8) is a well-characterized model for the age-related decline in acquisition and retention. Calcium-dependent protein kinase C (PKC) and calcium-calmodulin-dependent protein kinase (CAM K) have been implicated in these processes in the hippocampus. Therefore, the expression of hippocampal PKC and CAM K was determined in SAMP8 mice aged 4, 8, and 12 months. As measured by Western blotting, total hippocampal PKC-gamma protein declined linearly with age. In addition, the distribution of the PKC-gamma also changed with age. The amount of PKC in the particulate fraction declined linearly with age relative to the soluble PKC. The decline in total PKC and particulate PKC correlated with the previously reported decline in retention but not with the decline in acquisition. Western blotting revealed no consistent change in CAM KII protein levels. In addition to protein levels, Ca-dependent protein kinase activity may also be affected by changes in intracellular Ca concentration. Therefore, the levels of calbindin and the plasma membrane Ca pump, two proteins involved in maintaining low levels of intracellular Ca, were measured in the hippocampus. Calbindin protein declined progressively with age, but there was no significant change in total plasma membrane Ca pump expression. These studies demonstrate a decrease in the amount and distribution of hippocampal PKC-gamma in the SAMP8 between 4 and 12 months that is associated with decreased retention. PMID- 10526126 TI - Reciprocal circuits involved in nitroglycerin-induced neuronal activation of autonomic regions and pain pathways: a double immunolabeling and tract-tracing study. AB - This study uses tract-tracing protocols to determine the circuitry of specific nuclei involved in nitroglycerin-induced activation. Combined retrograde and anterograde tracers were injected into nuclei which consistently demonstrate robust Fos expression following our systemic nitroglycerin injection paradigm. The nuclei, which conform to these criteria, that we have evaluated in this study are the locus coeruleus, parabrachial nucleus and paraventricular nucleus of the hypothalamus. Dual Fos/tracer immunocytochemistry in treated animals documented the existence of a subset of autonomic nuclei which are activated by nitroglycerin injection and have reciprocal connections. From the nature of this rich interconnection we suggest that nitroglycerin activates autonomic responses involved in cardiovascular pressor mechanisms. Nuclei which show strong Fos labeling following nitroglycerin administration, but not traced in this study, include the nucleus trigeminalis caudalis and the ventrolateral column of the periaqueductal gray, both of which mediate nociceptive modalities. These data confirm and expand on our previous findings and demonstrate that nitroglycerin activates a complex set of structures that are functionally and structurally interconnected to articulate an integrated response. PMID- 10526127 TI - Activation of caspase-3 in beta-amyloid-induced apoptosis of cultured rat cortical neurons. AB - Amyloid beta protein (Abeta) has been thought to participate in the neurodegeneration associated with Alzheimer's disease. We here report on caspase 3 activation by Abeta-treatment of cultured neurons. Treatment of rat primary cortical culture with Abeta 25-35, an active fragment of Abeta, induced neuronal death as determined by a decrease in neuron-specific microtubule-associated protein 2 (MAP2)-like immunoreactivity and by the release of cellular lactate dehydrogenase (LDH). Abeta 25-35 also induced elevation of caspase-3-like Ac-DEVD MCA cleavage activity in advance of neuronal death with similar concentration dependency for neuronal death. Inhibitor sensitivity of the Abeta-induced proteolytic activity was similar to that of human recombinant caspase-3. Cleavage of pro-caspase-3 and cleavage of its endogenous substrates, poly (ADP-ribose) polymerase (PARP) and alpha-fodrin, were produced by Abeta-treatment. A caspase-3 inhibitor, Ac-DEVD-CHO, prevented Abeta-induced DNA fragmentation and cleavage of alpha-fodrin, but not of PARP. Caspase inhibitor of broad specificity, Z-VAD CH(2)-DCB, additionally prevented Abeta-induced cleavage of PARP and some early loss of cell membrane integrity measured by LDH release. However, Abeta-induced condensation of nuclear chromatin and most of the late disintegration of cell membranes were not prevented in the presence of these caspase inhibitors. These results suggest that activation of both caspase-3 and caspase(s) other than caspase-3 play distinct roles in Abeta-induced apoptosis of rat cortical neurons. Furthermore, in the presence of caspase inhibitors, Abeta-induced neuronal death still occurred with different morphological features. PMID- 10526128 TI - Importance of stimulation paradigm in determining facilitation and effects of neuromodulation. AB - Evoked synaptic activity within the CNS and at the neuromuscular junction in most in vivo preparations studied occurs not with single isolated stimuli, but with trains, or bursts, of stimuli. Although for ease in studying the mechanisms of vesicular synaptic transmission one often uses single discrete stimuli, the true mechanisms in the animal may be far more complex. When repetitive stimuli are present at a nerve terminal, often a heightened (i.e., facilitated) postsynaptic potential can be as a result. Facilitation is commonly used as an index of synaptic function and plasticity induced by chronic stimulation or by neuromodulation. The mechanisms that give rise to facilitation are thought to be the same that may underlie short-term learning and memory [C.H. Bailey, E.R. Kandel, Structural changes accompanying memory storage. Annu. Rev. Physiol. 55 (1993) 397-426.]. Differences in short term facilitation (STF) are seen depending on the conventional stimulation paradigm (twin pulse, train, or continuous) used to induce facilitation. Thus, a battery of paradigms should be used to characterize synaptic function to obtain a closer understanding of the possible in vivo conditions. PMID- 10526129 TI - Antagonism of the aconitine-induced inexcitability by the structurally related Aconitum alkaloids, lappaconitine and ajacine. AB - Aconitine, lappaconitine and ajacine are structurally related alkaloids occurring in several species of the Aconitum genus. While aconitine is known to activate the voltage-dependent sodium channel, lappaconitine has been reported to block this channel. To investigate a possible antagonism of the aconitine action on neuronal activity by lappaconitine and the closely related alkaloid ajacine, we have performed extracellular recordings of stimulus evoked population spikes and field excitatory postsynaptic potential (EPSP) in rat hippocampal slices. Aconitine (10-100 nM) diminished the amplitude of the orthodromic population spike in a concentration-dependent manner. When aconitine was applied in presence of 10 microM lappaconitine, the concentration-response curve was shifted to the right. Furthermore, the complete suppression of the population spike evoked by 100 nM aconitine was reversed by 10 microM lappaconitine. The action of lappaconitine was mimicked by ajacine, however, the latter alkaloid was less potent. Both lappaconitine and ajacine shifted the input-output relationship of the presynaptic fiber spike as function of the stimulation intensity and of the field EPSP as function of the presynaptic fiber spike to the right. After pharmacological isolation, the presynaptic fiber spike was decreased by both compounds in a frequency-dependent manner indicative for a use-dependent action. Thus, electrophysiologically these alkaloids seem to inhibit predominantly the excitability of the afferent fibres and, in consequence, neurotransmission between Schaffer collaterals and the CA1 neurons, thereby suppressing the firing of the latter. Ajacine and lappaconitine inhibited stimulus-triggered epileptiform population bursts in area CA1 elicited by omission of Mg(2+) as well as spontaneously occurring epileptiform discharges in area CA3 elicited by omission of Mg(2+) and elevation of K(+). It is concluded that the inhibitory and antiepileptiform effect of ajacine and lappaconitine is mediated by a frequency dependent inhibition of the voltage-dependent sodium channel, thereby decreasing the excitability which might be important for filtering high frequency bursts of action potentials characteristic for epileptiform activity in the hippocampus. Moreover, these alkaloids are naturally occurring antagonists of the sodium channel activator aconitine. PMID- 10526130 TI - pRb phosphorylation is regulated differentially by cyclin-dependent kinase (Cdk) 2 and Cdk4 in retinoic acid-induced neuronal differentiation of P19 cells. AB - The retinoblastoma protein (pRb) is a key regulator of cell growth, differentiation and survival. pRb(-/-) mice show abnormal neuronal cell death in the developing brain. The function of pRb is regulated by its phosphorylation state. In this study, the phosphorylation of pRb during retinoic acid (RA) induced neuronal differentiation of P19 cells was examined using site-specific antibodies against pRb phosphorylated at Ser601, Ser605 and Ser773. Although pRb was hyperphosphorylated in undifferentiated P19 cells, Ser601 and Ser773 were not phosphorylated. Upon exposure to RA, however, these two sites became strongly phosphorylated. Cdk4 kinase activity was almost undetectable in undifferentiated P19 cells, but was strongly activated on exposure to RA. In contrast, Cdk2 kinase activity and the phosphorylation of Ser605 were observed in undifferentiated cells as well as in RA-treated cells. These observations suggest that Cdk2 and Cdk4 may phosphorylate different sites of pRb in vivo and that the two sites of pRb examined here are newly phosphorylated during RA-induced neuronal differentiation in P19 cells. PMID- 10526131 TI - ATM immunolocalization in mouse neuronal endosomes: implications for ataxia telangiectasia. AB - Ataxia-telangiectasia (A-T) is a human disorder with pleiotropic manifestations that include neoplasms, immune dysfunction and neurodegeneration. The disorder is due to mutations in the gene known as ATM (A-T, mutated), which causes a deficiency in its protein product (Atm in mice) that is necessary for DNA damage surveillance. This nuclear function of Atm explains in principle the propensity to cancer and immunodeficiency in A-T, but not the neurodegeneration which results in the earliest clinical manifestations and causes progressive disability. Here we report ultrastructural evidence of cytoplasmic localization of Atm-like immunoreactivity (ALI) within endosomes in murine cerebellocortical neurons, one of the principal targets of A-T. The ALI was obtained with two separate monoclonal antibodies that recognize Atm specifically. By contrast, electron-dense endosomes that could be confused with ALI occur in negligible amounts in both wild-type mice and in mice deficient in Atm ("knockout" mice). Furthermore, there was a marked preferential distribution of Atm-immunopositive endosomes in the granule cell layer - where they are present in granule neurons - with a much lower density in the Purkinje and molecular layers. These observations suggest that endosome-bound Atm may be more important for the function of certain neurons than others - or that it is processed differently among them - and that this protein may be involved in molecular sorting in the cytoplasm. This is relevant to elucidating the role of Atm deficiency in the pathobiology of neurodegeneration in A-T. PMID- 10526132 TI - Light and electron microscopic immunocytochemical study on the innervation of the pineal gland of the tree shrew (Tupaia glis), with special reference to peptidergic synaptic junctions with pinealocytes. AB - Conventional and immunocytochemical, light- and electron-microscopic studies on the innervation of the pineal gland of the tree shrew (Tupaia glis) were made. Neuropeptide Y (NPY)-immunoreactive fibers, which were abundantly distributed in the gland, disappeared almost completely after superior cervical ganglionectomy, suggesting that these fibers are mostly postganglionic sympathetic fibers. By contrast, tyrosine hydroxylase (TH)-immunoreactive fibers, which were less numerous than NPY-fibers, remained in considerable numbers in ganglionectomized animals, indicating the innervation of TH-positive fibers from extrasympathetic sources. Bundles of substance P (SP)- or calcitonin gene-related peptide (CGRP) immunoreactive fibers, entering the gland at its distal end, were left intact after ganglionectomy. SP-fibers were numerous, but CGRP-fibers were scarce in the gland. SP-immunoreactive fibers were myelinated and nonmyelinated, and were regarded as peripheral fibers because of the presence of a Schwann cell sheath. NPY- and SP-immunoreactive fibers and endings were mainly localized in the pineal parenchyma. NPY-immunoreactive endings synapsed frequently, and SP-positive ones did less frequently, with the cell bodies of pinealocytes. The results suggest that NPY and SP directly control the activity of pinealocytes. Sections stained for myelin showed that thick and less thick bundles of myelinated fibers entered the gland by way of the habenular and posterior commissures, respectively. Under the electron microscope, the bundles were found to contain also unmyelinated fibers. A considerable number of nerve endings synapsing with the cell bodies of pinealocytes remained in ganglionectomized animals; these endings were not immunoreactive for TH or SP. Such synaptic endings may be the terminals of commissural fibers. PMID- 10526133 TI - Loss of cyclin D1 in necrotic and apoptotic models of cortical neuronal degeneration. AB - Recent evidence suggests that apoptosis in post-mitotic neurons involves an aborted attempt of cells to re-enter the cell cycle and it is characterized by increased expression of cyclins, such as cyclin D1, prior to death. Cyclin D1 increases to permit transition from growth phase (G0/G1) to synthesis phase (S) during normal development but there is controversy as to which of the cyclins are activated prior to apoptotic cell death. We looked at the expression of cyclin D1 in cortical neuronal cultures treated with either staurosporine to produce apoptotic death, or with glutamate, to produce a non-apoptotic death. Cyclin D1 immunoreactivity was observed in the cytoplasm and nucleus of virtually all neurons under control conditions. Following the addition of either staurosporine or glutamate, cyclin D1 immunoreactivity did not change within 4 h. The cyclin D1 immunoreactivity was lost by 6 h with the appearance of either staurosporine induced fragmented nuclei or glutamate-induced pyknotic nuclei. These immunocytochemical observations were confirmed with immunoblot analysis. Therefore, cyclin D1 is not a reliable indicator of apoptosis in cortical neuronal cultures and should not be used as an indicator of apoptotic cell death. PMID- 10526134 TI - Enhanced blood pressure buffering role of the brain nitrergic system in renin transgenic rats. AB - Previous studies provided evidence for an interaction between the brain nitrergic and vasopressinergic systems in normotensive and spontaneously hypertensive rats in regulation of the cardiovascular functions. The present study was designed to determine the role of the brain nitric oxide (NO) in regulation of basal blood pressure and its interaction with vasopressin (AVP) in rats with renin dependent transgenic hypertension TGRmRen2(27) (TGR). The experiments were performed on conscious hypertensive TGR and normotensive Sprague-Dawley (SD) rats. Both groups were chronically instrumented with the left cerebral ventricle cannula (LCV) and femoral arterial catheter. LCV application of 2.3 nmol (0.5 microg) of N(G)-nitro L-arginine (L-NNA) an inhibitor of NO synthesis significantly elevated blood pressure (MAP) in TGR but not in SD rats. In contrast administration of NO donor S-acetyl-N-penicillamine (SNAP) produced significant decrease of MAP only in SD rats. LCV application of AVP (10 ng) elicited comparable increases of MAP in TGR and SD rats. Pretreatment with L-NNA significantly potentiated pressor response to AVP in TGR rats but not in SD rats. The results provide evidence that increased production of intrabrain NO may play a significant blood pressure buffering role in TGR rats both under baseline conditions and during activation of the vasopressinergic system. PMID- 10526135 TI - Corticotropin releasing factor and substance P mediate the nucleus amygdaloideus centralis-nucleus ventromedialis-nucleus dorsomedialis pressor system. AB - Prolonged emotional stress is an important factor in the development of neurogenic hypertension, but its mechanism is still unclear. The purpose of the present study is to analyze the possible neural basis of hypertension induced by prolonged emotional stress. In the brain many nuclei are involved in emotional reaction, stress or defense response; among them the nucleus amygdaloideus centralis (AC) is the most important one which widely connects with other nuclei controlling emotion and stress, such as nucleus ventromedialis (NVM), nucleus dorsomedialis (NDM), nucleus paraventricularis (NPV) etc. These nuclei contain corticotropin releasing factor (CRF)- and substance P (SP)-immunoreactive cell bodies, nerve terminals and corresponding receptors. Our previous and present studies showed that microinjection of CRF or SP into these nuclei induced pressor responses. These data imply that excitation of the AC can activate many nuclei controlling emotion and stress via CRF and SP, and excessive activities of these nuclei may be the neural basis of hypertension induced by prolonged emotional stress. The present study revealed that (1) the AC pressor response to glutamate (Glu) could be reduced by preinjection of CRF antagonist (alpha-Helical CRF[9-41] or SP antagonist ([D-Pro(2), D-Phe(7), D-Trp(9)]-substance P) into bilateral NVM, (2) the NVM pressor response to Glu were decreased by pretreatment of the NDM with CRF- or SP-antagonist, (3) the AC-, NVM- or NDM-pressor responses were all attenuated by preinjection of CRF- or SP-antagonist into bilateral NPV or rostral ventrolateral medulla (RVL). The results indicate that excitation of the AC can indirectly activate the NPV and RVL to evoke pressor response via the NVM-NDM, CRF and SP are transmitters in each connection of this pathway; this is one component of the mechanism underlying the AC pressor response. Taken together with the findings of our previous studies, it provides neurophysiological basis for the above-mentioned implications. PMID- 10526136 TI - Use of in vitro superfusion to assess the dynamics of striatal dopamine clearance: influence of estrogen. AB - To determine the feasibility of assessing dopamine uptake using in vitro superfusion, striatal tissue from ovariectomized female rats was infused with dopamine (1 microM), nomifensine (1 mM), or a combination of dopamine and nomifensine. Treatment with nomifensine or dopamine/nomifensine increased the recovery of dopamine in the effluent samples as compared to treatment with dopamine alone. In Experiment 2, the striatal tissue was treated with varying concentrations (0, 3, 30 or 300 nM) estradiol throughout the superfusion and subsequently given a dopamine (1 microM) challenge. The recovery of dopamine was enhanced in the presence of 3 and 30 nM estradiol. These results show that (1) in vitro superfusion can be used to dynamically evaluate dopamine recovery, and (2) estradiol, like nomifensine, increases the recovery of exogenously applied dopamine from the striata of ovariectomized female rats. Such increases in dopamine recovery with estrogen and similarities to that obtained with nomifensine suggest that estrogen may be inhibiting dopamine uptake from these striatal tissue fragments. Moreover, the doses at which estrogen can exert these effects insinuates a physiological role for this process. Our data provide a clear functional demonstration for one of the mechanisms by which estradiol can modulate striatal dopamine neurons, that of an uptake inhibitor. Such a mechanism has important implications with regard to estradiol's capacity to function as a neuroprotectant of the nigrostriatal dopaminergic system through inhibition of uptake of neurotoxins which can produce neurodegeneration of striatal dopamine neurons. PMID- 10526137 TI - Histamine-immunoreactive neurons in the brain of the cockroach Leucophaea maderae. AB - Histamine is the neurotransmitter of insect photoreceptor cells but has also been found in a small number of interneurons in the insect brain. In order to investigate whether the accessory medulla (AMe), the putative circadian pacemaker of the cockroach Leucophaea maderae receives direct visual input from histaminergic photoreceptors, we analyzed the distribution of histamine-like immunoreactivity in the optic lobe and midbrain of the cockroach. Intense immunostaining was detected in photoreceptor cells of the compound eye, which terminated in the first optic neuropil, the lamina, and in a distal layer of the medulla, the second optic neuropil. Histamine immunostaining in parts of the AMe, however, originated from a centrifugal neuron of the midbrain. Within the midbrain 21-23 bilaterally symmetric pairs of cell bodies were stained. Most areas of the brain were innervated by one or more of these neurons, but the protocerebral bridge and the mushroom bodies were devoid of histamine immunoreactivity. The branching patterns of most histamine-immunoreactive neurons could be reconstructed individually. While the majority of identified neurons arborized in both brain hemispheres, five cells were local neurons of the antennal lobe. A comparison with other insect species shows striking similarities in the position of certain histamine-immunoreactive neurons, but considerable variations in the presence and branching patterns of others. The data suggest a role for histamine in a non-photic input to the circadian system of the cockroach. PMID- 10526138 TI - Facilitatory effect of unilateral lesion of the ventral tegmental area on locomotor response to stimulation of the contralateral ventral tegmental area: involvement of GABAergic transmission. AB - It was found previously that in the rat, unilateral electrolytic lesion of the ventral tegmental area (VTA) facilitated feeding induced by electrical stimulation of the homologous VTA tissue in the contralateral hemisphere. In the present work, VTA stimulation-induced locomotor response was tested in male Wistar rats using a latency to move/stimulation frequency curve shift paradigm in order to check for functional generality of the "contralateral facilitation effect" and also with the aim of elaborating an easy and reliable behavioral model to study this phenomenon. In a further step, the hypothesis was tested that enhancement of function of the intact VTA results from elimination of tonic GABAergic influence derived normally from the lesioned VTA. GABA(A) (bicuculline, doses 0, 0.5 and 5.0 ng) and GABA(B) (phaclofen, doses 0, 500 and 1000 ng) receptors antagonists, and for comparison, a GABA(A) receptor agonist (muscimol, doses 0, 12.5, 25. 0 and 50.0 ng), were injected unilaterally to VTA and their effect on locomotor response elicited by electrical stimulation of the contralateral VTA was tested in a latency/frequency paradigm. It was found that similar to feeding, locomotor response evoked by unilateral electrical stimulation of the VTA was facilitated after contralateral VTA lesion which manifested as a decrease of the locomotion threshold and a leftward shift of the function relating latency to move to stimulation frequency. The effect was immediate, long-lasting and specific to the VTA destruction; lesions outside the VTA area caused gradual impairment of the locomotor response to stimulation. The facilitatory effect of the electrolytic lesion could be replicated by bicuculline, which significantly facilitated stimulation-induced behavior. Phaclofen exerted slight facilitating influence only at a low dose. No effect of muscimol on the locomotion threshold was found. We conclude that "the contralateral facilitation effect" at the level of VTA reflects the interhemispheric regulation of activity of the dopaminergic (DA) cells in which GABA(A)-mediated interhemispheric communication plays a significant role. PMID- 10526139 TI - Neurotrophin-elicited short-term glutamate release from cultured cerebellar granule neurons. AB - Brain-derived neurotrophic factor (BDNF) has been suggested to play an important role in neuronal plasticity. In this study, we investigated the effects of BDNF on short-term transmitter release from cultured CNS neurons. Rapid and transient glutamate and aspartate releases induced by BDNF were observed from cultured cortical, hippocampal, striatal and cerebellar neurons. We furthermore investigated the mechanism of release induced by neurotrophins from cerebellar granule cells, since granule cells represent a large homogeneous glutamatergic population. NGF and NT-3 elicited neurotrophin-induced release of glutamate as well as BDNF from the cerebellar granule neurons. The release was dependent on intracellular Ca(2+) mobilization. Pretreatment with K252a and also TrkB-IgG completely blocked the glutamate and aspartate release elicited by BDNF, but not by NGF. The cerebellar granule neurons expressed trkB and p75 mRNAs at high levels, but not trkA mRNA. These results suggested that while BDNF induced release via TrkB, NGF-elicited release was not mediated by Trks. Furthermore, in the experiment using the styryl dye FM1-43, which selectively labels synaptic vesicles, neither BDNF nor NGF evoked dye loss, suggesting that neurotrophin induced excitatory amino acid release occurs through a non-exocytotic pathway. PMID- 10526140 TI - Copper levels are increased in the cerebral cortex and liver of APP and APLP2 knockout mice. AB - The pathological process in Alzheimer's disease (AD) involves amyloid beta (Abeta) deposition and neuronal cell degeneration. The neurotoxic Abeta peptide is derived from the amyloid precursor protein (APP), a member of a larger gene family including the amyloid precursor-like proteins, APLP1 and APLP2. The APP and APLP2 molecules contain metal binding sites for copper and zinc. The zinc binding domain (ZnBD) is believed to have a structural rather than a catalytic role. The activity of the copper binding domain (CuBD) is unknown, however, APP reduces copper (II) to copper (I) and this activity could promote copper-mediated neurotoxicity. The expression of APP and APLP2 in the brain suggests they could have an important direct or indirect role in neuronal metal homeostasis. To examine this, we measured copper, zinc and iron levels in the cerebral cortex, cerebellum and selected non-neuronal tissues from APP (APP(-/-)) and APLP2 (APLP2(-/-)) knockout mice using atomic absorption spectrophotometry. Compared with matched wild-type (WT) mice, copper levels were significantly elevated in both APP(-/-) and APLP2(-/-) cerebral cortex (40% and 16%, respectively) and liver (80% and 36%, respectively). Copper levels were not significantly different between knockout and WT cerebellum, spleen or serum samples. There were no significant differences observed between APP(-/-), APLP2(-/-) and WT mice zinc or iron levels in any tissue examined. These findings indicate APP and APLP2 expression specifically modulates copper homeostasis in the liver and cerebral cortex, the latter being a region of the brain particularly involved in AD. Perturbations to APP metabolism and in particular, its secretion or release from neurons may alter copper homeostasis resulting in increased Abeta accumulation and free radical generation. These data support a novel mechanism in the APP/Abeta pathway which leads to AD. PMID- 10526141 TI - The effect of ipsapirone and S(-)-pindolol on dopamine release in rat striatum and nucleus accumbens. AB - Serotonin (5-hydroxytryptamine, 5-HT)(1A) receptor agonism and 5-HT(2A) receptor antagonism are components in the action of some of the recently developed antipsychotic drugs, e.g., clozapine and ziprasidone. However, studies of the role of 5-HT(1A) receptor agonism in the ability of these drugs to modulate dopamine (DA) release in the nucleus accumbens (NAC), which may be relevant to antipsychotic action, are lacking. Thus, we examined the effect of clinically available agents, ipsapirone, a 5-HT(1A) receptor partial agonist, and the mixed 5-HT(1A/1B)/beta receptor antagonist S(-)-pindolol, on DA release in the NAC compared to the striatum (STR). Ipsapirone produced a biphasic effect; low dose (0.1 mg/kg) decreased, high dose (3 mg/kg) increased and intermediate doses (0.1 and 1 mg/kg) did not change DA release in the NAC, respectively. However, ipsapirone, at all doses (0.3, 1, 3, but not 0.1 mg/kg) increased striatal DA release. S(-)-pindolol (3, 10, but not 1 mg/kg) produced a comparable increase in DA release in the NAC and STR. These results suggest that the ability of lower dose of ipsapirone to decrease DA release in the NAC is more likely to be due to 5-HT(1A) receptor agonism. On the other hand, the effect of higher dose of ipsapirone on striatal DA release may be due to 5-HT(1A) receptor antagonism, as is the case with S(-)-pindolol. The mechanism and clinical significance of these results for developing antipsychotic drugs is discussed. PMID- 10526142 TI - Alkylamides that produce tingling paresthesia activate tactile and thermal trigeminal neurons. AB - Alkylamides isolated from the fruit of Xanthoxylum, Szechuan pepper, produce a strong tingling sensation in the mouth. In order to determine the peripheral basis of this sensation, extracellular nerve recordings were obtained from the lingual nerve of rats. The primary pungent compound, hydroxy-alpha-sanshool (HO alpha-S), altered the levels of spontaneous activity in cool-sensitive fibers as well as inducing activity in tactile fibers, cold nociceptors and silent fibers that were insensitive to innocuous thermal or tactile stimuli. Moreover, tactile or thermal sensitivity was induced in fibers that were initially insensitive to touch or cooling. The neuronal distribution of sensitivities to capsaicin and to HO-alpha-S indicate that this compound affects neurons mediating innocuous sensations. HO-alpha-S may be useful as a model stimulus for studies of paresthesia. PMID- 10526143 TI - Subtypes of metabotropic glutamate receptors in the nucleus of the solitary tract of rats. AB - We have determined the role of subgroups of metabotropic glutamate receptors (mGluRs) in the nucleus of the solitary tract (NTS) of normotensive Wistar rats. Unilateral microinjection of (S)-3, 5-dihydroxyphenylglycine (3,5-DHPG), an agonist of group I mGluRs, into the NTS significantly decreased mean arterial pressure (MAP), heart rate (HR), and renal sympathetic nerve activity (RSNA) ( 19. 4+/-2.6 mmHg, -16.4+/-5.1 beats/min, and -30.6+/-5.7% by 1 nmol). Microinjection of (R,S)-1-aminoindan-1,5-dicarboxylic acid (AIDA; 1 nmol), a putative antagonist of group I mGluRs, into the NTS caused transient decreases in MAP and RSNA, followed by sustained increases in MAP (+8.3+/-2.4 mmHg) and RSNA (+27.7+/-10.8%). Pretreatment with AIDA failed to prevent the cardiovascular and RSNA responses to microinjection of 3,5-DHPG. Unilateral microinjection of (S)-4 carboxy-3-hydroxyphenylglycine (4C3HPG), an agonist of group II mGluRs, into the NTS also significantly decreased MAP, HR, and RSNA, whose responses were not inhibited by pre-microinjection of (2S)-alpha-ethylglutamic acid (EGLU; 2 nmol), a putative antagonist of group II mGluRs. On the other hand, unilateral microinjection of L(+)-2-amino-4-phosphonobutyric acid (L-AP4), an agonist of group III mGluRs, into the NTS caused dose-related decreases in MAP (-8. 3+/-1.5 mmHg by 0.1 nmol and -45.1+/-3.4 mmHg by 0.3 nmol), HR, and RSNA (-21.3+/-3.9% by 0.1 nmol and -77.2+/-6.5% by 0.3 nmol), whose responses were suppressed by pre microinjection of (R, S)-alpha-cyclopropyl-4-phosphonophenylglycine (CPPG; 0.3 nmol), an antagonist of group III mGluRs. These results suggest that all subgroups of mGluRs participate in cardiovascular and sympathetic regulations in the NTS of rats, and that endogenous group I mGluRs in the NTS may contribute to tonic cardiovascular and sympathetic regulations. PMID- 10526144 TI - NMDA receptor-mediated control of GABA release from neurointermediate lobes of female and male rats. AB - The effect of glutamate (GLUT) and its ionotropic receptor agonists on K(+) evoked GABA release from the neurointermediate lobe (NIL) was investigated in diestrus, ovariectomized, ovariectomized-estrogenized female rats and intact male rats. GLUT and N-methyl-D-aspartate (NMDA) increased K(+)-evoked GABA release from the NIL in all the experimental groups. This stimulatory effect of NMDA was blocked by specific NMDA receptor antagonists but not by non-NMDA receptor antagonists. However, kainate did not modify evoked GABA release from the NIL in any of these groups. Neither GLUT nor NMDA modified nitric oxide synthase activity. These results indicate that GLUT, acting through NMDA receptors, stimulates evoked GABA release from the NIL of female and male rats. This effect is not influenced by gonadal status and does not appear to be mediated by nitric oxide production. PMID- 10526145 TI - Food intake elicited by central administration of orexins/hypocretins: identification of hypothalamic sites of action. AB - Orexin A and B, a recently identified pair of neuropeptides, are produced in perikarya located in the lateral and perifornical hypothalamus (LH and PFH). Immunoreactive fibers from these neurons innervate several nuclei in the hypothalamus. Orexin A and orexin B stimulate feeding when administered intracerebroventricularly to rats. To identify the specific sites of orexin action, orexin A and B were microinjected into a number of hypothalamic and extrahypothalamic sites in rats. Orexin A was found to enhance food intake when injected into four hypothalamic sites, the paraventricular nucleus (PVN), the dorsomedial nucleus (DMN), LH and the perifornical area, but was ineffective in the arcuate nucleus (ARC), the ventromedial nucleus (VMN), and the preoptic area (POA) as well as the central nucleus of the amygdala (CeA) and nucleus of the tractus solitarius (NTS). Orexin B was not effective at any site tested. These findings demonstrate that orexin A receptive sites for stimulation of food intake exist primarily in a narrow band of neural tissue within the hypothalamus that is known to be involved in control of energy homeostasis. PMID- 10526146 TI - Terfenadine prevents NMDA receptor-dependent and -independent toxicity following sodium channel activation. AB - Exposure of cultured cerebellar neurons to terfenadine prevented the N-methyl-D aspartate (NMDA) receptor-mediated early appearance (30 min) of toxicity signs induced by the voltage sensitive sodium channel (VSSC) activator veratridine. Delayed neurotoxicity by veratridine (24 h) occurring independently from NMDA receptor activation was also prevented by terfenadine. Terfenadine did not protect from excitotoxicity following direct exposure of neurons to glutamate. Our results suggest that terfenadine may modulate endogenous glutamate release following activation of VSSCs. PMID- 10526147 TI - Localization of opioid-binding cell adhesion molecule (OBCAM) in adult rat brain. AB - We investigated the tissue distribution and brain localization of opioid-binding cell adhesion molecule (OBCAM) in the adult rats by immunoblotting and immunohistochemistry using a monoclonal anti-OBCAM peptide antibody that is specific for OBCAM. OBCAM was preferentially expressed in the central nervous system (CNS) and at a very low level in the spleen. Within the brain, OBCAM was distributed in almost all the gray matter, but little or no immunoreactive OBCAM was found in the white matter. Morphologically, the distribution pattern of OBCAM immunoreactivity was very similar to that of synaptophysin, suggesting a role in the synaptic machinery. PMID- 10526148 TI - Striatal perfusion of indomethacin attenuates dopamine increase in immature rat brain exposed to anoxia: an in vivo microdialysis study. AB - Using in vivo microdialysis and HPLC, we examined the effects of indomethacin on extracellular dopamine (DA) in the striatum of immature rats submitted to anoxia. Rat pups in two indomethacin groups received intrastriatal perfusion of either 1 mM or 5 mM indomethacin throughout the experiment. The DA level reached 1185+/ 400% of the basal level during anoxia; in contrast, the peak levels of DA were only 307+/-63%, 153+/-35% in indomethacin groups (p<0.05). We consider that this suppression would be one of the mechanisms of the protective effect of indomethacin on hypoxic ischemic encephalopathy. PMID- 10526149 TI - Histochemical study of dopamine-degrading monoamine oxidase activity in dopaminergic neurons of rat brain. AB - We examined whether dopamine-degrading activity of monoamine oxidase (MAO) is present in dopaminergic neurons of the rat brain. We employed a double-labeling procedure combining immunohistochemistry for tyrosine hydroxylase (TH) and enzyme histochemistry for MAO activity using dopamine as a substrate. The following dopaminergic cell groups were examined: A16 (glomerular layer of the olfactory bulb), A14 (hypothalamic periventricular region), A13 (zona incerta), A12 (arcuate nucleus), A11 (periventricular gray matter of the caudal thalamus), A10 (ventral tegmental area), A9 (substantia nigra pars compacta, SNC) and A8 (retrorubral nucleus). Although no MAO activity was detected in any of the TH immunoreactive dopaminergic neurons, strong dopamine-degrading MAO activity was found in TH-positive neurons in the locus coeruleus (LC) (i.e., noradrenergic neurons). Our results indicate that dopamine-degrading MAO activity is very low in dopaminergic neurons compared to the MAO activity in LC noradrenergic neurons. PMID- 10526150 TI - Role of potassium channels in the central neurogenic neuroprotection elicited by cerebellar stimulation in rat. AB - Electrical stimulation of the cerebellar fastigial nucleus (FN) in spontaneously hypertensive (SHR), Wistar-Kyoto (WKY) and Fisher rats reduced, by approximately 50%, the infarctions produced by occlusion of the middle cerebral artery. Blockade of ATP-dependent potassium (K-ATP) channels with glibenclamide (i.c.v.) abolished salvage only in the SHR rat. While blockade of K-ATP channels failed to abolish salvage in WKY and Fisher rats, participation of potassium channels in neurogenic neuroprotection cannot be excluded. PMID- 10526151 TI - Direct evidence of cytoplasmic delivery of PKC-alpha, -epsilon and -zeta pseudosubstrate lipopeptides: study of their implication in the induction of apoptosis. AB - Protein kinases C (PKC) are serine/threonine kinase enzymes involved in the mechanism of cell survival. Their pseudosubstrate sequences are autoinhibitory domains, which maintain the enzyme in an inactive state in the absence of allosteric activators, thus representing an attractive tool for the modulation of different PKC isoforms. Here, we report the use of palmitoylated modified PKC alpha, -epsilon, and -zeta pseudosubstrate peptides, and determine their intracellular distribution together with their respective PKC isoenzymes. Finally, we propose that the differential distribution of the peptides is correlated with a selective induction of apoptosis and therefore argues for different involvement of PKC isoforms in the anti-apoptotic program. PMID- 10526152 TI - Identification and molecular characterization of BP75, a novel bromodomain containing protein. AB - We here describe the identification and characterization of a novel bromodomain containing protein, the bromodomain protein of 75 kDa (BP75). Initially, we identified BP75 in a two-hybrid screening for proteins that interact with the first PDZ (acronym for post-synaptic density protein PSD-95, Drosophila discs large tumor suppressor DlgA and the tight junction protein ZO-1) domain in protein tyrosine phosphatase-BAS-like (PTP-BL). We found that BP75 is expressed ubiquitously and show that both BP75 and a PTP-BL deletion mutant consisting of the first PDZ domain are located mainly in the nucleus, although cytoplasmic localization is also evident. Full-length PTP-BL, on the contrary, is predominantly localized in the cytoplasm, although some basal nuclear staining is observed. The described molecular interaction may reflect a mechanism of coupling submembraneous signalling events and nuclear events. PMID- 10526153 TI - Cathepsin J, a novel murine cysteine protease of the papain family with a placenta-restricted expression. AB - A novel mouse cysteine protease of the papain family was identified by searching the dbEST database. A 1.28 kb full-length cDNA was obtained which contains an open reading frame of 999 nucleotides and encodes a predicted polypeptide of 333 amino acids. The deduced polypeptide exhibits features characteristic of cysteine proteases of the papain type including the highly conserved residues of the catalytic triad, and was hence named cathepsin J. Cathepsin J represents the murine homologue of a previously described rat cathepsin L-related protein. Mature cathepsin J shows 59.3% identity to mouse cathepsin L and contains the characteristic ER(F/W)NIN motif within the propeptide indicating that this protease belongs to the subgroup of cathepsin L-like cysteine proteases. Northern blot analysis of various tissues revealed a placenta-restricted expression. This expression pattern may suggest a role of cathepsin J in embryo implantation and/or placental function. Ctsj was mapped to mouse chromosome 13 in the vicinity of cathepsin L suggesting that cathepsin J may have arisen by gene duplication from cathepsin L or a common ancestral gene. PMID- 10526154 TI - High-resolution immunogold cytochemistry indicates that AQP4 is concentrated along the basal membrane of parietal cell in rat stomach. AB - Gastric parietal cells secrete hydrochloric acid in stomach. Because the secreted HCl solution is isotonic with the plasma fluid, it should accompany the water transport across the membranes of parietal cells. Aquaporins (AQPs) are water channel proteins that play the central role in the cellular handling of water in various mammalian tissues. Using immunocytochemistry, we found that AQP4 was expressed only in parietal cells of rat gastric mucosa. Immunogold electron microscopy study further demonstrated that AQP4 was mostly localized at the basal membrane of parietal cells. In the basal membrane, AQP4 was prominently enriched on the portion contacting with the basement membrane surrounding gastric glands. These results suggest that the contact between basement membrane and basal membrane may generate the signal involved in the targeting of AQP4 in gastric parietal cells. PMID- 10526155 TI - PDMP blocks the BFA-induced ADP-ribosylation of BARS-50 in isolated Golgi membranes. AB - We reported that an inhibitor of sphingolipid biosynthesis, D, L-threo-1-phenyl-2 decanoylamino-3-morpholino-1-propanol (PDMP), blocks brefeldin A (BFA)-induced retrograde membrane transport from the Golgi complex to the endoplasmic reticulum (ER) (Kok et al., 1998, J. Cell Biol. 142, 25-38). We now show that PDMP partially blocks the BFA-induced ADP-ribosylation of the cytosolic protein BARS 50. Moreover, PDMP does not interfere with the BFA-induced inhibition of the binding of ADP-ribosylation factor (ARF) and the coatomer component beta-coat protein to Golgi membranes. These results are consistent with a role of ADP ribosylation in the action of BFA and with the involvement of BARS-50 in the regulation of membrane trafficking. PMID- 10526156 TI - Rho-specific binding and guanine nucleotide exchange catalysis by KIAA0380, a dbl family member. AB - Several guanine nucleotide exchange factors (GEFs) for Rho-GTPases have been identified, all of them containing a Dbl homology (DH) and pleckstrin homology (PH) domain, but exhibiting different specificities to the Rho family members, Rho, Rac and Cdc42. We report here that KIAA0380, a protein with a tandem DH/PH domain, an amino-terminal PDZ domain and a regulator of G protein signalling (RGS) homology domain, is a specific GEF for RhoA, but not for Rac1 and Cdc42, as determined by GDP release, guanosine 5'-O-(3-thio)triphosphate (GTPgammaS) binding and protein binding assays. When expressed in J82 cells, DH/PH domain containing forms of KIAA0380 induced actin stress fibers, whereas expression of the RGS homology domain prevented lysophosphatidic acid (LPA)-induced stress fiber formation. PMID- 10526157 TI - Creation of a functional S-nitrosylation site in vitro by single point mutation. AB - Here we show that in extrahepatic methionine adenosyltransferase replacement of a single amino acid (glycine 120) by cysteine is sufficient to create a functional nitric oxide binding site without affecting the kinetic properties of the enzyme. When wild-type and mutant methionine adenosyltransferase were incubated with S nitrosoglutathione the activity of the wild-type remained unchanged whereas the activity of the mutant enzyme decreased markedly. The mutant enzyme was found to be S-nitrosylated upon incubation with the nitric oxide donor. Treatment of the S nitrosylated mutant enzyme with glutathione removed most of the S-nitrosothiol groups and restored the activity to control values. In conclusion, our results suggest that functional S-nitrosylation sites can develop from existing structures without drastic or large-scale amino acid replacements. PMID- 10526158 TI - Loss of STAT1 expression confers resistance to IFN-gamma-induced apoptosis in ME180 cells. AB - Interferon gamma (IFN-gamma) induces apoptosis in many tumor cell lines and sensitizes tumor cells to apoptosis by tumor necrosis factor family members. IFN gamma induces the expression of many early response genes such as interferon regulatory factor-1 (IRF-1) by activation of signal transducer and activator of transcription (STAT) factor proteins. We found that ME180 cells became resistant to IFN-gamma-induced cell death after 4-5 passages in culture. These resistant cells were characterized by a loss of STAT1 expression and a loss of inducible IRF-1 expression. We describe for the first time the emergence of a STAT1 deficient ME180 cell line. PMID- 10526159 TI - Demonstration of glycosaminoglycans in Caenorhabditis elegans. AB - A considerable amount (approximately 1.6 microg from 1 mg of dried nematode) of non-sulfated chondroitin, two orders of magnitude less yet an appreciable amount of heparan sulfate, and no hyaluronate were found in Caenorhabditis elegans nematodes. The chondroitin chains were heterogeneous in size, being shorter than that of whale cartilage chondroitin sulfate. The disaccharide composition analysis of heparan sulfate revealed diverse sulfation including glucosamine 2-N sulfation, glucosamine 6-O-sulfation and uronate 2-O-sulfation. These results imply that chondroitin and heparan sulfate are involved in fundamental biological processes. PMID- 10526160 TI - The fMet-tRNA binding domain of translational initiation factor IF2: role and environment of its two Cys residues. AB - Mutations of the cysteines (positions 668 and 714) were generated in the IF2 C domain of Bacillus stearothermophilus translation initiation factor IF2. The corresponding proteins were characterized functionally and structurally. Most (yet not all) amino acid replacements at both positions resulted in severe reduction of the fMet-tRNA binding activity of IF2 C without grossly altering its structure. Our work demonstrates that: (a) both Cys residues are buried within an hydrophobic core and not accessible to protonation or chemical substitution, (b) neither Cys is functionally essential and (c) both Cys residues are located near the active site, probably without participating directly in fMet-tRNA binding. PMID- 10526161 TI - The regiochemical distribution of positive charges along cholesterol polyamine carbamates plays significant roles in modulating DNA binding affinity and lipofection. AB - We have quantified the effects of the regiochemical distribution of positive charges along the polyamine moiety in lipopolyamines for DNA molecular recognition. High affinity binding leads to charge neutralisation, DNA condensation and ultimately to lipofection. Binding affinities for calf thymus DNA were determined using an ethidium bromide displacement assay and condensation was detected by changes in turbidity using light scattering. The in vitro transfection competence of cholesterol polyamine carbamates was measured in CHO cells. In the design of DNA condensing and transfecting agents for non-viral gene therapy, the interrelationship of ammonium ions, not just their number, must be considered. PMID- 10526162 TI - Superoxide anion inhibits drug-induced tumor cell death. AB - Intracellular superoxide (O(2)*- was manipulated in M14 melanoma cells by overexpression or repression of Cu/Zn SOD using a tetracycline-inducible expression system. Scavenging intracellular O(2)*- increased tumor cell sensitivity to daunorubicin, etoposide, and pMC540, whereas expression of the antisense SOD mRNA significantly decreased cell sensitivity to drug treatment. Whereas Cu/Zn SOD overexpressing cells exhibited higher activation of the executioner caspase 3 upon drug exposure, caspase 3 activation was significantly lower when Cu/Zn SOD was repressed by antisense expression. These data show that intracellular O(2)*- regulates tumor cell response to drug-induced cell death via a direct or indirect effect on the caspase activation pathway. PMID- 10526163 TI - Alignment of a sparse protein signature with protein sequences: application to fold prediction for three small globulins. AB - A novel algorithm has been developed for scoring the match between an imprecise sparse signature and all the protein sequences in a sequence database. The method was applied to a specific problem: signatures were derived from the probable folding nucleus and positions obtained from the determined interactions that occur during the folding of three small globular proteins and points of inter element contact and sequence comparison of the actual three-dimensional structures of the same three proteins. In the case of two of these, lysozyme and myoglobin, the residues in the folding nucleus corresponded well to the key residues spotted by examination of the structures and in the remaining case, barnase, they did not. The diagnostic performance of the two types of signatures were compared for all three proteins. The significance of this for the application of an understanding of the protein folding mechanisms for structure prediction is discussed. The algorithm is generic and could be applied to other user-defined problems of sequence analysis. PMID- 10526164 TI - Kinetics of apolipoprotein E isoforms-binding to the major glycosaminoglycans of the extracellular matrix. AB - Apolipoprotein E (apoE), a key lipid transport protein, displays a heparin binding property that is critical in several apoE functions. The kinetics of the interaction between apoE isoforms and glycosaminoglycans (GAGs) were studied using surface plasmon resonance. The dissociation constant of equilibrium K(D) for apoE3-heparin interaction was estimated to be 12 nM for apoE3 and three common apoE isoforms revealed similar affinities for heparin. ApoE binds to GAGs in the following order: heparin>heparan sulfate>dermatan sulfate>chondroitin sulfate. The affinity parameter of the binding of low molecular weight heparins to apoE is correlated with the chain length. The effective number Z of electrostatic interactions between plasma apoE3 and heparin was assessed to be three. Metal chelators were able to diminish apoE-binding to heparin, suggesting some stabilizing effect of metal ions while reconstitution with lipids did not affect binding affinities for heparin, suggesting that the N-terminal heparin binding site is responsible for apoE-containing lipoprotein interactions with heparin. PMID- 10526165 TI - Autocatalytic processing of recombinant human procathepsin B is a bimolecular process. AB - Cathepsin B and other lysosomal cysteine proteinases are synthesized as inactive zymogens, which are converted to their mature forms by other proteases or by autocatalytic processing. Procathepsin B autoactivation was shown in vitro at pH 4.5 to be a bimolecular process with K(s) and k(cat) values of 2.1+/-0.9 microM and 0.12+/-0.02 s(-1)6.0. However, in the presence of 0.5 microg/ml of dextran sulfate, relatively rapid processing is observed even at pH 6.5 (t(1/2) approximately 90 min), suggesting that glycosaminoglycans are involved in in vivo processing of lysosomal cysteine proteases. PMID- 10526166 TI - A product of growth arrest-specific gene 6 modulates scavenger receptor expression in human vascular smooth muscle cells. AB - Although Gas6 is identified as a growth factor for vascular smooth muscle cells (VSMCs), its roles in these cells have not been clearly elucidated. To examine the role of Gas6 in atherosclerosis, we examined the effects of Gas6 on scavenger receptor family expression in VSMCs. Scavenger receptor class A, one of the scavenger receptor family members, was upregulated in VSMCs by Gas6. Furthermore, the atherogenic lipoprotein, oxidized LDL, induced Gas6 production in these cells. These results indicate that Gas6 plays an important role in foam cell formation in human VSMCs. PMID- 10526167 TI - The tolbutamide site of SUR1 and a mechanism for its functional coupling to K(ATP) channel closure. AB - Micromolar concentrations of tolbutamide will inhibit (SUR1/K(IR)6. 2)(4) channels in pancreatic beta-cells, but not (SUR2A/K(IR)6.2)(4) channels in cardiomyocytes. Inhibition does not require Mg(2+) or nucleotides and is enhanced by intracellular nucleotides. Using chimeras between SUR1 and SUR2A, we show that transmembrane domains 12-17 (TMD12-17) are required for high-affinity tolbutamide inhibition of K(ATP) channels. Deletions demonstrate involvement of the cytoplasmic N-terminus of K(IR)6.2 in coupling sulfonylurea-binding with SUR1 to the stabilization of an interburst closed configuration of the channel. The increased efficacy of tolbutamide by nucleotides results from an impairment of their stimulatory action on SUR1 which unmasks their inhibitory effects. The mechanism of inhibition of beta-cell K(ATP) channels by sulfonylureas during treatment of non-insulin-dependent diabetes mellitus thus involves two components, drug-binding and conformational changes within SUR1 which are coupled to the pore subunit through its N-terminus and the disruption of nucleotide dependent stimulatory effects of the regulatory subunit on the pore. These findings uncover a molecular basis for an inhibitory influence of SUR1, an ATP binding cassette (ABC) protein, on K(IR)6.2, a ion channel subunit. PMID- 10526168 TI - The lactose analog GalNAcbeta1-->4Glc is present in bovine colostrum. Enzymatic basis for its occurrence. AB - We have isolated from bovine colostrum the lactose analog GalNAcbeta1-->4Glc. The enzymatic basis for its occurrence was studied by assaying the activities of GlcNAcbeta-R beta4-N-acetylgalactosaminyltransferase (beta4-GalNAcT) and GlcNAcbeta-R beta4-galactosyltransferase (beta4-GalT) in primary milk and several lactating bovine mammary gland fractions. As the beta4-GalNAcT, which appears to be tightly membrane bound, is induced by the milk protein alpha-lactalbumin (alpha-LA) to act on Glc, it is concluded that beta4-GalNAcT is responsible for the synthesis of GalNAcbeta1-->4Glc in the gland. The comparatively low level (15 20 mg/l) at which this disaccharide is produced may be due to the relatively poor interaction of beta4-GalNAcT with alpha-LA as well as to the fact that alpha-LA does not inhibit the action of the enzyme on N-acetylglucosaminides. PMID- 10526169 TI - Up-regulation of multidrug resistance-associated protein 2 (MRP2) expression in rat hepatocytes by dexamethasone. AB - Regulation of multidrug resistance-associated protein (MRP2) expression in response to dexamethasone (DEX) was analyzed using mainly primary rat hepatocytes. Enhanced levels of MRP2 mRNAs associated with increased amounts of a 190 kDa MRP2 were found in cultured DEX-treated hepatocytes; similarly, administration of DEX to rats (100 mg/kg, i.p.) led to a marked increase of hepatic amounts of MRP2 mRNAs. Maximal induction of MRP2 expression in DEX treated primary hepatocytes was reached with 10(-5) M DEX, a concentration higher than that (10(-7) M) required for maximal up-regulation of tyrosine aminotransferase (TAT), a typical glucocorticoid receptor-regulated enzyme. In addition, the anti-glucocorticoid compound RU486 failed to inhibit MRP2 induction caused by DEX whereas it fully blocked that of TAT. These findings therefore demonstrate that DEX is a potent inducer of MRP2 expression in rat hepatocytes through a mechanism that seems not to involve the classical glucocorticoid receptor pathway. PMID- 10526170 TI - Purification and functional reconstitution of a truncated human Na(+)/glucose cotransporter (SGLT1) expressed in E. coli. AB - A truncated human Na(+)/glucose cotransporter (C(5), residues 407-664) was expressed and purified from Escherichia coli using a GST fusion vector and glutathione affinity chromatography. The truncated transporter (C(5)) was cleaved from GST-C(5) by Factor Xa proteolysis and purified by gel filtration chromatography. Up to 1 mg of purified GST-C(5) was obtained from 1 l bacterial culture. Reconstitution of both GST-C(5) and C(5) proteins into lipid vesicles resulted in 2.5-fold higher initial uptake rates of [(3)H]D-glucose into C(5) proteoliposomes than into liposomes. Transport was stereospecific, saturable, and inhibited by phloretin. These properties are similar to those obtained for C(5) in Xenopus laevis oocytes, and provide additional evidence that the five C terminal transmembrane helices in SGLT1 form the sugar translocation pathway. PMID- 10526171 TI - Suppression of okadaic acid-induced apoptosis by overexpression of calpastatin in human UV(r)-1 cells. AB - Proteolytic systems have various involvements in apoptotic pathways. To understand the role of calpain in apoptosis, calpastatin, a specific inhibitor of calpain, was overexpressed in human UV(r)-1 fibroblasts by transfection of its cDNA. The elevated expression of calpastatin resulted in decreased survival in the presence of okadaic acid (OA) but in no apparent alteration in the sensitivity toward other drugs such as 5-fluorouracil, mitomycin C and methotrexate. After treatment with OA, a typical apoptotic DNA ladder was observed in control vector-transfected cells but not in calpastatin-transfected cells. This indicates that OA-induced apoptosis was suppressed by overexpression of calpastatin. Further immunoblot analysis showed that the OA-induced hyperphosphorylation of c-Jun was inhibited in calpastatin-transfected cells. This might be involved in the resistance to OA-induced cell death in calpastatin overproducing cells. PMID- 10526172 TI - PcpA, which is involved in the degradation of pentachlorophenol in Sphingomonas chlorophenolica ATCC39723, is a novel type of ring-cleavage dioxygenase. AB - The pentachlorophenol (PCP) mineralizing bacterium Sphingomonas chlorophenolica ATCC39723 degrades PCP via 2,6-dichlorohydroquinone (2,6-DCHQ). The pathway converting PCP to 2,6-DCHQ has been established previously; however, the pathway beyond 2,6-DCHQ is not clear, although it has been suggested that a PcpA plays a role in 2, 6-DCHQ conversion. In this study, PcpA expressed in Escherichia coli was purified to homogeneity and shown to have novel ring-cleavage dioxygenase activity in conjunction with hydroquinone derivatives, and converting 2,6-DCHQ to 2-chloromaleylacetate. PMID- 10526173 TI - Conservation of the central proline-rich (PxxP) motifs of human immunodeficiency virus type 1 Nef protein during the disease progression in two hemophiliac patients. AB - The nef gene is considered to play a crucial role in the development of acquired immunodeficiency syndrome (AIDS). In this study, we analyzed the sequence of nef quasispecies obtained from replication-competent HIV-1 isolates from two Japanese hemophiliac patients infected with HIV-1. At least 10 nef clones were isolated at each time point and a total of 75 individual nef quasispecies were sequenced. We observed a gradual increase in genetic diversity of the nef gene over time. Among the various functional regions of Nef protein, myristoylation site and the central PXXP (SH3 ligand) motifs were well conserved. The scattered regions responsible for downregulation of CD4 and class I MHC were also conserved. These data suggest that these functions of Nef may be involved throughout the disease process. PMID- 10526175 TI - Mossbauer studies of Escherichia coli biotin synthase: evidence for reversible interconversion between [2Fe-2S](2+) and [4Fe-4S](2+) clusters. AB - The nature and properties of the iron-sulphur (Fe-S) cluster in as-prepared and reduced biotin synthase of Escherichia coli have been investigated by Mossbauer spectroscopy. Our data clearly demonstrate that in the as-prepared sample, the cluster is present as [2Fe-2S](2+) with isomer shift, delta = 0.29 mm/s and quadrupole splitting, DeltaE(Q) = 0.53 mm/s, indicating incomplete cysteinyl-S coordination. Anaerobic reduction by dithionite in the presence of 55% (v/v) glycerol converts this form to [4Fe-4S](2+) (delta = 0.45 mm/s and DeltaE(Q) = 1.11 mm/s) and is accompanied by some destruction to Fe(2+). This cluster conversion is reversible and when exposed to air, the [4Fe-4S](2+) cluster is quantitatively reconverted to the [2Fe-2S](2+) cluster without any further cluster degradation. PMID- 10526174 TI - Functional complementation of the Schizosaccharomyces pombe wis1 mutant by Arabidopsis MEK1 and non-catalytic enhancement by CTR1. AB - Arabidopsis thaliana MEK1 encodes a MAPKK homolog whose role in plants is currently unknown. High (but not low) expression of MEK1 rescued the Deltawis1 (MAPKK) mutant of the Schizosaccharomyces pombe Win1/Wis4-Wis1-Sty1 stress activated MAPK pathway. Rescue was dependent upon upstream and downstream components of the pathway, suggesting that MEK1 might function in a homologous MAPK pathway in plants. When MEK1 was expressed at a low level, rescue of Deltawis1 was achieved by co-expressing Arabidopsis CTR1 (a putative MAPKK kinase (MAPKKK)). CTR1 constructs alone did not rescue the pathway, indicating that CTR1 augmented MEK1 function. Further data indicated that this enhancement was not due to CTR1 kinase activity. PMID- 10526176 TI - Characterization of recombinant human glucuronyltransferase I involved in the biosynthesis of the glycosaminoglycan-protein linkage region of proteoglycans. AB - We characterized the recombinant glucuronyltransferase I (GlcAT-I) involved in the glycosaminoglycan-protein linkage region biosynthesis. The enzyme showed strict specificity for Galbeta1-3Galbeta1-4Xyl, exhibiting negligible incorporation into other galactoside substrates including Galbeta1-3Galbeta1-O benzyl, Galbeta1-4GlcNAc and Galbeta1-4Glc. A comparison of the GlcAT-I with another beta1,3-glucuronyltransferase involved in the HNK-1 epitope biosynthesis revealed that the two beta1,3-glucuronyltransferases exhibited distinct and no overlapping acceptor substrate specificities in vitro. Nevertheless, the transfection of the GlcAT-I cDNA into COS-1 cells induced the significant expression of the HNK-1 epitope. These results suggested that the high expression of the GlcAT-I gene rendered the cells capable of synthesizing the HNK-1 epitope. PMID- 10526177 TI - Inhibition of tau phosphorylating protein kinase cdk5 prevents beta-amyloid induced neuronal death. AB - The key target of this study was the tau protein kinase II system (TPK II) involving the catalytic subunit cdk5 and the regulatory component p35. TPK II is one of the tau phosphorylating systems in neuronal cells, thus regulating its functions in the cytoskeletal dynamics and the extension of neuronal processes. This research led to demonstration that the treatment of rat hippocampal cells in culture with fibrillary beta-amyloid (Abeta) results in a significant increase of the cdk5 enzymatic activity. Interestingly, the data also showed that the neurotoxic effect of 1-20 microM Abeta on primary cultures markedly diminished with co-incubation of hippocampal cells with the amyloid fibers plus the cdk5 inhibitor butyrolactone I. This inhibitor protected brain cells against Abeta induced cell death in a concentration dependent fashion. Moreover, death was also prevented by a cdk5 antisense probe, but not by an oligonucleotide with a random sequence. The cdk5 antisense also reduced neuronal expression of cdk5 compared with the random oligonucleotide. The studies indicate that cdk5 plays a major role in the molecular path leading to the neurodegenerative process triggered by the amyloid fibers in primary cultures of rat hippocampal neurons. These findings are of interest in the context of the pathogenesis of Alzheimer's disease. PMID- 10526178 TI - Analysis by atomic force microscopy of Med8 binding to cis-acting regulatory elements of the SUC2 and HXK2 genes of saccharomyces cerevisiae. AB - Med8 protein is a regulator that specifically binds to upstream activating sequences (UASs) of SUC2 promoter, to downstream repressing sequences (DRSs) of the HXK2 gene and to the carboxy-terminal domain of the RNA polymerase II. Atomic force microscopy has allowed for direct visualization of Med8 interactions with a 305 bp fragment of SUC2 promoter and with a 676 bp fragment of HXK2 gene, containing respectively the UASs and DRSs regulatory regions. This approach has provided complementary information about the position and the structure of the DNA-protein complexes. Med8 binding to DNA results in total covering of one of the two existing 7 bp motives (consensus, (A/C)(A/G)GAAAT) in the studied DNA fragments. No preference for binding either of the two UASs of SUC2 promoter as well as for the two DRSs of HXK2 gene has been found. We also discuss whether this protein works as dimer or as a monomer. PMID- 10526179 TI - Cloning and characterization of two novel aldo-keto reductases (AKR1C12 and AKR1C13) from mouse stomach. AB - In contrast to hepatic hydrosteroid dehydrogenases (HSDs) of the aldo-keto reductase family (AKR1C), little is known about a stomach one. From a mouse stomach cDNA library, we isolated two clones encoding proteins of 323 amino acid residues. They exhibited 93.2% amino acid sequence identity and 64-68% with any known HSDs. Recombinant proteins expressed in Escherichia coli reduced 9,10 phenanthraquinone with NAD(P)H as cofactor. The mRNAs were exclusively expressed in stomach, liver and ileum. The present study demonstrates that these proteins are new members of the HSD subfamily and they are named AKR1C12 and AKR1C13. Immunohistochemical analysis suggests that they are involved in detoxification of xenobiotics in the stomach. PMID- 10526180 TI - A stability transition at mildly acidic pH in the alpha-hemolysin (alpha-toxin) from Staphylococcus aureus. AB - The effects of mildly acidic conditions on the free energy of unfolding (DeltaG(u)(buff)) of the pore-forming alpha-hemolysin (alphaHL) from Staphylococcus aureus were assessed between pH 5.0 and 7.5 by measuring intrinsic tryptophan fluorescence, circular dichroism and elution time in size exclusion chromatography during urea denaturation. Decreasing the pH from 7.0 to 5.0 reduced the calculated DeltaG(u)(buff) from 8.9 to 4.2 kcal mol(-1), which correlates with an increased rate of pore formation previously observed over the same pH range. It is proposed that the lowered surface pH of biological membranes reduces the stability of alphaHL thereby modulating the rate of pore formation. PMID- 10526181 TI - Feedback inhibition of epithelial Na(+) channels in Xenopus oocytes does not require G(0) or G(i2) proteins. AB - Regulation of amiloride-sensitive epithelial Na(+) channels (ENaC) is a prerequisite for coordination of electrolyte transport in epithelia. Downregulation of Na(+) conductance occurs when the intracellular Na(+) concentration is increased during reabsorption of electrolytes, known as feedback inhibition. Recent studies have demonstrated the involvement of alphaG(0) and alphaG(i2) proteins in the feedback control of ENaC in mouse salivary duct cells. In this report, we demonstrate that Na(+) feedback inhibition is also present in Xenopus oocytes after expression of rat alpha,beta, gamma-ENaC. Interfering with intracellular alphaG(0) or alphaG(i2) signaling by coexpression of either constitutively active alphaG(0)/alphaG(i2) or dominant negative alphaG(0)/alphaG(i2) and by coinjecting sense or antisense oligonucleotides for alphaG(0) had no impact on Na(+) feedback. Moreover, no evidence for involvement of the intracellular G protein cascade was found in experiments in which a regulator of G protein signaling (RGS3) or beta-adrenergic receptor kinase (betaARK) was coexpressed together with alpha,beta, gamma-ENaC. Although some experiments suggest the presence of an intracellular Na(+) receptor, we may conclude that Na(+) feedback in Xenopus oocytes is different from that described for salivary duct cells in that it does not require G protein signaling. PMID- 10526182 TI - Evidence that fructose 1,6-bisphosphate specifically protects the alpha-subunit of pyrophosphate-dependent 6-phosphofructo-1-phosphotransferase against proteolytic degradation. AB - Pyrophosphate-dependent 6-phosphofructo-1-phosphotransferase (PFP) consists of alpha (regulatory) and beta (catalytic) subunits. The alpha-subunit was previously reported to be much more susceptible to tryptic digestion than the beta-subunit. In this study, ligand-induced protection of PFP subunits against proteolysis by subtilisin was investigated in vitro and the data obtained demonstrated that fructose 1,6-bisphosphate (Fru-1,6-P(2)), while exerting negligible effect on the beta-subunit, remarkably protected the alpha-subunit against proteolytic degradation. Western blot analysis revealed a good correlation between the Fru-1,6-P(2) concentration and the degree of corresponding protection on the alpha-subunit against proteolysis. In contrast, none of other examined ligands including fructose 2,6-bisphosphate, fructose 6 phosphate and pyrophosphate had such protection on the alpha-subunit. This finding (1) indicates that the stability of the alpha-subunit can be selectively increased by Fru-1,6-P(2), and (2) suggests that Fru-1,6-P(2) is likely a special effector of the alpha-subunit. PMID- 10526184 TI - The yeast Rgd1p is a GTPase activating protein of the Rho3 and rho4 proteins. AB - The RGD1 gene, identified during sequencing of the Saccharomyces cerevisiae genome, encodes a protein with a Rho-GTPase activating protein (GAP) domain at the carboxy-terminal end. The Rgd1 protein showed two-hybrid interactions with the activated forms of Rho2p, Rho3p and Rho4p. Using in vitro assays, we demonstrated that Rgd1p stimulated the GTPase activity of both Rho3p and Rho4p; no stimulation was observed on Rho2p. In addition, the rho3Deltargd1Delta double mutant exhibited a dramatic growth defect compared to the single mutants, suggesting that Rgd1p has a GAP activity in vivo. The present study allowed the identification of the first GAP of Rho3p and Rho4p. PMID- 10526183 TI - Cyclooxygenase-2 activity is necessary for the angiogenic properties of oncostatin M. AB - Macrophages play a major role in angiogenesis. We recently reported that oncostatin M (OSM), a cytokine of the interleukin (IL)-6 family secreted by macrophages, has a potent angiogenic activity on human microvascular endothelial cells (HMEC-1), but has no effect on macrovascular cells (human umbilical vein endothelial cells (HUVECs)). In this work, we show that in HMEC-1, OSM (0.5-2.5 ng/ml), leukemia inhibitory factor (LIF) (25 ng/ml), bFGF (25 ng/ml) and IL-1beta (5 ng/ml) induced production of cyclooxygenase (COX)-2. In contrast, in HUVECs, neither OSM nor LIF induced COX-2 mRNA, suggesting that COX-2 might be implicated in the angiogenic activity of OSM. This was confirmed by the inhibiting effect on OSM-induced HMEC-1 proliferation of specific COX-2 inhibitors. In vivo studies confirmed this findings. We conclude that induction of COX-2 by OSM is necessary for its angiogenic activity, but is not sufficient since IL-1beta, which also induces COX-2 in HMEC-1, has only a poor proliferative effect. PMID- 10526185 TI - The conserved undecapeptide shared by thiol-activated cytolysins is involved in membrane binding. AB - Thiol-activated cytolysins share a conserved hydrophobic, Trp-rich undecapeptide that is suggested to be involved in membrane binding and intercalation. The neutralizing monoclonal antibody PLY-5 recognizes all members of this toxin family and peptide mapping assigned its epitope to the undecapeptide motif. This antibody inhibited binding of the toxins to host cell membranes and the epitope was no longer available for binding when a preformed toxin/membrane complex was tested. These results confirm the model of cytolysin binding suggested by structural data. PMID- 10526186 TI - Antibody responses in the Guillain-Barre syndrome. PMID- 10526188 TI - Serum antibody against a peripheral nerve myelin ganglioside, LM1, in Guillain Barre syndrome. AB - Serum IgG antibody against LM1, the predominant ganglioside in the human peripheral nerve myelin, was found in 7 out of 140 patients with Guillain-Barre syndrome (GBS) in the acute phase, 1 out of 33 patients with chronic inflammatory demyelinating polyneuropathy (CIDP), and 2 out of 47 patients with Miller Fisher syndrome (MFS). Anti-LM1 IgM antibody was detected only in 2 patients, each with GBS and MFS. The clinical and electrophysiological features of the seven GBS patients with anti-LM1 IgG antibody in the serum were investigated. Six patients recovered to grade 1 within one month of the onset of neuropathy. Electrophysiological studies revealed demyelination in five patients, of which one had axonal damage in addition, whereas sufficient evidence of demyelination or axonal degeneration was not observed in the remaining two. Five had a respiratory tract infection before the onset of neuropathy, and also had serum anti-GQ1b IgG antibody. IgG antibody against LM1 might be involved in the pathogenetic mechanisms of GBS, as a possible demyelinating factor. Presence of both anti-GQ1b and anti-LM1 antibodies may be associated with some infectious agent(s) affecting the respiratory tract. PMID- 10526187 TI - Clinical presentation and outcome of Guillain-Barre and related syndromes in relation to anti-ganglioside antibodies. AB - We correlated the clinical features of 78 patients with Guillain-Barre syndrome (GBS) or related variants, with the presence of serum antibodies to the gangliosides GM1, GM2, GD1a, GD1b and GQ1b in order to determine whether these antibodies may influence the clinical presentation or outcome of GBS. Sixty-three patients had typical GBS (81%), nine a pure motor form (11%), three a paraparetic form (4%), and three had Miller Fisher syndrome (MFS). IgG or IgM (or both) anti ganglioside antibodies were found by ELISA in 37% of patients, including 36% with typical, 33% with pure motor and 100% with MFS. Beside the constant occurrence of anti-GQ1b antibodies in patients with MFS (P<0.00001), the other clinical forms were not associated with a specific anti-ganglioside reactivity. Anti-GM1 and anti-GD1a antibodies tended to be associated with a worse disability at 6 month than other or no reactivity and, similarly to anti-GM2 antibodies, with a more frequent respiratory impairment. Anti-GM2 and anti-GD1b antibodies were always associated with typical GBS and, in all but one patient, with a complete recovery; still they were found in only 13 and 3%, respectively, of the patients with this presentation. Anti-GQ1b antibodies, though always associated with ophthalmoplegia and ataxia in both MFS and GBS, were found in only 36 and 26%, respectively, of patients with these symptoms. Even if different anti-ganglioside antibodies tend to be associated with some clinical features possibly suggesting that they may influence the clinical presentation or outcome, with the exception of anti-GQ1b antibodies for ophthalmoplegia and ataxia, they do not permit to predict the clinical presentation or outcome in individual patients. PMID- 10526189 TI - A computer-based method for continuous single pulse analysis of intracranial pressure waves. AB - INTRODUCTION: The single pulse analysis of intracranial pressure waves provides valuable information about the autoregulative processes after head injury. This method has not been used for routine clinical assessment as yet. Current methods for evaluation of intracranial pressure waves are based on spectral analysis or related techniques. This imposes restrictions on the wave sequences available for the investigation of ICP attributes. Therefore, we have developed a computer based method, which enables continuous analysis of each pulse of the ICP wave, in any clinical setting. METHOD: Firstly, the raw data of the ICP wave is continuously recorded by the Multifunctional Anaesthetic Record System (MARS, Hewlett Packard). The recorded data is then subjected to single pulse wave analysis by our software. Each single pulse is identified by the algorithm. The maximum, minimum and mean value, as well as amplitude and gradient are calculated in each pulse pressure. All conceivable correlations of the listed parameters can be examined. RESULTS: We applied our software in 9 cases with head injury and evaluated the measurements over 59 days (1400 h). More than 7 million single pulse pressures have been analyzed off-line. The software proved to be accurate and easy to apply. It was possible to calculate correlations between the different wave attributes on a broad basis of data. Parameters of special clinical interest were the amplitude of the single pulse pressure and the gradient. CONCLUSION: This method is an improvement on ICP single pulse pressure analysis and facilitates its clinical application. It creates the possibility for continuous long-term analysis of the ICP wave attributes under any clinical condition without loss of data. There are indications that the amplitude and the gradient of the ICP pulse pressure could provide valuable additional information for clinical assessment. However, further evaluation is required. PMID- 10526190 TI - A prospective study on the natural history of multiple sclerosis: clues to the conduct and interpretation of clinical trials. AB - The study's objectives were to assess the predictive significance of different sets of demographic, clinical and extraclinical variables in identifying multiple sclerosis patients with various risk levels of worsening during the follow-up, in order to provide clues to inclusion criteria and selection of primary clinical end-points in therapeutic trials. Two hundred and twenty-four patients at their first diagnosis of multiple sclerosis admitted to our Department between 1983 and 1990 were prospectively followed-up until the end of 1996. We considered as end points time to reach non-reversible disability levels corresponding to EDSS scores of 4.0 and 6.0 and the beginning of a secondary progressive phase in the relapsing-remitting subgroup of patients. For the statistical treatment of our data we used the Kaplan-Meier survival curves and the Cox regression analysis. An initially progressive course and higher basal EDSS scores proved to be the best predictors of unfavorable prognosis; a greater number of functional systems involved at onset as well as higher residual deficits in pyramidal, visual, sphincteric and cerebellar systems were other factors predictive of a poor outcome, whereas sensory system involvement turned out to be favorable. In the relapsing-remitting subgroup, a longer first inter-attack interval was associated with a better prognosis; however, overall number of relapses in the first two years of the disease was of no prognostic value. The presence of oligoclonal banding in the cerebrospinal fluid and a cerebral MRI 'strongly suggestive' or 'suggestive' of MS in the early phases of the disease were associated with a higher probability of a worse outcome. PMID- 10526191 TI - Prolonged postexcitatory inhibition after transcranial magnetic stimulation of the motor cortex in patients with cerebellar ataxia. AB - Postexcitatory inhibition after transcranial magnetic stimulation of the motor cortex (silent period, SP) is supposed to be predominantly mediated by the activation of inhibitory cortical interneurons. Cortical excitability seems to be reduced in patients with cerebellar ataxia. Motor threshold, central motor conduction time and the duration of the silent period after a single magnetic stimulus to the motor cortex on both sides were measured in five patients with cerebellar ataxia of different origin and 18 healthy controls. Duration of SP was highly significantly prolonged in patients compared with controls (P<0.001) while motor threshold and central motor conduction times were not different. Fifteen of 18 control subjects showed late EMG responses after magnetic stimulation but none of the patients did (P<0.001). These findings support the hypothesis that cerebellar lesions activate inhibitory cortical interneurons or cause a disruption of a normally tonic cerebellar excitation to the motor cortex. Silent period measurement appears to be a sensitive diagnostic tool in the neurophysiological examination of cerebellar diseases. PMID- 10526192 TI - Normal cerebrospinal fluid glutathione concentrations in Parkinson's disease, Alzheimer's disease and multiple system atrophy. AB - We measured total glutathione concentrations in the cerebrospinal fluid (CSF) of non-demented Parkinson's disease patients (PD; n=71), demented PD patients (PDD; n=13), multiple system atrophy patients (MSA; n=10), Alzheimer's disease patients (AD; n=17) and age-matched controls (n=21). No statistically significant differences in the mean total CSF glutathione concentrations were found between groups and dopaminomimetic treatment was not found to have any effect on total CSF glutathione levels. Our main conclusion is that total glutathione is not useful as a CSF marker for assumed oxidative stress in patients with PD, MSA or AD. PMID- 10526193 TI - The neurotoxicant, cuprizone, retards the differentiation of oligodendrocytes in vitro. AB - The effects of oxalyldihydrazone (cuprizone) on weanling rodents provided an early protocol for toxic demyelination in vivo, in which degeneration of oligodendrocytes preceded disruption of the myelin sheath, and in which remyelination could take place. We administered cuprizone to oligodendrocyte enriched glial-cell cultures and to mixed glial-cell cultures from neonatal rat brains. The cultures were treated with cuprizone for 1 h and allowed to continue differentiating on subsequent days. Treated cultures and respective control cultures were fixed with 4% paraformaldehyde (w/v) and immunostained with double immunofluorescence. MAbO4 was used to mark precursors and mature oligodendrocytes, and anti-myelin basic protein (MBP) to mark mature oligodendrocytes (O4+/MBP+), as distinguished from precursors, which were O4+/MBP . Cell counts suggested that cuprizone inhibited the maturation of oligodendrocytes without diminishing the numbers of precursors, and appeared to affect the mitochondria in those cells. PMID- 10526194 TI - Acute motor axonal neuropathy and acute motor-sensory axonal neuropathy share a common immunological profile. AB - Griffin and colleagues (Griffin JW, Li CY, Ho TW, Tian M, Gao CY, Xue P, Mishu B, Cornblath DR, Macko C, McKhann GM, Asbury AK. Pathology of motor-sensory axonal Guillain-Barre syndrome. Ann Neurol 1996;39:17-28 [4]) proposed that acute motor axonal neuropathy (AMAN) and acute motor-sensory axonal neuropathy (AMSAN) are part of the spectrum of a single type of immune attack on the axon. In contrast, IgG anti-GM1 antibody is associated closely with AMAN, but whether other IgG anti ganglioside antibodies are associated with this neuropathy is not clear. We investigated whether IgG anti-ganglioside antibodies can be used as immunological markers to differentiate AMAN from acute inflammatory demyelinating polyneuropathy (AIDP) and whether these autoantibodies are present in AMSAN. The frequencies of anti-GM1, anti-GM1b, and anti-GD1a IgG antibodies in 21 AMAN patients were significantly higher than in 19 AIDP patients. Anti-GM1b and anti GD1a IgG, as well as anti-GM1 IgG antibodies, therefore are immunological markers for AMAN. The patients with AMSAN had anti-GM1, anti-GM1b, and anti-GD1a IgG antibodies, indicative that AMAN and AMSAN share a common immunological profile. PMID- 10526195 TI - Olfactory loss in multiple sclerosis. AB - The objectives of the present study were to test odor identification ability in patients with multiple sclerosis (MS) and to examine possible correlations between smell identification test scores and various clinical variables. We performed a case-control study comparing the Cross Cultural Smell Identification Test scores of 40 patients with definite multiple sclerosis with those obtained in 40 age-, sex- and smoking-habit-matched healthy controls. The neurological impairment, the disability, the cognitive performances and the psychological functioning were also assessed. Patients with multiple sclerosis scored significantly poorer than controls on the Cross-Cultural Smell Identification Test (P<0.001). Olfactory function was borderline normal in four (10%) and abnormal in five (12.5%) MS patients, whereas it was normal in all controls (P<0.02). Significant correlations between the smell identification score and symptoms of anxiety (r=-0.43, P=0.006), depression (r=-0.42, P=0. 008) and severity of neurological impairment (r=-0.32, P=0.05) were found. Only two (5%) patients with multiple sclerosis reported having episodes of smell loss, suggesting a low level of awareness of this problem. Although smell changes are rarely reported, olfactory function is impaired in a considerable number of patients with MS. The observed association between decreased odor identification ability and symptoms of anxiety and depression in our patients suggests that mood and anxiety disorders have to be considered in assessing olfaction in MS patients. Clearly, smell disturbances deserve greater attention from health professionals and caregivers dealing with such patients. PMID- 10526196 TI - Further study on the specificity and incidence of neutralizing antibodies to interferon (IFN) in relapsing remitting multiple sclerosis patients treated with IFN beta-1a or IFN beta-1b. AB - The development of neutralizing antibodies (NAbs) to interferon (IFN) is a common phenomenon of IFN beta therapy for relapsing-remitting multiple sclerosis (RRMS) patients. Here we examine the specificity of NAbs developed during therapy for RRMS with recombinant interferon (rIFN) beta-1a or rIFN beta-1b, and study the effect of switching from rIFN beta-1a to rIFN beta-1b on the incidence and specificity of NAbs. The relative ability to neutralize rIFN beta-1a and beta-1b was assayed in sera positive for NAbs derived from RRMS patients treated with either rIFN beta-1a (N=9) or rIFN beta-1b (N=16), while the incidence and specificity of NAbs to IFN beta developed during therapy were studied in 50 RRMS patients who were treated for two years with rIFN beta-1a followed by a further year either switching to rIFN beta-1b (N=34) or continuing treatment with rIFN beta-1a (N=16). The results show that all positive sera, independent of the source, may recognize both forms of rIFN beta and that a further year of treatment does not significantly affect the incidence and specificity of the NAbs developed during the first two years of treatment even if treatment is switched to a different type of IFN beta. The data then suggests that it is unlikely that the administration of rIFN beta-1b to anti-rIFN beta-1a NAbs-positive patients can overcome the inhibitory effect exerted by the serum antibodies (and vice versa), and that a further period of treatment with IFN beta-1b in patients previously treated with rIFN beta-1a does not significantly change the pattern of antibody response to IFN beta. PMID- 10526197 TI - Migratory basal ganglia lesions in subacute sclerosing panencephalitis (SSPE): clinical implications of axonal spread. AB - We report a boy with subacute sclerosing panencephalitis (SSPE) who exhibited parkinsonian symptoms four months after onset. The symptoms improved after administration of levodopa. One year after onset, bilateral symmetric lesions appeared in the substantia nigra and the putamen, as observed using magnetic resonance imaging. After a one-year interval, the lesions migrated to the bilateral caudate and the cerebellar dentate nuclei. The series of migratory legions, each of which was connected by axonal pathways originating from the substantia nigra, suggests axonal spread of the SSPE virus. PMID- 10526198 TI - Mutation at codon 210 (V210I) of the prion protein gene in a North African patient with Creutzfeldt-Jakob disease. AB - A point mutation at codon 210 of the prion protein gene (PRNP), resulting in the substitution of isoleucine for valine (V210I) has been found in a 54-year-old Moroccan patient affected with Creutzfeldt-Jakob disease (CJD). This patient is the first carrier of the PRNP V210I mutation reported from North Africa. The clinical presentation of the patient was rather similar to that seen in classical CJD, except that unusual early sensory symptoms were observed. The mother of the proband, aged 72, is a further example of an asymptomatic elderly carrier of the PRNP V210I mutation, suggesting an incomplete penetrance of the disease. PMID- 10526199 TI - Medial temporal lobe metabolic impairment in dementia associated with motor neuron disease. AB - In the course of their disease certain patients with frontotemporal dementia (FTD) develop clinical features compatible with a motor neuron disease (FTD-MND). Previous reports have suggested that the functional pattern is similar in FTD and FTD-MND. However, some neuropathological studies suggest greater involvement of medial temporal regions in FTD-MND than in FTD. Using statistical parametric mapping (SPM96), we compared the metabolic patterns obtained at rest with positron emission tomography in 10 FTD patients and three FTD-MND patients with those obtained from 46 healthy subjects (HS). Mean age, duration of illness and dementia stage did not differ statistically between the FTD and FTD-MND groups. In comparison with HS, both groups showed frontal and anterior temporal hypometabolism at P<0.001. When the FTD-MND group was compared to the FTD group, significant hypometabolism was only observed in bilateral amygdala, bilateral hippocampus, and bilateral enthorinal and parahippocampal regions (Brodmann's areas, BA 28/36) at P<0.005. We found no significant differences in regional glucose uptake when FTD patients were contrasted to FTD-MND patients. Our results suggest statistically comparable frontal and lateral temporal hypometabolism in both conditions but greater impairment of medial temporal lobe activity in FTD MND. Our results and a review of the literature support the hypothesis that there is a functional continuum between classical motor neuron disease (cMND), FTD-MND, and FTD. PMID- 10526200 TI - The base substitution fidelity of HIV-1 reverse transcriptase on DNA and RNA templates probed with 8-oxo-deoxyguanosine triphosphate. AB - We have used 8-O-dGTP, a mutagenic nucleotide generated by oxidative metabolism, to probe the misincorporation potential of HIV-1 reverse transcriptase (RT) during DNA synthesis templated by the same nucleotide sequence as either RNA or DNA. With either template, 8-O-dGMP was misincorporated opposite template A, yielding characteristic A-->C transversions. The error rate with DNA was similar to that with RNA, suggesting that base misincorporation by the RT during first strand and second-strand replication may contribute equally to the HIV-1 base substitution mutation rate. The rate of 8-O-dGMP misincorporation differed by more than 10-fold among the 20 adenines in the M13mp2 template where A-->C transversions can be detected. The transversion distribution was similar with the two templates, indicating that the effects of flanking nucleotides on misincorporation rates were similar. This is consistent with structural and biochemical data suggesting that HIV-1 RT binds RNA x DNA and DNA x DNA template primers in the same orientation. The similarities in error rates and distribution further indicate that, despite differences in the structures of free RNA x DNA and DNA x DNA duplexes (e.g., minor groove dimensions), the polymerase active site that assembles upon substrate binding establishes a similar degree of nucleotide selectivity with both types of template-primers. Comparison of the RT error distribution to that observed with two Pol I family DNA polymerases and a Pol alpha family polymerase revealed common hot and cold spots for misincorporation. This suggests that the local nucleotide sequence influences the nucleotide selectivity of four polymerases in a similar manner, despite their differences in structure, biochemical properties, and functions. PMID- 10526201 TI - Visual quantification of DNA double-strand breaks in bacteria. AB - In this paper, we describe a method for the visualization of double-strand breaks in a single electrostretched Escherichia coli DNA molecule. We also provide evidence that electrostretched or migrated DNA under neutral microgel electrophoresis conditions is made up of individual chromosomes. Using the neutral microgel electrophoresis technique, DNA migration (stretching) was measured and the number of DNA double-strand breaks were counted following exposure of E. coli cells to 0, 12.5, 25, 50, or 100 rad of X-rays. The use of an intense fluorescent dye, YOYO and custom-made slides have helped us in visualizing individual bacterial DNA molecules. Bacterial DNA appears similar in structure compared to electrostretched DNA from human lymphocytes. We were able to detect changes in DNA migration (stretching) induced by an X-ray dose as low as 12.5 rad and an increase in the number of DNA breaks induced by a dose as low as 25 rad. The extent of DNA migration and number of breaks were directly correlated to X-ray dosage. PMID- 10526202 TI - A comparison of gamma and neutron irradiation on Raji cells: effects on DNA damage, repair, cell cycle distribution and lethality. AB - The Comet assay (microgel electrophoresis) was used to study DNA damage in Raji cells, a B-lymphoblastoid cell line, after treatment with different doses of neutrons (0.5 to 16 Gy) or gamma rays (1.4 to 44.8 Gy). A better growth recovery was observed in cells after gamma-ray treatments compared with neutron treatments. The relative biological effectiveness (RBE) of neutron in cell killing was determined to be 2.5. Initially, the number of damaged cells per unit dose was approximately the same after neutron and gamma-ray irradiation. One hour after treatment, however, the number of normal cells per unit dose was much lower for neutrons than for gamma rays, suggesting a more efficient initial repair for gamma rays. Twenty-four hours after treatment, the numbers of damaged cells per unit dose of neutrons or gamma rays were again at comparable level. Cell cycle kinetic studies showed a strong G2/M arrest at equivalent unit dose (neutrons up to 8 Gy; gamma rays up to 5.6 Gy), suggesting a period in cell cycle for DNA repair. However, only cells treated with low doses (up to 2 Gy) seemed to be capable of returning into normal cell cycle within 4 days. For the highest dose of neutrons, decline in the number of normal cells seen at already 3 days after treatment was deeper compared with equivalent unit doses of gamma rays. Our present results support different mechanisms of action by these two irradiations and suggest the generation of locally multiply damaged sites (LMDS) for high linear energy transfer (LET) radiation which are known to be repaired at lower efficiency. PMID- 10526203 TI - Preservation of comet assay slides: comparison with fresh slides. AB - The single cell gel electrophoresis assay (comet assay) is an inexpensive, rapid and highly sensitive method for the determination of DNA damage, crosslinks, and alkaline-labile lesions in individual cells. A limitation of the procedure is that the microelectrophoretic gels must be scored rapidly as the comet configuration deteriorates on storage due to dehydration of the agarose and diffusion of DNA. The objectives of this study were firstly to evaluate drying regimes as rapid and simple methods of preservation of the microgels as close to their original fresh state as possible, and secondly to examine the effects of storage of the slides. Human hepatoma (HepG2) cells challenged for 30 min with hydrogen peroxide (H(2)O(2)) were used in the study. Microgel slides were prepared and evaluated immediately, or after drying with or without a methanol fixation step. Microgels that were dried at a variety of temperatures (22-50 degrees C) and re-hydrated did not differ in the values obtained for H(2)O(2) induced DNA damage when compared to fresh samples. Samples could also be continually dried and re-hydrated over a period of up to 3 months with no obvious loss of information. In conclusion, drying of microgels represents a simple and inexpensive method of preserving comet assay slides. PMID- 10526204 TI - Mutation spectra in Salmonella TA98, TA100, and TA104 of two phenylbenzotriazole mutagens (PBTA-1 and PBTA-2) detected in the Nishitakase River in Kyoto, Japan. AB - Previous studies have identified two potent aromatic amine mutagens in the Nishitakase River, a tributary of the Yodo River, which serves as the main drinking water supply for the Osaka area in Japan. The two potent mutagens are 2 [2-(acetylamino)-4-[bis(2-methoxyethyl)amino]-5-methoxyphenyl]-5-am ino-7-bromo-4 chloro-2H-benzotriazole (PBTA-1) and 2-[2-(acetylamino)-4-[N-(2 cyanoethyl)ethylamino]-5-methoxyphenyl]-5- amino-7-bromo-4-chloro-2H benzotriazole (PBTA-2). PBTA-1 and PBTA-2 are presumed to be formed from azo dyes discharged in a reduced form from dye factories to sewage treatment plants where they become chlorinated and are then discharged into the river. PBTA-1 and PBTA-2 account for 21% and 17% of the mutagenic activity of the Nishitakase River, respectively. Here we determined the mutation spectra induced by these two mutagens in TA98, TA100, and TA104 at 30-35, 8-10, and 2x, respectively, above the background. In TA98, the PBTA compounds produced identical mutation spectra, with 100% of the revertants containing the hotspot 2-base deletion of CG within the (CG)(4) sequence. In TA100, 73% of the revertants were GC-->TA transversions, with most of the remaining being GC-->AT transitions; the spectra produced by the two compounds in TA100 were not significantly different (p=0.8). In TA104, as in TA100, the majority (83%-87%) of the revertants were GC-->TA transversions, with most of the remaining revertants (11%-13%) being AT-->TA transversions. Thus, 83% 87% of the mutations induced by the PBTA compounds in TA104 were at G/C sites. The mutation spectra produced by the two compounds in TA104 were not significantly different (p0.08). PBTA-1 and PBTA-2 are structurally similar and have similar mutagenic potencies and mutation spectra in the respective strains. The mutation spectra produced by the PBTA compounds (100% hotspot deletion in TA98 and primarily GC-->TA transversions in TA100 and TA104) are similar to those produced by other potent aromatic amines, which is the class of compounds from which the PBTA mutagens derive. PMID- 10526205 TI - Plasmid-mediated expression of the UmuDC mutagenesis proteins in an Escherichia coli strain engineered for human cytochrome P450 1A2-catalyzed activation of aromatic amines. AB - The mutagenic actions of many chemicals depend on the activities of bacterial "mutagenesis proteins", which allow replicative bypass of DNA lesions. Genes encoding these proteins occur on bacterial chromosomes and plasmids, often in the form of an operon (such as umuDC or mucAB) encoding two proteins. Many bacterial strains used in mutagenicity testing carry mutagenesis protein genes borne on plasmids, such as pKM101. Our objective was to introduce mutagenesis protein function into Escherichia coli strain DJ4309. This strain expresses recombinant human cytochrome P450 1A2 and NADPH-P450 reductase and carries out the metabolic conversion of aromatic and heterocyclic amines into DNA-reactive mutagens. We discovered that many mutagenesis-protein plasmids severely inhibit the response of strain DJ4309 to 2-amino-3,4-dimethylimid-azo[4,5-f]quinoline (MeIQ), a typical heterocyclic amine mutagen. Among many plasmids examined, one, pGY8294, a pSC101 derivative carrying the umuDC operon, did not inhibit MeIQ mutagenesis. Strain DJ4309 pGY8294 expresses active mutagenesis proteins, as shown by its response to mutagens such as 1-nitropyrene and 4-nitroquinoline 1-oxide (4-NQO), and is as sensitive as the parent strain DJ4309 to P450-dependent mutagens, such as MeIQ and 1-aminopyrene. PMID- 10526206 TI - Telomere instability in a human cancer cell line. AB - Telomere maintenance is essential in immortal cancer cells to compensate for DNA lost from the ends of chromosomes, to prevent chromosome fusion, and to facilitate chromosome segregation. However, the high rate of fusion of chromosomes near telomeres, termed telomere association, in many cancer cell lines has led to the proposal that some cancer cells may not efficiently perform telomere maintenance. Deficient telomere maintenance could play an important role in cancer because telomere associations and nondisjunction have been demonstrated to be mechanisms for genomic instability. To investigate this possibility, we have analyzed the telomeres of the human squamous cell carcinoma cell line SQ-9G, which has telomere associations in approximately 75% of the cells in the population. The absence of detectable telomeric repeat sequences at the sites of these telomere associations suggests that they result from telomere loss. The analysis of telomere length by quantitative in situ hybridization demonstrated that, compared to the human squamous cell carcinoma cell line SCC-61 which has few telomere associations, SQ-9G has more extensive heterogeneity in telomere length and more telomeres without detectable telomeric repeat sequences. The dynamics of the changes in telomere length also demonstrated a higher rate of fluctuation in telomere length, both on individual telomeres and coordinately on all telomeres. These results demonstrate that telomere maintenance can play a role in the genomic instability seen in cancer cells. PMID- 10526207 TI - A genetic program for deletion of foreign DNA from the mammalian genome. AB - Mammalian genomes are in constant jeopardy of invasion by prokaryotic DNA sequences because of their extensive exposure to bacteria; however, mammalian genomes appear to be protected from horizontal transmission of bacterial DNA. Transgenic mice provide a convenient model system for investigating the capacity of mammalian genomes in vivo to retain, silence, and/or reject foreign DNAs. We have previously reported that bacterial genes encoding the Lac repressor (lacI) are subject to sequence-dependent methylation and silencing in the transgenic mouse. In this paper, we report that bacterially derived lacI transgenes, but not their mammalian counterparts, can also be eliminated from the somatic cell DNA of affected animals. This somatic instability is heritable, strain-dependent, and conferred in cis. Our data are consistent with a model of genome surveillance in the mouse which can lead to loss of foreign DNA and which may be analogous to restriction-modification systems that maintain the integrity of the bacterial genome. PMID- 10526208 TI - DNA strand methylation and sister chromatid exchanges in mammalian cells in vitro. AB - Among other targets, DNA demethylating agents are known to affect the sister chromatid exchange (SCE) frequency in mammalian cells in vitro. The SCE increase appears to be maintained for many (10-16) cell cycles after the end of the pulse in a given cell population, unlike SCEs induced by DNA damaging agents. Yet, epigenetic changes (such as demethylation) would not be expected to affect SCE at all. In the present report we challenge the working hypothesis of a relation between SCEs and demethylation by comparing SCE induction during different rounds of replication when the parental strands were normally methylated or demethylated. Azacytidine (AZA), ethionine (ETH), mitomycin-C (MMC), UV irradiation (UV) and hydrogen peroxide (H(2)O(2)) were tested for SCE induction in a Chinese hamster ovary cell line after a single pulse, one or two cell cycles before fixation. Whereas MMC, UV and H(2)O(2) induce SCE in both protocols, AZA and ETH show an effect on SCEs only if administered two cycles before fixation. Because two cell cycles are needed in order to achieve demethylation of the parental DNA strand, the data reported here support our working hypothesis that demethylation in the parental DNA strand, at the level of the replication fork (i.e., the region where SCEs are formed), is responsible for an increase in mistaken ligations of processed damage, eventually yielding an increase in SCEs. PMID- 10526209 TI - Genotoxicity of 3'-azido-3'-deoxythymidine in the human lymphoblastoid cell line, TK6: relationships between DNA incorporation, mutant frequency, and spectrum of deletion mutations in HPRT. AB - Perinatal treatment with 3'-azido-3'-deoxythymidine (AZT) has been found to reduce the rate of maternal-infant transmission of HIV; however, AZT is genotoxic in mammalian cells in vitro and induces tumors in the offspring of mice treated in utero. The purpose of the present study was to investigate the relationships between incorporation of AZT into DNA, and the frequency and spectrum of mutations at the HPRT locus of the human lymphoblastoid cell line, TK6, following in vitro exposures to AZT. Cells were cultured in medium containing 0 or 300 microM AZT for 1, 3, or 6 day(s) (n = 5/group). The effects of exposure duration on incorporation of AZT into DNA and HPRT mutant frequency were determined using an AZT radioimmunoassay and a cell cloning assay, respectively. AZT accumulated in DNA in a supralinear manner, approaching a plateau at 6 days of treatment (101.9 +/- 14.7 molecules AZT/10(6) nucleotides). After 3 days of AZT exposure, HPRT mutant frequency was significantly increased (1.8-fold, p = 0.016) compared to background (mutant frequency = 3.78 x 10(-6)). Multiplex PCR amplification of genomic DNA was used to determine the frequency of exon deletions in HPRT mutant clones from untreated cells versus AZT-treated cells. Molecular analyses of AZT induced mutations revealed a significant difference in the frequency of total gene deletions (44/120 vs. 18/114 in controls, p = 0.004 by the Mann-Whitney U statistic). In fact, the Chi-square test of homogeneity demonstrate that the differences between the control and AZT-treatment groups is attributed mainly to this increase in total gene deletion mutations (p = 0.00001). These data indicate that the primary mechanism of AZT mutagenicity in human TK6 cells is through the production of large deletions which occur as a result of AZT incorporation into DNA and subsequent chain termination. The data imply that perinatal chemoprophylaxis with AZT may put children of HIV-infected women at potential risk for genetic damage. PMID- 10526211 TI - Time to discontinue antigenotoxicity studies of dietary fiber? PMID- 10526210 TI - Induction of sister chromatid exchanges in human peripheral blood lymphocytes by bromoform: investigation of the role of GSTT1-1 polymorphism. AB - Brominated trihalomethanes (THMs) are disinfection by-products present frequently in chlorinated drinking water. Brominated THMs are mutagenic in a variety of systems and are carcinogenic in rodents. The metabolism of brominated THMs is thought to involve a GSH conjugation reaction leading either to formaldehyde or DNA-reactive intermediates via glutathione S-transferase-theta (GSTT1-1), which is polymorphic in humans. In the present study, we have determined the genotoxicity of one of the brominated THMs, bromoform (BF), by measuring its ability to induce sister chromatid exchanges (SCEs) in whole-blood (WB) cultures of human peripheral blood lymphocytes from GSTT1-1+ and GSTT1-1- donors. The results showed no differences in SCEs per cell by BF between GSTT1-1+ and GSTT1-1 individuals when the cells were exposed to 5 x 10(-3) M BF at the beginning of cell culturing (10.8+/-0.85 vs. 10.57+/-0.47, respectively), at the 16th (9.66+/ 0.91 vs. 9.57+/-0.07), or the 24th h (8.21+/-0.61 vs. 8.29+/-0.24) of cell growth. Although GSTT1-1 is expressed in the erythrocytes, the lack of expression of the GSTT1-1 gene in the target cells (lymphocytes) may account for this observation. PMID- 10526212 TI - Genetic interactions between error-prone and error-free postreplication repair pathways in Saccharomyces cerevisiae. AB - Evidence obtained from recent studies supports the existence of an error-free postreplication repair (PRR) and a mutagenesis pathway within the Saccharomyces cerevisiae RAD6 DNA repair group. The MMS2 gene is the only known yeast gene involved in error-free PRR that, when mutated, significantly increases the spontaneous mutation rate. In this study, the mutational spectrum of the mms2 mutator was determined and compared to the wild type strain. In addition, mutagenenic effects and genetic interactions of the mms2 mutator and rev3 anti mutator were examined with respect to forward mutations, frameshift reversions as well as amber and ochre suppressions. It was concluded from these results that the mms2 mutator phenotype is largely dependent on the functional REV3 gene. The synergistic effects of mms2 and rev3 mutations towards killing by a variety of DNA-damaging agents ruled out the possibility that MMS2 simply acts to suppress REV3 activity and favored the hypothesis that MMS2 and REV3 form two alternative subpathways within the RAD6 DNA repair pathway. Taken together, we propose that two pathways represented by MMS2 and REV3 deal with a similar range of endogenous and environmental DNA damage but with different biological consequences, namely, error-free repair and mutagenesis, respectively. PMID- 10526213 TI - Rad51/RecA protein families and the associated proteins in eukaryotes. PMID- 10526214 TI - Protein complexes in nucleotide excision repair. AB - The main pathway by which mammalian cells remove DNA damage caused by UV light and some other mutagens is nucleotide excision repair (NER). The best characterised components of the human NER process are those proteins defective in the inherited disorder xeroderma pigmentosum (XP). The proteins known to be involved in the first steps of the NER reaction (damage recognition and incision excision) are heterotrimeric RPA, XPA, the 6 to 9 subunit TFIIH, XPC-hHR23B, XPG, and ERCC1-XPF. Many interactions between these proteins have been found in recent years using different methods both in mammalian cells and for the homologous proteins in yeast. There are virtually no quantitative measurements of the relative strengths of these interactions. Higher order associations between these proteins in solution and even the existence of a complete "repairosome" complex have been reported, which would have implications both for the mechanism of repair and for the interplay between NER and other cellular processes. Nevertheless, evidence for a completely pre-assembled functional repairosome in solution is inconclusive and the order of action of repair factors on damaged DNA is uncertain. PMID- 10526215 TI - Radioadaptation in Indian muntjac fibroblast cells induced by low intensity laser irradiation. AB - Earlier reports have indicated that an adaptive, protective response to ionizing radiation is inducible by pre-treatment with low intensity laser irradiation (LILI). We have investigated the potential of LILI to induce an adaptive response against the damaging effects of ionizing radiation in Indian muntjac fibroblasts. LILI at 660, but not 820 nm, at 11.5 and 23.0 J/cm2, induced an apparent adaptive response in the form of a reduction in the frequency of radiation-induced chromosome aberrations, but not in cell survival. There was also a trend towards a reduction in the level of single-stranded and double-stranded DNA breaks induced by ionizing radiation when cells were preconditioned with LILI. However, this did not contribute to the reduced chromosome aberration frequency. Further analysis revealed that the reduced aberration frequency was caused by a laser induced extension of G2 delay. The adaptive response was therefore the result of cell cycle modulation by LILI, at a wavelength where there is no known DNA damaging effect to induce the checkpoint mechanisms that are normally responsible for altering cell cycle progression. PMID- 10526216 TI - Adaptive enhancement and kinetics of nucleotide excision repair in humans. AB - An adaptive response, low doses of a mutagen rendering cells more able to subsequently cope with higher doses of that or a related challenging mutagen, enhances nucleotide excision repair in human fibroblasts. After fibroblasts were flashed with 20 J/m2 of UVC, the cyclopyrimidine dimer frequency at any single dinucleotide position remained unchanged for several hours before abruptly displaying first order kinetics of repair. These kinetics were determined by ligation-mediated PCR along exon 9 of the human p53 gene. When a chronic dose of quinacrine mustard (QM) preceded the UVC challenge, the duration of the cyclobutane pyrimidine dimer (CPD) repair lags were reduced by a factor of three and the kinetic half-lives for CPD repair were reduced by a factor of three. The observed repair kinetics are consistent with the following model. The UVC dose required (K(m)) to generate a substrate concentration which half-saturates the cell's repair capacity is 3 J/m2 for the high affinity (6-4) photoproducts and greater than 100 J/m2 for the low affinity cyclobutane dimers. After 20 J/m2 of UVC, the repair enzyme is saturated with (6-4) photoproducts; these competitively inhibit CPD repair by binding all available repair enzyme. After the (6-4)s are repaired, the CPD concentration is less than K(m)(CPD) and so CPD repair kinetics initiate with first order kinetics. QM-induced enhancement, by increasing the concentration, Vmax, of repair enzyme, shortens the duration of (6-4) saturation and increases the rate constant for cyclobutane dimer repair. The data exactly fit the expectations from Michaelis kinetics. Transcription coupled repair is less amenable to Michaelis interpretations and enhanced global repair was almost as rapid as the slightly enhanced transcription coupled repair. We infer that repair enhancement is unable to proportionally increase the number of matrix attachment sites necessary for transcription coupled repair. Understanding competitive inhibition between adduct classes and adaptive enhancement of Vmax is important to understanding the effects of high doses of mutagen mixtures. PMID- 10526217 TI - Analysis of genomic damage in the mutagen-sensitive mus-201 mutant of Drosophila melanogaster by arbitrarily primed PCR (AP-PCR) fingerprinting. AB - DNA repair mechanisms are important to maintain the stability of the genome. In Drosophila melanogaster, the mus-201 gene is required in the excision repair process. To study the contribution of the mus-201 gene in the stability of the Drosophila genome, we have used the arbitrarily primed PCR fingerprinting method (AP-PCR). We have analysed the changes in the genomic DNA fingerprints from the progeny of wild-type males crossed with mus-201 repair-deficient or repair proficient females. After induction of DNA damage with 2-acetylaminofluorene (2 AAF) in the wild-type parental males, quantitative and qualitative differences in the AP-PCR fingerprints were detected between the two crosses, and the estimate of the genomic damage detected by AP-PCR has clearly shown that the mus-201 repair deficiency is associated with an increase of genomic damage. The predominant type of alterations detected by AP-PCR under the mus-201 repair deficient conditions agree with the results obtained in microsatellite PCR analysis, suggesting that the role of the mus-201 gene, necessary in excision repair, is not associated to the mismatch repair process. The work reported here demonstrates that the AP-PCR is a suitable technique to analyse genetic alterations in D. melanogaster and, consequently, can be used to compare the susceptibility to genomic damage of different DNA repair mutants. PMID- 10526218 TI - A UmuD,C-dependent pathway for spontaneous G:C to C:G transversions in stationary phase Escherichia coli mut Y. AB - In Escherichia coli trpA23 bacteria lacking the MutY glycosylase and incubated on plates in the absence of tryptophan, tryptophan-independent mutants continue to arise during incubation over many days. Their appearance is enhanced in umuD+,C+ strains in comparison with strains carrying a deletion through the umu operon and the umuD,C-dependent mutants were greater in number in uvrA bacteria (lacking nucleotide excision repair) than in uvr+ bacteria. Sequencing of mutations occurring in uvrA bacteria revealed the presence of G:C to C:G transversions but only in umuD+,C+ strains. There is thus a pathway in starved bacteria that generates G:C to C:G transversions and requires the inducible UmuD,C proteins. The data are consistent with the occurrence of a lesion, probably 8-oxoguanine, against which guanine may be incorporated during DNA synthesis by "dNTP stabilised" misalignment against the downstream template base. Upon realignment the configuration is substrate for MutY glycosylase which can remove the unmodified guanine. It is hypothesised that UmuD,C proteins are required for primer extension from the mismatch once formed. PMID- 10526219 TI - Deficiency in fast repair process of potentially lethal damage induced by X irradiation in fibroblasts derived from LEC strain rats. AB - The time course for the repair of PLD in LEC and WKAH rat cells irradiated at 5 Gy was examined. In the case of WKAH rat cells, the surviving fraction increased with increasing incubation times after X-irradiation. When hypertonic treatment was performed at each incubation time with 0.5 M NaCl for 20 min, increase in the surviving fractions was not shown. In contrast, no significant recovery of the surviving fraction in LEC rat cells was observed after incubation of irradiated cells with or without 0.5 M NaCl for 20 min. On dose-survival curves, hypertonic treatment with 0.5 M NaCl enhanced radiosensitivity of WKAH rat cells, but not LEC rat cells. Although the surviving fraction of the cells from backcross mice with normal radiosensitivity reduced by treatment with 0.5 M NaCl, the survival fraction was not affected in the cells from backcross mice with higher radiosensitivity by treatment with 0.5 M NaCl. When the cells were X-irradiated and incubated with or without 0.225 M NaCl, the radiosensitivities of LEC and WKAH rat cells treated with 0.225 M NaCl for 4 h were approximately two-fold higher than those of untreated cells. Treatment with caffeine also reduced the surviving fractions of both X-irradiated LEC and WKAH rat cells, compared with those of untreated cells. These results indicated that the slow repair of PLD occurred in LEC rat cells but not the fast repair of PLD. PMID- 10526221 TI - Drug sensitivity spectra in Fanconi anemia lymphoblastoid cell lines of defined complementation groups. AB - Fanconi anemia (FA) is one of several genetic diseases with characteristic cellular hypersensitivity to DNA crosslinking agents which suggest that FA proteins may function as part of DNA repair processes. At the clinical level, FA is characterized by bone marrow failure that affects children at an early age. The clinical phenotype is heterogeneous and includes various congenital malformations as well as cancer predisposition. FA patients are distributed into eight complementation groups suggesting a complex molecular pathway. Three of the eight possible FA genes have been cloned, although their function(s) have not been identified. FA cells are highly sensitive to DNA crosslinking agents (mitomycin C (MMC) and diepoxybutane), with some variability between cell lines. Sensitivity to monofunctional alkylating agents has been reported in some cases, although these studies were performed with genetically unclassified FA cells. To further analyse and characterize the newly identified FA complementation groups, we tested their sensitivity to UV radiation, monofunctional and bifunctional alkylating agents and to the X-ray mimetic drug bleomycin. We found that FA complementation groups D to H show increased sensitivity to the X-ray mimetic drug bleomycin. Furthermore, the single known FA-H cell line shows increased sensitivity to ethylethane sulfonate (EMS), methylmethane sulfonate (MMS) in addition to the characteristic sensitivity to crosslinking agents, suggesting a broader spectrum of drug sensitivities in FA cells. PMID- 10526220 TI - Oxidative DNA damage and mutations induced by a polar photosensitizer, Ro19-8022. AB - The oxidative DNA damage induced by the polar photosensitizer Ro19-8022 in the presence of light was studied and correlated with the associated mutagenicity. Both in isolated DNA and AS52 Chinese hamster ovary cells, photoexcited Ro19-8022 gave rise to a DNA damage profile that was similar to that caused by singlet oxygen: base modifications sensitive to the repair endonuclease Fpg protein, which according to high-performance liquid chromatography (HPLC) analysis were predominantly 8-hydroxyguanine (8-oxoG) residues, were generated in much higher yield than single-strand breaks, sites of base loss (AP sites) and oxidative pyrimidine modifications sensitive to endonuclease III. Fifty percent of the Fpg sensitive modifications were repaired within 2 h. Under conditions that induced 10 Fpg-sensitive modifications per 10(6) bp (six 8-oxoG residues per 10(6) bp), approximately 60 mutations per 10(6) cells were induced in the gpt locus of the AS52 cells. A rather similar mutation frequency was observed when a plasmid carrying the gpt gene was exposed to Ro19-8022 plus light under cell-free conditions and subsequently replicated in bacteria. Sequence analysis revealed that GC-->TA and GC-->CG transversions accounted for 90% of the base substitutions. A significant generation of micronuclei was detectable in AS52 cells exposed to the photosensitizer plus light as well. PMID- 10526222 TI - Gangliosides are the binding substances in neural cells for tetanus and botulinum toxins in mice. AB - We used the knockout mice lacking gangliosides and evaluated their response to tetanus and botulinum toxins. We found that tetanus toxin and botulinum type A or B toxin was less toxic in the knockout mice. We conclude that the toxins bind to the gangliosides on the synapses in the initial step of intoxication prior to penetration of the toxins into the neural cells. PMID- 10526223 TI - Membrane proteins implicated in long-chain fatty acid uptake by mammalian cells: CD36, FATP and FABPm. AB - Long-chain fatty acids can transfer passively across mammalian cell membranes. However, under physiological conditions of low fatty acid to albumin ratios in the circulation, the major fraction of uptake appears to be mediated by a saturable, protein-facilitated component. A simple diffusion process becomes significant at high molar ratios of fatty acid to albumin as the concentration of free fatty acid in solution is increased. Identification of the mammalian membrane fatty acid transporter(s) has been the focus of active investigation by several research groups. In this review we discuss three candidate proteins: FABPm, FAT/CD36 and FATP which have been cloned and are currently being characterized. Recent evidence arguing for an important role of the fatty acid transport step in general metabolism and linking these proteins to physiologic or metabolic abnormalities is described. PMID- 10526224 TI - Biliary phospholipid secretion is not required for intestinal absorption and plasma status of linoleic acid in mice. AB - Biliary phospholipids have been hypothesized to be important for essential fatty acid homeostasis. We tested this hypothesis by investigating the intestinal absorption and the status of linoleic acid in mdr2 Pgp-deficient mice which secrete phospholipid-free bile. In mice homozygous (-/-) for disruption of the mdr2 gene and wild-type (+/+) mice, dietary linoleic acid absorption was determined by 72 h balance techniques. After enteral administration, [(13)C] linoleic acid absorption was determined by measuring [(13)C]-linoleic acid concentrations in feces and in plasma. The status of linoleic acid was determined in plasma and in liver by calculating the molar percentage of linoleic acid and the triene:tetraene ratio. Although plasma concentration of [(13)C]-linoleic acid at 2 h after enteral administration was significantly lower in (-/-) compared to (+/+) mice (P1274; Cromie et al. 1999). Starting with three of these GEME-motif lethal mutations (G1271E, G1271V, M1273V), we have selected for intragenic suppressors, located within the same 3'-region, that prevent expression of the trans-dominant phenotype. RESULTS: We isolated a total of 24 missense mutants and a further 14 frameshift alleles (the latter generating a nested set of C-terminal deletions of the beta subunit) and studied the effect of the missense suppressors in vivo and in vitro. The majority of the second-site substitutions pinpoint highly conserved residues and were allele-specific. In contrast, one particular missense substitution (S1332P) acted on all three primary site mutations whilst not appreciably affecting assembly proficiency, suggesting motif-specific suppression. Two missense substitutions were found to perturb assembly of the beta subunit (M1232T and L1233P) and define a small conserved region (1228-->1233) adjacent to one of the active-site residues identified by affinity-labelling, H1237. The majority of primary mutations were located in three main clusters within the 116 amino acid region. CONCLUSIONS: The importance and functional co-operativity of the three main clusters pinpointed is supported by the present isolation of suppressors of three different GEME primary mutations in the same three regions (whereas the suppressors of G1271V and M1273V are located in all three clusters, those for G1271E are all C-terminal of this residue). Moreover, the location of the suppressors suggests that the GEME and HLVDDK regions are present as alpha-helices in holoenzyme, and that functional co operativity is through one particular face of each helix. PMID- 10526238 TI - Overproduction of elongation factor 1alpha, an essential translational component, causes aberrant cell morphology by affecting the control of growth polarity in fission yeast. AB - BACKGROUND: Elongation factor 1alpha (EF1alpha), an essential component of the eukaryotic translational machinery, has been shown to possess various biochemical and biological activities, including F-actin-binding and -bundling, microtubule- severing, and the activity of making fibroblasts highly susceptible to transformation. However, our understanding of the biological significance of EF1alpha with respect to these various biochemical or biological activities remains limited. Here we report the identification of EF1alpha-encoding genes as genes whose over-expression causes aberrant cell morphology in fission yeast. RESULTS: Overproduction of EF1alpha caused aberrant cell morphology-elliptic, curved or branched-and growth defects in yeast cells at high temperatures. EF1alpha-overproducing cells showed a supersensitivity to the actin inhibitor cytochalasin D and to the tubulin inhibitor thiabendazole. Genetic analyses using cdc mutants suggested that excess EF1alpha disturbed the establishment and the maintenance of growth polarity in the G1 phase by pre- venting the localization of F-actin to the polarized growing site and the organization of microtubules. Results from DNase I column chromatography indicated that EF1alpha was bound to G actin. Indeed, the fission yeast actin was immunoprecipitated along with EF1alpha. Moreover, the temperature sensitivity caused by the overproduction of EF1alpha was restored by co-overproduction of actin. CONCLUSIONS: Fission yeast EF1alpha has the ability to alter the cell morphology of yeast by affecting the control of actin and microtubule cytoskeletons. PMID- 10526239 TI - Multiple mammalian proteasomal ATPases, but not proteasome itself, are associated with TATA-binding protein and a novel transcriptional activator, TIP120. AB - BACKGROUND: SUG1 belongs to proteasomal ATPase. Previous studies have demonstrated that SUG1 is associated with TBP. It is assumed to be involved in transcriptional regulation in addition to proteolysis. In this study, we investigated the association of mammalian SUG1 with TBP in more detail. RESULTS: Pull-down experiments with TBP revealed multiple TBP-interacting proteins (TIPs) that were recovered dependent upon the presence of C-terminal conserved domain of TBP. By 2-D electrophoresis, we identified SUG1 in TIPs. By using far-Western analysis, we identified two proteins that could directly bind to TBP: SUG1 and another proteasomal ATPase (S4). Protein microsequencing and Western blotting identified all the remaining proteasomal ATPases (MSS1, TBP1, TBP7, and SUG2) in the TIP preparations. We present evidence that TBP and at least SUG1, MSS1, and S4 form a complex in the cell. However, no evidence of association of TBP with the 26S proteasome or its 19S regulatory unit was obtained. The molecular mass of the TBP/ATPases-complex, which also included a novel transcription regulatory factor, TIP120, was estimated to be approximately 800 kDa. CONCLUSION: These results suggest that there is a novel multisubunit complex containing TBP and proteasomal ATPases. Based on our findings, we hypothesize that proteasomal ATPases are involved in transcriptional regulation in addition to proteolysis. PMID- 10526240 TI - Requirement of cooperative functions of two repeated death effector domains in caspase-8 and in MC159 for induction and inhibition of apoptosis, respectively. AB - BACKGROUND: The death effector domain (DED), which functions as a domain for a homophilic protein interaction, plays a role in death receptor-mediated apoptosis. Two tandemly repeated DEDs in the prodomain of caspase-8 (Casp8NC-DED) and those in MC159 (viral FLIP) have been shown to positively and negatively regulate apoptosis, respectively, by binding to caspase-8 and/or Fas-associated death domain (FADD). However, characteristics of each DED in Casp8NC-DED and those in MC159 have not been well examined. RESULTS: We analysed deletion and chimera mutants of DEDs derived from Casp8NC-DED and MC159, and found that MC159 and Casp8NC-DED require the combined effects of the two repeated DEDs to exert their binding and biological activities. The carboxy-terminal DED of Casp8NC-DED (Casp8C-DED) has the potential to induce apoptosis, and the amino-terminal DED of MC159 showed a dominant inhibitory effect on apoptosis when combined with Casp8C DED. In addition, the two repeated DEDs in Casp8NC-DED and MC159 were shown to regulate the activities of caspase differently from the caspase recruitment domain (CARD) in the prodomains of caspase-2, -9 and Apaf-1. CONCLUSIONS: Although each of the DEDs in Casp8NC-DED and MC159 has the potential to stimulate or inhibit apoptosis, the combination of the two-repeated DEDs is necessary for the DED-containing proteins to stimulate or inhibit apoptosis. PMID- 10526241 TI - Vitamin D-binding protein prevents vitamin D deficiency and presents vitamin D for its renal activation. PMID- 10526242 TI - The crucial role of a phosphatase in insulin resistance and obesity. PMID- 10526243 TI - Effect of 1-year treatment with interferon-beta1b on thyroid function and autoimmunity in patients with multiple sclerosis. AB - OBJECTIVE: Interferon-beta (IFN-beta) is a widely used therapy for multiple sclerosis (MS), a demyelinating disease of the central nervous system. This study has evaluated the effect on thyroid function and autoimmunity of a 1-year treatment with IFN-beta1b in patients with MS. PATIENTS: We studied 31 patients (age 34+/-7 years, 21 women) with relapsing-remitting MS during IFN-beta1b treatment of 1 year duration. Systematic thyroid assessment and measurements of serum interleukin-6 (IL-6) levels were performed at baseline and every 3 months during treatment. RESULTS: Sixteen percent of the patients had autoimmune thyroiditis before IFN-beta1b, all positive for anti-peroxidase antibodies. The overall incidence of thyroid dysfunction was 33% over 1 year (10% hyperthyroidism, 23% hypothyroidism). Thyroid autoimmunity developed in 5/26 patients (19%), in one case without dysfunction. In addition to autoantibody positivity at baseline, female gender and the presence of an ultrasound thyroid pattern suggestive of thyroiditis were identified by multiple logistic regression as additional risk predictors for the development of thyroid dysfunction. During IFN-beta1b treatment, serum IL-6 levels rose in a consistent biphasic pattern; there was, however, no difference between patients with or without incident thyroid abnormalities. CONCLUSIONS: We conclude that IFN-beta1b therapy can induce multiple alterations in thyroid function, some of which are unrelated to thyroid autoimmunity. IL-6 measurement is not useful to identify patients prone to develop thyroid abnormalities. Though thyroid dysfunction is generally subclinical and often transient, systematic thyroid assessment should be performed during IFN-beta1b treatment. PMID- 10526244 TI - Standardized grey scale ultrasonography in Graves' disease: correlation to autoimmune activity. AB - OBJECTIVE: Graves' disease leads to thyroid enlargement and to reduction of tissue echogenicity. Our purpose was to correlate grey scale ultrasonography of the thyroid gland with clinical and laboratory findings in patients with Graves' disease. DESIGN: Fifty-three patients with Graves'disease were included in our study, 100 euthyroid volunteers served as control group. Free thyroxine (FT(4)), TSH and TRAb (TSH receptor antibodies) values were measured and correlated with sonographic echogenicity of the thyroid gland. METHODS: All patients and control persons underwent ultrasonographical histogram analyses under standardized conditions. Mean densities of the thyroid tissues were determined in grey scales (GWE). RESULTS: Compared with controls with homogeneous thyroid lobes of normal size (25.6 +/- 2.0GWE, mean +/- S.D.) echogenicity in patients with Graves' disease was significantly lower (21.3 +/- 3. 3GWE, mean +/- S.D., P < 0.0001). Among the patients with Graves' disease significant differences of thyroid echo levels were revealed for patients with suppressed (20.4 +/- 3.1 GWE, mean +/- S.D., n=34) and normalized TSH values (22.5 +/- 3.6GWE, mean +/- S.D., n=19, P < 0.02). Significantly lower echogenicities were also measured in cases of persistent elevated TRAb levels (19.9 +/- 2.9GWE, mean +/- S.D., n=31) in comparison with normal TRAb levels (22.9 +/- 3.5 GWE, mean +/- S.D., n=22, P < 0.0015). No correlation could be verified between echogenicity and either still elevated or already normalized FT(4) values or the thyroid volume. In coincidence of hyperthyroidism and Graves' ophthalmopathy (19.7 +/- 3.5GWE, mean +/- S.D., n=23) significantly lower echogenicity was measured than in the absence of ophthalmological symptoms (22.3 +/- 3.3GWE, mean +/- S.D., n=30, P < 0.016). Patients needing active antithyroid drug treatment revealed significantly lower thyroid echogenicity (20.3 +/- 3.1 GWE, mean +/- S.D., n=40) than patients in remission (23.7 +/- 3.4 GWE, mean +/- S.D., n=13, P < 0.001). Statistical evaluation was carried out using Student's t-test. CONCLUSIONS: Standardized grey scale histogram analysis allows for supplementary judgements of thyroid function and degree of autoimmune activity in Graves' disease. Whether these values help to estimate the risk of recurrence of hyperthyroidism after withdrawal of antithyroid medication should be evaluated in a prospective study. PMID- 10526245 TI - Changes in metabolism of TRH in euthyroid sick syndrome. AB - OBJECTIVE: The aim of this study was to examine the metabolism of a simple dose, intravenously administered TRH bolus of 200 microg, in patients with euthyroid sick syndrome (ESS). PATIENTS AND METHODS: A TRH test was performed on ten ESS patients and ten controls upon admission (d1) and after recovery (d2). Blood samples were collected at 0, 10, 20 and 30min after TRH injection. We analyzed the volume of distribution (V(d)), the plasma clearance rate (PCR), the fractional clearance rate (FCR), the half-life (t(1/2)) and the TSH response to the injection of TRH. RESULTS: All patients had lower tri-iodothyronine (T(3)) levels compared with controls (0.9 +/- 0. 1nmol/l vs 1.9 +/- 0.1 nmol/l; P < 0.0001; mean +/- S.D.; paired t-test). In addition, the V(d) (16.7 +/- 5.9/l vs 30.6 +/- 0.6/l; P < 0.0005) and PCR (2.0 +/- 0.80 l/min vs 3.3 +/- 0.25 l/min; P <0. 0005) were found statistically lowered in patients than in controls, whereas FCR (0.119 +/- 0.01 permin vs 0.110 +/- 0.01 per min; P < 0. 025) was found increased in patients as opposed to controls. The t(1/2) of exogenously administered TRH was increased in ESS compared with controls (7.2 +/- 0.7 min vs 6.3 +/- 0.6 min; P <0.005). TSH response to TRH was found significantly repressed at 10, 20 and 30 min after TRH injection. On d2, these findings had reverted to normal and no changes regarding the kinetics of TRH and the response of TSH could be detected between patients and controls. CONCLUSIONS: The results demonstrate an impairment of TRH metabolism in ESS. The findings may suggest altered enzymatic activity, responsible for TRH degradation in states of acute ESS. These changes might be involved in the pathogenesis of ESS and represent part of an adaptive mechanism to this syndrome. PMID- 10526246 TI - Effect of 6 months of GH treatment on myosin heavy chain composition in GH deficient patients. AB - OBJECTIVE: To investigate the effect of GH on myosin heavy chain (MHC) isoform composition, physical fitness and body composition in GH-deficient (GHD) patients. DESIGN: Twenty-two GHD patients were randomized in a double blind manner and half were treated with recombinant human GH (rhGH) and half were treated with placebo for 6 months. Twelve age-matched controls were also included in the study. METHODS: MHC isoform composition in biopsies obtained from the vastus lateralis muscle was determined using SDS-PAGE. Physical fitness was determined on a bicycle ergometer and body composition was determined using bioelectrical impedance analysis. RESULTS: More MHC IIX (28.9 +/- 4.1% and 10.0 +/- 3.1% in GHD and controls respectively (means +/- S.E.M.)) and less MHC I (36.2 +/- 2.4% and 51.7 +/- 3.9% in GHD and controls respectively (means +/- S.E.M.)) were present in the GHD patients compared with the controls. No significant difference in the amount of MHC IIA was detected. Linear regression was used to determine the relationship between variables. There were no significant relationships between the concentration of insulin-like growth factor I (IGF-I) or the body composition and the MHC composition. Maximal oxygen uptake (VO(2)max) per kg body weight (BW) (litres/min per kg) correlated significantly with the amount of MHC I (r=0.60) and MHC IIX (r=-0.72) but not with the amount of MHC IIA (r=0.35). Treatment of GHD patients with rhGH for 6 months increased the concentration of IGF-I, lean body mass and decreased fat mass but had no effect on MHC composition and physical fitness. CONCLUSIONS: We conclude that a major part of the differences in MHC composition between GHD patients and age matched controls can be explained by variation in physical fitness. The severity of the GHD and the body composition does not seem to be important for the MHC composition. Furthermore, treatment with GH for 6 months does not affect MHC composition in GHD patients. PMID- 10526247 TI - Population based study on serum ionised calcium, serum parathyroid hormone, and blood pressure. The Tromso study. AB - OBJECTIVE: To study associations between serum ionised calcium, serum parathyroid hormone (PTH) and blood pressure. DESIGN: A population based, cross-sectional study was used. METHODS: Blood pressure, body mass index, serum ionised calcium and serum PTH were measured in 460 males and 486 females in the Tromso study in 1994/1995. None were on medication for hypertension. The data were analysed with a multiple linear regression model. RESULTS: When looking at subjects with serum ionised calcium<1.39mmol/l, there was a significant negative association (P<0.01) between serum ionised calcium and PTH. There was no association between blood pressure and serum ionised calcium. In both sexes there was a significant positive association between age and serum PTH (P<0.01). For women, but not for men, there was a significant positive association between serum PTH and systolic and diastolic blood pressure (P<0.01). Within each age group there was a difference in both systolic and diastolic blood pressure of 3-10mmHg between the upper and lower serum PTH halves of the female population. Females with hypertension had significantly higher serum PTH levels than the normotensive females (P<0.01). CONCLUSION: Serum PTH is strongly and positively associated with blood pressure in women. PMID- 10526248 TI - Low levels of serum calcidiol in an African population compared to a North European population. AB - OBJECTIVE: To compare vitamin D status in an African population living at 10 degrees N with a Norwegian population living at 60 degrees N. DESIGN: Serum samples from 30 healthy young Ethiopians and 31 full term pregnant women from Addis Ababa were collected in September, and from 24 healthy Norwegians in March and 23 pregnant women from Oslo in February to June. METHODS: Serum (s) levels of calcidiol and intact parathyroid hormone (iPTH) were measured. RESULTS: The median values for s-calcidiol were significantly lower in Ethiopians compared with Norwegians (young Ethiopians 23.5nmol/l vs young Norwegians 81nmol/l, P<0.001; pregnant Ethiopians 25nmol/l vs pregnant Norwegians 36nmol/l, P<0.05) while those for s-iPTH were significantly higher (young Ethiopians 5.7pmol/l vs young Norwegians 2.4pmol/l, P<0.001; pregnant Ethiopians 4.8pmol/l vs pregnant Norwegians 2.8pmol/l, P<0.02). CONCLUSION: In spite of abundant availability of ultraviolet radiation, the population from Addis Ababa had a high rate of biochemical vitamin D deficiency compared with the Norwegian group. PMID- 10526249 TI - Comparison of finasteride versus flutamide in the treatment of hirsutism. AB - OBJECTIVE: To compare the effectiveness of finasteride and flutamide in the treatment of hirsutism in patients with polycystic ovary syndrome (PCOS) and with idiopathic hirsutism. DESIGN: Randomized study. PATIENTS: One hundred and ten hirsute patients were selected: 64 women with PCOS and 46 with idiopathic hirsutism. METHODS: Patients were assigned randomly to receive 5mg finasteride once daily or 250mg of flutamide twice daily, for 12 consecutive months. Hirsutism was evaluated at 12 months of therapy, with the Ferriman-Gallwey score and with measurement of the terminal hair diameters (microm) taken from four different body areas. Blood samples were taken for assessment of endocrine and hematochemical parameters. Side effects were monitored during the treatment. RESULTS: Both finasteride and flutamide induced a significant decrease in the hirsutism scores and hair diameters at the end of 12 months. Finasteride reduced the Ferriman-Gallwey score by 31.4% in the PCOS cases and by 34.2% in the idiopathic hirsutism cases, and hair diameter by 27.0-34.1% in PCOS and by 29.6 37.9% in idiopathic hirsutism. Flutamide reduced the Ferriman-Gallwey score by 56.7% in PCOS and by 50.9% in idiopathic hirsutism, and hair diameter by 50. 3 60.0% in PCOS and by 47.7-56.5% in idiopathic hirsutism. Flutamide did not induce hormone variations, while finasteride increased testosterone levels by 40% in PCOS and by 60% in idiopathic hirsutism and decreased 3alpha-androstanediol glucuronide (3alpha-diolG) by 66.7% in PCOS and by 69.5% in idiopathic hirsutism. No important side effects or changes in the hematochemical parameters were observed with finasteride, while two patients (3.6%) in the flutamide group expressed abnormal transaminase levels after 6 months of treatment. Dry skin also appeared significantly more with flutamide (67.3%) than with finasteride (23.6%). CONCLUSIONS: Both drugs are effective in the treatment of hirsutism but flutamide is more effective than finasteride. PMID- 10526250 TI - Serum reg protein level is not related to the beta cell destruction/regeneration process during early phases of diabetogenesis in type I diabetes. AB - OBJECTIVE: In type I diabetes mellitus, early markers of beta cell damage are needed in order to detect the infraclinical development of the disease. The reg protein may be a good candidate, as the reg gene has been proposed to play a role in the pancreatic beta cell destruction/regeneration process during diabetogenesis in animal models of autoimmune diabetes. The aim of this study was to test the hypothesis whether serum reg protein level could be representative of either the destructive or regenerative process at the beta cell level during the early phases of type I diabetes in humans. DESIGN AND METHODS: We used a highly specific immunoassay to measure serum reg protein level in controls and in three groups of either diabetes prone or diabetic subjects: recently diagnosed diabetic patients, long-standing diabetic patients and islet cell antibody-positive non diabetic subjects. RESULTS: We found no significant difference between the values observed in these three groups in comparison with control group (90.7+/ 18.1ng/ml, 83.1+/-5.6ng/ml, 98.7+/-24.5ng/ml vs 85.5+/- 5.6ng/ml respectively). Moreover, when the insulin reserve was evaluated at 6 months in the recently diagnosed group, serum reg protein levels were not different between patients with or without residual insulin secretion (at onset: 103+/-42 vs 70.3+/-8. 5ng/ml respectively; at 6 months: 79.7+/-25.8ng/ml vs 81.6+/-15ng/ml respectively). In contrast, trypsin levels were significantly lower in every group of diabetic patients. Results were expressed as means +/- S.E.M. and groups compared by Student's t-test (P<0.05). CONCLUSIONS: We conclude that serum reg protein level cannot be used as a marker for the progression of the diabetogenic process in type I diabetes. PMID- 10526251 TI - A point mutation in the albumin gene in a Chinese patient with familial dysalbuminemic hyperthyroxinemia. AB - Familial dysalbuminemic hyperthyroxinemia (FDH) is an autosomal dominant disorder characterized by euthyroid hyperthyroxinemia. However, FDH has not been reported in Chinese or African patients. Here, we report the first case of FDH in a Chinese patient. A 69-year-old Chinese man was found to have increased serum total T(4) concentrations (198-242nmol/l; normal range 58-148nmol/l) and free T(4) concentrations (>58pmol/l; T(4) analog method, normal range 9-28pmol/l). Serum total T(3) and TSH concentrations were normal. The patient was misdiagnosed as hyperthyroid and was later suspected to have a TSH-producing tumor by the finding of a pituitary microadenoma, which was eventually proven to be a non functional pituitary 'incidentaloma'. Electrophoretic analysis of the patient's serum proteins demonstrated enhanced albumin binding of [(125)I]T(4). Serum free T(4) concentrations were normal (16-19pmol/l, normal range 9-26pmol/l) when a two step method was used. Direct sequencing of the albumin gene showed a guanine to adenosine transition in the second nucleotide of codon 218, resulting in a substitution of histidine (CAC) for the normal arginine (CGC) in one of the two alleles in the patient. The point mutation was further confirmed by HphI digestion of exon 7 of the albumin gene. The patient's son was not affected. Our studies demonstrated that the point mutation of the albumin gene in a Chinese patient with FDH was similar to that found in western white families, but differed from that in a Japanese family in whom a guanine to cytosine transition at the same position was found. PMID- 10526252 TI - Neonatal diabetes mellitus with hypergalactosemia. AB - We report the case of a male, small-for-gestational-age newborn who presented with failure to thrive, severe fluctuation of blood glucose concentrations, and increased serum concentrations of galactose. The infant responded well to a lactose-free diet supplemented with fructose, inulin and corn starch. The metabolic disorder disappeared within 6 months. The transient course, and results of a molecular analysis of the glucose transporter 2 (Glut2) gene seem to rule out Fanconi-Bickel syndrome. PMID- 10526253 TI - Immunohistochemical analysis of Na+/I- symporter distribution in human extra thyroidal tissues. AB - 131Iodine concentration has been described in several extra-thyroidal tissues. Recent evidence has shown that iodine uptake is achieved by the recently cloned human Na(+)/I(-) symporter (hNIS) gene. However, conflicting results were observed in the expression of hNIS transcripts in extra-thyroidal tissues. In order to document further the distribution of hNIS, we investigated its expression using an immunohistochemical method, based on a polyclonal antibody raised against a synthetic peptide. Various extra-thyroidal tissues were examined, particularly from the digestive tract. Our results confirm that the salivary glands and the stomach express hNIS protein significantly. In contrast, hNIS was undetectable in the colon but the rectal mucosa, which has never been examined, exhibited positive immunohistochemical staining. Other digestive tissues, including the oesophagus, small intestine and appendix, were negative. Weak staining was observed in the mammary gland, indicating that hNIS is expressed in this tissue. The pancreas, skin, ovaries, spleen and kidney showed no positive immunostaining. PMID- 10526254 TI - Dopamine agonists both stimulate and inhibit prolactin release in GH4ZR7 cells. AB - Prolactin secretion from the anterior pituitary gland is regulated by multiple factors including prolactin-release inhibiting factors (PIFs) and prolactin releasing factors. PIFs, however, usually dominate to exert a tonic inhibition in the biological system, and the physiological PIF is believed to be dopamine. However, there is accumulating evidence that dopamine can not only inhibit but also stimulate prolactin release. Many investigators believe that this is achieved by activating inhibitory and stimulatory subtypes of dopamine receptors. We tried to demonstrate that one subtype of dopamine receptors is capable of both inhibiting or stimulating prolactin release using GH(4)ZR(7) cells. GH(4)ZR(7) cells express only a short form of dopamine D(2) receptors (D(2s)). Low concentrations of three well-established D(2) receptor agonists (dopamine, apomorphine and bromocriptine) stimulated prolactin release from GH(4)ZR(7) cells while high concentrations inhibited the release. Haloperidol, a D(2) receptor antagonist, blocked the inhibitory action, but was unable to block the dopamine induced stimulatory action. Pretreatment of cells with phenoxybenzamine, a receptor alkylating agent, abolished both the dopamine-induced stimulatory and inhibitory actions. Our results support the thesis that the stimulation of prolactin release induced by dopamine is mediated through dopamine D(2s) receptors since the GH(4)ZR(7) cells have only D(2s) receptors among dopamine receptors. We have concluded that the D(2s) receptor is capable of both stimulating and inhibiting prolactin release, probably via the activation of a G(s) protein by low concentrations and a G(i) protein by high concentrations of dopaminergic agents. PMID- 10526255 TI - Somatostatin and its analog lanreotide inhibit the proliferation of dispersed human non-functioning pituitary adenoma cells in vitro. AB - OBJECTIVE: Somatostatin is a powerful inhibitor of hormone secretion and cell proliferation. Treatment with somatostatin analogs in humans causes a reduction in size and secretory activity of some endocrine tumors, including somatotropic pituitary adenomas. Less studied are the effects of somatostatin agonists on non functioning pituitary adenomas (NFPAs). In this study we characterized the effects of somatostatin and its analog lanreotide on the proliferation of NFPAs in vitro and the intracellular mechanisms involved. DESIGN: Twenty-three NFPA post-surgical specimens were analyzed for somatostatin receptor (SSTR) expression and 12 of them were cultured in vitro to study somatostatin's effects on cell proliferation, assessed by means of [(3)H]thymidine uptake, and the intracellular signaling. RESULTS: One or more SSTR subtypes were expressed in 90% of the adenomas tested. Somatostatin and lanreotide treatment inhibited phorbol myristate acetate (PMA)-induced cell proliferation. Vanadate pretreatment reversed somatostatin and lanreotide inhibition of PMA-induced DNA synthesis suggesting an involvement of tyrosine phosphatase in this effect. In the only adenoma tested, somatostatin directly induced a tyrosine phosphatase activity. Somatostatin and lanreotide caused also a significant inhibition of voltage sensitive calcium channel activity induced by 40mmol/l K(+) depolarization in microfluorimetric analysis. CONCLUSIONS: These data show that somatostatin and lanreotide inhibit human NFPA cell proliferation in vitro, and suggest that activation of tyrosine phosphatases and inhibition of the activity of voltage dependent calcium channels may represent intracellular signals mediating this effect. PMID- 10526256 TI - Mutation analysis of protein kinase A catalytic subunit in thyroid adenomas and pituitary tumours. AB - OBJECTIVE: The adenylyl cyclase system plays an important role in the control of both thyroid follicular and anterior pituitary cell function. Activating mutations affecting important pathway components such as the TSH receptor and Gsalpha occur in the majority of autonomously functioning thyroid nodules. Only a small proportion of other types of thyroid tumours, however, have been reported to harbour these mutations. Activating mutations of Gsalpha have been reported to occur in up to 40% of pituitary somatotroph adenomas. As the majority of cold thyroid nodules and pituitary tumours are unaffected by these mutations, we have investigated the possibility of activating mutations occurring in protein kinase A (PKA), which is another key component of the adenylyl cyclase pathway. DESIGN: Genomic DNA and cDNA were analysed for the presence of PKA Calpha mutations by allele-specific oligonucleotide hybridisation and single strand conformation polymorphism analysis. PATIENTS: A total of 171 tissue samples were investigated. These comprised 66 benign and 24 malignant thyroid neoplasms, 21 somatotroph adenomas, 35 non-functioning pituitary adenomas, 2 corticotroph adenomas, 1 malignant prolactinoma, and 22 normal pituitary tissue samples. RESULTS: No mutations of PKA Calpha were identified using either allele-specific oligonucleotide hybridisation or single strand conformation polymorphism analysis. CONCLUSIONS: It appears that PKA Calpha mutations at the codons investigated do not represent an oncogenetic mechanism in the development of thyroid and pituitary neoplasms. PMID- 10526257 TI - Human lymphocytes express hGH-N gene transcripts of 22kDa, 20kDa and minor forms of GH, but not hGH-V gene. AB - Expression of human growth hormone (hGH) in lymphocytes was examined by reverse transcription polymerase chain reaction (RT-PCR) in five normal subjects. Transcripts of hGH-N gene, but not hGH-V gene, were detected. Sequence analysis revealed four kinds of transcripts: 22kDa GH, 20kDa GH and two other forms of variant GH. The 20kDa GH transcript was generated by alternative splicing within exon 3, resulting in a 45bp deletion. One of the variant GH transcripts was also generated by alternative splicing within exon 3, but at a different site, resulting in a 73bp deletion. Because of a frameshift, this variant GH transcript may encode a 6.6kDa protein (truncated GH) that structurally differs from that of 22kDa GH after residue 31. In the other variant GH mRNA, exons 3 and 4 were completely skipped. The proportions of expression of 22kDa GH, 20kDa GH and the truncated GH were 60.9+/-13.6 (+/-S.D.)%, 32.7+/-14.1% and 6.4+/-1.1% (n=5), respectively, by comparative RT-PCR. We conclude that human lymphocytes, like the pituitary gland, express hGH-N gene transcripts of mainly 22kDa GH, but also 20kDa GH and minor variant forms of GH. PMID- 10526258 TI - Adaptation of pancreatic islet B-cells during the last third of pregnancy: regulation of B-cell function and proliferation by lactogenic hormones in rats. AB - In rodents, placental lactogen (PL)-I is considered to be the first trigger to enhance pancreatic islet B-cell function, and after its secretion is diminished at mid-pregnancy, PL-II takes over this role. However, little information is available on the regulation of islet B-cell function and proliferation by lactogenic hormones during the last third of pregnancy. This was the focus of the present study using rats in which pregnancy was forcibly prolonged. This rat possesses unique characteristics in that PL-I is re-secreted during the prolonged period of pregnancy and the peak concentrations in maternal circulation are comparable with those observed during mid-pregnancy in normal-pregnancy rats. Pregnancy was prolonged by successive administration of pregnant mare's serum gonadotropin (30IU/rat, s.c. on day 12) and human chorionic gonadotropin (10IU/rat, i.v. on day 14). When the insulin secretory responses to 10mmol/l glucose in islets obtained from normal-pregnancy and prolonged-pregnancy rats were tested, each insulin secretory response correlated well with the values of plasma lactogenic activity throughout the period of pregnancy and lactation. Examination of B-cell proliferation in normal-pregnancy rats showed that 5-bromo 2'-deoxyuridine (BrdU) incorporation into dividing B-cells reached a maximum on day 15 and then decreased markedly towards term. No increase in B-cell proliferation was observed on day 19 when plasma lactogenic activity reached the maximum. In prolonged-pregnancy rats, BrdU incorporation also continued to decrease as observed in normal-pregnancy rats after day 15, and then no enhancement in B-cell proliferation was observed even when the plasma lactogenic activity, including re-secreted PL-I, reached maximum. These results suggest that, in the last third of pregnancy, B-cell proliferation is no longer stimulated by lactogenic hormones in contrast to the insulin secretory response which is sustained. PMID- 10526260 TI - Regulation of neuropeptide Y mRNA expression in cultured human pheochromocytoma cells. AB - The expression of the neuropeptide Y (NPY) gene varies considerably in human pheochromocytomas, but the mechanisms for this variation have not been clarified. To investigate the regulation pattern of the NPY gene in human pheochromocytomas, we screened 16 pheochromocytomas and 9 normal adrenal tissues with Northern blots. The expression level of NPY mRNA in normal adrenal medulla was low and relatively constant, while the pheochromocytomas showed a very wide variation in NPY mRNA levels in both malignant and benign tumors. This indicates that NPY gene expression is not correlated with malignancy in pheochromocytomas. In primary cultures of human pheochromocytoma cells, nerve growth factor treatment (causing neuronal differentiation) increased NPY mRNA accumulation 2- to 5-fold (P < 0.05). NPY mRNA levels were also induced by protein kinase modulators (Bu)(2)cAMP and staurosporine in the cultures (P < 0.05). In contrast, treatment with dexamethasone and IGF-II (causing or linked with chromaffin differentiation) reduced NPY mRNA accumulation (P < 0.05). These data show that the regulation pattern of NPY mRNA expression in cultured human pheochromocytoma cells is different from that previously described in rat pheochromocytoma PC12 cells. Regulation of NPY mRNA expression in primary cultures by these differentiating factors suggests that the expression of NPY mRNA in pheochromocytoma tissues may be associated with the neuronal differentiation of the tumor cells affected by multiple factors. PMID- 10526259 TI - Changes in uterine ornithine decarboxylase activity and steroid receptor levels during decidualization in the rat induced by CDRI-85/287. AB - CDRI-85/287 is an anti-estrogen and interferes with decidualization in the rat uterus. In this study, uterine estrogen receptor (ER) and progesterone receptor (PR) levels were determined during the inhibition of decidualization. The effect of 85/287 on uterine ornithine decarboxylate (ODC) activity (a marker enzyme for decidualization) was also studied, using immature ovariectomized rats divided into four different groups: control, 2.5mg/kg 85/287 only; 1 microg estradiol only; and 85/287 + estradiol. Pseudopregnant rats were administered 85/287 (2.5mg/kg p.o.) on day 3 post-coitum. Deciduoma was induced in one of the uterine horns on day 4 and animals were autopsied 18h post-traumatization. Both ERs and PRs showed an increase in traumatized horns compared with non-traumatized. In the 85/287-treated uterus, there was a reduction in cytosolic ERs in both traumatized horns. However, nuclear ER and PR levels increased in both horns under the influence of 85/287. Similarly, in a tamoxifen (0.03mg/kg)-treated group a decline was noticed in cytosolic ER with a mild increase in nuclear PR. Total ER content, expressed per 100 microg DNA, showed a decline in 85/287- or tamoxifen treated rats. However, no significant alterations were observed in total PR levels in non-traumatized horns. In an immature rat model, 85/287 caused a significant (>50%) reduction in estradiol-induced ODC activity. These findings suggest that the decidualization inhibitory activity of 85/287 may be attributed to inhibition of certain timed biochemical events and genomic/non-genomic actions of estrogens in the rat uterus. PMID- 10526261 TI - Depot-specific release of leptin from subcutaneous and omental adipocytes in suspension culture: effect of tumor necrosis factor-alpha and transforming growth factor-beta1. AB - OBJECTIVE: Leptin, the product of the ob gene, is overexpressed in human obesity and increased serum leptin levels are closely correlated with adipose tissue mass, but the regulation of leptin production is not completely understood. The aim of this study was to characterize the role of tumor necrosis factor (TNF) alpha and transforming growth factor (TGF)-beta1 in depot-specific secretion of leptin from cultured human adipocytes. DESIGN AND METHODS: We measured the leptin concentrations in the culture medium of omental and subcutaneous abdominal adipocytes taken from severely obese individuals and kept in suspension culture, and studied the effect of TNF-alpha and TGF-beta1 on leptin release. Leptin protein was measured by radioimmunoassay, leptin mRNA was assessed by reverse transcriptase (RT)-PCR relative to a housekeeping gene. RESULTS AND CONCLUSION: Leptin secretion from subcutaneous fat cells was 2- to 3-fold higher than that from omental fat cells after incubation for 2 and 24h respectively. A 2-h exposure of adipocytes to 1nmol/l TNF-alpha and 400pmol/l TGF-beta1 respectively did not significantly affect leptin secretion. Whereas a 24-h incubation with 1nmol/l TNF-alpha also did not influence leptin secretion from fat cells from both depots, exposure of omental fat cells to 400pmol/l TGF-beta1 for 24h resulted in a significant inhibitory effect (by 33%) on leptin secretion (P<0.05). A 24- and 48-h exposure of in vitro differentiated human adipocytes to TNF-alpha led to a significant decrease in leptin mRNA levels to 70 +/- 8% and 49 +/- 13% of controls respectively. Similarly, TGF-beta1 decreased leptin mRNA expression in newly differentiated human adipocytes to 77 +/- 12% after 24h and to 54 +/- 8% after 48h compared with control cultures. These data provide evidence that long-term exposure of human fat cells to TNF-alpha or TGF-beta1 may suppress leptin expression in human adipose tissue. The inhibitory effect of TGF beta1 appears to be more pronounced in omental as compared with subcutaneous adipocytes. PMID- 10526262 TI - Overexpression of focal adhesion kinase, a protein tyrosine kinase, in ovarian carcinoma. AB - BACKGROUND: Focal adhesion kinase (FAK) is a tyrosine kinase that is important to such key functions such as cell adhesion, motility, and invasion. A MEDLINE search of the years 1980-1998 found no previous reports of FAK expression in human ovarian carcinoma. The authors performed experiments to determine whether FAK expression is elevated in this disease. METHODS: Ten normal human ovarian tissue samples and 26 cancer samples from patients with Stage I-IV ovarian carcinoma were obtained. Two ovarian carcinoma cell lines were also analyzed. FAK expression was determined by Western blot analysis with the V39 anti-human FAK polyclonal antibody. The level of FAK protein expression was determined using densitometric scanning of the 125 kD band on autoradiographs of Western immunoblots. RESULTS: Serous cancers expressed fourfold-increased values of FAK relative to normal ovarian tissue (P < 0.0001), and nonserous adenocarcinomas expressed threefold- to fourfold-increased values of FAK (P < 0. 0006). Ovarian carcinoma cell lines also expressed increased values of FAK. With a cutoff of 40, an elevated FAK level was associated with a sensitivity of 93% and specificity of 100%. There was no significant difference in FAK expression with regard to grade or stage of tumor. CONCLUSIONS: FAK is significantly overexpressed in ovarian carcinoma, implying that FAK may play an important role in ovarian carcinogenesis. FAK expression may be useful as a screening tool to identify newly developed disease or as a tumor marker in confirmed cases of epithelial ovarian carcinoma. FAK may also serve as a potential target for therapeutic disruption of ovarian carcinoma progression. PMID- 10526263 TI - Intra-arterial preoperative cytostatic treatment versus preoperative irradiation: A prospective, randomized study of lingual and sublingual carcinomas. AB - BACKGROUND: For several decades, both preoperative intra-arterial chemotherapy and preoperative irradiation have been accepted treatments for patients with tumors of the head and neck. Unfortunately, arguments have often been put forward in favor of one or other of the two methods, but without the performance of an objective, randomized investigation. To resolve this situation, the authors have carried out a multicenter, randomized prospective study of selected patients with a view to deciding which method affords better results in complex tumor therapy from the aspects of survival and postoperative quality of life. METHODS: One hundred thirty-one patients with operable sublingual or lingual squamous cell carcinoma in stages T2NXM0 to T4MXM0 were randomized into 2 groups: 1 group participated in preoperative chemotherapy with cisplatin and epirubicin (total doses: 200 mg cisplatin, 120 mg epirubicin) via the external carotid artery, whereas the other group received preoperative radiation therapy (46 grays). Following subsequent radical surgery, the patients received regular follow-up for 5 years. RESULTS: By the end of the 5 years, 95 of the 131 patients had conformed to the protocol. Of those 95, 47 had received preoperative chemotherapy and 48 preoperative irradiation. After 5 years, 18 of the 47 patients who received chemotherapy and 15 of the 48 patients who received irradiation were still alive and tumor free. A few more patients had died of recurrence or regional metastasis in the chemotherapy group (23 patients) than in the irradiation group (20 patients). Occurrence of a second carcinoma was 3 times as frequent in the irradiation group (9 patients) as in the chemotherapy group (3 patients). Overall, the survival rates were by-and-large the same for the two groups. Regarding postoperative quality of life, the chemotherapy group presented a more favorable picture. CONCLUSIONS: The long term survival results subsequent to preoperative intra-arterial chemotherapy or preoperative radiotherapy were practically the same. Regarding postoperative quality of life, patients who underwent intra-arterial chemotherapy appeared to be in a slightly more favorable situation. The authors consider it important to stress these findings, as they are not aware of a similar randomized study of patients with tumors of the oral cavity. PMID- 10526264 TI - Development of a new staging system for recurrent oral cavity and oropharyngeal squamous cell carcinoma. AB - BACKGROUND: Approximately 33% of patients with squamous cell carcinoma of the oral cavity and oropharynx develop a recurrence. The management of recurrent tumors can be challenging to both physician and patient, at least in part due to the lack of an accurate and clinically applicable staging system for these patients. The purposes of this study were to examine the survival patterns of patients presenting with recurrent oral cavity and oropharyngeal tumors, to identify key factors affecting prognosis, and to combine these factors to create a new staging system to predict survival and aid in planning therapy. METHODS: The methods included a retrospective chart review of 641 patients with oral cavity and oropharyngeal squamous cell carcinoma who underwent their initial treatment at Washington University between 1980 and 1992. From this population, 249 patients (39%) developed a recurrence. RESULTS: The overall 2-year survival rate was 20% (50 of 249 patients). Six variables affected survival significantly: histologic differentiation, initial (prior to first therapy) TNM stage, initial treatment, time to recurrence, extent of recurrence, and treatment of recurrence. These six variables were entered into a logistic model to determine the individual prognostic significance of each variable. Two variables were found to be statistically significant: initial TNM stage (chi-square test = 7.67; P = 0.0056) and extent of recurrence (chi-square test = 11.75; P = 0.0006). Using the process of conjunctive consolidation, these two variables were combined to create a new staging system for recurrent tumors of the oral cavity and oropharynx. CONCLUSIONS: This staging system provides accurate estimates of prognosis, involves no new technology to implement, demonstrates statistically significant differences in survival by stage, and may aid both the physician and the patient in planning therapy. PMID- 10526265 TI - Prognostic significance of biologic factors in squamous cell carcinoma of the esophagus. AB - BACKGROUND: Esophageal carcinoma is one of the most lethal tumors. Therefore, it is important to identify prognostic factors for patients with this disease. The objective of this study was to clarify the relation between clinicopathologic and biologic factors in esophageal carcinoma and to determine the prognostic significance of different biologic factors. METHODS: DNA ploidy pattern, Ki-67 labeling index (LI), and cyclin D1 and p53 protein expression were examined and detailed pathologic examinations were conducted on tumors from 53 patients (46 males and 7 females with a mean age of 66 years [range, 47-85 years]) with surgically resected esophageal squamous cell carcinoma and the prognostic value of these factors was evaluated. RESULTS: Of the 53 esophagus carcinomas examined, 26 (49%) were classified as DNA diploid. The mean Ki-67 LI was 45 +/- 4. 9% (range, 10.5-86.1%). p53 expression was detected in 38 of the carcinomas (71.7%) and cyclin D1 expression was detected in 35 (66%). Various prognostic factors were examined using the Cox stepwise regression model, four of which were found to correlate with overall survival: tumor size (P = 0.0346), lymph node status (P = 0.0384), Ki-67 LI (P = 0.0161), and p53 expression (P = 0.001). Lower Ki-67 LI and a lower rate of p53 expression were detected in the long term survival group (> 3 years) compared with the short term survival group (P = 0.00045 and P = 0.0023, respectively). CONCLUSIONS: The biologic factors of Ki-67 LI and p53 expression, as well as clinicopathologic factors, may be used as independent prognostic factors for patients with esophageal carcinoma. However, the results of the current study do not support cyclin D1 expression as a prognostic factor. PMID- 10526266 TI - Concurrent chemoradiotherapy for esophageal carcinoma patients with malignant fistulae. AB - BACKGROUND: It remains controversial whether chemotherapy and/or radiotherapy are/is contraindicated for esophageal carcinoma patients with malignant fistulae. In some case reports, closure of fistulae by chemotherapy or radiotherapy has been reported. The current study investigated chemoradiotherapy for these patients using various primary treatments to manage the pulmonary complications. The aim of this study was to evaluate the efficacy and feasibility of chemoradiotherapy for patients with locally advanced esophageal carcinoma with malignant fistulae. METHODS: Patients with endoscopically or radiologically confirmed fistulae were treated with concomitant chemoradiotherapy. Closure of fistulae was assessed by esophagography or endoscopy. Oral food intake also was assessed before and after treatment. RESULTS: Of 202 esophageal carcinoma patients treated at National Cancer Center Hospital East between July 1992 and May 1998, 24 patients (11.9%) developed malignant fistulae. Twelve patients developed fistulae before treatment and the remaining patients developed fistulae during treatment. Closure of the fistulae after chemoradiotherapy was observed in 17 of these patients (70.8%), and 16 of these 17 patients (94.1%) had oral alimentation restored after successful treatment. The median survival time from the diagnosis of the fistula for all patients with fistulae was 198 days; in the patients whose fistulae were present before chemoradiotherapy, the median survival time was 238 days. CONCLUSIONS: These results suggest that the presence of malignant fistulae does not contraindicate chemoradiotherapy. Once the inflammation due to the fistula has been controlled, chemoradiotherapy should be utilized because it may provide the best chance for survival and palliation of severe dysphagia. PMID- 10526267 TI - Depressed adenoma of the duodenum in patients with familial adenomatous polyposis: endoscopic and immunohistochemical features. AB - BACKGROUND: Depressed neoplastic lesions of the colorectum have been specified in patients with familial adenomatous polyposis (FAP). The aim of this study was to characterize endoscopic, histologic, and immunohistochemical features of depressed adenoma of the duodenum in patients with FAP. METHODS: Duodenoscopy was performed on 25 patients with FAP, and the neoplastic nonampullary lesions were classified as polypoid or depressed adenomas. The grade of dysplasia, the proliferative activity determined by Ki-67 labeling index (LI), and the grade of p53 expression were compared between polypoid and depressed neoplasia. RESULTS: Ten subjects had depressed nonampullary adenoma, whereas polypoid adenoma was found in the remaining 15 subjects. Moderate dysplasia was more frequent in depressed adenoma than in polypoid adenoma (70% vs. 27%, P = 0.04). Whereas p53 expression was not different between the two adenoma groups, the LI was significantly higher in depressed adenoma than in polypoid adenoma (59.7 +/- 9.5 vs. 47.5 +/- 10.7, P < 0.01). CONCLUSIONS: Depressed adenoma of the duodenum is a distinctive phenotype of duodenal neoplasm in patients with FAP. The high proliferative activity of depressed adenoma suggests that there may be a need to survey FAP patients with such lesions intensively. PMID- 10526268 TI - Combined role of tumor angiogenesis, bcl-2, and p53 expression in the prognosis of patients with colorectal carcinoma. AB - BACKGROUND: The objective of this study was to evaluate intratumoral neoangiogenesis in Dukes Stage B and Stage C (AJCC/UICC Stage I and III) colorectal adenocarcinoma and its correlation with nuclear p53 oncoprotein accumulation and cytoplasmic bcl-2 expression as well as to assess the prognostic significance of these features in patient outcome. METHODS: Paraffin embedded specimens from 55 patients with Dukes Stage B (AJCC/UICC Stage I) and 51 patients with Dukes Stage C (AJCC/UICC Stage III) colorectal adenocarcinoma who were treated with surgery were assessed. Patients with lymph node involvement (Dukes Stage C [AJCC/UICC Stage III]) also were treated with postoperative pelvic radiotherapy and adjuvant chemotherapy with 5-fluorouracil and leucovorin with or without interferon-alpha. Immunohistochemistry was performed using the anti-CD31 monoclonal antibody (MoAb) for vessel staining, the DO7 MoAb for nuclear p53 expression, and the clone 124 for cytoplasmic/perinuclear bcl-2 expression. Patient follow-up ranged from 4-70 months (median, 28 months). RESULTS: High vascular grade (microvessel score [MS] >/= 40) was observed in 39 of 106 specimens (37%), a medium MS (16-39) was observed in 29 of 106 cases (27%), and a low MS (7-15) was observed in 38 of 106 cases (36%). Positive expression of the bcl-2 protein in > 10% of cells was observed in 33 of 106 cases (31%), whereas p53 nuclear oncoprotein accumulation in > 10% of cells occurred more frequently (44 of 106 cases [42%]). No correlation among p53 expression, bcl-2 expression, and vascular grade was observed. Stroma infiltration by CD31 positive lymphocytes was associated strongly with increased vessel density (P = 0.0001). In univariate analysis Dukes stage was the only significant prognostic parameter (P = 0.02), whereas p53 and vascular grade showed marginal prognostic significance (P = 0.07 and P = 0.09, respectively). In Dukes Stage C (AJCC/UICC Stage III) patients, high vascular grade was the only parameter that predicted a worse overall survival (P = 0.04). Double stratification showed that patients with high vascular grade and positive p53 expression had a poorer survival (P = 0.03). CONCLUSIONS: The results of the current study suggest that p53 mutations, loss of bcl-2 expression, and tumor angiogenesis are events linked to the processes of metastases and local invasion in patients with colorectal carcinoma. Increased vascularization appears to be the most important prognostic factor in patients with Dukes Stage C (AJCC/UICC Stage III) colorectal adenocarcinoma. PMID- 10526269 TI - Telomere shortening and the clinicopathologic characteristics of human colorectal carcinomas. AB - BACKGROUND: It has been reported that shortening of telomeres and strong activation of telomerase occur frequently in colorectal carcinomas. In the current study, the authors examined the correlations between the telomere length of colorectal carcinomas and their clinicopathologic characteristics as well as the activity of telomerase to clarify whether telomere length might represent the biologic behavior of tumors and the mode of tumor development. METHODS: Telomere length was examined by Southern blot analysis in 61 invasive colorectal carcinomas and corresponding normal mucosas. Telomerase activity was assayed by the telomeric repeat amplification protocol with minor modifications. RESULTS: Shortening of the telomere was detected in 38 (62.3%) and elongation in 3 (4. 9%) of the 61 carcinomas. The telomere shortening occurred more frequently in nonulcerating polypoid carcinomas than in ulcerating carcinomas (P = 0.0373) and also occurred more frequently in ascending colon carcinomas than in sigmoid colon or rectal carcinomas (P = 0.0259 and P = 0.0407, respectively). However, no significant correlation was found between the activity of telomerase and the length of telomere. CONCLUSIONS: The results of this study indicate that telomere length may represent the biologic behavior of individual tumors and possibly the mode of development of colorectal carcinomas. PMID- 10526270 TI - High prevalence of transfusion-transmitted virus among patients with non-B, non-C hepatocellular carcinoma. AB - BACKGROUND: Many patients with hepatocellular carcinoma are positive for hepatitis B surface antigen (HBsAg) or antibodies to hepatitis C virus (anti HCV). Recently, transfusion-transmitted virus (TTV) DNA was identified in the serum of patients with non-B, non-C posttransfusion hepatitis. In this study, the prevalence of TTV DNA in the serum of patients with non-B, non-C hepatitis associated hepatocellular carcinoma was evaluated. METHODS: Fifteen patients with hepatocellular carcinoma negative for HBsAg, antibodies to hepatitis B core antigen (anti-HBc), and anti-HCV antibodies were enrolled in this study (non-B, non-C group). Fifteen patients positive for HBsAg and negative for anti-HCV antibody (HBV group) and another group of patients negative for HBsAg but positive for anti-HCV antibody (HCV group) were also enrolled in this study. Data obtained from 27 healthy subjects negative for both HBsAg and anti-HCV antibody and normal levels of serum alanine aminotransferase represented controls. The healthy control group, the non-B, non-C group, and the HCV group were age matched. TTV DNA was detected by heminested polymerase chain reaction in which specific primers were used. RESULTS: TTV DNA was detected in 10 of 15 patients (67%) in the non-B, non-C group. This prevalence rate in the non-B, non-C group was significantly higher than that in the HBV group (3 of 15 patients, 20%) and the control group (9 of 27 patients, 33%), but it was not significantly different from that in the HCV group (7 of 15 patients, 47%). The noncancerous hepatic tissue samples of 10 TTV-DNA positive patients in the non-B, non-C group included 2 with chronic hepatitis and 8 with cirrhosis. CONCLUSIONS: This study showed that TTV DNA is frequently detected in the serum of patients with non-B, non-C hepatocellular carcinoma. This result suggests a potential pathogenetic association between hepatocellular carcinoma and TTV infection. PMID- 10526271 TI - K-ras mutations in duodenal aspirate without secretin stimulation for screening of pancreatic and biliary tract carcinoma. AB - BACKGROUND: K-ras mutations at codon 12 (KRM) have been detected in over 80% of tissues and pure pancreatic juice (PPJ) samples from patients with pancreatic carcinoma (PCa) and are promising genetic tumor markers. Aspirating PPJ not only requires technical skill, but is also exhausting for patients. The authors attempted to evaluate whether the detection of KRM in the duodenal aspirate (DA) obtained immediately after endoscopic retrograde cholangiopancreatography (ERCP), an easier sample-collecting method than collecting PPJ, could be useful for the diagnosis of PCa and biliary tract carcinoma (BTCa). METHODS: DA was collected endoscopically without secretin stimulation immediately after the ERCP procedure from 160 patients: 38 patients with PCa, 38 with chronic pancreatitis (CP), 22 with BTCa, 20 with adenomyomatosis of the gallbladder (AGB), 22 with cholecystolithiasis (CCL), and 20 control subjects. Mutant allele specific amplification (MASA), which is a highly sensitive method for detecting KRM, was performed, with the DNAs extracted from these samples by phenol-chloroform. RESULTS: The incidence of KRM in DA by MASA was 25 (66%) of the 38 PCa cases, 12 (32%) of the 38 CP cases, and 12 (55%) of the 22 BTCa cases. There was no patient with positive KRM in DA among the 20 cases of AGB, 22 of CCL, and 20 control subjects. The sensitivity was 62% and the specificity 88% in this study design. The KRM incidence was found to be relatively high for the patients with PCa and BTCa by MASA, which is a highly sensitive method, although the incidence of KRM in DA from the patients with PCa was not as high as the incidence in their PPJ with secretin stimulation. CONCLUSIONS: MASA showed a relatively high incidence of KRM even in the DA, which was easily obtained from the patients with PCa and BTCa without secretin stimulation immediately after ERCP. These results suggest that the detection of KRM in the DA by MASA is useful for the screening of both PCa and BTCa. PMID- 10526272 TI - Predictive value of Ki-67, p53 protein, and DNA content in the diagnosis of gastric carcinoma. AB - BACKGROUND: The ability to make a precise preoperative diagnosis is a valuable and effective method in improving the prognosis of patients with gastric carcinoma. The authors examined retrospectively whether preoperative histopathologic analysis with p53 protein, Ki-67 labeling index, and DNA ploidy along with preoperative radiographic and endoscopic findings led to a precise preoperative diagnosis of patients with gastric carcinoma. METHODS: Histopathologic analysis of p53 protein, Ki-67 labeling index, and DNA content was performed on formalin fixed, paraffin embedded tissue. Tissue sections from endoscopic and surgically resected specimens were stained immunohistochemically for p53 protein and Ki-67 labeling index, and the cell nuclear DNA content of the surgically resected primary lesion was measured using a microspectrophotometer. These analyses were performed on 16 patients with early gastric carcinoma (EGC) who were diagnosed with advanced gastric carcinoma (AGC) based on the preoperative imaging findings and on 15 patients with AGC who were diagnosed preoperatively with EGC. RESULTS: Overexpression of p53 in the AGC group was significantly more frequent compared with that in the EGC group (P = 0.0386). With regard to the correlation between lymph node metastases and p53 overexpression, there was no apparent relation in either the AGC group (P = 0.648) or the EGC group (P = 0.726). The AGC group had significantly higher Ki-67 labeling indices compared with the EGC group (P = 0.0195). There was complete concordance between endoscopic and surgically resected specimens with regard to the p53 and Ki-67 labeling index findings. DNA ploidy in the primary tumor did not differ between the AGC and EGC groups. The survival rates for the EGC group were significantly superior to those for the AGC group (P = 0.0312). CONCLUSIONS: The findings of the current study suggest that in routine clinical practice, the combination of preoperative imaging findings in addition to Ki-67 labeling indexes, and p53 protein analyses may be useful for the accurate diagnosis of EGC; however, DNA ploidy did not appear to reflect the growth potential of gastric carcinoma. PMID- 10526273 TI - The expression of transforming growth factor-beta1 is significantly correlated with the expression of vascular endothelial growth factor and poor prognosis of patients with advanced gastric carcinoma. AB - BACKGROUND: Transforming growth factors beta (TGFs beta) are involved in a variety of important cellular functions, including cell growth and differentiation, adhesion, migration, extracellular matrix formation, and immune function. Moreover, it has been reported that TGFs beta are correlated with angiogenesis. However, the role of TGF-beta as an angiogenic factor in gastric carcinoma is still unclear. METHODS: TGF-beta1 expression was determined in 101 patients with gastric carcinoma by immunohistochemical procedures, and this expression was compared in the current study with both the expression of vascular endothelial growth factor (VEGF), which is thought to be the most potent angiogenic factor, and microvessel density, to evaluate the effect of TGF-beta1 on the angiogenesis of gastric carcinoma tissues. RESULTS: TGF-beta1 expression was detected in 23 tumors (22.8%). TGF-beta1 expression was more frequent in differentiated than in undifferentiated gastric carcinoma. Furthermore, TGF-beta1 expression was significantly correlated with the depth of invasion and the stage of disease. There was a close correlation between TGF-beta1 expression and VEGF expression. There was no correlation between TGF-beta1 expression and microvessel density, whereas VEGF expression was significantly correlated with microvessel density. With regard to prognosis, the 5-year survival rate was 55.9% for patients with TGF-beta1 positive tumors and 67.0% in patients with TGF-beta1 negative tumors. Accordingly, the prognosis for patients with TGF-beta1 negative tumors was significantly better than that for patients with TGF-beta1 positive tumors. Multivariate analysis indicated that lymph node metastasis, tumor size, and TGF-beta1 expression were independent prognostic factors. CONCLUSIONS: These results suggest that TGF-beta1 might be associated with tumor progression by indirectly stimulating angiogenesis through the up-regulation of VEGF expression in gastric carcinoma. PMID- 10526274 TI - Gemcitabine plus vinorelbine in nonsmall cell lung carcinoma patients age 70 years or older or patients who cannot receive cisplatin. Oncopaz Cooperative Group. AB - BACKGROUND: Although the prevalence of nonsmall cell lung carcinoma (NSCLC) is high among elderly patients, few data are available regarding the efficacy and toxicity of chemotherapy in this group of patients. Recent reports indicate that single agent therapy with vinorelbine (VNB) or gemcitabine (GEM) may obtain a response rate of 20-30% in elderly patients, with acceptable toxicity and improvement in symptoms and quality of life. In the current study the efficacy and toxicity of the combination of GEM and VNB in elderly patients with advanced NSCLC or those with some contraindication to receiving cisplatin were assessed. METHODS: Forty-nine patients with advanced NSCLC were included, 38 of whom were age >/= 70 years and 11 were age < 70 years but who had some contraindication to receiving cisplatin. All patients were evaluable for response and toxicity. Treatment was comprised of VNB, 25 mg/m(2), plus GEM, 1000 mg/m(2), both on Days 1, 8, and 15 every 28 days. Patients received a minimum of three courses unless progressive disease was detected. RESULTS: One hundred sixty-five courses were administered, with a median of 3. 6 courses per patient. The overall response rate was 26% (95% confidence interval, 15-41%). Two patients attained a complete response (4%) and 11 patients (22%) achieved a partial response. Eastern Cooperative Oncology Group performance status improved in 35% of those patients with an initial value > 0, whereas relief of at least 1 symptom without worsening of other symptoms was noted in 27 patients (55%). The median time to progression was 16 weeks and the 1-year survival rate was 33%. Toxicity was mild. Six patients (12%) had World Health Organization Grade 3-4 neutropenia, 2 patients (4%) had Grade 3-4 thrombocytopenia, and 2 patients (4%) had Grade 3 neurotoxicity. Three patients with severe neutropenia (6%) died of sepsis. The median age of those patients developing Grade 3-4 neutropenia was significantly higher than that of the remaining patients (75 years vs. 72 years; P = 0.047). CONCLUSIONS: The combination of GEM and VNB is moderately active and well tolerated except in patients age >/= 75 years. This age group had an increased risk of myelosuppression. Therefore the prophylactic use of granulocyte-colony stimulating factor should be considered with this treatment. New chemotherapy combinations with higher activity and lower toxicity are needed for elderly patients with advanced NSCLC. PMID- 10526275 TI - Scintigraphic prediction of resistance to radiation and chemotherapy in patients with lung carcinoma: technetium 99m-tetrofosmin and thallium-201 dual single photon emission computed tomography study. AB - BACKGROUND: Various prognostic markers for lung carcinoma have been proposed, but to the authors' knowledge none is noninvasive and convenient for clinical use. The current study examined the utility of several radiotracers for the prediction of multidrug resistance (MDR) and radioresistance in patients with lung carcinoma. METHODS: Thirty patients with untreated lung carcinoma underwent a dual isotope single photon emission computed tomography (SPECT) scan at 10 minutes and 120 minutes after the injection of technetium-99m ((99m)Tc) tetrofosmin ((99m)Tc-TF) (370 megabecquerels [MBq]) and thallium-201 ((201)TlCl) (111 MBq). Retention of each tracer was evaluated semiquantitatively. Using radiation and chemotherapy (cisplatin plus etoposide), the patients either were treated sequentially (n = 12) or concurrently (n = 18). The relation between therapeutic response and retention of each tracer was analyzed. The detectability of radioresistance was examined. RESULTS: In patients treated with sequential therapy, the response to radiation was predicted by (99m)Tc-TF retention, whereas (201)Tl retention was found not to be predictive. Regardless of whether the sequential or concurrent protocol was applied, 14 of 18 tumors with high (99m)Tc TF retention (>/= 15%) exhibited a favorable response to chemoradiotherapy whereas all 12 tumors with low (99m)Tc-TF retention (/= 65 years can be screened less frequently than younger women. METHODS: A cost-utility analysis using a computer model that simulates the demography, epidemiology, and natural history of breast carcinoma to estimate expected life-years gained, extra incidence, extra life-years with disease, and costs incurred by different breast carcinoma screening programs in the general population was conducted. RESULTS: The estimated ratio of favorable/unfavorable effects was lower for longer screening intervals compared with shorter screening intervals. The cost-effectiveness ratio was much less favorable in shorter screening intervals. CONCLUSIONS: The results of the current analysis showed that although a longer sojourn time for preclinical breast carcinoma should not necessarily be accompanied by a longer screening interval, a shorter screening interval was not very efficient. PMID- 10526280 TI - Factors associated with axillary lymph node metastasis from breast carcinoma: descriptive and predictive analyses. AB - BACKGROUND: Although axillary lymph node metastasis is one of the most important prognostic determinants of breast carcinoma prognoses, the reasons why tumors vary in their capability to produce for axillary metastases remain unclear. METHODS: The authors used data from the nationwide Patient Care Evaluation (PCE) survey of the American College of Surgeons to evaluate the correlations between patient/tumor characteristics and lymph node status, and to explore the use of these factors, which are all known prior to axillary dissection, in predicting lymph node status. The PCE data set contained 18,025 breast carcinoma cases diagnosed in 1990 after exclusion of women older than 79 years or with fewer than 6 lymph nodes examined. RESULTS: In a multivariate logistic regression model, larger tumor size, young age, African American or Hispanic race, outer half tumor location, poor or moderate differentiation, aneuploidy, and infiltrating ductal histology were independently associated with a higher likelihood of one or more positive lymph nodes. Contrary to expectation, cases negative for estrogen receptor (ER) and progesterone receptor (PR) had a lower risk of positive lymph nodes when adjusted for other factors (odds ratio = 0.82; 95% confidence interval: 0.74-0.91) compared with cases positive for both receptors. This model accurately predicted lymph node status in 2 validation data sets (a 50% random sample of 1990 PCE data and 1992 data from the National Cancer Data Base), but was less accurate in a third, older data set (1983 PCE data). However, the percentage of cases (1990 validation set) with predicted probabilities less than 0.05 or greater than 0.95 were only 4.6% and <0.1%, respectively. CONCLUSIONS: The authors concluded that 1) most variation in axillary lymph node metastatic status can be explained by routinely available data, 2) ER and PR status may be involved in the mechanism of this behavior, and 3) the difficulty of using prediction models to avert axillary dissection should not be underestimated. PMID- 10526281 TI - Long term results of definitive radiotherapy for cervical carcinoma using four applications of high dose rate afterloading. AB - BACKGROUND: In definitive radiotherapy for cervical carcinoma, combined modality treatment using external beam radiotherapy and brachytherapy is standard. Although the optimal number of afterloading applications is controversial, the majority of authors recommend three applications. METHODS: In this study, the authors investigated the use of 4 applications with iridium-192 afterloading with a dose of 7.5 grays (Gy). Standardized radiotherapy doses were adapted to the individual tumor anatomy using a prospective schedule. RESULTS: In the 73 study patients, actuarial and tumor-related 5-year survival rates by T classification were: T1b: 30% (100% cause specific survival); T2a: 55% (76% cause specific survival); T2b: 50% (60% cause specific survival); T3a: 50% (67% cause specific survival); T3b: 39% (50% cause specific survival); and T4a: 40% (40% cause specific survival). Morbidity, graded according to the National Cancer Institute's Common Toxicity Criteria (CTC) were low: CTC Grade 1: 7 patients (7.6%); CTC Grade 2: 7 patients (7.6%); CTC Grade 3: 1 patient (1.4%); and CTC Grade 4: 1 patient (1.4%). In a multivariate analysis, the T classification was the only significant independent prognostic factor for actuarial survival, tumor related (cause specific) survival, local tumor control and freedom of metastatic disease. The number of afterloading applications was an independent prognostic factor for local tumor control. CONCLUSIONS: Based on the long term results of the current study, external beam radiotherapy combined with 4 afterloading high dose rate applications (total of 30 Gy) appears to be clinically feasible and results in satisfactory survival rates and few side effects. PMID- 10526282 TI - The importance of hemoglobin levels during radiotherapy for carcinoma of the cervix. AB - BACKGROUND: It is unclear whether blood transfusion can overcome the negative impact of anemia before or during radiotherapy (RT) in patients with carcinoma of the cervix. The objective of this retrospective study was to examine the impact of anemia and blood transfusion on 605 patients with carcinoma of the cervix treated with radical RT at 7 centers across Canada in 1989, 1990, and 1992. METHODS: The data collected included hemoglobin (Hgb) levels from the time of diagnosis to the end of therapy; blood transfusions administered; and identifiable patient-, tumor-, and treatment-related factors. Survival, disease free survival, and pelvic control analyses were evaluated by univariate and multivariate analysis. RESULTS: The median follow-up was 41 months (range, 0-92 months). Presenting Hgb level, average weekly nadir Hgb (AWNH) during RT, and blood transfusion were correlated significantly with local control, disease free survival, and overall survival on univariate analysis. However, the AWNH remained significant on multivariate analysis, whereas Hgb at presentation and blood transfusion did not. The 5-year survival was 74% for patients with an AWNH >/= 120 g/L, 52% for patients with AWNH levels 110-119 g/L inclusive, and 45% for patients with AWNH levels < 110 g/L (P < 0.0001). At each Hgb level, patients who were transfused and maintained a specific Hgb level had a survival rate that was not significantly different from patients who were at that level spontaneously. There was a significant reduction in both pelvic and distant recurrence (P < 0.0001 and P < 0.0006, respectively) in patients whose AWNH level during RT was >/= 120 g/L compared with < 120 g/L. A reduction in the rate of distant recurrence was observed in patients with and without pelvic recurrence. CONCLUSIONS: AWNH is highly predictive of outcome for patients treated with RT for carcinoma of the cervix. Blood transfusion appears to overcome the negative prognostic effects of low presenting Hgb levels and AWNH levels. PMID- 10526283 TI - Reversed CD4/CD8 ratios of tumor-infiltrating lymphocytes are correlated with the progression of human cervical carcinoma. AB - BACKGROUND: To investigate the clinical significance of tumor-infiltrating lymphocytes (TILs) within the tumor milieu of human cervical carcinoma, the authors quantitatively measured and compared the subpopulations of lymphocytes infiltrating the neoplastic cervix. METHODS: A total of 30 patients with Stage Ia IIa cervical carcinoma were enrolled. TILs were isolated from tissue specimens by means of a mechanical dispersal technique, and the immunocyte subsets were quantified with dual-color flow cytometry. Bulky tumor was defined as tumor size >4 cm in greatest dimension according to the 1995 staging of the International Federation of Gynecology and Obstetrics. RESULTS: The CD4/CD8 ratios of TILs were reversed in both cervical squamous cell carcinoma (n = 20) and cervical adenocarcinoma (n = 10). The proportion of CD4(+) T cells was significantly lower in tumors from patients with lymph node metastasis (n = 8) than in those from patients without lymph node metastasis (n = 22) (24.5 vs. 32.7, P = 0.001), as was the reversed CD4/CD8 ratio (0.50 vs. 0.81, P = 0.001). The proportion of CD4(+) T cells was much lower in bulky tumors (n = 5) than in nonbulky tumors (n = 25) (21.4 vs. 32.5, P < 0.001), reflecting in a more strongly reversed CD4/CD8 ratio (0.41 vs. 0.81, P = 0.001). CONCLUSIONS: Decreased proportions of tumor infiltrating CD4+ T cells with reversed CD4/CD8 ratios are highly correlated with rapid tumor growth and lymph node metastasis in cervical carcinoma. The regional immune escape is of prognostic importance with regard to cancer progression. PMID- 10526284 TI - Ovarian dysplasia in prophylactic oophorectomy specimens: cytogenetic and morphometric correlations. AB - BACKGROUND: Ovarian dysplasia, a potential precursor to ovarian carcinoma, has been described in ovarian tissue obtained by prophylactic oophorectomy and also adjacent to ovarian carcinoma. Women with a family history of ovarian carcinoma, especially those of Jewish Ashkenazi descent, often test positive for BRCA mutant genes. Prophylactically removed ovaries, generally described as normal on macroscopic examination, can exhibit a "preneoplastic phenotype" and unsuspected neoplasm. METHODS: Histologic slides of ovarian tissue from 54 Ashkenazi Jewish women were reviewed. All had a family history of ovarian carcinoma and all were tested for BRCA mutations. Forty-four women tested positive. Thirty-one women underwent prophylactic oophorectomy and 23 underwent oophorectomy for ovarian carcinoma. Normal, dysplastic, and ovarian carcinoma epithelial cells were analyzed morphometrically combining nuclear area measurements with chromatin texture assessment using a novel method based on the computation of autocorrelation coefficients and a derived parameter (Beta). Discriminant analysis between classificatory algorithms was used to obtain results. RESULTS: Ovarian dysplasia was identified in 77.6% of the prophylactic oophorectomy specimens. An unsuspected ovarian carcinoma was diagnosed in one prophylactic oophorectomy specimen. Of 10 women who underwent prophylactic oophorectomy and were negative for BRCA mutations, three had ovarian dysplasia. The average nuclear measurements of the dysplastic cells were similar to those published previously. The new autocorrelation-based method evaluating nuclear texture, as revealed by tridimensional surface plots, demonstrated high discriminatory potential. Discriminant analysis based on nuclear area and nuclear texture information resulted in the correct classification of nearly all the cases in the three diagnostic categories. CONCLUSIONS: Ovaries removed by prophylactic oophorectomy examined in their entirety often reveal ovarian dysplasia and occasionally ovarian carcinoma. The new morphometric method used was highly discriminatory in the evaluation of nuclear texture. Ovarian dysplasia in women with risk factors for ovarian carcinoma is significant in early ovarian carcinogenesis. PMID- 10526285 TI - Defining biochemical cure for prostate carcinoma patients treated with external beam radiation therapy. AB - BACKGROUND: The authors retrospectively reviewed their institution's long term experience with conventional external beam radiation therapy (RT) for localized prostate carcinoma to identify criteria associated with long term biochemical cure. METHODS: Between January 1987 and December 1994, 871 patients were treated with external beam RT alone for clinically localized prostate carcinoma at William Beaumont Hospital, Royal Oak, Michigan. All patients received only external beam RT to a median total dose of 66.6 grays (Gy) (range, 59.4-70.4 Gy). No patient received hormonal therapy unless treatment failure was documented. The median follow-up was 5.0 years (range, 0. 2-11.8 years). Biochemical failure was defined according to the American Society for Therapeutic Radiology and Oncology Consensus Panel definition. RESULTS: In the entire study group, 380 patients experienced biochemical failure at a median interval of 1.5 years after the completion of RT. The 5-year and 7-year actuarial rates of biochemical control were 50% and 48%, respectively. On multivariate analysis, a higher pretreatment prostate specific antigen (PSA) level, higher Gleason score, higher clinical T classification, higher nadir level, and shorter time interval to nadir all were associated significantly with biochemical failure (P < 0.001). The median intervals to biochemical failure for patients with pretreatment PSA levels /= 20.0 ng/mL were 2.2 years, 1.5 years, and 1.2 years, respectively (P < 0. 001). The median intervals to biochemical failure for patients with Gleason scores of 2-4, 5-7, and 8-10 were 1.8 years, 1.5 years, and 1.1 years, respectively (P < 0.001). Only 6 patients failed beyond 5 years after treatment even though 136 patients were at risk for failure beyond this point. When restricting analysis to 643 patients (74%) with >/= 3 years of PSA follow up, the median nadir level for biochemically controlled patients was 0.6 ng/mL and occurred at a median interval of 1.9 years after RT versus a median nadir level of 1.3 ng/mL (P = 0.002) occurring at a median interval of 1.0 years (P < 0.001) in those patients who experienced biochemical failure. Patients were divided into subgroups based on their PSA nadir level and time to nadir. The 5 year actuarial biochemical control rates for patients with nadir values of /= 4.0 ng/mL were 78%, 60%, 50%, 20%, and 9%, respectively (P < 0.001). The 5-year actuarial biochemical control rates for patients who reached their nadir at < 1.0 years, 1.0-1.9 years, 2.0-2.9 years, and >/= 3.0 years were 30%, 52%, 64%, and 92%, respectively (P < 0.001). All 52 patients who achieved a nadir of /= 2.0 years to reach this nadir had biochemically controlled disease. CONCLUSIONS: These results suggest that a patient has a high likelihood of biochemical cure after treatment for prostate carcinoma with conventional doses of external beam RT if he has not demonstrated biochemical failure within 5 years of treatment. Patients with lower pretreatment PSA levels and lower Gleason scores may require longer follow-up than those with less favorable characteristics to achieve the same certainty of cure. Patients who achieve a PSA nadir /= 2.0 years to reach this nadir have the highest probability of cure. PMID- 10526286 TI - Second primary tumors after prostate carcinoma. AB - BACKGROUND: Several large datasets have shown a reduced risk of all neoplasms after a diagnosis of prostate carcinoma but an increased incidence rate of urologic carcinoma has been suggested. METHODS: Data collected by the Cancer Registries of the Swiss Cantons of Vaud and Neuchatel (approximately 760,000 inhabitants) were used to estimate the incidence rate of a second primary tumor after a diagnosis of prostate carcinoma. A total of 4503 cases registered between 1974 and 1994 were followed until the end of 1996 (17,065 person-years). RESULTS: A total of 380 second primary neoplasms were observed versus 534.1 expected primary neoplasms (standardized incidence ratio [SIR] = 0.7; 95% confidence interval, 0.6-0.8). SIRs were significantly below unity for lung carcinoma (SIR = 0.7) and other major tobacco-related neoplasms, including those of the mouth or pharynx (SIR = 0.5), esophagus (SIR = 0.4), pancreas (SIR = 0.5), and larynx (SIR = 0.8). There was no excess rate of subsequent urologic carcinoma (SIR = 1.0) or colorectal carcinoma (SIR = 0.9). The reduced SIRs for lung carcinoma were stronger in elderly men (age >/= 75 years) and in patients with a shorter period since diagnosis (< 5 years). CONCLUSIONS: The incidence of all neoplasms was reduced significantly in men diagnosed with prostate carcinoma. Selection of the population, under-registration of second primary tumors, and reduced surveillance in elderly men with prostate carcinoma may, at least in part, explain this reduction in risk. No excess risk was observed for the complex of urologic neoplasms nor for tobacco-related neoplasms. This finding would not support an association between cigarette smoking and prostate carcinoma. PMID- 10526287 TI - Intratesticular leiomyosarcoma in a young man after high dose doping with Oral Turinabol: a case report. AB - BACKGROUND: Androgenic anabolic steroids have been suspected of activity as carcinogens in the development of carcinoma and angiosarcoma of the liver and adenocarcinoma of the prostate. Although the proliferation of smooth muscle cells is stimulated by sexual steroids, to the authors' knowledge a possible relation between androgenic anabolic steroids and the development of leiomyosarcoma has not previously been reported in humans. METHODS: A 32-year-old man underwent right radical orchiectomy for a tumor of the upper pole of the right testicle. Routine histopathologic examination and immunohistochemical staining were performed. RESULTS: The tumor was identified as an intratesticular leiomyosarcoma based on its typical growth pattern and the characteristic immunohistochemical staining profile. The patient reported a 5-year history of systematic use of high dose Oral-Turinabol (4-chloro-1-dehydro-17alpha-methylteststerone) that began at age 18 years and stopped approximately 9 years before presentation. CONCLUSIONS: The rarity of intratesticular leiomyosarcoma, the experimental induction of similar tumors in animals by androgens and estrogens, and the unusually young age at presentation of the patient in the current study support the hypothesis that high dose doping with androgenic anabolic steroids could have played a cocarcinogenic role in the development of the tumor in this case. PMID- 10526288 TI - Morphologic evidence that analgesic-induced kidney pathology contributes to the progression of tumors of the renal pelvis. AB - BACKGROUND: Whether phenacetin-containing analgesics cause renal pelvic tumors by virtue of the weak mutagenicity of phenacetin, or indirectly through local effects of analgesic-induced renal papillary scarring, is debated. Because phenacetin consumption ceased in New South Wales, Australia in 1975, cases of renal pelvic carcinoma seen 14-15 years later (many of which were associated with long-standing analgesic-induced renal papillary pathology) provided an opportunity to examine the temporal relation between phenacetin exposure and those histologic characteristics of the tumors and adjacent renal tissue that may implicate analgesics in their etiology. METHODS: The authors conducted a "blinded" histopathologic review of tumors of the renal pelvis and adjacent noncancerous renal tissue from 100 cases for which epidemiologic data regarding risk factor exposure (specifically phenacetin-containing analgesics, tobacco, infection, and kidney stones) had been obtained in a population-based case control study from New South Wales in 1989 and 1990. RESULTS: A history of consumption of phenacetin-containing analgesics was associated strongly with the presence and severity of diffuse renal papillary scarring, and less strongly with papillary calcification. The histologic grade of the renal pelvic tumors tended to rise significantly with consumption of phenacetin-containing analgesics in a dose-dependent fashion and with the degree of papillary scarring, but was not related to smoking. In multivariate analysis it was the degree of papillary scarring (to a greater extent than the amount of phenacetin consumption) that was associated significantly and strongly with a higher histologic grade. Only diffuse papillary calcification was associated significantly with squamous change in the renal pelvic tumors. CONCLUSIONS: Based on the results of the current study, the authors conclude that 1) in phenacetin-related tumors of the renal pelvis, the presence and severity of analgesic-induced renal papillary scarring correlates with tumor progression and 2) papillary calcification is a risk factor for squamous change in renal pelvic urothelioma. PMID- 10526289 TI - Predicting metastasis of pheochromocytomas using DNA flow cytometry and immunohistochemical markers of cell proliferation: A positive correlation between MIB-1 staining and malignant tumor behavior. AB - BACKGROUND: In the absence of metastases, there are no reliable microscopic features that distinguish malignant from benign pheochromocytomas. Because a common feature of malignancy is the loss of cell cycle regulation and normal growth arrest, the authors hypothesized that analysis of the cell cycle could be used to aid in the diagnosis of malignant pheochromocytoma. METHODS: Cell cycle analysis of archival samples of 51 pheochromocytomas (40 sporadic, 11 familial) from 45 patients, including 6 malignant and 45 benign tumors, was conducted. Flow cytometry data and immunohistochemistry for markers of cell proliferation (proliferating cell nuclear antigen [PCNA] and MIB-1 [Ki-67]) were correlated with the authors' clinical data base records, with a mean follow-up of 66 months. RESULTS: No correlation of DNA ploidy, S-phase fraction by flow cytometry, or PCNA with malignancy was observed. Staining for the MIB-1 nuclear proliferation marker was positive in 3 of 6 (50%) of the malignant pheochromocytomas and negative in all 45 benign tumors (P< 0.01). CONCLUSIONS: Contrary to some previous reports, a diploid DNA pattern does not necessarily predict benign behavior of pheochromocytoma. In this study, cell cycle analysis and, in particular, assessment of the MIB-1 nuclear proliferation marker was useful in the histologic evaluation of pheochromocytoma, as MIB-1 was expressed only in malignant tumors. PMID- 10526290 TI - Sequential chemotherapy (etoposide, vinblastine, and doxorubicin) and subtotal lymph node radiation for patients with localized Hodgkin disease and unfavorable prognostic features: A phase II Cancer and Leukemia Group B Study (9051). AB - BACKGROUND: The aim of this study was to evaluate a regimen of sequential chemotherapy and radiotherapy for patients with Hodgkin disease. METHODS: The Cancer and Leukemia Group B conducted a Phase II study of three cycles of etoposide, vinblastine, and doxorubicin (EVA) chemotherapy followed by subtotal lymph node radiation for patients with localized Hodgkin disease and unfavorable prognostic features. Fifty-nine patients were enrolled in the study. Fifty-three patients met all study eligibility criteria; 48 of them (91%) had mediastinal disease and 29 (55%) had bulky mediastinal disease. RESULTS: A complete response (CR) occurred in 35 of the patients (66%). Of all patients who had CR, 26% had the CR after the chemotherapy and before the radiation, and 74% after the chemotherapy and radiation. Twenty percent of the patients who had CR experienced disease progression; in these patients, the progression was outside the radiotherapy field in the lung and involved widespread disease. CONCLUSIONS: EVA offers a nonbleomycin-containing alternative for patients in whom preexisting pulmonary disease may be exacerbated by bleomycin and radiation therapy. EVA, as given in this study (in three cycles), was insufficient chemotherapy for patients who had disease in areas outside the radiation fields (occult disease). PMID- 10526291 TI - Propylthiouracil-induced hypothyroidism reduces xenograft tumor growth in athymic nude mice. AB - BACKGROUND: Thyroid hormones are endocrine modulators of several vital processes that are crucial to tumor growth and differentiation. Several anecdotal reports in the literature suggest that some histologic types of carcinoma may remain in a dormant state for prolonged periods of time in patients with hypothyroidism, with eventual progression of the disease once the decreased thyroid function is identified and corrected. METHODS: Oral propylthiouracil (PTU) was used to induce hypothyroidism in athymic nude mice that were subsequently inoculated with lung adenocarcinoma and prostate adenocarcinoma cells. Mice were also treated with a combination of PTU and thyroxine, which resulted in hyperthyroid levels of T(4). RESULTS: Subcutaneous lung and prostate xenografts grew significantly more slowly in hypothyroid mice treated with PTU than in euthyroid or hyperthyroid mice, regardless of treatment with PTU. Tumors grew well in groups of mice that were changed from a hypothyroid state to a euthyroid state by withdrawal of oral PTU. Administration of PTU 3 weeks after tumor inoculation also caused the tumor growth to slow significantly compared with tumors in mice that did not receive PTU. Mice that received PTU and thyroxine had tumors that grew as well as the tumors in euthyroid control animals. CONCLUSIONS: Our study indicates that human lung and prostate tumors do not grow well in hypothyroid nude mice, and that rendering these animals euthyroid has a significant impact on the growth rate of these tumors. Furthermore, in vitro and in vivo data indicated that this was not a result of an interaction of the tumor cells with PTU, but rather a result of the hypothyroid state. PMID- 10526292 TI - Metastases detected at the time of diagnosis of primary pediatric extremity osteosarcoma at diagnosis: imaging features. AB - BACKGROUND: The authors performed a retrospective study to estimate the incidence rate of metastatic disease at the time of diagnosis of extremity osteosarcoma (OS), to characterize its pattern of presentation, and to identify factors predictive of survival within a cohort of patients with pulmonary metastatic disease at diagnosis. METHODS: From the institutional solid tumor database, the authors identified all patients diagnosed with extremity OS since CT became available at the study institution (1977). The authors recorded patient demographics, the site of primary disease, the histologic subtype of OS, and the presence of metastases at diagnosis. In those patients with pulmonary metastases at diagnosis, the presence of calcifications, the primary tumor volume, the number of pulmonary lobes with disease, and the number of pulmonary nodules were recorded. RESULTS: Of an evaluable population of 215 patients, 32 (15%) had bone or pulmonary metastases at diagnosis, of whom original imaging from 28 patients was available for review. Osteoblastic histology correlated with lung metastases at diagnosis (P = 0.049). One of the 32 patients had a solitary bone metastasis without lung metastases. Four of 28 patients (14%) with original imaging available had calcifications within the pulmonary nodules. Both the number of nodules and the number of lobes involved were found to be significant predictors of survival (P = 0.0009 and P = 0. 04, respectively); multiple nodules were bilateral in 61% of patients. CONCLUSIONS: The rate of incidence of computed tomography detected pulmonary metastases was found to be 14% (31 of 215 patients) at diagnosis and 0.5% (1 of 215 patients) for bone metastases in patients with primary extremity OS. Pulmonary metastases usually are multiple and bilateral and infrequently calcify. The number of nodules and lobes involved are predictors of patient survival. PMID- 10526293 TI - Symptom reporting in cancer patients II: relations to social desirability, negative affect, and self-reported health behaviors. AB - BACKGROUND: Patients' appraisal of somatic symptoms is correlated with their negative affect. The authors have investigated whether social desirability is associated with patients' symptom and health behavior reporting. METHODS: One hundred fourteen surgical cancer patients who participated in either an outpatient or an inpatient follow-up care program filled out the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire C30, the New Social Desirability Scale, and a health behavior checklist. RESULTS: Patients' reports of somatic symptoms were correlated inversely with social desirability (r = -0.50) and positively with negative affect (r = 0.72). When objective health and demographic variables (e.g., prognosis, adjuvant therapy prior to follow-up, and gender) were entered first in hierarchical multiple regression analyses, social desirability and negative affect accounted for an additional 16% and 36% of the symptom variance, respectively. Similar results were found when global quality of life was the dependent variable. Self-reported health behaviors were explained only through the set of health and demographic variables (14%), and social desirability and negative affect did not account for additional variance. On the average, patients reported that they had a median of 4.7 (out of a list of 21) self-initiated health behaviors, and 11% of the patients admitted to having used unproven therapies. CONCLUSIONS: Symptom reports do not give a pure picture of patients' health status, but they are strongly correlated with social desirability and negative affect. Detection of such psychologic variables is essential to understanding the dynamics of quality of life. In applied settings, quality-of-life measures should be used together with conventional criteria. As practical experience and scientific understanding grow, the relative positioning of these patient-oriented versus clinic-oriented endpoints will become clear. PMID- 10526294 TI - Uptake of radiolabeled somatostatin analog is detectable in patients with metastatic foci of sarcoma. AB - BACKGROUND: Somatostatin receptors are present on many types of epithelial tumors, and ligands targeting these receptors are used to treat patients with neuroendocrine malignancies. Preclinical studies have demonstrated the presence of somatostatin receptors on a variety of mesenchymal tumors by in vitro receptor autoradiography. The use of radiolabeled somatostatin analogs to assess the presence of somatostatin receptors in vivo has been established, but use of this technique to evaluate human sarcomas has not been reported previously. METHODS: Seventeen patients (13 females and 4 males) with metastatic sarcoma underwent imaging via somatostatin-receptor scintigraphy. Scans were performed using indium -111 pentetreotide. Planar studies and single photon emission computed tomography imaging were performed at 4 and 24 hours, and results of scintigraphy were correlated with computed tomography findings. RESULTS: Twelve of 17 scans showed increased uptake in regions of known metastatic disease. There was no apparent correlation with scan positivity and patient age, histology, site of disease, or duration of diagnosis. CONCLUSIONS: Seventy-one percent of patients with advanced soft-tissue sarcomas had positive scintigraphy scans demonstrating tumor expression of somatostatin receptors subtype 2 in vivo. Imaging with indium-111 pentetreotide could be studied as an adjunct to conventional imaging modalities for assessment of sarcoma patients. Further research is needed to determine the prognostic implications of somatostatin receptor subtype 2 positivity, including larger studies to evaluate any potential correlation with metastatic behavior and other clinical outcomes. PMID- 10526311 TI - Effects of testosterone on the development of a sexually dimorphic neuromuscular system in ciliary neurotrophic factor receptor knockout mice. AB - Motoneurons in the spinal nucleus of the bulbocavernosus (SNB) innervate the perineal muscles, bulbocavernosus (BC), and levator ani (LA). Testosterone regulates the survival of SNB motoneurons and BC/LA muscles during perinatal life. Previous findings suggest that effects of testosterone on this system may be mediated by trophic factors-in particular, by a factor acting through the ciliary neurotrophic factor alpha-receptor (CNTFRalpha). To test the role of CNTFRalpha in the response of the developing SNB system to testosterone, CNTFRalpha +/+ and -/- mice were treated with testosterone propionate (TP) or oil during late embryonic development. BC/LA muscle size and SNB motoneuron number were evaluated on the day of birth. Large sex differences in BC and LA muscle size were present in newborn mice of both genotypes, but muscle volumes were reduced in CNTFRalpha -/- animals relative to same-sex, wild-type controls. Prenatal testosterone treatment completely eliminated the sex difference in BC/LA muscle size in wild-type animals, and eliminated the effect of the CNTFRalpha gene deletion on muscle size in males. However, the effect of TP treatment on BC and LA muscle sizes was blunted in CNTFRalpha -/- females. SNB motoneuron number was sexually dimorphic in oil-treated, wild-type mice. In contrast, there was no sex difference in SNB motoneuron number in oil-treated, CNTFRalpha knockout mice. Prenatal treatment with testosterone did not increase SNB motoneuron number in CNTFRalpha -/- mice, but also did not significantly increase SNB motoneuron number in newborn wild-type animals. These findings confirm the absence of a sex difference in SNB motoneuron number in CNTFRalpha -/- mice. Moreover, the CNTFRalpha gene deletion influences perineal muscle development and the response of the perineal muscles to testosterone. Prenatal TP treatment of CNTFRalpha -/- males overcomes the effects of the gene deletion on the BC and LA muscles without a concomitant effect on SNB motoneuron number. PMID- 10526312 TI - Role of cdk5 and tau phosphorylation in heterotrimeric G protein-mediated retinal growth cone collapse. AB - During axonal growth, repulsive guidance cues cause growth cone collapse and retraction. In the chick embryo, membranes from the posterior part of the optic tectum containing ephrins are original collapsing factors for axons growing from the temporal retina. We investigated signal transduction pathways in retinal axons underlying this membrane-evoked collapse. Perturbation experiments using pertussis toxin (PTX) showed that membrane-induced collapse is mediated via G(o/i) proteins, as is the case for semaphorin/collapsin-1-induced collapse. Studies with Indo-1 revealed that growth cone collapse by direct activation of G(o/i) proteins with mastoparan did not cause elevation of the intracellular Ca(2+) level, and thus this signal transduction pathway is Ca(2+) independent. Application of the protein phosphatase inhibitor okadaic acid alone induced growth cone collapse in retinal culture, suggesting signals involving protein dephosphorylation. In addition, pretreatment of retinal axons with olomoucine, a specific inhibitor of cdk5 (tau kinase II), prevented mastoparan-evoked collapse. Olomoucine also blocks caudal tectal membrane-mediated collapse. These results suggest that rearrangement of the cytoskeleton is mediated by tau phosphorylation. Immunostaining visualized complementary distributions of tau phospho- and dephosphoisoforms within the growth cone, which also supports the involvement of tau. Taking these findings together, we conclude that cdk5 and tau phosphorylation probably lie downstream of growth cone collapse signaling mediated by PTX-sensitive G proteins. PMID- 10526313 TI - Characterization of ATP release from cultures enriched in cholinergic amacrine like neurons. AB - Adenosine triphosphate (ATP) has been proposed to play a role as a neurotransmitter in the retina, but not much attention has been given to the regulation of ATP release from retinal neurons. In this work, we investigated the release of ATP from cultures enriched in amacrine-like neurons. Depolarization of the cells with KCl, or activation of alpha-amino-3-hydroxy- 5-methyl-4-isoxazole propionate (AMPA) receptors, evoked the release of ATP, as determined by the luciferin/luciferase luminescent method. The ATP release was found to be largely Ca(2+) dependent and sensitive to the botulinum neurotoxin A, which indicates that the ATP released by cultured retinal neurons originated from an exocytotic pool. Nitrendipine and omega-Agatoxin IVA, but not by omega-Conotoxin GVIA, partially blocked the release of ATP, indicating that in these cells, the Ca(2+) influx necessary to trigger the release of ATP occurs in part through the L- and the P/Q types of voltage-sensitive Ca(2+) channels (VSCC), but not through N-type VSCC. The release of ATP increased in the presence of adenosine deaminase, or in the presence of 1,3-dipropyl-8-cyclopentylxanthine (DPCPX), an adenosine A(1) receptor antagonist, showing that the release is tonically inhibited by the adenosine A(1) receptors. To our knowledge, this is the first report showing the release of endogenous ATP from a retinal preparation. PMID- 10526314 TI - Evidence that POU factor Brn-3B regulates expression of Pax-6 in neuroretina cells. AB - The Pax-6 gene encodes a transcriptional master regulator involved in the development of the eye. The quail Pax-6 gene is expressed in the neuroretina from two promoters, P0 and P1, and is regulated by an intragenic neuroretina-specific enhancer (EP enhancer). The activity of this enhancer is restricted to the P0 promoter, which is activated at the onset of neuronal differentiation. In this article, we show that the POU domain transcription factor Brn-3b, which is expressed in various regions of the brain including retina and sensory neurons, is one of the factors interacting with the EP enhancer. Brn-3b strongly activates the EP enhancer in neuroretina cells but not in other cell types. Interestingly, this activation appears to be specific for Brn-3b, as the closely related POU factors Brn-3a and Brn-3c do not show activation of the EP enhancer. Our results identify the Pax-6 gene as a new potential downstream effector of the POU transcription factor Brn-3b. PMID- 10526315 TI - Regulation of cyclic GMP elevation in the developing antennal lobe of the Sphinx moth, Manduca sexta. AB - In the moth, Manduca sexta, 3',5'-guanosine monophosphate (cGMP) is transiently elevated during adult development in about 100 neurons of the antennal lobe. We demonstrate that nearly all of these neurons are local interneurons of the lateral cluster I, that their capacity to show a strong cGMP response during development is regulated by the steroid hormone 20-hydroxyecdysone, and that in a subpopulation of these neurons cGMP elevation seems to be controlled directly by the gaseous messenger molecule nitric oxide (NO). Treatment with the acetylcholine esterase inhibitor eserine, antennal nerve transection, and electrical stimulation of the antennae suggest that NO/cGMP signaling during development is an activity-dependent process. Besides input from the antennae, input from the central brain and the ventral ganglia is involved in upregulating cGMP in the antennal-lobe neurons. Possible sources are centrifugal aminergic neurons, since application of serotonin and histamine enhances the GMP signal in local interneurons. Comparing the time course of cGMP elevation with events occurring during development leads us to the hypothesis that the NO/cGMP signaling pathway might be involved in synapse formation of a subset of antennal lobe neurons. PMID- 10526316 TI - Parvalbumin immunoreactivity is enhanced by brain-derived neurotrophic factor in organotypic cultures of rat retina. AB - The rodent retina undergoes considerable postnatal neurogenesis and phenotypic differentiation, and it is likely that diffusible neurotrophic factors contribute to this development and to the subsequent formation of functional retinal circuitry. Accordingly, perturbation of specific neurotrophin ligand-receptor interactions has provided valuable information as to the fundamental processes underlying this development. In the present studies we have built upon our previous observation that suppression of expression of trk(B), the high-affinity receptor for brain-derived neurotrophic factor (BDNF), in the postnatal rat retina results in the alteration of a specific interneuron in the rod pathway-the parvalbumin (PV)-immunoreactive AII amacrine cell. Here, we isolated retinas from newborn rats and maintained them in organotypic culture for up to 14 days (approximating the time of eye opening, in vivo) in the presence of individual neurotrophins [BDNF or nerve growth factor (NGF)]. We then examined histological sections of cultures for PV immunoreactivity. In control cultures, only sparse PV immunostained cells were observed. In cultures supplemented with NGF, numerous lightly immunostained somata were present in the inner nuclear layer (INL) at the border of the inner plexiform layer (IPL). Many of these cells had rudimentary dendritic arborizations in the IPL. Cultures supplemented with BDNF displayed numerous well-immunostained somata at the INL/IPL border that gave rise to elaborate dendritic arborizations that approximated the morphology of mature AII amacrine cells in vivo. These observations indicate that neurotrophins have specific effects upon the neurochemical and, perhaps, morphological differentiation of an important interneuron in a specific functional retinal circuit. PMID- 10526317 TI - Protein kinase C prevents oligodendrocyte differentiation: modulation of actin cytoskeleton and cognate polarized membrane traffic. AB - In a previous study, we showed that activation of protein kinase C (PKC) prevents oligodendrocyte differentiation at the pro-oligodendrocyte stage. The present study was undertaken to identify downstream targets of PKC action in oligodendrocyte progenitor cells. Activation of PKC induced the predominant phosphorylation of an 80-kD protein, identified as myristoylated alanine-rich C kinase substrate (MARCKS). Upon phosphorylation, MARCKS is translocated from the plasma membrane to the cytosol. Furthermore, PKC activation perturbed the organization of the actin cytoskeleton, causing a redistribution of actin filaments to the submembranous or cortical actin cytoskeleton. As a consequence, transport of a protein traffic marker, the vesicular stomatitis virus glycoprotein, from the trans-Golgi network to the plasma membrane becomes perturbed. The effect of disruption of the actin filament network by cytochalasin D perfectly matched the effect of PKC. These data thus favor the existence of a causal relationship between actin rearrangement and docking and/or fusion of proteins to the plasma membrane. Interestingly, neither in control cells nor in PKC-activated cells did another protein traffic marker, influenza hemagglutinin (HA), reach the cell surface. However, an eminent and specific accumulation of HA just underneath the plasma membrane became apparent upon PKC activation. Yet, this effect could not be simulated by cytochalasin D treatment. Therefore, these observations imply that although MARCKS represents a prominent PKC target site in regulating differentiation, another target involves the differential control of cognate polarized trafficking pathways, which are apparently operating in oligodendrocyte progenitor cells. PMID- 10526318 TI - Family of prohormone convertases in Lymnaea: characterization of two alternatively spliced furin-like transcripts and cell-specific regulation of their expression. AB - The majority of neuropeptides in Lymnaea stagnalis are proteolytically processed from larger precursors at sites composed of single or multiple basic amino acid residues. Previous studies have identified several putative prohormone convertases in the brain of Lymnaea. To characterize the complete family, we undertook three independent approaches: reverse-transcribed polymerase chain reaction screening, and low-stringency cDNA and genomic library screenings. The central nervous system cDNA library screening yielded two cDNAs encoding Lfurin1 and its variant form, Lfurin1-X. Both proteins show the characteristic organization of (human) furin with a putative catalytic domain, a P domain, a Cys rich domain, a transmembrane domain, and a cytoplasmic tail. Lfurin1 and Lfurin1 X are identical, apart from a putative alternatively spliced noncatalytic luminal protein domain, which is present exclusively in Lfurin1-X. In situ hybridization revealed that the Lfur1 gene is expressed throughout the Lymnaea brain, but that the level varies considerably from one neuron to another. Quantitative analysis of the expression level of the two alternatively spliced transcripts revealed that it is neuron type-specifically regulated. This probably indicates the functional importance of noncatalytic luminal protein domains in these enzymes. In addition, our findings suggest that apart from the identified convertases LPC2, Lfurin1/Lfurin1-X, and Lfurin2, additional prohormone convertase diversity is either not present or present only at low levels in the Lymnaea brain. Alternatively, additional prohormone convertases could exist with a lower degree of sequence conservation than the other Lymnaea prohormone convertase members. From our findings, it appears that the majority of prohormone processing in Lymnaea is carried out by the three thus far identified types of Kex2-related prohormone convertases despite the large number of neuropeptide precursors and diverse multiple basic cleavage sites hydrolyzed. PMID- 10526319 TI - Alcohol exposure during the first two trimesters equivalent alters granule cell number and neurotrophin expression in the developing rat olfactory bulb. AB - Although alcohol has been shown to affect brain development adversely, the underlying mechanism of alcohol's actions are poorly understood. The present study addressed the hypothesis that alcohol affects growth factor availability during critical periods of neural growth by measuring the mRNA expression of brain-derived neurotrophic factor (BDNF), a potent developmental growth factor. Multiple offspring of timed-pregnant rat dams given alcohol (6.0 g/kg per day) or control treatments during gestation were sacrificed at either embryonic (E) day 21 or E33 (usually postnatal day 10) when their olfactory bulbs were processed for molecular analyses or neuron counting. BDNF mRNA levels were measured by reverse-transcription-polymerase chain reaction, and DNA methylation of the BDNF gene was quantified by Southern blot analyses following digestion with methylation-sensitive enzymes. Estimates of total granule cell number were obtained by counting those cells using unbiased stereological techniques. There was a significant decrease in BDNF mRNA levels in the alcohol-exposed offspring of both ages compared with controls. In addition, the number of olfactory bulb granule cells significantly decreased in the E33 but not the E21 rat pups exposed to alcohol compared with their appropriate aged controls. Finally, BDNF DNA of alcohol-exposed animals was less susceptible to digestion with the methylation sensitive enzyme HpaII compared with controls, suggesting that the DNA of the alcohol exposed pups was hypermethylated. Our results indicate that exposure to alcohol during early brain development in the rat, a period equivalent to the first two trimesters in humans, can have a detrimental effect on normal development of the olfactory bulb by reducing the number of BDNF-synthesizing neurons. Although the exact mechanism for the alcohol-induced neuronal loss is unknown, the inappropriate transcription of the BDNF gene is one mechanism that may account for the complexity of effects observed in offspring exposed to heavy alcohol exposure in utero. PMID- 10526320 TI - Defective calmodulin-dependent rapid apical endocytosis in zebrafish sensory hair cell mutants. AB - Vertebrate mechanosensory hair cells contain a narrow "pericuticular" zone which is densely populated with small vesicles between the cuticular plate and cellular junctions near the apical surface. The presence of many cytoplasmic vesicles suggests that the apical surface of hair cells has a high turnover rate. The significance of intense membrane trafficking at the apical surface is not known. Using a marker of endocytosis, the styryl dye FM1-43, this report shows that rapid apical endocytosis in zebrafish lateral line sensory hair cells is calcium and calmodulin dependent and is partially blocked by the presence of amiloride and dihydrostreptomycin, known inhibitors of mechanotransduction channels. As seen in lateral line hair cells, sensory hair cells within the larval otic capsule also exhibit rapid apical endocytosis. Defects in internalization of the dye in both lateral line and inner ear hair cells were found in five zebrafish auditory/vestibular mutants: sputnik, mariner, orbiter, mercury, and skylab. In addition, lateral line hair cells in these mutants were not sensitive to prolonged exposure to streptomycin, which is toxic to hair cells. The presence of endocytic defects in the majority of zebrafish mechanosensory mutants points to a important role of apical endocytosis in hair cell function. PMID- 10526321 TI - Nasotemporal asymmetry during teleost retinal growth: preserving an area of specialization. AB - Teleost fish retinas grow throughout adult life through both cell addition and stretching. Cell division occurs at the periphery of the retina, resulting in annular addition of all cell types except rod photoreceptors, which are added in the central retina. Since many teleosts have a region of high cellular density at the temporal pole of the eye, we analyzed whether and how this specialized region of high visual acuity maintained its relative topographical position through asymmetric circumferential growth. To do this, we measured the pattern of long term retinal growth in the African cichlid Haplochromis burtoni. We found that the retina expands asymmetrically along the nasotemporal axis, with the nasal retina growing at a higher rate than the temporal, dorsal, or ventral retinae, whose growth rates are equal. This nasotemporal asymmetry is produced via significantly greater expansion of retinal tissue at the nasal pole rather than through differential cell proliferation. The mechanisms responsible for this differential retinal enlargement are unknown; however, such asymmetric expansion very likely minimizes disruption in vision during rapid growth. PMID- 10526322 TI - A mouse's tale that grew in the telling. PMID- 10526323 TI - On the neurology of morals. AB - Patients with medial prefrontal lesions often display irresponsible behavior, despite being intellectually unimpaired. But similar lesions occurring in early childhood can also prevent the acquisition of factual knowledge about accepted standards of moral behavior. PMID- 10526324 TI - UNC-13 and neurotransmitter release. AB - Genetic studies in mice, worms and flies indicate that the synaptic protein UNC 13 plays a central role in the regulation of vesicle fusion. PMID- 10526325 TI - A new savior for neurons. AB - How NGF promotes neuronal survival is unclear. New findings suggest that the mechanism involves induction of an anti-apoptotic protein called ITA. PMID- 10526327 TI - Creating teetotaler mice. PMID- 10526326 TI - News on views: pandemonium revisited. AB - How do we recognize objects from different viewpoints? A new model, based on the known properties of cortical neurons, may help resolve this long-standing debate. PMID- 10526328 TI - Brain Research for Policy Wonks. PMID- 10526329 TI - Reciprocal regulation of P/Q-type Ca2+ channels by SNAP-25, syntaxin and synaptotagmin. PMID- 10526330 TI - TIP39: a new neuropeptide and PTH2-receptor agonist from hypothalamus. PMID- 10526331 TI - A sex difference in the hypothalamus of the spotted hyena. PMID- 10526332 TI - Information theory and neural coding. AB - Information theory quantifies how much information a neural response carries about the stimulus. This can be compared to the information transferred in particular models of the stimulus-response function and to maximum possible information transfer. Such comparisons are crucial because they validate assumptions present in any neurophysiological analysis. Here we review information-theory basics before demonstrating its use in neural coding. We show how to use information theory to validate simple stimulus-response models of neural coding of dynamic stimuli. Because these models require specification of spike timing precision, they can reveal which time scales contain information in neural coding. This approach shows that dynamic stimuli can be encoded efficiently by single neurons and that each spike contributes to information transmission. We argue, however, that the data obtained so far do not suggest a temporal code, in which the placement of spikes relative to each other yields additional information. PMID- 10526333 TI - UNC-13 is required for synaptic vesicle fusion in C. elegans. AB - We analyzed the synaptic physiology of unc-13 mutants in the nematode C. elegans. Mutants of unc-13 had normal nervous system architecture, and the densities of synapses and postsynaptic receptors were normal at the neuromuscular junction. However, the number of synaptic vesicles at neuromuscular junctions was two- to threefold greater in unc-13 mutants than in wild-type animals. Most importantly, evoked release at both GABAergic and cholinergic synapses was almost absent in unc-13 null alleles, as determined by whole-cell, voltage-clamp techniques. Although mutant synapses had morphologically docked vesicles, these vesicles were not competent for release as assayed by spontaneous release in calcium-free solution or by the application of hyperosmotic saline. These experiments support models in which UNC-13 mediates either fusion of vesicles during exocytosis or priming of vesicles for fusion. PMID- 10526334 TI - Drosophila UNC-13 is essential for synaptic transmission. AB - The UNC-13 protein family has been suggested to be critical for synaptic vesicle dynamics based on its interactions with Syntaxin, Munc-18 and Doc 2alpha. We cloned the Drosophila homolog (Dunc-13) and characterized its function using a combination of electrophysiology and ultrastructural analyses. Dunc-13 contained a C1 lipid-binding motif and two C2 calcium-binding domains, and its expression was restricted to neurons. Elimination of dunc-13 expression abolished synaptic transmission, an effect comparable only to removal of the core complex proteins Syntaxin and Synaptobrevin. Transmitter release remained impaired under elevated calcium influx or application of hyperosmotic saline. Ultrastructurally, mutant terminals accumulated docked vesicles at presynaptic release sites. We conclude that Dunc-13 is essential for a stage of neurotransmission following vesicle docking and before fusion. PMID- 10526335 TI - AMPA receptor-PDZ interactions in facilitation of spinal sensory synapses. AB - Silent synapses form between some primary sensory afferents and dorsal horn neurons in the spinal cord. Molecular mechanisms for activation or conversion of silent synapses to conducting synapses are unknown. Serotonin can trigger activation of silent synapses in dorsal horn neurons by recruiting AMPA receptors. AMPA-receptor subunits GluR2 and GluR3 interact via their cytoplasmic C termini with PDZ-domain-containing proteins such as GRIP (glutamate receptor interacting protein), but the functional significance of these interactions is unclear. Here we demonstrate that protein interactions involving the GluR2/3 C terminus are important for serotonin-induced activation of silent synapses in the spinal cord. Furthermore, PKC is a necessary and sufficient trigger for this activation. These results implicate AMPA receptor-PDZ interactions in mechanisms underlying sensory synaptic potentiation and provide insights into the pathogenesis of chronic pain. PMID- 10526336 TI - The anti-apoptotic protein ITA is essential for NGF-mediated survival of embryonic chick neurons. AB - The avian ITA is homologous to the baculoviral and mammalian inhibitor of apoptosis (IAP) proteins, which can prevent apoptosis by inhibition of specific caspases. We investigated the role of ITA in embryonic chick sympathetic and dorsal root ganglionic neurons, which depend on nerve growth factor (NGF) for their survival. Within 6 hours, NGF upregulated ITA protein production more than 25-fold in sensory and sympathetic neurons. Overexpression of ITA in primary neurons supported survival of these cells in the absence of NGF, and ita antisense constructs inhibited NGF-mediated survival. Thus the induction of ITA expression seems to be an essential signaling event for survival of sympathetic and dorsal root ganglionic sensory neurons in response to NGF. PMID- 10526338 TI - In vivo dendritic calcium dynamics in deep-layer cortical pyramidal neurons. AB - Dendritic Ca2+ action potentials in neocortical pyramidal neurons have been characterized in brain slices, but their presence and role in the intact neocortex remain unclear. Here we used two-photon microscopy to demonstrate Ca2+ electrogenesis in apical dendrites of deep-layer pyramidal neurons of rat barrel cortex in vivo. During whisker stimulation, complex spikes recorded intracellularly from distal dendrites and sharp waves in the electrocorticogram were accompanied by large dendritic [Ca2+ ] transients; these also occurred during bursts of action potentials recorded from somata of identified layer 5 neurons. The amplitude of the [Ca 2+] transients was largest proximal to the main bifurcation, where sodium action potentials produced little Ca2+ influx. In some cases, synaptic stimulation evoked [Ca2+] transients without a concomitant action potential burst, suggesting variable coupling between dendrite and soma. PMID- 10526337 TI - Furin mediates enhanced production of fibrillogenic ABri peptides in familial British dementia. AB - The genetic lesion underlying familial British dementia (FBD), an autosomal dominant neurodegenerative disorder, is a T-A transversion at the termination codon of the BRI gene. The mutant gene encodes BRI-L, the precursor of ABri peptides that accumulate in amyloid deposits in FBD brain. We now report that both BRI-L and its wild-type counterpart, BRI, were constitutively processed by the proprotein convertase, furin, resulting in the secretion of carboxyl-terminal peptides that encompass all or part of ABri. Elevated levels of peptides were generated from the mutant BRI precursor. Electron microscopic studies revealed that synthetic ABri peptides assembled into irregular, short fibrils. Collectively, our results support the view that enhanced furin-mediated processing of mutant BRI generates fibrillogenic peptides that initiate the pathogenesis of FBD. PMID- 10526339 TI - Supersensitivity to allosteric GABA(A) receptor modulators and alcohol in mice lacking PKCepsilon. AB - Several of the actions of ethanol are mediated by gamma-aminobutyrate type A (GABA(A)) receptors. Here we demonstrated that mutant mice lacking protein kinase C epsilon (PKCepsilon) were more sensitive than wild-type littermates to the acute behavioral effects of ethanol and other drugs that allosterically activate GABA(A) receptors. GABA(A) receptors in membranes isolated from the frontal cortex of PKCepsilon null mice were also supersensitive to allosteric activation by ethanol and flunitrazepam. In addition, these mutant mice showed markedly reduced ethanol self-administration. These findings indicate that inhibition of PKCepsilon increases sensitivity of GABA(A) receptors to ethanol and allosteric modulators. Pharmacological agents that inhibit PKCepsilon may be useful for treatment of alcoholism and may provide a non-sedating alternative for enhancing GABA(A) receptor function to treat other disorders such as anxiety and epilepsy. PMID- 10526340 TI - Odor- and context-dependent modulation of mitral cell activity in behaving rats. AB - The projections and odor responses of mammalian olfactory receptor neurons, as well as the physiology of the bulb's principal neurons-the mitral cells (MCs)-are known from studies in slices and anesthetized animals. In behaving rats trained to discriminate between two odors associated with different reinforcers, we examined MC responses following alternated odor-reinforcer pairings. Whereas only 11% of the recorded MCs showed changes in odor-selective firing rate during the odor-sampling phase, 94% of MCs modulated activity during specific behaviors surrounding odor sampling. These cell- and odor-selective responses were not primary sensory responses; rather, they depended (reversibly) on the predictive value of each odor. MC activity thus depends critically on efferent influences linked to the animal's experience and behavior. PMID- 10526341 TI - The effects of color on brightness. AB - Observation of human subjects shows that the spectral returns of equiluminant colored surrounds govern the apparent brightness of achromatic test targets. The influence of color on brightness provides further evidence that perceptions of luminance are generated according to the empirical frequency of the possible sources of visual stimuli, and suggests a novel way of understanding color contrast and constancy. PMID- 10526342 TI - Neural strength of visual attention gauged by motion adaptation. AB - Single-cell and neuroimaging studies reveal that attention focused on a visual object markedly amplifies neural activity produced by features of the attended object. In a psychophysical study, we found that visual attention could modulate the strength of weak motion signals to the point that the perceived direction of motion, putatively registered early in visual processing, was powerfully altered. This strong influence of attention on early motion processing, beside complementing neurophysiological evidence for attentional modulation early in the visual pathway, can be measured in terms of equivalent motion energy, and thus provides a useful metric for quantifying attention's effects. PMID- 10526343 TI - Hierarchical models of object recognition in cortex. AB - Visual processing in cortex is classically modeled as a hierarchy of increasingly sophisticated representations, naturally extending the model of simple to complex cells of Hubel and Wiesel. Surprisingly, little quantitative modeling has been done to explore the biological feasibility of this class of models to explain aspects of higher-level visual processing such as object recognition. We describe a new hierarchical model consistent with physiological data from inferotemporal cortex that accounts for this complex visual task and makes testable predictions. The model is based on a MAX-like operation applied to inputs to certain cortical neurons that may have a general role in cortical function. PMID- 10526344 TI - Independent learning of internal models for kinematic and dynamic control of reaching. AB - Psychophysical studies of reaching movements suggest that hand kinematics are learned from errors in extent and direction in an extrinsic coordinate system, whereas dynamics are learned from proprioceptive errors in an intrinsic coordinate system. We examined consolidation and interference to determine if these two forms of learning were independent. Learning and consolidation of two novel transformations, a rotated spatial reference frame and altered intersegmental dynamics, did not interfere with each other and consolidated in parallel. Thus separate kinematic and dynamic models were constructed simultaneously based on errors computed in different coordinate frames, and possibly, in different sensory modalities, using separate working-memory systems. These results suggest that computational approaches to motor learning should include two separate performance errors rather than one. PMID- 10526345 TI - Impairment of social and moral behavior related to early damage in human prefrontal cortex. AB - The long-term consequences of early prefrontal cortex lesions occurring before 16 months were investigated in two adults. As is the case when such damage occurs in adulthood, the two early-onset patients had severely impaired social behavior despite normal basic cognitive abilities, and showed insensitivity to future consequences of decisions, defective autonomic responses to punishment contingencies and failure to respond to behavioral interventions. Unlike adult onset patients, however, the two patients had defective social and moral reasoning, suggesting that the acquisition of complex social conventions and moral rules had been impaired. Thus early-onset prefrontal damage resulted in a syndrome resembling psychopathy. PMID- 10526346 TI - Critical assessment of methods of protein structure prediction (CASP): round III. PMID- 10526347 TI - Automated large scale evaluation of protein structure predictions. AB - Evaluation and assessment are critical issues in CASP experiments. Automated procedures are necessary to compare a large number of predictions with the target folds. The evaluation has to reveal the maximum extent of similarity between predictions and targets, it should be applicable across prediction categories, and it should be transparent and accessible to a wide community. Here we present an automated evaluation scheme which is an attempt to meet these requirements. In the implementation and execution of this scheme we had to solve or circumvent problems of convergence, where algorithms fail to find optimum solutions, problems of ambiguity where no unique optimum solution exists, and problems in ranking and interpretation. Key features of this implementation are (1) the root mean square deviation of structure superimposition is kept close to a constant value throughout the evaluation and (2) all structural matches found between two folds are taken into account. We discuss these points in detail and describe the numerical criteria used in the CASP3 evaluation. PMID- 10526348 TI - RMS/coverage graphs: a qualitative method for comparing three-dimensional protein structure predictions. AB - Evaluating a set of protein structure predictions is difficult as each prediction may omit different residues and different parts of the structure may have different accuracies. A method is described that captures the best results from a large number of alternative sequence-dependent structural superpositions between a prediction and the experimental structure and represents them as a single line on a graph. Applied to CASP2 and CASP3 data the best predictions stand out visually in most cases, as judged by manual inspection. The results from this method applied to CASP data are available from the URLs http:/(/)PredictionCenter. llnl.gov/casp3/results/th/ and http:/(/)www.sanger.ac.uk/ approximately th/casp/. PMID- 10526349 TI - Processing and analysis of CASP3 protein structure predictions. AB - Livermore Prediction Center provides basic infrastructure for the CASP (Critical Assessment of Structure Prediction) experiments, including prediction processing and verification servers, a system of prediction evaluation tools, and interactive numerical and graphical displays. Here we outline the essentials of our approach, with discussion of the superposition procedures, definitions of basic measures, and descriptions of new methods developed to analyze predictions. Our primary focus is on the evaluation of three-dimensional models and secondary structure predictions. To put the results of the three prediction experiments held to date on the same footing, the latest CASP3 evaluation criteria were retrospectively applied to both CASP1 and CASP2 predictions. Finally, we give an overview of our website (http:/(/)PredictionCenter.llnl.gov), which makes the target structures, predictions, and the evaluation system accessible to the community. PMID- 10526350 TI - CASP3 comparative modeling evaluation. AB - We report our evaluation of the CASP3 comparative modelling competition. Our analysis covers the accuracy of the over-all fold, the bridging of insertions and deletions, and the adding of side-chains. We describe our attempts at automating aspects of the evaluation. PMID- 10526351 TI - Model building by comparison at CASP3: using expert knowledge and computer automation. AB - Ten models were constructed for the comparative modeling section of the Critical Assessment of Techniques for Protein Structure Prediction-3 (CASP3). Sequence identity between each target and the best possible parent(s) ranged between 12% and 64%. The modeling protocol is a mixture of automated computer algorithms with human intervention at certain critical stages. In particular, intervention is required to check sequence alignments and the selection of parameters for various computer programs. Seven of the targets were constructed from single-parent templates, and three were constructed from multiple parents. The reasons for such a high ratio of modeling from single parents only are discussed. Models constructed from multiple parents were found to be more accurate than models constructed from single parents only. A novel loop-modeling algorithm is presented that consists of fragment database searches, several fragment libraries, and mean-field calculations on representative fragment candidates. PMID- 10526352 TI - An iterative structure-assisted approach to sequence alignment and comparative modeling. AB - Correct alignment of the sequence of a target protein with those of homologues of known three-dimensional structure is a key step in comparative modeling. Usually an iterative approach that takes account of the local and overall structural features is required. We describe such an approach that exploits databases of structural alignments of homologous proteins (HOMSTRAD, http:/(/)www cryst.bioc.cam.ac.uk/ approximately homstrad) and protein superfamilies (CAMPASS, http:/(/)www-cryst.bioc.cam.ac.uk/ approximately campass), in which structure based alignments are analyzed and formatted with the program JOY (http:/(/)www cryst.bioc.cam.ac.uk/ approximately joy) to reveal conserved local structural features. The databases facilitate the recognition of a family or superfamily, they assist in the selection of useful parent structures, they are helpful in alignment of the target sequences with the parent set, and are useful for deriving relationships that can be used in validating models. In the iterative approach, a model is constructed on the basis of the proposed sequence alignment and this is then reexpressed in the JOY format and realigned with the parent set. This is repeated until the model and sequence alignment is optimized. We examine the case for comparison and use of multiple structures of family members, rather than a single parent structure. We use the targets attempted by our group in CASP3 to assess the value of such procedures. PMID- 10526353 TI - Modeling three-dimensional protein structures for amino acid sequences of the CASP3 experiment using sequence-derived predictions. AB - Homology or comparative modeling is aimed at modeling the three-dimensional structure of a target sequence of unknown structure using the framework of an already known fold. Traditionally, homology modeling has been applied to targets with clear sequence similarity to proteins of known structure. Because methods to identify increasingly distant relationships have been developed, homology models can now be built for a wider range of targets. The first challenge in homology modeling is to obtain an initial, accurate, sequence-structure alignment with the most compatible fold. In CASP3, the abilities of fold-recognition methods to fulfill this challenge were evaluated with a number of target sequences of unknown structure. Sequence-structure alignments for 33 of the CASP3 targets using the fold-recognition method SDP were submitted (Fischer and Eisenberg, Protein Sci 1996; 5:947-955). After the three-dimensional structures of the sequences were subsequently released, the quality of the predictions were evaluated. Here I describe three of the predictions for targets with little sequence similarity to proteins of known structure that were judged by the assessors to be of higher quality. For two of these predictions, the sequence structure alignment corresponded perfectly to the structural alignment (zero average shift), and for the third, the average shift was 0.1. This alignment accuracy entails an ideal starting point for homology modeling. PMID- 10526354 TI - Sequence to structure alignment in comparative modeling using PrISM. AB - PrISM (Protein Informatics System for Modeling) is a protein analysis and modeling system in which informatics, alignment, modeling, and assessment modules are integrated in a computational environment where protein analysis and modeling protocols can be designed and assessed interactively. It can then be used automatically and repetitively in response to a variety of protein analysis and modeling problems. PrISM was used to predict a single model for each of the 43 targets in the CASP3 experiment. In this paper, we present results for 13 target sequences, which we consider to be comparative modeling targets with clearly related structural templates. We emphasize the problem of aligning a target sequence to a template structure with various alignment methods. When more than one alignment method and/or parameter set are applied, the final alignment is chosen on the basis of a model ranking system also used in PrISM's fold recognition module. Advanced sequence-template alignment procedures in PrISM are useful in some cases when standard pairwise dynamic programming algorithm fail to make any reasonable global alignment. The same procedures, however, failed in other cases, corresponding to remotely related query-template pairs that involved extensive insertions and deletions. PMID- 10526355 TI - Addressing the issue of sequence-to-structure alignments in comparative modeling of CASP3 target proteins. AB - During a blind protein structure prediction experiment (the third round of the Critical Assessment of Techniques for Protein Structure Prediction; URL http://PredictionCenter.llnl.gov/casp3/) , four target proteins, T0047, T0048, T0055, and T0070, were modeled by comparison. These proteins display 62%, 29%, 24%, and 19% sequence identity, respectively, to the structurally homologous proteins most similar in sequence. The issue of sequence-to-structure alignment in cases of low sequence homology was the main emphasis. Selection of alignments was made by constructing and evaluating three-dimensional models based on series of samples produced mainly by automatic multiple sequence alignments. Sequence-to structure alignments were correct in all but two regions, in which significant changes in target structures compared with related proteins were the source of errors. Template choice is an important determinant of model quality, and a correct selection was made of a lower homology template for modeling of T0070; however, in the case of T0055, a template with 8% greater sequence homology proved deceptive. Loops and some ungapped template regions were assigned conformations taken from other proteins. Using fragments from homologous structures led to improvement over template backbone more often than cases in which nonhomologous structures were the source. The results also indicate that side-chain prediction accuracy depends not only on sequence similarity but also on accuracy of the backbone. PMID- 10526356 TI - Comparative modeling of CASP3 targets using PSI-BLAST and SCWRL. AB - We present results of comparative modeling on 11 targets from the CASP3 experiment. Our methods comprise the following steps: first, PSI-BLAST is used to find homologues of the target sequence in the nonredundant GenBank protein sequence database; second, after several iterations of PSI-BLAST, the resulting profile or position-specific similarity matrix is used to search a database of Protein Databank (PDB) sequences; third, from the list of hits resulting from the PDB search, a parent structure is chosen on the basis of the quality of the alignment and the quality of the experimental structure; fourth, this alignment is adjusted manually whenever insertions or deletions take place in secondary structure regions of the parent; fifth, the backbone is modeled from the parent structure and the alignment; and finally, the program SCWRL is used to replace nonconserved side chains onto the parent backbone given the target sequence. For comparison, we also produced structural models from the unaltered PSI-BLAST alignment, from an alignment from the nonprofile version of BLAST, and from the global sequence alignment program CLUSTAL W. Our results indicate that PSI-BLAST produced considerably better alignments than would be possible with either global or local pairwise sequence alignment algorithms and that manual adjustments were helpful. SCWRL, which uses a backbone-dependent rotamer library to predict side chain conformations, did well in comparison with other methods used in CASP3. PMID- 10526357 TI - Structure classification-based assessment of CASP3 predictions for the fold recognition targets. AB - The sequences of at least 23 of the 43 CASP3 targets showed no significant similarity to the sequences of known structures. The experimental structures of all but three of these 23 targets revealed substantial similarities to known structures, with at least eleven of the target structures likely being distantly homologous to known structures. Nineteen of the 23 target structures were available at the time of the final CASP3 meeting in Asilomar in December 1998, whereas the experimental data on the protein folds of the remaining four targets were obtained afterwards. The predicted three-dimensional structures for each of the 23 targets were analyzed to select those predictions sharing with the experimental structures a similar overall fold and/or having correctly folded a substantial fraction of the target sequence. Initially, predicted models were numerically evaluated and the evaluation results aided the selection process. Each target structure was then classified to identify a minimal set of structural features characteristic to its protein fold and evolutionary superfamily. The predictions containing this set were assessed comparatively to find the best predictions for each target. The predictions of new folds were assessed separately. The total number of the selected 'correct' predictions and the quality of these predictions were used to compare the performance of different predictor teams and different prediction methods in the fold prediction/recognition category. PMID- 10526358 TI - Successful recognition of protein folds using threading methods biased by sequence similarity and predicted secondary structure. AB - Analysis of our fold recognition results in the 3rd Critical Assessment in Structure Prediction (CASP3) experiment, using the programs THREADER 2 and GenTHREADER, shows an encouraging level of overall success. Of the 23 submitted predictions, 20 targets showed no clear sequence similarity to proteins of known 3D structure. These 20 targets can be divided into 22 domains, of which, 20 domains either entirely match a previously known fold, or partially match a substantial region of a known fold. Of these 20 domains, we correctly assigned the folds in 10 cases. PMID- 10526360 TI - Predicting protein structure using only sequence information. AB - This paper presents results of blind predictions submitted to the CASP3 protein structure prediction experiment. We made predictions using the SAM-T98 method, an iterative hidden Markov model-based method for constructing protein family profiles. The method is purely sequence-based, using no structural information, and yet was able to predict structures as well as all but five of the structure based methods in CASP3. PMID- 10526359 TI - Sustained performance of knowledge-based potentials in fold recognition. AB - We describe the results obtained using fold recognition techniques in our third participation in the CASP experiment. The approach relies on knowledge-based potentials for alignment production and fold identification. As indicated by the increase in alignment quality and fold identification reliability, the predictions improved from CASP1 to CASP3. In particular, we identified structural relationships in which no known evolutionary link exists. Our predictions are based on single sequences rather than multiple sequence alignments. Additionally, we voluntarily submitted only a single model for each target because, in our view, submission of a single model is the most stringent test. We describe the methods used, the strategy adopted in the predictions, and the prediction results and discuss future work. PMID- 10526361 TI - Cooperative approach for the protein fold recognition. AB - We, four independent predictors, organized a team and tackled blind protein structure predictions using fold recognition methods. We tried to assign the homologous or analogous folds in the protein structure database for a number of target sequences that showed no apparent sequence homology to the proteins of known folds. After primary analyses by conventional softwares, these sequences were threaded through the structural library using three different programs developed by ourselves, which employed different compatibility functions. Collecting the results of our individual analyses, and the available biological knowledge about the target, we held meetings and discussed all plausible structures for the target. For 25 target sequences, we submitted 56 models including NONE: This was the first time the fold was determined. At the time of the meeting (CASP3), 19 protein structures (21 domains) categorized as the threading targets were available. We succeeded in predicting eight out of 18 targets (20 domains) that we submitted; however, alignment accuracies were not satisfactory for some of the models. We often obtained correct answers even if some of us missed the right prediction; therefore it would appear that our threaders compensated each other. When all the information is managed effectively, the prediction gains more accuracy. PMID- 10526362 TI - Threading with explicit models for evolutionary conservation of structure and sequence. AB - We have attempted to predict the three-dimensional structures of 19 proteins for the CASP3 experiment, each showing less than 25% sequence identity with known structures. Predictions were based on a threading method that aligns the target sequence with the conserved cores of structural templates, as identified from structure-structure alignments of the template with homologous neighbors. Alternative alignments were scored using contact potentials and a position specific score matrix derived from sequence neighbors of the template. We find that this method identified the correct structural family for 11 of the 19 targets and predicted the remaining 8 targets to be similar to "none" of the templates, avoiding false positives. Threading alignments are relatively accurate for 10 of the 11 targets, including alignments for 6 of 7 identified at CASP3 as fold-recognition targets. These predictions were ranked "first place" by the CASP3 assessor when compared to fold-recognition predictions made by other methods. It appears that threading with family-specific models for structure and sequence conservation has improved threading prediction accuracy. PMID- 10526363 TI - Fold recognition using sequence and secondary structure information. AB - We applied a succession of sequence search and structure prediction methods to the targets in the fold recognition part of the CASP3 experiment. For each target, we expanded an initial sequence space, obtained through PSI-BLAST, by searching for statistically significant relationships to low-scoring sequences and then by searching for conserved sequence patterns. We then divided the proteins in the sequence space into families and built an alignment hierarchically, using the multiple alignment program MACAW. If no significant similarity to a protein of known structure was apparent at this point, we submitted the alignment to the Jpred server for consensus secondary structure prediction and searched the structure space using the secondary structure mapping program MAP. Failing this, we compared the structural properties that we believed we recognized in the aligned proteins to the folds in the SCOP database, using visual inspection. If all these methods failed to uncover a plausible match, we predicted that the target would adopt a novel fold. This procedure yielded correct answers for seven of twenty-one targets and a partly correct answer for one. A retrospective analysis shows that automating the sequence search procedures would have represented a significant improvement, with at least three additional correct predictions. PMID- 10526364 TI - Analysis and assessment of ab initio three-dimensional prediction, secondary structure, and contacts prediction. AB - CASP3 saw a substantial increase in the volume of ab initio 3D prediction data, with 507 datasets for fifteen selected targets and sixty-one groups participating. As with CASP2, methods ranged from computationally intensive strategies that attempt to recreate the physical and chemical forces involved in protein folding to the more recent knowledge-based approaches. These exploit information from the structure databases, extracting potentially similar fragments and/or distance constraints derived from multiple sequence alignments. The knowledge-based approaches generally gave more consistently successful predictions across the range of targets, particularly that of the Baker group (Bystroff and Baker, J Mol Biol 1998;281:565-577; Simons et al. Proteins Suppl 1999;3:171-176), which used a fragment library. In the secondary structure prediction category, the most successful approaches built on the concepts used in PHD (Rost et al. Comput Appl Biosci 1994;10:53-60), an accepted standard in this field. Like PHD, they exploit neural networks but have different strategies for incorporating multiple sequence data or position-dependent weight matrices for training the networks. Analysis of the contact data, for which only six groups participated, suggested that as yet this data provides a rather weak signal. However, in combination with other types of prediction data it can sometimes be a useful constraint for identifying the correct structure. PMID- 10526365 TI - Ab initio protein structure prediction of CASP III targets using ROSETTA. AB - To generate structures consistent with both the local and nonlocal interactions responsible for protein stability, 3 and 9 residue fragments of known structures with local sequences similar to the target sequence were assembled into complete tertiary structures using a Monte Carlo simulated annealing procedure (Simons et al., J Mol Biol 1997; 268:209-225). The scoring function used in the simulated annealing procedure consists of sequence-dependent terms representing hydrophobic burial and specific pair interactions such as electrostatics and disulfide bonding and sequence-independent terms representing hard sphere packing, alpha helix and beta-strand packing, and the collection of beta-strands in beta-sheets (Simons et al., Proteins 1999;34:82-95). For each of 21 small, ab initio targets, 1,200 final structures were constructed, each the result of 100,000 attempted fragment substitutions. The five structures submitted for the CASP III experiment were chosen from the approximately 25 structures with the lowest scores in the broadest minima (assessed through the number of structural neighbors; Shortle et al., Proc Natl Acad Sci USA 1998;95:1158-1162). The results were encouraging: highlights of the predictions include a 99-residue segment for MarA with an rmsd of 6.4 A to the native structure, a 95-residue (full length) prediction for the EH2 domain of EPS15 with an rmsd of 6.0 A, a 75-residue segment of DNAB helicase with an rmsd of 4.7 A, and a 67-residue segment of ribosomal protein L30 with an rmsd of 3.8 A. These results suggest that ab initio methods may soon become useful for low-resolution structure prediction for proteins that lack a close homologue of known structure. PMID- 10526366 TI - Ab initio folding of proteins using restraints derived from evolutionary information. AB - We present our predictions in the ab initio structure prediction category of CASP3. Eleven targets were folded, using a method based on a Monte Carlo search driven by secondary and tertiary restraints derived from multiple sequence alignments. Our results can be qualitatively summarized as follows: The global fold can be considered "correct" for targets 65 and 74, "almost correct" for targets 64, 75, and 77, "half-correct" for target 79, and "wrong" for targets 52, 56, 59, and 63. Target 72 has not yet been solved experimentally. On average, for small helical and alpha/beta proteins (on the order of 110 residues or smaller), the method predicted low resolution structures with a reasonably good prediction of the global topology. Most encouraging is that in some situations, such as with target 75 and, particularly, target 77, the method can predict a substantial portion of a rare or even a novel fold. However, the current method still fails on some beta proteins, proteins over the 110-residue threshold, and sequences in which only a poor multiple sequence alignment can be built. On the other hand, for small proteins, the method gives results of quality at least similar to that of threading, with the advantage of not being restricted to known folds in the protein database. Overall, these results indicate that some progress has been made on the ab initio protein folding problem. Detailed information about our results can be obtained by connecting to http:/(/)www.bioinformatics.danforthcenter.org/+ ++CASP3. PMID- 10526367 TI - Improved ab initio predictions with a simplified, flexible geometry model. AB - Structure predictions for nine targets from the CASP3 meeting are presented and compared with the experimental structure. These predictions are made using the simplified flexible geometry representation of protein structure and potentials which mimic the physical forces involved in protein folding with no help from multiple sequences. The major differences from the CASP2 potentials are identified, and the prediction successes and failures are related to the underlying potentials. Target T0065 was successfully folded from an extended chain to 3.8 A CA RMS of the native structure. The quality of the secondary structure component of all predictions was significantly improved, averaging a Q3 of 64%, which was spread evenly across the fold types, all alpha, alpha/beta mixed, and mainly beta. A number of the other predictions had a spatial arrangement of the secondary structure segments close to native, although the major problem with most predictions was the over-extension of secondary structure segments which often coalesced independent segments together. For target T0056 the ab initio prediction was the closest to the native structure of all methods including threading according to the Hubbard RMS coverage plots. PMID- 10526368 TI - Ab initio protein structure prediction using a combined hierarchical approach. AB - As part of the third Critical Assessment of Structure Prediction meeting (CASP3), we predict the three-dimensional structures for 13 proteins using a hierarchical approach. First, all possible compact conformations of a protein sequence are enumerated using a highly simplified tetrahedral lattice model. We select a large subset of these conformations using a lattice-based scoring function and build detailed all-atom models incorporating predicted secondary structure. A combined all-atom knowledge-based scoring function is then used to select three smaller subsets from these all-atom models. Finally, a consensus-based distance geometry procedure is used to generate the best conformations from each of the all-atom subsets. With this method, we are able to predict the global topology/shape for all or a large part of the sequence for six out of the thirteen proteins. For two other proteins, the topology/shape for shorter fragments are predicted. This represents a marked improvement in ab initio prediction since CASP was first instigated in 1994. PMID- 10526369 TI - Prediction of protein structure: the problem of fold multiplicity. AB - Three-dimensional (3D) models of four CASP3 targets were calculated using a simple modeling procedure that includes prediction of regular secondary structure, analysis of possible beta-sheet topologies, assembly of amphiphilic helices and beta-sheets to bury their nonpolar surfaces, and adjustment of side chain conformers and loops to provide close packing and saturation of the "hydrogen bond potential" (exposure of all polar groups to water or their involvement in intramolecular hydrogen bonds). It has been found that this approach allows construction of 3D models that, in some cases, properly reproduce the structural class of the protein (such as beta-barrel or beta-sandwich of definite shape and size) and details of tertiary structure (such as pairing of beta-strands), although all four models were more or less incorrect. Remarkably, some models had fewer water-exposed nonpolar side-chains, more hydrogen bonds, and smaller holes than the corresponding native structures (although the models had a larger water-accessible nonpolar surface). The results obtained indicate that hydrophobicity patterns do not unequivocally determine protein folds, and that any ab initio or fold recognition methods that operate with imprecise potential energy functions, or use crude geometrical approximations of the peptide chain, will probably produce many different nonnative structures. PMID- 10526370 TI - Calculation of protein conformation by global optimization of a potential energy function. AB - A novel hierarchical approach to protein folding has been applied to compute the unknown structures of seven target proteins provided by CASP3. The approach is based exclusively on the global optimization of a potential energy function for a united-residue model by conformational space annealing, followed by energy refinement using an all-atom potential. Comparison of the submitted models for five globular proteins with the experimental structures shows that the conformations of large fragments (approximately 60 aa) were predicted with rmsds of 4.2-6.8 A for the C alpha atoms. Our lowest-energy models for targets T0056 and T0061 were particularly successful, producing the correct fold of approximately 52% and 80% of the structures, respectively. These results support the thermodynamic hypothesis that protein structure can be computed solely by global optimization of a potential energy function for a given amino acid sequence. PMID- 10526372 TI - A measure of progress in fold recognition? AB - We present a retrospective analysis of CASP3 threading predictions, applying evaluation and assessment criteria used at CASP2. Our purpose is twofold. First, we wish to ask whether measures of model accuracy are comparable between CASP3 and CASP2, even though they have been calculated differently. We find that these quantities are effectively the same, and that either may be used to compare model accuracy. Secondly, we wish to assess progress in fold recognition by comparing the numbers of CASP2 and CASP3 models that cross specific accuracy thresholds. We find that the number of accurate models at CASP3 drops sharply as the targets become more difficult, with less extensive similarity to known structures, exactly the pattern seen at CASP2. CASP3 teams do not seem to have predicted accurate models for targets of greater difficulty, and for a given difficulty range the best CASP3 models seem no more accurate than the best models at CASP2. At CASP3, however, we find greater numbers of accurate models for medium difficulty targets, with extensive similarity to a known structure but no shared sequence motifs. Threading methods would appear to have become more reliable for modeling based on remote evolutionary relationships. PMID- 10526371 TI - CAFASP-1: critical assessment of fully automated structure prediction methods. AB - The results of the first Critical Assessment of Fully Automated Structure Prediction (CAFASP-1) are presented. The objective was to evaluate the success rates of fully automatic web servers for fold recognition which are available to the community. This study was based on the targets used in the third meeting on the Critical Assessment of Techniques for Protein Structure Prediction (CASP-3). However, unlike CASP-3, the study was not a blind trial, as it was held after the structures of the targets were known. The aim was to assess the performance of methods without the user intervention that several groups used in their CASP-3 submissions. Although it is clear that "human plus machine" predictions are superior to automated ones, this CAFASP-1 experiment is extremely valuable for users of our methods; it provides an indication of the performance of the methods alone, and not of the "human plus machine" performance assessed in CASP. This information may aid users in choosing which programs they wish to use and in evaluating the reliability of the programs when applied to their specific prediction targets. In addition, evaluation of fully automated methods is particularly important to assess their applicability at genomic scales. For each target, groups submitted the top-ranking folds generated from their servers. In CAFASP-1 we concentrated on fold-recognition web servers only and evaluated only recognition of the correct fold, and not, as in CASP-3, alignment accuracy. Although some performance differences appeared within each of the four target categories used here, overall, no single server has proved markedly superior to the others. The results showed that current fully automated fold recognition servers can often identify remote similarities when pairwise sequence search methods fail. Nevertheless, in only a few cases outside the family-level targets has the score of the top-ranking fold been significant enough to allow for a confident fully automated prediction. Because the goals, rules, and procedures of CAFASP-1 were different from those used at CASP-3, the results reported here are not comparable with those reported in CASP-3. Nevertheless, it is clear that current automated fold recognition methods can not yet compete with "human-expert plus machine" predictions. Finally, CAFASP-1 has been useful in identifying the requirements for a future blind trial of automated served-based protein structure prediction. PMID- 10526373 TI - An attempt to analyse progress in fold recognition from CASP1 to CASP3. AB - The Critical Assessment of Techniques for Protein Structure Prediction (CASP) experiment has been conducted for the third time. An obvious question is whether there has been progress from CASP1 to CASP3. An analysis depends on many variables, including prediction category, number and difficulty of targets, methods used to evaluate prediction success, and the rules for submission. It also depends on whether progress is measured in terms of all predictions submitted or in terms of the best predictions for each target. The progress made by individual groups is another interesting issue. In view of this complexity and the limited amount of data, an objective estimate of progress is difficult to obtain. Despite such difficulties, some estimate of progress is desirable. Here, we present an attempt to quantify progress in the fold-recognition category from CASP1 to CASP3. The numbers indicate clear progress from CASP1 to CASP2 but no improvement from CASP2 to CASP3. However, we argue that the targets in CASP3 are more difficult compared with CASP2, which translates into better performance of CASP3 over CASP2. PMID- 10526374 TI - Some measures of comparative performance in the three CASPs. AB - Performance in the three Critical Assessment of protein Structure Prediction (CASP) experiments has been compared in the areas of alignment accuracy for models based on homology and three-dimensional accuracy for models produced by using ab initio prediction methods. The homologous models span the comparative modeling and fold-recognition regimes. Each CASP target is assigned a relative difficulty based on the extent of sequence identity and the degree of structural overlap with the best available template. There is a clear improvement in alignment accuracy between CASP1 and CASPs 2 and 3 over much of the difficulty scale but no apparent improvement between CASP2 and CASP3. Encouragingly, the best ab initio models of small targets are clearly more accurate in CASP3 than in CASPs 1 and 2. PMID- 10526375 TI - Are children also fated to develop back pain? PMID- 10526376 TI - Gait analysis: matching the method to the goal. PMID- 10526377 TI - Back pain and spinal alignment abnormalities in schoolchildren. AB - OBJECTIVES: To study the prevalence of back pain and spinal alignment abnormalities in children aged 10 to 14 years; to define subsets of subjects with similar clinical profiles; and to identify factors associated with pain in the thoracic or lumbar spine. PATIENTS AND METHODS: 972 five- and nine-graders completed a back pain questionnaire at school and were examined by a school physician for spinal alignment abnormalities and for motion range limitation in the spine and/or lower limbs. Multivariate analysis was used to define clinical subsets and to identify factors associated with back pain. RESULTS: The point prevalence of back pain increased with age, from 14.3% in the ten-year-olds to 24% in the 14-year-olds. Girls were more likely than boys to report back pain, which was usually located in the low back. The prevalence of scoliosis increased with age and was higher in the girls. Multivariate analysis identified five clinical profiles: no spinal pain; nonserious spinal pain with no impact on medical service utilization or physical activities; spinal pain unrelated to an injury; injury-related spinal pain not treated by drugs or physical therapy; and injury-related spinal pain treated by drugs and physical therapy. Several factors associated with spinal pain were identified, with variations across the five groups. PMID- 10526378 TI - Nonspecific back pain in children. A search for associated factors in 14-year-old schoolchildren. AB - BACKGROUND: Nonspecific back pain in children is nearly as common as in adults but is associated with a number of age-specific risk factors including female gender, a family history of low back pain, a high level of physical activity, and prolonged sitting. OBJECTIVE: To investigate potential school-related risk factors for back pain in children, most notably schoolbag weight expressed as a percentage of body weight (relative schoolbag weight), whether the schoolbag is carried by hand or by a shoulder harness, how the child travels to and from school, and sitting positions. PATIENTS AND METHODS: 123 eighth-graders, 58 girls and 65 boys, with a mean age of 14 +/- 0.6 years, completed an anonymous self questionnaire during a school day involving six hours of classes. Their schoolbag was weighed on the same day. RESULTS: Most respondents traveled to and from school in a vehicle (70%), made one trip in each direction each day (75%), and carried their schoolbag by the shoulder harness (92%). The prevalence of back pain on the study day was 27.6%, whereas the cumulative prevalence for the last 12 months was 82.9% with 16.3% of respondents reporting a single episode of pain, 57.7% recurrent pain, and 8.9% chronic pain. A need for a physician visit for back pain was reported in 18.7% of cases, and 14.6% of respondents had missed school and/or sporting activities because of back pain. Female gender was associated with current back pain (odds ratio [OR], 2.7; 95% confidence interval [CI], 1.2-6.1). A relative schoolbag weight of 20% or more was associated with a history of back pain (OR, 3.1; 95% CI, 1.0-9.2), and this effect was larger in children who traveled to and from school on foot and in those who carried their schoolbag in their hand. Sitting on the edge of the chair while completing the questionnaire was significantly associated with a history of a physician visit for back pain (OR, 3.1; 95% CI, 1.0-9.5). Neither handedness nor the position of the questionnaire on the table were significantly associated with back pain in our study population. CONCLUSIONS: The findings from this cross-sectional study indicate a need for a longitudinal prospective study designed to identify etiologic and prognostic factors of back pain in adolescents, with the goal of devising preventive strategies likely to reduce the risk of low back pain in adulthood. PMID- 10526379 TI - Accelerometric gait analysis for use in hospital outpatients. AB - OBJECTIVES: To provide clinicians with a quantitative human gait analysis tool suitable for routine use. METHODS: We evaluated the reproducibility, sensitivity, and specificity of gait analysis based on measurements of acceleration at a point near the center of gravity of the body. Two accelerometers held over the middle of the low back by a semi-elastic belt were used to record craniocaudal and side to-side accelerations at a frequency of 50 Hz. Subjects were asked to walk at their normal speed to the end of a straight 40 meter-long hospital corridor and back. A 20-second period of stabilized walking was used to calculate cycle frequency, stride symmetry, and stride regularity. Symmetry and regularity were each derived from an auto-correlation coefficient; to convert their distribution from nonnormal to normal, Fisher's Z transformation was applied to the auto coefficients for these two variables. Intraobserver reproducibility was evaluated by asking the same observer to test 16 controls on three separate occasions at two-day intervals and interobserver reproducibility by asking four different observers to each test four controls (Latin square). Specificity and sensitivity were determined by testing 139 controls and 63 patients. The 139 controls (70 women and 69 men) were divided into five age groups (third through seventh decades of life). The 63 patients had a noninflammatory musculoskeletal condition predominating on one side. ROC curves were used to determine the best cutoffs for separating normal from abnormal values. RESULTS: Neither intra- nor interobserver variability was significant (P > 0.05). Cycle frequency was significantly higher in female than in male controls (1.05 +/- 0.06 versus 0.98 +/- 0.05 cycles/s; P < 0.001). Neither symmetry nor regularity were influenced by gender in the controls; both variables were also unaffected by age, although nonsignificant decreases were found in the 61 to 70-year age group, which included only nine subjects. In the ROC curve analysis, the area under the curve was high for all three variables (frequency, 0.81 +/- 0.04; symmetry, 0.85 +/- 0.03; and regularity, 0.88 +/- 0.03), establishing that there was a good compromise between sensitivity and specificity. CONCLUSION: Our gait analysis method offers satisfactory reproducibility and is sufficiently sensitive and specific to be used by clinicians in the quantitative evaluation of gait abnormalities. PMID- 10526380 TI - Magnetic resonance imaging changes in periarticular soft tissues during flares of medial compartment knee osteoarthritis. Preliminary study in 10 patients. AB - OBJECTIVE: To quantify changes in magnetic resonance imaging signals from the deep and superficial capsulo-ligamentous planes and the medial collateral ligament in flares of knee osteoarthritis. PATIENTS AND METHODS: Preliminary prospective study of ten patients with medial compartment knee osteoarthritis meeting American College of Rheumatology criteria and associated with a Lequesne index of 5 or more. A grid was used to evaluate signal changes as compared to the opposite (asymptomatic) knee. Magnetic resonance images were read independently by two radiologists blinded to clinical data. RESULTS AND DISCUSSION: In all ten patients the capsulo-ligamentous planes and medial collateral ligament generated low signal on T1 images and high signal on T2 images. No significant changes were seen in the asymptomatic knee. The evaluation grid produced satisfactory interobserver agreement. CONCLUSION: Flares of medial compartment knee osteoarthritis are associated with changes in magnetic signal as compared to the contralateral asymptomatic osteoarthritic knee. The grid developed for this study could be used to evaluate the effects of treatment in a larger number of patients. PMID- 10526382 TI - Synovial membrane metaplasia to secondary lymphoid organs: role in the pathogenesis of auto-immune arthritis. PMID- 10526381 TI - Causes of osteoporosis in males. A review of 160 cases. AB - BACKGROUND: The discovery of osteoporosis in a male requires a careful search for a cause. OBJECTIVE: To evaluate etiologic factors in male osteoporosis. PATIENTS AND METHODS: Males admitted to our department for osteoporosis were included if they had a nontrauma-related vertebral or peripheral fracture and/or a spinal or femoral neck bone mineral density value 2.5 standard deviations or more below the mean in young subjects. The study was retrospective from 1990 to 1995 and prospective from 1996 to 1997. During the prospective part of the study, each subject underwent a standardized battery of laboratory tests including renal tubular function parameters. Causes identified during these two periods were compared. RESULTS: Of the 160 patients included in the study, 28.1% had idiopathic osteoporosis, 22.5% had alcoholic osteoporosis, 19.4% had glucocorticoid-induced osteoporosis, 12.5% had osteoporosis due to moderate idiopathic proximal tubule dysfunction, and 8.8% had senile osteoporosis. The proportion of patients with idiopathic osteoporosis was 30% (23/76) during the retrospective part of the study and 26% (21/84) during the prospective part (nonsignificant difference). Moderate idiopathic proximal tubule dysfunction was found in 2.6% (2/76) and 21.4% (18/84) of patients during these two parts of the study, respectively, a difference ascribable to the routine determination of tubule function parameters during the second part of the study. CONCLUSION: An exhaustive search for a cause decreases the proportion of male osteoporosis cases that remain idiopathic. In our study, only 28% of cases were classified as idiopathic, a term that probably indicates involvement of multiple interrelated factors. PMID- 10526383 TI - Suspected role of ofloxacin in a case of arthalgia, myalgia, and multiple tendinopathy. AB - A 53-year-old woman on ofloxacin developed myalgia, arthralgia, and tendinopathy. Her symptoms resolved after ofloxacin discontinuation. Although tendinopathy is a well-documented complication of quinolone therapy, there have been few reports of muscle symptoms. Concomitant involvement of the tendons, muscles, and joints has been exceedingly rare. Inhaled glucocorticoid therapy and moderate hypothyroidism were probably precipitating factors in our patient. PMID- 10526384 TI - Acute calcific tendinitis in children. AB - Acute calcific tendinitis is uncommon in children. Clinical manifestations are similar to those in adults. The abrupt onset, functional impairment, and frequent presence of fever suggest an infection. Radiographic findings establish the diagnosis, obviating the need for further investigations. PMID- 10526386 TI - Dropped head syndrome. Three case-reports. AB - Dropped head syndrome is characterized by gradual forward sagging of the head due to weakness of the neck extensor muscles. We report three cases in elderly patients seen by rheumatologists at our institution. There was some evidence suggestive of a neurogenic process, whereas most reported cases of dropped head syndrome have been ascribed to myopathy. Dropped head syndrome can probably be produced by multiple causes. The close ties between dropped head syndrome and acquired camptocormia in adults are discussed. PMID- 10526385 TI - A case of intracranial dural arteriovenous fistula draining into the spinal medullary veins. AB - Intracranial dural arteriovenous fistulas draining into the spinal medullary veins (ICDAVFMs) are exceedingly rare lesions. Their diagnosis is difficult and is often made late. About twenty well documented cases have been published. We report a case in a 55-year-old woman who presented with persistent interscapular pain and neurological evidence of ascending myelopathy after therapy for cervicobrachial neuralgia. ICDAVFM should be considered by rheumatologists in patients with clinical and radiological findings suggestive of spinal cord disease, particularly if these findings indicate involvement of the medulla oblongata or cervical spinal cord. PMID- 10526387 TI - Successful lipid-complexed amphotericin B treatment of Candida arthritis in a lymphoma patient. AB - Fungal arthritis is uncommon but has been increasingly diagnosed over recent years, particularly in patients with immunodeficiency due for instance to hematological malignancies. Candida albicans is the most frequent causative agent, and the knee is the joint most often involved. Amphotericin B is the drug of choice, but is associated with significant toxicity. Recently developed lipid formulations of amphotericin B have been found as effective and less toxic than the conventional formulation. We report a new case of Candida arthritis that occurred after chemotherapy for nonHodgkin's lymphoma and was successfully treated with lipid-complexed amphotericin B. PMID- 10526388 TI - Septic hip arthritis after multiple injections into the joint of hyaluronate and glucocorticoid. PMID- 10526390 TI - [Year of the brain 1999]. PMID- 10526389 TI - Bilateral frozen shoulder at the same time in two brothers. PMID- 10526391 TI - What is new in movement disorders? AB - Movement disorders is a term applied for a heterogeneous group of diseases and syndromes sharing deficits of voluntary motor function or movement patterns. In clinical practice, the term movement disorders is usually employed to designate those syndromes and diseases that are linked to a pathology or dysfunction of cortico-basal ganglia circuits. The last years have witnessed a rapid expansion in our understanding of the etiological and pathophysiological factors underlying movement disorders such as Parkinson's disease or dystonia. The discovery of new gene mutations is bound to give rise to new insights into the molecular pathogenesis of movement disorders related to neurodegenerative processes. It is already becoming apparent that pathological protein aggregation may be a common link in the neuronal degeneration underlying such diverse entities as spinocerebellar ataxia, idiopathic torsion dystonia and Parkinson's disease. So far, these new findings have not been translated into new forms of symptomatic or preventive therapies. Nevertheless, symptomatic treatment of movement disorders, as evident in the field of Parkinson's disease, is one of the most rewarding and innovative areas of neurological therapy. PMID- 10526392 TI - [Current therapy of cerebrovascular infarct]. AB - In general, the acute treatment of ischemic stroke is intended to achieve three goals: a) reduce the volume of the infarct, b) prevent further embolism or thrombotic occlusion of vertebrobasilar or carotid arteries, and c) prevent or react quickly to complications. Methods to expedite clot lysis and restore circulation (thrombolysis) can limit the extent of brain injury and improve outcome. Further therapeutic aims are to optimize collateral perfusion in the penumbra--the area surrounding the core of the ischemic focus--and to reduce ischemia-related secondary injury mechanisms (neuroprotection). Patients at high risk of further embolism or those at risk of acute thrombotic occlusion of a major vessel may--under certain circumstances--be candidates for the application of high-dose heparin. PMID- 10526393 TI - What is new in degenerative dementia disorders? AB - Alzheimer's disease and other degenerative disorders--dementia with Lewy bodies, frontotemporal dementia, etc.--causing about 90% of dementias in advanced age, are a major health problem of increasing practical, scientific, and socio economic importance. Despite considerable progress in genetic, clinical and basic neurosciences, the aetiology and molecular mechanisms of these disorders are still unknown and their early diagnosis, due to lack of specific biomarkers, is still unsatisfactory. The epidemiology, risk factors, clinical and morphological diagnostic criteria, probable pathogenic factors, and molecular genetics of the major types of degenerative dementias are reviewed. Their management involves several pharmacologic, non-pharmacologic and psychosocial options. Modification of the disease by reducing known and presumable risk factors, cognitive enhancement with cholinomimetic drugs, and reduction of behavioural abnormalities with psychotropic drugs, together with informed community and private management are currently achievable goals that will serve to delay the progression of disease. In the future, these options will hopefully be replaced by more effective management strategies in order to improve the quality of life of both, patients and caregivers. PMID- 10526394 TI - [Current problems in epilepsy]. AB - Three new aspects of epilepsy are discussed: the mesiotemporal syndrome, vagus nerve stimulation, and epilepsy and driving fitness. In recent years mesiotemporal epilepsy has been recognised as the most frequent epileptic syndrome in adults. The main clinical features are febrile convulsions during childhood, followed by characteristic focal seizures in the second decade of life. The typical seizure is characterised by an aura, followed by loss of consciousness, with motor phenomena and automatisms followed by longer periods of postictal confusion. Atrophy of the hippocampus and sclerosis are observed in MRI. The syndrome is frequently drug resistant, however, 80% of the patients are free of seizure after surgical treatment. Vagus nerve stimulation is a new option in the treatment of patients with drug resistant epilepsy (partial seizures with or without secondary generalization, Lennox-Gastaut syndrome), especially when surgical intervention is not indicated. Worldwide a total of more than 4000 patients have been treated. More than 50% reduction in the frequency of seizures can be obtained in 35-40% of drug resistant patients. Complications are rare. Finally, the issue of driving fitness and epilepsy as well as provoked seizures are discussed. The current regulations and laws are taken into consideration and revised regulations for Austria are suggested. PMID- 10526395 TI - [New developments and perspectives of intensive neurology]. AB - In the last few years considerable advances have been made in the diagnosis and treatment of cerebrovascular diseases, neurotrauma, and infections or inflammatory diseases of the CNS. Although thrombolysis of medial cerebral artery infarction and craniotomy with dural replacement plastic in space-occupying medial cerebral artery infarction only concern a minority of "patients with acute cerebral infarction", these treatment strategies have now been accepted as justifiable methods in the acute management of vascular ischaemic cerebral events. In addition to invasive and non-invasive monitoring of intracranial pressure and perfusion pressure in acute craniocerebral injuries, the therapeutic possibilities include treatment with calcium antagonists in traumatic subarachnoid haemorrhage and early recognition of space-occupying hemorrhagic brain contusions, in particular those mainly concerning the frontal portion of the brain. In the last few years, remarkable progress has also been made in understanding the pathophysiology of intracranial processes in bacterial meningitis. In addition to consecutive monitoring and management strategies, it is now possible to make prognostic statements and to propose adjuvant treatment strategies based on pathophysiological variables. In the therapeutic management of most severe intensive neurological disorders (spontaneous subarachnoid haemorrhage, craniocerebral trauma, dexamethasone in bacterial meningitis, etc.), a series of carefully conducted prospective randomised double-blind studies are currently under way. Hence, further significant advancements in the understanding of pathophysiology, in diagnostic and prognostic measurements and, in particular, the application of causal and adjuvant treatment strategies, are anticipated in the near and nearest future. PMID- 10526396 TI - [Prevention of stroke]. AB - Stroke is an extremely frequent disorder and represents the third most frequent cause of death in industrial countries. It is associated with catastrophic sequelae not only for the patient but also for his family members, and imposes an enormous socioeconomic burden. The efficacy of treatment of modifiable risk factors in secondary prevention of stroke is beyond any doubt. Based on these experiences, a corresponding strategy for an efficient, economic, and low-risk primary prevention of stroke can be deduced, the efficacy of which again can not be seriously doubted. PMID- 10526397 TI - The immunopathogenesis of multiple sclerosis. A survey of recent advances and implications for future therapy. AB - The etiology of multiple sclerosis (MS) still remains elusive. However, the association of the disease with relevant genes, the characteristic white matter infiltrates, similarities with animal models, and the fact that MS can be treated with immunomodulatory and immunosuppressive therapies support the notion that this disorder is autoimmune in nature. During the last few years the knowledge about the mechanisms involved in the pathogenesis of MS increased rapidly. In this review, recent advances in our understanding about relevant pathomechanisms in inflammatory demyelinating diseases of the central nervous system will be discussed and the rational of some novel immunotherapies is explained in context of the current understanding of immunological principles. PMID- 10526398 TI - Optimization of adhesion mode atomic force microscopy resolves individual molecules in topography and adhesion. AB - The force sensor of an atomic force microscope (AFM) is sensitive enough to measure single molecular binding strengths by means of a force-distance curve. In order to combine high-force sensitivity with the spatial resolution of an AFM in topography mode, adhesion mode has been developed. Since this mode generates a force-distance curve for every pixel of an image, the measurement speed in liquid is limited by the viscous drag of the cantilever. We have equipped our adhesion mode AFM with a cantilever that has a low viscous drag in order to reach pixel frequencies of 65 Hz. Optimized filtering techniques combined with an auto-zero circuitry that reduces the drift in the deflection signal, limited high- and low frequency fluctuations in the height signal to 0.3 nm. This reduction of the height noise, in combination with a thermally stabilized AFM, allowed the visualization of individual molecules on mica with an image quality comparable to tapping mode. The lateral resolution in both the topography and the simultaneously recorded adhesion image are only limited by the size of the tip. Hardware and software position feedback systems allows individual molecules to be followed in time during more than 30 min with scan sizes down to 60 x 60 nm2. PMID- 10526399 TI - Salmonella landau as a live vaccine against Escherichia coli O157:H7 investigated in a mouse model of intestinal colonization. AB - The present study was performed to assess the potential of a humoral mucosal immune response directed against the O157 antigen of Escherichia coli O157:H7 to prevent intestinal colonization by the pathogen. To this end, mice were gavaged with inocula of Salmonella landau, a Salmonella strain that naturally expresses the O157 antigen. Salmonella landau was avirulent for mice. Despite this, mice exposed to S. landau developed high titres of serum and coproantibodies against the O157 antigen. These mice, compared with controls, demonstrated some ability to resist transient intestinal colonization by an oral inoculum of an isolate of E. coli O157:H7. These findings suggest that a local immune response directed against the O157 antigen might increase host resistance to this pathogen. PMID- 10526400 TI - Effect of subinhibitory concentrations of antimicrobial agents (quinolones and macrolide) on the production of verotoxin by enterohemorrhagic Escherichia coli O157:H7. AB - In Japan, antimicrobial agent therapy for patients with diarrhea due to enterovirulent organisms including enterohemorrhagic Escherichia coli (EHEC) is common, and norfloxacin (NFLX), fosfomycin, and kanamycin are recommended for EHEC treatment by the Japanese Ministry of Health and Welfare. The aim of this study was to analyze the effects of antimicrobial agents which have been used or recommended for the treatment of EHEC on the production of verotoxin (VT) in vitro. Subinhibitory concentrations of quinolones, NFLX, sparofloxacin (SPFX), and grepafloxacin (GPFX) markedly stimulated the productions of VT1 and VT2. The macrolide azithromycin (AZM), erythromycin (EM), and clarithromycin (CAM) did not stimulate the production of VT at a wide range of concentrations. These in vitro results indicate that when quinolones are prescribed for a patient infected with EHEC, the concentration of antimicrobial agents used in vivo and the susceptibility of the EHEC strains against quinolones should be taken into consideration. PMID- 10526401 TI - Survival of Cryptosporidium parvum oocysts in source separated human urine. AB - The survival of Cryptosporidium parvum in source separated urine was investigated as part of a broader study on microbial risks associated with the reuse of human urine for sustainable agriculture. A dye permeability assay and in vitro excystation were the primary methods used to assess viability. In the collected urine most of the nitrogen is present as ammonia and the pH is generally around 9. Parallel investigations were made in buffers to compare possible toxic effects of urine to actual pH effects. Oocysts in the untreated urine were inactivated below the detection limit (1/300) within 63 days. This inactivation rate was significantly higher (p < 0.01) than in urine adjusted to pH 5 or 7 according to the dye permeability assay. The corresponding difference between different pH values was not seen in buffers, suggesting that the antiprotozoan effect of urine was mediated by other factors besides pH. The Swedish practice of storing urine for six months before its use thus appears satisfactory for the inactivation of Cryptosporidium oocysts. PMID- 10526402 TI - Epitope mapping of monoclonal antibodies specific for the directly cross-linked mesodiaminopimelic acid peptidoglycan found in the anaerobic beer spoilage bacterium Pectinatus cerevisiiphilus. AB - Nineteen monoclonal antibodies (Mabs) were isolated based on reactivity with disrupted Pectinatus cerevisiiphilus cells. All of the Mabs reacted with cells from which the outer membrane had been stripped by incubation with sodium dodecyl sulphate, suggesting the peptidoglycan (PG) layer was involved in binding. Mab reactivity with purified PG confirmed this. Epitope mapping revealed the Mabs in total recognize four binding sites on the PG. Mabs specific for each of the four sites also bound strongly to disrupted Pectinatus frisingensis, Selenomonas lacticifix, Zymophilus paucivorans, and Zymophilus raffinosivorans cells, but weakly to disrupted Megasphaera cerevisiae cells. No antibody reactivity was seen with disrupted cells of 11 other species of Gram-negative bacteria. These results confirm that a common PG structure is used by several species of anaerobic Gram negative beer spoilage bacteria. These results also indicate that PG-specific Mabs can be used to rapidly detect a range of anaerobic Gram-negative beer spoilage bacteria, provided the bacterial outer membrane is first removed to allow antibody binding. PMID- 10526403 TI - Transfer RNA genes and their significance to codon usage in the Pseudomonas aeruginosa lamboid bacteriophage D3. AB - Using tRNAscan-SE and FAStRNA we have identified four tRNA genes in the delayed early region of the bacteriophage D3 genome (GenBank accession No. AF077308). These are specific for methionine (AUG), glycine (GGA), asparagine (AAC), and threonine (ACA). The D3 Thr- and Gly-tRNAs recognize codons, which are rarely used in Pseudomonas aeruginosa and presumably, influence the rate of translation of phage proteins. BLASTN searches revealed that the D3 tRNA genes have homology to tRNA genes from Gram-positive bacteria. Analysis of codon usage in the 91 ORFs discovered in D3 indicates patterns of codon usage reminiscent of Escherichia coli or P. aeruginosa. PMID- 10526404 TI - Development of a polymerase chain reaction diagnostic test for the detection of the biotrophic pathogen Plasmopara halstedii in sunflower. AB - The obligate parasitic fungus-like organism Plasmopara halstedii (Farl.) Berl. et De Toni, is the causal agent of downy mildew disease in sunflower (Helianthus annuus). New races of this economically important parasite are regularly detected throughout the world. In addition, fungicide-resistant isolates have been reported in Europe and North America. These observations of parasite evolution, as well as the risk of propagation of the disease by infected seeds, means that it is necessary to guarantee the absence of Plasmopara halstedii in seed shipments. We report here the development of a rapid assay that can be used to detect infection by Plasmopara halstedii in plant tissues. Based on the nucleotide sequence information obtained from one cloned random amplified polymorphic DNA fragment, specific oligonucleotides were designed and used as primers for in vitro DNA amplification by polymerase chain reaction. An amplification product was detected on agarose gel stained with ethidium bromide when DNA from various Plasmopara halstedii races was tested, whereas no amplified DNA was detected when DNA from other origins was tested, including DNA from the host plant. The sensitivity of the technique was evaluated. The assay successfully reveals the presence of Plasmopara halstedii in infected sunflower plants prior to sporulation. PMID- 10526405 TI - Biochemical and molecular basis of pesticide degradation by microorganisms. PMID- 10526406 TI - NMR exchange broadening arising from specific low affinity protein self association: analysis of nitrogen-15 nuclear relaxation for rat CD2 domain 1. AB - Nuclear spin relaxation monitored by heteronuclear NMR provides a useful method to probe the overall and internal molecular motion for biological macromolecules over a variety of time scales. Nitrogen-15 NMR relaxation parameters have been recorded for the N-terminal domain of the rat T-cell antigen CD2 (CD2d1) in a dilution series from 1.20 mM to 40 microM (pH 6.0, 25 degrees C). The data have been analysed within the framework of the model-free formalism of Lipari and Szabo to understand the molecular origin of severely enhanced transverse relaxation rates found for certain residues. These data revealed a strong dependence of the derived molecular correlation time tau c upon the CD2d1 protein concentration. Moreover, a number of amide NH resonances exhibited exchange broadening and chemical shifts both strongly dependent on protein concentration. These amide groups cluster on the major beta-sheet surface of CD2d1 that coincides with a major lattice contact in the X-ray structure of the intact ectodomain of rat CD2. The complete set of relaxation data fit well to an equilibrium monomer-dimer exchange model, yielding estimates of exchange rate constants (kON = 5000 M-1 s-1; kOFF = 7 s-1) and a dissociation constant (KD approximately 3-6 mM) that is consistent with the difficulty in detecting the weak interactions for this molecule by alternative biophysical methods. The self association of CD2d1 is essentially invariant to changes in buffer composition and ionic strength and the associated relaxation phenomena cannot be explained as a result of neglecting anisotropic rotational diffusion in the analysis. These observations highlight the necessity to consider low affinity protein self association interactions as a source of residue specific exchange phenomena in NMR spectra of macromolecular biomolecules, before the assignment of more elaborate intramolecular conformational mechanisms. PMID- 10526408 TI - Estimating the time scale of chemical exchange of proteins from measurements of transverse relaxation rates in solution. AB - Chemical (conformational) exchange on the ms--microsecond time scale is reliably identified by the observation of transverse relaxation rates, Rex, that depend upon the strength of the effective field (omega 1eff = gamma B1eff) used in spin lock or CPMG experiments. In order to determine if the exchange correlation time, tau ex, is the fast or slow limit, measurements of (i) signal line shape and (ii) temperature dependence of Rex have been commonly used in studies of stable, small molecules. However, these approaches are often not applicable to proteins, because sample stability and solubility, respectively, limit the temperature range and signal sensitivity of experiments. Herein we use a complex, but general, two-site exchange equation to show when the simple fast exchange equations for Rex are good approximations, in the case of proteins. We then present a simple empirical equation that approximately predicts Rex in all exchange regimes, and explains these results in a clear, straightforward manner. Finally we show how one can reliably determine whether tau ex is in the fast or slow exchange limit. PMID- 10526407 TI - Human replication protein A: global fold of the N-terminal RPA-70 domain reveals a basic cleft and flexible C-terminal linker. AB - Human Replication Protein A (hsRPA) is required for multiple cellular processes in DNA metabolism including DNA repair, replication and recombination. It binds single-stranded DNA with high affinity and interacts specifically with multiple proteins. hsRPA forms a heterotrimeric complex composed of 70-, 32- and 14-kDa subunits (henceforth RPA70, RPA32, and RPA14). The N-terminal 168 residues of RPA70 form a structurally distinct domain that stimulates DNA polymerase alpha activity, interacts with several transcriptional activators including tumor suppressor p53, and during the cell cycle it signals escape from the DNA damage induced G2/M checkpoint. We have solved the global fold of the fragment corresponding to this domain (RPA70 delta 169) and we find residues 8-108 of the N-terminal domain are structured. The remaining C-terminal residues are unstructured and may form a flexible linker to the DNA-binding domain of RPA70. The globular region forms a five-stranded anti-parallel beta-barrel. The ends of the barrel are capped by short helices. Two loops on one side of the barrel form a large basic cleft which is a likely site for binding the acidic motifs of transcriptional activators. Many lethal or conditional lethal yeast point mutants map to this cleft, whereas no mutations with severe phenotype have been found in the linker region. PMID- 10526409 TI - Resonance assignments, secondary structure and 15N relaxation data of the human transcriptional coactivator hMBF1 (57-148). AB - Multiprotein bridging factor 1 (MBF1) is a transcriptional coactivator that is thought to bridge between the TATA box-binding protein (TBP) and DNA binding regulatory factors, and is conserved from yeast to human. Human MBF1 (hMBF1) can bind to TBP and to the nuclear receptor Ad4BP, and is suggested to mediate Ad4BP dependent transcriptional activation. Here we report the resonance assignments and secondary structure of hMBF1 (57-148) that contains both TBP binding and activator binding residues. 15N relaxation data were also obtained. As a result, hMBF1 (57-148) was shown to consist of flexible N-terminal residues and a C terminal domain. The C-terminal domain contains four helices and a conserved C terminal region. PMID- 10526410 TI - 1H, 13C and 15N resonance assignments of the C-terminal domain of MutY: an adenine glycosylase active on G:A mismatches. PMID- 10526411 TI - Partial cross protection against Ichthyophthirius multifiliis in Gyrodactylus derjavini immunized rainbow trout. AB - Partial cross protection against a skin-parasitic ciliate has been recorded in rainbow trout previously immunized with an ectoparasitic platyhelminth. The susceptibility to infection by Ichthyophthirius multifiliis differed significantly between naive and Gyrodactylus derjavini immunized rainbow trout. Fish partly immune to the ectoparasitic monogenean G. derjavini became less infected and experienced lower mortality than naive fish when exposed to I. multifiliis infections. In vitro studies on immobilization of theronts using decomplemented (heat-inactivated) serum from G. derjavini immune or non-immune hosts showed no immobilization. However, untreated serum from both immune and non immune fish containing intact complement immobilized theronts (titre 128-256). In addition, non-specific priming of the host response with interleukin (IL-1), bacterial lipopolysaccharide (LPS), concanavalin A (Con A) or mannan did confer a partial resistance to I. multifiliis infection. This will suggest that non specific factors including complement could be partly responsible for the host response against infections with this ciliate. PMID- 10526412 TI - The parasite community infecting flounders, Platichthys flesus, in the tidal Thames. AB - The composition of the parasite fauna of the flounder, Platichthys flesus, retrieved from two locations in the tidal Thames is described in detail for the first time. The combined parasite species list of the flounders from Lots Road in the upper tideway and West Thurrock in the middle tideway consisted of one protozoan (Glugea stephani), one monogenean (Gyrodactylus sp.), four larval digeneans (Cryptocotyle concava, Timoniella imbutiforme, T. praeterita, and Labratrema minimus), five adult digeneans (Derogenes varicus, Lecithaster gibbosus, Podocotyle sp., Plagioporus varius, and Zoogonoides viviparus), one larval cestode (unidentified tetraphyllidean), one or possibly more larval nematodes (unidentified) plus five adult nematodes (Capillaria sp., Cucullanus heterochrous, C. minutus, Contracaecum sp. and Goezia sp.), two acanthocephalans (Pomphorhynchus laevis and Acanthocephalus anguillae), three copepods (Lepeophtheirus pectoralis, Acanthochondria sp. and Lernaeocera branchialis), and one mollusc (unidentified glochidia). The overall parasite community of flounders from Lots Road and West Thurrock were compared in terms of species richness and diversity. The parasite community in flounders from the former location in the upper tideway was found to be less diverse than that of its counterpart at West Thurrock in the middle estuary. The component community of Lots Road flounders was dominated by the acanthocephalan Pomphorhynchus laevis. PMID- 10526413 TI - Schistosome cercariae as the causative agent of swimmer's itch in Iceland. AB - During late summer in 1995 to 1997, repeated outbreaks of maculopapular skin eruptions were noted on the legs of children after wading in the pond in the Family Park in Laugardalur, Reykjavik, Iceland. Clinical symptoms developing on the legs resembled those of cercarial dermatitis. An examination of Lymnaea peregra snails from this pond and from the adjacent Lake Tjornin resulted in detection of previously undescribed schistosome cercariae. This is the first report of schistosomes in Iceland and also the most northern occurrence of these parasites in Europe. PMID- 10526414 TI - Influence of host weight, sex and reproductive status on helminth parasites of the wild rabbit, Oryctolagus cuniculus, in Navarra, Spain. AB - A study was carried out in Navarra (northern Spain) on the influence of the weight, sex and reproductive status (lactant, pregnant or lactant + pregnant females and testicular weight for males) of the wild rabbit (Oryctolagus cuniculus) on two cestodes species: Andrya cuniculi and Mosgovoyia ctenoides and four intestinal nematodes: Graphidium strigosum, Trichostrongylus retortaeformis, Nematodiroides zembrae and Dermatoxys hispaniensis. A significantly higher prevalence of A. cuniculi was detected in lactant + pregnant females compared with non-breeding females. Trichostrongylus retortaeformis and N. zembrae showed a significantly higher mean intensity in lactant and lactant + pregnant females than in non-reproductive females. Trichostrongylus retortaeformis presented a higher mean intensity in females than in males, and the mean intensity of the same parasite species was significantly lower in active and inactive males compared with lactant and lactant + pregnant females. There were no significant differences between sexes in the prevalence of helminth parasites. No significant correlation was detected between host weight and the intensity (of infection) of helminths studied. No significant differences in the prevalence and mean intensity of the two cestode species were observed in the three weight categories studied (kittens, juveniles and adults). The prevalence of G. strigosum and mean intensity of T. retortaeformis were significantly higher in older heavier animals than in juveniles. PMID- 10526415 TI - An enhanced humoral immune response against the swimbladder nematode, Anguillicola crassus, in the Japanese eel, Anguilla japonica, compared with the European eel, A. anguilla. AB - The humoral immune response in the two eel species, Anguilla japonica and Anguilla anguilla against two fractions of antigens in Anguillicola crassus were studied. Within species, both eel species showed significantly elevated titres compared with controls when immunized with antigens from Anguillicola crassus. In interspecific comparison, Anguilla japonica showed significantly elevated titres in comparison with Anguilla anguilla. Immunization of Anguilla anguilla caused a significantly decrease in the plasma levels of protein in comparison with control fish and all groups of Anguilla japonica. In contrast, Anguilla japonica showed significantly lower plasma levels of Ig in all groups compared with Anguilla anguilla. The different susceptibilities to Anguillicola crassus between the natural host, Anguilla japonica, and the naive, Anguilla anguilla, is partly due to differences in the ability of the two eel species to mount a humoral immune response. PMID- 10526416 TI - Predilection sites of Trichinella spiralis larvae in naturally infected horses. AB - A total of 120 muscle tissues from three horses naturally infected with Trichinella spiralis were examined. The head was the most infected site. In particular, the muscles harbouring the highest number of larvae were: musculus buccinator (12, 411 and 1183 larvae g-1), the tongue (11, 615 and 1749 larvae g 1), m. levator labii maxillaris (17,582 and 1676 larvae g-1), and the masseter (4.9, 289 and 821 larvae g-1). Compared with the diaphragm, the number of larvae per gram was from 3.5 to 6.8 times higher in the tongue, from 3.5 to 6.5 higher in m. levator labii maxillaris, and from 2.5 to 4.6 higher in m. buccinator. Of the examined muscles, the diaphragm had from the 6th to the 15th highest level of infection (3.1, 166 and 256 larvae g-1). Published data from experimentally infected horses confirm these results, suggesting that efforts to detect predilection sites should focus on the head muscles. PMID- 10526417 TI - Humoral immune responses induced by Gymnorhynchus gigas extracts in BALB/c mice. AB - The aim of this study was to determine if the plerocercoid larvae of Gymnorhynchus gigas, a common cestode of the ray's bream (Brama raii), possess antigenic compounds potentially capable of provoking anaphylactic episodes. A murine experimental model, using BALB/c mice, was developed to study the humoral immune response induced by G. gigas extracts. A highly specific humoral immune response was detected and cross-reactions were not observed between parasite and host antigens. The presence of IgM and IgG3 levels suggest the presence of thymus independent antigens in the parasitic extract. The IgG antibody class showed the highest levels, with the IgG1 the predominant subclass. These IgG1 levels are in accordance with the supposed presence of a type I allergic reaction after the ingestion of G. gigas plerocercoids parasitizing fish, as well as inducing anaphylaxia in fish. These results indicate that somatic products released from ingested larvae of G. gigas could induce the development of a Th2 response capable of causing allergic disorders. PMID- 10526418 TI - Influence of infection intensity on predilection sites in swine trichinellosis. AB - The muscular distribution of Trichinella spiralis or T. britovi was studied by digestion in 59 experimentally infected pigs and seven wild boars. Crus muscle was the predilection site in 89.3% of 28 heavily infected swine with 146-3634 larvae per gram (lpg), but in 51.6% of middle to light infections (0.005-59 lpg) the basis of the tongue showed higher larval densities than the crus muscle. The basis of the tongue was also the predilection site in 71.4% of wild boars. Highest counts in other muscles were found only in lightly infected pigs. The influence of intensity of infection, host species, and Trichinella species on muscle distribution is discussed. PMID- 10526419 TI - Biochemical profiles of hydatid cyst fluids of Echinococcus granulosus of human and animal origin in Libya. AB - A comparative study on the biochemical parameters in hydatid cyst fluids of sheep, goats, camels, cattle and human cystic forms of Echinococcus granulosus has been made in Libya. Quantitative variations in the levels of sodium, potassium, calcium, cholesterol, glucose, urea, creatinine and gamma glutamyl transpeptidase (gamma GT) were found in the cystic fluids of different host origins although these differences were statistically insignificant compared with the hydatid fluids of sheep. However, the concentration of triglycerides and proteins were significantly elevated in the cyst fluids of sheep compared with the other fluids studied. Similarities in the biochemical composition of different hydatid cyst fluids suggest the existence of sheep strains of E. granulosus in human and other domestic animal intermediate hosts in Libya. PMID- 10526420 TI - Lung nematodes of chamois, Rupicapra rupicapra tatrica, from the Tatra National Park, Slovakia. AB - A larvoscopic examination of faeces collected from localities inhibited by chamois in the Tatra National Park (TANAP) in 1997 demonstrated the presence of the lung nematodes Muellerius spp. (likely to be M. tenuispiculatus and M. capillaris) and Neostrongylus linearis. The overall prevalence of lung nematodes in chamois herds in TANAP was 48.4% with prevalences of 45.6% and 11.9% for Muellerius spp. and N. linearis, respectively. No significant differences in lung nematode prevalences were observed in the biotopes of TANAP with prevalence values of 44.9% being recorded in the High Tatras and 58.5% in the Belianske Tatras. Individual species were in equal proportion in both biotopes, although N. linearis was significantly less prevalent (11.2-13.8%). The prevalence of lung nematodes in the High Tatras varied from 25.0 to 84.2% within individual localities, while in the Belianske Tatras it was more proportionate (50.0-85.7%). In the High Tatras, the prevalence of lung nematodes in the chamois herds peaked during August, declining to its lowest in October. A similar prevalence was also recorded for Muellerius species, while the minimum prevalence of N. linearis was found in July. In the Belianske Tatras, the prevalence of lung nematodes including both species of Muellerius peaked in July and gradually decreased until October. On the other hand, N. linearis was most prevalent in October. The mean L1 count per gram faeces was low (7.6 +/- 13.2 larvae g-1). PMID- 10526421 TI - Polymorphism in the Dirofilaria immitis immunodominant antigen gene. AB - Dg2, a gene encoding a 34 kDa immunodominant antigen of Dirofilaria immitis was cloned and demonstrated to be specifically expressed in the larval stage. In this study, a newly constructed genomic DNA library was screened by hybridization with Dg2. One of the resulting positive clones was similar to Dg2 in the structure of its exonic regions but different in number, position, size and sequence of introns. This was designated DgK. Full-length cDNA was isolated using the rapid amplification of cDNA ends (RACE) method to study the transcript corresponding to DgK. Sequence analysis revealed that the mRNA corresponding to DgK is trans spliced during post-transcriptional processing because the 5' end of the amplified cDNA contains seven nucleotides of the nematode-spliced leader (SL) sequence. PMID- 10526422 TI - Recognition of deglycosylated larval proteins of Gnathostoma spinigerum by a monoclonal antibody and human gnathostomiasis antiserum. AB - The study on the recognition of 35S-labelled somatic antigens of Gnathostoma spinigerum advanced third-stage larva (aL3) has revealed that the mAb GN6/24 immunoprecipitated 26- and 24-kDa proteins from the undigested and N-glycosidase F-digested larval extracts, respectively. The recognition of the deglycosylated form of the glycoprotein indicated that the mAb reacted with the peptide epitope on the 26-kDa protein. Human gnathostomiasis antiserum immunoprecipitated most of the N-glycosidase F-digested larval proteins including the deglycosylated 26-kDa protein. PMID- 10526423 TI - Schistosoma japonicum infection in pregnant mice. AB - Ten 1-week and ten 2-weeks pregnant female NMRI mice were experimentally exposed to 70 Schistosoma japonicum cercariae. Ten littermice from each group were examined for worms by perfusion 4, 6 and 8 weeks post infection. Although the mothers (n = 15) were found infected with 15.5 +/- 13.4 worms at perfusion 6 and 7 weeks post infection, no worms were found in any of the examined littermice, as well as no detection of faecal or tissue eggs. Litter sizes did not differ from control groups and all littermice were healthy. The present study therefore suggests that congenital infection with S. japonicum does not occur in percutaneously infected mice and that infection of the mother during pregnancy does not seem to affect the offspring. PMID- 10526424 TI - Bacillus sphaericus interferes with the development of Brugia malayi in Aedes aegypti. AB - Aedes aegypti (black-eyed Liverpool strain) were exposed to a sublethal dose (LD25) of Bacillus sphaericus and were fed to Mastomys coucha infected with Brugia malayi. The development of the filarial parasite was found to be arrested mostly at the second larval stage. The infection (P < 0.05), infectivity rates (P < 0.001) and L3 load (P < 0.001) were found to be reduced significantly in the treated group. PMID- 10526425 TI - Update on fibromyalgia syndrome. PMID- 10526427 TI - Quality of life in patients with chronic myeloid leukemia after unrelated donor bone marrow transplantation. AB - A descriptive study was designed to assess the quality of life and quantify potential long-term physical and psychosocial problems resulting from unrelated donor bone marrow transplantation. A sample of 28 survivors of an average 41.2 months post-bone marrow transplantation showed that quality of life was good to excellent in most subjects, with a small fraction having impaired physical or psychosocial functioning. Psychosocial adjustment was most impaired in domains of life related to sexual relationships, vocational adjustment, social adjustment, and psychological distress. Females and older adults reported significantly higher dysfunction compared with males and younger subjects. Passage of time since bone marrow transplantation was unrelated to psychosocial improvement. Fatigue was the main symptom interfering with daily life in most (78.6%). Regression analysis showed that development of fatigue in our sample was mainly the result of the combined effects of anxiety, presence of pain, and infections (R2 = 0.75, p < .001), with weight loss showing a trend in relation to fatigue. Results indicate that unrelated donor bone marrow transplantation long-term survivors have an acceptable degree of quality of life, but rehabilitative services still need to be used for those who exhibit difficulties with their daily life. PMID- 10526426 TI - Spiraling out of control: one case of pathologic anxiety as a response to a genetic risk of cancer. AB - The threat of cancer can result in an existential crisis characterized by feelings of uncertainty and fear. Anxiety, the most common response to the threat of cancer, may be expressed in ways as varied as individual personalities and circumstances. It is a normal response to the threat of cancer, but in some it may deteriorate to pathologic anxiety and manifest somatic or avoidant patterns. Members of families diagnosed with genetic mutations that predispose to cancer are unique in that they experience a complex chain of life events. People affected with genetic mutations that increase the risk for the development of cancer may be at greater risk of manifesting abnormal anxiety. Little research exists that can guide the health professional in meeting the needs of these individuals, which leads health practitioners to approach their needs on the basis of combined theoretical assumptions about the needs of people with cancer, people who have family members with cancer, and people with recurrent cancer. Some factors may be assessed by the health professional as an aid in identifying an individual at increased risk of developing a psychopathology. These factors include age and developmental level, existence of a previous psychologic disorder, and family integrity. Advanced practice nurses may effectively intervene in the care of these patients by (a) accurately assessing the risk for and extent of the anxiety reaction in individuals and family members; (b) developing management plans that include ongoing support, education, psychotherapy, and pharmacotherapeutics for the individual; and (c) support and psychotherapy for the family. In this article, the pathologic anxiety experienced by one adolescent girl diagnosed with a genetic mutation that caused multiple endocrine neoplasia 2a is addressed, along with the treatment of her avoidant anxiety disorder--trichotillomania. PMID- 10526428 TI - The impact of hospice inpatient care on the quality of life of patients terminally ill with cancer. AB - This study explored the expectations and experiences of patients with terminal cancer in a hospice inpatient environment in an attempt to evaluate their quality of life and the impact of the care and services provided. A total of 52 patients terminally ill with cancer from 11 hospice units in Hong Kong participated in the study. Data were collected from patients by devising a Hospice Care Performance Inventory (HCPI), which was an interview schedule consisting of 25 items. The HCPI was developed after a review of the literature on the quality of life experienced by patients with advanced cancer and the aims of hospice units in Hong Kong. Each item was rated by the patient on a Likert scale in terms of its importance and the perceived effectiveness of the care provided. The study identified six issues in which expectations did not seem to match effectiveness. These issues indicated areas in which improvement could be attempted to enhance the quality of life for the patients. The most important was maximizing self-care and mobility. Two issues were identified in which effectiveness was high and importance to the patient relatively low. One of these issues was pain management, and the other was spiritual care. PMID- 10526429 TI - Cancer prevention education in developing countries: toward a model for nurse educators. AB - The National Cancer Institute, United States of America, funded a series of continuing education courses in cancer prevention between 1986 and 1994 for nurses from developing countries. The purpose of this program was to stimulate interest and facilitate an increase in the participants' knowledge of primary and secondary cancer prevention. The long-term objectives were to increase the number of nurses, internationally, prepared to engage in the prevention and the early detection of cancer in their countries, to expand the international cancer nursing network, and to have these nurses ultimately play a role in reducing the incidence of cancer in developing countries. More than 50 nations were represented. Participants were chosen for their demonstrated ability to influence nursing education and practice in their country. They completed a demographic data sheet, an attitude inventory, a program evaluation and pre- and postconference activities surveys. Before and after attending the conference, participants were asked to identify anticipated problems and obstacles to their goal achievement. These problems included a lack of screening facilities and a lack of primary prevention services. Although numerous differences existed in their education, experience, and personal attributes, the participants voiced common problems with cancer prevention programs. Results from the postconference survey showed a substantial increase in cancer-related activities conducted by the participants. Activities included an increase in cancer content in nursing education programs, an increase in public and professional presentations on cancer prevention, and improvement in the delivery of cancer care. PMID- 10526430 TI - Communication in cancer care: what science can and cannot teach us. AB - Although research in the cancer communication field has produced a body of information applicable to clinical practice, there is little evidence that the new knowledge has led to any significant improvements in the health care communication experiences of patients and their families. In this article, an analysis of the existing research-based knowledge provides a basis for critical analysis of the gaps and limitations within it. Insights from the current literature are contrasted with interpretations deriving from consumer-perspective research by this author and others. The discontinuity between the problems consumers identify and the issues that attract research attention is examined in the context of the orientation to science that drives much of the research. The case is made for challenging traditional notions of what counts as evidence in developing education and practice standards for cancer care. PMID- 10526431 TI - Women's approaches to decision making about mammography. AB - Health professionals have an obligation to understand women's decision making about mammography and to advocate for their active participation in health care decision making. Although mammography is a major screening measure for the second largest cancer killer of women, only approximately half of women older than age 50 years, and fewer older than age 70 years, undergo mammography in accordance with American Cancer Society (ACS) guidelines. Therefore, the purpose of this study was to identify women's overall decision-making approaches when considering mammography. Subjects were a purposive, convenience sample of 50 women in the community who had made a decision about mammography; they included those who chose to have mammograms and those who decided not to have mammograms according to the pre-1997 ACS guidelines. Subjects participated in audiotaped interviews. Results indicated that women approached the mammography decision differently, regardless of the decision they made. Three overall decision-making approaches to addressing risk factors, issues about mammography, or other factors before their decision were evident. The approaches were (1) thoughtful consideration; (2) cursory consideration; and (3) little or no consideration. Each approach has implications for nurses who assist women in making decisions about mammography. PMID- 10526432 TI - Breast cancer patients' out-of-pocket expenses. AB - Patients with cancer soon discover that there is much more to the cost of treatment than hospital and physician bills. Out-of-packet expenses for transportation, food supplements, over-the-counter medications, distractions, telephone bills, insurance premiums--at a time when employment may be out of the question--can be a significant drain on family finances. Women with regional breast cancer reported their estimates of out-of-pocket expenses incurred during 1 month of outpatient chemotherapy. All women in the study were covered by some form of insurance. Mean monthly out-of-pocket costs were $360 (SD = $346), and ranged from $36 to $1224. Additional costs for wigs, special events, gifts, and alternative treatment incurred since diagnosis, ranged from $20 to $3700. These costs excluded expenses for health providers. Very low and very high expenditures may indicate risks concealed from providers that may have subsequent impact on long-term capacity to sustain treatment. Nurses can help families anticipate expenses and prioritize referrals to community agencies. PMID- 10526434 TI - A message to my "live & then give" partners. PMID- 10526433 TI - Lyme disease. PMID- 10526435 TI - Initiative no. 9: for and against term limits. PMID- 10526436 TI - [Cathepsin A activity in the aqueous humor in patients with cataract, absolute glaucoma and intraocular tumors]. AB - PURPOSE: To evaluate cathepsin A activity in the aqueous humor of patients with cataract, absolute glaucoma and intraocular tumors. MATERIAL AND METHODS: The studies were performed on human aqueous humor taken from anterior chamber of eye balls of patients operated because of cataract, absolute glaucoma and intraocular tumors. Cathepsin A activity was determined by the ninhydrin method with synthetic substrate (N-Cbz-Phe-Ala) at its optimum pH 5.0. RESULTS: In the human aqueous humor of the eye with cataract cathepsin A activity was more than three times higher than in the eye with choroid tumors and absolute glaucoma. No differences of enzyme activity in aqueous humor between patients with glaucoma and intraocular tumors were found. CONCLUSION: The increasing proteolytic activity of cathepsin A in aqueous humor of patients with cataract suggests its importance in cataract pathogenesis. This implies that cathepsin A is involved in development of lens opacity and is found in the aqueous humor due to diffusion from cataractous lens in which the proteolytic process prevails. PMID- 10526437 TI - [Cathepsin A activity in the vitreous body of patients with absolute glaucoma and intraocular tumors]. AB - PURPOSE: To evaluate cathepsin A activity in the vitreous body of patients with absolute glaucoma and intraocular tumors. MATERIAL AND METHODS: The studies were performed on human vitreous body taken from eye balls which were enucleated because of absolute glaucoma (18 eyes) and intraocular tumors (14 eyes). Cathepsin A activity was determined by the ninhydrin method with synthetic substrate (N-Cbz-Phe-Ala) at its optimum pH 5.0. RESULTS: Cathepsin A activity in the human vitreous body in absolute glaucoma was twice as high as in intraocular tumors. CONCLUSION: Our results suggest that cathepsin A may participate in the pathogenesis of absolute glaucoma and that proteolysis may play a significant role in local destruction of the retina and the optic nerve. PMID- 10526439 TI - [Pars plana vitrectomy in the treatment of endophthalmitis]. AB - The aim of the study was to present and assess the results obtained after performing vitrectomy in patients with endophthalmitis after cataract surgery and injuries of the eyeball. MATERIAL: The study material comprised 22 patients: 16 patients with endophthalmitis after cataract surgery, 4 after eyeball injuries and 2 after fistula operations. EFFECTS: After vitrectomy we obtained in most patients an improvement of visual acuity and reduction of inflammatory symptoms. 10 days after operation in half of the patients the visual acuity was from 2/50 to 5/50, in 27.2% from 5/30 to 5/10. PMID- 10526438 TI - Can the functional outcome in macular hole surgery be improved by internal limiting membrane maculorhexis? PMID- 10526440 TI - [Pars plana vitrectomy in the treatment of endophthalmitis]. AB - PURPOSE: To assess the effectiveness of vitrectomy in the treatment of patients with endophthalmitis. MATERIAL AND METHODS: Ten patients with endophthalmitis were qualified for treatment with vitrectomy via pars plana of a cilliary body. Endophthalmitis was diagnosed as resulting from previous: ECCE + I in 3 patients, penetrating bulb injury in 3 patients, penetrating keratoplasty in 1 patient; 3 other patients suffered from endogenous endophthalmitis. In all patients prior to surgical procedure samples of the vitreous were collected for bacteriology and mycology tests. RESULTS: In 9 cases the surgery was followed by intravitreal administration of 1 or 2 antibiotics. In 1 case an antibiotic was added to the infusion solution. In all patients the following data were analyzed together with the result of a surgical procedure: cause of endophthalmitis, visual acquity directly before and after the procedure and over the follow-up period (from 6 months to 3 years), time delay between the onset of endophthalmitis and vitrectomy performed. CONCLUSION: Vitrectomy performed via pars plana of a cilliary body is an effective method of treatment in cases of endophthalmitis. In patients with endophthalmitis early performed vitrectomy, followed by adequate adjuvant pharmacotherapy determine the chances for good postoperative prognosis. PMID- 10526441 TI - [Scanning laser tomography in the diagnosis of juvenile glaucoma]. AB - The aim of our research was to evaluate the optic nerve head parameters of children with juvenile glaucoma with the scanning laser ophthalmoscope from Laser Diagnostic Technologies Inc. The material consisted of 46 eyes of 25 children aged from 7 to 18 years. There were 18 glaucomatous eyes (I group) and 28 eyes without glaucoma (II group) as a control group. The three examinations were repeated every three months. Mean values of Vol. B in the glaucomatous group were -0.327 +/- 0.098, first visit; -0.390 +/- 0.130, second visit; 0.344 +/- 0.096 mm3, third visit. These values in the control group were--0.234 +/- 0.73; -0.224 +/- 0.071; -0.252 +/- 0.069 mm3. The difference was statistically significant. Mean values of Volume A were significantly lower in the I group: 0.216 +/- 0.034, 0.207 +/- 0.033, 0.203 +/- 0.030 mm3 than in the II group: 0.269 +/- 0.039, 0.283 +/- 0.032, 0.271 +/- 0.042 mm3. CD ratio was higher in the glaucomatous eyes: 0.447 +/- 0.080; 0.491 +/- 0.069; 0.484 +/- 0.074 in comparison with the control group: 0.376 +/- 0.060; 0.368 +/- 0.067; 0.399 +/- 0.056. It was significant difference. More changes of biomorphometric parameters of the optic nerve head were observed in the glaucomatous group during follow-up period. The role of laser scanning ophthalmoscopic tomography is very important in the diagnosis of early forms of juvenile glaucoma in the youth. PMID- 10526442 TI - [Corneal endothelial cell density and corneal thickness after laser trabeculoplasty]. AB - PURPOSE: The evaluation of the influence of laser trabeculoplasty on the corneal thickness and central endothelial cell density. MATERIAL AND METHODS: Corneal thickness and central endothelial cell density were measured before and 3 months after laser treatment. Patients were divided into 2 groups with different energy applied: 550-800 mW and 850-1100 mW. RESULTS: Differences of corneal thickness and central endothelial cell density were not statistically significant neither within nor between the groups. CONCLUSION: Laser trabeculoplasty performed with energy up to 1100 mW has no influence on corneal thickness and central endothelial cell density. PMID- 10526443 TI - [Fluorescein angiography in children and adolescents with type I diabetes mellitus]. AB - PURPOSE: The evaluation of changes in ophthalmoscopic examination and fluorescein angiography in children and adolescents with insulin dependent diabetes mellitus. MATERIAL AND METHODS: 100 patients with type I diabetes mellitus aged 9.3-21.5 years (15.76 +/- 2.69), with diabetes duration of 1-13.5 years (6.57 +/- 2.52) were examined. RESULTS: Retinal changes in ophthalmoscopic examination were observed in 12 cases. Fluorescein angiography allowed to detect diabetic retinopathy in 29 patients. There was no correlation between the incidence of retinopathy and the age of patients. Vascular abnormalities were related to the duration of diabetes. Retinopathy was not found in children < 10 years of age. PMID- 10526444 TI - [The influence of treatment of progressive edematous infiltrative ophthalmopathy on intraocular pressure]. AB - PURPOSE: To evaluate the influence of systemic steroid therapy and retrobulbar irradiation on intraocular pressure (IOP) in patients with infiltrative- oedematous Graves' ophthalmopathy. MATERIAL AND METHODS: We examined 76 patients divided into 3 groups: I--treated by irradiation only (15 patients), II--treated by irradiation and oral prednisone therapy (26 patients), III--treated by irradiation and intravenous methylprednisolone pulse therapy (35 cases). All patients underwent full ophthalmological examination (including IOP measurement, perimetry and gonioscopy) before, during, immediately after and 2-20 months after treatment. RESULTS: Increased IOP (21-31 mm Hg) was observed in 54 patients (71%) before treatment. The iridocorneal angle was open in all eyes. Changes in perimetry were not characteristic for glaucoma. IOP was higher in patients with more severe ophthalmopathy. We recorded transient increase of IOP during treatment in only 3 patients. Increased IOP immediately after therapy was observed in 16 patients with severe symptoms and signs of ophthalmopathy: in group I--4/15 (27%), in group II--4/26 (15%), in group III--8/35 (23%). Higher IOP was recorded in 10 patients two to twenty months after completion of treatment: from group I--4/15 (27%), from group II--1/26 (4%) and from group III- 5/35 (14%). In 6 of these 10 persons we observed recurrence of ophthalmopathy, in 4 patients higher IOP was the only deviation, they needed local therapy. The mean values of IOP were lower in patients treated by steroid therapy in comparison to patients treated by irradiation only. The most rapid improvement of clinical status was observed in patients treated by methylprednisolone pulse therapy. CONCLUSIONS: The increase of IOP in patients with Graves' ophthalmopathy correlates with severity and duration of eye disease. Systemic steroid therapy is more efficient in reduction of IOP than irradiation of the retrobulbar tissue. Our results suggest that combined therapy is a preferable method of treatment of progressive ophthalmopathy, including cases with increased intraocular pressure. PMID- 10526445 TI - [Ocular manifestations in sarcoidosis]. AB - PURPOSE: The ophthalmic examination of patients with diagnosed systemic sarcoidosis. MATERIAL AND METHODS: 33 patients (17 women, 16 men), aged 22-60 years had ophthalmic examination (visual acuity, anterior and posterior segment evaluation, applanation tonometry). In 8 patients repeated examination was performed (duration of observation: 2-31 months). RESULTS: In 27 patients no characteristic features of ocular sarcoidosis were found. In 6 persons (18.2%) variety of ocular lesions was recognized (nodular infiltrations of the eyelids, chronic uveal inflammation, signs of anterior and posterior uveitis in the past, optic disc oedema). In 3 cases ocular lesions preceded the signs of systemic sarcoidosis. This emphasizes the importance of the routine ophthalmic examination of patients with suspected or proven sarcoidosis. PMID- 10526446 TI - [Effect of vaccination programs on the incidence of idiopathic optic neuritis]. AB - To evaluate the influence of vaccination programs on the incidence of idiopathic optic neuritis. MATERIAL: 359 questionnaired patients with idiopathic optic neuritis hospitalized in the Department of Ophthalmology in Bydgoszcz and in the province of Bydgoszcz in the period from 1978 to 1997. Control group included 264 persons with no history of optic neuritis. RESULTS: Vaccinated persons were significantly (p = 0.01) rarely affected by this disease. CONCLUSION: The study revealed the positive correlation between lack of vaccination (according to vaccination program) and the incidence of idiopathic optic neuritis. PMID- 10526447 TI - [Bilateral retinal detachment in toxemia of pregnancy]. AB - In this article, we present two cases of bilateral retinal detachment in toxemia of pregnancy. The first case describes 31-year old patient admitted to hospital due to toxemia of pregnancy in the 38th week of the second pregnancy. Medical examination following admission showed blood pressure 180/130 mm Hg oedema of face and legs. Accessory investigations displayed proteinuria, leukocyturia, bacteriuria. At the fundus of eye the features of initial retinopatia hypertonica were discovered. On the fifth day of the patient's hospitalization because of the worsening of general health condition and sudden blood pressure increase, the pregnancy was delivered by cesarean section. Some hours after awakening the patient reported the loss of vision of both eyes. Ophthalmological examination showed the occurrence of retinal detachment. Tearing wasn't found. When the protein deficiency was supplemented and antiinflammatory and oedema--reducing treatment was administered the blood pleasure stabilized retina attached itself and visual functions returned. In the second case 22-year old patient was admitted to hospital at term of labour with blood pressure 150/90 mm Hg and a little proteinuria. On the second day of hospitalization the pregnancy was delivered through natural passages. A few hours after the delivery the patient reported indistinct vision and blurring of the image. Through ophthalmological examination retinal detachment was stated. Tearing was not found. Thanks to bed regime and resorbing treatment total retinal attachment was gained and visual functions returned. This case suggests that retinal detachment of pregnant woman does not have to be proceeded by symptoms of toxemia of pregnancy and the period of delivery may accelerate and release mechanisms damaging choriocapillaries, which causes the flow of liquid from vessels of chorioidea to subretinal space. PMID- 10526448 TI - [A case report of interferon-associated retinopathy]. AB - The authors describe signs and symptoms of interferon-associated retinopathy in patient with hepatitis C virus, nephropathy and diabetes mellitus. Necessity to assess visual system before and after therapy was emphasised. PMID- 10526449 TI - [Secondary glaucoma in the course of arterio-cavernous fistula: a case report]. AB - PURPOSE: To present a case of 59-year-old woman with the symptoms of redness, proptosis of her left eye and temporary bruit in her head. METHODS: Basic ophthalmological examination, measurement of proptosis, tonometry, visual field examination (Humphrey 30-2 threshold) CT, MRI and carotid angiography were performed. CLINICAL SIGNS: Proptosis of the left eye, dilatation of the episcleral vessels, fundus examination--optic disc normal, enlarged and engorged venous vessels, intraretinal haemorrhages in posterior pole. Tonometry--13 mm Hg- right eye, 24 mm Hg--left eye. Perimetry--general reduction of sensitivity. CT scan--abnormal structure (0.9 x 0.4 cm) in the medial part of the left orbit, without enhancement after applying contrast. MRI examination--enlargement of extraocular muscles of the left eye. Carotid artery angiography revealed indirect carotid--cavernous fistula. CONCLUSION: Special investigations like CT and MRI showed different picture, final diagnosis could be based upon the clinical picture and confirmed by angiography. PMID- 10526450 TI - [The relation of the motion velocity of the real image to the velocity of illusory image in diplopia studied by the clock method]. AB - From a perspective of 34 years' experience of using the clock method to examine ophthalmo-plegia, the author assesses the utility of this method and discusses the ensuing principles. He gives a thorough analysis of the principle which states that if the real image traverses a circular (or semi-circular) path then the apparent image moves in the same time along an elliptic (or semi-elliptic) path. According to his calculations the velocity of the apparent image is greater than the velocity of the real image and proportional to the magnitude of the disparation. The velocity of the apparent image is also important in the process referred to by earlier authors as "diplophobia". The present author gives his own simplified formula for the calculation of the circumference (or surface area) of the ellipse. PMID- 10526451 TI - [The study of the NMDA receptor function in the visual cortex. Summary of habilitation thesis]. AB - PURPOSE: Finding the agents influencing the function of NMDA receptors in the visual cortex. MATERIAL AND METHODS: The experiments were performed on cats aged from three weeks to several years. Recordings were made from single neurons of the visual cortex after iontophoresis of N-methyl-D-aspartate acid, D-2-amino-5 phosphonovaleric acid, D-serine, 7-chlorokynurenic acid. RESULTS: It was found that: 1. Light is one of the factors which has an influence on the development of NMDA receptors in the visual cortex. Rearing cats in the dark is delaying the changes in the function of NMDA receptors, reducing the number of directional sensitive neurons, lowering the firing rate during the visual response. 2. Monocular deprivation is first creating the reduction of the NMDA receptors' contribution to the visual response and later the functional degradation of the synapse. 3. In the Brodmann's 17th area most glycine sites at the NMDA receptors are not saturated by endogenous glycine. PMID- 10526452 TI - [Swedish health care has lost the grasp on information technology. Incompatible systems result in duplication of work and unnecessary costs]. PMID- 10526453 TI - [Even laboratory measurements must be scrutinized for "adverse effects"]. PMID- 10526455 TI - [Future usefulness of school health services]. PMID- 10526454 TI - [Approved national guidelines for diabetic care. Diabetes Center, Sahlgrenska University Center, Gothenburg, Sweden]. PMID- 10526456 TI - [Academic specialist and continuing education!]. PMID- 10526457 TI - [The discussion on the choice of drugs requires more facts]. PMID- 10526458 TI - [Warning against the Babysense]. PMID- 10526459 TI - [There are still some unanswered issues in the survey on helicopters]. PMID- 10526460 TI - [Salt and hypertension. The most recent research on molecular biology adds to the understanding of the connection]. PMID- 10526461 TI - [Future anesthesiologists will be as much outside as inside the operating theatres]. AB - During the first decade of the new millennium the intense reorganisation of hospitals and of medical care will be replaced by stability and long-term goals. An anaesthesiologist is now as active outside as within the operating theatre, being a predominant resource in intensive care, pain management, and emergency and prehospital care. The anaesthesiologist will also have a key part to play in risk analysis of patients scheduled for various kinds of advanced treatment. Anaesthesiologists are now also more involved in primary home care where, together with other physicians and categories of health care providers, they offer qualified treatment of various diseases at home--the environment preferred by the patient. PMID- 10526462 TI - [Laparoscopic living-donor nephrectomy. A small study shows positive results, but the procedure still lacks satisfactory evaluation]. AB - Retrospective analysis and comparison of a small series of 12 laparoscopic live donor nephrectomy (LapLDN) procedures with 15 open live donor nephrectomies, all 27 performed in 1998, showed operating time to be significantly longer but sick leave shorter and hospital stay somewhat shorter in the LapLDN subgroup. One patient in the open procedure subgroup developed herniation and scar discomfort, and in one LapLDN procedure severe bleeding necessitated conversion to open nephrectomy. All kidneys in both subgroups manifested immediate resumption of function after transplantation. Though the LapLDN procedure has yet to be satisfactorily evaluated, the present findings were predominantly in its favour. PMID- 10526463 TI - [How reliable is the laboratory? Increased needs of patient-related quality assurance]. AB - Recent developments in medical care and research involve the increased use of immunochemical assays for hormones, tumour markers, vitamins and drugs. External quality assurance programmes using pooled human sera usually fail to detect analytical interference due to substances (e.g. anti-immunoglobulin or anti ligand antibodies) present in individual serum specimens. The article reports on experience gained during a three-year period when specimens from individual patients attending a thyroid unit were distributed to hospital laboratories in Sweden for analysis. Specimen selection criteria were based on contradictory findings at the initial clinical or laboratory evaluation. The programme has given rise to the formation of a network of the laboratories involved, under the co-ordination of EQUALIS (External quality assurance in laboratory medicine in Sweden). PMID- 10526464 TI - [Who needs all the information collected in computerized medical records? A computer crash shows that to ask the patient is often simpler and quicker]. PMID- 10526465 TI - [The linkage between computerized medical record systems and the national quality registries is not functioning. The catch of data "at the source" is prevented]. PMID- 10526466 TI - [A male contraceptive injection can be available within 5 years]. AB - After more than two decades of attempts to develop a safe male contraceptive, the goal now seems attainable. Spermatogenesis, which is dependent on endogenous testosterone production in the testes, may be controlled by such exogenous steroid hormones as testosterone, gestagens, or combinations of them. The recent development of gonadotrophin-releasing hormone (GnRH) antagonists has provided an added means of depressing testosterone production. Other targets for interference with male fertility are the germinal epithelium (e.g. using the cottonseed oil product, Gossypol, or Triptyrigeum Wilfordii extract), or the maturing sperm in the epididymis (using immunoactive substances). PMID- 10526467 TI - [Watch for cortisone abuse!]. PMID- 10526468 TI - [New initiatives promote clinical research]. AB - The problems of clinical research have now attracted the attention of leading research administrators in the USA. At a recent conference these problems were discussed and the Association for Patient Oriented Research (APOR) was created. One of the major problems was considered to be difficulties in recruiting clinical researchers, but another was the existence of economic obstacles to pathophysiological research based on observations in patients. The National Institutes of Health (NIH) are now making energetic efforts to improve this situation. Congress has approved a doubling of NIH resources over a limited period, a major part of the increase to be allotted to clinical and physiological research. The argument used to obtain Congress approval was that new and powerful advances in technology will facilitate an understanding of the pathogenesis of prevalent diseases, and thus improve treatment efficacy, though achieving these goals will necessitate substantial reinforcement of clinical and physiological research. PMID- 10526469 TI - [Life help or death help? Sedation is a veiled euthanasia during terminal care]. PMID- 10526470 TI - [Family practitioner vs. hospital physician. A debate on patient care continuity in the USA]. PMID- 10526471 TI - [The problem of "man -- environment" in the papers by the I.P. Pavlov's school]. AB - I.P. Pavlov and I.P. Razenkov made a great theoretical, methodological and practical contribution into specification of "human-environment" problem. The followers have successfully continued research in this sphere, enriching science and practice including occupational medicine. PMID- 10526472 TI - [Physiological-ergonomic mechanism of the skeletal muscles]. AB - The article deals with theoretical analysis of present data on physiology of muscular activity. This analysis helped to specify physiologic and ergonomic mechanism of skeletal muscles work. The authors prove this physiologic and ergonomic mechanism to result from known mechanic and chemical processes in muscular tissue structures. Those mechanic and chemical processes underlie well known molecular theories of muscular contraction. The physiologic and ergonomic mechanism includes change in elasticity basic for mechanic models and theories of muscular work, so this mechanism appears superior to other models and theories explaining muscular activity. Physiologic and ergonomic mechanism of muscular work appeared to be genetically linked with morphologic and functional differentiation of locomotory units, with presence of specific muscular zones different in contraction and tensile strain. PMID- 10526473 TI - [Ergonomic characteristics of professional work among violinists]. AB - Complex hygienic, ergonomic, mechanographic studies and video timing revealed that violinist occupational activities are characterized by high physical intensity simultaneously with extreme nervous strain--that is assigned to the second degree of the third class, according present hygienic classification. PMID- 10526474 TI - [Relief of visual fatigue in visual strain-related work]. AB - Work associated with visual strain results in visual fatigue. Laser radiation in red part of spectrum relieves visual fatigue, activating reduction-oxidation thiol disulfide system in eyes. PMID- 10526475 TI - [Computer monitoring of the effectiveness of the operator's adaptation to shift work]. AB - Shift work necessitates operators' reliable and effective activities equally in various parts of the shift despite of diurnal changes of functional activities, occurrence of fatigue and other unfavorable conditions. Computer monitoring of adaptation efficacy in operators could be based on evaluation of changes in test results during various parts of the shift. Analysis of the data in group revealed unfavorable factors of work management. Analysis of the individual data helps to diagnose those who need prophylactic or rehabilitation measures. Thus, computer monitoring is an effective diagnostic tool in work management and aimed to higher reliability of operators. PMID- 10526476 TI - [On the evaluation of the influence of cellular phones on their users]. AB - The authors studied influence of ultrahigh frequency radiation caused by cellular phones on functional state of central nervous, cardiovascular systems and local temperature changes in cellular phones users. The head area near the phone antenna appeared to be under the most intensive heating. Ultrahigh frequency radiation induces significant changes in local temperature and in physiologic parameters of central nervous and cardiovascular systems. PMID- 10526477 TI - [The health problems of computer operators]. AB - The data presented demonstrate that quality of PCs and protective filters is sometimes inadequate. Electric puncture examination was conducted to assess functional state of PC operators before and after the work and of other specialists working in the same conditions without exposure to radiation. Finding is that share of individuals with functional vascular disorders of breast and brain increases annually with constant work at PC. PMID- 10526478 TI - [Approaches to the evaluation of effects caused by industrial pollution on children]. AB - The article deals with results of up-to-date approach to evaluation of xenobiotics' quantitative contribution into development of common pediatric problems. Using multivariate statistics, the authors calculated attributive and relative risks of health disorders in children. Analysis of clinical, epidemiologic and statistic data helped to identify some pathogenetic features of ecologically induced disorders in children. PMID- 10526479 TI - [The evaluation of skin temperature points and heat flow in the area of the foot for the measurement of the footwear heat insulation]. AB - Analyzing results of the personal experiments, the authors present methodology to minimize number of sites for measurement of foot skin temperature and heat flow. Average foot temperature could be reliably measured through one or several sites. The studies revealed informativeness of each among 11 measurement sites and helped to specify corresponding equations for evaluation of foot heat conditions and heat insulation. PMID- 10526481 TI - [On the use of biodestructor "Disoil" for the purification of soil from petroleum wastes]. AB - Sanitation of territories polluted with oil products is possible and quite effective through biodestructor produced in Russia on the basis of nonpathogenic yeast lines. Oil content of the soil lowers by 60-78%. Self-clearance of the soil is preserved, toxicity level of the soil does not increase. PMID- 10526480 TI - [X-ray densitometry and biochemical parameters of bone tissue metabolism in patients with vibration disease]. AB - To reveal systemic and local osteoporosis, the authors studied biochemical markers of bone metabolism in vibration disease patients. The vibration disease patients appeared to have the most frequent and marked osteoporosis in peripheral bones--hands (in 90% of cases) and forearms (in 66.7%). Prevalence of systemic osteoporosis and osteopenia reached 11.7 and 48.3% in the select respectively. According to biochemical markers, bone reconstruction state was characterized by moderately intensified bone resorption and diminished bone formation. PMID- 10526482 TI - [Hygienic regulation of air concentration of byproducts of insecticide permethrin synthesis in the working area air]. PMID- 10526483 TI - [A new therapeutic approach to autoimmune diseases: the engraftment of hematopoietic stem cells]. PMID- 10526484 TI - [Self-measurement of blood pressure: history of a method with a future]. PMID- 10526485 TI - [Anatomy, physiology and clinical examination of the shoulder]. AB - Shoulder pain is usually caused by periarthritis. Impingement syndrome and frozen shoulder (adhesive capsulitis) are the more frequently clinical aspect observed. A shoulder standardised clinical examination often give a precise lesional diagnostic. Complementary examinations are useful to distinguish other cause of shoulder pain. PMID- 10526486 TI - [Imaging of the shoulder]. AB - Except for instability, a variety of causes has been described in the painful shoulder. A thorough clinical examination may help establish the cause of shoulder pain. Even if modern imaging modalities (ultrasonography, computed tomography, MR imaging) allowed a precise diagnostic and therapeutic approach, plain radiographs are still necessary. Performed by an experienced radiologist, ultrasonography may be useful for the assessment of rotator cuff tears which are a common problem. CT arthrography and MR imaging are only recommended in preoperative assessment. In calcific tendinitis plain radiographs provide evidence of calcifications. It is not difficult to establish clinically the diagnosis of adhesive capsulitis and arthrography is the only technique to reveal the articular capacity limitation reliably. In degenerative osteoarthritis of the glenohumeral joint, CT arthrography is necessary to assess the rotator cuff changes and lenoid osseus findings before undergoing shoulder arthroplasty. PMID- 10526488 TI - [Adhesive capsulitis of the shoulder]. AB - Adhesive capsulitis is a painful stiff shoulder due to the thickening of the capsule and synovium. The main observed changes are hyperhaemia of the synovium and a capsular fibrosis similar to that of Dupuytren's disease. Stiffness involves mainly flexion, lateral rotation and abduction. In most cases, a spontaneous healing is observed within 12 to 30 months. When the capsulitis is disabling and pain still present, a joint distension followed by rehabilitation can be indicated. When the disability is important and mainly due to stiffness, a manipulation under anesthesia with or without arthroscopic release of soft tissues can be indicated. PMID- 10526489 TI - [Arthritis of the shoulder]. AB - Diagnosis of shoulder arthritis should situate the precise location (glenohumeral, acromioclavicular, scapulothoracic) and rapidly determine the cause in order to eliminate the possibility of septic arthritis which requires urgent treatment. Good knowledge of shoulder symptomatology is essential. When there is joint effusion, the fluid should be sampled for rapid analysis (cells, crystals, germs), and the work-up for diagnosis must include clinical and laboratory analyses as well as imaging. Causes are especially microcrystallin arthritis (hydroxyapatite rheumatism, chondrocalcinosis, etc.) and inflammatory rheumatism (rheumatoid arthritis, ankylosis spondylitis, etc.). Septic arthritis is much less common but more rapidly destructive, which justifies their consideration whenever suspected. Treatment should be rapidly initiated in acute, particularly septic forms. In other forms, diagnosis guides treatment. Steps should be taken rapidly, both medically and sometimes surgically, to avoid destructive articular and periarticular lesions (rotator cuff). PMID- 10526487 TI - [Pathology of the rotator cuff]. AB - In daily practice painful shoulder is frequent. Rotator cuff pathology (traumatic, microtraumatic or both) is frequently involved. Treatment is closely related to diagnosis, which directly depends on features given by the triad "questioning, clinical exam, standard X-rays". In some very precise cases this triad will be completed by more complex exams (arthro CT, MRI). Rotator cuff pathology is diversified: either tendinopathies (calcifying or not) or tears (partial or transfixiant). Rotator cuff tear's natural evolution is a slow aggravation to a final subacromial arthrosis or an excentrated omarthrosis in a certain term which tolerance may vary. PMID- 10526490 TI - [Non-inflammatory osteoarthritis of the shoulder]. AB - Although rotator cuff disorder is by far the leading cause of shoulder pain, such pain can also reflect actual osteoarthritis. In shoulder pathology, non inflammatory osteoarthritis should be kept in mind, as it is often the cause of symptoms resembling those of degenerative rotator cuff, with which it is most often associated. Among types of non-inflammatory osteoarthritis of the shoulder, the most often observed are acromioclavicular osteoarthritis and osteonecrosis of the humeral head. PMID- 10526491 TI - [Shoulder instability]. AB - Shoulder instability is a sign described by the patient. The etiology of this instability is varied: sometimes it is the result of a traumatic luxation with tear of the gleno-humeral inferior ligament, sometimes it is the result of an abnormal hyperlaxity. The examiner must be able to do a difference between these two causes because treatment and consequence are very different. Nevertheless, these two factors can be associated at variable degrees. A careful history and examination, a precise x-rays and scanning can help the examiner to solve this difficult problem. PMID- 10526492 TI - [Surgery for non-traumatic shoulder lesions]. AB - The last decade has witnessed considerable development in shoulder surgery, particularly in three domains: anterior instability, rotator cuff disorders and glenohumeral degenerative osteoarthritis. The development of arthroscopy and improvement in implant design have been involved in this development. The basic influence of the trophicity of the periarticular muscles on the results of rotator cuff repair and of joint prostheses is a recent notion that has led to optimising therapeutic indications. The future of shoulder surgery essentially resides in perfecting arthroscopic technique and in improving knowledge of joint kinetics. PMID- 10526493 TI - [Evidence-based medicine: pluses and minuses (I)]. PMID- 10526494 TI - [Tremor. Diagnostic orientation]. PMID- 10526495 TI - [Gastro-esophageal reflux. Physiopathology, diagnosis, development and treatment]. PMID- 10526496 TI - [Squamous cell carcinoma of the tongue. Epidemiology, diagnosis, complications and long-term treatment]. PMID- 10526497 TI - [Horton's disease. Pathologic anatomy, diagnosis, evolution, treatment]. PMID- 10526498 TI - [Anxiolytics. Principles and rules of their utilization]. PMID- 10526499 TI - [Psoriasis. Diagnosis, evolution, principles of treatment]. PMID- 10526500 TI - [Micro-imagery MRI and ultrasonography of cartilage]. PMID- 10526501 TI - [Ultrastructural imagery of chondrocytes]. PMID- 10526502 TI - [Interleukin 1: Its role, its dosage, the difficulties in advances in arthritis. Results of a "pilot" study with diacerheine (ART 50) in gonarthrosis]. PMID- 10526503 TI - [Interleukin-1, nitric oxide synthase and cartilage]. PMID- 10526504 TI - [Clinical relevance of radiologic joint space measurements in coxarthrosis]. PMID- 10526505 TI - [The outcome of total hip replacement in the development of coxarthrosis]. PMID- 10526506 TI - [Interpretation of the complaints and expectations of arthritis patients]. PMID- 10526507 TI - [Preventive surgery in the course of secondary coxarthrosis]. PMID- 10526508 TI - [Digital arthritis: what diagnostic resources and what modalities for its management?]. PMID- 10526509 TI - [Rapidly destructive coxarthrosis]. PMID- 10526510 TI - Assessment of competence. PMID- 10526511 TI - Laparoscopy and acute appendicitis. PMID- 10526512 TI - Complete surgical training. PMID- 10526513 TI - Soft-tissue images. Abdominal aortic aneurysm causing lumbar plexus neuropraxis. PMID- 10526514 TI - Musculoskeletal images. Early bone changes in hyperparathyroidism detected on magnetic resonance imaging. PMID- 10526515 TI - Soft-tissue case 29. Adult ileocolic intussusception. PMID- 10526516 TI - Musculoskeletal case 6. High-grade osteosarcoma of the iliac bone. PMID- 10526517 TI - Hypothermia and the trauma patient. AB - Hypothermia has profound effects on every system in the body, causing an overall slowing of enzymatic reactions and reduced metabolic requirements. Hypothermic, acutely injured patients with multisystem trauma have adverse outcomes when compared with normothermic control patients. Trauma patients are inherently predisposed to hypothermia from a variety of intrinsic and iatrogenic causes. Coagulation and cardiac sequelae are the most pertinent physiological concerns. Hypothermia and coagulopathy often mandate a simplified approach to complex surgical problems. A modification of traditional classification systems of hypothermia, applicable to trauma patients is suggested. There are few controlled investigations, but clinical opinion strongly supports the active prevention of hypothermia in the acutely traumatized patient. Preventive measures are simple and inexpensive, but the active reversal of hypothermia in much more complicated, often invasive and controversial. The ideal method of rewarming is unclear but must be individualized to the patient and institution specific. An algorithm reflecting newer approaches to traumatic injury and technical advances in equipment and techniques is suggested. Conversely, hypothermia has selected clinical benefits when appropriately used in cases of trauma. Severe hypothermia has allowed remarkable survivals in the course of accidental circulatory arrest. The selective application of mild hypothermia in severe traumatic brain injury is an area with promise. Deliberate circulatory arrest with hypothermic cerebral protection has also been used for seemingly unrepairable injuries and is the focus of ongoing research. PMID- 10526518 TI - Ileoanal anastomosis with reservoirs: complications and long-term results. AB - OBJECTIVE: To determine the rate of complications of ileoanal pouch anastomosis, their treatment and their influence on a successful outcome. DESIGN: A computerized database and chart review. SETTING: Three academic tertiary care health centres. PATIENTS: All 239 patients admitted for surgery between 1981 and 1994 with a diagnosis of ulcerative colitis and familial adenomatosis coli. INTERVENTIONS: Sphincter-saving total proctocolectomy and construction of either S-type of J-type ileoanal reservoir. OUTCOME MEASURES: Indications, early and late complications, incidence of pouch excision. RESULTS: Of the 239 patients, 228 (95.4%) were operated on for ulcerative colitis and 11 (4.6%) for familial polyposis coli. One patient in each group was found to have a carcinoma not previously diagnosed. Twenty-eight patients had poor results: in 17 (7.1%) the ileostomy was never closed or was re-established because of pelvic sepsis or complex fistulas, sclerosing cholangitis or severe diarrhea; 11 (4.6%) patients required excision of the pouch because of anal stenosis, perirectal abscess fistula or rectovaginal fistula. Three patients died--of suicide, and complications of liver transplantation and HIV infection. Thus, 208 patients maintained a functioning pouch. The early complication rate (within 30 days of operation) was 57.7% (138 patients) and the late complication rate was 52.3% (125 patients). Pouchitis alone did not lead to failure or pouch excision. Emptying difficulties in 25 patients with anal stenosis were helped in 2 by resorting to intermittent catheterization. Patients with indeterminate colitis had a higher rate of anorectal septic complications, and all patients having Crohn's disease after pouch construction had complicated courses. CONCLUSIONS: The complication rate associated with ileoanal pouch anastomosis continues to be relatively high despite increasing experience with this technique. Overall, however, a satisfactory outcome was obtained in 87% of patients. PMID- 10526519 TI - Resident continuity of care experience in a Canadian general surgery training program. AB - OBJECTIVES: To provide baseline data on resident continuity of care experience, to describe the effect of ambulatory centre surgery on continuity of care, to analyse continuity of care by level of resident training and to assess a resident run preadmission clinic's effect on continuity of care. DESIGN: Data were prospectively collected for 4 weeks. All patients who underwent a general surgical procedure were included if a resident was present at operation. SETTING: The Division of General Surgery, Queen's University, Kingston, Ont. OUTCOME MEASURES: Preoperative, operative and inhospital postoperative involvement of each resident with each case was recorded. RESULTS: Residents assessed preoperatively (before entering the operating room) 52% of patients overall, 20% of patients at the ambulatory centre and 83% of patients who required emergency surgery. Of patients assessed by the chief resident, 94% were assessed preoperatively compared with 32% of patients assessed by other residents (p < 0.001). Of the admitted patients, 40% had complete resident continuity of care (preoperative, operative and postoperative). There was no statistical difference between this rate and that for emergency, chief-resident and non-chief-resident subgroups. Of the eligible patients, 58% were seen preoperatively by the resident on the preadmission clinic service compared with 54% on other services (p < 0.1). CONCLUSIONS: This study serves as a reference for the continuity of care experience in Canadian surgical programs. Residents assessed only 52% of patients preoperatively, and only 40% of patients had complete continuity of care. Factors such as ambulatory surgery and junior level of training negatively affected continuity experience. Such factors must be taken into account in planning surgical education. PMID- 10526520 TI - Home prophylactic warfarin anticoagulation program after hip and knee arthroplasty. AB - OBJECTIVE: To determine the efficiency of a program designed to maintain prophylactic oral anticoagulation within a target range for 6 weeks after hip and knee arthroplasty. DESIGN: A prospective continuous quality improvement indicator. SETTING: A tertiary care university hospital. PATIENTS: Patients who underwent hip and knee arthroplasty and had no indications for routine anticoagulation other than postoperative thromboembolism prophylaxis. INTERVENTION: An outpatient warfarin prophylaxis program, which included an information letter given to the patient. Home Care coordinated community laboratory services, communication with and anticoagulant dosage adjustment by the patient's personal family physician. OUTCOME MEASURES: The proportion of international normalized ratio (INR) values within, below and above the target range of 2.0 to 3.0. RESULTS: Sixty-two patients were enrolled over a 3-month period. On the day of hospital discharge, 64% of patients had INR values that were within the target range, 31% were below and 5% were above. After hospital discharge, 42% of the INR values were within the target range, 48% were below and 10% were above. CONCLUSION: Despite a program designed to address patient information, physician communication and laboratory testing, tight control of home INR values could not be achieved with the existing resources of Home Care and family physicians. PMID- 10526521 TI - Safety of the limited open technique of bone-transfixing threaded-pin placement for external fixation of distal radial fractures: a cadaver study. AB - OBJECTIVE: To examine the safety of threaded-pin placement for fixation of distal radial fractures using a limited open approach. DESIGN: A cadaver study. METHODS: Four-millimetre Schanz threaded pins were inserted into the radius and 3-mm screw pins into the second metacarpal of 20 cadaver arms. Each threaded pin was inserted in the dorsoradial oblique plane through a limited open, 5- to 10-mm longitudinal incision. Open exploration of the threaded-pin sites was then carried out. OUTCOME MEASURES: Injury to nerves, muscles and tendons and the proximity of these structures to the threaded pins. RESULTS: There were no injuries to the extensor tendons, superficial radial or lateral antebrachial nerves of the forearm, or to the soft tissues overlying the metacarpal. The lateral antebrachial nerve was the closest nerve to the radial pins and a branch of the superficial radial nerve was closest to the metacarpal pins. The superficial radial nerve was not close to the radial pins. CONCLUSION: Limited open threaded-pin fixation of distal radial fractures in the dorsolateral plane appears to be safe. PMID- 10526522 TI - Optimizing femorotibial alignment in high tibial osteotomy. AB - OBJECTIVE: To study factors that affect femorotibial (F-T) alignment after valgus closing wedge tibial osteotomy. STUDY DESIGN: A review of standardized standing radiographs. Femorotibial alignment was measured 1 year postoperatively for over- and under-correction. Changes in F-T alignment and in tibial plateau angle were measured. SETTING: An urban hospital and orthopedic clinic. PATIENTS: Eighty-two patients with osteoarthritis and varus femorotibial alignment underwent valgus closing wedge tibial osteotomy. Patients having a diagnosis of inflammatory arthritis or a prior osteotomy about the knee were excluded. RESULTS: A 1 degree wedge removed from the tibia resulted in an average correction F-T alignment of 1.2 degrees. A knee that had increased valgus orientation of the distal femur had a greater degree of correction, averaging 1.46 degrees in F-T alignment per degree of tibial wedge. This resulted in excessive postoperative valgus alignment for some patients who had increased valgus tilt of the distal femur. Optimal F-T alignment of 6 degrees to 14 degrees valgus occurred when the postoperative tibial inclination was 4 degrees to 8 degrees of valgus. CONCLUSIONS: There was a trend for knees with increased valgus orientation of the distal femur to have greater correction in F-T alignment after tibial osteotomy, likely because of a greater opening up of the medial joint space during stance. Surgeons need to account for this in their preoperative planning. PMID- 10526523 TI - Borrmann's type IV gastric cancer: clinicopathologic analysis. AB - OBJECTIVE: To determine whether there is a specific pattern of clinicopathological features that could distinguish Borrmann's type IV gastric cancer from other types of gastric cancer. DESIGN: A retrospective study of patients with advanced gastric cancer treated between 1985 and 1995. SETTING: The Department of Surgery, Sendai National Hospital, a 716-bed teaching hospital. PATIENTS: The clinicopathologic features of 88 patients with Borrmann's type IV carcinoma of the stomach were reviewed from the database of gastric cancer. The results were compared with those of 309 patients with other types of gastric carcinoma. MAIN OUTCOME MEASURES: Gender, age, tumour size, depth of invasion, histologic type, cancer-stromal relationship, histologic growth pattern, nodal involvement, lymphatic and vascular invasion, type of operation, cause of death and 5-year survival. RESULTS: Women were afflicted as commonly as men in the Borrmann's type IV group. These patients tended to be younger and to have larger tumours involving the entire stomach than patients with other types of cancer. Histologic type was commonly diffuse and scirrhous, and serosal invasion was prominent with infiltrative growth. Nodal involvement and lymphatic invasion were more common in patients with Borrmann's type IV than in those with other types of gastric cancer. The disease was advanced in most instances and a total gastrectomy was performed in 55% of the patients. The survival rate of patients with Borrmann's type IV tumour was lower than for patients with other types of gastric cancer (p < 0.005, log-rank test). CONCLUSIONS: In Borrmann's type IV gastric cancer, early detection and curative resection are crucial to extend the patient's survival. Aggressive postoperative chemotherapy is recommended when a noncurative resection is performed. PMID- 10526524 TI - A meta-analysis of laparoscopic versus open appendectomy in patients suspected of having acute appendicitis. AB - OBJECTIVE: To determine if any significant differences exist between laparoscopic appendectomy (LA) and open appendectomy (OA). DESIGN: A meta-analysis of randomized controlled trials (RCTs) comparing LA to OA. DATA SOURCES: An extensive literature search was conducted for appropriate articles published between January 1990 and March 1997. Articles were initially retrieved through MEDLINE with MeSH terms "appendicitis" or "appendectomy" and "laparoscopy". Additional methods included cross-referencing bibliographics of retrieved articles, hand searching abstracts from relevant meetings and consultation with a content expert. STUDY SELECTION: Only RCTs published in English in which patients had a preoperative diagnosis of acute appendicitis were included. DATA EXTRACTION: The outcomes of interest included operating time, hospital stay, readmission rates, return to normal activity and complications. The Cochrane Collaboration Review Manager 3.0 was used to calculate odds ratios (OR), weighted mean differences (WMD) and 95% confidence intervals (CI). The random-effects model was used for statistical analysis. DATA SYNTHESIS: Twelve trials met the inclusion criteria. Because there were insufficient data in some trials, operating time, hospitalization and return to work were assessed in only 8 trials. Mean operating time was significantly longer with LA (WMD 18.10 minutes, 95% CI 12.87 to 23.15 minutes). There were fewer wound infections in LA (OR 0.40, 95% CI 0.24 to 0.69), but no significant differences in intra-abdominal abscess rates (OR 1.94, 95% CI 0.68 to 5.58). There was no significant difference in the mean length of hospital stay (WMD -0.16 days, 95% CI -0.44 to 0.15 days) or readmission rates (OR 1.16, 95% CI 0.54 to 2.48). However, the return to normal activity was significantly earlier with LA (WMD -5.79 days, 95% CI -7.38 to -4.21 days). Sensitivity analyses did not affect the results. CONCLUSION: This meta analysis suggests that operating room time is significantly longer, hospital stay is unchanged but return to normal activities is significantly earlier with LA. PMID- 10526525 TI - Mechanical failure of a gamma nail in a patient with an impending pathologic subtrochanteric fracture. PMID- 10526526 TI - Endometrioma simulating inguinal hernia: case reports. PMID- 10526527 TI - Ruptured esophageal intramural pseudodiverticulum: an unusual cause of mediastinitis in a child. PMID- 10526528 TI - Cytogenetic study of eight new cases of radiation-induced solid tumors. AB - Radiation-induced tumors were selected according to the criteria defined by Cahan (1948) for sarcomas. Cell cultures and/or xenografts in nude mice were performed with biopsies obtained from second primary tumors. Karyotypes of eight tumors were established after R-banding. After comparison with literature data on 15 other cases, two distinct cytogenetic patterns could be distinguished. One was characterized by polyclonal karyotypes, of which a large proportion were simple and carriers of balanced translocations. Another one was characterized by monoclonal chromosome alterations observed in highly aneuploid and complex karyotypes, in which many deletions were observed. These two different patterns could be related to the modality of metaphase harvesting. Polyclonal karyotypes were preferentially observed after long-term cultures, and monoclonal karyotypes after short-term cultures or xenografts. The following scheme of radiation oncogenesis is proposed: a) induction of recessive gene mutations including that of tumor suppressor genes; b) accumulation of genomic alterations in the irradiated tissue with aging, including deletions or mutations of normal alleles from mutated tumor suppressor genes; and c) loss of tumor suppressor gene function and initiation of a multistage tumor development and progression. Polyclonal abnormalities are assumed to exist in noncancerous cells which acquired radiation-induced chromosome aberrations. PMID- 10526529 TI - Expression of HMGIY in three uterine leiomyomata with complex rearrangements of chromosome 6. AB - Uterine leiomyomata (UL) are a major public health problem, yet little is known about their etiology. Genetic factors likely influence UL development and growth; for example, approximately 40% of UL have chromosomal abnormalities detectable by conventional cytogenetic analysis, including t(12;14)(q15;q23-24), rearrangements involving the short arm of chromosome 6 and interstitial deletions of the long arm of chromosome 7. Two high-mobility group (HMG) protein genes, HMGIC and HMGIY, located at 12q15 and 6p21.3, respectively, are involved in rearrangements in various mesenchymal tumors including UL. In this study, we investigated HMGIY expression in three UL with complex cytogenetic rearrangements of 6p21.3 by reverse transcriptase-polymerase chain reaction (RT-PCR) and electrophoretic shift assay (EMSA). Our findings suggest that there are multiple mechanisms for HMGIY dysregulation, which may include post-translational modification of the hmgiy protein and dysregulation due to different translocation partners. Furthermore, the mechanism dysregulating HMGIY in UL with 6p21.3 and 14q23-24 rearrangements may be similar to the mechanism dysregulating HMGIC in UL characterized as t(12;14)(q15;q23-24), because of the common involvement of an HMG gene and a gene at 14q23-24. PMID- 10526531 TI - Comparison of chromosomal imbalances in neuroendocrine and non-small-cell lung carcinomas. AB - Lung carcinomas are represented by non-small-cell lung carcinomas (NSCLC) and neuroendocrine carcinomas (NE) which differ in their clinical presentation and prognosis. We used comparative genomic hybridization (CGH) to characterize and compare the chromosomal pattern of 11 NSCLC and 11 high-grade NE lung carcinomas. Overall, the total number of aberrations was higher in NSCLC than in high-grade NE lung tumors (p < 0.05) and gains predominated over losses in NSCLC (p < 0.0003). Gains common to both lung tumor phenotypes were detected in 1p, 1q, 3q, 5p, 6p, 8q, 12, 17q, 19p, 19q, 20p, 20q, and X, whereas common losses were found in 2q, 3p, 4p, 4q, 5q, 8p, 9p, 10p, 11p, 11q, 13q, and 17p. Major gains on 18q and losses on 2p and 16q were exclusively detected in high-grade NE lung tumors. On the other hand, major gains on 2p and 15q and losses on 21q were found only in NSCLC. Furthermore, gains within 22q11-q12 and 7p12-p15 were associated with NSCLC (p < 0.05). The differences in the pattern and distribution of genetic changes observed in NSCLC as opposed to high-grade NE lung carcinomas suggest the existence of distinct tumorigenic pathways between these two major classes of lung tumors. PMID- 10526530 TI - Insertion of the 5' part of BCR within the ABL gene at 9q34 in a Philadelphia negative chronic myeloid leukemia. AB - We report a chronic myeloid leukemia patient without evidence of a Philadelphia (Ph) chromosome in whom RT-PCR analysis performed in blast crisis demonstrated the existence of both common b3a2 and b2a2 BCR/ABL fusion transcripts. In situ hybridization studies with BCR- and ABL-specific probes showed location of the BCR/ABL fusion gene on chromosome 9, band q34, instead of at chromosome 22q11, and that it resulted from an insertion of the 5' side of BCR within the ABL gene on chromosome 9. The vast majority of cells showed a BCR/ABL fusion gene on both chromosomes 9, which is equivalent to a double Ph chromosome, thus reinforcing the notion that the critical event in CML is the formation of a functional BCR/ABL fusion gene. PMID- 10526532 TI - DNA copy number losses are more frequent in primary larynx tumors with lymph node metastases than in tumors without metastases. AB - Comparative genomic hybridization was performed on 38 primary laryngeal carcinomas divided into two groups according to the metastatic phenotype. DNA copy number changes were detected in 22 of the 38 cases (57.9%). Gains were most frequently observed at 3q, 8q, and 9q, and losses were found in decreasing order at 18q, 3p, and 4. The mean value of losses was 2.5 times as high in metastasizing primary tumors (23/38) as in nonmetastasizing tumors. The most frequent losses in metastasizing tumors were at 18q, 3p, and 5q. PMID- 10526534 TI - Karyotypic analyses of hepatoblastoma. Report of two cases and review of the literature suggesting chromosomal loci responsible for the pathogenesis of this disease. AB - Two cases of fetal hepatoblastoma with unique karyotypic changes are described. One was a 17-month-old boy with multiple unbalanced chromosomal translocations, resulting in four types of derivative chromosomes involving chromosomal loci at 1q21, 1q32, 2q23, 6q27, 7p22, and 21p12, partial tetrasomy of 1q, partial trisomy of 2q, and partial monosomy of 21p. The clonal karyotype of this tumor was 46,XY,der(2)t(1;2)(q32;q37), der(6)t(1;6)(q12;q27), der(7)t(2;7)(q23;p22), der(21)t(2;21) (q23;p12). In the other case, a 4-year-old girl, karyotypic analyses revealed trisomy 2 and 8, and the clonal karyotype of this case was 48,XX,+2,+8. Review of these cases together with previous reports suggested the significance of chromosomal changes including numerical abnormalities of 1q, 2(or 2q), 20, and 8 (or 8q), and breakage of 1q and 2q in the development of hepatoblastoma. The results presented herein underscore the significance of numerical abnormalities of chromosomal regions 1q and 2q and of chromosome 8 in the development of hepatoblastoma, in addition to abnormalities of 6q27, 7p22, and 21p12-13 as other chromosomal loci that may be responsible for the pathogenesis of this embryonal type of tumor. PMID- 10526533 TI - Clinical correlations of genetic changes by comparative genomic hybridization in Ewing sarcoma and related tumors. AB - Our previous comparative genomic hybridization (CGH) study of Ewing sarcoma and related tumors showed that DNA sequence copy number increases of 1q21-q22 and of chromosomes 8 and 12 were associated with trends toward poor survival (Armengol et al., Br J Cancer 1997, 75, 1403-1409). These trends were not statistically significant. In the present study, we analyzed 28 primary Ewing sarcomas and related tumors by CGH to study whether these (or other) changes have prognostic value in these tumors. Twenty-one tumors (75%) had changes with a mean of 1.9 changes per tumor. The most frequent aberration was gain of chromosome 8 in 10 tumors (36%). Five tumors (18%) had copy number increases at 1q21-22 and 5 had gain of 7q. Copy number increase of 6p21.1-pter, gain of chromosome 12, and loss of 16q were seen in 11%. Copy number increases of 1q21-q22 and of chromosomes 8 and 12 were associated with trends toward worse outcome, but the differences did not reach statistical significance. A novel finding is the association of copy number increase at 6p with worse distant disease-free (P = 0.04) and overall survival (P = 0.004). To confirm this finding and to see whether copy number increases of 1q21-q22 and of chromosomes 8 and 12 have definite prognostic value, a larger number of cases needs to be studied. PMID- 10526535 TI - Biphenotypic hematological malignancy with T-lymphoid and myeloid differentiation: association with t(3;12)(p25;q24.3). Case report and review of the literature. AB - Biphenotypic hematological malignancies of T-lymphoid and myeloid differentiation are relatively rare and have most commonly been associated with t(8;13). However, this entity is invariably associated with eosinophilia and generally progresses to acute leukemia within a year of diagnosis. We describe a case of a biphenotypic hematological malignancy with T-lymphoid and myeloid differentiation without associated eosinophilia; however, there was an association with t(3;12)(p25;q24.3) as a sole abnormality and progression to acute leukemia within 10 months of presentation. This association with such a malignancy has not previously been described. Additional cases need to be accrued to determine the prognostic significance and clinical implications of such an association. PMID- 10526536 TI - Chromosome 11 abnormalities in myelodysplastic syndromes. AB - Cytogenetic studies were performed in 140 patients with myelodysplastic syndrome (MDS) at diagnosis. Chromosome 11 anomalies were found in 7 cases (5%); 2 of these patients had refractory anemia (RA), 2 had refractory anemia with excess of blasts (RAEB), 1 had RAEB in transformation (RAEB-t), and 2 had chronic myelomonocytic leukemia (CMMoL) according to the French-American-British (FAB) Cooperative Group criteria. The chromosome 11 abnormalities comprised trisomy 11 (2 patients), monosomy 11 (1 patient), del(11)(q23) (2 patients), add(11)(p15) (1 patient), and der(11) t(3;11)(p21;q23) (1 patient). Abnormalities involving band q23 of chromosome 11 occurred in 3 cases and were the most common alteration. However, specific chromosomal alterations were not associated with any FAB classification group. These findings and their implications in the biology of MDS are discussed. PMID- 10526537 TI - Chromosomal aberrations in Bilharzial bladder cancer as detected by fluorescence in situ hybridization. AB - Cancer of the bladder is a frequent malignancy in Egypt and other developing countries in which bladder infection with the parasite Schistosoma haematobium is common. Several epidemiological, histopathological, and clinical characteristics of cancer of the Bilharzial bladder suggest that it is distinct from bladder cancer seen in other places in the world. No numerical aberrations of chromosomes that might be specific for Bilharzial bladder carcinoma have been established. In this study, we used fluorescence in situ hybridization (FISH) with centromere specific probes for chromosomes 3, 4, 7, 8, 9, 10, 11, 16, and 17 to detect numerical aberrations of these chromosomes in frozen-stored samples of 31 Egyptian patients affected with Bilharzial carcinoma. Among 5 types of chromosomes examined, imbalance was observed; the most common imbalance was a loss of chromosome 9 (48.4%), with numerical aberration of chromosome 17 being the second most-frequent anomaly (19.4%). The presence of such anomalies, especially losses of chromosome 9, are associated with a younger age group of patients, as well as with a lower grade tumor and negative pelvic node involvement by the disease. Fluorescence in situ hybridization analysis thus proved to be a useful method for detecting numerical aberrations of individual chromosomes, with application to touch preparations of frozen-stored tissue having the advantage of exact sampling of cancer foci. This result also suggests that the mechanism of genetic progression of bladder cancer is independent of its etiology. PMID- 10526539 TI - Fluorescence in situ hybridization analysis of complex translocations in two newly diagnosed Philadelphia chromosome-positive chronic myelogenous leukemia patients. AB - Two different complex translocations from newly diagnosed cases of Philadelphia chromosome-positive chronic myelogenous leukemia (CML) were characterized by G banding and fluorescence in situ hybridization (FISH) analysis. In one case, a unique balanced t(9;22;9;11) (q34;q11;p22;q23) was identified by G-banding, and confirmed by FISH using MBCR/ABL and painting probes. In the second case, an apparently balanced t(19;22) was identified by G-banding analysis. FISH using MBCR/ABL probe detected the fusion gene on the derivative chromosome 22, indicating the involvement of chromosome 9. Further FISH analysis with selected painting probes showed that the t(19;22) was a result of a complex translocation involving chromosomes 9, 19, 21, and 22. PMID- 10526538 TI - Translocation (10;11)(p13;p15) in an infant acute myeloid leukemia with MLL gene rearrangement. AB - Molecular rearrangements of the MLL gene at the 11q23 region have been identified in most cases of infant leukemia, regardless of the phenotype. We present a case of acute myeloid leukemia which coexpressed myeloid and lymphoid markers in a 12 month-old girl. Karyotype analysis revealed the presence of a thus far unreported translocation t(10;11)(p13;p15). Although no 11q23 abnormalities were cytogenetically detectable, an MLL gene molecular rearrangement was found. PMID- 10526540 TI - Structural abnormalities of chromosome 2 in benign thyroid tumors. Three new cases and review of the literature. AB - Here we report our cytogenetic findings on three cases of nodular goiter, all showing structural clonal abnormalities of chromosome 2. In the first case, we found a t(2;3)(q21;q27 or q28) in two nodules of the same patient. The second case revealed a t(2;20;3)(p21;q11.2;p25), and the third case showed a t(1;2)(p22;p13). When the data from the literature and the present cases are summarized, the results suggest the existence of at least three breakpoint clusters of chromosome 2 in benign thyroid tumors or hyperplasias. PMID- 10526541 TI - Translocation (X;5)(q13;q33) in essential thrombocythemia. AB - We report a novel chromosomal translocation (X;5)(q13;q33) in a woman with no history of prior chemotherapy or radiotherapy, found to have essential thrombocythemia. Aberrations in chromosome 5, mostly deletions of 5q, have been described in essential thrombocythemia; however, a t(X;5) translocation has not been reported. PMID- 10526542 TI - Epithelioid sarcoma of the proximal type with complex karyotype including i(8q) PMID- 10526544 TI - A novel t(1;10)(q32;p12) in acute myelomonocytic leukemia. PMID- 10526543 TI - Basophilic crisis of chronic myeloid leukemia with a novel chromosomal aberration. PMID- 10526545 TI - Laparoscopic management of hiatal hernia and gastroesophageal reflux. PMID- 10526546 TI - Responses of HMO medical directors to trust building in managed care. AB - Managed care organizations (MCOs) are facing intense criticism at national, state, and local levels and battling initiatives that would impose stricter regulation. Medical directors of HMOs were surveyed regarding their organizations' strategies of communication, the programs they have instituted to build trust, and their commitment to sponsoring family and patient support groups. The responses obtained from 252 directors indicate that nonprofit and free-standing organizations are more likely than either for-profit HMOs or organizations that are part of a chain to sponsor community activities and programs and to offer family and patient support groups. Staff- and group-model HMOs are more likely than other organizational configurations to initiate many types of "trust programs." The results indicate that more dispersed and "virtual type" organizations must explore ways to respond meaningfully to community concerns--and to public health, prevention, and health promotion needs as well- while continuing to improve their practice patterns. PMID- 10526547 TI - Interpersonal processes of care in diverse populations. AB - Persons of lower socioeconomic status and members of racial and ethnic minority groups experience poorer health and increased health risk factors. A framework of interpersonal processes of care specifies distinct components and incorporates the perspective of diverse racial and ethnic or socioeconomic groups. Its dimensions, each with several domains, are communication (general clarity, elicitation of and responsiveness to patient concerns, explanations, empowerment), decision making (responsiveness to patient preferences, consideration of ability and desire to comply), and interpersonal style (friendliness, respectfulness, discrimination, cultural sensitivity, support). All the domains, except cultural sensitivity, were validated through a survey of 603 ethnically diverse, low-income adults. Confirmation of the framework's usefulness should enable researchers to explore how interpersonal processes might account for observed ethnic and social class differences in health care and health. PMID- 10526548 TI - Methadone maintenance and state Medicaid managed care programs. AB - Coverage for methadone services in state Medicaid plans may facilitate access to the most effective therapy for heroin dependence. State Medicaid plans were reviewed to assess coverage for methadone services, methadone benefits in managed care, and limitations on methadone treatment. Medicaid does not cover methadone maintenance medication in 25 states (59 percent). Only 12 states (24 percent) include methadone services in Medicaid managed care plans. Moreover, two of the 12 states limit coverage for counseling or medication and others permit health plans to set limits. State authorities for Medicaid and substance abuse can collaborate to ensure that appropriate medication and treatment services are available for Medicaid recipients who are dependent on opioids and to construct payment mechanisms that minimize incentives that discourage enrollment among heroin-dependent individuals. PMID- 10526549 TI - Community care for people with chronic conditions: an analysis of nine studies of health and social service utilization in Ontario. AB - A series of studies conducted in the same region found that programmatic, community-based health and social service interventions have a positive impact on client well-being. These proactive interventions, designed to address the full range of health and social needs, were usually provided at the same--or even lower--costs as uncoordinated, illness-focused care. The results of this series suggest that across-the-board health care reduction, at least in a system of national health insurance, will produce poorer results, at higher cost, for people with chronic conditions living in the community. Policy planners need more research that concentrates on comparisons of outcomes between and within different models of health and social service delivery. The studies should be designed to help them determine who benefits from different service configurations carried out within a range of policy environments at various costs. PMID- 10526550 TI - What is right about the Canadian health care system? AB - Canadians tend to dwell on problems in their health care system, looking to the United States for magical fixes. Evidence on comparative system performance, which rarely surfaces in public debate, indicates that Canadians are healthier, not only because the social environment is more benign, but also because health care is allocated by need rather than ability to pay. Expenditures are much lower, but Canadians receive equivalent care because their system is more efficient. Although Canadian "waiting lists" are highly publicized, the United States avoids the issue by excluding those who cannot pay. Why, then, do American notions keep pushing north? All expenditures are someone's income. There is a great deal of money to be made by wrecking Canadian Medicare. PMID- 10526551 TI - What is wrong with the U.S. health care system?: It does not effectively exist for one of every five Americans. PMID- 10526552 TI - Canadian health care and its vulnerabilities. PMID- 10526553 TI - [Pneumococcal infections in intensive care. Retrospective 8 year study]. AB - OBJECTIVE: The aim of this study was to analyze the clinical presentations and severity of S. pneumoniae infections requiring hospitalization in an intensive care unit and evaluate the incidence and severity of infections caused by penicillin-resistant strains. PATIENTS AND METHODS: This retrospective study reviewed cases in our intensive care unit from January 1989 through December 1996 including all patients with pneumococcal infection. RESULTS: The study included 102 patients, mean age 59.6 years. Pneumonia was the most frequent (83 cases) followed by bacteriemia (31 cases) and meningitis (15 cases). Mortality was high (43%) and influenced by age, simplified severity score, and presence of shock at admission. Antibiotic resistance appeared in 1991 and increased over the years reaching, in 1996: 24% for penicillin, 38% for macrolides, 20% for sulfamides, 19% for tetracyclins, and 14% for phenicols. Penicillin-resistance was not found to modify clinical expression nor severity of infection. Amoxicillin and third generation cephalosporins were the most widely used antibiotics. CONCLUSION: Pneumococcal infections in intensive care patients are severe with high mortality. The emergence of more and more resistant strains has little clinical consequence on severity or treatment. PMID- 10526554 TI - [Black esophagus related to acute esophageal necrosis: a new case]. AB - BACKGROUND: Endoscopic discovery of a black esophagus due to acute necrosis is quite exceptional excepting cases of poisoning. CASE REPORT: A 48-year-old man suffered acute necrosis of the esophagus giving a black aspect at endoscopy. Soon after the exploration he developed a state of shock with hematemesis related to a bulbar ulcer. The patient was given a short regimen of parenteral nutrition and sodium pump inhibitors and the course was rapidly favorable both locally and generally. DISCUSSION: There have been 27 such cases reported in the literature. Ischemia appears to be the main pathogenic mechanism. Prognosis depends on the patientis general status rather than the aspect of the esophageal lesions. PMID- 10526555 TI - [Cerebral intravascular lymphoma during T CD4+ idiopathic lymphopenia syndrome]. AB - BACKGROUND: Intravascular lymphoma is a proliferation of lymphoid cells, usually B cells, in small vessels, predominantly in the nervous system and skin. CASE REPORT: We report a case of a man with a 3-year history of lymphopenia with no detectable etiology (all viral causes were ruled out) who developed intravascular lymphoma in the cerebral vessels. DISCUSSION: This case was particular as the patient had idiopathic CD4+ lymphopenia. It points out the probable role of immunodepression in the development of lymphomas, particularly in endovascular localizations. PMID- 10526556 TI - [A case of septicemia manifesting as a black esophagus]. PMID- 10526557 TI - [No N-acetylcysteine in the absence of verified paracetamol intoxication]. PMID- 10526558 TI - [Treatment of herpes zoster infections in patients infected with HIV]. PMID- 10526559 TI - [Muscle contractures. Essay on a physiopathological approach to clarify the nomenclature]. AB - There is a good deal of confusion in the international literature concerning the use of the term contracture. A pathophysiological approach would help in defining a more precise terminology. Antalgic contracture principally concerns a compensating phenomenon related to a polysynaptic reflex Painful contracture includes different varieties of cramps and metabolic-induced contractures. Painless contracture groups together myostatic and myostatic contractures. We suggest a simplified nomenclature. PMID- 10526560 TI - [Re-examination of the bioethics law of 1999: reflections and propositions. Group on Ethical Aspects of Transplantation]. AB - THE FRENCH LAW ON BIO-ETHICS BEING REVISITED: The GRET (Group on ethical aspects of transplantation) makes a series of proposals in order to update some articles of the law, this being considered as a considerable progress. BONE MARROW GRAF: The new concept of graft of "Hematopoietic stern cells" including their different sources, better reflects the current practice, remaining considered basically as an organ. ORGAN TRANSPLANTATION, WITH LIVING DONOR: The extension to new categories of donors is proposed, with, however, the creation of a multidisciplinary committee of experts and representatives of the society in charge of the evaluation of the motivations, with objective criteria. ORGAN TRANSPLANTATION, DECEASED DONOR: A better information of the public, together with the help of Associations is proposed, and the creation of help units in order to provide support for family members, who come in emergency as witnesses and greatly need support and help. PMID- 10526561 TI - [Zolmitriptan]. PMID- 10526562 TI - [Recommendations for drug treatment of type 2 diabetes (I). Normalizing glucose levels. French Agency for Health Product Safety]. PMID- 10526563 TI - [Concerning the recommendations for drug treatment of type 2 diabetes (I). An interview with Pr S. Halimi]. PMID- 10526564 TI - [Action of bisphosphonates on malignant osteolysis]. AB - MECHANISM OF BONE DESTRUCTION IN MALIGNANT OSTEOLYSIS: Many mechanisms have been described to explain the excessive osteoclastic activity: local stimulation factors (cytokines, lymphokines) are mainly found in breast cancer of myeloma; a general stimulation factor (PTH rP) is found predominantly in lung cancer, head and neck cancer and in cancer of the kidney or ovary. CONTRIBUTION OF BISPHOSPHONATES: The use of bisphosphonates is warranted to counteract the overactivity of the osteoclasts in humans, especially since these drugs could have a direct effect on cancer cells and also have their own therapeutic effect. There are four objectives for using bisphosphonates in cancer patients: lowering serum calcium levels, pain relief, treatment and prevention of bone metastasis. Four bisphosphonates have marketing approval in France for this indication. PROVEN AND TO BE PROVEN EFFECTS: The serum calcium lowering effect and the curative effect for the treatment of bone metastasis are well documented for all four formulations which all meet the clinical requirements for marketing approval. The preventive effect on the development of secondary bone localizations of primary cancers remains to be demonstrated and will require extensive testing in humans. There is however room for hope of substantial progress in cancerology. PMID- 10526565 TI - [Primary small-cell bronchial cancer: value of serum tumor markers in the prognostic evaluation]. AB - TUMOR MARKERS: We reviewed the biomedical literature searching for the most well documented serum markers with prognostic value independent of the usual radioclinical and histological parameters of small cell lung cancer. RELATIVE PROGNOSTIC VALUES: Neuron specific enolase (NSE) would have a pretherapeutic prognostic value better than LDH (lacto-dehydrogenase) and perhaps better than serum sodium, biocarbonates, and uric acid. The superiority of NSE over serum albumin remains to be proven. Although there are only a few reports, TK (thymine kinase), TPA (tissue polypeptide antigen), Cyfra 21-1 and/or IL-2 (interleukin-2) secretion might have better pretherapeutic prognostic value than NSE. We also found that iterative blood assays to follow therapeutic effects in patients with small-cell lung cancer have not been proven to provide independent prognostic information. CONCLUSION: As medical laboratory resources must be concentrated on priority exploitable assays, the currently available data would suggest that it is not necessary to routinely measure serum tumor markers, and in particular NSE, for the prognostic evaluation of small-cell lung cancer patients. PMID- 10526566 TI - [Cerebral venous thromboses]. AB - VARIABLE EXPRESSION: The clinical expression of cerebral venous thrombosis varies widely. The only manifestation may be an intracranial hypertension if the thrombus is limited to the superior longitudinal sinus or a predominant lateral sinus. Thrombosis of cortical veins, alone or in association with a sinus thrombus causes venous infarction. PROGNOSIS: Deep venous thrombosis generally leads to coma with signs of intracranial hypertension. However, partial or complete recovery is possible, even with a severe initial presentation, underlining the importance of early diagnosis and treatment. DIAGNOSTIC METHODS: Magnetic resonance imaging (MRI) coupled with magnetic resonance angiography (MRA) can usually confirm the diagnosis without conventional angiography. ETIOLOGICAL DIAGNOSIS: Cerebral venous thrombosis often follows prethrombotic conditions. Infectious causes only account for 10% of the cases and no cause can be identified in about 1 out of 5 cases. TREATMENT: Anticoagulation is indicated even in case of venous infarction with spontaneous bleeding. Thrombolysis can be proposed, particularly for deep venous thrombosis. PMID- 10526567 TI - [New food allergies]. AB - RISING INCIDENCE OF FOOD ALLERGIES: Food allergies are becoming more and more common, concerning 3 to 4% of the general population. One out of four persons allergic to nuts, the most frequent food allergen, have severe signs and symptoms. A CLASSICAL DIAGNOSIS: Certain diagnosis of food allergy is established on the basis of labial and oral tests. The dose required to induce a reaction is established by the oral test, giving information about the severity of the allergy and its progression. OTHER ALLERGENS: "Emerging" food allergens include spices and condiments, exotic fruits (kiwi, avocado, cashew and pecan nuts, Brazil nuts), sesame seeds, psyllium, sunflower seeds. Endurance exercise following ingestion of a food allergen can lead to severe anaphylactic reactions. Allergen associations "food-pollen", "latex-food", "mitessnails" have been described. INDISPENSABLE PREVENTION: Avoiding contact is essential. Many allergens are "masked" within prepared foods. Precise labeling, with particular attention to nut content, must be reinforced. Individualized counseling on food allergies should be available for school children. Persons with severe allergies should keep at hand an emergency kit with antihistamines, injectable rapid action corticoids and adrenalin (1 mg/ml). PMID- 10526568 TI - Introduction: STATs as essential intracellular mediators of cytokine responses. PMID- 10526569 TI - Interferons as a paradigm for cytokine signal transduction. AB - Characterization of the ability of interferons to induce immediate early genes led to the identification of the signal transducer and activators of transcription (STAT) signaling paradigm, where a single protein transduces signals directly from the receptor to the nucleus. Subsequent studies have determined that all cytokines transduce pivotal signals through at least one of the seven members of this STAT family. Notably, cytokines can be placed in functional subgroups based on the STATs they activate. PMID- 10526570 TI - The Janus kinase family of protein tyrosine kinases and their role in signaling. AB - In the early 1990s, the search for protein kinases led to the discovery of a novel family of non-receptor tyrosine kinases, the Janus kinases or JAKs. These proteins were unusual because they contained two kinase homology domains and no other known signaling modules. It soon became clear that these were not 'just another' type of kinase. Their ability to complement mutant cells insensitive to interferons and to be activated by a variety of cytokines demonstrated their central signaling function. Now, as we approach the end of the decade, it is evident from biochemical studies to knockout mice that JAKs play non-redundant functions in development, differentiation, and host defense mechanisms. Here, recent progress is reviewed, with particular emphasis on structure-function studies aimed at revealing how this family of tyrosine kinases is regulated. PMID- 10526571 TI - Transcription factor activity of STAT proteins: structural requirements and regulation by phosphorylation and interacting proteins. AB - The seven mammalian members of the signal transducer and activator of transcription (STAT) family share a common core structure which reflects their shared mechanism of activation, dimerization, and DNA binding. By contrast, the STAT C termini containing the sequences required for transcriptional activation are much less homologous, suggesting different ways by which individual STATs activate their target genes. This paper describes several important discoveries linked to mechanistic aspects of STAT transcription factor function. These include regulated serine phosphorylation of the transactivating domain, promoter dependent interactions of STATs with each other, or of STATs with other transcription factors, and with transcriptional co-activators. The basis, background, and implications of these molecular events will be summarized and discussed. PMID- 10526572 TI - The role of STATs in proliferation, differentiation, and apoptosis. AB - The spectrum of biological systems which makes use of the signal transducers and activators of transcription (STAT) paradigm extends beyond the interferon system in which it was first discovered to include many other cytokines and agonists. Having catalogued which STATs are activated by each stimulus, investigators have turned their attention to defining the biological processes and the genes regulated by the STAT pathway. These studies are in their early stages. Although many tools have been developed to probe the STAT pathway, e.g., mutant receptors, dominant-negative STATs, chemically dimerizable STATs, and mice lacking STAT proteins, more is known about the biological phenomenon affected than the molecular mechanism or the STAT-regulated genes involved. The cellular events currently believed to utilize STAT-dependent pathways can be grouped according to those which affect cell growth, differentiation, and apoptosis. PMID- 10526573 TI - Physiological significance of STAT proteins: investigations through gene disruption in vivo. AB - Signal transducers and activators of transcription (STATs) were discovered as mediators of type I interferon-induced gene expression. This family of transcription factors has been found in widespread signaling pathways, especially those involving cytokines regulating the immune response. Because a plethora and often confusing set of activators for STAT proteins was observed in cell culture models, it became important to define the physiologically relevant actions of these molecules. One approach to this question has been through the targeted disruption of STAT genes in transgenic mice. Now that all seven STAT genes have been disrupted, both the high degree of STAT selectivity as well as many surprising and unexpected complexities are beginning to be characterized. PMID- 10526574 TI - Negative regulators of cytokine signal transduction. AB - Enormous advances have been made over the last 10 years in unravelling cytokine signal transduction. This work has led to the recognition of the prime importance of Janus kinases (JAKs) and signal transducers and activators of transcription (STATs). More recently, the importance of negative regulators of this pathway has begun to be realised. There is now evidence for at least three families of proteins that inhibit JAK/STAT signalling. The suppressors of cytokine signalling (SOCS), protein inhibitors of activated STATs (PIAS) and the SH2-containing phosphatase (SHP-1). This review describes some of the key features of SOCS proteins and contrasts their actions with other negative regulators, the PIAS proteins and SHP-1. PMID- 10526575 TI - JAKs and STATs in invertebrate model organisms. AB - Invertebrate organisms provide systems to elucidate the developmental roles of Janus kinase (JAK)/signal transducers and activators of transcription (STAT) signaling pathways, thereby complementing research conducted with mammalian cells and animals. Components of the JAK/STAT protein pathway have been identified and characterized in the fruit fly Drosophila melanogaster and the cellular slime mold Dictyostelium discoideum. This review summarizes the molecular and genetic data obtained from these model organisms. In particular, a Drosophila JAK/STAT pathway regulates normal segmentation, cell proliferation, and differentiation, and hyperactivation of the pathway leads to tumor formation and leukemia-like defects. A Dictyostelium STAT regulates the development of stalk cells during the multicellular part of the life cycle. Future research utilizing these organisms should continue to provide insights into the roles and regulation of these proteins and their signaling pathways. PMID- 10526576 TI - Novel approaches to the treatment of small-cell lung cancer. AB - Small-cell lung cancer (SCLC) is characterized by its initial responsiveness to chemotherapy and the appearance of early metastases. Although combination chemotherapy, in some instances together with radiation, has improved the prognosis of this disease, in most patients SCLC ultimately recurs in a drug resistant form. Several new strategies for the eradication of SCLC are being explored at the preclinical level. The identification of selective target molecules on the surface of SCLC cells, together with the progress made in antibody engineering, have provided new generations of antibodies and immunoconjugates as well as growth factor antagonists and inhibitors. In addition, recent advances in understanding the biology of SCLC have stimulated new investigations searching to counter the molecular basis underlying the increased proliferation and the apoptosis deficiency of SCLC cells. This can be achieved using antisense oligodeoxynucleotides that repress the expression of growth factor receptors and anti-apoptosis genes, or by gene replacement to compensate for the loss or inactivation of tumor suppressor genes. PMID- 10526577 TI - Anthrax toxins. AB - Though its lethal effects were ascribed to an exotoxin almost half a century ago, the pathogenesis of anthrax has yet to be satisfactorily explained. Subsequent work has led to the molecular identification and enzymatic characterization of three proteins that constitute two anthrax toxins. Protective antigen binds an as yet unknown cell receptor and mediates the entry of the other two components to the cytoplasm via the endosomal pathway. Edema factor, so named for its ability to induce edema, is a Ca2+/calmodulin-dependent adenylate cyclase. Lethal factor, the dominant virulence factor associated with the toxin, proteolytically inactivates mitogen-activated protein kinase kinases, key players in signal transduction. We describe the fascinating work that has led to these discoveries and discuss their relevance to our understanding of the pathogenesis of anthrax. PMID- 10526578 TI - Interleukin-12, a key cytokine in Th1-mediated autoimmune diseases. AB - Interleukin 12 (IL-12) is a heterodimeric cytokine produced primarily by antigen presenting cells (APCs) which plays a key role in promoting type 1 T helper cell (Th1) responses. The powerful activity of IL-12 requires tight control, which is exerted at various levels. Primary control is exerted on IL-12 production by APCs, a major factor driving the response towards the Th1 or Th2 phenotype. Another level of control regulates expression of the IL-12 receptor (IL-12R), which is composed of two subunits, beta 1 and beta 2. The IL-12R beta 2 subunit has signal-transducing capacity and modulation of its expression is central to the regulation of IL-12 responsiveness. Endogenous IL-12 plays an important role in host defense against infection by a variety of intracellular pathogens. Its Th1-promoting activity, however, also favors Th1-mediated immunopathology and, in particular, the induction of Th1-mediated autoimmune diseases. PMID- 10526579 TI - Minisatellite instability and germline mutation. AB - Tandem-repeat DNA actively turns over in the genome by a variety of poorly understood dynamic mechanisms. Minisatellites, a class of tandem repeats, have been shown to cause disease by influencing gene expression, modifying coding sequences within genes or generating fragile sites. There has been recent rapid progress towards understanding molecular turnover processes at human minisatellites. Instability at GC-rich minisatellites appears to involve distinct mutation processes operating in somatic and germline cells. In the germline, complex conversion-like events occur, probably during meiosis. Repeat turnover appears to be controlled by intense recombinational activity in DNA flanking the repeat array, suggesting that minisatellites might evolve as by-products of localised meiotic recombination in the human genome. In contrast, AT-rich minisatellites appear to evolve by intra-allelic processes such as replication slippage. Curiously, minisatellites in other organisms appear to be more stable than their human counterparts, suggesting species-specific differences in turnover processes. Some yeast models display human-like minisatellite turnover processes at meiosis. However, all attempts to transfer human germline instability to transgenic mice have failed. Finally, tandem repeat instability in various species appears to be extremely sensitive to environmental agents such as radiation via a mechanism which remains enigmatic. PMID- 10526580 TI - Dexamethasone enhances CTLA-4 expression during T cell activation. AB - T cell activation is enhanced by the costimulatory interaction of B7 on antigen presenting cells and CD28 on T cells, resulting in long-term T cell proliferation, differentiation and production of large amounts of cytokines, such as interleukin (IL)-2. CTLA-4 is a co-stimulation receptor that shares 31% homology with CD28 and binds B7 family members with higher affinity. CTLA-4 is transiently expressed intracellularly and on the cell surface following activation of T cells. We have studied the kinetics of CTLA-4 expression and the effects of dexamethasone on CTLA-4 expression during T cell activation in cultures of mouse spleen cells stimulated by a mixture of immobilized anti-CD3 and anti-CD28 monoclonal antibodies (anti-CD3/CD28 mAb) or concanavalin A (ConA). CTLA-4 expression peaked on day 2 and returned to background levels after 7 days. Dexamethasone was found to potentiate CTLA-4 expression in a dose-dependent manner with an EC50 effective concentration 50%) of about 10(-8) M. In contrast, other immunosuppressive agents, such as rapamycin or cyclosporin A had no or an inhibitory effect on CTLA-4 expression, respectively. Dexamethasone also stimulated CD28 expression, but inhibited IL-2R expression during anti-CD3/CD28 mAb-induced mouse splenic T cell activation. Western blot analyses of lysates of activated mouse T cells showed that dexamethasone increased CTLA-4 protein levels twofold during anti-CD3/CD28 mAb-induced activation. Dexamethasone also enhanced CTLA-4 messenger RNA twofold as quantified by ribonuclease protection assay. The effects of dexamethasone on CTLA-4 expression were glucocorticoid-specific and completely inhibited by the glucocorticoid receptor antagonist mifepristone (RU486), indicating that the effect of dexamethasone on CTLA-4 expression is mediated through the glucocorticoid receptor. In conclusion, the immunosuppressive agent dexamethasone actually stimulates CTLA-4 expression, which is involved in downregulation of T cell activation. PMID- 10526581 TI - [Psychiatric consultation and treatment for mentally handicapped persons exhibiting behavioral changes]. AB - In three mentally handicapped people, two women aged 47 and 68 years respectively and a man aged 68, who suffered from behavioural changes that were not understood by the staff of the institution where the people lived, a psychiatric diagnosis was made by a consulting psychiatrist. The first woman had Down syndrome, she suffered from weight loss, loss of enjoyment and severe hallucinations. She was treated for a depressive disorder and recovered. The second woman yelled and threatened to hit the nursing staff. A bipolar condition was diagnosed and after unsuccessful drug treatment she was treated with electroconvulsion therapy upon which she recovered. The man had developed restlessness and verbal aggression with megalomanic episodes. A mood disorder was diagnosed which responded to valproic acid. In people with a mental handicap psychiatric disorders can be easily missed. The disorder can be complicated by an atypical presentation of symptoms, difficulty in obtaining information and limited knowledge and organization of the psychiatric services. Psychiatric consultation in people with mental retardation may lead to diagnosis and treatment of a psychiatric disorder. PMID- 10526582 TI - [Prenatal typing of Rh- and Kell- blood group system antigens]. AB - Rhesus (Rh) and Kell blood group immunisations are the most frequent causes of haemolytic disease of the newborn. Recently, the molecular bases of the Rh and Kell antigens have been elucidated. Subsequently, specific polymerase chain reactions (PCRs) could be developed to determine the RhD, RhC/Rhc and RhE/Rhe genotypes as well as the KI genotype (from the Kell blood group) with genomic DNA. The tests were applied to genomically determine the foetal Rh and Kell blood groups with DNA obtained from amniotic fluid cells. The genotypes obtained were compared with the Rh phenotypes established by cord blood red cell serology. The PCRs to determine the RhD, Rhc, RhE and Rhe and KI genotypes were found to be reliable. The test for RhC however, resulted in false-positive C genotypes. Indeed, more than half of the subsequently tested C-negative Negroid donors were false-positive with the DNA test. Thus, except for RhC, it is possible to reliably determine the Rh and KI genotypes of a foetus with DNA isolated from amniotic fluid cells. Amniocentesis, however, carries a risk for the pregnancy and therefore the tests will only be justified in pregnant women in whom an antibody has been detected and the father of the foetus is heterozygous for the specific antigen. Recently foetal RhD genotypes were determined in foetal DNA circulating in the plasma of RhD-negative pregnant women. This could eventually lead to the introduction of assays with which the foetal blood group can be determined without any risk to the foetus. PMID- 10526583 TI - [Antibiotic prophylaxis of hematogenous bacterial arthritis]. AB - The outcome of bacterial arthritis is generally poor: the mortality is 10-15% and there is loss of joint function in 25-50% of the survivors. Adverse prognostic factors are advanced age, a pre-existent joint disease and an infection of a prosthetic joint. The incidence of bacterial arthritis is low: 2-6 per 100,000 persons per year. Risk factors are advanced age, a joint disease--especially rheumatoid arthritis--diabetes mellitus and presence of a prosthetic joint. Situations that can lead to bacterial arthritis are mainly skin infections of the feet and only rarely invasive medical or dental procedures. Because of the severity of the disease, antibiotic prophylaxis of haematogenous bacterial arthritis in patients with prosthetic joints is advocated in guidelines. However, because of the rarity of the disease it is unclear whether the advantages of prophylaxis outweigh the disadvantages of the large-scale use of antibiotics, such as side effects, costs and increased resistance of bacteria. In a decision analysis of a large group of patients with joint diseases, antibiotic treatment of skin infections appeared to be (cost-)effective in the prevention of haematogenous bacterial arthritis, mainly in high-risk patients. On the other hand, prophylaxis around medical or dental procedures was not (cost-)effective, except possibly in a small group of patients with increased risk. PMID- 10526584 TI - [Physical examination--percussion of the thorax]. AB - Percussion of the chest cannot detect small lesions (< 3 cm in diameter) and lesions situated deep in the lung (5-7 cm away from the outside of the thoracic wall). The value of percussion as a test to detect a pathological pulmonary situation in comparison with chest radiography is evaluated in only a few studies. Chest percussion appears to be of limited value; especially the sensitivity is very low. Moreover, the inter- and intra-observer errors are high. The value of percussion to detect cardiomegaly is only investigated in one recent study in a selected patient population. In this study the predictive values as well as the reproducibility appear to be moderate. PMID- 10526585 TI - [Child with tubercular meningoencephalitis, etiologically related to a microepidemic of tuberculosis in Southern Limburg by DNA 'fingerprinting']. AB - MICROEPIDEMIC: In a child fatal tuberculous meningoencephalitis was diagnosed and the Regional Public Health Service Geleen, Limburg, the Netherlands, was notified. Source identification and contact tracing (ring investigation) did not reveal a source of the infection. In a person living in the same village pulmonary tuberculosis had been diagnosed, but there was no evident contact between both patients. When the mycobacteria from all patients with tuberculosis were typed by DNA fingerprinting, both patients belonged to the same cluster, thus identifying the infection source of the meningoencephalitis patient. In the management of the outbreak 950 persons were examined in a contact tracing survey. Of them 35 had recently been infected and four of these had recently acquired pulmonary tuberculosis. DISCUSSION: Highly infectious patients with tuberculosis are able to infect persons who cannot be found by conventional contact tracing survey as the transmission of tuberculosis is more subject to casual encounters than was hitherto believed. DNA fingerprinting is a very useful method in contact investigation of tuberculosis. In the Netherlands, therefore, the early diagnosis and treatment of symptomatic patients with infectious tuberculosis is more important to stop the transmission of Mycobacterium tuberculosis than the identification and screening of risk groups in the population. PMID- 10526586 TI - [Use of noninvasive mechanical ventilation to avoid intubation during acute respiratory insufficiency]. AB - OBJECTIVE: To determine the value of noninvasive mechanical ventilation in patients with acute respiratory insufficiency. DESIGN: Descriptive. METHODS: Noninvasive mechanical ventilation was considered in all patients with acute respiratory insufficiency in the intensive care unit of the Rijnstate Hospital, Arnhem, the Netherlands, between 1 June 1998 and 31 January 1999. Indication for mechanical ventilation was: respiratory frequency > or = 28/min, and PaCO2 > 6.0 kPa, pH < 7.35, and/or PaO2 < 8.0 kPa. Patients were intubated immediately in case of systolic blood pressure < 90 mmHg, cardiac or respiratory arrest, coma or a severely diminished consciousness. The other patients received noninvasive mechanical ventilation by nasal or full face mask (pressure support: 10-20 cmH2O; positive end expiratory pressure (PEEP): 0-5 cmH2O). Patients were intubated if the respiratory frequency or the level of consciousness or a blood gas value deteriorated. RESULTS: Of the 97 patients who needed ventilation support, 67 were immediately intubated. Noninvasive mechanical ventilation was administered in the other 30 (31%) patients, 22 men and 8 women with a mean age of 67 years (SD: 15). Causes for acute respiratory failure were: chronic obstructive pulmonary disease (COPD) (n = 12); pneumonia (n = 6); heart failure (n = 4); and other (n = 8). Median characteristics at baseline in the noninvasive mechanical ventilation group: acute physiology and chronic health evaluation (APACHE) II score: 17 (range: 1-25); respiratory frequency: 30/min (28-59); pH: 7.33 (6.99-7.54); PaCO2: 9.3 kPa (3.2-18.7); PaO2: 7.4 kPa (4.2-13.4); SaO2: 85% (63-95). Intubation was avoided in 9/30 (30%) of all patients and in 7/12 (58%) of the patients with COPD. Intubation was needed in 21/30 patients (70%): in 8/30 (27%) immediately because of clinical deterioration within 2 hours and in 13/30 (43%) after a mean period of stabilisation of 6 hours (2-16). CONCLUSION: Noninvasive mechanical ventilation prevented intubation in over half of the selected patients presenting with acute respiratory failure due to an exacerbation of COPD. The method appears to be less successful in acute respiratory failure due to other causes. PMID- 10526587 TI - [Two neonates with severe rhesus antagonism inspite of low values in recent tests of antibody dependent cellular cytotoxicity (ADCC)]. AB - In two female newborn babies severe haemolytic disease of the newborn developed due to anti-RhD antibodies of the mother. In both cases the results of the last antibody-dependent cellular cytotoxicity (ADCC) test had been < 10% (two and three weeks before parturition respectively) making haemolytic disease unlikely. The ADCC test is used to determine the destructive power of the responsible maternal antibodies. It is believed that with a low ADCC value (< 10%), there is no danger of clinically relevant haemolytic disease. The first neonate recovered after exchange transfusion, the second died notwithstanding extensive supportive therapy. A recent low ADCC value makes serious haemolytic disease unlikely, but does not completely exclude it, as the titre of the causative antibodies can rise very quickly. PMID- 10526588 TI - [Disclosure of medical information after a patient's death: principles and recent developments in jurisprudence]. AB - After a patient's death, the obligation of medical secrecy remains in force; the physician is still bound to confidentiality and when necessary should invoke his right to remain silent. However, it is generally accepted that circumstances can occur in which a doctor may disclose confidential information to third parties like the relatives Inspection of the medical file after the patient's death is subject to the following principles. Disclosure of medical data after a patient's death is--apart from the general exceptions on medical secrecy--justified when the wish of the deceased can be reconstructed, or (if this is impossible) if there are such important interests of third parties involved that the obligation to maintain confidentiality may be put aside. The relatives have no personal right of access to medical data of the deceased, and they cannot authorize disclosure of data to other parties (e.g. the attorney of the hospital). With regard to the ex-mentor it seems as if the traditional opinion on medical secrecy after a patient's death has been abandoned. In principle, the physician should inform this person, unless it can be proven that disclosure is in conflict with the wish of the deceased. PMID- 10526589 TI - [Foregoing artificial feeding and hydration for nursing home patients in the last phase of life]. PMID- 10526590 TI - [Foregoing artificial feeding and hydration for nursing home patients in the last phase of life]. PMID- 10526591 TI - [Foregoing artificial feeding and hydration for nursing home patients in the last phase of life]. PMID- 10526592 TI - [Ventilator-associated pneumonia; controversies with respect to diagnosis, pathogenesis, therapy and prevention]. PMID- 10526593 TI - [Selective decontamination of the digestive tract reduces mortality in intensive care patients]. PMID- 10526594 TI - [Rubber band ligation of hemorrhoids: symptoms almost gone after 6 weeks, but many patients need retreatment in the long run]. PMID- 10526595 TI - [Toxic shock-like syndrome caused by streptococci]. PMID- 10526596 TI - [Bilateral blood pressure measurement before and after coronary bypass surgery: an absolute necessity]. AB - Anginous symptoms and a difference in blood pressure between the two arms prompted angiography in two patients, men aged 66 and 50 years. The examination revealed coronary sclerosis and a stenosis in the left subclavian artery. The symptoms disappeared after percutaneous dilatation of the subclavian artery, followed by a coronary bypass operation (CABG) using an internal thoracic artery (a branch of the subclavian artery). In two other patients, men aged 61 and 71 years, who had undergone an arterial CABG 12 years previously, anginous symptoms were the manifestation of a narrowed subclavian artery. The symptoms disappeared after balloon dilatation of the subclavian artery and revascularization of the anterior interventricular branch (left artery descendens) and embolization of the internal thoracic artery graft (internal mammarian artery graft), respectively. Stenosis or occlusion of the proximal subclavian artery may attenuate the blood flow in the ipsilateral A. thoracica interna graft. The diagnosis can simply be made by bilateral blood pressure measurement. PMID- 10526597 TI - [Sexual side effects of antidepressants]. AB - Most antidepressant drugs have some sexual side effects. These side effects are caused by an increased neurotransmission of serotonin in the central nervous system, a stimulating effect at postsynaptic serotonin receptors and effects on the peripheral nervous system. Antidepressant-induced sexual disorders may impair patient compliance. Publications on sexual side effects are mainly case reports and few double blind placebo-controlled trials. This hampers generalisation of findings. The best treatment consists of prescribing another antidepressant which probably has hardly any or no sexual side effects. However, this entails the risks of reduced antidepressant effects and of non-sexual side effects. The extent of sexual side-effects among the serotonin reuptake inhibitor seems to differ. Antidepressants with few or no sexual side effects include those with noradrenergic and dopaminergic action (some tricyclic antidepressants, bupropion), as well as antidepressants with a postsynaptic receptor blocking action (nefazodone and mirtazapine). PMID- 10526598 TI - [Chronic hepatitis-b-virus infections: new options for antiviral therapy]. AB - The possibilities for antiviral treatment of patients with chronic hepatitis B virus (HBV) infections have been in flux for the last few years. Apart from interferon alpha, oral nucleoside analogues are given a place. The treatment focuses on the group of patients with active virus multiplication (hepatitis B e antigen (HBeAg) and HBV DNA are demonstrable), abnormal liver enzyme values and histologically demonstrated inflammatory activity in the liver. For the individual patient the pros and cons of protracted virus inhibition should be weighted by means of an orally administered nucleoside analogue such as lamivudine, immunostimulation with injections of interferon alpha or a combination of these two. PMID- 10526599 TI - [Physical examination--the significance of Homan's sign]. AB - Homans's sign is often used in the diagnosis of deep venous thrombosis of the leg. A positive Homans's sign (calf pain at dorsiflexion of the foot) is thought to be associated with the presence of thrombosis. However, Homans's sign has a very poor predictive value for the presence or absence of deep vein thrombosis, like any other symptom or clinical sign of this disease. PMID- 10526600 TI - [Hyperthermic intra-peritoneal chemotherapy (HIPEC) in patients with peritoneal pseudomyxoma or peritoneal metastases of colorectal carcinoma; good preliminary results from the Netherlands Cancer Institute]. AB - OBJECTIVE: To apply (HIPEC) using mitomycin in patients with peritoneal carcinomatosis. DESIGN: Descriptive. METHOD: The HIPEC treatment includes cytoreductive surgery and subsequent peritoneal lavage with exposure of the superficial tumour residues to a high concentration of a cytostatic drug at an intra-abdominal temperature of 40-42 degrees C. The HIPEC treatment was given to 24 patients with pseudomyxoma peritonei and to 29 patients with peritoneal carcinomatosis of colorectal origin. The adequate dose of mitomycin was determined in the Netherlands Cancer Institute in 26 patients: doses of 15 mg/m2 (n = 8), 25 mg/m2 (n = 3), 35 mg/m2 (n = 7) and 40 mg/m2 (n = 8) were administered. RESULTS: The maximal tolerable dose of mitomycin appeared to be 35 mg/m2, while unacceptable toxicity was recorded at a dose of 40 mg/m2. Therefore 35 mg/m2 was used in subsequent cases. Among the patients with pseudomyxoma peritonei 8 developed severe complications, two of which were fatal. After a median follow-up of 12 months, 21 patients were alive of whom 18 were free of disease. Among the patients with peritoneal carcinomatosis of colorectal origin one patient died from a treatment-related complication. After a median follow-up of 18 months, 18 patients were alive of whom 11 were free of disease. The actuarial 2 year survival was 59%. CONCLUSION: In the Netherlands Cancer Institute HIPEC treatment is considered the treatment of choice for pseudomyxoma peritonei. The results in cases of peritoneal carcinomatosis of colorectal origin are promising, but the results of a randomized trial are awaited. PMID- 10526602 TI - [Delayed (tension) pneumothorax after placement of a central venous catheter]. AB - Laborious attempts at introducing a central venous catheter for parenteral nutrition in two women, aged 36 and 62 years, were followed by shortness of breath after 32 and 10 hours, respectively. This symptom was due to a (tension) pneumothorax not visible on earlier roentgenograms. Thoracic drainage led to recovery. In all patients with a central venous catheter an undetected delayed pneumothorax can be present. Urgent chest X-ray examination should be performed in all patients with acute respiratory symptoms. Patients undergoing elective intubation with positive pressure breathing should be examined carefully, since they are at risk of developing a late (tension) pneumothorax. PMID- 10526601 TI - [Diurnal triglyceride profiles in 30 young healthy men as a function of diet, fasting triglyceride levels, body composition and insulin sensitivity]. AB - OBJECTIVE: To study predictors of diurnal capillary triglyceride (TG-c) profiles in healthy males. DESIGN: Observational, cross-sectional. SETTING: University Hospital Utrecht, Department of Internal Medicine, the Netherlands. METHOD: In 30 healthy males (20-34 years) TG-c was measured during three days, six times a day. Fasting blood was collected at inclusion. Body composition and HOMA ratio as insulin sensitivity index were determined. TG-c profiles were calculated as integrated area under the mean TG-c curve (TG-AUC). RESULTS: All subjects had normal fasting plasma and capillary TG and cholesterol concentrations. Diurnal TG c values were higher than fasting values. The average TG-AUC was 24.6 +/- 6.7 mmol/l over 14 hrs. Variables associated with TG-AUC were: fasting TG-c, relative fat mass, total protein and saturated fat intake. After correction for fasting TG c only diet and fat mass were correlated with TG-AUC. The relative fat mass was positively correlated with the HOMA ratio and fasting insulin concentrations, suggesting that decreased insulin sensitivity accompanied increased body fat. CONCLUSION: Triglyceride profiles provide information about the total diurnal TG load. The best determinant of diurnal triglyceride changes was fasting triglycerides. However, diet, body composition and insulin sensitivity are also important. Future investigations should address the question whether triglyceride profiles may be used to estimate more accurately the individual risk profile for coronary heart disease. PMID- 10526603 TI - [Milestones in prevention in the past century]. AB - The prevention of disease and premature death expanded enormously in the 20th century, greatly changing the pattern of disease and death in the Netherlands. This is most clearly apparent where infectious diseases are concerned. In part, the improvement of public health is not a direct result of specific programmes against specific health problems but of general improvement in socioeconomic circumstances. The other part of the improvement is mostly due to specific prevention both within and without the institutionalized health care. Within public health care this means, for instance, prenatal and adolescent health care. Outside health care it includes activity in the areas of hygiene, lifestyle and safety. PMID- 10526604 TI - [Discordant fetal growth in multiple pregnancy: intervention should be based on chorionicity]. PMID- 10526605 TI - [Tingling in the hands]. PMID- 10526606 TI - [Role of allergy in interstitial cystitis]. PMID- 10526607 TI - [The role of allergy in interstitial cystitis]. PMID- 10526608 TI - [Primary HIV-infection: infectious mononucleosis-like presentation with treatment options]. PMID- 10526609 TI - [Chromosomal deviations in subfertile men and their partners is often not a reason to refrain from intracytoplasmic sperm injection]. PMID- 10526611 TI - [Gastrointestinal surgery and gastroenterology. I. Introduction]. AB - The enormous increase in theoretical knowledge and technical possibilities in general surgery has led to differentiation and subspecialization. Gastrointestinal surgery is now recognized as a distinguished area of specific interest within the field of general surgery. It covers the surgical diagnosis and treatment of benign and malignant diseases of the digestive tract, including liver and pancreas. Most gastrointestinal diseases require a multidisciplinary approach in which the gastroenterologist and the gastrointestinal surgeon are key figures. This Journal is planning a series of articles highlighting the recent developments and current state of the art of gastrointestinal surgery, with special emphasis on the close connection with gastroenterology. PMID- 10526610 TI - [Pruritus in cancer: uncommon, but sometimes worse than the pain]. AB - Three patients, two females aged 45 and 56 years with metastasized breast carcinoma and one man aged 88 years with inoperable bronchial carcinoma, suffered from severe pruritus. This was only alleviated after treatment with paroxetine, a serotonin re-uptake inhibitor, or with tropisetron, a serotonin antagonist. The youngest woman then could be given chemotherapy, after which clinical recovery occurred, the other patients died, one week and 3 months, respectively, after start of the treatment. Pruritus is a relatively rare symptom in malignancies, but may be worse than pain. In the development and transmission of pruritus signals, in cholestatic icterus as well, serotonin appears to play a more important part than histamine. PMID- 10526612 TI - [Gastrointestinal surgery and gastroenterology. II. Transplantation of pancreas]. AB - In patients with type I diabetes mellitus, adequate blood glucose control prevents the development or aggravation of late complications. Apart from administration of insulin, transplantation of insulin-producing tissue is also a possibility. Transplantation of Langerhans islets which contain the insulin producing beta cells is still in its initial phase. Transplantation of the entire pancreas received a boost in the mid-eighties when it became possible to drain the secretion of the exocrine part of the pancreas to the bladder using the duodenum. Other important steps forward were prevention and treatment of rejection and improvement of the preservation fluid. Because pancreas transplantation makes lifelong immunosuppression necessary, it is performed mainly in patients subjected to kidney transplantation because of terminal renal failure. The one-year survival of the patients after simultaneous pancreas and kidney transplantation increased to over 90%, that of the grafted pancreas to 82% and that of the grafted kidney to 86-90%. The one-year survival after transplantation of the pancreas alone increased to 62%. A successful pancreas transplantation leads to independence from insulin treatment and to normal glucose and HbA1c values. Pancreas transplantation also reduces diabetes nephropathy and progression of coronary sclerosis. PMID- 10526613 TI - [Gastrointestinal surgery and gastroenterology. III. Diagnosis and treatment of esophageal carcinoma]. AB - Some 900 new cases of oesophageal carcinoma occur annually in the Netherlands. The proportion of adenocarcinomas has increased to 41% in 1989-1992, probably in connection with the increasing incidence of Barrett oesophagus. The diagnosis of oesophageal carcinoma is made on the basis of endoscopical biopsy and histological examination. Staging is done by means of endoscopical ultrasonography for the T and N stages, and CT and external ultrasonography for the M stage. Treatment with curative intent consists of surgical resection, which may be combined with chemotherapy and/or radiotherapy. The 5-year survival rate ranges from 70-90% for a stage I tumour to 5-11% for a stage IV tumour. If a short survival is expected, palliative therapy may improve the passage of food. The palliative therapies used most in the Netherlands are introduction of a self unfolding endoprosthesis or brachytherapy. PMID- 10526614 TI - [Thrombocyte receptors: current views and therapeutic options]. AB - In the action of thrombocytes during stemming of a bleeding after damage to a blood vessel, receptors on the thrombocyte membrane play an important part. Adhesion of platelets takes place via specific binding of receptors; the main binding is that of glycoprotein (Gp) Ib to Von Willebrand factor which is synthetized by endothelial cells. Activation of thrombocytes is stimulated by adhesion and by agonists. Weak agonists, through production of thromboxane A2 and release of agonists from granules cause a self-fortifying process of thrombocyte stimulation; strong agonists (like thrombin) lead also to activation of Gp IIb/IIIa receptors. Aggregation of thrombocytes occurs after activation of Gp IIb/IIIa receptors. During stimulation, a change of shape occurs which enables binding to suitable plasma proteins of which the main one is fibrinogen. Knowledge of thrombocyte receptors enhances the insight into the prognosis and efficacy of certain treatments in diseases in which platelet aggregation is pivotal. Of the six categories of antiplatelet drugs, antagonists of Gp IIb/IIIa receptors are the most potent. In clinical trials good results have been obtained in patients with coronary disease of the intravenously administered form added to acetylsalicylic acid. PMID- 10526615 TI - [Effects of articles published in the Dutch Journal of Medicine]. AB - OBJECTIVE: To inventory the effects of publishing in the NTvG. DESIGN: Retrospective, descriptive. METHODS: The first authors of the articles of the sections Clinical lessons, Capita selecta, For practice, Original articles, Case reports and Side effects of drugs, from the issues 27-53 of Volume 138 (1994) of the NTvG were approached for a written enquiry about the effects of their articles. Reactions in the form of letters to the editor were assessed by screening the relevant section. RESULTS: The results of the enquiry concerned 165 articles. The authors of 160 articles (97%) reported that they had been approached in person with reference to the publication, on average 15.7 times orally and 2.0 times in writing. The authors of 66 articles (40%) were invited to deliver a lecture, data from 62 articles (38%) were used by others and 54 articles (33%) resulted in (continuation) research. An approach by the media followed after 23 articles (14%). A positive effect on the number of relevant patient referrals was mentioned of 4 (44%) articles from section For practice and 7 (30%) articles from section Clinical lessons. Various other effects were reported of 48 articles (29%). Forty (20%) of the total of 197 articles involved in the study were followed by one or two letters to the editor, prompted mostly by Clinical lessons (33%). CONCLUSION: Virtually all articles produced an effect. Many articles were followed by personal reactions and many articles elicited a response indicating influence on medical teaching, science or clinical activity. PMID- 10526616 TI - [Influence of specific clinical lessons in the Dutch Journal of Medicine on clinical management]. AB - OBJECTIVE: To establish whether publication of an article propagating a laboratory test leads to measurable increase of the number of requests for that test. DESIGN: Retrospective. METHOD: From volume 138 (1994) of the Dutch Journal of Medicine (NTvG), three clinical lessons were selected that contained an unequivocal clinical message and a recommendation to request a specific laboratory test for particular patients. All laboratories performing the test in question were asked to report the number of requests per month in the months before, after and during publication of the article in question and during the same months of 1993. The difference between the number of requests in the period after publication of the article in 1994 and in the same period in 1993 was determined and tested postulating a Poisson distribution. RESULTS: Regarding two clinical lessons (one about determination of Coxsackie virus in neonates and one about examining arthritis patients for parvovirus B19) no significant difference in the numbers of requests before and after the publication was found, particularly also because the laboratories could not supply itemized data so that relevant information was lost in a flood of other data. The third clinical lesson (about determination of antibodies against Onchocerca in patients complaining of itching after a trip to the tropics) was followed by a significant increase of the number of requests (from 50 to 90; p < 0.001) in the 3 months following publication. CONCLUSION: Publication of a clinical lesson about a recommended laboratory test for onchocerciasis in the NTvG resulted in a significant rise of the number of requests for that test. PMID- 10526617 TI - [Implantable ECG recorder revealed the diagnosis in a baby with apparent life threatening events]. AB - A 14-month-old boy went through episodes of cyanosis and brief loss of consciousness. Extensive investigations failed to lead to a diagnosis, until an implanted ECG recorder revealed ECG abnormalities suggestive of strangulation. Interviews with the father and mother showed that this was indeed the case. The diagnosis of 'Munchhausen by proxy' was made. Psychiatric assistance and home help were called in. The child recovered well. If there is a suspicion of arrhythmia as the cause of apparent life-threatening events, prolonged ECG recordings are necessary. In a clinical environment it is possible to make continuous ECG recordings during a limited period. An insertable recorder allows continuous ECG recordings during a syncopal event and can be used for prolonged monitoring. The patient presented is the youngest infant in the world in whom such a device has been implanted. PMID- 10526618 TI - [Reports in the Dutch newspapers prompted by articles from medical scientific journals]. AB - OBJECTIVE: To gain an impression of the frequency and nature of reports in Dutch newspapers about publications in medical-scientific journals. DESIGN: Retrospective, descriptive. METHODS: Press reports appearing during February to July 1995 in the internal and external collections of newspaper cuttings of the Ministry of Public Health, Welfare and Sports, both composed of reports from, among other things, nine popular evening and morning newspapers, were examined for articles that had been prompted directly by medical publications; this study was carried out in the editorial offices of the Dutch Journal of Medicine (NTvG). The title of the publication and the report, the name of the newspaper and the name of the journal were recorded. RESULTS: In all, 34 reports prompted by 27 different articles were found, most of them (21%) written in reference to publications in The New England Journal of Medicine, followed by the NTvG (15%). The list was headed by socially relevant subjects such as cancer and aids (29% and 12%, respectively, of the reports). The Telegraaf and the Volkskrant contained the largest numbers of reports (both 18%) and nearly all newspapers used journals published in either English or Dutch. CONCLUSION: Dutch newspapers regularly use articles from various medical-scientific journals published in English and in Dutch as a direct source for reporting. Most attention was given to socially relevant subjects such as cancer and aids. PMID- 10526619 TI - [Headache and chronic sleep deprivation: an often missed relationship in children and also in adults]. AB - Three children, two girls aged 9 and 13 years and a boy aged 9 years, and a man aged 22, presented with headache of migrainoid character. The complaints resolved after attention had been given to sleeping habits, notably problems with falling asleep. The literature on the relationship between lack of sleep and headache is scarce. In history-taking doctors often neglect sleeping customs. Yet awareness of this connection can provide the clue to a successful treatment of headache. In more than half the children with headache who are referred to the outpatient clinic of Neurology, a serious lack of sleep is a major contributing factor to the problems. One should strongly advise against use of coffee and Cola drinks at all times and of tea in the evening. Patients are urged to create a restful atmosphere about bedtime: taking a walk or a hot shower, being told a story and so on. They should not read in bed, watch TV or play computer games in the bedroom. Prescription of clonidine 0.025-0.075 mg in children before bedtime may be useful. PMID- 10526620 TI - [Breastfeeding and contraception]. AB - In the Netherlands many women stop breastfeeding in the first few months postpartum. In 1997, only 16.9% of all 3-month-old babies received full breastfeeding. One of the causes may be insufficient support by the medical profession. A second factor is that often combined oral contraceptives are prescribed to breastfeeding women. As it has been shown that estrogens in these contraceptives inhibit lactation, this is probably one of the reasons why breastfeeding frequently fails in this country. WHO advises not to use estrogens during lactation. According to recent research the lactational amenorrhoea method of contraception (LAM) is highly effective during the first 4 months postpartum. In the 5th and 6th month the effectiveness is strongly dependent on the accuracy by which the conditions are met. The medical profession should pay more attention to the support of breastfeeding and contraception in relation to each other. PMID- 10526621 TI - [Glutamine as a key ingredient in protein metabolism]. AB - Glutamine has a number of unique properties which suggest that this amino acid plays an important role in health and disease. Glutamine is considered a conditionally essential amino acid, because during periods of severe metabolic stress the body's requirements of glutamine may exceed the individual's ability to produce sufficient amounts of the amino acid. Studies with glutamine-enriched nutrition show beneficial effects on nitrogen balance, muscle protein metabolism, gastrointestinal mucosa, and immune status. In certain patient categories addition of glutamine reduces the number of infectious complications, improves long-term survival, and shortens hospital stay, e.g. bone marrow transplantation patients, neonates with severely subnormal weight, patients with multiple organ failure, multi-trauma patients. More studies are needed to document the dose response and to identify the patients that are likely to benefit from glutamine supplementation. PMID- 10526622 TI - [Condylomata acuminata: a rare symptom of ubiquitous human papilloma virus and not a sign of risky sex behavior]. AB - In general, condylomata acuminata can be diagnosed and treated by the general practitioner. Condylomata are caused by certain types of human papillomavirus (HPV). According to their carcinogenicity HPVs are classified as high risk and low risk HPV. The benign condylomata are an infrequent sign of an infection with low risk HPV, while cervical cancer is a rare and late complication of an infection with high risk HPV. Because high and low risk HPV are different viruses, the risk of cervical cancer is not increased by condylomata. Anogenital HPVs are predominantly transmitted sexually. It is useful to discriminate between sexually transmitted diseases (STDs) that are ubiquitous, like infections with HPV or herpes simplex virus (HSV), and rare STDs like syphilis, gonorrhoea and HIV infection: infections with HPV and HSV are also common with unriskful sexual behaviour, while syphilis, gonorrhoea and HIV infection are almost exclusively associated with riskful sexual behaviour. It has been shown that double infections with HPV and Chlamydia trachomatis are not more frequent than may be expected by chance. The literature indicates that the presence of condylomata acuminata by itself is no reason to screen patients for other STDs. PMID- 10526623 TI - [National study of the spread of hepatitis B to provide a foundation for future vaccination policy]. AB - On May 1st, 1999, a study was started regarding the spread of hepatitis B in the Netherlands (the so-called BRON study). For one year, data will be collected on new cases of hepatitis B with respect to transmission route, source of infection and risk factors. In addition, by use of mailed questionnaires, information on risk factors will be collected within a large group of control subjects throughout the country. The BRON study is being carried out by the National Institute for Public Health and the Environment (RIVM) in co-operation with the municipal health services and medical laboratories. The results of this study should underpin future governmental policy decisions regarding universal or selective vaccination against hepatitis B. PMID- 10526624 TI - [Can intraocular lenses containing methylmethacrylate polymers cause hypersensitivity reactions?]. PMID- 10526626 TI - [Thiomersal in gammaglobulins for pregnant travelers may not be safe for the fetus]. PMID- 10526625 TI - [Irregular blood group antagonisms]. PMID- 10526627 TI - Comparison of different tracers in the follow up of differentiated thyroid carcinoma. AB - In the follow up of differentiated thyroid carcinoma (DTC) several scintigraphic methods are used in addition to the serum thyroglobulin and ultrasonography of the neck. Iodine-131 whole body scintigraphy (WBS), which is performed since many years, is able to detect iodine positive recurrence, lymph node metastases and distant metastases in a very specific way. However, the problem of I-131 WBS is the fact that only 67% of metastases from DTC accumulate iodine. Therefore non specific radionuclides like TI-201 or tracers such as Tc-99m Sestamibi or Tc-99m Tetrofosmin and new metabolic tracers like F-18 FDG were introduced in the diagnostic work up to detect iodine negative metastases as well. This study describes the comparison of different tracers in 35 patients with elevated thyroglobulin and suspicion of metastatic disease or already known metastases from DTC. PMID- 10526628 TI - Thyrotropin-releasing hormone (TRH), a signal peptide of the central nervous system. AB - Thyrotropin-Releasing Hormone (TRH; pyroGlu-His-Pro-NH2), originally isolated as a hypothalamic neuropeptide hormone, most likely acts also as a neuromodulator and/or neurotransmitter in the central nervous system (CNS). This interpretation is supported by the identification of a peptidase localized on the surface of neuronal cells which has been termed TRH-degrading ectoenzyme (TRH-DE) since it selectively inactivates TRH. Vice versa it also holds true that TRH is selectively inactivated only by TRH-DE and thus, this enzyme might be considered to be the terminator of TRH signals. In situ-hybridization histochemistry was used to study the TRHergic communication system by analyzing the gene expression of TRH-DE in relation to TRH and to the TRH receptors (TRH-R1 and TRH-R2). TRH mRNA is highly expressed in "thyrotropic" hypothalamic regions and in some selected brain areas. For TRH-R1 and TRH-R2, an almost exclusive mRNA distribution pattern was noticed in many brain regions. Interestingly, a widespread distribution of TRH-DE predominantly in neo- and allocortical regions was observed essentially overlapping the distribution patterns of TRH-R1 and TRH R2. These data support the hypothesis that TRH-DE is important in the TRH mediated modulation of sensory, locomotor and cognitive functions of the CNS and could be considered to be a marker to map TRHergic pathways. PMID- 10526629 TI - [Hypophyseal-hypothalamo-thyroid axis in affective disorders]. AB - For centuries there has been convincing evidence that diseases of the thyroid gland may produce psychiatric symptoms. Nowadays systematic data are available concerning a higher incidence of thyroid diseases in psychiatric patients and vice versa a higher incidence of psychiatric disorders in thyroid patients. A more subtle approach concerns challenge tests like the TRH test. It could be shown that depressive patients show a blunted TSH response to TRH in 30-40% of the cases. This might lead to a definition of a subgroup of depressives from a psychoneuroendocrinological point of view. But it also might have an impact on the prediction of treatment outcome in psychopharmacological treatment approaches. PMID- 10526630 TI - [Thyroid autoantibodies in depressive disorders]. AB - Whereas clinical relevant hypo- as well as hyperthyreosis are strongly suspected to induce psychiatric symptoms, there is a controversy about the relevance of only subclinical and autoimmune findings. We found autoantibodies (MAK, TAK, TRAK) in a high percentage (100 out of 144 = 70%) in severely depressed inpatients. Also we found a Hashimoto thyreoiditis in 5 patients. In the long run this may lead to relevant hypothyreosis which is regarded to be a risk factor for depression and for possible non-response in medical treatment. We conclude that in cases of repeated depressive episodes especially depression of the elderly and in nonresponders it seems necessary not only to get lab for TSH, T3 and T4 but also to assess the autoimmune status of the thyroid gland (autoantibodies). There is further need for controlled studies whether there is a better outcome in nonresponders to antidepressive medical treatment and positive autoantibody status after supplementation with triiodothyronine. PMID- 10526631 TI - [Administration of thyroid hormones in therapy of psychiatric illnesses]. AB - This review tries to give the state of the art of the therapeutic use of thyroid hormones in psychiatric disorders, mainly in depression. Four hypotheses suggest an effect in these ailments: 1) a local relative T4 excess present (or better postulated to be present) in the brain of depressed patients is lowered by triiodothyronine (T3), by lowering serum levels of thyroxine (T4), 2) the effect of a depression-mediated cerebral lack of catecholamines is compensated by the T3/T4 induced activation of beta-receptors, 3) a postulated depression-induced local cerebral hypothyroidism can be counteracted by T3 and T4. 4) thyroid hormones increase the cerebral content of serotonin. This may be beneficial in depression, where shortage of serotonin in the brain is accused to be etiologically important. Thyroid hormones have been used so far in the following ways: 1) as T3 monotherapy in depression; 2) initial additive T3 for acceleration of the response to treatment with tricyclic antidepressants (TCA); 3) additive T3 for augmentation of the response to TCA in therapy-resistant patients with depression, 4) as high-dose (250-500 micrograms/die) T4-treatment of "rapid cycling bipolar disorder". Low dose (5-50 mg/die) T3 "augmentation therapy" is the best documented form of treatment with thyroid hormones in depression. The results suggest a convincing benefit for a varying percentage of non responders to therapy with TCA. For the other forms of treatment placebo-controlled double blind studies are not yet available or give conflicting results. PMID- 10526632 TI - [Thyroid gland and sleep]. AB - A set of data suggests that the thyroid gland plays a role in the bi-directional interaction between the electrophysiological and the endocrine components of sleep, e.g. the nonREM-REM-cycle and the patterns of nocturnal hormone secretion, respectively. In detail thyroid-stimulating hormone (TSH) and thyroxin (T4) show circadian rhythms. A specific relationship was observed between TSH and REM sleep. Blunted TSH levels were found in healthy elderly subjects and, probably due to overactivity of corticotropin-releasing hormone in patients with depression in comparison to young normal controls. Pulsatile administration of thyrotropin-releasing hormone induced a decrease of sleep efficiency and an earlier occurrence of the cortisol rise in normal controls. Slow wave sleep was reduced in patients with hypothyroidism in comparison to normal controls. The sleep EEG normalised after therapy. PMID- 10526633 TI - 'Paracrine' control of spermatogenesis. AB - Spermatogenesis is a remarkably complex but precise process yielding highly differentiated haploid germ cells from diploid stem cells. Although many factors have been implicated in the paracrine control of spermatogenesis, functional proof is only available for a few regulators. Among those are androgens, growth factors and stem cell factor. Cell- and organ-specific genetargeting will provide important insights into the relevance of local factors controlling male gametogenesis. As testicular communication frequently occurs between rather remote cells and compartments, it is proposed that the term 'local' rather than 'paracrine' mediators/factors should be used, since the latter term refers to communication amongst neighbouring cells (and mainly via diffusion). PMID- 10526634 TI - Sperm dysfunction in subfertile patients with varicocele and marginal semen analysis. AB - A comparative analysis of potentially functional spermatozoa per ejaculate, progressive motility, hypo-osmotic swelling test, acrosome integrity and sperm viability (24 and 48 h) was carried out in a group of 40 subfertile patients with varicocele and marginal semen analysis and 40 fertile subjects, in order to identify subclinical abnormalities that may explain subfertility. Patients with varicocele had lower numbers of potentially functional spermatozoa per ejaculate, progressive motility, acrosome and membrane integrity and sperm viability. These abnormalities were not related to the grade of varicocele, testicular volume or serum FSH concentration. A positive correlation between the hypo-osmotic swelling test and progressive motility (r = 0.71) and between potentially functional spermatozoa and the hypo-osmotic swelling test (r = 0.69) was found in patients with varicocele. These data suggest that some of the deleterious effects produced by the varicocele might be related to sperm migration and viability in the female genital tract and others to sperm-zona interaction and/or sperm-egg fusion. PMID- 10526635 TI - Flow cytometric analysis of semen preparation, and assessment of acrosome reaction, reactive oxygen species production and leucocyte contamination in subfertile men. AB - Reactive oxygen species (ROS) production, poor acrosome reaction ability, and leucocyte and round cell contamination of semen are associated with diminished male fertility. In addition to the assessment of these variables, the number of abnormal and round cells in semen preparation was studied by flow cytometry. Men with varying degree of fertility despite normal concentrations and motility of spermatozoa were investigated. Group 1 consisted of semen donor candidates (n = 25), group 2 of patients with acceptable fertilization in vitro, but diminished fertility in vivo (n = 56), and group 3 of patients with poor (< 25%) fertilization in vitro (n = 32). The subfertile men in groups 2 and 3 had lower concentrations and poorer rapid progressive motility of spermatozoa than the men in group 1. Using flow cytometry, the number of events in the round cell gate in the Percoll-prepared semen sample was higher in the subfertile men than in the donors, independently of the differences in other semen parameters. However, there were no significant differences between the groups in the number of round cells in native semen by microscopic analysis, or in ROS production or acrosome reaction. In conclusion, flow cytometric assessment of round cells may be a useful method in the investigation of male fertility. PMID- 10526636 TI - Use of acridine orange to evaluate chromatin integrity of human spermatozoa in different groups of infertile men. AB - To study the sperm chromatin compactness various methods, such as acidic aniline blue or acridine orange staining, have been applied. Due to its metachromatic properties, acridine orange dye fluoresces green with double- and red with single stranded DNA. Samples (n = 181) were evaluated and grouped as follows: group I, normal recently fertile; group II, male having female partner with repeated early pregnancy loss; group III, male with varicocele; and group IV in-vitro fertilization and intrauterine insemination failures. Routine semen analyses were carried out in all the cases. Amorphous particulate matter as observed under phase contrast microscope was graded on the scale of nil to +4. Fixed smears were stained with an aqueous solution of acridine orange and viewed under a fluorescence microscope. Two hundred cells were counted and the percentage of fluorescence calculated. Groups II, III and IV exhibited significantly low green fluorescence compared with the control group. The study also indicates that increased amorphous particulate matter (indicating infection) might be one of the contributing factors to lower acridine orange stainability. Thus acridine orange staining can be used to evaluate the integrity of the nucleus, disorders of which can cause unexplained infertility or lower fertilization potential that may go undetected by routine analysis. PMID- 10526637 TI - Expression of IL-12, IL-10, PGE2, sIL-2R and sIL-6R in seminal plasma of fertile and infertile men. AB - The involvement of cytokines and other immunoregulatory factors in male infertility is still unclear. In the present study we compared the levels of IL 12, IL-10, PGE2, sIL-2R and sIL-6R in the seminal plasma (SP) of fertile and infertile men. Four groups were included: fertile donors (FERT), infertile men with azoospermia (AZOO), and infertile men with either oligoterato asthenoazoospermia (OTA), or OTA with genital infection (OTA-INF). Cytokines and cytokine-soluble receptors in semen were evaluated by specific ELISA commercial kits. The levels of IL-12, sIL-2R and sIL-6R were similar in SP of fertile and infertile men. The mean levels of IL-10 in the SP of FERT, OTA and AZOO were 5.6 +/- 0.9, 4 +/- 2.8 and 8 +/- 3.5 pg ml-1, respectively, and did not differ significantly. The mean level of IL-10 in SP from OTA-INF (0.9 +/- 0.5 pg ml-1) was significantly lower than that for FERT (5.6 +/- 1.9 pg ml-1; P = 0.02) and AZOO (8.2 +/- 3.4 pg ml-1; P = 0.05), but not significantly different from that for OTA (3.7 +/- 2.1 pg ml-1). The mean SP level of PGE2 was significantly lower in SP of OTA-INF than FERT (7.67 +/- 2.26 and 19.67 +/- 3.69 micrograms ml-1, respectively; P < 0.02). In conclusion, the seminal plasma from fertile and infertile men contained similar levels of IL-12, sIL-2R and sIL-6R. However, the levels of IL-10 were significantly lower in SP from OTA-INF compared to FERT and AZOO. Our results indicate that specific cytokines and their soluble receptors are involved in the male reproductive system. PMID- 10526638 TI - Why do we determine alpha-glucosidase activity in human semen during infertility work-up? AB - alpha-Glucosidase is a normal constituent of human semen, produced mainly in the epididymis. It is significantly correlated to sperm count. Its activity is low in cases of epididymal obstruction. We evaluated alpha-glucosidase activity in 653 semen samples of patients, who attended our department for marital infertility, with respect to associations to clinical and other seminal parameters. The normal range (mean +/- 2 SD) in samples with normal parameter values was 7.2-46.4 mU ml 1. The determination in patients with azoospermia revealed mean values of 7.7 +/- 9.5 mU ml-1 in obstructive azoospermia, and 15.8 +/- 11.5 mU ml-1 in nonobstructive azoospermia. The difference was not statistically significant in that the sensitivity of determination with respect to the presence of obstruction was only 0.66, and the specificity 0.83. A significant correlation (r = 0.34) of alpha-glucosidase activity with log sperm count was observed. The mean alpha glucosidase activity was not significantly different in groups formed according to sperm motility, according to leucocyte count or according to semen volume. A difference between smokers and nonsmokers with comparable sperm count, as reported in the literature, did not occur. We conclude from our results that the determination of alpha-glucosidase activity does not give additional information of the fertility status exceeding that of other clinical investigations or parameters of semen analysis. PMID- 10526639 TI - Which efforts towards conservative treatment of male infertility will be successful? Reactive oxygen species, antioxidants, and sperm phospholipids. PMID- 10526640 TI - Which efforts towards conservative treatment of male infertility will be successful? Antibiotic therapy. PMID- 10526641 TI - Which efforts towards conservative treatment of male infertility will be successful? Mast cell blockers. PMID- 10526642 TI - Which efforts towards conservative treatment of male infertility will be successful? Antiphlogistics and glucocorticoids. PMID- 10526643 TI - The testicular antiviral defence system: I. Interferon and interferon-induced proteins expression in the testis. PMID- 10526645 TI - Testicular macrophages and inflammation. PMID- 10526644 TI - Antisperm antibody detection: methods and standard protocol. PMID- 10526646 TI - Immunological aspects of male accessory gland infections. PMID- 10526647 TI - The sperm acrosomal antigen 1 (SAA-1) in male infertility. PMID- 10526648 TI - Results of ICSI in the treatment of male immunological infertility. PMID- 10526649 TI - Immunity to spermatozoa and male fertility. AB - Immunological characterization of semen and male fertility are discussed. The pH of seminal plasma is very similar in groups of fertile and infertile men. Nonspecific immunological factors such as lysozyme, C3c, alpha-1 antitrypsin and beta-2 microglobulin were significantly higher in 14% of infertile men. Seminal antibodies agglutinating spermatozoa are frequently observed in IgG and IgA isotypes. High levels of seminal anti-phospholipid antibodies in seminal plasma could influence epitopes on the endometrium, oocyte or early embryo in patients with repeated unexplained spontaneous miscarriages. PMID- 10526651 TI - Immunology of the epididymis. PMID- 10526650 TI - The zona pellucida 'receptors' ZP1, ZP2 and ZP3. AB - The male component that is necessary for successful reproduction depends on a large variety of biological processes working in concert. The sperm-egg interaction occurs through complementary molecules and is an obligatory process for successful fertilization. However, this complex phenomenon and its molecular mechanisms remain to be fully understood. The oocyte is protected by the zona pellucida, a network of various proteins which encloses the oocyte. Depending on the species, the zona pellucida consists of different glycoproteins that are proposed to function as 'receptors' for spermatozoa. In the mouse, ZP1 is the homodimeric filament crosslinker, held together by intermolecular disulphides. ZP2 is the 'secondary receptor', which is cleaved by egg proteases after egg activation. The mouse ZP3 protein appears to be the 'primary receptor', which is responsible for species-specific binding of spermatozoa to the oocyte and the induction of the acrosome reaction. To localize zona pellucida protein and to evaluate the function of ZP2 and ZP3, polyclonal antisera were raised against synthetic ZP2 or ZP3 peptides which are specific for human or for mouse zona pellucida proteins. It could be demonstrated that anti-synthetic peptide antisera detected their respective zona pellucida proteins in immunoblots, ovary sections and native hemizonae pellucidae. Functional assays with anti-ZP3 synthetic peptide antibodies revealed that the antisera did not inhibit sperm-zona pellucida binding, whereas one of the antisera against synthetic ZP2 peptides significantly inhibited binding of spermatozoa to the zona pellucida. PMID- 10526652 TI - Distribution and function of angiotensin II receptors in mouse spermatozoa. AB - Previous work indicates that angiotensin II (AngII) stimulates sperm motility and acrosomal exocytosis. Here we determined the distribution of AngII receptors on mouse sperm by immunocytochemistry and used Ca2+ probe photometry to examine their coupling to sperm regulatory pathways. We found both AT1 and AT2 receptors localized on the acrosomal region of the sperm head. The AT1 receptor, but not the AT2 receptor, is found also on the principal piece of the sperm tail. Local perfusion of motile but nonprogressive sperm with 0.1-1 microM of AngII evokes a rapid, substantial rise in intracellular [Ca2+]. This response is blocked by losartan, a specific antagonist of the AT1 receptor. These results indicate that sperm possess functional AT1 receptors that are distributed to sites that may allow selective control of motility and exocytosis. They also suggest that the AT2 receptors detected by immunoreactivity are either nonfunctional or are not coupled to Ca(2+)-mediated signalling mechanisms. PMID- 10526653 TI - Flow cytometric studies on sperm antibodies. PMID- 10526654 TI - The preparation of sperm antigens. PMID- 10526655 TI - Parental consanguinity as a cause for increased incidence of births defects in a study of 238,942 consecutive births. AB - The risk for birth defects in the offspring of first-cousin matings has been estimated to increase sharply compared to non consanguineous marriages. As a general decline in the frequency of consanguineous marriages was observed in this century, one wonders whether consanguinity is still a factor in the appearance of birth defects in developed countries. Based on our registry of congenital anomalies we tried to answer to this question. In the population studied in North Eastern France a consanguineous mating was known in 1.21% of the cases with congenital anomalies, vs. 0.27% in controls, (p < 0.001). The frequency of the malformations recorded paralleled the degree of consanguinity: out of 89 malformed children, 51 were seen in first-cousins mating (10.3 times more frequent than in offspring of non consanguineous couples), 17 in second-cousins marriages and 18 in more distant relatives mating. Three were uncle-niece marriage. Excluding known mendelian conditions these numbers were 73, 36, 17 and 17 respectively and the corresponding relative risk were 3.68, 3.01, 3.41 and 4.89 respectively. Therefore there is a negative dose-response effect between level of inbreeding and risk of congenital malformations. Consanguineous mothers were more often pregnant than non consanguineous mothers (p < 0.01) and they had more stillbirths than non consanguineous mothers. These results show that consanguinity is still a factor of birth defects and they must be taken into account for genetic counseling of inbred marriages, in developed countries. PMID- 10526656 TI - Familial high myopia: evidence of an autosomal dominant mode of inheritance and genetic heterogeneity. AB - High myopia, defined as a refractive error inferior to -6 diopters, often appears as a familial disease. In order to precise its genetic background, we performed a segregation analysis on 32 French families (320 subjects including 120 individuals with clinical data) containing at least one high myopic person in their genealogy. Under the assumption of a two-alleles single gene model, the autosomal dominant transmission mode showed a much greater likelihood than the autosomal recessive mode, which therefore was rejected. From the segregation model obtained, a two-point linkage analysis was made on 18 families (107 subjects), among the 32 used for the segregation analysis. Different candidate loci were tested: collagen genes including Stickler syndrome types 1 and 2, proteoglycan genes, Marfan 1 syndrome and a Marfan like disorder localised in 3p24.2-p25. No evidence of linkage was found with any of the studied markers. In addition, the absence of linkage with chromosome 18p11.31 markers, a locus linked to familial high myopia in 6 North American families and 1 family of Chinese descent, demonstrated the genetic heterogeneity of the disease. PMID- 10526657 TI - Genetic comparisons of patients with cystic fibrosis with or without meconium ileus. Clinical Centers of the French CF Registry. AB - Cystic fibrosis (CF) is an autosomal disorder caused by mutations in the cystic fibrosis transmembrane conductance regulator gene (CFTR). Neonatal meconium ileus (MI) occurs in 10-20 percent of newborns with CF. The purpose of this study was to determine the allelic frequencies of the CF mutation in French patients with and without MI and the incidence of MI in 7 homozygotes or compound heterozygotes for mutation of the CFTR gene. Our study confirms the positive association between delta F508, the most frequent CF mutation, G542X mutation and MI and a negative association with G551D. PMID- 10526659 TI - Partial trisomy 4q and monosomy 9p resulting from a familial translocation t(4;9)(q27;p24) in a child with choanal atresia. AB - A male infant with a deletion of 9p and concomitant duplication of 4q: 46,XY, der(9)t(4;9)(q27;p24), is described. Parental chromosome analysis showed a balanced maternal translocation. To our knowledge, the above cytogenetic and clinical abnormalities have not been described previously. A phenotype comparison is presented with previously reported cases concerning a deletion of 9p and a duplication of 4q. PMID- 10526658 TI - CTG instability in myotonic dystrophy: molecular genetic analysis of families from south-eastern France with characteristics of intergenerational variation in CGT repeat numbers. AB - We report clinical, genetical and genealogical findings in 149 French families from the Rhone-Alpes area studied over a 5-year period. There was a significant excess of DM females compared to DM males with (CTG) repeat sizes between 1-2 kb. The mean maternal (CTG) repeat size was higher than paternal repeat size. Anticipation phenomenom was significantly higher after maternal than after paternal transmission. A significant correlation between parental (CTG) repeat size and intergenerational variation both in paternal and maternal transmissions was observed. The anticipation phenomenom was more important for sons than daughters particularly after maternal transmission. The mean (CTG) repeat size in mothers of CDM cases was about twice that of mothers of NCDM children. The risk of giving birth to a CDM child increased considerably when the number of maternal (CTG) repeats was over 300 (CTG). A significant excess of DM females was observed. They had on average 24% fewer children than male patients. Paternal transmission (63.6%) of DM occurred more frequently than maternal transmission (52.7%). PMID- 10526660 TI - Pure partial trisomy 5q33-->5q35 resulting from the adjacent-1 segregation of a paternal (5;14)(q33;p12) translocation. AB - A 14-year-old male was referred for evaluation of mental retardation with short stature and dysmorphic features. His karyotype was 46,XY,der(14)t(5;14)(q33;p12)pat, resulting in a pure partial 5q33-q35 trisomy due to the adjacent-1 segregation of a paternal balanced translocation. Paternal blood karyotype revealed a balanced translocation t(5;14)(q33;p12) retaining Ag Nors. To date, only two cases of pure partial 5q trisomies spanning this region have been reported. Analysis of these cases and the one we report does not allow the delineation of a specific phenotype. PMID- 10526661 TI - Kyphomelic dysplasia: a report of a family with an autosomal dominant pattern. AB - Kyphomelic dysplasia (KD) is a rare autosomal recessive entity characterized by shortening and bowing of the limbs, skin dimples, abnormalities of methaphysis and ribs, a short trunk, a narrow thorax, neonatal respiratory distress, platyspondyly, and facial dysceptism with micrognathia, midfacial hypoplasia, and a broad nasal bridge. Some children die in early infancy. The survivors show normal hands, feet, cranium and psychomotor development. The condition varies in severity. The facial features and bowing improve during childhood, and stature remains short during adulthood. We report here a family with KD inherited as an autosomal dominant trait, which appears to be less severe than the autosomal recessive form, without facial and vertebral a favorable outcome and with involvement and final short stature. PMID- 10526662 TI - Familial coarctation of the aorta in three generations. AB - Four members in three generations of a family were affected by a coarctation of the aorta (CoA) which was mild or severe, either isolated or in association with other cardiac defects. This family suggests that a rare form of CoA could be the result of an autosomal dominant mutation with incomplete penetrance and variable expressivity rather than polygenic inheritance. PMID- 10526663 TI - Monitoring responses to antiangiogenic agents using noninvasive imaging tests. PMID- 10526664 TI - Cancer predisposition associated with defective DNA repair: studies with mutant mouse strains. PMID- 10526665 TI - Greater curability in advanced Hodgkin's disease? PMID- 10526666 TI - Complex decision-making for BRCA1/2 carriers. PMID- 10526667 TI - A survey of treatments used in patients with metastatic melanoma: analysis of 189 patients referred to the National Cancer Institute. AB - BACKGROUND: Few effective treatments exist for patients with metastatic melanoma. The United States Food and Drug Administration has approved the use of interferon alfa-2b after the resection of locoregional disease, and dacarbazine or interleukin-2 for the treatment of patients with metastatic melanoma beyond the locoregional area, although many additional agents and combinations of agents are currently in use. METHODS: Between January 1997 and June 1998, the Surgery Branch of the National Cancer Institute conducted a prospective analysis of 226 consecutive patients with metastatic melanoma referred for protocol evaluation. The previous systemic treatments these patients received both before and after the development of metastatic disease were tabulated, along with the association of these treatments with formal institutional protocols. Only the identity of the agents and not the dose or the schedule of treatments was considered in this analysis. Complete information could be obtained from 189 of the 226 patients. RESULTS: Of the 189 patients evaluated for this study, 135 (71%) received some form of systemic therapy before referral to the National Cancer Institute. Before the development of metastatic disease, 80 patients were administered 25 different systemic treatments, including 23 different agents. After the development of metastatic disease, 53 patients were administered 57 different systemic and regional treatments, including 37 different agents. After the resection of all metastatic sites, 23 patients were administered nine different systemic adjuvant treatments. Overall, 78 different systemic treatments were administered to these patients. The majority of treatments in each group were not associated with formal institutional protocols. CONCLUSIONS: This study has demonstrated that a large number of agents and different combinations of agents are currently being administered to patients before and after the development of metastatic melanoma, and frequently not within the context of an approved institutional protocol. These results indicate a need for more formal evaluation of treatments in prospective protocols and greater standardization of the treatment of patients with melanoma. PMID- 10526668 TI - Long-term results of an intensive regimen: VEBEP plus involved-field radiotherapy in advanced Hodgkin's disease. AB - PURPOSE: This pilot study was conducted to evaluate the efficacy and toxicity of a new intensive drug regimen, combined with involved-nodal-field radiotherapy, in advanced Hodgkin's disease not treated by chemotherapy. PATIENTS AND METHODS: From September 1990 to March 1993, 73 evaluable patients with newly diagnosed stage IIB, III (A and B), and IV (A and B) Hodgkin's disease or who were relapsing after primary subtotal or total nodal irradiation were treated with eight cycles of etoposide, epirubicin, bleomycin, cyclophosphamide, and prednisolone (VEBEP) followed by radiotherapy (30-36 Gy) to the nodal site or sites of pretreatment disease. The median duration of follow-up was 68 months. RESULTS: The complete remission rate was 94% (95% CI: 86-98). At 6 years, freedom from progression and overall survival rates were 78% (95% CI: 68-88) and 82% (95% CI: 73-91), respectively. There was one episode of fatal sepsis after bone marrow aplasia that occurred after VEBEP and extended-field irradiation. Hematologic toxicity during chemotherapy was acceptable; without the support of growth factors, grade IV leukopenia and grade IV neutropenia, as determined within cycles, occurred in 38% and 85% of patients, respectively, but was reversible in the vast majority of patients by the day of treatment recycle. No episodes of epidoxorubicin-related cardiomyopathy or symptomatic pulmonary toxicity were documented. Overt and/or subclinical hypothyroidism occurred in 38% of cases. Gonadal damage was evident in the large majority of male patients but reversible in half of them, whereas permanent sterility was observed in females at least 35 years of age. No secondary leukemia has been so far detected. DISCUSSION: VEBEP followed by involved-nodal-field radiotherapy is an effective treatment for chemotherapy-naive Hodgkin's disease and is associated to acceptable rates of acute and intermediate-term toxicity. This intensive regimen, which does not routinely require the support of hematopoietic growth factors and can be delivered in an outpatient setting, warrants a prospective comparison in a randomized trial versus one of the more effective standard-combination regimens. PMID- 10526669 TI - The quality of life associated with prophylactic treatments for women with BRCA1/2 mutations. AB - PURPOSE: This study was conducted to obtain and compare the preferences assigned to cancer states and prevention measures by women who had breast cancer, were at high risk for breast cancer, or had neither condition. PATIENTS AND METHODS: We administered a time trade-off questionnaire to 21 breast cancer patients, 28 women with a personal history of multiple breast biopsies or a family history of breast cancer, and 135 women without these conditions (the reference group). We stratified the reference group into two groups aged 20 to 32 years and 33 to 50 years, respectively. RESULTS: All four groups assigned higher preference to breast cancer than to ovarian cancer. Both reference groups preferred using a tamoxifen-like drug to having mastectomy or oophorectomy for cancer prevention; the high-risk and breast cancer groups did not. None of the four groups had a preference between prophylactic mastectomy and breast cancer. All the groups were willing to subtract more years from their life expectancy to protect offspring from genetic risk than to protect themselves. Reference group members in the 33- to 50-year age range had lower mean ratings than the breast cancer group for almost all the health states, and breast cancer patients were less willing than other respondents to trade time for health. Most of these differences were not statistically significant. The high-risk group was similar to the older reference group in time trade-off ratings. DISCUSSION: The time trade-off-based preferences of healthy women may be used to predict the treatment preferences of women with BRCA1/2 mutations. Obtaining healthy women's ratings of treatment outcomes may help health care policy makers envision the consequences of the difficult choices that high-risk women face. PMID- 10526670 TI - Treatment of Ras-induced cancers by the F-actin cappers tensin and chaetoglobosin K, in combination with the caspase-1 inhibitor N1445. AB - For transforming normal fibroblasts to malignant cells, oncogenic Ras mutants such as v-Ha-ras require Rho family GTPases (Rho, Rac, and CDC42) that are responsible for controlling actin-cytoskeleton organization. Ras activates Rac through a PI-3 kinase-mediated pathway. Rac causes uncapping of actin filaments (F-actin) at the plus-ends, through phosphatidylinositol 4,5 bisphosphate (PIP2), and eventually induces membrane ruffling. Several distinct F-actin/PIP2-binding proteins, such as gelsolin, which severs and caps the plus-ends of actin filaments, or HS1, which cross-links actin filaments, have been shown to suppress v-Ha-Ras-induced malignant transformation when they are overexpressed. Interestingly, an F-actin cross-linking drug (photosensitizer) called MKT-077 suppresses Ras transformation. Thus, an F-actin capping/severing drug might also have an anticancer potential. PURPOSE: This study was conducted to determine first whether Ras-induced malignant phenotype (anchorage-independent growth) is suppressed by overexpression of the gene encoding a large plus-end F-actin capping protein called tensin and second to test the anti-Ras potential of a unique fungal antibiotic (small compound) called chaetoglobosin K (CK) that also caps the plus-ends of actin filaments. METHODS AND RESULTS: DNA transfection with a retroviral vector carrying the tensin cDNA was used to overexpress tensin in v Ha-Ras-transformed NIH 3T3 cells. All stable tensin transfectants rarely formed colonies in soft agar, indicating that tensin suppresses the anchorage independent growth. The anti-Ras action of CK was determined by incubating the Ras-transformants in the presence of CK in soft agar. Two microM CK almost completely inhibited their colony formation, indicating that CK also suppresses the malignant phenotype. However, unlike tensin, CK causes an apoptosis of Ras transformed NIH 3T3 cells and, less effectively, of normal NIH 3T3 cells, indicating that CK has an F-actin capping-independent side effect(s). CK-induced apoptosis is at least in part caused by CK-induced inhibition of the kinase PKB/AKT. However, a specific ICE/caspase-1 inhibitor called N1445 completely abolished the CK-induced apoptosis by reactivating PKB, but without affecting the CK-induced suppression of Ras transformation. CONCLUSIONS: Like the F-actin cross linking drug MKT-077, the F-actin capping drug CK may be useful for the treatment of Ras-associated cancers if it is combined with the ICE inhibitor N1445, which abolishes the side effect of CK. Our observations that two distinct F-actin capping molecules (i.e., tensin and CK) suppress Ras-induced malignant phenotype strongly suggest, if not prove, that capping of actin filaments at the plus-ends alone is sufficient to block one of the Ras signaling pathways essential for its oncogenicity. This notion is compatible with the fact that Ras induces the uncapping of actin filaments at the plus-ends through the Rac/PIP2 pathway. PMID- 10526671 TI - Risk profiles to predict PSA relapse-free survival for patients undergoing permanent prostate brachytherapy. AB - PURPOSE: Combined risk profiles are helpful in assessing the likelihood of prostate-specific antigen relapse-free survival (PSA-RFS) for patients with localized prostate cancer. This retrospective analysis reports the 5-year biochemical outcome of a large cohort of patients according to prognostic factors and risk profiles for patients undergoing ultrasound-guided transperineal interstitial permanent prostate brachytherapy (TIPPB). PATIENTS AND METHODS: Seven hundred seventeen consecutive patients with clinically localized prostate cancer treated between June 1992 and November 1997 underwent TIPPB. Palladium-103 (103Pd) (n = 539) or iodine-125 (125I) (n = 178) sources were utilized to a prescribed dose of 120 Gy and 144 Gy, respectively. One hundred eleven patients received combined external-beam irradiation (41.4 or 45 Gy) and TIPPB (103Pd, 90 Gy, and 125I, 108 Gy). One hundred twelve patients received androgen ablation therapy in advance of TIPPB. Patients were grouped into risk profiles based on pretreatment factors that consisted of PSA < or = 10 and Gleason score < or = 6. Patients meeting both criteria were classified as favorable (n = 334). Those not fulfilling one criteria were classified as intermediate risk (n = 261), and those not fulfilling two criteria were classified as unfavorable risk (n = 261). PSA RFS was calculated based on the American Society of Therapeutic Radiology and Oncology consensus conference definition of PSA failure. The median follow-up was 41 months (range, 14-82 months). RESULTS: The actuarial PSA-RFS survival at 5 years was 82%. Multivariate Cox regression analysis identified PSA and Gleason score as highly significant factors predicting for biochemical outcome. Hazard risk ratios using a nadir definition of the PSA as < or = 1.0 ng/mL and < or = 0.5 ng/mL were each significant predictors of outcome with the 0.5 ng/mL nadir level only slightly better (risk ratio for nadir < or = 1.0 = 2.78; risk ratio for nadir < or = 0.5 = 3.57). Favorable-risk patients had a 5-year actuarial PSA RFS of 93%, whereas intermediate-risk and unfavorable-risk groups had 5-year PSA RFSs of 77% and 62%, respectively. Four hundred ninety-three patients underwent brachytherapy without external-beam irradiation or androgen ablation. From this group, the 5-year PSA-RFSs for the favorable-risk, intermediate-risk, and unfavorable-risk patients were 92%, 74%, and 55%, respectively. DISCUSSION: This retrospective study examining TIPPB demonstrated excellent 5-year actuarial PSA RFS rates. The assignment of risk profiles based on the results of multivariate analysis of prognostic factors identifies the expected PSA-RFS for patients undergoing TIPPB. PMID- 10526672 TI - Feasibility trial of postoperative radiotherapy and cisplatin followed by three courses of 5-FU and cisplatin in patients with resected head and neck cancer: a Southwest Oncology Group study. AB - BACKGROUND: Appropriate adjuvant chemotherapy for resected head and neck cancer patients has yet to be defined. Multiple trials have noted trends toward improved disease-free survival and local control. The Southwest Oncology Group undertook a feasibility trial of postoperative cisplatin and radiotherapy followed by three cycles of cisplatin and 5-fluorouracil. METHODS: Patients with resected stage III or IV head and neck cancer received cisplatin, 100 mg/m2, on days 1, 22, and 43 of radiotherapy. This therapy was followed by three cycles of cisplatin, 100 mg/m2 or last tolerated dose, and 5-fluorouracil, 1000 mg/m2, on days 1 to 4 every 21 days. RESULTS: Seventy-two patients from 22 institutions were registered; 68 were evaluable. Sixty-eight patients received radiotherapy. Only 25 of 68 patients (36.7%) were able to complete all six cycles of chemotherapy. Forty-three of 68 patients (63%) completed all three cycles with radiotherapy. Toxicities were tolerable. One toxic death occurred. CONCLUSIONS: It is not feasible to deliver six cycles of chemotherapy postoperatively in the sequence described. Compliance issues need further exploration to define effective adjuvant chemotherapy for head and neck patients. PMID- 10526673 TI - Essentials of immune response: the macrophage approach: the antibody approach. PMID- 10526675 TI - How to assess patient preference of migraine treatments. PMID- 10526676 TI - The difficulties of making rational treatment choices in migraine for the primary care physician. PMID- 10526677 TI - Is menstrually associated migraine difficult to treat? PMID- 10526678 TI - The use of isolates in migraine genetic research. PMID- 10526679 TI - A novel approach to the study of familial influences on evoked cortical responses in migraine. PMID- 10526680 TI - Measuring event-related potentials. PMID- 10526682 TI - The continuously evolving problems of acute ischemic coronary syndromes. PMID- 10526681 TI - Can factors that make a stable coronary artery plaque unstable be identified prospectively in the living patient? PMID- 10526683 TI - Crotchets (1999) PMID- 10526684 TI - Transvenous atrial defibrillation--techniques and clinical applications. AB - Atrial fibrillation (AF) is the most common arrhythmia requiring treatment. The most desirable therapy may be restoration and maintenance of sinus rhythm. Limitations of the current methods for cardioversion of AF have prompted the development of transvenous atrial defibrillation (TADF) as an alternative and more effective technique for converting AF. Recent advances in the technique of TADF, particularly in the design and configuration of the electrodes, and the use of an optimal biphasic shock waveform have dramatically improved the efficacy of TADF for the termination of all types of AF. The reduction in voltage and energy requirements for cardioversion by TADF may allow the procedure to be performed with little or no sedation and the risk of general anesthesia may be avoided. Both experimental and clinical studies have demonstrated the feasibility, safety, and efficacy of using TADF as a new temporary or "permanent" mode of electrical therapy for AF. It has several potential applications, from acute termination of AF in the electrophysiology laboratory and in patients who have failed to respond to external cardioversion, to its use as an implantable device for treating recurrent AF. This article reviews the current technique and clinical applications of TADF for treatment of AF. PMID- 10526685 TI - Effects of isosorbide dinitrate on electrocardiography, hemodynamics, and ventilation in patients with exercise-induced elevation of pulmonary artery wedge pressure. AB - BACKGROUND: The mechanisms underlying exertional hyperpnea in patients with coronary artery disease and transient left ventricular dysfunction are still not fully understood. HYPOTHESIS: The study was undertaken to investigate whether the ventilatory response to exercise reflects the effects of acute medical treatment of exercise-induced left ventricular dysfunction, and to evaluate mechanisms relevant to excessive exertional ventilation. METHODS: In 11 male patients, aged 65.2 +/- 6.0 years, all with pulmonary artery wedge pressure (PAWP) > 25 mmHg and ST depression > 2 mm during moderate supine exercise, ventilation (V), oxygen uptake (VO2), hemodynamics, electrocardiogram (ECG), and arterial and mixed venous blood gases were examined during supine rest and exercise, before and at hourly intervals after peroral intake of 30 mg isosorbide dinitrate (ISDN). Six similar patients were examined with the same protocol without ISDN administration and comprised a control group. RESULTS: Before administration of ISDN, exercise PAWP was 35.3 +/- 5.9 mmHg, ECG showed 2.77 +/- 1.06 mm ST depression, and V/VO2 was 31.8 +/- 4.8 l/l. One h after ISDN administration, exercise mean PAWP was 11.0 +/- 2.5 mmHg (p < 0.001), ST depression 0.59 +/- 0.8 mm (p < 0.001), whereas V/VO2 was unchanged, 30.1 +/- 5.3 l/l. Two h later, PAWP remained reduced and there were only minor ST depressions, while V/VO2 remained high. Exercise cardiac index (CI) and mixed venous oxygen tension (PvO2), initially 4.7 +/- 0.67 l/min/m2 and 3.54 +/- 0.35 kPa, respectively, remained at the same low level throughout the study. In the six nontreated patients, there were no significant changes in ST depression, exercise PAWP, or exertional ventilation. CONCLUSION: Isosorbide dinitrate treatment markedly improved exercise-induced left heart dysfunction, whereas excessive ventilatory response was unaffected, even after 3 h. Thus, measurements of the exercise hyperpnea did not properly reflect effective reduction of myocardial ischemia. PMID- 10526687 TI - The clinical utility of adenosine in difficult to diagnose tachyarrhythmias. AB - BACKGROUND: The use of intravenous adenosine to help differentiate the origin of tachyarrhythmias has been suggested to be beneficial. However, the benefit of this intervention to physicians with different levels of training in electrocardiographic (ECG) interpretation is unknown. HYPOTHESIS: The purpose of the study was to determine whether intravenous adenosine improved the diagnostic accuracy of difficult to diagnose tachyarrhythmias when used by physicians with different levels of training in ECG interpretation. METHODS: We studied 28 consecutive patients presenting with wide and narrow complex tachyarrhythmias, in whom adenosine was given specifically for diagnostic purposes. Two groups of physicians, attending (n = 14) and housestaff (n = 10), reviewed each ECG before and after the administration of adenosine. RESULTS: For narrow complex tachyarrhythmias, neither physician group derived diagnostic benefit from the use of adenosine. However, for wide complex tachyarrhythmias, the diagnostic accuracy of the housestaff group significantly improved with the use of adenosine (pre = 54%, post = 70%, p < 0.01), while the attending physician group had no significant improvement (pre = 61%, post = 71%, p = NS). CONCLUSION: This study suggests that adenosine provides useful diagnostic information to physicians less experienced in ECG interpretation when presented with patients having wide complex tachyarrhythmias of uncertain origin. PMID- 10526686 TI - The role of flumazenil in outpatient transesophageal echocardiography. AB - BACKGROUND: Benzodiazepines used for transesophageal echocardiography (TEE) sedation may be associated with postprocedural psychomotor effects that are undesirable. HYPOTHESIS: We hypothesize that flumazenil can reverse cognitive and motor effects from benzodiazepine, promoting earlier return to baseline function. METHODS: We prospectively evaluated the cognitive and motor function of patients who did or did not receive flumazenil following TEE. Patients' gait, level of drowsiness, and recall of items learned before and after benzodiazepine administration were evaluated before TEE, as well as immediately and 30 min after the procedure. RESULTS: Of 207 patients (123 men and 84 women), 93 (45%) were given flumazenil 0.2 or 0.4 mg intravenously, and 113 (55%) were not. The baseline characteristics of the patients who received flumazenil were not significantly different from those who did not receive flumazenil, with the exception of a higher mean dosage of midazolam administered to the flumazenil group. In addition, patients in the flumazenil group appeared more drowsy immediately following TEE. When adjusted for age and midazolam dosage, there were no differences, at any time, between the two groups in gait or recall of items learned prior to sedation. however, at 30 min following TEE, the flumazenil group was able to recall a larger number of new items learned immediately after the procedure (1.92/3 vs. 1.61/3, p = 0.02) than did patients in the group not receiving flumazenil. No adverse effects were encountered in any patient. CONCLUSION: Flumazenil appears safe and effective in reversing anterograde amnesic effects of benzodiazepine following TEE, but has no effects on retrograde amnesia and does not promote earlier return of motor function to baseline. It is useful in clinical situations where high dosages of benzodiazepine have been used and/or excessive drowsiness is evident following TEE. Routine use of the drug, however, is not necessary. PMID- 10526688 TI - Use of QT dispersion measured on treadmill exercise electrocardiograms for detecting restenosis after percutaneous transluminal coronary angioplasty. AB - BACKGROUND: Treadmill exercise electrocardiography (ECG) has been used to detect restenosis in patients following percutaneous transluminal coronary angioplasty (PTCA). However, the level of sensitivity achieved using conventional criteria of ST-segment depression is too low to be clinically useful in this population. HYPOTHESIS: QT dispersion is a sensitive method for detecting myocardial ischemia and may improve the accuracy of treadmill exercise ECG testing for detecting restenosis after PTCA. METHODS: We evaluated 104 patients who underwent PTCA for the treatment of single-vessel coronary artery disease and who had no history of myocardial infarction. Treadmill exercise ECG and coronary angiograms were performed 3 months after PTCA to determine the accuracy of diagnosis restenosis based on standard ST-segment depression and QT dispersion criteria. RESULTS: Restenosis was observed in 37 of the 104 patients (36%) 3 months after PTCA. QT dispersion immediately after exercise was significantly greater in patients with than in those without restenosis, as was the difference in QT dispersion before and immediately after exercise. The sensitivity, specificity, and accuracy of ST segment depression criteria were 59, 64, and 63%, respectively. Measurements of QT dispersion immediately after exercise (> or = 50 ms: positive, < 50 ms: negative) improved the sensitivity, specificity, and accuracy of treadmill ECG for predicting restenosis to 81, 87, and 85%, respectively. CONCLUSIONS: This novel diagnostic method using QT dispersion-based criteria significantly improves the clinical usefulness of treadmill exercise ECG for detecting the presence of restenosis after PTCA. PMID- 10526689 TI - Relation of ventricular repolarization to cardiac cycle length in normal subjects, hypertrophic cardiomyopathy, and patients with myocardial infarction. AB - BACKGROUND: Prolonged QT interval and QT dispersion have been reported to reflect an increased inhomogeneity of ventricular repolarization, which is believed to be responsible for the development of arrhythmic events in patients with long QT syndrome, coronary heart disease, and myocardial infarction, congestive heart failure, and hypertrophic cardiomyopathy (HC). HYPOTHESIS: This study was undertaken to determine whether an abnormal QT/RR dynamicity may reflect autonomic imbalance and may contribute to arrhythmogenesis in patients with heart disease. METHODS: The relation between QT, QTpeak (QTp), Tpeak-Tend (TpTe) intervals and cardiac cycle length was assessed in 70 normal subjects, 37 patients with HC, and 48 survivors of myocardial infarction (MI). A set of 10 consecutive electrocardiograms was evaluated automatically in each subject using QT Guard software (Marquette Medical Systems, Milwaukee, Wisc.). RESULTS: In patients with HC, all intervals were significantly prolonged compared with normals (p < 0.001 for QT and QTp; p < 0.04 for TpTc); in survivors of MI, this was true for the maximum QT and QTp intervals (p < 0.05). A strong linear correlation between QT, QTp, and RR intervals was observed in normals and in patients with MI and HC (r = 0.65-0.59, 0.82-0.77, 0.79-0.74, respectively, p < 0.0001). TpTe interval only showed a weak correlation with heart rate in normals (r = 0.24, p < 0.05) and was rate-independent in both patient groups (p = NS). Compared with normals, the slopes of QT/RR and QTp/RR regression lines were significantly steeper in patients with MI and HC (0.0990-0.0883, 0.1597-0.1551, 0.1653-0.1486, respectively). Regression lines were neither parallel nor identical between normals and patients (T > 1.96, Z > 3.07). There was no difference in steepness for TpTeR/RR lines between groups (0.0110, 0.0076, 0.0163, respectively). TpTe/QTp ratio was similar in normals and in patients with MI and HC (0.30 +/- 0.03, 0.31 +/- 0.07, 0.30 +/- 0.04, respectively), in the absence of any correlation between QTp and TpTe intervals, suggesting disproportional prolongation of both components of QT interval. CONCLUSION: Compared with normals, a progressive increase in QT and QTp intervals at slower heart rates in patients with MI and HC may indicate an enhanced variability of the early ventricular repolarization and may be one of the mechanisms of arrhythmogenesis. PMID- 10526690 TI - Global T-wave inversion: limited QT dispersion despite QTc prolongation--a correlate of benignity in patients with strikingly abnormal electrocardiograms. AB - BACKGROUND: The global T-inversion (GTI) electrocardiogram (ECG) is strikingly abnormal with major QTc prolongation, but with a surprisingly good prognosis by Kaplan-Meier curve. This contrasts with most significant QTc prolongations. HYPOTHESIS: This study was undertaken to ascertain QT interval dispersion (QTd) in global T wave inversion, a clinically benign long QTc ECG. METHODS: Longest and shortest QT intervals in all 12 leads in 35 consecutive patients with GTI were determined by two mutually blinded observers. QTd was determined by subtraction (maximum-minimum) and QTc was calculated using the Bazett formula. RESULTS: There was a 2:1 female preponderance QTc was prolonged and equal for men (0.471) and women (0.469). Observer variability of under 2% permitted averaging of QT measurements. Composite mean QTd was 55 ms. The literature revealed a range of QTd in normal subjects of 39 to 59 ms (mostly 49 to 59 ms). Patient series with abnormal QTd were well above this level. CONCLUSION: Despite a strikingly abnormal ECG with marked QTc prolongation, QT dispersion was limited in global T inversion, consistent with its previously demonstrated benignity. PMID- 10526691 TI - Prolongation of the QT interval in children with liver failure. AB - BACKGROUND: Alcoholic liver disease has been associated with QT prolongation and sudden cardiac death. HYPOTHESIS: We evaluated children with hepatic failure to determine whether they have abnormalities of ventricular repolarization. METHODS: Between October 1990 and January 1996, 38 pediatric patients (mean age 6.5 +/- 7.2 years) underwent evaluation for liver transplantation, including a 12-lead electrocardiogram and an echocardiogram. All patients had normal serum electrolytes, calcium, and magnesium at the time of cardiac evaluation and were not on any medications known to prolong repolarization. Follow-up electrocardiograms were performed on all survivors with QT prolongation following liver transplantation. RESULTS: Among those evaluated, seven (18%) were noted to have a prolonged QT interval corrected for rate (QTc > 450 ms; range 460-560 ms). All had a structurally normal heart, except one with an atrial and ventricular septal defect. When compared with patients with a normal QT interval, there was no significant difference in serum indices of liver function or indication for liver transplantation. None of the patients developed a ventricular arrhythmia. Two patients with a prolonged QTc died prior to transplant and another died immediately after surgery. All four survivors had normalization of the QTc following liver transplantation. CONCLUSION: QTc prolongation can be seen in a significant number of children with hepatic failure. While the mechanism is not known, it appears to be reversible following liver transplantation. PMID- 10526692 TI - Intrapulmonary artery infusion of urokinase for treatment of massive pulmonary embolism: a review of 26 patients with and without contraindications to systemic thrombolytic therapy. AB - BACKGROUND: Pulmonary emboli (PE) are a common event seen in over 600,000 patients a year. Occurring suddenly, PE often result in a high rate of mortality. To combat the high rate of mortality, more aggressive therapies including the use of thrombolytics are often indicated. The use of intrapulmonary artery infusion of urokinase has been shown to promote rapid resolution of emboli and restoration of normal pulmonary hemodynamics. HYPOTHESIS: The study was undertaken to review the effectiveness and safety of pulmonary artery infusion of urokinase in 26 patients with and without contraindications to the use of systemic thrombolytic therapy. METHODS: We reviewed the outcomes of 26 patients who received infusion of urokinase, using a usual loading dose of 4,000 U/kg body weight given as a bolus, followed by 4,000 U/kg/h for 12 to 24 h, using either/or unilateral or bilateral infusions. Pulmonary angiograms were obtained prior to and following the urokinase infusions. RESULTS: Intrapulmonary artery infusion of urokinase was given to 26 patients, 9 of whom had contraindications to the use of systemic thrombolytic therapy. Six patients were recent post operative, one was receiving oral anticoagulants, one was receiving chemotherapy with bleeding complications, and one had received cardiopulmonary resuscitation. Twenty of the patients returned to their baseline state (normal heart rate, blood pressure, and p02), one was minimally improved, and five deaths occurred. Of the five deaths, three occurred within 1 h of starting urokinase infusion, the remaining two died more than 36 h after treatment with urokinase as a result of their basic underlying disease. Minor bleeding occurred from puncture sites, two hematomas occurred at the puncture site, and there were two gastrointestinal bleeds, one of which occurred a week post urokinase therapy while the patient was receiving heparin and coumadin. No central nervous system bleeds occurred and no transfusions were required as a result of urokinase intrapulmonary artery infusions. The overall mortality rate in this series was 11.5%. CONCLUSIONS: Intrapulmonary artery infusion of urokinase in extensive pulmonary embolism is a safe and efficient treatment in patients with and without contraindication to the use of systemic thrombolytic therapy. With a usual loading dose of 4,000 U/kg body weight, followed by an infusion of 4,000 U/kg/h for 12 to 24 h, it produces significant and rapid resolution of pulmonary emboli with a low morbidity and mortality rate. In our series, the mortality rate was 11.5%, and none of the deaths was the direct result of urokinase therapy. PMID- 10526693 TI - Irregularity of the ventricular rhythm during atrial fibrillation: effect of slow atrioventricular nodal pathway ablation. AB - BACKGROUND: The contribution of dual atrioventricular (AV) nodal pathway physiology to the irregularity of the ventricular rhythm during atrial fibrillation has not been clarified. HYPOTHESIS: This study was performed to assess the effects of slow AV nodal pathway ablation on the irregularity of the ventricular rhythm during atrial fibrillation. METHODS: Irregularity of the ventricular rhythm was quantified using analysis of heart rate variability. In 20 patients with AV nodal reentrant tachycardia, absolute heart rate variability during atrial fibrillation was quantified before and after slow AV nodal pathway ablation by the standard deviation of all NN intervals (SDNN). Relative heart rate variability was determined by computing the coefficient of variation, SDNN normalized for the standard deviation of the mean ventricular cycle length (MVCL AF). RESULTS: The slope of the regression between MVCL-AF and SDNN was significantly more gradual after slow pathway ablation (slope 0.39 vs. 0.23, p < 0.001). Coefficient of variation increased in 12 patients with heart rates > 120 beats/min at baseline (18.6 +/- 3.9 vs. 22.1 +/- 2.7% MVCL-AF, p < 0.05), but decreased in 8 patients with heart rates < 120 beats/min at baseline (25.6 +/- 3.1 vs. 22.2 +/- 2.2% MVCL-AF, p = 0.05). Furthermore, coefficient of variation correlated with MVCL-AF only at baseline (slope 0.034, r = 0.66), but no relation was found after slow pathway ablation (slope 0, r = 0). CONCLUSIONS: Slow AV nodal pathway ablation alters the relation between absolute heart rate variability and mean ventricular rate during atrial fibrillation and eliminates cycle length dependency of relative heart rate variability. These data indicate that dual AV nodal pathway physiology contributes to the irregularity of the ventricular rhythm during atrial fibrillation. PMID- 10526694 TI - Pulmonary embolus in transit. PMID- 10526695 TI - Coil embolization of hepatoportal arteriovenous fistula in a neonate. PMID- 10526696 TI - Spontaneous multivessel coronary artery dissection: repeated presentation in a healthy postmenopausal woman. AB - Spontaneous coronary artery dissection is a rare cause of acute myocardial infarction which is infrequently diagnosed antemortem. Most previously reported cases were found in women of whom a significant proportion presented during pregnancy or the postpartum period. We describe the first antemortem case of spontaneous coronary artery dissection, unrelated to pregnancy or the postpartum state, which ultimately resulted in diffuse involvement of both the left and right coronary arteries over a period of 4 months. Pathophysiology and case management of this disorder are discussed. PMID- 10526697 TI - From hypertension to heart failure: what have we learned? AB - Hypertension is associated with an increased risk for heart failure, stroke, and end-stage renal disease. The mechanisms involved in progression from hypertension to heart failure have been the focus of many recent studies. In addition, we have learned much from epidemiologic studies that have helped identify risk factors for hypertension and have thus provided insight into mechanisms that are involved in the pathogenesis of this disease. This paper will consider the epidemiology of both hypertension and heart failure and their relationship with left ventricular hypertrophy. In addition, results of recent clinical trials of antihypertensive agents will be reviewed. PMID- 10526698 TI - Prevention of sudden cardiac death with beta blockers. AB - Beta blockers have been shown to reduce the risk of sudden cardiac death in more than 50 randomized trials involving more than 55,000 patients. Relative reductions (vs. placebo) in cardiac death in some of these trials ranged from 30 to 50%. These reductions are substantially greater than trials of other drug classes including angiotensin-converting enzyme inhibitors. However, not all beta blockers confer equal benefit to patients at risk of sudden cardiac death. Results from various trials suggest that lipophilic beta blockers--such as timolol, metoprolol, propranolol, bisoprolol, and carvedilol--may be more beneficial than hydrophilic beta blockers. Results of animal studies have indicated that sudden cardiac death is mediated, at least in part, by the central nervous system, which may account for why lipophilic agents have more pronounced clinical effects. Based on the results of numerous clinical and mechanistic studies, it is suggested that beta blockers should be given to all patients at risk for sudden cardiac death, including those patients with previous myocardial infarction, hypertension, or congestive heart failure. PMID- 10526699 TI - The cellular and physiologic effects of beta blockers in heart failure. AB - Enhanced and sustained cardiac adrenergic drive occurs in heart failure (HF) and contributes, in part, to the progression of left ventricular (LV) dysfunction and remodeling that are characteristic of this disease state. Enhanced sympathetic drive in HF can lead to downregulation and desensitization of cardiac beta adrenergic receptors with a consequent impairment of myocardial reserve and exercise tolerance. This sympathoadrenergic maladaptation can also lead to cellular abnormalities in the failing heart, manifested by defects in calcium handling of the sarcoplasmic reticulum, by defects in myocardial energetics, and by ongoing loss of cardiomyocytes through necrosis or apoptosis. Chronic treatment with beta blockers in patients with HF and in animals with experimentally induced HF has been shown to reverse, prevent, or, at the least, arrest many, if not all, of these adverse processes. Beta blockers improve function of the failing LV, prevent or reverse progressive LV dilation, chamber sphericity, and hypertrophy, and consequently have positive impact on cardiac remodeling. Beta blockers also reduce heart rate and LV wall stress, leading to reduced myocardial oxygen consumption, a clear benefit to the failing heart. Beta blockers can also improve the intrinsic contractile function of cardiomyocytes and have also been shown to improve myocardial energetics in HF, possibly through desirable changes in substrate utilization. Recent studies from our laboratories have also shown that beta blockers can attenuate cardiomyocyte apoptosis in HF. These benefits provide strong reinforcement to the clinical findings that beta blockers are highly beneficial for the management of patients with chronic HF and, when properly used, afford unequivocal reductions in mortality and morbidity in this patient population. At present, there is general agreement that increased cardiac sympathetic drive occurs in HF and may potentially be an important contributor to the progression of LV dysfunction and chamber remodeling that is characteristic of this disease state. Experimental studies in animal models of HF as well as clinical studies in patients with HF have suggested that chronic therapy with beta blockade is effective in preventing the progression of LV dysfunction and remodeling, the latter evidenced by reversal and/or prevention of progressive LV dilation and chamber sphericity. Results of recent multicenter clinical trials support these findings and have made it abundantly clear that long-term therapy with beta blockade inhibits clinical progression and has a major impact on mortality and morbidity in patients with HF that is at least as favorable, if not better, than that observed with angiotensin-converting enzyme (ACE) inhibitors. Beta blockers improve mortality and morbidity in HF and also improve LV ejection fraction (EF), a beneficial feature that, until recently, has only been attributed to positive inotropic agents. PMID- 10526700 TI - Experience with beta blockers in heart failure mortality trials. AB - Recent investigations have indicated that chronic heart failure can be reversed with agents that inhibit the reninangiotensin-aldosterone or sympathetic nervous system, such as angiontensin-converting enzyme (ACE) inhibitors and beta blockers. A meta-analysis of clinical trials of ACE inhibition in chronic heart failure reported reductions in mortality ranging from 13 to 33%, but as ACE inhibitors do not block chronic noradrenergic stimulation of the heart, mortality remains unacceptably high. Beta blockers have been shown to increase left ventricular ejection fraction, reduce end-systolic and end-diastolic cardiac dimensions, improve quality of life, and reduce mortality. All-cause mortality in the US Carvedilol trial was reduced 65%, and in MERIT-HF there was a 49% reduction in mortality from heart failure among patients receiving metoprolol CR/XL. MERIT-HF was ended early because of evidence of survival benefit. Although certain effects of beta blockers may be considered class effects, it is not yet clear whether there are differences between beta 1-selective antagonists and nonselective agents. The benefits conferred across differences in disease severity, race, and age should be answered as large ongoing and planned clinical trials of beta blockers are completed. PMID- 10526701 TI - The mortality effect of metoprolol CR/XL in patients with heart failure: results of the MERIT-HF Trial. AB - The study was undertaken to investigate the effect of metoprolol CR/XL on all cause mortality in patients with heart failure in New York Heart Association (NYHA) class II-IV. In all, 3,991 patients in NYHA class II-IV who were stable on standard medical treatment, including angiotensin-converting enzyme inhibitors, diuretics, and digitalis, were randomized to metoprolol CR/XL or placebo and uptitrated from 12.5 or 25 mg to 200 mg over an 8-week period and were planned to be followed for a period of 2 years. The study was stopped earlier than planned due to the significant benefit achieved with metoprolol CR/XL on all-cause mortality. Treatment with metoprolol CR/XL was associated with a 34% decrease in all-cause mortality, 38% decrease in cardiovascular mortality, 41% decrease in sudden death, and 49% decrease in death due to progressive heart failure. The average dose of metoprolol CR/XL at the end of the study was 159 mg, and 64% of the patients were receiving 200 mg of metoprolol CR/XL. There was no significant difference in the placebo and active treatment group with regard to permanent discontinuation. Treatment of patients in NYHA class II-IV with metoprolol CR/XL is associated with a significant decrease in total mortality. PMID- 10526702 TI - Trials of glycoprotein IIb-IIIa inhibitors in non-ST-segment elevation acute coronary syndromes: applicability to the practice of medicine in the United States. AB - Platelet-mediated thrombosis has been recognized as the primary pathophysiologic mechanism of acute coronary syndromes (ACS) and acute complications of percutaneous coronary intervention (PCI). Despite the clinical efficacy of the two most widely used antithrombotic agents, aspirin and heparin, each of them has significant therapeutic limitations. As a result, thrombosis and clinical events may occur despite the use of aspirin and heparin. The discovery that the platelet glycoprotein (GP) IIb-IIIa represents the final common pathway to platelet aggregation and the growing recognition of the key role of platelets in the progression of thrombosis prompted the development of several GP IIb-IIIa inhibitors as a potentially more effective form of antithrombotic therapy. Numerous trials of various GP IIb-IIIa inhibitors as adjuncts to PCI have strongly supported this hypothesis. The subject of this supplement is the review of more recent evaluations of GP IIb-IIIa inhibitors in the context of various treatment strategies for the management of patients with unstable angina or non ST-segment elevation myocardial infarction, collectively known as non-ST-segment elevation ACS. Appropriate translation of these trials into clinical practice requires not only the knowledge of the trials' results but also the understanding of the design of individual studies, most notably the entry criteria and patient management strategies. PMID- 10526703 TI - The secondary prevention of myocardial infarction. PMID- 10526704 TI - The history of liposuction and fat transplantation in America. AB - Dermatologists have played an important role in the development of both liposuction and fat transplantation in the United States. Dermatologists were among the earliest physicians to embrace these new procedures and have been responsible for a number of breakthroughs and refinements in these techniques. Klein's development of the tumescent technique has profoundly altered the way liposuction and fat transplantation are performed. These procedures continue to be increasingly popular among dermatologic surgeons. PMID- 10526705 TI - Preoperative evaluation of the liposuction patient. AB - Tumescent liposuction is a safe and effective procedure. It is best used in normal individuals with localized areas of adiposity and good overlying skin tone. Patients must have realistic goals and expectations. Meticulous surgical technique is essential, and care must be taken to remove the correct amount of fat in the appropriate plane. Careful patient selection following a preoperative assessment will result in a very satisfying procedure for both the patient and physician. PMID- 10526706 TI - Instrumentation for liposuction. AB - This article presents a comprehensive review of instrumentation for liposuction. It discusses equipment for achieving anesthesia and aspirating fat. In addition, techniques for maintaining sterility, patient monitoring, and postoperative care are presented. Emphasis is made on patient safety. PMID- 10526708 TI - Ultrasonic-assisted liposuction. Internal and external. AB - UAL has not provided the promised ideal of "fat dissolution without surgery." In extremely fibrous areas and second procedure liposuction, internal UAL may be a valuable tool once it is further perfected. UAL equipment continues to evolve and as it improves we hope to see a better safety profile, a mechanism for smaller entrance sites, and greater time efficiency in the procedure. PMID- 10526707 TI - Anesthetic formulation of tumescent solutions. AB - There is no standard or official recipe for the tumescent anesthetic solutions. The actual concentrations of lidocaine and epinephrine should depend on the areas to be treated and clinical situation. This article discusses the safe usage of tumescent solutions and the proper procedures and precautions to take when mixing these solutions. PMID- 10526709 TI - "Cook Weekend Alternative to the Facelift". Liposculpture of the face, neck, and jowls with laser dermal resurfacing and platysmal plication. AB - The "Cook Weekend Alternative to the Facelift" is a combined liposculpture and laser surgical procedure which produces excellent cosmetic results, in many cases comparable to the results of traditional surgical rhytidectomy, without the extensive surgical intervention and prolonged recovery time needed for rhytidectomy. The procedure consists of liposculpture of the face, neck, and jowls; laser resurfacing of the platysma and underside of the dermis; vaporization of subcutaneous fat; resection of a small ellipse of excess submental skin; separation of the neck septa; and plication of the platysma, with or without chin augmentation. Cosmetic results can be dramatic, and most patients return to normal activities approximately three days postoperatively. PMID- 10526710 TI - Liposuction of the arms. AB - Liposuction of arms when properly performed with realistic expectations is almost always a "patient pleaser." Patients routinely marvel at the degree of skin contraction that typically occurs. Before and after photos, even as early as 1 week, routinely show dramatic skin contraction when significant volumes of fat are removed although textural changes may evolve for weeks to months (Figs. 14 19). Only the lower abdomen and neck consistently obtain such profound and predictable contraction. I am convinced that historically liposuction of the arms has been performed in a substandard fashion. Inadequate fat removal will often produce irregularity and will always result in less than maximal skin contraction. As I have performed progressively larger-volume cases, the indications for brachioplasty, in my opinion, are nearly nonexistent. My present approach, except in the most extreme cases, is to initially recommend liposuction and possibly even a second liposuction prior to performing or recommending brachioplasty. Even massive arms with good skin tone will usually obtain an aesthetically pleasing result when treated properly. Massive arms with poor skin tone, however, may not. One does not "burn bridges," however, by performing liposuction alone in these questionable candidates. If brachioplasty is subsequently desirable, in spite of the major drawback of the resultant scar, it can be performed at a later date. In summary, the key concepts for maximizing the potential of liposuction of the arms are to perform thorough but gentle fat removal and to avoid immediate subdermal fat removal or trauma to the underside of the dermis. PMID- 10526711 TI - Liposuction of the chest and back. AB - Subcutaneous fat deposition on the chest and back is notoriously diet and exercise resistant. The "top heavy" appearance is a source of frustration for patients as well as surgeons. Fortunately, with advent of tumescent liposuction surgery the disparity between the upper and lower torso can now be reconciled. PMID- 10526712 TI - "Three-dimensional tumescent liposculpture" of the abdomen, waist, and flanks. AB - Treating the entire area as a cosmetic unit is the best approach to tumescent liposculpture of the abdomen, waist, and flanks. This "Three-Dimensional Tumescent Liposculpture" procedure is performed under tumescent local anesthesia with optional intraoperative external ultrasound. Areas treated are the abdomen, waist, flanks, and infrascapular fat pad if indicated. Postoperatively, patients show a flatter abdomen, a smaller and better-defined waist (the "Cook waist"), reduction of unsightly bulges, and a smoother, better proportioned and more attractive overall contour. Patient recovery is rapid with minimal complications. PMID- 10526713 TI - Contouring the female buttocks. Liposculpting the buttocks. AB - Body sculpting has progressed during the past decade to a point where cosmetic units may be sculpted to improve contours which blend imperceptibly and appropriately with adjacent cosmetic units. The buttocks is ideally suited for sophisticated contouring as its frame is determined by the hips, thighs, and lower back and its proportions are balanced by the anterior projection of the breasts. In addition, ethnic differences in the shape and proportions of the buttocks create a variety of aesthetically pleasing variations in size and shape. The article attempts to elucidate these considerations combined with a logical surgical approach to achieve pleasing results in body sculpting. PMID- 10526714 TI - Liposuction of the abdomen. The basics. AB - As one of the most frequent regions treated by liposuction in both men and women, the abdomen presents unique learning opportunities for the liposuction surgeon. Because of anatomic variation, the upper abdomen is more fibrous than the lower abdomen, requiring a slightly different approach to achieve optimal fat removal. The periumbilical area also provides unique challenges for the operator and care must be taken to avoid leaving a ring of residual fat. Potentially excellent skin retraction in the area, combined with relatively aggressive fat removal can lead to dramatic results including, in some cases, significant contraction of a long standing panniculus. In this article, the basic techniques and potential pitfalls of abdominal liposuction are presented in detail. PMID- 10526715 TI - Liposuction of the abdomen. An analysis of form. AB - This article presupposes that the surgeon has complete command of the process or craft of liposuction. This consideration of form does not presume to issue a commentary upon the ultimate fitness of the form; rather, form confines itself with shape. The final question that matters is, "has the surgeon used the craft to successfully alter the three-dimensional mass into an aesthetically pleasing end." PMID- 10526716 TI - Liposuction of the thigh. AB - This article discusses liposuction of the thigh. All aspects of preoperative evaluation, anesthesia, operative technique, and postoperative care are reviewed. Issues relating to safety are emphasized as are aesthetic dimensions of liposuction of the thigh. PMID- 10526717 TI - Liposuction of the knees, calves, and ankles. AB - Lipodystrophy of the knees, calves, ankles, and neck are somewhat unique in comparison to other sites because they are more difficult to camouflage. The calves and ankles, in particular, are less dependent on body weight and more resistant to diet and exercise. Calf and ankle lipodystrophy is usually present from early adolescence. PMID- 10526718 TI - Post-tumescent liposuction care. Open drainage and bimodal compression. AB - The goals of post-liposuction care must be to minimize edema, bruising, and patient discomfort. The postoperative pain and edema resulting from sutured incisions and prolonged post-liposuction compression is an irrational remnant from the days before the tumescent technique. This article discusses various issues involving post-liposuction care. PMID- 10526719 TI - Fat transplantation. AB - Fat transplantation has been used by physicians for over 100 years; however, this technique has been refined considerably since the development of liposuction in the late 1970s. Fat transplantation can be used for augmentation of subcutaneous defects throughout the body. It continues to be the safest and most versatile form of soft tissue augmentation. PMID- 10526720 TI - Morbidity and mortality related to liposuction. Questions and answers. AB - Reports of fatalities following liposuction have lead to investigations by state medical boards. The risk of complications and fatalities is clearly different for liposuction under local anesthesia and intravenous sedation. Thousands of patients have been treated with true tumescent liposuction as described by dermatologist Dr. Jeffrey A. Klein, with no reports of fatalities. Patients should seek physicians who are experienced in this extremely safe method of liposuction. PMID- 10526721 TI - Blood lipid levels in type 2 diabetes. What are the effects of diet? PMID- 10526722 TI - Effect of the fast-acting insulin analog lispro on the risk of nocturnal hypoglycemia during intensified insulin therapy. U.K. Lispro Study Group. AB - OBJECTIVE: To measure the effectiveness of insulin lispro, a fast-acting insulin analog, in reducing hypoglycemic episodes when used in a basal bolus regimen by patients with type 1 diabetes using intensive insulin therapy. RESEARCH DESIGN AND METHODS: In 11 diabetes outpatient clinics in the U.K., 165 subjects with type 1 diabetes were enrolled in a randomized crossover open-label study with a 2 month run-in period and then treated with a basal bolus regimen. Patients used human NPH insulin at night with either premeal insulin lispro for 4 months followed by human regular insulin for another 4 months or human regular insulin for 4 months followed by insulin lispro for another 4 months. The main outcome measures were the number of hypoglycemic episodes during both treatments and HbA1c level. RESULTS: A total of 135 patients were randomized, with 68 receiving insulin lispro and 67 receiving human regular insulin for the first 4 months. The data for the first 4 months of treatment only were compared as two independent groups because of a period effect and a treatment-period interaction. Glycemic control was equally tight during treatment with human regular insulin (HbA1c, 6.2 +/- 0.8%) and insulin lispro (6.0 +/- 0.9%). A total of 1,156 hypoglycemic episodes occurred during treatment with human regular insulin compared with 775 hypoglycemic episodes that occurred during treatment with insulin lispro (P = 0.04). This difference was chiefly because of a reduced number of nocturnal episodes (181 vs. 52, P = 0.001) in the insulin lispro group. CONCLUSIONS: The use of a fast-acting insulin analog, insulin lispro, as part of a basal bolus regimen reduces nocturnal hypoglycemia in patients with type 1 diabetes who maintain tight glycemic control during intensive insulin therapy. PMID- 10526723 TI - Improvements in diabetic care as measured by HbA1c after a physician education project. AB - OBJECTIVE: To measure the quality of diabetic care as indicated by HbA1c testing frequency and HbA1c values and to demonstrate improvement in care after an appropriate quality improvement intervention. RESEARCH DESIGN AND METHODS: The quality improvement project used computerized claims and laboratory data relating to HbA1c testing among the private practices of nine physicians caring for diabetic Medicare patients. Nine indicators evaluated three main areas: HbA1c testing frequency, HbA1c values, and frequency of office visits. A quality improvement intervention consisting of a physician component and a patient component was implemented. RESULTS: There were 835 patients and 4,367 visits studied. After the intervention, statistically significant improvements in HbA1c testing frequency and values were noted. Rates of seized opportunities for testing HbA1c improved from 17.7 to 33.9% (P < 0.0001). The percentage of patients with a current HbA1c value improved from 31.3 to 47.6% (P < 0.0001). The median HbA1c values fell from 8.5 to 7.8% (P < 0.006). Patients achieving good or fair control (HbA1c < or = 8%) improved from 43.8 to 56.9% (P = 0.007). The median time between physician visits fell from 70 days to 60 days (P < 0.0001). CONCLUSIONS: The study revealed that HbA1c testing was underused but that after a quality improvement initiative, a significant increase in testing use could be achieved. The quality improvement initiative also resulted in significant improvements in glycemic control. The techniques and interventions used in this study could be used to intervene in larger populations and practice settings to improve medical care for diabetic patients. PMID- 10526724 TI - Diabetes in urban African-Americans. XV. Identification of barriers to provider adherence to management protocols. AB - OBJECTIVE: To determine whether health care providers appropriately identify patients with poor glycemic control and to investigate reasons why providers may fail to intensify therapy in these patients. RESEARCH DESIGN AND METHODS: Our management protocol calls for providers to advance diabetes therapy in patients with fasting plasma glucose levels > 7.8 mmol/l or random plasma glucose levels > 10.0 mmol/l. During a 3-month period, providers completed a questionnaire at the end of individual patient visits by asking whether the patient was well controlled and whether therapy was advanced. If therapy was not advanced in patients perceived to have poor control, providers were asked to provide a justification. RESULTS: Providers appropriately identified 88% of well-controlled patients and 94% of patients with poor glycemic control. Out of 1,144 patient visits, control was reported to be good in 508 and poor in 636. In these 636 visits, therapy was advanced in 490 but not in 146 visits. The dominant reasons for failure to intensify therapy were the perception by the provider that control was improving (34%) or the belief that the patient was not compliant with diet or medications (25%). Less common reasons included acute illness, patient refusal, and recurrent hypoglycemia. Based on fasting glucose levels, protocol adherence was 55% before the questionnaire, 64% during the questionnaire (P = 0.006), and 63% afterwards. CONCLUSIONS: Providers in a specialty diabetes clinic appropriately classified patients according to glycemic control and tended to intensify therapy when indicated in most poorly controlled patients. Provider self-survey of behavior and decision making may be an effective strategy to improve adherence to management protocols. PMID- 10526725 TI - Short needles (8 mm) reduce the risk of intramuscular injections in children with type 1 diabetes. AB - OBJECTIVE: To study whether 8-mm needles can reduce the frequency of intramuscular injections in diabetic children. RESEARCH DESIGN AND METHODS: We conducted a prospective crossover study in 50 children whose BMI was < or = 60th percentile to compare two lengths of needles (12.7 and 8 mm) regarding the occurrence of intramuscular injections as assessed by ultrasonography. RESULTS: The frequency of intramuscular injections was 86% with the 12.7-mm needles and 38% with the 8-mm needles. The frequency of intramuscular injections was significantly reduced when using the 8-mm needles in the arms (P < 0.01) and thighs (P < 0.001). The efficiency of 8-mm needles, as defined by an intramuscular injection with a 12.7-mm needle and a subcutaneous injection with an 8-mm needle, was found for half of the children who injected in the arm and for two-thirds of the children who injected in the thigh. The subcutaneous tissue (SQT) thickness measured by ultrasonography with a skinfold was significantly higher (9.8 +/- 2.2 mm) in the group in which the 8-mm needles were efficient than in the group in which they were not efficient (6.8 +/- 2.1 mm, P < 0.0001). The efficiency of the 8-mm needle was not related to age, sex, BMI, percentile of BMI, injection device, or injection site. The sensibility and specificity of SQT thickness in predicting the efficiency of the 8-mm needles were both 79%. CONCLUSIONS: Needles that are 8 mm long significantly reduce the risk of intramuscular insulin injection in slim or normal-weight (BMI < or = 60th percentile) diabetic children and adolescents. PMID- 10526726 TI - Vitamin E supplementation and oxidative damage to DNA and plasma LDL in type 1 diabetes. AB - OBJECTIVE: To determine the effect of 400 IU/day of the antioxidant vitamin E on the susceptibility of plasma LDL and lymphocyte DNA to oxidative damage in type 1 diabetes. RESEARCH DESIGN AND METHODS: We studied 42 patients with type 1 diabetes and 31 age- and sex-matched control subjects in a randomized prospective double-blind placebo-controlled trial by using 400 IU/day of oral vitamin E for 8 weeks. Measurements were made of single-strand breaks in lymphocyte DNA at baseline and after hydrogen peroxide-induced stress (comet assay) and of copper induced LDL oxidization and plasma antioxidant profiles. RESULTS: Plasma LDL and lymphocyte DNA were more resistant to induced oxidative change in the type 1 diabetes group than in control subjects. Vitamin E supplementation reduced LDL oxidizability in the control subjects but not in the type 1 diabetes group and had no effect on oxidative DNA damage in either group. The type 1 diabetes group had a significantly poorer plasma antioxidant profile with lower mean serum concentrations of alpha-tocopherol and most carotenoids than control subjects. CONCLUSIONS: Plasma LDL and lymphocyte DNA appear to be more resistant to oxidative change in type 1 diabetic subjects than in control subjects, and there was no evidence of oxidatively induced DNA or LDL change in type 1 diabetes. This study does not support the hypothesis of oxidative damage in these patients, and a dose of vitamin E (400 IU/day) that reduced LDL oxidative susceptibility in control subjects did not do so in patients with type 1 diabetes. PMID- 10526727 TI - Heterogeneity in associations between macronutrient intake and lipoprotein profile in individuals with type 2 diabetes. AB - OBJECTIVE: To evaluate associations between macronutrient intake and lipoprotein profile among individuals with type 2 diabetes who participated in the San Luis Valley Diabetes Study (SLVDS) or the Insulin Resistance Atherosclerosis Study (IRAS). RESEARCH DESIGN AND METHODS: Diet was assessed by 24-h recall in the SLVDS (n = 421) and by validated food frequency interview in the IRAS (n = 437). Analyses adjusted for kilocalories, age, sex, and other covariates were conducted separately for the two study groups. For the SLVDS, repeated observations were included in mixed model analyses (865 observations). For the IRAS, standard regression analyses were conducted. Recent weight history and time of diabetes diagnosis were evaluated as possible modifiers of associations between nutrient intake and lipoprotein profile. RESULTS: Higher reported intake of total dietary fat was related to significantly higher levels of LDL cholesterol (P < 0.05) in both studies and in all subgroups. Reported intake of total and saturated fat was associated positively with total cholesterol, although statistical significance was not reached for all subgroups. Higher reported carbohydrate intake was associated with increased triglyceride concentrations (P < 0.01) only among individuals with previously undiagnosed diabetes in the SLVDS (n = 69) and only among individuals who gained weight (> 5 lb, n = 87) during the previous year in the IRAS. CONCLUSIONS: Toward the goal of optimizing the lipoprotein profile of individuals with diabetes, these results emphasize the potential importance of reducing fat intake while recognizing that individualized approaches to diet are important to minimize the risk of cardiovascular disease. PMID- 10526728 TI - Effects of exercise training on oxygen uptake kinetic responses in women with type 2 diabetes. AB - OBJECTIVE: Women with uncomplicated type 2 diabetes have both a decreased maximal oxygen consumption (VO2max) and slowed oxygen uptake (VO2) kinetics at the onset of exercise compared with nondiabetic women. These abnormalities are seen not only at maximal workloads, but also at the onset of low-level exercise. To evaluate the hypothesis that VO2max and VO2 kinetics would improve with exercise training in untrained people with type 2 diabetes, we measured these parameters in premenopausal sedentary women before and after 3 months of supervised exercise training. RESEARCH DESIGN AND METHODS: A total of 8 women with type 2 diabetes, 9 overweight nondiabetic women, and 10 lean nondiabetic women were studied. At baseline and after 3 months of exercise training, subjects underwent bicycle ergometer testing to obtain VO2max and VO2 kinetics data. RESULTS: On entry, women with type 2 diabetes had the lowest VO2max and slowest VO2 kinetics of the three groups. After exercise training, the women with type 2 diabetes improved their VO2max more than the lean and overweight control women: 28 vs. 5 and 8%, respectively (P < 0.05 for the diabetic group vs. both control groups). In the group with diabetes, VO2 kinetics improved by 39 and 22% at 20 and 30 W, respectively. For the control subjects, VO2 kinetics did not improve at any workload in either group. CONCLUSIONS: Despite beginning with the lowest VO2max and slowest VO2 kinetics, subjects with type 2 diabetes benefited more from an exercise training program than did control subjects. These findings suggest that in addition to its known metabolic effects, exercise training in individuals with type 2 diabetes may be an effective therapy to improve the cardiovascular response to exercise and to overcome low-level exercise impairment as reflected by improved VO2max and VO2 kinetics. If the ability to make circulatory adjustments at the beginning of exercise at low workloads is improved by an exercise training program, as suggested by the VO2 kinetics data, the clinical significance of exercise for people with type 2 diabetes is clear. PMID- 10526729 TI - Interaction of sulfonylureas and exercise on glucose homeostasis in type 2 diabetic patients. AB - OBJECTIVE: To determine whether the plasma glucose-lowering effects of sulfonylureas and acute submaximal exercise are additive and, accordingly, to determine whether they may increase the risk of hypoglycemia when combined in fasting patients. RESEARCH DESIGN AND METHODS: Eight postabsorptive type 2 diabetic patients were examined at three occasions: after oral sulfonylurea (7 mg glibenclamide), during 60 min of ergometer cycle exercise at 57 +/- 3% of VO2max, and during exercise after glibenclamide. RESULTS: Heart rate, VO2, and lactate responses to exercise were comparable (P > 0.05) on days with and without glibenclamide. Plasma insulin concentrations were always increased by glibenclamide, and they were lowered identically by exercise with and without glibenclamide. However, throughout exercise, absolute concentrations of insulin were lower on days without glibenclamide compared with days with glibenclamide (34.5 +/- 4.7 vs. 47.4 +/- 5.5 pmol/l; P < 0.05). At the start of exercise, glucose concentrations were similar between experiments (P > 0.05). The rate of decrease in glucose during exercise was higher (P < 0.05) on days with both glibenclamide and exercise, compared with days with glibenclamide alone and days with exercise alone (-0.035 +/- 0.009 vs. -0.016 +/- 0.002 and -0.022 +/- 0.005 mmol.l-1.min-1, respectively). Consequently, the glucose nadir was lower on days with glibenclamide and exercise than on days with glibenclamide or exercise alone (6.7 +/- 1.1 vs. 8.1 +/- 0.9 and 7.6 +/- 1.0 mmol/l, respectively; P < 0.05). During exercise, the rate of appearance of plasma glucose determined by 3 [3H]glucose infusion was lower on days with glibenclamide than on days without glibenclamide (2.3 +/- 0.1 vs. 2.9 +/- 0.1 mg.min-1.kg-1; P < 0.05). In contrast, glucose clearance was identical (P > 0.05). CONCLUSIONS: In postabsorptive type 2 diabetic patients, the hypoglycemic action of glibenclamide and exercise is enhanced when the treatments are combined. The interaction reflects an increased inhibition by glibenclamide-enhanced insulin levels of hepatic glucose production when hepatic glucose production is accelerated by exercise. PMID- 10526730 TI - A quantitative scale of acanthosis nigricans. AB - OBJECTIVE: To develop and validate a scale for acanthosis nigricans (AN). RESEARCH DESIGN AND METHODS: Subjects were participants from the San Antonio Family Diabetes Study and the San Antonio Family Heart Study. A total of 406 subjects were independently examined for AN by at least two observers. Five locations were examined: the neck, axilla, elbows, knuckles, and knees. Interobserver concordance and kappa statistics were calculated to determine replicability of the scale. Comparisons of diabetes-related risk factors by AN score were also calculated. RESULTS: Only the neck had consistently high kappa statistics, and thus, other locations were excluded from further analyses. Elevated AN was strongly associated with elevated fasting insulin and BMI in both diabetic and nondiabetic subjects. Elevated AN was also strongly associated with elevated fasting glucose, systolic blood pressure, and diastolic blood pressure, and with decreased HDL in nondiabetic subjects. In diabetic subjects, elevated AN was associated with elevated total cholesterol. CONCLUSIONS: We have developed a scale for AN that is easy to use, has high interobserver reliability in Mexican Americans, and correlates well with fasting insulin and BMI. This scale will permit longitudinal and cross-sectional evaluation of AN and will permit the evaluation of AN as a trait in genetic studies. PMID- 10526731 TI - Predicting expenditures for Medicare beneficiaries with diabetes. A prospective cohort study from 1994 to 1996. AB - OBJECTIVE: To describe health care expenditures and utilization patterns among older adults with diabetes and to examine factors associated with expenditures over a 3-year period. RESEARCH DESIGN AND METHODS: We conducted a prospective cohort study of health care expenditures and utilization by diabetic patients from a random nationwide sample of aged Medicare beneficiaries from 1994 to 1996. All services covered by the Medicare program were examined. Multivariate regression was used to assess the contribution of patient characteristics in 1994 on Part B, inpatient, and total expenditures in 1995 and 1996. RESULTS: Per capita expenditures for beneficiaries with diabetes (n = 169,613) were 1.7 times greater than those for those beneficiaries without diabetes (n = 968,832) in 1994. This ratio remained fairly constant over the 2 years of follow-up. Expenditures for beneficiaries with diabetes were highly skewed. However, few of these individuals remained in the highest expenditure quintile over the 2 years of follow-up. Using multiple regression analysis to adjust for demographic and clinical characteristics, we were able to explain 7% of the variation in total expenditures in 1995 and 6% of the variation in 1996. Using the same model, we were able to explain 10.7% of the variation in Part B expenditures in 1995 and 8% in 1996. CONCLUSIONS: Beneficiaries with diabetes are consistently more expensive than beneficiaries without diabetes. Demographic and clinical factors at baseline are able to predict only a small portion of future expenditures among this population, and the most expensive patients in one year were often not the most expensive in subsequent years. More work is necessary to assure equitable risk adjustment in the calculation of capitation rates for health plans and practitioners who specialize in the care of individuals with diabetes. PMID- 10526732 TI - Consequences of the new diagnostic criteria for diabetes in older men and women. DECODE Study (Diabetes Epidemiology: Collaborative Analysis of Diagnostic Criteria in Europe). AB - OBJECTIVE: To evaluate the prevalence of diabetes and the risk of death in older European men and women aged between 60 and 79 years at baseline using the new American Diabetes Association diagnostic criteria for diabetes. RESEARCH DESIGN AND METHODS: The analysis involved existing population-based European studies from the DECODE Study Group (Diabetes Epidemiology: Collaborative Analysis of Diagnostic Criteria in Europe) and involved baseline measures of fasting and 2-h glucose concentrations after a 75-g oral glucose tolerance test (OGTT) and follow up to determine deaths. This analysis included 4,032 men and 2,207 women who were not previously known to have diabetes and 383 men and 319 women who had established diabetes. RESULTS: More than half of the diabetic subjects had already been diagnosed, one sixth had a fasting hyperglycemia > or = 7.8 mmol/l (140 mg/dl), one-sixth had a fasting glucose level of 7.0-7.8 mmol/l (126-140 mg/dl), and one-sixth had an isolated postchallenge hyperglycemia (fasting glucose < 7.0 mmol/l and 2-h glucose > or = 11.1 mmol/l [200 g/dl]). Compared with non-diabetic subjects, the hazard ratios for death in diabetic subjects were close to 2 and did not differ significantly according to the method of diagnosis of diabetes, age-group, or sex. CONCLUSIONS: One-third of the older diabetic subjects who were undiagnosed at baseline had isolated postchallenge hyperglycemia. OGTT screening of the subjects with impaired fasting glucose (6.1 6.9 mmol/l) would reduce this fraction by half. The group with isolated postchallenge hyperglycemia had an elevated risk of mortality similar to that of other diabetic subjects. PMID- 10526733 TI - Implementing practice guidelines for diabetes care using problem-based learning. A prospective controlled trial using firm systems. AB - OBJECTIVE: A controlled trial with 15-month follow-up was conducted in two outpatient clinics to study the effects of using the problem-based learning technique to implement a diabetes clinical practice guideline. RESEARCH DESIGN AND METHODS: A total of 144 patients with type 2 diabetes aged 25-65 years in two internal medicine outpatient clinics were enrolled in the study. African Americans and Hispanics made up > 75% of the patients. Doctors and staff in one of the clinics were trained in the use of a clinical practice guideline based on Staged Diabetes Management. A problem-based learning educational program was instituted to reach consensus on a stepped intensification scheme for glycemic control and to determine the standards of care used in the clinic. HbA1c was obtained at baseline and at 9 and 15 months after enrollment. RESULTS: At 9 months, there was a mean -0.90% within-subject change in HbA1c in the intervention group, with no significant changes in the control group. The 15 month mean within-subject change in HbA1c of -0.62% in the intervention group was also significant. Among intervention patients, those with the poorest glycemic control at baseline realized the greatest benefit in improvement of HbA1c. The intervention group also exhibited significant changes in physician adherence with American Diabetes Association standards of care. CONCLUSIONS: Clinical practice guidelines are an effective way of improving the processes and outcomes of care for patients with diabetes. Problem-based learning is a useful strategy to gain physician support for clinical practice guidelines. More intensive interventions are needed to maintain treatment gains. PMID- 10526734 TI - Racial and ethnic differences in health insurance coverage for adults with diabetes. AB - OBJECTIVE: To evaluate the extent and types of health insurance coverage in a representative sample of adults with diabetes in the U.S. RESEARCH DESIGN AND METHODS: The Third National Health and Nutrition Examination Survey included national samples of non-Hispanic whites, non-Hispanic blacks, and Mexican Americans aged > or = 20 years. Information on medical history and treatment of diabetes was obtained to determine subjects who had been diagnosed with diabetes by a physician before the survey (n = 1,503) and subjects without diagnosed diabetes (n = 17,319). Information on health insurance coverage was obtained via a structured questionnaire for 96% of participants. RESULTS: A total of 93% of all adults with diabetes had some form of health insurance. Of these subjects, 73% had private insurance, 48% had Medicare coverage, 15% had Medicaid coverage, and 5% had Champus/Veterans Affairs coverage. Approximately 52% of adults with diabetes had multiple types of health insurance, and 54% had health care coverage through one or more government-sponsored programs. A greater proportion of non Hispanic whites (91%) and non-Hispanic blacks (89%) than Mexican-Americans (66%) had health insurance among subjects aged 20-64 years. For those aged > or = 65 years, coverage was virtually 100% for all racial and ethnic groups. Non-Hispanic whites had the highest rate of coverage through private insurance (81%), with non Hispanic blacks having an intermediate rate (56%) and Mexican-Americans having the lowest rate (45%). Rates of coverage were similar for adults with and without diabetes in each racial and ethnic group for any type of insurance and for private insurance. CONCLUSIONS: There are marked racial and ethnic differences in health insurance coverage for adults with diabetes, although these differences are similar to those for adults without diabetes. Whether these racial and ethnic disparities influence access to care, quality of care, or health outcomes for people with diabetes remains to be determined. PMID- 10526735 TI - High normal blood pressure, hypertension, and the risk of type 2 diabetes in Japanese men. The Osaka Health Survey. AB - OBJECTIVE: To investigate the relationship between high normal blood pressure or hypertension and the risk of developing type 2 diabetes in a large Japanese cohort. RESEARCH DESIGN AND METHODS: We enrolled 7,594 Japanese men aged 35-60 years who did not have diabetes or impaired fasting glucose at study entry. Type 2 diabetes was defined as a fasting plasma glucose level of > or = 126 mg/dl or a 2-h postload plasma glucose level of > or = 200 mg/dl. High normal blood pressure was defined as no history of hypertension and a systolic blood pressure of > or = 130 and < 140 mmHg or a diastolic blood pressure of > or = 85 and < 90 mmHg. Subjects were considered to have hypertension if they had a systolic blood pressure > or = 140 mmHg, if they had a diastolic blood pressure > or = 90 mmHg, or if they were taking anti-hypertensive medications. RESULTS: We confirmed 600 cases of type 2 diabetes during the 72,946 person-years of follow-up. Both high normal blood pressure and hypertension were associated with the risk of type 2 diabetes. Compared with normotensive men, men with high normal blood pressure had a multiple adjusted relative risk (RR) of 1.39 (95% CI 1.14-1.69), and men with hypertension had a multiple adjusted RR of 1.76 (1.43-2.16). Even among lean men (BMI < 22.7 kg/m2), men with high normal blood pressure had a multiple adjusted RR of 1.71 (1.20-2.42), and men with hypertension had a multiple adjusted RR of 2.02 (1.34-3.05) compared with normotensive men. CONCLUSIONS: High normal blood pressure and hypertension are associated with an increased risk of developing type 2 diabetes. PMID- 10526736 TI - LDL particle size in relation to insulin, proinsulin, and insulin sensitivity. The Insulin Resistance Atherosclerosis Study. AB - OBJECTIVE: LDL particles are heterogeneous in terms of size and density; small dense LDL particles are considered more atherogenic than larger LDL particles. The aim of this study was to investigate the interrelationships among LDL size, insulin, proinsulin (intact and split), and insulin sensitivity in a tri-ethnic population with varying degrees of glucose tolerance (n = 1,549) in the Insulin Resistance Atherosclerosis Study. RESEARCH DESIGN AND METHODS: Insulin sensitivity was assessed by a frequently sampled intravenous glucose tolerance test with minimal model analysis. Proinsulin levels were measured using highly sensitive assays without detectable cross-reactivity with insulin, and LDL size was determined by gradient-gel electrophoresis. RESULTS: In univariate analyses, LDL size was related to various features of the insulin resistance syndrome, including fasting insulin (r = -0.18), intact proinsulin (r = -0.24), split proinsulin (r = -0.24), the proinsulin-to-insulin ratio (r = -0.14), and insulin sensitivity (r = 0.21; all P < 0.0001). In a multivariate regression model (adjusted for age, BMI, ethnicity, and clinic), triglyceride levels (P = 0.0001), HDL cholesterol (P = 0.0001), sex (P = 0.002), and proinsulin (P = 0.01) were significantly related to LDL size. In the same model stratified by sex, LDL size was significantly inversely related to proinsulin in men (P = 0.005 and P = 0.04 after further adjustment for the glucose tolerance status), but not in women (P > 0.15). CONCLUSIONS: We found an inverse relation of proinsulin to LDL particle size in a large tri-ethnic population with varying degrees of glucose tolerance. This relation was independent of age, BMI, and triglyceride and HDL cholesterol concentrations, and was more pronounced in men than in women. PMID- 10526737 TI - Lack of association between early childhood immunizations and beta-cell autoimmunity. AB - OBJECTIVE: To determine whether early childhood immunization history affects the risk of developing the beta-cell autoimmunity that precedes type 1 diabetes. RESEARCH DESIGN AND METHODS: This article describes a case-control study whose participants were 317 children aged < or = 12 years who have a first-degree relative with type 1 diabetes. The children were enrolled in a prospective cohort study of the etiology of beta-cell autoimmunity, the Diabetes Autoimmunity Study in the Young, in Denver, Colorado. The main outcome measure was beta-cell autoimmunity as determined by persistent autoantibodies against insulin, GAD, or islet cell antibody (IA-2) 512. The number of cases with beta-cell autoimmunity was 25, and the number of control subjects (the remainder of the cohort) was 292. RESULTS: There was no difference between cases and control subjects in the proportion receiving hepatitis B (HBV), Haemophilus influenzae b (Hib), polio, or diphtheria tetanus pertussis (DTP) vaccines before 9 months of age; in the proportion receiving HBV at birth rather than later; or in the median age at first HBV, Hib, polio, or DTP vaccination. CONCLUSIONS: The results suggest that changing the early childhood immunization schedule would not affect the risk of developing beta-cell autoimmunity or type 1 diabetes. PMID- 10526738 TI - Perinatal risk factors for childhood type 1 diabetes in Europe. The EURODIAB Substudy 2 Study Group. AB - OBJECTIVE: To explore whether perinatal factors are associated with the development of childhood type 1 diabetes. RESEARCH DESIGN AND METHODS: We studied hospital records from 892 cases of childhood type 1 diabetes compared with 2,291 population-based control subjects in seven study centers in Europe. RESULTS: In a pooled analysis incorporating stratification by center, we confirmed the previous findings that older maternal age, maternal preeclampsia, neonatal respiratory disease, and jaundice caused by blood group incompatibility are significant risk factors for type 1 diabetes, whereas being a firstborn child, having a low birth weight, or having a short birth length were protective. Cesarean section delivery and neonatal infectious diseases were not significantly associated with the risk of type 1 diabetes in this study. The strongest association was found for blood group incompatibility (AB0 and Rh factor) with an odds ratio (OR) of 2.96 (95% CI 1.88-4.65). AB0 incompatibility (OR = 3.92) was a more common and also a stronger risk factor than Rh incompatibility (OR = 1.62). The effect of AB0 blood group incompatibility was independent of treatment effects in logistical regression analysis. CONCLUSIONS: Different perinatal events are associated with an increased risk of type 1 diabetes. The effect of maternal-child blood group incompatibility is strong and indicates a true effect that must be further explored. PMID- 10526739 TI - Effect of Bacillus Calmette-Guerin vaccination on new-onset type 1 diabetes. A randomized clinical study. AB - OBJECTIVE: We undertook this study to test whether Bacillus Calmette-Guerin (BCG) vaccine preserves beta-cell function and increases the remission rate in children with new-onset type 1 diabetes. RESEARCH DESIGN AND METHODS: This was a randomized double-blind placebo-controlled trial offered to children referred to the Barbara Davis Center for Childhood Diabetes or the Baystate Medical Center with a diagnosis of new-onset type 1 diabetes. There were 94 children aged 5-18 years who received either BCG or saline intradermally within 4 months of onset of symptoms and who were then evaluated at 3-month intervals for 2 years. The primary end point was remission, defined as insulin independence for 4 weeks. Secondary end points were C-peptide levels (fasting and in response to a mixed meal challenge), insulin dose, and HbA1c. RESULTS: Of the patients, 47 were randomized to each arm; 7 in the placebo group and 9 in the BCG group did not complete 1 year of the study and are not included in the analysis. One patient from each group achieved remission. Fasting and stimulated C-peptide levels did not differ by treatment arm but declined in both groups and were lower initially and during the entire 2-year period in younger children. Insulin requirements and HbA1c levels did not differ in the two groups. CONCLUSIONS: Vaccination with BCG at the time of onset of type 1 diabetes does not increase the remission rate or preserve beta-cell function. PMID- 10526740 TI - Correlation of fingerstick blood glucose measurements with GlucoWatch biographer glucose results in young subjects with type 1 diabetes. AB - OBJECTIVE: The purpose of this study was to compare measurements of glucose obtained via iontophoretic extraction with the GlucoWatch automatic glucose biographer (Cygnus, Inc., Redwood City, CA) with capillary blood glucose values that were determined 1) in a controlled outpatient clinic setting and 2) in a home setting. RESEARCH DESIGN AND METHODS: There were 76 GlucoWatch biographers used on 28 different young adults (21 women and 7 men) with type 1 diabetes (age 30.9 +/- 6.9 years and duration of diabetes 18.4 +/- 8.1 years [mean +/- SD]) in a controlled outpatient clinic setting. Some subjects participated on multiple days. Subjects wore two GlucoWatch biographers, each on the forearm (ventral aspect). Comparisons were made to HemoCue blood glucose analyzer (Aktiebolgat Leo, Helsingborg, Sweden) capillary blood glucose measurements. In addition, GlucoWatch biographers (one each day for 3 consecutive days) were used by 12 subjects (8 women, 4 men) in a home setting. Comparisons were made to capillary blood glucose values determined using the One Touch Profile meter (Johnson & Johnson, New Brunswick, NJ). RESULTS: GlucoWatch biographer glucose values correlated well with capillary blood glucose values determined using the HemoCue analyzer in the clinic setting (r = 0.90, 1,554 paired data points) and using the One Touch Profile meter in the home setting (r = 0.85, 204 paired data points). When 36 subjects wore two biographers simultaneously, the correlation between the two biographers was r = 0.94. The error grid analysis demonstrated that > 96% of biographer glucose values determined in the clinic or home setting were in the clinically acceptable A and B regions. CONCLUSIONS: This study confirms the accuracy and precision of glucose values as determined using the GlucoWatch biographer in clinic and home settings. PMID- 10526741 TI - Use of the Semmes-Weinstein monofilament in the strong heart study. Risk factors for clinical neuropathy. AB - OBJECTIVE: We used the Semmes-Weinstein 5.07 monofilament to assess the prevalence of foot insensitivity and its relationship to potential risk factors. RESEARCH DESIGN AND METHODS: There were 3,638 American Indian participants from Arizona, North and South Dakota, and Oklahoma who attended a study clinic on two occasions: baseline and follow-up, 4 years later. Oral glucose tolerance tests were performed at the visits for those who had not previously been diagnosed as having diabetes. A total of 2,051 participants were diagnosed with diabetes before the study or at the subsequent study visits. At the follow-up visit, participants were tested for their ability to sense the 5.07 (10 g) monofilament at 10 sites of the foot. The prevalence of foot insensitivity was ascertained, and its relation to characteristics of participants was assessed in both univariate and logistic regression analyses. RESULTS: Diabetic participants had a much higher prevalence of foot insensitivity (defined as greater than or equal to five incorrect responses) than nondiabetic participants (14 vs. 5%, respectively). However, marked foot insensitivity was uncommon within the first few years of diagnosis of diabetes. Among the diabetic participants, those diagnosed before study entry had the highest prevalence of foot insensitivity. The prevalence of foot insensitivity was highest in the Arizona Indians (22 vs. 9% in the Dakotas and 8% in Oklahoma). In a logistic regression analysis, foot insensitivity was significantly and independently related to center (Arizona versus others), age, duration of diabetes, and height. CONCLUSIONS: Marked foot insensitivity is prevalent in the diabetic American Indian population, especially in Indians in Arizona; however, this insensitivity is apparently uncommon for several years after the diagnosis of diabetes. The data show that Indians with diabetes are particularly vulnerable to the risk of foot ulceration and that the diagnostic screening of diabetes may lead to better prevention of sensory neuropathy and subsequent foot ulceration. PMID- 10526742 TI - Augmentation of central arterial pressure in type 1 diabetes. AB - OBJECTIVE: Atherosclerosis is more severe in individuals with diabetes. Whether diabetic subjects have accelerated arterial hardening (i.e., arteriosclerosis) is less clear. Arteriosclerosis increases pulse-wave velocity and can augment central arterial pressure due to early wave reflection. The aim of this study was to determine whether subjects with type 1 diabetes had evidence of increased arterial stiffness by using pulse-wave analysis. RESEARCH DESIGN AND METHODS: Radial artery pressure waveforms were obtained noninvasively by applanation tonometry (PWV Medical Blood Pressure Analysis System, Sydney). A central aortic waveform can be derived by using a transfer function used in previous studies during cardiac catheterization. A total of 89 subjects with type 1 diabetes (46 men and 43 women, aged 34.0 +/- 11.0 years, duration of diabetes 13.1 years [interquartile range 5.8-24.3], HbA1c 8.2 +/- 1.7%) and 95 control subjects (44 men and 51 women, aged 36.1 +/- 12.0 years) were studied. The central aortic waveform allowed the determination of 1) the aortic augmentation index (AAI), a parameter that reflects the degree to which central arterial pressure is augmented by wave reflection, and 2) the subendocardial viability ratio (SEVR), which is a measure of myocardial perfusion relative to cardiac workload. RESULTS: In multivariate analysis, diabetes was an important determinant of AAI (P = 0.001). The higher AAI was mainly evident in the men, for whom diabetes was a highly significant covariate (P = 0.006); this was not the case for diabetic women (P = 0.2). Nondiabetic men had a lower AAI than nondiabetic women (103.7 +/ 18.6 vs. 117.0 +/- 22.3%, respectively, P = 0.002), but this difference was abolished by diabetes (110.7 +/- 18.5 vs. 116.1 +/- 18.7%, respectively, P = 0.2). Subjects with type 1 diabetes had a significantly lower mean SEVR compared with control subjects (139.2 +/- 28.3 vs. 163.6 +/- 27.4%, respectively, P < 0.0001). In multivariate analysis, diabetes was an important determinant of SEVR (P = 0.001). A significant interaction between diabetes and age was evident (P = 0.0001), which suggests that the effect of age is modified by diabetes. CONCLUSIONS: These findings suggest that central systolic blood pressure is increased in relatively young individuals with type 1 diabetes, although myocardial perfusion related to cardiac workload is decreased. These changes can be explained by more rapid pulse-wave velocity resulting from arterial stiffening. PMID- 10526744 TI - Exploring and expanding the research agenda for diabetes in managed care. A report of a Centers for Disease Control and Prevention-Managed Care Workshop. AB - The objective of this article is to describe the proceedings of a workshop organized by the Centers for Disease Control and Prevention (CDC) to 1) summarize current preventive care practices for people with diabetes in managed care organizations (MCOs) in the U.S., 2) understand the strengths and barriers to diabetes-related health services research in MCOs, and 3) highlight the major research questions and approaches to improve diabetes-related research in these organizations. Review and synthesis of presentations and discussions are provided. MCOs have considerable potential to enhance diabetes health services research because of their access to large diverse populations, novel health service interventions, and availability of data systems that link patients, health care providers, health care use, and health outcomes. Barriers to improved MCO-based research include confounding by concurrent interventions and secular trends, lack of control groups for rigorous evaluation of interventions, and the heterogeneous structures of MCOs. Future research, particularly if conducted in a collaborative environment, has the potential to improve the understanding of trends in health services, improve the understanding of characteristics of MCOs, and evaluate complex interventions directed at patients, health care providers, and systems of care. PMID- 10526743 TI - Relationship between urinary albumin excretion, body composition, and hyperinsulinemia in normotensive glucose-tolerant adults. AB - OBJECTIVE: Elevated urinary albumin excretion (UAE) has been associated with insulin resistance and is suggested to be elevated in prediabetic individuals. Upper body obesity, especially visceral obesity, predicts insulin resistance and development of type 2 diabetes. We examined whether UAE clusters with obesity associated insulin resistance traits in healthy glucose-tolerant normotensive subjects. RESEARCH DESIGN AND METHODS: There were 49 volunteers with a wide range of body fat and body fat distribution studied. All had normal blood pressure and glucose tolerance and were maintained on a controlled diet for 2 weeks. UAE was assessed from three overnight urine collections, and body composition was assessed by whole-body dual-energy X-ray absorptiometry scanning and abdominal computed tomography scanning. RESULTS: Fasting insulin and insulin responses to oral glucose were significantly increased in obese subjects, who also tended to have more dyslipidemia, greater blood pressure, and more visceral fat than lean subjects. These differences were more apparent in upper body obese subjects. UAE was normal in obese and upper body obese subjects and not different from that of lean subjects. UAE ranged from 0.3 to 8.3 micrograms/min in lean subjects and from 0.2 to 7.2 micrograms/min in obese subjects. UAE was not significantly correlated with body composition, plasma insulin, glucose, or lipids. CONCLUSIONS: Obese subjects (even upper body obese subjects) with increased visceral and total body fat, high plasma insulin and triglycerides, and low HDL cholesterol concentrations do not have elevated UAE. This suggests that UAE is not closely associated with these characteristics and implies a later onset of abnormal albuminuria in the course of the insulin resistance syndrome. PMID- 10526745 TI - Cardiovascular disease in type 2 diabetes. PMID- 10526746 TI - Association of G82S polymorphism in the RAGE gene with skin complications in type 2 diabetes. PMID- 10526747 TI - Asymptomatic coronary artery disease is associated with cardiac autonomic neuropathy and diabetic nephropathy in type 2 diabetic patients. PMID- 10526748 TI - Are women with gestational diabetes more likely to have been bottle-fed? PMID- 10526749 TI - No effect of gluten-free diet on the metabolic control of type 1 diabetes in patients with diabetes and celiac disease. Retrospective and controlled prospective survey. PMID- 10526750 TI - Relationship of tumor necrosis factor-alpha plasma levels to metabolic control in type 1 diabetes. PMID- 10526751 TI - Nitrate in drinking water and risk of childhood diabetes in The Netherlands. PMID- 10526752 TI - Insulin lispro and retinopathy in pregnancy. PMID- 10526753 TI - Rapid increase in the prevalence of undiagnosed diabetes and impaired fasting glucose in asymptomatic Hong Kong Chinese. PMID- 10526754 TI - Effect of troglitazone on lipoprotein(a) levels in obese subjects. PMID- 10526755 TI - Tretinoin treatment of necrobiosis lipoidica diabeticorum. PMID- 10526756 TI - Effect of exercise training on doses of oral agents and insulin. PMID- 10526757 TI - Further data on the comparison between World Health Organization and American Diabetes Association diagnostic criteria DIAINF Study Group. PMID- 10526758 TI - Rethinking the diabetes prevention program clinical trial. PMID- 10526759 TI - The diabetes prevention program: evaluation and management of diabetes. Response to Adler and Turner and Singer et al The DPP Research Group. PMID- 10526760 TI - Cellular immune response to GAD in type 1 diabetes with residual beta-cell function. PMID- 10526761 TI - Anti-GAD65 antibody titer may be important in assessing T-cell response in anti GAD65+ diabetes with residual beta-cell function PMID- 10526762 TI - On calculating treatment satisfaction. PMID- 10526763 TI - Vertically transmitted enteroviruses and the benefits of neonatal immunization. PMID- 10526764 TI - Engaging multiproblem families in treatment: lessons learned throughout the development of multisystemic therapy. AB - Multisystemic therapy (MST) is a family-based treatment model that has achieved high rates of treatment completion with youths who present serious clinical problems, and their families. The success of MST in engaging challenging families in treatment is due to programmatic commitments to family collaboration and partnership as well as to a conceptual process that delineates barriers to family engagement, develops and implements strategies to overcome these barriers, and evaluates the success of these strategies. This article provides an overview of the nonspecific/universal engagement strategies used by MST therapists, frequently observed barriers to achieving therapist-family engagement, and specific strategies to overcome a sampling of these barriers. PMID- 10526765 TI - Challenges to family engagement: what can multisystemic therapy teach family therapists? PMID- 10526766 TI - Rebound from marital conflict and divorce prediction. AB - Marital interaction has primarily been examined in the context of conflict resolution. This study investigated the predictive ability of couples to rebound from marital conflict in a subsequent positive conversation. Results showed that there was a great deal of consistency in affect across both conversations. Also examined was the ability of affective interaction to predict divorce over a 4 year period, separately in each of the two conversations. It was possible to predict divorce using affective variables from each conversation, with 82.6% accuracy from the conflict conversation and with 92.7% accuracy from the positive rebound conversation. PMID- 10526767 TI - Predicting divorce among newlyweds from the first three minutes of a marital conflict discussion. AB - This study tested the hypothesis that how a discussion of a marital conflict begins--in its first few minutes--is a predictor of divorce. The marital conflict discussion of 124 newlywed couples was coded using the Specific Affect Coding System, and the data were divided into positive, negative, and positive-minus negative affect totals for five 3-minute intervals. It was possible to predict marital outcome over a 6-year period using just the first 3 minutes of data for both husbands and wives. For husbands this prediction improved as the groups diverged in the remaining 12 minutes; for wives the prediction remained equally powerful for the remaining 12 minutes as it had been in the first 3 minutes. PMID- 10526768 TI - Negotiating couplehood: the process of resolving the December dilemma among interfaith couples. AB - Christmas forces interfaith couples to address questions concerning holiday observances. The purpose of this investigation was to explore the experience of the "December dilemma," that is, the experience of Christmas and Hanukah among couples in which one partner is Jewish. A qualitative design based on the continuous comparison method of Grounded Theory analysis was used. Participants were solicited through interfaith couples' programs, referral, and snowballing. Unstructured interactive interviews of 22 couples were audiotaped, transcribed, and analyzed. The categories generated were: Ghosts of Christmas and Hanukah Past, Coming Together, and Holiday Observances as a Couple. The basic problem facing these couples was how to bridge religious backgrounds with differing holiday traditions in a way that integrated respect for each partner's needs, heritage, and identity. The basic social process of negotiating "couplehood," that is, moving from individuality to partnership emerged when mutual agreement could be reached to solve problems about how to celebrate the December holidays. The data indicated that exploration of the ways these couples managed the dilemmas created by the December holidays provided a window to how they negotiated other challenges in their relationships. PMID- 10526769 TI - Beyond different worlds: a "postgender" approach to relational development. AB - Approaches to gender in therapy either reinforce or challenge existing gender differences and inequalities. The authors suggest a way to help clients move beyond gender constructions from the past. They argue that perceived gender differences are rooted in power differences that limit relational development for both women and men, and perpetuate unequal relationship structures. As an alternative to the "two different worlds," gender-as-culture framework, they present an approach to therapy based on an expanded version of Bowen's notion of differentiation. The article helps therapists recognize four "gender traps" that interfere with relational development and suggests strategies for helping clients differentiate from old gender patterns. PMID- 10526770 TI - Relationships among parental reports of child, parent, and family functioning. AB - Most children with psychosocial problems do not present for treatment in mental health settings. They are managed by primary care physicians. Children with psychosocial problems often have parents and/or families with psychosocial distress. The present study measured associations between parental reports of child, parent, and family functioning in individuals in the general population. Participants were 226 parents of children, aged 2-16 years, who presented for routine primary care. Parents reported on the psychosocial functioning of themselves, their child, and their family. All correlations of measures were significant, ranging from .55 to .23. Similar to data from psychiatric samples, the psychological functioning of children, parents, and families were significantly correlated. Unlike in psychiatric settings, child mental health problems were not as closely related to parent or family distress as parent and family distress were related to each other and to child behavior problems. PMID- 10526771 TI - Disruption and reconstruction: narrative insights into the experience of family members caring for a relative diagnosed with serious mental illness. AB - The findings of a study investigating carers' accounts about serious mental illness occurring in their family are presented. The narrative form is a primary means of ordering, structuring, and communicating illness experiences, reflecting some of the processes that carers intend to master and understand. Psychotic episodes entail a frightening disruption that forces carers to face fundamental existential, moral, and psychological issues because they call into question the continuity of lives and life-projects. This study has explored how carers articulate the consequences of a devastating experience and turn it into a meaningful event that can in some way be incorporated into the course of their life. Two types of narrative structure were identified. In stories of restitution or reparation, the experience of the event is transformed into phenomena having meaning, occupying a place in carers' lives. In chaotic and frozen narratives, the illness remains a series of random events. The effects on coping of these two narrative types were explored, as well as gender-related themes and beliefs about mastery and control. Therapeutic implications are discussed and also possible connections to other research constructs (for example, Expressed Emotion). It is argued that the concept of illness must be approached from a systemic, multidetermined perspective that includes our narrative constructions. PMID- 10526772 TI - Physical "phantasies" and family functions: overcoming the mind/body dualism in somatization. AB - In this article, we examine some of the ways in which family therapists have conceptualized the experience of illness of unexplained physical origin. We argue that opinions about the etiology of somatic symptoms should not be the primary focus of therapeutic work with people who share the prototypical characteristics of what has been defined as "somatization disorder." We suggest that current research in neurobiology can expand the linguistic resources of clinicians and help them avoid perpetuating unhelpful dichotomies between the mind and the body. PMID- 10526773 TI - Effects of epoxycarotenoids, beta-carotene, and retinoic acid on the differentiation and viability of the leukemia cell line NB4 in vitro. AB - Three all-trans epoxides of beta-carotene (beta-Car), namely, 5,6-epoxy-beta carotene (5,6-EC), 5,8-epoxy-beta-carotene (5,8-EC) and 5,6,5',6'-diepoxy-beta carotene (5,6,5',6'-DEC) were synthesized by treatment of beta-carotene with 3 chloroperoxybenzoic acid, were purified chromatographically, and were characterized. The relative potencies (mean +/- S.D.) of 1 microM compounds in inducing the differentiation of NB4 cells, a cell line that contains the chromosomal transposition t(15;17) characteristic of acute promyelocytic leukemia, after 4 days of incubation were: RA: 1.35 +/- 0.16, 5,6-EC: 0.29 +/- 0.01, 5,8-EC: 0.22 +/- 0.05, 5,6,5',6'-DEC: 0.11 +/- 0.02, beta C: 0.09 +/- 0.01, and the control: 0.06 +/- 0.01. The same order of potencies existed at other concentrations tested and at other incubation times. P values for the differences between the inducing activities of successive pairs of compounds at 1 microM were: RA vs. 5,6-EC, < 0.001; 5,6-EC vs. 5,8-EC, < 0.01; 5,8-EC vs. 5,6,5',6' DEC, < 0.01; 5,6,5',6'-DEC vs. beta-Car, < 0.10; beta-Car vs. control, < 0.005. Similar P values were also obtained for studies at other concentrations and at other incubation times. The viable cell mass at 4 days was inversely proportional to the extent of differentiation (rs = -1.0). The inducing activities of all compounds were dose-dependent. Thus, the 5,6-monoepoxide of beta-carotene, which has not previously been studied as an inducer, showed higher activity in NB4 cell differentiation than the 5,8-monoepoxide, the 5,6,5',6'-diepoxide, or beta carotene. Possible explanations of these observations are discussed. PMID- 10526774 TI - Determination of retinol, antioxidant vitamins and homocysteine in skin puncture blood. AB - For determination of the vitamin status via mass screening, simple and rapid methods are required. Additionally, blood samples should be obtained using simple and low invasive sampling techniques. To fulfill this existing methods have been modified to analyze retinol, tocopherols, beta-carotene, vitamin C and homocysteine in 20 microliters plasma. Blood samples were obtained via skin punctures. HPLC measurements were carried out with isocratic separation and precolumn derivatization. Intra and interday variation coefficients were below 8% and regression coefficients better than 0.99 for all measurements. The difference between venous and capillary samples were < 5%. In conclusion, the methods employed proved satisfactory for the determination of important nutritional parameters in blood samples obtained via skin punctures. These methods are therefore well suited for mass screening, especially under field conditions in developing countries. PMID- 10526775 TI - Influence of dietary n-3 polyunsaturated fatty acids on plasma lipemic effect of vitamin B6 deficiency. AB - Since many connections exist between vitamin B6 and lipid metabolism, we aim to investigate the lipemic effect of different dietary intakes of polyunsaturated fatty acids in rats fed a vitamin B6 deficient diet. Diets were either vitamin B6 deficient (-B6) or vitamin B6 sufficient, pair-fed to the deficient group (PF) and ad libitum (N). The diets were combined with normal lipid (LC: soya bean coconut-palm oils) and fish oil (FO: soya bean-fish oil). The fish oil diet with sufficient vitamin B6 content caused an increase in n-3 long chain polyunsaturated fatty acids and a decrease in arachidonic acid. In the -B6 group fed a normal lipid diet, the arachidonic acid percentage decreased and the linoleic acid percentage increased; in the -B6 group fed fish oil these changes in fatty acid composition, already consequent upon dietary intake of n-3 long chain polyunsaturated fatty acids, did not show further variations. In the dietary condition of vitamin B6 deficiency, plasma cholesterol content increased in rats fed a lipid control diet, whereas no hypocholesterolemic effect was observed in those fed a fish oil diet. Plasma triglyceride contents were not influenced by dietary lipid quality because, in all conditions, the lower food intake of the PF groups caused a decrease and vitamin B6 deficiency caused an elevation in triglyceride contents which reached those of the ad libitum groups. The study highlights the interaction between vitamin B6 and polyunsaturated fatty acids and the opportunity of dietary intake of fish oil to counterbalance some effects of vitamin B6 deficiency. PMID- 10526776 TI - Epidemiologic correlates of serum folate and homocysteine levels among users and non-users of vitamin supplement. AB - Lower serum folate and higher serum homocysteine levels are known risk factors for various conditions. Thus, epidemiologic correlates with these measurements were studied for 256 multivitamin users and 230 non-users who were middle-aged women. Both serum folate and homocysteine levels increased with advancing age in both multivitamin users (P < 0.01 and P < 0.01) and non-users (P = 0.08 and P < 0.01). Among non-users, higher intake of vegetables, fruits, cold cereals and total protein were associated positively with serum folate and inversely with homocysteine levels. There were 25-74% increases in serum folate and 10-15% decreases in serum homocysteine between 1st and 4th quartiles of intake of these food/nutrients. In addition, 26% lower serum folate and 18% higher serum homocysteine were observed for those smoking 20 or more cigarettes per day compared with non-smokers. Among multivitamin users, body weight was correlated inversely with serum folate (P < 0.01) and positively with serum homocysteine levels (P = 0.04), while no correlates were found among lifestyle factors. Regular use of multivitamins increased serum folate about fourfold and decreased homocysteine twofold. These results suggest that multivitamin use can offset the effects of an unhealthy lifestyle on these serum markers, and that levels of serum folate and homocysteine can also be favorably influenced by healthier diet and abstinence from smoking. PMID- 10526777 TI - Effects of dietary crude palm oil, fish oil and their association on cholesterol and lipoprotein constants in rats which could be beneficial in humans. AB - The aim was first to examine the differential effects of crude and refined palm oil (CPO and RPO) on the lipid and lipoprotein constants of plasma in rats and to compare the effect of crude palm oil to that of fish oil. Secondarily, it was to know whether one can take advantage from the association of CPO with FO. Twenty four-day-old weaning rats were divided into five experimental groups, each receiving a purified diet containing 10% oil as either a single oil or an equal amount of two oils. After a feeding period of 36 days, the main results were as follows. As compared to the rats fed the RPO diet, those fed the CPO diet had lower total cholesterol, LDL-C, VLDL-C and apoB and higher HDL-C/LDL-C and apoA1/apoB ratios. Those fed the FO diet had only lower VLDL-C and triglycerides and higher HDL-C and HDL-C/LDL-C ratio. Whereas FO associated with RPO in the same diet had the same effect as FO alone, FO associated with CPO tends to reinforce the effect of CPO. This is particularly true for the effects on apoB and apoA1 which were found to be synergistically depressed and enhanced, respectively. Given the role played by these biological constants as predictors of CVD in humans, and in spite of the fact that these predictors are not relevant in rats, these results would suggest the potential interest of CPO or the association of CPO with FO in human nutrition. PMID- 10526778 TI - Effects of capsaicin on serum triglycerides and free fatty acid in olive oil treated rats. AB - Male rats were dosed with capsaicin after the administration of olive oil, and the serum was obtained for analysis of triglycerides and free fatty acids (FFAs). The serum triglycerides level was increased at 2 and 4 hours after olive oil treatment. On the other hand, capsaicin significantly lowered this increase after 4 hours of treatment in the dosage of 100 mg/kg. Total FFA level was also lowered which had previously been increased by the administration of olive oil after 2 hours of treatment, solely the increase in oleic acid (C18:1) levels was lowered among the FFA. The present results indicated that capsaicin decreased only the components of the dosed olive oil. Furthermore, elevation of the serum total FFA concentration was significantly inhibited 2 or 4 hours after the treatment but not at 8 hours, suggesting that a single high dose treatment with capsaicin may inhibit the absorption of lipid in the gastrointestinal tract. PMID- 10526779 TI - Hypocholesterolemic effect of naringin associated with hepatic cholesterol regulating enzyme changes in rats. AB - The effects of the citrus bioflavonoid naringin were tested by using it as a supplement in a high-cholesterol diet. Male rats were fed for 42 days with a 1% (wt/wt) high cholesterol diet either with or without naringin-supplementation (0.1%, wt/wt) to study the effect on plasma lipid levels, hepatic lipid contents, hepatic enzyme activity, and the excretion of fecal neutral sterols. Naringin did not significantly alter the levels of plasma triglycerides, however, the levels of plasma cholesterol (3.80 +/- 0.31 mmol/L vs. 2.61 +/- 0.30 mmol/L, mean +/- SE; p < 0.05) and hepatic cholesterol (70.3 +/- 4.3 mg/g vs. 54.3 +/- 3.8 mg/g, mean +/- SD; p < 0.05) were significantly lowered compared to those of the control. HMG-CoA reductase (2487.0 +/- 210.0 pmole/min/mg vs. 1879.0 +/- 236.0 pmole/min/mg, mean +/- SE; p < 0.05) and ACAT (806.0 +/- 105.0 pmole/min/mg vs. 643.0 +/- 80.0 pmole/min/mg, mean +/- SE; p < 0.05) activities were both substantially lower in the naringin-supplemented group than in the control. The naringin supplementation markedly decreased the excretion of fecal neutral sterols (204.7 +/- 28.5 mg/day) compared to the control (521.9 +/- 53.9 mg/day). The combination of the inhibited HMG-CoA reductase (-24.4%) and ACAT (-20.2%) activities as a result of naringin supplementation could account for the decrease of fecal neutral sterols. PMID- 10526780 TI - Dietary heme iron does not prevent postgastrectomy anemia but fructooligosaccharides improve bioavailability of heme iron in rats. AB - Gastrectomized rats exhibit iron deficiency anemia. We observed the effects of dietary heme-iron and short chain frucooligosaccharides (Sc-FOS) in relation to prevention of postgastrectomy anemia in rats. Twelve laparotomized (sham operated) rats were fed iron-citrate (control) as iron source diet without or with Sc-FOS (75 g/kg of diet) and twenty four totally gastrectomized (Bilroth II) rats, were fed a iron-citrate (control) or heme-iron (heme) as iron source diet without or with Sc-FOS (75 g/kg of diet) for 4 weeks. All rats received an intramuscular injection of vitamin B-12 every two weeks. Tail blood was collected every other week for determination of hematocrit and hemoglobin concentration. At the end of the experiment, the rats were killed and whole blood was collected. The total gastrectomy induced the postgastrectomy anemia. Dietary Sc-FOS increase iron absorption and thereby prevented completely this anemia in gastrectomized rats fed the control diet but this effect of Sc-FOS in rats fed heme diet was not complete. Dietary heme iron could not prevent postgastrectomy anemia itself, but fructooligosaccharides improve bioavailability of not only non-heme iron such as iron-citrate, but also heme-iron in rats. PMID- 10526781 TI - Flow-cytometric investigation of cellular metabolism during oxidative stress and the effect of tocopherol. AB - Many studies and scientific publications report on potentially beneficial effects of the lipophilic anti-oxidant vitamin E on cellular metabolic pathways. The present work presents data on the influence of tocopherol on different intracellular parameters of intact and living human skin fibroblasts by flow cytometric measurements. The parameters analysed were the intracellular pH, representing cell metabolism and cell function, intracellular glutathione, representing one of the cell's own radical scavenger enzyme systems, membrane potential and cell viability. In order to cause large numbers of free radicals cells were UVB-irradiated prior to measurement. The results of the flow cytometric measurements indicate that vitamin E has significant protecting effects on the measured biochemical parameters during oxidative stress. In the presence of the lipophilic radical scavenger a significant stabilizing effect on pH, intracellular glutathione levels and membrane potential could be observed. Furthermore, vitamin E administration was associated with increased cell viability after UVB irradiation. PMID- 10526782 TI - A microbiological assay on microtitre plates of thiamine in biological fluids and foods. AB - A microbiological method for the determination of thiamine in biological fluids and food using 96-well microtitre plates and automatic plate reader, suitable for using in routine clinical diagnosis is described. Thiamine was extracted from samples by acid digestion (acetate buffer, pH 4.5) at 110 degrees C for 20 minutes. Assay recovery and reproducibility were optimally evaluated. Results obtained from this method were compared with those obtained using traditional microbiological and HPLC methods. The values resulting from this method were more or less the same as those of the traditional microbiological method, but much higher than those of HPLC assay. However, the new assay presents many advantages: it reduces the use of serum volume, which is a main benefit for clinical analysis. It lowers also the reagent costs and increases the number of analyses as well as it is easy to perform for routine clinical laboratory. PMID- 10526783 TI - Solid-phase extraction: method development, sorbents, and coupling with liquid chromatography. AB - The objective of this review is to provide updated information about the most important features of the new solid-phase extraction (SPE) materials, their interaction mode and their potential for modern SPE. First, the recent developments are given in formats, phases, automation, high throughput purpose and set-up of new types of procedures. Emphasis is then placed on the large choice of sorbents for trapping analytes over a wide range of polarities, such as highly cross-linked copolymers, functionalized copolymers, graphitized carbons or some specific n-alkylsilicas. The method development is given which is based on prediction from liquid chromatographic retention data or solvation parameters in order to determine the main parameters of any sequence (type and amount of sorbent, sample volume which can be applied without loss of recovery, composition and volume of the clean-up solution, composition and volume of the desorption solution). Obtaining extracts free from matrix interferences in a few steps--one step when possible--is now included in the development of SPE procedure. New selective phases such as mixed-mode and restricted access matrix sorbents or emerging phases such as immunosorbents or molecularly imprinted polymers are reviewed. Selectivity obtained by combining two sorbents is described with the use of ion-exchange or ion-pair sorbents. Special attention is given to complete automation of the SPE sequence with its on-line coupling with liquid chromatography followed by various detection modes. This represents a fast, modern and reliable approach to trace analysis. Many examples illustrate the various features of modern SPE which are discussed in this review. They have been selected in both biological and environmental areas. PMID- 10526784 TI - Membrane-based sample preparation coupled on-line to chromatography or electrophoresis. AB - A review on the use of membranes for on-line sample preparation prior to chromatographic and electrophoretic analysis is provided. The current state-of the-art of four membrane-based techniques (dialysis, electrodialysis, filtration and membrane extraction) is described by reviewing their principles and applications. Possible future developments are discussed. PMID- 10526785 TI - Supercritical fluids in separation science--the dreams, the reality and the future. AB - The last 20 years have seen an intense interest in the use of supercritical fluids in separation science. This started with the introduction of commercial instruments first for packed and then for capillary chromatography and it looked as if this would be a technique to rival gas-liquid chromatography and HPLC. The activity developed quite rapidly into packed column supercritical fluid separations then into supercritical fluid extraction. However, in recent years there has been a decline in publications. These later techniques continue to be used but are now principally applied to a limited group of applications where they offer significant advantages over alternative techniques. This review looks back over this period and analyses how these methods were developed and the fluids, detectors and applications that were examined. It suggests why many of the initial applications have vanished and why the initial apparent promise was not fulfilled. The rise and fall of supercritical fluids represents a lesson in the way analysts approach new techniques and how we might view other new separation developments at the end of this millennium. The review looks forward to the future of supercritical fluids and their role at the end of the first century of separation science. Probably the most important idea that supercritical fluids have brought to separation science is a recognition that there is unity in the separation methods and that a continuum exists from gases to liquids. PMID- 10526786 TI - Recent developments in microcolumn liquid chromatography. AB - An overview of the most recent developments in microcolumn liquid chromatography (LC) is presented. A short theoretical discussion on chromatographic dilution and extracolumn bandbroadening is given and also the recent progress and advances in column technology and instrumentation are reviewed. However, the emphasis of this review is on miniaturized sample clean-up, sample introduction techniques and on both established and more recent detection techniques for microcolumn LC. The hyphenation of miniaturized LC columns with other techniques, specifically on multidimensional chromatography and the coupling of microcolumn LC to mass spectrometry is discussed in detail. Both the on-line and automated off-line interfacing to other separation and detection techniques will also be addressed. Finally, a number of typical microcolumn LC applications are presented in order to demonstrate the potential of microcolumn LC methods in a variety of scientific areas. PMID- 10526787 TI - Developments in sample preparation and separation techniques for the determination of inorganic ions by ion chromatography and capillary electrophoresis. AB - A review is presented of sample preparation and separation techniques for the determination of inorganic ions by ion chromatography (IC) and capillary electrophoresis (CE). Emphasis has been placed on those sample treatment methods which are specific to inorganic analysis, and the developments in separation methods which are discussed are those which enhance the capabilities of IC and CE to handle complex sample matrices. Topics discussed include solid-phase extraction for sample clean-up and preconcentration, dialytic methods, combustion methods, matrix-elimination IC, electrostatic IC, electrically polarised ion exchange resins, electromigration sample preparation in CE, chromatographic sample preparation for CE, use of high-ionic strength background electrolytes, buffering of background electrolytes in CE, use of capillary electrochromatography for inorganic determinations, and methods for the manipulation of separation selectivity in both IC and CE. Finally, some possible future trends are discussed. PMID- 10526788 TI - State-of-the-art in liquid chromatography-mass spectrometry. AB - Impressive progress has been made in the technology and application of combined liquid chromatography-mass spectrometry (LC-MS) in the past decennium. From a technique, that could only be used by a specialist, it has developed into a routinely applicable technique. LC-MS has become the method-of-choice of analytical support in many stages of drug development within pharmaceutical industries and has found its way into environmental, biochemical and other laboratories. This paper provides a perspective on the current technology, principles and applications of LC-MS. PMID- 10526789 TI - Liquid chromatography-nuclear magnetic resonance spectroscopy. AB - A general overview of the experimental set-up for performing analytical-scale and nanoliter-scale liquid chromatography-1H nuclear magnetic resonance spectroscopy (LC-1H-NMR) experiments is given. The high power of combining LC with 1H-NMR spectroscopy is demonstrated by two examples, where NMR acquisition was performed either in the continuous-flow mode on the analytical scale or in the stopped-flow mode on the nanoliter scale. Current developments employing the on-line coupling of capillary as well as supercritical fluid separation methods with 1H-NMR spectroscopy together with LC-13C-NMR spectroscopy are discussed. PMID- 10526790 TI - Liquid chromatography-Fourier-transform infrared spectrometry. AB - Over the past years the coupling of liquid chromatography (LC) and Fourier transform infrared spectrometry (FT-IR) has been pursued primarily to achieve specific detection and/or identification of sample constituents. Two approaches can be discerned in the combination of LC and FT-IR. The first and simpler approach is to use a flow cell through which the effluent from the LC column is passed while the IR spectra are continuously recorded. The second approach involves elimination of the LC solvent prior to IR detection using an interface which evaporates the eluent and deposits the analytes onto a substrate. This paper provides a general overview of flow-cell based IR detection and briefly discusses early solvent-elimination interfaces for LC-FT-IR. A more comprehensive description is given of interface systems which use spraying to induce rapid eluent evaporation, and which basically represent the state-of-the-art in LC-FT IR. Finally, the interface systems suitable for reversed-phase LC are summarized and the perspectives of LC-FT-IR are discussed. The overview indicates that flow cell LC-FT-IR has rather poor detection limits but can be useful for the specific and quantitative detection of major constituents of mixtures. Solvent-elimination techniques, on the other hand, provide much better sensitivity and enhanced spectral quality which is essential when unambiguous identification of low-level constituents is required. PMID- 10526791 TI - Liquid chromatography-inductively coupled plasma mass spectrometry. AB - The technique of coupling liquid chromatography to inductively plasma mass spectrometry (ICP-MS) is reviewed. A brief introduction to the ICP-MS instrument is given as well as methods to couple the two analytical instruments together. The various types of LC that have been used with ICP-MS detection are discussed and advantages over traditional methods of detection are highlighted, such as the improvements in sensitivity and selectivity. Several applications that have been described in the literature are reviewed. An outlook for the future of LC-ICP-MS, particularly with regard to elemental speciation is given. PMID- 10526792 TI - Programmed temperature vaporiser-based injection in capillary gas chromatography. AB - The application of programmed temperature vaporisation (PTV) in capillary gas chromatographic analysis is reviewed. The development of the different strategies as well as the state of the art are described. As the analytes are normally enriched in the PTV insert, the quoted papers are subdivided depending on whether the enrichment was carried out from organic solvents, from water or from gaseous media. Furthermore, the possibilities of PTVs for on-line coupling with sample preparation methods or other separation techniques and their use as thermoreactors are mentioned. PMID- 10526793 TI - On-line combination of aqueous-sample preparation and capillary gas chromatography. AB - Methods currently in use to combine the preparation of aqueous samples on-line with capillary gas chromatography (GC) comprise heartcut-orientated reversed phase liquid chromatography-GC and analyte-isolation-orientated analyte extraction-GC. These approaches either use techniques in which water is directly introduced onto the GC column, or an indirect approach in which water is eliminated, i.e., by solid-phase extraction, solid-phase microextraction or liquid-liquid extraction, prior to introduction of the analytes onto the GC column. The latter type of approach is much more successful and user-friendly, and many applications have been reported. PMID- 10526794 TI - High-speed gas chromatography: an overview of various concepts. AB - An overview is given of existing methods to minimise the analysis time in gas chromatography (GC) being the subject of many publications in the scientific literature. Packed and (multi-) capillary columns are compared with respect to their deployment in fast GC. It is assumed that the contribution of the stationary phase to peak broadening can be neglected (low liquid phase loading and thin film columns, respectively). The treatment is based on the minimisation of the analysis time required on both column types for the resolution of a critical pair of solutes (resolution normalised conditions). Theoretical relationships are given, describing analysis time and the related pressure drop. The equations are expressed in reduced parameters, making a comparison of column types considerably simpler than with the conventional equations. Reduction of the characteristic diameter, being the inside column diameter for open tubular columns and the particle size for packed columns, is the best approach to increase the separation speed in gas chromatography. Extremely fast analysis is only possible when the required number of plates to separate a critical pair of solutes is relatively low. Reducing the analysis time by reduction of the characteristic diameter is accompanied by a proportionally higher required inlet pressure. Due to the high resistance of flow of packed columns this seriously limits the use of packed columns for fast GC. For fast GC hydrogen has to be used as carrier gas and in some situations vacuum-outlet operation of capillary columns allows a further minimisation of the analysis time. For fast GC the columns should be operated near the conditions for minimum plate height. Linear temperature programmed fast GC requires high column temperature programming rates. Reduction of the characteristic diameter affects the sample capacity of the "fast columns". This effect is very pronounced for narrow-bore columns and in principle non-existing in packed columns. Multi-capillary columns (a parallel configuration of some 900 narrow-bore capillaries) take an intermediate position. PMID- 10526795 TI - Comprehensive two-dimensional gas chromatography: a hyphenated method with strong coupling between the two dimensions. AB - Comprehensive two-dimensional gas chromatography (GC x GC) provides a true orthogonal separation system. It is explained and demonstrated that it generates a peak capacity that is approximately equal to the product of the peak capacities of the two individual separation systems. The resulting peaks are ordered in a two-dimensional plane in bands of compounds with the same characteristics. Quantitation of the separated (groups of) components is fundamentally not different from one-dimensional gas chromatography, but the sensitivity is far better and true baseline is always available. The two co-ordinates of each peak in the plane make the identification more reliable. Instrumental considerations of GC x GC are discussed. The three designs of contemporary GC x GC systems are presented and compared. Although the technique is still very young, a number of applications on complex samples as petroleum and environmental samples have already been reported. Finally, the future perspectives of GC x GC are discussed. PMID- 10526796 TI - Gas chromatography with spectroscopic detectors. AB - In recent years, capillary gas chromatography (GC) with Fourier Transform Infrared (FT-IR) and/or mass spectral (MS) detection has become a primary analytical tool for qualitative and quantitative analysis of complex mixtures. Because of the wide range of applications, the analytical requirements have motivated a variety of chromatographic and detection developments. This review examines those, illustrating with applications that demonstrate the power of GC and multidimensional GC-MS, GC-FT-IR and GC-FT-IR-MS systems for solving a variety of analytical problems. In addition, the article discusses the integrated performance of such analytical systems with the aid of recent sample introduction and computer data analysis advances. PMID- 10526797 TI - Planar chromatography at the turn of the century. AB - An overview of the state-of-the-art of modern thin-layer chromatography (planar chromatography) is presented with emphasis on the complementary features of thin layer and column liquid chromatographic separations. The reasons for selecting thin-layer chromatography for a particular analysis are identified by its attributes: a disposable stationary phase; simultaneous parallel separations; static detection free of time constraints; storage device for chromatographic information; all sample components are observed in the chromatogram. Future prospects for improved separation performance in TLC using zone refocusing, forced flow and electroosmotic flow methods are discussed as well as increasing zone capacity by using two-dimensional development and coupling to column chromatographic methods. Advances in coupling thin-layer chromatography with spectroscopic methods for structural elucidation are also considered. Finally, some predictions are made for how thin-layer chromatography will be practiced in the future. PMID- 10526798 TI - The state of the art in thin-layer chromatography-mass spectrometry: a critical appraisal. AB - Thin-layer chromatography-mass spectrometry (TLC-MS) is a readily implemented technique that, in its simplest form, puts few demands on either chromatography or spectrometry. Nevertheless, compared to the situation with high performance liquid chromatography, it is much less highly developed. Currently, the bulk of the practical applications of TLC-MS are directed towards the use of fast atom, or ion bombardment. Recent developments, however, include the use of matrix assisted laser desorption ionisation (MALDI), surface assisted laser desorption (SALDI) and the development of a TLC-electrospray interface. Here, the state of the art of TLC-MS is described and future trends identified. PMID- 10526799 TI - Overview of capillary electrophoresis and capillary electrochromatography. AB - This paper provides an overview on the current status of capillary electrophoresis (CE) and capillary electrochromatography (CEC). The focus is largely on the current application areas of CE where routine methods are now in place. These application areas include the analysis of DNA, clinical and forensic samples, carbohydrates, inorganic anions and metal ions, pharmaceuticals, enantiomeric species and proteins and peptides. More specific areas such the determination of physical properties, microchip CE and instrumentation developments are also covered. The application, advantages and limitations of CEC are covered. Recent review articles and textbooks are frequently cited to provide readers with a source of information regarding pioneering work and theoretical treatments. PMID- 10526800 TI - Electrokinetic chromatography. AB - The important features of electrokinetic chromatography are critically reviewed. Special emphasis is given to systems using micelles as pseudostationary phase. Short and comprehensive overviews are given on the subjects of separation, comparison with capillary electrochromatography, on-line coupling with mass spectrometry, and developments that are expected in the future. A greater coverage on the subject of improvement of detection sensitivity, specifically by on-line concentration was also contributed. PMID- 10526801 TI - Coupling of biological sample handling and capillary electrophoresis. AB - The analysis of biological samples (e.g., blood, urine, saliva, tissue homogenates) by capillary electrophoresis (CE) requires efficient sample preparation (i.e., concentration and clean-up) procedures to remove interfering solutes (endogenous/exogenous and/or low-/high-molecular-mass), (in)organic salts and particulate matter. The sample preparation modules can be coupled with CE either off-line (manual), at-line (robotic interface), on-line (coupling via a transfer line) or in-line (complete integration between sample preparation and separation system). Sample preparation systems reported in the literature are based on chromatographic, electrophoretic or membrane-based procedures. The combination of automated sample preparation and CE is especially useful if complex samples have to be analyzed and helps to improve both selectivity and sensitivity. In this review, the different modes of solid-phase (micro-) extraction will be discussed and an overview of the potential of chromatographic, electrophoretic (e.g., isotachophoresis, sample stacking) and membrane-based procedures will be given. PMID- 10526802 TI - Analysis of DNA adducts using high-performance separation techniques coupled to electrospray ionization mass spectrometry. AB - Identification and quantitation of covalent carcinogen-DNA adducts, an important class of biomarkers, is an integral goal in toxicological research. Since these adducts are commonly present at very low levels in in vivo samples, sensitive and specific analytical methodologies are imperative for accurate detection, characterization and quantitation. High-performance separations coupled to electrospray mass spectrometry (ESI-MS) provide the sensitivity and specificity required for the analysis of DNA adducts. This review provides an overview over the research conducted in this area, focusing on the application of HPLC-ESI-MS and CE-ESI-MS techniques for structural analysis and quantitation of modified nucleosides, nucleotides and oligonucleotides. PMID- 10526803 TI - Respiratory flow measurements and anesthetic drugs: some in vitro observations. AB - STUDY OBJECTIVE: To illustrate the influence of anesthetic gases on respiratory flow measurements and to correct this influence. DESIGN: In vitro evaluation. SETTING: Laboratory. MEASUREMENTS AND MAIN RESULTS: An in vitro method using a 120-L Tissot water-seal spirometer was used along with a Bicore CP-100, designed for use in intensive care units, and a Datex Ultima, designed for use in the operating room. The flow transducer of one of the instruments being tested was placed in the gas inlet of the Tissot so that simultaneous measurements could be made. Timed flows (2 to 60 L/min) of various gases (O2 and air) and gas mixtures (halothane-O2, isoflurane-O2, N2O-O2, and N2O-O2-isoflurane) were used and the measurements made by the Tissot and the test instrument compared. The Datex Ultima, which includes software corrections for anesthetics, was able to accurately measure gas flows (2 to 60 L/min) of air, 100% oxygen, and anesthetic gas mixtures to within +/- 10% of measurements made by the Tissot. The Bicore CP 100, intended for use with mechanically ventilated patients, accurately measured air and 100% oxygen flow rate to within +/- 8% of the measurements made by the Tissot, but there were large errors (up to 40%) when anesthetics were used. CONCLUSIONS: This study illustrates the effects of anesthetic gases on measurements of ventilatory flow and the need to ascertain whether corrections are needed to improve the accuracy of flow transducers. PMID- 10526804 TI - Effects of sympathetic blockade on the efficiency of forced-air warming during combined spinal-epidural anesthesia for total hip arthroplasty. AB - STUDY OBJECTIVE: To evaluate if active cutaneous warming of the two upper limbs with reflex vasoconstriction is less effective in maintaining intraoperative normothermia than warming the vasodilated unoperated lower limb during combined spinal-epidural anesthesia (CSE). DESIGN: Prospective, randomized study. SETTING: Inpatient anesthesia at university departments of orthopedic surgery. PATIENTS: 48 ASA physical status I, II, and III patients, who were scheduled for elective total hip arthroplasty. INTERVENTIONS: Patients received CSE with intrathecal injection of 15 mg of 0.5% hyperbaric bupivacaine. All procedures started 8 to 10 AM, and operating room temperature was maintained between 21 degrees and 23 degrees C, with relative humidity ranging between 40% and 45%. For warming therapy, patients received active forced-air warming of either the two upper limbs (Group Upper body, n = 24), or the unoperated lower limb (Group Lower extremity, n = 24). Core temperature was measured before CSE placement (baseline), and then every 30 minutes until completion of surgery. Time for fulfillment of clinical discharging criteria from the recovery area was evaluated by a blinded observer. MEASUREMENTS AND MAIN RESULTS: Demographic data, duration of surgery, intraoperative blood losses, crystalloid infusion, and hemodynamic variables were similar in the two groups. Core temperature slightly decreased in both groups, but at the end of surgery the mean core temperature was 36.2 degrees +/- 0.5 degree C in Group Upper body and 36.3 +/- 0.5 in Group Lower extremity (NS). At recovery room arrival, seven patients in Group Upper body (29%) and three patients in Group Lower extremity (12.5%) had a core temperature less than 36 degrees C (NS). Shivering was observed in one patient in Group Upper body and in two patients in Group Lower extremity (NS). Clinical discharging criteria were fulfilled after 37 +/- 16 minutes in Group Upper body and 30 +/- 32 minutes in Group Lower extremity (NS). CONCLUSIONS: Forced-air cutaneous warming allows the anesthesiologist to maintain normothermia during CSE for total hip replacement even if the convective blanket is placed on a relatively small skin surface with reflex vasoconstriction. Placing the forced-air warming system on the vasodilated unoperated lower limb may be troublesome to the surgeons and does not offer clinically relevant advantages in warming efficiency. PMID- 10526805 TI - Remifentanil as an adjuvant during desflurane anesthesia facilitates early recovery after ambulatory surgery. AB - STUDY OBJECTIVE: To investigate the effect of using a remifentanil infusion during desflurane anesthesia on the early recovery profile and side effects. DESIGN: Randomized, single-blind study. SETTING: University-based ambulatory surgery unit. PATIENTS: 46 healthy, ASA physical status I and II women undergoing outpatient laparoscopic tubal ligation procedures. INTERVENTIONS: After premedication with midazolam 2 mg intravenously (IV), anesthesia was induced with propofol 2 mg.kg-1 i.v. and remifentanil 1 microgram.kg-1 i.v. Following tracheal intubation, anesthesia was maintained with desflurane 2% and nitrous oxide (N2O) 65% in both groups. During the maintenance period, hemodynamic stability was maintained using either a variable inspired concentration of desflurane, 2% to 8% (Control group), or a variable-rate infusion of remifentanil 0.05 to 0.2 microgram.kg-1.min-1 i.v. (Remi group). Ketorolac 30 mg i.v. and local anesthetic infiltration at the surgical portals were administered for preventive analgesia prior to skin closure. MEASUREMENTS AND MAIN RESULTS: Emergence times and times to achieving an Aldrete score of 10 (i.e., fast-tracking eligibility) were determined. Postoperative nausea and vomiting (PONV), as well as the need for analgesic and antiemetic rescue medications, were noted during the 24-hour follow up period. A structural questionnaire was used to assess intraoperative recall. Compared to the Control group, the Remi group had shorter emergence times and reduced times to achieving an Aldrete score of 10. There were no differences between the two groups with respect to the incidence of PONV and the requirements for postoperative analgesic and antiemetic drugs. None of the patients experienced intraoperative recall. CONCLUSIONS: The adjunctive use of a remifentanil infusion (0.07 +/- 0.03 microgram.kg-1.min-1) during desflurane-N2O anesthesia facilitated early recovery without increasing PONV, pain, or the need for rescue medication after laparoscopic surgery. PMID- 10526806 TI - Acute normovolemic hemodilution and nitroglycerin-induced hypotension: comparative effects on tissue oxygenation and allogeneic blood transfusion requirement in total hip arthroplasty. AB - STUDY OBJECTIVES: To study the comparative effects of acute normovolemic hemodilution and nitroglycerin-induced hypotension on tissue oxygenation and blood transfusion requirement. DESIGN: Prospective, randomized study. PATIENTS: 30 ASA physical status I and II patients scheduled for primary total hip arthroplasty. INTERVENTIONS: Patients were randomized to one of three groups of 10 patients each, to receive acute normovolemic hemodilutin (Group 1) or nitroglycerin-based hypotension (Group 2); Group 3 served as the control group. In Group 1, 2 U of blood was collected and replaced with an equal volume of hydroxyethyl starch (200/0.56%) immediately after anesthesia induction. In Group 2, nitroglycerin was infused at a rate sufficient to reduce mean arterial pressures to 60 to 65 mmHg before initiation of surgery. When hematocrit was reduced to 25%, at first autologous blood and then, if necessary, allogeneic blood was transfused to Group 1, and allogeneic blood was transfused to the other two groups, until hematocrit reached 30% for 5 days postoperatively. MEASUREMENTS AND MAIN RESULTS: Total transfused allogeneic units of blood were determined by the fifth postoperative day. Arterial oxygen content (CaO2), venous oxygen content (CvO2), and oxygen extraction ratios (EO2) were calculated by standard formulas. The mean allogeneic transfusion requirement was significantly lower in Group 1 (1.3 +/- 0.8 U) than in Group 2 (2.3 +/- 0.8 U) or Group 3 (2.7 +/- 1.1 U) (p < 0.05). In Group 1, CaO2 and CvO2 were decreased at all times, but EO2 was significantly increased from 15 +/- 3.9% to 33.3 +/- 5.3% (p < 0.001). As for the other two groups, although CaO2 and CvO2 were decreased, EO2 was not significantly increased. CONCLUSIONS: Acute normovolemic hemodilution is more effective than nitroglycerin-induced hypotension in reducing allogeneic blood transfusion requirement in total hip replacement surgery, without significant metabolic changes. PMID- 10526807 TI - Changes in respiratory pattern and arterial blood gases during sedation with propofol or midazolam in spinal anesthesia. AB - STUDY OBJECTIVE: To investigate changes in respiratory pattern and arterial blood gases during sedation with propofol or midazolam in spinal anesthesia. DESIGN: Randomized, placebo-controlled study. SETTING: Operating room of a university affiliated hospital. PATIENTS: 40 ASA physical status I and II patients who required spinal anesthesia. INTERVENTIONS: Spinal anesthesia with tetracaine and subsequent sedation with propofol (n = 15), midazolam (n = 15), or placebo (n = 10) was performed. MEASUREMENTS: Respiratory pattern [rib cage contribution to the tidal volume (%RC) and phase shift between rib cage and abdominal movements (PSrc-ab)] with a respiratory inductive plethysmograph (Respigraph) and arterial blood gas analysis (pH, pO2, and pCO2) were recorded. MAIN RESULTS: Spinal anesthesia per se increased %RC by 35% without changing PSrc-ab values (1.00). Sedation with propofol and midazolam decreased %RC by 60% and by 40%, respectively. PSrc-ab increased in both groups following sedation, and the increase in this parameter was higher in the propofol group (1.12) than in the midazolam group (1.04). In the placebo group, %RC decreased by 20% without any change in PSrc-ab. The decrease in pO2 was more significant in the propofol group (65.1 mmHg) than in the midazolam (74.2 mmHg) and placebo (83.1 mmHg) groups. CONCLUSION: Significant decreases in %RC and pO2 during propofol sedation seem to depend on paradoxical respiration due, in part, to upper airway obstruction; therefore, attention should be directed to the respiratory pattern during sedation, especially with propofol. PMID- 10526808 TI - Disseminating information using an anesthesiology consultant report: impact on patient perceptions of quality of care. AB - STUDY OBJECTIVE: To determine if providing an Anesthesiology Consultant Report (ACR) to patients would result in enhanced patients' perceptions of their knowledge about their care and improve their perception of the quality of their care. DESIGN: Randomized, unblinded study. SETTING: Outpatient center associated with tertiary care center. PATIENTS: 371 outpatients without adverse anesthetic events. INTERVENTIONS: Patients were randomized to receive either routine discharge instructions or routine instructions and an anesthesia discharge summary (ACR). MEASUREMENTS AND MAIN RESULTS: Short questionnaire with discharge packet regarding knowledge of anesthetic and questions regarding satisfaction and perceptions of quality of care was distributed. The patients in the group that received an ACR were more satisfied with the management of their pain and other symptoms (p < 0.05, by Wilcoxon rank sums) and were more satisfied overall with the quality of the anesthesia care (p < 0.01, by Wilcoxon rank sums). Taken another way, significantly more patients deemed the quality of their anesthetic care as excellent in the ACR group compared to control (83% vs. 67%, p < 0.01). CONCLUSIONS: Providing patients, with uneventful anesthetic courses, with information regarding their anesthetic care, in the form of the ACR, results in improved perceptions of the quality of care. PMID- 10526809 TI - Detection of fluid volume absorption by end-tidal alcohol monitoring in patients undergoing endoscopic renal pelvic surgery. AB - STUDY OBJECTIVE: To determine the risk of relevant fluid absorption (calculated volume above 500 ml) during endoscopic procedures of the renal pelvis. DESIGN: Prospective clinical investigation with implementation of statistical process control tools (SPC). SETTING: Nonuniversity teaching hospital. PATIENTS: 62 consecutive ASA physical status I and II patients scheduled for endoscopic renal pelvic surgery with general anesthesia. INTERVENTIONS: Intraoperative measurement of breath alcohol for detection of fluid absorption. Irrigation fluid (0.9% saline) with 1% alcohol for tracing the irrigation fluid. MEASUREMENTS AND MAIN RESULTS: Calculation of the amount of fluid absorbed using breath alcohol values. Process variability (numbers of patients with relevant fluid absorption) defined by SPC. The prevalence of fluid absorption in endoscopic renal pelvic surgery was 6%. Peak fluid absorption during a vascular route was detected by the monitoring. Monitoring was easily introduced into routine clinical practice. No relevant side effects due to the monitoring were seen in patients with relevant fluid absorption. There was no mortality, but two patients with detected severe fluid overload were admitted to the intensive care unit for treatment. CONCLUSION: Breath alcohol levels during general anesthesia for endoscopic renal pelvic surgery were technically simple to measure. Our results show the predictive value of alcohol monitoring, which has been previously demonstrated only for transurethral prostatectomy. The prevalence of relevant fluid absorption was 6% compared to 13% during transurethral resection of the prostate. PMID- 10526810 TI - Interaction modeling of propofol and sufentanil on loss of consciousness. AB - STUDY OBJECTIVES: To examine the possible pharmacodynamic interaction of propofol and sufentanil with respect to the induction of loss of consciousness. DESIGN: Prospective, randomized, double-blinded study. SETTING: University hospital. PATIENTS: 30 female, ASA physical status I and II patients undergoing elective gynecologic surgery. INTERVENTIONS: Patients were allocated randomly to receive an individual combination of propofol (1, 2, 3, 4, 5, or 6 micrograms/ml) and sufentanil (0.1, 0.2, 0.3, 0.5, or 1.0 ng/ml) target blood concentrations using target-controlled infusions. MEASUREMENTS AND MAIN RESULTS: Study endpoint was loss of consciousness, which was tested by response to verbal commands and classified as responder or nonresponder, as assessed by the anesthetist, who was blinded to the drugs' target blood concentrations. Nonlinear association (interaction) of both drugs was accomplished with logistic regression analysis using the maximum likelihood method, based principally on the hypothesis of interaction: In [p/(1-p)] = beta 0 + beta 1 x Cprop + beta 2 x Csuf + beta 3 x Cprop x Csuf with a p-value < 0.05 for coefficient estimates considered significant. In the logistic regression model, sufentanil and propofol showed no supra-additive interaction regarding loss of consciousness (p = 0.5916). CONCLUSIONS: Our results give no evidence of additional hypnotic properties of sufentanil compared to the other fentanyl congeners, although logistic regression may be of limited value in modeling interaction of hypnotic-analgesic combinations. PMID- 10526811 TI - Influences of age and gender on dose response and time course of effect of atracurium in anesthetized adult patients. AB - STUDY OBJECTIVE: To determine the influences of age and gender on the dose response and the time course of effect of atracurium. DESIGN: Prospective, nonrandomized, clinical comparison. SETTING: Operating room, Plastic Surgery Hospital of the Chinese Academy of Medical Sciences and Peking Union Medical College. PATIENTS: 72 adult ASA physical status I patients (38 male and 34 female), aged 15 to 59 years, scheduled for elective plastic surgery. INTERVENTIONS: Patients were divided into the three groups on the basis of age: Group 1, patients aged 15-29 years (n = 32); Group 2, patients aged 30-40 years (n = 21); and Group 3, patients aged 41-59 years (n = 19). Anesthesia was maintained with 60% nitrous oxide in oxygen, thiopental, and incremental doses of fentanyl as required. The dose-response relationship of atracurium was determined by a cumulative dose-response technique. MEASUREMENTS AND MAIN RESULTS: Neuromuscular function was assessed mechanomyographically with train-of-four stimulation at the wrist every 12 seconds and the percentage depression of first twitch (T1) response was used as the study variable. Age and gender significantly affected the dose-response relationship and time course of recovery of atracurium. Advancing age was associated with a reduced effective doses (ED50, ED90, and ED95) of atracurium and a longer duration of action. The effective doses of atracurium were greater, and its duration of action was shorter in men than in women. There were significant differences in the 50%, 90%, and 95% effective dose (ED50, ED90, and ED95) of atracurium, and clinical duration and total duration following administration of atracurium 400 micrograms/kg among the three age groups, and between men and women. CONCLUSIONS: Age and gender have significant effects on the dose response and time course of effect of atracurium. Older patients and women are more sensitive to atracurium-induced neuromuscular block than are young patients and men. PMID- 10526812 TI - Abdominal wall lift versus carbon dioxide insufflation for laparoscopic resection of ovarian tumors. AB - STUDY OBJECTIVE: To evaluate and compare changes in pulmonary mechanics and stress hormone responses between abdominal wall lift (gasless) and carbon dioxide (CO2) insufflation laparoscopic surgery during controlled general anesthesia. DESIGN: Prospective, randomized clinical study. SETTING: Operating rooms at a university medical center. PATIENTS: 12 ASA physical status I and II female patients undergoing laparoscopic resection of ovarian tumors. INTERVENTIONS: Patients were divided into two groups of six each: the abdominal wall lift group and the CO2 pneumoperitoneum laparoscopic group. Following induction of anesthesia, patients were paralyzed and the trachea was intubated. Anesthesia was maintained with isoflurane and nitrous oxide (N2O) in oxygen. Throughout the procedure, patients were mechanically ventilated with a tidal volume of 10 ml/kg and a respiratory rate of 10 breaths per minute. MEASUREMENTS AND MAIN RESULTS: During the laparoscopic procedure, arterial blood gases, acid-base balance, pulmonary mechanics, stress-related hormones, and urine output were measured and recorded. In the CO2 pneumoperitoneum group, arterial CO2 tension increased (p < 0.01), dynamic pulmonary compliance decreased (p < 0.01), peak inspiratory airway pressure increased (p < 0.01), and plasma epinephrine (p < 0.05), norepinephrine (p < 0.05), dopamine (p < 0.01), and antidiuretic hormones (p < 0.05) increased significantly during the laparoscopic procedure as compared to the abdominal lift group. Adrenocorticotropic hormone and cortisol increased as compared to baseline value in both groups (p < 0.05). Urine output was significantly less (p < 0.01) in the CO2 pneumoperitoneum group than in the abdominal wall lift group. CONCLUSIONS: Abdominal wall lift laparoscopic surgery is physiologically superior to CO2 pneumoperitoneum laparoscopic surgery as seen during the conditions of this study. Abdominal wall lift laparoscopic surgery provides normal acid-base balance and a lesser degree of hormonal stress responses, it maintains urine output, and it avoids derangement of pulmonary mechanics. PMID- 10526814 TI - Meralgia paresthetica: an unusual complication of post-cesarean analgesia. AB - Intramuscular (IM) injections may be associated with nerve injury, classically the sciatic nerve after intragluteal injection. We describe an unusual injury of the lateral femoral cutaneous nerve following an IM injection of 100 mg meperidine and 25 mg promethazine in the anterolateral right thigh. Although the thigh is advocated as a relatively safe site for IM injection, iatrogenic neuropathy may result. Awareness of the anatomy of the lateral femoral cutaneous nerve and avoiding injections into a partially anesthetized extremity may decrease the likelihood of recurrences. PMID- 10526813 TI - Dysphagia following intrathecal local anesthetic-opioid administration. AB - We describe the case of a parturient who developed dysphagia shortly after the intrathecal injection of a local anesthetic-opioid combination for cesarean section. Dysphagia was the only symptom of cephalad spread of the spinal anesthetic, and it was not associated with a "high" motor or sensory block. Although it resolved spontaneously, the dysphagia was extremely distressing to the patient. We conclude that dysphagia, even in the absence of an overtly "high spinal," should be added to the possible side effects of intrathecal local anesthetic-opioid administration in parturients. PMID- 10526815 TI - Prolonged perioperative myocardial ischemia in a young male: due to topical intranasal cocaine? AB - We present a case of prolonged myocardial ischemia in a young healthy male presenting for nasal polypectomy and tonsillectomy. Induction of anesthesia proceeded uneventfully. Immediately after surgical incision, the patient developed a sinus tachycardia with ST-segment depression in leads II and III, and ST elevation in leads aVR, aVL, aVF, and V. Depth of anesthesia was increased, esmolol was administered, which slowed the heart rate, and the procedure was terminated. However, myocardial ischemia only gradually resolved, leaving residual T-wave flattening in lead III by day 3 postoperatively. After extensive investigation to rule out other causes of ischemia, we considered cardiotoxicity due to intranasally administered cocaine with epinephrine to be the most likely precipitant. Nasal packing with gauze soaked in a solution containing cocaine 3 mg/kg and epinephrine 1 mg occurred just 40 minutes prior to induction of anesthesia. Topical intranasal cocaine is rapidly and reliably absorbed systemically, with peak plasma concentrations occurring within 30 to 60 minutes, corresponding to the time course of cocaine administration and surgical stimulation in this patient. Systemic absorption of topical intranasal cocaine has previously been reported to cause adverse cardiac sequelae, including myocardial infarction. This report reinforces the need for caution regarding the use of topical intranasal cocaine, particularly if used in combination with epinephrine. PMID- 10526816 TI - Hypotension and adrenal insufficiency. AB - This case conference describes two patients with hypotension who eventually were diagnosed with adrenal insufficiency. The first patient was initially believed to have a cardiac abnormality and the second patient sepsis, which was causing the continued hypotension. Both patients exhibited several clinical similarities; however, neither had the classic symptoms of adrenal insufficiency. This report discusses the causes of postoperative hypotension, diagnostic testing, and treatment for patients with adrenal insufficiency, and briefly reviews the literature. PMID- 10526817 TI - Anesthetic care for the child with congenital central alveolar hypoventilation syndrome (Ondine's curse). AB - Idiopathic congenital central alveolar hypoventilation syndrome, otherwise known as Ondine's curse, is a rare neuropathologic syndrome characterized by an inadequate respiratory drive with hypoventilation and periods of prolonged apnea resulting in hypercarbia and hypoxemia. Although no definite pathologic abnormality has been identified to account for the disorder, it is thought to represent a primary defect related to altered function of central chemoreceptors resulting in defective control of minute ventilation. Associated problems related to neural crest cell migration, including neuroblastoma formation and Hirschsprung's disease, suggest that the primary defect is defective neural crest cell migration and function. Problems that may impact on perioperative care include the defective central control of ventilation and defective control of upper respiratory musculature, which may lead to upper airway obstruction. Although many patients will have previously undergone tracheostomy and chronic mechanical ventilation, problems in other organ systems can impact on perioperative care. Cardiovascular issues include the possible presence of cor pulmonale and autonomic nervous system dysfunction. Central nervous system issues include the frequent occurrence of seizures and mental retardation. The preoperative work-up, premedication, and the intraoperative/postoperative care and monitoring of these patients is reviewed. PMID- 10526818 TI - Iatrogenic bacterial meningitis after spinal anesthesia for pain relief during labor. PMID- 10526819 TI - Diabetic silent hearts and anesthesia. PMID- 10526820 TI - Extubation in adult patients: who, what, when, where, how, and why? PMID- 10526821 TI - Tracheal extubation of adult surgical patients while deeply anesthetized: a survey of United States anesthesiologists. AB - STUDY OBJECTIVE: To examine current practice regarding the performance of tracheal extubation of adult surgical patients while deeply anesthetized (deep extubation). DESIGN AND SETTING: Survey comprised of an anonymous written questionnaire mailed to 1,000 randomly selected active American Society of Anesthesiologists members. MEASUREMENTS AND MAIN RESULTS: Questionnaires were mailed between February and April 1998. Five hundred eighty-three completed forms were returned, 538 of which were suitable for data analysis. Responses from anesthesiologists who infrequently or never administer general anesthetics to adult surgical patients were excluded. The overall frequency of deep extubation of adults was "never" for 106 respondents (19.7%), "rarely" for 87 (16.2%), and "more frequently" for 345 (64.1%). The most common reasons for never performing deep extubations were lack of necessity and concern regarding potential laryngospasm and aspiration. The most frequent indications for deep extubations were unclipped intracranial aneurysm, reactive airway disease, and open-globe eye surgery. The most frequent contraindications to deep extubations for those who otherwise perform the technique were difficult airway, aspiration risk, and obesity. After performing a deep extubation, 44.0% of respondents remain with the patient in the operating room until he or she is awake. Deep extubations were perceived to have no consistent effect on operating room turnover time by 61.6% of anesthesiologists who perform them. CONCLUSIONS: Most anesthesiologists in this survey perform deep extubations in adult surgical patients. Lack of necessity and potential respiratory complications were the main reasons cited by those who do not use the technique. Future investigations are necessary to examine the risk-to-benefit ratio of the technique in adults. Our results may be used to determine which potential indications should be examined in such studies and to help delineate the standard of care followed in this country. PMID- 10526822 TI - Efficacy of repeat intravenous dosing of ondansetron in controlling postoperative nausea and vomiting: a randomized, double-blind, placebo-controlled multicenter trial. AB - STUDY OBJECTIVES: To compare repeat intravenous (i.v.) dosing of ondansetron 4 mg with placebo for the treatment of postoperative nausea and vomiting (PONV) in patients for whom prophylactic, preoperative ondansetron 4 mg i.v. was inadequate DESIGN: Randomized, double-blind, placebo-controlled study. SETTING: Ten outpatient surgical centers in the United States. PATIENTS: 2,199 male and female ASA physical status I, II, and III patients > or = 12 years old scheduled to undergo outpatient surgical procedures and receive nitrous oxide-based general anesthesia. INTERVENTIONS: Ondansetron 4 mg i.v. was administered to all patients before induction of general anesthesia. Patients who experienced PONV or requested antiemetic therapy within 2 hours after discontinuation of inhaled anesthesia were randomized (1:1) to either a repeat i.v. ondansetron 4 mg dose or placebo. MEASUREMENTS AND MAIN RESULTS: Of the 2,199 patients prophylactically treated with ondansetron 4 mg before anesthesia induction, 1,771 (80.5%) did not experience PONV or request antiemetic therapy during the 2 hours following discontinuation of anesthesia. Of the 428 patients who experienced PONV or requested antiemetic therapy during the same period, and were randomized to additional treatment (214 randomized to ondansetron, 214 randomized to placebo), the incidence of complete response (no emesis, no rescue medication, no study withdrawal) was similar for both ondansetron-randomized and placebo-randomized groups for the 2-hour (34% and 43%, respectively, p = 0.074) and 24-hour (28% and 32%, respectively, p = 0.342) postrandomization study periods. Repeat ondansetron dosing was not more effective than placebo in controlling either postoperative emesis or the severity/duration of postoperative nausea. The administration of an additional dose of ondansetron 4 mg postoperatively did not result in an increased incidence of adverse effects. CONCLUSIONS: In patients for whom preoperative prophylaxis with ondansetron 4 mg i.v. is not successful, a repeat dose of ondansetron 4 mg i.v. in the postanesthesia care unit does not appear to offer additional control of PONV. PMID- 10526823 TI - Recovery and pharmacokinetic parameters of desflurane, sevoflurane, and isoflurane in patients undergoing urologic procedures. AB - STUDY OBJECTIVE: To compare the pharmacokinetics and the speed of recovery after inhalation anesthesia with desflurane, sevoflurane, and isoflurane in elective surgery. DESIGN: Prospective, randomized study. SETTING: University medical center. PATIENTS: 30 ASA physical status I and II adults presenting for elective surgery. INTERVENTIONS: Anesthesia was induced with etomidate and maintained with desflurane (n = 10), sevoflurane (n = 10), or isoflurane (n = 10) and nitrous oxide. The inhalation drugs were titrated until an adequate clinical depth of anesthesia was reached. At the end of anesthesia, the patients breathed oxygen via the endotracheal tube and after extubation via a face mask. MEASUREMENTS AND MAIN RESULTS: The groups were similar with respect to age, weight, duration of anesthesia, and mean arterial pressure. Mean end-tidal concentration (FA = FA0) at the end of anesthesia was 6.34 +/- 1.15% after desflurane, 1.85 +/- 0.42% after sevoflurane, and 1.10 +/- 0.24% after isoflurane. FA/FA0 decreased significantly faster with desflurane than with isoflurane, while there was little difference between desflurane and sevoflurane. As for the terminal half-life (t1/2), there were no differences among the groups (8.16 +/- 3.15 min after desflurane, 9.47 +/- 4.46 min after sevoflurane, and 10.0 +/- 5.57 min after isoflurane). The time until a command was followed for the first time was the same in all three groups (13.0 +/- 4.7 min after desflurane, 13.4 +/- 4.4 min after sevoflurane, and 13.6 +/- 3.4 min after isoflurane). There was no significant correlation between duration of anesthesia and the time until recovery. CONCLUSIONS: There are only minor differences with regard to the recovery phase in premedicated patients who receive clinically titrated inhalation anesthesia with desflurane, sevoflurane, or isoflurane. PMID- 10526824 TI - Effect of dexmedetomidine on the minimum alveolar concentration (MAC) of sevoflurane in adults age 55 to 70 years. AB - STUDY OBJECTIVE: To determine the effect of two target dexmedetomidine infusions (0.3 ng/ml and 0.6 ng/ml) on the minimal alveolar concentration (MAC) of sevoflurane in adults age 55 to 70 years. DESIGN: Prospective, randomized, placebo-controlled study. SETTING: University-affiliated hospital. PATIENTS: 45 ASA physical status I and II adults, age 55 to 70 years, undergoing elective surgery with at least a 3 inch skin incision. INTERVENTIONS: Patients were given a dexmedetomidine or placebo infusion for at least 15 minutes before anesthetic induction with sevoflurane in oxygen by face mask. After tracheal intubation, a target sevoflurane concentration was maintained for 15 minutes while the dexmedetomidine or placebo infusion continued to run. MEASUREMENTS AND MAIN RESULTS: At the time of skin incision, two observers independently determined movement or nonmovement to the incision. Blood samples for dexmedetomidine were taken before the infusion and at the time of skin incision. The dexmedetomidine plasma concentrations were 0 before infusion with all treatment groups. At the time of incision, they were 0 in the placebo group, 0.39 +/- 0.13 ng/ml in the 0.3 ng/ml target group, and 0.7 +/- 0.13 ng/ml in the 0.6 ng/ml target group. The MAC of sevoflurane was 1.83% in the placebo group, 1.78% in the 0.3 ng/ml target dexmedetomidine group, and 1.51% in the 0.6 ng/ml target dexmedetomidine group. CONCLUSIONS: Dexmedetomidine 0.7 ng/ml decreased the MAC of sevoflurane by 17%, whereas there was no difference between the placebo and the dexmedetomidine 0.39 ng/ml group. PMID- 10526825 TI - A bougie improves the utility of the UpsherScope. AB - STUDY OBJECTIVE: To assess the alignment of the tube-guide and visual-guide channels of the UpsherScope and to evaluate methods to improve the success of tracheal intubation. DESIGN: In-vitro observation followed by clinical series. SETTING: Tertiary care, academic medical institution. PATIENTS: 56 surgical patients. MEASUREMENTS AND MAIN RESULTS: In an in-vitro study, cross-marked concentric circles were used as the target for the tracheal tube. It confirmed that the tube-guide channel of the UpsherScope directs the tip of the tracheal tube posteriorly and to the right of the visual field when the tube was loaded into the guide channel, according to the manufacturer's recommendations. Of five other methods used for maneuvering the tracheal tube to the target, use of a bougie within the lumen of the tube resulted in the highest success rate. When this method was assessed in a clinical setting, it was successful in directing the tracheal tube into the trachea of 95% of the patient population. Two failures were due to secretions obscuring the view, and one, to a broken bougie. CONCLUSIONS: There is a functional misalignment between the long axes of the tube guide and visual-guide channels of the UpsherScope. Use of a bougie will minimize this potential problem. PMID- 10526826 TI - Cost analysis of xenon anesthesia: a comparison with nitrous oxide-isoflurane and nitrous oxide-sevoflurane anesthesia. AB - STUDY OBJECTIVE: To determine the cost of xenon (Xe) anesthesia in relation to the anesthetic duration by conducting a cost analysis of this relatively expensive inhaled anesthetic. DESIGN: Cost analysis based on the literature on Xe anesthesia. SETTING: Anesthetic simulation based on data obtained in the operating rooms at a university hospital. PATIENTS: A 40-year-old, ASA physical status I adult patient model weighing 70 kg, undergoing elective minor surgery with endotracheal intubation and mechanical ventilation. INTERVENTIONS: Anesthesia was given in the following four techniques: 1) closed-circuit technique with Xe; 2) closed-circuit technique with nitrous oxide (N2O) isoflurane; 3) semi-closed technique with N2O-isoflurane; and 4) semi-closed technique with N2O-sevoflurane. MEASUREMENTS AND MAIN RESULTS: Cost of each anesthetic technique was compared in U.S. dollars. The cost of Xe anesthesia was consistently higher than that of N2O-isoflurane or N2O-sevoflurane (for 240-min anesthesia; $356 with Xe, $52 with closed-circuit N2O-isoflurane, $94 with semi closed N2O-isoflurane, and $84 with semi-closed N2O-sevoflurane). The major cost of Xe anesthesia was a result of the cost of priming and flushing; the cost of Xe used for its anesthetic effects was comparable with the other semi-closed techniques after 240 minutes. CONCLUSIONS: For Xe to be widely used in routine anesthesia, the methods of minimizing the amount of Xe necessary for priming and flushing must be developed. PMID- 10526827 TI - High incidence of intravenous thrombi after short-term central venous catheterization of the internal jugular vein. AB - STUDY OBJECTIVE: To assess incidence and characteristics of intravenous (i.v.) thrombi associated with short-term central venous catheterization through the internal jugular vein. DESIGN: Prospective clinical study. SETTING: University hospital. PATIENTS: 81 patients undergoing cardiac surgery. INTERVENTIONS: A triple-lumen central venous catheter was inserted into the right internal jugular vein immediately before surgery and removed 3 to 4 days later. Heparin at an i.v. dose of 15,000 IU/24 hours was started 6 hours after surgery and continued until the first postoperative morning, followed by subcutaneous low molecular weight heparin 5,000 IU/day in combination with oral aspirin 100 mg/day. MEASUREMENTS AND MAIN RESULTS: Anatomy of the internal jugular vein and i.v. blood flow were studied using two-dimensional and color Doppler ultrasonography before insertion of the catheter and after its removal. Thrombi were found in 45 patients (56%). Twenty-five of these thrombi (56%) had the shape of a sleeve, and 20 thrombi (44%) were compact. Length of the thrombi was 1.4 +/- 0.8 cm (mean +/- SD). Half of the thrombi floated with venous blood flow and half were stable. Neither impaired venous blood flow nor clinical signs of embolism or sepsis was found. Follow-up studies in eight patients revealed that the thrombi had not disappeared 5 days after removal of the catheter but had become smaller. CONCLUSION: The incidence of i.v. thrombi associated with short-term catheterization of the internal jugular vein was high despite prophylactic anticoagulation. This finding reaffirms the importance of removing central venous catheters as soon as clinically possible. Additional studies using specific outcome tests are needed to thoroughly assess the clinical importance of this finding. PMID- 10526828 TI - Acute renal failure following laparoscopic cholecystectomy: a case report. AB - The carbon dioxide (CO2) pneumoperitoneum of laparoscopic surgery is a complex physiologic event associated with neuroendocrine, respiratory, cardiovascular, and renal disturbances, as well as compromised organ blood flow. A case is presented of a 67-year-old man with a history of chronic renal failure, renal tubular acidosis, and hypertension, who underwent an uneventful elective laparoscopic cholecystectomy that included 75 minutes of CO2 pneumoperitoneum of 15 mmHg pressure. Postoperatively, the patient developed acute renal failure from which he recovered within 2 weeks. In the absence of other evident precipitating factors, we suspect that the CO2 pneumoperitoneum played a causal role in the development of his acute renal failure. The potential seriousness of the physiologic insult of conventional CO2 pneumoperitoneum suggests that "minimal access" surgery is not necessarily "minimally invasive." PMID- 10526829 TI - Use of laryngeal mask airway in a patient requiring continuous positive airway pressure: a case report. AB - The successful use of a laryngeal mask airway over a 48-hour period is reported in a patient with partial upper airway obstruction who required continuous positive airway pressure. PMID- 10526831 TI - Behavior disturbances with repeated propofol sedation in a child. PMID- 10526830 TI - Bragdon v. Abbott. PMID- 10526832 TI - Organization of a comprehensive anesthesiology oral practice examination program: planning, structure, startup, administration, growth, and evaluation. AB - STUDY OBJECTIVE: To describe the planning, structure, startup, administration, growth, and evaluation of a comprehensive oral practice examination (OPE) program. SETTING: Midwest U.S. anesthesiology residency training program. MEASUREMENTS AND MAIN RESULTS: Committee planning involved consideration of formal and frequency of administration, timing for best resident and faculty availability, communication, forms design, clerical support, record keeping, and quality monitoring. OPE format was deliberately constructed to resemble that used by the American Board of Anesthesiology (ABA) to enhance resident familiarity with ABA style oral examination. Quality improvement tools consisted of regular examiner and examinee inservice sessions, liaison with ABA associate examiners, and review of examinee exit questionnaires. A set of OPE databases was constructed to facilitate quality monitoring and educational research efforts. A semiannual administration schedule on three to four consecutive Mondays optimally accommodated resident rotations and faculty work schedules. Continued administration of the OPE program required ongoing construction of a pool of guided case-oriented questions, selection of appropriate questions based on examinee training exposure, examination calendar publication, and scheduling of recurring examiner and examinee activities. Important issues that required action by the governing committee were examination timing, conflict with clinical demands, use of OPE results, and procurement of training resources. The OPE program grew from 56 examinations in the first year to 120 exams by year 3. It was perceived positively by the majority of residents. There were 90.2% of exit questionnaires that acknowledged specific learning about oral examination technique, while only 0.3% indicated lack of meaningful information exchange. Fewer than 10% of responses indicated misleading questions or badgering by examiners. Resident preparedness increased with repeat OPE exposure. CONCLUSIONS: A comprehensive mock oral examination program was successfully planned, initiated, and developed. It is well accepted by residents and faculty. Its inception was associated with an increase in resident preparedness. Now in its tenth year of existence it continues to be an asset and essential component of our training program. PMID- 10526833 TI - Evaluation, feedback, and remediation in anesthesiology residency training: a survey of 124 United States programs. AB - STUDY OBJECTIVE: To provide a review of evaluation, feedback, and remediation methods in United States residency programs during 1995 to 1996. The information gathered is to serve as a framework for discussions within and among programs regarding ways to enhance their current processes of evaluation, feedback, and remediation, and to serve as a baseline for future assessments. DESIGN, SETTING AND SUBJECTS: A three-page survey was mailed to program directors of each of the 145 anesthesiology programs listed in the Accreditation Council for Graduate Medical Education (ACGME/NRMP) Directory. MEASUREMENTS AND MAIN RESULTS: Quantitative and qualitative responses were sought about the resident evaluation process (including techniques of gathering information, frequency of evaluations, faculty compliance, and modes of offering feedback), departmental clinical competence committee, probation and remediation policies for problem residents, and the use of formal examinations. There was an 85.5% response rate. Frequency of evaluation of residents ranged from daily to quarterly: evaluations used both narrative comments and rating scales in 89% of institutions. Faculty compliance in the evaluation process was greater than 75% in 45.1% of programs. Only 25 (20.2%) programs offered formal training about resident evaluation to their faculty. Clinical competence committee meetings averaged five times annually. Ninety-five percent of committees were chaired by someone other than the department chairperson and 27% had resident members. A written policy regarding problem residents was used by 67.7% of programs, a formal probation policy by 82.2%. Standardized tests to provide feedback and guidance to residents existed in 48.3% of programs. CONCLUSIONS: There is a tremendous variety of techniques and methodologies employed among anesthesiology residency programs with regard to evaluation, feedback, and remediation, within the framework of the ACGME guidelines. Faculty training in the assessment of and feedback to residents is one area in which many programs can begin to strengthen their current procedures. PMID- 10526834 TI - Introduction to the special issue: work-family research in occupational health psychology. AB - In this introduction, the authors discuss work-family research in the context of occupational health psychology (OHP), describe the special contributions of articles in this special issue, and outline directions for the next generation of research in the field of OHP. PMID- 10526836 TI - Value attainment: an explanation for the negative effects of work-family conflict on job and life satisfaction. AB - Perceptions of work interfering with family life and family issues interfering with work are examined as 2 distinct constructs representing work-family conflict. Experienced work-family conflict is argued to reduce one's value attainment which, in turn, lowers both job and life satisfaction. This study examines value attainment as a mediating variable in the work-family conflict and satisfaction relationship. Responses from 270 hotel managers indicate that value attainment either partially or fully mediates the relationship between work interference with family and family interference with work and both job and life satisfaction. Value attainment is argued to be a meaningful explanatory variable for the negative relationship between work-family conflict and job-life satisfaction. PMID- 10526835 TI - Fit as a mediator of the relationship between work hours and burnout. AB - The authors studied number of hours worked and estimated its relationship to burnout in a nonrandom sample of 141 married physicians. It was hypothesized that this relationship is mediated by a process called fit, conceptualized as the extent to which workers realize the various components of their work-family strategies. Results of structural equation modeling supported the mediation hypothesis. Employees whose work hours are more or fewer than they and their partner prefer and whose work hours are distributed differently than they and their partner prefer will be more disengaged, distracted, and alienated at work than will their counterparts who are working their preferred schedules. Thus, the relationship between number of hours worked and burnout depends on the extent to which work schedules meet the needs of the worker, her or his partner, and their children, if any. PMID- 10526837 TI - Work-family conflict, spouse support, and nursing staff well-being during organizational restructuring. AB - This study examined work and family conflict, spouse support, and nursing staff well-being during a time of hospital restructuring and downsizing. Data were collected from 686 hospital-based nurses, the vast majority (97%) women. Nurses reported significantly greater work-family conflict than family-work conflict. Personal demographic but not downsizing and restructuring variables predicted family-work conflict; downsizing and restructuring variables but not personal demographics predicted work-family conflict. Spouse support had no effect on work family conflict but reduced family-work conflict. Both work-family conflict and family-work conflict were associated with less work satisfaction and greater psychological distress. PMID- 10526839 TI - Parents' job insecurity affects children's grade performance through the indirect effects of beliefs in an unjust world and negative mood. AB - The authors postulated a model in which children's perceptions of their parents' job insecurity indirectly affect their grade performance through the effects of beliefs in an unjust world and negative mood. A total of 127 undergraduate students (55 male, 72 female) completed questionnaires on their perceptions of their parents' job insecurity and their own beliefs in an unjust world and negative mood. The parents reported on their own job insecurity. In addition, students provided their course grades from the previous semester 3 months after completing the questionnaires. Support for the proposed model was provided using LISREL 8. PMID- 10526838 TI - The source, nature, and direction of work and family conflict: a longitudinal investigation. AB - The authors examine the source, nature, and direction of work and family conflict. Confirmatory factor analysis of a 22-item scale suggested the appropriateness of distinguishing between strain-based and time-based conflict and between family interfering with work (FIW) and work interfering with family (WIF). Six-month longitudinal survey data (N = 236) suggested that strain-based FIW is a precursor to both stress and turnover intentions. Strain-based WIF emerged as an outcome of stress. PMID- 10526840 TI - The impact of dependent-care responsibility and gender on work attitudes. AB - On the basis of a survey of 18,120 federal employees in dual-income households, six 5-stage hierarchical multiple regression analyses, controlling for 10 demographic variables, assessed the impact of child care, elder care, and gender on work-family balance and various facets of job satisfaction. Elder-care responsibility was associated with lower levels of satisfaction with perceived organizational support, pay, leave benefits, and work-family balance, whereas the negative main effects of child care were limited to leave benefits and work family balance. However, child-care responsibility also interacted with gender: Its negative influence was greater on women's work-family balance and leave satisfaction. Decrements in satisfaction associated with dependent care on the "sandwich generation" were additive, not interactive. PMID- 10526841 TI - The home as a workplace: work-family interaction and psychological well-being in telework. AB - Home-based telework is a growing phenomenon with great potential to affect employees' psychological well-being. Although prior studies show both positive and negative effects on work-family interaction, conclusions are limited by the way telework, well-being, and work-family interaction have been modeled. The authors present a conceptual framework that describes telework as a multidimensional phenomenon and separates the effects of the home environment from those of distance from the organization. Propositions concerning work-family interaction are developed from P. Warr's (1987) model of the environmental antecedents of well-being, prior telework studies, and the work-family literature. Spillover between work and nonwork domains of well-being is discussed, and suggestions for future research on this complex issue are presented. PMID- 10526842 TI - The work-family research agenda in changing contexts. AB - This article argues that research on the work-family interface has evolved against a backdrop of dramatic and ongoing social and workplace change and must continue to reflect current and future context. The article overviews current trends that have implications for work and family and considers some possible future scenarios. It identifies a number of research areas and questions that build on previous theoretical and practical developments in the work-family field and reflect current trends. It is argued that questions about the well-being and sustainability of workplace human resources, of families in their diverse forms, and of communities are of overriding significance for the work-family research agenda, particularly if current trends continue unabated. PMID- 10526843 TI - Work-related stress--it's time to act. PMID- 10526844 TI - The Tokyo Declaration on Work-Related Stress and Health in Three Postindustrial Settings--the European Union, Japan and the United States of America. PMID- 10526845 TI - Surfactant protein A in rabbit sinus and middle ear mucosa. AB - In the present study, pulmonary surfactant protein A (SP-A) messenger RNA (mRNA) and protein were characterized in adult rabbit middle ear and maxillary sinus. Fifteen adult rabbits were used for the study: 6 with evidence of acute middle ear infections and maxillary sinusitis, 6 with infections that were successfully treated with tetracycline, and 3 that were pathogen-free. We detected SP-A mRNA in maxillary sinus and middle ear tissues by Northern blot analysis and reverse transcriptase-polymerase chain reaction (RT-PCR). The RT-PCR also revealed the presence of SP-B and SP-C mRNA in middle ear and sinus tissues. We detected SP-A protein, of molecular weight approximately 29 and 70 kd, in middle ear and sinus tissues by immunoblot analysis. Unlike the SP-A protein present in the lung, the molecular weight of the SP-A protein present in the middle ear and paranasal sinus was not altered by digestion with an enzyme that cleaves N-linked carbohydrates. Immunostaining and in situ hybridization showed that SP-A protein and mRNA, respectively, were present in surface epithelial cells of the middle ear and in epithelial cells of submucosal glands in sinus tissues. These data provide the first evidence of the presence of pulmonary surfactant proteins in the paranasal sinuses and confirm previous reports of SP-A in the middle ear epithelium. PMID- 10526846 TI - Cervical vertebral anomalies in patients with anomalies of the head and neck. AB - Congenital head and neck anomalies can occur in association with vertebral anomalies, particularly of the cervical vertebrae. While the former are easily recognized, especially when part of a syndrome, the latter are often occult, thereby delaying their diagnosis. The presence of vertebral anomalies must be considered in pediatric patients with head and neck abnormalities to expedite management of select cases and to prevent neurologic injury. We present our experience with 5 pediatric patients who were referred to the Department of Otolaryngology-Head and Neck Surgery at the University of Iowa with a variety of syndromic anomalies of the head and neck. Each patient was subsequently also found to have a vertebral anomaly. The relevant embryogenesis of the anomalous structures is discussed, with highlighting of potential causes such as teratogenic agents and events and germ-line mutations. A review of syndromes having both head and neck and vertebral anomalies is presented to heighten awareness of otolaryngologists evaluating children with syndromic disorders. Finally, the findings on radiographic imaging studies, particularly computed tomography, are discussed to facilitate the prompt diagnosis of vertebral anomalies. PMID- 10526847 TI - Immortalization of chinchilla middle ear epithelial cells by adenovirus 12-simian virus 40 hybrid virus. AB - In order to study the cellular and molecular mechanisms of the pathogenesis of otitis media, a chinchilla middle ear epithelial cell line (CMEE-1) with differentiated cell characteristics was established by infection of a primary culture with the adenovirus 12-simian virus 40 (Ad12-SV40) hybrid. This cell line has been in continuous culture for 42 passages, whereas the parent cells underwent senescence and died at the 8th passage. The cell line also retains epithelial morphology and expresses cytokeratin polypeptides 4, 7, and 18, characteristic markers for epithelia. In Western blots of cell proteins, bands at 94 and 53 kd were labeled after binding antibodies against SV40 large T antigen and p53, respectively. Karyotype analysis showed that the cell line is derived from chinchilla epithelial cells. These findings confirm that the cell line is a chinchilla epithelial cell immortalized by the hybrid virus. PMID- 10526848 TI - Effect of carbogen inhalation on peripheral tissue perfusion and oxygenation in patients suffering from sudden hearing loss. AB - The effects of repeated carbogen inhalation on peripheral tissue perfusion and oxygenation were assessed in 5 patients suffering from sudden hearing loss by means of continuously measured subcutaneous tissue oxygen and carbon dioxide tension, transcutaneous oxygen tension, laser Doppler red cell flux, and fingertip temperature. The subcutaneous oxygen tension increased clearly during the carbogen inhalation periods, and also, a smaller increase in subcutaneous carbon dioxide tension was simultaneously noticed. The changes in transcutaneous oxygen tension were even greater and the latency was shorter as compared with the subcutaneous gas tensions. The laser Doppler measurements showed no signs of vasoconstriction during the study. In conclusion, carbogen inhalation increases peripheral tissue oxygenation without microvascular vasoconstriction and with only a minor retention of carbon dioxide. PMID- 10526849 TI - Discovering diagnostic rules from a neurotologic database with genetic algorithms. AB - Data on patients with Meniere's disease, vestibular schwannoma, traumatic vertigo, sudden deafness, benign paroxysmal positional vertigo, or vestibular neuritis were retrieved from the database of otoneurologic expert system ONE for the development and testing of a genetic algorithm (GA). The accuracy of the diagnostic rules in solving the test cases was 81%, 91%, 92%, 95%, 96%, and 98% for the respective diseases. The best rules retrieved from the GA were described by a set of questions with the most likely answers. The most important questions concerned the duration of hearing loss and the occurrence of head injury. The validity and structure of the rules created with a GA can be analyzed in detail. For rare diseases, some other reasoning process can be used, for example, case based reasoning. PMID- 10526850 TI - Cochlear function of guinea pigs with experimental chronic renal failure. AB - This study aimed to evaluate electrophysiologically the cochlear function of guinea pigs that underwent a five-sixths nephrectomy and, additionally, to explore the synergistic action between chronic renal failure (CRF) and noise. Cochlear potentials were recorded at 1, 2, and 3 months postoperatively. Slight changes in compound action potential and cochlear microphonics were seen at 1 month postoperative, while moderate and profound changes were seen at 2 and 3 months. Endocochlear potential showed no significant reduction. The results indicate that CRF may be an etiologic factor for cochlear dysfunction and that the hair cells seem likely to be a main site of the lesion. One-month postoperative animals were exposed to a broadband noise. In contrast to control animals, the test animals demonstrated no recovery from the decrease in compound action potential and cochlear microphonics that occurred immediately after noise exposure. This suggests a synergistic interaction between CRF and noise. PMID- 10526851 TI - Distribution of psammoma bodies in the internal auditory canal and its extended areas in the human temporal bone. AB - The internal auditory canal (IAC) and its extended areas of 27 normal human temporal bone specimens were investigated histologically for the distribution of psammoma bodies. A total of 145 +/- 25 (mean +/- SE) psammoma bodies were counted in series of every tenth 30-microm-thick section. Psammoma bodies were observed in the IAC and around the labyrinthine portion of the facial nerve (FN), the geniculate ganglion of the FN, and the posterior ampullary nerve in the singular canal. The number of psammoma bodies increases with age. We believe that psammoma bodies are a normal finding of aging in the IAC. The compression of the FN by psammoma bodies in the labyrinthine portion of the facial canal and the distribution of numerous psammoma bodies surrounding the posterior ampullary nerve in the narrow singular canal raise the questions of the involvement of psammoma bodies in the FN and in vestibular dysfunction and the presence of psammoma bodies in the subarachnoid space. PMID- 10526852 TI - Expression of NPY Y1 and CGRP1 receptors in human nasal mucosa: implications in allergic rhinitis. AB - Numerous nerve fibers containing neuropeptide Y (NPY) and calcitonin gene-related peptide (CGRP) have been found in the human nasal mucosa by means of immunocytochemistry. We detected NPY Y1 and CGRP1 receptors at the same location using reverse transcriptase-polymerase chain reaction. The possibility of local release in connection with specific target receptors suggests a role for endogenous NPY and CGRP in the regulation of vascular tone, glandular secretion, and epithelial functions. PMID- 10526853 TI - Mineralocorticoid receptors in the mammalian olfactory mucosa. AB - Mineralocorticoid hormones regulate secretion and absorption in a wide variety of epithelial tissues, although specific mechanisms in the olfactory mucosa are currently unknown. Utilizing reverse transcription-polymerase chain reaction (RT PCR) analysis, we have demonstrated the expression of mineralocorticoid (type I) receptor messenger RNA in the rodent olfactory mucosa. Amplification products of predicted size were obtained with nucleotide sequences corresponding to respective mineralocorticoid receptor (MR) kidney transcripts. Immunocytochemistry, using an antibody with known specificity for MRs, was then utilized in order to localize the cellular site(s) of MR protein expression in the olfactory mucosa. The highest levels of MR immunoreactivity were localized to the supranuclear region of sustentacular cells, as well as the acinar cells of the Bowman's glands. The respiratory regions of the nasal cavity were devoid of appreciable MR immunoreactivity. This study demonstrates both MR transcript and protein expression in the olfactory mucosa. We hypothesize that the mineralocorticoid hormones may have a role in modulation of olfactory secretion and/or sensory transduction in the peripheral olfactory system. PMID- 10526854 TI - Comparison of upper esophageal sphincter opening in healthy asymptomatic young and elderly volunteers. AB - Deglutitive upper esophageal sphincter opening (UES) in the elderly has been incompletely studied. Our aim was to determine in the elderly the temporal and dimensional characteristics of deglutitive UES opening; anterior and superior hyoid and laryngeal excursions as measures of distracting forces imparted on the UES; and hypopharyngeal intrabolus pressure (IBP). Fourteen healthy elderly and 14 healthy young volunteers were studied by concurrent videofluoroscopy and hypopharyngeal manometry during swallowing of 5- and 10-mL barium boluses. The anteroposterior UES diameter, as well as the anterior hyoid bone and laryngeal excursion, was significantly smaller in the elderly compared to the young (p < .05) for 5-mL barium boluses, but not for 10-mL boluses. The lateral diameter of UES opening was similar between groups for all boluses. The IBP for 5- and 10-mL swallows in the elderly was significantly higher than that in the young (p < .05). We conclude that anteroposterior deglutitive UES opening and hyoid bone and thyroid cartilage anterior excursion are reduced in the elderly. These changes are associated with increased IBP, suggesting a higher pharyngeal outflow resistance in the elderly compared to the young. PMID- 10526855 TI - Psychosocial adjustment after laryngeal cancer surgery. AB - The objective of the study was to assess the psychosocial adjustment of 111 patients, and 87 partners, after laryngeal cancer surgery. Sixty-nine patients were grouped as having had radical surgery (total or near-total laryngectomy), and 30 as having had functional surgery (horizontal supraglottic laryngectomy or cordectomy). The Psychosocial Adjustment to Illness Scale Self Report questionnaire was used as the primary outcome. No significant differences were found between groups when global adjustment or domain adjustment was compared. Patient and partner responses were almost identical. Work and family relationships were the domains with poorest adjustment for both patients and partners. Information about treatment expectations was negatively rated by functional surgery patients, especially by those who underwent a cordectomy. We conclude that patient perspectives should be considered and consulted to 1) evaluate patient opinion about treatment results, 2) identify patients with special support needs, and 3) inform patients according to other patients' priorities, based on their experience. PMID- 10526856 TI - Management of N0 neck in T1-T2 unilateral supraglottic cancer. AB - Early-stage supraglottic cancers (stage I and II) are treated with several different programs. Previously reported data have led us to design a therapeutic protocol in treatment of patients with early-stage squamous cell carcinoma of the supraglottic larynx. From 1991 to 1996, 39 patients with unilateral supraglottic carcinoma were treated according to this protocol. All patients underwent unilateral functional neck dissection and resection of the primary carcinoma in an en bloc fashion. Histopathologic studies showed that 9 (23%) of them had positive nodes, and they received planned adjuvant radiotherapy. None of the 30 patients with histopathologically NO necks received either adjuvant irradiation or contralateral neck dissection. The mean follow-up period was 34 months. All patients are alive, and none have developed any recurrence in either dissected or undissected sides of the neck. This treatment policy seems satisfactory and will avoid unnecessary therapeutic interventions. Routine bilateral neck dissection may not be necessary in the surgical treatment of all supraglottic laryngeal cancers. PMID- 10526857 TI - Experimental reinnervation of a strap muscle with a few roots of the phrenic nerve in rabbits. AB - In order to compare application of the roots of the phrenic nerve to the ansa hypoglossi for laryngeal muscle neurotization, 1 or more roots from the phrenic nerve were implanted into the right sternothyroid (RST) muscle of rabbits (n = 36). Controls were intact animals (in which RST innervation is provided by the ansa; n = 6) and denervated ones (n = 6). At 66 +/- 2 days (mean +/- SE) after neurotization, during quiet breathing, inspiratory electromyographic activity and isometric contraction force were observed in all reinnervated RST muscles (n = 24). During maximal inspiratory effort, electromyographic activity and force increased. In animals reinnervated by the C4 root alone, forces (46.22 +/- 7.8 g) were significantly higher than in intact animals (10.83 +/- 5.0 g). Retrograde labeling proved the phrenic origin of the neurotization. Electromyography of the diaphragm was recorded. We conclude that in rabbits, neurotization of a strap muscle by 1 or 2 roots of the phrenic nerve allows inspiratory contraction, even during quiet breathing. Such inspiratory activity is not observed in sternothyroid muscles of intact animals innervated by the ansa hypoglossi. PMID- 10526858 TI - Meningoencephalic herniation into the middle ear. PMID- 10526859 TI - George E. Shambaugh, Jr, 1903-1999. PMID- 10526860 TI - Objective calibration of bone conductors using otoacoustic emissions. AB - OBJECTIVE: To demonstrate a technique for objectively calibrating bone conductors on an individual basis using distortion product otoacoustic emissions (DPOAEs). DESIGN: Individual calibrations were obtained using DPOAEs recorded from a single ear of 21 normally hearing adults. Validity and robustness of the technique were investigated through subjective phase cancellation measurements and sensitivity analysis. RESULTS: Calibrations obtained using the DPOAE method were well supported by phase cancellation results. Intersession repeatability was good, and manipulation of the DPOAE data showed that the calculated calibration is relatively insensitive to small variations of emission magnitude. Bilateral stimulation through bone conduction did not display an apparent effect on emission magnitude in a single individual. CONCLUSION: Bone conductors can be accurately calibrated on an individual basis with good repeatability using DPOAEs. The technique is robust and offers an objective, noninvasive calibration method for research and specialized clinical applications. No training and only passive cooperation are required, making the procedure ideal for special groups such as children. A number of limitations will reduce the clinical utility of this technique. Important audiometric frequencies below 1 kHz cannot be tested because of noise, because individuals with significant hearing loss are unlikely to produce sufficient DPOAEs, and because commercial bone conductors typically have poor high-frequency response above 4 kHz. PMID- 10526861 TI - Effects of chronic tobramycin treatment on distortion product otoacoustic emissions. AB - OBJECTIVE: To investigate the effects of chronic tobramycin treatment on distortion product otoacoustic emission (DPOAE) latencies and response growth detection thresholds in human subjects to determine the sensitivity of these DPOAE features to ototoxic damage. DESIGN: Six groups of children in two different age ranges were tested: three groups in the 7 to 14 yr age range, i.e., six children with normal hearing, four cystic fibrosis (CF) patients who received no aminoglycosides, and eight CF patients who received low- to moderate cumulative doses of tobramycin (< 1250 mg/kg) for respiratory infections; and three groups of five subjects each in the 15 to 23 yr age range, i.e., the healthy group and the CF groups that received low- (< 285 mg/kg) and moderate (1000 to 2000 mg/kg) cumulative drug dosages. The aggregate drug dosages compiled longitudinally over the past 5 yr were used to group the drug-treated CF patients. All subjects showed normal audiometric profiles (< or = 25 dB HL in the conventional frequency region and age-appropriate thresholds as described by Osterhammel and Osterhammel [1979] in the high-frequency region) and DP-grams (absolute DPOAE and noise amplitudes being consistent with the normative data obtained with the CUBeDIS system at this institution). RESULTS: Even though the audiometric profiles and DP-grams of all drug-treated CF groups were identical to their healthy counterparts, the DPOAE latencies and growth function thresholds showed significant changes. Whereas low and low-to-moderate doses of tobramycin were related to DPOAE latency prolongations, higher cumulative drug doses of 1000 to 2000 mg/kg produced significant reductions in DPOAE latencies. Response growth detection thresholds at high frequencies showed significant elevations in all CF patient groups treated with tobramycin, regardless of drug dosages, as compared with the control subjects. CONCLUSIONS: DPOAE amplitudes may not reflect the earliest changes produced by chronic aminoglycoside treatment, suggesting that cochlear ototoxicity may be more effectively monitored through the assessment of latencies and response growth detection thresholds. These findings pertain at least to the early stages of ototoxicity development, specifically during chronic tobramycin treatment. In light of the small sample size, however, these outcomes must be considered as tentative. PMID- 10526862 TI - Tympanic pressure gradients and otoacoustic emissions. AB - OBJECTIVE: The purpose of this study was to investigate the immediate effects of tympanic over- and under-pressure, induced by variations in ambient pressure on click-evoked otoacoustic emissions (CEOAEs) in healthy individuals. It was of particular interest to elucidate whether changes in the CEOAE response in both spectral and time domains could be attributed not only to tympanic, but also to cochlear influence. DESIGN: Nine healthy subjects with normal hearing and middle ear pressure were exposed to ambient pressure changes in a pressure chamber. The pressure was progressively changed in 100 daPa steps to accomplish an increase and a decrease in tympanic pressure. Pressure equilibration of the middle ear was avoided. The relative tympanic over- and under-pressure (+/- 320daPa) was monitored by tympanometry and the CEOAEs recorded at every step of tympanic pressure change. RESULTS: There was a statistically significant reduction of the otoacoustic emission (OAE) response levels and reproducibility already at 100 daPa of ambient pressure change. The OAE response was progressively reduced by increased pressure gradients. The CEOAEs recorded during progressive tympanic over- and under-pressure also had increasingly shorter latencies. These changes of the OAE response characteristics were most pronounced in the 750 to 3000 Hz frequency bands. CONCLUSIONS: The progressive attenuation of the OAE response and the concomitant shortening of the OAE response latencies were observed during a combination of altered middle and inner ear pressure. Although the middle and inner ear influence cannot be separated we suggest, based on our findings, that the shortening of latencies may partly be caused by inner ear pressure changes and stiffening of the labyrinthine membranes. Further studies are needed to more specifically clarify the relative contribution of the tympanic and labyrinthine influence, respectively, for the various aspects of pressure influence on the OAE response. PMID- 10526863 TI - Morphological changes in serial auditory brain stem responses in 24 to 32 weeks' gestational age infants during the first week of life. AB - OBJECTIVE: The purpose of this investigation was to describe and quantify the sequential morphological changes in the auditory brain stem response (ABR) during the first postnatal week of life in very premature infants < or = 32 wk gestational age. These normative data could be useful in predicting neurological outcome in infants with perinatal risk factors. DESIGN: Sequential ABRs were recorded on a total of 135 infants on 5 out of the first 7 days of life. For analysis, data were grouped by gestational age in 2 wk intervals. In addition, a unique system was devised to categorize waveform response types in premature infants: type 1, a response with normal morphology and replicable waves III and V; type 2, a replicable response with either a wave III or wave V; type 3, a replicable response with neither a wave III or wave V; type 4, a response with no replicable waveform. RESULTS: The frequency of detection of waves improves over the first week of life with the detectability of waves III and V being more frequent than wave I at all gestational ages. There was a gradual improvement in response types in infants > 26 wk with the greatest improvement occurring during the 28 to 29 wk gestation. ABRs were predominantly types 3 and 4 at 24 to 25 wk, type 3 at 26 to 27 wk, type 2 at 28 to 29 wk, and types 1 and 2 at 30 to 31 wk. Absolute wave latencies and interwave latencies also progressively decreased during the first postnatal week. In some infants there was a transient increase in latencies or worsening of response type on the second to third test day. CONCLUSIONS: There is progressive improvement in frequency of detection of waves I, III, and V with increasing gestational age. Response types gradually mature over the first postnatal week, particularly in premature infants 28 to 32 wk gestational age. PMID- 10526864 TI - Cochlear implants in young children: the relationship between speech perception and speech intelligibility. AB - OBJECTIVE: To determine the relationship between measures of speech perception and speech production after cochlear implantation of young children with profound congenital and prelingual deafness. DESIGN: A prospective study was undertaken on a consecutive group of children with profound deafness. There were 126 children at the preimplantation interval and 71, 50, 26, and 20 children, respectively, at the 2, 3, 4, and 5 yr follow-up after implantation. Speech perception and speech intelligibility were assessed using hierarchical rating scales. Spearman rank correlation coefficients were used to determine the statistical correlations. All patients were either congenitally deaf or deafened before the age of 3 yr and were implanted before age 7 yr. The patients all received the Nucleus multichannel cochlear implant system with the most appropriate speech encoding strategy. RESULTS: Speech intelligibility at 5 yr was strongly correlated with speech perception at the 2, 3, 4, and 5 yr intervals after implantation (Spearman coefficients 0.77, 0.81, 0.58, 0.58; p < or = 0.01). Speech intelligibility at the 2, 3, and 4 yr intervals also correlated in a similar manner with earlier speech perception abilities (p < or = 0.01). CONCLUSIONS: The results suggest that speech intelligibility between 2 and 5 yr after implantation in young children with congenital and prelingual profound deafness can be predicted by measures of earlier speech perception. PMID- 10526865 TI - Altered phonatory physiology with short-term deactivation of children's cochlear implants. AB - OBJECTIVES: The purposes of this investigation were 1) to determine whether short term auditory deprivation results in systematic phonatory changes for prelingually deafened children who use cochlear implants (CIs) and 2) to determine whether such changes are similar to those that have been reported for postlingually deafened adults. DESIGN: Participants were two 6-yr-old children with CIs. Both children had been prelingually deafened, had good to excellent speech production and speech perception skills, and had been using their CIs for 2.5 yr. A single-subject design was used. Intraoral air pressure (Po), phonatory air flow (Vl), electroglottograph (EGG) cycle width, fundamental frequency (F0), and intensity were measured during syllable production over two baseline days and three experimental days. Data were collected twice on each baseline day while the children wore their CIs, with a 1 hr break between data collection sessions. On experimental days, data were collected while the children wore their CIs (ON condition) and after their CIs had been removed for 1 hr (OFF condition). RESULTS: Both children demonstrated highly variable phonatory behaviors in baseline. The child with the more proficient speech production and perception skills showed consistent and significant reductions in Po, F0, and intensity in the OFF condition. These findings were dissimilar to those that occurred with repeated testing in the baseline condition and so were attributed to the sudden loss of auditory feedback. The other child showed a consistent and significant increase in mean Vl in the OFF condition. However, this child exhibited a similar finding with repeated testing in the baseline condition. Therefore, increased Vl in the OFF condition may have represented a practice effect. She also showed a small and consistent decrease in F0 in the OFF condition when F0 was derived from acoustic data, but this effect was not reliable in another data set when F0 was derived from the EGG signal. Our results with prelingually deafened children were inconsistent with reports of increased intensity and F0 in the absence of auditory feedback for postlingually deafened adults with CIs. CONCLUSIONS: Some prelingually deafened children who are successful CI users appear to use auditory feedback to self-monitor phonation. We suggest that the participant in our investigation who showed systematic phonatory changes in response to diminished auditory feedback was using auditory feedback primarily to stabilize her phonatory behaviors. She may not have had adequate experience with auditory feedback or adequate flexibility in her use of feedback mechanisms to implement the phonatory compensations that late-deafened adults use when auditory feedback suddenly is diminished. We further suggest that the phonatory changes that she exhibited during short-term auditory deprivation reflected disruption of her typical speaking strategies or apprehension about speaking when her ability to self-monitor auditorily was compromised. PMID- 10526866 TI - The role of sulbactam-ampicillin/sultamicillin in mixed infections. PMID- 10526867 TI - Beta-lactamases and beta-lactamase inhibitors. AB - Penicillin, the first of the beta-lactam antibiotics, was introduced into medical practice in the 1940s. Since then, a large number of different beta-lactams, including penicillins, cephalosporins, monobactams, and carbapenems, have been developed, all of which are structurally related through the presence of a core beta-lactam ring. Resistance to beta-lactam antibiotics among target pathogens developed early in the history of their use. Of the mechanisms of resistance, the most widespread and most important is the destruction of the beta-lactam ring, which is mediated by beta-lactamases. The fact that these resistance enzymes may be coded on plasmids means that they are mobile within a bacterial community, and that they have spread widely. Resistance to beta-lactams mediated by beta lactamases can be overcome successfully with the use of beta-lactamase inhibitors. The combination of beta-lactams with beta-lactamase inhibitors restores the activity of the beta-lactams, allowing their continued clinical use. The development of beta-lactamase inhibitors allows clinicians to rely on the well-tolerated, clinically effective beta-lactam antibiotics to treat a variety of bacterial infections. PMID- 10526868 TI - In vitro efficacy of beta-lactam/beta-lactamase inhibitor combinations against bacteria involved in mixed infections. AB - Mixed infections are usually caused by a relatively limited range of bacteria, with the anaerobes and opportunistic pathogens contributing to their severity. In order to make the best therapeutic choice for a patient with a life-threatening infection, which is probably of mixed etiology, clinicians must be aware of the organisms that are likely to be involved, and the fact that most of them will produce beta-lactamase. Of the options available for empiric therapy, the beta lactam/beta-lactamase inhibitor combinations represent a good choice. Their antibacterial spectra include both aerobic and anaerobic pathogens. Five combinations are available in clinical practice: ampicillin-sulbactam, piperacillin-tazobactam, ticarcillin-clavulanic acid, amoxicillin-clavulanic acid, and cefoperazone-sulbactam. More strains of clinically important anaerobic bacteria are susceptible to ampicillin-sulbactam than to either piperacillin tazobactam or ticarcillin-clavulanic acid, which are also available widely and suitable for more life-threatening infections. In addition, sulbactam itself has the highest intrinsic activity of the beta-lactamase inhibitors against the opportunistic pathogen, Acinetobacter baumannii. Thus, ampicillin-sulbactam could be considered a drug of choice for the empirical treatment of mixed infections where there is a reasonable possibility of the presence of A. baumannii. PMID- 10526869 TI - The role of beta-lactam/beta-lactamase inhibitors in the management of mixed infections. AB - Microbiological studies show that the in vitro antimicrobial activity of sulbactam-ampicillin encompasses not only gram-positive and gram-negative aerobes, but also anaerobes. Such a broad spectrum of activity suggests its suitability as monotherapy for the empiric management of polymicrobial infections. Typical mixed infections, which are frequently life-threatening, include those occurring in the abdomen or pelvis, diabetic foot infections, and brain abscess. Numerous comparative clinical studies have revealed the clinical and bacteriological efficacy of sulbactam-ampicillin to be comparable to that of imipenem cilastatin and the second-generation cephalosporins cefoxitin and cefotetan. In addition, other studies have demonstrated that sulbactam-ampicillin monotherapy is cost-beneficial. A reduction in the duration of hospitalization, the lack of potentially toxic side-effects, and lower drug costs associated with monotherapy all contribute to the cost-effectiveness of sulbactam-ampicillin. PMID- 10526870 TI - Antibiotic prophylaxis in head and neck oncologic surgery: the role of gram negative coverage. AB - Many studies have elucidated the risk factors associated with peri-operative infection following head and neck cancer surgery (HNS), the implications of infection for total treatment cost, and the clinical benefits of successful antimicrobial prophylaxis. The most appropriate antibiotic use is achieved by focusing on patients with clean, contaminated wounds. Thereafter, the usefulness of an antibiotic agent depends on its antimicrobial spectrum, tolerability profile, and cost. Successful antimicrobial prophylaxis requires antimicrobial activity against gram-positive, gram-negative, and anaerobic organisms. The beta lactam/beta-lactamase inhibitor combination, sulbactam-ampicillin, has just such an antimicrobial spectrum. A double-blind, randomized clinical trial, involving patients undergoing HNS, recorded a lower post-operative infection rate among patients receiving peri-operative sulbactam-ampicillin 0.5 g/1.0 g i.v. q6h compared with those receiving clindamycin 600 mg i.v. q6h (13.3 vs. 27.1%; P = 0.02). Culture of strains from infected individuals indicated a significantly lower proportion of gram-negative organisms for sulbactam-ampicillin than for clindamycin (32 vs. 81%; P < 0.05). There was a significant difference in the median duration of surgery between infected and non-infected individuals (8.5 vs. 5.9 h; P < 0.0001). These data support the use of sulbactam-ampicillin to reduce the incidence of post-operative infection following HNS. PMID- 10526871 TI - Trends in self-reported use of mammograms (1989-1997) and Papanicolaou tests (1991-1997)--Behavioral Risk Factor Surveillance System. AB - PROBLEM/CONDITION: In 1999, an estimated 175,000 women will be diagnosed with breast cancer, and 43,300 will die from the disease. In the same year, an estimated 12,800 women will be diagnosed with invasive cervical cancer, and 4,800 will die from it. Early detection and timely treatment of breast cancer and cervical dysplasia can alter the progress of and reduce mortality from these diseases. REPORTING PERIOD COVERED: 1989-1997 for breast cancer screening and 1991-1997 for cervical cancer screening. DESCRIPTION OF SYSTEM: The Behavioral Risk Factor Surveillance System is a state-based telephone survey of the civilian, noninstitutionalized adult population (i.e., persons aged > or =18 years). In this report, responses for women aged > or =40 years are included for measures of breast cancer screening, and responses for women aged > or =18 years with an intact uterine cervix are included for measures of cervical cancer screening. RESULTS: The percentage of women aged > or =40 years who reported ever participating in breast cancer screening and the proportion who had participated within the previous 2 years increased during 1989-1997. The percentage of women aged > or =18 years who reported ever participating in cervical cancer screening and the proportion who had participated within the previous 2 years were stable during 1991-1997. For both types of screening, substantially fewer women had received screening within the previous 2 years than had ever been screened. INTERPRETATION: These findings may indicate that some women who participate in initial screening do not seek further screening. ACTIONS TAKEN: Initiatives to encourage women to receive initial screening should continue, but additional initiatives specifically aimed at promoting rescreening should be developed. Continued surveillance of the percentage of women who receive regular screening will help public health officials evaluate breast and cervical cancer prevention programs. PMID- 10526872 TI - Breast-conserving surgery for invasive cancer: a principle based on segmental anatomy. AB - As the incidence of breast cancer increases in Japan, breast-conserving surgery becomes an important issue in the light of quality of life. We have demonstrated by 3-D reconstruction studies that ductal carcinoma in situ (DCIS) originates from the terminal duct-lobular unit (TDLU). Normal mammary epithelium anatomically located in the TDLU was shown to be biologically associated with cancerous change, particularly in specimens from patients who subsequently developed invasive carcinoma. Atypical ductal hyperplasia as well as DCIS expressed breast cancer associated antigen, providing further biological evidence that the atypical lesion at the TDLU are premalignant. Intraductal spread of carcinoma was defined as "DCIS was present clearly extending beyond the TDLU, or present prominently within the large ducts," and was classified into 3 grades according to the distribution of carcinoma in the duct-lobular system. We have developed a breast-conserving surgery consisting of quadrantectomy and regional lymph node dissection and immediate volume replacement using lateral tissue-flap (LTF). The quadrantectomy was employed on the basis of segmental anatomy of the duct-lobular system in which breast carcinoma originates. Fairly good cosmetic outcome as well as local control were obtained in the patients who underwent the immediate volume replacement using LTF. It must be emphasized that the quadrantectomy is a radical procedure in the sense that it aims at removal of all the carcinoma cells of the primary tumor. PMID- 10526873 TI - Effect of mandibular advancement splint on psycho-intellectual derangements in patients with sleep apnea syndrome. AB - The mandibular advancement splint (MAS) was recently introduced for the management of sleep apnea syndrome (SAS), although its effects on psycho intellectual functions have not been elucidated yet. We examined psycho intellectual function before and after treatment with MAS in patients with SAS. Twenty patients with SAS underwent psycho-intellectual function testing before and after treatment with MAS for 3 to 4 weeks. The apnea index significantly decreased from 19.0+/-15.6 to 2.4+/-1.9. The state anxiety score significantly decreased from 44.6+/-12.1 to 33.7+/-11.1, the trait anxiety score significantly decreased from 46.2+/-13.4 to 37.6+/-13.8, and the depression scale score significantly decreased from 39.2+/-11.0 to 30.8+/-9.9 with MAS treatment. By the Cornell Medical Index and the Yatabe-Guilford test, the patients became less neurotic and less eccentric after treatment. By the Uchida-Kraepelin psychodiagnostic test, calculation ability significantly increased from 1247.4+/ 402.1 to 1950.2+/-651.9. We conclude that MAS treatment reduces apneic episodes and improves psycho-intellectual derangements in patients with SAS. PMID- 10526874 TI - Creatinine at the evaluation of urinary 1-methyladenosine and pseudouridine excretion. AB - The elevation of urinary modified nucleosides levels in urine is found in patients with cancers. In the present study, we have tested 616 urine samples randomly collected from non-malignant cases. Thirty-two percent (194/616) and 11% (68/616) had elevated levels of 1-methyladenosine and pseudouridine, respectively (They are designated as false-positive cases). To elucidate the cause on non specific elevation of the nucleosides, the correlation between creatinine excretion level and urinary nucleosides levels were determined. The result revealed that false-positive cases were frequently detected in patients with lower creatinine excretion levels. The mean creatinine levels of false-positive cases were significantly lower than those of negative cases. From these results, the false-positive of urinary 1-methyladenosine and pseudouridine might be due to the low creatinine excretion mainly caused by the renal dysfunction. Creatinine excretion in each individual should be taken into consideration in case of determining urinary modified nucleosides. PMID- 10526875 TI - Determining factors of mortality in the elderly with hip fractures. AB - We conducted a retrospective study of the influence of various factors on the mortality of 114 patients with hip fractures. The mortality rate one year after surgery was 18%, which was 2.5 times larger than that of the general population. It was related to age, ECG abnormality, and post-operative complications. PMID- 10526876 TI - Correlation between magnetic resonance imaging and clinical profiles of periventricular leukomalacia. AB - Magnetic resonance imaging (MRI) findings of 70 children with periventricular leukomalacia (PVL), examined between 1 year 2 months and 8 years of age (mean: 2 years 4 months of age), were analysed. Neurological assessments were made between 1 year 3 months and 15 years (mean: 4 years 9 months). The possible correlations between MRI findings and clinical profiles of PVL were investigated using three parameters of the MRI findings. The grade of ventriculomegaly correlated well with the severity of cerebral palsy (CP) but not with the severity of mental impairment. The grade of reduction of periventricular white matter correlated well with the severity of CP and mental impairment, and is the most reliable parameter for neurological prognosis. The degree of periventricular hyperintensity on T2-weighted images did not correlate well with severity of CP, but correlated to some degree with mental impairment. There was a significantly lower degree of periventricular hyperintensity in children at less than 28 weeks of gestation than at 28 or more weeks of gestation, but no significant difference in other parameters. The periventricular hyperintensity should be evaluated in view of the gestational age. PMID- 10526877 TI - Escape of parasympathetic vasodilatation from sympathetic attenuation in oro facial areas in the cat. AB - We examined the effects of concurrent repetitive stimulation of the cervical sympathetic trunk (CST) on the parasympathetically mediated reflex blood flow increase in the orofacial area of cats. In urethane plus alpha-chloralose anaesthetized cats, parasympathetic reflex vasodilatation in the ipsilateral lower lip was elicited by electrical stimulation of the central cut end of the lingual nerve (LN). This blood flow increase was attenuated in a frequency dependent manner when CST was stimulated concurrently at 0.5-10 Hz for 10 minutes. When we applied repeated LN stimulation (using identical parameters, each time) at intervals during a 30-minutes period of 10 Hz CST stimulation, the attenuation of the blood flow increase gradually weakened in a time-dependent manner even though the direct vasoconstrictor effect of CST stimulation showed no such decline. PMID- 10526878 TI - The effects of tone exposure on the inner ear functions in the guinea pig: impact tone vs. steady state tone. AB - The damage-risk criterion (DRC) for hearing supposes that sound exposure with equal energy implies equal risk for noise-induced hearing loss (NIHL). We measured cochlear microphonics (CM), compound action potential (CAP), endocochlear potential (EP) and K+ ion concentration in the scala media, to see if the same level of Leq24h (impact tone and steady state tone) induced the same physiological changes in the inner ear function or not. Regarding the equal energy principle (EEP), we also examined if the EEP is appropriate or not at exposure of moderate level tone. We also checked how the time interval between impact tones affects or not the inner ear functions at the same Leq24h tone exposure. Therefore we used exposure at 1 pulse/second or 1 pulse/3 seconds and steady state tone exposure at Leq24h=90, 85 and 80 dB. The results are the following. Both steady state and impact tone exposure causes change of the electrophysiological data. First, CM maximum output voltage after exposure to impact tone of 115 dB (Leq24h=90 dB) was lower than after exposure to a 8 kHz steady state tone of 90 dB. CAP threshold (below 10 microV) obtained after the 115 and 110 dB exposure of impact tone were 5-10 dB higher than that of steady state tone of 90 dB. The negative EP induced by impact tone exposures showed the same tendency as the CM experiments. Having more frequent pulses (1 pulse/second vs. to 1 pulse/3 seconds) showed more inhibition. The K+ concentration time course remained similar to the control when the Leq24h was low (80 dB). Impact tone exposure induced stronger effects to the inner ear at exposure of moderate level tone than that of steady state tone of Leq24h. PMID- 10526879 TI - Noncardiogenic pulmonary edema as the chief manifestation of a pheochromocytoma: a case report of MEN 2A with pedigree analysis of the RET proto-oncogene. AB - Pheochromocytomas are rare neoplasias of the adrenal medulla which generally present with paroxysmal or sustained hypertension. Cardiogenic pulmonary edema is a common feature of these tumors, but few cases have been described with noncardiogenic pulmonary edema. We report a pheochromocytoma with the principle manifestation of noncardiogenic pulmonary edema and characterize a genetic lesion associated with the disorder. A 30-year-old man was admitted with abdominal pain and breathlessness. x-Ray examination of the chest revealed a massive, diffuse infiltration of the left lung without cardiomegaly. No paroxysmal blood pressure fluctuations or heart failure were evident during the entire course, and the infiltrate and dyspnea resolved in three days without inotropic or diuretic agents. Serum norepinephrine and epinephrine levels were elevated twenty and fifty times above normal, respectively. The patient was ultimately diagnosed with multiple endocrine neoplasia type 2A (MEN 2A). Mutations in the RET proto oncogene have been described recently in patients with MEN 2A. Mutation analysis of selected RET exonic sequences identified a germline mutation at codon 634 in exon 11 of the RET proto-oncogene. The mutation introduces a transition encoding a non-conservative substitution from TGC (Cys) to CGC (Arg) and creates a novel restriction site recognized by HhaI. We further screened for this mutation among four of the proband's relatives by HhaI restriction analysis. One asymptomatic family member was identified who subsequently elected prophylactic total thyroid removal. Histological examination of this specimen confirmed the presence of medullary thyroid carcinoma. PMID- 10526880 TI - Synthesis of a template-associated peptide designed as a transmembrane ion channel former. AB - We describe the design and the Fmoc/tBu solid phase synthesis of a 20 residue long peptide containing five regularly distributed lysines. Cyclization of this peptide was achieved using BOP as coupling agent. After side-chain deprotection, all the basic residues were iodoacetylated and then allowed to react either with a C-terminal free COOH peptide or with peptides bearing a cysteamide group. The final pentameric templates were identified by mass and amino acid analysis which gave data compatible with the expected values. PMID- 10526881 TI - Synthesis of thiazole, imidazole and oxazole containing amino acids for peptide backbone modification. AB - Novel 5-ring heterocyclic building blocks are synthesized. These can be incorporated into analogs of peptide antibiotics such as microcin B17, which is a potent DNA-gyrase inhibitor that exhibits eight thiazole and oxazole moieties. In particular, the syntheses of imidazole and bisoxazole amino acids as novel peptidomimetics are reported, this includes a new procedure for the oxidative conversion of the intermediates oxazoline, imidazoline as well as oxazole oxazoline into the corresponding heteroaromatic compounds. A mixture of 1,8 diazabicyclo-[5.4.0.]-undec-7-ene carbon tetrachloride/acetonitrile and pyridine proved to be a very effective and mild agent. PMID- 10526882 TI - Pi-allyloxymethyl protection of histidine. AB - Experimental details are presented for the introduction and application of pi allyloxymethyl protection for histidine side-chains. PMID- 10526883 TI - The synthesis of 'difficult' peptides using 2-hydroxy-4-methoxybenzyl or pseudoproline amino acid building blocks: a comparative study. AB - A comparative study has been undertaken between Hmb-protected amino acid and pseudoproline building block analogues for use in the solid phase synthesis of 'difficult' peptides. Both of these derivatives act by blocking inter- and intramolecular hydrogen bonding, which has been shown to be a major cause of poor synthesis/quality/efficiency. While the two were shown to result in substantial improvements in the purity of crude peptides, pseudoproline incorporation was found to be superior to Hmb backbone protection. This was due to slow and incomplete coupling of the amino acid immediately following the Hmb amino acid. PMID- 10526884 TI - Solution structure of human beta-endorphin in helicogenic solvents: an NMR study. AB - Beta-endorphin is the largest natural opioid peptide. The knowledge of its bioactive conformation might be very important for the indirect mapping of the active site of opioid receptors. We have studied beta-endorphin in a variety of solution conditions with the goal of testing the intrinsic tendency of its sequence to assume a regular fold. We ran NMR experiments in water, dimethylsulfoxide and aqueous mixtures of methanol, ethylene glycol, trifluoroethanol, hexafluoracetone trihydrate and dimethylsulfoxide. The solvent in which the peptide is more ordered is the hexafluoracetone trihydrate/water mixture. The helical structure detected for beta-endorphin in this mixture at 300 K extends for the greater part of its address domain, hinting at a possible mechanism of interaction with opioid receptors: a two-point attachment involving an interaction of the helical part of the address domain (PLVTLFKNAIIKNAY) with one of the transmembrane helices and a classical interaction of the message domain (YGGF) with the receptor subsite common to all opioid receptors. PMID- 10526885 TI - Trends in European computational neuroscience. AB - Understanding the complex structure and function of the central nervous system requires the integration of many scientific disciplines and descriptive levels, and the development and application of adequate research tools. The fast growing field of computational neuroscience plays an important role within this integrative effort. In Europe, an active community is emerging that investigates structure-function relationships in the nervous system at subcellular, cellular, network, and behavioural levels. An increasing number of courses and conferences are offering opportunities for development and exchange of ideas between scientists interested in computational neuroscience. Thus, it is timely to present a spectrum of European activities in this field and to foster the communication between scientists across disciplines towards joint efforts. PMID- 10526886 TI - Contrast adaptation and infomax in visual cortical neurons. AB - In the primary visual cortex (V1) the contrast response function of many neurons saturates at high contrast and adapts depending on the visual stimulus. We propose that both effects - contrast saturation and adaptation - can be explained by a fast and a slow component in the synaptic dynamics. In our model the saturation is an effect of fast synaptic depression with a recovery time constant of about 200 ms. Fast synaptic depression leads to a contrast response function with a high gain for only a limited range of contrast values. Furthermore, we propose that slow adaptation of the transmitter release probability at the geniculocortical synapses is the underlying neural mechanism that accounts for contrast adaptation on a time scale of about 7 sec. For the functional role of contrast adaptation we make the hypothesis that it serves to achieve the best visual cortical representation of the geniculate input. This representation should maximize the mutual information between the cortical activity and the geniculocortical input by increasing the release probability in a low contrast environment. We derive an adaptation rule for the transmitter release probability based on this infomax principle. We show that changes in the transmitter release probability may compensate for changes in the variance of the geniculate inputs - an essential requirement for contrast adaptation. Also, we suggest that increasing the release probability in a low contrast environment is beneficial for signal extraction, because neurons remain sensitive only to an increase in the presynaptic activity if it is synchronous and, therefore, likely to be stimulus related. Our hypotheses are tested in numerical simulations of a network of integrate-and-fire neurons for one column of V1 using fast synaptic depression and slow synaptic adaptation. The simulations show that changing the synaptic release probability of the geniculocortical synapses is a better model for contrast adaptation than the adaptation of the synaptic weights: only in the case of changing the transmitter release probability does our model reproduce the experimental finding that the average membrane potential (DC component) adapts much more strongly than the stimulus modulated component (F1 component). In the case of changing the synaptic weights, however, the average membrane potential (DC) as well as the stimulus modulated component (F1 component) would adapt. Furthermore, changing the release probability at the recurrent cortical synapses cannot account for contrast adaptation, but could be responsible for establishing oscillatory activity often observed in recordings from visual cortical cells. PMID- 10526887 TI - Single cell and population activities in cortical-like systems. AB - Dynamics of single cells and large cell populations are the subject of investigation by using differently detailed models. Multicompartmental modeling techniques are used to systematically investigate the location-dependent effects of GABA-ergic inhibition on the firing patterns of hippocampal pyramidal cells. Appearance of stochastic resonance in a model of mitral and granule cells of the olfactory bulb is demonstrated by using a single-compartmental model approach. Spatial propagation of synchronized activities in hippocampal slices are studied by a model of large neural populations. PMID- 10526888 TI - Computational models of predictive and memory-related functions of the hippocampus. AB - We discuss the role of the hippocampus in information processing in the brain and hypothesise that the hippocampus monitors the stability of sensory cues it receives from the external world, using the current context to predict the next sensory event in the episodic sequence by learning from experience, and memorising these sequences of sensory events. Two computational models are presented here. The predictive theory and model are closely related to experimental evidence and use dynamic synapses with an asymmetric learning rule to develop predictive neural activity of a leaky integrate-and-fire model of a pyramidal CA3 cell. The oscillatory model of the hippocampus for memorising sequences of sensory events is developed as a chain of interacting neural oscillators forced by oscillatory inputs from the entorhinal cortex and from the medial septum. PMID- 10526889 TI - Using realistic models to study synaptic integration in cerebellar Purkinje cells. AB - This review presents an approach to modeling which we call "experiments in computo". The use of realistic models makes it possible to generate new predictions that can be confirmed experimentally. Several examples are given of how this approach has improved our understanding of synaptic integration by the Purkinje cell active dendrite. The computer model was constructed to replicate neuronal behavior which has no direct relevance to synaptic integration: it was tuned to reproduce the response of Purkinje cells to current injection in vitro, which consists of a high frequency, regular rhythm of somatic Na+ spikes, interrupted by spontaneous dendritic Ca2+ spikes. The in vivo firing behavior of Purkinje cells is quite different as it consists of highly irregular simple spike firing only, without spontaneous dendritic Ca2+ spikes. The computer model predicted-that the Purkinje cell needs to receive a continuous background inhibitory synaptic drive in addition to the excitation by parallel fibers to obtain this typical in vivo firing. This prediction was confirmed by blocking inhibition during in vivo intracellular recordings. More recently, we demonstrated that the net inhibitory drive to the Purkinje cell dendrite has to be larger than the excitatory synaptic drive. Inhibition hyperpolarizes the dendrite compared to the soma, making it act as a current sink during most of the spiking cycle. These predictions have been confirmed with the dynamic clamp method in the cerebellar slice preparation. Synchronous focal excitatory input by parallel fiber leads in the model to activation of voltage-gated Ca2+ channels which amplify the somatic response by a variable amount. The variability of this graded amplification is due both to position of the input, effectively canceling the cable attenuation, and to the effect of preceding background input. Differences between the model and experimental results in this aspect can be explained by the relative hyperpolarized state of Purkinje cells in the in vitro experimental preparation. These studies led to a new theory about the function of long-term depression in the cerebellum which can explain recent experimental results. In conclusion, our modeling approach generated predictions which contradicted prevalent ideas on how the cerebellum, or neurons in general, works and led to experiments which otherwise would not have been carried out. PMID- 10526890 TI - Towards an integration of biochemical and biophysical models of neuronal information processing: a case study in the nigro-striatal system. AB - The experimental and theoretical study of intracellular biochemical signaling mechanisms lags considerably behind our understanding of electrical processes of neuronal membranes. Both signaling processes, however, are extensively intertwined and can be analyzed and modeled using formally similar mathematical tools. With the nigro-striatal system as an example, we review various formal approaches to describe metabotropic signaling in dopamine- and calcium-dependent pathways and their interactions with electrical membrane processes. These demonstrate the feasibility of synthetic modeling and afford insights into a variety of specific signaling mechanisms. Extending and linking hitherto isolated models has the potential to transcend descriptive levels and to provide a fuller understanding of the molecular basis of macroscopic information processing in the central nervous system. PMID- 10526891 TI - Interaction of cortex and hippocampus in a model of amnesia and semantic dementia. AB - We describe a systems-level computational model, called TraceLink, that can explain the major characteristics of the neuropsychology of amnesia and of semantic dementia, a recently discovered syndrome in which there is a progressive loss of semantic memory. It also approximates the normal forgetting curve and presents an explanation of why spaced learning is more efficient than massed learning. A central assumption is that consolidation of memory takes place, probably during dream sleep. The model consists of three systems: trace system (certain parts of the neocortex), link system (includes the hippocampus), and the modulatory system (includes certain basal forebrain nuclei). Lesioning each of these causes a characteristic form of amnesia or semantic dementia. Lesioning the modulatory system causes anterograde amnesia only. Lesioning the link system causes a correlated degree of retrograde and anterograde amnesia. Retrograde amnesia shows the characteristic Ribot curve with relative sparing of remote memories but loss of recent ones. Lesioning the trace system causes semantic dementia. We also review the main sources of constraints for the model and discuss its status and function as well as its falsifiability. PMID- 10526892 TI - Properties of the evoked spatio-temporal electrical activity in neuronal assemblies. AB - Properties of neural computation were studied in two types of neuronal networks: isolated leech ganglia and neuronal cultures of dissociated cortical neurons from neonatal rats. With appropriate experimental set-ups it was possible to obtain a precise description of the spread of excitation induced by specific inputs. The evoked spatio-temporal electrical activity was characterized by large variability and the electrical activity of neurons activated by the same stimulation was found to be statistically independent to a high degree. The variability presumably originates from basic properties of synaptic transmission, which is stochastic in nature. As a consequence, the large variability of the evoked spatio-temporal electrical activity appears to be a general property of neural computation and a typical feature of neuronal assemblies. It is shown, however, that the observed statistical independence of co-activated neurons may be used to reduce the effects of variability by appropriately averaging or pooling the electrical activity. PMID- 10526893 TI - What can robots tell us about brains? A synthetic approach towards the study of learning and problem solving. AB - This paper argues for the development of synthetic approaches towards the study of brain and behavior as a complement to the more traditional empirical mode of research. As an example we present our own work on learning and problem solving which relates to the behavioral paradigms of classical and operant conditioning. We define the concept of learning in the context of behavior and lay out the basic methodological requirements a model needs to satisfy, which includes evaluations using robots. In addition, we define a number of design principles neuronal models should obey to be considered relevant. We present in detail the construction of a neural model of short- and long-term memory which can be applied to an artificial behaving system. The presented model (DAC4) provides a novel self-consistent implementation of these processes, which satisfies our principles. This model will be interpreted towards the present understanding of the neuronal substrate of memory. PMID- 10526894 TI - Nitric oxide and the renin-angiotensin system. Is there a physiological interplay between the systems? AB - Opposed actions for nitric oxide (NO) and angiotensin II (Ang II) in vascular contraction and vascular smooth muscle cell proliferation and apoptosis are well documented. In addition, various experimental approaches have shown that NO negatively modulates the renin-angiotensin system by inhibiting angiotensin converting enzyme (ACE) activity and down-regulating AT1 receptors. On the other hand, Ang II and Ang-(1-7) positively stimulate NO synthesis and release. In this review, we analyse the data suggesting a mutual regulation between the renin angiotensin and the nitric oxide-generating systems, and we propose a homeostatic interplay between both factors aimed at regulating cardiovascular function. PMID- 10526895 TI - Blood pressure in childhood and adolescence: the Italian normal standards. Study Group on Hypertension' of the Italian Society of Pediatrics'. AB - OBJECTIVES: To develop a national standard level of blood pressure (BP) for Italian children on the basis of a large sample of the population. DESIGN: We analyzed data available from 21 Italian studies conducted according to the recommendations of the American Task Force between 1988 and 1994. Percentile curves of systolic and diastolic BP were constructed by fitting a third-order polynomial model of BP on age and height using multiple regression analysis. PARTICIPANTS: BP was measured in 11 519 healthy individuals (6258 boys and 5261 girls) aged 5-17 years in various locations throughout Italy. All measurements were performed at school. RESULTS: Percentile curves (5th, 10th, 25th, 50th, 75th, 90th and 95th) of systolic and diastolic BP are reported by age and by height for males and females. CONCLUSIONS: With respect to the American standards, the levels in Italy for the 90th and 95th percentiles were 3-8 mmHg higher for systolic and diastolic BP in both sexes between 5 and 12 years of age, and 2-3 mmHg higher in older males. With respect to Northern Europe, in the lower ages, levels in Italy were quite similar, although slightly higher, whereas in late adolescence, the Northern European levels were much higher, especially in males, with differences of 4-5 mmHg for the mean values and 8-12 mmHg for the 95th percentile. PMID- 10526896 TI - Blood pressure and risk of myocardial infarction in elderly men and women: the Rotterdam study. AB - OBJECTIVE: To study the association between blood pressure and risk of myocardial infarction in elderly subjects. DESIGN: Prospective cohort study. SETTING: The Rotterdam Study, a Dutch population-based study. PARTICIPANTS: 6004 men and women aged > or = 55 years. MAIN OUTCOME MEASURES: Fatal or non-fatal myocardial infarction (n = 190) during a 4-year follow-up. RESULTS: After excluding participants using blood pressure-lowering medication and participants with a history of myocardial infarction, increasing levels of systolic blood pressure (SBP) were associated with increasing risk of first myocardial infarction (P for trend < 0.0001). The relative risk (RR) for an SBP of 160 mmHg or higher was 5.7 (95% confidence interval (CI) 1.9-17.1) compared with an SBP below 120 mmHg. Increasing diastolic blood pressure (DBP) was also associated with increasing risk of first myocardial infarction, with the RR reaching 2.5 (95% CI 1.4-4.5) in subjects with values of 80-90 mmHg compared with values below 70 mmHg (P for trend < 0.05). Analyses in subjects aged 70 years and over showed that the positive associations between SBP and DBP and risk of first myocardial infarction remained at older age. CONCLUSION: These findings in a relatively healthy cohort of elderly subjects do not provide evidence for a J- or U-shaped relation between SBP and DBP and risk of first myocardial infarction. They suggest that the risk of first myocardial infarction increases with increasing level of systolic and diastolic blood pressure and that this relationship persists into older age. PMID- 10526897 TI - A randomized trial on effects of hormone therapy on ambulatory blood pressure and lipoprotein levels in women with coronary artery disease. AB - OBJECTIVE: To investigate 1-year effects of hormone replacement therapy (HRT) on ambulatory blood pressure (ABP) and lipoprotein levels in postmenopausal women with coronary artery disease (CAD). METHODS: Sixty patients at a mean age (+/- SD) of 59 +/- 7 years were randomized into three groups: conjugated equine oestrogens (CEE) 0.625 mg daily (n = 20), 50 microg 17beta-oestradiol transdermally (TTSE) per 24 h (n = 20) or placebo (n = 20) for 18 days, then combined with medroxyprogesterone acetate 5 mg for 10 days. Each cycle of 28 days was repeated for one year. RESULTS: Night-time systolic ABP had decreased by 9.6% (P= 0.0075) in 15 of 18 women in the CEE group and by 22% in 12 of 13 women (P = 0.0034) in the placebo group after 1 year. In the CEE group, a 4.6% rise in daytime systolic ABP (P< 0.05) and a 4.2% rise in night-time systolic ABP (P< 0.05) appeared from baseline to 6 months in 13 of 18 women. In the CEE group (14 women analysed), high-density lipoprotein levels showed a 15.8% increase (P= 0.0018) in 13 women, low-density lipoprotein levels a 15.2% decrease (P= 0.0129) in 12 women and total cholesterol levels a 7.5% decrease (P = 0.057) in 11 women after 1 year. Triglyceride levels showed no changes. In the TTSE group and in the placebo group, with 12 and 13 women analysed respectively, no significant changes appeared. CONCLUSIONS: One year of HRT in patients with CAD does not influence ABP. Oral HRT induces beneficial effects on lipoprotein levels. PMID- 10526898 TI - A pressure-time index' for assessing the severity of essential hypertension. AB - OBJECTIVE: A new derivative of 24 h ambulatory blood pressure monitoring (ABPM) is introduced and its association with left ventricular mass index (LVMI) in essential hypertension is examined. PATIENT: population One hundred and fifty three previously untreated essential hypertension patients. METHODS: Patients underwent casual blood pressure (BP) readings, 24 h ABPM and left ventricular echocardiographic assessment The following 24 h awake and sleep ABP variables were calculated: mean systolic and diastolic BP, systolic and diastolic BP loads (percentage of systolic readings > 140/120 mmHg (day/ night) and diastolic readings > 90/80 mmHg (day/night)), standard deviation of systolic and diastolic ABP and nocturnal fall of systolic BP, as well as the integrated areas under the ABP curve. The area under the BP curve divided in horizontal slices was accurately modelled by a sigmoid curve. The parameters controlling the shape of the curve and in particular that regarding its 'slope' is hereafter called the 'pressure-time index'. RESULTS: 'Systolic pressure-time index 24 h' (SPTI24) is related to left ventricular mass index (multivariate analysis, P= 0.008). Using either partial correlation coefficients or a multivariate analysis, SPTI24 is related to left ventricular mass index, independently of age, casual blood pressure, mean systolic and diastolic ABP, systolic and diastolic BP loads, BP variability (standard deviation (SD), nocturnal fall of systolic BP) and integrated area under the curve (multivariate analysis, P= 0.004). CONCLUSIONS: In essential hypertension, the SPTI24 is related to LVMI independently of age, casual blood pressure, integrated area under the curve or any other derivative of 24 h ABPM, and might be used to assess the extent of hypertensive load. PMID- 10526899 TI - The effects of short-term passive smoke exposure on endothelium-dependent and independent vasodilation. AB - OBJECTIVE: There is limited information on the mechanisms mediating the deleterious effects of passive smoke exposure. Cross-sectional studies indicate that nonsmokers exposed chronically to passive smoke have impaired endothelium mediated vasodilation. We tested the hypothesis that acute exposure to sidestream (passive) smoke impairs endothelium-dependent vasodilation in healthy nonsmokers. METHODS AND RESULTS: We studied 12 healthy nonsmokers (aged 27 +/- 5 years, nine men and three women). We obtained measurements of blood pressure, heart rate, and bilateral forearm blood flow (FBF). Each individual was studied twice, following a randomized, placebo-controlled design. The effects of passive smoke were studied on one day and the effects of vehicle (room air) on a separate day. Acetylcholine (ACh) and sodium nitroprusside (SNP) were infused into the left brachial artery before and after 15 min of exposure to either passive smoke (carbon monoxide concentration between 20 and 40 p.p.m.) or vehicle (room air). The order of ACh and SNP, and smoke or vehicle, was randomized between individuals. Smoke exposure increased carboxyhemoglobin from 0.5 +/- 0.1 % to 0.8 +/- 0.1% (P= 0.002). Neither passive smoke nor vehicle changed baseline measurements of heart rate, blood pressure and forearm vascular resistance (FVR). The vasodilatory responses to ACh and SNP were very similar, both before and after exposure to passive smoke and before and after vehicle. CONCLUSION: Our data demonstrate that acute exposure to passive smoke does not alter either endothelium-dependent or independent vasodilatory responses in healthy nonsmoking individuals. Hence, impaired endothelial vasodilatory responses in nonsmokers chronically exposed to passive smoke most likely reflect chronic functional and/or structural changes in responses to cigarette smoke, rather than the acute effects of cigarette smoke toxicity on endothelial function. PMID- 10526900 TI - Differences in aortic response to vasoactive stimuli in Japanese and Lyon rats. The role of hypertension. AB - OBJECTIVE: We have previously shown that conduit arteries of normotensive (WKY) and hypertensive (SHR) Japanese rats differ from normotensive (LN) and hypertensive (LH) Lyon rats in terms of lower aortic thickness and higher collagen III content, whereas differences in vasoactive properties are unknown. METHODS: Aortic rings with (E+) and without (E-) endothelium were studied under resting and noradrenaline-stimulated conditions in the presence of N(omega)-nitro L-arginine (L-NNA) alone or in association with indomethacin, bosentan and/or BQ123. RESULTS: Under resting conditions, aortas of normotensive and hypertensive Japanese rats differed from Lyon rats by higher developed tension in the presence of L-NNA and endothelium. In the absence of endothelium, normotensives differed from hypertensives in terms of stronger developed tensions in the presence of L NNA in the two strains. Addition of indomethacin to L-NNA induced relaxation in E+ SHR and E- WKY and contraction in E-LH. By contrast, tensions were unchanged after addition of bosentan and BQ123. Under stimulated conditions, tensions were equally increased by L-NNA in E+ and unchanged in E- both in Japanese and Lyon rats whether they were normotensive or hypertensive, and indomethacin (but not bosentan) elicited higher response in Lyon than in Japanese rats in E+ and E- aorta. CONCLUSION: Under NO synthase inhibition, the vasoactive properties of Japanese and Lyon aorta differ in the presence of a cyclo-oxygenase blocker but not endothelin blockers. These results indicate that the aorta vasorelaxant tone is associated to prostanoid regulation in Lyon but not in Japanese rats. This observation appears dependent on the genetic and/or environmental background linked to the origin and not the presence of hypertension. PMID- 10526901 TI - Apoptosis in the muscular arteries from young spontaneously hypertensive rats. AB - OBJECTIVE: The purpose of this study was to test the hypothesis that a different incidence of apoptosis occurs in the mesenteric arteries of the spontaneously hypertensive rat (SHR) compared with its normotensive control the Wistar-Kyoto rat (WKY) at 1-2 weeks of age. DESIGN: We examined the incidence of apoptotic cells in the blood vessel wall of muscular arteries from the SHR and WKY at 1-2 weeks of age using two techniques of apoptosis measurement DNA laddering and 3' OH end labelling. We also measured the volume of the blood vessel wall components and lumen sizes with the confocal microscope to determine whether a differential incidence of apoptosis occurred between the two rat strains. METHODS: We used phenol/chloroform extraction to isolate genomic DNA and assess DNA fragmentation, with gel electrophoresis to determine DNA laddering, and 3'-OH end labelling, where the enzyme terminal deoxynucleotidyl transferase catalyses the addition of fluorescein-conjugated nucleotides to the cut ends of DNA, to detect in situ DNA fragmentation. The volume per unit length of the blood vessel structural components was measured by optical sectioning with the confocal microscope. RESULTS: We found that the SHR had a significantly decreased incidence of cellular apoptosis over WKY. This was true for both the electrophoretic method where SHR had significantly less fragmented DNA (molecular size < 600 bp) than WKY (P= 0.01), and for the microscopic method where SHR had fewer labelled cells in both the adventitia (P= 0.01) and the media (P= 0.0001) layers of large mesenteric arteries. The volumes of the adventitia, media and lumen in the large mesenteric arteries were similar between the two strains at this age. CONCLUSION: These findings suggest that a differential incidence of cellular apoptosis at the age of 1 -2 weeks may be responsible for the larger media volume found in older SHR and thus contributes to the development of hypertension in these animals. PMID- 10526902 TI - Low dose of eicosapentaenoic acid inhibits the exaggerated growth of vascular smooth muscle cells from spontaneously hypertensive rats through suppression of transforming growth factor-beta. AB - OBJECTIVE: To evaluate effects of eicosapentaenoic acid (EPA), an n-3 polyunsaturated fatty acid, on the exaggerated growth of vascular smooth muscle cells (VSMC) from spontaneously hypertensive rats (SHR). DESIGN: Cultured VSMC were prepared by an explant method from thoracic aortas in 8-week-old male Wistar Kyoto (WKY)/Izumo rats and SHR/Izumo. Effects of EPA on basal DNA synthesis, expression of growth factors and cyclin-dependent kinase 2 (cdk2) activity were examined in VSMC from WKY rats and SHR. METHODS: The cell cycles were synchronized with serum deprivation, then DNA synthesis in VSMC was measured by [3H]-thymidine incorporation. Fatty acid composition of the phospholipid fraction in VSMC was measured by gas chromatography. Expression of platelet-derived growth factor (PDGF) A-chain, transforming growth factor (TGF)-beta1 and basic fibroblast growth factor (bFGF) mRNAs was evaluated by reverse-transcription and polymerase chain reaction analysis. Cdk2 activity was determined by autoradiography after polyacrylamide gel electrophoresis of VSMC extracts that had been immunoprecipitated with anti-cdk2 antibody and protein A sepharose, and then incubated with 32P-ATP and histone H1. RESULTS: High concentrations (40 and 80 micromol/I) of EPA significantly inhibited basal DNA synthesis in VSMC from both rat strains. Low dose (20 micromol/l) of EPA significantly inhibited basal DNA synthesis in VSMC from SHR, whereas the same dose of EPA stimulated DNA synthesis in VSMC from WKY rats. In analysis of fatty acid composition, low dose of EPA was considerably incorporated in VSMC. Low dose of EPA significantly inhibited angiotensin II- and phorbol ester milisterol-stimulated DNA synthesis in VSMC from both rat strains, whereas EPA did not affect PDGF-AA-stimulated DNA synthesis in VSMC from either rat strain. Low dose of other polyunsaturated fatty acids such as docosahexaenoic acid, arachidonic acid and linoleic acid did not significantly affect basal DNA synthesis in VSMC from either strain. Low dose of EPA significantly inhibited expression of TGF-beta1 mRNA in VSMC from SHR, whereas EPA did not affect expression of PDGF A-chain and bFGF mRNAs in VSMC from SHR. Cdk2 activity in VSMC from SHR was higher than that from WKY rats. Low dose of EPA inhibited cdk2 activity in VSMC from SHR, whereas it stimulated the activity in VSMC from WKY rats. CONCLUSION: Low dose of EPA exerted specific inhibition of the exaggerated growth of VSMC from SHR through the suppression of TGF-beta. PMID- 10526904 TI - Linkage but lack of association for blood pressure and the alpha-adducin locus in normotensive twins. AB - BACKGROUND: alpha-adducin is a cytoskeletal protein involved with sodium-pump activity in the renal tubule. The alpha-adducin gene locus has been linked to hypertension and a polymorphism identified which is associated with hypertension; however, the role of the alpha-adducin gene locus in normal blood pressure regulation is not defined. We performed a combined linkage and association study in normotensive monozygotic (MZ) and dizygotic (DZ) twins and their parents to address this issue. METHODS: We studied 126 MZ and 70 DZ twin pairs and parents of DZ twins. Blood pressure values and responses to a cold pressor test were obtained. Cardiac dimensions were measured echocardiographically. Three microsatellites adjacent to the alpha-adducin gene were studied as well as the 460 Trp mutation in the alpha-adducin gene. RESULTS: We obtained strong evidence for linkage (P< 0.001) between the alpha-adducin gene locus and systolic blood pressure. However, we were not able to associate the 460 Trp mutation with higher blood pressures, cold pressor responses or cardiac dimensions. CONCLUSIONS: The alpha-adducin gene locus is relevant to blood pressure regulation in normal subjects. Failure to find an association between higher blood pressures and the 460 Trp mutation suggests that this mutation may become important only when hypertension is triggered, or that other variations in alpha-adducin are present which have not yet been discovered. PMID- 10526903 TI - Linkage analysis of endothelial nitric oxide synthase gene with human blood pressure. AB - OBJECTIVE: Endothelial nitric oxide exerts important effects on the regulation of vascular tone and structure. Variants of the endothelial nitric oxide synthase gene (eNOS) have been associated with hypertension and myocardial infarction, although some reports have shown negative linkage with hypertension. To examine whether the region encoding the eNOS gene is linked with physiological blood pressure variation, we undertook a linkage analysis of this region in the general population. DESIGN: In healthy volunteer families, we used two independent quantitative linkage analyses to examine the relationship between genotypes and phenotypes, with both parametric and non-parametric and single-locus and multi point methods. METHODS: We selected 260 families comprising mother and father (aged 40-70 years) and two natural offspring (aged 18-30 years) from the Victorian Family Heart Study. After standardized measurement of clinical data and extraction of DNA, all family members were genotyped at five microsatellite loci including the CA repeat in the eNOS gene by a PCR method. The quantitative linkage analyses were conducted according to two different analysis programs, the Genetic Analysis System (GAS) and the MAPMAKER/SIBS. RESULTS: With both linkage analyses, we found no linkage between any of the loci on chromosome 7q35-36 and the phenotypes systolic and diastolic blood pressure, mean arterial pressure, pulse pressure, pulse rate, weight, height and body mass index. CONCLUSION: Based on these results, we conclude that in this population the eNOS gene is not linked to the physiological variation of blood pressure and other related phenotypes. PMID- 10526905 TI - Delayed recovery of hypertension after single dose losartan in angiotensin II infused conscious rats. AB - OBJECTIVE: In a conscious unrestrained rat model, it takes approximately 1 week for angiotensin II to increase blood pressure to maximum levels. We investigated the time required for hypertension to fully recover after acute angiotensin II receptor blockade in this angiotensin II dependent hypertensive model. DESIGN: Conscious unrestrained rats (n = 8) infused with 10 ng/kg per min angiotensin II for 21 days received losartan (10 mg/kg) on day 17 of angiotensin II infusion. Mean arterial pressure (MAP) and heart rate were monitored continuously. The acute pressor response to 50 ng/kg per min angiotensin II was monitored for 2 h on days 15, 17, 18, 19 and 20 of angiotensin II infusion. Plasma renin concentration (PRC) was measured daily. RESULTS: Angiotensin II increased MAP acutely by 26 +/- 2 mmHg and by a further 23 +/- 4 mmHg between days 4 and 8. Losartan acutely reduced MAP by 75 +/- 2 mmHg; 24 h later MAP had partially recovered but remained suppressed by 47 +/- 3 mmHg. MAP had not fully recovered 4 days later. Some 2 h after losartan, the acute pressor response to angiotensin II had fallen from 24 +/- 2 mmHg to zero. This recovered to 13 +/- 5 and 28 +/- 2 mmHg 24 and 48 h post losartan. After losartan PRC rose from 0.1 +/- 0.05 to above 1 ng/ml per h for less than 24 h. CONCLUSION: A single dose of losartan reverses both the fast and slow pressor effects of continuous angiotensin II infusions. While losartan is metabolized, the fast vasoconstrictor effect recovers quickly but the slow pressor effect takes almost a week to build up again to maximum levels. Since the slow pressor effect is mediated via the AT1 receptor, any means of blocking the renin-angiotensin system is likely to keep blood pressure below maximum hypertensive levels for several days after the drug has disappeared from the circulation. PMID- 10526906 TI - Fasting insulin and leptin serum levels are associated with systolic blood pressure independent of percentage body fat and body mass index. AB - OBJECTIVE: To examine the relationship between leptin and insulin serum levels and systolic and diastolic blood pressure in young men. SETTING: Kobe University of Mercantile Marine, Kobe, Japan. PARTICIPANTS: One hundred and ninety-eight male students aged 18-20 years (comprising 100% of those eligible). DESIGN AND MEASUREMENTS: A cross-sectional survey of a sample of male college students was performed, with measurements to include anthropometry, blood pressure and blood tests after overnight fasting. RESULTS: Compared with 90 men with an optimal blood pressure, 56 men with high-normal and high blood pressure had an increase in body mass index (23.7 +/- 5.2 versus 20.4 +/- 2.2 kg/m2), percentage body fat (21.7 +/- 8.0 versus 16.3 +/- 4.2%) and serum leptin (3.7 +/- 4.7 versus 1.5 +/- 0.8 ng/ml). In addition, they had greater serum insulin (59 +/- 31 versus 43 +/- 12 pmol/l) despite there being no differences in plasma glucose, resulting in a reduction of the ratio of glucose to insulin (x 10(6)) (107 +/- 43 versus 126 +/ , which is an estimate of insulin sensitivity in a nondiabetic population. Furthermore, the 56 men had higher serum triglyceride levels, although there was no difference in low density lipoprotein-cholesterol and high density lipoprotein cholesterol between men with optimal and high-normal plus high blood pressure. Similar differences were found between men in a top versus low tertile of systolic and diastolic blood pressure. In multiple regression analysis, both log leptin and log insulin emerged as determinants for systolic blood pressure independent of body mass index and percentage body fat, but an association with diastolic blood pressure was only shown for log leptin. CONCLUSION: Hyperleptinemia and hyperinsulinemia may be regulators of arterial pressure, independent of body mass index or percentage body fat. PMID- 10526907 TI - 825T allele of the G-protein beta3 subunit gene (GNB3) is associated with impaired left ventricular diastolic filling in essential hypertension. AB - OBJECTIVE: Recently, a novel C825T polymorphism in the gene (GNB3) encoding for the G-protein beta3 subunit was identified. The 825T allele is associated with the generation of a novel splice variant, enhanced intracellular signal transduction, and arterial hypertension. In this study, we investigated the impact of the 825T allele on left ventricular structure and function in mild to moderate essential hypertensive subjects. METHODS: In 34 white patients with established mild to moderate essential hypertension (World Health Organization stage I or II, mean age 52 +/- 9 years) genotype analysis of GNB3 C825T polymorphism, insertion/deletion polymorphism of the ACE gene and 1166 A/C polymorphism of the AT1 receptor gene was performed. In each patient, 24 h ambulatory blood pressure measurement (SpaceLabs 90207) and two-dimensional guided M-mode echocardiography combined with Doppler sonography were performed. RESULTS: In our homogenous study group, the GNB3 825T allele was not associated with casual and 24 h ambulatory blood pressure (CC versus TC/TT: 144 +/- 13/92 +/ 8 versus 151 +/- 14/97 +/- 7 and 143 +/- 11/92 +/- 7 versus 150 +/- 16/ 96 +/- 9 mmHg, respectively) or parameters of left ventricular structure (relative wall thickness: CC versus TC/TT, 0.48 +/- 0.1 versus 0.46 +/- 0.1; left ventricular mass: CC versus TC/TT, 281 +/- 65 versus 299 +/- 80 g). However, transmitral flow variables reflecting left ventricular diastolic filling were impaired in patients expressing the TC/TT genotype (ratio of peak late (A) to early (E) velocities: CC versus TC/TT, 0.95 +/- 0.24 versus 1.2 +/- 0.26, P< 0.02; velocity time integrals A/E: CC versus TC/TT, 0.57 +/- 0.16 versus 0.76 +/- 0.23, P< 0.01) while all co variables such as age, body mass index, ambulatory blood pressure, heart rate and end-diastolic volume were similar between the two groups. If patients were stratified according to the I/D polymorphism of the ACE gene and the A1166C polymorphism of the AT1 receptor gene, no differences in blood pressure, left ventricular structure or systolic and diastolic function of the left ventricle were found between different genotypes. CONCLUSION: The GNB3 825T allele was associated with impaired left ventricular diastolic filling in hypertensive subjects in this study. Since alterations in left ventricular filling have been identified as an early marker of hypertensive heart disease, the GNB3 C825T polymorphism may influence cardiac adaptation to increased afterload. PMID- 10526908 TI - Detection of hypertensive patients at risk for paroxysmal atrial fibrillation during sinus rhythm by computer-assisted P wave analysis. AB - OBJECTIVE AND METHODS: To determine whether hypertensive patients at risk for paroxysmal atrial fibrillation (AF) could be detected while in sinus rhythm, a computer-based 12-lead surface electrocardiogram was recorded in 50 hypertensive patients with history of paroxysmal AF (group A) and in 60 hypertensive patients without history of AF (group B). The maximum P-wave duration (P(maximum)), the minimum P-wave duration (P(minimum)), P-wave dispersion (Pdispersion = Pmaximum Pminimum), adjusted P-wave dispersion (APdispersion = Pdispersion/square root of the number of measurable leads), mean P-wave duration (mean P) and the standard deviation of the P-wave duration in all measured leads (SDP) were calculated. RESULTS: Pdispersion, APdispersion and SDP were significantly higher in group A than in group B (Pdispersion, 52 +/- 19 versus 41 +/- 15 ms, P< 0.001; APdispersion, 15.2 +/- 5.5 versus 11.9 +/- 4.6 ms, P< 0.001; SDP, 16 +/- 5 versus 13 +/- 5 ms, P < 0.001). P(minimum), mean P and left ventricle ejection fraction (LVEF) were significantly lower in group A than in group B (Pminimum, 79 +/- 18 versus 91 +/- 13 ms, P < 0.001; mean P, 108 +/- 18 versus 116 +/- 13 ms, P= 0.005; LVEF, 64 +/- 5 versus 69 +/- 8%, P< 0.001). Pminimum, Pdispersion, mean P, SDP, APdispersion and LVEF were found to be significant univariate predictors of paroxysmal AF, whereas only Pminimum (P< 0.001) remained a significant independent predictor of paroxysmal AF in the multivariate analysis. CONCLUSION: Hypertensive patients at risk for paroxysmal AF could be detected while in sinus rhythm by computer-assisted electrocardiographic P-wave analysis. PMID- 10526910 TI - Vasoactive effects of potassium in kidneys of hypertensive rats fed a high potassium diet. AB - DESIGN AND METHODS: Levels of dietary and serum potassium are thought to correlate inversely with vascular resistance and blood pressure. This study examined renal vascular resistance in perfused rat kidneys partially preconstricted with 10 micromol/ phenylephrine, quantifying changes in the resistance when levels of potassium in the perfusate ([K+]o) were varied between 2 and 80 mmol/l. RESULTS: In kidneys from 17-week-old Wistar-Kyoto rats (WKY strain) fed a normal diet (American Institute of Nutrition AIN-76 diet), the resistance decreased when [K+]o was raised from 4 to 6-20 mmol/l, whereas resistance increased when [K+]o was either lowered to 2 mmol/l or raised above 25 mmol/l. The vasodilation that occurred at 13 mmol/l [K+]o was blocked by 100 micromol/l BaCl2 and 10 micromol/l ouabain in an additive manner, suggesting that both the inward rectifier K+ channel and the Na-K-ATPase underlie the dilation. Kidneys from spontaneously hypertensive rats (SHR strain) fed the AIN-76 diet displayed modestly enhanced vasodilations and vasoconstrictions as compared to WKY. A high-potassium diet (AIN-76 supplemented with 3.5% potassium citrate, provided for 8 weeks) led to exaggerated vasoconstrictive effects of [K+]o, and modestly enhanced vasodilations, in WKY rats. In contrast, the diet led to attenuated vasoconstrictions, and dramatically enhanced vasodilations, in the SHR strain. The diet did not affect the blood pressure increase or weight gain of either strain. CONCLUSIONS: Changes in the responsiveness of blood vessels to extracellular potassium might underlie some beneficial effects of high-potassium diets in hypertensive individuals. PMID- 10526909 TI - The impact of different echocardiographic diagnostic criteria on the prevalence of left ventricular hypertrophy in essential hypertension: the VITAE study. Ventriculo Izquierdo Tension Arterial Espana. AB - BACKGROUND: The prevalence of echocardiographic left ventricular hypertrophy in essential hypertension ranges from 12 to 96% depending on the threshold values used to define it, and on the selection bias. OBJECTIVE: To estimate the prevalence of echocardiographic left ventricular hypertrophy by different criteria in essential hypertensives seen in primary care centres. METHODS: Cross sectional study in a population-based sample of 946 essential hypertensives randomly selected in 39 primary care centres across Spain. Echocardiographic studies were performed in reference hospitals by trained observers (concordance Cohen kappa index > 0.7) and analysed by a single observer. RESULTS: Prevalence of left ventricular hypertrophy ranged from 59.2% [95% confidence interval (CI) 56.1 -62.3] by Framingham criteria to 72.7% (95% CI 69.9-75.6) using the criteria of De Simone et al. (J Am Coll Cardiol 1995; 25: 1056-1062). Prevalence was higher in males by the Cornell-Penn criteria, but higher in females when using Framingham or De Simone et al. criteria. Eccentric hypertrophy was more frequent (51.3-54.1%) independently of the criteria used, particularly when adjusting wall thickness-ratio for age (56.2-58.9%). Concentric remodelling was present in 6.5 11.4% and only 20.8-29.7% of patients had no evidence of left ventricular structural alterations. Factors independently associated with left ventricular hypertrophy in the logistic regression analysis were age, gender, systolic blood pressure, pulse pressure and body mass index. CONCLUSION: Prevalence of echo left ventricular structural alterations among essential hypertensives seen in primary care centres in Spain ranged from 70.3 to 79.2% depending on the threshold values used. Left ventricular hypertrophy ranged from 59.2 to 72.7% and age-adjusted concentric remodelling ranged from 6.5 to 11.4% depending on the criteria used. Only one-quarter of hypertensive patients were free from morphological alterations. PMID- 10526911 TI - Mibefradil prevents L-NAME-exacerbated nephrosclerosis in spontaneously hypertensive rats. AB - OBJECTIVE: To determine the potential renal protective effects of a novel calcium channel blocker mibefradil in chronic renal failure. METHOD: We compared the long term effects of mibefradil with an angiotensin-converting enzyme inhibitor cilazapril on blood pressure, proteinuria, renal function and histological alterations in N-nitro-L-arginine methylester (L-NAME)-treated spontaneously hypertensive rats (SHR). Three groups of SHR were studied for 45 days: group 1 (n = 14), treated with L-NAME only (50 mg/l in the drinking water); group 2 (n = 15) L-NAME plus co-treatment with mibefradil (30 mg/kg per day); group 3 (n = 15), L NAME plus co-treatment with cilazapril (10 mg/kg per day). RESULTS: Both mibefradil and cilazapril attenuated the increased systolic blood pressure, and prevented the development of proteinuria and the decreased creatinine clearance (Ccr) seen at day 42 in the group treated with L-NAME alone. Notably, mibefradil had similar effects to cilazapril on proteinuria and Ccr, despite a reduced antihypertensive effect All animals receiving mibefradil co-treatment remained alive throughout the experiment, whereas the mortality rate was 43% in SHR treated with L-NAME alone. Both mibefradil and cilazapril completely prevented renal structural damage as assessed by scoring glomerular, tubulo-interstitial and vascular lesions. CONCLUSIONS: Our data show that mibefradil prevented the development of hypertension and proteinuria, renal functional impairment and nephrosclerosis, and also improved animal survival. The renal protective effects of mibefradil were at least equivalent to those of an ACE inhibitor in this animal model of chronic renal failure. PMID- 10526912 TI - Effects of intensified antihypertensive treatment in diabetic nephropathy: mortality and morbidity results of a prospective controlled 10-year study. AB - OBJECTIVE: The aim of this study was to describe the effect of intensified antihypertensive therapy based on a structured teaching and treatment programme on the prognosis of hypertensive type 1 (insulin-dependent) diabetic patients with kidney disease. DESIGN: The study was a controlled, prospective, parallel, 10-year follow-up trial. PATIENTS AND INTERVENTIONS: A sequential sample of 91 hypertensive type 1 diabetic patients with overt diabetic nephropathy was prospectively followed for 10 years. Forty-five patients (intensified antihypertensive therapy; IT group) participated in an intensified antihypertensive therapy programme and 46 patients received routine antihypertensive treatment as provided by family physicians, consultants and local hospitals (routine antihypertensive therapy; RT group). OUTCOME MEASURES: The main endpoint was death; secondary endpoints were renal replacement therapy, blindness and amputation. RESULTS: Blood pressure was reduced in the IT group and increased in the RT group. During the follow-up period, 29 patients died, seven in the IT group and 22 in the RT group. The survival curves were significantly different (P = 0.0008). The main causes of death were cardiac. In a multiple Cox proportional hazards model, allocation to the IT group reduced the mortality risk [relative risk (RR) = 0.213; 95% confidence interval 0.089-0.509, P = 0.00051, while age (P = 0.0039) and mean blood pressure (P= 0.0113) increased this risk. In multiple Cox or multiple logistic regression models, the risks of dialysis (RR = 0.269, 95% confidence interval 0.110-0.656, P = 0.0039), blindness (odds ratio = 0.158, 95% confidence interval 0.037-0.684, P= 0.0136), and amputation (RR = 0.181, 95% confidence interval 0.047-0.703, P= 0.0135) were significantly lower in the IT group compared with the RT group (log rank P = 0.0008). CONCLUSION: We conclude that intensified antihypertensive treatment, based on a hypertension teaching and treatment programme, reduces long-term morbidity and mortality in patients with diabetic nephropathy. PMID- 10526913 TI - Flow dependence of forearm noradrenaline overflow, as assessed during mental stress and sodium nitroprusside infusion. PMID- 10526914 TI - Influence of smoking on baroreceptor function: 24 h measurements. PMID- 10526915 TI - The medical evaluation of the sexually abused child. PMID- 10526916 TI - Diagnostic imaging in obstetrics and gynecology: new developments. AB - This article reviews the most recent studies incorporating diagnostic imaging modalities into gynecologic and obstetric care. It describes studies evaluating the utility of various imaging modalities for the diagnosis of fetal abnormalities, uterine and tubal pathology, and staging of gynecologic malignancies. In addition, the article reviews some experimental studies and their potential clinical applications. PMID- 10526917 TI - Prevention of mother-to-child transmission of HIV-1. AB - Several recently published randomized trials have demonstrated that a substantial proportion of the mother-to-child transmission of HIV-1 can be prevented by antiretroviral therapy late in gestation and at delivery to mother and infant. The cost implications of these findings are considerable for resource-poor settings. Preliminary data also suggest very low rates of transmission among mothers receiving maximally suppressive combination drug regimens. Prophylactic cesarean delivery has also been shown to reduce transmission in women not receiving antiretroviral agents, and may play a role in selected patients. The avoidance of breast feeding with infant formula supplementation is clearly protective against HIV-1 transmission, but may not improve infant survival in some poorer settings because of associated increases in other infectious diseases and malnutrition. PMID- 10526918 TI - Recurrent pregnancy loss: an update. AB - This review highlights recent studies that investigate causes and treatments for recurrent pregnancy loss. Generally the causes of recurrent pregnancy loss are classified as genetic, endocrinologic, anatomic, immunologic, microbiologic, and environmental. The majority of recent work has focused on potential autoimmune and alloimmune causes; however, controversy still exists over appropriate testing and treatment. Reports have investigated the potential associations between autoimmune factors (antithyroid antibodies and antiphospholipid antibodies) and alloimmune factors (natural killer cells, cytotoxic T cells, and embryotoxic factors) and recurrent pregnancy loss. Increasingly, clinical reports are suggesting intravenous immunoglobulin as a potential treatment for these immunologic problems. Several lines of investigation have suggested certain hypercoagulable states as causative of recurrent pregnancy loss. New studies relating recurrent pregnancy loss to endocrinologic aberrations (hyperprolactinemia and hyperandrogenism) as well as social/environmental factors (stress, caffeine use, tobacco use, human immunodeficiency virus, and history of induced abortion) have been made. A summary of proposed evaluation and treatment options is presented. PMID- 10526919 TI - Developmental abnormalities of the female reproductive tract. AB - Developmental abnormalities of the female reproductive tract are a group of heterogeneous anomalies that may also affect other organ systems. Our review outlines the diverse abnormalities, etiologies, modes of diagnosis, and treatment options currently available. PMID- 10526920 TI - New developments in diagnosis and management of adolescents with sexually transmitted disease. AB - The battle against the epidemic of sexually transmitted infections continues but the clinician has new diagnostic and therapeutic weapons. This manuscript reviews new technologies, as well as novel applications of newer methodologies, such as utilization of amplified DNA techniques for urine specimens or patient-collected specimens. Newest treatment recommendations are also reviewed. PMID- 10526922 TI - Common problems in pediatric gynecology: new developments. AB - Physicians, particularly gynecologists, pediatricians and family practitioners, are often called upon to perform a gynecological evaluation of a child. The following article is a review of current developments in the area of pediatric and adolescent gynecology. It outlines the recent clinical information and offers a review of common gynecological disorders among children and adolescent girls. PMID- 10526921 TI - Update on pubertal development. AB - This review provides updated information relating to the timing of pubertal onset from a large study of girls seen in pediatric practices. In addition, new studies investigating the relationship of the hormone leptin to the onset of puberty are discussed, as well as new information on the neuroendocrine control of pubertal regulation. A provocative study documenting poor mental health, more behavior problems, and lower IQ in children with premature adrenarche when compared with controls raises the question of whether psychological stress triggers premature adrenarche or whether the early increase in adrenal hormone secretion causes psychosocial problems. Finally, significant advances in the management of central precocious puberty in girls have been made over the past year. PMID- 10526923 TI - Recent clinical issues related to the use of depot medroxyprogesterone acetate (Depo-Provera). AB - This report critically reviews recent original research articles concerning patient use of depot medroxyprogesterone acetate. Specifically, recent studies have been conducted on the following clinical issues: depression, galactorrhea, weight gain, bone mineral density, epithelial and mucus changes in the lower genital tract, and the acceptability of and continuation rates with the use of depot medroxyprogesterone acetate. PMID- 10526924 TI - A vaginal approach to Burch's retropubic urethropexy. PMID- 10526926 TI - Multichannel urodynamics: ambulatory versus standard urodynamics. AB - Standard urodynamics in the office setting has been the mainstay of urodynamic evaluation. Ambulatory urodynamics has previously been viewed as a research tool only. This article summarizes new studies on ambulatory urodynamics that highlight its potentially more practical and clinical use. PMID- 10526925 TI - The overactive bladder: neuropharmacological basis of clinical management. AB - The overactive bladder continues to pose a major challenge to clinicians treating lower urinary tract disorders, not least because our understanding of the pathogenesis of detrusor overactivity is still relatively limited. However, progress in understanding the basis of the overactive bladder is moving steadily forward, accompanied by a growing number of different forms of treatment. New pharmacological treatments and variations in the mode of delivery of older agents offer hope of efficacy with fewer side-effects. Neuromodulation is also offering a viable alternative to surgery in patients unresponsive to medical treatment. PMID- 10526927 TI - The surgical management of recurrent stress urinary incontinence. AB - Recurrent stress urinary incontinence is a distressing problem. Its causes are multifactorial and the literature continually provides suggestions for changes to the surgical approach. Over 200 surgical procedures exist for the treatment of stress urinary incontinence, and this leaves the practising surgeon with an overwhelming range of choice. This article will review current available techniques for the surgical treatment of recurrent stress urinary incontinence. PMID- 10526928 TI - Urological trauma in gynaecological surgery: diagnosis and management. AB - Lesions of the urinary tract being a rare, but typical, complication of gynaecological surgery need a skilled pelvic surgeon for prevention and, when they occur, knowledge of adequate techniques of repair or palliative solutions until final repair can take place. The increasing incidence of lesions of the urinary tract as a result of endoscopic surgery makes better training mandatory. PMID- 10526929 TI - Vaginal birth and natural outcome. AB - Vaginal delivery is reviewed from a urogynecologic perspective. The birthing process is recognized to be clearly traumatic to the pelvic floor. Directed study is suggested as a means by which predisposing factors for injury may be identified. Ultimately we may be better able to select the optimal mode of delivery for all women. PMID- 10526931 TI - Connective tissue in female urinary incontinence. AB - An effective closure of the female urethra in stress situations is dependent on an integrated action of various anatomical structures connected to the organ. The most important of these structures - from a functional aspect - are the suburethral vaginal wall, the pubourethral ligaments, the pubococcygeus muscles and the paraurethral connective tissues. In all these structures connective tissue is an essential ingredient. Hence, defects in the actual connective tissue - in particular the paraurethral connective tissue that connects the aforementioned structures to each other and to the urethra - will bring about an ineffective urethral closure. Female urinary incontinence may then be caused by defective connective tissue per se and/or by a disconnection of the aforementioned structures, whereby the urethra cannot be 'kinked' - that is, closed off in stress situations. PMID- 10526930 TI - Critical evaluation of electro-stimulation for management of female urinary incontinence. AB - Electro-stimulation has been reported to be effective in the relief of stress and urge urinary incontinence. The rates of cure and improvement brought about by pelvic floor electro-stimulation in patients with urinary incontinence are 30-50% and 60-90%, respectively. In clinical practice, vaginal, anal and surface electrodes are used for external, short-term stimulation, and sacral root stimulation for internal, chronic (long-term) stimulation. The effectiveness of electro-stimulation has been verified in a randomized, placebo-controlled study. However, its superiority over other conservative treatments, such as pelvic floor exercise, has not been confirmed. A long-term effect has also been reported. In conclusion, pelvic floor exercise together with electro-stimulation is the mainstay of conservative management for the treatment of stress incontinence. For urge and mixed stress plus urge incontinence, electro-stimulation may be the first choice alternative treatment to drug therapy. PMID- 10526932 TI - Bibliography. Current world literature. Adult and pediatric gynecology. PMID- 10526933 TI - Bibliography. Current world literature. Urogynecology. PMID- 10526934 TI - Ischemic strokes and the role of echocardiography. PMID- 10526935 TI - Nephrogenic diabetes insipidus: the era of gene therapy. PMID- 10526936 TI - CD30-positive anaplastic large cell lymphoma with HTLV-I proviral integration: a unique histologic subgroup of adult T-cell leukemia/lymphoma. PMID- 10526937 TI - Immunopathology of ANCA-associated vasculitis. AB - During the past few years remarkable progress has been achieved in the understanding of the pathogenic mechanisms leading to vascular inflammation and injury in ANCA-associated vasulitides (AAV): Wegener's granulomatosis (WG), microscopic polyangiitis (MPA) and Churg Strauss syndrome (CSS). In this paper we review the immunopathology of these diseases by describing the role of autoantibodies (ANCA), dysregulation and abnormalities at the cellular level, genetic background and environmental factors that predispose to autoimmune response. PMID- 10526939 TI - Dissociation of vascular tolerance and plasma norepinephrine adjustment during long-term nitrate therapy in human coronary arteries. AB - OBJECTIVE: We examined whether changes in plasma norepinephrine (NE) concentration contribute to the development of nitrate tolerance in human coronary arteries. METHODS: Patients with stable angina were randomized to receiving nitrate (isosorbide dinitrate: ISDN or nitroglycerin: TNG) infusion for 30 minutes (group A), 48 hours (group B), or 78 hours (group C). Coronary diameters were measured angiographically at baseline (CT), during maximum dilation by ISDN (N1), at the end of nitrate infusion (N2) and after additional ISDN (1 mg) injection (N3). RESULTS: In groups A and B, N1, N2, and N3 were greater than CT, and there was no significant difference between N1, N2, and N3 for each group. In group C, N1 and N3 were greater than CT, but there was no difference between CT and N2, in the development of nitrate tolerance. In group A, NE increased significantly during nitrate infusion (304+/-163 vs. 418+/-273 pg/ml). NE did not change in groups B and C. CONCLUSION: The change in NE concentration is not a primary contribution to the development of nitrate tolerance. PMID- 10526938 TI - Transesophageal echocardiographic detection of cardiac sources of embolism in elderly patients with ischemic stroke. AB - OBJECTIVE: The aim of this study was to clarify the role of transesophageal echocardiography in detecting cardiac sources of embolism in elderly stroke patients. METHODS: We performed transesophageal echocardiography in 77 patients > or = 70 years old (mean 76.9) with ischemic stroke and investigated embolic sources. Thirty-seven patients were in sinus rhythm (SR) and 40 in atrial fibrillation (Af). RESULTS: Left atrial spontaneous echo contrast was detected in 73% of Af and in 14% of SR (p<0.01). Left atrial thrombus was present in 10% of Af and none of SR (p<0.05). Patent foramen ovale, atrial septal aneurysm, and aortic atherosclerotic plaque > or = 4.0 mm in thickness in the proximal aortic arch were more commonly found in patients with SR. CONCLUSIONS: In elderly ischemic stroke patients, 1) Left atrial spontaneous echo contrast and thrombus are more commonly detected in patients with Af, reflecting left atrial enlargement and blood stasis, and 2) atrial septal aneurysm, patent foramen ovale and aortic atherosclerotic plaque > or = 4.0 mm in thickness in the proximal aortic arch are important findings in patients with SR. PMID- 10526940 TI - A clinicopathological study of lung cancer patients with occupational exposure to chrysotile asbestos fibers. AB - OBJECTIVE: To summarize the features of asbestos-related lung cancer. PATIENTS: Thirty-one lung cancer patients with occupational exposure to chrysotile asbestos fibers. They worked or had worked in one asbestos factory or its subcontracters. RESULT: All patients were male with mean age of 60.6 when diagnosed, and all except one were current or ex-'heavy' smokers. Histological types were fairly evenly divided into adeno-, squamous and small cell carcinoma and 24 (78%) of patients showed 'peripheral type' lung cancers. Regarding clinical stages, 20 patients (65%) were classified as III or IV (advanced stage). Tumor shadow(s) was detected on chest X-ray in 22 patients (71%), and in 5 patients with 'negative' chest X-ray, chest CT was necessary to recognize a primary tumor. Seventeen patients (55%) did not undergo periodical check-ups. CONCLUSION: Occupational asbestos exposure is interpreted as one of the important risks for lung cancer and frequent and accurate observation is necessary. PMID- 10526941 TI - Asymptomatic cerebrovascular lesions detected by magnetic resonance imaging in patients with systemic lupus erythematosus lacking a history of neuropsychiatric events. AB - OBJECTIVE: To clarify the extent of asymptomatic cerebrovascular involvement in systemic lupus erythematosus (SLE). PATIENTS AND METHODS: Cerebral magnetic resonance imaging (MRI) findings and ultrasonography findings of 100 patients with SLE lacking present or past clinical neurologic deficits were compared with 66 age-matched volunteers to determine the combined intima-media thickness (IMT) of the common carotid artery, and tests for anti-cardiolipin antibodies (aCL). RESULTS: Thirty-eight patients, but only 2 controls, showed imaging abnormalities. Among 23 SLE patients with cerebrovascular lesions by MRI who underwent single-photon emission computed tomography (SPECT), 14 showed hypoperfusion of the lesion. The IMT value and prevalence of aCL did not differ between the 55 SLE patients tested and controls. SLE disease activity index (SLEDAI) as assessed by a quantitative clinical index was significantly greater in patients with brain lesions than in those without. CONCLUSION: The prevalence of asymptomatic brain lesions in SLE patients is highs and shows a relationship to disease activity. PMID- 10526942 TI - Rapidly growing primary gastric B-cell lymphoma after eradication of Helicobacter pylori. AB - Helicobacter pylori (H. pylori) infection plays a decisive role in primary gastric B-cell lymphoma especially of mucosa-associated lymphoid tissue (MALT) type. We treated a 47-year-old male patient with primary gastric B-cell lymphoma associated with H. pylori infection. Although antibiotic therapy for eradication of H. pylori caused great improvement in the low-grade MALT lymphoma-like lesion, the small areas of high-grade lesion rapidly formed a new bulky mass in only 8 weeks. This suggests that eradication of H. pylori is not effective for high grade lymphoma. PMID- 10526943 TI - Hb Ube-2 in a diabetic case with an abnormally low HbA1C value. AB - A 69-year-old male diabetic patient had an abnormally low HbA1C value of 2.8%, which was inconsistent with his elevated fasting plasma glucose of 8.2 mmol/l. Hb analysis disclosed that the abnormal Hb was Hb Ube-2 [alpha68 (E17) Asn --> Asp] and it accounted for 21.5% of the total Hb. Since the glycated abnormal Hb emerged at the same position as did HbF on high performance liquid chromatography, the HbA1C value was falsely low. The present case demonstrates that Hb Ube-2 is one of the abnormal Hbs in which caution should be exercised when monitoring diabetic control. PMID- 10526944 TI - Adrenocortical insufficiency associated with long-term high-dose fosfestrol therapy for prostatic carcinoma. AB - A 59-year-old man was admitted to our hospital because of muscular pain, weakness, and anorexia. He had been treated with 600 mg/day of fosfestrol, a synthetic estrogen, for 10 years for prostatic carcinoma. Endocrinological studies demonstrated adrenocortical insufficiency due to inadequate ACTH secretion. After initiation of glucocorticoid replacement therapy, his symptoms subsided rapidly. To our knowledge, an association between estrogenic agents, including fosfestrol, and secondary adrenocortical insufficiency has not been previously reported. Physicians who treat patients with long-term and high-dose strong estrogenic agents should be cautious about the possible emergence of secondary adrenocortical insufficiency. PMID- 10526945 TI - A novel mutation in the vasopressin V2 receptor gene in a woman with congenital nephrogenic diabetes insipidus. AB - A 56-year-old Japanese woman with congenital nephrogenic diabetes insipidus (CNDI) is reported. She was diagnosed with CNDI accompanied by advanced gastric cancer. After total gastrectomy, approximately 500 ml fluid per hour was necessary to prevent dehydration. Urinary volume was decreased by administration of hydrochlorothiazide. We detected a novel mutation in the vasopressin V2 receptor gene of her chromosomal DNA. A substitution from G to A was found at the 631 nucleotide position, altering codon 12 from glycine (GGG) to glutamic acid (GAG) in the first extracellular domain. This missense mutation appeared to be the cause of her resistance to arginine vasopressin. PMID- 10526947 TI - Solitary squamous papilloma of the bronchus associated with human papilloma virus type 11. AB - A 79-year-old female presented with persistent dry cough, and a chest radiograph showed a mass shadow in the right upper lung. Bronchoscopic examination revealed that the right main bronchus was severely obstructed by a polypoid tumor, which was diagnosed pathologically as squamous papilloma. After the failure of the attempted endobronchial snare to remove the tumor, right upper lobectomy was performed. The polymerase chain reaction (PCR) examination showed the presence of human papilloma virus type 11 DNA in the resected tumor, suggesting that this virus was the cause of this solitary squamous papilloma of the lung. PMID- 10526948 TI - Multiple osteolysis of peripheral extremities in a patient with adult T cell leukemia/lymphoma. AB - A 67-year-old woman with severe pain in the peripheral extremities was admitted to our hospital. Radiography of the peripheral extremities revealed multiple osteolytic lesions. Antibody to human T cell leukemia virus type-I (HTLV-I) was positive, and right radial bone biopsy showed infiltration of adult T cell leukemic (ATL) cells. Irradiation therapy was effective in the osteolytic lesions. In the present case, elevation of IL-6 was detected, suggesting that IL 6 produced by ATL cells is related to their proliferation in the bone, and local osteolysis. PMID- 10526946 TI - Bronchiectasis with myeloperoxidase antineutrophil cytoplasmic antibody and bactericidal/permeability-increasing protein antineutrophil cytoplasmic antibody. AB - A 56-year-old woman was hospitalized for recurrent hemoptysis. She had been suffering from bronchiectasis for 4 years. Pseudomonas aeruginosa was persistently detected in her sputum. Serum was positive for Myeloperoxidase antineutrophil cytoplasmic antibody (MPO-ANCA) and bactericidal/permeability increasing protein antineutrophil cytoplasmic antibody (BPI-ANCA). She underwent lung resection. Histopathologically, the resected lung showed bronchiectasis with pulmonary fibrosis but did not show vasculitis. Her serum became negative for the ANCAs after the operation. To date, she has no recurrence of hemoptysis. We discuss this case of bronchiectasis with MPO-ANCA and BPI-ANCA and suggest a possible role for ANCAs in chronic airway infection. PMID- 10526950 TI - Successful treatment of cerebral aspergillosis with a high oral dose of itraconazole after excisional surgery. AB - We report a case of cerebral aspergillosis which originated from the sphenoid sinus, and involved a progressive decrease of visual acuity. The neurological signs indicated a cavernous sinus invasion. After extensive intracranial surgery we treated the residual aspergillosis with a high oral dose of itraconazole (800 mg/d for 4 months, followed by 400 mg/d for 5 months). The neurological impairments of the patient gradually subsided with the resolution of the fungal lesion shown on MRI. The successful therapy indicated that itraconazole has a significant role in the treatment of advanced cerebral aspergillosis if it is used in high doses (16 mg/kg/d for adults). PMID- 10526949 TI - Adult T-cell leukemia/lymphoma in which the pathohistological diagnosis was identical to that of Ki-1 positive anaplastic large cell lymphoma. AB - A 65-year-old man developed severe lumbago and a loss of appetite two months before presentation. A computerized tomograph at admission revealed soft tissue masses destroying the Th12, L4 and L5 vertebral bones. We diagnosed the lesions to be metastatic bone tumors, but the primary focus could not be determined. Just after the irradiation treatment, abnormal lymphocytes were detected in the peripheral blood cells. Under the suspicion of adult T-cell leukemia/ lymphoma (ATL), we thus performed a lymph node biopsy. The specimens were histologically composed of Ki-1 positive anaplastic large cell lymphoma (ALCL). The lymphoma cells demonstrated a biclonal integration of HTLV-1 proviral DNA. After 6 cycles of chemotherapy, the patient has demonstrated a partial and favorable remission from ATL. PMID- 10526951 TI - Peritonitis due to Mycobacterium fortuitum infection following gastric cancer surgery. AB - Mycobacterium fortuitum is a well-documented cause of nosocomial infection. However, no studies have reported peritonitis with M. fortuitum as a postoperative complication. We describe a case of peritonitis with M. fortuitum biovariant peregrinum following gastric cancer surgery. Gram-positive bacterial infection coexisted. Although the source of the infection was unclear, the patient was successfully treated with drainage tube exchange and combination therapy consisting of sparfloxacin, clarithromycin, and imipenem/cilastatin sodium. Thus for postoperative infectious pathogens, not only bacteria but also nontuberculous mycobacteria should be considered. PMID- 10526952 TI - Microcirculation and arterial hypertension. AB - A large part of the pressure gradient takes place in the microvascular network (which corresponds to vessels less than 150 microns in diameter). Most of the changes in the peripheral resistance associated with hypertension affect the microvascular network. From the brief review presented here, it appears that the functional characteristics of arterioles are significantly modified in hypertension. Sensitivity to numerous vasoconstrictive substances is increased. Local ACE activity is considerably higher, and endothelium-dependent dilation is lower, in genetically hypertensive animals than in control models. The myogenic response, which represents the vasoconstriction of arterioles in response to a stepped increase in pressure, is also amplified by mechanisms dependent on both prostanoids and endothelin. Changes also affect the structure of the microvascular network. Morphological alterations in the arteriolar wall are not observed for all types of hypertension. Conversely, arteriolar and capillary rarefaction appears to be the most commonly observed change affecting the structure of the microvascular network. The first stage of rarefaction is functional and affects the number of vessels perfused but not the total number of vessels of the microvascular network. At this stage, potent dilators can induce a recruitment of microvessels, which may cancel the difference between the number of perfused vessels in hypertensive and normotensive animals. The second stage is the anatomical rarefaction corresponding to a decrease in arterioles and/or the total number or density of capillaries. Microvascular rarefaction has also been described in patients even in the early stages of hypertension. This led us to consider the microvascular network not only as one of the putative factors responsible for increased pressure but also as a key target of hypertension. Consequently, antihypertensive drugs should also be assessed and differentiated in terms of their efficacy in preventing or reversing the microcirculatory damage associated with hypertension. PMID- 10526953 TI - Growth factors and endothelial dysfunction. AB - Endothelial dysfunction has been implicated in the pathogenesis of many cardiovascular diseases: experimental and clinical studies have shown that endothelial dysfunction may be a key factor in various processes, including abnormal arterial vasomotion, thrombosis or neointimial proliferation. Endothelial dysfunction has been shown to be a characteristic feature of atherosclerotic vessels, sites subject to mechanical injury or collateral vessels that develop in response to severe ischaemia. Fibroblast growth factor (FGF) and vascular endothelial growth factor (VEGF) are important growth factors for endothelial cells in vitro. While VEGF is specific for endothelial cells. FGFs are also potent growth factors for other cell types such as smooth muscle cells. Recent studies have demonstrated the feasibility of using endothelial cell growth factors in vivo. Basic FGF (bFGF) and VEGF have been shown to increase the development of collateral vessels in ischaemic models and to enhance the extent of endothelial regrowth following arterial injury. The marked anatomical improvement associated with the administration of endothelial cell growth factors has promoted questions concerning a possible role for these factors in endothelial dysfunction. In vivo administration of endothelial cell growth factors is associated with significant improvement in endothelium-dependent responses. This effect is observed with bFGF and VEGF in various animal models of endothelial dysfunction such as the collateral circulation, the regenerated endothelium following arterial injury and experimental atherosclerosis. While the precise mechanisms underlying this ubiquitous beneficial effect of endothelial cell growth factors are still to be determined, these results do support the concept of using such factors as a new therapeutic strategy in patients with vascular diseases. PMID- 10526954 TI - New techniques for clinical assessment of the peripheral microcirculation. AB - Current methods for clinical investigation of the cutaneous microcirculation in patients are based mainly on laser Doppler and capillary microscopy. The use of laser Doppler gives a semi-quantitative index of superficial tissue perfusion. The most recent devices are capable of analysing both the volumetric and velocimetric components. New instruments use two different frequencies to compare tissue perfusion at different depths beneath the skin surface. The combination of a laser probe and a small automate produces a 2-dimensional image, allowing the evaluation of spatial heterogeneity in tissue perfusion, an important pathophysiological concept in vascular diseases. Capillaroscopy has recently been improved by the emergence of the flexible videomicroscope, allowing easy exploration of not only the classical site of the nail-fold but also of the body skin surface. The use of this method was therefore extended--from peripheral vascular disease and connective tissue diseases to the whole spectrum of trophic changes in the skin of the extremities. Systems for digital image analysis allow quantification of the structure of the microvascular bed (quantitative appraisal of microangiopathies) and function (capillary haemodynamics and exchange). Laser Doppler and capillaroscopy can also be combined for the measurement of red blood cell velocity in single capillaries. PMID- 10526955 TI - Coronary microcirculation and cardiovascular pathology. AB - The recent arrival of new techniques for exploring the coronary microcirculation has facilitated assessment of both the incidence and consequences of disorders of this network in a large number of cardiovascular diseases. The microcirculation is affected in numerous cardiomyopathies in the presence of different cardiovascular risk factors and also following cardiac transplantation. Dysfunction of the microcirculation may correspond to a reduction in the surface of the maximum section of coronary arterioles, which involves multiple mechanisms, although this phenomenon does not appear to play a role in ischaemic heart disease. Reduced coronary flow is most frequently related to vascular rarefaction of multifactorial origin, including greater or lesser degrees of intimal proliferation, perivascular fibrosis, hypertrophy of the media and extrinsic compression. PMID- 10526956 TI - Angiogenesis and gene therapy in man: dream or reality? AB - Preclinical studies indicate that angiogenic growth factors can stimulate the development of collateral arteries in animal models of peripheral or myocardial ischaemia, a concept termed 'therapeutic angiogenesis'. The goal of this review is to provide a brief overview of the advantages and disadvantages of gene versus recombinant protein therapy for therapeutic angiogenesis. We also discuss different options for delivering genes to patients, including plasmids and modified viral vectors. Recently, the safety and potential utility of gene therapy for ischaemic disease were demonstrated in 3 clinical trials involving the delivery of plasmid DNA encoding the 165 amino acid isoform of human vascular endothelial growth factor (phVEGF165), a factor that specifically promotes the proliferation and migration of vascular endothelial cells. Two trials involved the administration of phVEGF165 for peripheral arterial disease. In one trial, the plasmid was administered to the arterial wall from a hydrogel-coated angioplasty balloon, while a second trial examined the direct injection of phVEGF165 into the skeletal muscle of the affected limb. More recently, phVEGF165 was directly injected into ischaemic myocardium. In all these trials, it appears that administration of phVEGF165 led to improvements in tissue perfusion. PMID- 10526958 TI - An evolutionary role of formaldehyde. AB - The evolution can be divided into three stages: chemical, prebiological and biological evolution. Most of the problems emerge when the development of cellular organization, the so-called prebiological evolution, is investigated. Here the possible evolutionary roles for formaldehyde as well as for the methylglyoxalase pathway are proposed. The theory, on the one hand, ascertaines a pathway serving as an anaplerotic route for the reductive citric acid cycle of surface metabolists and using formaldehyde as raw molecule. On the other hand, an explanation for the glyoxalase enigma is offered hoping that in this way a long lasting mystery of almost nine decades biochemical research can be solved. PMID- 10526957 TI - Arteriolar and capillary remodelling in hypertension. AB - The major haemodynamic abnormality underlying elevated blood pressure in hypertension is an increase in vascular resistance. The microcirculation is a key site of increased vascular resistance in hypertension. A reduced density of arterioles and capillaries is an important common characteristic of various microvascular beds in many forms of hypertension. This microvascular rarefaction has been observed even in very early stages of the development of hypertension. A hypothesis is discussed for early microvascular rarefaction being the result of genetic and fetal mechanisms of decreased small blood vessel growth. PMID- 10526959 TI - Related alphaN- and epsilonN-methyltransferases methylate the large and small subunits of Rubisco. AB - Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) is methylated at the ax amino position of the N-terminal methionyl residue of the processed and assembled form of the small subunit (SS), and is also methylated in some species at the epsilon-amino group of lysine-14 in the large subunit (LS). The gene (rbcMT-S) and cDNAs for the SS alphaN-methyltransferase (SSMT) from spinach (Spinach oleracea) have been cloned, sequenced, and expressed. The gene is closely related to a previously characterized LS methyltransferase (Rubisco LSMT) cDNA from pea (Rubisco LSMT) and a Rubisco LSMT gene from tobacco. Sequence analysis of the cDNA and transcript mapping experiments demonstrate that the rbcMT-S pre-mRNAs experience alternative 3' splice site selection, such that mRNAs for a long form with a four amino acid insertion and a short form are expressed at approximately equal abundance. The coding sequence of spinach SSMT includes a putative targeting presequence with sequence identity at a plastid processing site. A N terminal truncated form of spinach SSMT was expressed and purified from E. coli cells. Both long and short forms of the cDNAs were shown to catalyze methylation of the a amine of the N-terminal methionine of the SS of Rubisco. PMID- 10526960 TI - Methylation and gene mutation in eukaryotic DNA. AB - 5-methylcytosine (m5C) as a rare base exists in eukaryotic genomes, which is a normal constitution in many eukaryotic DNA and the existence of m5C is a feature of eukaryotic DNA. Under regular physiological conditions, cytosine of eukaryotic DNA is usually methylated. Up to the present, many people consider that the m5C may be mutation hotspots by the deamination leading to gene mutation. Our study indicated that the spontaneous mutation caused by the transition of G.C --> A.T, in eukaryotic DNA, may result from the tautomer changing of base pairs and may also be cause by other factor actions, however it could not be caused by the deamination of m5C. PMID- 10526962 TI - Effect of methionine enrichment on the biological activities of food proteins. AB - A special enzymatic technique, the enzymatic peptide modification (EPM) reactions have been worked out in our laboratory, by which proteins can be modified with respect to their biological activities. Enzymatic modification methods provide several advantages in producing proteins of desired nutritional properties. Proteolytic modification, particularly when combined with methionine enrichment, significantly altered the biological activities and leads to increased nutritional value. PMID- 10526961 TI - Role of formaldehyde in direct formation of glycine and serine in bean leaves. AB - In our early study [2] labelled formaldehyde, glyoxylate, glycine and serine were formed from 14CO2 during 1 min incubation. Continuing the detailed work by unlabelled HCO3- with short (1 min) incubation time in numerous experiments, there were significant changes in the amount of formaldehyde, glyoxylate, glycine and serine. The main product was glycine. In illuminated leaves after HCO3- intake, the formaldehyde content increased. However, in dark in the presence of 5 mM HCO3- the formaldehyde content decreased and the amount of glyoxylate, glycine and serine increased in green leaves. The transamination is improbable as the amount of glutamate and aspartate did not decrease during the glycine and serine formation. The change in the amount of the measured free amino acids and glyoxylate was hindered by (aminooxy)acetic acid (AOA) and amethopterin. AOA, which is an inhibitor of pyridoxal phosphate has no effect on formaldehyde alteration. In the presence of amethopterin which inhibits the activity of tetrahydrofolate the formaldehyde amount increased in dark kept CO2 containing green leaves instead of decreasing. This shows that tetrahydrofolate has a role in glyoxylate formation from formaldehyde and CO2. A direct formation of glycine and serine can be supposed in definite circumstances during photosynthesis, in which pyridoxal phosphate has an important role. PMID- 10526963 TI - L-carnitine as essential methylated compound in animal metabolism. An overview. AB - Certain kinds of N-methylated compounds proved to be essential for animals. L Carnitine is a small-molecular-weight quaternary amine which occurs naturally in micro-organisms, plants and animals. Generally, plants contain little carnitine compared to animals where especially high levels may be found in heart and skeletal muscle. The main function of L-carnitine is the translocation of long chain fatty acids from the extramitochondrial space to the mitochondrial space. It also facilitates the removal from mitochondria of short-chain and medium-chain fatty acids that accumulate as a result of normal and abnormal metabolism. This pathway provides a means to regenerate the intramitochondrial free coenzyme A under conditions where short-chain acyl-CoA esters are produced at a rate faster than they can be utilized. L-Carnitine is synthetized by most animals but its supplementation can be beneficial under certain condition including insufficient carnitine synthetic enzyme activity, metabolic abnormalities, dietary deficiencies or malnutrition. Several studies on pigs, fish, foal, quail and broiler chickens demonstrate a growth improvement by feeding extra dietary L carnitine. It has been also found that L-carnitine supplementation resulted in lowered muscle and liver lipid contents. Formaldehyde is involved in metabolism, methylation-demethylation processes of these compounds. PMID- 10526964 TI - Addition of CH2O to arginine--a theoretical study by ab initio method. AB - Arginine is an acid possessing a guanidine side group which is protonated at all biological pHs, R-NH-C(=NH2+)-NH2. We have performed computations on the neutral species by the ab initio method using MINI and 6-31G(d) basis sets. Geometry optimization at the HF SCF level has also been carried out using the 6-31 G(d) basis set. We investigated the addition of formaldehyde to both the NH2- and H N=C sites of the guanidine side group. The calculated exothermicities of the reactions with formaldehyde were found different for the above structures. We are planning to determine the energy barriers for these reactions in the near future. PMID- 10526965 TI - Formaldehyde cycle and the natural formaldehyde generators and capturers. AB - S-adenosyl-L-methionine serves as a methyl donor in virtually all of the vast number of enzymatic transmethylation reactions including DNA methylation. On the basis of our former experiences we questioned the formation of a methyl cation or methyl radical in the enzymatic transmethylation reactions. The formation of formaldehyde from the methyl moiety of S-adenosyl-L-methionine has been demonstrated. It became increasingly evident that there is a formaldehyde cycle in biological systems in which the formation of the methyl group of L-methionine takes place through formaldehyde and the formation of formaldehyde from S adenosyl-L-methionine is linked to different enzymatic transmethylation reactions. It is also known that during demethylation processes both formaldehyde and demethylated compound can be formed. The abnormalities of the originally controlled formaldehyde cycle and the uncontrolled enzymatic production of formaldehyde from endogenous and/or exogenous substrates may be potential risk factors in pathogenesis of different disorders. The formaldehyde generator and capturer molecules may potentially normalise these abnormal processes. Trans resveratrol (trans-3,5,4'-trihydroxystilbene), which is as phytoalexin, occurs naturally in grapes and a variety of medicinal plants. According to our present observations it is a natural concentration-dependent formaldehyde capture molecule. It would seem that elimination of the uncontrolled formaldehyde with resveratrol may exert a double effect in biological systems. The elimination of formaldehyde with resveratrol (first step) may cause a cardioprotective effect and the reaction products between resveratrol and formaldehyde (second step) may act as a chemopreventive factor against cancer. PMID- 10526966 TI - Formaldehyde in the plant kingdom. AB - Formaldehyde, at its dimedone adduct, formaldemethone, has been detected by thin layer and high-performance liquid chromatography in extracts of all species tested of marine algae, macrofungi, lichens, bryophytes, pteridophytes, gymnosperms and angiosperms. The yields of formaldehyde recorded in this study varied from 30 microg/g to 4060 microg/g, fresh weight. PMID- 10526967 TI - Plant tissue culture as a model for study of diversity in formaldehyde binding. AB - The effect of endogenous formaldehyde (HCHO) deprivation with dimedone - as abiotic stress - was investigated on the methylation-demethylation reactions in Datura innoxia Mill. callus cultures. The matrix assisted laser desorption/ionization mass spectrometric (MALDI MS) investigation of the culture extracts revealed characteristic differences between the tissues cultivated in light and dark. MALDI MS data show the presence of a precursor molecule and its mono- and tri-hydroxymethyl derivative, only in the cultures maintained in light. The relative amount of the different derivatives changed considerably, as a consequence of dimedone application in the culture medium and during the extraction. PMID- 10526968 TI - Determination of endogenous formaldehyde in plants (fruits) bound to L-arginine and its relation to the folate cycle, photosynthesis and apoptosis. AB - A very powerful nucleophilic reagent, hydralazine(1-hydrazino-phtalazine) proved to be suitable for determination of the endogenous formaldehyde level in biological samples. It was found that in different plants (vegetables, fruits, especially in red beet, cauliflower, kohlrabi, grapes) is a large amount of releasable endogenous formaldehyde (0.5-1.0 mM) bound to L-arginine mainly in the form of N(G)-trihydroxymethyl-L-arginine (TriHMA). N(G)-hydroxymethyl-L-arginines (HMA) were proved to transfer their hydroxymethyl groups to tetrahydrofolic acid producing N5,N10-methylene-tetrahydrofolate, the coenzyme of thymidylate synthase. HMA was found to inhibit the cell proliferation of HT-29 cell culture (human colon adenocarcinoma ATCC HT-B 38) causing apoptosis. Photosynthetic experiments produced confirmatory evidences that 14CH2O could be formed in photosynthesis already after 10 seconds of 14CO2 fixation in the seedlings of Zea mays L. (single cross) and the 14CH2O was immediately trapped by L-arginine mainly as TriHMA. PMID- 10526969 TI - The hydrazine derivative aminoguanidine inhibits the reaction of tetrahydrofolic acid with hydroxymethylarginine biomolecule. AB - Aminoguanidine (AG), a hydrazine derivative is known to inhibit the formation of Advanced Glycosylation Endproducts (AGE) and AG has been proposed as an agent in prophylaxis of diabetic complications. However, treatment with hydrazine produced liver and lung tumors by formation of N7- and O6-methylguanine in the DNA of rodents. The hydrazine derivative, isonicotinic acid hydrazide induced pulmonary tumors in mice. N(G)-hydroxymethyl-arginine (HMA) was synthesized by our research group and it showed anticancer effect against experimental tumors. HMA was found earlier in human blood and urine, and recently in many plants (in fruits and vegetables). We could demonstrate a reaction (pH = 7.5, 37 degrees C, 1h) between HMA and tetrahydrofolate (THF) producing N5,N10-methylene-tetrahydrofolate (CH2 THF), the coenzyme of thymidylate synthase (TS). In model experiments AG proved to react with formaldehyde (HCHO) and to eliminate the C1-fragment of HMA, but not that of CH2-THF. In the presence of AG burst chemiluminescence and a higher speed of the formylation and methylation reactions were found in the AG, HCHO, hydrogen peroxide (H2O2) and L-lysine system than without AG. CONCLUSIONS: HMA as a biomolecule is one of the compounds which are responsible for the endogenous HCHO level. The biochemical function of HMA may be the direct supply of C1 fragment for the folate cycle. AG can disturb the above function of HMA. The reaction between AG and HCHO seems to be dangerous for biological systems because of the possible presence of L-lysine and H2O2. The burst chemiluminescence indicates excited molecules with extreme high energy producing uncontrolled formylation and methylation reactions. Considering the results of the experiments with AG its use as a medicament seems to be questionable. PMID- 10526970 TI - Induction of resistance of wheat plants to pathogens by pretreatment with N methylated substances. AB - It has been established that the application of N-methylated compounds as N, N dimethyl-L-tyrosine and glycine-betaine can induce resistance to biotrophic fungi (Erysiphe graminis, Puccinia recondita) in wheat (Yubileynaya) at two different pretreatment concentrations. This dose-dependent "double immune response" of plants is the basic phenomenon of the biochemical immunization. These N methylated compounds are potential formaldehyde generators and the formaldehyde formed can take part in the induction of resistance of wheat plants. It seems that the dose-dependent "double immune response" of plants is in correlation with the biotransformation steps of formaldehyde cycle as a fundamental biochemical pathway. PMID- 10526971 TI - Identification and measurement of resveratrol and formaldehyde in parts of white and blue grape berries. AB - The phytoalexin resveratrol (3,5,4'-trihydroxy-trans-stilbene) and formaldehyde (as its dimedone adduct, formaldemethone) have been identified and measured in the extracts of parts of white and blue grapes as well as in white and red wines by overpressured layer chromatography (OPLC), high performance liquid chromatography (HPLC) and from matrix assisted laser desorption/ionization mass spectrometric data. It has been established that the level of resveratrol was very high in skin and in some cases in the stem. Blue grape varieties and red wines always contained a considerably higher amount of resveratrol than white grapes and wines. The measurable level of formaldehyde as well as the resveratrol content was always parallelly high in the same parts of the berries, however, the formaldehyde level was higher in white grapes than in blue ones. The simultaneous occurrence of resveratrol and formaldehyde gives a possibility for interaction between these two special molecules, consequently, hydroxymethyl derivatives of resveratrol can be formed. These resveratrol derivatives may be responsible for special biological activities of resveratrol in grapes and dietetically (cardioprotective effect and chemopreventive effect against cancer) in the human organism. PMID- 10526972 TI - Relationship between dimedone concentration and formaldehyde captured in plant tissues. AB - In different parts of water-melon plants (Citrullus vulgaris L.) formaldehyde (HCHO), in dimedone adduct form (formaldemethone), and fully N-methylated substances were identified and determined by OPLC as well as HPLC, capillary GC and GC-MS methods using authentic substances. The HCHO captured originates from dynamic methylation and demethylation processes in which this simplest aliphatic aldehyde is bound in the form of highly reactive hydroxymethyl groups. Dimedone will react with the small quantity of HCHO in equilibrium with the hydroxymethyl groups and it follows from this that the rate of HCHO captured as the dimedone adduct will increase parallel to the increasing concentration of dimedone applied, as a methanolic solution, until a plateau is reached. The level of HCHO was very high in the roots of water-melon seedlings. PMID- 10526973 TI - The use of HPLC for the detection and quantification of formaldehyde in the pteridophyta. AB - Formaldehyde, as its dimedone adduct, formaldemethone, has been detected and quantified in all species of Pteridophyta examined. The procedure involved the use of an Hypersil C-18 column, methanol-water (60 : 40 v/v) as the mobile phase and an UV detector set at 258 nm. Quantification was based on peak height. Yields varied from 30 microg/g fresh weight for Polystichum setiferum to 5370 microg/g fresh weight for Selaginella viticulosa. PMID- 10526974 TI - Drought stress, peroxidase activity and formaldehyde metabolism in bean plants. AB - In our investigations we have studied the changes of total peroxidase and isozyme activities, peroxidase isozyme pattern and the level of endogenous formaldehyde and trigonelline. We have examined the effect of drought stress resulted by Carbowax treatment (2, 5, 7, 10%) of two different snap bean (Phaseolus vulgaris L.) genotypes [drought tolerant (T) and drought sensitive (S)]. There were no detectable differences in the peroxidase isozyme patterns but the total peroxidase activity was increased. We have found similar steep enhancement in the peroxidase activities of one of three peroxidase fractions while the increase in the other isozyme fractions was not so significant in the two genotypes. Increasing peroxidase activities and changes in the amount of endogenous formaldehyde and trigonelline were detectable with increasing Carbowax treatment in both genotypes, but in different ways. According to our experiments, the total peroxidase and some isozyme activity correlated with the concentration of endogenous formaldehyde and trigonelline, playing an important role during drought stress. PMID- 10526975 TI - The increase of formaldehyde level in some rare pathological cases of teeth determined with the use of quantitative TLC. AB - In our previous works we proved that formaldehyde (HCHO) level in hard tissues of teeth could depend on their physiological state. In the current paper we presented the results of HCHO determinations in different dental pathologies, mainly the ones rarely encountered in dental practice, e.g. in the case of reinclusion. The determination of HCHO in the form of dimedone was performed by means of quantitative TLC. The obtained results were compared with HCHO levels in hard tissues of teeth presenting pathological changes. It proved that the highest HCHO level was found in reincluded teeth while it was lower in retained teeth, that is the ones which are not subjected to stress factors present in the mouth. The obtained results can constitute a contribution to the problems of dental pathologies, mainly caries which is a very common problem. PMID- 10526976 TI - Formaldehyde generators and capturers as influencing factors of mitotic and apoptotic processes. AB - There is a growing amount of evidence pointing to the fact that several endogenous and exogenous methylated compounds are potential formaldehyde generators in their biological reactions. N(G)-methylated lysines, N(G) methylated as well as hydroxymethylated arginines, and 1'-methyl-ascorbigen have been examined in this respect. The apoptosis-inducing effect of formaldehyde molecules formed from methyl groups was earlier first published by our group. Dimedone, an artificial capturer molecule for formaldehyde, has been found to prevent the apoptosis-inducing effect of 1'-methyl-ascorbigen as well as N(G) hydroxymethylated arginines. More recently resveratrol, present in grapes and wines, has been shown to have cardioprotective and cancer chemopreventive effect. Our group has been successful in demonstrating that this natural formaldehyde capturer molecule can also influence cell proliferation and apoptosis. The apoptosis-inducing or -preventing effect of formaldehyde generators and capturers seems to be dose-dependent and may be utilized in various disturbances of cell proliferation and active cell death. PMID- 10526977 TI - Reduction of apoptosis of in vitro cultured lymphocytes of HIV-positive persons by N(G)-hydroxy-methylated-L-arginine and 1'-methyl-ascorbigen. AB - Some formaldehyde generating chemicals due to reduction of apoptosis in lymphocytes may slow down the progress of immune decline of HIV-infected individuals. N(G)-hydroxy-methylated-L-arginine (MAX) and 1'-methyl-ascorbigen (MeAsc) could enter this way the biochemical pathway of cells and affect the apoptotic process. Separated peripheral blood lymphocytes of five asymptomatic HIV-positive persons were cultured. Unstimulated, IL-2 stimulated and IL-2 stimulated plus 0.1, 1.0, 10.0 microg/ml MAX or MeAsc treated lymphocytes were investigated for apoptosis morphologically (HE) and by flow cytometrical DNA fragmentation method. IL-2 stimulation lowered the apoptotic rate in lymphocytes of HIV-positive persons related to unstimulated ones. MAX and MeAsc reduced the apoptotic activity of stimulated lymphocytes in the least or the middle doses while in the higher dose did not. MAX and MeAsc reduced the apoptotic activity of stimulated lymphocytes originated from HIV-positive patients in vitro. This compounds may have the same effect in vivo and may prolong the symptomless period of HIV-infected patients. The role of methylation and production of formaldehyde in this process is discussed. PMID- 10526978 TI - Formaldehyde generation by N-demethylation. AB - Microsomal oxidation of exogenic compounds yields efferent metabolites with small molecular size. N-demethylation results in formaldehyde generation in addition to the nor-compound. Interesting changes in the level of formaldehyde elimination were observed after a single dose of either ( )-deprenyl or (+)-deprenyl. Urine elimination of the generated formaldehyde was determined using thin-layer chromatography after derivatization with dimedone. PMID- 10526979 TI - Formaldehyde-induced modification of hemoglobin in vitro. AB - Formaldehyde is known to react with proteins. The purpose of our experiments was to analyse in vitro the effect of formaldehyde on the physicochemical and biological properties of hemoglobin molecules. The effect of formaldehyde concentration, reaction time, pH and temperature on hemoglobin free amino groups was estimated. The modified hemoglobin was analysed using electrophoretic, potentiometric and spectrophotometric techniques. Reaction between formaldehyde and hemoglobin was accelerated by increasing concentration of formaldehyde and higher temperature. This reaction was most intensive during the first few hours at pH 7.4 so the amount of free amino groups of hemoglobin was significantly diminished by directly mixing formaldehyde with hemoglobin. The modified protein was characterized by the increase in electrophoretic mobility and the decrease in maximum absorption derived from porphyrin rings. Formaldehyde modified hemoglobin was less susceptible to the action of cathepsin D. PMID- 10526980 TI - Change of biotransformation steps of formaldehyde cycle in water-melon plants after infection with Fusarium oxysporum. AB - Recent experiments indicate that the measurable formaldehyde (HCHO) level is considerably elevated in the parts of water-melon plants immediately after a nonlethal infection with Fusarium oxysporum f. sp. niveum. At the same time the level of some quaternary ammonium compounds (N(epsilon)-trimethyl-L-lysine, choline) as potential HCHO generators (gene products) is considerably decreased. That is probably due to the fact that the alarm reaction phase of this biotic stress syndrome includes an intensive demethylation process. It has been proved that HCHO may play a role in dynamic methylation-demethylation processes that also may include the methylation of biotic stress proteins. In this paper we report on qualitative and quantitative changes in the biotransformation steps of the formaldehyde cycle in different parts of the water-melon plant after nonlethal infection (biotic stress) with Fusarium. In consequence of the infection identical quantitative changes, but to a different degree, of the compounds examined are observable in both varieties. The connections resulting from the depiction of the time-dependent quantitative changes of the measured methylated compounds due to infection show a picture similar to that of Selye's stress syndrome model. PMID- 10526981 TI - The effect of heat shock on the formaldehyde cycle in germinating acorns of European Turkey oak. AB - The effect of heat shock (40 degrees C) on the formaldehyde cycle has been studied in European Turkey oak (Quercus cerris L.) acorns germinated to a 10% increase in mass. Hydroxy-methyl groups bonded to sulfur, oxygen and nitrogen atoms were made to react with dimedone and the derivative obtained (formaldemethone), which represented the endogenous formaldehyde level, was determined by high performance liquid chromatography. Qualitative alterations of methyl donors and acceptors in the response of acorns to the heat shock have been mapped by MALDI (matrix assisted laser desorption ionization) mass analysis. In the first experiment the acorns were prevented from withering by wrapping them in aluminium foil and in the second they were not. The relatively high temperature of the acorns wrapped in aluminium foil was the dominant stress effect and the role of withering was subsidiary. Alteration of the endogenous formaldehyde level in the seed-leaves reflected the phases of the stress syndrome. If the withering were not hindered, two local minima in the alteration of endogenous formaldehyde level were found. First, the increase in temperature decreased the endogenous formaldehyde level and after a local maximum a repeated local minimum was observed as a delayed response. It is presumed that the second minimum was induced by the decreasing water amount becoming more and more significant in the seed-leaves. PMID- 10526982 TI - Changes in formaldehyde contents of germinating acorns of Quercus cerris L. under low temperature stress conditions. AB - Acorns of Quercus cerris L., after saturation with water and storage at -20 degrees C, were studied for changes in their contents of endogenous formaldehyde and its potential precursor and generator compounds. For the measurement of formaldehyde, after conversion to formaldemethone and some methyl acceptor and donor substances, high performance liquid chromatography (HPLC) were used. First, the amount of formaldehyde was drastically decreased. Having reached a minimum value within three to five days of the beginning of low temperature storage, a higher steady-state than the control acorns was recorded. Trigonelline and gamma amino-butyric acid in seedleaf extracts were identified by matrix assisted laser desorption-ionization mass spectrometry (MALDI-MS). PMID- 10526983 TI - Alteration of endogenous formaldehyde level following mercury accumulation in different pig tissues. AB - The effect of mercury accumulation on the formaldehyde cycle of different pig tissues resulted by a single dose of mercury (0.4 mg mercury in Hg(II)-chloride form, 500 kBq Hg-203/animal) has been studied. Daily mercury excretion was tracked, and having reached the steady-state mercury level of the body (10th day), samples were taken from the liver, kidney and muscle (musculus longissimus dorsi). After reaction with dimedone the endogenous formaldehyde levels in the samples were measured by high performance liquid chromatography. Our results show that the endogenous formaldehyde level decreased by more than 50% in the liver and the kidney, where the average mercury accumulation was the highest (1017 Bq/100 g and 625 Bq/100 g, respectively). In contrast, the muscle tissues, with a low mercury level (139 Bq/100 g), responded to the stress effect by about a 30% increase in their endogenous formaldehyde level. PMID- 10526984 TI - Investigation of some methylated compounds and peroxidase activity during plant ontogenesis in snap bean. AB - Changes in the level of endogenous formaldehyde (HCHO) and some N-methylated compounds were investigated in the leaves of snap bean (Phaseolus vulgaris L.) during ontogenesis. In addition, the activity and isozyme pattern of peroxidase enzymes were also examined. HCHO, as dimedone adduct, and fully N-methylated compounds were determined by overpressured layer chromatography in different development stages of snap bean plant. Peroxidase activities were measured by spectrophotometry and isozyme patterns were examined by isoelectric focusing. HCHO level decreased until blooming with aging of the plant being the highest in the youngest leaves all the time, but then increased again in old leaf tissues. At the same time the concentration of choline and trigonelline as potential HCHO generators (marker molecules or gene products) decreased considerably while peroxidase activity increased with aging of plants. PMID- 10526985 TI - Formaldehyde as a proof and response to various kind of stress in some Basidiomycetes. AB - The influence of cadmium chloride and/or high temperature on the level of selected parameters were examined in both medium and mycelium of some Basidiomycetes belonging to white-rot fungi: Abortiporus biennis, Trametes versicolor and Cerrena unicolor. We investigated changes in the formaldehyde (FA) level and in the level of superoxide radical anions (SR). Accordingly, the capacity of three enzymes was also studied: two enzymes of the cellular antioxidative system - superoxide dismutase (SOD; EC 1.15.1.1), and catalase (CAT; EC 1.11.1.6), and laccase (LAC; EC 1.10.3.2) the main lignin-modifying enzyme, which is produced by white-rot fungi. During the first 24 hours after application of separate stressors, or jointly with two stressors to 10-day-old cultivation, changes in all selected parameters were observed. Moreover, we found significant changes in the levels of extracellular SR, FA and extra- and intracellular activity of LAC. Simultaneous action of two stressors in the fungal cultures decreased the extracellular LAC level, intracellular CAT activity and the level of SR in the medium compared to the control values, while using the two stress factors separately would strongly make these values increase. Stressful conditions brought a rapid increase in the level of FA in all fungal species. The results of our study, carried out on selected strains of Basidiomycetes, seem to have shown that: (I) LAC, the lignin-modifying enzyme, may have an important application in fungal stress response, and take part in the cross-protection between responses to heat shock and cadmium resistance; (2) the oxidative burst as a result of cadmium and high temperature treatment is an additional factor to damage fungal cells; (3) FA may be a determining factor in the phases of fungal stress syndrome. PMID- 10526986 TI - Relationships between demethylase activity, formaldehyde and oxygen during incubation of Rhodococcus erythropolis with veratrate. AB - Relationships between demethylase activity, formaldehyde and oxygen were investigated. Demethylase activity was measured against the following substrates: veratric, vanillic, and isovanillic acids, as well as in the presence of guaiacol. The influence of ATP and GTP on demethylase activity was also checked. Demethylase activity was found to be dependent on the capability of the cells for endogenous oxygen uptake. In some cases ATP produced the opposite effect: instead of being taken up, oxygen was released, which suggested a reversibility of the demethylation reaction. Curiously enough, GTP demonstrated the same effect. Changes in enzyme activity were correlated with those occurring in the level of formaldehyde. The latter increased after addition of ATP, but decreased after addition of GTP. PMID- 10526987 TI - Comparative quantitative chromatographic determination of formaldehyde in different groups of physiological and pathological hard tissues of teeth. AB - Taking into consideration that HCHO level in cells of plant, animal and human tissues as well as in body fluids depends from physiological state of an organism in the current study it was decided to find out if there are changes of HCHO level in different physiological and pathological hard tissues of teeth. The obtained results showed in all 4 groups of teeth separately analysed that there were some regularities in the level of HCHO as far as similar physiological or pathological states are concerned. This was best seen when comparing the obtained results with mean HCHO level of the studied groups of teeth. PMID- 10526988 TI - Are the reductions in nematode attack on plants treated with seaweed extracts the result of stimulation of the formaldehyde cycle? AB - Soil application to the roots of tomato plants (Lycopersicon esculentum) of a commercially-available alkaline extract of the brown alga, Ascophyllum nodosum, resulted in a significant reduction in the number of second-stage juveniles of both Meloidogynejavanica and M. incognita invading the roots, compared to those of plants treated with water alone. Egg recovery from the seaweed extract treated plants was also significantly lower. The three major betaines found in the seaweed extract (gamma-aminobutyric acid betaine, delta-aminovaleric acid betaine and glycinebetaine) also led to significant reductions in both the nematode invasion profile and egg recovery when applied at concentrations equivalent to those present in the extract. This led to the conclusion that the betaines present in the seaweed extract play a major role in bringing about the observed effects. Treatment of Arabidopsis thaliana plants with seaweed extract also resulted in a significant decrease in the number of females of M. javanica which developed in the roots. Significant reductions in egg recovery were also achieved from plants treated with the seaweed extract and similar effects were produced with the betaines found in the seaweed extract. As the experiments were conducted under monoxenic conditions, it can be concluded that the results obtained with the application of either the seaweed extract or betaines are not dependent on microorganisms associated with the rhizosphere. PMID- 10526989 TI - Effect of 1-methylascorbigen on the resistance potential of plants to pathogens. AB - We have studied the effect of 1'-methylascorbigen, an immunostimulating substance in animal systems on the resistance potential of barley, bean and wheat plants to the fungal pathogens Erysiphe graminis, Uromyces phaseoli and Puccinia recondita, respectively. The effectiveness of protection depends - as in the case of other endogenous, N-methylated compounds - on the dosage of applied 1'-methylascorbigen and on the time interval between the chemical pretreatment and inoculation. The time- and dose-dependent double immune response was clearly demonstrated in case of the barley - Erysiphe graminis and bean - Uromyces phaseoli host-parasite relationships while in case of the wheat -Puccinia recondita relationship the relatively long-time interval between pretreatment and inoculation allowed manifestation of only a single immune response. PMID- 10526990 TI - Analogies and differences in the excited reactions of formaldehyde and D-glucose. AB - The investigations proved that D-glucose (as reducing sugar) can easily be activated in a ternary system (L-lysine: D-glucose: H2O2) similarly to formaldehyde at 20 degrees C, in pH = 7.4 forming chemiluminescence (CL) and singlet oxygen. The kinetic investigation showed that: CL lasted many hours (permanent emission) and had no bell-shaped curve differently from other aldehydes e.g. formaldehyde. The reason of the effect is that D-glucose exists mainly in ring form in water solution (Haworth ring form) and the open form (the aldehyde group) is slowly liberated during the excited reaction. These excited reactions may be important in human organism, because D-glucose and lysyl residues of proteins occur permanently in human body and endogenous formaldehyde and H2O2 may be liberated there, too. PMID- 10526991 TI - Microbial urea-formaldehyde degradation involves a new enzyme, methylenediurease. AB - The enzymic mechanism of metabolization of urea-formaldehyde condensation products (methyleneureas; MU) and the fate of the degradation products ammonium, urea and formaldehyde were studied in bacteria isolated from garden soil, which were able to use methyleneureas as the sole source of nitrogen for growth. An organism identified as Ochrobactrum anthropi completely degraded methylenediurea (MDU) and dimethylenetriurea (DMTU) to urea, ammonia, formaldehyde and carbon dioxide. An enzyme designated as methylenediurease (methylenediurea deiminase; MDUase) was responsible for the degradation of both MDU and DMTU as well as higher polymerized MU. Growth on MU as the nitrogen source specifically induced the synthesis of this enzyme, which seems to be located in the periplasm of the bacterium. Under these growth conditions, urease as well as NAD-specific formaldehyde and formiate dehydrogenase were expressed to high levels, efficiently using the products of MU degradation, and high-affinity transport systems for urea and ammonia were synthesized scavenging the environment for these products. PMID- 10526992 TI - Differential detection of N-heterocyclic compounds and their N-methylated derivatives by immunoanalysis. AB - Competitive enzyme-linked immunosorbent assay (ELISA) systems are proposed for the indirect monitoring of formaldehyde by the parallel detection of its N methylated precursors and the corresponding demethylated compounds. As an example for such immunoanalytical differentiation between an N-heterocyclic compound and its N-methylated derivative, the quantitative detection of the systemic triazole fungicide, myclobutanil, is discussed. Antibodies recognizing the non zwitterionic structure of 2-(4-chlorophenyl)-2-[(1,2,4-triazol-1-yl)-methyl] hexanonitril e (myclobutanil) showed only minor binding to corresponding N alkylated derivatives of myclobutanil. And vice versa, literature data indicate that antibodies raised against the pyridilium ionic structure of the herbicide paraquat, displayed only mediocre reactivity towards the corresponding dealkylated derivatives. Thus, both experimental and literature data suggest that immunoanalytical methods for differential detection of N-methylated heterocycles (potentially including formaldehyde precursors) and their non-methylated counterparts are possible to develop. PMID- 10526993 TI - Urea-formaldehyde resins and free formaldehyde content. AB - Specifications of wood adhesives must be in correlation with the requirements for the corresponding wood products, for which they will be used. Formaldehyde emission of wood products bonded with urea-formaldehyde resin based adhesives is strictly regulated by standards and there is a compromise is between formaldehyde emission and performance, such as strength, or water resistance. Since values of formaldehyde emission depend on the test method used, in Europe urea-formaldehyde resins for adhesives may generally be classified according to HCHO emission in the particleboard rating of Emission 0 to Emission I class (E-0 to E-1). According to DIN EN 120, particleboard quality E1 emits <6.5 mg/100 g dry article determined with the perforator method. Although a great number of factors effect the formaldehyde emission of the cured products, such as the hardener system, the type of wood etc., the emission of formaldehyde is in strict correlation with the free formaldehyde content of the resin before the curing process. E1 emission class can be achieved, if the free formaldehyde content of the resin is lower than 0.2% by mass. Urea-formaldehyde resins containing. higher than 0.5% free formaldehyde by mass exceed emission class E2, and are not accepted. Since the free formaldehyde content of the urea-formaldehyde resin effects the emission of formaldehyde in the cured product, low formaldehyde content must be ensured during resin synthesis. This can be achieved by properly selecting synthesis conditions as well as raw materials. The quality of raw materials is an essential and determining factor for the synthesis of urea formaldehyde resins. The principal changes which may take place in the formaldehyde solution on storage are the polymerization and precipitation of the polymer, Cannizzaro reaction, methylal formation, oxydation to formic acid, condensation to hydroxyaldehydes and sugars. Any of these reactions are detrimental to product quality. The state of formaldehyde is also an essential factor, since the reaction of poly(methylene glycol)s with urea leads to methylene ether linkages resulting in emission of formaldehyde during storage and later on during the process of curing. Hydrolysis, isomerisation and decomposition of urea may take place simultaneously during improper storage conditions, such as high humidity, high temperatures, industrial atmosphere. The side products formed affect the reaction with formaldehyde during the synthesis resulting in high free formaldehyde content of urea-formaldehyde resins. The relation between synthesis conditions, free formaldehyde content and performance of urea-formaldehyde resins are discussed in detail. PMID- 10526994 TI - How to select a frail elderly population? A comparison of three working definitions. AB - Aim of this study was to compare three different working definitions for selecting a frail elderly population. Frailty was defined as inactivity combined with (1) low energy intake (n = 29), (2) weight loss (n = 26), or (3) low body mass index (n = 26). In the Zutphen Elderly Study (n = 450; age 69-89 years) differences in health, functioning, and diet in 1990 and functional decline and mortality in the following 3 years between "frail" and "nonfrail" participants, according to the working definitions, were studied using logistic regression analysis. Differences according to the inactivity/weight loss criterium were more pronounced than according to the other two criteria. Inactivity/weight loss was associated with lower subjective health and performance and more diseases and disabilities in 1990. Three-year relative risks of mortality (odds ratio [OR]: 4.1, 1.8-9.4) and functional decline (OR: 5.2, 1.04-25.8 for disabilities, OR: 3.7, 0.8-16.2 for performance) were higher as well. Inactivity in combination with weight loss seems a practicable working definition for selecting a frail elderly population. PMID- 10526995 TI - Validity of an activities of daily living questionnaire among older patients in the emergency department. AB - The objective of this study was to determine the validity of French and English versions of the Older American Resources and Services (OARS) activities of daily living (ADL) questionnaire using a premorbid reference period among older emergency department (ED) patients. A sample of 404 ED patients aged 65 and over participating in a study of functional decline was invited to participate in a clinical assessment shortly after their ED visit. The OARS ADL questionnaire was administered either to the patient or a proxy informant at the ED visit. The clinical assessment was conducted by a nurse, blind to the OARS score, using the Functional Autonomy Measurement System (SMAF). Disability scores for the OARS and SMAF were computed, based on the patient's premorbid status. Assessments were conducted in 213 patients (52.7%). The OARS summary scores, a total and an ordinal score, were highly correlated with the SMAF total disability score (Spearman's r of 0.80 and 0.79, respectively). Similar correlations were found for French and English versions. The OARS ADL questionnaire with a premorbid reference period appears to be valid when administered in the ED, both in French and English. PMID- 10526996 TI - Use of a clustered model to identify factors affecting hospital length of stay. AB - Predictive models have been used to identify factors that may prolong hospital length of stay (LOS). However, because predictors of LOS are collinear, the proportion of variance associated with each factor in a multivariate stepwise regression model may not reflect its mathematical contribution in explaining LOS. In an attempt to model factor contribution to LOS more realistically, we evaluated a clinically based clustered model. This model uses classes of candidate predictors, that is, patient attributes, adverse events, treatment modality, and health provider identity. Clusters of variables are permitted to enter into the model in a theoretically based predetermined sequence, so that the additional contribution of each cluster of factors can be assessed while the contribution of preceding factors is preserved. The clustered model was tested and compared with a free stepwise multivariate analysis in a cohort of patients undergoing prostatectomy for benign prostatic hypertrophy. We found that both models explained a similar proportion of the variance in LOS (56%-57%). However, some important differences were evident. Prostate size, associated with 12% of the variance in the clustered model, was not an independent predictor in the free model. A higher proportion of variance was associated with process variables, such as treatment modality in the free model. We conclude that use of a clustered model may facilitate more realistic assessment of the relative contribution of factors to LOS. PMID- 10526997 TI - Cross-cultural adaptation of a psychometric instrument: two methods compared. AB - Cross-cultural adaptations of questionnaires are needed in multilingual research, but little is known about the effectiveness of specific translation methods. We compared properties of two French-language adaptations of the SF36 health survey: (a) a rapid translation developed over 3 months in Geneva in 1992 (Geneva version), based on three initial translations, one synthesis, and two pretests, and (b) a comprehensive adaptation developed by the International Quality of Life Assessment Project between 1991 and 1994 (IQOLA version), which involved back translations, focus groups, development of equidistant response options, item difficulty and quality ratings, and multiple pretests. Wordings of 34 of 36 items differed. These two instruments were administered 1 year apart to the same sample of 946 young adults. Ceiling effects were somewhat lower for the IQOLA than for the Geneva version (means 30.4% and 35.5%), and missing scores slightly less frequent (IQOLA: mean 0.5%; Geneva: 1.2%). Floor effects (means 2.7% and 2.4%), proportions of consistent respondents (93.4% and 94.0%), and internal consistency coefficients (IQOLA: 0.78-0.89, Geneva: 0.80-0.92) were similar. Factor analysis supported the existence of two main aspects of health (physical and mental) for both versions. A majority of known-group comparisons were compatible with theory, for both versions. In conclusion, the two French-language versions of the SF36 had similar psychometric properties, despite extensive differences in the development process. This suggests that a moderately resource-intensive translation may produce adequate results. More empirical research is needed to understand what translation methods yield the best results. PMID- 10526998 TI - Are health states "timeless"? The case of the standard gamble method. AB - The standard gamble method, as currently recommended for use in health care program evaluation, provides an individual's preference score or "utility weight" for living in a given health state for the rest of the individual's life. Many researchers interpret this value as a time-independent or "timeless" one and order health states on a scale of zero (death) to one (full health), regardless of the time spent in the health state. This article examines whether preference scores for a severe pain health state are "timeless," or in other words whether the utility independence assumption is satisfied. Our study results suggest that for the majority of respondents, the preference scores are not independent of time. PMID- 10526999 TI - Relation of environmental tobacco smoke to diet and health habits: variations according to the site of exposure. AB - It has been postulated that the relationship of environmental tobacco smoke (ETS) exposure to cancer or cardiovascular diseases may be confounded by social class or diet because women exposed to ETS by their smoker spouse belong to lower social classes and have an unhealthy diet. In a population survey in Geneva, Switzerland, 914 female never-smokers were interviewed about sociodemographic factors, health habits including a semiquantitative food frequency questionnaire, and exposure to ETS according to the site (home, work, leisure). Compared to women unexposed to ETS, those exposed to ETS at work ate less fibers, cereals, vegetables, lean meat, had a lower intake of iron and beta-carotene, and had a lower total energy intake; women exposed during leisure time ate less cereals, drank less skim milk, and had a lower intake of complex carbohydrates. But the diet of women exposed at home did not differ from the diet of those unexposed to ETS. Thus, "living with a smoker" in Geneva does not necessarily imply adopting his health and dietary habits. We conclude that confounding factors of the association of ETS and disease vary according to site and populations and therefore should not be invoked as a systematic source of bias in all studies. PMID- 10527000 TI - Assessment of functional status, low back disability, and use of diagnostic imaging in patients with low back pain and radiating leg pain. AB - We analyzed data from outpatients with chronic low back pain (LBP) in the Veterans Health Study (n = 563) to examine the relationship between localized LBP intensity and radiating leg pain in assessing patient functional status, low back disability, and use of diagnostic imaging. Based on the localized LBP intensity, the study subjects were divided into tertiles (low, moderate, and high intensity). The study subjects were also stratified by the extent of radiating leg pain. Using analysis of variance and multiple regression analysis, we compared the relative importance of localized LBP intensity and radiating leg pain in explaining the variability in the means of the SF-36 scales and low back disability days, and in the proportion of patients who had used diagnostic imaging. The results of the study indicate that measures of localized LBP intensity and radiating leg pain contribute separately to the assessment of patient functional status, low back disability, and use of diagnostic imaging. These results suggest that localized LBP intensity and radiating leg pain may represent two different approaches in assessing back pain severity. Future epidemiological and health services research should consider both measures in assessing the impact of LBP on patient functional status, low back disability, and use of diagnostic imaging. PMID- 10527001 TI - The two by two diagram: a graphical truth table. AB - The two by two table is widely used in statistics, and in particular in the medical literature, to present the results of experimental and clinical studies in which two different operators (such as a reference standard and a new diagnostic test) sort a sample into two groups. The four cells of the table reflect the four possible categories of results. Despite the simplicity of the table, the interplay of its contents is surprisingly complex, and description of this interplay can be confusing if rendered with the conventional descriptive terms alone, such as sensitivity and predictive value. We present a graphical transformation of the table, comprised of a rectangular box in a special coordinate system. This diagram is offered as a flexible and subtle conceptual tool to help physicians, authors, and students to understand, plan, and present clinical research studies. PMID- 10527002 TI - Repeated significance tests on accumulating survival data. AB - The aim of the study was to compare the properties of two well-known group sequential methods and to demonstrate the effect of performing interim analyses on accumulating survival data without making appropriate adjustments of the nominal significance level. The properties of a group sequential method with fixed nominal significance level (Pocock stopping boundaries) and a method with increasing nominal level with each interim analysis (O'Brien-Fleming boundaries) were compared by stochastic simulation. Simulation experiments with survival times sampled from a breast cancer trial and from exponential distributions were performed. The true overall significance level with unplanned interim analyses increased from 5% to 14% when a maximum of five tests were performed. Both group sequential methods maintained the desired overall significance level. The O'Brien Fleming method had higher power than Pocock's method. It also reduced the risk of early stopping based on immature data and should usually be preferred. PMID- 10527004 TI - The influence of noncognitive factors on the Mini-Mental State Examination in older Mexican-Americans: findings from the Hispanic EPESE. Established Population for the Epidemiologic Study of the Elderly. AB - Mini-Mental State Examination data from the Hispanic Established Population for the Epidemiologic Study of the Elderly baseline survey, a population-based study of community-dwelling Mexican Americans aged 65 and older, were used to examine the relationship between cognitive impairment, sociodemographics, and health related characteristics. The rate of cognitive impairment found in this group of older Mexican Americans, using the conventional cut point of 23/24 on the MMSE, was 36.7%. Using a more conservative cut point of 17/18 indicated an overall rate of severe cognitive impairment of 6.7%. Rates of impairment varied significantly with age, education, literacy, marital status, language of interview, and immigrant status and were associated with high and moderate levels of depressive symptoms, and history of stroke. Importantly, although education was strongly related to poor cognitive performance, it was not a significant predictor of severe cognitive impairment. Multivariate analyses further indicated that as a screen for cognitive impairment in older Mexican Americans, the MMSE is strongly influenced by these noncognitive factors. Scores may reflect test bias, secondary to cultural differences or the level of education in this population. PMID- 10527003 TI - Gender-related differences in scores of the Barthel Index and Frenchay activities index in randomly sampled elderly persons living at home in Japan. AB - The purpose of this study was to examine for gender-related differences in activities of daily living (ADL) and lifestyle of elderly persons living at home, and to support our hypothesis that the gender-related difference in lifestyle of stroke patients derives from their lifestyle prior to the stroke. Participants were randomly sampled elderly persons living at home. Questionnaire sheets including subject profile, Self-Rating Barthel Index (disability index), and Self Rating Frenchay Activities Index (activity index) were mailed and collected, and the data were analyzed with the t-test and General Linear Model (factorial model with interaction). A total of 752 subjects were recruited, and their average age was 67.1 years. No significant gender-related differences were evident in the disability index including self-care and mobility domains (t-test, P > 0.05). In contrast gender-related differences in the activity index were significant (t test, P < 0.05) for three factors; gender, age group, and living conditions, and in a covariate disability index (GLM, P < 0.05). Because randomly selected elderly persons in this study exhibited a prominent gender-related difference in lifestyle, we believe the lifestyle difference in stroke patients that we have previously described derives primarily from their premorbid attitude to daily life. PMID- 10527005 TI - Cardiovascular drug prescriptions and risk of depression in diabetic patients. AB - Our aim was to investigate the association of calcium channel blocker (CCB), beta blocker, and ACE inhibitor medications with the risk of depression in diabetic patients. A case-control study was performed using an automated database (MediPlus, IMS) of 400 primary care practices in Germany including 972 diabetic cases with newly diagnosed depression in 1996 (index date) and 972 diabetic controls, matched for age, sex, and index date. The odds ratios (95%-confidence intervals) for depression, adjusted for type of practice, number of visits and prescriptions, hospitalization, cardiovascular diagnoses, and renal failure, were 2.2 (95% CI: 1.2-4.2) for exposure to CCB 6 months prior to index date, 2.6 (95% CI: 1.1-7.0) for beta-blockers, and 1.3 (95% CI: 0.8-2.2) for ACE inhibitors, respectively. Adjusted odds ratio for CCB (4.3; 95% CI: 1.7-13.5) and beta blockers (4.5; 95% CI: 1.2-29.5) were higher with daily dosages above the median. Prescriptions of CCB and beta-blockers among diabetic patients may increase the risk of depression. Because this association may alternatively be explained by cardiovascular comorbidity, further studies will be necessary to investigate the link between these cardiovascular medications and depression. PMID- 10527006 TI - Patient knowledge, awareness, and delay in seeking medical attention for malignant melanoma. AB - We investigated the relationship between patient knowledge, awareness, and delay in seeking medical attention for melanoma. The study population was comprised of 255 cases with cutaneous melanoma newly diagnosed during January 15, 1987 to May 15, 1989, who were part of a population-based case control study. Personal interviews were conducted to obtain information on patient's knowledge of melanoma signs and symptoms, skin awareness, delay in seeking medical attention, and related covariates. The adjusted odds ratio for the association between skin awareness and delay was 0.30 (95% confidence interval 0.12-0.71). Odds ratios ranged from 0.43 to 0.81 for knowledge and delay. Awareness of skin changes was associated with a reduced Breslow depth for stage I melanomas. Individuals who are aware of skin changes and abnormalities appear to be less likely to delay seeking medical attention for melanoma. Knowledge of melanoma signs and symptoms may also contribute to a decreased delay in melanoma diagnosis. PMID- 10527007 TI - The effects of lottery incentive and length of questionnaire on health survey response rates: a randomized study. AB - Maximizing the response rate of self-administered questionnaires is key in survey research. We aimed to evaluate the effects of lottery incentive and length of questionnaire on health survey response rates when used in isolation or combined. A random sample of 440 residents in Western Sydney, Australia was randomly allocated to four equal groups to receive or not receive an instant lottery ticket and a long (seven page) or short (one page) questionnaire. The overall response rate was 71.8%. The final response rates were higher among those receiving the short, rather than the long, questionnaire (75.6% versus 68.2%) (P = 0.08); and among those receiving the lottery incentive compared with those not receiving the incentive (75% versus 68.2%) (P = 0.09). By logistic regression analysis, the success of obtaining a completed questionnaire without any follow up reminders was significantly associated with the lottery incentive but not the questionnaire length (P = 0.03 and P = 0.54, respectively). The difference between lottery and no lottery groups decreased gradually during the follow-up. A lottery incentive is associated with an increased response after the first mailing. A small up-front cost for a lottery ticket may be worthwhile, since it can save further costs by obviating the need for repeated follow-ups. PMID- 10527008 TI - Pacemaker-related patient mortality. PMID- 10527009 TI - Incidence and significance of chronotropic incompetence in patients with indications for primary pacemaker implantation or pacemaker replacement. AB - This prospective study was undertaken to evaluate the incidence and significance of chronotropic incompetence in 211 patients [age 71.1 6 10.6 years (mean 6 SD)] by means of maximum exercise test in order to determine the indication for rate responsive pacing before primary pacemaker implantation (147 patients) or pacemaker replacement (64 patients). There were 112 (53%) patients with second- or third-degree AV block, 63 (30%) with sick sinus syndrome, and 36 (17%) with chronic atrial fibrillation. Chronotropic incompetence was defined as maximum heart rate lower than age-adjusted norm calculated by the formula: 0.7x(220 - age) and its significance as the difference between the two rates. The overall incidence of chronotropic incompetence was 42%. The incidence was significantly higher in patients with atrial fibrillation (67%, P<0.0005) and sick sinus syndrome (49%, P<0.012) than in those with AV block (30%). The mean difference between maximum heart rate and the age-adjusted norm was 18% (range 2%-63%). The mean difference was significantly higher in patients with atrial fibrillation (27%, range 8-63%) than in those with sick sinus syndrome (19%, range 2%-45%, P<0.01), or with AV block (12%, range 6%-26%, P<0.000001). The rate-responsive pacemakers were implanted in 44% of 211 patients studied and in 43% of 196 patients excluded from the study due to the apparent (contra)indication of rate responsive pacing (NS). Thus, chronotropic incompetence seems to be common in the pacemaker patient population. The highest incidence and significance was found in patients with chronic atrial fibrillation. Systematic evaluation of chronotropic competence can double the rate of implantation of rate-responsive pacemakers; however, further studies are needed to clarify relation between the significance of chronotropic incompetence and functional benefit of rate-responsive pacing. PMID- 10527010 TI - Value of magnetocardiographic QRST integral maps in the identification of patients at risk of ventricular arrhythmias. AB - It has been shown that regional ventricular repolarization properties can be reflected in body surface distributions of electrocardiographic QRST deflection areas (integrals). We hypothesize that these properties can be reflected also in the magnetocardiographic QRST areas and that this may be useful for predicting vulnerability to ventricular tachyarrhythmias. Magnetic field maps were obtained during sinus rhythm from 49 leads above the anterior chest in 22 healthy (asymptomatic) control subjects (group A) and in 29 patients with ventricular arrhythmias (group B). In each subject, the QRST deflection area was calculated for each lead and displayed as an integral map. The mean value of maximum was significantly larger in the control group A than in the patient group B (1,626+/ 694 pTms vs. 582+/-547 pTms, P<0.0001). To quantitatively assess intragroup variability in the control group A and intergroup variability of the control and patient groups, we used the correlation coefficient r and covariance sigma. These indices showed significantly less intragroup than intergroup variation (e.g., in terms of sigma, 28.0x10(-6)+/-12.3x10(-6) vs. 3.4x10(-6)+/-12.5x10(-6), P<0.0001). Each QRST integral map was also represented as a weighted sum of 24 basis functions (eigenvectors) by means of Karhunen-Loeve transformation to calculate the contribution of the nondipolar eigenvectors (all eigenvectors beyond the third). This percentage nondipolar content of magnetocardiographic QRST integral maps was significantly higher in the patient group B than in the control group A (13.0%+/-9.1 % vs. 2.6%+/-2.0%, P<0.0001). Discriminations between control subjects and patients with ventricular arrhythmias based on magnitude of the maximum, covariance sigma, and nondipolar content were 90.2%, 90.2%, and 86.3% accurate, with a sensitivity of 89.7%, 93.1%, and 75.9%, and a specificity of 90.9%, 86.4%, and 100%. We have shown that magnitude of the maximum and indices of variability and nondipolarity of the magnetocardiographic QRST integral maps may predict arrhythmia vulnerability. This finding is in agreement with earlier studies that used body surface potential mapping and suggests that magneticfield mapping may also be a useful diagnostic tool for risk analysis. PMID- 10527011 TI - Quality-of-life six months after CABG surgery in patients randomized to ICD versus no ICD therapy: findings from the CABG Patch Trial. AB - ICDs can affect a patient's perceived quality-of-life (QOL). This article describes the QOL in patients who participated in The CABG Patch Trial. This trial evaluated the potential benefit of empiric ICD implantation in patients with an increased risk of arrhythmic cardiac death as determined by reduced ejection fraction (<0.36) and an abnormal signal-averaged ECG. Patients were randomized to control (no ICD) or treatment (ICD) limbs. QOL was measured using the SF-36 and other measures among 490 (68%) of 719 patients available at 6-month follow-up. Analysis was performed on 228 control patients (those without ICDs) and 262 patients with ICDs. RESULTS: Six months after having CABG surgery, patients in the ICD group had lower levels of psychological well-being than those in the control group. In addition, compared to controls, patients whose ICDs had delivered therapy reported feeling less healthy, had reduced physical and emotional role functioning, and had lower levels of psychological well-being. CONCLUSION: Strategies aimed at easing patients' adjustment to ICDs should be developed and tested for efficacy in the setting of ICD prophylaxis. PMID- 10527012 TI - Results from the use of a preshaped lead for single-pass VDD/DDD stimulation. AB - Main criticisms about single-pass VDD stimulation in patients with AV block and normal sinus node function concern atrial undersensing in a lead with floating atrial electrodes, and loss of AV synchrony if sinus node dysfunction develops after implantation. We evaluated the concept of a preshaped single-pass lead designed to place the atrial ring electrodes in a constant position close to, or in contact with, the atrial wall. A preshaped lead (Model 2775, Medtronic Inc.) was implanted in 14 patients and followed for up to 2 years. Mean P wave amplitudes (PWAs) were 3.1 mV at implantation, 1.2 mV at predischarge, and 1.3 mV after 12 months. In all patients, minimal PWAs were well above maximal atrial sensitivity of the pacemaker in all body positions during the complete follow-up; atrial undersensing was not observed. Effective atrial stimulation was possible in all patients at implantation (mean stimulation threshold 2.5 V at 0.50 ms), in 11 patients on the first day after implant (mean stimulation threshold 0.22 ms at 5.0 V), in 10 patients after 1 month (mean stimulation threshold 0.57 ms at 5.0 V), and in 10 patients after 1 year (mean stimulation threshold 0.65 ms at 5.0 V). Intermittent phrenic nerve stimulation could be provoked in six patients. In conclusion, the concept of a preshaped single-pass lead facilitated implantation, improved atrial sensing performance, and allowed atrial stimulation in some patients. Still, further improvements are necessary to decrease the atrial stimulation thresholds to acceptable values in all patients. PMID- 10527013 TI - Paradigm shift in lead design. AB - During the past 30 years there has been a tremendous development in electrode technology from bulky (90 mm2) to pin-sized (1.0 mm2) electrodes. Simultaneously, impedance has increased from 110 Ohms to >1 kOhms, which has been termed a "paradigm shift" in lead design. If current is responsible for stimulation, why is its impedance a key factor in saving energy? Further, what mechanism is behind this development based on experimental findings and what conclusion can be drawn from it to optimize electrode size? If it is assumed that there is always a layer of nonexcitable tissue between the electrode surface and excitable myocardium and that the electric field (potential gradient) produced by the electrode at this boundary is reaching threshold level, then a formula can be derived for the voltage threshold that completely describes the electrophysiology and electrophysics of a hemispherical electrode. Assuming that the mean chronic threshold for porous steroid-eluting electrodes is 0.6 V with 0.5-ms pulse duration, thickness of nonexcitable tissue can be estimated to be 1.5 mm. Taking into account this measure and the relationship between chronaxie and electrode area, voltage threshold, impedance, and energy as a function of surface area can be calculated. The lowest voltage for 0.5-ms pulse duration is reached with r(o) = 0.5 d, yielding a surface area of 4 mm2 and a voltage threshold of 0.62 V, an impedance of 1 kOhms, and an energy level of 197 nJ. It can be deduced from our findings that a further reduction of surface areas below 1.6 mm2 will not diminish energy threshold substantially, if pulse duration remains at 0.5 ms. Lowest energy is reached with t = chronaxie, yielding an energy level <100 nJ with surface areas < or =1.5 mm2. It is striking to see how well the theoretically derived results correspond to the experimental findings. It is also surprising that the hemispheric model so accurately approximates experimental results with differently shaped electrodes that it can be concluded that electrode shape seems to play a minor role in electrode efficiency. Further energy reduction can only be achieved by reducing the pulse duration to chronaxie. A real paradigm shift will occur only if the fundamentals of electrostimulation in combination with electrophysics are accepted by the pacing community. PMID- 10527014 TI - Spectrotemporal mapping of high-resolution ECGs in experimental myocardial infarction: comparison with time-domain analysis and epicardial electrograms. AB - The study was undertaken to evaluate the relationship of signal-averaged ECG (SA ECG) readings in the frequency domain (STM) and epicardial electrograms (EE) recorded before and after acute myocardial infarction (AMI) in pigs and to compare the changes with findings in time-domain analysis (TDA). In 20 pigs the left anterior descending artery (LAD) was ligated. Prior to ligation, a SA-ECG was recorded (method of Simson) and bipolar electrodes were used to register EE in the areas supplied by the LAD and the circumflex artery (CIRC). Five minutes after LAD ligation, all measurements were repeated. Time-domain parameters were QRS duration (QRS D) and the duration of the signal below 30 microV (LAS 30). Beginning at a point of 20 ms before the QRS end, the frequency spectra (0-200 Hz) of 25 segments of 80-ms duration at the QRS end were analyzed. The volumes below the 25 curves were analyzed separately for 0-50 Hz, 51-100 Hz, 101-150 Hz, and 151-200 Hz. After AMI, five pigs died within 7 minutes. In 15 pigs, QRS D as well as LAS 30 lengthened significantly (P<0.05; P<0.001). Spectrotemporal mapping (STM) showed a significant decrease of the frequencies above 50 Hz (51 200 Hz) in the entire group and in the animals with late potentials (P<0.05). EE of the LAD area were significantly prolonged (P<0.01); this did not correlate with the changes in STM parameters. In pigs acute myocardial infarction causes a shift towards lower frequencies in the STM which most likely reflects the slowed depolarisation in the ischemic area. PMID- 10527015 TI - Usefulness of echocardiography to predict inappropriate atrial sensing in single lead VDD pacing. AB - Reliable atrial sensing is the prerequisite for restoration of atrioventricular synchrony in patients with single-lead VDD pacing systems. To determine echocardiographic variables associated with inappropriate atrial sensing, 21 consecutive patients with symptomatic second- or third-degree AV block and normal sinus node function were studied. Prior to implantation echocardiographic measurements of end-systolic and end-diastolic dimensions and volumes of the right atrium and right ventricle were performed. All patients underwent implantation of a Medtronic Thera VDD(d) pacemaker with a bipolar Medtronic Capsure electrode. A minimal amplitude of the unfiltered atrial electrocardiogram of > or =0.5 mV was required for permanent lead position and the atrial sensitivity was programmed below the lowest recorded value. Appropriate atrial sensing (atrial triggered ventricular paced complexes/total number of ventricular paced complexes) was assessed during 24-hour Holter monitoring and treadmill exercise testing 3 to 6 weeks after implantation. Inappropriate atrial sensing (<95% correct atrial synchronization during Holter registration and/or <97.5% during exercise testing) was present in nine patients. Right atrial volumes and the right ventricular end-diastolic volume was significantly higher, as compared to patients without inappropriate sensing (12 patients). The right atrial and diastolic volumes had the highest correlation with correct atrial sensing r = 0.83, P<0.0001). Using a postdefined cut-off value of > or =80 mL for the end diastolic right atrial volume, sensitivity and specificity for inappropriate sensing was 100% and 92%, respectively. These findings show that preimplant echocardiography can identify patients with inappropriate sensing during VDD pacing, in whom DDD pacing should be considered. PMID- 10527016 TI - Intravascular extraction of problematic or infected permanent pacemaker leads: 1994-1996. U.S. Extraction Database, MED Institute. AB - Of the 400,000-500,000 permanent pacemaker leads implanted worldwide each year, around 10% may eventually fail or become infected, becoming potential candidates for removal. Intravascular techniques for removing problematic or infected leads evolved over a 5-year period (1989-1993). This article analyzes results from January 1994 through April 1996, a period during which techniques were fairly stable. Extraction of 3,540 leads from 2,338 patients was attempted at 226 centers. Indications were: infection (27%), nonfunctional or incompatible leads (25%), Accufix or Encore leads (46%), or other causes (2%). Patients were 64+/-17 years of age (range 5-96); 59% were men, 41% women. Leads were implanted 47+/-41 months (maximum 26 years), in the atrium (53%), ventricle (46%), or SVC (1%). Extraction was attempted via the implant vein using locking stylets and dilator sheaths, and/or transfemorally using snares, retrieval baskets, and sheaths. Complete removal was achieved for 93% of leads, partial for 5%, and 2% were not removed. Risk of incomplete or failed extraction increased with implant duration (P<0.0001), less experienced physicians (P<0.0001), ventricular leads (P<0.005), noninfected patients (P<0.0005), and younger patients (P<0.0001). Major complications were reported for 1.4% of patients (<1% at centers with >300 cases), minor for 1.7%. Risk of complications increased with number of leads removed (P<0.005) and with less experienced physicians (P<0.005); risk of major complications was higher for women (P<0.01). Given physician experience, appropriate precautions, and appropriate patient selection, contemporary lead removal techniques allow success with low complication rates. PMID- 10527017 TI - Usefulness of intravenous propofol anesthesia for radiofrequency catheter ablation in patients with tachyarrhythmias: infeasibility for pediatric patients with ectopic atrial tachycardia. AB - General anesthesia is sometimes required during radiofrequency catheter ablation (RFCA) of various tachyarrhythmias because of an anticipated prolonged procedure and the need to ensure stability during critical ablation. In this study, we examine the feasibility of using propofol anesthesia for RFCA procedure. There were 150 patients (78 male, 72 female; mean age 30 years, range 4-96 years) in the study. Electrophysiologic study was performed before and during propofol infusion in the initial 20 patients and was performed only during propofol infusion in the remaining 130 patients. In the initial 20 patients, propofol infusion increased the sinus rate and facilitated AV nodal conduction. The accessory pathway effective refractory period, as well as the sinus node recovery time, atrial effective refractory period, and ventricular effective refractory period were not significantly changed. There were 152 tachyarrhythmias in 150 patients (24 atrial flutter, 31 AV nodal reentrant tachycardia, 68 AV reciprocating tachycardia, 12 ventricular tachycardia, and 17 atrial tachycardia). Most (148/152) tachycardias remained inducible after anesthesia and RFCA was performed uneventfully. However, in four of the seven pediatric patients with ectopic atrial tachycardia, the tachycardia terminated after propofol infusion and could not be induced by isoproterenol infusion. Consequently, RFCA could not be performed. Intravenous propofol anesthesia is feasible during RFCA for most tachyarrhythmias except for ectopic atrial tachycardia in children. PMID- 10527018 TI - Prediction of optimal atrioventricular delay in patients with implanted DDD pacemakers. AB - In patients with an implanted DDD pacemaker (PM), the atrial contribution may be interrupted by too short an atrioventricular (AV) delay, and filling time may be shortened by too long an AV delay. The AV delay at which the end of the A wave on transmitral flow coincides with complete closure of the mitral valve may be optimal. The subjects were 15 patients [70.3+/-12.3 (SD) years old] with an implanted DDD PM. Cardiac output (CO) and pulmonary capillary wedge pressure (PCWP) were measured by Swan-Ganz catheter. Transmitral flow was recorded by pulsed Doppler echocardiography. AV delay was prolonged stepwise by 25 msc. When the AV delay was set at 155+/-26 ms, the end of the A wave coincided with complete closure of the mitral valve. When the AV delay was prolonged 25, 50, 75, and 100 ms from this AV delay, the interval between the end of the A wave and complete closure of mitral the valve was prolonged 16+/-5, 39+/-6, 65+/-4 and 88+/-5 ms, respectively (r = 0.97, P<0.0001) and diastolic mitral regurgitation was observed during this period. Thus, the optimal AV delay may be predicted as follows: the slightly prolonged AV delay minus the interval between the end of the A wave and complete closure of the mitral valve. When the AV delay was set at 215 ms, there was a significant positive correlation between the predicted optimal AV delay (166+/-23 ms) and the optimal AV delay (CO: 161+/-26 msec, r = 0.93, P<0.0001, PCWP: 161+/-28 msec, r = 0.95, P<0.0001). In conclusion, optimal AV delay can be predicted by this simple formula: slightly prolonged AV delay minus the interval between end of A wave and complete closure of mitral valve at the AV delay setting. PMID- 10527019 TI - Altered cardiac histology following apical right ventricular pacing in patients with congenital atrioventricular block. AB - Previous studies have demonstrated that right ventricular apical pacing inherently alters ventricular contraction, regional blood flow, wall stress, and predisposes to diminished function. However, histological consequences of chronic apical pacing potentially contributing to the observed ventricular dysfunction remain conjectural. Previous canine studies have demonstrated histopathological cellular abnormalities with apically initiated ventricular pacing that may result in the observed diminished ventricular function. To determine if comparable adverse changes also occur in the clinical setting, 16 endomyocardial biopsies were obtained from 14 age-matched patients with congenital complete atrioventricular block (CCAVB) and otherwise normal anatomy, divided into two groups: eight biopsies (median patient age 15.5 years) from patients prior to pacemaker implant and another eight biopsies (median patient age 16 years) from patients following 3-12 years (median 5.5) of chronic ventricular pacing. In one patient, biopsy samples were obtained before and after pacing. Results demonstrated a significant (P<0.05) increase in histopathological alterations among the patient biopsy samples following pacing, consisting of myofiber size variation, fibrosis, fat deposition, sclerosis, and mitochondrial morphological changes. These findings indicate that chronic apical right heart ventricular pacing may adversely alter myocellular growth, especially among the young, on the cellular and subcellular level, potentially contributing to the diminished function observed clinically. PMID- 10527020 TI - Effects of adenosine on local stimulus-response latency and induction of atrial fibrillation by premature stimuli. AB - Premature atrial stimuli delivered during the relative refractory or "vulnerable" period exhibit increased local stimulus-response latency and may occasionally induce atrial arrhythmias. The use of adenosine to treat supraventricular tachycardias may also provoke atrial arrhythmias. In this study we investigated the effects of adenosine on the latency of premature complexes in relation to repolarization and induction of atrial arrhythmias in 14 patients without structural heart disease. A monophasic action potential catheter was used for recording in the right atrium and introducing premature stimuli (S2) at twice diastolic threshold after eight paced (S1) complexes. At short coupling intervals, S2 latency increased relative to S1 latency. S2 was delivered repeatedly at a fixed coupling interval (producing maximal local response latency) and adenosine (6 mg) was given intravenously. Adenosine decreased S2 latency significantly (23+/-5 to 11+/-3 ms, P<0.01), to values similar to S, latency. However, despite the decrease in S2 latency, the combination of adenosine and S2 more often resulted in transient atrial arrhythmias (11 of 14 patients vs 2 of 14 patients without adenosine, P<0.05). Adenosine had no effect on S, latency (9+/-2 vs. 9+/-2 ms) but decreased monophasic action potential duration from 202+/-37 to 158+/-38 ms (P<0.01). Adenosine was also given to 10 patients with S2 introduced at a coupling interval 40-50 ms less than the baseline effective refractory period. This resulted in a decrease in atrial refractoriness and capture of S2 in all cases. Latency for S2 was significantly greater than Si latency (21+/-12 vs. 9+/-2 ms, P<0.01) and transient atrial arrhythmias were induced in 9 of 10 patients. We conclude that for a given S2 coupling interval, adenosine decreases local stimulus-response latency but increases atrial vulnerability to transient atrial arrhythmias. Decreased latency may be related to a shift in the zone of relative refractoriness associated with an adenosine-mediated decrease in monophasic action potential duration. Induction of atrial arrhythmias in the presence of adenosine occurs independently of increased latency and is therefore not dependent on S2 falling within the relative refractory period at the site of stimulation. PMID- 10527021 TI - The costs of recurrent syncope of unknown origin in elderly patients. AB - Although syncope has been shown to reduce quality-of-life, its impact on resource costs has not been documented. The objective of this study was to quantify the annual medical costs of caring for elderly patients with syncope, especially recurrent syncope of unknown origin. Administrative data from the Health Care Financing Administration were obtained on 7,959 Medicare patients who had at least one inpatient admission with a diagnosis of syncope in 1993. The costs of any inpatient admissions, outpatient procedures, or physician visits with an ICD CM-9 diagnosis for syncope were summed for a 365-day period from the date of the initial hospitalization for syncope. Patients who had at least two hospitalizations with admission and discharge diagnosis of syncope were deemed to have recurrent syncope of uncertain origin. To better estimate syncope costs for those whose syncope costs could have been attributed to other diagnoses, a regression analysis was performed including variables representing the most frequent secondary diagnoses. The average annual costs of those who were admitted with syncope but who were discharged with another diagnosis was $4,942 in 1993. The average annual cost of patients with recurrent syncope deemed to be of unknown origin was $5,165. For those patients with secondary diagnoses of atherosclerosis, urinary tract infections, or hypokalemia, the annual costs of syncope averaged $6,820, $7,013, or $7,949, respectively. PMID- 10527022 TI - 2:1 atrioventricular block during tachycardia. PMID- 10527024 TI - Biomaterials Access Assurance Act of 1998: a legislative success. PMID- 10527023 TI - Comparison of formulae for heart rate correction of QT interval in exercise electrocardiograms. AB - The study investigated the differences in five different formulae for heart rate correction of the QT interval in serial electrocardiograms recorded in healthy subjects subjected to graded exercise. Twenty-one healthy subjects (aged 37+/-10 years, 15 male) were subjected to graded physical exercise on a braked bicycle ergometer until the heart rate reached 120 beats/min. Digital electrocardiograms (ECG) were recorded on baseline and every 30 seconds during the exercise. In each ECG, heart rate and QT interval were measured automatically (QT Guard package, Marquette Medical Systems, Milwaukee, WI, USA). Bazett, Fridericia, Hodges, Framingham, and nomogram formulae were used to obtain QTc interval values for each ECG. For each formula, the slope of the regression line between RR and QTc values was obtained in each subject. The mean values of the slopes were tested by a one-sample t-test and the comparison of the baseline and peak exercise QTc values was performed using paired t-test. Bazett, Hodges, and nomogram formulae led to significant prolongation of QTc intervals with exercise, while the Framingham formula led to significant shortening of QTc intervals with exercise. The differences obtained with the Fridericia formula were not statistically significant. The study shows that the practical meaning of QT, interval measurements depends on the correction formula used. In studies investigating repolarization changes (e.g., due to a new drug), the use of an ad-hoc selected heart rate correction formula is highly inappropriate because it may bias the results in either direction. PMID- 10527025 TI - Anomalous "VVI" pacing of a dual chamber pacemaker when programmed in DDI(R) mode. AB - We describe the case of a dual chamber rate responsive pacemaker (Relay, model 294-03, Intermedics, Angleton, TX, USA) implanted in a 68-year-old male for sick sinus syndrome, which was not working properly when programmed in the DDIR mode, thus determining occasionally a sort of "VVI" pacing. However, the pacemaker performed well when programmed in the DDDR mode. We discovered that this was not a malfunction of a single device but rather a general behavior of this family of Intermedics dual chamber pacemakers (also not rate responsive), caused by a software problem. PMID- 10527026 TI - Transesophageal echocardiographic visualization of left ventricular malpositioned pacemaker electrodes: implications for lead extraction procedures. AB - Two cases of malpositioned LV electrodes are presented. Transesophageal echocardiography (TEE) allowed for a careful inspection of left-sided leads and for tracking their course. One LV and two right-sided leads were safely retrieved with TEE monitoring. One chronic LV lead was left in place as it was heavily fibrosed. TEE was helpful in the inspection and monitoring of the extraction and also in guided traction efforts. This is the first published report of echocardiographic visualization of the lead retrieval procedure. PMID- 10527027 TI - Association between nonreentrant supraventricular tachycardia and atrioventricular node reentrant tachycardia: a presentation of dual AV node physiology. AB - Persistent simultaneous conduction of P waves over a fast and a slow nodal pathway defines the nonreentrant type of supraventricular tachycardia, usually not associated with reciprocating movements. We report a unique association between this uncommon tachycardia and a usual AV nodal reentrant tachycardia, made possible by the existence of three different nodal pathways. PMID- 10527028 TI - Supraventricular electrical interaction in conjoined twins with common coronary sinus. AB - Conjoined twins with echocardiographic evidence of continuity of the coronary sinuses had identical heart rates on ECG. Both had broad, polyphasic QRS complexes, and various imaging modalities were unable to determine whether there was ventricular myocardial continuity. Administration of adenosine demonstrated that the broad polyphasic complexes were a "fusion" of the twins' individual QRS complexes, which could be clearly distinguished after administration of the drug. Ventricular pacing resulted in dissociation of the individual QRS complexes, thus demonstrating an absence of ventricular myocardial continuity. This was confirmed when the twins were successfully separated at the age of 10 months. PMID- 10527029 TI - Twiddler's syndrome with transvenous defibrillators in the pectoral region. AB - Advances in technology have enabled the implantation of defibrillators in the pectoral region. Complications encountered with pacemakers may also be observed with defibrillators. We describe two cases of twiddler's syndrome in patients with defibrillators implanted subcutaneously in the left pectoral region. PMID- 10527030 TI - The role of hospitalists in medical education. PMID- 10527031 TI - Association of panic disorder and panic attacks with hypertension. AB - PURPOSE: Previous studies of the association between hypertension and panic disorder were uncontrolled or involved small numbers of patients. PATIENTS AND METHODS: We compared the prevalence of panic disorder and panic attacks in 351 patients with documented hypertension who were randomly selected from all hypertensive patients registered in one primary care practice with age- and gender-matched normotensive patients from the same practice and with hypertensive patients attending a hospital clinic. All three groups completed questionnaires for panic disorder based on standard criteria, as well as the Hospital Anxiety and Depression scale. RESULTS: The prevalence of current (previous 6 months) panic attacks was significantly greater in primary care patients with hypertension (17%, P <0.05) and hospital-based hypertensive patients (19%, P <0.01) than in normotensive patients (11%). Similar results were seen for lifetime panic attacks (35% versus 39% versus 22%; both P for comparisons with normotensive patients <0.001). The prevalence of panic disorder was significantly greater in primary care patients with hypertension (13%) than normotensive patients (8%, P <0.05). Anxiety scores were significantly higher in both hypertensive groups than in normotensive patients. Depression scores were significantly higher in hospital-based hypertensive patients than in the other two groups. The reported diagnosis of hypertension antedated the onset of panic attacks in a large majority of patients (P <0.01). CONCLUSIONS: Physicians caring for patients with hypertension should be aware of the significantly greater prevalence of panic attacks in these patients. PMID- 10527032 TI - Older persons' perceptions of home and hospital as sites of treatment for acute illness. AB - PURPOSE: Home care is increasingly being used as a substitute for hospital care. This study examined older patients' perceptions of the home and of the hospital as treatment sites for acute illness and the patient characteristics that are associated with these perceptions. SUBJECTS AND METHODS: A series of questions derived from open-ended interviews supplemented by literature review were administered by telephone in a cross-sectional, descriptive study to community dwelling persons age 65 years or older who had been hospitalized 2 months earlier with congestive heart failure, chronic obstructive pulmonary disease, or pneumonia. RESULTS: Among 246 participants, nearly equal proportions agreed with statements that the home and the hospital would be comfortable sites of care (54% versus 55%), that the home and the hospital would provide rapid recovery (41% versus 37%), and that home treatment and hospital treatment would be burdensome on family and friends (40% versus 33%). Although 93% would feel safe in the hospital, only 42% would feel safe at home. Perceptions were not associated with sociodemographic characteristics, primary diagnosis, self-rated health, depression, or social support. Functionally dependent patients had more positive perceptions of treatment at home. CONCLUSIONS: Evaluation of perceptions of home and hospital can facilitate assessing the acceptability of shifting acute care from hospital to home. Our findings suggest that successful expansion of acute home care will require flexibility in the use of home and hospital as well as education to change perceptions about the safety and efficacy of treatment at home. PMID- 10527033 TI - Use of critical pathways to improve the care of patients with acute myocardial infarction. AB - PURPOSE: While critical pathways have become a popular strategy to improve the quality of care, their effectiveness is not well defined. The objective of this study was to investigate the effect of a critical pathway on processes of care and outcomes for Medicare patients admitted with acute myocardial infarction. SUBJECTS AND METHODS: A retrospective cross-sectional and longitudinal cohort study was made of Medicare patients aged 65 years and older hospitalized at 32 nonfederal Connecticut hospitals with a principal diagnosis of myocardial infarction during two periods: June 1, 1992, to February 28, 1993, and August 1, 1995, to November 30, 1995. The main endpoints of the cross-sectional analyses for the 1995 cohort were the proportion of patients without contraindications who received evidence-based medical therapies, length of stay, and 30-day mortality. Hospitals with specific critical pathways for patients with myocardial infarction were compared with hospitals without critical pathways. The main endpoints of the longitudinal analyses were change between 1992-93 and 1995 in the proportion of patients receiving evidence-based medical therapies, length of stay, and 30-day mortality. RESULTS: Ten hospitals developed critical pathways between 1992-93 and 1995. Eighteen of 22 nonpathway hospitals employed some combination of standard orders, multidisciplinary teams, or physician champions. Patients admitted to hospitals with critical pathways did not have greater use of aspirin within the first day, during hospitalization, or at discharge; beta-blockers within the first day or at discharge; reperfusion therapy; or use of angiotensin-converting enzyme inhibitors at discharge in 1995. The mean (+/- SD) length of stay in 1995 was not significantly different between pathway (7.8 +/- 4.6 days) versus nonpathway hospitals (8.0 +/- 4.2 days), and the change in length of stay between 1992-93 and 1995 was 2.2 days for pathway hospitals and 2.3 days for nonpathway hospitals. Patients admitted to critical pathway hospitals had lower 30-day mortality in 1995 (8.6% versus 11.6% for nonpathway hospitals, P = 0.10) and in 1992-93 (12.6% versus 13.8%, P = 0.39), but the differences were not statistically significant. CONCLUSIONS: Hospitals that instituted critical pathways did not have increased use of proven medical therapies, shorter lengths of stay, or reductions in mortality compared with other hospitals that commonly used alternative approaches to quality improvement among Medicare patients with myocardial infarction. PMID- 10527034 TI - Adult health status of women with histories of childhood abuse and neglect. AB - PURPOSE: Several recent studies have found associations between childhood maltreatment and adverse adult health outcomes. However, methodologic problems with accurate case determination, appropriate sample selection, and predominant focus on sexual abuse have limited the generalizability of these findings. SUBJECTS AND METHODS: We administered a survey to 1,225 women who were randomly selected from the membership of a large, staff model health maintenance organization in Seattle, Washington. We compared women with and without histories of childhood maltreatment experiences with respect to differences in physical health status, functional disability, numbers and types of self-reported health risk behaviors, common physical symptoms, and physician-coded ICD-9 diagnoses. RESULTS: A history of childhood maltreatment was significantly associated with several adverse physical health outcomes. Maltreatment status was associated with perceived poorer overall health (ES = 0.31), greater physical (ES = 0.23) and emotional (ES = 0.37) functional disability, increased numbers of distressing physical symptoms (ES = 0.52), and a greater number of health risk behaviors (ES = 0.34). Women with multiple types of maltreatment showed the greatest health decrements for both self-reported symptoms (r = 0.31) and physician coded diagnoses (r = 0.12). CONCLUSIONS: Women with childhood maltreatment have a wide range of adverse physical health outcomes. PMID- 10527035 TI - The value of chest roentgenography in the diagnosis of pneumothorax after thoracentesis. AB - PURPOSE: We sought to assess the yield of chest roentgenography for the detection of pneumothorax among hospitalized patients with pleural effusion who have undergone diagnostic or therapeutic thoracentesis. SUBJECTS AND METHODS: We performed a prospective study of 506 thoracentesis procedures in 370 patients. After the procedure, each operator filled out a note recording patient data and the characteristics of the thoracentesis. A chest radiograph was performed within 12 hours after the procedure in all patients. RESULTS: Eighteen (4%) pneumothoraces occurred in 17 patients, 9 (2%) of which required chest tube drainage. Of the 488 patients without symptoms, only 5 (1%) developed a pneumothorax, only 1 of which required chest tube drainage. By contrast, of the 18 patients with symptoms, 13 developed a pneumothorax, 8 of which required chest tubes. There were two independent predictors of pneumothorax: presence of symptoms (odds ratio [OR] = 250; 95% confidence interval [CI]: 65 to 980) and male gender (OR = 5.4; 95% CI: 1.9 to 69). CONCLUSIONS: Among the symptom-free patients in our sample, the risk of developing pneumothorax with clinical consequences was so low that the practice of routine chest roentgenography may not be justified. PMID- 10527036 TI - Local insulin infusion stimulates expression of plasminogen activator inhibitor-1 and tissue-type plasminogen activator in normal subjects. AB - PURPOSE: Plasma levels of plasminogen activator inhibitor-1 are increased in obesity, hypertension, and diabetes. Their correlation with insulin levels supports the hypothesis that hypofibrinolysis may affect the development of atherosclerotic complications in patients with insulin resistance. To investigate the effect of insulin on fibrinolysis, we evaluated levels of plasminogen activator inhibitor-1 (PAI-1) and tissue plasminogen activator (tPA) antigens during insulin infusion in the forearm vascular beds of 8 healthy subjects. MATERIALS AND METHODS: Insulin was infused in the brachial artery of each subject to raise local venous concentrations to approximately 100 microU/mL. Blood samples were obtained from the brachial artery and vein at baseline, after 30, 60, 90, and 120 minutes of infusion, and 30 minutes after the end of the infusion. RESULTS: Following intra-arterial infusion of insulin, forearm blood flow (mean +/- SD) increased progressively from 2.7 +/- 0.6 to 4.0 +/- 0.6 mL/dL/min (P <0.01) and did not return to baseline after the end of the infusion. Plasminogen activator inhibitor-1 balance increased (345 +/- 160 versus 8 +/- 152 fmol/dL/min, P <0.02) at 60 minutes, reaching baseline levels after the end of the infusion. After 90 minutes, tPA balance increased (40 +/- 26 versus 7 +/- 29 fmol/dL/min, P <0.01) with a profile similar to forearm blood flow. CONCLUSIONS: Local hyperinsulinemia induces regional vasodilation and expression of PAI-1 and tPA antigens. An alteration of this physiological process could be involved in the development of hypofibrinolysis and atherosclerosis in states of insulin resistance. PMID- 10527037 TI - Foreign body asphyxiation--an autopsy study. AB - PURPOSE: Food asphyxiation is a common problem whenever and wherever people eat. A knowledge of predisposing factors might help to prevent this problem. SUBJECTS AND METHODS: We reviewed 34,476 consecutive autopsies done during a 14-year period (1984 to 1997) at the Institute of Forensic Medicine, Vienna. Demographic features and predisposing factors were determined for the 191 cases of fatal foreign body asphyxiation. RESULTS: Old age, poor dentition, and alcohol consumption were frequent findings. Other risk factors included chronic disease, sedation, and eating risky foods. On 120 (63%) of the 191 occasions, observers were present at the time of the incident and subsequently called the Emergency Service. In 110 (92%) cases, neither the observers nor the majority of the emergency medical technicians and physicians who would have been able to intervene recognized the definite diagnosis. Only 10 cases were correctly identified during cardiopulmonary resuscitation. CONCLUSIONS: These fatal accidents could be prevented easily. Effective prevention depends on understanding the nature and frequency of accidental deaths due to asphyxiation and the factors that lead to their occurrence and having a high degree of suspicion. PMID- 10527038 TI - The risk of serious cardiac arrhythmias among cisapride users in the United Kingdom and Canada. AB - PURPOSE: Serious, although rare, ventricular arrhythmias and deaths have been reported in patients taking cisapride monohydrate. Without quantification of the risk involved, it is impossible to develop rational therapeutic guidelines. SUBJECTS AND METHODS: Arrhythmic events (sudden deaths and other events compatible with serious ventricular arrhythmias) were sought among 36,743 patients prescribed cisapride in the United Kingdom and Saskatchewan, Canada. Prescriptions and cases were identified from computerized medical claims data and physicians' office records. We compared rates of events between periods of recent cisapride use and nonrecent use, using cohort analysis. Potential confounding factors, including concomitant treatment with agents that inhibit CYP3A4 metabolism or that prolong the QT interval, were assessed in a nested case control study. RESULTS: In the cohort analysis, the incidence of the arrhythmic events was 1.6 times greater (95% confidence interval [CI]: 0.9 to 2.9) in periods of recent use. With adjustment for clinical history, use of CYP3A4 inhibitors, and use of drugs that prolong the QT interval, the odds ratio for cisapride and cardiac outcomes was 1.0 (95% CI: 0.3 to 3.7). There was no identifiable increase in risk when cisapride was dispensed at about the same time as QT-prolonging drugs or CYP3A4 inhibitors. QT-prolonging agents were associated with a 2.5-fold increase in the risk of arrhythmic events (95% CI: 1.1 to 5.8). CONCLUSIONS: Serious rhythm disorders were not associated with cisapride use, although the upper confidence bounds do not rule out an increase in risk. PMID- 10527039 TI - A meta-analysis of the effects of ipratropium bromide in adults with acute asthma. AB - PURPOSE: To review the literature to determine whether inhaled ipratropium bromide provides additive benefits to adults with acute asthma who are being treated with beta-agonists in an emergency department. SUBJECTS AND METHODS: English-language studies, both published (1978 to 1999) and unpublished, were retrieved using Medline, Science Citation Index, Current Contents, bibliographic reviews of primary research, review articles, consultation with experts, and the register of Medical Editors' Trial Amnesty. Only randomized, double-blind, controlled trials that enrolled patients having an exacerbation of asthma were included. The main outcome measure was pulmonary function; hospital admission rate was also evaluated. RESULTS: Ten studies including 1,483 adults with acute asthma were selected (mean age 32 +/- 13 years, 36% men). The overall effect size in SD units of pulmonary function showed a significant benefit from ipratropium (effect size 0.14, 95% confidence interval [CI]: 0.04 to 0.24, P = 0.008). Study specific effect sizes ranged from 0.03 to 0.63. This pooled effect size was equivalent to a 10% (95% CI: 2% to 18%) increase in forced expiratory volume in 1 second (FEV1) or peak expiratory flow in the ipratropium group compared with the control group. Analysis of the four studies that included patients with extreme obstruction (FEV1 or peak flow <35% of predicted at presentation) showed substantial improvement with ipratropium therapy (effect size 0.38, 95% CI: 0.09 to 0.67). In the five trials (1,186 patients) that studied the effect of ipratropium administration on hospital admissions, pooled results revealed that ipratropium reduced admission rates significantly (odds ratio 0.62, 95% CI: 0.44 to 0.88, P = 0.007). CONCLUSIONS: The addition of ipratropium to beta-agonist therapy offers a statistically significant, albeit modest, improvement in pulmonary function, as well as a reduction in the rate of hospital admissions. PMID- 10527040 TI - Cell adhesion molecules and the glomerulopathies. PMID- 10527041 TI - Low-dose dopamine does not prevent acute renal failure in patients with septic shock and oliguria. NORASEPT II Study Investigators. PMID- 10527042 TI - Bisoprolol-induced rapid eye movement sleep behavior disorder. PMID- 10527043 TI - Hypersensitivity pneumonitis induced by intranasal heroin use. PMID- 10527044 TI - Et tu, critical pathways? PMID- 10527045 TI - The lifelong legacy of childhood abuse. PMID- 10527046 TI - Adult respiratory distress syndrome due to pheochromocytoma as the initial presentation of multiple endocrine neoplasia type IIA syndrome. PMID- 10527047 TI - Autoimmune chronic gastritis and iron deficiency anemia. PMID- 10527049 TI - The mineral content of bottled water and other beverages: implications for health and disease. PMID- 10527048 TI - A patient with chronic lymphoid leukemia and recurrent necrotic herpetic lymphadenitis. PMID- 10527050 TI - Lumbar puncture complicated by retroperitoneal hemorrhage. PMID- 10527051 TI - Generalist versus pulmonologist care for severe chronic obstructive pulmonary disease. PMID- 10527052 TI - Effective management of gout. PMID- 10527053 TI - Thyroid disease mediated by molecular defects in cell surface and nuclear receptors. PMID- 10527054 TI - Spontaneous multivessel coronary artery dissection in a pregnant woman treated successfully with stent implantation. PMID- 10527055 TI - Source memory and divided attention: reciprocal costs to primary and secondary tasks. AB - Source memory, in comparison with item memory, is more sensitive to frontal lesions and may require more strategic processing. Divided attention was used to restrict attentional resources and strategic processing on memory tasks. Participants encoded and retrieved items (i.e., words) and source (i.e., voice or spatial location) while concurrently performing a finger-tapping (FT) or visual reaction-time (VRT) task. Memory accuracy costs under divided attention were greater for retrieval of source than item and were greater with VRT than FT. Similarly, costs to the secondary task were greater when concurrently retrieving source as opposed to item and were greater for VRT than FT. Effects were stronger when spatial location was used as the source task. Findings support the idea that processing source information requires more attentional resources and effort than processing item information. Furthermore, concurrent performance of VRT produced greater interference with a task that was more dependent on intact frontal functioning and better simulated the performance of patients with frontal dysfunction. PMID- 10527056 TI - Effects of bilateral stimulation and stimulus redundancy on interhemispheric interaction. AB - Recent visual laterality studies have included trials in which critical stimulus information is presented simultaneously in both visual half-fields and, thereby, simultaneously to both cerebral hemispheres. To investigate interhemispheric interaction, researchers compare performance on bilateral redundant trials with performance on unilateral trials in which a single copy of the target is presented to one hemisphere or the other. The authors used the identification of nonword letter trigrams to examine the relationship between unilateral and bilateral performance when the 2 types of trials were equated for the number of locations stimulated (Experiment 1) and the number of redundant copies of the target (Experiment 2). Results suggest that when the number of stimulated locations is held constant, each of 2 copies of a target stimulus can be processed with the same efficiency and the same strategy as it would have been had it been the only copy. PMID- 10527057 TI - Hemispheric dissociations in controlled lexical-semantic processing. AB - Cognitive mechanisms of semantic priming in individuals with intact cerebral hemispheres were studied using the visual half-field method and lexical-decision tasks. In Experiment 1, unidirectionally associated word pairs were presented in a forward direction (e.g., BEAVER-TAIL) to isolate priming attributable to automatic activation or expectancy-based processing. Forward priming was restricted to the right visual field-left hemisphere, and it occurred only when expectancy-based processing was encouraged in the instructions. Experiments 2 and 3 found backward priming (e.g., TAIL-BEAVER) only in the left visual field, indicating that the right hemisphere contributes to retrospective semantic matching of the target back to the prime. The results suggest that the 2 hemispheres have different roles in controlled processing of semantic relations. PMID- 10527058 TI - Cross-language tests of hemispheric strategies in reading nonwords. AB - Four experiments explored the effects of specific language characteristics on hemispheric functioning in reading nonwords using a lateralized trigram identification task. Previous research using nonsense consonant-vowel-consonant (CVC) trigrams has shown that total error scores reveal a right visual field (RVF) advantage in Hebrew, Japanese, and English. Qualitative error patterns have shown that the right hemisphere uses a sequential strategy, whereas the left hemisphere uses a more parallel strategy in English but shows the opposite pattern in Hebrew. Experiment 1 tested whether this is due to the test language or to the native language of the participants. Results showed that native language had a stronger effect on hemispheric strategies than test language. Experiment 2 showed that latency to target letters in the CVCs revealed the same asymmetry as qualitative errors for Hebrew speakers but not for English speakers and that exposure duration of the stimuli affected misses differentially according to letter position. Experiment 3 used number trigrams to equate reading conventions in the 2 languages. Qualitative error scores still revealed opposing asymmetry patterns. Experiments 1-3 used vertical presentations. Experiment 4 used horizontal presentation, which eliminated sequential processing in both hemispheres in Hebrew speakers, whereas English speakers still showed sequential processing in both hemispheres. Comparison of the 2 presentations suggests that stimulus arrangement affected qualitative errors in the left visual field but not the RVF for English speakers and in both visual fields for Hebrew speakers. It is suggested that these differences result from orthographic and morphological differences between the languages: Reading Hebrew requires attention to be deployed to all the constituents of the stimulus in parallel, whereas reading English allows sequential processing of the letters in both hemispheres. Implications of cross-language studies for models of hemispheric function are discussed. PMID- 10527059 TI - Dissociation between two forms of conceptual priming in Alzheimer's disease. AB - Patients with Alzheimer's disease (AD) and healthy control participants performed 2 conceptual repetition priming tasks, word-associate production and category exemplar production. Both tasks had identical study-phases of reading target words aloud, had the most common responses as target items, and required production of a single response. Patients with AD showed normal priming on word associate production but impaired priming on category-exemplar production. This dissociation in AD suggests that conceptual priming is not a unitary form of memory but rather is mediated by separable memory systems. PMID- 10527060 TI - Longitudinal analysis of phonemic clustering and switching during word-list generation in Huntington's disease. AB - Two characteristics of word-list generation performance are forming clusters (i.e., contiguous words from the same subcategory) and switching among them. Patients with frontal lobe pathology show reduced switching on letter-cued word generation tasks, and clustering has been associated with temporal lobe functioning. Letter-cued word generation was examined in 72 patients with Huntington's disease (HD) and 41 healthy participants of equivalent age and education. As predicted, the patients showed reduced switching but normal clustering. In addition, switching but not clustering correlated inversely with disease severity, as measured by both movement and mental status scales. Furthermore, 5-year longitudinal analysis revealed a monotonic decrease in switching over time, whereas clustering performance remained stable. Control participants performed uniformly over time on both measures. These results are consistent with a progressive reduction in cognitive flexibility attributed to disruption of frontal-subcortical circuits secondary to neostriatal pathology in HD. PMID- 10527061 TI - Depression does not aggravate the episodic memory deficits associated with Alzheimer's disease. AB - In a population-based study of persons between 75 and 96 years of age, normal old adults (n = 296), patients with Alzheimer's disease (AD; n = 45), and patients with concomitant AD and depression (AD-D; n = 9) were compared on free recall and recognition of slowly and rapidly presented words and digit span. With the exception of forward digit span, the normal old group outperformed the 2 AD groups across all tasks. In free recall, only the normal old group performed better as task pacing decreased; however, all groups benefited from more study time in recognition. This suggests that both AD and AD-D patients have deficits in the ability to use more study time for remembering. Of most importance, the 2 AD groups were indistinguishable for all task variables. This lack of comorbidity effects is discussed relative to the view that depression, much like many other individual-difference variables that affect memory performance in normal aging, may be overshadowed by the influence of the neurodegenerative process in AD. PMID- 10527062 TI - Deterioration of frontal lobe function in normal aging: influences of fluid intelligence versus perceptual speed. AB - A group of young participants were compared with 2 groups of older participants (young-old, 65-74 years and old-old, 75 years or over) on a range of frontal lobe tasks. They were also assessed on the Digit Symbol Substitution Test (DSST), a test of digit cancellation (DC), the AH4 test of fluid intelligence, and the National Adult Reading Test (NART)-a measure of crystalized intelligence. Reliable age differences on all frontal measures except word fluency (FAS) were found. However, age effects were radically attenuated when either DSST speed or Alice Heim 4 (AH4) performance was used as a covariate. In contrast, DC and NART attenuated age-related variance to a much lesser degree. The authors conclude that a large proportion of age-related variance on measures of frontal lobe function may be attributed to a more general factor characterized jointly by DSST and AH4 performance. PMID- 10527063 TI - Depression in multiple sclerosis: relationship to working memory capacity. AB - Recent research has shown that depression in multiple sclerosis (MS) is associated with deficits on cognitively demanding tasks. One explanation for this relationship is that depressed MS patients may have reduced working memory capacity. The present study was designed to test this hypothesis. Depressed MS patients were compared with nondepressed MS patients and nondepressed healthy controls on a task of working memory capacity (reading span) and a short-term memory task not taxing working memory capacity (word span). In support of the capacity-reduction model, compared with the nondepressed groups, depressed MS patients performed significantly worse on reading span (p<.001) but not on word span. Additionally, reading span was significantly correlated with capacity demanding tasks shown to be impaired in depressed MS patients in previous reports. Results suggest that depressed MS patients are characterized by limited working memory capacity and that the central executive component of the working memory system may be most affected. PMID- 10527064 TI - Relative memory deficits in recurrent versus first-episode major depression on a word-list learning task. AB - Although memory deficits are associated with major depressive disorder, few studies have identified which patient characteristics predict impairment. Because recurrent depression appears related to more severe cerebral dysfunction, the present study tested whether recurrent depressed individuals have worse memory function than first-episode depressed individuals. Two groups of young-adult, nonpsychotic, depressed inpatients (20 single episode [SE] and 46 recurrent episode [RE]) were administered the California Verbal Learning Test within a broader battery of neuropsychological tests. The groups were equivalent in age, education, estimated IQ, severity of depression, and demographic composition. The RE group demonstrated memory deficits relative to both the SE group and published norms, but no other significant difference was found across the battery. Data indicate that abnormal memory performance is associated with recurrent depression, whereas memory deficits are not prominent in first-episode depressed individuals. PMID- 10527065 TI - Striatal activation during acquisition of a cognitive skill. AB - The striatum is thought to play an essential role in the acquisition of a wide range of motor, perceptual, and cognitive skills, but neuroimaging has not yet demonstrated striatal activation during nonmotor skill learning. Functional magnetic resonance imaging was performed while participants learned probabilistic classification, a cognitive task known to rely on procedural memory early in learning and declarative memory later in learning. Multiple brain regions were active during probabilistic classification compared with a perceptual-motor control task, including bilateral frontal cortices, occipital cortex, and the right caudate nucleus in the striatum. The left hippocampus was less active bilaterally during probabilistic classification than during the control task, and the time course of this hippocampal deactivation paralleled the expected involvement of medial temporal structures based on behavioral studies of amnesic patients. Findings provide initial evidence for the role of frontostriatal systems in normal cognitive skill learning. PMID- 10527066 TI - Visual localization in dyslexia. AB - Individuals with specific reading disability (SRD) may exhibit visual psychophysical abnormalities that include prolonged visual persistence, decreased luminance contrast sensitivity, lower flicker fusion thresholds, abnormal metacontrast masking, and lower motion detection sensitivity. These abnormalities could result from impairment of the magnocellular division of the visual afferent pathway to the cortex. The authors predicted that an impairment of this pathway would also cause abnormalities in ability to localize visual stimuli. This prediction was tested in 2 experiments. Results of both experiments showed that adults who reported a history of SRD and who currently had lower reading performance were less able than non-SRD participants to report the locations of small visual stimuli that were briefly flashed at positions similar to the ends of lines of text. PMID- 10527067 TI - Postconcussion syndrome following sports-related head injury: expectation as etiology. AB - Mild head trauma is often complicated by a persistent set of symptoms known as postconcussion syndrome (PCS). Past research has suggested that an expectancy guided, retrospective-recall bias may account for much of the variance in PCS symptom reporting. The present study examined the influence of symptom expectations on mild head trauma symptom reports among participants in contact sports. Head-injured athletes reported symptom rates that did not differ from those of uninjured athletes but consistently underestimated the preinjury incidence of symptoms. Athletes with no head trauma history overestimated the expected degree of pre- to postinjury change in symptom status. Results suggest that individuals with mild head injury tend to overestimate postconcussion symptom change in a manner consistent with their symptom expectations. A cognitive-behavioral model that explains the persistence of PCS is proposed. PMID- 10527068 TI - Sex differences in episodic memory: the impact of verbal and visuospatial ability. AB - The impact of verbal and visuospatial ability on sex differences in episodic memory was investigated. One hundred men and 100 women, 2040 years old, participated in a series of verbal and visuospatial tasks. Episodic memory was assessed in tasks that, to a greater or lesser extent, were verbal or visuospatial in nature. Results showed that women excelled in verbal production tasks and that men performed at a superior level on a mental rotation task. In addition, women tended to perform at a higher level than men on most episodic memory tasks. Taken together, the results demonstrated that (a) women perform at a higher level than men on most verbal episodic memory tasks and on some episodic memory tasks with a visuospatial component, and (b) women's higher performance on episodic memory tasks cannot fully be explained by their superior performance on verbal production tasks. PMID- 10527070 TI - The development of awareness of the carcinogenic hazard of inhaled iron. AB - Numerous studies have observed that workers in ferriferous industries have an elevated risk of respiratory tract neoplasia. Research at the cellular and animal model levels indicates that iron compounds, per se, are carcinogenic. However, some investigators have suggested that inhaled iron compounds are merely carriers of other carcinogens. Evidence is presented that iron apparently is a principal carcinogenic hazard in inhalation of silicon dioxide, asbestos, and tobacco smoke. Included in the discussion are unresolved questions concerning the precise role of inhaled iron as a carcinogen. PMID- 10527069 TI - Peripartum hypoxic risk and cognitive outcome: a study of term and preterm birth children at early school age. AB - The authors examined the relationships between gestational maturity, perinatal hypoxic risk, and intellectual outcome in early school-age children. The sample was composed of 48 children whose arterial pH obtained within 3 hr after delivery was between 7.3 (the lower end of the normal range) and 7.1 (the lower end of the moderately acidotic range). Gestational maturity did not account for a significant proportion of variance in outcome, whereas arterial pH was found to be significantly related to subsequent intellectual performance. The observed relationship between peripartum arterial pH and cognitive performance is especially noteworthy because the arterial pH range was restricted. The authors conclude that a "dose-response" relationship can be observed between arterial pH and intellectual outcome at early school age, even when the lower end of the acidotic range is truncated above the pH level that is thought to reflect severe asphyxia neonatorum. PMID- 10527071 TI - Short-term cultures of clinical tumor material: potential contributions to oncology research. AB - The culture of surgical tumor specimens has long been considered as a potential approach to the tailoring of chemotherapy and radiotherapy to the individual patient, and to the development of improved therapy. Recent work highlighting the importance of cell-cell interactions in the growth and survival of cancer tissue, as well the demonstrated importance of drug- or radiation-induced loss of tumor cells (for instance by apoptosis), points to a need to reexamine the question of what information might be derived from such cultures. In this commentary, we consider whether the short-term culture of human tumor tissue as small cellular aggregates, preserving to some extent the three-dimensional organization of tumors in vivo, can be used to obtain information on the behavior of cancer cells before and after therapy. Using [3H]thymidine incorporation as an end-point, we show how the shapes of dose-response curves might be used to estimate two key cytokinetic properties of the cultured cells, proliferation rate, and susceptibility to drug- or radiation-induced cell death. We have illustrated this discussion with our studies of melanoma, ovarian cancer, and lung cancer samples. We consider how application of culture methods may lead not only to the discovery of new antitumor drugs, but also to improved choice of patients' treatment. PMID- 10527072 TI - Induction of apoptosis in T98G glioblastoma cells by transfection of GML, a p53 target gene. AB - Expression of the glycosyl-phosphatidylinositol-anchored molecule-like protein (GML) gene, a p53 target, correlates with the sensitivity of some cancer cell lines to anticancer drugs and ionizing radiation. To investigate the function of GML further, we introduced the GML cDNA into various cancer cell lines under control of the tetracycline-regulated system. When we introduced GML into human glioblastoma cell line T98G, which lacks wild-type p53 and expresses no endogenous GML, we observed significant growth suppression accompanied by G2/M arrest in two independent, stable cell lines. We confirmed induction of apoptosis by fluorescence-activated cell sorting (FACS) analysis and nuclear staining. Our results indicated that GML could induce apoptosis of T98G without functional p53, and implied that GML plays a crucial role in the apoptotic pathway in some cancer cells. PMID- 10527073 TI - Antitumor effect of vaccinia virus in glioma model. AB - The ability of certain viruses to lyse cancer cells suggests that they may have potential as oncolytic agents. We investigated the effect of vaccinia virus (VV) and its recombinant derivatives (recVV2, rVV-p53) on growth of C6 rat glioma cells that form fast growing tumors in athymic nude mice. VV effectively infected C6 cells in vitro, inducing high level of foreign gene expression. Most of C6 cells infected in vitro with rVV-p53 expressing the tumor suppressor p53 protein showed apoptosis specific morphological changes in DAPI-stained nuclei and DNA fragmentation pattern on gel electrophoresis; infection with VV induced low level of cell apoptosis. In an ex vivo experiment, VV-infected C6 cells were implanted s.c. in athymic nude mice and tumor development was monitored. In contrast to the control PBS group, most of mice implanted with infected cells remained tumor free until the end of the observation period. In an in vivo experiment, injection of VV or rVV-p53 after the C6 cells had been implanted in nude mice induced effective inhibition of tumor growth in comparison with control PBS groups. The oncolytic effect was greater with rVV-p53, apparently due to overexpressed p53 and p53-mediated cell apoptosis. In study of virus virulence we did not observe disease symptoms in athymic mice infected with a high dose of VV. Experimental results indicate that vaccinia virus itself and vaccinia-mediated delivery of therapeutic genes represent novel potential strategies for tumor therapy. PMID- 10527074 TI - Biological evaluation on different human cancer cell lines of novel colchicine analogs. AB - Three new 7-0-substituted deacetamidothiocolchicine derivatives have been evaluated for their antitumor activity against various human tumor cell lines, some of which express the multidrug resistance (MDR) phenotype, for their impact on the cell cycle and their binding to tubulin. Colchicine and thiocolchicine were used as reference compounds. Thiocolchicine was the most active agent on MDR negative cells in terms of growth inhibition, whereas for multidrug-resistant cells, thiocolchicone was the most active compound (IC50 = 14 nM). As indicated by statistical analysis, a perfect agreement for the potency order (IC50 values) of the compounds between all the MDR-negative cancer cells (k = 1.00), a poor agreement between MDR-positive and MDR-negative cancer lines, and a moderate agreement (k = 0.50) between the two resistant cancer cells MCF-7 ADRr and CEM VBL were observed. To gain further insight into the mechanism of the antitumor activity of colchicinoids, the most active compounds, colchicone and thiocolchicone, were selected to evaluate their effect on cell cycle, apoptosis, and tubulin interaction. The highest recruitment activity into the G21/M phase of the cell cycle was detected in thiocolchicone-treated breast cancer cells. Interestingly, after 72 h of culture, when the cell cycle block subsided, a consistent amount of DNA fragmentation, a hallmark of apoptosis, was evident. Morphological analysis of MCF-7 ADRr cells confirmed this hypothesis and revealed that thiocolchicone was able to induce apoptosis in this MDR-bearing model. We also demonstrated, using flow cytometry, that thiocolchicone interacts with alpha and beta-tubulin, thereby affecting the expression of both subunits. PMID- 10527075 TI - In vivo studies of adenovirus-mediated p53 gene therapy for cis-platinum resistant human ovarian tumor xenografts. AB - We have recently reported that mutations of the tumor suppressor p53 gene are associated with the development of resistance to cis-platinum in human ovarian cancer cells, and that adenovirus-mediated reintroduction of the wild-type p53 (wtp53) gene in ovarian tumor cells resulted in the sensitization of tumor cells to cis-diamminedichloroplatinum (II) (CDDP). The purpose of this study was to evaluate whether i.p. treatment of CDDP-resistant tumor cells expressing mutant p53 (mutp53) with a recombinant adenovirus expressing wtp53 (Adwtp53) would result in the sensitization of resistant cells to CDDP. In order to determine whether i.p. injection of a recombinant adenovirus would result in expression of the transgene in tumor cells growing intraperitoneally, we first injected A2780/CP cells in nude mice and 10 days later the mice were injected i.p. with a recombinant adenovirus expressing beta-galactosidase (Ad beta-gal). Twenty-four hours following i.p. injection of Ad beta-gal, tumors were removed and stained for beta-gal. While tumors showed extensive staining for beta-gal, indicating internalization of adenovirus and the expression of the transgene in tumors, no expression of beta-gal protein was detected in liver. I.p. treatment of A2780/CP tumor xenografts with Adwtp53 caused extensive tumor cell death, which was further enhanced by CDDP. Treatment with Adwtp53 (5 x 10(7) pfu/day, 3-5 treatments) resulted in a significant decrease in tumor volume and increase in animal survival compared to either no treatment or treatment with vector alone without p53 gene. Additional therapy with CDDP (1 mg/kg/day x 3-4) further reduced tumor volume and increased survival (30-40%), suggesting that combination therapy of Adwtp53 and CDDP was better than single agents alone. Our results indicate that i.p. dosing with adenovirus-mediated wtp53 gene therapy could be beneficial in combination with CDDP for the treatment of ovarian tumors expressing mutp53. PMID- 10527077 TI - Osteoporosis in hemiplegic stroke patients as studied with dual-energy X-ray absorptiometry. AB - OBJECTIVES: To compare bone mineral densities (BMDs) of the affected and unaffected limbs in stroke patients at multiple sites; to study longitudinal changes during a 3-month rehabilitation program; and to relate BMDs to demographic, impairment, and disability variables. DESIGN: Descriptive study. SETTING: Tertiary rehabilitation center. PATIENTS: One hundred four consecutive hemiplegic inpatients, 69 men, age 56.5 +/- 13.2 yrs, 47 with left-sided brain lesion. Median days from onset to admission and median length of stay days were 83 and 105.5, respectively. MAIN OUTCOME MEASURES: BMDs of proximal humerus, distal radius, femoral neck, and calcaneus bilaterally, and third lumbar vertebra, measured with dual-energy x-ray absorptiometry (DXA), were compared between affected and unaffected sides at admission and discharge. RESULTS: Stroke Impairment Assessment Set (SIAS) motor scores, Functional Independence Measure (FIM) scores, grip strength, and awake/sleep heart rate counts (activity index) improved significantly at discharge. Affected/unaffected BMD ratios were 88.3% to 98.4% at admission and 79.6% to 98.8% at discharge, lowest for the humerus. Discharge/admission ratios were 89.1% to 97.8% for the affected and 97.4% to 100% for the unaffected side. All BMDs were intercorrelated (R = .438 to .873). They correlated significantly with age, body weight, grip strength, FIM scores, and activity index. Factors selected to explain BMD with multiple regression analysis differed according to the site and timing of the measurement. CONCLUSIONS: BMDs of the affected side were lower and most marked in the humerus. Longitudinally, not only the affected but the unaffected BMDs decreased. Age, sex, duration of stroke, anthropometric measurements, motor paralysis, muscle strength, and activity level contributed differently to bone loss according to the site and timing of the measurement. PMID- 10527076 TI - Muscle strengthening and physical conditioning to reduce impairment and disability in chronic stroke survivors. AB - OBJECTIVE: To evaluate the impact of a program of muscle strengthening and physical conditioning on impairment and disability in chronic stroke subjects. DESIGN: A randomized pretest and posttest control group, followed by a single group pretest and posttest design. SUBJECTS: Thirteen community-dwelling stroke survivors of at least 9 months. INTERVENTION: A 10-week (3 days/week) program consisting of a warm-up, aerobic exercises, lower extremity muscle strengthening, and a cool-down. MAIN OUTCOME MEASURES: Peak isokinetic torque of the major muscle groups of the affected lower limb, quadriceps and ankle plantarflexor spasticity, gait speed, rate of stair climbing, the Human Activity Profile (HAP), and the Nottingham Health Profile (NHP) were recorded twice for the treatment group and three times for the control group. RESULTS: Significant improvements were found for all the selected outcome measures (HAP, NHP, and gait speed) for the treatment group (p < .001). In terms of overall training effects, the 13 subjects demonstrated increases in strength of the affected major muscle groups, in HAP and NHP profiles, and in gait speed and rate of stair climbing without concomitant increases in either quadriceps or ankle plantarflexor spasticity. CONCLUSIONS: The 10-week combined program of muscle strengthening and physical conditioning resulted in gains in all measures of impairment and disability. These gains were not associated with measurable changes of spasticity in either quadriceps or ankle plantarflexors. PMID- 10527078 TI - Balance and physical impairments after stroke. AB - OBJECTIVE: To investigate the relationship among laboratory and clinical balance measures and physical impairments. DESIGN: A descriptive correlational study. SETTING: Research laboratory. PARTICIPANTS: Thirty subjects with stroke, recruited through convenience sampling. MAIN OUTCOME MEASURES: Postural sway was calculated in terms of center of pressure (COP) parameters including spectral characteristics. Clinical balance was measured using the Balance Scale. The assessed physical impairments included stages of lower limb motor recovery, ankle proprioception, and passive dorsiflexion range of the involved limb. RESULTS: The Balance Scale was correlated with COP speed (r = -.57), COP root mean square speed (r = -.50), and COP mean frequency (r = -.50) in the anterior-posterior direction only. Moderate to high correlations were found among most of the COP parameters except spectral characteristics. Significant differences in postural sway were found among different stance in eyes-open (p = .00 to .02) and eyes closed conditions (p = .00 to .04). Subjects with impaired ankle proprioception had significantly increased postural sway and decreased Balance Scale scores when compared with the subjects with intact ankle proprioception. CONCLUSIONS: Some of the clinical and laboratory balance assessments were related, indicating that some components of the tests are similar, but some measured different aspects of balance. Postural sway was related to visual condition, stance position, and proprioception. PMID- 10527079 TI - Neurolysis of the musculocutaneous nerve with alcohol to treat poststroke elbow flexor spasticity. AB - OBJECTIVE: To evaluate the effectiveness of alcohol in neurolysis of the musculocutaneous nerve for the treatment of elbow flexor spasticity in individuals with a stroke. DESIGN: Case series. SETTING: Outpatient clinic of a tertiary rehabilitation facility. PARTICIPANTS: Twenty patients with a mean age of 62.8 years and poststroke duration of 12.3 months with elbow flexor spasticity. INTERVENTION: Musculocutaneous nerve block of the hemiplegic upper extremity with 50% ethyl alcohol. OUTCOME MEASURES: The severity of spasticity as assessed by the modified Ashworth scale (MAS) score and the elbow passive range of motion (PROM). RESULTS: The mean baseline MAS score was 3.7 +/- 0.6, and this improved to 1.7 +/- 1.0, 2.0 +/- 0.8, and 2.1 +/- 0.8 at 4 weeks, 3 months, and 6 months postneurolysis, respectively. The elbow PROM was 87.3 degrees +/- 20.2 degrees, 104.3 degrees +/- 20.1 degrees, 103.8 degrees +/- 18.9 degrees, and 101.6 degrees +/- 19.7 degrees, respectively. These improvements were statistically significant (p < .05). Four subjects had concomitant improvement of finger flexor spasticity and another four had relief of shoulder pain. Three subjects developed temporary dysesthetic pain over the lateral forearm. CONCLUSION: Neurolysis of the musculocutaneous nerve with alcohol provides good relief of elbow flexion spasticity in hemiplegic individuals. PMID- 10527080 TI - Powered feeding devices: an evaluation of three models. AB - OBJECTIVE: To evaluate and compare three powered feeding devices (Beeson, Handy 1, Winsford) as perceived by disabled individuals who require assistance with eating. DESIGN: Subjects and assistants were surveyed after using each device and serving their own controls. The order in which the devices were used was balanced. SETTING: Place of subjects' residence. SUBJECTS: Twelve subjects, ages 11 to 42 years, and their feeding assistants. INTERVENTION: Each device trial covered a 4-day period. Day 1 focused on training to use the device, Days 2 and 3 focused on using the device at home, and on Day 4 subjects returned to the laboratory for debriefing, completing questionnaires, and videotaping. MAIN OUTCOME MEASURE: Subjects and assistants answered questionnaires including Likert like rankings and yes/no responses regarding functional and esthetic characteristics of each feeding device. RESULTS: Significant differences were found among three powered feeding devices regarding specific design characteristic. Great percentages of both subjects and their feeding assistants responded that the devices were an improvement over how they were currently being fed and that they would use such a device on a daily basis. CONCLUSION: Individuals dependent on others for feeding may benefit from the use of a powered feeding device. PMID- 10527081 TI - Pain in persons with cerebral palsy. AB - OBJECTIVE: To examine the nature and scope of pain in persons with cerebral palsy (CP). DESIGN: Standardized interviews to assess demographics, pain experiences, and the impact of pain on activities. SUBJECTS: Ninety-three adults with CP recruited from medical clinics at the University of Washington and local residential and community housing for persons with developmental disabilities. MAIN OUTCOME MEASURES: Weekly and 3-month pain intensities, chronic pain grade, interference in daily activities caused by pain, and pain-exacerbating and pain relieving factors. RESULTS: Sixty-two subjects (67%) reported one or more areas of pain of > or =3 months' duration. Lower extremity pain and back pain were the most common complaints. Fifty-six percent of the subjects reporting pain indicated it occurred daily. Mean average pain intensity, graded on a scale of 0 (no pain) to 10 (pain as bad as could be), was 3.16 (SD = 2.45) in the preceding week and 4.45 (SD = 2.34) in the previous 3 months. Approximately 53% of subjects reporting pain indicated their average pain was of moderate to severe intensity (average pain rated as > or =5). Using Von Korff's Chronic Pain Grade classification system, the majority of subjects who reported pain fell into either grade I (low disability, low pain intensity; 51%) or grade II (low disability, high pain intensity; 39%). Subjects reported many factors that exacerbate pain (eg, stress or weather) or decrease it (eg, exercise or rest). CONCLUSIONS: The data suggest that pain is common in adults with CP. In many subjects, pain levels were moderate to intense. PMID- 10527082 TI - Development of fatigue during repeated eccentric-concentric muscle contractions of plantar flexors in patients with stroke. AB - OBJECTIVE: To better understand the mechanisms behind fatigue in muscles affected by a neuromuscular disease. DESIGN: Patients recruited by convenience compared to age-matched healthy subjects from a population study. SETTING: University hospital laboratory. METHODS: Repetitive eccentric-concentric plantar flexions at 60 degrees/sec were performed on a dynamometer until exhaustion. The mean power frequency and root mean square of the electromyogram were recorded, and work was calculated. SUBJECTS: Both legs of seven patients with upper motor neuron lesion from stroke and one leg of healthy men were tested. RESULTS: There were no significant (p > .05) differences in number of cycles performed or decrease of work between any of the tested legs. There was a significant (p > .05) difference in work performed by the affected leg and the reference group. Mean power frequency decreased significantly (p > .05) for the gastrocnemius muscle in the nonaffected leg and for the reference group, while no such decreases were found in the affected leg. The statistical methods used were the nonparametric tests: the Wilcoxon one-sample for differences between paired observations, and the Mann Whitney U for differences between groups. CONCLUSION: A reduction in work in high intensity dynamic muscle activity may not be associated with a reduction in mean power frequency, especially in patients with altered supraspinal control. There may be peripheral fatigue factors not reflected in the electromyographic activity. PMID- 10527083 TI - Neoplastic versus traumatic spinal cord injury: an outcome comparison after inpatient rehabilitation. AB - OBJECTIVE: To compare outcomes of patients with neoplastic spinal cord compression (SCC) to outcomes of patients with traumatic spinal cord injury (SCI) after inpatient rehabilitation. DESIGN: A comparison between patients with a diagnosis of neoplastic SCC admitted to an SCI rehabilitation unit and patients with a diagnosis of traumatic SCI admitted to the regional Model Spinal Cord Injury Centers over a 5-year period, controlling for age, neurologic level of injury, and American Spinal Injury Association impairment classification. SETTING: Tertiary university medical centers. PATIENTS: Twenty-nine patients with neoplastic SCC and 29 patients with SCI of traumatic etiology who met standard rehabilitation admission criteria. MAIN OUTCOME MEASURES: Acute and rehabilitation hospital length of stay (LOS), Functional Independence Measure (FIM) scores, FIM change, FIM efficiency, and discharge rates to home. RESULTS: Patients with neoplastic SCC had a significantly (p < .01) shorter rehabilitation LOS than those with traumatic SCI (25.17 vs 57.46 days). No statistical significance was found in acute care LOS. Motor FIM scores on admission were higher in the neoplastic group, but discharge FIM scores and FIM change were significantly lower. Both groups had similar FIM efficiencies and community discharges. CONCLUSIONS: Patients with neoplastic SCC can achieve rates of functional gain comparable to those of their counterparts with traumatic SCI. While patients with traumatic SCI achieve greater functional improvement, patients with neoplastic SCC have a shorter rehabilitation LOS and can achieve comparable success with discharge to the community. PMID- 10527084 TI - Quantitative sensory testing in patients with incomplete spinal cord injury. AB - OBJECTIVE: To examine the utility of quantitative sensory testing (QST) to characterize sensory dysfunction in patients with spinal cord injury (SCI). DESIGN: Perceptual thresholds to warm, cold, cold pain, and vibratory stimuli were investigated using a modified method of "limits." METHOD: Three QST trials were administered to six lower leg dermatomes, on two different days, to estimate the reliability of measurement. SETTING: Regional Spinal Cord Injury Rehabilitation Center in Ontario, Canada. SUBJECTS: Twenty-one SCI patients with incomplete neurologic deficits and 14 able-bodied controls of similar age. RESULTS: ANOVA revealed significantly (p < .05) reduced perceptual threshold values (hypoesthesia) for warm, cold, and vibratory sensation in the SCI group. There were no differences between group mean values for cold pain because of the inclusion of patients with hypoalgesia and hyperalgesia. Intraclass correlation coefficient estimates of reliability revealed large between-subject variability in the SCI patients associated with relatively small trial-to-trial variability within each day of testing, and appreciable between-day variances. CONCLUSIONS: With QST in SCI there is a need for repeated measurements across days to establish stable baseline measures or outcomes following intervention. QST is a useful adjunct to clinical examination for assessment of preserved sensation. PMID- 10527085 TI - Salbutamol effect in spinal cord injured individuals undergoing functional electrical stimulation training. AB - OBJECTIVE: Preliminary study to investigate possible changes in skeletal muscle morphology and function, as well as hormonal and metabolic effects, after treatment with a selective beta2-adrenergic receptor agonist. DESIGN: Double blind, placebo-controlled trial. PARTICIPANTS: Three individuals with spinal cord injury (SCI). INTERVENTION: Two-week treatment with salbutamol (2mg) or placebo (ascorbic acid, 50mg) twice a day. Program of functional electronic stimulation (FES) cycling for 30 minutes twice a week. MAIN OUTCOME MEASURES: Body weight, three measures of leg circumference (gluteal furrow, one third of subischial height up from tibial-femoral joint space, and minimum circumference above the knee), muscle fiber area, and total work output per session. RESULTS: There were increases in body weight (2.30 +/- .70kg), leg circumferences (gluteal furrow 1.70 +/- .27cm, one third subischial height 1.53 +/- 1.65cm, minimum circumference above the knee .43 +/- .04cm), and muscle (vastus lateralis) cross sectional area (1,374 +/- 493 to 2,446 +/- 1,177microm2) after salbutamol treatment, whereas quadriceps muscle contractile function was not modified. Total work output during FES cycling sessions was increased more during salbutamol treatment (64%) compared with training alone (27%). Salbutamol treatment was associated with a large decrease in skeletal muscle beta-adrenergic receptor density. CONCLUSION: Although some side effects were noted, these results suggest that a short treatment with the beta2-adrenergic receptor agonist salbutamol during a training program with FES cycling could be beneficial in patients with SCI. PMID- 10527086 TI - Posttraumatic erectile potential of spinal cord injured men: how physiologic recordings supplement subjective reports. AB - OBJECTIVE: To investigate by means of a neurophysiologic model the remaining erectile function in spinal cord injured men. DESIGN: A nonrandomized control trial. SETTING: A Referred Care Center. SUBJECTS: Forty-seven spinal cord injured men and 7 noninjured controls. INTERVENTION: The subject penile responses were recorded by a penile strain gauge during two sessions--one to obtain baseline responses, and one with reflexogenic stimulation (masturbation) and psychogenic stimulation (film). MEASURES: Average tumescence, maximal tumescence, percentage rigidity, and duration of tumescence and rigidity. RESULTS: Significant results were found for subjects with lower lesions using psychogenic stimulation as their optimal mode compared with reflexogenic stimulation as an alternate mode, and for subjects with higher lesions using reflexogenic stimulation as their optimal mode, compared with psychogenic stimulation as an alternate mode. The responses with optimal stimulation modes were comparable to those achieved by controls. CONCLUSION: The findings validate the neurophysiologic model of posttraumatic erectile potential as a function of the lesion type and stimulation source. The results were comparable to those of noninjured subjects; the potential for normal function is present and may be amenable to sexual rehabilitation or use in conjunction with new oral drug treatments for impotence. PMID- 10527087 TI - The value of electrodiagnostic consultation for patients with upper extremity nerve complaints: a prospective comparison with the history and physical examination. AB - OBJECTIVES: To determine whether electrodiagnostic testing changes diagnostic certainty compared with a detailed history and physical examination, and whether interactions between medical information, the extent of testing, and diagnostic certainty imply a need for advanced medical knowledge on the part of the tester. DESIGN: Prospective observation. SETTING: University orthopedic department and small community hospital electrodiagnostic laboratories. PATIENTS: Two hundred fifty-five consecutive referrals for upper extremity nerve complaints. OUTCOME MEASURES: Diagnosis, diagnostic confidence, and severity of neurologic lesion were coded after standardized history and physical and after electrodiagnostic testing. RESULTS: Electrodiagnostic testing substantially altered 42% of diagnoses, confirmed 37%, and did not clarify 21%. The extent of testing correlated with the size of the differential diagnosis, the number of previous hospitalizations, and the number of other medical problems. Confidence in final diagnoses correlated positively with severity of the lesion, but negatively with the size of the differential diagnosis and the number of painful body areas. Hospitalizations and medical problems also tended towards negative correlations. CONCLUSIONS: This study, in which all electrodiagnostics, histories, and physical examinations were performed by a single physician, suggests that electrodiagnosis substantially alters clinical impressions in a large percentage of patients. The complex relationship between clinical information, the extent of testing, and final diagnostic certainty suggests that specialized medical knowledge is required for accurate electrodiagnosis. PMID- 10527088 TI - Persistent functional and social benefit 5 years after a multidisciplinary arthritis training program. AB - OBJECTIVE: To assess the sustainable benefits of a professional, multidisciplinary training program for patients with rheumatoid arthritis. DESIGN: Two studies with different observation periods. Study I was a prospective, randomized trial for 1 year. Study II was a noncontrolled observation over 5 years. SETTING: The 9-day program for eight patient groups encompassed a multidisciplinary cooperation between rheumatologists, orthopedists, physicotherapists, psychologists and social workers. PATIENTS: Sixty-eight consecutive patients with rheumatoid arthritis participated in an arthritis training program either immediately after enrollment in the program or after 1 year. INTERVENTIONS: The program covered the following fields: pathogenesis of rheumatoid arthritis, drug therapy, physicotherapy, practical exercise in remedial gymnastics, use of joint protection devices, orthopedic perspectives, psychological counseling, dietetics, information about unproven cures and social assistance. MAIN OUTCOME MEASURES: Clinical outcome was assessed by self-report questionnaires: (1) Stanford Health Assessment Questionnaire, (2) Freiburg Questionnaire of Coping with Illness, (3) Beck Depression Inventory, and (4) a 21-point scale to evaluate cognitive-behavioral and environmental impact. RESULTS: A significant and persistent improvement of all investigated parameters was demonstrated in the 1-year controlled trial. Between the end-point of the 1 year study and the 5-year evaluation, this improvement increased even more for functional status and coping with illness, whereas depression returned to baseline values. These effects were seen even without reinforcement of the training. CONCLUSION: A professional, multidisciplinary approach to educate patients with rheumatoid arthritis leads to a significant and sustained improvement of the clinical outcome and is an approach that should be established as a part of conventional therapy. PMID- 10527089 TI - Oxygen uptake during peak graded exercise and single-stage fatigue tests of wheelchair propulsion in manual wheelchair users and the able-bodied. AB - OBJECTIVE: To determine if a single-stage, submaximal fatigue test on a wheelchair ergometer would result in higher than expected energy expenditure. DESIGN: An experimental survey design contrasting physiologic responses during peak graded exercise tests and fatigue tests. SETTING: A rehabilitation science laboratory that included a prototypical wheelchair ergometer, open-circuit spirometry system, and heart rate monitor. PARTICIPANTS: Nine able-bodied non wheelchair users (the NWC group: 6 men and 3 women, mean +/- SD age 30 +/- 7yrs) and 15 manual wheelchair users (the WC group: 12 men and 3 women, age 40 +/- 9yrs, time in wheelchair 16 +/- 9yrs). No subject had any disease, medication regimen, or upper body neurologic, orthopedic, or other condition that would limit wheelchair exercise. MAIN OUTCOME MEASURES: Peak oxygen uptake (VO2) for graded exercise testing and during fatigue testing, using a power output corresponding to 75% peak aerobic capacity on graded exercise test. RESULTS: In the WC group, VO2 at 6 minutes of fatigue testing was not significantly different from peak VO2. In the NWC group, VO2 was similar to the expected level throughout fatigue testing. CONCLUSION: Energy expenditure was higher than expected in the WC group but not in the NWC group. Fatigue testing may provide a useful evaluation of cardiorespiratory status in manual wheelchair users. PMID- 10527090 TI - Osteoarthritis of the knee: isokinetic quadriceps exercise versus an educational intervention. AB - OBJECTIVE: To evaluate the effects of isokinetic exercise versus a program of patient education on pain and function in older persons with knee osteoarthritis. DESIGN: A randomized, comparative clinical trial, with interventions lasting 8 weeks and evaluations of 12 weeks. SETTING: An outpatient Veterans Affairs Medical Center clinic and an affiliated university hospital. PATIENTS: One hundred thirteen men and women between 50 and 80 years old with diagnosed osteoarthritis of the knee; 98 completed the entire assigned treatment. INTERVENTION: Patients received either a regimen of isokinetic exercise of the quadriceps muscle three times weekly over 8 weeks or a series of 4 discussions and lectures led by health care professionals. MAIN OUTCOME MEASURES: Variables studied for change were isokinetic and isometric quadriceps strength, pain and function determined by categorical and visual analog scales, and overall status using physician and patient global evaluations by the Arthritis Impact Scale, version 2, Western Ontario McMaster's Arthritis Index, and Medical Outcome Study Short Form 36. RESULTS: Both treatment groups showed significant strength gains (p < .05), which occurred over a wider velocity spectrum for the exercise group. Exercised patients also had improved pain scores for more of the variables measured than those receiving education. Both groups had positive functional outcomes and slightly improved measures of overall status. CONCLUSIONS: Isokinetic exercise is an effective and well-tolerated treatment for knee osteoarthritis, but a much less costly education program also showed some benefits. PMID- 10527091 TI - Pain Patient Profile: a scale to measure psychological distress. AB - OBJECTIVE: To evaluate the construct validity of the Pain Patient Profile (P-3), a brief self-report instrument designed to measure anxiety, depression, and somatization in patients presenting with pain. DESIGN: Comparison of P-3 scores with previously established measures of depression, anxiety, and somatization, and comparison of P-3 scores of pain patients with those of patients with diabetes. SETTINGS: Hospital-based outpatient pain clinic, family practice clinic, diabetes education group. PATIENTS: Seventy pain patients and 40 patients with diabetes. RESULTS: High positive correlations (.69 to .90) were found between the P-3 scales of Depression, Anxiety, and Somatization and the corresponding measures of these constructs, and high intercorrelations were found among the three P-3 scales. Significant differences were found between pain patients and diabetes patients for the P-3 Depression and Somatization scale scores, but not for the P-3 Anxiety scale scores. CONCLUSIONS: The P-3 is a useful instrument for initial screening of psychological distress in pain patients. Some patients may show elevations on more than one of the clinical scales, which either indicates that the P-3 does not distinguish well among these constructs or reflects the well-established comorbidity of these constructs. PMID- 10527092 TI - The Community Integration Questionnaire revisited: an assessment of factor structure and validity. AB - OBJECTIVE: To investigate the factor structure and concurrent validity of the Community Integration Questionnaire (CIQ), using a large sample of persons with traumatic brain injury (TBI). DESIGN: Principal components analysis with varimax rotation was performed on CIQ items completed through interview with patients at 1 year after injury. Correlational analyses compared CIQ scores to scores on other widely used outcome measures. SETTING: Outpatient clinics affiliated with four TBI Model System rehabilitation centers funded by the National Institute on Disability and Rehabilitation Research. PARTICIPANTS: Three hundred twelve patients with medically documented TBI who were enrolled in the TBI Model Systems Project. The majority of patients were Caucasian males with severe TBI. MAIN OUTCOME MEASURES: CIQ; Functional Independence Measure (FIM); Functional Assessment Measure (FAM); Disability Rating Scale (DRS). RESULTS: Three factors emerged: Home Competency, Social Integration, and Productive Activity. The financial management item was moved from Social Integration to Home Competency, and the travel item was moved from Productive Activity to Social Integration. Each CIQ scale score showed significant correlations in the expected direction with the FIM+FAM and DRS items. CONCLUSIONS: The results provide further evidence for the validity of the CIQ and improve the scoring system. The factor structure is clinically and theoretically meaningful. The subscale and total scores show significant relationships with other widely used measures of outcome. Future research should focus on increasing the range of questions, accounting for changes from preinjury functioning, and obtaining normative data on the new factors. PMID- 10527093 TI - Global aphasia: an innovative assessment approach. AB - OBJECTIVE: To provide an alternative language comprehension assessment strategy for patients unable to be tested with traditional verbally/behaviorally based methods. DESIGN: Event-related brain potentials were recorded from three midline scalp locations to visually and aurally computer-presented sentences, 50% of which were semantically appropriate and 50% semantically incongruous. SETTING: A rehabilitation hospital. PATIENT: A 21-year-old man with a traumatic brain injury. RESULTS: The patient exhibited brain response patterns to aurally presented congruous and incongruous sentences indicative of intact semantic processing capabilities. These findings resulted in reinstatement of individualized rehabilitative intervention, with a successful outcome. CONCLUSIONS: This innovative technique provides new opportunities for assessing intellectual function in noncommunicative patients who were patients previously unable to be tested. PMID- 10527094 TI - Strength assessment in postpolio syndrome: validity of a hand-held dynamometer in detecting change. AB - OBJECTIVES: To investigate the validity, the intraexaminer and interexaminer reproducibility, and the ability to detect change of a hand-held dynamometer (HHD) in strength measurements in former polio subjects. DESIGN: HHD measurement of knee extensor strength was compared with the criterion standard of a chair dynamometer measurement in 49 subjects. The "break" method was used for HHD measurements. Reproducibility was studied for six lower extremity muscle groups in 28 subjects. The measurements were performed by one experienced and one inexperienced examiner on two separate occasions, with an interval of 1 week. The examiners were blinded to each other's and to previous results. SETTING: University hospital. SUBJECTS: Volunteer sample of former polio subjects. RESULTS: For knee extension, the forces that could be measured with the HHD were limited to approximately 200N. Although the intraclass correlation coefficients were high (.75 to .98), the 95% limits of agreement between measurements showed large intervals for differences between two measurements (ratio intervals ranging from .76-1.52 to .52-2.77). The intraexaminer reproducibility for the experienced examiner was superior to that of the inexperienced examiner. The reproducibility of the inexperienced examiner showed systematic bias, with significantly higher strength values for the second session measurement of three muscle groups. CONCLUSION: The device has good validity in the lower force range. However, because the agreement between measurements was poor, it has limited ability to detect a change in muscle strength. Therefore, this method is unable to detect small changes in lower extremity muscle strength in former polio patients. PMID- 10527095 TI - Normative values of agonist-antagonist shoulder strength ratios of adults aged 20 to 78 years. AB - OBJECTIVE: To determine normative values for isometric flexion/extension, abduction/adduction, and external/internal rotation strength ratios about the shoulder and to determine if these ratios are affected by age or gender. STUDY DESIGN: A cross-sectional study of 120 healthy volunteers (60 men, 60 women) aged 20 to 78 years. SETTING: Orthopedic research laboratory. METHODS: Flexion and extension strengths were measured isometrically using a Cybex II dynamometer at arm flexion angles of 30 degrees, 60 degrees, and 90 degrees. Abduction and adduction strengths were measured at 30 degrees, 60 degrees, and 90 degrees abduction. Internal and external rotation strengths were measured (1) with the arm abducted 15 degrees and neutral external/internal rotation and (2) with the arm abducted 90 degrees and externally rotated 30 degrees above the transverse plane. OUTCOME MEASURES: Isometric strength ratios for flexion/extension, abduction/adduction, and external/internal rotation. RESULTS: No statistically significant differences in agonist/antagonist strength ratios were found between dominant and nondominant sides or between genders. Age was associated with changes in strength ratios for measurements taken with the arm flexed or abducted 90 degrees. Posture was found to affect strength ratios. CONCLUSIONS: These data can serve as a normative reference for clinical use. PMID- 10527096 TI - Healing of open stump wounds after vascular below-knee amputation: plaster cast socket with silicone sleeve versus elastic compression. AB - OBJECTIVE: To assess the effect of a plaster cast socket on the healing of open wounds and on temporary prosthesis fitting after below-knee amputation because of arterial occlusive disease. DESIGN: Randomized controlled trial. SETTING: Rehabilitation center, university hospital. PATIENTS: All included patients had undergone recent (in the previous 3 months) below-knee amputation because of arterial disease and initially had an open stump. Patients were randomly assigned to two groups of 28 subjects each. The sizes of the amputation scars were 8 to 24 cm2. Ischemia of the stump was eliminated as a probable cause of delayed wound healing by the inclusion criterion of transcutaneous oxygen tension (TcPO2) of >35 mmHg. The average age in group I (the experimental group) was 65.2 +/- 12.4 (SD) years and in group II (the control group) 66.8 +/- 10.8 years (not significant). INTERVENTION: A plaster cast (supracondylar-type) socket was fitted on the stumps of group I patients, interposed with a silicone sleeve. The patients were gradually trained to wear this cast for up to 5 hours a day. They were provided with elastic compression bandages for the remainder of the time. Patients in group II wore elastic compression bandages, which were only removed for dressing changes. MAIN OUTCOME MEASURES: Time required for stump healing, length of time between amputation and ability to walk wearing a contact socket, and length of hospital stay. RESULTS: Group I had a quicker average healing time (71.2 +/- 31.7 [SD] days compared to the control group's 96.8 +/- 54.9 days) and a shorter average length of hospital stay (99.8 +/- 22.4 days compared to the control group's 129.9 +/- 48.3 days). CONCLUSION: Use of a plaster cast socket leads to more rapid healing of the open stump and to a shorter hospitalization. If there is no stump ischemia, this plaster cast technique is safe. PMID- 10527098 TI - Functional recovery after bilateral pallidotomy for the treatment of early-onset primary generalized dystonia. AB - This report describes the successful treatment of dystonia musculorum deformans with bilateral stereotactic pallidotomy in a 14-year-old girl in whom the dystonia was diagnosed when she was 7 years old. The patient presented with dystonia of the right upper extremity that progressed to generalized dystonia. Preoperatively, she required maximal assistance with all activities of daily living and transfers. She was not a functional ambulator. Postoperatively, she had remarkable functional recovery. At discharge, she was at modified independence level for all basic activities of daily living and required supervision for household ambulation. No postoperative complications were noted. We propose that bilateral stereotactic lysis of globus pallidus interna may be an alternative treatment for dystonia musculorum deformans. The technique of bilateral pallidotomy and theories of its effectiveness are discussed. PMID- 10527097 TI - Male infertility and erectile dysfunction in spinal cord injury: a review. AB - OBJECTIVE: To review the pathophysiology, evaluation, and management of erectile dysfunction and infertility in spinal cord injury (SCI). STUDY SELECTIONS: Studies that covered various treatment options and their contraindications, complications, or side effects, including sildenafil (Viagra), intracavernosal injection therapy, topical medications and a urethral delivery system, a vacuum erection device, and penile prostheses. Other studies covered the effects of SCI on reproduction: spermatogenesis and testicular function, and seminal constituents. In addition, assisted reproductive techniques were compared: external vibratory stimulation, electroejaculation, testicular sperm aspiration, and intracytoplasmic sperm injection. CONCLUSION: Goal-directed therapy is the mainstay of treatment of erectile dysfunction in men with or without SCI. The choice of therapy is often defined more by the needs of the patient's sexual relationship than by his erectile dysfunction. The majority of men with SCI are infertile because of a combination of ejaculatory dysfunction, impaired spermatogenesis, and poor semen quality. Although many technological advances have evolved to overcome ejaculatory dysfunction, the sperm density, motility, and function remain poor. Until these parameters are improved, men with SCI will have to pursue more financially and emotionally taxing procedures. Further studies to elucidate the cellular and molecular mechanisms of diminished sperm quality are needed. Addressing the issues of erectile dysfunction and male infertility may help to preserve the relationship between the patient and his partner. PMID- 10527099 TI - False positive perfusion lung scintiscans in tetraplegic patients: a case series. AB - An accurate diagnosis of pulmonary embolism is essential to prevent excessive morbidity and mortality from either inappropriate therapy or failure to institute anticoagulation. The diagnosis of pulmonary embolism in tetraplegic spinal cord injury patients is complicated by frequent inability to perform the ventilation portion of the ventilation-perfusion scintiscan (V/Q scan) and by controversy regarding classification of defects on perfusion-only scans, as well as by coexisting pulmonary disease, systemic illness, related injuries, and the tendency for tetraplegic patients to have unexplained fever. This report describes three tetraplegic ventilator-dependent patients with hypoxic respiratory failure and normal chest radiographs who had large defects on perfusion-only lung scans. Ventilation scintiscans were not performed because the patients were ventilator-dependent with tracheostomies. Pulmonary angiography findings were normal in all patients, and all three responded to aggressive pulmonary toilet. Even large defects on perfusion-only scans despite normal chest radiographs should not be used to establish a diagnosis of pulmonary embolism in tetraplegic patients, and further diagnostic imaging is warranted. PMID- 10527100 TI - Ulnar conduction block at the wrist. AB - Two cases of ulnar nerve lesions at the wrist are reported. The lesions had an acute onset and exclusively impaired the ulnar motor deep branch. The coexistence of carpal tunnel syndrome in each case allowed an early diagnosis but was somewhat misleading. In both cases, the use of classic motor and sensory conduction studies did not provide clear abnormalities that would have precisely determined the site of the nerve lesion. In both cases, only palmar stimulation of the ulnar motor deep branch showed an important conduction block. This electrodiagnostic finding showed definitively the site of the ulnar nerve lesion at the wrist and excluded proximal ulnar nerve lesions or C8-T1 radiculopathy. In both cases recovery occurred without surgery. PMID- 10527101 TI - Mobility challenges and solutions for fibrodysplasia ossificans progressiva. AB - Fibrodysplasia ossificans progressiva (FOP) is a rare genetic disorder characterized by progressive soft tissue ossification. Although signs may be present at birth, the first appearance of ectopic bone typically occurs in early childhood. The primary target is the axial musculature. Eventually ectopic bone also occurs in ligaments, fascia, aponeurosis, tendons, and joint capsules of the appendicular skeleton with a proximal to distal predilection. As the disease advances, mobility becomes restricted, and affected individuals are typically limited to bed or chair by their early 30s. This report describes a 30-year-old woman with advanced FOP. She had a fused spine and a fixed pelvis, with hips and knees locked in flexion and feet in plantarflexion. Her upper limb mobility was similarly restricted. She was not able to stand upright or sit independently. The modification of a commercially available power wheelchair that allowed the patient to maintain her employment as a preschool teacher and custom shoes are described. Creative physiatric intervention is essential to liberate human potential for people with FOP. PMID- 10527102 TI - Spotter strap for the prevention of wheelchair tipping. AB - Injuries caused by wheelchair rear-tipping accidents are common. This article reports on the safety and effectiveness of a spotter strap that attaches to the cross-brace or frame below the center of gravity of an occupied wheelchair. We videotaped five therapists spotting 89 wheelchair users while the users each performed six tasks that were designed to induce rear instability. We induced 16 episodes of complete rear tipping. In all cases, the spotter strap allowed the spotter to stay out of the way during the task, but step in easily when necessary to prevent the wheelchair user from being injured. In one instance, the spotter needed assistance lifting a heavy subject to the upright position after catching the subject with the strap. In summary, the spotter strap is a safe and effective device. We recommend its use when there is a high risk of a rear-tipping accident. PMID- 10527103 TI - Independent living centers, medical rehabilitation centers, and managed care for people with disabilities. PMID- 10527104 TI - Contraction-induced H-reflex amplitude ratio. PMID- 10527105 TI - Neurophysiologic mechanisms of attention deficits in schizophrenia. AB - BACKGROUND: Despite advances in the pharmacologic treatment of schizophrenia, the neurophysiologic mechanism(s) of disordered attention in schizophrenia remain elusive. OBJECTIVE: The goal of the present study was to assess specific components of attention, including disengagement, movement, re-engagement, and the inhibitory processes involved their control. METHODS: Thirteen chronic schizophrenics from the inpatient and outpatient units of the Veterans Administration Medical Center (New Orleans, LA) and thirteen normal control subjects were administered a saccadic eye movements task. Saccade latency was measured in the presence of contra-lateral distracter stimuli that preceded the target onset (Distracter-before), followed the target onset (Distracter - after) or in the absence of a distracter (No-distracter). In order to assess the interactive process of fixation disengagement and target selection, fixation was either offset before the target (Gap) or it remained on in the presence of the target (Overlap). RESULTS: Repeated measures analysis of variance revealed that saccadic latency in patients with schizophrenia is prolonged to a greater extent than in normal control subjects in the presence of distracter stimuli. Patients with schizophrenia are also characterized by a greater percentage of error saccades directed to the distracter, and require a longer latency to "issue" corrective saccades following error saccades. CONCLUSIONS: The findings suggest that patients with schizophrenia are required to invoke volitional control under distracter conditions, whereas normal control subjects require minimal volitional control. The results are interpreted in terms of the inhibitory mechanisms that regulate attention. PMID- 10527106 TI - Smooth pursuit tracking deficits of patients with schizophrenia at specific within-sine wave bins. AB - BACKGROUND: Early information processing deficits are consistently reported for patients with schizophrenia on smooth pursuit tracking tasks. A growing number of studies have applied a transient (magnocellular) or sustained (parvocellular) explanation to account for deficient processing of briefly presented visual stimuli, moving stimuli, and particularly, stimuli requiring smooth tracking eye movements in patients with schizophrenia. OBJECTIVE: Although the preponderance of findings offer support for transient (where is it?) as opposed to sustained (what is it?) deficit, a need remains for specific depiction of the deficit. This was accomplished by applying a unique analytic method to a smooth pursuit tracking task. METHODS: Fourteen patients with schizophrenia and fifteen normal control subjects were tested on smooth pursuit tracking performance at five different "within-wave" dot velocity frequencies that ranged from .3 to 1.1 hz. Performance data was extracted from each of the five frequencies and then separated into 12 discrete components that corresponded to light velocity (i.e., 12 bins). RESULTS: A repeated measures multivariate analysis of covariance indicated that the performance of patients with schizophrenia was significantly poorer than that of their normal counterparts for three separate analyses of the time in smooth pursuit, F(11,594) = 8.99; p <0.00001, percentage of time in smooth pursuit, F(11,594) = 3.06; p <0.0005, and time in saccade eye movement, F(11,594) = 3.11; p <0.0004. A regression analysis revealed that the medication dosage was not significantly associated with performance on any of the critical measures, although trends were observed. CONCLUSIONS: The findings provide support for an early information processing deficit in patients with schizophrenia. In addition, the results support the current neurophysiologic model for abnormal smooth pursuit tracking in patients with schizophrenia, specifically implicating a transient channel deficiency. PMID- 10527107 TI - Late-onset paranoid psychosis as a distinct clinicopathologic entity: magnetic resonance imaging data in elderly patients with paranoid psychosis of late onset and schizophrenia of early onset. AB - OBJECTIVE: The aim of this study is to equalize the influence of age-related changes and to test the hypothesis that specific structural brain changes are mediating the development of unique clinical features in late-onset paranoid psychosis (LOPP). BACKGROUND: Findings of unique white matter lesions have been recently described in patients with LOPP. These findings have not been consistent, however, when age-matched normal subjects have been used as a control group. METHOD: Magnetic resonance imaging data were compared in 13 patients with LOPP, mean age 66.33, and 35 elderly patients with early-onset paranoid schizophrenia (PSCH), mean age 63.89. Patients in the LOPP group differed from the PSCH group by the mild degree or absence of negative symptoms, the absence of formal thought disorders, and by prevalence of female patients. RESULTS: Analysis of the magnetic resonance imaging data revealed statistically significant differences between the LOPP and PSCH groups. White matter hyperintensity was almost threefold more frequent in LOPP than in PSCH groups, 69.2% versus 22.9% respectively. Ventricular enlargement and cortical atrophy were more frequent in the PSCH group, reaching, for moderate to severe abnormalities, 28.6% for ventricular enlargement and 22.9% for cortical atrophy; moderate to severe abnormalities were absent in all 13 patients of the LOPP group. CONCLUSIONS: These data point to the possibility that late-onset paranoid psychosis is a distinct clinicopathological entity, with white matter hyperintensity mediating the development of LOPP in a significant percentage of the cases. The vascular origin of white matter lesions in LOPP is suggested. PMID- 10527108 TI - The crucial role of frontostriatal circuits for depressive disorders in the postacute stage after stroke. AB - OBJECTIVE: This study analyzes lesion configuration in patients in the post-acute stage after first single unilateral stroke who suffered from depressive disorders. BACKGROUND: Recent studies indicate a biological origin of poststroke depressive disorders. Due to differences in times of investigation, methods applied, and patient selection, most data are not comparable. Furthermore, only a few studies of poststroke depression report detailed neuropsychologic assessments. METHODS: We investigated 20 consecutive patients who were diagnosed as depressive according to DSM-III-R criteria and exhibited no other severe illness, had no history of neurologic or psychiatric disease, and who were either not aphasic, or only mildly aphasic. A structured clinical interview, self-based and observer-based depression rating scales, a comprehensive neuropsychologic and neurologic examination and ADL-measurement were applied. Neuroradiologic analysis was based on standardized computed tomography scans. RESULTS: Nine of 10 subjects with left hemisphere strokes exhibited a major depression and 7 of 10 subjects with right hemisphere infarcts a minor depression. The most prominent neuropsychologic deficits were found in frontal lobe associated tasks. Type and severity of depression were not related to the severity of neurologic symptoms or impairment in activities of daily living. For both major and minor depression the maximal overlap of lesions was found in subcortical areas, including parts of the caudate nucleus, posterior parts of the putamen, and the deep white matter. CONCLUSIONS: The findings support the theory that poststroke depression is related to the dysfunction of (cortico-) striato-pallido-thalamic-cortical projections that modulate cortico-thalamo-cortical loop systems. PMID- 10527110 TI - Emotional versus nonemotional lexical perception in patients with right and left brain damage. AB - OBJECTIVE: This study examined lexical emotional perception in patients with unilateral brain damage. BACKGROUND: Hypotheses pertaining to laterality and emotion were tested. More specifically, we were interested in whether the right hemisphere is dominant for verbally-presented emotion. In addition, we examined whether emotional content improves the performance of patients with left brain damage (LBD) and language deficits. METHOD: Subjects were 11 patients with right brain damage (RBD), 10 patients with LBD, and 15 normal control adults. The subject groups did not differ significantly on demographic or basic cognitive variables; the patient groups were similar on neurologic variables. Parallel emotional experimental and nonemotional control tasks included word identification (or recognition), sentence identification, and word discrimination. There were eight emotional categories (e.g., happiness) and eight nonemotional categories (e.g., vision). RESULTS: A significant interaction among Group, Condition, and Task revealed that patients with RBD were significantly impaired relative to patients with LBD and normals within the emotional condition, particularly for the identification tasks. Furthermore, the performance of patients with LBD and language deficits was improved by emotional content for the sentence identification task. CONCLUSIONS: These findings suggest that the right hemisphere has a unique contribution in the identification of lexical emotional stimuli. Implications for rehabilitation of patients with LBD and language deficits and patients with RBD by means of emotion-based strategies are discussed. PMID- 10527109 TI - Anatomic asymmetries of the posterior superior temporal lobes: a postmortem study. AB - OBJECTIVE: To examine for structural asymmetries in the posterior superior temporal lobe at the microscopic level in an effort to explain the gross anatomical and functional asymmetries of this brain region. BACKGROUND: The posterior superior temporal lobe is typically larger on the left and damage to this area frequently results in an aphasia. This has led to the hypothesis that the structural asymmetry determines the functional asymmetry, but no definite confirmation of this hypothesis exists. METHODS: Sixteen men were studied at postmortem. Posterior superior temporal lobe dimensions, gray matter volume, white matter volume, SMI-32 immunopositive neuronal density, and glia cell volume were measured for both the left and right hemispheres. In a subset of eight subjects, myelin sheath and axon diameters were measured with electron microscopy. RESULTS: Posterior superior temporal lobe white matter volume was greater on the left (p = 0.003, t test for dependent samples). This asymmetry did not appear to be the result of an isolated proliferation of glia (p = 0.46, t test for dependent samples), nor the density of cortical to cortical projections neurons in the overlying cortex (p = 0.71, t test for dependent samples). In a subset of eight subjects studied with electron microscopy, axons of the left posterior superior temporal lobe were more thickly myelinated (57 nm [SD = 27] left, 46 nm [SD = 24], p < 0.001, ANOVA). CONCLUSIONS: As axons with thicker myelin sheaths conduct faster and require a greater volume, these results suggest asymmetry of myelination as an explanation for both a left hemisphere dominance for rapid sensory signal processing, leading to a functional asymmetry for language, and a larger left planum temporale. PMID- 10527111 TI - Explicit and implicit learning in patients with Alzheimer disease and Parkinson disease with dementia. AB - OBJECTIVE: To examine the differential impairment of implicit and explicit memory systems in cortical and subcortical dementias. BACKGROUND: Whereas verbal priming was reported to be impaired in patients with Alzheimer Disease (AD), patients with Parkinson Disease (PD) may be relatively more impaired on tasks of motor skill learning. METHODS: We examined 15 patients with Alzheimer disease, 10 patients with Parkinson disease and dementia (PD-D), 15 patients with PD but no dementia, and 24 age-comparable normal control subjects with a neuropsychologic battery that included tests of explicit memory (Buschke Selective Reminding Test, Benton Visual Retention Test, Digits Span), and tests of implicit memory (Word Stem Completion task and the Maze Test). RESULTS: AD and PD-D groups showed similar deficits on all measures of explicit memory, and performed significantly worse than PD patients without dementia and normal control subjects. On the other hand, there were no significant between-group differences in any of the measures of implicit memory. CONCLUSIONS: Our study demonstrated preserved implicit learning in the context of severe explicit learning deficits in patients with dementia, but could not demonstrate a different profile of memory deficits between so-called cortical and subcortical dementias. PMID- 10527112 TI - Telephone adaptation of the Modified Mini-Mental State Exam (3MS). The Cache County Study. AB - OBJECTIVE: To examine the concurrent validity of a newly developed telephone adaptation of the Modified Mini-Mental State Exam. BACKGROUND: Longitudinal studies of cognition may be advantaged by availability of assessment instruments that can be used over the telephone, as well as in person. METHOD: Subjects were 263 noninstitutionalized elderly residents of a rural community in southern Idaho, aged 65 to 93, who had little or no cognitive difficulty. At an average interval of four weeks, we administered the Modified Mini-Mental State Exam (3MS) and the newly adapted Telephone Modified Mini-Mental State Exam (T3MS). Order of administration was randomly assigned. RESULTS: Agreement between scores on the two instruments was good (r = 0.82, p < 0.001). When we applied various cutoff scores to the instruments, thereby generating assignments of individuals to "screen positive" and "screen negative" groups, the percent agreement in screening results ranged from 80% to 96% as we reduced the cutoff scores from 90 to 74 (100 points possible). CONCLUSIONS: At least among subjects without major cognitive syndromes, the Telephone Modified Mini-Mental State Exam provides a reasonable substitute for the more costly in-person 3MS. The telephone instrument should now be tested over a broader range of cognitive abilities in order to assess its validity in more impaired subjects, e.g., by studying an institutionalized sample. PMID- 10527113 TI - The animal scientist in a changing society. AB - Despite the lack of any credible scientific evidence to oppose the use of animal performance-enhancing agents, acceptance of performance enhancers seems no closer than it was a decade ago--at least among the European Community and its major trading partners. Consumers are suspicious of new technologies, and politicians are wary of legalizing growth promoters when the relative price of animal products has never been cheaper. Among the factors that have recently re-fuelled consumer concerns over farming methods are: bovine spongiform encephalopathy, cloning of farm animals, and genetic manipulation of crops. Meanwhile, politicians try to balance the interests of the environmentalist, farming, and welfare lobbies with the politico-economic realities of an expanding European Community and the demands of the GATT agreement. In the United States, where corporate influence over political actions is more overtly established than in Europe, some new technologies have been introduced. This has further antagonized many consumers. As scientists with a direct interest in animal performance enhancers, we need to re-assess our positions--if for no other reason than to protect our research (and personal) incomes. We could probably better protect our own interests--and those of the farming community--if we raised our eyes from the microscope to look at the wider view. There are two challenges for animal production scientists: to identify truly acceptable ways of enhancing animal performance and to be highly active in bringing scientific consensus to the attention of both the public and the political establishments. PMID- 10527114 TI - Bovine somatotropin and lactation: from basic science to commercial application. AB - Bovine somatotropin (bST) results in increased milk yield and an unprecedented improvement in efficiency. Beginning in the 1930s to present day, investigations have examined animal-related factors such as nutrition, bioenergetics, metabolism, health and well being and consumer-related factors such as milk quality, manufacturing characteristics, and product safety. Overall, bST is a homeorhetic control involved in orchestrating many physiological processes. Direct effects involve adaptations in many tissues and the metabolism of all nutrient classes--carbohydrates, lipids, protein, and minerals. Mechanisms include alterations in key enzymes, intracellular signal transduction systems, and tissue response to homeostatic signals. Indirect effects involve the mammary gland and are thought to be mediated by the insulin-like growth factor (IGF) system. Specific changes include increased cellular rates of milk synthesis and enhanced maintenance of secretory cells. Indirect effects are modulated by environment and management factors, especially nutritional status. This modulation is a central component in allowing ST to play a key role in regulating nutrient utilization across a range of physiological situations. U.S. commercial use began in 1994, and adoption has been extensive. From a consumer perspective, bST was unique, and special interest groups loudly predicted dire consequences. However, introduction of bST had no impact on milk consumption, and milk labeled as recombinant bST-free occupies a minor niche market. From a producer perspective, commercial use verified scientific studies and enhanced net farm income. Overall, ST is a key homeorhetic control regulating nutrient partitioning, and the ST/IGF system plays a key role in animal performance and well being across a range of physiological situations. PMID- 10527115 TI - Growth hormone and mammary development. AB - Classic studies in rodents conducted in the 1950s showed that growth hormone (GH) is essential for mammary development both in the pubertal phase and during pregnancy. Since then, a considerable number of experiments have been carried out in ruminants to investigate the role of GH for regulation of normal mammary development and to examine the possibility of enhancing mammary growth by administration of GH. The available evidence demonstrates that GH treatment stimulates mammary growth before puberty, but the data do not convincingly support the idea that the effect is translated into increased milk yield. GH treatment during late pregnancy seems to stimulate both mammary growth and milk yield during lactation. The limited data concerning the effect of GH on mammary growth during lactation indicate that mammary growth is unaffected by GH treatment in early lactation, whereas GH seems to increase the amount of mammary parenchyma in mid-lactation. The mechanism of action of GH remains a puzzle, but the effect of exogenous GH most likely involves insulin-like growth factor-I (IGF I). Full understanding of the role of endogenous GH for regulation of normal mammary development requires more knowledge about the interaction between GH and IGF-I and the interplay between the GH-IGF-I axis and locally produced factors, including receptors, binding proteins, and growth factors. PMID- 10527116 TI - Manipulation of milk production and quality by use of somatotropin in dairy ruminants other than cow. AB - The ability of recombinant bovine somatotropin (BST) to enhance milk production is well established in cows and in other dairy ruminants. In dairy ewes, we found increased milk yield (20-30%) following treatment with BST, which did not negatively affect the gross composition or coagulating properties of milk, except in the advanced stage of lactation, when the percentages of milk protein and fat were reduced and the coagulation time was improved (shorter) compared with untreated animals. In dairy goats, administration of BST increased overall milk yield by 14-29%. Our studies and those of others on the Italian river buffalo showed that BST treatment increased milk yield by about 17%, or more, when associated with dietary protected fat, without affecting milk protein content. In general, studies on dairy ruminants show that treatment with BST increases milk production in the short term (immediate postinjection period) and that there is also a medium to long term effect on persistency of lactation. There is evidence that mammary gland involution can be at least partially reversed by BST administration, and this could be due to limitation in the decrease in mammary parenchyma as lactation progresses and/or to modulation of the plasmin plasminogen system. PMID- 10527117 TI - Candidate gene markers associated with somatotropic axis and milk selection. AB - One of the obstacles to progress in dairy cattle selection is that milk production traits are only expressed after the first calving. However, the use of the quantitative trait loci (QTL) technology will improve the efficiency of dairy industry with a positive image for the consumers. QTL are part of the genome showing a preponderant action and explaining the major part of variation of the trait production. At the present time, the two major strategies developed to detect such QTL are the candidate gene approach and the positional genetics approach. The somatotropic axis contains the most promising candidates in this respect, as it strongly regulates milk production. Then, the identification of favorable QTL associated with the somatotropic axis that are significantly correlated with genetic merits for milk production could lead to more effective selection programs. PMID- 10527118 TI - Modulation of the inflammatory reaction and neutrophil defense of the bovine lactating mammary gland by growth hormone. AB - This review is focused on the possible interactions of prolactin and somatotrope hormone in the modulation of inflammation of the mammary gland. Several different models are examined: Escherichia coli, Streptococcus uberis, and endotoxin mastitis. Subsequently, the release of growth hormone and insulin-like growth factor during fever and mastitis, the immunophysiological effects of GH on E. coli mastitis, S. uberis and endotoxin mastitis, the galactopoietic action of rBST on healthy and mastitis cows as well as the immunologic effects of GH on leukocytes in healthy and diseased cows are discussed. It can be concluded that the underlying regulation of the neuro-endocrine network is fundamental in the normal function of the immune system. PMID- 10527119 TI - Presence of thyroxine deiodinases in mammary gland: possible modulation of the enzyme-deiodinating activity by somatotropin. AB - Thyroid hormones (TH) and somatotropin (ST) play critical role in lactation. One explanation of their multiple physiological actions is based on the functional interrelationships among ST, TH, and thyroxin deiodinase (5'D). This enzyme is present in the mammary tissue, milk cellular components, and whole milk and is responsible for intramammary production of triiodothyronine (T3). In rats in which the 5'D isozymes in the mammary gland and in the liver are similarly of type I (5'D-I), an enhancement of mammary 5'D-I causes a reduction of hepatic 5'D I activities. This opposite rearrangement in the mammary and hepatic deiodinating activities is thought to be a factor of a homeorhetic response characterized by an increased and compartmentalized energy expenditure of the mammary gland. In the cow, the mammary 5'D is the type II (5'D-II) deiodinase. The 5'D-II, owing to its high catalytic efficiency, secures T3 production, making tissues relatively independent from the circulatory levels of TH and from variations in the hepatic 5'D-I activity. No significant alterations of 5'D-II isozymes were found during a low T3 syndrome. Location of tissue deiodinases in the cow, the 5'D-II in the mammary gland, and the 5'D-I in the liver make it so that T3 production in these two tissues can be dissociated in time to secure better local requirement for T3 supporting lactation. To date, attempts to evidence that the alterations in iodothyronines blood levels and in tissues' 5'Ds activity during lactation are due to ST action have not received clear experimental support in either cows or rats. PMID- 10527120 TI - Emerging strategies for enhancing growth: is there a biotechnology better than somatotropin? AB - During the past 20 years, there have been many impressive advances in a number of scientific disciplines that have led to the discovery and development of exciting new biotechnologies that offer the potential to improve productive efficiency of animal agriculture. Some technologies have been developed from advances made in our understanding of how the endocrine system regulates growth and lactation. This information then has been used to devise viable strategies that alter circulating hormone concentration(s) to enhance animal production and productive efficiency. The most notable success to date using this approach has been bovine somatotropin, which has been adopted for use in the dairy industry in certain countries. Advances in transgenic biology, gene therapy, "knock-out" gene technologies, and cloning may lead to other novel products/strategies that enhance productive efficiency. The purpose of this paper is to discuss what future strategies might emerge that will increase meat and milk production and the efficiency of these processes. PMID- 10527121 TI - New insights into the mechanism and actions of growth hormone (GH) in poultry. AB - Despite well documented anabolic effects of GH in mammals, a clear demonstration of such responses in domestic poultry is lacking. Recently, comprehensive dose response studies of GH have been conducted in broilers during late post-hatch development (8 to 9 weeks of age). GH reduced feed intake (FI) and body weight gain in a dose-dependent manner, whereas birds pair-fed to the level of voluntary FI of GH-infused birds did not differ from controls. The reduction in voluntary FI may involve centrally mediated mechanisms, as hypothalamic neuropeptide Y protein and mRNA were reduced with GH, coincident with the maximal depression in FI. Growth of breast muscle was also reduced in a dose-dependent manner. Circulating IGF-I was not enhanced by GH, despite evidence that early events in the GH signaling pathway were intact. A GH dose-dependent increase in circulating 3,3',5-triiodothyronine(T3) paralleled decreases in hepatic 5D-III monodeiodinase activity, whereas 5'D-I activity was not altered. This confirms that a marked hyperthyroid response to GH occurs in late posthatch chickens, resulting from a decrease in the degradative pathway of T3 metabolism. This secondary hyperthyroidism would account for the decreased skeletal muscle mass (52) and lack of enhanced IGF-I (53) in GH-treated birds. Based upon these studies, it is now evident that GH does in fact have significant effects in poultry, but metabolic responses may confound the anabolic potential of the hormone. PMID- 10527122 TI - Growth factors controlling muscle development. AB - The enlarged muscles of certain breeds of cattle, such as the Belgian Blue, have been shown to result from a marked increase in the number of normal sized muscle fibers. Originally insulin-like growth factors (IGFs) were implicated in this myofiber hyperplasia, as IGFs have been shown to stimulate myoblast proliferation as well as maintain fiber differentiation. Recently it has been reported that mice lacking a myostatin gene, a member of the TGFbeta superfamily, have enhanced skeletal mass resulting from increased muscle fiber number and size. Mutations in this gene have been found in double-muscled cattle, indicating that myostatin is an inhibitor of muscle growth. Myostatin is expressed early in gestation and then maintained to adulthood in certain muscles. Myostatin expression in bovine muscle is highest during gestation when muscle fibers are forming and some of the myogenic regulatory factors have elevated expression over the same period as myostatin. Molecular expression of the IGF axis does not differ between Belgian Blue and normal muscled cattle, and IGF-II mRNA is increased throughout formation of secondary fibers in both breeds. However, myostatin and MyoD expression in muscle differ between normal and hypertrophied muscle cattle breeds. This evidence strongly suggests that lack of myostatin is associated with an increase in fiber number which then results in a marked increase in potential muscle mass in double-muscled cattle. PMID- 10527123 TI - Role of melatonin in the control of growth and growth hormone secretion in poultry. AB - The pineal hormone melatonin controls reproduction of photoperiodic mammals and is an integral part of the circadian organization in birds. Recent findings indicate an involvement of this hormone also in more basic physiological processes, including growth, development, and aging. Melatonin may modulate growth in poultry through interaction with transcriptional factors, through interaction with hormones involved in growth control, and by modulation of energy metabolism and decreasing physical activity. Our studies showed that a single melatonin injection increased plasma growth hormone (GH) concentrations in the Japanese quail. Specific serotonin receptor blocker ketanserin did not preclude a stimulatory action of melatonin on GH synthesis. Serotonin agonist quipazine increased GH levels but failed to enhance the stimulatory effect of melatonin. Pretreatment with melatonin in drinking water did not affect the magnitude of the GH response to subcutaneous (s.c.) administration of thyrotropin releasing hormone (TRH) that considerably stimulated GH secretion. Present data suggest that melatonin modulates rather central neural pathways involved in the control of GH synthesis at the hypothalamic level than the sensitivity of the pituitary gland. PMID- 10527124 TI - Regulation of protein and energy metabolism by the somatotropic axis. AB - The somatotropic axis plays a key role in the co-ordination of protein and energy metabolism during postnatal growth. This review discusses the complexity of the regulation of protein and energy metabolism by the somatotropic axis using three main examples: reduced nutrition, growth hormone (GH) treatment and insulin-like growth factor-1 (IGF-1) treatment. Decreased nutrition leads to elevated GH secretion, but it reduces hepatic GH receptor (GHR) number and plasma levels of IGF-1; it also changes the relative concentrations of IGF binding proteins (IGFBPs) in plasma. GH treatment improves the partitioning of nutrients by increasing protein synthesis and decreasing protein degradation and by modifying carbohydrate and lipid metabolism. However, these well-established metabolic responses to GH can change markedly in conditions of reduced nutritional supply or metabolic stress. Short-term infusion of IGF-1 in lambs reduces protein breakdown and increases protein synthesis. However, long-term IGF-1 administration in yearling sheep does not alter body weight gain or carcass composition. The lack of effect of IGF-1 treatment can be explained by activation of feedback mechanisms within the somatotropic axis, which lead to a reduction in GH secretion and hepatic GHR levels. The somatotropic axis has multiple levels of hormone action, with complex feedback and control mechanisms, from gene expression to regulation of mature peptide action. Given that GH has a much wider range of biologic functions than previously recognized, advances in research of the somatotropic axis will improve our understanding of the normal growth process and metabolic disorders. PMID- 10527125 TI - Endocrine and metabolic aspects in milk-fed calves. AB - In neonatal calves besides adaptations in organ function there are marked metabolic and endocrine changes. The growth hormone (GH)-insulin-like growth factor (IGF) axis is basically functioning, but needs maturation. Various metabolic and endocrine traits do not exhibit marked ontogenetic changes after the first week of life, but others remain different from the adult stage. Thus, plasma oxytocin or an oxytocin-like substance and nitrate concentrations are elevated for months. The ability to digest colostrum (C) and milk involves great alterations in structure and function of the gastrointestinal (GI) tract. C intake is important for passive immunity, provision of nutrients, minerals and vitamins, and contains biologically active substances. IGF-I, present in C in high amounts, appears to enhance GI tract development and function. For sufficient absorption not only of immunoglobulins, but also of fatty acids and fat-soluble vitamins, C should be ingested immediately after birth. The amino acid pattern and the glutamine/glutamate ratio depends greatly on whether C is fed or not. Effects on insulin, IGF-I, and IGF binding proteins depend on time point and amounts of C fed. After the colostral period calves are almost exclusively fed milk and milk substitutes or weaned. Low iron intake, required for the production of pale meat, besides anemia causes metabolic and endocrine adaptations, such as enhanced insulin-dependent glucose utilization and appears to reduce IGF-I responses to GH. Metabolic and endocrine changes, such as insulin resistance and disturbed glucose metabolism, can be observed in part in association with high feeding intensity in veal calves. PMID- 10527126 TI - Genetic models for the study of insulin-like growth factors (IGF) and muscle development in birds compared to mammals. AB - IGFs are important positive modulators of overall body and muscle growth in different species. Genetic variation in IGF-I gene expression exists as shown by the possibility of genetic selection for high or low circulating IGF-I concentrations in mice and the associated variations in growth potential. Targeted over-expression of IGF-I in transgenic mice results in muscle hypertrophy, but it is yet unknown whether genetic variability in muscle IGF-I gene expression exists. Much less data are available in birds. This review is focussed on the potential role of IGFs on chicken muscle development. Apart from the absence of a type 2 IGF receptor (IGF-II receptor), the general characteristics of the chicken IGF system seem to be similar to mammalian species. In different genetic models with altered growth rates or body composition, differences in the IGF system are observed suggesting its importance in regulating body growth in those species. The components for a paracrine action of IGF are also present in chicken muscle but it has not yet been demonstrated if they contribute to differences in muscle development. PMID- 10527127 TI - The role of the somatotrophic axis in the metabolism of the chicken. AB - As it is for mammalian species, growth hormone (GH) is indispensable for normal growth and development of avian species. In contrast to mammals, exogenous GH administration has little, if any, potential for improving the growth rate and feed efficiency of rapidly growing broilers; it is more likely to do so in older birds. This is at least partly because of age-related changes in tissue GH binding activity and GH-receptor mRNA expression. The effects of GH on lipid deposition depends on the age of the bird and pattern of GH administration. Pulsatile, but not continuous, GH administration to older broilers seems to reduce fat deposition. As in rats, the bioactivity of GH might also depend on the pulse-induced cyclicity in GH receptors and GH-binding proteins. In chickens, GH is also a very potent lipolytic hormone, but seems to have no diabetogenic effect, which is reported in mammalian species. Both insulin-like growth factors have apparently no growth-promoting effects in normal growing broilers, but seem to have opposite effects on fat deposition. In contrast to GH, both insulin-like growth factors have a marked hypoglycemic effect. Whether all these effects are direct effects, or are mediated by secondary mechanisms, awaits further investigations. PMID- 10527128 TI - Regulation of porcine adipogenesis in vitro, as compared with other species. AB - In vitro studies, mostly performed on murine cell lines, allowed us to identify the role played by hormonal agents, second-messenger pathways, extracellular matrix proteins, and transcription factors in adipose conversion. Some information has also been reported when studies were conducted on primary cultures that originated from various species. However, because of conflicting results, probably caused, at least in part, by species specificity, developing cultures of preadipose cells from economically important species appeared necessary to better understand and control the animals' fat development. We reviewed our current knowledge concerning the regulation of cultured porcine preadipose cells by hormones, second-messenger pathways, and extracellular matrix proteins. The results clearly demonstrate that such primary cultures are essential to avoid the establishment of hazardous concepts originated from rodent and aneuploid cell lines in particular. PMID- 10527129 TI - The effects of exogenous growth hormone on follicular steroid secretion and ovulation rate in sheep. AB - Growth hormone (GH) has diverse actions in many tissues, including the follicle. This paper summarizes three experiments that examined the effects of GH and insulin-like growth factor (IGF)-I on the ovary. Ewes given oGH and pregnant mane serum gonadotrophin were compared with control and pregnant mane serum gonadotrophin-treated ewes. Ewes, with synchronized cycles, were given varying doses of pregnant mane serum gonadotrophin and/or oGH to determine if oGH is able to augment ovulation rate (Experiment 1). Experiments 2 and 3 used the ovarian autotransplant model. Ewes were infused via the ovarian artery with oGH (Experiment 2) or insulin-like growth factor I (IGF-I) (Experiment 3). Both were administered for 12 hr on Day 10. In Experiment 2, ewes were given intravenous gonadotropin releasing hormone (150 ng i.v.) at -2.5 and 10.5 hr relative to infusion. Ovarian and jugular venous blood was collected every 15 min from -30 to 150 min relative to gonadotropin releasing hormone. In Experiment 3, luteolysis was induced at the end of infusion. Ovarian and jugular venous blood was collected every 3 hr from before and until 84 hr after the infusion. Estradiol and androstenedione were assayed in ovarian venous plasma and GH in jugular venous plasma. In Experiment 1, treatment with oGH increased the jugular venous concentration of GH. However, in Experiment 2 treatment with oGH via the ovarian artery did not increase jugular venous GH but did increase ovarian venous GH. Treatment with oGH had no effect on ovulation rate (Experiment 1) or the secretion of androstenedione and estradiol (Experiment 2). Infusion of IGF-I (Experiment 3) increased the secretion of estradiol during the follicular phase. These data show that short-term treatment of sheep with GH had no in vivo effects on the follicle and that IGF-I was a potent stimulator of follicular steroidogenesis in vivo. PMID- 10527130 TI - Possible role of growth hormone, IGFs, and IGF-binding proteins in the regulation of ovarian function in large farm animals. AB - The aim of the study and short review was to present evidence that growth hormone (GH), locally produced insulin-like growth factors (IGFs), and IGF-binding proteins (IGFBPs) may have an important role in the control of ovarian function. There is clear evidence for a distinct GH-receptor mRNA expression and protein production in follicles (oocytes and granulosa-cumulus cells) and corpus luteum (CL). In hypophysectomized ewes, GH and LH are necessary for normal CL development. IGF-1 mRNA in the follicles is expressed in theca interstitial cells (TIC) and granulosa cells (GC) with already higher levels in the TIC before follicle selection. In contrast, IGF-2 is mainly expressed in the TIC. The IGFR-1 mRNA is expressed in both the TIC and GC, with increasing levels in GC during the final development of dominant follicles. IGF-1 is a very potent stimulator of progesterone and oxytocin release in GC. IGFBP-1, -2, -3, -4, -5, and -6 have been isolated from follicular fluid or ovarian tissue. Studies indicate that IGFBP expression and production in the developing follicle is dependent on both cell type and follicle size and is regulated by IGF-1 and gonadotropins. The highest expression of IGF-1 and IGFR-1 mRNA was demonstrated during the early luteal phase. Distinct receptors for IGF-1 and IGF-2 were present in CL membrane preparations at all stages investigated. Intense immunostaining for IGF-1 was observed mainly in bovine large and small luteal cells and in a limited number of endothelial cells. In contrast, IGF-2 protein was localized in perivascular fibroblast and pericytes of the capillaries. With the use of a microdialysis system, we found that in vitro and in vivo IGF-1, IGF-2, and GH stimulated the release of progesterone in cultures of luteal cells or intact tissues. In conclusion, there is clear evidence for a central role of the IGFs, IGFBPs, and GH in follicular development and CL function. PMID- 10527131 TI - Effects of exogenous somatotropin (ST) on gonadal function in ruminants and swine. AB - During the past 15 years, many investigators have examined the effects of somatotropin (ST) on growth and lactation in farm animals. Throughout this period, concerns about potential effects of ST on reproduction have been expressed. The objective of the present review will be to focus on the effects of exogenous ST on the hypothalamic-pituitary-gonadal axis. Plasma progesterone is increased when recombinant bovine (rb)ST is given to cattle, early in lactation, and also to sheep. Also, the release of progesterone from cultured swine and human luteal cells is increased by ST. Treatment with rbSt increases the numbers of small follicles, but does not increase the ovulatory rate of ruminants. Doses of ST similar to those used to increase milk production do not affect the secretion of testosterone or spermatogenesis in rams or bulls. Stimulatory and inhibitory effects of exogenous ST on reproductive function of gilts have been reported. Daily injections of porcine ST (pST) delayed puberty and expression of estrus after puberty. Daily administration of pST increased the number of small follicles, but not of medium follicles, whereas administration of pST by using a sustained release implant increased the number of medium follicles. Size and weight of reproductive organs and concentration of testosterone are not affected when pST is administered for at least 42 d. However, pST enhanced testicular development and spermatogenesis when given to neonatal boars. In summary, administration of exogenous ST at doses known to alter milk production and carcass composition may have subtle positive and/or negative effects on the reproductive systems of cattle and swine; however, these effects appear to be transient. PMID- 10527132 TI - Insulin-like growth factors in the regulation of avian ovarian functions. AB - In the past three decades, overwhelming evidence has accumulated to show that insulin-like growth factor (IGF)-I and -II, their receptors and binding proteins (IGFBP) (the IGF system), have major roles to play in the regulation of ovarian function in mammals. Although studies in birds did not start until 5-6 years ago, the limited information thus far available suggests that the IGFs act as autocrine/paracrine regulators of follicular growth and differentiation, just as observed in mammals. The genes for IGF-I and -II, type-I IGF receptor, IGFBP-2, and IGFBP-5 are expressed in both granulosa and theca cells of the chicken ovary. The mechanisms by which the IGF system controls ovarian function in the avian species are complex and involve interactions with the gonadotrophins (LH and FSH), growth hormone, and even other growth factors. Effects are different between strains and nutritional status. PMID- 10527133 TI - Control of ovarian follicles activity in the ewe. AB - During the ovine estrous cycles, three waves of follicular growth, closely associated with the FSH secretion pattern, were observed. The parameters of these follicular waves and the ability of follicles to produce steroids in vitro were studied in various conditions. In vivo, the follicular events were similar between the breeding season and the anestrus, except for the lack of ovulation; but at the end of the breeding season and in anestrus, the follicles lose a big part of their aromatization ability. In ewes carrying the Booroola fecundity gene or Cambridge fecundity gene, the reduction in follicular atresia seems to be one of the main follicular features implicated in the control of high ovulation rate. In vitro, the most relevant difference is an early acquisition of estrogen production ability of small follicles in Booroola fecundity gene barring ewes. Fluoro-gestone-acetate (FGA) pessaries reduced the number of growing follicles; despite this effect disappearing after the sponge withdrawal, the ovulation rate is significantly reduced. But an equine chorionic gonadotrophin (eCG) treatment restores the ovulation rate (OR) by reducing the atresia rate of pre-ovulatory follicles. In similar conditions, a pretreatment of the ewes with melatonin again reduced the atresia rate of large follicles and resulted in an increased ovulation rate. In vitro, FGA blocked aromatization ability, and melatonin inhibited both androstenedione and estradiol production, but a further treatment with eCG partly restores the steroid secretion. Immunization against androstenedione leads to a higher OR, owning to a reduced atresia of large follicles. Daily growth hormone injections for a hole cycle resulted in an increased follicular population and ovulation rate, while FSH plasma levels decreased and the follicle sensitivity to gonadotrophins was reduced. PMID- 10527134 TI - MR cholangiopancreatography techniques. AB - The tissue contrast principles and the technical aspects involved in the design of the imaging protocols currently used for clinical MR cholangiopancreatography are reviewed using a neutral terminology that is applicable to most of the high field MRI equipment available from the major manufacturers. Furthermore, the technical discussions that follow are accompanied by a comprehensive set of tables listing the pulse sequence parameters used by the authors of the other articles in this issue. The tables are organized according to groups of parameters that determine the fundamental features of the protocols and of the generated images, specifically motion artifact reduction technique, scan geometry, image contrast, and recommended image post processing algorithm. PMID- 10527135 TI - Benign diseases of the biliary tract: evaluation with MR cholangiography. AB - MR cholangiography (MRC) is a noninvasive, rapid means of evaluating the biliary tract that, in many instances, may replace invasive procedures such as diagnostic endoscopic retrograde cholangiography and percutaneous transhepatic cholangiography. This article describes and illustrates the MRC features of benign diseases of the biliary tract such as choledocholithiasis; intrahepatic bile duct calculi; congenital anomalies, including aberrant bile ducts and choledochal cysts; postsurgical strictures; and strictures related to chronic pancreatitis. PMID- 10527136 TI - Bile duct stones: diagnosis with MR cholangiography and helical CT. AB - MR cholangiography (MRC) is a highly accurate, noninvasive method for diagnosing bile duct stones. Various breath-hold and non-breath-hold techniques for MRC have been used successfully to depict stones in the intrahepatic and extrahepatic biliary tree. Although detection of stones with MRC is usually straightforward, attention to technical details is important to avoid false-negative and false positive interpretations. With the advent of helical CT, other options for noninvasive imaging of the bile ducts, such as CT cholangiography, are now available. CT techniques are especially useful when MRI is unavailable or contraindicated, or when the quality of MRC images is suboptimal. PMID- 10527137 TI - MR cholangiopancreatography in malignant biliary obstruction. AB - Malignant lesions of the biliary tract are a frequent occurrence, typically presenting with clinical findings of obstructive jaundice. The authors discuss the role of MR cholangiopancreatography (MRCP) as a second level diagnostic technique, which can provide information regarding not only the location, but also the cause of the obstruction. This can be obtained if MRCP is considered as part of a complete study of the upper abdomen, with acquisition of T1- and T2 weighted images. The "all-in-one" approach may provide the identification, characterization, and staging of the lesion, giving the clinician all the information necessary for the planning of adequate treatment. Typical MR features of cholangiocarcinoma are provided, as well as conventional MR and MRCP findings in pancreatic carcinoma, periampullary carcinoma, and biliary obstruction secondary to hilar lymphadenopathy and metastatic lesions. PMID- 10527138 TI - MR pancreatography. AB - Recent innovations in the field of ultrafast MR imaging have increased the robustness of MR cholangiopancreatography (MRCP). Its complete noninvasiveness and flexible physiological approaches in detecting pancreaticobiliary pathologic conditions are gaining the acceptance of many clinicians. The procedure is also safer and more comfortable both for physicians and patients compared with direct pancreatography or cholangiography. Because of its cost effectiveness and safety, optimized MRCP technologies will gradually replace the diagnostic use of endoscopic retrograde cholangiopancreatography (ERCP). It is also notable that MRCP techniques can be used to obtain physiological/dynamic information that ERCP cannot provide. This article addresses recent advances in MRCP from technological and clinical aspects, focusing on its unique features as a hydrographic technique, and also refers to its limitations. PMID- 10527139 TI - Secretin-enhanced MR pancreatography. AB - MR cholangiopancreatography has now emerged as a noninvasive diagnostic technique that can replace diagnostic endoscopic retrograde cholangiopancreatography in many instances. Recent technical issues include the use of fast single-shot T2 weighted single-slice projections in combination with a negative oral contrast agent and secretin stimulation for assessment of pancreatic flow dynamics and duodenal filling. Potential clinical applications include the evaluation of patients with recurrent pancreatitis and inconclusive CT examination as well as the evaluation of chronic pancreatitis complications whenever endoscopic treatment is suggested. In combination with cross-sectional MR sequences, secretin-enhanced MR pancreatography offers the possibility of a comprehensive examination of the pancreas that provides parenchymal, ductal, and functional information within a single diagnostic modality. PMID- 10527140 TI - Because "it is necessary to affix right ideas to words". Thomas Paine, The Age of Reason, 1794. PMID- 10527141 TI - Facial warming increases the threshold for shivering. AB - A decrease of 1-2 degrees C core temperature provides protection against cerebral ischemia. However, shivering usually prevents reduction in core temperature in unanesthetized patients. Therefore, it was tested whether facial and airway heating increases the shivering threshold and enables core cooling in unanesthetized patients. Nine trials were performed on seven healthy male volunteers. Each subject was positioned supine on a circulating-water mattress (8 15 degrees C) with a convective-air coverlet (15-18 degrees C) extending from the neck to the feet. A dynamic study protocol governed by individualized physiological responses was used. Focal facial (and airway) warming was employed to suppress involuntary motor activity (muscle tensing, shivering) and, thereby, enabling noninvasive cooling to lower the core temperature. The following parameters were monitored: 1) heart rate, 2) blood pressure, 3) core temperature (tympanic, axilla, and rectal), 4) cutaneous temperatures, and 5) a subjective shiver index (scale 1-10). In three, electromyograms and infrared thermographs were also obtained. Upon cooling without facial and airway warming, involuntary motor activity increased until it was widespread. This vigorous motor activity prevented any significant lowering of core temperature or caused it to slightly increase. Subsequently, in all subjects, within seconds after the application of facial focal warming, motor activity was suppressed almost completely, and within minutes core temperatures significantly decreased. Preliminary studies described here indicate that focal facial warming applied during active whole body cooling to initiate mild hypothermia might minimize the need to pharmacologically suppress involuntary motor activity. Such a procedure might be useful for initiating as soon as possible (such as during emergency transport), cerebral mild hypothermia in order to maximize protection and thus improve outcome in neurologically injured patients (head trauma, stroke). PMID- 10527142 TI - Cerebral oxygenation in patients after severe head injury: monitoring and effects of arterial hyperoxia on cerebral blood flow, metabolism and intracranial pressure. AB - Early impaired cerebral blood flow (CBF) after severe head injury (SHI) leads to poor brain tissue oxygen delivery and lactate accumulation. The purpose of this investigation was to elucidate the relationship between CBF, local dialysate lactate (lact(md)) and dialysate glucose (gluc(md)), and brain tissue oxygen levels (PtiO2) under arterial normoxia. The effect of increased brain tissue oxygenation due to high fractions of inspired oxygen (FiO2) on lact(md) and CBF was explored. A total of 47 patients with SHI were enrolled in this studies (Glasgow Coma Score [GCS] < 8). CBF was first assessed in 40 patients at one time point in the first 96 hours (27 +/- 28 hours) after SHI using stable xenon computed tomography (Xe-CT) (30% inspired xenon [FiXe] and 35% FiO2). In a second study, sequential double CBF measurements were performed in 7 patients with 35% FiO2 and 60% FiO2, respectively, with an interval of 30 minutes. In a subsequent study, 14 patients underwent normobaric hyperoxia by increasing FiO2 from 35 +/- 5% to 60% and then 100% over a period of 6 hours. This was done to test the effect of normobaric hyperoxia on lact(md) and brain gluc(md), as measured by local microdialysis. Changes in PtiO2 in response to changes in FiO2 were analyzed by calculating the oxygen reactivity. Oxygen reactivity was then related to the 3-month outcome data. The levels of lact(md) and gluc(md) under hyperoxia were compared with the baseline levels, measured at 35% FiO2. Under normoxic conditions, there was a significant correlation between CBF and PtiO2 (R = 0.7; P < .001). In the sequential double CBF study, however, FiO2 was inversely correlated with CBF (P < .05). In the 14 patients undergoing the 6-hour 100% FiO2 challenge, the mean PtiO2 levels increased to 353 (87% compared with baseline), although the mean lact(md) levels decreased by 38 +/- 16% (P < .05). The PtiO2 response to 100% FiO2 (oxygen reactivity) was inversely correlated with outcome (P < .01). Monitoring PtiO2 after SHI provides valuable information about cerebral oxygenation and substrate delivery. Increasing arterial oxygen tension (PaO2) effectively increased PtiO2, and brain lact(md) was reduced by the same maneuver. PMID- 10527143 TI - Extracranial contribution to cerebral oximetry in brain dead patients: a report of six cases. AB - The near infrared spectroscopy offers a noninvasive method to monitor regional brain oxygenation. The problem with the technique appears to be possible extacranial contribution to the measurements. As a part of another study, we monitored regional saturation (rSO2) in six brain dead patients either during the test for spontaneous respiration or in those not eligible for organ donation, after discontinuation of mechanical ventilation. Relatively normal rSO2 values were obtained after brain death, and the values decreased concomitantly with the hemoglobin saturation of oxygen (SpO2) after the discontinuation of mechanical ventilation. A corresponding decrease in SpO2 and rSO2 suggests extracranial contribution to the measured rSO2. The diagnosis of brain death cannot be made based on this technology; furthermore the presence of extracranial contribution may limit its potential value even in other applications. PMID- 10527145 TI - Monitoring brain PO2, PCO2, and pH during graded levels of hypoxemia in rabbits. AB - Brain ischemia and hypoxia are of concern when they occur following traumatic brain injury because they frequently result in potentially preventable secondary brain damage. In this study, we examined the ability of an implantable catheter (Paratrend 7; Diametrics Medical, St. Paul, MN) to continuously measure brain tissue pH, PCO2, and PO2 during graded levels of hypoxia. Values obtained from this catheter were compared with simultaneous measurements of arterial and sagittal sinus blood. As expected, there was a good correlation between the changes in pH, PCO2, and PO2 in brain tissue and sagittal sinus blood. Brain tissue PO2 was numerically lower than sagittal sinus blood at all inspired levels of oxygen. These data suggest that the Paratrend 7 may be useful in monitoring brain tissue oxygen tension in patients at risk for regional cerebral ischemia and hypoxia. PMID- 10527146 TI - Effects of morphine on cerebral blood flow autoregulation CO2-reactivity in experimental subarachnoid hemorrhage. AB - Previous reports show that naloxone improves ischemic deficits and clinical conditions in patients after subarachnoid hemorrhage (SAH). These observations have raised concern about the routine use of morphine in the treatment of severe headache after SAH. The present study was carried out to investigate the effects of morphine on cerebral vasoreactivity after experimental SAH. Cerebral blood flow (CBF) autoregulation was studied in two groups of eight rats each with experimental SAH. A bolus intravenous injection of morphine, 1 mg/kg, was administered in one group and the other was used as a control group. During eucapnia, CBF was measured by the intracarotid 133Xenon method during decreasing mean arterial blood pressure (MABP). CO2-reactivity was investigated in two corresponding groups where CBF was measured at decreasing PaCO2 levels during constant MABP. Morphine decreased mean baseline CBF by 34% and 26% in the study of autoregulation and CO2-reactivity, respectively. Cerebral blood flow autoregulation was found impaired in both controls and the morphine group. However, the mean slope of the linear regression lines of CBF/MABP was 0.49 +/- 0.32 ml/100g/min/mm Hg in the morphine group, which was significantly lower than 1.24 +/- 0.59 ml/100g/min/mm Hg in the controls (p < 0.05). Also the mean CO2 reactivity was significantly lower, 0.64 +/- 0.53 %/0.1kPa, in the morphine group, compared to 2.36 +/- 0.87 %/0.1kPa in the controls (p < 0.001). The results show that in rats with SAH, morphine partially restores CBF autoregulation but attenuates CO2-reactivity. PMID- 10527144 TI - Median EEG frequency is more sensitive to increases in sympathetic activity than bispectral index. AB - Sympathetic heart rate variability is correlated with the increase in plasma catecholamines during rapid opioid detoxification. We evaluated whether the bispectral index, median frequency, or 95% spectral edge of the electroencephalogram are sensitive to the sympathetic response seen during reversal of opioid dependence. Eight patients undergoing rapid opioid detoxification gave informed consent. Two-channel frontal electroencephalogram was measured. Sympathetic heart rate variability was measured in 256 second segments by Fourier transform of continuous heart rate and the low frequency segment (0.02-0.13 Hz) analyzed for sympathetic function. Patients were anesthetized with propofol infusion. After a 30-60 min steady state, naloxone was infused intravenously at a rate of 25 mg/30 min, followed by an infusion of 1 mg/hr. During induction of anesthesia, sympathetic heart rate variability decreased from 1.80 to 0.3, bispectral index from 86 to 47, median frequency from 10.2 to 3.4, spectral edge from 23.5 to 16.7 (all P<.05). During naloxone infusion, the median percent increase in sympathetic heart rate variability was 487% (P<.05), median frequency increased 163% (P<.05), bispectral index (10%), and spectral edge (7%) did not significantly change. The increase in median frequency was delayed compared to sympathetic heart rate variability and median frequency remained elevated after sympathetic heart rate variability returned to anesthetized baseline in 5 of 8 cases. Our results show that median frequency and sympathetic heart rate variability increase during opioid detoxification, but the time course of each response is different. Median frequency is a more sensitive electroencephalogram indicator of opioid reversal than bispectral index or spectral edge. PMID- 10527147 TI - Albumin or hetastarch improves neurological outcome and decreases volume of brain tissue necrosis but not brain edema following closed-head trauma in rats. AB - The present study examined whether hemodilution with 20% human serum albumin (HSA) or 10% hydroxyethyl starch (HES) improved the outcome from closed-head trauma (CHT) in rats. Rats anesthetized with halothane were given one of three hemodilution solutions (i.e., 20% HSA, 10% HES, or control [0.9% saline]) after CHT or sham surgery. CHT was delivered using a weight drop impact of 0.5 J onto the closed cranium. The hemodilution solution (volume = 1% of body weight) was given just after determining the neurological severity score (NSS) at 1 hour following CHT. The NSS was determined again at 24, 48, and 72 hours following CHT. At 72 hours, brains were removed, and brain edema and brain tissue necrosis volume were determined. Solutions of 20% HSA and 10% HES significantly improved brain tissue necrosis volume (143 +/- 72 mm3 and 104 +/- 53 mm3 as compared to 271 +/- 65 mm3 in controls, mean +/- SD) and NSS (12 +/- 2 and 9 +/- 2 as compared to 15 +/- 2 in controls at 72 hours, median +/- range) but not brain edema. The hematocrit decreased similarly in all groups during hemodilution. Hemodilution with 20% HSA and 10% HES following CHT in rats did not decrease brain edema but did decrease brain tissue necrosis volume and NSS (improved neurological function), suggesting that the beneficial effect of hemodilution resulted not from decreased edema formation but rather from effects not measured in this study such as improved perfusion of the salvageable brain tissue surrounding the core injury. PMID- 10527148 TI - Recovery from anesthesia and postoperative extubation of neurosurgical patients: a review. AB - The most feared complications after intracranial surgery are development of an intracranial hematoma and major cerebral edema. Both may result in cerebral hypoperfusion and brain injury. Arterial hypertension via catecholamine release or sympathetic stimulation and hypercapnia may be predisposing factors. Other systemic secondary insults to the brain such as hypoxia and hypotension may exacerbate neuronal injury in hypoperfused areas of the brain. Thus, the anesthetic emergence of a neurosurgical patient should include maintenance of stable respiratory and cardiovascular parameters. Minimal reaction to the endotracheal tube prevents sympathetic stimulation and increases in venous pressure. On one hand, a delayed emergence and later extubation in the intensive care unit (ICU) might be recommended to achieve better thermal and cardiovascular stability after major intracranial procedures. On the other hand, the timely diagnosis of neurosurgical complications is required to limit brain damage; the diagnosis of complications relies on rapid neurological examination after early awakening. After uncomplicated surgery, normothermic and normovolemic patients generally recover from anesthesia with minimal metabolic and hemodynamic changes. Thus, early recovery and extubation in the operating room is the preferred method when the preoperative state of consciousness is relatively normal and surgery does not involve critical brain areas or extensive manipulation. In the complicated or unstable patient, the risks of early extubation may outweigh the benefits. It is, however, often possible to perform a brief awakening of the patient without extubation to allow early neurological evaluation, followed by delayed emergence and extubation. Close hemodynamic and respiratory monitoring are mandatory in all cases. The availability of ultrashort intravenous anesthetic agents and adrenergic blocking agents has added to the flexibility in the immediate emergence period after intracranial surgery. PMID- 10527149 TI - Radiation safety audits. AB - Radiation Safety Management Audits are an important tool for maintaining a good radiation safety program. They serve as a useful mechanism for program evaluation and improvement and can be instrumental in correcting problems, communicating strengths and weaknesses to manangement and involving the Radiation Safety Committee in the program operation. Effective audits can and should be done regardless of the size of the program. They need not be costly and, if organized properly, can be performed with a minimal time commitment. The remainder of this article will cover some successful audit tips. PMID- 10527150 TI - Radioisotope laboratory audits. AB - Elsewhere in this issue, Robin Elliott outlined a successful program for performing annual radiation safety management program audits. The results of such audits provide a useful means of evaluating and correcting deficiencies or weaknesses in a radiation safety program. At our institution we also perform an annual audit of each radioisotope lab oratory for the same reasons. PMID- 10527151 TI - The first year as a neophyte RSO at a liberal arts college. AB - Union College is a small liberal arts college in the northeast with a good reputation in science. It is also one of the oldest schools in the nation, having been founded in 1795. Langmuir and Steinmetz taught here as did Chandler, the founder of the American Chemical Society and Ellery, one of the longest tenured Secretaries of Sigma Xi. Arthur and Carter became U.S. Presidents. We claim a Nobel Prize winner, the inventor of transistors, the person who developed the methods for producing quartz crystals and fiber optics. The presidents of du Pont and Kodak attended Union College. We are referred to as the "Mother of Fraternities," and we had the first radio station on a college campus (although not through the airways). Still, until recently, our reputation has been less than admirable among radiation safety experts. This article highlights some of the experiences of a new RSO at this well-acknowledged institution. PMID- 10527152 TI - Detection of radon decay products in rainwater. AB - The Argonne National Laboratory-East (ANL-E) Environmental Radiation Monitoring System measures and records ambient radiation levels and provides detection capability for radon decay products in rain clouds. These decay products in rainwater tracked into a facility on the shoes of workers can cause false alarms from hand and shoe monitors. The monitors at ANL-E can easily detect the radon decay products, and the 19.6 and 26.8 min half-lives of the beta-particle emitters are long enough in many cases for sufficient activity to still be present to initiate a contamination alarm when the shoes are checked for radioactivity. The Environmental Radiation Monitoring System provides a warning when precipitation contains elevated levels of radon decay products. It is based on a prototype developed at the Super Collider Laboratory. During its first year of operation there were nine alarms from radon decay products with an alarm trigger point set at 30% greater than background. The alarms occurred at both monitoring stations, which are approximately 1,000 m apart, indicating large diameter radon clouds. The increases in background were associated with low atmospheric pressure. There was no correlation with radon released from the coal burning steam plant on the site. Alarms also occurred when short-lived accelerator-produced radioactivity in the exhaust stack plume passed over the NaI(Tl) detector in one of the stations. The 450 MeV proton accelerator near the station produced 12C, 13N, and 15O by spallation of air nuclei. The gamma-ray spectrum from the plume from the accelerator exhaust stack was dominated by the 511 keV annihilation gamma rays from decay of these radionuclides. These gamma rays were easily distinguished from the 609 keV, 1,120 keV, and 1,764 keV gamma rays emitted by the radon decay products. PMID- 10527153 TI - Lessons learned in decommissioning medical facilities. AB - In decommissioning medical research buildings at this institution, most areas surveyed were not contaminated above NRC 1993 release limits. Most contamination found was fixed. Wipe tests were inefficient at assessing removable contamination. An initial approach to surveying such a facility should begin with limited sampling emphasizing fixed contamination detection. Recent NRC guidance on decommissioning suggests that the requirements are becoming less stringent for medical facilities. This seems to support our limited decommissioning survey strategy for medical research buildings where radioisotopes have been used. PMID- 10527154 TI - Basic radiation protection considerations in dental practice. AB - This article provides basic guidance to radiation safety professionals unfamiliar with dental radiography on how to optimize the use of x-ray equipment and maintain occupational and non-occupational doses ALARA. Topics discussed in this article include basic protective measures commonly used to minimize the patient and operator's exposure to radiation, recommendations for the development of operating procedures, and the performance of compliance audits in dental practice. PMID- 10527155 TI - Reflections on the benefits of participating in ACURI. AB - There are many benefits to participating in the activities of a low-level radioactive waste generators group. Among them are the sharing of ideas, experience, and successes; ready access to information on all aspects of radioactive waste disposal; and the ability to have input into the regulatory and decision making processes. PMID- 10527156 TI - Protocols for implementing DOE authorized release of radioactive scrap metals. AB - A process to implement the U.S. Department of Energy's (DOE) policy for authorized release of radioactive materials from DOE facilities is provided in the Draft Handbook for Controlling Release for Reuse or Recycle of Property Containing Residual Radioactive Material, published by DOE in 1997 and distributed to DOE field offices for interim use and implementation. The authorized release of such property is intended to permit its beneficial use across the entire DOE complex. A computerized management tool--P2Pro(RSM)--has been developed to aid in carrying out the release process for radioactive metals. It contains protocols for the authorized release process and relevant information to facilitate the evaluation of scrap metals for reuse and recycle. The P2Pro(RSM) protocols provide DOE and its contractors with an effective, user friendly tool for managing authorized release activities P2Pro(RSM) is designed to be used in the Windows environment. The protocols incorporate a relational database coupled with a graphic-user interface to guide the user through the appropriate steps so authorized release limits can be developed. With the information provided in the database, an as-low-as-reasonably-achievable (ALARA) optimization process can be easily set up and run for up to 10 alternatives for disposition of radioactive scrap metals. The results of the ALARA optimization process can be printed in a series of reports and submitted as part of the application for the authorized release of the radioactive scrap metals. PMID- 10527157 TI - The LANL model 8823 whole-body TLD and associated dose algorithm. AB - The Los Alamos National Laboratory Model 8823 whole-body TLD has been designed to perform accurate dose estimates for beta, photon, and neutron radiations that are encountered in pure calibration, mixed calibration, and typical field radiation conditions. The radiation energies and field types for which the Model 8823 dosimeter is capable of measuring are described. The Model 8823 dosimeter has been accredited for all performance testing categories in the Department of Energy Laboratory Accreditation Program for external dosimetry systems. The philosophy used in the design of the Model 8823 dosimeter and the associated dose algorithm is to isolate the responses due to beta, photon, and neutron radiations; obtain radiation quality information; and make functional adjustments to the elemental readings to estimate the dose equivalent at 7, 300, and 1,000 mgcm(-2), representing the required repoting quantities for shallow, lens-of-the eye, and deep dose, respectively. PMID- 10527158 TI - The AL-R8 SI: the next generation staging container for plutonium pits at the USDOE Pantex Plant. AB - The AL-R8 SI (sealed insert) is the next generation staging container for plutonium pits at the U.S. DOE Pantex Plant. The sealed insert is a stainless steel container that will be placed inside a modified AL-R8 container to stagepits. A pit is a hollow sphere of plutonium metal which is the primary fissionable material in nuclear weapons (warheads and bombs). It is hermetically sealed by a cladding material, which is usually stainless steel. Personnel exposures to ionizing radiation from the pits in storage are expected to decrease due to the attenuation provided by the new SI. All personnel exposures to ionizing radiation at Pantex Plant are As Low As Reasonably Achievable (ALARA). Pantex Plant secures the common defense and national security of the United States by safely staging plutonium pits in a manner that protects the health and safety of employees, the public, and the environment. PMID- 10527159 TI - Extension of Fong and Alvarez: when is a lower limit of detection low enough? AB - The work of Fong and Alvarez is easily extended for counting when paired counting is not employed. It remains true in the general case that the precision of a counting method can be no less than about 30% at the LLD and so it is desirable to have decision levels at least several times larger than the LLD so that measurements have sufficient precision to make valid decisions. PMID- 10527160 TI - Confidence intervals for low-level, paired counting. AB - Neyman-Pearson principles are briefly discussed and 95% confidence intervals of the form [0, ##.##] are presented. Use is made of the fact that the probability of the difference of two random variables, each with a Poisson distribution, can be expressed in terms of modified Bessel functions of integral order and elementary functions. The validity of the values is discussed. PMID- 10527161 TI - Improved derivation of results of D. E. Allen: two-count method for stripping short-lived activity out of an air sample. AB - An improved derivation of the critical levels and detection limits for a two count method that attempts to measure long-lived activity in the presence of short-lived, naturally occurring progeny of radon is presented. A limitation of the two-count method is also mentioned. PMID- 10527162 TI - Science-based views of drug addiction and its treatment. PMID- 10527163 TI - Hopes for HIV eradication dim as stopping HAART allows resurgence. PMID- 10527164 TI - Like trying to get blood from a . . . federal agency. PMID- 10527165 TI - Illicit drug users not idle; report says 70% go to work. PMID- 10527166 TI - From the Centers for Disease Control and Prevention. Epidemiology of measles- United States, 1998. PMID- 10527167 TI - From the Centers for Disease Control and Prevention. Menigococcal disease--New England, 1993-1998. PMID- 10527168 TI - From the Centers for Disease Control and Prevention. Carbon monoxide poisoning deaths associated with camping--Georgia, March 1999. PMID- 10527169 TI - Parkinson disease in twins. PMID- 10527170 TI - Parkinson disease in twins. PMID- 10527171 TI - Moxibustion for breech presentation. PMID- 10527172 TI - Moxibustion for breech presentation. PMID- 10527173 TI - Health care service quality. PMID- 10527174 TI - Health care service quality. PMID- 10527175 TI - Medical futility in end-of-life care. PMID- 10527176 TI - A new potential hazard of ear piercing? PMID- 10527177 TI - Some implications of the prion paradigm: caveat denaturor. PMID- 10527178 TI - Prenatal and postnatal methylmercury exposure and neurodevelopmental outcomes. PMID- 10527179 TI - Modified directly observed therapy for treatment of human immunodeficiency virus. PMID- 10527180 TI - Factors correlated with progression-free survival after high-dose chemotherapy and hematopoietic stem cell transplantation for metastatic breast cancer. AB - CONTEXT: Women with breast cancer are the most frequent recipients of high-dose chemotherapy followed by autologous hematopoietic stem cell transplantation (autotransplants) in North America. Despite widespread use, controversy exists about the benefits of and appropriate patients for this therapy. OBJECTIVE: To determine factors associated with disease progression or death after autotransplantation in women with metastatic breast cancer. DESIGN: Analysis of data collected retrospectively (January 1989 to 1992) and prospectively (1992 through January 1995) for the Autologous Blood and Marrow Transplant Registry. SETTING: Sixty-three hospitals in North America, Brazil, and Russia. PARTICIPANTS: A total of 1188 consecutive women aged 18 to 70 years receiving autotransplants for metastatic or locally recurrent breast cancer, with a median follow-up of 291/2 months. MAIN OUTCOME MEASURE: Time to treatment failure (disease progression, disease recurrence, or death) after autotransplantation. RESULTS: Factors associated with significantly (P<.05) increased risk of treatment failure in a Cox multivariate analysis included age older than 45 years (relative hazard, 1.17; 95% confidence interval [CI], 1.02-1.33), Karnofsky performance score less than 90% (1.27; 95% CI, 1.07-1.51), absence of hormone receptors (1.31; 95% CI, 1.15-1.51), prior use of adjuvant chemotherapy (1.31; 95% CI, 1.10-1.56), initial disease-free survival interval after adjuvant treatment of no more than 18 months (1.99; 95% CI, 1.62-2.43), metastases in the liver (1.47; 95% CI, 1.20-1.80) or central nervous system (1.56; 95% CI, 0.99 2.46 [approaches significance]) vs soft tissue, bone, or lung, 3 or more sites of metastatic disease (1.32; 95% CI, 1.13-1.54), and incomplete response vs complete response to standard-dose chemotherapy (1.65; 95% CI, 1.36-1.99). Receiving tamoxifen posttransplantation was associated with a reduced risk of treatment failure in women with hormone receptor-positive tumors (relative hazard, 0.60; 95% CI, 0.47-0.87). Women with no risk factors (n = 38) had a 3-year probability of progression-free survival of 43% (95% CI, 27%-61 %) vs 4% (95% CI, 2%-8%) for women with more than 3 risk factors (n = 343). CONCLUSION: These data indicate that some women are unlikely to benefit from autotransplantation and should receive this treatment only after being provided with prognostic information and in the context of clinical trials attempting to improve outcome. PMID- 10527181 TI - Effects of risedronate treatment on vertebral and nonvertebral fractures in women with postmenopausal osteoporosis: a randomized controlled trial. Vertebral Efficacy With Risedronate Therapy (VERT) Study Group. AB - CONTEXT: Risedronate, a potent bisphosphonate, has been shown to be effective in the treatment of Paget disease of bone and other metabolic bone diseases but, to our knowledge, it has not been evaluated in the treatment of established postmenopausal osteoporosis. OBJECTIVE: To test the efficacy and safety of daily treatment with risedronate to reduce the risk of vertebral and other fractures in postmenopausal women with established osteoporosis. DESIGN, SETTING, AND PARTICIPANTS: Randomized, double-blind, placebo-controlled trial of 2458 ambulatory postmenopausal women younger than 85 years with at least 1 vertebral fracture at baseline who were enrolled at 1 of 110 centers in North America conducted between December 1993 and January 1998. INTERVENTIONS: Subjects were randomly assigned to receive oral treatment for 3 years with risedronate (2.5 or 5 mg/d) or placebo. All subjects received calcium, 1000 mg/d. Vitamin D (cholecalciferol, up to 500 IU/d) was provided if baseline levels of 25 hydroxyvitamin D were low. MAIN OUTCOME MEASURES: Incidence of new vertebral fractures as detected by quantitative and semiquantitative assessments of radiographs; incidence of radiographically confirmed nonvertebral fractures and change from baseline in bone mineral density as determined by dual x-ray absorptiometry. RESULTS: The 2.5 mg/d of risedronate arm was discontinued after 1 year; in the placebo and 5 mg/d of risedronate arms, 450 and 489 subjects, respectively, completed all 3 years of the trial. Treatment with 5 mg/d of risedronate, compared with placebo, decreased the cumulative incidence of new vertebral fractures by 41 % (95% confidence interval [CI], 18%-58%) over 3 years (11.3 % vs 16.3%; P= .003). A fracture reduction of 65% (95% CI, 38%-81 %) was observed after the first year (2.4% vs 6.4%; P<.001). The cumulative incidence of nonvertebral fractures over 3 years was reduced by 39% (95% CI, 6%-61 %) (5.2 % vs 8.4%; P = .02). Bone mineral density increased significantly compared with placebo at the lumbar spine (5.4% vs 1.1 %), femoral neck (1.6% vs -1.2%), femoral trochanter (3.3% vs -0.7%), and midshaft of the radius (0.2% vs -1.4%). Bone formed during risedronate treatment was histologically normal. The overall safety profile of risedronate, including gastrointestinal safety, was similar to that of placebo. CONCLUSIONS: These data suggest that risedronate therapy is effective and well tolerated in the treatment of women with established postmenopausal osteoporosis. PMID- 10527182 TI - Prevalence of attempting weight loss and strategies for controlling weight. AB - CONTEXT: Overweight and obesity are increasing in the United States. Changes in diet and physical activity are important for weight control. OBJECTIVES: To examine the prevalence of attempting to lose or to maintain weight and to describe weight control strategies among US adults. DESIGN: The Behavioral Risk Factor Surveillance System, a random-digit telephone survey conducted in 1996 by state health departments. Setting The 49 states (and the District of Columbia) that participated in the survey. PARTICIPANTS: Adults aged 18 years and older (N = 107 804). MAIN OUTCOME MEASURES: Reported current weights and goal weights, prevalence of weight loss or maintenance attempts, and strategies used to control weight (eating fewer calories, eating less fat, or using physical activity) by population subgroup. RESULTS: The prevalence of attempting to lose and maintain weight was 28.8% and 35.1 % among men and 43.6% and 34.4% among women, respectively. Among those attempting to lose weight, a common strategy was to consume less fat but not fewer calories (34.9% of men and 40.0% of women); only 21.5% of men and 19.4% of women reported using the recommended combination of eating fewer calories and engaging in at least 150 minutes of leisure-time physical activity per week. Among men trying to lose weight, the median weight was 90.4 kg with a goal weight of 81.4 kg. Among women, the median weight was 70.3 kg with a goal weight of 59.0 kg. CONCLUSIONS: Weight loss and weight maintenance are common concerns for US men and women. Most persons trying to lose weight are not using the recommended combination of reducing calorie intake and engaging in leisure-time physical activity 150 minutes or more per week. PMID- 10527183 TI - Unintended pregnancy among adult women exposed to abuse or household dysfunction during their childhood. AB - CONTEXT: Studies have identified childhood sexual and physical abuse as a risk factor for adolescent pregnancy but the relationship between exposure to childhood abuse and unintended pregnancy in adulthood has, to our knowledge, not been studied. OBJECTIVE: To assess whether unintended pregnancy during adulthood is associated with exposure to psychological, physical, or sexual abuse or household dysfunction during childhood. DESIGN AND SETTING: Analysis of data from the Adverse Childhood Experiences Study, a survey mailed to members of a large health maintenance organization who visited a clinic in San Diego, Calif, between August and November 1995 and January and March 1996. The survey had a 63.4% response rate among the target population for this study. PARTICIPANTS: A total of 1193 women aged 20 to 50 years whose first pregnancy occurred at or after age 20 years. MAIN OUTCOME MEASURE: Risk of unintended first pregnancy by type of abuse (psychological, physical, or sexual abuse; peer sexual assault) and type of household dysfunction (physical abuse of mother by her partner, substance abuse by a household member, mental illness of a household member). RESULTS: More than 45% of the women reported that their first pregnancy was unintended, and 65.8% reported exposure to 2 or more types of childhood abuse or household dysfunction. After adjustment for confounders (marital status at first pregnancy and age at first pregnancy), the strongest associations between childhood experiences and unintended first pregnancy included frequent psychological abuse (risk ratio [RR], 1.4; 95% confidence interval [CI], 1.2-1.6), frequent physical abuse of the mother by her partner (RR, 1.4; 95% CI, 1.1-1.7), and frequent physical abuse (RR, 1.5; 95% CI, 1.2-1.8). Women who experienced 4 or more types of abuse during their childhood were 1.5 times (95% CI, 1.2-1.8) more likely to have an unintended first pregnancy during adulthood than women who did not experience any abuse. CONCLUSIONS: This study indicates that there may be a dose-response association between exposure to childhood abuse or household dysfunction and unintended first pregnancy in adulthood. Additional research is needed to fully understand the causal pathway of this association. PMID- 10527184 TI - Tube feeding in patients with advanced dementia: a review of the evidence. AB - Patients with advanced dementia frequently develop eating difficulties and weight loss. Enteral feeding tubes are often used in this situation, yet benefits and risks of this therapy are unclear. We searched MEDLINE, 1966 through March 1999, to identify data about whether tube feeding in patients with advanced dementia can prevent aspiration pneumonia, prolong survival, reduce the risk of pressure sores or infections, improve function, or provide palliation. We found no published randomized trials that compare tube feeding with oral feeding. We found no data to suggest that tube feeding improves any of these clinically important outcomes and some data to suggest that it does not. Further, risks are substantial. The widespread practice of tube feeding should be carefully reconsidered, and we believe that for severely demented patients the practice should be discouraged on clinical grounds. PMID- 10527185 TI - Users' Guides to the Medical Literature: XIX. Applying clinical trial results B. Guidelines for determining whether a drug is exerting (more than) a class effect. PMID- 10527186 TI - High-dose chemotherapy and breast cancer. PMID- 10527187 TI - Lack of evidence about tube feeding--food for thought. PMID- 10527188 TI - JAMA Patient Page: osteoporosis. PMID- 10527189 TI - Uterine fibroid embolization: another paradigm shift for interventional radiology? PMID- 10527190 TI - Initial results from uterine fibroid embolization for symptomatic leiomyomata. AB - PURPOSE: To evaluate the safety and short-term efficacy of uterine fibroid embolization (UFE) in patients with symptomatic uterine fibroids. MATERIALS AND METHODS: Bilateral UFE was performed in 61 patients with symptomatic uterine leiomyomata during a 16-month period. Imaging was performed before the procedure and at 3 months and 1 year after the procedure. Questionnaires were obtained at regular intervals after the procedure to assess patient outcome. RESULTS: All procedures but one were technically successful. Mean clinical follow-up was 8.7 months. Minor complications occurred in five patients during the follow-up period. All were treated without permanent sequelae. Menstrual bleeding was improved in 89%, with 81% of patients moderately to markedly improved. Pelvic pain and pressure was improved in 96% of patients, with moderate to marked improvement in 79%. At initial imaging follow-up (mean, 4.4 months postprocedure), median uterine volume decreased 34% (P = .0001) and the median dominant fibroid volume decreased 50% (P = .0001). Imaging at 1 year (mean, 12.3 months) after the procedure showed continued reduction with a median uterine volume reduction of 48% (P = .0002) and median dominant fibroid volume decrease of 78% (P = .0002). CONCLUSION: In the authors' initial clinical experience, UFE appears effective in controlling symptoms and substantially reducing fibroid volume with few complications. PMID- 10527191 TI - Uterine artery embolization for the treatment of uterine leiomyomata midterm results. AB - INTRODUCTION: The authors review their midterm experience with uterine artery embolization for the treatment of uterine fibroids. MATERIALS AND METHODS: Sixty patients were referred for permanent polyvinyl alcohol (PVA) foam particle uterine artery embolization during an 18-month period. Detailed clinical follow up and ultrasound follow-up were obtained. RESULTS: Bleeding was a presenting symptom in 56 patients and pain was a presenting symptom in 47 patients. All patients underwent a technically successful embolization. One of the patients underwent unilateral embolization. Fifty-nine patients underwent bilateral embolization. Of all patients undergoing bilateral embolization, at last follow up (mean, 16.3 months), 81% had their uterus and had moderate or better improvement in their symptoms. Ninety-two percent of these patients also had reductions in uterine and dominant fibroid volumes. Overall, the mean uterine and dominant fibroid volume reduction were 42.8% and 48.8%, respectively (mean follow up, 10.2 months). One infectious complication that necessitated hysterectomy occurred. CONCLUSION: Uterine artery embolization for the treatment of uterine fibroids is a minimally invasive technique with low complication rates and very good clinical efficacy. PMID- 10527192 TI - Ovarian artery supply of uterine fibroids as a cause of treatment failure after uterine artery embolization: a case report. PMID- 10527193 TI - A clinical failure of uterine fibroid embolization due to adenomyosis. PMID- 10527194 TI - Embolotherapy of persistent endoleaks after endovascular repair of abdominal aortic aneurysm with the ancure-endovascular technologies endograft system. AB - PURPOSE: Endoleak is a potential complication after endovascular repair of abdominal aortic aneurysm (AAA). It may result in continued growth of the aneurysm and potentially result in aneurysm rupture. The authors present their experience with embolotherapy in patients with persistent perigraft flow treated with the Ancure-Endovascular Technologies endograft system. MATERIALS AND METHODS: Between February 1996 and August 1998, 54 patients underwent successful repair of AAA with use of the Ancure system. All underwent operative angiography and discharge computed tomography (CT). Follow-up included CT at 6, 12, and 24 months, and CT was also performed at 3 months if an endoleak was present on the discharge CT. Persistent endoleak was defined as perigraft flow still present on the 6-month CT. Seven of 21 initial endoleaks persisted at 6 months. Six patients returned for embolization of the perigraft space and outflow vessels including lumbar arteries and the inferior mesenteric artery (IMA). RESULTS: Five of the six patients had leaks from the proximal (n = 1) or distal attachment sites (n = 4) of the Ancure system with outflow into lumbar arteries and/or the IMA; one leak was caused by retrograde IMA flow. The six patients underwent nine embolization procedures with only one minor complication. Follow-up CT showed complete resolution of endoleak and decrease in size of the aneurysm sac in all patients. CONCLUSIONS: Although endoleak is commonly seen initially with the Ancure system, persistent leak occurred in 13% of the patients in the study. Persistent flow in most patients arises from a graft attachment site combined with patent outflow vessels such as the IMA or lumbar arteries. Persistent endoleaks can be effectively and safely embolized with use of a combination of coil embolization of the perigraft space and embolization of outflow vessels. Such intervention resulted in a decrease in size of the aneurysm sac. PMID- 10527195 TI - Accuracy and safety of carbon dioxide inferior vena cavography. AB - PURPOSE: The purpose of this study was to assess the accuracy of carbon dioxide compared to iodinated contrast material for determining inferior vena cava (IVC) diameter prior to filter placement, and to assess the safety of CO2 when used for this purpose. PATIENTS AND METHODS: Consecutive patients undergoing inferior vena cavography prior to filter placement were prospectively evaluated with use of both CO2 and iodinated contrast material. The diameter of the IVC was measured and compared in the same four locations in each patient for both agents. The diameter was corrected for magnification and pin-cushion distortion. The ability of CO2 to correctly classify IVC diameter as < or =28 mm or >28 mm, based on the IVC diameter with iodinated contrast material, was determined. A consensus panel assessed renal vein visualization with CO2 and iodinated contrast material. Blood pressure and arterial oxygen saturation were measured immediately before and after CO2 injection. RESULTS: Among 30 patients, there was no significant difference in the measured diameter of the IVC with CO2 versus iodinated contrast material after correction for magnification and pin-cushion distortion. One of 30 patients (3.3%) in this study was misclassified as having an IVC < or =28 mm with CO2 when, in fact, the IVC diameter was >28 mm based on iodinated contrast material. This could be clinically significant for certain IVC filters. Forty seven percent of renal veins identified on contrast venography were identified by CO2 vena cavography. There was no significant difference in the blood pressure or oxygen saturation values measured before and after CO2 injection. However, one patient with pulmonary artery hypertension did experience transient, symptomatic hypotension after CO2 injection. CONCLUSIONS: In most patients, CO2 vena cavography accurately evaluated IVC diameter prior to filter placement. In 3.3% of patients, the discrepancy in measurements between CO2 and iodinated contrast material could be clinically significant, depending on the type of filter placed. CO2 was less accurate than iodinated contrast material in identifying renal veins. Although CO2 vena cavography is safe in the majority of patients, it should be used with caution in patients with pulmonary hypertension. PMID- 10527196 TI - Skin radionecrosis after percutaneous transluminal coronary angioplasty: dosimetric and biological assessment. PMID- 10527197 TI - Comparison of the angiojet rheolytic catheter to surgical thrombectomy for the treatment of thrombosed hemodialysis grafts. Peripheral AngioJet Clinical Trial. AB - PURPOSE: To compare the clinical effectiveness of the AngioJet F105 rheolytic catheter to that of surgical thrombectomy for the treatment of thrombosed hemodialysis grafts. MATERIALS AND METHODS: This was a multicenter, prospective, randomized trial comparing technical success, primary patency, and complication rates. A total of 153 patients were enrolled: 82 patients in the AngioJet group and 71 patients in the surgical thrombectomy group. Patient follow-up was performed 24-48 hours, 1 month, and 6 months after the procedures. RESULTS: Technical success, as defined by the patient's ability to undergo hemodialysis treatment, was 73.2% for the AngioJet group and 78.8% for the surgical thrombectomy group (P = .41). The primary patency rates of the AngioJet group were 32%, 21%, and 15% at 1, 2, and 3 months, respectively. The primary patency rates for the surgical group were 41%, 32%, and 26% at 1, 2, and 3 months, respectively. This difference approached statistical significance (P = .053). The groups had similar complication rates-14.6% in the AngioJet group and 14.1% in the surgery group-although the surgery group had more major complications (11.3%). In the AngioJet group, there was a transient increase in plasma-free hemoglobin, which normalized within 24-48 hours. CONCLUSIONS: The AngioJet F105 catheter provides similar clinical results when compared to surgical thrombectomy for the treatment of thrombosed hemodialysis grafts. The difference in patency rates between these two techniques approached statistical significance. In addition, results of both thrombectomy methods were inferior to those suggested by the Dialysis Outcomes Quality Initiative guidelines. PMID- 10527199 TI - Placement of a flexible endovascular stent across the femoral joint: an in vivo study in the swine model. AB - PURPOSE: To investigate the effects of joint motion on the structural integrity of periarticular stents and on the development of neointimal hyperplasia within these devices. MATERIALS AND METHODS: In four juvenile farm swine, Wall-stents were implanted in the common femoral arteries and contralateral common femoral veins, centered at the point of maximal conformational change during passive hip flexion. Control stents were placed in the aortae and iliac veins. Angiography and transcatheter blood pressure measurements were obtained across each stent, with periarticular stents studied in flexion and extension. Two animals underwent repeated evaluation after 1 month, the others after 3 months. Findings were correlated with gross and histopathologic findings in the harvested stents. RESULTS: No stent fractures occurred. One femoral vein was injured during stent placement and was occluded 1 month later at follow-up. Hemodynamically significant stenoses were identified in one arterial stent and one venous stent at 3 months. The amount of neointimal hyperplasia was greater in periarticular stents than in controls and greater in animals studied at 1 month than in those studied at 3 months. The pattern of neointimal hyperplasia within mobile arteries was circumferentially asymmetric and thicker at the distal ends of the stents. Venous neointimal hyperplasia was thicker and markedly different in character than that seen in arterial stents from the same animals. CONCLUSIONS: Periarticular Wallstents and the underlying vascular anatomy remained structurally intact despite the stresses of repetitive motion during a 3-month period. Stents deployed across joints or in venous locations may be at greater risk for neointimal hyperplasia development and eventual occlusion than those deployed in immobile vessels and arteries. Neointimal hyperplasia may decrease in thickness after an initial period of exuberant development. Additional studies are necessary to determine long-term outcomes. PMID- 10527198 TI - Experimental evaluation of cellulose acetate NF and ethylene-vinyl alcohol copolymer for selective arterial embolization. AB - PURPOSE: Studies were conducted in rabbits to evaluate two new liquid polymeric compounds developed for selective arterial embolization. MATERIALS AND METHODS: The compounds consisted of cellulose acetate NF (Embolyx C) or ethylene-vinyl alcohol copolymer (Embolyx E) dissolved in anhydrous dimethyl sulfoxide (DMSO) containing 30% tantalum powder. Acute renal embolization was performed to determine an optimal method of administration and level of embolization. Kidneys were embolized with and without flow around the catheter. DMSO was also injected in the same manner. Tissue sections were examined radiographically and microscopically. Tumor embolization was performed to evaluate the efficacy of the polymers and compare their embolic effects with polyvinyl alcohol (PVA) particles and gelatin sponge (Gelfoam) powder. An embolic agent, saline, or DMSO was injected into the deep femoral artery feeding an intramuscular VX2 carcinoma. Animals were followed up for 3 weeks. RESULTS: Viscosity and administration technique affected polymer distribution and depth of penetration. Embolization with the test polymers was quicker and more easily achieved than with PVA or Gelfoam, and no recanalization occurred. Both polymers were as effective as PVA particles for tumor ablation, but DMSO caused some vascular damage. CONCLUSION: Although use of DMSO has some drawbacks, the results of this study warrant further investigation of the Embolyx polymers for tumor embolization. PMID- 10527200 TI - Percutaneous transjugular kidney biopsy in swine with use of a side-cutting needle with a blunt-tipped stylet. AB - PURPOSE: To evaluate a new 19-gauge blunt-tipped, side-cutting, single throw transjugular biopsy needle for transvenous kidney biopsies. MATERIALS AND METHODS: Transjugular transvenous kidney biopsies were performed with a modified 70-cm biopsy needle utilizing fluoroscopic guidance in nine swine. Three tissue specimens were obtained with four biopsy device passes in five animals and three biopsy device passes in four animals. Renal arteriography and venography were performed immediately before and after renal biopsy. Five animals were killed immediately after biopsy. Four animals were allowed to recover and underwent arteriography and venography prior to being killed, which varied from 1 to 6 weeks. Gross and histologic examinations of the biopsied kidney were performed after euthanasia. A pathologist reviewed all biopsy specimens for quality based on the number of glomeruli present. RESULTS: Results of immediate and delayed arteriography and venography were normal in all cases. Histologic evaluation of all biopsy specimens demonstrated a range of two to 13 glomeruli per sample (mean, 6.5), with successful acquisition of the cortex. In one animal killed immediately after biopsy, a small subcapsular hematoma was present. CONCLUSION: The 19-gauge, side-cut biopsy needle with a blunt-tip stylet proved to be efficacious for obtaining renal cortical samples in right swine kidneys via a transjugular approach. PMID- 10527201 TI - Transcaval TIPS: indications and anatomic considerations. PMID- 10527202 TI - Embolization of colonic hemorrhage with use of a flow-assisted catheter. PMID- 10527203 TI - Retrieval of a fractured Guglielmi detachable coil with use of the Goose Neck snare "twist" technique. PMID- 10527204 TI - Management of chylothorax by percutaneous catheterization and embolization of the thoracic duct: prospective trial. AB - PURPOSE: To prospectively assess the efficacy of percutaneous transabdominal thoracic duct catheterization and embolization in the management of patients with high-output chylothoracic effusions. MATERIALS AND METHODS: Eleven consecutive patients (four women and seven men; mean age, 53 years) were referred with chylothorax secondary to esophagectomy (n = 4), lobectomy (n = 1), lung transplant (n = 1), coronary artery bypass (n = 1), aortic graft (n = 2), lymphangioleiomyomatosis (n = 1), and gunshot wound (n = 1). Two patients were brought by ambulance and referred back to their hospital on the same day. Pedal lymphography was used to opacify the cisterna chyli or major retroperitoneal lymphatic trunks. When patent, these were punctured under local anesthesia with a fine needle and the thoracic duct was catheterized over a microguide wire with use of a 3-F catheter; the duct was embolized with platinum coils. Patients were followed up for decrease in thoracic drainage output and morbidity. RESULTS: There were no retroperitoneal ducts suitable for catheterization in six patients because of previous abdominal surgery, trauma, or lymphangioleiomyomatosis; the thoracic duct was successfully catheterized in five patients, a 45% technical success rate. Thoracic duct embolization was performed in four patients, with cure of effusion in two. In the other two patients, one with lymphangioleiomyomatosis and the other with nonchylous pleural fluid, continued effusion was successfully treated by means of pleurodesis. Of two patients with previous thoracic duct ligation, one was found to have the duct incompletely tied. The authors were surprised to find that previous major abdominal surgery, chronic aortic dissection, and lymphangioleiomyomatosis could obliterate major retroperitoneal lymphatic ducts and the cisterna chyli. Percutaneous study of the thoracic duct with aqueous contrast medium was more sensitive than lymphography with iodinated oil. There was no morbidity. CONCLUSIONS: Catheterization of the thoracic duct was possible in all patients who had patent major retroperitoneal lymphatic trunks. Thoracic duct embolization was curative in patients with demonstrable duct leakage. Previous abdominal surgery, aortic dissection, and lymphangioleiomyomatosis can lead to silent occlusion of retroperitoneal lymphatic trunks. Percutaneous thoracic duct catheterization and embolization is safe and can replace surgical ligation in some patients. PMID- 10527205 TI - Porous and nonporous polycarbonate urethane stent-grafts for TIPS formation: biologic responses. AB - PURPOSE: To evaluate the biologic response to transjugular intrahepatic portosystemic shunts (TIPS) lined with polycarbonate urethane endografts and the effects of different porosity formulations. MATERIALS AND METHODS: Seventeen TIPS were created in non-modified portal hypertensive miniswine with use of porous (n = 6), nonporous (n = 7) polycarbonate urethane stent-grafts, and control Wallstents TIPS (n = 4). Eight-week venography, histology, scanning electron microscopy, and immunohistochemical analyses were performed. RESULTS: The mean 8 week percent parenchymal tract shunt stenosis was 75%, 46%, and 26% in the control, porous, and nonporous groups, respectively. Occlusions developed in one control, one porous, and two nonporous shunts. The biologic response to porous grafts included marked inflammation and encapsulation and permeation of the grafts by a thick fibrous pseudointima. Nonporous grafts evoked little inflammation or pseudointima. Mature thrombus lined the occluded shunts (under which little luminal pseudointima or endothelium was present). The control group showed typical pseudointimal hyperplasia enveloping the intraparenchymal portions of the stents. CONCLUSIONS: The healing response of the porous and nonporous grafts markedly differed. Unlike the porous grafts and control stents, the nonporous endografts elicited little inflammation or luminal pseudointimal hyperplasia, although sporadic thrombosis was problematic in this normotensive model. Graft use in high-flow situations (ie, human TIPS, possibly in concert with antiplatelet agents) may allow desired shunt patency prolongation. PMID- 10527206 TI - Comparison of hepatic damage from direct injections of iodinated contrast agents and carbon dioxide. AB - PURPOSE: This study guides the choice of contrast agent for localization of portal veins during transjugular intrahepatic portosystemic shunt (TIPS) placement or use in percutaneous transhepatic cholangiography (PTC) by providing gross anatomic and histologic comparison of effects from parenchymal injections of iodinated contrast agents and carbon dioxide. MATERIALS AND METHODS: Eighteen New Zealand White rabbits received direct injections of 2-5 mL of either the nonionic contrast agent iohexol 300 mgI or the ionic contrast agent diatrizoate meglumine 60% into one lobe of the liver and the same volume of CO2 into the other lobe. The rabbits were killed at 2-7 days for gross and histologic evaluation of the livers. RESULTS: At the time of injection, the diatrizoate and iohexol sites showed persistent dark discoloration, whereas CO2 sites showed minimal visible changes. On gross examination at death, all diatrizoate sites showed severe scarring and also commonly showed areas of necrosis. CO2 and iohexol sites showed only minimal discoloration and needle-puncture scars (P < .0001). The histologic grade for diatrizoate sites was significantly more severe than paired CO2 sites (P < .016). Iohexol sites showed mild histologic changes similar to paired CO2 sites (P = .375). CONCLUSION: Iohexol and CO2 produce less severe hepatic damage and are preferred to meglumine diatrizoate for hepatic injection. PMID- 10527207 TI - Ultrasound-guided percutaneous coil embolization of incompetent perforating veins: not effective for treatment of venous ulcers and recurrent varicosities. AB - PURPOSE: To investigate the feasibility of ultrasound-guided percutaneous coil embolization of incompetent perforating veins as minimally invasive treatment for venous ulcers and recurrent varicosities in the lower leg. This could be an alternative to surgical ligation of perforating veins. MATERIALS AND METHODS: In 15 patients (six women, nine men; mean age, 50 years), 18 incompetent perforating veins in the lower leg were treated by ultrasound-guided percutaneous placement of embolization coils. RESULTS: Successful vein occlusion with one or more coils was achieved in 12 of the 18 veins (technical success rate, 67%). Clinical symptoms improved in only three of the 15 patients (clinical success rate, 20%). During follow-up (2-12 months), recanalization of coil embolized veins occurred in nine of the 12 initially occluded veins. CONCLUSION: Percutaneous ultrasound guided coil embolization does not appear to be as effective as subfascial endoscopic perforator surgery in the treatment of incompetent perforator veins. PMID- 10527208 TI - Effectiveness of preoperative transarterial chemoembolization in presumed inoperable hepatoblastoma. AB - PURPOSE: To evaluate the effectiveness and therapeutic role of preoperative transarterial chemoembolization (TACE) of hepatoblastoma. MATERIALS AND METHODS: Four patients (one boy, three girls) with unresectable hepatoblastoma were treated twice with preoperative TACE in an effort to improve the surgical and clinical outcome. The patients ranged in age from 8 to 27 months (mean, 15 months). The first TACE was performed superselectively in tumor feeding arteries. The second TACE was performed 3 weeks later. Surgical hepatic resection was performed 1 month after the second TACE. Contrast-enhanced computed tomography (CT) was used to evaluate changes in size, volume, internal texture, and margin of the masses. The toxicity of the chemotherapeutic drugs was evaluated by blood chemistry analysis (AST/ALT, alpha-FP) performed before and after TACE, and after surgery. RESULTS: TACE allowed subsequent surgical resection in all four patients, who remained disease free 16-52 months after operation. There were no major problems related to TACE. There was no chemotherapeutic agent toxicity from TACE. The average largest diameters and volumes of the tumors decreased by 31% (8.3 to 5.6 cm) and 69% (317 to 93 cm2), respectively. CONCLUSION: TACE provided subsequent successful surgical resection and good long-term results in all four patients. The hepatoblastomas were initially considered inoperable because of extensive hepatic involvement and indistinct margins. PMID- 10527209 TI - Re: long-term follow-up of upper extremity implanted venous access devices in oncology patients. PMID- 10527210 TI - Re: treatment of acute aortorenal bypass graft thrombosis by means of primary stent placement and adjunctive thrombolysis. PMID- 10527211 TI - Surface EMG of proximal leg muscles in neuromuscular patients and in healthy controls. Relations to force and fatigue. AB - In an effort to find parameters to evaluate patients with neuromuscular disorders, surface electromyography (SEMG) of proximal leg muscles was performed in 33 patients with myotonic dystrophy (MyD), 29 patients with Charcot-Marie Tooth (CMT) disease and 20 healthy controls. The root mean square (RMS) of the SEMG amplitude (microV) was calculated at different torque levels. Endurance (seconds) and median frequency (Fmed) of the SEMG power spectrum, used as parameters of fatigue, were determined at 80% of MVC. Maximum voluntary contraction (MVC) was found to be decreased in patients; the ratio between RMS values of antagonists and agonists was increased and torque-EMG ratios (Nm/microV) were decreased. These differences with respect to controls were more pronounced in MyD than in CMT. The initial Fmed value was lowest in CMT. The greatest decrease in Fmed was found in MyD. SEMG data in relation to force have not been determined before in groups of MyD or CMT patients. In both disorders, parameters differed from controls, which means that adding SEMG to strength measurements could be useful in studying the progress of the disorder and the effects of interventions. PMID- 10527212 TI - Estimating net lumbar sagittal plane moments from EMG data. The validity of calibration procedures. AB - In the present study the validity of EMG based methods to estimate the net moment working at the lumbar spine was investigated. Eight subjects performed a series of static and dynamic tasks. EMG was recorded from 8 locations over the back muscles. At the same time force platform and kinematic data for a linked segment analysis were collected. The net moment at the lumbar spine was calculated from the latter data and compared to EMG based estimates of the same moment. These estimates were derived from a linear regression between the EMG amplitudes and the net moments obtained during static ramp calibrations. It appeared that calibration in several postures, covering the range occurring in the tasks studied, and in a posture in the middle of this range, yielded estimates of the group averaged 10th, 50th, and 90th percentile of the net moments which were within 10% of the real value. The explained variance obtained in the calibration procedure proved not to be a good indicator of the validity of the procedure. PMID- 10527213 TI - Upper trapezius muscle activity patterns during repetitive manual material handling and work with with a computer mouse. AB - Firstly, upper trapezius EMG activity patterns were recorded on the dominant side of 6 industrial production workers and on the side operating a computer mouse of 14 computer-aided design (CAD) operators to study differences in acute muscular response related to the repetitiveness of the exposure. The work tasks were performed with median arm movement frequencies ranging from 5 min(-1) to 13 min( 1) and were characterized by work cycle times ranging from less than 30 sec to several days. However, the static and median EMG levels and EMG gap frequencies were similar for all work tasks indicating that shoulder muscle loads may be unaffected by large variations in arm movement frequencies and work cycle times. An exposure variation analyses (EVA) showed that the EMG activity patterns recorded during production work were more repetitive than during CAD work, whereas CAD work was associated with more static muscle activity patterns, both may be associated with a risk of developing musculoskeletal symptoms. Secondly, upper trapezius EMG activity patterns recorded on the mouse side of the CAD operators were compared with those recorded on the non-mouse side to study differences in muscular responses potentially related to the risk of developing shoulder symptoms which were more prevalent on the mouse side. The number of EMG gaps on the mouse side were significantly lower than the values for the upper trapezius on the non-mouse side indicating that more continuous activity was present in the upper trapezius muscle on the mouse side and EVA analyses showed a more repetitive muscle activity pattern on the mouse side. These findings may be of importance to explain differences in the prevalence of shoulder symptoms. PMID- 10527214 TI - Reduction in subluxation and improved muscle function of the hemiplegic shoulder joint after therapeutic electrical stimulation. AB - Seventeen hemiplegic patients with chronic shoulder subluxation secondary to a cerebrovascular accident (CVA) were divided into three groups, two of which were subjected to 6 weeks of therapeutic electrical stimulation (TES) for 15 minutes twice a day, in order to assess the effectiveness of the treatment in reducing subluxation, and in improving shoulder abduction function. The third group was used as a control (C group). After 6 weeks of electrical stimulation of the supraspinatus (S group) and deltoid (D group), a significant (p<0.05) reduction in subluxation was observed in both groups when compared to the C group. The maximal force of shoulder abduction showed a tendency to increase in the S group (p<0.10). A significant increase in maximal force was also observed in the D group. In most of the TES-treated muscles, the interference pattern of EMG at maximum voluntary contraction increased. The amplitude of the EMG activity of the stimulated muscle also increased. Thus, we concluded that electrical stimulation therapy of the supraspinatus and the deltoid muscle is an effective treatment modality for shoulder subluxation and shoulder abduction function in hemiplegic patients. PMID- 10527215 TI - Time-frequency methods applied to muscle fatigue assessment during dynamic contractions. AB - This paper discusses the assessment of the electrical manifestations of muscle fatigue during dynamic contractions. In the past, the study of muscle fatigue was restricted to isometric constant force contractions because, in this contraction paradigm, the myoelectric signal may be considered as wide sense stationary over epochs lasting up to two or three seconds, and hence classic spectral estimation techniques may be applied. Recently, the availability of spectral estimation techniques specifically designed for nonstationary signal analysis made it possible to extend the employment of muscle fatigue assessment to cyclic dynamic contractions, thus increasing noticeably its possible clinical applications. After presenting the basics of time-frequency distributions, we introduce instantaneous spectral parameters well suited to tracking spectral changes due to muscle fatigue, discuss the issues of quasi-stationarity and quasi cyclostationarity, and present different strategies of signal analysis to be utilized with cyclic dynamic contractions. We present preliminary results obtained by analyzing data collected from paraspinal muscles during repetitive lift movements, from the first dorsal interosseus during abduction-adduction movements of the index finger, and from knee flexors and extensors during isokinetic exercise. In conclusion, data herein reported demonstrate that the described techniques allow for evidencing the electrical manifestations of muscle fatigue in different paradigms of cyclic dynamic contractions. We believe that the extension of the objective assessment of the electrical manifestations of muscle fatigue from static to dynamic contractions may increase considerably the interest of researchers and clinicians and open new application fields, as ergonomics and sports medicine. PMID- 10527216 TI - Reproducibility of surface EMG variables and peak torque during three sets of ten dynamic contractions. AB - The interpretation of the electromyogram (EMG) of dynamic contractions might be difficult because the movement per se introduces additional factors that could affect its characteristics. There is a lack of studies concerning the reproducibility of surface EMG registrations during dynamic contractions. The aim was to investigate the during-the-day reproducibility (using intra-class correlation; ICC) of the peak torque (PT) and the EMG variables (without removing the electrodes) of dynamic contractions. Ten healthy subjects performed three sets of 10 dynamic maximum right-knee extensions with a one-hour interval in between, using an isokinetic dynamometer and the PT was determined. EMG signals were recorded from the right vastus lateralis, rectus femoris and vastus medialis muscles using surface electrodes and the mean frequency of the power spectrum (MNF [Hz]) and the signal amplitude (RMS [microV]), were computed. The ability to relax in-between the maximum extensions was calculated as a ratio of the RMS during the passive flexion phase and the RMS during the active extension phase of each contraction cycle: the signal amplitude ratio (SAR). Both PT (ICC = 0.99) and RMS (ICC = 0.83-0.98) had good reproducibility. The reproducibility of MNF was good for all muscles when the mean of contraction nos.: 1-10 was used. Vastus lateralis had the highest ICC among the three muscles. The reproducibility of SAR was generally poor (ICC < 0.60). The present study showed good reproducibility for common EMG variables (MNF and RMS) obtained during maximum isokinetic contractions. PMID- 10527217 TI - Evaluation of arterial supply to the spinal cord: different approaches in different settings. PMID- 10527218 TI - Assessment of ventricular function in critically ill patients: limitations of pulmonary artery catheterization. Institutions of the McSPI Research Group. AB - OBJECTIVE: To determine the accuracy of conventional hemodynamic assessment using pulmonary artery catheter-derived data in critically ill patients. DESIGN: Cohort study. SETTING: Kaiser Permanente and Veterans Affairs Medical Centers. PARTICIPANTS: Twenty-five consecutive patients who had undergone elective aortocoronary bypass surgery. MEASUREMENTS AND MAIN RESULTS: In the intensive care unit, conventional assessment (CA) was performed hourly by clinicians using conventional (radial artery and pulmonary artery) hemodynamic measurements from which left ventricular (LV) function and intracardiac volume were estimated. Simultaneously, transesophageal echocardiography (TEE) data were recorded on videotape, blinded to the clinicians, and quantitatively analyzed off-line. TEE determined LV function was classified as either normal (ejection fraction > or =40%) or abnormal (ejection fraction <40%) and intracardiac volume as normal (end diastolic area = 8 to 22 cm2), low (end-diastolic area <8 cm2), or high (end diastolic area >22 cm2). CONCLUSION: Evaluable data included 130 of 150 (87%) observations of simultaneously collected CA and TEE data, averaging 5.6+/-4.4 observations per patient. The overall predictive probability for conventional clinical assessment of normal ventricular function was 98% (118/121), whereas for abnormal ventricular function it was 0% (0/9). For CA of volume, the overall predictive probabilities for hypovolemia, normovolemia, and hypervolemia were 50% (3/6), 60% (69/115), and 22% (2/9). Although conventional clinical assessment of normal LV function in the intensive care unit correlates well with echocardiographic assessment, both LV dysfunction and extremes of preload (hypovolemia or hypervolemia) are assessed poorly by clinicians using conventional clinical monitoring with pulmonary artery catheterization. PMID- 10527219 TI - Recurrent laryngeal nerve palsy after cardiovascular surgery: relationship to the placement of a transesophageal echocardiographic probe. AB - OBJECTIVE: To examine the relationship between the incidence of recurrent laryngeal nerve palsy after cardiovascular surgery and the placement of a transesophageal echocardiographic probe. DESIGN: A prospective clinical study. SETTING: A single-institutional study in a university hospital. PARTICIPANTS: One hundred sixteen patients undergoing cardiovascular surgery. INTERVENTIONS: All patients were assigned into one of two groups: 64 patients in whom transesophageal echocardiography (TEE) was performed and 52 patients in whom TEE was not performed during surgery. The incidence of recurrent laryngeal nerve palsy was examined and compared between the two groups. MEASUREMENTS AND MAIN RESULTS: Five of 64 patients (7.8%) in whom TEE was monitored and 3 of 52 patients (5.8%) in whom TEE was not monitored were diagnosed with recurrent laryngeal nerve palsy postoperatively. There was no statistically significant difference between the incidence of recurrent laryngeal nerve palsy in patients with intraoperative TEE monitoring, and patients without it. The durations of surgery, anesthesia, and cardiopulmonary bypass were significantly longer in patients with nerve palsy than those without it. CONCLUSION: These results suggest that placement of the transesophageal echocardiographic probe is not responsible for postoperative recurrent laryngeal nerve palsy. It seems likely that surgical manipulation itself and the durations of surgery, cardiopulmonary bypass, and tracheal intubation are related to the incidence of laryngeal nerve palsy. PMID- 10527220 TI - Computed tomography-based tracheobronchial image reconstruction allows selection of the individually appropriate double-lumen tube size. AB - OBJECTIVES: To determine whether individualized selection of double-lumen tubes or alternatives based on three-dimensional reconstruction of the tracheobronchial image from routine preoperative computed tomography (CT) scans leads to clinically appropriate choices. DESIGN: Prospective observational study; comparison to historic controls. SETTING: Anesthesia and radiology facilities of a university medical center. PARTICIPANTS: Forty-nine patients undergoing thoracic surgery requiring one-lung ventilation. INTERVENTIONS: Three-dimensional image reconstruction of individual tracheobronchial anatomy was performed from routine preoperative spiral CT scans as well as from scans of five left-sided and four right-sided double-lumen tubes. Results of image-based tube size selection were compared with literature recommendations. Prospectively, individualized tube selection was performed by superimposition of printed transparencies of tubes over the tracheobronchial system and was validated using bronchoscopic and clinical criteria (n = 24). MEASUREMENTS AND MAIN RESULTS: Three-dimensional reconstruction visualized individual anatomy with good accuracy and resolution. Correlations between patient morphology and tracheobronchial dimensions were weak (height versus mainstem bronchial diameters: r < 0.50). In 11 of 48 patients (23%). CT-fitted double-lumen tube sizes would have differed from a conventional height-based and gender-based selection. Individual, prospective, CT-based double lumen tube selection was associated with (1) good fit and positioning confirmed by fiberoptic bronchoscopy, (2) adequate bronchial cuff seal volumes, (3) complete lung separation, and (4) oxygenation and ventilation parameters during one-lung ventilation similar to those with conventional size selection. In one patient, three-dimensional CT study allowed noninvasive evaluation of a tracheal stenosis precluding double-lumen tube placement. CONCLUSION: Individualized selection of double-lumen tube size using CT-based reconstructions of tracheobronchial anatomy leads to clinically appropriate choices. Risks resulting from variations in tracheobronchial morphology are recognized in advance. PMID- 10527221 TI - Somatosensory-evoked potentials during aortic coarctation repair. AB - OBJECTIVE: To determine the incidence of somatosensory-evoked potential (SSEP) changes and the interventions based on these changes during aortic coarctation repair. DESIGN: Retrospective review. SETTING: Single-institution, university hospital. PARTICIPANTS: Eighty-four children who had undergone surgical repair of aortic coarctation from January 1984 to May 1996. INTERVENTIONS: SSEPs were monitored in all patients throughout the procedure. A persistent decrease in amplitude greater than 50% from baseline was considered significant. Duration of SSEP changes in relation to the time course of surgical repair and whether a surgical or anesthetic intervention resulted from a change in SSEPs were documented. MEASUREMENTS AND MAIN RESULTS: Eighty-four patients underwent 87 surgical procedures. SSEP changes occurred in 40% of the procedures: 38.5% with repair and 15% with test clamp, with 9% occurring during both test clamp and repair. Interventions, which included repositioning the aortic cross-clamp, elevating blood pressure, and aborting surgery, occurred in 26.4% of all procedures based on SSEP changes. No patient sustained a neurologic deficit. CONCLUSION: This is the largest series to date describing the use of SSEPs in aortic coarctation repair. These SSEP changes were often immediately amenable to changes in surgical and anesthetic management. SSEP changes and interventions based on these changes occurred with a considerable frequency. PMID- 10527223 TI - The association between preoperative patient characteristics and both clinical and economic outcomes after abdominal aortic surgery. AB - OBJECTIVE: To evaluate the association between patient characteristics and both clinical and economic outcomes in patients having abdominal aortic surgery in Maryland between 1994 and 1996. DESIGN: Retrospective study using an administrative data set. SETTING: All Maryland hospitals that performed abdominal aortic surgery from 1994 through 1996 (n = 46). PARTICIPANTS: All patients who had abdominal aortic surgery in Maryland from 1994 through 1996 (n = 2,987). INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: The authors obtained discharge abstracts from the Maryland Health Services Cost Review Commission for patients with a primary procedure code for abdominal aortic surgery. Primary outcome variables were in-hospital mortality, hospital length of stay, and intensive care unit (ICU) days. The authors evaluated the following groups of independent variables: demographic characteristics, severity of illness, comorbid disease, and preoperative admission days. In multivariate analysis, independent predictors of in-hospital mortality were age 61 to 70 years (odds ratio [OR], 3.1; confidence interval [CI], 1.4 to 6.9), age 71 to 84 years (OR, 7.2; CI, 3.7 to 14.1), age 85 years or older (OR, 9.3; CI, 3.9 to 21.9), ruptured aneurysm (OR, 5.3; CI, 3.5 to 8.2), urgent operation (OR, 2.3; CI, 1.1 to 5.2), emergent operation (OR, 3.0; CI, 1.9 to 4.7), mild liver disease (OR, 4.6; CI, 2.0 to 10.9), and chronic renal disease (OR, 6.9; CI, 3.9 to 12.1). Hospital admission 1 to 2 days preoperatively was not associated with a difference in in-hospital mortality but was associated with a 31% increase in hospital days (CI, 23% to 40%) and a 38% increase in ICU days (CI, 19% to 60%). CONCLUSION: In patients having aortic surgery, several patient characteristics such as mild liver disease and chronic renal failure, were associated with increased in-hospital mortality and length of stay. The practice of admitting patients to the hospital 1 to 2 days before surgery should be reevaluated because this was not associated with reduced in-hospital mortality but was associated with increased hospital and ICU stay. PMID- 10527222 TI - Cerebral oxygenation during cardiopulmonary bypass measured by near-infrared spectroscopy: effects of hemodilution, temperature, and flow. AB - OBJECTIVE: To determine the effects of hemodilution, PaCO2, PaO2, arterial pressure, and temperature on cerebral oxygenation during mild hypothermic cardiopulmonary bypass (CPB). PARTICIPANTS: Fourteen patients electively scheduled for cardiac surgery. INTERVENTIONS: Oxyhemoglobin (HbO2), deoxyhemoglobin (Hb), hemoglobin differential (Hb-diff = HbO2-Hb), and oxidized cytochrome aa3 (CtO2) were measured with near-infrared spectroscopy (NIRS) during CPB. RESULTS: With onset of CPB, a significant decrease in HbO2 (median, -4.55 micromol/L; 25th to 75th percentile, -5.5 to -3.1; p < 0.05), Hb-diff (median, 3.88 micromol/L; 25th to 75th percentile, -4.7 to -1.9; p < 0.05), and CtO2 (median, -0.05 micromol/L; 25th to 75th percentile, -0.15 to 0; p < 0.001) occurred. The simultaneous decrease in arterial hemoglobin concentration (from 11.7 to 8.5 g/100 mL, p < 0.005) correlated significantly with changes in HbO2 (r2 = 0.71; p < 0.001), Hb-diff (r2 = 0.59; p < 0.005), and CtO2 (r2 = 0.57; p < 0.005). After 24 minutes of CPB, the largest decline in HbO2 (-5.03 micromol/L) and Hb-diff (-5.68 micromol/L) was recorded, whereas CtO2 showed no changes during cooling. During CPB, Hb and Hb-diff significantly correlated with the duration of CPB, PaO2 and PaCO2. CONCLUSIONS: In early stages of CPB, a diminished cerebral oxygen supply was found, which may be caused by acute hemodilution. Despite an increased extraction of oxygen as demonstrated by the decrease in Hb-diff, cerebral energy balance reflected by CtO2 was maintained within a safe range during cooling. Because NIRS measures regional cerebral oxygenation, it is useful as an adjunct to global measures in the early noninvasive detection of cerebral hypoxia. PMID- 10527224 TI - The safety and effectiveness of esmolol in the perioperative period in patients undergoing abdominal aortic surgery. AB - OBJECTIVES: To determine (1) if perioperative use of esmolol in major vascular surgery patients provides strict heart rate (HR) control, (2) what doses of esmolol are required to do this, and (3) does this control influence myocardial ischemia or result in adverse consequences. DESIGN: Prospective study of 40 patients randomized to two groups: The HR was controlled to either less than 80 beats/min (group 80) or less than 110 beats/min (group 110) using esmolol. Patients were monitored continuously for electrocardiographic changes perioperatively. HR control began after induction of anesthesia and continued for 48 hours thereafter. SETTING: Operating room and intensive care unit. PATIENTS: Patients undergoing abdominal vascular surgery involving aortic cross-clamping. INTERVENTIONS: Esmolol was titrated until the target HR was met. MEASUREMENTS AND RESULTS: Only one patient demonstrated an adverse effect. The median infusion rates were 100 and 12.5 microg/kg/min for groups 80 and 110. Target HR was met less in group 80 than in group 110, primarily in the postoperative period. Ischemia patterns were not significantly different between groups. CONCLUSION: Using esmolol for HR control in the intraoperative period for abdominal vascular surgery patients is effective and safe. HR control was much less effective in the postoperative period, but esmolol is safe when used at recommended doses. Further study with a larger number of patients is necessary to determine whether strict HR control with esmolol affects the incidence of myocardial ischemia or infarction in this patient population. PMID- 10527225 TI - Duration of preoperative amiodarone treatment may be associated with postoperative hospital mortality in patients undergoing heart transplantation. AB - OBJECTIVE: To assess the effect of preoperative amiodarone treatment on patient mortality after heart transplantation. DESIGN: Retrospective study. SETTING: Single-institution university hospital. PARTICIPANTS: One hundred six consecutive patients with heart transplants. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Patients were grouped according to duration of preoperative amiodarone treatment, and posttransplant mortality before hospital discharge was compared with patients not treated with amiodarone. The authors collected cardiovascular data in the preoperative and postoperative periods. There was a significant increase in posttransplant mortality before hospital discharge in patients treated with amiodarone for more than 4 weeks in the preoperative period (p < 0.05). Patients treated with amiodarone had significantly lower (p < 0.05) heart rates (mean heart rate, 103+/-19 beats/min) in the early postoperative period than patients not treated with amiodarone (mean heart rate, 111+/-15 beats/min), but there was no relationship with mortality (p = not significant). Patients who died had a significantly lower (p < 0.05) postoperative cardiac index (2.2+/-0.7 to 2.5+/-0.7) in the first 24 hours after cardiopulmonary bypass compared with patients who survived to hospital discharge (3.0+/-0.7 to 3.1+/-0.9), but there was no relationship to amiodarone treatment (p = not significant). CONCLUSION: Preoperative amiodarone treatment for more than 4 weeks may be associated with a significant increase in postoperative mortality in patients undergoing heart transplantation. Therefore, the indications for amiodarone must be carefully considered and, if needed, the maintenance dose should be kept to a minimum. PMID- 10527226 TI - Rocuronium versus vecuronium during fentanyl induction in patients undergoing coronary artery surgery. AB - OBJECTIVE: To evaluate the neuromuscular, ventilatory, and cardiovascular effects of rocuronium and vecuronium. DESIGN: Randomized, prospective, blinded study. SETTING: Tertiary care teaching center, single institution. PARTICIPANTS: Patients undergoing elective coronary artery bypass graft procedure. INTERVENTIONS: Patients received rocuronium, 1.0 mg/kg (n = 17), or vecuronium, 0.15 mg/kg (n = 15), during fentanyl induction of anesthesia. MEASUREMENTS AND MAIN RESULTS: Measures consisted of time to visual loss of orbicularis oculi twitches in response to facial nerve stimulation, ease of mask ventilation, hemodynamics, need for vasoactive drugs, and tracheal intubating conditions. Median time to twitch loss was faster (p < 0.05) after rocuronium (60 s) than after vecuronium (>84 s). Within 45 seconds, only 3 of 17 patients in the rocuronium group had moderate-to-severe difficulty with mask ventilation versus 12 of 15 patients in the vecuronium group (p < 0.05). Tracheal intubating conditions were excellent in all patients after rocuronium. In the vecuronium group, intubating conditions were excellent in 46%, good in 27%, and poor in 27% (p < 0.05 vrocuronium). Patients receiving vecuronium were more likely to require ephedrine and phenylephrine for hypotension (10/15 patients v 5/17 patients for rocuronium, p < 0.05). There were no clinically important differences in hemodynamic variables, oxygen metabolism, or myocardial ischemia between groups. CONCLUSION: During narcotic induction of anesthesia, rocuronium was associated with lower requirement for vasopressors, faster onset of neuromuscular blockade, and better conditions for mask ventilation and tracheal intubation compared with vecuronium. PMID- 10527227 TI - Intrathecal morphine for coronary artery bypass graft procedure and early extubation revisited. AB - OBJECTIVE: To determine the dose of intrathecal (IT) morphine (along with the intraoperative baseline anesthetic) that provides significant analgesia yet does not delay extubation in the immediate postoperative period in patients undergoing cardiac surgery and early extubation. DESIGN: Prospective, randomized, double blinded, placebo-controlled clinical study. SETTING: Single university hospital. PARTICIPANTS: Forty patients undergoing elective coronary artery bypass graft procedure and early extubation. INTERVENTIONS: Twenty patients received 10 microg/kg of IT morphine, and 20 patients received IT placebo. Perioperative anesthetic management was standardized and included postoperative patient controlled morphine analgesia. MAIN RESULTS: Of the patients tracheally extubated during the immediate postoperative period, mean time to extubation was similar in patients who received IT morphine (6.8+/-2.8 h) or IT placebo (6.5+/-3.2 h). Four patients who received IT morphine had extubation substantially delayed because of prolonged ventilatory depression. There was no difference between groups in postoperative patient-controlled morphine analgesia use. CONCLUSION: Even when used in conjunction with an intraoperative baseline anesthetic that allows early extubation, IT morphine (10 microg/kg) was unable to provide substantial postoperative analgesia. The risks of using IT morphine in patients undergoing cardiac surgery and early extubation may outweigh the potential benefits. PMID- 10527228 TI - Thoracic epidural anesthesia reduces infarct size in a canine model of myocardial ischemia and reperfusion injury. AB - OBJECTIVE: To determine the effects of thoracic epidural anesthesia on myocardial infarct size, regional myocardial blood flow (RMBF), and plasma norepinephrine in an anesthetized canine model of ischemia reperfusion injury with infarction. DESIGN: Blinded, randomized, placebo-controlled animal study. SETTING: Experiments were performed in the cardiothoracic research laboratory at Wake Forest University Baptist Medical Center. PARTICIPANTS: Anesthetized, open-chest mongrel dogs were used in these studies. METHODS: Dogs were instrumented for measurement of aortic pressure (AP) and left ventricular systolic pressure (LVSP), dP/dt, and RMBF Epidural catheters were inserted at thoracic segment T5. Three groups received epidural 0.5% bupivacaine: low-dose (n = 7; 0.3 mg/kg bolus, 0.15 mg/kg/ h), mid-dose (n = 7; 0.6 mg/kg bolus, 0.3 mg/kg/h), high-dose (n = 7; 1.2 mg/kg bolus, 0.6 mg/kg/h). The vehicle (VEH) group received epidural saline. Bolus followed by maintenance infusions began 30 minutes before the onset of ischemia (60 min) and continued through reperfusion (180 min). RESULTS: Myocardial infarct size was significantly reduced in the high-dose group versus the VEH and low-dose groups (p < 0.05). After initiation of the mid and high dose, AP, LVSP, and dP/dt decreased 7% to 16% (high vVEH; p < 0.05). VEH dogs showed a 130% increase from control in early postischemic RMBF. There was a dose dependent attenuation in this reflow response: 72%, 31%, and 6% increase in RMBF in the low, mid, and high groups, relative to controls (p < 0.05 high v VEH). Although there was no significant difference in plasma norepinephrine, fewer surges occurred in the high-dose group. CONCLUSIONS: Thoracic epidural anesthesia reduces infarct size and postischemic hyperemia in a model of ischemia reperfusion injury. PMID- 10527229 TI - Imaging of the anterior spinal artery by transesophageal color Doppler ultrasonography. PMID- 10527230 TI - Transesophageal echocardiography as an aid to surgical decision-making during resection of a rare thoracic neoplasm. PMID- 10527232 TI - Continuous percutaneous paravertebral block for minimally invasive cardiac surgery. PMID- 10527233 TI - Hypotension associated with clozapine after cardiopulmonary bypass. PMID- 10527231 TI - An adult atrial blood cyst. PMID- 10527234 TI - Neurophysiologic monitoring and outcomes in cardiovascular surgery. AB - The first step to make in improving neurologic outcome is to recognize and accept neurologic injury occurs in all patient groups undergoing CPB. Fortunately, that stage has now been passed. Accurate detection and documentation of the incidence of brain injury is the next progression. At the same time, the cause of the injury needs to be established. Since the introduction of CPB, numerous improvements and refinements have been achieved, making it the acceptable, everyday clinical tool that has enabled the development of cardiac surgery. Despite these improvements, CPB-related morbidity persists. The advent of new technologic advances drives the quest for new techniques. New protective strategies for many end organs, including the heart, kidney, and brain, are evolving. No organ system should be viewed in isolation; otherwise, organ specific protective strategies may arise in conflict. A strategy that confers absolute myocardial protection would be ideal, but at what cost to the protection of the kidneys, intestines, and brain? A neuroprotective strategy would ideally eliminate brain injury and be beneficial for all organs. The only way to continue to make progress is by the scientific evaluation of new techniques. The use of appropriate monitoring and outcome measures is fundamental to this process. PMID- 10527235 TI - Transesophageal echocardiography for port-access surgery. PMID- 10527236 TI - Case 3--1999. Intraoperative coronary thrombosis in association with low-dose aprotinin therapy. PMID- 10527237 TI - Pro: transesophageal echocardiography should be routinely used during pediatric open cardiac surgery. PMID- 10527238 TI - Con: transesophageal echocardiography should not be used routinely for pediatric open cardiac surgery. PMID- 10527239 TI - A complex echocardiographic diagnosis. PMID- 10527240 TI - An unusual echocardiograph after heart transplantation. PMID- 10527241 TI - Pro and con of heparin-bonded circuits for cardiopulmonary bypass. PMID- 10527242 TI - Is interpleural analgesia better than thoracic epidural analgesia after thoracotomy? PMID- 10527243 TI - Effect of temperature on the oxyhemoglobin dissociation curve. PMID- 10527244 TI - Intra-aortic balloon pump may cause arterial blood pressure discrepancy independently of cardiopulmonary bypass. PMID- 10527245 TI - Benign mediastinal emphysema during laparoscopic Nissen fundoplication. PMID- 10527246 TI - Tracheoesophageal fistula with total anomalous pulmonary venous drainage. PMID- 10527247 TI - Efficiency of inhaled nitric oxide as rescue therapy during severe ARDS: survival and factors associated with the first response. AB - PURPOSE: The purpose of this study was to determine if the response to inhaled nitric oxide (NO) as salvage therapy is an independent factor for survival in adult respiratory distress syndrome (ARDS) patients and to identify the factors that predict the response to inhaled NO during ARDS. MATERIALS AND METHODS: This was a multicenter, 2-year retrospective, clinical study in five university surgical or medical intensive care units, including all consecutive patients with ARDS in whom inhaled NO was tried. Clinical data (medical history, diagnoses), general severity scores (SAPS II, OSF), biological data, radiological and hemodynamic data at admission, at the beginning of the ARDS, and under treatment with inhaled NO were recorded. The NO response was defined as the variation of PaO2/Fio2 ratio before initiation and after 30 minutes of NO inhalation (VarPaO2/FiO2). RESULTS: Ninety-three patients aged 49 +/- 18 years were studied. Mean SAPS II was 45 +/- 16. Before the beginning of inhaled NO, PaO2/Fio2 ratio was 95 +/- 53 mm Hg and lung injury score 2.7 + 0.3. VarPao2/Fio2 when NO was started (11 +/- 4 ppm) was 26 +/- 44.5 mm Hg (median 17 mm Hg). Intensive care unit mortality was 74%. None of the parameters studied were predictors of response to inhaled NO, although there was a tendency for the youngest patients with the more severe hypoxemia to have a better response. Response to first inhaled NO test (VarPaO2/FiO2) was univariately associated with survival (Survivors: 45 +/- 44 mm Hg vs. Nonsurvivors: 20 +/- 43 mm Hg, P = .01), but this difference disappeared after adjusting for other prognostic factors (P = .16) selected by multivariate analysis. Finally, inhaled NO was continued for more than 1 day for 75 patients, and definitively stopped for 18 patients. Intensive care unit mortality (73% vs. 78%) was not different between these groups (P = .25, Log-rank test). CONCLUSIONS: We conclude that (1) efficacy of inhaled NO in improving oxygenation was moderate and difficult to predict, (2) response to first NO inhalation was not associated with prognosis, and (3) treatment of the most severe ARDS patients with inhaled NO did not influenced their intensive care unit survival. PMID- 10527248 TI - Nosocomial pneumonia with isolation of anaerobic bacteria in ICU patients: therapeutic considerations and outcome. AB - PURPOSE: Evaluate the influence of the anti-anaerobic antimicrobial therapy in the outcome of patients with nosocomial pneumonia. MATERIALS AND METHODS: The population study included 53 intensive care unit patients with nosocomial pneumonia in whom, using a protected specimen brush, anaerobic bacteria were isolated, which were associated or not with aerobes. Current and empirical antibiotherapies were retrospectively analyzed, regarding their efficacy against anaerobic bacteria. Since it was debated, sensitivity to cefotaxime, ceftazidime, and ciprofloxacin was determined in 38 strains of Prevotella species. Outcome was evaluated 10 days after the day of protected specimen brushes. Improvement was defined as a decrease of Murray score or ventilator weaning. RESULTS: The most frequently isolated bacteria were Prevotella species, which were more frequently resistant to cefotaxime (37%), ceftazidime (50%), and ciprofloxacine (32%) than usually reported in the literature. Sixty-six percent of these strains produced beta-lactamase. The effect of empirical anti-anaerobic antibiotherapy on the outcome at day 10 was evaluable in 39 patients. Twenty-nine patients were improved and 10 patients worsened. Interestingly, patients who had received well adapted antibiotics against anaerobes had a better outcome after 10 days (P < .02). CONCLUSIONS: This study suggests that specific antianaerobic therapy may be considered in the choice of empirical antibiotherapy in patients with nosocomial pneumonia. PMID- 10527249 TI - Adapting prognostic respiratory variables of ARDS in children to small-scale community needs. AB - PURPOSE: The clinical literature on the incidence and subsequent mortality of adult respiratory distress syndrome (ARDS) has come primarily from the experiences of large tertiary referral centers, particularly in Western Europe and North America. Consequently, very little has been published on the incidence, management, and outcome of ARDS in smaller community-based intensive care units. We aimed to delineate early clinical respiratory predictors of death in children with ARDS on the modest scale of a community hospital. MATERIALS AND METHODS: A retrospective chart review of children with ARDS needing conventional mechanical ventilation admitted to our pediatric intensive care unit from 1984 to 1997. The diagnosis of ARDS was based on acute onset of diffuse, bilateral pulmonary infiltrates of noncardiac origin and severe hypoxemia defined by partial pressure of oxygen <200 mm Hg during positive end-expiratory pressure (PEEP) of 6 cm H2O or greater for a minimum of 24 hours. Demographic, clinical, and physiological data including PaO2/ FIO2, A-aDo2, and ventilation index were retrieved. RESULTS: Fifty-six children with ARDS aged 8 +/- 5.5 years (range, 50 days to 21 years) were identified. The mortality rate was 50%. Early predictors of death included the peak inspiratory pressure (PIP), ventilation index, and PEEP on the third day after diagnosis: Nonsurvivors had significantly higher PIP (35.3 +/- 10.5 cm H2O vs 44.4 +/- 10.7 cm H2O, P < .001), PEEP (8 +/- 2.8 cm H2O vs 10.7.0 +/- 3.5 cm H2O, P < .01), and ventilation index (49.14 +/- 20.4 mm Hg x cm H2O/minute vs 61.6 +/- 51.1 mm Hg cm H2O/minute) than survivors. In contrast, PAO2/FIO2 and A-a DO2 were capable of predicting outcome by day 5 and thereafter. CONCLUSIONS: A small-scale mortality outcome for ARDS is comparable to large tertiary referral institutions. The PIP, PEEP, and ventilation index are valuable for predicting outcome in ARDS by the third day of conventional therapy. The development of a local risk profile may assist in decision-making of early application of supportive therapies in this population. PMID- 10527250 TI - Myocardial oxygen consumption during dobutamine infusion in endotoxemic pigs. AB - PURPOSE: Dobutamine infusion is used to increase whole-body oxygen delivery in septic patients to satisfy unmet oxygen demand of hypoxic tissues. However, dobutamine infusion also increases myocardial work and myocardial oxygen consumption. Our goal was to determine the importance of this effect as a fraction of the increase in whole-body oxygen consumption, in a porcine model of septic shock. MATERIALS AND METHODS: Four hours after a 50 microg/kg infusion of Escherichia coli endotoxin (0111: B4, Sigma) in eight anesthetized pigs, whole body oxygen delivery and myocardial oxygen delivery and consumption were calculated from blood flow and arterial and venous oxygen content measurements. We directly measured whole-body oxygen consumption by analysis of inhaled and exhaled gases using a metabolic cart. Then dobutamine 10 and 20 microg/kg/min was infused and measurements were repeated. RESULTS: Dobutamine infusion increased whole-body oxygen delivery but did not increase metabolic cart measured whole body oxygen consumption. Dobutamine infusion of 10 and 20 microg/kg/min increased myocardial oxygen consumption by 7.0 +/- 0.6 (80 +/-10%) and 12.0 +/- 2.0 mL O2/min (142 +/- 30%), respectively (P < .01). CONCLUSIONS: In this porcine model of sepsis, dobutamine infusion significantly increases myocardial oxygen consumption. Because whole-body oxygen consumption does not change, dobutamine infusion may fail to increase and may decrease oxygen consumption by other organs. PMID- 10527252 TI - Nitric oxide and nitrogen dioxide concentrations during in vitro high-frequency oscillatory ventilation. AB - PURPOSE: The purpose of this study was to measure nitric oxide (NO) and nitrogen dioxide (NO2) concentrations, at various ventilatory settings and sampling sites, during in vitro inhaled NO and high-frequency oscillatory ventilation therapy [iNO-HFOV]. MATERIALS AND METHODS: We used a high-frequency oscillatory ventilator (model 3100A, SensorMedics, Yorba Linda, CA), a test lung (model VT-2A Ventilator Tester, Bio-Tek Instruments, Inc., Winooski, VT), nitric oxide delivery and NO/NO2 monitoring (Pulmonox II, Pulmonox, Tofield, Canada), and scavenging systems in this study. The ventilator frequency, amplitude, and inspired oxygen concentration were systematically changed at a fixed flow of NO. The concentrations of NO and NO2, sampled at four sites, were determined by an electrochemical method (Pulmonox II). The NO and NO2 concentrations were measured at the proximal part of the inspiratory limb (site 1), near the Y-piece (site 2), the carina of the test lung (site 3), and the bellows of the test lung (site 4). RESULTS: The concentration of NO decreased significantly (P < .001) from the proximal port (site 11 of the inspiratory circuit (86.16 +/- 0.38 ppm) through the lung bellows (site 4) (70.08 +/- 0.23 ppm). The concentration of NO2 increased significantly (P < .001) from site 1 (3.25 +/- 0.04 ppm) through site 4 (19.4 +/- 0.19 ppm). However, the total concentration of NO + NO2 (NOx) remained unchanged at both site 1 and site 4. Increasing the frequency and amplitude of the ventilator significantly altered NO and NO2 concentrations. The NO2 concentration increased significantly (P < .0001) from 5.6 ppm to 18.1 ppm at site 4 when the fraction of inspired oxygen was increased from 0.25 to 0.93. The NO2 concentration also increased significantly (P < .0001) from 0.6 ppm to 18.7 when NO concentrations were independently increased from 12 ppm to 80 ppm. CONCLUSIONS: During HFOV, the concentrations of NO and NO2 vary between sampling sites and also are influenced by the frequency, amplitude, and inspired oxygen concentration. NO2 concentrations in the lung were significantly increased above commonly accepted toxic concentrations during ventilation with high concentrations of NO (80 ppm) and high fractional concentrations of oxygen. The excessive increase in NO2 concentration at the "alveolar" level in our test lung model warrants confirmation in an in vivo model. PMID- 10527251 TI - Selective vasodilation by nitric oxide inhalation during sustained pulmonary hypertension following recurrent microembolism in pigs. AB - PURPOSE: This study establishes a new model of sustained pulmonary hypertension induced by recurrent microembolism in pigs and evaluates the effects of nitric oxide (NO) inhalation in this model. MATERIALS AND METHODS: Fourteen pigs were embolized under general anesthesia with 300-microm microspheres intravenously three times over a period of 7 weeks. Four pigs served as untreated controls. Hemodynamic and gas exchange measurements were performed on days 1 and 7 after the last embolization. RESULTS: Recurrent microembolism caused sustained pulmonary hypertension (mean pulmonary artery pressure [MPAP] 26 +/- 2 and 18 +/- 1 mm Hg on days 1 and 7, respectively) compared with the control group (MPAP 13 +/- 1 mm Hg each for days 1 and 7; P < .05, respectively). Right heart hypertrophy was present at autopsy as indicated by an increase in minimal myocyte diameter. Inhaled NO (5 and 40 parts per million [ppm]) was administered on days 1 and 7. On both days, inhaled NO significantly reduced MPAP and pulmonary vascular resistance without affecting systemic hemodynamics. There were no differences in responses to 5 and 40 ppm inhaled NO. CONCLUSION: We conclude that recurrent microembolization in pigs provides a reliable model of sustained pulmonary hypertension. In this model inhaled NO is a selective pulmonary vasodilator, indicating that active vasoconstriction significantly contributes to sustained pulmonary hypertension after recurrent microembolism. PMID- 10527253 TI - Review of antibiotic prophylaxis recommendations for office-based urologic procedures. AB - Prophylactic antibiotic recommendations for urologic procedures are not well established. Any assessment of the need for antibiotics entails thorough cost benefit analysis. The subject of this article is an evaluation of the role of antimicrobial prophylaxis for outpatient office-based diagnostic procedures, including diagnostic flexible cystoscopy, transrectal ultrasound biopsy, and urodynamics. Relevant studies were identified using MEDLINE database searches and review of selected bibliographies. Studies of infections after transrectal ultrasound and biopsy suggest that periprocedure antibiotics are indicated, but that the exact course and timing have not been defined. Most evidence suggests that outpatient cystoscopy is associated with minimal infectious risk and that the routine administration of oral antibiotics is not indicated. Support in the literature for the use of prophylactic antibiotics at the time of urodynamic evaluation is equivocal. The current prophylactic regimens at the University of Michigan are presented as recommendations, but optimization of antimicrobial prophylaxis will require multicenter studies with large numbers of patients. PMID- 10527254 TI - In situ slings with concurrent cystocele repair. AB - Stress urinary incontinence (SUI) is commonly associated with varying degrees of genitourinary prolapse; therefore, it is customary to perform surgical corrections of both problems simultaneously. The type of surgical correction is based on the surgeon's discretion. We present a series of patients who underwent in situ vaginal wall slings as well as anterior vaginal wall (cystocele) repairs. Eighteen patients treated between 1994 and 1998 were evaluated. The average age was 61 years (range 35 to 74). Urodynamic evaluation was performed preoperatively. Postoperatively, the patients were assessed with objective testing as well as physician-performed Medical, Epidemiologic, and Social Aspects of Aging questionnaires. Follow-up ranged from 6 months to 4 years. SUI cure was defined as a patient who is completely dry and voiding. Sixteen (89%) of 18 patients were cured of both their cystocele and SUI; 2 of 18 had recurrent SUI with no evidence of recurrent cystocele. Fifty-six percent of the patients with good results had preoperative leak point pressures (LPP) of 50 to 100, and 44% had LPP > 100. None of the patients who were cured had an LPP <50, and only one patient in the failure group had an LPP <50. Seventeen percent of the patients had de novo urgency. In situ vaginal wall slings are a good procedure to use in combination with cystocele repairs in patients with LPP >50. PMID- 10527255 TI - Reconstruction or substitution of the pediatric urethra with buccal mucosa: indications, technical aspects, and results. AB - The main indication for using buccal mucosa in the urinary tract is in those who require complex secondary hypospadias surgery. Twenty-two children had inner lower lip mucosa onlay patch to reconstruct the urethra after 1 to 20 prior failed hypospadias/epispadias repairs. The mucosa was harvested from the lower inner lip by microsurgical dissection using optical magnification. The donor site was not sutured but was sealed with fibrin glue. The patches were anastomosed to the urethral plate using 7/0 polyglactin suture. The neourethra was covered with a vascularized dartos fascia or tunica vaginalis graft tunneled under the penile shaft skin. Special attention was given to closure of the glans and positioning of the meatus to the tip. Follow-up was between 12 and 72 months (mean 44). Complications included meatal stenosis in 1, fistula in 6, and wound infection in 1. The fistula rate decreased after changing the suture material and with increased experience. Lip mucosa was easy to harvest, and healing of the donor site was uncomplicated. PMID- 10527256 TI - Predictability and significance of multifocal prostate cancer in the radical prostatectomy specimen. AB - Multifocal prostate cancer has been reported in 50-76% of all cases of radical retropubic prostatectomy (RRP) specimens, but the clinical and prognostic significance of this finding is still unclear. A retrospective analysis of patients who underwent RRP between 1993 and 1997 was performed. Preoperative screening parameters and 4-mm RRP specimen sections were examined. The location, Gleason score, and extracapsular extension of the tumor recorded. Three hundred eight cases were reviewed. Mean follow-up was 4.2 +/- 1 years (range 2-6 years). Two hundred six patients (66.9%) had multifocal prostate cancer and 102 (33.1%) had unifocal prostate cancer. Of those with multifocal disease, 63% had two foci and 37% had three or more foci. There were statistical significant differences between both groups with respect to preoperative prostate-specific antigen (PSA) density of the transition zone (PSA-TZ), free/total (f/t) PSA, as well as percentage of patients with organ confined disease, high-grade tumors, and local recurrence. PSA-TZ (p = .001) and f/t PSA (p = .004) were significantly different between patients with unifocal and multifocal disease (0.9 vs. 2.2 ng/mL/cc and 18% vs. 6.5%, respectively). However, preoperative PSA (11.2 vs. 12.8 ng/mL; p = .09) and PSA density (0.17 vs. 0.19 ng/mL/cc; p = .07) were not able to predict unifocality or multifocality. These data suggest that multifocal prostate cancer is associated with higher grade, stage, and recurrence rate than unifocal prostate cancer. Preoperative PSA-TZ (> 1.5 ng/mL/cc) and f/t PSA (<9%) may predict multifocality in the RRP specimen. PMID- 10527257 TI - Needle imaging during ultrasound-guided permanent prostate implants. AB - As demonstrated by water phantom experiments and clinical observations, the reverberation artifact associated with the ultrasound needle image during permanent prostate implants is extremely useful in determining precise radioactive seed positioning. It also serves as an independent quality assurance check of the number of seeds in the strand. PMID- 10527258 TI - Repetitive prostatic massage therapy for chronic refractory prostatitis: the Philippine experience. AB - Patients frustrated with failure of traditional therapy for prostatitis have traveled to the Philippines and elsewhere for repetitive prostatic massage combined with antibiotic therapy. The aim of our study was to evaluate prospectively the response of patients who traveled to Manila to undergo this treatment. Twenty-six patients consented and were registered by the Prostatitis Foundation (B.H.) and subsequently evaluated (J.C.N., J.D.) prior to and following treatment (A.E.F.). Evaluation at baseline and after treatment consisted of standardized history and previously validated prostatitis-specific Symptom Frequency Questionnaire (SFQ) and Symptom Severity Index (SSI), International Prostate Symptom Score (I-PSS) and Quality of Life (QoL) questionnaire, the O'Leary Sexual Function Inventory (SFI), and a Subjective Global Assessment (SGA). Treatment in Manila consisted of triweekly prostatic massage combined with specific culture directed and/or empirical antimicrobial therapy for 6 to 12 weeks. Twenty-two patients completed at least one follow-up assessment and 12 patients completed 2-year assessment (average follow-up of 17 months in 22 patients). There was a significant decrease in average symptom severity (SSI) by 4 months that continued for 2 years, but less improvement in symptom frequency (SFQ) and quality of life (QoL), and no significant improvement in voiding symptoms (I-PSS) or sexual function (SFI) at time of last assessment. Forty-six percent of the 22 evaluable patients had >60% decrease (significant improvement) in symptom severity (SSI), whereas 27% had similar significant improvement in frequency of symptoms (SFQ) when last assessed. Thirty-three percent reported marked subjective improvement (SGA) at last evaluation. Of the 12 patients who completed the 2-year follow-up, 5 of the original 26 had a significant and sustainable improvement in objective and subjective measurements of frequency and severity of symptoms. The combination of prostatic massage and antibiotics for treating difficult refractory cases of prostatitis may be promising, but its ultimate value needs to be confirmed. Studies in patients with less refractory and shorter duration disease may allow us to predict who will respond to this therapeutic approach. PMID- 10527259 TI - Use of the laparoscopic knot introducer in urethrovesical anastomosis following radical prostatectomy. AB - One of the most delicate stages of retropubic radical prostatectomy intervention is urethrovesical anastomosis, especially if it is performed in a deep bony pelvis with a short urethral stump. Correct knot tying is essential to avoid the risk of postoperative anastomotic leakage. In such conditions, the urethrovesical anastomosis is performed with six 4-0 Monocril sutures and the knots are tied under vision using the laparoscopic knot introducer. In a 2-year period we performed 106 retropubic radical prostatectomies in our urology department. In four cases (3.8%) we performed urethrovesical anastomosis with the aid of the described technique. Two weeks after catheter placement, the radiologic control showed no leakage or contracture of the anastomosis. Mean follow-up is 14.8 months (range 8 to 20); all of the patients are continent. A second radiologic study 6 months after the intervention documented in all cases a correct new anatomical repair without stenoses or contracture. This technique is a minor modification of the direct urethrovesical anastomosis that facilitates the anastomosis between the urethral stump and the bladder neck and reduces the risk of anastomotic leakage due to incorrect suture knot positioning and tying. PMID- 10527260 TI - Combination therapy in the treatment of patients with staghorn calculi. AB - The treatment of patients with staghorn calculi remains a challenging problem. Combination therapy using percutaneous nephrolithotomy and extracorporeal shock wave lithotripsy has been recommended as the best option for most patients. Using this technique, 10 (83%) of 12 renal units with partial or complete staghorn calculi were rendered stone-free, with no significant septic episodes or serious complications. Blood transfusion was necessary in three patients. "Sandwich" therapy using initial percutaneous debulking followed by extracorporeal shock wave lithotripsy and/or "second-look" nephroscopy offers patients a high likelihood of achieving a stone-free state while avoiding the morbidity and lengthy recovery associated with open surgery. PMID- 10527261 TI - Magnetic resonance urography in the evaluation of acute flank pain. AB - We report our clinical experience with magnetic resonance urography in 13 patients. This noninvasive relatively new application of magnetic resonance to initial radiologic evaluation of patients with acute flank pain offers valuable diagnostic information and may provide additional support in defining the need for possible further therapeutic intervention in selected cases. PMID- 10527262 TI - Right intrathoracic renal ectopia: a case report and review of the literature. AB - We report a case of right intrathoracic renal ectopia associated with a previously unreported congenital anomaly (trisomy 21) and review of the literature. PMID- 10527264 TI - Hand-assisted laparoscopic donor nephrectomy versus standard laparoscopic donor nephrectomy: a comparison study in the canine model. AB - The aim of this study was to compare live donor nephrectomy by hand-assisted laparoscopy to standard laparoscopy in a canine model. Fourteen dogs underwent a left laparoscopic nephrectomy; a standard laparoscopic nephrectomy technique was utilized in seven dogs. In a second group of seven dogs, a hand-assisted laparoscopic technique was used with a Dexterity Pneumo Sleeve hand port. All nephrectomies were performed as "donor" nephrectomies, dividing the vessels last. Total blood loss, operative warm ischemia, time and organ retrieval times were assessed for each group. The average operative time was significantly shorter for hand-assisted laparoscopic donor nephrectomy (32 +/- 8 minutes vs. 61 +/- 8 minutes; p = .02) than for the standard technique. The average warm ischemia (86 +/- 24 seconds vs. 224 +/- 52 seconds; p = .03) and average organ delivery times (4 +/- 3 seconds vs. 45 +/- 9 seconds; p < .01) also were shorter using the hand assisted laparoscopic technique. No significant differences in average blood loss were found between the two groups (9 +/- 2 cc vs. 6 +/- 1 cc; p = 0.16, NS). Good parenchymal, ureteral, and vascular preservation was achieved by both techniques. Hand-assisted laparoscopy permits shorter operating times and warm ischemia times than standard laparoscopy in a canine model of donor nephrectomy. Hand assistance makes donor laparoscopic nephrectomy technically easier and significantly quicker to perform. If hand-assisted laparoscopy donor nephrectomy is confirmed to be a rapid and safe technique for removing an intact organ, laparoscopic nephrectomy will be a more widely accepted technique among urologists who participate in living related donor kidney transplantation. PMID- 10527263 TI - Endovascular stent graft for management of ureteroarterial fistula after orthotopic bladder substitution. AB - We describe the first case of an ureteroarterial fistula developing after orthotopic neobladder substitution and its minimally invasive management using endovascular stent grafting. We outline the risk factors for the development of ureteroarterial fistulae and trace the evolution of diagnostic and therapeutic modalities used in the management of these life-threatening complications. Minimally invasive management with endovascular stent grafting and exclusion of two pseudoaneurysms in the iliac artery system was performed successfully. After successful endovascular exclusion of two pseudoaneurysms, the patient's hematuria resolved and he recovered fully. Three-dimensional computed tomography performed 3 months later documented a patent aortoiliac arterial system without evidence of pseudoaneurysm or endovascular leak. Ureteroarterial fistula after orthotopic bladder substitution was managed with an endovascular stent graft without the need for extra-anatomical vascular bypass. Early recognition, stabilization, and angiographic evaluation followed by this minimally invasive technique may avoid open operative repair and attendant morbidity. PMID- 10527265 TI - Interaction of the protein C/protein S anticoagulant system, the endothelium and pregnancy. AB - Normal pregnancy is associated with significant changes in haemostasis, lipid metabolism and endothelial function. This suggests that maternal adaptation in these systems is required for successful pregnancy outcome. A number of acquired and heritable prothrombotic abnormalities are associated with complications in pregnancy. A common feature of these abnormalities is their ability to alter endothelial function or the protein C/protein S system and increase thrombin generation. In this review the normal function of the endothelium and the protein C/protein S system is detailed. The changes which characterize normal and complicated pregnancies are outlined and the evidence for the impact of heritable and acquired disorders of the protein C/protein S system on pre-eclampsia and fetal loss are discussed. PMID- 10527266 TI - AIDS-related lymphoma. AB - The incidence of intermediate and high grade B-cell non-Hodgkin's lymphomas in HIV-infected individuals is approximately 60 times greater than in the general population. These AIDS-related lymphomas (AIDS-NHL) are a late manifestation of HIV infection and may increase in frequency as patients live longer with highly active antiretroviral therapy and effective prophylaxis of opportunistic infections. AIDS-NHL have unique clinical and pathological features that are different from non-Hodgkin's lymphomas in the general population. Histologically AIDS-NHL are either high (2/3) or intermediate (1/3) grade lymphomas. Clinically AIDS-NHL have a preponderance for extranodal involvement with central nervous system being the most common site for this. In addition to the clinical and pathological features of AIDS-NHL, a current knowledge of their pathogenesis and treatment options are presented in this review. PMID- 10527267 TI - Immune regulation in multiple myeloma: the host-tumour conflict. AB - Multiple myeloma (MM) remains essentially incurable by conventional anti-tumour therapy. This has led to increased interest in the possibility that forms of immune therapy might be effective. The successful use of donor lymphocyte infusions (DLI) in a few cases of MM relapse following allogeneic bone marrow transplantation have added weight to this view. MM is characterized by several defects in the host's immune system. The influence of the malignant clone on the function of the immune effector cells results from both passive and active suppression. Despite an array of functional adhesion molecules and HLA class I and II molecules on their surface and the secretion of a tumour-specific peptide, they fail to express adequate levels of co-stimulatory molecules thus inducing anergy in potentially tumour-specific T cells. In addition to this passive evasion of immune regulation, MM tumour cells are capable of producing a number of immunologically active agents which can induce immunosuppression such as transforming growth factor-beta, Fas ligand (FasL), vascular endothelial growth factor and Muc-1. It is postulated that these agents may be produced by the tumour cell to influence the microenvironment to support growth and differentiation of the clone but may have the additional benefit of altering the function of the host immune effector cells and thus preventing tumour rejection. This duality of function is important if clinicians are to design immunotherapy strategies which will achieve the true potential and result in improved survival in MM. PMID- 10527268 TI - Splenic irradiation in myelofibrosis with myeloid metaplasia: a review. AB - Morbidity from myeloid metaplasia and myelofibrosis arises from progressive anemia and abdominal discomfort related to massive splenomegaly, which may be associated with hypercatabolic symptoms. To date, no therapy, other than allogeneic bone marrow transplantation, has been shown to cure or to prolong the survival of these patients. Thus, current management strategies are palliative and include red cell transfusional support and androgen therapy for anemia; chemotherapeutic agents for control of thrombocytosis, leukocytosis, and hypermetabolic symptoms; and splenectomy or splenic irradiation for symptomatic splenomegaly. The major indication for splenic irradiation is left upper quadrant discomfort related to massive splenomegaly, usually in patients for whom splenectomy is contraindicated or has been declined. In most patients, it provides relief from abdominal pain and a moderate reduction in splenic size. Although responses are transient, some patients may experience prolonged relief. Splenic irradiation can result in prolonged myelosuppression in certain patients. This calls for cautious dosing, because individual sensitivity is variable and cannot be predicted. The use of splenic irradiation does not preclude subsequent splenectomy; however, the increased risk of postoperative hemorrhage should discourage consideration of splenic irradiation as an alternative or a temporizing measure before splenectomy when indicated. PMID- 10527269 TI - Perfluorinated blood substitutes and artificial oxygen carriers. AB - Blood transfusion is a remarkably safe, routine clinical procedure. However, the need for sophisticated blood processing, storage and cross-matching, coupled with increasing concerns about the safety of blood products, has fuelled the search for safe and efficacious substitutes. Candidate materials based on modified haemoglobin (including recombinant molecules) or highly inert, respiratory gas dissolving perfluorinated liquids (perfluorochemicals) have been developed. The latter are immiscible in aqueous systems and must, therefore, be injected as emulsions. Second-generation perfluorochemical emulsions are available and in clinical trials as temporary intravascular oxygen carriers during surgery, thereby reducing patient exposure to donor blood. One commercial product is currently under Phase III clinical evaluation, with regulatory approval expected within 1 2 years. Other biomedical applications for perfluorochemicals and their emulsions include their use as pump-priming fluids for cardiopulmonary bypass, lung ventilation fluids, anti-cancer agents, organ perfusates and cell culture media supplements, diagnostic imaging agents and ophthalmologic tools. Novel applications for perfluorochemicals as immunomodulating agents are also being explored. PMID- 10527270 TI - Transoral carbon dioxide laser resection of supraglottic carcinoma. AB - Between 1981 and 1994, 34 patients with squamous cell carcinoma of the supraglottis were treated by transoral carbon dioxide laser resection, 12 of them palliatively. Additional treatment included neck dissection in 21 patients and radiotherapy in 24 patients. The 3-year overall survival was 62%, and the actuarial survival 80%. The overall survival for T1 and T2 tumors was 71%, and that for T3 and T4 tumors was 47%. The overall 3-year survival for the early stages, I and II, was 88%, and that for the advanced stages, III and IV, was 50%. These results are comparable to the outcome after conventional open partial resection. Given the significantly lower morbidity (only 7 patients required tracheostomy), we do not observe an age limit anymore. The transoral method can be recommended as curative treatment in T1 and T2 tumors and in selected T3 and T4 tumors in concert with neck dissection and/or radiotherapy. In patients with advanced inoperable tumors, laser surgery is an excellent alternative to tracheostomy and palliative radiotherapy. Prerequisites for successful application of the transoral carbon dioxide laser resection are adequate resection techniques. PMID- 10527271 TI - Management of the neck in cancer of the larynx. AB - In the Department of Otolaryngology at the University of Pittsburgh School of Medicine, cancer of the larynx is usually treated by primary surgery. Radiotherapy is used as adjuvant treatment in certain patients who have cancer that has adverse histologic features such as perineural, vascular, and/or cartilage invasion. With this approach, patients rarely develop local recurrence. Patient survival is therefore unlikely to be improved by changes to the management of the primary tumor. Survival may, however, be improved by reducing the incidence of recurrence in the neck, as well as distant. Hence, we have adopted an aggressive surgical approach to the cN+ as well as the N0 neck. The theoretical basis for this aggressive surgical approach to the neck will be considered under the following headings: staging, regional control, distant metastasis, survival, choice of neck dissection, and the pathologically positive elective neck dissection. PMID- 10527272 TI - Minimally invasive device to effect vocal fold lateralization. AB - Despite many operative procedures focused on vocal fold lateralization, none has achieved an acceptable level of dependability. Bilateral vocal fold abductor paralysis is treated by arytenoidectomy, cordotomy, suture lateralization, or partial cordectomy. Tracheotomy remains the gold standard for maximizing the airway and preserving phonatory function. We have developed a device that is minimally invasive, tunable, and reversible, with the potential for lateralization or medialization of the vocal process. The device consists of a polyethylene collar, a Vitallium cam, and a double-helix core for engaging soft tissue. It is introduced through a circular opening in the thyroid cartilage by a modified thyroplasty approach. Both the first and second iterations of this device have been evaluated for clinical effectiveness in 9 sheep by means of photographic and video documentation. Effectiveness in humans is currently being assessed. The results of the animal study permit us to have substantial optimism with respect to the clinical application of this device. PMID- 10527273 TI - Subglottic stenosis: correlation between computed tomography and bronchoscopy. AB - The evaluation of subglottic stenosis has been limited by the lack of standardized methods for determining the cross-sectional area and length of the stenotic segment. A rabbit model was used to prospectively evaluate the correlation between computed tomography (CT) and bronchoscopy in the evaluation of this disease. Subglottic stenosis was produced in 39 New Zealand White rabbits by a transoral endoscopic technique. The animals were evaluated 3 weeks later with spiral CT, rigid bronchoscopy, and open laryngotracheal exploration. Spiral CT was performed with the location, degree, and length of subglottic stenosis being determined by a blinded observer. Each animal then underwent rigid bronchoscopy and open laryngotracheal exploration for determination of the same measurements. Data were analyzed to determine the correlation between the radiographic and surgical techniques in evaluating the airway stenosis. With regard to the degree of stenosis, 94% of the rabbits were determined to have CT and bronchoscopic measurements that were within 15% (Pearson correlation .94, p < .05). With regard to the length of stenosis, 94% of animals had a measurement on CT that was within 2 mm of that observed upon open exploration (Pearson correlation .81, p < .05). The CT evaluation of subglottic stenosis correlated well with the currently used method of visual inspection at bronchoscopy in evaluating tracheal stenosis in this animal model. These data suggest that CT could serve as a useful adjunct in the evaluation of tracheal stenosis, especially when serial examinations are required. PMID- 10527274 TI - Palliative treatment for tracheal stenoses using carbon dioxide laser and the Gianturco stent. Long-term results. AB - Between September 1992 and March 1998, the self-expandable Gianturco prosthesis was inserted in 23 patients suffering from tracheal stenosis. After radial incision and dilation of the stenosis as described by Shapshay, the positioning of the stent was performed during an endoscopic procedure under optical control. The prosthesis used was a double-ring stent 50 mm long and 20 mm in diameter. The follow-up period ranged between 0.5 and 67 months with an average of 31 +/- 18 months. Pulmonary function tests showed an average improvement of the peak expiratory flow (50%) from preoperative results of 1.06 +/- 0.60 L/s to short term postoperative results of 2.08 +/- 0.78 L/s and long-term postoperative results of 2.11 +/- 0.78 L/s. The mean peak inspiratory flow (50%) improved from 1.43 +/- 0.85 L/s to 2.40 +/- 1.29 L/s at short term and to 2.56 +/- 1.20 L/s at long term. Eight patients out of the 23 had to undergo a second endoscopic procedure: 3 patients for granuloma vaporization; 1 patient to change a malpositioned stent; 2 patients to add a second stent because of insufficient tracheal enlargement; and 2 patients to resect mucosal membranes between the 2 stent rings and to place a second stent. Optical control of the accurate positioning and use of this model of Gianturco prosthesis helped to avoid the severe complications described in the literature (migration, extrusion, fracture, wall erosion. and hemorrhage). The follow-up must particularly target the prevention of granulomas. The self-expandable Gianturco prosthesis can be advocated for long-term palliative treatment of tracheal stenoses that are inoperable by an external surgical approach. PMID- 10527275 TI - Expression of Epstein-Barr virus latent membrane proteins leads to changes in keratinocyte cell adhesion. AB - Epstein-Barr virus (EBV) has 3 latent membrane proteins (LMPs)--LMP1, LMP2a, and LMP2b--which are expressed in nasopharyngeal carcinoma. Using keratinocyte cell lines expressing LMP2a and LMP2b and coexpressing LMP1/LMP2a, we grew organotypic raft cultures to analyze changes in morphology and expression of the cell adhesion molecule ICAM-1; alpha2, alpha3, alpha5, beta1, and alpha6beta4 integrins; laminin 5; E-cadherin; and desmoplakin. Cells expressing LMP2a or LMP2b were defective in their ability to mature and progress through normal squamous stratification when compared to the parental cell lines. Cells coexpressing LMP1/LMP2a additionally demonstrated "pseudoinvasion" into the raft dermal equivalent. There was a consistent and dramatic up-regulation in the suprabasal expression of laminin 5 and alpha6beta4 and beta1 integrins in the LMP expressing cell lines. ICAM-1, not expressed in the control cell lines, was up regulated in the LMP-expressing cell lines. Expression of alpha3 and alpha5 integrins was also up-regulated in the LMP-expressing cell lines, while alpha2 demonstrated a loss of the normal basal layer expression. E-cadherin and desmoplakin expression patterns were essentially unchanged. We conclude that LMP2a and LMP2b singly, and LMP1/LMP2a coexpressed, are capable of altering keratinocyte cell adhesion molecule expression consistent with nasopharyngeal carcinoma. PMID- 10527276 TI - Distant lymphatic metastasis from head and neck cancer. AB - A predictable pattern of metastasis based on tumor histology and site of origin has been well documented for most cancers that arise in the head and neck region. The current study demonstrates that this predictable pattern of metastasis can be significantly impacted by previous therapy, resulting in unusual patterns of metastasis in patients with recurrent tumors. A retrospective case series of 5 patients with head and neck carcinomas who developed metastases to distant lymph nodes is presented. All patients underwent surgery and radiotherapy to the primary tumor and regional lymphatics at the time of their initial treatment. All of the patients developed a local recurrence less than a year before the detection of distant lymphatic metastases. Cytology or excision confirmed metastases to the axillary, inguinal, or anterior intercostal lymph nodes. All of the patients underwent aggressive surgery for attempted cure of the local recurrence shortly before the presence of distant lymphatic metastases was clinically recognized. The metastatic workup of patients with carcinomas of the head and neck frequently includes examination of the regional lymph nodes as well as chest radiography, liver function tests, and serum calcium determination. This evaluation may fail to detect metastases to distant lymph nodes in patients who present with recurrent or second primary cancers. Such patients should undergo careful examination of all major lymph node-bearing regions of the body when being evaluated for the presence of distant metastases. PMID- 10527277 TI - Montgomery salivary bypass tube in the reconstruction of the hypopharynx. Cost benefit study. AB - This study analyzes the postoperative complications and the functional results in 61 patients who underwent total laryngectomy with partial or total (circumferential) pharyngectomy reconstructed with a pectoralis major myocutaneous flap, in relation to the use of the Montgomery Salivary Bypass Tube (MSBPT). There were no significant differences regarding frequency of postoperative cervical complications in relation to the use of the MSBPT. The median hospital stay for patients without the MSBPT (36 days) was significantly higher than that for patients with the MSBPT (25 days). Although the MSBPT did not modify the rate of complications at the cervical level, it did reduce their severity. A financial study showed the cost-effectiveness of using the MSBPT. Systematic use of the MSBPT is recommended after total laryngectomy with partial or total pharyngectomy reconstructed with a pectoralis major myocutaneous flap. PMID- 10527278 TI - Histochemical and immunocytochemical study of nitrergic innervation in human nasal mucosa. AB - Nitric oxide (NO) is a free radical gas that has been found to be produced in neuronal cells by the action of the enzyme brain nitric oxide synthase (bNOS). The aim of this study was to identify NO-containing nerve structures in the human nasal mucosa by localizing bNOS and to find out whether NO production is attached to the parasympathetic system. For this purpose, immunocytochemistry with antibodies to bNOS and neurofilament was performed. Additionally, nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d), an enzyme that correlates with the localization of NO synthase, and acetylcholinesterase were visualized in a histochemical double staining technique on frozen sections. The NADPH-d and bNOS reactions were found in axons of nerve bundles and in subepithelial, glandular, and vascular nerve fibers. Arteries showed a distinctly developed nitric innervation, whereas no activity was found in nerve fibers supplying veins. A high coexistence of NADPH-d in parasympathetic nerves could be detected. These findings suggest that NO takes part in the nerve control functions of the human nasal mucosa. PMID- 10527279 TI - Unilateral tonsillar enlargement and tonsillar lymphoma in children. AB - The clinical presentation and surgical and pathological findings of 46 children with unilateral tonsillar enlargement (UTE; age range 2 to 13 years, mean age 6.5) who underwent tonsillectomy for biopsy purposes between 1975 and 1995 were compared with those of 7 children who received treatment for tonsillar lymphoma (TL; age range 2 to 9 years, mean age 4.8) during the same period. There was no history of rapid tonsillar enlargement in children in the UTE group, and only 20 (43%) were symptomatic. Symptoms included recurrent sore throats in 10 patients (22%), snoring in 5 (11%), nasal obstruction in 4 (9%), and dysphagia in 1 (2%). No children had systemic symptoms or significant cervical lymphadenopathy. In contrast, tonsillar enlargement was observed to occur within a 6-week period in all children with TL, and 6 (86%) children had symptoms at presentation that included dysphagia in 5 (71%), snoring in 3 (43%), night sweats in 2 (29%), and fever and rigors in 2 (29%). Cervical lymphadenopathy greater than 3 cm was present in 6 (86%) children, while 1 child (14%) had hepatosplenomegaly. There was no histopathologic evidence of neoplasia in the UTE group, and a true discrepancy in size between the two tonsils was confirmed in only 21 of 44 (48%) cases. All 7 patients in the TL group had non-Hodgkin's lymphoma. All received chemotherapy, with 5 of the 7 cured and 2 dying of disease. The data suggest that tonsillectomy should be performed for biopsy purposes in UTE where there is a history of progressive enlargement, significant upper aerodigestive tract symptoms, systemic symptoms, suspicious appearance of the tonsil, cervical lymphadenopathy, or hepatosplenomegaly. The diagnosis of TL should also be considered when UTE is present in an immunocompromised child or one with a previous malignancy, when acute tonsillitis is asymmetric and unresponsive to medical treatment, or when rapid bilateral tonsil enlargement occurs. Observation is appropriate management for other cases of UTE. PMID- 10527280 TI - Management of persistent tracheocutaneous fistula in the pediatric age group. AB - Different surgical methods have been advocated for closure of persistent tracheocutaneous fistula (TCF) in children. The objective of this study was to compare different methods of repair and postoperative care that were used for management of TCF in children. The charts of 98 children with persistent TCF who were surgically managed in our department between January 1990 and April 1997 were reviewed retrospectively. Excision of the fistulous tract and healing by secondary intention was employed in 18 patients. Eighty patients were managed by tract excision followed by primary closure. Sixty-three patients remained intubated for 18 to 24 hours postoperatively, while 17 patients were extubated in the recovery room. One patient had a large tracheal granuloma on follow-up endoscopy. Three patients needed a second procedure. No significant correlation was found between the method of surgical repair or the length of postoperative intubation and outcome. In our experience, TCF repair, either by primary closure or secondary intention, is a relatively safe and effective procedure in the pediatric age group. Preoperative evaluation and possible indications for selecting the method of repair are discussed. PMID- 10527281 TI - Otalgia and aversive symptoms in temporomandibular disorders. AB - The term Costen's syndrome has been used in the dentomedical literature to describe a constellation of craniofacial symptoms. Since some of the same complaints have been reported in patients with "generalized" psychological distress, symptoms associated with the syndrome may not be useful in differential diagnosis of temporomandibular disorders. The present study investigated whether some somatic complaints, particularly tinnitus and dizziness, were pathognomonic in patients with chronic temporomandibular pain. Illness behavior and personality factors were studied for possible interrelationships with these symptoms. Factor analysis revealed that tinnitus and dizziness loaded on separate factors. Tinnitus loaded with nasal stuffiness, tearing, and itching of the eyelids and nose, while dizziness loaded with complaints of altered taste and smell and blurred vision. Neither was consistently related to measures of pain or to indices of illness behavior or somatic focus. PMID- 10527282 TI - Otoendoscopy for improved pediatric cholesteatoma removal. AB - Our objective was to determine the usefulness of intraoperative rigid endoscopy in detecting incompletely removed cholesteatomas, and to learn whether "second look" procedures are still needed in children. We used 30 degrees, 2.7-mm endoscopes to evaluate the middle ears of 14 children (29 procedures) with cholesteatomas once all visible disease had been removed under the operating microscope. If residual cholesteatoma was seen, removal continued until all disease visualized with the endoscope was removed. If the cholesteatoma was not removed intact, planned exploratory surgery was performed. The rigid endoscope detected incompletely removed cholesteatomas at surgery in 7 of the 29 cases (24%). In 2 of the 11 cases (18%) judged free of cholesteatoma by both otomicroscopy and otoendoscopy, residual disease was found at planned exploratory procedures. While otoendoscopy is clearly useful in detecting incompletely removed cholesteatoma, a substantial rate of residual disease following "complete" removal suggests the continued need for planned exploratory procedures. PMID- 10527283 TI - Management of abducens palsy in patients with petrositis. AB - Our objective was to discuss the management and outcome of abducens nerve palsy in patients with Gradenigo's syndrome. In a retrospective analysis of patients with Gradenigo's syndrome at a tertiary-care center in Houston, Texas, from 1987 to 1995, we identified 2 patients with Gradenigo's syndrome, both female. One had bilateral involvement, so that the total was 3 ears. Both patients had complete recovery of their abducens nerve palsy. In 2 ears with chronic mastoiditis, sixth nerve palsies failed to respond to medical therapy alone, but resolved after mastoidectomy with drainage of the petrous apex. We recommend that patients with Gradenigo's syndrome and evidence of chronic mastoiditis be treated with aggressive medical and surgical care. PMID- 10527284 TI - Coexpression of neurotrophic growth factors and their receptors in human facial motor neurons. AB - Neuronal development and maintenance of facial motor neurons is believed to be regulated by neurotrophic growth factors. Using celloidin-embedded sections, we evaluated immunoreactivity of 11 neurotrophic factors and their receptors in facial nuclei of human brain stems (4 normal cases, and 1 from a patient with facial palsy and synkinesis). In the normal subjects, positive immunoreactivity of the growth factor neurotrophin-4 and acidic fibroblast growth factor (aFGF) was observed in facial motor neurons, as was positive immunoreactivity against ret, the receptor shared by glial cell line-derived neurotrophic factor and neurturin. Immunoreactivity was moderate for the receptor trkB and strong for trkC. In the case of partial facial palsy, surviving cells failed to show immunoreactivity against neurotrophins. However, immunoreactivity of aFGF was up regulated in both neuronal and non-neuronal cells in this patient. Results suggest that these trophic growth factors and their receptors may protect facial neurons from secondary degeneration and promote regrowth of the facial nerve after axotomy or injury. PMID- 10527285 TI - FDG PET imaging of malignant paraganglioma of the neck. PMID- 10527286 TI - Universal modular glottiscope system: the evolution of a century of design and technique for direct laryngoscopy. AB - Since Kirstein introduced formal direct examination (autoscopy) of the glottis in 1895, a great number of laryngoscopes have been produced to view the vocal folds; however, none have had universal appeal. The primary goals for the designs have been to optimize exposure and to facilitate instrumentation of the glottis. An analysis of more than 50 laryngoscopes was done to assess key design characteristics that would ideally be present in a laryngoscope for optimally viewing the musculomembranous vocal folds. The Pro/Engineer and Pro/Mechanica computer programs were used to model the universal modular glottiscope. This new laryngoscope comprises a plurality of specially designed examining tubes that are bivalved proximally to improve utilization of hand instrumentation, and form a single tube distally to achieve internal distention of the supraglottal tissues. The distal lumen has an arcuate isosceles-triangular conformation to optimally expose the glottis. The base of the tube is detachable for the difficult intubation or the placement of bronchoscopes. The examining tubes vary in size and dimension to accommodate the diversity of human anatomy. The tubes are easily attachable to and detachable from an ergodynamically designed universal handle that can be joined to fulcrum laryngoscope holders or suspension gallows. The universal modular glottiscope evolved from the selective integration of optimal 20th-century design modifications of Kirstein's original autoscope. This new laryngoscope is ideally suited for phonomicrosurgery as well as for difficult intubation. PMID- 10527287 TI - Pathologic findings in a transplant donor. PMID- 10527288 TI - Magnesium and type 2 diabetes mellitus. PMID- 10527289 TI - Is integrative medicine the medicine of the future? A debate between Arnold S. Relman, MD, and Andrew Weil, MD. PMID- 10527290 TI - A community-based study of chronic fatigue syndrome. AB - BACKGROUND: Most previous estimates of the prevalence of chronic fatigue syndrome (CFS) have derived largely from treated populations, and have been biased by differential access to health care treatment linked with sex, ethnic identification, and socioeconomic status. OBJECTIVE: To assess the point prevalence of CFS in an ethnically diverse random community sample. DESIGN AND PARTICIPANTS: A sample of 28,673 adults in Chicago, Ill, was screened by telephone, and those with CFS-like symptoms were medically evaluated. MAIN OUTCOME MEASURES AND ANALYSES: Self-report questionnaires, psychiatric evaluations, and complete medical examinations with laboratory testing were used to diagnose patients with CFS. Univariate and multivariate statistical techniques were used to delineate the overall rate of CFS in this population, and its relative prevalence was subcategorized by sex, ethnic identification, age, and socioeconomic status. RESULTS: There was a 65.1% completion rate for the telephone interviews during the first phase of the study. Findings indicated that CFS occurs in about 0.42% (95% confidence interval, 0.29%-0.56%) of this random community-based sample. The highest levels of CFS were consistently found among women, minority groups, and persons with lower levels of education and occupational status. CONCLUSIONS: Chronic fatigue syndrome is a common chronic health condition, especially for women, occurring across ethnic groups. Earlier findings suggesting that CFS is a syndrome primarily affecting white, middle class patients were not supported by our findings. PMID- 10527291 TI - Duration of, and temporal trends (1994-1997) in, prehospital delay in patients with acute myocardial infarction: the second National Registry of Myocardial Infarction. AB - BACKGROUND: Extent of delay in seeking medical care in persons with acute myocardial infarction (AMI) is receiving increasing attention, given the time dependent benefits associated with early administration of coronary reperfusion therapy. OBJECTIVE: To examine recent data, and temporal trends therein, about duration of prehospital delay in a large (N = 364,131) cross-sectional sample of patients included in the second National Registry of Myocardial Infarction. METHODS: The medical records of patients hospitalized with AMI in 1624 US hospitals from June 1, 1994, to October 31, 1997, were reviewed for information about duration of prehospital delay. RESULTS: There was evidence of a slight decline in average delay times in patients hospitalized in 1997 (5.5 hours) compared with those hospitalized in 1994 (5.7 hours). Median delay times (2.1 hours) did not change. Approximately 20% of patients presented to the hospital within 1 hour of acute symptom onset, and slightly more than two thirds presented within 4 hours. Delay times were more prolonged for older patients, women, nonwhite patients, and patients with a history of diabetes or hypertension vs respective comparison groups. Patients in cardiogenic shock exhibited shorter delay times than less severely ill patients. Patients with previous AMI or who had undergone previous coronary angioplasty presented to the hospital with shorter delay times, as did individuals hospitalized in the Mountain and Pacific regions. CONCLUSIONS: These results provide insights into recent delay times and into groups at risk for prolonged delay. PMID- 10527292 TI - Serum and dietary magnesium and the risk for type 2 diabetes mellitus: the Atherosclerosis Risk in Communities Study. AB - BACKGROUND: Experimental studies in animals and cross-sectional studies in humans have suggested that low serum magnesium levels might lead to type 2 diabetes; however, this association has not been examined prospectively. METHODS: We assessed the risk for type 2 diabetes associated with low serum magnesium level and low dietary magnesium intake in a cohort of nondiabetic middle-aged adults (N = 12,128) from the Atherosclerosis Risk in Communities Study during 6 years of follow-up. Fasting serum magnesium level, categorized into 6 levels, and dietary magnesium intake, categorized into quartiles, were measured at the baseline examination. Incident type 2 diabetes was defined by self-report of physician diagnosis, use of diabetic medication, fasting glucose level of at least 7.0 mmol/L (126 mg/dL), or nonfasting glucose level of at least 11.1 mmol/L (200 mg/dL). RESULTS: Among white participants, a graded inverse relationship between serum magnesium levels and incident type 2 diabetes was observed. From the highest to the lowest serum magnesium levels, there was an approximate 2-fold increase in incidence rate (11.1, 12.2, 13.6, 12.8, 15.8, and 22.8 per 1000 person-years; P = .001). This graded association remained significant after simultaneous adjustment for potential confounders, including diuretic use. Compared with individuals with serum magnesium levels of 0.95 mmol/L (1.90 mEq/L) or greater, the adjusted relative odds of incident type 2 diabetes rose progressively across the following lower magnesium categories: 1.13 (95% CI, 0.79 1.61), 1.20 (95% CI, 0.86-1.68), 1.11 (95% CI, 0.80-1.56), 1.24 (95% CI, 0.86 1.78), and 1.76 (95% CI, 1.18-2.61) (for trend, P = .01). In contrast, little or no association was observed in black participants. No association was detected between dietary magnesium intake and the risk for incident type 2 diabetes in black or white participants. CONCLUSIONS: Among white participants, low serum magnesium level is a strong, independent predictor of incident type 2 diabetes. That low dietary magnesium intake does not confer risk for type 2 diabetes implies that compartmentalization and renal handling of magnesium may be important in the relationship between low serum magnesium levels and the risk for type 2 diabetes. PMID- 10527293 TI - Clinical and economic assessment of the omeprazole test in patients with symptoms suggestive of gastroesophageal reflux disease. AB - OBJECTIVE: To evaluate the diagnostic accuracy of a trial of a high-dose proton pump inhibitor (the omeprazole test) in detecting gastroesophageal reflux disease (GERD) in patients with heartburn symptoms. DESIGN: A randomized, double-blind, placebo-controlled, crossover trial. PATIENTS AND SETTING: Forty-three consecutive patients with symptoms suggestive of GERD were enrolled at a Veterans Affairs medical center. MAIN OUTCOME MEASURES: Symptom response to the omeprazole test vs placebo in GERD-positive and GERD-negative patients; sensitivity, specificity, and positive and negative predictive values of the omeprazole test; and cost per correct diagnosis achieved with the omeprazole test compared with traditional diagnostic strategies. RESULTS: Of 42 patients (98%) who completed the study, 35 (83%) were classified as GERD positive and 7 (17%) as GERD negative. Twenty-eight GERD-positive and 3 GERD-negative patients responded to the omeprazole test, providing a sensitivity of 80.0% (95% confidence interval, 66.7%-93.3%) and a specificity of 57.1% (95% confidence interval, 20.5%-93.8%). Economic analysis revealed that the omeprazole test saves $348 per average patient evaluated, and results in a 64% reduction in the number of upper endoscopies performed and a 53% reduction in the use of pH testing. CONCLUSIONS: The omeprazole test is sensitive and fairly specific for diagnosing GERD in patients with typical GERD symptoms. This strategy could result in significant cost savings and decreased use of invasive diagnostic tests. PMID- 10527294 TI - Tea flavonoids may protect against atherosclerosis: the Rotterdam Study. AB - BACKGROUND: Epidemiological studies have indicated a protective role of dietary flavonoids in cardiovascular disease, but evidence is still conflicting. Tea is the major dietary source for flavonoids in Western populations. We studied the association of tea intake with aortic atherosclerosis in a general population. METHODS: The present analysis formed part of the Rotterdam Study, a prospective study of men and women 55 years and older. Dietary intakes were assessed at baseline by a trained dietician who used a semiquantitative food frequency questionnaire. Calcified plaques in the abdominal aorta were radiographically detected after 2 to 3 years of follow-up. Aortic atherosclerosis was classified as "mild," "moderate," or "severe," according to the length of the calcified area (<1 cm, 1-5 cm, and >5 cm, respectively). The association of tea intake with severity of aortic atherosclerosis was studied in 3454 subjects who were free of cardiovascular disease at baseline. Data were analyzed by logistic regression, adjusting for age, sex, body mass index (calculated as weight in kilograms divided by the square of height in meters), smoking, education, and intake of alcohol, coffee, vitamin antioxidants, total fat, and total energy. RESULTS: Multivariable analyses showed a significant, inverse association of tea intake with severe aortic atherosclerosis. Odds ratios decreased from 0.54 (95% confidence interval [CI], 0.32-0.92) for drinking 125 to 250 mL (1-2 cups) of tea to 0.31 (CI, 0.16-0.59) for drinking more than 500 mL/d (4 cups per day). The associations were stronger in women than in men. The association of tea intake with mild and moderate atherosclerosis was not statistically significant. CONCLUSION: This study indicates a protective effect of tea drinking against ischemic heart disease. PMID- 10527295 TI - Lifetime health and economic consequences of obesity. AB - BACKGROUND: Obesity is an established risk factor for several chronic diseases. The lifetime health and economic consequences of obesity for individual patients have not been documented. OBJECTIVE: To estimate the lifetime health and economic consequences of obesity. METHODS: We developed a dynamic model of the relationship between body mass index and the risks and associated costs of 5 obesity-related diseases: hypertension, hypercholesterolemia, type 2 diabetes mellitus, coronary heart disease, and stroke. The model was estimated using data from the Third National Health and Nutrition Examination Survey, the Framingham Heart Study, and other secondary sources. We used this model to estimate (1) risks of hypertension, hypercholesterolemia, and type 2 diabetes mellitus at future ages; (2) lifetime risks of coronary heart disease and stroke; (3) life expectancy; and (4) expected lifetime medical care costs of these 5 diseases for men and women aged 35 to 64 years with body mass indexes of 22.5, 27.5, 32.5, and 37.5 kg/m2 (nonobese and mildly, moderately, and severely obese, respectively). RESULTS: Disease risks and costs increase substantially with increased body mass index. The risk of hypertension for moderately obese 45- to 54-year-old men, for example, is roughly 2-fold higher than for their nonobese peers (38.1% vs 17.7%), whereas the risk of type 2 diabetes mellitus is almost 3-fold higher (8.1% vs 3.0%). Lifetime risks of coronary heart disease and stroke are similarly elevated (41.8% vs 34.9% and 16.2% vs 13.9%, respectively), whereas life expectancy is reduced by 1 year (26.5 vs 27.5 years). Total discounted lifetime medical care costs for the treatment of these 5 diseases are estimated to differ by $10,000 ($29,600 vs $19,600). Similar results were obtained for women. CONCLUSIONS: The lifetime health and economic consequences of obesity are substantial and suggest that efforts to prevent or reduce this problem might yield significant benefits. PMID- 10527296 TI - Self-reported exercise tolerance and the risk of serious perioperative complications. AB - BACKGROUND: Impaired exercise tolerance during formal testing is predictive of perioperative complications. However, for most patients, formal exercise testing is not indicated, and exercise tolerance is assessed by history. OBJECTIVE: To determine the relationship between self-reported exercise tolerance and serious perioperative complications. METHODS: Our study group consisted of 600 consecutive outpatients referred to a medical consultation clinic at a tertiary care medical center for preoperative evaluation before undergoing 612 major noncardiac procedures. Patients were asked to estimate the number of blocks they could walk and flights of stairs they could climb without experiencing symptomatic limitation. Patients who could not walk 4 blocks and climb 2 flights of stairs were considered to have poor exercise tolerance. All patients were evaluated for the development of 26 serious complications that occurred during hospitalization. RESULTS: Patients reporting poor exercise tolerance had more perioperative complications (20.4% vs 10.4%; P<.001). Specifically, they had more myocardial ischemia (P = .02) and more cardiovascular (P = .04) and neurologic (P = .03) events. Poor exercise tolerance predicted risk for serious complications independent of all other patient characteristics, including age (adjusted odds ratio, 1.94; 95% confidence interval, 1.19-3.17). The likelihood of a serious complication occurring was inversely related to the number blocks that could be walked (P = .006) or flights of stairs that could be climbed (P = .01). Other patient characteristics predicting serious complications in multivariable regression analysis included history of congestive heart failure, dementia, Parkinson disease, and smoking greater than or equal to 20 pack-years. CONCLUSION: Self-reported exercise tolerance can be used to predict in-hospital perioperative risk, even when using relatively simple and familiar measures. PMID- 10527297 TI - Brief physician- and nurse practitioner-delivered counseling for high-risk drinkers: does it work? AB - BACKGROUND: There is a need for primary care providers to have brief effective methods to intervene with high-risk drinkers during a regular outpatient visit. OBJECTIVE: To determine whether brief physician- and nurse practitioner-delivered counseling intervention is efficacious as part of routine primary care in reducing alcohol consumption by high-risk drinkers. METHODS: Academic medical center-affiliated primary care practice sites were randomized to special intervention or to usual care. From a screened population of 9772 patients seeking routine medical care with their primary care providers, 530 high-risk drinkers were entered into the study. Special intervention included training providers in a brief (5- to 10-minute) patient-centered counseling intervention, and an office support system that screened patients, cued providers to intervene, and made patient education materials available. The primary outcome measures were change in alcohol use from baseline to 6 months as measured by weekly alcohol consumption and frequency of binge drinking episodes. RESULTS: Participants in the special intervention and usual care groups were similar on important background variables and potential confounders except that special intervention participants had significantly higher baseline levels of alcohol usage (P = .01). At 6-month follow-up, in the 91% of the cohort who provided follow-up information, alcohol consumption was significantly reduced when adjusted for age, sex, and baseline alcohol usage (special intervention, -5.8 drinks per week; usual care, -3.4 drinks per week; P = .001). CONCLUSIONS: This study provides evidence that screening and very brief (5- to 10-minute) advice and counseling delivered by a physician or nurse practitioner as part of routine primary care significantly reduces alcohol consumption by high-risk drinkers. PMID- 10527298 TI - Cross-classification of JNC VI blood pressure stages and risk groups in the Framingham Heart Study. AB - BACKGROUND: The recently published Sixth Report of the Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure (JNC VI) includes a classification of blood pressure stages and a new risk stratification component. Patients with high-normal blood pressure or hypertension are stratified into risk group A (no associated cardiovascular disease risk factors, no target organ damage or cardiovascular disease); group B (> or =1 associated cardiovascular disease risk factor excluding diabetes, no target organ damage or cardiovascular disease); or group C (diabetes or target organ damage or cardiovascular disease). OBJECTIVE: To examine the prevalence of risk groups and blood pressure stages in a community-based sample. METHODS: We evaluated 4962 subjects from the Framingham Heart Study and Framingham Offspring Study examined between 1990 and 1995. We cross-classified men and women separately according to their JNC VI blood pressure stages and risk groups. RESULTS: In the whole sample, 43.7% had optimal or normal blood pressure and 13.4% had high-normal blood pressure; 12.9% had stage 1 hypertension and 30.0% had stage 2 or greater hypertension or were receiving medication. As blood pressure stage increased, the proportion of subjects in group A decreased, whereas the proportion in group C increased. Among those with high-normal blood pressure or hypertension, only 2.4% (all women) were in risk group A, 59.3% were in group B, and 38.2% were in group C. In the high-normal or hypertensive group, 39.4% qualified for lifestyle modification as the initial intervention according to JNC VI recommendations, whereas 60.6% were eligible for initial drug therapy or were already receiving drug therapy. Nearly one third of high-normal subjects were in risk group C, in which early drug therapy may be needed. Among those in stage 1, only 4.0% were in group A, in which prolonged lifestyle modification is recommended. CONCLUSIONS: These results provide a foundation for estimating the number of individuals with hypertension who fall into different risk groups that require different treatment approaches. With nearly 50 million individuals with hypertension in the United States, there are important implications for clinicians and policymakers if JNC VI recommendations are widely adopted in clinical practice. PMID- 10527299 TI - Impediments to writing do-not-resuscitate orders. AB - BACKGROUND: Physicians are frequently unaware of their patients' desires regarding end-of-life care. Consequently, opportunities to implement do-not resuscitate (DNR) orders are often missed. OBJECTIVE: To determine the reasons attending physicians do not write DNR orders when patients face increased mortality. METHODS: Over 4 months, the medical records of all inpatients on the General Medicine Service were reviewed at the time of discharge to identify patients with conditions predicting increased mortality. These cases were presented to a 5-member panel who decided if a DNR order was indicated. Reasons for missing DNR orders were discussed with the attending physicians. RESULTS: Of 613 consecutive admissions, the panel identified 149 patients (24%) for whom DNR orders were indicated. In 88 (59%) of these, DNR orders were absent. The lack of a DNR order did not correlate with age (P = .95), sex (P = .61), or race (P = .80). The attending physicians' explanations for not writing DNR orders in these 88 cases included the belief that the patient was not in imminent danger of death (n = 49 [56%]), the belief that the primary physician should discuss DNR issues (n = 43 [49%]), and the lack of an appropriate opportunity to discuss end-of-life issues (n = 38 [43%]). In 11 (12%) of the 88 cases, patients or their families did not accept the recommendation for a DNR order. No physicians expressed concerns regarding the morality of DNR orders, discomfort discussing end-of-life issues, or the threat of litigation as reasons for not writing a DNR order. CONCLUSIONS: Limitations in the extent and depth of the physician-patient relationship appear to be the most frequent impediments to writing DNR orders in our institution. PMID- 10527301 TI - Fulminant toxic epidermal necrolysis induced by trovafloxacin. PMID- 10527300 TI - Life-threatening interactions between HIV-1 protease inhibitors and the illicit drugs MDMA and gamma-hydroxybutyrate. AB - Human immunodeficiency virus 1 (HIV-1) protease inhibitors have dramatically reduced the morbidity and mortality due to HIV-1 infection. However, most of these antiretrovirals are also potent inhibitors (and occasionally inducers) of hepatic and intestinal cytochrome P450 systems and, therefore, have the potential to alter the elimination of any substance that utilizes these metabolic pathways. We describe a patient infected with HIV-1 who was treated with ritonavir and saquinavir and then experienced a prolonged effect from a small dose of methylenedioxymetamphetamine (MDMA or ecstacy) and a nearly fatal reaction from a small dose of gamma-hydroxybutyrate (GHB). We also discuss the potential for HIV 1 protease inhibitors to alter the metabolism of other abusable prescribed and illicit substances. PMID- 10527302 TI - Smoking is an important component in the analysis of heart failure. PMID- 10527303 TI - Assessing cardiac risk assessment. PMID- 10527304 TI - Spontaneous reports of ticlopidine-associated thrombotic thrombocytopenic purpura from 2 Italian regions. PMID- 10527305 TI - Magnetic resonance imaging evaluation of muscle usage associated with three exercises for rotator cuff rehabilitation. AB - PURPOSE: Methods of determining muscle usage for exercises involving rotator cuff muscles are limited. Therefore, this investigation used magnetic resonance imaging (MRI) to evaluate the effect of three different exercises used for rehabilitation of the rotator cuff. METHODS: Five normal volunteer subjects (3 men, 2 women, mean age 31.4 yr) were studied. The exercises were scaption with internal rotation (SIR), military press (MP), and side-lying 45 degrees abduction (SLA). MR imaging was performed immediately before and after exercise using a "fast" spin echo STIR sequence and oblique coronal plane imaging. Changes in signal intensity pre- and post-exercise were measured at comparable section locations for the MR images of the supraspinatus, infraspinatus, teres minor, subscapularis, deltoid, and trapezius. RESULTS: The SLA showed the greatest increase in signal intensity in all the muscles (percent change, P < 0.01) except for the trapezius, which was used more by the MP and SIR. None of the exercises activated the teres minor (percent change, P = not significant). CONCLUSION: These findings have important implications in efficacy of physical rehabilitation of the rotator cuff and avoidance of subacromial impingement exercise motions. PMID- 10527306 TI - Recognizing and treating common cold-induced injury in outdoor sports. AB - We briefly review the physiology of cold exposure, the spectrum and prevention of common cold-induced injuries (especially in athletes participating in outdoor sports), and the potentially harmful side effects of localized cryotherapy. Severe cold affects all organ systems and especially the central nervous and cardiovascular systems; many biochemical reactions and pathways become distorted or slowed at low body core temperatures and can thus affect athletic performance. Low body shell temperature, too, can interfere with athletic ability by weakening and slowing muscle contractions, by delaying nerve conduction time, and by facilitating injury. Cold-induced injuries may be local or systemic, but they can usually be prevented by knowledge, good physical condition, appropriate nutrition and equipment, and avoidance of moisture. PMID- 10527308 TI - Weight control in wrestling: eating disorders or disordered eating? AB - PURPOSE: Several recent studies have pointed out that the weight loss techniques used by wrestlers to make weight are similar to the behavior of bulimics. The purpose of this study was to determine whether an increased risk of bulimia nervosa existed for a group of junior high and high school wrestlers. METHODS: Wrestlers (N = 85) completed the Eating Disorder Inventory (EDI) once during the season, and once during the off-season. A comparison group of nonwrestlers (N = 75) also completed the questionnaire. RESULTS: No significant differences were found between the number of in-season wrestlers and nonwrestlers classified as "at risk" for bulimia nervosa. Significant differences were revealed, however, between in-season wrestlers and nonwrestlers, and between in-season wrestlers and off-season wrestlers, on the Drive for Thinness subscale. In both cases, significantly more in-season wrestlers scored above the "at risk" cutoff on the subscale. CONCLUSIONS: These results indicate that although in-season wrestlers are more weight conscious than nonwrestlers, these feelings and attitudes are transient. All subjects classified as "at risk" also participated in an interview which followed the format of the Eating Disorder Examination. Interviews with in season wrestlers revealed that their concerns with weight were due entirely to the demands of wrestling, and did not meet the severity level required for a diagnosis of bulimia nervosa. PMID- 10527307 TI - Etiologic factors associated with Achilles tendinitis in runners. AB - PURPOSE: The purpose of this study was to determine whether relationships exist between selected training, anthropometric, isokinetic muscular strength, and endurance, ground reaction force, and rearfoot movement variables in runners afflicted with Achilles tendinitis. METHODS: Specifically, we examined differences in selected measures between a noninjured cohort of runners (N = 58) and a cohort of injured runners with Achilles tendinitis (N = 31). Isokinetic, kinetic, and kinematic measures were collected using a Cybex II+ isokinetic dynamometer (Medway, MA), AMTI force plate (500 Hz), and Motion Analysis high speed videography (200 Hz), respectively. Separate discriminant function analyses were performed on each of the five sets of variables to identify the factors that best discriminate between the injured and control groups. RESULTS: Years running, training pace, stretching habits (injured runners were less likely to incorporate stretching into their training routine), touchdown angle, plantar flexion peak torque at 180 degrees x s(-1) and arch index were found to be significant discriminators. CONCLUSION: A combined discriminant analysis using the above mentioned significant variables revealed that plantar flexion peak torque, touchdown angle, and years running were the strongest discriminators between runners afflicted with Achilles tendinitis and runners who had no history of overuse injury. PMID- 10527309 TI - The effect of prolonged exercise on lipid peroxidation in eumenorrheic female runners. AB - PURPOSE: Recently a protective role has been demonstrated for estrogens as free radical scavengers. In this study, lipid peroxidation was evaluated in eumenorrheic runners before and after participation in a half-marathon. METHODS: Seven female runners who participated in regular training (average 25 miles x wk( 1) and reported regular menses (12/yr) served as subjects. Subjects were all in a low estrogen phase of their menstrual cycle as confirmed by menstrual record and plasma estradiol level (42.71 +/- 21.65 pg x mL(-1). Low density lipoprotein oxidation (formation of conjugated dienes) was determined 2 h prerace and 5 min after subject's completion of the race. RESULTS: Results showed a significant increase in lag phase time of conjugated dienes after prolonged exercise (28.43 +/- 4.89 min vs postrace 35.21 +/- 4.32 min, P < 0.05). No correlation between mean levels of estradiol and mean lipid peroxidation levels at rest, 5 min after exercise, or difference (prepost) was observed. CONCLUSION: Prolonged endurance exercise does not appear to increase potential for lipid peroxidation in trained eumenorrheic runners during a low estrogen phase of the menstrual cycle. PMID- 10527310 TI - Shoulder proprioception: latent muscle reaction times. AB - PURPOSE: The purpose of this study was to identify electromyographic (EMG) differences in the latent muscle reaction timing (LMRT) of the rotator cuff between trained overhead throwers and control subjects in response to sudden internal rotation perturbation (P < or = 0.05). METHODS: Subjects included 15 trained overhead throwers (male intercollegiate baseball players) and 15 untrained subjects (males not active in competitive throwing sports). Subjects were tested while seated, with their dominant glenohumeral joint positioned in 90 degrees abduction/external rotation (scapular plane), their elbow flexed to 90 degrees, and their forearm placed in the perturbation device. Rotator cuff LMRT was assessed as they tried to decelerate a variably timed, sudden internal rotation force. EMG sampling (2000 Hz, 2-s duration) began immediately before perturbation. RESULTS: Trained throwers had slower infraspinatus (P = 0.011) and teres minor (P = 0.024) LMRT and decreased supraspinatus (P = 0.001) and posterior deltoid (P = 0.0001) muscle activation duration compared with control subjects. CONCLUSIONS: These results suggest that the rotator cuff muscles of trained throwers may be downregulated in response to sudden internal rotation perturbation. Although these adaptations would enable greater internal rotation velocities during overhead throwing, they may also contribute to glenohumeral joint pathology. The identification of changes in rotator cuff LMRT in response to sudden internal rotation perturbation suggests an area of acquired neuromuscular imbalance warranting consideration by those involved in the rehabilitation and conditioning of the overhead throwing athlete. PMID- 10527311 TI - Quantitative MR measures of three-dimensional patellar kinematics as a research and diagnostic tool. AB - PURPOSE: A three-dimensional (3D) study of normal patellar-femoral-tibial (knee) joint kinematics was performed using Cine Phase Contrast Magnetic resonance imaging (Cine-PC MRI) to determine the utility of this technique as a diagnostic tool in defining alterations in patellar tracking. METHODS: Cine-PC MRI was originally developed to measure heart motion and blood flow and has now been adapted to the study of the musculoskeletal system. Thus, for the first time knee joint kinematics can be studied three-dimensionally, noninvasively, and in vivo during dynamic volitional leg extensions under load. Cine-PC MRI provides one anatomic and three orthogonal velocity images (vx, vy, and vz) for each time frame within the motion cycle. Bone displacements are calculated using integration and are then converted into both 3D orientation angles and 2D clinical angles. RESULTS: The 3D patellar tilt and 2D clinical patellar tilt angle were nearly identical, even though these two angles have distinct mathematical definitions. The precision of the 2D clinical patellar tilt angle (N = 3) was approximately 2.4 degrees. CONCLUSIONS: Since the overall subject (N = 18) variability for clinical patellar tilt angle and medial/lateral patellar displacement was low (SD = 2.9 degrees and 3.3 mm, respectively), Cine-PC MRI could prove to be a valuable tool in studying subtle changes in patellar tracking. PMID- 10527312 TI - Plasma-electrolytes in natives to hypoxia after marathon races at different altitudes. AB - PURPOSE: It is well known that altitude natives differ from sea level natives in aspects of fluid and electrolyte homeostasis. METHODS: To evaluate exercise and environmental influences on the electrolyte and water status in hypoxia adapted subjects, we investigated 11 well-trained marathon runners (33.7 +/- 0.7 yr, 60.5 +/- 1.9 kg), native to an altitude above 2600 m, before and after two marathon races. One competition was held at moderate altitude (AM, 2650 m, 14 degrees C, 55% RH, running time 3 h 6 min +/- 22 min) and another under tropical conditions (HM, 470 m, 28 degrees C, 70% RH, running time 2 h 54 min +/- 30 min). Blood samples were taken 3 d before, immediately after, 1 h after, and 24 h after the races. RESULTS: The loss in body fluid was calculated to be 2.15 L during AM and 5.05 L during HM, respectively. It was compensated mostly by ingested fluids without electrolyte content and by metabolically produced water, which led to hyponatremia during AM (plasma [Na+] from 144.3 +/- 0.7 to 131.7 +/- 2.1 mmol x L(-1)). Severe dehydration without significant changes in plasma [Na+] could be detected after HM. Serum antidiuretic hormone concentrations and serum aldosterone concentrations significantly increased during both races and remained at a high level for at least 1h after both competitions. Serum atrial natriuretic peptide (ANP) concentrations were at a high level at rest, increasing during HM, and decreasing during AM. CONCLUSION: Under tropical conditions, we found a severe state of dehydration characterized by an extended ANP-response, which was not prevented by water intake during the race. Under hypoxic conditions, however, we found that hyponatremia had developed. This can be partly explained by pure water intake and metabolically produced water, and also, possibly, by a special hypoxia-induced effect. PMID- 10527313 TI - Short-term effects of marathon running: no evidence of cardiac dysfunction. AB - PURPOSE: The purpose of this study was to analyze the short-term effects of a marathon race (Madrid Marathon) on both markers of cardiac damage and echocardiographic parameters in a group of 22 runners (17 male and 5 female; 34 +/- 5 yr; VO2max: 55.7 +/- 9.1 mL x kg(-1) x min(-1) with a wide range of fitness levels. METHODS: Venous blood samples were collected from each subject 48 h before the race, at race finish, and 6, 24, and 48 h postexercise for the determination of myoglobin, total creatine kinase catalytic activity (total CK), mass concentration of creatine kinase isoenzyme MB (CK-MB mass), and cardiac isoforms of troponin T and I (TnT-c and TnI-c, respectively). In addition, echocardiographic parameters (M-mode two-dimensional and Doppler analysis) indicative of both left ventricular (LV) systolic and diastolic function were obtained three times from each runner: 2-5 d before the race, at race finish, and 24-36 h after exercise. RESULTS: Except in one subject, levels of TnT-c and TnI-c were within normal limits (<0.1 ng x mL(-1)) in all the samples collected before or after the race. Overall LV systolic function was not altered by marathon running. Finally, LV diastolic function was transiently altered after the race since the ratio between peak early and late transmitral filling velocities (E/A) was significantly reduced at race finish (P < 0.01) and returned to resting levels after 24-36 h. CONCLUSIONS: Our findings suggest that marathon running does not adversely affect the hearts of healthy individuals independently from their training status. PMID- 10527314 TI - Ventilatory responses during experimental cycle-run transition in triathletes. AB - PURPOSE AND METHODS: To determine the effects of cycling on a subsequent triathlon run, nine male triathletes underwent four successive laboratory trials: 1) an incremental treadmill test, 2) an incremental cycle test, 3) 30 min of cycling followed by 5 km of running (C-R), and 4) 30 min of running followed by 5 km of running (R-R). Before and 10 min after the third and fourth trials, the triathletes underwent pulmonary function testing including spirometry and diffusing capacity testing for carbon monoxide (DL(CO)). During the C-R and R-R trials, arterialized blood samples were obtained to measure arterial oxygen pressure (PaO2). During all trials, ventilatory data were collected every minute using an automated breath-by-breath system. RESULTS: The results showed that 1) the oxygen uptake (VO2) observed during subsequent running was similar for the C R and R-R trials; 2) the ventilatory response (VE) during the first 8 min of subsequent running was significantly greater in the C-R than in R-R trial (P < 0.05); 3) only the C-R trial induced a significant increase (P < 0.05) in residual volume (RV), functional residual capacity (FRC), and the ratio of residual volume to total lung capacity (RV/TLC); and 4) although a significant decrease (P < 0.05) in DL(CO) was noted after C-R, no difference between the two exercise trials was found for the maximal drop in PaO2. CONCLUSIONS: We concluded that 1) the C-R trial induced specific alterations in pulmonary function that may be associated with respiratory muscle fatigue and/or exercise-induced hypoxemia, and 2) the greater VE observed during the first minute of running after cycling was due to the specificity of cycling. This reinforces the necessity for triathletes to practice multi-trial training to stimulate the physiological responses experienced during the swim-cycle and the cycle-run transitions. PMID- 10527315 TI - Ventilatory response to erect and supine exercise. AB - PURPOSE: To test the hypothesis that altering the ventilation-perfusion ratio of the lung by changing the body position from erect to supine would alter the ventilatory response to exercise as described by the slope of the relationship between minute ventilation and carbon dioxide production. METHODS: Ten normal subjects volunteers (5 female, 5 male: average age 22 yr; range 19-25 yr; height (SD) 173.5 (3.8) cm; weight 68.0 (3.3) kg) performed in random order erect and supine incremental cycle exercise with metabolic gas exchange measurements to determine peak oxygen consumption (VO2) and the slope of the relation between ventilation and carbon dioxide production (VE/VCO2 slope). RESULTS: Subjects reached a higher peak VO2 when erect (mean (SEM))(39.2 (2.4) vs 35.7 (2.0); P < 0.05). Heart rate, ventilation, and VO2 were higher at each stage in the erect position. The respiratory exchange ratio was the same in each position at matched workloads and at peak exercise. The VE/VCO2 slope was unchanged (27.8 (2.2) erect vs 27.7 (1.9) erect). CONCLUSION: Cycle exercise in the erect position is associated with an increase in exercise capacity compared with supine exercise but with no associated changes in ventilatory response to carbon dioxide production. PMID- 10527316 TI - Evidence for restricted muscle blood flow during speed skating. AB - INTRODUCTION: We have previously hypothesized restricted muscle blood flow during speed skating, secondary to the high intramuscular forces intrinsic to the unique posture assumed by speed skaters and to the prolonged duty cycle of the skating stroke. METHODS: To test this hypothesis, we studied speed skaters (N = 10) during submaximal and maximal cycling and in-line skating, in both low (knee angle = 107 degrees) and high (knee angle = 112 degrees) skating positions (CE vs SkL vs SkH). Supportive experiments evaluated muscle desaturation and lactate accumulation during on-ice speed skating and muscle desaturation during static exercise at different joint positions. RESULTS: Consistent with the hypothesis were reductions during skating in VO2peak (4.28 vs 3.83 vs 4.26 L x min(-1)), the VO2 at 4 mmol x L(-1) blood lactate (3.38 vs 1.93 vs 3.31 L x min(-1)), and cardiac output during maximal exercise (33.2 vs 25.3 vs 25.6 L x min(-1)). The reduction in maximal cardiac output was not attributable to differences in HRmax (197 vs 192 vs 193 b x min(-1)) but to a reduction in SVmax (172 vs 135 vs 134 mL x beat(-1)). The reduction in SV appeared to be related to an increased calculated systemic vascular resistance (354 vs 483 vs 453 dynes x s(-1) x cm( 1)). During maximal skating there was also a greater % O2 desaturation of the vastus lateralis based on near infrared spectrophotometry (50.3 vs 74.9 vs 60.4% of maximal desaturation during cuff ischemia). The results were supported by greater desaturation with smaller knee angles during static exercise and by greater desaturation and accelerated blood lactate accumulation during on-ice speed skating in the low vs high position. The results of this study support the hypothesis that physiological responses during speed skating are dominated by restriction of blood flow, attributable either to high intramuscular forces, the long duty cycle of the skating stroke, or both. PMID- 10527317 TI - Simplified deceleration method for assessment of resistive forces in cycling. AB - PURPOSE: The purpose of this study was to develop and test a simplified deceleration technique for measurement of aerodynamic and rolling resistances in cycling. METHODS: Coast-down tests were performed in level hallways with an experienced cyclist as the rider. Average initial velocities were 2.5-12.8 m x s( 1)) The deceleration technique was simplified by the use on only three switches and a derivation that did not require an assumption that deceleration is constant. The effective frontal area (AC(D)) and coefficient of rolling resistance (CR) were then calculated through a derivation from the equation for resistive forces opposing motion. Method reproducibility was tested by comparison of results for four tests of 30 trials under identical conditions. Method sensitivity was tested by performing 30 trials with three different rider head positions and four different transported mass conditions. RESULTS: Analysis of variance revealed that there were no differences among the results in the reproducibility study for either AC(D) or C(R). Furthermore, the reproducibility tests revealed mean errors of only 0.66% and 0.70% for AC(D) and CR, respectively. ANOVA identified a significant increase (P < 0.001) in rolling resistance with external loading and a significant effect (P < 0.001) of head position on AC(D). Mean (+/-SD) values for AC(D) and C(R) from tests in a racing aeroposture with the head up, the head in line with the trunk, and the head in an intermediate position were 0.304 +/- 0.011, 0.268 +/- 0.010, and 0.262 +/- 0.013 m2, respectively. C(R) averaged 0.00368 in the three head positions. CONCLUSIONS: The findings indicate that this simplified deceleration technique is satisfactorily reproducible and sensitive for measurement of aerodynamic and rolling resistances in cycling. PMID- 10527318 TI - Architectural characteristics of muscle in black and white college football players. AB - PURPOSE: The purpose of this study was to determine whether architectural characteristics of skeletal muscle differ by race. METHODS: Skeletal muscle architectural characteristics and body composition were studied in 13 black and 31 white male college football players. Fat-free mass (FFM) and percentage body fat (% fat) were determined by hydrostatic weighing technique. Muscle thickness (MTH) was measured by B-mode ultrasound at 13 anatomical sites. Isolated MTH and muscle pennation angle (PANG) of the triceps long head, vastus lateralis, and gastrocnemius medialis muscles were measured by ultrasound, and fascicle length was estimated. RESULTS: There were no significant differences between blacks and whites in isolated MTH, PANG, and fascicle length in the triceps long head, vastus lateralis, and gastrocnemius medialis muscles. On average, % fat and FFM of black and white football players were 18.8 +/- 4.6% and 17.2 +/- 5.6% for % fat, and 89.9 +/- 15.6 kg and 89.1 +/- 10.4 kg for FFM, respectively. Blacks had a significantly greater, 30%-quadriceps (P < 0.05), 50%-hamstrings (P < 0.05), biceps (P < 0.01), and abdomen (P < 0.01) MTH than those of whites. Standing height and body weight were similar between blacks and whites, but the ratio of leg length to standing height was significantly greater in blacks compared with whites. CONCLUSIONS: It appears that although there may be race differences in anatomical stature, muscle architecture is likely independent of race. PMID- 10527319 TI - Physiological responses to upper body exercise on an arm and a modified leg ergometer. AB - PURPOSE: The present study was undertaken to compare cardiorespiratory, metabolic, and perceptual responses to upper body exercise on an arm ergometer (AE) and a modified leg ergometer (LE). METHODS: Seventeen male and seven female subjects completed two experimental trials. During each trial, the subjects performed two successive 8-min steady-state arm crank exercises on either an AE or an LE. The crank frequency was kept constant at 50 rev x min(-1) during all exercise bouts. The two power outputs selected were 50 and 75 W for male subjects and 25 and 50 W for female subjects. To achieve these power outputs, the brake resistance was set at 1, 2, and 3 kg at a power output of 25, 50, and 75 W, respectively, for the AE and 0.5, 1, and 1.5 kg at a power output of 25, 50, and 75 W, respectively, for the LE. Oxygen uptake (VO2), heart rate (HR), respiratory exchange ratio (RER), expired ventilation (VE), gross efficiency (GE), and ratings of perceived exertion (RPE) were measured every minute during the last 2 min of each exercise bout. RESULTS: In male subjects, VO2, HR, RER, VE, and RPE were higher (P < 0.05), whereas GE was lower (P < 0.05) during arm crank exercise on an AE than an LE at power outputs of 50 and 70 W. In female subjects, similar differences in these variables between the two ergometers were also observed when exercise was performed at 50 W. However, VO2, RER, VE, and GE did not differ between the two ergometers when exercise was performed at 25 W. CONCLUSIONS: Upper body exercise elicits greater cardiorespiratory, metabolic, and perceptual responses on an AE than an LE at the same power output when power output is computed according to the manufacturer's instructions. PMID- 10527320 TI - Effects of a proposed challenge on effort sense and cardiorespiratory responses during exercise. AB - PURPOSE: Highly trained endurance athletes train and race at relatively high intensities and are often confronted with challenges throughout a running event. The purpose of this study was to examine the effects of the anticipation of a proposed challenge on effort sense, heart rate (HR), ventilation ([dotVE), and ventilatory equivalent VE/VO2), a measure of ventilatory efficiency. METHODS: Highly trained endurance athletes (VO2max = 68.46 +/- 1.47 mL x kg(-1) x min(-1) ran two sessions at approximately 75% of VO2max for 35 min in a control condition and a proposed challenge condition. During the control condition, the subjects ran on a treadmill while simultaneously viewing a video depicting a runner exercising at 75% of VO2max and were told the run would continue at a speed that elicited 75% of VO2max. During the proposed challenge condition, subjects completed the same exercise protocol but viewed a video of a struggling runner and were told that the treadmill speed would be increased to "an extremely difficult" 95% of VO2max matching the intensity of the runner on the video. However, after data assessment at 17 min, subjects were told that the treadmill was malfunctioning and the treadmill speed could not be altered. The same intensity was maintained in both conditions. RPE, HR, VE, and VE/VO2 were assessed during the treadmill runs at 10, 17, 25, and 35 min. RESULTS: The effects of the manipulation were represented by a significant increase in state anxiety immediately following the video proposing the 95% challenge. RPE, HR, and VE increased similarly under both conditions, while VE/VO2 did not change. CONCLUSION: These findings suggest that for highly trained endurance athletes, anticipation of proposed challenge during running does not influence cardiorespiratory responses; thus these athletes demonstrate a "physiologically toughened" response. PMID- 10527321 TI - Bioimpedance assessment of hypohydration. AB - PURPOSE: This study examined the utility of bioimpedance spectroscopy (BIS) for assessing total body water (TBW) changes associated with moderate (6-7% TBW), hypertonic (HH), and isotonic (IH) hypohydration. METHODS: The TBW of nine men was measured using BIS (TBWBIS) when euhydrated (EU) and during HH and IH. These measurements were compared with TBW measurements obtained using isotope dilution (deuterium oxide; TBWD20) during EU, and the estimated TBWD20 during hypohydration calculated from body weight change. RESULTS: Body weight loss was similar (P > 0.05) for HH (3.4 +/- 0.7 kg) and IH (2.9 +/- 0.7 kg). Plasma osmolality was higher (P < 0.05) on HH (292 +/- 4 mOsmol x kg(-1)) than EU (280 +/- 4 mOsmol x kg(-1)) and IH (284 +/- 3 mOsmol x kg(-1)), and higher (P < 0.05) during IH than EU. The measurements reflected a similar decrease (P < 0.05) in TBW during HH (TBWD20: 45.4 +/- 7.3 L, TBWBIS: 42.2 +/- 5.1 L) and IH (TBWD20: 45.8 +/- 7.5 L, TBWBIS: 42.0 +/- 4.9 L), compared with EU (TBWD20: 48.8 +/- 7.5 L, TBWBIS: 44.3 +/- 5.0 L), but TBWD20 was consistently higher (3.9 +/- 4.0 L, P < 0.05) than TBWBIS during all tests. TBWD20 and TBWBIS were correlated (P < 0.05) during EU (r = 0.87), HH (r = 0.84), and IH (r = 0.84). The change in TBW from EU during HH was greater (P < 0.05) for TBWD20 (3.5 +/- 0.8 L) than TBWBIS (2.1 +/- 0.9 L), but during IH the change in TBW reflected by TBWD20 (3.0 +/- 0.6 L) and TBWBIS (2.3 +/- 0.8 L) did not differ. The change in TBWD20 and TBWBIS between EU and hypohydration was correlated for HH (r = 0.77, P < 0.05), but not IH. CONCLUSION: These findings indicate that BIS is sufficiently sensitive to detect moderate hypohydration; however, the resolution of this technique diminished with isotonic fluid loss. PMID- 10527322 TI - Physiological effects of constant versus variable power during endurance cycling. AB - PURPOSE: Previous theoretical research found that varying power slightly to counter external conditions may result in improved performance during cycling time trialing, but it is not known whether such power variations result in added physiological stress. Thus, the purpose of this study was to determine whether variable power (VP) cycling produced greater physiological stress than constant power (CP) cycling of the same mean intensity. METHODS: Eight trained male cyclists (age 28 +/- 2 yr, mass 74.4 +/- 2.3 kg, VO2max 4.24 +/- 0.13 L x min( 1), weekly training 277 +/- 44 km) performed three 1-h ergometer trials. The first trial was performed at a self-paced maximal effort. The mean power from that trial was used to determine the power for the CP trial (constant effort at mean power) and the VP trial (alternating +/- 5% of mean power every 5 min). RESULTS: No differences were found between the CP and VP trials in mean VO2 (CP 3.33 +/- 0.11 L x min(-1), VP 3.26 +/- 0.12 L x min(-1)), mean heart rate (CP 158 +/- 3 min(-1), VP 159 +/- 3 min(-1)), mean blood lactate concentration (CP 4.2 +/ 0.7 mM, VP 4.3 +/- 0.7 mM), or mean RPE (CP 13.9 +/- 0.4, VP 14.1 +/- 0.4). CONCLUSION: Therefore, during a strenuous 1-h effort (78% of VO2max), subjects experienced no additional physiological stress by varying power +/- 5% compared with that during a constant power effort. PMID- 10527323 TI - The power output/heart rate relationship in cycling: test standardization and repeatability. AB - PURPOSE: The purpose of this study was to update and standardize the test for determining the power output/heart rate (PO/HR) relationship in cycling. METHODS: The current protocol was developed in the laboratory using a wind-load cycling simulator. Five hundred incremental tests were carried out by 290 male cyclists during a 2-yr period (1995-1997). The subjects' own bicycles, equipped with a standard crankset with a built-in power measuring system, were used for testing. The test protocol consisted of time-based increments in cadence that were uniform up to submaximal speeds and progressively greater in the final phase. RESULTS: The PO/HR relationship obtained was linear at low to submaximal PO and curvilinear from submaximal to maximal PO. A method was developed for the mathematical identification of the point of transition from the linear to the curvilinear phase (deflection point or heart rate break point). In 484 of the 500 tests performed, the deflection was independent of the final acceleration (PO at deflection 318.4 +/- 42.4 W, PO at final acceleration 351.6 +/- 43.2 W, P < 0.001), whereas in 16 tests the deflection and the start of the final acceleration coincided. To evaluate test repeatability and precision, 15 subjects repeated the test twice within a few days. No significant differences were found for the heart rate at deflection, power output at deflection, or slope of the linear part of the PO/HR relationship obtained in the two tests. CONCLUSION: It is concluded that the deflection point obtained by determining the PO/HR relationship on a wind-load simulator is not an artifact dependent on the incremental test protocol but rather a repeatable physiological phenomenon. PMID- 10527324 TI - Comparison of calcaneal ultrasound and DXA in young women. AB - PURPOSE: The purpose of the study was to assess quantitative ultrasound (QUS) parameters in collegiate female gymnasts, a population whose training incorporates high-impact loading, which is particularly osteogenic, and to determine the discriminative capacity of this relatively new radiation-free technique compared with bone densitometry in a young healthy population. METHODS: We studied 19 collegiate gymnasts and 23 healthy controls undergoing regular weight-bearing activity, matched for age (gymnasts 19.2 +/- 1.2, controls 19.9 +/ 1.6 yr) and body weight (gymnasts 56.7 +/- 3.7, controls 57.7 +/- 7.8 kg). QUS parameters of the calcaneus (broadband ultrasound attenuation (BUA), bone velocity (BV), and speed of sound (SOS)) were measured by a Walker Sonix UBA 575+. Bone mineral density (BMD; g x cm(-2)) of the lumbar spine, hip (femoral neck, trochanter, Ward's triangle) and whole body was assessed by dual energy x ray absorptiometry (DXA, Hologic QDR 1000/W). Data analysis included unpaired two tailed Student's t-tests, analysis of variance, Pearson product-moment, and Spearman rank-order correlations. RESULTS: Regional and whole body BMD of gymnasts was greater than controls (P < 0.001), with the difference being 7-28%. Average QUS parameters of the right and left calcaneus were also higher (P < 0.001) in the gymnasts. BUA, BV, and SOS were significantly (P < 0.001) correlated to each bone site with r = 0.54-0.79. Analysis of receiver operating characteristic (ROC) curves indicated no significant difference in sensitivity and specificity for QUS and DXA measures. CONCLUSIONS: These results indicate that QUS parameters of the calcaneus are higher in young women gymnasts compared to individuals who undergo regular weight-bearing activity and that QUS parameters are able to discriminate between these two groups in a similar manner as does regional and whole body BMD. PMID- 10527325 TI - Surgery, passion, and the medical student. PMID- 10527326 TI - Severe local hypothermia from laparoscopic gas evaporative jet cooling: a mechanism to explain clinical observations. AB - BACKGROUND AND OBJECTIVES: Explanations for laparoscopic-induced hypothermia fail to explain clinical observations. It is possible that water evaporation occurs from the jet stream of gas inflation resulting in tissue surface super-cooling leading to tissue damage and drying. METHODS: Theoretical calculations based on thermal conductivity, mass transfer effects and heat flux considerations correlated closely with synthetic and tissue experiments. Thermocouple measurements at a rate of 15 data points per second were performed. RESULTS: Cooling rates of 10 to 25 degrees centigrade per second for high flow rates were found based on gas flow rate and effective size of gas delivery site. These rapid temperature drops extended beyond a 2 cm2 diameter. CONCLUSIONS: Evaporative cooling accounts for significant hypothermia. The cooling is dependent on the lack of water vapor in the gases currently used during laparoscopy. Cooling rates are independent of height from tissue and geometry of delivery port. Heating and hydrating the gas to a physiologic condition eliminates hypothermia and tissue dessication. PMID- 10527327 TI - Laparoscopic management of ovarian dermoid cysts: ten years' experience. AB - OBJECTIVE: To determine the safety and efficacy of laparoscopic management of ovarian dermoid cysts based upon our ten years' experience. METHODS: Charts of 81 patients who underwent laparoscopic removal of dermoid cysts since March 1988 at Stanford University Medical Center or the Center for Special Pelvic Surgery in Atlanta were reviewed retrospectively. RESULTS: Ninety-three dermoid cysts with a mean diameter of 4.5 cm were removed in 81 patients. Operative techniques used were cystectomy for 70 cysts, salpingooophorectomy for 14, and 9 salpingo oophorectomy with hysterectomy. Fifty-three cysts were treated via enucleation followed by cystectomy or salpingo-oophorectomy and removal through a trocar sleeve. Twenty-two were treated via enucleation and removal within an impermeable sack. Nine were treated via enucleation and removal by posterior colpotomy. Nine were removed via colpotomy following hysterectomy. We had a total of 39 spillages. Spillage rates varied with removal method: 32 (62%) for trocar removal without an endobag, 3 (13.6%) for removal within an endobag, and 4 (40%) with colpotomy removal. No spillage occurred for the nine patients who had a colpotomy done for hysterectomy. Mean hospital stay after surgery was 0.98 days, and there were no intraoperative complications. In one case, there was a postoperative complication of an incisional infection in the umbilicus. CONCLUSION: Including this and 13 other studies, review of the literature reveals a 0.2% incidence of chemical peritonitis following laparoscopic removal of dermoid cysts. Thus, we conclude that laparoscopic management of dermoid cysts is a safe and beneficial method in selected patients when performed by an experienced laparoscopic surgeon. PMID- 10527328 TI - Total laparoscopic hysterectomy using the harmonic scalpel. AB - Total laparoscopic hysterectomy (TLH) is the complete hysterectomy including transection of the uterine vessels and opening/closure of the vaginal vault performed laparoscopically. This procedure can be performed as an alternative to total abdominal hysterectomy in many cases. We previously found use of the harmonic scalpel to be extremely helpful in performing laparoscopically assisted vaginal hysterectomies. In this series, the harmonic scalpel was used to facilitate performing TLH. Our experience has shown this can be performed without major complications in a cost-effective manner. PMID- 10527329 TI - Non-traumatic acute abdomen: videolaparoscopic approach. AB - BACKGROUND AND OBJECTIVE: Although videolaparoscopy has been considered a safe method for many elective procedures, its use in traumatic and non-traumatic acute abdomen needs to be evaluated. The aim of this article is to evaluate the role of videolaparoscopy in non-traumatic acute abdomen as a method of diagnosis and treatment. METHODS: Between January 1992 and December 1996, 462 patients' charts were reviewed, retrospectively. Patients were admitted to the emergency room of Sao Rafael Hospital with symptoms of non-traumatic acute abdomen. Routine investigation of abdominal pain was performed in all patients, followed by videolaparoscopy. The laparoscopic procedures were done with four main purposes: diagnosis (ie, enteritis); diagnosis and treatment (ie, appendicitis); treatment only, when the diagnosis was known (ie, acute cholecystitis); and in cases where the conversion to conventional laparotomy was necessary, indicating the best incision. RESULTS: The vast majority of patients had inflammatory causes of acute abdomen (82.03%); others causes were hemoperitoneum (11.03%), bowel obstruction (3.25%), perforation of a hollow viscera (1.74%), vascular occlusion (1.3%), and negative laparoscopy (0.65%). CONCLUSIONS: This study shows that laparotomy was necessary in only 7.14% of the patients. The videolaparoscopic approach was used for diagnosis (99.35%) and treatment (92.86%) of patients with acute abdomen. PMID- 10527330 TI - Laparoscopic Roux-en-Y gastric bypass for morbid obesity. AB - Surgery is currently the only effective treatment for morbid obesity. The two most commonly accepted operations are the Roux-en-Y gastric bypass and vertical banded gastroplasty. Although multiple authors have reported on a laparoscopic approach to gastric banding, the Roux-en-Y gastric bypass is a complex operation to be replicated using laparoscopic techniques. In this article, we describe our technique of the Roux-en-Y gastric bypass using a laparoscopic approach in four cases. PMID- 10527331 TI - Esophageal manometry and 24-hour pH monitoring to evaluate laparoscopic Lind fundoplication in gastroesophageal reflux disease. AB - Laparoscopic and thoracoscopic techniques have provided a new dimension in the correction of functional disorders of the esophagus. Therapeutic success, however, depends on the confirmation of esophageal disease as a cause of the symptoms, on understanding the basic cause of dysfunction and on identifying the surgical patient. This study is a retrospective study of patients submitted to surgery using the Lind procedure for gastroesophageal reflux disease (GERD). The purpose of this study is to establish the value of the routine use of esophageal manometry and 24-hour pH monitoring in order to select patients and perform pre and postoperative functional evaluation. Forty-one patients (68.3%) had a hypotonic lower esophageal sphincter. The average pressure was 9.2 mm Hg preoperatively and 15.2 mm Hg postoperatively, with an increase of 6.0 mm Hg. This increase was 8.8 mm Hg in hypotonics and 4.3 mm Hg in the normotonics. There was a certain degree of hypomotility of the esophageal body in 14 patients (23.3%) and, of this group, 4 (28.5%) improved postoperatively. Pathological acid reflux was found in 51 cases (85.0%) by pH monitoring. The mean of the preoperative DeMeester score was 31.4, later dropping to 3.2. Esophageal manometry and 24-hour pH monitoring are effective methods for revealing the level of functional modification established by anti-reflux surgery and for helping to objectively perform the selection. PMID- 10527332 TI - Peptic ulcer disease and thoracoscopic left truncal vagotomy. AB - BACKGROUND: This study illustrates our experience in treating duodenal ulcer by means of thoracoscopy and laparoscopy over a period of six years. MATERIALS AND METHODS: From October 1991 to October 1998, we submitted 38 patients (31 males and 7 females), average age 51 years (range 22-78 years), with duodenal ulcer to vagotomy with minimally invasive access: 23 Hill-Barkers, 2 Taylors, 9 thoracoscopic truncal vagotomies and 4 laparoscopic truncal vagotomies. The patients submitted to thoracoscopic truncal vagotomy had previous gastric surgery (5 ulcers of the neostoma in patients who had undergone gastric resection, 3 hemorrhagic gastritis of the gastric neostoma and 1 incomplete abdominal vagotomy). RESULTS: The average time required for the thorascopic approach was 30 minutes (range 20-40 minutes) with return to normal feeding in 1 day, without any difficulty, and discharge on day 3 (range 2-5 days). The patients were followed for 3-54 months. Twenty-two patients (91.3%) out of 23 submitted to anterior superselective and posterior truncal vagotomy, and the patients submitted to thoracoscopic vagotomy, were pain free without medical therapy. One patient (4.3%) was lost to the follow-up. There was only one relapse (4.3%) after seven months where the patient underwent left thorascopic truncal vagotomy. We had no mortality and no intraoperative or postoperative complications. CONCLUSIONS: In our opinion, minimally invasive treatment of peptic ulcer disease may represent the "gold standard." It is simple, quick, effective and delivers the same excellent results of open surgery but with minimum trauma. PMID- 10527333 TI - Laparoscopic exploration in the management of retroperitoneal masses. AB - BACKGROUND AND OBJECTIVES: The isolated finding of a retroperitoneal mass (RM) often represents a diagnostic challenge. Image-guided biopsy is frequently inadequate for diagnosis. With increasing experience, the use of laparoscopy for exploration of an indeterminate RM may provide a minimally invasive alternative to open exploration. Herein, we present a retrospective review of our initial four laparoscopic explorations, comparing our experience to four contemporary open explorations for an RM. PATIENTS AND METHODS: From July 1995 to January 1998, four patients, aged 50 to 62 years old, with an RM of undetermined etiology underwent laparoscopic exploration. Another four patients underwent open exploration at the same hospital. The medical records of these patients were reviewed. RESULTS: The tumors were smaller in the laparoscopic group, averaging 3.7 cm (range 2-6 cm) vs 6.5 cm (range 1-10 cm) in the open group. A definitive diagnosis was obtained for all eight patients. Postoperative complications were observed in one of the laparoscopic explorations, and in three of the open explorations; there was no operative mortality. The blood loss (90 vs 440 ml), fall in hematocrit (5.1 vs 7.8%), time to resumption of a regular diet (3 vs 5 days), amount of morphine sulfate equivalents required for analgesia (128 mg vs 161 mg), time to ambulation (2.3 vs 6 days) and hospital stay (4.8 vs 6 days) were all less among the laparoscopy patients. However, the operative time was longer for the laparoscopic procedure; this time included stent placement and patient repositioning in addition to the time for laparoscopic excision of the mass (7.8 vs 4.3 hours). CONCLUSION: Laparoscopic exploration appears to be a viable alternative to open exploration in patients presenting with a retroperitoneal mass. It is as effective as an open procedure and provides benefits with regard to patient morbidity and convalescence. However, operative time for this laparoscopic procedure is lengthy. PMID- 10527334 TI - Laparoscopy in the management of children with chronic recurrent abdominal pain. AB - BACKGROUND AND OBJECTIVES: The purpose of the present study was to evaluate the results of diagnostic laparoscopy in children with chronic recurrent abdominal pain. PATIENTS AND METHODS: Thirteen children with chronic recurrent abdominal pain were subjected to diagnostic laparoscopy. Ages varied from 10 to 17 years. There were six males and seven females. Abdominal pain was present from 3 weeks to 12 months (mean, 2 months). Extensive laboratory and imaging studies did not contribute to the diagnosis. In all patients, the pain was disabling and severe enough to warrant repeated visits to the pediatrician, emergency room visits, or hospital admissions, as well as absence from school. RESULTS: All children recovered uneventfully. Laparoscopic findings that identified the cause of abdominal pain were obtained in 12 of 13 patients. Laparoscopic appendectomy was done in all patients. There were no operative complications. One child presented three months later with incomplete small bowel obstruction, which resolved with conservative management. There were no other postoperative complications. Follow up varied from six months to three years. Abdominal pain resolved in ten patients. One patient presented eight months later with biliary dyskinesia. She improved following laparoscopic cholecystectomy and later on sphincterotomy, but her pain has not yet completely resolved. One patient presented six months later with abdominal pain secondary to intestinal adhesions. Her pain completely resolved after laparoscopic lysis of adhesions. A third patient who developed lower abdominal pain six months after laparoscopy improved with conservative management and antibiotics for pelvic inflammatory disease. CONCLUSIONS: Diagnostic laparoscopy is a valuable procedure in the management of children with chronic recurrent abdominal pain. In the present study, laparoscopic examination revealed the cause of abdominal pain in most patients, and this pain resolved in most cases. Based on our experience, we recommend diagnostic laparoscopy early in the course of debilitating chronic recurrent abdominal pain in children. Appendectomy should be done when no other significant cause of abdominal pain has been identified, even if the appendix looks normal. PMID- 10527335 TI - Chronic acalculous cholecystitis: changes in patient demographics and evaluation since the advent of laparoscopy. AB - BACKGROUND AND OBJECTIVE: To analyze patients with chronic acalculous cholecystitis over ten years, during which laparotomy was replaced by laparoscopy as the dominant operation for cholecystectomy in regard to patient demographics, diagnostic evaluations, follow-up symptoms, and additional operations. METHODS: Of 7181 cholecystectomies from June 1985 to June 1995, 301 patients had chronic acalculous cholecystitis. All subsequent hospital admissions and emergency room visits were reviewed through May 1997. Office records were available for review in 158 cases. Two eras were defined, the open era from June 1985 through May 1990, and the laparoscopic era from June 1990 through June 1995. RESULTS: Twice as many patients with chronic acalculous disease underwent cholecystectomy after the advent of laparoscopy. Patients with chronic acalculous disease were significantly younger than patients with cholelithiasis in both open and laparoscopic cases. The percentage of white women increased from 64.7% in the open to 75.7% in the laparoscopic era (p<0.05). The numbers of preoperative diagnostic tests performed decreased from 4.7+/-2.4 in the open to 3.2+/-1.8 in the laparoscopic era (p<0.05). Twenty-two percent of patients had continued symptoms postoperatively, and 8 patients (2.7%) required other abdominal operations within one year of cholecystectomy. CONCLUSION: Chronic acalculous cholecystitis is a disease of white females, doubling in frequency over the decade of review. Of these, 78% of patients had resolution of their symptoms on long-term follow-up. PMID- 10527336 TI - Laparoscopic-assisted resection of a bleeding gastrointestinal stromal tumor. AB - The authors report a case of a 29-year-old male patient with a severe lower gastrointestinal hemorrhage in whom a successful laparoscopic diagnosis and resection (assisted) of an ileal gastrointestinal stromal tumor (GIST) was performed. Laparoscopy can be very useful in the diagnosis and treatment of selected cases of lower gastrointestinal bleeding. PMID- 10527337 TI - Endometriosis ascites: a case report. AB - This is a case presentation of an usual nature, a 43-year-old Hispanic female, multigravida presenting with physical findings of massive ascites. In most instances, the presence of massive ascites is associated with malignancies, tuberculosis or perforated visous. In this case, the diagnosis of extensive endometriosis with ascites is reported as a very rare complication of the disease. PMID- 10527339 TI - Click c@refully before you quote: citing internet-based sources. AB - At the end of the 20th century, access to information provided by the World Wide Web (WWW) is changing as never before. The fast availability of current medical literature and the availability of tools for easy access to information, as well as for the easy production of information, have confronted research physicians, scholars, and students with new kinds of problems, many of which concern us personally. Quality control, difficulty establishing basic citation components, lack of standard guidelines for citing, as well as the short lifetime of Internet addresses concern us deeply. Some of these problems could be solved by the concept of an "Online-Library of Medicine" presented in the following paper. Since, however, at the present time there are no good answers to the problems regarding citing Internet-based sources, a Web surfer must keep in his or her mind the motto "caveat lector" (let the reader beware) - or, rather, in the spirit of our time: click c@refully before you cite. PMID- 10527338 TI - Pneumomediastinum as a complication of extraperitoneal laparoscopic inguinal hernia repair. AB - A 52-year-old man with left indirect groin hernia was admitted for elective inguinal repair using the totally extraperitoneal (TEP) approach. After an uneventful intubation, TEP repair of the hernia was performed with three midline trocars. Immediately after extubation, the patient noted severe chest pain. There was a decrease in PaO2 saturation, and neck subcutaneous emphysema was detected. There was no emphysema of the abdomen or of the back. A chest film and thoracic computed tomographic (CT) scan confirmed the presence of pneumomediastinum without pneumothorax. The patient was discharged without complications. PMID- 10527340 TI - Tailoring indicators to the surgical service. PMID- 10527341 TI - Helicopter transport: can physicians save lives? PMID- 10527342 TI - Clinical indicators in orthopaedic surgery. PMID- 10527343 TI - Current trends in radiation management of localized prostatic carcinoma. PMID- 10527344 TI - Addition of physicians to paramedic helicopter services decreases blunt trauma mortality. AB - BACKGROUND: The authors hypothesized that the addition of critical care physicians to the flight crew of paramedic helicopter services would decrease mortality in blunt trauma, and that this would be due to the greater procedural capability and clinical judgement of the physician. METHODS: Retrospective comparison was undertaken of patients flown directly from the accident scene over a 28-month period by the paramedic-staffed Westpac Hunter region helicopter to John Hunter Hospital, and the physician-staffed NRMA CareFlight helicopter to Westmead or Nepean Hospitals. Inclusion criteria were blunt trauma and an Injury Severity Score of > 10. Mortality was compared by trauma score-injury severity score (TRISS) methodology. RESULTS: There were 140 patients in the paramedic treatment group and 67 in the physician group. There were no significant differences between the groups in age, mechanism of injury, distance transported, response, scene or transport times. Physicians intubated a greater proportion of patients (51 vs 10%; P < 0.001) including all patients with a Glasgow Coma Score of < 9. Physicians gave significantly greater volumes of fluids to hypotensive patients (median: 5035 vs 1475 mL: P < 0.001) and performed thoracic decompressions on a larger proportion of patients (12 vs 1%; P < 0.01). The Z statistic for the physician treatment group was 2.72 (P < 0.01 ) compared with 1.16 (P = 0.25) in the paramedic group. The adjusted W statistic was 13.44 (95% CI: 7.80-19.08) suggesting that there would be between eight and 19 extra survivors per 100 patients treated in the physician group compared with the paramedic group. CONCLUSIONS: Physicians perform a greater number of procedures at accident scenes without increasing scene time. This results in significantly lower mortality. Critical care physicians should be added to paramedic helicopter services for scene response to blunt trauma. PMID- 10527345 TI - Should New South Wales hospital disaster teams be sent to major incident sites? AB - BACKGROUND: The aim of the present review was to assess the suitability of hospital disaster medical teams' training, personal safety and medical equipment for site casualty work at multiple casualty incidents (MCI), and to compare this with retrieval teams who routinely provide pre-hospital trauma care. The options for the provision of a site medical response based upon international and Australian disaster planning guidelines are also reviewed. METHODS: A questionnaire was mailed to all doctors dispatched to the 1997 Thredbo disaster as part of trauma service (TS) hospital medical teams, medical commanders or Helicopter Emergency Medical Service (HEMS) crew. Doctors with Sydney retrieval services (SRS) experience were compared with those without SRS experience in regard to the reported level of relevant training and experience as defined by current Australian guidelines and the Education and Training in Disaster Medicine Curriculum, Scientific Committee of the International Society of Disaster Medicine. Familiarity with medical equipment was assessed, as was level of compliance with Australian guidelines for personal protective clothing and equipment. RESULTS: Responses were obtained from all 25 doctors. Nine had SRS experience. None of the 16 doctors without SRS experience met the criteria of the Education and Training Curriculum, compared with four of nine doctors with SRS experience (44%). All six SRS doctors using SRS equipment had personally used or checked their equipment within 2 weeks prior to dispatch to Thredbo, compared with none of the 19 doctors using hospital equipment. Of the 11 areas of personal safety equipment and clothing assessed, all SRS doctors using SRS equipment complied with the guidelines in five areas (45%). There was no area assessed in which all the doctors using hospital equipment complied. CONCLUSION: Hospital medical teams suffer from the same problems of inadequate training, experience and personal safety equipment that are identified in previous reports from disasters overseas. The continued focus on hospital medical teams in counter disaster planning as the primary source of on-site medical services is inappropriate because, with the exception of retrieval doctors who routinely provide pre-hospital trauma care, appropriately trained and experienced doctors are unlikely to be available from within the hospital system. PMID- 10527346 TI - Curative external beam radiotherapy for prostate carcinoma: results in 231 patients treated in Lyon. AB - BACKGROUND: Radical prostatectomy and external beam radiation therapy (EBRT) are the mainstays of treatment of prostate cancer with curative intent. The possible development of radiation proctitis and rectal bleeding are major concerns when using EBRT. Recently, conformal radiotherapy has been introduced in an attempt to improve the results of EBRT. This paper presents an overview of the Lyon experience using standard EBRT with doses of 68 Gy, and reports the preliminary results of a study of conformal radiotherapy with dose escalation. METHODS: From 1981 to 1995, EBRT was used to treat 231 patients with localized adenocarcinomas of the prostate. The dose of EBRT was 68 Gy/34 fractions/7 weeks using a four field box technique with 18-MeV photons. A feasibility study of conformal radiotherapy was commenced in 1996. To date, 145 patients have been treated with doses escalating from 68 to 80 Gy. RESULTS: In the EBRT group of 231 patients, the 5-year overall survival was 80.3%. Anorectal function was scored as excellent in 90% of patients. Rectal bleeding was seen in 14.3% of patients and required local treatment in only seven. In the group treated with conformal radiotherapy, the preliminary results indicate good early tolerance. CONCLUSION: The curative treatment of patients with prostate cancer using EBRT gives good long-term survival with low rectal toxicity. Conformal radiotherapy appears to be an interesting approach to improve local control and perhaps survival. PMID- 10527347 TI - A prospective survey of current methicillin-resistant Staphylococcus aureus control measures. AB - BACKGROUND: Methicillin-resistant Staphylococcus aureus (MRSA) is now endemic in tertiary referral hospitals among the developed world. By prospective survey, the effect of two measures aimed to reduce the spread of MRSA was determined. First, a surgical ward with persistently high levels of MRSA detection was cleaned and renovated. Second, the medical records of all MRSA-colonized patients were electronically flagged, facilitating immediate application of control measures on readmission. METHODS: Data were collected for 995 newly colonized patients admitted between 1 July 1995 and 31 December 1997. Methicillin-resistant Staphylococcus aureus detection was determined before and after implementation of the interventions, along with the likely place of MRSA acquisition and the monthly incidence of MRSA detection for all inpatients. Chi-squared testing with odds ratios and 95% confidence intervals determined associations between the effect of control measures studied and MRSA detection rates. RESULTS: New MRSA detection was 21.6 per 1000 admissions before refurbishment compared with 20.4 per 1000 admissions to the surgical ward after refurbishment. New MRSA detection averaged 6.4 per 1000 hospital admissions before the introduction of record flagging and patient cohorting, compared with 6.2 per 1000 admissions after. CONCLUSION: Neither ward refurbishment, nor introduction of flagging, significantly reduced rates of colonization during the study period. In hospitals that receive MRSA-colonized patients and provide intensive care facilities, spread of MRSA is a major problem. Effective containment demands separate wards for MRSA-colonized and non-colonized patients. The need for such containment should be considered in design of the modern hospital. PMID- 10527348 TI - Management of biliary tract complications following liver transplantation. AB - BACKGROUND: A review of biliary tract complications was performed in 32 patients who underwent liver transplantation by the Western Australian Liver Transplantation Service during a 2-year period. METHODS: A review was made of patient data collected prospectively, and confirmed by retrospective casenote review. RESULTS: A total of 30 patients (31 grafts) survived more than 2 days after transplantation, and of these 28 had an end-to-end biliary anastomosis. Analysis of these 28 patients found that eight of 17 patients with T-tubes had complications: three leaks at T-tube removal; two strictures and leaks; and three strictures. Six of 11 patients without a T-tube had complications: one leak; three strictures and leaks; and two strictures. Predisposing factors were present in eight of the 14 patients with biliary tract complications: hepatic artery stenosis in three; and one each with hepatic artery thrombosis; biliary calculi; donor-recipient bile duct mismatch; severe cellular rejection: and prolonged postoperative hypotension. Acute rejection, steroid-resistant rejection and cytomegalovirus infection were all significantly more common in those patients with biliary tract complications compared with those without. There was no difference in cold ischaemic time or donor age. Twelve of the 14 patients with biliary complications required endoscopic stenting with or without balloon dilation, and eight patients required radiological percutaneous drainage of bile collections. Only one patient required biliary reconstruction and two patients required re-transplantation. One patient died of uncontrolled infection. Of three patients who underwent choledochojejunostomy, biliary leak developed in two patients, both of whom required operative biliary and hepatic repair. One of the three patients died from disseminated Aspergillus infection. The median total hospital stay of patients with biliary complications was 61 days (range: 30-180 days) compared with 33.5 days (range: 22-70 days) for patients without. Of patients with end-to-end biliary anastomosis, 50% had biliary tract complications and more than half of these had predisposing factors. The majority of biliary complications were managed without the need for surgery. CONCLUSION: A total of 50% of patients with end-to-end biliary anastomosis had biliary tract complications. Biliary strictures presented later than leaks, and the majority of these complications were managed without the need for surgery. PMID- 10527349 TI - How to predict return to the community after fractured proximal femur in the elderly. AB - BACKGROUND: There are numerous studies about morbidity and mortality, technical complications and in-hospital factors after proximal femoral fracture surgery in the elderly. Although experienced clinicians are often able to make an accurate prediction, little information is available about the factors that allow early determination of whether a patient may return to the community. The present study aimed to provide that information and, hence, allow better use of health resources. METHODS: At Westmead Hospital a prospective study was conducted of 304 patients with a proximal femoral fracture who were previously residing at home. A number of different factors were analysed using statistical methods to determine their effect on outcome, which was defined simply in terms of whether the patient was able to return to the community or whether they needed institutional care. RESULTS: Factors that had an adverse influence on return to the community were: a low activities of daily living (ADL) score on admission; increasing age; dementia; the use of regional analgesia; and the occurrence of postoperative complications. Other factors such as gender, fracture type, delay to surgery and length of stay were not significant. CONCLUSIONS: Consideration of the ADL score, age and mental state at the time of admission to hospital is all that is needed to determine return to the community. This is helpful to the patient and their family, and allows an early and appropriate referral pattern to either community services or a nursing home. PMID- 10527350 TI - Donor site morbidity in the iliac crest bone graft. AB - BACKGROUND: Open iliac crest bone grafting is a common surgical procedure with recognized short-term complications. The present paper documents the medium- to long-term complications and level of patient satisfaction following the procedure. METHODS: Seventy-three patients undergoing an open iliac crest bone graft over the past 6 years at Wellington Hospital were retrospectively reviewed. All patients completed a postal questionnaire which assessed their current pain, sensory changes in and below the scar, scar appearance and overall appearance with the bone graft donor site. RESULTS: Sixteen patients (21.9%) reported pain, 11 patients (15%) stated that their scar was sensitive to touch and 19 patients (22%) reported a degree of sensory change below the scar. Six patients (8.2%) felt that the scar appearance was totally unacceptable. Overall satisfaction, however, was high with 70 patients (95.8%) 'fairly satisfied' or 'totally satisfied' with their iliac crest donor site. CONCLUSIONS: Bone grafting from the iliac crest is a relatively benign procedure in terms of patient satisfaction, and the most significant morbidity is pain. PMID- 10527351 TI - Peritonectomy and intraperitoneal chemotherapy in appendiceal and colorectal cancer. AB - BACKGROUND: Peritoneal spread of gastrointestinal malignancies has been regarded as an incurable disease, and treatment has been aimed at short-term palliation. The use of cytoreductive surgery, including peritonectomy procedures and intraperitoneal chemotherapy, has been proposed with the intention of prolonging survival, and perhaps curing patients with peritoneal carcinomatosis from appendiceal and possibly colon cancers. A series of eight patients who have undergone this procedure at St George Hospital is presented, and the results obtained by other groups are reviewed. METHOD: Eight patients fitted the criteria for peritoneal carcinomatosis between January 1996 and November 1998. In seven patients this was secondary to appendiceal or colon cancer. and one patient had signet ring cancer of the uterus. The surgical treatment involved removing all macroscopic evidence of disease, and this was followed by early postoperative intraperitoneal chemotherapy. RESULTS: The eight patients (seven female, one male) ranged in age from 25 to 67 years. There were seven complications, including two patients with pelvic abscesses, and one patient who developed Tenchkoff catheter occlusion. There were three deaths, one due to pelvic sepsis after 30 days, and the other two were due to metastatic disease. Of the remaining five patients, two have developed recurrence and three remain disease-free. CONCLUSION: The results of peritonectomy and intraperitoneal chemotherapy for appendiceal tumours are encouraging. The role in colorectal cancer is less clear, although there are some reports that suggest a benefit. PMID- 10527352 TI - Familial fatal and near-fatal third ventricle colloid cysts. AB - BACKGROUND: Despite having a presumed congenital origin, familial cases of colloid cysts have been reported only rarely. The first case of a brother and sister with colloid cysts is reported here, and the relevant literature is reviewed. METHODS: A 25-year-old man presented with a 24-h history of headache and vomiting. He rapidly became unconscious and fulfilled the criteria for brain death on arrival at hospital. No surgical intervention was performed. RESULTS: The patient's sister presented at the age of 41 with headaches and rapidly became unconscious. The sister had urgent bilateral ventriculostomies. followed by transcallosal removal of a colloid cyst. CONCLUSIONS: These cases support the hypothesis that colloid cysts are congenital lesions and provide some evidence of a possible genetic predisposition to their formation. Sudden death remains a real risk for patients harbouring a colloid cyst. PMID- 10527353 TI - Pre-operative chemoradiotherapy in locally advanced rectal cancer. AB - BACKGROUND: The aim of the present study was to investigate the effectiveness and toxicity of pre-operative chemoradiation in locally advanced rectal cancer (T3 T4). METHODS: Forty-seven patients were assessed (38 T3 and nine T4 tumours). Pre operative pelvic radiotherapy was delivered in four fields, 45 Gy in 25 fractions over 5 weeks. Bolus 5-fluorouracil (5-FU) was delivered 500 mg/m2 on days 1, 2, 3 and days 22, 23, 24. Total mesorectal excision of the rectal tumour either by anterior or abdomino-perineal resection was planned at 4-6 weeks from completion of pre-operative treatment. Response to therapy was assessed by fresh macroscopic measurement of the surgical specimen. RESULTS: All patients undergoing chemoradiation completed therapy as planned, with no treatment-related interruptions. The regimen had a low acute toxicity profile with an estimated 50% or greater response in 38 out of 47 patients (four patients had complete responses). Forty-three (97%) of 44 patients who underwent surgery were operable. Patients who were operated on between 4 and 7 weeks had a statistically better response then those who were operated on after 7 weeks (P = 0.013; Fisher's exact test). Eight of 10 patients who were considered to be inoperable prior to the treatment underwent total mesorectal excision with negative radial margins. Anastomotic leakage occurred in four patients (9%); one required surgical intervention. Wound infection occurred in three patients (6%); one patient required re-exploration for haemorrhage. Delayed complications occurred in three patients (6%); one requiring surgery for a stomal stricture. After a median follow-up of 20 months, two patients (4%) had developed local recurrence. CONCLUSION: The pre-operative chemoradiation regimen employed had a low acute toxicity profile and all patients completed therapy. The majority of patients considered inoperable prior to receiving this treatment underwent successful excision. Appropriately fractionated pre-operative chemoradiotherapy is a reasonable option in this disease and deserves further evaluation. PMID- 10527354 TI - Laparoscopic radical deroofing of hepatic cysts using the ultrasonic scalpel. AB - BACKGROUND: There is some debate regarding the role of laparoscopy in deroofing of liver cysts. METHODS: Three patients underwent laparoscopic deroofing by means of an ultrasonic scalpel. RESULTS: All three patients underwent treatment successfully with minimal postoperative morbidity and hospital stay. CONCLUSIONS: The ultrasonic scalpel has a useful role in the laparoscopic treatment of hepatic cysts. PMID- 10527355 TI - What's new in plastic surgery? AB - A review is made of the latest developments in the field of plastic and reconstructive surgery. The subspecialty divisions are more pronounced in the modern era, but are still linked by a unified approach to soft-tissue management. Although cornerstone areas such as microsurgery have seen refinement, other fields, including aesthetic surgery, have been totally redefined by laser technology and the spread of endoscopic techniques. PMID- 10527356 TI - An easy and effective method of cholangiography in laparoscopic cholecystectomy: laparoscopic cholecystocholangiography: comment. PMID- 10527357 TI - Guidelines for the surgical management of breast cancer: comment. PMID- 10527358 TI - Giant leiomyosarcoma of the transverse mesocolon. PMID- 10527359 TI - Sarcoidosis presenting as hoarseness and dysphagia. PMID- 10527360 TI - Familial ulnar nerve instability. PMID- 10527361 TI - The role of sulindac in familial adenomatous polyposis patients with ileal pouch polyposis. PMID- 10527362 TI - Pott's puffy tumour: a clinical variant. PMID- 10527363 TI - Peripheral blood progenitor cell cycle kinetics following priming with pIXY321 in patients treated with the "ICE" regimen. AB - PURPOSE: Treatment with hematopoietic growth factors increases the percentage of hematopoietic progenitor cells in cell cycle. Following withdrawal of certain growth factors, preclinical data suggest that there is a transient fall in the percentage of progenitor cells in cycle below the baseline, thus providing a window to administer chemotherapy with reduced risk of myelotoxicity. PATIENTS AND METHODS: Patients with histologically confirmed, previously untreated neoplasia, were treated with pIXY321 by subcutaneous injection at a dose of 375 microg/m2 twice daily (total dose 750 microg/m2/day) for seven days (days -8 to 2), followed by a two-day rest (days -1 to 0). Patients received ICE (ifosfamide, carboplatin and etoposide) on days 1 to 3. On day 4, pIXY321 was resumed until hematologic recovery. Peripheral blood was collected on days -8, -2, -1, 1, and cell cycle distribution was determined using flow cytometry. RESULTS: Twenty patients were treated in this study and received a total of 54 cycles. Partial responses were observed in three of 13 patients with non-small cell lung cancer (23 percent) and two of five patients with small cell lung cancer (40 percent). Six of 15 patients had an increased number of cells in S+G2/M on day 1 of ICE following seven days of pIXY321 and two days off (days -1 to 0). The average increase was 63 percent (range 6-253). Seven patients had a decreased number of cells in S+G2/M. The average decrease was 55 percent (range 6.3-78). There were no significant differences among the fifteen patients with regards to the observed toxicity of the chemotherapy. DISCUSSION: pIXY321 in this schedule did not consistently decrease the percentage of cycling progenitor cells in the peripheral blood. Future studies should define whether other growth factors and/or schedules can synchronize progenitor cell cycling and protect the marrow compartment from cycle specific chemotherapy. PMID- 10527365 TI - Guillain-Barre syndrome: rehabilitation outcome and recent developments. AB - Guillain-Barre syndrome is the most common polyneuropathy causing major disability and respiratory failure. Respiratory complications are the main cause of death. Improved respiratory care and new treatment strategies such as plasmaphoresis and immunoglobulin have been shown to improve outcome. We studied the course and outcome of 37 patients with Guillain-Barre syndrome who were admitted to a rehabilitation and respiratory care facility over a 10-year period. There were 21 males and 16 females with a mean age of 62+/-3 years. Fourteen patients developed respiratory failure requiring endotracheal intubation and mechanical ventilation. The mean duration of mechanical ventilation was 38+/-10 days. All patients were successfully liberated from the ventilator. However, 83 percent of the patients were moderately to severely disabled at the time of discharge. Thirteen out of 37 (35 percent) developed long-term disability. None of the patients died over the period of follow-up. These results indicate that early recognition and treatment of respiratory complications in Guillain-Barre syndrome could reduce the morbidity and mortality of this condition. PMID- 10527366 TI - Efficacy of promethazine suppositories dispensed to outpatient surgical patients. AB - Postoperative nausea and vomiting frequently complicate outpatient anesthesia and surgery. The duration of treatment for this complication must occasionally extend beyond discharge from the hospital. In this study, we evaluated the commonly used anti-emetic promethazine for its efficacy in the post-discharge period. Adult outpatient surgical patients who had excessive postoperative nausea and vomiting in the recovery room, or who were at risk for postoperative nausea and vomiting following discharge were given two promethazine suppositories (25 mg) for home use. All patients were contacted by our recovery room nurses on the first business day after their surgery and questioned as to their use of the suppositories and, if used, their efficacy. We found that 55 percent of patients given promethazine suppositories for home use had nausea and vomiting in the post discharge period. Of the patients given promethazine, 89 percent used the suppositories. All of these patients reported improvement in their symptoms following use of the suppositories. None reported adverse effects from the promethazine suppositories. In conclusion, we found promethazine suppositories to be an inexpensive and efficacious treatment for nausea and vomiting in adult outpatient surgical patients following discharge from the hospital. Side-effects were minimal, and our patients voiced no complaints about this mode of therapy. We recommend this therapy for treatment of nausea and vomiting after hospital discharge following adult outpatient surgery. PMID- 10527367 TI - A century of pathology at Yale: personal reflections. AB - This history is largely about the players on the stage of the Yale Pathology Department acting out their roles as observed by the author in over a half century as a member of the department and as associate dean of the medical school. PMID- 10527368 TI - Grating orientation as a measure of tactile spatial acuity. AB - Recent studies have used grating orientation as a measure of tactile spatial acuity on the fingerpad. In this task subjects identify the orientation of a grooved surface presented in either the proximal-distal or lateral-medial orientation. Other recent results have suggested that there might be a substantial anisotropy on the fingerpad related to spatial sensitivity. This anisotropy was revealed using a task in which subjects discriminated between a smooth and a grooved surface presented at different orientations on the fingerpad. The anisotropy was substantial enough that it might permit subjects to discriminate grating orientation on the basis of intensive rather than spatial cues. The present study examined the possibility that anisotropy on the fingerpad might provide cues in a spatial acuity task. The ability of subjects to discriminate between a smooth and a grooved surface was measured under conditions that are typically used in grating orientation tasks. No evidence of anisotropy was found. Also, using a grating orientation task, separate estimates were made of sensitivity in the proximal-distal and lateral-medial orientations. Again no evidence of anisotropy was found. Consistent with changes in the density of innervation, grating orientation sensitivity was found to vary as a function of location on the fingerpad. The results support the view that grating orientation is a valid measure of spatial acuity reflecting underlying neural, spatial mechanisms. PMID- 10527364 TI - Aging and reproductive potential in women. AB - Reproductive potential in women declines with age. Age-related changes in the ovary account for most of this loss of reproductive function. Oocytes, all of which are present at birth, decline in number and quality with age. The endocrine function of the ovary also declines with age, and the ovary becomes unable to sustain its normal function in the neuroendocrine axis. The neuroendocrine axis may be further affected by primary changes occurring in the hypothalamus and pituitary during aging, although this has not been established in humans. Aging also affects the function of the uterus as the endometrium loses its ability to support implantation and growth of an embryo. Diminished uterine function during aging may be due to changes in the uterine vasculature or to changes in the hormone-dependent development of the endometrium. Finally, aging increases a woman's risk of developing medical, gynecologic or obstetric conditions that may impair her fertility. Knowledge of these affects of aging on a woman's reproductive function is essential to advise and treat the growing number of women seeking pregnancy at advanced reproductive age. PMID- 10527369 TI - Descending projections arising from the parafascicular nucleus in rats: trajectory of fibers, projection pattern and mapping of terminations. AB - The organization of the descending projections from the intralaminar parafascicular nucleus was studied using biocytin as an anterograde tracer in rats. After biocytin injection into the lateral parafascicular nucleus, three bundles of fibers descending throughout the brainstem were seen. Terminal fields were found in several structures, for example the lateral geniculate nucleus, nucleus reticularis thalami, subthalamus, zona incerta, substantia nigra, red nucleus, periaqueductal gray, superior colliculus, reticular formation, raphe nuclei, pontine nuclei, trigeminal complex, and ventral horn of the spinal cord. Different types of labeled terminals (small terminal boutons, en passant varicosities, large claw-like terminals) were observed, particularly in the substantia nigra and reticular formation where the density of terminals was highest. The organization of these extensive descending connections to both motor and sensory structures does not allow functions to be conclusively attributed to the parafascicular nucleus neurons. Further investigations are required to resolve this question. PMID- 10527370 TI - Estimation of the number and size of female adult rat C4, C5 and C6 dorsal root ganglia (DRG) neurons. AB - In previous studies primary sensory neurons of adult rats have been counted in lumbar dorsal root ganglia. However, different counting methods have given very different results and at the cervical level, recent data are scarce. In the present study, the number of neurons in C4, C5 and C6 adult rat ganglia was determined using two previously calibrated techniques. The stereological tool was preferred because it directly identifies neurons instead of nucleoli and is more efficient. The C4, C5 and C6 dorsal root ganglia were found to contain 7508+/ 299, 6825+/-950 and 6858+/-923 neurons, respectively, and statistical analysis indicated that there was no significant difference between the three levels. There was, however, a great interindividual variation, which was also found at other levels of the spinal cord. The mean diameter of neurons in the C4, C5 and C6 dorsal root ganglia was determined and was 17.52, 20.16 and 20.68 microm, respectively. It is important to know more about the organization of the sensory systems in the normal rat. Once established, the number of neurons in these dorsal root ganglia could be compared with different pathological situations or experimental treatments such as developmental conditions, nerve section or ganglion transplantation. PMID- 10527371 TI - Vibrotactile temporal summation: probability summation or neural integration? AB - Temporal summation, a decrease in the detection threshold that occurs when either the duration of a stimulus or the number of stimuli in a sequence is increased, has been attributed to the operations of either the mechanism of neural integration or of probability summation. Our experiments indicate that under certain conditions, both mechanisms may operate, but that the process of neural integration is an exclusive characteristic of the Pacinian (P) channel. The P channel was isolated by applying 250 Hz stimuli through a 1.5 cm2 contactor to the thenar eminence of the hand and the NPII channel was isolated by applying the stimuli through a 0.01 cm2 contactor. The finding that the slopes of the psychometric functions were the same within both channels indicated that probability summation could not account for temporal summation for stimulus durations less than 1 s. The finding that the threshold for the detection of multiple-pulse stimuli increased as the interpulse interval increased indicated that, for time intervals less than 800 ms, temporal summation results from neural integration. But for interstimulus intervals greater than 800 ms, probability summation accounts for temporal summation. PMID- 10527372 TI - Influence of segmental and extra-segmental conditioning, stimuli on cortical potentials evoked by painful electrical stimulation. AB - This study evaluated the effects of two different types of segmental/extra segmental conditioning stimuli (tonic muscle pain and non-painful vibration) on the subjective experience (perceived pain intensity) and on the cortical evoked potentials to standardized test stimuli (cutaneous electrical stimuli). Twelve subjects participated in two separate sessions to investigate the effects of tonic muscle pain or cutaneous vibration on experimental test stimuli. The experimental protocol contained a baseline registration (test stimuli only), a registration with the test stimuli in combination with the conditioning stimuli, followed by a registration with the test stimuli only. In addition, the effects of the conditioning stimuli were examined at two anatomically separated locations (segmental and extra-segmental). Compared with the test stimulus alone, the perceived pain intensity and peak-to-peak amplitudes of the evoked potentials were unchanged in the presence of non-painful conditioning stimuli at either location. In contrast, a significant decrease of the perceived pain intensity and peak-to-peak amplitudes was found in the presence of painful conditioning stimuli at the extra-segmental sites. Moreover, the topographic maps of the 32-channel recordings suggested that the distribution of the scalp evoked potentials was almost symmetrical around the vertex Cz in the baseline registration. The evoked potentials were generally decreased during hypertonic saline infusion at the extra-segmental sites, but the distribution of the topographic maps did not appear to change. Vibration has previously been shown to inhibit pain, but in the present study the perceived intensity of phasic painful electrical stimuli was unchanged. The reduced perceived pain intensity and the smaller peak-to-peak amplitude of the evoked potential in the presence of extra-segmental conditioning pain are in accordance with the concept of diffuse noxious inhibitory control. PMID- 10527373 TI - Biomechanical model predicts directional tuning of spindles in finger muscles facilitates precision pinch and power grasp. AB - Humans have a sense of static limb position derived primarily from the output of secondary muscle spindle endings. The features of finger pose these proprioceptors signal best were predicted by singular value decomposition of a kinematic model of the human long finger and the six muscles that actuate it. The analysis indicated that muscle spindles signal the location of the fingertip with less error than they signal angles of individual finger joints. The fingertip displacements for which proprioceptors have greatest sensitivity were also predicted. These fingertip displacements seem to correspond to the fine positioning of an object pinched between the fingertip and distal phalanx of the thumb. The analysis also predicted the directions in which subjects can displace the fingertip most rapidly. The directions seem to correspond to rapid closure of precision pinch or power grasp. PMID- 10527374 TI - Psoriasis and the arachidonic acid cascade. AB - Arachidonic acid (5.8,11,14-eicosatetraenoic acid C20:4, n-6) is released from the cell membrane by the action of phospholipases on membrane phospholipids. Metabolites of arachidonic acid, which are generically termed eicosanoids, including prostaglandins, thromboxane, leukotrienes and hydroxyeicosatetraenoic acids, have been implicated as mediators or modulators of a number of physiological functions and pathological conditions in both normal and diseased human skin. Particularly, eicosanoids have been suspected to play an important role in the pathogenesis of psoriasis, because a number of phenomena observed in psoriasis can be explained, at least in part, by the action of eicosanoids. This review will focus on recent progress regarding the significance of eicosanoids in the pathogenesis of psoriasis. Recent developments in the molecular biology in the eicosanoids have renewed interest in the role of eicosanoids in psoriasis. New understanding of the etiology of psoriasis and advances in its treatment due to recent progress in eicosanoid biology will also be presented. PMID- 10527375 TI - Dermatological applications of skin potential level measurements. AB - Decreasing of the negativity of skin potential level (SPL) on the palmar surface of healthy subjects associated with the process of falling asleep, with a return to increased negativity on awakening, was reported by Nishimura and Nagumo (Ergonomics 1985;28(6):905-913). To clarify the dermatological significance of SPL measurements, the SPL of normal skin at various paired sites in the right upper limb, right lower limb and trunk in healthy volunteers and also at the sites of lesions in patients with psoriasis, herpes zoster and nummular eczema were measured. Fitting curves for the decreasing showed exponential characteristics on the palm and the heel where many sweat glands are involved. SPL differences on other sites did not show the exponential decreasing, but kept almost constant values. The values are negative on normal skin, but positive on lesions sites. We also studied the effects of water content in the horny layer on SPL differences and the biorhythmical changes of SPL differences. We discussed the reasons why these results were obtained. The present study provides the promise of an objective, noninvasive and easy method of testing dermatological conditions. PMID- 10527377 TI - Characteristics in adherence of streptococci and Staphylococcus aureus isolated from various infective skin lesions: serum IgA decreases adherence of Streptococcus pyogenes but not Staphylococcus aureus. AB - We characterized adherence of streptococci and Staphylococcus aureus strains isolated from various infective skin lesions in terms of hydrophobicity, negative charge, tube adherence, slime production, and influence on adherence to coverslips by plasma and serum immunoglobulins. High hydrophobicity was more frequently observed in Streptococcus pyogenes strains than in Streptococcus agalactiae strains (P < 0.01) and S. aureus strains (P < 0.001) and slime production was more frequently observed in S. agalactiae strains than in S. pyogenes strains (P < 0.05). Serum IgA decreased adherence to coverslips of S. pyogenes strains but not that of S. aureus strains. PMID- 10527376 TI - Complementary peptides against the major epitope in the NC16A domain of BP180 show no specificity as vaccines to bullous pemphigoid. AB - A stretch of 14 amino acids (542-555) (MCW-1) in the NC16A domain of BP180 has been shown to be an immunogenic and pathogenic epitope for bullous pemphigoid (BP). Therefore, it provides an excellent target for treatment through a complementary peptide approach, which has been established in other autoimmune diseases, including experimental autoimmune myasthenia gravis. We examined two synthetic complementary peptides BP3CP5 and BP5CP3 against this region. These peptides were derived, respectively, by reading the antisense RNA of this region of BP180 in 3'-5' and 5'-3' directions. We found evident complementarities in hydropathic scores between MCW-1 and both complementary peptides. However, by enzyme-linked immunosorbent assay (ELISA), the complementary peptides BP3CP5 and BP5CP3 did not bind to either synthetic peptide BPNP or glutathione-S-transferase (GST) fusion proteins BP180NC16a and GST-BP-1050. BPNP, BP180NC16a and GST-BP 1050 cover the MCW-1 region of BP180 and were used as the natural peptides in this study. In addition, neither BP3CP5 nor BP5CP3 blocked the reaction between BPNP and anti-BPNP antibody, nor did they block immunofluorescent staining of the basement membrane zone by BP sera. Pre-incubation with BP3CP5 and BP5CP3 did not block the binding of BP sera to the BP18NC16a fusion protein in immunoblotting. Furthermore, rabbit antisera raised against BP3CP5 and BP5CP3 did not bind BP sera in ELISA. Pre-incubation with these rabbit antisera did not inhibit or reduce the binding of BP sera to the autoanltigen in either imnmunoblotting or immunofluorescence. Thus, we concluded that complementary peptides against this particular epitope in BP180 NC16A domain showed no specificity as vaccines to BP, although this approach should be tried for other epitopes in various autoimmune bullous diseases. PMID- 10527378 TI - Antigenic characterization in ampiroxicam-induced photosensitivity using an in vivo model of contact hypersensitivity. AB - Ampiroxicam (APX), a prodrug of piroxicam (PXM), has been reported to induce photosensitivity. Antigenic characterization of these photosensitivities, however, is still insufficient. The purpose of the present study was to elucidate further mechanism of photosenstivity induced by APX and PXM using an in vivo model of contact hypersensitivity in guinea pigs. Animals sensitized with ultraviolet-A (UVA)-irradiated 1% APX showed positive reaction in the patch testing to UVA-irradiated 1% APX and 1% thiosalicylate (TOS), while they were negative in challenge with UVA-irradiated 1% PXM, non-irradiated APX and PXM, whereas none of UVA-irradiated or non-irradiated APX and PXM showed positive patch test reaction in animals sensitized with UVA-irradiated 1% PXM or control vehicles. Animals sensitized with 1% TOS were successfully challenged by 1% TOS and cross-reacted with UVA-irradiated 1% APX; however, they failed to react with UVA-irradiated PXM, non-irradiated APX and PXM. Indeed, the in vitro study revealed that the concentration of APX was easily reduced by the increase of UVA irradiation dose, as compared with that of PXM. Interestingly, absorption spectrum of UVA-irradiated APX was similar to that of TOS, which is thought to be an active hapten of PXM. In the present study, we succeeded in the development of a novel animal model reflecting the clinical observations. Furthermore, these results suggested that contact hypersensitivity induced by UVA-irradiated APX is developed by photoproducts of APX itself, but not by the biotransformation of APX to PXM. PMID- 10527379 TI - Differences between fibroblasts cultured from oral mucosa and normal skin: implication to wound healing. AB - It is generally agreed that oral mucosa heals faster with less scar than skin does, and hypertrophic scar or keloid is very rare in the oral cavity. Fibroblasts are thought to play an important role in wound healing and scar formation, whose control is mediated by growth factors. We have studied whether there are any differences in the cellular behavior of fibroblasts between oral mucosa and skin, and in their response to growth factors. Oral mucosal fibroblasts proliferated slightly more than dermal fibroblasts on average. Dermal fibroblasts in collagen gel possessed greater contraction potency than oral mucosa fibroblasts, irrespective of the presence of growth factors; however, oral mucosa fibroblasts showed an earlier collagen gel contraction with or without TGF beta1. There were no differences in basal collagen synthetic rate between dermal and oral mucosal fibroblasts, while the latter synthesized more collagen than dermal fibroblasts when they were stimulated with TGF-beta1. Our study showed that oral mucosal fibroblasts and dermal fibroblasts had selective differences in cellular behavior and in their responses to growth factors, which seems to contribute to the differences in wound healing. PMID- 10527380 TI - A study of anti-carbonic anhydrase II antibodies in rheumatic autoimmune diseases. AB - Autoantibodies to human carbonic anhydrase II (CAII) were screened by ELISA in 109 sera from Asian Japanese patients with systemic lupus erythematosus (SLE), primary Sjogren's syndrome (Sjs), progressive systemic sclerosis (PSS) and dermatomyositis (DM). Anti-CAII antibodies were positive in 24.1% of SLE, 20.0% of primary Sjs, 16.7% of PSS and 25.0% of DM. On the other hand, sera from atopic dermatitis, bullous pemphigoid and psoriasis patients showed no activity for anti CAII antibodies. CAII could be a common exonuclear autoantigen in subsets of rheumatic autoimmune diseases. PMID- 10527382 TI - In vitro immunization with a recombinant antigen carrying the HIV-1 RT248-262 determinant inserted at different locations results in altered TCRVB region usage. AB - Immunodominance or cripticity of a peptide-borne determinant may be influenced by the protein context in which the epitope is embedded. In this frame, we previously showed that certain human T cell clones, derived from different donors, may differentially recognize the RT248-262 helper determinant depending on whether it is provided to the presenting cells as a synthetic peptide or as a recombinant carrier protein to which the sequence of interest is fused. We now report that, upon in vitro immunization of human PBL with autologous APC, the epitope-specific TCRVB repertoire obtained when selection is applied by pulsing the APC with the cognate synthetic peptide is different from that found when a recombinant protein is used in which the antigenic sequence is placed at either a N-terminal or C-terminal location of the GST carrier. As the TCRVB distribution is not a function of the APC used, we propose that processing of different recombinant molecules containing the same epitope may generate MHC/peptide complexes which, being antigenically diverse, may recruit distinct TCR specificities. These findings may be relevant for evaluating and predicting the immunogenic potential of subunit vaccines based on synthetic peptides or on recombinant proteins as compared to the native antigen. PMID- 10527383 TI - Induction and tolerization of anti-male CD8+ cytotoxic T lymphocytes by in vivo immunization with an H-Y-derived peptide. AB - We have analyzed the immune response induced by a 9mer synthetic peptide derived from the male histocompatibility antigen H-Y and containing Db-binding motifs in C57BL/6 mice. In this study we report that a single, subcutaneous injection of the peptide emulsified in IFA gave rise to the development of male-specific CD8+ T cells which displayed H-Y-specific proliferative response in vitro and showed a Tc1-type pattern of cytokine production (i.e. they secreted IFN-gamma and IL-2, but not IL-4 and IL-10). Development of a strong cytotoxic activity required in vitro stimulation with specific peptide and IL-2: under these culture conditions, we were able to generate potent CD8+ CTLs that lysed both male cells and peptide pulsed female cells. Continuous administration of soluble peptide, delivered over a 7-day period by a mini-osmotic pump implanted subcutaneously, inhibited proliferative and cytotoxic responses and IFN-gamma production in lymph node cells from C57BL/6 mice subsequently primed with peptide in adjuvant. This decreased responses were associated with a strong increase in the secretion of IL 4 by antigen-specific CD8+ T lymphocytes. Subcutaneous administration of the H-Y peptide in adjuvant significantly accelerates rejection of male skin graft, while continuous administration of peptide in soluble form did not modify the time course of rejection. PMID- 10527381 TI - Analysis of the mutant HLA-A*0201 heavy chain H74L: impaired TAP-dependent peptide loading. AB - A mutation of the HLA-A*0201 heavy chain at position 74 from histidine to leucine (H74L) resulted in a molecule with an interesting phenotype. H74L-expressing targets were recognized by peptide-specific HLA-A*0201-restricted cytotoxic T lymphocytes at lower peptide concentrations than wild type HLA-A*0201. H74L's improved ability to sensitize cells for tysis was due to its enhanced capability to bind exogenous peptide. Furthermore, this phenotype of improved exogenous binding and functional recognition was not peptide-specific. In contrast, the H74L molecule failed to present the HIV- HLA-A2-restricted pol peptide when expressed and processed endogenously. The inability to bind endogenous pol could be rescued by preceding the pol peptide with a signal sequence. The defect affecting endogenous presentation, therefore, appeared to be limited to the TAP dependent pathway. Surprisingly, the H74L heavy chain was able to enter the defined MHC class I pathway and associate with beta2M, calreticulin, tapasin, and TAP. Despite the presence of the H74L heavy chain at the TAP complex, H74L was functionally inefficient at loading TAP-dependent peptides. H74L may help elucidate further steps in the process of loading TAP-dependent peptides into the class I cleft. PMID- 10527384 TI - Immune response to a recombinant human TNFR55-IgG1 fusion protein: auto antibodies in rheumatoid arthritis (RA) and multiple sclerosis (MS) patients have neither neutralizing nor agonist activities. AB - A substantial number of patients enrolled in clinical studies of TNFR55-IgG1 in TNF-neutralizing treatment of rheumatoid arthritis and multiple sclerosis developed antibodies to the recombinant human protein. To enable more detailed investigation subgroups of patients donated small blood samples. TNFR55-IgG1 reactive antibodies were affinity purified from plasma; IgM and IgG class antibodies reactive with TNFR55-IgG1 were found which varied considerably in titer and kinetics of appearance among individual patients. The affinity purified antibody fractions included specificities to the receptor moiety of TNFR55-IgG1, but also rheumatoid factor and other pre-existing antibodies directed to the IgG1 moiety. The antibodies bound to Fc receptors, but not detectably to TNFR55 at the human cell surface. No agonistic nor neutralizing activities of these antibodies were detected. Major linear epitopes clustered in the TNFR55 sequence in close proximity to the IgG1 fusion site. The relative content of antibodies to linear and conformational epitopes was highly variable among patients. Route and frequency of administration rather than underlying disease appeared to influence the major linear B cell epitopes selected. PMID- 10527385 TI - The expression of killer cell inhibitory receptors on natural killer cells and activation status of CD4+ and CD8+ T cells in the decidua of normal and abnormal early pregnancies. AB - The establishment of the human placenta in early pregnancy is characterized by the presence of large numbers of natural killer cells within the maternal decidua. These NK cells have an unusual phenotype, CD3- CD16- CD56(bright), distinguishing them from peripheral blood NK cells. They may control trophoblast migration and placentation. Using a panel of monoclonal antibodies to several members of the KIR family and flow cytometry, we found that KIRs are expressed on decidual NK cells. There is variation in both the percentage of cells expressing a particular receptor and the density of receptor expression between decidual NK cells from different individuals. In anembryonic pregnancy, the proportions of decidual NK cells with a particular KIRs (GL183 and EB6) decreased significantly when compared with normal pregnancy (p = 0.01 and 0.01, respectively), raising the possibility that these NK receptors may be involved in recognition of the allogeneic fetus by the mother at the implantation site. In the decidua, more CD4+ and CD8+ T cells expressed CD69 and HLA-DR than in blood, indicating that T cells are regionally activated during early pregnancy. When compared with normal pregnancy, decidual HLA-DR+CD4+CD3+, CD69+CD8+CD3+ and HLA-DR+CD8+CD3+ T lymphocytes are significantly increased in anembryonic pregnancy. The over activation of decidual T cells during anembryonic pregnancy may thus contribute to the increased NK cytotoxicity activity. PMID- 10527386 TI - Tracking antigen-specific human T lymphocytes in rheumatoid arthritis by T cell receptor analysis. AB - The aim of this study was to use TCR sequencing as a tool to address the frequency of antigen specific T cells in different T cell compartments from a rheumatoid arthritis patient. We have previously established a clear link between T cell recognition of a specific Mhsp60 epitope and the amino acid sequence in the CDR3 region of the TCRB chain. This information was used to determine the frequency of these characteristic sequences in unmanipulated synovial fluid (SF), peripheral blood (PB) and hyperplastic lymph node of the same patient by amplification and sequencing. TCRBV sequences identical to those seen in antigen specific clones, and closely related sequences, were readily identified in SF, where they represented approximately 1% of all T cells, but were absent from PB or lymph node. The prevalence of putative Mhsp60 specific T cells within the SFMC is much greater than previously suggested by limiting dilution assays. Thus, amplification and sequencing may prove a superior technique for tracking the frequency of antigen-specific T cells in different tissues and in a longitudinal fashion. PMID- 10527387 TI - T cell repertoire in the liver of patients with autoimmune hepatitis. AB - Despite a large number of T cells infiltrating into the liver of patients with autoimmune hepatitis (AIH), little is known about their roles or target antigens. To investigate the roles of these T cells in the pathogenesis of AIH, we have studied the clonality of alphabeta T cell populations in liver tissue by size spectratyping the complementarity-determining region (CDR)3 size lengths of T cell receptor (TCR) Vbeta-chain transcripts. Analysis of nine AIH patients who had the HLA DR4 haplotype showed clonal expansion in all samples. More than two T cell clones expanded in most patients. Although the expression of the TCR Vbeta genes was different among the nine patients, clonal expansion of T cells expressing either TCR Vbeta2, 3, 4, 16, or 22 was observed in two patients or more. TCR Vbeta4 clones expanded in 5 cases. Cloning and sequencing of TCR Vbeta CDR3 from PCR products revealed no whole CDR3-shared clones among different patients. In conclusion, several T cell clonotypes first recognize target antigens, then expand and accumulate in the liver of AIH patients. These suggest heterogeneity of autoantigens and the complexity of AIH immunopathogenesis in individual patients. PMID- 10527389 TI - Antibodies to endothelial cells identify myocardial damage and predict development of coronary artery disease in patients with transplanted hearts. AB - BACKGROUND: Transplant-induced coronary artery disease is a leading cause of graft failure in cardiac allograft recipients after the first year of transplantation, but there presently is no test to identify patients at high risk for developing the disease. Our research is focused on development of a predictive test to identify patients at high risk of developing the disease. METHODS: Sixty-eight cardiac allograft recipients transplanted and followed at Methodist Hospital between 1982 and 1996 were studied. Serial annual angiograms were used to diagnose coronary artery disease, and serial endomyocardial biopsies were used to detect cellular infiltrates and microvascular disease. Biopsy matched serum samples were used for cardiac troponin-T determinations as measures of myocardial damage, and serum antibodies to endothelial cells were determined by using flow cytometry, enzyme-linked immunosorbent assay and immunoblotting techniques. The endothelial antibody data were evaluated statistically for associations with angiographic changes, biopsy findings and biochemical evidence of myocardial damage. FINDINGS: Antibodies to endothelial cells were identified by all three techniques, and significant associations were found for the amount of antibody identified by Western immunoblotting with histological rejection grades in biopsies, which were confirmed immunocytochemically as macrophages (p<0.01) and T lymphocytes (P = 0.03). These antibodies also associated significantly with vascular antithrombin depletion (p = 0.02), biochemical evidence of myocardial damage (p = 0.005) and subsequent development of coronary artery disease (p = 0.03). INTERPRETATION: The significant association of anti endothelial antibodies with cellular infiltrates, depletion of vascular antithrombin and myocardial damage suggests a role for antibody in the development of transplant-induced arteriopathy. The significant association of antiendothelial antibodies with the future development of coronary artery disease further suggests that assessment of these antibodies may provide a non-invasive test to predict the development of transplant-induced coronary artery disease. PMID- 10527388 TI - Complementation between HLA-DR4 (DRB1*0401) and specific H2-A molecule in transgenic mice leads to collagen-induced arthritis. AB - We generated transgenic mice with DRB1*0401 gene with mutation in the beta2 domain (aa 110 and 139) for better interaction with mCD4. The DR4 transgene was introduced into H2-Aq (B10RQB3) and H2-Af (B10RFB3) to examine the role of DR4 in collagen arthritis. The HLA-DR molecules in these mice were found to be functional on the basis of their positive/negative selection of the Vbeta T cell repertoire. H2-Aq mice are resistant to porcine CII-induced arthritis. The RQB3/DR4 mice (H2Aq/DR4) developed severe collagen induced arthritis (CIA) when immunized with Porcine type II collagen while the negative littermates were resistant. RQB3.DR4 mice were also highly susceptible to CIA induced by Human CII while negative littermates got only mild disease. However, RFB3/DR4 mice (H2Af/ DR4) did not get CIA with any type II collagen. Therefore, the DR4 gene in the context of H2-Aq predisposes to severe arthritis but not in the context of H2-Af. Antibodies to renatured cyanogen bromide (CB) cleaved fragments of PII in RQB3/DR4 mice and negative littermates suggest that the presence of DR4 does not result in any differences in specificity of antibody response to CB fragments. These results indicate that a specific gene complementation occurring between DR4 and H2.Aq but not DR4 and H2Af promotes the induction of arthritis with PII and HII in these mice. A similar interaction may be involved between DR and DQ molecules in human RA. PMID- 10527390 TI - Positivity in a modified mixed leukocyte reaction test correlates with molecular HLA-C disparity in prediction of unrelated bone marrow transplantation outcome. AB - The modified mixed leukocyte reaction (MMLR) test consists of the standard MLR (SMLR) test to which interleukin-4 (IL-4) has been added. It is a sensitive procedure capable of detecting alloreactivity not detected by the SMLR. In the present study we applied the MMLR test to unrelated bone marrow transplantation (BMT) in an attempt to predict graft versus host disease (GVHD) and graft rejection (GR) by detecting alloreactivity between recipient/donor pairs otherwise found to be fully matched (HLA class I A and B tested by serology; class II DRB1 and DQB1 by sequence specific oligonucleotide probes [SSOP]) and by studying the relationship of MMLR alloreactivity and HLA-C disparity in the prediction of transplant related complications. Thirty-five patients transplanted from unrelated donors were included in the study. The MMLR test was seen to correlate with the incidence of transplant related complications, as of the 19 positive, cases 12 (63%) developed acute GVHD and 7 (37%) GR, while of the 16 negative cases only 5 (31%) developed GVHD (4 acute, 1 chronic) (p = 0.0001) and 2 (12.5%) GR. No such correlation was seen between the SMLR and the incidence of transplant related complications: the SMLR test was positive in only 4 (11%) cases (all of which developed GVHD or GR) but of the 31 negative cases 22 (71%) also developed GVHD or GR. Reactivity in the MMLR also correlated with molecular HLA-C disparity (p = 0.015): While of the 19 positive cases 10 (53%) had molecular HLA-C disparity, of the 16 cases with negative MMLR, 14 (87.5%) were matched for molecular HLA-C. Two-way analysis confirmed that patients with positive MMLR transplanted from HLA-C mismatched donors were more likely to develop post BMT complications, including GVHD and GR, than patients with negative MMLR transplanted from HLA-C matched donors (r = +0.70) (p = 0.001). We conclude that the MMLR test may be a useful tool in the prediction of transplant related complications such as GVHD and GR, post unrelated BMT. Moreover, the MMLR test, in conjunction with molecular HLA-C typing, may improve unrelated donor selection. PMID- 10527391 TI - Allergens induce apoptosis in lymphocytes from atopic patients. AB - Repeated stimulation of immune cells may induce an "activation-induced cell death" (AICD) program. Allergy is characterized by the cyclic activation of allergen-reactive immune cells. To study the effects of allergen stimulation in cell proliferation and apoptosis in atopic subjects, peripheral blood mononuclear cells (PBL) from 40 atopic patients with positive reactivity to the allergens Olea Europaea (OE) and Lollium Perenne (LP) (20 without immunotherapy and 20 with specific immunotherapy) and 10 normal subjects were cultured with the allergens OE and LP. PBL from atopic patients proliferate more vigorously than cells from normal subjects after culture in vitro with both allergens, although PBL from atopic subjects without immunotherapy proliferate more than PBL from atopic subjects with immunotherapy. The study of cell proliferation shows that in atopic patients PBL mainly exhibit the CD4/CD45RO phenotype. This preferential proliferation is more evident in PBL from atopic patients treated without immunotherapy. Cell culture with specific allergens induces apoptosis in PBL from atopic patients. The percentage of apoptosis increased when atopic patients had been previously treated with immunotherapy. In addition to the observed increase in cell proliferation, apoptosis mainly occurs in the CD45RO cells that support the involvement of these cells in allergy. Furthermore, results obtained in cells from immunized patients suggest that an AICD process may partly at least explain the mechanism of action of allergen immunotherapy. PMID- 10527393 TI - Comparison of crossmatch results obtained by ELISA, flow cytometry, and conventional methodologies. AB - The complement-dependent lymphocytotoxicity crossmatch (CXM) is the presently accepted standard for detection of donor-reactive alloantibodies in transplant patients. However, the newer flow cytometric (FXM) and ELISA (EXM) crossmatch technologies are increasingly used as substitutes for the CXM. We have compared the sensitivity and reproducibility of FXM vs. EXM and, in general, find them to be quite similar. However, when we compared the agreement of FXM vs. EXM in 112 donor/recipient combinations, we found that they identified different subsets of donor-specific alloantibodies in about 35% of the tests. When compared to the standard CXM method, the EXM correlated much better than did the FXM, yielding a much lower rate of false positive (2.5% vs. 8%) and false negative (7% vs. 18.5%) results. The reduction in time required to obtain a result (3 h) and the cost of materials ($25/test) was identical for the EXM and FXM. We conclude that the ELISA method for crossmatching has advantages over the flow cytometric method as a substitute for the present standard complement-dependent lymphocytotoxicity method. PMID- 10527392 TI - Autoimmune markers and neurological complications in non-insulin-dependent diabetes mellitus. AB - To verify whether autoimmune markers related to nervous system structures and other autoimmunity indexes present in diabetes mellitus are associated with subclinical neuropathy, we examined 48 non-insulin-dependent diabetic patients with and without neuroelectrophysiological alterations. Nerve conduction velocity at the external sciatic-popliteal nerve, at the sural nerve, at the median and ulnar nerves level has been evaluated. Autoimmunity was investigated by evaluating glutamic acid decarboxylase (GAD-Ab), insulin (IAA), GM3, GD3 and GT1b gangliosides, pancreatic islet cell (IC-A) and anti-nervous-tissue autoantibody presence. Nerve conduction velocities were decreased in 72.9% of diabetic patients. Anti-insulin antibodies were detected in seven non-insulin created diabetic patients and in higher amount in subjects with (17.1%) than in those without (7.7%) asymptomatic neuropathy. Anti-GM3 antibodies were detected in four diabetic patients all of whom presented neurological complication. A significant correlation has been found between neurological damage and presence of anti insulin antibodies (p<0.05). In the case of GM3 autoantibody, a similar result was obtained, but the data failed to reach statistical significance. Our data demonstrate that autoimmunity might play a role in the development of peripheral neuropathy. PMID- 10527394 TI - Matching for TNF microsatellites is strongly associated with matching for other non-HLA MHC sequences in unrelated bone marrow donor-recipient pairs. AB - The use of unrelated donors for bone marrow transplantation is associated with an increased morbidity and mortality when compared with HLA identical siblings. We have demonstrated previously that matching of unrelated donors and recipients for TNFa microsatellites is correlated with lower CTLp frequencies. Matching of unrelated donors and recipients for other non-HLA sequences in the major histocompatibility complex has been reported to result in less graft-versus-host disease and improved survival. It has been argued that matching for non-HLA sequences in the MHC in addition to the HLA genes themselves results in matching for the entire MHC and is therefore the equivalent of providing an HLA identical sibling donor. In order to test this hypothesis we have examined TNFa microsatellites of unrelated donor recipient pairs in whom matching for HLA loci, non-HLA sequences near HLA B (beta-block markers) and non-HLA sequences near DRB1 (delta-block markers) had been determined. All 17 patients who were matched for HLA and non-HLA markers were also matched for TNF microsatellites. This data supports the idea that matching for HLA genes and non-HLA markers results in matching at all other loci in the MHC. PMID- 10527395 TI - The influence of T cell receptor and cytokine genes on sarcoidosis susceptibility in African Americans. AB - The pathogenesis of sarcoidosis, a multisystem granulomatous disorder, is mediated through immunoregulatory pathways. While sarcoidosis clusters in families, inherited risk factors remain undefined. In search of possible sarcoidosis susceptibility genes, we examined anonymous polymorphic genetic markers tightly linked to six different candidate gene regions on chromosomes 2q13, 5q31, 6p23-25, 7p14-15, 14q11 and 22q11. These candidate regions contain T cell receptor, interleukin (IL) and interferon regulatory factor (IRF) genes. Our study population consisted of 105 African-American sarcoidosis cases and 95 unrelated healthy controls. The allelic frequency distribution of two out of the six markers, IL-1 alpha marker (p = 0.010) on 2q13 and the F13A marker (p = 0.0006) on 6p23-25, was statistically significantly different in cases compared with controls. The two alleles most strongly associated with sarcoidosis were IL 1 alpha*137 (Odds Ratio (OR) = 2.60; 95% confidence interval (CI) = 1.36-4.98) and F13A*188 (OR = 2.42; 95% CI = 1.37-4.30). Individuals that had both of these alleles were at a six-fold increased risk for sarcoidosis (OR = 6.19; 95% CI = 2.54-15.10). Restricting the analysis to cases with at least one first or second degree relative affected with sarcoidosis increased the OR to 15.38. IL-1 levels are elevated in sarcoidosis and the F13A marker is tightly linked to a gene that codes for a newly identified interferon regulatory factor protein (IRF-4), which is thought to play a role in T cell effector functions. Our results suggest genetic susceptibility to sarcoidosis may be conferred by more than one immune related gene that act synergistically on disease risk. PMID- 10527397 TI - IDDM9 and a locus for rheumatoid arthritis on chromosome 3q appear to be distinct. AB - Markers near a locus for type 1 diabetes on chromosome 3q22-q25 (IDDM9) demonstrate linkage to rheumatoid arthritis, however it is not clear whether these two loci overlap. Sex-specific linkage analysis may be of interest for rheumatoid arthritis on chromosome 3q since linkage of type 1 diabetes to IDDM9 derives predominantly from affected female sibpairs, and rheumatoid arthritis is more common in females than males. Using data from a recent genome scan for rheumatoid arthritis and sex-specific linkage analysis we show that linkage of rheumatoid arthritis to chromosome 3q peaks approximately 30 cM centromeric to IDDM9. Furthermore, there is no evidence for linkage to IDDM9 in females with rheumatoid arthritis. PMID- 10527396 TI - Strong association of HLA class II sequences in Mexicans with Vogt-Koyanagi Harada's disease. AB - Vogt-Koyanagi-Harada's syndrome (VKH) is an autoimmune disease prevalent in Mongoloids with evident participation of HLA. The aim of this study was to identify the class II DNA sequences involved in the etiopathogenesis of VKH in Mexican Mestizos. This study included 46 VKH patients and 170 controls. 75% were females (mean age at onset of 33.5 years). The disease evolved to chronicity (68%) and 25% of the patients were unresponsive to corticotherapy. DNA typing of HLA-DRB1, DQA1 and DQB1 was done following the 12th International Histocompatibility protocols. VKH was strongly dependent of DRB1 gene; DRB1*04 was found in 78.2% of the patients vs. 50.6% of the controls (p = 0.001). No particular DRB*04 subtype was significantly increased, suggesting that residues E 9 V-11; H-13; H-33 and Y-37 shared by all DR4s are implicated in susceptibility to VKH. However DRB1*0101 (p = 0.009, OR = 4.2) was clearly associated. This allele shares the motif LLEQRRAAG located at position 67-74 and 86 of DRB1 with *0405 associated in Japanese. Two HLA associated mechanisms may be triggering the autoimmune phenomena. One involving critical polymorphic residues expressed in different alleles. Secondly, some peptides may anchor to the conserved residues leaving other sequences to bind to the T cell receptor. PMID- 10527398 TI - HLA-DMB gene and HLA-DRA promoter region polymorphisms in Australian multiple sclerosis patients. AB - The MHC region has been shown to contain a susceptibility locus for multiple sclerosis (MS). While the strongest association to date has been between HLA DRB1*1501 and MS, the exact nature of the MHC association in MS remains unclear. Two candidate polymorphic loci within the MHC class II region, the HLA-DMB gene and the HLA-DRA promoter, which lie close to HLA-DRB1, were therefore examined in an Australian MS population. The HLA-DMB*0103 phenotype was increased in the MS patients (46% vs. 30%) and the frequency of the HLA-DRA promoter A allele was also increased (81% vs. 68%). When the subjects were stratified into HLA-DRB*1501 positive and negative individuals these associations were not significantly different. This is a result of the strong linkage disequilibrium between HLA DRB*1501 and both HLA-DMB*0103 and the HLA-DRA promoter A allele. The complete linkage between DRB1*1501 and the HLA-DRA promoter A allele indicates that the MS susceptibility haplotype (DRB1*1501-HLA-DQB1*0602-HLA-DQA1* 0102) can be extended out to promoter of the HLA-DRA locus. Interactions between both HLA-DMB and the HLA-DRA promoter and other reported MS susceptibility loci were examined (TCRBV polymorphisms, HLA-DQA1 and HLA-DQB1). Some interactions between specific TCRBV polymorphisms and the HLA-DRA promoter were observed, which is consistent with other published reports suggesting an epistatic interaction between TCRBV and HLA DRB1. PMID- 10527399 TI - HLA microsatellite associations with insulin-dependent diabetes mellitus in Singaporean Chinese. AB - Singaporean Chinese with insulin-dependent diabetes mellitus (IDDM) have previously been shown to be associated with the DRB1*0301 haplotype and the joint occurrence of DRB1*0301/*0901 and DRB1*0301/*04. The present study extended previous HLA associations by investigating the HLA region using four microsatellites (TNFa, D6S273, TAP1, DQCARII). Seventy-five IDDM patients and 80 healthy controls were studied. TNFa*3 (RR = 2.26), TNFa*12 (RR = 3.30), TAP1*9 (RR = 2.55) showed increased frequencies while TNFa*11 (RR = 0.29), TAP1*4 (RR = 0.50) showed decreased frequencies in patients compared to controls. Linkage analysis suggested that the positive associations of TNFa*3 and TAP1*9 were secondary to that of DRB1*0301. However, TNFa*12 appeared to provide additional risks to IDDM besides the DRB1*0301 haplotype, whereas TNFa*11 and TAP1*4 conferred an independent protective effect against IDDM. Our findings reinforce the notion that susceptibility to and protection against IDDM may include TNF region. In the present study, TNFa*12 seemed to be the primary association in the DRB1*0405 haplotype and may play an independent role in the pathogenesis of IDDM through TNF-alpha function. PMID- 10527400 TI - High resolution HLA-DQB1 typing by combination of PCR-RFLP and PCR-SSCP. AB - A reliable method for high-resolution HLA-DQB1 typing using a combination of PCR- restriction fragment length polymorphism (RFLP) and PCR-single strand conformation polymorphism (SSCP) analysis is described. The second exon of the DQB1 gene was subjected to PCR using generic primers and digested with two restriction enzymes, MspA1I and HaeIII, and the DQB1 alleles were divided into seven groups. According to the RFLP patterns, appropriate group specific primers for DQ5, 6 and DQ2, 3, 4 groups were used to selectively amplify the alleles and the SSCP technique was used to distinguish the individual alleles. A total of 88 quality control samples of various ethnic groups distributed in the International Cell Exchange and HLA DNA Exchange programs and the ASHI/CAP Proficiency Tests were investigated by the PCR-RFLP/SSCP method. The concordance between our typing results and the consensus results of the surveys were 100%, and a total of 14 DQB1 alleles in 49 homozygous and heterozygous combinations were all correctly identified by the method described. This method is accurate, economical and relatively easy to interpret and well suited for routine clinical and research uses. PMID- 10527401 TI - A susceptibility region for myasthenia gravis extending into the HLA-class I sector telomeric to HLA-C. AB - We have analyzed a series of HLA region markers in 207 UK Caucasoids with early onset myasthenia gravis (EOMG, onset before age 40), where there is a strong female bias. The well known associations with HLA-DR3 and -B8 have now proved to be significantly stronger in the 165 females than in the 42 males. In patients (of either sex) lacking -DR3, there was also a significant increase in HLA-DR2. Although the muscle weakness in EOMG is clearly mediated by autoantibodies, the associations are consistently stronger with HLA-B8 (in class I) than with HLADR3 (in class II), as confirmed here. We therefore typed 87-137 cases for polymorphisms at four loci in the intervening class III region, and also at three in the adjacent stretch of class I. At each locus, one allele tended to co-occur with HLA-B8 and showed strong and highly significant associations in the patients. There appeared to be a region of maximal susceptibility extending from HSP70 (in class III) past HLA-B and HLA-C at least 600 kb telomerically into the class I region, which is now being mapped in detail. Any candidate genes here that act shortly after puberty may allow more precise localization of susceptibility. PMID- 10527402 TI - Nomenclature for factors of the HLA system, update April 1999. WHO Nomenclature Committee for factors of the HLA system. PMID- 10527403 TI - Nomenclature for factors of the HLA system, update May 1999. WHO Nomenclature Committee for factors of the HLA system. PMID- 10527404 TI - Antiplatelet factor 4--heparin antibodies in patients with antiphospholipid antibodies. AB - Antibodies directed against platelet factor 4-heparin are present in patients with heparin-induced thrombocytopenia (HIT). Additionally, it has been suggested that heparin can be an antigenic target of antiphospholipid antibodies (aPL). We investigated the presence of heparin-platelet factor 4-induced antibodies (HPIA) in 33 patients with aPL. There were 30 patients with lupus anticoagulant, 25 with anticardiolipin antibodies, 21 with anti-beta2 glycoprotein I, and 18 with antiprothrombin antibodies. 20 patients had a history of thrombosis and 19 had received heparin during the last 60 months. We found 7 (21.2%) who had HPIA; 5 of them also had anti-beta2 glycoprotein I antibodies. Four patients had severe thrombocytopenia and suspicion of HIT. Among them, two presented high positive HPIA results, one of them with positive platelet aggregation test. The third patient showed grey zone HPIA and borderline aggregation test and the fourth one had negative results. Among patients without a history of HIT, 2 who had never received heparin presented high positive, one a moderate positive, and one a grey zone HPIA result; all of them with negative aggregation tests. Five positive sera samples were incubated with cardiolipin liposomes in the presence of beta2 glycoprotein I, and whereas an inhibition greater than 50% was achieved in anticardiolipin and anti-beta2 glycoprotein I activities, HPIA results did not change. We demonstrate that HPIA could be frequently found in patients with aPL. They are responsible for HIT in some cases but can also be found in patients who have not received heparin. Whether they predispose patients with aPL to HIT is not known; nevertheless, a close follow-up of heparin treatment in these patients seems to be mandatory. PMID- 10527405 TI - Treatment of hyperhomocysteinemia with folic acid and vitamins B12 and B6 attenuates thrombin generation. AB - The effect of homocysteine-lowering treatment on thrombin generation was investigated in 17 subjects with hyperhomocysteinemia (aged 22-60 years), 11 of whom had symptomatic atherosclerotic vascular disease. All subjects had fasting total homocysteine levels above 16 micromol/L. The formation of thrombin was assessed by measuring thrombin-antithrombin III complexes and prothrombin fragment 1+2 in peripheral venous blood and in the bleeding time blood collected at 30-second intervals from skin incisions on a forearm. All the tests were performed before and after an 8-week treatment with folic acid p.o. 5 mg/day, vitamin B6 p.o. 300 mg/day, and vitamin B12 i.m. 1000 microg given on a weekly basis. Following the 8-week therapy, the median plasma homocysteine concentration became significantly reduced from 20 to 10 micromol/L, while plasma levels of fibrinogen, prothrombin, and antithrombin III as well as activity of protein C, S, and factor VII showed no changes. Vitamin treatment was associated with a significant fall in thrombin-antithrombin III complexes and prothrombin fragment 1+2 concentrations in peripheral venous blood. Bleeding time became prolonged by about 60 seconds. At sites of hemostatic plug formation, plasma concentrations of both thrombin markers significantly decreased. Compared with pretreatment values, significantly less thrombin was produced during the first 3 minutes of bleeding after homocysteine-lowering therapy. In subjects with hyperhomocysteinemia a reduction of plasma fasting homocysteine concentration by folic acid and vitamins B12 and B6 administration is associated with attenuation of thrombin generation both in peripheral blood and at sites of hemostatic plug formation. PMID- 10527406 TI - Changes of platelet surface antigens in patients suffering from abdominal septic shock. AB - Sepsis and related syndromes account for a high morbidity and mortality caused by the development of multiorgan failure. Pathogenesis of sepsis is complex, involving humoral as well as cellular factors. Since the role of platelets is still undefined in this concern, we investigated CD63, CD62P, CD36, and CD31 expression on platelets of patients in septic shock (n = 18) using a flow cytometric assay in whole blood. Samples were drawn within 24 hours of onset. We found thrombocytopenia accompanied by a significantly higher expression of CD63, CD62P, and CD31 and a significant downregulation of CD36 in comparison to healthy volunteers (n = 18). Changes in CD63 and CD62P expression indicates platelet activation. Because CD62P, CD36, and CD31 mediate interaction of platelets with leukocytes, subendothelial matrix and probably endothelial cells as well as platelet adhesion/aggregation, our findings suggest an involvement of platelets in leukocyte/endothelial cell interaction in septic shock. We suspect that thrombocytopenia is not due to bone marrow depression, but rather is due to consumption of highly activated platelets in the microcirculation. We feel that our observations may offer a rationale for potentially beneficial effects of antiplatelet therapy in sepsis; however, further studies have to evaluate its beneficial impact as well as its potential risk for bleeding complications. PMID- 10527407 TI - Phenotypic consequence of the gene abnormality in the platelet glycoprotein IX gene observed in a patient with Bernard-Soulier syndrome through mammalian cell expression system. AB - We previously reported a patient with Bernard-Soulier syndrome (BSS) who had a homozygous missense mutation in the GPIX gene Phe55 (TTT) to Ser (TCT) replacement in the leucine-rich motif (LRM) of the GPIX polypeptide, as a probable cause of BSS phenotype. To study the effect of this mutation on the surface expression of the GPIb/IX complex and GPIX, we introduced this mutation into the cDNA of GPIX by site-directed mutagenesis and performed in vitro transfection studies with plasmid for mutant GPIX and other plasmids for GPIb/IX complex. Mutant GPIX could not increase the surface expression of GPIb-alpha, but also surface expression of GPIX itself. Immunostaining of the CHO-K1 cells transfected with two plasmids for mutant GPIX and GPIb-beta showed that mutant GPIX was weakly but certainly detected in the cytoplasm of the transfected cells. These findings indicate that this substitution is responsible for BSS phenotype and that the LRM of GPIX is a critically important element for an efficient expression of the GPIb/IX complex. They also suggest that the failure of GPIb/IX complex expression can be caused by a mechanism other than synthetic defect of the mutant protein. PMID- 10527408 TI - Effects of ethanol on platelet responses associated with adhesion to collagen. AB - Adhesion of platelets to collagen in damaged blood vessels or ruptured atherosclerotic plaques is important in hemostasis and arterial thrombosis. Adhesion to collagen results in secretion of granule contents and formation of thromboxane A2; thromboxane A2 and released ADP synergistically promote aggregation around platelets adherent to collagen. Ethanol inhibits collagen induced platelet aggregation, secretion, arachidonate mobilization, and thromboxane A2 formation but does not inhibit platelet adhesion to de endothelialized rabbit aortae. We investigated whether ethanol affects the initial signalling events and responses of platelets adherent to collagen, independent of the actions of secondary agonists. Suspensions of washed human platelets, labelled by incorporation of [3H]oleate into phospholipids, were used to measure platelet adhesion to collagen by a filtration method; studies were done in the presence of an ADP-removing system and blockers of receptors for thromboxane A2, platelet-activating factor, serotonin, and fibrinogen. Ethanol (87 mM) did not affect the rate or extent of platelet adhesion to collagen or secretion of [14C]serotonin from prelabelled platelets adherent to collagen, but ethanol did inhibit thromboxane A2 formation. Previous studies showed that ethanol does not affect platelet stimulation by arachidonate, leading to the suggestion that reduced mobilization of arachidonate, rather than inhibition of its conversion to thromboxane A2, is responsible for inhibition by ethanol of thromboxane A2 formation. Here, we show by a gel mobility shift assay and immunoblotting, that ethanol delays the collagen-induced increase in the phosphorylation of cytosolic phospholipase A2, the enzyme responsible for arachidonate mobilization. However, ethanol has no effect on collagen-induced tyrosine phosphorylation of phospholipase Cgamma2, determined by immunoprecipitation and immunoblotting. Thus, ethanol's effect on signal transduction in collagen-adherent platelets occurs distal to phosphorylation of phospholipase Cgamma2 but proximal to phosphorylation of cytosolic phospholipase A2. PMID- 10527410 TI - Evidence of a phospholipid binding species within human fibrinogen preparations. AB - Fibrinogen has been reported to interact with phospholipid; however, the properties of this binding interaction have not been characterized. Purified preparations of human fibrinogen bound to small unilamellar vesicles containing phosphatidylserine (PS) as measured by light scattering and radioisotope filtration. Binding to 100% PS was saturable (apparent Kd=5 microM, Bmax=1.9 g protein/g lipid), reversible, and involved a minor subfraction of the fibrinogen preparation (3-6% of total protein). Fibrinogen interacted minimally with phosphatidylinositol, and not at all with pure phosphatidylcholine (PC) or PC vesicles containing 5% glycosphingolipid (lactosylceramide, ganglioside GM3, ganglioside GD3). Binding efficiency decreased as the PS content of vesicles was diluted with PC. Calcium chloride (2 mM) enhanced protein binding to PS, which was reversed by EDTA. Fibrin clot formation almost quantitatively precipitated the PS binding activity. PS, but not PC, increased the final turbidity of fibrin clots. Computerized sequence analysis of fibrinogen revealed three candidate acidic phospholipid binding motifs located at position 143-210 in the alpha chain, and positions 59-77 and 101-139 in the beta chain. Further study of the PS binding activity of fibrinogen may lead to new insights about fibrinogen function. PMID- 10527409 TI - Effects of cathepsin G pretreatment of platelets on their subsequent responses to aggregating agents. AB - Cathepsin G, a proteolytic enzyme from activated leukocytes, can interact with platelets during inflammation and thrombosis. Platelets that have been exposed to cathepsin G in thrombi may recirculate if they are freed during fibrinolysis. To determine whether some of the subsequent functions of such platelets would be impaired, we investigated the responses of cathepsin G-pretreated platelets to agonists that they would encounter in the circulation. Suspensions of washed human platelets were labeled with [14C]serotonin and resuspended in Tyrode albumin solution (with 2 mM Ca2+ and apyrase). After 15 minute incubation with 400 nM cathepsin G at 37 degrees C, 52+/-3% of [14C]serotonin had been released, and glycoprotein Ib was degraded. The platelets were washed and resuspended in fresh medium to remove cathepsin G and released materials. Ristocetin-induced agglutination was abolished, indicating that the binding site for von Willebrand Factor on glycoprotein Ib had been removed. Aggregation and release of residual [14C]serotonin in response to 0.1-1.0 U/mL thrombin was blocked or greatly reduced by the cathepsin G pretreatment. This inhibition is probably largely due to cleavage by cathepsin G of some of the protease-activated receptors at the C terminal side of Ser42 so that the tethered ligand is lost. Pretreatment with cathepsin G did not affect responses to ADP or a low concentration of platelet activating factor in the presence of fibrinogen, indicating that receptors for these agonists were unaffected and that the function of the fibrinogen receptor, GPIIb/IIIa was unchanged. Responses to cathepsin G, the thrombin receptor activating peptide SFLLRN, collagen, or the thromboxane A2 mimetic U46619 were partially inhibited, even in the presence of added fibrinogen. Platelet adhesion to a collagen-coated surface was 51+/-7% inhibited, which may indicate cleavage of a collagen receptor or receptors; this may partly account for strong inhibition of collagen-induced aggregation and release of granule contents; additionally, as shown by inhibition of responses to U46619, the function of the thromboxane A2 receptor may be compromised. Thus, although cathepsin G activates platelets, if they recirculate after interaction with it, their subsequent adhesion to damaged vessel walls, aggregation, and release of granule contents induced by thrombin and collagen will be diminished. PMID- 10527411 TI - Orally administered acetylsalicylic acid decreases protein incorporation into the cytoskeleton of thrombin-stimulated platelets. PMID- 10527412 TI - Role of secreted adenosine diphosphate in the synergistic effects of cathepsin G on human platelets. PMID- 10527413 TI - Recurrent multiple thrombosis in a patient with abnormal plasminogenemia and Behcet's disease. PMID- 10527414 TI - Relationship between venous thromboembolism and lipid or lipoprotein disorders. PMID- 10527415 TI - LINEs, SINEs and repetitive DNA: non-LTR retrotransposons in plant genomes. AB - Retroelements and remnants thereof constitute a large fraction of the repetitive DNA of plant genomes. They include LTR (long terminal repeat) retrotransposons such as Ty1-copia and Ty3-gypsy retrotransposons, which are widespread in plant genomes and show structural similarity to retroviruses. Recently, non-LTR retrotransposons, lacking the long terminal repeats and subdivided into LINEs (long interspersed nuclear elements) and SINEs (short interspersed nuclear elements), have been discovered as ubiquitous components of nuclear genomes in many species across the plant kingdom. LINEs are probably the most ancient class of retrotransposons in plant genomes, but the evolutionary borders between non LTR retrotransposons, LTR retrotransposons and retroviruses are indistinct as shown by the detection of intermediate forms in other eukaryotic taxa. Transposition of non-LTR retrotransposons is only rarely observed in plants indicating that the majority of these retroelements are inactive and/or under regulation of the host genome. Transposition is poorly understood, but experimental evidence from other genetic systems, in particular from insect and mammalian species, shows that LINEs are able to transpose autonomously, while non autonomous SINEs depend on the reverse transcription machinery of other retrotransposons. Fluorescence in situ hybridization demonstrated that different classes of retrotransposons differ largely in their chromosomal organization and are often excluded from blocks of rapidly homogenizing tandem repeats. In particular, LINEs contribute considerably to the repetitive DNA of nuclear plant genomes. PMID- 10527416 TI - Differential expression of rice alpha-amylase genes during seedling development under anoxia. AB - The unique capability of rice (Oryza sativa L.) seedlings to grow under anoxic conditions may result in part from their ability to express alpha-amylase and maintain the supply of sugar needed for energy metabolism. Previous studies have demonstrated that under aerobic conditions the Amy1 and Amy2 subfamily genes are regulated primarily by phytohormones while the Amy3 subfamily genes are induced during sugar starvation. The expression patterns for these alpha-amylase genes were considerably different in anoxic vs. aerobic rice seedlings. The level of total alpha-amylase mRNA under anoxic conditions was decreased in aleurone layers while it increased in the embryo. Anoxic conditions greatly diminished the expression of the Amy1A gene in aleurone. Conversely, expression of many Amy3 subfamily genes was up-regulated and prolonged in embryo tissues under anoxic conditions. PMID- 10527417 TI - Elicitor- and A23187-induced expression of WCK-1, a gene encoding mitogen activated protein kinase in wheat. AB - Wheat cultured cells were used to study the role of Ca2+ in regulating protein kinases during the induction of defense-related genes by fungal elicitor treatments. Manipulation of intracellular Ca2+ concentrations by treatment with calcium ionophore A23187 in the presence of high extracellular Ca2+ resulted in the induction of mRNA expression of WCK-1, a gene encoding mitogen-activated protein (MAP) kinase. The induction of WCK-1 mRNA by A23187 did not occur when extracellular Ca2+ was chelated by 1,2-bis(2-aminophenoxy)ethane-N,N,N',N' tetraacetic acid (BAPTA). The WCK-1 mRNA was also induced by Typhula ishikariensis-derived elicitors, suggesting a possible involvement of WCK-1 in the plant defense response against pathogens. BAPTA and a calcium channel blocker, La3+, inhibited the elicitor-induced expression of the WCK-1 mRNA. A recombinant fusion protein of WCK-1 (GST-WCK-1) autophosphorylated at the Tyr residue and exhibited an autophosphorylation-dependent protein kinase activity towards myelin basic protein. Alteration of Tyr-196 in the conserved 'TEY' motif in GST-WCK-1 to Phe by site-directed mutagenesis abolished the autophosphorylation. The GST-WCK-1 protein was activated by elicitor-treated wheat cell extracts but not by the control extract. These results suggest that fungal elicitors activate WCK-1, a specific MAP kinase in wheat. Furthermore, the results suggest a possible involvement of Ca2+ in enhancing the MAP kinase signaling cascade in plants by controlling the levels of the MAP kinase transcripts. PMID- 10527418 TI - Negative effect of the 5'-untranslated leader sequence on Ac transposon promoter expression. AB - Transposable elements are used in heterologous plant hosts to clone genes by insertional mutagenesis. The Activator (Ac) transposable element has been cloned from maize, and introduced into a variety of plants. However, differences in regulation and transposition frequency have been observed between different host plants. The cause of this variability is still unknown. To better understand the activity of the Ac element, we analyzed the Ac promoter region and its 5' untranslated leader sequence (5' UTL). Transient assays in tobacco NT1 suspension cells showed that the Ac promoter is a weak promoter and its activity was localized by deletion analyses. The data presented here indicate that the core of the Ac promoter is contained within 153 bp fragment upstream to transcription start sites. An important inhibitory effect (80%) due to the presence of the 5' UTL was found on the expression of LUC reporter gene. Here we demonstrate that the presence of the 5' UTL in the constructs reduces the expression driven by either strong or weak promoters. PMID- 10527419 TI - A leucine-rich repeat receptor-like protein kinase (LRPKm1) gene is induced in Malus x domestica by Venturia inaequalis infection and salicylic acid treatment. AB - A cDNA clone encoding a leucine-rich repeat (LRR) receptor-like protein kinase (LRPKm1) of Malus x domestica cv. Florina has been isolated using as a heterologous probe a cloned gene encoding a polygalacturonase-inhibiting protein (PGIP) of Phaseolus vulgaris L. A genomic clone containing the 5'-regulatory region and a 5' portion of the open reading frame of the LRPKm1 gene has also been isolated. An open reading frame of 2997 nt (999 amino acids) was present in the cDNA clone, encoding a receptor-like protein comprising a 21 amino acid signal peptide for secretion, a leucine zipper, 23 LRRs, a putative membrane spanning region and a serine/threonine protein kinase domain. LRPKm1 shows homology to the A. thaliana receptor-like protein kinase RLK5 and, to a minor extent, to PGIP. The LRPKm1 region from +5 to +600 exhibits an alternative reading frame that encodes a product corresponding to a proline-rich protein fragment homologous to several hydroxyproline-rich proteins. Southern blot analysis showed that LRPKm1 belongs to a multigene family and that there is length polymorphism of the hybridizing restriction fragments among different M. x domestica cultivars. Northern blot analysis was carried out on mRNA extracted from infected leaves of either cv. Florina (resistant to Venturia inaequalis) or cv. Golden Delicious (susceptible to V. inaequalis), and from tissues treated with salicylic acid. A 3500 bp transcript hybridizing at high stringency with the LRPKm1 cDNA accumulated in response to infection or salicylic acid treatment. Transcript accumulation was more intense in the incompatible interaction than in the compatible one. The possible involvement of this receptor-like protein kinase in resistance of apple to phytopathogenic fungi is discussed. PMID- 10527420 TI - The mat-r open reading frame is transcribed from a non-canonical promoter and contains an internal promoter to co-transcribe exons nad1e and nad5III in wheat mitochondria. AB - The expression of the mat-r locus (mat-r-nad1e-nad5III) was studied in wheat mitochondria. Transcription initiation sites were mapped by S1 protection, primer extension and capping experiments. Two different transcription initiation sites were found. One, non-canonical promoter of low expression level generates a transcript containing the complete mat-r open reading frame (orf), suggesting that this form is the maturase-reverse transcriptase mRNA. A second transcription initiation site, found within the coding region of the mat-r orf, directs the transcription of an abundant co-transcript containing the carboxy-terminal region of the mat-r orf, exon e of the nad1 gene, exon III of the nad5 gene and their respective trains-introns. The co-transcript promoter carries the consensus motif of plant mitochondrial promoters. Analysis of transcript sequences reveals the presence of editing sites in analogous positions in both nad1e and nad5III trans introns, suggesting that RNA editing is necessary for the trans-splicing process. PMID- 10527421 TI - A cDNA from tobacco codes for an inhibitor of virus replication (IVR)-like protein. AB - We have shown previously that localization of tobacco mosaic virus (TMV) in tobacco is associated with a ca. 23 kDa protein that inhibits replication of several plant viruses. This protein, named 'inhibitor of virus replication' (IVR), was purified from the medium of TMV-inoculated protoplasts derived from Nicotiana tabacum cv. Samsun NN. IVR was shown to be present also in induced resistant leaf tissue of N. tabacum cv. Samsun NN. We prepared an expression cDNA library from such induced-resistant tissue and screened it with a polyclonal antibody raised against the IVR protein. A 1016 bp clone (named NC330) containing a 597 bp open reading frame, coding for a 21.6 kDa polypeptide, was isolated. The NC330 clone hybridized with RNA from induced-resistant tissue from N. tabacum cv. Samsun NN but not with RNA from non-induced tissue. Likewise, it did not hybridize with RNA from infected or uninfected tissue of N. tabacum cv. Samsun nn. Similarly, the NC330 cloned probe hybridized with the RT-PCR products from RNA of the induced-resistant tissue only. In Southern blot hybridization the NC330 DNA probe detected several genomic DNA fragments in both N. tabacum cv. Samsun NN and Samsun nn. The size of the DNA fragments differed in Samsun NN and Samsun nn. We suggest that DNA encoding the IVR-like protein is present in resistant and susceptible N. tabacum genotypes, but is expressed only in NN. We have inserted the NC330 into the expression vector pET22b and a 21.6 kDa protein was produced in Escherichia coli that reacted in immunoblots with the IVR antibody. This protein greatly reduced replication of TMV in N. tabacum cv. Samsun nn leaf disk assays. PMID- 10527422 TI - Construction and characterization of a common bean bacterial artificial chromosome library. AB - We have constructed a common bean (Phaseolus vulgaris L.) bacterial artificial chromosome (BAC) library consisting of 33,792 clones and an estimated 3- to 5 fold coverage of the common bean genome. Leaf nuclei were used as the source for high-molecular-weight DNA, and an endonuclease/methylase competition assay was employed to partially cleave the DNA. The library was screened with a number of nuclear and mitochondrial probes. Each nuclear probe identified at least two BACs with an average insert size of ca. 100 kb. Only 26 clones were identified after hybridizing with mitochondrial probes, indicating contamination with organellar sequences is low. Numerous clones could be identified after screening the library with two repetitive probes flanking the nuclear fertility restorer Fr. Intriguingly, 12 clones appeared to hybridize to both markers, and restriction analysis of these clones revealed that they can be assembled into maximally four contigs, suggesting that these repetitive probes may be useful for the physical mapping of the Fr locus. PMID- 10527423 TI - Expression of two PIP genes in rapidly growing internodes of rice is not primarily controlled by meristem activity or cell expansion. AB - Membrane intrinsic proteins facilitate movement of small molecules often times functioning as water channels. We have identified two genes from rice which encode proteins with characteristic features of plasma membrane intrinsic proteins (PIP). They possess six membrane-spanning domains, an NPA repeat, overall high sequence homologies and characteristic C- and N-terminal hallmark motifs which allowed assignment of OsPIP1a to the PIP1 subfamily and of OsPIP2a to the PIP2 subfamily. OsPIP1a and OsPIP2a showed similar but not identical expression patterns. The two genes were expressed at higher levels in seedlings than in adult plants and expression in the primary root was regulated by light. In internodes of deepwater rice plants which were induced to grow rapidly by submergence, transcript levels were slightly induced in the intercalary meristem (IM) and slightly reduced in the elongation zone (EZ) after 18 h. In internodes of GA-induced excised stem sections transcript levels transiently declined in the IM and EZ after 1 h and subsequently recovered to elevated levels after 18 h. GA also induced OsPIP expression in non-growing tissue after 18 h. In the IM of submergence-induced stem sections transcript levels remained constitutive. The different growth-promoting treatments showed no direct correlation between growth rate and OsPIP gene expression in dividing or expanding cells. In fact, treatment of excised stem sections with ABA or drought stress induced similar changes in OsPIP expression in the growing zone during the first 6 h as GA did. We conclude that regulation of OsPIP1a and OsPIP2a expression is not primarily controlled by growth. GA-induced growth may however change the water status of cells which in turn results in altered PIP abundance. PMID- 10527424 TI - Dimerisation of maize glutathione transferases in recombinant bacteria. AB - Two cDNAs encoding novel type III maize (Zea mays) GST subunits, ZmGST VI and ZmGST VII, have been cloned in addition to the previously described ZmGST V. Together with the type I GSTs ZmGST I and ZmGST III, these subunits were expressed in Escherichia coli, both individually and in tandem combinations using a customised pET vector. The GST dimers formed were then characterised. When type I GSTs were co-expressed only the respective homodimers were formed rather than the ZmGST I-III heterodimer. The failure to form this heterodimer, together with the negligible herbicide-detoxifying activity associated with recombinant ZmGST III-III, suggests that the identity of herbicide-detoxifying isoenzymes described in maize as being composed of ZmGST III subunits requires re-evaluation. In contrast, co-expression of the type III GSTs ZmGST V and ZmGST VI resulted in the formation of ZmGST V-V, ZmGST VI-VI and ZmGST V-VI dimers in the ratio 1:1:2 as predicted for random subunit association. ZmGST V-VI had kinetic characteristics intermediate between those of the two homodimers, indicating that the subunits were catalytically independent of one another. Co-expression of ZmGST V and ZmGST VII resulted in the formation of ZmGST V-VII and this isoenzyme was subsequently identified in maize plants. Attempts to dimerise type I GST subunits with type III GST subunits proved unsuccessful. These results demonstrate the utility of co expressing recombinant GSTs to explore the potential of subunit-subunit associations and to help unravel the complexity of homodimeric and heterodimeric GSTs in plants. PMID- 10527425 TI - Methyl jasmonate induces expression of a novel Brassica juncea chitinase with two chitin-binding domains. AB - We have cloned a 1.3 kb Brassica juncea cDNA encoding BjCHI1, a novel acidic chitinase with two chitin-binding domains that shows 62% identity to Nicotiana tabacum Chia1 chitinase. BjCHI1 is structurally unlike Chia1 that has one chitin binding domain, but resembles Chia5 chitinase UDA1, the precursor of Urtica dioica agglutinin: however there is only 36.9% identity between them. We propose that BjCHI1 should be classified under a new class, Chia7. The spacer and the hinge region of BjCHI1 are proline-rich, like that of Beta vulgaris Ch1, a Chia6 chitinase with half a chitin-binding domain. Northern blot analysis showed that the 1.3 kb BjCHI1 mRNA is induced by wounding and methyljasmonate (MeJA) treatment but is unaffected by ethylene, salicylic acid (SA) or abscisic acid (ABA). This is the first report on MeJA induction of chitinase gene expression and further suggests that wound-related JA-mediated signal transduction is independent of that involving SA. Western blot analysis using polyclonal antibodies against BjCHI1 showed a cross-reacting band with an apparent molecular mass of 37 kDa in wounded tissues of B. juncea, revealing that, unlike UDA1, BjCHI1 is not cleaved post-translationally at the hinge. Expression of recombinant BjCHI1 in Escherichia coli BL21(DE3) inhibited its growth while crude extracts from E. coli JM109 expressing recombinant BjCHI1 showed chitinase activity. Results from polymerase chain reaction (PCR) suggest that genes encoding chitinases with single or double chitin-binding domains exist in B. juncea. PMID- 10527426 TI - A starch-branching enzyme gene in wheat produces alternatively spliced transcripts. AB - A wheat gene, denoted Sbe1, encoding a type I starch-branching enzyme (SBEI) was isolated from a genomic library and shown to comprise 14 exons distributed over a 5.7 kb DNA region. Analyses of kernel RNA by 5' rapid amplification of cDNA ends (5'-RACE) and reverse transcription-polymerase chain reaction (RT-PCR) demonstrated a considerable sequence variation at the 5' ends of SBEI gene transcripts. DNA sequence alignments between the 5'-RACE products and the Sbe1 genomic DNA indicated that the first two exons and first intron were differentially processed to generate three classes of the mature transcript. One form of the SBEI gene transcript in 12-day old kernels contained the exon I+II+III combination at the 5' end, whereas other forms differed by inclusion of intron 1 or exclusion of exon II sequences. RT-PCR analysis of Sbe1-uidA::nptII chimeric mRNA produced in transgenic wheat cultured cells confirmed that the isolated Sbe1 was able to produce all three forms of SBEI gene transcripts by alternative splicing of the primary mRNA. The variants of processed Sbe1 mRNA were potentially translated into N-terminal variants of the SBEI precursor with different transit peptide sequences. PMID- 10527427 TI - Semi-quantitative RT-PCR analysis of photoregulated gene expression in marine diatoms. AB - The low cell densities of diatoms and other phytoplankton in culture has precluded the use of classical RNA analysis techniques for routine studies of gene expression in large numbers of samples. This has seriously hampered studies of the basic biology of such organisms. To circumvent this problem, we have developed a high-throughput semi-quantitative RT-PCR-based protocol and used it to monitor expression of a gene encoding a fucoxanthin, chlorophyll a/c-binding protein (FCP) in the centric planktonic diatom Thalassiosira weissflogii. Analysis of FCP gene expression in dark-adapted diatom cultures revealed that mRNA levels increase 5- to 6-fold in response to white light irradiation and peak around 6 to 8 h. To determine the photoreceptors involved in this response action spectra of FCP gene expression were determined using the Okazaki large spectrograph. Responses consistent with the presence of cryptochrome-, rhodopsin- and phytochrome-type receptors could be detected. The apparent presence of phytochrome-mediated responses is of particular interest given the low fluences of red and far-red light wavelengths in the marine environment. PMID- 10527428 TI - Importance of the B2 domain of the Arabidopsis ABI3 protein for Em and 2S albumin gene regulation. AB - Genetic and molecular studies have shown that the Arabidopsis ABSCISIC ACID INSENSITIVE3 (ABI3) protein plays a prominent role in the control of seed maturation. The ABI3 protein and its orthologues from various other plant species share four domains of high sequence identity, including three basic domains designated as B1, B2 and B3. The leaky abi3-1 mutation is a single amino acid substitution within the B3 domain. A new abi3 allele, abi3-7, was generated by mutagenizing abi3-1 seeds. The abi3-7 line contains, in addition to the abi3-1 mutation, a point mutation that converts residue Ala-458 into Thr within the B2 domain of the ABI3 protein. This Ala residue is absolutely conserved in all known ABI3 orthologues. Abi3-7 seeds display reductions in dormancy and in sensitivity to abscisic acid which are intermediate between those of the leaky abi3-1 and of the severe abi3-4 and abi3-5 mutants. Accumulation and distribution of At2S1 and At2S2 albumin mRNA as well as of AtEm1 and AtEm6 late embryogenesis-abundant proteins and mRNA have been analyzed. Both At2S1 and At2S2 mRNA are reduced in abi3-7, but distribution of At2S2 is spatially restricted. Accumulation of AtEm6 protein is more sensitive to abi3-7 mutation than AtEm1. However both mRNAs are considerably reduced in this mutant. Their distribution is also differentially affected. These results provide genetic evidence for the importance of the conserved B2 domain for ABI3 function in vivo. PMID- 10527429 TI - As Chlamydomonas reinhardtii acclimates to low-CO2 conditions there is an increase in cyclophilin expression. AB - When exposed to low CO2 levels, Chlamydomonas reinhardtii acquires the ability to accumulate CO2 to increase photosynthetic carbon fixation. A cDNA library has been constructed and screened to facilitate the identification of the different genes and proteins involved in this acclimation to low-CO2 conditions. The differential cDNA library screening led to the identification of several cDNAs up regulated under low-CO2 conditions. One such cDNA shows homology to cyclophilins, a class of immunophilins with a peptidyl-prolyl cis-trans isomerase activity. This is the first report of an algal cyclophilin. In this report we study the changes in the C. reinhardtii cyclophilin transcript and protein levels during low-CO2 adaptation. Possible reasons for the increased cyclophilin expression in response to the drop in CO2 concentration are discussed. PMID- 10527430 TI - Cloning and characterization of a class II DNA photolyase from Chlamydomonas. AB - Damage to DNA induced by ultraviolet light can be reversed by a blue light dependent reaction catalyzed by enzymes called DNA photolyases. Chlamydomonas has been shown to have DNA photolyase activity in both the nucleus and the chloroplast. Here we report the cloning and sequencing of a gene, PHR2, from Chlamydomonas encoding a class II DNA photolyase. The PHR2 protein, when expressed in Escherichia coli, is able to complement a DNA photolyase deficiency. The previously described Chlamydomonas mutant, phr1, which is deficient in nuclear but not chloroplast photolyase activity was shown by RFLP analysis not to be linked to the PHR2 gene. Unlike the recently reported class II DNA photolyase from Arabidopsis, the protein encoded by PHR2 is predicted to contain a chloroplast targeting sequence. This result, together with the RFLP data, suggests that PHR2 encodes the chloroplast targeted DNA photolyase. PMID- 10527432 TI - The declining interest in surgical careers, the primary care mirage, and concerns about contemporary undergraduate surgical education. PMID- 10527431 TI - The HD-Zip gene ATHB6 in Arabidopsis is expressed in developing leaves, roots and carpels and up-regulated by water deficit conditions. AB - Homeodomain-leucine zipper (HD-Zip) proteins are transcription factors as yet found only in plants. We have characterized one HD-Zip gene, ATHB6, from Arabidopsis thaliana. ATHB6 was expressed constitutively in seedlings, but significantly up-regulated in seedlings subjected to water deficit, osmotic stress or exogenous treatment with abscisic acid (ABA), an induction being detectable within 30 min. The ATHB6 induction was impaired in the two ABA insensitive mutants, abi1 and abi2, but unaffected in the abi3 mutation. The induction was ABA-dependent, since no increase in ATHB6 transcript was detected in the ABA-deficient mutant aba-3 subjected to drought treatment. These results suggest that ATHB6 may act downstream to both ABI1 and ABI2 in a signal transduction pathway mediating a drought stress response. A translational fusion of the ATHB6 promoter with the reporter gene GUS (ATHB6::GUS) in transgenic A. thaliana plants showed high-level expression in leaf primordia. Expression in developing cotyledons, leaves, roots and carpels was restricted to regions of cell division and/or differentiation. The expression in the cotyledons was detectable in the epidermis and high in the stomatal cells. In mature cotyledons and leaves the marker gene was expressed only in the vascular tissue. These expression data suggest ATHB6 to have a function related to cell division and/or differentiation in developing organs. PMID- 10527433 TI - Surgical internet at a glance: the American Online Health Channel. PMID- 10527434 TI - Surgical decision making for carotid endarterectomy and contemporary magnetic resonance angiography. AB - BACKGROUND: Benefit from carotid endarterectomy (CEA) centers on patient selection and percent stenosis as determined by cerebral angiography. However, angiography remains expensive and poses risks. Validated carotid duplex ultrasonography has proven to be an accurate tool for selecting patients for CEA. However, the role of another noninvasive test-magnetic resonance angiography (MRA)-remains uncertain. Because of recent advances in MRA hardware and software, we hypothesized that clinically appropriate patients could be accurately selected for CEA based on MRA alone. METHODS: Fifty-four carotid arteries in 29 patients (with and without symptoms) underwent both three-dimensional time-of-flight MRA (1.5 Tesla) with multiple overlapping thin slab acquisition and biplanar intra arterial digital subtraction angiography. All patients undergoing both tests over a 24-month period were included. The majority of these patients did not undergo carotid duplex ultrasound owing to the clinical practice of the hospital's neurosurgery service. Staff radiologists interpreted each study. The accuracy of patient selection based on MRA was calculated using angiography as the standard (NASCET method). Since operative thresholds vary depending on clinical history, we considered four commonly used ranges of percent stenosis for CEA. RESULTS: Patient selection accuracy of MRA alone was low, but increased as percent stenosis increased. Out of 10 occluded arteries by angiography, 5 were interpreted as patent with stenosis (70% to 99%) by MRA. One patent artery was misread as occluded on MRA. CONCLUSION: Reliance solely on contemporary MRA for surgical decision making cannot be justified in view of low accuracy, which leads to high rates of error in patient selection for CEA. PMID- 10527435 TI - Perigraft air, fever, and leukocytosis after endovascular repair of abdominal aortic aneurysms. AB - BACKGROUND: The postimplantation syndrome of fever and leukocytosis after endovascular repair of infrarenal aortic aneurysms has not been previously characterized and its etiology is not known. METHODS: We studied the first 12 patients who underwent successful endovascular repair of infrarenal aortic aneurysms with Dacron-covered stent-grafts, as part of an ongoing phase II clinical trial. Sepsis syndrome evaluations (physical examination, urinalysis, chest radiograph, urine cultures, and blood cultures) were performed for all patients with postoperative temperature (T) greater than 101.4 degrees F. Computed tomography scans of the abdomen were performed, as part of the clinical protocol, on postoperative days 2 and 30. RESULTS: Fever (T > 101.4 degrees F) was seen in 8 of 12 (67%) patients (P < 05). An additional 2 of 12 (17%) patients had low-grade fevers (100.3 degrees F, 100.6 degrees F). Only 2 of 12 (17%) patients remained afebrile postoperatively. Leukocytosis with counts over 11,000 white blood cells (WBC)/dL was observed in 7 of 12 (58%) patients (P < 05). Sepsis evaluations failed to identify any source of infection in 11 of 12 (97%) patients. Computed tomography scan evidence of perigraft air was noted in 8 of 12 (67%) patients. All patients were afebrile, had normal white blood cell counts, and were discharged within 1 week postoperatively. There has been no evidence of graft infection after 1 to 6 months of follow-up. CONCLUSIONS: Fever and leukocytosis after stent-graft repair of aortic aneurysms does not represent evidence of systemic or graft infection and is not clearly related to nonspecific causes of postoperative fever and leukocytosis. Moreover, the finding of early postoperative perigraft air is not necessarily an indication of graft infection even when concurrently present with fever and leukocytosis. PMID- 10527436 TI - Diagnosis, treatment, and outcome of blunt carotid arterial injuries. AB - BACKGROUND: Blunt carotid injuries are rare, and present late with devastating strokes. A sizeable single-institution descriptive report could help characterize the injury and its diagnosis and treatment. METHOD: We performed a retrospective review of blunt carotid artery injuries from May 1988 to December 1997 at a level I trauma center. Chart review consisted of demographics, mechanism of injury, associated injuries, diagnostic modalities, initial neurologic status, treatment, and outcome. Discharge outcome was classified as "good" (normal-mild deficit), "fair" (needing daily assistance), "poor" (institutionalized), or "dead." RESULTS: During the study period 16 patients sustained a carotid artery injury. Mean age was 35 years and 63% were female. Vehicular trauma was the most common mechanism of injury (81%), followed by assaults (13%). Dissection was the most common injury (75%), with one quarter having an associated pseudoaneurysm. Initial neurologic presentation was normal in 31% and Glasgow Coma Score was < 13 in 31% (including 13% in coma). Eventual hemispheric symptoms developed in 81%. Associated injuries were present in 94%, commonly head (44%) and chest (50%). Duplex ultrasound accurately identified the injury in all patients (5 of 5) when used. Anticoagulation (88%) had no complications. Observation and therapeutic embolization each resulted in 1 fatal stroke. A third patient, with worsening deficits on heparin, died after carotid ligation, for an overall mortality of 19%. There were no deaths in the 13 patients treated by anticoagulation alone. Six patients (38%) had a "good" neurologic outcome, five (31%) "fair," and two (13%) "poor." Initial neurologic presentation, associated injuries, and mechanism of injury did not appear to correlate with these outcome categories. CONCLUSIONS: These uncommon injuries should be suspected in the presence of head and/or chest injuries, basilar skull fracture, or coma (particularly if the computed tomography scan is unremarkable). Presentation may be varied, but most patients eventually develop hemispheric symptoms. Duplex ultrasound detects many of these injuries, but this does not demonstrate its utility as a screening tool. Anticoagulant therapy appears to be associated with a better outcome than expectant or occlusive therapy. PMID- 10527437 TI - The prevalence of carotid artery stenosis in patients undergoing aortic reconstruction. AB - BACKGROUND: Coronary artery disease occurs frequently in patients undergoing aortic reconstruction, and it has been presumed that internal carotid artery occlusive disease is also common. This has led to the practice of screening for and repairing significant carotid lesions in asymptomatic patients prior to aortic reconstruction. The purpose of this study was to determine the true prevalence of internal carotid artery disease in these patients. METHODS: The records of 240 patients who underwent duplex ultrasound screening for carotid artery disease prior to aortic reconstruction were reviewed. Surgery was performed for aortic aneurysm (AA) or aorto-iliac occlusive disease (AO). The prevalence of hyperlipidemia and coronary artery disease was similar between the two groups, but tobacco use, hypertension, and diabetes mellitus differed. RESULTS: Internal carotid artery stenosis > or = 50% occurred in 26.7% of the total group (64 of 240 cases). Stenosis > or = 50% was more common in the AO group (40 of 101 cases, 39.6%) than the AA group (24 of 139 cases, 17.3%, P = 0.0001). Severe disease (70% to 99%) was also more common in the AO group than the AA group (9.9% versus 3.6%, P = 0.0464). CONCLUSION: Internal carotid artery disease occurs commonly in patients undergoing aortic reconstruction, and screening is worthwhile. Significant disease is more common in patients with aorto-iliac occlusive disease than in those with aortic aneurysm, although atherosclerotic risk factors occur with varying frequency in the two groups. These findings suggest that additional factors may contribute to the higher prevalence of internal carotid artery stenosis in aorto-iliac occlusive disease. PMID- 10527438 TI - Long-term follow-up of reoperative carotid surgery. AB - BACKGROUND: We examined our long-term results of carotid reoperation to identify risk factors for morbidity and secondary recurrence. METHODS: Medical record review revealed 27 patients had reoperative surgery for recurrent stenosis. Demographics, operative details, pathology, clinical outcome, and follow-up imaging results were reviewed. RESULTS: No neurologic deficits and no mortalities were noted perioperatively. Long-term follow-up (average 54 months) revealed an 85% 5-year and 29% 10-year estimated survival. The 5- and 10-year estimated neurologic event rates were 15% and 35%, respectively. These included 3 ipsilateral strokes and 1 ipsilateral TIA; only the TIA involved secondary restenosis. Follow-up imaging revealed a 21% incidence of secondary restenosis, occurring more frequently in patients with hyperlipidemia (P < 0.05) and previous contralateral endarterectomy (P < 0.05). CONCLUSIONS: (1) Reoperation provides long-term protection from stroke due to recurrent stenosis. (2) Secondary restenosis rates appear higher than those for primary surgery. (3) Hyperlipidemia and contralateral endarterectomy are risk factors for secondary restenosis. PMID- 10527439 TI - The impact of an occluded internal carotid artery on the mortality and morbidity of patients undergoing coronary artery bypass grafting. AB - PURPOSE: To evaluate whether the presence of stenosis or an occluded internal carotid artery (ICA) influences perioperative stroke and mortality rates in patients subjected to coronary artery bypass grafting (CABG). MATERIAL AND METHODS: Between January 1995 and July 1998, 3,344 patients (59% males; 41% females) had CABG performed at our institution. Preoperative carotid duplex scans performed by registered vascular technologists at an ICAVL accredited laboratory were available for review in all patients. Of these, 3,101 (92.7%) had < 60% ICA stenosis (group A), 182 (5.4%) had 60% to 99% ICA stenosis (group B), and the remaining 61 (1.8%) had a occluded ICA (group C). In the latter group, 53 patients (87%) had < 60% contralateral ICA stenosis, while 8 (13.1%) had significant (60% to 99%) contralateral stenoses. Concomitant carotid endarterectomies (CEAs) were performed in 70 patients in group B (40%) and in 2 patients in group C (3.2%). Age, indications for surgery, prevalence of diabetes, hypertension, and smoking were similar in all groups. The mean pump time for groups A, B and C were 132, 138, and 125 minutes, respectively. The aortic cross clamp time for group A, B, C were 78, 75, and 75 minutes, respectively. Statistical analyses were performed using the chi-square, Fisher's exact test, and unpaired t test. RESULTS: Perioperative stroke rates (30 days) were 1.6%, 3.8%, and 6.5% for groups A, B, and C, respectively. Group A results varied significantly from groups B (P < 0.03) and C (P < 0.003). No statistically significant difference was noted between groups B and C (P = 0.6). The presence of a contralateral ICA stenosis in group C patients was predictive of a perioperative stroke (25% versus 3.8%; P < 0.0001). Concomitant CEAs for contralateral severe ICA stenosis in group C were associated with higher stroke rate (100%) when compared with those in group B patients (4.2%; P < 0.02). Perioperative (30 days) mortality rates for groups A, B, and C were 3.6%, 6.6%, and 8.6%, respectively. The mortality rate for group A was lower than for groups B (P < 0.05) and C (P < 0.05). CONCLUSION: The presence of an ICA occlusion increases the morbidity and mortality in patients undergoing CABG. To the best of our knowledge, this is the first reported large series of patients that investigates the role of carotid occlusions. PMID- 10527440 TI - Mobile atheroma of the aortic arch and the risk of carotid artery disease. AB - BACKGROUND: Mobile atheromas of the aortic arch are associated with otherwise unexplained strokes and transient ischemic attacks (TIA). They are associated with increased perioperative strokes in patients undergoing coronary artery bypass surgery. Peripheral embolization is an additional risk. Transesophageal echocardiography (TEE) accurately identifies mobile atheroma. Anticoagulant therapy may have therapeutic considerations in the management of this condition. However, the risk of significant carotid artery disease associated with mobile atheromas is unknown. METHODS: Between March 1994 and July 1998, 40 patients with mobile atheromas by TEE and evidence of embolization were studied. All patients were captured prospectively in a vascular registry and were retrospectively reviewed. Carotid artery disease was evaluated using carotid duplex imaging in an accredited vascular laboratory. All patients with significant carotid disease, 70% or greater stenosis, underwent arteriography. Patients with significant carotid artery stenosis then underwent carotid endarterectomy. All patients with mobile atheromas were maintained on anticoagulation. RESULTS: Forty patients with mobile atheromas of the aortic arch were diagnosed with TEE. All 40 patients had evidence of embolization. Patient age ranged from 57 to 73 years (mean 68.4). There were 22 men and 18 women. Twenty of 40 (50%) patients presented with symptoms of TIA. Eleven of 40 (28%) patients presented with diffuse atheroembolization (lower extremity embolization and renal insufficiency). Six of 40 (15%) patients presented with a completed stroke. Three of 20 (7%) patients presented with acute extremity ischemia secondary to a peripheral embolus. Twenty three of 40 (58%) of patients had significant carotid artery stenosis, 70% or greater stenosis. These 23 patients underwent both arteriography and carotid endarterectomy without complication. All patients were treated with anticoagulation and have remained anticoagulated. Clinical follow-up between 2 to 48 months (mean 18) has demonstrated no further evidence of systemic embolization in these 40 patients. Repeat TEE was performed in 6 of 40 patients. These follow up studies no longer visualized mobile atheromas. CONCLUSIONS: Mobile atheromas are recognized sources for embolization. Routine carotid duplex imaging should be performed in patients found to have mobile atheromas of the aortic arch. Carotid endarterectomy appears to be safe in patients who have combined carotid artery stenosis and mobile atheromas. Anticoagulation may have therapeutic considerations in the management of this condition. PMID- 10527441 TI - Complications associated with percutaneous closure devices. AB - BACKGROUND: In an effort to reduce time to hemostasis after angiography, several closure devices have been marketed. We report some of their complications. METHODS: A retrospective review was conducted. RESULTS: Over an 8-month study period, 2,181 diagnostic and interventional procedures were performed. Closure devices were used in 408 (19%) of these patients. The Angio-Seal closure device is composed of a collagen sponge and an absorbable polymer anchor that compresses the hole in the arteriotomy. The ProstarXL sealing device consists of a rotating barrel that deploys 4 needles through the arteriotomy, and then individual knots are extracorporeally tied. The Duett device consists of a balloon occluding catheter and injectable collagen and thrombin. Ten of the patients developed a complication from the closure device (2.5%). Four of these were subcutaneous abscesses. Two of these patients had expanding pseudaneurysms, 2 had lower extremity ischemia, and 1 patient had an acute bleed. These complications were all managed surgically. Another patient developed a retroperitoneal bleed that was managed nonoperatively. CONCLUSIONS: As the use of these devices increase, these complications will become more common. PMID- 10527442 TI - Special iliac artery considerations during aneurysm endografting. AB - BACKGROUND: The feasibility of endograft exclusion of abdominal aortic aneurysms (AAA) has been established. However, the technical challenges of graft delivery through tortuous or diseased iliac arteries and the treatment of associated iliac aneurysmal disease have received little attention. METHODS: Over 19 months, 74 patients underwent endoluminal repair of AAA and/or iliac artery aneurysms. Iliac anatomy that required special consideration during endografting was reviewed. RESULTS: Of the 74 patients, 35 (47%) had iliac anatomy that required special attention. Thirteen patients (18%) had aneurysmal involvement of a common iliac artery. Eleven of these patients required endograft extension into the external iliac artery (EIA) and hypogastric coil embolization due to the proximity of the aneurysm to the hypogastric origin. Eleven patients with ectatic, nonaneurysmal iliac arteries required aortic cuffs to achieve a distal seal in these oversized vessels. Iliac artery tortuosity or stenosis were complicating factors in 27 of the 74 patients (36%), requiring the use of brachial guidewire tension in 2 patients to facilitate tracking of the delivery device. Five patients with severely splayed aortic bifurcations required crossed placement of the iliac limbs to prevent kinking of the endograft. Occlusive atherosclerotic disease of the EIA mandated preprocedural dilatation and stenting in 3 patients and postprocedural surgical EIA reconstruction in another 5 patients. Three patients who underwent successful endograft placement required subsequent endovascular repair of traumatized EIAs. CONCLUSIONS: Iliac artery anatomy plays a significant role in the endoluminal treatment of infrarenal abdominal aortic aneurysms, complicating the procedure in up to 47% of patients with otherwise suitable anatomy. A variety of supplemental procedures, both surgical and endovascular, may be required to facilitate endograft placement. A special understanding of these constraints and proper planning is required for optimal therapy. PMID- 10527443 TI - Infectibility of endovascular stents following antibiotic prophylaxis or after arterial wall incorporation. AB - BACKGROUND: Case reports of endovascular stent infection have been accumulating in the last several years. We sought to determine if prophylactic antibiotics would prevent stent/artery complex infections in a swine model if given before a bacterial challenge at the time of stent placement and 4 weeks following deployment. We also investigated whether arterial wall incorporation protected the stent against infection without antibiotic prophylaxis. METHODS: Balloon expandable Palmaz stents were placed in the iliac arteries of 42 swine. At the same time, angioplasty was performed on the contralateral iliac artery as a control. In group A, prophylactic cefazolin was given to 12 swine at the time of stent deployment followed by an intraaortic bacterial challenge of Staphylococcus aureus. In group B, 10 swine received prophylactic cefazolin followed by intravenous S aureus 4 weeks after iliac stenting and angioplasty. In group C, 3 months following iliac stent placement and angioplasty an intravenous bacterial challenge was administered with S aureus. All swine were euthanized, and the iliac stent/artery complex and the contralateral angioplastied iliac artery were harvested and sent for culture and pathology. Experimental groups were compared with results from our previously published swine infection model using the Fisher's exact test. P values were considered significant if less than 0.05. RESULTS: Group A: Two of the 12 (17%) stent/artery complexes in the antibiotic treatment group had positive cultures. This compares with 7 of 10 (70%) in the control group (P = 0.016). In addition, there was one infection in an angioplastied vessel contralateral to one of the two stent infections. Molecular strain typing verified that the positive cultures were the same strain that was used to challenge the animals. No vessel thrombosis occurred in the stented arteries even in the presence of infection. Group B: One of 10 (10%) stented iliac arteries had a culture positive infection. This compares with 7 of 14 (50%) positive cultures in the control group (P = 0.04). In addition, one angioplastied vessel did have mild S aureus growth on culture. Both positive cultures were verified to be the same as the injected strain by molecular strain typing. There were no thrombosed or occluded vessels. Group C: One of 15 patent stents had growth of S aureus on culture and evidence of acute inflammation on histopathologic examination. The stent infection rate of 1 of 15 (7%) patent stents in this study was significantly less than the infection rates with bacterial challenge at placement (7 of 10, 70%; P < 0.01) and at 1 month postplacement (7 of 14, 50%; P = 0.0142). Five stents occluded without evidence of infectious cause. CONCLUSIONS: The results of this study support a recommendation that antibiotic prophylaxis should be used at the time of arterial stent placement and early after placement at times of anticipated bacteremia, but indefinite prophylaxis may be unnecessary due to arterial wall incorporation of the stent. PMID- 10527445 TI - Minimal access surgery for cholelithiasis induces an attenuated acute phase response. AB - BACKGROUND: Some benefits of laparoscopic (LC) and minilaparotomy (MC) cholecystectomy may reflect attenuation of the acute phase response. The authors examined components of this response. METHODS: Patients were randomized to LC (n = 11) or MC (n = 11). C-reactive protein (CRP), alpha-1-antitrypsin (AAT), retinol-binding protein (RBP), transferrin, and albumin were measured preoperatively and on postoperative days 1, 2, 4, and 7. Interleukin-1 receptor antagonist (IL-1ra), IL-6, and tumor necrosis factor (TNF-alpha) were measured more frequently perioperatively. Peak expiratory flow rate, forced expiratory volume in 1 second, and forced vital capacity were measured daily. RESULTS: The IL-6 increase was more persistent and marked in the MC patients from hour 8 to day 7 postoperatively (P < 0.05). Alterations in CRP, AAT, and albumin were similar. Postoperative deficits of respiratory function correlated with the magnitude of acute phase protein alteration. CONCLUSIONS: Minimal access surgery induces an acute phase response that is less prominent after a laparoscopic technique. PMID- 10527444 TI - Algorithm for the diagnosis and treatment of endoleaks. AB - BACKGROUND: Endoluminal grafting of abdominal aortic aneurysms (AAA) has shown promising early results. However, endoleaks present a new and challenging obstacle to successful aneurysm exclusion. We report our experience with primary, persistent endoleaks and provide an algorithm for their diagnosis and management. METHODS: Over a 19-month period, 73 patients underwent endoluminal repair of their AAAs using a modular bifurcated endograft as part of a US FDA Investigational Device Exemption trial. Spiral computed tomography (CT) scanning was performed prior to discharge after repair to evaluate for complete aneurysm exclusion. If no endoleak was present on that initial CT scan, color-flow duplex scanning was performed at 1 month, with repeat CT scanning at 6 months and 1 year. If the initial CT scan revealed the presence of an endoleak, repeat CT scanning was performed at 2 weeks, 1 month, and 3 months, or until the endoleak resolved. Any patient with an endoleak that persisted beyond 3 months underwent angiographic evaluation to localize the source of the leak. RESULTS: At 1 month, 62 patients (85%) had successful aneurysm exclusion. The remaining 11 patients (15%) had primary endoleaks, 8 (11%) of which persisted beyond 3 months, prompting angiographic evaluation. In 2 patients the endoleak was related to a graft-graft or graft-arterial junction. One was from the endograft terminus in the common iliac artery and was successfully embolized, along with its outflow lumbar artery. The other required placement of an additional endograft component across a leaking graft-graft junction to successfully exclude the aneurysm. The remaining six endoleaks were due to collateral flow through the aneurysm sac. In 4 cases this was lumbar to lumbar flow fed by hypogastric artery collaterals to the inflow lumbar artery. In the remaining 2 patients the endoleak was found to be due to flow between a lumbar and inferior mesenteric artery. Resolution of the endoleak by coil embolization of the feeding hypogastric artery branch in 1 patient was unsuccessful due to rapid recruitment of another hypogastric branch. Two of the six collateral flow endoleaks have resolved spontaneously without treatment, while the remaining cases have been followed up without evidence of aneurysm expansion. CONCLUSION: Systematic postoperative surveillance facilitates proper diagnosis and treatment of endoleaks. This involves serial CT scans to detect the presence of endoleaks, followed by angiography to determine their etiology and guide treatment, if clinically indicated. PMID- 10527446 TI - Traumatic injury to the superior mesenteric artery. AB - BACKGROUND: Superior mesenteric artery (SMA) injuries are rare and devastating injuries incurring very high mortality rates. It is the purpose of this study to review our experience with these injuries, to analyze Fullen's classification based on anatomical zone and injury grade for its predictive value, and to correlate the American Association for the Surgery of Trauma-Organ Injury Scale (AAST-OIS) for abdominal vascular injury with mortality. METHODS: Retrospective study was made over a 65-month period of all patients sustaining SMA injuries in an urban level I trauma center. RESULTS: Thirty-five patients, mean age 31, had a mean Revised Trauma Score of 5.86 and a mean Injurity Severity Score of 23. Mechanisms of injury were penetrating 27 (77%) and blunt 8 (23%). Mean admission systolic blood pressure was 85 mm Hg. Mean estimated blood loss was 8,500 mL and mean total fluid replacement 17,000 mL. Operating room findings were retroperitoneal hematoma in 34 (97%) and "black bowel" in 2 (6%). Number of associated injuries was nonvascular, mean 4.2, and vascular, mean 1.5. Surgical management consisted of ligation in 18 (51%), primary repair in 14 (40%), and interposition graft in 2 (6%). Overall mortality was 19 of 35 (54%). Mortality versus Fullen's zones was zone I, 100%, zone II, 43%, and zones III and IV, 25%. Mortality versus Fullen's ischemia grade was grade 1, 89%, grade 2, 58%, grade 3, 100%, and grade 4, 19%. Mortality versus AAST-OIS: was grade 1, 0%, grade II, 20%, grade III, 0%, grade IV, 59%, and grade V, 88%. CONCLUSIONS: SMA injuries are highly lethal. Most deaths are due to exsanguination. A higher number of associated vascular injuries increases mortality. "Black bowel" is an uncommon finding. Both Fullen's anatomical zones and the AAST-OIS for abdominal vascular injuries correlate with mortality. Fullen's ischemia grade does not. PMID- 10527447 TI - Factors affecting the incidence of postoperative septic complications and recurrence after strictureplasty for jejunoileal Crohn's disease. AB - BACKGROUND: This retrospective study was undertaken to examine the long-term outcome of strictureplasty for Crohn's disease and factors affecting the incidence of postoperative septic complications and recurrence. METHODS: Eighty seven patients who underwent 245 primary jejunoileal strictureplasties for jejunoileal Crohn's disease between 1980 and 1997 were reviewed. RESULTS: Septic complications (fistula/abscess) occurred in 7 patients (all at strictureplasty site). Only intra-abdominal sepsis with peritoneal contamination at laparotomy was significantly associated with these complications. After a median follow-up of 104 months, 49 patients (56%) developed symptomatic recurrence. In 11 patients, symptomatic recurrence was successfully managed by medical treatment. Thirty-eight patients (44%) required further surgery for recurrence. Only young age (< or = 37 years) was associated with high incidence of reoperation for recurrence. Preoperative steroid use, nutritional status, synchronous bowel resection, and number, site, or length of strictureplasties did not affect the incidence of septic complications and recurrence requiring reoperation. CONCLUSIONS: Intra-abdominal sepsis with peritoneal contamination increased the incidence of septic complications. Only young age was associated with the increased risk of recurrence requiring reoperation. PMID- 10527448 TI - The presentation of gallstones and results of biliary surgery in a spinal cord injured population. AB - BACKGROUND: Since spinal cord injured patients lack visceral sensation, their clinical manifestations of gallstones could be relatively occult. A higher proportion of these individuals may present with advanced biliary disease compared with the general population. Prophylactic cholecystectomy for asymptomatic stones may therefore be justified. METHODS: All spinal cord injured patients seen at the Seattle Veterans Hospital over a 5-year period were retrospectively surveyed to define a set of patients who had undergone a cholecystectomy. The operative indications and results were compared with those from a series of cholecystectomies in neurologically intact patients. RESULTS: The presentation of biliary disease in spinal cord injured patients was not more advanced than that of neurologically intact patients. Patients with high cord injuries presented in a similar fashion to those with low injuries. CONCLUSIONS: Since most spinal cord injured patients with biliary disease present with typical findings, prophylactic removal of gallstones in these patients is not warranted. PMID- 10527449 TI - Long-term results after esophagectomy for squamous cell carcinoma of the esophagus associated with head and neck cancer. AB - BACKGROUND: Esophageal squamous cell carcinomas are frequently associated with head and neck cancers. The poor prognosis of each cancer, and their proximity, often limit the treatment options. This study was conducted to determine the characteristics and long-term outcome of such dual cancers. PATIENTS AND METHODS: We included 75 patients with esophageal carcinoma, of whom 25 had a synchronous head and neck malignancy. Curative treatment was possible in every case. The patients were divided into "solitary cancer" and "synchronous cancer" groups. RESULTS: The gender distribution, tumor location, and histological findings were similar in the two groups. Patients in the synchronous cancer group were younger than those in the solitary group (P < 0.0042). The operative mortality and pulmonary morbidity rates were not significantly different in the two groups. The rate of cervical anastomotic leaks was higher in the synchronous group (P < 0.05). The mean follow-up was 83 +/- 50 months. Five-year survival rates were not significantly different in the two groups (14.3% +/- 5.7% in the solitary group and 17.5% +/- 7.9% in the synchronous group). CONCLUSIONS: With aggressive treatment, the survival of patients with synchronous esophageal and head and neck cancers was similar to that of patients with isolated esophageal cancer. PMID- 10527450 TI - Intraoperative lavage for cytological examination in 1,297 patients with gastric carcinoma. AB - BACKGROUND: This study examined the clinical value of intraoperative peritoneal lavage for cytological examination in patients with gastric cancer. Peritoneal dissemination is the most frequent mode of recurrence for this tumor. METHODS: A retrospective of lavage findings, other factors, and outcome was performed in 1,297 patients with gastric cancer who underwent intraoperative peritoneal lavage. RESULTS: The 5-year survival rate of patients with positive lavage cytology was only 2%. Patients who underwent curative resection and had negative cytology had a significantly better 5-year survival rate (P < 0.001). Even among patients with macroscopic peritoneal dissemination, the survival rate was significantly better with negative cytology, which reflected fewer free cancer cells in the peritoneal cavity. Serum concentrations of carcinoembryonic antigen and carbohydrate antigen 19-9 were significantly higher in patients with positive cytology. Multivariate analyses indicated that intraoperative cytological findings was an independent prognostic factor for survival, and was the most important factor for predicting peritoneal recurrence. CONCLUSIONS: Intraoperative peritoneal lavage cytology is important in predicting survival and peritoneal recurrence in gastric cancer. PMID- 10527451 TI - Additional microvascular anastomosis in reconstruction after total esophagectomy for cervical esophageal carcinoma. AB - BACKGROUND: Maintaining sufficient blood flow to the substitute organ after total esophagectomy is essential for decreasing the risk of anastomotic leakage. Additional venous, or arterial and venous, anastomoses between the vessels of the gastric tube and the vessels in the neck after total esophagectomy are described for 11 patients with cervical esophageal carcinoma. METHODS: The tissue blood flow was measured by laser Doppler flowmetry before and after anastomosis. Venous anastomosis was performed for all 11 patients, and arterial anastomosis was added for 7 patients. RESULTS: A significant increase in tissue blood flow was observed after venous anastomosis alone (mean, 19%; P < 0.05) and after arterial and venous anastomoses (mean 43%; P < 0.01). There was no anastomotic leakage or hospital death. CONCLUSIONS: This procedure may reduce the risk of anastomotic leakage especially in the case of pharyngogastrostomy following total esophagectomy. PMID- 10527452 TI - Intrathecal synthesis of immunoglobulins in Neisseria meningitidis and echovirus 6 meningoencephalitis. AB - Infectious diseases of the central nervous system (CNS) constitute the main cause of death amidst the neurological diseases in pediatric patients in Cuba. This factor, among others, made us study the immune response in children with such disorders. Cerebrospinal fluid (CSF) and serum samples were collected from 69 patients: 26 with Neisseria meningitidis meningoencephalitis, and 43 with echovirus 6. IgA, IgM, IgG, and albumin were measured by immunodiffusion in serum and CSF. Local synthesis of immunoglobulins was calculated using Reiber's formula. The detection of intrathecal synthesis of IgG represents an important event in the majority of cases. Detection depends on the acuity of the process, the nature of the biologic agent and its virulence, and the immunological status of the patient. Local IgG synthesis is a very frequent finding in Neisseria meningoencephalitis and echovirus 6 meningoencephalitis. Intrathecally synthesized IgA in patients suffering from N. meningitidis meningoencephalitis is an important indication. PMID- 10527454 TI - Amyloid beta peptide 25-35 modulates hydrolysis of phosphoinositides by membrane phospholipase(s) C of adult brain cortex. AB - Phosphoinositide-specific phospholipase C (PLC) is a key enzyme in signal transduction. A subset of muscarinic cholinergic receptors are linked to G proteins that activate phospholipase C. Cholinergic pathways are important in learning and memory, and deficits in cholinergic transmission have been implicated in Alzheimer's disease (AD). AD is also associated with increased beta amyloid plaques. In the present study, we have investigated the effect of the amyloid beta (A beta) synthetic peptide homologous to residue 25-35 of A beta in nonaggregated and aggregated forms on the degradation of inositol phospholipids. Synaptic plasma membranes (SPM) and the cytosolic fraction from rat brain cortex served as a source of enzymes. The studies were carried out with radioactive inositol phospholipids in the presence of endogenous and 2 mM CaCl2. The enzyme(s) activity was evaluated by determination of the product formation of [3H]inositol-1-phosphate (IP1) or [3H]inositol-1,4,5-trisphosphate (IP3). Results show that the PI-PLC activity was significantly higher in cytosol compared to SPM, and this enzyme was stimulated by 2 mM CaCl2, but not by GTPgammaS or carbachol, a cholinergic receptor agonist. Activity of the SPM-bound PIP2-PLC was similar to that in cytosol and was not activated by 2 mM CaCl2. The SPM PIP2-PLC was significantly stimulated by GTPgammaS together with the cholinergic agonist, carbachol. Fresh-water-soluble A beta 25-35 activated PI-PLC in SPM markedly by two- to threefold, but this effect was absent in the presence of 2 mM CaCl2. Moreover, A beta 25-35 had no effect on basal PIP2-PLC activity and cytosolic PI PLC and PIP2-PLC. The aggregated form of A beta 25-35 significantly inhibited PIP2-PLC only in the presence of endogenous CaCl2. It also inhibited the carbachol and GTP(gamma)S-stimulated PIP2-PLC. Our findings show that depending on the aggregation state and Ca2+ concentration, A beta modulates phosphoinositide degradation differently and exclusively in brain synaptic plasma membranes. Our data suggested that aggregated A beta peptide may be responsible for the significant impairment of phosphoinositide signaling found in brain membranes during AD. PMID- 10527453 TI - Involvement of lipid mediators on cytokine signaling and induction of secretory phospholipase A2 in immortalized astrocytes (DITNC). AB - Our previous studies demonstrated the ability of proinflammatory cytokines, such as tumor necrosis factor-alpha (TNF-alpha) and interleukin 1beta (IL-1beta), to stimulate NFkappaB/DNA binding and synthesis of secretory phospholipase A2 (sPLA2) in immortalized astrocytes (DITNC). In this study, we examined possible involvement of lipid mediators in the cytokine action. Using [14C]serine to label sphingomyelin and ceramide in these cells, subsequent exposure of cells to cytokines did not result in alteration of sphingomyelin/ceramide ratio. Furthermore, neither exogenous sphingomyelinase nor cell-permeable ceramides could stimulate NFkappaB/DNA binding. On the other hand, C-2 ceramide (0.3 microM) as well as other lipid mediators, such as lysophosphatidylcholine and arachidonic acid, were able to elicit a small increase in sPLA2 and potentiate the induction of sPLA2 by TNF-alpha. When DITNC cells were prelabeled with [32P]Pi, an increase in labeled phosphatidic acid (PA) was observed on treatment of cells with IL-1beta (200 U/mL). However, despite the ability of phorbol myristate acetate (PMA) to stimulate phospholipase D (PLD) and synthesis of phosphatidylethanol (PEt) in these cells, PLD activity was not affected by IL 1beta. With the [32P]labeled cells, however, PA-phosphohydrolase inhibitors, such as chlorpromazine and propranolol, could elicit large increases in labeled PA, indicating active PA metabolism in these cells. Cytokines also caused an increase in levels of diacylglycerol (DG) in these cells, although the source of this lipid pool is presently not understood. Taken together, these results provide evidence for the participation of PA and DG in cytokine signaling activity. Furthermore, although cytokines did not cause the release of ceramide, lipid mediators, such as lysophospholipids, and AA could modulate cytokine-mediated induction of sPLA2 in astrocytes. PMID- 10527455 TI - Gangliosides and neutral glycolipids in ependymal, neuronal and primitive neuroectodermal tumors. AB - Neutral glycolipid and ganglioside compositions were determined on 11 ependymal tumors, 12 medulloblastomas, 6 other neuronal tumors of the brain, 4 peripheral neuroblastomas, 1 cerebral primitive neuroectodermal tumor (PNET), and 1 PNET of the thoracic wall. Within the group of tumors that can demonstrate neuronal phenotypes, there was an association between the degree of neuronal differentiation usually demonstrated by these tumors and the proportions of both GD1a and 1b-pathway gangliosides. The amount of globoside also correlated with the amount of 1b pathway gangliosides. Patients with medulloblastomas whose 1b gangliosides made up over 15% of the total gangliosides survived longer that those with lower proportions of 1b gangliosides. The only gangliosides in the choroid plexus papilloma were GM3 and GD1a, but other ependymal tumors had significant amounts of GD1b and its metabolic precursors. Ependymoma and anaplastic ependymoma had similar neutral glycolipid compositions, which were different from subependymoma, which lacked ceramide monohexoside and ceramide dihexoside. These differences in glycolipid compositions suggest that there may be fundamental biological differences between these types of ependymal tumors. PMID- 10527456 TI - Immunocytochemical localization of cPLA2 in rat and monkey spinal cord. AB - Rat spinal cord contains a high level of calcium-dependent cytosolic phospholipase A2 (PLA2) activity. A dense immunoreactivity is present in motor neurons from cervical, thoracic, lumbar, and sacral regions of rat spinal cord. Under normal conditions, this enzyme liberates arachidonic acid, a polyunsaturated fatty acid that is a second messenger itself, and a precursor for eicosanoids. However, under pathological conditions during spinal cord injury, intracellular calcium increases so the cytosolic PLA2 may also be involved in the release and accumulation of arachidonic acid, eicosanoids, and lipid peroxides. PMID- 10527457 TI - Identification of 3'UTR region implicated in tau mRNA stabilization in neuronal cells. AB - Tau, a neuronal microtubule-associated protein (MAP) plays an important role in the formation and maintenance of neuronal polarity. Tau mRNA is a stable message and exhibits a relatively long half-life in neuronal cells. The regulation of mRNA stability is a crucial determinant in controlling mRNA steady-state levels in neuronal cells and thereby influences gene expression. The half-lives of specific mRNAs may be dependent on specific sequences located at their 3'untranslated region (UTR), which in turn, may be recognized by tissue-specific proteins. To identify the sequence elements involved in tau mRNA stabilization, selected regions of the 3'UTR were subcloned downstream to c-fos reporter mRNA or to the coding region of the tau mRNA. Using stably transfected neuronal cells, we have demonstrated that a fragment of 240 bp (H fragment) located in the 3'UTR can stabilize c-fos and tau mRNAs. Analysis of stably transfected cells indicated that the transfected tau mRNAs are associated with the microtubules of neuronal cells, suggesting that this association may play a role in tau mRNA stabilization. This step may be a prerequisite in the multistep process leading to the subcellular localization of tau mRNA in neuronal cells. PMID- 10527459 TI - Human hematopoietic stem/progenitor cells generate CD5+ B lymphoid cells in NOD/SCID mice. AB - The nonobese diabetic/severe combined immunodeficient (NOD/SCID) xenotransplantation model is increasingly utilized to study both human lymphohematopoietic stem/progenitor cells and committed cell types. Human B lymphoid cells develop and proliferate in this model. We found high numbers of CD19+CD5+ B lymphoid cells in the bone marrows and spleens of NOD/SCID mice transplanted with human CD34+ stem/progenitor cells. The CD5+ cells accounted for a particularly large percentage of the B lymphoid cells in the spleens of chimeras analyzed three months after transplantation. CD19+CD5+ cells from all the analyzed chimeras coexpressed HLA-DR, surface IgM, CD20, CD38, CD43, and CD45. However, CD19+CD5+ cells were negative for kappa light chain, CD10, CD11a, CD11b, CD15, CD21, CD22, CD23, CD25, CD34, CD35, CD44, CD62L, CD69, and CD71. Cell surface expression of the lambda light chain, surface IgD, CD9, and CD40 antigens was detected in some but not all chimeras. Thus, the CD19+CD5+ cell population detected in our study has the phenotype of previously described CD5+ B lymphoid cells in humans and other species. The origin and role of the B lymphoid cells which express CD5 cell surface glycoprotein are poorly understood. The malignant cells in B lymphoid chronic lymphocytic leukemia express CD5, and the numbers of CD5+ B lymphoid cells are elevated in several autoimmune conditions. The human-NOD/SCID chimera system may provide an in vivo model to investigate the maturation and development of this cryptic human CD5+ B lymphoid cell subpopulation. PMID- 10527458 TI - Effect of HIV-1 gp41 peptides on secretion of beta-amyloid precursor protein in human astroglial cell line, T98G. AB - To understand the mechanism underlying cognitive deficits in AIDS patients, we examined the influence of gp41 peptides on the expression and the secretion of Alzheimer's amyloid precursor protein (APP) in human astroglial cell line T98G. Western blotting analyses demonstrated that treatment of glial cells with a putative immunosuppressive domain (aa 583-599) of gp41 remarkably downregulated the interleukin 1beta- (IL-1beta) induced elevation of the secreted form of APP (sAPP alpha) containing Kunitz-type protease inhibitor (KPI) domain without significant changes of the expression pattern of APP mRNAs as revealed by reverse transcriptase polymerase chain reaction (RT-PCR) analysis. Recombinant gp41 protein encoding for ectodomain, including aa 583-599 residues, also elicited a similar dose-dependent inhibitory effect, whereas the control peptides resulted in little change. The molecular mechanism underlying this gp41-mediated reduction of sAPP alpha secretion appears not to be owing to the difference in the function of extracellular proteases based on the finding of similar proteolytic activities responsible for APP metabolism in vitro present in the conditioned media from the cultures treated with or without gp41 peptide. However, the known PKC inhibitors such as H-7 or staurosporine, partially inhibited the elevation of sAPP alpha secretion in response to protein kinase C (PKC) agonist phorbol 12,13-dibutyrate (PdBu) as well as to IL-1beta, mimicking the immunosuppressive gp41 peptide. These observations implicate that part of the neurodegenerative cascade in AIDS brains may involve the inhibitory effect of gp41 on secretion of sAPP alpha, a potent glial neurotrophic factor, through impaired PKC response. PMID- 10527460 TI - Maturation rate of mouse neutrophilic granulocytes: acceleration by retardation of proliferation, but no detectable influence from G-CSF or stromal cells. AB - Our purpose was to examine the possible influence of stromal and humoral mediators on granulocytic maturation rates. Sorted immature murine progenitor (Lin-Sca-1+) cells were cultured in peritoneal diffusion chambers (DCs) with or without a confluent layer of irradiated bone marrow stromal cells on one of the micropore membrane walls. In other experiments, 10 microg/kg/d recombinant G-CSF (rhG-CSF) was administered continuously into DC host mice through s.c. implanted osmotic minipumps. Operationally, maturation rate was assessed as the ratio between the number of polymorphonuclear cells (PMN) and proliferative granulocytes (PG) in short-term cultures, based on the differential cell counts, and supported by flow cytometric measurement of a granulocytic differentiation marker; and by the emergence time of PMN in the DCs, obtained by extrapolation. Also, increased maturation is associated with increased cell density, as reflected by the positioning of the granulocytes during centrifugation in a discontinuous Percoll gradient. This method, as well as the conversion rate of 3H thymidine labeled PG into the heavier non-PG maturational stages, were also used as indicators of maturation rate. After five, six, and seven days of culture in the peritoneal cavity, DC cells were harvested. Their proliferative status, based on measurement of incorporated bromodeoxyuridine, was determined, and their maturation rates were evaluated. Proliferation of immature granulocytic progenitor cells was apparently inhibited by direct contact with bone marrow stromal cells, and stimulated by G-CSF during the early stage of culturing. However, the subsequent maturation rate, which could be accelerated by increasing culture cellularity, thus decreasing PG proliferation rate, was not detectably influenced by either stromal cells or G-CSF. PMID- 10527461 TI - Correlation between IL-3 receptor expression and growth potential of human CD34+ hematopoietic cells from different tissues. AB - CD123 (alpha-subunit of IL-3 receptor) expression on primitive and committed human hematopoietic cells was studied by multicolor sorting and single-cell culture. The sources of cells included fetal liver (FLV), fetal bone marrow, umbilical cord blood, adult bone marrow and mobilized peripheral blood. Three subsets of CD34+ cells were defined by the levels of surface CD123: CD123negative, CD123low, and CD123bright. Coexpression of lineage markers showed that a majority of CD34+CD123bright cells were myeloid and B-lymphoid progenitors, while erythroid progenitors were mainly in the CD34+CD123negative subset. The CD34+CD123low subset contained a heterogeneous distribution of early and committed progenitor cells. Single CD34+ cells from the CD123 subsets were cultured in a cytokine cocktail of stem cell factor, interleukin 3 (IL-3), IL-6, GM-CSF, erythropoietin, insulin-like growth factor-1, and basic fibroblast growth factor. After 14 days of incubation, a higher cloning efficiency (CE) was observed in the CD34+CD123negative and CD34+CD123low fractions (37+/-23% and 44+/ 23%, respectively) than in the CD34+CD123bright fraction (15+/-21%). Using previously published criteria that colonies containing dispersed, translucent cells (dispersed growth pattern, DGP) were derived from primitive cells and that colonies composed solely of clusters were from committed cells, early precursors were distributed evenly in the CD34+CD123negative and CD34+CD123low subsets. When CD38 and CD90 (Thy-1) were used for further characterization of CD34+ cells from FLV, CE increased from 37+/-23% in CD123negative to 70+/-19% in CD123negativeCD38 and from 44+/-23% in CD123low to 66+/-19% in CD123lowCD38-. No significant increase in CE or DGP progenitors was observed when CD34+ cells were sorted by CD90 and CD123. We concluded that: A) high levels of CD123 were expressed on B lymphoid and myeloid progenitors; B) early erythroid progenitors had little or no surface CD123, and C) primitive hematopoietic cells are characterized by CD123negative/low expression. PMID- 10527462 TI - The reduction of in vitro radiation-induced Fas-related apoptosis in CD34+ progenitor cells by SCF, FLT-3 ligand, TPO, and IL-3 in combination resulted in CD34+ cell proliferation and differentiation. AB - Recovery from radiation-induced (RI) bone marrow aplasia depends on appropriate cytokine support. The early effects of exogenous cytokines at the hematopoietic stem and progenitor cell (HSPC) level following irradiation are still largely unknown, especially those of survival factors such as stem cell factor (SCF) and Flt-3 ligand (FL). This study was aimed at A) clarifying Fas/Fas-Ligand (Fas-L) implication in RI apoptosis of CD34+ cells and B) assessing the capacity of a combination of cytokines to mitigate RI apoptosis in HSPCs in vitro. We showed that most of in vitro gamma-irradiated CD34+ HSPCs incubated in a medium devoid of cytokines underwent progressive apoptosis-related changes from 6 h (i.e., decreased CD34 antigen expression, Annexin V binding); then Fas/Fas-L coexpression occurred from 10 h on. A strong DNA fragmentation, as assessed by TUNEL assay and propidium iodide staining, was observed at 24 h. Within a 2.5- to 6-Gy dose range, the RI apoptotic process finally led to 97% CD34+ cell death within 48 h with a complete loss of functionality. Unirradiated cells incubated in the same conditions displayed a significantly reduced apoptotic pattern. The early addition of a combination of SCF, FL, thrombopoietin, and interleukin 3 (4F) after cell irradiation prevented 15% (2.5 Gy) and 12% (4 Gy) of HSPCs, respectively, from RI apoptosis, whereas these cytokines used as single factors were inefficient. Furthermore, irradiated HSPCs (2.5 Gy) incubated with 4F in a serum-free culture system for seven days proliferated, giving rise to an increase in the number of total cells (x5.6-fold) and CD34+ cells (x4.2-fold) and to megakaryocytic and granulomonocytic precursors. These results show that the prevention of apoptosis in in vitro irradiated HSPCs depends on an early combination cytokine support. These data suggest that the early therapeutic administration of anti-apoptotic cytokines may be critical for preserving functional HSPCs from in vivo radiation damage. PMID- 10527463 TI - Comparison of hematopoietic activities of human bone marrow and umbilical cord blood CD34 positive and negative cells. AB - Although the hematopoietic activities of human CD34+ bone marrow (BM) and cord blood (CB) cells have been well characterized, the phenotype of nonobese-diabetic severe combined immunodeficient (NOD/SCID) mice repopulating cells (SRCs) in CB and BM has not yet been fully examined. To address this issue, various hematopoietic activities were compared in terms of total and CD34+ CB and BM cells. Clonal culture of fluorescence-activated cell sorter (FACS) CD34+ CB and BM cells revealed a higher incidence of colony-forming cells with greater proliferation capacity in CB over BM CD34+ cells. CB CD34+ cells also demonstrated higher secondary plating efficiency over BM cells. In addition, we demonstrated that mice transplanted with CB mononuclear cells (MNCs) showed significantly higher levels of chimerism than those transplanted with BM MNCs. However, recipients of FACS-sorted CD34+ CB cells showed significantly lower levels of chimerism than those that received total CB MNCs, suggesting a role of facilitating cells in the CD34- cell population. To further analyze the role of CD34- cells, the NOD/SCID repopulating ability of FACS-sorted CB CD34-c-kit+Lin- and CD34-c-kit-Lin- cells were examined. However, SRCs were not detected in those cells. Taken together, these data suggest that CB is a better source of hematopoietic stem cells and that there are cells in the CD34- fraction that facilitate repopulation of hematopoiesis in the NOD/SCID environment. PMID- 10527464 TI - Maintenance of CD34 expression during proliferation of CD34+ cord blood cells on glycosaminoglycan surfaces. AB - Recent studies have indicated that glycosaminoglycan (GAG) interactions with hematopoietic progenitors play a significant role in the regulation of hematopoiesis. However, the details of these interactions are not clear. In this study, we examined the role of soluble and immobilized GAGs in the proliferation of CD34+ cells. Chitosan, a cationic polysaccharide, was used to immobilize GAGs in ionic complex membranes. The GAGs studied were heparin, hyaluronate, and chondroitin sulfates A, B, and C. CD34-enriched umbilical cord blood cells were seeded onto tissue culture plates coated with the GAG-chitosan complex membranes. Cultures were maintained in medium supplemented with stem cell factor and interleukin 3 for up to six weeks, during which total and CD34+ cell numbers were determined by flow cytometry. Total cell number expansion ranged from 25-fold to 40-fold after six weeks. However, only heparin and chondroitin sulfate B (CSB) surfaces retained a significant CD34+ fraction. All other surfaces exhibited declines in CD34 expression, with negligible CD34+ percentages remaining after four weeks. In contrast, heparin and CSB surfaces exhibited CD34+ fractions as high as 90% after four weeks. GAG desorption studies indicated that the observed effects were partly mediated by desorbed GAGs in a concentration dependent manner. Subsequent studies showed that sustained high (160 microg/ml) heparin levels had toxic effects, while the same concentration of CSB exhibited more rapid early proliferation of CD34+ cells. In conclusion, this culture system has demonstrated the ability to produce simultaneous proliferation and CD34+ cell enrichment of a partially purified cord blood population by controlling the nature and levels of GAG moieties to which the cells are exposed. The results indicate that specific GAGs can significantly influence the growth and differentiation characteristics of cultured CD34+ cells. PMID- 10527465 TI - Apoptosis. AB - Mechanisms in Hematology is a book with an accompanying interactive CD-ROM designed to assemble basic concepts that underlie clinical understanding and progress. It is presented as a concise text with a series of diagrams that distill diffuse information into a compact form. The interactive CD, in particular, brings many of the processes "to life" as details of the more complex pathways are conveyed in clear visual images. The text begins with the basic molecular biology that underlies hematological and oncological physiology/pathology-cell signaling, adhesion molecules, and apoptosis. This is followed by sections, among others, on hematopoiesis, iron, B12, and folate metabolism, neutrophil function, immunoproteins, chemotherapy, and coagulation. With the permission of the authors and publisher, Stem Cells has reproduced the section on apoptosis, which we think our readers will enjoy. PMID- 10527466 TI - The molecular perspective: methotrexate. PMID- 10527467 TI - Maternal vitamin A nutriture and the vitamin A content of human milk. AB - Because of the many functions of vitamin A in human physiology, deficiency or excess of the vitamin in lactating women or their infants can adversely affect their health. Infants are born with low body stores of vitamin A, and rely on vitamin A in milk to meet their needs. The vitamin A content of milk is related to maternal vitamin A status and maternal dietary vitamin A intake during lactation. Low-income lactating women in non-industrialized countries have lower milk vitamin A concentrations than lactating women in industrialized countries. Supplementation of lactating women in non-industrialized countries with vitamin A or beta-carotene has resulted in increased milk vitamin A concentrations. However, the optimal timing and dose for sustaining adequate levels of vitamin A in milk throughout the lactation period has not been determined. Further research is needed to understand factors affecting the transfer of vitamin A to milk, and to evaluate various strategies for improving the vitamin A status of mothers and infants. PMID- 10527468 TI - Zinc transfer to the breastfed infant. AB - Zinc is a micronutrient which is critical to normal growth and development. Zinc concentrations in human milk decline sharply during the early months post partum, regardless of maternal zinc intake. Milk zinc concentrations do not increase in response to increased maternal zinc intake if maternal zinc status is adequate. The mechanism of zinc secretion into milk is not fully understood. A mutation in the gene for a zinc transporter protein may account for abnormally low milk zinc concentrations associated with severe zinc deficiency in breastfed infants. The zinc requirements of breastfed infants are generally met with exclusive breastfeeding through 5-6 months of age, due to the favorable bioavailability of the zinc in human milk. Because of declining milk zinc concentrations and intake, zinc status in exclusively breastfed infants is likely to become marginal beyond 6 months of age, and may become suboptimal for some infants if exclusive breastfeeding continues. The choice of complementary foods is important to maintain adequate zinc status in breastfed infants after 6 months. PMID- 10527470 TI - Interrelation among dietary energy and fat intakes, maternal body fatness, and milk total lipid in humans. AB - The relationship between maternal dietary intakes of energy or fat maternal body composition and the milk fat concentration is an important element in understanding the role of breast-milk in infant nutrition. In most studies in both developing and developed countries, no relation between maternal energy intake and milk fat content was observed. In only one published study, in which maternal fat intake comprised a very low 5% of calories, was a short term reduction in milk lipid observed in some subjects. On the other hand, a positive relation between maternal fatness and milk fat is evident in both well-nourished and under-nourished women when appropriate methodologies have been used. Low milk fat concentrations are associated with higher milk volumes probably because infant demand determines milk intake, compensating, at least partially, for low milk fat. No impairment of infant growth was associated with low milk fat, in studies where it has been measured. The mechanism for the relationship between body fat and milk fat is a fertile field for additional investigation. PMID- 10527469 TI - Polyunsaturated fatty acids in human milk and their role in early infant development. AB - The lipid fraction of human milk represents the main source of energy for the newborn infant and supplies essential nutrients such as fat-soluble vitamins and polyunsaturated fatty acids (PUFA). The essential fatty acids linoleic and alpha linolenic acids are precursors of long-chain polyunsaturated fatty acids (LC PUFA), such as arachidonic (C20:4 n-6) and docosahexaenoic (C22:6 n-3) acids, present in human milk in considerable amounts. LC-PUFA are indispensable structural components of all cellular membranes, and they are incorporated in relatively large amounts during early growth of the brain and the retina. Moreover, some LC-PUFA are precursors of eicosanoids, molecules with potent biological activity that modulates various cellular and tissue processes. The supply of long-chain fatty acids has been associated with functional outcomes of the recipient infants such as visual acuity and development of cognitive functions during the first year of life. Here we discuss the PUFA composition of human milk, factors which determine and modulate milk PUFA content, and possible effects of milk LC-PUFA on infant growth and development. PMID- 10527471 TI - Effects of nutrients in human milk on the recipient premature infant. AB - As the rate of survival of premature infants is increasing, more attention is necessarily focused on improving the quality of survival through optimal nutritional management. The nutritional needs of the premature infant are greater than at any other time in the life cycle. The benefits of human milk for term infants are well known. Emerging data suggest that human milk may especially benefit the premature infant. The human milk-fed premature infant may experience improved health (such as lower rates of infection and necrotizing enterocolitis), gastrointestinal function, and neurodevelopment. These factors may outweigh the concerns about adequate growth, nutrient accretion, and biochemical indices of nutritional status attributed to the lower nutrient content of human milk compared with preterm formula. Some of the nutritional concerns may be met by the use of multinutrient supplements during the time infants receive tube-feeding, generally the time prior to attaining complete oral feeding in-hospital. The available data suggest that the quality of survival of premature infants can be improved, both in the short-term and long-term, through the feeding of human milk. PMID- 10527472 TI - Impact of lactation on maternal body weight and body composition. AB - Women worldwide generally lose weight and body fat during lactation. This loss, although increased by longer, more intensive breastfeeding, is modest and may be reduced by increased food intake and decreased activity. Higher parity and older age are associated with greater weight loss postpartum among poorly nourished women. Well-nourished women or those who breastfeed only for a limited time may not return to their prepregnant weight or body composition by the end of the lactation period. Those who are overweight or obese may have difficulty initiating or maintaining lactation. For the majority of women in the world, lactation is unlikely to represent a threat to their health. To advise women on how to optimize their health and lactational performance, one must consider all of the changes in maternal nutritional status that occur during a reproductive cycle, which may or may not compensate for the modest decreases in body weight associated with lactation. PMID- 10527474 TI - Hematopathologists and cytopathologists: enemies or allies? PMID- 10527473 TI - Hormonal and dietary regulation of changes in bone density during lactation and after weaning in women. AB - Lactating women secrete approximately 250 mg of calcium in breast milk each day. Some of the calcium used for milk production comes from bone as women experience a transient 3-9% decrease in bone density during lactation. This loss appears to be obligatory and under hormonal regulation as lactation-induced bone loss occurs even when calcium intake is high. Bone mineral is recovered after lactation ceases or menses resume. Recovery of bone mineral appears to be complete even when pregnancies and lactations are closely spaced, and lactation does not increase future risk of osteoporotic fracture. Current data point to estrogen and parathyroid hormone-related peptide as regulating bone mobilization during lactation. The typical calcium regulatory hormones, parathyroid hormone, calcitriol and calcitonin, do not appear to stimulate bone resorption during lactation. Restoration of ovarian hormone production and decreased production of PTHrP2 are likely to result in the recovery of bone mineral after lactation has ceased. PMID- 10527476 TI - Analysis of cerebrospinal fluid cells by flow cytometry and immunocytochemistry in inflammatory central nervous system diseases: comparison of low- and high density cell surface antigen expression. AB - The examination of cerebrospinal fluid (CSF) continues to play an important role in the diagnosis of inflammatory diseases of the central nervous system (CNS). Immunocytochemistry and flow cytometry are the most commonly used methods for analysis of surface markers on CSF cells. We here compared these methods in the examination of CSF cells from a total of 68 patients with acute and chronic inflammatory CNS diseases. Expression of costimulatory molecules CD80 (B7-1) and CD86 (B7-2) as activation markers that are present at low density on the cell surface was analyzed in comparison to CD22 (B-cells) and CD4 (T-cell subset), that show high staining intensities. For CD22 and CD4, the results obtained with both methods were similar and reliable. Using flow cytometry, CD80 expression was detected in 6% of CSF cells in patients with chronic inflammatory CNS disease, as compared to 2% using immunocytochemistry, where the reliability of the data was found to be higher. We conclude that for examination of low-density surface markers on CSF cells, particularly with low cell counts, immunocytochemistry may be more reliable. PMID- 10527475 TI - Multinucleated giant cells in fine-needle aspiration of thyroid nodules: their diagnostic significance. AB - Multinucleated giant cells (MNGCs) are reported in many thyroid lesions. This study examines whether their quantity and quality can help in the differential diagnosis. All fine-needle aspirations (FNAs) of the thyroid with a "significant" number of MNGCs were reviewed from 1995 -1998. There were 23 cases (<1% of thyroid FNAs): 8 papillary carcinomas (PC), 4 subacute thyroiditis (ST), 3 granulomas, 7 adenomatous goiters (AG), and one Hurthle-cell adenoma (HA). MNGCs with dense cytoplasm were seen exclusively in PC, ST, and granulomas. They had angulated shapes. They were most numerous, largest, and with the highest number of nuclei in ST and granulomas. MNGCs with foamy cytoplasm were seen in AG and HA and 80% of the other cases (PC, ST, and granulomas). In PC, rare MNGCs had intranuclear inclusions and grooves. The accompanying cell population was characteristic of each disease. The quantity and quality of MNGCs in thyroid FNA may be helpful in narrowing the differential diagnosis. Diagn. Cytopathol. 1999;21:307-312. PMID- 10527477 TI - Aspiration cytology of extramammary tumors metastatic to the breast. AB - This study was carried out to examine the cytomorphologic features of metastatic breast tumors and to assess the utility of fine-needle aspiration cytology (FNAC) in diagnosing these tumors. The study group comprised five females and one male, all presenting with a breast mass. Their ages ranged between 35 and 65 years. FNAC of the breast mass was done in all cases. Three of the cases were previously diagnosed as squamous cell carcinoma (SCC) of the cervix, mucinous cystadenocarcinoma (MCA) of the ovary, and melanoma. Three cases presented initially with a breast mass. These included melanoma, non-Hodgkin's lymphoma (NHL), and plasmacytoma. The diagnosis of NHL was confirmed on histology. The patient with plasmacytoma presented primarily with a breast lump but subsequently developed multiple myeloma, and in one case of melanoma the primary tumor was detected after breast metastases. Preoperative FNAC of extramammary tumors metastatic to the breast is invaluable because the management of the patient differs entirely from that of a primary neoplasm. An accurate diagnosis can be made with the help of clinical and radiological correlation. If available, a perusal of previous history and biopsy material may prove useful. PMID- 10527478 TI - Fine-needle aspiration cytology of renal-cell adenocarcinoma metastatic to the breast: A report of three cases. AB - Metastases to the breast from extramammary primary malignancies, including renal adenocarcinoma, are rare. Fine-needle aspiration biopsy (FNA) is a useful, noninvasive, and rapid procedure to evaluate these mammary lesions. This study describes the cytomorphology of 3 cases of renal-cell adenocarcinoma metastatic to the breast. All patients had a prior history of renal-cell adenocarcinoma treated with radical nephrectomy, and they presented with a solitary mammary mass. The cytologic findings showed irregular clusters and dispersed single cells with eccentric nuclei and abundant, vacuolated cytoplasm in a hemorrhagic background. The nuclei were round to oval, with fine granular chromatin and a single, prominent nucleolus. All aspirates were interpreted initially and correctly as consistent with metastatic renal-cell adenocarcinoma. In summary, a cytologic diagnosis of renal-cell adenocarcinoma metastatic to the breast can be made by correlating clinical and cytologic findings. The distinction between metastatic extramammary malignancies to the breast and primary mammary carcinoma is critical to avoid unnecessary surgery and to ensure appropriate chemotherapy or radiation therapy. PMID- 10527479 TI - Cyst fluid cytology and chemical features in a case of lymphoepithelial cyst of the pancreas: A rare and difficult preoperative diagnosis. AB - Most pancreatic cysts (90%) are inflammatory pseudocysts. Approximately 10% of pancreatic cysts are neoplasms, including serous cystadenomas, and mucinous tumors, some of which are malignant. Analysis of pancreatic cyst fluid obtained by percutaneous or endoscopic fine-needle aspiration is increasingly being used for the preoperative diagnosis of pancreatic or peripancreatic cysts. However, cyst fluid chemical and cytologic features in less common types of pancreatic cysts have not been reported. Lymphoepithelial cyst of the pancreas is exceedingly rare, and only occasional individual reports have described cyst fluid findings. We report on a case of lymphoepithelial cyst of the pancreas developing in a middle-aged man. Cyst fluid aspirated under radiological guidance showed elevated levels of carcinoembryonic antigen (CEA), CA19-9, CA 125, and amylase, and a viscosity greater than that of serum. A cell block preparation of a fine-needle aspiration showed tissue fragments with a keratinized squamous lining and a lymphocytic infiltrate in the wall, and abundant background keratinous debris. The cytologic and biochemical findings in this case exhibit similarities to the findings reported in other reports, and may represent a recognizable pattern on cyst fluid analysis. PMID- 10527480 TI - Diagnostic intraocular fine-needle aspiration biopsy--An experience in three cases of retinoblastoma. AB - Ophthalmoscopically guided intraocular fine-needle aspiration biopsy (FNAB) under general anesthesia was performed in three suspected cases of retinoblastoma (Rb). The first case presented with uveitis and the second with unusual radiological features. The third case had classical clinical features of bilateral Rb but the parents refused enucleation. After a cytologic diagnosis of retinoblastoma, two cases were subjected to enucleation and a histologic confirmation was obtained. The present article discusses the technique of guided intraocular FNAB and its limitations as well as utility in the preoperative diagnosis of retinoblastoma (Rb). The authors are of the opinion that FNAB should be considered a viable diagnostic option in selected cases of leucocoria when conventional investigative modalities prove inconclusive or for tissue diagnosis when a patient of suspected Rb refuses enucleation. PMID- 10527481 TI - Chondroid chordoma: fine-needle aspiration cytology with histopathological, immunohistochemical, and ultrastructural study of two cases. AB - Chondroid chordoma is a controversial and confusing entity that was originally described by Heffelfinger et al. (Cancer 1973; 32:410-420) as a biphasic malignant neoplasm possessing elements of both chordoma and cartilaginous tissue. Fine-needle aspiration (FNA) cytology of chondroid chordoma has not been described. The aim of our investigation was to characterize the chondroid area of chondroid chordoma and to compare the FNA features with those of well differentiated chondrosarcoma. Clival and cervical spine chondroid chordomas were studied with light microscopy, immunohistochemistry, and electron microscopy. Chondroid chordomas demonstrated an epithelial nature by immunohistochemistry and ultrastructural studies. The FNA smears showed low cellularity, with loosely arranged or dispersed round cells in a myxoid background. Although the smears were similar to those of well-differentiated chondrosarcomas, they showed a positive reaction for epithelial markers. These findings reveal that chondroid chordoma is a variant of chordoma which possesses a hyaline matrix. Immunohistochemical demonstration of epithelial markers is useful to distinguish it from chondrosarcoma. Diagn. Cytopathol. 1999; 21:335-339. PMID- 10527482 TI - Cytologic features of metastatic sebaceous carcinoma: report of two cases with comparison to three cases of basal cell carcinoma. AB - The cytologic findings of two cases of metastatic sebaceous carcinoma are described and compared to three cases of locally recurrent basal cell carcinoma. Morphological findings for sebaceous carcinoma in fine-needle aspiration biopsy (FNAB) smears included cellular, loosely cohesive cell clusters with central necrosis, squamous pearl formation, and adjacent keratin debris. The tumor cells had moderate amounts of vacuolated cytoplasm, round to oval vesicular nuclei with clumped chromatin, nucleoli, some nuclear overlap, and numerous mitotic figures. An interesting finding was the presence of numerous multinucleated giant cells, probably responding to extravasated lipid or keratin material. In contrast, the FNAB smears of basal cell carcinoma typically were less cellular, with more tightly cohesive and smaller clusters of uniform hyperchromatic basaloid cells with high nuclear to cytoplasmic ratios, and a narrow rim of cytoplasm without vacuolization. The morphologic features of sebaceous carcinoma in FNAB smears appear to be distinct from those of basal cell carcinoma. FNAB can be a useful preoperative diagnostic technique to distinguish these two cutaneous malignancies. PMID- 10527483 TI - Cytologic features of lymphoepithelial cyst of the pancreas: two preoperatively diagnosed cases based on fine-needle aspiration. AB - We describe the cytologic features seen in fine-needle aspiration (FNA) specimens from two cases of preoperatively diagnosed lymphoepithelial cyst (LEC) of the pancreas. Pancreatic LEC is a rare, true cyst of uncertain histogenesis that may clinically and radiologically mimic a pseudocyst or cystic neoplasm. Both our patients were middle-aged men who presented with vague abdominal pain. Computed tomography (CT) of the abdomen revealed a mass in or around the pancreas, and CT guided percutaneous FNA (patient 1) and endoscopic ultrasound-guided FNA (patient 2) yielded paste-like yellow-gray material. Cytologic smears showed numerous anucleated squamous cells in a background of keratinous and amorphous debris. A few benign nucleated squamous cells and plate-like cholesterol crystals were also seen. Unlike LEC of the head and neck region, only rare lymphocytes and histiocytes were present. Pancreatic LEC was diagnosed based on these cytologic findings and was histologically confirmed following cyst enucleation (patient 1) and partial pancreatectomy (patient 2). We conclude that preoperative FNA and recognition of the characteristic cytologic pattern will enable conservative surgical management of pancreatic LEC. Diagn. Cytopathol. 1999;21:346-350. PMID- 10527484 TI - ThinPrep vs. conventional smears in fine-needle aspirations of sarcomas: A morphological and immunocytochemical study. AB - Very limited data exist describing the characteristics of sarcomas sampled by fine-needle aspiration (FNA) and processed by the ThinPrep (TP) method. We compared the cytopathological and immunocytochemical features of sarcoma aspirates prepared using both the conventional and TP method. We reviewed 70 sarcoma FNAs. Samples were first used to prepare conventional smears and the remainder of the specimen was rinsed in Cytolyt. The average number of slides examined per case was two for the TP method and five for the conventional technique. Immunocytochemistry for different markers was performed in a subset of cases. Sixty-five cases were positive for sarcoma both by conventional and TP methods. Five cases were positive by one method only. Cellularity was higher on conventional slides. In terms of cytoarchitecture, TP slides revealed fewer thick clusters, more single cells that were more evenly distributed, and sometimes distortion of expected cellular arrangements and architectural patterns. Cytomorphological and nuclear details were better preserved on TP slides. The background of TP slides revealed a reduction of blood but also some loss of necrosis and characteristic background tumor features. Immunocytochemical staining revealed superior results on TP slides. TP and conventional slides showed good correlation. TPs were excellent for immunocytochemistry and represent an alternative to conventional smears when expertise in slide preparation is not available. However, TPs may require additional experience in the interpretation of sarcomas, mainly related to the loss of tumor-specific background features. They are useful as an adjuvant to conventional smears in sarcoma diagnoses, particularly when special studies are needed. Diagn. Cytopathol. 1999;21:351-354. PMID- 10527485 TI - AutoPap system detection of infections and benign cellular changes: results from primary screener clinical trials. AB - Primary screening devices for cervical cytology must show performance data for the detection of infectious organisms and benign cellular changes (BCC) for cytologists who routinely report these findings. The data on infection and BCC from the AutoPap primary screening clinical trials are presented herein. The presence of infectious organisms (candida, trichomonas, shift in bacterial flora, herpes, actinomyces) and BCC were noted in each of the clinical trial arms (current practice, CP; AutoPap-assisted practice, AP). For the purposes of these analyses, a report of infection or BCC from either arm was considered to be "truth." In 25,124 slides analyzed, there were 2,925 cases of infection identified. Of these, CP identified 2,141, and AP identified 1,985. The overall detection results are statistically equivalent. Of 17 cases of actinomyces, CP detected 8, while AP detected 12. Of 1,282 cases of candida, CP detected 983, and AP detected 865. Of 1,375 cases of shift of bacterial flora, CP detected 897, and AP detected 869. Of 14 cases of herpes, CP detected 9, and AP detected 11. Of 343 cases of trichomonas, CP detected 293, and AP detected 275. There were 5,156 cases of BCC identified in the trial. CP detected 3,431, and AP detected 3,276. The detection rates for BCC are statistically equivalent. The results show that the AutoPap-assisted practice for the primary screening of conventional cervical cytology slides is equivalent to the current practice for the detection of cervical infections and benign cellular changes. PMID- 10527486 TI - Malignant histiocytosis in childhood: A case report. AB - A case of malignant histiocytosis (MH) in a 12-year-old male child is described. Fever and wasting were the most prominent symptoms. Bilateral cervical and axillary lymphadenopathy was present along with hepatosplenomegaly. The haematological, cytological and histopathological features are described. The diagnostic dilemma presented during the diagnosis of this case is also discussed. Diagn. Cytopathol. 1999;21:359-361. PMID- 10527487 TI - Renal-cell carcinoma: cytologic diagnosis in a child. PMID- 10527488 TI - FNA cytology of epithelial-myoepithelial carcinoma. PMID- 10527489 TI - Comparison of ThinPrep and conventional preparations: urine cytology evaluation. PMID- 10527490 TI - The fluorescent cationic dye rhodamine 6G as a probe for membrane potential in bovine aortic endothelial cells. AB - The membrane potential of cultured bovine aortic endothelial cells was assessed by a fluorescent probe as an alternative to direct methods. We used the fluorescent cationic dye rhodamine 6G, a lipophilic probe with high permeability in cell membranes. A linear relationship was obtained between fluorescence intensity (F.I.) and membrane potential (Em) as a function of the extracellular Na(+) concentration in the presence of the ionophore gramicidin. From the equation derived from the linear relationship F.I. = -0.004 Em + 0. 03 (P < 0.001), the fluorescence measurements could be converted to membrane potential. The resting plasma membrane potential obtained was -65 +/- 7 mV. Nigericin (27 microM), ouabain (1 mM), and bradykinin (20 nM) induced a decrease in F.I. (depolarization), while ATP (25-100 microM) induced an increase in F.I. (hyperpolarization). Mitochondrial membrane potential inhibitors myxothiazol (3 microM) and oligomycin (4 microM) did not influence F. I. measured in the cultured bovine aortic endothelial cells. The results indicate that rhodamine 6G can be used as a sensitive and specific dye in studies of substances that affect the membrane potential of endothelial cells. PMID- 10527491 TI - Detection of DNA hybridization using the TISPR-1 surface plasmon resonance biosensor. AB - Biotinylated oligonucleotide probes were immobilized to the gold sensor surface of the TISPR-1 miniature integrated surface plasmon resonance liquid sensor system for the purpose of detecting specific DNA hybridization. The immobilization of the oligonucleotide capture probes was carried out through streptavidin-biotin binding technology. The sensor detected the immobilization of unlabeled DNA through shifts in index of refraction as the molecules entered and remained selectively bound to the surface in the vicinity of the exponentially decaying surface plasmon resonance wave. The surface immobilization chemistry was proven to be stable for long periods of time, reproducible, and practical for detecting DNA hybridization with the TISPR-1. DNA hybridization was detected as a slow, positive, and small (when compared to protein-protein or antibody-antigen binding experiments) increase in the measured index of refraction under passive hybridization conditions by the TISPR-1 sensor. The DNA hybridization signal was significant (index of refraction change of 0.001) when large fragment PCR amplified DNA products were hybridized to the oligonucleotide probes (S/N = 6 10). The DNA hybridization techniques were demonstrated using DNA sequences from the HIV genome which encode the Tat and Rev genes. PMID- 10527492 TI - Quantitative immunoblots of proteins resolved from brain homogenates: underestimation of specific protein concentration and of treatment effects. AB - Estimation of the concentration of a specific protein in a biological sample often is obtained by analysis of immunoblots. We used this technique to estimate the concentration of three proteins present in homogenates of brain: glial fibrillary acidic protein (GFAP), myelin basic protein (MBP), and synapsin I. Homogenates prepared from rat brains known to contain more than 6-fold increases in GFAP, based on a GFAP enzyme-linked immunosorbent assay (ELISA), showed only small relative increases in this protein when the same samples were subjected to immunoblot analysis with polyclonal or monoclonal anti-GFAP; quantification was based on PhosphorImager analysis of [(125)I] protein A bound to the antibodies. Estimates of GFAP in the GFAP-enriched samples approached the expected 6-fold increase when the total protein load per gel lane was reduced from 30 to 1 microgram. Pure GFAP run as standard was not affected by 10-fold increases in protein load, but spiking brain homogenates with pure GFAP "quenched" the values obtained for the standard run alone. Examination of the quenching potential of pure brain tubulin, a protein that nearly comigrates with GFAP on SDS gels, showed that it may be one component of brain homogenates that contributes to masking of immunodetection of GFAP. The effect of total brain homogenate proteins on the signal obtained for a specific protein was not limited to GFAP; similar effects were observed for MBP and synapsin I. The data indicate that estimates of the concentration of a specific protein, whether as a function of its relative amount in a given protein mixture or its relative amount in one mixture compared to another, are influenced by other homogenate proteins present in the mixture. PMID- 10527493 TI - Binding study of semotiadil and levosemotiadil with alpha(1)-acid glycoprotein using high-performance frontal analysis. AB - High-performance frontal analysis (HPFA) was used to investigate the binding properties of human alpha(1)-acid glycoprotein (AGP) with semotiadil ((R)-isomer, Ca-channel blocker) and its antipode levosemotiadil ((S)-isomer, Ca- and Na channel blockers). An on-line HPLC system consisting of a HPFA column, an extraction column, and an analytical HPLC column was used to determine the unbound concentrations of these enantiomers, and the experimental data were subsequently subjected to the Scatchard analyses to estimate their binding parameters. The binding affinity of the (R)-isomer (K = 3.17 x 10(7) M, n = 0.74) is approximately 1.2 times stronger than that of (S)-isomer (K = 2.59 x 10(7) M, n = 0.74). An enantioselective competitive binding study indicated that both enantiomers are bound at the same site on AGP molecules. PMID- 10527494 TI - Enzymatic cycling assay for D-carnitine determination. AB - An enzymatic method for d-carnitine determination using the enzyme d-carnitine dehydrogenase is described. The assay is based on the amplified signal produced during NAD(+) cycling in the presence of a tetrazolium salt and using phenazine methosulfate as electron carrier. Optimum assay conditions were studied with two tetrazolium salt pairs: 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT)/MTT-formazan and 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyl tetrazolium chloride (INT)/INT-formazan. The first pair (MTT) showed higher sensitivity. The calibration curve was linear from 0.1 to 5 mM d-carnitine, with a quantification limit of 0.1 mM and a relative standard deviation of 1.51%. The procedure is simple, rapid, accurate, and easily automated. It was satisfactorily applied to following d-carnitine levels during the microbial transformation of d carnitine into l-carnitine and to determining the d-carnitine content of pharmaceutical preparations. PMID- 10527495 TI - Chemically synthesized ubiquitin extension proteins detect distinct catalytic capacities of deubiquitinating enzymes. AB - We have used solid-phase chemistry to synthesize proteins equivalent to a human ubiquitin precursor (ubiquitin-52-amino-acid ribosomal protein fusion; UBICEP52) and representative of isopeptide-linked ubiquitin-protein conjugates [ubiquitin (epsilonN)-lysine]; these proteins were precisely cleaved by a purified recombinant Drosophila deubiquitinating enzyme (DUB), UCH-D. Along with the previously synthesized ubiquitin-(alphaN)-valine, these synthetic proteins were used as substrates to assess the catalytic capacities of a number of diverse DUBs expressed in Escherichia coli: human HAUSP; mouse Unp; and yeast Ubps 1p, 2p, 3p, 6p, 11p, and 15p and Yuh1p. Distinct specificities of these enzymes were detected; notably, in addition to UCH-D, isopeptidase activity [ubiquitin (epsilonN)-lysine cleavage] was only associated with Yuh1p, Unp, Ubp1p, and Ubp2p. Additionally, human placental 26S proteasomes were only able to cleave UBICEP52 and ubiquitin-(epsilonN)-lysine, suggesting that 26S proteasome associated DUBs are class II-like. This work demonstrates that the synthetic approach offers an alternative to recombinant methods for the production of small proteins in vitro. PMID- 10527496 TI - "Linkage region" sequences of heparins and heparan sulfates: detection and quantification by nuclear magnetic resonance spectroscopy. AB - The (13)C NMR spectra of most heparin and heparan sulfate preparations display minor signals not attributable to the glycosaminoglycan chains of these polysaccharides. These signals have been "concentrated" in oligosaccharides isolated from an acid hydrolyzate of heparin and shown to arise from the sequence GlcA-Gal-Gal-Xyl of the "linkage region" (LR) connecting the carbohydrate chains to the peptide chains in the original proteoglycans. Mono- and two-dimensional (1)H and (13)C NMR analysis of the major oligosaccharide (LR-OLIGO) indicated the prevalent structure GlcA-GlcNAc-GlcA-Gal-Gal-Xyl, where GlcNAc is partially 6-O sulfated. (13)C NMR signals at 84.6 and 85.0 ppm, arising from C-3 of the two Gal residues, lend themselves to easy detection and quantification of the linkage region in heparins and heparan sulfates and can be used to assess the importance of the LR in the modulation of various biological activities of these glycosaminoglycans. PMID- 10527497 TI - Simultaneous analysis of hemoglobin adducts of acrylamide and glycidamide by gas chromatography-mass spectrometry. AB - Acrylamide (AA) is a carcinogen in experimental animals. Glycidamide (GA), formed by metabolic epoxidation of AA, is believed to be responsible for the carcinogenicity of AA. Occupational exposure to AA has been assessed earlier by measurement of its adducts with N-terminal valine in hemoglobin. A background of AA adducts [N-(2-carbamoylethyl)valine (AAVal), about 30 pmol/g globin] was found in individuals without known exposure to the compound. The method previously available for adducts of GA only allowed analysis of samples from highly exposed individuals and showed similar levels of AAVal and adducts of GA [N-(2-hydroxy-2 carbamoylethyl)valine (GAVal)]. We have developed a sensitive method for simultaneous quantification of adducts of GA and AA, which is suitable down to low exposure levels. The method is based on the so-called modified Edman method, where globin is reacted with pentafluorophenyl isothiocyanate under neutral conditions. The valine adducts are then extracted in the form of pentafluorophenylthiohydantoin (PFPTH) derivatives. The analytical procedure included reaction of the PFPTH derivatives with acetic anhydride in order to protect the hydroxyl group of GAVal. The PFPTH derivatives of AAVal and GAVal were analyzed by gas chromatography/tandem mass spectrometry. ((2)H(3))AAVal PFPTH was used as the internal standard. The method was applied to samples from 11 workers at an AA production plant, 1 nonexposed nonsmoker, and a few participants of a smoking cessation program. AAVal levels were in the range 27 1854 pmol/g globin. Recorded levels of GAVal were 3-12% of those of AAVal, suggesting that previous measurements of GAVal overestimate GAVal at low levels of exposure to AA. PMID- 10527498 TI - Chromogranin A from bovine adrenal medulla: molecular characterization of glycosylations, phosphorylations, and sequence heterogeneities by mass spectrometry. AB - Chromogranin A (CGA) is a member of a family of acidic glycoproteins present in endocrine and neuroendocrine tissues. One of its suggested physiological roles is being a precursor molecule for several peptide hormons. Further interest in this protein has recently originated from its potential role in pathophysiological processes of Alzheimer's disease. The concentration of CGA in the brain has been used for diagnosis of this disease, and CGA as an insoluble deposit has been found in the extracellular beta-amyloid plaques. By developing a new purification procedure we were able to isolate abundant CGA in high purity from bovine chromaffin cells. A MALDI-MS analysis of the intact protein revealed a heterogeneous molecular mass of ca. 50 kDa, indicating several structure modifications. By use of several subsequent proteolytic/chemical cleavage steps, HPLC isolation, a newly developed deglycosylation procedure, and several MS and MS-MS fragmentation approaches, the complete primary structure of CGA including four sequence heterogeneities, two O-glycosylations, five phosphorylations, and one disulfide bridge could be characterized. For both glycans six different forms could be identified. Ser167 was found to be mainly glycosylated by a trisaccharide, and Thr231 was found to be mainly glycosylated by a tetrasaccharide. Ser81, Ser124, and Ser297 residues were partially phosphorylated, whereas Ser372 and Ser377 were found completely phosphorylated. Sequence heterogeneities were identified in positions 293 (H/R), 301 (K/E), and 373 (Q/R) and at the partly missing C-terminal residue. Furthermore, a disulfide bridge between Cys17 and Cys38 was ascertained. PMID- 10527500 TI - A quantitative analysis of serum sulfatide by matrix-assisted laser desorption ionization time-of-flight mass spectrometry with delayed ion extraction. AB - Matrix-assisted laser desorption ionization time-of-flight mass spectrometry with delayed ion extraction (DE MALDI-TOF MS) was applied for the first time for the quantitation of sulfatide content in serum at the picomole level. The total lipids extracted by n-hexane:isopropanol (3:2, v/v) from 100 microliter of serum were saponified to convert sulfatide to its lyso form, and then the lysosulfatide was directly determined using DE MALDI-TOF MS in the presence of other degraded lipids. Hydrogenated N-acetyl lysosulfatide was used as an internal standard. The relative peak height of sulfatide was calculated and plotted versus its contents. This plot showed linearity between 2 pmol and 1 nmol of sulfatide (regression coefficient r > 0.95). Sulfatide contents of normal human sera and rabbit serum were quantitated by this method. The results corresponded well to the reported data determined by gas-liquid chromatography. This new approach was found to be sensitive, convenient, and reliable. It is expected to be applied to quantitate sulfatide from other small amounts of body fluids or tissues and to clinical examination. It is also expected to be applicable to quantitate other glycosphingolipids. PMID- 10527499 TI - Development and validation of an immunoreceptor assay for simulect based on surface plasmon resonance. AB - Simulect is a chimeric human/mouse antibody directed against interleukin-2 (IL-2) receptor. A combined immuno- and receptor assay has been developed and validated to characterize the production of Simulect batches. This assay is based on surface plasmon resonance (SPR) technology. In each experiment two successive interactions were monitored: the direct binding of Simulect to an anti-human IgG antibody, followed by the direct binding of IL-2-soluble receptor to the preformed anti-human IgG antibody/Simulect complex. Based on the first interaction a direct immunoassay for Simulect was optimized and validated. Based on the second interaction a direct receptor assay for Simulect biological activity was optimized and validated. The assays were validated by performing three independent assays on 3 different days. The intra- and interday variations of the immunoassay (expressed as % CV) were, respectively, 1.7 and 1.6%. The overall accuracy for the immunoassay was 98.5% +/- 1. The intra- and interday variations of the receptor assay (% CV) were, respectively, 1.6 and 3.7%. The overall accuracy of the receptor assay was 100% +/- 2. Four batches of Simulect were compared to a reference batch. The results did not show significant differences for the immunoreactivity. However, the results of the receptor assay showed accuracies which were apparently higher than 100%. This was explained by a slight degradation of the reference batch after few years of storage. These results demonstrate the advantage of this method combining evaluation of the immunological and biological integrity of the drug and a high reproducibility in accuracy and precision of the biosensor-based technology. PMID- 10527501 TI - Critical increment of Lewis blood group antigen in serum by cancer found by photothermal immunoassay. AB - Lewis blood group antigen levels in human sera were assayed with a highly sensitive photothermal immunoassay which is based on laser-induced photothermal detection. Comparison of 32 colon cancer patients' sera and 34 healthy persons' sera showed that cancer patients' sera contained more Lewis antigens than healthy persons' sera. Le(a) antigen level in Le(a-) type persons and Le(b) level in Le(b ) type persons differentiated healthy persons and colon cancer patients. Furthermore, it was found that in Lewis blood phenotype (a-) several cancer patients' sera specimens changed to (a+). Many reports demonstrated that Lewis phenotype of erythrocytes changed with various conditions, including carcinomas, but they dealt mostly with erythrocytes and salivas or showed immunohistochemical evidence, and there are no reports on the quantitative analysis of ordinary (noncancerated) Lewis antigen levels in human sera. This is because Lewis antigens in sera, unlike those found in saliva, are too small to quantify with conventional immunoassay and there has been no highly selective method to measure Lewis antigens in sera. The increase of Lewis antigen in cancer patients' sera is presumed to antecede the blood type change. Our assay presented here, a highly sensitive assay of Lewis antigens, will greatly contribute to an early detection or diagnosis of cancers. PMID- 10527502 TI - Radiolabeling of receptor ligands by chemical incorporation of phosphorylation sites. AB - A chemical reagent, N-acetyl-cys((succinimidyl-6-(thioacetyl)amino) hexanoate) ser-arg-arg-ala-ser-val-tyr-amide ("phosite NHS ester"), allowing the introduction of phosphorylation sites into proteins, peptides, or small molecules, has been synthesized and characterized. The phosite reagent enables the enzymatic radiolabeling of any protein, peptide, or small molecule containing a reactive amine using [(32)P] or [(33)P]ATP and protein kinase A. The utility of the reagent has been demonstrated in cytokine and G protein-coupled radioligand receptor binding assays using whole cell and immobilized receptor formats. Use of the reagent does not require genetic manipulation of the target ligand. PMID- 10527503 TI - Sensitive high-performance liquid chromatographic method for profiling phytoestrogens using coulometric electrode array detection: application to plasma analysis. AB - An HPLC method for profiling 13 phytoestrogens and their metabolites using coulometric electrode array detection was developed. Sensitivity of the method was slightly less than that of our GC-MS method, but significantly higher compared to the HPLC methods using diode-array or UV detection. Detection limits varied from 3.4 (secoisolariciresinol) to 40.3 (genistin) pg on column. Signal linearities ranged from the detection limits to 61 ng on column. Resolution values for the peak pairs varied from 1.1 (O-desmethylangolensin anhydrosecoisolariciresinol) to 16 (daidzin-genistin). Intra- and interassay retention time variations were negligible and detector response variation was eliminated by frequent calibration. Chromatographic method was applied to plasma analyses and 6 of the 13 compounds were detected. Method accuracy for those six analytes varied from 69% (enterodiol) to 118% (genistein). Intraassay precision CVs ranged from 1.5% (enterolactone, 12.4 nmol/liter) to 14% (genistein, 245 nmol/liter) and interassay precision CVs ranged from 9.9% (daidzein, 67.4 nmol/liter) to 44% (enterodiol, 1.20 nmol/liter). PMID- 10527504 TI - Influence of the antibody purification method on immunoassay performance: hapten antibody binding in accordance with the structure of the affinity column ligand. AB - The effects of ligands for immunoaffinity chromatography on the immunoassay were investigated with three goat anti-methamphetamine (anti-MA) antibodies (Abs). An N-4-aminobutyl derivative of methamphetamine (4-ABMA) was conjugated with proteins and used as immunogens. All the antisera produced were purified by affinity chromatography with various ligands of 4-ABMA-proteins and of haptens as well as protein G: 4-ABMA-bovine serum albumin (4-ABMA-BSA), 4-ABMA-keyhole limpet hemocyanine (4-ABMA-KLH), 4-ABMA-ovalbumin (4-ABMA-OVA), MA, 4-ABMA, and amphetamine were used as ligands. Enzyme-linked immunosorbent assay (ELISA) was conducted to examine characteristics of the purified Abs with the 4-ABMA-OVA competitor coated. The results obtained revealed that characters of the purified Abs were closely related with chemical structures of ligands used. The Abs from the MA and the amphetamine columns showed better sensitivities than those from the others in each antiserum. Particularly, the Ab from the amphetamine column gave the best results in terms of sensitivity and specificity. The recognition or the affinity of the Ab selected was considered to be affected by the structure of the ligand concerned. These results suggest that the Ab purification method should be considered as an important parameter which has great influence on the performance of immunoassays with polyclonal Abs. PMID- 10527505 TI - Analysis of glutathione and glutathione disulfide in whole cells and mitochondria by postcolumn derivatization high-performance liquid chromatography with ortho phthalaldehyde. AB - A method is described for the detection of glutathione (GSH) and glutathione disulfide (GSSG) based on a HPLC postcolumn reaction with ortho-phthalaldehyde (OPT) at pH 12 followed by fluorescence detection. Although similar methods have been reported, the high pH of the postcolumn reaction adds considerable selectivity and sensitivity to the measurement of GSH and glutathione disulfide. The limit of detection approaches 100 fmol, which is sufficient to detect whole cell glutathione disulfide in 10,000 cells or mitochondrial glutathione disulfide in 20 million cells. Using this method, glutathione and glutathione disulfide were measured in human lymphocytes, granulocytes, and cultured Jurkat T cells, as well as in the corresponding samples of mitochondria. The percentage of glutathione disulfide to total glutathione in whole-cell extracts was approximately 1%. In contrast, the percentage was relatively high in mitochondria, with the mitochondria of granulocytes having the highest (25%) followed by those of lymphocytes (15%) and finally by cultured Jurkat T cells (9%). This method extends the analysis of glutathione and glutathione disulfide to mitochondria obtained from a relatively small number of cells. PMID- 10527506 TI - A continuous assay for the spectrophotometric analysis of sulfotransferases using aryl sulfotransferase IV. AB - We have developed a continuous spectrophotometric coupled-enzyme assay for sulfotransferase activity. This assay is based on the regeneration of 3' phosphoadenosine-5'-phosphosulfate (PAPS) from the desulfated 3'-phosphoadenosine 5'-phosphate (PAP) by a recombinant aryl sulfotransferase using p-nitrophenyl sulfate as the sulfate donor and visible spectrophotometric indicator of enzyme turnover. Here recombinant rat aryl sulfotransferase IV (AST-IV) is expressed, resolved to the pure beta-form during purification, and utilized for the regeneration. The activity of betaAST-IV to catalyze the synthesis of PAPS from PAP and p-nitrophenyl sulfate is demonstrated via capillary zone electrophoresis, and the kinetics of this reverse-physiological reaction are calculated. betaAST IV is then applied to the coupled enzyme system, where the steady-state activity of the commercially available Nod factor sulfotransferase is verified with an enzyme concentration study and substrate-specificity assays of N-chitoses. The potential applications of this assay include rapid kinetic determinations for carbohydrate and protein sulfotransferases, high-throughput screening of potential sulfotransferase substrates and inhibitors, and biomedical screening of blood samples and other tissues for specific sulfotransferase enzyme activity and substrate concentration. PMID- 10527508 TI - A continuous spectrophotometric linked enzyme assay for transglutaminase activity. PMID- 10527507 TI - Preparation of recombinant histone H3 as a substrate for protein kinase assays. PMID- 10527509 TI - Detection of proteins after biotinylation within polyacrylamide gels (BIG method). PMID- 10527510 TI - Charge heterogeneity of commercial, red-shifted recombinant green fluorescent protein, revealed by capillary zone electrophoresis under nondenaturing conditions. PMID- 10527511 TI - Protein analysis using enzymes immobilized to paramagnetic beads. AB - A new method combining protein chemistry with enzymes immobilized to paramagnetic beads is presented. The immobilized enzymes can substitute for regular enzymes in a number of protein chemistry protocols, resulting in faster reaction times, less sample contamination, and improved interfacing to modern procedures, like mass spectrometry. Trypsin was used as a model enzyme to test the amount of protein coupled to glass beads and the degree of autodigestion when analyzed by MALDI-MS and HPLC. Immobilization of trypsin resulted in digestions comparable with standard solution digestions using fetuin as a model substrate. Furthermore, fetuin was used to test the stability of the enzyme-coated beads. No apparent loss of enzyme activity was observed after 10 times reuse of trypsin-coated beads. Immobilization of exo- and endoglycosidases to paramagnetic beads resulted in high sensitivity, faster sequential glycosidase digestion of glycopeptides, and reduced sample contamination. All digestions could be performed in less than 24 h, when a tryptic glycopeptide from human lung proteinosis surfactant protein A was used as model compound. PMID- 10527512 TI - Application of mercury cold vapor atomic fluorescence spectrometry to the characterization of mercury-accessible -SH groups in native proteins. AB - A new analytical approach has been applied to the determination and characterization of mercury-accessible -SH groups in pure native protein samples (ovalbumin, hemoglobin, glyceraldehyde-3-phosphate dehydrogenase, aldolase, pyruvate kinase, hexokinase, lactate dehydrogenase, alcohol dehydrogenase, creatine phosphokinase, lysozyme, and cytochrome c). The method is based on the selective reduction of Hg(II) in the presence of Hg(II)-thiol complexes with alkaline sodium tetrahydroborate, to give Hg(0) in a continuous flow reaction system coupled with atomic fluorescence spectrometric (AFS) detection. The method is fast and specific and allows one to work with nanomole amounts of a single protein without any preliminary incubation and without any separation of Hg(II) from thiol-complexed mercury. The meaning of the results obtained in the determination of the accessible -SH groups in native proteins by using chemical probes is discussed. PMID- 10527513 TI - Direct analysis of the products of sequential cleavages of peptides and proteins affinity-bound to immobilized metal ion beads by matrix-assisted laser desorption/ionization mass spectrometry. AB - Consecutive enzymatic reactions on analytes affinity-bound to immobilized metal ion beads with subsequent direct analysis of the products by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry have been used for detecting protein synthesis errors occuring at the N-terminus. The usefulness of this method was demonstrated by analyzing two commercially available recombinant HIV proteins with affinity tags at the N-terminus, and histatin-5, a peptide with multiple histidine residues. The high specificity, sensitivity, and speed of analysis make this method especially useful in obtaining N-terminal sequencing information of histidine-tagged recombinant proteins. PMID- 10527514 TI - The efficient removal of endotoxins from insulin using quaternized polyethyleneimine-coated porous zirconia. AB - The synthesis and use of a zirconia-based, alkali-stable strong anion-exchange stationary phase are described for the removal of pyrogenic lipopolysaccharides (LPS) from insulin. The strong anion-exchange material is produced by deposition of polyethyleneimine (PEI) onto porous zirconia particles, followed by cross linking with a novel reagent, 1,2-bis-(2-iodoethoxy) ethane, and quaternization with iodomethane. Physical characterization of the chromatographic support shows that it has an ion-exchange capacity of 0.6 mmol/g, and 82% of the amine sites on the surface are in quaternized form. Isocratic elution of small benzoic acid derivatives shows good column efficiency. The two primary virtues of this material are its chemical stability under alkali conditions (up to pH 13) and its lower hydrophobicity compared to previously described alkali-stable PEI-coated zirconia supports cross-linked with 1,10-diiododecane. Using this new zirconia based phase, a purification protocol is developed for the efficient removal of Escherichia coli 0111:B4 LPS from bovine insulin samples. An endotoxin clearance rate of up to 1.3 x 10(8) was attained. Endotoxin levels were reduced to less than 5 endotoxin units/ml even at initial contamination levels as high as 5.0 x 10(6) endotoxin units/ml. Furthermore, endotoxin adsorbed to the porous zirconia column may be easily removed (depyrogenated) using alkali for repeated purification cycles. PMID- 10527515 TI - Comparison of the tetrazolium salt assay for succinate dehydrogenase with the cytosensor microphysiometer in the assessment of compound toxicities. AB - The cytosensor microphysiometer (a biosensing instrument for detecting cellular metabolism) was compared to the established tetrazolium salt assay as a chemosensitivity test. Two coumarin compounds, 7-hydroxycoumarin and esculetin, were examined to determine their effect on the cellular metabolism of A431 cells over a 24-h exposure period. In the tetrazolium salt assay, 7-hydroxycoumarin caused suppression of the succinate dehydrogenase activity at concentrations greater than 10 microg/ml. Esculetin exerted a more serious effect on succinate dehydrogenase, with decreases in activity observed at greater than 1 microg/ml. The observed effect was dose-dependent for both compounds examined. The metabolic activities of cells exposed to 100 microg/ml of drug were 90.37 +/- 2.8 and 71.62 +/- 2.96 (n = 3), of control values, for 7-hydroxycoumarin and esculetin, respectively. Using the cytosensor microphysiometer to assess metabolic activities, a similar pattern of inhibition was observed, with esculetin more detrimental to cellular metabolism than 7-hydroxycoumarin. The effect was dose- and time-dependent for both compounds. 7-Hydroxycoumarin (100 microg/ml) caused the cellular metabolic rate to drop to 44.21 +/- 5.34% (n = 4) of the control metabolic rate, while 100 microg/ml esculetin caused the metabolic rate to fall to 21.5 +/- 4.54% (n = 4) of the control rate. The cytosensor method proved to be superior to the tetrazolium salt assay for a number of reasons, which are discussed in this paper. PMID- 10527516 TI - Development of a continuous, fluorometric coupled enzyme assay for thyrotropin releasing hormone-degrading ectoenzyme. AB - Thyrotropin-releasing hormone degrading-ectoenzyme (TRH-DE) (EC 3.4. 19.6), removes the N-terminal pyroglutamyl residue of thyrotropin-releasing hormone (TRH). Discontinuous assays have been used to measure TRH-DE activity; however, a continuous assay is needed to make reliable measurements of initial rates and facilitate kinetic studies. Presented is a continuous, coupled enzyme assay for TRH-DE in which TRH-DE hydrolyzed the substrate, pyroglutamyl-histidyl prolylamido-4-methyl coumarin (TRHMCA), to give His-ProMCA, which was then cleaved by dipeptidyl peptidase IV (EC 3.4.14.5) to give 7-amino-4-methyl coumarin (MCA). Reaction progress was monitored continuously by measuring the increase in MCA fluorescence. This assay should be especially useful for rapid screening of potential TRH-DE inhibitors. A previously reported discontinuous assay, where nonenzymatic cyclization at 80 degrees C was used to liberate MCA from His-ProMCA, was found to underestimate the amount of product formed. A modified procedure that avoids this is presented. Initial rates and kinetic parameters for TRHMCA hydrolysis by TRH-DE determined using this modified assay correspond with those determined by the continuous assay. Discontinuous and continuous assays gave K(m) values for TRHMCA of 3.4 +/- 0.7 microM (n = 5) and 3.8 +/- 0.5 microM (n = 5), respectively. K(i) values determined by the discontinuous assay for TRH and TRH-OH were 35 +/- 4 microM (n = 3) and 311 +/- 31 microM (n = 5), respectively. PMID- 10527517 TI - Characterization of the carbohydrate binding specificity and kinetic parameters of lectins by using surface plasmon resonance. AB - An accurate, rapid, and sensitive method for characterizing the carbohydrate binding properties of lectins using a BIAcore apparatus and the detection method of surface plasmon resonance is described. As a model study, the sialic acid binding lectins from Sambucus nigra and Maackia amurensis, which are specific for the epitopes Neu5Ac(alpha2-6)Gal and Neu5Ac(alpha2-3)Gal, respectively, were chosen as suitable candidates. Two systems, one for the analysis of oligosaccharides and the other for glycoproteins, were developed after a rigorous analysis and evaluation of such parameters as binding conditions, buffers, and regeneration conditions. The systems take into account nonspecific binding, using the respective denatured lectin as negative blank, and avoid loss of activity: regeneration of the surface using either 10 mM NaOAc (pH 4.3) buffer (oligosaccharide system) or 20 mM HCl (glycoprotein system). The specificity of the lectins is well illustrated, while the kinetics parameters are shown to be sensitive to subtle changes in the recognized epitopes, and to be affected by steric hindrance. Surface plasmon resonance is a suitable technique for the analysis and characterization of lectins. PMID- 10527518 TI - Identification of isoflavone metabolites dihydrodaidzein, dihydrogenistein, 6'-OH O-dma, and cis-4-OH-equol in human urine by gas chromatography-mass spectroscopy using authentic reference compounds. AB - The metabolic products of daidzein and genistein, the principal isoflavones of soy, were examined. Six volunteers included soy into their normal diet for a 2 week period and urine samples were analyzed before and after soy consumption. Isolation and characterization of the urinary metabolites were carried out with absorption chromatography on Sephadex LH-20 and gas chromatography-electron ionization mass spectrometry (GC-EIMS). The structures of the isoflavones isolated were confirmed by using authentic reference compounds. Dihydrogenistein, 6'-OH-O-desmethylangolensin, and cis-4-OH-equol were identified, in addition to known isoflavonoids daidzein, genistein, glycitein, and the known metabolites equol, O-desmethylangolensin, and dihydrodaidzein, by comparing the retention times and the spectra of the urinary compounds with those of the synthesized reference standards. The mammalian lignans enterolactone and enterodiol were also identified. Derivatization of the isoflavones for GC-MS was examined by comparing two silylating reagents, N, O-bis-(trimethylsilyl)-trifluoroacetamide (BSTFA) and pyridine:hexamethyldisilazan:trimethylchlorosilane (QSM), both used for the derivatization of these compounds. The silylation experiments revealed significant differences in the compositions of the derivatization products. Some corrections were made concerning the earlier published data of dihydrogenistein and 6'-OH-O-dma. PMID- 10527519 TI - Assay for IkappaB kinases using an in vivo biotinylated IkappaB protein substrate. AB - IkappaB kinases (IKK)-1 and -2 are related kinases that are induced by stimuli such as TNF or IL-1 to phosphorylate serines 32 and 36 of IkappaBalpha, the regulatory subunit of the transcription factor NF-kappaB. A procedure for an IKK protein kinase assay is described that uses an in vivo biotinylated IkappaB protein substrate, [gamma-(33)P]ATP, and capture onto a streptavidin membrane. Residues 1-54 of the IkappaBalpha substrate were expressed as a fusion with glutathione S-transferase (GST) and a short (22 amino acid) biotinylation sequence that allowed modification during bacterial expression. Using the streptavidin capture assay the phosphorylation activities of recombinant IKK-1 and -2 were characterized. The assay provided a convenient way to compare IKK protein and peptide substrate preferences; biotinylated GST-IkappaBalpha(1-54) was more readily phosphorylated by both IKK-1 and IKK-2 compared to biotinylated myelin basic protein or a 20-mer biotinylated peptide containing serines 32 and 36 of IkappaBalpha. IKK-1 had 83-fold less activity than IKK-2, and the IKK-1+2 complex had approximately 2-fold more activity than IKK-2. IKK-1+2 and IKK-2 had similar K(m) values for ATP and GST-biotin-IkappaB(1-54) and were similarly inhibited by staurosporine and two of its analogues K252a and K252b, suggesting that most of the IkappaBalpha kinase activity in the IKK-1+2 complex may be attributed to IKK-2. Several features of the assay including the broad linear binding range of the streptavidin membranes for the protein substrate GST-biotin IkappaB(1-54) (1-4000 pmol of protein/cm(2)), the low background, and its capacity for both biotinylated peptides and proteins make it a useful tool for quantitating IKK activity. These factors and the ease of expressing in vivo biotinylated GST fusions will make this assay approach suitable for a wide variety of protein kinases. PMID- 10527520 TI - A pyridylamination method aimed at automatic oligosaccharide analysis of N-linked sugar chains. AB - The procedure for preparation of pyridylaminated sugar chains from glycoproteins was improved with a view to its eventual automation. Following on the coupling reaction improvement already reported [N. Kuraya and S. Hase (1992) J. Biochem. 112, 122-126], two further aspects were improved in this study. Instead of sodium bicarbonate-acetic anhydride, volatile reagents were adopted for the re-N acetylation of hexosamine residues after hydrazinolysis to give rapid removal of excess reagents. Subsequent to the pyridylamination reaction, excess reagents were removed by cation-exchange to isolate the pyridylaminated oligosaccharides in place of gel filtration. These alterations rendered a one-pot reaction possible and resulted in a large reduction in the amount of time needed compared with other methods so far reported. The procedure was successfully applied to the detection of sugar chains from Taka-amylase A and human erythrocyte membranes. PMID- 10527521 TI - Quantitative analysis of plasmid forms by agarose and capillary gel electrophoresis. AB - Plasmids may appear in different forms: circular with different degrees of coiling, partially cleaved or linear, and multimeric as concatamers or catenates. Capillary gel electrophoresis (CGE) of plasmid samples allows the determination of plasmid form distribution. Monomeric and dimeric plasmid DNA forms were separated by both CGE and agarose gel electrophoresis (AGE). The pattern of isoform bands from AGE was compared to the corresponding peak pattern from CGE, and differences in the relative mobility of the plasmid forms between the two methods were found. The comparison of AGE and CGE allows the assignment of AGE bands to CGE peaks. Additionally, the different isoforms can now be quantified by CGE. Routine plasmid form analysis by CGE may be automated, allowing easy, fast, and highly reliable quantification. CGE also offers high resolution and the amount of DNA required is very low. Therefore this method is very useful for the analysis of therapeutics based on plasmid DNA during their production, isolation, and formulation. PMID- 10527522 TI - A nonradioisotope biomedical assay for intact oligonucleotide and its chain shortened metabolites used for determination of exposure and elimination half life of antisense drugs in tissue. AB - Rigorous extraction methods coupled with capillary gel electrophoresis (CGE) provide a basis for a nonradiolabel assay for quantitation of intact antisense drug and its numerous chain-shortened metabolites. As part of the validation of the CGE method, we compared the quantitation of unlabeled ISIS 3521 (ISI 641A) and its chain-shortened metabolites with total radioactivity of [(35)S]-ISIS 3521. ISIS 3521 was labeled on the fifth nucleotide linkage from the 5'-end with (35)S by well-established methods. Multiple tissues collected from rats after administration of [(35)S]-ISIS 3521 were assayed by both radiolabel (liquid scintillation spectroscopy) and CGE methods. The CGE method provided accurate quantitation of the drug and its metabolites in kidney cortex and liver tissues. The correlation between methods for multiple tissues over time was excellent with 88.5% of the measurements being statistically equivalent. These data suggest that CGE is an accurate means of quantitating oligonucleotide in tissue and that it compares favorably with traditional radiochemical techniques. Clearance half lives for total measurable oligonucleotides were equivalent to clearance of total radioactivity in both liver and kidney with the longest clearance half-life associated with the kidney. PMID- 10527523 TI - A direct method for the correction of pressure-induced scrambling of polarized fluorescence intensities. AB - A simple and direct method for the simultaneous correction of steady-state polarized fluorescence intensities, depolarized (or scrambled) by the effects of applied hydrostatic pressure, is described. In the method discussed here, it is not necessary to first determine the scrambling factors from a separate experiment with a dye immobilized in a rigid medium. Rather correction for depolarizing effects of the high-pressure spectroscopy cell windows is achieved by direct recalculation of the measured polarized data obtained for the sample of interest at the time of data collection. This method of correction is tested for common fluorescent dyes 1, 6-diphenyl-1,3,5-hexatriene (DPH) and 9,10 diphenylanthracene in glycerol where their rotational behavior is well understood. In addition, the pressure-induced "melt" profile for the more complicated biologically relevant system of DPH imbedded within dipalmitoylphosphatidylcholine small unilamellar vesicles has been reexamined. While the method discussed here is used for the correction of steady-state polarized data, it may be easily adapted for use in time-resolved polarized fluorescence measurements. Advantages and limitations of the new correction method are discussed. PMID- 10527525 TI - Alkaline density gradient floatation of membranes: polypeptide composition of the mammalian peroxisomal membrane. AB - A method for purification of the peroxisomal membrane from rat liver is described. The procedure consists of floating the (contaminated) peroxisomal membranes through an alkaline sucrose density gradient. A good resolution between the peroxisomal membrane and other membrane systems is achieved. Using these floated peroxisomal membranes we have determined that only 7.8 +/- 0.9% of the total peroxisomal protein is alkali resistant. The polypeptide composition of these highly pure peroxisomal membranes was analyzed by SDS-PAGE. Bands corresponding to polypeptides with apparent molecular masses of 15, 18, 22, 24, 26, 29, 35, 36, 38, 40, 52, 55, 70, 74-77, and 88 kDa are detected upon Coomassie blue staining of polyacrylamide gels. The identity of several of these polypeptides was determined by N-terminal sequencing and Western blotting analysis. PMID- 10527524 TI - A fluorescence-based high-performance liquid chromatographic assay to determine acid ceramidase activity. AB - Acid ceramidase (N-acylsphingosine amidohydrolase) is the lysosomal enzyme required to hydrolyze the N-acyl linkage between the fatty acid and sphingosine moieties in ceramide. A deficiency of acid ceramidase activity results in the lipid storage disorder, Farber disease. This study reports a new assay method to detect acid ceramidase activity in vitro using Bodipy or lissamine rhodamine conjugated ceramide (C12 ceramide; dodecanoylsphingosine). Using mouse kidney extracts as the source of acid ceramidase activity, this new method was compared with an assay using radioactive C12 ceramide (N-[(14)C]-dodecanoylsphingosine) as a substrate. The Bodipy C12 ceramide substrate provided data very similar to those of the radioactive substrate, but under the experimental conditions tested, it was significantly more sensitive. Using Bodipy C12 ceramide, femtomole quantities of the product, Bodipy dodecanoic acid, could be detected, providing an accurate measure of acid ceramidase activity as low as 0.1 pmol/mg protein/h. Acid ceramidase activities in skin fibroblasts and EBV-transformed lymphoblasts from Farber disease patients were around 7.8 and 10% of those in normal cells, respectively, confirming the specificity of this new assay method. Based on these results, we suggest that this fluorescence-based, high-performance liquid chromatographic technique is a reliable, rapid, and highly sensitive method to determine acid ceramidase activity, and that it could be useful wherever the in vitro detection of acid ceramidase activity is of importance. PMID- 10527526 TI - Microscale purification of proteins exhibiting anomalous electrophoretic migration: application to the analysis of GAP-43 phosphorylation. AB - Quite often, in vivo analysis of posttranslational protein modifications is complicated by the lack of specific antibodies or unsatisfactory immunoprecipitation efficiency. Here, we present an alternative method to immunoprecipitation that takes advantage of the anomalous electrophoretic behavior exhibited by GAP-43. This method can be applied to other proteins which show similar characteristics, such as myristoylated alanine-rich C kinase, NAP 22, and Neurogranin, among others. All these proteins display relative mobility values that depend on the concentration of polyacrylamide used in the resolving gel. Cell extracts or tissue homogenates are first separated by SDS-PAGE in 15% polyacrylamide gels, and the bands containing GAP-43 are identified, excised from the gel, and rerun on a second SDS-PAGE in 7.5% polyacrylamide/6 M urea gels. To quickly identify the position of GAP-43 in the first gel, a small amount of fluorescein-labeled recombinant GAP-43 was added to the initial extracts. The method, applied to the analysis of GAP-43 phosphorylation in rat hippocampal slices, can be typically completed in less than 4 h. The excellent yields of purification obtained contributed to a greater accuracy and increased reliability of the radioactivity measurements. It also allowed further processing of the samples, including the analysis of the different phosphorylation sites by proteolytic digestion and peptide mapping. PMID- 10527527 TI - Quantitation of adenovirus DNA and virus particles with the PicoGreen fluorescent Dye. AB - A microplate assay for the rapid quantitation of adenovirus DNA has been developed using the fluorescent dye PicoGreen, which selectively binds double stranded DNA. The method was first applied to extracted adenoviral DNA and then extended to samples of intact, purified adenovirus after lysis of the viral capsid with the ionic detergent SDS. Utilizing the stoichiometric relationship between adenovirus DNA and intact particles, a physical particle count of intact virus is then derived for the sample. This PicoGreen-based assay has excellent reproducibility, linearity, and sensitivity. In its present form, this assay has a limit of quantitation of 10.3 ng/ml viral DNA, predicted to correspond to 2.6 x 10(8) virus particles/ml. This procedure was compared to a widely utilized spectroscopic method, in which samples are lysed with SDS and absorbance is read at 260 nm, and found to be 10- to 20-fold more sensitive. The dye binding assay also uses considerably less sample volume (<20%) than that needed for the spectroscopic method. Particle count values generated by the PicoGreen procedure are consistently lower (typically 1.5- to 2-fold) than this spectroscopic method. The applications and limitations of this method in the analysis of adenovirus samples are discussed. PMID- 10527529 TI - Adolescent tobacco use PMID- 10527528 TI - Purification procedure and monoclonal antibodies: two instruments for research on vertebrate porins. AB - On Western blots of skeletal muscle preparations of different vertebrate classes, four monoclonal anti-human type 1 porin antibodies recognize one single band of either 30.5 or 31 kDa, respectively. To confirm that it is eukaryotic porin which is labeled by the antibodies, we used a purification procedure developed for human type 1 porin for porins from skeletal muscle of shark, frog, and turkey. Applied to different mammalian species and tissues, this procedure exclusively provides type 1 porin. However, applied to shark skeletal muscle, it provides two porin isotypes in nearly equal amounts. In the case of frog skeletal muscle, the procedure provides mainly type 2 porin and a lower amount of type 1 porin. Applied to turkey skeletal muscle, the method provides exclusively type 2 porin. As demonstrated by two-dimensional Western blots, both shark and frog porin isotypes and the turkey type 2 porin are recognized by our antibodies. Furthermore, we elucidated the entire amino acid sequence of frog type 2 porin. PMID- 10527530 TI - The effectiveness of policy and health education strategies for reducing adolescent smoking: a review of the evidence. AB - This paper identifies the most effective measures to prevent smoking among adolescents. A review was made of the international literature concerning a ban on tobacco advertising, restrictions on sales to youth, product regulation, price increase of cigarettes and educational strategies. It is concluded that isolated measures produce little effect. Most effect may be expected from a combination of a complete ban on tobacco advertising, increasing prices, restricting tobacco product sales to tobacconists, mass media education aimed at youth and intensifying school education. A less effective measure is limiting the age at which adolescents are allowed to buy cigarettes. PMID- 10527531 TI - Adolescents and smoking: evidence from France and Spain. AB - Until recently detailed evidence on smoking and its reduction was found mainly in the U.S.A. and the U.K. Now there is active research in a much wider range of countries which increases the potential for international comparison and international learning. In this paper we focus on the recent evidence now available from France and Spain for the smoking behaviour of adolescents and young people. PMID- 10527532 TI - Teenage smoking in China. AB - China's production and consumption of cigarettes have ranked first in the world. One of every three cigarettes manufactured in the world is consumed in China. Three of every five Chinese smokers begin smoking at the age of 15-20 years. Teenage smoking is increasingly becoming a problem in modern China. At least 50 million of the children now living in China will be killed by smoking. Therefore China's top priority in control of smoking is to educate the youth against smoking so as to prevent them from starting and reduce the overall number of new smokers. Adults smoke; children follow. Thus a major feature of China's smoking control efforts has been the mobilization of primary school children to advise their parents to stop smoking. The goal of the Chinese Association on Smoking and Health is to achieve a male (age 15+) smoking rate below 58% and a female (age 15+) smoking rate below 5% by the year 2000. Although the number of smoke-free schools is on the increase and many more teenagers are quitting, China still has a long way to go. PMID- 10527533 TI - Cigarette smoking among schoolboys in Beijing, China. AB - The current study was designed to assess the recent rate of cigarette smoking and to examine the association of cigarette smoking with individual factors and problem behaviors using data collected in December 1997 from 323 middle school students (43% females) in Beijing, China. About 15% of the study sample (23% males and 5% females) reported having ever smoked. The data in the current study confirmed findings from our earlier study (Li et al., 1996, Substance Use and Misuse, 31, 545-563) that the prevalence of cigarette smoking among Chinese adolescents was higher among males, increased with advancing age and was associated with participation in other problem behaviors and with poorer self perceived academic performance. The data underscore the need for smoking prevention programs targeting Chinese children and adolescents. PMID- 10527534 TI - Knowledge, attitudes and preventive efforts of Malaysian medical students regarding exposure to environmental tobacco and cigarette smoking. AB - A longitudinal study was conducted to determine changes in knowledge, attitudes and preventive efforts of Malaysian medical students concerning cigarette smoking and environmental exposure to tobacco smoke from their first pre-clinical year in medical school until their final clinical year. There were significant improvements in knowledge about cigarette smoking and in knowledge, attitudes and efforts concerning environmental exposure to tobacco smoke. Overall attitudes concerning cigarette smoking did not change over this period. The same pattern was found for male non-smokers. Women improved on all five scales; male smokers improved on none over the 3-year period. Male non-smokers had better scores on these scales than male smokers in both beginning and ending years. Women excelled in comparison to male non-smokers on smoking attitudes in the pre-clinical year and on all scales except preventive efforts in the final clinical year.Although medical students experienced no changes in the amount of pressures not to smoke from family and friends, there was a significant increase in the amount of prohibition on smoking from their teachers. Male non-smokers alone accounted for this increase. Women experienced more pressure than men not to smoke from their teachers in both years, but the male smokers and non-smokers did not differ in teacher pressure for either year. PMID- 10527535 TI - Smoking behaviour: predisposition or adaptation? AB - The analysis makes use of a panel survey of 11-15-year olds containing their own responses to questions on smoking, smoking behaviour, and psychological well being, along with information obtained from parents on household composition, income and parental education. The panel covers four waves, enabling comparison of smoking behaviour, attitudes and psychological states at the start and end of the period. The data show that a significant part of adolescent smoking can be considered "experimental". It does not necessarily result in subsequent adoption. Changes in smoking behaviour are then related to changes in beliefs about the dangers of smoking which indicate that while the latter inhibit smoking they are also pliable when behaviour changes for other reasons. Finally, logistic regression is used to help explain change in smoking behaviour. The outcomes suggest that while young people, in terms of the impact of family background, can be seen as predisposed to smoke, some of the key explanatory factors in take-up are temporary, psychological states. This confirms that adolescent smoking is in large measure an adaptive response to immediate concerns and feelings. PMID- 10527536 TI - Starting smoking: girls' explanations of the influence of peers. AB - A questionnaire survey of cigarette smoking completed in six secondary schools by 4773 pupils aged 11-16 years included five items concerning the circumstances of smoking initiation: (1) age at first cigarette; (2) source of supply; (3) location of first smoke; (4) persons present; and (5) perceived degree of coercion. Although there was a highly significant gender difference on the item concerning age at first cigarette, with the average for boys lower than that for girls, the remaining items showed no gender effects. In order to explore further the reasons for the observed higher prevalence of cigarette smoking among teenage girls compared to boys, 33 focus groups were undertaken with girls in Years 7 and 9. These qualitative data provide evidence of the social representations of different groups and of smoking and non-smoking identities. Any understanding of smoking among girls needs to take account the dynamics of girls' membership in groups of never, experimental and regular smokers in determining subsequent smoking behaviour. PMID- 10527538 TI - Smoking behaviour in youth: the problem of low self-esteem? AB - The study considers the relationship between self-esteem and smoking in youth. Research has emphasized the need for ecological perspectives on health behaviours, for example, the context and meaning of cigarette smoking in young people's lives. Recent Scottish research, utilizing a range of methodologies, has examined the peer group context and smoking. The convergence of findings is striking. It would appear that different social groupings exist within the peer context, where these are tied to peer status and associated with distinctive lifestyle practices, and dispositions, including smoking. In the case of self esteem and smoking the results from conventional, survey-based research have often been inconclusive, the suggestion being that global measures of self-esteem are insufficient, since feelings of self-esteem are domain or context specific. However, the present study analyses survey data from two Scottish samples of 13 14-year-olds, conducted some 10 years apart, one national (n=2100, 1987) and the other rural (n=800, 1996) to show that even with the bluntest of research instruments, i.e. self-report questionnaire survey data and general measures, it is possible to elaborate on the relationship between self-esteem and cigarette smoking in youth. PMID- 10527537 TI - Why may teenage girls persist in smoking? AB - Teenage girls often smoke cigarettes, recognizing that it protects them from the impulse to binge eat with its feared weight-gain consequences. Evidence is marshalled from our studies of a female eating-disordered population, teenage females (London, U.K. and Ottawa, Canada) and middle-aged women (London and rural England) in the general population. Teenage female data analysis reveals links between smoking and body-weight/shape concerns. Those who smoked were likely to be moderately overweight. Smoking was also related at all ages to being postmenarchal. Sensitivity to shape is largely and qualitatively prompted by the development of body fat in puberty. Smoking by the London schoolgirls in particular also independently revealed an association with greater weight loss since puberty. Smoking was powerfully linked with vomiting undertaken as another defence against weight gain and may also be further reinforced as a behaviour by it. The eating-disordered population showed these latter associations most strikingly. Since smoking amongst older women is associated with below average body-weight it may indeed be effective in curbing weight gain and therefore promoting desired weight loss. Our studies provide little evidence of association between smoking and generalized or social anxiety. We propose that preventative psychological approaches to teenage female smoking should include attention to these matters. PMID- 10527539 TI - Continuities and changes: teenage smoking and occupational transition. AB - Rates of cigarette smoking among young people in the U.K. remain high and may be increasing. However, few studies have explored smoking behaviour during the mid- to late teens. This paper reports the third wave of a longitudinal study that followed 106 15-year-olds from their last compulsory year at school for 22 months. Using a mixture of qualitative and quantitative methods, the study shows that this is a period of considerable flux in smoking behaviour. Becoming a regular smoker is not a straightforward progressive process. The role of friendship groups and social context is highlighted. Smoking prevention programmes should be developed to meet the needs of young people in this transitional period. PMID- 10527541 TI - Reviewers PMID- 10527540 TI - Saying "no" to cigarettes: a reappraisal of adolescent refusal skills. AB - The principles of social inoculation developed in the late 1940s and later presented as a theory combined with Bandura's self-efficacy construct in the 1970s led to a series of smoking-prevention programmes for young people based on refusal skills. The present study examines refusal skills developed by young people who have not been taught such a programme. The survey was carried out by self-completed questionnaires administered to whole classes of 11-15-year-olds in two secondary schools in northern England. Harter's Self-Perception Profile for Children was used. Responses from 743 (365 boys and 378 girls) were analysed. Smoking prevalence reflected the national prevalence pattern for the age group at the time of the survey. Girls were at greater risk than boys of being repeatedly offered a cigarette and more likely than boys to accept it after more than two offers. Girls with high self-perception scores for all domains except social competence were at lowest risk of being offered a cigarette. For boys this only applied in the context of behavioural conduct. However, the factor most strongly related to multiple offers of cigarettes was having a best friend who smoked. Never smokers were most likely to have simply said "No, thank you" to proffered cigarettes but most had used several responses, boys generally using more refusal mechanisms than girls did. Cigarette refusal among young people is a complex process and programmes must be variable in order to meet specific circumstances, such as refusing a cigarette from one's best friend. PMID- 10527542 TI - Terminal differentiation and left-handed Z-DNA: a review. AB - Nucleic acids control the expression of genes, and different conformations of DNA structure may regulate cell death. Left-handed Z-DNA, which is speculated to function as a transcriptional enhancer, may be directly influenced by the destructive effects of terminal differentiation. The nicking-denaturation of double-stranded Z-DNA could possibly initiate and enhance terminal differentiation within specific tissues. PMID- 10527543 TI - A Na(+)-dependent system A and ASC-independent amino acid transport system stimulated by glucagon in rat hepatocytes. AB - The effects of glucagon on amino acid transport in rat hepatocytes are not fully understood. We examined the effect of this hormone on alanine, serine and cysteine preferring system (system ASC)-mediated amino acid transport in rat hepatocyte monolayers using 2-aminoisobutyric acid (AIB) and l -cysteine. Glucagon induced a time and protein synthesis-dependent stimulation of Na(+) dependent alanine preferring system (system A)-independent AIB transport. The glucagon-induced increase in transport activity was not modified by substrate starvation and not related to changes in the intracellular pool of amino acids. Glucagon did not modify system ASC activity measured by l -cysteine. Therefore the transport activity of AIB independent of system A stimulated by glucagon cannot be attributed to system ASC. This suggests a Na(+)-dependent transport system in rat hepatocytes not identified until now. PMID- 10527544 TI - Effects of cAMP on ERK mitogen-activated protein kinase activity in hepatocytes do not parallel the bidirectional regulation of DNA synthesis. AB - Previous studies have indicated that cAMP has bidirectional effects on epidermal growth factor (EGF)-induced DNA synthesis in cultured hepatocytes, acting to stimulate soon after plating (early G(1)) and to inhibit at later stages (nearer the G(1)/S transition). In this study we examined the role of the extracellular signal-regulated kinase (ERK) subgroup (p42/p44) of the mitogen activated protein (MAP) kinases both at growth-stimulatory and growth-inhibitory conditions. When added at low concentrations early during culturing, glucagon and 8 chlorophenylthio-cAMP (8-CPT-cAMP) did not increase MAP kinase activity, but enhanced the subsequent DNA synthesis. However, when administered at 24 h, glucagon and 8-CPT-cAMP decreased basal and EGF-induced MAP kinase activity and also inhibited EGF-induced DNA synthesis. Thus, although MAP kinase might play a role in the growth-inhibitory effect, it does not seem to be involved in growth promoting regulation by cAMP in hepatocytes. PMID- 10527545 TI - Differential alterations in the distribution of three phosphatase enzymes during the plasma membrane transformation of uterine epithelial cells in the rat. AB - Ultrastructural and light microscopic cytochemical methods were used to study the distribution and changes in distribution of three phosphatase enzymes: 5' nucleotidase (5N); thiamine pyrophosphatase (TPP); and adenosine triphosphatase (ATP) in the rat endometrium during early pregnancy up to the time of blastocyst attachment. The authors were particularly interested in changes in the apical plasma membrane and reaction product for all three enzymes was clearly localized along this membrane especially on day 1 of pregnancy. However, the three enzymes showed markedly different patterns of organization of reaction product at later times during early pregnancy. 5N, while showing a continuous lining along the microvilli on day 1 was virtually undetectable by day 6. TPP was also strongly present apically on day 1, but reaction product was not always found as a continuous lining. Again, by day 6, there was no presence of this enzyme along the apical surface. ATP differed from the other two in that it produced a strong, and relatively unchanged reaction product along the apical plasma membrane from day 1 through to day 6 of pregnancy. The changes in distribution of these enzymes was particularly obvious at the electron microscopic level and we consider their contribution to the process of 'plasma membrane transformation' of early pregnancy. PMID- 10527546 TI - Osteogenesis in vitro in rat tibia-derived osteoblasts is promoted by the homeopathic preparation, FMS*Calciumfluor. AB - We studied the effects of in vitro treatment of differentiating osteogenic cells with FMS*Calciumfluor, to determine whether it caused changes in proliferative or differentiation potential of osteoblasts. FMS*Calciumfluor was developed for the therapy of post-menopausal and age-related osteoporosis on the basis of the principles of resonance homeopathy and VTR Vega test. Its daily prescribed therapeutical usage is about 30,000-fold less in fluoride concentration than that recommended for NaF associated with calcium monophosphate. Rat tibial osteoblast (ROB) primary cultures represent populations of early osteoblasts and their derivative cultures of more than 60 cumulative population doubling (CPD) represent more mature osteogenic cells. Both these populations were shown to undergo in vitro differentiation, as monitored by the sequential expression of markers that define the stages of the osteogenic progression. Here we report that continual treatment of ROB during osteogenesis with FMS*Calciumfluor modulated the expression of critical osteogenic markers: alkaline phosphatase (AP), an indicator of osteoblast maturation, and(45)Ca incorporation into the matrix and nodule formation, events of the last phase of osteogenesis and a measure of osteoid mineralization. Treatment did not affect proliferation, or expression and activation of metalloproteinases (MMP). AP activity and levels of AP mRNA were increased by treatment with FMS*Calciumfluor; the incorporation of radiolabelled Ca into the matrix was also increased and the formation of nodules occurred in a shorter time and with higher frequency than in untreated control cultures. The effects of FMS*Calciumfluor were concentration dependent and specific for its modalities of preparation, and were observed at a concentration about three orders of magnitude lower than similar effects reported in the literature by treatment of osteoblast cultures in vitro with NaF. PMID- 10527547 TI - Tissue transglutaminase expression in quail epiphyseal chondrocytes. AB - Tissue transglutaminase (tTGase) is a GTP-binding Ca(2+)-dependent enzyme which catalyses the post-translational modification via epsilon(gamma-glutamyl)lysine bridges. The physiological role of tTGase is not fully understood. It has been shown that in cartilage the expression of tTGase correlates with terminal differentiation of chondrocytes. Recent evidence suggests that the GTP-binding activity of tTGase may play a role in the control of cell cycle progression thus explaining some of the suggested roles for the enzyme.tTGase activity is present in primary cultures of epiphyseal chondrocytes and increases transiently upon retinoic acid (RA) treatment. Increase in enzyme activity occurs upon RA addition and is accompanied by a parallel increase in protein and mRNA levels. Stimulation of tTGase expression by RA correlates with suppression of cell growth and occurs independently of cell adhesion and cell differentiation.tTGase expression is not observed in MC2, a permanent chondrocyte cell line derived from retrovirus infected chondrocytes. RA treatment fails to activate tTGase expression in MC2 cells and to completely suppress cell proliferation. Our findings lend support to the idea that tTGase might play a role in non-dividing cultured chondrocytes. PMID- 10527548 TI - Lovastatin induces mitotic abnormalities in various cell lines. AB - We examined the effects of lovastatin, a common anti-atherosclerotic drug and a blocker of the cell cycle, on the process of mitosis. It is known that lovastatin induces an arrest or a retardation of the cell cycle in many cell types not only at the G(1)phase, but also at the G(2)/M transition. After 24-48 h incubation of epithelial PtK(2), T24, HeLa cells and fibroblastic L929 cells in the presence of 1. 0-60.0 microm lovastatin, diverse mitotic perturbations have been observed. The most noteworthy phenomena recorded were prometaphase retardation and chromosome lagging during metaphase and anaphase. After the recovery in lovastatin-free media, the cells continued mitosis without any disturbances. Mevalonic acid prevented the effects of lovastatin. We conclude that the effects were specific for lovastatin-induced inhibition of mevalonic acid synthesis. Immunofluorescence studies with anticentromeric antibodies suggested that one of the possible causes of the lovastatin-induced mitotic disorder could be an interference with the development and function of the centromeres. PMID- 10527549 TI - Protein kinase C affects reformation of endothelial junctions in xenopus XTH-2 cells. AB - Endothelial cells can reversibly be forced to suppress the formation of endothelial junctions (EJ) by cultivation in a low calcium medium. The authors localized vinculin and cadherin as marker proteins of EJ and actin as a cytoskeletal component by fluorescence microscopy, and used this cell model to study the reformation of endothelial junctions under conditions of activation and inhibition of protein kinase C (PKC). Inhibition of PKC by H-7 leads to an acceleration of EJ reformation, while constitutive activation by TPA inhibits the reformation process. PMID- 10527550 TI - Protein domains involved in nuclear transport of Fos. AB - The protein encoded by the proto-oncogene c-fos is constitutively nuclear in most cell types analyzed. It has a predicted molecular weight of about 55 kDa. Proteins with a molecular weight above 40 kDa cannot enter the nucleus passively. Our interest was to study which regions in the protein are involved in the nuclear transport. We prepared a series of deletions and point mutations of the protein and cloned the mutated genes into a eukaryotic expression vector. Cos-1 cells were used to express the mutants transiently. Using indirect immunofluorescence we studied the subcellular localization, analyzing the percentage of cells containing the protein in the nucleus, the cytoplasm, or both locations. Our studies showed that the Fos protein contains several regions which can act independently to translocate the protein into the nucleus. PMID- 10527551 TI - Short communicationa simple way to obtain sufficient amounts of arterial endothelial cells from human umbilical cords. PMID- 10527552 TI - The development of mathematical cognition: arithmetic. PMID- 10527553 TI - The roles of estimation and the commutativity principle in the development of third graders' mental multiplication. AB - In this training experiment, pretesting identified 36 third graders (mean age = 8 years 5 months, SD = 4 months) with negligible mastery of multiplication combinations involving factors from 3 to 9. Participants were randomly assigned to 2 groups, which practiced different subsets of combinations, and were then retested. The results were inconsistent with R. S. Siegler's (1988) proposal that item-specific computational practice is necessary to promote changes in error patterns and combination mastery. The results were consistent with the hypotheses that children devise increasingly flexible and accurate estimation strategies and use relational knowledge such as the commutativity principle to master combinations. Because even fast mental-arithmetic errors and correct responses may be due to strategies other than retrieval, researchers need to craft ways of distinguishing between retrieval and nonretrieval strategies. PMID- 10527554 TI - Children's understanding of the relation between addition and subtraction: inversion, identity, and decomposition. AB - In order to understand addition and subtraction fully, children have to know about the relation between these two operations. We looked at this knowledge in two studies. In one we asked whether 5- and 6-year-old children understand that addition and subtraction cancel each other out and whether this understanding is based on the identity of the addend and subtrahend or on their quantity. We showed that children at this age use the inversion principle even when the addend and subtrahend are the same in quantity but involve different material. In our second study we showed that 6- to 8-year-old children also use the inversion in combination with decomposition to solve a + b - (b + 1) problems. In both studies, factor analyses suggested that the children were using different strategies in the control problems, which require computation, than in the inversion problems, which do not. We conclude that young children understand the relations between addition and subtraction and that this understanding may not be based on their computational skills. PMID- 10527556 TI - Working memory impairments in children with specific arithmetic learning difficulties. AB - Working memory impairments in children with difficulties in arithmetic have previously been investigated using questionable selection techniques and control groups, leading to problems concluding where deficits may occur. The present study attempted to overcome these criticisms by assessing 9-year-old children with difficulties specific to arithmetic, as indicated by normal reading, and comparing them with both age-matched and ability-matched controls. A battery of 10 tasks was used to assess different aspects of working memory, including subtypes of executive function. Relative to age-matched controls, children with poor arithmetic had normal phonological working memory but were impaired on spatial working memory and some aspects of executive processing. Compared to ability-matched controls, they were impaired only on one task designed to assess executive processes for holding and manipulating information in long-term memory. These deficits in executive and spatial aspects of working memory seem likely to be important factors in poor arithmetical attainment. PMID- 10527555 TI - Numerical and arithmetical cognition: patterns of functions and deficits in children at risk for a mathematical disability. AB - Based on performance on standard achievement tests, first-grade children (mean age = 82 months) with IQ scores in the low-average to high-average range were classified as at risk for a learning disability (LD) in mathematics, reading, or both. These at-risk children (n = 55) and a control group of academically normal peers (n = 35) were administered experimental tasks that assessed number comprehension and production skills, counting knowledge, arithmetic skills, working memory, and ease of retrieving information from long-term memory. Different patterns of intact cognitive functions and deficits were found for children in the different at-risk groups. As a set, performance on the experimental tasks accounted for roughly 50% and 10% of the group differences in mathematics and reading achievement, respectively, above and beyond the influence of IQ. Performance on the experimental tasks thus provides insights into the cognitive deficits underlying different forms of LD, as well as into the sources of individual differences in academic achievement. PMID- 10527557 TI - Simple mental addition in children with and without mild mental retardation. AB - The speeded performance on simple mental addition problems of 6- and 7-year-old children with and without mild mental retardation is modeled from a person perspective and an item perspective. On the person side, it was found that a single cognitive dimension spanned the performance differences between the two ability groups. However, a discontinuity, or "jump," was observed in the performance of the normal ability group on the easier items. On the item side, the addition problems were almost perfectly ordered in difficulty according to their problem size. Differences in difficulty were explained by factors related to the difficulty of executing nonretrieval strategies. All findings were interpreted within the framework of Siegler's (e.g., R. S. Siegler & C. Shipley, 1995) model of children's strategy choices in arithmetic. Models from item response theory were used to test the hypotheses. PMID- 10527558 TI - Murine T cell determination of pregnancy outcome. AB - At the fetomaternal interface, maternal effector cells come in intimate contact with fetal trophoblast cells which express paternal antigens. Failure of fetal trophoblast cells to activate maternal Th1 immune responses has been attributed in part to the absence of classical Class I and Class II major histocompatibilty complex (MHC) antigen expression and elaboration of factors which reduce TcR expression and shift any immune responses which may occur to Th2. Classical TcR alphabeta(+) T cells have not been found to be able to respond to trophoblasts. Recently, TcR gammadelta(+) T cells have been characterized in the low-abortion rate pregnant C57Bl/10 mouse decidua, and the Vgamma1(+) subset may be able to respond to trophoblasts in a non-MHC-dependent manner. Trophoblast-recognizing T cells with Vgamma1 receptors are also present in the decidua of CBA/J mice pregnant by DBA/2, an abortion-prone mating combination. To test the role of the Vgamma1 subset of decidual gammadelta T cells in abortion-prone pregnancies, we altered this subset by injecting monoclonal anti-Vgamma1.1 antibody on gestation day 5.5, 1 day after implantation. This reduced detectability of a Vgammadelta subset producing TNF-alpha and reduced the abortion rate. Anti-Vgamma2, which reacts with a similar proportion of decidual gammadelta T cells as anti Vgamma1.1, failed to prevent abortions. Vdelta6.3(+) cells are prominent at the fetomaternal interface, and anti-Vdelta6 antibody injected on day 5.5 prevented abortions. TGF-beta2(+) gammadelta cells first appear on day 8.5 of pregnancy; anti-Vgamma1.1 antibody injection on day 8.5 depleted these cells and boosted abortions; anti-Vdelta6.3 given on day 8.5 boosted abortions to the same level. These results suggest that two populations of Vgamma1.1(+)delta6.3(+) T cells may arise in the decidua: an early population that is Th1, abortogenic, and present during the time of implantation, and a Th2/3 cell subset that is present in the decidua later during pregnancy and which is pregnancy-protective. PMID- 10527559 TI - Cloning of a novel MHC-encoded serine peptidase highly expressed by cortical epithelial cells of the thymus. AB - Antigen presentation by cortical thymic epithelial cells (cTEC) during the positive selection of T cells has been shown to differ from that of other antigen presenting cells. In the case of MHC class II presentation, cathepsin L as opposed to cathepsin S is responsible at least in part for the degradation of invariant chain. Other proteases, however, must be involved. We have identified a putative serine protease that is specifically expressed in the thymus. Encoded within the class I major histocompatibility complex (MHC) region, this gene has sequence homology with lysosomal prolylcarboxypeptidase, suggesting that it is a serine protease. We have, therefore, designated this gene thymus-specific serine protease (TSSP). In situ hybridization and immunofluorescence staining reveal that TSSP is expressed exclusively by cortical thymic epithelial cells, with the strongest staining noted around vessels and the thymic capsule. The identification of TSSP further supports the theory that MHC class II antigen processing and presentation in the thymic cortex involves a proteolytic milieu that differs from that of other antigen-presenting cells. PMID- 10527560 TI - Cytotoxic properties of CD4(+) T-cell clones which lyse HLA class II negative autologous non-small-cell lung cancer cells. AB - In this report, a subset of CD4(+) cells which kills autologous HLA class I positive and HLA class II negative non-small-cell lung cancer (NSCLC) cells is described. Killing was performed both by direct cytolytic mechanisms and by apoptosis. The peculiar characteristic of these effectors was their capability of lysing only interferon (IFN)-gamma-treated NSCLC target cells, expressing high numbers of HLA class I molecules. Analysis at the clonal level confirmed that cytolytic capability was distributed clonotypically. The addition of anti-HLA class I monoclonal IgM abrogated the susceptibility to lysis. Notably, the CD4 mediated cytolytic effect on IFN-gamma-treated targets was incomplete. Thus cytofluorimetric DNA and HLA class I expression analyses of target cells were performed: "resistant" targets consisted of large, aneuploid cells expressing a low number of HLA class I molecules. These findings suggested a direct role of HLA class I expression in the recognition of autologous cancer cells mediated by cytolytic CD4(+) cells. PMID- 10527561 TI - The role of CD80 and CD86 in enhancing CD8(+) cell suppression of HIV replication. AB - CD8(+) cells activated in the presence of autologous macrophages (Mphi) have an increased ability to suppress HIV replication compared to the same cells stimulated in the absence of Mphi. Blocking the B7 molecules decreases the ability of Mphi to increase CD8(+) cell antiviral activity. In the present study CD8(+) cells exposed to purified forms of both the CD80 and the CD86 molecules during stimulation with anti-CD3 antibodies (Ab) had a greater ability to suppress HIV replication than CD8(+) cells exposed to anti-CD3 Ab alone. The addition of anti-CD86 blocking Ab, but not anti-CD80 blocking Ab, to Mphi decreased their ability to enhance CD8(+) cell suppression of HIV replication. Moreover, anti-CD86 Ab and not anti-CD80 Ab blocked the production of IL-2 by CD8(+) cells stimulated in the presence of Mphi. The incapacity of anti-CD80 Ab to block the enhanced antiviral activity and IL-2 production of CD8(+) cells stimulated in the presence of Mphi was not due to the inability of this Ab to function since anti-CD80 Ab are able to block proliferation of CD8(+) cells cultured in the presence of Mphi. Thus, while both B7 molecules can deliver a costimulatory signal sufficient to increase CD8(+) cell antiviral activity, CD86 appears to be the molecule that serves as the costimulatory molecule on Mphi to enhance CD8(+) cell suppression of HIV replication. The difference in use of CD86 over CD80 molecules on Mphi by CD8(+) cells mediating the antiviral suppressing activity most likely results from a higher number of Mphi expressing the CD86 molecule compared with the CD80 molecule. This information offers a possible therapeutic approach to increase CD8(+) cell anti-HIV response. PMID- 10527562 TI - Protein kinase C-alpha is essential for Ramos-BL B cell survival. AB - The Ramos-Burkitt lymphoma (BL) B cell line is driven into growth arrest and apoptosis by cross-linking surface immunoglobulin. We demonstrate that protein kinase C (PKC) activity is required for Ramos B cell proliferation and survival. A variety of PKC inhibitors trigger a significant decrease in [(3)H]thymidine incorporation with a concomitant increase in cell death. Antisense depletion of expression of the PKC-alpha isoform is sufficient to trigger cell death in the absence of any other signal, demonstrating a requirement for this isoform for survival of Ramos-BL B cells. Cross-linking surface immunoglobulin also leads to depletion of PKC-alpha levels, suggesting that this may be one mechanism by which this signals for cell death in Ramos-BL B cells. PMID- 10527563 TI - Induction of CD4(+) and CD8(+) Bordetella pertussis toxin subunit S1 specific T cells by immunization with synthetic peptides. AB - In this study two synthetic peptides from the Bordetella pertussis toxin subunit S1 were conjugated to human anti-idiotypic antibodies and used as an immunogen in cancer patients to induce immunity. The aims of the present report are to explain why no carrier or adjuvant effect of the conjugated pertussis peptides could be established regarding induction of responses against the anti-idiotype and to explore the type and quality of induced anti-pertussis immune responses. The lack of carrier and adjuvant effect of the peptides might be related to the fact that the anti-idiotypic antibodies by themselves include helper epitopes and that none of the patients had a detectable T cell response against any of the selected peptides before immunization, which might be a requirement for an adjuvant effect. However, three of four immunized patients mounted a humoral as well as cellular response against the pertussis peptides used. The induced T cell immunity was restricted to one of the two peptides in responding patients. Established T cell lines and MHC blocking studies indicated that the T cell epitopes of the two peptides had a different MHC restriction. The type of T cell response induced seemed to govern the humoral response. The only durable antibody response was accompanied by the presence of a CD4(+) T cell response against the same peptide. Immunization with an anti-idiotype conjugated to synthetic peptides might thus induce both a B and a T cell response against the peptides and the type of induced T cells (CD4 or CD8) governs the quality of the humoral response. Moreover, the possibility of boosting or inducing a response against the antigen from which the peptide sequences were deduced also seemed feasible. PMID- 10527564 TI - Maternal Th1- and Th2-type reactivity to placental antigens in normal human pregnancy and unexplained recurrent spontaneous abortions. AB - Spontaneous abortion is the most common complication of pregnancy, but the etiology of a significant proportion of abortions is still unknown. We have examined the production of Th1- and Th2-type cytokines by women with unexplained recurrent spontaneous abortion (RSA) since it appears that successful murine pregnancy occurs in a Th2-dominant situation and that Th1-type immunity is associated with pregnancy failure. We have compared maternal reactivity toward placental antigens in women with a history of successful pregnancy with that in women with a history of RSA. This was done by coculturing maternal peripheral blood mononuclear cells (PBMC) with autologous placental cells and also by stimulating maternal PBMC with antigens from a choriocarcinoma cell line of trophoblastic origin. We detected significantly greater levels of the Th2 cytokines IL-6 and IL-10 in normal pregnancy compared to unexplained RSA and significantly higher levels of the Th1 cytokine IFN-gamma in RSA compared to normal pregnancy. These results suggest that women with normal pregnancy have a higher Th2 bias, while women with a history of RSA evince a bias toward Th1-type reactivity. PMID- 10527566 TI - Adsorption of Nitrogen on Thermally Treated Peat Soils: The Role of Energetic and Geometric Heterogeneity. AB - We investigate adsorption isotherms of nitrogen at 80 K on several peat soils. In addition to natural soil samples we also study samples thermally treated at 50, 100, and 150 degrees C. The experimental adsorption isotherms are used to evaluate the surface fractal dimension and the energy distribution functions. Moreover, for some samples we have also determined the pore size distributions from mercury intrusion data. We compare the surface fractal dimensions evaluated from the mercury intrusion data and from adsorption isotherms and discuss how the thermal treatment changes the energetic heterogeneity of the samples. Copyright 1999 Academic Press. PMID- 10527565 TI - Apoptotic cells are generated at every division of in vitro cultured T cell lines. AB - Long- and short-term T cell lines form the backbone of many assays for T cell function and also represent important tools for use in human immunotherapy. Despite much study concerning the requirements for T cell activation and growth in culture there is relatively little information about the kinetics of proliferation and cell death in such cultures. Here we studied these parameters in a long-term CD8(+) T cell line using a tetrameric MHC reagent and the fluorescent dye CFSE. We observed proliferation of the T cells within 24 h of restimulation with antigen and IL-2 and the cells continued to divide once every 12 h on average. Interestingly, a proportion of cells entered apoptosis with each cell division, showing that a degree of programmed cell death occurred constantly in vitro, not merely at the end of the culture period when antigen or the necessary growth factors became limiting. This information should assist in the design of more efficient protocols for generating large numbers of specific T cells for clinical use. PMID- 10527567 TI - Excess of Solubilization and Curvature in Nonionic Microemulsions. AB - We measure separately the amount of solute dissolved in a surfactant monolayer and the average curvature of the relevant sample to establish a link between these two quantities. The model system chosen involves the common hydrophobic pesticide lindane (gamma-C(6)H(6)Cl(6)) in a nonionic surfactant solution of the ethylene oxide type. Excess solubilization, defined as the solubilization in the surfactant film by comparison with bulk oil, is quantified by the interfacial composition lambda (molar ratio solute/surfactant) within the interfacial film. A linear relationship between the amount of solute adsorbed on the film and the induced variation in curvature of the surfactant film is deduced from the phase diagram, dosage, and small-angle scattering experiments in the case of micellar, Winsor I, and several Winsor III domains at equilibrium in the same ternary system. We discuss the linear relationship obtained with constraints set by molecular packing. Copyright 1999 Academic Press. PMID- 10527568 TI - Synthesis of Porous Zirconia Spheres-Mechanism and Prospects for Multistep Processing. AB - We have previously reported the polymerization-induced colloid aggregation process (1, 2) to synthesize monodisperse, porous zirconia particles. One drawback is that it is difficult to simultaneously achieve a larger particle size and a higher zirconia yield. In this paper, we develop strategies to achieve these dual goals. We elucidate the interplay between the polymerization reaction and the various polymer-colloid interactions which result in the formation of the monodisperse particles. We explain the role of the polymer and the colloid in the process and suggest reasons for the limitations observed with a single-step process in achieving the dual goals of larger particle size and increased zirconia yield. Finally, we demonstrate a multistep process to overcome these limitations and achieve these dual goals. Copyright 1999 Academic Press. PMID- 10527569 TI - Enthalpy-Entropy Compensation Phenomenon Observed for Different Surfactants in Aqueous Solution. AB - Based on previously reported thermodynamic data such as changes of the Gibbs energy (DeltaG(m)( degrees )), the enthalpy (DeltaH(m)( degrees )), and the entropy (DeltaS(m)( degrees )) on micelle formation of more than 15 species of surfactants (including nonionic, anionic, and cationic surfactants), plots of DeltaH(m)( degrees ) vs DeltaS(m)( degrees ) (not of DeltaS(m)( degrees ) vs DeltaH(m)( degrees ), as is usually done) were made. For each surfactant, a linear relation having almost the same slope (1/307 K(-1)) within a small error (+/-2.3%) but a different intercept (varsigma) depending on the surfactant species was obtained, i.e., DeltaS(m)( degrees ) = (1/307)DeltaH(m)( degrees ) + varsigma, where 1/307 (K(-1)) means that the so-called compensation temperature (T(C)) is 307 K. Strictly speaking, T(C) ranges from 299 to 315 K, depending on the species. The intercept corresponds to the entropy change at a specific temperature giving DeltaH(m)( degrees ) = 0, at which the driving force of micelle formation comes only from the entropy term; this temperature is characteristic of the surfactant species. On the other hand, the compensation temperature has no significant meaning other than a mean temperature studied. Copyright 1999 Academic Press. PMID- 10527570 TI - Numerical Analysis of the Concentration Polarization in Colloidal Suspensions: Comparison with Theoretical Predictions. AB - The equation system based on the standard model for a charged spherical particle suspended in an electrolyte solution and under the action of a static applied field is numerically solved. Equilibrium and nonequilibrium properties-electric potential, ion concentrations, adsorption coefficients, dipolar coefficient, stored electrostatic and Gibbs free energies, conductivity increment, and permittivity increment-are deduced, discussed, and compared with theoretical predictions of the Shilov and Dukhin model. Theoretical predictions for the conductivity properly reproduce the numerical results for low values of the surface potential. But for highly charged particles important deviations appear even for large particle radius to Debye screening length ratios. Theoretical predictions for the permittivity are generally better than for the conductivity. This leads to the conclusion that in many situations the theoretical expression is sufficiently good for the interpretation of experimental results in terms of the standard model. But its range of applicability is determined not only by the particle radius to Debye screening length ratio but also by the surface potential. Copyright 1999 Academic Press. PMID- 10527571 TI - A Comparative Study of Surface Acid-Base Characteristics of Natural Illites from Different Origins. AB - The acid-base characteristics of naturally occurring illites, collected from different locations, were investigated by potentiometric titrations. The experimental data were interpreted using the constant capacitance surface complexation model. Considerable release of Al and Si from illite samples and subsequent complexation or precipitation of hydroxyl aluminosilicates generated during the acidimetric forward titration and the alkalimetric back titration, respectively, were observed. Therefore, the acidimetric supernatant, rather than the neutral one, was regarded as the system blank for each illite suspension to yield the surface site concentrations. In order to describe the acid-base chemistry of aqueous illite surfaces, two surface proton-reaction models, introducing the corresponding reactions between the dissolved aluminum species and silicic acid, as well as a surface Al-Si complex on homogeneous illite surface sites, were proposed as follows: The K(f2) constant in Model II was obtained by simulating the complex formation between the dissolved aluminum species and silicic acid that occurred in acidimetric supernatant when the hydroxide was added. Additionally, the following cation exchange reaction was also considered for a special case, where a large amount of K(+) is released during the corresponding acidimetric titration, in which a high concentration of protons are consumed. Optimization results indicated that both models could provide a good description of the titration behavior for all aqueous illite systems in this study. The intrinsic acidity constants for the different illites were similar in Model I, showing some generalities in their acid-base properties. Model I may be considered as a simplification of Model II, evident in the similarities between the corresponding constants. In addition, the formation constant for surface Al-Si species (complexes or precipitates) is relatively stable in this study. Copyright 1999 Academic Press. PMID- 10527572 TI - Experimental Investigation on the Resonance of a Liquid Column in a Capillary Tube. AB - Using a visualization technique, we observed the resonance of a water column trapped in a vertically oriented capillary tube due to acoustic excitation. The analysis of the quasi-static response suggests that the upper nonvisible meniscus followed the imposed flow by means of a sliding contact line without changing its shape. We compared the experiments with a previously developed theoretical model that addresses dissipation by assuming an axially symmetric and incompressible flow field that is spatially constant along the tube axis. Whereas the model agrees well with the measured quasi-static response, the deviations in the dynamic response reveal shortcomings of the model due to the simplified treatment of the viscous dissipation. Copyright 1999 Academic Press. PMID- 10527573 TI - Analysis of Fine Bubble Attachment onto a Solid Surface within the Framework of Classical DLVO Theory. AB - Fine bubble attachment onto a solid surface in an impinging jet flow was analyzed within the framework of DLVO theory. The effects of hydrodynamic convection, van der Waals (VDW) interaction, electrostatic double-layer (EDL) interaction, and gravitational force on bubble attachment rate (in terms of the Sherwood number) were examined in detail. The analyses showed that due to large Peclet number and gravity number for gas bubbles the behavior of the bubble attachment is significantly different from that of colloidal particle deposition in some aspects. Specifically, it was demonstrated that within a certain range of physicochemical conditions, gas bubbles can attach onto a solid surface despite the existence of repulsive VDW interaction force and the fact that the surfaces of both the bubble and the solid collector carry the same sign of electrostatic potentials. This is attributed to the role played by the short-range attractive asymmetric EDL interaction and the strong hydrodynamic and gravity forces, without any need for the so-called hydrophobic interaction force. In addition, it was also shown that the models derived for the impinging jet system can be used to evaluate transport of fine gas bubbles onto a large particle surface, suggesting that the information extracted from the impinging jet geometry can be applied to the analysis of flotation processes. Copyright 1999 Academic Press. PMID- 10527574 TI - Controlled Formation of Low-Volume Liquid Pillars between Plates with a Lattice of Wetting Patches by Use of a Second Immiscible Fluid. AB - We describe a method for forming an array of microdroplets between two plates, at least one of which is patterned with a lattice of wetting patches, using a second immiscible fluid to control droplet formation. The method may be useful for performing multiple, small-volume biochemical reactions in parallel. We analyze the forces responsible for droplet formation, describe results of a computer simulation using Surface Evolver, and derive an analytic criterion for droplet formation in terms of the contact angles of the droplet:second fluid interface on the wetting patches and surrounding surface, the diameter of the wetting patches, the distance between wetting patches, and the distance between the plates. Copyright 1999 Academic Press. PMID- 10527575 TI - Foam Destabilization by Mechanical and Ultrasonic Vibrations. AB - The need to destabilize foams and control their formation arises in many industrial applications. Vibrations are known to affect the structure and rheology of various soft solids but have not been exploited in destroying persistent foams. Being noninvasive, they offer elegant potential alternatives to chemical and mechanical foam breaking techniques. This paper reports an experimental study on the effectiveness of mechanical and ultrasonic vibrations in foam destruction. Mechanical vibration is effective at breaking static foams generated from non-Newtonian shear-thinning liquids, by enhancing liquid drainage and film breakage. Drainage is increased due to enhanced shear thinning that leads to a reduction in yield stress and shear viscosity of the nonNewtonian liquid. High intensity ultrasonic vibration is efficient at destabilizing static foams but is also effective at controlling dynamic foam heads and would be suitable for use in processes that require continuous defoaming. Destabilization of the foam films is attributed to possible squeezing mode surface wave phenomena. Copyright 1999 Academic Press. PMID- 10527576 TI - A Novel Fast Technique for Measuring Dynamic Surface and Interfacial Tension of Surfactant Solutions at Constant Interfacial Area. AB - A novel, fast formed drop technique is developed for measuring dynamic surface and interfacial tension of surfactant solutions. It employs the basis of the capillary pressure methods. A new way of formation of a fresh interface is the key point in the proposed technique. It is based on the fact that a jet of liquid flowing out of a capillary breaks very fast after a sudden stop of the flow, but a small drop with a fresh interface remains at the capillary tip. Once the drop is formed, its size (and thereby its area) remains constant, while the capillary pressure is measured with a pressure transducer. The dynamic surface (or interfacial) tension is calculated by means of the Laplace equation for capillarity. In this way, measurements under constant interfacial area in a time domain from about 50 ms up to several minutes or more can be made. The measurements with pure liquids give constant values of the dynamic surface and interfacial tension equal to the corresponding equilibrium values reported in literature. The dynamic surface tension of Triton X 100 and Triton X 405 solutions measured by the new technique are in a good agreement with those obtained by the static drop weight method. The applicability of the new technique for measuring the dynamic interfacial tension is demonstrated. Copyright 1999 Academic Press. PMID- 10527577 TI - Effect of Lauryl Alcohol on Morphology of Uniform Polystyrene-Poly(methyl methacrylate) Composite Microspheres Prepared by Porous Glass Membrane Emulsification Technique. AB - Fairly uniform poly(styrene)-poly(methyl methacrylate) (PST-PMMA) composite microspheres were prepared by employing an SPG (Shirasu Porous Glass) membrane emulsification technique. A mixture of PST, PMMA, and cosurfactant [lauryl alcohol (LOH)] dissolved in dichloromethane (DCM) was used as the dispersed phase, and an aqueous phase containing poly(vinyl alcohol) and sodium lauryl sulfate was used as the continuous phase. It is necessary to add LOH to obtain uniform particles with the SPG emulsification technique. The effects of the volume of LOH on the morphology of the final particles were investigated by varying the volume of LOH from 0 to 2 ml (per 1.2 g polymer). A three-component model was developed for different PMMA/PST ratios and LOH/polymer ratios, based on Sundberg's theory; and the calculation on morphology was carried out by using the three-component model. Agreement was obtained between experimental and calculated results. When 2 ml of LOH was added, it was found that LOH can engulf the polymer particles completely; a hemicore (HCP1P2P3) morphology, where PST and PMMA formed a hemisphere core inside a LOH shell, was observed when the PMMA/PST ratio was high, while core-shell-shell (CSP1P2P3) morphology, where PMMA formed a core and PST and LOH formed an inner shell and an outer shell, respectively, was observed when the PMMA/PST ratio was low. When the volume of LOH was below 1 ml (per 1.2 g polymer), however, LOH could not always engulf the inner polymer particles completely; cored hemisphere (CHSP3P2P1), core-shell-shell (CSP2P1P3), and hemicore (HCP1P2P3) morphologies were observed, depending on the PMMA/PST ratio and volume of LOH. Copyright 1999 Academic Press. PMID- 10527578 TI - Coalescence of Lipid Emulsions in Floating and Freeze-Thawing Processes: Examination of the Coalescence Transition State Theory. AB - Lipid emulsions, triacylglycerol droplets covered with single surface monolayers of phospholipid in aqueous medium, were prepared by high-pressure emulsification and successive ultracentrifugation. Egg yolk phosphatidylethanolamine did not stabilize triolein (TO) droplets in aqueous medium. Phosphatidylcholine (PC) of various long fatty acyl chains dispersed TO and tricaprylin (TC) well in aqueous medium as emulsion droplets of 110 nm. After ultracentrifugation, however, oil separation was observed in the floating creamy layer of TO/dioleoyl-PC emulsions. Dioleoyl-PC contains two bulky unsaturated fatty acyl chains, as well as oleoyl chains, and it has a lower spontaneous curvature than egg PC. Egg PC and dipalmitoyl-PC gave stably dispersed emulsion droplets of TO even after the freeze-thawing (F-T) procedure. On the other hand, TO/egg PC emulsion droplets containing cholesterol of 83 mol% PC and TC/egg PC emulsion droplets coalesced after the F-T procedure. Cholesterol and a medium chain triacylglycerol, TC, were distributed into the surface PC monolayers and occupied an appreciable fraction of the surface area. These neutral lipids have small polar groups and thus decrease the mean spontaneous curvature of surface lipids. The relationship between the droplet coalescence and the spontaneous curvature was discussed on the basis of the coalescence transition state theory recently developed by Kabalnov et al. in 1996. In addition, effects of maltose on the emulsion coalescence in the freeze state were briefly discussed. Copyright 1999 Academic Press. PMID- 10527579 TI - Correlation between Generated Shear Stress and Generated Permittivity for the Electrorheological Response of Colloidal Silica Suspensions. AB - Electrorheological response was experimentally studied by the use of silicone oil suspensions containing submicrometer-sized and supermicrometer-sized silica particles with different amounts of adsorbed water. The simultaneous measurements of dielectric permittivity and shear stress of the suspensions were carried out after the application of alternating current voltage under steady shear in ranges of shear rate (150-1400 s(-1)), electric field strength (0-4 kV/mm), its frequency (30-1000 Hz), and particle volume fraction (0.1-0.3). For the particles with small amounts of adsorbed water, steady shear stress was attained within several minutes after the application of electric field. The steady-state data for both the particles at each electric field strength showed that the shear stress generated by the application of electric field, Deltatau, varied correlatively with the generated permittivity, Deltaepsilon(r) (= epsilon(r) - epsilon(r,oil)), where epsilon(r) and epsilon(r,oil) are the permittivities of the suspension and the silicon oil, respectively. Under a wide variety of experimental conditions, the steady-state data for both the particle sizes could be correlated with a simple relationship, Deltatau ~ (Deltaepsilon(r)E)(2), where E is electric field strength. For the particles with large amounts of adsorbed water, steady state was not attained, and the evolutions of shear stress and permittivity of the suspensions were measured after the application of electric field. Remarkably, the transient values of Deltatau varied with (Deltaepsilon(r)E)(2) and fell along the same correlation line as the steady state data. Copyright 1999 Academic Press. PMID- 10527580 TI - Adsorption and Two-Dimensional Condensation of Thiabendazole at a Mercury/Solution Interface. AB - The adsorption behavior of the fungicide thiabendazole [2-(4'-thiazolyl) benzimidazole] was studied with out-of-phase ac voltammetry using a hanging mercury drop electrode in 0.3 M KCl solutions at various temperatures and thiabendazole concentrations. A state of "normal" adsorption and a condensed two dimensional layer at moderately charged and rather negatively charged surfaces, respectively, have been characterized. Copyright 1999 Academic Press. PMID- 10527581 TI - Monolayer Coverage-Sorbate Area Relationship for Mixtures of Sorbents. AB - The adsorption of molecules of different sizes, at monolayer coverages, was determined for binary mixtures of montmorillonite and hydroxy-Al montmorillonite in order to test the applicability of the general equation (NA(z) = k) relating the number of sorbate units at monolayer coverage (N) with their areas (A). The effect of the proportions of the components in the mixture on the value of the exponent z was also evaluated. The data obtained confirm the validity of the equation and indicate that the value of z for the mixtures assumes intermediate values between the two z values of the components, depending on their proportions. It is also demonstrated that the z exponent could have a value of unity (a value supposed to indicate surface uniformity) for certain mixtures of sorbents having z values different from unity, i.e., possessing nonuniform surfaces. Copyright 1999 Academic Press. PMID- 10527582 TI - Binding and Ion-Exchange Analysis in the Process of Adsorption of Anionic Polyelectrolytes on Barium Sulfate. AB - The adsorption of poly(acrylic acid) and poly(styrenesulfonic acid) (PSS) is studied on a barium sulfate powder. Comparison of the polyelectrolytes shows that the difference in binding strength corroborates the difference between the ligand strengths with barium ions. The surface excess dependence on pH correlates with the density of the ionized groups. The electrokinetic potential follows the surface coverage and the ionization ratio of the polymer up to the onset of the adsorption plateau, but continues to increase above that point. This peculiarity is explained by the release of barium ions from the adsorption layer into the solution. The phenomenon is attributed to the complexing power of unadsorbed molecules. Analysis of the displacement of small ions (Na(+), SO(2-)(4)) shows that adsorbed PSS releases sulfate ions from the surface and sodium counterions from the polymer. The displacement ratio for sulfate ions (SO(2-)(4)/PSS monomer units) remains constant over the adsorption isotherm, but that for sodium ions is constant only up to about two-thirds of the maximum coverage. From the data we deduced that about half of the monomer units of the adsorbed PSS molecules bind with surface barium ions. The other half form barium and sodium sulfonates whose ratio varies with the concentration of unadsorbed molecules. Copyright 1999 Academic Press. PMID- 10527583 TI - Effect of Sublytic Concentrations of Sodium Cholate on Phospholipase C Hydrolysis of Phospholipid Bilayers. AB - Phospholipase C activity has been assayed with phosphatidylcholine as substrate in the presence of sodium cholate at concentrations well below those producing lipid solubilization. With short-chain phosphatidylcholine, which exists in monomeric form in aqueous solution, cholate has little or no effect. However, when the substrate is egg phosphatidylcholine in the form of bilayers, small cholate concentrations (below 1 mM, corresponding to an effective surfactant:lipid ratio below 0.05) increase the maximum enzyme rates by about threefold, while decreasing drastically the latency periods of enzyme activity. Previous studies from this laboratory have associated the phospholipase enhancing activity of a variety of amphiphiles to their ability to facilitate the formation of inverted hexagonal phospholipid structures, yet sodium cholate has the opposite effect, stabilizing the lamellar versus the inverted hexagonal phase. This suggests that cholate is activating phospholipase C through a hitherto undescribed mechanism. Sodium cholate concentrations above 1 mM decrease further the enzyme lag time, but they are less effective in enhancing enzyme rates. These observations may be pertinent in the analysis of biochemical data with purified lipases, as well as in physiological studies of biliary function. Copyright 1999 Academic Press. PMID- 10527584 TI - Lipid-Drug Interaction and Colligative Properties in Phospholipid Vesicles. AB - Imipramine penetration into the lipid core of a membrane was demonstrated through measurements on lipid monolayers (surface pressure and surface potential). The surface pressure measurements allow us to calculate the intrinsic binding constant (partition coefficient) for the lipid-Imipramine interaction. This latter value is in correct agreement with the results obtained by electrophoretic mobility measurements on liposomes. In addition, it was observed that the same mole fraction of "lipid-soluble drug" (Chlorpromazine or Imipramine) incorporated in a given lipidic phase (DPPC) induced the same shift in the transition temperature. Copyright 1999 Academic Press. PMID- 10527585 TI - Dispersive Stabilization of Liquid Crystal-in-Water with Acrylamide Copolymer/Surfactant Mixture: Nematic Curvilinear Aligned Phase Composite Film. AB - The effect of nonionic surfactant, (H(OCH(2)-CH(2))(8)-OC(6)H(4)-C(9)H(19)), on the dispersion stabilization of liquid crystal (LC)-in-water with acrylamide copolymer containing the related nonylphenyl groups was studied. It was observed that the addition of nonionic surfactant increases the stability of LC dispersions and improves the electrooptical properties of the nematic curvilinear aligned phase (NCAP) composite film. On the basis of the surface tension, reduced viscosity, cloud point, and coalescence time measurements, it was proposed that formation of an integrated structure induced by interactions between hydrophobic groups in the polymer chains is probably important to fabrication of a polymer composite film made of LC and polymer matrix. Copyright 1999 Academic Press. PMID- 10527586 TI - Application of the Replica Ornstein-Zernike Equations to Study Submonolayer Adsorption on Energetically Heterogeneous Surfaces. AB - Weapply the replica methodology and the replica Ornstein-Zernike equations to calculate adsorption isotherms on an energetically heterogeneous surface. The surface is modeled by quenching a two-dimensional "fluid of centers." After the quenching, we study gas adsorption using the replica Ornstein-Zernike equation with the hypernetted chain closure. The results of theoretical predictions are compared with the data obtained from the Grand Canonical Monte Carlo simulation, and the obtained agreement is satisfactory. Copyright 1999 Academic Press. PMID- 10527587 TI - Probing Polyethylene Glycol-Phospholipid Membrane Interactions Using Enzymes. AB - Interaction of polyethylene glycols (PEGs) with phospholipid membranes leads to aggregation and fusion of membranes. The structural basis of these events in membranes, especially in contact with low-molecular-weight PEGs, is uncertain. Using phsopholipases, a class of interfacially active enzymes, we demonstrate enhanced accessibility of lipid hydrophobic portions in the presence of PEGs. All three phospholipases, i.e., A(2), C, and D, show enhancement of activity in the presence of PEGs. Enhancement of activity does not depend on the size of the vesicle or the presence of proteins in the membrane. Fluorescence quenching of probes buried in the membrane supports the phospholipase data. The utility of phospholipases as probes to monitor local and fine structural changes in the membranes is discussed. Copyright 1999 Academic Press. PMID- 10527588 TI - Surface Acidity and Hydrophilicity of Coprecipitated Al(2)O(3)-SiO(2) Xerogels Prepared from Aluminium Nitrate Nonahydrate and Tetraethylorthosilicate. AB - Amorphous aluminosilicate xerogels with various chemical compositions were prepared by coprecipitation, and their surface acidity and hydrophilicity were investigated by NH(3) gas temperature programed desorption (TPD), water vapor adsorption-desorption isotherms, and (27)Al magic angle spinning nuclear magnetic resonance (MAS NMR). The xerogels were synthesized by adding conc. NH(4)OH to an ethanol solution of calculated amounts of aluminium nitrate nonahydrate and tetraethylorthosilicate, and calcined at 300 degrees C for 4 h. All the NH(3) TPD spectra of the xerogels showed similar asymmetric peak profiles at around 200 degrees C tailing to the higher temperature side. The amount of acidity evaluated from the peak area of the TPD spectra showed a maximum at around 10 mol% Al(2)O(3) composition. The change as a function of composition showed a good correlation with the total amount of four and five coordinated Al atoms in the xerogels deduced from the (27)Al MAS NMR spectra. The water vapor adsorption isotherms of the xerogels were all of type IV irrespective of the composition. The maximum amounts of water vapor adsorbed by these xerogels were about 600-700 ml(STP)/g and were relatively high compared with those for various other adsorbents reported so far. Since the thickness of the adsorbed water vapor layer of the xerogels in the low relative pressure region increased with increasing Al(2)O(3) content, the surface of the xerogels is considered to become more hydrophilic with increasing Al(2)O(3) content of the xerogels. Copyright 1999 Academic Press. PMID- 10527589 TI - Synthesis, Characterization, and Catalytic Uses of Mg(3)(PO(4))(2)/MgO Systems. AB - Various Mg(3)(PO(4))(2)/MgO systems containing different proportions of both components were synthesized. The solids were characterized in structural terms by X-ray diffraction, thermal gravimetric analysis, and diffuse reflectance infrared spectroscopy; also, their surface properties were determined from N(2) adsorption desorption isotherms and their chemical properties were measured with various titrants. An increasing P content in the solids was found to result in decreasing specific surface area and basicity. The solids were used as catalysts for the transformation of 2-hexanol and the alkylation of aniline with methanol. Dehydrogenation selectivity in the former was found to be highly correlated with basicity of the solid; also, alkylation activity and selectivity toward N,N' dimethylaniline in the latter were found to increase with increasing P content. Copyright 1999 Academic Press. PMID- 10527590 TI - Separation Rate Dependence of Adherence Strength. AB - The adherence energy measured during the separation of adhesive/substrate assemblies is a complex function of the adhesion energy and also of the dissipative properties of both adhesive and substrate. Dissipative phenomena occurring during an assembly separation are reflected in the dependence of the measured energy on separation rate. Up to now, different relationships have been used to describe the dependence of the adherence energy G on separation speed V. However, these models assume that only the separation speed affects the dissipation function; the adhesion energy W is supposed to remain constant. This paper proposes that the "real" adhesion energy is also rate dependent. Indeed, the number of interfacial interaction points under stress is able to increase with speed. This effect could be a consequence of the influence of separation rate on the adhesive elastic modulus. Copyright 1999 Academic Press. PMID- 10527591 TI - Study of the Dissolution of the Barium Sulfate (001) Surface with Hydrochloric Acid by Atomic Force Microscopy. AB - Noncontact atomic force microscopy (NC-AFM) has been used to investigate the morphological changes of a freshly cleaved (001) surface of barium sulfate (barite) etched with an aqueous solution of 0.1 M HCl at room temperature. Shallow triangular etch pits with a height of 3.6 A were developed in atomically flat (001) terraces. The etching of the surface was found to proceed in a layer by-layer dissolution process. Because the crystal structure of barite exhibits a two-fold screw axis parallel to the c axis, "alternating" etch pits were formed, with any two consecutive etch pits pointing opposite to each other. These etch pits became deeper and more elongated along the b axis with time. Copyright 1999 Academic Press. PMID- 10527592 TI - Current treatment for colorectal cancer metastatic to the liver. AB - Surgery is currently the only available treatment option which offers the potential for cure for patients with liver metastases from colorectal cancer. Of those who undergo a potentially curative operation for their primary tumour but subsequently recur, almost 80% will develop evidence of metastatic disease within the liver. Greater experience and improvements in technique in liver surgery, with an increasingly aggressive surgical approach to metastatic colorectal cancer to the liver, has resulted in prolonged disease-free survival with 5-year rates varying from 21% to 48%. In order to increase these numbers further and to treat patients not eligible for surgical therapy, new treatment modalities and strategies have been developed. This review presents an update of the current treatment for colorectal disease metastatic to the liver. PMID- 10527593 TI - Margin assessment by cavity shaving after breast-conserving surgery: analysis and follow-up of 543 patients. AB - AIMS: To analyse cavity shaving as a method of assessing completeness of surgical excision after breast-conserving surgery. METHODS: Shavings were taken from the wall of the cavity remaining in the breast after breast-conserving surgery in 543 women. Each shaving was extensively sectioned and the presence and type of microscopic disease recorded. Disease in cavity shavings (tumour bed positivity) was correlated with clinicopathological factors as well as overall survival. RESULTS: Tumour bed positivity (TBP) was found in 37% of patients (16% with invasive disease). Patients were selected for further surgery according to the extent of positivity, which varied widely. A total of 15% of patients underwent re-excision or mastectomy. TBP was significantly associated with high tumour grade, presence of an extensive intraduct component, young age and large tumour diameter. It was also associated with a significantly shorter overall survival when compared to patients who were tumour bed negative. CONCLUSIONS: Cavity shaving is a practical and sensitive method of assessing completeness of excision after breast-conserving surgery. In addition it may provide useful prognostic information. PMID- 10527594 TI - Immediate breast reconstruction after mastectomy for cancer. AB - AIMS: The oncological, surgical and cosmetic results, patient satisfaction and psychological morbidity of immediate breast reconstruction following mastectomy for breast cancer were evaluated. METHODS: From 1980 to 1994, 79 immediate breast reconstructions were performed in Malmo. From 1985 immediate breast reconstruction was performed in 21% of mastectomies among patients 500,000) is a versatile and widely expressed cytolinker protein. In striated muscle it is predominantly found at the Z-disc level where it colocalizes with the intermediate filament protein desmin. Both proteins show altered labeling patterns in tissues of muscular dystrophy patients. Moreover, mutations in the plectin gene lead to the autosomal recessive human disorder epidermolysis bullosa simplex with muscular dystrophy, and defects in the desmin gene have been shown to cause familiar cardiac and skeletal myopathy. Since intermediate filaments (IFs) in striated muscle tissue have been found to be intimately associated with mitochondria, we investigated whether plectin is involved in this association. Using postembedding immunogold labeling of Lowicryl sections and immunogold labeling of ultrathin cryosections, we show that plectin is associated with desmin IFs linking myofibrils to mitochondria at the level of the Z-disc and along the entire length of the sarcomere. The localization of plectin label at the mitochondrial membrane itself was consistent with a putative linker function of plectin between desmin IFs and the mitochondrial surface. In mitochondrion-rich muscle fibers, both plectin and desmin were part of an ordered arrangement of mitochondrial side branches, which wound around myofibrils adjacent to the Z-discs and were anchored into a filamentous network transversing from one fibril to the other. The association of mitochondria with plectin and IFs was seen also in tissues without regular distribution patterns of mitochondria, such as heart muscle and neonatal skeletal muscle tissues. These data were supplemented with in vitro binding assays showing direct interaction of plectin with desmin via its carboxy-terminal IF-binding domain. As a cytolinker protein associated with mitochondria and desmin IFs, plectin could play an important role in the positioning and shape formation, in particular branching, of mitochondrial organelles in striated muscle tissues. PMID- 10527639 TI - Nature of the critical labile event that controls RB phosphorylation in the cyclic AMP-dependent cell cycle of thyrocytes in primary culture. AB - This study addresses the nature of the critical labile event that couples at restriction point mitogenic cascades with the autonomous part of the cell cycle. In primary cultures of dog thyroid epithelial cells, kinetic experiments indicate that a labile cAMP-dependent event positively controls a late G1 commitment to DNA replication and RB phosphorylation. As previously shown in this system, the cAMP-dependent mitogenic pathway differs from the most generally envisaged growth factor cascades as it stimulates the accumulation of p27(kip1) but not of cyclins D. Nevertheless, cyclin D3 and CDK4 activity are essential in the cAMP-dependent mitogenesis, and cAMP unmasks the DCS-22 epitope of cyclin D3 and induces the nuclear translocations and assembly of cyclin D3 and CDK4 in a complex that also contains p27(kip1). Unexpectedly, the washing out of forskolin rapidly arrested S phase entry and the accumulation of hyperphosphorylated RB, but did not reverse any of the above events associated with cyclin D3-CDK4 activation. This implies that even after induction of stable nuclear cyclin D3-CDK4 complexes, dog thyrocytes still depend on cAMP for RB phosphorylation and commitment to DNA synthesis, which suggests that a key labile event responsible for a last control of restriction point passage remains to be uncovered, in the cAMP-dependent cell cycle of dog thyrocytes and possibly other systems. PMID- 10527640 TI - P-glycoprotein system as a determinant of drug interactions: the case of digoxin verapamil. AB - Digoxin, which has a very narrow therapeutic window, is one of the most commonly prescribed drugs in the treatment of congestive heart failure. In some cases of atrial fibrillation digoxin is used in combination with verapamil. Verapamil can increase the plasma concentration of digoxin up to 60-90%. So far the precise mechanism of this pharmacokinetic drug-drug interaction is not known. Many studies suggest that verapamil reduces the renal clearance of digoxin. The energy dependent membrane-bound transport enzyme, P-glycoprotein, may also be involved. Reports from oncology research show that verapamil can interact with P glycoprotein as a modulator. Also taking into account that digoxin, like many anticancer drugs, is a substrate for P-glycoprotein, it is likely that P glycoprotein modulation accounts for the digoxin-verapamil interaction. Current knowledge suggest that the non-competitive digoxin-verapamil interaction is due to inhibition of P-glycoprotein activity by verapamil resulting in a decreased renal tubular elimination of digoxin. PMID- 10527641 TI - The importance of androgen on noradrenergic responses of vas deferens isolated from acutely stressed rats. AB - This study investigated the importance of androgen on responses to alpha and beta (norepinephrine) and alpha(1)(phenylephrine and methoxamine) agonists in vasa deferentia isolated from adult, immature, cryptorchid, and castrated rats submitted to swimming-induced acute stress. The participation of adrenergic nervous terminals was also investigated. Acute stress was shown to induce a significant subsensitivity to norepinephrine only in vas deferens from adult rats with normal levels of androgens. In addition, sympathetic denervation of the vas deferens prevented the appearance of subsensitivity. Subsensitivity was not seen when the experiments were carried out using phenylephrine and methoxamine. This shows that subsensitivity to norepinephrine in this acute stress situation may depend on other factors such as neuronal uptake, but not on alpha(1)-adrenoceptor response. Thus, when animals are exposed to acute stressogenic situations, this subsensitivity requires physiological levels of androgens to establish, and may also be involved in body homeostasis. PMID- 10527642 TI - Propofol's biphasic effect on GABA(A)-receptor-mediated response of the isolated guinea pig ileum. AB - This study examines the effect of the anaesthetic agent propofol on the guinea pig ileum and whether this effect derives from an interaction of the drug with GABA receptors. Propofol produced a biphasic effect consisting of a dose dependent contractile effect (EC(50)=2.2x10(-5)m) followed by an 'after relaxation'. This propofol effect was similar to the one produced by GABA and was bicuculline-sensitive (ID(50)=3.2x10(-7)m). Propofol, at concentrations of 10( 7)and 10(-6)m, potentiated the ileum contractile responses to GABA, but only at the lower dose range of applied GABA, while at a concentration of 10(-5)m, it inhibited the contractile effect over the entire dose range of applied GABA. In addition, while the contractile response of the ileum to exogenously applied acetylcholine was not influenced by propofol at concentrations of up to 7x10( 6)m, it was antagonised by higher concentrations of propofol. In conclusion, the above data permit us to suggest that propofol's contractile effect on the guinea pig ileum is mediated by an interaction of the drug with GABA(A)-receptors. PMID- 10527643 TI - Effect of amlodipine upon the protective activity of antiepileptic drugs against maximal electroshock-induced seizures in mice. AB - Amlodipine, a calcium channel antagonist of the dihydropyridine class, up to 10 mg kg(-1)(i.p.) did not significantly affect the threshold for electroconvulsions. However, this calcium channel antagonist (10 mg kg(-1)) enhanced the anticonvulsive activity of carbamazepine, valproate and phenobarbital against maximal electroshock-induced seizures in mice. Furthermore, amlodipine (5 mg kg(-1)) intensified the protection offered by carbamazepine. This effect was associated with the increased free plasma level of carbamazepine in the presence of amlodipine. Amlodipine did not influence the free or total plasma level of phenobarbital and valproate, so a pharmacokinetic interaction is not probable for valproate and phenobarbital. The anticonvulsive action and free plasma level of diphenylhydantoin was not modified by amlodipine. The combined treatment of the calcium channel antagonist and antiepileptics caused motor impairment (evaluated in the chimney test). Long-term memory (assessed in the passive avoidance test) in case of combinations of amlodipine with carbamazepine or diphenylhydantoin was not affected. The combination of amlodipine with valproate or phenobarbital significantly influenced the retention in this test. A possible usefulness of amlodipine as add-on therapy in epileptic patients may be limited by its considerable adverse effect revealed by behavioural tests. The pharmacokinetic interaction between carbamazepine and amlodipine might have some clinical importance for patients treated with these drugs. PMID- 10527644 TI - Effects of buprenorphine on motility in morphine post-dependent rats. AB - The effects of buprenorphine (0.0125-0.2 mg kg(-1)) on the locomotor activity of rats were determined 1-2 months after ceasing a chronic treatment with morphine (20 mg kg(-1)for 28 days). In control animals buprenorphine exhibited both depressive and stimulatory actions, as repeatedly described for morphine. In post dependent animals buprenorphine showed a depressive effect similar to that observed in naive ones. On the contrary, a persistent sensitization to the excitatory effect was observed; the degree of cross-sensitization was similar to that of morphine itself (20 mg kg(-1)). The results are discussed in terms of persistent changes in the locomotor response to opioids, possibly correlated to both drug craving and relapse. PMID- 10527645 TI - Acamprosate does not antagonise the discriminative stimulus properties of amphetamine and morphine in rats. AB - Acamprosate (calcium acetylhomotaurinate) is a GABA derivative that prevents drinking relapses in a significant number of alcoholics. Since little is known about the interaction of acamprosate with other addictive drugs, we studied the effects of this agent (as sodium salt) in two groups of rats trained to discriminate, respectively, morphine (1.7 mg kg(-1)i.p.) or amphetamine (0.5 mg kg(-1)i.p.) from solvent in a two-lever fixed ratio 30 operant behaviour reinforced by water access. Accordingly to the finding that acamprosate inhibits the action of excitatory aminoacids, its effects were compared with those of dizocilpine (MK-801), an NMDA antagonist. Results show that acamprosate (170 and 320 mg kg(-1)i.p. ) produced a slight, and not significant, shift to the left of generalization curves of both morphine and amphetamine without affecting response rates. In contrast, MK-801 potentiated response rate effects of both morphine and amphetamine without affecting their generalization curves. As far as discriminative stimuli participate in the relapsing process of addiction, our results do not predict a role of acamprosate in the prevention of amphetamine or morphine abuse relapsing. PMID- 10527646 TI - Effect of dehydroleucodine in experimental colitis in rats and mice. AB - Dehydroleucodine (DhL), a sesquiterpene lactone (SQL) of the guaianolide type isolated from Artemisia douglasiana Besser, shows a pharmacological cytoprotective effect and significantly prevents the formation of gastric and duodenal lesions induced by various necrotising agents in rodents. The effects of DhL, on two models of experimental colitis were examined. Colitis was produced in male Wistar rats by rectal instillation of 5 and 10% acetic acid, following the methods of Eliakim et al. and Le Duc et al., respectively. In mice colitis was produced by rectal instillation of 0.1 ml of 2,4,6-trinitrobenzene sulphonic acid (5 mg in 50% ethanol) (TNB) as previously described by Chin et al. In this study, the administration of DhL 40 mg kg(-1)(1 h before the induction of colitis) significantly decreased mucosal damage. This effect was consistent in both models. The protection provided by DhL was accompanied by significant decreases in diarrhoea and colon weight; and histologically normal mucosa without ulceration and mucus production were observed. This study shows that both TNB and acetic acid colitis can be pharmacologically controlled by DhL. Our results suggest that the protective activity of DhL in experimental colitis is mediated, at least in part, through the increase of glycoprotein synthesis, anti inflammatory effect and inhibition of COX-2 induction, and by inhibiting the degranulation of cells containing monoamines. PMID- 10527647 TI - N -Acetyl-cysteine reduces homocysteine plasma levels after single intravenous administration by increasing thiols urinary excretion. AB - A decrease of plasma homocysteine (Hcy) may represent a therapeutic promise for reducing the impact of atherosclerosis. N -Acetyl-cysteine (NAC) is a thiol containing compound interfering with endogenous thiols, cysteine (Cys) and Hcy, by forming with them mixed disulphides with a possibly more efficient renal clearance. The aim of this work was to assess the effect of NAC intravenous infusion on plasma levels of different forms of Hcy and particularly to verify the effect on Hcy renal excretion. We collected basal blood samples at 0.5, 1, 2, 5, 8 and 24 h after the beginning of NAC infusion (50 mg kg(-1)body wt.) and also 24-h urine samples of the day of NAC infusion and of the day before and of the day after the infusion in ten healthy subjects (mean age 73+/-15). Urinary and plasma thiols (Hcy, Cys and NAC) were assayed by HPLC. Both total plasma Hcy (approx. 69%vs basal values) and Cys (approx. 40%vs basal values) fell progressively, reaching a minimum 5 h after infusion start; total free (i.e. not bound to proteins) Hcy (2.2+/-1.8 down from 4.4+/-4.2 nmol ml(-1)) and Cys (70.4+/-39.8 down from 113. 3+/-61.2 nmol ml(-1)) decreased as well. Reduced (thiolic-free form) Hcy and Cys decreased during infusion, though not as pronounced as for the other forms. Percentagewise, out of the total plasma levels, Hcy and Cys total free form and reduced form tended to increase over infusion as well as their difference (i.e. the plasma mixed disulphide moiety), thus supporting the idea that excess NAC displaces thiols from their plasma binding sites forming mixed disulphides. Urinary total Cys and Hcy excretion significantly increased at the end of the day of NAC infusion (tenfold for Cys and fivefold for Hcy) and reduced appreciably on the following day. Also urinary excretion of the free form of Cys and Hcy increased at the end of the day of NAC infusion, although in a lower amount with respect of total amounts, meaning a reduction of percentage Cys and Hcy excreted as the free form; for none of the patients had proteinuria, the 'free' form of urine thiols has to be identified in the 'reduced' form, the difference between the total and free form reflecting the 'mixed disulphide' moiety. NAC intravenous administration induces an efficient and rapid reduction of plasma thiols, particularly of Hcy; our data support the hypothesis that NAC displaces thiols from their binding protein sites and forms, in excess of plasma NAC, mixed disulphides (NAC-Hcy) with an high renal clearance. This effect may represent the start of an alternative approach in the treatment of hyperhomocysteinaemic conditions. PMID- 10527648 TI - Effect of rifabutin on ethambutol pharmacokinetics in healthy volunteers. AB - The effect of repeated administration of rifabutin on the pharmacokinetics and metabolism of ethambutol was evaluated in ten healthy volunteers. The subjects received a single oral administration of 1200 mg ethambutol on days 1 and 10 and a single daily oral dose of 300 mg rifabutin from days 3 to 9. No statistically significant difference was found in plasma pharmacokinetics (C(max), t(max), AUC, half-life and MRT) and in the renal clearance, whereas a significant decrease in the amount of unchanged ethambutol excreted in urine was observed. The decrease observed in ethambutol urinary excretion may be accounted for by taking into consideration the variability of the urinary excretion of ethambutol reported in the literature. However, a slight, likely not clinically relevant, induction or activation of kidney alcohol and/or aldehyde dehydrogenase isoenzymes by rifabutin cannot be ruled out at present. Evidence exists in the present study for autoinduction of rifabutin metabolism; this is shown by the lower plasma concentrations obtained 24 h after the seventh dose as compared to the theoretical concentrations. PMID- 10527649 TI - Open questions on bioequivalence: some problems and some solutions. AB - This paper focuses on some specific situations where bioequivalence requires careful attention and tailored protocols in order to overcome intrinsic difficulties either marginally covered or fully neglected by operating guidelines. Some problems congregate with serious difficulties, namely high variability, very poorly absorbed drugs and endogenous substances with their own baseline. With endogenous substances, the dilemma faced is whether to subtract baseline from post-dose values in assessing bioequivalence. Either approach has intrinsic problems and is somewhat puzzling. In an attempt to resolve other existing problems, the most appropriate approach should be selected on a case-by case basis, ensuring that the adopted procedure does not conflict with operating guidelines and scientific literature on the matter. Problematic cases include the management of trials with a predominant active metabolite, the absence of a reliable analytical bioassay, the availability of various strengths of the same drug on the market, a wide acceptability titre range, the management of studies on topical drugs that are devoid of systemic activity, the management of drugs that cannot be given for ethical reasons to healthy subjects or that may cause adverse events, especially when a steady state design is required. The parallel group study design appears to be more appropriate than the cross-over or the individual bioequivalence design in assessing drugs with a long half-life. Some pharmacokinetic and statistical analysis-related issues are also discussed such as the sequence/period interaction sometimes encountered in these trials, which, in the absence of the carry-over effect, does not bias the bioequivalence results and the need to process data with non-compartmental pharmacokinetic analysis. PMID- 10527650 TI - Drug glucuronidation and hepatic lipid microsomal membrane profile in cholestatic rats followed paracetamol intoxication. AB - The uridin-diphosphoglucuronyl-transferase (UDP-GT) is a membrane-bound enzyme responsible for glucuronidation of endogenous and exogenous compounds. This work established the UDP-GT activity and its lipid membrane microenvironment in two experimental models: acute paracetamol intoxication, and cholestasis followed by acute paracetamol intoxication. Cholestasis was performed by bile duct ligation. After 7 days animals were injected with paracetamol (BDL-APAP group). Sham operated rats were injected at day 7 with paracetamol (APAP group). Cholestatic and sham-operated rats injected with vehicle (BDL and control groups). UDP-GT activity was measured by a kinetic method for different substrates. Microsomal membrane phospholipid composition, cholesterol content and ultrastructure were determined. BDL-APAP group showed an increment in the UDP-GT activity except for chloramphenicol, morphine and paracetamol if compared to controls and to BDL group. The same increment was observed when BDL-APAP was compared to APAP except for chloramphenicol and lorazepam. Between BDL and APAP groups similar levels of activity were detected except for paracetamol. Microsomal phospholipid profile: phosphatidylcholine showed the lowest content in the BDL group, with a significant recovery in the BDL-APAP and APAP groups. Phosphatidylserine was markedly decreased in the APAP group compared to the rest and phosphatidylinositol was decreased in all the groups if compared to control values. An increment of phosphatidylethanolamine was seen in the APAP and BDL APAP groups if compared to BDL and control values. A significant increment of microsomal cholesterol content was seen in BDL. Under these conditions, a different lipid microenvironment is produced, resulting in an increment of the enzyme activity for a variety of substrates. PMID- 10527651 TI - Chloroacetonitrile-induced toxicity and oxidative stress in rat gastric epithelial cells. AB - Chloroacetonitrile (CAN), is a disinfectant by-product of chlorination of drinking water. Epidemiological studies indicate that exposure to CAN via drinking water might present a potential hazard to human health. The objective of the present work was to investigate the cytotoxic effects as well as the oxidative stress induced by CAN in cultured rat gastric epithelial cells (GECs). GECs were exposed in vitro to different concentrations of CAN (5-40 microm) for 60 min. Also, GECs were incubated with CAN (10 microm) for different time intervals extending to 180 min. Cytotoxicity was determined by assessing cell viability and lactate dehydrogenase (LDH) release, glutathione (GSH) level and lipid peroxidation as indicated by malondialdehyde (MDA) production. Exposure of GECs CAN (10 microm) for 60 min caused a 50% decrease in cell viability and induced an eightfold increase of LDH leakage. In the same experiment, CAN caused a decrease in cellular GSH content to approximately 50% and significantly enhanced MDA accumulation (approx. sevenfold). These toxic responses to CAN were dependent on both concentration and duration of exposure to CAN. There was a good correlation between LDH release and GSH depletion (r =0.96, P<0.05). Treatment of GECs with 5 m mN -acetyl- l -cysteine (NAC) prior to exposure to CAN afforded some degree of protection as indicated by a significant decrease in the LDH leakage (32% of total leakage) and lipid peroxidation (54%) as compared to CAN alone-treated cells. Also, pretreatment of GECs with vitamin C (1 m m) or vitamin E (10 microm) significantly inhibited LDH leakage (20 and 36% of total leakage, respectively). Preincubation with 1 m m desferroxiamine (DFO), a ferric iron chelator, or 10 microm phenanthroline (PHE), a ferrous iron chelator, diminished CAN-induced LDH leakage by 16 and 21% of total leakage, respectively and MDA production by 40 and 44%, respectively. In conclusion, our results suggest that CAN has a potential cytotoxic effect in rat GECs; and thiol group-donors, antioxidants and iron chelators can play a critical role against CAN-induced cellular damage. PMID- 10527652 TI - Modulation of oxidant status by meloxicam in experimentally induced arthritis. AB - Meloxicam is a new non-steroidal anti-inflammatory drug, that possesses a selective inhibition of the inducible isoform of cyclooxygenase enzyme (COX-2) relative to the constitutive one, COX-1. Oxidative stress has been documented to be involved in the aetiology of many pathological conditions. The present study aims to further explore the relationship between free radical generation and the inflammatory process, and extends more to investigate the effect of meloxicam on the oxidant status in experimentally induced arthritis, namely, Freund's adjuvant induced arthritis in rats. Results of the present investigation revealed that animals inoculated with Freund's complete adjuvant showed a biphasic response regarding changes in the right hind paw oedema volume. During the chronic phase of the disease, arthritic animals showed an elevated plasma level of lipid peroxides, enhanced blood glutathione peroxidase activity, with depletion of plasma total thiols and albumin; while no significant effects have been observed on erythrocytic superoxide dismutase activity and plasma total proteins content, as compared to normal untreated rats. Long-term administration of meloxicam, at two dose levels, produced significant antioedemetous effect and succeeded in modulating the altered parameters affected during arthritis. The selected dose regimens of meloxicam did not show any apparent lesions in the gastric mucosa. The results of the present investigation lend further support to the reported observations concerning selective COX-2 inhibitors. The modulatory influence of meloxicam on the oxidant status, particularly on lipid peroxidation and thiols might be a relevant effect accounting for its anti-inflammatory properties. PMID- 10527653 TI - Oxidative stress and pulmonary inflammation: pharmacological intervention with antioxidants. AB - Reactive oxygen and nitrogen species are generated by several inflammatory and structural cells of the airways. These oxidant species may have important effects on different lung cells as regulators of signal transduction, activators of key transcription factors, and modulators of gene expression and apoptosis. Thus, an increased oxidative stress accompanied by reduced endogenous antioxidant defences may have a role in the pathogenesis of a number of inflammatory pulmonary diseases including asthma. Although antioxidant drugs could play a useful role in the therapy of inflammatory lung diseases, their clinical impact is relatively modest at present. Rigorous clinical investigation with the existing antioxidants and development of new drugs with improved lung bioavailability are necessary in the future.pc 1999 Academic Press@p$hr PMID- 10527654 TI - Inhibition of triiodothyronine production by fenugreek seed extract in mice and rats. AB - The effects of fenugreek seed extract on the alterations in serum thyroid hormone concentrations were studied in adult male mice and rats. Simultaneously, hepatic lipid peroxidation (LPO) and the activities of the antioxidant enzymes, viz superoxide dismutase (SOD) and catalase (CAT) were examined. Administration of methi seed extract (0.11 g kg body wt.(-1) day(-1) for 15 days) to both mice and rats significantly decreases serum triiodothyronine (T(3)) concentration and T(3)/T(4) ratio, but increases thyroxine (T(4)) levels and body weight. While hepatic LPO and CAT activities were not altered, a significant decrease in SOD activity was observed in both the animal models. These findings suggest that fenugreek seed extract induced inhibition in T(4)to T(3) conversion is not peroxidation-mediated and the inhibition in SOD activity could be the result of a decrease in the protein anabolic hormone, T3. PMID- 10527655 TI - The effect of eltanolone on pulmonary vascular resistance in landrace swine. AB - This paper reports on the haemodynamic effects of eltanolone observed in Landrace swine during the investigation of the drug with respect to safety in malignant hyperthermia-susceptible individuals. Pigs were sedated with intramuscular ketamine, followed by induction of anaesthesia employing thiopentone administered via an ear-vein. After intubation, anaesthesia was maintained using nitrous oxide in oxygen. A total of eight pigs were then further anaesthetised on two separate occasions using one of two dose schedules. A bolus of 1.5 mg kg(-1) of eltanolone was administered, followed by a continuous infusion at either 2 or 10 mg kg(-1) h(-1). There were no significant changes in heart rate, mean arterial pressure, cardiac output or systemic vascular resistance following eltanolone. In all cases eltanolone induced marked rises in pulmonary artery pressure and pulmonary vascular resistance (P<0.01) at all measuring points and in right ventricular stroke work at 6-10 min after drug exposure. We conclude that the selective influence of eltanolone on the pulmonary vasculature is probably species specific, but may have clinical significance in patients with pulmonary hypertension. PMID- 10527657 TI - 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002) inhibits nitric oxide production in cultured murine astrocytes. AB - The level of nitrite accumulation in the culture media of astrocytes activated with lipopolysaccharide (LPS) and interferon-gamma (IFN) was decreased by pretreatment of cells with LY294002, a quercetin derivative developed for phosphatidylinositol-3-kinase (PI3K) inhibitor, in a dose-dependent manner. The expression of iNOS mRNA in the astrocytes was inhibited by LY294002, as revealed by reverse transcriptional polymerase chain reaction and agarose gel electrophoresis. The catalytic activity of astrocytic iNOS was also inhibited by LY294002. On the other hand, wortmannin which was known to enhance endotoxin induced NO production in macrophages by inhibiting PI3K did not cause any significant change in the NO production and iNOS mRNA expression of the astrocytes. These results suggest that LY294002 suppresses NO production in the astrocytes through not only the inhibition of iNOS mRNA expression but also the inhibition of the iNOS activity and that PI3K is not involved in the inhibitory actions of LY294002.pc 1999 Academic Press@p$hr PMID- 10527656 TI - Inhibitory effects of cannabinoid receptor ligands on electrically-evoked responses in rat isolated tracheal ring segments. AB - We have examined the possible existence of cannabinoid receptors in the isolated rat tracheal ring segments by studying the effects of some cannabinoid receptor ligands on electrically-induced contractions. Anandamide (10(-8)-3 x 10(-5)m), an endogenous ligand for cannabinoid receptors, and WIN 55,212-2 (10(-9)-3 x 10( 5)m), a moderately selective CB(2)agonist, inhibited electrically evoked contractions of the rat tracheal ring segments in a concentration-related manner. Addition of phentolamine (10(-6)m) to Krebs Henseleit solution to block alpha(2) adrenoceptors did not affect anandamide-induced inhibition of the electrically evoked contractions. The EC(25)(-log m) values were 5.25+/-0.2 and 5.8+/-0. 4 for anandamide and WIN 55,212-2, respectively. The maximal inhibition produced by the highest concentration of the agonists used was 51.4+/-5.8% for anandamide and 35.1+/-19.5% for WIN 55, 212-2. WIN 55,212-3 also produced a concentration dependent inhibition of the electrically evoked contractions. The maximal inhibition produced by WIN 55,212-3 was 15.8+/-2.4. The inhibitory effects of anandamide and WIN 55,212-2 were not attenuated by SR141716A (10(-6)m), a selective CB(1)receptor antagonist. Anandamide (10(-8)-3 x 10(-5)m) did not relax rat tracheal ring segments pre-contracted with carbachol (10(-6)m). These results suggest that anandamide and WIN 55,212-2 produce pre-junctional inhibitory effects in the rat trachea and that these effects were likely mediated through cannabinoid CB(2)receptors. These effects were probably non-cannabinoid receptor mediated considering the high concentrations of the agents required to produce inhibitory responses and the effectiveness of WIN 55,212-3. PMID- 10527658 TI - Cellular toxicity of N-substituted 2,2'-dicarboxamidodiphenyldisulphides with high antimicrobial activity. AB - In the present paper we evaluate the cellular toxicity of some N-substituted 2,2' dicarboxamidodiphenyldisulphides with high antimicrobial activities, in view of their potential application in humans or animals. The toxicological studies have been conducted in the murine cell line of 3T3 fibroblasts as eucaryotic cellular model. Our results have allowed the identification of a series of derivatives exhibiting antimicrobial activity and low cellular toxicity. Structure-cytotoxic activity relationships are also discussed. PMID- 10527659 TI - ACEA-1328, a NMDA receptor/glycine site antagonist, acutely potentiates antinociception and chronically attenuates tolerance induced by morphine. AB - The effect of ACEA-1328, a competitive and systemically bioavailable NMDA receptor/glycine site antagonist, was studied on morphine-induced antinociception and tolerance in CD-1 mice using the tail flick test. To study the effect of acute administration of ACEA-1328 on morphine-induced antinociception, mice were injected with either ACEA-1328 (1, 5, and 10 mg kg(-1)) or Bis-Tris (0.2 m) immediately followed by an injection of morphine and tested for antinociception 30 min later. ACEA-1328 significantly increased the antinociceptive potency of morphine. To study the effect of chronic administration of ACEA-1328 on morphine induced antinociception and tolerance, mice were treated, either once per day for 9 days or twice daily for 4 days, with ACEA-1328 or with the vehicle. Mice were then, within 1 min, injected daily with either morphine or saline. On the day of the test, mice were injected with only morphine and tested for antinociception 30 min later. In comparison to the acute effect of ACEA-1328, chronic treatment with the NMDA receptor/glycine site antagonist did not affect the antinociceptive potency of morphine. Chronic treatment with morphine, by both methods, produced a significant degree of tolerance. Concurrent administration of ACEA-1328 with the opioid analgesic completely blocked morphine tolerance. Our results demonstrate that acute, but not chronic, treatment with ACEA-1328 increased the antinociceptive potency of morphine. Furthermore, co-administration of the NMDA receptor antagonist with morphine abolished the development of tolerance. Overall, the data support a growing body of evidence showing that activation of the NMDA receptor plays a functional role in opioid-induced antinociception and tolerance. PMID- 10527661 TI - Improved gene transfer efficiency in liver with vesicular stomatitis virus G protein pseudotyped retrovirus after partial liver resection and thymidine kinase ganciclovir pre-treatment. AB - Liver-directed gene therapy is a promising alternative for the treatment of various liver diseases. Pseudotyped (VSV-G) retroviruses can be produced in high titres which is essential to overcome the problem of low gene transfer efficiency detected previously with first generation Moloney murine (MMLV) retroviruses and plasmid vectors. We compared the lacZ gene transfer efficiency of MMLV retroviruses and VSV-G retroviruses in Watanabe heritable hyperlipidaemic rabbit liver using an intraportal administration route. Hepatocyte proliferation was stimulated by a partial (10%) liver resection and a thymidine kinase-ganciclovir treatment. We also studied the safety of the gene transfer by clinical chemistry, tissue pathology and PCR analysis of lung, kidney, spleen and gonads. Gene transfer efficiency with the VSV-G retrovirus was significantly higher than with the traditional MMLV-based retrovirus (9.5+/-5.26 vs 0.21+/-0.10 positive hepatocytes mm(-2), P<0.05). After a 12-month follow-up period no lacZ expression was detected in liver samples. No transgene was detected in plasma or in lung, kidney, spleen and gonads by PCR analysis 7 days after gene transfer. Transient increases were found in plasma c-reactive protein, aspartyl aminotransferase and alanine aminotransferase levels shortly after the operation with both types of retroviruses. VSV-G retrovirus was well tolerated and may become an efficient new tool in liver gene therapy. The absence of transgene in systemic circulation or in extrahepatic tissues including gonads is an important safety feature required for in vivo gene therapy. PMID- 10527660 TI - Muscarinic M(2) receptors are not primarily involved in the contraction of guinea pig gallbladder smooth muscle. AB - The presence of M(1)-M(4) receptors in guinea-pig gallbladder smooth muscle cells has been reported recently. The majority of these receptors are said to be of M(2) subtype. However, there are controversial reports about the functional muscarinic receptors that mediate contraction in this tissue. Similar to gallbladder, it was claimed that M(4) receptors mediate guinea-pig uterine contractions, but these receptors have appeared to be of M(2) subtypes later. Therefore, the antagonistic affinities of three M(2)-selective muscarinic antagonists were determined in contraction and radioligand binding experiments in guinea-pig gallbladder in the present study. The antagonistic affinity values (p K(i)) of gallamine, tripitramine and imperialine were as follows, respectively: 6.28+/-0.15, 8.65+/-0.10 and 6.55+/-0.07 against 0.250 n m [(3)H]QNB binding. All three antagonists displaced the concentration- response curves to carbachol to the right in parallel without affecting the maximum responses. The p A(2) values obtained from constrained Schild plots (-log K(B)) were 4.14+/-0.18 for gallamine, 6.79+/-0.09 for tripitramine, and 7.02+/-0.09 for imperialine. The antagonistic affinity values of gallamine, tripitramine and imperialine for M(2) receptors are reported to be 6. 3, 9.6, 7.7, respectively. The p A(2) values obtained in this study clearly indicate that the primary muscarinic receptors involved in carbachol-induced guinea-pig gallbladder contraction are not of M(2) subtype. The poor correlation between the antagonistic affinity values of these antagonists obtained at radioligand binding (p K(i)) and contraction (p A(2)) experiments also support the conclusion that the majority of muscarinic receptors which have been reported to be of M(2) do not mediate the contractile responses. PMID- 10527662 TI - Is there treatment for "genetic" disease? PMID- 10527663 TI - The structural basis of phenylketonuria. AB - The human phenylalanine hydroxylase gene (PAH) (locus on human chromosome 12q24.1) contains the expressed nucleotide sequence which encodes the hepatic enzyme phenylalanine hydroxylase (PheOH). The PheOH enzyme hydroxylates the essential amino acid l-phenylalanine resulting in another amino acid, tyrosine. This is the major pathway for catabolizing dietary l-phenylalanine and accounts for approximately 75% of the disposal of this amino acid. The autosomal recessive disease phenylketonuria (PKU) is the result of a deficiency of PheOH enzymatic activity due to mutations in the PAH gene. Of the mutant alleles that cause hyperphenylalaninemia or PKU 99% map to the PAH gene. The remaining 1% maps to several genes that encode enzymes involved in the biosynthesis or regeneration of the cofactor ((6R)-l-erythro-5,6,7,8-tetrahydrobiopterin) regenerating the cofactor (tetrahydrobiopterin) necessary for the hydroxylation reaction. The recently solved crystal structures of human phenylalanine hydroxylase provide a structural scaffold for explaining the effects of some of the mutations in the PAH gene and suggest future biochemical studies that may increase our understanding of the PKU mutations. PMID- 10527664 TI - Molecular mechanisms of holoprosencephaly. AB - Holoprosencephaly (HPE) is the most common developmental defect of the forebrain in humans. Several distinct human genes for holoprosencephaly have now been identified. They include Sonic hedgehog (SHH), ZIC2, and SIX3. Many additional genes involved in forebrain development are rapidly being cloned and characterized in model vertebrate organisms. These include Patched (Ptc), Smoothened (Smo), cubitus interuptus (ci)/Gli, wingless (wg/Wnt, decapentaplegic (dpp)/BMP, Hedgehog interacting protein (Hip), nodal, Smads, One-eyed pinhead (Oep), and TG-Interacting Factor (TGIF). However, further analysis is needed before their roles in HPE can be established. Here we present an overview of the presently known genes causing human holoprosencephaly and describe candidate genes involved in forebrain development identified in other systems. A model is discussed for how these genes may interact within and between several different signaling pathways to direct the formation of the forebrain. PMID- 10527665 TI - Craniosynostosis syndromes: from genes to premature fusion of skull bones. PMID- 10527666 TI - Making CENs of mammalian artificial chromosomes. AB - Mammalian artificial chromosomes (MACs) hold the promise of providing autonomous vectors for gene therapy in dividing cells. They would not require insertion into the genome and could include sufficient genomic sequences that surround the therapeutic gene to ensure proper tissue-specific and temporal regulation. Several groups have reported successful formation of MACs in human cells using transfection strategies that included alpha satellite DNA, the primary DNA found at normal human centromeres. These results, although extremely encouraging, have limitations such as unpredictable chromosome formation and success thus far in only one transformed human cell line. Examination of other cells where alpha satellite DNA has integrated into ectopic chromosomal locations, as well as naturally occurring dicentric and neocentromere-containing cell lines, suggests that alpha satellite DNA may not be necessary or sufficient for centromere formation. Overall, these results suggest that epigenetic modifications of centromeric DNA are required for efficient centromere formation. Models for this centromere-specific epigenetic modification include a specialized chromatin structure and differential replication timing of centromeric DNA. Thus, further investigation of these centromere-specific epigenetic modifications may suggest strategies for increasing the efficiency of generating human artificial chromosomes for use as gene therapy vectors. PMID- 10527667 TI - Novel aspects of the insulin-like growth factor binding proteins. AB - The insulin-like growth factors (IGFs), IGF binding proteins (IGFBPs), and IGFBP proteases regulate somatic growth and cellular proliferation both in vivo and in vitro. IGFs are potent mitogens whose actions are determined by the availability of free IGFs to interact with IGF receptors. IGFBPs comprise a family of six proteins that bind IGFs with high affinity and specificity and thereby regulate IGF-dependent actions. IGFBPs have also recently emerged as IGF-independent regulators of cell growth. Several IGFBP association proteins have been discovered recently which can affect IGFBP action. Cleavage of IGFBPs by specific proteases modulates levels of free IGFs and IGFBPs and thereby their actions. IGFBP-related proteins (IGFBP-rPs) are an emerging group of proteins which bind IGFs with low affinity and also play important roles in cell growth and differentiation. The IGFBPs appear to have emerging roles in the mechanisms underlying human cancer. The GH-IGF-IGFBP axis is complex and powerful. Future research on its physiology promises exciting insights into cell biology as well as advancements in the treatment of a wide range of disease states including cancer, diabetes, vascular disease, asthma, and growth disorders. PMID- 10527668 TI - The progesterone receptor and its isoforms in mammary development. PMID- 10527669 TI - The molecular basis of human hypogonadotropic hypogonadism. AB - Patients with hypogonadotropic hypogonadism (HH) present with delayed puberty, infertility, and low serum gonadotropins. The molecular basis for most cases of HH is unknown, but single gene mutations have been described for some hypothalamic and pituitary genes. Kallmann syndrome due to KAL gene mutations and adrenal hypoplasia congenita/HH caused by AHC gene mutations are both X-linked recessive disorders. Mutations in the gonadotropin releasing hormone receptor, leptin, and the leptin receptor cause autosomal recessive HH. In addition, isolated deficiencies of follicle stimulating hormone and luteinizing hormone in the corresponding specific beta-subunit genes and PROP1 gene mutations represent pituitary deficiency states, resulting in a phenotype of HH. Despite these remarkable advances in our understanding of human HH, the cause of approximately 90% remains unknown. PMID- 10527671 TI - Type 2 gaucher disease: an expanding phenotype. PMID- 10527670 TI - Leber congenital amaurosis. AB - Leber's congenital amaurosis (LCA) is the earliest and most severe form of all inherited retinal dystrophies responsible for congenital blindness. Genetic heterogeneity of LCA has been suspected since the report by Waardenburg of normal children born to affected parents. In 1995, we localized the first disease causing gene, LCA1, to chromosome 17p13 and confirmed the genetic heterogeneity. In 1996, we ascribed LCA1 to mutations in the photoreceptor-specific guanylate cyclase gene (retGC1). RetGC1 is an essential protein implicated in the phototransduction cascade, especially in the recovery of the dark state after the excitation process of photoreceptor cells by light stimulation. In 1997, mutations in a second gene were reported in LCA, the RPE65 gene, which is the first specific retinal pigment epithelium gene. The protein RPE65 is implicated in the metabolism of vitamin A, the precursor of the photoexcitable retinal pigment (rhodopsin). Finally, a third gene, CRX, implicated in photoreceptor development, has been suspected of causing a few cases of LCA. Taken together, these three genes account for only 27% of LCA cases in our series. The three genes encode proteins that are involved in completely different physiopathologic pathways. Based on these striking differences of physiopathologic processes, we reexamined all clinical physiopathological discrepancies and the results strongly suggested that retGC1 gene mutations are responsible for congenital stationary severe cone-rod dystrophy, while RPE65 gene mutations are responsible for congenital severe but progressive rod-cone dystrophy. It is of tremendous importance to confirm and to refine these genotype-phenotype correlations on a large scale in order to anticipate the final outcome in a blind infant, on the one hand, and to further guide genetic studies in older patients on the other hand. PMID- 10527672 TI - Phosphomannomutase deficiency: the molecular basis of the classical Jaeken syndrome (CDGS type Ia). PMID- 10527673 TI - Mitochondria: ignition chamber for apoptosis. PMID- 10527674 TI - Is the Laron mouse an accurate model of Laron syndrome? PMID- 10527675 TI - Somatic mutation as mechanism for cyst formation in autosomal dominant polycystic kidney disease. PMID- 10527676 TI - Recent developments in the investigation of inherited metabolic disorders using cultured human cells. AB - Thepurpose of this paper is to share experience with our systems and review recent "in vitro" methods using intact cells (fibroblasts, amniocytes) in which entire metabolic pathways can be probed for inherited metabolic defects reflected by elevations of intermediates determined by tandem mass spectrometry, HPLC, or gas chromatography-mass spectrometry. Currently, one can explore the integrity of mitochondrial fat oxidation, peroxisomal degradation of methyl-branched fatty acids (e.g., pristanate), and the mitochondrial degradation of the branched chain amino acids (leucine, valine, and isoleucine). For many of the diseases, the specific defect can be recognized from the acylcarnitine profile resulting from incubation of the intact cells with stable-isotope-labeled precursors to the particular pathway. This approach has also been successful in identifying new inherited metabolic disorders, biochemical correlation with clinical phenotypes of individual defects, and sequential oxidation of fatty acids by peroxisomal mitochondrial interaction. PMID- 10527677 TI - Glucose regulation of the expression of the glucagon receptor gene. AB - The glucagon receptor gene is a member of a gene family, the expression of which is strongly upregulated by glucose. This review deals with the structure of both the glucagon receptor gene and its promoter. Attention is focused on the glucose regulatory element that we discovered in the promoter of this gene. Regulation by glucose of genes implicated in glucose homeostasis represents one mechanism contributing to the control of fuel utilization. Its deficiency or imbalance could potentially lead to or participate in pathological situations such as diabetes mellitus. On the other hand, the regulatory element of the glucagon receptor gene promoter could be used as a tool for the glucose-regulated expression of other genes. Indeed, an analysis of the glucagon receptor gene promoter demonstrated that only a short fragment of the genomic DNA, easy to subclone, contains all required elements for activation by glucose. Its potential use for gene therapy is also considered, therefore, in this report. PMID- 10527678 TI - Immune response to enzyme replacement therapy in lysosomal storage disorder patients and animal models. AB - The lysosomal storage disorders (LSD) are a group of severe multiple pathology disorders characterized by enzyme deficiencies which cause the lysosomal accumulation of undegraded or partially degraded macromolecules. Enzyme replacement therapy (ERT) has been developed as a therapy for LSD patients. However, immune responses to ERT have been reported in some individuals from LSD animal model and LSD human patient studies. Antibodies can have adverse effects during ERT, which include hypersensitivity/anaphylactic reactions, enzyme inactivation, altered targeting, and increased enzyme turnover. The monitoring of antibody production during replacement therapy is an important consideration for patient management, as high-titer antibodies can affect the safety and efficacy of the therapy. PMID- 10527679 TI - Gene targeting for gene therapy: prospects. AB - Ideally, gene therapy involves the correction of genetic defects through the natural means of gene targeting. This therapy possesses a number of conceptual advantages. However, a major obstacle to successful gene therapy is the relative inefficiency of the targeting process in mammalian cells. Gene targeting may be accomplished by two different mechanisms: the homologous recombination and the mismatch correction of DNA heteroduplexes. Based on the model of homologous recombination for the well-studied prokaryotic and the less studied eukaryotic systems, three approaches have been employed to improve the efficiency and accuracy of homologous recombination events. These are: (1) artificial double strand breaks in both the exogenous and the chromosomal DNA, (2) a contiguous long homology between the exogenous and chromosomal DNA, and (3) a transient overproduction of an active recombinase, the bacterial RecA or mammalian RecA like proteins, in mammalian cell nuclei. Combining these approaches can result in more effective gene targeting protocols. The second mechanism has been improved based on recent observations of recombinogenic activity of oligonucleotides and, especially, specifically designed chimeric RNA/DNA oligonucleotides. The use of RecA-like proteins to stimulate searching for homology and forming stable DNA heteroduplexes between oligonucleotides and chromosomal DNA remains an attractive idea for additional improvement of gene targeting events. PMID- 10527680 TI - Clinical, molecular, and cell biological aspects of Chediak-Higashi syndrome. AB - Chediak-Higashi syndrome (CHS) is a rare autosomal recessive disorder characterized by variable degrees of oculocutaneous albinism, easy bruisability, and bleeding as a result of deficient platelet dense bodies, and recurrent infections, with neutropenia, impaired chemotaxis and bactericidal activity, and abnormal NK cell function. Neurologic involvement is variable, but often includes peripheral neuropathy. Most patients also undergo an "accelerated phase," which is a nonmalignant lymphohistiocytic infiltration of multiple organs resembling lymphoma. Death often occurs in the first decade from infection, bleeding, or development of the accelerated phase. The hallmark of CHS is the presence of huge cytoplasmic granules in circulating granulocytes and many other cell types. These granules are peroxidase-positive and contain lysosomal enzymes, suggesting that they are giant lysosomes or, in the case of melanocytes, giant melanosomes. The underlying defect in CHS remains elusive, but the disorder can be considered a model for defects in vesicle formation, fusion, or trafficking. Because the beige mouse demonstrates many characteristics similar to those of human CHS patients, including dilution of coat color, recurrent infections, and the presence of giant granules, it is considered the animal homologue of CHS. The beige gene, Lyst, was mapped and sequenced in 1996, prompting identification of the human LYST gene on chromosome 1q42. Lyst and LYST show 86.5% sequence homology. LYST encodes a 429 kDa protein with a function that remains unknown, but the source of extensive speculation among students of cell biology. PMID- 10527681 TI - Dystrophin and the retina. PMID- 10527682 TI - The ABCR gene: a major disease gene in macular and peripheral retinal degenerations with onset from early childhood to the elderly. PMID- 10527683 TI - Genotype-phenotype correlations in disorders of peroxisome biogenesis. AB - Genetically determined human peroxisomal disorders are subdivided into two major categories: disorders of peroxisome biogenesis (PBD), in which the organelle is not formed normally, and those that involve a single peroxisomal enzyme. Twelve PBD have been identified, and the molecular defects have been defined in 10. All involve defects in the import of proteins into the organelle. Factors required for this import are now referred to as peroxins (PEX) and form the basis of a new and preferred classification system. The PBD are associated with four clinical phenotypes, named before their association with the organelle was recognized: Zellweger syndrome (ZS), neonatal adrenoleukodystrophy (NALD), infantile Refsum disease (IRD), and rhizomelic chondrodysplasia punctata (RCDP). The first three are associated with 9 of the 10 PEX defects that have been defined so far, and represent a clinical continuum with variant severity, with ZS the most severe, NALD intermediate, and IRD the least severe. RCDP is associated with PEX7. Genotype-phenotype correlations are complicated by the fact that the clinical manifestations of the ZS-NALD-IRD continuum can be mimicked by disorders that affect single enzymes of peroxisomal fatty acid oxidation, and PEX7 by disorders of plasmalogen synthesis enzymes. Furthermore, clinical manifestations of each of the PEX disorders may vary. Phenotypic expression varies with the nature of the mutation, the milder phenotypes being associated with mutations that do not abolish function completely, or with mosaicism. Definition of the molecular defects is of great value for genetic counseling and may be of aid in establishing prognosis. PMID- 10527684 TI - The G-protein-coupled, extracellular Ca(2+)-sensing receptor: expression in pancreatic islet B-cells and possible role in the regulation of insulin release. PMID- 10527685 TI - Anti-adhesion molecule therapy as an interventional strategy for autoimmune inflammation. AB - Functional inactivation of leukocyte adhesion molecules has been used to intervene in the development of tissue injury in experimental models of postperfusion infarction as well as autoimmune inflammation. We investigated the use of humanized monoclonal antibodies (mAb) against CD18 in the treatment of five patients with vasculitic tissue injury sufficient to threaten infarction or gangrene. The treatment was monitored in three ways: (i) whole-body gamma camera scintiscanning of autologous indium-labeled PMN, (ii) an index of the therapeutic inhibition of adhesion derived from comparison pre, during, and post mAb treatment of the ability of patients' PMN to be aggregated after activation by fMLP, and (iii) flow cytometric analysis of PMN CD18 expression. Four of five patients given anti-CD18 at 20 mg/day for up to 3 weeks showed prompt clinical improvement, with healing of the ulceration and restoration of limb function within 4 weeks, which was sustained. The fifth patient, who was not doing well clinically, decided to withdraw from all active treatment: at autopsy there was no evidence of the underlying vasculitis evident pretreatment. Our findings suggest that anti-adhesion molecule treatment might be an effective immediate treatment in severe vasculitis especially when tissue viability is threatened by progressive infarction and/or development of gangrene. PMID- 10527686 TI - Blocking ICAM-1 (CD54) and LFA-1 (CD11a) inhibits experimental allergic conjunctivitis. AB - Cell adhesion molecules are critical for the homing and migration of leukocytes into inflamed tissues. We investigated the role of ICAM-1 and LFA-1 in a previously described experimental model of ragweed (Rw)-induced allergic conjunctivitis. SWR/J mice were treated intraperitoneally 6 and 1 h prior to topical challenge with Rw with injections of anti-ICAM-1 monoclonal antibody (mAb), anti-LFA-1 mAb, both anti-ICAM-1 and anti-LFA-mAbs, or rat IgG. Blocking ICAM-1 or LFA-1 reduced the clinical signs of allergic conjunctivitis. Treatment with anti-ICAM-1 or anti-LFA-1 mAbs also significantly inhibited cellular infiltration into the conjunctiva. The greatest inhibitory effect was achieved with the combination of antibodies against both cell adhesion molecules. Since antibodies against ICAM-1 and LFA-1 significantly inhibit the development of the clinical and histologic signs of allergic conjunctivitis, they may be useful for treating patients with ocular allergy. PMID- 10527687 TI - Aberrant expression of CD27 and soluble CD27 (sCD27) in HIV infection and in AIDS associated lymphoma. AB - CD27 is a member of the tumor necrosis factor receptor superfamily that is expressed primarily on T cells, as well as on subsets of B cells and NK cells. CD70, which is expressed on activated B and T cells, but not on resting lymphocytes, is a ligand for CD27. Cell surface CD27 can be proteolytically cleaved to produce a 32-kDa soluble CD27 (sCD27) molecule. Elevated levels of sCD27 are seen in a number of disease states and malignancies. Although it has been reported that cerebrospinal fluid sCD27 levels were elevated in people who had AIDS dementia, little is known about CD27 expression in HIV disease. To determine if sCD27 levels were elevated in those with HIV infection, and/or in those with AIDS-associated non-Hodgkin's lymphoma (AIDS-NHL), sCD27 levels were measured in HIV-negative and HIV-positive subjects as well as in people who developed AIDS-NHL. Serum sCD27 levels were seen to be elevated in HIV+ subjects. Furthermore, sCD27 levels were particularly elevated in those subjects who went on to develop AIDS-NHL, with serum sCD27 levels in AIDS-NHL subjects being significantly higher than those in HIV+ subjects who did not develop lymphoma. Most AIDS-NHL cell lines and primary AIDS-NHL tumor specimens expressed both CD27 and its ligand, CD70. The proportion of circulating B cells that expressed cell surface CD27 was substantially reduced in those with HIV infection, and B cells from HIV-infected subjects produced decreased levels of sCD27 in culture. Together, these results indicate that CD27/sCD27 expression is abnormal in HIV infection and suggest that this molecule merits further examination as a potential marker for AIDS-NHL. PMID- 10527688 TI - Peptide T blocks GP120/CCR5 chemokine receptor-mediated chemotaxis. AB - We previously reported that certain short gp120 V2 region peptides homologous to vasaoactive intestinal peptide (VIP), such as "peptide T," were potent inhibitors of gp120 binding, infectivity, and neurotoxicity. The present study shows that synthetic V2-region-derived peptides have potent intrinsic chemotaxis agonist activity for human monocytes and also act as antagonists of high-affinity (0.1 pM) gp120-mediated monocyte chemotaxis. Selectivity is shown in that peptide T is more potent at suppressing M-tropic than T-tropic gp120 chemotaxis. Peptide T was also able to suppress monocyte chemotaxis to MIP-1beta, a chemokine with selectivity for CCR5 chemokine receptors, while chemotaxis of the more promiscuous ligand RANTES was not inhibited, nor was chemotaxis mediated by SDF 1alpha. In order to determine if peptide T mediated its gp120 antagonistic effects via modulation of CCR5 receptors, RANTES chemotaxis was studied using a CCR5 receptor-transfected HOS cell line. In this case, RANTES chemotaxis was potently inhibited by V2-region-derived short peptides. Peptide T also partially suppressed (125)I-MIP1-beta binding to human monocytes, suggesting action at a subset of MIP1-beta receptors. The V2 region of gp120 thus contains a potent receptor binding domain and synthetic peptides derived from this region modulate CCR5 chemokine receptor chemotactic signaling caused by either gp120 or chemokine ligands. The results have therapeutic implications and may explain recent clinical improvements, in that HIV/gp120 actions at CCR5 receptors, such as occur in the brain or early infection, would be susceptible to peptide T inhibition. PMID- 10527690 TI - Expression of extracellular matrix ligands and receptors in the muscular tissue and draining lymph nodes of mdx dystrophic mice. AB - The mdx mouse, an animal model of Duchenne muscular dystrophy, develops an X linked recessive inflammatory myopathy. During onset of disease and height of myonecrosis, mdx mice also display important changes in the microenvironment of lymphoid tissues. Draining lymph nodes showed reduced cellularity and atrophy accompanied by intense immunolabeling for fibronectin, laminin, and type-IV collagen. Following clinical amelioration of dystrophy, mdx mice showed enhanced cellularity and a consistent increase in the absolute numbers of CD4(+) and CD8(+) cells expressing alpha4(high) and alpha5(high) extracellular matrix receptors. Furthermore, infiltrating cells in the proximity of myonecrosis expressed alpha4, alpha5, and alpha6 integrin chains during both height of myonecrosis and muscular tissue regeneration. Such results indicate that during distinct phases of muscular dystrophy, altered expression of extracellular matrix ligands and receptors may be influencing myonecrosis by promoting adhesion and migration of mononuclear cells into the altered skeletal muscle and toward local draining lymphoid tissue. PMID- 10527689 TI - Immunoglobulin VH gene expression in human aging. AB - Immune responses change in aging humans, but it is not known whether there is an age-associated change in the expressed B cell repertoire. We compared Ig VH cDNA libraries from circulating B cells of five elderly and three young human adults. As in young persons, nearly two-thirds of the cDNA clones from older subjects had zero to three V(H) mutations, although there was more individual variation among the elderly. V(H)4 family expression increased in older subjects, both in unmutated and in mutated cDNA clones, whereas V(H)3 family expression predominated in young adults. To test for bias toward activated cells in the cDNA libraries, we studied two older persons by both cDNA library analysis and single cell RT-PCR. In one subject, more than 85% of VH segments were unmutated by either analysis. In the second, mutated Ig segments were much more frequent in cDNA clones than in consecutive single cells; however, V(H) family usage and high representation of particular genes were similar in both analyses. While aging humans continue to produce naive B cells with unmutated Ig genes, a shift to greater use of the V(H)4 family members and expression of particular genes may reflect changes in selection of developing B cells before affinity maturation toward reactivity with foreign antigen. PMID- 10527691 TI - Curcumin causes the growth arrest and apoptosis of B cell lymphoma by downregulation of egr-1, c-myc, bcl-XL, NF-kappa B, and p53. AB - It has been well known that curcumin is a powerful inhibitor of proliferation of several tumor cells. However, the molecular basis of the anti-proliferative effect of curcumin has not been investigated in detail. In this paper, we present evidence to show that curcumin inhibited proliferation of a variety of B lymphoma cells. At low concentrations curcumin inhibited the proliferation of BKS-2, an immature B cell lymphoma, more effectively than that of normal B lymphocytes and caused the apoptosis of BKS-2 cells in a dose- and time-dependent manner. Furthermore, curcumin downregulated the expression of survival genes egr-1, c myc, and bcl-X(L) as well as the tumor suppressor gene p53 in B cells. In addition, NF-kappaB binding activity was also downregulated almost completely by curcumin. Stimulation with CpG oligonucleotides or anti-CD40 overcame growth inhibition induced by low concentrations of curcumin. Our results suggest that curcumin caused the growth arrest and apoptosis of BKS-2 immature B cell lymphoma by downregulation of growth and survival promoting genes. PMID- 10527692 TI - Gene conversion events contribute to the polymorphic variation of the surrogate light chain gene lambda 5/14.1. AB - Normally occurring and experimentally induced models of immunodeficiency indicate that B cell development and antibody production are influenced by genetic factors. It is highly likely that polymorphic variants in genes that encode receptors for growth and differentiation factors, signal transduction molecules, and components of the B cell and pre-B-cell receptor complex contribute to this genetic control. We have identified a surprisingly large number of polymorphic variants in lambda5/14.1. Together with VpreB, lambda5/14.1 forms the surrogate light chain in the pre-B-cell receptor complex. Thirteen variant alleles of lambda5/14.1 were found in 134 unrelated individuals. Nine of these variants result in changes in the amino acid sequence of this small protein. The majority of the single base pair substitutions in lambda5/14.1 could be attributed to gene conversion events in which donor sequences from the lambda5 pseudogenes, 16.1, 16.2, and Glambda1, replace the wild-type sequence in the lambda5/14.1 functional gene. These findings indicate that gene conversion events play a major role in generating diversity that could affect stability or expression of the pre-B-cell receptor complex. PMID- 10527693 TI - Lymphocyte activation in angina pectoris. AB - To determine the role of immune responses in the etiology of coronary angioplasty, the distribution of blood lymphocytes and levels of soluble immune factors in sera of patients with primary unstable angina were determined at pre and post coronary angioplasty. Our data showed (1) an increase in the numbers of lymphocytes bearing lymphocyte activating gene-3 (LAG-3) and CD40 in the blood and (2) an increase in levels of sIL2-R and sVCAM-1 in the sera of patients with unstable angina, compared with normal subjects. In contrast, there were no changes in these values in blood or sera of patients shortly after coronary angioplasty. However, levels of sCD8 in the sera of patients, which were similar to those of normal subjects, significantly increased post coronary angioplasty. These results indicate that peripheral blood lymphocytes of patients with unstable angina are immunologically activated and are producing soluble factors which may allow their interaction with endothelial cells in areas of inflammation. This may play a role in antigen presentation and T-B cell interactions which can lead to potentiation of heart disease. PMID- 10527694 TI - Enhancement of the contact hypersensitivity reaction by acute morphine administration at the elicitation phase. AB - The present study investigated the effects of morphine on the irritant contact sensitivity (ICS) and contact hypersensitivity (CHS) reaction. ICS was induced by croton oil application on the pinnae of naive rats. Morphine injected prior to croton oil application did not affect the ICS response when assessed by measurements of pinnae thickness. CHS was induced by applying the antigen 2,4 dinitro-1-fluorobenzene (DNFB) to the pinnae of rats sensitized to DNFB. Rats received an injection of morphine prior to either initial antigen exposure (sensitization) or antigen reexposure (challenge). Morphine prior to challenge, but not sensitization, resulted in a pronounced enhancement of the CHS response as measured by pinna thickness. Quantitative PCR also showed increased IFN-gamma mRNA levels in the inflamed tissue of morphine-treated rats. Naltrexone blocked the morphine-induced enhancement of the CHS response. The differential effects of morphine suggest that opioids have a more pronounced effect on in vivo immune responses that involve immunological memory. PMID- 10527696 TI - Letter to the editor PMID- 10527695 TI - Letter to the editor regarding the use of intravenous immunoglobulin (IVIG) in asthma. PMID- 10527697 TI - Enhanced apoptosis mediates inhibition of EBV-transformed lymphoblastoid cell line proliferation by curcumin. AB - BACKGROUND: Epstein-Barr virus (EBV)-associated B-cell lymphomas occur more frequently in immunodeficient states such as organ transplantation and HIV infection. We have previously reported that B cell immortalization with EBV was promoted by cyclosporin A (CyA) and that curcumin (Cur), a natural phenol with known antioxidant and antitumor properties, blocked EBV-induced B cell immortalization. In the following experiments we show that Cur inhibits the proliferation of EBV-transformed lymphoblastoid cell lines (LCL) via enhanced apoptosis. METHODS: LCL were generated by infecting freshly isolated human B cells with EBV (B95-8) for 12 h and coculturing with predetermined optimal concentrations of CyA (500 ng/ml) for 4 weeks. LCL were then either frozen for future use or propagated for immediate experiments. These cells were then plated in 96-well plates with 20 microM Cur or 0.1% DMSO (vehicle control). The number of immortalized colonies/well, cell count, and (3)H uptake were used as an index of immortalization. To assess apoptosis rate LCL were cultured with 0.1% DMSO or Cur (20 microM) for 0, 18, and 42 h in culture flasks and then stained with MC540 and H33342, as markers for apoptosis, and analyzed by FACS. RESULTS: A profound inhibition of proliferation was seen in the LCL with 20 microM curcumin compared to 0.1% DMSO control. The colony count reduced from 34.5 +/- 3.4 to 0/well (P = 0.005), cell number reduced from 101,250 +/- 12,093 to 3750 +/- 1500/well (P = 0.002), and (3)H uptake reduced from 40,889 +/- 3669 to 70 +/- 5.2/well (P = 0.001). The apoptosis rate of LCL in the DMSO control at 24.07 and 16.87% increased significantly with 20 microM Cur to 76.4 and 95.1% at 18 and 42 h, respectively (P = 0.02). CONCLUSION: Cur is a potent inhibitor of EBV-transformed LCL. This effect appears to be mediated through enhanced apoptosis. A further investigation of this effect may be useful in prevention and therapy of B-cell lymphoma in immunodeficient patients. PMID- 10527698 TI - Effects of anastomosis of tissue-engineered neointestine to native small bowel. AB - BACKGROUND: Our laboratory is investigating the tissue engineering of small intestine using intestinal epithelial organoid units seeded onto highly porous biodegradable polymer matrices. This study investigated the effects of anastomosis of tissue-engineered intestine to native small bowel alone or combined with small bowel resection on neointestinal regeneration. METHODS: Intestinal epithelial organoid units harvested from neonatal Lewis rats were seeded onto biodegradable polymer tubes and implanted into the omentum of adult Lewis rats as follows: (1) implantation alone (n = 9); (2) implantation followed by anastomosis to native small bowel at 3 weeks (n = 11); and (3) implantation after small bowel resection and anastomosis to native small bowel at 3 weeks (n = 8). All constructs were harvested at 10 weeks and examined by histology. Morphometric analysis of the neomucosa was obtained using a computer image analysis program. RESULTS: Cyst development was noted in all animals. All anastomoses were patent at 10 weeks. Histology revealed the development of a vascularized tissue with a neomucosa lining the lumen of the cyst with invaginations resembling crypt-villus structures. Morphometric analysis demonstrated significantly greater villus number, villus height, crypt number, crypt area, and mucosal surface length in groups 2 and 3 compared with group 1, and significantly greater villus number, villus height, crypt area, and mucosal surface length in group 3 compared with group 2 (P < 0.05, ANOVA, Tukey test). CONCLUSION: Intestinal epithelial organoid units transplanted on biodegradable polymer tubes can regenerate into complex tissue resembling small intestine. Anastomosis to native small bowel combined with small bowel resection and anastomosis alone contribute significant regenerative stimuli for the morphogenesis and differentiation of tissue-engineered neointestine. PMID- 10527699 TI - Cyclopentyladenosine improves cell proliferation, wound healing, and hair growth. AB - BACKGROUND: N(6)-Cyclopentyladenosine (CPA), a structural analog of adenosine, is a vasodilator with extensive pharmacological effects. However, little is known about the effect of CPA on wound healing and hair growth. METHODS: Cellular responses to CPA were measured in vitro by tetrazolium dye reduction and in vivo by bromodeoxyuridine (BrdU) uptake. The effect of CPA on healing of incisional and excisional wounds on the dorsum of diabetic (db/db, n = 94) and nondiabetic (db/+, n = 20) mice and hair growth along the wound margin was evaluated with wound breaking strength, wound closure rate, and quantitative histology. RESULTS: CPA stimulated proliferation of BALB/3T3 fibroblasts and human dermal microvascular endothelial cells in both quiescent and nonquiescent phases. Wounds treated with CPA at 10 microM showed a significant increase in the number of BrdU labeled cells, including keratinocytes, fibroblasts, endothelial cells, and cells in sebaceous glands and the outer root sheath of hair follicles, compared with controls (P < 0.05). CPA application (5.1 microg/daily for 12 days) significantly increased the breaking strength of incisional wounds at day 24 postwound (P < 0.05). Excisional wound closure rate in the CPA-treated group (3.4 microg/daily for 15 days) was accelerated starting at day 10 postwound compared with controls (P < 0.01). Tissue sections from CPA-treated wounds showed a sevenfold increase in hair follicle number, compared with controls (P < 0.01). Enhanced hair growth along the wound margin was revealed in CPA-treated groups. CONCLUSION: CPA stimulated proliferation of many cell types in vivo and in vitro and enhanced wound healing and hair growth. Therefore, CPA could be an interesting candidate for clinical application. PMID- 10527700 TI - ICAM-1 affects reperfusion injury and graft function after cardiac transplantation. AB - BACKGROUND: The effects of increased expression of intercellular adhesion molecule (ICAM-1), an important mediator of neutrophil-mediated reperfusion injury (RI), were assessed in donor cardiac allografts in a heterotopic rat transplantation model. METHODS: At -24 h, PVG donors were untreated (n = 35) or treated (n = 37) with lipopolysaccharide (LPS, 5 mg/kg ip). Hearts were procured at 0 h, stored at 4 degrees C for 45 min, and grafted heterotopically into ACI recipients pretreated with vehicle or anti-ICAM-1 (1A29) mAb. Intracardiac balloons (n = 8 per group) were used to measure allograft left ventricular function (dP/dt) prior to harvest and following reperfusion. RI was assessed at 6, 12, and 24 h by myeloperoxidase (MPO) levels, percentage wet weight (%w/w), and percentage contraction band necrosis (%CBN). RESULTS: At 12 h, LPS-pretreated grafts showed increased ICAM-1 expression by Northern blot (n = 3) and immunohistochemistry (n = 3) and significantly increased MPO (0.33 +/- 0.2 U/mg vs 0.05 +/- 0.04 U/mg at 12 h), %w/w (81.7 +/- 1.8% vs 79.2 +/- 0.7% at 12 h), and %CBN (15.2 +/- 1. 9% vs 11.4 +/- 2.0% at 24 h). LPS pretreatment had no effect on graft function at early time points (baseline to 2 h) but led to depressed dP/dt at later time points with trends toward significance at 12 h (2101 +/- 1653 mmHg/s vs 173 +/- 201 mmHg/s, P = 0.06, ANOVA). Recipient 1A29 treatment (n = 6 per group) reversed the effects of LPS pretreatment in all three RI parameters and significantly improved functional recovery. CONCLUSIONS: Alteration of cardiac graft phenotype to that likely seen in clinical organ donors leads to increased delayed-onset myocardial RI following transplantation in this model. The blockade of this increased RI following 1A29 mAb treatment supports a central role for ICAM-1 in this process. PMID- 10527701 TI - In vivo effects of monoclonal antibodies against rat beta(2) integrins on kidney ischemia-reperfusion injury. AB - BACKGROUND: Ischemia-reperfusion (IR) involves adhesion of leukocytes to the activated endothelium, leading to tissue damage. CD11/CD18 beta(2) integrins interact with their ligands on endothelial cells and may therefore represent a therapeutic target for the prevention of IR. We investigated the effects of three monoclonal antibodies (mAbs) that recognize epitopes of heavy or light chain of the beta(2) integrins on IR in kidneys. METHODS: Uninephrectomized Fischer rats were subjected to 45 or 60 min of renal ischemia, treated with intravenously anti beta(2) integrin monoclonal antibodies (anti-CD11a, anti-CD11b, and anti-CD18) 5 min prior to reperfusion, and compared to a nontreated group. Serum creatinine, blood urea nitrogen (BUN), and kidney histopathological damages were assessed at 1, 2, and 7 days after ischemia. RESULTS: After 45 and 60 min of ischemia, serum creatinine and BUN were significantly higher in the control than in animals treated with anti-CD11a and anti-CD18 at 24 and 48 h. Administration of anti CD11b had a beneficial effect on renal function after 45 min but not after 60 min of ischemia. Histologic and immunostaining studies demonstrated mild tubular necrosis and less leukocyte infiltration in the anti-CD11a- and anti-CD18-treated groups compared to the control group. CONCLUSION: These results indicate that selected antibodies to CD11a/CD18 may decrease kidney IR injury when administered prior to reperfusion. PMID- 10527702 TI - Calcium antagonists improve cardiac mechanical performance after thermal trauma. AB - Burn trauma initiates a pathophysiologic cascade, which includes cardiac dysfunction and intramyocyte calcium accumulation. This study examined the hypothesis that therapeutic interventions which limit intracellular cardiac Ca(2+) accumulation after burn trauma will improve cardiac function. Guinea pigs were anesthetized (methoxyflurane), burned over 43% of total body surface area, and fluid resuscitated (FR) for 24 h. Burn guinea pigs were randomly divided into three groups: Group 1, FR alone, Group 2, FR plus dantrolene (10 mg/kg body wt, IV, 30 min, 8 and 22 h postburn), a drug which inhibits the Ca(2+) release channel (ryanodine receptor) of the cardiac sarcoplasmic reticulum, and Group 3, FR plus diltiazem (0.20-0.22 mg/kg given IV as a slow infusion over 6 h postburn), a drug which specifically blocks Ca(2+) slow channels; sham burn guinea pigs were given vehicle (Group 4), dantrolene (Group 5), or diltiazem (Group 6) as described above (respective controls). Cardiac dysfunction was impaired in fluid-treated burns (Group 1) compared to sham burns (Group 4) as indicated by reduced developed left ventricular pressure (LVP) (86 +/- 2 vs 52 +/ 3 mm Hg, P < 0.05), rate of LVP rise, (+dP/dt max, 1379 +/- 64 vs 909 +/- 44 mm Hg/s, P < 0.05), and LVP fall (-dP/dt max, 1184 +/- 31 vs 881 +/- 40 mm Hg/s, P < 0.05), and time to peak pressure (110 +/- 2 vs 102 +/- 2 ms, P < 0.05). In addition, [Ca(2+)](i) rose in cardiomyocytes harvested from fluid-treated burns (Group 1, 307 +/- 29 nM) compared to vehicle-treated controls (Group 4, 152 +/- 6 nM, P < 0.05). Neither calcium antagonist altered ventricular function or [Ca(2+)](i) in sham burns (Groups 5 and 6). In contrast, antagonists given after burn injury reduced cardiomyocyte [Ca(2+)](i) (Group 2, dantrolene-treated burns: 196 +/- 8 nM, and Group 3, diltiazem treated burns: 216 +/- 8 nM) and improved cardiac performance compared to that measured in burns given FR alone. Our data suggest that calcium antagonists given after burn trauma restored intracellular Ca(2+) homeostasis, decreased cardiac cell injury, and improved cardiac contractile function. PMID- 10527703 TI - Exposure to thrombus diminishes endothelial derived relaxation in the rabbit carotid artery. AB - PURPOSE: Thrombus is believed to be deleterious to intimal function. However, few studies have directly examined this effect. This study examines the effect of thrombus on endothelial-dependent and -independent vasorelaxation in the rabbit carotid artery. METHODS: Twelve male New Zealand white rabbits (3.5-4.5 kg) were divided into two groups of six. Thrombosis was induced in group I by segmental right carotid artery ligation. Group II underwent segmental right carotid ligation immediately followed by removal of thrombus with normal saline flush through an arteriotomy. The left carotid arteries were exposed in both groups and served as internal controls. After 4 h, left and right carotid arteries were harvested, sectioned into 6-mm rings, and mounted on isometric force transducers in a physiologic bath. Thrombus was removed from the arteries in group I during the ring preparation process. Neither group I nor group II had thrombus in contact with endothelium during ex vivo testing. The arterial rings were constricted with norepinephrine (1 x 10(-4) M). Endothelium-dependent and independent vasorelaxation to acetylcholine (Ach) and s nitrosoacetylpenicillamine, respectively, were measured in a dose-response manner. Results were expressed as a percentage of vasorelaxation. Statistical analysis was performed using an analysis of variance. RESULTS: Endothelial dependent vasorelaxation, which tests for endothelial cell function, was decreased in the thrombus and endothelial ischemia group (I) compared to control as noted by vasorelaxations of 22% vs 34% at 1 x 10(-4) molar concentration Ach, and 33% vs 48% at 1 x 10(-3) molar concentration Ach, respectively (P = 0.05). By comparison, there was no difference in the endothelial-dependent vasorelaxation of the endothelial ischemia group (II) versus control. Endothelial-independent vasorelaxation, which tests for smooth muscle function, was not affected by either the thrombus and endothelial ischemia group (I) or the endothelial ischemia group (II) compared to the control group. The controls in group I and group II were slightly different. When this difference was removed, the resulting comparison of treatments in group I and group II approached significance at molar concentrations of 1 x 10(-4), 1 x 10(-5), and 1 x 10(-6) (P = 0.07, 0.06, 0.06). CONCLUSIONS: The presence of thrombus within the rabbit carotid artery for a period of 4 h decreases endothelial-dependent relaxation. Four hours of endothelial ischemia without thrombus did not change endothelial-dependent vasorelaxation. Neither thrombus nor ischemia alone had any effect on the endothelium-independent vasorelaxation. We conclude that thrombus is deleterious to endothelial function independent of smooth muscle function in the acute setting as measured by endothelial-dependent vasorelaxation. PMID- 10527704 TI - Gelatin-sealed polyester resists Staphylococcus epidermidis biofilm infection. AB - BACKGROUND: The purpose of this study was to show that gelatin-impregnated polyester grafts inhibit Staphylococcus epidermidis biofilm infection in a canine model of aortic graft interposition. A clinically native species and two engineered strains, which differed in slime and adhesin antigen components, were compared to determine differential gelatin and slime interactions. METHODS: In vitro bacterial graft colonization was validated by immersion of graft segments in inoculating solutions (10(6) colony forming units/ml) of a clinically native species RP62A and two genetically engineered S. epidermidis species, M187sn3 (SN3: slime and adhesin negative) or M187sp11 (SP11: slime and adhesin positive), for 18 h at 23 degrees C. The grafts were washed, sonicated, and cultured to assess in vitro bacterial graft adherence. Grafts similarly inoculated were placed as aortic interposition grafts in dogs. Three sterile grafts were implanted as controls. Grafts were excised after 6 weeks and cultured for bacterial growth as in the in vitro study. Infection was defined by a positive culture in the excised grafts. Data were analyzed with nonparametric statistical methods. RESULTS: In vitro bacterial graft adherence in colony forming units per milliliter was similar at 18 h postsonication for RP62A (8 x 10(4) +/- 1 x 10(4)), SN3 (7 x 10(4) +/- 2 x 10(4)), and SP11 (6 x 10(4) +/- 2 x 10(4)) (P = NS). Only one of five grafts inoculated with RP62A was culture positive after 6 weeks. No grafts inoculated with the engineered strains SN3 or SP11 were culture positive after explanation. CONCLUSION: In vitro bacterial inoculation of gelatin impregnated polyester was similar among the species and not dependent upon the presence of slime and adhesin components. Gelatin-impregnated polyester grafts demonstrated in vivo resistance to coagulase-negative staphylococcal biofilm infection. PMID- 10527705 TI - Alterations in chemokine mRNA expression in animals receiving portal vein immunization and renal allo- or xenotransplantation precede altered cytokine production. AB - We have analyzed chemokine mRNA expression in graft tissue of C3H/HEJ mice receiving allogeneic (C57BL/6) or xenogeneic [Lewis (LEW) rat donors] kidney grafts and correlated this with graft survival. Since donor-specific portal vein (pv) immunization is known to increase allo- and xenograft survival, in some cases recipients also received pretransplant pv or intravenous (iv) immunization; other animals received the antioxidant N-acetylcysteine (NAc) to examine the role of ischemia/reperfusion injury in the changes observed. Graft tissue and lymph nodes draining the respective grafts were obtained at various times posttransplantation and used for quantitative polymerase chain reaction analysis of mRNAs for different chemokines. In addition, lymphocytes were restimulated in culture with donor antigen and supernatants assayed for different cytokines. We observed that early increases in mRNA for MCP-1 preceded a polarization to type 2 cytokine production. Infusion of NAc twice daily for 4 days following transplantation further altered chemokine mRNA expression (increased MCP-1 and RANTES; decreased CINC); led to more enhanced type 2 cytokine production relative to control animals; and further increased xenograft survival. By use of heteroantibodies to different chemokines, anti-MCP-1 alone, but not antibodies to MIP-1alpha or RANTES, abolished this early polarization in cytokine production, implying a causal link between MCP-1 production and polarization in cytokine production. We conclude that manipulation of chemokine production early after transplantation might indirectly modify graft outcome by modifying cytokine production. PMID- 10527706 TI - Effect of intracerebroventricular injection of tumor necrosis factor alpha on gut mucosal protein turnover in mice fed enterally. AB - OBJECTIVE: The objective of this study was to determine whether mucosal protein turnover is enhanced by intracerebroventricular (ICV) injection of tumor necrosis factor alpha (TNF-alpha) in mice fed enterally. METHODS: Male B6C3F1 mice (n = 37, 20.0-25.0 g) were catheterized in the ICV space using stereotaxic coordination, and subsequently a catheter was inserted into the stomach for enteral feeding on Day 0. The animals were fed a standard mouse diet and water ad libitum for 4 days. On Day 4, enteral feeding was begun with a standard liquid diet, and either artificial cerebrospinal fluid (control) or recombinant human (Rh) TNF-alpha (5 ng/h) was continuously given via the ICV catheter using an osmotic pump. On either Day 5 or 7, the mice were sacrificed 10 min after injection of [U-(14)C]phenylalanine (5 microCi/mouse, ip). Fractional synthetic rates (FSRs) of jejunal mucosa and muscle were measured by the pool flooding technique. Interleukin-6 (IL-6) levels in the plasma were measured by ELISA. RESULTS: Body weight was significantly reduced by ICV injection of TNF-alpha, because muscle FSR was decreased with TNF-alpha. There were no differences in IL 6 levels in the plasma between the two groups. The FSR of jejunal mucosa was elevated by infusion of TNF-alpha as compared with control mice on both Days 5 and 7 [Day 5: (control) 99.5 +/- 17. 6%/day vs (TNF) 150.5 +/- 20.5%/day, P < 0.05; Day 7: (control) 54.8 +/- 8.8%/day vs (TNF) 102.7 +/- 19.1%/day, P < 0.05). CONCLUSION: injection of TNF-alpha enhanced the mucosal protein synthesis rate without elevating IL-6 levels as compared with control mice, suggesting that the central nervous system links to the gut to regulate mucosal protein turnover through cytokine in the brain, but plasma IL-6 is not involved in this linkage. PMID- 10527707 TI - Leukocyte-endothelium interaction in arterioles after ischemia and reperfusion. AB - Leukocyte-endothelium interaction in postcapillary venules plays an important role in reperfusion injury, inflammation, shock, and sepsis. This phenomenon is poorly described in precapillary arterioles. In fact, many researchers have reported no evidence of leukocyte adherence in arterioles whatsoever. Most research has focused on venules of larger rodents, in which observation of the microcirculation, especially arterioles, is limited. We have developed a model which provides a clearer view of these microvessels using the mouse cremaster muscle. This muscle has an approximate thickness of 100 microm allowing images produced by transillumination to be very clear. After vascular isolation, the right cremaster muscle was subjected to 4 h of ischemia, followed by 2 h of reperfusion. The left muscle was not rendered ischemic, thereby allowing it to serve as the animal's own internal control. We observed leukocyte rolling in arterioles in both the ischemic and the nonischemic muscles. Leukocyte sticking was seen in arterioles and venules on both sides, except in control arterioles. The number of rolling and sticking leukocytes on the ischemic side was significantly higher than in controls (P < 0.05) for both arterioles and venules. During the reperfusion period, this number did not change significantly. Transmigration of leukocytes was observed only in venules, but not in arterioles. The number of perfused capillaries was reduced on the ischemic side compared to controls and did not change significantly during the 2 h of reperfusion. Our results demonstrate that leukocyte-endothelium interaction occurs in muscle arterioles of mice. This phenomenon is more pronounced after ischemia and reperfusion, i.e., depends on the extent of tissue insult. PMID- 10527708 TI - Confirmation of translocated gastrointestinal bacteria in a neonatal model. AB - PURPOSE: The hypothesis that enteric bacteria translocate from the gastrointestinal (GI) tract to extraintestinal sites has been extensively studied. However, definitive evidence that spontaneous bacterial translocation and dissemination from the GI tract to extraintestinal sites occur in a neonatal model has been lacking. The aim of this study was to confirm this phenomenon by tracking enterally administered, plasmid-labeled bacteria to extraintestinal sites. MATERIALS AND METHODS: Escherichia coli 07:K1 (E. coli K1) with and without a nontransferable, ampicillin resistance plasmid (pGEM-7) were used in this study. Newborn New Zealand white rabbit pups were separated into three treatment groups: transformed E. coli K1 (E. coli K1 + pGEM-7, n = 20), nontransformed E. coli K1 (n = 12), and control pups (no bacteria, n = 7). Pups were enterally fed 10% Formulac solution supplemented with a suspension of bacteria respective to their group. After the pups fed twice daily for 2 days, representative tissue specimens from the small bowel (SB), mesenteric lymph nodes (MLNs), spleen (SPL), and liver (LIV) were aseptically harvested and tested for culture growth in ampicillin-supplemented medium. RESULTS: Positive growths of plasmid-induced ampicillin-resistant bacteria were detected in tissue specimens harvested from rabbits fed transformed E. coli K1, but were not detected in the other groups. CONCLUSION: This experiment demonstrated conclusively that transformed E. coli K1 fed to healthy rabbit pups spontaneously translocated from the intestinal lumen and subsequently disseminated to the mesenteric lymph nodes, spleen, and liver. PMID- 10527709 TI - Roles of platelet-activating factor, interleukin-1beta and interleukin-6 in intestinal barrier dysfunction induced by mesenteric arterial ischemia and reperfusion. AB - BACKGROUND: Platelet-activating factor (PAF), cytokines, proteases, and other factors are probably involved in the development of gut barrier dysfunction following intestinal ischemia and reperfusion (I/R), although the act underlying pathophysiological mechanisms has not yet been fully clarified. The aim of the present study was to clarify the relationship of intestinal barrier integrity to systemic levels of interleukin-1beta, interleukin-6, and protease inhibitor levels and local leukocyte accumulation in a rat model of intestinal ischemia for 40 min followed by 3 or 12 h reperfusion, with or without treatment with a PAF inhibitor. METHODS: Myeloperoxidase (MPO) content in the small intestinal mucosa, serum levels of interleukin-1beta and -6, and plasma protease inhibitors, and intestinal endothelial and epithelial permeability were assessed, with or without treatment with the PAF antagonist lexipafant. RESULTS: Intestinal I/R resulted in intestinal barrier dysfunction with pronounced plasma leakage to the intestinal lumen, the leakage being aggravated following a longer reperfusion period. Proteolytic plasma activity was evident by low levels of the plasma protease inhibitors measured. MPO content increased significantly after I/R, as did serum levels of interleukin-1beta and -6, without difference between the two periods of reperfusion. Treatment with the PAF inhibitor lexipafant partly, though not fully, restored the changes caused by I/R. CONCLUSION: PAF seems to be involved in the release of cytokines, such as interleukin-1 and -6, consumption of protease inhibitors, and impaired intestinal barrier integrity seen following intestinal I/R. Treatment with a PAF antagonist was effective in restoring the changes caused by intestinal I/R, though not reaching complete normal levels. PMID- 10527710 TI - Effects of angiotensin type-I receptor blockade on pericardial fibrosis. AB - BACKGROUND: Reoperative cardiac surgical procedures are associated with a significantly greater complication rate than that of the initial procedure. Enhanced collagen synthesis can occur due to increased production of angiotensin II (Ang-II) and subsequent activation of Ang AT(1) receptor. Accordingly, the goal of the current study is to test the hypothesis that increased Ang AT(1) receptor activity following pericardiotomy contributes to pericardial thickening and fibrosis. MATERIALS AND METHODS: Adult pigs were randomly assigned to three protocols: (1) pericardiotomy with 28-day recovery period (n = 5); (2) pericardiotomy with Ang AT(1) receptor blockade instituted throughout the 28-day recovery period using 60 mg/day valsartan (n = 5); and (3) sham controls (n = 6). Pericardium samples were collected and analyzed by biochemical and histomorphometrical methods. Pericardial fibrosis occurred postpericardiotomy as indicated by increased hydroxyproline content from normal value of 50 +/- 3 microg/mg to 75 +/- 4 microg/mg (P < 0. 05). RESULTS: Pericardial thickness was increased postpericardiotomy to 2.7 +/- 0.4 mm compared to normal values of 0.4 +/- 0.05 mm (P < 0.05). Ang AT(1) receptor blockade reduced pericardial thickness by 50% and the relative degree of fibrosis was comparable to that of the normal group. CONCLUSIONS: The results from this pericardial fibrosis animal model suggest that Ang AT(1) receptor activation contributes to the development of pericardial thickening and collagen accumulation in the postoperative period. Thus, Ang AT(1) receptor inhibition may provide a novel therapeutic strategy to prevent pericardial fibrosis that follows cardiac surgical procedures. PMID- 10527711 TI - Antiangiogenic effects of the oral administration of liquid cartilage extract in humans. AB - BACKGROUND: The antiangiogenic properties of shark cartilage extracts have been demonstrated in animal models but there are no data in human subjects. MATERIALS AND METHODS: A placebo or one of two doses of a liquid shark cartilage extract was orally administered daily, from Day 1 to Day 23 of the study protocol, to 29 healthy male volunteers randomized into three groups. On Day 12, a polyvinyl alcohol sponge threaded in a perforated silicone tubing was inserted subcutaneously on the anterior side of the arm and removed on Day 23. Evaluation of endothelial cell density, with factor VIII immunostaining, an indirect measurement of angiogenesis, was performed on histological sections of the implant using a semiquantitative numerical scale ranging from 1 (low density) to 5 (high density). The hydroxyproline content of the sponges was measured by HPLC. RESULTS: The mean endothelial cell density was significantly lower in groups that had received the liquid cartilage extract: grades 2.24 +/- 0.10, 2.47 +/- 0.10, and 3.15 +/- 0.11 for 7 and 21 ml liquid cartilage extract and placebo, respectively (P < 0.01 for both comparisons). No grade 1 was observed in the placebo group, whereas 9 treated subjects received a grade 1. Hydroxyproline content of the sponges did not differ between groups and there was no significant correlation between hydroxyproline content and endothelial cell density in the sponges. CONCLUSIONS: These results demonstrate that the liquid cartilage extract contains an antiangiogenic component bioavailable in humans by oral administration. This is the first report of an inhibition of wound angiogenesis in healthy men. PMID- 10527712 TI - Radiofrequency ablation lesions in a pig liver model. AB - BACKGROUND: Radiofrequency (RF) ablation has been reported as a means of liver tumor destruction. This study evaluates the use of ultrasound monitoring of radiofrequency lesion creation and describes the morphology, histologic characteristics, and vascular effects of radiofrequency ablations in a pig liver model. MATERIALS AND METHODS: Hemodynamic monitoring was established and laparotomies were performed in 50-kg pigs. Under ultrasound guidance, radiofrequency needle probes were placed in the liver at predetermined locations. Radiofrequency energy was applied over 15 min to generate lesions 3 cm in diameter. Eighty lesions were generated in 10 animals. At the completion of the experiment, the lesions were examined with ultrasound and then excised for CT, gross, and histologic examination. RESULTS: There were no adverse systemic effects. Ultrasound imaging demonstrated the size, shape, and position of the lesions. Gross examination demonstrated a core of ablated tissue with a surrounding 1- to 2-mm hemorrhagic perimeter. Lesion volumes averaged 12.8 cc(3) (range 5-34 cc(3)). Final lesion shape and size were frequently altered by the cooling effect of local blood flow. Histologic stains demonstrated microvascular thrombosis and coagulative necrosis within the lesions. There appeared to be 100% cellular destruction within the lesion by cytochemical staining. CONCLUSIONS: We demonstrated that RF ablation is capable of killing large volumes of normal liver tissue; however, local vasculature plays a significant a role in defining the ultimate size and shape of the lesion created. This may interfere with the utility of radiofrequency ablation as a modality for local tumor control. PMID- 10527713 TI - Dose-dependent limitation of arterial enlargement by the matrix metalloproteinase inhibitor RS-113,456. AB - BACKGROUND: Arterial diameter changes in response to flow. Chronic flow-mediated arterial enlargement may be mediated through metalloproteinase activity in the extracellular matrix of the arterial wall. We examined flow-mediated enlargement in the setting of increasing competitive matrix metalloproteinase (MMP) inhibition and with respect to gelatinase A and B expression and activity. METHODS: Left common femoral arteriovenous fistulas (AVFs) were created in dose response (52) and time course (34) cohorts of rats. Dose-response rats received either vehicle alone or 12.5, 25, or 37. 5 mg/kg b.i.d. RS 113,456, a competitive MMP inhibitor. Heart rate, blood pressure, and weight were measured at intervals following AVF construction. Aortic and common iliac diameters were measured on postoperative day (POD) 21. Untreated time course rats were sacrificed on PODs 0 (no AVF), 3, 7, 14, and 21. Aortic diameter was measured and the vessels were harvested for tissue analysis. Equal amounts of aortic RNA underwent reverse transcription and polymerase chain reaction with primers for MMP-2, MMP-9, and GAPDH. Zymography was performed on iliac artery tissue to measure gelatinolytic activity. RESULTS: A significant, stepwise reduction in flow-mediated aortic and left common iliac enlargement following left femoral AVF creation was noted with progressively higher doses of RS 113,456 without apparent hemodynamic or toxic effects. Right common iliac diameter was unchanged. Over 21 days following AVF creation, there was an upward trend in expression and activity for MMP-2 not evident for MMP-9. CONCLUSION: Flow-mediated arterial enlargement is limited by competitive MMP inhibition in a dose-dependent fashion. MMP-dependent flow mediated enlargement may involve differential expression and activity of MMP-2 and MMP-9. PMID- 10527714 TI - mRNA levels of dipeptidyl peptidase IV decrease during intestinal adaptation. AB - BACKGROUND: Glucagon-like peptide 2 (GLP-2) has recently been shown to be a potent enterotrophic factor that may mediate mucosal hyperplasia during intestinal adaptation. The intestinal brush-border protease dipeptidyl peptidase IV (DPP IV) cleaves GLP-2 to an inactive form. It has been postulated that DPP IV activity limits the enterotrophic activity of GLP-2 in rats and humans. Massive small bowel resection (MSBR) in rats is an animal model of intestinal adaptation that has been used successfully to characterize factors involved in the modulation of adaptation. METHODS: Total RNA was extracted from normal terminal ileum or terminal ileum post-MSBR from Sprague-Dawley rats which were sacrificed 2, 4, and 7 days postresection. A partial rat DPP IV clone was isolated by reverse transcription polymerase chain reaction, and Northern blot analysis of rat DPP IV mRNA levels in normal small bowel and small bowel post-MSBR was performed. RESULTS: Within normal small bowel, DPP IV mRNA levels were greatest in the terminal ileum; levels in the duodenum and jejunum were approximately 50% of those in the terminal ileum. DPP IV mRNA levels decreased in terminal ileum post-MSBR 2, 4, and 7 days after resection. CONCLUSION: The decreased DPPIV gene expression suggests a novel mechanism by which the effects on mucosal growth of GLP-2 may be further enhanced, and further that GLP-2 may be a more useful therapeutic agent in humans than currently anticipated. PMID- 10527715 TI - Transient exposure to tumor necrosis factor-alpha inhibits collagen accumulation by cultured hypertrophic scar fibroblasts. AB - BACKGROUND: Hypertrophic scars (HS) are frequent consequences of deep dermal injury, such as deep partial-thickness burns and abrasions, and are characterized by overproduction of collagen. In vitro studies have shown that cultured HS fibroblasts produce elevated levels of collagen and insulin-like growth factor binding protein 3 (IGFBP-3). Additionally, histological studies have indicated HS contain fewer tumor necrosis factor alpha (TNF-alpha)-positive infiltrating cells and express lower levels of TNF-alpha mRNA, suggesting TNF-alpha, which can inhibit collagen expression in some systems, may function to deactivate the wound healing process in scars. HS also exhibit increased levels of transforming growth factor beta (TGF-beta), a factor that stimulates collagen and extracellular matrix deposition by fibroblasts and also stimulates IGFBP-3 expression. In some systems, IGFBP-3 mediates the effects of TGF-beta. The present study sought to determine the effects of continuous and transient TNF-alpha exposure on collagen and IGFBP-3 expression by cultured HS fibroblasts and to investigate the role of IGFBP-3 in collagen accretion by HS fibroblasts. MATERIALS AND METHODS: Superficial and deep dermal HS fibroblasts from four patients were cultured. Fibroblasts were cultured in serum-free medium and exposed to 0-2 ng/ml TNF-alpha for 0, 1, 4, or 72 h. After 72 h of culture, medium samples were processed for Western blot analysis of type I collagen accumulation or for ligand blot analysis of IGFBP-3 accumulation. The effects of an anti-IGFBP-3 neutralizing antibody on collagen accumulation were also assessed. RESULTS: Treatment of superficial and deep HS fibroblasts with TNF-alpha resulted in dose-dependent decreases in accumulation of both type I collagen and IGFBP-3 in the culture medium (P < 0.01). However, using the anti-IGFBP-3 neutralizing antibody, a causal relationship between decreased IGFBP-3 and decreased collagen accumulation could not be demonstrated. Transient exposure of cultured HS fibroblasts to TNF-alpha for as little as 1 h was as effective as continuous exposure to TNF-alpha for 72 h in inhibiting collagen accumulation. CONCLUSIONS: These results support the hypothesis that TNF-alpha functions as a wound healing deactivation signal that is deficient in HS. Although TNF-alpha inhibited accretion of both collagen and IGFBP-3, the role of IGFBP-3 in HS remains unresolved. This study suggests that transient TNF-alpha exposure may be used to inhibit collagen overaccumulation in HS and that the timing of TNF-alpha exposure following dermal injury may not be critical for this inhibition. PMID- 10527716 TI - A dynamic model of the behaviour of sitka spruce in high winds AB - This paper describes the development, verification and use of a mathematical model to describe the dynamic behaviour of typical forest trees in high winds. The model assumes (a) that the trunk can be represented by a vertical tapered cantilever with specified stiffness and mass distributions; (b) that the canopy can be represented by a cylindrical body of a different density at the top of the trunk; (c) that the wind loading can be represented by a spatially constant wind distribution applied to the upper part of the canopy, that is varying in time with realistic spectral properties; (d) that the damping of the oscillations of the tree is caused solely by aerodynamic damping.The resulting complex, fourth order differential equations are solved using numerical methods. This model is used to predict the transfer functions relating tree displacement spectra to wind spectra, and it is shown that the model is able to represent experimental spectra well, particularly with regard to the prediction of the primary natural frequency. Wind speeds for tree failure by both trunk snapping and uprooting are then pictured, and reasonably realistic values obtained. The need for a better understanding of the relationship between extreme and mean wind gusts is apparent. Through a discussion of the sensitivity of the results to variations in the different model parameters some general conclusions are drawn about the effects of the different parameters on tree stability. Copyright 1999 Academic Press. PMID- 10527717 TI - Numerical simulation of motility patterns of the small bowel. II. Comparative pharmacological validation of a mathematical model. AB - A model of a locus of the small bowel, described earlier by the authors (Miftakhov et al., 1999) was validated in a comparison of the results of numerical simulations of pharmacological compounds to their effects in biological studies. The actions of the following four classes of drugs were simulated, those: (i) acting on the sarcoplasmic reticulum, (ii) altering the permeability of L- and T-type Ca(2+)channels on the smooth muscle membrane, (iii) motilides, and (iv) benzodiazepines. The strong qualitative resemblance between the theoretical and experimental results supports the robustness of the model. PMID- 10527718 TI - Facts and fictions regarding post-natal neurogenesis in the developing human cerebral cortex. AB - In a recent paper (Shankle et al., 1998a), post-natal neurogenesis in the human cerebral cortex was discussed. Based on re-calculations of morphometric data from the literature, the authors concluded an average 1.1% monthly increase in post natal cortical neuron number between post-natal months 15-72. The present paper makes clear by discussing four main assumptions done by Shankle et al., i.e. shrinkage of the tissue, morphometric features of the neurons under study, conversion of cell densities per area to number per unit volume and estimation of coefficients of variation, that their final conclusion about an increase in neuron number is unsound. Furthermore, five points are discussed here that Shankle et al. had mentioned in order to demonstrate that the pulse thymidine labeling method is less reliable than some have assumed. The present paper refute these assumptions point by point. Thus, the Shankle et al. paper does not provide scientifically valid evidence of a post-natal neurogenesis in the developing human cerebral cortex. PMID- 10527719 TI - Evidence for nonlinear capacitance in biomembrane channel system. AB - The electrophysiological properties of voltage-dependent anion channels from mitochondrial membrane have been studied in a bilayer membrane system. It was observed that the probability of opening of the membrane channel depends on externally applied voltage and the plot is a bell-shaped curve symmetric around probability axis. A scheme of conformational energy levels under varying externally applied voltage was formulated. Assuming that the probability follows Boltzmann distribution, we arrive at an expression of change in energy containing a separate term identical to the energy of a capacitor. This fact indicates the possibility of existence of an added capacitance due to the channel protein. Further it was shown that the aforesaid channel capacitor could be a function of voltage leading to nonlinearity. We have offered a general method of calculating nonlinear capacitance from the experimental data on opening probability of a membrane channel. In case of voltage-dependent anion channel the voltage dependence of the capacitor has a power 0.786. The results have been interpreted in view of the structural organization of the channel protein in the membrane. Our hypothesis is that the phenomenon of capacitor behaviour is a general one for membrane channels. PMID- 10527720 TI - The continuous prisoner's dilemma: I. linear reactive strategies. AB - We present a general model for the Prisoner's Dilemma in which variable degrees of cooperation are possible, and payoffs are scaled accordingly. We describe a continuous strategy space, and divide this space into strategy families. We derive the payoff function for these families analytically, and study the evolutionary outcome when a wide range of strategies play against each other. Our results show that the initial degree of cooperation offered by a strategy is a decisive factor for evolutionary robustness: the most successful strategies in our model offer full cooperation as an initial move, but thereafter cooperate fully only if their opponent does the same. These strategies gradually raise the stakes when playing a strategy which is initially reticent to cooperate, but differ from the strategies predicted by other continuous models in that they are not only generous, but are also consistently optimistic and uncompromising. PMID- 10527721 TI - The continuous Prisoner's dilemma: II. Linear reactive strategies with noise. AB - We present a general model for the Continuous Prisoner's Dilemma and study the effect of errors. We find that cooperative strategies that can resist invasion by defectors are optimistic (make high initial offers), generous (always offer more cooperation than the partner did in the previous round) and uncompromising (offer full cooperation only if the partner does). A necessary condition for the emergence of cooperation in the continuous Prisoner's Dilemma with noise is b (1 p)>c, where b and c denote, respectively, the benefit and cost of cooperation, while p is the error rate. This relation can be reformulated as an error threshold: cooperation can only emerge if the probability of making a mistake is below a critical value. We note, however, that cooperation in the continuous Prisoner's Dilemma with noise does not seem to be evolutionarily stable: while it is possible to find cooperative strategies that resist invasion by defectors, such cooperators are generally invaded by more cooperative strategies which eventually yield to defectors. Thus, the long-term evolution of the continuous Prisoner's Dilemma is either characterized by unending cycles or by stable polymorphisms of cooperators and defectors. PMID- 10527723 TI - Testing the phylogenetic stability of early tetrapods PMID- 10527722 TI - Is there an epigenetic component in long-term memory? PMID- 10527724 TI - Paul A. Srere (1925-1999) PMID- 10527725 TI - Protein interactions. PMID- 10527726 TI - Characterization of heterologous protein-protein interactions using analytical ultracentrifugation. AB - Methods for quantitative characterization of heterologous protein-protein interactions by means of analytical ultracentrifugation (AUC) include sedimentation equilibrium, tracer sedimentation equilibrium, sedimentation velocity, and analytical band sedimentation. Fundamental principles governing the behavior of macromolecules in a centrifugal field are summarized, and the application of these principles to the interpretation of data obtained from each type of experiment is reviewed. Instrumentation and software for the acquisition and analysis of data obtained from different types of AUC experiments are described. PMID- 10527727 TI - Isothermal titration calorimetry of protein-protein interactions. AB - The interaction of biologicalmacromolecules, whether protein-DNA, antibody antigen, hormone-receptor, etc., illustrates the complexity and diversity of molecular recognition. The importance of such interactions in the immune response, signal transduction cascades, and gene expression cannot be overstated. It is of great interest to determine the nature of the forces that stabilize the interaction. The thermodynamics of association are characterized by the stoichiometry of the interaction (n), the association constant (K(a)), the free energy (DeltaG(b)), enthalpy (DeltaH(b)), entropy (DeltaS(b)), and heat capacity of binding (DeltaC(p)). In combination with structural information, the energetics of binding can provide a complete dissection of the interaction and aid in identifying the most important regions of the interface and the energetic contributions. Various indirect methods (ELISA, RIA, surface plasmon resonance, etc.) are routinely used to characterize biologically important interactions. Here we describe the use of isothermal titration calorimetry (ITC) in the study of protein-protein interactions. ITC is the most quantitative means available for measuring the thermodynamic properties of a protein-protein interaction. ITC measures the binding equilibrium directly by determining the heat evolved on association of a ligand with its binding partner. In a single experiment, the values of the binding constant (K(a)), the stoichiometry (n), and the enthalpy of binding (DeltaH(b)) are determined. The free energy and entropy of binding are determined from the association constant. The temperature dependence of the DeltaH(b) parameter, measured by performing the titration at varying temperatures, describes the DeltaC(p) term. As a practical application of the method, we describe the use of ITC to study the interaction between cytochrome c and two monoclonal antibodies. PMID- 10527728 TI - Quantification of protein-protein interactions using fluorescence polarization. AB - Quantitative determinations of the dissociation constants of biomolecular interactions, in particular protein-protein interactions, are essential for a detailed understanding of the molecular basis of their specificities. Fluorescence spectroscopy is particularly well suited for such studies. This article highlights the theoretical and practical aspects of fluorescence polarization and its application to the study of protein-protein interactions. Consideration is given to the nature of the different types of fluorescence probes available and the probe characteristics appropriate for the system under investigation. Several examples from the literature are discussed that illustrate different practical aspects of the technique applied to diverse systems. PMID- 10527729 TI - Fluorescence fluctuation spectroscopy. AB - The analysis of the intensity fluctuation of a fluorescence signal from a relatively small volume and from a few molecules contains information about the distribution of different species present in the solution and about kinetic parameters of the system. The same information is generally averaged out when the fluorescence experiment is performed in a much larger volume, typically a cuvette experiment. The fundamental reason for this difference is that the fluctuations of the fluorescence signal from a few molecules directly reflect the molecular nature of the matter. Only recently, with the advent of confocal microscopy and two-photon excitation, it has become practical to achieve small excitation volumes in which only a few fluorescent molecules are present. We introduce the concept of fluctuation spectroscopy and highlight some of the technical aspects. We discuss different analysis methods used in fluctuation spectroscopy and evaluate their use for studying protein-protein interactions. PMID- 10527731 TI - Electrophoretic methods for studying protein-protein interactions. AB - Protein-protein interactions are involved in many biological processes ranging from DNA replication, to signal transduction, to metabolism control, to viral assembly. The understanding of those interactions would allow the effective design of new drugs and further manipulation of those interactions. Several useful analytical methods are available for the study of protein-protein binding, and among them, electrophoresis is commonly used. We describe two types of electrophoresis: gel electrophoresis and capillary electrophoresis. Gel electrophoresis is a well-established method used to study protein-protein interactions and includes overlay gel electrophoresis, charge shift method, band shift assay, countermigration electrophoresis, affinophoresis, affinity electrophoresis, rocket immunoelectrophoresis, and crossed immunoelectrophoresis. These techniques are briefly described along with their advantages and limitations. Capillary electrophoresis, on the other hand, is a relatively new method and affinity capillary electrophoresis has demonstrated its value in the measurement of binding constants, the estimation of kinetic rate constants, and the determination of stoichiometry of biomolecular interactions. It offers short analysis time, requires minute amounts of protein samples, usually involves no radiolabeled compounds, and, most importantly, is carried out in solution. We summarize the principles of affinity capillary electrophoresis for studying protein-protein interactions along with current limitations and describe in depth its application to the determination of stoichiometries of tight and weak binding protein-protein interactions. The protocol presented in the experimental section details the use of affinity capillary electrophoresis for the determination of stoichiometry of protein complexes. PMID- 10527730 TI - Exploring biomolecular recognition using optical biosensors. AB - Understanding the basic forces that determine molecular recognition helps to elucidate mechanisms of biological processes and facilitates discovery of innovative biotechnological methods and materials for therapeutics, diagnostics, and separation science. The ability to measure interaction properties of biological macromolecules quantitatively across a wide range of affinity, size, and purity is a growing need of studies aimed at characterizing biomolecular interactions and the structural elements that drive them. Optical biosensors have provided an increasingly impactful technology for such biomolecular interaction analyses. These biosensors record the binding and dissociation of macromolecules in real time by transducing the accumulation of mass of an analyte molecule at the sensor surface coated with ligand molecule into an optical signal. Interactions of analytes and ligands can be analyzed at a microscale and without the need to label either interactant. Sensors enable the detection of bimolecular interaction as well as multimolecular assembly. Most notably, the method is quantitative and kinetic, enabling determination of both steady-state and dynamic parameters of interaction. This article describes the basic methodology of optical biosensors and presents several examples of its use to investigate such biomolecular systems as cytokine growth factor-receptor recognition, coagulation factor assembly, and virus-cell docking. PMID- 10527732 TI - Chromatographic methods to study protein-protein interactions. AB - Proteins and enzymes are now generally thought to be organized within the cell to form clusters in a dynamic and versatile way, and heterologous protein-protein interactions are believed to be involved in virtually all cellular events. Therefore we need appropriate tools to detect and study such interactions. Chromatographic techniques prove to be well suited for this kind of investigation. Real complexes formed between proteins can be studied by classic gel filtration. When enzymes are studied, active enzyme gel chromatography is a useful alternative. A variant of classic gel filtration is gel filtration equilibrium analysis, which is similar to equilibrium dialysis. When the association formed is only dynamic and equilibrates very rapidly, either the Hummel-Dryer method of equilibrium gel filtration or large-zone equilibrium filtration sometimes allows the interactions to be analyzed, both qualitatively and quantitatively. Very often, however, interactions between enzymes and proteins can only be evidenced in vitro in media that mimic the intracellular situation. Immobilized proteins are excellent tools for this type of research. Several examples are indeed known where the immobilization of an enzyme on a solid support does not affect its real properties, but rather changes its environment in such a way that the diffusion becomes limiting. Affinity chromatography using immobilized proteins allows the analysis of heterologous protein-protein interactions, both qualitatively and quantitatively. A useful alternative appears to be affinity electrophoresis. The latter technique, however, is exclusively qualitative. All these techniques are described and illustrated with examples taken from the literature. PMID- 10527733 TI - Substrate channeling. AB - Substrate channeling is the process in which the intermediate produced by one enzyme is transferred to the next enzyme without complete mixing with the bulk phase. This process is equivalent to a microcompartmentation of the intermediate, although classic diffusion occurs simultaneously to varying extents in many of these cases. This microcompartmentation and other factors of channeling provide many potential biological advantages. Extensive examples of channeling can be found in the cited reviews. The choice of methods to detect and characterize substrate channeling depends extensively on the type of enzyme associations involved, the constants of the system, and, to some extent, the mechanism of channeling. Thus it is important to distinguish stable, dynamic, and catalytically induced enzyme associations as well as recognize different mechanisms of substrate channeling. We discuss the principles, experimental details, and limitations and precautions of five rather general methods. These use measurements of transient times, isotope dilution or enhancement, competing reaction effects, enzyme buffering kinetics, and transient-state kinetics. These encompass methods applicable to studies in vitro, in situ, and in vivo. None of these methods is applicable to all systems. They are also susceptible to artifacts without proper attention to precautions. Transient-state kinetic methods clearly excel in elucidating molecular mechanisms of channeling. However, they are often not the best method for initial detection and characterization of the process and they are not applicable to many complex systems. Several other methods that have been successful in indicating substrate channeling are briefly described. PMID- 10527734 TI - Antigen mimicry by anti-idiotypic antibodies: study of interactions between complementary surfaces in macromolecules. AB - Anti-idiotypic antibodies were obtained from New Zealand White rabbits injected with affinity-purified rabbit anti-TrpR antibodies. In gel mobility shift studies, such immunoglobulin preparations were shown to contain one or more species able to form specific complexes with DNA molecules bearing a trp operator. In competitive ELISA assays, the binding of anti-idiotypic antibodies to operator-bearing DNA was reversed by TrpR. The demonstration that the immune repertoire contains information for operator-specific DNA-binding proteins may be relevant to the etiology of certain autoimmune diseases. PMID- 10527735 TI - Applications of the yeast two-hybrid system. AB - In recent years, the yeast two-hybrid system has become the method of choice for detection and analysis of protein-protein interactions in an in vivo context. This system, which capitalizes on the significant genetic history and ease of protocols for manipulation of Saccharomyces cerevisiae, is accessible to most laboratories and is applicable to the pursuit of a large variety of experimental goals. To date, the two-hybrid system has seen widespread application for identification of interaction partners by screening methods using a particular protein of interest as a "bait." Large-scale ventures are also in progress, for example, a cataloging of interactions among the cellular proteins in yeast. However, this method also has tremendous potential for more focused analyses of specific proteins and should become more routine as an alternative or adjunct approach for many structure-function investigations. PMID- 10527737 TI - Microimaging at 14 tesla using GESEPI for removal of magnetic susceptibility artifacts in T(2)(*)-weighted image contrast. AB - In magnetic resonance imaging (MRI), T(2)(*)-weighted contrast is significantly enhanced by extremely high magnetic field strength, offering broad potential applications. However, the T(2)(*)-weighted image contrast distortion and signal loss artifact arising from discontinuities of magnetic susceptibility within and around the sample are also increased, limiting utilization of high field systems for T(2)(*)-weighted contrast applications. Due to the B(0) dependence of the contrast distortions and signal losses, and the heterogeneity of magnetic susceptibility in biological samples, magnetic susceptibility artifacts worsen dramatically for in vivo microimaging at higher fields. Practical applications of T(2)(*)-sensitive techniques enhanced by higher magnetic fields are therefore challenged. This report shows that magnetic susceptibility artifacts dominate T(2)(*)-weighted image contrast at 14 T, and demonstrates that the GESEPI (gradient echo slice excitation profile imaging) technique effectively reduces or eliminates these artifacts at long TE in the highest field (14 T) currently available for (1)H imaging. PMID- 10527736 TI - Genetic approaches to the study of protein-protein interactions. AB - This article describes genetic approaches to the study of heterologous protein protein interactions, focusing on the yeast Saccharomyces cerevisiae as a useful eukaryotic model system. Several methods are described that can be used to search for new interactions, including extragenic suppression, multicopy suppression, synthetic lethality, and transdominant inhibition. Strategies for screening, genetic characterization, and clone identification are described, along with recent examples from the literature. In addition, genetic methods are discussed that can be used to further characterize a newly discovered protein-protein interaction. These include the creation of mutant libraries of a given protein by chemical mutagenesis or polymerase chain reaction, the production of dominant negative mutants, and strategies for introducing these mutant alleles back into yeast for analysis. Although these genetic methods are quite powerful, they are often just a starting point for further biochemical or cell biological experiments. PMID- 10527738 TI - Maps of self-diffusion coefficients by radiofrequency field gradient NMR microscopy AB - The methods of measurement of spatially resolved diffusion coefficients using radiofrequency field gradient (E. Mischler et al., J. Magn. Reson. B 106, 32, 1995; R. Kimmich et al., J. Magn. Reson. A 112, 7, 1995) produce 1D profiles whose amplitude is not only a function of the local self-diffusion coefficient but also is modulated by cosine functions of spatial coordinates. Due to this modulation diffusion-weighted images cannot be obtained unless cumbersome data processing is used. Here, we present a new sequence which avoids this modulation and yields in a straightforward manner true self-diffusion coefficient maps; this is in contrast with conventional methods which use static field gradients and which are therefore altered by background gradients. The feasibility and the reliability of the method are demonstrated with phantoms; it is also applied to different systems of interest such as solvent swelled rubber, membranes, and plants. Copyright 1999 Academic Press. PMID- 10527739 TI - Analysis of dipolar-coupling-mediated coherence transfer in a homonuclear two spin-12 solid-state system AB - Homonuclear dipolar-mediated coherence transfer (DCT), a through-space transfer of magnetization between like spins, can yield otherwise difficult-to-obtain structural information for macromolecules by measuring the internuclear distances between two sites of interest. The behavior of a spin-12 system under DCT is analyzed in detail by computing the time development of the density matrix using the product operator formalism. The effect of coherence transfer (CT) via the homonuclear isotropic scalar coupling on DCT is examined. Analytical and computational results that yield useful information on the frequencies, first maxima, and first-zero of CT for a uniaxially oriented or a single-crystal solid state system are presented. The results predict that the evolution of the spin angular momentum operators under the homonuclear dipolar coupling Hamiltonian leads to "cylindrical mixing" unlike "isotropic mixing" due to the strong scalar coupling Hamiltonian. These results will find relevance in both the design of RF pulse sequences for the structural studies of uniaxially oriented biological solids and the interpretation of solution NMR results from proteins embedded in partially oriented bicelles. Copyright 1999 Academic Press. PMID- 10527740 TI - Magnetic field gradients in solid state magic angle spinning NMR. AB - Magnetic field gradients have proven useful in NMR for coherence pathway selection, diffusion studies, and imaging. Recently they have been combined with magic angle spinning to permit high-resolution measurements of semi-solids, where magic angle spinning averages any residual dipolar couplings and local variations in the bulk magnetic susceptibility. Here we show the first examples of coherence pathway selection by gradients in dipolar coupled solids. When the gradient evolution competes with dipolar evolution the experiment design must take into account both the strength of the dipolar couplings and the means to refocus it. Examples of both homonuclear and heteronuclear experiments are shown in which gradients have been used to eliminate the need for phase cycling. PMID- 10527741 TI - MUSIC in triple-resonance experiments: amino acid type-selective (1)H-(15)N correlations AB - Amino acid type-selective triple-resonance experiments can be of great help for the assignment of protein spectra, since they help to remove ambiguities in either manual or automated assignment procedures. Here, modified triple-resonance experiments that yield amino acid type-selective (1)H-(15)N correlations are presented. They are based on novel coherence transfer schemes, the MUSIC pulse sequence elements, that replace the initial INEPT transfer and are selective for XH(2) or XH(3) (X can be (15)N or (13)C). The desired amino acid type is thereby selected based on the topology of the side chain. Experiments for Gly (G-HSQC); Ala (A-HSQC); Thr, Val, Ile, and Ala (TAVI-HSQC); Thr and Ala (TA-HSQC), as well as Asn and Gln (N-HSQC and QN-HSQC), are described. The new experiments are recorded as two-dimensional experiments and therefore need only small amounts of spectrometer time. The performance of the experiments is demonstrated with the application to two protein domains. Copyright 1999 Academic Press. PMID- 10527742 TI - HN(alpha/beta-COCA-J) experiment for measurement of (1)J(C'C(alpha)) couplings from two-dimensional AB - Anew method for measurement of one-bond (13)C'-(13)C(alpha) scalar and dipolar couplings from a two-dimensional [(15)N, (1)H] correlation spectrum is presented. The experiment is based on multiple-quantum coherence, which is created between nitrogen and carbonyl carbon for simultaneous evolution of (15)N chemical shift and coupling between (13)C' and (13)C(alpha). Optional subspectral editing is provided by the spin-state-selective filters. The residual dipolar dipolar contribution to the (13)C'-(13)C(alpha) coupling can be measured from these simplified [(15)N, (1)H]-HSQC-like spectra. In this way, without explicit knowledge of carbon assignments, conformational changes of proteins dissolved in dilute liquid crystals can be probed conveniently, e.g., in structure activity relationship by NMR studies. The method is demonstrated with human cardiac troponin C. Copyright 1999 Academic Press. PMID- 10527744 TI - Phase-modulated rotating-frame NQR techniques for spatial encoding AB - The rotating-frame method of localization for spatially resolved spectroscopy and imaging in the pure quadrupole regime relies on a gradient B(1) field in which spins experience a flip angle dependent on their position in the B(1) field strength. So far, the techniques have been implemented as amplitude-modulated methods, i.e., the spatial nuclear quadrupole distribution is encoded in the amplitude of the free-induction decay signals. In this work, we describe the implementation of phase-modulated variants of both two-dimensional and rapid rotating-frame imaging techniques. The experiments are discussed for both single crystalline and powder samples. The phase-modulated experiment offers some advantages over the amplitude-encoding technique: It enables one to distinguish the sign of the spatial coordinate and the signal-to-noise ratio is higher than for the simplest amplitude-encoding method. Copyright 1999 Academic Press. PMID- 10527743 TI - An in vivo evaluation of the effects of local magnetic susceptibility-induced gradients on water diffusion measurements in human brain. AB - The effect of possible susceptibility-induced gradients on measurements of water diffusion along the transverse and longitudinal axes of white matter fibers in the brain was investigated in vivo at 1.5 T. Measurements obtained with sequences sensitive and insensitive, respectively, to susceptibility-induced gradients indicated that these gradients do not contribute significantly to diffusion anisotropy in brain white matter. Furthermore, diffusion measurements were unaffected by the presence of known susceptibility-induced gradients at the interface between the petrous bone and brain parenchyma. These results agree with those obtained on in vitro samples and appear to support the hypothesis that interactions between the diffusing water molecules and the cellular environment constitute the principal mechanism for diffusion anisotropy in brain white matter at 1.5 T. This, in turn, simplifies the interpretation of diffusion time dependent measurements in terms of membrane separation and permeability. PMID- 10527745 TI - Invariant and orthonormal scalar measures derived from magnetic resonance diffusion tensor imaging AB - A diffusion tensor is a mathematical construct used to describe water diffusion in complicated biological structures. It describes a process which occurs in all directions simultaneously. It is difficult to comprehend or graphically display the information in the diffusion tensor. This paper describes a coordinate system approach for producing scalar measures which characterize key aspects of the diffusion tensor. The eigenvalues of the diffusion tensor are introduced as the three elements of a point in a Cartesian coordinate system. The Cartesian coordinates are then expressed in cylindrical and spherical coordinates. The orthonormal coordinates of the spherical system are particularly useful scalar measures of attributes of the diffusion tensor: One coordinate contains all the information about the overall magnitude of diffusion. Another contains all of the anisotropy information. The third coordinate contains all of the information about skewness. No information is lost when transforming the original eigenvalues to spherical coordinates. Copyright 1999 Academic Press. PMID- 10527746 TI - Magnitudes and orientations of interaction tensors determined from rotational resonance MAS NMR lineshapes of a four-(13)C-spin system AB - Possibilities and limitations of iterative lineshape fitting approaches for the complete determination of magnitudes and orientations of NMR interaction tensors in a four-(13)C-spin system from MAS NMR experiments are investigated. The availability of fast and numerically accurate computational methods is an important prerequisite. The model compound chosen for this investigation is the monoammonium salt of maleic acid. Various selectively and fully (13)C-labeled versions of this compound permit a stepwise reduction of the number of unknown parameters, necessary to fully describe the four-(13)C-spin system in the uniformly (13)C-labeled maleate moiety. This stepwise procedure allows one to monitor reliability and accuracy of multiparameter fits of the four-(13)C-spin system itself, as well as to characterize limitations and requirements for such fitting procedures. Satisfactory (1)H-decoupling performance is an essential experimental requirement; TPPM decoupling yields n = 1, 2 rotational resonance (13)C MAS NMR lineshapes suitable for analysis by iterative lineshape fitting methods. It is demonstrated that assumptions about "typical" chemical shielding tensor orientations, even if not deviating much from the real orientations, lead to severe errors in internuclear distance determinations. Copyright 1999 Academic Press. PMID- 10527747 TI - Analysis of cross-polarization dynamics between two abundant nuclei, (19)F and (1)H, based on spin thermodynamics theory AB - The phenomenological theory of spin thermodynamics based on the spin temperature hypothesis was employed to describe the cross-polarization (CP) dynamics between two abundant nuclei, (19)F and (1)H, when the number of fluorine atoms is not substantially less than the number of hydrogens. The influence of T(1rho)'s of both nuclei and the relative magnitude (heat capacity) of the two spin baths must be incorporated explicitly into the analysis in order to derive values for the parameters involved in the CP dynamics. Numerical calculations were performed to clarify the difference in the evolution of magnetization in variable contact time CP experiments between the (1)H --> (13)C and (1)H --> (19)F cases. A new type of CP-drain experiment was developed for observing the residual (1)H magnetization after (1)H --> (19)F CP. (19)F direct polarization magic-angle spinning (MAS), (1)H --> (19)F CP, and (1)H --> (19)F CP-drain MAS NMR spectra have been measured for a fluorinated polyimide, 6FDA/ODA. The CP dynamics between (1)H and (19)F for the polyimide were analyzed on the basis of the spin thermodynamics theory. The constant for polarization transfer (T(HF)) was determined by the analysis using the effective CP parameters, which were directly obtained from the CP and CP drain experiments, together with independently measured values of T(H)(1rho) and T(F)(1rho). Copyright 1999 Academic Press. PMID- 10527749 TI - Line narrowing of I = 12 spins coupled to quadrupolar nuclei in liquids: effects of weak decoupling fields AB - In this paper an exact description of the observed transverse magnetization of spin 12 nuclei, coupled to quadrupolar spins which are subjected to RF irradiation, is presented. It is shown that for on-resonance CW decoupling at weak to intermediate irradiation levels, the transverse decay of the spin 12 magnetization is modulated with a period of 1/nu(2), where nu(2) is the amplitude of the decoupling irradiation. When the spin 12 signal is created as a spin echo, and the quadrupolar resonance continuously irradiated during the echo evolution, the echo amplitudes experience much stronger modulation with period 2/nu(2). In previous treatments of such spin systems, the regime of weak decoupling power was usually neglected, and approximate analytical expressions seeking to define the "adequate" or "minimal" decoupling power, necessary to achieve the collapse of the spin 12 multiplet into a single narrow line, were derived. It is demonstrated here, both by experiments and by simulations using a full Redfield formalism, that simple analytical predictions for the T(2) decay of the spin 12 magnetization are still possible, even when the scalar relaxation is not in the "fast exchange" limit and the transverse decay is considerably modulated due to insufficient decoupling power. In this case, the expected single exponential decay rate is obtained for the nonmodulated component of the signal. The theoretical solution for spin I = 12 coupled to S = 3 is derived, and results for the proton decay in (10)B-enriched sodium borocaptate in aqueous solution are presented. The effects of irradiation by several composite pulse decoupling sequences are also considered. Copyright 1999 Academic Press. PMID- 10527748 TI - Toward an in vivo neurochemical profile: quantification of 18 metabolites in short-echo-time (1)H NMR spectra of the rat brain. AB - Localized in vivo (1)H NMR spectroscopy was performed with 2-ms echo time in the rat brain at 9.4 T. Frequency domain analysis with LCModel showed that the in vivo spectra can be explained by 18 metabolite model solution spectra and a highly structured background, which was attributed to resonances with fivefold shorter in vivo T(1) than metabolites. The high spectral resolution (full width at half maximum approximately 0.025 ppm) and sensitivity (signal-to-noise ratio approximately 45 from a 63-microL volume, 512 scans) was used for the simultaneous measurement of the concentrations of metabolites previously difficult to quantify in (1)H spectra. The strongly represented signals of N acetylaspartate, glutamate, taurine, myo-inositol, creatine, phosphocreatine, glutamine, and lactate were quantified with Cramer-Rao lower bounds below 4%. Choline groups, phosphorylethanolamine, glucose, glutathione, gamma-aminobutyric acid, N-acetylaspartylglutamate, and alanine were below 13%, whereas aspartate and scyllo-inositol were below 22%. Intra-assay variation was assessed from a time series of 3-min spectra, and the coefficient of variation was similar to the calculated Cramer-Rao lower bounds. Interassay variation was determined from 31 pooled spectra, and the coefficient of variation for total creatine was 7%. Tissue concentrations were found to be in very good agreement with neurochemical data from the literature. PMID- 10527750 TI - Determination of coupling constants by deconvolution of multiplets in NMR AB - The structures of multiplets in one- and two-dimensional NMR spectra can be simplified by recursive deconvolution in the frequency domain. Deconvolution procedures are described for in-phase and antiphase doublets of delta functions. Recursive simplification is illustrated by applications to double-quantum filtered correlation spectra (DQF-COSY) and selective correlation spectra (soft COSY). Coupling constants can be measured reliably even if signals of opposite signs lead to partial cancellation. Copyright 1999 Academic Press. PMID- 10527751 TI - Dual processing of two-dimensional exchange data in magic angle spinning NMR of solids. AB - We discuss procedures for processing data in rotor-synchronized two-dimensional magic angle spinning (2D MAS) NMR exchange measurements for both structural and dynamical studies. We show, both mathematically and experimentally, that there are two distinct data processing procedures that lead to 2D MAS exchange spectra with purely absorptive crosspeaks. One procedure is that described previously by Hagemeyer, Schmidt-Rohr, and Spiess (HSS). The other procedure is related, but different, and leads to crosspeak intensities given by the formulae of Herzfeld, Roberts, and Griffin (HRG). In 2D MAS exchange experiments on doubly (13)C labeled l-alanylglycylglycine, we demonstrate that the HSS and HRG crosspeak intensities can be extracted separately from the same data set and contain independent information. Processing and analysis of 2D MAS exchange data with both the HSS and the HRG procedures may enhance utilization of the information content of 2D MAS exchange measurements. PMID- 10527752 TI - Simultaneous micromechanical and electromagnetic detection of electron paramagnetic resonance AB - The peculiar advantages of simultaneous observation by electromagnetic and micromechanical methods in EPR spectroscopy are discussed. The development of a novel apparatus with the capability of this simultaneous detection is described. Experiments at 23 GHz show the performance of the apparatus. The problems related to the sensitivity and to the spatial resolution are analyzed. Future prospects are presented. Copyright 1999 Academic Press. PMID- 10527753 TI - Methylene-only subspectra in (13)C CPMAS using a new double quantum filtering sequence AB - Methodology for the assignment of (13)C CPMAS spectra is still in its infancy. Previous methods of CPMAS spectral editing have utilized differences in the strength of the (13)C-(1)H dipolar interaction or the rate and spin thermodynamics of crosspolarization from protons to carbon, to differentiate between quaternary, tertiary, and methylene carbons. We introduce a different approach, which is based on the fact that double-quantum coherence develops between the protons of a methylene group considerably faster than between most other proton spin pairs in an organic solid. We generate this coherence, filter it, convert it back to single quantum, and then crosspolarize selectively to carbon, followed by a short period of reversed crosspolarization to null out unwanted coherence generated from longer distance spin pairs. The sequence has been named DQCP. While the signal-to-noise of this method is poorer than ordinary CP, it is comparable to previous methods for generating methylene-only spectra, and the technique is straightforward and easy to implement. Copyright 1999 Academic Press. PMID- 10527754 TI - Off-resonance multiple-pulse dynamics in solid-state NMR spectroscopy: A revised coherent averaging theory analysis AB - The standard coherent averaging theory treatment of the resonance offset interaction is compared to exact calculations for multiple-pulse solid-state NMR experiments. Significant differences between coherent averaging approximations and exact results are revealed, even for idealized conditions. A revision of the standard analysis is proposed as a way of improving the description of the off resonance dynamics in these experiments. We use the insights obtained from this investigation to derive a qualitatively new type of homonuclear decoupling sequence, and evaluate the performance and advantages of this sequence with both simulations and experimental tests. Copyright 1999 Academic Press. PMID- 10527755 TI - Transverse-relaxation-optimized (TROSY) gradient-enhanced triple-resonance NMR spectroscopy. AB - Two modifications to sensitivity-enhanced gradient-selected TROSY-based triple resonance NMR experiments are proposed that reduce the overall duration of the pulse sequences and minimize radiation damping effects on water-flipback solvent suppression. The modifications are illustrated for the HNCO-TROSY experiment, but are applicable to all triple-resonance experiments that detect proton magnetization after a reverse polarization transfer step from a (15)N spin. The methods are applied to yeast triosephosphate isomerase, a symmetric dimer with 248 amino acid residues per monomer. PMID- 10527756 TI - A pure-phase homonuclear J-modulated HMQC experiment with tilted cross-peak patterns for an accurate determination of homonuclear coupling constants AB - A new HMQC-based experiment is presented which allows for an efficient determination of accurate homonuclear coupling-constant values. Pure absorption lineshapes with tilted cross-peak patterns are obtained by a combination of the active-coupling-pattern tilting (ACT) scheme with J-scaling. Characteristic features include separate heteronuclear echo and antiecho acquisition with a BIRD(y) pulse positioned before or after the t(1) period, respectively, to refocus I-spin homonuclear coupling evolution. Additionally, due to the incorporation of J-scaling the relative spacing of the S-spin chemical-shift differences and I-spin homonuclear coupling splittings in the F(1) domain is largely under experimental control. The most important advantage of the proposed method is that the I-spin homonuclear coupling evolution occurs simultaneously with the evolution of the heteronuclear zero and double-quantum coherences, which exhibit a slower transverse relaxation than I-spin single-quantum coherences. The effectiveness of the new sequence is demonstrated by a determination of the (3)J(HN,Halpha) couplings in a peptide sample. Additionally, the broadband property of the new sequence is verified with a sucrose sample. Copyright 1999 Academic Press. PMID- 10527757 TI - Observation of long-range small-molecule NOEs using a neoteric sensitivity enhancement scheme. AB - A new method to increase the sensitivity of the 1D transient NOE experiment for molecules in the positive NOE regime is presented. This method, the reverse NOE, simply replaces the conventional relaxation delay between scans. Transient positive NOE enhancements from all other spins are used to accelerate the recovery of the target resonance toward its equilibrium intensity. In favorable cases, the intensity of the target peak at the start of an experiment can actually be increased beyond its equilibrium value. There is also a sensitivity enhancement in the rapid pulsing regime, where recovery is always incomplete. This sensitivity enhancement is illustrated with the one-dimensional double pulsed field gradient spin echo NOE experiment to observe a "fourth-order" NOE. Sensitivity gains of 30% are demonstrated. PMID- 10527759 TI - Proposition for assessment of quantitative whole-body autoradiography. AB - To assess recent improvements in quantitative whole-body autoradioluminography (QWBA), the entire QWBA procedure was divided into five processes. Each process was then investigated carefully to determine whether there were any problems in defining a clear standard operating procedure. Results show that use of two instruments, Macro-Cut, Leica, Germany, and the Bioimaging Analyzer with IP, Fuji Photo Film, was essential to produce macroautoradiographs for QWBA data. The remaining problems include the process for freezing the animal carcass and the process for freeze-drying or lyophilizing the frozen sections of the biomaterials. In addition, a desirable standard operating procedure (SOP) must be developed for assessing QWBA. This article proposes satisfactory SOPs with sufficient clarification and experimental proofs to ensure regulatory compliance for the QWBA technique. PMID- 10527758 TI - Effect of 1,25(OH)(2)-vitamin D(3) on the activation of natural killer cells: role of protein kinase C and extracellular calcium. AB - As a first approach for studying the implication of PKC and the steroid hormone 1,25(OH)(2)-vitamin D(3) [1,25(OH)(2)D(3)] on natural killer cell (NK) activity, we analyzed in the YT NK cell line the expression of PKC isoforms and the effects of 1, 25(OH)(2)D(3) on BLT-esterase (a marker of NK lytic granules) activity. Western blot and RT-PCR showed a greater extent of PKC alpha, beta, delta, zeta, epsilon, theta, and lambda and lower levels of PKC mu and eta. In a dose dependent manner 1, 25(OH)(2)D(3) induced significant increases in BLT-esterase and PKC activities and the stimulatory effect on BLT-esterase activity was mimicked and blocked, respectively, by the PKC activator phorbol ester PMA and PKC inhibitors (H7, PKC(19-36), and N-myristoylated PKC(19-31) peptides). Moreover, the effects of 1,25(OH)(2)D(3) on BLT-esterase could be blocked in a Ca(2+)-free (+EGTA) medium and mimicked by the Ca2+ ionophore A23187. The results suggest that 1, 25(OH)(2)D(3) is a stimulatory factor of NK activity acting through a mechanism involving PKC and extracellular Ca2+. PMID- 10527760 TI - Protection against cellular damage in the rat heart by hyperosmotic solutions. AB - In this comparative study, rat hearts were perfused at 37 degrees C with three clearly defined protocols: the Ca2+ paradox, the O(2) paradox, and with 20 mM caffeine. Each protocol involved an initial priming (Ca(2+)(0) depletion or anoxia; stage 1) and subsequent full activation (Ca(2+)(0) repletion, caffeine or reoxygenation; stage 2) of the damage system of the sarcolemma. Creatine kinase release in stage 2 was completely inhibited (P < 0.001) in all three protocols when 420 mOsm was added to the perfusion medium throughout the experiments, or only during stage 1, or only during stage 2. Increasing the perfusion pressure in the Ca2+ paradox significantly (P < 0.001) exacerbated creatine kinase release, although this was still completely inhibited at 28 degrees C. Amiloride (1 mM) inhibited creatine kinase release completely at 40 cm of water pressure but only some 50% at 80 cm of water pressure. It is suggested that the transmembrane damage system needs to be uncoupled or deactivated by modifying its relationship with the cytosol or with the underlying cytoskeleton by hyperosmotic cell shrinkage for only one of the stages in all three protocols to block the damage pathway. Increased perfusion pressure has the opposite effect and exacerbates damage. PMID- 10527761 TI - Treatment of experimental avascular necrosis of the femoral head with hyperbaric oxygen in rats: histological evaluation of the femoral heads during the early phase of the reparative process. AB - The healing of vascular deprivation-induced necrosis of the femoral head of rats exposed to hyperbaric oxygen was compared with that in untreated rats. The amount of necrotic bone, extent of osteoneogenesis, degree of remodeling, and changes of the articular cartilage were histologically graded on a semiquantitative scale of 0 to 3+. On the 2nd, 7th, and 21st postoperative days, there were no differences between the two groups. Newly formed appositional and intramembranous bone was more abundant and remodeling was more advanced in the femoral heads of the hyperbaric oxygen-treated than untreated rats sacrificed on the 42nd postoperative day; also there was less necrotic debris in the femoral heads of the treated rats. There were no differences in the severity of the degenerative changes of the articular cartilage of the treated and untreated rats. Exposure of rats to hyperbaric oxygen does not preserve tissue viability after all arteries supplying the femoral head are severed. Yet, resulting in an increased oxygen tension of the tissues, it seems to provide the optimal settings for reparative processes. The results suggest that hyperoxygenation-mediated relief of ischemia enhances the fibroblastic, angioblastic, osteoblastic, and osteoclastic activities such that healing of the rats' necrotic femoral heads is expedited. PMID- 10527762 TI - Assessment of chromosomal trisomies in prostate cancer using fluorescent in situ hybridization. AB - In a previous study, we observed a low frequency of HER-2/neu oncogene amplification in prostate cancer using fluorescent in situ hybridization (FISH). In our continued effort to identify prognostic biomarkers in prostate cancer, we analyzed 74 cases of prostate cancer to assess the presence of chromosomal trisomies in this cohort of patients. Previous results from this laboratory have implicated a role of chromosomal trisomies in various cancers. FISH using a chromosome 7 and a chromosome 8 centromere probe was utilized to study abnormal chromosome copy numbers together with data from a chromosome 17 control. The frequency of trisomy 7 was found to be 58.1% (43 of 74 informative cases), while the frequency of trisomy 8 was found to be 9.5% (7 of 74 informative cases). The frequency of cells showing chromosome 17 trisomy was 18.5% (15 of 81 cases successfully studied). While chromosome 8 trisomy did not seem to play as significant a role here as in other cancers that we studied, the results of chromosome 7 trisomy are consistent with those reported in the literature. Further exploration of selected trisomies as biomarkers in prostate cancer using a larger study sample size is warranted to establish their clinical utilities. PMID- 10527764 TI - Interstitial fluid pressure and capillary diameter distribution in human melanoma xenografts. AB - Tumors have been shown to differ substantially in interstitial fluid pressure (IFP), but the biological properties of tumors governing the intertumor heterogeneity in IFP have not been identified conclusively. The purpose of the work reported here was to investigate whether the IFP of tumors is influenced significantly by the diameter distribution of the capillaries and hence by the geometric resistance of the capillary network to blood flow. Tumors of three human melanoma xenograft lines (D-12, R-18, U-25) showing similar capillary densities were included in the study. IFP was measured using the wick-in-needle technique. Capillary diameter distribution was determined by stereological analysis of histological sections. The lines differed significantly in tumor IFP (P < 0.05) and capillary diameter distribution (P < 0.05). Mean IFP was 6 mm Hg (D-12), 17 mm Hg (R-18), and 11 mm Hg (U-25). The mean of the mean capillary diameter was 13.1 microm (D-12), 10.9 microm (R-18), and 12.0 microm (U-25). The sequence of the lines from low to high IFP was the same as the sequence of the lines from large to small mean capillary diameter: D-12, U-25, R-18. Also, individual tumors of the same line differed substantially in IFP and in mean capillary diameter. IFP ranged from 2 to 15 mm Hg (D-12), from 2 to 36 mm Hg (R 18), and from 4 to 30 mm Hg (U-25). Mean capillary diameter ranged from 11.0 to 14.6 microm (D-12), from 9.5 to 11.7 microm (R-18), and from 10.4 to 13.0 microm (U-25). Inverse linear correlations between tumor IFP and mean capillary diameter were found for each of the melanoma lines [P < 0.05, R(2) = 0.85 (D-12); P < 0.05, R(2) = 0.86 (R-18); P < 0.01, R(2) = 0.93 (U-25)]. Moreover, the IFP and mean capillary diameter of individual tumors varied with tumor size in all lines. IFP decreased during tumor growth whereas mean capillary diameter increased with increasing tumor volume (P < 0.001). Taken together, these data suggest that the diameter distribution and hence the geometric resistance of the capillary network exerts significant influence on the IFP of tumors. PMID- 10527763 TI - Androgenetic alopecia: in vivo models. AB - Androgenetic alopecia is the most common form of balding in humans. There is great interest in finding a reliable animal model to study the pathogenesis and treatment of this abnormality. The sump-tailed macaque (Macaca artoides) has been the standard model and appears to be useful homologue. These primates are reasonably good predictors of compound efficacy. Due to reduced size and expense, rodent models have been sought. Testosterone inducible models require more development but offer potential. Xenografts of human skin to immunodeficient mice, notably nude or severe combined immunodeficiency, are small, relatively inexpensive, and easy to work with if a source of human tissue is available. Xenografts to double mutant mice for severe combined immunodeficiency and a number of hormone receptor null mutations offer new refinements to these xenograft models. PMID- 10527765 TI - Suppression of sodium-dependent glucose uptake by captopril improves high-glucose induced morphological and functional changes of cultured bovine retinal pericytes. AB - The effects of captopril on glucose uptake, as well as morphological and functional changes of retinal pericytes, in a high-glucose medium were examined. Retinal pericytes were incubated in medium with 5 and 30 mM glucose and 30 mM glucose with 10(-6) to 10(-3) M captopril. Captopril decreased the cellular uptakes of d-glucose and alpha-methyl glucoside in the presence, but not in the absence, of sodium. The cellular size and contents of glucose, sorbitol, and fructose were increased in 30 mM glucose concomitant with the decreased thymidine, cellular DNA content, and ratios in glucose to sorbitol and to fructose, compared with those in 5 mM glucose. These changes observed in 30 mM glucose were reversed by 10(-4) M captopril. These data suggest that the suppression of d-glucose uptake through a sodium-coupled glucose transporter by captopril may attenuate the swelling and loss of pericytes observed in the early stage of diabetic retinopathy. PMID- 10527766 TI - Angiopoietin-1 and its receptor Tie-2 participate in the regulation of capillary like tubule formation and survival of endothelial cells. AB - Angiopoietin-1 (Ang-1) and its receptor Tie-2, a trans-membrane tyrosine kinase uniquely expressed by endothelial cells, are shown by null mutation studies to be essential to developmental angiogenesis. The phenotypic abnormalities in these knockout animals suggest that Tie-2 signaling is necessary for the maintenance and expansion of the primitive capillary network. We present in vitro evidence indicating that the Ang-1/Tie-2 system participates in the regulation of capillary tubule formation and is necessary for the survival of confluent endothelial cells. Although recombinant Ang-1, which induces Tie-2 phosphorylation, has no effect on the proliferation of endothelial cells, treatment of confluent adult bovine aortic endothelial cells (ABAE) cells grown on collagen gels with Ang-1 (100 ng/ml) causes the cells to migrate into the collagen gel and form capillary-like tubules. The tubule-forming effect of Ang-1 is similar to the effect caused by FGF-2. A soluble form of the Tie-2 extracellular domain, in fivefold molar excess, blocks Ang-1-induced tubule formation. Specific elimination of Tie-2 protein expression in cultured ABAE cells as a result of transfection with an antisense oligonucleotide causes cell death in a dose-dependent manner (IC(50) = 50 nM). The antisense treatment has no effect on cells that do not express Tie-2. Cells treated with antisense oligonucleotide show a sixfold increase in the rate of apoptosis as assessed by in situ end labeling of fragmented DNA. These findings are consistent with the view that Ang-1/Tie-2 signaling is essential for both angiogenesis and endothelial cell survival. PMID- 10527767 TI - Adenovirus-mediated acidic fibroblast growth factor gene transfer induces angiogenesis in the nonischemic rabbit heart. AB - Most patients with severe coronary artery disease have normal baseline myocardial blood flow. Therefore, interventions aimed at inducing therapeutic angiogenesis in these patients should cause new blood vessel growth in the heart in the absence of chronic ischemia. It was examined whether adenovirus-mediated gene transfer of recombinant, secreted acidic fibroblast growth factor (sp+aFGF(1 154)), next to a major epicardial artery, may induce neovascularization and reduce the risk region for myocardial infarction upon coronary ligation near the injection site. Fifteen days prior to coronary artery occlusion, rabbits were treated with intramyocardial injections of AdCMV.sp+aFGF(1-154), the control vector AdCMV.NLSbetagal (1 x 10(9) plaque-forming units), or saline. Messenger RNA transcripts for aFGF(1-154) were present up to 12 days after injection in the tissues exposed to AdCMV.aFGF(1-154). Following coronary artery occlusion rabbits treated with AdCMV. sp+aFGF(1-154) showed a 50% reduction of the risk region for myocardial infarction (P < 0.01 vs control). Histologic analysis showed a twofold increase in length density of intramural coronary arterioles (P < 0.01 vs control) and a 17% increase in length density of the capillary network (P < 0.001) in the myocardium exposed to AdCMV.sp+aFGF(1-154). Thus, gene therapy with AdCMV. sp+aFGF(1-154) can induce angiogenesis in the absence of chronic ischemia. The newly formed collateral blood vessels provide an anatomical basis for the reduction in the risk region for myocardial infarction upon subsequent occlusion of the coronary artery in proximity of the site where angiogenesis was induced. PMID- 10527768 TI - Experimental studies on the endothelium ultrastructure of heart capillaries under moderate (28-30 degrees) and deep (22-24 degrees) hypothermia without perfusion. AB - Ultrastructural changes in endothelial cells (EC) of myocardial capillaries were studied in 24 dogs which underwent hypothermia without perfusion. Biopsy specimens for electron microscopy were taken from the left ventricle of each dog in the control group, during anesthesia (prior to active cooling), and at the end of moderate (28-30 degrees ) and deep (22-24 degrees ) artificial body cooling. The following morphological types of the EC were identified both in the control group and in all test groups: those with moderately dense cytoplasm, light, dark, and irreversibly damaged cells. Dark cells showed increased numbers of plasmalemmal vesicles and appeared to be more transport-specialized as opposed to other types. In all stages of the experiment the amount of dark cells continuously increased (to 23.80, 34.62, and 47.17%, respectively). On cooling to 28-30 degrees, subcellular manifestation of reduced synthetic activity of organelles (nucleus, Golgi complex, and rough endoplasmic reticulum) was observed in all types of the EC. These changes persisted, or even increased, at the end of deep hypothermia. The transport activity of the EC changed differently in three experimental groups in all cell types. Micropinocytotic activity increased under spontaneous mild hypothermia (34-35 degrees ) during anesthesia and tended to decrease with subsequent artificial lowering of the temperature to 22-24 degrees. These ultrastructural changes seem to make up an integral part of the process of capillary endothelium adaptation to body surface cooling, and they might contribute to the development of tolerance to subsequent ischemic exposure during cardiac arrest. PMID- 10527769 TI - Exercise skeletal muscle blood flow is related to peripheral microvascular stiffness in idiopathic dilated cardiomyopathy. AB - Peripheral microvascular function plays an important role in congestive heart failure (CHF). Decreased exercise blood flow and microvascular dysfunction have been described in CHF and both factors are regarded as parameters that might influence exercise capacity in these patients. Whether these factors are related to or can be characterized in clinical severity of CHF has not been elucidated in this population. Skeletal muscle blood flow (SMBF) was measured continuously noninvasively, by means of the local isotope washout technique using (133)Xenon, in musculus tibialis anterior during graded maximal supine bicycle exercise. The distensibility in skeletal muscle was measured in a papaverine-relaxed vascular bed using (99m)Tc-pertechnetate. The investigation included 20 patients with moderate CHF (NYHA II), 11 patients with severe CHF (NYHA III, IV) due to idiopathic dilated cardiomyopathy (IDCM), and 31 age-matched healthy subjects. The maximal SMBF level was significantly lower in severe CHF (3.6 +/- 2.5 (ml x (100 g x min)(-1))) compared with moderate CHF (8.6 +/- 5.1 (ml x (100 g x min)( 1)); P < 0.005) and controls (11.0 +/- 4.1 (ml x (100 g x min)(-1)); P < 0.0001), but similar between moderate CHF and controls. Distensibility in skeletal muscle was decreased in severe CHF (12 +/- 8%) compared with controls (44 +/- 17%; P < 0.0001 vs severe CHF) and decreased with increasing severity of CHF (moderate CHF, 23 +/- 14%; P < 0.0005 vs controls). In CHF patients, a relationship was demonstrated between skeletal muscle distensibility and the maximal SMBF (P < 0.0001; r = 0.70). Moreover, maximal SMBF correlated directly to exercise time (P < 0.005; r = 0.54). Patients with CHF have reduced exercise SMBF, which may be a limiting factor for the reduced maximal exercise capacity. Moreover, microvascular distensibility in skeletal muscle is reduced and correlates to maximal exercise SMBF. Furthermore, maximal SMBF correlates to exercise time. This implies that increased skeletal muscle microvascular stiffness may contribute to the reduced blood flow during exercise and SMBF may partly limit exercise performance in CHF patients due to IDCM. PMID- 10527770 TI - A new view of Starling's hypothesis at the microstructural level. AB - In this paper we quantitatively investigate the hypothesis proposed by Michel (Exp. Physiol. 82, 1-30, 1997) and Weinbaum (Ann. Biomed. Eng. 26, 1-17, 1998) that the Starling forces are determined by the local difference in the hydrostatic and colloid osmotic pressure across the endothelial surface glycocalyx, which we propose is the primary molecular sieve for plasma proteins, rather than the global difference in the hydrostatic and oncotic pressure across the capillary wall between the plasma and tissue, as has been universally assumed until now. A spatially heterogeneous microstructural model is developed to explain at the cellular level why there is oncotic absorption at low capillary pressures in the short-lived transient experiments of Michel and Phillips (J. Physiol. 388, 421-435, 1987) on frog mesentery capillary, but a small positive filtration once a steady state is achieved. The new model also predicts that the local protein concentration behind the surface glycocalyx can differ greatly from the tissue protein concentration, since the convective flux of proteins through the orifice-like pores in the junction strand will greatly impede the back diffusion of the proteins into the lumen side of the cleft when the local Peclet number at the orifice is >1. The net result is that the filtration in the capillaries is far less than heretofore realized and there may be no need for venous reabsorption. PMID- 10527771 TI - A single high dose of vitamin C counteracts the acute negative effect on microcirculation induced by smoking a cigarette. AB - Cigarette smoking is associated with marked acute changes in microcirculation including reduced blood flow. We tested the hypothesis that the reduced blood flow velocity is due to the imbalance between prooxidants and antioxidants that occurs as a consequence of smoking and that it can be reduced by an antioxidant. The effect of smoking a single cigarette on nail-fold microcirculation was analyzed in 24 healthy subjects with varying smoking habits. Vital capillary microscopy was used and the blood cell flow velocity in the capillaries was evaluated before and 1-30 min after smoking. Smoking induced a marked decrease in microcirculatory blood flow in 23 of the 24 subjects (40-50% decrease 1-5 min after smoking). This change was reduced by more than 50% in the same subjects after intake of 2 g of vitamin C 2 h before smoking (P < 0.0001 by ANOVA test) with smokers responding similarly to nonsmokers in these experiments. Intake of 1 g of vitamin C had no significant effect on the smoking-induced changes in most of the subjects tested (n = 11). Pretreatment with aspirin had little or no effect on the response to smoking (n = 9). Our results show that treatment with a single high dose of vitamin C can reduce and in some individuals even completely abolish the negative acute effect on microcirculation induced by smoking a single cigarette. This effect of vitamin C is not likely to be mediated by the cyclooxygenase system. PMID- 10527772 TI - Microvessel organization and structure in experimental brain tumors: microvessel populations with distinctive structural and functional properties. AB - We studied microvessel organization in five brain tumor models (ENU, MSV, RG-2, S635cl15, and D-54MG) and normal brain, including microvessel diameter (LMVD), intermicrovessel distance (IMVD), microvessel density (MVD), surface area (S(v)), and orientation. LMVD and IMVD were larger and MVD was lower in tumors than normal brain. S(v) in tumors overlapped normal brain values and orientation was random in both tumors and brain. ENU and RG-2 tumors and brain were studied by electron microscopy. Tumor microvessel wall was thicker than that of brain. ENU and normal brain microvessels were continuous and nonfenestrated. RG-2 microvessels contained fenestrations and endothelial gaps; the latter had a maximum major axis of 3.0 microm. Based on anatomic measurements, the pore area of RG-2 tumors was estimated at 7.4 x 10(-6) cm(2) g(-1) from fenestrations and 3.5 x 10(-5) cm(2) g(-1) from endothelial gaps. Increased permeability of RG-2 microvessels to macromolecules is most likely attributable to endothelial gaps. Three microvessel populations may occur in brain tumors: (1) continuous nonfenestrated, (2) continuous fenestrated, and (3) discontinuous (with or without fenestrations). The first group may be unique to brain tumors; the latter two are similar to microvessels found in systemic tumors. Since structure function properties of brain tumor microvessels will affect drug delivery, studies of microvessel function should be incorporated into clinical trials of brain tumor therapy, especially those using macromolecules. PMID- 10527773 TI - Pressure-permeability relationships in crosslinked basement membranes. PMID- 10527774 TI - Analysis of Rotational and Rovibrational Spectra of SiDF(3) in the Ground and v(4) = 1 Excited States. AB - This paper presents the analysis of the spectra of SiDF(3) in its vibrational ground and v(4) = 1 states. The pure rotational spectrum of the ground state was measured up to 903 GHz (J" = 65). Rotational, quartic, and sextic centrifugal distortion constants were accurately determined. Furthermore the parameter ||h(3) || = 7.64(11) x 10(-4) Hz was derived from the observation of A(1)-A(2) splittings of six K = 3 lines. The assignment of the v(4) = 1 spectrum was performed by combining 140 pure rotational frequencies and more than 2300 rovibrational transitions. Among them, because of the strong (+/-2, -/+4) interaction, there are more than 90 A(1)-A(2) resolved split transitions between the (l = 0, k = +/-3) ground state levels and the (l = +/-2, k = -/+1) v(4) = 1 levels. Although the v(4) band is located at about 994.3 cm(-1), the energies of the other fundamental bands are at a distance of more than 140 cm(-1). The assumption of the isolated character of the state was confirmed by the small differences between the v(4) = 1 state and the ground state parameters and by the possibility of using the D and Q reduction schemes proposed by E. I. Lobodenko, O. N. Sulakshina, V. I. Perevalov, and Vl. G. Tyuterev [J. Mol. Spectrosc. 126, 159-170 (1987)]. Copyright 1999 Academic Press. PMID- 10527775 TI - Laser-Excited Fluorescence Spectra of Yttrium Monoiodide. AB - The rotational structure of the laser-excited fluorescence of yttrium monoiodide, recorded on a Fourier transform spectrometer, was analyzed, enabling the characterization of two excited states at T(e) approximately 20 690 and approximately 24 100 cm(-1). Observation of the ground state X(1)Sigma(+) is extended up to v = 15. A scheme for the electronic structure is proposed, involving the X(1)Sigma(+), B(1)Pi, D(1)Pi, Omega1, and possibly A((1)Delta) states. Copyright 1999 Academic Press. PMID- 10527776 TI - Determination of the Line-Shift and Line-Broadening Coefficients in the nu(3) Band of ()NO(2) Perturbed by ()O(2), N(2), H(2), D(2), and CO(2). AB - A high-resolution three-channel diode-laser spectrometer was used to record a large number of spectra with (14)N(16)O(2) and foreign gas pressures ranging from 0.1 to 0.4 and 20 to 150 Torr, respectively, at room temperature. The 6.2-um region corresponding to the nu(3) band of NO(2) was analyzed and the broadening and shift coefficients were derived in the case of collisions between NO(2) and H(2), D(2), O(2), N(2), CO(2), and SO(2) for 13 lines with 18 O(2) + O shows very good agreement with experimental data. The new potential energy surface was used to predict the band centers of the isotopomers (17)O(3) and (18)O(3). Copyright 1999 Academic Press. PMID- 10527782 TI - The High-Resolution Infrared Spectrum of 1,2,4-Triazine Vapor between 550 and 1700 cm(-1). AB - The Fourier transform gas-phase IR spectrum of 1,2,4-triazine between 550 and 1700 cm(-1) was measured with a resolution of ca. 0.003 cm(-1). Comparing with the liquid-phase IR and Raman spectra and using ab initio predictions, most of the fundamental bands of 1,2,4-triazine below 1600 cm(-1) were assigned. From the high-resolution gas-phase spectra, 12 of the fundamental bands were analyzed by the Watson Hamiltonian model to yield upper state spectroscopic constants. A number of local resonances were identified and explained. From a simultaneous ground state combination difference analysis of four of the bands, a set of ground state rotational and centrifugal distortion constants were obtained. Copyright 1999 Academic Press. PMID- 10527783 TI - Rotational Analysis of the LiHg 2(2)Pi(3/2)-X(2)Sigma(+)(1/2) (v' = 0 <-- v" = 0, 1, 2) Vibronic Bands. AB - We present a complete analysis of the previously reported LiHg 2(2)Pi(3/2) X(2)Sigma(+) excitation spectrum [X. Li, P. Pircher, D. Gruber, and L. Windholz, Chem. Phys. Lett. 263, 463 (1996)]. The v' = 0 - v" = 0, 1, 2 vibronic bands are analyzed and the rotational transitions are identified. The assigned rotational transitions are initially fitted on a band-to-band basis. A global fit is performed afterwards for all the known vibronic transitions of the LiHg molecules. In the fit the ground state molecular constants are separated into spin-rotation coupling terms and Dunham-type coefficients. The obtained Dunham coefficients are used to derive the molecular potential for the ground electronic state using the inverted perturbation approach (IPA). The equilibrium bond length was found to be 2.9528(6) A for the LiHg ground state. Copyright 1999 Academic Press. PMID- 10527784 TI - Absolute Intensity of the NH(3) nu(2) Band. AB - Absolute line intensities of NH(3) in the nu(2) band were measured for a number of rovibrational lines using a high-resolution Fourier transform infrared spectrometer. The transition dipole moments calculated from the absolute intensities showed significant and systematic deviations from the usual rotational dependencies predicted from the J, K dependence of the direction cosine matrix elements. The anomalous rotational dependencies may be explained by vibration-rotation interactions, and the Herman-Wallis-type rotational correction factors for a symmetric top molecule were determined experimentally for the first time in the present study. The self-broadening coefficients were determined as well. Copyright 1999 Academic Press. PMID- 10527785 TI - Rovibrational Constants for the nu(6) and 2nu(9) Bands of HCOOD by Fourier Transform Infrared Spectroscopy. AB - The Fourier transform infrared spectrum of the nu(6) and 2nu(9) bands of deuterated formic acid (HCOOD) was recorded with an apodized resolution of 0.004 cm(-1) in the frequency range of 930-1040 cm(-1). These two bands with band centers 40 cm(-1) apart were mutually coupled by Coriolis and Fermi interactions. By fitting a total of 1076 infrared transitions of both nu(6) and 2nu(9) with a standard deviation of 0.00075 cm(-1) using a Watson's A-reduced Hamiltonian in the I(r) representation with the inclusion of c-type Coriolis and a Fermi resonance term, two sets of rovibrational constants for v(6) = 1, and v(9) = 2 states were derived for the first time. Both nu(6) and 2nu(9) bands are A type with band centers at 972.8520 +/- 0.0001 and 1011.6766 +/- 0.0001 cm(-1), respectively. Copyright 1999 Academic Press. PMID- 10527786 TI - Direct High-Resolution Determination of the Singlet-Triplet Splitting in NH Using Stimulated Emission Pumping. AB - A completely resolved spectrum of the strongly forbidden NH (a(1)Delta --> X(3)Sigma(-)) transition is observed. The NH radicals in the excited a(1)Delta state are exclusively generated in a Nd:YAG laser photolysis of hydrazoic acid at a wavelength of 266 nm. The NH (a(1)Delta --> X(3)Sigma(-)) intercombination transition around 794 nm is used to produce NH (X(3)Sigma(-)) applying the stimulated emission pumping technique. The ground state radicals are detected by laser-induced fluorescence (LIF). The energy splitting between the NH (X(3)Sigma( ), v = 0, J = 1, N = 0) state and the NH (a(1)Delta, v = 0, J = 2) state is determined with an accuracy of 0.1 cm(-1) to DeltaE = 12 687.8(65) cm(-1). In addition, the radiative lifetime tau of the NH (a(1)Delta --> X(3)Sigma(-)) transition was estimated by a determination of the saturation intensity to be tau approximately 12.5 s. Copyright 1999 Academic Press. PMID- 10527787 TI - High-Resolution Infrared Laser Study of the nu(4) Absorption Band of cis CHCl&dbond;CHF. AB - cis-1-Chloro-2-fluoroethylene was synthesized, and the gas-phase infrared spectrum was investigated in the nu(4) band region between 1321-1350 cm(-1), at a resolution of about 0.002 cm(-1), employing a tunable diode-laser spectrometer. This vibration of symmetry species A' gives rise to an a/b-hybrid band, even though our analysis pointed out that the intensity of the a-type component is predominant. Most of the J and K structure was resolved in different subbranches, and the rovibrational analysis led to the assignment of about 1900 a-type lines with J X(3)Sigma(-) and a(1)Delta --> X(3)Sigma(-) Transitions of SO. AB - Emission spectra of the b(1)Sigma(+) --> X(3)Sigma(-) and a(1)Delta --> X(3)Sigma(-) transitions of SO in the near-infrared spectral region were studied at high spectral resolution with a Fourier-transform spectrometer. The 0-0 band of the b --> X system was measured at high signal/noise and four magnetic dipole branches were observed in addition to the previously known five electric dipole branches. From the relative line intensities and the radiative lifetime of the b state, the electric and magnetic transition moments were determined to be u(0) = +/-0.0042 ea(0), u(1) = -/+0.0047 ea(0), and M = 0.16 u(B). All nine electric dipole branches were observed in the 0-0 band of the a --> X transition. Fixing the rotational constants of the X(3)Sigma(-) ground state to microwave values, the following parameters were obtained for the a(1)Delta state (in cm(-1)): v(00) = 5862.1853(1), B(0) = 0.71033746(37), D(0) = 1.16814(46) x 10(-6) (with H(0) fixed to -3.99 x 10(-13)), where the numbers in parentheses are the standard deviations of the parameters. Copyright 1999 Academic Press. PMID- 10527792 TI - 19F Nuclear Spin-Rotation Constant of Yttrium Monofluoride. PMID- 10527791 TI - Near-Infrared Laser Spectroscopy of the C(3)Delta-X(3)Delta Transition of TiS. AB - The (0, 0), (1, 0), (2, 0), (3, 0), and (4, 0) bands of the C(3)Delta-X(3)Delta transition of TiS, between 743-863 nm, were studied using the technique of laser vaporization/reaction with supersonic cooling and laser-induced fluorescence (LIF) spectroscopy. The linewidth of the LIF spectrum obtained was about 250 MHz. The P, Q, and R branches of each DeltaOmega = 0 subband were observed. A merged least-squares fit of the measured line positions yielded molecular parameters of the v = 0-4 levels of the C(3)Delta state and the v = 0 level of the X(3)Delta state. Copyright 1999 Academic Press. PMID- 10527793 TI - On "Transition Dipole Moments for the Vibrational Fundamentals of HNC Determined from the Herman-Wallis Effect" PMID- 10527794 TI - The 2nu(1) + 5nu(3) Triad of (12)CO(2). PMID- 10527795 TI - Observation of Magnetic-Dipole Transitions in the CN B(2)Sigma(+)-A(2)Pi Band System. PMID- 10527796 TI - An Accurate Equilibrium Structure and CCSD(T) Spectroscopic Constants for Linear C(3)Si(2). PMID- 10527797 TI - First Spectroscopic Observation of Gaseous Diatomic CuGe. PMID- 10527798 TI - New Low-Lying Electronic States of LaF. PMID- 10527799 TI - Neurobiology of disease in the 21st century PMID- 10527800 TI - Experimental models of amyotrophic lateral sclerosis. AB - Amyotrophic lateral sclerosis (ALS) is a chronic neurodegenerative disease characterized by the progressive loss of motor neurons, leading to profound weakness and eventual death of affected individuals. For the vast majority of patients with ALS, the etiology of the disorder is unknown, and although multiple clinical trials of various therapeutic agents have been undertaken, truly effective therapy is not currently available for the disease. The selection of treatments used in ALS clinical trials frequently has its basis in promising data obtained from experimental model systems in which the proposed agent has shown some effect in protecting motor neurons from a particular insult. The likelihood of a successful clinical outcome for a given treatment in ALS would therefore depend on two principal factors, including the similarity of the model to the disease and the biologic action of the potential therapeutic agent. Partly because early experimental models of ALS failed to replicate the disease process, treatment success in these models did not carry over into human trials. Recently, however, a variety of newer model systems have been developed and utilized to investigate motor neuron degeneration as related to ALS. For example, in this issue, Corse et al. use a rat spinal cord organotypic slice subjected to glutamate excitotoxicity as a model system to test the effectiveness of neurotrophic factors in preventing motor neuron degeneration. This review will assess the strengths and weaknesses of differing ALS model systems that have been used to preclinically test potential drug efficacy in ALS. PMID- 10527801 TI - Targeted disruption of the Cln3 gene provides a mouse model for Batten disease. The Batten Mouse Model Consortium [corrected]. AB - Batten disease, a degenerative neurological disorder with juvenile onset, is the most common form of the neuronal ceroid lipofuscinoses. Mutations in the CLN3 gene cause Batten disease. To facilitate studies of Batten disease pathogenesis and treatment, a murine model was created by targeted disruption of the Cln3 gene. Mice homozygous for the disrupted Cln3 allele had a neuronal storage disorder resembling that seen in Batten disease patients: there was widespread and progressive intracellular accumulation of autofluorescent material that by EM displayed a multilamellar rectilinear/fingerprint appearance. Inclusions contained subunit c of mitochondrial ATP synthase. Mutant animals also showed neuropathological abnormalities with loss of certain cortical interneurons and hypertrophy of many interneuron populations in the hippocampus. Finally, as is true in Batten disease patients, there was increased activity in the brain of the lysosomal protease Cln2/TPP-1. Our findings are evidence that the Cln3-deficient mouse provides a valuable model for studying Batten disease. PMID- 10527802 TI - Preclinical testing of neuroprotective neurotrophic factors in a model of chronic motor neuron degeneration. AB - Many neurotrophic factors have been shown to enhance survival of embryonic motor neurons or affect their response to injury. Few studies have investigated the potential effects of neurotrophic factors on more mature motor neurons that might be relevant for neurodegenerative diseases. Using organotypic spinal cord cultures from postnatal rats, we have demonstrated that insulin-like growth factor-I (IGF-I) and glial-derived neurotrophic factor (GDNF) significantly increase choline acetyltransferase (ChAT) activity, but brain-derived neurotrophic factor (BDNF), neurotrophin-4 (NT-4/5), and neurotrophin-3 (NT-3) do not. Surprisingly, ciliary neurotrophic factor (CNTF) actually reduces ChAT activity compared to age-matched control cultures. Neurotrophic factors have also been shown to alter the sensitivity of some neurons to glutamate neurotoxicity, a postulated mechanism of injury in the neurodegenerative disease, amyotrophic lateral sclerosis (ALS). Incubation of organotypic spinal cord cultures in the presence of the glutamate transport inhibitor threo-hydroxyaspartate (THA) reproducibly causes death of motor neurons which is glutamate-mediated. In this model of motor neuron degeneration, IGF-I, GDNF, and NT-4/5 are potently neuroprotective, but BDNF, CNTF, and NT-3 are not. The organotypic glutamate toxicity model appears to be the best preclinical predictor to date of success in human clinical trials in ALS. PMID- 10527803 TI - Neurons undergo apoptosis in animal and cell culture models of diabetes. AB - Recent clinical trials indicate that the severity of diabetic neuropathy is correlated with the level of patient glycemic control. In the current study, hyperglycemia induces apoptotic changes in dorsal root ganglion neurons and Schwann cells in vivo both in streptozotocin-treated diabetic rats and in rats made acutely hyperglycemic with infused glucose. Typical apoptotic nuclear and cytoplasmic changes are observed. In addition mitochondrial changes recently reported to occur as part of the apoptotic cascade, such as ballooning of mitochondria and disruption of the internal cristae, are seen in diabetic dorsal root ganglion neurons and Schwann cells. Similar changes have been reported in neurons in the presence of oxidative stress. In order to study the neurotoxic effects of high glucose we developed an in vitro model using rat dorsal root ganglion neurons. In dorsal root ganglion cultured in defined medium, addition of moderate glucose levels results in neurite degeneration and apoptosis. These changes are coupled with activation of caspase-3, dependent on the concentration of glucose. The apoptotic changes observed in vitro are similar to those observed in vivo. In contrast, addition of IGF-I, even at physiological concentrations, prevents activation of caspase-3 and neuronal apoptosis in vitro. We suggest that oxidative stress may promote the mitochondrial changes in diabetic animals and lead to activation of programmed cell death caspase pathways. These results imply a new pathogenetic mechanism for diabetic sensory neuropathy. PMID- 10527804 TI - Mutant huntingtin forms in vivo complexes with distinct context-dependent conformations of the polyglutamine segment. AB - Huntington's disease (HD) is caused by an expanded glutamine tract, which confers a novel aggregation-promoting property on the 350-kDa huntingtin protein. Using specific antibodies, we have probed the structure of the polyglutamine segment in mutant huntingtin complexes formed in cell culture from either truncated or full length protein. Complexes formed by a mutant amino terminal fragment most frequently entail a change in conformation that eliminates reactivity with the polyglutamine-specific mAb 1F8, coincident with production of insoluble aggregate. By contrast, complexes formed by the full-length mutant protein remain soluble and are invariably 1F8-reactive, indicating a soluble polyglutamine conformation. Therefore, aggregates in HD may form by different biochemical mechanisms that invoke different possibilities for the pathogenic process. If pathogenesis is triggered by a truncated fragment, it probably involves the formation of an insoluble aggregate. However, the observation of soluble complexes in which an HD-specific pathogenic conformation of the glutamine tract remains accessible suggests that pathogenesis could also be triggered at the level of full-length huntingtin by abnormal aggregation with normal or abnormal protein partners. PMID- 10527805 TI - Alzheimer's disease associated presenilin 1 interacts with HC5 and ZETA, subunits of the catalytic 20S proteasome. AB - Proteolytic processing and degradation tightly regulate the amount of stable, functional presenilin 1 (PSEN1) in the cell. The approximately 46-kDa PSEN1 holoprotein is cleaved into a approximately 30-kDa N-terminal fragment (NTF) and a approximately 20-kDa C-terminal fragment (CTF) by an unknown protease. The fragments are stabilized in a high molecular weight complex and nonincorporated fragments and excess holoprotein are degraded by the 26S proteasome. The tight balance between, on the one hand, processing and incorporation into the stable complex and, on the other hand, proteolytic degradation of excess PSEN1, indicates that minor changes in one of these two processes could be pathologically relevant. Here we demonstrate the direct physical interaction between PSEN1 and two subunits, HC5 and ZETA, of the 20S proteasome. These interactions were identified using an interaction trap screening and were further established in an in vitro binding assay. Furthermore, we were able to coimmunoprecipitate the transfected binding partners, as well as the endogenous PSEN1 and ZETA proteins from HEK 293T cells. Finally, degradation of ubiquitinated wild-type and mutant PSEN1 by the 26S proteasome was demonstrated. In conclusion, we report a direct interaction between PSEN1 and subunits of the 20S catalytic particle of the 26S proteasome, further establishing the involvement of proteasomal degradation in the regulation of PSEN1 turnover. PMID- 10527806 TI - T cell receptor BV gene rearrangements in the spinal cords and cerebrospinal fluid of patients with amyotrophic lateral sclerosis. AB - Amyotrophic lateral sclerosis (ALS) is a fatal disorder whose etiology and pathogenesis remain unknown. Recent studies, however, have demonstrated the presence of inflammatory infiltrates within ALS spinal cord and suggested the possibility of an immune-mediated process in motor neuron degeneration. We have analyzed the diversity of T-cells in the spinal cord in ALS. Reverse transcriptase polymerase chain reaction (RT-PCR) with variable (V) region sequence specific oligonucleotide primers was used to amplify T-cell receptor (TCR)BV transcripts from spinal cords obtained at autopsy from patients with ALS, patients who died without inflammatory disease of the central nervous system, brains from patients with ALS, and brains from patients who died with inflammatory CNS disease. Sequencing was then performed on the amplified transcripts. An overall increase in the level of TCRBV 2 transcripts was detected in ALS specimens when compared to controls. This result was independent of the HLA genotype of the individual. Furthermore, enrichment of TCRBV2-positive T cells could be demonstrated in cerebrospinal fluid derived from patients with ALS, using PCR analysis and a T cell stimulation assay with toxic shock syndrome toxin-1 (TSST-1), a Vbeta2-specific superantigen. Our results suggest that an immunological process involving the specific expansion of Vbeta2 TCR-positive T cells may be important in the pathogenesis of ALS. PMID- 10527807 TI - Intrinsic physiological and morphological properties of principal cells of the hippocampus and neocortex in hamsters infected with scrapie. AB - Scrapie is a transmissible spongiform encephalopathy, or "prion disease." We investigated the effects of intracerebral Sc237 scrapie inoculation in hamsters on the physiology and morphology of principal cells from neocortical and hippocampal slices. Scrapie inoculation resulted in increased branching of basal dendrites of hippocampal CA1 pyramidal cells (Sholl analysis), reduced amplitudes of medium and late afterhyperpolarizations (AHPs) in CA1 pyramidal cells and layer V neocortical cells, loss of frequency potentiation of depolarizing afterpotentials (DAPs), and double action potentials in synaptically evoked CA1 pyramidal cell responses. Postsynaptic double action potentials could also be evoked in normal hamster CA1 pyramidal cells by acute pharmacological block of AHPs, suggesting that the depressed AHPs in scrapie-infected hamsters caused the action potential doublets. Both the AHP and the DAP potentiations depend on increased intracellular calcium, which suggests that the underlying deficit, in hamsters infected with Sc237 scrapie, may lie in calcium entry and/or homeostasis. Fast IPSPs, passive membrane properties, and density of dendritic spines remained unchanged. These last two results differ markedly from recent studies on mice infected with ME7 scrapie, indicating diversity of pathophysiology in this group of diseases, perhaps associated with the progressive and substantial neuronal loss found in the ME7, and not the Sc237, model. PMID- 10527808 TI - Molecular interpretation of expanded RED products in bipolar disorder by CAG/CTG repeats located at chromosomes 17q and 18q. AB - Previously we provided evidence that the anticipation observed in bipolar (BP) disorder may be explained by expanded CAG/CTG triplet repeats. Data were generated with the repeat expansion detection (RED) method in a BP case-control sample showing a significant association of BP disorder with expanded CAG/CTG repeats (RED products of 120 bp). In this study we demonstrated that 86% of the RED expansions could be accounted for by the ERDA1 and CTG18.1 CAG/CTG repeats located respectively on chromosomes 17 and 18. Further, significantly different allele distributions were observed for ERDA1, with a larger proportion of BP patients (34.7%) carrying one or two expanded ERDA1 alleles (CAG/CTG repeats >40) than controls (19.2%) (P = 0.032). Also, a negative correlation was observed for ERDA1 between CAG/CTG length and age at onset in affected offspring of eight BP families. Although interesting, these data should be interpreted with caution since the ERDA1 association did not remain significant after correcting for multiple testing. Also, no linkage was observed between BP disorder and expanded ERDA1 alleles in the families. PMID- 10527809 TI - The T102C polymorphism of the 5-HT2A-receptor gene in fibromyalgia. AB - Based on a possible involvement of serotonergic dysfunction in the pathophysiology of fibromyalgia (FM) and on preliminary reports of a possible genetically driven vulnerability for this disorder we investigated the silent T102C polymorphism of the 5-HT2A-receptor gene in 168 FM patients and 115 healthy controls. Our results showed a significantly different genotype distribution in FM patients with a decrease in T/T and an increase in both T/C and C/C genotypes as compared to the control population (Fisher's Exact test, two-sided, P = 0.008). However, the increase in allele-C102 frequency felt short of significance (P = 0.07). Correlation of genotypes to clinical parameters revealed no influences on age of onset, duration of disease or psychopathological symptoms, measured with the Beck Depression Inventory and the symptom checklist SCL-90-R. In contrast to that the pain score, being a self reported information on pain severity, was significantly higher in patients of the T/T genotype (Mann-Whitney U test, P = 0.028). This suggests that the T102-allele might be involved in the complex circuits of nociception. However, the T102C polymorphism is not directly involved in the aetiology of FM but might be in linkage dysequilibrium with the true functional variant, which has to be unravelled. PMID- 10527812 TI - Frames of Reference in Quantity Estimations by Groups and Individuals. AB - The superiority of group performance over performance of the average individual is relatively greater on world knowledge tasks than on quantity estimation tasks. Previous research on quantity estimations has involved judgments without an explicit frame of reference. We propose that a frame of reference converts a quantity estimation into a world knowledge inference by embedding the estimation in a larger cognitive structure. Individuals first estimated 30 pairs of quantities, such as the length of the Ohio River and the length of the Arkansas River, given either 2 statements as a frame of reference (the Mississippi River is 2340 miles long; the Colorado River is 1450 miles long), 1 of these statements as a frame of reference, or no frame of reference. Then they made the same 30 pairs of estimations again as 3-person groups or as individuals under the same frame-of-reference conditions. As predicted, group estimations were more accurate than individual estimations, both group and individual estimations were more accurate with either a 2-statement or a 1-statement frame of reference than without a frame of reference, and the frame of reference improved group estimations relatively more than individual estimations. Copyright 1999 Academic Press. PMID- 10527811 TI - PMP22 accumulation in aggresomes: implications for CMT1A pathology. AB - Peripheral myelin protein 22 (PMP22) is a 22-kDa glycoprotein mainly expressed by Schwann cells (SCs). Duplication or deletion of the PMP22 gene locus is associated with heritable peripheral neuropathies suggesting that the correct level of PMP22 protein is essential for SC functioning. Previously we reported that in SCs the majority (80%) of newly synthesized PMP22 is rapidly degraded, possibly due to inefficient folding. Here we show that inhibition of the proteasome pathway results in a marked accumulation of PMP22 in the perinuclear cytoplasm. Double immunolabeling with an anti-ubiquitin antibody and various organelle markers indicates that the accumulated PMP22 is found in unique intracellular inclusions, called aggresomes. Moreover, overexpression of PMP22 in SCs can induce perinuclear accumulation of the protein. Together, these studies suggest that the proteasome pathway is critical for the regulation of PMP22 protein levels and raise the possibility that aggresomes may be involved in the pathogenesis of PMP22-associated peripheral neuropathies. PMID- 10527810 TI - Neuronal apoptosis induced by beta-amyloid is mediated by caspase-8. AB - The Alzheimer disease-associated beta-amyloid peptide has been shown to induce apoptotic neuronal death. In the present study, we test the hypothesis that the apoptotic pathway activated by beta-amyloid is similar to the pathway activated by the Fas/TNFR family of death receptors, which requires caspase-8 activity and adaptor proteins such as FADD. We demonstrate that the selective caspase-8 inhibitor IETD-fmk blocks neuronal death induced by beta-amyloid. Furthermore, using viral-mediated gene delivery, we show that neurons expressing dominant negative FADD are protected from apoptosis induced by beta-amyloid. Together these results indicate that the apoptotic pathway activated by beta-amyloid requires both caspase-8 activity and FADD. These findings further support the hypothesis that beta-amyloid might initiate apoptosis by cross-linking death receptors of the Fas/TNFR family. PMID- 10527813 TI - The Disjunction Effect and Reason-Based Choice in Games. AB - This paper reports an experiment that extends previous find ings of the disjunction effect, sometimes described as a violation of Savage's sure-thing principle. Evidence of the disjunction ef fect is observed using elicited beliefs about others' actions (rather than controlled beliefs) in a prisoners' dilemma studied by Shafir and Tversky (1992) as well as in an asymmetric version of it and in a nondilemma game with a unique equilibrium. Debiasing tech niques as well as implications for these extensions are discussed. Copyright 1999 Academic Press. PMID- 10527814 TI - The Effects of Subject-Defined Categories on Judgmental Accuracy in Confidence Assessment Tasks. AB - The accuracy of confidence judgments can be determined using measures of discrimination and calibration. The present paper utilizes a new assessment methodology that decomposes the confidence assessment task, allowing us to investigate discrimination and calibration skills in greater depth than has been done in previous studies. Researchers investigating the goodness of confidence judgments have typically grouped forecasters' assessments into experimenter defined categories, generally in equal widths of.10. In the present research, subjects created their own categories and later assigned confidence judgments to the categories, separating the tasks of discriminating categories (discrimination) and assigning numbers to categories (calibration). Further, the typical assessment procedure assumes that subjects are able to discriminate equally across the confidence scale. Since subjects in the present study defined their own assessment categories, they could locate those categories at any point on the scale. A final issue of interest was whether subjects were able to determine accurately the number of categories into which they could discriminate. Sixty subjects performed 1 of 2 tasks, general knowledge or forecasting, in both relatively easy and relatively hard conditions. Results showed a trade-off in performance: Calibration generally became worse as the number of categories increased, while discrimination generally improved. Overall accuracy was not affected by the number of categories used. Further, subjects partitioned categories more at the high end of the scale. Finally, measures showed that subjects were not accurate in their beliefs about their own discrimination ability. Copyright 1999 Academic Press. PMID- 10527815 TI - The Directionality of Verbal Probability Expressions: Effects on Decisions, Predictions, and Probabilistic Reasoning. AB - Verbal phrases denoting uncertainty are of two kinds: positive, suggesting the occurrence of a target outcome, and negative, drawing attention to its nonoccurrence (Teigen & Brun, 1995). This directionality is correlated with, but not identical to, high and low p values. Choice of phrase will in turn influence predictions and decisions. A treatment described as having "some possibility" of success will be recommended, as opposed to when it is described as "quite uncertain," even if the probability of cure referred to by these two expressions is judged to be the same (Experiment 1). Individuals who formulate their chances of achieving a successful outcome in positive terms are supposed to make different decisions than individuals who use equivalent, but negatively formulated, phrases (Experiments 2 and 3). Finally, negative phrases lead to fewer conjunction errors in probabilistic reasoning than do positive phrases (Experiment 4). For instance, a combination of 2 "uncertain" outcomes is readily seen to be "very uncertain." But positive phrases lead to fewer disjunction errors than do negative phrases. Thus verbal probabilistic phrases differ from numerical probabilities not primarily by being more "vague," but by suggesting more clearly the kind of inferences that should be drawn. Copyright 1999 Academic Press. PMID- 10527816 TI - Studies of mesenchymal cells from 1st trimester human placenta: expression of cytokeratin outside the trophoblast lineage. AB - A fibroblast cell strain that expressed cytokeratins 8 and 18 was isolated from explanted first trimester placenta. Its properties were consistent with an origin in the villous fibroblast-myofibroblast lineage, including expression of vimentin, smooth muscle alpha-actin and fibroblast surface protein. The cells grew rapidly in vitro, and exhibited tightly aligned bipolar morphology at confluence, absence of multi-nucleated cells, lack of secreted chorionic gonadotrophin, and absence of HLA G, placental alkaline phosphatase and pregnancy specific beta-1 glycoprotein (SP1). On these criteria it was concluded they were not trophoblasts. On the basis of their morphology, cytokeratin expression and absence of CD34 and endoglin it was concluded they were not endothelial cells. They lacked desmin and smooth muscle myosin and so were not vascular smooth muscle cells. Further mesenchymal cell isolates were studied to determine the generality of these findings. Phenotypic heterogeneity was a consistent characteristic, but cytokeratin-positive cells were always present both in first trimester and term strains. Desmin was absent from almost all the cells isolated using the protocols employed, despite its occurrence in a significant subpopulation of cells in the villous stroma. Cytokeratins 8 and 18 can be observed in the stromal compartment of both first trimester and term placental villi. Cytokeratin has hitherto been regarded as a highly reliable marker for cells of the trophoblast lineage in vitro. These observations suggest that care should be taken in characterization of placental cell isolates; trophoblasts should be identified by the presence of cytokeratin 7 in preference to cytokeratin 8/18. The functional significance of cytokeratin expression in placental mesenchymal cells remains to be established. PMID- 10527817 TI - Connexin expression patterns in human trophoblast cells during placental development. AB - This study focuses on the gap junction expression pattern in trophoblast cells during human placental development in vivo and in vitro. Investigations of cell cell communication properties within the subpopulations of trophoblast responsible for invasion, placental growth and feto-maternal transport seem of special interest because the intercellular channels are believed to coordinate proliferation and differentiation processes. From all gap junction connexins (Cx) investigated (Cx26, Cx31, Cx32, Cx37, Cx40, Cx43), Cx40 was the only connexin clearly detected within the cytotrophoblast of human placenta, and was restricted to the extravillous trophoblast of cell islands and cell columns. Most intense staining was found in the juxtastromal area correlated to the proliferating extravillous trophoblast cells. Connexin protein expression was missing during trophoblast migration into the decidua but was re-expressed in trophoblast aggregates within the decidua. Cx40 expression decreased with progressing pregnancy and no connexins could be detected in villous or extravillous trophoblast of mature placentae. In parallel, isolated trophoblast cells of first and second trimester placentae revealed Cx40 expression and, in contrast to the situation in vivo, Cx43 was also found. In isolated cells of mature placentae, expression of both Cx40 and Cx43 transcripts was decreased to low levels and Cx40 immunoreactivity was absent. Cx43 protein, however, was still detectable in trophoblast cultures of term placentae. Our studies suggest that Cx40 is the characteristic channel for the proliferating cell population of cell islands and cell columns of first and second trimester placentae and isolated trophoblast and is probably involved in regulation and coordination of the invasive pathway. PMID- 10527818 TI - Placental anionic and cationic amino acid transporter expression in growth hormone overexpressing and null IGF-II or null IGF-I receptor mice. AB - The role of growth hormone (GH), insulin-like growth factor (IGF)-II and the IGF I receptor (IGF-Ir) in the regulation of the in vivo expression of Na(+)-coupled anionic [System X-AG; GLAST1 (EAAT1), GLT1 (EAAT2), EAAC1 (EAAT3), EAAT4; where the human homologues of amino acid transport proteins first cloned in the rat are given in parentheses] and Na(+)-independent cationic (System y(+);CAT1) amino acid transport proteins was evaluated by comparing transporter expression in day 17 placentae of mice that overexpressed bovine GH (GH+) or that carried null gene mutations for IGF-II or IGF-Ir. Northern analysis revealed no apparent difference in the mRNA content of GLAST1 (EAAT1), EAAC1 (EAAT3), or EAAT4, in homogenates of GH+ placentae, but levels of GLT1 (EAAT2) and CAT1 mRNA were increased. Immunoblot analysis revealed that whole-placental steady-state GLAST1 (EAAT1), EAAC1 (EAAT3), and EAAT4 protein levels were not affected by GH+, whereas GLT1 (EAAT2) levels were increased. Immunohistochemical analysis showed that the cell specific expression of the anionic and CAT1 transporters was not affected by overexpression of GH. Similar analyses of null IGF-II placentae demonstrated increases in GLAST1 (EAAT1), EAAT4 and CAT1 mRNAs. Parallel immunoblot analysis demonstrated decreased expression of GLT1 (EAAT2), GLAST1 (EAAT1) and EAAC1 (EAAT3) protein, but an increased expression of EAAT4. In null IGF-II and IGF-Ir placentae, however, GLT1 (EAAT2) and EAAC1 (EAAT3) protein content was decreased in junctional zone cells, whereas CAT1 content was increased in junctional and labyrinth zone cells. These data indicate that an excess level of GH stimulates GLT1 (EAAT2) expression and that a normal level of IGF-II is required for typical expression of GLT1 (EAAT2), GLAST1 (EAAT1) and EAAC1 (EAAT3), but that IGF-II downregulates the expression of EAAT4 and CAT1. PMID- 10527819 TI - Characterization of glucose transport and glucose transporters in the human choriocarcinoma cell line, BeWo. AB - In this study we have characterized 2-deoxyglucose (2DG) transport and hexose transporter expression in the human choriocarcinoma cell line, BeWo. 2DG uptake in BeWo cells displayed saturable kinetics (V(max), 29+/-1.5 nmol/min/mg protein;K(m), 1.5+/-0.02 m m) and was significantly inhibited in the presence of 2-deoxyglucose, mannose and 3- O -methyl glucose (all at a competing concentration of 30 m m) by up to 97 per cent, but not by galactose or fructose. Glucose uptake was not Na(+)-dependent, but was inhibited by cytochalasin B (by approx 85 per cent) indicating that hexose uptake was mediated via a facilitative glucose transport mechanism. Northern and immunoblot analyses revealed that BeWo cells expressed GLUT1 and GLUT5, but not GLUT2 or GLUT3. On immunoblots, GLUT1 migrated as a broad protein band on SDS-gels (average M(r)of 55 kDa) and treatment with N -glycanase resulted in a significant shift in its electrophoretic mobility; the core protein migrating as a 40 kDa band indicating that the carrier was heavily glycosylated. GLUT5 was detected as a discrete 60 kDa band and like GLUT1, the observed immunoreactive signal was lost when using antiserum that had been pre-adsorbed with the antigenic peptide. Our findings indicate that BeWo cells express a facilitative glucose transport system with characteristics broadly similar to those reported in isolated human placental membrane vesicles and that they are likely to serve as a useful experimental system for studying the regulation of placental glucose transport and transporter expression. PMID- 10527820 TI - The effects of angiogenic growth factors on extravillous trophoblast invasion and motility. AB - There is accumulating evidence that deficient trophoblast invasion of the placental bed spiral arteries is crucial to the pathogenesis of pre-eclampsia and intrauterine growth restriction. However, the factors which regulate the process of trophoblast invasion remain unclear. We have investigated whether extravillous trophoblast invasion and motility are mediated by the angiogenic growth factors, vascular endothelial growth factor (VEGF) and placental growth factor (PlGF). The SGHPL-4 extravillous trophoblast cell line was utilized. Expression of mRNA for the receptors of VEGF and PlGF (KDR and flt-1) was determined using the reverse transcriptase polymerase chain reaction. An in vitro model of invasion assessed the number and length of trophoblast processes invading into an extracellular matrix. The motility of cells under standard culture conditions was also quantified. The effect of the addition of VEGF and PlGF (+/-heparin) on trophoblast invasion and motility was determined. The effect of VEGF and PlGF (+/ heparin) on SGHPL-4 cell proliferation was assessed by cell counts at 24, 48 and 72 h post-addition of growth factor. The SGHPL-4 cells expressed mRNA for the flt 1 but not the KDR receptor. The addition of VEGF resulted in a significant decrease in the number of trophoblast processes formed (P< 0.02); this effect was not influenced by the addition of heparin. However, there was no effect on the length of processes formed in response to VEGF (+/-heparin). The addition of PlGF had no effect on either the number or the length of processes formed. The addition of VEGF increased the motility of the SGHPL-4 cells (P< 0.002); the addition of heparin prevented this VEGF-induced increase in motility. The addition of PlGF had no effect on SGHPL-4 motility (+/-heparin). Neither growth factor had any effect on the proliferative ability of SGHPL-4 cells. Contrary to our hypothesis, we did not find that the angiogenic growth factors, VEGF and PlGF, mediated the in vitro invasion of trophoblast cells into an extracellular matrix. However, VEGF did increase trophoblast motility. Our findings of an effect of VEGF on trophoblast motility (and possibly invasion) suggests the presence of functional receptors, which can mediate the actions of VEGF. Caution must be exercised before any extrapolation to the in vivo situation, however, it could be speculated that the increased motility in response to VEGF may be an initial response to attract trophoblast cells to the decidua, and that VEGF might then limit the degree to which trophoblast cells invade. PMID- 10527821 TI - Oestradiol stimulates morphological and functional differentiation of human villous cytotrophoblast. AB - Trophoblast differentiation is a complex process involving interactions of cytotrophoblastic cells with their evolutive milieu. During pregnancy, the feto placental unit produces large amounts of steroids. Progesterone and oestradiol are increasingly produced when the syncytiotrophoblast is highly differentiated. Furthermore, receptors to these hormones are expressed by the trophoblast. This led us to test the hypothesis that steroid production could affect the morphological and functional differentiation of the trophoblast during gestation. The fusion of cytotrophoblastic cells into syncytiotrophoblast was assessed using fluorescence recovery after photobleaching for gap junctional communication analysis (gap-FRAP), desmoplakin immunostaining and connexin 43 expression. In parallel, functional differentiation was assessed by beta-human chorionic gonadotrophin (betahCG) production and human chorionic somatomammotropin (hCS) expression analysis. The presence of oestradiol, 1 microm, increased the percentage of coupled cells (3. 8-fold), connexin 43 expression and stimulated the syncytium formation. In parallel, oestradiol (1, 3 and 5 microm) induced a significant increase in the daily hCG production. The steroid action was specific, as the stimulatory effects were inhibited by tamoxifen. Oestradiol also stimulated hCS expression (51 per cent compared to control after 3 days). As trophoblastic differentiation is specifically stimulated by hCG, oestradiol could act via the stimulation of hCG production or via a direct action. In the presence of an efficient concentration of hCG antibody, oestradiol still stimulated hCS expression, suggesting a self-sufficient effect of the steroid. Physiological concentrations of progesterone were ineffective in modulating trophoblast differentiation. In conclusion, oestradiol could be implicated in the maturation and aging of the trophoblast. PMID- 10527822 TI - Leptin receptor in human term placenta: in situ hybridization and immunohistochemical localization. AB - In the present study, we investigated the expression and localization of leptin receptors in human term placentae. On human term placenta tissue slices, digoxigenin-UTP labelled RNA-probe detected the long form of the leptin receptor ObR(L)mRNA in syncytiotrophoblasts of the villi, whereas the haematological subtype of the leptin receptor ObR/B219.1 was detected in blood cells of the intervillous space and fetal vessels. Immunohistochemistry, with two polyclonal antibodies to the N-terminus recognizing ObR(L)and ObR(S)of the leptin receptors and one to the C-terminus recognizing the long form of the leptin receptor ObR(L), localized leptin receptor protein at the apical membrane of the syncytiotrophoblasts. Our results show that the long form of the leptin receptor ObR(L)is expressed in human term placentae. We localized the long form of leptin receptor mRNA to the cytoplasm of syncytiotrophoblasts and leptin receptor proteins in human term placentae to the apical membrane of syncytiotrophoblasts. We conclude that in term placentae, leptin could mediate a growth promoting effect in the fetoplacental unit through the long form of the leptin receptor localized in the syncytiotrophoblasts. In contrast, the haematological subtype of the leptin receptor is not expressed in placental cells, but solely by blood cells in the intervillous space and fetal vessels. PMID- 10527823 TI - Stimulation of DNA synthesis in trophoblasts and human umbilical vein endothelial cells by hepatocyte growth factor bound to extracellular matrix. AB - Hepatocyte growth factor (HGF) promotes the growth not only of hepatocytes but also of several other types of cells such as cytotrophoblasts and endothelial cells. Recent studies have revealed that HGF is trapped in the extracellular (ECM) matrix through heparan sulphate in vivo, thereby acting as a mitogen for hepatocytes in cooperation with heparan sulphate. In this study, we detected HGF protein in chorionic tissue and placental tissue extracts, and found that HGF and heparan sulphate were co-distributed in the endothelial basement membrane and trophoblast basement membrane on immunohistochemical examination. The rates of DNA synthesis in primary cultured cytotrophoblasts and human umbilical vein endothelial cells (HUVEC) cultured on HGF-bound Matrigeltrade mark were 6-8 times those of control cytotrophoblasts and HUVEC. When Matrigeltrade mark dishes were pretreated with heparinase and heparitinase prior to binding of HGF, stimulation of DNA synthesis was markedly decreased. A considerable decrease in stimulation of DNA synthesis was observed following washing of HGF-bound Matrigeltrade mark with 1 m acetic acid, 1 m NaCl and 0.1 per cent trypsin, but not following treatment with chondroitinase ABC. These observations suggest that HGF can be trapped in ECM in vivo, thereby acting as a mitogen for cytotrophoblasts and placental vein endothelial cells in cooperation with heparan sulphate. PMID- 10527824 TI - Effect of proline rich polypeptide from ovine colostrum on virus replication in human placenta and amniotic membrane at term; possible role of endogenous tumour necrosis factor alpha. AB - Freshly prepared organ cultures of human placentae and amniotic membranes at term show different sensitivity to vesicular stomatitis virus (VSV) infection. In six of 16 amniotic membranes and seven of 17 placentae VSV replicated to relatively high titres (10(3)-10(6)TCID(50)/ml). The others were partially or completely resistant to virus infection (<10(1)-10(2)TCID(50)/ml). Addition of the immunomodulating agent, proline-rich-polypeptide (PRP) from ovine colostrum to explants freshly obtained from the organs, influenced VSV replication in a manner dependent on the innate immune state of the organ culture. In cultures resistant to the virus, PRP at a concentration of 10 microg/ml increased 10-10 000 times the VSV titre. In contrast, treatment of highly sensitive cultures by PRP hardly influenced viral replication at all. The effect of virus stimulation by PRP was abolished by specific anti-TNF antibodies. The results indicate that endogenous TNF may be one of the mediators of virus stimulation by PRP. Antibodies against TNFalpha, added to VSV infected organ cultures sensitive to the virus reduced viral replication. The antibodies caused stimulation of virus replication in VSV infected resistant organ cultures. The results indicate the double role of endogenous TNF in viral replication in placenta and the amniotic membrane. PMID- 10527825 TI - Isolation and sequence analysis of a cDNA encoding human placental tissue protein 13 (PP13), a new lysophospholipase, homologue of human eosinophil Charcot-Leyden Crystal protein. AB - Expression of placental tissue protein 13 (PP13) in different human tissues was investigated by chemiluminescence Western blot analysis using monospecific anti PP13 serum. In term placentae we detected a 16 kDa single protein band immunochemically identical to the purified PP13 antigen. After investigation of 26 types of human fetal and adult tissue, PP13 was also found in certain other normal and tumorous tissue extracts. It is not secreted into circulation as we could not find PP13 in sera of pregnant women. A full length cDNA with 578 bp insert was isolated by screening a human placental cDNA library with anti-PP13 serum. The open reading frame of the cDNA encodes for a 139-residue-long protein with a predicted molecular mass of 16.118 kDa, identical to the previously isolated and characterized PP13 antigen described in 1983. By alignment search of the protein databank PP13 is highly homologous (69 per cent) to the 16.5 kDa human eosinophil Charcot-Leyden Crystal protein, a unique dual-function lysophospholipase, a member of the beta-galactoside binding S-type animal lectin superfamily. Northern blot analysis revealed a 600 bp PP13 mRNA, detected only in placental tissue from 16 types of human healthy adult tissue. Lysophospholipase activity of PP13 was confirmed by(1)H and(31)P nuclear magnetic resonance (NMR) measurements. PMID- 10527826 TI - Localization and significance of urokinase plasminogen activator and its receptor in placental tissue from intrauterine, ectopic and molar pregnancies. AB - Urokinase plasminogen activator receptor (uPAR) is a membrane-anchored protein with urokinase plasminogen activator (uPA) as the ligand. This complex induces proteolysis and remodelling of maternal decidua during placental implantation. The presence of uPAR on trophoblasts is supposed to promote adhesion, migration and invasion. In cancer tissue, high levels of uPAR are correlated with a poor prognosis. This immunohistochemical study shows the localization of uPA and uPAR in a prospective design with stereological sampling of fetal and maternal tissue from normal, ectopic and hydatidiform molar (HM) pregnancies. Cytokeratin and Ki67 were used as markers for trophoblasts and proliferating cells. Membrane bound uPAR was observed on villous non-proliferating intermediate trophoblasts (IT) within cell columns in intrauterine and ectopic pregnancies. The corresponding proliferating IT with cytological atypia sprouting from the chorionic villi in HM was uPAR-negative. uPA but not uPAR was observed in anchoring distal IT at the attachment-point to the basal plate. In the placental bed, extravillous interstitial trophoblasts were uPA-positive but uPAR-negative. The trophoblast giant cells were both uPA- and uPAR-negative. In relation to the maternal vessels, a focal distribution for uPA and uPAR was present in the endovascular and perivascular trophoblasts. The intraluminal trophoblasts overlying endothelial cells were uPAR-positive only. In maternal tissue from intrauterine and molar pregnancies, uPAR was seen in the decidual cells in a zone facing the anchoring villi and the fibrinoid lesions with embedded trophoblasts. In contrast, the stromal cells of the fallopian tube without a decidual reaction facing the implanted gestation were uPAR-negative. Non-invaded decidual, myometrial and muscular tissue of the pregnant uterus and fallopian tube was extensively positive for uPA whereas 'pseudodecidual' cells from the intrauterine evacuate in patients with an ectopic pregnancy only showed a focal and scanty reaction for uPA. When trophoblast invasion of the decidua was present, the decidual cells were uPA-negative. A semi-quantitative assessment of the receptor was estimated in villous IT within cell columns in normal and molar pregnancies but, in conclusion, quantitative evaluation of uPAR cannot be used to predict development of post-molar persistent trophoblastic disease (PTD). PMID- 10527827 TI - The neonatal implications of a high placental ratio in small-for-gestational age infants. AB - An increased placental ratio has been associated with small-for-gestational age (SGA) infants. A retrospective study on 252 singleton SGA infants without major anomalies born within a 1-year period was performed to determine the relationship between placental ratio and maternal/infant characteristics, and perinatal complications. The cases were categorized into three groups according to the placental ratio (<1 sd below the mean, within 1 sd of the mean, >1 sd above the mean) based on our previous data. There were more infants with a high ratio (32.9 per cent) than with a low ratio (15.5 per cent). While there was no difference in the maternal characteristics or antenatal complications, there was a significant trend in decreasing birthweight and an increasing placental weight in relation to an increasing placental ratio. The infants with a high ratio had increased incidence of meconium stained liquor, hypocalcaemia, hypomagnesaemia and phototherapy, a trend that was consistent even after exclusion of the preterm infants. Our data indicated that a high placental ratio in SGA infants was due to both increased placental size and decreased birthweight, and this was associated with increased neonatal morbidity. PMID- 10527828 TI - SHORT COMMUNICATION maternal thyroid dysfunction and c- fos and c- jun expression in rat placenta. AB - Maternal thyroid dysfunction is associated with perturbed fetal brain development and neurological deficits in adulthood in rat and human. To investigate whether these effects occur secondary to placental dysfunction, c- fos and c- jun expression in placenta from normal (euthyroid) and moderately hypothyroid rat dams were investigated by Northern hybridization analysis. In normal placenta, c- fos expression increased by 74 per cent between 16 and 21 days of gestation (dg) whereas c- jun expression declined by 46 per cent. Moderate maternal hypothyroidism depressed placental c- fos expression by 32 per cent at 19 dg, but elevated c- fos and c- jun expression by 139 and 86 per cent, respectively, at 21 dg. Maternal hypothyroidism may therefore induce c- fos/c- jun -related placental dysfunction, but only relatively late in gestation when fetal thyroid function is already established. PMID- 10527829 TI - CONTENTS OF VOLUME 20. PMID- 10527830 TI - Protection of MLV vector particles from human complement. AB - Murine cell-derived MLV vector particles usually are highly sensitive to human complement-mediated lysis. Expression of the human complement inhibitor CD59 on murine packaging cells resulted in partial protection of these cells from lysis caused by human complement proteins. Furthermore, CD59 was incorporated into MLV vector particles released by these packaging cells, leading to an improved resistance of the virions against human complement-mediated inactivation. PMID- 10527831 TI - Real-time study of interactions between a composite DNA regulatory region (HIV-1 LTR NRE) and several transcription factors of nuclear extracts. AB - Here we describe the first real-time study of nuclear protein interaction with a composite DNA regulatory region. We studied the interplay between the three target sites of the negative regulatory element (NRE) of HIV-1 LTR, comprising a noncanonical GATA site overlapping two negative regulatory regions, USF and NFIL 6, and their corresponding transcription factors in nuclear extracts. By bandshift analysis, no GATA binding activity could be detected between LTR NRE and different nuclear extracts, although evidenced by in vitro footprinting. Additionally, the LTR NRE and a USF oligonucleotide showed identical retarded complexes. BIAcore study of these interactions revealed the binding of huGATA-3, as well as USF, to the immobilized LTR NRE oligonucleotide. Competition analyses, performed with GATA, USF, and NFIL-6 oligonucleotides, clearly showed that this regulatory region could bind both huGATA-3 and USF factors. Finally, the presence of USF and huGATA-3 proteins in the complexes formed with LTR NRE was ascertained using specific anti-huGATA-3 and anti-USF2 polyclonal antibodies. PMID- 10527832 TI - Presence and expression of a novel variant form of ST2 gene product in human leukemic cell line UT-7/GM. AB - A novel variant cDNA from the human ST2 gene other than ST2 or ST2L was identified and tentatively named ST2V. Alternative splicing inserts a new exon which leads to a change in the C-terminal portion of ST2, causing it to gain a hydrophobic tail instead of losing the third immunoglobulin-like domain. ST2V is expressed in human leukemic cell line UT-7 and its sublines UT-7/GM, UT-7/EPO, and UT-7/TPO, in addition to human helper T cell line 5C10. The amount of ST2V mRNA is greatly diminished when UT-7/GM cells are induced to differentiate into either erythroblastic or megakaryoblastic phenotypes. The possible roles of the ST2V in growth and differentiation are intriguing. PMID- 10527833 TI - Bovine lactoferrin peptidic fragments involved in inhibition of herpes simplex virus type 1 infection. AB - Bovine lactoferrin (BLf) prevents the infection of some enveloped and naked viruses. To identify BLf sequences responsible for the antiviral activity, we tested 31 HPLC fractions, derived from tryptic digestion of BLf, toward herpes simplex virus type 1 (HSV-1). Only a few HPLC purified fragments were active against HSV-1, even if at lower extent than the native undigested BLf. Two large fragments, one corresponding to the C-lobe (amino acid sequence 345-689) and the other corresponding to a large portion of the N-lobe (1-280), were inhibitors of HSV-1 infection, while a smaller part of the N-lobe (86-258) was ineffective. Among the low-molecular-weight fragments, only two small peptides, which coeluted in a single chromatographic peak, were effective towards HSV-1. These peptides, both present in the N-lobe, were identified as peptides 222-230 (ADRDQYELL) and 264-269 (EDLIWK). The same peptides, chemically synthesised, were able to inhibit HSV-1 infection only when they were assayed in association. PMID- 10527834 TI - Cloning, sequencing, and characterization of ribosomal protein and RNA polymerase genes from the region analogous to the alpha-operon of escherichia coli in halophilic archaea, halobacterium halobium. AB - A determination was made of the nucleotide sequence of the 3215-bp region of a ribosomal protein gene cluster (HS13, HS4, HS11, and HeL18), RNA polymerase (RNA poly D), and tRNA genes (tRNAser and tRNAarg) of halophilic Archaea Halobacterium halobium, which is analogous to the alpha-operon of Escherichia coli (tRNAser HS13-HS4-HS11-RNA poly D-tRNAarg-HeL18). The seven-gene string was preceded by a pseudoknot-like structure similar to the proposed S4 ribosomal protein binding site of the alpha-operon mRNA leader in E. coli. Using an inducible expression system H. halobium HS4 was produced in large amounts in E. coli, and immunoblot analysis showed the S4 to constitute a 21-kDa polypeptide component of the ribosome. Analysis of the deduced amino acids sequence revealed that the HS13, HS4, and HS11 sequences including the RNA polymerase subunit are more similar to their eukaryotic than to their bacterial counterparts. HeL18, located downstream of the gene cluster analogous to the E. coli alpha-operon (S13-S11-S4-RNA poly D L17), was similar to both the eukaryotic (eL18) and eubacterial ribosomal protein L15 located in the spc-operon, but not to L17 positioned as the terminal gene of the bacterial alpha-operon. PMID- 10527835 TI - Expression of cholecystokinin type A receptor gene correlates with DNA demethylation during postnatal development of rat pancreas. AB - Cholecystokinin stimulates pancreatic amylase secretion, gallbladder contraction, and pancreatic growth, etc. by binding with high affinity to a cholecystokinin type A receptor (CCKAR). To better understand the expression of CCKAR mRNA in terms of tissue specificity and postnatal development, we determined the methylation status of BssHII sites (5'-B sites) in the rat CCKAR gene promoter. The 5'-B sites in adult pancreas expressing CCKAR mRNA were much less extensively methylated than those in fetal pancreas not expressing the mRNA. In brain, liver, and kidney of adult rats not expressing CCKAR mRNA, the 5'-B sites were methylated. In pancreas, the demethylation level of the sites increased at 21 days after birth. Concomitant with the DNA demethylation level in the 5'-B sites, the mRNA level rose rapidly in 21 days. These results demonstrate that methylation and expression of the CCKAR gene reveal a good inverse correlation. PMID- 10527836 TI - Cocaine potentiates the switch between latency and replication of Epstein-Barr virus in Raji cells. AB - This paper shows that cocaine amplifies Epstein-Barr virus (EBV) reactivation in Raji cells. Its effect on early viral protein synthesis was maximal when it was added with 12-O-tetradecanoyl phorbol-13-acetate (TPA) plus n-butyrate, but nil when added alone. The enhancing effect of cocaine on early replicative stages of latent EBV was associated with an increase of Ca(2+) mobilization induced by the drug and with an induction of cellular protein phosphorylation in chemicals and cocaine-treated Raji cells. Cocaine also acted synergistically with TPA and n butyrate to induce Z Epstein-Barr replication activator (ZEBRA), a nuclear phosphoprotein responsible for the activation of early viral gene expression. These findings provide the first evidence that cocaine may represent an important co-factor in the reactivation of early stages of latent EBV infection. PMID- 10527837 TI - Unique characteristics of ubiquitin-bonded complex play a pathological role in dentatorubral-pallidoluysian atrophy. AB - Abnormal complex formation of dentatorubral-pallidoluysian atrophy (DRPLA) protein and pathological ubiquitination of abnormal complex are two pathological processes involved in DRPLA neurodegeneration. Pathological ubiquitination and solubility in SDS and reducing agent are two unique characteristics of the DRPLA protein complex. Ubiquitination of abnormal DRPLA protein complex in DRPLA brain tissue is heat-resistant and stronger than that in control brain tissue. Pathological ubiquitination of DRPLA protein complex correlates with the onset of symptoms and the size of an expanded glutamine repeat in brain tissue of patients with DRPLA. Pathological ubiquitination plays an important role in DRPLA pathology. DRPLA protein complex is water-insoluble but soluble in SDS and reducing agent, and displays no difference in water insolubility between control and DRPLA brain tissue. Abnormal insoluble complex formation is not developed by a qualitative change in water insolubility of DRPLA protein complex but is developed by a spontaneous accumulation of an abnormally large amount of the DRPLA protein complex. PMID- 10527838 TI - Attachment of C-terminus of SDF-1 enhances the biological activity of its N terminal peptide. AB - The N-terminus of stromal cell-derived factor 1 (SDF-1) is known to be a critical site for CXCR4 receptor binding and signaling. However, the functional role of other regions, in particular the C-terminal helix of SDF-1, has yet to be defined. In this study, we designed and synthesized a peptide model of SDF-1 containing its N- and C-terminal regions. The attachment of the C-terminus of SDF 1, which by itself had no activity in receptor binding and signaling, dramatically increased the effect of the N-terminal fragment in inducing chemotaxis and intracellular Ca(2+) influx in sup T1 cells compared with the peptide containing only the N-terminal sequence. The enhancement in activity was not due to the increase in receptor affinity as the N,C-terminal peptide did not show higher CXCR4 binding than the N-terminal peptide. On the other hand, the intracellular Ca(2+) influx activated by the N,C-terminal peptide, but not the N terminal peptide, was completely abolished by the addition of heparin, suggesting that the C-terminal fragment of the peptide binds glycosaminoglycans (GAGs) and exerts an effect to modulate biological activity. These data raise the possibility that the C-terminus in native SDF-1 is one of interaction sites with GAGs and may be associated with biological function of SDF-1. Furthermore, this study demonstrates an approach for the design of novel agonists or antagonists of other chemokine receptors that possess enhanced biological activity. PMID- 10527839 TI - Association between an alpha(2) macroglobulin DNA polymorphism and late-onset Alzheimer's disease. AB - An association between a five-base-pair deletion/insertion DNA polymorphism at the alpha(2) macroglobulin gene (A2M) and late-onset Alzheimer's disease (LOAD) has been recently described. We developed a PCR assay to analyze this polymorphism in 190 LOAD patients (older than 65 years) and 400 controls from Spain. Controls were stratified into three groups: <65 years (n = 200), 65 to 80 years (n = 100), and 81 years or older (n = 100). We found a significantly higher frequency of carriers of the D allele in patients older than 81 years compared to controls older than 81 years (p = 0.0012). In addition, the frequency of the D allele was significantly lower in controls older than 81 years compared to controls younger than 65 (p = 0.048). Our work suggests that the D allele confers an age-dependent increased risk to develop late-onset Alzheimer's disease. PMID- 10527840 TI - Era, an essential Escherichia coli small G-protein, binds to the 30S ribosomal subunit. AB - Era is an essential G-protein in Escherichia coli identified originally as a homologue protein to Ras (E. coli Ras-like protein). It binds to GTP/GDP and contains a low intrinsic GTPase activity. Its function remains elusive, although it has been suggested that Era is associated with the cytoplasmic membrane, cell division, energy metabolism, and cell-cycle check point. Recently, a cold sensitive phenotype was found to be suppressed by the overexpression of 16S rRNA methyltransferase, suggesting Era association with the ribosome. Here we demonstrate that Era specifically binds to 16S rRNA and the 30S ribosomal subunit. Both GTP and GDP, but not GMP, inhibit Era binding to ribosomal component. Involvement of Era in protein synthesis is suggested by the fact that Era depletion results in the translation defect both in vitro and in vivo. PMID- 10527841 TI - BSMAP, a novel protein expressed specifically in the brain whose gene is localized on chromosome 19p12. AB - Using the sequence of cosmids derived from chromosome 19p12, we have identified a gene encoding a novel protein, BSMAP (brain-specific membrane-anchored protein) and cloned cDNA encoding the full-length open reading frame. Northern blot analysis revealed that BSMAP mRNA is preferentially expressed at a high level in the brain. BSMAP has a putative transmembrane domain and is predicted to be a type-I membrane glycoprotein. Genomic sequence analysis revealed that the gene encoding BSMAP consists of eight exons spanning approximately 8 kb and lies 6 kb away from the gene encoding CLF-1 in a reverse orientation. Although no candidate genetic disorders were found to map either to this precise region of chromosome 19 or to the syntenic region of the mouse genome, the highly specific expression of BSMAP mRNA suggests a role for the protein in CNS function. PMID- 10527842 TI - Selective lysis of virus-infected cells by cobra snake cytotoxins: A sendai virus, human erythrocytes, and cytotoxin model. AB - By using a Sendai virus-human erythrocyte model, this work found that virus infected cells were 10-fold more susceptible to lysis in two of five examined cobra venoms. Four cytotoxins were isolated from the venom of the cobra Naja nigricollis that also showed 10-fold higher cytotoxicity toward virus-infected cells than to untreated cells. As selective destruction of virus-infected cells is of immense importance in clinical practice, this work demonstrates the potential of cobra cytotoxins to serve as leading compounds for the generation of derivatives or fractions with high cytotoxic specificity toward virus-infected cells. PMID- 10527843 TI - Hierarchical order of critical residues on the immunity-determining region of the Im7 protein which confer specific immunity to its cognate colicin. AB - The directed mutagenesis study of the Im7 protein of colicin E7 revealed that three residues, D31, D35, and E39, located in the loop 1 and helix 2 regions of the protein were critical for initiating the complex formation with its cognate colicin E7. Interestingly, the importance of these three critical residues in conferring specific immunity to its own colicin was exhibited in a hierarchical order, respectively. Moreover, we found that existence of the three critical residues was common among the DNase-type Im proteins. Most likely the three residues of the DNase-type immunity proteins are critical for initiating the unique protein-protein interactions with their cognate colicin. In addition, replacement of the helix 2 of Im7 by the corresponding region of Im8 produced a phenotype of the mutant protein very similar to that of Im8. This result suggests that the DNase-type Im proteins indeed share a "homologous-structural framework" and evolution of the Im proteins may be engendered by minor amino acid changes in this specific immunity-determining region without causing structural alteration of the proteins. PMID- 10527844 TI - An expanded CTG trinucleotide repeat causes trans RNA interference: a new hypothesis for the pathogenesis of myotonic dystrophy. AB - Here we report a novel mechanism for the pathogenesis of myotonic dystrophy (DM). The DMPK mRNA with expanded CTG trinucleotide repeats interacts with other transcripts having expanded CAG repeats. This "trans RNA interference" occurs in vitro only when the number of CTG repeats is over 140 and the number of target CAG repeats exceeds 35. The trans RNA interference can explain all the phenomena previously reported about DM. PMID- 10527845 TI - A tetratricopeptide repeat-containing protein gene, tpis, whose expression is induced with differentiation of spermatogenic cells. AB - The tetratricopeptide repeat (TPR) is a degenerate 34-amino-acid sequence which forms scaffolds to mediate protein-protein interactions. We have isolated a cDNA named tpis from mouse embryonic skin and found that the deduced 529-amino-acid sequence contained 5 TPRs. In addition to skin, the transcript of tpis was detected in tissues with stratified squamous epithelium, e.g., tongue, esophagus, and forestomach. tpis was most strongly expressed in testis among adult tissues examined. The transcript of tpis from testis was longer, encoding 372 additional amino acid residues at the 5'-side with 3 more TPRs. In situ hybridization revealed specific expression of tpis at a distinct differentiation stage of spermatogenic cells, indicating involvement of tpis in spermatogenesis. Chromosomal localization of the tpis gene was determined as 18.10 cM of chromosome 15. PMID- 10527846 TI - Molecular cloning of a human MafF homologue, which specifically binds to the oxytocin receptor gene in term myometrium. AB - The US-2 DNA-binding element (ggaatgattactcagctaga) in the promoter of the human oxytocin receptor (OTR) gene has been shown to bind specifically nuclear proteins from human myometrium at parturition. To elucidate the molecular mechanisms involved in OTR gene upregulation at term, the US-2 element was used in a yeast one-hybrid system to screen a cDNA library derived from term human myometrium. Positive clones were further screened by electrophoretic mobility shift assay for their ability to bind the human OTR gene promoter, containing the US-2 motif. A 2.3-kb full-length cDNA encoding a human homologue of chicken MafF (hMafF) was isolated. hMafF represents an 18-kDa protein and contains an extended leucine zipper structure, but lacks a transactivation domain. Furthermore, Northern hybridization showed strong hMafF mRNA expression in the kidney and in term myometrium only, but not in nonpregnant myometrium. The hMafF protein is also preferentially expressed in term myometrium, as shown by specific binding to the OTR promoter. The highly specific binding of hMafF to the US-2 motif in the human OTR gene, together with its pattern of expression, supports a role for hMafF in OTR gene upregulation at term. PMID- 10527847 TI - Inhibition of adipogenesis by a COOH-terminally truncated mutant of PPARgamma2 in 3T3-L1 cells. AB - Peroxisome proliferator-activated receptor gamma (PPARgamma) is a nuclear receptor that is thought to be an important regulator of adipocyte differentiation. This ligand-dependent transcription factor is also activated by thiazolidinediones, a new class of synthetic antidiabetic drugs, resulting in a marked adipogenic response in cultured cells and enhanced insulin sensitivity in vivo. The importance of the COOH-terminal region of PPARgamma2 in thiazolidinedione-induced adipogenesis has now been investigated by expression of a mutant protein (PPARgamma2-DeltaC) that lacks the COOH-terminal 16 amino acids of full-length PPARgamma2. The mutant protein failed to bind a thiazolidinedione ligand, but its ability to bind the peroxisome proliferator response element was similar to that of the wild-type protein. Expression of PPARgamma2-DeltaC inhibited the thiazolidinedione-induced increase in trans-activation activity of endogenous PPARgamma in CV-1 cells. Furthermore, the mutant protein prevented thiazolidinedione-induced adipogenesis in 3T3-L1 cells, whereas expression of recombinant wild-type PPARgamma2 promoted adipogenesis. These data show not only that the COOH-terminal region of PPARgamma2 is indispensable for thiazolidinedione-induced adipogenesis mediated by this protein in 3T3-L1 cells, but also that the PPARgamma2-DeltaC mutant acts in a dominant negative manner by interfering with the access of endogenous PPARgamma to the peroxisome proliferator response element of target genes. PMID- 10527848 TI - Hemolysate induces tyrosine phosphorylation and collagen-lattice compaction in cultured fibroblasts. AB - Hemolysate, a proposed causative agent for cerebral vasospasm after subarachnoid hemorrhage, produces contraction of cerebral arteries by activation of tyrosine kinases. In addition, hemolysate increases fibroblast-collagen compaction that could play a role in cerebral vasospasm. We studied the effect of hemolysate on tyrosine phosphorylation and fibroblast-collagen compaction in cultured canine basilar and human dermal fibroblasts using tyrosine kinase inhibitors and tyrosine antibodies. Hemolysate enhanced tyrosine phosphorylation of two proteins, 64 and 120 kDa, in cultured canine basilar artery and human dermal fibroblast cells. The effect of hemolysate was time-dependent and concentration dependent. Oxyhemoglobin and ATP, the two major components of hemolysate, produced similar tyrosine phosphorylation, however, with a different time course. Tyrosine kinase inhibitors genistein and tyrphostin A51 abolished the effect of hemolysate in both cerebral and dermal fibroblasts. Hemolysate increased fibroblast-populated collagen-lattice compaction and tyrosine kinase inhibitors genistein and tyrphostin A51 attenuated the effect of hemolysate. We conclude that hemolysate activates tyrosine kinase that leads to the increase of fibroblast compaction. This effect of hemolysate may contribute to cerebral vasospasm. PMID- 10527849 TI - Farnesyl pyrophosphate synthase is the molecular target of nitrogen-containing bisphosphonates. AB - Bisphosphonates (Bps), inhibitors of osteoclastic bone resorption, are used in the treatment of skeletal disorders. Recent evidence indicated that farnesyl pyrophosphate (FPP) synthase and/or isopentenyl pyrophosphate (IPP) isomerase is the intracellular target(s) of bisphosphonate action. To examine which enzyme is specifically affected, we determined the effect of different Bps on incorporation of [(14)C]mevalonate (MVA), [(14)C]IPP, and [(14)C]dimethylallyl pyrophosphate (DMAPP) into polyisoprenyl pyrophosphates in a homogenate of bovine brain. HPLC analysis revealed that the three intermediates were incorporated into FPP and geranylgeranyl pyrophosphate (GGPP). In contrast to clodronate, the nitrogen containing Bps (NBps), alendronate, risedronate, olpadronate, and ibandronate, completely blocked FPP and GGPP formation and induced in incubations with [(14)C]MVA a 3- to 5-fold increase in incorporation of label into IPP and/or DMAPP. Using a method that could distinguish DMAPP from IPP on basis of their difference in stability in acid, we found that none of the NBps affected the conversion of [(14)C]IPP into DMAPP, catalyzed by IPP isomerase, excluding this enzyme as target of NBp action. On the basis of these and our previous findings, we conclude that none of the enzymes up- or downstream of FPP synthase are affected by NBps, and FPP synthase is, therefore, the exclusive molecular target of NBp action. PMID- 10527850 TI - Molecular cloning of the gene for p85 that regulates the initiation of cytokinesis in Tetrahymena. AB - Tetrahymena p85 is localized to the presumptive division plane before division furrow formation; its molecular weight in SDS-polyacrylamide gel electrophoresis differs in wild-type and temperature-sensitive cell-division-arrest mutant cdaA1 cells. At the restrictive temperature, p85 localization and division furrow formation are not observed in cdaA1 cells. In this study, we purified p85 and cloned a wild-type p85 cDNA. The deduced amino acid sequence of p85 was composed mainly of two kinds of repeat sequences. One of these contained regions homologous to a calmodulin-binding site and a part of actin, and the other contained a region homologous to a part of a cdc2 kinase homologue. Moreover, we cloned a cDNA encoding the cdaA1 p85. There was no difference in the predicted amino acid sequences of wild-type and cdaA1 p85, suggesting that the difference in molecular weight between p85 in wild-type and mutant cells is caused by a disorder of posttranslational-modification mechanisms of p85 in the cdaA1 cell. PMID- 10527851 TI - Human chromosomal localization, tissue/tumor expression, and regulatory function of the ets family gene EHF. AB - Ets factors are members of an ancient multigene family of transcription factors including oncoproteins and possibly tumor suppressors. We previously characterized a novel divergent ets gene, Ehf (ets homologous factor) in mice. Here we report the cDNA sequence, chromosomal location, and tissue/tumor expression patterns of the human EHF gene and the regulatory activity of the EHF protein. EHF maps to 11p12, which is deleted in many prostate, breast, and lung carcinomas and is a hot spot for inherited deletion- or amplification-associated developmental defects. EHF is differentially expressed in normal tissues and carcinomas and between tumor stages and is most highly expressed in the organs known to form carcinomas upon 11p12 deletion. EHF protein represses the ETS-2 induced activity of both stromelysin-1 and collagenase-1 promoters. These data suggest that EHF may contribute to human development and carcinogenesis and is a candidate for the 11p12 tumor suppressor gene. PMID- 10527852 TI - The beta4 integrin subunit rescues A431 cells from apoptosis through a PI3K/Akt kinase signaling pathway. AB - To study whether alpha6beta4 integrin regulates apoptosis, human A431 cells were plated on bacteria plates in the presence or absence of mAb beta4. In the absence of mAb beta4, A431 cells demonstrated morphological characteristics of apoptosis by 24 h and most cells died by 48 h. In contrast, in the presence of mAb beta4, cells remained viable, and at the end of 48 h, 70-80% of cells survived. Treatment of A431 cells with mAb beta4 resulted in tyrosine phosphorylation of the p85 subunit of PI3 kinase; PI3 kinase activity increased within 15 min and peaked at 60 min. Stimulation of beta4 in A431 cells resulted in a time-dependent phosphorylation of Akt with a concomitant and parallel phosphorylation of Bad. Inactivation of PI3 kinase with inhibitors blocked the anti-apoptotic effect induced by mAb beta4. These are the first results to suggest that ligation of alpha6beta4 integrin protects cells from apoptosis through a PI3K/Akt kinase signaling pathway. PMID- 10527853 TI - In vitro effects of angiopoietins and VEGF on hematopoietic and endothelial cells. AB - To investigate the role of TIE-2/angiopoietins (Angs) in postnatal hematopoiesis, we cultured bone marrow cells in the presence of Ang-1 and -2 with or without VEGF. While either Ang-1 or Ang-2 alone had hematopoietic effects on unseparated bone marrow cells and no effect on proliferation of endothelial cells, both enhanced the growth of endothelial cells and hematopoietic progenitor cells in the presence of VEGF. FACS analysis showed that Lin(-)TIE-2(+)Flk-1(+) cells cocultured with OP9 stromal cells gave rise to endothelial and hematopoietic cells in the presence of VEGF and Ang-1. Ang-1 promoted the adhesion of sorted primary Lin(-)TIE-2(+) cells to fibronectin, and this adhesion enhanced proliferation of hematopoietic progenitor cells synergistically with stem cell factor (SCF). Our findings suggest that TIE-2/angiopoietins may act as critical regulators of proliferation of hematopoietic progenitors and endothelial cells in a synergistic manner. PMID- 10527854 TI - The cell death regulatory protein bak is expressed in endothelial cells in inflamed tissues and Is induced by IFN-gamma in vitro. AB - In the present report, we examined the endothelial expression of the anti- and pro-apoptotic proteins Bcl-2 and Bak in situ and in vitro. Endothelial cells (EC) in regular tissue of the bowel and the skin were essentially negative for both Bcl-2 and Bak. In contrast, EC within the walls of fistulas and abscesses in these organs stained distinctly for Bak, but remained Bcl-2-negative. In tissue culture both unstimulated human dermal microvascular endothelial cells (HDMEC) and human umbilical vein endothelial cells (HUVEC) expressed Bcl-2 and Bak constitutively. Exposure of EC to 200-1000 IU IFN-gamma downregulated Bcl-2 but upregulated Bak. This opposing regulation of Bcl-2 and Bak in vitro and the expression of Bak in EC adjacent to necrotic tissue areas suggests that this pro apoptotic protein may play a decisive role in regulation of EC survival in vivo. PMID- 10527855 TI - CRM1 mediates nuclear export of nonstructural protein 2 from parvovirus minute virus of mice. AB - The nonstructural protein 2 (NS2) from parvovirus minute virus of mice (MVMp) is a 25-kDa polypeptide which localizes preferentially to the cytoplasm and associates with cellular proteins in cytoplasm. These lines of evidence suggest that NS2 is positively exported from the nucleus to cytoplasm and functions in cytoplasm. We report here that nuclear export of NS2 is inhibited by leptomycin B (LMB), a drug that specifically blocks nuclear export signal (NES)-chromosomal region maintenance 1 (CRM1) interactions. CRM1 binds specifically to the 81- to 106-amino-acid (aa) region of NS2, and the region of NS2 actually functions as a NES. Interestingly, this region appears to be distinct from a typical NES sequence, which consists of leucine-rich sequences. These results indicate that NS2 protein is continuously exported from the nucleus by a CRM1-dependent mechanism and suggest that CRM1 also exports to distinct type of NESs. PMID- 10527856 TI - Molecular cloning of a novel Xenopus spalt gene (Xsal-3). AB - The sal (spalt) gene family is characterized by unique double zinc finger motifs and is conserved among various species from Drosophila to humans. Here we report a new Xenopus member of this family, Xsal-3. It is 38% homologous at the amino acid level to the previously reported Xenopus homologue of the spalt gene, Xsal 1. Alternatively spliced Xsal-3 transcripts give rise to RNAs coding either two or three double zinc fingers, and the longer form is expressed maternally. Xsal-3 is expressed in the neural tube, the mandibular, hyoid, and branchial arch, and the pronephric duct, which is different from the expression pattern of Xsal-1. These findings suggest that Xsal-3 may have distinct roles in early Xenopus development. PMID- 10527857 TI - Identification of aquaporin 5 (AQP5) within the cochlea: cDNA cloning and in situ localization. AB - Dysfunction of fluid and electrolyte homeostasis is considered to cause variety of inner ear disorders. One group of candidate proteins that may play a critical role in the inner ear fluid homeostasis is the aquaporins, a family of proteins whose members have well defined roles in fluid transport in variety of organs. This study reports the identification of AQP5, a member of the aquaporin family, within the rat inner ear and its in situ localization. AQP5 was initially identified within rat cochlear RNA via RT-PCR and sequence analysis of the amplified fragments. Immunoblot of cochlear homogenate yielded a predominant AQP5 immunoreactive band of M(r) 35 kDa. The anti-AQP5 immunoreactivity, indicating expression of the AQP5 polypeptide, was localized within the cochlea in situ to the cell types that form the lateral wall of the cochlear duct-the external sulcus (ES) cells and the cells of the spiral prominence. Expression of AQP5 was observed in the apical turn but not the basal turn of the cochlea; nor was it observed in the vestibular neuroepithelia or its supporting cells. The restricted expression of AQP5 to the apical turns of the cochlea suggests its potential role in low frequency hearing. PMID- 10527859 TI - Phenylalanine 138 in the second intracellular loop of human thromboxane receptor is critical for receptor-G-protein coupling. AB - Eicosanoid receptors exhibit a highly conserved ERY(C)XXV(I)XXPL sequence in the second intracellular loop. The carboxyl end of this motif contains a bulky hydrophobic amino acid (L,I,V, or F). In human thromboxane A2 receptor (TXA(2)R), phenylalanine 138 is located at the carboxyl end of this highly conserved motif. This study examined the function of the F138 in G protein coupling. F138 was mutated to aspartic acid (D) and tyrosine (Y), respectively. Both mutants F138D and F138Y showed similar ligand binding activity to that of the wild type TXA(2)R. The Kd and Bmax values of either mutant were comparable to those of the wild type receptor. However, both mutants showed significant impairment of agonist induced Ca(2+) signaling and phospholipase C activation. These results suggest that the F138 plays a key role in G protein coupling. PMID- 10527858 TI - Novel association of the src family kinases, hck and c-fgr, with CCR3 receptor stimulation: A possible mechanism for eotaxin-induced human eosinophil chemotaxis. AB - The chemokine eotaxin is a potent and relatively eosinophil-specific chemoattractant implicated in the cell migration to inflammatory sites in allergic diseases. Eotaxin exerts its activity solely through the CCR3 receptor, but the signaling pathways are poorly defined. In this study, we show that eotaxin induces an increase in tyrosine phosphorylation of multiple cellular proteins in normal human eosinophils. Eotaxin-dependent tyrosine phosphorylation was detected 1 min after stimulation and increased for at least 15 min with kinetics similar to those of eotaxin-induced cell shape changes. Herbimycin A, a tyrosine kinase inhibitor, blocked both eotaxin-induced tyrosine phosphorylation and cell shape changes as well as chemotaxis. Immunofluorescence microscopy analyses showed that eotaxin-induced cell shape changes were accompanied by redistribution of tyrosine-phosphorylated proteins and F-actin reorganization that were sensitive to herbimycin A. Coimmunoprecipitation studies revealed that binding of eotaxin to CCR3 greatly enhanced association of the Src family kinases, Hck and c-Fgr, with CCR3 after internalization of CCR3. These results may indicate that recruitment of Hck and c-Fgr to CCR3 in a compartment triggers tyrosine phosphorylation, leading to rapid cell shape changes required for cell migration. PMID- 10527860 TI - Study of DFF45 in its role of chaperone and inhibitor: two independent inhibitory domains of DFF40 nuclease activity. AB - CAD/DFF40, the nuclease responsible for DNA fragmentation during apoptosis, exists as a heterodimeric complex with DFF45/ICAD. This study determines the molecular mechanisms of regulation of DFF40 via the chaperone and inhibition activities of DFF45. We analyze proteins corresponding to the fragments (D1, D2, and D3) of DFF45 generated by cleavage at the caspase consensus sites in DFF45. Either D1 or D2, as an isolated domain, is capable of inhibiting DFF40 nuclease activity while double domain fragments D1-2 and D2-3, as well as full-length DFF45, bind to DFF40 with high affinity and are much more effective inhibitors. The chaperone activity of DFF45 resides in part in its ability to maintain DFF40 as a soluble protein. In addition, D1 of DFF45 was found to be critical for the expression of active DFF40 in vivo, suggesting a role for DFF45 in binding nascent DFF40. PMID- 10527861 TI - Multiple domains of DFF45 bind synergistically to DFF40: roles of caspase cleavage and sequestration of activator domain of DFF40. AB - CAD/DFF40, the nuclease responsible for DNA fragmentation during apoptosis, exists as a heterodimeric complex with DFF45/ICAD. The study presented here augments the accompanying inhibition and chaperone study with an analysis of specific binding strengths and locations of DFF45 binding sites within DFF40. This allows us to show that DFF40/45 interaction is mediated by binding of three functional domains (D1, D2, and D3) of DFF45 to two domains (activator and catalytic) of DFF40. D1 binds exclusively to the activator domain and D2 binds to the catalytic domain of DFF40. Inhibition of DFF40 nuclease activity arises independently from D1 functional sequestration of the activator domain and D2 blockage of the catalytic domain of DFF40. The mechanism of caspase activation of DFF40 is the disruption of the synergistic binding activity of DFF45 domains to DFF40 after caspase recognition and cleavage of DFF45 in the context of a DFF45/40 complex. PMID- 10527862 TI - Regulated expression of the nifM of Azotobacter vinelandii in response to molybdenum and vanadium supplements in Burk's nitrogen-free growth medium. AB - Azotobacter is a diazotrophic bacterium that harbors three genetically distinct nitrogenases referred to as nif, vnf, and anf systems. The nifM is an accessory gene located in the nif gene cluster and is transcriptionally regulated by the NifA. However, Azotobacter mutants that lack NifA are known to synthesize functional NifM and this accessory protein is known to be needed for the activity of nitrogenase-2 and nitrogenase-3. To determine how the transcription of nifM is regulated when Azotobacter is grown under conditions in which nitrogenase-2 or nitrogenase-3 is expressed, we generated an Azotobacter vinelandii strain that carries a nifM:lacZ-kanamycin resistance gene cassette in its chromosome. In this strain the nifM open reading frame was disrupted by the presence of a lacZ kanamycin resistance gene cassette so that it could not produce active NifM. Moreover, the lacZ gene was placed under the transcriptional control elements of the nifM gene so that the lacZ expression could be used as a marker to determine the extent of expression of the nifM gene under different growth conditions. Our results show that this strain was unable to grow in Burk's nitrogen-free medium supplemented with either molybdenum or vanadium or lacking both metals suggesting that in the absence of functional NifM none of the nitrogenases were active. It was also found that the nifM expression was differentially regulated when the A. vinelandii cells were grown under conditions that activate nitrogenase-2 and nitrogenase-3, as determined by liquid beta-galactosidase activity measurements. These results suggest that the transcriptional activators, VnfA and AnfA, may regulate the nifM expression. PMID- 10527863 TI - Identification of two penicillin-binding multienzyme complexes in Haemophilus influenzae. AB - Dansyl-labeled penicillin, reversed-phase chromatography, and peptide mapping have been used to detect, separate, and study penicillin-binding proteins (PBPs) and PBP multienzyme complexes of H. influenzae. The cross-linking of proteins in the multienzyme complex was accomplished with the aid of cyanogen, a salt-bridge specific cross-linking agent. The chromatographic profile of the PBPs clearly showed a dramatic change in the number and identity of peaks after treatment of the bacterial cells with cyanogen. The disappearance of all seven peaks corresponding to the PBPs was accompanied by the emergence of two new peaks with molecular weights between 400 kDa and 600 kDa. The results hint at the existence of two penicillin-binding multienzyme complexes, each containing subunits that interact via salt-bridges. Chromatographic active site peptide mapping of PBPs and PBP complexes was used to determine the identity of PBPs involved in each complex. It is postulated that one multienzyme complex containing PBP 2 may be involved in cell elongation while the other complex containing PBP 3 may be responsible for cell division. PMID- 10527864 TI - Evidence that histidine-163 is critical for catalytic activity, but not for substrate binding to Escherichia coli agmatinase. AB - Agmatinase (agmatine ureohydrolase, EC 3.5.3.11) from Escherichia coli was inactivated by diethyl pyrocarbonate (DEPC) and illumination in the presence of Rose bengal. Protection against photoinactivation was afforded by the product putrescine, and the dissociation constant of the enzyme-protector complex (12 mM) was essentially equal to the K(i) value for this compound acting as a competitive inhibitor of agmatine hydrolysis. Upon mutation of His163 by phenylalanine, the agmatinase activity was reduced to 3-5% of wild-type activity, without any change in K(m) for agmatine or K(i) for putrescine inhibition. The mutant was insensitive to DEPC and dye-sensitized inactivations. We conclude that His163 plays an important role in the catalytic function of agmatinase, but it is not directly involved in substrate binding. PMID- 10527865 TI - Characterization of recombinant fungal phytase (phyA) expressed in tobacco leaves. AB - The phyA gene from Aspergillus ficuum coding for a 441-amino-acid full-length phytase was expressed in Nicotiana tabacum (tobacco) leaves. The expressed phytase was purified to homogeneity using ion-exchange column chromatography. The purified phytase was characterized biochemically and its kinetic parameters were determined. When the recombinant phytase was compared with its counterpart from Aspergillus ficuum for physical and enzymatic properties, it was found that catalytically the recombinant protein was indistinguishable from the native phytase. Except for a decrease in molecular mass, the overexpressed recombinant phytase was virtually the same as the native fungal phytase. While the temperature optima of the recombinant protein remain unchanged, the pH optima shifted from pH 5 to 4. The results are encouraging enough to open the possibility of overexpressing phyA gene from Aspergillus ficuum in other crop plants as an alternative means of commercial production of this important enzyme. PMID- 10527866 TI - Conversion of lysine to N(epsilon)-(carboxymethyl)lysine increases susceptibility of proteins to metal-catalyzed oxidation. AB - Metal-catalyzed oxidation (MCO) of proteins leads to the conversion of some amino acid residues to carbonyl derivatives, and may result in loss of protein function. It is well documented that reactions with oxidation products of sugars, lipids, and amino acids can lead to the conversion of some lysine residues of proteins to N(epsilon)-(carboxymethyl)lysine (CML) derivatives, and that this increases their metal binding capacity. Because post-translational modifications that enhance their metal binding capacity should also increase their susceptibility to MCO, we have investigated the effect of lysine carboxymethylation on the oxidation of bovine serum albumin (BSA) by the Fe(3+)/ascorbate system. Introduction of approximately 10 or more mol CML/mol BSA led to increased formation of carbonyls and of the specific oxidation products glutamic and adipic semialdehydes. These results support the view that the generation of CML derivatives on proteins may contribute to the oxidative damage that is associated with aging and a number of age-related diseases. PMID- 10527867 TI - EPR detection of reactive oxygen species in hemolymph of Galleria mellonella and Dendrolimus superans sibiricus (Lepidoptera) larvae. AB - The formation of reactive oxygen species (ROS) in hemolymph and hemocytes of Galleria mellonella and Dendrolimus superans sibiricus larvae was studied by ESR spectroscopy using spin-trap 1-hydroxy-3-carboxy-pyrrolidine (CP-H). The background level of ROS formation was detected in the intact hemolymph. The addition of dihydroxyphenylalanine (DOPA) into free cells of the hemolymph increased CP-H oxidation about two times for G. mellonella and about four times for D. superans sibiricus. This increase was completely inhibited by a specific inhibitor of phenoloxidase, phenylthiourea. The presence of exogenous superoxide dismutase (SOD) did not change CP-H oxidation in the hemolymph. The data obtained in hemocytes showed only a DOPA-induced increase in CP-H oxidation. Phagocytosis activators did not affect ROS formation in hemocytes of both insect species. SOD decreased DOPA-induced CP-H oxidation 20-30% in suspension of hemocytes of D. superans sibiricus only. Our results are in agreement with the contribution of superoxide radical and DOPA-derived quinones/semiquinones in the immune response of insects. PMID- 10527868 TI - A key driving force in determination of protein structural classes. AB - The three-dimensional structure of a protein is uniquely dictated by its primary sequence. However, owing to the very high degenerative nature of the sequence structure relationship, proteins are generally folded into one of only a few structural classes that are closely correlated with the amino-acid composition. This suggests that the interaction among the components of amino acid composition may play a considerable role in determining the structural class of a protein. To quantitatively test such a hypothesis at a deeper level, three potential functions, U((0)), U((1)), and U((2)), were formulated that respectively represent the 0th-order, 1st-order, and 2nd-order approximations for the interaction among the components of the amino acid composition in a protein. It was observed that the correct rates in recognizing protein structural classes by U((2)) are significantly higher than those by U((0)) and U((1)), indicating that an algorithm that can more completely incorporate the interaction contributions will yield better recognition quality, and hence further demonstrate that the interaction among the components of amino acid composition is an important driving force in determining the structural class of a protein during the sequence folding process. PMID- 10527869 TI - Fragments from alpha-actinin insert into reconstituted lipid bilayers. AB - Recent experiments have indicated that alpha-actinin interacts with phospholipid membranes. Using computer analysis methods we determined two possible lipid binding sites capable of membrane attachment/insertion, residues 281-300 and 720 739 of the primary amino acid sequence on smooth muscle alpha-actinin. Having expressed these regions as fusion proteins with schistosomal GST (glutathione S transferase), we used differential scanning calorimetry (DSC) to investigate their interaction with mixtures of zwitterionic (dimyristoyl-l-alpha phosphatidylcholine, DMPC) and anionic (dimyristoyl-l-alpha-phosphatidylglycerol, DMPG) phospholipids in reconstituted lipid bilayers. Calorimetric measurements showed that as fusion protein concentration increased, the main chain transition enthalpy decreased and chain melting temperatures shifted, which is indicative of partial protein insertion into the hydrophobic region of the lipid membranes. Centrifugation assay and subsequent SDS/Page chromatography confirmed this finding. PMID- 10527870 TI - Estradiol-17beta stimulates the renewal of spermatogonial stem cells in males. AB - In this work, we examined the functions of the female hormone "estrogen" on spermatogenesis of the Japanese eel (Anguilla japonica). Estradiol-17beta (E(2)), a natural estrogen in vertebrates, was present in the serum and its receptor was expressed in the testis during the whole process of spermatogenesis. Spermatogonial stem cell renewal was promoted by E(2) implantation but was suppressed by tamoxifen (an antagonist of estrogen). In vitro, 10 pg/ml of E(2) was sufficient to induce spermatogonial stem cell division in cultured testicular tissue, therefore confirming the in vivo observations. These results clearly show that estrogen is an indispensable "male hormone" in the early spermatogenetic cycle. PMID- 10527871 TI - Molecular cloning of chick UCH-6 which shares high similarity with human UCH-L3: its unusual substrate specificity and tissue distribution. AB - A full-length cDNA encoding ubiquitin C-terminal hydrolase-6 (UCH-6) was isolated from the chick skeletal muscle cDNA library. The sequence of two peptides generated from purified UCH-6 matched perfectly with the predicted amino acid sequence. Nucleotide sequence analysis of the cDNA containing an open reading frame of 690 base pairs revealed that the protease consists of 230 residues with a calculated molecular mass of 26,315 Da. UCH-6 belonged to members of the UCH family containing highly conserved Cys, His, and Asp domains and showed 86% amino acid identity to human UCH-L3. Interestingly, most tissues examined contained significant amounts of UCH-6 mRNA, while human UCH-L3 is expressed only in the brain, lungs, and red cells. Moreover, UCH-6, unlike other UCH family enzymes including UCH-L3, could release free ubiquitin from ubiquitin-beta-galactosidase fusion proteins both in vivo and in vitro. The ubiquitous expression pattern and unusual substrate specificity of UCH-6 suggest that the enzyme may represent a distinct subfamily of UCH-L3. PMID- 10527872 TI - Serine phosphorylation of the sarcoplasmic reticulum Ca(2+)-ATPase in the intact beating rabbit heart. AB - Recent studies have demonstrated that Ca(2+)/calmodulin-dependent protein kinase phosphorylates the Ca(2+)-pumping ATPase of cardiac sarcoplasmic reticulum (SR) in vitro. Also, evidence from in vitro studies suggested that this phosphorylation, occurring at Ser(38), results in stimulation of Ca(2+) transport. In the present study, we investigated whether serine phosphorylation of the SR Ca(2+)-ATPase occurs in the intact functioning heart. Hearts removed from anesthetized rabbits were subjected to retrograde aortic perfusion of the coronary arteries with oxygenated mammalian Ringer solution containing (32)P(i) and contractions were monitored by recording systolic left ventricular pressure development. Following 45-50 min of (32)P perfusion, the hearts were freeze clamped, SR isolated, and analyzed for protein phosphorylation. SDS polyacrylamide gel electrophoresis and autoradiography showed phosphorylation of several peptides including the Ca(2+)-ATPase and Ca(2+) release channel (ryanodine receptor). The identity of Ca(2+)-ATPase as a phosphorylated substrate was confirmed by Western immunoblotting as well as immunoprecipitation using a cardiac SR Ca(2+)-ATPase-specific monoclonal antibody. The Ca(2+)-ATPase showed immunoreactivity with a phosphoserine monoclonal antibody indicating that the in situ phosphorylation occurred at the serine residue. Quantification of Ca(2+) ATPase phosphorylation in situ yielded a value of 208 +/- 12 pmol (32)P/mg SR protein which corresponded to the phosphorylation of approximately 20% of the Ca(2+) pump units in the SR membrane. Since this phosphorylation occurred under basal conditions (i.e., in the absence of any inotropic intervention), a considerable steady-state pool of serine-phosphorylated Ca(2+)-ATPase likely exists in the normally beating heart. These findings demonstrate that serine phosphorylation of the Ca(2+)-ATPase is a physiological event which may be important in the regulation of SR function. PMID- 10527874 TI - Enhancement of dendrimer-mediated transfection using synthetic lung surfactant exosurf neonatal in vitro. AB - Pulmonary surfactants enhance adenovirus-mediated gene transfer but inhibit cationic liposome-mediated transfection in lung epithelial cells in vitro. This study examines the effect of the synthetic lung surfactant Exosurf on dendrimer mediated transfection in eukaryotic cells. Exosurf significantly enhanced dendrimer-luciferase plasmid transfection in a number of cell lines and was very effective in primary cells. Luciferase expression increased up to 40-fold in primary normal human bronchial/tracheal epithelial cells (NHBE). FACScan analysis demonstrated that the transfection rate of the human T cell leukemia Jurkat cell line has significantly improved from 10 to 90% of cells at 24 h after transfection. Analysis of the components of Exosurf revealed that the nonionic surfactant tyloxapol was responsible for the enhancement of dendrimer-mediated gene transfer. The tyloxapol effect was due to increased cell membrane porosity and DNA uptake. Our results demonstrate that Exosurf and its component, tyloxapol, constitute a powerful enhancer for dendrimer-mediated gene transfer in vitro. PMID- 10527873 TI - Proline-rich synapse-associated proteins ProSAP1 and ProSAP2 interact with synaptic proteins of the SAPAP/GKAP family. AB - We have recently isolated a novel proline-rich synapse-associated protein-1 (ProSAP1) that is highly enriched in postsynaptic density (PSD). A closely related multidomain protein, ProSAP2, shares a highly conserved PDZ (PSD-95/discs large/ZO-1) domain (80% identity), a ppI domain that mediates the interaction with cortactin, and a C-terminal SAM (sterile alpha-motif) domain. In addition, ProSAP2 codes for five ankyrin repeats and a SH3 (Src homology 3) domain. Transcripts for both proteins are coexpressed in many regions of rat brain, but show a distinct expression pattern in the cerebellum. Using the PDZ domains of ProSAP1 and 2 as bait in the yeast two-hybrid system, we isolated several clones of the SAPAP/GKAP (SAP90/PSD-95-associated protein/guanylate kinase-associated protein) family. The association of the proteins was verified by coimmunoprecipitation and cotransfection in HEK cells. Therefore, proteins of the ProSAP family represent a novel link between SAP90/PSD-95 bound membrane receptors and the cytoskeleton at glutamatergic synapses of the central nervous system. PMID- 10527875 TI - Formation of oxygen radicals in solutions of 7,8-dihydroneopterin. AB - Neopterin and 7,8-dihydroneopterin, two compounds which are secreted by activated macrophages, have been shown to interfere with radicals generated by cellular and certain chemical systems. Reduced pterins were reported to scavenge whereas aromatic pterins promoted or reduced radical mediated reactions or had no effect. However, recently it was found that high concentrations of 7, 8-dihydroneopterin enhanced luminol dependent chemiluminescence and T-cell apoptosis, suggesting an enhancement of free radical formation. In this study hydroxylation of salicylic acid was used for detection of hydroxyl radicals. It is shown that in solutions of 7,8-dihydroneopterin hydroxyl radicals were formed in the absence of any radical source. The presence of EDTA chelated iron enhanced hydroxyl radical formation. Whereas the addition of iron accelerated the hydroxylation reaction, 7,8-dihydroneopterin was responsible for the amount of hydroxylation products. In the presence of superoxide dismutase or catalase, as well as by helium purging, hydroxylation was inhibited. Our data suggest that in solutions of 7, 8 dihydroneopterin superoxide radicals are generated which are converted to hydroxyl radicals by Fenton or Haber-Weiss type reactions. While superoxide might be generated during autoxidation of ferrous iron, dihydroneopterin seems to be involved in regeneration of ferrous iron from the ferric form. PMID- 10527877 TI - Stimulation of macrophages by mucins through a macrophage scavenger receptor. AB - We found that phorbol ester-primed THP-1 cells (a human monocyte cell line), which express a scavenger receptor, were stimulated by mucins through the macrophage scavenger receptor, resulting in enhanced secretion of IL-1beta. The activity was abolished by treatment of the mucins with sialidase, indicating that sialic acid is involved in binding. (125)I-Labeled ovine submaxillary mucin could bind to COS 7 cells transfected with cDNA encoding the scavenger receptor. Binding was inhibited by mucins, fucoidan, and polyinosinic acid but not by polycytidylic acid, this being consistent with the characteristics of the scavenger receptor. When phorbol ester-primed THP-1 cells were cocultured with colon cancer cells producing mucins, IL-1beta secreted from the THP-1 cells increased significantly. Adhesion between colon cancer cells and a scavenger receptor transfectant was observed, and binding was inhibited partly by mucins and ligands for the scavenger receptor. PMID- 10527876 TI - Interaction of measles virus (Halle strain) with CD46: evidence that a common binding site on CD46 facilitates both CD46 downregulation and MV infection. AB - CD46 acts as a cellular receptor for vaccine strains of measles virus (MV). The MV/CD46 interaction-mediated by the MV attachment glycoprotein, the hemagglutinin (H)-not only facilitates infection but also induces CD46 downregulation. A conflict of opinion exists as to whether a single MVH binding site on CD46, or two separate sites, facilitates the two phenomena. To investigate this conundrum we first tested and compared a panel of CD46-specific monoclonal antibodies (mAbs) for their capacity to block both processes. One (mAb 13/42) abrogated both MV fusion and CD46 downregulation. Mutation of an amino acid (arg59 in the SCR1 of CD46) essential for the epitope of mAb 13/42 resulted in the abrogation of both CD46 downregulation and viral fusion. This strongly suggests that the same MV binding site on CD46 is responsible for both CD46 downregulation and MV infection. PMID- 10527878 TI - Effects of tryptophan residues of porcine myeloid antibacterial peptide PMAP-23 on antibiotic activity. AB - PMAP-23 is a 23-residue antimicrobial peptide from porcine myeloid cells. In order to determine the effects of two Trp residues in positions 7 and 21 of PMAP 23 on antibacterial activity and phospholipid vesicle interacting property, two analogues in which Ala is substituted for Trp residue in position 7 or 21 were synthesized. A(21)-PMAP-23 exhibited reduced antibacterial activity and phospholipid vesicle disrupting activity when compared to those of PMAP-23 and A(7)-PMAP-23. PMAP-23 readily interacted with model lipid membrane and induced membrane destabilization. Therefore antibacterial activity induced by PMAP-23 is due to the interaction of cell membrane with peptide followed by membrane perturbation. A significant structural change on the SDS micelle was not found by Ala substitution of the Trp residue of PMAP-23. Also, there is a good correlation between hydrophobic interaction on RP-HPLC, expressed as retention time on RP HPLC, and antibacterial activity. The vesicle titration experiment indicated that Trp residues located at near C-terminus are accessible to hydrophobic tail of phospholipid vesicle. This result suggests that the C-terminal end of PMAP-23 penetrates into the lipid bilayer in the course of the interaction with phospholipid membranes and is important for its antibacterial activity. PMID- 10527879 TI - Residues within the HFRIGC sequence of HIV-1 vpr involved in growth arrest activities. AB - HIV-1 Vpr is a virion-associated protein that can cause growth arrest when produced inside the cell but when added externally it can cause cell death. Employing the yeast model system, the C-terminal domain, in particular the sequence HFRIGCRHSRIG (Vpr(71-82)), is essential for both the growth arrest and cytocidal activities. Conservation of this sequence in HIV-2 and SIV suggests that these residues may be functionally important. Using site-directed mutagenesis we show that the most highly conserved aa residues, His71 and Gly75, were important for the cell cycle inhibitory effects. In contrast, we show that the wild-type Vpr(71-82) peptide and three variants of this peptide with Gly75 changed to Ser, Ala, and Ile all exhibited the same cytocidal activity suggesting that the intracellular and extracellular effects are unrelated. PMID- 10527880 TI - Cloning and immunolocalization of a rat pancreatic Na(+) bicarbonate cotransporter. AB - In the rat, pancreatic HCO(-)(3) secretion is believed to be mediated by duct cells with an apical Cl(-)/HCO(-)(3) exchanger acting in parallel with a cAMP activated Cl(-) channel and protons being extruded through a basolateral Na(+)/H(+) exchanger. However, this may not be the only mechanism for HCO(-)(3) secretion by the rat pancreas. Recently, several members of electrogenic Na(+)/HCO(-)(3) cotransporters (NBC) have been cloned. Here we report the cloning of a NBC from rat pancreas (rpNBC). This rpNBC is 99% identical to the longer, more common form of NBC [pNBC; 1079 amino acids (aa); 122 kDa in human heart, pancreas, prostate, and a minor clone in kidney]. The longer NBC isoforms are identical to the rat and human kidney-specific forms (kNBC; 1035 aa; 116 kDa) at the approximately 980 C-terminal aa's and are unique (with different lengths) at the initial N-terminus. Using polyclonal antibodies to the common N- and C termini of rat kidney NBC, a approximately 130-kDa protein band was labeled by immunoblotting of rat pancreas homogenate and was enriched in the plasma membrane fraction. Immunofluorescence and immunoperoxidase light microscopy of rat pancreatic tissue with both antibodies revealed basolateral labeling of acinar cells. Labeling of both apical and basolateral membranes was found in centroacinar cells, intra- and extralobular duct, and main duct cells. The specificity of the antibody labeling was confirmed by antibody preabsorption experiments with the fusion protein used for immunization. The data suggest that rpNBC likely plays a more important role in the transport of HCO(-)(3) by rat pancreatic acinar and duct cells than previously believed. PMID- 10527881 TI - Identification and cDNA cloning of headpin, a novel differentially expressed serpin that maps to chromosome 18q. AB - Differential display was used to identify a novel serpin (headpin) underexpressed in squamous cell cancers of the oral cavity. Headpin cDNA encoding a complete open reading frame was cloned and sequenced. Headpin is expressed in normal oral mucosal tissue, skin, and cultured keratinocytes. Using Northern analysis and relative reverse-transcription polymerase chain reaction (relative RT-PCR), downregulation of headpin mRNA expression was demonstrated in oral cavity squamous carcinomas. Northern blot analysis identified a 3. 3-kb headpin mRNA transcript. Headpin is a 391-amino-acid protein with a theoretical molecular weight of 44 kDa. Hinge region homology at the reactive site loop suggests that headpin belongs to the inhibitory class of serine protease inhibitors. Headpin was mapped to 18q21.3/18q22. This region includes the ovalbumin serpins (ov serpins) maspin, SCCA1, SCCA2, and PAI-2. Furthermore, 18q is recognized as a region for frequent loss of heterozygosity (LOH) in head and neck cancer and other malignancies. PMID- 10527882 TI - The N-terminal half of NPM dissociates from nucleoli of HeLa cells after anticancer drug treatments. AB - NPM (nucleophosmin/B23) is a nucleolar phosphoprotein abundant in tumor cells. It dissociates from nucleoli of cells after treatments with various anticancer drugs. To determine the domain of NPM responsible for nucleolar binding, the N- and C-terminal halves of NPM were fused to GFP (green fluorescent protein) and introduced into HeLa cells. The N-terminal half (aa 1-150) of NPM (GFP-NPM(N)) was found localized in the nucleoli. A stable transformant of GFP-NPM(N) in HeLa cells was prepared and tested for association to nucleoli after anticancer drug treatments. GFP-NPM(N) dissociates from nucleoli after treatments with daunomycin, actinomycin D, camptothecin, and toyocamycin. The dissociation is time- and dose-dependent, and correlates with the cytotoxicity induced by the drugs. These results indicate that a stable transformant of GFP-NPM(N) in HeLa cells may be useful for the screening of anticancer drugs. PMID- 10527883 TI - Identification and validation of tumor suppressor genes. AB - Cancers are associated with frequent deletions of genetic material that select for the loss of genes regulating normal cellular physiology. Although several cancer suppressor genes have been identified from these areas of deletion, the identities of the vast majority remain unknown, making approaches leading to their localization, identification, and validation an important continuing endeavor. Those currently characterized cancer suppressors include regulators of aspects of the cell cycle, growth and transcriptional regulators, DNA repair enzymes, differentiation factors, cell motility elements, and regulators of signal transduction. Several inherited cancer predisposition genes have been mapped and cloned using meiotic genetic linkage mapping but less success has been achieved identifying those genes involved in nonfamilial cancer. The future localization, identification, and validation of these genes are likely to involve a combination of complementary position-oriented and function-driven approaches, some of which are detailed in this article. PMID- 10527884 TI - Inhibition of polymorphonuclear leukocyte activation by acetylcholinesterase. AB - Previously we had shown that basement membrane collagen (COL IV), and specifically residues 185-203 of the non-collagenous domain of the alpha3(IV) chain, inhibits PMN activation. Since acetylcholinesterase (AchE) possesses collagenous and non-collagenous domains, we tested its effect on PMN activation. Whole AchE and the AchE recombinant catalytic subunit inhibited PMN superoxide anion (O2-) generation. AchE synthetic peptides, residues 139-154 and 252-266 of the catalytic subunit, with sequence homology to that of the alpha3(IV) peptide also inhibited O2- production by PMN. Reactive pAb and mAb to the alpha3(IV) 185 203 peptide abolished the inhibitory effect of the AchE. The data show that the non-collagenous domain of the AchE down-regulates O2- production by PMN. We suggest that this inhibitory activity may serve as a protective mechanism against PMN-mediated injury at the level of vessel wall and the neuromuscular junction. PMID- 10527885 TI - Molecular characterization of mitochondria in asexual and sexual blood stages of Plasmodium falciparum. AB - Molecular mechanisms that regulate gene expression during development of asexual stage to sexual stage of Plasmodium falciparum in the human erythrocyte are largely unknown. There were apparent variations in ultrastructural characteristics of the mitochondrion between the two developing stages. The asexual stage's mitochondrion had developed less than that of the sexual stage. The respiratory complexes of the mitochondrial electron transport system in the asexual stage were approximately 8-10 times less active than those in the sexual stage. Using quantitative polymerase chain reaction to amplify the cytochrome b gene encoding a subunit of mitochondrial cytochrome c reductase, the amount of the cytochrome b gene of the sexual stage was calculated to be approximately 3 times higher than that obtained from the asexual stage. Moreover, using quantitative reverse-transcription polymerase chain reaction, a relatively high level of approximately 1.3-kb transcript mRNA of the cytochrome b gene was observed in the sexual stage compared to the asexual stage. A known single-copy chromosomal dihydrofolate reductase gene was found to have a similar amount in the two stages. These results suggest that the copy number of the mitochondrial gene, including transcriptional and translational mechanisms, plays a major regulatory role in differential expression during the development of the asexual to sexual stage of P. falciparum in the human cell. PMID- 10527886 TI - Structural features and expression of an alternatively spliced growth factor type I receptor for follitropin signaling in the developing ovary. AB - The pleiotropic actions of pituitary follitropin (FSH), regulate the expression of many cell cycle genes controlling ovarian follicular development and differentiation. In this study we asked the question whether different receptor motifs are created by the alternative splicing of the single large 80-100 Kb receptor gene. A 1.2 Kb transcript identified from a cDNA library of hormone primed (immature) sheep ovaries, codes for a putative protein lacking the seven transmembrane segment. The receptor of 259 amino acids designated FSH-R3 is derived from a transcript comprising the first eight exons of the Gs coupled larger FSH receptor (R1) spliced to another DNA segment. This event produces a different carboxyl terminus at the junction creating a novel receptor motif with a single membrane spanning domain, assigning it to the growth factor type I receptor family. In transfected cells the expressed receptor localizes on the cell surface and specific antibodies directed against the unique C-terminal portion (residues 242-259) of FSH-R3 demonstrate the presence of the receptor protein in solubilized ovarian and testicular membrane preparations. FSH binding to the transfected cells induced [Ca2+]i identifying coupling of the R3 receptor to calcium signaling pathways. Thus, a growth factor type I receptor for FSH may be implicated in the growth promoting actions of FSH in the ovary. This is the first documentation of alternative splicing of a G protein coupled receptor gene creating a different signaling motif for cellular signaling. PMID- 10527887 TI - Identification of Src as a novel atypical protein kinase C-interacting protein. AB - Atypical protein kinase C-zeta (PKC-zeta) participates in nerve growth factor (NGF) signaling and is required for NGF-induced differentiation of PC12 cells. The biological activity of PKC-zeta is likely mediated by interaction of PKC-zeta with specific proteins. Affinity column chromatography employing the PKC-zeta regulatory domain coupled to glutathione-agarose was used to search for proteins that bound PKC-zeta. Two proteins (59/60 kDa) were recovered from NGF-stimulated PC12 cell lysates that bound the matrix. Western blot analysis of pooled column fractions identified these proteins as tubulin and src, respectively. Using purified preparations of src and tubulin, PKC-zeta was shown to interact with both proteins using blot overlay. To demonstrate a functional interaction in vivo, PC12 cells expressing a temperature-sensitive v-src were shifted to the permissive temperature (37 degrees C), followed by immunoprecipitation. At the permissive temperature where src was active, PKC-zeta was tyrosine phosphorylated and coassociated with src in vivo; by comparison, at the nonpermissive temperature (40 degrees C) PKC-zeta was not tyrosine phosphorylated. Taken together, these findings support a novel role for the interaction of src and atypical PKC in vivo, which is dependent upon the activity of src and the tyrosine phosphorylation state of PKC-zeta. PMID- 10527888 TI - Novel polymorphism of the 5-lipoxygenase activating protein (FLAP) promoter gene associated with asthma. AB - The human 5-lipoxygenase activating protein (FLAP) gene is one of the key genes involved in the production of the cysteinyl-leukotrienes. We studied novel polymorphism of the FLAP promoter gene and attempted to clarify the relationship between this polymorphism and asthma. We sequenced the FLAP promoter region, containing the -170 to +46-bp sequence from the translational start codon, and found two homozygotes of novel alleles in the polyadenyl region which showed 21 A repeats and 18 A repeats, respectively. The frequency of the 21 A repeats was 52/71 (73.2%) in asthmatics and 39/71 (54.9%) in control subjects. The difference between these frequencies was statistically significant (P = 0.035). This is the first report of FLAP promoter gene polymorphism associated with asthma. Our data suggest that FLAP promoter gene polymorphism might play a crucial role in the pathogenesis of asthma. PMID- 10527889 TI - A crucial role of caspase 3 and caspase 8 in paclitaxel-induced apoptosis. AB - The anticancer drug paclitaxel is well known as an inhibitor of microtubule depolymerization, resulting in mitosis arrest. We investigated the mechanism underlying antitumor effects of paclitaxel on the lung adenocarcinoma cell line LC-2-AD. Less than 10 microg/ml paclitaxel induced mitosis arrest upon LC-2-AD, followed by apoptosis, but more than 30 microg/ml paclitaxel induced apoptosis without mitosis arrest. LC-2-AD with less than 1 microg/ml paclitaxel showed a loss of mitochondrial transmembrane potential (deltapsim), which correlated with antitumor effects. However, LC-2-AD with more than 10 microg/ml paclitaxel showed slight changes in the loss of deltapsim in spite of its ability to induce apoptosis significantly. The cleavage of caspase 3, caspase 8, and DFF45/ICAD was also observed in paclitaxel-induced apoptosis, and the inhibitor of caspase 3 and caspase 8 inhibited both antitumor effects and apoptosis induced by paclitaxel. These results suggest that activation of caspase 3 and caspase 8 plays a crucial role in paclitaxel-induced apoptosis under any concentrations of paclitaxel. PMID- 10527890 TI - Direct visualization of gram-negative bacterial lipopolysaccharide self-assembly. AB - Bacterial lipopolysaccharides (LPS) are outer cell wall components of gram negative bacteria that may cause septic shock in mammals. The exact morphology of LPS when interacting with macromolecular complexes of the septic shock pathway in blood is still uncertain. Here, the geometry and morphology of hydrated bacterial LPS, dispersed in solution, at and below its the critical aggregate concentration, were directly examined by tapping mode atomic force microscopy (TMAFM) and dynamic light scattering. High-resolution phase-shift TMAFM images of hydrated LPS of Salmonella minnesota Re595, Salmonella typhimurium, and Escherichia coli 0111:B4 adsorbed on mica surfaces unveiled nanosized lipidic particles with a species-specific organization. The complex hydrodynamic geometry exhibited by LPS in dilute suspensions may have consequences for the interpretation of LPS biological activity in the host immune response. PMID- 10527891 TI - The human thrombospondin 3 gene: analysis of transcription initiation and an alternatively spliced transcript. AB - Thrombospondin 3 (TSP3) is a member of a family of modular, extracellular proteins that have been implicated in a diverse number of important biological processes. To contribute to an understanding of the precise roles of human TSP3, aspects of TSP3 gene transcription have been investigated. The TSP3 gene (THBS3) shares a promoter/intergenic region of 1.4 kb with the divergently transcribed metaxin gene, and the existence of TSP3 transcription initiation sites in the TSP3/metaxin intergenic region was investigated by a PCR procedure. Transcripts were detected which initiate in the intergenic region, up to several hundred bases upstream from the major transcription start site. An alternatively spliced transcript of TSP3 was also detected by the PCR procedure. This includes a new exon, exon A', which replaces exon A. Exon A' is located in the TSP3/metaxin intergenic region, 1 kb 5' of exon A. In addition, transcripts of metaxin were found with extended 5' ends; these overlap the 5' end of the TSP3 alternative transcript. The complexities of TSP3 transcription initiation revealed by this study could contribute to the tissue-specific expression and diverse functions of TSP3. PMID- 10527892 TI - Natriuretic peptides and cGMP modulate K+, Na+, and H+ fluxes in Zea mays roots. AB - Recent evidence suggests that in plants, as in vertebrates, natriuretic peptides (NPs) regulate homeostasis. In this study noninvasive ion-selective vibrating microelectrodes were used to measure net fluxes of K+, Na+, and H+ in Zea mays root conductive tissue. Immunoreactant plant natriuretic peptides (irPNP) cause immediate net H+ influx and delayed net K+ and Na+ uptake. Delayed net K+ influx was also observed in response to 8-Br-cGMP, however, not accompanied by significant changes in net H+ fluxes. Furthermore, 8-Br-cGMP does not stimulate the plasma membrane H+-ATPase implying that cGMP directly affects cation channels. The data are consistent with NP and cGMP-dependent stimulation of nonselective cation channels with P(K) > P(Na) and point to a complex role for NPs in plant homeostasis. PMID- 10527893 TI - Interactions between Candida albicans and the human extracellular matrix component tenascin-C. AB - Tenascins are large multimeric proteins that contain repeated structural motifs that include epidermal growth factor (EGF)-like repeats, fibronectin type III repeats and a globular fibrinogen-like domain, and are involved in tissue and organ morphogenesis, as well as in adhesion and migration of cells. C. albicans germ-tubes, but not blastospores, were able to bind to soluble human tenascin-C as revealed by an indirect immunofluorescence assay. However, materials present in cell wall extracts from both morphologies attached to tenascin-C immobilized in wells of a microtiter plate. The binding specificity was demonstrated by the inhibitory effect of antibodies against C. albicans cell wall components and an anti-tenascin antibody, but not anti-laminin antibody. Fibronectin, but not fibrinogen, inhibited binding, thus indicating a role of the fibronectin type III repeats in the interaction between the fungus and tenascin-C. Binding of C. albicans cell wall materials to tenascin was RGD- and divalent cation independent. PMID- 10527894 TI - Signal transduction in human macrophages by gp83 ligand of Trypanosoma cruzi: trypomastigote gp83 ligand up-regulates trypanosome entry through protein kinase C activation. AB - We found that Trypanosoma cruzi trypomastigote cloned surface ligand (gp83 trans sialidase) signals macrophages to up-regulate parasite entry by activating protein kinase C (PKC). Incubation of r-gp83 ligand with macrophages activates PKC and this activation is abolished when r-gp83 is depleted by immunoprecipitation with anti-r-gp83 antibodies, which recognize the secreted gp83 of trypomastigotes by immunoblotting. This activation is seen as early as 15 min with maximal activity at 60 min and correlates with the concentration of macrophage cell cytosol. Bisindolylmaleimide I, a PKC inhibitor, abolished the activation of PKC induced by r-gp83 ligand. Incubation of macrophages with r-gp83 ligand significantly enhanced the number of trypanosomes per cell. Bisindolylmaleimide I also inhibited the enhancement of trypomastigote uptake by macrophages induced by the r-ligand. These results demonstrate that T. cruzi uses a novel mechanism to signal cells in the process of trypanosome entry, via a secreted trypanosome ligand which signals macrophages through activation of PKC. PMID- 10527895 TI - Disulfide-cross-linked tau and MAP2 homodimers readily promote microtubule assembly. AB - The neuronal proteins Tau and MAP2 use homologous C-terminal MT-binding regions (MTBRs) to interact with microtubules, F-actin, and intermediate filaments. Although Tau-MTBR is the principal component of pronase-treated Alzheimer paired helical filaments, both Tau and MAP2 form filaments in vitro from disulfide linked homodimers. That the critical thiol lies within a domain needed for MT binding raised the question: Does disulfide formation block Tau-Tau or MAP2-MAP2 dimer binding to microtubules, thereby acting to divert dimers toward filament formation? We now report that cross-linked Tau and MAP2 homodimers readily promote tubulin polymerization and that monomer and dimer affinity for MTs is surprisingly similar. Therefore, disulfide cross-bridging into homodimers is unlikely to be a drive force for filament formation in Alzheimer's disease. PMID- 10527896 TI - Metal cluster labeling. PMID- 10527897 TI - Synthesis of undecagold labeling compounds and their applications in electron microscopic analysis of multiprotein complexes. PMID- 10527898 TI - Undecagold cluster labeling of proteins at reactive cysteine residues. AB - Undecagold cluster labeling of reactive cysteine residues in numerous proteins has allowed the labeled sites to be identified by electron microscopy, providing high-resolution information on the location and orientation of subunits in oligomeric enzymes, virus capsids, crystalline sheets of membrane proteins, and muscle thin filaments. The range of applications of undecagold cluster labeling has been greatly extended by the availability of site-directed mutagenesis to introduce cysteine residues at sites of interest. In this paper I discuss factors that can influence the extent and specificity of labeling, methods for biochemical analysis of undecagold-labeled proteins, and the effects of undecagold cluster labeling on biological activity. PMID- 10527899 TI - Review: chemical aspects of the use of gold clusters in structural biology. PMID- 10527900 TI - Gold-tagged RNA-A probe for macromolecular assemblies. AB - Ribonucleic acids (RNAs) play a key role in many fundamental life processes. These polymers are often found complexed with proteins in extremely large particles whose molecular mass may reach several millions of daltons (e.g., ribosomes, spliceosomes, and viruses). Structural studies of such RNA-protein complexes should help elucidate their mode of action. For the structural analyses of many macromolecular assemblies, electron microscopy (EM) has served an instrumental role. However, localization by EM of RNA within biological complexes is not yet a straightforward undertaking. Here we describe a methodology for the covalent tagging of RNA molecules with gold clusters, thereby enabling their direct visualization by microscopical methods. Our strategy involves transcription in vitro of RNAs that carry free thiol groups, using ribonucleoside triphosphate analogs containing a substituent with a terminal thiol group on their heterocyclic ring. This synthesis is followed by coupling of gold clusters to the thiolated transcript through a maleimido group. Visualization of such gold tagged RNAs by transmission electron microscopy showed spots of gold clusters, with a diameter of 1-2 nm, arranged at nearly regular distances on an imaginary curve that presumably corresponds to the RNA chain. This assignment was corroborated by atomic force microscopy that exhibited images of RNA chains in which knob-like structures, whose height corresponds to the diameter of the gold clusters, were clearly seen. This study demonstrates the potential use of nucleic acids that are covalently labeled with gold clusters for the structural characterization of protein-RNA complexes. PMID- 10527901 TI - Gold-ATP. AB - The synthesis, purification, and chemical analysis of two covalent conjugates between ATP and undecagold are described, one in which gold is attached to the ribose moiety of ATP and the other in which it is attached to the N-6 position of the adenine base. The former probe was then used to bind to two ATP binding proteins, the helicase DnaB and the chaperone DnaK. After purification from unbound gold by column chromatography, binding was measured by UV-Vis spectroscopy, then the protein and gold were visualized by scanning transmission electron microscopy. Binding was observed with the conjugates, and virtually no binding occurred in the control of undecagold without the ATP attached. This new probe may be useful for studying nucleotide binding sites on proteins or for labeling nucleic acids or oligonucleotides directly. PMID- 10527902 TI - Recollections of the electron crystallographic heavy atom derivative search of purple membrane: the quest for EM structure determination. AB - The use of multiple isomorphous replacement in protein electron crystallography for phase determination has been systematically studied only for purple membrane, even though the use of heavy atoms or heavy atom clusters has been used on many occasions in electron microscopy for locating domains or subunits in protein assemblies. The background behind the structure determination of bacteriorhodopsin, the protein component of purple membranes, is summarized and an evaluation of the strengths and weaknesses of using isomorphous replacement in electron crystallography is discussed. PMID- 10527903 TI - Metal compounds as tools for the construction and the interpretation of medium resolution maps of ribosomal particles. AB - Procedures were developed exploiting organometallic clusters and coordination compounds in combination with heavy metal salts for derivatization of ribosomal crystals. These enabled the construction of multiple isomorphous replacement (MIR) and multiple isomorphous replacement combined with anomalous scattering medium-resolution electron density maps for the ribosomal particles that yield the crystals diffracting to the highest resolution, 3 A, of the large subunit from Haloarcula marismortui and the small subunit from Thermus thermophilus. The first steps in the interpretation of the 7. 3-A MIR map of the small subunit were made with the aid of a tetrairidium cluster that was covalently attached to exposed sulfhydryls on the particle's surface prior to crystallization. The positions of these sulfhydryls were localized in difference Fourier maps that were constructed with the MIR phases. PMID- 10527904 TI - Metallosomes. AB - Structures and ordered arrays containing organometallic particles have potential application in nanofabrication, smaller computer components, optical devices, sensors, and membrane probes and as detection agents. Here, we describe construction of gold clusters covalently attached to lipids and their use in forming typical lipid structures: micelles, liposomes ("metallosomes"), and sheets on an air-water interface. Two sizes of gold clusters were used, undecagold, with an 11-gold atom core 0.8 nm in diameter, and the larger Nanogold, with a 1.4-nm gold core. The morphology of the structures formed was determined by electron microscopy at a resolution at which single gold-lipid molecules were visualized. Further modification by additional catalytic metal deposition enhanced detectability. The approach is flexible and permits a wide variety of metal particle structures to be created using known lipid structures as templates. Additionally, these gold-lipids may serve as useful membrane labels. PMID- 10527905 TI - Visualizing "greengold" clusters in the STEM. AB - "Greengold" is a large metallic cluster thought to contain 75 gold atoms in a compact 1.4-nm-diameter core surrounded by an organic shell. Scanning transmission electron microscope imaging shows uniform mass and size distributions with an apparent mass of 24 kDa, unaffected by radiation damage. The signal-to-noise ratio is adequate for visualization at low dose and in the presence of a relatively thick biological matrix. Under some conditions these clusters have a slight tendency to form linear chains and 2-D hexagonal arrays with a spacing of 2.6 nm. The parameters presented permit estimation of the feasibility of proposed labeling experiments. PMID- 10527906 TI - Tetrairidium, a four-atom cluster, is readily visible as a density label in three dimensional cryo-EM maps of proteins at 10-25 A resolution. AB - Heavy metal clusters derivatized to bind to designated chemical groups on proteins have great potential as density labels for cryo-electron microscopy. Smaller clusters offer higher resolution and penetrate more easily into sterically restricted sites, but are more difficult to detect. In this context, we have explored the potential of tetrairidium (Ir(4)) as a density label by attaching it via maleimide linkage to the C-terminus of the hepatitis B virus (HBV) capsid protein. Although the clusters are not visible in unprocessed cryo electron micrographs, they are distinctly visible in three-dimensional density maps calculated from them, even at only partial occupancy. The Ir(4) label was clearly visualized in our maps at 11-14 A resolution of both size variants of the HBV capsid, thus confirming our previous localization of this site with undecagold (Zlotnick, A., Cheng, N., Stahl, S. J., Conway, J. F., Steven, A. C., and Wingfield, P. T., Proc. Natl. Acad. Sci. USA 94, 9556-9561, 1997). Ir(4) penetrated to the interior of intact capsids to label this site on their inner surface, unlike undecagold for which labelling was achieved only with dissociated dimers that were then reassembled into capsids. The Ir(4) cluster remained visible as the resolution of the maps was lowered progressively to approximately 25 A. PMID- 10527908 TI - Ni-NTA-gold clusters target His-tagged proteins. AB - Addition of six histidines to recombinant proteins has proved useful in their purification by nickel-affinity columns. This technology was adapted by synthesizing the chelator for nickel (nitrilotriacetic acid, NTA) onto the surface of gold clusters. These Ni-NTA-gold clusters were shown to specifically target the 6His region of tagged proteins. Results were verified by column chromatography, dot and overlay blots, UV-Vis spectroscopy, and scanning transmission electron microscopy. A 6His-tagged adenovirus "knob" protein was also shown to maintain receptor binding activity after gold labeling. Two types of gold clusters were used: 1.4-nm Nanogold and a new 1.8-nm "PeptideGold" coated with an NTA-dipeptide-thiol. These novel labels should be useful in site-specific high-resolution EM labeling, as well as in metallographic development, detection in the light microscope, or direct visualization. PMID- 10527907 TI - Giant platinum clusters: 2-nm covalent metal cluster labels. AB - A new class of covalently linkable platinum cluster reagents with core diameters close to 2 nm has been prepared. The new label offers the performance advantages and versatility of the 1.4-nm Nanogold in a larger label which is more clearly visualized against electron-dense regions of a specimen. Large platinum clusters were prepared by the reduction of platinum(II) acetate in the presence of 1,10 phenanthroline ligands which had been synthetically modified to include solubilizing and reactive cross-linkable functional groups. These were then conjugated site-specifically to antibody IgG molecules and to Fab' fragments and visualized by scanning transmission electron microscopy. The resulting conjugates may be autometallographically enhanced and show sensitivity similar to that of Nanogold conjugates in immunoblotting experiments. In preliminary experiments, they have also exhibited labeling of tissue antigens and penetrate to access nuclear targets. PMID- 10527909 TI - Cell-type-specific activation of p38 protein kinase cascades by the novel tumor promoter palytoxin. AB - Palytoxin is a potent non-12-O-tetradecanoylphorbol-13-acetate (TPA)-type skin tumor promoter. We used COS7 and HeLa cells to investigate the protein kinase cascades by which palytoxin activates the mitogen-activated protein kinase (MAPK) p38. Three p38 kinases have been identified: stress-activated protein kinase/extracellular signal-regulated kinase kinase-1 (SEK1), MAPK kinase 3 (MKK3), and MKK6. SEK1 phosphorylates and activates both p38 and c-Jun NH(2) terminal kinase (JNK), whereas MKK3 and MKK6 selectively phosphorylate and activate p38. Although transiently overexpressed SEK1 activates p38 in cells, the importance of endogenous SEK1 for the activation of p38 by specific types of stimuli is unclear because some agents, such as sorbitol, can activate p38 in cells derived from SEK1 knockout mice. Because we previously showed that palytoxin activates JNK through an SEK1-dependent pathway, we investigated whether SEK1 also mediates the activation of p38 by palytoxin. The results presented here demonstrate that endogenous SEK1 does play an important role in the activation of p38 by palytoxin in specific cell types. In COS7 cells, palytoxin stimulated the phosphorylation of SEK1 and MKK6, and expression of dominant negative mutants of either SEK1 or MKK6 inhibited palytoxin-stimulated p38 activation. In HeLa cells, palytoxin stimulated the phosphorylation of MKK3 in addition to SEK1 and MKK6. In contrast to COS7 cells, in HeLa cells expression of a dominant negative mutant of SEK1 did not inhibit palytoxin-stimulated activation of p38, although expression of dominant negative mutants of either MKK3 or MKK6 did inhibit palytoxin-stimulated p38 activation in this cell type. These studies indicate that the importance of SEK1 in the activation of p38 by palytoxin depends on the ability of palytoxin to activate MKK3 and MKK6. PMID- 10527910 TI - Metabolism of chloroform by cytochrome P450 2E1 is required for induction of toxicity in the liver, kidney, and nose of male mice. AB - Chloroform is a nongenotoxic-cytotoxic liver and kidney carcinogen and nasal toxicant in some strains and sexes of rodents. Substantial evidence indicates that tumor induction is secondary to events associated with cytolethality and regenerative cell proliferation. Therefore, pathways leading to toxicity, such as metabolic activation, become critical information in mechanism-based risk assessments. The purpose of this study was to determine the degree to which chloroform-induced cytotoxicity is dependent on the cytochromes P450 in general and P450 2E1 in particular. Male B6C3F(1), Sv/129 wild-type (Cyp2e1+/+), and Sv/129 CYP2E1 knockout (Cyp2e1-/- or Cyp2e1-null) mice were exposed 6 h/day for 4 consecutive days to 90 ppm chloroform by inhalation. Parallel control and treated groups, excluding Cyp2e1-null mice, also received an i.p. injection (150 mg/kg) of the irreversible cytochrome P450 inhibitor 1-aminobenzotriazole (ABT) twice on the day before exposures began and 1 h before every exposure. Cells in S-phase were labeled by infusion of BrdU via an implanted osmotic pump for 3.5 days prior to necropsy, and the labeling index was quantified immunohistochemically. B6C3F(1) and Sv/129 wild-type mice exposed to chloroform alone had extensive hepatic and renal necrosis with significant regenerative cell proliferation. These animals had minimal toxicity in the nasal turbinates with focal periosteal cell proliferation. Administration of ABT completely protected against the hepatic, renal, and nasal toxic effects of chloroform. Induced pathological changes and regenerative cell proliferation were absent in these target sites in Cyp2e1-/- mice exposed to 90 ppm chloroform. These findings indicate that metabolism is obligatory for the development of chloroform-induced hepatic, renal, and nasal toxicity and that cytochrome P450 2E1 appears to be the only enzyme responsible for this cytotoxic-related metabolic conversion under these exposure conditions. PMID- 10527911 TI - Role of Ca(2+) in the regulation of nickel-inducible Cap43 gene expression. AB - We have recently cloned a gene, Cap43, that was significantly induced by exposure to nontoxic levels of both water-soluble and -insoluble Ni(2+) compounds. In this paper, we utilized the expression levels of this gene as a tool to identify second messengers involved in nickel-inducible transcription. We report here that the Ca(2+) ionophore A23187 substantially stimulated Cap43 gene expression. In addition, we found that BAPTA-AM, a specific chelator of free intracellular Ca(2+), consistently attenuated the induction of Cap43, indicating that elevation of intracellular Ca(2+) was essential for this response. TPEN, a chelator of heavy metals, such as Ni(2+) with a very low affinity for Ca(2+), did not attenuate Cap43 induced by Ni or calcium ionophore, suggesting that elevations of Ca(2+) but probably not elevations of other metal ions were involved in the induction of Cap43. A direct measurement of Ca(2+) levels using the fluorescent probe Fluo-3 AM showed elevations of free intracellular Ca(2+) in Ni-treated cells. A strong induction of Cap43 by okadaic acid suggested the involvement of a serine/threonine phosphorylation in a signaling pathway that was presumably activated by Ni and that led to enhanced Cap43 gene expression. However, calcium dependent protein kinase(s) involved in the nickel-activated signaling pathway remains to be identified. PMID- 10527912 TI - Spontaneous and thiazolidinedione-induced B6C3F1 mouse hemangiosarcomas exhibit low ras oncogene mutation frequencies. AB - Hemangiosarcomasare uncommon malignant endothelial cell tumors in humans and experimental animal species. The mechanisms giving rise to these tumors are poorly understood even though the histotypes are comparable between humans and rodents. Activating mutations in cellular ras protooncogenes have been detected in sporadic and chemically induced human and rodent hemangiosarcomas. Ras activation significantly modulates tumor angiogenesis, suggesting that mutations in ras genes might be causally related to vascular tumorigenesis. To more clearly define the role of ras in experimental vascular tumorigenesis, mutations in the Ki- and Ha-ras genes were characterized in 63 hemangiosarcomas that arose unexpectedly in control and treated B6C3F1 mice during a two-year carcinogenicity study of the thiazolidinedione troglitazone. DNA was extracted from paraffin sections of mouse hemangiosarcomas, control liver, or positive control hepatocellular carcinomas with defined mutations in the Ki- or Ha-ras genes. Exons 1 and 2 of the Ki- and Ha-ras genes were independently amplified using primer extension preamplification/locus-specific heminested PCR, and PCR amplicons were directly sequenced to identify mutations in codons 12, 13, or 61. Activating mutations were detected in 3 of 63 hemangiosarcomas: a single G-->A transition in the second position of Ki-ras codon 13 in a tumor from a treated animal and two G-->T transversions in the second position of Ha-ras codon 13, one in a single tumor from a control animal and one in a tumor from a treated animal. These mutations are consistent with endogenous mutagenesis arising from oxidative DNA damage. The low frequency of mutation (<5%) indicates that ras mutations did not contribute significantly to hemangiosarcoma development and suggests that mutational ras activation may not be a necessary step in vascular tumorigenesis in mice. PMID- 10527913 TI - Role of Fas-Fas ligand interactions in 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD)-induced immunotoxicity: increased resistance of thymocytes from Fas deficient (lpr) and Fas ligand-defective (gld) mice to TCDD-induced toxicity. AB - 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a highly toxic environmental pollutant well known for its toxicity to the thymus. Recent studies from our laboratory demonstrated that TCDD induces apoptosis in thymocytes. In the current study, we investigated the mechanism of TCDD-induced apoptosis. Administration of a single dose of TCDD at 0.1, 1, 5, and 50 microg/kg body wt intraperitoneally, into C57BL/6 +/+ (wild-type) mice caused a dose-dependent decrease in thymic cellularity. In contrast, a similar treatment with TCDD, in Fas-deficient C57BL/6 lpr/lpr (lpr) or Fas-ligand defective C57BL/6 gld/gld (gld), mice failed to induce thymic atrophy at 0.1-5 microg/kg body wt of TCDD. In lpr and gld mice, significant thymic atrophy was seen only at 50 microg/kg body wt of TCDD. Injection of TCDD caused apoptosis only in wild-type but not in lpr or gld mice. The sera from TCDD-treated wild-type mice exhibited increased levels of soluble Fas ligand, inasmuch as incubation of Fas(+), but not Fas(-) cells with the sera, triggered apoptosis. Also, TCDD-induced apoptosis in thymocytes was inhibited both in vitro and in vivo by caspase inhibitors. TCDD treatment caused significant up-regulation in the expression of FasL but not Fas mRNA in the thymocytes of wild-type mice. Also, such thymocytes exhibited marked alterations in the surface markers, characteristic of cells undergoing apoptosis. In contrast, TCDD treatment caused minimal phenotypic changes in thymocytes from lpr and gld mice. Together, the current study demonstrates that Fas-Fas ligand interactions play an important role in TCDD-mediated induction of apoptosis and immunotoxicity. PMID- 10527914 TI - Distinct immunomodulation by autoimmunogenic xenobiotics in susceptible and resistant mice. AB - HgCl(2) and diphenylhydantoin (DPH) are prototype chemicals associated with diverse (auto)immune effects in genetically susceptible individuals. Both chemicals activate T cells, and the balance of Th1 versus Th2 activation may influence the clinical outcome of exposure. It is unknown which chemically created neoantigens are responsible for Th activation. We therefore investigated the effect of DPH and HgCl(2) on specific responses to TNP-ovalbumin, in mouse strains with varying sensitivity for the adverse effects. HgCl(2) was found to enhance Th2-driven antibody responses in susceptible B10.s, but protective type 1 responses in resistant B10.d2 mice. This was chemical-specific, as DPH enhanced type 2 responses in both strains. DBA/2 mice were relatively unresponsive to HgCl(2), whereas DPH stimulated type 1 responses in these mice. Interestingly, prior exposure to HgCl(2), but not DPH, facilitated IC deposition in B10.s mice only. Thus, we demonstrate that, depending on MHC-II and background genes, HgCl(2) and DPH preferentially adjuvate type 1 or type 2 responses. In case of HgCl(2), the type of response corresponds with susceptibility to antibody mediated autoimmunity induced by this chemical. In addition, we demonstrate that, within one strain, different autoimmunogenic chemicals can enhance distinct responses to the same antigen. PMID- 10527915 TI - Effects of microsomal enzyme inducers on thyroid-follicular cell proliferation, hyperplasia, and hypertrophy. AB - The microsomal enzyme inducer (MEI), phenobarbital (PB), has been proposed to promote thyroid tumors by increasing the biotransformation and elimination of T(4), resulting in an increase in serum thyroid-stimulating hormone (TSH). In turn, TSH stimulates thyroid gland function, growth, and ultimately neoplasia. The dose-dependent effects of MEI on thyroid-follicular cell proliferation, a measure of thyroid gland growth, has not been reported. In the present study, it was hypothesized that MEIs that increase TSH would stimulate thyroid-follicular cell proliferation and the total number of thyroid-follicular cells. Male Sprague Dawley rats were fed either a basal diet or a diet containing PB (at 300, 600, 1200, or 2400 ppm), pregnenolone-16alpha-carbonitrile (PCN) (at 200, 400, 800, or 1600 ppm), 3-methylcholanthrene (3MC) (at 50, 100, 200, or 400 ppm), or Aroclor 1254 (PCB) (at 25, 50, 100, or 200 ppm) for 7 days. PB and PCN increased TSH 65% and 95%, respectively, whereas 3MC and PCB did not appreciably affect TSH. PB and PCN increased thyroid-follicular cell proliferation 625% and 1200%, respectively, whereas 3MC and PCB did not have a consistent or appreciable effect. The total number of thyroid-follicular cells was not significantly increased by MEI treatment. In conclusion, small increases in TSH by PB and PCN produced large increases in thyroid-follicular cell proliferation, which did not result in a comparable increase in the total number of thyroid-follicular cells. Furthermore, MEI that did not increase TSH did not consistently or appreciably increase thyroid-follicular cell proliferation or cell number. PMID- 10527916 TI - Effects of smokeless tobacco and tumor promoters on cell population growth and apoptosis of B lymphocytes infected with epstein-barr virus types 1 and 2. AB - The effects of smokeless tobacco purified products 4-(N-methyl-N-nitrosamine)-1-3 pyridinyl)-1-butanone (NNK) and N-nitrosonornicotine (NNN), smokeless tobacco extracts (dry snuff, moist snuff, and loose leaf), and the tumor promoters 12-O tetradecanoyl phorbol-13-acetate (TPA) and n-butyrate on cell population growth, cell death, and apoptosis were studied in B lymphocyte cell lines harboring Epstein-Barr virus (EBV) type 1 (Raji and X50-7) or type 2 (HR-1K and AG876) and in an EBV-uninfected control lymphocyte cell line (Ramos). Spontaneous apoptosis was present in all EBV-infected cell lines, but at varying levels. Spontaneous and induced apoptosis were generally greater by Student-Newman-Keuls tests in cells harboring EBV type 2 compared to EBV type 1. The greatest effects on cell population growth, cell death, and apoptosis on cells harboring lytic EBV type 1 (X50-7) was with each of the three smokeless tobacco extracts. The greatest effects on cells harboring EBV type 2 was with TPA and n-butyrate. There were no effects of smokeless tobacco extracts on the Raji cell line that harbors EBV type 1 incapable of lytic replication. Smokeless tobacco purified products, NNN and NNK, had no discernible effects. At the concentrations used in these experiments, there appears to be an EBV type-specific response to chemical induction, with greater susceptibility of lytic EBV type 1 to smokeless tobacco extracts and lytic EBV type 2 to TPA and n-butyrate. This EBV type-specific susceptibility to the effects of smokeless tobacco extracts is another phenotypic difference between EBV types. The use of smokeless tobacco products may affect B lymphocytes infected with replication-capable EBV in the oropharynx. The absence of significant effects with NNK and NNN suggests that these properties reside with other compounds present in tobacco extracts. PMID- 10527917 TI - Tissue distribution of cadmium in rats given minimum amounts of cadmium-polluted rice or cadmium chloride for 8 months. AB - To investigate the relationship between cadmium (Cd) toxicity, intestinal absorption, and its distribution to various tissues in rats treated orally with minimum amounts of Cd, 14 female rats per dose group per time point were given diets consisting of 28% purified diet and 72% ordinary rice containing Cd polluted rice (0. 02, 0.04, 0.12, or 1.01 ppm of Cd) or CdCl(2) (5.08, 19.8, or 40.0 ppm of Cd) for up to 8 months. At 1, 4, and 8 months after the commencement of Cd treatment, seven rats per group were euthanized for pathological examinations to determine the Cd concentrations in the liver and kidneys and metallothionein (MT) in the liver, kidneys, intestinal mucosa, serum, and urine. One week before each period of 1, 4, and 8 months, the remaining seven rats in each group were administered a single dosage of (109)Cd, a tracer, to match the amounts of the designated Cd doses (about 1.2 to 2400 microg/kg body wt). They were euthanized 5 days later to determine the distribution of Cd to various tissues. No Cd-related toxic changes were observed. The concentrations of Cd in the liver and kidneys at any time point and MT in the liver, kidney, serum, and urine at 4 and 8 months increased dose-dependently, whereas MT in the intestinal mucosa did not alter markedly at any time point. The distribution rates of Cd to the liver increased dose-dependently (40% at lower doses to 60% at higher doses), whereas those to the kidney decreased dose-dependently (20% at lower doses to 10% at higher doses). The Cd retention rates 5 days after (109)Cd administration (amounts of Cd in various tissues/amounts of Cd administered) ranged from 0.2 to 1. 0% at any time point. These results suggest that the distribution of Cd to the liver and kidneys after the oral administration vary depending on the dosage levels of Cd. The difference of the distribution pattern of Cd to the liver and kidney is probably due to the difference in the form of the absorbed Cd, i.e., free ion or Cd-MT complex, although not closely related to the MT in the intestinal mucosa. PMID- 10527918 TI - Decrease in protein kinase and phosphatase activities in the liver nuclei of rats exposed to carbon tetrachloride. AB - The alteration in protein kinase and phosphatase activities in the liver nuclei of rats administered carbon tetrachloride (CCl(4)) was investigated. Rats received a single oral administration of CCl(4) (1 ml/100 g body wt of 5, 10, and 25% CCl(4) in corn oil), and 5, 24, and 48 h later they were euthanized by bleeding. The administration of CCl(4) (10 and 25%) caused a significant decrease in protein kinase activity in the liver nuclei. The enzyme activity in the liver nuclei from normal and CCl(4)-administered rats was significantly increased by the addition of Ca(2+) (0.5 mM) and calmodulin (10 microg/ml) in the reaction mixture, suggesting that Ca(2+)/calmodulin-dependent protein kinase activation is not suppressed by CCl(4) treatment. Liver nuclear phosphatase activity toward phosphotyrosine, but not phosphoserine and phosphothreonine, was markedly decreased by CCl(4) (5, 10, and 25%) administration. This decrease was seen 5 h after CCl(4) administration. The presence of vanadate (10(-4) M) in the reaction mixture caused a significant decrease in phosphotyrosine phosphatase activity in the liver nuclei from normal and CCl(4)-administered rats, whereas the enzyme activity was not decreased by okadaic acid (10(-5) M) or sodium fluoride (10(-3) M). The effect of anti-regucalcin antibody (100 ng/ml) in increasing phosphotyrosine phosphatase activity was seen in the liver nuclei of CCl(4) administered rats, suggesting that regucalcin-sensitive phosphatase activity is decreased by CCl(4) administration. The present study demonstrates that CCl(4) administration induces a decrease in protein kinase and tyrosine phosphatase activities, which are involved in signaling factors in the liver nuclei of rats. PMID- 10527920 TI - Parade of the Little Millions. PMID- 10527919 TI - Vicinal-thiol-containing molecules enhance but mono-thiol-containing molecules reduce nickel-induced DNA strand breaks. AB - Several thiol-containing molecules (TCM) are currently used as antidotes for nickel, and vicinal TCM seem to be more effective in mobilizing tissue nickel than are mono TCM. Using single cell alkaline electrophoresis, we have shown that the vicinal TCM, meso-2, 3-dimercaptosuccinic acid (DMSA), 2,3-dimercaptopropane 1-sulfonate, and 2,3-dimercaptopropanol markedly enhanced, whereas the mono TCM, D-penicillamide, glutathione, beta-mercaptoethanol, and diethyl dithiocarbomate, reduced nickel chloride (Ni)-induced DNA breaks in a human leukemia cell line, NB4 cells. Ni or TCM alone did not induce plasmid DNA breaks in test tubes and neither did Ni plus mono TCM; however, Ni plus vicinal TCM did. Vicinal TCM did, but mono TCM did not generate H(2)O(2) in solution. H(2)O(2) alone did not, but H(2)O(2) plus Ni induced plasmid DNA breaks. Although Ni plus glutathione did not break DNA, Ni plus glutathione plus H(2)O(2) did. The Ni-DMSA-induced DNA breaks in NB4 cells, as well as in plasmids, were completely prevented by d-mannitol or partially prevented by several antioxidants. Therefore, the DNA breaks induced by Ni plus vicinal TCM seem to be due to the complex of Ni with TCM in concert with the H(2)O(2) produced by the vicinal TCM. The results that DMSA at a concentration as low as 5 microM enhanced the Ni-induced DNA breaks suggest a further evaluation of the TCM as nickel chelators is needed. PMID- 10527921 TI - The Diversity of Eukaryotes. AB - The discipline of evolutionary protistology has emerged in the past 30 yr. There is as yet no agreed view of how protists are interrelated or how they should be classified. The foundations of a stable taxonomic superstructure for the protists and other eukaryotes lie in cataloging the diversity of the major monophyletic lineages of these organisms. The use of common patterns of cell organization (ultrastructural identity) seems to provide us with the most robust hypotheses of such lineages. These lineages are placed in 71 groups without identifiable sister taxa. These groups are here referred to as "major building blocks." For the first time, the compositions, ultrastructural identities, synapomorphies (where available), and subgroups of the major building blocks are summarized. More than 200 further lineages without clear identities are listed. This catalog includes all known major elements of the comprehensive evolutionary tree of protists and eukaryotes. Different approaches among protistologists to issues of nomenclature, ranking, and definitions of these groups are discussed, with particular reference to two groups-the stramenopiles and the Archezoa. The concept of "extended in group" is introduced to refer to in-groups and the most proximate sister group and to assist in identifying the hierarchical location of taxa. PMID- 10527922 TI - Ultrastructure as a Control for Protistan Molecular Phylogeny. AB - A variety of molecular sequences and treeing methods have been used in attempts to unravel early protistan evolution and the origins of "higher" eukaryotic taxa. How does one know which approach is closest to the real phylogenetic tree? Obviously it is the robustness of its resulting trees, the coherence with other data sets, both structural and molecular, that is the test. Simply put: it should make biological sense. It seems evident, comparing morphology, especially ultrastructure, with ribosomal DNA trees, that the major lineages have now been confirmed. In particular, the remarkably conservative mitochondrial crista type in protists is coherent with mitochondrial DNA sequences. Several amitochondrial groups, presumed to be primitive on the basis of SSU ribosomal DNA, show alarming positional volatility when other genes are used. In addition, the presence of mitochondrial genes in the nucleus of several amitochondrial flagellates raises doubts about them being primordially amitochondrial. Consequently, the root of the eukaryote tree is still in question. A disturbing question arises: can loss of features in parasitism mimic primitiveness not only in a morphological but also in a molecular way, evolving more rapidly and creating long branches that methodologically place them basal in the trees? Conflicting molecular phylogenies cannot be resolved by molecular data alone. Morpholological, especially ultrastructural, data are an essential component of phylogenetic reconstruction. PMID- 10527923 TI - The Tangled Web: Gene Genealogies and the Origin of Eukaryotes. AB - Accessing data from the genomes of organisms (individual genes) and analyzing these data using sophisticated alignment and phylogenetic methods led to the expectation that we would be able to paint a clear picture of the evolution of eukaryotes. Previous analyses based on morphology and ultrastructure failed to pinpoint both the sister taxon to eukaryotes and the branching order of eukaryotic lineages. However, the expectation that molecular data would provide resolution has not been met since a growing number of gene genealogies present conflicting hypotheses for the origin and diversification of eukaryotes. Instead of reconstructing a simple bifurcating tree of life, these gene genealogies have generated a complex picture of eukaryotic genomes whereby ancient lateral transfers (of individual genes or perhaps even entire genomes) has tangled the evolutionary history of eukaryotes. Resolution of these conflicting genealogies comes in recognizing that eukaryotes are chimeric, containing genetic information from multiple ancestral lineages. PMID- 10527924 TI - Reconstructing Early Events in Eukaryotic Evolution. AB - Resolving the order of events that occurred during the transition from prokaryotic to eukaryotic cells remains one of the greatest problems in cell evolution. One view, the Archezoa hypothesis, proposes that the endosymbiotic origin of mitochondria occurred relatively late in eukaryotic evolution and that several mitochondrion-lacking protist groups diverged before the establishment of the organelle. Phylogenies based on small subunit ribosomal RNA and several protein-coding genes supported this proposal, placing amitochondriate protists such as diplomonads, parabasalids, and Microsporidia as the earliest diverging eukaryotic lineages. However, trees of other molecules, such as tubulins, heat shock protein 70, TATA box-binding protein, and the largest subunit of RNA polymerase II, indicate that Microsporidia are not deeply branching eukaryotes but instead are close relatives of the Fungi. Furthermore, recent discoveries of mitochondrion-derived genes in the nuclear genomes of entamoebae, Microsporidia, parabasalids, and diplomonads suggest that these organisms likely descend from mitochondrion-bearing ancestors. Although several protist lineages formally remain as candidates for Archezoa, most evidence suggests that the mitochondrial endosymbiosis took place prior to the divergence of all extant eukaryotes. In addition, discoveries of proteobacterial-like nuclear genes coding for cytoplasmic proteins indicate that the mitochondrial symbiont may have contributed more to the eukaryotic lineage than previously thought. As genome sequence data from parabasalids and diplomonads accumulate, it is becoming clear that the last common ancestor of these protist taxa and other extant eukaryotic groups already possessed many of the complex features found in most eukaryotes but lacking in prokaryotes. However, our confidence in the deeply branching position of diplomonads and parabasalids among eukaryotes is weakened by conflicting phylogenies and potential sources of artifact. Our current picture of early eukaryotic evolution is in a state of flux. PMID- 10527925 TI - Tracing the Thread of Plastid Diversity through the Tapestry of Life. AB - Plastids (chloroplasts) are endosymbiotic organelles derived from previously free living cyanobacteria. They are dependent on their host cell to the degree that the majority of the proteins expressed in the plastid are encoded in the nuclear genome of the host cell, and it is this genetic dependency that distinguishes organelles from obligate endosymbionts. Reduction in the size of the plastid genome has occurred via gene loss, substitution of nuclear genes, and gene transfer. The plastids of Chlorophyta and plants, Rhodophyta, and Glaucocystophyta are primary plastids (i.e., derived directly from a cyanobacterium). These three lineages may or may not be descended from a single endosymbiotic event. All other lineages of plastids have acquired their plastids by secondary (or tertiary) endosymbiosis, in which a eukaryote already equipped with plastids is preyed upon by a second eukaryote. Considerable gene transfer has occurred among genomes and, at times, between organisms. The eukaryotic crown group Alveolata has a particularly complex history of plastid acquisition. PMID- 10527926 TI - A Search for the Origins of Animals and Fungi: Comparing and Combining Molecular Data. AB - Green plants, animals, and fungi have long held our interest as complex, largely multicellular eukaryotes of indeterminate origin. Considerable progress has now been made toward understanding the evolutionary relationships among these taxa as well as identifying their closest protistan relatives. An exclusive animal-fungal clade (the Opisthokonta) is now widely accepted based on an insertion in the protein synthesis elongation factor 1alpha (EF-1alpha) and molecular phylogenies of ribosomal RNAs and the conservative proteins actin, alpha-tubulin, beta tubulin, and EF-1alpha. Protein data also suggest that the cellular (dictyostelid) and acellular (myxogastrid) slime molds are a close outgroup to the animal-fungal clade. Subsequent sequencing and phylogenetic analysis of EF 1alpha sequences very strongly support a monophyletic slime mold clade (the Mycetozoa or Eumycetozoa), which also includes the lesser-known protostelid slime molds. Monophyly of the opisthokont and mycetozoan clades, exclusive of green plants, is suggested by individual analyses of EF-1alpha and actin and given strong support by concatenated protein data. Neither the monophyly of the slime molds nor their close relationship to animals and fungi are consistently supported by ribosomal RNA data. Thus, it appears unlikely that any single molecule will accurately reconstruct all higher-order taxonomy. PMID- 10527927 TI - The malonyl-CoA-long-chain acyl-CoA axis in the maintenance of mammalian cell function. AB - Long-chain acyl-CoA esters have potent specific actions (e.g. on gene transcription, membrane trafficking) as well as non-specific ones (e.g. on phospholipid bilayers). They are synthesized on the cytosolic aspects of several intracellular membranes, to give rise to (a) cytosolic pool(s) to which a variety of enzymes and processes have access, including some localized in the nucleus. Their concentration in cells is highly regulated, interconversion with corresponding acylcarnitines being the most important mechanism involved. This reaction is catalysed by cytosol-accessible carnitine long-chain acyl (palmitoyl) transferase activities that are themselves located on multiple membrane systems. Regulation of these activities is through the inhibitory action of malonyl-CoA. Hence the existence of a potent malonyl-CoA-acyl-CoA axis through which many processes involved in the maintenance of mammalian cell function are regulated. The molecular, topographical and physiological interactions that make this possible are described and discussed. PMID- 10527930 TI - Sulphation of lithocholic acid in the colon-carcinoma cell line CaCo-2. AB - High levels of bile acids in the colon may correlate with an increased risk of colon cancer, but the underlying mechanisms are not known. Proteoglycan structures have been shown to change when human colon cells differentiate in vitro. The expression of [(35)S]sulphated molecules was used as a phenotypic marker to study the effects of bile acids on the human-colon-carcinoma cell line CaCo-2. [(35)S]sulphated compounds were isolated from the medium of cell fractions of cells metabolically labelled with [(35)S]sulphate in the absence and presence of cholic acid, deoxycholic acid, chenodeoxycholic acid and lithocholic acid (LA). Labelled molecules were analysed by gel chromatography, HPLC and SDS/PAGE in combination with chemical and enzymic methods. The expression of (35)S-labelled proteoglycans was not affected by any of the bile acids tested. However, the level of sulphated metabolites increased 7-18-fold in different experiments during a 22 h labelling period in the presence of an LA concentration of 10 microg/ml (26.6 nmol/ml) compared with controls. Further analyses showed that this was due, at least in part, to the sulphation of LA itself. This sulphation of LA was a rapid process followed by secretion back to the medium. Brefeldin A did not reduce the sulphation of LA, indicating that this conversion takes place in the cytosol, rather than in the Golgi apparatus of the CaCo-2 cells. LA in colon may be sulphated efficiently by the colonocytes to reduce the toxic effects of this particular bile acid. Sulphation may possibly be an important protective mechanism in the colon. PMID- 10527929 TI - The role of tyrosine-9 and the C-terminal helix in the catalytic mechanism of Alpha-class glutathione S-transferases. AB - Glutathione S-transferases (GSTs) play a key role in the metabolism of drugs and xenobiotics. To investigate the catalytic mechanism, substrate binding and catalysis by the wild-type and two mutants of GST A1-1 have been studied. Substitution of the 'essential' Tyr(9) by phenylalanine leads to a marked decrease in the k(cat) for 1-chloro-2,4-dinitrobenzene (CDNB), but has no affect on k(cat) for ethacrynic acid. Similarly, removal of the C-terminal helix by truncation of the enzyme at residue 209 leads to a decrease in k(cat) for CDNB, but an increase in k(cat) for ethacrynic acid. The binding of a GSH analogue increases the affinity of the wild-type enzyme for CDNB, and increases the rate of the enzyme-catalysed conjugation of this substrate with the small thiols 2 mercaptoethanol and dithiothreitol. This suggests that GSH binding produces a conformational change which is transmitted to the binding site for the hydrophobic substrate, where it alters both the affinity for the substrate and the catalytic-centre activity ('turnover number') for conjugation, perhaps by increasing the proportion of the substrate bound productively. Neither of these two effects of GSH analogues are seen in the C-terminally truncated enzyme, indicating a role for the C-terminal helix in the GSH-induced conformational change. PMID- 10527928 TI - Oligomerization properties of fragile-X mental-retardation protein (FMRP) and the fragile-X-related proteins FXR1P and FXR2P. AB - The absence of fragile-X mental-retardation protein (FMRP) results in fragile-X syndrome. Two other fragile-X-related (FXR) proteins have been described, FXR1P and FXR2P, which are both very similar in amino acid sequence to FMRP. Interaction between the three proteins as well as with themselves has been demonstrated. The FXR proteins are believed to play a role in RNA metabolism. To characterize a possible functional role of the interacting proteins the complex formation of the FXR proteins was studied in mammalian cells. Double immunofluorescence analysis in COS cells over-expressing either FMRP ISO12/FXR1P or FMRP ISO12/FXR2P confirmed heterotypic interactions. However, Western-blotting studies on cellular homogenates containing physiological amounts of the three proteins gave different indications. Gel-filtration experiments under physiological as well as EDTA conditions showed that the FXR proteins were in complexes of >600 kDa, as parts of messenger ribonuclear protein (mRNP) particles associated with polyribosomes. Salt treatment shifted FMRP, FXR1P and FXR2P into distinct intermediate complexes, with molecular masses between 200 and 300 kDa. Immunoprecipitations of FMRP as well as FXR1P from the dissociated complexes revealed that the vast majority of the FXR proteins do not form heteromeric complexes. Further analysis by [(35)S]methionine labelling in vivo followed by immunoprecipitation indicated that no proteins other than the FXR proteins were present in these complexes. These results suggest that the FXR proteins form homo multimers preferentially under physiological conditions in mammalian cells, and might participate in mRNP particles with separate functions. PMID- 10527931 TI - Novel bimodal effects of the G-protein tissue transglutaminase on adrenoreceptor signalling. AB - Tissue transglutaminase (tTG) is a novel G-protein that previous studies showed can couple ligand-bound activated alpha(1B) adrenoreceptors to phospholipase C delta, resulting in phosphoinositide (PI) hydrolysis. In human neuroblastoma SH SY5Y cells we found that although endogenous tTG can facilitate alpha(1B) adrenoreceptor-stimulated PI hydrolysis, its contribution is minor compared with the classical heterotrimeric G-protein G(q/11). Further, we show that the alpha(1B) adrenoreceptor recruits tTG to the membrane and that this recruitment is enhanced by agonist occupancy of the receptor. In addition, the effects of tTG on signalling are bimodal. At low expression levels, tTG enhanced alpha(1B) adrenoreceptor-stimulated PI hydrolysis, whereas at higher expression levels tTG attenuated significantly this response. These findings are the first to demonstrate that a protein can both facilitate and attenuate receptor-stimulated PI hydrolysis. PMID- 10527932 TI - Site-directed mutagenesis establishes cysteine-110 as essential for enzyme activity in human gamma-glutamyl hydrolase. AB - Gamma-glutamyl hydrolase (GH), which hydrolyses the gamma-glutamyl conjugates of folic acid, is a key enzyme in the maintenance of cellular folylpolyglutamate concentrations. The catalytic mechanism of GH is not known. Consistent with earlier reports that GH is sulphydryl-sensitive, we found that recombinant human GH is inhibited by iodoacetic acid, suggesting that at least one cysteine is important for activity [Rhee, Lindau-Shepard, Chave, Galivan and Ryan (1998) Mol. Pharmacol. 53, 1040-1046]. Using site-directed mutagenesis, the cDNA for human GH was altered to encode four different proteins each with one of four cysteine residues changed to alanine. Three of the mutant proteins had activities similar to wild-type GH and were inhibited by iodoacetic acid, whereas the C110A mutant had no activity. Cys-110 is conserved among the human, rat and mouse GH amino acid sequences. The wild-type protein and all four mutants had similar intrinsic fluorescence spectra, indicating no major structural changes had been introduced. These results indicate that Cys-110 is essential for enzyme activity and suggest that GH is a cysteine peptidase. These studies represent the first identification of the essential Cys residue in this enzyme and provide the beginning of a framework to determine the catalytic mechanism, important in defining GH as a therapeutic target. PMID- 10527933 TI - A method for S- and O-palmitoylation of peptides: synthesis of pulmonary surfactant protein-C models. AB - A method for O- and S-palmitoylation of non-protected peptides has been developed. The peptides are treated with excess of palmitoyl chloride in 100% trifluoroacetic acid for 10 min at room temperature. The acidic conditions prevent acylation of amino groups, which is only significant after prolonged treatment (hours to days). The tripeptides Gly-Cys-Phe and Gly-Ser-Phe were converted into the respective S- and O-palmitoylated compounds, and the hydrophobic pulmonary surfactant protein-C model peptides, LRIPCCPVNLKRLLVVV [SP C(1-17)] and FGIPSSPVLKRLLILLLLLLLILLLILGALLMGL [SP-C(Leu)] were converted into their respective S,S- and O,O-dipalmitoylated peptides. The reactions were virtually quantitative, and the palmitoylated peptides were isolated in about 75 80% yield after reversed-phase HPLC purification. CD spectroscopy showed that S, S-dipalmitoylation of SP-C(1-17) affects the peptide secondary structure (substantial increase in the alpha-helix content) in dodecylphosphocholine micelles. PMID- 10527934 TI - An extremely thermostable aldolase from Sulfolobus solfataricus with specificity for non-phosphorylated substrates. AB - Sulfolobus solfataricus is a hyperthermophilic archaeon growing optimally at 80 85 degrees C. It metabolizes glucose via a novel non-phosphorylated Entner Doudoroff pathway, in which the reversible C(6) to C(3) aldol cleavage is catalysed by 2-keto-3-deoxygluconate aldolase (KDG-aldolase), generating pyruvate and glyceraldehyde. Given the ability of such a hyperstable enzyme to catalyse carbon-carbon-bond synthesis with non-phosphorylated metabolites, we report here the cloning and sequencing of the S. solfataricus gene encoding KDG-aldolase, and its expression in Escherichia coli to give fully active enzyme. The recombinant enzyme was purified in a simple two-step procedure, and shown to possess kinetic properties indistinguishable from the enzyme purified from S. solfataricus cells. The KDG-aldolase is a thermostable tetrameric protein with a half-life at 100 degrees C of 2.5 h, and is equally active with both d- and l-glyceraldehyde. It exhibits sequence similarity to the N-acetylneuraminate lyase superfamily of Schiff-base-dependent aldolases, dehydratases and decarboxylases, and evidence is presented for a similar catalytic mechanism for the archaeal enzyme by substrate dependent inactivation by reduction with NaBH(4). PMID- 10527935 TI - GLUT4 trafficking in insulin-stimulated rat adipose cells: evidence that heterotrimeric GTP-binding proteins regulate the fusion of docked GLUT4 containing vesicles. AB - Agents that activate the G-protein G(i) (e.g. adenosine) increase, and agents that activate G(s) [e.g. isoprenaline (isoproterenol)] decrease, steady-state insulin-stimulated glucose transport activity and cell-surface GLUT4 in isolated rat adipose cells without changing plasma membrane GLUT4 content. Here we have further examined the effects of R(s)G(s) and R(i)G(i) ligands (in which R(s) and R(i) are G(s)- and G(i)-coupled receptors respectively) on insulin-stimulated cell-surface GLUT4 and the kinetics of GLUT4 trafficking in these same cells. Rat adipose cells were preincubated for 2 min with or without isoprenaline (200 nM) and adenosine deaminase (1 unit/ml), to stimulate G(s) and decrease the stimulation of G(i) respectively, followed by 0-20 min with insulin (670 nM). Treatment with isoprenaline and adenosine deaminase decreased insulin-stimulated glucose transport activity by 58%. Treatment with isoprenaline and adenosine deaminase also resulted in similar decreases in insulin-stimulated cell-surface GLUT4 as assessed by both bis-mannose photolabelling of the substrate-binding site and biotinylation of the extracellular carbohydrate moiety when evaluated under similar experimental conditions. After stimulation with insulin in the absence of G(s) and the presence of G(i) agents, a distinct sequence of plasma membrane events took place, starting with an increase in immunodetectable GLUT4, then an increase in the accessibility of GLUT4 to bis-mannose photolabel, and finally an increase in glucose transport activity. Pretreatment with isoprenaline and adenosine deaminase before stimulation with insulin did not affect the time course of the increase in immunodetectable GLUT4 in the plasma membrane, but did delay both the increase in accessibility of GLUT4 to photolabel and the increase in glucose transport activity. These results suggest that R(s)G(s) and R(i)G(i) modulate insulin-stimulated glucose transport by influencing the extent to which GLUT4 is associated with occluded vesicles attached to the plasma membrane during exocytosis, perhaps by regulating the fusion process through which the GLUT4 in docked vesicles becomes exposed on the cell surface. PMID- 10527937 TI - The third chitinase gene (chiC) of Serratia marcescens 2170 and the relationship of its product to other bacterial chitinases. AB - The third chitinase gene (chiC) of Serratia marcescens 2170, specifying chitinases C1 and C2, was identified. Chitinase C1 lacks a signal sequence and consists of a catalytic domain belonging to glycoside hydrolase family 18, a fibronectin type III-like domain (Fn3 domain) and a C-terminal chitin-binding domain (ChBD). Chitinase C2 corresponds to the catalytic domain of C1 and is probably generated by proteolytic removal of the Fn3 and ChBDs. The loss of the C terminal portion reduced the hydrolytic activity towards powdered chitin and regenerated chitin, but not towards colloidal chitin and glycol chitin, illustrating the importance of the ChBD for the efficient hydrolysis of crystalline chitin. Phylogenetic analysis showed that bacterial family 18 chitinases can be clustered in three subfamilies which have diverged at an early stage of bacterial chitinase evolution. Ser. marcescens chitinase C1 is found in one subfamily, whereas chitinases A and B of the same bacterium belong to another subfamily. Chitinase C1 is the only Ser. marcescens chitinase that has an Fn3 domain. The presence of multiple, divergent, chitinases in a single chitinolytic bacterium is perhaps necessary for efficient synergistic degradation of chitin. PMID- 10527936 TI - Osmotic cell swelling-induced ATP release mediates the activation of extracellular signal-regulated protein kinase (Erk)-1/2 but not the activation of osmo-sensitive anion channels. AB - Human intestine 407 cells respond to hypo-osmotic stress by the rapid release of ATP into the extracellular medium. A difference in the time course of activation as well as in the sensitivity to cytochalasin B treatment and BAPTA-AM [1,2-bis (2-aminophenoxy)ethane-N,N,N',N'-tetra-acetic acid acetoxymethyl ester] loading suggests that ATP leaves the cell through a pathway distinct from volume regulated anion channels. To evaluate a putative role for nucleotides as autocrinic/paracrinic factors in osmotic signalling, the effects of extracellular ATP on the regulation of volume-sensitive anion channels as well as on the hypotonicity-induced activation of extracellular signal-regulated protein kinases (Erk-1/2) were investigated. Micromolar concentrations of ATP were unable to elicit an isotope efflux from (125)I(-)-loaded cells by itself, but strongly potentiated the hypotonicity-provoked anion efflux through a Ca(2+)-dependent mechanism. The order of potency of nucleotides (ATP = UTP = ATP[S] > ADP = AMP >> adenosine = cAMP) indicated the involvement of P2Y(2) receptors. In contrast, millimolar concentrations of ATP markedly inhibited both the osmotically induced isotope efflux and whole-cell Cl(-) currents. Inhibition of whole-cell Cl(-) currents, not only by millimolar ATP but also by the purinoceptor antagonists suramin and reactive blue, was observed most prominently at depolarizing holding potentials, suggesting a direct interaction with volume-sensitive Cl(-) channels rather than interaction with purinoceptors. Both ATP and UTP, at submicromolar levels, were found to act as potent activators of Erk-1/2 in intestine 407 cells. Addition of the ATP hydrolase apyrase to the bath greatly reduced the hypotonicity-induced Erk-1/2 activation, but did not affect the swelling-induced isotope efflux or whole-cell Cl(-) currents. Furthermore, pre-treatment with suramin or reactive blue almost completely prevented the hypo-osmotic activation of Erk-1/2. The results indicate that extracellularly released ATP functions as an autocrinic/paracrinic factor that mediates hypotonicity-induced Erk-1/2 activation but does not serve as an activator of volume-sensitive compensatory Cl(-) currents. PMID- 10527938 TI - Purification of multiple functional leaf-actin isoforms from Phaseolus vulgaris L. AB - Plant actins show diversity in their gene sequences, protein isovariants and tissue distribution in eukaryotes. Besides general difficulties with the isolation of proteins from plant material (i.e. the presence of a cell wall and high proteolytic activity), the actin concentration in any vegetative plant tissue is much lower than in cytoplasmic animal tissues. In this study, we adapted a deoxyribonuclease I-Sepharose affinity purification scheme and we were able to enrich and isolate multiple functional plant actin isovariants from common bean leaves (Phaseolus vulgaris). Urea (4 M) elution proved that the DNase I column was able to bind at least eight actin isoforms with pI values ranging from 5.5 to 5.9, as observed by two-dimensional Western blots. Three of the most acidic actin isoforms, with pI values of approximately 5.6-5.7, were eluted partially with 0.75 M urea. The purified actin was also able to bind leaf and rabbit muscle profilin, phalloidin and DNase I. Moreover, the protein could polymerize into filaments that contained the main isoforms eluted from the column. The average actin recovery using this procedure was approximately 4-8 microg from 20 g of fresh tissue, of which at least 80% was able to form filaments. This is the first report of the purification of multiple plant-actin isoforms that are functional by the criteria of both binding to other ligands and polymerization. PMID- 10527939 TI - NADPH as a co-substrate for studies of the chlorinating activity of myeloperoxidase. AB - The reinvestigation of the kinetics of myeloperoxidase (MPO) activity with the use of NADPH as a probe has allowed us to determine the effects of H(2)O(2), Cl( ) ion and pH on the MPO-dependent production of HOCl. The chlorination rate of NADPH did not depend on NADPH concentration and was entirely related to the rate of production of HOCl by MPO. The overall oxidation of NADPH occurred similarly in the absence of O(2) and was insensitive to scavengers of the superoxide radical anion. Experiments performed on the direct oxidation of NADPH by MPO in the presence and the absence of H(2)O(2) showed that neither the rate nor the stoichiometry of the reaction could interfere in the NADPH oxidation process involved in the steady-state chlorination cycle. The oxidation of NADPH was characterized by a decrease in the A(339) of the reduced nicotinamide with the concomitant appearance of a new chromophore with absorbance maximum at 274 nm, characterized by isosbestic points at 300 and 238 nm. The reaction product did not possess any enzymic properties with dehydrogenases and led to a metabolite other than NADP(+). Its amount accounted for a stoichiometric conversion of H(2)O(2) into HOCl. Analyses of the NADPH reaction allowed the determination of both kinetic (k(cat) and K(m)) and thermodynamic (K(d)) parameters. When the values of kinetic parameters were compared with previously published ones, the main discrepancy was found with data obtained with the chlorination of monochlorodimedon and a better agreement with diethanolchloramine formation or H(2)O(2) consumption. Variations in the extent of NADPH oxidation with Cl(-) concentration enabled us to determine the dissociation constant for the enzyme Cl(-) complex. In the course of titration studies, the spectral properties of NADPH reacting with either HOCl or the MPO/H(2)O(2)/Cl(-) system were quantitatively similar in terms of stoichiometry and absorbance coefficient and thus led to identical chlorinated products. However, no spectral modification occurred with NADP(+) and adenine nucleotide analogues under the same conditions. A quantitative comparison of difference spectra obtained with NADPH and NMNH indicated that chlorination occurred on the nicotinamide part of the molecule. PMID- 10527940 TI - Mitogen-activated protein kinase mediates erythropoietin-induced phosphorylation of the TAL1/SCL transcription factor in murine proerythroblasts. AB - Ectopic expression of the basic helix-loop-helix transcription factor TAL1 (or SCL) is the most frequent gain-of-function mutation in T-cell acute lymphoblastic leukaemia. Gene-knockout studies in mice have demonstrated that TAL1 is required for embryonic and adult haematopoiesis, and considerable evidence suggests it also has important functions in terminal erythroid differentiation. We reported previously that TAL1 phosphorylation is stimulated by erythropoietin in splenic proerythroblasts isolated from mice infected with the anaemia-inducing strain of Friend virus and show here the signalling pathway responsible. Erythropoietin was found to stimulate nuclear mitogen-activated protein kinase activity in addition to TAL1 protein phosphorylation, both of which were quantitatively inhibited by the mitogen-activated protein kinase kinase inhibitor PD 098059 and the phosphatidylinositol 3-kinase inhibitor wortmannin. Tryptic phosphopeptide analysis of radiolabelled TAL1 immunoprecipitated from nuclear extracts of Friend virus-induced proerythroblasts revealed that phosphorylation of Ser(122), shown previously to be a substrate for the mitogen-activated protein kinase ERK1 (extracellular signal-regulated protein kinase) in vitro, was specifically, although not exclusively, increased by erythropoietin and inhibited by wortmannin and PD 098059. These results are consistent with an erythropoietin-stimulated signalling pathway in which there is direct activation of a mitogen-activated protein kinase kinase by phosphatidylinositol 3-kinase and identify TAL1 as one of its nuclear targets. These data suggest, in addition, a specific mechanism by which the principal regulator of erythroid differentiation could enhance TAL1 function, in addition to increasing its expression. PMID- 10527941 TI - Opposite roles of trehalase activity in heat-shock recovery and heat-shock survival in Saccharomyces cerevisiae. AB - A variety of results has been obtained consistent with activation of neutral trehalase in Saccharomyces cerevisiae through direct phosphorylation by cAMP dependent protein kinase (PKA). A series of neutral trehalase mutant alleles, in which all evolutionarily conserved putative phosphorylation sites were changed into alanine, was tested for activation in vitro (by PKA) and in vivo (by glucose addition). None of the mutations alone affected the activation ratio, whereas all mutations combined resulted in an inactive enzyme. All mutant alleles were expressed to similar levels, as shown by Western blotting. Several of the point mutations significantly lowered the specific activity. Using this series of mutants with different activity levels we show an inverse relationship between trehalase activity and heat-shock survival during glucose-induced trehalose mobilization. This is consistent with a stress-protective function of trehalose. On the other hand, reduction of trehalase activity below a certain threshold level impaired recovery from a sublethal heat shock. This suggests that trehalose breakdown is required for efficient recovery from heat shock, and that the presence of trehalase protein alone is not sufficient for efficient heat-stress recovery. PMID- 10527942 TI - Glycosylphosphatidylinositol-anchor intermediates associate with triton-insoluble membranes in subcellular compartments that include the endoplasmic reticulum. AB - Glycosylphosphatidylinositol (GPI)-anchored proteins are resistant to solubilization with Triton X-100 at 4 degrees C, and they can be recovered in Triton-insoluble membranes (TIMs) that float to a characteristic buoyant density. Because the GPI structure itself has been shown to target GPI-anchored proteins to TIMs, we investigated the association of GPI-anchor intermediates with TIMs. GPI-anchor biosynthesis involves a pathway of some 10 steps that take place in the endoplasmic reticulum (ER). These intermediates include glucosaminyl acylphosphatidylinositol [GlcN-(acyl)PI] and later mannosylated GPIs, denoted H6, H7 and H8, that are present not only in the ER but also in other cell compartments, including the plasma membrane. At least two-thirds of the GlcN (acyl)PI in HeLa D cells and mannosylated GPIs in K562 cells were found in TIMs. Although previous reports have considered TIMs to be derived primarily from the plasma membrane, we recovered TIMs from subcellular fractions enriched in ER membranes. The ER marker calnexin and GPI-anchored proteins as well as N acetylglucosaminyl-phosphatidylinositol and mannosylated GPIs were present in ER TIMs. Interestingly, GlcN-PI and H7 were less enriched in ER-TIM than the other GPIs, suggesting that ER-TIMs might reflect a compartmentalization of the GPI anchor biosynthetic pathway in the ER. PMID- 10527943 TI - Identification of protein kinase C phosphorylation sites in the angiotensin II (AT1A) receptor. AB - Protein kinase C (PKC) phosphorylates the C-terminus of the type 1 angiotensin II receptor (AT(1)), although the exact site(s) of phosphorylation are unidentified. In the present study, we examined the phosphorylation of epitope-tagged wild-type AT(1A) receptors, transiently expressed in Chinese hamster ovary K1 cells, in response to angiotensin II (AngII) and following selective activation and inhibition of PKC. This phosphorylation was compared with mutant receptors where C-terminal serine residues (Ser(331), Ser(338) and Ser(348)) within three putative PKC consensus sites were replaced with alanine, either individually or in combination. Stimulation by AngII or the phorbol ester PMA to activate PKC induced an increase in phosphorylation of the wild-type AT(1A) receptor, which was prevented by truncation of the receptor C-terminus to remove the last 34 amino acids, including Ser(331), Ser(338) and Ser(348). Whereas single alanine mutation (Ser(331)Ala, Ser(338)Ala and Ser(348)Ala) resulted in decreased receptor phosphorylation, no single mutant completely inhibited either AngII- or PMA-induced phosphorylation. Combined mutation of the three PKC consensus sites caused an approximately 70% reduction in PMA-mediated phosphorylation. The approximately 60% reduction in AngII (1 microM)-induced phosphorylation of this triple mutant and the partial inhibition of wild-type receptor phosphorylation by bisindolylmaleimide, a specific PKC inhibitor, suggest a significant contribution of PKC to agonist-stimulated regulation. The ratio of PKC to total receptor phosphorylation was greatest at low doses of AngII (1 nM), consistent with the idea that PKC phosphorylates and regulates receptor function at low levels of stimulation, whereas phosphorylation by other kinases is more prevalent at high levels of agonist stimulation. To determine if a single PKC site is favoured when the contribution of PKC varies, the phosphorylation of wild-type and mutant receptors was examined over a range of AngII concentrations (0, 1, 10 and 100 nM). At all AngII concentrations, single mutation of Ser(331), Ser(338) or Ser(348) was incapable of completely preventing receptor phosphorylation, suggesting no clear preference for PKC consensus-site utilization. Together, these results indicate a redundancy in PKC phosphorylation of the AT(1A) receptor, whereby all three consensus sites are utilized to some degree following homologous (AngII) and heterologous (PMA) stimulation. The contribution of PKC phosphorylation to receptor regulation is unclear, but multiple PKC phosphorylation of the AT(1A) receptor may allow independent and/or complementary events to occur at the three separate sites of the C-terminus. PMID- 10527944 TI - Structural characterization of human and bovine lung surfactant protein D. AB - Human and bovine surfactant proteins D (SP-D) were purified from late amniotic fluid and bronchioalveolar lavage on the basis of its Ca(2+)-dependent affinity for maltose. The molecular mass of a trimeric subunit was determined by matrix assisted laser desorption ionization MS to lie in the range 115-125 kDa for human SP-D and 110-123 kDa for bovine SP-D. A single polypeptide chain was determined at 37-41 and 36-40 kDa for the human and bovine species respectively. The major parts of the primary structures of both SP-D molecules were determined by a combination of MS and Edman degradation. The heterogeneity in SP-D was caused mainly by a high number of post-translational modifications in the collagen-like region. Proline and lysine residues were partly hydroxylated and lysine residues were further O-glycosylated with the disaccharide galactose-glucose. A partly occupied N-linked glycosylation site was characterized in human SP-D. The carbohydrate was determined as a complex type bi-antennary structure, with a small content of mono-antennary and tri-antennary structures. No sialic acid residues were present on the glycan, but some had an attached fucose and/or an N acetylglucosamine residue linked to the core. Bovine SP-D was determined as having a similar structure. PMID- 10527945 TI - Conserved charged residues in the leucine-rich repeat domain of the Ran GTPase activating protein are required for Ran binding and GTPase activation. AB - GTPase activating proteins (GAPs) for Ran, a Ras-related GTPase participating in nucleocytoplasmic transport, have been identified in different species ranging from yeast to man. All RanGAPs are characterized by a conserved domain consisting of eight leucine-rich repeats (LRRs) interrupted at two positions by so-called separating regions, the latter being unique for RanGAPs within the family of LRR proteins. The cytosolic RanGAP activity is essential for the Ran GTPase cycle which in turn provides directionality in nucleocytoplasmic transport, but the structural basis for the interaction between Ran and its GAP has not been elucidated. In order to gain a better understanding of this interaction we generated a number of mutant RanGAPs carrying amino acid substitutions in the LRR domain and analysed their complex formation with Ran as well as their ability to stimulate the intrinsic GTPase activity of the G protein. We show that conserved charged residues present in the separating regions of the LRR domain are indispensable for efficient Ran binding and GAP activity. These separating regions contain three conserved arginines which could possibly serve as catalytic residues similar to the arginine fingers identified in GAPs for other small GTPases. However, mutations in two of these arginines do not affect the GAP activity and replacement of the third conserved arginine (Arg91 in human RanGAP) severely interferes not only with GAP activity but also with Ran binding. This indicates that RanGAP-stimulated GTP hydrolysis on Ran does not involve a catalytic arginine residue but requires certain charged residues of the LRR domain of the GAP for mediating the protein-protein interaction. PMID- 10527946 TI - Expression of proteoglycan core proteins in human bone marrow stroma. AB - Heparan sulphate proteoglycans (HSPGs) present on the surface of bone marrow stromal cells and in the extracellular matrix (ECM) have important roles in the control of adhesion and growth of haemopoietic stem and progenitor cells. The two main groups of proteoglycans which contain heparan sulphate chains are members of the syndecan and glypican families. In this study we have identified the main surface membrane and matrix-associated HSPGs present in normal human bone marrow stroma formed in long-term culture. Proteoglycans were extracted from the adherent stromal layers and treated with heparitinase and chondroitinase ABC. The core proteins were detected by Western blotting using antibodies directed against syndecans-1-4, glypican-1 and the ECM HSPG, perlecan. Stromal cell expression at the RNA level was detected by Northern blotting and by reverse transcription PCR. Glypican-1, syndecan-3 and syndecan-4 were the major cell-membrane HSPG species and perlecan was the major ECM proteoglycan. There was no evidence for expression of syndecan-1 protein. Syndecan-3 was expressed mainly as a variant or processed 50-55 kDa core protein and in lower amounts as the characteristic 125 kDa core protein. These results suggest that syndecan-3, syndecan-4 and glypican-1 present on the surface of marrow stromal cells, together with perlecan in the ECM, may be responsible for creating the correct stromal 'niche' for the maintenance and development of haemopoietic stem and progenitor cells. The detection of a variant form of syndecan-3 as a major stromal HSPG suggests a specific role for this syndecan in haemopoiesis. PMID- 10527947 TI - Properties of the 40 kDa antigen of Mycobacterium tuberculosis, a functional L alanine dehydrogenase. AB - The 40 kDa antigen of Mycobacterium tuberculosis is the first antigen reported to be present in the pathogenic M. tuberculosis, but not in the vaccine strain Mycobacterium bovis BCG. It is a functional L-alanine dehydrogenase (EC 1.4.1.1) and hence one of the few antigens possessing an enzymic activity. This makes the 40 kDa antigen attractive for potential diagnostic and therapeutic interventions. Recently, we developed a strategy to purify quantities of the recombinant protein in active form, and here we describe the biochemical properties of this enzyme. In the oxidative-deamination reaction, the enzyme showed K(m) values of 13. 8 mM and 0.31 mM for L-alanine and NAD(+), respectively, in a random-ordered mechanism. K(m, app) values in the reductive-amination reaction are 35.4 mM, 1.45 mM and 98.2 microM for ammonium, pyruvate and NADH, respectively. The enzyme is highly specific for all of its substrates in both directions. The pH profile indicates that oxidative deamination virtually may not occur at physiological pH. Hence L-alanine most likely is the product of the reaction catalysed in vivo. The enzyme is heat-stable, losing practically no activity at 60 degrees C for several hours. PMID- 10527949 TI - The role of amino acid alpha38 in the control of oxygen binding to human adult and embryonic haemoglobin Portland. AB - The role of the amino acid at position alpha(38) in haemoglobin has been probed using site-directed mutagenesis. When the Thr residue at position alpha(38) (which is totally conserved in all mammals) is changed to a Gln, the equilibrium properties of the protein are significantly altered. Equilibrium and kinetic data show that the R-state properties of the protein are essentially unaffected by the mutation whilst the allosteric equilibrium and T-state properties are changed. Mutation of the naturally occurring Gln(38) of the human embryonic haemoglobin zeta-chain (the only known non-Thr containing globin) to a Thr residue shows the converse change in properties produced by the adult mutation, although in this case the situation is complicated by significant chain heterogeneity in the T state. An extension of the two-state model of co-operativity is presented to describe quantitatively the equilibrium ligand binding in the presence of T-state chain heterogeneity. A molecular model is described in which the putative interaction of alphaGln(38) and betaTyr(145) is identified which make a significant contribution to the previously reported unusual ligand-binding properties of the zeta-chain containing human embryonic haemoglobins. PMID- 10527948 TI - Evidence for an interaction of the metalloprotease-disintegrin tumour necrosis factor alpha convertase (TACE) with mitotic arrest deficient 2 (MAD2), and of the metalloprotease-disintegrin MDC9 with a novel MAD2-related protein, MAD2beta. AB - Metalloprotease-disintegrins are a family of transmembrane glycoproteins that have a role in fertilization, sperm migration, myoblast fusion, neural development and ectodomain shedding. In the present study we used the yeast two hybrid system to search for proteins that interact with the cytoplasmic domain of two metalloprotease-disintegrins, tumour necrosis factor alpha convertase (TACE; ADAM17) and MDC9 (ADAM9; meltrin gamma). We have identified mitotic arrest deficient 2 (MAD2) as a binding partner of the TACE cytoplasmic domain, and a novel MAD2-related protein, MAD2beta, as a binding partner of the MDC9 cytoplasmic domain. MAD2beta has 23% sequence identity with MAD2, which is a component of the spindle assembly (or mitotic) checkpoint mechanism. Northern blot analysis of human tissues indicates that MAD2beta mRNA is expressed ubiquitously. The interaction of the TACE and MDC9 cytoplasmic domains with their binding partners has been confirmed biochemically. The independent identification of MAD2 and MAD2beta as potential interacting partners of distinct metalloprotease-disintegrins raises the possibility of a link between metalloprotease-disintegrins and the cell cycle, or of functions for MAD2 and MAD2beta that are not related to cell cycle control. PMID- 10527950 TI - Delineation of the insulin-responsive sequence in the rat cytosolic aspartate aminotransferase gene: binding sites for hepatocyte nuclear factor-3 and nuclear factor I. AB - Expression of the rat cytosolic aspartate aminotransferase gene is stimulated by glucocorticoids and repressed by insulin in the liver. The regulation by insulin and part of the glucocorticoid effect are mediated by a distal region in the promoter. A 142 bp fragment (-1844 to -1702) confers hormonal sensitivity to the heterologous thymidine kinase promoter in transient-transfection assays in H4IIEC3 hepatoma cells. Footprinting and gel-shift assays showed that several nuclear proteins bind to this region at conserved CCAAT-enhancer binding protein (C/EBP), activator protein (AP-1) and E-box sequences. Hepatocyte nuclear factor 3alpha (HNF-3)alpha and beta bind to sequences upstream of a glucocorticoid responsive element (GRE) half-site as demonstrated by supershift experiments. Nuclear factor I (NFI)-like proteins bind downstream of the GRE half-site. These sites around the GRE motif overlap with five insulin responsive element (IRE) like sequences (TG/ATTT). The effect of insulin was not prevented by any single mutation in the IRE-like sites. However, mutation of two IRE sites (namely IREc and d) prevented the insulin effect although only marginally affecting the glucocorticoid effect. The results suggest that the effect of insulin is due to a complex interplay of factors requiring the synergistic contribution of at least two sites and underline the contribution of HNF-3 and NFI-like proteins. PMID- 10527951 TI - A novel chaperone-activity-reducing mechanism of the 90-kDa molecular chaperone HSP90. AB - The 90-kDa heat shock protein (HSP90) acts as a specific molecular chaperone in the folding and regulates a wide range of associated proteins such as steroid hormone receptors. It is known that HSP90 possesses two different chaperone sites, both in the N- and C-domains, and that the chaperone activity of HSP90 is blocked by binding of geldanamycin (GA) to the N-domain, the same as the ATP binding site. Here we show that Cisplatin [cis-diamminedichloroplatinum (II), CDDP], an antineoplastic agent, associates with HSP90 and reduces its chaperone activity. In order to analyse the binding proteins, bovine brain cytosols were applied to a CDDP-affinity column and binding proteins were eluted by CDDP. In the elutants, only 90-kDa protein bands were detected on SDS/PAGE, and the protein was cross-reacted with the anti-HSP90 antibody on immunoblotting. No protein bands were detected in the elutants from the control column on SDS/PAGE. These results indicated that CDDP has a high affinity for HSP90. On CD spectrum analysis, the binding of CDDP to HSP90 resulted in a conformational change in the protein. Although HSP90 inhibited the aggregation of citrate synthase as a molecular chaperone in vitro, the activity was suppressed almost completely in the presence of CDDP. Mg/ATP has an influence on the chaperone activity to some extent. The CDDP binding region of HSP90 is near the C-terminal which is quite different from the GA-binding site. Our results suggest that the chaperone activity of HSP90 may be inhibited by the binding of CDDP or GA by different mechanisms. PMID- 10527954 TI - Bronchodilation using combined urodilatin - albuterol administration in asthma: a randomized, double-blind, placebo-controlled trial. AB - Urodilatin, the renal form of natriuretic peptide type A, induces bronchodilation, increasing intracellular cyclic guanosine monophosphate (cGMP), whereas the bronchorelaxant effect by b2-agonists is triggered by cyclic adenosine monophosphate (cAMP). The objective of this investigation is to demonstrate the efficacy of urodilatin in inducing bronchodilation, and to show this activity alone or in combination with albuterol. Therefore, a randomized, double-blind, placebo-controlled, dose-finding study with cross-over design was carried out including 12 stable, mild to severe (step 2 to 4, definition by NIH/NHLBI guideline 1997) asthmatics. 96 treatments were thus performed. The intervention was comprised of an intravenous infusion of urodilatin (0, 10, 30, or 60 ng/kg/min) combined with inhaled albuterol (0 or 200 microg). As primary objective, the increase in forced expiratory volume in one second (FEV subset1) was measured. - The trial shows that urodilatin at all applied doses or 200 microg albuterol significantly increases FEV subset1 (p < 0.05). Combination of urodilatin and albuterol treament significantly improves FEV subset1 (p < 0.05) compared to either monotherapy and results in maximum bronchodilation. - From the results, the following conclusions can be drawn. In stable asthmatics, the combined activation of cGMP- and cAMP-mediated pathways results in a significantly improved, maximal bronchodilation in comparison to either type of monotherapy. This shows that urodilatin combined with albuterol improves lung function and ameliorates the therapy in asthmatics. PMID- 10527953 TI - Fibroblast growth factor protects the kidney against ischemia-reperfusion injury. AB - Ischemia-reperfusion injury, a common source of renal dysfunction in adults, is associated with tubular epithelial cell damage. Since fibroblast growth factors (FGF) attenuated tissue injury after transient myocardial ischemia, we hypothesized that acidic fibroblast growth factor (aFGF; FGF-1) would attenuate renal ischemia-reperfusion injury. We studied the effects of FGF-1 in a rat model of acute renal failure induced by bilateral renal ischemia (60 min) and 1, 2 or 7 days reperfusion. After FGF-1 administration at the onset of renal reperfusion, there was less functional impairment of the kidneys. The histological changes were not as severe as in controls. Increases in serum creatinine and blood urea nitrogen 24 h after reperfusion were attenuated by 35% (p< 0.01) and by 53% (p< 0.001), respectively, in FGF-1-treated animals compared to vehicle-treated rats. The ischemia/reperfusion-induced increase in tissue myeloperoxidase, a marker of neutrophil infiltration, was mitigated (67% reduction, p< 0.05) with FGF-1 treatment. As shown by histology, neutrophil infiltration and tubular cell necrosis in medulla were less pronounced (p< 0.0001 and p< 0.05, respectively) in animals receiving FGF-1. Furthermore, ischemia-induced apoptosis, prevalent in tubular cells of the cortex, was also attenuated by FGF-1-treatment (83% reduction, p< 0.0001). Pretreatment of animals with Nw-nitro-L-arginine (L-NNA), an inhibitor of nitric oxide synthase, abolished the attenuating effects of FGF-1 on neutrophil infiltration, suggesting that nitric oxide might participate in the anti-inflammatory effects of FGF-1 in this experimental design. Our data support a role for FGF-1 in attenuation of renal damage or failure after ischemia reperfusion injury of the kidney, in part at least by inhibition of neutrophil infiltration. PMID- 10527955 TI - Immunologic and virologic studies in long-term nonprogressors with HIV-1 infection. AB - BACKGROUND: It is not known whether clinical latency in long-term nonprogressors (LTNP) with HIV-1 infection is due to a strong HIV-1 specific immune response of the host or to virologic factors. - METHODS: Peripheral blood mononuclear cells (PBMC) of 6 LTNP were analyzed for their phenotype, proliferation rates, natural killer (NK) cell activity, antibody dependent cellular cytotoxicity (ADCC), and CCR5 chemokine receptor genotype. Furthermore sequence analyses of the HIV-1 gene were performed. - RESULTS: Phenotypic analyses of lymphocyte subsets revealed increased CD8+ as well as HLA-DR+ expressing cells in LTNP and patients with progressive disease (PRO). Proliferation assays in LTNP and PRO showed a reduction of stimulation by polyclonal mitogens (PHA, ConA, PWM) of up to 60%. NK activity was within normal ranges in LTNP but reduced in PRO. 1 LTNP exhibited heterozygosity for CCR5-D32. A mutant HIV nef gene was not discovered by PCR in any of the LTNP. HIV-V3 loop PCR in 5 LTNP revealed the HIV-1B (NSI) subtype. In 2 patients further sequence analyses of the HIV-1 genome showed homozygous mutations in the Sp1 and NF-kB binding sites. CONCLUSION: The non-progression of HIV-1 infection in some LTNP seems to be due to single mutations in the viral genome resulting in a less replicative HIV-1 subtype or to a mutant chemokine receptor leading to a reduced HIV-1 entry into CD4+ cells. NK cell activity might be an additional contributing factor in controlling viremia. PMID- 10527956 TI - Influence of folic acid, pyridoxal phosphate and cobalamin on plasma homocyst(e)ine levels and the susceptibility of low-density lipoprotein to ex vivo oxidation. AB - Mild hyperhomocyst(e)inaemia is a risk factor for atherosclerotic vascular disease. In-vitro studies have shown that autooxidation of homocyst(e)ine is accompanied by the generation of oxygen radicals. This may lead to oxidative modification of low-density lipoproteins (LDL) and promote atherosclerotic vascular lesions. In male patients with peripheral arterial occlusive disease we determined fasting and post methionine load homocyst(e)ine levels by high performance liquid chromatography and the susceptibility of their LDL particles to ex-vivo oxidation by continously measuring the conjugated diene production induced by incubation with copper ions. Oxidation resistance (expressed as lag time), maximal oxidation rate, and extent of oxidation (expressed of total diene production) of LDL from patients with normal or mildly elevated homocyst(e)ine levels did not differ significantly. Folic acid, pyridoxal phosphate and cobalamin supplementation significantly decreased plasma homocyst(e)ine levels in hyperhomocyst(e)inaemic patients. This went along with a significant decrease in the extent of LDL oxidation and additionally increased HDL-cholesterol levels. The clinical relevance of these findings for the long-term course of atherosclerotic vascular disorders has to be determined by intervention studies. PMID- 10527957 TI - Acute effects of LDL-apheresis on cholesterol oxidation products and antioxidants in plasma and lipoproteins of patients with familial hypercholesterolemia. AB - Regular LDL-apheresis treatment of hypercholesterolemic patients has proven to reduce the formation of atherosclerotic lesions. Regarding the underlying mechanisms, cholesterol oxidation products (COP) may play a detrimental role. Therefore, COP levels were determined before and after regular LDL-apheresis treatment in ten patients with familial hypercholesterolemia. - The patients had approximately twofold elevated plasma and LDL COP concentrations on the average as compared to healthy subjects. LDL-apheresis treatment efficiently removed COP from the circulation. As a consequence of a smaller reduction of the COP content (- 52 %) than of the total cholesterol content (-71 %) in LDL, the LDL COP:cholesterol ratio increased. Lipid-soluble antioxidants in the plasma of the hypercholesterolemics decreased to a comparable extent as did plasma lipids. In contrast to nearly stable vitamin C concentrations, plasma selenium concentrations also decreased, resulting altogether in a decreased but still normal serum total antioxidant capacity. - In conclusion, LDL-apheresis treatment effectively reduced potentially atherogenic COP from the plasma. With normal plasma antioxidant concentrations before LDL-apheresis in long-term treated hypercholesterolemics, the observed acute decrease in lipid-soluble antioxidants and selenium by treatment seems not to be as meaningful. The higher LDL COP:cholesterol ratio after treatment needs further elucidation. PMID- 10527952 TI - Regulation of intestinal Na+-dependent phosphate co-transporters by a low phosphate diet and 1,25-dihydroxyvitamin D3. AB - In a study of the rat intestinal P(i) transport system, an activator protein for rat Na/P(i) co-transport system (PiUS) was isolated and characterized. We also investigated the effects of restriction of vitamin D and P(i) (two of the most important physiological and pathophysiological regulators of P(i) absorption in the small intestine) on intestinal P(i) transport activity and the expression of Na/P(i) co-transporters that are expressed in rat small intestine. Rat PiUS encodes a 424-residue protein with a calculated molecular mass of 51463 Da. The microinjection of rat PiUS into Xenopus oocytes markedly stimulated Na(+) dependent P(i) co-transport activity. In rats fed with a low-P(i) diet, Na(+) dependent P(i) co-transport activity was increased approx. 2-fold compared with that of rats fed a normal P(i) diet. Kinetic studies demonstrated that this increased activity was due to an elevation of V(max) but not K(m). The PiUS mRNA levels showed an approximate doubling in the rats fed with the low-P(i) diet compared with those fed with the normal P(i) diet. In addition, after the administration of 1, 25-dihydroxyvitamin D(3) [1,25-(OH)(2)D(3)] to vitamin D deficient animals, the P(i) uptake was significantly increased in the Na(+) dependent component in the brush border membrane vesicle (BBMV) at 24 and 48 h. In addition, we found a further high-affinity Na/P(i) co-transport system in the BBMV isolated from the vitamin D-replete animals. The levels of type III Na/P(i) co-transporter PiT-2 mRNA were increased 24 and 48 h after 1,25-(OH)(2)D(3) administration to vitamin D-deficient animals, whereas PiUS and the type IIb Na/P(i) co-transporter mRNA levels were unchanged. In conclusion, we first cloned a rat activator protein, PiUS, and then studied its role along with that of other type III Na/P(i) co-transporters. PiUS and PiT-2 might be important components in the regulation of the intestinal P(i) transport system by P(i) restriction and 1,25-(OH)(2)D(3). PMID- 10527958 TI - Influence of methotrexate and azathioprine on radiologic progression in rheumatoid arthritis. AB - We investigated 43 patients with seropositive rheumatoid arthritis (RA) under a therapy with methotrexate (MTX) or azathioprine (AZA). All patients fulfilled the American Rheumatism Association criteria. It has been a retrospective study over 2.6 years (MTX: 26 patients, average age 54 +/- 14.6 years, female/male 19:7, AZA: 17 patients, average age 59.4 +/- 12.5 years, female/male 12:5). The mean duration of disease was 8 years (+/- 5.3) in the MTX-group and 7 years (+/- 7.3) in the AZA-group. The mean dose of MTX was 13. 3mg/week and of azathioprine 142.6mg/ daily. The drugs were administered orally. - Radiographs of hands, wrists and feet obtained at enrollment and at review were scored by a rheumatologist according to a modified Larsen score. Radiographic damages were counted in 25 joints of each hand and in 25 joints of each foot. The change in radiological score was calculated by subtracting the joint damage scores at follow up with first damage score at inclusion. The rate of radiological progression (YP) was calculated by dividing the change in radiological score by the number of years during the period of study. - Only 3 patients with MTX showed no radiologic progression. All other patients of both groups showed progressive radiological changes. - The study demonstrated no significant difference in the rate of radiologic progression between the different treatment-groups. However, there was a trend that MTX treated patients (YP 5 +/- 4.4) had a slower radiographic progression compared with those treated with AZA (YP 8.5 +/- 7.7). MTX may be more effective in patients at an earlier stage of rheumatoid arthritis. - When we started a therapy with AZA or MTX in a later period of disease we revealed a better influence of radiologic progression under AZA and a trend towards an increase of the radiologic progression under MTX. - Probably there is a decreasing effect of MTX in later periods of the disease. - The corticoid dose reduction was higher under AZA (AZA: Reduction about 58.6%, MTX: 25%) over the study duration. - Our investigation demonstrated a trend towards reduced radiological progression in MTX treated patients compared to AZA, however statistic analysis showed no significant difference in the rate of radiologic progression. At an earlier stage of the disease there is a better influence of MTX in radiologic progression, at the later stage we showed a slowing of radiological deterioration in AZA treated group. PMID- 10527959 TI - Severe OHSS: decreasing the risk of severe ovarian hyperstimulation syndrome. PMID- 10527960 TI - Antisperm antibodies. Antisperm antibodies and infertility: an unsolvable question? PMID- 10527961 TI - Antisperm antibodies. Do antisperm antibodies bound to spermatozoa alter normal reproductive function? PMID- 10527962 TI - WHO manual...who should care? PMID- 10527963 TI - Do human concepti have the potential to enter into diapause? AB - Although there is no direct evidence as to whether human concepti have the potential to enter into diapause before implantation, the possibility that human concepti may be capable of following this developmental pathway if exposed to an appropriate environment cannot be ruled out. Direct evidence remains elusive because of the ethical restraints associated with research activities within this area of knowledge. If conceptus diapause has evolved in primates and persists at the present time despite its apparent limited or no adaptive advantage, artificial induction of diapause in humans may have clinical implications for increasing: (i) the viability of concepti after biopsy, freezing-thawing or any other experimental procedure that tends to decrease cell numbers or division rate of concepti; and (ii) the relatively low implantation rates obtained at the present time after uterine transfer of human concepti fertilized in vitro. Furthermore, conceptus diapause may be a good paradigm to understand the interplay between the different genetic/molecular components of both the conceptus and endometrium at implantation. PMID- 10527964 TI - Prediction of severe ovarian hyperstimulation syndrome by free serum vascular endothelial growth factor concentration on the day of human chorionic gonadotrophin administration. AB - In this prospective study the concentration of circulating vascular endothelial growth factor (VEGF) was followed in 10 patients with severe ovarian hyperstimulation syndrome (OHSS) after ovarian stimulation and in 15 patients without OHSS. VEGF was assayed by means of two different commercially available kits as either free or total VEGF in serum. The concentration of free VEGF was significantly higher on the days of human chorionic gonadotrophin (HCG) administration (309.4 +/- 165.0 versus 190.3 +/- 127.8 pg/ml, P < 0.05) and embryo transfer (315.0 +/- 125.2 versus 209.3 +/- 137. 2 pg/ml, P < 0.05) in the OHSS compared to the control group. No such difference existed with respect to total circulating VEGF. In addition, there was no significant rise in the free or in the total serum VEGF concentration in the OHSS patients or the controls from the day of HCG administration up to the days of oocyte retrieval or embryo transfer. A cut-off concentration of 200 pg/ml free serum VEGF concentration on the day of HCG treatment resulted in a sensitivity of 90% and a specificity of 80% for the prediction of OHSS development. This is the first report on the parallel measurement of free and total VEGF in serum following ovarian stimulation. The value of the proposed cut-off concentration should be confirmed in a study of a larger group of women. PMID- 10527966 TI - Factor V leiden and factor II G20210A mutations in patients with recurrent abortion. AB - Recurrent abortion (RA) represents an intriguing problem in obstetric practice in which genetic and acquired factors may play a role. In the present investigation we sought to assess the possibility that inherited thrombophilia might determine the risk of RA. We therefore investigated the prevalence of two genetic abnormalities frequently associated with venous thrombosis [factor V Leiden (FVL) and factor II G20210A] in 56 patients with primary or secondary abortion and in 384 healthy control women. Polymerase chain reaction amplification followed by digestion with the restriction enzymes MnlI and HindIII was used to define the FVL and FII G20210A genotypes respectively. FVL was found in 4/56 patients (7.1%) and in 6/384 controls (1.6%), yielding an odds ratio (OR) for RA related to FVL of 4.9 [95% confidence interval (CI): 1.3-17.8]. FII G20210A was detected in 2/56 (3.6%) patients and in 4/384 (1%) controls (OR for RA: 3.5, CI: 0.6-19.7). In conclusion, FVL and FII G20210A mutations in patients with RA were more prevalent in comparison with controls. These data support a role for both mutations as determinants of the risk of RA and strengthen the notion that thrombophilia plays a role in this clinical entity. PMID- 10527965 TI - A prospective, randomized clinical trial comparing 150 IU recombinant follicle stimulating hormone (Puregon((R))) and 225 IU highly purified urinary follicle stimulating hormone (Metrodin-HP((R))) in a fixed-dose regimen in women undergoing ovarian stimulation. AB - A prospective, randomized, open, multicentre (n = 3) study was conducted to compare the efficacy and efficiency of a fixed daily dose of 150 IU (3x50 IU) recombinant follicle stimulating hormone (recFSH, Puregon((R))) and 225 IU (3x75 IU) highly purified urinary FSH (uFSH-HP, Metrodin-HP((R))) in women undergoing ovarian stimulation prior to in-vitro fertilization treatment. A total of 165 women were treated with FSH, 83 subjects with recFSH and 82 subjects with uFSH HP. In the recFSH group a mean number of 8.8 oocytes were retrieved, compared with 9.8 in the uFSH-HP group (not statistically significant). In the recFSH group, a significantly lower total dose was required compared to the uFSH-HP group, 1479 versus 2139 IU, respectively (P < 0.0001; 95% confidence interval 747 to -572). Treatment with recFSH resulted in a significantly higher embryo development rate (69.6 versus 56.2%; P = 0.003) and more embryos accessible for the embryo freezing programme (3.3 versus 2.0; P = 0.02) compared to uFSH-HP. The vital pregnancy rate per cycle started was 30.2 versus 28.3% in the recombinant and urinary FSH group, respectively. It is concluded that treatment outcome of a fixed daily dose of 150 IU recFSH is comparable to a fixed daily dose of 225 IU uFSH-HP. However, a significantly lower total dose was needed in the recFSH group (nearly 700 IU less). PMID- 10527967 TI - No mutations found in candidate genes for dystocia. AB - Dystocia is a disorder characterized by prolonged or dysfunctional labour. Delivery that starts late or not at all, leads to an increased risk for Caesarean section, infant morbidity and mortality. Familial aggregations of dystocia suggest a polygenic background. We have studied three candidate genes for dystocia, i.e. the genes for testosterone 5-alpha reductase type 1, prostaglandin F2alpha receptor and endothelin 1 and performed mutational screening in 23 women with dystocia, of which 12 have affected relatives. No mutations were found, making it unlikely that any of these genes represent a major cause of dystocia in man. PMID- 10527968 TI - Familial idiopathic premature ovarian failure: an overrated and underestimated genetic disease? AB - The incidence of familial cases of premature ovarian failure varies from 4 to 31%. Recall bias may explain part of the variance. Thorough evaluation of alleged affected relatives showed a lower incidence than the original family history suggested. In the present study the incidence of familial cases was 12.7%. Pedigree studies on affected families showed a mode of inheritance suggestive of autosomal dominant sex-limited transmission or X-linked inheritance with incomplete penetrance. An adequate family history can distinguish between familial or sporadic premature ovarian failure. The risk of female relatives developing premature ovarian failure may be as high as 100% in familial premature ovarian failure, or as low as 1% in sporadic cases. PMID- 10527969 TI - Ultrasound evaluation of uterine wound healing following laparoscopic myomectomy: preliminary results. AB - The purpose of our work was to study the evolution of the uterine scar following laparoscopic myomectomy, as imaged by ultrasonography and Doppler velocimetry of the uterine arteries. We prospectively studied 30 patients. In the first phase, 15 patients were submitted to two-dimensional (2D) endovaginal ultrasound on day 1, 1, 7, 30 and 60 (surgery = day 0). In the second phase an additional 15 patients were studied by both 2D ultrasound and by Doppler velocimetry. The resistance index (RI) was calculated from the flow velocity waveform of the uterine arteries, at the origin of their ascending branch. Only one ultrasonic pattern was found, which was a dense echogenic area having an ill-defined, heterogeneous texture. In one case a small anechoic area (1 cm) was detected in the scar, possibly due to a haematoma. The evolution of uterine healing showed a progressive reduction in the size of the scar. On day 1 its mean diameter was 37.04% less than the myoma diameter and on day 30 71.7% less. The difference was significant at P < 0.001. A further significant (P < 0.001) reduction was found at day 60 in the 15 patients studied in phase I. On both day 1 and day 30 following surgery, there was no correlation between the sizes of the myoma and the scar. There was a statistically significant increase (P < 0.01) in the RI value of the ipsilateral uterine artery from 0.64 on day -1 to 0.79 on day 1. On day 30, 12/15 (80%) cases had RI values ranging between 0.80 and 0.98, while in three cases there was absence of end diastolic flow. The RI values of the contralateral uterine artery were high (0.90) before surgery and did not change afterwards. There was no correlation between the size of the myoma and the increase in the uterine artery RI value following surgery. Considering the velocimetric findings, 30 days are a reference point for assessing the healing process. Ultrasound imaging and Doppler velocimetry can be used for studying the evolution of the uterine scar following myomectomy. PMID- 10527970 TI - Hysterectomy techniques used for benign pathologies: results of a French multicentre study. AB - The objective of this study was to assess the techniques by which hysterectomies are carried out and to determine the rate of total laparoscopic hysterectomy (TLH). A transversal multicentre study was conducted in 23 gynaecology and obstetrics departments of French University Hospital Centres. The study population comprised only those patients for whom hysterectomy was indicated for benign disease without genital prolapse or urinary stress incontinence. Whereas the rates of performance of hysterectomy by laparotomy and by the vaginal route are comparable [respectively 40.0% (94 patients) and 46.8% (110 patients)], the rate of performance of TLH is only 13.2% (31 patients). All 23 centres (100%) carried out hysterectomy by laparotomy and 21 centres (91.3%) carried out vaginal hysterectomy; however, only nine centres (39.1%) carried out TLH. Only seven centres (30.4%) performed all three types of operation. Of the eight centres whose rate of vaginal hysterectomy was >60%, six (75%) did not carry out TLH. The study suggests that the usage of the TLH technique appears to be limited. The extent of surgical training is a major factor in the choice of technique for hysterectomy. PMID- 10527971 TI - Spontaneous bilateral cornual uterine dehiscence early in the second trimester after bilateral laparoscopic salpingectomy and in-vitro fertilization: case report. AB - A bilateral cornual uterine dehiscence is reported, which occurred 14 weeks after in-vitro fertilization (IVF) in a patient having a medical history of previous bilateral salpingectomy via laparoscopy. Uterine rupture is a rare obstetric complication usually occurring during the third trimester of pregnancy within a uterus which has previously undergone an operation. Ectopic pregnancy is a well known complication of IVF. Post-salpingectomy cornual localization with rupture has also been published. Possible causes are discussed and the attention of the counselling physician is directed to the necessary awareness of such a complication in this high risk population. The reported case is an extreme rarity: a similar case has not been previously published in the literature. PMID- 10527972 TI - Conventional in-vitro fertilization versus intracytoplasmic sperm injection in sibling oocytes from couples with tubal infertility and normozoospermic semen. AB - An auto-controlled study was conducted in couples with tubal infertility and normozoospermic semen. The fertilization rates and embryonic development in sibling oocytes treated, using the same semen sample, either by conventional in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) at the same time were compared. Sibling oocyte-cumulus complexes (OCC) of 56 different couples with tubal infertility and normozoospermic semen were randomly divided in order of retrieval into two groups inseminated either by conventional IVF or by ICSI. Of the retrieved OCC in the same cohort, 53.0 +/- 31.2 and 62.0 +/- 26.6% showed two distinct pronuclei after conventional IVF and ICSI respectively (not significant). Complete fertilization failure occurred after conventional IVF in 12.5% (7/56 couples). After ICSI, the comparable figure was 3.6% (2/56). The number of cases was too small to apply a statistical test to this difference. Total cleavage rates were quite similar: 86.7 +/- 28.0 and 90.1 +/- 21% of the zygotes developed into transferable embryos after IVF and ICSI respectively (not significant). Similarly, no difference in embryo quality was observed. Although injection and insemination of the oocytes were performed at the same time in the two groups, at 42 h post-insemination more embryos were at the four-cell stage after ICSI (P < 0.001) than after conventional IVF, where more embryos were still at the two-cell stage (P < 0.02). Embryo transfer was possible in all 56 couples, resulting in 16 positive serum human chorionic gonadotrophin tests (28.6% per embryo transfer), from which a clinical pregnancy resulted in 15 couples. The best embryos were selected for transfer independently of the insemination procedure, but preferably from the same origin. There appeared to be no difference in implantation potency of the embryos obtained with either technique after the non-randomized transfers. PMID- 10527974 TI - A quantitative evaluation of alpha1, alpha4, alphaV and beta3 endometrial integrins of fertile and unexplained infertile women during the menstrual cycle. A flow cytometric appraisal. AB - The expression of integrin molecules alpha1beta1, alpha4beta1 and alphaVbeta3 within endometrial tissue has been proposed as a marker of uterine receptivity during the implantation window. The present investigation examines by flow cytometric analysis the concentrations of alpha1, alpha4, alphaV and beta3 integrin subunits in endometrial stromal (ESC) and epithelial cells (EEC) in two groups of women throughout the menstrual cycle: normal fertile women (n = 27) and women with unexplained infertility (n = 26). Integrin concentrations in endometrial cells were calculated in relative fluorescence units against a negative cellular control. The assessment of integrin subunits detected the protein in ESC and EEC from the late proliferative to the late secretory phase. In both groups of women, the alpha1 was the highest integrin expressed in ESC and EEC throughout the menstrual cycle. All women exhibited low concentrations of alpha4-EEC at the time of the implantation window. Infertile women expressed lower concentrations of the alpha4-ESC during the proliferative and early secretory phase while lower concentrations of the alpha1-ESC were seen during the late secretory phase. Interestingly, the infertile women expressed lower concentrations of beta3-EEC in the early, mid-secretory and late secretory phases (P < 0.05). Infertile women also expressed lower concentrations of alpha1-EEC and alphaV-EEC during the late secretory phase (P < 0.05). It can be concluded that the quantitative determination of beta3-EEC by flow cytometry confirmed its potential feature as a marker of endometrial receptivity at the time of the implantation window. In addition, the defective expression of the alpha1-ESC found in the late secretory phase might be associated with the poor fertility outcome of women with unexplained infertility. PMID- 10527973 TI - A prospective randomized study of electro-acupuncture versus alfentanil as anaesthesia during oocyte aspiration in in-vitro fertilization. AB - The aim of the present study was to evaluate the anaesthetic effect during oocyte aspiration of a paracervical block (PCB) in combination with either electro acupuncture (EA) or intravenous alfentanil. In all, 150 women undergoing in-vitro fertilization (IVF) and embryo transfer were randomized to receive either EA plus PCB or alfentanil plus PCB. Visual analogue scales (VAS) were used to evaluate subjective experiences during oocyte aspiration, and IVF outcome parameters were recorded. No differences in pain directly related to oocyte aspiration, adequacy of anaesthesia during oocyte aspiration, abdominal pain, or degree of nausea were found between the two groups in the VAS ratings. Before oocyte aspiration, the level of stress was significantly higher in the EA group than in the alfentanil group (P < 0.05), and the EA group experienced discomfort for a significantly longer period during oocyte aspiration (P < 0. 01). Compared with the alfentanil group, the EA group had a significantly higher implantation rate (P < 0.05), pregnancy rate (P < 0.05), and take home baby rate (P < 0.05) per embryo transfer. In conclusion, EA has been shown to be as good an anaesthetic method as alfentanil during oocyte aspiration, and we suggest that EA may be a good alternative to conventional anaesthesia during oocyte aspiration. PMID- 10527976 TI - Successful in-vitro fertilization in a natural cycle after four previously failed attempts in stimulated cycles: case report. AB - A case is reported of successful in-vitro fertilization (IVF) and pregnancy in a natural cycle after four previously failed attempts with stimulated cycles. The patient began treatment at the age of 36 years and underwent four stimulated IVF cycles, each time with three embryos of good quality transferred. In one attempt, three cryopreserved embryos were transferred in a natural cycle. The patient failed to conceive. At the age of 38 years, the patient was entered into a natural cycle IVF programme. The patient conceived twice in each of her first two attempts but unfortunately aborted. In her third natural cycle of IVF, again with one oocyte obtained and one embryo transferred, the patient conceived and had a full term gestation. It is concluded that IVF in a natural cycle is a viable option for infertile women with blocked Fallopian tubes who have normal ovulatory menstrual cycles. PMID- 10527975 TI - The best donor. AB - Oocyte donation has become a common treatment modality for a large spectrum of infertility conditions. The purpose of this study was to assess the success rate of a shared egg donation programme, and to define the profile of a successful 'donor-recipient' couple in view of the limitations imposed by the shared programme. The results of all consecutive cycles of egg donation from 1st January 1995 to 31st December 1996 were analysed. A total of 383 donor cycles were matched with 946 recipient cycles; clinical pregnancy rates were 23. 5 and 16.7% respectively. With the exception of endometriosis, which significantly reduced the pregnancy rate in both groups, similar pregnancy rates were obtained in both groups for all the other infertility aetiologies of the donors. The donor's age had no impact on pregnancy rate of the recipient, but pregnancy rate was significantly decreased in donors >35 years. Recipients >50 years had significantly reduced pregnancy rates and those >45 years a significantly increased abortion rate. Recipients with severe male factor infertility, who had intracytoplasmic sperm injection treatment, showed pregnancy rates equivalent to those recipients who had regular in-vitro fertilization. We conclude that in a shared egg donation programme, the recipients' pregnancy rate and outcome are dependent only on the donors' infertility aetiologies and on recipients' ages. PMID- 10527977 TI - Norplant((R)) implants and progesterone vaginal rings do not affect maternal bone turnover and density during lactation and after weaning. AB - Bone density and turnover was assessed in a longitudinal study of healthy lactating women who initiated use of Norplant((R)) implants (NOR, n = 29), progesterone vaginal rings (PVR, n = 28) or Copper T 380A intrauterine devices (T Cu, n = 51, control group) around day 60 postpartum. Bone density, serum calcium, phosphorus, alkaline phosphatases, parathyroid hormone (PTH), follicle stimulating hormone (FSH), oestradiol and prolactin, and urinary hydroxyproline and creatinine were measured at postpartum months 1 (PM1), and 12 (PM12) and 6 or 12 months after weaning; at month 6 postpartum (PM6) serum and urine tests alone were performed. Baseline characteristics and lactation performance were similar between groups. Biochemical markers of bone turnover were higher at PM1, PM6 and PM12 than after weaning, with no differences between groups. Bone density in the lumbar spine (L2-L4) and femoral neck at PM1 and PM12 ( approximately 1.11 g/cm(2)) was similar in three groups. Lumbar spine values were found to be lower in lactating women than those present in non-lactating women, but increased after weaning to similar values. The two progestin-only contraceptives studied appear to have no deleterious effect upon bone density and metabolism in healthy lactating women. PMID- 10527978 TI - DNA flow cytometry of human semen. AB - The aim of this study was the evaluation of DNA flow cytometry for the analysis of male infertility. 171 ejaculates from 155 patients with fertility problems were analysed by flow cytometry and by conventional microscopical procedures. Using flow cytometry, it was possible to determine the relative proportions of the various cell populations: mature haploid and abnormal diploid mature spermatozoa, cellular fragments, immature germ cells (haploid round spermatids, diploid cells, S phase and 4C cells), and of leukocytes as indicators of infection. A linear association was observed between sperm concentration in semen as quantified by light microscopy and by flow cytometry, even with fewer than 20x10(6) spermatozoa/ml. Eight classes of histograms, each with differing fractions of spermatozoa and other particles, were obtained and correlated with the results of the spermiograms. During the 10 year follow-up, the two patient groups with a low sperm concentration or a high concentration of cellular debris exhibited significantly impaired fertility. The two patient groups with >/=5% diploid spermatozoa and with malcondensed sperm chromatin were also subfertile. No ovulatory disorders were revealed in the 155 female partners. DNA flow cytometry thus provides an additional dimension to semen analysis not easily gained by other methods and has the advantage of being rapidly performed and interpreted. We therefore recommend application of this technique in the diagnosis of male infertility. PMID- 10527979 TI - Antioxidant capacity of the epididymis. AB - The human epididymis provides an optimal environment for the storage and maturation of spermatozoa. However, the ability of the epididymis to protect spermatozoa from oxidative attack whilst stored at this site, through the local actions of antioxidants, has not thus far been well studied. This study assessed the contribution of the epididymis to seminal plasma antioxidant activity, by comparing the semen of normozoospermic and vasectomized men. Total seminal plasma antioxidant activity was measured, as were concentrations of urate, ascorbate and thiols, antioxidants that are abundant in human semen. Thiobarbituric acid reactive species (TBARS) were measured to indicate lipid peroxidation. Total antioxidant activity and thiol content were significantly lower (P < 0.05) in the plasma from vasectomized men compared with that of normozoospermic donors. Ascorbate and urate were found at similar concentrations in the plasma of both groups. The concentration of TBARS was significantly higher (P < 0.001) in the semen from vasectomized individuals compared with the normozoospermic group. The results indicate that the epididymis contributes to the antioxidant capacity of seminal plasma and possesses region-specific antioxidant activity, which may potentially protect spermatozoa from oxidative attack during storage at this site. PMID- 10527980 TI - Failure of fertilization after intracytoplasmic sperm injection in a patient with Kartagener's syndrome and totally immotile spermatozoa: case report. AB - Patients with Kartagener's syndrome (KS) are invariably infertile with totally immotile spermatozoa. Intracytoplasmic sperm injection (ICSI) is considered to be the treatment of choice for patients with immotile spermatozoa. We report the second KS case in the literature from whom immotile spermatozoa from the ejaculate failed to fertilize mature oocytes after ICSI. The role of micromanipulation in the treatment of KS patients is discussed. PMID- 10527981 TI - Human primordial, primary and secondary ovarian follicles in long-term culture: effect of partial isolation. AB - Ovarian cortical tissue, donated by 20 women aged 25-43 years during gynaecological laparoscopies or laparotomies, was first cultured for 7-9 days as tissue slices, 0.1-0.3 mm in thickness, in extracellular matrix, to initiate the growth of the primordial and primary follicles. It was then divided into two parts, one of which was cultured further as slices, and the other one used for enzymatic (collagenase at 1, 0.5 or 0.25 mg/ml; 17 patients) or mechanical (four patients) partial isolation of the follicles. The tissue slices and the partially isolated follicles were cultured for a further 1-3 weeks in the matrix. After approximately 2 weeks in culture, some oocytes began to extrude from the follicles, which were usually at the secondary stage. They were small, 20-80 micrometer in diameter, and had a thin or absent zona. Polar bodies and meiotic chromosomes could be seen in these naked oocytes. This premature extrusion probably resulted from sub-optimal culture conditions. It occurred sooner in follicles that had been partially isolated using collagenase. Histologically, larger numbers of oocytes were observed in non-isolated slice cultures than in the partially isolated cultures. Initiation of growth of the follicles occurred during the first 7-9 days in culture within slices. In non-isolated slices and following mechanical partial isolation there were significantly more secondary follicles after 11-18 days in culture than following isolation with collagenase. The proportion of atretic follicles increased during all cultures, and it was significantly higher after partial isolation. Because partial isolation did not improve the survival or development of the follicles the optimal method for human ovarian follicles could be to culture them non-isolated within small tissue slices. PMID- 10527982 TI - Dimeric inhibins and activin A in human follicular fluid and oocyte-cumulus culture medium. AB - The concentrations of inhibin A, inhibin B and activin A in follicular fluid and oocyte culture medium were analysed to investigate the production of these peptide hormones by ovarian granulosa cells and oocyte-cumulus complexes, as well as their potential as possible biochemical markers for oocyte quality and fertilizing capacity. Follicular fluids were collected from individual follicles during oocyte retrieval for in-vitro fertilization (IVF). Oocyte-cumulus culture media were collected after in-vitro insemination. The concentrations of dimeric inhibin A, inhibin B and activin A were measured using two-site enzyme-linked immunosorbent assays in the follicular fluid and matched oocyte culture medium. Hormone concentrations were compared with oocyte quality and fertilizing capacity. The concentration of inhibin A in follicular fluid increased while that of inhibin B decreased with increasing follicle size. Follicular fluid concentrations of inhibin A inhibin B and activin A were not significantly different in follicles with differing oocyte quality. Oocyte culture medium concentrations of activin A were significantly higher in morphologically good quality oocytes. There was no relationship between the concentrations of the three hormones and oocyte fertilizing capacity. This study confirms that follicular fluid concentrations of inhibin A may prove to be a marker of follicular growth and maturation. Higher concentrations of activin A produced by good quality oocyte-cumulus complexes suggest that activin A may play a role in oocyte maturation. PMID- 10527983 TI - Human DAZL1 encodes a candidate fertility factor in women that localizes to the prenatal and postnatal germ cells. AB - The human DAZ (Deleted in AZoospermia) gene family contains a cluster of DAZ genes on the Y chromosome and a single autosomal homologue, DAZL1 (DAZ-Like) that maps to chromosome 3p24. Although the role of the Y chromosome gene family in determining fertility and the expression of the gene family has been well explored in men, little is known of the role of the DAZL1 gene in determining the fertility of women. In mice, loss of function of the homologue of DAZL1 results in the loss of male and female germ cells. In mice, Dazl1 protein is localized to prenatal and postnatal follicles. Here we demonstrate using two antisera that recognize human DAZL1 that the protein is expressed embryonically in germ cells of girls and in mature oocytes. This pattern of expression suggests that the DAZL1 gene is a candidate fertility factor in women and that it would be appropriate to search for mutations in the DAZL1 gene in peripheral blood DNA from women with primary amenorrhoea or premature ovarian failure. PMID- 10527984 TI - Effects of a nitric oxide donor and nitric oxide synthase inhibitors on luteinizing hormone-induced ovulation in the ex-vivo perfused rat ovary. AB - The aim of this study was to investigate the role of nitric oxide (NO) in ovulation and ovarian steroidogenesis by the use of NO synthase (NOS) inhibitors and an NO donor administrated to the luteinizing hormone (LH)-stimulated ex-vivo perfused pre-ovulatory rat ovary. The ovaries were stimulated with LH (0.2 microgram/ml) alone or in combination with the phosphodiesterase inhibitor IBMX (200 micromol/l). The presence of both endothelial NOS (eNOS) and inducible NOS (iNOS) in the perfused rat ovary were detected by immunoblotting and a clear increase in amount of iNOS protein was seen after LH+IBMX stimulation. The addition of a non-selective NOS inhibitor, N(G)-monomethyl-L-arginine (L-NMMA; 300 micromol/l), to the perfusate significantly decreased ovulation numbers (median = 4. 0, range = 1-14) as compared with LH + IBMX stimulated control (12.0, 6-17). In contrast, an inhibitor with relative selectivity towards iNOS, aminoguanidine bicarbonate (AG, 300 micromol/l and 1 mmol/l), did not change the ovulation rate (11.5, 6-18 and 11.0, 7-15 respectively). In perfusions with only LH, a lower ovulation rate was seen but with similar effects (0.0, 0-8 for L NMMA; 7.5, 3-12 for control and 7.0, 1-15 for AG 300 micromol/l). The administration of an NO donor, spermine NONOate, resulted in similar ovulation numbers as in LH-stimulated controls. The NO inhibitors did not affect steroid concentrations in the perfusion media, while 100 micromol/l NONOate increased progesterone production. PMID- 10527985 TI - A macaque model for studying mechanisms controlling oocyte development and maturation in human and non-human primates. AB - A model to study mechanisms controlling nuclear and cytoplasmic maturation of primate oocytes is being developed in our laboratory. The high incidence of pregnancy failure in women following in-vitro fertilization (IVF) may be partly attributed to inadequate cytoplasmic maturation of oocytes. Advancement of knowledge of mechanisms controlling primate oocyte maturation would have important implications for treatment of human infertility, and would potentially increase numbers of viable non-human primate embryos for biomedical research. Use of a non-human primate model to study oocyte and embryo biology avoids legal, ethical and experimental limitations encountered in a clinical situation. Using this model, the meiotic and developmental capacity of oocytes from three sources have been compared: (i) in-vivo matured oocytes from monkeys stimulated with follicle-stimulating hormone (FSH) and human chorionic gonadotrophin, (ii) in vitro matured oocytes from monkeys primed with FSH, and (iii) in-vitro matured oocytes from non-stimulated monkeys. This work demonstrates that oocyte developmental competence is likely acquired both during follicle development, before meiotic resumption, and during meiotic progression, concurrent with nuclear maturation. Potential causes of developmental failure of in-vitro matured oocytes, implications for human infertility, and future strategies to study the regulation of primate oocyte maturation are discussed. PMID- 10527986 TI - Murine embryos as a direct target for some human autoantibodies in vitro. AB - The involvement of one or another autoantibody in reproductive failure have long been thought to be through post-implantation thrombosis and/or peri-implantation trophoblast dysfunction and/or maternal hormonal imbalance. It can be postulated that the embryo may be a direct target for some autoantibodies prior to implantation. Mouse embryos have been labelled and cultured with affinity purified immunoglobulin (IgG) and IgA from positive for antiphospholipid antibody sera, as well as IgG from positive for antinuclear antibody sera and positive for antithyroid antibody sera. Intact IgG and IgA from healthy individuals were used as controls. All embryos cultured with purified antiphospholipid IgG or IgA, and anti-nuclear IgG exhibited strong immunofluorescence. No difference in fluorescent intensity was observed whether antiphospholipid or anti-nuclear antibodies were used, but the pattern of antibody distribution seemed to be different. Antiphospholipid IgG was more dominant on the zona pellucida, while antiphospholipid IgA and antinuclear IgG had predominant distribution on the embryonic cells. None of the embryos cultured with antithyroid IgG or with control immunoglobulins showed strong immunofluorescence. Embryos cultured with purified antiphospholipid and antinuclear immunoglobulins experienced significant growth impairment or death compared to those cultured with antithyroid or control immunoglobulins. PMID- 10527987 TI - Shortened exposure of oocytes to spermatozoa improves in-vitro fertilization outcome: a prospective, randomized, controlled study. AB - A prospective, randomized study of 158 patients undergoing in-vitro fertilization (IVF) and embryo transfer was conducted to evaluate whether a shortened exposure of oocytes to spermatozoa enhances oocyte development, and subsequently influences the IVF outcome. A comparison was made between conventional treatment time and shorter exposure of retrieved oocytes to spermatozoa. Fertilization and cleavage rates, embryo quality, implantation and pregnancy rates in the study group (short exposure) versus controls (standard IVF procedure) were evaluated. Fertilization (56 versus 61%) and cleavage rates (96 versus 92%) were similar in the two groups respectively. However, embryo quality was significantly higher in the study group (P < 0.05). Moreover, the pregnancy and implantation rates were significantly increased (42.4 versus 26% per embryo transfer, and 16 versus 10% respectively; P < 0.05). Our results demonstrated that shorter exposure of oocytes to spermatozoa is superior to the standard time in IVF and may have a favourable effect on implantation rates by improving embryo quality. PMID- 10527988 TI - Intracytoplasmic sperm injection: position of the polar body affects pregnancy rate. AB - A prospective study on intracytoplasmic sperm injection (ICSI) was performed to evaluate the effect of the position of the polar body relative to the opening of the injection needle during sperm injection, and of the person who performs the injections on fertilization, cleavage, and pregnancy rates. This study included 173 couples undergoing 313 ICSI cycles from September 1995 to December 1997. All injections were performed by two persons. For each injected oocyte the person who performed the injection was recorded as well as the position of the polar body during injection (6 o'clock: animal pole towards the opening of the needle; 12 o'clock: animal pole away from the opening of the needle). Of 2630 oocytes retrieved, 2232 were injected. Significantly more oocytes developed two pronuclei after injection with the polar body at 6 o'clock versus 12 o'clock (P = 0.01; 51 versus 45% respectively) and after injection by person 1 versus person 2 (P = 0.02; 50 and 45% respectively). Higher pregnancy rate (P = 0.046) was found after transfer of embryos from oocytes injected with the polar body at 6 o'clock (36%) versus 12 o'clock (18%). This was the result of a significant interaction (P = 0.03) between the position of the polar body and the person performing the injections. Given the higher fertilization rate in the 6 o'clock group, it is recommended that oocytes be injected with the polar body at 6 o'clock. The higher pregnancy rate as a result of polar body position and the interaction between polar body position and the operator suggest variations in injection technique. PMID- 10527989 TI - The influence of in-vitro culture versus stimulated and untreated oviductal environment on mouse embryo development and implantation. AB - A prospective randomised study was performed to evaluate stimulated versus natural oviductal environment in comparison with in-vitro culture for the developmental capacity of mouse embryos. Therefore, embryos of superovulated F1 hybrid CBAxC57Bl females were collected at 17, 22, 41 and 46 h after human chorionic gonadotrophin treatment and randomly divided into five groups. They were either transferred immediately to untreated pseudopregnant females, cultured in vitro for 5, 24 or 29 h before transfer, or cultured in vitro for 96 h to blastocysts. The transfers resulted in an impaired implantation (P < 0.001) and a lower numbers of living fetuses (P < 0.001) when embryos had been exposed longer to the stimulated oviductal environment. Similar results were obtained after a longer period of in-vitro culture (P < 0.05). However when embryos were flushed earlier from the superovulated mice and cultured longer in-vitro until the transfer was performed, the implantation rate was improved (P < 0.01). Blastocyst development, however, was better (P < 0.001) when embryos were flushed later. In conclusion, the stimulated oviductal environment impairs the developmental capacity of embryos in comparison with untreated pseudopregnant females. In-vitro culture is also suboptimal but better than the stimulated oviductal environment. PMID- 10527990 TI - Fetal development after transfer is increased by replacing protein with the glycosaminoglycan hyaluronan for mouse embryo culture and transfer. AB - The effect of macromolecules on mouse embryo development and viability after culture in sequential media was investigated. It was found that high rates of viable blastocysts could be obtained in the absence of any macromolecule. Blastocyst cell numbers were increased when bovine serum albumin was present in the culture medium, although this benefit was not manifest after blastocyst transfer. Rather, the highest rates of implantation and fetal development after blastocyst transfer were observed when hyaluronan was the macromolecule in the culture media. Subsequent analysis revealed that the beneficial effects of hyaluronan were due to its presence in the transfer medium. As the highest cell numbers and hatching rates obtained in this study occurred when both serum albumin and hyaluronan were present in the same medium, it is proposed that embryo culture media should contain both serum albumin and hyaluronan, while the transfer medium need only contain hyaluronan. PMID- 10527991 TI - Prevention of twin pregnancy after in-vitro fertilization or intracytoplasmic sperm injection based on strict embryo criteria: a prospective randomized clinical trial. AB - A prospective randomized study comparing single embryo transfer with double embryo transfer after in-vitro fertilization or intracytoplasmic sperm injection (IVF/ICSI) was carried out. First, top quality embryo characteristics were delineated by retrospectively analysing embryos resulting in ongoing twins after double embryo transfer. A top quality embryo was characterized by the presence of 4 or 5 blastomeres at day 2 and at least 7 blastomeres on day 3 after insemination, the absence of multinucleated blastomeres and <20% cellular fragments on day 2 and day 3 after fertilization. Using these criteria, a prospective study was conducted in women <34 years of age, who started their first IVF/ICSI cycle. Of 194 eligible patients, 110 agreed to participate of whom 53 produced at least two top quality embryos and were prospectively randomized. In all, 26 single embryo transfers resulted in 17 conceptions, 14 clinical and 10 ongoing pregnancies [implantation rate (IR) = 42.3%; ongoing pregnancy rate (OPR) = 38.5%] with one monozygotic twin; 27 double embryo transfers resulted in 20 ongoing conceptions with six (30%) twins (IR = 48.1%; OPR = 74%). We conclude that by using single embryo transfer and strict embryo criteria, an OPR similar to that in normal fertile couples can be achieved after IVF/ICSI, while limiting the dizygotic twin pregnancy rate to its natural incidence of <1% of all ongoing pregnancies. PMID- 10527992 TI - Pronuclear orientation, polar body placement, and embryo quality after intracytoplasmic sperm injection and in-vitro fertilization: further evidence for polarity in human oocytes? AB - Three hypotheses were tested: (i) the distance between first and second polar bodies (PB) may relate to embryo morphology, (ii) that the orientation of pronuclei (PN) relative to PB may relate to embryo morphology, (iii) that the placement of a spermatozoon in a fixed plane relative to the first PB [intracytoplasmic sperm injection (ICSI)] may alter PN/PB orientation relative to in-vitro fertilization (IVF). A total of 251 two pronuclear (2PN) embryos (124 ICSI, 127 IVF) from 64 patients was studied. Angles were measured between the PN axis and the nearest PB (alpha), the furthest PB (beta), and between the two PB (gamma). On day 2, the morphological grades of embryos were recorded. gamma ranged from 0 to 150 degrees and was not significantly different for ICSI or IVF embryos of different grades; however, an unusual distribution of gamma suggested different populations of oocytes. The first hypothesis was rejected. alpha and beta ranged from 0 to 90 degrees : alpha did not relate significantly to embryo grade, but beta increased significantly with decreasing quality of ICSI embryos (P < 0.05) and the total group (P < 0.01), supporting hypothesis (ii). The difference in beta between ICSI and IVF embryos was not significant, so hypothesis (iii) was unproven. Significant differences between ICSI and IVF embryos in PN positions, irregular cleavage, and cleavage failure were noted. PMID- 10527993 TI - A prospective randomized study comparing intramuscular with intravaginal natural progesterone in programmed thaw cycles. AB - A simple programmed thaw cycle is described, during which the endometrium is prepared with oral oestradiol, followed by a natural progesterone source. This method involves minimal blood tests and uses inexpensive medications. Originally, an i.m. progesterone-in-oil preparation was used. Although highly successful in achieving high serum progesterone concentrations, the daily injections of the oily compound were found to be problematic from the patients' perspective. To examine the possibility of replacing the injectable progesterone with a vaginal preparation we conducted a prospective randomized study of 1 year's duration, during which time 170 and 184 thawing cycles were done with injectable and vaginal progesterone respectively. Although the progesterone blood concentrations obtained with the injectable preparation were more than twice those obtained with the vaginal progesterone, the clinical pregnancy rates (defined as percentage thawing cycles with gestational sac(s)/embryo transfer as demonstrated on ultrasound, 4 weeks after embryo transfer) were similar for both groups (15.9 and 16.8% respectively). Implantation and abortion rates were also comparable. These results agree with previous reports of higher uterine progesterone concentrations after the vaginal application. We conclude that the combination of oral oestradiol and vaginal progesterone is an effective, simple and inexpensive approach for programmed thaw cycles. PMID- 10527994 TI - Saline infusion contrast intrauterine sonographic assessment of the endometrium with high-frequency, real-time miniature transducer in normal menstrual cycle: a preliminary report. AB - Normal endometrial texture was visualized using saline infusion contrast intrauterine sonography with a specially developed 20 MHz flexible catheter-based high-resolution, real-time miniature (2.4 mm outer diameter) ultrasound transducer in primary infertile women (n = 15) with a normal menstrual cycle. All the women had <2 years infertility duration and were studied in proliferative, and early or mid-secretory phases. Before intrauterine sonography, transvaginal sonographic assessment of the endometrium was conducted. The overall image clarity was subjectively compared between intrauterine and transvaginal sonography. Most endometrial textures in both proliferative and secretory phases were viewed more easily with intrauterine rather than transvaginal sonography, and this was especially true with an intrauterine saline infusion technique. Moreover, it was possible to obtain finer image quality of very small endometrial interfacial and internal textures with intrauterine sonography. However, the depth of penetration of the ultrasound beam is only approximately 2 cm, therefore examination of larger pathological endometrial lesions is markedly limited because of the shallow scanning range of the high-frequency transducer. Intrauterine sonography may be a valuable tool in imaging endometrial texture in normal menstrual cycle, and possibly in infertility practice, complementing and not replacing transvaginal sonography. PMID- 10527995 TI - Reproducibility of transvaginal three-dimensional endometrial volume measurements during ovarian stimulation. AB - Before estimating the clinical role of a method, the reproducibility has to be evaluated precisely. This study aimed to document the reproducibility of the endometrial volume measurement by three-dimensional (3D)-ultrasound. The volume measurements were done on 57 consecutive in-vitro fertilization (IVF) patients with either the full planar (contour) or the three distance method. A paired t test provided no statistically significant difference between the two methods. Linear regression analysis, using the full planar method as independent and the other as dependent variables, yielded the following results: intercept = 0.348, not statistically different from 0; slope = 0.962, statistically different from 1 (P < 0.01). Interobserver reliability for the three distance method was 0.6667 and for the full planar method was 0.9565. Intra-observer reliability for the three distance method was 0.8426 and for the full planar method 0.9394. The correspondence between and within observers seemed to be good. Both methods are reliable, but the full planar method seems to provide slightly better reproducibility in regard to endometrium volume measurement. PMID- 10527996 TI - Endometrial histomorphometry of trimegestone-based sequential hormone replacement therapy: a weighted comparison with the endometrium of the natural cycle. AB - Histomorphometric changes in the endometrium were evaluated under the effect of a trimegestone-based sequential hormone replacement therapy (HRT) regimen, and the findings were compared to those in endometrium of the natural cycle. Endometrial samples were taken from postmenopausal women who completed a randomized, double blind, dose-ranging study of oral trimegestone (0.05, 0.1, 0.25 and 0.5 mg per day) from day 15 to day 28 with continuous micronized oestradiol 2 mg daily for six treatment cycles. The HRT-treated endometrium, irrespective of the dose, had a smaller mean total glandular area, smaller average glandular diameter, smaller mean total vascular space area and diameter than the luteal phase. Stromal cellularity was similar in the four dose groups. There were reduced glandular secretions in the endometrium from the high dose group. The relative weighting of these histological parameters was evaluated by linear discriminant analysis. The weighted values were dose independent, and may therefore represent either a specific effect of trimegestone, number of days administered, or both. We have constructed an equation to assign a value for a histological parameter which determines the position on linear discriminant functions. These assigned values can be used in other sequential HRT regimens to determine the relative influence of a given progestogen on endometrial morphology in relation to different phases of the natural cycle. PMID- 10527997 TI - Obstetric outcome of pregnancies after the transfer of cryopreserved and fresh embryos obtained by conventional in-vitro fertilization and intracytoplasmic sperm injection. AB - This study reports the obstetric outcome of pregnancies obtained after the transfer of cryopreserved or fresh embryos where the initial procedure was standard in-vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI). Pregnancies obtained after frozen IVF (n = 245) or frozen ICSI (n = 177) were compared with a control group of pregnancies after fresh embryo transfer in standard IVF (n = 245) and ICSI (n = 177) cycles were selected as controls. The controls were matched according to maternal age, parity and date of embryo transfer. In the standard IVF group, the biochemical pregnancy rates in the cryopreserved and fresh groups were 18.8 and 9.8% respectively (P < 0.01). In the ICSI group, the biochemical pregnancy rates in the cryopreserved and fresh groups were 16.4 and 6.8% respectively (P < 0.01). The miscarriage rates were comparable between the cryopreserved and fresh groups. However, in the frozen ICSI group the miscarriage rate (26.0%) was significantly higher than in the frozen conventional IVF group (13.1%) (P = 0.001). The frequencies of preterm deliveries, infants with very low birthweight and intrauterine deaths were similar in the groups. The low birthweight rates in the frozen IVF (16.1%) and ICSI (12.1%) groups were significantly lower than those in the fresh IVF (32.2%) and ICSI (32.7%) groups (P < 0.001). The major malformation rates in the frozen IVF (2.4%) and ICSI (2.9%) groups were not different from the major malformation rates in the fresh IVF (4.5%) and ICSI (2.4%) groups. In conclusion, the cryopreservation process had no negative impact on the outcome of pregnancies over 20 weeks of gestation. Long-term follow-up studies are needed in order to prove the safety of the freezing-thawing process. PMID- 10527998 TI - Hepatocyte growth factor concentration in the early second-trimester amniotic fluid does not predict fetal growth at birth. AB - The purpose of this study was to evaluate whether hepatocyte growth factor (HGF) concentrations in the early second-trimester amniotic fluid predict fetal growth at birth. HGF and insulin-like growth factor-I (IGF-I) concentrations in the early second-trimester amniotic fluid were measured in 12 pregnancies with small for gestational age (SGA) infants, 84 pregnancies with appropriate for gestational age (AGA) infants, and eight pregnancies with large for gestational age (LGA) infants. HGF concentrations were measured from the early second trimester amniotic fluid samples using an enzyme-linked immunosorbent assay. IGF I concentrations were measured from the early second-trimester amniotic fluid samples using an immunoradiometric assay. Maternal age in AGA group (34.2 +/- 5.5 years) was significantly lower than in SGA (37.9 +/- 3.0 years) and LGA (37.6 +/- 3.3 years) groups (P < 0.05). There were no significant differences for parity or gestational age at amniocentesis among the groups. There were significant differences for birth age, birth weight, neonatal height, and placental weight among the groups (P < 0.05). HGF concentrations in SGA, AGA and LGA groups were 16.9 +/- 6.6, 16.7 +/- 9.0 and 20.2 +/- 14.8 ng/ml respectively (not significant). There was no correlation between amniotic fluid HGF concentrations and birth weight, height or placental weight. There were also no significant differences for amniotic fluid IGF-I concentrations among the three groups. These results suggest that differences in HGF concentrations in the early second trimester amniotic fluid do not predict fetal growth at birth. Further study is needed to clarify the role of high HGF concentrations in early second-trimester amniotic fluid during pregnancy. PMID- 10527999 TI - Genetic sonography as the preferred option of prenatal diagnosis in patients with pregnancies following intracytoplasmic sperm injection. AB - The option of prenatal diagnosis with nuchal translucency measurement at 10-14 weeks of gestation and second trimester targeted ultrasound including fetal echocardiography (genetic sonography) is reported in patients after intracytoplasmic sperm injection (ICSI). From January 1995 to December 1998, 153 consecutive patients, with a mean age of 32.3 years (+/-4.1) and 29. 6% >/= 35 years, who had become pregnant after ICSI, were studied. They attended our unit for first and second trimester sonography. Of these, 67.8% of primigravid and 80.9% of nulliparous women were included. Multiple pregnancy rate was 19.7%; 189 fetuses were screened in total. Due to the introduction of genetic sonography in 1995, the rate of invasive prenatal diagnosis decreased from 74% in 1995, to 48, 36 and 19% in 1996, 1997, and 1998 respectively. Two inherited numerical and structural chromosomal anomalies in clinically healthy children at birth (1.0%) and four major malformations in all liveborn children and late abortions (2.1%) were recorded. The results demonstrate that especially in women of advanced reproductive age with a long history of infertility, detailed genetic sonography may be a reasonable and highly accepted alternative to avoid even the relatively low risks associated with invasive screening procedures. PMID- 10528000 TI - Comparison of alterations in fetal regional arterial vascular resistance in appropriate-for-gestational-age singleton, twin and triplet pregnancies. AB - The objective of this longitudinal study was to evaluate alterations in fetal vascular resistance of fetal peripheral arteries with advancing gestation in singleton appropriate-for-gestational-age (S-AGA), twin appropriate-for gestational-age (Tw-AGA) and triplet appropriate-for-gestational-age (Tri-AGA) infants. Colour Doppler flow imaging and pulsed Doppler ultrasonographic examinations were performed on 35 S-AGA, 52 Tw-AGA and 12 Tri-AGA fetuses. The pulsatility index for middle cerebral artery (MCAPI), umbilical artery (UAPI), descending aorta (DAPI), splenic artery (SAPI), renal artery (RAPI) and femoral artery (FAPI) was measured as vascular resistance every 2 weeks after 15 weeks of menstrual age until delivery. Optimal models and normal ranges for pulsatility index for each artery in each group were generated. The alterations in various fetal regional arterial pulsatility indices with advancing gestational age showed no significant differences in S-AGA, Tw-AGA and Tri-AGA infants, respectively. These results suggest that there is no significant difference for regional arterial vascular resistance in AGA fetuses among singleton, twin, and triplet pregnancies, whereas there was a slight difference in fetal growth pattern among singleton, twin, and triplet pregnancies described in our previous investigation. PMID- 10528001 TI - Evaluation of ectopic pregnancy by magnetic resonance imaging. AB - Patients (n = 37) suspected of ectopic pregnancy were prospectively evaluated with magnetic resonance (MR) imaging to assess the capability of MR imaging in the diagnosis of ectopic pregnancy. Five levels of confidence were defined: diagnostic, suspicious, equivocal, questionable, and negative. Tubal wall enhancement and presence of tubal haematoma or gestational sac-like structure were considered diagnostic findings. There were 21 diagnostic, two suspicious, eight equivocal, and six negative findings. MR findings were compared with the surgical findings in 18 patients. Surgical confirmation was obtained in 12 diagnostic, two suspicious, and four equivocal studies. Using the MR diagnostic criteria for tubal pregnancy, MR had 12 true positive, three true negative, three false negative, and no false positive results for the diagnosis of tubal pregnancy. Retrospective analysis of the signal intensity of haematoma and ascites was performed for these 18 surgically confirmed cases. The predominant signal intensity of tubal haematoma was an intermediate signal on T1-weighted image (WI) and a low signal on T2WI. Ascites showed signal intensity higher than that of urine on T1WI in 100% of 13 cases. In conclusion, MR imaging with use of intravenous contrast material allows a specific diagnosis of tubal pregnancy, recognizing tubal wall enhancement and fresh tubal haematoma. PMID- 10528002 TI - Cumulative delivery rates after intracytoplasmic sperm injection: 5 year follow up of 498 patients. AB - The use of life-table analysis for infertility data has the advantages of clarity and ease of application. Success rates per cycle have been reported, but not cumulative delivery rates for intracytoplasmic sperm injection (ICSI). We selected retrospectively 498 Belgian patients <37 years old, who had their first ICSI cycle between July 1992 and December 1993. Follow-up was till the end of October 1997. Outcome measure was any delivery >25 weeks. These couples underwent 963 ICSI cycles using fresh ejaculated spermatozoa. The indications for ICSI were long-standing severe male infertility or fertilization failure after conventional in-vitro fertilization (IVF). Cumulative delivery rates were calculated by life table analysis and compared according to age groups and sperm quality. There were 298 deliveries within a mean rate per cycle of 31%. The average number of cycles required for a delivery was 3.15 (CI 2.88; 3.43). Twenty-three (4.6%) spontaneous pregnancies occurred after the patients had finished therapy. There was no significant difference between the sperm quality groups but delivery rates decreased significantly with increasing female age. The real delivery rate after six cycles was 60%, while the expected cumulative delivery rate was 86%. This life-table analysis may provide a means by which to counsel couples on the likelihood of a delivery following ICSI. PMID- 10528003 TI - Long-term follow-up of children born after inadvertent administration of a gonadotrophin-releasing hormone agonist in early pregnancy. AB - Our objective was to evaluate long-term outcome of children born after inadvertent administration of a gonadotrophin-releasing hormone agonist (GnRHa) in early pregnancy, compared to a control group of children born to matched women undergoing in-vitro fertilization and children born after spontaneous pregnancies. Six children from six pregnancies, exposed to a long-acting gonadotrophin agonist, comprised the study group and 20 children were included in the control groups. Pre-, peri- and postnatal data were collected and the children were followed and examined at a mean age of 7.8 +/- 2.0 years. All children underwent physical and neurological examination, and psychological tests. In the study group, one child was born with a major congenital malformation (cleft palate), and four children subsequently demonstrated neurodevelopmental abnormalities, including epileptic disorder (n = 1), attention deficit hyperactivity disorder (n = 3), motor difficulties (n = 3) and speech difficulties (n = 1). In the control groups, one child had attention deficit hyperactivity disorder. This observation of neurodevelopmental abnormalities in four of six children in the study group justifies the need for long-term follow up of more children previously exposed to gonadotrophin-releasing hormone agonist. PMID- 10528004 TI - Non-functioning pituitary tumour after long-term treatment with gonadotrophin releasing hormone agonists in a patient with vaginal agenesis who underwent neovaginoplasty and cauterization of endometriosis under laparoscopy. AB - Vaginal agenesis combined with a functional uterus is a rare condition in which treatment modalities that preserve reproductive function are controversial. A 21 year old female presented with congenital vaginal agenesis combined with cervical atresia. She was treated with gonadotrophin-releasing hormone (GnRH) agonists for a total period of over 5 years when a non-functioning pituitary tumour was detected by brain magnetic resonance imaging (MRI). A laparoscopically assisted reconstruction of a neovagina and neoendocervical canal was performed utilizing lyophilized porcine dermal skin to line the neovagina. Endometriosis of the pelvis was revealed and adhesiolysis and cauterization were also carried out under laparoscopy. The GnRH agonist was discontinued and the patient resumed cyclic menses with no abdominal pain. The pituitary tumour decreased in size 6 months after the cessation of GnRH agonists. We raise the question as to whether pituitary MRI should be performed for patients who need long-term administration of GnRH agonists. PMID- 10528005 TI - Preliminary experience of the use of a gonadotrophin-releasing hormone antagonist in ovulation induction/in-vitro fertilization prior to cancer treatment. AB - Therapeutic regimens for the treatment of malignant disease may compromise future fertility. One approach to circumvent this is the cryopreservation of embryos created before treatment for the malignancy. Conventional regimens using gonadotrophin-releasing hormone (GnRH) agonists are time consuming, requiring pituitary down-regulation before gonadotrophin administration, thus the duration of treatment is approximately 20-30 days. GnRH antagonists, however, do not cause an initial stimulation of gonadotrophin secretion and can thus be administered during the later stages of follicular maturation to prevent premature luteinization and ovulation. The duration of ovulation induction/in-vitro fertilization (IVF) treatment is thus reduced. In this study, case histories are reported of six women with newly diagnosed malignancies who requested ovulation induction/IVF prior to chemotherapy or surgery in which we have used the GnRH antagonist Cetrorelix. Gonadotrophin administration was started in the early follicular phase, and Cetrorelix (0.25 mg s.c. daily) was added from day 6 of treatment. Subsequent to human chorionic gonadotrophin (HCG) administration oocytes were recovered and successful fertilization and embryo cryopreservation was achieved in all cases. The median duration of treatment was 12 days (range 8 13, including induction of luteolysis in two patients). These results illustrate the potential use and advantages of a GnRH antagonist in ovulation induction/IVF when the need for immediate initiation of treatment and its duration are critical factors. PMID- 10528006 TI - The male partner involved in legal abortion. AB - This study comprises 75 men who have been involved in legal abortion. The men answered a questionnaire concerning living conditions and attitudes about pregnancy and abortion. Most men were found to be in stable relationships with good finances. More than half clearly stated that they wanted the woman to have an abortion while 20 stressed that they submitted themselves to their partner's decision. Only one man wanted the woman to complete the pregnancy. Apart from wanting children within functioning family units, the motivation for abortion revealed that the desire to have children depended on the ability to provide qualitatively good parenting. More than half the men had discussed with their partner what to do in event of pregnancy and half had decided to have an abortion if a pregnancy occurred. More than half expressed ambivalent feelings about the coming abortion, using words such as anxiety, responsibility, guilt, relief and grief. In spite of these contradictory feelings, prevailing expectations concerning lifestyle make abortion an acceptable form of birth control. A deeper understanding of the complexity of legal abortion makes it necessary to accept the role of paradox, which the ambivalence reflects. Obviously, men must constitute a target group in efforts to prevent abortions. PMID- 10528007 TI - Cervical pregnancy-a conservative stepwise approach. PMID- 10528008 TI - Which factors are important for successful embryo transfer after in-vitro fertilization? PMID- 10528009 TI - Which factors are important for successful embryo transfer after in-vitro fertilization? PMID- 10528010 TI - MEMORANDUM FOR: science writers and editors on the journal press list : A viral gene may Be involved in the rapid growth of Kaposi's sarcoma lesions PMID- 10528011 TI - MEMORANDUM FOR: science writers and editors on the journal press list : high blood levels of vitamin E associated with lower incidence of lung cancer PMID- 10528012 TI - Human herpesvirus 8 latent-state gene expression and apoptosis in Kaposi's sarcoma lesions. PMID- 10528013 TI - Scientific interest in Newcastle disease virus is reviving. PMID- 10528014 TI - Viruses and cancer. PMID- 10528015 TI - ECCO 10: Quality-of-life issues bumped into the spotlight. PMID- 10528016 TI - In coping with cancer, gender matters. PMID- 10528017 TI - Stat bite: Incidence of uterine cancer in U.S. women age 50 and older. PMID- 10528019 TI - NCI addresses "inconsistencies" in cancer care quality. PMID- 10528018 TI - Leeches latch on after reconstructive surgery. PMID- 10528020 TI - NCI funds cancer trials support unit PMID- 10528021 TI - Prostate cancer practice patterns and quality of life: the Prostate Cancer Outcomes Study. PMID- 10528022 TI - Expression of K13/v-FLIP gene of human herpesvirus 8 and apoptosis in Kaposi's sarcoma spindle cells. AB - BACKGROUND: Human herpesvirus 8 (HHV8) infection is associated with all forms of Kaposi's sarcoma (KS). The HHV8 genome locus ORFK13-72-73 (ORF = open reading frame) encodes proteins that may be important in HHV8-mediated pathogenesis, i.e., the latency-associated nuclear antigen (encoded by ORF73), viral-cyc-D (v cyc-D), a viral homologue of cellular cyclin D (encoded by ORF72), and viral-FLIP (v-FLIP), a homologue of the cellular FLICE (Fas-associated death domain-like interleukin 1 beta-converting enzyme) inhibitory protein (encoded by ORFK13; is an inhibitor of apoptosis [programmed cell death]). Through differential splicing events, this locus expresses individual RNA transcripts that encode all three proteins (tricistronic transcripts) or just two of them (v-FLIP and v-cyc-D; bicistronic transcripts). We examined expression of these transcripts in KS tissues. METHODS: We collected tissues from patients with KS of different stages. By use of an optimized in situ hybridization procedure, we examined different ORFK13-72-73 locus transcripts in HHV8-infected cells in skin lesions and in one adjacent lymph node. Apoptosis in KS lesions was determined by use of an in situ assay. RESULTS AND CONCLUSIONS: Our results indicate the following: 1) Transcripts from the ORFK13-72-73 locus appear to be spliced differentially in latently infected KS cells in skin lesions and in HHV8-infected cells in lymph nodes; specifically, ORFK13-ORF72 bicistronic transcripts were expressed abundantly in KS cells, whereas ORFK13-ORF72-ORF73 tricistronic transcripts were detected only in lymph node cells. 2) Sequences encoding the antiapoptotic protein v-FLIP are expressed at very low levels in early KS lesions, but expression increases dramatically in late-stage lesions. 3) The increase in expression of v-FLIP-encoding transcripts is associated with a reduction in apoptosis in KS lesions. IMPLICATIONS: These data suggest that functional v-FLIP is produced in vivo and that antiapoptotic mechanisms may be involved in the rapid growth of KS lesions, where only a few cells undergoing mitosis are generally observed. PMID- 10528023 TI - Prostate-specific antigen testing of older men. AB - BACKGROUND: Elevated serum prostate-specific antigen (PSA) levels are predictive of a future diagnosis of prostate cancer. To test the hypothesis that older men with low PSA levels may require less intensive PSA testing because of a reduced prostate cancer detection rate, we evaluated the association between age, baseline PSA level, and prostate cancer detection. METHODS: We conducted a prospective cohort study among participants in a study of aging who had serial PSA measurements taken from age 60 or 65 years until they either were diagnosed with prostate cancer (cancer case subjects) or reached the age of 75 years (subjects without prostate cancer). The time of cancer detection among cancer case subjects was defined as the measurement date on which a PSA level above 4.0 ng/mL was detected (i.e., PSA conversion). Cancer case subjects and subjects without prostate cancer were analyzed according to baseline PSA level and age. RESULTS: All cancer case subjects in the 60-year-old cohort had baseline PSA levels above 0.5 ng/mL, and 14 of 15 cancer cases that would have been detected by a PSA conversion among the 65-year-old cohort were associated with baseline PSA levels of 1.1 ng/mL or more. If PSA testing were discontinued in men aged 65 years with PSA levels of 0.5 ng/mL or less, 100% (95% confidence interval [CI] = 78%-100%) of the cancers would still be detected by age 75 years; if PSA testing were discontinued in men aged 65 years who had PSA levels of 1.0 ng/mL or less, 94% (95% CI = 70%-100%) of the cancers would still be detected by age 75 years. CONCLUSIONS: These data suggest that a decrease in the intensity of screening among older men with low PSA values may not lead to an increase in undetected prostate cancer. PMID- 10528024 TI - Serum alpha-tocopherol and subsequent risk of lung cancer among male smokers. AB - BACKGROUND: Higher blood levels of alpha-tocopherol, the predominant form of vitamin E, have been associated in some studies with a reduced risk of lung cancer, but other studies have yielded conflicting results. To clarify this association, we examined the relationship between prospectively collected serum alpha-tocopherol and lung cancer in the Alpha-Tocopherol, Beta-Carotene Cancer Prevention (ATBC) Study cohort. METHODS: The ATBC Study was a randomized, clinical trial of 29 133 white male smokers from Finland who were 50-69 years old and who had received alpha-tocopherol (50 mg), beta-carotene (20 mg), both, or neither daily for 5-8 years. Data regarding medical histories, smoking, and dietary factors were obtained at study entry, as was a serum specimen for baseline alpha-tocopherol determination. alpha-Tocopherol measurements were available for 29 102 of the men, among whom 1144 incident cases of lung cancer were diagnosed during a median observation period of 7.7 years. The association between alpha-tocopherol and lung cancer was evaluated with the use of multivariate proportional hazards regression. RESULTS: A 19% reduction in lung cancer incidence was observed in the highest versus lowest quintile of serum alpha-tocopherol (relative risk = 0.81; 95% confidence interval = 0. 67-0.97). There was a stronger inverse association among younger men (<60 years), among men with less cumulative tobacco exposure (<40 years of smoking), and possibly among men receiving alpha-tocopherol supplementation. CONCLUSIONS: In the ATBC Study cohort, higher serum alpha-tocopherol status is associated with lower lung cancer risk; this relationship appears stronger among younger persons and among those with less cumulative smoke exposure. These findings suggest that high levels of alpha-tocopherol, if present during the early critical stages of tumorigenesis, may inhibit lung cancer development. PMID- 10528025 TI - Tumor-specific gene delivery using recombinant vaccinia virus in a rabbit model of liver metastases. AB - BACKGROUND: Several approaches to gene therapy for cancer have yielded promising results in rodent models. The translation of these results to the clinical realm has been delayed by the lack of tumor models in large animals. We investigated the pattern of transgene (i. e., foreign or introduced gene) expression and virus vector elimination after systemic gene delivery using a thymidine kinase-negative vaccinia virus in a rabbit model of disseminated liver metastases. METHODS: VX-2 rabbit carcinoma cells were maintained by serial transplantation in the thigh muscles of New Zealand White rabbits, and disseminated liver metastases were established by direct injection of tumor cells into the portal vein of the animals. Different doses of a recombinant thymidine kinase-negative vaccinia virus vector encoding the firefly luciferase reporter gene (i.e., transgene) were injected into tumor-bearing rabbits. Transgene activity in tumors and other organs was measured at multiple time points thereafter. The pattern of development of antibodies against the vaccinia virus vector was also examined. Two-tailed Student's paired t test was used for comparisons of transgene activity. RESULTS: Transgene expression was increased in tumors by at least 16 fold in comparison with expression in other tissues by day 4 after vector injection (all P<. 001) and was maintained for approximately 1 week, providing evidence of tumor-specific gene delivery in this model. Rapid elimination of the circulating vector by the host immune system was observed. Anti-vector antibodies were detectable in serum as early as day 6 and were maintained for more than 3 months. CONCLUSIONS: Tumor-specific gene delivery is possible after systemic injection of a thymidine kinase-negative vaccinia virus vector in a model of rabbit liver metastases. Although the period of transgene expression appears limited because of a rapid immune response, the therapeutic window might be sufficient for an enzyme/prodrug gene therapy approach in clinical application. PMID- 10528026 TI - Dietary fat and protein in relation to risk of non-Hodgkin's lymphoma among women. AB - BACKGROUND: Non-Hodgkin's lymphoma occurs more frequently in individuals with suppressed immune status, and some types of dietary fat and protein have been associated with decreased immune responses. In this study, we examined the intake of specific types of dietary fat and protein in relation to the risk of non Hodgkin's lymphoma. METHODS: We documented 199 incident cases of non-Hodgkin's lymphoma in a cohort of 88 410 women, who were enrolled in the Nurses' Health Study and were aged 34-60 years in 1980, during 14 years of follow-up. Relative risks of the disease and 95% confidence intervals (95% CIs) were calculated. All P values are two-sided and were considered to be statistically significant for P<.05. RESULTS: Intake of saturated fat was associated with an increase in risk that was not statistically significant; the multivariate relative risk for the highest versus the lowest quintiles of intake was 1.4 (95% CI = 0.7-3.0; P for trend =.42). Intake of beef, pork, or lamb as a main dish was associated with a statistically significantly increased risk of non-Hodgkin's lymphoma; the multivariate relative risk for consumption of these meats at least once per day as compared with less than once per week was 2.2 (95% CI = 1.1-4.4; P for trend =.002). Higher intake of trans unsaturated fat was also statistically significantly associated with an increased risk of the disease; the multivariate relative risk for the highest versus the lowest quintiles was 2.4 (95% CI = 1.3 4.6; P for trend =.01). Higher intake of red meat cooked by broiling or barbecuing-but not by roasting, pan-frying, or boiling or stewing-was associated with an increase in risk that was not statistically significant. CONCLUSIONS: Greater dietary intake of certain meats and fats was associated with a higher risk of non-Hodgkin's lymphoma. These relationships and their potential mechanisms deserve further examination. PMID- 10528027 TI - Hormone therapy failure in human prostate cancer: analysis by complementary DNA and tissue microarrays. AB - BACKGROUND: The molecular mechanisms underlying the progression of prostate cancer during hormonal therapy have remained poorly understood. In this study, we developed a new strategy for the identification of differentially expressed genes in hormone-refractory human prostate cancer by use of a combination of complementary DNA (cDNA) and tissue microarray technologies. METHODS: Differences in gene expression between hormone-refractory CWR22R prostate cancer xenografts (human prostate cancer transplanted into nude mice) and a xenograft of the parental, hormone-sensitive CWR22 strain were analyzed by use of cDNA microarray technology. To validate the data from cDNA microarrays on clinical prostate cancer specimens, a tissue microarray of specimens from 26 prostates with benign prostatic hyperplasia, 208 primary prostate cancers, and 30 hormone-refractory local recurrences was constructed and used for immunohistochemical detection of protein expression. RESULTS: Among 5184 genes surveyed with cDNA microarray technology, expression of 37 (0.7%) was increased more than twofold in the hormone-refractory CWR22R xenografts compared with the CWR22 xenograft; expression of 135 (2.6%) genes was reduced by more than 50%. The genes encoding insulin-like growth factor-binding protein 2 (IGFBP2) and 27-kd heat-shock protein (HSP27) were among the most consistently overexpressed genes in the CWR22R tumors. Immunohistochemical analysis of tissue microarrays demonstrated high expression of IGFBP2 protein in 100% of the hormone-refractory clinical tumors, in 36% of the primary tumors, and in 0% of the benign prostatic specimens (two-sided P =.0001). Overexpression of HSP27 protein was demonstrated in 31% of the hormone-refractory tumors, in 5% of the primary tumors, and in 0% of the benign prostatic specimens (two-sided P =.0001). CONCLUSIONS: The combination of cDNA and tissue microarray technologies enables rapid identification of genes associated with progression of prostate cancer to the hormone-refractory state and may facilitate analysis of the role of the encoded gene products in the pathogenesis of human prostate cancer. PMID- 10528028 TI - Breast-feeding and risk of childhood acute leukemia. AB - BACKGROUND: Breast-feeding is well known to have a protective effect against infection in infants. Although the long-term effects of breast-feeding on childhood cancer have not been studied extensively, a protective effect against childhood Hodgkin's disease and lymphoma has been suggested previously from small investigations. In this study, we tested the hypothesis that breast-feeding decreases the risk of childhood acute leukemia. METHODS: A total of 1744 children with acute lymphoblastic leukemia (ALL) and 1879 matched control subjects, aged 1 14 years, and 456 children with acute myeloid leukemia (AML) and 539 matched control subjects, aged 1-17 years, were included in the analysis. Information regarding breast-feeding was obtained through telephone interviews with mothers. All leukemias combined, histologic type of leukemia (ALL versus AML), immunophenotype of ALL (early pre-B cell, pre-B cell, or T cell), and morphology of AML were assessed separately in the data analysis. RESULTS: Ever having breast fed was found to be associated with a 21% reduction in risk of childhood acute leukemias (odds ratio [OR] for all types combined = 0.79; 95% confidence interval [CI] = 0.70-0.91). A reduction in risk was seen separately for AML (OR = 0.77; 95% CI = 0.57-1.03) and ALL (OR = 0.80; 95% CI = 0.69-0.93). The inverse associations were stronger with longer duration of breast-feeding for total ALL and AML; for M0, M1, and M2 morphologic subtypes of AML; and for early pre-B-cell ALL. CONCLUSION: In this study, breast-feeding was associated with a reduced risk of childhood acute leukemia. If confirmed in additional epidemiologic studies, our findings suggest that future epidemiologic and experimental efforts should be directed at investigating the anti-infective and/or immune-stimulatory or immune modulating effects of breast-feeding on leukemogenesis in children. PMID- 10528029 TI - Risk of transfusion-associated transmission of human herpesvirus 8. PMID- 10528030 TI - Preliminary sibpair linkage analysis of percent mammographic density. PMID- 10528032 TI - Re: Measure once or twice--does it really matter? PMID- 10528031 TI - Membranoproliferative glomerulonephritis following gemcitabine and vinorelbine chemotherapy for peritoneal mesothelioma. PMID- 10528033 TI - RESPONSE: re: measure once or twice-does It really matter? PMID- 10528034 TI - A 10-year prospective study of primary hyperparathyroidism with or without parathyroid surgery. AB - BACKGROUND AND METHODS: In the United States, most patients with primary hyperparathyroidism have few or no symptoms. The need for parathyroidectomy to treat all patients with this disorder has therefore been questioned. We studied the clinical course and development of complications for periods of up to 10 years in 121 patients with primary hyperparathyroidism, 101 (83 percent) of whom were asymptomatic. There were 30 men and 91 women (age range, 20 to 79 years). During the study, 61 patients (50 percent) underwent parathyroidectomy, and 60 patients were followed without surgery. RESULTS: Parathyroidectomy in patients with or without symptoms led to normalization of serum calcium concentrations and a mean (+/-SE) increase in lumbar-spine bone mineral density of 8+/-2 percent after 1 year (P=0.005) and 12+/-3 percent after 10 years (P=0.03). Bone mineral density of the femoral neck increased 6+/-1 percent after 1 year (P=0.002) and 14+/-4 percent after 10 years (P=0.002). Bone mineral density of the radius did not change significantly. The 52 asymptomatic patients who did not undergo surgery had no change in serum calcium concentration, urinary calcium excretion, or bone mineral density. However, 14 of these 52 patients (27 percent) had progression of disease, defined as the development of at least one new indication for parathyroidectomy. All 20 patients with symptoms had kidney stones. None of the 12 who underwent parathyroidectomy had recurrent kidney stones, whereas 6 of the 8 patients who did not undergo surgery did have a recurrence. CONCLUSIONS: In patients with primary hyperparathyroidism, parathyroidectomy results in the normalization of biochemical values and increased bone mineral density. Most asymptomatic patients who did not undergo surgery did not have progression of disease, but approximately one quarter of them did have some progression. PMID- 10528036 TI - A preliminary study of long-term treatment with interferon gamma-1b and low-dose prednisolone in patients with idiopathic pulmonary fibrosis. AB - BACKGROUND AND METHODS: Patients with idiopathic pulmonary fibrosis have progressive scarring of the lung and usually die within four to five years after symptoms develop. Treatment with oral glucocorticoids is often ineffective. We conducted an open, randomized trial of treatment with a combination of interferon gamma-1b, which has antifibrotic properties, and an oral glucocorticoid. We studied 18 patients with idiopathic pulmonary fibrosis who had not had responses to glucocorticoids or other immunosuppressive agents. Nine patients were treated for 12 months with oral prednisolone alone (7.5 mg daily, which could be increased to 25 to 50 mg daily), and nine with a combination of 200 microg of interferon gamma-1b (given three times per week subcutaneously) and 7.5 mg of prednisolone (given once a day). RESULTS: All the patients completed the study. Lung function deteriorated in all nine patients in the group given prednisolone alone: total lung capacity decreased from a mean (+/-SD) of 66+/-8 percent of the predicted value at base line to 62+/-6 percent at 12 months. In contrast, in the group receiving interferon gamma-1b plus prednisolone, total lung capacity increased (from 70+/-6 percent of the predicted value at base line to 79+/-12 percent at 12 months, P<0.001 for the difference between the groups). In the group that received interferon gamma-1b plus prednisolone, the partial pressure of arterial oxygen at rest increased from 65+/-9 mm Hg at base line to 76+/-8 mm Hg at 12 months, whereas in the group that received prednisolone alone it decreased from 65+/-6 to 62+/-4 mm Hg (P<0.001 for the difference in the change from baseline values between the two groups); on maximal exertion, the value increased from 55+/-6 to 65+/-8 mm Hg in the group that received combined treatment and decreased from 55+/-6 mm Hg to 52+/-5 mm Hg in the group given prednisolone alone (P<0.001). The side effects of interferon gamma-1b, such as fever, chills, and muscle pain, subsided within the first 9 to 12 weeks. CONCLUSIONS: In a preliminary study, 12 months of treatment with interferon gamma 1b plus prednisolone was associated with substantial improvements in the condition of patients with idiopathic pulmonary fibrosis who had had no response to glucocorticoids. PMID- 10528035 TI - Lamivudine as initial treatment for chronic hepatitis B in the United States. AB - BACKGROUND AND METHODS: Although the nucleoside analogue lamivudine has shown promise in patients with chronic hepatitis B, long-term data on patients from the United States are lacking. We randomly assigned previously untreated patients with chronic hepatitis B to receive either 100 mg of oral lamivudine or placebo daily for 52 weeks. We then followed them for an additional 16 weeks to evaluate post-treatment safety and the durability of responses. The primary end point with respect to efficacy was a reduction of at least 2 points in the score on the Histologic Activity Index. On this scale, scores can range from 0 (normal) to 22 (most severe abnormalities). RESULTS: Of the 143 randomized patients, 137 were included in the efficacy analysis: 66 in the lamivudine group and 71 in the placebo group. The other six patients were excluded at the base-line visit because of the absence of a documented history of hepatitis B surface antigen for at least six months. After 52 weeks of treatment, lamivudine recipients were more likely than placebo recipients to have a histologic response (52 percent vs. 23 percent, P<0.001), loss of hepatitis B e antigen (HBeAg) in serum (32 percent vs. 11 percent, P=0.003), sustained suppression of serum hepatitis B virus (HBV) DNA to undetectable levels (44 percent vs. 16 percent, P<0.001), and sustained normalization of serum alanine aminotransferase levels (41 percent vs. 7 percent, P<0.001), and they were less likely to have increased hepatic fibrosis (5 percent vs. 20 percent, P=0.01). Lamivudine recipients were also more likely to undergo HBeAg seroconversion, defined as the loss of HBeAg, undetectable levels of serum HBV DNA, and the appearance of antibodies against HBeAg (17 percent vs. 6 percent, P=0.04). HBeAg responses persisted in most patients for 16 weeks after the discontinuation of treatment. Lamivudine was well tolerated. Self-limited post-treatment elevations in serum alanine aminotransferase were more common in lamivudine recipients: 25 percent had serum alanine aminotransferase levels that were at least three times base-line levels, as compared with 8 percent of placebo recipients (P=0.01). The clinical condition of all patients remained stable during the study. CONCLUSIONS: In U.S. patients with previously untreated chronic hepatitis B, one year of lamivudine therapy had favorable effects on histologic, virologic, and biochemical features of the disease and was well tolerated. HBeAg responses were generally sustained after treatment. PMID- 10528037 TI - Clinical consequences of electrocardiographic artifact mimicking ventricular tachycardia. PMID- 10528038 TI - Images in clinical medicine. Tremor as a cause of pseudo-ventricular tachycardia. PMID- 10528039 TI - Signaling pathways for cardiac hypertrophy and failure. PMID- 10528040 TI - Behcet's disease. PMID- 10528041 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 32-1999. A 44-year-old man with tracheal narrowing and respiratory stridor. PMID- 10528042 TI - Treatment of primary hyperparathyroidism. PMID- 10528045 TI - Correction: Whither Continuity of Care? PMID- 10528043 TI - Interferon gamma-1b for the treatment of idiopathic pulmonary fibrosis. PMID- 10528044 TI - Putting power into patient choice. PMID- 10528046 TI - Long-term care for frail elderly people--the On Lok model. PMID- 10528047 TI - Editorial PMID- 10528048 TI - Dual-energy X-ray absorptiometry: a review. AB - Dual-energy X-ray absorptiometry (DXA) has been recently used for body composition analysis in dialysis patients. It is based on the principle that X rays passed through various body tissues have different attenuation and, therefore, can be differentiated. By using X-rays at two different energy levels, better tissue differentiation is possible compared with single energy systems. This article will review the evolution of DXA scanners and the role DXA has in assessing body composition in dialysis patients. Overall, this technique has excellent precision and holds promise for use in the serial evaluation of body composition and nutritional evaluation of dialysis patients. PMID- 10528049 TI - Correlates of poor appetite among hemodialysis patients. AB - OBJECTIVE: Poor appetite is an important barrier to adequate nutrition among hemodialysis patients. We sought to determine the relationship between overall appetite and (1) patient demographic and medical characteristics and (2) appetite for specific foods. DESIGN: Cross-sectional study. SETTING: All 22 chronic hemodialysis units in northeast Ohio. PATIENTS: Two hundred ninety-eight randomly selected patents. INTERVENTION: Interview and chart abstraction. MAIN OUTCOME MEASURE: Overall patient appetite. RESULTS: Of all patients, 12% reported poor overall appetite, 20% reported fair appetite, and 69% reported good appetite. There was no relationship between overall appetite and patient demographic (age, gender, race, education, health insurance) or medical (cause of renal failure, years on dialysis, number of comorbid conditions, Kt/V) characteristics. There was little relationship between overall appetite and appetite for 20 specific foods. Of 34 patients with poor overall appetite, 29 (85%) identified at least three specific protein foods for which they had good appetite and 28 (82%) identified at least three nonprotein foods for which they had good appetite. CONCLUSION: Poor overall appetite is not associated with patient characteristics or appetite for specific foods. Most patients with poor overall appetite have good appetite for several specific foods. Helping patients increase the intake of these foods may be beneficial in improving nutritional status. PMID- 10528050 TI - The effects of parathyroidectomy on nutritional and biochemical status of hemodialysis patients with severe secondary hyperparathyroidism. AB - The nutritional and biochemical parameters of 15 chronic hemodialysis (HD) patients with severe secondary hyperparathyroidism who had undergone total parathyroidectomy (PTX), with a forearm implant, were retrospectively studied at 1, 3, 6, and 12 months pre- and post-PTX. The concentrations of serum calcium and phosphorous significantly decreased in the first 6 months post-PTX. The concentration of blood urea nitrogen significantly increased at 6 and 12 months post-PTX. In contrast with control chronic HD patients who had no weight gain after nonparathyroid surgery, there was a progressive weight gain leading to a significantly higher dry weight at 12 months post-PTX. There was no significant change in serum potassium, albumin, cholesterol, transferrin, bicarbonate, hematocrit, normalized protein catabolic rate (nPCR), or erythropoietin dose at any time point post-PTX. The biochemical parameters of the 8 patients who had more than 5% weight gain, during 12 months post-PTX, were not statistically different from the remaining 7 cases who had weight gain less than 5% (or had lost weight) in the same time period. The same was true for 4 patients with weight gain of more than 10% versus the latter group. In conclusion, HD patients with severe secondary hyperparathyroidism are prone to progressive weight gain post-PTX, which reaches significance by the twelfth month. In 53% of the patients, the weight gain is more than 5% above the baseline. The nutritional and biochemical parameters pre-PTX were not helpful in distinguishing those who developed significant weight gain post-PTX. PMID- 10528051 TI - Bioelectrical impedance measurement: errors and artifacts. AB - Bioelectrical impedance analysis (BIA) allows simple noninvasive estimation of body water, and it could potentially be a very useful technique for clinical monitoring and study of abnormalities of body water. It has been shown that the total body impedance is dominated by the arm (46%) and leg (44%). The trunk, which represents an average of 46% of the body weight, accounts for only 10% of the total impedance. The objective of the current study was to determine the errors in prediction of body composition from BIA when applied to dialysis patients with measurement on the nondominant arm, postural changes, muscular contractions or cramps, monolateral lymphoedema, arteriovenous fistula, central venous catheter, or vascular graft. We studied 20 healthy subjects, 20 uremics on chronic hemodialysis, 3 uremics with fever (body temperature >38.5 degrees C), 3 uremics with cramps, 3 patients with monolateral lymphoedema of an arm, and 3 patients with a prosthetic fistula on an arm. The results of our study show different values of total body water (TBW) derived by BIA measurements effected on supine or standing position (percentage rate variation = 1.1% to 1.6%), or effected during fever (6%), during cramps (-0.73%), with lymphoedema (25%), or in presence of a native arteriovenous fistula, a catheter in a central vein, or a graft (between -24% and +4%). We concluded that a significant error may occur in the measurement of body composition from whole body BIA when performed in the reported cases. PMID- 10528052 TI - A review of food provision to a renal ward and the proposed appointment of feeding assistants. AB - Malnutrition among renal patients has been widely documented and is associated with increased morbidity and mortality. Advances in dialysis technology and transplantation have helped to increase patient long-term survival, and as more elderly patients commence dialysis programs, the problem of malnutrition is escalating. Hospitalized renal patients are at a greater risk, as dietary intakes may be reduced for a number of reasons. A multidisciplinary team decided to review the existing cook-chill plated meal system and all food provision to the renal ward. The aim of this review was to assess the nutritional intake of renal inpatients and to gauge patients' and relatives' attitudes towards hospital food provision. From these results, we hoped to go on and implement some changes to help improve the situation. Results showed that 34% of the patients ate half or less of the hospital food provided, and 80% of patients surveyed relied on food brought in by relatives and friends. Actual dietary intakes were compared to Dietary Reference Values (DRVs; Department of Health [DoH], UK, 1995). One hundred percent of the patients did not achieve the DRVs for energy, iron, potassium, zinc, folate, B6, and riboflavin. Sixty-six percent of the patients did not achieve the DRV for protein. These results were discussed by the multidisciplinary group, and it was decided to trial a cook-chill bulk trolley to replace the existing plated meal system. Unfortunately, to implement a bulk trolley system, the ward needs someone to serve the food. This could be the job of a "feeding assistant" or "ward hostess." A bid has been put forward to the hospital Trust Board to obtain funding for these "feeding assistants, " and the bulk trolley can be acquired from existing funds. It is hoped that the creation of these new posts will go some of the way towards improving the patients' dietary intake while in the hospital. PMID- 10528053 TI - Gastroparesis and jejunal feeding. AB - A kidney transplant patient with diabetic gastroparesis was effectively treated by jejunal feeding. The patient, a 31-year-old woman, has a complicated medical history, with insulin-dependent diabetes mellitus. Complications include kidney failure followed by transplantation, bilateral knee amputations, and being registered blind. She was admitted with nausea and vomiting for the previous 6 days; the provisional diagnosis was diabetic gastroparesis. Various treatments were tried, including several prokinetic drugs and total parenteral nutrition. The total parenteral nutrition provided most of the patient's nutritional requirements, and, only slight weight loss was observed. Nothing seemed to improve the symptoms of vomiting. An endoscopic retrograde cholangiopancreatography, a radiographic examination of the bile and pancreatic ducts, was performed to exclude obstruction. At the same time, having found nothing, a gastrostomy was placed with a jejunal extension. Feeding was established within 3 days. Her weight remained stable after 7 weeks of jejunal feeding. She had started to increase her oral intake of solid foods and fluids. By 8 weeks, she was taking a full oral diet and fluids. Now, 14 weeks after the placement of the gastrostomy tube with the jejunal extension, she is doing well. Her weight remains stable and her oral intake is excellent. Her diabetes is under control. After 22 weeks, the gastrostomy was removed. After this success with jejunal feeding when all other treatments had failed, this treatment could be used to treat future diabetic gastroparesis. Slow introduction of the feed seems to help toleration. PMID- 10528056 TI - Literature review PMID- 10528055 TI - The rehab exercise "E": a natural role for renal dietitians. AB - Quality of life and physical well-being are significant concerns in end-stage renal disease (ESRD). The complications of ESRD combined with patient illness and inactivity result in deconditioning and disability. There has been a recent movement toward rehabilitation for these individuals in the past several years. The following information was presented at the Forty-eighth Annual Meeting of the National Kidney Foundation on October 25, 1998 in Philadelphia, PA. The author presents exercise programs and equipment options for use within the ESRD facility and discusses the crucial role the registered dietitian plays in such a program. PMID- 10528054 TI - Successful intradialytic parenteral nutrition after abdominal "Catastrophes" in chronically hemodialysed patients. AB - OBJECTIVE: To assess the therapeutic contribution of intradialytic parenteral nutrition (IDPN) in four acutely ill, hypercatabolic, hemodialysed patients. All underwent major surgery, complicated by infection and malnutrition. DESIGN: A retrospective clinical study. SETTING: An in-center hemodialysis unit, at a tertiary referral hospital. PATIENTS: Patient 1: a young woman, with a good renal transplant. Developed gastric lymphoma, which required gastrectomy. After cessation of immunosuppression, "lost" her kidney and returned to hemodialysis. Received IDPN for 4 months and recovered well from severe malnourishment. Patient 2: an elderly, malnourished man, on continuous ambulatory peritoneal dialysis (CAPD). Developed biliary peritonitis and bacteremia. In a 3-month period, the patient had four operations. Maintained on IDPN for 4 months. Patient 3: a young and obese man, who suffered from life-threatening staphylococcal aureus peritonitis, resulting in widespread bowel adhesions. Underwent repeated aspirations of purulent ascites, laparoscopy, and explorative laparotomy. IDPN was administered for 4 months and stopped on the patient's request. Patient 4: a young man, who after cadaveric renal transplantation remained hospitalized for 6 months because of acute rejection and peritoneal and retroperitoneal abscesses. Had major surgery performed seven times. Received IDPN for 6 months, and is now well. RESULTS: All four patients benefited from 4 to 6 months of IDPN, as an integral part of intensive supportive and nutritional treatment. Weight loss was halted, as patient appetite returned and oral nutrition became adequate. Estimated daily protein intake reached 1.2 g/kg, while caloric intake rose to nearly 30 kcal/kg/d (Table 3). Mean serum albumin levels increased from 25.5 g/L +/- 0.9 g/L to 38.0 g/L +/- 1.5 g/L. No adverse side effects were seen from IDPN. CONCLUSION: IDPN is a worthwhile part of treatments used in the catabolic, postoperative hemodialysed patient. It is safe and efficient when used over a 6 month period in trying to attenuate existing, or worsening malnutrition in these patients. It should be commenced at an early stage in these patients, after attempts at oral nutritional support have been deemed inadequate. PMID- 10528058 TI - Message from the chairperson PMID- 10528057 TI - Holiday eating guidelines PMID- 10528059 TI - Subject index PMID- 10528060 TI - Preface PMID- 10528061 TI - Minimally invasive approaches in venous surgery. PMID- 10528062 TI - Endoscopic vein harvest techniques for coronary and infrainguinal bypass. AB - Endoscopic saphenous vein harvest represents a minimally invasive approach to obtain a suitable bypass conduit for coronary or extremity revascularization. Endoscopic vein harvest has been designed to reduce wound complications in a population typically at risk for problematic wound healing. Most studies have shown a reduction in such wound healing complications and improved patient comfort, which may result in fewer postoperative visits. The technique of endoscopic saphenous vein harvest is described, and the current limitations of the procedure are discussed. PMID- 10528063 TI - Endovascular in situ bypass. AB - The history of arterial bypass dates back to the turn of the century. Recent advances have reduced the significant morbidity of some of these procedures. PMID- 10528064 TI - Laparoscopically assisted abdominal aortic aneurysm repair. AB - Since the advent of laparoscopy, the sweeping changes seen in general surgery have not been paralleled in vascular surgery. However, the application of laparoscopic techniques to intraabdominal vascular procedures has now progressed from the animal laboratory to the clinical arena. Initial experience with laparoscopically assisted aortic bypasses for occlusive disease has led to the development of procedures for aneurysmal disease. This article reviews the current clinical experience in the evolving technique of laparoscopically assisted abdominal aortic aneurysm repair. PMID- 10528065 TI - Totally laparoscopic abdominal aortic aneurysm repair. AB - Current experience with totally laparoscopic aortic aneurysm repair is reviewed with particular attention to the techniques of surgery. Vascular surgery has been slow to enter the field of minimally invasive surgery because of the unique difficulties of managing arterial anatomy with minimal access techniques. Laparoscopic instrumentation has undergone a stunning evolution, and surgeon experience with minimally invasive surgery has grown exponentially. This dramatic revolution has allowed several groups to perform laparoscopic aortic vascular surgery. The surgical approach that each group has taken has varied. The approaches have included both laparoscopically assisted and totally laparoscopic aortic surgery with both transperitoneal and retroperitoneal approaches to the aorta. A review of these varied techniques will be discussed and include our experience with totally laparoscopic aortic surgery. This experience includes both transperitoneal and retroperitoneal approaches to infrarenal aortic aneurysms. An extended discussion of our surgical technique for aneurysm bypass is included. Patient selection, patient positioning, and trocar placement are described. The pattern of surgery for both techniques is enumerated, and postoperative care is discussed. However, the world experience with minimally invasive vascular surgery remains small, therefore a wider acceptance will require a prospective, randomized trial that shows an equally as safe surgical approach as provided open vascular surgery. With its acceptance, minimally invasive vascular surgery should show the patient benefits that befall minimally invasive surgery patients. PMID- 10528066 TI - Technological advances in laparoscopic aorto-occlusive surgery. AB - Minimally invasive surgery (MIS) has been recognized as increasingly beneficial to patients undergoing various cardiovascular surgical procedures. Cardiac applications with MIS techniques and technologies are being shown as beneficial in heart valve replacement and in coronary artery bypass. In vascular surgery, benefits are being reported for endoscopic saphenous vein harvesting as well as endoscopic ligation of incompetent perforators. Since 1993, applications of laparoscopy to aortic surgery have been reported. Until these reports, percutaneous interventional procedures have been the mainstay of MIS vascular work for aortoiliac disease. Reported laparoscopic techniques have ranged from laparoscopically assisted techniques to procedures performed completely laparoscopically. Several studies show that laparoscopic aortic surgery is feasible. These show the known advantages of MIS for patients, with decreased use of analgesics, shortened ileus, earlier ambulation, and shortened length of stay. Laparoscopy has been showing a growing role in the armamentarium of the modern vascular surgeon. PMID- 10528067 TI - Design of infection-resistant antibiotic-releasing polymers: I. Fabrication and formulation. AB - Biomaterials-related infections are often observed with prosthetic implants and in many cases result in the failure of the devices. To design a biomedically useful polymer that is intrinsically infection-resistant, we have developed a ciprofloxacin-loaded polyurethane (PU) matrix that releases antibiotic locally at the implant surface, thereby minimizing bacterial accumulation. We report here the methods of fabrication and formulation for making such antibiotic-loaded devices, as well as evidence of their bactericidal properties. Specifically, various pore-forming agents and drug loadings were examined. An optimum formulation consisting of BIOSPAN PU, poly(ethylene glycol) and ciprofloxacin offered the longest effective period of sustained release (5 days). The bactericidal efficacy of the released ciprofloxacin against Pseudomonas aeruginosa (PA) was four times that of the control PU without antibiotics. This bactericidal efficiency was due to an increase in the PA detachment from the surface. These observations suggested that the released ciprofloxacin was biologically active in preventing the bacteria from permanently adhering to the substratum, and thus decreasing the possibility of biofilm-related infection. PMID- 10528068 TI - Design of infection-resistant antibiotic-releasing polymers. II. Controlled release of antibiotics through a plasma-deposited thin film barrier. AB - In the first paper in this series, we described the methods to synthesize an antibacterial polyurethane (PU) incorporating ciprofloxacin as the releasable antibiotic and poly(ethylene glycol) as the pore-forming agent. Here, we report that a thin, RF-plasma-deposited, n-butyl methacrylate (BMA) overlayer on this drug-loaded PU can act as a rate-limiting barrier to achieve a constant, sustained release of ciprofloxacin. Deposition power and deposition time during the coating process were optimized to give an appropriate crosslinked coating barrier that yielded desirable release rates, above the minimum required killing rate, N(kill). Electron spectroscopy for chemical analysis (ESCA), also known as X-ray photoelectron spectroscopy (XPS), was used to characterize the coating, and its crosslinking degree was indirectly related to the C/O ratio. Increasing either deposition power (10-60 W) or duration (5-25 min) resulted in increased C/O ratios and decreased ciprofloxacin release rates. The correlation between increased C/O ratios and reduced release rates is believed to be due to the increased crosslinking, increased hydrophobicity and increased thickness of the coating. The optimal plasma conditions to attain an appropriate crosslinked plasma-deposited film (PDF) required argon etching, pre-treatment of the matrices with an 80W-BMA plasma for 1 min, followed by immediate BMA plasma deposition at 40 W and 150 mT for 20 min. By using these plasma deposition protocols, we eliminated the initial burst effect, significantly reduced the release rates, and closely approached the zero order release kinetics for at least five days. In this study, we also showed that ESCA could be used as a powerful tool to explain the release behavior of molecules through the plasma-deposited films (PDFs). PMID- 10528069 TI - Microstructure and release characteristics of the minipellet, a collagen-based drug delivery system for controlled release of protein drugs. AB - We have developed the minipellet, a matrix-type system for the sustained delivery of protein drugs using collagen as a biodegradable drug carrier. In this study, we analyzed the microstructure and release profile of the minipellet containing human serum albumin (HSA) as a model drug.The findings suggest that the minipellet has a structure in which collagen fibers are strongly oriented in the direction of extrusion from the nozzle in the molding process of the minipellet, and that HSA exists as fine particulate clusters which are homogeneously distributed among the collagen fibers in the minipellet. During release, the HSA clusters dissolve and HSA is retained within the collagen matrix as a solution.The results of release experiments indicate that HSA release from the minipellet is mainly controlled by diffusion in the collagen matrix, and that sustained release is achieved by the dense structure of the collagen matrix which is formed in the manufacture process. In addition, more detailed study suggests that the minipellet has unique directional release behavior caused by its microstructure. PMID- 10528070 TI - Induction of cytotoxic T-cell responses following oral immunization with synthetic peptides encapsulated in PLG microparticles. AB - CTL responses play a critical role in clearing viral infections. We have investigated the potential of poly(lactide-co-glycolide) (PLG) microparticles as an oral delivery system for peptides representing CTL epitopes from measles virus nucleoprotein. Oral administration of CTL epitopes encapsulated in 50:50 PLG microparticles, resulted in vivo priming of splenic peptide-specific CTL responses. However, the observed CTL lysis was low and cofeeding of encapsulated peptide with cholera toxin as a mucosal adjuvant did not result in any significant enhancement of the observed CTL responses. The pronounced immunostimulatory effect of microparticles, combined with their excellent tissue compatibility and biodegradability makes them a valuable delivery system for synthetic peptide immunogens. However, further work is needed to improve their efficiency via the oral route. PMID- 10528071 TI - Phase inversion dynamics of PLGA solutions related to drug delivery. Part II. The role of solution thermodynamics and bath-side mass transfer. AB - The role of solvent properties and bath-side composition on the phase inversion dynamics and in vitro protein release kinetics of polylactic-co-glycolic acid (PLGA) solutions has been examined using dark ground imaging, in vitro release rate, and SEM techniques. Thermodynamic phase diagrams for three model systems (PLGA in 1-methyl-2-pyrrolidinone (NMP), triacetin, and ethyl benzoate) suggest two general classes of precipitation behavior, depending on the relative solvent strength and water miscibility. Drug release from the NMP-based system is primarily governed by the dynamics of phase inversion and exhibits a distinct burst region followed by a much slower release. Alternatively, depots with low solvent/water affinity (PLGA in triacetin or ethyl benzoate) undergo much slower phase inversion, resulting in a less porous, more fluid, two-phase structure that also releases protein more uniformly. Addition of a small chain triglyceride or organic salt to the aqueous receptor bath also evokes a significant increase in the mass transfer rate of protein from the low solvent/non-solvent affinity depots. An interpretation of these results in terms of a qualitative model for the protein release mechanism is also given. PMID- 10528072 TI - Synthesis of a single-tailed cationic lipid and investigation of its transfection. AB - Single-tailed cationic lipids were originally reported to have low transfection efficiency and high toxicity in plasmid delivery. We hypothesized that particular single-tailed cationic lipids may also function in plasmid transfection. To test this hypothesis, we synthesized a new cationic lipid-oleoyl ornithinate (OLON). To decrease cytotoxicity, we then introduced a potential biodegradable ester bond in the tail of lipid yielding 6-lauroxyhexyl ornithinate (LHON). The data demonstrated that the cytotoxicity of LHON was lower than that of 1,2-dioleoyl-3 trimethylammonium-propane (DOTAP) or OLON. To investigate the transfection activity of the new lipids and determine the cellular uptake of DNA/liposome complexes, we compared the transfection of liposomes produced from double-tailed 1',2'-dioleyl-sn-glycero-3'-succinyl-1, 6-hexanediol ornithine conjugate (DOGSHDO) with an ornithine headgroup, single-tailed OLON with an ornithine head group, double-tailed DOTAP with quaternary amine group, and single-tailed cetyltrimethylammonium bromide (CTAB) with a quaternary amine group. At the optimal ratios as defined in transfection experiments, OLON/DOPE had more than 10 times the transgene expression than other liposomes even though the DNA uptake was not necessarily greater. In the experiments comparing the release of DNA from DNA/liposome complexes by anionic substances, a greater fraction of DNA was released from DNA/OLON/DOPE complexes than that from DNA/DOTAP/DOPE complexes. PMID- 10528073 TI - In vitro percutaneous absorption enhancement of propranolol hydrochloride through porcine epidermis by terpenes/ethanol. AB - The purpose of this study was to investigate the mechanism(s) of percutaneous absorption enhancement of propranolol hydrochloride (PHCL) across porcine epidermis by terpenes (e.g. menthone and limonene) in combination with ethanol. The in vitro percutaneous absorption experiments were performed using Franz diffusion cells. The solubility of PHCL in control and enhancer solutions was determined through high-performance liquid chromatography. Partitioning of PHCL to powdered SC from control or enhancer solutions was also determined in order to investigate the binding of PHCL to the SC from the SC/enhancer system. Fourier transform infrared spectroscopy (FT-IR) was employed to study the biophysical changes in stratum corneum (SC) lipids. The in vitro macroscopic barrier properties were investigated by measuring transepidermal water loss (TEWL) using Tewameter. Five percent menthone or limonene in combination with ethanol (EtOH) (menthone/EtOH or limonene/EtOH) significantly increased (P<0.05) the flux of PHCL through porcine epidermis in comparison to the control (EtOH). The partitioning of PHCL to the SC from the SC/enhancer system was also significantly greater than the SC/control system. The above enhancers showed a decrease in the peak heights and areas for both asymmetric and symmetric C-H stretching absorbances in comparison with the untreated SC, indicating the SC lipids extraction. Menthone/EtOH and limonene/EtOH enhanced (P<0.05) the in vitro TEWL through the epidermis in comparison to the control. Thus, an enhancement in the flux of PHCL by menthone/EtOH and limonene/EtOH is due to SC lipid extraction, macroscopic barrier perturbation, and improvement in the partitioning of the drug to the SC. PMID- 10528074 TI - Elasticity of vesicles affects hairless mouse skin structure and permeability. AB - One of the possibilities for increasing the penetration rate of drugs through the skin is the use of vesicular systems. Currently, special attention is paid to the elastic properties of liquid-state vesicles, which are supposed to have superior properties compared to gel-state vesicles with respect to skin interactions. In this study, the effects of vesicles on hairless mouse skin, both in vivo and in vitro, were studied in relation to the composition of vesicles. The interactions of elastic vesicles containing the single chain surfactant octaoxyethylene laurate-ester (PEG-8-L) and sucrose laurate-ester (L-595) with hairless mouse skin were studied, in vivo, after non-occlusive application for 1, 3 and 6 h. The skin ultrastructure was examined by ruthenium tetroxide electron microscopy (TEM) and histology. The extent, to which vesicle constituents penetrated into the stratum corneum, was quantified by thin layer chromatography (TLC). The interactions of the elastic vesicles containing PEG-8-L and L-595 surfactants were compared with those observed after treatment with rigid vesicles containing the surfactant sucrose stearate-ester (Wasag-7). Furthermore, skin permeability experiments were carried out to investigate the effect of treatment with PEG-8-L micelles, elastic vesicles (containing PEG-8-L and L-595 surfactants) or rigid Wasag-7 vesicles on the 3H(2)O transport through hairless mouse skin, in vitro, after non-occlusive application. Treatment of hairless mouse skin with the elastic vesicles affected the ultrastructure of the stratum corneum: distinct regions with lamellar stacks derived from the vesicles were observed in intercellular spaces of the stratum corneum. These stacks disrupted the organization of skin bilayers leading to an increased skin permeability, whereas no changes in the ultrastructure of the underlying viable epidermis were observed. Treatment with rigid Wasag-7 vesicles did not affect the skin ultrastructure or skin permeability. TLC measurements showed that after 1 h of non-occlusive application of elastic or rigid vesicles, a six-fold increased amount of elastic vesicle material was present within the stratum corneum compared to rigid vesicle material. After 3 and 6 h of application the amount of PEG-8-L vesicle material in SC decreased to approximately three- and two-fold, respectively, compared to Wasag-7 vesicle material. Pretreatment of the hairless mouse skin with the elastic vesicles containing 70 mol% PEG-8-L increased the diffusion of 3H(2)O with an optimum application dose of 2.5 mg lipids/cm(2) compared to PBS pretreatment. No significant difference in the enhancement of the 3H(2)O-diffusion was observed between PEG-8-L micelles or elastic vesicles containing 30 or 70 mol% PEG-8-L. Pretreatment with the rigid Wasag-7 vesicles decreased the diffusion rate of 3H(2)O, most probably by the formation of a lipid layer on the skin surface. The effect of the elastic vesicles on the skin permeability is supported by the ultrastructural changes observed by TEM in the intercellular lipid domains. The elastic vesicles containing 70 mol% PEG-8-L disorganize the lipid bilayers thereby creating or modifying pathways for possible drug penetration. PMID- 10528075 TI - Poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) (Pluronic)/poly(epsilon-caprolactone) (PCL) amphiphilic block copolymeric nanospheres. I. Preparation and characterization. AB - Amphiphilic block copolymers based on PEO-PPO-PEO block copolymer (Pluronic) and poly(epsilon-caprolactone) were synthesized by bulk polymerization. The structural analysis of Pluronic/PCL block copolymer was carried out using FT-IR, 1H NMR, GPC, WAXD, DSC and TGA measurements. To prepare copolymeric nanospheres with a micellar structure, Pluronic/PCL amphiphilic block copolymers were dialyzed against water. The size and size distribution of Pluronic/PCL block copolymeric nanospheres were examined by dynamic light scattering measurement. They showed an average diameter of 116 to 196 nm depending on the type of copolymer. All the nanosphere samples exhibited a narrow size distribution. The critical micelle concentrations of Pluronic/PCL amphiphilic block copolymers determined by fluorescence spectroscopy were lower than that of common low molecular weight surfactants. We confirmed the formation of stable copolymeric nanospheres through the solution behavior of amphiphilic block copolymer in selective solvents. PMID- 10528076 TI - A controlled release system for proteins based on poly(ether ester) block copolymers: polymer network characterization. AB - The properties of a series of multiblock copolymers, based on hydrophilic poly(ethylene glycol) (PEG) and hydrophobic poly(butylene terephthalate) (PBT) blocks were investigated with respect to their application as a matrix for controlled release of proteins. The degree of swelling, Q, of the copolymers increased with increasing PEG content and with increasing molecular weight of the PEG segment. Within the composition range tested, Q varied from 1.26 for polymers with PEG segments of 600 g/mol and a PBT content of 60 weight.% up to 3.64 for polymers with PEG segments of 4000 g/mol and a PEG/PBT weight ratio of 80:20. Equilibrium stress (compression)-strain measurements were performed in order to estimate mesh sizes. The mesh size of the copolymers ranged from 38 to 93 A, which was experimentally confirmed by diffusion of vitamin B(12) (hydrodynamic diameter d(h)=16.6 A), lysozyme (d(h)=41 A) and bovine serum albumin (d(h)=72 A). The in vitro degradation of PEG/PBT copolymers with a PEG block length of 1000 g/mol and PEG/PBT weight ratios of 70:30, 60:40 and 40:60 was studied. Matrices with increasing PEG contents exhibited a faster weight loss in phosphate-buffered saline (pH 7.4) at 37 degrees C. Over a degradation period of 54 days, M(n) decreased by about 35-45%, while the composition of the matrices, determined by NMR, remained almost constant. PMID- 10528077 TI - Transmucosal sustained-delivery of chlorpheniramine maleate in rabbits using a novel, natural mucoadhesive gum as an excipient in buccal tablets. AB - The objective of this study was to evaluate the gum from Hakea gibbosa (Hakea) as a sustained-release and mucoadhesive component in buccal tablets following their application to the buccal mucosa of rabbits. Flat-faced core tablets containing either 22 or 32 mg of Hakea and 40 or 25 mg of chlorpheniramine maleate (CPM) per tablet with either sodium bicarbonate or tartaric acid in a 1:1.5 molar ratio were formulated using a direct compression technique and were coated with Cutina(R) on all but one face. The resulting plasma CPM concentration versus time profiles were determined following buccal application of the tablets in rabbits. The strength of mucoadhesion of the tablets was also quantitated in terms of the force of detachment as a function of time. Following the application of the mucoadhesive buccal tablets, the following values for several pharmacokinetic parameters were obtained. The force of detachment for the mucoadhesive buccal tablets containing 22 mg of Hakea and either 25 and 40 mg CPM, and 32 mg Hakea and 40 mg CPM increased from 1.64+/-0.47 to 7.32+/-0.34 N, 1.67+/-0.30 to 7.21+/ 0.36 N, and 2.93+/-0.73 to 7.92+/-0.60 N, respectively from 5 to 90 min following application to excised intestinal mucosa. Addition of either sodium bicarbonate or tartaric acid, as well as higher amounts of CPM, did not affect the mucoadhesive bond strength. These results demonstrate that the novel, natural gum, H. gibbosa, may not only be used to sustain the release of CPM from a unidirectional-release buccal tablet, but also demonstrate that the tablets are sufficiently mucoadhesive for clinical application. The mucoadhesive strength as measured by the force of detachment, can be modulated by altering the amount of Hakea in the tablet. The mucoadhesive buccal tablets evaluated represent an improved transbuccal delivery system for conventional drug substances. PMID- 10528078 TI - An in vitro investigation into the potential for bimodal drug release from pectin/chitosan/HPMC-coated tablets. AB - The influence of disintegrant on the water uptake and subsequent disintegration force developed was investigated in a simple tablet formulation. The results indicated that a reasonable correlation existed between water uptake and disintegration force for the disintegrants screened with cross linked polyvinyl pyrrolidone (PVP XL) showing a proportionally higher disintegration force for the amount of water imbibed. Two tablet formulations, intended to promote accelerated drug release in the colon, were prepared, with and without PVP XL, and film coated with a mixture of pectin, chitosan and HPMC. The two systems showed different drug release rates which were influenced by the pH of the dissolution medium. In the presence of pectinolytic enzyme, drug release was faster when compared to release in buffer alone for both systems although the mechanism differed for each. Drug release in simulated gastrointestinal conditions showed a bimodal profile with the increased drug release rate being triggered by the action of pectinolytic enzymes. PMID- 10528079 TI - Effect of amphotericin B lipid formulation on immune response in aspergillosis. AB - The immune response against Aspergillus fumigatus has been studied during infection and therapy in order to understand the mechanism of pathogenesis and the effect of treatment with amphotericin B. With this in view an animal model of aspergillosis was developed in Balb/c mice by intravenous injection of an optimized dose of 3. 6x10(6) A. fumigatus spores. Infection due to Aspergillus was well established by histopathological examination and fungal load in the animal. Lesions and eosinophil infiltration was observed in the infected tissues which indicated the involvement of a Type I hypersensitivity response. Evaluation of serological parameters indicated high levels of interleukin-4 (IL-4) and A. fumigatus specific IgG antibodies. The reduction in fungal load and modulation of immune response in the infected mice was studied following treatment with amphotericin B/cholesterol hemisuccinate vesicles (ABCV). The results clearly indicated significant reduction in the fungal load, disappearance of eosinophils and lesions with the appearance of macrophages and neutrophils in the infected lung tissue, a decrease in IL-4 (fourfold) and a concomitant increase of interferon-gamma (IFN-gamma; twofold) with an improvement in general condition of mice. In the non-treated mice, the rise of IL-4 level indicated the association of T(H)2 cell response with susceptibility to infection while the increase of IFN gamma in the treated group suggested that T(H)1 cell response may be involved in resistance to Aspergillus infection. PMID- 10528080 TI - A centrifuge technique for the evaluation of the extent of water movement in wet powder masses. AB - A centrifuge method has been applied to the assessment of water retention in pharmaceutical powders. Five drug models and microcrystalline cellulose (MCC) were each mixed with different amounts of water and centrifuged at different speeds. The amount of water retained by the wet mass was evaluated by drying the powders to constant weight. Binary mixtures of each of the five model drugs, MCC and water were also processed in the same way. From the amount of water extracted the moisture retention capacity (MRC) was calculated. The MCC retained water more strongly than the different drug models over a wider range of initial water contents. The five drug models, although similar in their chemical structure, were divided into two groups, in terms of their MRC values. 4-HBA and propyl gallate recorded higher MRC values than methyl, propyl and butyl paraben. For the drug models mixed with MCC, the MRC values recorded were similar, though it was still possible to divide the drugs into the two subgroups. A correlation between the MRC value recorded for the different systems and the hydrogen bonding solubility component was found. The application of different centrifuge speeds indicated that within the same material there were different mechanisms of water retention. PMID- 10528081 TI - Enhanced hepatocyte uptake and liver targeting of methotrexate using galactosylated albumin as a carrier. AB - Liver targeting of drugs has wide therapeutic implications due to numerous liver related diseases. Using conjugates of methotrexate (MTX) to variously galactosylated bovine serum albumin (BSA), we studied whether we could enhance the liver targeting of MTX, a model drug, via galactose receptors selectively abundant on the hepatocytes. Here, we report that the galactosylation of the carrier protein BSA significantly enhanced the hepatocyte uptake and liver targetability of MTX. In vitro, the amount of MTX taken up by rat hepatocytes was positively correlated with the galactose content in BSA. MTX conjugates were relatively stable in plasma, but released MTX with time in liver homogenates. These results imply that the conjugates would exert low toxicity in the blood, but have therapeutic activity in the liver by liberating MTX. In vivo, MTX galactosylated BSA conjugates (MTX-L(24)BSA) showed significantly different pharmacokinetics from free MTX or MTX-BSA conjugates. The plasma level of free MTX rapidly declined in a biexponential fashion with an apparent terminal half life of 0.35 h. MTX-BSA conjugates showed the slowest decline with an apparent terminal half-life of 6 h, whereas MTX-L(24)BSA showed a biphasic pattern; a rapid distributive phase with a half-life of 0.567 h and a slow terminal phase. MTX-L(24)BSA showed the highest liver targetability, when evaluated in terms of two indices based on the area under the total amount of radioactivity-time curve (AUQ); Te*(liver), % AUQ(liver) to total AUQ, and te*, the ratio of AUQ(liver) to AUQ(kidney). Compared with free MTX and MTX-BSA, MTX-L(24)BSA showed about twofold higher Te*(liver) of 87.5%. The te* of MTX-L(24)BSA was 25- and fourfold higher than those of free MTX and MTX-BSA, respectively. Moreover, MTX-L(24)BSA showed a gradual increase in the therapeutically active intact form of MTX in the liver while showing the lowest level of intact MTX in the kidney. These results suggest that galactosylated BSA has a great potential as an hepatocyte-directed and more effective liver targeting carrier of drugs for liver diseases. PMID- 10528083 TI - The filling of granules into hard gelatine capsules. AB - Four different granule size fractions of Sorbitol instant(R) were filled into hard gelatine capsules on a tamp filling (Bosch) and a dosator nozzle machine (Zanasi) to allow comparison of the filling principles. An acceptable filling performance was always achieved and was independent of the machine type employed. Tamp filling was found to be slightly better for the coarser granule size fractions, because it does not seem to rely on a firm plug formation. A direct relationship between the angle of internal flow (Varthalis and Pilpel, 1976) and the coefficient of fill weight variation was found for both systems. Using the dosator nozzle machine, the plug formed was always denser than the maximum bulk density, whereas on the tamp filling machine for smallest granule size the maximum plug density could not be achieved with the settings employed. The results suggest that in situations where a low plug density is an essential prerequisite for drug dissolution and bioavailability the tamp filling machine appears the more suitable filling principle. However, if a greater extent of compression is required in order to fill large dose drugs or to use a smaller capsule size, the dosator nozzle principle might work more successfully for granules. PMID- 10528082 TI - Adriamycin release from flower-type polymeric micelle based on star-block copolymer composed of poly(gamma-benzyl L-glutamate) as the hydrophobic part and poly(ethylene oxide) as the hydrophilic part. AB - Star-block copolymer based on PBLG as the hydrophobic part and PEO as the hydrophilic one (as abbreviated GEG) was synthesized and characterized. Polymeric micelle was prepared by the diafiltration method. From the measurement of photon correlation spectroscopy, the nanoparticle sizes of GEG-1, GEG-2 and GEG-3 were 106.5+/-59.2, 43.8+/-0.7 and 13.5+/-1.0 nm in number average, respectively, indicating of the formation of polymeric micelle. Also, the nanoparticle sizes were dependent on the PBLG chain length, i.e. the more PBLG content in the copolymer, the larger the particle size. From the observation of transmission electron microscope(TEM), GEG-2 block copolymer had almost spherical shapes with size range about 20-70 nm, that was similar to particle size measurement. Fluorescence spectroscopy measurement indicated that GEG block copolymers associated in water to form polymeric micelles and critical micelle concentration (CMC) values of the block copolymers decreased with increasing PBLG chain length in the block copolymer. Characteristic peaks of the protons of the benzyl group in the PBLG and the methylene protons adjacent to the benzyl group of the PBLG segment in the GEG-2 nanoparticles appeared in 7.2 approximately 7.4 and 5.0 approximately 5.2 ppm, respectively, and disappeared in D(2)O, indicating the restricted motions of these protons within the micellar core and the very rigid structure of the PBLG core in the GEG polymeric micelles. Release of ADR from the polymeric micelles in vitro was slower in longer PBLG chain length and higher loading contents of ADR. PMID- 10528084 TI - Controlled release of dual drug-loaded hydroxypropyl methylcellulose matrix tablet using drug-containing polymeric coatings. AB - A dual drug-loaded hydroxypropylmethylcellulose (HPMC) matrix tablet simultaneously containing drug in inner tablet core and outer coated layer was formulated using drug-containing aqueous-based polymeric Eudragit RS30D dispersions. Effects of coating levels, drug loadings in outer layers, amount and type of five plasticizers and talc concentration on the release characteristics were evaluated on the characteristics in simulated gastric fluid for 2 h followed by a study in intestinal fluids. Melatonin (MT) was selected as a model drug. The surface morphology of dual drug-loaded HPMC tablets using scanning electron microscope (SEM) was smooth, showing the distinct coated layer with about 75 microm coating thickness at the 15% coating level. Unlike the uncoated and conventionally coated HPMC tablet, the dual drug-loaded HPMC matrix tablet gave a biphasic linear release, showing a zero-order for 4 h (first) followed by another zero-order release when fitted using linear regression (r(2) = 0.99). As the coating levels (15, 25%) increased, the release rate was further decreased. The biphasic release profiles of dual drug-loaded HPMC matrix tablet was unchanged except when 25% coating level containing 0.5% drug concentration was applied. As the drug concentration in polymeric coating dispersion increased (0.25-1.0%), the amount of drug released increased. The time for the first linear release was also advanced. However, the biphasic release pattern was not changed. The biphasic release profiles of dual drug-loaded HPMC matrix tablet were highly modified, depending on the amount and type of five plasticizers. Talc (10-30%) in coating dispersion as an anti-sticking material did not affect the release profiles. The current dual drug-loaded HPMC matrix tablet, showing biphasic release profiles may provide an alternative to deliver drugs with circadian rhythmic behaviors in the body but needs to be further validated in future in human studies. The dual drug-loaded coating method is also interesting for the modified release of poorly water-soluble drugs because solubilizers and other additives can be added in drug containing polymeric coating dispersions. PMID- 10528085 TI - In vitro transport and uptake of protohypericin and hypericin in the Caco-2 model. AB - The intestinal absorption characteristics of protohypericin, a protonaphthodianthrone present in Hypericum extract, were studied and compared with those of hypericin. The Caco-2 model was used as a model of the intestinal mucosa to assess transepithelial transport and cell uptake. The experimental work was performed in specific light conditions that prevented both the photoconversion of protohypericin into hypericin and the photosensitization of the cells. Following application of the individual compounds (80-200 microM) to the apical side of the monolayers, the appearance in the basolateral compartment was found to be very low (<0.5%/5 h), but was comparable for both compounds. A lag-time of 2-3 h was observed, suggesting gradual saturation of binding sites on the membrane or inside the cells. Uptake experiments of protohypericin and hypericin by Caco-2 cells revealed a very significant cellular accumulation (4 8%); uptake was characterised by saturation after 3 h. The findings of this study suggest that protohypericin has comparable absorption characteristics as hypericin and may contribute to the beneficial effect of Hypericum extract after oral dosing. PMID- 10528086 TI - Factors affecting the size distribution of liposomes produced by freeze-thaw extrusion. AB - This paper describes the development of a protocol for the production of liposomes using a freeze-thaw extrusion methodology. Laser diffraction particle size analysis showed that the median diameter of freeze-thawed egg phosphatidylcholine multilamellar vesicles (eggPC MLVs) was increased when cholesterol was included in the bilayers. Using a freeze-thaw cycle of 3 min freezing in liquid nitrogen at -196 degrees C followed by 3 min thawing at 50 degrees C resulted in an anomalously large particle size for eggPC/cholesterol formulations. When liposomes were repeatedly freeze-thawed a maximum size was achieved after five freeze-thaw cycles. Dispersion of liposomes in sodium chloride solutions promoted size increases following freeze-thawing, suggesting that vesicles had aggregated or fused. Poloxamers P338 and P407 inhibited the size increases observed during freeze-thawing for eggPC MLVs dispersed in 1.0 M NaCl, probably through steric prevention of aggregation and fusion. PMID- 10528087 TI - An opportunistic stability strategy; simulation with real data. AB - A multivariate modelling procedure is proposed in order to identify factors influencing stability, to estimate shelf-life, to select new batches for further stability testing and to evaluate changes in new batches. A model developed by the proposed procedure predicts the degradation rate constant as a function of storage temperature, pH, concentration and volume. The predicted rate constants were compared with prospectively measured rate constants, primarily from new batches stored under stress conditions, which emphasised batch differences earlier than storage under normal conditions. Strong deviations from expected rate constants led to extended testing of the batches concerned. The new data were used to upgrade the multivariate model. The procedure proposed led to the formulation of an opportunistic stability strategy (OSSY). Of the 15 batches of injectable solutions, nine batches are proposed tested by using OSSY. This led to an approximately 75% reduction in analytical measurements. Hold samples are recommended for storage under several stability conditions for back up analysis. In general, a multivariate stability model should be based on scientific data obtained from early studies, such as preformulation and formulation studies to provide both a qualitative and quantitative understanding of the mechanisms involved. PMID- 10528088 TI - A multivariate method to predict the water vapour diffusion rate through polypropylene packaging. AB - Semipermeable containers typically allow water to diffuse upon storage. The water diffusion rate of the widely distributed X-ray contrast agent Visipaque in polypropylene bottles and MR contrast agent Omniscan in polypropylene syringes has been evaluated. The goal was to develop a mathematical method for estimating the rate constant for water diffusion through the polypropylene wall as a function of several variables. The method presents an opportunity to measure the effect of the variables influencing product stability and thereby predicting the shelf-life at an early stage of the stability study. The effect of temperature, humidity, surface area, wall thickness, concentration of active ingredient and fill volume on the logarithmic rate constant for diffusion was estimated by partial least squares regression. The predictive ability of the cross-validated models was good (r = 0.99) for the two contrast agents. The models were used to predict shelf-life for relevant combinations of temperature and humidity, for the four defined climatic zones. PMID- 10528090 TI - The determination of a diffusional pathlength through the stratum corneum. AB - The stratum corneum possesses a very heterogenous structure. As such a diffusing molecule can access a number of different pathways. It is probable that the excellent barrier properties of the stratum corneum result from a tortuous diffusional pathway around the dead cells. However, there are considerable problems in designing diffusion experiments and analysing the data to prove, without doubt, which is the predominant pathway. The mathematical problems posed are discussed in this article. PMID- 10528089 TI - Influence of the formulation composition on the in vitro characteristics of hot stage extrudates. AB - The aim of this study was to evaluate the influence of the formulation composition on the characteristics of starch based hot stage extrudates in order to obtain high quality and high efficacy matrices for controlled drug delivery. Their characteristics were compared to those of a reference formulation, for which in vivo data were available. The influence of the starch type, the plasticizer and the lubricant were investigated. The extrudates were produced with a twin screw co-rotating extruder equipped with a twin screw powder feeder and a 3 mm cylindrical die. The extrudates were manually cut and dried for 48 h at 60 degrees C prior to analysis. They were characterized by Karl Fischer titration, Hg-porosimetry, 4-point bending and dissolution testing. Changing the formulation composition did not affect the water content or the porosity of the extrudates but had an influence on their mechanical strength and dissolution profiles. These characteristics were particularly dependent on the plasticizer and lubricant concentration used and on the nature of the starch component and the lubricant. Expansion, depending on starch conversion and amylose-lipid complex formation, played a major role in the explanation of the results obtained. All the formulations tested showed a slow drug release profile in vitro. However, the differences with the reference formulation--which was tested in vivo in earlier work--were probably too small to give a distinct improvement of its in vivo behaviour. PMID- 10528091 TI - Kinetics and mechanism of degradation of klerval, a pseudo-tetrapeptide. AB - The degradation of klerval (I) was studied as a function of pH. The extent and routes of degradation were found to be pH-dependent. Under strongly acidic conditions (pH<2), the drug predominantly undergoes specific acid-catalyzed hydrolysis of the side-chain amide bond yielding II8), the drug undergoes specific base-catalyzed hyrolysis yielding II and epimerization generating D epimer. The epimerization appears to occur via the succinimide intermediate in neutral pH region. With increasing pH, however, the epimerization rate increases due to direct epimerization of the peptides. PMID- 10528092 TI - Effect of additives on the physicochemical properties of liquid suppository bases. AB - To investigate the effects of additives on the physicochemical properties of in situ gelling and mucoadhesive liquid suppository base, gelation temperature, gel strength and bioadhesive force of liquid suppository base, poloxamer 407 (P 407) and poloxamer 188 (P 188) (15/15%) were evaluated in the presence of following additives: solvent (ethanol, propylene glycol, glycerin), ionic strength controlling agent (sodium chloride) and pH-controlling agent (hydrochloric acid, sodium monohydrogen phosphate, sodium dihydrogen phosphate). Among the additives studied, sodium chloride, sodium monohydrogen phosphate and sodium dihydrogen phosphate increased to a great extent the gel strength and the bioadhesive force of P 407/P 188 (15/15%) with a decrease in gelation temperature. Glycerin slightly decreased the gelation temperature and slightly increased the gel strength and bioadhesive force. However, the addition of 1% of sodium chloride, sodium monohydrogen phosphate or sodium dihydrogen phosphate caused a greater than 60-fold increase in gel strength and over a tenfold increase in bioadhesive force with 2-4 degrees C decrease of gelation temperature within optimal range, compared with P 407/P 188 (15/15%) alone. On the other hand, ethanol, propylene glycol and hydrochloric acid increased the gelation temperature and slightly decreased the gel strength and the bioadhesive force. Taken together, these findings indicate that the effect of additives on the physicochemical properties of liquid suppository bases depends on their bonding capacities, in that additives such as sodium chloride, sodium monohydrogen phosphate and sodium dihydrogen phosphate having strong cross-linking bonds with the components of liquid suppository base increase the strength and bioadhesive force of a gel compared to liquid suppository base alone, while additives such as ethanol, propylene glycol and hydrochloric acid having weaker hydrogen bonding result in a weaker response. Thus, sodium chloride and sodium phosphates appear to be promising additives for in situ gelling and mucoadhesive liquid suppository base, if used in adequate amounts. PMID- 10528093 TI - Development of a rectal nicotine delivery system for the treatment of ulcerative colitis. AB - The aims of this investigation were: i. to develop a rectal nicotine delivery system with bioadhesives for the treatment of ulcerative colitis and ii. to evaluate nicotine transport and cytotoxicity of the delivery system using Caco-2 cell culture systems. Rectal nicotine suppository formulations were prepared in semi-synthetic glyceride bases (Suppocire AM and AI, Gattefosse Inc.) by fusion method. The in vitro release of nicotine was carried out in modified USP dissolution apparatus 1. Differential scanning calorimetry (DSC) and powder X-ray diffraction were used to study the polymorphic changes if any in the formulations. An LC method was used for the assay of nicotine. The effect of bioadhesives (glyceryl monooleate (GMO), and Carbopol) on the nicotine flux was evaluated using Caco-2 cell permeability studies and Caco-2 cell viability was determined using the MTT toxicity assay. In vitro release studies indicated that the low melting AI base was superior to that of the AM base. Presence of GMO in the formulation enhanced the release of nicotine whereas Carbopol showed an opposite effect. The enhanced release of nicotine in the presence of GMO was found to be partly due to the melting point lowering effect of this compound. Caco-2 cell absorption studies showed that there was a decrease in the flux of nicotine in the presence of both the bioadhesives. The flux of the fluorescein marker which is used to study the integrity of the cell monolayers was found to be slightly higher only in the presence of 10% (w/w) Carbopol. Nicotine, Carbopol, and GMO do not have any cytotoxic effect on these cell monolayers within the concentration range used in the formulations. Rectal nicotine formulations containing bioadhesives were developed and characterized. Both in vitro release and cell culture studies have indicated that one can manipulate the nicotine release from these rectal delivery systems by incorporation of various bioadhesives or the use of different bases in the formulation. Nicotine concentration below 2% (w/v) and bioadhesive concentration below 10% (w/w) do not have any cytotoxic effect on Caco-2 cells. PMID- 10528094 TI - Systemic absorption of insulin from a Gelfoam ocular device. AB - In previous reports (Lee et al., 1997b; Lee and Yalkowsky, 1999), it has been shown that insulin, delivered by an acidified Gelfoam (absorbable gelatin sponge, USP) based ocular device, can be efficiently absorbed into the systemic circulation without the aid of an absorption enhancer. The role of acid in the enhancer-free absorption of insulin is investigated in this report. Gelfoam ocular devices containing 0.2 mg of sodium insulin prepared with either water or 10% acetic acid were evaluated in rabbits. The results suggest that a change in the Gelfoam upon treatment with acid is responsible for the efficient systemic absorption of insulin from these enhancer-free devices. PMID- 10528096 TI - Size-dependent extravasation and interstitial localization of polyethyleneglycol liposomes in solid tumor-bearing mice. AB - We have examined the size dependence of extravasation and interstitial localization of polyethyleneglycol-coated liposomes (PEG-liposomes) in the solid tumor tissue by means of electron microscopic observation. Liposomes composed of distearoyl phosphatidylcholine, cholesterol and distearoylphosphatidylethanolamine derivative of polyethyleneglycol (PEG) were prepared in various size ranges. PEG-liposomes with an average diameter of 100 200 nm showed the most prolonged circulation time and the greatest tumor accumulation in all the solid tumors employed in this experiment. Although large PEG-liposomes with a diameter of 400 nm showed a short circulation time in normal mice, the results in splenectomized mice indicated that they do have an intrinsic prolonged circulation character in vivo. However, large PEG-liposomes could not extravasate into solid tumor tissue. These results indicate that the size of liposomes is critical for extravasation. The electron microscopic observations revealed the almost exclusive engulfment of extravasated liposomes by tumor associated macrophages; very few were taken up by tumor cells. PMID- 10528095 TI - Solubility profiles of some isoxazolyl-naphthoquinone derivatives. AB - Water, ethanol and n-hexane solubility and pH-solubility behavior of a homologous series of isoxazolyl-naphthoquinone derivatives, which exhibit important biological activity, were studied. Their pK(a) values were determined spectrophotometrically as well as from their pH-solubility profiles. Also, the molar aqueous solubility of these compounds was estimated with the equations developed by Yalkowsky and Valvani using the data of their melting points and octanol/water partition coefficients. PMID- 10528098 TI - Preparation and evaluation of liposomal formulations of tropicamide for ocular delivery. AB - Tropicamide, a mydriatic, cycloplegic drug was entrapped in liposomes. Liposomes were investigated by laser counting studies, transmission electron microscopy and differential scanning calorimetry for characterization. The precorneal clearance of liposomes was compared with solution by gamma-scintigraphy in the rabbit. The neutral liposomes failed to demonstrate significant enhancement in precorneal retention in comparison with aqueous solution. The potential of liposomes as an ophthalmic drug delivery system was investigated by comparing pupil dilatory effect of tropicamide by topical instillation, in the rabbit eye, of the solution and various drug-loaded liposomal forms, i.e. neutral liposomes, positively charged liposomes and neutral liposomes dispersed in 0.25% (w/v) polycarbophil gel. The positively charged liposomal formulation and liposomes dispersed in polycarbophil gel were found to be more effective than neutral liposomal dispersion when data were statistically treated at the 5% level of significance. PMID- 10528097 TI - A comparison of the bronchodilator effect of salbutamol inhaled via Turbuhaler as two consecutive doses or as two divided doses at different time intervals. AB - The aim of the study was to compare the bronchodilatory effect of 2x50 microg salbutamol inhaled via Turbuhaler(R) as two consecutive doses compared with two divided doses (50+50 microg) at different time intervals. The study was of a randomized, double-blind, crossover design. The patients inhaled two doses of 50 microg salbutamol immediately after each other and with a time interval between the doses of 1.5, 3, 5, or 10 mins. Forced expiratory volume in 1 s (FEV(1)) was measured before the first inhalation and at 1, 2. 5, 4.5, 9.5, 15, 20, 30, and 60 min after the first inhalation. Seventeen asthmatic patients (8 men) with a mean age of 32 years (range: 19-49 years), mean FEV(1) of 2.9 l (range: 1.7-3.9 l) and a mean FEV(1) in percentage of predicted normal value of 77% (range: 63-91%) participated. The mean reversibility 15 min after inhaling 100 microg salbutamol from pMDI was 23% (range: 16-35%). The mean maximum increase in FEV(1) from baseline ranged between 18.6% (consecutive doses) and 21.2% (1.5 min between doses), corresponding to a difference between the treatments of 0.06 l. There were no significant differences in maximum FEV(1) or time to reach maximum FEV(1) between the treatments. We were not able to show any clinically relevant differences in maximal bronchodilating effect, assessed as FEV(1), in stable asthmatics, when therapeutic doses of salbutamol were given via Turbuhaler either as two consecutive doses or as two divided doses separated by different time intervals. PMID- 10528100 TI - Investigation of the phase behaviour of systems containing lecithin and 2-acyl lysolecithin derivatives. AB - A series of modified phospholipids (m-PC) possessing different acyl chains in position 2, from butanoyl to hexadecanoyl, were prepared by partial synthesis from soybean lysolecithin. They were used with soybean lecithin to construct phase diagrams containing ethanol as cosolvent, water and medium chain triglycerides (MCT) or isopropyl myristate (IPM) as oils. The weight ratios lecithin:m-PC and surfactants:ethanol were kept constant at 1:1. The results indicate that the m-PCs have a strong effect on the microemulsion (L) and liquid crystalline (LC) domains in the water-rich/oil-poor part of the phase diagrams, although all diagrams correspond to a single lecithin:m-PC ratio. On decreasing the acyl chain length, and thus increasing the hydrophilicity of the surfactant, there was a corresponding increase in the L area, which moved towards the aqueous corner of the phase diagrams. The LC phase was detected only in the presence of the hexadecanoyl derivative for the systems containing MCT, and it was not detected only in the presence of the butanoyl derivative for the systems containing IPM. The use of a second hydrophilic surfactant to adjust the packing properties of the lecithin-alcohol systems, and/or to increase the fluidity of the surfactant film, increased the region of existence of the isotropic systems. This may be of importance in the formulation of drug delivery systems, especially those which are diluted by biological fluids upon administration. PMID- 10528099 TI - Screening of high shear mixer melt granulation process variables using an asymmetrical factorial design. AB - The effects of process conditions on the granulometric characteristics of a placebo formulation prepared in a 10 l high shear mixer by single-step melt granulation were studied. The factors under investigation were: binder grade, mixer load, presence of the deflector (all of analysed at two levels), binder concentration, impeller speed, massing time, type of impeller blades (these four at three levels) and jacket temperature (considered at four levels). Two granule characteristics were analysed: the geometric mean diameter and the percentage of particles finer than 315 microm. In order to screen simultaneously the above mentioned factor levels, an asymmetrical factorial design was adopted, which allowed the reduction in the number of runs from 2592 to 25. Additionally, this technique permitted the selection of the factor levels which have the major 'weight' on the two granule characteristics under study. Two additional trials were performed to attest the screening validity. PMID- 10528101 TI - Application of viscometry and solubility parameters in miconazole patches development. AB - Nine binary mixtures of seven different methacrylic copolymer systems (Plastoid((R)) E 35 L (PLE) and Plastoid((R)) L 50 (PLL); Eudragit((R)) (Eu) NE, RL, RS, L, S) were tested as components of monolayer patches containing miconazole. Only three mixtures (PLE:EuNE, PLE:EuRL and PLE:EuRS) were suitable for the preparation of placebo matrices. Miconazole patches with good technological characteristics were obtained by using mixtures of PLE:EuNE and PLE:EuRL. The in vitro miconazole release rate from the two patches and from the patch prepared using only PLE were significantly different. The amounts of drug released in 24 h were quite satisfactory. A mathematical model based on capillary viscometry data was used for the evaluation of interactions between copolymers. This was useful to predict and understand the mechanisms related to the instability of the prepared mixture. The solubility parameters of the drug and of the matrix were also calculated. Miconazole release was faster when the difference between the solubility parameters of the matrix and of the drug was higher. A relationship between miconazole release rate and the difference of drug and matrix solubility parameters was found. Therefore, the solubility parameter could be applied in formulation studies of patches. PMID- 10528102 TI - Stability and compatibility of an aerosol mixture including N-acetylcysteine, netilmicin and betamethasone. AB - The physicochemical stability and the compatibility between N-acetylcysteine (1 g/5 ml), betamethasone (4 mg/1 ml) and netilmicin (100 mg/1 ml) were studied at room temperature (25+/-2 degrees C) over 1 h. During this study, drug concentrations were measured using three separate HPLC methods with UV detection at t=0, 5, 10, 20, 30, and 60 min. The pH of the mixture was determined. Degradation products of the drugs were assayed using HPLC. This study demonstrates the stability and compatibility of the mixture over 1 h at room temperature. The pinkish non-remnant coloration observed when pouring N acetylcysteine into a recipient has no effect on the stability of the drug. PMID- 10528103 TI - Stability of rifampicin in dissolution medium in presence of isoniazid. AB - Rifampicin (RIF) hydrolyzes in acidic medium to form insoluble and poorly absorbed 3-Formyl rifamycin SV (3-FRSV). This study describes development of two principally different methods, Dual Wavelength UV-Vis. spectrophotometry (DW spectrophotometry) and HPTLC, to determine 3-FRSV in presence of RIF. Using DW spectrophotometry, RIF was estimated by using wavelengths 475.0 and 507.0 nm and 3-FRSV was estimated using 457.0 and 492.0 nm. In HPTLC method, a mixture of chloroform:methanol:water (80:20:2.5 v/v) was used as the mobile phase to resolve 3-FRSV from RIF and 3-FRSV was quantified at 333 nm. The linearity range for 3 FRSV was 2-10 microg/ml and 50-250 ng/spot for DW spectrophotometric method and HPTLC method, respectively, and 5-50 microg/ml for RIF using DW spectrophotometric method. Both the methods were found to be specific, accurate and reproducible. The proposed methods were successfully applied to determine the rate of degradation of RIF to 3-FRSV in dissolution medium (0.1 N HCl) and also in presence of isoniazid (INH). The rate of degradation of RIF in presence of INH was almost two times more than that of RIF alone. These methods were utilized to study the stability of RIF in market formulations of RIF and RIF with INH in dissolution medium. It has been observed that RIF degrades by 12.4% to form 3 FRSV (RIF formulations) while in presence of INH the degradation is catalyzed to about 21.5% (RIF+INH formulations), in 45 min. Thus, lower concentration of RIF may be available for absorption leading to poor bioavailability of RIF from combination dosage forms (RIF+INH) as compared to formulations containing only RIF. It is proposed that specific analytical method should be used to measure RIF in presence of 3-FRSV in a dissolution study. PMID- 10528104 TI - Mature and developing visual system of Ceratitis capitata (Diptera, Tephritidae): histochemical evidence of nitric oxide synthase in the wild type and the white eye mutant strains. AB - Nitric oxide (NO) is acknowledged as a messenger molecule in the nervous system. It has a role in the modulation of the chemosensory information and seems implicated also in visual processes and visually guided behaviour of some insects. In the present study, we used two different strains of the medfly Ceratitis capitata (Diptera, Tephritidae), a wild type eye colour and a white eye mutant line, as models to clarify the involvement of NO in the mature and developing visual system. The comparison between the pattern of enzyme histochemical localization of NO synthase (NOS), through NADPH diaphorase (NADPHd) staining, in the optic lobes of the two strains revealed for adults a stronger intensity of reaction in all the neuropiles and the sub-retinic monopolar cell layer of the wild type flies, with respect to the white eye mutant correspondent areas. Anti-NOS immunocytochemistry correlated with these results, underlying reactivity both in fine fibres and varicosities and in cell bodies and supporting the idea of presence of NOS also in the retina of the medfly optic lobes. NADPHd reactivity was present in the first developmental stages of the white eye mutant also, but at lower intensity than wild type, and it decreased in some areas during the transition to adult fly stage both in the wild type and in the white eye mutant. All these observations together indicate that changes in the NO system of C. capitata could be related to the visual information processing, when the visual response or discrimination are altered. Furthermore, NO may be involved in the establishment of the retinal projection pattern and in the control of optic lobes morphogenesis. PMID- 10528105 TI - Selective opioid delta agonists elicit antinociceptive supraspinal/spinal synergy in the rat. AB - A multiplicative antinociceptive interaction of morphine activity at supraspinal and spinal sites has been clearly established and is thought to be responsible, in part, for the clinical utility of this compound in normal dose-ranges. While synergistic actions of mu-opioid receptor agonists have been shown, it is unclear whether a similar interaction exists for opioid agonists acting via delta-opioid receptors. Responses to acute nociception were determined with the 52 degrees C hot plate, 52 degrees C warm-water tail-flick and the Hargreaves paw-withdrawal tests. The peptidic opioid delta(1) agonist [D-Pen(2),D-Pen(5)]enkephalin (DPDPE) or delta(2) agonist [D-Ala(2),Glu(4)]deltorphin (DELT) were given into the rostral-ventral medulla (RVM), intrathecally (i.th.) or simultaneously into both the RVM and i.th. (1:1 fixed ratio). Both of the opioid delta agonists produced dose-dependent antinociception in all tests. With the exception of DPDPE in the hot plate test, isobolographic analysis revealed that the supraspinal/spinal antinociceptive interaction for both DPDPE and DELT were synergistic in all nociceptive tests. These data suggest that opioid delta agonists exert a multiplicative antinociceptive interaction between supraspinal and spinal sites to acute noxious stimuli and suggest possibility that compounds acting through delta-opioid receptors may have sufficient potency for eventual clinical application. PMID- 10528106 TI - Evaluation of lipid peroxidation, cathepsin L and acid phosphatase activities in experimental brain ischemia-reperfusion. AB - This investigation was conducted in rat brain tissues to elucidate the free radical induced cellular and subcellular membrane injuries in two different depth of global ischemia. Global moderate (penumbral) ischemia was performed on rat brains by bilateral vertebral arteries cauterization and temporary occlusion of the bilateral carotid arteries. Global severe ischemia was produced by a neck tourniquet in addition to four vessel occlusion. Somatosensory evoked potentials (SSEPs) were used as a feed back parameter to monitor electrophysiologically the ischemia. At the end of ischemic insult (0 min reperfusion) or various reperfusion periods (20, 60 and 240 min), all rats were decapitated and brains were frozen in liquid nitrogen. The brain tissues were prepared for the determination of cathepsin L (CL) and acid phosphatase (AP) activities in the supernatant (cytosolic) fraction (SF) and the fraction enriched with lysosomes (FEL). Further the level of thiobarbituric acid reactive substances (TBARS) of lipid peroxidation was assessed by the spectrophotometric methods. Severe ischemia-reperfusion was accompanied by a significant increase in TBARS levels and the SF/FEL ratio for CL and AP activities compared to the sham operated group and the concurrent reperfusion groups of moderate ischemia (p<0.05). There were no significant differences between the sham operated and moderate ischemia reperfusion groups for the same parameters. Our data clearly demonstrate that; in rat brain although severe ischemia-reperfusion causes lipid peroxidation in cellular membranes and redistribution of lysosomal enzymes from lysosomes to cytoplasm due to lysosomal membrane injury, there are no changes in lysosomal membrane stability in moderate ischemia-reperfusion. PMID- 10528108 TI - The period of latency before a muscle receptor generates an action potential as a response to a muscle stretch. AB - Six primary (Ia) and seven secondary (II) muscle spindle afferents and eight Golgi tendon organ afferents (Ib) from the tibial anterior muscle of the cat, recorded at the dorsal roots, were subjected to a sinusoidal stretch of the host muscle, the frequency of which increased linearly from 2 to 80 Hz over four different lengths of time. Both the amplitude of the sinusoidal stretch and the prestretch of the muscle were varied. The phase of the action potentials was determined. The phase of the action potential, driven 1:1, increased linearly with frequency. From the gradient of the phase of this action potential the muscle-muscle receptor latency was determined, i.e., the period of latency between the stretch of the muscle and the occurrence of the action potential at the muscle nerve where it enters the muscle. The muscle-muscle receptor latency had values lying between 3 and 8 ms: it was dependent on the experimental parameters and became shorter as the conduction velocity of the afferent fiber increased. In three experiments the muscle latency was determined, i.e., the period of latency before the stretch was transferred from the tendon of the muscle to the proximal third of the muscle belly. The muscle was stretched sinusoidally under the same varying parameters as given above. The length changes occurring in the proximal third of the muscle were measured with a piezo element. The muscle latency was determined from the slope of the phase of the zero points of the sinusoidal piezo length changes; the phase increases linearly with frequency. The muscle latency had values lying between 6 and 15 ms: it was dependent on the experimental parameters. The muscle spindle latency, i.e., the period of latency between the stretch of the polar parts of the intrafusal muscle fibers and the recording of the action potentials from the spindle nerve near the spindle capsule, was determined from 5 Ia fibers and 1 II fiber of isolated muscle spindles. The isolated muscle spindle was stretched under the same varying parameters as given above. The muscle spindle latency was determined from the slope of the phase of the phase-locked action potential. The muscle spindle latency as measured by our method proved to be 0 ms. The latencies of the three elements and their dependence on the experimental parameters are discussed in the light of the transfer properties of the muscle and the muscle receptors. PMID- 10528107 TI - Calbindin 28kD and parvalbumin immunoreactive neurons receive different patterns of synaptic input in the cat superior colliculus. AB - Recent evidence suggests that neurons containing the calcium binding proteins calbindin 28kD (CB) and parvalbumin (PV) have differing distributions which match respectively the distribution of W and Y retinal ganglion cell inputs to the cat superior colliculus (SC). In this study we have used electron microscope immunocytochemistry to study directly the synaptic inputs to neurons containing CB and PV. Aspiration lesions of areas 17-18 of visual cortex were made 4 days prior to sacrifice in order to identify degenerating cortical terminals (CT). Retinal terminals (RTs) were identified by their characteristic morphology including large round synaptic vesicles and pale mitochondria. We photographed RTs and CTs that were in contact with immunoreactive profiles sampled in both the superficial gray and optic layers (ol) of SC. CB immunoreactive (ir) dendrites were usually of small to medium caliber and were found to receive synaptic input from RTs. These RTs were all small profiles forming a single synaptic contact with asymmetric densifications. CBir profiles also received other synaptic input, including from terminals with dark mitochondria that contained flattened synaptic vesicles (F profiles). No CBir dendrites were found to receive CT input even though degenerating CTs were found in the vicinity of CBir profiles. By contrast, both RT and CT were found to contact PVir dendrites. RT terminals contacting PVir dendrites were both small and larger profiles with round synaptic vesicles and asymmetric synaptic densifications. CT were undergoing electron dense degeneration but still sometimes formed asymmetric synaptic densifications with PV neurons. PV cells also received F profile synaptic input. We conclude that CB neurons receive small RT synapses that are probably of W origin, while PV neurons receive both RT and CT synapses which are likely related to the Y pathway. PMID- 10528109 TI - Toxicity induced by a polyglutamated folate analog is attenuated by NAALADase inhibition. AB - Folates have been shown to be neurotoxic and convulsive. Endogenously, folates exist in the brain in a polyglutamated form with 1-7 terminal glutamates (approx. 1 microM). The brain enzyme N-acetylated alpha-linked acidic dipeptidase (NAALADase) has been shown to remove sequentially the gamma-linked glutamates from folic acid polyglutamates. We report that, at high concentrations (300 microM-30 mM), a folic acid hexaglutamate analog is dose-dependently toxic to dissociated rat cortical cultures and that this toxicity is reversed by 2-PMPA, a potent and selective NAALADase inhibitor. These data suggest a new mechanism for folic acid toxicity. PMID- 10528110 TI - Cellular co-localization of phosphorylated tau- and NACP/alpha-synuclein-epitopes in lewy bodies in sporadic Parkinson's disease and in dementia with Lewy bodies. AB - The precursor of the non-Abeta-component of Alzheimer's disease (AD) amyloid (NACP, alpha-synuclein) aggregates into insoluble filaments of Lewy bodies (LBs) in Parkinson's disease (PD) and dementia with LBs (DLB). The microtubule associated protein tau is an integral component of filaments of neurofibrillary tangles (NFTs). NFTs are occasionally found in brains of PD and DLB; however, the presence of NFTs or tau-epitopes within LB-containing neurons is rare. Double immunofluorescence study and peroxidase-immunohistochemical study in serial sections, performed to examine the co-localization of tau- and NACP-epitopes in the brainstem of PD and DLB, demonstrated that four different epitopes of tau including phosphorylation-dependent and independent ones were present in a minority of LBs, but more often than previously considered. A tau (tau2)-epitope was localized to filaments in the outer layers of brainstem-type LBs by immunoelectron microscopy. Therefore, we conclude that tau is incorporated into filaments in certain LBs. Extensive investigation has enabled us to classify this co-localization into four types: type 1, LBs with ring-shaped tau immunoreactivity; type 2, LBs surrounded by NFTs; type 3, NACP- and tau immunoreactive filamentous and granular masses; and type 4, NACP- and tau immunoreactive dystrophic neurites. This study raises a new question whether aggregation and hyperphosphorylation of tau in PD and DLB are triggered by the collapse of intraneuronal organization of microtubules due to NACP-filament aggregation in neuronal perikarya and axons. PMID- 10528111 TI - GABA immunoreactivity in mouse barrel field after aversive and appetitive classical conditioning training involving facial vibrissae. AB - We have previously reported that a classical conditioning paradigm involving stimulation of a row of facial vibrissae produced an expansion of the cortical representation of the "trained row", labeled with 2-deoxyglucose (2DG), in layer IV of the barrel field. The present study has examined the pattern of GABA immunoreactivity (GABA-IR) in the cortical representation of row B of the facial vibrissae after (i) 3 days of aversive training, and (ii) 2 months of appetitive training, where stimulation of row B of vibrissae on one side of the snout was used as a conditioned stimulus. The most notable observation was a greater density of GABA-IR cells concentrated in the hollows of the "trained row" B barrels compared to the hollows in the barrel field of the opposite hemisphere in the same mouse. After aversive training, we noted a 2-fold increase in the density of GABA-IR neurons in the hollows of row B; after reward training, the increase amounted to 49%. In contrast, GABA-IR was unchanged in the control groups, which received only stimulation of vibrissae without the unconditioned stimulus. The classification of labeled neurons according to size revealed that the increase in density of GABA-IR neurons was confined to the small (12-15 microm) diameter group. We concluded that the GABAergic system undergoes up regulation, after both associative learning paradigms, and that the population of small, GABAergic neurons plays an active role in use-dependent plasticity. PMID- 10528112 TI - Analysis of MEG signals of spreading cortical depression with propagation constrained to a rectangular cortical strip. I. Lissencephalic rabbit model. AB - Magnetic fields arising from the rabbit cortex during spreading cortical depression (SCD) were measured in order to study the currents in the neocortex during SCD. SCD was constrained to propagate in a rectangular cortical strip perpendicular to the midline. This simplified in vivo cortical preparation enabled us to correlate magnetoencephalographic (MEG) signals to their underlying currents within the cortical strip. The propagation of SCD was monitored with an array of electrodes placed along the strip. The propagation speed for SCD in the lissencephalic rabbit brain was 3. 5+/-0.3 mm/min (mean+/-S.E.M., n=14). Slow, quasi-dc, MEG signals were observed as the SCD entered into the longitudinal fissure. The currents giving rise to the MEG signals were perpendicular to the cortical surface and directed from the surface to deeper layers of the cortex. A distributed dipolar source model was used to relate the data to the underlying cortical current. The moment of the single equivalent current dipole source was 38+/-9 nA-m (n=17). This study clarified the nature of the cortical currents during SCD in a lissencephalic in vivo preparation. PMID- 10528113 TI - Analysis of MEG signals of spreading cortical depression with propagation constrained to a rectangular cortical strip. II. Gyrencephalic swine model. AB - Currents produced during spreading cortical depression (SCD) in a gyrencephalic species (swine) were studied with magnetoencephalography (MEG) and electrocorticography (ECoG). SCD, initiated using electrical stimulation of the cortex, was constrained to propagate within a rectangular cortical strip in order to simplify the interpretation of the underlying currents. The ECoG signals monitored along the strip revealed that SCD propagated from an initiation site on the gyrus at a rate of 7.9+/-3.2 mm/min (n=23), entered the deep coronal sulcus and in most cases emerged from the other side of the sulcus, continuing to propagate across the next gyrus at a rate of 5.9+/-2.7 mm/min (n=22). The apparent propagation velocity within the sulcus was reduced to 1.7+/-0.8 mm/min (n=21). Strong MEG signals were observed as SCD entered the sulcus. The direction of magnetic field was opposite for SCD's on opposite banks of the sulcus. The currents were directed from a superficial layer to deeper layers of the cortex. The characteristics of SCD and associated MEG patterns from a gyrencephalic species may be similar to those in human patients during migraine aura. PMID- 10528114 TI - Apolipoprotein E and beta-amyloid levels in the hippocampus and frontal cortex of Alzheimer's disease subjects are disease-related and apolipoprotein E genotype dependent. AB - The epsilon4 allele of apolipoprotein E (apoE) is associated with increased risk for the development of Alzheimer's disease (AD), possibly due to interactions with the beta-amyloid (Abeta) protein. The mechanism by which these two proteins are linked to AD is still unclear. To further assess their potential relationship with the disease, we have determined levels of apoE and Abeta isoforms from three brain regions of neuropathologically confirmed AD and non-AD tissue. In two brain regions affected by AD neuropathology, the hippocampus and frontal cortex, apoE levels were found to be decreased while Abeta(1-40) levels were increased. Levels of apoE were unchanged in AD cerebellum. Furthermore, levels of apoE and Abeta(1 40) were found to be apoE genotype dependent, with lowest levels of apoE and highest levels of Abeta(1-40) occurring in epsilon4 allele carriers. These results suggest that reduction in apoE levels may give rise to increased deposition of amyloid peptides in AD brain. PMID- 10528115 TI - Cellular localization of dopamine-releasing protein (DARP) in rat C6 glioma and primary mesencephalic cell cultures. AB - Dopamine-releasing protein (DARP) is a multisubunit protein shown to have dramatic effects on development, recovery, and function of the rat catecholaminergic (CA) system. This study details efforts to determine if glial cells are responsible for the production of DARP in the central nervous system (CNS). Enzyme-linked immunosorbent assays (ELISA), Western blotting, and immunocytochemical techniques were employed to measure DARP levels and identify DARP immunoreactive proteins in rat C6 glioma cells and medium, respectively. ELISA analysis of serum-free C6 culture media revealed a maximal concentration of DARP by culture day 1. However, ELISA analysis of C6 cultures grown in F 12K/serum medium revealed that maximal levels of DARP were detected on culture day 6 with a 108% increase in DARP immunoreactivity from culture day 1. These values were determined using a polyclonal antibody generated against DARP-36aa (anti-DARP-36aa), a synthetic peptide with dopamine (DA) releasing activity, and anti-DARP B9-B10, a monoclonal antibody generated against partially purified DARP. Western blot analysis revealed that anti-DARP B9-B10 recognized proteins of approximately 60, 50, and 45 kDa in C6 cell homogenates while anti-DARP-36aa had immunoreactivity with the 60-kDa protein alone. Immunocytochemical studies demonstrated that anti-DARP-36aa and anti-DARP B9-B10 had strong immunoreactivity with proteins throughout the cytosol and in several processes of C6 cells. These results reveal that DARP is detected in glioma cells and secreted in a time dependent fashion during culture. Primary rat mesencephalic cultures were also examined using immunocytochemistry. Incubation with DARP antibodies and antisera against glial fibrillary acidic protein (GFAP) revealed that DARP and GFAP immunoreactivity co-localized in primary mesencephalic cultures. However, the majority of DARP immunoreactivity was localized to cells without GFAP staining. These findings reveal that DARP is detected in astrocytes although the majority of DARP immunoreactivity is found in non-astrocyte type cells. PMID- 10528116 TI - A rat model of endothelin-3-induced middle cerebral artery occlusion with controlled reperfusion. AB - Surge hyperemia and mechanical damage to the cerebrovascular endothelium may serve to exacerbate the neuropathological outcome in animal models of focal cerebral ischemia. We have modified an existing model of endothelin-1-induced middle cerebral artery (MCA) occlusion to enable controlled reperfusion without damage to the cerebral vasculature. Endothelin-1 (ET-1) and endothelin-3 (ET-3) were injected via a double-injection cannula into brain parenchyma adjacent to the MCA of anesthetized rats to produce focal cerebral ischemia. ET-1 and ET-3 produced large ischemic lesions that were restricted to those cortical and subcortical structures supplied by the MCA. The volume of ischemic damage produced by 100 pmol of ET-1 and ET-3 was similar. The endothelin-A (ET(A)) receptor antagonist FR139317 (3 or 30 nmol) injected 10 min after ET-1 did not significantly alter the volume of damage. By contrast, the lesion produced by ET 3 was completely inhibited by FR139317 at the 10 min time-point. FR139317 partially attenuated the ET-3-induced lesion when administered 30 min post occlusion, but injection 90 min following ET-3 produced a lesion not different to that produced by ET-3 alone. These findings were supported by laser Doppler flowmetry which determined FR139317 induces reperfusion when injected 10 or 90 min following ET-3. ET-3-induced MCA occlusion is therefore amenable to reversal by the ET(A) receptor antagonist FR139317, and this model may offer a means to investigate the neuropathology of reperfusion without the procedure-related artifacts associated with some reperfusion models. PMID- 10528117 TI - Induction of catecholamine synthesis in human neuroblastoma cells by replication inhibitors and sodium butyrate. AB - Various inhibitors of DNA synthesis induced neurite extension in human neuroblastoma cells. In order to clarify morphology-function relationship in differentiation of neuroblastoma cells, the effect of the replication inhibitors on inducibility of catecholamine synthesis was examined. The reagents alone did not affect production of dopamine and noradrenaline, but joint administration of each inhibitor and sodium butyrate considerably promoted the catecholamine synthesis, without additional change in neurite profile. Although inactive in neurite extension, sodium butyrate was moderately active in catecholamine production. The promoting effect of thymidine (or hydroxyurea) and sodium butyrate was repealed by alpha-amanitin, actinomycin D or cycloheximide. PMID- 10528118 TI - Cyclooxygenase-2 selective inhibitors aggravate kainic acid induced seizure and neuronal cell death in the hippocampus. AB - Cyclooxygenase-2 (COX-2) in the brain is expressed constitutively and also increased in pathological conditions such as seizure, cerebral ischemia, and inflammation. This study examined the role of COX-2 in kainic acid-induced seizure and in the following neuronal death by using selective inhibitors. Systemic kainate injection (50 mg/kg; i.p.) in mice evoked seizure within 15 min and led to 29% mortality within 2 h. TUNEL-positive neuronal death peaked at 3 days after injection and was prominent in CA(3a) regions of the hippocampus. NS 398 or celecoxib (10 mg/kg, COX-2 selective inhibitor) and indomethacin (5 mg/kg, nonselective inhibitor) exaggerated kainic acid-induced seizure activity and mortality. COX-2 selective inhibitors induced the seizure at earlier onset and more severe mortality within the first hour than indomethacin and aspirin. NS-398 also aggravated kainic acid-induced TUNEL positive neuronal death and decreased Cresyl violet stained viable neurons, and extended lesions to CA(1) and CA(3b). Kainic acid increased the levels of PGD(2), PGF(2a) and PG E(2) in the hippocampus immediately after injection. Indomethacin attenuated the production of basal and kainic acid-induced prostaglandins. In contrast, NS-398 failed to reduce until the first 30 min after kainic acid injection, during which the animals were severely seizured. It has been challenged the endogenous PGs might have anticonvulsant properties. Thus, COX-2 selective inhibitor, including nonselective inhibitor such as indomethacin, aggravated kainic acid-induced seizure activity and the following hippocampal neuronal death even with variable prostaglandin levels. PMID- 10528119 TI - Effects of restraint stress on alpha(1) adrenoceptor mRNA expression in the hypothalamus and midbrain of the rat. AB - We examined the effects of restraint stress on alpha(1) adrenoceptor mRNA expression in the rat brain using reverse transcriptase-polymerase chain reaction (RT-PCR). After rats had been restrained for 10, 30, 60, 120 or 240 min, the hypothalamus and midbrain were removed immediately and alpha(1) adrenoceptor mRNA levels in these regions were determined by RT-PCR. Blood samples were also collected for simultaneous measurement of serum adrenocorticotropic hormone (ACTH) and corticosterone. Restraint stress resulted in a variety of changes in the hypothalamus and midbrain. In the hypothalamus, 30 and 60 min of stress resulted in a significant fall in the level of alpha(1) adrenoceptor mRNA relative to the control. This was associated with a rise in serum ACTH and corticosterone. In the midbrain, significant elevation of alpha(1) adrenoceptor mRNA was noted after 60, 120 and 240 min of restraint stress. Our findings indicated that the influence of restraint stress on alpha(1) adrenoceptor mRNA level in the hypothalamus is different to that of the midbrain region in rats. PMID- 10528120 TI - Developmental and age-related changes of dopamine transporter, and dopamine D1 and D2 receptors in human basal ganglia. AB - The developmental and age-related changes of the dopamine transporter (DAT), and the dopamine D1 and D2 receptor (D1R and D2R) subtypes were investigated in basal ganglia (BG) of human brain. DAT immunostaining was mainly observed in the neuropil, neurons, and glia of the striatum. The DAT-positive neuropil was detectable at 32 GW, a peak being reached at 9-10 years of age, with a decrease to 50-63 years of age. The developmental pattern of DAT immunoreactivity in neuron was similar to that of the neuropil. DAT-positive glia were observed in the BG at 32 GW, which increased slightly at 38-40 GW, and then did not obviously change until 6-8 months after birth. D2R-positive neurons were clearly observed at 19 GW, a peak being reached at 32 GW and 1-3 months of age in the globus pallidus and striatum, respectively, with a decrease after 9-10 years of age. D1R was expressed as early as D2R, but decreased after 6-8 months. Our results suggest that D1R and D2R expression is an intrinsic property of striatal neurons and is independent of dopaminergic innervation. D1R may play a more important role in neuronal maturation of the BG than D2R. D2R may be closely correlated with late neuronal development. The higher expression of DAT during adolescence may be related to function of the BG which learns complex behavioral patterns. The significance of the age-related decreases in DAT, D1R and D2R in the BG remains to be further investigated. PMID- 10528122 TI - "Gating" of human short-latency somatosensory evoked cortical responses during execution of movement. A high resolution electroencephalography study. AB - The present study aimed at investigating gating of median nerve somatosensory evoked cortical responses (SECRs), estimated during executed continuous complex ipsilateral and contralateral sequential finger movements. SECRs were modeled with an advanced high resolution electroencephalography technology that dramatically improved spatial details of the scalp recorded somatosensory evoked potentials. Integration with magnetic resonance brain images allowed us to localize different SECRs within cortical areas. The working hypothesis was that the gating effects were time varying and could differently influence SECRs. Maximum statistically significant (p<0. 01) time-varying gating (magnitude reduction) of the short-latency SECRs modeled in the contralateral primary motor and somatosensory and supplementary motor areas was computed during the executed ipsilateral movement. The gating effects were stronger on the modeled SECRs peaking 30-45 ms (N30-P30, N32, P45-N45) than 20-26 ms (P20-N20, P22, N26) post stimulus. Furthermore, the modeled SECRs peaking 30 ms post-stimulus (N30-P30) were significantly increased in magnitude during the executed contralateral movement. These results may delineate a distributed cortical sensorimotor system responsible for the gating effects on SECRs. This system would be able to modulate activity of SECR generators, based on the integration of afferent somatosensory inputs from the stimulated nerve with outputs related to the movement execution. PMID- 10528121 TI - Heterogeneous expression of voltage-gated potassium channels of the shaker family (Kv1) in oligodendrocyte progenitors. AB - Outwardly rectifying K(+) channels determine the membrane conductance and influence the proliferation rate of glial progenitor cells. To analyze the molecular identity and the functional role of K(+) channels in glial progenitors of mouse brain, expression of shaker-type Kv1 genes was studied at three levels: (1) presence of Kv1 mRNAs, (2) biosynthesis of channel proteins and (3) electrophysiological and pharmacological properties of K(+) currents. mRNA expression of Kv1.1 to Kv1.6 genes was studied by single-cell reverse transcription-mediated polymerase chain reaction (RT-PCR) using degenerate primers to amplify the six Kv1 transcripts. Most cells expressed several mRNA combinations simultaneously. In more than half of the cells, messages for Kv1.2, Kv1.5 and Kv1.6 were found, while Kv1.1, Kv1.3 and Kv1.4 were detected in only a minority of cells. In contrast, at the level of protein expression - employing immunocytochemistry with subtype-specific antibodies - Kv1. 2 and Kv1.3 were undetectable (<2%), while almost all cells expressed Kv1.4 (85%), Kv1.5 (99%) and Kv1.6 (99%). Kv1.1 was present in a minor cell population (10%). Functional contribution of Kv1 proteins to progenitor membrane conductance was determined by analyzing the voltage-dependence of K(+) current activation and inactivation as well as their current sensitivities to the subtype-preferring blockers and toxins tetraethylammonium (TEA), 4-aminopyridine (4-AP), charybdotoxin (CTX), alpha dendrotoxin (DTX) and mast-cell degranulating peptide (MCDP). From these results, it is concluded: first, glial progenitor cells can express all transcripts of the six Kv1 genes, but do not express all proteins; second, Kv1.4, Kv1.5 and Kv1.6 proteins are most abundant and were found in the majority of cells; and third, K(+) currents flow predominantly either through heteromeric channel complexes or through homomeric Kv1.5 ion pores, but not through homomeric Kv1.4 or Kv1.6 channels. PMID- 10528123 TI - Comparison of brain metabolic activity patterns induced by ketamine, MK-801 and amphetamine in rats: support for NMDA receptor involvement in responses to subanesthetic dose of ketamine. AB - Subanesthetic doses of NMDA receptor antagonists induce positive, negative and cognitive schizophrenia-like symptoms in healthy humans and precipitate psychotic reactions in stabilized schizophrenic patients. These findings suggest that defining neurobiologic effects induced by NMDA antagonists could guide the formulation of experimental models relevant to the pathophysiology of schizophrenia and antipsychotic drug action. Accordingly, the effects of subanesthetic doses of the non-competitive NMDA antagonists ketamine and MK-801 were examined on regional brain [14C]-2-deoxyglucose (2-DG) uptake in rats. The effects of these drugs were compared to those of amphetamine, in order to assess the potential role of generalized behavioral arousal, motor activity and dopamine release in brain metabolic responses to the NMDA antagonists. Subanesthetic doses of MK-801 and ketamine induced identical alterations in patterns of 2-DG uptake. The most pronounced increases in 2-DG for both NMDA antagonists were in the hippocampal formation and limbic cortical regions. By contrast, amphetamine treatment did not increase 2-DG uptake in these regions. In isocortical regions, ketamine and MK-801 reduced uptake in layers 3 and 4, creating a striking shift in the laminar pattern of 2-DG uptake in comparison to control conditions. After amphetamine, the fundamental laminar pattern of isocortical labeling was similar to saline-treated rats. Administration of ketamine and MK-801 decreased 2-DG uptake in the medial geniculate and inferior colliculus, whereas amphetamine tended to increase uptake in these regions. Since ketamine induced similar effects on regional 2-DG uptake as observed for the selective antagonists MK-801, the effects of ketamine are likely related to NMDA antagonistic properties of the drug. The distinct differences in brain 2-DG uptake induced by amphetamine and NMDA antagonists indicate that generalized behavioral arousal, and increased locomotor activity mediated by dopamine release, are not sufficient to account for the alterations in brain metabolic patterns induced by ketamine and MK-801. Thus, the dramatic alteration in regional 2-DG uptake induced by ketamine and MK 801 reflects a state selectively induced by reduced NMDA receptor function. PMID- 10528124 TI - Influence of dietary fats on c-Fos-like immunoreactivity in mouse hypothalamus. AB - The hypothalamus is a brain region of major importance in regulation of energy balance via autonomic nervous control of both intake and expenditure. There is substantial evidence that diets high in saturated fats lead to obesity while diets equally high in polyunsaturated fats (PUFAs) do not. Using c-Fos as a marker, this study aimed to investigate hypothalamic neuronal response in mice fed high fat diets (58% of calories as fat) emphasising saturated, n-3 or n-6 polyunsaturated fatty acids, or a low fat (10% of calories) diet over periods of 1 and 7 weeks. In addition, a 4-week "reversal" intervention with n-3 polyunsaturated or low fat diet was undertaken in saturated fat-fed mice. Food intake and body weight were measured over the feeding periods. At 1, 7 and 11 weeks mice were killed, epididymal fat pad were weighed and brains were removed for quantitation of hypothalamic c-Fos-like immunoreactive (FLI) neurons. Weight gain, and epididymal fat pad weight, were highest on the saturated fat diet and lowest on the n-3 diet despite similar food intakes (epididymal fat weight at week 7: saturated fat, 622+/-48 mg; n-6 fat 423+/-69; low fat 387+/-10, n-3 fat 225+/-26). Compared to a low fat diet, FLI neurons in the dorsal part of lateral hypothalamic (dLH) area was dramatically increased by saturated fat feeding (+367% at 1 week) while ventromedial hypothalamic (VMH) activity was decreased. In contrast with n-6 and n-3 feeding dLH FLI neuronal activity was unchanged but actually increased in the VMH. Paraventricular nucleus (PVN) FLI neurons increased in the high saturated group only at 7 and 11 weeks, after substantial fat accumulation. Substitution of saturated fat diet with the n-3 diet partially reversed (48%) the increase in FLI neurons in PVN of saturated fat-fed mice, while it significantly increase FLI neurons in arcuate nucleus (+400%). In summary, this study demonstrates that dietary saturated fat modulates hypothalamic neuronal activity in a pattern (high lateral, reduced ventromedial activity) consistent with its obesogenic effects. In contrast, diets equally high in PUFA (particularly of the n-3 class) neither increase adiposity nor derange the lateral/medial neuronal activity balance. PMID- 10528125 TI - Potentiation of transmitter release from NMB human neuroblastoma cells by kappa opioids is mediated by N-type voltage-dependent calcium channels. AB - The selective kappa-opioid agonist trans-(+/-)-3, 4-dichloro-N-methyl-N-[2-(1 pyrrolidinyl) cyclohexyl] benzenacetamidemethansulfonate (U50,488) potentiates both basal and depolarization-evoked [3H]dopamine release from NMB cells. The potentiation of dopamine release by U50,488 is mediated by N-type voltage dependent calcium channels since it is blocked by omega-conotoxin, and is resistant to pertussis toxin (PTX)-treatment. When the stimulation of release by U50,488 is blocked by the N-channel antagonist omega-conotoxin, an inhibitory effect on dopamine release is revealed, suggesting that stimulatory and inhibitory effects of U50,488 are exerted in parallel. PMID- 10528126 TI - Measured increase in intracellular Ca(2+) during stimulated release of endogenous glutamate from human cerebrocortical synaptosomes. AB - Presynaptic terminals (synaptosomes) prepared from guinea pig and rat cerebral cortex release endogenous glutamate in a Ca(2+)-dependent manner in response to membrane depolarisation. In the present study, synaptosomes were prepared from human cerebral cortex removed in association with temporal lobe resections in epileptic patients. The cytosolic free Ca(2+) concentration increased from 474+/ 66 before to 649+/-89 nM after 2 min depolarisation. The basal level of free cytosolic Ca(2+) is higher and the increase in response to depolarisation is more pronounced in human synaptosomes than observed in animal experiments. The Ca(2+) dependent glutamate release, estimated as the difference between total - and the Ca(2+)-independent glutamate release, increased from 0 to 5.4+/-1.9 nmol/mg protein. The released amount of glutamate is larger than reported in animal models. These results demonstrate that membrane depolarisation of synaptosomes from human brain evokes a rapid rise in cytosolic free Ca(2+) and a more prolonged rise in synaptic, Ca(2+)-dependent glutamate release. PMID- 10528128 TI - The partial agonist properties of levocabastine in neurotensin-induced analgesia. AB - The antihistaminic drug levocabastine is a ligand for the low affinity neurotensin receptor (NTS2). Its intracerebroventricular administration to mice induced a significant analgesia in the writhing test but not in the hot plate test. In the writhing test, levocabastine decreased neurotensin-induced analgesia to a level not significantly different from the effects of levocabastine alone. In the hot plate test, levocabastine had no analgesic effect but completely reversed the neurotensin-induced analgesia. Mepyramine, another antihistaminic drug, did not share these levocabastine effects. Neither levocabastine nor mepyramine modified the colonic temperature or reversed the neurotensin-induced hypothermia. Thus, levocabastine behaves as a partial agonist at neurotensin NTS2 receptors, which are involved in visceral nociception, but not at yet unidentified neurotensin receptors involved in hypothermia. PMID- 10528127 TI - The GABA(B) receptor antagonist CGP36742 improves learned helplessness in rats. AB - Effects of 3-aminopropyl-n-butyl-phosphinic acid (CGP36742), a GABA(B) receptor antagonist, in the learned helplessness paradigm were examined in rats in comparison with those of imipramine and endo-8-methyl-8-azabicyclo[3,2,1]oct-3-ol indol-3-yl-carboxylate hydrochloride (ICS205-930). Rats were treated with CGP36742, imipramine or ICS205-930 for 14 days. On day 14, the rats were subjected to 90 inescapable shocks. On day 15, the rats received the 40-trial escape test. The inescapable shocks increased escape failures in the escape test. CGP36742, imipramine and ICS205-930 dose-dependently improved the escape failures induced by the inescapable shocks. Baclofen attenuated the escape failures improving effect of CGP36742, imipramine and ICS205-930. Although the action of imipramine and ICS205-930 was attenuated by 1-(m-chlorophenyl)-biguanide (mCPBG), mCPBG failed to influence the CGP36742 action. Therefore, it is suggested that CGP36742 may have an antidepressant profile and that the mechanisms of CGP36742 in antidepressant action may be different from those of imipramine and ICS205 930. PMID- 10528129 TI - Differential haemodynamic effects of endothelin receptor antagonist, SB 209670, in conscious hypertensive and normotensive rats. AB - Endothelin has been implicated in the pathogenesis and/or maintenance of hypertension. Endothelin receptor antagonists lower blood pressure in the spontaneously hypertensive rat (SHR), but the regional haemodynamic effects of such drugs in the SHR remain unknown. The aim of this study was to examine the regional haemodynamic effects of the endothelin receptor antagonist, (+/-) (1S,2R,3S)-3-(2-carboxymethoxy-4-methoxyphenyl)-1-(3, 4-methylenedioxyphenyl)-5 (prop-1-yloxy)-indane-2-carboxylic acid (SB 209670), in SHR and Wistar-Kyoto (WKY) rats. Rats underwent a two-stage operation for implantation of Doppler flow probes and intravascular catheters. Recordings were made of mean arterial pressure (MAP), heart rate (HR) and renal (Ren), mesenteric (Mes) and hindquarters (HQ) blood flows and conductances (Cond). SHR and WKY received 4.5 h infusions of saline or SB 209670 (5 mg/kg priming dose+1 or 5 mg/kg/h, i.v.). SB 209670 lowered blood pressure in both SHR (-23+/-2 mm Hg) and WKY rats (-13+/-1 mm Hg). In addition, a lower dose infusion of SB 209670 also had an antihypertensive effect in SHR (-15+/-5 mm Hg). In SHRs which received the higher dose of antagonist, Ren, Mes and HQ Cond were significantly increased as was the HQ Cond in a low-dose group. In WKY rats, SB 209670 decreased Ren blood flow whilst increasing Mes and HQ blood flows and Cond. SB 209670 also attenuated the regional vasoconstrictor effects of endothelin-1, except in the Mes circulation in SHR. This study illustrates that SB 209670 causes differential haemodynamic effects in SHR and WKY rats. In SHR, the antihypertensive effect of SB 209670 was accompanied by a generalised vasodilatation in the Ren, Mes and HQ vascular beds. In WKY rats, the hypotensive effect of SB 209670 was accompanied by Mes and HQ vasodilatation, but with Ren vasoconstriction. Thus, endothelin is involved in the maintenance of blood pressure and vascular tone in both SHR and WKY rats, but the haemodynamic profiles of these effects differ between the two strains. PMID- 10528130 TI - In vivo and in vitro characterisation of a nonpeptide vasopressin V(1A) and V(2) receptor antagonist (YM087) in the rat. AB - This paper reports the in vitro and in vivo characterisation of a nonpeptide, orally active, vasopressin V(1A) and V(2) receptor antagonist, YM087 (methyl 1,4,5,6-tetrahydroimidazo[4, 5-d][1]benzoazepine-6-carbonyl)-2-phenylbenzanilide monohydrochloride) in the rat. YM087 dose dependently displaced the vasopressin V(1A) receptor antagonist radioligand, 125I-labelled [d(CH(2))(5),sarcosine(7)]vasopressin at vasopressin V(1A) receptors in liver and kidney medulla membranes and caused a concentration dependent displacement of the vasopressin V(2) receptor antagonist radioligand [3H]desGly-NH(2)(9)[d(CH(2))(5), D-Ile(2), Ile(4)]vasopressin at vasopressin V(2) receptors in kidney medulla membranes. In vitro binding kinetic studies showed YM087 acted as a competitive antagonist at liver V(1A) and kidney V(1A) and V(2) vasopressin receptors. Oral administration of YM087 (0.1-3 mg/kg) dose dependently inhibited vasopressin binding to liver V(1A) and kidney V(1A) and V(2) vasopressin receptors over 24 h. Oral YM087 (1-3 mg/kg/day) for 7 days in normotensive rats caused a dose dependent aquaresis with no effect on systolic blood pressure. These results show that YM087 is an orally effective vasopressin V(1A) and V(2) receptor antagonist that may be useful in the treatment of conditions characterised by vasoconstriction and fluid retention such as congestive heart failure. PMID- 10528131 TI - Cardiac effects of hexarelin in hypopituitary adults. AB - Growth hormone (GH)-releasing peptides possess specific pituitary, hypothalamic, and myocardial receptors. Seven adult male patients with GH deficiency (GHD) (age, mean+/-S.E.M.: 42.0+/-4.0 year) were studied by equilibrium radionuclide angiocardiography after i.v. administration of hexarelin, a peptide GH secretagogue. Data for these patients were compared with those for nine adult male controls (37.0+/-2.7 year). The GH response to hexarelin was negligible in patients with GHD compared to control subjects (CS) (peak: 1.9+/-0.9 vs. 45.7+/ 3.6 microg/l, P<0.001). Basal left ventricular ejection fraction (LVEF) in patients with GHD was lower than that in CS (50+/-1% vs. 63+/-2%, P<0.001). Hexarelin administration increased LVEF both in patients with GHD and in CS (peak: 57+/-2 vs. 70+/-2, respectively, P<0.05 vs. baseline) without changing catecholamine levels, mean blood pressure (MBP), or cardiac output in either group. In conclusion, the acute administration of hexarelin exerts a short lasting positive inotropic effect in humans, probably GH-independent and mediated by specific myocardial receptors for GH secretagogues. PMID- 10528132 TI - Different roles of two types of endothelin receptors in partial ablation-induced chronic renal failure in rats. AB - Recent work has drawn attention to endothelin as a likely contributor to renal pathogenesis. To elucidate the mechanism of progressive renal disease, we investigated the mRNA expression of endothelin and endothelin receptors, and the effect of endothelin ET(A), and/or ET(B) receptor antagonists on disease progression in the remnant kidney model. Proteinuria progressively increased in rats subjected to 5/6 nephrectomy (Nx) after 8 weeks (from 25+/-3 to 221+/-28 microg min(-1) kg(-1)). Creatinine clearance (Ccr) after renal ablation gradually decreased by 8 weeks (from 5.04+/-0.42 to 2. 68+/-0.26 ml min(-1) kg(-1)). Together with maximal proteinuria and decreased renal function, there was an increase in cortical mRNA expression of prepro endothelin-1 and endothelin ET(A) receptor expression, but a decrease in endothelin ET(B) receptor expression and in urinary excretion of endothelin-1. Administration (1-3 mg/day) of S-0139, (+) disodium 27-[(E)-3-[2-[(E)-3-carboxylatoacryloylamino]-5-hydroxyphenyl]a crylay loxy]-3-oxoolean-12-en-28-oate, an endothelin ET(A) receptor-specific antagonist, had a beneficial effect on the evolution of the disease, preventing the appearance of intense proteinuria (113+/-11) and decreased Ccr (3.97+/-0.33). High blood pressure was observed in rats with 5/6 Nx and was decreased by S-0139 administration. To examine whether treatment modalities that decrease endothelin ET(B) receptor signaling have a deleterious effect on the kidney remnant, the effect of 97-618, an endothelin ET(B) receptor-specific antagonist, 4-tert-butyl N-[5-(2-methoxyphenoxy)-6-(4-oxobutoxy)pyromidine+ ++-4-yl]b enzenesulfonamide, was also examined on the action of S-0139. Concomitant administration of S-0139 and 97-618 reversed the beneficial effect of S-0139 alone in the remnant kidney on proteinuria and renal functional impairment. These findings indicate that endothelin participates in the pathogenesis of proteinuria and glomerular injury and that an endothelin ET(A) receptor-specific antagonist could be useful in the treatment of some forms of human nephritis. The loss of endothelin ET(B) receptor seems to be important in the progression of renal disease. PMID- 10528133 TI - Opposing effects of rapamycin and cyclosporin A on activation-induced Ca(2+) release. AB - Insofar as Ca(2+) plays a major role in T cell activation, we investigated the effect of the immunosuppressants cyclosporin A and rapamycin on T cell proliferation and on the activation-induced increase in [Ca(2+)](i). Both cyclosporin A and rapamycin inhibited mitogen (concanavalin A and phytohemagglutinin) and ionomycin+phorbol myristate acetate (PMA)-driven T cell proliferation (Ca(2+)-dependent). However, only rapamycin suppressed T cell proliferation stimulated by anti-CD28 antibody (Ab)+PMA, and recombinant interleukin-6-stimulated proliferation of the interleukin-6 dependent B9 cells (Ca(2+)-independent). These differences were associated with a different effect of both drugs on Ca(2+) release, as cyclosporin A attenuated while rapamycin augmented the mitogen-induced elevation in [Ca(2+)](i). Collectively, this supports the notion that Ca(2+) is required in early stages of T cell activation, and that cyclosporin A blocked only Ca(2+)-dependent while rapamycin blocked both Ca(2+)-dependent and -independent events of T cell activation. PMID- 10528134 TI - Effects of differentiation-inducing factors of Dictyostelium discoideum on human leukemia K562 cells: DIF-3 is the most potent anti-leukemic agent. AB - DIF-1 (differentiation-inducing factor-1; 1-(3,5-dichloro-2, 6-dihydroxy-4 methoxyphenyl)hexan-1-one) is a putative morphogen that induces stalk cell formation in the cellular slime mold, Dictyostelium discoideum. It has been previously reported that DIF-1 exhibits anti-tumor activity in mammalian cells. In this study, we examined the effects of six DIF analogues on DNA synthesis, cell growth, erythroid differentiation, and cytosolic free calcium concentration ([Ca2+]i) in human leukemia K562 cells. The DIF analogues used here were DIF-1, DIF-2 (which has pentanone in place of hexanone), DIF-3 (dechlorinated form of DIF-1), 2-MIDIF-1 (2-methoxy isomer of DIF-1), DMPH (dechlorinated form of DIF 3), and THPH (4-hydroxy substitution of DMPH). DIF-3 proved to be the most potent anti-leukemic agent among them, and the order of potency for causing growth inhibition, erythroid induction, and increases in [Ca2+]iTHPH in all the categories tested. The present results suggest new routes for the development of more potent and effective anti-tumor agents. PMID- 10528135 TI - Inhibition of plasma membrane monoamine transporters by beta-ketoamphetamines. AB - Methcathinone and methylone, the beta-ketone analogues of methamphetamine and 3,4 methylenedioxymethamphetamine (MDMA), respectively, were tested for neurotransmitter uptake inhibition in vitro. The beta-ketones were threefold less potent than the nonketo drugs at inhibiting platelet serotonin accumulation, with IC(50)'s of 34.6+/-4.8 microM and 5.8+/-0.7 microM, respectively. Methcathinone and methylone were similar in potency to methamphetamine and MDMA at catecholamine transporters individually expressed in transfected glial cells. For dopamine uptake, IC(50)'s were 0.36+/-0.06 microM and 0.82+/-0.17 microM, respectively; for noradrenaline uptake, IC(50) values were 0.51+/-0.10 microM and 1. 2+/-0.1 microM, respectively. In chromaffin granules, IC(50)'s for serotonin accumulation were 112+/-8.0 microM for methcathinone and 166+/-12 microM for methylone, 10-fold higher than the respective values for methamphetamine and MDMA. Our results indicate that methcathinone and methylone potently inhibit plasma membrane catecholamine transporters but only weakly inhibit the vesicle transporter. PMID- 10528136 TI - Troglitazone but not pioglitazone affects ATP-sensitive K(+) channel activity. AB - We compared the effects of the two thiazolidinedione derivatives, troglitazone and pioglitazone, on ATP-sensitive K(+) (K(ATP)) channel activities. Pancreatic beta-cell type and cardiac type K(ATP) channels were reconstituted in COS-1 cells (SV 40-transformed African green monkey kidney (AGMK) cells) by heterologously expressing sulfonylurea receptor 1 (SUR1) plus Kir6.2 and sulfonylurea receptor 2A (SUR2A) plus Kir6.2, respectively. Troglitazone inhibited [86Rb(+)] efflux in both K(ATP) channel types in the presence of metabolic inhibitors, which was confirmed by electrophysiological techniques. The [86Rb(+)] efflux increased by the channel openers diazoxide and pinacidil was abolished by troglitazone. In contrast, pioglitazone did not affect these channel activities in either type K(ATP) channel. These results suggest that troglitazone modulates the various cellular functions including insulin secretion by inhibiting the K(ATP) channels, while pioglitazone has no effect on K(ATP) channel activity. PMID- 10528137 TI - Structural determinants of phorbol ester binding in synaptosomes: pharmacokinetics and pharmacodynamics. AB - The present study used structurally distinct phorbol esters to investigate the relationship between their pharmacokinetics of binding to protein kinase C (PKC) in rat brain cortex synaptosomes, their affinity for PKC in synaptosomes and ability to enhance noradrenaline release from rat brain cortex. Affinity binding studies using [3deoxyphorbol 13-tetradecanoate (dPT)=PDB&z. Gt;12-deoxyphorbol 13 acetate (dPA)=phorbol 12,13-diacetate (PDA). In intact synaptosomes PDB, dPA and PDA rapidly displaced bound [3H]PDB whereas PMA and dPT were comparatively slow. However, the displacement rates for all the phorbol esters were equally rapid in synaptosomal membranes or synaptosomes permeabilised with Staphylococcus alpha toxin. These results suggest that the lipophilic phorbol esters (dPT and PMA) are slower to displace [3H]PDB binding because they are hindered by the plasma membrane. In brain cortex slices it was found that the rate of displacement of [3H]PDB binding was closely correlated with the degree of elevation of transmitter noradrenaline release. Thus kinetic characteristics may determine biological responses and this may be particularly evident in events which occur rapidly or where there is fast counter-regulation. PMID- 10528138 TI - The site of general anesthesia and cytochrome P450 monooxygenases: occupation of the enzyme heme pocket by xenon and nitrous oxide. AB - In previous studies, cytochrome P450 monooxygenases were shown to be appropriately sensitive to structurally diverse compounds varying widely in anesthetic potencies and to increasing carbon-number series of straight chain primary and secondary alcohols and rigid cyclic alcohols. We now report that xenon and nitrous oxide, at one atmosphere, occupy the P450 heme cavity and competitively inhibit catalytic activity. The heme enzymes appear to be the most relevant model of the site of general anesthesia, thus far identified. PMID- 10528140 TI - Cocaine discrimination and time-course effects in male and female Wistar rats. AB - Previously, sex differences have been observed in the behavioral effects of acute and chronic cocaine administration. In the present experiment, male and female rats were trained to discriminate intraperitoneal injections of 10.0 mg/kg cocaine from its vehicle. It was hypothesized that the subjective effects of cocaine might differ between male and female rats. It was further hypothesized that generalization gradients between male and female rats might differ as a function of the time since cocaine administration. In addition, we were interested to see whether multiple generalization gradients could be determined within the same experimental session. For that purpose, two different types of generalization tests were conducted in extinction, one in which subjects were tested both 10 min and 30 min following cocaine administration (vehicle, 1.0, 3.0, 5.6, 10 or 17 mg/kg) and one in which subjects were only tested 30 min after cocaine administration. The generalization gradients obtained 30 min following drug administration were shifted to the right of the gradient obtained 10 min following drug administration. The two 30-min gradients were not different from one another, showing that multiple generalization gradients can be obtained within the same experimental session. PMID- 10528139 TI - Dopaminergic modulation of rat pup ultrasonic vocalizations. AB - The dopamine D(1) receptor agonist, R(+)-6-chloro-7, 8-dihydroxy-1-phenyl-2,3,4,5 tetrahydro-1H-3-benzazepine hydrobromide (SKF 81297), the dopamine D(2)/D(3) receptor agonist, trans-(-)-4aR-4,4a,5,6,7,8,8a,9-octahydro-5-propyl-1H pyrazolo[3, 4-g]quinoline hydrochloride (quinpirole), and the dopamine D(3) receptor agonist, (+/-)-7-hydroxy-dipropylaminotetralin hydrobromide (7-OH-DPAT) all reduced the frequency of isolation-induced infant rat ultrasonic vocalizations and lowered body temperature when compared to saline-injected controls. Ultrasonic vocalization rate was not affected by either the dopamine D(1) receptor antagonist, R(+)-2,3,4, 5-tetrahydro-8-iodo-3-methyl-5-phenyl-1H-3 benzazepin-7-ol hydrochloride (SCH 23390) or the dopamine D(2)/D(3) receptor antagonist, S(-)-raclopride-L-tartrate (raclopride) when given alone, nor did these antagonists block the ultrasonic vocalization reductions caused by the dopamine D(1) receptor agonist or the dopamine D(2)/D(3) receptor agonist. The dopamine D(2)/D(3) receptor antagonist but not the dopamine D(1) receptor antagonist blocked the dopamine D(3) receptor agonist's ultrasonic vocalization reduction. SKF 81297 reduced general activity while quinpirole and 7-OH-DPAT increased activity. Raclopride reversed quinpirole's reduction in body temperature, as well as 7-OH-DPAT's effects on body temperature, ultrasonic vocalizations, and activity. These results indicate that dopamine D(1), D(2)/D(3), and D(3) receptor agonists all reduce ultrasonic vocalizations by as yet undetermined mechanisms. PMID- 10528141 TI - Suramin inhibits the toxic effects of presynaptic neurotoxins at the mouse motor nerve terminals. AB - Clinically available chemical antagonists of snake neurotoxins still await to be identified. In this study, we demonstrate that an anti-trypanosomiasis agent, suramin, is an effective inhibitor of beta-bungarotoxin isolated from the venom of Formosan Krait snake. Following intraperitoneal injection (12 ng/g) of beta bungarotoxin in mice, the time to paralysis (loss a limb withdrawal reflex, 21. 8+/-3.4 h, n=4) was significantly prolonged after intravenous injection (16 microg/g) of suramin (35.9+/-4.0 h, n=4, P<0.05). The mechanism of this inhibitory effect of suramin was analyzed at the mouse nerve terminals. beta Bungarotoxin (1 microg/ml) produces an irreversible blocking effect of nerve evoked muscle contractions of mouse phrenic nerve-diaphragm (blocking time 135+/ 6 min, n=6). Pretreatment with suramin (0.3 mM) significantly prolonged the blocking time by three-fold. This selective inhibitory effect of suramin was further confirmed when suramin was shown to delay the neuromuscular blocking effect of another presynaptic neurotoxin, crotoxin (from American rattlesnake venom), but not that of the postsynaptic neurotoxin, alpha-bungarotoxin. Furthermore, suramin inhibited beta-bungarotoxin in blocking transmitter release as revealed by prolonging the time to abolish the end-plate potential amplitude (with suramin, 391+/-8 min; without treatment, 141+/-5 min). K(+) current was measured in the mouse triangularis sterni preparation; suramin (0.3 mM) had no significant effect on beta-bungarotoxin in inhibiting K(+) current (77+/-3% of control; with suramin 75+/-3% of control, respectively). These findings clearly show that suramin is an inhibitor of presynaptic neurotoxins, mediated by interrupting the toxins in blocking the releasing mechanism of transmitter at the motor nerve terminals. The implication of these findings is that suramin and related compounds can become useful agents in management of snakebites. PMID- 10528142 TI - Delayed protection against ventricular arrhythmias by monophosphoryl lipid-A in a canine model of ischaemia and reperfusion. AB - Bacterial endotoxin reduces the severity of ventricular arrhythmias which occur when a coronary artery is occluded several hours later. We have now examined in anaesthetised dogs the effects on ischaemia and reperfusion-induced arrhythmias, of a non-toxic derivative component of the endotoxin molecule of the lipid-A (monophosphoryl lipid-A). This was given intravenously, in doses of 10 and 100 microg kg(-1), 24 h prior to coronary artery occlusion. Arrhythmia severity was markedly reduced by monophosphoryl lipid-A. During ischaemia, ventricular premature beats were reduced from 315+/-84 in the vehicle controls to 89+/-60 (with the lower dose of monophosphoryl lipid-A) and 53+/-23 (P<0.05) with the higher dose. The incidence of ventricular tachycardia was reduced from 75% to 25% (P<0.05) and 31% (P<0.05), and the number of episodes of ventricular tachycardia from 13.4+/-4.9 per dog to 1.1+/-1.1 (P<0.05) and 1. 2+/-0.9 (P<0.05) after doses of 10 and 100 microg kg(-1), respectively. The incidence of ventricular fibrillation during occlusion and reperfusion in the control group was 96% (15/16), i.e., only 6% (1/16) dogs survived the combined ischaemia-reperfusion insult. Monophosphoryl lipid-A (100 microg kg(-1)) significantly reduced the incidence of occlusion-induced ventricular fibrillation (from 50% to 7%; P<0.05), and increased survival following reperfusion to 54% (P<0.05). Monophosphoryl lipid-A also significantly reduced ischaemia severity as assessed from ST-segment elevation recorded from epicardial electrodes as well as the degree of inhomogeneity of electrical activation within the ischaemia area. There were no haemodynamic differences prior to coronary occlusion between vehicle controls and monophosphoryl lipid-A-treated dogs. These results demonstrate that monophosphoryl lipid-A reduces arrhythmia severity 24 h after administration. Although the precise mechanisms are still unclear, there is some evidence that nitric oxide and prostanoids (most likely prostacyclin) may be involved because the dual inhibition of nitric oxide synthase and cyclooxygenase enzymes by administration of aminoguanidine and meclofenamate abolished the marked antiarrhythmic protection resulted from monophosphoryl lipid-A treatment 24 h previously. PMID- 10528143 TI - Alpha-adrenoceptor-mediated pressor responses in pithed rats fed diets with different calcium contents. AB - Results of many clinical and experimental studies indicate an inverse relationship between dietary calcium and the prevalence of hypertension. Our study was designed to evaluate the alterations in arterial blood pressure and the changes in alpha-adrenoceptor-mediated vascular reactivity in normotensive Sprague-Dawley and spontaneously hypertensive rats (SHR) fed from weaning (3 weeks of life) three diets: normal calcium (Ca 1%), low calcium (Ca 0.1%), and high calcium (Ca 2.5%). The systolic and the diastolic arterial blood pressures were measured weekly by the tail cuff method. The plasma calcium levels in the animals were also measured regularly by colourimetric methods, and the alpha adrenoceptor-mediated vascular reactivity was evaluated by measuring the pressor responses to alpha-adrenoceptor agonists in pithed rats. These determinations were carried out at the end of the feeding periods (9 weeks of life in Sprague Dawley rats and 20 weeks of life in SHR) and also at the moments when maximal differences in arterial blood pressure were observed between the conscious animals fed the normal calcium diet and those fed the other two diets. Dietary calcium deficiency increased arterial blood pressure in both strains but calcium supplements were effective to lower this only in hypertensive animals. The plasma calcium levels were altered in both strains when calcium administration was not normal. The low-calcium diet did not modify the pressor responses to either the alpha(1)-adrenoceptor agonist, methoxamine, or the alpha(2)-adrenoceptor agonist, B-HT 920 (5-allyl-2-amino-5,6,7, 8-tetrahydro-4H-thiazolo-(4,5-D)-acepin dihydrochloride, talixepole), in the normotensive and the hypertensive rats. On the contrary, the high-calcium diet caused a definite decrease in alpha(1)- and alpha(2)-adrenoceptor-mediated vascular reactivity in both strains. The changes in the alpha-adrenoceptor-mediated vasoconstrictor responses were observed in pithed 9-week old Sprague-Dawley rats and in pithed 20-week old SHR, but none were observed in pithed 15-week old SHR, although at this age maximal differences in arterial blood pressure between the animals fed the high- and the normal calcium diet were observed. The results of this study suggest that the mechanisms implicated in the effects of dietary calcium supplements on arterial blood pressure are clearly different from the mechanisms, which bring about changes in arterial blood pressure when the diet is deficient in calcium. The results of this study also show that calcium administration causes variations in alpha adrenoceptor-mediated vascular reactivity, but this is probably not the only mechanism implicated in the calcium effect on arterial blood pressure. PMID- 10528144 TI - Effects of alniditan on neurogenic oedema in the rat dura mater and on contraction of rat basilar artery. AB - The non-indole 5-HT receptor agonist, alniditan (R 91274), was tested and compared to sumatriptan in an in vivo model of neurogenic inflammation within the meninges of rats and in rat basilar artery in a Mulvany-Halpern chamber in vitro. Alniditan dose dependently attenuated the neurogenic inflammation and was more potent than sumatriptan. The alniditan response was blocked by the 5-HT(1B/D) receptor antagonist, GR 127935 (2'-methyl-4'-(5-methyl-[1,2, 4]oxadiazol-3-yl) biphenyl-4-carboxylic acid [4-methoxy-3-(4-methyl-piperazin-1-yl)-phenyl]-amide), but not by ketanserin, indicating that the effect is mediated through 5-HT(1B/D) receptors. Alniditan did not attenuate substance P-induced inflammation, suggesting that the mediating receptors are located prejunctionally. In vitro alniditan exhibited less vasoconstrictive effects on the rat basilar artery than did sumatriptan, although at a very high concentration (1 mM), alniditan caused intensive constriction, most likely through a mechanism independent from 5-HT receptor activation. PMID- 10528145 TI - Nitric oxide-releasing compounds inhibit neutrophil adhesion to endothelial cells. AB - In the present work, we demonstrated that chemically different nitric oxide (NO) releasing compounds inhibit tumor necrosis factor alpha (TNF-alpha)-induced polymorphonuclear leukocyte adhesion to endothelial cells in vitro. Two mesoionic oxatriazole derivatives GEA 3162 (1,2,3,4-oxatriazolium,5-amino-3(3, 4 dichlorophenyl)-chloride) and GEA 3175 (1,2,3,4-oxatriazolium, -3-(3-chloro-2 methylphenyl)-5-[[(4-methylphenyl)sulfonyl]amino]-, hydroxide inner salt) were compared to the earlier-known NO donor SIN-1 (3-morpholino-sydnonimine). GEA 3162 (3-10 microM) and GEA 3175 (10-30 microM) inhibited human polymorphonuclear leukocyte adhesion to B(4) endothelial cells in a dose-dependent manner being more potent than SIN-1. In the present model, leukocytes rather than endothelial cells seemed to be the target of the effect of NO. Flow cytometric analysis showed that NO-releasing compounds did not alter TNF-alpha induced CD11/CD18 surface expression in polymorphonuclear leukocytes. The inhibitory action of NO releasing compounds on adhesion paralleled with the increased synthesis of cGMP in polymorphonuclear leukocytes. Analogues of cGMP inhibited polymorphonuclear leukocyte adhesion indicating a role for cGMP in the action of NO donors. The results suggest that exogenous NO in the form of NO-releasing compounds inhibits polymorphonuclear leukocyte adhesion to endothelial cells, which may be implicated in the regulation of leukocyte migration and leukocyte-mediated tissue injury. PMID- 10528146 TI - Enhanced striatal dopamine D(2) receptor-induced [35S]GTPgammaS binding after haloperidol treatment. AB - Dopamine receptor-G protein coupling and dopamine D(2) receptor density were assessed in rats treated for 3 weeks with either haloperidol (2 mg/kg; i.p.) or vehicle. After 3 days of withdrawal, agonist-induced guanosine 5'-O-(gamma [35S]thio)triphosphate ([35S]GTPgammaS) and [3H]spiperone binding were determined in striatal homogenates. Maximal [3H]spiperone binding was increased (24.8%, P<0.01) following haloperidol treatment. The efficacy of dopamine and the dopamine D(2) receptor agonist R(-)-10, 11-dihydroxy-N-n-propylnorapomorphine (NPA) to induce [35S]GTPgammaS binding were found to be increased by 24.1% (P<0.01) and 44.6% (P<0. 001), respectively. When measured in the presence of a saturating concentration of a dopamine D(2) receptor antagonist, the response to dopamine was not significantly affected by haloperidol treatment. In addition, the measurement of haloperidol-induced catalepsy confirmed that the efficient dopamine receptor blockade was followed by a progressive development of dopaminergic supersensitivity. Taken together, these results indicate that a functional pool of dopamine D(2) receptors is increased after prolonged haloperidol administration. PMID- 10528147 TI - Structure-activity relationship of gramine derivatives in Ca(2+) release from sarcoplasmic reticulum. AB - 5,6-Dibromo-1,2-dimethylgramine evoked Ca(2+) release from skeletal muscle sarcoplasmic reticulum through ryanodine receptors in a concentration-dependent manner with an EC(50) of 22.2 microM. Since the EC(50) of caffeine was 0.885 mM, 5,6-dibromo-1,2-dimethylgramine was 40 times more sensitive than caffeine. Among 14 gramine derivatives having different substituents at N-1, C-2, C-5 or C-6 of the indole skeleton, we found that five derivatives were effective. Study of the structure-activity relationship for Ca(2+) release indicated that 1-methylation and/or both 5- and 6-bromination are important for Ca(2+) release. Thus, gramine derivatives are useful tools for the investigation of Ca(2+) release from sarcoplasmic reticulum. PMID- 10528148 TI - The 5-HT(1A) receptor antagonist robalzotan completely reverses citalopram induced inhibition of serotonergic cell firing. AB - 5-HT(1A) receptor antagonists have been suggested to increase the efficacy of selective serotonin (5-hydroxytryptamine; 5-HT) reuptake inhibitors in the treatment of depression by enhancing the increase in brain 5-HT induced by 5-HT reuptake blockade. Here, the novel 5-HT(1A) receptor antagonist robalzotan [(R)-3 N, N-dicyclobutylamino-8-fluoro-3, 4-dihydro-2H-1-benzopyran-5-carboxamide hydrogen (2R, 3R) tartrate monohydrate] (12.5, 25, 50, 100 microg/kg, i.v.) was found to completely reverse the acute inhibitory effect of citalopram (300 microg/kg i.v.) or paroxetine (100 microg/kg, i.v.) on the activity of 5-HT neurons in the dorsal raphe nucleus in rats. Robalzotan (5, 50 microg/kg, i.v.) by itself increased the firing rate of the majority of 5-HT cells studied. The present results suggest that robalzotan may indeed augment the increases in 5-HT output induced by selective 5-HT reuptake inhibitors by antagonizing the feedback inhibition of 5-HT cell firing produced by such drugs. Thus, robalzotan may be effective by enhancing the action of selective 5-HT reuptake inhibitors or as monotherapy in the treatment of depression. PMID- 10528149 TI - Chronic haloperidol treatment impairs glutamate transport in the rat striatum. AB - High-affinity, Na(+)-dependent transport of glutamate into neurons and glial cells maintains the extracellular concentration of this neurotransmitter at a sub toxic level. Chronic blockade of dopamine D(2) receptors with haloperidol elevates extracellular glutamate levels in the striatum. The present study examines the effect of long-term haloperidol treatment on glutamate transporter activity using an assay based on measuring the uptake of D-[3H]aspartate in striatal synaptosomes prepared from male Wistar rats. The maximal rate of glutamate transport in the striatum is reduced by 63% following 27 weeks of haloperidol treatment. This impairment of glutamate transport may be important in chronic neuroleptic drug action. PMID- 10528150 TI - The role of lats in cell cycle regulation and tumorigenesis. PMID- 10528151 TI - CDK-independent activities of D type cyclins. PMID- 10528152 TI - TCF transcription factors: molecular switches in carcinogenesis. AB - Although originally cloned as lymphoid transcription factors, members of the T cell factor (Tcf) family are now well recognized as key activators/repressors in many developmental processes. Transcriptionally inert Tcf factors become potent transactivators upon interaction with the Wnt signaling product beta-catenin or its Drosophila counterpart Armadillo. In contrast, Tcf proteins mediate repression when bound to members of the Groucho family of transcriptional repressors, CBP and CtBP. Recently, Tcf factors have been reported as tumor inducers, aberrantly activating their target genes as a result of elevated beta catenin levels in many types of cancer. These abnormal beta-catenin levels are usually caused by stabilizing mutations in beta-catenin itself or truncating mutations in the adenomatous polyposis coli (APC) tumor suppressor gene. In this review, we will give a chronological overview of the Tcf factors and the phenotypes of Tcf mutant mice, as well as Tcf-binding partners. We will discuss Tcf signaling upon interaction with different partners, resulting in activator and repressor roles of Tcf factors in the light of carcinogenic events. PMID- 10528153 TI - Chromophore-assisted laser inactivation (CALI) to elucidate cellular mechanisms of cancer. AB - Chromophore-assisted laser inactivation (CALI) is a new technology for acute protein inactivation in living cells. It targets laser energy to specific proteins via non-function-blocking antibodies that are labeled with the dye malachite green. Excitation of the dye generates short-lived free radicals that damage the bound protein without affecting other cellular components. The wavelength of laser light used (620 nm) is not readily absorbed by cells such that non-specific light damage does not occur. CALI provides an alternative to other inactivation strategies and has the advantages of high spatial and temporal resolution. The ultimate value of this technology for cancer research will be assessed by how effective CALI is in ascribing in situ function during cancer relevant processes and in identifying and validating protein targets for drug discovery. Recent work using CALI on ezrin and pp60-c-src, two proteins that may be involved in cancer, suggests its potential. Further application of CALI will likely be of utility for understanding cellular mechanisms of cancer and developing cancer therapeutics. PMID- 10528154 TI - 1999 keystone symposium on B lymphocyte biology and disease: B cell malignancy II session. PMID- 10528155 TI - Many faces of ATM: eighth international workshop on ataxia-telangiectasia. PMID- 10528156 TI - 11th Lorne Cancer Conference, Lorne, Victoria, Australia, 11-14th February 1999. PMID- 10528157 TI - Presidential address to the American Association of Immunologists. Stimulating naive T cells. PMID- 10528158 TI - Cutting edge: Ig heavy chain 3' HS1-4 directs correct spatial position independent expression of a linked transgene to B lineage cells. AB - The Ig H chain locus is regulated by a set of cis-acting elements. Hypersensitive sites (HS) located 3' of the IgH, HS1-4, has been suggested to act as a locus control region (LCR) in cell lines. To assess the proposed role of HS1-4 acting as an LCR, we generated transgenic mice harboring a VH promoter-beta-globin reporter gene linked to the Ig H chain HS1-4 3'regulatory sequences. Transgene expression is strictly confined to B lymphocytes, with no detectable expression outside the B cell lineage in all transgenic founder lines. Furthermore, reporter gene activity is integration independent but not copy number dependent. Thus, additional sequences are required to allow the HS1-4 regulatory region to act as a classical LCR in mice. Our data are discussed in the context of tissue-specific gene expression in B lineage cells. PMID- 10528159 TI - Cutting edge: Role of C-C chemokine receptor 5 in organ-specific and innate immunity to Cryptococcus neoformans. AB - After intratracheal inoculation of the AIDS-associated pathogen Cryptococcus neoformans, 12-wk survival was >90% for CCR5+/+ mice but <25% for CCR5-/- mice. There were no defects in lung leukocyte recruitment (wk 5), pulmonary clearance, or delayed-type hypersensitivity in CCR5-/- mice. However, CCR5-/- mice had defects in leukocyte recruitment into the brain and, strikingly, in elimination of cryptococcal polysaccharide from the brain. In nonimmune CCR5-/- mice, there was a significant defect in macrophage recruitment after challenge with shed cryptococcal products (C. neoformans filtrate Ag) but not other nonspecific stimuli. Thus, CCR5 plays specific roles in innate immunity and organ-specific leukocyte trafficking during host defense against C. neoformans. PMID- 10528160 TI - Cutting edge: Cross-talk between cells of the innate immune system: NKT cells rapidly activate NK cells. AB - alpha-Galactosylceramide (alpha-GalCer) is a glycolipid with potent antitumor properties that binds to CD1d molecules and activates mouse Valpha14 and human Valpha24 NKT cells. Surprisingly, we found that, as early as 90 min after alpha GalCer injection in vivo, NK cells also displayed considerable signs of activation, including IFN-gamma production and CD69 induction. NK activation was not observed in RAG- or CD1-deficient mice, and it was decreased by pretreatment with anti-IFN-gamma Abs, suggesting that, despite its rapid induction, it was a secondary event that depended on IFN-gamma release by NKT cells. At later time points, B cells and CD8 T cells also began to express CD69. These findings identify a high-speed communication network between the innate and adaptive immune systems in vivo that is initiated upon NKT cell activation. They also suggest that the antitumor effects of alpha-GalCer result from the sequential recruitment of distinct innate and adaptive effector lymphocytes. PMID- 10528161 TI - Cutting edge: KAP10, a novel transmembrane adapter protein genetically linked to DAP12 but with unique signaling properties. AB - Transmembrane adapter proteins are a class of molecules that mediate signals from an extracellular receptor to the cytoplasm of the cell. We have cloned a novel transmembrane adapter protein called KAP10, a approximately 10-kDa protein that is encoded within 100 bp of the DAP12 locus on human chromosome 19. KAP10 is predominantly expressed in immune cells, including NK cells, T cells, and monocytes. We show that KAP10, unlike other transmembrane adapter proteins, binds phosphatidylinositol-3 kinase following phosphorylation of a cytoplasmic YINM motif, which results in activation of Akt. In addition, we identify KAP10 as being able to bind the adapter protein Grb2. Based on our data, we suggest that this molecule is involved in stimulation and costimulation in cells of both myeloid and lymphoid origin. PMID- 10528162 TI - Mitochondria connects the antigen receptor to effector caspases during B cell receptor-induced apoptosis in normal human B cells. AB - We have previously reported that CD40 stimulation sensitizes human memory B cells to undergo apoptosis upon subsequent B cell receptor (BCR) ligation. We have proposed that activation stimuli connect the BCR to an apoptotic pathway in mature B cells and that BCR-induced apoptosis of activated B cells could serve a similar function as activation-induced cell death in the mature T cell compartment. Although it has been reported that caspases are activated during this process, the early molecular events that link the Ag receptor to these apoptosis effectors are largely unknown. In this study, we report that acquisition of susceptibility to BCR-induced apoptosis requires entry of memory B cells into the S phase of the cell cycle. We also show that transduction of the death signal via the BCR sequentially proceeds through a caspase-independent and a caspase-dependent phase, which take place upstream and downstream of the mitochondria, respectively. Furthermore, our data indicate that the BCR-induced alterations of the mitochondrial functions are involved in activation of the caspase cascade. We have found both caspases-3 and -9, but not caspase-8, to be involved in the BCR apoptotic pathway, thus supporting the notion that initiation of the caspase cascade could be under the control of the caspase-9/Apaf 1/cytochrome c multimolecular complex. Altogether, our findings establish the mitochondria as the connection point through which the Ag receptor can trigger the executioners of apoptotic cell death in mature B lymphocytes. PMID- 10528163 TI - Paired Stat6 C-terminal transcription activation domains required both for inhibition of an IFN-responsive promoter and trans-activation. AB - The cytokines IL-4 and IFN-gamma exert biologically antagonistic effects that in part reflect opposing influences on gene transcription. While the molecular mechanisms for IL-4-mediated transcription activation have been extensively studied, little is known about molecular mechanisms required for IL-4 inhibition of IFN-gamma signaling. We have investigated IL-4 inhibition of the IFN-gamma inducible promoter for IFN regulatory factor-1 (IRF-1). In a cell line with low endogenous Stat6, increasing levels of activated Stat6 at constant doses of IFN gamma and IL-4 leads to inhibition of the IRF-1 promoter. The Stat1-dependent IFN gamma activation sequence element of the IRF-1 promoter is a target for Stat6 mediated inhibition despite apparently normal Stat1 DNA binding. However, our data are inconsistent with competition between Stat1 and Stat6 for access to the IRF-1 IFN-gamma activation sequence or for an essential coactivator as a mechanism for this Stat6-mediated inhibition. Instead, the data demonstrate that a threshold of Stat6 transcription activation domains is required for IL-4 dependent inhibition. The findings provide evidence of a novel mechanism in which the Stat6 transcription activation domains play a critical role in the IL-4 mediated inhibition of an IFN-gamma-inducible promoter. PMID- 10528164 TI - TH cells primed during influenza virus infection provide help for qualitatively distinct antibody responses to subsequent immunization. AB - The quality of the primary Ab-forming cell (AFC) response in cervical lymph nodes and mediastinal lymph nodes of mice to intranasal influenza virus was strongly influenced by viral replicative capacity. IgA secretors were prominent in the early AFC response to infectious virus in mediastinal lymph nodes, while IgG expression was more frequent among isotypically switched AFC in cervical lymph nodes of the same mice; this pattern was reversed in the response to inactivated virus. Influenza viruses A/Puerto Rico/8/34 (A/PR8) and A/X-31 share six of eight genome segments, differing only in hemagglutinin (H1 in A/PR8, H3 in A/X-31) and neuraminidase (N1 in A/PR8, N2 in A/X-31) genes. These viruses therefore elicit extensively cross-reactive TH populations, though their glycoproteins are serologically unrelated. Mice recovered from an A/X-31 infection thus mount a primary B cell response against A/PR8 glycoproteins, when challenged with the latter virus, though this response can call upon memory TH cells. To assess the impact of memory TH populations on a primary Ab response, we compared the AFC response to inactivated A/PR8 in naive mice and mice that had cleared an A/X-31 infection. A/X-31 immune mice mounted a more vigorous AFC response against A/PR8 H1 and N1 glycoproteins than naive animals, when immunized intranasally with inactivated A/PR8. However the distribution of isotypes among H1/N1-specific AFC in lymph nodes of A/X-31-primed mice resembled that of naive mice. Evidently, in this functional context, memory TH cells retained the ability to help Ab responses different in quality from that generated during their primary reaction. PMID- 10528165 TI - Granzyme B-induced loss of mitochondrial inner membrane potential (Delta Psi m) and cytochrome c release are caspase independent. AB - CTLs kill targets by inducing them to die through apoptosis. A number of morphological and biochemical events are now recognized as characteristic features of the apoptotic program. Among these, the disruption of the inner mitochondrial transmembrane potential (Delta Psi m) and the release of cytochrome c into the cytoplasm appear to be early events in many systems, leading to the activation of caspase-3 and, subsequently, nuclear apoptosis. We show here that, in Jurkat targets treated in vitro with purified granzyme B and perforin or granzyme B and adenovirus, Delta Psi m collapse, reactive oxygen species production, and cytochrome c release from mitochondria were observed. Loss of Delta Psi m was also detected in an in vivo system where green fluorescent protein-expressing targets were attacked by a cytotoxic T cell line that kills predominantly through the granzyme pathway. DNA fragmentation, phosphatidylserine externalization, and reactive oxygen species production were inhibited in the presence of the caspase inhibitors benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (zVAD-fmk) and benzyloxycarbonyl-Asp-Glu-Val-Asp-fluoromethyl ketone (zDEVD-fmk) in our in vitro system. Importantly, in either the in vitro or in vivo systems, these inhibitors at concentrations up to 100 microM did not prevent Delta Psi m collapse. In addition, cytochrome c release was observed in the in vitro system in the absence or presence of zVAD-fmk. Thus the granzyme B dependent killing pathway in Jurkat targets involves mitochondrial alterations that occur independently of caspases. PMID- 10528167 TI - Antigen presentation determines the fate of the T memory response in vivo after sublethal gamma-irradiation. AB - The survival of memory T cells is critical to vaccination strategies for infectious diseases and cancer, whereas their elimination may be crucial for treatment of autoimmune states. We examined the consequences of gamma irradiation, which induces apoptosis of memory T cells in vitro, on the memory response to MHC class I alloantigen in vivo. Sublethal gamma-irradiation of primed mice eliminated accelerated rejection of skin allografts but failed to induce tolerance. Accelerated rejection was restored in irradiated mice by infusion of bone marrow cells expressing the priming alloantigen on immunostimulatory APCs (dendritic cells), whereas the memory response was not restored by infusion of bone marrow cells expressing the priming alloantigen on nonstimulatory APCs (B cells). Strikingly, irradiated mice infused with nonstimulatory bone marrow APCs exhibited long-term survival or tolerance to skin grafts expressing the priming MHC class I alloantigen. The mechanism of tolerance in this setting is explored. PMID- 10528166 TI - Natural killing of xenogeneic cells mediated by the mouse Ly-49D receptor. AB - NK lymphocytes lyse certain xenogeneic cells without prior sensitization. The receptors by which NK cells recognize xenogeneic targets are largely uncharacterized but have been postulated to possess broad specificity against ubiquitous target ligands. However, previous studies suggest that mouse NK cells recognize xenogeneic targets in a strain-specific manner, implicating finely tuned, complex receptor systems in NK xenorecognition. We speculated that mouse Ly-49D, an activating NK receptor for the MHC I ligand, H2-Dd, might display public specificities for xenogeneic target structures. To test this hypothesis, we examined the lysis of xenogeneic targets by mouse Ly-49D transfectants of the rat NK cell line RNK-16 (RNK. Ly-49D). Of the xenogeneic tumor targets tested, RNK.Ly-49D, but not untransfected RNK-16, preferentially lysed tumor cells derived from Chinese hamsters and lymphoblast targets from rats. Ly-49D-dependent recognition of Chinese hamster cells was independent of target N-linked glycosylation. Mouse Ly-49D also specifically stimulated the natural killing of lymphoblast targets derived from wild-type and MHC-congenic rats of the RT1lv1 and RT1l haplotypes, but not of the RT1c, RT1u, RT1av1, or RT1n haplotypes. These studies demonstrate that Ly-49D can specifically mediate cytotoxicity against xenogeneic cells, and they suggest that Ly-49D may recognize xenogeneic MHC encoded ligands. PMID- 10528168 TI - Switching to IgG3, IgG2b, and IgA is division linked and independent, revealing a stochastic framework for describing differentiation. AB - LPS was used to induce switching of B cells to IgG3 and, in the presence of TGF beta, to IgG2b and IgA. Switching to all three isotypes increased with division number according to a consistent relationship that was independent of time in culture. The mode of activation altered the relationship with division, as CD40 ligand increased switching to IgA and decreased switching to IgG2b and IgG3 when measured per division. This division-linked switching behavior could be described by Gaussian probability distributions centered around a mean division number. The divisions at which switching to IgG3 and IgG2b occurred overlapped, raising the possibility that the two switching mechanisms were linked. However, when IgG3+ and IgG3- B cells were sorted and placed back in culture, they switched to IgG2b at an equivalent rate, indicating that alternative switching decisions were made independently within a single cell. As a consequence, isotype switching could be predicted at the population level by standard probability laws. Therefore, division number provides a framework for a stochastic description of differentiation that may be widely applicable. PMID- 10528169 TI - Macrophage inflammatory protein-3 beta enhances IL-10 production by activated human peripheral blood monocytes and T cells. AB - We report that the addition of human macrophage inflammatory protein-3 beta (MIP 3 beta) to cultures of human PBMCs that have been activated with LPS or PHA results in a significant enhancement of IL-10 production. This effect was concentration-dependent, with optimal MIP-3 beta concentrations inducing more than a 5-fold induction of IL-10 from LPS-stimulated PBMCs and a 2- to 3-fold induction of IL-10 from PHA-stimulated PBMCs. In contrast, no significant effect on IL-10 production was observed when 6Ckine, the other reported ligand for human CCR7, or other CC chemokines such as monocyte chemoattractant protein-1, RANTES, MIP-1 alpha, and MIP-1 beta were added to LPS- or PHA-stimulated PBMCs. Similar results were observed using activated purified human peripheral blood monocytes or T cells. Addition of MIP-3 beta to nonactivated PBMCs had no effect on cytokine production. Enhancement of IL-10 production by MIP-3beta correlated with the inhibition of IL-12 p40 and TNF-alpha production by monocytes and with the impairment of IFN-gamma production by T cells, which was reversed by addition of anti-IL-10 Abs to the cultures. The ability of MIP-3 beta to augment IL-10 production correlated with CCR7 mRNA expression and stimulation of intracellular calcium mobilization in both monocytes and T cells. These data indicate that MIP 3 beta acts directly on human monocytes and T cells and suggest that this chemokine is unique among ligands binding to CC receptors due to its ability to modulate inflammatory activity via the enhanced production of the anti inflammatory cytokine IL-10. PMID- 10528170 TI - In vivo priming by DNA injection occurs predominantly by antigen transfer. AB - DNA vaccines can stimulate both humoral and cytolytic immune responses. Although bone marrow-derived elements present the expressed Ag, the mechanisms for acquiring immunogenic peptides have yet to be fully elucidated. APCs may become directly transfected by plasmid DNA or process extracellular proteins produced by other transfected cells. Using a transactivating plasmid system and bone marrow chimeras, we show that both mechanisms appear to be involved; however, the bulk of the immune response is dependent on expression of Ag by nonlymphoid tissues and transfer to APCs. These in vivo studies are the first to define the role of transfected nonlymphoid cells in generating Ag for presentation by bone marrow derived APCs after needle injection with plasmid DNA. PMID- 10528171 TI - Normal induction of oral tolerance in the absence of a functional IL-12-dependent IFN-gamma signaling pathway. AB - There is considerable evidence that regulatory cytokines play an important role in mediating the systemic tolerance found after oral administration of protein Ags. Although most existing work has focused on cytokines such as IL-4, IL-10, and TGF-beta, recent evidence from TCR transgenic systems suggests that the induction of oral tolerance is accompanied by priming of Ag-specific IFN-gamma production. IFN-gamma has also been implicated as a mediator of T cell tolerance in other models in vivo and in vitro, including that induced by aerosol administration of protein. We show here that feeding tolerogenic doses of OVA primes for IFN-gamma production in the spleen of mice with a normal T cell repertoire. However, depleting IFN-gamma at the time of feeding OVA had no effect on the induction of tolerance. In addition, tolerance was induced normally in both IFN-gamma receptor knockout (IFN-gammaR-/-) and IL-12 p40 knockout (IL-12-/ ) mice. This was the case for all components of the systemic immune response and also with a variety of feeding protocols, including those believed to induce distinct regulatory mechanisms. We conclude that IL-12-dependent IFN-gamma mediated regulation does not play an essential role in oral tolerance. PMID- 10528172 TI - Protein kinase C inhibits CD95 (Fas/APO-1)-mediated apoptosis by at least two different mechanisms in Jurkat T cells. AB - We have recently reported that activation of protein kinase C (PKC) plays a negative role in CD95-mediated apoptosis in human T cell lines. Here we present data indicating that although the PKC-induced mitogen-activated protein kinase pathway could be partially implicated in the abrogation of CD95-mediated apoptosis by phorbol esters in Jurkat T cells, the major inhibitory effect is exerted through a PKC-dependent, mitogen-activated protein kinase-independent signaling pathway. Furthermore, we demonstrate that activation of PKC diminishes CD95 receptor aggregation elicited by agonistic CD95 Abs. On the other hand, it has been reported that UV radiation-induced apoptosis is mediated at least in part by the induction of CD95 oligomerization at the cell surface. Here we show that activation of PKC also inhibits UVB light-induced CD95 aggregation and apoptosis in Jurkat T cells. These results reveal a novel mechanism by which T cells may restrain their sensitivity to CD95-induced cell death through PKC mediated regulation of CD95 receptor oligomerization at the cell membrane. PMID- 10528173 TI - Age-associated rapid and Stat6-independent IL-4 production by NK1-CD4+8- thymus T lymphocytes. AB - The source of IL-4 required for priming naive T cells into IL-4-secreting effectors has not been clearly identified. Here we show that upon TCR stimulation, thymus NK1-CD4+8- T cells produced IL-4, the magnitude of which was inversely correlated with age. This IL-4 production response by Th2-prone BALB/c mice was approximately 9-fold that of Th1-prone C57BL/10 mice. More than 90% of activated NK1-CD4+8- thymocytes did not use the invariant V alpha 14-J alpha 281 chain characteristic of typical CD1-restricted NK1+CD4+ T cells. Stat6-null NK1 CD4+8- thymocytes produced bioactive IL-4, with induction of IL-4 mRNA expression within 1 h of stimulation. Our results support the possibility that TCR repertoire-diverse conventional NK1-CD4+ T cells are a potential IL-4 source for directing naive T cells toward Th2/type 2 CD8+ T cell (Tc2) effector development. PMID- 10528174 TI - Presence of CpG DNA and the local cytokine milieu determine the efficacy of suppressive DNA vaccination in experimental autoimmune encephalomyelitis. AB - We here study the adjuvant properties of immunostimulatory DNA sequences (ISS) and coinjected cytokine-coding cDNA in suppressive vaccination with DNA encoding an autoantigenic peptide, myelin basic protein peptide 68-85, against Lewis rat experimental autoimmune encephalomyelitis (EAE). EAE is an autoaggressive, T1 mediated disease of the CNS. ISS are unmethylated CpG motifs found in bacterial DNA, which can induce production of type 1 cytokines in vertebrates through the innate immune system. Because ISS in the plasmid backbone are necessary for efficient DNA vaccination, we studied the effect of one such ISS, the 5'-AACGTT 3' motif, in our system. Treatment with a DNA vaccine encoding myelin basic protein peptide 68-85 and containing three ISS of 5'-AACGTT-3' sequence suppressed clinical signs of EAE, while a corresponding DNA vaccine without such ISS had no effect. We further observed reduced proliferative T cell responses in rats treated with the ISS-containing DNA vaccine, compared with controls. We also studied the possible impact of coinjection of plasmid DNA encoding rat cytokines IL-4, IL-10, GM-CSF, and TNF-alpha with the ISS-containing DNA vaccine. Coinjection of IL-4-, IL-10-, or TNF-alpha-coding cDNA inhibited the suppressive effect of the DNA vaccine on EAE, whereas GM-CSF-coding cDNA had no effect. Coinjection of cytokine-coding cDNA with the ISS-deficient DNA vaccine failed to alter clinical signs of EAE. We conclude that the presence of ISS and induction of a local T1 cytokine milieu is decisive for specific protective DNA vaccination in EAE. PMID- 10528175 TI - p38 mitogen-activated protein kinase regulates human T cell IL-5 synthesis. AB - Involvement of p38 mitogen-activated protein (MAP) kinase in human T cell cytokine synthesis was investigated. p38 MAP kinase was clearly induced in human Th cells activated through the TCR. SB203580, a highly selective inhibitor of p38 MAP kinase, inhibited the induction of p38 MAP kinase in human Th cells. Major T cell cytokines, IL-2, IL-4, IL-5, and IFN-gamma, were produced by Der f 2 specific Th clones upon stimulation through the TCR. IL-5 synthesis alone was significantly inhibited by SB203580 in a dose-dependent manner, whereas the production of IL-2, IL-4, and IFN-gamma was not affected. The proliferation of activated T cells was not affected. IL-5 synthesis of human Th clones induced upon stimulation with rIL-2, phorbol ester plus anti-CD28 mAb, and immobilized anti-CD3 mAb plus soluble anti-CD28 mAb was also suppressed by SB203580 in the same concentration response relationship. The results clearly indicated that IL-5 synthesis by human Th cells is dependent on p38 MAP kinase activity, and is regulated distinctly from IL-2, IL-4, and IFN-gamma synthesis. Selective control of IL-5 synthesis will provide a novel treatment devoid of generalized immune suppression for bronchial asthma and atopic dermatitis that are characterized by eosinophilic inflammation. PMID- 10528176 TI - Selective up-regulation of phosphatidylinositol 3'-kinase activity in Th2 cells inhibits caspase-8 cleavage at the death-inducing complex: a mechanism for Th2 resistance from Fas-mediated apoptosis. AB - In this study the mechanism of differential sensitivity of CD3-activated Th1- and Th2-type cells to Fas-mediated apoptosis was explored. We show that the Fas associated death domain protein (FADD)/caspase-8 pathway is differentially regulated by CD3 activation in the two subsets. The apoptosis resistance of activated Th2-type cells is due to an incomplete processing of caspase-8 at the death-inducing signaling complex (DISC) whereas recruitment of caspase-8 to the DISC of Th1- and Th2-like cells is comparable. Activation of phosphatidylinositol 3'-kinase upon ligation of CD3 in Th2-type cells blocked caspase-8 cleavage to its active fragments at the DISC, thereby preventing induction of apoptosis. This study offers a new pathway for phosphatidylinositol 3'-kinase in mediating protection from Fas-induced apoptosis. PMID- 10528177 TI - Visualization, fate, and pathogenicity of antigen-specific CD8+ T cells in the graft-versus-host reaction. AB - To follow the fate of alloreactive T cell effectors in graft-vs-host disease, Ld specific CD8+ T cells from C57BL/6 2C TCR-transgenic donors were transplanted into sublethally irradiated (750 cGy) Ld+ or Ld- recipients. In Ld- C57BL/6 or (BALB/c-dm2 x C57BL/6)F1 recipients, naive 2C T cells engrafted and survived long term, but did not acquire effector function. In Ld+ (BALB/c x C57BL/6)F1 recipients, 2C T cells engrafted, expanded, became cytolytic, destroyed host B cells and double-positive thymocytes, and later disappeared. Despite marked damage to lymphoid and hemopoietic cells by 2C T cells, no significant pathology was detected in other organs, and recipients survived. Ld+ (BALB/c x C57BL/6)F1 recipients died when LPS/endotoxin was administered on day 7 after cell transfer, while Ld- (BALB/c-dm2 x C57BL/6)F1 recipients survived. Our findings show that under certain conditions, a CD8+ T cell population recognizing an extremely limited repertoire of Ags can initiate graft-vs-host disease. PMID- 10528178 TI - Emergence of T, B, and myeloid lineage-committed as well as multipotent hemopoietic progenitors in the aorta-gonad-mesonephros region of day 10 fetuses of the mouse. AB - We investigated the developmental potential of hemopoietic progenitors in the aorta-gonad-mesonephros (AGM) region, where the definitive type hemopoietic progenitors have been shown to emerge before the fetal liver develops. By using an assay system that is able to determine the developmental potential of individual progenitors toward the T, B, and myeloid lineages, we show that not only multipotent progenitors but also progenitors committed to the T, B, or myeloid lineage already exist in this region of day 10 fetuses. Bipotent progenitors generating myeloid and T cells or those generating myeloid and B cells were also detected, suggesting that the commitment to T and B cell lineages is in progress in the AGM region. The numbers of these progenitors, however, were only 1/200-1/1000 of those in fetal liver of day 12 fetuses. Such small numbers of progenitors suggest that hemopoiesis has just started in the AGM region of day 10 fetuses. Although most of T cell lineage-committed progenitors in the AGM region generated only a small number of immature T cells, some were able to generate a large number of mature T cells. The detection of various types of lineage-committed progenitors strongly suggests that the AGM region is not only the site of stem cell emergence, but also the site of hemopoiesis, including lineage commitment. The T cell progenitors found in the AGM region may represent the first immigrants to the thymus anlage. PMID- 10528179 TI - Peripheral CD4+ T cell maturation recognized by increased expression of Thy 1/CD90 bearing the 6C10 carbohydrate epitope. AB - The SM6C10 IgM autoantibody recognizes a surface determinant, 6C10, that is highly expressed on all immature thymocytes. In contrast, its expression on peripheral T cells appears developmentally regulated, i.e., absent from most naive T cells in spleen of neonatal mice, but expressed on 40-80% of naive CD4+ T cells in adult. In this paper, we demonstrate that SM6C10 recognizes a carbohydrate epitope on the Thy-1 glycoprotein using immunoprecipitation analysis, by binding to affinity-purified Thy-1 in an ELISA, and by sensitivity to N-glycosidase-F treatment. Retroviral Thy-1 gene transduction experiments into Thy-1- variant T cell lines and a pro-B cell line provide evidence that 6C10 glycosylated Thy-1 expression is not restricted to T cells but depends on the recipient cell. Therefore, differences in 6C10 levels among Thy-1+ T cells in mice likely reflect developmental regulation of posttranslational modification of the Thy-1 glycoprotein. The ability of naive CD4+ T cells to respond to anti-Thy 1 stimulation increases from neonate to adult, and 6C10- naive cells from adult mice respond poorly compared with 6C10+ cells, similar to the cells in neonatal mice. These results suggest that there is functional maturation by peripheral CD4+ T cells that coincides with 6C10 glycosylated Thy-1 up-regulation, and natural autoantibody recognizes this 6C10 carbohydrate epitope. PMID- 10528180 TI - CD40 ligand blockade induces CD4+ T cell tolerance and linked suppression. AB - The CD40-CD40 ligand (CD40L) interaction is a key event in the initiation of an adaptive immune response, and as such the therapeutic value of CD40L blockade has been studied in many experimental models of tissue transplantation and autoimmune disease. In rodents, transplantation of allogeneic tissues under the cover of anti-CD40L Abs has resulted in prolonged graft survival but not tolerance. In this report, we show that failure to induce tolerance probably results from the inability of anti-CD40L Abs to prevent graft rejection elicited by the CD8+ T cell subset. When the CD8+ T cell population is controlled independently, using anti-CD8 Abs, then tolerance is possible. Transplantation tolerance induced by anti-CD4 mAbs can often be associated with dominant regulation, manifested as infectious tolerance and linked suppression, both of which are mediated by CD4+ T cells. We show here that CD4+ T cells rendered tolerant using anti-CD40L therapy exhibit the same regulatory property of linked suppression, as demonstrated by their ability to accept grafts expressing third party Ags only if they are expressed in conjunction with the tolerated Ags. This observation of linked suppression reveals a hitherto undocumented consequence of CD40L blockade that suggests the tolerant state is maintained by a dominant regulatory mechanism. Our results suggest that, although anti-CD40L Abs are attractive clinical immunotherapeutic agents, additional therapies to control aggressive CD8+ T cell responses may be required. PMID- 10528181 TI - Differential requirements for CD4 in TCR-ligand interactions. AB - The coreceptor molecule, CD4, plays an integral part in T cell activation; it is involved in both extracellular Ag recognition and intracellular signaling. We wanted to examine the functional role of CD4 in the recognition of agonist and altered peptide ligands (APLs). We generated two CD4-deficient T cell lines expressing well-characterized TCRs specific for Hb(64-76)/I-Ek. Although the responsiveness of the T cell lines to the agonist peptide was differently affected by the loss of CD4 expression, the recognition of APLs was in both cases dramatically reduced. Nearly full responsiveness to the agonist peptide was achieved by expression of a CD4 variant that did not associate with p56lck; however, the stimulation by APLs was only partially restored. Importantly, the expression of a CD4 variant in which domains interacting with MHC class II molecules have been mutated failed to restore the reactivity to all ligands. CD4 deficient T cells were able to be antagonized by APLs, indicating that CD4 was not required for antagonism. Overall, these findings support the concepts that CD4 is an integral part of the initial formation of the immunological synapse, and that the requirement for different CD4 functions in T cell activation varies depending upon the potency of the ligand. PMID- 10528182 TI - Mutual regulation between B7-1 (CD80) expressed on T cells and IL-4. AB - We have used T cells from B7-1-deficient TCR transgenic DO11.10 mice to demonstrate a functional role for B7-1 on T cells. B7-1-deficient DO11.10 T cells produce more IL-4 than wild-type DO11.10 T cells, suggesting that B7-1 expressed by T cells regulates the differentiation of IL-4-producing cells. In addition, we found that IL-4 inhibits B7-1 expression by wild-type DO11.10 T cells. Our results suggest that there is a reciprocal relationship between B7-1 expressed on T cells and IL-4 production, which results in a modulatory feedback loop. When high levels of IL-4 are produced by T cells, B7-1 expression by T cells is inhibited, which allows amplification of IL-4 production by these T cells. When low levels of IL-4 are produced by T cells, B7-1 expression by these T cells is increased, and a further reduction in IL-4 production follows. However, in addition to being influenced by IL-4, B7-1 expression by T cells is affected by peptide concentration and by B7 costimulation from APCs. The studies presented here demonstrate that B7-1 on T cells as well as on APCs regulates IL-4 production. However, whereas B7-1 expression on APCs can promote IL-4 production, IL-4 production is inhibited by B7-1 on T cells. PMID- 10528183 TI - Defective CD8+ T cell activation and cytolytic function in the absence of LFA-1 cannot be restored by increased TCR signaling. AB - Signaling through the TCR as well as engagement of costimulatory molecules are required for efficient T cell activation and progression into differentiated effector cells. The beta2 integrin LFA-1 (CD11a/CD18) has been implicated in TCR costimulation as well as in cell-cell adhesion function, but its exact role is still ambiguous. The present study focuses on the requirement for LFA-1 in CD8+ T cell activation and effector function using LFA-1-deficient cells expressing the 2C transgenic TCR as a model system. The lack of LFA-1 expression in 2C T cells resulted in severely diminished proliferative response toward allogeneic BALB/c splenocytes. Increase in TCR signaling alone by pulsing stimulators with high affinity peptides, p2Ca or QL9, had minimal effects in restoring proliferation. Addition of exogenous IL-2, however, enhanced the effect of peptide pulsing on proliferation of LFA-1-deficient 2C T cells. LFA-1-deficient 2C CTLs generated from alloantigen stimulation exhibited a defective cytotoxic activity when tested on a variety of target cells. Cytolysis could be improved, but not fully rectified by peptide pulsing of target cells. Thus, in the 2C TCR model, LFA-1 has a requisite role for optimal CD8+ T cell activation and effector function, which cannot be overcome by increasing peptide/MHC density on either the APCs or target cells, respectively. PMID- 10528184 TI - Analysis of 4-1BB ligand (4-1BBL)-deficient mice and of mice lacking both 4-1BBL and CD28 reveals a role for 4-1BBL in skin allograft rejection and in the cytotoxic T cell response to influenza virus. AB - 4-1BB ligand (4-1BBL) is a member of the TNF family expressed on activated APC. 4 1BBL binds to 4-1BB (CD137) on activated CD4 and CD8 T cells and in conjunction with strong signals through the TCR provides a CD28-independent costimulatory signal leading to high level IL-2 production by primary resting T cells. Here we report the immunological characterization of mice lacking 4-1BBL and of mice lacking both 4-1BBL and CD28. 4-1BBL-/- mice mount neutralizing IgM and IgG responses to vesicular stomatitis virus that are indistinguishable from those of wild-type mice. 4-1BBL-/- mice show unimpaired CTL responses to lymphocytic choriomeningitis virus (LCMV) and exhibit normal skin allograft rejection but have a weaker CTL response to influenza virus than wild-type mice. 4-1BBL-/-CD28 /- mice retain the CTL response to LCMV, respond poorly to influenza virus, and exhibit a delay in skin allograft rejection. In agreement with these in vivo results, allogeneic CTL responses of CD28-/- but not CD28+/+ T cells to 4-1BBL expressing APC are substantially inhibited by soluble 4-1BB receptor as is the in vitro secondary response of CD28+ T cells to influenza virus peptides. TCR transgenic CD28-/- LCMV glycoprotein-specific T cells are insensitive to the presence of 4-1BBL when a wild-type peptide is used, but the response to a weak agonist peptide is greatly augmented by the presence of 4-1BBL. These results further substantiate the idea that different immune responses vary in their dependence on costimulation and suggest a role for 4-1BBL in augmenting suboptimal CTL responses in vivo. PMID- 10528185 TI - The Th1/Th2 nature of concurrent immune responses to unrelated antigens can be independent. AB - We tested the independence hypothesis, namely that the Th1/Th2 nature of concurrent immune responses, generated in the same secondary lymphoid organ to non-cross-reacting Ags, can be independently determined. Some infectious agents and some adjuvants contain modulatory molecules that affect the Th1/Th2 nature of immune responses in a non-Ag-specific manner. We therefore excluded infectious agents as Ags and the use of adjuvants to generate immune responses. We first show that the dose of xenogeneic RBC administered i.v. determines the Th1/Th2 nature of the splenic immune response. Low doses generate a virtually exclusive Th1 response, whereas a higher dose induces either a mixed Th1/Th2 or a predominantly Th2 response, and stimulates the production of specific Abs. We immunized individual mice simultaneously with a low dose of one kind of xenogeneic RBC and with a higher dose of another non-cross-reacting xenogeneic RBC and assessed the Th1/Th2 nature of the immune responses generated in the spleen to each kind of RBC. The Th1/Th2 nature of the response to each RBC in doubly immunized mice was indistinguishable from that of the corresponding immune response in singly immunized mice. We discuss the significance of our findings for understanding immune class regulation, and the possible reasons why such independence is not always seen. PMID- 10528186 TI - Thymocyte antigens do not induce tolerance in the CD4+ T cell compartment. AB - Thymocytes fail to tolerize the developing T cell repertoire to self MHC class I (MHC I) Ags because transgenic (CD2Kb) mice expressing H-2Kb solely in lymphoid cell lineages reject skin grafts mismatched only for H-2Kb. In this study, we examined why thymocytes fail to tolerize the T cell repertoire to self MHC I Ags. The ability of CD2Kb mice to reject H-2Kb skin grafts was age dependent because CD2Kb mice older than 20 wk accepted skin grafts. T cells from younger CD2Kb mice proliferated, but did not develop cytotoxic functions in vitro in response to H 2Kb. Proliferative responses were dominated by H-2Kb-specific, CD4+ T cells rather than CD8+ T cells. Representative CD4+ T cell clones from CD2Kb mice were MHC II restricted and recognized processed H-2Kb. TCR transgenic mice were generated from one CD4+ T cell clone (361) to monitor development of H-2Kb specific immature thymocytes when all thymic cells or lymphoid cell lineages only expressed H-2Kb. Thymocyte precursors were not eliminated and mice were not tolerant to H-2Kb when Tg361 TCR transgenic mice were intercrossed with CD2Kb mice. In contrast, all thymocyte precursors were eliminated efficiently in thymic microenvironments in which all cells expressed H-2Kb. We conclude that self MHC I Ags expressed exclusively in thymocytes do not induce T cell tolerance because presentation of processed self MHC I Ags on self MHC II molecules fails to induce negative selection of CD4+ T cell precursors. This suggests that some self Ags are effectively compartmentalized and cannot induce self-tolerance in the T cell repertoire. PMID- 10528187 TI - 4-1BB ligand, a member of the TNF family, is important for the generation of antiviral CD8 T cell responses. AB - 4-1BB (CD137) is a costimulatory molecule expressed on activated T cells and interacts with 4-1BB ligand (4-1BBL) on APCs. To investigate the role of 4-1BB costimulation for the development of primary immune responses, 4-1BBL-deficient (4-1BBL-/-) mice were infected with lymphocytic choriomeningitis virus (LCMV). 4 1BBL-/- mice were able to generate CTL and eliminate acute LCMV infection with normal kinetics, but CD8 T cell expansion was 2- to 3-fold lower than in wild type (+/+) mice. In the same mice, virus-specific CD4 Th and B cell responses were minimally affected, indicating that 4-1BB costimulation preferentially affects CD8 T cell responses. This result contrasts with our earlier work with CD40L-deficient (CD40L-/-) mice, in which the CD8 T cell response was unaffected and the CD4 T cell response was markedly impaired. When both 4-1BBL- and B7 dependent signals were absent, CD8 T cell expansion was further reduced, resulting in lower CTL activity and impairing their ability to clear LCMV. Altogether, these results indicate that T cells have distinct costimulatory requirements: optimal CD8 responses require 4-1BBL-dependent interactions, whereas CD4 responses are minimally affected by 4-1BB costimulation, but require CD40-CD40L and B7-dependent interactions. PMID- 10528188 TI - CD34+ cell-derived CD14+ precursor cells develop into Langerhans cells in a TGF beta 1-dependent manner. AB - Langerhans cells (LC) are CD1a+E-cadherin (E-cad)+Birbeck granule+ but CD11b-CD36 factor XIIIa (FXIIIa)- members of the dendritic cell (DC) family. Evidence holds that LC originate from CD1a+CD14- rather than CD14+CD1a- progenitors, both of which arise from GM-CSF/TNF-alpha-stimulated CD34+ stem cells. The CD14+CD1a- progenitors, on the other hand, can give rise to a separate DC type characterized by its CD1a+CD11b+CD36+FXIIIa+E-cad-BG- phenotype (non-LC DC). Although GM CSF/TNF-alpha are important for both LC and non-LC DC differentiation, TGF-beta 1 is thought to preferentially promote LC development in vitro and in vivo. However, the hemopoietic biology of this process and the nature of TGF-beta 1 responsive LC precursors (LCp) are not well understood. Here we show that CD14+ precursors in the presence, but not in the absence, of TGF-beta 1 give rise to a progeny that fulfills all major criteria of LC. In contrast, LC development from CD1a+ progenitors was TGF-beta 1 independent. Further studies revealed that CD14+ precursors contain a CD11b+ and a CD11b- subpopulation. When either subset was stimulated with GM-CSF/TNF-alpha and TGF-beta 1, only CD14+CD11b- cells differentiated into LC. The CD11b+ cells, on the other hand, acquired non-LC DC features only. The higher doubling rates of cells entering the CD14+ LCp rather than the CD1a+ LCp pathway add to the importance of TGF-beta 1 for LC development. Because CD14+CD11b- precursors are multipotent cells that can enter LC or macrophage differentiation, it is suggested that these cells, if present at the tissue level, endow a given organ with the property to generate diverse cell types in response to the local cytokine milieu. PMID- 10528189 TI - T cell infiltration into class II MHC-disparate allografts and acute rejection is dependent on the IFN-gamma-induced chemokine Mig. AB - Direct evidence that cytokines with chemoattractant properties for leukocytes, chemokines, recruit alloantigen-primed T cells into transplanted allografts has been lacking. We present evidence that neutralization of a single chemokine inhibits T cell infiltration into class II MHC-disparate murine allografts and acute rejection. The chemokines IFN-gamma-inducible protein-10 and monokine induced by IFN-gamma (Mig) are expressed in allogeneic skin grafts during the late stages of acute rejection. Survival of class II MHC-disparate B6.H-2bm12 allografts is prolonged from day 14 to day 55 posttransplant when C57BL/6 recipients are given a short course treatment with an antiserum to Mig. This treatment also inhibits T cell and macrophage infiltration into the allografts. B6.H-2bm12 allografts are also not rejected by IFN-gamma-/- C57BL/6 recipients. Injection of Mig directly into B6.H-2bm12 grafts on IFN-gamma-deficient recipients restores T cell infiltration and rejection. Therefore, the inability of IFN-gamma-deficient recipients to reject the class II MHC-disparate allografts is due to the lack of intraallograft Mig production and alloantigen-primed T cell recruitment to the graft. These results indicate for the first time the potential utility of chemokine neutralization strategies in preventing T cell infiltration into allografts and abrogating acute rejection. PMID- 10528190 TI - A "stealth effect": adenocarcinoma cells engineered to express TRAIL elude tumor specific and allogeneic T cell reactions. AB - BALB/c mammary adenocarcinoma cells engineered to express TNF-related apoptosis inducing ligand (TRAIL)/APO-2 ligand (APO-2L) on their membrane (TSA-TRAIL) grow with kinetics similar to that of parental cells (TSA-pc) in vitro and in nu/nu mice. In contrast, TSA-TRAIL cells grow faster than TSA-pc in normal BALB/c mice. In DBA/2 mice, which differ from BALB/c mice at minor histocompatibility Ags, they also grow faster and display a higher percentage of tumor takes than TSA-pc. In fully histoincompatible C57BL/6 (B6) mice, TSA-TRAIL cells form evident tumors that are slowly rejected by most mice, but outgrow in a few. In contrast, TSA-pc cells are rejected at once by B6 mice. Since TRAIL/APO-2L induces apoptosis by interacting with a variety of specific receptors, this rapid growth in both syngeneic and allogeneic mice may be the result of an immunosuppressive mechanism. The following evidence supports this hypothesis: 1) TSA-TRAIL cells overcome the strong immunity against TSA-pc cells elicited in BALB/c mice by preimmunization with TSA cells engineered to release IL-4; 2) their rejection by B6 mice does not prime a CTL-mediated memory; 3) thymidine uptake by T lymphocytes unstimulated or stimulated by allogeneic cells is inhibited when TSA TRAIL cells are added as third party cells; 4) CTL kill TSA-pc but not TSA-TRAIL cells in 48-h assays; and 5) activated lymphocytes interacting with TSA-TRAIL cells in vivo and in vitro undergo apoptosis. PMID- 10528191 TI - Influence of lymphocytes on the presence and organization of dendritic cell subsets in the spleen. AB - Studies were undertaken to clarify the roles of individual leukocyte populations in maintaining the presence and organization of splenic dendritic cells (DCs). Using Abs specific for DC subsets, we found that the distinct types of DC maintained appropriate compartmentalization within the white pulp of lymphocyte deficient mice despite an unusual overall distribution of DCs. Even in mice lacking both B and T lymphocytes, the central arteriole remained the structure around which T area DCs were organized. Marginal zone area DCs remained in a peripheral sheath excluded from the T area DCs. Additionally, we revealed an important role for splenic B cells in the presence and organization of marginal zone cells. B-deficient or B- and T-deficient mice lacked sialoadhesin+ marginal zone macrophages and lacked MAdCAM-1 expression in marginal zone reticular endothelial cells. Adoptive transfer of B lymphocytes induced MAdCAM-1 expression but failed to recruit marginal zone macrophages. Taken together, our results demonstrate that the arrival, localization, and persistence of DCs in spleen are events not solely dependent upon signals from the mature B and T cells or marginal zone macrophages. We suggest that specific stromal elements in the vicinity of the central arteriole are primarily responsible for providing directional cues to the DC. PMID- 10528192 TI - A novel role for H-Ras in the regulation of very late antigen-4 integrin and VCAM 1 via c-Myc-dependent and -independent mechanisms. AB - Despite extensive studies on the crucial functions of Ras and c-Myc in cellular proliferation and transformation, their roles in regulating cell adhesion are not yet fully understood. Involvement of Ras in modulating integrin activity by inside-out signaling has been recently reported. However, in contrast to R-Ras, H Ras was found to exhibit a suppressive effect. Here we show that ectopic expression of a constitutively active H-Rasv12, but not c-Myc alone, in a hemopoietic cell line induces activation of very late Ag-4 (VLA-4, alpha4beta1) integrin without changing its surface expression. Intriguingly, coexpression of H Rasv12 and c-Myc in these cells results in not only the activation of VLA-4, but also the induction of expression of VCAM-1, the counterreceptor for VLA-4, thereby mediating a marked homotypic cell aggregation. In addition, H-Rasv12 induced VLA-4 activation appears to be partly down-regulated by coexpression with c-Myc. Our results represent an unprecedented example demonstrating a novel role for H-Rasv12 in the regulation of cell adhesion via c-Myc-independent and dependent mechanisms. PMID- 10528193 TI - In vivo evidence that caspase-3 is required for Fas-mediated apoptosis of hepatocytes. AB - Caspase-3 is essential for Fas-mediated apoptosis in vitro. We investigated the role of caspase-3 in Fas-mediated cell death in vivo by injecting caspase-3 deficient mice with agonistic anti-Fas Ab. Wild-type controls died rapidly of fulminant hepatitis, whereas the survival of caspase-3-/- mice was increased due to a delay in hepatocyte cell death. Bcl-2 expression in the liver was dramatically decreased in wild-type mice following anti-Fas injection, but was unchanged in caspase-3-/- mice. Hepatocytes from anti-Fas-injected wild-type, but not caspase-3-/-, mice released cytochrome c into the cytoplasm. Western blotting confirmed the lack of caspase-3-mediated cleavage of Bcl-2. Presumably the presence of intact Bcl-2 in caspase-3-/- hepatocytes prevents the release of cytochrome c from the mitochondria, a required step for the mitochondrial death pathway. We also show by Western blot that Bcl-xL, caspase-9, caspase-8, and Bid are processed by caspase-3 in injected wild-type mice but that this processing does not occur in caspase-3-/- mice. This study thus provides novel in vivo evidence that caspase-3, conventionally known for its downstream effector function in apoptosis, also modifies Bcl-2 and other upstream proteins involved in the regulation of Fas-mediated apoptosis. PMID- 10528194 TI - CD44-deficient mice develop normally with changes in subpopulations and recirculation of lymphocyte subsets. AB - Cell adhesion molecules are considered to be pivotal elements required for proper embryo development. The transmembrane glycoprotein CD44, which is expressed in numerous splice variants on the surface of many different cell types and tissues, has been suggested to be involved in several physiological processes such as cell cell interactions, signal transduction, and lymphocyte homing and trafficking during embryogenesis and in the adult organism. Some splice variants are thought to play an important role in tumor progression. To investigate the physiological roles of CD44 in vivo, we abolished expression of all isoforms of CD44 in mice by targeted insertion of a lacZ/neo cassette into the reading frame of the leader peptide. CD44-deficient mice are viable without obvious developmental defects and show no overt abnormalities as adults. However, CD44-deficient lymphocytes exhibit impaired entry into the adult thymus, although lymphocyte development is apparently unaltered. Our data indicate that all splice variants of CD44 are dispensable for embryonic development and implicate a critical function for CD44 in lymphocyte recirculation. PMID- 10528195 TI - Distinct mechanisms target stress and extracellular signal-activated kinase 1 and Jun N-terminal kinase during infection of macrophages with Salmonella. AB - The interaction between bacteria and macrophages is central to the outcome of Salmonella infections. Salmonella can escape killing by these phagocytes and survive and multiply within them, giving rise to chronic infections. Cytokines produced by infected macrophages are involved in the early gastrointestinal pathology of the infection as well as in the induction and maintenance of the immune response against the invaders. Jun N-terminal kinases (JNK) are activated by inflammatory stimuli and play a role in cytokine production. We have investigated the signaling routes leading to JNK activation in Salmonella infected macrophages and have discovered that they differ radically from the mechanisms operating in epithelial cells. In particular, activation of the JNK kinase stress and extracellular-activated kinase 1 (SEK1) and of JNK in macrophages occurs independently of actin rearrangements and of the GTPases Cdc42 and Rac, essential mediators in other cells. Activation of JNK is effected by a novel pathway comprising tyrosine kinase(s), phosphoinositide 3-kinase and, likely, atypical protein kinase C zeta. SEK1 is stimulated by a distinct mechanism involving phosphatidylcholine-phospholipase C and acidic sphingomyelinase. Dominant-negative SEK1 can block JNK activation by LPS, but not by Salmonella. These data demonstrate that SEK1 and JNK are activated independently in Salmonella-infected macrophages and offer experimental support for the concept that incoming signals can direct the selective coupling of downstream pathways to elicit highly specific responses. Inhibitors of stress kinase pathways are receiving increasing attention as potential anti-inflammatory drugs. The precise reconstruction of stimulus-specific pathways will be instrumental in predicting/evaluating the effects of the inhibitors on a given pathological condition. PMID- 10528197 TI - Quantitative alleles of CR1: coding sequence analysis and comparison of haplotypes in two ethnic groups. AB - The quantitative expression of complement receptor type 1 (CR1) on erythrocytes is regulated by two CR1 alleles that differ in having genomic HindIII fragments of either 7.4 or 6.9 kb and that determine high (H allele) or low (L allele) CR1 expression, respectively, across a 10-fold range. To investigate whether the product of the L allele may contain amino acid substitutions that make it more susceptible to proteolysis, cDNA sequence spanning the CR1 coding region was analyzed in two donors who were homozygous for the H and L alleles and differed by 7-fold in their mean numbers of CR1 per erythrocyte. Sequence differences were detected at 10 nucleotide positions, including 6 that would cause amino acid substitutions. The HindIII RFLP and 3 of the latter 6 sites were analyzed in genomic DNA of 85 Caucasians and 75 African Americans; sites encoding the other amino acid substitutions were analyzed less extensively. Two major haplotypes defined prototypic H and L alleles in both ethnic groups, suggesting that these alleles existed before the African and European populations diverged. Decreased erythrocyte CR1 expression is associated with impaired clearance of immune complexes from blood. Persistence of the L allele in all populations that have been analyzed may suggest a compensatory survival advantage, perhaps related to malaria or another infectious disease. PMID- 10528196 TI - Cloning and characterization of a novel activating Ly49 closely related to Ly49A. AB - The majority of the known Ly49 family members have been isolated from either C57BL/6 (B6) or BALB/c mice. Interestingly, the anti-Ly49 Ab reactivities observed in 129/J mice are different from those of B6 mice. Furthermore, immunoprecipitation of 129/J NK cell lysates with YE1/32 and YE1/48, Abs specific for the inhibitory Ly49A in B6, resulted in detection of the activation associated DAP12 molecule. These results indicated a need for a more detailed study of this strain. Therefore, a cloning strategy was devised to isolate Ly49 cDNAs from 129/J mice. An immunoreceptor tyrosine-based inhibitory motif containing, Ly49D-related clone was discovered that we have named Ly49O, and one immunoreceptor tyrosine-based inhibitory motif-lacking, Ly49A-related clone was discovered that we have named Ly49P. No anti-Ly49 mAb reacted with Ly49O, whereas the molecule encoded by the Ly49P cDNA was found to react with YE1/32 and YE1/48. Ly49P was found to associate with mouse DAP12, and Ab-mediated cross-linking of Ly49P resulted in mouse DAP12 phosphorylation and Ca2+ mobilization, indicating that Ly49P is a competent activation receptor. Ly49P, therefore, represents a novel member of the Ly49 activating receptor subfamily. PMID- 10528198 TI - Systematic mutagenesis of TCR complementarity-determining region 3 residues: a single conservative substitution dramatically improves response to both multiple HLA-DR alleles and peptide variants. AB - To define the relative contributions of HLA and peptide contacts with TCR complementarity-determining region (CDR) 3 residues in T cell recognition, systematic mutagenesis and domain swapping was conducted on two highly similar TCRs that both respond to the influenza hemagglutinin (HA) peptide, HA307-319, but with different HLA restrictions. Despite the primary sequence similarity of these TCRs, exchange of as little as two CDR3 residues between them completely abrogated responsiveness. At position 95 within CDR3alpha, various substitutions still allowed for some degree of recognition. One modest substitution, alanine for glycine (essentially the addition of a methyl group), significantly broadened the specificity of the TCR. Transfectants expressing this mutant TCR responded strongly in the context of multiple HLA-DR alleles and to HA peptide variants with substitutions at each TCR contact residue. These results suggest that the conformations of CDR3 loops are crucial to TCR specificity and that it may not be reliable to extrapolate from primary sequence similarities in TCRs to similarities in specificity. The ease with which a broad specificity is induced in this mutant TCR has implications for the mechanisms and frequency of alloreactivity and promiscuity in T cell responses. PMID- 10528199 TI - Molecular defects in variant forms of mannose-binding protein associated with immunodeficiency. AB - Distinct molecular mechanisms underlying immunodeficiency caused by three different naturally occurring point mutations within the collagen-like domain of human mannose-binding protein (MBP; also known as mannose-binding lectin) have been revealed by introduction of analogous mutations into rat serum MBP. The change Arg23-->Cys results in a lower proportion of the large oligomers most efficient at activating the complement cascade. The presence of cysteine at position 23, which forms aberrant interchain disulfide bonds, causes disruption of the normal oligomeric state. The deficiency in MBPs containing Gly25-->Asp and Gly28-->Glu substitutions also results in part from reduced formation of higher oligomers. However, decreased ability to interact with downstream components of the complement cascade due to changes in both the N-terminal disulfide-bonding arrangement and the local structure of the collagenous domain make more important contributions to the loss of activity in these mutants. PMID- 10528200 TI - Immune-associated nucleotide-1 (IAN-1) is a thymic selection marker and defines a novel gene family conserved in plants. AB - Positive selection of thymocytes is a complex and crucial event in T cell development that is characterized by cell death rescue, commitment toward the helper or cytotoxic lineage, and functional maturation of thymocytes bearing an appropriate TCR. To search for novel genes involved in this process, we compared gene expression patterns in positively selected thymocytes and their immediate progenitors in mice using the differential display technique. This approach lead to the identification of a novel gene, mIAN-1 (murine immune-associated nucleotide-1), that is switched on upon positive selection and predominantly expressed in the lymphoid system. We show that mIAN-1 encodes a 42-kDa protein sharing sequence homology with the pathogen-induced plant protein aig1 and that it defines a novel family of at least three putative GTP-binding proteins. Analysis of protein expression at various stages of thymocyte development links mIAN-1 to CD3-mediated selection events, suggesting that it represents a key player of thymocyte development and that it participates to peripheral specific immune responses. The evolutionary conservation of the IAN family provides a unique example of a plant pathogen response gene conserved in animals. PMID- 10528201 TI - An essential role for antibody in neutrophil and eosinophil recruitment to the cornea: B cell-deficient (microMT) mice fail to develop Th2-dependent, helminth mediated keratitis. AB - Invasion of the corneal stroma by neutrophils and eosinophils and subsequent degranulation disrupts corneal clarity and can result in permanent loss of vision. In the current study, we used a model of helminth-induced inflammation to demonstrate a novel role for Ab in mediating recruitment of these inflammatory cells to the central cornea. C57BL/6 and B cell-deficient (microMT) mice were immunized s. c. and injected intrastromally with Ags from the parasitic helminth Onchocerca volvulus (which causes river blindness). C57BL/6 mice developed pronounced corneal opacification, which was associated with an Ag-specific IL-5 response and peripheral eosinophilia, temporal recruitment of neutrophils and eosinophils from the limbal vessels to the peripheral cornea and subsequent migration to the central cornea. In contrast, the corneas of microMT mice failed to develop keratitis after intrastromal injection of parasite Ags unless Ags were injected with immune sera. Eosinophils were recruited from the limbal vessels to the peripheral cornea in microMT mice, but failed to migrate to the central cornea, whereas neutrophil recruitment was impaired at both stages. With the exception of IL-5, T cell responses and peripheral eosinophils were not significantly different between C57BL/6 and microMT mice. Taken together, these findings not only demonstrate that Ab is required for the development of keratitis, but also show that recruitment of neutrophils to the cornea is Ab dependent, whereas eosinophil migration is only partially dependent upon Ab interactions. PMID- 10528203 TI - Neutralization of endogenous granulocyte-macrophage colony-stimulating factor subverts the protective immune response to Histoplasma capsulatum. AB - We examined the influence of endogenous GM-CSF on the course of primary and secondary pulmonary histoplasmosis. A high proportion (>/=75%) of C57BL/6 mice given mAb to GM-CSF did not survive primary infection, whereas 88-94% of infected controls survived. Analysis of leukocytes revealed significantly fewer CD4+ and CD8+ cells in lungs, but not airways, of anti-GM-CSF-treated mice as compared with infected controls. However, the histopathology was similar between the two groups. Lungs of mice given mAb to GM-CSF manifested depressed levels of TNF alpha, IFN-gamma, and reactive nitrogen intermediates and elevated levels of IL-4 and IL-10. Administration of mAb to IL-4, to IL-10, or both restored protective immunity in GM-CSF-neutralized mice. In secondary infection, administration of mAb to GM-CSF exacerbated infection but did not alter survival over 30 days. The character of the inflammatory response was similar, and no differences were detected in Th1 or Th2 cytokine production between the two groups. Thus, endogenous GM-CSF is essential for survival in primary but not secondary infection, and blockade perturbs protective immunity. These findings reveal a new mechanism whereby GM-CSF contributes to host protection and demonstrate differences in control of primary and secondary histoplasmosis. PMID- 10528204 TI - A cyclophilin B gene encodes antigenic epitopes recognized by HLA-A24-restricted and tumor-specific CTLs. AB - We have studied Ags recognized by HLA class I-restricted CTLs established from tumor site to better understand the molecular basis of tumor immunology. HLA-A24 restricted and tumor-specific CTLs established from T cells infiltrating into lung adenocarcinoma recognized the two antigenic peptides encoded by a cyclophilin B gene, a family of genes for cyclophilins involved in T cell activation. These two cyclophilin B peptides at positions 84-92 and 91-99 induced HLA-A24-restricted CTL activity against tumor cells in PBMCs of leukemia patients, but not in epithelial cancer patients or in healthy donors. In contrast, the modified peptides at position 2 from phenylalanine to tyrosine, which had more than 10 times higher binding affinities to HLA-A24 molecules, could induce HLA-A24-restricted CTL activity against tumor cells in PBMCs from leukemia patients, epithelial cancer patients, or healthy donors. PHA-activated normal T cells were resistant to lysis by the CTL line or by these peptide induced CTLs. These results indicate that a cyclophilin B gene encodes antigenic epitopes recognized by CTLs at the tumor site, although T cells in peripheral blood (except for those from leukemia patients) are immunologically tolerant to the cyclophilin B. These peptides might be applicable for use in specific immunotherapy of leukemia patients or that of epithelial cancer patients. PMID- 10528202 TI - Nitric oxide and the Th2 response combine to prevent severe hepatic damage during Schistosoma mansoni infection. AB - During infection with Schistosoma mansoni, NO production increases following the deposition of parasite eggs in the liver. In wild-type C57BL/6 mice, NO levels peak during the sixth week of infection and are subsequently down-regulated. Inducible NO synthase (iNOS) mRNA was found in diseased liver tissue along with TNF-alpha and IFN-gamma, which are known promoters of iNOS expression. Mice treated with aminoguanidine, a selective inhibitor of iNOS, exhibited cachexia and exacerbated liver pathology, suggesting that NO limits hepatocyte damage when the liver is first exposed to eggs. Hepatic iNOS is up-regulated in SCID mice, indicating that NO production is part of an innate response. Studies with infected highly susceptible IL-4-/- mice revealed that prolonged NO production is in itself deleterious and that a major function of the Th2 response, which is severely compromised in the absence of IL-4, is to regulate NO production. In these animals, plasma NO levels are high compared with those in infected wild type mice and remain elevated until death. Nevertheless, the underlying importance of NO is illustrated by the finding that aminoguanidine treatment leads to more severe liver disease and reduced time to death in infected IL-4-/- mice. PMID- 10528205 TI - Nematode infection enhances survival of activated T cells by modulating accessory cell function. AB - The type of immune response generated following exposure to Ag depends on a variety of factors, including the nature of the Ag, the type of adjuvant used, the site of antigenic entry, and the immune status of the host. We have previously shown that infection of rodents with Nippostrongylus brasiliensis (Nb) shifts the development of type 1 allo-specific responses toward type 2 immunity, suggesting nematode modulation of T cell activation. In this report we explore the immunomodulatory effects of Nb on T cell activation. We found that spleen cells from Nb-infected mice exhibited dramatically increased proliferation in response to Con A and anti-CD3. This hyperproliferation could be transferred in vitro to naive splenocytes by coculture with mitomycin C-treated cells from Nb infected animals. The transfer was mediated by non-T accessory cells and supernatants derived from Con A-activated non-T cells, suggesting the involvement of a soluble factor secreted by accessory cells. The accessory cells secreted high levels of IL-6, and anti-IL-6 treatment abrogated the supernatant-induced hyperproliferation, thus confirming that IL-6 was mediating the effect. Further, spleen cells from Nb-infected mice were more resistant to activation-induced cell death (AICD) following mitogenic stimulation. Reduced AICD was also transferable and IL-6 dependent. Thus, the hyperproliferation was in part due to enhanced activated T cell survival. These phenomena mediated by accessory cells may contribute to the powerful polyclonal activation of type 2 immunity caused by nematode infection. PMID- 10528206 TI - Neutrophil dysfunctions, IL-8, and soluble L-selectin plasma levels in rapidly progressive versus adult and localized juvenile periodontitis: variations according to disease severity and microbial flora. AB - We used flow cytometry to analyze the expression of adhesion molecules and the oxidative burst of whole-blood polymorphonuclear neutrophils (PMN) from 26 patients with periodontitis. Three different clinical entities were studied: adult periodontitis (AP), localized juvenile periodontitis (LJP), and rapidly progressive periodontitis (RPP). Unstimulated PMN from the patients showed reduced Lewis x, sialyl-Lewis x, and L-selectin expression relative to those from healthy control subjects. These alterations were present whatever the severity of periodontal disease. However, PMN from RPP patients showed increased basal H2O2 production and decreased L-selectin shedding. These latter impairments, which correlated with increased IL-8 plasma levels, could contribute to initial vascular damage. In addition, decreased IL-8 priming of H2O2 production by PMN from RPP patients could account for a lower bactericidal capacity of PMN, leading to the large number of bacteria in the subgingival region of RPP patients. Soluble L-selectin plasma levels were also decreased in the RPP group, indicating more severe or diffuse endothelial damage. These abnormalities were not found in the patients with less destructive forms of periodontitis (AP and LJP). Porphyromonas gingivalis, a bacterial pathogen known to increase IL-8 production by PMN, was found in the periodontal pockets of RPP patients only. These results show links among PMN abnormalities, the clinical form of periodontitis, and the gingival bacterial flora. PMID- 10528207 TI - T cell-derived IL-10 promotes lung cancer growth by suppressing both T cell and APC function. AB - We have found previously that human lung cancers potently induce T lymphocyte IL 10 production in vitro. To assess the impact of enhanced T cell-derived IL-10 on antitumor immunity in vivo, we utilized transgenic mice expressing IL-10 under the control of the IL-2 promoter. We have shown previously that Lewis lung carcinoma cells (3LL) have more aggressive growth potential in IL-10 transgenic mice compared with control littermates. In this study, we show that transfer of T cells from IL-10 transgenic mice to control littermates transferred the IL-10 immunosuppressive effect and led to enhanced 3LL tumor growth. In addition to changes in T cell-mediated immunity, professional APC from IL-10 transgenic mice were found to have significantly suppressed capacity to induce MHC alloreactivity, CTL responses, and IL-12 production. Tumor Ag-pulsed dendritic cells from IL-10 transgenic mice also failed to generate antitumor reactivity. These results suggest that increased levels of T cell-derived IL-10 severely impair antitumor immunity in vivo, due to defects in both T cell and APC function. PMID- 10528208 TI - Relative contribution of LFA-1 and Mac-1 to neutrophil adhesion and migration. AB - To differentiate the unique and overlapping functions of LFA-1 and Mac-1, LFA-1 deficient mice were developed by targeted homologous recombination in embryonic stem cells, and neutrophil function was compared in vitro and in vivo with Mac-1 deficient, CD18-deficient, and wild-type mice. LFA-1-deficient mice exhibit leukocytosis but do not develop spontaneous infections, in contrast to CD18 deficient mice. After zymosan-activated serum stimulation, LFA-1-deficient neutrophils demonstrated activation, evidenced by up-regulation of surface Mac-1, but did not show increased adhesion to purified ICAM-1 or endothelial cells, similar to CD18-deficient neutrophils. Adhesion of Mac-1-deficient neutrophils significantly increased with stimulation, although adhesion was lower than for wild-type neutrophils. Evaluation of the strength of adhesion through LFA-1, Mac 1, and CD18 indicated a marked reduction in firm attachment, with increasing shear stress in LFA-1-deficient neutrophils, similar to CD18-deficient neutrophils, and only a modest reduction in Mac-1-deficient neutrophils. Leukocyte influx in a subcutaneous air pouch in response to TNF-alpha was reduced by 67% and 59% in LFA-1- and CD18-deficient mice but increased by 198% in Mac-1 deficient mice. Genetic deficiencies demonstrate that both LFA-1 and Mac-1 contribute to adhesion of neutrophils to endothelial cells and ICAM-1, but adhesion through LFA-1 overshadows the contribution from Mac-1. Neutrophil extravasation in response to TNF-alpha in LFA-1-deficient mice dramatically decreased, whereas neutrophil extravasation in Mac-1-deficient mice markedly increased. PMID- 10528209 TI - A novel LPS-inducible C-type lectin is a transcriptional target of NF-IL6 in macrophages. AB - C-type lectins serve multiple functions through recognizing carbohydrate chains. Here we report a novel C-type lectin, macrophage-inducible C-type lectin (Mincle), as a downstream target of NF-IL6 in macrophages. NF-IL6 belongs to the CCAAT/enhancer binding protein (C/EBP) of transcription factors and plays a crucial role in activated macrophages. However, what particular genes are regulated by NF-IL6 has been poorly defined in macrophages. Identification of downstream targets is required to elucidate the function of NF-IL6 in more detail. To identify downstream genes of NF-IL6, we screened a subtraction library constructed from wild-type and NF-IL6-deficient peritoneal macrophages and isolated Mincle that exhibits the highest homology to the members of group II C type lectins. Mincle mRNA expression was strongly induced in response to several inflammatory stimuli, such as LPS, TNF-alpha, IL-6, and IFN-gamma in wild-type macrophages. In contrast, NF-IL6-deficient macrophages displayed a much lower level of Mincle mRNA induction following treatment with these inflammatory reagents. The mouse Mincle proximal promoter region contains an indispensable NF IL6 binding element, demonstrating that Mincle is a direct target of NF-IL6. The Mincle gene locus was mapped at 0.6 centiMorgans proximal to CD4 on mouse chromosome 6. PMID- 10528210 TI - IL-1 alpha beta blockade prevents cartilage and bone destruction in murine type II collagen-induced arthritis, whereas TNF-alpha blockade only ameliorates joint inflammation. AB - Anti-TNF-alpha treatment of rheumatoid arthritis patients markedly suppresses inflammatory disease activity, but so far no tissue-protective effects have been reported. In contrast, blockade of IL-1 in rheumatoid arthritis patients, by an IL-1 receptor antagonist, was only moderately effective in suppressing inflammatory symptoms but appeared to reduce the rate of progression of joint destruction. We therefore used an established collagen II murine arthritis model (collagen-induced arthritis(CIA)) to study effects on joint structures of neutralization of either TNF-alpha or IL-1. Both soluble TNF binding protein and anti-IL-1 treatment ameliorated disease activity when applied shortly after onset of CIA. Serum analysis revealed that early anti-TNF-alpha treatment of CIA did not decrease the process in the cartilage, as indicated by the elevated COMP levels. In contrast, anti-IL-1 treatment of established CIA normalized COMP levels, apparently alleviating the process in the tissue. Histology of knee and ankle joints corroborated the finding and showed that cartilage and joint destruction was significantly decreased after anti-IL-1 treatment but was hardly affected by anti-TNF-alpha treatment. Radiographic analysis of knee and ankle joints revealed that bone erosions were prevented by anti-IL-1 treatment, whereas the anti-TNF-alpha-treated animals exhibited changes comparable to the controls. In line with these findings, metalloproteinase activity, visualized by VDIPEN production, was almost absent throughout the cartilage layers in anti-IL-1 treated animals, whereas massive VDIPEN appearance was found in control and sTNFbp-treated mice. These results indicate that blocking of IL-1 is a cartilage- and bone-protective therapy in destructive arthritis, whereas the TNF-alpha antagonist has little effect on tissue destruction. PMID- 10528211 TI - C1q and C4b bind simultaneously to CR1 and additively support erythrocyte adhesion. AB - Previously, we showed that soluble C1q bound specifically to CR1 on transfected cells. If the CR1-C1q interaction were to participate in immune complex clearance, then this interaction should support E adhesion. Using a tip plate adhesion assay, we found that immobilized C1q mediated adhesion of human E. E binding to C1q was specifically inhibited by polyclonal anti-CR1 Fab fragments. Intact C1 was not efficient as an adherence ligand until it was treated with EDTA or the C1 inhibitor to remove the C1r2C1s2 complex from C1, leaving C1q. Titration of C1q alone, C4b alone, and C1q + C4b indicated that the two complement ligands were additive in their ability to support CR1-mediated adhesion of E. Analysis of binding to immobilized CR1 using a BIAcore instrument documented that C1q, C4b, and C3b binding were independent events. Additionally, C1q-dependent binding of immune complexes and heat-aggregated IgG to E was documented. These experiments confirm that the immune adherence receptor in humans, CR1, is the single receptor for all of the opsonic ligands of complement, provide evidence for a single C1q binding site on LHR-D of CR1, and suggest that C1q may participate in immune clearance. PMID- 10528212 TI - Proteinase-activated receptor-2-activating peptides induce leukocyte rolling, adhesion, and extravasation in vivo. AB - Proteinase-activated receptor 2 (PAR2) has been suggested to play a role in inflammatory reactions. Because leukocyte-endothelial cell interactions are critical events during inflammatory reactions, and because PAR2 is expressed both on endothelium and leukocytes, we have examined the effects of PAR2-activating peptides (PAR2-APs) on leukocyte rolling and adhesion in mesenteric venules and on leukocyte recruitment into the peritoneal cavity. Using intravital microscopy, leukocyte rolling, flux, and adhesion in rat mesenteric postcapillary venules were quantified. Topical addition of PAR2-APs (10 microM) for 1 min to the superfused venule induced a significant increase in leukocyte rolling and adherence. The increase in leukocyte adherence was not affected by pretreatment with a mast cell stabilizer (sodium cromoglycate) nor by prior degranulation of mast cells with compound 48/80. Nonetheless, both leukocyte rolling and adhesion were completely inhibited by pretreatment with a platelet-activating factor receptor antagonist (WEB 2086). Intraperitoneal injections of a selective PAR2-AP (SLIGRL-NH2) caused a significant increase in leukocyte migration into the peritoneal cavity. The effect of SLIGRL-NH2 on peritoneal leukocyte infiltration was completely inhibited by WEB 2086. These data suggest that PAR2 activation could contribute to several early events in the inflammatory reaction, including leukocyte rolling, adherence, and recruitment, by a mechanism dependent on platelet-activating factor release. PMID- 10528213 TI - L-selectin ligands expressed by human leukocytes are HECA-452 antibody-defined carbohydrate epitopes preferentially displayed by P-selectin glycoprotein ligand 1. AB - Leukocytes express L-selectin ligands critical for leukocyte-leukocyte interactions at sites of inflammation. The predominant leukocyte L-selectin ligand is P-selectin glycoprotein ligand-1 (PSGL-1), which displays appropriate sialyl Lewis x (sLex)-like carbohydrate determinants for L-selectin recognition. Among the sLex-like determinants expressed by human leukocytes is a unique carbohydrate epitope defined by the HECA-452 mAb. The HECA-452 Ag is a critical component of L-selectin ligands expressed by vascular endothelial cells. However, HECA-452 Ag expression on human leukocyte L-selectin ligands has not been assessed. In this study, the HECA-452 mAb blocked 88-99% of neutrophil rolling on, or attachment to, adherent cells expressing L-selectin in multiple experimental systems. A function-blocking anti-PSGL-1 mAb also inhibited L selectin binding to neutrophils by 89-98%. In addition, the HECA-452 and anti PSGL-1 mAbs blocked the majority of P-selectin binding to neutrophils. Western blot analysis revealed that PSGL-1 immunoprecipitated from neutrophils displayed HECA-452 mAb-reactive determinants and that PSGL-1 was the predominant scaffold for HECA-452 Ag display. Leukocyte L-selectin ligands also contained sulfated determinants since culturing ligand-bearing cells with NaClO3 abrogated L selectin binding. Consistent with this, human neutrophils expressed mRNA encoding five different sulfotransferases associated with the generation of selectin ligands: CHST1, CHST2, CHST3, TPST1, and HEC-GlcNAc6ST. Therefore, the HECA-452 defined carbohydrate determinant displayed on PSGL-1 represented the predominant L-selectin and P-selectin ligand expressed by neutrophils. PMID- 10528214 TI - Src-regulated extracellular signal-related kinase and Syk-regulated c-Jun N terminal kinase pathways act in conjunction to induce IL-1 synthesis in response to microtubule disruption in HL60 cells. AB - A microtubule reorganization is often observed during cellular contacts that are associated to IL-1 production. Here, we show that in HL60 cells, vincristine, a microtubule-disrupting agent that induces a strong production of IL-1, triggers the activation of both extracellular signal-related kinase (ERK) and c-Jun N terminal kinase (JNK-1). While ERK activation is rapid and transient, peaking at 10 min, the JNK1 activation is delayed and more sustained reaching a maximum at 2 h. ERK activation was blocked by CP 118556, indicating it is regulated by a Src like kinase, while JNK1 was inhibited by piceatannol, revealing an upstream regulation by Syk. Each kind of the nonreceptor tyrosine kinase blockers efficiently inhibits the vincristine-induced IL-1 production and diminishes the level of IL-1 transcripts, indicating that the ERK and JNK pathways act coordinately to elicit the transcription of the IL-1 gene. Furthermore, we found that pertussis toxin, a blocker of Go/Gi proteins, abrogated the vincristine induced activation of both Src and Syk. Our data support a model where the status of microtubule polymerization influences the activity of Go or Gi proteins that control, in turn, two independent Src/ERK and Syk/JNK1 cascades that are both necessary to sustain IL-1 synthesis. PMID- 10528215 TI - Tissue specificity of E- and P-selectin ligands in Th1-mediated chronic inflammation. AB - The demonstrated role of E- and P-selectin ligands in the recruitment of Th1 cells raises the question of tissue specificity determination by pathogenic T cells. We took advantage of the fact that chronic Th1-mediated inflammation in the scid/scid CD4+CD45RBhigh T cell transfer model can occur at multiple tissue sites, resembling inflammatory bowel disease in the colon and psoriasis in the skin. We show that the majority of infiltrating effector T cells from psoriatic skin expresses high levels of functional P-selectin ligand (87 +/- 3%), detected by P-selectin-Ig (PIg), while a significantly smaller subset of T cells from colitic lesions expresses this ligand (24 +/- 2%). Similarly, E-selectin ligand is preferentially expressed on CD4+ T cells infiltrating the skin (24 +/- 2%), but only on very few CD4+ T cells infiltrating the colon (CIT; 1.3 +/- 0.8%). In contrast, CD4+ T cells infiltrating the skin express alpha4beta7 at a significantly lower level than CIT (mean fluorescence intensity, 28 vs 61, respectively), although, interestingly, alphaEbeta7 was expressed at high levels on both populations. Analysis of the disease-inducing potential of PIg+ and PIg- CD4+ CIT cells revealed that both populations not only express similar levels of the gut-homing molecule alpha4beta7 (mean fluorescence intensity, 50 vs 56, respectively), but do not differ in their capacity to express IFN-gamma. Furthermore, CIT depleted of cells expressing functional P-selectin ligand were able to induce colitis upon transfer, suggesting that induction of colitis in this model may be independent of E- and P-selectin. These results indicate that adhesion molecule expression and the homing pattern of inflammatory T cells are regulated by the local environment independently of their inflammatory capacity. PMID- 10528216 TI - Role of IL-6 in the pleurisy and lung injury caused by carrageenan. AB - In the present study we used IL-6 knockout mice (IL-6KO) to evaluate the role of IL-6 in the inflammatory response caused by injection of carrageenan into the pleural space. Compared with carrageenan-treated IL-6 wild-type (IL-6WT) mice, carrageenan-treated IL-6KO mice exhibited a reduced degree of pleural exudation and polymorphonuclear cell migration. Lung myeloperoxidase activity and lipid peroxidation were significantly reduced in IL-6KO mice compared with those in IL 6WT mice treated with carrageenan. Immunohistochemical analysis for nitrotyrosine and poly(A)DP-ribose polymerase revealed a positive staining in lungs from carrageenan-treated IL-6WT mice. No positive staining for nitrotyrosine or PARS was found in the lungs of the carrageenan-treated IL-6KO mice. Staining of lung tissue sections obtained from carrageenan-treated IL-6WT mice with an anti-cyclo oxygenase-2 Ab showed a diffuse staining of the inflamed tissue. Furthermore, expression of inducible nitric oxide synthase was found mainly in the macrophages of the inflamed lungs from carrageenan-treated IL-6WT mice. The intensity and degree of the staining for cyclo-oxygenase-2 and inducible nitric oxide synthase were markedly reduced in tissue sections obtained from carrageenan-treated IL-6KO mice. Most notably, the degree of lung injury caused by carrageenan was also reduced in IL-6KO mice. Treatment of IL-6WT mice with anti-IL-6 (5 microg/day/mouse at 24 and 1 h before carrageenan treatment) also significantly attenuated all the above indicators of lung inflammation. Taken together, our results clearly demonstrate that IL-6KO mice are more resistant to the acute inflammation of the lung caused by carrageenan injection into the pleural space than the corresponding WT mice. PMID- 10528217 TI - Recombinant human (rh)IL-4-mediated apoptosis and recombinant human IL-6-mediated protection of recombinant human stem cell factor-dependent human mast cells derived from cord blood mononuclear cell progenitors. AB - Although stem cell factor (SCF) appears to be the major growth factor for human mast cells, other factors undoubtedly play important roles in the development, survival, and function of these cells. The current study examined the effects of recombinant human (rh) IL-4 and rhIL-6 on rhSCF-dependent development and survival of human mast cells derived in vitro from cord blood progenitor cells. After 4-8 wk of culture with rhSCF and various amounts of rhIL-4, a dramatic decline in mast cell numbers was observed with rhIL-4, the EC50 being about 0.1 ng/ml. Numbers of other cell types remained high. Mast cells derived from cord blood progenitors after 7 wk of culture with rhSCF alone displayed an MCT phenotype and expressed Kit, FcepsilonRI, and IL-4R on their surface. Mast cells examined after purification by immunomagnetic sorting became apoptotic within hours after exposure to rhIL-4, a phenomenon blocked by anti-IL-4 Ab. Because rhIL-4-dependent apoptosis but not the loss of mitochondrial membrane potential was prevented by the pan-caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-(Z-VAD) fluoromethylketone, mitochondrial perturbation most likely preceded caspase activation. Consistent with this conclusion was the observation that both apoptosis and loss of mitochondrial membrane potential (Deltapsim) were inhibited by cyclosporin A in combination with aristolochic acid. rhIL-6 protected cord blood mast cells from rhIL-4-induced apoptosis. Thus, IL-4 can cause both maturation and apoptosis of human mast cells, the latter effect being abrogated by IL-6. PMID- 10528218 TI - Preferential role for NF-kappa B/Rel signaling in the type 1 but not type 2 T cell-dependent immune response in vivo. AB - T cell function is a critical determinant of immune responses as well as susceptibility to allergic diseases. Activated T cells can differentiate into effectors whose cytokine profile is limited to type 1 (IFN-gamma-dominant) or type 2 (IL-4-, IL-5-dominant) patterns. To investigate mechanisms that connect extracellular stimuli with the regulation of effector T cell function, we have measured immune responses of transgenic mice whose NF-kappa B/Rel signaling pathway is inhibited in T cells. Surprisingly, these mice developed type 2 T cell dependent responses (IgE and eosinophil recruitment) in a model of allergic pulmonary inflammation. In contrast, type 1 T cell responses were severely impaired, as evidenced by markedly diminished delayed-type hypersensitivity responses, IFN-gamma production, and Ag-specific IgG2a levels. Taken together, these data indicate that inhibition of NF-kappa B can lead to preferential impairment of type 1 as compared with type 2 T cell-dependent responses. PMID- 10528219 TI - Granulocyte-colony stimulating factor treatment of lupus autoimmune disease in MRL-lpr/lpr mice. AB - G-CSF not only functions as an endogenous hemopoietic growth factor for neutrophils, but also displays pro-Th2 and antiinflammatory properties that could be of therapeutic benefit in autoimmune settings. We evaluated the effect of treatment with G-CSF in a murine model of spontaneous systemic lupus erythematosus, a disease in which G-CSF is already administered to patients to alleviate neutropenia, a common complication. Chronic treatment of lupus-prone MRL-lpr/lpr mice with low doses (10 microg/kg) of recombinant human G-CSF, despite the induction of a shift toward the Th2 phenotype of the autoimmune response, increased glomerular deposition of Igs and accelerated lupus disease. Conversely, high-dose (200 microg/kg) treatment with G-CSF induced substantial protection, prolonging survival by >2 mo. In the animals treated with these high doses of G-CSF, neither the Th1/Th2 profile nor the serum levels of TNF-alpha and IL-10 were modified. Despite the presence of immune complexes in their kidney glomeruli, no inflammation ensued, and serum IL-12 and soluble TNF receptors remained at pre-disease levels. This uncoupling of immune complex deposition and kidney damage resulted from a local down-modulation of FcgammaRIII (CD16) expression within the glomeruli by G-CSF. Our results demonstrate a beneficial effect of high doses of G-CSF in the prevention of lupus nephritis that may hold promise for future clinical applications, provided caution is taken in dose adjustment. PMID- 10528220 TI - B cell repertoire diversity and clonal expansion in multiple sclerosis brain lesions. AB - Multiple sclerosis (MS) lesions in the CNS are characterized by disseminated demyelination with perivascular infiltrates of macrophages, T cells, and B cells. To investigate the origin and characteristics of the B cell population found in MS plaque tissue, we performed molecular studies in 10 MS patients and 4 non-MS control samples. Ig transcripts from the perivascular infiltrated brain lesions were analyzed by complementary-determining region 3 spectratyping to ascertain the B cell heavy chain gene rearrangement repertoire expressed in MS brains. Significant rearrangement diversity and deviation from the normal Ig heavy (H) chain repertoire was observed. The cloning and sequencing of RT-PCR products from families VH1 and VH4 showed a correlation with the profiles obtained by spectratyping. Generally, restricted spectratyping patterns concurred with repetition of in-frame complementary-determining region 3 identical sequences. The analysis of heavy chain variable (VH), diversity (D), and joining (JH) gene segments revealed the increased usage of VH1-69, VH4-34, and VH4-39. Similarly, gene segments from families D2, D3, and JH4 were over-represented. The presence of restricted patterns of rearranged Ig mRNA within the plaque lesion suggests that Ab production in the demyelinating plaque is a local phenomenon and supports the idea that in MS an Ag-driven immune response might be responsible for demyelination. PMID- 10528221 TI - Photochemical treatment with S-59 psoralen and ultraviolet A light to control the fate of naive or primed T lymphocytes in vivo after allogeneic bone marrow transplantation. AB - Donor leukocyte infusions after allogeneic bone marrow transplantation can provide a curative graft-vs-leukemia (GVL) effect, but there is a significant risk of graft-vs-host (GVH) disease. A simple and effective method for controlling the fate of naive or primed T-lymphocytes in vivo without eliminating their beneficial properties is needed. In this report, photochemical treatment (PCT) ex vivo with a synthetic psoralen (S-59) and UVA light was evaluated as a pharmacological approach to limiting the proliferation and GVH potential of naive and primed donor T cells in vivo. S-59 rapidly intercalates into and cross-links DNA on UVA illumination. The effects of PCT on T cells were found to be both S-59 and UVA dose dependent. With selected PCT regimens, treated T cells still expressed activation markers (CD25 and CD69) and secreted IL-2 on activation, but they showed limited proliferative capacity in vitro and in vivo. Clonal expansion of CTL in MLR was reduced after PCT, but short term lytic activity of primed CTL was not affected. In a murine model of MHC-mismatched bone marrow transplantation, the addition of PCT-treated T cells to T-depleted bone marrow facilitated donor engraftment and complete chimerism without causing acute or chronic graft-vs-host disease. Allospecific GVL reactivity was reduced but not eliminated after PCT treatment. In an MHC-matched model using host-presensitized donor T cells, PCT significantly reduced GVH-associated mortality without eliminating GVL reactivity. Thus, PCT ex vivo offers a simple, rapid, and inexpensive method by which to control the fate of naive and primed T cells in vivo. PMID- 10528222 TI - Induction of experimental autoimmune Graves' disease in BALB/c mice. AB - We immunized BALB/c mice with M12 cells (H-2d) expressing either mouse (mM12 cells) or human thyrotropin receptor (TSHR) (hM12 cells). Immunized mice developed autoantibodies to native TSHR by day 90 and, by day 180, showed considerable stimulatory Ab activity as measured by their ability to enhance cAMP production (ranging from 6. 52 to 20.83 pmol/ml in different treatment groups relative to 1.83 pmol/ml for controls) by TSHR-expressing Chinese hamster ovary cells. These mice developed severe hyperthyroidism with significant elevations in both tetraiodothyronine and triiodothyronine hormones. Tetraiodothyronine levels in different experimental groups ranged from a mean of 8.66-12.4 microg/dl, relative to 4.8 microg/dl in controls. Similarly, mean triiodothyronine values ranged from 156.18 to 195.13 ng/dl, relative to 34.99 ng/dl for controls. Next, we immunized BALB/c mice with a soluble extracellular domain of human TSHR (TBP), or TBP expressed on human embryonic kidney cells (293 cells) (293-TBP cells). These mice showed severe hyperthyroidism in a manner very similar to that described above for mice immunized with the mouse TSHR or human TSHR, and exhibited significant weight loss, with average weight for treatment groups ranging from 20.6 to 21.67 g, while controls weighed 24.2 g. Early after onset of the disease, histopathological examination of thyroids showed enlargement of colloids and thinning of epithelial cells without inflammation. However, later during disease, focal necrosis and lymphocytic infiltration were apparent. Our results showed that conformationally intact ectodomain of TSHR is sufficient for disease induction. Availability of a reproducible model in which 100% of the animals develop disease should facilitate studies aimed at understanding the molecular pathogenesis of Graves' disease. PMID- 10528223 TI - Gene therapy using SV40-derived vectors: what does the future hold? AB - Effective genetic therapy requires both a fragment of genetic material to be used therapeutically and a means to deliver it. We began to study simian virus-40 (SV40) as a vector for gene transfer because available gene delivery vehicles did not provide for the full range of therapeutic uses. Other vectors are variably limited by immunogenicity, difficulties in production, restricted specificity, low titers, poor transduction efficiency, etc. In theory recombinant viral vectors based on SV40 (rSV40) should not, on the other hand, be similarly constrained. rSV40 vectors are easily manipulated and produced at very high titer, stable, lacking in immunogenicity, and capable of providing sustained high levels of transgene expression in both resting and dividing cells. The principle limitation of SV40-derived vectors is the size of the packageable insert (95%) and the increased release of 3-MH into the medium in response to Dex (>95%). Northern hybridization studies demonstrated that Dex also elicited similar time- and concentration-dependent increases in the expression of mRNA encoding two components (14 kDa E(2) Ub-conjugating enzyme and Ub) of the ATP-Ub-dependent pathway. The data demonstrate that Dex stimulates preferential hydrolysis of myofibrillar proteins in C(2)C(12) myotubes and suggests that the ATP-Ub dependent pathway is involved in this response. PMID- 10528232 TI - Possible involvement of heat shock protein 25 in the angiotensin II-induced glomerular mesangial cell contraction via p38 MAP kinase. AB - In the rat kidney, mesangial cells (MCs), especially those in the extraglomerular mesangium (EGM) region of the juxtagomerular apparatus, express high amounts of heat shock protein 25 (HSP25). Because MCs are contractile in vivo and HSP25 is known to modulate polymerization/depolymerization of F-actin and to be involved in smooth muscle contraction, it is possible that HSP25 participates in the contraction process of MCs. We analyzed a permanent mouse MC line using Northern and Western blot analyses, and observed that similar to the MCs in the glomerulus, these cells also express high amounts of HSP25 constitutively. Exposure of these cells to angiotensin II (ANG II: 2 x 10(-7) M) evoked contraction and a concomitant increase in HSP25 phosphorylation, while the cytoplasmic fraction of HSP25 was transiently reduced. Because phosphorylation of HSP25 is essential for its actin-modulating function, we suppressed the activity of p38 MAP kinase, the major upstream activator of HSP25 phosphorylation, with the specific inhibitor SB 203580. This maneuver reduced HSP25 phosphorylation dramatically, abolished cell contraction, and prevented the decrease of the cytoplasmic HSP25 content. This suggests that HSP25 might be a component of the contraction machinery in MCs and that this process depends on p38 MAP kinase mediated HSP25 phosphorylation. The decrease of cytoplasmic HSP25 content observed after ANG II exposure is probably the result of a transient redistribution of HSP25 into a buffer-insoluble fraction, because the whole cell content of HSP25 did not change, a phenomenon known to be related to the actin modulating activity of HSP25. The fact that this function requires phosphorylation of HSP25 would explain the observation that HSP25 does not redistribute in SB 203580-pretreated cells. PMID- 10528233 TI - Glucagon-like peptide-1 does not mediate amylase release from AR42J cells. AB - In this study, AR42J pancreatic acinar cells were used to investigate if glucagon like peptide-1 (GLP-1) or glucagon might influence amylase release and acinar cell function. We first confirmed the presence of GLP-1 receptors on AR42J cells by reverse trasncriptase-polymerase chain reaction (RT-PCR), Western blotting, and partial sequencing analysis. While cholecystokinin (CCK) increased amylase release from AR42J cells, GLP-1, alone or in the presence of CCK, had no effect on amylase release but both CCK and GLP-1 increased intracellular calcium. Similar to GLP-1, glucagon increased both cyclic adenosine monophosphate (cAMP) and intracellular calcium in AR42J cells but it actually decreased CCK-mediated amylase release (n = 20, P < 0.01). CCK stimulation resulted in an increase in tyrosine phosphorylation of several cellular proteins, unlike GLP-1 treatment, where no such increased phosphorylation was seen. Instead, GLP-1 decreased such protein phosphorylations. Genestein blocked CCK-induced phosphorylation events and amylase secretion while vanadate increased amylase secretion. These results provide evidence that tyrosine phosphorylation is necessary for amylase release and that signaling through GLP-1 receptors does not mediate amylase release in AR42J cells. J. Cell. Physiol. 181:470-478, 1999. Published 1999 Wiley-Liss, Inc. PMID- 10528234 TI - Regulation of expression of collagenase-3 in normal, differentiating rat osteoblasts. AB - We investigated the regulation of collagenase-3 expression in normal, differentiating rat osteoblasts. Fetal rat calvarial cell cultures showed an increase in alkaline phosphatase activity reaching maximal levels between 7-14 days post-confluence, then declining with the onset of mineralization. Collagenase-3 mRNA was just detectable after proliferation ceased at day 7, increased up to day 21, and declined at later ages. Postconfluent cells maintained in non-mineralizing medium expressed collagenase-3 but did not show the developmental increase exhibited by cells switched to mineralization medium. Cells maintained in non-mineralizing medium continued to proliferate; cells in mineralization medium ceased proliferation. In addition, collagenase-3 mRNA was not detected in subcultured cells allowed to remineralize. These results suggest that enhanced accumulation of collagenase-3 mRNA is triggered by cessation of proliferation or acquisition of a mineralized extracellular matrix and that other factors may also be required. After initiation of basal expression, parathyroid hormone (PTH) caused a dose-dependent increase in collagenase-3 mRNA. Both the cyclic adenosine monophosphate (cAMP) analogue, 8-bromo-cAMP (8-Br-cAMP), and the protein kinase C (PKC) activator, phorbol myristate acetate, increased collagenase-3 expression, while the calcium ionophore, ionomycin, did not, suggesting that PTH was acting through the protein kinase A (PKA) and PKC pathways. Inhibition of protein synthesis with cycloheximide caused an increase in basal collagenase-3 expression but blocked the effect of PTH, suggesting that an inhibitory factor prevents basal expression while an inductive factor is involved with PTH action. In summary, collagenase-3 is expressed in mineralized osteoblasts and cessation of proliferation and initiation of mineralization are triggers for collagenase-3 expression. PTH also stimulates expression of the enzyme through both PKA and PKC pathways in the mineralizing osteoblast. PMID- 10528235 TI - Protein kinase C activation downregulates the expression and function of the basolateral Na+/K+/2Cl(-) cotransporter. AB - The basolateral Na+/K+/2Cl(-) cotransporter (NKCC1) has been shown to be an independent regulatory site for electrogenic Cl(-) secretion. The proinflammatory phorbol ester, phorbol 12-myristate 13-acetate (PMA), which activates protein kinase C (PKC), inhibits basal and cyclic adenosine monophosphate (cAMP) stimulated NKCC1 activity in T84 intestinal epithelial cells and decreases the steady state levels of NKCC1 mRNA in a time- and dose-dependent manner. The levels of NKCC1 protein also fall in accordance with the NKCC1 mRNA transcript and these levels are unaffected by 4alpha-phorbol, which does not activate PKC. Inhibition of maximal (cAMP-stimulated) NKCC1 functional activity by PMA was first detected by 1 h, whereas decreases in the steady state levels of NKCC1 mRNA were not detectable until 4 h. NKCC1 mRNA expression recovers toward control levels with extended treatment of cells with PMA suggesting that the PMA effects on NKCC1 expression are mediated through activation of PKC. Although NKCC1 mRNA and protein levels return to control values after extended PMA exposure, NKCC1 functional activity does not recover. Immunofluorescence imaging suggest that the absence of functional recovery is due to failure of newly synthesized NKKC1 protein to reach the cell surface. We conclude that NKCC1 has the capacity to be regulated at the level of de novo expression by PKC, although decreased NKCC1 expression alone cannot account for either early or late loss of NKCC1 function. PMID- 10528236 TI - Skeletal muscle satellite cell proliferation in response to members of the fibroblast growth factor family and hepatocyte growth factor. AB - Fibroblast growth factors (FGF) have the ability to regulate satellite cell proliferation in culture and in muscle tissue, but the specific FGF receptors (FGFR) expressed by adult rat muscle satellite cells and the action of members of the FGF family have not been assessed. Therefore, the expression of FGF receptors 1-4 was examined in proliferating satellite cells in culture, and the effects of eight members of the fibroblast growth factor family (FGFs1, 2, 4, 5, 6, 7, 8, and 9) on adult rat muscle satellite cells were evaluated. In addition, the interactions of FGFs with hepatocyte growth factor (HGF) were described. Of the eight FGFs evaluated, 1, 2, 4, 6, and 9 significantly (P < 0.05) stimulated proliferation above control. FGFs5, 7, and 8 displayed no mitogenic activity. Furthermore, combinations of HGF with FGFs2, 4, 6, or 9 stimulated satellite cell proliferation above that of optimal concentrations of HGF alone. Expression of four FGFR genes was detected in satellite cell cultures by reverse-transcription polymerase chain reaction (RT-PCR). FGFR1 and FGFR4 were the most prominent forms expressed, and FGFR2 was only expressed at low levels. FGFR3 was difficult to detect. FGFR1 and FGFR2 were also expressed in muscle-derived fibroblasts, but FGFR4 and FGFR3 were not. In proliferating cultures of satellite cells, HGF, insulin-like growth factor I (IGF-I) and FGF1 stimulated significantly (P < 0.05) higher levels of FGFR1 message content, relative to control conditions, and platelet-derived growth factor-BB (PDGF-BB) and insulin-like growth factor (IGF II) significantly (P < 0.05) depressed FGFR1 expression. During the activation period of satellite cell growth in culture (0-48 h), FGFR1 message content significantly (P < 0.05) increased from less than 1,000 copies per cell to approximately 5,000 copies per cell between 18 and 48 h, and HGF treatment significantly (P < 0.05) accelerated the accumulation of FGFR1 message during this period. PMID- 10528238 TI - Penile neurofibromas. AB - Unless omitted and underreported, penile neurofibromas are rare. Between January 2, 1982 and December 31, 1997 through the USF Regional Genetics Program we evaluated 566 propositi with suspected or clinically diagnosed neurofibromatosis (NF1, NF2, segmental NF=NF5, NF/Noonan syndrome, familial cafe-au-lait macules, and solitary neurofibroma, NF). These index cases were part of 32,715 families evaluated during the period. NF1 was the diagnosis in 361; 2 of them had penile NFs. A toddler presented with congenital plexiform NF of the penile shaft and another propositus developed two small subcutaneous NFs, on the penile shaft and on the left scrotal wall, respectively. A review documented 26 additional patients with penile NF. As to the pathogenesis of the NF1 lesions, a paracrine growth model including the multiple levels of regulation of expression of the NF1 gene appeared more plausible than the loss of heterozygosity (LOH) model, which ignores the complexity of the paracrine growth mechanism. PMID- 10528237 TI - MET(PRC) mutations in the Ron receptor result in upregulation of tyrosine kinase activity and acquisition of oncogenic potential. AB - Ron and Met are structurally related receptor tyrosine kinases that elicit a complex biological response leading to invasive growth. Naturally occurring point mutations activate the Met kinase in papillary renal carcinomas (MET(PRC) mutations). By site-directed mutagenesis, we generated homologous amino acid substitutions in the Ron kinase domain and analyzed the biochemical and biological properties of the mutant receptors. Among the mutations studied, D(1232)H and M(1254)T displayed transforming activity in NIH3T3 cells, inducing focus formation and anchorage-independent growth. The D(1232)H and M(1254)T substitutions resulted in increased Ron autophosphorylation both in vivo and in vitro and constitutive binding to intracellular signal transducers. Both mutations yielded a dramatic increase in catalytic efficiency, indicating a direct correlation between kinase activity and oncogenic potential. Molecular modeling of the Ron D(1232)H mutation suggests that this single amino acid substitution favors the transition of the kinase from the inactive to the active state. These data demonstrate that point mutations can confer transforming activity to the Ron receptor and show that RON is a potential oncogene. PMID- 10528239 TI - Proximal 5p trisomy resulting from a marker chromosome implicates band 5p13 in 5p trisomy syndrome. AB - We describe an infant with trisomy of (5)(p10p13.1) resulting from a de novo marker chromosome. The marker's origin was identified by chromosome microdissection and reverse in situ hybridization. The clinical findings are compared to those of other partial and complete 5p duplications. This case further defines the critical region of 5p trisomy syndrome to proximal 5p. PMID- 10528240 TI - Somatic mosaicism of a greater than 1.7-Mb deletion of genomic DNA involving the entire NF1 gene as verified by FISH: further evidence for a contiguous gene syndrome in 17q11.2. AB - We report on a third case with neurofibromatosis type 1 (NF1) due to mosaicism for a gross deletion in 17q11.2 covering the entire NF1 gene. The deletion was suspected in Giemsa banded chromosomes and was confirmed by fluorescence in situ hybridization using the cosmids CO919 from the 5' region, GO2121 from the central, H10410 from the 3' region of the NF1 gene, and the 1.7-Mb YAC 947G11 spanning the entire 350-kb genomic DNA of the NF1 gene. The deletion was present in 33% of peripheral blood lymphocytes and 58% of fibroblasts. The clinical manifestations in this 6-year-old male patient were especially severe and extended beyond the typical features of NF1. The patient also displayed facial anomalies, severe and early-onset psychomotor retardation, seizures, spasticity, and microcephaly. These features differ from other large-deletion NF1 patients, even nonmosaic cases. The complex phenotype could be explained by the involvement of coding sequences flanking the NF1 gene, thus supporting the existence of a contiguous gene syndrome in 17q11.2. PMID- 10528241 TI - Subtle overlapping deletions in the terminal region of chromosome 6q24.2-q26: three cases studied using FISH. AB - Interstitial deletions in the terminal region of chromosome 6 are rare. We describe three new cases with subtle interstitial deletions in the q24-q26 region of the long arm of chromosome 6. The karyotypes were analyzed at a 550 band level. Patient1 is a 9-month-old boy with an interstitial deletion, del(6)(q24.2q25.1), developmental delay, low birth weight, hypotonia, heart murmur, respiratory distress, craniofacial and genital anomalies. This is the first report of a case with deletion del(6)(q24.2q25.1). Patient 2 is a 17-year old young man with an interstitial deletion del(6)(q25.1q25.3), developmental delay, short stature, mental retardation, autism, head, face, chest, hand and feet anomalies and a history of seizures. For the first time autism was described as a manifestation in 6q deletions. Patient 3 is baby boy with a de novo interstitial deletion, del(6)(q25.1q26), anomalies of the brain, genital organs, limbs and feet. This is the first report of a case with deletion, del(6)(q25.1q26). In all three patients, fluorescence in situ hybridization (FISH) using chromosome 6 painting probe ruled out an insertion. The ESR (6q25.1) and TBP (6q27) probes were used to confirm the breakpoints. Since TBP signal is present in all cases, it confirmed an interstitial deletion proximal to this probe. Patient 1 has a deletion of the ESR locus; Patient 2 and 3 have signals for the ESR locus on both chromosomes 6. Therefore the deletion in Patients 2 and 3 are between ESR and TBP loci distal to that of Patient 1. FISH validated the deletion breakpoints assessed by conventional cytogenetics. PMID- 10528242 TI - Linkage disequilibrium of MTHFR genotypes 677C/T-1298A/C in the German population and association studies in probands with neural tube defects(NTD). AB - A number of studies have demonstrated that the common polymorphism 677C-->T in the gene encoding 5, 10-methylenetetrahydrofolate reductase (MTHFR) leads to a thermolabile variant with decreased enzyme activity and to mildly elevated plasma homocysteine. 677TT homozygosity was shown to be more frequent in NTD probands compared with controls in some studies. Recently, another polymorphism, 1298A- >C, in the MTHFR gene was described and combined heterozygosity 677CT/1298AC was suggested to be an additional risk factor for NTD. The present study examines the genotype and haplotype distribution of the two polymorphisms in the German population and evaluates the impact on NTD individuals and their relatives. To determine the haplotype of all individuals tested, we developed an easy-to perform ARMS-RFLP test. Our data show that the two polymorphisms are in linkage disequilibrium in the general population and in NTD individuals. There was no statistically significant difference in allele and genotype frequency between probands (patients, fetuses) and controls (P > 0.10) and between observed and expected values for mother-child pairs (P > 0.80). Taking into account gender, an increased rate of 677CT heterozygotes was found in affected and unaffected males compared to affected and unaffected females. A family-based association study using a multiallelic transmission disequilibrium test (TDT) also shows that transmission rates do not deviate significantly from equilibrium (P > 0.50). Thus, our data provide no evidence for an association between NTD phenotype and MTHFR 677C/T-1298A/C genotypes and haplotypes. PMID- 10528243 TI - Late-onset familial Mediterranean fever (FMF): a subset with distinct clinical, demographic, and molecular genetic characteristics. AB - To determine the prevalence and characterize demographic, clinical, and genetic features of familial Mediterranean fever (FMF) of late onset, all patients experiencing their first FMF attack at age 40 years or more were identified using the computerized registry of our FMF clinic, and then thoroughly interviewed and examined. The control group consisted of 40 consecutive FMF patients, who arrived at the FMF clinic for their regular follow-up visit and were 40 years of age or older at the time of the examination. The severity of the disease in patients and controls was determined using a modified score, developed previously. Mutational analysis in the FMF gene was performed using a commercial kit. Only 20 of 4000 (0.5%) patients had late-onset FMF. These patients were mostly men, of non-North African origin, P < 0.05 compared to controls. All had abdominal attacks and in most these were the only manifestation of their disease, P < 0.001. None had chronic or prolonged manifestations of FMF, for example, amyloidosis, chronic arthritis, or protracted myalgia, P < 0.001. The response to treatment was good despite using low colchicine dose, P < 0.05. The overall severity score indicated a mild disease, P < 0.001. Mutational analysis revealed absence of M694V homozygosity, P < 0.01, compared to our regular FMF population. We conclude that the onset of FMF in a late age defines a milder form of disease with typical clinical, demographic, and molecular genetic characteristics. PMID- 10528244 TI - Congenital hypoplastic anaemia in a patient with a new multiple congenital anomalies-mental retardation syndrome. AB - We report on a girl with congenital hypoplastic anaemia, "coarse" face, generalized hypertrichosis with scalp hypotrichosis, short fifth finger, hypoplastic toenails, and mental retardation. A sister of the proposita, who died at the age of 1 year, had severe congenital anaemia, hypoplastic fingernails, low birth weight, failure to thrive, and repeated upper respiratory tract infections. Based on family history, we suspect that hypoplastic anaemia and the same multiple congenital anomalies-mental retardation syndrome (MCA/MR) were also present in this sister. To the best of our knowledge, this patient represents the first report of congenital hypoplastic anaemia and such a complex MCA/MR syndrome, probably inherited as an autosomal recessive trait. PMID- 10528245 TI - Molecular cytogenetic resources specific for chromosome 12. AB - We have generated a panel of 20 somatic cell hybrids retaining fragments of human chromosome 12. Each hybrid was characterized cytogenetically by reverse fluorescence in situ hybridization (FISH) and molecularly by 24 sequence tagged sites (STSs) spaced evenly along the chromosome. The panel can be exploited to map subregionally DNA sequences on chromosome 12 and to generate partial chromosome paints useful in the characterization of chromosomal rearrangements involving this chromosome. Furthermore, a panel of 58 yeast artificial chromosomes (YACs) mapping to chromosome 12 was characterized by FISH experiments on normal human metaphases. A subset of this panel is recognized by the STSs used in the somatic cell hybrid characterization. In this way a correlation between the genetic and the physical maps of this chromosome can be established. PMID- 10528246 TI - Trisomy 2p syndrome: a fetus with anencephaly and postaxial polydactyly. AB - We report on a male fetus with partial trisomy 2p21-2pter and monosomy 15q26 15qter due to t(2,15)(p21;q26). This fetus had a typical trisomy 2p phenotype including minor facial anomalies, musculoskeletal defects and two unusual findings: polydactyly and anencephaly. The observation of anencephaly adds support to the theory that genetic material mapping to chromosome band 2p24 is involved in neural tube development. In addition, we propose that a gene on 2p23 may play a role in the morphogenetic patterning of hands and feet. PMID- 10528247 TI - Maternal isodisomy of chromosome 9 with no impact on the phenotype in a woman with two isochromosomes: i(9p) and i(9q). AB - We describe a 34-year-old healthy woman with isochromosomes for the short and long arm of chromosome 9 who was ascertained because of repeated spontaneous abortions. Molecular analysis demonstrated maternal uniparental isodisomy for the whole chromosome 9, thus the origin of the isochromosomes was maternal. Because the patient had a normal phenotype, the maternal isodisomy supports the previous assumption that there are no maternally imprinted genes on chromosome 9. PMID- 10528248 TI - Exclusion of angiotensinogen gene in molecular basis of human hypertension: sibpair linkage and association analyses in Australian anglo-caucasians. AB - Linkage with essential hypertension has been claimed for a microsatellite marker near the angiotensinogen gene (AGT; chromosome 1q42), as has association for the AGT variants M235T, G(-6)A and A(-20)C. To more rigorously evaluate AGT as a candidate gene for hypertension we performed sibpair analysis with multiple microsatellite markers surrounding this locus and using more sophisticated analysis programs. We also performed an association study of the AGT variants in unrelated subjects with a strong family history (two affected parents). For the linkage study, single and multiplex polymerase chain reaction (PCRs) and automated genescan analysis were conducted on DNA from 175 Australian Anglo Celtic Caucasian hypertensives for the following markers: D1S2880-(2.1 cM)-D1S213 (2.8 cM)-D1S251-(6.5 cM)-AGT-(2.0 cM) -D1S235. Statistical evaluation of genotype data by nonparametric methods resulted in the following scores: Single-point analysis - SPLINK, P > 0.18; APM method, P > 0.25; ASPEX, MLOD < 0.28; SIB-PAIR, P > 0. 24; Multipoint analysis - MAPMAKER/SIBS, MLOD < 0.24; GENEHUNTER, P > 0.35. Exclusion scores of Lod -4.1 to -5.1 were obtained for these markers using MAPMAKER/SIBS for a lambda(s) of 1.6. The association study of G(-6)A, A(-20)C and M235T variants in 111 hypertensives with strong family history and 190 normotensives with no family history showed significant linkage disequilibrium between particular haplotypes, but we could find no association with hypertension. The present study therefore excludes AGT in the etiology of hypertension, at least in the population of Australian Anglo-Celtic Caucasians studied. PMID- 10528249 TI - The 3C syndrome: evolution of the phenotype and growth hormone deficiency. AB - The 3C syndrome (cranio-cerebello-cardiac dysplasia or the Ritscher-Schinzel syndrome) is a recently delineated condition involving abnormalities of the cranium (large head with prominent forehead), cerebellum (Dandy-Walker cyst and vermis hypoplasia), and cardiac (primarily septal) defects. At least 20 individuals with this condition have been reported in the past 11 years. We report on a girl with the 3C syndrome who at 13 years of age is the oldest patient reported to date. She has been followed since birth, allowing us to show the evolution of her phenotype over time. In addition, she has documented growth hormone deficiency. We suggest that growth hormone deficiency should be considered as a possible cause of the short stature often seen in this condition. PMID- 10528250 TI - Spinal muscular atrophy variant with congenital fractures. AB - A single report of brothers born to first-cousin parents with a form of acute spinal muscular atrophy (SMA) and congenital fractures suggested that this combination represented a distinct form of autosomal recessive SMA. We describe a boy with hypotonia and congenital fractures whose sural nerve and muscle biopsies were consistent with a form of spinal muscular atrophy. Molecular studies identified no abnormality of the SMN(T) gene on chromosome 5. This case serves to validate the suggestion of a distinct and rare form of spinal muscular atrophy while not excluding possible X-linked inheritance. PMID- 10528251 TI - Novel nonsense mutation of the endothelin-B receptor gene in a family with Waardenburg-Hirschsprung disease. AB - Waardenburg syndrome (WS) comprises sensorineural hearing loss, hypopigmentation of skin and hair, and pigmentary disturbances of the irides. Four types of WS have been classified to date; in WS type IV (WS4), patients additionally have colonic aganglionosis (Hirschsprung disease, HSCR). Mutations in the endothelin-3 (EDN3), endothelin-B receptor (EDNRB), and Sox10 genes have been identified as causative for WS type IV. We screened a family with a combined WS-HSCR phenotype for mutations in the EDNRB locus using standard DNA mutation analysis and sequencing techniques. We have identified a novel nonsense mutation at codon 253 (CGA-->TGA, Arg-->STOP). This mutation leads to a premature end of the translation of EDNRB at exon 3, and it is predicted to produce a truncated and nonfunctional endothelin-B receptor. All affected relatives were heterozygous for the Arg(253)-->STOP mutation, whereas it was not observed in over 50 unrelated individuals used as controls. These data confirm the role of EDNRB in the cause of the Waardenburg-Hirschsprung syndrome and demonstrate that in WS-HSCR there is a lack of correlation between phenotype and genotype and a variable expression of disease even within the same family. PMID- 10528252 TI - Multiple congenital anomalies syndrome: growth and mental retardation, microcephaly, preauricular skin tags, cleft palate, camptodactyly, and distal limb anomalies. Report on two unrelated Brazilian patients. AB - We report on 2 unrelated Brazilian patients, born to non-consanguineous parents, both with multiple anomalies including growth and mental retardation, microcephaly, telecanthus, cleft palate, preauricular skin tags/pit, camptodactyly, and foot anomalies. To our knowledge, this is a previously undescribed formal genesis syndrome. Clinical and genetic aspects are discussed. PMID- 10528253 TI - Blepharophimosis, minor facial anomalies, genital anomalies, and mental retardation: report of two sibs with a unique syndrome. AB - We report on two sibs, a 2.5-year-old girl and a 10-month-old boy, with a hitherto unreported combination of congenital anomalies: blepharophimosis, ptosis, midface hypoplasia, abnormal palate, low anterior and posterior hairlines, displaced hair whorl, apparently low-set and abnormally shaped ears, trigonocephaly, dental anomalies, laryngomalacia, sensorineural hearing loss, genital anomalies, hypotonia, and mental retardation. The occurrence of a similar pattern of anomalies in two sibs of opposite sex suggests autosomal recessive inheritance. To our knowledge, this combination of anomalies has not been reported previously, and thus we propose it to be a formal genesis syndrome. PMID- 10528254 TI - Esophageal, duodenal, rectoanal and biliary atresia, intestinal malrotation, malformed/hypoplastic pancreas, and hypospadias: further evidence of a new distinct syndrome. PMID- 10528255 TI - Culture of fetal erythroid cells from maternal blood using a two-phase liquid system. PMID- 10528256 TI - Screening of the C43G mutation in the promoter region of the XIST gene in females with highly skewed X-chromosome inactivation. PMID- 10528257 TI - Autosomal recessive congenital cerebellar hypoplasia and short stature in a large inbred family. PMID- 10528258 TI - Expansion of a 27 CAG repeat allele into a symptomatic huntington disease producing allele. PMID- 10528259 TI - The golden decade of molecular floral development (1990-1999): A cheerful obituary AB - Cloning of genes involved in the specification of floral meristem and organ identity and in the transition to flowering in some model plants such as Arabidopsis, Antirrhinum, and Petunia during the last decade represents an unprecedented step forward towards an understanding of floral development. Most of these genes belong to conserved and widespread gene families encoding transcription factors, such as the MADS-box genes, FLO-like, and AP2-like genes. Current work on the molecular genetic basis of floral development still focuses on a deeper understanding of the classical model systems, which are all higher eudicots. However, in order to apply the current knowledge about floral developmental genetics to plant breeding and evolutionary biology, flowering plant diversity is now also seriously taken into account. In the next decade, developmental control genes will be studied less and less individually, but rather as components of complex gene regulatory networks. The necessary technology is currently being developed. Learning to understand the origin and evolution of these gene networks will also help to clarify the origin and diversification of flowers, one of the most "abominable" and long-standing mysteries of botany. Copyright 1999 Wiley-Liss, Inc. PMID- 10528260 TI - The gibberellic acid biosynthesis mutant ga1-3 of Arabidopsis thaliana is responsive to vernalization. AB - The Arabidopsis mutant ga1-3 contains a deletion in an enzyme that catalyzes an early step in the synthesis of gibberellic acid. It has been shown that ga1-3 mutant plants cannot flower under 8-h short-day (SD) conditions, even after vernalization. In this article, we present data demonstrating that the ga1-3 mutation does not block the response to vernalization in intermediate photoperiods or in long-day conditions in a late-flowering, vernalization responsive background. Thus, GA may not have a direct role in the vernalization response in Arabidopsis, but it may be required for an alternate pathway that promotes flowering in noninductive photoperiods. PMID- 10528261 TI - The inflorescence architecture of Petunia hybrida is modified by the Arabidopsis thaliana Ap2 gene. AB - We have introduced the Apetala2 (Ap2) gene of Arabidopsis thaliana into Petunia hybrida. Four out of 10 Ap2 transgenic plants flowered and exhibited an altered inflorescence architecture. Internode elongation suggests that the transition from the vegetative to the inflorescence phase does occur, although flower formation is delayed and the cymose branching pattern is not established. Instead, the inflorescence continues to produce bracts and eventually terminates in an aberrant flower with an excess of floral organs. New inflorescence branches then develop from the axillary meristems of the bracts, repeating the formation of a number of bracts before conversion into a terminal, aberrant flower. These results indicate that the Ap2 gene plays a role in the determination of inflorescence meristem identity, but not as a typical A-like function, adding to the existing doubt about the general role of Ap2 gene(s) in floral development. PMID- 10528262 TI - The ASK1 gene regulates development and interacts with the UFO gene to control floral organ identity in Arabidopsis. AB - Normal flower development likely requires both specific and general regulators. We have isolated an Arabidopsis mutant ask1-1 (for -Arabidopsis skp1-like1-1), which exhibits defects in both vegetative and reproductive development. In the ask1-1mutant, rosette leaf growth is reduced, resulting in smaller than normal rosette leaves, and internodes in the floral stem are shorter than normal. Examination of cell sizes in these organs indicates that cell expansion is normal in the mutant, but cell number is reduced. In the mutant, the numbers of petals and stamens are reduced, and many flowers have one or more petals with a reduced size. In addition, all mutant flowers have short stamen filaments. Furthermore, petal/stamen chimeric organs are found in many flowers. These results indicate that the ASK1 gene affects the size of vegetative and floral organs. The ask1 floral phenotype resembles somewhat that of the Arabidopsis ufo mutants in that both genes affect whorls 2 and 3. We therefore tested for possible interactions between ASK1 and UFO by analyzing the phenotypes of ufo-2 ask1-1 double mutant plants. In these plants, vegetative development is similar to that of the ask1-1 single mutant, whereas the floral defects are more severe than those in either single mutant. Interior to the first whorl, the double mutant flowers have more sepals or sepal-like organs than are found in ufo-2, and less petals than ask1-1. Our results suggest that ASK1 interacts with UFO to control floral organ identity in whorls 2 and 3. This is very intriguing because ASK1 is very similar in sequence to the yeast SKP1 protein and UFO contains an F-box, a motif known to interact with SKP1 in yeast. Although the precise mechanism of ASK1 and UFO action is unknown, our results support the hypothesis that these two proteins physically interact in vivo. PMID- 10528263 TI - Ectopic expression of AINTEGUMENTA in Arabidopsis plants results in increased growth of floral organs. AB - AINTEGUMENTA (ANT) was previously shown to be involved in floral organ initiation and growth in Arabidopsis. ant flowers have fewer and smaller floral organs and possess ovules that lack integuments and a functional embryo sac. The present work shows that young floral meristems of ant plants are smaller than those in wild type. Failure to initiate the full number of organ primordia in ant flowers may result from insufficient numbers of meristematic cells. The decreased size of ant floral organs appears to be a consequence of decreased cell division within organ primordia. Ectopic expression of ANT under the control of the constitutive 35S promoter results in the development of larger floral organs. The number and shape of these organs is not altered and the size of vegetative organs is normal. Microscopic and molecular analyses indicate that the increased size of 35S::ANT sepals is the result of increased cell division, whereas the increased sizes of 35S::ANT petals, stamens, and carpels are primarily attributable to increased cell expansion. In addition, 35S::ANT ovules often exhibit increased growth of the nucellus and the funiculus. These results suggest that ANT stimulates cell growth in floral organs. PMID- 10528264 TI - OsMADS13, a novel rice MADS-box gene expressed during ovule development. AB - MADS-box genes have been shown to play a major role in defining plant architecture. Recently, several MADS-box genes have been reported that are highly expressed in the ovule. However, only for the Petunia genes FBP7 and FBP11 has a function in defining ovule identity been shown. We have isolated a rice MADS-box gene named OsMADS13. Expression analysis has shown that this gene is highly expressed in developing ovules. In order to facilitate a detailed characterization of rice ovule-expressed genes, a comprehensive morphological description of ovule development in rice has been performed. The predicted amino acid sequence of OsMADS13 shows significant homology with ZAG2, a maize MADS-box gene, which is also expressed mainly in the ovule. Mapping of the gene in the rice genome showed that it is located on chromosome 12, which is syntenic to two maize regions where ZAG2 and its paralogous gene ZMM1 have been mapped. Our results suggest that OsMADS13 is the ortholog of ZAG2 and ZMM1 and might play a role in rice ovule and seed development. PMID- 10528265 TI - A DEF/GLO-like MADS-box gene from a gymnosperm: Pinus radiata contains an ortholog of angiosperm B class floral homeotic genes. AB - The specification of floral organ identity during development depends on the function of a limited number of homeotic genes grouped into three classes: A, B, and C. Pairs of paralogous B class genes, such as DEF and GLO in Antirrhinum, and AP3 and PI in Arabidopsis, are required for establishing petal and stamen identity. To gain a better understanding of the evolutionary origin of petals and stamens, we have looked for orthologs of B class genes in conifers. Here we report cDNA cloning of PrDGL (Pinus radiata DEF/GLO-like gene) from radiata pine. We provide phylogenetic evidence that PrDGL is closely related to both DEF- and GLO-like genes of angiosperms, and is thus among the first putative orthologs of floral homeotic B function genes ever reported from a gymnosperm. Expression of PrDGL is restricted to the pollen strobili (male cones) and was not detected in female cones. PrDGL expression was first detected in emergent male cone primordia and persisted through the early stages of pollen cone bud differentiation. Based on the results of our phylogeny reconstructions and expression studies, we suggest that PrDGL could play a role in distinguishing between male (where expression is on) and female reproductive structures (where expression is off) in radiata pine. We speculate that this could be the general function of DEF/GLO like genes in gymnosperms that may have been recruited for the distinction between stamens and carpels, the male and female reproductive organs of flowering plants, during the evolution of angiosperms out of gymnosperm-like ancestors. PMID- 10528266 TI - MADS-box genes active in developing pollen cones of Norway spruce (Picea abies) are homologous to the B-class floral homeotic genes in angiosperms. AB - The reproductive organs of conifers, the pollen cones and seed cones, differ in morphology from the angiosperm flower in several fundamental respects. In this report we present evidence to suggest that the two plant groups, in spite of these morphological differences and the long evolutionary distance between them, share important features in regulating the development of the reproductive organs. We present the cloning of three genes, DAL11, DAL12, and DAL13, from Norway spruce, all of which are related to the angiosperm B-class of homeotic genes. The B-class genes determine the identities of petals and stamens. They are members of a family of MADS-box genes, which also includes C-class genes that act to determine the identity of carpels and, in concert with B genes specify stamens in the angiosperm flower. Phylogenetic analyses and the presence of B-class specific C-terminal motifs in the DAL protein sequences imply homology to the B class genes. Specific expression of all three genes in developing pollen cones suggests that the genes are involved in one aspect of B function, the regulation of development of the pollen-bearing organs. The different temporal and spatial expression patterns of the three DAL genes in the developing pollen cones indicate that the genes have attained at least in part distinct functions. The DAL11, DAL12, and 13 expression patterns in the pollen cone partly overlap with that of the previously identified DAL2 gene, which is structurally and functionally related to the angiosperm C-class genes. This result supports the hypothesis that an interaction between B- and C-type genes is required for male organ development in conifers like in the angiosperms. Taken together, our data suggests that central components in the regulatory mechanisms for reproductive organ development are conserved between conifers and angiosperms and, thus, among all seed plants. PMID- 10528267 TI - Morphological and molecular analysis of a double-flowered mutant of the dioecious plant white campion showing both meristic and homeotic effects AB - Many double-flowered plants, in which petals replace stamens, are highly valued by the horticultural industry. These mutants exhibit a homeotic conversion of floral organs and frequently also a meristic increase in floral organ number. By gamma irradiation we generated a novel double-flowered mutant, Sl-dfl, in a male genetic background of the dioecious plant white campion (Silene latifolia). This mutant shows a homeotic conversion of stamens to petals, together with uncontrolled growth and division of second and third whorl floral organ primordia, causing a proliferation of petal and chimeric petal-stamen organs. We characterize this mutant developmentally by scanning electron microscopy and demonstrate that the effects of the mutation commence following the formation of a correctly partitioned floral meristem with a wild-type arrangement of organ primordia. We have commenced a molecular investigation of the Sl-dfl mutant by testing the expression and genomic organization of the known white campion putative MADS-box floral homeotic genes. These studies indicate four MADS-box genes to be unlikely to be mutated in the double-flowered mutant. The possibility that a putative C-function MADS-box gene may cause the mutant phenotype has not currently been excluded, though our morphological studies suggest that a C function mutation is not involved in this case. We conclude that a number of different classes of double-flowered mutation exist, not all of which are currently known from model plant species. This may be indicative of important developmental differences between species and may also emphasize a need for comparative studies of floral development. Copyright 1999 Wiley-Liss, Inc. PMID- 10528268 TI - Flower development in pisum sativum: from the war of the whorls to the battle of the common primordia AB - The ontogeny of pea (Pisum sativum L.) flowers, as in many legume and nonlegume plant species, proceeds through a very different sequence of events from the same process in Antirrhinum majus and Arabidopsis thaliana. Using scanning electron microscopic analysis, we have characterized the early development of wild-type pea flowers and selected morphological characters or markers to break it down into different developmental stages. We used these markers as tools to characterize early alterations in flower development of several pea floral homeotic mutants. These mutants display phenotypes resembling those of: (1) floral meristem identity mutations, frondosus (brac); (2) class A mutations, calix carpellaris (cc); (3) class B mutations, stamina pistilloida (stp-1 and stp 2); and (4) class C mutations, petalosus (pe). According to the homeotic transformations observed in the pea floral mutants, it would appear feasible that the identity and developmental pattern of the four organ types in pea flowers are governed by at least the same three developmental functions, A, B, and C, proposed for the two model systems. However, our results suggest that, in pea, although these functions do have a similar role in the specification of organ identity shown by their counterparts in Arabidopsis or Antirrhinum, they may differ in the control of other processes, such as floral determinacy, organ number, or leaf development. The more remarkable features of pea flower ontogeny were the existence of four common primordia to petals and stamens, the early carpel primordium initiation, and the abaxial-adaxial unidirectional initiation of organ primordia within each different floral whorl, in contrast to the centripetal and sequential floral ontogeny in other plants. Organ differentiation within each of these common primordia appears to be a complex process that plays a central role in the ontogeny of pea flowers. Analysis of flower developmental pea homeotic mutants suggests that A, B, and C functions are necessary for the correct differentiation of organs from common primordia and that, in addition to its role in the specification of petals and stamens, B function, would be involved in conferring common primordia identity. Copyright 1999 Wiley-Liss, Inc. PMID- 10528269 TI - Computer-aided craniofacial surgical planning implemented in CAD software. AB - Accurate presurgical planning is imperative for successful cranial surgery. This article introduces a simulation program developed in a computer-aided design environment. The neurocranium is introduced as a mathematical surface, since this is the part on which the actual operation will be performed. The viscerocranium, which serves as reference, is visualized using small triangular surfaces. The development of the program commenced with a classification of the different surgical techniques mentioned in the literature into six basic actions. The use of mathematically described surfaces has the advantage that the program can simulate actions which change the shape of a surface and perform an on-line estimation of the fracture risk during bending. Three-point bending tests were carried out to provide the necessary data to perform the mathematical check, as these data are not available in the literature. A database with reference distances was introduced to guide the surgeon to obtain the best possible results. During one clinical trial, the computer was taken into the operating room so that the surgical plan developed with the simulation program could be applied to the actual operation. PMID- 10528270 TI - AnatomyBrowser: A novel approach to visualization and integration of medical information. AB - In this article, we present a novel technique for visualization of three dimensional (3D) surface models, as well as its implementation in a system called AnatomyBrowser. Using our approach, visualization of 3D surface models is performed in two separate steps: a pre-rendering step, in which the models are rendered and saved in a special format, and an actual display step, in which the final result of rendering is generated using information from the prerendering step. Whereas prerendering requires high-end graphics hardware, the final image generation and display can be implemented efficiently in software. Moreover, our current implementation of AnatomyBrowser interface uses the Java programming language and can therefore be readily run on a wide range of systems, including low-end computers with no special graphics hardware. In addition to visualization of 3D models and 2D slices, AnatomyBrowser provides a rich set of annotation and cross-referencing capabilities. We demonstrate several possible applications for AnatomyBrowser, including interactive anatomy atlases, surgery planning, and assistance in segmentation. PMID- 10528271 TI - Impingement simulation of the hip in SCFE using 3D models. AB - OBJECTIVE: Affecting as it does the geometry of adolescent hips, slipped capital femoral epiphysis (SCFE) and its evaluation represent a major three-dimensional problem. The current methods of clinical assessment-geometric measurements of the femur on plain radiographs or on axial computed tomographic (CT) cross-sections address only one of the two joint components. MATERIALS AND METHODS: We have developed a system to simulate motion of hip joints with physiologic joint contact. In our system, CT-based computer models of the femur, pelvis, etc., are fitted with oriented bounding boxes (OBBs) and manipulated. Collision detection algorithms control the hip motion, which, in this virtual joint, is based on the surface geometry of the joint partners rather than on a predefined fixed rotation center. RESULTS: An illustrative case is presented to show the advantages of the new biomechanical evaluation method over conventional radiological assessments for SCFE. The proposed system provides remarkably high speed, and the necessary data can be prepared in a reasonable time. CONCLUSION: The range-of-motion assessment provides the surgeon with information about the site and the impact of nonphysiologic contact in the hip joint. The information thus obtained can be valuable for indication and planning of corrective surgery in cases of SCFE. PMID- 10528272 TI - Abstracts from the 4(th) symposium on computer assisted orthopaedic surgery (CAOS'99), held in davos, switzerland on march 18(th) and 19(th), 1999* PMID- 10528273 TI - First announcement and call for papers, 5th international symposium on CAOS, computer assisted orthopaedics surgery, february 17-19, 2000, davos, switzerland PMID- 10528274 TI - Focus groups. PMID- 10528275 TI - Normal development of a zona-free oocyte to the blastocyst stage following ICSI. PMID- 10528276 TI - [What is the effect of pars plana vitrectomy in diabetic traction detachment with macular involvement on the social status of the patient?]. PMID- 10528277 TI - [Myopia due to bedroom lighting in young children?]. PMID- 10528278 TI - [What is the value of transpupillary thermotherapy in treatment of flat posterior choroid melanomas? A systematic review of the literature?]. AB - BACKGROUND: Transpupillary thermotherapy is a relatively new method for the treatment of choroidal melanomas. We present a systematic survey of the current literature. METHOD: A temperature rise in the tumor ranging from 45-60 degrees C is achieved by an infrared laser beam delivered through the dilated pupil. With a modified delivery system beam widths between 1 and 3 mm and exposure times of one minute are generated. Thus, tumors of up to 4 mm thickness are treatable. TTT can be used as a single treatment procedure or in combination with brachytherapy. RESULTS: Several studies presented in the literature show a satisfactory local tumor control. However, there is a significant risk of vision threatening side effects like retinal vascular occlusion or retinal traction in selected cases. CONCLUSION: The TTT is a minimal invasive procedure for the treatment of flat choroidal melanomas of the posterior pole which is capable of achieving a good local tumor control. Studies with more patients and longer follow-up will demonstrate if TTT is also beneficial in the longterm management of choroidal melanomas. PMID- 10528279 TI - [Expulsive hemorrhage in perforating keratoplasty--incidence and risk factors]. AB - BACKGROUND: Expulsive hemorrhage is a severe complication of intraocular surgery. In a retrospective study we looked up surgical records of all penetrating keratoplasties (PK) performed in our department between 1989 and 1998. In all patients suffering from expulsive or preexpulsive choroidal hemorrhage we intended to find out possible risk factors and to report on the final outcome of these eyes. PATIENTS AND METHODS: Between January 1989 and November 1998 a total of 2421 PKs were performed. Nine preexpulsive and three expulsive hemorrhages occurred. The group of patients with preexpulsive hemorrhage consists of four females and five males (mean age 57 +/- 6.5 years). Three patients (one female, 2 males; mean age 67 +/- 8.5 years) suffered from expulsive hemorrhage. RESULTS: Incidence of expulsive hemorrhage was 0.1% (preexpulsive hemorrhage 0.4%). All twelve operations were performed under general anesthesia. Risk factors for preexpulsive hemorrhage were: previous ocular surgery (three patients), ocular trauma (2 patients) and internal diseases (five patients): arterial hypertension, coronary heart disease, diabetes. Preoperative visual acuity was light perception to 4/20, visual acuity at the last postoperative examination after a mean follow up of 41.0 +/- 22.6 months ranged from light perception to 12/20. Risk-factors for expulsive hemorrhage were: previous ocular surgery (two patients), primary open angle glaucoma (two patients), coronary heart disease (one patient) und asthma bronchiale (one patient). One of the patients awoke from general anesthesia during the "open-sky" situation. Preoperative visual acuity was light perception to 2/400, visual acuity at the last postoperative examination (mean follow-up 14.0 +/- 1.0 months) was light perception in all eyes. CONCLUSION: The incidence of an expulsive hemorrhage was 0.1% in 2412 Pks. Risk factors are ocular and internal predispositions which can hardly be controlled, although arterial blood pressure was not significantly elevated during opening of the globe. PMID- 10528280 TI - [Minimizing induction of astigmatism in preoperative spherical cornea a. by mini incision surgery with foldable IOL and b. by corneal tunnel incision with limbal relaxing incision]. AB - BACKGROUND: In case of a spheric cornea preoperatively the refractive effect of a clear corneal cataract incision is undesirable. We studied two actual techniques to minimize the surgically induced astigmatism. PATIENTS AND METHODS: Temporal clear corneal incision was performed in 77 patients with practically spherical cornea (0.2 +/- 0.1 D). 27 patients with 4.1-mm clear corneal stretch incision and 5 mm PMMA lens implantation served as control. 25 further patients were operated on with the same technique, but 2 additional limbal relaxing incisions (LRI) of 0.55-mm depth and 8 mm length at 6 and 12 o'clock were performed. In 25 patients a foldable acrylic lens (Acrysof) was implanted through a 3.2-mm temporal clear corneal incision. Corneal topography results were evaluated in all patients by the Jaffe and the Holladay analysis. RESULTS: The surgically induced astigmatism of 0.8 +/- 0.5 dpt in the control group was reduced to 0.4 +/- 0.3 dpt by LRI and by reduction of the incision size as well in the treatment groups. With-the-wound-change (WTW) in the Holladay analysis was 0.6 +/- 0.7 dpt in the control group and around 0 in the groups with astigmatism reducing techniques. CONCLUSION: To preserve a spherical cornea in clear corneal-tunnel incision, compensating limbal relaxing incisions (LRI) or ultra-small incisions with foldable lens implantation should be performed. PMID- 10528281 TI - [Calculating the localization and dimension of the real pupil in keratoconus with ray tracing of corneal topography data]. AB - BACKGROUND: It is crucial to center surgical procedures for optical indications on the pupil or the optical axis of the eye. In keratoconus the pupil appears to be dislocated due to optical aberrations of corneal topography. The purpose of this study was to evaluate the real pupil structure from the virtual image using exact raytracing techniques. PATIENTS AND METHODS: Eighty-eight patients with keratoconus (46 with mild and 42 with severe clinical signs) and a control group of 40 normal subjects were included in this study. Topographic height data were calculated from refraction data of a commercially available topographer (TMS-1) using a local approximation algorithm and a convex surface was modelled using a subdivision scheme. For the posterior corneal surface we postulated an aspherical surface with a central radius of curvature of 6.5 mm using Navarro's model eye. At the virtual pupil outline a bundle of parallel rays were intersected with the anterior and posterior corneal surface and refracted into the anterior chamber. The intersections of these rays with the pupil plane was defined as the real pupil outline. We assessed the amount and direction of pupil dislocation, the ratio between the virtual and real pupil size for each group and correlated these parameters with the central corneal power. RESULTS: The size of the virtual pupil exceeded the reference value of the real pupil in the normal group by 11%, in the group with mild keratoconus by 19% and in the group with severe keratoconus by 35%. The center of the virtual pupil was decentered 0.06 mm in the normal group, 0.49 in the group with mild keratoconus and 1.24 mm in the group with severe keratoconus. Whereas the direction of decentration was randomly in the normal group, we measured a preferred decentration to the inferior quadrants in mild keratoconus and a systematic decentration to the temporal inferior quadrant in severe keratoconus. Correlation of the optical dislocation did not correlate with central corneal power in any group. CONCLUSIONS: In keratoconic eyes the pupil outline is distorted and dislocated due to optical aberrations of the cornea. Exact raytracing technique allows the calculation of the real pupil outline from the virtual image and the topographic height of both corneal surfaces. Knowledge about the real pupil position may have an impact on adequate centration of keratorefractive surgery and penetrating keratoplasty. PMID- 10528282 TI - [Transplant endothelium and measuring corneal thickness after non-high-risk keratoplasty with briefly or long-term preserved corneal donor tissue]. AB - PURPOSE: The corneal endothelial cell density is essential for the pump function and the transparency of grafts after penetrating keratoplasty (PK). The purpose of this study was to assess corneal endothelial cell density after non-high-risk PK and to check for possible correlations with storage parameters of the donor corneas using two different storage methods. PATIENTS AND METHODS: Endothelial cell density (specular microscope EM 1100, TOMEY, Erlangen) and central corneal thickness (ultrasonic pachymetry SP-2000, TOMEY, Erlangen) were assessed 6 weeks, 3, 6, 9 months and one year postoperatively in 168 non-high-risk PKs. Short-term preserved donor corneas were used in 89 patients, whereas in 79 patients organ cultured corneas were used. The donor trephination was performed from the epithelial side using an artificial anterior chamber. The postoperative treatment with topical steroids was standardized. The mean donor post-mortem time was 9.6 +/- 8.0 hours for short-term-preserved and 17.6 +/- 10.5 hours for organ-cultured corneas (p < 0.0001). The storage time was 71 +/- 49 and 380 +/- 167 hours (p < 0.0001), respectively. RESULTS: Endothelial cell density did not differ significantly between the two storage methods (p > 0.05). At 6 weeks postoperatively, the mean endothelial cell density was 2042 +/- 675 cells/mm2 for short-term-preserved corneas and 1972 +/- 522 cells/mm2 for organ-cultured corneas (p = 0.7). Endothelial cell density did not decrease significantly (p > 0.05) within the observation period of 12 months in both groups (after 12 months: 1868 +/- 957 cells/mm2 and 1638 +/- 643 cells/mm2, respectively). The mean corneal thickness was 542 +/- 50 microns for short-term-preserved and 541 +/- 55 microns for organ-cultured corneas and remainded unchanged during the follow-up of 12 months (542 +/- 42 microns and 521 +/- 43 microns, respectively). Neither the group of short-term-preserved corneas nor organ-cultured corneas showed a significant correlation between endothelial cell density or central cornea thickness with post-mortem time or with storage time of the donor corneas at any postoperative stage (p > 0.1). CONCLUSION: During the first year after PK, only a small decrease in endothelial cell density was observed in comparison with the 6 weeks finding. The storage method does not seem to affect the short-term changes of endothelial cell density. Further long-term studies are necessary to assess the clinical relevance of these observations. PMID- 10528283 TI - [Clinical significance of objective vision assessment using visually evoked cortical potentials induced by rapid pattern sequences of different spatial frequency]. AB - BACKGROUND: In patients where reliable subjective assessment of visual acuity is impossible, further diagnostics should be enhanced by an objective method. PATIENTS AND METHODS: A group of 34 patients was examined by objective assessment of visual acuity using visual evoked potentials (VEP) as described by Hajek and Zrenner in 1988. The presentation of five checkerboards with different spatial frequency in repetitive sequences on a TV-monitor elicits a series of transient visual evoked potentials. Shape and amplitude of each wavelet depends on check size and directly reflect a spatial tuning function with a low- and high frequency cut-off. This amplitude is described by a polynomial fit (2nd order). The function's intersection with the x-axis at higher spatial frequencies leads to an estimation of the visual acuity. RESULT: This result is compared to the subjectively determined visual acuity. In the majority of the presented cases the suspected malingering was confirmed. CONCLUSION: Patients with suspected malingering represent the primary indication of the described method. PMID- 10528284 TI - [Initial clinical experiences with the Dresden 3D display in conjunction with the Heidelberg Retina Tomograph (HRT)]. AB - BACKGROUND: For years the Heidelberg Retina Tomograph has been an established method to diagnose early glaucomatous damages at the optic nerve head. The major difficulty consists in defining the outlayer of the optic nerve head in a 2 dimensional reflective or topographic picture. A 3-dimensional presentation of the ocular fundus could ease the defining very much. The Dresden 3D-display tested provides a true 3-dimensional presentation of the HRT-values measured. METHODS: For the study 5 groups of prediagnosed follow-up examinations were formed, which in their course showed various progression. The examination firstly was carried out by manual defining of the papilla contour on the Dresden 3D display and secondly by the new automatic contour-finding of the 3D-software. RESULTS: Through the 3-dimensional presentation a more correct position of the contour can be found when drawn manually. As a result some diagnoses of the 2 dimensional HRT-pictures had to be revised. The results proved considerable differences between the automatic outlayer-finding and the probably real position of the papilla borders. CONCLUSION: The 3-dimensional presentation of the HRT pictures represents an subjective improvement regarding the accuracy of manual contour definition. In particular, the fact that it is feasible to make the pictures rotate around the axes offers completely new insights to the morphology of the disc. PMID- 10528285 TI - [Continuous intraocular pressure measurement over several days with the Codman Microsensor--a case report]. AB - BACKGROUND: In eyes with irregular corneal surface (e.g. following bullous keratopathy, irregular astigmatism, edema or scars and following perforating keratoplasty), applanation tonometry often cannot be performed or results do not correlate with the clinical findings. In these cases, intraocular measurement of intraocular pressure is necessary. By puncturing of the anterior chamber, single measurements can be done for a short time period. Data of the course of intraocular pressure for a long period of time can not be assessed by this method. We report on two patients whom we implanted a continuously measuring probe into the anterior chamber for up to 96 hours instead of puncturing the anterior chamber. PATIENTS AND METHODS: A neurosurgical micro sensor (CODMAN, Norderstedt) was placed into the anterior chamber via a 1.2 mm wide and 4 mm long scleral tunnel. The data were transmitted to the ICP Express Display Monitor (CODMAN, Norderstedt) and displayed. From there, the data were transmitted to the multifunctional monitor DINAMAP Plus 8720 (CRITIKON, Norderstedt). After analog digital transformation, the data were recorded on a personal computer with Pentium processor for analysis (patient #1: one measurement per minute, patient #2: one measurement per 10 seconds). In the first patient, implantation of the probe was indicated by enormous deviation of applanation tonometric measurements (12 to 20 mmHg) from the measurement results with the finger tips (25 to 30 mmHg). Clinical findings correlated to the higher intraocular pressure. Due to a decompensation of the corneal transplant, a re-keratoplasty was necessary. Within this operation, the micor sensor for continuous measurement of the intraocular pressure was implanted. The probe was explanted the next day. In the second patient, an primary chronical open-angle glaucoma in both eyes was known. In 1997, corneal transplantation has been performed in both eyes due to corneal dystrophy. Postoperatively, intraocular pressure stayed high. Applanation tonometry gave measurements of 16 to 20 mmHg although the measurement results with the finger tips exceeded 30 mmHg. To find out the real intraocular pressure and to have a basis for a rational therapy, we implanted the intraocular measurement probe for five days to determine the intraocular pressure at night and day. After measuring the baseline values, the efficiency of several antiglaucomatous drugs was tested to find out the drugs with the highest effect to prescribe it to the patient after the removal of measuring probe. RESULTS: The intraocular measurement with the CODMAN micro sensor could confirm in both patients that the measurements by applanation tonometry were wrongly too low. The measurement results with finger tips were confirmed by the intraocular measuring. The data had essential implications for the patients. Meanwhile, in both patients pressure lowering surgery was performed. The probe did not cause intraocular problems (1 day respectively 5 days). An irritation of the anterior chamber did not appear. In the first patient, the measuring probe moved from its position with following external filtration. So the probe was explanted only the next day. A movement of the probe tip can be avoided be appropriate subconjunctival suture fixation. CONCLUSION: Continuous measuring and recording of the intraocular pressure may be indicated, if applanation tonometry gives unreliable or even wrong results. Via a long scleral tunnel, a water-proof implantation into the anterior chamber is possible. Because a postoperative irritation could not be seen, we think that the probe only causes a minor falsification of the intraocular pressure. The described pressure measuring system allows measuring intraocular pressure continuously and assessing the individual effect of different antiglaucomatous drugs. Before using the probe as routine procedure, some improvements are necessary, e.g. smaller tip of the probe. The transmission wire to the PMID- 10528286 TI - [Orbital hemorrhage as a sequela of heparin-induced thrombocytopenia]. AB - BACKGROUND: Spontaneous orbital hemorrhage is a rare but sight threatening incident. Several causative factors are known. Infrequently, hemorrhagic ocular complications can be associated with heparin therapy. The mechanism can involve a immunologically caused thrombocytpenia. In every hemorrhagic ocular complication which occurs in patients treated with heparin Heparin-induced thrombocytopenia (HIT) must be taken into consideration. PATIENT: A 76-year-old woman who underwent hip surgery three weeks prior to a spontaneous orbital hemorrhage had been treated with subcutaneous heparin. CT scan of the right orbit showed peribulbar hemorrhage with the main focus retro-lateral of the bulbus. Retrobulbar expansion of the hemorrhage was only slight with no compression of the optic nerve. There was no sign of tumor or anomaly of blood vessels. Blood cell count revealed a marked thrombocytopenia with 8000/microliter. After discontinuation of heparin therapy, thrombocytes' cell counts rised rapidly with 114,000/microliter after three days. Parallel to the normalization of the thrombocytes' cell counts the clinical symptoms disappeared quickly. CONCLUSIONS: Heparin-induced thrombocytopenia as a cause for spontaneous orbital hemorrhage has never been reported before. For differential diagnosis in patients with hemorrhagic ocular complications, HIT must taken into account. The most important measure is counting thrombocytes. In thrombocytopenia, heparin therapy must be discontinued before hemorrhagic or thromboembolic complications occur. PMID- 10528287 TI - [Mycophenolate mofetil after penetrating high risk keratoplasty. A pilot study]. AB - AIM: Mycophenolate mofetil (MMF, CellCept) has become a successful part of the standard immunosuppression regimes after solid organ transplantation. It was the aim of this study to compare the efficacy and the safety of MMF after penetrating high-risk keratoplasty with our standard immunosuppression, i.e. systemic cyclosporin A (CSA), in a pilot study. PATIENTS AND METHODS: Sixteen patients after penetrating high-risk keratoplasty were randomized to be treated either with MMF or with CSA for six months postoperatively. MMF was administered in an oral dose of two times 1 g daily whereas the CSA dose varied according to the blood trough levels of 120-150 ng/ml (monoclonal TDx) between 100 and 500 mg daily. RESULTS: During this first follow-up period of 11.4 (5-18) months neither in the MMF- nor in the CSA-group irreversible graft failure was observed. One serious acute endothelial immune reaction was observed in the CSA-group after systemic immunomodulation had been tapered. It was treated successfully with systemic and topical corticosteroids. In one patient CSA-prophylaxis had to be stopped five months postoperatively because of elevated liver enzymes. Side effects did not occur in the MMF-group. CONCLUSIONS: Up to now MMF has been evaluated to be as efficacious as CSA and safe. If these results are confirmed in the long run in this study MMF may become an armament to avoid immune reactions in high-risk penetrating keratoplasty patients who must not receive systemic CSA. PMID- 10528288 TI - [Dermatitis and conjunctivitis after contact with Euphorbia myrsinites (wolf's milk extract)--a case report]. AB - BACKGROUND: Fresh sap of euphorbiaceae leads to a toxic burn of the skin and the eyes. Since years the sap of euphorbiaceae has been used in the treatment of different kinds of verrucas. PATIENTS: After contact with the sap of Euphorbia myrsinites three children developed a toxic dermatitis. In addition, the youngest girl showed a conjunctivitis and an occlusion of the right eye. Phorbolesters are considered to be responsible for the toxicity of the euphorbiaceae. All three children have resulted in a restitutio ad integrum. CONCLUSION: This case report is demonstrating the danger of toxic burn of this kind of plant. PMID- 10528289 TI - Recombinant human thyrotropin (rhTSH) in the management of differentiated thyroid carcinoma. PMID- 10528291 TI - Can reduced imaging times be used for scintimammography? AB - A reduction in acquisition times from 10 to 5 min for scintimammography does not reduce the diagnostic value of the study when imaging for detection of breast lesions. The test showed an overall sensitivity of 96%, specificity of 100% and accuracy of 98% for breast lesions visualized on both 5 and 10 min acquisitions. Even if lymph node detection is the primary concern of the study, longer scan times do not increase the sensitivity of the test (40% on both 5 and 10 min). Scintimammography is poor at detecting lymph nodes (sensitivity 40%, specificity 69-82%) and is not useful for assessing lymph node involvement. A reduction in imaging times appeared to be consistent on both camera systems tested, which have very different display outputs. This would indicate that other departments may be able to reduce times on their systems without affecting the quality of the study. The detection of breast lesions was also consistent between reporters. PMID- 10528290 TI - Enhanced uptake of 99Tcm-MDP in skeletal metastases from prostate cancer following initiation of hormone treatment: potential for increasing delivery of therapeutic agents. AB - Following androgen ablation therapy, skeletal metastases from prostate cancer appear in some instances to show an increase in 99Tcm-methylene diphosphonate (99Tcm-MDP) uptake. Such a phenomenon could represent a mechanism to increase delivery of bone-seeking therapeutic agents to skeletal metastatic sites. The aim of this study was to characterize more precisely the potential increase in 99Tcm MDP in skeletal metastases from prostate cancer following initiation of hormone therapy. Baseline bone scans were performed within 1 week of onset of hormone therapy in patients with stage D2 prostate cancer followed by multiple repeat bone scans for up to 4-6 weeks. The count density within metastatic lesions was divided by the average count density from several areas of normal bone to obtain a lesion to normal bone uptake ratio (L/N) for each lesion in each scan. Altogether, 61 skeletal metastases were identified on bone scans from five subjects. Eighty-four percent (51/61) of these lesions showed an increase in 99Tcm-MDP activity relative to normal bone following initiation of hormone therapy with a mean peak increase of 39%. Thirty-nine of these 51 metastatic lesions showed maximum uptake at 3 weeks post-onset of hormone treatment. From our findings, it appears that approximately 3 weeks following initiation of hormone blockade, most skeletal metastases from prostate cancer will demonstrate significantly enhanced 99Tcm uptake relative to normal bone. Consequently, it may be possible to improve the uptake and effectiveness of therapeutic bone-seeking radiopharmaceuticals by administering these agents following hormone therapy in patients with prostate cancer metastases. PMID- 10528292 TI - The additive values of left ventricular function and extent of myocardium at risk to dipyridamole perfusion imaging for optimal risk stratification prior to vascular surgery. AB - Although the increased risk of cardiac complications in surgical patients with diminished left ventricular ejection fraction (LVEF) is well-established, this method has been supplanted in recent years by assessment of ischaemic burden using myocardial perfusion imaging (MPI). This study was conducted to determine if MPI and LVEF determination provide complementary or redundant information in preoperative evaluation of vascular surgery patients. A total of 101 patients were studied with dipyridamole MPI and radionuclide ventriculography before surgery. Single photon emission tomographic MPI images were scored for defect severity and categorized as either fixed or reflecting ischaemia. Resting left ventricular cavity was also categorized as normal or dilated. LVEF was subdivided into normal (> or = 50%) and abnormal (< 50%). Seventeen patients had cardiac events. Events were more frequent in patients with ischaemia, in patients with a LVEF < 50% and in those with dilated left ventricular chambers. The mean number of ischaemic segments was also higher in the cardiac event group. Higher event rates were seen when a combination of these factors was present. A history of myocardial infarct, congestive heart failure or coronary artery disease was also a significant predictor of subsequent events. Thus, both abnormal left ventricular function and extent of ischaemic myocardium have independent and complementary predictive power for cardiac events in vascular surgery patients. PMID- 10528293 TI - A report system for PET assessment of myocardial viability. AB - Coronary bypass surgery can improve the prognosis of patients with heart failure due to coronary artery disease. However, these patients have a high operative risk and should be operated on only if they have a sizeable amount of viable tissue (i.e. asynergic myocardium) that can recover contractile function following coronary revascularization. In the clinical setting, regional wall motion is usually evaluated by two-dimensional echocardiography, whereas retained metabolic activity assessed by positron emission tomography (PET) and 18F fluorodeoxyglucose is a well-established means for the evaluation of myocardial viability. Unfortunately, the two-dimensional echocardiography and PET reports are often different, and this renders the matching of information difficult and the estimation of the risk-benefit ratio of the operation unreliable. In this paper, we present a report system for the evaluation of myocardial viability with PET. We divided the left ventricle into 16 segments following the proposal of the American Society of Echocardiography for wall motion analysis by two-dimensional echocardiography. Following this partition, three portions of the left ventricle are identified along the long axis: basal, mid and apical. Each plane of the basal and mid portions is automatically divided into six segments with the super imposition of a radial divider over the short-axis images. Similarly, each plane of the apical portion is automatically divided, but into four segments. This partition of the left ventricle permits a precise match between the information on wall motion obtained with two-dimensional echocardiography and that on viability obtained with PET. PMID- 10528294 TI - Planar scintigraphy and SPET with 99Tcm-HMPAO-labelled leukocytes in patients with median sternotomy: normal patterns. AB - The aim of this study was to determine the normal planar and SPET patterns of the thoracic distribution of 99Tcm-hexamethylpropylene amine oxime (99Tcm-HMPAO) in 20 patients who had undergone a previous median sternotomy and without infectious complications at follow-up. The study included anterior and oblique anterior planar views at 4 and 20 h. SPET of the chest was also carried out at 4 and 20 h. At 4 h, the planar views showed low background vascular activity in the lungs and cardiac region in addition to the sternal uptake, which showed two patterns: homogeneous in five patients and heterogeneous in 15. A long and narrow defect of uptake along the sternal midline was the most characteristic finding. At 4 h, in addition to the background vascular activity in the lungs and cardiac region, the greatest uptake on SPET was in the sternum anteriorly and the marrow spine posteriorly without any focal uptake, allowing visualization of the mediastinum free of focal activity. At 20 h, both the planar and SPET images showed a higher organ-to-background ratio. Knowledge of these post-surgical patterns will make it easier to interpret planar and SPET images when 99Tcm-HMPAO-labelled leukocytes are used in the diagnosis of mediastinitis and sternal infections in patients who had previously undergone median sternotomy. Planar views were better for the assessment of sternal uptake, but SPET views were better for the direct visualization of the mediastinum by eliminating overlapping sternal uptake. PMID- 10528295 TI - The relation between the degree of gastro-oesophageal reflux and the rate of gastric emptying. AB - In this study, we evaluated the relationship between the degree of gastro oesophageal reflux and the rate of gastric emptying and determined the variability of gastric emptying in children. The reproducibility of radionuclide imaging for the presence and grading of gastro-oesophageal reflux was also examined. Twenty-eight children less than 2 years of age participated in the study. For assessment of variability, all subjects underwent two scintigraphic studies. For each study, the number of reflux episodes and gastric emptying half times were recorded. The amount of reflux was graded according to the classification suggested by Blumhagen. Patients with grade 1 reflux were considered low-grade refluxers, while patients with grade 2 or 3 reflux were considered high-grade refluxers. The level of reflux for each patient was based on the highest reflux grade recorded in either study. Of the 28 patients, 19 had reflux in at least one study. Ten patients had high-grade and nine patients low grade reflux. All patients but one with high-grade reflux had the same grade of reflux in both studies (90%). Of nine patients with low-grade reflux, three had the same grade in both studies. The mean half-time was significantly higher for high-grade than for low-grade refluxers (P < 0.05). For subjects with low-grade reflux, this value did not differ significantly from that of non-refluxers (P > 0.05). Our results show that patients with high-grade gastro-oesophageal reflux had prolonged gastric emptying. The inter- and intra-subject variability of gastric emptying in children appeared to be low. Reproducibility for the presence and grading of gastro-oesophageal reflux by the radionuclide method was good, with the highest value being for the diagnosis of high-grade gastro-oesophageal reflux. PMID- 10528296 TI - Estimation of risk based on biological dosimetry for patients treated with radioiodine. AB - A multicentre study was undertaken to assess the cytogenetic damage to peripheral blood lymphocytes in 31 patients treated with 131I for thyrotoxicosis using the cytokinesis-blocked micronucleus assay. The results were compared to those for eight thyroid carcinoma patients using the same method. For each patient, blood samples were taken immediately before and 1 week after iodine administration. The first blood sample was divided into three fractions and each fraction was subsequently irradiated in vitro with 0, 0.5 and 1 Gy 60Co gamma rays, respectively. After blood culture for 70 h, cells were harvested, stained with Romanowsky-Giemsa and the micronuclei scored in 1000 binucleated cells. For both patient groups, a linear-quadratic dose-response curve was fitted through the data set of the first blood sample by a least squares analysis. The mean increase in micronuclei after 131I therapy (second blood sample) was fitted to this curve and the mean equivalent total body dose (ETBD) calculated. Surprisingly, in view of the large difference in administered activity between thyroid carcinoma patients and thyrotoxicosis patients, the increase in micronuclei after therapy (mean +/- S.D.: 32 +/- 30 and 32 +/- 23, respectively) and the equivalent total body dose (0.34 and 0.32 Gy, respectively) were not significantly different (P > 0.1). The small number of micronuclei induced by 131I therapy (32 +/- 29), compared with external beam radiotherapy for Hodgkin's disease (640 +/- 381) or cervix carcinoma (298 +/- 76) [1], gave a cancer mortality estimate of less than 1%. This also explains why late detrimental effects in patients after 131I treatment have not been reported in the literature. PMID- 10528297 TI - Radiation protection in radioguided surgery of breast cancer. AB - The protocols for sentinel lymph node biopsy and radioguided occult lesion localization could potentially be of great value in the management of breast cancer patients. Both involve the injection of a 99Tcm-labelled radiopharmaceutical close to or into the lesion, localization of the sentinel lymph node or occult lesion by scintigraphy, and surgical removal with the aid of a hand-held gamma-ray detector. We present dosimetric data on patients and hospital personnel involved in these procedures. For evaluation of radiation protection, we measured the absorbed dose and air kerma rate. Activity levels in excised tissues and surgical instruments were also determined. For patients, the mean absorbed dose to the abdomen was 0.45 mGy, which is low compared to doses received from other diagnostic examinations. For surgeons after 100 operations, the mean absorbed dose to the hands was 0.45 mGy and the mean effective dose 0.09 mSv. Absorbed doses to all hospital personnel involved in the procedures were very low compared to recommended annual limits stipulated by the International Commission on Radiological Protection. We conclude that these procedures, performed according to protocols laid down by the European Institute of Oncology, Milan, are safe from the point of view of radiological protection and that only routine precautions are necessary. PMID- 10528298 TI - Kinetic parameter estimation from compartment models using a genetic algorithm. AB - Kinetic parameters were estimated from a three-compartment fluorodeoxyglucose model with three rate constants using a genetic algorithm. The performance of the genetic algorithm was investigated by simulation studies, in which brain time activity data (TAD) were generated using cited mean values of rate constants and the plasma TAD obtained from positron emission tomographic studies. The accuracy of kinetic parameter estimation using the genetic algorithm was compared with that using the non-linear least-squares (NLSQ) method. The margin of error in the parameters estimated using the genetic algorithm tended to be smaller than that obtained by the NLSQ method. Although not statistically significant at a noise level of 5% in the brain TAD, the difference between the two methods became significant for all parameters at a noise level of 15% or higher. Our results suggest that the genetic algorithm is a promising means of estimating kinetic parameters from compartment models, because it is more robust against statistical noise than the NLSQ method and it can be rendered highly parallel for processing. PMID- 10528299 TI - 13C-urea versus 14C-urea breath test--which is the safer? PMID- 10528300 TI - [Neuropathological findings in early-onset Alzheimer's disease (E280a-PS1 mutation)]. AB - INTRODUCTION AND MATERIAL: Nine brains belonging to early onset Alzheimer disease (E280A-PS1 mutation) affected individuals from Antioquia, Colombia, were analyzed by neuropathological standard techniques. All individuals were ascertained from genealogies descendents from a common ancestor that shows a dominant autosomical pattern of inheritance. RESULTS: All cases analyzed were carriers to the E280A PS1 mutation. This type of mutation produce beta-amyloid deposits of 42 aminoacids in the CNS. The mean of onset age was 48.4 years with an average of evolution time of 7.55 years and a mean of death age of 56.55. Although, all the cases showed symmetrical atrophy and them was more severe in the hippocampal region, a definitive anterior pattern (temporo-frontal) was showed. The higher the time of evolution of disease the lower the brain weight. CONCLUSIONS: All types of senile plaques and abundant neurofibrillary tanggles were found. In the stem, similar lesions were found but they were in lower number. Only the mesencephalic region showed a significative positive correlation between the number of senile plaques and the number of neurofibrillary tanggels (p < 0.05, r = 0.76). Only the parietal region showed a significant positive correlation between the number of senile plaques and the disease evolution time (p < 0.02, r = 0.74). Particularly, the cerebellum only showed senile plaques but neurofibrillary degeneration was not observed. With the exception of the Hirano bodies, all findings traditionally described were observed. PMID- 10528301 TI - [Image characterization of Alzheimer's disease associated with the E280A-PS1 mutation. Case-control study: MRI findings]. AB - INTRODUCTION AND METHODS: In order to compare the magnetic resonance image characteristics of individuals belonging to pedigrees carrying the mutation E280A PS1 associated to early onset Alzheimer disease, coming from Antioquia, Colombia, 78 individuals were studied. 47 of them were carriers of the mutation, 23 of those presented symptoms and 31 individuals being controls (non carriers of the mutation). RESULTS: In summary, significative differences were appreciated between symptomatic individuals and those asymptomatic. There was not significant difference between asymptomatic carriers and the controls. The presence of the perihippocampal fissure constituted a difference statistically significant between the symptomatic individuals and those carriers asymptomatic and between the symptomatic ones and the controls. The interuncal distance increased significantly was another difference between symptomatic and asymptomatic individuals and among symptomatic and control group. The lobar atrophy and the ventriculomegaly were found in symptomatic individuals and they correlate with the disorder graveness. There was not significance in the presence of infarcts and/or hippocampal hyperintensities. CONCLUSION: These results corroborate the statement that magnetic resonance image is very useful in the diagnosis and follow-up of individuals affected by early onset Alzheimer disease. PMID- 10528302 TI - [Serum pro-oxidant and antioxidant factors and risk of Parkinson's disease: population study]. AB - INTRODUCTION: Several studies suggested a role of 'oxidative stress' (increased production of prooxidants, antioxidants deficiencies or both) in the pathogenesis of Parkinson's disease. In this study we have measured the serum levels of a number of prooxidant and antioxidant substances to evaluate their possible relation with the risk for Parkinson's disease. PATIENTS AND METHODS: We assessed the serum levels of iron, ferritin, ansferrin, ceruloplasmine, vitamin A, alpha carotene, beta-carotene, and alpha-tocopherol, in 28 patients with Parkinson's disease and 85 matched controls. All of them were recruited from a population study. RESULTS: None of the values studied differed significantly between the two study groups, and none of them were correlated with age at onset, duration of the disease, scores of the Unified Parkinson Disease Rating Scale or the Hoehn and Yahr staging in the Parkinson's disease group. CONCLUSIONS: These results confirm the previous findings of classic case-control studies, suggesting the absence of relationship of the studied values with the risk for Parkinson's disease. PMID- 10528303 TI - [The Neuropsychiatric Inventory. Psychometric properties of its adaptation into Spanish]. AB - OBJECTIVE: The purpose of this study was to examine the reliability and validity of the Spanish (Spain) version of the Neuropsychiatric Inventory. PATIENTS AND METHODS: The Neuropsychiatric Inventory was administered to the caregivers of 63 subjects from the Dementia Unit, Hospital Santa Caterina, Girona. All patients had detailed neuropsychological assessment, and their non-cognitive symptoms were also examined with the CAMDEX. RESULTS: There was a high level of internal consistency reliability. The Neuropsychiatric Inventory subscores correlated with those of the CAMDEX, indicating an acceptable level of validity. The most frequent symptom was apathy (56%), followed by irritability (38%), depression/dysphoria (35%), aberrant motor behavior (30%), agitation/aggression (29%), anxiety (27%), disinhibition (24%), delusions (19%), hallucinations (14%) and euphoria (3%). CONCLUSIONS: This study showed that the Spanish version of the Neuropsychiatric Inventory is a reliable instrument, which can briefly assess non cognitive symptoms in demented patients. The Neuropsychiatric Inventory is a useful instrument for research and clinical practice in different culture around the world. PMID- 10528304 TI - [Subclinical attention changes in transient ischemic attacks in the vertebrobasilar region]. AB - INTRODUCTION: A significant number of patients who have had cerebrovascular illness apparently recover their former abilities completely but return to normal life with subtle cognitive deficits which may affect their daily lives. Such is the situation of patients with transitory ischemic accidents who present with sustained, undiagnosed attention deficits. OBJECTIVES: To identify subclinical alterations due to attention deficits in patients with transitory ischemic accidents, and to contribute to the study of the physiopathological mechanisms involved in the integration of this function. PATIENTS AND METHODS: We examined 44 persons, divided into three groups for this study: one group had vertebro basilar transitory ischemic accidents, a second group had supratentorial infarct and a third was healthy. All were given a specially designed computerized test of continuous work to evaluate the sustained attention component. RESULTS: Significant differences were found between the transitory ischemic accidents and healthy groups, regarding the variables including correct answers, omissions and indications of attention. This was not seen with the variables involving reaction time and number of errors. This demonstrated the existence of attention disorders involving omission in the group of patients with transitory ischemic accidents. CONCLUSION: These findings suggest the hypothesis that in the vertebro-basilar region there are important mechanisms involved in the process of sustained attention. PMID- 10528305 TI - [Incidence of stroke in central Dalmatia during the war in the Republic of Croatia (1991-1995)]. AB - INTRODUCTION AND METHOD: A retrospective study was made of the incidence of stroke in the Dalmatian region (Split) during the war in the Republic of Croatia (1991-1995), to determine whether the war had affected the incidence of stroke in the population at risk. The data obtained was compared with data regarding the incidence of strokes in the pre-war period (1986-1990). RESULTS AND CONCLUSIONS: There was a small but significant increase in the incidence of strokes during the war period. There was also a considerable increase in patients with primary intracerebral hemorrhage during this period. The possible causes of this increase in patients with primary intracerebral hemorrhage are discussed. PMID- 10528306 TI - [Cognition, emotion, and behavior. Neuropsychosomatisms and non-neurological paroxysms]. AB - INTRODUCTION: We present neuropsychosomatic disorders diagnosed and treated during the period 1994-1997. PATIENTS AND METHODS: A total of 83 cases, 24 boys and 59 girls, were selected according to suspected diagnosis. Their ages were in relationship with the psychosomatic disorder. This 83 cases is 10% of neuropediatric assistance in the period. A protocol was designed for disclosing any organic pathology. The psychopedagogic method is based on the PASS theory of intelligence and emotion processing theory of masquerade behavior. The success was defined after a period of, at least, two years of follow-up. RESULTS: Cephalalgia was the most frequent diagnosis. Language and learning difficulties, attention deficit disorder and pseudo-epilepsy were also frequent. Other diagnoses were: amblyopia, paralysis, pseudo-autism, tic, sphincter disorder, vertigo, mutism and sleep disorder. DISCUSSION: Concerning differential diagnosis, it must be emphasized that complex partial epilepsy of frontal lobe can mimic psychosomatics disorder as short, less than one minute, automatism. Partial epilepsy of temporal lobe may also mimic psychosomatic disorder but epilepsy does not respond to psychopedagogic remediation. Tumor and migraine must be also disclosed in case of cephalalgia but they do not respond to psychopedagogic remediation. Neurological scientific bases of emotion processing theory are widely explained. CONCLUSION: Cognition and emotion are functions of the central nervous system so they are competency of neuropediatrician. On the other hand, it is convenient for neuropediatrician to know about behavioral analysis in order to improve diagnosis and economical cost. PMID- 10528307 TI - [Bilateral cyclic sciatica caused by endometriosis. Apropos of a case]. AB - INTRODUCTION: Cyclical sciatica due to implantation of endometrial tissue in the sciatic nerve in the region of the sciatic notch is a very unusual cause of sciatica. It occurs in women of childbearing age, as episodes of pain in the distribution of the sciatic nerve, which present in a cyclic manner and coincide with menstruation. If it is not treated, a sensomotor mononeuropathy of the sciatic nerve develops. CLINICAL CASE: The patient had complained of right-sided sciatic pain from the age of 36 years. Over the years a motor deficit had slowly and progressively appeared causing foot drop. The painful crises were related to her menstrual periods. At the age of 44 years a pyramidal muscle syndrome was diagnosed and treated surgically. This was followed by increase in the crises of sciatic pain. A year later, she started to have sciatic pain on the left side, which was similar to that of the right side. The clinical, imaging and electrophysiological findings are reported. The patient improved. She is still being treated with depot medroxyprogesterone and her pain has disappeared. CONCLUSIONS: Cyclical sciatica due to endometriosis is little known and may lead to permanent disability. Computerized axial tomography of the pelvis using contrast material is very useful for diagnosis. The use of depot medroxyprogesterone seems to be a satisfactory treatment in some patients. PMID- 10528308 TI - [Severe polyneuropathy after using nitrous oxide as an anesthetic. A preventable disease?]. AB - INTRODUCTION: Nitrous oxide is a commonly used anaesthetic agent. One complication of this is due to its capacity to inactivate cobalamin. Therefore, in patients with poor reserves of vitamin B12, neurological and hematological alterations may be induced after a short period of exposure to nitrous oxide. CLINICAL CASE: A 69 year old man was anesthetized for three hours with 50% nitrous oxide during a surgical operation. Two weeks later he complained of severe mixed, mainly sensory polyneuropathy and was unable to walk. On diagnostic studies, vitamin B12 levels were found to be 18 pg/ml. The Shilling test confirmed that there was lack of intrinsic factor. In the preoperative studies a striking increase in motor conduction velocity was observed. Neurophysiological studies showed that there was mixed polyneuropathy, predominantly axonal. After starting treatment with hydroxycobalamin there was marked improvement and the patient became able to walk unaided. CONCLUSION: Since nitrous oxide may cause serious neurological alterations in patients with subclinical deficits of cobalamin, which may not always be accompanied by hematological changes, we consider the need for determination of plasma levels of vitamin B12 and if possible, of methylmalonic acid and homocysteine in elderly patients who are to have general anesthetics involving nitrous oxide. PMID- 10528309 TI - [Reflex benign myoclonic epilepsy of childhood. Apropos of a new case]. AB - INTRODUCTION: Recently a series of cases has been reported characterized by myoclonic crises similar to those occurring in benign myoclonic epilepsy of childhood. However, these crises only occurred after unexpected tactile or auditory stimuli. These clinical conditions represent a new epileptic syndrome, which is age-dependent and has been called benign myoclonic epilepsy of childhood. CLINICAL CASE: We present the case of a 12 month old girl with myoclonic crises which occurred only after auditory or tactile stimuli. The myoclonia could be set off whilst awake or asleep. No other types of crises or neurological changes were seen. A brother of the patient had had febrile convulsions. The EEG recorded during the crises showed generalized brief spike and-wave discharges at 3 cycles/second. The intercritical EEG was normal whilst awake, but during sleep showed brief generalized discharges. After treatment with valproate was started the crises became less frequent. CONCLUSIONS: The case we describe is similar to those described by Ricci et al in 1995. We, therefore, consider it to fit the concept of reflex myoclonic epilepsy of childhood of benign character. We consider that this condition should be differentiated from other reflex epilepsies and epileptic syndromes with a predominance of myoclonia, including benign myoclonic epilepsy of childhood. PMID- 10528310 TI - [Neurological clinical findings as the initial manifestation of a peripheral lymphoma]. AB - INTRODUCTION: Lymphomas of the central nervous system include primary and secondary lymphomas of Hodgkin and non-Hodgkin types. For a long time central nervous system involvement during the course of systemic non-Hodgkin lymphoma has been considered unusual, to occur late on in the disease process and usually to be located in the meninges. CLINICAL CASE: We describe the case of a 27 year old man initially diagnosed as having a primary cerebral lymphoma after having had repeated convulsive crises. Two months later he was found to have a retroperitoneal mass. On anatomopathological study of the mass, a peripheral T lymphoma was confirmed. CONCLUSIONS: Dissemination of systemic lymphomas to the central nervous system is usually seen in persons with advanced systemic disease. It is atypical to find a peripheral T lymphoma with initial clinical findings that were neurological, and even less frequent that these neurological findings were due to an intraparenchymatous lesion. In this article we describe a patient with these characteristics, and conclude that it is necessary to study patients with a diagnosis of primary cerebral lymphoma very fully to establish where the primary focus is. PMID- 10528311 TI - [Severe form of juvenile type II glycogenosis in a compound-heterozygous boy (Tyr 292--> Cys/Arg-854-->Stop)]. AB - INTRODUCTION: Type II glycogenosis is a glycogen storage disease inherited as an autosomal recessive trait. This molecular and clinically heterogeneous condition is due to a deficiency in a lysosomal acid 1,4-alpha-glucosidase. OBJECTIVE: To report the clinical, enzymatic and molecular characterization of a mulatto child, born to healthy Dominican mother and Caucasian father, affected by the juvenile phenotype form of type II glycogenosis. CLINICAL CASE: This 16-month-old male presented from 10 months with motor delay, limb-girdle hypotonia, prominence of calves, increased CPK value, mixed myotonic/myopathic pattern on EMG, intense glycogen muscle storage (775 micrograms/mg of protein) and severe deficiency of 1,4-alpha-glucosidase activity (< 0.03 mU/mg of protein). At 24 months he developed a rapidly progressive hypotonia and respiratory failure. The patient was found to be compound heretozygote for the novel mutation Tyr-292-->Cys, coming from the father, and Arg-854-->Stop, coming from the mother and previously reported in two Afroamerican patients, one with the adult phenotype and the other with the juvenile one. CONCLUSIONS: The present case corresponds to a severe form of type II juvenile glycogenosis. The severity of this condition may be related to the observed mutations and the associated extremely low muscle enzymatic activity. PMID- 10528312 TI - [Juvenile amyotrophic lateral sclerosis. Apropos of a case]. AB - INTRODUCTION: There are scanty reports on juvenile forms of amyotrophic lateral sclerosis, specially amyotrophic lateral sclerosis and deafness, and it is known as Madras pattern of motor neurone disease. CLINICAL CASE: We describe an sporadic case of juvenile amyotrophic lateral sclerosis with deafness in a young person who started with hearing loss at 21 years old, loss of strength in upper limbs and muscular atrophy. He was seen by a neurologist when he was 25 years old, there were evident generalized fasciculation activity in proximal and distal muscles in the four limbs and the tongue, with swallowing troubles, and increased tendon reflexes in lower limbs with abnormal plantar extensor responses. All the paraclinical test were normal, except the electromyogram, showing a classical pattern of lower motor neuron disease, and the auditory brain stem response with absence of the main components of this evoked response, as expression of VIII cranial nerve damage. DISCUSSION: Patients like this one were first described in Madras (India), and the evolution of this kind of juvenile form of amyotrophic lateral sclerosis is chronically progressive and relative benign, in relation to the classical form of amyotrophic lateral sclerosis and other forms of motor neurone disease which begin in childhood, adolescence or young adulthood. CONCLUSION: Its recognition is very important in order to diminish misleading therapies in these patients. PMID- 10528313 TI - [Diagnostic strategy for mitochondrial diseases]. AB - OBJECTIVE: The variability of both phenotypic and genotypic expression in mitochondrial diseases makes clinical diagnosis difficult, which is essential to establish therapy, aid in genetic counselling or for performing prenatal diagnosis. We have therefore proposed a strategy to help determine correct diagnosis of these alterations, in an attempt to rationalize the number of tests and, whenever possible, avoid tissue biopsy and minimize the size of the biopsy when indicated. DEVELOPMENT: Based on mitochondrial metabolism and molecular bases, as well as their alterations, a preliminary metabolic examination is carried out including at least one study of cytoplasmatic (lactate/pyruvate) and mitochondrial oxide reduction (hydroxibutirate/acetoacetate) in basal conditions or, if required, following glucose overload or an effort test. Metabolic study, in addition to clinical exploration, are the screening tests used to determine the need for tissue biopsy in which biochemical (pyruvate dehydrogenase, free and total carnitine, beta oxidation enzymes and respiratory chain complexes), genetic (mitochondrial DNA or nuclear alterations) and histologic tests are carried out to confirm diagnosis. CONCLUSIONS: a) Metabolic exploration may discard mitochondrial disease and many cases, avoid the use of an invasive procedure such as tissue biopsy. b) Biochemical study of tissue biopsy is the only useful key in the confirming of the diagnosis when no mitochondrial and/or nuclear DNA are observed. PMID- 10528315 TI - [Cognitive changes in normal aging: nosology and current status]. AB - INTRODUCTION AND OBJECTIVES: During the last 15 years several diagnostic categories have appeared to describe a group of adults with cognitive impairment compared to their age-matched standardized norms but without dementia. In this work, the main studies relating to these categories are reviewed and compared in order to establish if they define similar or different aged populations. DEVELOPMENT: Differences in prevalence or in prognostic values among studies are probably due to the selection of diagnostic categories or the differences in the application of inclusion/exclusion criteria. Genetic and neuroimaging data have contributed to reinforce the validity of the proposed classifications to identify the age related cognitive decline. CONCLUSIONS: The criteria used seems to be very important in the inclusion of subjects closer to normal aging or to dementia. In this respect further longitudinal studies and a consensus from previous described categories are need to reliably identify aged population with lower cognitive function compared to their age norms but different from patients in the initial stages of dementia. PMID- 10528314 TI - [Ubiquinone: metabolism and functions. Ubiquinone deficiency and its implication in mitochondrial encephalopathies. Treatment with ubiquinone]. AB - OBJECTIVE: Review of ubiquinone-10 metabolism and functions in humans, focusing its implication in the pathogenesis and physiopathology of mitochondrial encephalomyopathies. DEVELOPMENT: Ubiquinone-10 is an endogenously synthesized lipid with a wide distribution in tissues. Tyrosine and acetil-CoA are involved in ubiquinone biosynthesis. This molecule has several biological functions in cells: it is a movil electron carrier in the mitochondrial respiratory chain and also acts as antioxidant. Owing to its implication in these functions, ubiquinone deficiency may cause important deletereous effects in tissues. Several authors reported ubiquinone deficient status in some physiological and pathological conditions. Mitochondrial encephalomyopathies may be related to a primary or secondary ubiquinone deficient status, or even to an altered function of ubiquinone in the respiratory chain. Moreover, some relevant aspects about ubiquinone therapy in mitochondrial disorders are reported. CONCLUSIONS: According to recent reports about ubiquinone implication in several diseases, its determination in different biological samples seems very useful to elucidate the physiopathological mechanisms involved and even the to start a therapy in cases with ubiquinone deficiency. PMID- 10528316 TI - [First Virtual Ibero-American Congress in Neurology. A critical analysis]. PMID- 10528317 TI - [The influence of oleic acid on the aminopeptidase activity in astrocytes of the rat]. AB - INTRODUCTION: Changes in fatty acid composition of membrane lipids induce modifications on the activity of several enzymes and membrane transporters. Glial cells possess aminopeptidases which are located in the plasma membranes. Aminopeptidases are generally zinc-metalloenzymes which hydrolyze peptide bonds near the N-terminal end of peptides and polypeptides. The importance of these enzymes is based on their major role in protein metabolism and in the regulation of circulating hormones and biologically active peptides. OBJECTIVE: We study the effects of oleic acid on several aminopeptidase activities in primary cultures of rat astroglia, using aminoacyl-beta-naphthylamides as substrates. RESULTS: Oleic acid inhibits Ala-, Cys-, Leu- and Tyr-aminopeptidase activities, but not modifies Arg- and pGlu-aminopeptidase activities. CONCLUSIONS: Oleic acid modulates aminopeptidase activities in astrocytes. This could be related with intercellular communication and molecular transport processes, in which the astrocyte function has been involved. Furthermore, oleic acid might modulate the action of opioid peptides and steroid hormones on astroglial cells. PMID- 10528318 TI - [Progressive myoclonic epilepsy: clinical characteristics of 18 patients]. AB - INTRODUCTION: The term progressive myoclonic epilepsy (PME) includes a groups of heterogeneous conditions, with genetic causes, characterized by having different types of seizures, basically myoclonic, and other neurological findings due to a progressive lesion of the central nervous system. OBJECTIVE: To demonstrate the aetiology and clinico-encephalographic changes seen in patients with PME. PATIENTS AND METHODS: A retrospective, descriptive study was done of patients attended for PME in the Instituto de Neurologia y Neurocirugia de Cuba between 1990 and 1995. Eighteen patients were included. All were interviewed and had a physical examination, EEG and the specific complementary tests for each aetiology. RESULTS: There was a predominance of neural ceroid lipofuschinosis in 10 patients (55.5%), and in 9 of these the illness started before the age of 9 years. The second most frequent condition was myoclonic epilepsy with red-torn fibres (16.6%) and Unverricht-Lundborg disease (16.6%). The latter began in late childhood or adolescence. The most marked clinical characteristics were epilepsy, which was difficult to control and intellectual deterioration in 100%, followed by cerebellar signs in 88.8%. Myoclonias were the commonest type of seizures (94.4%) and many children presented with prior tonic-clonic seizures (88.8%). CONCLUSION: Response to treatment was poor but the best results were obtained using valproate either alone or associated with benzodiazepines. PMID- 10528319 TI - [A study of Gobbi's syndrome in Spanish population]. AB - INTRODUCTION: The association of epilepsy, occipital cerebral calcifications and coeliac disease was recognized for the first time in 1992 by Gobbi as an independent syndrome. He emphasized that it occurred almost exclusively in persons of Italian origin. OBJECTIVE: To define the prevalence of this syndrome in the Spanish population with a view to confirming its probably genetic etiopathogenesis. PATIENTS AND METHODS: Neurological studies were done in 44 coeliac patients, as were cranial CT scans. All cases of Spanish origin described in the literature were noted. RESULTS: In the patients with coeliac disease there was an increased incidence of crises and generalized epilepsy, but no patient was found to have occipital calcifications. Only 12 cases of Spanish origin were found, mainly in the Canary Isles and Mediterranean littoral, in spite of the high incidence of coeliac disease in Spain. No familial cases were seen. CONCLUSIONS: We suspect a genetic etiopathogenesis, associated with different environmental factors, to be the origin of the syndrome. This is supported by the common geographical origin of most cases and the anatomopathological findings described in those cases studied. PMID- 10528320 TI - [Carotid stenosis and lacunar infarct]. AB - INTRODUCTION: Perforating artery disease is still the main cause of lacunar infarcts (LI), while the relationship between this type of ischemia and carotid stenosis (CS) is controversial. OBJECTIVE: To assess the association between CS and LI in patients with criteria of localized ischemia in the carotid territory separately analyzing isolated LI and multiples LIs. PATIENTS AND METHODS: Three hundred and thirty patients with a first episode of cerebral infarct in the area supplied by the carotid artery were registered prospectively. There were 205 LI (135 isolated and 70 multiple) which could be differentiated from 125 non-lacunar infarcts (NLI). The vascular risk factors were determined and the degree of carotid stenosis measured by duplex-colour ultrasound exploration. RESULTS: In the isolated LI group the frequency of presentation of CS greater than 50% was 22% for the artery ipsilateral to the LI and 8% for the contralateral artery. For significant CS (> 70%) frequencies of 14% (ipsilateral) and 8% (contralateral) were observed. On comparative analysis with the NLI group, the presence of stenosis greater than 50 to 70% was significantly higher in these for both ipsilateral and contralateral arteries. In the LI group, isolated CS was significantly more frequent on the side which was homolateral to the ischemia as compared to its isolated presence on the contralateral side (22/30 vs 4/12; OR: 5.5 [95% CI: 1.2-23]). Ipsilateral CS greater than 70% behaved as a factor which was significantly associated with the multiple LI subtype with a unilateral pattern. CONCLUSION: CS should be considered a risk factor for LI. PMID- 10528321 TI - [Haptoglobin in cerebrospinal fluid as a marker of infectious process in central nervous system]. AB - INTRODUCTION: Haptoglobin is a transport protein and protects organism against iron loss and it should be involved in central nervous system infectious process. PATIENTS AND METHODS: Simultaneous serum and cerebrospinal fluid were obtained of 39 pediatric patients, 14 suffering from viral meningoencephalitis and 25 from bacterial meningoencephalitis. Five control cases were examined too. Haptoglobin, IgG and albumin were quantified in both fluids by radial immunodiffusion. Haptoglobin cerebrospinal fluid/serum ratio, haptoglobin index and haptoglobin/IgG index were calculated. Local IgG intrathecal synthesis was determined by reibergram. RESULTS: Haptoglobin index was higher not statistically significant in viral meningoencephalitis in comparison with bacterial disease but both were statistically significant with respect to control group. Increased haptoglobin/IgG index were statistically significant in bacterial meningoencephalitis in relation with viral meningoencephalitis. There were no association between haptoglobin and polymorphonuclear cells count and globular sediment speed. CONCLUSION: Haptoglobin should be considered a relevant marker of central nervous system infectious process. PMID- 10528322 TI - ["Floppy infant" syndrome in twins secondary to the use of benzodiazepines during pregnancy]. AB - INTRODUCTION: Due to the pharmacological characteristics of benzodiazepines, they have been use in a wide variety of disorders, as well as during pregnancy and labor. Benzodiazepine intake during early pregnancy may be teratogenic, and their intake during late pregnancy may be associated with neonatal withdrawal syndrome and 'floppy infant' syndrome. CLINICAL CASE: A hypotonic syndrome in twins secondary to the use of diazepam in the last month of pregnancy is described. Their neurological improvement occur in the first two weeks of life, in relation with the normalization in diazepam blood levels. CONCLUSIONS: The list of possible diagnosis for floppines in newborn is extensive, and we must remember some drugs use during pregnancy. Diazepam show a rapid placental transfer with significant uptake of the drug; by this, high single doses and repeated and prolonged administration of benzodiazepines have to be avoided during pregnancy. PMID- 10528323 TI - [Late onset polyneuropathy due to exposure to organophosphates]. AB - INTRODUCTION: Organophosphate esters are widely used both in agriculture and in industry and may give rise to acute intoxication due to their direct anticholinesterase effect. Less often a polyneuropathic syndrome of late onset may occur. This is predominantly motor, symmetrical, with muscle atrophy and pyramidal signs. These neurotoxic features are due to inhibition of the target esterase of this neuropathy. Histopathological studies show axon degeneration of 'dying back' type. CLINICAL CASE: We present the case of a 24 year old man with severe sensomotor neuropathy, predominantly distal, with marked atrophy of the interosseus muscles, claw hand and foot drop, together with increased deep tendon reflexes. This patient had worked on the land, cultivating maize, from the age of 14, when his symptoms began and became progressively worse. On electromyography there was sensomotor peripheral polyneuropathy, which was primarily axonal and predominantly motor and distal. Peripheral nerve biopsy confirmed the presence of 'dying back' type axonopathy. Extensive biochemical investigations ruled out other causes of polyneuropathy. Cerebral resonance study was normal. CONCLUSIONS: Agricultural workers chronically exposed to organophosphate insecticides, without adequate protection, have an increased risk of developing late onset neuropathy due to organophosphates. The risk of developing late onset neuropathy is not necessarily related to a history of acute intoxication, since chronic sublethal exposure may mean that previous acute anticholinergic symptoms pass undetected. PMID- 10528324 TI - [Menkes disease: experience in copper salts therapy]. AB - INTRODUCTION: Menkes disease is an X linked genetic disorder of copper intracellular transport. The clinical effects are explainable by impaired activity of various copper-dependent enzymes. The classic form of the disease is characterized by: low temperature, impairment of growth, characteristic hair abnormalities (thin, breakable and grey-haired) and seizures with severe developmental retardation from birth. Treatment with various forms of copper salts has been used, but none of them effective. CLINICAL CASE: We report a patient diagnosed of Menkes disease at 12 months of age. Copper treatment was commenced at 14 months with 100 micrograms/kg daily intravenous infusion of copper sulphate. On the basis of copper and ceruloplasmin plasma levels, doses of treatment were controlled until a maintained dose of 1,000 micrograms intramuscular of copper sulphate every 17 days, until the age of 6 years. The effects of the treatment were marked by the normalization of the biochemical parameters, the growth development and the hair characteristics. However, no significant effect was shown on the seizures or the neurologic impairment. The patient died at the age of 7 years old. CONCLUSION: Parenteral therapy for Menkes disease with various inorganic copper salts has been shown to get longer clinical course, but no successful effect on the devastating neurodegenerative progression of the disease. PMID- 10528325 TI - [Multifocal motor neuropathy with block of nerve conduction. Report of three cases]. AB - INTRODUCTION: Multifocal motor neuropathy is a severe chronic demyelinating, disimmune, crippling and potentially treatable disease. The clinical features are characterized by a syndrome of progressive motor deficiency with muscle atrophies, fasciculations, areflexia, myokimias and cramps, that sometimes we misdiagnosed as motor neuron disease. Electrophysiologically it is characterized by the occurrence of multifocal conduction blocks of motor fibres. CLINICAL CASES: Three patients with asymmetric multifocal motor neuropathy are reported in whom diagnosis was made by nerve conduction studies. All the patients had developed an asymmetric chronic progressive motor deficit that involved mostly upper limbs. One patient exhibited mild tactile sensory disturbances and a sural nerve biopsy showed moderated involvement of myelinated fibers. All the patients showed electrophysiologically, multifocal conduction blocks of motor fibers with spared conduction through sensory fibers at the same segments where motor conduction was blocked. The response to high doses intravenous cyclophosphamide therapy followed by 100 mg orally for six months, was satisfactory in all three patients. There was no response to previous trials with prednisone, azathioprine, and intravenous immunoglobulin. One patient was also submitted to several sessions of plasma exchange without benefit. The patient had been previously diagnosed as having motor neuron disease. CONCLUSION: The importance of early diagnosis of this disorder and the possibility of successful treatment with high doses of intravenous cyclophosphamide is stressed. PMID- 10528327 TI - [8th course of Spanish of the American Academy of Neurology]. PMID- 10528326 TI - [Progressive multifocal leukoencephalopathy in elderly immunocompetent patients. Report of 2 cases]. AB - INTRODUCTION: Progressive multifocal leukoencephalopathy is a disorder which is rare in immunocompetent patients. OBJECTIVES: We report the cases of two elderly patients with serology, in one case positive for hepatitis C, and in the other with anti-DNA antibodies, and discuss the part these might play in causing progressive multifocal leukoencephalopathy. CLINICAL CASES: Case 1. An 86 year old man had been found on serology investigations to be positive for hepatitis C virus. In November 1996 he complained of dysarthria and left hemi-negligence following an accidental fall. Since his clinical condition became worse he was admitted to hospital for further investigation. On neuroimaging studies the intracerebral lesions were increased. The only other finding confirmed was that of positive serology for hepatitis C virus. The patient deteriorated progressively and died 50 days after admission. Case 2. A 70 year old woman began to show progressive cognitive impairment and left hemiparesia in June 1996. Se was investigated in another centre and provisionally diagnosed as having vasculitis of the CNS, in view of her positive anti-DNA antibody and right frontoparietal hypodense lesion. Treatment had been started with corticosteroids. She was admitted to our hospital when her neurological deficits worsened. The immunological alterations were confirmed. On MRI the lesions in the white matter were seen to have progressed. The patient slowly improved. She was discharged from hospital in February 1997 in a semiconscious state, able to follow persons and things with her eyes, with global aphasia and with spastic tetraparesia which was mainly left-sided. She remains stable. CONCLUSION: Progressive multifocal leukoencephalopathy is a condition which should be remembered when dealing with immunocompetent patients. PMID- 10528328 TI - [Recent advances in Parkinson disease]. AB - OBJECTIVE: To evaluate the latest advances in the diagnosis and symptomatic pharmacological treatment of patients with Parkinson's disease at different stages. DEVELOPMENT: In this article we summarize the criteria for clinical diagnosis of Parkinson's disease based on recent clinico-pathological studies. We evaluate the different therapeutic options at early and late stages of the illness in view of the results of recent, controlled, double-blind studies in which the efficacy of new dopaminergic agonists (carbergolin, pramipexol and ropinirol) and catechol-O-methyl-transferase enzyme inhibitors (COMT; entacapone and tolcapone) are compared with placebo and with established treatments (e.g. levodopa and bromocryptin). CONCLUSIONS: Levodopa is still the most important advance yet made in the treatment of Parkinson's disease, but new dopaminergic agonists and COMT inhibitors are useful additions to the possibilities available for the treatment of early and late stages of the illness and may be included in modern guidelines for treatment. PMID- 10528329 TI - [Advances in the surgical treatment of Parkinson disease and other movement disorders]. AB - OBJECTIVE: To present the latest advances in surgery for patients with Parkinson's disease and other movement disorders, emphasizing pallidotomy and deep cerebral stimulation (DCS), thalamic, pallidal and subthalamic, their indications and complications. DEVELOPMENT: Advances in neurophysiology and radiology have led to greater understanding of the physiology of the basal ganglia. This has led to the revival of old surgical techniques (pallidotomy) and the clinical application of new variants on these techniques. Pallidotomy is indicated in patients with advanced Parkinson's disease who are under the age of 65 years and have no psychiatric complications or drug-induced debilitating dyscinesias. In most of these patients the daily dose of levodopa is reduced by approximately 30%. Any complications which occur are transient. The versatility of adjustment of the parameters of deep cerebral stimulation allow them to be adjusted to the patient's needs. Complications include transitory paraesthesia of the side contralateral to the stimulation, transitory tonic contraction of the face and limbs and in some cases dyskinesias which are similar to those induced by drugs. In the United States of America, the Food and Drugs Administration has authorized the use of thalamic DCS for treatment of essential/familial or Parkinsonian tremor of the hands. It is expected that by the end of this year pallidal-DCS will have been approved for the treatment of rigidity, tremor and bradykinesia in Parkinson's disease. CONCLUSION: Pallidotomy and DCS seem to be safe, useful techniques for the treatment of late complications of Parkinson's disease and possibly other movement disorders also. PMID- 10528331 TI - [Indications and management of botulinum toxin]. AB - INTRODUCTION: Botulinus toxin (BTX) is the most potent biological toxin yet known. It is produced by Clostridium botulinium, a Gram positive bacteria. DEVELOPMENT: Type A Botulinus toxin is the most widely used in human drug trials. It has become the treatment of choice for blepharospasm, hemifacial spasm, cervical dystonia and laryngeal dystonia. It may also be used in the treatment of patients with oromandibular dystonia and limb dystonia, especially writer's cramp, and has been used successfully in the treatment of spasticity and cerebral paralysis. There are many benefits from this treatment, including improved walking, improved posture of wheelchair patients, improvement of patients with spasms and easier extension of their arms and knees. The toxin also alleviates pain and may be used in therapeutic trials for prediction of the response to surgical elongation. PMID- 10528330 TI - [Cognitive and behavioral disorders in Parkinson disease]. AB - INTRODUCTION: Most patients with Parkinson's disease have cognitive and behavior disorders during the course of their illness. OBJECTIVE: In this study we consider practical aspects of the treatment of patients with Parkinson's disease who have such problems. DEVELOPMENT: Better understanding of both normal and abnormal physiology of the basal ganglia and their connections makes it possible to suggest hypotheses regarding the cause of these disorders. The combination of lesions of multiple subcorticocortical systems with different degrees of direct cortical pathology due to Lewy bodies and neuritic plaques may explain most of the changes, from depression to dementia. CONCLUSIONS: It is necessary to test these concepts using studies based on specific predictions derived from pathophysiological models. Until this is done, the clinical treatment of cognitive and compartmental disorders in Parkinson's disease will continue to be symptomatic, complex and controversial. PMID- 10528332 TI - [Myasthenia gravis: diagnosis and treatment]. AB - INTRODUCTION: Myasthenia gravis is an autoimmune disorder characterized by fluctuating muscle weakness and fatigue of different muscle groups. Myasthenia gravis may affect persons of all ages, but especially women aged 20 to 40 years. DEVELOPMENT AND CONCLUSIONS: The ocular, facial and bulbar muscles are most often involved in this disease. The muscle weakness of patients with myasthenia gravis becomes worse with intercurrent episodes of infection, fever and physical or emotional exhaustion. Respiratory infection (bacterial or viral) is the most frequent trigger factor. The presence of antibodies to acetylcholine receptors in a patient with the clinical features of myasthenia gravis, confirms the diagnosis. Treatment is controversial. Each patient therefore has to be treated individually, as no single treatment is suitable for all patients. Treatment may include anticholinesterase drugs, corticosteroids, plasmapheresis, immunoglobulin, immunosuppressive drugs and thymectomy. PMID- 10528333 TI - [Transcranial magnetic stimulation]. AB - INTRODUCTION: Transcranial magnetic stimulation (TMS) permits stimulation of the cerebral cortex in humans without requiring open access to the brain and is one of the newest tools available in neuroscience. There are two main types of application: single-pulse TMS and repetitive TMS. DEVELOPMENT: The magnetic stimulator is composed of a series of capacitors that store the voltage necessary to generate a stimulus of the sufficient intensity of generate an electric field in the stimulation coil. The safety of TMS is supported by the considerable experience derived from studies involving electrical stimulation of the cortex in animals and humans, and also specific studies on the safety of TMS in humans. CONCLUSIONS: In this article we review historical and technical aspects of TMS, describe its adverse effects and how to avoid them, summarize the applications of TMS in the investigation of different cerebral functions, and discuss the possibility of using TMS for the treatment of neuropsychiatric disorders. PMID- 10528334 TI - [Treatment of acute polyneuropathies]. AB - OBJECTIVE: To describe the treatment of a group of polyneuropathies of different aetiologies, characterized by acute onset. DEVELOPMENT: We review the treatment of the Guillain-Barre syndrome, whose mortality has been significantly reduced by the use of intensive care units. Plasmapheresis and immunoglobulin have been shown to be effective and speed clinical recovery of these patients. The use of corticosteroids is controversial, since clinical studies have not shown them to be effective in treating this disease. In this article we also describe the treatment of other neuropathies such as neuropathies secondary to vasculitis, vasculitis limited to the peripheral nervous system and vasculitis associated with infections such as HIV and hepatitis B. PMID- 10528335 TI - [Chronic inflammatory neuropathies]. AB - INTRODUCTION: The group of chronic inflammatory neuropathies is made up of a variety of neuropathies of autoimmune or infectious origin. DEVELOPMENT: We describe three types of neuropathies and their treatment: demyelinating chronic neuropathy, motor multifocal neuropathy and demyelinating neuropathy associated with monoclonal gammapathy. These three types of neuropathy have uniform characteristics and features, which permit precise identification and suggest that they are separate entities. CONCLUSIONS: In the management of any type of neuropathy it is essential to make the correct diagnosis. Initial investigations should include detailed electrophysiological examination to detect the presence of demyelination, lumbar puncture and, if pleocytosis is found, the possibility of associated acquired immunodeficiency syndrome should be investigated. Also, before immunosuppressive treatment is begun, a systematic medical examination should be done to detect any other coexisting illness which might affect use of the different drugs involved. PMID- 10528336 TI - [Plasmapheresis, immunotherapy and chemotherapy in polyneuropathies]. AB - INTRODUCTION: Although the precise cause of many of the polyneuropathies evaluated clinically is unknown, there is substantial evidence that many are caused by autoimmune disorders. Currently treatment is directed towards production of immunomodulation or immunosuppression. DEVELOPMENT AND CONCLUSIONS: The degree of subjective symptoms such as fatigue, pain or other sensory symptoms are not good criteria for determination of the need for immunosuppressive treatment. Treatment should be orientated towards serious neuropathic defects, not minimal deficits. It is therefore important to record the defect precisely, so as to be able to determine objectively the effect of therapy. For this, tables or diagrams using established scales, such as that of the Medical Research Council, are recommended. Similarly, evaluation and follow-up by the physical therapy and occupational therapy departments should be considered when deciding the treatment to be used for autoimmune neuropathies such as corticosteroids, plasmapheresis, ev immunoglobulin, cytopathic agents and alpha interferon. PMID- 10528337 TI - [Important aspects of the diagnosis and management of peripheral neuropathies]. AB - INTRODUCTION: The peripheral neuropathies represent a diverse group of disorders of the peripheral nervous system of very varied aetiology. It is therefore important to classify them, so as to reduce the number of possible diagnoses and to identify the correct cause. OBJECTIVE: To review the most important principles of the initial examination and the management of peripheral neuropathies. DEVELOPMENT: The clinical features of these diseases may be debilitating and incapacitating, and may be part of a symptom complex of very serious diseases. It is necessary to carry out systematic evaluation of the symptoms, signs and laboratory tests so as to establish the correct diagnosis and begin treatment. Some of the most important elements used in the process of classification are: functional findings, topography or the anatomical pattern of the deficits, and the physiological or pathological findings. Treatment of the peripheral neuropathies may be divided into four groups: elimination of the casual agent, treatment of symptoms, modulation of the immune system and rehabilitation. CONCLUSION: The possibilities of diagnosis and treatment continue to increase as the mechanisms responsible become clearer. PMID- 10528339 TI - [Fronto-temporal dementia: findings on transcranial Doppler]. PMID- 10528338 TI - [Metastatic neurological complications in patients with cancer]. AB - INTRODUCTION: The neurological complications of patients with cancer may be classified as metatatic or non-metastatic. DEVELOPMENT: The development of cerebral metastases (CM) is not only one of the complications most feared by patients, but also often leads doctors to withdraw the systemic treatment of cancer, although early diagnosis and treatment usually alleviates or resolves the neurological deficits, and improves the quality of life and prolongs it. Most CM are originated by embolization of neoplastic cells to the brain. They are therefore found in the region where grey and white matter meet. The commonest symptoms of CM are headache, convulsive crises, altered mental functions, motor deficits, ataxia and speech disorders. Cerebral MR and CT are the most useful investigations in CM. The tumours most frequently involved in metastatic spinal compression are cancer of the lung, prostate, breast and kidney, and non-Hodgkin lymphoma. MR is the investigation of choice, but if this cannot be done, or is contraindicated, myelography or medullary CT using subarachnoid contrast material should be used. CONCLUSION: In this article we review the diagnosis and management of metastatic neurological complications, specifically cerebral metastases and spinal compression. PMID- 10528340 TI - [Physiology and pathology of cerebral ischemia]. AB - In this article the pathophysiology of cerebral ischemia is reviewed. The concept of penumbra is explained, and a distinction is made between ischemic necrosis, which occurs in the severely ischemic regions and is associated with loss of calcium and glutamate homeostasis, and programmed cell death, or apoptosis, which is more likely to occur in the moderately ischemic regions, evolves more slowly and depends on the activation of a sequence of genes. Despite this knowledge, it is pointed out that currently only a small percentage of stroke patients can access therapies. Some thoughts are provided on future research directions and therapies. PMID- 10528341 TI - [The pathophysiology of acute cerebral ischemia: clinical approach using physiologic imaging]. AB - Physiologic imaging has recently allowed major new insights into the pathophysiology of acute ischemic stroke in man. The main observation concerns the major heterogeneity in pathophysiological patterns, with three situations being encountered: 1) cortical ischemic penumbra persisting up till 16 hrs post onset, which would represent an ideal therapeutic target; 2) early spontaneous reperfusion, predictive of favourable clinical and tissular outcome, and a priori requiring no specific therapy; and 3) early extensive necrosis, predictive of malignant hemispheric infarction. Physiologic imaging should soon become a fundamental tool in the evaluation of the individual pathophysiologic situation before a therapeutic decision about entry into trials is made. PMID- 10528342 TI - Neuroprotective therapy. AB - Neuroprotective therapy for stroke remains unproven despite its ability to substantially reduce injury in animal stroke models. Based on an understanding of the cascade of biochemical events that follow interruption of blood flow to the brain, various neuroprotective drugs have been tested in clinical studies, but none have been shown to improve clinical outcome. Progress depends on designing our clinical trials to better simulate the experimental conditions under which these drugs have been found to be effective. PMID- 10528343 TI - [Brain CT scan for acute cerebral infarction: early signs of ischemia]. AB - Computed tomography (CT) is widely used for early evaluation of acute strokes. Most importantly, CT excludes acute hemorrhage or other diseases mimicking ischemia. Therefore, CT is the main imaging examination in patients with brain ischemia and when antithrombotic agents are being considered. During the first hours after acute ischemic stroke, the CT does not usually show much in the first 24 hours. However, early abnormal findings on CT scan have been described such as the hyperdense middle cerebral artery sign (HMCAS), and reduced contrast attenuation of the cerebral parenchyma. HMCAS reflects arterial occlusion. Early parenchymal abnormalities, the attenuation of lentiform nucleus (ALN), loss of the insular ribbon (LIR) or hemispheric sulcus effacement (HSE) occur less frequently and they are positive criteria for cerebral in progress. Early parenchymal abnormalities might also predict subsequent infarct extension and hemorrhagic transformation. Therapeutic trials of ischemia in MCA territory involved decision making when the CT may not show obvious ischemic changes. Finally, initial CT findings may also help to predict response to therapy. PMID- 10528344 TI - [Anti-platelet and anticoagulant therapy in acute cerebral ischemia]. AB - Therapeutical trials in the acute phase of stroke have showed a moderate benefit of administration of aspirin in prevention of death or recurrent cerebral events. This benefit was obtained despite a small increase in systemic and cerebral haemorrhages. Heparin used at high dosage, without any control of coagulation test, induces an excess of cerebral and systemic haemorrhage which overset its benefit in prevention of recurrent cerebral events. Similar results have been observed with heparinoid and nadroparine used at high dosage. The only benefit of anticoagulation is the prevention of total and fatal pulmonary embolism which has been observed in all recent studies. The antithrombotic treatment which offers the best ratio benefit-risk in the acute phase of stroke is aspirin at a minimum dosage of 160 mg by day and, if risk factors are present, heparin at an adequate dosage to prevent venous thrombo-embolism. Explicative studies are required to explore the potential benefit of heparin in patients with a high risk of recurrent cerebral ischemic events. PMID- 10528345 TI - Update in thrombolytic therapy. AB - Several studies have proven the usefulness of thrombolytic agents in the therapy of ischemic stroke. Data from open and small placebo-controlled trials show a relationship between recanalization and an improved clinical outcome. Three large scaled studies have examine the effect of streptokinase treatment. Because of an increasing mortality the results of these studies were negative, and so streptokinase should not be used to treat acute ischemic stroke. In contrast, three rt-PA studies with together more than 2000 patients showed a significant benefit. Recent meta-analyses including data of all these trials show an odds ratio for death and disability of 0.67 (95 p. 100 CI 0.56; 0.8). The number needed to treat to prevent one death or disability is 11 in a 6 hour time window and 7 in a 3 hour time window. These are impressive numbers that are rarely found in other areas of internal or neurological medicine. The most important risk of thrombolytic therapy is the occurrence of intracerebral hemorrhage, which is reported as between 0-18 p. 100, but which is not associated with increased morbidity or mortality. At present, thrombolytic therapy cannot be recommended for all patients with acute stroke. Careful selection and experiences with this therapy and its risks are necessary. PMID- 10528346 TI - [Role of emergency cerebrovascular units in the network of care]. AB - Stroke units can improve outcome after stroke as demonstrated by a meta-analysis which evidenced a reduction of the odds ratio for mortality (18 +/- 8 p. 100) and for poor outcome, death, or institutional care (25 +/- 10 p. 100), and death or dependancy (29 +/- 11 p. 100). Improved quality of life and shorter hospital stay have not been so clearly established. Stroke units are helpful in organizing acute neurology care. Moreover, stroke units are involved in teaching and research. Their development should be promoted in hospitals. PMID- 10528347 TI - [Arterial hypertension and cerebrovascular accident: the need for improved prevention strategies]. AB - It is well demonstrated that the antihypertensive treatment is effective, particularly for the primary prevention of stroke. However, benefits of treatment are rather small in certain groups of patients. The explicit assessment of absolute cardiovascular risks and likely treatment benefits in patients with hypertension can usefully guide treatment decisions and provide a more rational basis for initiating therapy than blood pressure levels alone. This approach highlights the generally greater cardiovascular risk and potential treatment benefits in older compared with younger hypertensive patients. Some specific questions remain still unanswered. Evidence is accumulating concerning protective effect of antihypertensive treatment against dementia. Trials are in progress to investigate the effect of treatment on stroke incidence in hypertensive patients over the age of 80 years. Finally, despite the worldwide use of calcium antagonists and converting enzyme inhibitors, solid evidence of their safety and efficacy compared with the references drugs (beta-blockers and diuretics) is still lacking. PMID- 10528348 TI - [Cardiopathies associated with a low embolic risk]. AB - Advances in cardiac imaging techniques have resulted in the recognition of several potential cardioembolic sources (such as patent foramen ovale, atrial septal aneurysm, strands, mitral valve prolapse), but it is often not clear whether the findings, which often have a substantial prevalence in patients without stroke, are merely an associated abnormality or a direct cause of the stroke. Overall, these cardiopathies are associated with a low risk of stroke. Studies are needed to better define the risk and mechanisms of stroke associated with these cardiac abnormalities, to determine which patients are at increased risk of stroke and to assess the potential benefits from therapeutic interventions. Unless we can define strict indications for therapeutic interventions, we risk ending up with exposing some patients to unnecessary complications of treatment. PMID- 10528349 TI - [Vascularization of the cerebral cortex]. AB - Three levels are described for the study of cortical blood vessels: the main leptomeningeal arteries and veins, the fine pial network and the intracortical vessels. If the main leptomeningeal arteries and veins are well known, the functional anatomy of the fine cortical blood supply is still obscure. Pial arteries and veins form a dense superficial network. The anastomoses between arteries and between veins are numerous but no arteriovenous anastomoses were found. Numerous smooth vascular formations are suspected into the vascular walls and would regulate the pial (and intracortical) blood flow. The intracortical network is divided into arteries, veins and capillaries. Five types of arteries and veins are described according to their degree of intracortical penetration. The capillary network may be described into four layers according to their density. The lay-out of arteries and veins and their branching would allow to suspect three types of intracortical flows. Intracortical vascular units were also described each composed of a central vein surrounded by peripheral arteries. No arterial or venous anastomoses were found in the cortex whereas precapillary arteriovenous shunts would be suspected. Vascular deformities may be due to aging, are present in the cortex and in the subcortical region. PMID- 10528350 TI - [Prevention of cerebral ischemia: anti-platelet agents]. AB - Besides the optimal management of risk factors for stroke and carotid surgery, antiplatelet agents are the cornerstone for prevention of cerebral ischaemia. The aim of this overview is to determine their role in the prevention of cerebral ischaemia, from available literature. In primary prevention, the benefit of aspirin has been established only for patients with non-valvular atrial fibrillation and a low risk of cardioembolism, or as an alternative choice of warfarin, and in subjects at high risk of atherosclerosis. In secondary prevention, antiplatelet agents are effective to reduce the risk in patients with ischaemic stroke due to atherosclerosis: aspirin (50 to 1300 mg), ticlopidine (500 mg), clopidogrel (75 mg) and dipyridamole (400 mg) are effective, but the higher levels of risk reduction are obtained with clopidogrel, ticlopidine and the association aspirin--dipyridamole. Aspirin is recommended in most other causes of cerebral ischaemia, except in high risk cardiopathies when anticoagulation is possible. Other domains should still be explored: are antiplatelet agents also effective to reduce the risk of cerebral ischaemia in patients with other causes, especially lipohyalinosis of the deep perforators leading to lacunar infarcts? In daily practice, does prescription follow recommendations? Will it be possible to reproduce the results of the European Stroke Prevention Study (ESPS)2? Are antiplatelet agents other than aspirin effective in non-valvular atrial fibrillation? Are other associations of antiplatelet agents more effective than these agents alone? Finally, what will be the role of new antiplatelet agents in the future? PMID- 10528351 TI - Anticoagulant treatment in stroke prevention. AB - This review aims to summarise the value of long-term oral anticoagulant treatment in stroke prevention. Oral anticoagulation is the treatment of first choice in patients with atrial fibrillation (AF) and vascular risk factors and in AF patients with recent cerebral ischemia. The treatment also substantially reduces the risk of stroke in patients after myocardial infarction. The optimal target intensity of anticoagulation in stroke prevention is an International Normalized Ratio (INR) between 2.0 and 3.0. The treatment has been found to be hazardous at INR intensities between 3.0 and 4.5 in patients with transient ischemic attack (TIA) or minor stroke of presumed arterial origin. The value of the treatment in lower intensity in such patients still has to be established. PMID- 10528352 TI - Carotid surgery. AB - Carotid endarterectomy may well be the most studied operation ever. Unlike many if not most surgical interventions, there are several randomised trials, and now meta-analyses, to inform clinical practice. On balance, accepting the small but definite risk of stroke as a result, surgery-is a reasonable option for many patients with severe and recently symptomatic carotid stenosis who have such a high risk of stroke without surgery. Although operating on asymptomatic stenosis is safer, the unoperated risk of stroke is--on average--so low that surgery is generally not worthwhile. The challenge now is to make surgery even safer, and perhaps angioplasty will be safer still and as durable although so far there is no good randomised evidence. At the same time we must identify, if we can, those few patients with severe symptomatic stenosis who will have a stroke and focus surgery just on them, and not offer it to all the patients with severe symptomatic stenosis who might have a stroke. This will require the development and validation of mathematical models of risk. But, however well focused surgery becomes and so however small we can make "the numbers-needed-to-treat", it will have a negligible impact on stroke incidence because so few strokes are preceded by transient ischaemic attacks in patients with severe carotid stenosis. PMID- 10528353 TI - [Angioplasty of atherosclerotic carotid stenoses]. PMID- 10528354 TI - From therapeutic trials to current practice. AB - Ideally, there is continuous interaction between clinical reality and the methods of controlled clinical trials. The design of a proper trial ought to reflect the physician's and especially the patient's point of view, and conversely a familiarity with clinical trials improves the methods of daily practice. In other words, every methodological principle is rooted in clinical practice. Some examples are: randomisation (with constructive doubt as its practical equivalent), independence (a healthy mistrust of drug companies), informed consent (sharing uncertainty with patients), type I error (no false optimism after a single trial), type II error (no false pessimism after a single trial), choosing the right measure of outcome (relevance outweighs precision), the intention-to-treat principle (pragmatic analysis), and the dangers of subgroup analysis (the "my-last-patient syndrome"). PMID- 10528355 TI - Cortical plasticity after stroke: implications for rehabilitation. AB - While adaptive processes in the cerebral cortex have long been thought to contribute to functional recovery after stroke, the precise neuronal structures and mechanisms underlying these processes have been difficult to identify. Over the past 15 years, a large number of studies conducted in human stroke patients and in experimental animal models have contributed to a more coherent picture of the brain's adaptive capacity after injury. These studies suggest that the cerebral cortex undergoes significant and functional structural plasticity for at least several weeks to months following injury. Adaptive changes have been demonstrated in the intact tissue surrounding the lesion, as well as in other cortical motor areas remote from the site of injury. Recent results from non human primate studies of cortical reorganization after stroke demonstrate marked functional changes in the intact cortical tissue adjacent to the infarct in the weeks following an ischemic lesion. Further, intensive task-specific practice with the impaired limb has a modulatory effect on the inevitable cortical plasticity. Taken together with parallel studies of forced use in human stroke patients, it is likely that use of the impaired limb can influence adaptive reorganizational mechanisms in the intact cerebral cortex, and thus, promote functional recovery. PMID- 10528356 TI - [Mechanisms of motor recovery after cerebrovascular accident]. AB - Recovery from motor deficit after a stroke remains a puzzling scientific question as well as public health problem. The natural history of deficits after stroke is given to us through published series of patients and we know from them that neurological deficits, spontaneously but most of the time partially, recover. Neuroimaging modern techniques (PET scan, fMRI, evoked potentials) allowed us to identify the main aspects of the post-stroke intracerebral reorganisation. Reorganisation of basal cerebral metabolism, changes in the somatotopia of primary motor cortex, recruitment of remote cortices, participation of associative cortices are clearly part of the rearrangement processes. It is likely that such mechanisms represent the basis of clinical recovery of our patients. However, despite those important advances, very few is known about the effect of treatments on the recovery phenomenon. Some lines of evidence appear now to give rationale to rehabilitation procedures and to drugs suspected to improve clinical recovery. PMID- 10528357 TI - [Mechanisms of aphasia recovery and brain imaging]. AB - Aphasia recovery may depend on right hemisphere or non-lesioned left hemisphere structures, pre-morbid brain language organization, and de novo learning of language. Here we review the brain imaging evidence supporting these different hypotheses. CT-scan studies have investigated the prognosis value of size and site of left hemisphere lesions. The size of the lesion is a global but not an individual predictor of the initial severity and subsequent recovery of aphasia. Studies on the site of the lesion have given different results for verbal expression and comprehension. There is no consensus on a single critical site for recovery of verbal expression in non-fluent aphasia, which may depend on sub cortical more than cortical extend of the lesion. Conversely the extend of the lesion in the superior temporal gyrus emerges as a critical negative factor for comprehension recovery. Rest measurements of brain metabolism have consistently shown that aphasia severity depends much more on the degree of dysfunction of language-related areas in the left hemisphere than on the site of the lesion it self. This suggests that aphasia recovery may depend on metabolic dysfunction recovery in peri-lesional structures. More recently, activation studies have shown consistent right hemisphere activation during language tasks in aphasic subjects, but their role in recovery remains debated. It is likely limited, and may depend on atypical pre-morbid language lateralization. Left hemisphere activations are also found in aphasic patients. They are often relocalized in peri-lesional areas, and emerge in most studies as the main factor of aphasia recovery. PMID- 10528358 TI - Pharmacological approach to functional reorganization: the role of norepinephrine. AB - Pharmacological studies in laboratory animals show that systemically administered drugs that modulate the levels of specific central neurotransmitters can influence post brain injury behavioral recovery. Several lines of evidence support an important role of central norepinephrine. Selective lesions of central noradrenergic pathways impair recovery after a subsequent injury to the cerebral cortex. Drugs that deplete central norepinephrine, block alpha 1-adrenergic receptors, or decrease norepinephrine release (alpha 2-adrenergic receptor agonists) impede recovery whereas drugs that increase norepinephrine release (alpha 2-adrenergic receptor antagonists) or sympathomimetics (amphetamine) facilitate recovery. These studies also provide insights into the potential neurobiological processes underlying these drug effects. PMID- 10528359 TI - [Validation of procedures of rehabilitation after cerebrovascular accident]. AB - There are several methodological difficulties in the assessment of the effectiveness of rehabilitation in stroke patients. Obviously, double-blind studies are not possible, and only single-blind procedures can used. It is difficult to control for non specific effects, such as spontaneous recovery, patient's and therapist's motivation, social environment. It is necessary to assess the effect non only at the level of impairments, but also on disability and handicap. However, there are now accumulating data in the literature suggesting that stroke rehabilitation has a significant, although mild, effectiveness. Patients treated in specialized stroke rehabilitation units obtain a better outcome, in terms of independence in daily-life activities, than those treated in general wards. They also have shorter hospitalisation durations and are more frequently able to return home. Treatment effectiveness is related to intensity and duration of rehabilitation, and also to stroke severity. Patients with moderate impairments seem to benefit more from treatment than patients with mild or severe deficits. However, significant improvements can still be obtained in very severe cases, and even late (up to two years) post stroke. Similarly, rehabilitation of cognitive deficits (aphasia and unilateral neglect) has also been found efficient in most studies, even if the beneficial effect is relatively small. Aphasic patients treated by speech therapists improve more than patients treated by non specialized therapists or by family members who received a short training. One limitation of neglect rehabilitation is the inconsistent generalisation of treatment effects to daily-life situations. These data are encouraging but further research is needed to find out what precisely works, and how, in the "black box" of rehabilitation. PMID- 10528360 TI - [Arterial hypertension and cognitive decline]. AB - Arterial hypertension is the leading risk factor for all stroke subtypes. However, its relationship with cognitive decline and dementia is more complex than a simple causal relationship. Cognitive functions are worse in patients with arterial hypertension, especially when the level of education is lower, age higher and arterial hypertension more severe. Arterial hypertension is an independent factor of cognitive decline. It also leads to white matter changes which contribute to the cognitive decline. Longitudinal studies have shown that a higher blood pressure at the age of 70 years is associated with an increased risk of dementia (vascular or Alzheimer) 10 to 15 years later, but blood pressure spontaneously decreases as dementia occurs. Treatments of arterial hypertension decrease the incidence of stroke, but clinical trials are still necessary to determine if they also decrease the incidence of dementia. Preliminary results obtained in elderly subjects with systolic arterial hypertension, support this hypothesis. PMID- 10528361 TI - [Risk factors and mechanisms of post-stroke dementia]. AB - Stroke significantly increases the risk of dementia in subjects aged 55 years or more. Twenty to 25 p. 100 of patients are demented 5 years after a stroke. Age and supratentorial location of the vascular lesion are risk factors for post stroke dementia. Volume, left side of the lesion, large middle cerebral artery infarction, lesions of the frontal lobe, second stroke, diabetes, aphasia, clinical features expressing the severity of the stroke event in the acute phase, mitral valve prolapse, atrial fibrillation, depression, concomitant hypoxic/ischemic disorders, and white matter changes have also been found as predictors of dementia. There are many different mechanisms of vascular pathology that may lead to dementia: ischemic or hemorrhagic lesions, large vessel disease including multi-infarct and strategic single infarct, small-vessel disease including lacunes and white matter changes, hypoperfusion.... Post-stroke dementia may not be due only to vascular lesion. Some post-stroke dementias have a progressive onset and course. The cognitive decline may pre-exist to the stroke, even when a dementia is not diagnosed. This suggests a degenerative process. Alzheimer's disease is frequent in ages when the majority of strokes occur. Alzheimer's and vascular diseases share common risk factors such as age, APOE4, hypertension, and smoking. Patients with low MMS scores and AD patients are at risk for stroke. Moreover, white matter changes are associated with stroke and Alzheimer's disease and may contribute to the cognitive decline. Many post stroke dementias could be multifactorial. Even when vascular lesions and degenerative changes (mainly Alzheimer changes) are not severe enough, no their own, to be the cause of dementia, their summation may reduce the preclinical stage of the degenerative process. PMID- 10528362 TI - Pathophysiology of vascular dementia and white matter changes. AB - Vascular dementia (VaD) can be defined as a dementia syndrome likely to be the consequence of lesions of the brain, vascular in origin, irrespective of their ischemic, hemorrhagic or hypoxic nature. Six subtypes (1) multi-infarct dementia, (2) strategic single infarct dementia, (3) small-vessel disease with dementia, (4) hypoperfusion dementia, (5) hemorrhagic dementia and other VaD, have been proposed, indicating the broad clinical spectrum of this disorder. Major determinants of the dementia syndrome are (i) stroke characteristics--i.e., lacunar infarcts, localization in the left hemisphere and/or in strategic regions -, (ii) white matter changes frequently seen in stroke patients, especially in those who have lacunes or deep hemorrhages, represent a strong predictor for risk of dementia--and (iii) associated Alzheimer pathology--Alzheimer and vascular lesions are frequently associated. Finally, it should also be considered the role of the summation of various lesion types, since many cases of dementia occurring in stroke patients are multifactorial. PMID- 10528363 TI - Fuel metabolism during pregnancy. AB - This article reviews carbohydrate and fat metabolism in both healthy pregnant women and women with gestational diabetes. Emphasis is placed on more recent investigations that have utilized stable, nonradioactive isotopes with insulin clamps to study gestational fuel metabolism. In early pregnancy, glucose stimulated insulin secretion is increased, insulin sensitivity is unchanged or enhanced, and glucose tolerance is normal or slightly improved. Late gestation is characterized by accelerated fetal growth, rising concentrations of several diabetogenic hormones, and increased insulin resistance. The increased resistance reduces maternal glucose utilization, sparing carbohydrates for the rapidly growing fetus. The inhibitory effect of insulin on the rate of lipolysis is also significantly reduced during the third trimester of pregnancy. An earlier than normal switch from carbohydrate to fat utilization serves to promote the use of lipids as a maternal energy source. Women with gestational diabetes have been reported to have either comparable or increased insulin resistance during late gestation with several studies also demonstrating reduced insulin secretory capacity. PMID- 10528364 TI - Spontaneous abortions and major malformations in women with diabetes mellitus. AB - Women with insulin dependent diabetes mellitus are at increased risk for both first trimester spontaneous abortions and major congenital malformations when they become pregnant. The magnitudes of both of these risks depend upon the degree of metabolic control of their diabetes in the first trimester. The risks differ in the degree of control necessary to minimize them and the degree to which they can ultimately be reduced. A stricter degree of metabolic control is necessary to avoid spontaneous abortions than major malformations. Although the risks for both complications can be reduced by improved metabolic control, the risk for major malformations remains elevated, when compared to the risk for non diabetic women, despite good to excellent control. In contrast, good to excellent control does reduce the risk for spontaneous abortions to a rate comparable to that seen in non-diabetic women. Women with insulin dependent diabetes mellitus who are planning pregnancies should be encouraged to achieve the best possible degree of metabolic control prior to and throughout pregnancy. They should be re assured, however, that perfect control is not necessary to avoid dramatically increased risks for spontaneous abortions and major malformations. PMID- 10528365 TI - Diabetes and preimplantation events of embryogenesis. AB - From animal and human studies is it clear that mammalian embryos are vulnerable to injury during the earliest preimplantation stages of development. Exposure to some agents during this brief period has resulted not only in fetal loss but also in malformations. Thus, the potential for maternal induced embryotoxicity exists earlier than previously expected. This period is marked by a drastic change in metabolic needs at the late morula stage. Glucose becomes the predominant exogenous energy substrate and enters the blastocyst via one of three facilitative glucose transporters, GLUT1, GLUT2, and GLUT3. It has been shown that maternal diabetes adversely affects the preimplantation embryo. Recent work has revealed that hyperglycemia leads to a downregulation of the GLUTs at the blastocyst stage in the mouse. This downregulation results in decreased glucose transport into the blastocyst of diabetic mice and thus lower intraembryonic free glucose levels. The embryos are starving themselves of the key substrate necessary for survival. Maternal diabetes also causes a decrease in the number of cells in rat blastocyst and recently hyperglycemia has been shown to induce apoptosis in the mouse blastocyst via cell death effector pathways involving BAX and caspases. Significant loss of key progenitor cells from the blastocyst may predispose these diabetic embryos to later developmental deficiencies manifesting as dysmorphogenesis, fetal loss or early growth delay. The hypothesis that the hyperglycemia-induced decrease in glucose transport triggers apoptosis is presented and discussed as a novel mechanism to explain preimplantation diabetic embryopathy. PMID- 10528366 TI - Genotoxicity and diabetic embryopathy: impaired expression of developmental control genes as a cause of defective morphogenesis. AB - Since the advent of insulin therapy for diabetes mellitus, the survival of mothers with diabetes prior to pregnancy and their offspring has greatly improved. Nevertheless, the observation that the earliest stages of organogenesis can be impaired in the offspring of women with diabetes raises the question of how abnormal fuel metabolism disturbs embryogenesis. Research into this process has been made possible in recent years by advances in molecular biology which makes it possible to study gene expression in early embryos, and by the availability of genetically engineered mutant mouse strains. Using these approaches, a model is emerging in which elevated glucose, by disturbing expression of genes which regulate embryonic development and cell cycle progression, causes premature cell death of emerging organ structures, thereby causing defective morphogenesis. Investigation into the signaling mechanisms by which excess glucose metabolism exhibits toxic effects on embryo gene expression will explain how diabetic embryopathy occurs on a molecular and cellular level, as well as increase our understanding of the role of metabolic homeostasis in proper embryonic development. PMID- 10528368 TI - The role of prostanoids in the development of diabetic embryopathy. AB - In infants of diabetic mothers, congenital anomalies occur about two-three times as often as in normal population. Many etiologic factors have been proposed regarding the mechanism of diabetes related birth defects. The metabolic alterations associated with hyperglycemia include myo-inositol and arachidonic acid deficiency, and as a result disturbed metabolism of prostaglandins. Recent studies provide evidence that a deficiency in prostaglandins adversely affects membranogenesis and membrane function. These changes in membrane function permit the influx of high levels of glucose into the cells, inducing the generation of free oxygen radicals that cause morphologic damage of the embryo, involving aberrant mitochondrial function and enhanced peroxidation of embryonic lipids. The functional deficiency of prostaglandins at a critical time of fetal development can cause embryonic malformations. This paper reviews the role of prostanoids in the development of diabetic embryopathy. PMID- 10528367 TI - Signaling pathways and diabetic embryopathy. AB - Diabetic embryopathy is the leading cause of neonatal death and/or congenital malformations in infants of diabetic mothers. Because the development of the embryo critically depends on the maternal and the embryonic signaling pathways, a defective signaling mechanism between the maternal and the embryonic tissues appears to be involved in the etiology of diabetic embryopathy. Analyses of the recent studies from different laboratories suggest a "multifactorial" basis for diabetic embryopathy. These studies suggest that a wide variety of signal transducers converge towards the regulation of elcosanoid signaling pathway which appears to be the critical pathway involved in diabetic embryopathy. The characterization of the regulatory components of this pathway is likely to identify the signaling loci susceptible for the therapeutic intervention. PMID- 10528369 TI - Maternal fuels, diabetic embryopathy: pathomechanisms and prevention. AB - Congenital malformations remain the major cause of morbidity and mortality among the offspring of women with diabetes. Animal and human studies indicate that these anomalies occur very early in pregnancy and result from derangements of the maternal metabolic fuels which support embryogenesis. The mechanism for induction of dysmorphogenesis in experimental diabetic pregnancy has been shown to include deficiency states of membrane lipids (myoinositol, arachidonic acid, etc.), alteration in the prostaglandin cascade, and the generation of excess free oxygen radicals. These biochemical alterations result in characteristic morphological and molecular changes which are considered to be the basis of diabetic embryopathy. This article not only discusses the pathomechanism, but also reviews both clinical and experimental strategies to prevent diabetes-associated birth defects. PMID- 10528370 TI - Lumbar spinal fusion using recombinant human bone morphogenetic protein in the canine. A comparison of three dosages and two carriers. AB - STUDY DESIGN: A randomized, prospective and controlled animal study. OBJECTIVE: To evaluate lumbar spinal fusion using recombinant human bone morphogenetic protein 2 in a canine model. SUMMARY OF BACKGROUND DATA: Spinal fusion using autogenous bone grafting is associated with donor site morbidity and a nonunion rate of 5% to 35%. The use of recombinant human bone morphogenetic protein 2 as a bone graft substitute would eliminate donor site morbidity and perhaps augment the rate of successful fusion. METHODS: Mature beagles underwent bilateral paraspinal exposure at L4-L5, followed by transverse process decortication and randomization into one of six groups using differing doses of recombinant human bone morphogenetic protein 2 implanted using either a Type I collagen carrier or a polylactic acid carrier. Two control groups were used: one group without recombinant human bone morphogenetic protein 2 and another group using autogenous rib graft alone. RESULTS: Groups treated with recombinant human bone morphogenetic protein 2 demonstrated complete fusion in all animals. Animals treated with collagen carrier alone (no recombinant human bone morphogenetic protein 2) demonstrated complete absence of fusion. Successful fusion occurred in one of three canines in the autogenous bone graft group. Fusion masses in the recombinant human bone morphogenetic protein 2 treatment groups were significantly larger in size at 3 months than in the autogenous bone graft group. The collagen carrier was more biocompatible and biodegradable because residual polylactic acid carrier was seen with adjacent multinucleated giant cells. There was no evidence of spinal canal or nerve root encroachment in the recombinant human bone morphogenetic protein 2 treatment groups. CONCLUSIONS: The use of recombinant human bone morphogenetic protein 2 implanted using a Type I collagen carrier resulted in 100% fusion without adverse effects. PMID- 10528371 TI - Presence and distribution of antigen-antibody complexes in the herniated nucleus pulposus. AB - STUDY DESIGN: Herniated tissue was studied by immunohistochemistry in eight patients with lumbar disc herniation. The results were compared with those of control subjects. OBJECTIVE: To assess the presence and distribution of possible antigen-antibody complexes in herniated disc tissue. SUMMARY OF BACKGROUND DATA: It has been suggested that the nucleus pulposus may be recognized as a foreign body by the immune system and that this will lead to secondary nerve root disturbance. Such immunologic events should be initiated by binding of antibodies to a specific antigen in the disc tissue. However, the presence of antigen antibody complexes in the herniated disc tissue has not been assessed. METHODS: Amplification of the peroxidase reaction produced in avidin-biotin-peroxidase complex immunostaining by diaminobenzidine was used to visualize antigen-antibody complexes in the herniated tissue. The authors used herniated tissue from eight patients with lumbar disc herniation and nucleus pulposus from five control subjects with nonlumbar disc herniation. Thin paraffin sections, prefixed in 4% paraformaldehyde, were incubated with anti-human IgG antibody to allow visualization of antigen-antibody complexes in the specimens. RESULTS: A brown deposit, indicating antigen-antibody complexes, could be observed in the pericellular capsule in herniated disc tissue but not in control discs or in the residual discs of the herniation patients. CONCLUSION: Antigen-antibody complexes seem to be commonly present in herniated disc tissue, but not in healthy discs. However, the pathophysiologic and clinical significance of this observation has to be elucidated further. PMID- 10528372 TI - Pathogenesis of idiopathic scoliosis. Experimental study in rats. AB - STUDY DESIGN: A radiographic examination of pinealectomized rats to observe the development of scoliosis and halt the condition by administration of melatonin. OBJECTIVES: To discover whether pinealectomy has the same effect in mammals as shown in the chicken, and to determine whether the bipedal condition is important for development of scoliosis. SUMMARY OF BACKGROUND DATA: Pinealectomizing chickens shortly after hatching consistently resulted in scoliosis closely resembling human idiopathic scoliosis. It has not been determined whether this phenomenon is restricted solely to chickens, or if this experimental model is applicable to other animals, especially those more closely related to humans. METHODS: A sham operation in five bipedal rats served as the control in this study. Pinealectomy was performed in 10 quadrupedal rats, pinealectomy in 20 bipedal rats, and pinealectomy with implantation of melatonin pellet in 10 bipedal rats. Spinal radiographs were used to measure the degree of scoliosis at 3 months after surgery. RESULTS: Scoliosis developed only in pinealectomized bipedal rats and not in quadrupedal rats. It developed in none of the sham operation group and in only 1 of 10 pinealectomized bipedal rats with melatonin treatment. CONCLUSIONS: Melatonin deficiency secondary to pinealectomy alone does not produce scoliosis if the quadrupedal condition is maintained. The bipedal condition, such as that in chickens or humans, plays an important role in the development of scoliosis. The findings suggest a critical influence of a postural mechanism for the development of scoliosis. PMID- 10528373 TI - Cervical kyphosis in diastrophic dysplasia. AB - STUDY DESIGN: An evaluation of cervical kyphosis in diastrophic dysplasia from newborn to adult life. OBJECTIVES: To discover the prevalence and natural history of cervical kyphosis in diastrophic dysplasia. SUMMARY OF BACKGROUND DATA: Typical findings in this rare skeletal dysplasia are sport-limbed short stature, multiple joint contractures, early degeneration of joints, and spinal deformities such as cervical kyphosis, scoliosis, and exaggerated lumbar lordosis. In diastrophic dysplasia, spontaneous resolution of cervical kyphosis has been reported, but so have severe forms causing medullar compression leading to quadriplegia and death. The prevalence and clinical outcome of the kyphosis are not known. METHODS: The radiographic natural history of the cervical spine was studied in 120 patients. They varied in age from newborns to 63-year-olds. The average follow-up time in 26 living patients with cervical kyphosis was 10.0 years. RESULTS: Midcervical kyphosis was noted in 29 patients (24%) in their first radiograph. In 25 patients, the first radiographs were taken before the age of 18 months, and 24 of these patients (96%) had cervical kyphosis. The most severe case was that of a 32-year-old patient with a 165 degrees kyphosis. In the 24 patients, the kyphosis resolved spontaneously at an average age of 7.1 years. Three patients with a severe kyphosis died; one patient is alive. One patient, a 4-year-old child has mild resolving deformity. CONCLUSIONS: Cervical kyphosis in diastrophic dysplasia usually is shown at the time of birth. It resolves spontaneously during growth and seldom needs treatment. Careful follow-up study and treatment, if necessary, are important tools for avoiding the neurologic problems and fatal outcome. PMID- 10528374 TI - Soft cervical disc herniation. Influence of cervical spinal canal measurements on development of neurologic symptoms. AB - STUDY DESIGN: In 100 consecutive patients who underwent surgery because of soft cervical disc herniation, the sagittal and transverse diameters, the area of the bony cervical spinal canal, the sagittal diameter of the hernia, and the minimal bony intervertebral foramen diameter were measured by computed tomography. The data were compared with measurements from a control group of 35 matched healthy individuals. OBJECTIVES: To evaluate the relation between the severity of concurrent neurologic symptoms and the sagittal and transverse diameters, the cross-sectional area of the bony spinal canal, the sagittal diameter of the hernia, and diameter of the minimal bony intervertebral foramen in patients with soft cervical disc herniation. SUMMARY OF BACKGROUND DATA: Traumatic injury and spondylotic changes have a far greater impact on the spinal cord and nerve roots if the sagittal diameter of the bony cervical spinal canal is small. However, in the case of soft cervical disc herniation, no computer tomographic measurements are available for sagittal and transverse diameters, cross-sectional area of the bony spinal canal, sagittal diameter of the hernia, and diameter of the minimal bony intervertebral foramen in relation to the severity of concurrent neurologic symptoms. METHODS: Computed tomography was used to measure sagittal and transverse diameters, cross-sectional area of the bony cervical spinal canal, sagittal diameter of the hernia, and diameter of the minimal bony intervertebral foramen in 100 patients with symptomatic monosegmental cervical soft disc herniation. All patients had undergone an anterior discectomy with removal of the hernia and subsequent interbody fusion using an autologous bone graft taken from the iliac crest. RESULTS: A mean sagittal diameter of the bony cervical spinal canal of 12.9 mm was found, indicating a certain degree of developmental stenosis. Patients with motor disturbances had a significantly smaller sagittal diameter of the bony spinal canal than did patients without motor disturbances. There was a linear correlation between the sagittal diameter of the bony cervical spinal canal and that of the hernia. The sagittal diameter, the area of the bony spinal canal, and diameter of the minimal bony intervertebral foramen were significantly smaller in patients with soft cervical disc herniation than in the control group. CONCLUSIONS: Results from this study strongly suggest that the degree and severity of neurologic symptoms accompanying cervical soft disc herniation are inversely related to the sagittal diameter and the area of the bony cervical spinal canal. The latter area is reduced in cases of developmental stenosis or because of soft disc herniation. Moreover, patients with soft cervical disc herniation have a significantly smaller sagittal diameter of the bony spinal canal, a significantly smaller minimal bony intervertebral foramen diameter, and a significantly smaller cross-sectional area of the bony cervical canal than do healthy matched individuals. PMID- 10528375 TI - Nerve root pressure in lumbar disc herniation. AB - STUDY DESIGN: The contact pressure between the nerve root and lumbar disc herniation was measured and compared with clinical features. OBJECTIVE: To assess levels of actual compression to the nerve root in clinical cases. SUMMARY OF BACKGROUND DATA: Actual levels of pressure to the nerve root of lumbar disc herniation in clinical cases is unknown. METHODS: The study was performed on 34 patients who had lumbar disc herniation. All of them had been treated by open discectomy. After laminotomy, nerve root pressure was measured by inserting a transducer between the nerve root and the disc herniation. The magnitude of pressure was compared with clinical features. RESULTS: Nerve root pressures before discectomy were varied from 7 mm Hg to 256 mm Hg (mean, 53 mm Hg). After discectomy, the contact pressure was 0 mm Hg in all cases. There were no significant correlations between the magnitude of nerve root pressure and limits to the degree of straight leg raising, duration of symptoms, and age of the patients. However, the magnitude of the pressure in patients with neurologic deficits and trunk list was significantly higher than in the absence of these findings. CONCLUSIONS: The contact pressure exerted by lumbar disc herniation on the nerve roots was recorded during surgical intervention, and the mean pressure was 53 mm Hg. The magnitude of nerve root pressure was not correlated with the degree of straight leg raising, but with the severity of neurologic deficits. PMID- 10528376 TI - Idiopathic scoliosis. The clinical value of radiologists' interpretation of pre- and postoperative radiographs with interobserver and interdisciplinary variability. AB - STUDY DESIGN: A retrospective analysis of radiographic reports of 161 consecutive patients with idiopathic scoliosis at the authors' institution. OBJECTIVES: To compare various radiographic findings that directly affect surgical decision making and the evaluation of postsurgical outcomes to determine the usefulness of information gathered from radiologists' multiple duplicate reading of films. SUMMARY OF BACKGROUND DATA: To the authors' knowledge, there are no previous studies on the readings of scoliosis films by radiologists and surgeons. METHODS: The patient pool was drawn from the private practices of two board-certified orthopaedic surgeons. Each set of radiographs was read by one of seven board certified radiologists and by one of the two surgeons. The two reports of each radiograph were compared. The factors included in the reports were scoliosis deformity, scoliosis type, curve progression, curve magnitude, levels of the curve, kyphosis, lordosis, the presence of instrumentation, and the presence of a fusion. RESULTS: The radiologists and orthopedic surgeons mentioned the presence of scoliosis in 95% and 99.4% of their reports, respectively. The type of scoliosis was mentioned in 5% of reports by radiologists and in 99.4% by orthopedists. Progression of the curve was documented in 16.7% of the radiologists' reports and in 98.4% of orthopedists' reports. The magnitude of the curve was stated in 12.6% of the radiologists' reports, compared with 98.1% of the orthopaedists' reports. The levels of the curve were documented in 10.6% and 95.6% of reports by the radiologists and orthopedists, respectively. Radiologists mentioned kyphosis and lordosis in 28% and 26.5% of reports, respectively. These same two entities were mentioned in 98.2% and 79.4% of reports by the orthopedists. Finally, the radiologists noted the presence of instrumentation and of a fusion in 77.8% and 68.3% of reports, respectively. Orthopedists mentioned these same two entities in 84.4% and 100% of reports, respectively. In the radiologists' reports on the presence of instrumentation, 20% were mislabeled or improperly identified. Seven percent of the fusions documented by the radiologists were incorrect because they were recorded before biologic fusion could have taken place. In all these categories, the radiologists provided information in excess of the orthopedic reports a total of 1.9% of the time. Of this 1.9% additional information, 36.8% was incorrectly read or mislabeled. The other 63.2% of the additional information (1.9% of the total) did not elucidate anything of real clinical significance that was missed by the orthopedic surgeons (e.g., a tumor in the lung). CONCLUSIONS: These findings show that the attending orthopedic spine surgeons gained little useful information from the radiologists' multiple duplicate reading of films. PMID- 10528377 TI - Intraoperative comparison of two instrumentation techniques for the correction of adolescent idiopathic scoliosis. Rod rotation and translation. AB - STUDY DESIGN: A prospective and controlled comparative study of two instrumentation techniques used for the correction of adolescent idiopathic scoliosis. OBJECTIVE: To measure the three-dimensional intraoperative correction obtained with a rotation maneuver as compared with that obtained with a translation maneuver of the first instrumentation rod inserted to determine the difference, if any, in the two techniques for achieving three-dimensional correction. SUMMARY OF BACKGROUND DATA: Adequate three-dimensional correction of scoliotic deformities has been reported with the Cotrel-Dubousset instrumentation using the rod-rotation maneuver. More recently, however, authors of studies with newer instrumentation systems have claimed that better correction can be obtained using a translation technique. So far, no report has clearly demonstrated the three-dimensional changes obtained with this more recent instrumentation technique. METHODS: The changes in position of thoracic and lumbar vertebrae exposed during surgery were documented using a three-dimensional magnetic digitizer in 70 adolescents with idiopathic scoliosis undergoing correction by a posterior approach. Vertebral positions were measured intraoperatively before and after the surgical maneuver in 39 patients with the Cotrel-Dubousset instrumentation (rod rotation) and in 31 patients with the Colorado instrumentation (translation). RESULTS: In both groups, adequate three dimensional correction of the scoliotic deformities was documented, with significant changes in the frontal and sagittal planes and in the orientation of the plane of maximum deformity for thoracic and lumbar curves. On the other hand, no significant differences were documented between the two procedures except in the frontal plane where a tendency for greater correction was observed for thoracic curves with the translation technique. CONCLUSIONS: The two instrumentation techniques are equally able to achieve a comparable and effective three-dimensional correction of the scoliotic deformities. The use of either a rotation maneuver or a translation technique during surgery does not result in any significant measurable difference in three-dimensional correction. PMID- 10528378 TI - Recovery from exercise-induced desaturation in the paraspinal muscles in idiopathic scoliosis. AB - STUDY DESIGN: A study using near-infrared light spectroscopy to measure recovery from exercise-induced desaturation in the paraspinal muscles of patients with idiopathic scoliosis. OBJECTIVES: To measure oxygenation of the paraspinal muscles and obtain differences between the convex and concave sides. SUMMARY OF BACKGROUND DATA: Authors of previous studies have reported that some patients experience pain on the convex side of the paraspinal muscles. The muscles on the convex side are more stretched and stressed than those on the concave side. The current authors investigated the degree of stress by measuring oxygenation and blood volume changes. METHODS: Paraspinal muscle spectral properties at L3 were investigated using near-infrared light spectroscopy. Thirty-six patients (8 men and 28 women) underwent this procedure. To assess a level of peripheral adaptations to exercise, the half-time of Oxyhemoglobin/Myoglobin recovery was measured, which indicates the recovery from energy deficit after exercise. RESULTS: The average half-time recovery on the convex side was 3.38 seconds (range, 1.5-5.5 seconds), whereas that on the concave side was 1.51 seconds (range, 0.7-4.0 seconds). The average difference between the convex side and the concave side was 1.87 seconds (range, 0.9-3.5 seconds). CONCLUSIONS: Half-time recovery on the convex side in the patients with idiopathic scoliosis was slower than that in healthy adults (P < 0.05). Half-time recovery on the concave side in such patients was faster than that on the convex side (P < 0.01). The authors consider half-time recovery an indication of back muscle stress. PMID- 10528379 TI - Successful monitoring of neurogenic mixed evoked potentials elicited by anterior spinal cord stimulation through thoracoscopy during spine surgery. AB - STUDY DESIGN: Neurogenic mixed evoked potentials were recorded after thoracoscopic spinal cord stimulation in patients undergoing video-assisted spine surgery. OBJECTIVE: To demonstrate the feasibility and value of thoracoscopic spinal cord monitoring. SUMMARY OF BACKGROUND DATA: Video-assisted thoracic surgery recently has been proposed as a new technique for thoracic spine surgery. It can be used for anterior spinal release of patients with severe spinal deformities and for thoracic hernia removal. METHODS: Five patients undergoing video-assisted thoracic surgery for spinal fusion were studied. Neurogenic mixed evoked potentials were elicited by electrodes seated into intervertebral discs through thoracoscopy and recorded from peripheral nerves of the lower limbs. Moreover, the study included the case of a patient with a thoracic hernia who underwent video-assisted thoracic surgery with combined thoracoscopic neurogenic mixed evoked potential and standard somatosensory evoked potential monitoring. RESULTS: Neurogenic mixed evoked potentials were recorded consistently after spinal cord stimulation in all patients. For the patient with a thoracic hernia, neurogenic mixed evoked potentials suddenly disappeared, whereas somatosensory evoked potentials were not significantly modified, leading to surgery interruption. Neurogenic mixed evoked potentials progressively reappeared after a 30-minute delay. Postoperation examination revealed a Brown-Sequard's syndrome with incomplete right motor deficit. CONCLUSIONS: Neurogenic mixed evoked potentials evoked by anterior stimulation through thoracoscopy are of interest for spinal cord monitoring when posterior electrical stimulation is impossible, and they provide reliable information regarding spinal motor pathways. PMID- 10528380 TI - Intraoperative experience with neuromotor evoked potentials. A review of 60 consecutive cases. AB - STUDY DESIGN: A review of 60 surgical cases for correction of scoliosis, during which neuromotor evoked potentials and somatosensory evoked potentials were monitored. OBJECTIVES: To determine the validity and reliability of intraoperative neuromotor evoked potential monitoring in cases of scoliosis, where damage to the motor tracts of the spinal cord can occur. SUMMARY OF BACKGROUND DATA: Recently, the validity of neuromotor evoked potentials monitoring has been challenged, suggesting that the responses are not necessarily neuromotor evoked responses, but a combination of neuromotor and somatosensory evoked responses. This theory rendered the responses a potentially invalid measure of motor ability. However, despite controversy surrounding this topic, many professionals consider neuromotor evoked potential monitoring to be a successful and reliable measure of motor spinal cord function. METHODS: The results of neuromotor evoked potential testing were reviewed in 60 consecutive cases of children who underwent surgery for scoliosis. A standard protocol described in 1995 by Owen was used. RESULTS: Clinically useful neuromotor evoked potentials were obtained for 54 patients (90%). Inability to obtain neuromotor evoked potentials occurred in six patients (10%). CONCLUSIONS: This two-part study demonstrated the efficacy and reliability of neuromotor evoked potential monitoring during scoliosis surgery and examined the sources of difficulty in achieving accurate and valid results. PMID- 10528381 TI - Pain sensitivity as a correlate of clinical status in individuals with chronic low back pain. AB - STUDY DESIGN: A cross-sectional study of baseline correlates of clinical pain and functional status in consecutive patients being treated for chronic low back pain. OBJECTIVES: To determine if an individual's global pain sensitivity, measured by experimental pain threshold to pressure at various regions of the body, is associated with baseline measures of clinical pain and physical functioning. SUMMARY OF BACKGROUND DATA: Previous studies have demonstrated that in individuals with chronic low back pain, clinical pain and functional status are significantly associated with demographic, structural, and psychosocial factors. However, a large portion of variance remains unexplained. Because pain sensitivity (tenderness) has been shown to occur as a continuum in the population, the authors sought to determine if such sensitivity might be associated with clinical status in chronic low back pain, beyond what is known regarding demographic, structural, and psychosocial factors. METHODS: Forty-five patients with chronic low back pain were assessed for a variety of demographic, structural, and psychosocial factors, which previously have been shown to contribute to clinical status. In addition, all patients underwent testing for pain tolerance and threshold at various areas of the body. RESULTS: Age, degree of structural abnormality observed on magnetic resonance imaging, and depressive symptoms were all significantly correlated with either clinical pain or functional status. Pain sensitivity, the target of this investigation, accounted for significant proportions of variance in functional status and pain, even after controlling for demographic, structural, and psychosocial variables. CONCLUSIONS: These pilot data suggest that an individual's experimental pain threshold (a measure of tenderness) is associated with baseline functional status and pain in cases of chronic low back pain and may represent an important domain warranting further investigation. PMID- 10528382 TI - Management of chronic disabling low back pain with 360 degrees fusion. Results from pain provocation test and concurrent posterior lumbar interbody fusion, posterolateral fusion, and pedicle screw instrumentation in patients with chronic disabling low back pain. AB - STUDY DESIGN: A follow-up study conducted by an independent observer was performed on the authors' first 29 consecutive patients treated with concurrent posterior lumbar interbody fusion, posterolateral fusion, and pedicle screw instrumentation, for whom at least 2 years had transpired since the operation. OBJECTIVE: To evaluate the results of concurrent instrumented posterior lumbar interbody fusion and posterolateral fusion used to manage chronic disabling low back pain. SUMMARY OF BACKGROUND DATA: Patients chosen for surgery all had a history of chronic disabling low back pain exceeding 2 years and a sick leave period in excess of 6 months (average, 3.4 years). METHODS: From 1989 to 1993, 29 consecutive patients were surgically treated with fusion. The level of fusion was chosen depending on radiologic changes and results from a intradiscal injection provocation test. Bone union was verified by computed tomography scan with 1-mm thin slices and sagittal reformation, and by a "second look" in all but three patients. All patients were evaluated subsequently by an independent observer in November 1995, 4.7 years after surgery on the average. RESULTS: Bone fusion was obtained in 27 of the 29 patients (93%). There was a highly significant reduction in back and leg pain measurements. Of the 29 patients, the results were excellent in 9 patients (31%), good in 6 patients (21%), fair in 6 patients (21%), and poor in 8 patients (27%). A total of 18 patients (62%) had returned to work. CONCLUSION: The authors consider posterior lumbar interbody fusion with concurrent posterolateral fusion and pedicle screw instrumentation a possible method for managing chronic disabling low back pain. PMID- 10528383 TI - Kinematic magnetic resonance imaging of the upper cervical spine using a novel positioning device. AB - STUDY DESIGN: The development of a novel positioning device for magnetic resonance imaging of the upper cervical spine and an evaluation of motion patterns of the craniovertebral junction in asymptomatic volunteers as a part of the whole cervical spine motion. OBJECTIVES: To design and construct a positioning device that enables magnetic resonance imaging of the cervical spine in rotation, lateral bending, flexion, and extension in a horizontally open magnetic resonance scanner, and to define reference values for movements of the occiput (C0), the atlas (C1), and the axis (C2) in asymptomatic volunteers. SUMMARY OF BACKGROUND DATA: In previously used devices, the direction of motion is limited usually to flexion-extension, or the position of the head and neck are adjusted without a positioning device using semihard wedges or pillows. METHODS: Magnetic resonance imaging of the upper cervical spine in 20 asymptomatic individuals (10 men and 10 women) was performed in a horizontally open 0.23-T magnetic resonance imager in progressive steps during rotation, lateral bending, and flexion-extension using axial, coronal, and sagittal imaging planes, respectively. The positions of C0, C1, and C2 were measured, and pattern of motions between segments analyzed. Lateral displacement of the atlas during lateral bending and cranial migration distance during flexion-extension were assessed. RESULTS: The nonferromagnetic positioning device was designed and constructed. The motion patterns of the craniovertebral junction during rotation did not differ between the men and women, but in lateral bending there was a small difference between genders at C1-C2. In men, the position of C1 during flexion-extension was consistently more extended in relation to C0 and C2 than in women. CONCLUSIONS: The new positioning device allows magnetic resonance imaging of the upper cervical spine during flexion, extension, rotation, and lateral bending. To assess the relationship between C0-C1 and C1-C2 in flexion and extension, separate reference values for men and women are recommended. PMID- 10528384 TI - The anatomic relation of lateral mass screws to the spinal nerves. A comparison of the Magerl, Anderson, and An techniques. AB - STUDY DESIGN: Analysis of the anatomic relation of the Magerl, Anderson, and An screws to the spinal nerve. OBJECTIVES: To compare the potential incidence of nerve root (ventral and dorsal ramus) injury caused by the Magerl, Anderson, and An techniques. SUMMARY OF BACKGROUND DATA: Posterior plating with lateral mass screw fixation is a common procedure for managing an unstable cervical spine. Comparative study of the Roy-Camille and Magerl techniques has been reported. However, the risk of nerve root injury for the Anderson and An techniques is not known. METHODS: Three lateral mass screw insertion techniques were performed in this study: Magerl, Anderson, and An. Each technique involved two specimens and 20 screws inserted from C3 through C7. A 20-mm-long screw was used to overpenetrate the ventral cortex. The anterolateral aspect of the cervical spine was carefully dissected to allow observation of the screw-ramus relationship. RESULTS: The overall percentage of nerve violation was significantly higher with the Magerl (95%) and Anderson (90%) techniques than with the An (60%) technique (P < 0.05). The largest percentages of nerve violation for the Magerl, Anderson, and An screws were found at the dorsal ramus (50%), the bifurcation of the ventral dorsal ramus (45%), and the ventral ramus (55%), respectively. CONCLUSIONS: The results of this study indicate that the potential risk of nerve root violation is higher with the Magerl and Anderson techniques than with the An technique. PMID- 10528385 TI - Management of incidental durotomy without mandatory bed rest. A retrospective review of 20 cases. AB - STUDY DESIGN: A retrospective review of 20 patients with incidental durotomy treated without mandatory bed rest. OBJECTIVES: To determine whether patients with incidental durotomy can be treated effectively without multiple days of bed rest. SUMMARY OF BACKGROUND DATA: Incidental durotomy can cause postural headaches, nausea, vomiting, dizziness, photophobia, tinnitus, and vertigo. These symptoms are believed to result from a decrease in cerebrospinal fluid pressure, leading to traction on the supporting structures of the brain. Traditional management includes bed rest for up to 7 days to eliminate traction and reduce hydrostatic pressure during the healing process. METHODS: Twenty incidental durotomies were repaired intraoperatively with dural stitches and fibrin glue. Patients were allowed to ambulate according to the natural course after surgery without mandatory bed rest. Symptoms were monitored closely for 1 week, and long term follow-up assessments were obtained at a minimum of 10 months. RESULTS: Of the 20 patients in this study, 75% had no symptoms after repair of the incidental durotomy. Each of the dural tears was 1-3 mm in length. Two patients reported headache, two reported nausea, and one reported tinnitus; no patients experienced vomiting. One patient (5%) had stitch loosening requiring revision surgery. There were no additional serious complications. CONCLUSIONS: This study has shown that the majority of patients with incidental durotomy can be treated effectively with dural stitches and fibrin glue. Patients can be permitted to ambulate immediately after surgery but should be cautioned to lay flat if they develop symptoms. This will reduce the costs related to the hospital stay and missed work. PMID- 10528386 TI - Thoracic disc herniation mimicking acute lumbar disease. PMID- 10528387 TI - Comparison of anterior and posterior instrumentation for correction of adolescent thoracic idiopathic scoliosis. PMID- 10528388 TI - [Risk management program in a health facility using a project approach: transfusion risk management]. AB - Risk management in the hospital, which is one of the referentiels of the ANAES accreditation manual, may be considered on two levels. Firstly, risk management may be approached globally, in the same way as it is tackled in the accreditation process. Secondly, risk management may be more definite. A specific risk chosen in accordance with the priorities of a particular plan may be dealt with individually. In this respect, the tranfusion process allows the risk management method to be tested and developed. f1 PMID- 10528389 TI - [Good transfusion safety practices. Groupe Recherche et Demarche Qualite. Societe Francaise de Transfusion Sanguine]. PMID- 10528390 TI - [Autologous transfusion: when best should not be the enemy of good enough...]. AB - Autologous blood transfusion has been shown to decrease allogeneic transfusion in patients undergoing elective procedures, in adults as well as in children. However, its indication must be carefully discussed for each patient, since, on the one hand, viral risks associated with allogeneic blood are greatly reduced, while on the other hand, adverse events may occur in some patients with poor physical condition. An assessment of the ratio 'benefit-risk' has to be made for each patient. PMID- 10528391 TI - C-H...O interactions in dimethyl 6,6'-dimethoxy-3,3',5,5'-tetramethylbiphenyl 2,2'-dicarboxylate. AB - The title compound, C22H26O6, crystallized in the centrosymmetric space group Pbcn with half of the molecule as the asymmetric unit (the molecular symmetry is 2). The H atoms of each of the four methyl groups in the asymmetric unit are disordered over two sites. Nine leading C-H...O interactions are present, with H...O distances ranging from 2.50 to 2.87 A and C...O distances ranging from 3.157 (3) to 3.567 (3) A. The various interaction chains from a three-dimensional network. The intramolecular angle between the biphenyl ring planes (twist angle) is 71.3 (1) degree. PMID- 10528392 TI - Hydrogen bonding and C-H...X interactions in two triclinic phases of 4 carboxyquinolinium chloride monohydrate. AB - The title substance, C10H8NO2+.Cl-.H2O, crystallized in the centrosymmetric space group P1 in two phases, alpha and beta, each with one organic cation, one Cl- ion and one water molecule in the asymmetric unit. The principle structural difference in the two asymmetric units lies in the relative orientation of the water molecule. Three hydrogen bonds in the alpha phase have donor-acceptor distances (N...Cl or O...Cl) ranging from 3.052 (1) to 3.189 (2) A, while one has an O...O distance of 2.603 (3) A. Three hydrogen bonds in the beta phase have donor-acceptor distances (N...Cl or O...Cl) ranging from 3.044 (2) to 3.206 (2) A, while two have O...O distances of 2.586 (2) and 3.147 (3) A. The H atoms in all these hydrogen bonds are ordered. Each of these phases has five leading C H...X interactions, for which the H...X distances are less than, or at most slightly greater than, the corresponding van der Waals radius sums. Taken together, these hydrogen bonds and C-H...X interactions form a three-dimensional network of interactions in each phase. The dihedral angle between the best-fit quinoline core plane and the carboxyl group plane is 25.9 (1) degree for the alpha phase and 20.0 (1) degree for the beta phase. PMID- 10528393 TI - Hydrogen bonding and C-H...O interactions in bis(8-dimethylamino-1 dimethylammonionaphthalene) [(DMANH+)2] 4,8-dicarboxynaphthalene-1,5 dicarboxylate dihydrate. AB - The title substance, 2C14H19N2+.C14H6O8(2-).2H2O, crystallized in the centrosymmetric space group P1 with one monopositive dimethylamine dimethylammonio cation, half of a dinegative acid anion and one water molecule in the asymmetric unit, with the anion located on a center of symmetry. There are four intermolecular hydrogen bonds, of which three have Odonor...Oacceptor distances of 2.577 (1), 2.946 (2) and 2.764 (2) A, while the fourth has an Ndonor...Oacceptor distance of 3.154 (2) A. There is a single intramolecular hydrogen bond, in which the Ndonor...Nacceptor distance is 2.620 (2) A. In each case, the O, or N, and H atoms are ordered. The Ndonor-H distance in the intramolecular hydrogen bond, 1.06 (1) A, is among the shortest seen in this cation at room temperature. Moreover, the antiplanar orientation of the H atom in the hydrogen bond between acid anions is noteworthy. There are, in addition, 12 significant intermolecular C-H...O interactions. In these, the C...O distances range from 3.229 (2)-3.671 (2) A, while the C-H...O angles range from 121-165 degrees for the fixed H atoms involved. A three-dimensional network of hydrogen bonding interactions is generated. PMID- 10528394 TI - Bis(1,3-dioxan-2-yl)arenes: precursors to linked porphyrins. AB - The crystal structures of two linked-prophyrin precursors have been determined. In the crystalline state, 1,4-bis[2-(1,3-dioxan-2-yl)phenyl]butane, C24H30O4, (I), exists in a stair-like conformation with dioxane rings in chair conformations. The molecule is positioned on an inversion center. In the crystalline state, 1,2-bis(1,3-dioxan-2-yl)benzene, C14H18O4, (II), is arranged in zigzag chains. In each chain, molecules of (II) are tilted 106.2 (1) degree relative to each other. They are also oriented to form a weak C-H...O contact (H...O 2.43 A). PMID- 10528395 TI - Phylogenetic analyses of Bradyrhizobium strains nodulating soybean (Glycine max) in Thailand with reference to the USDA strains of Bradyrhizobium. AB - To elucidate the phylogenetic relationships between Thai soybean bradyrhizobia and USDA strains of Bradyrhizobium, restriction fragment length polymorphism (RFLP) analysis using the nifDK gene probe and sequencing of the partial 16S rRNA gene were performed. In our previous work, Thai isolates of Bradyrhizobium sp. (Glycine max) were separated clearly from Bradyrhizobium japonicum and Bradyrhizobium elkanii based on the RFLP analysis using the nodDYABC gene probe. RFLP analysis using the nifDK gene probe divided 14 Thai isolates and eight USDA strains of B. japonicum into different groups, respectively, but categorized into the same cluster. All of seven strains within these Thai isolates had the same sequence of the partial 16S rRNA gene, and it was an intermediate sequence between those of B. japonicum USDA 110 and B. elkanii USDA 76T. Furthermore, three USDA strains of B. japonicum, USDA of (B. japonicum ATCC 10324T), USDA 115 and USDA 129, had the same partial 16S rRNA gene sequence that seven Thai isolates had. These results suggest that Thai isolates of Bradyrhizobium sp. (Glycine max) are genetically distinct from USDA strains of B. japonicum and B. elkanii, but also indicate a close relationship between Thai isolates and USDA strains of B. japonicum. PMID- 10528396 TI - Insertional inactivation of the Listeria monocytogenes cheYA operon abolishes response to oxygen gradients and reduces the number of flagella. AB - The nucleotide sequence of a region downstream of the Listeria monocytogenes flagellin gene, flaA, revealed two putative chemotaxis genes, cheY and cheA. These genes have been shown to be transcribed as a bicistronic unit. In this study Tn916 delta E mutagenesis was used to generate two mutants, PF10 and PF16, which contain transposon inserts in the promoter region of this operon. These mutants were motile in liquid, but had reduced flagellin expression and were unable to burrow or swarm on soft agar plates. Complementation of the single transposon-copy mutant PF16 with cloned cheY and cheA in trans partially restored microaerotaxis and swarming on soft agar. The complemented strain did not exhibit any increase in flagellin production. Both PF10 and PF16 appear deficient in their ability to attach to the mouse fibroblast cell line 3T3. PMID- 10528397 TI - Enhanced benzaldehyde formation by a monokaryotic strain of Pycnoporus cinnabarinus using a selective solid adsorbent in the culture medium. AB - A monokaryotic strain of the white-rot fungus Pycnoporus cinnabarinus was shown to produce, in a 2-L bioreactor culture, 100 mg.L-1 benzaldehyde (bitter almond aroma) from L-phenylalanine with a productivity of 33 mg.L-1.day-1. The addition of HP20 resin, a styrene divinylbenzene copolymer highly selective for benzaldehyde, enabled an eightfold increase in the production of benzaldehyde and a twofold increase in productivity. In the presence of HP20 resin, the production of 790 mg.L-1 benzaldehyde was concomitant with the synthesis of cinnamic acid derivatives of high organoleptic notes such as cinnamaldehyde, cinnamyl alcohol, and methyl cinnamate. PMID- 10528398 TI - The porin OmpC of Salmonella typhimurium mediates adherence to macrophages. AB - Macrophages recognize, adhere to, and phagocytose Salmonella typhimurium. The major outer membrane protein OmpC is a candidate ligand for macrophage recognition. To confirm this we used transposon mutagenesis to develop an ompC deficient mutant in a known virulent strain of S. typhimurium; mutant and wild type were compared in macrophage adherence and association assays. Radiolabeled wild type S. typhimurium bound to macrophages at five-fold higher levels than did the ompC mutant. In association assays, macrophages in monolayers bound and internalized three-fold more wild type than mutant, while macrophages in suspension bound and internalized 40-fold more wild type than mutant. The ompC gene of our test strain of S. typhimurium contains several discrete differences compared with the ompC genes of Salmonella typhi and Escherichia coli. The deduced OmpC amino acid sequence of S. typhimurium shares 77 and 98% identity with OmpC amino acid sequence of E. coli and S. typhi, respectively. Evidence from this study supports a role for the OmpC protein in initial recognition by macrophages and distinguishes regions of this protein that potentially participate in host-cell recognition of bacteria by phagocytic cells. PMID- 10528399 TI - Bacterial surface antigen-specific monoclonal antibodies used to detect beer spoilage pediococci. AB - Fourteen monoclonal antibodies (Mabs) were isolated that react with surface antigens of Pediococcus beer spoilage organisms, including P. damnosus, P. pentosaceous, P. acidilactici, and unspeciated isolates. Immunoblotting, enzyme immunoassays (EIAs) of protease- and neuraminidase-treated surface antigen extracts, carbohydrate competition EIAs, and cardiolipin EIAs were used to characterize the bacterial antigens involved in Mab binding. Antigen stability in situ was tested by protease treatment or surface antigen extraction of washed bacteria. In most cases, the Mabs bind to Pediococcus surface antigens that appear to be covalently bound cell wall polymers resistant to alteration or removal from the bacterial surface. These bacterial surface antigen reactive Mabs show good potential for rapid, sensitive, and specific immunoassay detection of Pediococcus beer spoilage organisms. PMID- 10528400 TI - Trichosporon guehoae sp.nov., an anamorphic basidiomycetous yeast. AB - A morphological and physiological description of an anamorphic basidiomycetous yeast species, named Trichosporon guehoae (CBS 8521T), is presented. The ability to assimilate several aliphatic and aromatic compounds as sole source of carbon and energy is reported. The phylogenetic position within the genus, based on nuclear base sequencing of the D1/D2 region of the large subunit of rDNA is discussed. PMID- 10528401 TI - The effect of water, ascorbic acid, and cranberry derived supplementation on human urine and uropathogen adhesion to silicone rubber. AB - In this study, urine was collected from groups of volunteers following the consumption of water, ascorbic acid, or cranberry supplements. Only ascorbic acid intake consistently produced acidic urine. Photospectroscopy data indicated that increased water consumption produced urine with lower protein content. Surface tension measurements of the collected urine showed that both water and cranberry supplementation consistently produced urine with surface tensions higher than the control or urine collected following ascorbic acid intake. These urine samples were also employed to study uropathogen adhesion to silicone rubber in a parallel plate flow chamber. Urine obtained after ascorbic acid or cranberry supplementation reduced the initial deposition rates and numbers of adherent Escherichia coli and Enterococcus faecalis, but not Pseudomonas aeruginosa, Staphylococcus epidermidis, or Candida albicans. Conversely, urine obtained from subjects with increased water intake vastly increased the initial deposition rates and numbers of adherent E. coli and E. faecalis (P < 0.05). PMID- 10528402 TI - Glycerol production by yeasts under osmotic and sulfite stress. AB - The yeasts Saccharomyces cerevisiae, Candida boidinii, Pichia augusta, and Pichia anomala were tested for glycerol production both under osmotic stress and by addition of a sulfite-steering agent. The osmotic pressure was increased by employing glucose concentrations from 50 to 200 g/L and by supplementing with NaCl (40 g/L). Of all the yeasts, S. cerevisiae exhibited the highest level of osmotolerance. The increased osmotic pressure affected glycerol formation the most in C. boidinii. In both Pichia species, glycerol formation was not sufficiently induced when exposed to sugar and salt stress. The addition of 40 g/L Na2SO3 to the medium containing 100 g/L glucose shifted the metabolism of all yeasts towards glycerol formation. Saccharomyces cerevisiae achieved 68.6%, while C. boidinii reached 25.5% of the theoretical glycerol yield, respectively. The highest glycerol yield, 82.3% of the theoretical, was produced by S. cerevisiae under microaerophilic conditions. PMID- 10528403 TI - Poor efficacy of residual chlorine disinfectant in drinking water to inactivate waterborne pathogens in distribution systems. AB - To evaluate the inactivating power of residual chlorine in a distribution system, test microorganisms (Escherichia coli, Clostridium perfringens, bacteriophage phi X 170, and poliovirus type 1) were added to drinking water samples obtained from two water treatment plants and their distribution system. Except for Escherichia coli, microorganisms remained relatively unaffected in water from the distribution systems tested. When sewage was added to the water samples, indigenous thermotolerant coliforms were inactivated only when water was obtained from sites very close to the treatment plant and containing a high residual chlorine concentration. Clostridium perfringens was barely inactivated, suggesting that the most resistant pathogens such as Giardia lamblia, Cryptosporidium parvum, and human enteric viruses would not be inactivated. Our results suggest that the maintenance of a free residual concentration in a distribution system does not provide a significant inactivation of pathogens, could even mask events of contamination of the distribution, and thus would provide only a false sense of safety with little active protection of public health. Recent epidemiological studies that have suggested a significant waterborne level of endemic gastrointestinal illness could then be explained by undetected intrusions in the distribution system, intrusions resulting in the infection of a small number of individuals without eliciting an outbreak situation. PMID- 10528404 TI - Biosynthesis of trehalose from maltooligosaccharides in Rhizobia. AB - Previously, the enzymes for trehalose synthesis that are present in Escherichia coli were demonstrated in Bradyrhizobium japonicum and B. elkanii. An alternative mechanism recently reported for the synthesis of trehalose from maltooligosaccharides was considered based on the fact that high concentrations of sugars in liquid culture stimulated the accumulation of trehalose. An assay for the synthesis of trehalose from maltooligosaccharides using crude, gel filtered protein preparations was developed. Analysis of a variety of the Rhizobiaceae indicates that the "maltooligosaccharide mechanism" is present in B. japonicum, B. elkanii, Rhizobium sp. NGR234, Sinorhizobium meliloti, R. tropici A, R. leguminosarum bv viciae, R. I. bv trifolii, and Azorhizobium caulinodans. Synthesis of trehalose from maltooligosaccharide could not be detected in R. tropici B or R. etli. PMID- 10528405 TI - The pharmacological flexibility of the insect voltage gated sodium channel: toxicity of AaIT to knockdown resistant (kdr) flies. AB - AaIT is an insect selective neurotoxic polypeptide shown to affect insect neuronal sodium conductance by binding to excitable sodium channels. In the present study the paralytic potency of AaIT to wild type and various mutant strains of houseflies (Musca domestica) and fruitflies (Drosophila melanogaster) was examined and it has been shown that: On the basis of body weight when compared to published data on Sarcophaga falculata blowflies, the Musca and Drosophila flies reveal at least two orders of magnitude decreased susceptibility to the AaIT. When compared to wild type flies the toxicity of AaIT is greatly altered in knockdown resistant fly strains which are mutated in their para gene encoding the voltage gated sodium channel. Several strains, with genetically mapped para mutations conferring pyrethroid resistance, exhibited opposing response to AaIT. The para ts2 Drosophila strain, with a point of mutation in domain I of the para gene conferring a 6-fold resistance to deltamethrin also showed about 15-fold tolerance to AaIT. On the other hand the Musca kdr and super kdr flies, with a single or a double point mutation, respectively in domain II of the para gene, are about 9- and 14-fold more susceptible to AaIT, respectively. The above data are interpreted in terms of the pharmacological diversity and flexibility ("allosteric coupling") of voltage gated sodium channels and their implications for the management of pesticide resistance are discussed. PMID- 10528406 TI - Insulin stimulates ecdysteroid production through a conserved signaling cascade in the mosquito Aedes aegypti. AB - Selective activators and inhibitors of insulin signaling cascades in mammalian cells were tested for their effects on insulin stimulated steroidogenesis by ovaries of Aedes aegypti. Bovine insulin in the concentration range of 1.7 microM to 85 microM stimulated ecdysteroidogenesis in vitro. Pervanadate, an inhibitor of tyrosine kinase phosphatase, stimulated ecdysteroid production at concentrations of 250 microM to 1 microM. Okidaic acid, a serine/threonine phosphatase inhibitor, stimulated steroidogenesis with an ED50 of 77.39 nM. A selective inhibitor of tyrosine kinase activity, HNMPA-(AM3), inhibited ecdysteroid production with an IC50 of 14.2 microM. Two selective inhibitors of phosphatidylinositol 3-kinase, wortmannin and LY294002, inhibited ecdysteroid production at low concentrations (IC50 = 1.6 nM and 30 nM, respectively). These concentrations are similar to those inhibiting insulin action in mammalian cells. A selective inhibitor of mitogen-activated protein kinase, PD098059, had no effect on ecdysteroid production even up to 100 microM. Thus, insulin stimulation of ecdysteroid production by ovaries in vitro appears to be controlled by the tyrosine kinase activity of the mosquito insulin receptor and the signaling cascade involving phosphatidylinositol 3-kinase and protein kinase B. PMID- 10528407 TI - Antisera against Periplaneta americana Cu,Zn-superoxide dismutase (SOD): separation of the neurohormone bursicon from SOD, and immunodetection of SOD in the central nervous system. AB - In an effort to characterize the insect molting hormone bursicon from the cockroach, Periplaneta americana, amino acid sequences with high identity of Cu,Zn-superoxide dismutase (SOD) of Drosophila virilis were identified. Antisera against a conserved region of SOD, and a sequence unique to Periplaneta SOD were produced and used to test whether bursicon might be a form of SOD. Western blots of one- and two-dimensional gels revealed that the dimeric form of SOD and bursicon have a similar molecular mass (30 kDa). The two proteins can be separated, however, according to their different isoelectric points. Bursicon is identified in two-dimensional gels by elution from four unique spots not labeled by the anti-SOD antisera. In sections of Periplaneta nerve cords the antisera labeled glial material surrounding neuronal somata close to the neural sheath. Bursicon, however, is contained in unique cell pairs in the ganglia of the ventral nerve cord. These neurons were labeled with new antisera produced against novel sequences of one of the four above-mentioned bursicon active spots. The results show unequivocally that SOD and bursicon are distinctly different proteins. Furthermore, the anti-SOD antisera provided a tool to isolate and sequence bursicon. PMID- 10528408 TI - Amplified detection, using a monoclonal antibody, of an aphid-specific epitope exposed during digestion in the gut of a predator. AB - Monoclonal antibodies are invaluable tools for identifying and quantifying prey remains in the fore-guts of predators. However, they must be target-specific, detect an epitope that is well replicated within the prey (to enhance assay sensitivity) and, critically, recognise a site that can resist digestion. A monoclonal antibody is reported that proved to be aphid-specific and capable of detecting, and accurately identifying, as little as 16.5 ng of aphid protein within a heterologous mixture of invertebrate material. The antibody was selected by screening hybridoma lines for antibodies that bound with semi-digested aphid proteins. The antibody detected an epitope that was found, against expectation, to significantly increase in concentration with time (by approx. 50% over 6 h) in the gut of the carabid predator Pterostichus melanarius. The resultant extended antigen detection period and half-life, and the high specificity of this antibody, showed it to be an enhanced tool for studying interactions between aphids and their predators in the field. It was concluded that the antibody was initially generated to a surface epitope on a high molecular weight native protein (> 200 kD). This epitope, however, was then either replicated on internal sites progressively revealed by digestion, or new epitopes became available as the conformation of the protein changed during digestion. PMID- 10528409 TI - Expression of lacunin, a large multidomain extracellular matrix protein, accompanies morphogenesis of epithelial monolayers in Manduca sexta. AB - Morphogenesis is a complex process operating at several levels of organization- organism, tissues, cells, and molecules. Complex interactions occur between and within these levels. Many of the molecules that mediate these interactions are predictably turning out to be large multidomain proteins. Here we describe one such novel protein associated with remodeling of epithelial monolayers in embryos and developing wings of the moth Manduca sexta. On the basis of its sequence and its expression pattern along lacunae of developing wings, we propose the name lacunin for this extracellular matrix protein that contains nine different types of domains, most of which are present in multiple copies. These include domains of various types: Kunitz proteinase inhibitors, thrombospondin type I, immunoglobulin-like, and several newly defined domains of unknown function (PAL, PLAC, and lagrin domains). This rich patchwork of distinct domains probably exerts multiple effects on a variety of cell behaviors associated with the complex phenomenon of epithelial morphogenesis. PMID- 10528410 TI - Production and characterization of monoclonal antibodies to two new mosquito Aedes aegypti salivary proteins. AB - Mosquito salivary proteins, which are fundamental to the process of blood feeding, also facilitate disease transmission and cause allergic reactions. The identification and characterisation of these proteins have been hampered by the difficulty of obtaining them in purified form. In this report, we describe the production of mouse monoclonal antibodies (mAbs) against mosquito salivary proteins. BALB/c mice were immunised with Aedes aegypti saliva proteins. Hybridomas were produced by fusion of spleen cells with a mouse myeloma cell line. Positive clones were selected using a saliva-capture ELISA and further identified using immunoblotting. Three mAbs reacted with a 44 kDa protein (Aed a X1) in the saliva-immunoblotting, and did not react with 2 recombinant salivary proteins, rAed a 1 (apyrase) and rAed a 2 (D7), in both immunoblotting and ELISA. Two other mAbs reacted with a 37 kDa protein in saliva-immunoblotting, but failed to react with the 37 kDa rAed a 2 in either immunoblotting or ELISA, suggesting that there is a second 37 kDa protein (Aed a X2) which is recognised by the two mAbs. The 44 kDa and 37 kDa proteins have not been previously identified. These mAbs provide a means to purify proteins, to isolate new genes from the salivary gland cDNA library, and to standardise mosquito extracts, facilitating studies of disease transmission by mosquitoes and of mosquito allergy. PMID- 10528411 TI - Transcriptional activation of the cloned Heliothis virescens (Lepidoptera) ecdysone receptor (HvEcR) by muristeroneA. AB - Ecdysteroids play an important role during insect development. We report here the isolation and characterisation of an Ecdysone receptor (EcR) homologue from Heliothis virescens (HvEcR) and present evidence supporting the HvEcR active role as an active component of the native insect receptor. Alignment of the deduced amino acid sequence of HvEcR with those of EcRs from other species confirmed its membership of this family and showed that it is closely related to the B1 isoform of Drosophila melanogaster. Northern blot analysis showed that two transcripts (6.0 and 6.5 kb) were recognised by a probe spanning the DNA and ligand binding domains of the HvEcR. Genomic Southern blots showed that the HvEcR is encoded by a single copy gene. Two lines of evidence towards the functional activity of the HvEcR are presented. In vitro transcribed and translated HvEcR showed specific binding to hsp27 and pall response elements in the presence of CfUSP. Stable expression of HvEcR in 293 cells induced reporter gene activity in the presence of muristeroneA in a dose dependant manner while dexamethasone failed to activate. PMID- 10528412 TI - Functional characterization of two Ultraspiracle forms (CtUSP-1 and CtUSP-2) from Chironomus tentans. AB - Two forms, CtUSP-1 and CtUSP-2, of the Chironomus tentans homolog of Ultraspiracle (new nomenclature: Chironomus NR2B4) were described and verified as components of the functional ecdysteroid receptor. The two forms differed from each other in the most N-terminal regions of the A/B domain and were tested for several properties. Both forms showed the ability to heterodimerize with CtEcR and interact with a variety of direct repeat and palindromic EcREs, and both conferred specific ligand binding when heterodimerized with EcR. CtUSP-2 showed a twofold higher ponasterone-binding potential than CtUSP-1. Both USP forms demonstrated the ability to activate ecdysteroid-inducible transcription in HeLa cells and the variations in the A/B domain of these forms were not associated with detectable differences in transcriptional activation. Thus, the two forms function similarly. Among species for which USP forms have been reported, Chironomus is the most closely related one evolutionarily to Drosophila. Despite this proximity, a variety of structural differences were noted in both the A/B and E domains of USP between the two species. The Chironomus USP forms lack many of the amino acid residues associated with the ligand-dependent AF2 transactivation function found in all other RXRs and USPs reported so far. PMID- 10528413 TI - Functional binding of a vertebrate hormone, L-3,5,3'-triiodothyronine (T3), on insect follicle cell membranes. AB - A vertebrate hormone, L-3,5,3'-triiodothyronine (T3), induces volume reduction in the follicle cells of Locusta migratoria and Rhodnius prolixus. The effect of T3 on locust follicle cells is inhibited by ouabain and by antibodies raised against a membrane binding protein for juvenile hormone (JH). [125I]-T3 binds to membrane preparations of vitellogenic follicles in a specific and saturable fashion, with a KD in the low nanomolar range. T3 and JH III exhibited equivalent abilities to compete for the T3 binding site. These findings strongly suggest that T3 and JH act via the same receptor in follicle cells. PMID- 10528414 TI - [Effect of anesthetics on the function of the gastrointestinal tract]. AB - In spite of numerous interactions between the gut and the entire organism, today's knowledge in this field is still limited. In intensive care patients, reduced gastrointestinal perfusion and motility result in sequestration of fluids and translocation of bacteria and endotoxins, and the immunological function of the gut is depressed. To prevent gastrointestinal organ failure, early restitution of enteral nutrition is a main goal in intensive care medicine. Thus, the influence of anaesthetics on gut function is of special importance in analgosedation of intensive care patients. Pharmacological data of common anaesthetics allow judgement of their global effects on the gut. Interactions with opioid receptors of the enteral nerve system and systemic effects on the vegetative nerve system are of special interest. The results of in vitro and clinical studies show profound negative effects of opiods on gastrointestinal motility. Piritramide seems to be excluded from this judgement, but further studies with equipotent analgetic doses, when compared with fentanyl, are necessary. Ketamine is an analgetic alternative without relevant negative effects on gastrointestinal motility. Among the sedative components of analgosedation, midazolam, gamma-hydroxy butyric acid and probably propofol are useful, whereas barbiturates seem to have negative effects. Epidural anaesthesia with local anaesthetics is of additional benefit. PMID- 10528415 TI - [Practical aspects of early enteral feeding]. AB - "Gut injury" and a corresponding impaired gut barrier function are thought to have a high impact on the development of multiple organ failure (MOF) in the critically ill. Mucosal lesions and increased intestinal permeability can provoke translocation of bacteria and endotoxins and initiate local and/or systemic immune-inflammatory response, bearing the risk of development of multiple organ failure. Enteral nutrition using the physiological pathway provides the intestinal mucosa with nutrients, which is thought to reduce bacterial translocation and septic complications. Considerable gastric reflux and delayed bowel motility are the principal problems of enteral nutrition. Therefore, in the early postoperative period at least a nasoduodenal or--jejunal feeding tube or feeding jejunostomy is required. The commonly used enteral formulas are well tolerated. So-called "immunonutrition" includes special formulas supplemented with immunemodulating substances like arginine, omega-3-fatty acids, ribonucleic acids and glutamine. Some beneficial effects of immune-enhancing diets have recently been reported for immune response, infectious complication rate, systemic inflammatory response syndrome (SIRS), multiple organ failure (MOF), antibiotic usage and length of hospital stay, especially in patients after trauma or surgery. However, the definite role is still unknown and indications have still to be defined. Enteral feeding should start with small volumes, the amount being gradually increased according to a patient's individual tolerance. Common problems are gastric reflux, diarrhoea and distension, but usage of a suitable formula, a gradual increase or reduction in the amount of enteral feeding and, additionally, parenteral nutrition can help to overcome such complications. Clinical examination of the enterally fed patient should be performed carefully. Standard nutritional monitoring of electrolytes, glucose, triglycerides, cholinesterase, albumin, differential blood count, urine-glucose and nitrogen retention to assess the catabolic state should be performed routinely. Although only little data from randomised trials are available, enteral nutrition has advantages and is cheaper than total parenteral nutrition. In the critically ill, the goal of enteral feeding is not coverage of total caloric requirements, but continuous administration of at least a small amount in order to prevent gut mucosa atrophy. Nutrition is an important aspect in critical care medicine, and enteral feeding should be attempted at least partially. PMID- 10528416 TI - [Quality of washed autologous erythrocytes from drainage-suction pumps]. AB - High sub-pressure in high-vacuum suction bottles falls as the bottles fill up. Suction pumps with reservoir have a constant low suction level and decisive advantages. The question is: does the use of a suction pump before processing and retransfusion influence the quality of the erythrocytes? The randomized, controlled, prospective study presented here deals with drainage blood and washed autologous red blood cells (warbc) from 60 patients after hip endoprosthesis surgery. In a comparison between suction pump and redon bottle, the following parameters were studied: haematological-parameter (haemoglobin, haematocrit, erythrocyte count, leukocyte count, thrombocyte count, MCV, MCH, MCHC), vitality (osmotic fragility, 2,3-DPG) and haemolysis parameter (GOT, LDH, plasma haemoglobin, potassium). Control samples were taken immediately after operation: sample one from drainage blood before processing and sample two from warbc before retransfusion. There were no significant statistical differences between the groups. The osmotic fragility of the retransfused red blood cells was slightly above normal values, while the 2,3-DPG was normal. "Old" erythrocytes were haemolysed. The concentration of plasma haemoglobin was clearly above the normal range. In the "redon group" GOT and LDH were clearly increased. The quality of erythrocytes from suction pump reservoirs is not decisively impaired. PMID- 10528417 TI - [Suicidal Tachmalcor poisoning--a case report]. AB - In a case report, a Tachmalcor intoxication with a dose of 18 mg/kg body weight is described. This dose caused a ventricular flutter in the patient which lasted for a total of 10 hours, despite intensive treatment. The treatment began approximately three hours after the intoxication and concentrated on therapy of the ventricular tachycardia. The use of Xylocitin 2%, defibrillation, glucagon and sodium chloride is recommended with such symptoms. Additionally, we used hemoperfusion for drug elimination. Despite the cardiac rhythm disorder of such duration, no neurological deficiencies were observed in the patient. Intoxications caused by these drugs in normal intensive therapies are extremely rare and for this reason treatment can often be very problematic. The following article reports on the successful therapy of one such patient. PMID- 10528418 TI - [Expert systems in medicine: when and for whom? (editoria; comment)]. PMID- 10528419 TI - [Non-insulin-dependent diabetes mellitus associated with nonalcoholic liver cirrhosis: an evaluation of treatment with the intestinal alpha-glucosidase inhibitor acarbose]. AB - Non-insulin-dependent diabetes mellitus not responding to diet only in patients with non-alcoholic liver cirrhosis is characterized by high post-prandial hyperglycemia. The aim of this study was to evaluate the safety and efficacy of 24 weeks of treatment with 300 mg acarbose per day in 76 consecutive outpatients affected by type 2 diabetes and well-compensated liver cirrhosis. The study design was double-blind cross-over vs placebo. All patients tolerated both treatments well, and no significant variations in liver function tests were observed (< 5% vs pre-treatment). A significant reduction of several parameters was observed only after acarbose: fasting glycemia (19 +/- 6 vs 2 +/- 0.5%; p < 0.01), post-prandial glycemia (41 +/- 9 vs 3 +/- 0.6%; p < 0.01), mean glycemia (30 +/- 8 vs 14 +/- 5%; p < 0.01), daily glycemic variation (52 +/- 8 vs 8 +/- 1%; p < 0.01), HbA1c (16 +/- 1 vs 2 +/- 0.5; p < 0.05), incremental area of C peptide after a standard meal (80 +/- 19 vs 200 +/- 36 ng/mL/300 min; p < 0.01). After acarbose a significant increase of intestinal voiding/week (98 vs 28%; p < 0.01) and a parallel reduction of blood ammonia levels (52 +/- 9 vs 9 +/- 5%; p < 0.01) were observed. Results clearly document the good tolerability and the absence of toxic effects of acarbose on the liver, due to a theoretic absence of both absorption by the gut and hepatic metabolism of the drug. In fact, acarbose increases peristaltic movement of the gut, stimulates the proliferation of saccharolytic bacteria and simultaneously reduces proteolytic bacterial proliferation, thus actively reducing blood ammonia levels. These unexpected effects of acarbose may be used to advantage for the treatment of type 2 diabetes mellitus in patients with well-compensated liver cirrhosis. PMID- 10528420 TI - Factor V Leiden in patients with acute coronary syndromes. AB - A common mutation in coagulation factor V gene gives rise to factor V Leiden, which is resistant to the proteolytic activity of activated protein C. This mutation is known to be a major inherited risk factor for venous thrombosis. In order to investigate the possible pathogenetic role of factor V Leiden in coronary thrombosis, we screened patients with acute ischemic heart disease for this genetic mutation. We conducted a case-control study on 114 unrelated patients referred to the Cardiology Department of a University Hospital for coronary angiography. Fifty-three case patients with myocardial infarction or unstable angina were compared with 61 control patients seen for a different coronary syndrome or for stable angina pectoris. The two groups did not differ with respect to the well established risk factors for ischemic heart disease. For each patient genomic DNA was extracted from whole blood by standard techniques; the DNA region carrying the mutation was amplified by polymerase chain reaction, and the allele-specific restriction digestion was used in order to detect the mutation itself. The frequency of factor V Leiden was 0.028 among cases and 0.008 among controls (p = 0.5). According to the data presented, factor V Leiden cannot be considered as a risk factor for acute coronary events. PMID- 10528421 TI - [HEPASCORE: a decision-support system for the identification, clinical staging and functional assessment of hepatopathies]. AB - A decision support system (HEPASCORE) has been developed to optimize the application of objective criteria for qualitative and quantitative assessment of liver function; clinical and laboratory data are automatically processed, and conclusions are explained. Early recognition of abnormal liver states is performed according to a sequential approach, based at first on clinical rules utilizing data from history and physical examination, then confirming or denying the hypothesis by means of selected laboratory tests. Once an abnormal condition is defined, clinical severity can be evaluated by use of suitable scores, either prognostic or focused on major clinical complications. In addition, selected sets of biochemical tests can be used to score one or more functional aspects. Lastly, whenever quantitative estimates of residual liver function are requested, dynamic tests can be applied to measure meaningful parameters such as functioning liver mass and functional hepatic plasma flow. HEPASCORE has been successfully applied to exclude liver abnormalities in subjects at risk, to follow up liver patients, to predict the natural outcomes of severe liver diseases, to foresee the adverse effects of drugs undergoing first-pass liver extraction and the side effects of invasive procedures. While the proposals contained in the system could be further modified for specific needs, they reflect a satisfactory methodological approach, and the program serves as a useful support to decisions regarding the identification and functional evaluation of hepatopathies. The system was developed with Microsoft Access 7.0 and runs on a personal computer under Windows 95. PMID- 10528422 TI - [Disorders of eating behavior: recent research and controversial results]. AB - The aim of this review is to provide a comprehensive profile of research developments in the field of eating disorders. Eating disorders may be defined as a multidisciplinary field, not only because diverse specialist disciplines are required for treatment, but also because the need to establish ad hoc Eating Disorders Units comprised of psychiatrists, internists, dietologists and endocrinologists has become evident. New research data, collected by use of informatics-based systems, are presented. PMID- 10528423 TI - Magnesium and cardiovascular drugs: interactions and therapeutic role. AB - Numerous experimental, epidemiological and clinical studies have pointed out a relevant role for magnesium deficiency in the development of many cardiovascular diseases. Some pharmacological treatments may interfere with magnesium turnover, and magnesium deficiency may alter the pharmacokinetics and pharmacodynamics of some cardiovascular drugs. Loop and thiazide-like diuretics increase magnesiuresis, and total bodily magnesium deficiency may appear during prolonged treatment with diuretically active doses of these drugs. The potassium retaining agents, such as amiloride, triamterene and spironolactone, tend to retain magnesium but they are not magnesium-retaining substances to the extent to which they are potassium-retaining diuretics. The interaction between magnesium and digitalis is complex. Magnesium, acting as an indirect antagonist of digoxin at the sarcolemma Na(+)-K(+)-ATPase pump, reduces cardiac arrhythmias due to digoxin poisoning. Recent controlled studies have shown that treatment with magnesium significantly reduces the frequency and complexity of ventricular arrhythmias in digoxin-treated patients with congestive heart failure without digoxin toxicity. Magnesium improves the efficacy of digoxin in slowing the ventricular response in atrial fibrillation. Digoxin reduces tubular magnesium reabsorption, and in patients with congestive heart failure this interaction may be cumulative with other causes of magnesium deficiency (diuretics, diet, poor intestinal absorption). The complex and potentially life-threatening interactions between magnesium and some cardiovascular drugs suggest that magnesium status should be carefully monitored in patients receiving such drugs. Therapy with magnesium is rapidly acting, has a safe toxic-therapeutic ratio, is easy to administer and titrate. The correction of magnesium deficit should therefore always be considered for patients with cardiopathy. PMID- 10528424 TI - [The antiphospholipid syndrome during chronic lymphatic leukemia. An association with anti-factor VIII antibodies]. AB - Activated partial thromboplastin time may be prolonged as the result of either of two different autoimmune complications of chronic lymphocytic leukemia: the development of antiphospholipid antibodies, such as lupus anticoagulant or anticardiolipin antibodies, or anti-factor VIII inhibitors, such as acquired hemophilia A. In the rare simultaneous occurrence of both inhibitors, differential diagnosis of a prolonged activated partial thromboplastin time poses a number of problems during laboratory work-up, due to mutual interference of the commonly performed tests. Only careful clinical follow-up can disclose the significance of the laboratory findings. We report the case of concurrent antiphospholipid antibodies (lupus anticoagulant positivity, anticardiolipin antibodies; IgM 3880 MPL/mL and IgG 265 GPL/mL) and anti-factor VIII antibodies (46.8 Bethesda Units) in a patient with chronic B-cell lymphocytic leukemia who had prolonged activated partial thromboplastin time (78.8 s). The relationship between lymphoproliferative and antiphospholipid syndrome, laboratory work-up in the case of the association of antiphospholipid and anti-factor VIII antibodies, and related problems that occur during clinical management of the patient are also discussed. PMID- 10528425 TI - [The scombroid syndrome, a potentially serious ichthyotoxicosis of uncertain pathogenesis: personal experience]. AB - The aim of this report is to point out the potential seriousness of the scombroid syndrome which, on the basis of our experience, can be characterized by extremely serious symptoms. We describe 12 cases of scombroid syndrome: two-thirds of the patients presented with rapid worsening of their clinical condition and hypotension severe enough to require use of plasma-expanders and hospitalization in an Internal Medicine Department. In the youngest patient, hypotension and symptoms were so marked that intravenous administration of epinephrine, and hospitalization in the Intensive Care Unit were required. Thus, in contrast to reports in the literature, the scombroid syndrome should be considered as a potentially serious ichthyotoxicosis. The pathogenetic role played by histamine, poorly absorbed by the intestine and rapidly metabolized by the liver, should be reevaluated. The potential onset of serious clinical symptoms warrants prolonged observation of the patient in an environment equipped to deal with the not infrequent emergencies that can arise, even in young and healthy subjects. PMID- 10528426 TI - Hypoparathyroidism secondary to Riedel's thyroiditis. A case report and a review of the literature. AB - Riedel's thyroiditis is a rare condition in which the thyroid gland is replaced by fibrous tissue. Fibrosis in various distant sites is a possible concomitant event. We report a case of Riedel's thyroiditis complicated by mediastinal fibrosis, a tumefactive fibro-inflammatory lesion of the neck and primary hypothyroidism. A review of the literature in which only 8 previous cases of hypoparathyroidism secondary to Riedel's thyroiditis have been recounted concludes the report. PMID- 10528427 TI - Localized AL amyloidosis of the colon and clinical features of intestinal obstruction. A case report. AB - The term amyloidosis refers to a group of disorders characterized by the extracellular accumulation of insoluble, fibrillar proteins (i.e. amyloid) in various organs and apparatuses. Local deposition of amyloid without systemic involvement is a rather uncommon form that gives rise to an amyloid pseudo-tumor or "amyloidoma". This paper describes the clinical features of an intestinal subocclusion observed in an elderly patient with localized primary amyloidosis of the transverse colon. The endoscopic picture indicated a stenosing neoplasm. Segmentary colectomy, however, followed by histological examination (characteristic green color in polarized light after staining with Congo red) and immunohistochemical analysis of the resected tissue, revealed massive deposits of amyloid composed of lambda light chains in the interstitial connective, and perivascular tissue and muscular tunic, and resulted in a diagnosis of AL amyloidosis. This was successfully treated with a combination of melphalan and prednisone. PMID- 10528428 TI - [Therapeutic monitoring of drugs]. PMID- 10528429 TI - [35th Congress of the American Society of Clinical Oncology, Atlanta, USA, 15-18 May 1999. Non-small-cell lung tumors]. PMID- 10528430 TI - [35th Congress of the American Society of Clinical Oncology, Atlanta, USA, 15-18 May 1999. Neoplasms of the ovary and gastrointestinal system]. PMID- 10528431 TI - [IL-6, p53 and proto-oncogene c-myc play different roles as biological markers of plasma cell dyscrasias]. AB - PURPOSE: To determine the role of serum levels of IL-6 and p53 mutant protein as well as of c-myc proto-oncogene alterations: a) in discriminating between benign (MGUS) and malignant Plasma cell dyscrasias (Multiple and Microsecreting Myeloma, Plasmocytoma); b) in monitoring the clinical course of malignant forms of this disease. PATIENTS AND METHODS: Eighty-eight patients affected by Plasma cell dyscrasias (58 MGUS, 24 MM and 6 PLC) entered this study. Using commercially available ELISA kits, serum levels of IL-6 and p53 have been determined in all the patients. In addition, a selected group of patients (n = 30) was also analyzed for structural c-myc gene alterations by Southern blot technique. RESULTS: The results show that, conversely from p53 protein, IL-6 and c-myc gene may represent useful diagnostic markers for discriminating benign from malignant forms of Plasma cell dyscrasia. On the contrary, preliminary findings of the same work indicate a potential role for the mutant p53 protein in monitoring the response to chemotherapy of patients affected by MM or PLM. CONCLUSIONS: Overall, these data suggest that the combined use of IL-6, p53 and c-myc may provide a new approach for a more rational management of Plasma cell dyscrasia patients. PMID- 10528432 TI - [Iatrogenic sequelae of pulmonary tuberculosis]. AB - OBJECTIVES: The purpose of this study is to underline how topical is the chapter of the sequelae of pulmonary tuberculosis and to try to make a classification. Pulmonary tuberculosis can be cured definitely or hesitate in disease (BK negative) that is totally independent from tuberculosis about their pathogenesis and clinical features. They are called sequelae. MATERIALS AND METHODS: We made a statistical analysis that investigate a group of 110 patients without active infection (BK negative) admitted in the hospital because of a sequela of pulmonary tuberculosis. Patients were treated in the past by collapse-therapy or by antibiotic-therapy until their spittle became negative for BK. RESULTS: A significant (p < 0.05) relationship between each kind of sequela, among the most important ones (fibrothorax, interstitial fibrosis, bronchiectasis, empyema with or without pleural fistula, parafibrotic emphysema), and type of treatment, results. CONCLUSIONS: The sequelae of tuberculosis of the lung are highly disadvantageous for people who are affected; are observed frequently; are closely dependent on what kind of treatment the patient has received; are classified in iatrogenic, not iatrogenic or mixed. PMID- 10528433 TI - Pharmacokinetic of intraarterial mitomycin C with extra corporeal detoxification in humans. AB - PURPOSE: A study has been performed in 15 patients with liver metastases from colorectal cancer to evaluate the pharmacokinetic of mitomycin C and the effectiveness of drug removal techniques during high-dose locoregional chemotherapy. PATIENTS AND METHODS: Haemofiltration and/or haemodialysis of post hepatic venous blood were performed during 22 intra-arterial infusions of mitomycin C by the use of a double lumen catheter surgically introduced in the inferior vena cava. Mitomycin C levels were measured by high performance liquid chromatography in the extracorporeal circuit blood, in the peripheral venous blood and ultrafiltrate. RESULTS: The mean reduction of mitomycin C bioavailability in the extracorporeal circuit blood was 42.3 per cent using haemofiltration, 58.9 per cent using haemofiltration plus haemodialysis and 59.3 per cent with haemodialysis alone. The resulting mean mitomycin C plasmatic half life was 40.2 minutes using haemofiltration, 24.9 minutes using haemofiltration plus haemodialysis and 24.1 minutes using haemodialysis alone. CONCLUSIONS: The detoxification of posthepatic venous blood during intra-arterial hepatic chemotherapy is an effective method to increase mitomycin C concentrations and oncological response and limit extra hepatic toxicity. PMID- 10528434 TI - [An observational and longitudinal study on patients with kidney stones treated with Fiuggi mineral water]. AB - PURPOSE: To evaluate the development of clinical conditions particularly the possible expulsion of renal stones, in patients with nephrolithiasis this treated with Fiuggi Mineral Water. PATIENTS AND METHODS: This study was carried out in 614 patients, by administering them an appropriate questionnaire 6 month after drinking Fiuggi Mineral Water. RESULTS: The percentage of patients eliminating one or more renal stones was higher in those who drank mineral water at home (55.9%) compared to those who just drank water in Fiuggi (38.5%). Renal stones were eliminated more frequently in case of consecutive treatments than in case of cyclic treatments (54.3% vs 44.4%). Stone elimination was also related to quantity of the water which was daily drunk (up to 1 L/day, 52%; > 2 L/day, 72%). In 29% of the patient a stone relapse after operation or lithotripsy or both was observed. CONCLUSIONS: These data in agreement with those of other investigations show that surgical treatment or lithotripsy are not able to avoid stone relapse. Other treatments, such as mineral water drinking, may be useful in this situation. PMID- 10528435 TI - [Helicobacter pylori and gastric cancer]. AB - Several unresolved issues still cast doubts on the epidemiological data which point to an association between infection from Helicobacter pylori (Hp) and gastric cancer. These are: a) the male/female ratio of gastric cancer ranges from 4 to 1.5 in all studies, whereas the prevalence of Hp infection is the same in both sexes; b) the prevalence of Hp infection is as high as 90% in several developing countries where the frequency of gastric cancer is very low; c) the acquisition of the infection at a young age, considered very important with regard to the risk for cancer, varies from 4.2% to 83% in several countries in which the mortality for stomach cancer is, on the average, 10/100,000; d) the incidence of cancer in patients with a duodenal ulcer is half of that of the general population but Hp infects up to 100% of these patients. In the sequence of events that lead to gastric cancer, Hp appears to play a role only in the very initial steps, as a causative agent of chronic inflammation. The further events which lead to cancer are multifactorial, involving environmental agents and the host response. It is therefore inappropriate to consider Hp a direct carcinogen for humans. This also applies to specific strains of the bacterium such as the ones expressing the cagA gene. In a study we conducted in an area with a low incidence of gastric cancer (Latina), the prevalence of Hp infection was equal to 78.6% and, among the positives, 81% resulted cagA positive. This data, if compared with a similar research that took place in another area with a high incidence of gastric cancer (San Marino) where the prevalence of Hp infection was 51% and cagA 69%, further demonstrates the inconsistency of associating Hp and cancer of the stomach. PMID- 10528436 TI - [Prophylaxis of toxic effects on the peripheral venous system associated with intravenous administration of vinorelbine]. AB - Vinorelbine is effective in the treatment of a number of malignancies. However, beside the haematologic and not haematologic toxicity as thrombocytopenia, granulocytopenia, fatigue, paresthesias, nausea and vomiting, one of the most common side effects is the local irritation with thrombophlebitis. The side effect, reported in about 10-26% of patients receiving vinorelbine infusions, is due to the vescicant and irritant action of the drug. Many studies have been performed in order to reduce the incidence of venous irritation either with concomitant administration of anti-inflammatory drugs as defibrotide or ketorolac, or changing infusion schedule from bolus infusion to a 20-30 minute infusion. Aim of this review is to define peripheral venous system toxicity of vinorelbine and the best way of administration. PMID- 10528438 TI - [A case of atrial myxoma with connectivitis-like onset]. AB - A case of cardiac myxoma is reported. This tumor, although histologically benign, may be potentially lethal if an early diagnosis and an adequate surgical treatment is not performed. In an early stage, the disease may be asymptomatic or a large array of aspecific symptoms may be present. In this case, an initial diagnosis of polymyositis was made. This paraneoplastic syndrome is frequently associated with tumors, and should trigger a full evaluation for cancer. PMID- 10528437 TI - The role of granulocyte growth factors in the treatment of non small cell lung cancer. AB - In the past years granulocyte growth factors have been introduced in clinical practice. Their use is intended to reduce the risk of infection related to chemotherapy and to increase the dose-intensity of chemotherapy agents. Very few randomized trials have been reported in advanced non small cell lung cancer on chemotherapy plus or minus granulocyte-macrophage colony-stimulating factor or granulocyte colony-stimulating factor. No benefit for granulocyte growth factors use was observed in terms of response rate and survival. Recently, several investigators used growth factors to support new promising drug combinations including vinorelbine, gemcitabine, taxol or taxotere. However, outside controlled clinical trials the role of granulocyte growth factors in the treatment of non small cell lung cancer should be within the American Society of Clinical Oncology guidelines. PMID- 10528439 TI - [Development of cancer chemotherapy. Discovery of new active drugs. IV]. PMID- 10528440 TI - [Surveillance of mosquito-borne viruses in Breclav after the flood of 1997]. AB - In July 1997, devastating floods occurred after heavy rains in Moravia, Czech Republic. Mosquito populations increased abruptly in the flooded area thereafter. We carried out a surveillance for mosquito-borne virus infections in the Breclav area, South Moravia, including serosurveys of inhabitants. A total of 11,334 female mosquitoes in 117 pools (9,100 Aedes vexans, 917 A. cinereus, 11 A. cantans, 1,074 A. sticticus and 232 Culex pipiens pipiens) were examined by virological methods. Seven virus isolates were obtained: six of them Tahyna virus (Aedes vexans 5, A. cinereus 1), while one was West Nile (WN) virus (Culex p. pipiens--first isolation in the Czech Republic). Sera of 619 local inhabitants were examined in plaque-reduction neutralization test, and antibodies to Tahyna virus were detected in 333 (53.8%) and to WN virus in 13 (2.1%). In 72 individuals, paired sera were sampled: a significant increase of antibody titre was detected once against Tahyna virus (a subclinical infection) and 4 times against WN virus (two children had an illness compatible with WN fever). Of the nine remaining WN seroreactors, three other revealed clinical symptoms compatible with WN fever in summer 1997. The data indicate WN virus activity in the Breclav area, and describe the first cases of WN fever in Central Europe. The WN virus should not be underestimated as a potential agent of local epidemics even in the temperate climate of Central Europe. Environmental factors including human activities which enhance vector population densities (heavy rains followed by floods, irrigation, higher temperature) can produce an increased incidence of mosquito-borne diseases, including WN fever. PMID- 10528441 TI - [A tularemia epidemic in western Slovakia 1995-1996]. AB - The activation of natural foci of tularaemia in West Slovakia during 1994-1996 led to an epidemic outbreak in 1995-1996--mean annual morbidity 6.2 per 10(5) population. In comparison with the mean annual morbidity rate in the preceding period (1980-1994), a more than sevenfold increase was recorded. Of 213 notified cases of the disease 156 cases occurred in 1995--morbidity 8.9 per 10(5) population--being the highest morbidity recorded in this endemic region since the period of epidemic occurrence in the 60s. The highest proportion of cases (59.2%) was recorded in the districts of Nitra and Nove Zamky. The activation of natural foci along with changing social conditions, caused also marked changes of some epidemiological characteristics of tularaemia in Slovakia, such as seasonal and professional occurrence, as well as clinical forms of the disease. The impaired epidemiological situation in the occurrence of tularaemia is pointing to the importance of systematic surveillance to improve the diagnosis of the disease and ensure effective preventive measures. PMID- 10528442 TI - [Toxoplasma gondii antibodies in pregnant women in the Ceske Budejovice District]. AB - In 1984-1986 in the district of Ceske Budejovice in the southern part of the Czech Republic pregnant women were subjected to serological examinations for antibodies against Toxoplasma gondii. A total of 3,392 women were examined within the age bracket 16-54 years. For the serological examination parallel examinations were made using Sabin-Feldman test (SFT) and the complement fixation test (CFT). In SFT the basic serum dilution was 1:4, in CFT 1:10. Pregnant women were examined once or repeatedly. During the first examination which was usually made between the second and third month of pregnancy the total (SFT and CFT) prevalence was 37%. In the SFT antibodies were detected in 35%, in CFT in 25%. The second blood sample was taken during the 4th-5th month of pregnancy and subsequent samples during the 8th-9th month of pregnancy. In women who were examined twice or repeatedly (a total of 1,409 women), the dynamics of prevalence of antibodies were recorded. 64% women were permanently negative, 33% women were permanently positive with the same or a slightly varying titre and in 3% women during pregnancy, seroconversion was observed or a significant rise of antibodies. In 20 women where seroconversion or a significant rise of antibodies was found data were collected to find out whether in their children from birth to the age of 12-13 years toxoplasmosis was diagnosed. The toxoplasmosis was not diagnosed in these children. PMID- 10528443 TI - [Study of leukocyte phagocytic activity using flow cytometry and hydrophilic fluorochrome-labeled microspheric particles]. AB - The phagocytic activity was determined in human polymorphonuclear leucocytes using the flow cytometry technique with microspheric hydrophilic particles labelled with fluorescein isothiocyanate. Non-specific adherence of particles is discussed and normal values of phagocytic activity are presented. The advantages of this method include simplified sample preparation using whole blood, rapidity of the test, precise phagocytosis quantification and possibility of archiving all findings in a computer. PMID- 10528444 TI - [Properties of Salmonella isolates in the Czech Republic]. AB - Based on a grant project "Use and importance of epidemiological markers in Salmonella enteritidis and Salmonella typhimurium in the spread of salmonelloses in children under two years of age" implemented in 1995 to 1997, the authors investigated epidemiological markers in 1,186 salmonella isolates; the strains were isolated from faeces of 838 sick children, from 266 faeces of their contacts, from 49 specimens of incriminated foods and from 33 smears from the children's environment. Of 1,186 Salmonella isolates 999 were strains of S. enteritidis, 39 strains of S. typhimurium and 148 strains were not identified. The markers of Salmonella isolates were investigated from the aspect of biotyping -98% S. enteritidis were formed by the biovar Jena. 2% by biovar Essen; sensitivity to antibiotics--94.5% Salmonella strains were sensitive to 12 selected antibiotics, 2.9% were resistant and in 2.6% the resistance was in the intermediate zone; phagotyping--in 808 strains of S. enteritidis PT 8--88% predominated, in S. typhimurium DT 104 and DT 141; assessment of plasmid profiles -in strains of S. enteritidis plasmid 55 kb predominated, in three strains of S. typhimurium a plasmid size 95 kb; virulence--was compared in 43 strains isolated from hospitalized children with a severe clinical course with 39 strains from children treated at home. In vitro tests revealed that hospitalization of affected children was associated with virulence of the strains (SE phagotype 8) and not with age. The presented results are discussed with regard to the epidemiological situation in the Czech Republic and in the world. PMID- 10528445 TI - [Elastase activity of gram-negative non-fermenting bacteria and two variations of a simple method for their detection]. AB - The ability to produce elastase was examined in 1,168 strains of 11 representative genera of Gram-negative non-fermenting bacteria isolated from weakened patients in the course of eight years. Detection of the elastase activity was performed parallelly using the conventional method and its pulverisation variant. The difference between the results accomplished by the classical method and its pulverisation variant (53.9% versus 60.5% detected elastase-positive strains) was highly significant (p < 0.05). Among the examined strains 500 (42.8%) displayed elastase activity. The highest proportion of the elastase positive strains was found in Pseudomonas aeruginosa (84.5%), P. fluorescens (60.5%), P. stutzeri (56.4%), Burkholderia cepacia (43.8%) and Shewanella putrefaciens (39.7%). Elastase activity detected in Burkholderia cepacia, Pseudomonas alcaligenes, P. mendocina, P. putida, P. stutzeri and in Shewanella putrefaciens is apparently described for the first time. PMID- 10528446 TI - [The velvet revolution in health care]. PMID- 10528447 TI - Ethics and politics of research. PMID- 10528448 TI - Clarifying the concept of normalization. AB - PURPOSE: To refine and develop the concept of normalization. While firsthand accounts, clinical observation, and numerous studies suggest that parents of children with chronic conditions often strive to lead a normal family life, the distinguishing characteristics of normalization need to be understood before evaluating the feasibility and consequences. Conceptualizing these efforts as normalization, researchers have identified cognitive and behavioral strategies used by parents to normalize family life. SOURCES: A total of 33 articles representing 14 studies were selected through computer-assisted searches of the topic from 1966-1997, hand searches of nursing journals from 1970-1997, and analysis of reference lists. Key words in the searches included: adaptation, psychological; chronic disease; disabled persons; family; child; adolescence; parent-child relations; models, psychological; and nursing theory. Articles (N = 19) that applied and expanded the concept comprised the sample used in the analysis. A normalization construct was used. METHODS: Attributes for normalization were inductively derived based on the most recent methods for refining and developing concepts, with special attention to how family and illness affect manifestations of normalization. FINDINGS: The attributes identified in 1986 remained relevant, but required revision and expansion to reflect a contemporary understanding of normalization. Unique manifestations of normalization were identified within certain illness and family contexts. CONCLUSIONS: Knowledge synthesized from previous studies can enhance how normalization is used in future qualitative and quantitative research and in theory development. Findings indicate the need for researchers to build on the current state of knowledge and continue to further develop the concept. Understanding the findings can also sensitize clinicians to the complex process of normalizing when a child has a chronic condition. PMID- 10528449 TI - Clarification and integration of similar quality of life concepts. AB - PURPOSE: To identify major problems that obscure understanding of quality of life; to differentiate quality of life from other closely related concepts, and to offer a definition of quality of life. Quality of life is a term frequently used, but seldom defined, in nursing research. Multiple interpretations and measures make evaluating research difficult. The need to clarify concepts, develop nursing theory, and enhance communication is significant. Concept clarification is also essential for research instrument development and evaluation. ORGANIZING CONSTRUCT AND SCOPE: Concept analysis as proposed by Walker and Avant. Preconceived theories were avoided to keep the analysis as unbiased as possible. The analyst considered many descriptions of quality of life, but eventually included only those in the scientific literature. SOURCES: CINAHL, Medline, Psych-Info, ERIC, Social Science Abstracts, and reference lists from published articles in nursing, medicine, psychology, and sociology were used. Twenty-seven theory-based articles and book chapters published since 1989 and a convenience sample of 88 research articles published in 1997 were used. This sample was selected from 16,480 articles published 1993 to 1998. METHODS: Concept clarification comparisons to identify the major attributes of several closely related concepts, including well-being, satisfaction with life, and functional status. Major attributes were validated by a panel of cohorts who met weekly for five consecutive weeks for the purpose of analyzing various concepts. FINDINGS: Quality of life is comprised of subjective indicators such as well being and satisfaction with life and objective indicators, such as functional status. Continued dialog and research is necessary to distinguish between the concepts of health and quality of life. CONCLUSIONS: A model showing the relationships among quality of life, well-being, functional status, and satisfaction with life is provided and a definition of quality of life is offered. Researchers and authors should include the definition of quality of life used in their work. Instruments should reflect the theoretic definition used. Care should be taken to clarify if functional status, well-being, satisfaction with life are the concepts under consideration as reflecting a part of overall quality of life. PMID- 10528450 TI - Depression, social support, and quality of life in older adults with osteoarthritis. AB - PURPOSE: To develop an understanding of the quality of life of older adults with osteoarthritis (OA) with varying levels of depression and social support as a basis for nursing interventions. Osteoarthritis in the United States is the number one chronic disease in late life and the major cause of disability in older adults. In addition to the functional disability and economic effect of OA, older people with this disease experience suffering, depression, and diminished quality of life. DESIGN: For this cross-sectional survey, a convenience sample of 50 older adults with OA was recruited from two U.S. hospital-based arthritis clinics in northern Ohio for 3-months during 1995. METHODS: During face-to-face interviews, the Arthritis Impact Scales, Center for Epidemiological Studies Depression Scale, Social Support Questionnaire, and Quality of Life Survey, were used to measure osteoarthritis severity, depression, informal social support, and quality of life. FINDINGS: Although few formal social support services were used, high levels of satisfaction from the subjects' large informal networks of family and friends were reported. In addition, satisfaction with subjects' quality of life was extremely high despite depression, co-morbid conditions, pain, and functional limitation. CONCLUSIONS: Social support appeared to play an important role in moderating the effects of pain, functional limitation, and depression on these subjects' quality of life. Nurses who work with older adults are in a unique position to help them adjust to living with osteoarthritis by providing them the support needed to help them manage their disease. PMID- 10528451 TI - The rhythm of relating in a paradigm of wholeness. AB - The importance of relating to the rhythm of another person's interactive pattern is integral to helping people move through illness and disruptive events. At times of chaos, a person's rhythm may be irregular and difficult to sense, but transformation to higher levels of organization often occurs at far-from equilibrium states. Nurses should develop a tolerance for ambiguity and uncertainty and "hang in there" with clients until a new rhythm emerges. PMID- 10528452 TI - A model of chronic dyspnea. AB - PURPOSE: To present an overview of dyspnea, differentiate chronic dyspnea from acute dyspnea, critique models of dyspnea found in the nursing literature, and propose a new model of chronic dyspnea to guide the care and evaluation of chronic dyspnea in patients living with chronic obstructive pulmonary disease (COPD). Dyspnea is the major symptom that impairs quality of life for nearly 16 million Americans who have COPD. METHODS AND SOURCES: Review of scholarly literature on dyspnea by searching CINAHL and MEDLINE (1980-1998) using dyspnea and chronic obstructive lung disease as key words. The search produced studies conducted by a variety of health care professions including those in nursing, medicine, exercise physiology, and respiratory therapy. FINDINGS: The existing models fail to differentiate between acute and chronic dyspnea. These models were found to be inadequate for guiding interventions to decrease the long-term adverse consequences of chronic dyspnea. CONCLUSIONS: A useful model of chronic dyspnea defines chronic dyspnea as distress with varying levels of intensity and long-term physical, psychologic, and sociocultural consequences. The proposed model has implications for both research and clinical practice by identifying the consequences of chronic dyspnea as outcome measures of the effectiveness of treatment. PMID- 10528453 TI - Testing a model of avoiding environmental tobacco smoke in young adults. AB - PURPOSE: To test an explanatory model of gender, self-efficacy, situational influences, and other health-promoting behaviors on the avoidance of environmental tobacco smoke (ETS) in young adults. ETS is a cause of lung cancer, pulmonary disease, and cardiovascular disease. Although young adults are at increased risk for ETS exposure, there are few behavioral studies of ETS exposure and no reported studies of ETS exposure in young adults. Although college students are often exposed to ETS, the college environment offers a setting in which the opportunities for change are substantial. DESIGN: Model-testing. Data are reported on a convenience sample of 136 nonsmoking 18- to 25-year old students in one mid-atlantic U.S. university. This sample of nonsmokers was drawn from a larger sample of 241 smokers and nonsmokers in 1995. Model constructs were based on Pender's health promotion model (HPM). METHODS: The General Self efficacy Scale, Health Promotion Lifestyle Profile, and ETS Avoidance Scale were used along with items measuring ETS-avoidance efficacy and Living with Smoke. Path analysis was used to test the model. FINDINGS: The trimmed explanatory model showed that 26% of the variance in avoiding ETS was accounted for by gender, having self-efficacy, and ETS-avoidance efficacy, not living with people who smoke, and performing other healthy behaviors. Being female and general self efficacy indirectly influenced ETS avoidance through their effects on related health promoting behaviors. CONCLUSIONS: The explanatory model of ETS avoidance can provide useful information for the development of interventions to prevent exposure to passive smoke. Given the occurrence of ETS exposure in young adults, longitudinal research using this explanatory model is yielding promising results. Enhancing the self-efficacy of young adults and encouraging healthy lifestyle behaviors may be an important factor in their avoidance of ETS. PMID- 10528455 TI - Reachable moment. PMID- 10528454 TI - Adolescent resilience. AB - PURPOSE: To explore what resilience means to adolescents and whether the Resiliency Scale can accurately measure resilience. Researchers have identified that resilience in children and adolescents may lead to psychosocial maladaption and psychopathology in adulthood. Further research is necessary in order to understand what resilience is in adolescence and to identify those at risk for psychosocial problems. DESIGN: A triangulated research design was used to explore the concept of resilience in adolescents. This small pilot study, conducted in 1997, had a purposive sample of 51 10th-grade and 11th-grade volunteers from one inner-city, vocational high school in New England. METHODS: A researcher developed demographic tool was used to explore the environment of adversities to which students were exposed. Wagnild and Young's Resiliency Scale was used to measure adolescents' perceptions of their resilience. Focus groups, structured and unstructured interviews, and written stories were used to gather phenomenologic data. FINDINGS: Despite the traumatic and violent world in which participants lived, the adolescents ranked themselves as "resilient." These adolescents believed that being resilient was to be (a) disconnected from others because they could not trust, (b) isolated because they had inadequate or no support systems, and (c) insulated because the emotional pain was too much to bear. CONCLUSIONS: This study of resilience in adolescence shows the need for further research; the investigators question whether resilience is really a healthy state, and wonder if similar interventions are necessary for both "resilient" and "vulnerable" adolescents. PMID- 10528456 TI - Developing a regional nursing research website. PMID- 10528457 TI - Victor Fuchs on health care, ethics, and the role of nurses. Interview by Joanne Spetz. PMID- 10528458 TI - Ethics, policy, and practice: interview with Emily Friedman. Interview by Carolyn Brown. PMID- 10528459 TI - Ethical and policy considerations for centenarians--the oldest old. AB - PURPOSE: To illustrate the incongruence of ethical standards and fiscal and policy constraints on quality care for the oldest old. As the fastest growing demographic segment in the United States, care needs of the oldest old are a special challenge to the health care system. DESIGN: Narrative analysis of interviews with centenarians who used nursing home services. The sample was three participants of the Georgia Centenarian Study who had been community dwelling and cognitively intact at the onset of participation (between 1988 and 1997). Interviews were conducted in nursing homes or after discharge. METHODS: Case histories were constructed from interviews in 1997 to improve understanding of quality of care. FINDINGS: Less-than-optimal care was provided for these elders, and little consideration was given to their input to care decisions and prospects for medical improvement. Appropriate consideration was not given to providing least-restrictive environments, appropriate restraint use, and options for community care. CONCLUSIONS: Six policy reforms are suggested for meeting the needs of the oldest-old before and after institutionalization. These include: integration of resident involvement in care decisions; development of alternate models of care; greater input from nurses concerning nursing care of special populations; more effective family and community involvement in the caring of elderly populations; increased research to promote function and independence; and increased education of personnel and nursing students to allow for more accurate assessment of cognitive and physical status. PMID- 10528460 TI - Physicians' neonatal resuscitation of extremely low-birth-weight preterm infants. AB - PURPOSE: To examine the perceptions of physicians who make delivery room decisions to resuscitate extremely low-birth-weight (ELBW) neonates at marginal viability. Nurses, parents, economists, and ethicists have questioned resuscitation of ELBW neonates, many of whom experience high levels of morbidity and mortality. Yet no systematic studies were found that addressed physicians' perceptions and delivery room decisions. DESIGN: Descriptive, using naturalistic inquiry. A national U.S. convenience sample was obtained in 1996-1997 of 54 physicians in five perinatal subspecialties who resuscitated ELBW neonates. METHODS: Tape-recorded and transcribed interviews were analyzed using NUD*IST software and line-by-line constant comparison. FINDINGS: Despite awareness of the high morbidity and mortality, 96% of the physicians offered resuscitation to all ELBW neonates in the delivery room. The main factors affecting their decisions were "the role of physician;" having been "trained to save lives;" the belief that "if called, I resuscitate;" the inability to determine gestational age; requests from parents to "do everything;" and the need to move from a "chaotic" delivery room to a controlled neonatal intensive care unit. Six major themes were: role expectation, uncertainty, awareness, internal and external forces, burden, and continuing quandaries. Physicians were burdened by the devastated and dying babies, by their inability to predict which neonates had a chance for intact survival, and by conflicts with colleagues about viability. Statistical probability of survival, legal constraints, and cost of care did not appear to affect greatly their decisions. Physicians asked that society and national policy makers set parameters for resuscitation. CONCLUSIONS: The American Academy of Pediatrics' Neonatal Resuscitation Protocol needs revision to delineate the ethical criteria for resuscitation. Early prenatal education for families which clearly teaches the margins of viability and outcomes of early deliveries is also recommended. Physicians must be supported in changing the recessitation paradigm. PMID- 10528461 TI - Lucian Leape on the causes and prevention of errors and adverse events in health care. Interview by Peter I. Buerhaus. PMID- 10528462 TI - Language plus for international graduate students in nursing. AB - PURPOSE: To provide information about an English-language support program that focuses on the needs of international graduate nursing students. The growing presence of these students coincides with the increasing numbers of universities committed to world health. Crucial social and language competence affect the success and progress of international students in graduate nursing programs. DESIGN: Reviewed literature was 1980 to 1998, in nursing and applied linguistic research including second-language acquisition, phonology, discourse analysis, and language pragmatics to identify social and language phenomena. FINDINGS: Investigators suggest essential elements such as conventions of academic writing, reading comprehension, vocabulary, and pronunciation skills be included in the supportive Language Plus program. CONCLUSIONS: Ongoing development of the Language Plus program can promote collaboration between nurses and linguists and increase the success of international graduate nursing students. PMID- 10528463 TI - Community development specialists in nursing for developing countries. PMID- 10528464 TI - Focus on nurse midwives. PMID- 10528465 TI - The SCAR Working Group on Human Biology and Medicine: 25 years on. PMID- 10528466 TI - Anthropometric characteristics of men in Antarctica. AB - Thirty anthropometric and ten physiological parameters were evaluated over a 10 month period during 1985-86 in 66 polar explorers at an Antarctic station (Mirny observatory), all of them males aged 25-61 years. The evaluations were made in the months of April, September and January, which corresponded to the following Antarctic seasons: the beginning of the polar night, an intermediate period, and the beginning of the polar day; the necessary measurements were performed on subjects belonging to three occupational groups, namely: administrative, scientific, and manual workers. Significant changes in the pattern of skinfold thickness were observed using ANOVA with repeated measurements during the winter period (p < 0.05). Despite the fact that body weight and BMI of subjects remained unchanged, the mean sum of skinfold thickness and subcutaneous fat mass increased over the studied period at the expense of muscle mass. In participants engaged in high levels of outdoor physical activity (e.g. construction workers, drivers, technicians), an increase in fat mass, significant fall in muscle mass on wrist dynamometry, and protracted time of the simple motor response time was documented. Systolic blood pressure showed a downward trend during the winter in the group of manual workers, while significant rises in the diastolic pressure (p < 0.05) were found in the group of scientists at the end of the polar night. The present findings may be interpreted as evidence for destabilization in the studied individuals, and for an adaptation response to the Antarctic environment, which results in apparent increase in body fat and decrease in muscle mass. PMID- 10528468 TI - Psychological health, physical development and autonomic nervous system (ANS) activity in Siberian adolescents. AB - Cultural and biological influences on psychological health were examined in 256 schoolchildren, aged 13 to 17 years, in Novosibirsk, Russia. Children's competence and problems were assessed using the Youth Self-Report (YSR), the Child Behavior Checklist (CBCL) and the Teacher's Report Form (TRF). Compared with Americans, Siberian children scored lower on competencies and higher on somatic complaints. Height predicted mothers' ratings for children's Total Competence and Delinquent Behaviour and was negatively related to boys' Anxious/Depressed problems. Triceps skinfold thickness predicted children's self ratings for somatic complaints. In girls, self-ratings for Delinquent Behaviour were related to heart period variability and subscapular skinfold thickness and self-ratings for Attention Problems were related to triceps skinfold thickness. Systolic blood pressure was negatively associated with behavioural problems, thought problems and somatic complaints. Overall, the findings suggest culture bound and demographic influences on competence and problems in Siberian adolescents and variation related to physical development and autonomic nervous system activity. PMID- 10528467 TI - Changes in the serum lipid profile in man during 24 months of arctic residence. AB - The influence of the severe climate and geographical conditions at the Svalbard archipelago (78-79 degrees N) on serum lipid levels were measured in Caucasian miners who had arrived from the southern part of Ukraine and Russia (48 degrees N). The persons included in the study were randomly divided in five groups according to their time of living (1, 3, 6, 12 and 24 months) at Svalbard. Blood sampling took place during a two week period in January, when the Svalbard archipelago is into its polar night. General elevated levels of triglycerides were found in group I-III (1, 3 and 6 months stay), whereas the values measured in group IV and V (12 and 24 months stay) were somewhat lower. This apparent decline in triglycerides was paralelled by generally elevated levels of HDL cholesterol. The serum level of phospholipids was similar in all groups. All the level of free fatty acids was apparently higher in groups IV and V, particularly 18:3 and 16:1. These results may be indicative of a rise in triglyceride consumption after about a 12 month stay in the archipelago. Besides, the elevated levels of 18:3 and 16:1 fatty acids imply dietary modifications of the serum fatty acids. PMID- 10528469 TI - Mental disorders in the Greenlandic population. A register study. AB - In a register study of all psychiatric first hospitalizations (1974-93) of persons born and resident in Greenland rates for all admissions as well as for the separate diagnostic groups, hierarchically organized, were compared with corresponding figures for the population in Denmark. Relative mortality rates for the psychiatric patients compared with the general population were computed for the Greenlandic and Danish populations respectively. No significant differences in the total pattern of hospitalization was found, but Greenlandic men 15-24 years old and Greenlandic women 25-34 years old had significantly higher and older age groups lower first admission rates than Danish men and women, respectively. The rates for affective psychoses were low especially among men in Greenland, whereas the rates for schizophrenia among men were comparatively high. The relative mortality risk compared to the general population was much higher among Danish than Greenlandic psychiatric patients, especially regarding suicide. A probable explanation for that is that the suicide rate in the Green-landic general population is very high. PMID- 10528470 TI - Surveying mortality in northern Finland: two towns with contrasting trends. AB - Mortality was compared in two major towns in northern Finland (Kemi and Oulu), which according to previous studies have high and low mortalities, respectively. In 1991-1995 life expectancy at birth in Oulu exceeded the national average by 0.7 years in males and 0.4 years in females but in Kemi it fell short of the national average in both sexes by 2.5 years. The shorter life-time in Kemi resulted from a slower-than-average decline in male mortality since the 1960's and an increase in female mortality by a factor of 1/5 in 1981-1995. In the 1990's, Kemi as marked by ischaemic heart disease (excess over national mortality 10% in males, 18% in females), pneumonia among the elderly (excess 2-fold), cancer in females (excess 30%), accidents (excess 24% in males, 82% in females) and male suicides (excess 76%), most of which in Oulu were below the national figures. Surveys are underway to clarify reasons for this mortality difference between the two towns. PMID- 10528471 TI - Hepatocellular carcinoma not related to hepatitis B virus infection among Alaska natives. AB - Chronic infection with hepatitis B or C viruses is a common underlying condition in patients with hepatocellular carcinoma worldwide. We studied serum and liver tissue from a cohort of Alaska natives with hepatocellular carcinoma (HCC) for evidence of hepatitis B, C and G viral infection using conventional serological tests as well as the sensitive polymerase chain reaction. Evidence of HBV infection was found in 25 and possible HCV infection in two cases. Among the remaining 11 patients, four had a history of recent or remote alcoholism while seven had no recognizable risk factors for HCC. Only one was seropositive for HGV RNA and that was an individual with a history of alcoholism. Non-tumorous liver tissue was available for study in six of these seven cases. Histological features of chronic hepatitis were present in five. Thus, at least five of 38 (13%) Alaska natives with HCC appeared to have chronic hepatitis not related to HBV or HCV infection, suggesting the possibility of some form of previously unrecognized chronic liver disease predisposing to HCC. PMID- 10528472 TI - High blood levels of persistent organic pollutants are statistically correlated with smoking. AB - Persistent Organic Pollutants (11 pesticides and 14 PCB-congeners), and heavy metals (Cd, Cu, Hg, Pb, Se, and Zn) were determined in 175 pregnant women and 160 newborn infants (umbilical cord blood) from Disko Bay, Greenland, 1994-96. Among these, 135 women filled out questionnaires about drinking, smoking and intake of traditional Inuit food. Multiple linear regression analyses showed highly significant positive associations between the mothers' smoking status (never, previous, present) and plasma concentrations of all the studied organic pollutants both in maternal blood and umbilical cord blood. Traditional food and not the tobacco is known to be the source of the contaminants. But smoking may influence the enzymatic turnover of toxic substances. PMID- 10528473 TI - Carpal tunnel syndrome between two centuries. AB - Compression neuropathy is one of the most frequently encountered conditions in a hand surgeon's daily practice. Median nerve compression at the wrist or carpal tunnel syndrome (CTS) is the most common of all nerve compression syndromes of the upper extremity1.3 and CTS has been used as a model for studying compression neuropathy (CN). This compressive lesion of the median nerve occurs in a tunnel located within the wrist called the carpal tunnel. Severe chronic inappropriately treated CTS can cause prolonged symptoms and lead to permanent nerve damage. Carpal tunnel release (CTR) is currently among the most commonly performed surgical procedures on the hand in the United States. The profound economic impact of this trend is reflected by the escalating tangible and intangible costs associated with its management. This article contains current information and cumulative knowledge up to the turn of the century on the basic science, etiology, diagnosis, and management of CTS. PMID- 10528474 TI - Substance use disorders in physician training programs. AB - Physician impairment due to substance use occurs in all age groups of physicians. Risk factors are similar for age-matched controls, but choice of substance may be influenced by specialty and narcotic permit. Alcohol is by far the most commonly abused drug involved. One of the last areas of performance affected is work. Other areas may provide earlier clues. Intervention and referral to appropriate treatment centers is important. Death rates are relatively high in this disease. We report a favorable outcome of 75 percent at The University of Oklahoma Health Sciences Center, Tulsa (OUHSC-T), but a death rate of 16.7 percent in our small sample. Aftercare and monitoring are essential to a successful outcome. The vast majority of physicians in training can complete their training with appropriate treatment and monitoring and go on to successful careers. PMID- 10528475 TI - Caveat lector: getting quality out of the Internet. PMID- 10528476 TI - Folic acid to prevent recurrence of neural tube defects. PMID- 10528477 TI - The rules have to change. PMID- 10528478 TI - [Laparoscopic cholecystectomy. An analysis of the reasons for a conversion to conventional surgery in an elective surgery department]. AB - BACKGROUND: To assess which factors determined conversion to laparotomy in patients undergoing laparoscopic elective cholecystectomy. SETTING: department of General Surgery. University of Genoa. Italy. METHODS: Two hundred sixty-four consecutive laparoscopic cholecystectomies were performed in our Department. INTERVENTIONS: laparoscopic cholecystectomy was performed according to Dubois's technique. Duration of the procedure was not considered a reason for conversion. RESULTS: 121 patients showed "difficult intraoperative situations" with further conversion risk factor. Conversion to laparotomy was necessary in 11 patients (4.16%). Five patients underwent conversion in the first 50 cases (10%), while six in the last 214 (2.8%). We had to convert to open cholecystectomy only in eleven patients, despite the high rate of technical difficulties and anatomic anomalies even in cases which, in the past, represented a contraindication to this kind of technique. The use of new instruments and new surgical techniques has reduced to only factors of increased risk in those situations that in the past were considered as contraindications to laparoscopic cholecystectomy. CONCLUSIONS: Conversion to open cholecystectomy is based on the surgeon's decision and the safety should be the main consideration in performing laparoscopic cholecystectomy. The use of a careful dissection could avoid the conversion in many patients. PMID- 10528479 TI - [Our experience in the transanal excision of tubulovillous adenomas of the rectum]. AB - BACKGROUND: Personal experience on transanal excision of rectal adenomas without affecting sphincteric function is reported. METHODS: From 1985 to 1997, 27 patients suffering from rectal adenomatous polyps underwent surgery; the sites of lesions were within 3 to 10 cm from anal orifice in the whole series; the age of patients ranged from 30 to 81 years. Two different procedures were employed: the Parks' technique and the electroresection by traction flap technique according to Faivre. RESULTS: Any postsurgical complication such as hemorrhage, stenosis or incontinentia occurred; surgical mortality was absent. Histological examination disclosed severe dysplasia as well as in situ carcinoma in 6 patients (22.2%) and malignant polyps in 9 patients (33.3%). Only in a case a palliative excision was performed since the poor general conditions of this patient did not permit a more extended treatment; a local relapse of the tumour associated with liver metastases led the patient to death 22 months after surgery. Three patients were lost to follow-up and 2 patients died because of other causes, 6 and 8 years after surgical excision, respectively. CONCLUSIONS: The conclusions are is drawn that either Park's and Faivre's procedures are useful and safe for the surgical treatment of rectal villous polyps extended up to 8-12 cm from anal orifice, in spite of the presence of malignant foci within their mass. These surgical procedures are simple and relatively poor traumatic; for this reason they are more suitable than other transabdominal or abdomino-perineal approaches for older patients and other at risk-patients. It is underlined that the treated patients require a long-term follow-up aimed at the early diagnosis of possible relapses of adenomatosis. PMID- 10528480 TI - [The surgical therapy of hemorrhoidal pathology performed in one-day surgery]. AB - BACKGROUND: The surgical procedure in the haemorrhoidal disease, performed in one day surgery, using a local anaesthesia (posterior perineal block), where suggested and possible, presents many advantages: for the patient whose stay in the hospital is shorter, for the hospital which is facilitated in the organization and planning of admissions, for the surgeon who is gratified by results. Personal experience is reported and the operation performed during one year, the surgical, anaesthetic and therapeutic technique used, and the results obtained are analyzed. METHODS: 32 selected patients with haemorrhoidal disease affections: (haemorrhoidal nodules by third and fourth degree). We have been examined. The operation was performed in all patients in one day surgery. The technique used was haemorrhoidectomy according to Milligan Morgan modified Phillips, using a coagulator, and without ligation of the pedicles. Anaesthesia was performed by Back perineal block according to M. C. Marti; particular attention was paid to the post-operative control that, in personal opinion, must be done weekly, for at last 4 weeks. RESULTS AND CONCLUSIONS: Results, which seem satisfactory, and approval manifested by the patients, encourage to continue in this direction. PMID- 10528481 TI - [Gallbladder carcinoma. Personal case histories]. AB - BACKGROUND AND AIM: Carcinoma of the gallbladder is a rare pathology which is extremely malignant with a fatal prognosis owing to its difficult and late diagnosis. It represents 3% of all digestive tumours and is predominant in females. The major risk factor is gallbladder stones. METHODS: A total of 14 gallbladder neoplasms were diagnosed in 798 cholecystectomies performed from October 1990 to October 1995, with a male/female ratio of 6: 1 and a mean age of 68 years and 11 months. Biliary colic and jaundice represented approximately 93% of symptoms present in this series. Fourteen cholecystectomies were performed in association with other forms of surgery (1 GEA, 2 drainage acc. Kehr, 1 hemicolectomy, 1 resection of the hepatic bed, 1 hepaticojejunostomy). RESULTS: The operative mortality was nil. Mean survival was 1 year, 4 months and 15 days, and was obviously higher in the early stages. Four patients (all female) are still alive (mean survival 2 years 10 months 9 days). The five-year survival rate reported in the literature does not exceed 5-10%. Surgery does not currently give satisfactory results. In the light of present knowledge it is important to intervene on the risk factors in the form of laparoscopic cholecystectomies for patients with lithiasis or gallbladder polyps. CONCLUSIONS: Diagnosis must be as early as possible during the "early cancer" phase in order to ensure that surgery is oncologically radical and therefore also remedial. PMID- 10528482 TI - [The surgical treatment of lung metastases. The prognostic factors and the indications for the surgical approach]. AB - BACKGROUND: After the liver, the lungs represent the most frequent site of metastasis from primary tumours. Surgical treatment of lung secondary neoplasms leads to a significant improvement in survival. METHODS: Between 1960-1997, 178 patients with lung metastases underwent surgery at the Thoracic Surgery Department of Turin University in a total of 193 operations. A retrospective study was made in order to identify the prognostic factors which influenced final survival in this population. RESULTS: Overall survival was 47% after 2 years and 20% after five years. Prognosis was not influenced by the size of metastases, the type of surgery, adjuvant therapy and the number of operations on the same patient. On the other hand, useful prognostic factors were found to be the histological type of the primary tumour, the original site of the neoplasm, the number of metastases and, above all, the disease-free interval (DFI). CONCLUSIONS: Lung metastasectomy is an important therapeutic aid in selected patients, whereas the preoperative evaluation of the above prognostic factors enables a reasonably precise prognosis to be made in most patients. PMID- 10528483 TI - [The one-time surgical repair of postintubation esophagotracheal fistulae]. AB - Personal experience in the treatment of the tracheal-esophageal non-neoplastic fistula is reported. In the last years, three cases of FTE, concerning some cannula tracheal-stomachal beare patients from 14, 2, 1 months have been examined. In two cases the patients were in spontaneous ventilation, on the contrary a mechanical ventilation was employed in the third. In two patients the fistular way was located correspondingly of the decubitus point of the tracheal stomachal cannula, on the membranaceous pars, and it was not associated with concomitant tracheal stenosis. The first stage of the treatment was removal of the nasogastric probe, supporting the decubitus phenomenon subtending the establishment and the extension of the FTE, then the preparation of a gastrostomy to assure the drainage of secretions under the fistula and a jejunum anastomosis to allow a suitable feeding and recovery of the patients. In these three cases the restoring operation was accomplished by a cervicotomy with a direct opening of the fistula, a suture of the esophageal wall, a suture of the membranaceous pars on the healthy tissue and then a protection of these sutures by interposition, between trachea and esophagus, of the prethyroid muscles transposed and fixed to the prevertebral band. In two cases the post-operative course did not present complications, while the patients kept in assisted ventilation during the postoperative course showed a relapse of the FTE on the twelfth day and then the progressive establishment of a septic state and the exitus on the twentieth day. It is underlined how the success of the reparation of the fistula is largely conditioned by the respiratory autonomy of the patient that guarantees the recurrence of decubitus and infection phenomena causing the lesion. PMID- 10528484 TI - [The laparoscopic treatment of a rare case of intestinal obstruction due to an ectopic pregnancy]. AB - A rare case of intestinal occlusion due to primary abdominal pregnancy is described. Laparoscopy revealed normal appendix and bowel obstruction by adhesion between the last ileal loop and cecum. The adhesion started from a neoformation (diameter 2 cm) localised on the mesenteric side of the ileum, about 30 cm from the ileocecal valve. A resection of the adhesion and dissection of the neoformation were performed. Laparoscopic procedures lasted 30 minutes. Histologic examination of the specimen revealed to be an ectopic pregnancy. The laparoscopic technique permitted to verify the diagnosis and perform the treatment of the abdominal pregnancy in absolute conditions of safety, maintaining the fertility of the patient (actually she is presenting a regular pregnancy). PMID- 10528486 TI - [Peritoneal cystic mesothelioma. A clinical case and review of the literature]. AB - Multicystic peritoneal mesothelioma is an extremely rare benign neoplastic disease with high tendency to recur locally, but no tendency to malignancy. Correct diagnosis can be made with histopathologic examination and always with immunohistochemical and ultrastructural evaluation. A case in a twenty-eight-year old woman is reported and the anatomo-clinical characteristics of multicystic peritoneal mesothelioma from sixty-nine cases described in the literature are specified, discussing the management of this disease and emphasizing the importance of a nondemolitive approach. PMID- 10528487 TI - [Lymphatic cyst of the mesentery]. AB - Lymphatic cyst of the mesentery is a rare lesion; one case per 100,000 hospital admissions is reported. A case of lymphatic cyst in a sixty-one-year-old woman is presented. The symptoms are extremely variable, not characteristic and correlated to the location and size of the cyst. Abdominal ultrasonography and computed tomography may lead to a correct diagnosis, which is regularly made at the time of abdominal exploration. In this case the cyst was enucleated from the mesentery with open surgery. PMID- 10528485 TI - [Stomach rupture due to barotrauma (a report of the 13th case since 1969)]. AB - The thirteenth case of rupture of the stomach after a diving accident since 1969 is reported. This rare event was caused by equipment failure and panic reaction, which induced swallowing air during diving and consequential gas expansion in gastric cavity meanwhile the rapid ascent. Peritoneal decompression by paracentesis quickly improved the patient's condition and the following surgical laparotomy revealed a gastric tear along the lesser curvature, which was closed by suturing. The patient presented a postoperative splenic abscess two months later; literature demonstrated that rupture of a filled stomach may lead to septic complications. PMID- 10528488 TI - [Perineal hernia]. AB - The authors base this study on a case of perineal hernia referred to their attention. In the light of the scant international literature on this subject, they focus on the topographical anatomy of the pelvic floor in order to gain a clearer understanding of this pathology, as well as their classification into median, lateral, anterior and posterior forms. Above all, the authors draw attention to the importance of the differential diagnosis of perineal hernia from Bartholin cysts or vulvar tumours in relation to anterior perineal hernia, and perianal abscesses in relation to posterior hernia. They underline the value of ultrasonography or TAC during the diagnostic procedure. Lastly, they examine the channels of aggression for this type of hernia which may be abdominal, perianal or combined (abdominal and peri-anal), as well as the repair techniques used, varying from direct suture with non-absorbable material to the use of prolene mesh or flaps if the hernia breech is very large. PMID- 10528489 TI - [Pneumothorax secondary to pulmonary histiocytosis X]. AB - The authors report a case of pneumothorax in a young man suffering from pulmonary histiocytosis. They take this observation as a starting point for discussing about the so-called "histiocytosis X". The term "histiocytosis" is a term for a variety of proliferative disorders of histiocytes or macrophages. There is a small cluster of conditions characterized by proliferation of a special type of histiocyte called the "Langerhans' cell". The proliferation of Langerhans' cells results from disturbances in immunoregulation. Particularly, there are three clinicopathologic entities: *Letterer-Siwe disease (acute disseminated Langerhans' cells histiocytosis), *multifocal eosinophilic granuloma or Hand Schuller-Christian disease (multifocal Langerhans' cells histiocytosis), *unifocal eosinophilic granuloma (unifocal Langerhans' cells histiocytosis). In the past, these were collected in the not specific term of histiocytosis X. Most patients with Langerhans' cell granulomatosis have diseases limited to the lung, only a small proportion have extrapulmonary involvement or a systemic disorder. Typically, there are expanding, erosive accumulations of Langerhans' cells within the medullary cavities of bones. In this reported case, a patient, two years before, was affected by osteolytic bone lesion in the left femur. Pulmonary histiocytosis is often seen in smokers in the third and fourth decade of life. The 20% of patients are asymptomatic and found to have an abnormal chest radiograph; a lesser proportion presents with pneumothorax. Some patients develop progressive fibrotic lung disease and respiratory failure. Chest radiographs may show bilateral reticular or reticulonodular infiltrates, sometimes with cystic changes, often with sparing of the costophrenic angles, but high resolution computerized tomography scans may be virtually diagnostic based on the predilection of the cysts and nodules for the upper lung zones. Sometimes the typical Langerhans' cells may be recovered in bronchoalveolar lavage fluid of patients with active disease. The combination of the characteristic CT findings and Langerhans' cells in bronchoalveolar lavage fluid should be diagnostic, decreasing the number of cases requiring for diagnosis open lung biopsy. The differential diagnosis is with sarcoidosis, tbc, diffuse metastases, lymphoma, eosinophilic pneumonia, fibrosing interstitial pneumonia. PMID- 10528490 TI - [An experimental study of the use of synthetic meshes in large abdominal eventrations]. AB - BACKGROUND: The aim of this study is to evaluate the biocompatibility of Angimesh mesh (polypropilene) in rats, usually used to replace or to strengthen the abdominal fasciae. METHODS: We made an abdominal longitudinal incision of skin and muscles in 20 rats was carried out and two muscular semilunae removed. The incision was occluded with Angimesh mesh fixed by ethilcyanoacrilate glue (first group of rats) or with suture stitches (second group of rats). The animals were sacrificed after 80 days and the stability of the mesh and the presence of endoperitoneal adhesion were evaluated macroscopically, and microscopically the fibrogenesis and inflammation answer. RESULTS: Two rats of the first group and seven of the second group died before 30 days. The mesh was more steady when fixed by suture. Adhesion in the contact sites between the mesh and the peritoneum was found. Histology did not show any inflammation reaction but showed a fibrogenic answer. CONCLUSIONS: The cyanoacrilate non-toxicity has been confirmed also in this application. Angimesh mesh showed good biocompatibility and resulted ideal for the synthesis of large abdominal resections. PMID- 10528491 TI - [Our experience in internal medioposterior sphincterotomy with anoplasty]. AB - BACKGROUND: The technique of the posterior medial internal sphincterotomy according to Arnous is described and advice from personal experience reported. METHODS: From 1981 to 1995 posterior medial internal sphincterotomy with anoplasty was performed in 270 patients (132 males and 138 females) affected with chronic anal fissure, alone (112 cases) or associated with hemorrhoids (80 cases); moreover, the Arnous's operation was performed as well for hemorrhoids with posterior subsidiary packet and internal sphincter hypertonia (52 cases), relapsed hemorrhoids with postoperative anal stenosis (24 cases) and postoperative anal stenosis with painful fissure and residual internal sphincter hypertonia (2 cases). RESULTS: The results have been excellent; the average stay in hospital has been 7 days and the complete recovery occurred after 4-6 weeks without early or late complications. CONCLUSIONS: The importance of anorectal manometry for the preoperative valuation of the sphincteric hypertonia is emphasized: in this manner it is possible to modulate the sphincterotomy avoiding too economic sphincteric sections with next residue hypertonia or, in the contrary, too plentiful sphincteric section with problems of continence. Finally, the internal lateral-left sphincterotomy is mentioned, which is efficacious in the treatment of acute anal fissure. However, the proctological surgeon, on the basis of his experience, will propose the most convenient technique. PMID- 10528493 TI - Erythema chronica migrans, ticks and Lyme disease in Missouri. PMID- 10528492 TI - [The nasojejunal tube in early postoperative nutrition]. AB - BACKGROUND: To verify the effectiveness of the nasojejunal tube inserted during operation as an alternative to jejunostomy to perform early enteral feeding. METHODS: EXPERIMENTAL DESIGN: Prospective study. SETTING: Department of Surgery, General Hospital. PATIENTS: 27 patients undergoing laparotomy because of a gastric pathology. INTERVENTIONS: In 18 patients before construction of the distal jejunum anastomosis the tube was inserted by nasal route and advanced into the jejunum ansa until the end reached 15 to 20 cm down the anastomosis (group A); 9 patients underwent a jejunostomy according to Delany (group B). All the patients started enteral feeding 24 hours after operation and had the same polymeric diet, given to them using the same procedures. MEASURES: postoperative complications, tube intolerance, intestinal tolerance. RESULTS: The degree of non acceptance of the tube was: absent in 3 patients of group A and in 7 patients of group B (p > 0.05); slight in 6 patients of group A and in 2 patients of group B (p > 0.05); medium in 9 patients of group A and in no one of the group B (p no measurable); high in neither groups. The intestinal tolerance was similar in both groups. CONCLUSIONS: Nasojejunal tube is an effective alternative to jejunostomy to perform early postoperative enteral feeding. PMID- 10528494 TI - Erythema chronica migrans, ticks and Lyme disease in Missouri. PMID- 10528495 TI - Teaching students managed care. PMID- 10528496 TI - Dinner series to incorporate EPEC Project. Education for Physicians on End-of Life Care. PMID- 10528497 TI - Toward the survival of private healthcare delivery. AB - PREMISE: a major cause of the rising real cost of healthcare is the lack of coordinated management. The task of the delivery of care falls onto three autonomous systems: hospitals, physicians, and insurers. If all these entities were interdependent subsystems within an encompassing task system, the redundancies and duplications, which impair progress and efficiency, could be modified or eliminated. The establishment of internal and external feedback mechanisms, seriously lacking at present, would improve function and allow adaptation to the present accelerating changes in the healthcare environment. PMID- 10528498 TI - Radiology quiz. Basilar skull fracture. PMID- 10528499 TI - The importance of mammography. PMID- 10528500 TI - Tuberculosis screening in Kansas City homeless shelters. AB - Voluntary tuberculin skin testing, coupled with on-site radiographic examination of persons with indurations > or = 10 mm, was conducted in five home-less shelters in Kansas City. Of 856 skin tests administered, 654 were read and 89 (13.6%) were positive. Males were nearly four times as likely to have a positive skin test than females. None of the positive individuals had abnormal chest radiographs. Of 42 persons who initiated preventive therapy, only eight completed the course of treatment. PMID- 10528501 TI - The development and implementation of a population-based intervention model for public health nursing practice. AB - Mandatory enrollment of Medicaid recipients into managed care organizations (MCOs) in 1994 resulted in a major reduction in primary medical care provided by the Peninsula Health District (PHD) in southeastern Virginia. This article describes how the PHD responded proactively by changing from a predominantly primary care clinic practice to a population-based public health practice and changed the role of public health nursing. A participatory planning process involving all PHD employees carefully defined this new direction over several years. As public health care resources were redirected into preventive programs to serve the health needs of the entire community rather then the individual, a new intervention model evolved. The intervention model became the framework for program planning and implementation and has now been in use by the PHD for 3 years. The planning process used by the PHD to devise a population-based intervention model is described in this article. PMID- 10528502 TI - Predictors of health promotion lifestyle among three ethnic groups of elderly rural women in Taiwan. AB - The purpose of this study was to examine the predictors of health promotion lifestyle (HPL) and examine the similarities and differences among three ethnic groups of elderly rural women in Taiwan. Pender's Health Promotion Model was used as the conceptual framework of this study. A convenience sample of 599 elderly rural women was recruited from three rural areas: Kao-Shu, San-Di-Men, and Ma Chia. Ho-Lo, Hakka, and aboriginal people are the three main ethnic groups in these areas. Of the 599 elderly women, 391 completed all of the interview questions. Subjects ranged from 65 to 91 years old. All instruments used in this study have been evaluated for their content validity. The interrater reliability and alpha coefficient reliability of all instruments were greater than 0.70. A survey-interview method was used to collect data. Findings showed that the predictors of HPL have differences and similarities among elderly rural women from different ethnicities. In the group of elderly Ho-Lo women, age, education, living arrangements, and perceived barriers to health promotion lifestyle (PBaHPL) were significant predictors and they explained 41.9% of total variance in HPL. In the group of elderly Hakka women, education, number of chronic health problems, PBaHPL, and perceived benefits of health promotion lifestyle (PBeHPL) were significant predictors in explaining 53.9% of total variance in HPL. Finally, in the group of elderly aboriginal women, living arrangements, PBaHPL, and PBeHPL were significant predictors in explaining 70.0% of total variance in HPL. Community nurses can use their understanding of different ethnic groups to assess, identify, and use effective health promotion interventions for elderly rural women. PMID- 10528503 TI - Issues in balancing teenage clients' confidentiality and reporting statutory rape among Kansas Title X clinic staff. AB - Through Federal welfare reform, Congress directed states to aggressively enforce statutory rape laws. Family planning professionals deal with many adolescent clients, and their support for such enforcement or willingness to report is unclear. The authors of this study examined current attitudes and practices of family planning program managers (FPPMs) about statutory rape law enforcement, including current reporting practices. In 1997, all 77 local Kansas Title X FPPMs were surveyed. Structured telephone interviews were conducted with 10 FPPMs to add detail to quantitative responses. Sixty-eight FPPMs responded to the written survey (88%). Of these, 79% supported aggressive enforcement, and 43% thought enforcement would reduce adolescent pregnancy rates. With increased enforcement, 38% believed teenagers would be discouraged from seeking reproductive health care, compared to 41% who believed they would not. Among key informants, all of whom were FPPMs, willingness to report cases was mixed, with those who would report wanting the flexibility to judge on a case-by-case basis. For those not reporting cases, confidentiality concerns overrode beliefs in any positive outcome of enforcement. Kansas Title X FPPMs strongly supported aggressive enforcement, but had mixed beliefs about negative consequences. Among those interviewed, there were also mixed beliefs and practices about reporting. Reporting from FPPMs will be sporadic and arbitrary unless protocols are developed and laws are clarified. PMID- 10528504 TI - Surveillance of suicidal behavior in Kitsap County, Washington: a retrospective study. AB - Suicide is a major source of preventable morbidity and mortality in Kitsap County, Washington State. This article describes a study of suicidal behavior to identify risk groups in order to design intervention strategies. A retrospective study was conducted by reviewing the charts of individuals exhibiting suicidal behavior who had presented to the county's only civilian hospital emergency department over a 7 month period. Frequencies were calculated to identify at-risk populations and determine risk factors. One hundred forty-five charts were reviewed. Subjects were mostly female (69%), and ages ranged from 10 to 80 years with 73% between 15 and 44 years. Two-thirds of the subjects were not working. More than half had previously exhibited suicidal behavior and more than 75% had previous mental health encounter(s). Most admissions (67.6%) occurred between 4:00 p.m. and 4:00 a.m. The core public health functions of assessment, policy development, and assurance provided the framework for this community to explore the finding that suicide was a major source of preventable morbidity and mortality. Community-based intervention strategies have been developed in an effort to reach the Healthy People 2000 objective of reducing suicide deaths to no more than 10.5 per 100,000 residents. PMID- 10528505 TI - Wellness profile of midlife women with a chronic illness. AB - Among issues important to women's health are their wellness profiles including indicators such as activity level, weight status, breakfast and snacking patterns, health status perceptions, and alcohol and tobacco use. This is particularly true for midlife women with a long-term illness. The purpose of this study is to identify the wellness profile of a group of midlife women with multiple sclerosis (MS), and to compare their life-style indicators with national health statistics. Overall, the women in the study group indicated a lower perception of their health status and were less active. However, the women in this study group demonstrated healthier body weights, used less tobacco and alcohol, had better breakfast patterns, and comparable snacking patterns, indicating that may be more attuned to their bodies than women without chronic illnesses. Identifying women's wellness profiles can assist practitioners in addressing the issues of health for women managing a long-term illness. PMID- 10528506 TI - An oral health promotion module for the primary health care nursing course in Acornhoek, South Africa. AB - The severe personnel shortage in the health professions in most developing communities is well documented. This dearth of health workers and the widespread adoption of the primary health care approach (PHCA), requires that health professionals be trained to understand and effectively utilize the skills offered by other disciplines in the health care field. Nurses are expected to play active roles in the promotion of health including oral health, particularly in the rural under-served communities. There is a paucity of oral health education in nursing curricula. This article describes a 4-day module on oral health promotion developed and delivered to 36 nurses as part of a 12-month primary health care nursing (PHCN) training course. The module utilized a variety of educational methods and materials to facilitate and encourage both individual and group learning. The module was evaluated using both student academic attainment and perceptions as outcome measures. The evaluation and the experience of facilitating this module show that an oral health promotion module of short duration can sensitize nurses to the importance of oral health and increase oral health knowledge and skills. PMID- 10528507 TI - Public health nurses' responses to domestic violence: a report from the Enhanced Domestic Abuse Intervention Project. AB - Public health nurses (PHNs) can play an important role in the detection of domestic violence. This study examines whether the introduction of a domestic violence assessment protocol by public health nurses in a maternal and child health visiting program increases the identification and referral rates of women experiencing domestic violence. Data collected from case files during the baseline year prior to the initiation of the protocol were compared to case file information after the protocol had been implemented. When the protocol was used, there was a higher rate of identification, although the difference was not statistically significant. Significantly more women, however, were provided with information about domestic violence resources after the protocol was in place, and significantly more women were referred to services in the second year after the protocol had been implemented. This study provides support for the use of a domestic violence protocol to improve the public health nursing response to domestic violence. PMID- 10528508 TI - Peer education project with persons who have experienced homelessness. AB - This paper describes an unconventional health education project implemented by nurse practitioners in a nurse-managed clinic serving persons who are homeless. The nurse practitioners perceived that there were a number of potential barriers to providing health education to the homeless patients. These barriers included the fact that this patient population is part of a marginalized subculture affected by a variety of overwhelming social problems. An additional barrier was that the nurses often differed from their homeless patients in terms of race, gender, socioeconomic status, formal education, culture, and life experience. The nurse practitioners designed the Peer Health Education Project (PHEP) to try to overcome some of these barriers. The purpose of the PHEP was to increase the health education knowledge and empowerment of persons who had experienced homelessness so that they could serve as peer health educators with others who were living on the streets. The project model was based on the philosophy of Paulo Freire (1973). The peer health educators served as both leaders and participants in each education session. The educators set the agenda and the nurses acted as facilitators. The project was successful in preparing peer educators. Other indicators of the success of the project included increased empowerment, self-esteem, dignity, hope self-confidence, and community participation of the peer educators. PMID- 10528509 TI - [Obstetrical paralysis. Spontaneous remission, primary treatment of the nerve lesion and secondary treatment by muscle transfers]. PMID- 10528510 TI - [Palliative surgery by muscle transfer in the treatment of footdrop]. PMID- 10528511 TI - [Decubitus ulcers, alternative therapies]. PMID- 10528512 TI - [Reconstruction of facial and neck defects after surgical excision of skin tumors]. PMID- 10528513 TI - [Radionecrosis, physiopathology and treatment. Review of sixteen cases]. PMID- 10528514 TI - [Hypertrophic scars and keloids: which therapeutic options today?]. AB - Hypertrophic scars and keloids are a hyperproliferative response of connective tissue to trauma. Histologically the difference between the two is that keloids invade normal tissue whereas hypertrophic scars remain confined within the original wound. A variety of treatments have been proposed, which we will review according to their efficiency. Prevention of pathological scarring will also be discussed, and we will present our current attitude to treat these scars. As a surgical treatment for keloids, we have been using the intralesional technique which we think gives better results. PMID- 10528516 TI - [Principles of physics and application of the laser in plastic surgery]. AB - Recent advances in laser technology have expanded the surgeon's possibilities to treat various cutaneous lesions, as well as proposing a new tool for skin rejuvenation. Review of the basic physical principles of laser energy then overview of the different lasers used in plastic surgery as well as some of their clinical applications. PMID- 10528515 TI - [Breast augmentation: indications, types of prostheses, surgical techniques, complications, results]. AB - Augmentation mammaplasty is one of the most frequent surgical procedures in plastic surgery. The indications and techniques are precise and should be closely observed. Silicone gel implants are still used in Europe and multiple epidemiologic studies have failed to demonstrate an association between silicone breast implants and autoimmune disease. After a short historical review, we will present the indications, techniques and complications. As capsular contracture is the main complication, we shall discuss the latest issues. Data was collected on a group of patients using a systematic questionnaire and clinical follow-up. Patients' satisfaction is optimal in the absence of capsular contracture. We are convinced that silicone breast implants have an appropriate texture and give excellent aesthetic results. We are satisfied with silicone breast implants and continue to think that this device has an appropriate texture and gives excellent aesthetic results. PMID- 10528517 TI - [Revision of the carpal tunnel, study of 60 cases]. PMID- 10528518 TI - [Hand infections]. PMID- 10528519 TI - [Indication for toe transfer in congenital malformations of the hand]. PMID- 10528520 TI - Estimation of chronic hepatitis C activity based on ultrastructural changes of hepatocytes. AB - Seventy four liver bioptic specimens of 66 patients with chronic hepatitis C were analyzed ultrastructurally and by light microscopy. According to histological activity of the disease, the patients were divided in groups with minimal activity (CPHa), mild activity (CPHb-CAHa), and moderate and severe activity (CAHb,c), respectively. The presence of lamelar, simple and complex lipid inclusions in the cytoplasm, as well as the presence of nuclear bodies and morphology of the nucleolus were analyzed by classic methods of transmission electron microscopy. Cytoplasmic lamelar inclusions were found in 84.6% of patients, and simple and complex lipid inclusions were noted in 85.1% of patients, without the differences related to histological and enzyme activity. Unaltered nucleolus was most frequently observed in patients with minimal disease activity (84.6%), viral changed in patients with mild activity (48.4%) and synthetic active in patients with moderate and severe activity (66%). Serum aminotransferase activity was significantly higher in patients with viral changed and synthetic active nucleolus. These results can indicate that HCV induces cytopathogenic and immunologic damage of hepatocytes. The presence of virus acts as toxic agent that damages hepathocytes' metabolism, thus resulting in occurrence of cytoplasmic lamelar, simple and complex lipid inclusions, respectively. Immunologic damages occur when the virus alters cell membrane of hepatocytes. PMID- 10528521 TI - [Experimental study of the pathogenesis of frostbite. Part III. Significance of energy status of muscle tissue in the pathogenesis of frostbite]. AB - The authors have investigated energy status of muscle tissue of the lower legs of Wistar rats subjected to freezing, as well as the dependence of ultrastructure changes in muscle tissue mitochondria on the intensity of freezing. The animals were divided into three experimental groups: two groups were exposed to cryoinjury of different intensity, while the third was the control one. Cryoinjury was applied over an experimental after thawing were taken samples of muscle tissue for the analysis of content of energy phosphates and carbohydrates' metabolites, respectively, as well as for morphometric analysis of mitochondria volume out of frozen right and unfrozen left lower legs. Results obtained from different experimental groups were compared mutually and with control group that was not exposed to cryoinjury, respectively. It was found that the freezing in muscle tissue caused hydrops degeneration of mitochondria, depletion of energy reserves, reduction of energy status and the activation of mechanism of anaerobic metabolism. All these alterations were proportional to the intensity of freezing and considerably influenced contractility function of muscle cells, causing prolonged spasm of arterioles in thawed tissue, i.e., rigor congelationis that was primarily defined as an important factor in the pathogenesis of ischemic impairment of thawed tissues. PMID- 10528522 TI - [Evaluation of perfusion of transplanted microvascular flaps using radionuclide arteriography]. AB - Microvascular flap by its autonomous circulation, following the transplantation in recipient region, provides the quantity of blood sufficient for nutritive needs of flap, but it also provides additional blood supply in the surrounding of recipient region. The evaluation of the relation of the tissues perfusion of transferred microvascular flaps and the tissues of surrounding recipient region was done in 33 patients, 15.24 months after the transplantation of microvascular flaps, by radionuclide angiography using Tc-99m in the form of MAA HAS or pertechnetate at the Clinic for Plastic Surgery and Burns of Military Medical Academy (MMA) and at the Institute for Nuclear Medicine of MMA. Cutaneous, myocutaneous and osteocutaneous microvascular flaps were transferred to the foot, lower leg and in the head region. The obtained results revealed that the perfusion of the tissue of transferred flaps was better than the perfusion of the recipient region surrounding (232%). Perfusion of myocutaneous flaps related to the type of microvascular flap was the highest and came to 244% of the perfusion of the recipient region surrounding. Accumulation of radiopharmaceuticals in the tissue of the flap was homogenous in 19 (57.57%) flaps. PMID- 10528523 TI - [In vivo study of spontaneous and therapy-induced apoptosis in patients with chronic lymphocytic leukemia treated with chlorambucil]. AB - The efficiency of Chlorambucil in the induction of apoptosis was investigated in the study, and measurable apoptosis parameters were compared to the other prognostic factors with the aim of possible prediction of clinical response to the therapy in the patients with CD5 + B-cell chronic lymphocytic leukemia (B CLL). Seven newly diagnosed patients, initially treated with daily high-doses of Chlorambucil (HD-CLB) were analyzed. Quantitative analysis of apoptosis parameters on semi-fine sections obtained from peripheral blood was performed prior and during the first five days of therapy. The level of spontaneous apoptosis (SA) was determined, as well as the maximal response by apoptosis (MAR), and the time needed to establish maximal response by apoptosis (TMAR), respectively. The results revealed that the level of SA in the studied group of patients was 11.39%-20.50%. In three patients with achieved criteria for complete remission (CR) was observed high level of SA, TMAR 2-4 days and MAR 23.42-26.36%, respectively. All patients with CR were with negative LDT, non-diffuse involvement of bone marrow and clinical stage B. Criteria for partial remission (PR) were achieved in 4 patients. Within this group, all three measurable parameters of apoptosis could have been determined in only one patient, while in the rest was noticed the increased percentage of apoptotic cells on the last day of follow-up. In all patients was observed negative LDT, diffuse bone marrow involvement, and 2 out of 4 patients had CLPL of cytomorphological type and clinical stage B. By comparing the obtained values of measurable apoptotic parameters with the clinical response to the applied therapy with HD-CLB, it is possible to divide our patients into two groups: patients who have achieved CR have the highest percentage of cells dying due to the therapy-induced apoptosis, as well as the higher values of measurable parameters compared to the certain parameters of the patients with the criteria for PR. Our preliminary results of therapeutic response to the apoptosis might be useful for the timely decision upon the duration of therapy and change of modality of treatment for every patient during the follow-up period. PMID- 10528524 TI - [Effect of the conus telescope system on the periodontal status of abutment teeth in Kennedy class I partial edentulousness]. AB - Periodontal status of abutment teeth of conus telescope system was investigated in the production of telescope prosthesis on two and four abutment teeth in Kennedy class I edentulousness. In the first case, abutment teeth of conus telescope system were the first lower premolars, and in the second case the first lower premolars and canines. All investigations were performed on first lower premolars, and included the follow-up of following indexes and parameters: gingival index, periodontal pocket depth, bone resorption index, abutment teeth mobility and dental plaque index. Investigations were performed in two groups of patients: before and after three months and one year after the prosthetic management. The results of clinical investigations revealed the improvement of all investigated parameters three and twelve months later compared to the state prior to the production of telescope prosthesis. PMID- 10528525 TI - [Manufacture of lithium carbonate tablets using direct compression]. AB - The test results of lithium carbonate tablets made by direct compression were presented. The content of lithium carbonate, tablets weight variation, hardness, friability, disintegration, as well as the lithium carbonate dissolution rate were determined. The best properties were observed in tablets made with microcrystalline cellulose (Avicel PH 101), spray dried lactose and corn starch. PMID- 10528526 TI - [Stable and unstable coronary atherosclerotic plaque]. PMID- 10528527 TI - [Education of the radiologist in the 21st century]. PMID- 10528528 TI - [Importance of determination of autoantibodies in autoimmune diseases]. PMID- 10528529 TI - [Liver damage in systemic connective tissue diseases]. PMID- 10528530 TI - [Pregnancy, puerperium and neurologic complications]. PMID- 10528531 TI - [Complications of percutaneous transluminal renal angioplasty--case report]. AB - Complications of percutaneous transluminal renal angioplasty (PTRA) most frequently occur on dilated artery, and rarely on peripheral punctured artery. In the study was presented 24-year old female patient in whom the painful tumor developed five days after PTRA at the spot of axillary artery punction. Tumor caused the lesion of brachial plexus by the compression, which was the reason for urgent surgery. PMID- 10528532 TI - [Stress ulcer in a patient with a subacute subdural hematoma]. AB - During the onset and development of pathologic events, numerous changes occur in the body as the result of an attempt of dynamic equilibrium maintenance. Results of such sequence of events may lead to organic, as well as psychic impairment of the body that is often referred to as the onset of psychosomatic disease. This paper deals with the disorder that could provide better insight in the possible physiopathologic mechanisms of psychosomatic disease or disorders. In a patient, chronic alcoholic subdural hematoma was diagnosed. Peptic ulcer hemorrhage occurred after surgical removal of the hematoma. Possible physiopathologic mechanisms in the origin of this disorder have been described. PMID- 10528534 TI - Induced hypocalcaemia by Na2EDTA infusion. A review. AB - Disodium ethylenediaminetetraacetate (Na2EDTA) has been used in infusion studies in cows and other species for more than 35 years, primarily for the induction of hypocalcemia as a model for milk fever. This paper reviews such studies and discusses blood calcium kinetics, toxicology, changes in various blood parameters and the effect on blood circulation, cardiac function and smooth muscle motility in the gastro-intestinal tract and in the pregnant uterus. It is concluded that Na2EDTA infusion may serve as a valid model for spontaneous hypocalcemia. However, experimental results may vary with factors such as the choice of method for blood total calcium analysis, and the rate of EDTA infusion. Standardization of these and certain other experimental conditions may greatly improve the comparability of results obtained in EDTA infusion studies. For cows, an infusion rate of 1.2 ml/kg/h of a 5% (w/v) solution, corresponding to 0.25 mmol/kg/min, has been suggested as a standard infusion. PMID- 10528533 TI - [Lipedema of the leg associated with dermatomyositis]. AB - Lipedema represents a form of lipodistrophy, which consists of abnormal accumulation of fat in subcutaneous tissue of the lower limbs with consecutive development of lymphostasis and lymphedema. The aim of this article was to review one clear case of lower limbs lipedema, of unusual occurrence and appearance, which was associated with dermatomyositis. A moderately manifested lipedema in 8 years old little girl was reported with its expressive segmental distribution to upper and lower legs, without significant increase in its size during last 10 years and without signs of lymphostasis. The hereditary influence was not confirmed. Histological examination of lipedematous tissue revealed significant presentation of immune component of the disease. According to the available literature, association between lipedema and dermatomyositis, lower limbs lipedema with segmental distribution as noticed above and its appearance as a consequence of corticosteroid therapy have not yet been published. PMID- 10528535 TI - Relationships amongst liver bile salt clearance, bile secretion and infusion of lipids in calves. AB - In order to study the influence of portal lipid loading on the extraction rate of bile salts by the liver, four cholecystectomized calves (mean body weight 103 kg) were fitted with permanent cannulae to the common bile duct, duodenum and portal vein. A venflon catheter was also set up in the jugular vein to collect blood for analysis of fatty acids (FA) and bile salts (PBS) in plasma. The experiments were divided into two parts. In the first part sodium taurocholate (TCHNa) was infused for at least 2 h at a rate of 25 mumol/min into the duodenum to stabilize the bile flow and bile salt output in bile and the concentration in plasma. In the second part, as well as TCHNa, Intralipid (Itlp) (infusible 10% of lipid compounds) was also infused into the portal vein. Itlp was infused for 40 min, starting at a rate of 3 ml/min at the beginning of the 3rd hour of TCHNa infusion followed by a rate of 6 ml/min at the beginning of the 4th hour of TCHNa infusion. During TCHNa infusion the plasma bile salt concentrations were in the range 15.69-20.21 mumol/l, similar to that of the pre-infusion period. Introduction of Itlp to the infusion of TCHNa resulted in a significant (P < 0.05) increase of PBS, about 2 times higher at an Itlp infusion rate of 3 ml/min, and 3 times higher (62.82 +/- 16.42 mumol/l) at 6 ml/min. Under Itlp infusion, all common plasma FA increased, but the largest increases were in levels of linolenic, palmitic and oleic acids. During TCHNa infusion, the bile flow and the content of bile salts in bile did not change. The infusion of TCHNa with Itlp at the rate of 6 ml/min caused a 2-fold decrease both of the bile flow and of the output of bile salts from 18.58 +/- 3.04 microliters/min/kg and from 0.58 +/- 0.07 mumol/min/kg observed at the beginning of both infusions to 9.51 +/- 2.95 microliters/min/kg and 0.28 +/- 0.05 mumol/min/kg, respectively, at the end of the collecting period. When only TCHNa was infused, almost all of it was secreted to the bile, while with the additional infusion of Itlp only about half of the infused TCHNa was secreted to the bile. These results indicate that the extraction rate of PBS by the liver is decreased by loading of the portal blood by lipids, allowing more bile salts to escape into the systemic circulation, and thus reducing bile production. PMID- 10528536 TI - Age determination in mini-Shetland ponies and donkeys. AB - The accuracy of ageing mini-Shetland ponies and donkeys was assessed by correlating the appearance of specific dental features with the known ages of 106 mini-Shetland ponies and 63 donkeys. The ages of the animals ranged between 2 days and 26 years. In both species the eruption of the deciduous and permanent incisors occurred later than in horses. On the other hand, the appearance of the dental stars on the permanent incisors of mini-Shetlands and donkeys was seen at a younger age than in horses. As in most horse breeds, the disappearance of the cups, the clinical crown lengths, the presence of hooks on the upper corner and the presence of a Galvayne's groove are unreliable features for dental age determination in the pony and the donkey. Specific dental characteristics of both the mini-Shetland pony and the donkey are discussed. PMID- 10528537 TI - Studies on a local effect of boar seminal plasma on ovulation time in gilts. AB - Previous studies showed that intrauterine infusion of seminal plasma at the onset of oestrus could advance ovulation in pigs, possibly to enhance the chances of fertilization by optimizing the chronological events of fertilization. This effect has been attributed to a local unilateral mechanism whereby infusion into a single uterine horn advances ovulation in the adjacent ovary. The present study was designed to elucidate possible mechanisms of local signal transduction. In a series of five experiments using 43 gilts, the ovarian response was investigated after infusion of seminal plasma at different sites of the female reproductive tract. The time of ovulation was detected sonographically at 4- or 2-h intervals. Single uterine horn infusion of 100 ml seminal plasma advanced ovulation on the ipsilateral ovary by 9.3 h (mean) compared with the contralateral ovary. Dissection of the ipsilateral isthmus abolished the unilateral seminal plasma effect. Unilateral infusion of 50 microliters or 1 ml seminal plasma or 50 microliters of the concentrated 1-10 kDa fraction in the lower isthmus was ineffective. Application of 5 ml seminal plasma into the tip of a ligated uterine horn lead to 3.6 h (mean) earlier ovulation on the adjacent ovary. In contrast, the infusion of 5 ml NaCl showed no effect. Application of 5 ml seminal plasma in the middle of the uterine horn between two ligatures was ineffective. It is concluded that, for the transduction of the local signal involved in the advancement of ovulation, contact of seminal plasma with the epithelium of the utero-tubal junction is essential. PMID- 10528538 TI - Pharmacological and dietary treatment of canine malabsorption syndrome: a retrospective study of 17 clinical cases. AB - The present study deals with the symptomatology, diagnosis (by means of gastroduodenoscopy and biopsy) and response to a combined dietary/immunosuppressive pharmacological treatment of 17 dogs with malabsorption syndrome. Clinical signs, body weight and serum protein levels evolved favourably in all individuals included in this study during the 150 days following initiation of treatment. PMID- 10528540 TI - Isolates of fungi from symptomatic carthorses in Awassa, Ethiopia. AB - Samples were collected from clinically infected carthorses in Awassa. Fungus species affecting the carthorses were identified. Eight genera of fungal groups were isolated from swabs and skin scrapes taken from symptomatic horses. These included Aspergillus spp., Histoplasma spp., Penicillium spp., Microsporum spp., Trichophyton spp., yeast cells of Candida spp., Cryptococcus spp. and Geotrichum spp. The most frequent isolates were from the genera Aspergillus (48%), Penicillium (39.2%) and Trichophyton (31.6%). Clinical findings are reported, the economic, zoonotic and pathogenetic importance of fungi causing dermatomycoses is discussed, and further studies are recommended. PMID- 10528539 TI - Epidemiology of Streptococcus uberis intramammary infections in a dairy herd. AB - From 1987 to 1991, almost 36,000 quarter samples of mammary secretion representing 1790 lactations of 510 dairy cows from a research herd were collected for bacteriological examination. The percentage of cows infected with Streptococcus uberis ranged from 12 to 16% of cows/year. S. uberis was isolated from 14.2% of lactations over the 5-year period. The prevalence of S. uberis intramammary infection (IMI) was significantly higher in cows with > or = 4 lactations than in cows with 3 or fewer lactations. Regardless of lactation number, the prevalence of S. uberis was highest before parturition, during early lactation and near drying off. The prevalence of S. uberis infected quarters ranged from 1.3 to 2.3% of quarters/year; the prevalence rate for the 5-year period was 2% of quarters. The quarter prevalence of S. uberis was lowest in cows with < or = 3 lactations, increased significantly with lactation number and was highest in cows with > or = 6 lactations. The percentage of quarters infected with S. uberis varied significantly by year. The majority (95%) of S. uberis IMI were subclinical. The ratio of subclinical IMI to clinical IMI was lowest during early lactation, and increased with days in milk, and with lactation age except for cows in their 5th and 6th lactations. Results of this epidemiological investigation suggest that opportunities exist where suitable control measures could be applied to reduce the impact of S. uberis infections in the dairy herd. PMID- 10528541 TI - Restriction endonuclease analysis of BHV-1 and BHV-5 strains isolated in Argentina. AB - The genomes of 10 bovine herpesvirus 1 and 5 strains isolated in Argentina from 1989 to 1994, recovered from animals showing different clinical signs, and two reference strains (Los Angeles and A663) were compared by restriction endonuclease analysis. Four restriction enzymes, HindIII, BamHI, EcoRI and PstI, were used and analysis of the restriction patterns used to assign the isolate to either the BHV-1.1, BHV-1.2 or BHV-5 genotype. There was a correlationship between restriction pattern and clinical signs in six out of ten Argentinian isolates. PMID- 10528542 TI - Serological cross-sectional study of paratuberculosis in cattle in Austria. AB - From 1995 to 1997, the prevalence of serum antibodies against Mycobacterium avium subspecies paratuberculosis (M. av. ssp. ptbc.)--the causal agent of paratuberculosis (Johne's Disease)--was examined in 11,028 Austrian cattle. Samples from the four oldest cattle on 2757 farms were collected according to a specific sampling schedule for this epidemiological study. District, age and breed of animals were included as variables in this study. For antibody screening against M. avium subspecies paratuberculosis, a modified, commercially available ELISA (ALLIED Monitors, Fayette, USA) was employed. A total of 2253 samples that were found to be positive or questionable were subjected to further testing with a more specific ELISA (Institute of Microbiology and Infectious Animal Diseases). Results of this study were used for statistical analysis. The average prevalence of antibodies to M. avium subspecies paratuberculosis was 1.99% in Austria. The highest prevalence was seen in 6-year-old cattle (2.84%) and Holstein Frisian cattle (3.51%). Sero-positive animals were found on 6.96% of farms tested, and the prevalence was highest in Vorarlberg, followed by Salzburg, the Tyrol, Styria and Carinthia. This study is unique in Europe in the use of an adequate random sampling plan for an investigation of this magnitude. PMID- 10528543 TI - Ecology of genus Porphyromonas in canine periodontal disease. AB - Asaccharolytic pigmented Porphyromonas species, including P. endodontalis, P. gingivalis, P. circumdentaria and unclassified species, were isolated from the plaque of adult dogs, but not from any oral sites of puppies and adolescent dogs. With age-dependency, the proportion of Porphyromonas species in the flora of plaque increased. Isolation of the genus Porphyromonas was clearly associated with the progress of periodontol disease. We suggested that Porphyromonas is the exogenous organism and obligate pathogen for canine periodontal diseases. PMID- 10528544 TI - Cow-calf herds in eastern Germany: status quo of some parasite species and a comparison of chemoprophylaxis and pasture management in the control of gastrointestinal nematodes. AB - Infections with gastrointestinal parasites (Eimeria spp., Cryptosporidium spp., Buxtonella sulcata, Fasciola hepatica, Moniezia spp. and trichostrongyles) and lungworms were monitored in five cow-calf herds in the north German lowlands. Estimated prevalences of infections with Eimeria spp. (predominantly Eimeria bovis) ranged between approximately 2 and 48%. The highest prevalences were found during late summer and autumn. On one farm Cryptosporidium spp. were detected in July and August (prevalence: 8.5 +/- 2.7% and 6.7 < 2.0%). The latter finding coincided with diarrhoea in many calves. Buxtonella sulcata was found during the entire study period in highly variable estimated prevalences ranging between zero and 73%, but without any obvious association with clinical disease. Fasciola hepatica was detected on four out of five farms at estimated prevalences of approximately 1-20%. Lungworm infections played a minor role in at least three of five study herds. The estimated prevalence of trichostrongyle infections rose from August until November whereas the intensity of infection did not change significantly. No difference in the intensity of infection could be detected between one farm on which infections with gastrointestinal nematodes were controlled only by moving the animals to an uninfected pasture in July, and three other herds on which strategic anthelmintic control was in place. PMID- 10528545 TI - Does porcine reproductive and respiratory syndrome virus potentiate classical swine fever virus infection in weaner pigs? AB - Fifteen 6-week-old crossbred weaners weighing about 12 kg each were randomly divided into three groups of five animals each. One group of pigs was inoculated first with porcine reproductive and respiratory syndrome (PRRS) virus and then 3 days later with CSF virus. The second group received classical swine fever (CSF) virus, while the third group was inoculated with PRRS virus only. The aim of the experiment was to determine whether a primary PRRS virus infection influences the clinical outcome of experimentally induced CSF in young pigs. The PRRS virus infected weaners developed mild respiratory symptoms and recovered completely. All five weaners which were inoculated with CSF virus only showed severe clinical signs typical of the acute form of CSF. One pig had to be killed 15 days post inoculation (p.i.); the remaining four died between the 18th and 22nd day p.i. The clinical course of the animals inoculated with both viruses was slightly different from that of the pigs that received only CSF virus. Four out of five pigs from the PRRS/CSF group became febrile and viraemic earlier than the animals which received CSF virus only. These pigs had to be killed 15-17 days post CSF virus inoculation. One animal in this group survived the acute phase of CSF and recovered completely. It was concluded that the observed divergences of the clinical courses would not have been noticed under field conditions. Therefore these findings cast doubt on the relevance of PRRS virus infection potentiating significantly the clinical outcome of CSF in young pigs. PMID- 10528546 TI - Nine cases of idiopathic toxic epidermal necrolysis in cattle in Israel. AB - Nine sporadic cases of apparent toxic epidermal necrolysis in nine cattle herds in Israel are presented. The clinical symptoms were characterized by full thickness epidermal exfoliation. No underlying disease or drug administration could be detected in the affected animals, and so all cases were classified as idiopathic. Recovery was noted in all affected animals. The clinical and histological findings are discussed in the light of the pertinent literature. PMID- 10528547 TI - Typing by polymerase chain reaction of buffalo rotaviruses isolated in Italy. AB - Six group A rotaviruses were isolated in Italy from buffaloes during different outbreaks of calf diarrhoea occurring in the same dairy herd over a 5-year period of observation. The isolates were characterized by polymerase chain reaction assay for G- and P-type antigens. G8 and P1 were the types most frequently isolated. PMID- 10528548 TI - Perceived sources of stress in dental students. AB - A 36-item questionnaire was used to investigate the stress perceived by students at Manchester Dental School to potential stressors, grouped under the headings of living accommodation, personal factors, educational environment, academic work and clinical factors. The stressors producing the highest ratings for perceived stress varied throughout the course but high values were allocated to examinations, fear of failing the course or year, shorter and fewer holidays than other university students and--for clinical students--approachability of staff and completing the required quantity and variety of work within a limited time. Female students experienced greater stress than their male counterparts. Since high levels of perceived stress can reduce student performance, dental schools should consider courses in stress management. PMID- 10528549 TI - Orthodontics in the adult patient, with special reference to the periodontally compromised patient. AB - There is increasing demand from adult patients for orthodontic treatment, either purely for aesthetics, to improve aesthetics or function following previous disease, or to facilitate the stabilization, restoration or replacement of teeth. Orthodontics may have a major role in the rehabilitation of patients suffering the effects of advanced periodontal disease, but there are a number of important factors to be considered in the management of such patients if the optimal outcome is to be obtained. This paper summarizes important aspects of treatment and the potential complications and how to avoid them. PMID- 10528550 TI - Inhalation sedation: a viable alternative to general anaesthesia? AB - This paper reports three cases in which young children were treated successfully using inhalational sedation instead of general anaesthesia. PMID- 10528551 TI - Props and pitfalls in oral health promotion in the practice. AB - Practitioners often become demoralized and demotivated with regard to promoting the oral health of their patients as, so often, their efforts appear to be fruitless. This paper examines the published evidence and draws on behavioural theory in order to help practitioners understand what help and support they can offer their patients in order to improve oral health. PMID- 10528552 TI - Is lower incisor extraction treatment a compromise? AB - In this paper orthodontic treatment involving extraction of a lower incisor is described. This has traditionally been regarded as a compromise as loss of a lower incisor renders it impossible to achieve true ideal occlusion. However, with careful case selection, treatment planning and visualization of the post treatment occlusion, excellent functional and aesthetic results can be achieved. The various factors that govern the decision to treat certain malocclusions in this way are discussed and a clinical case is presented to demonstrate this type of treatment. PMID- 10528553 TI - Orofacial disease: update for the dental clinical team: 3. White lesions. AB - White lesions usually contain an increased amount of keratin. Some are rare congenital conditions, such as white sponge naevus and dyskeratosis congenita, unlikely to be seen in general dental practice. Inflammatory causes include candidosis and hairy leukoplakia, both now common in HIV disease. Non-infective causes include the common lesion of lichen planus, and the less common condition lupus erythematosus. Neoplastic and possibly preneoplastic causes include carcinoma, keratoses and leukoplakia. This article discusses the more common causes of oral white lesions. The first article in this series presented several general observations on diagnosis and treatment which should be borne in mind in relation to this article. PMID- 10528554 TI - A viewpoint on the coming impact of emerging diseases. AB - Infectious disease is now the third leading cause of death in the United States, and the No. 1 cause in the Third World. These diseases are the most important public health crisis facing the health care community. As part of that community, dentists must be armed with the knowledge necessary to take their part in the war against these infectious agents. PMID- 10528555 TI - Know thy hepatitis: A through TT. AB - Several viruses have been identified as causative agents of hepatitis in humans. Other hepatotropic viruses have been implicated as potentially disease-causing. This article reviews hepatitis A virus through the newly discovered hepatitis TT virus and their implication for the profession of dentistry. PMID- 10528556 TI - Antibiotic resistance and maxillofacial pathogens: emerging treatment issues. AB - The practice of using antibiotics to treat and control microbial infections is a little more than 50 years old. Widespread administration of multiple classes of antibiotics over the years has had the unfortunate secondary effect of inducing the emergence of an increasing array of drug-resistant microbial strains. This article will discuss the evolution of certain forms of antibiotic resistance, as well as the mechanisms by which bacteria render numerous antimicrobials ineffective. Special emphasis is placed on emerging issues relating to organisms making up portions of the normal oral microflora. PMID- 10528557 TI - The 1997 prevention of bacterial endocarditis recommendations by the American Heart Association: questions and answers. AB - Since the publication of the American Heart Association 1997 recommendations for the prevention of bacterial endocarditis, questions have arisen regarding the application of these guidelines. It is impossible for any such recommendations to include all conceivable clinical situations that might arise, and therefore questions are appropriate. Frequently asked questions are included in this article. Answers provided for the questions are the opinions of the authors, who participated in the formulation of these guidelines, and are not intended to supplant the judgment of the dental health professional who is privy to all the facts when the individual clinical decision is made. PMID- 10528558 TI - Current status of fenfluramine/dexfenfluramine-induced cardiac valvulopathy. AB - Since publication of the U.S. Department of Health and Human Services' interim recommendations in November 1997 for the management of patients having taken certain appetite suppressants, a number of studies evaluating the prevalence of cardiac valvular pathology in such individuals have been published. These studies generally support the association of fenfluramine/dexfenfluramine with cardiac valvulopathy but with significant differences in risk assessment. The analysis of these studies has produced two new guidelines for the management of such patients, including the appropriate use of antibiotic prophylaxis in these individuals. These studies are presented along with a comparison of the three present recommendations and their impact on dental practice. PMID- 10528560 TI - Dental treatment and bacterial endocarditis. PMID- 10528559 TI - Clostridium difficile-associated diarrhea and colitis. AB - Clostridium difficile-induced diarrhea (CDAD) and colitis (CDAC) are important nosocomial (hospital)-acquired infections resulting almost exclusively from antibiotic therapy and certain host factors. The severity of these disorders may range from simple diarrhea that can be resolved easily with antibiotic cessation to fulminant pseudomembranous colitis with fever, severe dehydration, abdominal pain and distention, and plaque formation over part or all of the colon. Community-acquired CDAD and CDAC are far less problematic but nevertheless may affect 20,000 or more people in the United States every year. Knowledge of the risk factors for CDAD and CDAC, including certain antibiotics, and recognition of the entire spectrum of signs and symptoms of this disorder are imperative for good dental practice. Likewise the prevention of recurrence of CDAD by judicious use of antibiotics in its immediate posttreatment period is an important consideration. PMID- 10528561 TI - Periapical diseases: spectrum and differentiating features. AB - There are a variety of lesions besides the typical granulomas and cysts that can appear at the apices of teeth. These other lesions must receive consideration in the diagnosis of periapical disease because of their potential impact on patient treatment and outcome. This paper will review the spectrum of diseases that may present in periapical tissues, the pathogenesis, of periapical inflammatory disease, and the signs and symptoms that separate periapical inflammatory disease from neoplastic disease. PMID- 10528562 TI - The diagnosis and management of chronic nonspecific mucosal lesions. AB - Chronic lichenoid or leukoplakic oral mucosal lesions are a common cause of morbidity and concern. Many of these are reactive or inflammatory lesions; but they can also represent other disorders, including dysplasia. These lesions are caused by a variety of irritants and allergens such as systemic drugs, dental restorations and prostheses, oral health care products, foods, habits, and candida. Indiscriminate use of steroids to treat these lesions empirically is contraindicated; management should be aimed at discovery and removal of the cause. The proposed sequence of investigation is intended initially to eliminate any obvious etiology and, if that is unsuccessful, to rule out candida or determine a histologically diagnosable disease. If a biopsy shows nonspecific or lichenoid mucositis and the lesions are symptomatic, the investigation is directed to the more obscure and speculative causes that require expensive and time-consuming trial-and-error elimination of various agents. PMID- 10528564 TI - Oral manifestations of gastrointestinal disease. AB - A variety of gastrointestinal diseases can be associated with lesions of the oral cavity. The lesions usually correlate to active intestinal disease, but they may present prior to any other evidence of the disease and even be used to initiate diagnosis and treatment. This paper reviews the more common oral manifestations of gastrointestinal disease and their dental management. PMID- 10528563 TI - Proliferative verrucous leukoplakia: report of two cases and a discussion of clinicopathology. AB - Proliferative verrucous leukoplakia (PVL) is a recently delineated but poorly recognized form of multifocal leukoplakia that is premalignant and of unproven origin. PVL generally presents as a simple benign form of hyperkeratosis that tends to spread and become diffuse. Although slow-growing, the disease is persistent and irreversible. Clinically, PVL often presents as an exophytic wart like form of leukoplakia that appears to be resistant to nearly all forms of therapy. PVL of the oral cavity is best-defined as a continuum of oral epithelial disease with hyperkeratosis at one end of a clinical and microscopic spectrum and verrucous carcinoma or squamous cell carcinoma at the other. The microscopic findings associated with PVL are dependent on the stage of the disease and the adequacy of the biopsy. Microscopic findings can be markedly variable. PVL is a clinicopathologic disorder that includes the microscopic entity known as verrucous hyperplasia as a component of its histopathologic progression. This article reports on two cases of PVL, describes the clinicopathology of the disease process, and presents therapeutic and etiologic considerations. PMID- 10528565 TI - Prevention and detection of lip cancer--the dentist's role. AB - With their attention to the oral area, dentists are in an excellent position not only to diagnose lip cancer, but also to counsel patients in its prevention. Patients need to be educated on the dangers of ultraviolet radiation and the measures available to decrease exposure to it. This article discusses the circumstances that increase the chance of developing lip cancer, the variety of ways to decrease that chance, and the recognition and treatment of premalignant and malignant lip lesions. PMID- 10528566 TI - When the president vanished. PMID- 10528567 TI - Oral health status of special athletes in the San Francisco Bay Area. AB - A standardized oral health screening protocol was developed for assessing the oral health status of athletes participating in annual Special Olympics events at sites across the country. This paper reports on results at the San Francisco Bay Area Special Olympics event, where 385 athletes participated in the oral health screening. Trained dental screeners determined the presence or absence of edentulism, untreated decay, filled teeth, missing teeth, tooth injury, fluorosis, and gingival signs, as well as treatment urgency. The frequency of mouth cleaning, having a mouth guard, use of tobacco, and presence or absence of pain were self-reported. Overall, child athletes 9-20 years of age had more untreated decay and substantially more missing permanent teeth than 9-20-year-old children represented in the 1986-87 National Institute of Dental Research Survey of U.S. School Children. Prevalence of missing teeth among adult athletes compared favorably with data from the Third National Health and Nutrition Examination Survey and the Behavioral Risk Factor Surveillance System Survey. Approximately one-third of child and adult athletes were determined to need dental care. Continued use of a standardized screening protocol could allow state specific data to be available on the oral health status of this population; trends could be tracked; and needs could be identified, with strategies developed to meet those needs. PMID- 10528568 TI - Prevalence of spit tobacco use across studies of professional baseball players. AB - A review of published data, together with previously unpublished information, shows that the use of spit tobacco among professional baseball players continues to be alarmingly high. In spite of efforts to make players aware of the harmful effects, approximately 35 percent to 40 percent of professional baseball players still use spit tobacco, and about half of those have associated lesions. However, current efforts of the National Spit Tobacco Education Program, Major League Baseball, the Professional Baseball Athletic Trainers Society and the Major League Baseball Players Association are expected to result in a significant reduction in spit tobacco use in this population in the next decade. PMID- 10528569 TI - A dental-based, athletic trainer-mediated spit tobacco cessation program for professional baseball players. AB - During 1997 spring training, the National Spit Tobacco Education Program provided a spit (smokeless) tobacco intervention program to 16 professional baseball clubs. The program consisted of an awareness-raising presentation and an opportunity to discuss quitting spit tobacco use with an expert cessation counselor. For two clubs, however, a more extensive intervention was pilot-tested for feasibility and acceptability among their major- and minor-league teams during their regularly scheduled health examinations at the beginning of spring training. The intervention included an oral exam by a dentist who advised spit tobacco users to stop and pointed out any spit tobacco-associated lesions in the player's mouth, brief cessation counseling by a specially trained dental hygienist, and ongoing support and follow-up by the certified athletic trainer to promote cessation. Findings from this pilot study indicate that this intervention, which is dependent upon involvement of dental professionals, was feasible to implement during spring training and appeared to be well-received by the athletes. Dental professionals are in an excellent position to advise and help spit tobacco users to quit and can have an important role in helping youth overcome this rapidly growing addiction. PMID- 10528570 TI - A contemporary perspective on dental sealants. AB - In spite of significant improvements in the oral health of Americans, dental caries still affects a majority of school-aged children. Its distribution is uneven, with a small proportion of the children experiencing a greater burden of the disease. In addition, caries in children's permanent teeth is predominantly a disease of the pits and fissures. The use of dental sealants has the potential to significantly reduce the disease burden. Although sealants are safe and effective, their use continues to be low. Efforts are needed to make sealants a covered benefit under all insurance plans and to encourage their appropriate use. This paper provides a review of the changes in the prevalence and distribution of dental caries, the effectiveness of sealants, and guidelines for the appropriate use of sealants in public health programs and private practice. PMID- 10528571 TI - Addressing the needs of underserved populations: one organization's experience. AB - Dental decay is the most prevalent and preventable chronic disease of childhood. Underserved populations are at a health disadvantage with greater unmet needs. This article will discuss the components of oral health promotion programs and facilities designed to meet the needs of underserved populations. These components include organization, needs assessment, resource assessment, priority setting and planning, oral health intervention, and monitoring and evaluation. Examples from the experience of the San Diego Children's Dental Health Association will be presented in the discussion of each component. PMID- 10528572 TI - Dientes! Community dental clinic: dental care for low-income residents of Santa Cruz County. AB - Dientes! is a private nonprofit community dental clinic that was established in 1994 to provide dental care for low-income residents of Santa Cruz County. Its founders were successful in securing support from a diverse group of community agencies, including city and county governments, philanthropic foundations, the dental community, and corporate and individual donors. Dientes! provides approximately 250 visits per month in a three-chair clinic in Santa Cruz; a school-based program in Watsonville began March 1998. The major challenge facing Dientes! is to establish a reliable financial base that will allow the program to better meet the needs of low-income county residents over the long term. PMID- 10528573 TI - Dental care for the underserved children of Monterey County: meeting the challenge. AB - With its expansive area, and the special needs of agricultural workers, Monterey County held significant challenges for setting up a children's health clinic. Part of the solution to addressing the county's unmet dental needs was the establishment of the Children's Miracle Network dental center in 1995. But working in the fields leaves little time for travel to appointments, so the dental center expanded to a mobile unit that can go where the need is. Understanding the special needs of one's community is crucial to establishing programs that can successfully address the state's needs for children's dental care. PMID- 10528574 TI - The Children's Dental Center--a community resource to meet a community need. AB - Growing numbers of children of working poor families in California have limited access to dental care. This article presents a unique solution to this problem: the Children's Dental Center. The center, a nonprofit corporation, emphasizes quality multidisciplinary care, aggressive preventive dental practices, and education programs for parent and child. Through behavioral change, coupled with dental care of urgent problems, the family's immediate needs are addressed while creating a future of diminished dental need and greater self-esteem. PMID- 10528575 TI - San Gabriel Valley Foundation for Dental Health: a hand up, not a handout. AB - The San Gabriel Valley Foundation for Dental Health Clinic was established to offer reduced-fee health care to the needy. The basic tenets of the clinic are to minimize dental disease by teaching prevention and treat the dental needs of the disadvantaged, while teaching responsibility for the cost of dental care. Beneficiaries of the clinic include patients, dental assisting students, volunteer dentists, and organized dentistry. PMID- 10528577 TI - Chartbook. Quality checkup. AB - With hospital quality programs, the reach is broad, not deep. So says the National Hospital Quality Improvement Survey, conducted by health care researcher Stephen Shortell for Arthur Andersen and the American Hospital Association. Though 93 percent of responding hospitals and health systems use CQI and TQM methods is some fashion, they've taught them to only 35 percent of employees and a scant 22 percent of doctors on their medical staffs. Those poor showings leave health care lagging behind most other industries. PMID- 10528576 TI - Improving oral health for people with special needs through community-based dental care delivery systems. AB - A community-based dental care delivery system is described. This system has been used in a number of communities in California to improve oral health for people with special needs. It includes oral health assessment, coalition building, development and networking of local resources, training of dental professionals, and utilization of preventive dentistry training materials. Also discussed are challenges of the future that will need to be met to continue to make oral health a priority and reality for people with special needs in California. PMID- 10528578 TI - Fast food. Interview by Chris Serb. PMID- 10528579 TI - The future of the future. PMID- 10528580 TI - The ratings slide. Are we headed for a capital crisis? AB - On hospital credit ratings, the news is tough and tougher--what many industry watchers call a capital crisis in the making. Rating agencies contend the situation has been building for years, even while hospital margins reached historic highs as recently as 1997. "Then everything fell apart at the seams," says one bond rating pro. PMID- 10528581 TI - Old tech gets hip again. AB - Telemedicine goes deep in the heart of Texas. Pagers get more persistent in St. Louis. And an e-network spans Spokane. From idea to method to gadget, innovative communication in today's digital economy typically involves all three. In each of our three profiles, organizations have pushed old boundaries--and bumped against new ones--in their search for a speedier response, better care, and lower costs. Each added value by putting existing technology to a new use, not the other way around. They all broke new ground by pairing developers with the people taking advantage of their innovations. And all agree: They're hardly finished yet. PMID- 10528582 TI - What you see/what you get. PMID- 10528583 TI - More than medicine. Interview by Chris Serb. AB - For all the business savvy demanded by their work as hospital CEO, info tech chief, or entrepreneur, these doctors say it's their medical background that gives them an edge. They know the needs and routines of physicians firsthand. They come to their jobs with built-in credibility as clinicians. And they understand what it means to care for patients. "It's hard for me to know how we're doing," says one hospital CEO, "if I'm sitting in my office all the time." PMID- 10528584 TI - Revenge of the reviewed. AB - The managed care backlash has shifted to a new target: HMO medical directors. Federal law shields employer-sponsored health plans from most state oversight and lawsuits, but angry enrollees have begun taking a back route, turning state medical boards on doctors in charge of utilization review. One attorney calls them "the soft underbelly of managed care." PMID- 10528585 TI - Selling brand MD. AB - Health care is more than local, of course: It's personal. Recognizing the connections between doctors and patients as a potent form of brand loyalty, health systems are looking for ways to leverage those bonds--and encourage new ones--with special marketing campaigns. PMID- 10528586 TI - [Can we really reach our goals? Arguments for aggressive lipid lowering]. PMID- 10528587 TI - [51st Ophthalmology Session on the occasion of commemoration of the centenary of the death of Alfred Graefe (1830-1899). University Clinic of Ophthalmology, Halle, 20 March 1999]. PMID- 10528588 TI - [Quality management system of the Ophthalmology Clinic at the Polyclinic of the Erlangen-Nurnberg Friedrich Alexander University, Erlangen. Certification according to DIN EN ISO 9001]. PMID- 10528589 TI - [Parkinson disease. Evidence based medicine for Levodopa]. PMID- 10528590 TI - [Health economy. Social and economic aspects of depression therapy]. PMID- 10528591 TI - [14th Congress European Association of Urology, Stockholm, Sweden. Therapy of advanced cancer of the bladder]. PMID- 10528592 TI - Critical analysis of injuries sustained in the TWA flight 800 midair disaster. AB - BACKGROUND: Previous reports of commercial airline disasters have reviewed incidents occurring at takeoff and landing. The purpose of the present study, which represents the first analysis of aviation injuries incurred during a midflight incident, was to examine the injuries sustained by the victims of the TWA Flight 800 disaster and to determine any correlation of injuries with structural damage and seat location. METHODS: Complete autopsy records, toxicology screening, and forensic analysis were reviewed. Injuries were assessed by anatomic region and severity by using the Abbreviated Injury Scale. The National Transportation Safety Board report of the investigation was applied to correlate individual injuries with seat location and structural damage. A comparison was performed against injury data from takeoff and landing incidents. RESULTS: All 230 passengers of TWA Flight 800 were recovered as fatalities. Head, thoracic, and abdominal injuries were multiple and severe, contributing to the mortality of the occupants. Analysis revealed that the severity of injury and anatomic injury pattern did not generally correlate with seating position or structural damage. A comparison of these injuries with those of takeoff and landing crashes showed differences in injury pattern and severity. CONCLUSION: Passengers of Flight 800 sustained instantaneous fatal blunt force injury. Analysis of the data revealed no global correlation between seat position and pattern of injury. In contrast to injuries incurred during crashes at takeoff and landing, these midflight injuries were too extreme to warrant a reappraisal of current passenger protective safety measures or standards. PMID- 10528593 TI - Evisceration after abdominal stab wounds: is laparotomy required? AB - OBJECTIVES: To determine the incidence of intra-abdominal injury requiring laparotomy after an abdominal stab wound with evisceration. To identify clinical signs that increase the likelihood of an intra-abdominal injury in the presence of such a wound. METHODS: Information was collected prospectively over an 8-year period on all patients who presented to our urban level I trauma center with an abdominal stab wound and evisceration. This information included which organ eviscerated, presence of other indications for laparotomy, organs injured, and postoperative complications. All comparisons used the Fisher's exact chi2. RESULTS: A total of 81 patients were admitted with evisceration after an abdominal stab wound. Sixty-one patients (75%) had eviscerated omentum, 18 patients (22%) had eviscerated small bowel, and 2 patients (2%) had eviscerated colon. Sixty-two patients (76%) had evisceration as the sole indication for laparotomy, the remaining 19 patients (24%) had another indication such as hypotension or peritonitis. Overall, 63 patients (78%) had an intra-abdominal injury that required repair. This was true regardless of organ eviscerated (omentum = 77% vs. viscus = 80%, not significant) or clinical presentation (no other indication = 76% vs. another indication = 84%, not significant). CONCLUSION: The majority of patients who present with an evisceration after a stab wound to the abdomen require a laparotomy. This is true regardless of what has eviscerated or the presence of other clinical indications to operate. Evisceration should continue to prompt operative intervention. PMID- 10528594 TI - Use of an Objective Structured Clinical Examination (OSCE) for the assessment of physician performance in the ultrasound evaluation of trauma. AB - BACKGROUND: A reliable means of assessing physician competency in performing ultrasound (US) is critical for training and credentialing. Objective Structured Clinical Examinations (OSCE) have been used successfully to assess clinical competency in other areas of surgical education but have not been applied previously to trauma ultrasound training. The objectives of this study were to assess physician performance in the focused abdominal sonography in trauma (FAST) examination by using a specifically designed OSCE, and to determine whether the OSCE detects differences in two determinants of competency (knowledge acquisition and clinical interpretation skills). METHODS: Eighty-two physicians in surgery (n = 49) and emergency medicine (n = 33) at a Level I trauma center were evaluated. All participated in a FAST course consisting of didactic sessions on US physics, indications, and technique, FAST examination videos, and a hands-on session with human models. The OSCE consisted of two parts: written examination that assessed factual knowledge, and videotape of real-time US examinations that assessed interpretation skills. The OSCE was administered before and after the FAST course. RESULTS: Significant improvements in postcourse OSCE scores were observed for factual knowledge (52.5 +/- 2.0 vs. 87.5 +/- 1.1, p < 0.001) and interpretation skills (27.2 +/- 1.4 vs. 62.9 +/- 1.3, p < 0.007). Scores for US interpretation were significantly lower than those for factual knowledge at both precourse (27.2 +/- 1.4 vs. 52.5 - 2.0, p < 0.001) and postcourse (62.9 +/- 1.3 vs. 87.5 +/- 1.1, p < 0.01). No performance differences were observed between surgeons and emergency medicine physicians and no effect of training level on test scores was observed. CONCLUSION: Knowledge acquisition and US interpretation skills can be assessed reliably with a specifically designed OSCE. Although both skills improved after participation in a FAST course, US interpretation scores were consistently lower than those for factual knowledge. This study supports the use of the objective structured clinical examination in both the design of ultrasound teaching programs and the assessment of physician competency. PMID- 10528595 TI - Prospective evidence of the superiority of a sonography-based algorithm in the assessment of blunt abdominal injury. AB - BACKGROUND: Although the routine use of FAST (focused assessment with sonography for trauma) in the evaluation of trauma victims is increasing, to our knowledge, a prospective comparison of contemporary adult trauma victims managed with and without FAST has not been reported in North America. METHODS: Adult victims of blunt trauma for whom there was a suspicion of abdominal injury were managed with one of two diagnostic algorithms, FAST or no-FAST. The two algorithms were compared for diagnostic accuracy, cost, time, and delayed diagnoses. RESULTS: Among 706 patients (mean Injury Severity Score, 23), 460 were managed with FAST and 246 with no-FAST. The two groups were similar with respect to age, Injury Severity Score, prehospital time, and mortality (p = not significant). There were 3 of 460 (0.7%) delayed diagnoses in the FAST group and 4 of 246 (1.6%) in the no FAST group (p = not significant). The diagnostic accuracy for the FAST and no FAST algorithms was 99% and 98%, respectfully. The FAST and no-FAST algorithms led to similar rates of laparotomy, 13% and 14%, respectfully, but nonoperative management was more common in the no-FAST group (p < 0.01). The mean diagnostic cost for the FAST algorithm was $156, compared with $540 with the no-FAST algorithm (p < 0.0001) and the mean time required for diagnostic work-up was 53 minutes with the FAST algorithm, compared with 151 minutes with the no-FAST algorithm (p < 0.0001). CONCLUSION: This study has provided prospective evidence that a FAST-based algorithm for blunt abdominal injury was more rapid, less expensive, and as accurate as an algorithm that used computed tomography or diagnostic peritoneal lavage only. Trauma centers are encouraged to incorporate a FAST-based algorithm into their initial management of blunt trauma victims. PMID- 10528596 TI - Early detection of arterial bleeding in acute pelvic trauma. AB - OBJECTIVE: To determine the accuracy of intravenous contrast-enhanced computerized tomography (CECT) in the detection of potentially life-threatening retroperitoneal hemorrhage in patients sustaining pelvic fractures, acetabular fractures or both. DESIGN: Retrospective review of sequential patients identified over a 1-year period by using a prospectively collected trauma database at two Level I trauma centers. MATERIALS AND METHODS: A group of patients admitted to one of two Level I trauma centers with pelvic or acetabular injuries between September 1, 1995, and September 30, 1996, was identified by using a prospectively collected trauma database. From this cohort, we selected those individuals who had undergone intravenous CECT scanning within 24 hours after admission and who had an Abbreviated Injury Score more than 3 because of their pelvic injury. Those individuals who required arterial embolization for uncontrolled hemodynamic shock were categorized as having "significant arterial bleeding" attributable to their pelvic injury. Individuals who regained hemodynamic ,stability without embolization were categorized as having "no significant arterial bleeding." Two observers who were blinded to clinical information and the results of angiography reviewed all injury radiographs and computed tomographic scans. The presence or absence of contrast extravasation on intravenous CECT was recorded. Each case was then categorized into a 2 x 2 table depending on the presence of contrast extravasation on CECT and the need for arterial embolization to determine the accuracy of the "contrast extravasation sign." RESULTS: Of the 192 eligible patients, 111 met the inclusion criteria. Eleven patients required an angiogram for ongoing hemodynamic instability. The sensitivity of extravasation on contrast enhanced computed tomography representing a significant arterial bleeding was 80%, and the specificity was 98%. The predictive value of a positive contrast "extravasation sign" was 80%, whereas the predictive value of a negative test was 98%. The likelihood ratio of a positive test was 40.4, and the likelihood ratio of a negative test was 0.204. CONCLUSION: The finding of contrast extravasation on CECT is highly suggestive of significant arterial bleeding that requires early angiographic embolization to restore hemodynamic stability. PMID- 10528597 TI - Urban free falls and patterns of renal injury: a 20-year experience with 396 cases. AB - OBJECTIVE: To determine the distribution and stage of renal injuries from free falls and to determine the appropriate methods for their evaluation and management. MATERIALS AND METHODS: We reviewed the records of 423 patients with renal injuries after a fall from height. Twenty-seven patients did not survive their injuries and were removed from the study. RESULTS: Based on the American Association for the Surgery of Trauma grading scale, 372 of the renal injuries (94%) were grade 1, whereas 24 injuries (6%) were grade 2 to 4. None of the injuries was grade 5. Of the patients with grade 2 to 4 renal injuries, nine patients had grade 2, three patients had grade 3, nine patients had grade 4, and one patients had a forniceal rupture, as well as two patients with ureteropelvic junction disruptions (one bilateral), four with segmental vascular injuries, and two with hilar vessel injuries. Mean height of free fall was 23.1 feet (range, 10 60 feet) and mean Injury Severity Score was 20.6. Neither the degree of renal injury nor the Injury Severity Score statistically correlated to the height of the free fall. Patients with grade 2 to 4 were more likely than patients with grade 1 renal injuries to be in shock and to have intra-abdominal injuries, gross hematuria, and higher Injury Severity Score(33%, 34%, 62%, 24.6 vs. 6%, 9%, 14%, 20.1, respectively). The degree of hematuria and the grade of renal injury, however, did not correlate. Grade 2 to 4 renal injuries had microscopic hematuria and no shock in 8.3% (2 of 24 patients) and no hematuria in 20.8% (5 of 24 patients). Thus, standard selection criteria for renal imaging of blunt trauma, namely gross hematuria or microhematuria and shock would have missed 7 or 29% of our grade 2 to 4 renal injuries, or 1.8% of all grade 1 to 4. Half of the patients with grade 2 to 4 renal injuries had associated multiple-system injuries, and half had flank ecchymosis or tenderness. Of the patients with grade 2 to 4 injuries, 9 patients (37%) underwent surgical exploration and repair of injury. All renal units were preserved and underwent successful reconstruction. Six of the nine patients initially were explored because of associated intra abdominal injuries. No major urological sequelae were noted postoperatively or in follow-up of all renal injuries. CONCLUSION: The height of the free fall cannot reliably predict the degree of the resulting renal injury. Despite the absence of hematuria or shock, vertical deceleration injuries, in particular those associated with multiple-system injuries and/or physical signs of potential renal injury (e.g., flank ecchymosis), demand renal imaging. After a fall from height, the ureteropelvic junction and renal vasculature should also be imaged for potential injury. PMID- 10528598 TI - Overuse of splenic scoring and computed tomographic scans. AB - BACKGROUND: As the most commonly injured abdominal organ in blunt trauma, the management of splenic injury has undergone evolution. The risk of blood transfusions administered in an attempt to save the spleen has lowered the threshold for operation and also expanded the limits for nonoperative management. An in-depth analysis was carried out of risk factors on patients requiring immediate surgery and those who fail non-operative management based on organ injury scaling grading by computed tomographic (CT) scan and operation. The application of nonoperative management in the elderly population and the use of follow-up CT scanning and sonography in the outpatient setting was also examined. METHODS: Between January of 1991 and June of 1996, 226 consecutive blunt splenic trauma, injured patients at a Level I trauma center were evaluated. All subsequent CT scans and sonograms in the inpatient and outpatient setting were analyzed. The Student's t test, Pearson chi2 analysis with Yates correction, and analysis of variance were used to compare between and among groups. RESULTS: There were 153 men (67.7%), an average age of 34.8 years, an average Injury Severity Score of 24.4, and 28 deaths (12%). There was a significant difference with respect to Injury Severity Score, Glasgow Coma Scale score, Revised Trauma Score, units of packed red blood cells transfused, length of stay, intensive care unit length of stay, mean splenic injury grade, and cost between patients observed initially and those operated on initially. There was no significant difference in age between the two groups. Of 170 patients, 37 patients (22%) who had an initial CT scan underwent immediate exploratory laparotomy. The remaining 133 patients (78%) had nonoperative management; however, 15 patients (11%) failed the period of observation. Five in this group had a laparotomy secondary to other causes and another six were operated on within 24 hours of their injury for their splenic injury. Thus, only four of the nonoperative management patients (3%) actually failed nonoperative splenic management after 24 hours of injury. There were 100 second CT scans obtained. Three of these patients, who had developed hemodynamic instability, required operation for a bleeding spleen. The subsequent CT scan was confirmatory in these three patients who resided in the intensive care unit. All other CT scans and sonograms for clinically unremarkable patients failed to yield any alteration in care based on the scans. CONCLUSION: Blunt splenic injured patients can be safely observed; however, there are certain risk factors in those requiring immediate surgery and those failing nonoperative management. The CT scan underestimates injury, possibly related to a progression of bleeding found at the time of operation. No outpatient studies altered the course of management. Age also did not influence outcome. Thus, in the dedicated trauma center, nonoperative management of blunt splenic injury patients does not lead to undue morbidity or mortality. Once discharged, follow-up radiographs in asymptomatic patients are not necessary. PMID- 10528599 TI - Use of hypertonic saline/acetate infusion in treatment of cerebral edema in patients with head trauma: experience at a single center. AB - BACKGROUND: Hypertonic saline (HS) recently has been introduced as a new form of hyperosmolar treatment in patients with brain injury from diverse causes. We reviewed our experience with the use of continuous hypertonic saline/acetate infusion in patients with cerebral edema attributable to head trauma. METHODS: We performed a retrospective chart review of all patients admitted with severe head injury, defined as admission Glasgow Coma Scale score of 8 or less, in the neurocritical care unit of a University hospital. Intravenous infusion of 2% or 3% saline/acetate for treatment of cerebral edema was introduced in the unit in April of 1993. The clinical characteristics, interventions required, and outcomes in patients who received HS were compared with patients who received 0.9% saline infusion only. Multivariate analyses were used to evaluate the impact of HS use on in-hospital mortality and Glasgow Outcome Scale score at discharge. RESULTS: Thirty-six patients with cerebral edema caused by head trauma received infusion of HS initiated within 48 hours of admission for a mean period of 72 +/- 85 hours. Compared with 46 patients who did not receive HS, there were no differences observed in age and admission Glasgow Coma Scale scores. Patients who received HS were more likely to have a penetrating injury (p = 0.07) and a mass lesion on initial computed tomographic scan (p = 0.07). There was no difference between frequency of use of hyperventilation, mannitol, cerebrospinal fluid drainage, and vasopressors between the two groups. The requirement for pentobarbital coma was higher in HS group (n = 7 patients) versus control group (n = 2,p = 0.04). After adjusting for differences between both groups, infusion of HS was associated with higher in-hospital mortality (OR, 3.1; 95% CI, 1.1 10.2). CONCLUSION: HS administration as prolonged infusion does not seem to favorably impact on requirement for other interventions and in-hospital mortality in our experience. Further efforts should be directed toward use of HS as bolus administrations or short infusions. PMID- 10528600 TI - Changes in left ventricular performance in patients with severe head injury during and after mild hypothermia. AB - OBJECTIVE: To evaluate left ventricular (LV) performance in patients with severe head injury during and after mild hypothermia. PATIENTS AND METHODS: Seven consecutive patients who underwent therapeutic mild hypothermia (age, 15 to 70 years; Glasgow Coma Scale score on admission, 4 to 8). LV performance was assessed by using M-mode, color tissue Doppler imaging tracings and pulsed Doppler echocardiography. LV contraction and relaxation were evaluated by using the peak velocity of LV posterior wall movement during systole (Smax) and diastole (Dmax), respectively, in addition to the conventional echocardiographic indices. RESULTS: Mild hypothermia increased LV ejection time and reciprocally reduced LV filling period as indicated by temperature-dependent shortening of the early diastolic filling and the total diastolic inflow time. The indices depending on temporal factors such as ejection time, Smax, or Dmax were significantly affected by mild hypothermia, whereas those depending on spatial factors such as fractional shortening or stroke volume index were not. The attenuated Smax was compensated for the prolonged ejection time resulting in the relatively consistent fractional shortening regardless of body temperature. There was no compensatory mechanism for the decreased Dmax during diastole. CONCLUSION: The effect of mild hypothermia seemed to be predominantly negatively chronotropic. LV diastolic function was more vulnerable to mild hypothermia than LV systolic function was. PMID- 10528601 TI - Adenosine-triphosphate in trauma-related and elective hypothermia. AB - BACKGROUND: In trauma patients, hypothermia is a frequent event. According to the literature, the majority of trauma patients are presenting a core temperature of less than 34 degrees C at admission. In contrast to the benefit of hypothermia in elective surgery, clinical experience with hypothermia in trauma patients has identified hypothermia to be one major cause of severe posttraumatic complications. It was hypothesized that this diverse effect of hypothermia is related to depletion of high-energy phosphates like adenosine triphosphate (ATP) in trauma patients. To verify this hypothesis, the relation of ATP plasma levels and hypothermia was examined in a clinical study. METHODS: Three different groups of patients were under study. The first group (group A, normothermic control group) included patients (n = 15) undergoing elective surgery of the lower limb with a mean operation time of 113 minutes. The second study group (group B, hypothermic control) was composed of patients (n = 15) who were subjected to elective coronary artery bypass operation under hypothermia (31 degrees C for 48 minutes, mean total operation time being 205 minutes). The third study group (group C) included trauma patients (n = 23, mean Injury Severity Score [ISS] of 24.7). At the time of admission, 10 patients presented a core temperature more than or equal to 34 degrees C (group C1, mean ISS, 25.2; mean T(A), 34.5 degrees C), 13 patients presented a T(A) less than 34 degrees C (group C2, mean ISS, 26.0; mean T(A), 32.9 degrees C). In both groups of surgical patients, the ATP plasma level was measured preoperatively, at 2, 4, and 24 hours postoperatively. For trauma patients, this measurement was performed at admission and 24 hours later. Within the same schedule, body core temperature was recorded and the clinical course was documented as well. RESULTS: Elective limb surgery in normothermic patients resulted only in a transient decrease in ATP plasma levels (preoperative, 87.8 micromol/dL; 4 hours postoperative, 52.0 micromol/dL). At 24 hours, the ATP plasma level (62.6 +/- 10.0 micromol/dL) has increased toward baseline level. Elective hypothermia in patients subjected to coronary bypass also resulted only in a transient decrease in ATP plasma levels. During the operation period, including hypothermia, the ATP plasma level was comparable (50.4 micromol/dL) to group A and also returned back toward normal values at 24 hours (58.2 micromol/dL). All trauma patients revealed a significant low ATP plasma level at admission compared with both control groups. Looking at subdivided groups the most significant drop in ATP plasma level (28.5 micromol/dL) was noted in patients presenting an initial core temperature less than 34 degrees C and ISS more than 30. Even 24 hours later, the ATP level of this subgroup was significantly diminished, despite a rise up to 44.4 micromol/dL. In contrast, only a moderate drop in ATP plasma concentration (59.2 micromol/dL) was noted in the group of T(A) more than or equal to 34 degrees C and ISS less than 20. This group revealed almost normal values (68.3 micromol/dL) 24 hours after trauma. In addition to hypothermia, the metabolic state, reflected by the plasma lactate levels, significantly influenced the ATP plasma levels, as high lactate levels were paralleled by low ATP levels. Also, the overall outcome was related to injury severity and hypothermia. CONCLUSION: Hypothermia in elective surgery, established by active cooling, preserves the ATP storage and maintains an aerobic metabolism, which both contribute to the beneficial effect of hypothermia in ischemia/reperfusion in cardiovascular surgery. However, in trauma patients hypothermia is caused by insufficient heat production due to utilization of ATP under anaerobic metabolic conditions. Low ATP plasma levels combined with hypothermia seem to be a predisposition for post-traumatic complications like organ failure. PMID- 10528602 TI - National survey of the incidence of cervical spine injury and approach to cervical spine clearance in U.S. trauma centers. AB - BACKGROUND: The overall incidence of cervical spine injury (CSI) has been estimated from small studies; the incidence of specific injury types is less well established. The approach to screening for CSI has not been well studied; variation may exist based on Trauma Center (TC) level and type (academic vs. nonacademic). We attempted to define the incidence of different types of CSI and determine whether a national standard for cervical spine clearance (CSC) could be identified. We hypothesized a significant variation in incidence of CSI and approach to CSC based on TC level and type. METHODS: In a survey of 615 TC, institutions were asked to describe themselves as academic/nonacademic and provide a Level I-IV. Questions concerned demographics, Injury Severity Score, incidence of CSI, clinical resources, and approach to CSC. Methods of CSC included protocols, use of flexion-extension films, computed tomography, magnetic resonance imaging, and cervical collars. Clinical scenarios examined indications and technique for CSC. RESULTS: A total of 637 surveys were sent to 615 TC (25 follow-ups), and 165 TC (25%) responded. A total of 156 TC provided data for type: academic 44 (28%), nonacademic 112 (72%). A total of 142 TC provided data for level: 49 (34%) Level I, 75 (53%), Level II, 18 (13%), Level III. A total of 111,219 patients were entered into the trauma registries of these TC. The overall incidence of all types of CSI was 4.3%, CSI without spinal cord injury was 3.0%, spinal cord injury without fracture was 0.70%, and delayed diagnosis of all types of CSI was 0.01%. There was no difference in the incidence of CSI overall or by subtype based on TC level or type. Injury Severity Score correlated with incidence of CSI without cord injury (r = 0.387, p < 0.01). Regarding approach to CSC, differences existed by TC level and type for responsibility for CSC and protocols for CSC (p < 0.05). Level II TC felt early flexion-extension views were potentially harmful (60%); Level I TC did not (39%) (p < 0.05). Regarding indications for CSC, there was agreement on 10 of 11 clinical scenarios. For three of five clinical scenarios examining radiographic approach to CSC there was a broad distribution of approaches to patients with normal radiographs and cervical pain, altered mental status, coma. CONCLUSION: Incidence of CSI is uniform by TC level and type. Incidence of spinal cord injury without fracture is low: 0.7%. Reported rate of missed CSI is very low: 0.01%. There is good agreement (>78%) among TC on indications for CSC but less agreement on radiographic approach to CSC. PMID- 10528603 TI - Multiple organ dysfunction after remote circulatory arrest: common pathway of radical oxygen species? AB - OBJECTIVES: Cardiovascular, respiratory, and vascular dysfunction can follow trauma-induced no-flow-reflow states: hemorrhage, blunt trauma, or neurogenic shock. Liver ischemia-reperfusion (IR) induces remote lung damage by means of xanthine oxidase (XO) pro-oxidant activity. This damage was not proven in the heart, neither was the independent role of radical oxygen species (ROS) established in such cases. We investigated whether multiple organ dysfunction after a trauma-like IR is XO and ROS related and whether clinically used ROS scavengers could be beneficial. METHODS: A controlled, randomized trial in which isolated rat livers, hearts, lungs, and aortic rings were perfused with Krebs Henseleit solutions. After stabilization, livers were either perfused or made ischemic (2 hours). Then, pairs of liver plus heart, lung, or ring were reperfused in series (15 minutes), and then the second organ circulated alone for 45 minutes. Remote organ protection against the pro-oxidant hepatic-induced toxicity was evaluated by using allopurinol (1 mmol/L, heart), mannitol (0.25 g/kg, lung), or methylene blue (40 mg/kg, ring). RESULTS: IR liver effluents typically contained high lactate dehydrogenase, XO, and uric acid concentrations compared with control organs. IR was associated with doubled lung peak inspiratory pressure and reduced static compliance. Myocardial velocity of contraction and relaxation decreased by one third of baseline, and rings contracted abnormally and responded inadequately to phenylephrine. Wet-weight to dry-weight ratios in the remote organs increased as well. Most remote reperfusion injuries were attenuated by the drugs. CONCLUSION: Liver no-flow-reflow directly induces myocardial, pulmonary, and vascular dysfunction. These are likely mediated by XO and ROS. The tested drugs protected against these pro-oxidants, even in the presence of circulating XO. PMID- 10528604 TI - Low-dose vasopressin in the treatment of vasodilatory septic shock. AB - BACKGROUND: Despite appropriate therapy, refractory hypotension often occurs in septic shock. A double-blinded placebo controlled clinical trial was performed to assess the role of low-dose vasopressin (VP) as a pressor agent in septic shock. METHODS: Patients admitted to a trauma intensive care unit with vasodilatory septic shock were randomized to receive either VP at 0.04 U/min (n = 5) or placebo (n = 5). Vasodilatory septic shock was defined as a need for catecholamine agents to maintain a mean arterial pressure more than or equal to 70 mm Hg, despite a cardiac index more than 2.5 L/min and a minimal pulmonary artery wedge pressure more than 12 mm Hg. After 1 hour of initiation of the study drug, attempts to discontinue norepinephrine, phenylephrine, and/or dopamine, in respective order, were undertaken provided that the mean arterial pressure remained more than or equal to 70 mm Hg. RESULTS: A vasopressin infusion increased systolic arterial pressure (98 +/- 5 to 125 +/- 8 mm Hg, p < 0.008) because of peripheral vasoconstriction (systemic vascular resistance increased from 878 +/- 218 to 1,190 +/- 213 dynes/s per cm(-5) p < 0.05). Arterial pressure and systemic vascular resistance were statistically unaffected in the placebo group. Before study termination, measured at 24 hours after drug initiation, two patients in the placebo group died of refractory hypotension. However, all patients receiving VP survived the 24-hour study period and had all other catecholamine pressors withdrawn and blood pressure maintained solely with a low dose VP infusion. CONCLUSION: A VP infusion improved arterial pressure and permitted the withdrawal of catecholamine vasopressors. VP is a useful agent in the treatment of refractory septic shock. PMID- 10528605 TI - Differential alterations in systemic and regional oxygen delivery and consumption during the early and late stages of sepsis. AB - BACKGROUND: Studies have indicated that regional changes in oxygen utilization during sepsis cannot be predicted from the changes in the whole body oxygen delivery (DO2) and consumption (VO2). The aim of this study, therefore, was to determine whether differential alterations in systemic and regional DO2 and VO2 occur during the early and late stages of sepsis. METHODS: Adult male Sprague Dawley rats were subjected to sepsis by cecal ligation and puncture (CLP). At 5 hours (i.e., the early, hyperdynamic phase of sepsis) or 20 hours (i.e., the late, hypodynamic phase) after CLP, cardiac output, and organ blood flow were measured by radioactive microspheres. Systemic and regional DO2 and VO2 were determined and plasma levels of lactate were measured. RESULTS: Cardiac output and blood flow to the liver, small intestine, and kidneys increased at 5 hours and decreased at 20 hours after CLP. Although both systemic DO2 and VO2 increased at 5 hours after CLP, systemic DO2 but not VO2 decreased at 20 hours. At 5 hours after CLP, intestinal and renal DO2 increased. However, DO2 in all the tested organs decreased at 20 hours after CLP. VO2 increased in the liver, small intestine, and kidneys at 5 hours after CLP but decreased only in the liver and small intestine at 20 hours after the onset of sepsis. Moreover, plasma lactate levels increased at the late stage of sepsis. CONCLUSION: Because hepatic and intestinal VO2 but not systemic and renal VO2 decreased at 20 hours after CLP, the liver and small intestine seem to be more vulnerable to the hypoxic insult during the hypodynamic stage of polymicrobial sepsis. PMID- 10528606 TI - Direct actions of endothelin-1 on single vessel hydraulic permeability. AB - BACKGROUND: There is evidence that endothelin-1 (ET-1) increases extravasation of fluid and protein into vascular beds. The present study was designed to determine the direct effects of ET-1 on hydraulic permeability (Lp) when microvascular hydraulic and oncotic pressures are controlled. METHODS: Postcapillary venules in the rat mesentery were perfused in situ and paired measurements of Lp obtained by using the modified Landis micro-occlusion method. Lp measured after a 15-minute perfusion with Ringer's albumin solution (control) was compared with Lp after a subsequent 15-minute perfusion with one of three treatments: control (n = 4), 8 pM ET-1 (n = 6), or 80 pM ET-1 (n = 6). RESULTS: Baseline L for all vessels averaged (+/- SE) 8.1 +/-0.8 x 10(-8) cm x sec(-10 x cm H2O(-1) and was not significantly different between groups. Perfusion with either control or 8 pM ET 1 did not significantly change the Lp of any of the vessels. Significant decreases in Lp of 40 to 60% were observed in venules perfused with 80 pM ET-1. The average Lp in this group was 9.9 +/- 1.4 during baseline and decreased to 5.0 +/- 0.7 during ET-1 perfusion (p = 0.003). Washout of 80 pM ET-1 for periods of up to 15 minutes did not return Lp to baseline values. CONCLUSION: Low-dose ET-1 did not directly increase Lp in postcapillary venules. ET-1 at 80 pM, however, significantly decreased Lp. These data implicate factors other than a direct permeability-increasing effect in ET-1. At higher concentrations, ET-1 may have a protective effect on endothelial barrier function. PMID- 10528607 TI - Systemic activation of tissue-factor dependent coagulation pathway in evolving acute respiratory distress syndrome in patients with trauma and sepsis. AB - BACKGROUND: Extravascular coagulation and fibrin deposition coupled with perturbations of intravascular coagulation occurs in association with acute respiratory distress syndrome (ARDS). To evaluate the pathogenetic role of an extrinsic coagulation pathway in the intravascular coagulation of ARDS patients and to explore the time course of the changes of tissue factor levels, platelet counts, and disseminated intravascular coagulation (DIC), we performed a prospective cohort study. METHODS: The study subjects consisted of 113 patients: 27 patients with ARDS, 31 patients at risk for but not developing the syndrome, and 55 patients without ARDS. According to the underlying disease, the patients were further subdivided into two groups: patients with trauma (n = 76) and patients with sepsis (n = 37). Ten normal healthy volunteers served as control subjects. Plasma tissue factor antigen (tissue factor) levels and platelet counts were measured on the day of admission and on days 1 through 4 after admission. Simultaneously, the DIC scores were determined. RESULTS: The values of tissue factor in the patients with ARDS were significantly more elevated than those measured in the other two groups (p < 0.001) and control subjects (p < 0.001) on the day of admission. The values continued to be markedly high up to day 4 of admission. On the day of admission, the platelet counts in the ARDS patients showed significantly lower values (p < 0.05) than those in the other two groups. The incidence of DIC and the DIC scores in ARDS patients were significantly higher than those in the other two groups. The tissue factor levels (r(s) = 0.428, p < 0.0001) and DIC scores (r(s) = 0.357, p < 0.0002) correlated significantly with Lung Injury Score. When the patients were subdivided into two subgroups, i.e., trauma and sepsis, some differences of the tissue factor levels were noted between the two groups. CONCLUSION: We demonstrated that tissue-factor dependent coagulation pathway of plasma is extensively activated in patients with ARDS, followed by intravascular coagulation and platelet consumption. We further provide precise information on the time course of tissue factor levels and DIC in patients with ARDS and those at risk for developing this syndrome. PMID- 10528608 TI - Below-knee amputations as a result of land-mine injuries: comparison of primary closure versus delayed primary closure. AB - BACKGROUND: Antipersonnel land mines are designed to maim by mutilating the lower extremities, and these injuries are at higher risk for infection than injuries from other weapon systems. METHODS: The results of 474 unilateral traumatic below knee amputations as a result of land-mine injuries were reviewed. If the delay in evacuation between the injury and arrival to the battle field hospital was less than 6 hours, 392 amputation stumps (group I) were closed primarily after meticulous debridement. Open amputation was performed after debridement in the remaining 82 amputation stumps (group II), because there was a suspicion of ineffective debridement, although they were evacuated in less than 6 hours or delay was more than 6 hours. RESULTS: Eleven patients in group I (2.8%) were reoperated because of wound sepsis of the stump. Wound sepsis was not encountered in group II. A total of 87.4% of stumps in group I and 81.2% of stumps in group II had healed without a problem. No gas gangrene or tetanus was encountered in any cases. CONCLUSION: Our results reveal that primary closure may be done in traumatic below-knee amputations caused by land-mine injuries with an acceptable infection rate, if the evacuation time is less than 6 hours, and if there is meticulous debridement. PMID- 10528609 TI - Argentine Jewish community institution bomb explosion. AB - BACKGROUND: Descriptive study of physical injuries and implemented organization from a nearby, unwarned university hospital after the July 18, 1994, bombing of the seven-story Argentine Israeli Mutual Association (AMIA) building in Buenos Aires. Data were obtained from hospital medical records. RESULTS: A total of 86 victims arrived at the emergency department, 2 victims were dead on arrival, 41 victims were admitted, and 43 victims with minor injuries were assisted and allowed to go home. The explosion caused a total of 86 deaths and left more than 200 people injured. Mortality rate among hospitalized survivors was 8.3% and among critically injured victims was 28.6%. CONCLUSION: The total collapse of a multiple-story building immediately kills most of its occupants. In the present study, the few surviving victims were located at the lower floors. The majority of hospitalized victims were outside the building at the moment of the blast. Rapid overcrowding of the emergency department with minor and moderate injuries that do not require hospitalization should be anticipated by disaster management plans. Centralization of severely injured patients in critical areas seems appropriate, because this method keeps major cases from spreading through different wards. PMID- 10528610 TI - Unarmed violence-related injuries requiring hospitalization in Sweden from 1987 to 1994. AB - BACKGROUND: Physical abuse and assault are common problems in the Western hemisphere. The aims of this study were to investigate the injury incidence, distribution of injuries, the age and sex distribution, and the geographical differences in all patients admitted to Swedish hospitals between 1987 and 1994 because of injuries related to unarmed assault. METHODS: Patients admitted to hospitals in Sweden between 1987 and 1994 after physical abuse were included in the Swedish Hospital Discharge Register. A description of the types of injuries, surgical procedures, and lengths of hospital stay are presented. The change in the incidence of hospital admissions for unarmed violence-related injuries was evaluated. Linear regression analysis was used to correlate population density with incidence of hospital admission and to evaluate the change in age standardized incidence of hospital admissions over time. RESULTS: Information was available on 17,453 persons, of whom 79% were males. The mean age was 30 years. Craniocerebral injury was the most common (72%) followed by injuries to the extremities (10%), thorax (5%), and abdomen (3%). The mean in-hospital stay was 3 days. Thirty-eight people (0.2%) died of their injuries. The age-standardized incidence of hospital admissions increased significantly over the years in males, but not in females. No correlation was detected between population density and incidence of injury. CONCLUSION: Young males are at the greatest risk of incurring physical injuries from assaults that warrant hospital admission, and the incidence in this group has increased significantly. Injuries to the head are the most common. Fatal injuries are rare. The in-hospital stay is usually brief. The frequencies of assaults are similar in urban and rural areas. PMID- 10528611 TI - Free medialis pedis flap as a coverage and flow-through flap in hand and digit reconstruction. AB - BACKGROUND: Skin and soft-tissue defects of the hand and digit present a challenge for the hand surgeon especially in meeting the reconstructive needs of thickness, texture, color matching, and sensation. It becomes an even bigger challenge to reconstruct the defect in a devascularized finger with segmental loss of the neurovascular bundle. We use the relatively new flap, the medialis pedis flap, to solve the above conditions and compare it with traditional flaps. METHODS: From May of 1994 to March of 1997, the free medialis pedis flap was used to reconstruct 19 digit and hand defects; the flap sizes ranging from 1.5 x 3.0 cm to 3.0 x 9.0 cm. Sixteen flaps were used for simple coverage of digit defects, including 12 for single-digit and 4 for multiple-digit reconstruction. The remaining three flaps were used as coverage and a flow-through flap for devascularized fingers. RESULTS: All 19 flaps survived and achieved a good protective sensation. The appearance was very satisfactory, and the donor-site scars were without sensory problems. CONCLUSION: Compared with traditional flaps, the free medialis pedis has the following advantages: it provides good thickness, texture, and color matching for hand and digit resurfacing; it can be used as a flow-through flap and as coverage for a devascularized finger in a one-stage procedure; the size of the feeding vessels of the flap matches those of the digital vessels well; and it consists of glabrous skin rich in nerve endings, so it has good potential for sensory recovery. Because of all of these characteristics, the free medialis flap may become a better consideration for hand surgeons. PMID- 10528612 TI - Cortical revascularization after reamed and unreamed intramedullary nailing in the rabbit femur: a microangiographic histometric analysis. AB - BACKGROUND: Intramedullary nailing leads to a reduction in cortical bone blood flow. The relative effect of reamed versus unreamed nailing on the degree of avascularity and on revascularization of the cortex remains controversial. We compared the effects of reamed versus unreamed intramedullary nailing on cortical revascularization and the time course for its recovery in the unfractured rabbit femur. METHODS: A total of 56 New Zealand white rabbits were used as experimental animals. Reamed nailings with Kirschner wire (diameter, 3 mm) were performed in the right femora (group R, n = 49), and unreamed nailings with Kirschner wire (diameter, 2 mm) were performed in the left femora (group UR, n = 49) of the same animals after a standard surgical procedure. Microangiography that used Indian ink was performed for each killed animal at one of seven postoperative time periods: 2 hours, 3 days, and 1, 2, 3, 4, and 5 weeks after surgery. The right femora and the left femora of seven rabbits were used as the postoperative day 0 controls of group R and those of group UR, respectively. We evaluated the vascularization ratio (VR) in cross-sections according to the following formula: VR = the number of enhanced vessels with Indian ink/total cortical vessels. Each cross-section was divided into an inner and outer half, which were in turn divided into quarters, so that the entire cross-sectional are is represented by eight regions. The VR was calculated for each regions and then comparison was made between group R and group UR at various time periods and areas. RESULTS: No statistically significant differences were observed in VR at any time period between group R and group UR in total cortical area and total periosteal side. However, on the total medullary side, the VR of group UR was significantly higher than that of group R at postoperative day 3 (p = 0.04). Statistically significant differences were observed between the VR on the periosteal side and that on the medullary side in both groups at all time periods aside from the 5-week period (p < 0.05). Revascularization of the cortex occurred 4 weeks after intramedullary nailing in both group R and group UR. CONCLUSION: The periosteal circulation was maintained better than the medullary circulation, irrespective of whether the canal was reamed or not. The present study failed to detect any statistically significant differences in cortical revascularization between reamed nailing and unreamed nailing. Thus, we concluded that reamed and unreamed nailing are not differentially advantageous in the unfractured rabbit femur, in terms of impairment of cortical blood supply. PMID- 10528613 TI - Hyperbaric oxygen therapy mitigates the adverse effect of cigarette smoking on the bone healing of tibial lengthening: an experimental study on rabbits. AB - OBJECT: We investigated whether -intermittent hyperbaric oxygen (HBO) therapy can mitigate the adverse effects of cigarette smoking on the bone healing of tibial lengthening by using a previously validated rabbit model. METHODS: Eighteen male rabbits were randomly divided into three groups of six animals each. Group 1 (smoking plus HBO) went through intermittent cigarette smoke inhalation and hyperbaric oxygen therapy, group 2 (control) did not go through intermittent cigarette smoke inhalation or hyperbaric oxygen therapy and group 3 (smoking) went through intermittent cigarette smoke inhalation. Each animal's right tibia was lengthened 5 mm by using an uniplanar lengthening device. Bone mineral density (BMD) study was performed for all the animals at 1 day before operation and 3, 4, 5, and 6 weeks after operation. All of the animals were killed at 6 weeks postoperatively for biomechanical testing. RESULTS: By using the preoperative BMD as an internal control, we found that the BMD of group 1 (smoking plus HBO)and group 2 (control) was superior to that of group 3 (smoking). The mean %BMD at 3, 4, 5, and 6 weeks were 58.6%, 66.6%, 73.7%, and 83.8%, respectively, in group 1, whereas the mean %BMD were 52.0%, 64.3%, 70.1%, and 76.2%, respectively, in group 2, and the mean %BMD were 46.2%, 54.0%, 64.9%, and 69.4%, respectively, in group 3 (two-tailed t test, p > 0.05, p > 0.05, p > 0.05, and p < 0.05 at 3, 4, 5, and 6 week respectively between group 1 and group 2, p < 0.01,p < 0.01,p < 0.01, and p < 0.01 at 3, 4, 5, and 6 week, respectively, between group 1 and group 3 and p < 0.05, p < 0.05, p < 0.05, and p < 0.05 at 3, 4, 5, and 6 week respectively between group 2 and group 3). By using the contralateral nonoperated tibia as an internal control, we found that the torsional strength of group 1 (smoking plus HBO) and group 2 (control) was superior to that of group 3 (smoking). The mean percentage of maximum torque was 80.9% in group 1 (smoking plus HBO) and was 78.0% in group 2 (control), whereas the mean percentage of maximum torque was 59.6 % in group 3 (smoking) (two-tailed t test, p < 0.05 between groups land 3 and between groups 2 and 3, whereas p > 0.05 between groups 1 and 2). CONCLUSION: This study suggests that smoke inhalation delays the bone healing in tibial lengthening; however, HBO mitigates the delayed healing effect of smoke inhalation and, thus, helps the smoking animal in achieving an expeditious bone healing in tibial lengthening. PMID- 10528614 TI - Internal fixation of distal radius fractures with dorsal dislocation: pi-plate or two 1/4 tube plates? A prospective randomized study. AB - BACKGROUND: Severely comminuted distal radius fractures can be treated by different methods. Our routine procedure in dorsal dislocated fractures is the dorsal stabilization with two 1/4 tube plates. The new pi-plate is an other device that matches optimally the anatomy of the distal radius and allows a near half-circumferential dorsal buttress of comminuted intraarticular and extra articular radial fractures. METHODS: In a prospective randomized study, comminuted distal radius fractures with dorsal displacement were stabilized either with two 1/4 tube plates or with the pi-plate. All patients were reviewed at 1, 3, and 6 months after surgery by thorough clinical examination and standard radiographs of both wrists. Results were analyzed and compared in both groups. RESULTS: Subjective and objective results in the pi-plate group are disappointing. Although optimal anatomic results were achieved, the complication rate was high (14.3%) and the range of motion was limited. At final review, extension and flexion of the injured wrist had recovered to an average of 67% of the normal, contralateral side. Radial and ulnar deviation were limited to 64%, whereas pronation and supination reached 89% and 87%, respectively. Overall, results were good to excellent only in 56%. In a comparable group of patients with similar fractures and stabilization with two 1/4 tube plates, 82% of patients achieved excellent to good results, wrist motion was significantly better (p < 0.05), and no complications occurred. CONCLUSION: With open reduction, cancellous bone grafting, and internal plate fixation in comminuted distal radial fractures, excellent results can be achieved. In our experience, we cannot recommend the 7pi-plate in its current shape and prefer to stabilize distal radius fractures and dorsal fragment dislocations with two 1/4 tube plates. PMID- 10528615 TI - Toxic lead levels treated with 2,3-dimercaptosuccinic acid and surgery. PMID- 10528616 TI - Late complications of arrow and spear wounds to the head and neck. PMID- 10528617 TI - Arteriovenous fistula presenting as airway hemorrhage after percutaneous tracheostomy. PMID- 10528618 TI - Fate of the vascularized fibular graft: report of two cases with 15-year follow up. PMID- 10528619 TI - Intra-articular dislocation of the patella: two cases and literature review. AB - Intra-articular dislocation of the patella remains uncommon and is generally thought to be a problem of young adolescent males. This report reminds emergency physicians that it can occur in the arthritic knee, and it should be considered in the differential diagnosis of locked knee in the elderly. Closed reduction should be attempted in these cases, because the dislocation is liable to be held in place by osteophytes rather than impaction of the patella deep in the intercondylar notch and a good functional outcome can be expected. PMID- 10528620 TI - Bilateral external iliac artery dissections after pelvic fracture: case report. PMID- 10528621 TI - Descending aortic cannulation during emergent thoracotomy for blunt traumatic cardiac arrest. PMID- 10528623 TI - Pericardioscopy in the management of suspected hemopericardium. PMID- 10528622 TI - Repair of combined traumatic rupture of the brachiocephalic trunk and left common carotid artery by using hypothermic circulatory arrest. PMID- 10528624 TI - Compartment syndrome of the liver. PMID- 10528625 TI - Minimally invasive treatment of distal femur fractures: report of a technique. PMID- 10528626 TI - Rural trauma: the challenge for the next decade. AB - Improving the care of trauma patients in a rural environment requires that several important issues be addressed. First, a universal definition of what constitutes "rural" must be established. We propose that a combined effort of the Federal Government and the Committee on Trauma of the American College of Surgeons develop this definition. Second, data on rural trauma demographics and outcome must be collected in a national database. We propose that this database be incorporated in the "TRACS" database of the Committee on Trauma of the American College of Surgeons. Such a database will allow a "needs assessment analysis of existing care in rural environments and facilitate planning and implementation of efficient systems of care. Funding for the rural database should come from the federal government. Finally, increased public awareness of problems unique to rural trauma care is necessary. The rural trauma subcommittee of the ACSCOT should go from an ad hoc committee to a standing committee with the American College of Surgeons Committee on Trauma. We propose a national conference on rural trauma care hosted by the federal government for the purpose of addressing these issues and simultaneously increasing public awareness. PMID- 10528627 TI - Complete division of the ulnar nerve at the level of the medial epicondyle in the cubital tunnel. PMID- 10528628 TI - Oxymetholone: I. Evaluation in a comprehensive battery of genetic toxicology and in vitro transformation assays. AB - Oxymetholone is generally assumed to be a nongenotoxic carcinogen. This assumption is based primarily on the results of an Ames test, existing data in repeat-dose toxicology studies, and the predicted results of a 2-yr National Toxicology Program (NTP) rat carcinogenicity bioassay. To provide a comprehensive assessment of its genotoxicity in a standard battery of mutagenicity assays, oxymetholone was tested in microbial and mammalian cell gene mutation assays, in an in vitro cytogenetics assay (human lymphocytes), and in an in vivo micronucleus assay. Oxymetholone was also tested in an in vitro morphologic transformation model using Syrian hamster embryo (SHE) cells. These studies were initiated and completed prior to the disclosure of the results of the NTP bioassay. Oxymetholone was tested at doses up to 5,000 microg/plate in the bacterial plate incorporation assay using 4 Salmonella strains and the WP2 uvrA (pKM101) strain of Escherichia coil. There was no induction of revertants up to the highest dose levels, which were insoluble as well as toxic. In the L5178Y tk+/- mouse lymphoma assay, doses up to 30 microg/ml reduced relative survival to approximately 30% with no increase in mutants. Male or female human lymphocytes were exposed in vitro to oxymetholone for 24 hr without S9 or 3 hr with S9 and evaluated for the induction of chromosomal aberrations. There was no increase in aberration frequency over control levels and no difference between male and female cells. Peripheral blood from Tg.AC transgenic mice treated dermally for 20 wk with 0, 1.2, 6.0, or 12.0 mg/day of oxymetholone and from p53 transgenic mice treated orally by gavage for 26 wk with 125, 625, or 1,250 mg/kg/day of oxymetholone was evaluated for micronuclei in polychromatic and normochromatic erythrocytes. There was no difference in micronuclei frequency between control and treated animals. These results confirm that oxymetholone is not genotoxic in a comprehensive battery of mutagenicity assays. In the SHE assay, oxymetholone produced a significant increase in morphologically transformed colonies at dose levels of 13-18 microg/ml. The lack of genotoxicity of oxymetholone, the positive response in the in vitro transformation assay, and the results of transgenic mouse carcinogenicity assays will provide an interesting perspective on the results of an on-going NTP rat carcinogenicity bioassay. PMID- 10528629 TI - Oxymetholone: II. Evaluation in the Tg-AC transgenic mouse model for detection of carcinogens. AB - Several rodent models are under examination as possible alternatives to the classical 2-yr carcinogenicity bioassay. The Tg.AC transgenic mouse has been proposed as a shorter term model offering the possibility of detecting nongenotoxic and genotoxic carcinogenic agents. Retrospective studies of chemicals with established carcinogenic potential have revealed a close correlation between classical bioassay results and the production of skin tumors in the Tg.AC mouse model. Oxymetholone is a synthetic testosterone derivative that is a suspected carcinogen but has shown no evidence of genotoxic activity in a comprehensive battery of genetic toxicity assays. It currently is being tested by the National Toxicology Program (NTP) in a 2-yr rat carcinogenicity bioassay. Because of its nongenotoxicity and the ongoing chronic bioassay, oxymetholone was considered an ideal candidate for a prospective evaluation of the predictive validity of the Tg.AC dermal carcinogenicity model. Consequently, a 6-mo dermal study with oxymetholone in the Tg.AC mouse model was initiated and completed prior to disclosure of the NTP rat bioassay results. In this study, male and female hemizygous Tg.AC mice, 7-8 wk old, were housed individually in suspended plastic cages. An area of dorsal skin was shaved to accommodate dermal applications of 200-microl doses of vehicle control (acetone), drug (1.2, 6.0, or 12 mg oxymetholone in dimethylsulfoxide:acetone, 20:80), or positive control (1.25 microg 12-o-tetradecanoyl-phorbol-13-acetate [TPA]) solutions. Mice received oxymetholone or acetone daily or TPA twice weekly for 20 wk followed by a 6-wk recovery period. The acetone control groups exhibited low spontaneous incidences of papillomas, whereas dermal application of oxymetholone produced dose-related increases in the numbers of papilloma-bearing mice and the numbers of papillomas per animal. Females showed a somewhat greater response to the androgen than did the males. TPA caused an unequivocal increase in papillomas, with males exhibiting a greater response than females. The results of this study indicate that this nongerotoxic androgenic compound possesses proliferative properties. The results predict that chronic systemic administration of oxymetholone will most likely be associated with increased incidences of neoplasms. PMID- 10528630 TI - Oxymetholone: III. Evaluation in the p53+/- transgenic mouse model. AB - Oxymetholone has been identified as a suspected nongenotoxic carcinogen and has recently completed testing in a conventional National Toxicology Program (NTP) 2 yr rodent bioassay program. As a synthetic androgen with a limited historical database in toxicology, oxymetholone is an ideal candidate for prospective examination of the performance of short-term transgenic mouse models in the detection of carcinogenic activity. In the present series of 3 articles, studies are described where oxymetholone was evaluated prior to disclosure of the results of the NTP 2-yr bioassay. The accompanying articles provide evidence showing that oxymetholone is devoid of mutagenic activity yet elicits a positive carcinogenic response in the Tg.AC transgenic mouse model. In the present study, oxymetholone was administered by oral gavage to p53 heterozygous male and female mice for 26 wk at doses of 125, 625, and 1,250 mg/kg/day. The vehicle was 0.5% aqueous methylcellulose. Positive controls consisted of mice treated daily by oral gavage with 200 or 400 mg/kg/day of p-cresidine in corn oil. The oxymetholone-treated females showed significantly increased body weight gain and clitoral enlargement attributable to drug treatment. In addition, significant alterations in kidney, liver, and testis weights were attributable to oxymetholone. However, there were no neoplastic lesions that were attributable to oxymetholone in either sex. p Cresidine produced unequivocal bladder neoplasms in both sexes at the high dose and in males at the lower dose. The absence of a neoplastic response with oxymetholone is consistent with the selectivity of the p53-/- mouse model for detecting carcinogens that act by genotoxic mechanisms. PMID- 10528631 TI - Transponder-induced sarcoma in the heterozygous p53+/- mouse. AB - Heterozygous p53+/- transgenic mice are being studied for utility as a short-term alternative model to the 2-yr rodent carcinogenicity bioassay. During a 26-wk study to assess the potential carcinogenicity of oxymetholone using p-cresidine as a positive control, glass/polypropylene microchips (radio transponder identification devices) were subcutaneously implanted into male and female p53+/- mice. During week 15, the first palpable mass was clinically observed at an implant site. This rapidly growing mass virtually quadrupled in size by week 25. Microscopic examination of all implant sites revealed that 18 of 177 animals had a subcutaneous histologically malignant sarcoma. The neoplasms were characterized as undifferentiated sarcomas unrelated to drug treatment, as indicated by the relatively even distribution among dose groups, including controls. An unusual preneoplastic mesenchymal change characterized by the term "mesenchymal dysplasia" was present in most groups and was considered to be a prodromal change to sarcoma development. The tumors were observed to arise from dysplastic mesenchymal tissue that developed within the tissue capsule surrounding the transponder. The preneoplastic changes, including mesenchymal dysplasia, appeared to arise at the transponder's plastic anchoring barb and then progressed as a neoplasm to eventually surround the entire microchip. Capsule membrane endothelialization, inflammation, mesenchymal basophilia and dysplasia, and sarcoma were considered unequivocal preneoplastic/neoplastic responses to the transponder and were not related to treatment with either oxymetholone or p cresidine. PMID- 10528632 TI - Dorsal skin reactions of hairless dogs to topical treatment with corticosteroids. AB - Dorsal skin reactions to continuous topical treatment with different types of corticosteroids were histologically investigated in hairless descendants of Mexican hairless dogs. The preparations tested were prednisolone (ST-1; weak), fluocinolone acetonide (ST-2; moderate), diflucortolone valrerate (ST-3; strong), and mometasone furoate (ST-4; very strong). Grossly, the sites treated with ST-3 and ST-4 showed moderate inflammatory reactions. After completion of the corticosteroid treatment, both sites were less pigmented and had a thin texture. The severity of histologic changes in the skin was dependent on the efficacy of the corticosteroids. The epidermis was prominently thinned from 1 wk after treatment with the corticosteroids, resulting in a flat dermis-epidermis junction. By the end of the corticosteroid treatment, these lesions became progressively more severe. At 2 wk after completion of topical treatment, the epidermal thickness in the sites treated with ST-1 and ST-2 began to return to normal values, whereas the epidermis of the skin treated with ST-3 and ST-4 became thinner. At 3-4 wk after topical treatment with ST-3 and ST-4, the dermis showed hyalinization of collagen bundles. These dermatologic findings in hairless dogs are in accordance with steroid-induced skin atrophy of human beings. These results suggest that the skin of hairless dogs responds sensitively to topical corticosteroids and that these animals are a useful model for investigating the efficacy and adverse effects of cutaneous topical corticosteroids. PMID- 10528633 TI - Comparisons of the intraocular tissue distribution, pharmacokinetics, and safety of 125I-labeled full-length and Fab antibodies in rhesus monkeys following intravitreal administration. AB - Access of recombinant proteins to the retina following intravitreal administration is poorly understood. A study was conducted in male Rhesus monkeys (15 to 28 mo of age; 2.8-3.3 kg) in order to compare the intraocular tissue distribution, pharmacokinetics, and safety of 125Iodine (I)-labeled full-length humanized rhuMAb HER2 antibody (148 kD) and of 125I-labeled humanized rhuMAb vascular endothelial growth factor Fab antibody (48.3 kD) following bilateral bolus intravitreal injection on day 0 (5 animals/group). The dose administered to each eye was 25 microg (9-10 microCi) in 50 microl. Animals were euthanatized on day 0 (1 hr postdose) and on days 1, 4, 7, and 14. Safety assessment included direct ophthalmoscopy, intraocular pressure measurements, clinical observations, body weight, and hematology and clinical chemistry panels. Blood and vitreous samples were collected daily (blood only) and at necropsy for pharmacokinetics and analysis for antibodies to the test materials; the ocular tissue distribution of the test material was evaluated by microautoradiography. All animals completed the study. Microautoradiography demonstrated that the full-length antibody did not penetrate the inner limiting membrane of the retina at any of the time points examined. In contrast, the Fab antibody fragment diffused through the neural retina to the retinal pigment epithelial layer at the 1-hr time point and persisted in this location for up to 7 days. Systemic exposure to test material was low but variable: the highest plasma concentration of the full-length antibody was 20.3 ng/ml, whereas plasma concentrations for the Fab antibody remained below the limit of quantitation (i.e., <7.8 ng/ml). An immune response to the test material was not evident in either treatment group. The half-life in vitreous was 5.6 days for the full-length antibody and 3.2 days for the Fab antibody. The shorter intravitreal half-life of the Fab antibody is related to its smaller size and its significant diffusion through the retinal layers. The differences in pharmacokinetics and tissue distribution that are noted between the full-length and Fab antibodies in this study identify potential therapeutic approaches that may be exploited in specific disease conditions. PMID- 10528634 TI - Troglitazone-induced heart and adipose tissue cell proliferation in mice. AB - Troglitazone, a thiazolidinedione, is a novel agent for the oral treatment of non insulin-dependent (Type II) diabetes mellitus; it works by increasing cell sensitivity to available insulin. Previous studies have shown that rodents treated with high doses of troglitazone develop increased heart weight and increased interscapular brown fat. This study investigated cellular proliferation in heart and brown fat of troglitazone-treated mice as well as possible interactions with an angiotensin-converting enzyme inhibitor (quinipril). B6C3F1 female mice were treated daily with either vehicle control, 125 mg/kg quinipril, 1,200 mg/kg troglitazone, or troglitazone/quinipril combination per os for up to 14 days. Four days before necropsy, mice were dosed with bromodeoxyuridine (BrdU) using osmotic pumps. Cell proliferation in heart, brown fat, and retroperitoneal white fat was investigated by means of light microscopic anti-BrdU immunolabeling techniques. Immunoelectron microscopy was used to determine the cell phenotypes and cellular distribution of BrdU label in heart and brown fat. Treatment with troglitazone for 2 wk resulted in increased heart and brown fat weights but in decreased white fat weight. Combination treatment with troglitazone and quinipril also resulted in decreased white fat weight compared with controls. Histologically, brown fat adipocytes in troglitazone- and troglitazone/quinipril treated mice had coalescent lipid vacuoles and increased eosinophilia of the cytoplasm. White fat adipocytes in troglitazone- and troglitazone/quinipril treated mice had decreased cell size and increased cytoplasmic eosinophilia. BrdU labeling revealed increased cell proliferation in troglitazone-treated hearts after 1 wk but did not reveal increased cell proliferation in quinipril- or troglitazone/quinipril-treated animals. Brown fat BrdU labeling after 1 wk was increased in troglitazone- and troglitazone/quinipril-treated mice. Ultrastructural anti-BrdU immunogold labeling demonstrated that troglitazone treated heart and brown fat had greater populations of BrdU-labeled cells that were identified as endothelial cells. These results demonstrated that troglitazone-induced increased cardiac weight in mice can be prevented by quinipril and that increased cardiac weight coincides with early increased endothelial cell proliferation. PMID- 10528635 TI - Liver tumor promoting effects of fenbendazole in rats. AB - In order to examine whether fenbendazole has tumor-promoting activity, a total of 70 male Fischer 344 rats were initiated with a single intraperitoneal injection of 100 mg/kg of diethylnitrosamine (DEN) or were given the saline vehicle alone; beginning 1 wk later, rats were given a diet containing 3,600; 1,800; 600; 200; 70; or 0 ppm of fenbendazole for 8 wk. Subgroups of 5 rats each from the DEN+ 1,800; DEN+0; 1,800; and 0 ppm groups were euthanatized after 1 wk of fenbendazole treatment, and the remaining animals were euthanatized at 8 wk. After 1 wk, relative liver weights (ratios to body weights) were significantly increased in the DEN+ 1,800 and 1,800 ppm groups, and based on light microscopy, periportal hepatocellular hypertrophy was evident in these groups. After 8 wk, relative liver weights were significantly increased in the groups given > or =600 ppm with or without DEN initiation. Periportal hepatocellular hypertrophy, characterized by a marked increase in smooth endoplasmic reticulum, was observed in the groups given > or =600 ppm with or without DEN initiation. Induction of cytochrome P-450 (CYP) 1A2, 2B1, or 4A1 was noted in the fenbendazole-treated groups with or without DEN initiation; that associated with CYP 1A2 was most marked. Positive immunostaining for anti-CYP 1A1/2 or CYP 2B1/2 was observed diffusely in the livers of animals in the DEN+1,800 and DEN+3,600 ppm groups. The numbers and areas of connexin 32 (Cx32)-positive spots per square centimeter in centrilobular hepatocytes were significantly decreased in an almost dose dependent manner with fenbendazole treatment after DEN initiation. In situ hybridization for Cx32 mRNA revealed a remarkable decrease in its expression in the centrilobular hepatocytes in the DEN+70 ppm group. The numbers of glutathione S-transferase placental-form positive single cells (plus mini foci) were significantly increased in the DEN+ 1,800 and DEN+3,600 ppm groups. Since those agents that induce CYP 2B1/2 isozymes and reduce Cx32 in centrilobular hepatocytes have been suggested to be liver tumor promoters, the present results indicate that fenbendazole may be a liver tumor promoter. PMID- 10528637 TI - Emerging harmful algal blooms and human health: Pfiesteria and related organisms. PMID- 10528636 TI - Prompt recovery of damaged adrenal medullae induced by salinomycin. AB - The morphologic changes in the adrenal medullae of rats treated with an ionophore antibiotic, salinomycin, are described. Male rats of approximately 7 wk of age were treated orally with a single dose of salinomycin at 80 mg/kg body weight. Following this treatment, the adrenal glands were examined, using immunohistochemistry, for neurofilament, laminin, fibronectin, and S-100 protein; the glands were also examined using transmission electron microscopy. One hour after the treatment, a karyopyknosis was observed in the clusters of affected chromaffin cells in which the neurofilament, laminin, and fibronectin were present. The lesions became progressively conspicuous between hours 5 and 10. Ultimately, the outcome was cell lysis. Five hours after salinomycin treatment, unaffected chromaffin cells strongly stained to tyrosine hydroxylase. At 10 hr, new chromaffin cells, which were irregular in shape with electron-dense cytoplasm (dark cell), that were strongly stained for tyrosine hydroxylase appeared at the basement membrane site of the necrotic clusters, and these cells contained very few immature catecholamine granules of less than 80 nm. At 17 hr, the catecholamine granules increased in number and size to about 200 nm. The newly formed chromaffin cells grew within the clusters to fill in the medulla by 24 hr, and cytoplasmic granules progressively increased in number and size. The interstitial tissue was seen to be edematous at 5 hr. New capillaries were found in the adrenal medullae of both control and salinomycin-treated rats. The protruding chromaffin cells (protruding cells), which we previously described in normal rats, were also observed in salinomycin-treated rats, which suggests that holocrine secretion is performed in the adrenal medullae. The results indicated that the rat adrenal medullae have the ability to make a rapid recovery after an insult by salinomycin. PMID- 10528638 TI - Hepatic oxidative stress following prolonged sublethal microcystin LR exposure. AB - Microcystin LR (MCLR) is a naturally occurring protein phosphatase inhibitor and potent hepatotoxin produced by strains of Microcystis aeruginosa. Although its acute toxicity has been well characterized, little is known about its chronic effects. In this study, we sought to acquire evidence that oxidative stress may play a role in the pathogenesis of prolonged sublethal MCLR toxicity. Twelve rats (3 per group) weighing on average 185 g were exposed to 1 of 3 different concentrations of MCLR (16, 32, and 48 microg/kg/day) or to saline via intraperitoneal osmotic pumps for 28 days. Histologic evidence of dose-dependent hepatic inflammation was seen, including infiltration of centrilobular regions by lymphocytes, macrophages, and neutrophils, centrilobular fibrosis, apoptosis, and steatosis. Analysis of lipid peroxidation products revealed a dose-dependent increase in malondialdehyde concentrations with an approximate 4-fold increase in the livers of the high-dose rats over those of the saline-treated controls. Livers from MCLR-exposed rats were more sensitive than those of controls to the cytotoxic effects of the organic oxidizing agent tert-butyl hydroperoxide, based on an MTT (3-[dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) viability assay. These histopathologic and biochemical findings indicate that oxidative stress may play a significant role in the pathogenesis of chronic MCLR toxicosis. PMID- 10528639 TI - Examination of low-incidence brain tumor responses in F344 rats following chemical exposures in National Toxicology Program carcinogenicity studies. AB - Neoplasms in the brain are uncommon in control Fischer 344 (F344) rats; they occur at a rate of less than 1% in 2-yr toxicity/carcinogenicity studies. Furthermore, only 10 of nearly 500 studies conducted by the National Toxicology Program (NTP) showed any evidence of chemically related neoplastic effects in the brain. Generally, the brain tumor responses were considered equivocal, because the characteristics of potential neurocarcinogenic agents (such as statistically significant increased incidences, decreased latency and/or survival, and demonstration of dose-response relationships) were not observed. A thorough examination, including comparisons with a well-established historical database, is often critical in evaluating rare brain tumors. Chemicals that gave equivocal evidence of brain tumor responses were generally associated with carcinogenicity at other sites, and many chemicals were mutagenic when incubated with metabolic activating enzymes. Other factors that were supportive of the theory that marginal increases in brain tumor incidence were related to chemical exposure were that (a) some of the tumors were malignant, (b) no brain neoplasms were observed in concurrent controls from some studies, and/or (c) brain tumors were also seen following exposure to structurally related chemicals. In 2-yr studies in F344 rats (studies conducted by the NTP), equivocal evidence of carcinogenicity was observed for the following 9 chemicals: isoprene, bromoethane, chloroethane, 3,3'-dimethylbenzidine dihydrochloride, 3,3' dimethoxybenzidine dihydrochloride, furosemide, C.I. direct blue 15, diphenhydramine hydrochloride, and 1-H-benzotriazole. Glycidol was the only chemical evaluated by the NTP with which there was clear evidence of brain tumor induction in F344 rats. Clarification of the potential neurocarcinogenic risks of chemicals that produce equivocal evidence of a brain tumor response in conventional 2-yr rodent studies may be aided by the use of transgenic mouse models that exhibit genetic alterations that reflect those present in human brain tumors as well as by the use of in utero exposures. PMID- 10528640 TI - A congenital ciliated epithelial cyst on the stomach of a B6C3F1 mouse. AB - Congenital anomalies of the alimentary tract are rare lesions in laboratory animals. We describe a congenital cyst attached to the greater curvature of the forestomach in a B6C3F1 mouse. The inner surface of the cyst was mostly covered with cuboidal or pseudostratified ciliated epithelium and was focally covered with the flat cuboidal epithelium. The base of the cyst appeared to be inserted between the layers of the outer longitudinal muscle layer of the forestomach, although no smooth muscle layer was evident in the free surface of the cyst wall. The cyst resembled duplication of the alimentary tract, as it was lined with ciliated epithelium and had developed at the greater curvature of the forestomach. Since the smooth muscle layer did not completely cover the whole wall and the cyst did not communicate with the gastric lumen, the cyst was not thought to be a standard duplication but rather a simple congenital cyst. PMID- 10528641 TI - Hypertensive nephrosclerosis: pathogenesis and prevalence. Essential hypertension is an important cause of end-stage renal disease. PMID- 10528642 TI - Glomerular IgA deposition in liver disease. PMID- 10528643 TI - Insulin-mediated sympathoexcitation in obesity and type 2 diabetes. PMID- 10528644 TI - Neuropathic pain in diabetic nephropathy--update on analgesic strategies. PMID- 10528645 TI - Haemodialysis catheter-related infection: time for action. PMID- 10528646 TI - Hypertension and the risk of intracerebral haemorrhage: special considerations in patients with renal disease. PMID- 10528647 TI - Hypercoagulable state and graft rejection--is there a link? PMID- 10528648 TI - Renal allograft thrombosis: can thrombophilia explain the inexplicable? AB - Renal allograft thrombosis remains a preventable cause of early allograft thrombosis. It should not be considered simply an unpredictable and poorly understood consequence of surgery. Extrapolated data from the general population and early data from renal patients supports the concept that the interplay of non inherited hypercoagulability of renal disease with inherited thrombophilia, and the altered environmental milieu of transplantation predisposes to thrombosis (summarized in Figure 2). We should not accept the inevitability of a constant attrition of grafts to thrombosis and need to continue to identify risk factors and confirm appropriate screening and interventions for its prevention, almost certainly requiring collaborative multicentre trials. In the future, just as we now expand the specificity of HLA gene typing with molecular biology, genotyping for recognized thrombophilia genes in patients at risk will expand our ability to recognize and prevent thrombosis with targeted interventions drawn from the increasing array of anticoagulants now available. The contribution of thrombophilia to non-immune mechanisms of chronic allograft loss is also a potentially important but neglected area of research. PMID- 10528650 TI - Organization of organ donation--concepts and experiences in Niedersachsen/Ostwestfalen. PMID- 10528649 TI - Prophylaxis of cytomegalovirus infection in renal transplantation: new data for an old problem. PMID- 10528651 TI - The impact of television on attitudes towards organ donation--a survey in a German urban population sample. AB - BACKGROUND: The stagnation or decrease in organ donation rates since 1992 in Germany has partly been attributed to the negative impact of reports about organ donation periodically presented by German television between 1992 and 1997. This study was performed to elucidate the impact of the media on the public's attitudes towards organ donation. METHODS: A questionnaire concerning different aspects of organ donation was sent to the parents of pupils of a high school in a German city in 1994 and 1998. RESULTS: In 1994, 940 adults could be identified who had (TV+, n = 546) or had not (TV-, n = 394) followed at least one television discussion about the topic. In 1998, the group consisted of 756 (TV+, n = 443 and TV-, n = 313) adults. The discriminating question was of sufficient strength to reveal significant differences between TV(+) and TV(-) respondents. Contrary to an assumed negative impact of TV, differences between the groups were detectable mainly in questions regarding information, but not in those which dealt with personal fears and concerns. The main results obtained in both surveys were identical. Furthermore, from 1994 to 1998 there was a trend in favour of information and organ donation for TV(+) but not for TV(-) respondents. CONCLUSION: The assumption that TV has had a negative impact on donation rates must be rejected. Therefore, the stagnation/decline in donation rates must be attributed to other factors. PMID- 10528652 TI - Cerebral aneurysms in patients with autosomal dominant polycystic kidney disease- to screen, to clip, to coil? PMID- 10528653 TI - Spontaneous resolution and recurrence of hypercalcemia in primary hyperparathyroidism--anecdotal observations with potential implications for parathyroid pathophysiology in renal disease. PMID- 10528654 TI - Exclusion of the candidate genes ACE and Bcl-2 for six families with nephronophthisis not linked to the NPH1 locus. AB - BACKGROUND: Nephronophthisis (NPH) is an autosomal recessively transmitted kidney disease, characterized by cyst formation at the cortico-medullary junction, and a sclerosing tubulointerstitial nephropathy. Juvenile nephronophthisis (NPH1) is the most common genetic cause of renal failure in children and maps to chromosome 2q12-q13. The responsible gene NPHP1 has been identified and encodes for nephrocystin. Not all families with NPH demonstrate linkage to that locus. METHODS: We studied six families with NPH without linkage to the NPH1 locus. In order to attempt identification of a new causative gene, the candidate genes ACE (angiotensin converting enzyme) and Bcl-2 (B cell leukaemia/lymphoma 2 gene) originating from mouse models, were examined. For the six families highly polymorphic microsatellites covering the whole candidate gene regions were haplotyped and linkage analysis was performed. RESULTS: Haplotype analyses of all families examined were incompatible with linkage of the disease status to ACE or Bcl-2. Linkage analysis excluded both candidate gene regions with a LOD-score of < -2. CONCLUSIONS: This study excluded the candidate genes ACE and Bcl-2 for NPH. Additional linkage studies need to be performed in order to identify further genes responsible for nephronophthisis. PMID- 10528655 TI - Epidemiological data of treated end-stage renal failure in the European Union (EU) during the year 1995: report of the European Renal Association Registry and the National Registries. AB - BACKGROUND: The new Centre Questionnaire, mainly based on the collection of epidemiological data, was launched in 1996 and the overall response rate of centres for the 15 countries constituting the European Union (EU) reached 82.2% (66-100%) for 1995. RESULTS: We could derive the following information for a general population of 372.6 million. In 1995, the incidence of new end-stage renal failure (ESRF) patients (Ni/P) was 120 p.m.p. (per million population) with a clear north to south/west gradient (69 in Ireland, 131 in Italy and 163 in Germany). The incidence of ESRF deaths (No/P) was 67 p.m.p. (from 35 in Ireland to 89 in Germany). The net increase of patients was therefore 53 p.m.p. (from 13 in Greece to 74 in Germany). The point prevalence of treated ESRF patients (Ns/P) alive on 31 December 1995 was 644 p.m.p. (from 444 in Finland to 773 in Italy). The mean increase in the stock of ESRF patients was +8.2% (4.6 to 13.0) as a linear rate and +0.085 as a fractional rate (exponential). The first treatment of new patients (Ni) was haemodialysis (HD; 81%), peritoneal dialysis (PD; 18%) and pre-emptive renal transplantation (Tx; 1%). The latest treatment for patients still alive was HD (58.5%), PD (9%) or functional Tx (32.5%). The number of Tx was 30 p.m.p. (from 14 in Greece to 45 in Spain). The death rate was 10.4% for all those with ESRF, with 14.4% for those dialysed and 2.2% for transplanted patients. In 1995, 6.5% of dialysed patients received a graft and 4.0% of transplant patients returned to dialysis. The linear expansion rate of the dialysis pool and the transplant pool was respectively 8.3% and 7.9%. CONCLUSIONS: This data shows considerable disparities among countries of the EU which merit further evaluation. Also this analysis by the ERA Registry provides data of value for health and economic purposes. PMID- 10528656 TI - Deaths within 90 days from starting renal replacement therapy in the ERA-EDTA Registry between 1990 and 1992. AB - BACKGROUND: Patients who die within 90 days of commencing renal replacement therapy (RRT) may be recorded by some centres and not others, and hence data on mortality and survival may not be comparable. However, it is essential to compare like with like when analysing differences between modalities, centres and registries. It was decided, therefore, to look at the incidence of deaths within 90 days in the ERA-EDTA Registry, and to try to define the characteristics of this group of patients. METHODS: Between 1 January 1990 and 31 December 1992, 78 534 new patients started RRT in 28 countries affiliated to the ERA-EDTA Registry. Their mean age was 54 years and 31% were over 65 years old. Eighty-two per cent of the patients received haemodialysis (HD), 16% peritoneal dialysis (PD) and 2% had preemptive transplantation as first mode of treatment. RESULTS: From January 1990 to March 1993 the overall incidence of deaths was 19% and 4% of all patients died within 90 days from the start of RRT. Among those dying within 90 days 59% were over 65 years compared to 53% over 65 years in those dying beyond this time (P<0.0001). The modality of RRT did not influence the distribution of deaths before and after 90 days. Vascular causes and malignancy were more common in those dying after 90 days, while there were more cardiac and social causes among the early deaths. Mortality from social causes was twice as common in the elderly, who had a significantly higher chance of dying from social causes within 90 days compared to those aged under 65 years. The overall incidence of deaths within 90 days was 3.9% but there was a wide variation between countries, from 1.8% to 11.4%. Finally, patient survival at 2 years was markedly influenced in different age groups when deaths within 90 days were taken into account. CONCLUSIONS: The incidence of deaths within 90 days from the start of RRT was 3.9%, with a marked variation between countries ranging from 1.8% to 11.4%, which probably reflects mainly differences in reporting these deaths, although variable selection criteria for RRT may contribute. Deaths within 90 days were significantly more frequent in elderly patients with more early deaths resulting from cardiac and social causes, while vascular causes of death and malignancy were more common in those dying after 90 days. Patient survival analyses should take into account deaths within 90 days from the start of RRT, particularly when comparing results between modalities, countries and registries. PMID- 10528657 TI - Improved survival in renal replacement therapy in Europe between 1975 and 1992. An ERA-EDTA Registry study. AB - BACKGROUND: The prevalence of Renal Replacement Therapy (RRT) is rising steadily, worldwide and in Europe. One reason for this is an increasing number of patients starting RRT, but improving survival on RRT may also be contributing. MATERIAL AND METHODS: In an ERA-EDTA Registry study we have examined survival of patients with Standard Primary Renal Disease, or Diabetes, aged 20 to 75 years, who started RRT with haemodialysis (HD) or peritoneal dialysis (PD) between 1975 and 1992. Altogether close to a quarter of a million patients were included in the analysis which included conventional survival analysis of comparable subgroups of the whole cohort as well as Cox regression. RESULTS: After accounting for age, mode of initial treatment, and diagnosis, an improvement in survival of RRT patients was evident. From Cox regression it was calculated the risk for death decreased by about 5% annually during the time period 1975 1992. Patients who started RRT using PD experienced a higher mortality than those starting with HD. According to Cox regression the relative risk ratio for death was 1.25 for the whole period. The difference in survival between patients starting with PD or HD diminished during the observation period (1975-1992). DISCUSSION: The survival prospects of a patient presenting with end stage renal disease were considerably better in the early 1990s compared to the mid 1970s. This is reassuring despite the fact that mortality on RRT remains high. The higher mortality of RRT patients who started with PD is probably an 'historical' observation as the techniques of this treatment modality have improved considerably since the 1980s which was the time period from which came most of the data for the analysis. PMID- 10528658 TI - Insulin's acute effects on glomerular filtration rate correlate with insulin sensitivity whereas insulin's acute effects on proximal tubular sodium reabsorption correlation with salt sensitivity in normal subjects. AB - BACKGROUND: Insulin induces sodium retention by increasing distal tubular sodium reabsorption. Opposite effects of insulin to offset insulin-induced sodium retention are supposedly increases in glomerular filtration rate (GFR) and decreases in proximal tubular sodium reabsorption. Defects in these opposing effects could link insulin resistance to blood-pressure elevation and salt sensitivity. METHODS: We assessed the relationship between the effects of sequential physiological and supraphysiological insulin dosages (50 and 150 mU/kg/h) on renal sodium handling, and insulin sensitivity and salt sensitivity using the euglycaemic clamp technique and clearances of [131I]hippuran, [125I]iothalamate, sodium, and lithium in 20 normal subjects displaying a wide range of insulin sensitivity. Time-control experiments were performed in the same subjects. Salt sensitivity was determined using a diet method. RESULTS: During the successive insulin infusions, GFR increased by 5.9% (P = 0.003) and 10.9% (P<0.001), while fractional sodium excretion decreased by 34 and 50% (both P<0.001). Distal tubular sodium reabsorption increased and proximal tubular sodium reabsorption decreased. Insulin sensitivity correlated with changes in GFR during physiological (r = 0.60, P = 0.005) and supraphysiological (r = 0.58, P = 0.007) hyperinsulinaemia, but not with changes in proximal tubular sodium reabsorption. Salt sensitivity correlated with changes in proximal tubular sodium reabsorption (r = 0.49, P = 0.028), but not in GFR, during physiological hyperinsulinaemia. Neither insulin sensitivity or salt sensitivity correlated with changes in overall fractional sodium excretion. CONCLUSIONS: Insulin sensitivity and salt sensitivity correlate with changes in different elements of renal sodium handling, but not with overall sodium excretion, during insulin infusion. The relevance for blood pressure regulation remains to be proved. PMID- 10528659 TI - An evaluation of the Banff classification of early renal allograft biopsies and correlation with outcome. AB - BACKGROUND: The Banff classification for assessment of renal allograft biopsies was introduced as a standardized international classification of renal allograft pathology and acute rejection. Subsequent debate and evaluation studies have attempted to develop and refine the classification. A recent alternative classification, known as the National Institutes of Health Collaborative Clinical Trials in Transplantation (NIH-CCTT) classification, proposed three distinct types of acute rejection. The 1997 Fourth Banff meeting appeared to move towards a consensus for describing transplant biopsies, which incorporated both approaches. Patients who received a renal allograft at the Oxford Transplant Centre were managed by a combination of protocol and clinically indicated biopsies. We have undertaken a retrospective analysis of the biopsies correlated with the clinical outcome to test the prognostic value of the original Banff (Banff 93-95) and NIH-CCTT classifications. METHODS: Three hundred and eighty-two patients received renal allografts between May 1985 and December 1989, and were immunosuppressed using a standard protocol of cyclosporine, azathioprine and steroid. Adequate 5-year follow-up data were available on 351 patients, and of these, 293 had at least one satisfactory biopsy taken between days 2 and 35 after transplantation, the latter patients forming the study group. The D2-35 biopsies taken from these patients, which were not originally reported according to the Banff classification, were re-examined and classified according to the Banff 93 95 protocols. For each patient the biopsy found to be the most severely abnormal was selected, and the Banff and NIH-CCTT grading compared with the clinical outcome. RESULTS: Seven hundred and forty-three biopsies taken from 293 patients between days 2 and 35 after transplantation were examined and the patients categorized on the basis of the 'worst' Banff grading as follows. Normal or non rejection, 20%; borderline, 34%; acute rejection grade I (AR I), 18%; AR IIA, 6%; AR IIB, 14%; AR III, 1%; AR IIIC, 3%; widespread necrosis 3%. The clinical outcome for the last two groups combined was very poor with 18% of grafts functioning at 3 months and 6% at 5 years. The other groups with vascular rejection (AR IIB and AR III) had an intermediate outcome, graft survival being 78% at 3 months and 61% at 5 years. The remaining four groups (normal, borderline, cellular AR I and AR IIA) had the best outcome: graft survival 95% at 3 months and 78% at 5 years with virtually no difference between the four groups. Three forms of acute rejection, namely tubulo-interstitial, vascular and transmural vascular, were identified, but only the latter two categories were associated with a poor outcome. CONCLUSIONS: The eight sub-categories of the Banff classification of renal allograft biopsies are associated with three different prognoses with respect to graft survival in the medium term. These three prognostic groups correspond to the three NIH-CCTT types. The data provide support for the consensus developed at Banff 97 separating tubulo-interstitial, vascular and transmural vascular rejection (types I, II and III acute rejection). PMID- 10528660 TI - A spectrum of clinicopathological features of nephropathy associated with POEMS syndrome. AB - BACKGROUND: In POEMS syndrome, substantial involvement of the kidney can occur and is reflected by proteinuria, haematuria, renal dysfunction, and renal failure requiring dialysis therapy. The mechanism by which renal dysfunction is induced and progresses to end-stage renal disease remains obscure. A pathogenic role of cytokines and growth factors has recently been implicated. METHODS: We reviewed cases of 52 Japanese patients with confirmed renal pathology who were reported in the literature, and personally analysed renal tissues from 22 subjects including nine patients of our own. Interleukin-6 (IL-6) and vascular endothelial growth factor (VEGF) were measured in our cases. RESULTS: Despite relatively mild renal symptomatology, about half of the cases had azotaemia with creatinine levels above 1.5 mg/dl and the BUN/creatinine ratio markedly raised by volume contraction or wasting. One-tenth of patients were placed on haemodialysis because of advanced or end-stage renal disease. Bilateral and unilateral contracted kidneys were found in four and two cases respectively. Pathological analyses disclosed two major changes: glomerular alterations and endarteritis like lesions of renal small arteries. The former included glomerular enlargement, cellular proliferation, mesangiolysis and marked swelling of endothelial mesangial cells. This structural disorganization led to a reduction in renal function to some degree by impairing the glomerular circulation. Vasculopathy of the small artery probably resulted in progressive renal damage and ultimately to kidney contraction. Serum IL-6 was elevated in about 40% of cases. IL-6 levels were found to be high in the ascites of three patients who were examined. In different studies, an increased level of VEGF was found in the peripheral blood (75-100%; overall 92.3%), but no apparent correlation with glomerular alterations was observed. CONCLUSION: POEMS nephropathy can be one cause of end-stage renal disease with variable intrarenal pathological changes of a microangiopathic nature which have differential influences on renal function. A pathogenic role for VEGF in POEMS syndrome appears to be likely, but its causal relation to the nephropathy awaits further investigation. PMID- 10528661 TI - The short- and long-term outcomes of membranous nephropathy treated with intravenous immune globulin therapy. Kanazawa Study Group for Renal Diseases and Hypertension. AB - BACKGROUND: A considerable diversity in prognosis is seen with membranous nephropathy (MN). A recent report showed beneficial effects of immune globulin (Glb) therapy in Heymann nephritis, a rat model of MN. However, the early and late clinical effects of Glb in human MN have remained unclear. METHODS: We studied retrospectively 86 patients with primary MN from 1965 to 1988 who were followed for at least 5 years, or until renal or actual death. Thirty patients were non-randomly treated with 1-3 courses of intravenous immune globulin, 5-10 g/day (100-150 mg/kg/day) for 6 consecutive days. Based on electron microscopic (EM) findings, the patients were divided into two subtypes, i.e. homogeneous type with synchronous electron-dense deposits, and heterogeneous type with various stages of dense deposits, due to their different clinical outcomes. RESULTS: There was no difference in the initial clinicopathological states between Glb (n = 30) and non-Glb group (n = 56) (70 vs. 68% in nephrotic state; 37 vs. 39% in female, 50 vs. 52% in homogeneous type, 50 vs. 48% in heterogeneous type respectively). For the homogeneous type, at 6 months post-treatment, Glb therapy had induced earlier remission as compared to non-Glb treatments with corticosteroid alone or together with cyclophosphamide (57 vs. 10% respectively, P = 0.006). However, there was no significant difference in the early therapeutic effect for the heterogeneous type (13% for Glb vs. 5% for non-Glb in remission after 6 months), or in the final outcome for all groups (18% for Glb vs. 10% for non-Glb in renal death after 15 years). No adverse effects were recorded during or after Glb therapy. CONCLUSIONS: Our results suggest that short-term relatively low-dose intravenous Glb therapy has a beneficial effect in the earlier induction of remission in a subgroup of MN, the homogeneous type with EM findings of synchronous electron-dense deposits, but does not alter the long-term outcome of human MN. PMID- 10528663 TI - Dyslipidaemia and the progression of renal disease in chronic renal failure patients. AB - BACKGROUND: Dyslipidaemia is common in patients with chronic renal failure (CRF), and there is increasing evidence to support the role of dyslipidaemia as a contributing factor in the progression of chronic renal disease. However, few prospective studies have been carried out which address the possible relationship between dyslipidaemia and the rate of progression of renal disease in patients with renal failure. METHODS: Between January 1985 and December 1997, we prospectively assessed the risk of CRF progression to dialysis in a cohort of 138 patients. Forty CRF patients reached end-stage renal disease (ESRD) and had to start supportive therapy during the follow-up period [group ESRD(+)]. The remaining 98 CRF patients served as controls [group ESRD(-)]. Potential clinical and laboratory risk factors for more rapid CRF decline to dialysis, including lipid abnormalities and baseline creatinine clearance were determined at the start of the follow-up period. RESULTS: Several significant differences were found in univariate analysis between the two groups of CRF, ESRD(+) and ESRD(-), namely a shorter follow-up period, a lower level of baseline creatinine clearance, a faster rate of creatinine clearance decline, a higher level of serum triglycerides, fibrinogen, total homocyst(e)ine and proteinuria, and a lower level of serum high-density lipoprotein in the ESRD(+) group than in the ESRD(-) group. However, by multivariate Cox analysis proteinuria [relative risk (95% confidence interval) 1.32 (1.16-1.50) for each g/day P = 0.001], baseline creatinine clearance [0.53 (0.40-0.70) for each 10 ml/min, P = 0.001] and chronic interstitial nephritis and hypertensive nephrosclerosis [0.38 (0.17-0.84) for presence, P = 0.005] were the only significant risk factors for CRF progression to dialysis. Hypertriglyceridaemia and male gender were selected in the final model, but were of borderline significance. CONCLUSIONS: These results suggest a limited role for dyslipidaemia in the progression of chronic renal disease to dialysis in CRF patients, in contrast with the powerful influence of proteinuria, baseline creatinine clearance and nephropathy type in predicting this progression. PMID- 10528662 TI - Continuous veno-venous haemodialysis with a novel bicarbonate dialysis solution: prospective cross-over comparison with a lactate buffered solution. AB - OBJECTIVE: To compare acid-base balance, lactate concentration and haemodynamic parameters during continuous veno-venous haemodialysis (CVVHD) using bicarbonate or a lactate buffered dialysate. METHODS: DESIGN: prospective randomized cross over design; SETTING: Multicentre combined adult surgical and medical intensive care units. Patients; 26 critically ill patients starting CVVHD for acute renal failure. INTERVENTIONS: Each patient to receive 48 h of bicarbonate dialysate and 48 h of lactate dialysate with the order of the 48 h block randomized at trial entry. RESULTS: The serum bicarbonate increased from baseline in both the lactate and bicarbonate groups over the first 48 h of treatment (16.3+/-1.53 to 22.2+/ 1.41 mmol/l and 18.9+/-2.02 to 22.2+/-1.18 mmol/l, respectively) and continued to rise towards normal over the next 48 h after cross-over to the other dialysate. The H+ and pCO2 only trended higher in the lactate group. Unlike the acid base parameters, serum lactate levels varied depending on the dialysate composition. The patients initially randomized to the lactate dialysate had higher serum lactate levels and these tended to increase further after 48 h of dialysis from 2.4+/-0.8 to 2.6+/-0.4 mmol/l. However, in the following 48 h the lactate levels fell to 1.8+/-0.6 (P = 0.039) while patients were being treated with the bicarbonate dialysate. Similar results were seen in the patients initially randomized to the bicarbonate dialysate. Serum lactate remained stable over the first 48h (1.4+/-0.2 to 1.5+/-0.1 mmol/l) but after cross-over to the lactate dialysate increased to 3.1+/-0.7 mmol/l (P = 0.051). Overall, lactate levels were significantly higher during dialysis with lactate buffered solution than bicarbonate buffered solution (2.92+/-0.45 vs. 1.61+/-0.25 mmol/l P = 0.01). Mean arterial pressure trended higher during bicarbonate dialysis but did not reach statistical significance (lactate vs. bicarbonate; 71.1+/-3.1 vs. 81.3+/-5.8 mm Hg). Subgroup analysis of the patients with abnormal liver indices or increased lactate levels at initiation of dialysis (n = 15) revealed only a trend toward better bicarbonate control (lactate vs. bicarbonate; 22.00+/-1.73 vs. 22.86+/ 1.09, P = 0.2). However, in this group with hepatic insufficiency elevations in serum lactate were even greater during lactate compared to the bicarbonate dialysis (3.39+/-0.68 vs. 1.78+/-0.42 P = 0.036). Patients who had elevations of lactate during lactate dialysis had a high mortality (6 of 7). These patients had an even greater disparity in lactate levels (4.3+/-1.4 vs. 1.3 +/-0.3) and blood pressure (68.0+/- 7.7 vs. 87.2+/-17.1) between lactate and bicarbonate dialysis. Due to small patient numbers these comparisons did not achieve statistical significance. CONCLUSION: During continuous veno venous haemodialysis a bicarbonate buffered dialysis solution provided equal acid-base control but maintained more normal lactate levels than a lactate buffered dialysis solution. PMID- 10528664 TI - The PTH-calcium curve and the set point of calcium in primary and secondary hyperparathyroidism. AB - BACKGROUND: The regulation of PTH secretion by calcium is altered in patients with primary hyperparathyroidism (HPT). A similar abnormality may occur in secondary HPT, but comparisons of PTH secretion in normal subjects and those with secondary HPT have given contrasting results. Differences in baseline serum ionized calcium (ICa) may partly account for these conflicting results. The aim of the present study was to evaluate whether the regulation of PTH secretion by calcium differs from normal in patients with primary and secondary HPT and to determine whether serum calcium concentration per se can affect the set point of calcium and the PTH-calcium relationship. METHODS: The PTH-ICa relationship and the set point of ICa were evaluated in 19 patients with primary HPT (1-HPT), 16 normocalcaemic patients with secondary HPT (2-HPT; PTH 344+/-191 pg/ml), 19 hypercalcaemic patients with secondary HPT (3-HPT; PTH 806+/-254 pg/ml) and 14 healthy volunteers, by inducing hypocalcaemia and hypercalcaemia in order to maximally stimulate or inhibit PTH secretion. In five 1-HPT patients the PTH-ICa curve was restudied after normalization of serum ICa by pamidronate. Parathyroid gland volume was determined by measuring gland size at parathyroidectomy or by means of high-resolution color Doppler ultrasonography. RESULTS: In 1-HPT patients the PTH-ICa curve, constructed using maximal PTH secretion induced by hypocalcaemia as 100%, was shifted to the right, the set point of ICa was increased, and the slope of the curve was reduced when compared to normal subjects. After normalization of baseline serum ICa by pamidronate, a shift of the PTH-ICa curve towards normal and a reduction in the set point of ICa was observed. However, basal PTH and maximal PTH secretion induced by hypocalcaemia increased, minimal PTH secretion induced by hypercalcaemia remained increased and the slope of the curve did not change significantly. The alterations in the PTH ICa relationship in hypercalcaemic patients with secondary HPT were similar to those found in 1-HPT patients. In normocalcaemic patients with secondary HPT baseline PTH, maximal and minimal PTH secretion and parathyroid gland size were reduced compared to 3-HPT patients. Compared to normal subjects, 2-HPT patients showed greater calcium-induced minimal PTH secretion. The increase in non suppressible PTH secretion resulted in a rightward shift of the PTH-ICa curve and an increase in the set point of ICa. A strong correlation was found, in both primary and secondary HPT, between the set point of ICa and baseline serum ICa, and between parathyroid gland size and baseline PTH, maximal PTH and minimal PTH. Multivariate regression analysis showed that baseline serum ICa was the main determinant of the set point of ICa in both primary and secondary HPT. CONCLUSIONS: (i) The regulation of PTH secretion by calcium is abnormal in secondary as well as in primary HPT. (ii) Parathyroid gland enlargement in secondary HPT is associated with reduced sensitivity to serum ICa and resistance of parathyroid gland to calcium-mediated PTH suppression, resulting ultimately in PTH hypersecretion, despite hypercalcaemia. (iii) The set point of calcium is strongly dependent on baseline serum calcium, and the PTH-ICa relationship can be affected by variations in serum ICa concentrations. Thus, when the set point of calcium and the PTH-ICa relationship are evaluated, possible differences in baseline serum ICa concentration among the patients should be taken into account. PMID- 10528665 TI - Improved clearance of iohexol with longer haemodialysis despite similar Kt/V for urea. AB - BACKGROUND: The efforts to improve the quality of haemodialysis (HD) has renewed the interest in the consequences of blood-flow distribution for removal of solutes. METHODS: To test the effects of HD time per se, 10 patients were studied in a cross-over fashion with HD for 3 h and 1 week later for 6 h, with similar blood urea Kt/Vs, achieved by adjusting the blood flow rate to 290 and 120 ml/min respectively. Injections of iohexol (MW 821 Dalton) were given 2 days prior to the dialysis sessions. Blood samples were taken before, during (6/HD), 1 and 24 h after the HD and analysed for concentrations of urea and iohexol. A urea on-line monitor (Gambro) was used for continuous recordings and sampling of dialysate. RESULTS: According to the study design the blood Kt/V for urea (Daugirdas II) was similar for 3 and 6 h HD, close to 1.0 (n.s), while the removed mass of urea showed that Kt/V was slightly and significantly higher for the 6 h HD. The 'apparent' mass of iohexol, defined as plasma concentration times estimated distribution volume, fell to 29% and 21% of pre-dialysis levels after 3 h and 6 h HD, respectively (P<0.01), but increased after HD, and more so after the short dialysis, reaching 46% of the predialysis mass 24 h after 3 h HD vs. 36% after 6 h HD (P<0.05). The removed mass of iohexol was 920+/-110 mg with 6h HD and 700+/ 81 mg with 3h HD, (P<0.01). Thus, the longer dialysis removed 32% more iohexol despite similar blood Kt/V for urea. CONCLUSION: The treatment time per se affects solute removal despite similar blood Kt/V for urea. This is particularly true for an intermediate-size molecule like iohexol. PMID- 10528666 TI - PDGF-AB release during and after haemodialysis procedure. AB - BACKGROUND: During haemodialysis blood membrane contact causes the release of the content of platelet alpha-granules, which contain platelet-derived growth factor (PDGF). In view of its possible role in accelerated atherosclerotic processes, we evaluated the intra- and post-dialytic changes in PDGF-AB serum levels during haemodialysis sessions performed using a cellulosic membrane. METHODS: Using the ELISA method, PDGF-AB, platelet factor-4 (PF4) and beta-thromboglobulin (beta-TG) levels were determined in peripheral blood, as well as in arterial and venous haemodialyser lines, in 10 patients each of whom underwent five consecutive dialysis sessions with a CU membrane. Blood samples were taken at 0, 15, 30, 60, 120, 180 and 240 min during dialysis and at 1, 4 and 20 h after the end of the session. In the same group of patients the levels of the same molecules were also determined after a heparin bolus injection of 4500 IU, blood samples were taken at 0, 15 and 30 min after injection of the bolus. RESULTS: PDGF-AB serum levels increased, remained consistently high during the haemodialysis session (in particular +134+/-20% after 30 min, P<0.001, and +140+/-5% after 240 min, P<0.001) and returned to basal values only after 20 h following the end of the session. PF4 and beta-TG showed a similar trend to PDGF. The heparin bolus injection caused only a small increase (+15+/-5% at 30 min) in PDGF-AB serum levels. CONCLUSIONS: PDGF-AB is released during dialysis mainly as consequence of the blood-membrane contact and it returns only slowly to basal values. PMID- 10528667 TI - Whole blood production of monocytic cytokines (IL-1beta, IL-6, TNF-alpha, sIL-6R, IL-1Ra) in haemodialysed patients. AB - BACKGROUND: The production of monocytic cytokines by isolated mononuclear cells after stimulation by phytohaemagglutinin (PHA) and lipopolysaccharide (LPS) is generally increased in haemodialysed (HD) patients. We performed whole blood (WB) cultures to evaluate cytokine production by blood cells inside their complex cellular and humoral network. METHODS: Diluted whole blood from HD patients (collected before dialysis) and controls was cultured alone with PHA (2.5 microg/ml) or LPS (1 and 3 microg/ml). Supernatants were collected after 24 and 48 h of culture, and concentrations of IL-1 beta, IL-6, TNF-alpha, sIL-6R and IL 1Ra were determined by ELISA. RESULTS: The low spontaneous production of IL 1beta, IL-6 and TNF-alpha in both patients and controls was not significantly modified by PHA. The lower dose of LPS (1 microg/ml) induced a significant but lower increase in production of IL-1beta, IL-6 and TNF-alpha in patients than in controls. In contrast, while it did not further increase their production in controls, the higher concentration of LPS (3 microg/ml) still increased their production in patients to the same level than in controls. The plasma concentrations of sIL-6R were higher in patients than in controls. In both groups, the sIL-6R concentration did not vary during the culture period whether the cells were stimulated or not with LPS or PHA. This suggests that the increased plasma levels of sIL-6R were not produced by blood cells. Despite a similar significant LPS and PHA induced production of IL-1Ra, the IL-1Ra/IL-1beta ratio was always higher in patients than in controls. CONCLUSION: Monocytes from HD patients in WB cultures are hyporesponsive to PHA and LPS for their IL-1beta, TNFalpha and IL-6 production in contrast to isolated monocytes that demonstrate signs of activation. If it reflects the in vivo situation it could partly explain the immune defect in uraemic and haemodialysed patients. Higher sIL-6R/IL-6 and IL-1Ra/IL-1beta ratios could also participate to the complex immune disturbances of HD patients by reducing the biological activity of two cytokines playing a major role in the immune and inflammatory network. PMID- 10528668 TI - Altered abdominal fat distribution and its association with the serum lipid profile in non-diabetic haemodialysis patients. AB - BACKGROUND: Disturbances of lipid and carbohydrate metabolism may be associated with the distribution of abdominal adiposity. However, little is known about the alteration of abdominal adiposity and its association with the serum lipid profile in haemodialysis patients. METHODS: We evaluated the distribution of abdominal adiposity by using computed tomography and examined its relationship with the serum lipid profile in 92 non-diabetic haemodialysis patients and 80 control subjects with normal renal function. Since the mean body mass index (BMI) and total body fat mass were significantly lower in the haemodialysis patients than in the control subjects, the subcutaneous abdominal fat area and the visceral fat area were standardized by body mass index and compared between the haemodialysis patients and the control subjects. RESULTS: Mean subcutaneous fat area/body mass index (SFA/BMI) was significantly lower, and mean visceral fat area/body mass index (VFA/BMI) was significantly higher in the haemodialysis patients (SFA/BMI, 2.40+/-0.12; VFA/BMI, 2.28+/-0.15) than in the control subjects (SFA/BMI, 3.75+/-0.21, P<0.01; VFA/BMI, 1.65+/-0.15, P<0.01). Consequently, visceral fat area/ subcutaneous fat area ratio was significantly higher in the haemodialysis patients (1.05+/-0.07) than in the control subjects (0.46+/-0.04, P<0.01). A scattered plot of visceral fat area relative to BMI revealed that visceral fat area was higher in the haemodialysis patients than in the control subjects at any BMI level. A simple regression analysis showed that BMI, total body fat mass, subcutaneous fat area and visceral fat area were all associated with serum triglycerides and the atherogenic index, (total cholesterol HDL cholesterol)/HDL cholesterol. Furthermore, a multiple regression analysis indicated that the visceral fat area was the best predictor for either the atherogenic index or triglycerides among these fat components. CONCLUSIONS: These data indicate that haemodialysis patients exhibited a visceral fat accumulation irrespective of BMI, and this shift of abdominal adiposity might be associated with disturbance of the serum lipid profile in non-diabetic haemodialysis patients. PMID- 10528669 TI - Improved bacteriological surveillance of haemodialysis fluids: a comparison between Tryptic soy agar and Reasoner's 2A media. AB - BACKGROUND: Accurate microbiological surveillance in haemodialysis centres is important as end-stage renal patients can suffer from pyrogenic reactions due to bacterial contamination of dialysis fluids. To evaluate the microbiological quality of haemodialysis fluids, special nutrient-poor culture techniques are necessary. Although the Association for the Advancement of Medical Instrumentation (AAMI) recommends Tryptic soy agar (TSA) as the standard agar, several studies have resulted in a general preference for Reasoner's 2A (R2A) agar, as it appeared to be more sensitive in demonstrating contamination of typical haemodialysis associated bacteria. In the Netherlands TSA is still used for culturing dialysate, while dialysis water is cultured on R2A. Therefore, the aims of our study were to evaluate bacterial yields of dialysis fluids on both media, and to qualify their use in routine microbiological monitoring within our haemodialysis centre. METHODS: Between April 1995 and March 1996, 229 samples of pre-treated and final purified dialysis water, and samples of dialysates were collected. The specimens were aseptically taken from the tap, various points of the reverse osmosis (RO) water-treatment system, and the effluent tubes of 32 bicarbonate haemodialysis machines. Samples of 0.1 ml were inoculated in duplicate on spread plates with TSA and R2A agars. After 10 days of incubation at 25+/-2 degrees C, the numbers of colonies were quantified. The ranges of spread were taken 0-100 and 0-200 colony-forming units per milliliter (c.f.u./ml). RESULTS: The R2A agar had significantly higher colony counts than TSA agar for both dialysis water and dialysates. Considering 100 c.f.u./ml as the upper allowable bacterial limit for all dialysis fluids, microbiological non-compliance (bacterial growth) would be missed in 16% when using only TSA media (TSA < or =100 c.f.u./ml and R2A >100 c.f.u./ml), while this was 3% when using only R2A (TSA >100 c.f.u./ml and R2A < or =100 c.f.u./ml, P<0.0001). Considering 200 c.f.u./ml as the upper limit, non-compliance would have been missed in 10% when using only TSA (TSA < or =200 c.f.u./ml and R2A >200 c.f.u./ml), and 2% when using R2A (TSA > 200 c.f.u./ml and R2A < or =200 c.f.u./ml, P = 0.0011). CONCLUSIONS: Microbiological surveillance of haemodialysis fluids, including pre treated dialysis water samples collected from RO treatment systems, can be performed more precisely with R2A media than TSA, when incubated at 25+/-2 degrees C for 10 days. PMID- 10528670 TI - Dialysis in The Netherlands: the clinical condition of new patients put into a European perspective. NECOSAD Study Group. Netherlands Cooperative Study on the Adequacy of Dialysis phase 1. AB - BACKGROUND: The unadjusted annual mortality rate among prevalent Dutch dialysis patients increased from 1981 to 1992. Part of this increase may be attributed to the ageing of the dialysis population, but hardly any data were available on other important prognostic features of new Dutch dialysis patients, such as co morbidity and other aspects of their clinical condition. The aim of the present study was to obtain these data and to put them into a European perspective. METHODS: Two hundred and fifty consecutive new patients were included in this prospective multi-centre study. Data were collected 3 months after start of dialysis. Multivariate linear regression analysis was used to explain the variability of parameters of nutritional state and blood pressure. RESULTS: Mean age was 57 years, co-morbid conditions were present in 51%, diabetes mellitus in 18%, and cardiovascular disease in 28%. Decreased protein intake was related to diminished residual renal function. Our patients did not have more co-morbidity than Dutch patients participating in a European study some years earlier. Comparison with other studies was complicated by the use of different definitions of co-morbidity and of selected patient populations. CONCLUSIONS: Despite the fact that Dutch dialysis patients have become older and the incidence of diabetic nephropathy has increased, no conclusions could be drawn on a concomitant increase in co-morbidity. This patient group may serve as a reference population to study future changes in patient case-mix within the Netherlands. Furthermore, the use of common international definitions of co-morbidity is needed to be able to make comparisons of survival data. PMID- 10528671 TI - Living donor kidney transplants: a biopsy study 1 year after transplantation, compared with baseline changes and correlation to kidney function at 1 and 3 years. AB - INTRODUCTION: Chronic changes in biopsies from long-term stable kidney allografts have been reported to correlate with graft prognosis. Morphological changes in baseline ('zero-hour') biopsies have been described as well, but their importance for long-term prognosis have been less clear. The aim of the present study was to evaluate biopsy changes from baseline to 1 year after transplantation in patients receiving kidneys from living donors, and to assess the possible prognostic implications of these findings. METHODS: Light microscopical changes in 18 gauge full-core biopsies were scored semi-quantitatively in 33 patients 1 year after transplantation, and compared to baseline changes previously reported [1]. All cases were also examined with transmission electron microscopy. The semi quantitative data from baseline and at 1 year were correlated with kidney function 1 and 3 years after transplantation. The reproducibility of baseline findings regarding arteriosclerosis and arteriolar hyalinosis was tested by comparison with biopsies 1 week after transplantation (n = 43). RESULTS: We found a significant increase in mesangial glomerular sclerosis (P<0.001), interstitial fibrosis/tubular atrophy (if/ta) (P = 0.002), and mononuclear cell interstitial infiltration (P = 0.003) after 1 year, compared to baseline changes. There was an increase of arteriosclerosis (P = 0.028) and arteriolar hyalinosis (P = 0.006) when compared to biopsies taken 1 week after transplantation, but not when compared to the 'zero-hour' findings. Electron microscopy revealed one case of recurrent immune-complex glomerulonephritis and another case of recurrent light chain deposition kidney disease. Comparing 1-week vascular findings with baseline gave a low level of reproducibility, probably due to sampling error. Baseline biopsy findings could not predict long-term kidney function. In the 1-year biopsy, if/ta was significantly correlated with serum creatinine (P = 0.007) and glomerular filtration rate (GFR) (P<0.001) at 1 year, with serum creatinine at 3 years (P = 0.011), and with the first-year cumulative dose of methylprednisolone (P = 0.004). Serum creatinine at 1 year, however, was found to be the most accurate predictor of 3-year kidney function (P<0.001). Donor age was correlated to kidney function at 3 years (P = 0.013) but not at 1 year after transplantation. CONCLUSION: Morphological changes in baseline biopsies of living donor kidneys tend to become more pronounced in well-functioning allografts during the first year after transplantation. In the 1 year biopsy, if/ta seems to be the most reliable variate for grading of chronic changes. However, 1-year serum creatinine predicted long-term kidney function more precisely than did the biopsy scores. Based on the results of the present study, a protocol 1-year biopsy does not seem warranted in the management of the graft recipient with a stable kidney function. PMID- 10528672 TI - Impaired kidney transplant survival in patients with antibodies to hepatitis C virus. AB - BACKGROUND: With a few exceptions, most published studies do not show an influence of antibodies to the hepatitis C virus (HCV) on the success of a kidney transplant. METHODS: We studied all our renal transplant recipients who had received kidneys from cadaver donors (n = 335) and had been treated with quadruple immunosuppression (steroids, azathioprine, and antilymphocyte antibodies, followed by cyclosporin). We had information on the status of the hepatitis C antibodies before and/or after the transplant in 320 cases (95.5%; in 300, pre-transplant). Patients with HCV antibodies before and/or after the transplant were considered to be HCV positive (HCV+). RESULTS: The HCV+ patients had more time in dialysis and a greater number of transfusions, hyperimmunized cases, and re-transplants. The evolution in the first post-transplant year was similar in both groups, but afterwards, the HCV+ patients had proteinuria more often as well as worse kidney function. The survival rate of the graft was significantly less in the HCV+ cases: 90.6, 68.3 and 51.0% at respectively 1, 5 and 10 years, compared with 91.5, 84.7 and 66.5% in HCV-patients (P<0.01). The patient survival rate was: 96.4, 87.0, and 71.9% in the HCV+ patients at 1, 5, and 10 years, compared with 98.2, 96.0 and 90.0% in the HCV- cases respectively (P<0.01). The differences remained the same in stratified studies according to time spent in dialysis or pre/post-transplant evolution of HCV antibodies, even when immunologically high-risk patients were excluded. In multivariant analysis, the presence of HCV antibodies acted as a independent prognostic factor for the survival of the kidney and the patient: 3.0 (1.8-5.0) and 3.1 (1.2-7.8) odds ratio (95% of the confidence interval), respectively. The main cause of death among HCV+ patients was cardiovascular; there was no apparent increase in mortality rate due to infections or chronic liver disease. The loss of organs was mainly due to chronic nephropathy or death with a functioning kidney. CONCLUSION: The presence of hepatitis C antibodies, before or after transplantation, is associated with a worse long-term survival rate for both the patient and the transplanted kidney in our patients treated with quadruple therapy. PMID- 10528673 TI - Chronic graft-versus-host disease complicated by membranous glomerulonephritis. PMID- 10528674 TI - Renal and other organ failure caused by germanium intoxication. PMID- 10528675 TI - Thrombotic microangiopathy in a patient with chronic myelocytic leukaemia treated with alpha-interferon. PMID- 10528676 TI - Haemolytic-uraemic syndrome following human parvovirus infection in a previously fit adult. PMID- 10528677 TI - Combination of APC resistance and acquired protein S deficiency in a haemodialysis patient with recurrent A-V shunt thrombosis. PMID- 10528678 TI - AA amyloidosis in Takayasu's arteritis--long-term survival on maintenance haemodialysis. PMID- 10528679 TI - Serious bleeding in a haemodialysis patient treated with recombinant hirudin. PMID- 10528680 TI - Survival of a human immunodeficiency patient with nucleoside-induced lactic acidosis--role of haemodialysis treatment. PMID- 10528681 TI - Successful twin pregnancy in a patient on long-term haemodialysis. PMID- 10528682 TI - Tacrolimus (FK506)-induced severe and late encephalopathy in a renal transplant recipient. PMID- 10528683 TI - The transplant recipient with elevated creatinine, and normal ultrasonography- detection of ureteral stenosis by magnetic resonance tomography. PMID- 10528684 TI - Post-renal transplant azathioprine-induced pancreatitis. PMID- 10528685 TI - A case of acute spontaneous epidural haematoma in a chronic renal failure patient undergoing haemodialysis: successful outcome with surgical management. PMID- 10528686 TI - Renal colic in a patient with anti-phospholipid antibodies and factor V Leiden mutation. PMID- 10528687 TI - Hyperkalaemia in a patient with hepatic cirrhosis. PMID- 10528688 TI - Bilateral thickening of the pericapsular renal area in a patient with refractory oedema of the legs. PMID- 10528689 TI - The loop of Henle, a turning-point in the history of kidney physiology. PMID- 10528691 TI - Health policies and epidemiology of diabetes among dialysed patients in France. PMID- 10528690 TI - Iconographic archives of European nephrology: arteriocapillary fibrosis. PMID- 10528692 TI - Survival comparisons in haemodialysis between France and USA. PMID- 10528693 TI - Ringed Gore-Tex for haemodialysis access. PMID- 10528694 TI - Insulin resistance in patients with adult polycystic kidney disease. PMID- 10528695 TI - Severe nephrotic syndrome without oedema in a patient with HIV associated nephropathy. PMID- 10528696 TI - Haemolytic uraemic syndrome after gemcitabine treatment for pancreatic carcinoma. PMID- 10528697 TI - Ancient wisdom on volume control. PMID- 10528698 TI - Increasing of renal replacement therapy (RRT) in diabetic patients in Madrid. PMID- 10528699 TI - Nephrotic syndrome following cefixime therapy in a 10-month-old girl: spontaneous resolution without corticosteroid treatment. PMID- 10528700 TI - Leptinaemia in patients dialysed with different buffers and dialysis membranes. PMID- 10528701 TI - The effect of vitamin C on laboratory tests in haemodialysis patients: is there a relationship between the administered vitamin C dose and serum uric acid levels? PMID- 10528702 TI - Intravenous analgesic use in dialysis patients. PMID- 10528703 TI - Sodium citrate for filling haemodialysis catheters. PMID- 10528705 TI - Healthcare systems. An international review. Introduction. PMID- 10528704 TI - Is tissue factor a mediator of fibrin deposition in glomerular pathology? PMID- 10528706 TI - Healthcare systems--an international review: an overview. AB - Based on the source of their funding, three main models of healthcare can be distinguished. The first is the Beveridge model, which is based on taxation and has many public providers. The second is the Bismarck 'mixed' model, funded by a premium-financed social insurance system and with a mixture of public and private providers. Finally, the 'Private Insurance model' is only in existence in the US. The present report explores the impact of these healthcare models on the access to, quality and cost of healthcare in selected European countries. Access is nearly 100% in countries with a public provider system, while in most of the 'mixed' countries, the difference from 100% is made up by supplementary private insurance. No differences are seen between public and mixed provider systems in terms of quality of care, despite the fact that the countries with the former model spend, in general, less of their Gross National Product on healthcare. The Private Insurance/private provider model of the US produces the highest costs, but is lowest in access and is close to lowest ranking in quality parameters. PMID- 10528707 TI - Healthcare systems and end-stage renal disease (ESRD) therapies--an international review: access to ESRD treatments. AB - Assessment of healthcare technology and economics can be used to assess the access to healthcare, its quality and efficacy as well as its cost and cost efficiency. This report addresses these issues for the provision of care for end stage renal disease (ESRD) patients. An international comparison of access to ESRD treatment modalities was made with reference to the healthcare provider structure in a range of industrial countries. The countries were grouped into 'public' (Beveridge model), 'mixed' (Bismarck model) and 'private' (Private Insurance model). In 'public' provider countries, 20-52% of dialysis patients are treated with home therapies (haemodialysis and peritoneal dialysis), and the number of patients with renal transplants is 45-81% of all ESRD patients. In 'mixed' provider countries, only 9 17% of all dialysis patients are treated with home therapies, and 20-48% of ESRD patients have renal transplants. In 'private' provider countries, 17% of US and 6% Japanese dialysis patients are treated with home therapies. Japan has 0.3% and the US has 26% of ESRD patients who receive renal transplants. It thus seems that provider structure influences access to and choice of ESRD treatment. With a growing elderly population and longer life expectancy, there will be an increased requirement for ESRD treatments in all industrial countries. Equal access to, and quality of ESRD care in the future will require adequate funding and reimbursement strategies in a cost-constrained healthcare environment. growing elderly population, new and innovative healthcare technologies, increasing expectations of the population and the dilemma of economic constraints. Therefore, new disciplines such as health technology assessment and healthcare economics are developing to support the needs of health policy decision makers. Their main objective is to create a balance between the three key factors of a healthcare system: access to healthcare (equity for all), quality of healthcare (efficacy) and finally the cost or cost efficiency of healthcare provision [1; see also Lameire et al., this issue]. This report will assess access to healthcare in a very specific and very costly area that of end stage renal disease (ESRD). An international comparison of access to ESRD treatment for patients from a series of industrial countries will be used as a means for evaluation of this access. PMID- 10528708 TI - The pattern of referral of patients with end-stage renal disease to the nephrologist--a European survey. AB - Much attention has been paid recently to an early or 'healthy' start of chronic dialysis in patients suffering from end-stage renal disease (ESRD). It is hoped that earlier initiation of dialysis will reduce the increased morbidity and mortality that is observed during the early months in patients who are referred too late to the nephrological unit. This report deals with a survey of the referral pattern of patients with ESRD in 14 European centres. Late referral was defined as those patients presenting to the renal unit within 1 month before chronic dialysis was needed. Between 1993 and 1995, 2236 ESRD patients started dialysis in these 14 centres. A total of 583 patients, representing 26%, were late referrals as defined above. A high variability between the individual centres, even within the same country, was observed. More detailed surveys in both Flemish and European centres revealed that late referrals more frequently started on haemodialysis, had significantly greater co-morbidity and mortality in the first year of dialysis and were transplanted less frequently. It can also be deduced from these results that late referral has important economic consequences, leading to increased costs because of longer initial hospitalization times, the lack of choice for the cheaper form of peritoneal dialysis the lower transplantation rates in the late referrals. PMID- 10528709 TI - Outcomes in peritoneal dialysis and haemodialysis--a comparative assessment of survival and quality of life. AB - Ever since the introduction of peritoneal dialysis (PD) as a therapy for managing patients with end-stage renal disease, there has been considerable debate about how it compares with outcomes on haemodialysis (HD) especially in terms of survival and quality of life. Whilst earlier results in the 1980s were certainly not comparable, data now emerging show that survival on PD is equivalent to that on HD. Recent registry data from the Canadian Organ Replacement Register show that survival of patients on PD is equivalent to that on HD and may well be better in the first few years of therapy. There have been numerous quality of life studies in patients on PD and HD. Health-related quality of life has been assessed using health profile measurements (both generic and disease-specific instruments) or preference-based measurements. The former approach has been used to analyse 14 different comparative studies. These studies suggest that patients on home HD and CAPD show better quality of life than patients on centre HD. Only a few studies found statistical differences between groups, and only in seven studies were results adjusted for patient differences. There is a need for longitudinal studies with more accurate information on health. Similar data are available for preference-based measurements and studies. Overall, the analysis suggests that PD and HD are equivalent therapies. On this basis, it is hard to explain the wide variation seen in the use of the two therapies. PMID- 10528710 TI - Healthcare systems and end-stage renal disease (ESRD) therapies--an international review: costs and reimbursement/funding of ESRD therapies. AB - BACKGROUND: In healthcare economics, the cost factor plays a leading role, particularly for chronic diseases such as end-stage renal disease because of the growing number of patients. OBJECTIVES: An international comparison was made of the costs and reimbursement/funding of a selection of key dialysis modalities- centre haemodialysis (CHD), limited care haemodialysis (LCHD), home haemodialysis (home HD), continuous ambulatory peritoneal dialysis (CAPD) and automated peritoneal dialysis (APD)--in various industrial countries. The focus was on treatment costs plus erythropoietin medication and reimbursement of transportation costs. RESULTS: Reimbursement/funding of dialysis is different from country to country, with some healthcare system-specific commonalities: in 'public' systems, the funding is based more on global budgets, whereas in mixed public and private countries it is based mainly on reimbursement rates per treatment. Only in the 'private system' of the US is there one DRG (diagnostic related group)-type rate for dialysis. By comparing the costs (in public countries) or reimbursements (in mixed countries) of treatment modalities within each country, we could see similar curves: the costs were the highest for public CHD, followed by private CHD. They were lower on LCHD and the lowest for home HD and CAPD, which were at nearly the same level. The cost level for APD was almost the same as that of LCHD. The reimbursements followed the cost pattern. Some countries introduced increases for CAPD and APD with the intention of increasing the share of home care. The costs and reimbursement patterns in the majority of countries (except the US and Japan) were very similar and therefore did not explain the different distribution of modalities in these countries. One explanation could be, however, the difference in microeconomics, CHD being a treatment with high fixed costs (personnel and structure) and CAPD being a treatment with low fixed costs, but high variable costs (supplies) and a low need for investments. DISCUSSION: The choice of treatment modality seems to be influenced strongly by the provider's perspective, being either public with limited HD capacity or private having invested in HD capacity. For public providers (and healthcare payers), CAPD is less expensive than CHD and offers a number of potential savings. In many countries, two CAPD patients could be treated for the same costs as one CHD patient. The microeconomics of private centres, however, are meant to use the investments maximally for CHD. Only if capacity limits are reached, is PD, with mainly supply costs, interesting. The future with constantly increasing numbers of patients and growing cost constraints will force all providers to make the best use of their resources by also offering home therapies such as PD to patients. The latter are cost efficient and offer comparable survival and quality of life. PMID- 10528711 TI - Development and use of polymerase chain reaction for the specific detection of Salmonella Typhimurium in stool and food samples. AB - Salmonella Typhimurium is one of the most important Salmonella serovars that may cause foodborne disease and human salmonellosis infection. Detection of this organism in the clinical samples of persons with gastroenteritis and the food samples associated with such persons may allow us to trace the cause of disease. Because malic acid dehydrogenase, an enzyme of the citric acid cycle, is common to organisms, the gene (mdh) coding for this enzyme was selected for the design of Salmonella Typhimurium-specific polymerase chain reaction (PCR) primers. By comparison of the mdh gene sequences of Salmonella Typhimurium and other Salmonella serotypes and of some isolates of other genera, two oligonucleotides were designed and used as PCR primers for the specific detection of Salmonella Typhimurium. The molecular weight of the PCR product was 261 bp as expected. Salmonella serovars other than Salmonella Typhimurium and isolates of other genera in the Enterobacteriaceae that is closely related to Salmonella did not generate any false-positive results. When this primer pair was used for the detection of Salmonella Typhimurium cells artificially inoculated into human stool specimens and food samples, such as milk and raw chicken meat, levels as low as 10(0) CFU per 0.1 g of stool specimen or per ml of milk or food homogenate could be detected if an 8- to 12-h preculture step using combined lactose tetrathionate broth was performed prior to the PCR. PMID- 10528712 TI - Response surface models for effects of temperature and previous temperature on lag time and specific growth rate of Salmonella Typhimurium on cooked ground chicken breast. AB - Response surface models were developed for effects of temperature (16 to 34 degrees C) and previous temperature (pretemperature; 16 to 34 degrees C) on lag time (lambda) and specific growth rate (mu) of Salmonella Typhimurium on cooked ground chicken breast. The primary objective was to determine whether pretemperature is a major factor affecting growth of Salmonella Typhimurium. Growth curves for model development (n = 32) and model testing (n = 18) were fit to a two-phase linear equation that directly estimated lambda and mu. Response surface models for ln lambda and ln mu as a function of temperature and pretemperature were obtained by regression analysis. Lag time and mu of Salmonella Typhimurium were affected by temperature but not pretemperature. Models were tested against data not used in their development. Prediction error (model accuracy) was 13.4% for lambda and 11.3% for mu, whereas the median relative error of predictions (model bias) was -3.0% for lambda and 6.8% for mu. Results indicated that the models provide reliable predictions of lambda and mu of Salmonella Typhimurium on cooked ground chicken breast within the matrix of conditions modeled. In addition, pretemperature (16 to 34 degrees C) is not a major factor affecting growth of Salmonella Typhimurium. PMID- 10528713 TI - Occurrence and survival of verocytotoxin-producing Escherichia coli O157 in meats obtained from retail outlets in The Netherlands. AB - In 1996 and 1997, 2,941 fresh and processed meat products obtained from supermarkets and butcher shops in The Netherlands were examined for the presence of verocytotoxin-producing Escherichia coli of serogroup O157 (O157 VTEC). Additionally, the fate of O157 VTEC in raw meat products stored at low temperatures and the effect of different additives were evaluated. O157 VTEC strains were isolated from 6 (1.1%) of 571 samples of raw minced beef, 2 (0.5%) of 402 samples of raw minced mixed beef and pork, 1 (1.3%) of 76 samples of raw minced pork, 1 (0.3%) of 393 samples of other raw pork products, and 1 (0.3%) of 328 samples of cooked or fermented ready-to-eat meats. Other raw beef products (n = 223) and meat samples originating from poultry (n = 819), sheep or lamb (n = 46), or wild animals (n = 83) were all found to be negative for O157 VTEC. For the survival experiments we used tartaar (minced beef with a fat content of less than 10%) and filet americain (tartaar mixed with a mayonnaise-based sauce [80 to 20%]). The O157 VTEC strain tested was able to survive in tartaar and filet americain stored at -20, 0, 5, or 7 degrees C for 3 days. At both 7 and at 15 degrees C, O157 VTEC counts in tartaar and filet americain remained virtually unchanged throughout a storage period of 5 days. Addition of acetic acid (to pH 4.0), sodium lactate (1 and 2% [wt/wt]), or components of the lactoperoxidase thiocyanate-hydrogen peroxide system to filet americain did not result in a reduction of viable O157 VTEC cells during storage at 7 or 15 degrees C. It was concluded that raw meat contaminated with O157 VTEC will remain a hazard even if the meat is held at low or freezing temperatures. PMID- 10528715 TI - Behavior of Listeria monocytogenes and Aeromonas spp. on fresh-cut produce packaged under equilibrium-modified atmosphere. AB - Storage experiments were conducted to follow the behavior of pathogens on fresh cut vegetables (trimmed brussels sprouts, grated carrots, shredded iceberg lettuce, and shredded chicory endives) packaged under an equilibrium-modified atmosphere (EMA) (2 to 3% O2, 2 to 3% CO2, and 94 to 96% N2) and stored at 7 degrees C. As a comparison, fresh-cut vegetables were also packaged in a perforated high-barrier film (air conditions) and stored at 7 degrees C. In a first step, the shelf life of the vegetables in the two kinds of packages was determined by evaluating the microbiological quality as well as the sensorial quality (appearance, taste, and odor). In general, sensorial properties were faster in limiting the shelf life than microbiological criteria. The shelf life of the vegetables stored under an EMA was extended by 50% or more, compared with the air-stored vegetables. In a second storage experiment, the four fresh-cut vegetables were inoculated with a cocktail of psychrotrophic pathogens (Listeria monocytogenes, Aeromonas caviae [HG4]) and A. bestiarum (HG2) before packaging under an EMA and air at 7 degrees C. The inoculated pathogens were more influenced by the type of vegetable than by the type of atmosphere. No growth was detected on the brussels sprouts or on carrots (L. monocytogenes). Aeromonas spp. had a higher growth rate than L. monocytogenes on the shredded chicory endives and shredded iceberg lettuce at 7 degrees C. PMID- 10528714 TI - Reduction of Listeria monocytogenes and Escherichia coli O157:H7 numbers on vacuum-packaged fresh beef treated with nisin or nisin combined with EDTA. AB - Nisin or nisin combined with EDTA was used to treat fresh beef. Beef cubes (2.5 by 2.5 by 2.5 cm) that were inoculated with approximately 7 log CFU/ml of Listeria monocytogenes Scott A or Escherichia coli O157:H7 505 B were dipped in the following solutions: (i) H2O, (ii) HCl, (iii) nisin, (iv) EDTA, or (v) nisin combined with EDTA, respectively, for 10 min each, with an exception of one set of control beef samples without treatment. Beef samples were then drip-dried for 15 min, vacuum packaged, and stored at 4 degrees C for up to 30 days. The pH on beef after different treatments was not a key factor in preventing bacterial growth. Treatment with nisin or with nisin combined with EDTA reduced the population of L. monocytogenes by 2.01 and 0.99 log CFU/cm2 as compared to the control, respectively, under the conditions of vacuum package and storage at 4 degrees C for up to 30 days. However, the effect of nisin and nisin combined with EDTA against E. coli O157:H7 505 B was marginal at 1.02 log CFU/cm2 and 0.8 log CFU/cm2 reductions, respectively. PMID- 10528716 TI - Irradiation and modified atmosphere packaging for the control of Listeria monocytogenes on turkey meat. AB - When radiation-sterilized ground turkey meat was inoculated with Listeria monocytogenes, packaged under mixtures of nitrogen and carbon dioxide, and irradiated with gamma-radiation doses of 0 to 3.0 kGy, there was a statistically significant (P < 0.05), but probably not a biologically significant, lower (0.39 log) predicted bacterial survival in the presence of 100% carbon dioxide than in the presence of 100% nitrogen. Possibly because all atmospheres contained oxygen and because a response surface design was used, gamma-radiation resistance was not significantly (P < 0.05) different in air than in modified atmosphere packaging (MAP) mixtures containing 5% O2 or containing 20, 40, 60, and 80% CO2 and balance N2. The antilisterial effects of MAP mixtures containing 17.2, 40.5, and 64% CO2 and balance N2 were compared to those associated with air and vacuum packaging on turkey inoculated with approximately 5 x 10(3) CFU/g. Samples were irradiated to doses of 0, 0.5, 1.0, 1.5, 2.0, and 2.5 kGy and were stored at 7 degrees C for up to 28 days. Irradiation treatments were significantly more lethal in the presence of air packaging than in either vacuum packaging or MAP, and in those samples that received >1.0 kGy, there was a concentration-dependent CO2 inhibition of L. monocytogenes multiplication and/or recovery. PMID- 10528717 TI - A predictive model to determine the effects of pH, milkfat, and temperature on thermal inactivation of Listeria monocytogenes. AB - Listeria monocytogenes is a foodborne pathogen of significance because of its comparatively high heat resistance, zero tolerance in ready-to-eat foods, and growth at refrigeration temperatures. A 3 x 3 x 3 factorial study was done to determine the effects of milkfat (0%, 2.5%, 5.0%), pH (5.0, 6.0, 7.0), and processing temperature (55 degrees C, 60 degrees C, 65 degrees C) on the thermal resistance of L. monocytogenes in a formulated and homogenized milk system. Data were fit to a modified Gompertz equation where parameter estimates characterized three regions of a survival curve: the shoulder, maximum slope, and tail. Statistical analysis was done for each of the 27 individual treatment sets to visualize individual effects on parameter estimates and to evaluate how well the Gompertz equation represented the data. A regression model for the Gompertz equation was generated to predict the logarithmic surviving fraction of L. monocytogenes based on all 27 treatments and their single and interactive effects. The shoulder region of the survival curve was affected by pH; however, the maximum slope was affected by temperature, milkfat, and the interaction of temperature x milkfat. Validation of the model suggests that the predictions are best suited for processing above 62 degrees C. Trends over time for a 4-log reduction in cells (4D values) were evaluated using results from the 27 individual treatment sets, the regression model for the Gompertz equation, and a linear equation. At lower temperatures, 4D values by the three methods varied by twofold. At higher temperatures, all methods gave similar 4D values, suggesting that death became more linear. Based on this study all three factors affect heat resistance for specific regions of a survival curve, and a predictive model was developed that can be used as a preliminary estimate for L. monocytogenes inactivation. PMID- 10528718 TI - A 23S rDNA-targeted polymerase chain reaction-based system for detection of Staphylococcus aureus in meat starter cultures and dairy products. AB - A polymerase chain reaction-based system for detection of Staphylococcus aureus was developed. The system consisted of the following components: (i) selective enrichment, (ii) DNA isolation, (iii) amplification of DNA with primers targeted against the 23S rRNA gene, and (iv) evaluation of the specificity of the polymerase chain reaction by Southern hybridization and nested polymerase chain reaction. The method achieved a high degree of sensitivity and unambiguity as required for the detection of contaminants in food starter preparations. The method permitted detection of Staphylococcus aureus in preparations of meat starter cultures containing Staphylococcus carnosus either alone or in combination with lactobacilli, pediococci, and/or Kocuria varians. Detection limits were sufficiently low to show within 12 h the presence of 10(0) CFU of S. aureus in starter preparations containing 10(10) CFU of S. carnosus. The system was also applied to dried skim milk and cream. For detection without selective enrichment, a protocol was developed and permitted detection of 120 CFU of S. aureus in 1 ml of cream within 6 h. With nested polymerase chain reaction, the detection limit was decreased by one order of magnitude. PMID- 10528719 TI - Effect of pH and CO2 on growth and toxin production by Clostridium botulinum in English-style crumpets packaged under modified atmospheres. AB - The effect of pH and CO2 on both growth of and toxin production by Clostridium botulinum in English-style crumpets, packaged under modified atmospheres was investigated using a 2 x 2 factorial experiment. English-style crumpets (water activity, 0.990; pH 6.5 and 8.3) were inoculated with C. botulinum spores types A and proteolytic B (500 spores/g), packaged in either 60% CO2 (balance N2) or 100% CO2, stored at ambient temperature (25 degrees C), and monitored daily for toxicity. Toxin was detected after 4 days in crumpets packaged in 60% CO2, irrespective of initial product pH. Toxin production was delayed 1.5 to 3 days in crumpets packaged under 100% CO2. Analysis of variance indicated a significant interaction effect of pH and %CO2 on time of earliest toxin detection. Delay of toxin production was greatest for high pH (8.3) crumpets. All products were organoleptically acceptable at the time of toxigenesis, and therefore, high moisture-high pH bakery products, if contaminated with spores of C. botulinum, could become hazardous if packaged in atmospheres containing CO2. PMID- 10528720 TI - The improvement of color and shelf life of ham by gamma irradiation. AB - This study was undertaken to determine if gamma irradiation can circumvent the need for sodium nitrite to obtain and maintain the desired color of pork loin ham. A dose of 5 kGy was observed to be as effective as the use of 200 ppm of sodium nitrite to provide and maintain the desired color of the product for 30 days. Peroxidation of the product was reduced with addition of sodium nitrite but increased with irradiation. However, organoleptic quality of the irradiated ham without added sodium nitrite was acceptable. PMID- 10528721 TI - Chlorophyll pigment composition in table olives (cv. Gordal) with green staining alteration. AB - Metallochlorophyllic complexes of copper are present in green table olives, showing the alteration known as green staining. They were formed stepwise in such a way that new metallochlorophyll derivatives were detected as the fruits became more altered. Cu-15-glyoxylic acid pheophytin a was the first compound formed, followed in order by Cu-pheophytin a, Cu-15-glyoxylic acid pheophytin b, Cu-15 formyl-pheophytin a, and Cu-pyropheophytin a. Pigment analysis in fruits classified according to the surface area affected by green staining showed a progressive increase in the concentration of all copper complexes with the course of the alteration. The metallochlorophyll derivatives of copper were present both in the part of the fruit affected by the alteration and in the remainder part, although the amount was significantly greater in the former. On the other hand, not all the copper present in the fruits was accessible for the formation of such compounds, since the concentration of copper complexes found in olives with the maximum degree of alteration observed did not exceed 20% of the total copper of the fruit. PMID- 10528722 TI - Identification of oxidized chlorophylls and metallochlorophyllic complexes of copper in table olives (cv. Gordal) with green staining alteration. AB - In Spanish green table olives showing the alteration known as green staining, new chlorophyll derivatives have been identified: the copper complexes of 3(1),3(2) didehydrorhodochlorin-15-glyoxylic acid-17(3)-phytyl ester a and b (Cu-15 glyoxylic acid pheophytins a and b), and 3(1),3(2)-didehydro-15(1)-hydroxy-15(1) hydroxyrhodochlorin- 15-acetic acid delta-lactone-15(2)-methyl-17(3)-phytyl ester a and b (Cu-15(1)-OH-lactone-pheophytins a and b). These compounds were isolated by normal phase thin-layer chromatography and identified from their UV-visible and mass spectra and by co-chromatography with authentic standards. The chromatographic and spectroscopic characteristics and the molecular mass for a new allomerized chlorophyll derivative and its copper complex are reported. The corresponding molecular structure according to the molecular mass has been proposed. The characterization of the latter compounds enables their possible detection in processed fruits and vegetables. The present work gathers new advances in the study of the color alteration in table olives and this is the first time that copper complexes of oxidized chlorophylls are detected in foodstuffs. PMID- 10528723 TI - Indirect enzyme-linked immunosorbent assay for the identification of sole (Solea solea), European plaice (Pleuronectes platessa), flounder (Platichthys flesus), and Greenland halibut (Reinhardtius hippoglossoides). AB - Polyclonal antibodies produced against soluble muscle protein extracts from sole (Solea solea), European plaice (Pleuronectes platessa), flounder (Platichthys flesus), and Greenland halibut (Reinhardtius hippoglossoides) were used in an indirect enzyme-linked immunosorbent assay for the specific identification of fillets from these flatfish species. The assay was performed in two different formats: microtiter plates and immunostick tubes. Immunorecognition of antibodies adsorbed to their specific fish samples was made with goat antirabbit immunoglobulins conjugated to the enzyme horseradish peroxidase. Subsequent enzymatic conversion of the substrate allowed unequivocal identification of all flatfish species studied. PMID- 10528724 TI - Evaluation of the Delvo-X-Press assay for detecting antibiotic residues in milk samples from individual cows. AB - Performance of the Delvo-X-Press beta-lactam antibiotic assay was examined using bulk-tank milk samples and milk samples from individual cows. Bulk-tank milk samples fortified with bovine lactoferrin at a concentration of 1 mg/ml or more consistently tested positive. False-positive results were also obtained from bulk tank milk samples fortified with bovine plasma at concentrations of 20 and 40%. The assay yielded positive results for milk with antibiotic concentrations as low as 2 ppb. Individual milk samples were collected from 144 healthy lactating cows and from 34 cows with chronic Staphylococcus aureus mastitis. Specificity estimates for samples from healthy and mastitic cows were 0.88 (95% confidence interval [CI], 0.82, 0.93) and 0.94 (95% CI, 0.86, 1.00), respectively. Individual milk samples were collected from three cows with experimentally induced mastitis for 21 consecutive days. False-positive results occurred as late as 12 days postchallenge. A moderate but significant (P < 0.01) positive linear correlation (r = 0.61) was observed between test result and somatic cell count (SCC) values in milk samples with SCCs of >10(6)/ml. PMID- 10528725 TI - Development of a polymerase chain reaction-based assay for the detection of Alternaria fungal contamination in food products. AB - Alternaria sp. are important fungal contaminants of vegetable, fruit, and grain products, including Alternaria alternata, a contaminant of tomato products. To date, the Howard method, based on microscopic observation of fungal filaments, has been the standard examination for inspection of tomato products. We report development of a polymerase chain reaction (PCR)-based method for detection of Alternaria DNA. PCR primers were designed to anneal to the internal transcribed regions ITS1 and ITS2 of the 5.8S rRNA gene of Alternaria but not to other microbial or tomato DNA. We demonstrate use of the PCR assay to detect Alternaria DNA in experimentally infested and commercially obtained tomato sauce and tomato powder. Use of the PCR method offers a rapid and sensitive assay for the presence of Alternaria DNA in tomato products. The apparent breakdown of DNA in tomato sauce may limit the utility of the assay to freshly prepared products. The assay for tomato powder is not affected by storage time. PMID- 10528726 TI - Production of cyclopiazonic acid by molds isolated from Taleggio cheese. AB - Twenty-seven strains of Penicillium were isolated from the rind of Taleggio, a typical Italian cheese, so that we could test their capacity to produce cyclopiazonic acid (CPA); all strains produced CPA. The production was strongly influenced by the strain variety and growth conditions. Strains incubated at 25 degrees C for 7 days always produced CPA in mannitol broth, with concentrations ranging from 0.02 to 1 microg/ml, whereas only 33% of strains grown in yeast extract broth produced CPA, with a maximum value of 0.1 microg/ml. In milk, maximum production (1.6 microg/ml) was observed after 14 days of incubation at 25 degrees C. In order to evaluate the presence of the toxin and its capacity for migrating into the cheeses, the rind, the cheese near the rind, and the cores from six Taleggio cheeses were analyzed. CPA was present in five cheeses, with a maximum concentration of 0.25 mg/kg in one rind, and in one cheese, the toxin migrated to the core. A positive correlation between CPA production and surface mold was found. PMID- 10528727 TI - Repair and recovery of thermally injured cells of Yersinia enterocolitica in milk. AB - One standard strain of the organism MTCC-861 and the two culture isolates VRW-22 and CRW-15 were exposed to batch pasteurization (62.8 degrees C for 30 min) in brain heart infusion broth, skim milk, and whole milk. The trials were repeated three times. None of the isolates survived pasteurization treatment. However, the studies were further directed toward the repair and recovery of thermally injured cells, if any, in peptone sorbitol bile broth, skim milk, and whole milk. The results revealed that only a low number of test cultures were recovered in peptone sorbitol bile broth after 8 days of incubation at 10 degrees C. On the other hand, the recovery was still slower in skim and whole milk, with a detection of the test isolates only on 10 days of incubation under similar conditions. PMID- 10528728 TI - Effects of iron and selenium on the production of catalase, superoxide dismutase, and listeriolysin O in Listeria monocytogenes. AB - Listeria monocytogenes 19112, Scott A, and 10403S were grown in tryptic soy broth (TSB) and TSB supplemented with 25 to 100 microg/ml of iron (Fe) and 0.5 to 2.5 microg/ml selenium (Se) to examine the effects on catalase (CA), superoxide dismutase (SOD), and listeriolysin O (LLO) activities. Growth in TSB supplemented with Fe resulted in significant increases in CA, SOD, and LLO activities in all three strains when compared to growth in TSB. The addition of 0.5 microg/ml Se to TSB resulted in significantly higher CA and LLO activities in L. monocytogenes 19112 but showed no effect on Scott A or 10403S. These results suggest that Fe plays a role in increasing the activities of CA, SOD, and LLO. PMID- 10528729 TI - A multiplex reverse transcription polymerase chain reaction method for the detection of foodborne viruses. AB - A multiplex reverse transcription polymerase chain reaction (RT-PCR) method was developed for the simultaneous detection of the human enteroviruses, hepatitis A virus (HAV) and Norwalk virus (NV). Poliovirus type 1 (PV1) was chosen as a model for the human enterovirus group. Three different sets of primers were used to produce three size-specific amplicons of 435 bp, 270 bp, and 192 bp for PV1, NV, and HAV, respectively. RT-PCR products were separated by agarose gel electrophoresis, and amplicon identity was confirmed by Southern transfer followed by DNA hybridization using nonradioactive, digoxigenin-labeled internal probes. When tested on mixed, purified virus suspensions, the multiplex method achieved detection limits of < or = 1 infectious unit (PV1 and HAV) or RT-PCR amplifiable unit (NV) for all viruses. With further streamlining efforts such as single tube amplification and liquid hybridization, multiplex PCR offers advantages over cell culture methodology and monoplex PCR because it allows for rapid and cost-effective detection of several human enteric viruses in a single reaction tube. PMID- 10528730 TI - Saccharomyces cerevisiae thermal inactivation kinetics combined with ultrasound. AB - Inactivation kinetics of Saccharomyces cerevisiae during thermal treatments at moderate temperatures (45.0, 47.5, 50.0, 52.5, or 55.0 degrees C) combined with application of 20 kHz of ultrasound were evaluated. S. cerevisiae inactivation under the combined effects of heat and ultrasound followed first-order reaction kinetics, with decimal reduction times (D) that varied from 22.3 to 0.8 min. D values in treatments that combined heat and ultrasound were significantly smaller (P < 0.05) than D values obtained for thermal treatments and were more noticeable at temperatures below 50 degrees C. The dependence of the D value on temperature had a significantly (P < 0.05) greater z value for combined treatments. Yeast heat inactivation kinetics revealed decreased thermal resistance caused by ultrasound. PMID- 10528731 TI - Fumonisin B1 and hydrolyzed fumonisin B1 (AP1) in tortillas and nixtamalized corn (Zea mays L.) from two different geographic locations in Guatemala. AB - Fumonisin B1 (FB1) is a common contaminant of corn worldwide and is responsible for several diseases of animals. In the preparation of tortillas, corn is treated with lime (producing nixtamal) that when heated hydrolyzes at least a portion of the FB1 to the aminopentol backbone (AP1), another known toxin. This study analyzed the amounts of FB1 and AP1 in tortillas and nixtamal from two communities in the central highlands of Guatemala where corn is a major dietary staple (Santa Maria de Jesus, Sacatepequez, and Patzicia, Chimaltenango). The amounts of FB1 and AP1 in tortillas from Santa Maria de Jesus were, respectively, 0.85 +/- 2.0 and 26.1 +/- 38.5 microg/g dry weight (mean +/- SD), and from Patzicia were 2.2 +/- 3.6 and 5.7 +/- 9.4 microg/g dry weight. Less than 6% of the tortillas from both locations contained > or = 10 microg FB1/g dry weight; whereas, 66% of the samples from Santa Maria de Jesus and 29% from Patzicia contained > or = 10 microg AP1/g dry weight. The highest amount of AP1 (185 microg/g dry weight) was found in tortillas from Santa Maria de Jesus. The highest amounts of FB1 were 6.5 and 11.6 microg/g dry weight in tortillas from Santa Maria de Jesus and Patzicia, respectively. The mean concentration of FB1 in nixtamal was significantly higher in Santa Maria de Jesus compared to Patzicia. Surprisingly, AP1 was not detected in any of the nixtamal samples. The human impact of exposure to these amounts of fumonisins is not known. However, based on findings with other animals, where corn is a dietary staple, long-term consumption of FB1 and AP1 (especially at > or = 10 microg/g of the diet) may pose a risk to human health. PMID- 10528732 TI - Possible synergistic effect of nisin and propionic acid on the growth of the mycotoxigenic fungi Aspergillus parasiticus, Aspergillus ochraceus, and Fusarium moniliforme. AB - The effects of nisin and propionic acid (PA) on aflatoxin production and on mycelial growth and spore germination of the mycotoxigenic fungi Aspergillus parasiticus, A. ochraceus, and Fusarium moniliforme were investigated. The growth of A. ochraceus was completely inhibited on media containing PA with nisin in concentrations of 0.05% PA with 1,000 ppm nisin, and 0.1% PA with 500 or 1,000 ppm nisin. The growth of both F. moniliforme and A. parasiticus was completely inhibited by PA with nisin at a concentration of 0.1% PA with 1,000 ppm nisin. Nisin alone caused a significant increase in mycelial growth when applied to A. ochraceus at 500 or 1,000 ppm and when applied to A. parasiticus at 1,000 ppm. Spore germination of A. ochraceus was completely inhibited on media containing 0.1% PA with 500 or 1,000 ppm nisin. Spores of F. moniliforme failed to germinate in 0.05% PA with 500 or 1,000 ppm nisin, whereas spores of A. parasiticus did not germinate on media containing 0.1% PA with 1,000 ppm nisin. For all three fungi tested, the inhibitory effect on mycelial growth was found to be fungistatic rather than fungicidal. The combined treatment of PA with nisin produced better fungistatic activity than treatment involving either material alone. Nisin, applied alone, did not stimulate aflatoxin production (expressed by microg toxin/mg mycelium), but the combined treatment at certain concentrations was inhibitory to aflatoxin B1 or G1. The production of aflatoxin G1, but not of B1, was stimulated in 0.05% PA with 1,000 ppm nisin and on media containing 0.1% PA with 100 ppm nisin. Nisin is currently applied in foods to prevent spoilage induced by bacteria but not by mold. The results of the present study indicate that a combined treatment of nisin in small concentrations of PA might be useful in preventing mold damage in certain foods and stored grain. PMID- 10528733 TI - Program assessment and distance education in nursing. PMID- 10528734 TI - Competency model development. PMID- 10528735 TI - Creating academic/service partnerships through nursing competency models. PMID- 10528736 TI - Development of standards for differentiated competencies of the nursing workforce at time of entry/advanced beginner. PMID- 10528737 TI - A model for differentiated entry-level nursing practice by educational program type. PMID- 10528738 TI - The shifting sands of health care delivery: curriculum revision and integration of community health nursing. AB - The health care delivery system in the United States is moving from an institutionally driven and controlled medical care model toward the Primary Health Care model described by the Alma-Alta Conference (World Health Organization [WHO], 1978) as community-driven comprehensive health care. However, nursing education still remains institutionally based, anchored in a medical model. Dynamic curricula must be developed that prepare nurses to practice in an ever-changing health care delivery system that is becoming more community based. The purpose of this article is twofold: to provide an overview of the revised curriculum of Brigham Young University's College of Nursing (BYU-CON) as one example of a faculty's attempt to develop a program that prepares graduates skilled in providing health care in the 21st century; and to examine closely the integration of community health nursing into the curriculum. PMID- 10528739 TI - Steering an academic department through a paradigm shift: the case of a new paradigm for nursing. PMID- 10528740 TI - A nontraditional curriculum for the preparation of nurse educators. AB - The purpose of this article is to describe a bold and innovative program for the preparation of nurse educators in a developing country. This program was designed to anticipate the competing demands for radical change in both the health care system and the nursing educational system. Consideration was given to the need for a curriculum model that was flexible and broad enough to accommodate the educational and professional needs of a very diverse group of students regarding academic ability, culture, and language. The aim of the curriculum was to achieve the desire for meaningful, lifelong learning and personal growth, while maintaining academic excellence within a transformative and democratizing context. PMID- 10528741 TI - Peer review of teaching: instituting a program in a college of nursing. AB - Institutions of higher education can lose sight of the importance of teaching. Teaching, in some settings, needs to be restored to its place of primacy in the faculty role and its quality needs to be evaluated. The American Association for Higher Education (AAHE) national project, "From Idea to Prototype: The Peer Review of Teaching," is an attempt to accomplish that purpose. Involvement in the AAHE national peer review project has been the impetus for the Kent State University College of Nursing to initiate a faculty-developed program of peer review. The program has been well received primarily because it was owned and controlled by the faculty. Its voluntary nature, administrative sanction, and strong support have contributed to its success. Faculty explored many possible methods to accomplish peer review. However, the use of a task force, classroom and clinical observations, and teaching circles have been the approaches best suited to this particular departmental culture. PMID- 10528742 TI - A curriculum self-study of writing assignments and reflective practice in nursing education. AB - Reflective practice is an ongoing process of purposeful thinking about one's clinical practice to develop understanding, insight, and clinical judgment. It can be enhanced through careful use of writing assignments that require reflection, evaluation, and thoughtful analysis. The uses of writing assignments in the undergraduate nursing programs at a midwestern college of nursing were studied over a 2-year period, using survey methodology. A purposive sample of all faculty who taught in baccalaureate or associate of science (ASN) programs (n=21) completed a questionnaire and were interviewed about the number and purpose of writing assignments in courses they taught. Quantification of the results in tabular form allowed the faculty to look at all writing assignments required of students across both programs with regard to the number, length, and nature of those assignments. Results were then recategorized by type of assignments, changes needed, and faculty assessment of the usefulness of these writing assignments. The summary of written assignments was then cross-referenced according to program, semester, and program level. This produced a working document that illustrated the quantity and type of writing assignments that each student, in each program, at a given level and semester must complete. Information was used by faculty teaching across courses to make changes that more effectively linked course writing assignments by association, themes, concepts, or areas of study. This process enables faculty to collaboratively develop writing assignments that facilitate the adult student's linking and associating concepts across courses, as a true exercise in critical thinking. PMID- 10528743 TI - Inducible nitric oxide synthase promotes cytokine expression in cardiac allografts but is not required for efficient rejection. AB - BACKGROUND: Inducible nitric oxide synthase (iNOS) is enhanced during acute rejection. Pharmacologic inhibition of nitric oxide synthase (NOS) activity has had variable effects on graft survival in a number of animal models. To further characterize the requirement and effects of iNOS during acute allograft rejection, we examined rejection responses of mice completely deficient of iNOS. METHODS: Heterotopic cardiac allografts were performed using wild-type and iNOS deficient mice (iNOS[-/-]) as recipients. Graft survival was determined by abdominal palpation. At days 3 and 7 following transplantation, grafts were harvested and analyzed histologically. Cytokine messenger RNA (mRNA) expression was measured by ribonuclease protection assay. RESULTS: Mean survival time of cardiac allografts did not differ between wild-type (18 +/- 3 days) and iNOS(-/-) recipients (16 +/- 2 days). At 3 days, findings of moderate acute rejection were seen in both recipients groups, although modestly reduced in iNOS(-/ -) mice. By 7 days, allografts in both groups demonstrated severe rejection. Within grafts at day 3, there was a 3-fold reduction in IL-1beta expression and a 4-fold reduction in IL-1RA in iNOS(-/-) recipients (p = 0.03 andp = 0.04, respectively) compared to wild-type recipients. Expression of other proinflammatory cytokines was detected in the grafts from both recipients, but was not significantly different. Finally, rejection responses to iNOS(-/-) cardiac allografts were nearly identical to wild-type allografts. CONCLUSIONS: Rejection of cardiac allografts by iNOS(-/-) mice occurs in a similar fashion to wild-type recipients, with extensive inflammation and proinflammatory cytokine production. While iNOS may play a role in cytokine induction by macrophages, these studies suggest that iNOS is not required for efficient cardiac graft rejection. PMID- 10528744 TI - Transforming growth factor beta (TGF-beta) and obliterative bronchiolitis following pulmonary transplantation. AB - BACKGROUND: Obliterative bronchiolitis (OB) characterised by small-airway fibrosis is a major cause of morbidity and mortality after lung transplantation. TGF-beta has been implicated in the pathogenesis of fibrosis. METHODS: We immunohistochemically examined 380 transbronchial biopsies (from 91 pulmonary transplants) using TGF-beta polyclonal antibodies. OB and interstitial fibrosis were diagnosed and graded in all biopsies. Other potential histologic and clinical risk factors for OB were analysed. RESULTS: Procedures were heart and lung (n = 32), bilateral sequential lung (n = 18), and single lung transplantation (n = 41). The incidence of OB in this group was 28.5%. In all patients with OB, TGF-beta was immunolocalized in the airways and lung parenchyma. TGF-beta expression was greater in OB patients (median score 8, range 5-12) in comparison to patients without OB (median score 4, range 1-13), p < .0001. Positive TGF-beta staining preceded the histologic confirmation of OB by 6 to 18 months. The development of OB was associated with two HLA mismatches at the A locus (p = .02); recurrent acute rejection episodes (p < .0005); lymphocytic bronchiolitis (p = .0001); and tissue eosinophilia, regardless of the rejection grade (p < .0001). CONCLUSIONS: Increased expression of TGF-beta is a risk factor for the development of OB. Other risk factors are recurrent acute rejection, lymphocytic bronchiolitis, tissue eosinophilia, and two mismatches at the HLA-A locus. This suggests that the pathogenesis of progressive small airway fibrosis characteristic of OB may be inflammatory damage, followed by an aberrant repair process due to excessive TGF-beta production following allograft injury. Hence, modulation of TGF-beta levels or function by antagonists may represent an important approach to control OB. PMID- 10528745 TI - Functional significance of cardiac reinnervation in heart transplant recipients. AB - BACKGROUND: There is accumulating evidence of structural sympathetic reinnervation after human cardiac transplantation. However, the functional significance of reinnervation in terms of exercise capacity has not been established as yet; we therefore investigated the influence of reinnervation on cardiopulmonary exercise testing. METHODS: After orthotopic heart transplantation 35 patients (mean age, 49.1 +/- 8.4 years) underwent positron emission tomography with scintigraphically measured uptake of C11-hydroxyephedrine (HED), lung function testing, and cardiopulmonary exercise testing. Two groups were defined based on scintigraphic findings, indicating a denervated group (n = 15) with a HED uptake of 5.45%/min and a reinnervated group (n = 20) with a HED uptake of 10.59%/min. RESULTS: The two study groups did not show significant differences with regard to anthropometric data, number of rejection episodes, preoperative hemodynamics, and postoperative lung function data. The reinnervated group had a significant longer time interval from transplantation (1625 +/- 1069 versus 800 +/- 1316 days, p < .05). In transplant recipients with reinnervation, heart rate at maximum exercise (137 +/- 15 versus 120 +/- 20 beats/min, p = .012), peak oxygen uptake (21.0 +/- 4 versus 16.1 +/- 5 mL/min/kg, p = .006), peak oxygen pulse (12.4 +/- 2.9 versus 10.2 +/- 2.7 mL/min/beat, p = .031), and anaerobic threshold (11.2 +/- 1.8 versus 9.5 +/- 2.1 mL/min, p = .046) were significantly increased in comparison to denervated transplant recipients. Additionally, a decreased functional dead space ventilation (0.24 +/- 0.05 versus 0.30 +/- 0.05, p = .004) was observed in the reinnervated group. CONCLUSIONS: Our study results support the hypothesis that partial sympathetic reinnervation after cardiac transplantation is of functional significance. Sympathetic reinnervation enables an increased peak oxygen uptake. This is most probably due to partial restoration of the chronotropic and inotropic competence of the heart as well as an improved oxygen delivery to the exercising muscles and a reduced ventilation-perfusion mismatching. PMID- 10528746 TI - Return to work after heart transplantation: 12-year follow-up. AB - BACKGROUND: Current statistics show that 50% of heart transplantation recipients survive 10 years or more. Emphasis on cost containment has renewed interest in the employment status of these long-term survivors. METHODS: We have identified 62 patients operated on at one United Kingdom center and evaluated their employment and clinical status by interview at 1, 5, and 12 years from surgery. RESULTS: Employment pattern was good, with 69%, 69% (of 55 survivors), and 57% (of 35 survivors) of the group working at 1, 5, and 12 years, respectively. Blue collar work status per se was not a deterrent to employment, but older patients with a poor presurgical work history were less successful in finding work, and trade and legislative restrictions forced some, who were willing and capable of returning to their previous employment, into lower paid, less satisfactory jobs. CONCLUSIONS: Subgroups of patients who are likely to be unemployed can be identified prior to surgery. Steps can be taken prior to surgery to anticipate future problems, and establish realistic employment goals. PMID- 10528747 TI - Long-term heart preservation using a new portable hypothermic perfusion apparatus. AB - OBJECTIVE: Perfusion storage is not often used clinically compared with simple immersion because of complicated circuits and demanding management. We developed a new apparatus for preservation combined with simple immersion and continuous coronary perfusion. METHODS: The main characteristics of this apparatus are as follows: (1) hypothermic storage, (2) does not require any energy source, (3) variable perfusion pressure, and (4) portability. The perfusion apparatus is composed of a storage chamber, a cooling chamber, and metal bars from which a perfusate bag is suspended. Adult mongrel dogs were divided into two groups: the coronary perfusion group (CP, n = 6) and the simple immersion group (SI, n = 6). Coronary vascular beds of the dog were washed out with a University of Wisconsin (UW) solution following cardiac arrest obtained using a GIK solution. The hearts were then excised. In the CP group, the heart graft, which was immersed in a 4 degrees C UW solution, was perfused with the same solution at a flow rate of 35 approximately 50 ml/hr. In the SI group, the heart graft was immersed in a 4 degrees C UW solution only. The heart graft was preserved for 12 hours in both groups. Beta-adenosine triphosphate (beta-ATP), phosphocreatine (Pcr), and inorganic phosphate (Pi) levels were measured immediately after excision of the heart, and at 3, 6, and 12 hours after preservation. Beta-ATP, Pcr, and Pi values were expressed as a percentage of control values, which had been obtained immediately after excision of the heart. Water content of the myocardium was measured prior to and after 12-hour preservation. The preserved graft was then evaluated through orthotopic transplantation. RESULTS: Beta-ATP/Pi levels at 6 and 12 hours after preservation were significantly higher in the CP group than in the SI group (62 +/- 5 versus 39 +/- 7%, 48 +/- 5 versus 22 +/- 8%, respectively, p < 0.05). Pcr/Pi levels at 6 and 12 hours after preservation were 30 +/- 9% and 22 +/- 8%, respectively in the CP group, while Pcr/Pi levels in the SI group were detected in only one case. There was no significant difference in water content either prior to or after 12-hour preservation between the two groups. Histopathologically, irregular expansion and/or contraction of myocardial fibers were more severe in the SI group than in the CP group. The recovery rate of hemodynamic parameters 2 hours after heart transplantation was significantly (p < 0.05) higher in the CP group than in the SI group. CONCLUSION: Stable and safe long-term canine heart preservation with continuous coronary perfusion associated with immersion is possible using this new apparatus, and may have broad clinical application. PMID- 10528748 TI - Combined treatment with endothelin- and PAF-antagonists reduces posttransplant lung ischemia/reperfusion injury. AB - BACKGROUND: Pathophysiologic changes of posttransplant lung ischemia/reperfusion injury are mediated by redundant cellular and humoral mechanisms. We investigated the protective effect of combined administration of platelet activating factor (PAF) and endothelin (ET) antagonists after prolonged ischemia in a small animal lung transplantation model. METHODS: Orthotopic left lung transplantation was performed after 20 hours cold ischemia in male Fischer (F344) rats weighing 200 250 g. Group I served as control. In Group II, donors received 1 mg/kg body weight of the endothelin antagonist TAK-044, and recipients 2 mg/kg. Group III was treated with the PAF antagonist TCV-309 (donor: 50 microg/kg; recipient: 100 microg/kg) (Takeda Chemicals Ltd.). Group IV received a combined treatment with both substances at the same dosage. Twenty-four hours after reperfusion, the native contralateral lung was occluded to assess gas exchange of the graft only, and 5 minutes later the thoracic aorta was punctured for arterial blood gas analysis (n = 5). In other animals (n = 5), lung tissue was frozen 24 hours after reperfusion and assessed for myeloperoxidase activity (MPO) and thiobarbituric acid reactive substances. RESULTS: Combined inhibition of PAF and ET-1 at the receptor level resulted in significantly improved graft function as compared to controls (Group I), and to groups treated with either TAK-044 or TCV-309. This was determined by a higher arterial oxygen content (112 +/- 9 mmHg, p = .00061 vs control, 48 +/- 5 mmHg), reduced MPO activity (0.35 +/- 0.02 deltaOD/mg/min, p = .000002 vs control, 1.1 +/- 0.1 deltaOD/mg/min) and reduced lipid peroxidation (59.5 +/- 2.5 pmol/g, p = .011 vs control, 78.5 +/- 4.1 pmol/g). The improvement of arterial oxygen (Group II 77 +/- 10 mmHg, p = .027 vs control; Group III 84 +/ 8 mmHg, p = .0081 vs control) and reduction of MPO activity (Group II 0.85 +/- 0.061 deltaOD/mg/min, p = .017; Group III 0.92 +/- 0.079 deltaOD/mg/min, p = .058) in groups treated with either a PAF antagonist or an ET antagonist was significantly less than in Group IV. CONCLUSIONS: Combined donor and recipient treatment with an ET antagonist and a PAF antagonist results in superior posttransplant graft function 24 hours after reperfusion, suggesting a synergistic role of ET-1 and PAF in the mediation of reperfusion injury in this model. Single treatment with either of the antagonists revealed only a slight improvement compared to untreated controls. PMID- 10528749 TI - Sudden death and tailored medical therapy in elective candidates for heart transplantation. AB - BACKGROUND: Due to the shortage of donor organs there is a long waiting time for heart transplantation. As a consequence, a high mortality rate on the waiting list diminishes the potential benefit of the procedure. Tailored medical therapy optimized according to the individual patients demands was introduced to select responding HTx candidates for continued management without transplantation. The development of modes of death over time (heart failure, sudden arrhythmic) in this population is unknown. METHODS: In 434 elective candidates for heart transplantation, submitted to our institution in the years 1984-1997 (50% coronary artery disease, mean age 51.6 +/- 12 years, 86% males) medical therapy was adjusted according to the results of repeated right heart catherizations. Adjuncts to conventional therapy with ACE inhibitors, digitalis and diuretics were amiodarone, beta-blockers, spironolactone, oral anticogulants, molsidomine or nitrates. Only patients not responding to these measures were processed to HTx. Clinical events (death, mode of death, HTx, resuscitation) were noted and analyzed by the Kaplan-Meier method and related to patients characteristics by multivariance analysis. RESULTS: During the mean follow-up of 2.36 +/- 2.4 years only 113 patients (25%) received a donor heart. One hundred-sixteen patients (26%) died without transplantation. Eighty-three (72%) of the deaths were sudden, 24 (20%) due to progression of heart failure and 9 (8%) due to other reasons. A shift from heart failure to sudden death was observed. Including 8 successful resuscitations due to documented VT/VF, there is a 20% risk of having a major arrhythmic event during the first two years of observation. Long-term (>1 year) medical responders had better hemodynamics at entry. Patients who died suddenly had similar clinical and hemodynamic data at entry than patients who needed an early transplant, but were in a comparable NYHA stage before death than long-term medical responders (2.15 +/- 0.8 vs 1.82 +/- 0.6, NS). Patients dying suddenly had significant more ventricular premature beats (1.6 +/- 2.9%/24 hours vs 1.06 +/- 2.8%/24 hours, p < .01) and complex ventricular arrhythmias (7.3 +/- 2.7/24 hours vs 1.98 +/- 5.6/24 hours, p < .01) than long-term responders. Seventy-five percent of all sudden death occurred during the first 2 observation years. CONCLUSIONS: The rate of heart failure death in elective candidates for heart transplantation under optimized medical therapy is low when patients are followed closely and transplant can be done rapidly after deterioration is recognized. Sudden death represents the highest risk for most patients. This event occurred predominantly in stable patients under tailored medical therapy without indication for HTx at that time. Our results strongly demand strategies for risk stratification and the investigation of prophylactic measures in this population. PMID- 10528750 TI - Acetylcholine but not sodium nitroprusside exerts vasodilation in pulmonary hypertension secondary to chronic congestive heart failure. AB - BACKGROUND: Reduced endothelium-dependent vasodilation contributes to the development of pulmonary hypertension in chronic congestive heart failure (CHF). We investigated pulmonary endothelium-dependent and independent vasodilation in patients with CHF. METHODS: We studied 42 patients with CHF (age 55 +/- 10, NYHA Classes II-III, left ventricular ejection fraction 27 +/- 10%, mean PAP 29 +/- 12 mmHg). The endothelial vasodilator capacity of pulmonary resistance vessels was assessed by the infusion of acetylcholine into a pulmonary artery branch while measuring the blood flow velocity with a Doppler flow wire. For comparison endothelium-independent vasodilation was measured with the response to sodium nitroprusside. The conductance vessel diameter (4.4 +/- 0.2 mm) was determined by intravascular ultrasound. Acetylcholine was administered at concentrations of 10( 6) to 10(-4) mol/l, sodium nitroprusside was administered at concentrations of 0.125 and 0.25 microg/kg per min. The effects on conductance vessel diameter were investigated in 12 patients by the measurement of diameter and flow velocity following the administration of acetylcholine and sodium nitroprusside. RESULTS: Acetylcholine markedly increased blood flow velocity (+39 +/- 7% at 10(-4) mol/l; p < .05). This correlated with the baseline PAP (r = 0.58; p < .05) and pulmonary vascular resistance (r = 0.58; p < .05). Sodium nitroprusside caused a small increase in the flow velocity (5 +/- 2% at 0.125, 12 +/- 4% at 0.25 microg/kg per minute; p < .05) that was accompanied by systemic vasodilation. The conductance vessel diameter was unchanged after acetylcholine was administered and was only marginally decreased after the administration of sodium nitroprusside. CONCLUSIONS: In CHF acetylcholine reveals preserved receptor-mediated endothelial vasodilation, that is positively correlated to pulmonary hypertension, and cannot be reproduced by sodium nitroprusside. PMID- 10528751 TI - Interstitial lung disease clinics for the management of idiopathic pulmonary fibrosis: a potential advantage to patients. Greater Manchester Lung Fibrosis Consortium. AB - BACKGROUND: Idiopathic pulmonary fibrosis (IPF) has a relatively poor prognosis with limited therapeutic intervention. This has led to varying practices, from a nihilistic approach at one end to more aggressive management at the other. However, a dedicated clinic with a multidisciplinary approach may offer advantages to patients with IPF. METHODS: A retrospective observational study was performed to compare patients with a diagnosis of IPF attending a general respiratory clinic between January 1988 and December 1996 to those attending a interstitial lung disease (ILD) clinic between January 1992 and December 1996. The notes were reviewed for (1) confirmation of diagnosis, (2) method of diagnosis, (3) date of initial consultation, and (4) survival. The end point was death, and the number of deaths that occurred up to October 1997 was recorded. RESULTS: The study identified 148 patients with a diagnosis of IPF. Ten patients underwent transplantation and were excluded. Of the remaining 138, 84 patients attended the general respiratory clinic (mean age 65 years: 53 men [63%], 31 women), and 54 attended the ILD clinic (mean age 56 years: 26 men [48%], 28 women). Patients attending the ILD clinic had a median survival of >3714 days. Patients attending the general respiratory clinic had a median survival of 1796 +/- standard error 437 days (CI 940-2652), p = .032 Breslow. Age was an important determinant of outcome. For patients less than 60 years of age (n = 60), the median survival for the ILD clinic was >3700 days, compared to 2535 +/- 577 days (CI 1404-280) in the general respiratory clinic, p = .037 Breslow. There was no difference in survival of patients over 60 years of age. CONCLUSIONS: The study suggests that a dedicated multidisciplinary clinic may result in an improved outcome for patients with IPF, particularly for patients younger than 60 years. This has implications that may facilitate the development of suitably powered therapeutic trials and may affect patient referral for transplantation. PMID- 10528752 TI - Comorbidities in end-stage lung disease. PMID- 10528753 TI - Detection of humoral rejection in human cardiac allografts by assessing the capillary deposition of complement fragment C4d in endomyocardial biopsies. AB - BACKGROUND: There are no well-established diagnostic criteria to detect humoral rejection in organ transplantation. The value of commonly used markers in immunohistochemistry, such as C1q, C3c, IgG, IgM and fibrinogen, is questioned by some groups. Complement fragment C4d is a more stable marker of complement activation as it is covalently bound to graft capillaries. C4d has been shown to identify clinically relevant, but otherwise undetectable humoral anti-graft reactions in human kidney transplants. METHODS: Immunohistochemical techniques were used to evaluate 155 endomyocardial biopsies from 56 heart transplant recipients less than 3 months post transplantation for the presence of capillary C4d staining. In a subset of patients, C4d staining was compared with C1q, C3c, IgM and fibrin staining and was correlated with clinical outcome. RESULTS: Within 3 months 9 of 56 patients died. Five of these nonsurvivors had prominent C4d staining (p < .05), whereas C1q, C3c and IgM showed no correlation with clinical outcome. Presence of fibrin correlated with clinical outcome and C4d staining (p < .05). CONCLUSIONS: The capillary deposition of complement split product C4d in human endomyocardial biopsies was significantly associated with graft loss. Determination of fibrin deposition may yield additional information to establish a diagnosis of humoral rejection. The immunohistochemical assessment of capillary deposition of C4d and fibrin appears to be an appropriate tool for the identification of patients, who may require additional or alternative immunosuppressive therapy targeted against the humoral immune system. PMID- 10528754 TI - Nefazodone and cyclosporine drug-drug interaction. AB - Depression is a significant post-transplant complication often necessitating drug therapy. Many of the newer selective serotonin reuptake inhibitor (SSRI) antidepressants are metabolized by the same cytochrome P450IIIA isoenzyme system that is responsible for the metabolism of cyclosporine, and these agents pose an interactive risk in transplant patients. We have observed nearly a 10-fold increase in whole blood cyclosporine concentrations in a cardiac transplant patient shortly after the addition of nefazodone antidepressant therapy. We suggest there is a clinically significant drug-drug interaction between nefazodone and cyclosporine due to inhibition of cytochrome P-450 IIIA4 isoenzymes by nefazodone. PMID- 10528755 TI - Immunohistochemical analysis of vascular prostheses implanted with the left ventricular assist system. AB - BACKGROUND: Dacron vascular prostheses are associated with thromboembolic complications and inflammatory responses; impregnation with bovine collagen reportedly stimulates additional inflammatory/immunologic complications. The Novacor (Baxter Healthcare Corp., Oakland, CA, USA) left ventricular assist system uses Dacron inflow and collagen-impregnated Dacron outflow prostheses. METHODS: Explanted inflow and outflow prostheses were evaluated for inflammatory/immunologic, hemostatic, anticoagulant, and fibrinolytic pathways. Non-implanted prostheses immersed in whole blood or plasma were used as controls. RESULTS: Immunoglobulins and complement components were observed in all prostheses with activated macrophages being present only in implanted prostheses. Antithrombin III was observed in all prostheses whereas fibrin, tissue plasminogen activator, and alpha-2 plasmin inhibitor were present only in implanted prostheses. Endothelial and smooth muscle cells associated with vascular structures containing collagen type IV and laminin were observed solely in implanted prostheses. CONCLUSION: An inflammatory response occurs and key components of hemostatic, anticoagulant, and fibrinolytic pathways are present within implanted prostheses. These processes are accompanied by endothelial and smooth muscle cell infiltration which appear to lay the foundation for neovessel development. PMID- 10528756 TI - Acquired atrial septal defect after heart transplantation. PMID- 10528757 TI - Reversal of myocardial interleukin-6-mRNA expression following long-term left ventricular assist device support for myocarditis-associated low output syndrome. PMID- 10528758 TI - Mitral valve myxoma: an unusual reason for rejecting a donor heart. PMID- 10528759 TI - Evolving trends in the treatment of anorectal diseases. AB - During the first half of this century, the safe and effective treatment of benign anorectal disorders perhaps did more to establish our specialty as a viable and distinct entity than anything else. A thorough understanding of anorectal anatomy and physiology, improved methods of local anesthesia, and an appreciation of proper postoperative care made the care of patients with diseases of the rectum and anus the domain of true specialists. Hirschman stated, "It is the action of the profession itself which has created the special field of proctology--the anus and rectum being organs peculiar to themselves and being subject to many medical and surgical diseases in the same way as the eye, the ear, the nose, the genital and urinary organs; and call for just as much special medical and surgical care. The general surgeon knows nothing about, and cares less for, the medical treatment of these organs; and the general practitioner who is able to treat the medical conditions is not, as a rule, properly equipped to do so. Thus, the proctologist came into existence--a man who, by special study of this particular region of the body, is able to give special care of either a surgical or medical nature, and often both in the same case, as may be required." PMID- 10528760 TI - Effect of delivery on anal sphincter morphology and function. AB - PURPOSE: Anal sphincter injury is a serious complication of childbirth, which may result in persistent anal incontinence. Occult injuries, visualized with endoanal ultrasonography, have previously been reported in up to 35 percent of females in a British study. The aim of the present study was to study anal sphincter morphology and function before and after delivery in primiparous females in the United States. METHODS: Thirty-eight primiparous patients (mean age, 31 years) were evaluated with endoanal ultrasonography, anal manometry, and pudendal nerve terminal motor latency during pregnancy and after delivery. Bowel function before and after delivery was recorded according to set questionnaires. Cesarean section was performed in three patients. RESULTS: Clinical sphincter tears, requiring primary repair, occurred in 15 percent of the patients. After delivery endoanal ultrasonography revealed disruptions in the external anal sphincter in six patients, but no patient had disruption in the internal anal sphincter. One patient had slight scarring in the external sphincter. Of the seven patients with pathologic findings at endoanal ultrasonography, the left pudendal latency increased after delivery (P < 0.05), and manometric results were reduced. Three of these seven patients had a third-degree or fourth-degree tear during delivery. All investigations were normal in the three patients who underwent cesarean section. CONCLUSIONS: The present study demonstrates a significant frequency of sphincter injuries (20 percent) after vaginal delivery. Obstetricians should be aware of this risk and explicitly inquire about incontinence symptoms at follow up after delivery. PMID- 10528761 TI - Do routinely measured delivery variables predict anal sphincter outcome? AB - PURPOSE: Trauma to the anal sphincter is a recognized complication of primiparous childbirth. This damage may be compounded during subsequent deliveries leading to symptoms. Earlier work is inconclusive as to which delivery variables are associated with such damage and may prove useful in predicting its occurrence, thereby allowing the potential for intervention in these later deliveries to protect the traumatized anal sphincter. The purpose of the present study was to determine whether routinely recorded obstetric variables can be correlated to anal sphincter damage in a consecutive series of females. METHODS: A prospective study was undertaken in a single maternity unit. Patients delivering were assessed before discharge using a symptom questionnaire and endoanal ultrasound. Delivery data were collected prospectively and analyzed statistically to see if a significant difference existed in the presence of an anal sphincter defect. RESULTS: A total of 159 patients were assessed. Endosonography revealed sphincter injuries in 8.7 percent of the normal vaginal delivery group and 83 percent of the forceps delivery group. No correlation was found between head circumference, baby weight, maternal body mass index, epidurals, episiotomy, length of each stage of labor, and duration of active pushing. Forceps delivery was the only factor to be significantly associated with sphincter trauma. CONCLUSION: Besides forceps delivery, commonly measured delivery variables are not useful predictors of anal sphincter trauma. Normal vaginal deliveries do not warrant routine postnatal anorectal assessment, but this should be routine for all instrument deliveries. PMID- 10528762 TI - Rectal excision and colonic pouch-anal anastomosis for rectal cancer: oncologic results at five years. AB - PURPOSE: Preservation of the anal sphincter is now accepted as a primary aim in surgical treatment of rectal cancer. The use of colonic J-pouch-anal anastomosis after complete rectal excision is one method that permits retention of continence without compromising oncologic principles. This study aimed to assess carcinologic results of rectal excision followed by colonic J-pouch anal anastomosis, with particular reference to rate of locoregional recurrence. METHOD: From 1984 to 1990 complete rectal excision and colonic pouch-anal anastomosis were performed in 167 patients for cancer of the middle or low rectum. A total of 154 patients were followed for this study for a minimum of five years, with evaluation of the frequency of locoregional recurrence. RESULTS: Sixty-five patients died during the period of surveillance, giving a five-year survival rate of 68.8 percent. Twenty patients (13 percent) presented with locoregional recurrence at an average of 31 months after surgery. In 11 cases (7 percent) the local recurrence was not associated with metastatic disease, and six of these patients underwent further curative surgery. CONCLUSIONS: These results confirm that coloanal anastomosis after complete rectal excision is a valuable option in the surgical treatment of rectal cancer and is accompanied by a frequency of isolated locoregional recurrence of less than 7 percent, of which half underwent surgical resection with curative intent. PMID- 10528763 TI - Coloanal anastomosis for distal third rectal cancer: prospective study of oncologic results. AB - PURPOSE: Jeopardizing cure and risking high local recurrence have served as arguments against sphincter-saving resection for patients with distal third rectal cancer. This prospective study examines and compares the local recurrence and survival rates in patients with distal third rectal cancer treated by either coloanal anastomosis or abdominoperineal resection. METHODS: Between 1977 and 1993, 174 patients underwent coloanal anastomoses and 38 patients underwent abdominoperineal resection. All tumors were located 4 to 7 cm from the anal verge. One hundred ninety-three patients (91 percent) underwent rectal excision with a curative intent. Mean follow-up was 66 months after sphincter-saving resection and 65 months after abdominoperineal resection. RESULTS: Mean anastomotic height from the anal verge was 2.3 cm after sphincter-saving resection. Overall local recurrence rate was 7.9 percent after sphincter-saving resection and 12.9 percent after abdominoperineal resection. The five-year actuarial survival rate was 78 percent after sphincter-saving resection and 74 percent after abdominoperineal resection. CONCLUSION: Local recurrence and survival are not compromised in patients with distal third rectal cancer when treated by sphincter-saving resection, provided that oncologic principles are not violated. Coloanal anastomosis can be performed with an acceptable morbidity. PMID- 10528764 TI - Preoperative combined radiotherapy and chemotherapy for rectal cancer does not affect early postoperative morbidity and mortality in low anterior resection. AB - PURPOSE: It is not yet known whether preoperative combined radiotherapy and chemotherapy for rectal cancer affects postoperative mortality and morbidity. We therefore evaluated early postoperative complications in patients given adjuvant radiotherapy and chemotherapy before surgery for middle and lower rectal adenocarcinoma. METHODS: Between 1994 and 1998, 41 patients underwent combined preoperative pelvic radiotherapy and chemotherapy at our institution. Most of the patients had 45 Gy (1.8 Gy/day/25 fractions) during five weeks plus 5 fluorouracil (350 mg/m2/day) and low-dose leucovorin (10 mg/m2/day) bolus on Days 1 to 5 and 29 to 33. Surgery was performed four to six weeks after completion of adjuvant therapy. The 41 patients (Group A) were retrospectively compared with 30 patients (Group B) who, in the same period, underwent surgery without preoperative adjuvant therapy. The groups were homogeneous for age, gender, preoperative risk factors, operating surgeon, and pathologic stage. Mean distance of the tumor from the anal verge was shorter in Group A patients (P = 0.031). RESULTS: There were seven major postoperative complications in each group. No significant differences were found between the groups for morbidity and mortality rates. Considering all patients, more postoperative complications were found in patients scored as American Society of Anesthesiologists 3, in those with a preoperative hemoglobin value < 10 g/dl, and in those without a diverting stoma (P = 0.0048, P = 0.0453, and P = 0.0033, respectively). At multivariate analysis, independent predictors of major complications were American Society of Anesthesiologists score (relative risk, 343; P = 0.022), diverting stoma (relative risk, 159; P = 0.010), type of surgical procedure (relative risk, 38.9; P = 0.048), preoperative hemoglobin value (relative risk, 9.72; P = 0.061), and intraoperative blood loss (relative risk, 1; P = 0.027). In Group A patients, the absence of diverting stomas was associated with major postoperative complications (P = 0.0307), and independent predictors of major complications were American Society of Anesthesiologists score (relative risk, 56; P = 0.111) and absence of a diverting stoma (relative risk, 22.42; P = 0.222). CONCLUSION: Early postoperative complications after resection for middle and lower rectal adenocarcinoma are affected by intraoperative and preoperative risk factors and absence diverting stomas, but not by preoperative adjuvant therapy. PMID- 10528765 TI - Hepatic lymph node involvement in resected cases of liver metastases from colorectal cancer. AB - PURPOSE: Lymph node metastasis in the hepatoduodenal ligament is known as one of the most significant prognostic factors after liver resection for colorectal metastasis. However, there have been very few articles on the clinical features of node-positive patients and on detailed distribution of positive nodes. Further, there has been no established strategy on how to handle hepatic lymph nodes during liver resection. To address these subjects, a retrospective study was conducted. METHODS: During the period of 1980 through April 1998, 182 hepatic resections were performed for metastatic colorectal carcinoma. Of these, 78 cases had hepatic lymph node sampling during the operation. Distribution of positive nodes, location of liver metastasis, stage of the primary lesion, and outcome after liver resection were analyzed. RESULTS: Nine cases (12 percent) had secondary lymph node metastases in the hepatoduodenal ligament. The incidence was slightly higher (13.5 percent) in the most recent 44 consecutive cases. There was a tendency for liver metastases in the right lobe to metastasize to No. 12b (or node of the foramen of Winslow, lymph nodes along the common bile duct) and liver metastases in the left lobe to metastasize to No. 8a (anterosuperior group of the lymph nodes along the common hepatic artery). Outcome of node-positive patients (n = 9) was extremely poor (P < 0.001) compared with that of node-negative patients (n = 66), and the most common site of recurrence in the node-positive patients was remnant liver and hepatic lymph nodes. Preoperatively, there were no significant predicting factors for positive hepatic lymph nodes. CONCLUSIONS: No. 8a and No. 12b nodes are principal nodes that should be palpated and sampled during liver resection to check the secondary lymphatic spread from liver metastases. Hepatic nodal involvement indicates the progression of disease beyond simple liver metastases and may not be the indication for simple surgical removal. Further study, including hepatoduodenal dissection and systemic adjuvant chemotherapy, may elucidate the survival benefit, if any, of liver resection in node-positive patients. PMID- 10528766 TI - Hypothermia in open and laparoscopic colorectal surgery. AB - PURPOSE: Perioperative hypothermia has been shown to be an important determinant of outcome after open colorectal resections. The degree of hypothermia occurring with laparoscopic-assisted colorectal surgery is, however, unknown, and the effectiveness of standard warming measures is untested. This study was designed to assess hypothermia in open and laparoscopic-assisted colonic resections using a standardized warming protocol. METHODS: A prospective, nonrandomized study was performed with temperature measurements recorded every ten minutes. Statistical analysis was based on repeated measures analysis of variance models with significance set at the conventional 95 percent (two tailed). RESULTS: A total of 107 patients were entered into the trial; 68 had open and 39 had laparoscopic colectomies. The groups were well matched for age, weight, and duration of surgery, with a median operating time of 180 minutes in each group. The average drop in temperature from commencement of surgery to lowest point was 0.68 degrees C (standard deviation, 0.08) in the open group, compared with 0.53 degrees C (standard deviation, 0.06) in the laparoscopic group (P = 0.126). CONCLUSIONS: Laparoscopic-assisted colorectal surgery is not associated with a higher incidence of perioperative hypothermia than open colorectal surgery using a standard warming regimen for both groups. On the basis of these results, standard temperature conservation is adequate, even for long, complex laparoscopic procedures. PMID- 10528767 TI - Sphincter preservation of leiomyosarcoma of the rectum and anus with local excision and brachytherapy. AB - PURPOSE: The aim of this study was to determine the outcome of patients with leiomyosarcoma of the rectum or anus treated with local excision and brachytherapy. METHODS: Eight patients with leiomyosarcoma of the rectum (7 patients) or anus (1 patient) were treated with a transanal excision followed by a temporary iridium-192 interstitial implant to 4,500 cGy. Median tumor size was 4.2 (range, 1.5-5) cm. Margins were positive in six patients, negative in one patient, and close in one patient. RESULTS: With a median follow-up of 53 months, median survival time was 53 months and the three-year actuarial survival rate was 71 percent. The cumulative incidence of failure as a component of failure for local was 25 percent (2/8), for abdominal was 0 percent (0/8), and for distant was 25 percent (2/8). Four patients eligible for functional analysis all had excellent sphincter function (1-2 bowel movements per day, no soilage). CONCLUSION: In selected patients the use of conservative surgery followed by brachytherapy is a reasonable alternative to an abdominoperineal resection. However, more experience and longer follow-up are needed before this approach can be recommended routinely. PMID- 10528768 TI - Estrogen replacement therapy and colorectal cancer risk in elderly women. AB - PURPOSE: Colorectal cancer is the fourth most common incident cancer in the United States and causes more cancer deaths than any site except lung. Twenty-two epidemiologic studies have examined the relationship of estrogen replacement therapy and colon and rectal cancers with inconsistent results. However, recent studies suggest a reduced risk among current users. The purpose of the present study was to analyze the Leisure World Cohort for possible association of estrogen replacement therapy with colorectal cancer risk. METHODS: A cohort of 7,701 female members who were initially free of cancer and self-reported their use of estrogen replacement therapy were followed up from June 1981 through December 1995 for development of colorectal cancer. RESULTS: We observed 249 incident colorectal cancer cases and 89 colorectal cancer deaths. Women who had used estrogen replacement therapy had an age-adjusted colorectal cancer incidence rate of 2.67 per 1,000 person-years compared with 3.30 per 1,000 person-years among lifetime nonusers (relative risk = 0.81; 95 percent confidence interval, 0.63 to 1.04). Among recent users the incidence was one-third lower than among lifetime nonusers (relative risk = 0.66; 95 percent confidence interval, 0.44 to 0.98). Risk did not differ by duration of estrogen replacement therapy, usual dose of conjugated estrogen, or route of estrogen administration. The effects of current estrogen replacement therapy on colon cancer incidence (relative risk = 0.70; 95 percent confidence interval, 0.45 to 1.09), right-sided colon cancer incidence (relative risk = 0.75, 95 percent confidence interval, 0.38 to 1.48), left-sided colon cancer incidence (relative risk = 0.76; 95 percent confidence interval, 0.41 to 1.41), rectal cancer incidence (relative risk = 0.52; 95 percent confidence interval, 0.21 to 1.31), and colorectal cancer mortality (relative risk = 0.82; 95 percent confidence interval, 0.44 to 1.54) were similar. CONCLUSION: A reduced risk of colorectal cancer may be an additional benefit of recent estrogen replacement therapy use, which should be considered by postmenopausal women when deciding whether to use hormones. PMID- 10528769 TI - Long-term results of lateral internal sphincterotomy for chronic anal fissure with particular reference to incidence of fecal incontinence. AB - PURPOSE: Lateral internal sphincterotomy is the procedure of choice for chronic anal fissure because it relieves symptoms and heals the fissure in nearly all patients. However, there is evidence that fecal incontinence complicates lateral internal sphincterotomy. The aim of this study was to examine the outcome of lateral internal sphincterotomy in terms of fissure healing and incidence of fecal incontinence. METHODS: Between 1984 and 1996, 585 patients underwent lateral internal sphincterotomy and were surveyed by questionnaire. Eighty-three percent (487/585) responded. The mean follow-up was 72 (range, 6-145) months. RESULTS: Fissures had healed by a median of three weeks after surgery in 96 percent of patients. Recurrent fissures occurred in 8 percent. Two thirds of the recurrent fissures healed on conservative management alone. Ninety-eight percent of patients were satisfied with the outcome of surgery, but some degree of fecal incontinence occurred in fully 45 percent of patients at some time in the postoperative period. Incontinence occurred in 53.4 percent of women and 33.3 percent of men (P < 0.05). Incontinence to flatus, mild soiling, and gross incontinence occurred in 31, 39, and 23 percent of patients, respectively. However, by the time of survey (a mean of >5 years after lateral internal sphincterotomy) 6 percent reported incontinence to flatus, 8 percent had minor fecal soiling, and 1 percent experienced loss of solid stool. Importantly, only 3 percent of patients stated that incontinence had ever affected their quality of life. CONCLUSION: Although lateral internal sphincterotomy heals and relieves symptoms of chronic anal fissure in nearly all patients (96 percent), incontinence occurs frequently. Most episodes of incontinence are indeed minor and transient, but in a small subgroup, incontinence seems to be permanent. PMID- 10528770 TI - Pattern of mucosal adaptation in acute and chronic pouchitis. AB - PURPOSE: Variant pathological changes have been observed in ileoanal pouches, including inflammation, villous atrophy, and crypt hyperplasia. Therefore, we investigated the type and degree of mucosal adaptation in patients with ulcerative colitis and familial adenomatous polyposis. METHODS: Forty-two patients with ulcerative colitis and 14 patients with familial adenomatous polyposis with ileoanal pouches were assessed. Samples were taken from three months to eight years after creation of an ileoanal pouch. Mucosal architecture was examined by morphometry after microdissection. RESULTS: Structural changes of the mucosa can be categorized into three groups. Compared with preoperative values, patients without pouchitis (73 percent) has only minor decrease of villous length (402 microm vs. 540 microm) and increase in crypt depth (274.5 microm vs. 177 microm). In patients with acute pouchitis (20 percent), a slight increase in villous length (477 microm vs. 402 microm) and pronounced crypt hyperplasia (376 microm vs. 274.5 microm) was observed compared with noninflamed ileoanal pouches. In contrast, in patients with chronic pouchitis (7 percent), severe villous atrophy (62.5 microm) and crypt hyperplasia (543 microm) was found. CONCLUSIONS: Minor structural changes of ileoanal pouch mucosa develop early as an adaptive response to a new environment. Only in a small group of patients with chronic pouchitis does severe villous atrophy and crypt hyperplasia of the ileoanal pouch mucosa develop, most likely as a consequence of mucosal inflammation. PMID- 10528771 TI - Effects of restorative proctocolectomy on renal and adrenal function. AB - PURPOSE: Restorative proctocolectomy is a standard procedure in the surgical treatment of ulcerative colitis and familial adenomatous polyposis. The radical removal of the colorectum with construction of an ileostomy often results in high stoma losses. These may lead to changes in the electrolyte and acid-base balance and to alterations in renal and suprarenal gland function. METHODS: In this study 33 patients who received an ileoanal pouch before and after proctocolectomy were investigated at different time intervals for electrolyte changes, alteration of the acid-base balance, kidney function, and hormonal changes of the suprarenal glands. Measurements were performed before proctocolectomy, ten days after proctocolectomy with ileal pouch-anal anastomosis under protective loop ileostomy, before ileostomy closure, and 6 to 12 months after ileostomy closure. Neither acute renal failure nor other vital complications were observed. RESULTS: Statistical analysis showed a significant decrease of urine pH to 5.4 +/- 0.22 (before ileostomy closure) and metabolic acidosis (pH 7.32 +/- 0.04; base excess 1.3 +/- 5.6 (before ileostomy closure)). Likewise, we found a decrease in renal clearance to 86 ml/minute (before ileostomy closure) without signs of tubular damage. The most important change during the phase with ileostomy was a functional secondary hyperaldosteronism with aldosterone levels of 63.2 +/- 70.8 ng/dl (before ileostomy closure). In comparison with preoperative levels, there was a ten-fold increase in mineralocorticoid adrenal activity. Additionally, during the period with protective ileostomy, the hepatic synthesis of aldosterone 18-glucuronide was only slightly increased, and the cortisol/cortisone ratio was extremely decreased. CONCLUSIONS: These results show that restorative proctocolectomy with ileal pouch-anal anastomosis and protective loop ileostomy significantly influences fluid, electrolyte, and acid-base balance. Functional secondary hyperaldosteronism is of central importance for subsequent renal recompensation. Approximately one-half year after ileostomy closure, the endogenous hormones with mineralocorticoid effects returned to normal levels. PMID- 10528773 TI - Antiperistaltic ileostomy using the long terminal ileal segment. AB - PURPOSE: This study was undertaken to determine whether reversed terminal ileal segments can be used to decrease ileostomy output in patients who have undergone total proctocolectomy and ileostomy for ulcerative colitis or familial adenomatous polyposis. METHODS: An approximately 25-cm length of terminal ileum was reversed in an antiperistaltic manner, and the new terminal ileal end was used for the ileostomy constructed in the usual manner. Six patients underwent this procedure and were compared with six patients who had conventional total proctocolectomy and ileostomy. Variables studied included weight of ileostomy output and the weight of the filtered fluid component. Data were obtained on seven different occasions during a two-month period beginning three months after the operation. Analysis was done using Student's t-test. RESULTS: There was a statistically significant decrease in the weight of the average 24-hour ileostomy effluent in those patients undergoing reversed antiperistaltic loop procedures. There was also a statistically significant decrease in the filterable liquid proportions. CONCLUSIONS: The antiperistaltic ileostomy is effective in reducing the daily amount of ileostomy effluent and facilitates stoma care, owing to its diminished liquid component. PMID- 10528772 TI - C-reactive protein and leukocyte count in the diagnosis of acute appendicitis in children. AB - PURPOSE: The aim of this study was to analyze the diagnostic accuracy of C reactive protein and its possible advantage, if any, over leukocyte counts in acute appendicitis in children. METHODS: We performed a retrospective study of 124 children (72 males) with a mean age of 9.3 (range, 2-14) years operated on under a clinical diagnosis of acute appendicitis. The diagnosis of acute appendicitis, confirmed by pathologic examination of the removed appendix, was then correlated with C-reactive protein, leukocyte count, and a combination of both C-reactive protein and leukocyte count, with a logistic regression model. C reactive protein serum measurements were performed by an immunoturbidimetric test. The patients were divided into two groups according to the pathologic features of the removed appendix: Group A (n = 104), patients with acute appendicitis, and Group B (n = 20), patients without acute appendicitis. To assess the accuracy of C-reactive protein, leukocyte counts, and a combination of both parameters, receiver operating characteristic curves were used. The areas under the curve were compared using the maximum likelihood estimation method. RESULTS: There were 95 cases (76.6 percent) of nonperforated appendicitis, 9 cases (7.3 percent) of perforated appendicitis and 20 cases (16.1 percent) of normal appendix. Mean C-reactive protein in Group A was 4.3 (standard deviation, 6.6) and in Group B was 1.2 (standard deviation, 1.7; P = 0.03). The C-reactive protein and leukocyte count values were correlated with the pathologic diagnosis of acute appendicitis. Mean C-reactive protein values increase as the pathologic inflammation type progresses (P = 0.007). The C-reactive protein receiver operating characteristic curve shows that the C-reactive protein value with highest accuracy was 1.7 mg/dl. The sensitivity, specificity, and accuracy rates calculated in the 1.7 cutoff were 58, 80, and 83.8 percent, respectively. A comparison of the respective receiver operating characteristic curves demonstrates that C-reactive protein, leukocyte count, and the combination of both tests all have a good diagnostic value but without any significant difference (P = 0.2). CONCLUSIONS: In children, 1) serum C-reactive protein is increased in acute appendicitis; 2) such increase is related to the severity of the appendiceal inflammation; and 3) although serum C-reactive protein has an adequate diagnostic accuracy, neither individually nor in combination with the leukocyte count is it significantly better than the leukocyte count alone. PMID- 10528774 TI - Technical manual for manufacturing autologous fibrin tissue adhesive. AB - PURPOSE: The aim of this article is to provide a concise and simple technical manual for manufacturing autologous fibrin tissue adhesive derived from the precipitation of fibrinogen using a combination of ethanol and freezing for surgery. METHODS: All materials and equipment needed to manufacture ethanol-based autologous fibrin tissue adhesive are listed. In addition, step-by-step instructions are provided to allow for easy and rapid fibrin adhesive production. RESULTS: Ethanol-based autologous fibrin tissue adhesive can be manufactured in under 60 minutes. Furthermore, at our institution the startup cost for manufacturing ethanol-based autologous fibrin tissue adhesive was under $2,500.00. CONCLUSION: Ethanol-based autologous fibrin tissue adhesive is a safe, reliable, and easily manufactured autologous fibrin tissue adhesive that can be made by a trained technician in any blood bank, pharmacy, or surgical laboratory. PMID- 10528775 TI - Rectal Dieulafoy's lesion: report of a case and review of the literature. AB - Dieulafoy's lesion is an uncommon cause of gastrointestinal bleeding that occurs after rupture of an exposed submucosal artery. The vast majority of lesions are found in the stomach, but cases have been described in the esophagus, small intestine, colon, and rectum. We describe an elderly patient who presented with severe lower gastrointestinal bleeding caused by a rectal Dieulafoy's lesion. This is the first report of a rectal Dieulafoy's lesion treated successfully with endoscopic epinephrine injection followed by thermocoagulation. We review the physiopathology, clinical presentation, diagnosis, and treatment of this disease. PMID- 10528776 TI - Treatment of grade 3 anal intraepithelial neoplasia by complete anal mucosal excision without fecal diversion: report of a case. AB - PURPOSE: The aim of this study was to remove completely the risk of malignant transformation without permanent or temporary fecal diversion in a patient with extensive anal intraepithelial neoplasia. METHODS: All anal canal mucosa and the lowest 1.5 cm of rectal mucosa were excised and the adjacent rectal mucosa and submucosa advanced to the anal verge skin. RESULTS: The patient achieved normal continence within a month after the operation. Multiple anal canal biopsies at 12 months after the operation revealed normal rectal mucosa. CONCLUSIONS: Total anal mucosal excision offers a relatively simple means of removing the malignant risk of anal intraepithelial neoplasia without fecal diversion in selected patients. PMID- 10528777 TI - Isolated splenic metastasis from colorectal carcinoma: report of a case and review of the literature. AB - In general, surgical resection for metastatic colorectal adenocarcinoma rarely benefits more than a small percentage of patients long term. In the case of isolated splenic metastasis without evidence of other metastatic disease, splenectomy may increase survival times in patients with previously resected colorectal adenocarcinoma. Currently there are only five case reports involving isolated splenic metastases in the English-language literature. This article presents a sixth case and a review of the previous five cases in the literature, with a discussion on the possible diagnostic and therapeutic approaches to these rare but important cases and the apparently significant survival advantage of early diagnosis and treatment. PMID- 10528778 TI - Current trends in restorative proctocolectomy: introduction of an ultrasonically activated scalpel. AB - PURPOSE: We evaluated the usefulness of an ultrasonically activated scalpel (Harmonic Scalpel) for mucosal proctocolectomy and ileal J-pouch-anal anastomosis. METHODS: Seventy-four patients with ulcerative colitis (70 patients) and familial adenomatous polyposis (4 patients) underwent mucosectomy using the Harmonic Scalpel since 1997. We compared the clinical and functional results with those of the monopolar electrocoagulator (forceps coagulation technique, 86 patients with colitis and 7 with polyposis). RESULTS: We performed graduated mucosal proctectomy by using the Harmonic Scalpel. The operative time (Harmonic Scalpel, 42 minutes vs. forceps coagulation technique, 85 minutes) and blood loss (Harmonic Scalpel, 33 ml vs. forceps coagulation technique, 86.8 ml) were significantly reduced by this method. The Harmonic Scalpel enabled restorative proctocolectomy by the synchronous approach within three hours. The functional results and complications were not significantly different between the two groups. CONCLUSION: The Harmonic Scalpel shortened the operative time, decreased blood loss, and was useful for restorative proctocolectomy in our study. PMID- 10528779 TI - Intraoperative localization of colorectal tumors in the early stages using a marking clip detector system. AB - In laparoscopic colectomy the identification of the site of a tumor is often difficult. The topical injection of india ink or blue dye by preoperative colonoscopy is the most prevalent method to mark the tumor site; however, such a procedure also includes the intrinsic danger of possibly injecting dye into the peritoneal cavity. In addition, the injected marker may also spread so widely that the intended site may become obscure. A marking clip detector system was used to detect metallic marking clips in the luminal side that had been applied to the mucosa adjacent to the lesion during the course of preoperative colonoscopy. This method was able to identify the marked site in 40 percent of cases in which only one clip was applied to the mucosa. However, when the lesion sites were marked with two or three clips, then the detection rate increased to 100 percent. Based on our findings, this procedure was found to be a safe and reliable method for identifying lesions during laparoscopic-assisted colectomy. PMID- 10528780 TI - Sodium fusidate. Preface PMID- 10528781 TI - Fusidic acid adverse drug reactions. AB - Unlike trials conducted today on new antimicrobials, the introduction of fusidic acid was not accompanied by extensive studies on toxicity and side effects. The early studies on small numbers of patients reported fusidic acid to be a nontoxic drug with the main side effects being gastrointestinal tract discomfort, diarrhoea and headache. Case reports of hepatotoxicity were reported from 1972. A retrospective analysis showed this to be reversible and mainly associated with intravenous administration. Rarely reported side effects are granulocytopenia, thrombocytopenia and venous spasm. Skin reactions, including contact dermatitis, are uncommon. After many years of use the most common side effects reported for fusidic acid are minor and relate to the gastrointestinal tract. PMID- 10528782 TI - Fusidic acid in other infections. AB - Fusidic acid, both systemic and topical, has been used for a wide variety of less common infections. Efficacy for oral fusidic acid has been demonstrated in the treatment of Clostridium difficile colitis and in staphylococcal infections in patients with cystic fibrosis. Topical fusidic acid gel is also effective in bacterial conjunctivitis and other minor external eye infections, and may be effective in reducing bacterial flora in the conjunctival sac prior to eye surgery. Studies suggest a potential role for fusidic acid in neurosurgical prophylaxis, as adjunctive therapy in bacterial endophthalmitis and Legionella pneumonia, and in leprosy. Topical fusidic acid has no effect in the treatment of chlamydial conjunctivitis or the prevention of staphylococcal infections in patients on continuous ambulatory peritoneal dialysis. PMID- 10528783 TI - Fusidic acid in septicaemia and endocarditis. AB - The in vitro activity of fusidic acid against Staphylococcus aureus is confirmed in clinical studies which demonstrate that this antibiotic in combination with other agents, particularly beta-lactams, is efficacious in non-MRSA septicaemia. Some reports suggest that fusidic acid when used in combination, may significantly diminish the risk of relapse after cessation of therapy. However, in spite of over 30 years of use and its recommendation in a number of guidelines, published evidence is insufficient to allow reliable comment on the efficacy of the antibiotic in the treatment of endocarditis. PMID- 10528784 TI - Fusidic acid pharmacology, pharmacokinetics and pharmacodynamics. AB - Fusidic acid comes in a variety of formulations for oral, intravenous and topical use. After oral administration of 500 mg Cmax values range from 14.5-3.3 mg/l and an elimination half-life of 8.9-11.0 h. Similar values are obtained with intravenous administration of the sodium salt, although peaks tend to be higher. Bioavailability for the new film-coated tablet is approximately 91% while that of the suspension formulation appears to be much lower. Repeated dosing results in substantial drug accumulation when given 8-hourly, and to a variable extent depending on dose when administered 12-hourly. One study has demonstrated a modest dose-dependency for pharmacokinetics, with decreased clearance at higher doses. Fusidic acid is primarily eliminated by non-renal mechanisms, and a proportion of the drug is metabolised to seven or more breakdown products that can be detected in bile. Hypoalbuminaemia increases fusidic acid clearance, while clearance is decreased in the presence of severe cholestasis, and essentially unchanged in renal failure. Fusidic acid is highly protein-bound (91-98S), but has good penetration to a number of tissues including skin blisters, burns, infected bone and joints. Topical application of fusidic acid results in poor penetration through skin but good penetration into aqueous and vitreous humour. Little is known about the pharmacodynamics of fusidic acid, apart from the fact that it is slowly bactericidal against Staphylococcus aureus, and produces moderate post-antibiotic effects in vitro. PMID- 10528785 TI - Resistance to fusidic acid. AB - Resistance to fusidic acid is determined by a number of mechanisms. The best described are alterations in elongation factor G, which appear in natural mutants that are harboured at low rates in normal populations of staphylococci (10(6) to 10(8)). Altered drug permeability has also been described, and appears to be plasmid-borne. Binding by chloramphenicol acetyltransferase type I and efflux are other described mechanisms of resistance whose prevalence is unclear. A large number of studies have examined rates of fusidic acid resistance in staphylococci. Most show low levels of resistance. Studies where high levels of resistance have been seen are from areas of the hospital where cross infection is common. Rates of resistance have tended to be slightly higher in methicillin resistant strains of Staphylococcus aureus. Studies on the evolution of resistance have shown no major trends to the emergence of resistance. In one case this is despite increasing use of both systemic and topical fusidic acid over more than 24 years. Selection for resistant variants during treatment was recognised early in vitro and in vivo. However, evidence suggests that it does not occur at high frequency in clinical practice. Nevertheless, accumulated experience is that fusidic acid in combination with other agents results in less resistance emergence. PMID- 10528786 TI - Fusidic acid in vitro activity. AB - Fusidic acid is a narrow spectrum agent that acts to inhibit protein synthesis by inhibition of elongation factor G at the level of the ribosome. Because of high protein binding susceptibility testing in vitro is affected by the presence of blood or serum. In addition, there is a modest inoculum effect in vitro. A breakpoint of 1 or 2 mg/l is most widely used for defining resistance to systemic treatment with fusidic acid. Fusidic acid activity is principally directed at staphylococci, both Staphylococcus aureus and coagulate-negative species which are highly susceptible. It is also active against Gram-positive anaerobic activity, and shows in vitro activity against Neisseria spp., Bordetella pertussis and Moraxella catarrhalis. It has no activity against other aerobic Gram-negative species. Modest activity (MICs just above breakpoint values) is seen with Streptococcus and Enterococcus spp. as well as Gram-negative anaerobic bacteria. Fusidic acid is defined as bacteriostatic. For staphylococci MBC values are generally 8--32-fold that of the MIC. Interaction studies with other antibiotics give varying results depending on methodology. However, interaction with beta-lactams is generally indifferent, as it is with rifampicin, while aminoglycosides and macrolides appear to be synergistic and fluoroquinolones antagonistic. Fusidic acid appears to inhibit the function of neutrophils and T lymphocytes at clinically achieved concentrations. PMID- 10528787 TI - Fusidic acid in skin and soft tissue infections. AB - Skin and soft skin tissue infections are usually caused by Staphylococcus aureus and Streptococcus pyogenes. In vitro data show good activity of fusidic acid against staphylococci but the minimal inhibitory concentrations for streptococci are relatively high indicating marginal activity. A limited number of clinical trials have been performed using oral fusidic acid and although all have methodological problems the difference in susceptibility of these two organisms is apparent. The end of study cure rates for these studies were 91-99% for S. aureus and 75-85% for S. pyogenes. Topical therapy has been used in a number of forms and for different skin infections. Comparative studies have been conducted with mupirocin, trimethoprim/polymixin cream, hydrogen peroxide and combination steroid preparations. For most of these studies fusidic acid was equivalent to the comparator agent except where there was a proven S. pyogenes infection. Studies with topical fusidic acid have also been reported in specific disease states such as acne, erythrasma, and abscesses with good results. PMID- 10528788 TI - Fusidic acid in the treatment of methicillin-resistant Staphylococcus aureus. AB - The emergence of MRSA in the 1960s coincided with the introduction of fusidic acid. Since that time, the antibiotic has been widely used against this organism, both in the 1960s and 1970s and against the more modern multi-resistant version of the 1980s and 1990s. Although showing potent in vitro activity, experimental in vivo and in vitro investigations have demonstrated disappointing results. Clinical efficacy has been demonstrated in a series of small studies and case reports, particularly when fusidic acid is used in combination for the treatment of MRSA carriage. Given the widespread use of fusidic acid against MRSA, published evidence supporting its efficacy is limited and does not support the use of the agent as monotherapy. PMID- 10528789 TI - Fusidic acid non-antibacterial activity. AB - In 1978 the first report was published indicating that fusidic acid had an immunomodulatory effect. This study describing an in vitro effect on white cells was followed by an in vivo study showing improved survival in heart transplanted mice. In 1990 these immunomodulatory effects were shown to be related to suppression of cytokine production in animal models of septic shock and insulin dependent diabetes. Disease modulation has been investigated in a number of immunologically mediated diseases, HIV infection, Behcet's disease, Crohn's disease, uveitis and scleroderma. However whether fusidic acid has any useful function as an immunomodulating agent has yet to be fully explored. PMID- 10528790 TI - Fusidic acid in bone and joint infections. AB - The prominence of staphylococci as the causative agent in bone and joint infections suggests that fusidic acid (FA) has a potentially important role in their treatment. FA has been studied in a broad range of orthopaedic infections, mostly in combination with other antimicrobials. For susceptible organisms, particularly Staphylococcus aureus, it has demonstrable efficacy in acute osteomyelitis, chronic osteomyelitis, specialised forms of osteomyelitis such as calcaneal and vertebral infection, septic arthritis, prosthetic and other device related infections. A small number of studies have also examined the use of FA alone for the treatment of bone infections, with evidence of good efficacy, as well as the local application of FA in plaster-of-Paris (POP) beads, or incorporated into bone cement, again with promising results. Further studies are required to confirm the efficacy of FA in the treatment of orthopaedic infections caused by methicillin-resistant strains of S. aureus. PMID- 10528792 TI - Genetics of rheumatoid arthritis (RA): two separate regions in the major histocompatibility complex contribute to susceptibility to RA. AB - We analyzed HLA-DR antigens and microsatellite Bat2 alleles in 97 adult caucasian patients with classical seropositive rheumatoid arthritis (RA) and 95 normal healthy controls. The results demonstrate that the prevalence of microsatellite Bat2 138 allele was significantly higher in RA-susceptibility DRB1 QKRAA/QRRAA epitope-negative patients as compared with normal controls. Analysis of the data suggested that Bat2 138 allele has primary association with RA-susceptibility in QKRAA/QRRAA epitope-negative patients. The Bat2 138 allele thus provides an additional risk in RA-susceptibility. In addition, microsatellite Bat2 138 allele showed a highly significant positive association with microsatellite D6S273 138 allele, which has similar (identical) association with RA development in DRB1 QKRAA/QRRAA epitope-negative patients. The present data demonstrate that DRB1 QKRAA/QRRAA epitope and microsatellite Bat2 138/D6S273 138 alleles more completely define the risk for development of RA. The results in the present study therefore suggest that two regions in MHC, class II (DRB1) and class III (Bat2 and D6S273 in HSP70-Bat2 region), contribute to susceptibility to RA. PMID- 10528791 TI - A nucleoside analogue, 7-thia-8-oxoguanosine stimulates proliferation of thymocytes in vitro. AB - 7-thia-8-oxoguanosine (immunosine) is a nucleoside analogue with immunoenhancing activity. In this work, its effects on proliferation of thymocytes in vitro were studied. It was found that immunosine stimulated proliferation of thymocytes both of mice and rats. The stimulatory effect depended on antigen presenting cells (APC), since thymocytes depleted of accessory cells did not proliferate to immunosine. In addition, pretreatment of APC with immunosine for 24 h significantly increased proliferation of thymocytes. Immunosine stimulated interleukin 2 (IL-2) production and the expression of activation markers (CD25 and CD71). The upregulation of CD25 (alpha subunit of IL-2R) was detected both on thymocytes and thymic dendritic cells. Proliferation of thymocytes in the presence of immunosine was predominantly mediated by IL-2 since blocking IL 2Ralpha by specific monoclonal antibodies inhibited cell proliferation by 65-85%. PMID- 10528793 TI - Atomic force microscopy to study the degranulation in rat peritoneal mast cells after activation. AB - We have studied the degranulation in rat peritoneal mast cells by atomic force microscopy (AFM). Although AFM has advantages close to electron microscopy (EM) in spatial resolution, visualization of surface topography in peritoneal mast cells has not been reported yet. In the present paper we have succeeded in visualizing the degranulation process of rat peritoneal mast cells by AFM. AFM images showed that secretory granules were about 1 microm in diameter and that they were densely packed in the cytoplasm surrounding the nucleus. After stimulation with compound 48/80 (5 microg/ml), the packed granules began to loosen through formation of irregular cracks, as if polymerization of actin filaments was changed; 3 min after stimulation with compound 48/80 at 37 degrees C, the packed granules were completely loosened and the nucleus was no longer covered by the granules. In addition, AFM images showed clear fringes at the edges of the spreading cells. The fringes were 30-50 nm in height and there existed many small matrixes which were secreted to the outside of the cell. The matrixes were about 80 nm in diameter and 40 nm in height. These kinds of secretory matrixes were observed here first by AFM. PMID- 10528794 TI - Broad clonal heterogeneity of antigen-specific CD4+ T-cells localizing at the site of disease during tuberculosis. AB - The repertoire of CD4+ T-lymphocytes was investigated in six patients affected by tuberculosis, who had a negative PPD skin test at diagnosis. Polyclonal CD4+ T cell lines from the peripheral blood failed to proliferate to PPD and to the 16- or 38-kDa proteins of Mycobacterium tuberculosis, while CD4+ T-cell lines from the site of disease responded to PPD, and to the 16- and 38-kDa proteins, and derived epitopes in vitro. The repertoire of CD4+ T-cells accumulating at the site of disease was found to be widely heterogeneous as demonstrated by the finding that at least seven different peptides from the 16- and 38-kDa proteins were recognized by every patient. These results indicate that CD4+ T-cells localized at the site of disease in tuberculosis recognize a vast array of M. tuberculosis epitopes. PMID- 10528795 TI - Are natural antibodies involved in tumour defence? AB - Natural antibodies (NAb) are found in the serum of healthy individuals. These antibodies are produced without any apparent specific antigenic stimulation. They are one part of the circulating immunoglobulins and are found in virtually all vertebrate species. NAb react to various self- and non-self antigens. A protective function in different infection models could be demonstrated. Several groups have reported the ability of NAb to bind to tumour cells. Their possible role in tumour defence is documented in mice. The present status of attempts to characterise the role of NAb in tumour defence is discussed, particularly as regards the human immune system. This paper focuses on antibody cell interactions and discusses the genetic background of the Nab-producing B-cells. PMID- 10528796 TI - Comparative studies of modulatory effect to the function of rat peritoneal neutrophils treated with new quinolones. AB - Some of the immunological effects of a variety of new quinolones on adhesion, phagocytosis, and production of reactive oxygen intermediators in neutrophils were studied. Ofloxacin, lomefloxacin, fleroxacin, and levofloxacin potentiated the phagocytosis of Escherichia coli in neutrophils. Moreover, lomefloxacin, and sparfloxacin significantly potentiated adhesion of neutrophils. In contrast, tosufloxacin was effective in significantly and persistently potentiating the production of superoxide anion, whereas the other agents markedly inhibited such production. Furthermore, tosufloxacin was effective in significantly potentiating the production of hydrogen peroxide, whereas sparfloxacin markedly inhibited such production. These results suggest that the new quinolones at a therapeutic concentration may affect functions such as phagocytosis, and production of superoxide anion in neutrophils. PMID- 10528797 TI - Regression and prevention of autochthonous tumors induced by 3-methylcholanthrene after injection of a T-cell receptor alpha /beta positive and CD4/CD8 double negative T-cells. AB - Both the therapeutic and preventative effects of a murine T-cell line, tMK-2, with T-cell receptor (TCR) alpha/beta positive and CD4-/8- double negative (DN) phenotype against autochthonously tumors induced by subcutaneous (s.c.) injection of 3-methylcholanthrene (MC) were examined. Complete regression of the tumor was observed when administration of tMK-2 cells was begun on tumors 5 mm in diameter. The tumor mass in five out of five mice was reduced in size after the administration of tMK-2 cells regardless of the routes of administration: s.c. injection of tMK-2 cells (5 x 10(7) cells) once a week around tumors, intraperitoneal (i.p.) injection (5 x 10(7) cells), or intravenous (i.v.) injection (1 x 10(7) cells). The tumors regressed to the status of a scar within 1 month of initial injection, and this status was maintained throughout the remainder of the 3 months period of tMK-2 cell injection. One month after discontinuation of tMK-2 cell administration, the diameter of the tumors had not increased regardless of the route of injection. The control groups consisted of either untreated mice, mice with i.v. injection of 1 microg of recombinant murine interleukin (IL)-12 once a week, or mice with s.c. injection of autologous splenocytes (5 x 10(7)) from BALB/c mice once a week. Continuous growth of tumors was observed in each group and all control mice died due to bleeding ulcerations of the tumors. Tumor development was effectively prevented when tMK-2 cells were administrated 1 week after the s.c. injection of MC. In the groups receiving s.c., i.p., and i.v. injection of tMK-2 cells, no MC-induced tumors developed, whereas four out of five of the control mice developed autochthonous tumors. The tMK-2 cells also exerted in vitro NK-like cytotoxic activity, and their killing activity was strongly increased in the presence of both IL-2 and IL-12. These results suggest that the injected T-cells with TCR alpha/beta positive and CD4- /8- DN phenotype and NK-like activity are important in the therapy as well as the prevention of tumor development. PMID- 10528798 TI - Cytokine production by rabbit alveolar macrophages: differences between activated and suppressor cell phenotypes. AB - The differences between cytokine-producing profiles of activated macrophages (A-M phi) and suppressor macrophages (S-M phi) were examined. A-M phi, which exhibited cytotoxicity against RK-13 cells, were generated from resident rabbit alveolar M phi by treatment with lymphokine solution (culture fluids of rabbit spleen cells stimulated with concanavalin A [Con A]). S-M phi, which were able to inhibit cellular proliferations of rabbit spleen cells stimulated with Con A, were generated from resident alveolar M phi by treatment with 1-methyladenosine (an immunosuppressive molecule in tumourous ascites fluids). When A-M phi were stimulated with lipopolysaccharide (LPS) in vitro, the cells produced significantly more interleukin (IL)-1 (approximately 1.4 times), IL-6 (approximately 2.1 times), IL-12 (approximately 60 times), and tumour necrosis factor-alpha (TNF-alpha) (approximately 37 times) than did resting macrophages (R M phi) stimulated with LPS as control cells. After the stimulation with LPS, both A-M phi and R-M phi did not produce transforming growth factor-beta (TGF-beta). In contrast, when S-M phi were stimulated with LPS in vitro, the cells produced significantly more TGF-beta (approximately 1.6 times) and significantly less IL-6 (approximately 1.8 times) than did control cells. Also, S-M phi did not produce IL-1, IL-12, and TNF-alpha into their culture fluids after the stimulation with LPS. These results show the differences between cytokine-producing profiles of A M phi and S-M phi, and characteristics of their cytokine-producing profiles are analogous to T cell subsets. Differences displayed in the cytokine profiles may contribute to the effector (A-M phi) or the suppressor (S-M phi) functions of alveolar M phi. PMID- 10528799 TI - Naturally occurring alpha-galactosyl antibodies in human sera display polyreactivity. AB - Anti-gal is a dominant autoantibody constituting nearly 1% of total circulating IgG in humans and old world primates. Raised levels of anti-gal have been demonstrated in parasitic diseases such as malaria, leishmaniasis and Chagas disease and in a variety of autoimmune diseases. It has also been implicated as a primary cause of rejection of xenogeneic cells and organs transplanted in old world primates since Gal-alpha 1,3 Gal is thought to be the major antigenic epitope to which xenoreactive natural antibodies bind. Since polyreactive antibodies have also been widely implicated in xenotransplantation and anti-gal is yet to be demonstrated to be polyreactive, we have attempted to study this property of anti-gal antibodies. Anti-gal levels were assayed in 72 human sera and compared with DNA-binding antibodies. A significant positive correlation was found between anti-gal and DNA-binding antibodies. Absorption of sera with fresh rabbit erythrocytes (which express abundant alpha-galactose on their surface) resulted in significant removal of both anti-gal and DNA-binding antibodies. Affinity purified anti-gal were found to be reactive to DNA, actin, myosin and tubulin indicating the polyreactive nature of naturally occurring anti-gal antibodies in human sera. The observed polyreactivity was not an exclusive feature of sera collected from tropical countries-anti-gal affinity purified from sera of North Americans were also found to react with DNA. The demonstration of polyreactivity of anti-gal indicates a much wider biological role for this autoantibody in humans and old world primates. PMID- 10528800 TI - Immunogenicity of the GPGRAFY epitope on synthetic peptides. PMID- 10528801 TI - Serum procalcitonin concentrations in transplant patients with acute rejection and bacterial infections. AB - Procalcitonin (PCT) represents a new marker of systemic inflammatory reactions of the body to infections. PCT is selectively induced by severe bacterial infections leading to sepsis or multiorgan dysfunction syndrome. The aim of our study was to test PCT as a postoperative infection marker in heart and kidney transplant patients compared with healthy subjects and patients with localized lung inflammatory processes without a manifest systemic response. PCT concentrations were measured by an immunoluminometric assay (ILMA) in a total of 419 serum samples. Normal serum levels were in the range of 0.08-0.6 ng/ml. Operative trauma associated with heart (not kidney) transplantation induced a transient increase in PCT levels to 7-10 ng/ml with a decline to normal levels within 2-3 days in most patients. Severe bacterial infections dramatically augmented serum PCT concentrations reaching values of 46-297 ng/ml in the most critical periods. Good response to antibiotic therapy was associated with a decline in serum PCT concentrations. Acute rejection or cytomegalovirus infections did not significantly increase the serum PCT levels. Localized pulmonary infections showed either no, or only a limited increase, in the serum PCT levels (max. 7 ng/ml). We conclude from our data that PCT can be used as a sensitive marker to differentiate systemic bacterial infections from other complications in organ transplantation. PMID- 10528802 TI - Effect of Pseudomonas aeruginosa exotoxin A on IFN-gamma synthesis: expression of costimulatory molecules on monocytes and activity of NK cells. AB - The aim of the study was (1) to evaluate the effect of Pseudomonas aeruginosa Exotoxin A (P-ExA) on the production of IFN-gamma in anti-CD3 induced human peripheral blood mononuclear cells (PBMC) and (2) to establish the effect of P ExA on the IFN-gamma dependent cellular activities such as the expression of costimulatory molecules on monocytes and cytotoxicity of NK cells. The toxin in a high dose (100 ng/ml) inhibited IFN-gamma synthesis. Inhibitory effect of P-ExA was abolished by IL-1alpha which in a combination with P-ExA exerted a strong synergistic effect on IFN-gamma synthesis. Other monokines such as IL-1beta, IL 6, TNF-alpha neither reversed the inhibitory effect of P-ExA nor induced production of IFN-gamma. P-ExA also inhibited IFN-gamma-induced cellular events: (1) expression of costimulatory molecules on monocytes (CD80, CD86, ICAM-1, HLA DR); (2) cytotoxic activity of NK cells. Inhibition of NK cells activity by P-ExA was not reversed by cytokines such as IL-2, IFN-alpha and TNF-alpha, which are known to enhance effector functions of NK cells. From these results we conclude that: (1) inhibition of IFN-gamma synthesis, as well as IFN-gamma-induced expression of costimulatory molecules and NK-cell effector functions may lead to suppression of specific and non-specific defense mechanisms, respectively, which are necessary for elimination of PA bacteria; (2) enhancement of IFN-gamma synthesis induced by P-ExA in a combination with IL-1alpha may cause harmful, Th1 cells dependent, inflammatory reactions of the host (septic shock, tissue damage) during infection with Pseudomonas aeruginosa. PMID- 10528803 TI - Modulation of the putative human immunodeficiency virus type 1 gp41 receptor expression on human T-lymphocytes by canavalin A, phytohemagglutinin and phorbol myristate acetate. PMID- 10528804 TI - The human immunoglobulin variable lambda locus IGLV9 gene is a monomorphic marker in the urban Brazilian population. AB - The physical map of the human immunoglobulin variable lambda locus (IGLV) located on chromosome 22q11.1-q11.2 shows the existence of 52 functional V-lambda genes distributed among three V-clusters. The IGLV9S1 gene, located in the V-B cluster, is a sequence tagged site and is a useful marker for restriction fragment length polymorphism (RFLP) population studies. The V-lambda genes are associated in the genome with EcoRI fragments detectable in Southern blots of genomic DNA samples. We have analysed DNA samples of an urban Brazilian population by Southern-EcoRI RFLP using an IGLV9 gene segment. Among 75 unrelated individuals analysed, we detected a single 6.0 kb EcoRI fragment containing the IGLV9 gene at 100% frequency. Reverse transcription followed by polymerase chain reaction (RT-PCR) of peripheral blood leukocyte total RNA from unrelated individuals showed that IGLV9S1 is a functional gene contributing to the B lymphocyte repertoire. These data represent evidence for monomorphism of the IGVL9S1 gene in this urban population. We demonstrate that IGLV9S1 is a functional single copy gene and is an important marker in the IGLV locus. PMID- 10528805 TI - Onset of T-cell receptor Vbeta8.1 and Dbeta2.1 V(D)J recombination during thymus development of inbred mouse strains. AB - The assembling of T-cell receptor (TCR alpha/beta and gamma/delta) genes depends on the V(D)J recombination occurring in early thymocytes during thymus ontogeny. The V(D)J recombination reaction is directed by a recombinase complex from the RAG-1 and RAG-2 genes, and is modulated by several other gene products. Due to the essential role of the TCRbeta in thymocyte differentiation, it is important to define with precision the temporal emergence of the TCRbeta recombination in normal non-manipulated mouse strains. We analysed the onset of V(D)J recombination between TCRVbeta8.1 and Jbeta2.1 gene segments during fetal development of the thymus in three non-manipulated inbred strains of mice; BALB c, C57BL/6 and CBA. We show that the emergence of the V(D)J recombination at the TCRbeta locus differs among strains, suggesting an in vivo role of the different genetic backgrounds in driving gene rearrangements. PMID- 10528807 TI - The misuse of alcohol and other drugs by doctors: a UK report and one region's response. AB - This Commentary reviews the report from the Working Group on the Misuse of Alcohol and Other Drugs by Doctors, and considers the response of a Scottish region to it. The report confirms that alcohol and drug misuse in doctors is a threat to patients and that the problem in doctors is not being addressed satisfactorily. Support for the establishment of dedicated services is outlined. Local enquiries confirmed that the reported conclusions were universally supported, but extensive work was required to incorporate the issues into student teaching, postgraduate training, continuing professional education, and future revalidation procedures. A proposal to establish a local, confidential, anonymous, informal contact point is outlined to promote early intervention and limit the development of entrenched morbidity relating to substance misuse. The intention is to monitor the utility of that arrangement by audit. PMID- 10528806 TI - Trait markers for alcoholism: clinical utility. AB - Because alcoholism is a multi-factorial psychiatric disorder, with both psychosocial and biochemical/genetic factors leading to its manifestation in any one individual, the presence of biochemical/genetic factors alone may not lead to the manifestation of the disorder. There are numerous difficulties associated with identification of a trait abnormality in a disorder that requires suitable socio-cultural permissiveness with distinct behavioural characteristics to manifest a disorder that may not require that predisposing trait abnormality in order to develop. Numerous studies have been performed in the past to potentially identify a biochemical or genetic trait abnormality in alcoholism, and not all of them have addressed significant methodological flaws in this type of research. This review addresses some of the difficulties inherent in this research, and aims for a comprehensive review of the highlights of the search for a clinically useful trait abnormality. Some series of investigations hold promise that a trait marker for a particular subset of alcoholics may be developed, e.g. severe alcoholism and the dopamine D2 receptor gene; the level of reaction to alcoholism in family history-positive alcoholics; beta-endorphin abnormalities in specific family groups of alcoholics; reduced P3 wave event-related potentials as markers and predictors of development of substance abuse in predisposed youths; reduced growth hormone response to apomorphine as a predictor of relapse to alcoholism in early abstinence; abnormal adenylyl cyclase activity in certain defined subgroups of alcoholics; and abnormal platelet monoamine oxidase levels in subjects with a behavioural predisposition to addictive disorders. The review concludes that while there has not yet been an identification of a comprehensive trait marker for alcoholism, there is hope for identification subgroups of alcoholics with consistent biological markers within that subgroup that may well prove fruitful over time. It will then be up to a future generation of clinicians to take that information and develop prevention programmes that can incorporate this information to help the predisposed individual avoid alcohol problems. PMID- 10528808 TI - Exploring young men's drinking using the AUDIT questionnaire. AB - This paper reports the findings of an AUDIT questionnaire administered to a sample of young (16-24-year-old) white males. Of the sample, 65% were drinking at potentially harmful levels. Averaging 45 U/week, 18-21 years was the age of highest alcohol consumption. Compared with 16-17- and 22-24-year-olds, this group recorded the highest proportion of hazardous drinkers and most negative consequences of drinking. PMID- 10528809 TI - Chronic alcohol consumption increases serum levels of circulating endothelial cell/leucocyte adhesion molecules E-selectin and ICAM-1. AB - A group of 30 chronic alcoholics without alcohol-related diseases and 30 controls (teetotallers) were selected to measure serum levels of endothelial adhesion molecules (AMs) (ICAM-1, VCAM-1, and E-selectin). ICAM-1 and E-selectin serum levels were higher in alcoholics, whereas VCAM-1 serum levels were similar in both groups. There was a significant correlation between daily alcohol intake and serum level of ICAM-1 (r = 0.49, P = 0.003) and E-selectin (r = 0.41, P = 0.02). A significant positive correlation between E-selectin and total lifetime dose of ethanol was also observed (r = 0.52, P = 0.003). These changes in serum levels of endothelial AMs of chronic alcoholics may reflect endothelial and/or immune activation, and could interfere with the reactions between immune cells and the endothelium. PMID- 10528810 TI - Operant responding for ethanol in rats with a long-term history of free-choice ethanol drinking. AB - Wistar rats were allowed to drink ethanol in a two-bottle (water vs 2-8% v/v ethanol) and then in a three-bottle choice paradigm (water vs 8% ethanol vs 16% ethanol, v/v). After 7 months of free access to alcohol, the subjects were trained to respond for 8% ethanol in an operant procedure. No relationship was found between prior alcohol drinking and lever pressing for ethanol. PMID- 10528811 TI - Effects of Hypericum perforatum extraction on alcohol intake in Marchigian Sardinian alcohol-preferring rats. AB - The present study investigated the effect of acute intragastric (i.g.) administration of dry Hypericum perforatum extract (HPE), containing 0.3% hypericin, on ethanol intake in genetically selected Marchigian Sardinian alcohol preferring (msP) rats. The i.g. administration of HPE, 125 or 250 mg/kg, induced a 30-40% reduction in ethanol intake in rats offered 10% (v/v) ethanol for 2 h/day. The effect of these doses was selective, since they modified neither food intake nor food-associated drinking; neither did the same doses modify the rat's gross behaviour in the open-field test. A dose of 500 mg/kg frequently induced immobility and a general suppression of ingestive behaviour. In rats offered 10% ethanol for 12 h/day, ethanol intake following treatment with 250 mg/kg HPE was significantly lower than that of controls for up to 10 h. The effect on ethanol intake was not related to the antidepressant-like effect of HPE revealed in the forced swimming test. In this regard, the effect on ethanol intake was observed after a single administration of 125 mg/kg, whereas the antidepressant effect was observed only after repeated treatment with doses higher than 125 mg/kg HPE. The i.g. administration of HPE, 250 mg/kg, did not affect blood-alcohol levels following i.g. treatment with 0.7 g/kg ethanol, the amount usually ingested in a single drinking episode; thus, the effect is not related to changes in the pharmacokinetics of ethanol. The present study shows that HPE markedly reduces ethanol intake in msP rats, without significantly modifying food intake. PMID- 10528812 TI - Attenuation of alcohol intake by extract of Hypericum perforatum (St. John's Wort) in two different strains of alcohol-preferring rats. AB - Extract of the common plant Hypericum perforatum L. (St John's Wort, SJW) has been used successfully for the treatment of mild to moderate depression since ancient times and has recently been studied clinically. Depression and alcoholism have some neurochemical similarities, such as low brain serotonin activities. Thus, we hypothesized that SJW extract, which contains 0.22% hypericin and 4.05% hyperforin, also may be effective in suppressing alcohol intake. To test this hypothesis, the effects of SJW extract on voluntary alcohol intake were studied in two different genetic animal models of human alcoholism: fawn-hooded (FH) and high-alcohol drinking (HAD) rats. FH and HAD rats received a single oral administration (5 ml/kg) of either vehicle or one of the five doses (100, 200, 400, 600, and 800 mg/kg) of SJW extract. The oral administration of SJW extract significantly (P < 0.0001) reduced alcohol intake in both FH and HAD rats. In a third study, FH rats did not develop tolerance to the suppressant effects of SJW on alcohol intake and preference following oral administration of (400 mg/kg) of the extract for 15 consecutive days. These promising findings suggest that SJW extract should be evaluated clinically as a potential therapeutic agent in the treatment of alcoholism. PMID- 10528813 TI - Effects of ageing and intermittent ethanol exposure on rat locus coeruleus and ethanol-withdrawal symptoms. AB - In this study, the effects of ethanol and age on the morphology of the locus coeruleus (LC) and on the severity of ethanol-withdrawal symptoms were studied during a 5-week intermittent ethanol exposure. Young (3-4 months) and old (29-30 months) male Wistar rats were given highly intoxicating doses of ethanol by intragastric intubations for 4 days, followed by a 3-day ethanol-withdrawal period. This 7-day cycle of ethanol exposure and withdrawal was repeated five times. A non-treated group and a sucrose-fed group of both ages were used as control groups. The severity of ethanol-withdrawal symptoms (rigidity, tremor, irritability, hypoactivity) was rated up to 62 h after the last dose of ethanol. The intoxication level was higher in the old, compared with the young, rats, despite the smaller doses of ethanol given to the old animals. There was no significant difference between the age groups in the severity of the ethanol withdrawal syndrome. The LC quantitative studies were performed using unbiased stereological methods. The results showed that there was no difference between the age groups in the LC total neuron numbers of the non-treated control groups. The 5-week intermittent ethanol exposure significantly reduced the LC neuron numbers and LC neuronal density in the old ethanol-exposed animals, compared with the sucrose-fed control animals. In the young rats, the ethanol-induced neuron loss did not reach statistical significance. According to the ANCOVA, the difference in the ethanol-induced LC neuronal loss between the age groups may be due to the difference in the intoxication levels. Interestingly, the sucrose intubations were also found to decrease the LC neuronal numbers in the young rats, compared with the non-treated young control group. It was concluded that ageing did not significantly affect the severity of ethanol-withdrawal symptoms or ethanol-induced loss of LC neurons in Wistar rats. PMID- 10528814 TI - Mental well-being in alcohol withdrawal: relationship to alpha2-adrenoceptor function. AB - The possible relationship between postsynaptic alpha2-adrenoceptor function, as assessed by the growth hormone (GH) response to clonidine (CLON) and aspects of mental well-being by self-report of mood using the Swedish Mood Adjective Check List, was investigated in alcohol-dependent patients in the early withdrawal period. Patients had blunted GH responses to CLON and worse mental well-being than control subjects immediately after the end of alcohol intake. No relation was found between mental well-being and postsynaptic alpha2-adrenoceptor function. After 1 week, the GH responses to CLON remained blunted, whereas the state of mental well-being had improved to levels similar to those of control subjects. The results further support a downregulated alpha2-adrenoceptor function during 1 week of alcohol withdrawal. Furthermore, even if subsensitive postsynaptic alpha2-adrenoceptor function was not generally related to state of mood, patients with the lowest postsynaptic alpha2-adrenoceptor function reported the highest levels in the dimensions pleasantness/unpleasantness and activation/deactivation when sober. PMID- 10528815 TI - Platelet serotonergic binding sites in alcohol-dependent patients. AB - The serotonin (5-hydroxytryptamine, 5-HT) uptake sites assessed with both [3H]imipramine and [3H]paroxetine, and the 5-HT2A receptors were simultaneously measured in platelets from 24 male subjects meeting the American Psychiatric Association's DSM-IV criteria for alcohol dependence and admitted for inpatient detoxification. Blood samples from alcoholic patients were collected during acute alcohol intoxication (day 0), during withdrawal (day 1), and after 2 weeks of abstinence (day 14). All patients met the criteria for type II alcoholism. Alcohol misuse was found to be associated with an increased number and a lower affinity of [3H]paroxetine binding in comparison to the control values. Abstinence from alcohol for 2 weeks (day 14) resulted in a decrease in the number of 5-HT uptake sites labelled with [3H]paroxetine compared to normal values, together with a significant decrease in the number of 5-HT2A binding sites. The present data indicate that altered serotonergic function existing in alcoholic patients is a reversible phenomenon that normalizes after detoxification and withdrawal. PMID- 10528816 TI - The development of alcohol consumption and problem drinking in Rotterdam 1980 1994: more problem drinking amongst the young and the middle aged. AB - In 1980/1981 and in 1994, two surveys on problem drinking were conducted in the city of Rotterdam. This article presents data on changes in alcohol consumption and alcohol-related problems between 1981 and 1994. Special attention has been paid to possible shifts in groups at risk and to shifts in the kind of problems experienced. It was found that, in 1994, compared to 1981, problem drinking had become more prevalent amongst the young and the middle aged. PMID- 10528817 TI - Frequency selective effects of alcohol on auditory detection and frequency discrimination thresholds. AB - In the first of two experiments, the effects of ethyl alcohol on monaural and binaural thresholds for pure tones were measured for a range of frequencies. The results showed a frequency-specific effect in which low frequencies were more severely affected than higher ones. Also, monaural thresholds tended to be more affected by alcohol than binaural ones. The second experiment extended this exploration by measuring frequency discrimination at several different frequencies. In this case, we also obtained a frequency-dependent effect: the increase in discrimination thresholds above 1000 Hz was three times greater than that for lower frequencies. The data suggest that the choice of stimuli may influence the ability to detect changes in auditory performance after alcohol and may account in part for the differences among earlier studies. The results are consistent with the hypothesis that alcohol is acting centrally, at the level of mechanisms involved in the temporal and binaural summation of auditory signals, rather than influencing peripheral structures. PMID- 10528818 TI - Psychopathology in alcohol withdrawal: relationship to alpha2-adrenoceptor function. AB - The possible relationship between postsynaptic alpha2-adrenoceptor function, as assessed by growth hormone (GH) response to clonidine (CLON; 1.5 or 2.0 microg/kg i.v.), and psychopathology was investigated in 30 patients with alcohol dependence in the early withdrawal period. Excluding patients with high baseline GH, 23 of the 26 patients had blunted GH responses to CLON and 57% moderate or severe depression at day 1 after the end of alcohol intake. After 1 week, the GH responses to CLON remained blunted in 20 of 21 retested patients, whereas the depression and anxiety remitted in all but two patients. The results do not support any relationship between postsynaptic alpha2-adrenoceptor function and symptoms of psychopathology in alcohol withdrawal. PMID- 10528819 TI - Influence of age, alcohol consumption and abstinence on the sensitivity of carbohydrate-deficient transferrin, gamma-glutamyltransferase and mean corpuscular volume. AB - Duration of abstinence before blood test, alcohol consumption and age was examined in 177 male alcohol-dependent patients as factors influencing serum carbohydrate-deficient transferrin (CDT), serum gamma-glutamyltransferase (GGT) and mean corpuscular volume (MCV). The strongest influence on all markers was the factor 'duration of abstinence before blood test'. In patients who had been abstinent for >4 days before the blood test, the markers had low sensitivities (GGT, 33%; CDT, 14%; MCV, 42%), whereas in patients with < or = 4 days of abstinence the markers had reasonably good sensitivities (GGT, 72%; CDT, 56%; MCV, 48%). GGT was more sensitive than CDT (P < 0.05) and MCV (P < 0.001). The combined use of CDT and GGT had sensitivity of over 90%. Mean alcohol consumption in the 30 days prior to the blood test had a significant effect on CDT and GGT, but not on MCV. Age did not have a clear effect on CDT and GGT. For MCV, a significant and linear increase with age was shown. We conclude that GGT is the most sensitive of these three markers. Using GGT and CDT combined, sensitivity can be enhanced to over 90%. The period of abstinence before the blood test has a strong influence on CDT and GGT. If a longer period of abstinence is suspected, MCV should also be measured, in order to detect evidence of earlier heavy drinking. PMID- 10528820 TI - How would you label your own drinking pattern overall? An evaluation of answers provided by 181 high functioning middle-aged men. AB - The self-rating of drinking habits was compared to DSM-III-R diagnoses of alcohol abuse and dependence in 181 men with an average age (+/- SD) of 38.7 +/- 1.91 years. Results indicate that the 150 subjects without alcohol-related diagnosis (Group 1) rated themselves as 'non-problem drinker', in categories from 'non drinker' to 'heavy drinker'. Among the 15 individuals with alcohol abuse (Group 2), none rated their drinking pattern as 'problem drinker'. Two (12.5%) subjects in the group of 16 individuals with alcohol dependence (Group 3) rated themselves as 'problem drinker', while most did not consider their drinking patterns as problematic. Within subjects who identified themselves as the same type of drinker (e.g. 'infrequent drinker', 'moderate drinker', etc ...), the quantity, frequency, and number of alcohol-related problems were higher in Groups 2 and 3, compared to Group 1. The self-rating of drinking habits using a single question failed to identify over 90% of the subjects diagnosed with alcohol use disorder (100% of those with alcohol abuse and 87.5% of those with alcohol dependence), and did not differentiate between levels of alcohol intake and number of alcohol related problems for subjects who identified as a particular drinking type. PMID- 10528821 TI - Patterns of alcohol consumption in the Seychelles Islands (Indian Ocean). AB - Self-reported drinking habits were examined in a random sample of 1067 persons aged 25-64 years in the Seychelles, a country in epidemiological transition where consumption of home-brewed, mostly unregistered beverages has been traditionally high. Alcohol consumption was calculated from respondents reporting at least one drink per week ('regular drinkers'). Among men, 51.1% were regular drinkers and had average intake of 112.1 ml alcohol a day. Among women, 5.9% were regular drinkers and had 49.7 ml alcohol a day. Frequency of drinking, but not amount per drinker, was slightly less in the 25-34-year than older-age categories. Home brews (mostly palm toddy and fermented sugar cane juice) were consumed by 52% of regular drinkers and accounted for 54% of the total alcohol intake reported by all regular drinkers. Based on the reported consumption by regular drinkers only, the average annual alcohol consumption amounted respectively to 20.7 litres and 1.2 litres per man and woman aged 25-64 years, or, using extrapolation, 13.2 litres and 0.8 litres per man and woman respectively of the total population. These values may underestimate the true figures by half, since reported beer consumption accounted for 53% of beer sales. Socio-economic status was associated strongly and inversely with home-brew consumption, but slightly and positively with consumption of commercially marketed beverages. Alcohol intake was associated with smoking, high-density lipoprotein cholesterol, carbohydrate deficient transferrin and blood pressure, but not with age and body mass index. In conclusion, these data show high alcohol consumption in the Seychelles with an important gender difference, a large proportion of alcohol derived from home brews, and opposite tendencies for the relationships between socio-economic status and home-made or commercially marketed beverages. PMID- 10528822 TI - Estimating prevalence of alcohol abuse and dependence in one general hospital: an approach to reduce sample selection bias. AB - Prevalence estimates of alcohol abuse or dependence in general hospitals are often limited to single wards, small data collecting periods or insufficient diagnostic procedures. Therefore, the present study aimed to ascertain alcohol abuse or dependence in one general hospital, to compare prevalence data for all the 11 wards and 6 intake months, to establish if screening is sufficient or if a two-step diagnostic procedure is needed, and to determine whether information for an alcohol diagnosis on suspicion is available. A sample of 1309 medical or surgical in-patients were screened by questionnaires or medication for withdrawal, and, if screening-positive, were interviewed with the alcohol section of a standardized psychiatric interview. In screening-negative patients, a diagnosis on suspicion was given if medication to treat withdrawal had been used, or if there was evidence of single criteria of alcohol dependence, somatic disorders from alcohol drinking, raised laboratory parameters on grounds of alcohol drinking or of self-reported high alcohol consumption. Of the medical and surgical in-patients, 20.7 and 16.0% respectively were alcohol abusers or dependents, with a range of prevalence rates of alcohol abuse or dependence among wards of 11.1-32.9% and among intake months between 11.3 and 28.7%. Of the medical department in-patients, 1.9%, and of the surgical in-patients, 2.1%, were screened as false-positive cases. In addition, 5.5% of the medical and 12.0% of the surgical patients were given a diagnosis on suspicion. It is concluded that all general wards and different intake months should be taken into account when estimating prevalence of alcohol abuse or dependence in a general hospital. PMID- 10528823 TI - Changes in attitudes and practices in primary health care with regard to early intervention for problem drinkers. AB - During an intervention period of 1 to 2 months, a project team supported general practitioners (GPs) and nurses in four primary health care centres in Sweden in introducing new routines for detection and treatment of problem drinkers. After the implementation of the new methods, the GPs reported increased involvement in early detection and intervention significantly more often than the nurses did. A majority in both groups reported perceived improvement in skills. There was a significant positive change of the attitudes concerning working with alcohol related problems in the nurses reaching the same level as the GPs. In the nurses, attitudes and self-perceived intervention skills were improved, but to a lesser extent than their practice. The results indicate that future efforts concerning improvement of primary health care staff involvement in alcohol interventions should focus on training, supervision, and giving positive examples, rather than on changing an already positive attitude towards alcohol intervention. The potential role of nurses is still uncertain and not utilized sufficiently. PMID- 10528824 TI - Toxicology: past, present, and future. AB - Toxicology is the study of poisons and poisoning and has an ancient and venerable history. Although there have been numerous notorious poisonings throughout the ages and rather astute descriptions of toxic agents, the scientific study of toxicology did not commence until the 19th century. There was rapid development of analytical methods in the late 19th century and then an acceleration of both method and scientific development in the latter half of the 20th century. Toxicology today can be subdivided into clinical toxicology, forensic toxicology, industrial or occupational toxicology, environmental toxicology, pharmaceutical toxicology, experimental toxicology, and workplace drug testing. The historical development of these overlapping areas of toxicology will be discussed, culminating in a prediction as to what the future may bring. PMID- 10528825 TI - Evolution of chemotherapy with platinum compounds. AB - Cancer is a major disease entity and cause of death in the human population. The discovery of cisplatin has revolutionized the chemotherapy of human cancer. The full therapeutic potential of cisplatin has not been realized due to the serious side effects and emergence of cisplatin-resistant tumor cells associated with its usage. Protective methods such as extensive hydration, improved schedules of administration, alternate routes of administration, and use of protective agents against specific side effects have allowed the use of higher doses of cisplatin against cisplatin-resistant tumors and has extended the list of tumor systems responsive to cisplatin chemotherapy. Incorporation of cisplatin into a number of cisplatin-based anti-cancer drug combinations has enhanced its effectiveness and allowed the use of lower doses of cisplatin, thus reducing its toxic side effects. Finally, the availability of cisplatin analogues, such as carboplatin and others with reduced toxicity, but increased effectiveness against cisplatin resistant tumors, has expanded the potential scope and therapeutic promise of the platinum anti-cancer agents. The evolution of chemotherapy with the platinum antitumor compounds is ongoing. PMID- 10528826 TI - Multifunctional roles of thrombin. AB - Thrombin is an unique molecule that functions both as a procoagulant and anticoagulant. In its procoagulant role it activates platelets through its receptor on the platelets. It regulates its own generation by activating coagulation factors V, VIII and even XI resulting in a burst of thrombin formation. It activates factor XI, thus preventing fibrin clots from undergoing fibrinolysis. Thrombin not only cleaves fibrinogen to fibrin, but also through the activation of factor XIII effects the cross-linking of fibrin monomers to produce a firm fibrin clot. Thrombin's role as an anticoagulant is mediated through binding to thrombomodulin, a receptor protein on the endothelial membrane of the blood vessel, initiating a series of reactions that leads to fibrinolysis. Thrombin has chemotactic properties enabling it to exert its effects during inflammation and vascular injury. It has a mitogenic effect stimulating growth of mammalian cells, fibroblasts and macrophage-like tumor cell lines. It has also been implicated in brain development. A molecule with multifunctional roles such as thrombin has its activity in vivo modulated by only a few endogenous inhibitors. PMID- 10528827 TI - Transmission electron microscopy: evaluation of damage in human oviducts caused by different surgical instruments. AB - Different wave length lasers (CO2, Nd-YAG, KTP-532), electrocautery and radiofrequency instruments were used to assess the degree of tissue damage in human oviducts. Excision of the human fallopian tube for tuboplasty is achieved most often by electrocautery, scalpel and, more recently, the Carbon Dioxide (CO2) Laser. There is not enough evidence at the present time to scientifically justify using one particular mode of incision which would show minimal damage to surrounding healthy tissue, especially heat lateral damage. The present study compares the tissue damage produced by the microelectrocautery, the CO2, the KTP 532, the Nd-YAG lasers and the radiofrequency surgical instrument, utilizing different power densities on the fallopian tubes freshly removed at the isthmic portion, taken from healthy women ages 30-42. Transmission electron microscopy sections at the cellular level show that the electrosurgical radiofrequency surgical instrument produces the least damage to surrounding healthy tissue. The CO2 laser with intermittent superpulse mode showed the second lowest amount of damage. The most damage was observed with the Nd-YAG laser at high power densities. PMID- 10528828 TI - Anionic charge sites in the kidney: comparison of two diabetic rat models. AB - The distribution and appearance of anionic charge sides (ACS) in the glomerular basement membrane (GBM) and mesangium were studied in two different animal models of diabetes mellitus (DM), the obese Zucker rat as an animal model of type 2 diabetes mellitus (DM) and streptozocin-induced Sprague-Dawley rat as an animal model of type 1 DM. Four obese Zucker rats (ZR) and four Sprague-Dawley rats were analyzed for the following parameters: number of ACS per length of lamina rara externa (LRE), (ACS/LRE); number of ACE per length of lamina rara interna (LRI) (ACS/LRI); percent of mesangial matrix as ACS (%MMACS); percent of LRE as ACS (%LREACS); percent of LRI as ACS (%LRIACS); length of ACS in LRI (LACSLRI); length of ACS in LRE (LACSLRE); width of ACS in LRI (WACSLRI); width of ACS in the LRE (WACSLRE); area of ACS in the LRI (AACSLRI); and the area of ACS in the LRE (AACSLRE). Statistical analyses include a one-way analysis of variance (ANOVA), Pearson's correlation coefficient, and stepwise multiple regression. This study confirms that a loss of ACS occurs as proteinuria develops in a variety of animal models. The majority of the ACS were more prominently localized, and thus lost form the LRE of the GBM in DN. This study also demonstrated that defined alterations in the glomerular ACS can be identified early in the evolution of DN in both animal models, and that the similarity of the changes in the ACS suggest that a common pathophysiologic mechanism induced the changes in both animal models. PMID- 10528829 TI - Clinical efficacy of the Abbott Tacrolimus II assay for the IMx. AB - Monitoring tacrolimus is essential to maintain therapeutic concentrations. Performance of the new Abbott Tacrolimus assay (FK II) was evaluated and compared to the original tacrolimus assay (FK I). 189 trough whole blood samples from transplant cases were included in the study. Samples (n = 117) with FK I concentrations > 5 ng/mL were reanalyzed with the FK II assay. Patient samples (n = 43) that had FK I concentration < 5 ng/mL with apparent mean and range of 3.1 ng/mL and 0.7 to 4.5 ng/mL, respectively, were also reanalyzed with FK II to yield a mean of 5.9 ng/mL with a range of 2.9 to 10.8 ng/mL. Checking for patient compliance, samples (n = 10) with a FK I concentration of 0 ng/mL were re analyzed. With one exception of a mislabeled cyclosporine sample, all samples (n = 9) showed FK506 levels greater than 2 ng/mL with the FK II assay. The FK II assay was shown to be a clinically efficacious assay, with improved sensitivity and acceptable precision versus the previous FK I assay. PMID- 10528830 TI - Pleomorphic large cell sarcoma of the spleen with rhabdomyosarcomatous differentiation. AB - An unusual case is reported of pleomorphic large cell sarcoma of the spleen with rhabdomyosarcomatous differentiation in a 34-year old male. According to our knowledge, such a neoplasm has never been reported in the literature. PMID- 10528831 TI - Differential diagnosis of endometrial hyperplasia and carcinoma by computerized image cytometry of cell proliferation, apoptosis and Bcl-2 expression. AB - BACKGROUND: The differential diagnosis between atypical endometrial hyperplasia and endometrial carcinoma is often difficult and based on controversial criteria. Cell kinetic parameters may be helpful. DESIGN: Cell proliferation, apoptosis and Bcl-2 expression were evaluated in benign endometrium, non atypical and atypical endometrial hyperplasia and endometrial carcinoma. The results were compared by one way analysis of variance and Bonferroni T tests. RESULTS: Cell proliferation was significantly higher (p < 0.01) in endometrial adenocarcinoma (25.6 percent) than in atypical hyperplasia (17.1 percent) and non-atypical hyperplasia (7.5 percent) of the endometrium. Apoptosis was observed in 12.3 percent of endometrial adenocarcinomas and less frequently in atypical hyperplasia (7.4 percent) and non-atypical hyperplasia of the endometrium (5.8 percent). Bcl-2 expression was significantly lower (p < 0.002) in endometrial adenocarcinoma (1.7 percent) than in atypical hyperplasia (4.2 percent) and non-atypical hyperplasia (5.3 percent) of the endometrium. In benign endometrium, cell proliferation and Bcl-2 expression were significantly higher during the proliferative phase while the rate of apoptosis was significantly higher during the secretory phase. CONCLUSIONS: Our data suggests that cell proliferation, apoptosis and Bcl-2 expression could be helpful when distinguishing endometrial carcinoma from non atypical or atypical endometrial hyperplasia. PMID- 10528832 TI - Identifying delayed separation in plasma homocysteine specimens. AB - The delayed separation of plasma from the cellular components of blood can lead to falsely elevated homocysteine results. The incidence of delayed separation in patient specimens was examined by using the ratio of arginine to the sum of arginine plus ornithine [arg/(arg + orn)], a ratio <0.50 being consistent with delayed separation. Two groups were examined: 1) low homocysteine (<10 micromol/L, n = 10), and 2) elevated (>18 micromol/L, n = 12). Specimens in the low group showed an average ratio of 0.47, with 6 of 10 <0.50. In contrast, specimens in the elevated group showed an average ratio of 0.31, with 11 of 12 <0.50. Characteristics consistent with delayed separation occurred frequently, but were significantly more frequent (p = 0.009) in the elevated group. This suggests that many homocysteine results may be falsely elevated due to preanalytic collection problems. PMID- 10528834 TI - CT-guided brain biopsy using a modified Pelorus Mark III stereotactic system: experience with 50 dogs. AB - This report describes the results of CT-guided stereotactic brain biopsies performed on 50 consecutive dogs using a modified Pelorus Mark III Stereotactic System. Based on available histopathologic samples (stereotactic biopsy [n = 50], surgery [n = 17], necropsy [n = 9]) the patient population consisted of 34 dogs with primary brain tumors, 2 with invasive nasal adenocarcinomas, and 13 with non neoplastic brain lesions. Brain tissue was not obtained from one dog. In 22 dogs a final diagnosis was made from tissue subsequently obtained from surgical resection or at necropsy. The final diagnosis was in agreement with the stereotactic biopsy diagnosis in 20 of these 22 dogs. In 17 other dogs without follow-up, stereotactic biopsy provided a diagnosis of a specific primary brain tumor subtype. Postoperative complications associated with the biopsy procedure were assessed in 41 dogs. The other 9 dogs either went directly to surgery (n = 7) or were killed (n = 2) immediately after the biopsy procedure. Thirty-six dogs recovered without apparent clinical complications. Postoperative clinical complications in the remaining 5 dogs included transient epistaxis (1 dog), transient exacerbation of cerebellar signs (1 dog), obtundation progressing to coma (1 dog), and medically uncontrollable seizures (2 dogs). The latter 3 dogs with severe neurologic complications all had large primary brain tumors and had been receiving high doses of phenobarbital and glucocorticoids to control seizures at the time of biopsy. These results suggest that this CT-guided biopsy procedure can provide an accurate pathologic diagnosis of brain lesions detected by CT and MR neuroimaging. Further refinement of both technique and case selection is expected to reduce the rate of clinical complications and to improve the accuracy of the procedure. PMID- 10528833 TI - Modification and application of a Pelorus Mark III stereotactic system for CT guided brain biopsy in 50 dogs. AB - The Pelorus Mark III Stereotactic System is a commercially available device for CT-guided stereotactic brain biopsy in people. With relatively minor modifications, this device was used to safely and accurately perform CT-guided stereotactic brain biopsies in 50 dogs with intracranial lesions. Modifications were necessary to accommodate a 90 degree shift in orientation of the canine head compared to the human head during the imaging phase of the procedure, and to facilitate other phases of the biopsy procedure that are affected by the uneven and variable topography of the canine skull. Description of a typical CT-guided brain biopsy procedure in dogs using the modified Pelorus Mark III Stereotactic System is provided. Accuracy of biopsy needle placement was determined by comparing the x, y and z coordinates of the biopsy target site with the actual coordinates of the needle tip on CT images. Mean needle placement error was 3.5 +/- 1.6 mm. Needle placement error was not significantly related to operator experience, dog size (body weight), or needle path length, however, needle placement error was significantly affected by lesion location. PMID- 10528835 TI - Computed tomographic findings in ovine coenurosis. AB - Computed tomographic imaging was conducted in twenty ewes with cerebral coenurosis. CT imaging allowed precise evaluation of the size and location of the cyst, which appeared as a hypoattenuating structure with a mass effect. No meaningful correlation between clinical signs and the location of parasitic cyst was detected. PMID- 10528836 TI - Correlative imaging findings in seven dogs and one cat with spinal arachnoid cysts. AB - Information regarding 7 dogs and 1 cat with a spinal arachnoid cyst is presented. All patients were evaluated with survey radiographs and myelography. Computed tomography (CT) following myelography, magnetic resonance (MR) imaging, and sonography, were used in some of the patients. These imaging techniques were evaluated to determine their efficacy in diagnosing arachnoid cysts, ascertaining the extent and internal cyst architecture and detecting associated spinal cord abnormalities. Survey radiographs were nondiagnostic in all patients. Myelographically, the arachnoid cyst was visible in all patients, with partial blockage to flow of contrast medium. CT provided additional information on localization and lateralization of the cyst, and allowed measurement of the degree of spinal cord compression. MR imaging enabled identification of an associated syringomyelia. Sonography was useful for defining the cyst wall and characterizing the internal architecture of the cyst cavity and adjacent spinal cord. Measurements of the degree of spinal cord compression could be made and were similar to measurements made from CT. Additionally, sonography was considered a useful technique for orientating the surgeon to the location and extent of the cyst. In the absence of the availability of CT or MR imaging for evaluating patients with an arachnoid cyst, sonography is considered a valuable technique for directly assessing the spinal cord for associated disease. Decompressive surgery was performed on 4 dogs and 1 cat, all with successful outcomes. PMID- 10528837 TI - Canine intracranial epidermoid cyst. AB - A 7-year-old intact male pitbull presented with a 2-month history of progressive dysequilibrium. Cerebrospinal fluid analysis was indicative of a central inflammatory or neoplastic disorder. A cerebellar cystic structure was identified on magnetic resonance imaging which was found to be an epidermoid cyst on histopathology. PMID- 10528838 TI - Hip dysplasia: a feline population study. AB - The study population consisted of cats presented to the University of Missouri Columbia Veterinary Medical Teaching Hospital from January 1, 1991 through December 31, 1995. Ventrodorsal radiographs including the pelvic region were evaluated for radiographic evidence of hip dysplasia. Each radiograph was evaluated independently by three board-certified veterinary radiologists and a consensus normal of dysplastic evaluation was determined. There were 684 cats from 12 breeds. The data derived from this study indicate the frequency of feline hip dysplasia in this population to be about 6.6% (45/684) and that the incidence appears to be breed dependent. Also, the radiographic appearance of hip dysplasia in cats is different than in dogs. A shallow acetabulum with remodeling and proliferation involving the cranio-dorsal acetabular margin were the most common radiographic signs. Minimal remodeling of the femoral neck was seen. PMID- 10528839 TI - Assessment of barium impregnated polyethylene spheres (BIPS) as a measure of solid-phase gastric emptying in normal dogs--comparison to scintigraphy. AB - Barium impregnated polyethylene spheres (BIPS) are radiopaque markers used for investigation of a variety of gastrointestinal disorders. One proposed use of the small (1.5 mm) marker is quantitative assessment of solid-phase gastric emptying, which may offer a simple, inexpensive alternative to nuclear medicine studies. In this study the rate and pattern of gastric emptying of a radiolabeled meal containing 30 small BIPS was evaluated in normal dogs by simultaneous comparison of the radiopaque marker method and a scintigraphic method. Serial scintigraphic images and radiographs were obtained for 8 hours or until 95% of the markers had left the stomach. Emptying curves were constructed and statistical analyses performed. There were significant differences in gastric emptying times and lag phase characteristics between the BIPS and scintigraphic studies. These results indicate that in normal dogs there are differences in both the rate and the pattern of solid-phase gastric emptying of a radiolabeled meal as assessed by scintigraphy and the gastric emptying of small BIPS. PMID- 10528840 TI - Development of reference intervals for the large intestinal transit of radiopaque markers in dogs. AB - Ten healthy dogs were fed 30 1.5 mm and 10 5 mm radiopaque markers (BIPS, MedID, Grand Rapids) mixed with sufficient quantities of a high fibre diet to meet 25% of their estimated daily caloric requirements. Abdominal radiographs were made at two hour intervals until 90% of the small and large markers had left the colon. The mean residence times (MRT) of each size of marker in the proximal, distal and total colon were calculated using kinetic analysis. The MRT's of the small markers were 4.9 hours (SD 4.4), 7.1 hours (SD 3.3) and 12.0 hours (SD 7.1) respectively. The MRT's of the large markers were not significantly different from the small markers except in the proximal colon where they were significantly shorter (3.2 hours, SD 2.3). Reference colonic filling and colonic transit curves for both sizes of markers were constructed. These may be useful to detect abnormal colonic transit in dogs. PMID- 10528841 TI - Radiographic diagnosis--gastrobronchial fistula in a dog. PMID- 10528842 TI - The dorsoproximal-dorsodistal projection of the distal carpal bones in horses: an evaluation of different beam-cassette angles. AB - To estimate the extent of the third carpal bone (C3) visible for evaluation in the dorsoproximal-dorsodistal oblique projection of the distal row of carpal bones, 13 forelimbs collected at post mortem from 7 horses were examined radiographically. The limbs were frozen with the carpal joints flexed then radiographed using fixed beam-cassette angles of 15 degrees to 45 degrees, at 5 degree intervals. The influence of beam-cassette angle on; the depth of the proximal articular surface examined, the radiographic appearance of C3 and the assessment of subchondral sclerosis was evaluated. Beam-cassette angles of 25 degrees to 40 degrees produced subjectively acceptable radiographs and did not appear to influence assessments of sclerosis. The mean depth of the examined proximal articular surface of the C3 increased significantly with each 5 degree increase in beam-cassette angle up to 40 degrees. The use of beam-cassette angles >35 degrees is recommended for the DPr-DDiO projection. PMID- 10528843 TI - Magnetic resonance imaging of the normal feline abdomen: an anatomic reference. AB - Magnetic resonance images of two adult domestic short-haired cats were obtained with a whole body scanner. Images of the abdomen were compared with cross sectional anatomy cadaver specimens from the same two cats. Anatomic structures were first identified on the cadaver specimens with the aid of anatomy texts and references and were then identified and labeled on the magnetic resonance images. Results from this project provide an atlas of normal cross-sectional MRI anatomy of the feline abdomen. PMID- 10528844 TI - M-mode echocardiographic reference values in cats in the first three months of life. AB - Thirty-five young cats were studied by echocardiography from the 2nd to 12th weeks of life to analyze correlation between body weight, body surface area, age and heart rate with fourteen echocardiographic parameters. There was a positive linear correlation (r = 0.49-0.78) between the independent variables (body weight, body surface area, age) and left ventricular wall thickness and diameter, aortic diameter and left atrial diameter, whereas there was a negative correlation (r = -0.39 and r = -0.43) between the heart rate and left ventricular diameter during systole and diastole. No linear dependence of the fractional shortening, ejection fraction, percentage thickening of the interventricular septum and left ventricular posterior wall, LA/AO ratio, and the ratio IVSED/LVWED to the independent variables was observed. PMID- 10528846 TI - Percutaneous ultrasound-guided trans-splenic catheterization of the portal vein in the dog. AB - The purpose of this investigation was to develop a safe and reliable technique for percutaneous catheterization of the portal vein via a major splenic vein using ultrasound guidance. Three separate catheter systems were evaluated on five anesthetized dogs. At least five attempts at catheterization of the splenic vein and subsequently the portal vein were attempted on each animal. Following the procedure the dogs were necropsied to assess for intrasplenic and intraabdominal hemorrhage. A technique using an introducer system and a large catheter was not successful on seven attempts. A technique using an over the needle catheter was successful in gaining access to the splenic vein on two out of five attempts; however the catheter could not be advanced into the portal vein. A technique utilizing 19 or 17 gauge needles with 22 or 19 catheter through-the-needle catheters was successful in catheterization of the splenic vein and advancement to the portal vein on twelve of fifteen attempts. The smaller gauge needle and catheter system provided for easier access to the splenic vein and subsequent catheter manipulation facilitating access to the portal circulation. PMID- 10528845 TI - Agreement between A-mode and B-mode ultrasonography in the measurement of ocular distances. AB - It is generally accepted that A-mode ultrasonography is more accurate than B-mode in measuring intraocular dimensions. Consequently, A-mode ultrasonography is the procedure of choice in ocular biometry while B-mode ultrasonography is used principally for diagnostic purposes. In this study, we investigated the agreement between measurement of intraocular distances using A- and B-mode ultrasonography on freshly enucleated camel eyes. Our results suggest that relative to average A mode values, B-mode overestimates corneal thickness (bias = 0.06 mm) and anterior chamber depth (bias = 0.03 mm), while it underestimates lens thickness (bias = 0.11 mm), vitreous chamber depth (bias = -0.32 mm) and axial length (bias = -0.40 mm). In general, difference between A- and B-mode values is larger for deeper intraocular dimensions. This implies that the two methods are more likely to give different readings for measurements of lens thickness, vitreous chamber depth and axial length. PMID- 10528847 TI - Ultrasound diagnosis: intra-abdominal wood foreign body. PMID- 10528848 TI - Palliative radiotherapy of appendicular osteosarcoma in 95 dogs. AB - Ninety-five dogs with either a presumptive (n = 24) or biopsy confirmed diagnosis (n = 71) of osteosarcoma received palliative radiotherapy using 60Co photons. Parallel opposed beams were used with each dog receiving either 10 Gy on days 0, 7 and 21 (n = 58) or 8 Gy on days 0 and 7 (n = 37). The 8 Gy fractionation scheme was given with the intent of retreating upon relapse from pain relief. Only 9 of 37 (24%) dogs in the 8 Gy group returned for retreatment. Forty-seven of the 95 dogs (49%) received concurrent or sequential chemotherapy. Seventy of the 95 dogs (74%) experienced pain relief following treatment. In dogs experiencing pain relief the median duration of response was 73 days. Numerous clinical variables were evaluated as predictors of response. The only variable significantly related to achieving a response was the use of chemotherapy. The following variables were significantly related to the duration of response: extent of bone lysis, chemotherapy use, length of bone involved and tumor site (humerus). In a multivariate analysis (n = 73 dogs), after adjusting for chemotherapy use, extent of bone involvement (p = 0.01) and tumor site (p = 0.02) retained statistical significance, while degree of bone lysis did not (p = 0.11). No difference in response incidence or duration was found between 3 fractions of 10 Gy vs. 2 fractions of 8 Gy. Administration of a low initial dose with the intent of retreatment was not a successful strategy. PMID- 10528849 TI - Images in medicine. A rare example of an autosagittogram. PMID- 10528851 TI - Closing time for CATCH22. PMID- 10528850 TI - Mutational analysis using oligonucleotide microarrays. AB - The development of inexpensive high throughput methods to identify individual DNA sequence differences is important to the future growth of medical genetics. This has become increasingly apparent as epidemiologists, pathologists, and clinical geneticists focus more attention on the molecular basis of complex multifactorial diseases. Such undertakings will rely upon genetic maps based upon newly discovered, common, single nucleotide polymorphisms. Furthermore, candidate gene approaches used in identifying disease associated genes necessitate screening large sequence blocks for changes tracking with the disease state. Even after such genes are isolated, large scale mutational analyses will often be needed for risk assessment studies to define the likely medical consequences of carrying a mutated gene. This review concentrates on the use of oligonucleotide arrays for hybridisation based comparative sequence analysis. Technological advances within the past decade have made it possible to apply this technology to many different aspects of medical genetics. These applications range from the detection and scoring of single nucleotide polymorphisms to mutational analysis of large genes. Although we discuss published scientific reports, unpublished work from the private sector could also significantly affect the future of this technology. PMID- 10528852 TI - A molecular investigation of true dominance in Huntington's disease. AB - Huntington's disease (HD) is thought to show true dominance, since subjects with two mutant alleles have been reported to have similar ages at onset of disease compared to heterozygous sibs. We have investigated this phenomenon using a cell culture model. Protein aggregate formation was used as an indicator for pathology, as intraneuronal huntingtin inclusions are associated with pathology in vitro and in vivo. We showed that cytoplasmic and nuclear aggregates are formed by constructs comprising part of exon 1 of huntingtin with 41, 51, 66, or 72 CAG repeats, in a rate that correlates with repeat number. No inclusions were seen with 21 CAG repeat constructs. Mutant and wild type huntingtin fragments can be sequestered into inclusions seeded by a mutant huntingtin. Wild type huntingtin did not enhance or interfere with protein aggregation. The rate of protein aggregation was dose dependent for all mutant constructs tested. These experiments suggested a model for the dominance observed in HD; the decrease in the age at onset of a mutant homozygote may be small compared to the variance in the age at onset for that specific repeat number in heterozygotes. Our experiments also provide a model, which may explain the different repeat size ranges seen in patients and healthy controls for the different polyglutamine diseases. PMID- 10528853 TI - A highly accurate, low cost test for BRCA1 mutations. AB - The hereditary breast and ovarian cancer syndrome is associated with a high frequency of BRCA1 mutations. However, the widespread use of BRCA1 testing has been limited to date by three principal concerns: the fear of loss of health and life insurance, the uncertain clinical value of a positive test result, and the current lack of an inexpensive and sensitive screening test for BRCA1 mutations. We have developed an inexpensive system for gene mutational scanning, based on a combination of extensive multiplex PCR amplification and two dimensional electrophoresis. The efficiency of this system, as a screening test for BRCA1 mutations, was evaluated in a panel of 60 samples from high risk women, 14 of which contained a previously identified mutation in BRCA1. All 14 mutations were identified, as well as an additional five that had previously escaped detection. In addition to the 19 mutations, a total of 15 different polymorphic variants were scored, most of which were recurring. All were confirmed by nucleotide sequencing. The cost of screening per sample was calculated to be approximately US$70 for the manual technique used in this study, and may be reduced to approximately US$10 with the introduction of commercially available PCR robotics and fluorescent imaging. Implementation of this method of mutation screening in the research and clinical setting should permit rapid accrual of quantitative data on genotype-phenotype associations for the evaluation of diagnostic testing. PMID- 10528854 TI - Two unrelated patients with inversions of the X chromosome and non-specific mental retardation: physical and transcriptional mapping of their common breakpoint region in Xq13.1. AB - Two unrelated mildly retarded males with inversions of the X chromosome and non specific mental retardation (MRX) are described. Case 1 has a pericentric inversion 46,Y,inv(X) (p11.1q13.1) and case 2 a paracentric inversion 46,Y,inv(X) (q13.1q28). Both male patients have severe learning difficulties. The same chromosomal abnormalities were found in their mothers who are intellectually normal. Fluorescence in situ hybridisation mapping showed a common area of breakage of each of the inverted chromosomes in Xq13.1 near DXS131 and DXS162. A detailed long range restriction map of the breakpoint region was constructed using YAC, PAC, and cosmid clones. We show that the two inverted chromosomes break within a short 250 kb region. Moreover, a group of ESTs corresponding to an as yet uncharacterised gene was mapped to the same critical interval. We hypothesise that the common inversion breakpoint region of the two cases in Xq13.1 may contain a new MRX gene. PMID- 10528855 TI - X linked severe mental retardation, craniofacial dysmorphology, epilepsy, ophthalmoplegia, and cerebellar atrophy in a large South African kindred is localised to Xq24-q27. AB - To date over 150 X linked mental retardation (XLMR) conditions have been documented. We describe a five generation South African family with XLMR, comprising 16 affected males and 10 carrier females. The clinical features common to the 16 males included profound mental retardation (100%), mutism despite apparently normal hearing (100%), grand mal epilepsy (87.5%), and limited life expectancy (68.8%). Of the four affected males examined, all had mild craniofacial dysmorphology and three were noted to have bilateral ophthalmoplegia and truncal ataxia. Three of 10 obligate female carriers had mild mental retardation. Cerebellar and brain stem atrophy was shown by cranial imaging and postmortem examination. Linkage analysis shows the gene to be located between markers DXS424 (Xq24) and DXS548 (Xq27.3), with a maximum two point lod score of 3.10. PMID- 10528856 TI - Microdeletions in FMR2 may be a significant cause of premature ovarian failure. AB - Genetic causes of premature ovarian failure (POF) include X chromosome deletions and fragile X (FRAXA) premutations. While screening a cohort of women with POF for FRAXA premutations, a more distal trinucleotide repeat, FRAXE, was also tested. We found an unexpected excess of FRAXE alleles with apparently fewer than 11 repeats in the POF group. However, sequence analysis of these alleles showed that the excess was caused by three females who carry cryptic deletions in FMR2, the gene associated with FRAXE. We propose that microdeletions within FMR2 may be a significant cause of premature ovarian failure, being found in 1.5% of women with the condition, and in only 0.04% of the general female population. The deletions may affect transcription of either FMR2 or an adjacent gene. PMID- 10528857 TI - Specific polymorphisms in the RET proto-oncogene are over-represented in patients with Hirschsprung disease and may represent loci modifying phenotypic expression. AB - Hirschsprung disease (HSCR) is a common genetic disorder presenting with functional intestinal obstruction secondary to enteric aganglionosis. HSCR can be familial or sporadic. Although five putative susceptibility genes have been identified, only germline mutations in the RET proto-oncogene account for a significant minority (up to 50%) of familial HSCR; 3% of sporadic HSCR in a population based series carry RET mutations. From 1998 to February 1999, we prospectively ascertained 64 cases of sporadic HSCR from the western Andalusia region. To determine if polymorphic sequence variants within RET could act as low penetrance predisposing alleles, we examined allelic frequencies at seven polymorphic loci in this population based series. Whether allele frequencies differed from those in the control population were determined by either chi squared analysis or Fisher's exact test. For two sequence variants, A45A (c 135G- >A) (exon 2) and L769L (c 2307T-->G) (exon 13), the rarer polymorphic allele was over-represented among HSCR cases versus controls (p<0.0006). In contrast, two other polymorphisms, G691S (c 2071C-->A) (exon 11) and S904S (c 2712C-->G) (exon 15), were under-represented in the HSCR patients compared to controls (p=0.02). Polymorphisms in the RET proto-oncogene appear to predispose to HSCR in a complex, low penetrance fashion and may also modify phenotypic expression. PMID- 10528858 TI - Unreported RSK2 missense mutation in two male sibs with an unusually mild form of Coffin-Lowry syndrome. AB - An unreported missense mutation of the ribosomal S6 kinase 2 (RSK2) gene has been identified in two male sibs with a mild form of Coffin-Lowry syndrome (CLS) inherited from their healthy mother. They exhibit transient severe hypotonia, macrocephaly, delay in closure of the fontanelles, normal gait, and mild mental retardation, associated in the first sib with transient autistic behaviour. Some dysmorphic features of CLS (in particular forearm fullness and tapering fingers) and many atypical findings (some of which were reminiscent of FG syndrome) were observed as well. The moderate phenotypic expression of this mutation extends the CLS phenotype to include less severe mental retardation and minor, hitherto unreported signs. The missense mutation identified may be less deleterious than those previously described. As this mutation occurs in a protein domain with no predicted function, it could be responsible for a conformational change affecting the protein catalytic function, since a non-polar amino acid is replaced by a charged residue. PMID- 10528859 TI - Defective PEX gene products correlate with the protein import, biochemical abnormalities, and phenotypic heterogeneity in peroxisome biogenesis disorders. AB - Peroxisome biogenesis disorders (PBD) comprise three phenotypes including Zellweger syndrome (ZS) (the most severe), neonatal adrenoleucodystrophy, and infantile Refsum disease (IRD) (the most mild), and can be classified into at least 12 genetic complementation groups, which are not predictive of the phenotypes. Several pathogenic genes for PBD groups have been identified, but the relationship between the defective gene products and phenotypic heterogeneity has remained unclear. We identified a mutation in the PEX2 gene in an IRD patient with compound heterozygosity for a missense mutation and the known nonsense mutation detected in ZS patients. In transfection experiments using the peroxisome deficient CHO mutant, Z65 with a nonsense mutation in the PEX2 gene, we noted the E55K mutation had mosaic activities of peroxisomal protein import machinery and residual activities of peroxisomal functions, including dihydroxyacetone phosphate acyltransferase and beta oxidation of very long chain fatty acids. The nonsense mutation severely affects these peroxisomal functions as well as the protein import. These data suggest that allelic heterogeneity of the PEX gene affects the peroxisomal protein import and functions and regulates the clinical severity in PBD. PMID- 10528860 TI - Maternal uniparental disomy for chromosome 14 in a boy with a normal karyotype. AB - We report on a boy with a maternal uniparental disomy for chromosome 14 (UPD(14)). At 7 years of age he was referred to us by the paediatrician because of symptoms of Prader-Willi syndrome (PWS). He showed short stature, obesity, mild developmental delay, cryptorchidism, and some mild dysmorphic features. The history further indicated intrauterine growth retardation at the end of the pregnancy. His mother was 44 years of age at the time of his birth. After birth he showed hypotonia with poor sucking, for which gavage feeding was needed. Motor development was delayed. After 1 year he became obese despite a normal appetite. Recurrent middle ear infections, a high pain threshold, and a great skill with jigsaw puzzles were reported. There were no behavioural problems or sleep disturbance. Chromosomal analysis was normal (46,XY). DNA analysis for Prader Willi syndrome showed no abnormalities. Two years later he was re-examined because we thought his features fitted the PWS-like phenotype associated with maternal UPD(14). At that time precocious puberty was evident. DNA analysis showed maternal heterodisomy for chromosome 14. In all the previously described 11 cases with maternal UPD(14), a Robertsonian translocation involving chromosome 14 was detected cytogenetically before DNA analysis. This is the first report of diagnosis of maternal UPD(14) based on clinical features. This finding underlines the importance of DNA analysis for maternal UPD(14) in patients with a similar PWS-like phenotype even without previous identification of a Robertsonian translocation involving chromosome 14. PMID- 10528861 TI - Schimke immuno-osseous dysplasia: case report and review of 25 patients. AB - Immuno-osseous dysplasia is characterised by spondyloepiphyseal dysplasia, lymphopenia with defective cellular immunity, and progressive renal disease. We describe a patient with a severe form of the disease, review the features of another 24 patients, and discuss the previous classification. The differences between the two groups are not striking, and although similarities are greater between affected sibs, the same diagnosis of Schimke immuno-osseous dysplasia should apply to them all. The aetiology and physiopathology of this rare osteochondrodysplasia of presumed autosomal recessive inheritance remain unknown. PMID- 10528863 TI - A PCR test for the detection of hypermethylated alleles at the retinoblastoma locus. PMID- 10528864 TI - Frequency and predictive value of 22q11 deletion. PMID- 10528862 TI - A missense mutation in both hMSH2 and APC in an Ashkenazi Jewish HNPCC kindred: implications for clinical screening. PMID- 10528865 TI - Cardiac and skeletal actin gene mutations are not a common cause of dilated cardiomyopathy. PMID- 10528866 TI - Subclinical cognitive impairment in autosomal dominant "pure" hereditary spastic paraplegia. PMID- 10528867 TI - Acute lymphoblastic leukaemia in a patient with cardiofaciocutaneous syndrome. PMID- 10528868 TI - Molecular detection of enteroviruses from an outbreak of hand, foot and mouth disease in Malaysia in 1997. AB - Enterovirus 5'UTR sequences were detected by RT-PCR in 22 out of 47 suspected hand, foot and mouth disease (HFMD) patients during an outbreak of the disease with incidences of fatal brainstem encephalomyelitis in Malaysia in 1997. Genetic and phylogenetic analyses of the isolates 5'UTR sequences suggest the presence of predominantly enteroviruses with high sequence similarities to Echovirus 1 and Coxsackievirus A9 in the Malaysian peninsula. No fatal cases, however, were associated with these isolates. The remaining isolates, including all (4/4) isolates of the fatal cases from the Malaysian peninsula and Sarawak shared very high sequence identity with enterovirus 71MS (EV71). These findings suggest that several enteroviruses were circulating in Malaysia during the outbreak period, with only EV71 causing fatal infections. PMID- 10528869 TI - Long-term pattern of HIV-1 RNA load in perinatally infected children. AB - The objective of this study was to describe the natural history of HIV-1 RNA load in vertically HIV-1-infected children. HIV-1 RNA in 156 plasma or serum samples (1-14, median 4 from each child) from 32 vertically HIV-1-infected children was detected with the NASBA technique (Organon Teknika, The Netherlands). Twenty-one children were prospectively followed from birth, and 11 were identified and included at the age of 7-89 (median 61) months. The highest numbers of HIV-1 RNA copies were seen at 1.5-3 months of age. A quadratic curve model showed a reduction of HIV-1 RNA with increasing age up to approximately 8 years, and thereafter increasing numbers, p(age) = 0.002, p(age2) = 0.008. This pattern was not typical for individual children in whom a great variation in HIV-1 RNA numbers was seen over time. The interval from birth to the first HIV-1 RNA peak ranged from 1.5 months to more than 2 years. The HIV-1 RNA levels remained relatively high and fluctuating over the years in symptomatic as well as in long term asymptomatic children. This makes HIV-1 RNA determination in children more difficult to use than in adults, as the only tool for prediction of disease progression and for initiation of therapy. PMID- 10528870 TI - Low frequency of the ccr5delta32 HIV-resistance allele in mainland China: identification of the first case of ccr5delta32 mutation in the Chinese population. AB - A 32-bp deletion on the CCR5 gene (ccr5delta32) confers resistance to HIV-1 infection. This deletion is common in Caucasians, but rare in Asians. Since the frequency of the ccr5delta32 allele of Chinese in mainland China has been unknown we investigated the ccr5delta32 mutation in a cohort of 407 Chinese people in this area. A 225-bp fragment of CCR5 encompassing the 32-bp region was analysed by PCR, hybridization and sequencing. Only 1 out of 407 subjects was heterozygous for ccr5delta32 and no homozygotes were detected. The frequency of ccr5delta32 in this cohort is thus 0.00123, i.e. much lower than that of Caucasians. The ccr5delta32 heterozygote is a healthy young man. To our knowledge this is the first ccr5delta32 mutant found in Chinese people. The results indicate that ccr5delta32 does exist in Chinese people, but at very low frequency. This suggests that ccr5delta32 is not a significant factor for the genetic resistance to HIV-1 in Chinese people. PMID- 10528871 TI - SPECT with 99mTc-HMPAO in subjects with HIV infection: cognitive dysfunction correlates with high uptake. AB - We prospectively studied a cohort of 25 HIV-1 infected individuals with no clinical signs of encephalopathy with 99mTc-HMPAO-SPECT. The findings were correlated with magnetic resonance imaging (MRI), neuropsychological testing and clinical staging aiming at the early diagnosis of HIV encephalopathy by single photon emission computed tomography (SPECT). A total of 25 matched seronegative controls were subject to neuropsychological testing only. A total of 24 patients and controls were monitored for 6-46 months (mean and median 26 months). No patients developed AIDS dementia complex during the study; 3 patients developed minimal symptoms (MSK classification stage 0.5). There was a significant decline in 99mTc-HMPAO uptake over time and neuropsychological abnormalities progressed. Unexpectedly, there was a correlation of high cortical and subcortical 99mTc HMPAO uptake and low performance in cognitive dysfunction tests, indicating a possible inflammatory reaction in the brain with increased blood flow due to HIV infection. We conclude that, in non-demented HIV-infected individuals, both the 99mTc-HMPAO uptake and functional level slowly decrease over time, but the regional cerebral blood flow decrease could be masked by a direct HIV-induced inflammatory reaction in the brain, which gives a 99mTc-HMPAO hyperfixation. PMID- 10528872 TI - Chronic hepatitis C in Sweden: genotype distribution over time in different epidemiological settings. AB - Hepatitis C virus (HCV) strains are divided into 6 genotypes and several subtypes. Recent studies reported a change in the relative frequency of genotypes within certain regions. We studied the HCV genotype in 312 Swedish patients with chronic hepatitis C, using a core region primer-specific PCR, and grouped the patients according to parenteral risk factors. The date of infection could be estimated in 127 cases. Genotypes 1a (35%) and 3 (31%) were the most common genotypes, followed by genotype 2 (17%), while only 6% had genotype 1b. Genotype 3 was relatively more frequent among subjects infected sexually or by intravenous drug use. The genotype distribution was different from that in studies from other parts of the world, with a lower frequency of genotype 1 (especially 1b) and a higher frequency of genotype 3. The frequency of genotype 1b has decreased and genotype 3 increased over time. The reasons for a different distribution of genotypes in Sweden, compared with other countries, might be a relatively recent introduction of HCV into the population, or a different pattern of transmission. PMID- 10528873 TI - Long-term re-treatment with interferon and ribavirin combination therapy in patients with chronic hepatitis C who are non-responders to interferon alone: a preliminary study. AB - We evaluated the efficacy and tolerance of ribavirin and IFN-alpha combination therapy over 12 months in 28 patients who were non-responders to IFN-alpha alone. Of 24 patients who have finished the therapy, 6 (25%) obtained a complete response (normal ALT and negative HCV RNA) at the end of treatment and maintained a sustained response 27% (5/18). PMID- 10528874 TI - Lack of association between mannose-binding lectin, acute otitis media and early Epstein-Barr virus infection among children in Greenland. AB - Low serum levels of mannose-binding lectin (MBL) have been associated with recurrent infections in early childhood. Otitis media (OM) is frequent in Greenlandic children and the first episode of acute OM (AOM) occurs early, as is the case also with Epstein-Barr virus (EBV) infection. We have therefore investigated the association between MBL genotypes, episodes of AOM, and early EBV infection in 82 community-based, unselected children in Greenland. Nasopharyngeal aspirations for EBV and MBL genotype examination, nasopharyngeal bacterial cultures, and history of AOM episodes were obtained. MBL genotypes were established in 73 specimens: 68% of these were homozygous for normal wildtype (AA), and 32% were homozygous or heterozygous for variant alleles that are associated with absence or low MBL serum level. The allele frequencies were: A = 0.88, B = 0.08 (codon 54) and D = 0.04 (codon 52). EBV was found in 41 specimens, more often with increasing age, and significantly related to ethnicity. Presence of variant MBL alleles or EBV infection was not associated with AOM, recurrent AOM (rAOM) or age at first AOM episode and EBV positive children with homozygosity for the normal MBL genotype did not have significantly more episodes of AOM, rAOM or earlier age at the first AOM episode. MBL genotypes and EBV infection alone or in interplay are not associated with the high prevalence of OM in Greenlandic children. The study suggests that low MBL level does not by itself predispose to AOM in community-based, unselected children. PMID- 10528875 TI - Lower lipoteichoic and teichoic acid CSF concentrations during treatment of pneumococcal meningitis with non-bacteriolytic antibiotics than with ceftriaxone. AB - In the rabbit model of Streptococcus pneumoniae meningitis, treatment with rifabutin, quinupristin-dalfopristin, moxifloxacin and trovafloxacin led to smaller increases of the CSF concentrations of the pro-inflammatory cell wall components lipoteichoic and teichoic acids (LTA and TA) than did treatment with ceftriaxone. Low doses of moxifloxacin were associated with higher LTA and TA concentrations in CSF than were high doses. PMID- 10528876 TI - Leuconostoc species: a case-cluster hospital infection. AB - Leuconostoc species are members of the Streptococcacae family. They are generally regarded as non-pathogenic culture contaminants and are thought to be an uncommon cause of infection. We present a study of a case-cluster nosocomial infection due to Leuconostoc spp. Three patients were hospitalized at the time of the infection with significant underlying diseases and all had a compromised skin and mucous barriers. Two had received previous antibiotic therapy. This report highlights the importance of Leuconostoc spp. as an emerging pathogen, even though the modes of transmission and reservoirs of Leuconostoc spp. are as yet unknown. PMID- 10528877 TI - Meningitis caused by streptococci other than Streptococcus pneumoniae: a retrospective clinical study. AB - We reviewed the medical records of 26 patients (median age 62 years, range 5-76 years) admitted to our institution during 1978-98 with acute bacterial meningitis (ABM) caused by streptococci other than Streptococcus pneumoniae (comprising 1.9% of all patients with ABM). 19 cases were community-acquired and 7 were nosocomial. 73% had comorbid or predisposing conditions and 73% had an identifiable extracerebral focus; only in 2 patients no comorbid disease, primary focus or predisposing condition was present. Five patients had cerebral abscesses, and 5 had endocarditis. Beta-haemolytic streptococci were grown in 14 cases (serotype A: 4, B: 5, C: 1, G: 4) and were predominant among patients with endocarditis, whereas alpha- or non-haemolytic strains grew in 12 cases (S. mitis: 4, S. constellatus: 2, E. faecalis: 2, S. bovis: 1, unspecified: 3) and were predominant in patients with a brain abscess. Staphylococcus aureus grew together with a streptococcus in 2 cases. Blood culture was positive in 9 cases (35%). Neurologic complications occurred in 11 patients (42%) and extraneurologic complications in 18 patients (69%). Adverse outcomes occurred in 10 patients (38%), including 3 patients who died. Occurrence of seizures at any time of disease was significantly associated with an adverse outcome; no other clinical or paraclinical features appeared to affect outcome. PMID- 10528878 TI - Free-living amoebae protecting Legionella in water: the tip of an iceberg? AB - Bacteria are a main food source for free-living amoebae inhabiting aquatic systems. Some bacteria however, have the ability to prevent intracellular destruction and can survive and grow in amoebic cells as endosymbionts. Free living amoebae are well adapted to their hostile environmental conditions and are resistant to both desiccation, elevated temperatures and various disinfectants. For their endosymbionts, amoebae represent perfect vectors, providing both protection against adverse environmental conditions and transportation. There is increasing interest in the potential role of free-living amoebae as reservoirs and vectors of pathogenic bacteria. The best known of such pathogenic bacteria is Legionella, and several studies provide evidence for the importance of the amoeba bacterium relationship in the biology and epidemiology of pneumonia caused by this pathogen. Although the relative importance of endosymbiosis of this kind is unknown when it comes to other human bacterial infections and the exact role of amoebic hosts in bacterial survival, multiplication and transmission in the environment is still poorly understood, naming free-living amoebae the "Trojan horses" of the microbial world is appropriate. PMID- 10528879 TI - Clinical presentation of tuberculosis and the degree of immunodeficiency in patients with HIV infection. AB - A retrospective study of 80 HIV patients diagnosed with tuberculosis was carried out in order to evaluate the clinical presentation of tuberculosis (CPT) in relation to the degree of HIV-induced immunodeficiency, as determined by the CD4 lymphocyte count and reactivity to the tuberculin and delayed-type hypersensitivity reaction (DHR) skin tests by applying 2TU of RT-23 tuberculin, and the Muiltest IMC Merieux, respectively. CPT classification was undertaken on the basis of the location of the disease: pulmonary tuberculosis (PT), distinguishing between typical pulmonary (TP) and atypical (AP) according to the radiological pattern; extrapulmonary (ET); mixed tuberculosis (MT) pulmonary and extrapulmonary; and miliary tuberculosis. The CD4 lymphocyte count was 264.6 +/- 226.8, the TP had the highest number (505), MT had 132 (p < 0.001) and the miliary tuberculosis had 148 (p < 0.001), the lowest. The tuberculin skin test was positive in 35%, of which 11% were MT (p < 0.05). In the delayed-type hypersensitivity reaction, 67% were non-normoergic, of which 95% and 100% were MT and miliary tuberculosis, respectively (p < 0.05). There was a good overlap between CD4 depletion and skin tests. TP exhibited moderate immunodeficiency, AP severe immunodeficiency, and mixed and miliary TB extremely high immunodeficiency. PMID- 10528880 TI - Evaluation of IS1245-based PCR for detection of mycobacterium avium bacteraemia in AIDS patients. AB - A PCR method based on the repetitive IS1245 sequence was evaluated for the detection of Mycobacterium avium bacteraemia in AIDS patients. Two blood preparation methods were applied: lysis of erythrocytes using a hypotonic buffer and Ficoll density centrifugation. Results were compared with culture and PCR amplification of the non-repetitive pMav22 sequence. Fifty-one of 251 tested samples grew M. avium. Bacterial densities lower than 10 cfu/ml blood were frequent, they occurred in 59% of blood samples from patients with mycobacteraemia. Inhibitory substances were detected more frequently with the lysis method (36%) than the Ficoll processed samples (19%). While specificity of PCR was high, sensitivities varied according to the methods used and the load of infection in the bloodstream. False-negative PCR results were attributable to low level bacteraemia and inhibition of PCR. Moreover, 1 of 186 and 9 of 51 M. avium strains investigated lacked the IS1245 and the pMav22 sequence, respectively. Sensitivities of culture, IS1245- and pMav22-based PCR were 88, 64 and 58%, within the lysis processed samples and 95, 86 and 50% using Ficoll prepared samples, respectively. Thus, we conclude that IS1245-based PCR is a rapid and specific method for diagnosing M. avium bacteraemia and shows a higher sensitivity than single copy gene targets, but that the sensitivity is inferior to culture. PMID- 10528881 TI - Antibiotic susceptibility of staphylococci isolated in blood cultures in relation to antibiotic consumption in hospital wards. AB - A total of 510 isolates of Micrococcaceae, 500 of staphylococci and 10 micrococci, detected in 485 (3.3%) of 14,860 consecutive blood cultures obtained from patients at a Swedish university hospital and 2 local hospitals were identified to species level and investigated for antibiotic susceptibility. The 5 most frequently isolated species were Staphylococcus epidermidis (54.8%), S. aureus (28.0%), S. hominis (3.4%), S. warneri (3.2%) and S. haemolyticus (2.8%). All isolates of S. aureus were oxacillin sensitive. Great diversity in antibiotic resistance among coagulase negative staphylococci between hospitals and different ward units in the university hospital was observed. The frequency of antimicrobial resistance among S. epidermidis correlated with the antibiotic consumption at different ward units, in particular for ciprofloxacin (p < 0.001) and co-trimoxazole (p < 0.004). The study emphasizes the importance of monitoring antibiotic consumption and resistance patterns of nosocomial staphylococci in order to avoid emergence and spread of multi-resistant bacteria within the hospital environment. PMID- 10528882 TI - Plasma nitrate as an index of nitric oxide formation in patients with acute infectious diseases. AB - In humans, the role of nitric oxide (NO) in host defence is controversial. We prospectively studied plasma levels of nitrate, the stable end-product of NO formation, during acute infection in 43 patients controlled with regard to dietary nitrate/nitrite. During acute gastroenteritis the mean plasma nitrate level was significantly increased compared with at recovery 4-5 weeks later (118 vs. 32.5 micromol/l; p < 0.001), in contrast with the findings in patients with acute pneumonia (PN; 34.6 vs. 42.8 micromol/l) or febrile urinary tract infection (UTI; 27.7 vs. 31.3 micromol/l). In a second group of 20 retrospectively studied patients with severe PN or UTI, of whom 70% were bacteraemic, no significantly increased nitrate levels could be demonstrated during the acute stage of infection. These findings indicate that increased NO production, as measured by plasma nitrate, is not a general finding in patients with acute infectious diseases, but may rather be associated with certain pathogens or sites of infection. PMID- 10528883 TI - Recurrent endocarditis caused by Streptococcus pneumoniae. AB - Pneumococcal endocarditis most often presents as an ulcerative endocarditis causing rapid destruction of the normal aortic valve, leading to aortic insufficiency and acute heart failure. Alcoholism is the most frequent underlying medical condition. This case illustrates that pneumococcal endocarditis can reoccur and is able to attack healthy, as well as previously damaged, heart valves. It also illustrates that vaccination of certain groups should be considered. The importance of repeated heart stethoscopy in patients with pneumococcaemia is emphasized. PMID- 10528884 TI - Whipple's disease confined to the central nervous system: case report and review of the literature. AB - Whipple's disease confined exclusively to the CNS without systemic involvement appears to be very rare, with only 8 cases reported in the literature. We present here a further case of primary cerebral Whipple's disease in which the neurological symptoms were seen in the absence of systemic involvement and emphasize the importance of diagnosing this treatable disease. PMID- 10528885 TI - Pancreatic infection with Candida parapsilosis. AB - Candida species other than C. albicans have been implicated as pathogens in intravascular (bloodstream, intravascular devices, endocarditis) and extravascular (arthritis, osteomielitis, endophtalmitis) infections. C. parapsilosis, however, is rarely implicated in intra-abdominal infections (peritonitis during peritoneal dialysis, complicating surgery or solid-organ transplantation). We describe a case of a 48-y-old male with acute pancreatitis who had a pancreatic abscess produced by primary C. parapsilosis infection. Although he received adequate treatment with antifungal medication and surgical drainage, the outcome was fatal. Because the clinical findings are indistinguishable from bacterial abscesses, Candida species should be considered in cases of complicated pancreatitis, in order to establish a prompt adequate treatment. PMID- 10528886 TI - Tuberculous cellulitis in a child demonstrated by magnetic resonance imaging. AB - The increasing prevalence of extrapulmonary tuberculosis means that it is important for clinicians to review their knowledge of unusual presentations of mycobacterial infections. Involvement of subcutaneous tissue and skeletal muscle is rare in tuberculosis. Occasionally, infection of soft tissue may be the sole manifestation of tuberculosis. Apart from cases of tuberculous lymphadenitis, the diagnosis of extrapulmonary tuberculosis may be difficult. Modern imaging techniques, such as magnetic resonance imaging, may be helpful in making a differential diagnosis. We present here a case of tuberculous cellulitis in an immunocompetent child and discuss the contribution of MRI in diagnosis. PMID- 10528887 TI - Seroprevalence of tick-borne Lyme borreliosis in a defined population in Estonia. AB - Sera from 200 randomly selected individuals living in Karksi Nuia, south Estonia, near an area endemic for tick-borne encephalitis and Lyme borreliosis (LB), were tested for antibodies to Borrelia burgdorferi. Antibodies were detected by enzyme linked immunosorbent assay in 6 individuals (3%; 95% CI: 1-5%), who were middle aged, asymptomatic anti-nuclear and anti-smooth muscle antibody negative. Our data show that there is low seroprevalence rate of antibodies to B. burgdorferi in an unselected south Estonian population. PMID- 10528888 TI - Primary HIV infection presenting with acute pancreatitis. PMID- 10528889 TI - Etiology of multiple myeloma: what's new. AB - Both the tumor cells and the microenvironment of dendritic stromal cells appear to be involved in the etiology of multiple myeloma. Switch translocations in myeloma tumor cells often involve oncogenes. These translocations have a clearly established role in the etiology of lymphoma and may prove to have a role in the transformation process of myeloma. Dendritic stromal cells infected with human herpesvirus-8 may provide a growth and antiapoptosis advantage for myeloma bone marrow stromal cells via viral interferon regulatory factor expression. In addition, increased vascular endothelial growth factor expression secondary to viral interleukin-8 receptor gene expression stimulates angiogenesis and inhibits development of uninfected dendritic cells, providing an advantage to infected dendritic cells. These recent advances in the understanding of the etiology of multiple myeloma provide potential new genetic, viral, and cytokine targets for therapy of this fatal malignancy. PMID- 10528890 TI - Advances in the biology of multiple myeloma: therapeutic applications. AB - Recent advances in the biology of multiple myeloma cell growth and survival have suggested new avenues for treatment and potential cure of this disease. Adhesion molecules on the myeloma cell surface mediate their localization in the bone marrow via binding to extracellular matrix proteins and stromal cells. Stromal cell to tumor cell contact and the secretion of transforming growth factor by tumor cells triggers interleukin-6 secretion from stromal cells and paracrine tumor cell growth. CD40 activation of myeloma cells changes their cell surface phenotype, triggers autocrine interleukin-6 secretion, and can regulate myeloma cell cycle in a p53-dependent fashion. Interleukin-6 is both a growth and survival factor for myeloma cells, and delineation of the signaling cascades mediating its effects permits the development of novel therapies either to interrupt growth or trigger apoptosis. New immune therapies offer the opportunity to treat minimal residual disease after stem cell transplantation, thereby improving outcome. Selected donor lymphocyte infusions after allografting and infusion of activated autologous T cells following autografting are examples of adoptive immunotherapy. Myeloma cell to dendritic cell fusions have been used in a vaccination strategy both to prevent and treat myeloma in an animal model, providing the rationale for similar clinical trials in humans. For the first time, a variety of novel treatment strategies derived from advances in understanding the disease pathogenesis offer the potential to achieve long-term disease-free survival in patients with multiple myeloma. PMID- 10528892 TI - Novel approaches in myeloma therapy. AB - High-dose melphalan (200 mg/m2) followed by one or more autologous peripheral blood stem cell transplantations is a safe and effective treatment regimen for multiple myeloma. This treatment regimen is as effective as standard therapy for myeloma in older (>65 years) patients and in patients with renal failure. However, advanced age (>50 years), duration of prior standard therapy (> 12 months), and a low CD34 mobilization potential (<20 x 10(6)/kg) are associated with a higher incidence of cytogenetic myelodysplasia. Future efforts directed at curing multiple myeloma should incorporate the best remission induction regimens presently available and should use consolidation/maintenance treatment (eg, idiotype/dendritic cell vaccination and dexamethasone/cyclophosphamide/etoposide/cisplatin combination chemotherapy) to enhance sustained complete remission. Other options to improve the treatment of myeloma include novel adjunctive therapies that target the myeloma cell microenvironment (eg, bisphosphonates, thalidomide, other antiangiogenesis agents) and allogeneic transplantation techniques to induce a graft-versus myeloma effect. PMID- 10528891 TI - Drug resistance in multiple myeloma: approaches to circumvention. AB - Resistance mechanisms to chemotherapy in multiple myeloma include (1) reduced drug concentrations at the target site of action, (2) alterations in the drug target, and (3) inhibition or prevention of drug-induced apoptosis. Recent advances in understanding resistance mechanisms have resulted in the investigation of novel therapies for the treatment of patients with multiple myeloma. P-glycoprotein is a drug transport protein that decreases intracellular drug concentrations at the target site. Valspodar, a third-generation cyclosporine analog, is an inhibitor of P-glycoprotein that currently is being evaluated to potentially overcome this mechanism of drug resistance. P glycoprotein inhibitors (also known as chemosensitizers) are being investigated for use in combination with chemotherapeutic agents to enhance the apoptotic effect and prevent resistance at the target site. Other novel approaches involve blocking pathways that result in the expression of antiapoptosis factors. Interleukin-6 is an important growth factor in myeloma and has been implicated in drug resistance via an antiapoptosis effect. In vitro blocking of an interleukin 6-dependent pathway with either a JAK inhibitor (tyrphostin, AG490) or STAT3 dominant negative (STAT3-DN) reduced expression of Bcl-xL (an antiapoptosis protein), increased spontaneous apoptosis, and enhanced sensitivity to Fas mediated apoptosis. In conclusion, several cellular mechanisms reduce the response to drug therapy in multiple myeloma. Future treatment approaches for this condition most likely will involve combinations of agents to enhance response or prevent resistance. PMID- 10528893 TI - Maintenance therapy and supportive care for patients with multiple myeloma. AB - Although multiple myeloma remains incurable, several drug therapies are valuable in the management of patients suffering from this condition. Interferon-alpha2b and prednisone are two agents that have been shown to modestly prolong disease free survival when each is used as monotherapy and when used in combination. Bone resorption and anemia are among several complications that adversely affect health and quality of life in patients with multiple myeloma. Bone resorption leads to skeletal complications, such as pathologic fractures, pain, spinal cord compression, and hypercalcemia. Bisphosphonates are specific inhibitors of osteoclastic activity that can be used to treat bone resorption. Intravenous pamidronate is effective in preventing skeletal complications and also may exert a direct inhibitory effect on myeloma cells. Exogenous epoetin alfa is helpful in treating more than half of patients with anemia during the plateau state of multiple myeloma. Overall, the management of patients with multiple myeloma is complex and should focus on the treatment of the disease process and the associated complications. PMID- 10528894 TI - Introduction: colorectal cancer: past accomplishments and future directions I. PMID- 10528895 TI - Hereditary colorectal cancer. AB - The question, "Is cancer hereditary?" has been answered beyond any doubt through the discovery of germ-line cancer-causing mutations in a subset of colorectal cancers (CRCs). Clearly, this authentication of the role of genetics was not solely dependent on molecular genetic studies, since hereditary cancer syndromes such as familial adenomatous polyposis (FAP) had been known for at least 100 years, but molecular advances are clarifying and refining clinical impressions. Have clinicians acted on the importance of hereditary factors in cancer so that this knowledge might be translated into patient benefit? Data showing that 59% of patients with FAP still die of metastatic CRC suggest that the answer is no. PMID- 10528896 TI - Screening and surveillance for colorectal cancer. AB - Colorectal cancer is the second most common cause of cancer death among American men and woman. Currently available screening and surveillance techniques are effective in detecting early-stage colorectal cancer and its premalignant precursor lesion, the adenomatous polyp (adenoma). Removal of adenomas by colonoscopic polypectomy significantly reduces the incidence of colorectal cancer. Appropriate screening and surveillance recommendations should be based on the individual's colorectal cancer risk stratification. High-risk groups, such as patients with hereditary nonpolyposis colorectal cancer (HNPCC) and familial adenomatous polyposis (FAP), should be offered genetic counseling and specialized screening recommendations for colorectal and associated extracolonic malignancies. PMID- 10528897 TI - Chemoprevention of colorectal cancer through inhibition of cyclooxygenase-2. AB - Two events in the last decade have set the stage for the large-scale clinical testing of chemopreventive agents for colorectal cancer in people at low to moderate risk for this disease. One of these is the discovery of a cause-effect relationship between the activities of cyclooxygenases (COX) and carcinogenesis in the colon, which can be interdicted by inhibitors of the enzymes. The other is the development of selective inhibitors of COX-2. These agents, when used in animals, also inhibit carcinogenesis in the colon. Additionally, they appear to be safe enough in humans to allow large-scale testing in healthy people. We review the key data implicating a causal relationship between the activity of COX and carcinogenesis and its possible mechanisms of action. We also emphasize work that points to other molecular targets for chemoprevention of colorectal cancer, which is emerging from studies of the link between COX and carcinogenesis. PMID- 10528899 TI - Surgical therapy of hepatic colorectal metastasis. AB - The liver is a common site of metastases from colorectal cancer. The data are convincing that liver resection is a safe and effective therapy for such metastatic disease. Even extensive resections can be performed with a less than 5% mortality rate at major centers and a 5-year survival rate of 30% to 40%. This review examines factors important for patient selection for liver resection and proposes a prognostic scoring system that may help in patient selection. For unresectable disease, many methods for ablating tumors are now available that will likely prove to be useful adjuncts to current treatment. PMID- 10528898 TI - Current issues in colorectal cancer surgery. AB - Increased understanding of the pathology and natural history of colorectal cancer has led to significant advances in the management of this disease. Surgical management of rectal cancer currently includes a spectrum of operative procedures ranging from radical operations to innovative sphincter-preserving techniques. At one end of the spectrum, 5% to 10% of patients present with small, superficially invasive rectal cancers amenable to a curative local excision. At the other end, a subset of patients present with locally advanced rectal cancers that require multimodality therapy, including sharp pelvic dissection with en-bloc resection of involved organs. However, the majority of rectal cancer patients present with nonfixed, yet deeply invasive, lesions requiring either a low anterior resection (LAR) or an abdominal perineal resection (APR). This report will address the controversies in the surgical management of colorectal cancer. PMID- 10528900 TI - Hepatic arterial chemotherapy in metastatic colorectal patients. AB - Hepatic metastases are a major cause of morbidity and mortality for patients with colorectal cancer (CRC). The rationale for hepatic arterial chemotherapy has both an anatomical and pharmacological basis. Several randomized clinical studies of fluoropyrimidine showed higher response rates in all trials when the drug was given as an hepatic arterial infusion (HAI) versus systemic administration. However, the studies did not accurately define survival for the following reasons: (1) some allowed a crossover; (2) some were too small; and (3) some used inadequate systemic chemotherapy. Patients who have failed to respond to previous systemic chemotherapy have an approximately 50% response rate with HAI treatment. Hepatic toxicity, especially biliary sclerosis, is the dose-limiting toxicity, occurring in 6% to 25% of patients. Adding dexamethasone to HAI fluoropyrimidine decreases the hepatobiliary toxicity. The therapeutic benefit of HAI in one study was also demonstrated by an increased time with normal quality of life. To truly define the role of regional therapy in patients with CRC confined to the liver, the current Cancer and Leukemia Group B (CALGB) study is randomizing patients to HAI versus systemic therapy without a crossover to demonstrate if HAI improves survival and/or quality of life in addition to response rates. PMID- 10528901 TI - Intraperitoneal chemotherapy in the management of colon cancer. AB - Intraperitoneal delivery of cytotoxic agents active in the treatment of colon cancer has the potential to significantly increase the exposure to these drugs to cancer present within the peritoneal cavity or liver. To date, few well controlled clinical trials have been conducted to test the relative efficacy of this strategy compared with systemic drug delivery. A major issue is the small number of useful cytotoxic agents in colon cancer, and the limited ability of the drugs to penetrate directly into tumor tissue following regional delivery. Intraperitoneal antineoplastic drug administration has its greatest therapeutic potential as an adjuvant treatment strategy for individuals with colon cancer and a high risk for recurrence in the peritoneal cavity or liver. PMID- 10528902 TI - Adjuvant therapy of rectal cancer. AB - For patients with clinically resectable stage T3 and/or node-positive rectal cancer, adjuvant radiation therapy decreases the rate of local recurrence. The addition of systemic chemotherapy further enhances local control and improves survival. Compared with postoperative therapy, preoperative treatment may have less toxicity and may increase the chance of sphincter preservation. The acute and long-term toxicity of pelvic radiation is related to the technique of delivery. Therefore, appropriate conventional radiation doses and techniques are recommended. Randomized trials comparing preoperative and postoperative combined modality therapy are in progress. PMID- 10528903 TI - Adjuvant therapy of colon cancer. AB - Colon cancer is an important cause of cancer-related mortality. A series of clinical trials of adjuvant systemic therapy have been performed in attempt to establish means to improve outcome in this disease. By the early 1990s, a role for 5-fluorouracil (5-FU)-based chemotherapy in stage III colon cancer had been firmly established. The precise role for chemotherapy in stage II disease remains under investigation. Progress continues toward optimizing the schedule and duration of systemic therapy, allowing for maximal efficacy with a minimum of toxicity. It appears that approximately 6 months of 5-FU and leucovorin are as effective as more prolonged regimens. Levamisole does not appear to add to the benefit of 5-FU and leucovorin. Several newer agents such as the oral fluorinated pyrimidines, irinotecan (CPT-11) and oxaliplatin have demonstrated activity in metastatic colon cancer and hold promise as potentially effective drugs to be tested in the adjuvant setting. PMID- 10528904 TI - Carcinoembryonic antigen screening: pros and cons. AB - The appropriate use of plasma carcinoembryonic antigen (CEA) levels in the management of patients with colorectal cancer has been debated over the last 30 years. It is clear from the very low sensitivity of this test in normal populations that there is no role for CEA assessment as a screening tool for colon cancer. Although in patients receiving chemotherapy for metastatic colon cancer, elevations of CEA generally indicate disease progression, while decreases are indicative of improvement, there is no convincing evidence that CEA monitoring significantly affects either survival or quality of life. The area of most interest for CEA monitoring has been the potential for its use after curative resection. The purpose of postoperative CEA monitoring would be to detect recurrence of cancer at an early, surgically curable stage. There is good evidence that routine CEA monitoring postresection of colon cancer detects metastatic disease on average 5 months before routine follow-up evaluation without CEA monitoring detects recurrence. Also, studies demonstrate that some patients with recurrent cancer detected by CEA monitoring may be cured by surgical resection of metastases. However, the overall cost-effectiveness of this approach is not clear, and convincing definition of the role of postoperative CEA monitoring awaits the results of large randomized clinical trials. PMID- 10528905 TI - Costs and cost-effectiveness of colorectal cancer prevention and therapy. AB - For cancer, the evaluation of new prevention and therapeutic strategies has traditionally focused almost exclusively on safety and efficacy. However, comparison of the costs and cost-effectiveness of medical interventions is increasingly being recognized as an important goal. Cancer care is a prime target for scrutiny because US cancer treatment consumes over $40 billion per year or approximately 12% of total health care expenditures. Colorectal cancer (CRC) treatment costs over $6.5 billion per year and, among malignancies, is second only to breast cancer at $6.6 billion per year. Nonetheless, there are relatively few published studies addressing the economic consequences of CRC. This review describes the strengths and limitations of the major types of health economic analyses, as well as the existing literature on the costs and cost-effectiveness of CRC prevention and treatment. Although standard approaches to both CRC screening and treatment appear cost-effective when compared with no intervention, the relative cost-effectiveness of different screening, treatment, and posttherapeutic surveillance strategies remains uncertain. As databases and information systems able to integrate comprehensive cost and treatment data grow in availability and sophistication, it should become easier to compare the impact of various approaches in terms of both traditional and economic outcomes. Over the next few years, the results of the first clinical trials that prospectively assess economic end points in CRC are anticipated; the experience resulting from these efforts should stimulate and enhance future studies. PMID- 10528906 TI - Advances in cross-sectional imaging of colorectal cancer. AB - Imaging of colorectal cancer is primarily accomplished with computed tomography (CT) and magnetic resonance imaging (MRI). These two modalities have been used for more than a decade in imaging both primary and metastatic colorectal cancer. Recent advances in the CT and MRI technologies are redefining the precise role of these imaging modalities in the work-up and evaluation of patients with colorectal cancer. PMID- 10528907 TI - Positron emission tomography imaging of colorectal cancer. AB - Continued improvements in the diagnosis and treatment of colorectal cancer are based on a clearer understanding of the pathophysiological processes underlying the disease and how it affects the patient. The addition of information derived from fluorine-18-labeled deoxyglucose (FDG) positron emission tomography (PET) scans to the cross-sectional imaging data enables a clearer understanding of the pathophysiology of the disease process. In particular, FDG PET scans are capable of demonstrating disease that mimics normal structures on conventional imaging, as well as finding disease in otherwise normal-sized lymph nodes. Abnormal areas of FDG uptake in the setting of known colorectal cancer are almost always due to recurrent disease. In addition, FDG PET offers great promise in the evaluation of treatment response, particularly as more targeted therapies become available. This report covers the basic principles underlying PET imaging, including the biochemistry of FDG and methods of PET analysis. We then, review the clinical indications for FDG PET scanning in colorectal cancer. PMID- 10528908 TI - Future strategies toward the cure of indolent B-cell malignancies: introduction. PMID- 10528909 TI - Future strategies toward the cure of indolent B-cell malignancies. Molecular genetic approaches. AB - Cancer is a genetic disease caused by somatic mutation(s) in essential genes that regulate cell growth or survival. Advances in genetics have identified molecular defects that can be targeted by novel pharmaceutics that exploit the altered genetic machinery of tumor cells. Gene transfer also can be used to potentiate the host immune response to weak tumor antigens that may arise during the process of neoplastic transformation. Advances in our understanding of the mechanics of immune activation have led to development of immune-based strategies for gene therapy of cancer. Preliminary findings in a phase I trial of one such approach in patients with chronic lymphocytic leukemia (CLL) are encouraging. Conceivably, we soon may see the advent of effective gene therapy for patients with indolent B cell malignancies. PMID- 10528910 TI - Future strategies toward the cure of indolent B-cell malignancies. Immune recognition and antiself. AB - Tumor cells from patients with B-cell malignancies exhibit a number of immunophenotypic characteristics that may be responsible for their survival. It has been shown, for example, that down-modulation of CD154 (CD40 ligand), which allows B cells to respond to T cells, may account for some of the immune defects of these cells in patients with chronic lymphocytic leukemia. New strategies are being developed in an attempt to enable the host immune system to recognize these cells as non-self and to augment the host response against tumor cells. PMID- 10528911 TI - Future strategies toward the cure of indolent B-cell malignancies. New biologic therapies. AB - Although chronic lymphocytic leukemia (CLL) cells provide an excellent target for antibody therapy, to date this approach has met with limited success in patients with CLL. Promising data are emerging with a new generation of antibodies, such as Campath-1H, which may offer effective therapeutic options not only as single agents but also in combination with such drugs as fludarabine. Other new antibodies, including rituximab and several new radioimmunoconjugates, have provided impressive results in the indolent non-Hodgkin's lymphomas and warrant further investigation. PMID- 10528912 TI - Stem-cell transplantation for indolent lymphoma. AB - Stem-cell transplantation (SCT) has become the treatment of choice for patients with relapsed aggressive non-Hodgkin's lymphoma (NHL). The role of SCT in the management of patients with low-grade NHL remains controversial, although increasing numbers of patients with advanced-stage follicular lymphoma, mantle cell lymphoma, and chronic lymphocytic leukemia (CLL) are now undergoing SCT. There is increasing concern regarding toxicity of autologous SCT, especially the long-term risk of development of myelodysplastic syndrome, which is higher than was expected. This has led to renewed interest in the role of allogeneic SCT for patients with NHL. A major advantage of allogeneic SCT is the potential for exploiting a graft-versus-lymphoma effect, and many studies under way are exploring the possibility of manipulating donor cells and the host to maximize T cell responsiveness against lymphoma. PMID- 10528913 TI - New chemotherapeutics strategies for the treatment of indolent lymphoid malignancies. AB - During the past few decades, clinical trials in indolent lymphoid malignancies have focused to a large extent on comparisons of combinations and permutations of conventional drugs, including alkylating agents and anthracyclines. Despite an enormous expenditure of financial and patient resources, there has been no major impact on survival, and these tumors remain incurable. Innovative approaches are clearly needed, including the development of new and more effective agents. PMID- 10528914 TI - Two pathways for the induction of apoptosis in human lymphocytes. AB - PURPOSE: To assess the roles of cell membranes and DNA as targets in radiation induced apoptosis. MATERIALS AND METHODS: Peripheral blood lymphocytes from normal human donors were exposed to different types of apoptosis-inducing agents. Several measures of apoptosis were used to compare the kinetics of the processes induced, as well as to correlate the processes with DNA damage and membrane oxidation. RESULTS: Two kinetically distinct processes were observed. DNA damaging agents, such as ionizing radiation, bleomycin, cisplatin and the topoisomerase inhibitor m-amsacrine, induced apoptosis by a kinetically slow process initiated by DNA damage and dependent on protein synthesis, but which did not correlate with membrane oxidation. Conversely, the agents t-butyl hydroperoxide and cumene hydroperoxide induced apoptosis by a kinetically fast process independent of protein synthesis and which did correlate with membrane oxidation. CONCLUSIONS: Slowly repaired or unrepairable DNA lesions, such as some of those produced by ionizing radiation exposure, trigger apoptosis by a kinetically slow process. This slow apoptotic pathway is distinct from a fast process not induced by radiation but which is induced by membrane-oxidizing agents. PMID- 10528915 TI - Apoptosis and the adaptive response in human lymphocytes. AB - PURPOSE: To determine whether the sensitivity of human lymphocytes for apoptosis induced by either a membrane oxidizing agent or a DNA damaging agent is modified by an adaptive response. MATERIALS AND METHODS: Peripheral blood lymphocytes from normal human donors were exposed to low doses of the DNA damaging agent gamma radiation, or the membrane oxidizing agent t-butyl hydroperoxide (t-BuOOH), incubated for various times and then tested for their sensitivity to induction of apoptosis by a subsequent exposure to a high dose of either agent. Apoptosis was measured using a fluorescent assay of DNA unwinding or a terminal deoxynucleotide transferase assay. RESULTS: The results show that Go lymphocytes pre-exposed to an adapting dose of radiation or DNA strand breaking agent are not protected but can become sensitized to subsequent apoptosis induced by radiation (a kinetically slow process). Inter- and intraindividual variations were observed. However, neither pre-exposure to radiation nor to a membrane oxidizing agent sensitized lymphocytes from any donor to apoptosis induced by a membrane oxidizing agent (a kinetically fast process). CONCLUSIONS: Since an increase in the elimination of genetically damaged cells by apoptosis could reduce the risk of cancer from exposure to radiation or other DNA damaging agents, this cellular sensitization for apoptosis may represent a novel adaptive response mechanism. PMID- 10528916 TI - Restoration by glycerol of p53-dependent apoptosis in cells bearing the mutant p53 gene. AB - PURPOSE: Effective heat-induced cell death in cultured cells bearing a mutant p53 (mp53) gene was sought by glycerol treatment which led to conformational change from mp53 to wild-type p53 (wtp53) in p53-null murine fibroblasts transfected with mp53. MATERIALS AND METHODS: Heat sensitivity was measured using a colony forming assay. For heat-induced apoptosis, gel electrophoresis was applied to detect DNA fragmentation and Hoechst33342 staining for apoptotic bodies. Glycerol (0.6 M) was applied to the cultured cells 48 h before heating at 44 degrees C in a water bath. RESULTS: Wtp53 transfectants (MT158/wtp53-1) were sensitive to heat stress compared with mp53 transfectants (MT158/mp53-2 cells), and the combined treatment with glycerol enhanced cell killing only in the MT158/mp53-2 cells. After glycerol pretreatment for 48 h, the subsequent heat treatment enhanced DNA fragmentation and apoptosis in MT158/mp53-2 cells, while DNA fragmentation and apoptotic bodies were not enhanced with heat treatment alone in these cells. In contrast, DNA fragmentation or apoptotic bodies were clearly observed in MT158/wtp53-1 cells 3-24h after heat treatment. Treatment with glycerol alone did not induce apoptosis in the transfectants. CONCLUSIONS: Glycerol appears to function as a chemical chaperone that restores mp53 to wtp53 function. PMID- 10528917 TI - Effects of intracellular calcium chelator BAPTA-AM on radiation-induced apoptosis regulated by activation of SAPK/JNK and caspase-3 in MOLT-4 cells. AB - PURPOSE: To examine the roles ofintracellular calcium in radiation-induced apoptosis of MOLT-4 cells, the effects of intracellular calcium chelator BAPTA-AM on the induction of apoptosis and the activation of apoptosis-relating enzymes SAPK/JNK and caspase-3 were studied. MATERIALS AND METHODS: MOLT-4 cells pretreated with 5 microM BAPTA-AM were exposed to X-rays. DNA fragmentation, the expression of phosphorylated SAPK/JNK and the activation of caspase-3 and calcium concentration were measured by agarose gel electrophoresis, Western blotting and spectrofluorometry. RESULTS: Time-dependent ladder-like DNA fragmentation was observed at 4h, 5 h and 6 h after exposure to 15 Gy of X-rays. This fragmentation was significantly attenuated by pretreatment with BAPTA-AM up to 5 h after irradiation, but the attenuation due to BAPTA-AM was no longer detectable at 6 h. Activation of SAPK/JNK and caspase-3 was observed at 1 and 4 h after X irradiation, respectively, and BAPTA-AM retarded the activation for 2 h. The pretreatment with BAPTA-AM was found to suppress the increase of calcium concentration for 6h after irradiation. CONCLUSION: These results revealed that chelation of calcium merely delayed the onset of the radiation-induced apoptosis regulated by the activation of SAPK/JNK and caspase-3, and calcium was not essential for the induction of apoptosis in X-irradiated MOLT-4 cells. PMID- 10528918 TI - Combined FISH with pan-telomeric PNA and whole chromosome-specific DNA probes to detect complete and incomplete chromosomal exchanges in human lymphocytes. AB - PURPOSE: To combine FISH with pan-telomeric peptide nucleic acid (PNA) and whole chromosome-specific DNA probes to detect complete and incomplete chromosome exchanges in human lymphocytes. MATERIALS AND METHODS: Human lymphocytes were irradiated in vitro with 0.9 Gy low dose-rate (0.019 Gy/h) tritium beta-rays. Metaphase spreads were treated with RNase, fixed in 1:3 acetic acid:methanol, and then further treated with KCl, proteinase K and fixed in 4% paraformaldehyde. Slides were denatured, hybridized for 1.5 h with an FITC-labelled telomeric PNA probe, and rehybridized overnight with a spectrum-orange whole-chromosome probe specific for chromosomes 1, 2 and 4. Hybridized spreads were washed with 70% formamide/20 x SSC and counterstained with DAPI. RESULTS: All three pairs of labelled chromosomes together with 92 telomeres were readily visible after hybridization. The whole chromosomes 1, 2 and 4 were painted orange, and all telomeres were stained green. Unpainted chromosomes were counterstained blue. In the observed 680 chromosome aberrations induced by tritium beta-rays in human lymphocytes after 52 h of culture, no evidence of telomere addition was detected. Incomplete and hidden complete exchanges and terminal deletions were definitively discriminated. CONCLUSION: The simultaneous detection of telomeres and specific whole chromosomes allows for the first time accurate analysis of complete and incomplete chromosome exchanges involving painted chromosomes in human lymphocytes. PMID- 10528919 TI - Simple chromosome exchanges are not linear with dose. AB - PURPOSE: To check whether simple chromosome exchanges are linearly related to radiation dose. MATERIALS AND METHODS: Go-irradiated lymphocytes were cultured to produce metaphase preparations. Chromosomes 1 and 2 were painted different colours and the remaining chromosomes counterstained. Cells containing a colour junction involving both chromosomes 1 and 2 were scored fully, so that simple and complex rearrangements were distinguished. RESULTS: At doses of 2 Gy and below very few complex rearrangements were seen. About 90% of exchanges were simple. At these doses the linear component of the dose-effect curve accounts on average for only approximately 30% of the observed yield. CONCLUSIONS: There is a square-law dose component to the yield of simple exchanges in addition to the linear term. PMID- 10528920 TI - Induction and persistence of chromosomal exchanges in mouse bone marrow cells following whole-body exposure to X-rays. AB - PURPOSE: To investigate the induction and persistence of chromosome aberrations in mouse bone marrow cells after X-ray exposure and to detect differential involvement of individual chromosomes in translocations. MATERIALS AND METHODS: Male and female Swiss mice were exposed to 1 and 3 Gy of X-rays. Chromosome aberrations in bone marrow cells were analysed at 1, 7, 21 and 100 days following irradiation by means of fluorescence in situ hybridization (FISH) with mouse chromosome-specific DNA libraries (#1,13; #2,8; #6,15 and X,Y). In total, 38% of mouse genome was painted and examined. RESULTS: Pooled data indicate that the frequencies of dicentrics and fragments decreased with time and reached to the control level at day 21 after exposure. Following exposure to 1 Gy of X-rays, the frequencies of translocations were not significantly lower between days 7 and 100 than observed at day 1. However, the frequencies of translocations for the 3 Gy group were significantly (about 40%) lower at day 7, then remained constant up to day 100. After exposure to 3Gy of X-rays, the frequencies of non-reciprocal translocations decreased with time, whereas reciprocal translocations between days 7 and 100 were not significantly less frequent than at day 1. A comparison of observed and expected numbers of translocations involving individual chromosomes showed that at day 1 after irradiation, distribution of X-ray-induced translocations among the painted chromosomes was proportional to their DNA content. However, at day 100 after exposure, the observed translocations involving chromosome 2 were more frequent than expected, those involving chromosomes 8 and 15 were less frequent than expected, while chromosomes 1, 6, X and Y were involved as frequently as expected. CONCLUSION: Among induced translocations, non-reciprocal translocations are relatively unstable, especially after exposure to high-dose X-rays. While the initial distribution of X-ray induced translocations is proportional among the painted chromosomes, the persistence of these translocations is heterogeneous. PMID- 10528921 TI - Chromatid break rejoining and exchange aberration formation following gamma-ray exposure: analysis in G2 human fibroblasts by chemically induced premature chromosome condensation. AB - PURPOSE: To analyse the kinetics of chromatid break induction, rejoining, and misrejoining after y-irradiation in G2 phase human cells using premature chromosome condensation induced by calyculin A. MATERIALS AND METHODS: Human fibroblast AG1522 cells were irradiated with gamma-rays and chromosomes were then prematurely condensed by calyculin A. The number of chromatid breaks and chromatid exchanges in G2 chromosomes were scored, and fitted curves were calculated. RESULTS: Calyculin A induced premature chromosome condensation in cells immediately after irradiation. Kinetics of rejoining of chromatid breaks demonstrated two exponential components with rapid and slow time constants. Within 5 min after irradiation, the number of chromatid breaks fell rapidly to about one-half, then gradually decreased. Chromatid exchanges were formed very quickly, reaching a plateau within 20 min from exposure. CONCLUSIONS: Chemically induced premature chromosome condensation technique allows a simple, rapid and precise analysis of chromatid breakage and rejoining. The rapid kinetic component was particularly well characterized. PMID- 10528922 TI - Cell cycle checkpoint evasion and protracted cell cycle arrest in X-irradiated small-cell lung carcinoma cells. AB - PURPOSE: To determine the longevity and dose-dependence of acute X-irradiation induced cell cycle perturbations in a panel of seven small-cell lung carcinoma (SCLC) cell lines (COR-L32B, COR-L51B, COR-L88B, COR-L96C, COR-L103, COR-L266B, COR-L279), assessed for TP53 tumour suppressor gene status and showing characteristically long population doubling periods. MATERIALS AND METHODS: Cell lines were screened for abnormalities in TP53. Cell cycle arrest and nuclear fragmentation were determined by flow cytometry under culture conditions that minimized the propensity of SCLC cells to form multicellular aggregates. A faster growing SCLC cell line (NCI-H69) and two breast tumour cell lines were used as controls. RESULTS: NCI-H69 and five of the COR-SCLC cell lines showed clear evidence of TP53 abnormalities and the cycle arrest responses of the breast tumour cell lines established the effects of TP53 mutation on G1/S checkpoint loss. All SCLC lines, at 24 h after low dose irradiation, showed abrogation of the G1/S checkpoint together with a range of expression of a protracted G2/M delay. G2/M delay progressed in all panel cell lines up to 48 h post-irradiation while NCI-H69 showed significant recovery for the dose range 75-600cGy. Only NCI H69 and one panel line showed dose-dependent progression to complete nuclear DNA fragmentation. CONCLUSIONS: The culture method permits the measurement of cell cycle effects that reflect the TP53 status of SCLC cells. G1/S checkpoint failure, long-term radiation-induced G2 arrest, highly muted apoptotic responses and delayed recovery appear to be typical responses of the recently derived COR SCLC lines. The results imply that low levels of unrepaired DNA damage, induced at clinically relevant doses, can persist for days in SCLC cells with long cell cycle traverse times, and can remain capable of checkpoint activation with implications for S phase-targeted therapies. PMID- 10528924 TI - Potential of radiation-induced chromosome aberrations to predict radiosensitivity in human tumour cells. AB - PURPOSE: To validate whether the number of aberrations could be used as a measure of the radiosensitivity of human tumour cells. If so, this would potentially provide a more rapid method than the colony assay to predict radiocurability in human tumour biopsy material. MATERIALS AND METHODS: A panel of 13 human tumour cell lines was investigated, covering a wide range of radiosensitivities. Fluorescence in situ hybridization (FISH) employing whole chromosome probes was used to detect aberrations. RESULTS: A dose-dependent increase in radiation induced chromosome aberrations was observed in all cell lines. A good correlation (r=0.90) was found between cell survival and total chromosome aberrations in 12 of the 13 cell lines (92%), with one exception. A poorer correlation was observed between cell survival and stable- (r=0.85) and unstable-type aberrations (r=0.81). Survival-aberration correlations for individual radiation doses were worse, although statistically significant. The exceptional cell line showed significantly more aberrations for a given level of cell kill than expected based on data for the other lines. CONCLUSION: This study indicates that radiation induced chromosome aberrations can be used as a potential predictor of intrinsic radiosensitivity for the majority of human tumours when more than one dose level is tested. This could aid the design of radiotherapy schedules for each individual patient, or in the decision of whether to use an alternative therapy. PMID- 10528923 TI - No association between radiosensitivity and TP53 status, G1 arrest or protein levels of p53, myc, ras or raf in human melanoma lines. AB - PURPOSE: First, to investigate whether TP53 status and/or radiation-induced G1 arrest are associated with radiosensitivity, and, second, to detect possible associations between protein levels of p53, myc, ras or raf and radiosensitivity and to investigate whether hypoxia-induced changes in the levels of these proteins are related to hypoxia-induced changes in radiosensitivity in human melanoma lines. MATERIALS AND METHODS: Radiosensitivity was assessed by clonogenic assays. TP53 status was investigated at the genomic level by constant denaturant gel electrophoresis and at the cDNA level by sequencing. G1 arrest was investigated by flow cytometric analysis of DNA. Protein expression of hypoxia treated and untreated cells was assessed by flow cytometric measurements and Western blotting. RESULTS: Considerable differences in radiosensitivity were detected among melanoma lines with wild-type TP53. Only a fraction of the melanoma cells, differing between the lines, was arrested in G1. No association between the fraction of arrested cells and radiosensitivity was detected. Protein levels of p53, myc, ras or raf were not associated with radiosensitivity. Hypoxia induced changes in p53, ras and raf levels were detected in all cell lines. Changes in the level of myc protein were detected for two of the four cell lines, while hypoxia-induced changes in radiosensitivity were observed only for one. CONCLUSIONS: Differences in radiosensitivity among melanoma lines cannot be elucidated by TP53 status, differences in G1 arrest or different levels of p53, myc, ras or raf proteins. Hypoxia-induced changes in p53, myc, ras or raf levels do not seem to be related to hypoxia-induced changes in radiosensitivity. PMID- 10528925 TI - Free radicals from lyophilized 'dry' DNA bombarded with heavy-ions as studied by electron spin resonance spectroscopy. AB - PURPOSE: To investigate the number and species of free radicals from dry DNA after bombardment with heavy-ions at low temperature in comparison with X irradiation at 77 K by electron spin resonance (ESR) spectroscopy. MATERIALS AND METHODS: Solid DNA samples were either X-irradiated at 77 K as cylinder samples or heavy-ion-bombarded (LET range 1500-12400keV/microm) at the Gesellschaft fur Schwerionenforschung (GSI Darmstadt) as very thin tablets. Data acquisition was on a Bruker ESP 380 ESR-spectrometer (X-band, 9.5 GHz). Data analysis involved computer treatment of spectra. RESULTS: Spectra from heavy-ion-bombarded samples were found to reveal the same free radical species with about the same relative contributions for most components as those from X-irradiated samples at comparable doses. Only two components, a thymine allyl radical and a Cl' deoxyribose located species, were enhanced after heavy-ion bombardment. Dose effects in both cases involved a higher relative amount of deoxyribose-derived free radicals. The analysis of G (total free radicals, taken from dose-response curves) were typically smaller than those for X-rays but showed no clear dependence on LET. CONCLUSIONS: The differing biological response to high-LET particle bombardment compared with low-LET irradiation is not strongly reflected by the chemical structure and total number of initial free radicals. It might rather derive from an inhomogeneous distribution of energy deposition (resulting in 'clustered damages') or from effects at higher chemical and biological levels (e.g. product formation and repair) which is not strongly apparent in the free radical characteristics obtained by ESR-spectroscopy. PMID- 10528926 TI - Hydroxyl radical scavenging by carnosine and Cu(II)-carnosine complexes: a pulse radiolysis and spectroscopic study. AB - PURPOSE: To obtain a wider insight into the general properties of carnosine and to provide support to its anti-oxidative role. This property is hypothesized to be linked to various mechanisms including free-radical scavenging and metal chelation (i.e. Cu(II)). METHODS: Pulse-radiolysis experiments were performed by a 12 MeV electron linear accelerator (LINAC) on carnosine/copper(II) (2:1) and carnosine aqueous solutions at different pH. Raman spectra of solid samples were obtained by a Bruker IFS 66 spectrometer. RESULTS: As well as for free carnosine, in the presence of copper ions the interaction of carnosine with *OH radicals involves the imidazole group of the molecule. The oxidation of copper (II) carnosine system by *OH radicals is related to the pH-dependent structure of the copper(II)-carnosine complex. Raman spectra indicate that at alkaline pH the formation of a dimeric species containing two carnosine molecules complexed to two Cu2+ ions takes place. This structure can address the *OH attack more selectively than carnosine itself to different sites of the imidazole ring. The formation of at least two different *OH-radical adducts occurs and positions C(2) and C(5) of the imidazole ring are the preferential sites for the *OH attack, as the heterocyclic ring is mainly present as its N(1)-protonated tautomeric form. CONCLUSION: These studies provide further evidence about the formation of carnosine-copper complexes and the predominance of a dimeric structure at slightly basic pH. The chelation of Cu(II) is not detrimental to the scavenging ability of carnosine. Raman spectra are helpful in identifying the structure of the copper(II)-carnosine complexes and in predicting the preferential sites for the *OH attack to the carnosine-copper system. PMID- 10528928 TI - Recent advances in the genetics of epilepsy: insights from human and animal studies. AB - Progress in understanding the genetics of epilepsy is proceeding at a dizzying pace. Due in large part to rapid progress in molecular genetics, gene defects underlying many of the inherited epilepsies have been mapped, and several more are likely to be added each year. In this review, we summarize the available information on the genetic basis of human epilepsies and epilepsy syndromes, and correlate these advances with rapidly expanding information about the mechanisms of epilepsy gained from both spontaneous and transgenic animal models. We also provide practical suggestions for clinicians confronted with families in which multiple members are afflicted with epilepsy. PMID- 10528927 TI - Induction of radical scavenging ability and protection against radiation-induced damage to microsomal membranes following low-dose irradiation. AB - PURPOSE: To investigate changes in rat liver cytosolic radical scavenging ability and in microsomal membrane function following low doses of radiation. MATERIALS AND METHODS: Wistar rats were irradiated with 1-50 cGy of X-rays and liver cytosolic radical scavenging ability was determined using DPPH, a stable free radical. Liver microsomal drug metabolizing activity was determined using hexobarbital as a substrate. Cytosolic antioxidants and microsomal enzymes were spectrophotometrically determined. RESULTS: Irradiation of rats at around 5-10 cGy induced liver cytosolic radical scavenging ability and a considerable increase in this ability at 5 cGy was observed for 3 days after irradiation. Glutathione reductase was suggested as a candidate of cytosolic antioxidants. Radiation-induced damage to liver microsomal drug metabolizing enzyme activity was suppressed by preirradiation with 5 cGy, mainly by protecting cytochrome P 450. CONCLUSIONS: Low doses of radiation increased cytosolic radical scavenging ability and resulted in protection of microsomal membrane function which is easily damaged by radiation-induced free radicals. PMID- 10528929 TI - Effects of pregnancy on the pharmacokinetics of lamotrigine in dogs. AB - PURPOSE: This study was designed to evaluate the effects of pregnancy on the kinetics of lamotrigine (LTG). METHODS: Five pregnant dogs were given a daily dose of LTG (100 mg) for a period of 1 week. Two months after parturition, the same subjects were given the LTG dose (100 mg) over the same period. On both occasions, plasma LTG concentrations were determined by a sensitive, high performance liquid chromatographic (HPLC) method, over a 30-h period after the last dose. RESULTS: The mean maximum plasma concentration (Cmax), volume of distribution (Vd/F), and oral body clearance (Cl/F) for LTG (+/- SD) during pregnancy were 7.63+/-2.46 microg/ml 1.74+/-0.29 L/kg, and 0.19+/-0.04 L/h/kg, respectively. After pregnancy, the same variables were 6.12+/-2.24 microg/ml, 2.36+/-1.10 L/kg, and 0.30+/-0.13 L/h/kg, respectively. None of these pharmacokinetic parameters was found to be significantly different between the two groups. CONCLUSIONS: The apparent lack of change in the relevant pharmacokinetic parameters of LTG during pregnancy may indicate that pregnancy has little or no effect on glucuronidation; the principal pathway for the drug's elimination. PMID- 10528930 TI - Age-specific N-methyl-D-aspartate-induced seizures: perspectives for the West syndrome model. AB - PURPOSE: With intraperitoneal N-methyl-D-aspartate (NMDA; 15-200 mg/kg) administration, we attempted to develop an animal model of age-specific West syndrome to serve for testing of putative anticonvulsant drugs and to determine the mechanisms of this disorder. METHODS: Experiments were performed in 12-, 18-, and 60-day-old (adult) rats. The effects of systemic pretreatment with hydrocortisone (5-25 mg/kg), pyridoxine (20-250 mg/kg), and sodium valproate (VPA; 200 and 400 mg/kg) against the NMDA-induced automatisms, emprosthotonic (hyperflexion), and clonic-tonic seizures were determined. NMDA-induced EEG changes and alterations of the performance in horizontal bar, rotorod, open field, and elevated plus-maze tests were recorded. RESULTS: In young rats, hydrocortisone had proconvulsant effects. High doses of pyridoxine induced epileptiform activity independent of and distinct from that induced by NMDA. Only VPA had moderate effects against the NMDA-induced syndrome. EEG consisted of periods of suppression mixed with ictal activity of serrated waves and high voltage chaotic EEG activity. In adult rats, EEG alterations involved spike and spike-and-wave activity. NMDA also deteriorated performance of young rats in the open field, rotorod, and elevated plus maze tests. CONCLUSIONS: NMDA syndrome in rats fulfills some, but not all, criteria of the West syndrome model, such as occurrence of flexion seizures, nonspecific diffuse EEG changes, refractoriness to antiepileptic therapy (but a response to VPA), as well as long-term alteration of behavioral tasks. However, NMDA-induced seizures represent an acute model without the occurrence of spontaneous seizures, whereas in the clinical situation, both the seizures and neurologic deterioration are chronic. Further, in the West syndrome and the NMDA seizure model, there is an incongruent response to therapy with antiepileptic drugs. PMID- 10528932 TI - Distribution of epilepsy syndromes in a cohort of children prospectively monitored from the time of their first unprovoked seizure. AB - PURPOSE: To assess the distribution of epilepsy syndromes and their stability in children. METHODS: A cohort of 407 children with a first unprovoked seizure was prospectively recruited and followed up for a mean of 9.4 years. Etiology and epilepsy syndromes were classified by using the International League Against Epilepsy (ILAE) guidelines in the 182 children with two or more seizures. Classification was done both at time of second seizure and at last follow-up. Two year terminal remission also was analyzed by etiology and epilepsy syndrome. RESULTS: Etiology of epilepsy syndromes was idiopathic in 45 (25%), cryptogenic in 89 (49%), and remote symptomatic in 48 (26%). In the initial classification, 114 (63%) children had a localization-based epilepsy syndrome including idiopathic in 26, cryptogenic in 34, and symptomatic based on localization or etiology in 54. Twenty-one (12%) children had a generalized epilepsy syndrome, including 19 with primary generalized epilepsy. Forty-seven (26%) cases were in the category of undetermined if focal or generalized. At last follow-up, there was a change in either etiology (n = 16) or the final epilepsy syndrome classification (n = 33) or both (n = 15) in 34 (19%) cases. At time of last follow-up, 144 (79%) of the children with epilepsy were in 2-year terminal remission, and 108 (59%) were in 2-year terminal remission without medications. Factors associated with a favorable prognosis included an idiopathic or cryptogenic etiology and having a localization-based idiopathic epilepsy syndrome. CONCLUSIONS: After two seizures, childhood-onset epilepsy can be classified by etiology and epilepsy syndrome. Prognosis is favorable in the majority of cases. However, the apparent syndrome may change with longer follow up. The ability to classify these cases early in the clinical course is important if they are to be used for prognostic purposes. PMID- 10528931 TI - Limbic seizures alter reproductive function in the female rat. AB - PURPOSE: Reproductive dysfunction and endocrine disorders are common among women with temporal lobe epilepsy. This study used the kindled rat model to test the hypothesis that limbic seizures directly contribute to reproductive dysfunction. METHODS: Kindling electrodes were implanted in the basolateral amygdala in adult female rats. Females were kindled by either brief, daily, suprathreshold stimulations with a bipolar electrode or sham-kindled (controls). Electrographic and behavioral seizures were monitored. Estrous cycles also were monitored with daily vaginal smears. RESULTS: Seizures arrested ovarian cyclicity in all (n = 42) kindled animals, the rats exhibiting persistent vaginal cornification (PVC). In these animals PVC was associated with high serum estradiol, increased pituitary weight, and polyfollicular ovaries consisting of many cystic follicles, as well as follicles in various stages of growth and atresia. In 93% of females, this effect occurred after the development of stage 5 motor seizures, when focal seizures had secondarily generalized. In contrast, only five (21%) of 24 sham kindled controls exhibited PVC. A single injection of progesterone (P4) temporarily restored cyclicity in five (18%) of 28 kindled females exhibiting PVC. In contrast, P4 administration restored cyclicity in all five sham-kindled controls that had spontaneously stopped cycling. P4 treatment to kindled females in PVC resulted in a different endocrine profile than that in non-P4-treated, kindled rats in PVC. P4-treated rats had high serum estradiol, testosterone, and prolactin levels; they showed an increase in pituitary weight; and their ovaries contained numerous corpora lutea and cystic follicles surrounded by markedly overdeveloped thecal cell layers. CONCLUSIONS: Seizures initiated in the amygdala result in impairment of the hypothalamic-pituitary axis, resulting in loss of ovarian cyclicity. PMID- 10528933 TI - Epidemiologic study of epilepsy in young Singaporean men. AB - PURPOSE: This survey of 20,542 Singaporean men born in 1974 studied the clinical features of young men diagnosed with epilepsy on preenlistment screening. METHODS: All male citizens in Singapore are medically screened at age 18 years before enlistment for compulsory military service. Patients suspected to have epilepsy are then referred to government hospitals for further management. We interviewed the patients and their parents and reviewed their hospital records. RESULTS: Eighty-nine patients with epilepsy were identified, indicating a lifetime prevalence of 4.9/1,000 males by age 18 years. The lifetime prevalence of epilepsy among Chinese, Malays, and Indians were 5.2, 2.8, and 6.4/1,000, respectively; these differences were not statistically significant. The mean age of seizure onset was 11.1 years. Generalized seizures (65.2%) were commoner than partial seizures (34.8%); common seizure types included generalized tonic-clonic seizures (52.8%), complex partial seizures with secondary generalization (24.7%), and myoclonic seizures (5.6%). Common epileptic syndromes included temporal lobe epilepsy (16.9%), juvenile myoclonic epilepsy (5.6%), and frontal lobe epilepsy (2.2%). Eighty-four (94.4%) patients sought medical treatment, and seven (7.9%) patients sought additional traditional treatment. Although 70 (78.7%) patients responded to medication, 14 (15.7%) patients remained refractory to treatment. CONCLUSIONS: The lifetime prevalence of epilepsy in young Singaporean men was 4.9/1,000. The majority (65.2%) had generalized seizures. Temporal lobe epilepsy was the commonest (16.9%) defined epilepsy syndrome. More patients with epilepsy (94.4%) sought medical treatment, although 15.7% remained refractory to medication. PMID- 10528934 TI - Short-term mortality after a first epileptic seizure: a population-based study. AB - PURPOSE: To determine the short-term mortality in a prospective incidence cohort of patients included after any kind of first afebrile epileptic seizure (i.e., provoked and unprovoked). METHODS: Information on death occurring within the first year of follow-up was collected in a cohort of 804 patients with a first seizure between March 1, 1984, and February 28, 1985, in southwest France. The variables analyzed were the etiology of seizure, cause of death, interval between seizure and death, and age of patients. RESULTS: By the end of the 1-year follow up, there were 149 deaths among these patients as compared with 16 expected deaths [standardized mortality ratio (SMR), 9.3; 95% confidence interval (CI), 7.9-10.9]. There were no deaths in patients with idiopathic seizures. Patients with cryptogenic seizures had slightly increased mortality (SMR, 1.6; 95% CI, 0.4 4.1). Mortality was increased for patients with remote symptomatic seizures (SMR, 6.5; 95% CI, 3.8-10.5), provoked seizures (SMR, 10.1; 95% CI, 8.1-12.4), and seizures due to a progressive neurologic condition (SMR, 19.8; 95% CI, 14.0 27.3). Causes of death were underlying pathology (64%), unrelated condition (20%), unknown cause (9%), seizure-related death (6%), and one suicide. CONCLUSIONS: Early mortality clearly differed according to the etiology of the first seizure. The highest mortality was associated with provoked seizures and with seizures caused by progressive central nervous system disorders. Patients died far more often from underlying or unrelated conditions than from seizures. PMID- 10528935 TI - Correlations between granule cell dispersion, mossy fiber sprouting, and hippocampal cell loss in temporal lobe epilepsy. AB - PURPOSE: Correlations between granule cell dispersion (GCD), collateral mossy fiber (MF) sprouting, and hippocampal cell loss were studied to assess the relation between GCD and synaptic reorganization in the dentate gyrus of patients with epilepsy. METHODS: Twenty specimens from patients with medically intractable temporal lobe epilepsy (TLE) were studied along with two control specimens. GCD was considered to be present when the stratum granulosum was wider than 120 microm, the close apposition between the granule cell (GC) soma was lost, and GCs were scattered in the molecular layer (ML). Patterns of MF sprouting were differentiated as wide or narrow according to the area of neo-Timm's staining in the ML. GC loss and volumetric cell-density decreases in the different subfields were assessed. RESULTS: MF sprouting was observed in 16 (80%) and GCD in nine (45%) cases. A significant correlation was found between MF sprouting and cell loss in all the subfields except the cornu Ammonis field 2 (CA2). A wide band of MF sprouting was associated with severe cell loss. Cases with GCD had a wide band of MF sprouting and also a higher degree of cell loss than cases without GCD. CONCLUSION: GCD is associated with a specific pattern of MF sprouting, but cell loss was found to be a major determinant for MF reorganization. PMID- 10528936 TI - The effect of menopause and perimenopause on the course of epilepsy. AB - PURPOSE: The purpose of this study was to obtain preliminary information about the effect of menopause and perimenopause on the course of epilepsy, and to determine whether seizure type, use of hormone-replacement therapy (HRT), or a history of catamenial seizure pattern would influence this course. METHODS: We performed a questionnaire study of women with epilepsy currently in menopause and perimenopause, requesting information regarding the course of their epilepsy and treatment. Statistical analysis was performed by using Pearson chi2 with 95% confidence limits. RESULTS: Forty-two menopausal women (ages 41-86 years) responded. Twelve subjects reported no change in seizures at menopause, 17 reported a decrease in seizure frequency, and 13 reported an increase. Sixteen (38%) took synthetic HRT. Sixteen (38%) additional subjects (having some overlap with the HRT group) reported having a catamenial seizure pattern before menopause. HRT was significantly associated with an increase in seizures during perimenopause (p = 0.001). A history of catamenial seizure pattern was significantly associated with a decrease in seizures at menopause (p = 0.013). Thirty-nine perimenopausal women (ages 38-55 years) responded. Nine subjects reported no change in seizures at perimenopause, five reported a decrease in seizure frequency, and 25 reported an increase. Eight (15%) subjects took synthetic HRT, and 28 (72%) reported having a catamenial seizure pattern before menopause. HRT had no significant effect on seizures; however, a history of catamenial seizure pattern was significantly associated with an increase in seizures at perimenopause (p = 0.02). CONCLUSIONS: These pilot data suggest that synthetic HRT may be associated with an increase in seizure frequency in menopausal women with epilepsy. A catamenial seizure pattern may be associated with seizure decrease during menopause but with an increase during perimenopause. PMID- 10528937 TI - Stereotactic amygdalohippocampotomy for the treatment of medial temporal lobe epilepsy. AB - PURPOSE: This study was carried out to assess the safety and efficacy of stereotactic ablation of the amygdala and hippocampus for the treatment of medial temporal lobe epilepsy. METHODS: Twenty-two stereotactic amygdalohippocampotomies were performed in 19 patients with unilateral temporal lobe seizures by using magnetic resonance imaging (MRI) localization for target planning and radiofrequency techniques for lesion production. Seizure frequency was assessed at 3-monthly follow-up visits. Two lesion groups were defined. In group I, four to 11 (mean, 6.4) discrete lesions were made, encompassing the amygdala and anterior 13-21 mm (mean, 16.8 mm) of the hippocampus. In group II, a large number of confluent lesions were made (mean, 26.0; range, 12-54) encompassing the amygdala and anterior 15-34 mm (mean, 21.5 mm) of the hippocampus. MRI scanning was carried out 24 h and 6-9 months after surgery. RESULTS: In five group I patients, one (20%) experienced a favorable seizure outcome. Of 15 group II patients, one of whom had previously undergone limited lesioning and was also analyzed as part of group I, nine (60%) experienced a favorable seizure outcome, with two seizure free. MRI scans at 6- to 9-months' follow-up disclosed discrete areas of atrophy in the amygdala and hippocampus, interspersed with preserved brain in the group I patients. More uniform and complete destruction of amygdala and hippocampus was evident in group II patients. All lesions were confined to the amygdala and hippocampus, sparing the parahippocampal gyrus (PHG). CONCLUSIONS: The extensive amygdalohippocampal ablation in group II patients improved seizure outcome compared with more limited ablation in group I, but these results were not so good as those from temporal lobectomy in a similar patient group. When considered together with the results of selective amygdalohippocampectomy, and temporal resections that spare hippocampus or amygdala (all producing similar outcomes, and all involving resection of the entorhinal cortex), this study suggests a pivotal role of the entorhinal cortex in temporal epileptogenesis. PMID- 10528938 TI - Longitudinal follow-up in 145 patients with medically refractory temporal lobe epilepsy treated surgically between 1984 and 1995. AB - PURPOSE: There are few studies of prolonged longitudinal follow-up after temporal resections. METHODS: We analyzed 145 consecutive patients with temporal lobe epilepsy treated surgically. Patients had a comprehensive presurgical evaluation, including video-EEG, psychometric testing, magnetic resonance imaging (MRI), single-photon emission computed tomography (SPECT), intracarotid amobarbital procedure (IAP), and recently, volumetric head MRIs and F-fluorodeoxyglucose positron emission tomography (FDG-PET) scans. Most had en bloc temporal resections, and a few had lesionectomies and resection of the epileptogenic zone. There was no surgical mortality. Longitudinal follow-up data of the seizure outcome were analyzed by actuarial analysis. Patients were followed up at 6 weeks, 3 months, 6 months, and then on a yearly basis. The mean follow-up was 5.6 years. RESULTS: Sixty-six percent were seizure free at 1 year, 63% at 2 years, 60% at 5 years, and 55% at 10 years follow-up. Moreover, 85%, became seizure free for > or =2 at the time of last follow-up or had rare seizures. Patients who were seizure free for 1 and 2 years after surgery, had an 83% and 92% probability, respectively, of remaining seizure free at the time of last follow-up. Ninety-one percent of patients with small tumors and cavernous angiomas became seizure free compared with 69% of patients with hippocampal sclerosis. CONCLUSIONS: Actuarial analysis showed that the long-term surgical outcome of temporal lobe epilepsy remains favorable. Follow-up at 1 and 2 years is highly predictive of the long term outcome. Patients with discrete lesions had the best outcome. Most of the patients with late recurrences had hippocampal sclerosis or temporal lobe gliosis. Some patients with postoperative seizures eventually became seizure free, reflecting the running-down phenomenon. PMID- 10528939 TI - Combined measurements of hippocampal N-acetyl-aspartate and T2 relaxation time in the evaluation of mesial temporal lobe epilepsy: correlation with clinical severity and memory performances. AB - PURPOSE: In this study we tried to find a correlation between the clinical severity and memory performances, by comparing proton magnetic resonance (MR) spectroscopy and T2 relaxation time measurements in the hippocampi, in a homogeneous group of 27 patients with unilateral mesial temporal lobe epilepsy with ipsilateral hippocampal sclerosis on MR imaging, with a view to answer the following questions: (a) how sensitive is this approach for the assessment of the apparently normal contralateral hippocampus, (b) do the results relate to the clinical severity, and (c) does it allow evaluation of the degree of hippocampal dysfunction. METHODS: Volume-selective proton MR spectroscopy of the head of both hippocampi was performed at 3 T, by using the PRESS sequence, with an echo time of 135 ms, to estimate NAA/(Cho + Cr) ratios. The relaxation times were measured at 0.28 T, by using a conventional Carr-Purcell-Meiboom-Gill sequence, with a repetition time of 2,000 ms, an echo time of 15 ms, and 48 echoes. RESULTS: The combination of NAA/(Cho + Cr) ratio and T2 relaxation time values was allowed to classify contralateral hippocampus abnormalities in two groups: first, decreased NAA/(Cho + Cr) ratio with strongly increased T2 relaxation time values corresponding to abnormalities observed in sclerotic ipsilateral hippocampi; and second, decreased NAA/(Cho + Cr) ratio with normal or slightly increased T2 relaxation time values. Whereas the NAA/(Cho + Cr) ratio or T2 relaxation time value alone was not correlated with memory performances, their association shows that left hippocampal injury evaluated both by NAA and T2 relaxation time measurements was clearly correlated with verbal memory scores, and right hippocampal injury, with visual memory scores. On the other hand, the maximal seizure frequency reported by the patients was correlated with ipsilateral NAA/(Cho + Cr) ratio and T2 relaxation time values but not with contralateral results. CONCLUSIONS: We showed that the combination of NAA and T2 relaxation time measurements can be used to examine the degree of ipsi- and contralateral hippocampal dysfunction or injuries and their relations with memory performances in the presurgical evaluation of patients. PMID- 10528940 TI - Effects of vigabatrin on the GABAergic system as determined by [123I]iomazenil SPECT and GABA MRS. AB - PURPOSE: To evaluate effects of vigabatrin (VGB) by using [123I]iomazenil single photon emission computed tomography (SPECT) to estimate central gamma aminobutyric acid (GABA(A))/benzodiazepine receptors (BZRs), and magnetic resonance spectroscopy (MRS) to assess tissue GABA levels. METHODS: Six patients with partial seizures had both SPECT and MRS before and 25-84 days after starting VGB (3 g p.o., q.d.). SPECT was acquired by using the constant-infusion method and, after nonuniform attenuation correction, coregistered with T1-weighted MR Imaging (MRI) A volume of interest (VOI) of 3 x 2 x 2 cc over the occipital cortex, used for MRS acquisition, was positioned on both MRI and coregistered SPECT. Occipital activity was divided by either total plasma activity or plasma [123I]iomazenil concentration to estimate BZR distribution volume (V(T)-p and V'(T), respectively). Wilcoxon's test was used for VOI differences in GABA levels, BZR V(T)-p or V'(T). SPM96 (either no global normalization or proportional scaling) was used to compare BZR V(T)-p changes in the patients with and without VGB with test-retest data in eight healthy age-matched controls. RESULTS: Occipital GABA levels were increased threefold (without VGB, 1.1+/-0.1 micromol/g; with VGB, 2.9+/-0.5 micromol/g; p = 0.027). BZR distribution volumes showed no change, when estimated by either V(T)-p (without VGB, 6.00+/-0.91 ml/g; with VGB, 5.86+/-0.44 ml/g; p = 0.92) or V(T) (without VGB, 41.1+/-11.2 ml/g; with VGB, 41.2+/-9.9 ml/g; p = 0.75). No significant changes were detected by SPM96. CONCLUSIONS: A clinically effective dose of VGB caused a threefold increase in tissue GABA levels but was not associated with a substantial BZR downregulation. PMID- 10528941 TI - The long-term use of gabapentin, lamotrigine, and vigabatrin in patients with chronic epilepsy. AB - PURPOSE: To compare the long-term retention of gabapentin (GBP), lamotrigine (LTG), and vigabatrin (VGB) by patients with chronic epilepsy and the reasons for treatment discontinuation. To assess the likelihood of seizure freedom, seizure related injury/hospital admission and mortality after these drugs were commenced. METHODS: This was a retrospective case-records survey in five tertiary referral epilepsy centres in the U.K. The retention times on treatment (from initiation to discontinuation) for the different antiepileptic drugs (AEDs) were compared by using Kaplan-Meier survival analysis and Cox regression. Incidences of seizure freedom and seizure-related injury/hospital admissions and standardised mortality ratios were calculated. RESULTS: There were 1,375 patients with chronic epilepsy included; 361 were taking GBP, 1,050 LTG, and 713 VGB. The retention of GBP, LTG, or VGB was <40% at 6 years. Fewer than 4% of patients become seizure free while taking one of the drugs. There was no reduction in mortality or seizure-related injury/admission. CONCLUSIONS: The impact of these new AEDs on chronic epilepsy can be described only as modest. This view may be revised, however, as more experience is gained with new drugs in previously untreated patients. PMID- 10528942 TI - Two visual mechanisms of photosensitivity. AB - PURPOSE: Photosensitive epilepsy is the most common of the "reflex" epilepsies. Precipitated by television viewing, flickering light, or specific visual patterns, it is the cause of seizures in 10% of young people with epilepsy. Photosensitivity is associated with two types of EEG abnormalities: photoparoxysmal responses (PPRs) and occipital spikes (OSs). It is unclear whether these abnormalities are mediated by different mechanisms, and furthermore, the clinical significance of OS is unknown. METHODS: By using our previously established population of patients with photosensitive epilepsy, all showing EEG abnormalities on intermittent photic stimulation or pattern stimulation, we examined the effects of pattern contrast, spatial and counterphase temporal frequency, and colour on these abnormalities. RESULTS: PPRs and not OSs show linear contrast dependency and are elicited by stationary stimuli and by non-colour-opponent isoluminant stimuli. CONCLUSIONS: PPRs and OSs are generated independently by the parvocellular and magnocellular visual systems, respectively. The results add support to the hypothesis that only PPRs and not OSs are clinically significant. PMID- 10528943 TI - Bradycardia and asystole with the use of vagus nerve stimulation for the treatment of epilepsy: a rare complication of intraoperative device testing. AB - PURPOSES: A 56-year-old man with mild mental retardation, right congenital hemiparesis, and refractory partial seizures was referred for vagus nerve stimulation (VNS). METHODS: Routine lead diagnostic testing during the surgical procedure (1.0 mA, 20 Hz, and 500 micros, for approximately 17 s) resulted, during the initial two stimulations, in a bradycardia of approximately 30 beats/min. A third attempt led to transient asystole that required atropine and brief cardiopulmonary resuscitation. RESULTS: The procedure was immediately terminated, the device removed, and the patient recovered completely. A postoperative cardiologic evaluation, including an ECG, 24-h Holter monitor, echocardiogram, and a tilt-table test, was normal. CONCLUSIONS: Possible mechanisms for the bradycardia/asystole include stimulation of cervical cardiac branches of the vagus nerve either by collateral current spread or directly by inadvertent placement of the electrodes on one of these branches; improper plugging of the electrodes into the pulse generator, resulting in erratic varying intensity of stimulation; reverse polarity; and idiosyncratic-type reaction in a hypersusceptible individual. The manufacturer reports the occurrence rate in approximately 3,500 implants for this intraoperative event to be approximately one in 875 cases or 0.1%. PMID- 10528944 TI - Ictal video-EEG recording of three partial seizures in a patient with the benign infantile convulsions associated with mild gastroenteritis. AB - PURPOSE: In infants, benign convulsions can be triggered by febrile illness or mild diarrhea such as Rotavirus gastroenteritis. The triggering mechanism of these convulsions is still unknown. In spite of several reports concerning clinical features, the ictal EEG recordings were rarely analyzed by a video-EEG monitoring system. To reveal a clue for the triggering mechanism of these convulsions, we analyzed the correlation of clinical manifestations and the EEG discharges during the ictal events and compared with previous reports. METHODS: The ictal EEG of a cluster of three afebrile convulsions associated with mild gastroenteritis was recorded by an EEG closed-circuit TV (EEG-CCTV) monitoring system in a 6-month-old healthy female infant. RESULTS: All seizures began as complex partial seizures (CPSs), which exhibited a motionless stare with or without leftward deviation of both eyes, and evolved to secondarily generalized tonic-clonic seizures (SGTCSs) for approximately 90 s. Each of three ictal discharges began from the right occipital, right centroparietotemporal, and left occipital regions, respectively. CONCLUSIONS: Although initiating sites of ictal discharges of benign infantile convulsions associated with mild gastroenteritis (BICE) were previously reported to be variable among patients, these results indicated that those differ among seizures even in a same infant. PMID- 10528945 TI - Multimodal MR imaging: functional, diffusion tensor, and chemical shift imaging in a patient with localization-related epilepsy. AB - PURPOSE: To demonstrate the integration of complementary functional and structural data acquired with magnetic resonance imaging (MRI) in a patient with localization-related epilepsy. METHODS: We studied a patient with partial and secondarily generalized seizures and a hemiparesis due to a malformation of cortical development (MCD) in the right hemisphere by using EEG-triggered functional MRI (fMRI), diffusion tensor imaging (DTI), and chemical shift imaging (CSI). RESULTS: fMRI revealed significant changes in regional blood oxygenation associated with interictal epileptiform discharges within the MCD. DTI showed a heterogeneous microstructure of the MCD with reduced fractional anisotropy, a high mean diffusivity, and displacement of myelinated tracts. CSI demonstrated low N-acetyl aspartate (NAA) concentrations in parts of the MCD. CONCLUSIONS: The applied MR methods described functional, microstructural, and biochemical characteristics of the epileptogenic tissue that cannot be obtained with other noninvasive means and thus improve the understanding of the pathophysiology of epilepsy. PMID- 10528946 TI - Relationship between cysticercosis and epilepsy. PMID- 10528947 TI - Problem of managing patients with epilepsy in developing countries. PMID- 10528948 TI - Change of editor--changing times. PMID- 10528949 TI - Clinical interventions and outcomes of One-to-One midwifery practice. AB - BACKGROUND: Changing Childbirth became policy for the maternity services in England in 1994 and remains policy. One-to-One midwifery was implemented to achieve the targets set. It was the first time such a service had been implemented in the Health Service. An evaluation was undertaken to compare its performance with conventional maternity care. METHODS: This was a prospective comparative study of women receiving One-to-One care and women receiving the system of care that One-to-One replaced (conventional care) to compare achievement of continuity of carer and clinical outcomes. The evaluation took place in The Hammersmith Hospitals NHS Trust, the Queen Charlotte's and Hammersmith Hospitals. This was part of a larger study, which included the evaluation of women's responses, cost implications, and clinical standards and staff reactions. The participants were all those receiving One-to-One midwifery practice (728 women), which was confined to two postal districts, and all women receiving care in the system that One-to-One replaced, in two adjacent postal districts (675 women), and expecting to give birth between 15 August 1994 and 14 August 1995. Main outcome measures were achievement of continuity of care, rates of interventions in labour, length of labour, maternal and infant morbidity, and breastfeeding rates. RESULTS: A high degree of continuity was achieved through the whole process of maternity care. One-to-One women saw fewer staff at each stage of their care, knew more of the staff who they did see, and had a high level of constant support in labour. One-to-One practice was associated with a significant reduction in the use of epidural anaesthesia (odds ratio (OR) 95 per cent confidence interval (CI) = 0.59 (0.44, 0.80)), with lower rates of episiotomy and perineal lacerations (OR 95 per cent CI = 0.70 (0.50, 0.98)), and with shorter second stage labour (median 40 min vs 48 min). There were no statistically significant differences in operative and assisted delivery or breastfeeding rates. CONCLUSIONS: This study confirms that One-to-One midwifery practice can provide a high degree of continuity of carer, and is associated with a reduction in the rate of a number of interventions, without compromising safety of care. It should be extended locally and replicated in other services under continuing evaluation. PMID- 10528950 TI - Hip fracture in Hong Kong over the last decade--a comparison with the UK. AB - BACKGROUND: Hip fracture is a major public health problem in Asia and the UK. The objectives of this study were to describe the trends of hip fracture in Hong Kong over the last decade, and to compare the incidence in Hong Kong with that from the Wessex Health Region of the UK in 1995. METHODS: The number of hip fractures was calculated using hospital discharge records for all public hospitals in Hong Kong in 1991 and 1995. Age-specific incidence rates were then calculated using the mid-year census population for the two years. These rates were presented with previously reported age-specific rates for Hong Kong in 1966 and 1985. These age specific rates for Hong Kong in 1995 were compared with rates for the Wessex Health Region of the UK. The total number of hip fracture expected in 2010 was calculated by applying the age-specific rates of 1995 to the projected population for 2010. RESULTS: In 1995, a total of 1138 men and 2782 women in Hong Kong fractured their hip. The age-specific rates had remained static from 1985 to 1995, after substantial rise from 1966 to 1985. In 1995, the rates of hip fracture rates were 11/1000 in women and 5/1000 in men who were 70 years and older. These rates were almost identical to those observed in the Wessex Health Region of the UK. CONCLUSION: The age-specific incidence rates of hip fracture had not risen in Hong Kong in the last decade. The incidence of hip fracture in Hong Kong was similar to that in the UK in 1995. The total number of patients with hip fracture in Hong Kong will increase substantially in the future, as a result of the ageing of the population. PMID- 10528951 TI - Controlled management of public relations following a public health incident. AB - This paper describes the management of public relations following an outbreak of multidrug resistant TB at a London hospital. Eight patients were involved, all of the secondary cases occurred in HIV seropositive patients, and three cases died. The paper describes how the the Incident Committee undertook to recall contacts of the cases for screening, inform the general practitioners of all of the contacts about their patients' exposure, warn other organizations and professionals interested or involved in the management of HIV in the London area as to the nature of the incident, and establish a helpline, before informing a wider audience through the EPINET, Communicable Disease Report and national press. PMID- 10528952 TI - Short Form 36 (SF-36) Health Survey questionnaire: which normative data should be used? Comparisons between the norms provided by the Omnibus Survey in Britain, the Health Survey for England and the Oxford Healthy Life Survey. AB - BACKGROUND: Population norms for the attributes included in measurement scales are required to provide a standard with which scores from other study populations can be compared. This study aimed to obtain population norms for the Short Form 36 (SF-36) Health Survey Questionnaire, derived from a random sample of the population in Britain who were interviewed at home, and to make comparisons with other commonly used norms. METHODS: The method was a face-to-face interview survey of a random sample of 2056 adults living at home in Britain (response rate 78 per cent). Comparisons of the SF-36 scores derived from this sample were made with the Health Survey for England and the Oxford Healthy Life Survey. RESULTS: Controlling for age and sex, many of mean scores on the SF-36 dimensions differed between the three datasets. The British interview sample had better total means for Physical Functioning, Social Functioning, Mental Health, Energy/Vitality, and General Health Perceptions. The Health (interview) Survey for England had the lowest (worst) total mean scores for Physical Functioning, Social Functioning, Role Limitations (physical), Bodily Pain, and Health Perceptions. The postal sample in central England had the lowest (worst) total mean scores for Role Limitations (emotional), Mental Health and Energy/Vitality. CONCLUSION: Responses obtained from interview methods may suffer more from social desirability bias (resulting in inflated SF-36 scores) than postal surveys. Differences in SF-36 means between surveys are also likely to reflect question order and contextual effects of the questionnaires. This indicates the importance of providing mode specific population norms for the various methods of questionnaire administration. PMID- 10528954 TI - Genetically modified organisms: an analysis of the regulatory framework currently employed within the European Union. AB - BACKGROUND: Genetic engineering technology is starting to bring many commercial products to the market. These genetically modified organisms (GMOs) and their derived products are subject to topical debate as to their benefits and risks. The strengths and weaknesses of the regulatory framework that controls their development and application is central to the question of whether this technology poses significant risk to the public health during this critical phase of its evolution. METHODS: A critical review was carried out of the legal framework regulating the contained use, deliberate release and some aspects of consumer protection relevant to the control of GMOs in Europe and the United Kingdom. RESULTS: The current legal framework is failing to provide a speed of adaptation commensurate with the development of the science of genetic engineering; failing to properly respond to democratic control; failing to resolve significant conflict between the protection of free markets and protection of public health and the environment; and failing to implement obligations on biodiversity. CONCLUSION: The present legal framework must be replaced. Current European Union proposals for new standards of regulation are welcome, but provide only for further incremental change, and do not address some significant fundamental flaws in our current laws. PMID- 10528953 TI - Surveillance of problem drug use in the UK: a review of a Regional Drug Misuse Database. AB - BACKGROUND: We report detailed findings of the first systematic validation of a Regional Drug Misuse Database (RDMD); such databases constitute the main investment in routine drug statistics in the UK by the Department of Health. METHODS: A retrospective case-finding study in a stratified random sample of one in three specialist drug agencies was carried out. Agency records of clients attending during 1994 were matched with reports (episodes) to the North Thames RDMD to assess the level of under-reporting, and the relationship between RDMD reports (episodes) and the number of problem drug users in contact with agencies. Under-ascertainment of cases was estimated using two-sample capture-recapture. RESULTS: Under-reporting was associated with agency records missing full date of birth or initials (attributers), and agency type. Compared with drug dependency units (DDU) the odds of under-reporting were 3-18 times higher by the other specialist drug agencies. Even after excluding episodes with missing attributers the odds ratio (OR) of not being reported was significantly higher among needle exchanges (OR 2.7), non-statutory community based drug teams (OR 3.2), statutory community based drug teams (OR 4.9) and residential rehabilitation units (OR 8.7) compared with DDUs. Overall database episodes represented 60 per cent of the number of clients attending specialist agencies as a result of a mixture of under reporting and the proportion of clients retained in treatment, which also varied by agency type. A total of 727 individuals (16 per cent) had never been reported. CONCLUSIONS: Surveillance of drug misuse through RDMDs does not yet fulfil its objectives. It is essential that a system of following up reports is introduced to improve their utility, and to contribute to the monitoring of the UK Government's new drugs strategy, and wider European surveillance. PMID- 10528955 TI - Social determinants of GHQ score by postal survey. AB - BACKGROUND: To develop interventions to reduce the morbidity associated with depression and anxiety, more information is needed about the social and demographic determinants of these disorders and the relative contributions of different potential predictors. METHODS: Using stratified sampling from the Family Health Services Authority (FHSA) register, postal surveys were sent to 61,000 adults across the North Western Regional Health Authority. Psychological morbidity was assessed using the 12-item General Health Questionnaire (GHQ). Nine potential predictors of morbidity were rated, including socio-demographic details and the presence of longstanding limiting physical illness and of a confidante. Logistic regression analyses were used to consider each of the nine potential predictors separately and in combination. RESULTS: A total of 38,014 questionnaires were returned (63 per cent). After adjustment for all other variables the strongest predictors of a high GHQ score were the absence of a confidante (odds ratio (OR) 3.64), longstanding limiting physical illness (OR 2.93), unemployment (OR 1.91), being a student (OR 1.78), being female (OR 1.64), single parenthood (OR 1.55) and living alone (OR 1.32). GHQ scores were highest in the 18-34 age range. Ethnicity exerted no significant effect after adjustment for other variables. CONCLUSION: In keeping with other research the data suggest that sociodemographic factors are strong predictors of depression and anxiety. The most vulnerable population groups are those with longstanding limiting physical illness and no-one to talk to. This should help in identifying high-risk individuals and informing preventive strategies. PMID- 10528956 TI - A national facility for small area disease mapping and rapid initial assessment of apparent disease clusters around a point source: the UK Small Area Health Statistics Unit. AB - BACKGROUND: Reports of disease clusters are often received by district health authorities and are, in some cases, associated with concerns about a pollution source. The Small Area Health Statistics Unit (SAHSU) has developed a Rapid Inquiry Facility, which will produce an estimated relative risk for any given condition for the population within defined areas around a point source, relative to the population in a local reference region. The system can also facilitate the production of annual reports and other health studies for Departments of Public Health Medicine through the creation of ward-level maps to illustrate disease variation across small areas. METHODS: The facility uses routinely collected morbidity, mortality and population data at a small area scale, together with the computing facilities and expertise necessary to run such analyses quickly and efficiently. Using this facility SAHSU can supply a report within three working days. To aid interpretation, smoothed small area maps that account for sampling variability in the observed data can also be produced. RESULTS: The paper reports on two case studies where the pilot system has been utilized by health authorities for both point source analyses and small area disease mapping. CONCLUSIONS: We believe that this facility would be of considerable use to districts. The local knowledge and expertise of the local public health specialist is essential in the interpretation and presentation of the facility's output. Feedback from public health specialists is helping SAHSU refine the output of the facility, so as to make the information presented as comprehensive and as useful as possible. PMID- 10528957 TI - Clinical information for research; the use of general practice databases. AB - General practice computers have been widely used in the United Kingdom for the last 10 years and there are over 30 different systems currently available. The commercially available databases are based on two of the most widely used systems -VAMP Medical and Meditel. These databases provide both longitudinal and cross sectional data on between 1.8 and 4 million patients. Despite their availability only limited use has been made of them for epidemiological and health service research purposes. They are a unique source of population-based information and deserve to be better recognized. The advantages of general practice databases include the fact that they are population based with excellent prescribing data linked to diagnosis, age and gender. The problems are that their primary purpose is patient care and the database population is constantly changing, as well as the usual problems of bias and confounding that occur in any observational studies. The barriers to the use of general practice databases include the cost of access, the size of the databases and that they are not structured in a way that easily allows analysis. Proper utilization of these databases requires powerful computers, staff proficient in writing computer programs to facilitate analysis and epidemiologists skilled in their use. If these structural problems are overcome then the databases are an invaluable source of data for epidemiological studies. PMID- 10528958 TI - Evaluating computerized health information systems: hardware, software and human ware: experiences from the Northern Province, South Africa. AB - Despite enormous investment world-wide in computerized health information systems their overall benefits and costs have rarely been fully assessed. A major new initiative in South Africa provides the opportunity to evaluate the introduction of information technology from a global perspective and assess its impact on public health. The Northern Province is implementing a comprehensive integrated hospital information system (HIS) in all of its 42 hospitals. These include two mental health institutions, eight regional hospitals (two acting as a tertiary complex with teaching responsibilities) and 32 district hospitals. The overall goal of the HIS is to improve the efficiency and effectiveness of health (and welfare) services through the creation and use of information, for clinical, administrative and monitoring purposes. This multi-site implementation is being undertaken as a single project at a cost of R130 million (which represents 2.5 per cent of the health and welfare budget on an annual basis). The implementation process commenced on 1 September 1998 with the introduction of the system into Mankweng Hospital as the pilot site and is to be completed in the year 2001. An evaluation programme has been designed to maximize the likelihood of success of the implementation phase (formative evaluation) as well as providing an overall assessment of its benefits and costs (summative evaluation). The evaluation was designed as a form of health technology assessment; the system will have to prove its worth (in terms of cost-effectiveness) relative to other interventions. This is more extensive than the traditional form of technical assessment of hardware and software functionality, and moves into assessing the day-to-day utility of the system, the clinical and managerial environment in which it is situated (humanware), and ultimately its effects on the quality of patient care and public health. In keeping with new South African legislation the evaluation process sought to involve as many stakeholders as possible at the same time as creating a methodologically rigorous study that lived within realistic resource limits. The design chosen for the summative assessment was a randomized controlled trial (RCT) in which 24 district hospitals will receive the HIS either early or late. This is the first attempt to carry out an RCT evaluation of a multi-site implementation of an HIS in the world. Within this design the evaluation will utilize a range of qualitative and quantitative techniques over varying time scales, each addressing specific aims of the evaluation programme. In addition, it will attempt to provide an overview of the general impact on people and organizations of introducing high-technology solutions into a relatively unprepared environment. The study should help to stimulate an evaluation culture in the health and welfare services in the Northern Province as well as building the capacity to undertake such evaluations in the future. PMID- 10528959 TI - What is the cost of getting the price wrong? AB - BACKGROUND: The objective of this study was to compare differences in cost estimates for paediatric HIV hospital service provision based on hospital prices with cost estimates obtained through a research-based service-specific costing exercise. METHODS: Activity data on the use of hospital services of children by stage of HIV infection were collected from case-notes for 118 HIV antibody positive children, managed at St Mary's Hospital NHS Trust, London, 1 January 1986-31 December 1994. Hospital unit prices were obtained from the Hospital Trust Finance Department; unit cost estimates were obtained from relevant hospital departments through a research-based service-specific costing exercise. Financial data related to the 1993-1994 financial year, and were indexed to 1995-1996 prices. The main outcome measures were cost estimates per patient-year by stage of HIV infection. Three cost scenarios were calculated: first by linking activity data with hospital prices (Trust Prices); second by linking activity data with routinely available hospital prices plus units costs from the costing exercise where no relevant hospital prices existed (Supplemented Trust Prices); third, by linking activity data exclusively with unit costs from the hospital-specific costing exercise (Unit Costs). RESULTS: There were substantial differences between unit cost estimates per patient-year based on Trust Prices and Supplemented Trust Prices compared with those based on Unit Costs. Differences increased with more intense use of services. The deficit based on Trust Prices compared with Unit Costs ranged from Pound Sterling 432 per patient-year for HIV negative children, Pound Sterling 574 for asymptomatic HIV-infected children, Pound Sterling 1288 for indeterminate children, Pound Sterling 1814 for children with symptomatic non-AIDS to Pound Sterling 7418 per patient-year for children with AIDS. CONCLUSIONS: In this hospital, reliance on generic hospital prices to derive cost estimates for paediatric HIV services produced considerable underestimates of the cost of service provision compared with data derived through the costing exercise. If this occurs across all or most areas of service provision, this can lead to substantial financial deficits, which in turn may mean that the needs of specific client populations may not be met. PMID- 10528960 TI - A clinical trial of 'AM', a Ugandan herbal remedy for malaria. AB - BACKGROUND: Mortality and morbidity from malaria is still high in Africa, and may further increase as resistance to antimalarial drugs spreads. Many people rely on herbal medicines as the first line of treatment. Yet there has been very little clinical research into their effectiveness. METHODS: Patients being treated for malaria at a herbalists' clinic in South-West Uganda were followed up and their response to a particular herb, 'AM', was monitored. Eighty-eight patients were enrolled; 72 were followed up for at least 2 days, and were questioned about side effects. Nineteen patients infected with Plasmodium falciparum had initial parasite counts sufficiently high for parasite clearance to be assessed. RESULTS: No severe adverse reactions were observed, although about 50 per cent experienced minor side-effects. Although complete parasite clearance was achieved in only one case, the geometric mean of parasite counts had declined significantly by day 7. There was also a marked symptomatic improvement in 17 of the 19 patients. CONCLUSIONS: AM appears safe, although it is not always well tolerated. Significant symptomatic improvement and a reduction of parasite counts were observed in patients taking AM. There is a need for further research, such as a randomized controlled trial, to assess the efficacy of this treatment. PMID- 10528961 TI - Entitlement to military healthcare--limitations of the NHS model. AB - The Defence Medical Services provide to a British population healthcare services that are funded from taxation and are free at the point of delivery. This paper reviews some principles for determining entitlement to healthcare for the population cared for by the Defence Medical Services. The starting point for entitlement uses the principles under which the National Health Service (NHS) was established. These are then extended to acknowledge the limitations of an NHS model when considering occupational health issues and geographical variations in healthcare provision. PMID- 10528962 TI - The impact of climate on the prevalence of respiratory tract infections in early childhood in Lahore, Pakistan. AB - BACKGROUND: Respiratory tract infections are a major health problem in developing countries. The aim of this study was to analyse the impact of the climate on the prevalence of upper respiratory tract infection (URTI) and lower respiratory tract infection (LRTI) in four socioeconomically different groups in a developing country. METHODS: A prospective cohort study was conducted among children in four socioeconomically different groups in Lahore, Pakistan. Monthly observations were made on 1476 infants born during 1984-1987 and followed for 24 months. Prevalence of URTI and LRTI was analysed according to age, area of living, family size, time of birth, the season of the year and climate variables such as rain, temperature and humidity. RESULTS: Low monthly average minimum day temperature was associated with high prevalence of URTI and LRTI. For LRTI the impact of temperature was larger for boys, children living in larger families and children living in the poorer areas. This pattern was not seen for URTI. A peak in prevalence for LRTI was shown at 5-6 months of age for LRTI and at 10-12 months of age for URTI. CONCLUSIONS: Temperature is related to prevalence of URTI and LRTI in a developing society. The effect of temperature on health varies between different subgroups. These effects should be considered in planning health actions to prevent respiratory tract infections. PMID- 10528963 TI - How does the prevalence of specific morbidities compare with measures of socio economic status at small area level? AB - BACKGROUND: Evidence from other studies has show large, systematic differences between the health of social groups. It is not clear whether this relationship applies equally to all areas of health need. We assess whether a variety of areas of ill health show positive correlations with increasing socioeconomic disadvantage, and whether there are indicators of socio-economic disadvantage that are better than others at predicting the prevalence of specific morbidities at a population level. METHODS: The prevalence of a range of common morbidities was determined by a postal questionnaire sent to 16,750 subjects (response rate 79 per cent), and compared with socio-economic information obtained from the 1991 Census. RESULTS: There was substantial variation in the degree to which the various morbidities were related to the socioeconomic variables. When compared with socio-economic variables, long-term limiting illness, respiratory conditions and depression had high correlations of +0.8 or more. Cardiovascular conditions were less related (r = +0.60 to +0.79). None of the disorders of the gastrointestinal system showed a high correlation with socio-economic variables. There was also substantial variation in the degree of correlation of the socio economic measures with each area of morbidity. The measures that showed the highest correlations were in respect of household characteristics such as car ownership and single parent households. Variables describing household amenities such as lacking a bath or central heating were least related to the morbidity measures. CONCLUSIONS: Some areas of morbidity show strong associations with socio-economic disadvantage, but others show only modest or no relationship. The optimum choice of socio-economic variable as a proxy for health need depends on the area of illness being considered. PMID- 10528964 TI - Quarterly communicable disease review January to March 1999. From the PHLS Communicable Disease Surveillance Centre. Public Health Laboratory Service. PMID- 10528965 TI - Delay in diagnosis of tuberculosis: of remaining concern in England and Wales. PMID- 10528966 TI - Quality indicators for general practice. PMID- 10528967 TI - Objectives of genetic counselling: differing views of purchasers, providers and users. PMID- 10528968 TI - Episiotomy and peritoneal tears in low-risk UK primigravidae. PMID- 10528969 TI - Occupation and risk of hip fracture. PMID- 10528970 TI - Screening: completeness of population registers a problem for some sub-groups. PMID- 10528971 TI - Clinical testing of restorative materials: some historical landmarks. AB - OBJECTIVES: The aim of the study was to review the historical background of the methodologies currently in use for clinical investigations of contemporary tooth coloured dental restorative materials and techniques, and to attempt to identify the likely general direction of such research projects in the future. DATA SOURCES: Published articles obtained as a result of a wide ranging literature search of both dental and medical journals. STUDY SELECTION: Historical and contemporary accounts of clinical trial management, upon which an overview of the key events important in the evolution of clinical methods of evaluation of dental restorative materials, and a prediction of possible future trends in this area, could be based. CONCLUSIONS: Publication of dental restoratives clinical research has not kept up with that in medicine, where editors of leading journals require that clinical trials be run in accordance with the Consolidated Standards of Reporting Trials (CONSORT) guidelines. The current trend towards evidence based treatment in dentistry can be expected to initiate more statistically powerful clinical studies of dental restoratives, and the merit of involving general dentists in practice based research can also be anticipated to become increasingly recognised in the future. PMID- 10528972 TI - A 4-year clinical study on amalgam, resin composite and resin-modified glass ionomer cement restorations in overdenture abutments. AB - OBJECTIVE: To assess the performance of three different filling materials in overdenture abutment teeth. METHODS: In 49 patients amalgam, resin composite and resin-modified glass ionomer cement were used to seal the root canal orifices of 155 overdenture abutment teeth. After initial preparation of the cavities, the three restorative materials were randomly assigned to the abutment teeth using a number of balancing criteria. All patients were reviewed every six months and received the same preventive regimen. Survival was assessed at two levels: Sorig (survival of the restoration independent from eventual maintenance treatments) and Scomp (restorations survived even without maintenance treatments). RESULTS: The calculated overall survival percentage of the original restorations (Sorig) after four years was 63 +/- 6% (mean +/- SE). Calculation for the overall complete survival (Scomp) revealed a percentage survival of 57 +/- 6%. At both levels, the differences between the survivals of the investigated materials were not statistically significant (p-values > 0.05). Two abutments were lost, severe caries was the reason for one extraction and another abutment tooth was extracted for periodontal reasons. CONCLUSIONS: The results of this study did not point out a superior restorative material for the seal of root canal orifices of overdenture abutments. The distribution of failures over the patients indicates a certain patient dependency. PMID- 10528973 TI - Human pulp response after an adhesive system application in deep cavities. AB - OBJECTIVES: To evaluate the pulpo-dentin complex response to a dentin adhesive application in deep cavities performed in human teeth. METHODS: Deep class V cavities were prepared on the buccal surface of 46 premolars. The remaining dentin of the axial wall received 10% phosphoric acid and dentin adhesive (group DA), or was protected before the acid and dentin adhesive application with calcium hydroxide cement (group CH). Half of the teeth, which received the acid application directly over the axial wall, were contaminated prior to the procedures with dental plaque collected from the patient's own teeth (group DAC). The plaque was placed on the dentin for 5 min and then the cavity was washed. All teeth were restored with a light-cured composite resin. The teeth were extracted after 7, 30 or 60 days and prepared according to normal histologic techniques. Serial sections were stained with H/E, Masson's trichrome and Brown & Brenn technique for demonstration of bacteria. RESULTS: The histopathologic evaluation showed that in groups DA and DAC, the inflammatory response was more evident than in group CH. Also, the intensity of the pulp reaction increased as the remaining dentin thickness decreased. There was no statistical difference in the inflammatory response between the groups DA and DAC. CONCLUSION: Based on the experimental conditions, we concluded that the All Bond 2 adhesive system, when applied on dentin in deep cavities, showed an acceptable biocompatibility. However, the intensity of the pulpo-dentin complex response depends on the remaining dentin thickness. PMID- 10528974 TI - Factors influencing the selection of panoramic radiography in general dental practice. AB - OBJECTIVES: To identify factors influencing dentists' decisions to take panoramic radiographs and to determine dentists' perceptions of the value of panoramic radiographs in the diagnosis of common dental pathologies. METHODS: Questionnaire of dentists with access to panoramic radiography equipment in 22 randomly selected Family Health Service Authorities in England and Wales. Dentists were asked to score 17 factors for their influence upon panoramic use, compare the relative diagnostic value of panoramic and intraoral radiographs for diagnosis of common dental pathologies and state their principal reasons for taking panoramic radiographs. RESULTS: The response rate to the questionnaire was 73.3%. The factors most likely to influence dentists to take a panoramic radiograph were 'planning oral surgery', 'facial trauma', 'periodontal disease', 'heavily restored dentition' and 'patient first attendance'. Dentists' opinions on the diagnostic usefulness of panoramic radiographs were in broad agreement with those in the scientific literature. The main reasons for taking panoramic radiographs were as a 'general screen' and as a 'view for unerupted or impacted teeth'. CONCLUSIONS: There are areas where dentists' prescription of panoramic radiographs is in disagreement with recent guidelines. Successful implementation of these guidelines would be likely to lead to a substantial reduction in the numbers of panoramic radiographs taken by GDPs. PMID- 10528975 TI - Torque, power and efficiency characterization of dental air turbine handpieces. AB - OBJECTIVES: In a previous paper the flow and free running speed characteristics of dental air turbine handpieces were discussed. The present work was to continue the analysis and characterization of these handpieces by addressing the issues of torque, power generation, efficiency and the specification of figures of merit to aid comparative testing and selection. METHODS: Using the principle of the rope brake, torque was determined from stall to free running at a variety of supply pressures for fourteen models of handpiece. Using previous results for air flow rate and free running speed, power and efficiency were calculated. RESULTS: Stall torque was demonstrated to depend on rotor position. Dynamic torque was found to closely approach the expected linear relationship over the range from stall to free running. Several figures of merit (performance indices) were identified which might be used for routine rating and specification of handpieces. CONCLUSIONS: Standardized testing can now be performed and effective figures of merit derived to characterize the behaviour of dental air turbine handpieces. PMID- 10528976 TI - The influence of clinically induced variability on the bi-axial fracture strength of aluminous core porcelain discs. AB - OBJECTIVE: The proportions of the constituents required to produce a porcelain slurry used in crown construction are assessed by experience so that variability is inevitable. The present study investigated the strength dependence of porcelain specimens on slurry consistency. METHODS: Vitadur-N core porcelain was formed into discs and the mean fracture strength, standard deviation and associated Weibull Moduli (m) determined as a function of slurry consistency using bi-axial flexure (ball-on-ring) by fracturing sets of 30 specimens prepared to different powder contents for a constant 0.4 ml of liquid. RESULTS: The strength data for the porcelain discs showed variation in magnitude and consistency ranging from 145 +/- 16.5 MPa (m = 9.7 +/- 1.8) at 1.2 g to 155 +/- 13.8 MPa (m = 12.3 +/- 2.2) at 1.4 g and 142 +/- 13.7 MPa (m = 11.7 +/- 2.1) at 1.6 g of powder. Increasing or decreasing the powder content of the slurry (related to specimens condensed using 1.4 g of powder) resulted in an increase in the apparent porosity and a decrease in the apparent solid density. CONCLUSIONS: There was no significant decrease in the mean bi-axial flexure strengths for the slurry consistencies investigated. However, the reliability of the fracture strength data did vary significantly. The results suggested that an optimum consistency exists (wherein consistent reproducibility of test results was achieved) and appears to have implications for laboratory testing of materials since a comparison between materials can only be achieved if specimen preparation occurs consistently between test centres for all the materials under investigation. PMID- 10528977 TI - Particle-induced X-ray emission and scanning electron microscopic analyses of the effects of CO2 laser irradiation on dentinal structure. AB - OBJECTIVES: The purpose of this study was to analyze the physico-chemical changes present on the dentinal surface after using CO2 laser irradiation, and to determine whether or not it is possible to seal the dentinal tubules. METHODS: Thirty human-extracted first premolars were obtained for this study. A Class V cavity was prepared on the buccal surface of all the specimens with a carbide pear-shaped bur, using a conventional high speed handpiece. Fifteen premolars (experimental group) were irradiated with a CO2 laser (with a wavelength of 10.6 microm, 2 W, 10 J, 0.2 s, 25 pulses). The remaining 15 premolars were used as the control group. RESULTS: Scanning electron microscopy showed that the effect of laser energy on dentin varied from charring, cratering, poring, fissuring, fracturing and cracking up to melting; also, the dentinal tubules were not sealed, in contrast with the control group in which the dentinal surfaces were more homogeneous. Particle-induced X-ray emission results showed that the irradiated dentinal surface presented a decrease in calcium content and an increase in phosphorous content, possibly due to a vaporization process which occurred during the irradiation. CONCLUSION: The physicochemical changes observed on the irradiated dentinal surface suggest that changes in the hydroxyapatite crystal structure take place, and that these structural changes may be responsible for the observed effects. PMID- 10528978 TI - Relationships between the modulus of elasticity and tensile strength for pharmaceutical drugs and excipients. AB - The relationship between the modulus of elasticity and tensile strength for a variety of pharmaceutical drugs and excipients has been explored as a means of generating algorithms for use in computer simulations and expert systems. From literature data two equations have been developed to enable the formulator, from knowledge of one property, to predict the other with a reasonable degree of accuracy. Such relationships are invaluable for use in computer simulation programs or formulation expert systems, enabling formulators to screen potential formulations quickly without the need for extensive experimental work. PMID- 10528979 TI - A dissolution test for a pressure-controlled colon delivery capsule: rotating beads method. AB - The rotating beads method is a new in-vitro dissolution test proposed for drugs formulated as pressure-controlled colon delivery capsules (PCDCs). The apparatus consisted of a glass vessel (500 mL) containing 4-mm (i.d.) glass beads (5000, 10000 or 15000) and dissolution medium (0-067 M phosphate buffer, pH 7; 25, 50 or 100 mL) containing polyvinyl alcohol (PVA; 5, 10 or 20 w/v%) to simulate the viscosity of the colon. The vessel was rotated at 5, 10 or 25 rev min(-1) and the temperature was maintained at 37 degrees C. Fluorescein was used as a model drug to explore the optimized conditions under which differences in the drug dissolution rate are detected between colon delivery systems. Fluorescein was formulated in four types of colon delivery systems. One was a tablet coated with an enteric polymer, Eudragit S-100, and the other three were PCDCs prepared with different thicknesses of ethylcellulose coating membrane (type I, II, III). The dissolution behaviour of fluorescein from the PCDC formulation was significantly different from that of the Eudragit S-100-coated tablets, when the dissolution conditions were as follows: rotation speed, 10 rev min(-1); bead number, 10000; dissolution medium, 50mL with 10% PVA. This dissolution method was applied to acetaminophen sustained-release tablets and two other drugs having low solubility in the colon, tegafur and 5-aminosalicylic acid. Similarly, significant differences in the dissolution rates of drugs from the PCDC formulation and the enteric tablet were detected. There was good correlation between the in-vitro dissolution rates and in-vivo absorption rates using T50 (the time for half of the amount of drug to be released from the preparation) and Cmax/Tmax (enteric tablet) or Cmax/(Tmax-Ti) (PCDC), where Ti is the first appearance time in the systemic circulation. The rotating beads method is a valuable technique for evaluating the dissolution rate of drugs formulated in PCDC. PMID- 10528980 TI - Studies on the co-encapsulation, release and integrity of two subunit antigens: rV and rF1 from Yersinia pestis. AB - In the development of combination or multiple sub-unit vaccines, determination of the encapsulation, release and integrity of two or more proteins co-encapsulated within microspheres is an important issue. A new extraction method, which exhibits excellent protein recovery, has been developed which enables samples to be used for sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and subsequent measurement of individual antigens encapsulated within microspheres. Using the new method, the protein loading of poly-(L-lactide) microspheres co-encapsulating two plague sub-unit antigens was found to be 1.22% (w/w) for recombinant V antigen (rV) and 1.24% (w/w) for recombinant F1 (rF1) by SDS-PAGE. The total protein loading was 2.49% (w/w) by bicinchoninic acid assay. The individual release of the two subunit antigens from the co-encapsulated microspheres was determined by SDS-PAGE analysis and rF1 was found to have a higher burst release than rV. The integrity and immunological activity of both rF1 and rV antigens was shown to be unaffected by the microencapsulation process. This study shows that encapsulation of more than one antigen within poly-(L lactide) microspheres is a viable method for the delivery of intact proteins. PMID- 10528981 TI - The maximum pharmacodynamic effect as a response parameter: pharmacokinetic considerations. AB - In previous human in-vivo studies measuring the maximum pharmacodynamic response to characterize cutaneously applied ointment preparations, it was observed that differences between various formulations caused by penetration enhancement led to different enhancement factors depending on the method used for determination of these factors from activity-response curves. To clarify this discrepancy, pharmacokinetic simulations have been performed based on an open one-compartment model with either first- or zero-order drug penetration kinetics and first-order elimination kinetics. Under the assumption that the maximum pharmacodynamic response corresponds to the maximum effective drug concentration in the receptor compartment, which represents the difference between the maximum drug concentration and the threshold concentration, drug concentration vs time profiles and dose-response curves were simulated. In addition, maximum effective concentrations were calculated and plotted against the logarithm of the thermodynamic drug activity to obtain activity-response curves. Relative bioavailability and enhancement factors were determined either from the horizontal distance between the curves of a standard and a test preparation, or as the ratio of the maximum effective concentration of test and standard formulations. A significant difference between the first-order and the zero-order input kinetics with regard to the evaluation of bioavailability and drug penetration enhancement was shown. Under finite dose conditions, i.e. first-order input kinetics from solution-type preparations, a misestimation of the factors usually occurs. Only under infinite dose conditions, i.e. if large preparation volumes are applied to achieve zero-order input kinetics, is the determination of bioavailability and enhancement factors from dose- and activity-response curves accurate. PMID- 10528982 TI - Age-related alteration of carbamazepine-serum protein binding in man. AB - To determine whether biological maturation influences the kinetics of carbamazepine-serum protein binding, the carbamazepine free fraction (%) was investigated in the serum of 66 patients, ranging from 4 to 83 years, with epilepsy or trigeminal neuralgia, treated with carbamazepine alone or carbamazepine in combination with phenytoin, phenobarbital, and/or valproic acid, over a relatively long period. Biochemical parameters such as levels of albumin and non-glycated albumin showed a significant relationship with carbamazepine free fraction (r = -0.521, P < 0.001 for albumin; r = -0.700, P < 0.001 for non glycated albumin). Non-glycated albumin was more strongly correlated with carbamazepine free fraction. The biochemical parameters showed a significant relationship with age (r =-0.243, P < 0.1 for albumin; r =0.666, P < 0.001 for glycated albumin; r = -0.459, P < 0.001 for non-glycated albumin; r = 0.640, P < 0.001 for carbamazepine free fraction). Glycated albumin (%), non-glycated albumin and carbamazepine free fraction (%) were strongly correlated with age, whereas albumin showed only a weak correlation with age. To evaluate the effects of ageing on carbamazepine-serum protein binding, the patients were divided into three groups according to age: children, 4-15 years; adults, 16-64 years; elderly, 65-83 years. Albumin and non-glycated albumin were much lower, and glycated albumin (%) and carbamazepine free fraction (%) much higher in the elderly group than in the other two groups. The results of this study showed that the major ligand of carbamazepine in the serum was non-glycated albumin, which decreased with age. These observations suggested that in elderly patients, the elevation of free carbamazepine concentrations in the serum caused by reduced non glycated albumin levels, induces increases in the sensitivity of the pharmacological effects of carbamazepine and the risk of drug interactions. PMID- 10528983 TI - Enhancement of blood-brain barrier permeability by sodium caprate. AB - We examined the effect of sodium caprate on the integrity of the tight junctions of brain capillary endothelial cells, using a modified in-situ brain perfusion technique. Model hydrophilic compounds used were [3H]mannitol, [14C]sucrose, [3H]PEG900, [3H]PEG4000, FITC-dextran 4000, FITC-dextran 20000 and FITC-dextran 70000. The brain distribution volume of [14C]sucrose was significantly increased by sodium caprate in a concentration-dependent manner. The effective minimum concentration was 10mM. Furthermore, the effects of sodium caprate on the distribution volumes of hydrophilic compounds, [3H]mannitol, [14C]sucrose, [3H]PEG900, [3H]PEG4000, FITC-dextran 4000, FITC-dextran 20000 and FITC-dextran 70000, showed a molecular weight dependence. A plot of apparent permeation clearance against diffusion coefficient values suggested that 20 mM sodium caprate induced pores so large that the above compounds could pass through the blood-brain barrier with negligible friction within the pore. Our results showed that intracarotid sodium caprate perfusion could enhance the permeation of hydrophilic compounds through the blood-brain barrier. PMID- 10528984 TI - Renal handling of iodobenzoates in rats. AB - Renal elimination pathways of three positional isomers of iodobenzoic acid (2 iodobenzoate, 3-iodobenzoate and 4-iodobenzoate radiolabelled with 125I) were compared using the perfused rat kidney in-situ. All agents were eliminated both in a parent form (involving all renal elimination mechanisms i.e. glomerular filtration, tubular secretion, and tubular reabsorption) and also metabolized to a large extent in the kidney. After 3-iodobenzoate and 4-iodobenzoate administration, the major fractions of radioactivity found in urine were in the form of their metabolites, whereas 2-iodobenzoate was eliminated into urine mostly as the parent compound. Proportions of the individual metabolites in the urine of the perfused rat kidney were similar to those in intact rats for all agents. The results suggest that the kidney is the major organ for both the excretion and metabolism of iodobenzoates in rats. The principal renal metabolic reaction for all compounds under study was conjugation with glycine to produce the corresponding hippuric acid derivatives. PMID- 10528985 TI - Gastrointestinal absorption of a new reversible proton pump inhibitor, YJA-20379 8, and its pharmacokinetics after oral administration in acetic acid-induced gastric ulcer in rats. AB - The absorption of YJA-20379-8 (3-butyryl-4-[5-(R)-(+)-methylbenzylamino]-8-ethoxy 1,7-naphthyrid ine) from various rat gastrointestinal segments was evaluated using in-situ closed-loops. The pharmacokinetics of the drug were also evaluated after oral administration to rats with acetic acid-induced gastric ulcer (AIURs). The concentrations of YJA-20379-8 in the biological samples were analyzed by HPLC. The absorption of YJA-20379-8 from stomach and jejunum was fast, but approximately 50% of the drug was recovered from each segment at 24 h. The total areas under the plasma concentration-time curves from time zero to 24h (AUC(0 24h)) were 161, 392, 233, 365, and 226 microg min mL(-1) for stomach, duodenum, jejunum, ileum, and colon, respectively. After oral administration of the drug, the plasma concentrations and the resultant AUC (0- 12h) were not significantly different between control and AIURs. The detection limits of YJA-20379-8 in human plasma and urine were 50 and 100 ng mL(-1), respectively. The results suggest that modification of the oral dose of YJA-20379-8 may not be required in gastric ulcer patients if the present rat pharmacokinetic data could be extrapolated to man. PMID- 10528986 TI - Gastrointestinal first-pass effect of YJA-20379-8, a new reversible proton pump inhibitor, in rats. AB - Since low bioavailability of YJA-20379-8 (3-butyryl-4-[5-R-(+)-methylbenzylamino] 8ethoxy-1,7-naph thy ridine), a new reversible proton pump inhibitor, has been reported after oral administration of the drug to rats, the first-pass organ of the drug was investigated in rats. YJA-20379-8, 50 mg kg(-1), was infused over 1 min via the jugular vein (n=5) or the portal vein (n=5), or was instilled directly into the stomach (n=5) or the duodenum (n=5). After intravenous or intraportal infusion of the drug, the total body clearance of YJA-20379-8 (18.1 and 19.7 mL min(-1) kg(- 1) based on plasma data) was considerably lower than the reported cardiac output (296 mL min(-1) kg(-1) based on blood data) in rats. This data indicated that the first-pass effect of YJA-20379-8 by the lung and heart was negligible. The areas under the plasma concentration-time curve from time zero to time infinity (AUC) after intravenous or intraportal administration of YJA-20379-8 (2760 and 2540 microg min mL(-1)) were not significantly different, indicating that the hepatic first-pass effect of the drug was also negligible in rats. After intragastric or intraduodenal instillation of YJA-20379-8, the extent of absolute oral bioavailability was 18.2 and 33.8%, respectively. Based on gastrointestinal recovery studies, approximately 86.5 and 91.2% of YJA-20379-8 was absorbed from rat gastrointestinal tract after intragastric or intraduodenal instillation, respectively. The data indicated that gastrointestinal and intestinal first-pass effects of YJA-20379-8 were approximately 68% (86.5-18.2) and 57% (91.2-33.8), respectively. The AUC(0-24h) values of YJA-20379-8 were significantly different between intragastric and intraduodenal instillation, indicating that the gastric first-pass effect of the drug was approximately 10% in rats. Therefore, it could be concluded that the low F value of YJA-20379-8 after oral administration of the drug could be due to a considerable (approx. 60%) intestinal first-pass effect in rats. PMID- 10528987 TI - The influence of plasma binding on absorption/exsorption in the Caco-2 model of human intestinal absorption. AB - The Caco-2 cell monolayer has become an increasingly useful in-vitro model of human intestinal absorption. In this study we have determined the effect of plasma on the basolateral side on the absorption as well as exsorption of several drugs that are highly bound to plasma proteins. The drugs used included propranolol and quercetin, which both use the transcellular route of absorption, and taxol and oestradiol 17 beta-D-glucuronide, which are thought to undergo efflux by P-glycoprotein and the multidrug resistance protein MRP, respectively. All experiments were carried out under sink conditions to mimic normal absorption. It was necessary to use heparin anticoagulation for generation of the plasma, as EDTA was found to make the monolayers very leaky. The apparent permeability (P(app)) values for absorption were 1.54 x 10(-6) cm s(-1) for oestradiol 17 beta-D-glucuronide, 3.33 x 10(-6) cm s(-1) for taxol, 20.8 x 10(-6) cm s (-1) for quercetin, and 35.3 x 10(-6) cm s(-1) for propranolol. For these four compounds, plasma on the basolateral side had no influence on absorption. However, plasma on the basolateral side significantly reduced the efflux of oestradiol 17 beta-D-glucuronide by 66%, taxol by 75%, propranolol by 82%, and quercetin by 94%. Failure to consider the effect of plasma binding can result in an overestimate of basolateral to apical efflux and result in misleading net flux calculations. PMID- 10528988 TI - Antisecretory and relaxatory effects of tachykinin antagonists in the guinea-pig intestinal tract. AB - Existing models used to study the mechanism of action and antagonism of tachykinergic effects on intestinal contraction and secretion suffer from technical problems and have not been fully characterized using specific tachykinin antagonists. Contraction of ileal segments by substance P, colonic circular muscle by beta-alanine-neurokinin A, and longitudinal muscle by senktide were used as models for neurokinin-induced contraction in the guinea-pig. Guinea pig colonic epithelial tissue was stimulated by substance P and senktide to assess NK1- and NK3-mediated secretion. Using these models the potency of therapeutically useful compounds was determined. NK1 and NK2 activation directly contracted smooth muscle, while NK1-mediated secretion was nerve-mediated. NK3 stimulation of contraction and secretion was neurally mediated, involving cholinergic nerves and 5-HT release. NK1-mediated contraction and secretion were antagonized by SR140333 (pD'2 = 9.29 and pKb = 8.53); NK2-mediated contraction was antagonised by SR48968 (pD'2 = 8.35) and NK3-mediated contraction and secretion were antagonized by SB223412 (pKb = 8.97 and 8.79). The mixed antagonist MDL103392 blocked NK1- and NK2-mediated contraction with pKb values of 7.92 and 6.71 respectively and NK1-mediated secretion with a pKb value of 6.57. This data characterizes existing tachykinin antagonists, and should orientate the development of improved compounds as therapies for intestinal disease. PMID- 10528989 TI - Inhibition of Na+ channel or Na+/H+ exchanger attenuates the hydrogen peroxide induced derangements in isolated perfused rat heart. AB - The effect of tetrodotoxin, a specific inhibitor of the Na+ channel, and 5-(N,N dimethyl)-amiloride, a specific inhibitor of the Na+/H+ exchanger, on the mechanical and metabolic derangements induced by hydrogen peroxide (H2O2) was studied in the isolated perfused rat heart. The isolated rat heart was perfused aerobically at a constant flow rate and driven electrically. H2O2 (600 microM) decreased the left ventricular developed pressure and increased the left ventricular end-diastolic pressure (i.e. mechanical dysfunction), decreased the tissue levels of adenosine triphosphate and adenosine diphosphate (i.e. metabolic derangement), and increased the tissue level of malondialdehyde (i.e. lipid peroxidation). These mechanical and metabolic derangements induced by H2O2 were significantly attenuated by tetrodotoxin (3 microM) or 5-(N,N-dimethyl)-amiloride (15 microM). Neither tetrodotoxin nor 5-(N,N-dimethyl)-amiloride modified the tissue malondialdehyde level, which was increased by H2O2. In the normal (H2O2 untreated) heart, neither tetrodotoxin nor 5-(N,N-dimethyl)-amiloride affected the mechanical function and energy metabolism. These results suggested that inhibition of the Na+ channel or Na+/H+ exchanger was effective in attenuating the H2O2-induced mechanical dysfunction and metabolic derangements in the isolated perfused rat heart. PMID- 10528990 TI - Inhibitory effects of the antihistamines epinastine, terfenadine, and ebastine on potassium currents in rat ventricular myocytes. AB - We examined and compared the inhibitory effects of three non-sedating antihistamines, terfenadine, ebastine, and epinastine, on delayed rectifier potassium current (IK) and transient outward potassium current (Ito) of rat isolated ventricular myocytes, using a patch clamp technique. Terfenadine, ebastine and epinastine were found to inhibit IK with IC50 values of 5.96, 15.3 and 145 microM, respectively. Ito was suppressed by epinastine with an IC50 value of 69.5 microM. The order of arrhythmogenicity, assessed by the inhibition of IK, was ranked as terfenadine > ebastine > epinastine, consistent with that of the potencies of each drug for QT prolongation reported in rats. PMID- 10528991 TI - The effect of Oren-gedoku-to on experimental colitis in rats. AB - In Japan and China, Oren-gedoku-to (a complex mixture of ingredients derived from plants) has been used as a herbal medicine in the treatment of inflammatory and ulcerative diseases. In other countries salicylazosulfapyridine has been used to treat inflammatory bowel disease. In this study, we have compared the effect of Oren-gedoku-to with salicylazosulfapyridine on trinitrobenzene-sulphonic acid induced colonic damage in rats, a model representative of ulcerative colitis and Crohn's disease in man. Oren-gedoku-to was administered orally for one or two weeks over a range of doses. Tissue damage scores, body weight, spleen weight, colon wet weight and colon wall thickness were measured, and colonic tissue levels of interleukin-8 (IL-8), leukotriene B4 (LTB4), prostaglandin E2 (PGE2), and myeloperoxidase activity were examined. The results indicated that Oren gedoku-to was effective in the treatment of inflammatory bowel disease in the rat model. Histological observation showed a quicker healing process of the lesions, and a reduction of inflammatory cell infiltration following administration of Oren-gedoku-to. The precise mechanism of action for Oren-gedoku-to is still unclear; however, the reduction of IL-8, LTB4, and PGE2 observed suggests that the mechanism may be different from salicylazosulfapyridine (which has no effect on IL-8). There may be a potential benefit in offering combination therapy for the treatment of inflammatory bowel disease. PMID- 10528993 TI - Effects of Sho-saiko-to extract on liver fibrosis in relation to the changes in hydroxyproline and retinoid levels of the liver in rats. AB - To examine the effects of Sho-saiko-to extract on liver fibrosis, the drug was administered to rats with dimethylnitrosamine-induced liver-injury at various doses. Hydroxyproline and retinoid levels in the liver were measured as indicators of liver function. In liver-injured rats, the hydroxyproline level in the liver (957+/- 154nmol g(-1)) was about 4.16-times that found in normal liver (230+/-11 nmol g(-1)), but administration of Sho-saiko-to extract (0.75%, 1.5% or 3%) reduced the hydroxyproline level significantly (554+/-58, 356+/-51, 374+/ 66nmol g(-1), P<0.01). Single administration of the active constituents of Sho saiko-to extract, glycyrrhizin, baicalin or baicalein, decreased the hydroxyproline level significantly compared with the ordinary food group (P < 0.05), but the decrease was smaller compared with the Sho-saiko-to extract group. The liver retinoid level was higher in the Sho-saiko-to extract group than the ordinary food group and the value increased dose-dependently. A significant negative correlation, r=-0.814 (P<0.001) was detected between the hydroxyproline level and retinoid level in the liver of liver-injured rats. Significant negative correlations, r =-0.728 (P < 0.001) and r= -0.873 (P < 0.001), were also detected between the liver hydroxyproline level and the liver concentrations of the active constituents (glycyrretic acid, baicalin and baicalein) in the liver-injured rats. From these findings, it was considered that the liver concentrations of hydroxyproline and retinoid as well as the active constituents were involved in the improvement of liver fibrosis in the liver-injured rats administered Sho saiko-to extract. Administration of Sho-saiko-to extract inhibited collagen production while an increase in retinoid level inhibited activation of Ito cells leading to inhibition and prevention of liver fibrosis. PMID- 10528992 TI - Antioxidant activity of tannin components from Vaccinium vitis-idaea L. AB - Reactive oxygen molecules have been implicated as important pathological mediators in many clinical disorders and periodontal disease. To provide possible alternative treatment of periodontal disease, six tannins isolated from Vaccinium vitis-idaea L. were evaluated for anti-lipid peroxidation, anti-superoxide formation and free radical scavenging activity. The results showed that cinnamtannin B1 displayed the strongest anti-lipid peroxidation activity, proanthocyanidin A-1 displayed the strongest superoxide scavenging activity, and epicatechin-(4beta--> 6)-epicatechin-(4beta-->8, 2beta-->O--> 7)-catechin had the strongest anti-superoxide formation effect. We conclude that tannins isolated from V. vitis-idaea L. exhibited multiple antioxidant activity, and could be used for the treatment of periodontal disease. PMID- 10528994 TI - Hypophysectomy and/or peroxisome proliferators strongly influence the levels of phase II xenobiotic metabolizing enzymes in rat testis. AB - The objectives of the present work were to determine the influence of hypophysectomy and/or peroxisome proliferators (PP) on certain xenobiotic metabolizing enzyme activities, i.e. glutathione transferases (GST), glutathione peroxidase (GPX), phenol sulphotransferases (pSULT), phenol UDP-glucuronosyl transferases (pUGT), catalase, NADP(H) quinone oxidoreductase (QR) and epoxide hydrolases (EH) in the rat testes. Adult male rats, hypophysectomized and their sham-operated controls, were treated for 10 days with clofibrate (0.5%), perfluorooctanoic acid (0.05%, PFOA), acetylsalicylic acid (1%, ASA) and di(2 ethylhexyl)phthalate (2%, DEHP) in their diet. The results show that, in addition to both body and testis weight, hypophysectomy caused dramatic changes in most of the xenobiotic-metabolizing enzyme activities, which have been measured here. The most pronounced effects were seen in cytosolic QR (2.2-fold increase), pUGT (95% reduction), pSULT (75% reduction), mitochondrial catalase (75% reduction), microsomal EH (70% reduction) and microsomal GST (55% reduction). Treatment with PP, i.e. perfluorooctanoic acid (PFOA), clofibrate, acetyl salicylic acid (ASA) and di(2-ethylhexyl)phthalate (DEHP) showed varied effects on the xenobiotic metabolizing enzyme activities, the highest effects (10-60% reduction) were seen in sham-operated animals. These effects were not so pronounced or were not seen in hypophysectomized rats except for the case of PFOA treatment, which caused increases of enzyme activities. The highest increases were seen with microsomal GST (70%), GPX (75%) and cytosolic EH (75%). It is concluded from these experiments that the regulation of several xenobiotic-metabolizing enzymes in the rat testis is affected by the pituitary and/or pituitary hormones and that different peroxisome proliferators have variable effects on the levels of these xenobiotic-metabolizing enzymes. The general trend of reduction in enzyme activities implies that the testis is less protected under conditions that can perturb hormonal status. PMID- 10528995 TI - Protection against 12-O-tetradecanoylphorbol-13-acetate induced skin-hyperplasia and tumor promotion, in a two-stage carcinogenesis mouse model, by the 2,3 dimethyl-6(2-dimethylaminoethyl)-6H-indolo-[2,3-b]quinoxaline analogue of ellipticine. AB - The effects of topical applications of 2,3-dimethyl-6(2-dimethylaminoethyl)-6H indolo-[2,3-b]quinoxaline (B-220), on 12-O-tetradecanoylphorbol-13-acetate (TPA) or benzoylperoxide (BPO) induced promotion of skin tumors and hyperplasia were studied in female SENCAR mice. Papillomas were induced by initiation with 7,12 dimethylbenz[a]anthracene (DMBA), followed by promotion biweekly with TPA or BPO. Administration of B-220 1 h before TPA promotion resulted in a prolonged latency period of tumor appearance and a significantly reduced (up to 15% of positive controls) papilloma yield at 20 weeks. Moreover, if B-220 treatment was terminated after 20 weeks and TPA treatment continued, papilloma development resumed indicating that initiated tumor cells were still present but were unable to grow with B-220 present. If B-220 pretreatment was not given during the first 10 weeks of TPA promotion, incidence at 20 weeks was not reduced but tumor multiplicity was still decreased. In addition a marked reduction of the TPA induced sustained epidermal hyperplasia was observed in the long term experiment. Neither the inflammatory response nor the increase in the number of apoptotic cells seen in short term experiment after a single TPA treatment were inhibited by B-220. B-220 administration before BPO promotion had no effect on the appearance of BPO induced papillomas or epidermal hyperplasia, suggesting that TPA and BPO promote tumor formation via at least partially different mechanisms. In experiments where B-220 was applied topically 1 h before DMBA initiation, little or no effect was seen. No morphological changes in mouse skin due to long term exposure (two times/week, 39 weeks) to B-220 were found. In conclusion, we present evidence that B-220 is a potent inhibitor of mouse skin tumor promotion by TPA, but has little effect on the initiation step or the survival of initiated cells. PMID- 10528996 TI - Effect of cyanamide on toxicity and glutathione depletion in rat hepatocyte cultures: differences between two dichloropropanol isomers. AB - The effect of aldehyde dehydrogenase inhibition by cyanamide pre-treatment in vitro on dichloropropanol-dependent toxicity and glutathione depletion was investigated in 24 h rat hepatocyte cultures. Cyanamide pre-treatment had no effect on nitrophenol hydroxylase, 7-methoxy-, 7-ethoxy- or 7-benzyloxyresorufin O-dealkylase activities in 24 h cultures from untreated rats, and had no effect on intracellular glutathione content in cultures from untreated rats, or in cultures from isoniazid-treated rats in which cytochrome P4502E1 (CYP2E1) is increased. In cultures from untreated animals the primary alcohol, 2,3 dichloropropanol, was not toxic and did not significantly deplete glutathione. Cyanamide pre-treatment however, potentiated both toxicity and glutathione depletion. Induction of CYP2E1 also potentiated the toxicity of 2,3 dichloropropanol, and in these cultures cyanamide pre-treatment significantly increased both toxicity and glutathione depletion. Cyanamide did not alter the toxicity or glutathione depletion due to the secondary alcohol, 1,3 dichloropropanol, irrespective of CYP2E1 induction. These results indicate that the primary alcohol isomer is metabolised to an aldehyde intermediate which depletes glutathione. Under basal conditions this metabolite appears to be effectively detoxified, but increased CYP2E1 activity and/or decreased aldehyde dehydrogenase activity promotes accumulation of metabolite, and therefore increases glutathione depletion and toxicity. PMID- 10528997 TI - The proximate carcinogen trans-3,4-dihydroxy-3,4-dihydro-dibenz[c,h]acridine is oxidized stereoselectively and regioselectively by cytochrome 1A1, epoxide hydrolase and hepatic microsomes from 3-methylcholanthrene-treated rats. AB - Metabolism of the proximate carcinogen trans-3,4-dihydroxy-3,4 dihydrodibenz[c,h]acridine has been examined with rat liver enzymes. The dihydrodiol is metabolized at a rate of 2.4 nmol/nmol of cytochrome P450 1A1/min with microsomes from 3-methylcholanthrene-treated rats, a rate more than 10-fold higher than that observed with microsomes from control or phenobarbital-treated rats. Major metabolises consisted of a diastereomeric pair of bis-dihydrodiols (68-83%), where the new dihydrodiol group has been introduced at the 8,9 position, tetraols derived from bay region 3,4-diol-1,2-epoxides (15-23%), and a small amount of a phenolic dihydrodiol(s) where the new hydroxy group is at the 8,9-position of the substrate. A highly purified monooxygenase system reconstituted with cytochrome P450 1A1 and epoxide hydrolase (17 nmol of metabolites/nmol of cytochrome P450 1A1/min) gave a metabolite profile very similar to that observed with liver microsomes from 3-methylcholanthrene-treated rats. Study of the stereoselectivity of these microsomes established that the (+) (3S,4S)-dihydrodiol gave mainly the diol epoxide-1 diastereomer, in which the benzylic 4-hydroxyl group and epoxide oxygen are cis. The (-)-(3R,4R)-dihydrodiol gave mainly diol epoxide-2 where these same groups are trans. The major enantiomers of the diastereomeric bis-dihydrodiols are shown to have the same absolute configuration at the 8,9-position. Correlations of circular dichroism spectra suggest this configuration to be (8R,9R). The (8R,9S)-oxide may be their common precursor. PMID- 10528998 TI - Down-regulation of the rat hepatic sterol 27-hydroxylase gene by bile acids in transfected primary hepatocytes: possible role of hepatic nuclear factor 1alpha. AB - In vitro and in vivo studies have shown that the sterol 27-hydroxylase (CYP27) gene is transcriptionally repressed by hydrophobic bile acids. The molecular mechanism(s) of repression of CYP27 by bile acids is unknown. To identify the bile acid responsive element (BARE) and transcription factor(s) that mediate the repression of CYP27 by bile acids, constructs of the CYP27 5'-flanking DNA were linked to either the CAT or luciferase reporter gene and transiently transfected into primary rat hepatocytes. Taurocholate (TCA), taurodeoxycholate (TDCA) and taurochenodeoxycholate (TCDCA) significantly reduced CAT activities of the 840/+23, -329/+23, and -195/+23 mCAT constructs. A -76/+23 construct showed no regulation by bile acids. When a DNA fragment (-110/-86) from this region was cloned in front of an SV 40 promoter it showed down-regulation by TDCA. 'Super' electrophoretic mobility shift assays (EMSA) indicated that both HNF1alpha and C/EBP bind to the -110 to -86 bp DNA fragment. Recombinant rat HNF1alpha and C/EBPalpha competitively bound to this DNA fragment. 'Super'-EMSA showed that TDCA addition to hepatocytes in culture decreased HNF1alpha, but not C/EBP, binding to the -110/-86 bp DNA fragment. A four base pair substitution mutation ( 103 to -99) in this sequence eliminated TCA and TDCA regulation of the (-840/+23) construct. The substitution mutation also eliminated (>95%) HNF1alpha, but not C/EBP, binding to this DNA fragment. We conclude that bile acids repress CYP27 transcription through a putative BARE located between -110 and -86 bp of the CYP27 promoter. The data suggest that bile acids repress CYP27 transcriptional activity by decreasing HNF1alpha binding to the CYP27 promoter. PMID- 10528999 TI - Conditional modulation of glucocorticoid receptor activities by CREB-binding protein (CBP) and p300. AB - Coactivators of nuclear receptors are integral components of the signal transduction pathways of steroid hormones. Here, we show that one of the major coactivators of the glucocorticoid receptor (GR), CREB-binding protein (CBP), can also function conditionally as a negative regulator of its activities. Indeed, CBP suppressed the responsiveness of the mouse mammary tumor virus (MMTV) promoter to dexamethasone in a dose-dependent fashion in HeLa and A204 cells. Similarly, this protein suppressed the responsiveness of several glucocorticoid responsive element (GRE)-containing synthetic promoters. Using deletion mutants of CBP, we localized the repressor effect of this protein to its N-terminal domain and showed that it was independent of the histone acetyltransferase and coactivator-binding domains but dependent upon its GR-binding domain. We also demonstrated functional differentiation between CBP and other coactivators, including SRC-1 and the CBP-related protein p300, both of which influenced GR signaling in a positive fashion. In fact, p300 completely antagonized the suppressive effects of CBP in a dose-dependent fashion, probably by competing with this protein at the level of the transcription complex. These findings suggest that CBP and p300 may function additively or antagonistically to each other depending on their relative concentrations and type of target tissue, to influence the sensitivity of tissues to glucocorticoids. PMID- 10529001 TI - Control of sulfatase and sulfotransferase activities by medrogestone in the hormone-dependent MCF-7 and T-47D human breast cancer cell lines. AB - In the present study, we explored the effect of the progestin medrogestone on the sulfatase and sulfotransferase activities in the hormone-dependent MCF-7 and T 47D human breast cancer cell lines. After 24 h incubation at 37 degrees C of physiological concentrations of estrone sulfate ([3H]-E1S: 5x10(-9) mol/l), it was observed that this estrogen was converted in a great proportion to E2 in both cell lines. Medrogestone significantly inhibits this transformation, at all the concentrations tested (5x10(-8) to 5x10(-5) mol/l), in both cell lines. The IC50 values were 1.93 micromol/l and 0.21 micromol/l in MCF-7 and T-47D cells, respectively. In another series of studies, after 24 h incubation at 37 degrees C of physiological concentrations of estrone ([3H]-E1: 5x10(-9) mol/l), the sulfotransferase activity was detectable in both cell lines. Estrogen sulfates (ES) are found exclusively in the culture medium, which suggests that as soon as they are formed they are excreted into the medium. Medrogestone has a biphasic effect on sulfotransferase activity in both cell lines. At low doses: 5x10(-8) and 5x10(-7) mol/l, this compound stimulates the enzyme by +73.5 and 52.7%, respectively, in MCF-7, and by 84.5 and 62.6% in T-47D cells. At high concentrations: 5x10(-6) and 5x10(-5) mol/l, medrogestone has no effect on MCF-7 cells, but inhibits the sulfotransferase activity in T-47D cells by -31.4% at 5x10(-5) mol/l. In conclusion, the inhibitory effect provoked by medrogestone on the enzyme involved in the biosynthesis of E2 (sulfatase pathway) in estrogen dependent breast cancer, as well as the stimulatory effect on the formation of the inactive ES, support a probable anti-proliferative effect of this progestin in breast tissue. Clinical applications of these findings can open new therapeutic possibilities for this disease. PMID- 10529000 TI - Ligand structure influences autologous downregulation of estrogen receptor-alpha messenger RNA. AB - A series of A- and D-ring substituted estrogen analogues have been examined for their effect on estrogen receptor-alpha (ERalpha) mRNA downregulation. Recently it has been proposed that ERalpha autologous downregulation occurs via transcriptional repression exerted by the binding of the ERalpha-ligand complex to the 5' region of the coding region of the ERalpha gene. Placement of the phenolic hydroxyl group on the various carbons of the aromatic A-ring of estratrien-17betaol (carbons 1-3) produced ligands which diminished the steady state level of ERalpha mRNA in relation to their affinity for receptor. 4 Hydoxyestratrien-17betaol, on the other hand, was inactive in the downregulation of ERalpha mRNA. Although this A-ring isomer brought about apparent processing of the nuclear receptor, the ERalpha reappeared in the cytosol within 24 h. Unlike the stimulation of genes regulated via estrogen response elements, maximum autologous negative regulation of the ERalpha gene required the presence of an hydroxyl group on carbon 17 of the D-ring. These results suggest that the conformational alterations elicited in the ERalpha molecule by various ligands create surfaces capable of interacting with other transcription factors in a manner which is different when the receptor functions via a response element mechanism relative to interactions during autologous negative regulation of the ERalpha gene. PMID- 10529002 TI - Changes of hypothalamic and plasma vasopressin in rats with deoxycorticosterone acetate induced salt appetite. AB - Mineralocorticoids play a predominant role in development of salt appetite and hypertension. Since vasoactive peptides could mediate the central effects of mineralocorticoids, we evaluated changes of immunoreactive (IR) arginine vasopressin (AVP) in the paraventricular (PVN) and supraoptic (SON) hypothalamic nucleus during DOCA-induced salt appetite. In one model, rats having free access to water and 3% NaCl during 9 (prehypertensive stage) or 21 days (hypertensive stage) received DOCA (s.c., 10 mg/rat/in alternate days). A decrease in the IR cell area, number of IR cells and staining intensity was obtained in magnocellular PVN of rats treated during 9 days. After 21 days IR cell area and number of cells in the PVN also decreased, but staining intensity of remaining cells was normal. The same parameters were unchanged in the SON. In another model, animals treated with DOCA during 9 days had only access to 3% NaCl or water. The IR cell area in PVN and SON significantly increased in mineralocorticoid-treated and control animals, both drinking 3% NaCl. Staining intensity (PVN and SON) and number of IR cells (PVN) also augmented in DOCA treated animals drinking salt respect of a group drinking water. Plasma AVP in rats treated with DOCA and offered salt and water, exhibited a 2-2.5 fold increase at the time of salt appetite induction. Plasma AVP was substantially higher in rats drinking salt only, while the highest levels were present in salt drinking DOCA-treated rats. Thus, peptide depletion in the PVN may be due to increased release, because reduced levels of hypothalamic and posterior pituitary AVP were measured in this model. In rats drinking salt only the substantial increase of IR AVP in the PVN and SON, may be due to dehydration and hyperosmosis. Because DOCA-salt treated rats showed higher AVP levels in the PVN compared to untreated rats drinking salt only, it is possible that DOCA sensitized PVN cells to increase AVP production. The results suggest the vasopressinergic system could mediate some central functions of mineralocorticoids. PMID- 10529003 TI - MR 20492 and MR 20494: two indolizinone derivatives that strongly inhibit human aromatase. AB - In this study, we describe the synthesis of a new family of indolizinone derivatives designed to fit an extrahydrophobic pocket within the active site of aromatase and to strongly inhibit human aromatase. This could help improve the specificity of the inhibitors. Equine aromatase, very well characterized biochemically, is used as a comparative model. Indeed, in a previous comparison between both human and equine aromatases, we described the importance of the interaction between the inhibitor and this pocket for the indane derivative MR 20814. MR 20492 and MR 20494 are more potent inhibitors of human aromatase (Ki/Km: 1.0+/-0.3 and 0.5+/-0.3, respectively). The Ki/Km for MR 20494 is slightly higher than that obtained for fadrozole (0.1+/-0.0) and Ki/Km for both indolizinone derivatives are lower than those obtained for 4 hydroxyandrostenedione (1.9+/-0.8) and MR 20814 (8.1+/-.7). These new compounds are not enzyme inactivators. Moreover, as indicated by the higher Ki/Km values obtained with equine enzyme (9.0+/-0.6 and 6.1+/-1.6 for MR 20492 and MR 20494, respectively), both human and equine aromatase active sites appear to be structurally different. Difference absorption spectra study (350-500 nm) revealed that MR20492 and MR20494 were characterized by a combination of type-I and -II spectra with both enzymes. This result could be due to the isomerization of the molecule in polar solvent (Z and E forms). The evaluation of these new molecules, as well as 4-hydroxyandrostenedione and fadrozole, on aromatase activity in transfected 293 cell cultures evidenced a strong inhibition (IC50: 0.20+/-0.03 microM, 0.20+/-0.02 microM and 0.50+/-0.40 microM for MR 20494, fadrozole and 4 OHA, respectively) except for MR 20492 (3.9+/-0.9 microM) and MR 20814 (10.5+/ 0.6 microM). These results proved that these molecules formed part of a promising family of potent inhibitors and that they penetrate 293 cells, without evidencing any cytotoxicity in Hela cells with MTT assay. This is thus encouraging for the development of new drugs for the treatment of estrogen-dependent cancers, these molecules also constitute new tools for understanding the aromatase active site. PMID- 10529004 TI - Estradiol induction of cAMP in breast cancer cells is mediated by foetal calf serum (FCS) and sex hormone-binding globulin (SHBG). AB - Plasma sex hormone-binding globulin (SHBG or SBP), the specific carrier for estradiol and androgens, after binding to its membrane receptor (SHBG-R), causes a significant increase of cAMP in the presence of estradiol, in both breast (MCF 7) and prostate (LNCaP) cancer cells maintained in serum-free medium. On the other hand, it has been proposed that estrogens, in addition to the well-known nuclear receptor pathway, exert their biological effect inducing cAMP, as a consequence of a direct membrane action, in breast cancer and uterine cells. The aim of the present study was to clarify this controversial issue by verifying if the cAMP increase in MCF-7 cells was a direct effect of estradiol, or if it was mediated by FCS proteins, such as bovine sex hormone-binding globulin; and to reevaluate the effect of human SHBG on cAMP induction in the presence of FCS. MCF 7 cells were maintained in DCC-FCS (treated with DCC to remove steroids), in SHBG FREE/DCC-FCS (treated with DCC and with a specific affinity chromatography to remove bovine sex hormone-binding globulin), or in serum-free medium (SFM). It was observed that estradiol determined a significant time-dependent increase of cAMP only in MCF-7 cells maintained in 10% DCC-FCS. When cells were maintained in 10% SHBG-FREE/DCC-FCS, estradiol had no detectable effect. However, its ability to increase cAMP was observed again after the addition of human SHBG, in doses ranging from 5 to 50 nM. Moreover, in the presence of 10% SHBG-FREE/DCC-FCS, SHBG, even in the absence of estradiol, caused a significant increase of cAMP. In conclusion, the data reported in the present study suggest that the ability of estradiol to induce cAMP in MCF-7 cells is not due to a direct membrane effect of the hormone, but rather it is mediated by FCS. SHBG is one of the serum factors mediating estradiol action. Lastly, it was proven that SHBG triggers the cAMP pathway in MCF-7 cells in a physiologic culture condition and at physiologic concentrations. PMID- 10529005 TI - Prenatal melatonin exposure influences the maturation of gonadotropin and prolactin estradiol-benzoate feedback system. AB - Gonadotropin and prolactin response to estrogen feedback in female rat offspring of control and melatonin treated (150 microg/100 g BW) mother rats during pregnancy (MEL-offspring) were studied at these periods: infantile, prepubertal and pubertal. In controls negative or absent LH feedback developed after estradiol benzoate (EB) injection up to 30 days of age indicating that the onset of puberty had not occurred. The positive feedback was established from day 33 on. However, in MEL-offspring the first activation of gonadotropin secretion during afternoon, 31 h after EB, was observed at 25 days of age, representing the first neuroendocrine sign of the onset of puberty. This positive response disappeared on day 30 in MEL-offspring. At 33 days of age, the LH positive response to EB was found in both groups, indicating a more advanced sexual development. In controls, this response increased at 35 days of age while in MEL offspring it was highly depressed. FSH secretion in response to EB showed a negative feedback effect from infantile to the end of prepubertal period in both groups. The positive feedback was observed earlier in MEL-offspring (at 33 days of age) than in controls (at 35 days of age), but at this age it was absent in MEL-offspring. A positive prolactin response to EB at all ages in controls was observed. The typical pulsatility with higher values in the afternoon appeared by the first time at 30 days of age. However, in MEL-offspring no pulsatile response was observed throughout any age. These data suggest that prenatal melatonin administration altered gonadotropin and prolactin response to EB inducing precocious sensitivity during prepubertal period but depressed response during the pubertal period. PMID- 10529006 TI - Rainbow trout, Oncorhynchus mykiss, as a model for aromatase inhibition. AB - The feasibility of utilizing rainbow trout, Oncorhynchus mykiss, as an alternative model for studying the inhibition of aromatase (CYP 19) was investigated. The suppression of estrogen-dependent tumors by aromatase inhibitors has been important in the treatment of breast cancer. Estrogens, estrogen precursors and xenoestrogens have been found to promote liver cancer in the trout model. A steroid, 4-hydroxy-4-androstene-3,17-dione (4-OHA), and non steroids, aminoglutethimide (AG) and Letrozole (CGS 20267), all of which are known aromatase inhibitors in rats and humans, were examined in vitro for activity in trout ovarian microsomes. Aromatase activity was quantified as the release of 3H2O from the conversion of [3H]-4-androstene-3,17-dione to 17beta estradiol and estrone. Trout ovarian microsomes exhibited activity between 39-60 fmol mg(-1) min(-1) with a calculated Vmax of 71.1 fmol mg(-1) min(-1) when incubated at 25 degrees C with 200 nM 4-androstene-3,17-dione (K(M) = 435 nM). Significant inhibition by 4-OHA up to 80% was seen at 1.5 microM. At 2000 microM, AG decreased aromatase activity by up to 82%. Letrozole reduced aromatase activity a maximum of 90% in a dose-dependent manner, but the Ki (2.3 microM) was 1000-fold higher than reported in human trials. Indole-3-carbinol and some of its derivatives, two DDE isomers and four flavones (except alpha-naphthoflavone) at 1000 microM did not significantly inhibit aromatase in vitro. Letrozole and clotrimazole, fed to juvenile rainbow trout at doses up to 1000 ppm for 2 weeks, were not effective in suppressing dehydroepiandrosterone (DHEA) induced increases in vitellogenin and 17beta-estradiol levels. These results document that trout aromatase is sensitive to inhibition in vitro by known inhibitors of the mammalian enzyme. The mechanism(s) for lack of inhibition in vivo is currently unknown and must be further investigated in order to develop a trout model for studying the role of aromatase in carcinogenesis. PMID- 10529007 TI - The length of the polyoxyethylene chain in the Triton X detergents modulates the apparent activation of neurosteroid sulfatase in bovine brain. AB - The effect of the Triton X series on the solubilization and enzyme activity of neurosteroid sulfatase (NSS) in the bovine midbrain was investigated. Triton X 100 and X165 stimulated NSS activity in the bovine midbrain, while Triton X-305 did not. This apparent activation was attributed to the action of the detergents, and not to the latency of the enzyme or the removal of some inhibitory substance from the microsomes. The maximum stimulation was obtained when the length of the polyoxyethylene chain of the detergent was 16. PMID- 10529008 TI - Differential glucuronidation of bile acids, androgens and estrogens by human UGT1A3 and 2B7. AB - In this work, UDP-glucuronosyltransferases (UGTs), UGT1A3, 2B7(H268) and 2B7(Y268), stably expressed in human embryonic kidney cells (HK293) were used to assess glucuronidation activities with a variety of steroid hormone and bile acid substrates. The rate of synthesis of carboxyl- and hydroxyl-linked glucuronides was determined under optimal reaction conditions. Expressed UGT1A3 catalyzed bile acid glucuronidation at high rates exclusively at the carboxyl moiety for all compounds tested. In contrast, UGT1A4 catalyzed bile acid glucuronidation at very low rates exclusively at the 3alpha-hydroxyl function. Both UGT2B7 allelic variants glucuronidated the bile acid substrates at both carboxyl and hydroxyl moieties, however, the 3alpha-hydroxyl position was preferentially conjugated compared to the carboxyl function. Similarly, androsterone, a 3alpha-hydroxylated androgenic steroid, was glucuronidated at very high rates by expressed UGT2B7. Of the estrogenic compounds tested, UGT2B7 catalyzed the glucuronidation of estriol at rates comparable to those determined for androsterone. Other structural discrimination was found with UGT2B7 which had activity toward estriol and estradiol exclusively at the 17beta-OH position, yielding the cholestatic steroid D-ring glucuronides. PMID- 10529009 TI - Progesterone metabolism in human saliva in vitro. AB - Human salivary glands are known to be able to metabolize progesterone as well as other steroid hormones. The rate of progesterone metabolism in the salivary glands is so low that it is not thought to affect salivary progesterone concentrations. On the other hand it is usually recommended that saliva should be frozen quickly after the collection to prevent any kind of metabolism in saliva. When saliva is collected at home e.g. delayed freezing or partial thawing during to transport to laboratory may create circumstances where progesterone metabolism may occur. However, it is not known to which extent progesterone metabolism continues in saliva and whether this continued metabolism of progesterone affects salivary hormone levels. Paraffin-stimulated salivary samples were collected from female (N = 6) and male (N = 6) dental students and perimenopausal women (N = 8). The salivary samples were incubated with 14C-progesterone for 2 h at 37 degrees C in a shaking water bath. Metabolites were analyzed using thin-layer chromatography and autoradiography and quantified by liquid scintillation counting. Human saliva was found to be able to metabolize progesterone, but its metabolic activity was very low, 9.3 and 6.8 pmol/ml/h in young adults and perimenopausal women, respectively. Metabolic activity was higher in whole saliva than in the corresponding activities of the supernatant or sonicated fraction of the same saliva. The supernatant fraction, which was thought to be mainly representative of glandular saliva, was metabolically least active. The polar metabolites of progesterone predominated in all incubations. The metabolic activity of saliva is probably mainly due to its cellular content and the contribution of this activity to salivary progesterone concentrations is not significant. PMID- 10529010 TI - Relation of transmembrane potential to cell survival following hyperthermia in HeLa cells. AB - Hyperthermia induces cell death and the usual endpoint to study this is the ability of the cells to form colonies. Hyperthermia is also known to alter membrane characteristics, especially transmembrane potential and this has been correlated with duration and degree of heating. The aim of the present study was to see the correlation between changes in membrane potential and clonogenic ability of HeLa cells after heat treatment of 41-44 degrees C. Membrane potential was measured by the fluorescence polarization of the plasma membrane probe 3,3' dipentyloxacarbocyanine by flow cytometry. Cell survival was assessed by colony formation assay. The fluorescence intensity increased and cell survival decreased with an increase in temperature. The fall in survival following heat treatment closely paralleled the increase in fluorescent intensity, especially heat treatments of 60 min or more. After 2 h of heating at 44 degrees C, the surviving fraction decreased to 1% and the fluorescence intensity increased to 154.84% of the unheated controls. This study suggests that measurement of membrane potential by flow cytometry may potentially be an alternative to colony forming assay for assessing cell survival. Since the results of membrane potential measurements are available immediately, this has implications for its potential use as a predictive assay of thermosensitivity. PMID- 10529011 TI - Microvascular endothelium dysfunction in rats bearing 1,2-dimethylhydrazine induced colon tumors. AB - Functional parameters (adhesivity, permeability, thrombogenicity and thromboresistance) of the mesenteric microvasculature endothelium in rats bearing 1,2-dimethylhydrazine (DMH) induced colon tumors were studied using a complex method, based upon computer-assisted microscopic video image processing. It was shown that paraneoplastic endothelial dysfunction in rat mesenteric microvessels is characterized by an increase in leukocyte adhesion to the endothelium, microvessel permeability and thrombogenic properties. It was demonstrated that the observed changes are the consequences of tumor growth and are related to tumor size and the duration of the process. PMID- 10529012 TI - Dim light during darkness stimulates tumor progression by enhancing tumor fatty acid uptake and metabolism. AB - Tumor linoleic acid uptake and metabolism, and growth are suppressed by melatonin, the synthesis of which is inhibited by light. Linoleic acid, via its mitogenic metabolite 13-hydroxyoctadecadienoic acid (13-HODE) is an important growth stimulant of rat hepatoma 7288CTC. Here we compared the effects of an alternating light:dark cycle (12L:12D), dim light (0.25 lux) present during the dark phase of a diurnal light cycle, and constant light on growth and fatty acid metabolism in hepatoma 7288CTC. Our results show that dim light suppressed melatonin release by the pineal gland, increased tumor linoleic acid uptake and 13-HODE production, and promoted tumor growth as effectively as did constant light. PMID- 10529013 TI - Anticlastogenic effects of centchroman and its enantiomers in Swiss albino mice. I. Acute study and their comparison with tamoxifen. AB - Centchroman (CC), a non steroidal oral contraceptive and a candidate drug for breast cancer, has been reported to exhibit partial to complete remission of lesions in 40.5% of breast cancer patients. Recently, we have reported the antimutagenic effects of CC in multiple mutational assays. The potent antioestrogenic activity, negligible side effects, anti-breast cancer activity and antimutagenic effects of CC prompted us to evaluate the anticlastogenic effects of CC and two of its enantiomers. i.e. D-centchroman (DC) and L centchroman (LC) in the acute in vivo studies in female Swiss albino mice as measured by chromosome aberrations (CA) and sister chromatid exchange (SCE) assays against two known positive mutagen compounds, i.e. dimethylbenz[a]anthracene (DMBA) and cyclophosphamide (CP). The results of anti mutagenicity assays of CC and its enantiomers have been compared to the known breast cancer drug tamoxifen (TM). CC and LC reduced both DMBA and CP induced CA when compared with the group treated with only DMBA and CP. DC did not reduce the DMBA-induced CA when compared with the DMBA-treated group alone. It reduces only the CP induced CA. TM also reduces both DMBA and CP induced CA when compared with group received only DMBA or CP. SCE were carried out only for LC. A weak but significant decrease in SCE was observed in both LC plus DMBA- and LC plus CP treated groups when compared with respective positive controls alone. Thus the overall results indicate that both CC and LC are more effective in reducing the genotoxic effects of DMBA and CP than DC. PMID- 10529014 TI - High incidence of synchronous cancer of the oral cavity and the upper gastrointestinal tract. AB - A high incidence of synchronous esophageal or gastric carcinoma in preoperative patients with carcinoma of the oral cavity was reported. Esophageal carcinoma was found in seven out of 56 patients (12.5%) and gastric cancer in five patients (8.9%) by videoendoscopy aided with lugol staining in the esophagus and indigocarmine solution in the stomach, although all patients were completely asymptomatic for these lesions. All patients were male, regular drinkers and heavy smokers. The depth of invasion of such tumors was limited to either mucosa or submucosa. Those esophageal and gastric lesions beside the primary oral cancers were positive for p53 protein by immunohistochemistry. Careful preoperative evaluation of not only the esophagus but also the stomach should be a routine procedure in patients with carcinoma of the oral cavity. PMID- 10529015 TI - Human urinary bladder carcinoma cell lines respond to treatment with alkylphosphocholines. AB - Alkylphosphocholines (APC) constitute a new group of antineoplastic agents without haematological toxicity. Their first clinically available derivative hexadecylphosphocholine (miltefosine) is locally used to control skin metastases of breast cancer. Since intravesical chemotherapy represents a form of topical treatment we investigated whether a new APC with a long alkyl chain would be active against 5637 and EJ bladder cancer cell lines. Their antineoplastic activity was inversely related to the alkyl chain length of the respective APC. Erucylphosphocholine and its congener with modified phosphocholine head erucylphospho-N,N,N-trimethylpropanolamine were the most effective derivatives. APC with alkyl chains over 16 carbons in length induced programmed cell death in both cell lines, as determined by oligonucleosomal DNA fragmentation and morphology. The distinct antineoplastic effects lead us to predict that urinary bladder instillation of APC will be of therapeutic benefit for patients with urinary bladder neoplasia. PMID- 10529016 TI - Expression of nitric oxide synthase in gastric cancer. AB - The expression of endothelial nitric oxide synthase (eNOS) and inducible nitric oxide synthase (iNOS) in samples of normal gastric mucosa and gastric cancer were examined by reverse transcriptase-polymerase chain reaction (RT-PCR) and semi quantitative Western blot. In normal gastric mucosa, eNOS protein was found in all samples examined (mean, 70.2 +/- 60.1), relative to a standard protein. In gastric cancer specimens, eNOS protein was also detected in all samples, but the quantity (86.5 +/- 76.6) was not different from that found in samples of normal mucosa. The quantity of eNOS in gastric cancer tissues was negatively correlated with serosal invasion. iNbS mRNA, detected in nine of 18 cases, was slightly related to massive lymph node metastasis (n1-3 vs. n4). Neither tumor necrosis factor alpha (TNF-alpha) mRNA nor interleukin-6 (IL-6) mRNA was related to the expression of iNOS mRNA. These results suggest that iNOS not eNOS plays a role in gastric cancer tumor extension, but iNOS mRNA appears not to be induced by either TNF-alpha or IL-6. PMID- 10529017 TI - In vivo administration of genistein has no effect on small intestinal epithelial proliferation and apoptosis, but a modest effect on clonogen survival. AB - The soya metabolite genistein possesses anti-proliferative, pro-apoptotic activities in intestinal epithelial cells in vitro and may reduce epithelial cancer incidence. This could involve cell cycle arrest/apoptosis in the proliferative or clonogenic cells. We investigated the effects of genistein on the small intestinal epithelium in vivo. No effect on the number or distribution of proliferative cells in the crypts was detected. Similarly, no change in spontaneous apoptotic cell incidence or the characteristic stem cell apoptotic response following low dose irradiation was observed. Genistein afforded a modest decrease in clonogen radiosensitivity. Hence, using a range of dosing protocols, sub-cutaneous administration of genistein for periods of up to 1 week did not alter intestinal epithelial homeostasis. PMID- 10529018 TI - Comparison of 1-(2-deoxy-2-fluoro-4-thio-beta-D-arabinofuranosyl)cytosine with gemcitabine in its antitumor activity. AB - The antitumor activity of a novel nucleoside, 1-(2-deoxy-2-fluoro-4-thio-beta-D arabinofuranosyl)cytosine (4'-thio-FAC) was compared with that of 2'-deoxy-2',2' difluorocytidine (gemcitabine). 4'-Thio-FAC showed potent antitumor effects against various solid cancer cell lines in vitro. Also, gemcitabine showed remarkable in vitro antitumor effects, even more potent than 4'-thio-FAC. However, 4'-thio-FAC inhibited tumor growth more strongly than gemcitabine did at the same dose against human cancer cells implanted s.c. in nude mice. In addition, 4'-thio-FAC suppressed the tumor growth by oral administration. The toxicity of 4'-thio-FAC was weaker than that of gemcitabine in nude mice in both consecutive and intermittent administration. Accordingly, clinical usefulness of 4'-thio-FAC is expected. PMID- 10529019 TI - Topographic quantitative EEG sequelae of chronic marihuana use: a replication using medically and psychiatrically screened normal subjects. AB - In two previous studies it was reported that chronic marihuana (THC) use was associated with unique quantitative EEG features which were present in the non intoxicated state. THC users, as contrasted with controls, had significant elevations of Absolute Power, Relative Power, and Coherence of alpha activity over the bilateral frontal cortex. Furthermore, a quantitative EEG discriminant function analyses permitted a 95% correct user versus non-user classification. However, because all of the THC users and 58% of the non-user controls were psychiatric inpatients, diagnostic and medication effects, if any, were uncontrolled. In the present study the same quantitative EEG methods were used to study daily THC users and non-user controls who underwent a rigorous screening process to insure that they were medically and psychiatrically healthy. The results of previous studies were replicated and an additional EEG correlate of chronic THC exposure (reduced alpha frequency) was identified. PMID- 10529020 TI - Naltrexone shortened opioid detoxification with buprenorphine. AB - This double-blind, randomized, placebo-controlled clinical trial evaluated the impact on withdrawal symptoms of (i) combining naltrexone with a 4-day buprenorphine taper for short opioid detoxification (NB Group), compared to (ii) using a 4-day buprenorphine taper alone, followed by naltrexone on day 8 (PB Group). Sublingual buprenorphine was administered on days 1-4 (26 mg total). For the NB Group (n = 32) escalating doses of oral naltrexone were given on days 2-8 (placebo day 1). For the PB Group (n = 28) placebo was given on days 1-7 and naltrexone on day 8. Main outcome measures were Observed Opioid Withdrawal scores (OOW, 0-30) and use of medications to treat opioid withdrawal. Of 32 patients in the NB group, 59% experienced clinically relevant withdrawal (defined as OOW > or = 5) on day 2, but, after day 5, none experienced withdrawal. In the PB group, the number of patients experiencing withdrawal increased over time. The first naltrexone dose induced comparable withdrawal in both groups: peak OOW scores were (mean +/- SD) 5.2 +/- 3.3 on day 2 for the NB group, and 4.0 +/- 3.9 on day 8 for the PB group (NS), though, on day 2, 7 patients dropped out in the NB group and none in the PB group, while only one patient dropped out in the PB group on day 8. Throughout the 8-day study, patients in both groups received similar amount of adjunct medication: 0.64 +/- 0.07 mg (NB group) of clonidine vs 0.73 +/ 0.15 mg (PB group; NS). Only 25% of patients required use of sedatives (up to 20 mg diazepam). Starting naltrexone on day 2 appeared to abolish withdrawal symptoms after day 5 and, thus, to shorten the duration of withdrawal symptoms. Peak withdrawal symptoms after naltrexone were of moderate intensity, suggesting that naltrexone combined with buprenorphine is an acceptable and safe treatment for shortened opioid detoxification and induction of naltrexone maintenance. PMID- 10529021 TI - Opioid reinforcement in heroin-dependent volunteers during outpatient buprenorphine maintenance. AB - This study examined the reinforcing effects of hydromorphone (HYD) (0, 4, 8, and 16 mg/70 kg i.m.) in heroin-dependent outpatient volunteers maintained on buprenorphine (BUP) at doses of 2, 4, and 8 mg, each for 2 weeks. Following a week of maintenance at each dose, volunteers received injections of one of the four HYD doses under double-blind conditions. Eight volunteers (abstainers) were heroin-free during HYD test weeks, whereas six volunteers remained heroin positive (nonabstainers). Among abstainers, HYD had minimal reinforcing value, whereas in nonabstainers there were marked dose-related increases in HYD reinforcing value, which were not attenuated by increasing doses of BUP. A similar pattern was found for HYD subjective agonist effects. Heroin craving among nonabstainers was significantly higher compared with abstainers, and was reduced in a dose-related manner by HYD. Although BUP and HYD produced dose related miosis, abstinence status had no differential effect. In summary, BUP effects on opioid reinforcement were consistent from outpatient setting (heroin abstinence) to laboratory setting (decreased HYD reinforcement), supporting the validity of this laboratory model. PMID- 10529022 TI - Cigarettes, alcohol, marijuana, other risk behaviors, and American youth. AB - Increases in adolescent marijuana and other drug use have created widespread concern. One theory argues that increased use of cigarettes and alcohol among younger adolescents leads to greater use of marijuana which, in turn, leads to subsequent use of other drugs (e.g. cocaine, heroin, hallucinogens). Detractors of this theory claim that use of these substances is a symptom of a larger set of destructive behaviors (e.g. violence, suicide, promiscuous sex), and marijuana has no independent effect on the use of other more serious drugs. The authors examined whether, for high school seniors, early use of cigarettes, alcohol and marijuana has an independent effect on more serious drug use even when other behaviors are considered. Using the 1995 Youth Risk Behavior Survey (n = 2871) and logistic analysis, after accounting for selected other behaviors, seniors using cigarettes before age 13 were 3.3 (95% C.I. 2.3,4.6) times likelier to have used marijuana than ones who never smoked; for alcohol, the odds ratio was 4.5 (2.6,7.7). Seniors using marijuana before the age of 14 were 7.4 times (4.0,13.6) likelier to have used other drugs. Though no causal effect is demonstrated, cigarette and alcohol use was associated with the likelihood of marijuana use; marijuana use was associated with the likelihood of other drug use, even after selected other risk and protective behaviors were considered. PMID- 10529023 TI - Smoking cessation and mortality trends among two United States populations. AB - The long-term impact of smoking cessation on mortality is assessed among two U.S. populations: a large cohort of U.S. veterans aged 55-64 at entry and followed from 1954 through 1979 and the NHANES I Epidemiologic Followup Study (NHEFS) cohort of a national sample of U.S. adults aged 55-74 at entry and followed from 1971 through 1992. Direct and indirect survey data indicate that 50-70% of those who were current cigarette smokers at entry had quit smoking during the 19- to 26 year follow-up periods. The impact of smoking cessation on mortality among the cigarette smokers as a whole has been assessed by determining the time trend of the relative risk (RR) of death and 95% confidence interval (CI) for the cigarette smokers compared with never-smokers over the entire follow-up period in both cohorts. The total death rates for the 1954/57 U.S. veteran smokers as a whole (63,159 males) have converged only slightly toward those of never-smokers, from RR = 1.65 (1.58-1.72) during 1954-1959 to RR = 1.61 (1.58-1.63) during 1954 1979. The lung cancer death rates for 1954/57 smokers as a whole have not converged toward those of never-smokers, with RR = 10.89 (7.70-15.41) during 1954 1959 and RR = 11.10 (9.78-12.61) during 1954-1979. The total death rates for the 1971-1975 NHEFS smokers as a whole (694 males and 1116 females) have not converged toward those of never-smokers. For males, RR = 1.92 (1.46-2.52) during 1971-1982 and RR = 1.96 (1.63-2.36) during 1971-1992; for females, RR = 1.79 (1.31-2.46) during 1971-1982 and RR = 1.79 (1.47-2.17) during 1971-1992. The lung cancer death rates have diverged, based on small numbers of deaths. For males, RR = 15.76 (2.06-120.61) during 1971-1982 and RR = 22.20 (5.31-92.92) during 1971 1992; for females, RR = 2.92 (0.57-15.06) during 1971-1982 and RR = 4.74 (1.94 11.59) during 1971-1992. These trends are contrary to the substantial convergence predicted by the death rate trends among U.S. veterans who were former smokers at the beginning of follow-up. While these results confirm that those former smokers who survive for at least 5 years experience death rates that converge toward those of never-smokers, they also indicate that a cohort of cigarette smokers that undergoes substantial cessation experiences a death rate that does not converge toward the death rate of never-smokers. The fact that there has been no convergence for lung cancer is quite surprising, as this is the disease most strongly linked to smoking and smoking cessation and less likely to be influenced by other lifestyle factors. Further investigation is needed for a complete understanding of the impact of smoking cessation. PMID- 10529024 TI - Rebuttal to the paper by Enstrom. PMID- 10529025 TI - A novel form of ascertainment bias in case-control studies of cancer screening. AB - In case-control studies of cancer screening, some have generally admonished investigators against case definitions based on diagnosis dates because of lead time bias. However, perhaps partly due to vagueness, the admonitions have been frequently ignored. A recurrence-time model simulates case ascertainment when diagnosis must occur within a specific calendar period. The model depends on screening test sensitivity and rate, age-specific preclinical incidence rates, and preclinical duration time and survival time distributions. For one study of sigmoidoscopic screening for colorectal cancer, when the true odds ratio is 1, its estimate is 0.50 to 0.75 under plausible assumptions. This bias can affect any observational study wherein case definition depends on diagnosis times (e.g., health-plan enrollment data). To avoid bias in observational investigations of cancer screening wherein the case definition depends on the diagnosis date, one must ensure that both screening and preclinical incidence do not occur before the case definition period. PMID- 10529026 TI - The repeatability of three methods for measuring prospective patients' values in the context of treatment choice for end-stage renal disease. AB - In the context of the choice of treatment for end-stage renal disease (ESRD), three approaches to value assessment were examined for their repeatability over time within subjects. If formal decision analyses are to be used to advise patients about treatment choice, then repeatable value assessment methods, an essential component of such analyses, are needed. The methods assessed were standard gamble (SG), time trade-off (TTO), and visual analogue (VA). Sixty-six nephrology clinic patients were interviewed on two occasions, 10 days apart, by one of two interviewers. An information session was conducted 1 week before the first interview. Subjects were taught about the treatments using an information package developed expressly for the study and a video produced by a pharmaceutical company for use in this decision context. Patients differed widely in the values provided for the various treatments of ESRD, with responses that ranged across the entire scale (0 to 100). The repeatability of the three methods was poor, with the coefficients of repeatability (95% range of differences from one occasion to the next) observed as +/- 27.4 for SG, +/- 38.4 for TTO, and +/- 36.5 for VA. When subsets defined by characteristics that may have improved the repeatability were analyzed, the magnitude of the error did not vary substantially. The poor repeatability of these methods raises questions about their use for decision analyses applied to the individual context. PMID- 10529027 TI - Further evidence supporting an SEM-based criterion for identifying meaningful intra-individual changes in health-related quality of life. AB - This study used the standard error of measurement (SEM) to evaluate intra individual change on both the Chronic Respiratory Disease Questionnaire (CRQ) and the SF-36. After analyzing the reliability and validity of both instruments at baseline among 471 COPD outpatients, the SEM was compared to established minimal clinically important difference (MCID) standards for three CRQ dimensions. A value of one SEM closely approximated the MCID standards for all CRQ dimensions. This SEM-based criterion was then validated by cross-classifying the change status (improved, stable, or declined) of 393 follow-up outpatients using the one SEM criterion and the MCID standard. Excellent agreement was achieved for all three CRQ dimensions. Although MCID standards have not been established for the SF-36, the one-SEM criterion was explored in these change scores. Among SF-36 scales demonstrating acceptable reliability and reasonable variance, the percent of individuals within each change category was consistent with those seen in the CRQ dimensions. These results replicate previous findings where a value of one SEM also closely approximated MCIDs for all dimensions of the Chronic Heart Disease Questionnaire among cardiovascular outpatients. The one-SEM criterion should be explored in other health-related quality of life instruments with established MCIDs. PMID- 10529028 TI - Test-retest reliability of the GO-QOL: a disease-specific quality of life questionnaire for patients with Graves' ophthalmopathy. AB - To assess the test-retest reliability of a recently developed disease-specific quality of life questionnaire for evaluative studies in patients with Graves' ophthalmopathy (the GO-QOL), measuring visual functioning and psychosocial consequences of changed appearance. Ninety-three patients were included and completed the GO-QOL. Additional information on general quality of life and disease characteristics was obtained. Construct validity and internal consistency of the two subscales was determined, based on principal component analyses, Cronbach's alpha's and correlations with MOS-24, three subscales of the SIP, and clinical measures. Eighty-nine patients completed a second GO-QOL after two weeks including four additional questions about perceived changes in health status. Test-retest reliability was assessed by calculating intraclass correlation coefficients (ICCs) and limits of agreement, using several definitions of stable patients. Slight modifications from the original questionnaire were evaluated for their effect on the validity and reliability. The construct validity of the two subscales was confirmed and Cronbach's alpha's were 0.89 for visual functioning and 0.87 for appearance. The substantial ICCs found for both scales of the GO-QOL (ICCs above 0.80) reflect that the errors of measurement were relatively small, which supports the value of this questionnaire for clinical studies with relatively small sample sizes. The modification of the appearance scale improved the validity of the scale and resulted in less missing values. Following the recommendations of the joint committee of thyroid associations, we recommend the inclusion of HRQL-measures in clinical studies that evaluate treatments for patients with GO. The GO-QOL is a promising tool for this purpose. PMID- 10529029 TI - A comparison of C/B ratios from studies using receiver operating characteristic curve analysis. AB - In receiver operating characteristic (ROC) curve analysis, the optimal cutoff value for a diagnostic test can be found on the ROC curve where the slope of the curve is equal to (C/B) x (1-p[D])/p[D], where p[D] is the disease prevalence and C/B is the ratio of net costs of treating nondiseased individuals to net benefits of treating diseased individuals. We conducted a structured review of the medical literature to examine C/B ratios found in ROC curve analysis. Only two studies were found in which a C/B ratio was explicitly calculated; in another 11 studies, a C/B ratio was based on a so-called holistic estimate, an all-encompassing educated estimate of the relative costs and benefits relevant to the clinical situation. The C/B ratios ranged from 0.0025 (tuberculosis screening) to 2.7 (teeth restoration for carious lesions). Clinical scenarios that are directly life threatening but curable had C/B ratios of less than 0.05. This analysis led us to construct a table of ordered C/B ratios that may be used by investigators to approximate C/B ratios for other clinical situations in order to establish cutpoints for new diagnostic tests. PMID- 10529030 TI - Finding bile duct injuries using record linkage: a validated study of complications following cholecystectomy. AB - Laparoscopic cholecystectomy was introduced to Western Australia in 1991 and has become the method of choice for this procedure, although there are concerns about complications, particularly bile duct injuries. Previous studies have investigated this problem but have not confirmed the accuracy of coded information. We used Record Linkage to link operative admissions to subsequent admissions for all people who underwent cholecystectomy between 1988 and 1994. Using ICD9-CM discharge codes, we identified patients with an associated complication. We validated these patients' medical notes to obtain the proportion of complications in the period encompassing the introduction of laparoscopic cholecystectomy. We found 48 bile duct injuries in 413 patients. Of these 43% were found using complication codes on the operative admission, 79% using linked records of subsequent admissions, and 90% by adding lists of complicated cases from the three teaching hospitals. Any epidemiological research that uses surgical complication codes from operative admissions, particularly in the absence of a specific ICD9-CM code, will lead to significantly underestimating the prevalence of complications. By using record linkage, and validating medical records, we captured a significant proportion of complications. PMID- 10529031 TI - Uses of ecologic studies in the assessment of intended treatment effects. PMID- 10529032 TI - Oral health and nutrition. PMID- 10529033 TI - Does everyone in heart failure need echocardiography? PMID- 10529034 TI - Osteoporosis: fact, fiction, fallacy and the future. PMID- 10529035 TI - Factors predictive of outcome on admission to an acute geriatric ward. AB - AIM: To investigate which factors predict outcome of elderly patients on discharge and at 6 months. METHODS: A prospective study in an acute geriatric ward. Within 48 h of admission, patients were assessed for social factors, geriatric problems, admission diagnoses, medication, function and mental ability. Outcome measures were mortality, length of stay, institutionalization, readmissions and attendance at accident and emergency within 6 months. RESULTS: 353 patients were studied, with a mean age of 81.8 years. Logistic regression analyses showed that variables predicting hospital mortality were Barthel index on admission, pre-morbid disability and polypharmacy. The only variable independently predictive of prolonged stay in hospital was a Barthel score of <45 on admission. Functional disability on admission was predictive of institutionalization on discharge. Variables predicting mortality within 6 months of discharge were Barthel index on admission <65, presence of pressure sores, malnutrition and polypharmacy. Variables independently predictive of institutionalization were mental state and a low pension. Those who took more than five drugs on admission were more likely to attend accident and emergency and be readmitted. CONCLUSION: Limited activities of daily living and geriatric problems on admission are the strongest predictive factors of outcome, independent of diagnoses. PMID- 10529036 TI - Observing the process of care: a stroke unit, elderly care unit and general medical ward compared. AB - BACKGROUND AND PURPOSE: Patients on stroke units have better outcomes but it is not known why. We investigated the process of care on a stroke unit, an elderly care unit and a general medical ward. METHODS: Comparison of the three settings was by non-participant observation of 12 patients in each. Data were analysed using multi-level modelling methods. RESULTS: Stroke unit patients spent more time out of bed and out of their bay or room, and had more opportunities for independence than patients on the medical ward. There were more observed attempts on the stroke unit than on the general medical ward to interact with drowsy, cognitively- or speech-impaired patients. Stroke unit patients spent more time with visitors. Most of these aspects of care were also found on the elderly care unit, where patients also spent less time asleep or 'disengaged', more time interacting with nurses, and were given appropriate help more often than those elsewhere. Stroke unit patients received less eye contact, were ignored and treated in a dehumanizing way more frequently and had more negative interactions or activities than those elsewhere. CONCLUSIONS: We have identified some aspects of the process of care which may help explain the improved outcomes on stroke units. These aspects were also observed in the elderly care unit. PMID- 10529037 TI - The influence of randomized trials on the use of anticoagulants for atrial fibrillation. AB - INTRODUCTION: Anticoagulants and anti-platelet drugs have been shown in randomized trials to reduce the risk of stroke in patients with atrial fibrillation (AF). We therefore investigated their use in patients known to be in AF before a stroke, transient ischaemic attack (either cerebral or ocular) or retinal artery occlusion to assess the influence of trials on clinical practice. METHODS: Inpatients and outpatients with acute stroke, transient ischaemic attack or retinal artery occlusion were prospectively identified by a stroke physician from 1990 to 1997. The presence or absence of AF before the vascular event, and prior use of anticoagulant and anti-platelet drugs were recorded at the time of the assessment and verified using information from general practitioner and hospital case notes. RESULTS: Of 1934 patients with stroke or retinal artery occlusion, 191 (10%) were in AF before their ischaemic event. Anticoagulants had been used in 40 (21%) of these, but only in 32 (2%) of the 1743 patients in sinus rhythm [odds ratio (OR) 14.2, 95% confidence interval (CI) 8.6-23.2]. Anti platelet drugs had been used in 62 (32%) of those with AF compared with 500 (30%) of those in sinus rhythm (OR 1.2, 95% CI 0.9-1.64). Of the 161 patients in AF without contraindications to anticoagulants, only 36 (22%) were taking them. Although there was a statistically significant increase in anticoagulant use from 8% in 1990 to 23% in 1996, this could be explained solely by a fall in the age of the patients referred to our hospital. CONCLUSION: Anticoagulation is probably under-used in AF. We found no conclusive evidence that anticoagulation trials have influenced clinical practice. This raises issues about the dissemination and implementation of trial results. PMID- 10529038 TI - Evaluation of echocardiography in the management of elderly patients with heart failure. AB - OBJECTIVE: To determine the validity of a clinical diagnosis of systolic dysfunction in elderly patients with heart failure and assess the contribution of echocardiography to their management. SUBJECTS: 61 elderly patients with a diagnosis of heart failure in a geriatric assessment unit setting. METHODS: Prospective study determining sensitivity, specificity and predictive values of a clinical and radiological diagnosis compared with echocardiographic standard. Proposed management was compared before and after echocardiography. RESULTS: Clinical assessment was highly sensitive (93%) but lacked specificity (32%). Combining radiological and clinical diagnoses increased specificity to 58%. Echocardiography revised the lead cardiac diagnosis for 28% of patients and influenced patient management plans for 41%. CONCLUSION: For elderly patients with heart failure, echocardiography improves diagnostic accuracy and identifies those patients with potential to benefit from angiotensin-converting enzyme inhibitors. PMID- 10529039 TI - Amlodipine lowers blood pressure without affecting cerebral blood flow as measured by single photon emission computed tomography in elderly hypertensive subjects. AB - AIM: To evaluate the effect of amlodipine on blood pressure and cerebral blood flow in elderly subjects with mild to moderate hypertension. METHODS: A double blind, parallel group study of 26 patients. After a 4-week placebo run-in period, amlodipine (5-10 mg) or matching placebo was given once daily for 8 weeks. RESULTS: Amlodipine significantly reduced blood pressure compared with baseline. Diastolic blood pressure was significantly reduced by amlodipine compared with placebo (P< 0.02 to P< 0.01). Ambulatory blood pressure monitoring showed that blood pressure control was sustained over the 24-h dosing interval. Relative regional cerebral blood flow, assessed using single photon emission computed tomography, was not significantly affected by amlodipine. Three placebo patients, but no amlodipine patients, withdrew because of adverse events. CONCLUSION: Amlodipine was a well-tolerated and effective antihypertensive agent, and did not reduce regional cerebral blood flow in elderly hypertensive patients. PMID- 10529041 TI - Relationship between oral health and nutrition in very old people. AB - OBJECTIVE: To evaluate the relationship between oral health status and nutritional deficiency. DESIGN: Cross-sectional clinical study. SUBJECTS: 324 institutionalized frail older adults (mean age 85). MEASUREMENTS: Structured oral examination including an evaluation of mucosa, periodontal state, caries prevalence and denture quality. The nutritional status was assessed using serum albumin concentration and the body mass index. Physical dependence was assessed using the Barthel index. To identify oral health disorders associated with markers of malnutrition we performed the Pearson chi2 test separately for edentulous and dentate patients. Subjects with at least one of the identified oral disorders were classified as having compromised oral functional status. RESULTS: About two-thirds of the subjects were functionally dependent and half had either a body mass index <21 kg/m2 or serum albumin <33 g/l. Among the edentulous, wearing dentures with defective bases or not wearing dentures at all were the factors most associated with malnutrition. In dentate subjects, corresponding identifiers were the number of occluding pairs of teeth (five or fewer, either natural or prosthetic), the number of retained roots (four or more), and the presence of mobile teeth. According to these criteria, 31% of the subjects had a compromised oral functional status. This was more frequently found in dependent subjects (37%) than semi-dependent subjects (18%; odds ratio, 2.6; 95% confidence interval, 1.4-4.8). Those with compromised oral functional status had a significantly lower body mass index and serum albumin concentration. CONCLUSION: Specific detrimental oral conditions are associated with nutritional deficiency in very old people. PMID- 10529040 TI - Nutritional differences in patients with proximal femoral fractures. AB - BACKGROUND: The reason why elderly human hips tend to break in one of two anatomical regions is uncertain. Nutritional factors may affect the site of fracture. OBJECTIVE: To assess possible nutritional differences in patients with proximal femoral fractures. DESIGN: Prospective observational cohort study. SETTING: University teaching hospital. SUBJECTS: 119 consecutive patients over the age of 65 with a hip fracture admitted to the trauma wards in a single centre. METHODS: One researcher measured triceps, biceps and supra-iliac skinfold thickness, and mid upper arm circumference on admission and on the fifth post operative day. Body mass index was calculated for each patient, and used to classify patients as severely, moderately or mildly malnourished, normal, overweight or obese. Logistic regression was used to determine the influence of various factors on fracture site. RESULTS: According to their body mass index, 31% of patients were classified as malnourished and 11% as severely malnourished. Patients with intracapsular fractures were significantly more malnourished than patients with trochanteric fractures (P < 0.008). Nutritional status was not related to post-operative complications. Ability to weigh a patient on the fifth post-operative day was the single most important prognostic indicator for complications. CONCLUSIONS: Patients with intracapsular fractures are more malnourished. Those with trochanteric fractures tend to be overweight. PMID- 10529042 TI - Energy intake and micronutrient intake in elderly Europeans: seeking the minimum requirement in the SENECA study. AB - OBJECTIVE: To examine energy intake of elderly people participating in the Survey in Europe on Nutrition and the Elderly, a Concerted Action (SENECA) study in relation to the adequacy of micronutrient intake. DESIGN: Data from eight countries on 486 men and 519 women who were 74-79 years old. Dietary intakes of energy, iron, thiamine, riboflavin and pyridoxine were calculated. RESULTS: There was inadequate intake of one or more nutrients in 23.9% of men and 46.8% of women. The prevalence of inadequate intakes decreased gradually with higher energy intakes. Of all people with energy intakes exceeding 1500 kcal, 19% of men and 26% of women still had an inadequate intake of at least one micronutrient. CONCLUSION: We found no single criterion ensuring level of energy intake with an adequate micronutrient supply. The prevalence of an inadequate intake of micronutrients was high at all energy intake levels, especially in women. PMID- 10529043 TI - Morbidity in older people with self-reported asthma. AB - OBJECTIVE: To investigate the differences in physical and psychological morbidity in older people with and without self-reported asthma and whether these are associated with use of more medication and hospital services. DESIGN: Cross sectional study of changes in health services for older people. SETTING: South Wales in 1990 and 1992. SUBJECTS: A population-based random sample of 2818 people aged 65 years and over. MAIN OUTCOME MEASURES: Prevalence of self-reported asthma; assessment of disability, anxiety, depression and memory using standardized measures; mobility; use of prescribed medication and hospital services. RESULTS: 231 subjects with self-reported asthma were identified. The prevalence of asthma was 8%, which was not significantly different between the sexes (P = 0.88), age groups (P = 0.06) or social classes (P = 0.108). There was a significant relationship between asthma and functional and physical disability (severe disability 29% vs 16%, P < 0.0001), mobility (housebound 7% vs 4%, P < 0.05), anxiety (37% vs 20%, P < 0.0001), depression (19% vs 10%, P < 0.001), poor perceived health status (23% vs 9%, P < 0.0001), number of different medications (seven or more, 13% vs 4%, P < 0.0001) and inpatient (P < 0.0001) and outpatient (P < 0.05) use of hospital services. CONCLUSIONS: There is excess psychological and physical morbidity and poorer perceived health status in older people with self-reported asthma. PMID- 10529044 TI - A comparison of large volume spacer, breath-activated and dry powder inhalers in older people. AB - AIMS: To see if elderly people can use the breath-activated (Easi-breathe) and dry powder (Turbohaler) inhalers as effectively as the metered-dose inhaler and Volumatic system. METHODS: 102 inhaler-naive patients (aged 75-101, mean 84 years), without cognitive impairment, were randomly allocated one of Easi breathe, Turbohaler or metered-dose inhaler and Volumatic placebo inhalers. Standardized tuition was done on enrolment and at 6 h review. Inhaler technique was assessed immediately after enrolment tuition and at 6 and 24 h. Assessment was by scoring (0 = poor, 1 = moderate, 2 = perfect) five aspects of technique. RESULTS: Mean total scores were significantly (P < 0.005) higher for Turbohaler and Easi-breathe than metered-dose inhaler and Volumatic on enrolment and 6 h and at 24 h (P < 0.045). Fewer patients achieved excellent scores of 9 or 10 when using metered-dose inhaler and Volumatic. The main difficulties were in assembling the metered-dose inhaler and Volumatic and detecting when the metered dose inhaler and Volumatic or Easi-breathe was empty. CONCLUSIONS: Breath activated and dry powder inhalers were more likely to be used correctly than metered-dose inhalers with large volume spacers. PMID- 10529045 TI - Age, deprivation and rates of inguinal hernia surgery in men. Is there inequity of access to healthcare? AB - OBJECTIVES: To study trends in hospital admissions for inguinal hernia surgery in men, examining relationships between age, deprivation and rate of surgery. DESIGN: graphical analyses of hospital discharge data and demographic information, guided by three hypotheses on urgency of surgery, age and evidence of discordance between population prevalence of disease and rates of surgery. SETTING AND SUBJECTS: Men undergoing inguinal hernia surgery in Scotland in 1982 4, 1987-9 and 1992-4. MAIN OUTCOME MEASURES: Rate of operation per 100 000 population. RESULTS: Over the study period, there has been (i) a marked increase in the rate of elective hernia operations in the over-65s, (ii) a stable rate of non-elective operations in all age groups, (iii) a lower rate of elective surgery in patients from deprived areas than in patients from affluent areas. CONCLUSIONS: During the period studied there has been decreasing inequity on the grounds of age but persisting inequity on the grounds of deprivation. These techniques of analysis are potentially applicable to many conditions and may be useful in equity audit in patients of all ages. PMID- 10529046 TI - Changes in and factors related to loneliness in older men. The Zutphen Elderly Study. AB - AIM: To investigate (i) whether loneliness increases in old age, and if so, whether it relates to ageing itself, to time trends or to cohort effects and (ii) the relationship between changes in institutionalization, partner status and health and loneliness. METHODS: 939 men born between 1900 and 1920 completed the De Jong-Gierveld Loneliness Scale, and answered questions about their partner status, health and institutionalization in 1985, 1990 and 1995. RESULTS: For the oldest group (born between 1900 and 1910) loneliness scores increased, but not for the younger groups. The increase in loneliness was attributable to ageing. No birth cohort or time effects were found. Loneliness was related to changes in institutionalization, partner status and subjective health but not to limitations in activities of daily living or cognitive function. CONCLUSIONS: the increased loneliness experienced by very old men is influenced by loss of a partner, moving into a care home or not feeling healthy. PMID- 10529047 TI - Hip fracture with correctly positioned external hip protector. PMID- 10529048 TI - Outcomes of percutaneous endoscopic gastrostomy feeding. PMID- 10529050 TI - Readmission of patients discharged from emergency departments. PMID- 10529049 TI - Endoscopic retrograde cholangiopancreatography in elderly patients. PMID- 10529051 TI - Parallel funeral services. PMID- 10529052 TI - Dialysis in old age--are we really doing all we should? PMID- 10529053 TI - Histological and immunohistological investigation of lymphoid tissue in the platypus (Ornithorhynchus anatinus). AB - The gross and histological appearance and the distribution of T and B lymphocytes and plasma cells are described for lymphoid tissues obtained from 15 platypuses. The spleen was bilobed and surrounded by a thick capsule of collagen, elastic fibres and little smooth muscle. White pulp was prominent and included germinal centres and periarterial lymphoid sheaths. Red pulp contained haematopoietic tissue. A thin lobulated thymus was located within the mediastinum overlying the heart. The cortex of lobules consisted of dense aggregates of small and medium lymphocytes, scattered macrophages and few reticular epithelial cells. In the medulla, Hassall's corpuscles were numerous, lymphocytes were small and less abundant, and reticular cells were more abundant than in the cortex. Lymphoid nodules scattered throughout loose connective tissue in cervical, pharyngeal, thoracic, mesenteric and pelvic sites measured 790 +/- 370 microm (mean +/- S.D., n = 39) in diameter, the larger of which could be observed macroscopically. These consisted of single primary or secondary follicles supported by a framework of reticular fibres. Macrophages were common in the germinal centres. The platypus had a full range of gut-associated lymphoid tissue. No tonsils were observed macroscopically but histologically they consisted of submucosal follicles and intraepithelial lymphocytes. Peyer's patches were not observed macroscopically but histologically they consisted of several prominent submucosal secondary follicles in the antimesenteric wall of the intestine. Caecal lymphoid tissue consisted of numerous secondary follicles in the submucosa and densely packed lymphocytes in the lamina propria. Bronchus-associated lymphoid tissue was not observed macroscopically but was identified in 7 of 11 platypus lungs assessed histologically. Lymphoid cells were present as primary follicles associated with bronchi, as aggregates adjacent to blood vessels and as intraepithelial lymphocytes. The distribution of T lymphocytes, identified with antihuman CD3 and CD5, and B lymphocytes and plasma cells, identified with antihuman CD79a and CD79b and antiplatypus immunoglobulin, within lymphoid tissues in the platypus was similar to that described in therian mammals except for an apparent relative paucity of B lymphocytes. This study establishes that the platypus has a well developed lymphoid system which is comparable in histological structure to that in therian mammals. It also confirms the distinctiveness of its peripheral lymphoid tissue, namely lymphoid nodules. Platypus lymphoid tissue has all the essential cell types, namely T and B lymphocytes and plasma cells, to mount an effective immune response against foreign antigens. PMID- 10529054 TI - Inhibition of dendrite formation in mouse melanocytes transiently transfected with antisense DNA to myosin Va. AB - In mice a molecular motor of the myosin V class (designated myosin Va) is known to be the product of the dilute locus, where a mutation prevents melanosome transport in melanocytes. There is conflicting evidence about whether it has a role in dendrite outgrowth. We investigated its role by transiently transfecting antisense oligonucleotides to inhibit its expression in a melanocyte cell line. We demonstrated mRNA and protein expression of myosin Va in 3 mouse melanocyte lines and 1 human melanoma cell line, using RT-PCR and immunoblotting. Two splice variants were found in human cells whilst only the longer transcript, containing an additional exon, was present in mouse melanocyte lines. The shorter variant was detected in other mouse tissues. Myosin Va protein levels were similar in 3 melanocyte lines with differing amounts of pigmentation, indicating that expression of myosin Va is not tightly coupled to expression of melanin. Immunocytochemistry showed 2 types of myosin Va localisation. A punctate pattern of staining concentrated in the perinuclear region was indicative of organelle association, and the observation of occasional linear punctate staining aligned with F-actin bundles supported the idea that myosin Va has a role in transporting melanosomes along actin filaments. Staining was also intense at tips of dendrites and at sites of dendrite-cell contact, consistent with a possible role in dendrite growth. Transient transfection of antisense phosphorothioate oligodeoxynucleotides targeted against myosin Va mRNA reduced expression of myosin Va protein in cultured mouse melan-a melanocytes by over 70 % 20 h after transfection whereas a control (shuffled sequence) oligonucleotide did not. Upon trypsinisation and replating these cells the capacity of the transfected cells to extend new dendrites was reduced in the cells containing the specific antisense oligonucleotides but unaffected by the control oligonucleotide. Image analysis confirmed that the effect of transfection on morphology was statistically significant (P < 0.01). In contrast when cells were not trypsinised and replated following transfection so that previously existing dendrites could persist, the normal dendritic morphology continued to be observed. We conclude that in addition to its involvement in melanosome transport, myosin Va has a role in the extension of new dendrites by melanocytes but not in maintenance of pre-existing dendrites. PMID- 10529055 TI - Relationship between accessory foramina and tumour spread in the lateral mandibular surface. AB - The spread of tumour cells to the mandible has been well recognised and invasion of the edentulous alveolar ridge by tumour through accessory foramina has been documented. Tumour infiltration can also occur through the lateral cortical plate, but the number and distribution of accessory foramina on this surface has not been reported. Lateral surfaces of 89 mandibles were examined and accessory foramina which showed a direct communication with the underlying cancellous bone were charted. It was found that the number of accessory foramina varied greatly from specimen to specimen. Only 70.8 % of mandibles showed foramina in the coronoid, sigmoid and condylar sections; of these 93.7 % exhibited foramina in the condylar section, 23.8% in the coronoid and only 19 % in the sigmoid section. This finding confirms that the current practice of conserving part of the ascending ramus posterior to the coronoid process following surgery is sound. Similarly in the rest of the lateral surface, foramina were present in the upper third section in 97.8 % of mandibles, 61.8% in the lower third and 58.4 % in the middle third sections. This result justifies the principle of rim resection in appropriate cases and the recognition that the alveolar section is commonly invaded before the rest of the body. The number and distribution of foramina may be of greater significance following radiotherapy when the foramina could provide multiple direct channels for invasion of tumour cells from the lateral surface to the medulla. PMID- 10529056 TI - A densitometric analysis of the human first metatarsal bone. AB - Bone responds to the stresses placed on it by remodeling its structure, which includes shape, trabecular distribution and density distribution. We studied 49 pairs of cadaveric human 1st metatarsal bones in an attempt to establish the pattern of density distribution and to correlate it with the biomechanical function of the bone. We found that the head is denser than the base, the dorsal portion of the whole metatarsal is denser than the plantar portion and the lateral portion of the whole metatarsal is denser than the medial aspect. The same pattern of density with respect to dorsal vs plantar and lateral vs medial was also seen in the head when it was examined alone. When we compared the 4 portions of the head with the same portion of the metatarsal as a whole we found that only the medial portion of the head was less dense than its respective portion of the whole metatarsal. All of these patterns of density distribution are consistent with respect to age, sex and laterality. We have also hypothesised as to the relationship between density distribution seen both in the whole metatarsal and in the metatarsal head and their biomechanical function in the gait cycle. PMID- 10529057 TI - Mechanism of change in the orientation of the articular process of the zygapophyseal joint at the thoracolumbar junction. AB - The orientation of the superior articular processes in thoracic and lumbar vertebrae differs. The present study was undertaken to investigate the possible mechanism for the change from a posterolaterally facing superior articular surface in the thoracic region to a posteromedially facing curved articular surface in the lumbar region. The material of the study consisted of dry macerated bones of 44 adult human vertebral columns. The orientation of the superior articular process and its relation to the mamillary tubercle (process) was examined between T9 and L5 vertebrae in each column. An abrupt change from the thoracic to lumbar type of articular process was observed in 3 columns (7 %). Forty-one (93 %) columns showed a gradual change extending over either 2 or 3 successive vertebrae. The present study suggests that the change in the orientation of the superior articular process, from the coronal to the sagittal plane (sagittalisation), occurs due to the change in the direction of weight transmission through zygapophyseal joints at the thoracolumbar junction. It was observed that the gradual sagittalisation of the superior articular process in the transitional zone brought it close to the mamillary tubercle which eventually fused with it. Thus the study suggests that the characteristic posteromedially facing concave superior articular process of lumbar vertebrae may have formed because of the fusion of the articular process and the mamillary tubercle. PMID- 10529058 TI - A study of motoneuron groups and motor columns of the human spinal cord. AB - Eight normal human spinal cords were studied for motoneuron (Mn) groups and columns. Spinal segments (C1 to Coc.) were identified and embedded in paraffin wax. Serial cross sections were cut at 25 microm and stained by cresyl violet. Cross-sectional profiles of the spinal cord were traced for each segmental level and the outlines of the various Mn groups superimposed. These charts (maps) were used to examine intra and intersegmental changes in the relative positions of the columns. An attempt was made to provide topographical picture of Mn groups of individual segments. In the cervical region neuronal groups were more numerous but smaller and less distinct, while in the lumbosacral region they were fewer, larger and at many levels better circumscribed. The average number of Mn groups at any segmental level was 3-4 and never exceeded 5. C4, C5, C6, C7, L4, L5 and S1 contained numerous Mn groups. Maximum intrasegmental changes were noted at C3, C4, C7, T1, and S2, while at C5, C6, all thoracic, L1 L2 and L3, the pattern was constant throughout the segment. Eleven motor columns were traced in the human spinal cord. Column 1 belonged to the medial division and columns 2-11 to the lateral division of the ventral grey horn. Columns 1 and 2 were the most extensive as they were traceable from the lower medulla to S3 segment. Columns 3 8 were confined to cervical segments (including T1), while columns 9-11 were traced in lumbosacral segments. In general, motor columns followed a definite mode for their appearance and disappearance. Many of them showed rotation from a dorsal to a ventromedial direction. PMID- 10529059 TI - Microvascular features and ossification process in the femoral head of growing rats. AB - In the epiphysis of long bones, different patterns of development of ossification processes have been described in different species. The development of the vascularisation of the femoral head has not yet been fully clarified, although its role in the ossification process is obvious. Our aim was to investigate ossification and vascular proliferation and their relationship, in growing rat femoral heads. Male Wistar rats aged approximately 1, 5 and 8 wk and 4, 8 and 12 mo were used. Light microscopy frontal sections and vascular corrosion casts observed by scanning electron microscopy were employed. In the rat proximal femoral epiphysis, ossification develops from the medullary circulation of the diaphysis, quickly extending to the neck and the base of the head. Hypertrophic chondrocytes occupy the epiphyseal cartilage, and a physeal plate with regular cell columns is present. Starting from about the end of the third month one or more points of fibrovascular outgrowth, above the physeal line, can be observed in each sample. They are often placed centrally or, sometimes, peripherally. The fibrovascular outgrowths penetrate deeply into the cartilage and extend laterally. At age 8 mo, large fibro-osseous peduncles connect the epiphysis to the diaphyseal tissue. At 12 mo, the entire epiphysis appears calcified with an almost total absence of residual cartilage islands. This situation differs in man and in other mammals due both to differing thickness of the cartilage and to the presence of more extensive sources of blood vessels other than the diaphyseal microcirculation, as supplied by the teres ligament and Hunter's circle. In young rats, subchondral vessels and the synovial fluid could play a role in feeding the ossifying cartilage. Later, a loss of resistance of the physis due to marked degeneration of the cell columns, and extensive chondrocyte hypertrophy permit fibrovascular penetration starting from diaphyseal vessels rather than neighbouring vascular territories, such as those of the periosteum and capsule. PMID- 10529060 TI - Gross and microscopic visceral anatomy of the male Cape fur seal, Arctocephalus pusillus pusillus (Pinnipedia: Otariidae), with reference to organ size and growth. AB - The gross and microscopic anatomy of the Cape fur seal heart, lung, liver, spleen, stomach, intestine and kidneys (n = 31 seals) is described. Absolute and relative size of organs from 30 male seals are presented, with histological examination conducted on 7 animals. The relationship between log body weight, log organ weight and age was investigated using linear regression. Twenty five animals were of known age, while 6 were aged from counts of incremental lines observed in the dentine of tooth sections. For the range of ages represented in this study, body weight changes were accurately described by the exponential growth equation, weight = w(o)r(t), with body weight increasing by 23 % per annum until at least 9-10 y of age. Organ weight increased at a rate of between 25 % and 33 % per annum until at least 9-10 y of age, with the exception of the intestines, where exponential increase appeared to have ceased by about 7 y. The relationship between body weight and organ weight was investigated using logarithmic transformations of the allometric equation, y = ax(b), where the exponent b is 1 if organ weight is proportional to body weight. Most organs increased in proportion to the body. However, the heart, liver and spleen had exponents b > 1, suggesting that these organs increased at a faster rate than the body. The basic anatomical features of the viscera were similar to those of other pinnipeds, with some exceptions, including the arrangement of the multilobed lung and liver. Apart from the large liver and kidneys, relative size of the organs did not differ greatly from similar sized terrestrial carnivores. The histological features of the organs were generally consistent with those previously described for this species and other otariids. The heart, as in other pinnipeds, was unlike that of cetacea in not having unusually thick endocardium or prominent Purkinje cells. Notable histological features of the lungs included prominent fibrous septa, prominent smooth muscle bundles, cartilage extending to the level of the alveolar sacs and ample lymphoid tissue. The spleen had a thick capsule, well developed trabeculae and plentiful plasma cells. Abundant parietal cells were present in the fundic glands and lymphoid follicles were present in the gastric lamina propria, particularly in the pyloric region. Small intestinal villi were very long but this could have resulted from underlying chronic inflammation. Lymphoid follicles were prominent in the colon. The kidney reniculi each had a complete cortex, medulla and calyx, but a sportaperi medullaris musculosa was not identified. PMID- 10529061 TI - Specialised sympathetic neuroeffector associations in immature rat iris arterioles. AB - Sympathetic nerve-mediated vasoconstriction in iris arterioles of mature rats occurs via the activation of alpha(1B)-adrenoceptors alone, while in immature rat iris arterioles, vasoconstriction occurs via activation of both alpha1- and alpha2-adrenoceptors. In mature rats the vast majority of sympathetic varicosities form close neuroeffector junctions. Serial section electron microscopy of 14 d iris arterioles has been used to determine whether restriction in physiological receptor types with age may result from the establishment of these close neuroeffector junctions. Ninety varicosities which lay within 4 microm of arteriolar smooth muscle were followed for their entire length. Varicosities rarely contained dense cored vesicles even after treatment with 5 hydroxydopamine. 47 % of varicosities formed close associations with muscle cells and 88 % formed close associations with muscle cells or melanocytes. Varicosities in bundles were as likely as single varicosities to form close associations with vascular smooth muscle cells, although the distribution of synaptic vesicles in single varicosities did not show the asymmetric accumulation towards the smooth muscle cells seen in the varicosities in bundles which were frequently clustered together. We conclude that restriction of physiological receptor types during development does not appear to correlate with the establishment of close neuroeffector junctions, although changes in presynaptic structures may contribute to the refinement of postsynaptic responses. PMID- 10529062 TI - Expression of three oligosaccharide conjugates by neonatal rat dorsal root ganglion neurons: comparison with CGRP and GAP43 immunoreactivity. AB - Adult dorsal root ganglion neurons express oligosaccharides conjugated to lipids that may be involved in cell-cell recognition, and consequently in the laminar organisation of their central terminations. This paper describes an immunohistochemical study of the developmental expression of 2 lactoseries (LA4 and LD2) and 1 globoseries (SSEA4) oligosaccharide conjugates in rats from embryonic d 19 to postnatal d 60. The expression of calcitonin gene related peptide and the growth associated protein GAP43 was also examined for comparative purposes. We found that these oligosaccharide conjugates begin to be expressed after birth, suggesting that they may be involved in maturation of the central or peripheral terminations, rather than axonal guidance. PMID- 10529063 TI - Electron microscopic stereological study of collagen fibrils in bovine articular cartilage: volume and surface densities are best obtained indirectly (from length densities and diameters) using isotropic uniform random sampling. AB - Results obtained by the indirect zonal isotropic uniform random (IUR) estimation were compared with those obtained by the direct point and interception counting methods on vertical (VS) or IUR sections in a stereological study of bovine articular cartilage collagen fibrils at the ultrastructural level. Besides comparisons between the direct and indirect estimations (direct IUR vs indirect IUR estimations) and between different sampling methods (VS vs IUR sampling), simultaneous comparison of the 2 issues took place (direct VS vs indirect IUR estimation). Using the direct VS method, articular cartilage superficial zone collagen volume fraction (Vv 41%) was 67% and fibril surface density (S(v) 0.030 nm2/nm3) 15% higher (P < 0.05) than values obtained by the indirect IUR method (V(v) 25 % and Sv 0.026 nm2/nm3). The same was observed when the direct IUR method was used: collagen volume fraction (Vv 40 %) was 63 % and fibril surface density (Sv 0.032 nm2/nm3) 21 % higher (P < 0.05) than those obtained by the indirect IUR technique. Similarly, in the deep zone of articular cartilage direct VS and direct IUR methods gave 50 and 55% higher (P < 0.05) collagen fibril volume fractions (Vv 43 and 44% vs 29%) and the direct IUR method 25% higher (P < 0.05) fibril surface density values (Sv) 0.025 vs 0.020 nm2/nm3) than the indirect IUR estimation. On theoretical grounds, scrutiny calculations, as well as earlier reports, it is concluded that the direct VS and direct IUR methods systematically overestimated the Vv and Sv of collagen fibrils. This bias was due to the overprojection which derives from the high section thickness in relation to collagen fibril diameter. On the other hand, factors that during estimation tend to underestimate Vv and Sv, such as profile overlapping and truncation ('fuzzy' profiles), seemed to cause less bias. As length density Lv and collagen fibril diameter are minimally biased by the high relative section thickness, the indirect IUR method, based on utilisation of these estimates, is here regarded as representing a 'gold standard'. The sensitivity of these 3 methods was also tested with cartilage from an in vitro loading experiment which caused tissue compression. In the superficial zone of articular cartilage Vv and Sv of collagen fibrils increased (P < 0.05). This difference in the stereological estimates was only detected by the indirect IUR estimation but not by the direct VS or direct IUR methods. This indicated that the indirect IUR estimation was more sensitive than the direct VS or direct IUR estimations. On the basis of these observations, the indirect zonal IUR estimation can be regarded as the technique of choice in the electron microscopic stereology of cartilage collagen. PMID- 10529064 TI - CT examination of the manipulation system in the giant panda (Ailuropoda melanoleuca). AB - The manipulation mechanism of the giant panda (Ailuropida melanoleuca) was examined by means of CT (computed tomography) and 3-dimensional (3-D) Volume Rendering techniques. In the 3-D images of the giant panda hand, not only the bones but also the muscular system was visualised. Sections of the articulated skeleton were obtained. It was demonstrated that the hand of the panda is equipped with separately moulded manipulation units as follows: (1) the radial sesamoid (RS), the radial carpal, and the first metacarpal (R-R-M) complex; and (2) the accessory carpal (AC) and the ulnar (A-U) complex. When the giant panda grasps anything, the R-R-M complex strongly flexes at the wrist joint, the RS becomes parallel with the AC, and the phalanges bend and hold the object. It is shown that the well-developed opponens pollicis and abductor pollicis brevis muscles envelop and fix the objects between the R-R-M complex and the phalanges during grasping. PMID- 10529065 TI - The posterior sacral foramina: an anatomical study. AB - The vascular and nervous structures and their relations with the spinal nerve roots were examined in the 2nd, 3rd and 4th posterior sacral foramina in relation to percutaneous needle insertion for neuromodulation. A foraminal branch provided by the lateral sacral artery to each foramen entered the inferior lateral quadrant of each foramen, adjacent to the nerve root medially. Facing the posterior sacral aperture and around the sacral nerves, there was no venous plexus. A venous plexus was sometimes present near the median line, and always around the proximal part of the spinal ganglion. The sacral nerve roots, especially the 3rd, had a long extradural course in the foramen, presenting a potential risk of nerve lesions during procedures involving needle insertion. PMID- 10529066 TI - Skeletal maturity in Pakistani children. AB - Skeletal maturity in 750 normal Pakistani children (400 males, 350 females) aged 1-18 y was determined by the Greulich-Pyle atlas system. Male children during first year and female children during first 2 y of life matured in conformity with Greulich-Pyle standards. After that period mean bone ages were lower than the American standards up to 15 y in males and 13 y in females (at or around puberty), which may be due to malnutrition, ill health or other environmental factors. After puberty bone ages were higher than the American standards indicating earlier maturity in Pakistani than Western children. Hence for the proper evaluation of skeletal age in a given region, a longitudinal study on individuals in that region to establish normal standards is necessary. PMID- 10529067 TI - Freeze-dried specimens for gross anatomy teaching. PMID- 10529068 TI - A Reinke-like inclusion within Leydig cells of the marmoset monkey (Callithrix jacchus) PMID- 10529069 TI - Predictors of an acute antidepressant response to fluoxetine and sertraline. AB - Sertraline and fluoxetine have different pharmacologic and pharmacokinetic profiles which may be of clinical relevance in the determination of response in different subtypes of depression. A randomized, double-blind, 6-week study comparing sertraline (50-100 mg/day) with fluoxetine (20-40 mg/day) in 286 outpatients with major depression, who had demonstrated comparable efficacy and tolerability for the two drugs, was analysed by subgroups of patients at baseline with melancholia, severe depression, single depressive episode, multiple depressive episodes, high anxiety, low anxiety, psychomotor retardation and psychomotor agitation. Multiple logistic regression with regressors including treatment-by-subgroup variables revealed that, within certain subgroups, the efficacy might differ substantially from that of the whole treatment group. However, the only treatment-by-subgroup interaction term that was significant was anxiety (P < 0.05). There was no evidence of interaction in single or recurrent episode subgroups, and these were not included in subsequent analyses. Subsequent two-sample statistical comparison tests of response (i.e. Hamilton Depression Scale reduction > or = 50%) rates at study endpoint between treatment groups demonstrated that patients with melancholic depression and those with symptoms of psychomotor agitation yielded a significantly greater proportion of responders with sertraline compared to fluoxetine (P < 0.05). Response rates in sertraline- and fluoxetine-treated patients, respectively, were: overall study 59%, 51%; melancholia 59%, 44%; severe depression 59%, 41%; low anxiety 71%, 55%; high anxiety 47%, 48%; psychomotor retardation, 48%, 46%; and psychomotor agitation 62%, 39%. Multiple logistic regression adjusting for possible confounding factors, that included a treatment by anxiety interaction term, also led to similar findings. In particular, the analysis showed that significant differences existed in favour of sertraline in patients with low anxiety in the melancholia and severe depression subgroups (P < 0.05), indicating that these characteristics predicted a superior response to 6 weeks of treatment with sertraline relative to fluoxetine. Sertraline also demonstrated advantages over fluoxetine on parameters such as sleep and weight disturbance in severely depressed patients, and sleep disturbance, weight, cognitive disturbance and retardation in melancholic patients. PMID- 10529070 TI - The comparative prophylactic efficacy of lithium and carbamazepine in patients with bipolar I disorder. AB - In a randomized prospective clinical trial with an observation period of 2.5 years, a subgroup analysis was carried out for the 114 patients with bipolar I disorder (DSM-IV) regarding the prophylactic efficacy of lithium and carbamazepine. Treatment outcome was evaluated taking rehospitalization, recurrence, subclinical recurrence, concomitant medication and severe adverse effects into consideration. Special interest was paid to the enzyme-inducing properties of carbamazepine, which might lessen the efficacy of psychotropic comedication. Lithium was superior to carbamazepine in bipolar I patients for various outcome criteria. Analyses in patients without psychotropic comedication indicate that the superiority of lithium is not the result of carbamazepine reducing plasma levels of concomitant drugs. PMID- 10529071 TI - Lithium versus carbamazepine in the maintenance treatment of bipolar II disorder and bipolar disorder not otherwise specified. AB - In a randomized clinical trial (MAP study), the prophylactic efficacy of lithium and carbamazepine was compared in a subgroup of patients (n = 57) who presented either a bipolar II disorder or a bipolar disorder not otherwise specified (DSM IV). During the observation period of 2.5 years, no significant differences between the drugs were found considering hospitalization, recurrences, subclinical recurrences, concomitant medication and severe side-effects. PMID- 10529072 TI - Tolerance and rebound insomnia with rapidly eliminated hypnotics: a meta-analysis of sleep laboratory studies. AB - Differences in development of tolerance and occurrence of rebound insomnia have been well established between rapidly and slowly eliminated benzodiazepine hypnotics. Based on meta-analytic methodology, this study assesses whether there are such differences among the rapidly eliminated benzodiazepine and benzodiazepine-like hypnotics (brotizolam, midazolam, triazolam, zolpidem and zopiclone). All sleep laboratory studies of these drugs (n = 137) published from 1966 to 1997 were obtained, mainly through a MEDLINE search. Rigorous selection criteria resulted in the inclusion of 75 studies employing 1276 individuals (804 insomniacs and 472 healthy volunteers). Using a mixed effects regression model, reliable estimation of the effects on insomniacs of the recommended dose of each drug could be obtained. All five rapidly eliminated hypnotics showed statistically significant initial efficacy. Tolerance with intermediate and long term use was clearly developed with triazolam and was only marginal with midazolam and zolpidem; it could not be estimated for brotizolam or zopiclone because of insufficient data. Rebound insomnia on the first withdrawal night was intense with triazolam and mild with zolpidem; data were unavailable for brotizolam and inadequate for midazolam and zopiclone. In conclusion, there are differences among the rapidly eliminated hypnotics with respect to tolerance and rebound insomnia suggesting that, in addition to short elimination half-life, other pharmacological properties are implicated in the mechanisms underlying these side-effects. PMID- 10529073 TI - Human leukocyte antigen typing, response to neuroleptics, and clozapine-induced agranulocytosis in jewish Israeli schizophrenic patients. AB - The atypical antipsychotic agent clozapine is known to be effective in schizophrenic patients refractory to other medications; however, it induces agranulocytosis in approximately 1-2%. In Jews, this complication is associated with the haplotype HLA B38,DR4,DQ3. The aim of the present study was to determine which human leukocyte antigen (HLA) antigens are involved in clozapine-induced agranulocytosis. We performed HLA typing in 88 Jewish Israeli schizophrenic patients and in 127 ethnically matched healthy individuals. Thirty-eight patients responsive to standard antipsychotic medications were treated with haloperidol, and 50 refractory patients received clozapine. A trend was noted for elevated rates of HLA B38 among control individuals and clozapine-treated patients of Ashkenazi origin compared to individuals of non-Ashkenazi origin, but the findings failed to reach statistical significance. No association was found between HLA class I antigens and the response to haloperidol or clozapine. Neutropenia developed in two clozapine-treated patients and agranulocytosis in one. Two of these three patients were of Ashkenazi origin, and both demonstrated the HLA B38 phenotype. Although the findings did not reach a statistical significance because of the small number of patients, they may support an association between clozapine-induced neutropenia/agranulocytosis and Ashkenazi origin and the HLA B38 phenotype. The rate of agranulocytosis in our sample (2%) is similar to the usual cumulative risk of agranulocytosis but in contrast to its high frequency among Jewish American patients. One possible explanation for this difference is the high rate of Ashkenazi patients in the American sample and the preponderance of non-Ashkenazi patients in our population. PMID- 10529074 TI - Tryptophan depletion: a predictor of future depressive episodes in seasonal affective disorder? AB - Patients with seasonal affective disorder (SAD) do not necessarily experience depressive episodes every winter. We assessed whether the behavioural response to tryptophan depletion in summer when patients are fully remitted and off therapy is capable of predicting a future depressive episode of SAD. In a prospective study design, we followed up 11 consenting SAD patients who had undergone tryptophan depletion during summer. We evaluated how many of these patients would develop a depressive episode in the subsequent fall/winter. Seven out of eight patients who relapsed during tryptophan depletion in summer developed a depressive episode in the subsequent winter. Two out of the three patients who did not relapse during tryptophan depletion remained well during the follow-up period. Our preliminary findings suggest that those SAD patients who develop depressive symptoms during tryptophan depletion when they are fully remitted and off therapy remain at high risk to experience a depressive episode of SAD also in the subsequent winter. PMID- 10529075 TI - Risperidone-induced absence of ejaculation. AB - The absence of ejaculation in two patients treated with risperidone is described. This side-effect has rarely been reported. The mechanism responsible for ejaculation dysfunction in risperidone treated patients is unclear, but might be due to the alpha1-adrenergic antagonist action of the drug. PMID- 10529077 TI - Cardiovascular advantages of a third generation calcium antagonist. Symposium on lacidipine. Foreword. PMID- 10529076 TI - Role of a third generation calcium antagonist in the management of hypertension. AB - Calcium antagonists are uniquely suitable for managing hypertension by virtue of their efficacy, metabolic neutrality and their ability to countervail counterregulatory adaptive changes, thereby enhancing blood pressure lowering. Recent evidence has accrued underscoring the concept that calcium antagonists are heterogeneous and consist of chemically dissimilar agents. The difference in formulations and pharmacokinetics affect clinical events including the effect on blood pressure, heart rate and the degree with which sympathetic activity is activated. Lacidipine is a new calcium antagonist that is the prototype of the lipophilic dihydropyridines. Of great importance, lacidipine has a slow onset of vasodilator/antihypertensive effect and does not promote an excessive sympathetic drive. These attributes commend its selection as an antihypertensive agent. PMID- 10529078 TI - Impact of calcium antagonists on the cardiovascular system: experience with lacidipine. AB - The evaluation of haemodynamic patterns in hypertensive patients by radionuclide techniques and a tomographic gamma camera has revealed differences between older and younger patients. In younger hypertensive patients, the hyperkinetic state is reflected in an increase in heart rate and, consequently, an increased cardiac index and left ventricular ejection fraction (LVEF) in comparison with normotensive controls. Older hypertensive patients, however, show a different haemodynamic pattern, with reduced systolic and diastolic function at rest compared with normotensive elderly people, and marked depression of cardiac reserve during exercise. Elderly hypertensive patients also show strikingly higher hyperresistance and reduced peripheral perfusion in comparison with younger hypertensive patients. These haemodynamic differences need to be taken into account when considering antihypertensive treatment. In a study in elderly hypertensive patients, lacidipine treatment (4 mg/day for 90 days) produced a significant decrease in total peripheral resistance and blood pressure, together with a reduction in left ventricular (LV) afterload and an increase in cardiac output and LVEF (tending towards normal values). The LV peak filling rate was also increased, and evaluation of systolic and diastolic cardiac reserve during exercise showed positive changes in cardiac performance. The haemodynamic changes with lacidipine were similar to those produced by other long-acting dihydropyridines [nifedipine gastrointestinal therapeutic system (GITS) and nitrendipine], but changes occurring with nisoldipine were less significant. The reduction in left ventricular hypertrophy (LVH) is an obvious goal of antihypertensive therapy, and several studies have demonstrated the effectiveness of lacidipine treatment in decreasing LVH in hypertensive patients. In hypertensive patients with associated LV dysfunction, favourable effects on global parameters of LV function similar to those with amlodipine have been noted with lacidipine. Myocardial blood flow was strikingly increased during lacidipine treatment and coronary resistance was significantly decreased, both at baseline and after maximal vasodilatation with dipyridamole. Thus, lacidipine's vasodilatory and anti-ischaemic profile makes it an appropriate choice for the treatment of hypertension. PMID- 10529079 TI - Coronary artery vasomotion and post-stenotic coronary artery blood flow after intracoronary lacidipine in patients with ischaemic heart disease: a pilot study. AB - BACKGROUND: The calcium antagonist lacidipine has been shown to be highly vasoselective and to improve myocardial perfusion in hypertensive patients. However, its effects on coronary artery vasomotility and on post-stenotic coronary flow reserve in patients with atherosclerotic heart disease are unknown. OBJECTIVES: This study was designed to investigate the acute direct effects of repeated infusions of lacidipine on epicardial coronary artery vasomotion and on post-stenotic coronary artery blood flow in patients with stable angina pectoris and angiographic evidence of coronary heart disease. METHODS: In 8 patients with stable angina and moderate to severe stenosis of the left coronary artery, measurements of epicardial dimensions (quantitative angiography) and of coronary blood flow (Doppler guidewire) distal to a stenosis were performed at baseline and after 3 repeated intracoronary boluses of 12 microg of lacidipine. Results were compared with those obtained after 10 mg of intracoronary papaverine. RESULTS: The intracoronary administration of lacidipine was well tolerated, without any adverse effects. Lacidipine significantly increased the minimal luminal diameter of the lesion (peak relative increase of 43.7%), without significant changes in heart rate and systolic aortic pressure. Intracoronary lacidipine caused a dose-dependent increase in coronary flow reserve. Maximal vasodilatory effects were equivalent to those obtained with intracoronary papaverine. CONCLUSIONS: These results suggest that lacidipine acts directly as a potent vasodilator in stenotic epicardial vessels and improves myocardial perfusion distal to a moderately severe stenosis in patients with stable angina. PMID- 10529080 TI - Safety profile of lacidipine: update from a clinical trials database. PMID- 10529081 TI - A neural network methodology of quadratic optimization. AB - According to the basic optimization principle of artificial neural networks, a novel kind of neural network model for solving the quadratic programming problem is presented. The methodology is based on the Lagrange multiplier theory in optimization and seeks to provide solutions satisfying the necessary conditions of optimality. The equilibrium point of the network satisfies the Kuhn-Tucker condition for the problem. The stability and convergency of the neural network is investigated and the strategy of the neural optimization is discussed. The feasibility of the neural network method is verified with the computation examples. Results of the simulation of the neural network to solve optimum problems are presented to illustrate the computational power of the neural network method. PMID- 10529082 TI - Asymptotic hyperstability of a class of neural networks. AB - This paper is concerned with the asymptotic hyperstability of recurrent neural networks. We derive based on the stability results necessary and sufficient conditions for the network parameters. The results we achieve are more general than those based on Lyapunov methods, since they provide milder constraints on the connection weights than the conventional results and do not suppose symmetry of the weights. PMID- 10529083 TI - An experimental comparison of neural algorithms for independent component analysis and blind separation. AB - In this paper, we compare the performance of five prominent neural or adaptive algorithms designed for Independent Component Analysis (ICA) and blind source separation (BSS). In the first part of the study, we use artificial data for comparing the accuracy, convergence speed, computational load, and other relevant properties of the algorithms. In the second part, the algorithms are applied to three different real-world data sets. The task is either blind source separation or finding interesting directions in the data for visualisation purposes. We develop criteria for selecting the most meaningful basis vectors of ICA and measuring the quality of the results. The comparison reveals characteristic differences between the studied ICA algorithms. The most important conclusions of our comparison are robustness of the ICA algorithms with respect to modest modeling imperfections, and the superiority of fixed-point algorithms with respect to the computational load. PMID- 10529084 TI - Color image correction for scanner and printer using B-spline CMAC neural networks. AB - The process of eliminating the color errors from the gamut mismatch, resolution conversion, and nonlinearity between scanner and printer is usually recognized as an essential issue of color reproduction. This paper presents a new formulation based on the generalized inverse plant control for the color error reduction process. In our formulation, the printer input and scanner output correspond to the input and output of a system plant, respectively. Obviously, if the printer input equals the scanner output, then there are no color errors involved in the entire system. In other words, the plant becomes an identity system. To achieve this goal, a plant generalized inverse should be identified and added to the original system. Since the system of a combination of both scanner and printer is highly nonlinear, CMAC-based neural networks, which have the capability to learn arbitrary nonlinearity, are applied to identify the plant generalized inverse. CMAC network is a perceptron-like feedforward structure with associative memory properties. Its memory requirements can be greatly reduced by the use of hash coding techniques. In order for CMAC networks to construct high-order, smooth, nonlinear plant inverse, more general CMAC addressing schemes have been proposed in conjunction with use of B-spline receptive functions. It is shown that B spline CMAC networks learn orders of magnitude more rapidly than typical implementations of back propagation in the multilayered neural networks, due to the local nature of its weighting updating and the finite support of B-spline receptive field functions. Finally, a number of test samples are conducted to verify the effectiveness of the proposed method. PMID- 10529085 TI - Modular neural networks: a survey. AB - Modular Neural Networks (MNNs) is a rapidly growing field in artificial Neural Networks (NNs) research. This paper surveys the different motivations for creating MNNs: biological, psychological, hardware, and computational. Then, the general stages of MNN design are outlined and surveyed as well, viz., task decomposition techniques, learning schemes and multi-module decision-making strategies. Advantages and disadvantages of the surveyed methods are pointed out, and an assessment with respect to practical potential is provided. Finally, some general recommendations for future designs are presented. PMID- 10529086 TI - Recognition of seed varieties using a temporal organisation map analysis of electrophoretic images. AB - This paper presents a method for seed varieties recognition using one-dimensional electrophoresis gels. It employs a neural network basically constituted of temporal organisation maps (TOM). The TOM model is a neural net which was initially developed for speech recognition. It can be trained to recognise words in speech by reference to the sound pattern over a sequence of time steps. Electrophoresis creates a set of bands in the gel, caused by migration of protein from the seed. Each seed variety generates a characteristic pattern. The bands are made visible by staining. They can then be imaged and digitised to create an input to a TOM, which treats the variation with distance along the lane in the same way as the time sequence for which it was originally employed. In this way the characteristic signature of a seed variety can be recognised. A set of 50 images--each containing 10 to 15 lanes--was used to train and test the performance of a neural network in recognising 75 cereal varieties. The network could achieve a recognition rate of 98%, provided that the gel was not distorted or cracked during heating or drying. Details of the design and training of the network are given. PMID- 10529088 TI - Effect of dietary docosahexaenoic acid on desaturation and uptake in vivo of isotope-labeled oleic, linoleic, and linolenic acids by male subjects. AB - The effect of dietary docosahexaenoic acid (22:6n-3, DHA) on the metabolism of oleic, linoleic, and linolenic acids was investigated in male subjects (n = 6) confined to a metabolic unit and fed diets containing 6.5 or <0.1 g/d of DHA for 90 d. At the end of the diet period, the subjects were fed a mixture of deuterated triglycerides containing 18:1n-9[d6], 18:2n-6[d2], and 18:3n-3[d4]. Blood samples were drawn at 0, 2, 4, 6, 8, 12, 24, 48, and 72 h. Methyl esters of plasma total lipids, triglycerides, phospholipids, and cholesterol esters were analyzed by gas chromatography-mass spectrometry. Chylomicron triglyceride results show that the deuterated fatty acids were equally well absorbed and diet did not influence absorption. Compared to the low-DHA diet (LO-DHA), clearance of the labeled fatty acids from chylomicron triglycerides was modestly higher for subjects fed the high DHA diet (HI-DHA). DHA supplementation significantly reduced the concentrations of most n-6[d2] and n-3[d4] long-chain fatty acid (LCFA) metabolites in plasma lipids. Accumulation of 20:5n-3[d4] and 22:6n-3[d4] was depressed by 76 and 88%, respectively. Accumulations of 20:3n-6[d2] and 20:4n 6[d2] were both decreased by 72%. No effect of diet was observed on acyltransferase selectivity or on uptake and clearance of 18:1n-9[d6], 18:2n 6[d2], and 18:3n-3[d4]. The results indicate that accumulation of n-3 LCFA metabolites synthesized from 18:3n-3 in typical U.S. diets would be reduced from about 120 to 30 mg/d by supplementation with 6.5 g/d of DHA. Accumulation of n-6 LCFA metabolites synthesized from 18:2n-6 in U.S. diets is estimated to be reduced from about 800 to 180 mg/d. This decrease is two to three times the amount of n-6 LCFA in a typical U.S. diet. These results support the hypothesis that health benefits associated with DHA supplementation are the combined result of reduced accretion of n-6 LCFA metabolites and an increase in n-3 LCFA levels in tissue lipids. PMID- 10529087 TI - Olestra formulation and the gastrointestinal tract. AB - Olestra is a mixture of compounds comprising sucrose esterified with 6-8 long chain fatty acids. It is not hydrolyzed by pancreatic lipase and as a result is not absorbed from the small intestine. Olestra in general has physical properties similar to those of a triacylglycerol with the same fatty acid composition. Foods made with olestra are virtually identical in taste and texture to those made with typical triacylglycerols. Olestra consumption does not generate hydrolytic products in the small intestine and, therefore, does not generate some of the signals that alter motility in the gastrointestinal tract. A reduction in gastroesophageal reflux with olestra, in contrast to triacylglycerols, is consistent with a lack of effect on stomach emptying. Unlike triacylglycerols that are absorbed in the proximal small intestine, olestra is distributed throughout the small intestine during transit and passes into the colon. In the colon, olestra's effects depend on its physical properties. Liquid nondigestible lipids result in separation of oil from the fecal matrix. Olestra formulations made with specific fatty acid compositions, particularly those containing a solid sucrose polyester component including behenic acid, possess appropriate rheology to hinder separation of oil from the rest of the fecal matrix, thereby reducing gastrointestinal symptoms. PMID- 10529089 TI - Fatty acid analysis of blood serum, seminal plasma, and spermatozoa of normozoospermic vs. asthenozoospermic males. AB - Docosahexaenoic acid (DHA; 22:6n-3) is found in extremely high levels in human ejaculate with the majority occurring in the spermatozoa. However, the relative concentration of DHA and other fatty acids, in blood serum, seminal plasma, and spermatozoa of asthenozoospermic vs. normozoospermic individuals is not known. We analyzed the phospholipid fatty acid composition of blood serum, seminal plasma, and spermatozoa of normozoospermic men and asthenozoospermic men in order to determine if DHA levels, as well as the levels of other fatty acids, differed. The serum phospholipid DHA levels were similar in the two groups, suggesting similar intakes of dietary DHA. On the other hand, seminal plasma levels of DHA (3.0 vs. 3.7%) and total polyunsaturated fatty acids (PUFA) (11.8 vs. 13.5%) were significantly lower in asthenozoospermic vs. normozoospermic men, respectively, while 18:1 (19.0 vs. 16.8%) and monounsaturated fatty acids (MUFA) (24.2 vs. 21.7%) were significantly higher in the asthenozoospermic vs. the normozoospermic men. Spermatozoa from asthenozoospermic men had higher levels of 18:1, 20:0, 22:0, 22:1, and 24:0 than sperm from normozoospermic men, and lower levels of 18:0 and DHA (8.2 vs. 13.8%). Furthermore, total MUFA (19.3 vs. 16.5%) was higher and total PUFA (19.0 vs. 24.0%), n-3 fatty acids (9.3 vs. 14.6%), and the ratio of n-3 to n-6 fatty acids (1.0 vs. 1.6) were lower in the asthenozoospermic men. Therefore, in asthenozoospermic individuals, lower levels of DHA in the seminal plasma, but not in the blood serum, mimic the decreased concentrations of DHA in the spermatozoa. This suggests that the lower concentrations of spermatozoon DHA in these individuals are due not to dietary differences but to some type of metabolic difference in the asthenozoospermic men. PMID- 10529090 TI - Increased diaphragm expression of GLUT4 in control and streptozotocin-diabetic rats by fish oil-supplemented diets. AB - Dietary fat intake influences plasma glucose concentration through modifying glucose uptake and utilization by adipose and skeletal muscle tissues. In this paper, we studied the effects of a low-fat diet on diaphragm GLUT4 expression and fatty acid composition in control and streptozotocin-induced diabetic rats. Control as well as diabetic rats were divided into three different dietary groups each. Either 5% olive oil, 5% sunflower oil, or 5% fish oil was the only fat supplied by the diet. Feeding these low-fat diets for 5 wk induced major changes in fatty acid composition, both in control and in diabetic rats. Arachidonic acid was higher in diabetic olive and sunflower oil-fed rats with respect to fish oil fed, opposite to docosahexaenoic acid which was higher in diabetic fish oil-fed rats with respect to the other two groups. Animals receiving a fish oil diet had the lowest plasma glucose concentration. GLUT4 expression in diaphragm, as indicated by GLUT4 protein and mRNA, is modulated both by diabetes and by diet fatty acid composition. Diabetes induced a decrease in expression in all dietary groups. Plasma glucose levels correlated well with the increased amount of GLUT4 protein and mRNA found in fish oil-fed groups. Results are discussed in terms of the influence that arachidonic and n-3 polyunsaturated fatty acids may exert on the transcriptional and translational control of the GLUT4 gene. PMID- 10529091 TI - Apolipoprotein B mRNA editing and apolipoprotein gene expression in the liver of hyperinsulinemic fatty Zucker rats: relationship to very low density lipoprotein composition. AB - We previously demonstrated increased apolipoprotein B (apoB) mRNA editing, elevated levels of mRNA for the catalytic component of the apoB mRNA editing complex, apobec-1, and increased secretion of the product of the edited mRNA, apoB48, in very low density lipoproteins (VLDL) in primary cultures of Sprague Dawley rat hepatocytes following insulin treatment. In order to determine the effect of in vivo hyperinsulinemia on these processes, we determined apoB mRNA editing, apobec-1 expression, hepatic expression of mRNA for apoB and other VLDL apoproteins, and the quantity and composition of plasma VLDL in the hyperinsulinemic fatty Zucker rat. Total apoB mRNA content of the livers of the fatty rats and lean littermates did not differ; however, edited apoB message coding for hepatic apo B48, and abundance of mRNA for the catalytic subunit of the apoB mRNA editing complex, apobec-1, was increased by 1.7- and 3.3-fold, respectively, in fatty rats. ApoCIII mRNA abundance was increased in livers of fatty rats as well, but the abundance of hepatic apoE mRNA in the fatty animal was not different from that of the lean rat. Hepatic apoAI mRNA abundance was also increased in the fatty rats. Associated with increased apoB mRNA editing, was the 1.7-fold increase in the fraction of apoB in plasma as apoB48 in fatty rats. VLDL-triglyceride and -apoB in plasma were 15- and 3-fold higher, respectively, in fatty Zucker rats compared to lean littermates, indicating both enrichment of VLDL with triglycerides and increased accumulation of VLDL particles. Increased hepatic expression of mRNA for apoCIII and apoAI was associated with increased content of apoC (and relative depletion of apoE) in VLDL of fatty rats, and plasma apoAI was increased in fatty Zucker rats, primarily in the HDL fraction. The current study provides further evidence that chronic exposure to high levels of insulin influences both the quantity of and lipid/apoprotein composition of VLDL in plasma. The increased apoC and decreased apoE (as well as increased triglyceride) content of VLDL in the fatty Zucker rat observed in the current study may affect VLDL clearance and therefore may be a factor in the observed accumulation of VLDL in the plasma of the fatty hyperinsulinemic Zucker rats. PMID- 10529092 TI - Effects of cholestyramine on hepatic cholesterol 7alpha-hydroxylase and serum 7alpha-hydroxycholesterol in the hamster. AB - Cholestyramine increases activities of hepatic cholesterol 7alpha-hydroxylase and serum levels of 7alpha-hydroxycholesterol. To examine if serum 7alpha hydroxycholesterol levels parallel with enzyme activity, 0, 0.5, 1, 2, 4, and 10% of cholestyramine was administered to female golden Syrian hamsters for 28 d in the dose-dependent study, and 2% cholestyramine for 0, 1, 3, 7, 14, 21, and 28 d in the time-dependent study. In the dose-dependent study, hepatic and serum cholesterol levels were significantly decreased dose-dependently when more than 0.5% of cholestyramine was fed for 28 d. Cholestyramine increased the cholesterol 7alpha-hydroxylase activity in a dose-dependent manner, while the serum 7alpha hydroxycholesterol level was essentially unchanged. No correlation was found between the serum level and the hepatic enzyme activity. In the time-dependent study, hepatic and serum cholesterol levels markedly decreased when 2% cholestyramine was fed for longer than 3 d. The serum triglyceride level increased significantly for the first 7 d and then decreased. Cholesterol 7alpha hydroxylase activity increased significantly as early as day 1, reached maximum activity level on day 7, and then kept the significantly high values until day 28. The serum 7alpha-hydroxycholesterol level significantly increased for the first 7 d and decreased to the pretreatment level thereafter. 7Alpha hydroxycholesterol levels significantly correlated with serum cholesterol and triglyceride levels. We conclude that the serum 7alpha-hydroxycholesterol level does not always reflect the activity of hepatic cholesterol 7alpha-hydroxylase, when cholesterol metabolism is severely disturbed by cholestyramine. PMID- 10529093 TI - Incorporation of [1-13C]oleate into cellular triglycerides in differentiating 3T3L1 cells. AB - Oleate is one of the most abundant dietary fatty acids, and much remains to be learned about its metabolism in fat cells. We studied the incorporation of exogenous [1-13C]oleate into triglycerides (TG) in differentiating 3T3L1 preadipocytes using 13C NMR spectroscopy. The quantity of oleate incorporated into TG was found to increase as preadipocytes differentiated into fat cells. The ratio of unesterified [1-13C]oleate to total stored fatty acids was higher in less differentiated cells, and declined at later stages of differentiation as cells accumulated fatty acids through de novo synthesis. When added as the only exogenous fatty acid, oleate was largely esterified at the sn-2 position. When equimolar unlabeled linoleate was co-provided at the same time, the ratio of [1 13C]oleate esterified at the sn-1,3 position increased, implying competition between linoleate and oleate for esterification, especially at the sn-2 position. When cells pre-enriched with [1-13C]oleate (esterified to TG) were treated with isoproterenol, a lipolytic agent, most of the [1-13C]oleate was still found in TG, despite a high rate of lipolysis determined by measuring glycerol release. This implies extensive re-esterification of the oleate released by lipolysis. PMID- 10529094 TI - The effect of conjugated linoleic acid on the antioxidant enzyme defense system in rat hepatocytes. AB - Short-term effects of physiological concentrations of conjugated linoleic acid (CLA) on membrane integrity, metabolic function, cellular lipid composition, lipid peroxidation, and antioxidant enzymes were examined using rat hepatocyte suspension cultures. Incubation with CLA (5-20 ppm) for 3 h decreased the ability of hepatocyte plasma membranes to exclude trypan blue by approximately 25%, and caused leakage of cytosolic lactate dehydrogenase (LDH) into the medium. The significant decrease (P< 0.02) in hepatocyte viability as measured by LDH leakage during cell incubation with 10 and 20 ppm CLA was not associated with significant changes in cellular ATP content. Protein synthesis in hepatocytes was elevated (P < 0.05) in the presence of 5 and 10 ppm CLA, but at a higher concentration (20 ppm), protein synthesis was similar to that of control cells. Gluconeogenesis was maintained in cells incubated with lower concentrations of CLA (5 and 10 ppm) but was decreased (P < 0.02) at the higher concentration. Incubation with 20 ppm CLA for 3 h did not affect the specific activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase, the rate-limiting enzyme of cholesterol synthesis. Both cis 9,trans-11/trans-9,cis-11, and cis-10,trans-12/trans-10,cis-12 isomers of CLA were incorporated to a similar level into hepatocytes. Levels ranged from 3.9 to 4.1%, respectively, of total fatty acids in neutral lipids, and from 0.7 to 0.8%, respectively, of total fatty acids in phospholipids. Cellular lipid peroxidation remained unchanged in the presence of CLA (5-20 ppm), despite significant inhibition (P < 0.05) of superoxide dismutase. Catalase activity was maintained near control levels in the presence of 5 and 10 ppm CLA but was significantly decreased in the presence of 20 ppm CLA. Glutathione peroxidase activity was significantly decreased in the presence of 10 ppm CLA. The apparent sensitivity of the antioxidant enzyme defense system of liver cells to CLA, coupled with the lack of effect of CLA on lipid peroxidation in cells, suggests that cytotoxic effects of CLA as described by LDH leakage and decreased gluconeogenesis were not mediated by a prooxidant action in hepatocytes. PMID- 10529095 TI - Characterization of the ybdT gene product of Bacillus subtilis: novel fatty acid beta-hydroxylating cytochrome P450. AB - We have characterized the gene encoding fatty acid alpha-hydroxylase, a cytochrome P450 (P450) enzyme, from Sphingomonas paucimobilis. A database homology search indicated that the deduced amino acid sequence of this gene product was 44% identical to that of the ybdT gene product that is a 48 kDa protein of unknown function from Bacillus subtilis. In this study, we cloned the ybdT gene and characterized this gene product using a recombinant enzyme to clarify function of the ybdT gene product. The carbon monoxide difference spectrum of the recombinant enzyme showed the characteristic one of P450. In the presence of H2O2, the recombinant ybdT gene product hydroxylated myristic acid to produce beta-hydroxymyristic acid and alpha-hydroxymyristic acid which were determined by high-performance liquid chromatography (HPLC) and gas chromatography-mass spectrometry. The amount of these products increased with increasing reaction period and amount of H2O2 in the reaction mixture. The amount of beta-hydroxyl product was slightly higher than that of alpha-hydroxyl product at all times during the reaction. However, no reaction products were detected at any time or at any concentration of H2O2 when heat-inactivated enzyme was used. HPLC analysis with a chiral column showed that the beta-hydroxyl product was nearly enantiomerically pure R-form. These results suggest that this P450 enzyme is involved in a novel biosynthesis of beta-hydroxy fatty acid. PMID- 10529096 TI - Identification of acylated glycoglycerolipids from a cyanobacterium, Synechocystis sp., by tandem mass spectrometry. AB - Acylated glycoglycerolipids were identified in the total lipid extract from cyanobacterium Synechocystis sp. PCC 6803. These compounds have a palmitoyl group esterified to the hydroxyl group at the C-6 position of the terminal glycosyl moiety of digalactosyl monoacylglycerol and digalactosyl diacylglycerol. Their structural elucidation was accomplished by tandem mass spectrometry coupled with fast atom bombardment ionization. Acylated digalactosyl monoacylglycerol has a structure of 1-hydroxy-2-palmitoyl-3-O-[(6-O-palmitoyl)-alpha-D-galactopyranosyl (1-->6)-beta-D-galactopyranosyl]-sn-glycerol. This compound has not been reported previously. PMID- 10529097 TI - Delta5-olefinic acids in the seed lipids from four Ephedra species and their distribution between the alpha and beta positions of triacylglycerols. Characteristics common to coniferophytes and cycadophytes. AB - The fatty acid compositions of the seed lipids from four Ephedra species, E. nevadensis, E. viridis, E. przewalskii, and E. gerardiana (four gymnosperm species belonging to the Cycadophytes), have been established with an emphasis on delta5-unsaturated polymethylene-interrupted fatty acids (delta5-UPIFA). Mass spectrometry of the picolinyl ester derivatives allowed characterization of 5,9- and 5,11-18:2; 5,9,12-18:3; 5,9,12,15-18:4; 5,11-20:2; 5,11,14-20:3; and 5,11,14,17-20:4 acids. Delta5-UPIFA with a delta11-ethylenic bond (mostly C20 acids) were in higher proportions than delta5-UPIFA with a delta9 double bond (exclusively C18 acids) in all species. The total delta5-UPIFA content was 17-31% of the total fatty acids, with 5,11,14-20:3 and 5,11,14,17-20:4 acids being the principal delta5-UPIFA isomers. The relatively high level of cis-vaccenic (11 18:1) acid found in Ephedra spp. seeds, the presence of its delta5-desaturation product, 5,11-18:2 acid (proposed trivial name: ephedrenic acid), and of its elongation product, 13-20:1 acid, were previously shown to occur in a single other species, Ginkgo biloba, among the approximately 170 gymnosperm species analyzed so far. Consequently, Ephedraceae and Coniferophytes (including Ginkgoatae), which have evolved separately since the Devonian period (approximately 300 million yr ago), have kept in common the ability to synthesize C18 and C20 delta5-UPIFA. We postulate the existence of two delta5-desaturases in gymnosperm seeds, one possibly specific for unsaturated acids with a delta9 ethylenic bond, and the other possibly specific for unsaturated acids with a delta11-ethylenic bond. Alternatively, the delta5-desaturases might be specific for the chain length with C18 unsaturated acids on the one hand and C20 unsaturated acids on the other hand. The resulting hypothetical pathways for the biosynthesis of delta5-UPIFA in gymnosperm seeds are only distinguished by the position of 11-18:1 acid. Moreover, 13C nuclear magnetic resonance spectroscopy of the seed oil from two Ephedra species has shown that delta5-UPIFA are essentially excluded from the internal position of triacylglycerols, a characteristic common to all of the Coniferophytes analyzed so far (more than 30 species), with the possibility of an exclusive esterification at the sn-3 position. This structural feature would also date back to the Devonian period, but might have been lost in those rare angiosperm species containing delta5 UPIFA. PMID- 10529098 TI - Positional analysis of triglycerides and phospholipids rich in long-chain polyunsaturated fatty acids. AB - Four sources of long-chain polyunsaturated fatty acids (LCP) differing in their chemical structure (triglycerides or phospholipids) and in their origin (tuna triglycerides, fungal triglycerides, egg phospholipids, and pig brain phospholipids) were analyzed to determine the distribution of the component fatty acids within the molecule. Lipase and phospholipase A2 hydrolysis was performed to obtain 2-monoacylglycerols and lysophospholipids, respectively, which allowed us to determine the distribution of fatty acids between the sn-2 and sn-1,3 positions of triglycerides or between the sn-1 and sn-2 position of phospholipids. Fatty acids in the LCP sources analyzed were not randomly distributed. In tuna triglycerides, half of the total amount of 22:6n-3 was located at the sn-2 position (49.52%). In fungal triglycerides, 16:0 and 18:0 were esterified to the sn-1,3 (92.22% and 91.91%, respectively)18:1 and 18:2 to the sn-2 position (59.77% and 62.62%, respectively), and 45% of 20:3n-6 and only 21.64% of 20:4n-6 were found at the sn-2 position. In the lipid sources containing phospholipids, LCP were mainly esterified to the phosphatidylethanolamine fraction. In egg phospholipids, most of 20:4n-6 (5.50%, sn-2 vs. 0.91%, sn-1) and 22:6n-3 (2.89 vs. 0.28%) were located at the sn-2 position. In pig brain phospholipids, 22:6n-3 was also esterified to the sn-2 (13.20 vs. 0.27%), whereas 20:4n-6 was distributed between the two positions (12.35 vs. 5.86%). These results show a different fatty acid composition and distribution of dietary LCP sources, which may affect the absorption, distribution, and tissue uptake of LCP, and should be taken into account when supplementing infant formulas. PMID- 10529099 TI - Preparation, separation, and confirmation of the eight geometrical cis/trans conjugated linoleic acid isomers 8,10- through 11,13-18:2. AB - Conjugated linoleic acid (CLA) mixtures were isomerized with p-toluenesulfinic acid or I2 catalyst. The resultant mixtures of the eight cis/trans geometric isomers of 8,10-, 9,11-, 10,12-, and 11,13-octadecadienoic (18:2) acid methyl esters were separated by silver ion-high-performance liquid chromatography (Ag+ HPLC) and gas chromatography (GC). Ag+-HPLC allowed the separation of all positional CLA isomers and geometric cis/trans CLA isomers except 10,12-18:2. However, one of the 8,10 isomers (8cis, 10trans-18:2) coeluted with the 9trans,11cis-18:2 isomer. There were differences in the elution order of the pairs of geometric CLA isomers resolved by Ag+-HPLC. For the 8,10 and 9,11 CLA isomers, cis,trans eluted before trans,cis, whereas the opposite elution pattern was observed for the 11,13-18:2 geometric isomers (trans,cis before cis,trans). All eight cis/trans CLA isomers were separated by GC on long polar capillary columns only when their relative concentrations were about equal. Large differences in the relative concentration of the CLA isomers found in natural products obscured the resolution and identification of a number of minor CLA isomers. In such cases, GC-mass spectrometry of the dimethyloxazoline derivatives was used to identify and confirm coeluting CLA isomers. For the same positional isomer, the cis,trans consistently eluted before the trans,cis CLA isomers by GC. High resolution mass spectrometry (MS) selected ion recording (SIR) of the molecular ions of the 18:1, 18:2, and 18:3 fatty acid methyl esters served as an independent and highly sensitive method to confirm CLA methyl ester peak assignments in GC chromatograms obtained from food samples by flame-ionization detection. The high-resolution MS data were used to correct for the nonselectivity of the flame-ionization detector. PMID- 10529100 TI - Synthesis of 9Z,11E-octadecadienoic and 10E,12Z-octadecadienoic acids, the major components of conjugated linoleic acid. AB - Linoleic acid was efficiently converted into the two major components of conjugated linoleic acid, 9Z,11E-octadecadienoic (1a) and 10E,12Z-octadecadienoic acid (1b) using either the superbase (n-butyllithium/potassium tert-butoxide) or by simply refluxing with KOH in 1-butanol. In turn, 1a and 1b were separated from each other using the lipase from Aspergillus niger via stereoselective esterification in 1-butanol. This enzyme has a preference for the 9Z,11E isomer, 1a, and has excellent selectivity. This method has allowed the ready preparation of gram quantities of 1a and 1b in their highly purified forms, which are not readily accessible by current methods. PMID- 10529101 TI - Paramyxovirus reverse genetics is coming of age. PMID- 10529103 TI - Ribotyping of Vibrio parahaemolyticus isolates obtained from food poisoning outbreaks in Taiwan. AB - Vibrio parahaemolyticus is a prevalent food-borne pathogen in Taiwan, Japan and other Asian countries. This work presents a novel ribotyping method for the molecular epidemiological examination of this pathogen. Genomic DNA was fragmented by HindIII digestion and hybridized with cDNA probe for Escherichia coli 16S and 23S RNA genes. A total of 121 isolates obtained from outbreaks during 1992 and 1994 in Taiwan were characterized by this ribotyping method. Four to seventeen restricted fragments were visualized in these isolates. After hierarchical cluster analysis, these isolates were grouped into thirty different ribotypes. In addition, A3, A7, E3 and F1 were the major ribotypes, consisting of 22.3, 13.2, 9.1, and 8.3% of the isolates, respectively. A, E, F, G and B were the major groups, consisting of 46.2, 14.0, 9.1, 6.7, and 6.7% of the isolates, respectively. The discriminatory ability of this ribotyping method, as determined by Simpson's index of diversity, was 0.93, which closely resembled that of a previously reported pulsed-field gel electrophoresis method. PMID- 10529104 TI - The production of nitric oxide and tumor necrosis factor by murine macrophages infected with mycobacterial strains differing by hemolytic activity. AB - In this study, we compared the secretion of nitric oxide (NO) and tumor necrosis factor (TNF-alpha) by murine macrophages infected in vitro with hemolytic or unhemolytic mycobacteria isolates. We observed that unhemolytic mycobacteria induced more intensive NO production by macrophages and were more susceptible to bactericidal effect of mononuclear phagocytes than hemolytic mycobacterial strains. In contrast, the high-virulence hemolytic isolates induced significantly stronger TNF-alpha production by infected macrophages than the low-virulence unhemolytic bacilli. PMID- 10529102 TI - Differences in genomic macrorestriction patterns of arabinose-positive (Burkholderia thailandensis) and arabinose-negative Burkholderia pseudomallei. AB - We reported previously two biochemically and antigenically distinct biotypes of Burkholderia pseudomallei. These two distinct biotypes could be distinguished by their ability to assimilate L-arabinose. Some B. pseudomallei isolated from soil samples could utilize this substrate (Ara+), whereas the other soil isolates and all clinical isolates could not (Ara-). Only the Ara isolates were virulent in animals and reacted with monoclonal antibody directed at the surface envelope, most likely the exopolysaccharide component. In the present study, pulsed-field gel electrophoresis was employed for karyotyping of these previously identified B. pseudomallei strains. We demonstrate here that the DNA macrorestriction pattern allows the differentiation between B. pseudomallei, which can assimilate L-arabinose, and the proposed B. thailandensis, which cannot do so. Bacterial strains from 80 melioidosis patients and 33 soil samples were examined by genomic DNA digestion with NcoI. Two major reproducible restriction patterns were observed. All clinical (Ara-) isolates and 9 Ara- soil isolates exhibited macrorestriction pattern I (MPI), while 24 soil isolates (Ara+) from central and northeastern Thailand displayed macrorestriction pattern II (MPII). The study here demonstrated pulsed-field gel electrophoresis to be a useful tool in epidemiological investigation possibly distinguishing virulent B. pseudomallei from avirulent B. thailandensis or even identifying closely related species of Burkholderia. PMID- 10529105 TI - The expression of the pathogenic yeast Candida albicans catalase gene in response to hydrogen peroxide. AB - The catalase gene of the pathogenic yeast Candida albicans was cloned and its expression was examined. Activity of the catalase was detected when cells which were in the early logarithmic stage were treated with hydrogen peroxide. Additionally, activity was detected without any treatment to cells in the late logarithmic and stationary phases. When cells were cultured in galactose, glycerol, or ethanol, catalase activity was always observed without the hydrogen peroxide treatment, suggesting that glucose represses the induction of catalase expression. To elucidate the molecular mechanism of catalase expression, the putative gene for catalase and its 5' untranscribed region were cloned. Sequences of the gene and its potential regulatory region revealed several motifs, including a GC box-like element and stress-responsive element (STRE), which could be involved in the transcriptional regulation. Northern analysis showed that hydrogen peroxide and sorbitol activated transcription of the catalase. On the other hand, treatment of glucose strictly repressed the expression of the catalase even when co-treated with hydrogen peroxide. The expression of catalase against treatment with hydrogen peroxide took place very quickly and decreased slowly in the experimental condition adopted here. From these results, we assumed that the expression of the catalase in Candida albicans is regulated by various environmental conditions via motifs for transcriptional activation as in other yeast catalases. PMID- 10529106 TI - Apoptosis observed in BALB/3T3 cells having ingested Staphylococcus aureus. AB - Staphylococcus aureus was previously shown to be internalized by murine fibroblast. We examined the intracellular events of S. aureus ingested by BALB/3T3 cells. After uptake of strains A191 and A151, isolates from atopic lesion, and a laboratory strain, Cowan I, for 1 hr, BALB/3T3 cells were incubated with 1.25 microg/ml lysostaphin. Laddering of the DNA in multiples of approximately 180 bp occurred within 4 hr following bacterial addition in BALB/3T3 cells infected with A191 and within 18 hr in BALB/3T3 cells infected with A151: histochemical staining by the terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling method revealed that the rate of the fragmentation of nucleic DNA in Cowan I-infected BALB/3T3 cells at 21 hr following bacterial addition was 0.52 +/- 0.25%, significantly higher than that in the control cells. Transmission electron micrographs of BALB/3T3 cells at 4 hr following A191 addition showed that the apoptotic features, including electron dense nucleus and plasma membrane blebbing, occurred in some cells in which many staphylococci escaped the endosome and went on to cell division. At the same time, A151 organisms enclosed with endosome membrane were static in the intact BALB/3T3 cells. The significant increase of A191 was confirmed by counting intracellular live bacteria during 2- to 6-hr incubation. These results suggest that internalized S. aureus escapes the endosome, multiplies and induces apoptosis in the fibroblast cell. PMID- 10529107 TI - Development of an enzyme-labeled oligonucleotide probe for detecting the Escherichia coli attaching and effacing A gene. AB - Enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic Escherichia coli (EHEC) can produce attaching and effacing (AE) lesions on intestinal epithelium in vitro and in vivo. A gene necessary to cause the AE lesion has been identified and designated Escherichia coli attaching and effacing A (eaeA) gene. In this study, an alkaline phosphatase (ALP)-conjugated oligonucleotide probe for the eaeA gene was developed and used to detect the eaeA gene among 163 strains of classical EPEC and 25 strains of EHEC O157. The prevalence rates of eaeA gene in the strains of classical EPEC and EHEC O157 were 51.5 and 100%, respectively. The eaeA-positive rate (60.0%) in strains of class I EPEC serogroups (O26, O55, O86, O111, O119, O125, O126, O127, O128ab, and O142) was significantly higher than that (22.9%) in strains of the class II EPEC serogroups (O18, O44, O114) (P<0.01). A total of 109 eaeA-positive classical EPEC and EHEC O157 were positive for fluorescent actin staining (FAS) assay, whereas 79 eaeA-negative classical EPEC were negative. Both the sensitivity and specificity of the eaeA probe versus the FAS assay positivity were 100%. Thus, use of the ALP-conjugated oligonucleotide probe for the eaeA gene would be specific and reliable in identifying the adherence capability of EPEC and EHEC. PMID- 10529108 TI - Repetitive sequence of Leptospira interrogans serovar icterohaemorrhagiae strain Ictero No. 1: a sensitive probe for demonstration of Leptospira interrogans strains. AB - A 4.8-kilobase (kb) repetitive sequence element generated with KpnI digestion was cloned from the Leptospira interrogans serovar icterohaemorrhagiae strain Ictero No. 1. The sequence, repeated in tandem, was located on the 280-kb fragment between the FseI and AscI sites on the chromosome by hybridization using the 4.8 kb fragment as a probe. We cloned the fragment containing the element for the Ictero No. 1 strain in a lambda EMBL3 bacteriophage DNA, and one out of 5 clones was sequenced. Within the sequenced 9-kb segment that partially repeated, 9 putative open-reading frames and 2 transfer RNA genes, for alanine and isoleucine, were identified. A similarity search for the products deduced from the sequenced data revealed that the repeated sequence includes both beta oxidation enzymes, acyl-CoA dehydrogenase and enoyl-CoA hydratase, and hydroxythiazole kinase protein homologues. Hybridization experiments against different leptospiral strains using the element as a probe showed a similar sequence in the strains of L. interrogans and L. kirschneri, but not in any strains of L. borgpetersenii, L. weillii, L. meyeri or L. biflexa. Results indicated that the highly repeated element in the Ictero No. 1 strain exists as a well conserved sequence, though at a moderate level of repetition, in certain strains of L. interrogans and L. kirschneri. PCR amplification targeting the repetitive element was successful and indicated that the procedure provides a sensitive and specific probe to detect leptospires. PMID- 10529109 TI - Borna disease virus infection in two family clusters of patients with chronic fatigue syndrome. AB - A high rate of Borna disease virus (BDV) infection has been demonstrated in patients with chronic fatigue syndrome (CFS). Herein, we focused on BDV infection in two family clusters of patients with CFS: a father, mother, two sons and one daughter (family #1); and a father, mother, two daughters and one son (family #2). All members, except for the elder son in family #1 and the father and son in family #2, were diagnosed with CFS. The results supported that all the family members with CFS were infected with BDV, as evidenced by the presence of antibodies to viral p40, p24 and/or gp18 and BDV p24 RNA in peripheral blood mononuclear cells. The healthy members, except for the father of family #2 who was positive for antibody to p24, were all negative by both assays. Follow-up studies in family #1 continued to reveal BDV antibodies and BDV RNA, except in the mother, who lost the RNA upon slight recovery from the disease. PMID- 10529110 TI - Arginine carboxypeptidase (CPR) in human plasma determined with sandwich ELISA. AB - There are two types of carboxypeptidases present in human blood, carboxypeptidase N (CPN) and arginine carboxypeptidase (CPR). CPR is generated during coagulation from a precursor (proCPR) which can be converted to the active form by trypsin in vitro. Since it is difficult to distinguish the two types of carboxypeptidases in human blood by the measurement of enzyme activity, we established a quantitative sandwich ELISA by which CPR can be quantitated. The amount of CPR in plasma, fresh serum and heated serum were essentially the same. Therefore the ELISA assay does not distinguish proCPR, activated CPR and inactivated CPR. With the ELISA method, CPR was quantitated in plasma from fifty patients with rheumatoid arthritis and eleven patients with severe hepatitis as well as healthy individuals. The amount of CPR in plasma obtained from patients with rheumatoid arthritis was not found to be lower than that of normal subjects. Furthermore, the patients who suffered severe hepatitis and had very low levels of CPR-total were fatal. This suggests that a decrease of CPR level might be a good indication of a patient's prognosis to death by hepatitis. PMID- 10529111 TI - The characterization of Shiga toxin-non-producing Escherichia coli serotype O157:H7 isolated from carcasses of cattle at a slaughter house. AB - A bacteriological investigation of Shiga toxin (Stx)-producing Escherichia coli (STEC) O157:H7 was performed on 298 carcasses of cattle at slaughter houses between July 1996 and January 1997 in Gifu Prefecture, Japan. As a result, four Stx-non-producing Escherichia coli O157:H7 strains were isolated from two slaughtered carcasses of cattle. The purpose of this study was to examine the characterization of isolates. Isolates possessed the E. coli attaching and effacing gene (eaeA), and hemolysin gene (hlyA), and harbored 3.0-MDa and 60-MDa plasmids. The Xba I pulsed-field gel electrophoresis (PFGE) pattern showed three similar patterns. Consequently, a closely related genotype of Stx-non-producing E. coli O157:H7 may widely exist in cattle. PMID- 10529112 TI - Salt-Sensitive growth of Staphylococcus aureus: stimulation of salt-induced autolysis by multiple environmental factors. AB - The growth of Staphylococcus aureus 209P became extremely sensitive to a high NaCl concentration following lowered temperature, reduced air-supply, and decreased Ca2+ concentration in the medium. Cells in high-NaCl and low-Ca2+ concentration media either autolyzed or transformed into protoplast-like forms during growth when grown standing below 37 degrees C. The abnormal growth, however, was invariably avoided by preliminary supplementation with polyanetholesulfonate (autolysin inhibitor) in the growth media. These results suggested that the autolytic activity of this organism was precisely controlled by multiple environmental factors such as ionic strength, temperature, air supply, and the concentration of Ca2+. PMID- 10529113 TI - Phylogenetic analysis of saccharolytic oral treponemes isolated from human subgingival plaque. AB - A total of 74 strains of oral treponemes, which were isolated from subgingival plaque samples from patients with periodontitis, were taxonomically studied on the basis of biochemical characteristics, DNA-DNA hybridization, and 16S rRNA gene sequences. These organisms fermented carbohydrates and required rumen fluid or short-chain volatile fatty acids for growth. The isolates were divided into seven subgroups based on their biochemical characteristics. The levels of DNA relatedness among the representative strains of each subgroup and Treponema socranskii (including three subspecies) were greater than 78%, while the levels of DNA relatedness among these strains and other Treponema species, including T. denticola and "T. vincentii", were less than 15%. DNA-DNA hybridization indicated that all subgroups belonged to T. socranskii. This result correlated well with the cluster on the phylogenetic trees based on 16S rRNA sequences. PMID- 10529114 TI - Phenotypic and genotypic homogeneity of the strains of Rickettsia japonica isolated from patients with Oriental spotted fever. AB - Nine pathogenic strains of Rickettsia japonica isolated from patients with Oriental spotted fever were compared phenotypically and genotypically. Constitution and antigenicity of the proteins demonstrated to be the same among strains. Polymerase chain reaction (PCR) amplification of the two major outer membrane protein genes (ompA and ompB) and an intracellular spotted fever group common antigen protein gene (rps120) produced the same sizes of products for all strains. Restriction fragment length polymorphism of the PCR products showed the same pattern among strains with each endonuclease. Thus, these strains belong to a single type, the same as the type strain YH (=ATCC VR-1363). PMID- 10529115 TI - Human IgM monoclonal antibody to ganglioside GM2 and complement suppress virus propagation in ex vivo cultures of lymphocytes from HIV-1 infected patients. AB - HIV-1 infection induces aberrant ganglioside GM2 expression on infected cell lines, and human IgM anti-GM2 monoclonal antibody (L55 Ab) together with normal fresh human serum (FHS) as a source of complement causes complement mediated cytolysis of HIV-1 infected cells as well as HIV-1 particles. We report here that high expression of GM2 was also detected on HIV-1 infected lymphocytes from HIV-1 seropositive patients. L55 Ab effectively suppressed the generation of HIV in the presence of FHS in primarily cultured lymphocytes from HIV-1 infected patients in ex vivo experiments, and the suppression was enhanced additively by AZT. These data suggest that L55 Ab may increase the therapeutic effect of chemotherapy. PMID- 10529116 TI - Human immunodeficiency virus associated spondyloarthropathy: lessons from the Third World. PMID- 10529117 TI - Is HIV the culprit? Spondyloarthropathy in the Third World. PMID- 10529118 TI - OMERACT: economic evaluations and health policy. PMID- 10529119 TI - Statistical analysis of cost effectiveness data. PMID- 10529120 TI - Seventy-five years of microscopic polyangiitis--what have we learned? PMID- 10529121 TI - Nitric oxide synthase is expressed in the lymphomononuclear cells of synovial fluid in patients with rheumatoid arthritis. AB - OBJECTIVE: To investigate the expression of inducible nitric oxide synthase (iNOS) in subpopulations of peripheral blood and synovial fluid (SF) leukocytes in patients with rheumatoid arthritis (RA). METHODS: iNOS was detected in peripheral blood and SF samples after cell permeabilization, by 2 color immunofluorescence flow cytometry. Samples from 14 patients with RA and 8 with osteoarthritis (OA) were studied. Nitrite concentration was determined by Griess reaction, interleukin 1beta and tumor necrosis factor alpha by an immunoenzymatic assay, and C-reactive protein (CRP) by an immunonephelometric method. RESULTS: In SF, iNOS was detected in 11 of 14 patients with RA and 2 of 8 with OA. In blood cells, iNOS was detected in 8 of 14 patients with RA and none of the OA group. iNOS was consistently detected in monocytes and was not detected in granular cells. In RA, there was no correlation between the number of iNOS positive mononuclear cells and cytokine concentrations. CRP concentration was correlated with the number of iNOS positive mononuclear cells in RA SF samples. CONCLUSION: SF mononuclear cells from patients with RA express iNOS and are involved in NO production in the joint. The number of positive cells is correlated with CRP concentration, suggesting the implication of NO production in the inflammatory process. PMID- 10529122 TI - Analysis of serial synovial biopsies in patients with rheumatoid arthritis: description of a control group without clinical improvement after treatment with interleukin 10 or placebo. AB - OBJECTIVE: Analysis of serial synovial biopsy specimens is increasingly used as an outcome measure for the evaluation of therapeutic interventions. However, observations in placebo treated groups are scarce. We describe the immunohistologic features of the synovium in placebo treated patients with RA and in those who received interleukin 10 (IL-10). METHODS: Ten patients with active RA received dosages of either placebo (n = 7) or 5 microg/kg (n = 1) or 10 microg/kg (n = 2) of recombinant human IL-10 (rhIL-10; SCH 52000, Schering Plough, Kenilworth, NJ, USA) daily for 28 consecutive days. Synovial biopsy specimens from the knee joint were obtained by needle arthroscopy before and 4 weeks after initiation of treatment. Immunohistochemistry was performed using monoclonal antibodies specific for the following surface markers and cytokines: CD3, CD4, CD8, CD38, CD68, CD55, IL-1beta, IL-6, and tumor necrosis factor-alpha. RESULTS: No patient exhibited clinical improvement after treatment with placebo or any rhIL-10 dosage. Microscopic analysis of synovial tissue revealed no significant change in the scores for infiltration by inflammatory cells or in the scores for the expression of cytokines after treatment. CONCLUSION: Studies of serial synovial biopsies from patients treated with placebo or IL-10 revealed no changes in immunohistologic scores. This suggests that the biopsy procedure itself has no effect on the features of the synovium. PMID- 10529123 TI - Synovium infiltrating T cells induce excessive synovial cell function through CD28/B7 pathway in patients with rheumatoid arthritis. AB - OBJECTIVE: To clarify involvement of synovial T cells in the development of synovial inflammation in patients with rheumatoid arthritis (RA), we analyzed cellular interactions between synovial cells and infiltrating T cells via CD28/B7 1 and B7-2. METHODS: Synovial cells and infiltrating T cells were recovered separately from RA synovial tissues. Expression of CD28, B7-1, and B7-2 of synovial cells was analyzed by immunohistochemical staining and immunofluorescence analysis. Interleukin 1beta (IL-1beta), IL-6, and matrix metalloprotease 3 (MMP-3) secreted by synovial cells in the presence of infiltrating T cells were measured by ELISA. Nuclear transcription factor CD28 responsive complex was detected by a gel shift assay. RESULTS: Both CD28+ T cells and B7-1/B7-2+ cells were found accumulating in the mononuclear cell infiltrate of RA synovial tissues and B7-1/B7-2+ cells were mainly LeuM3+ synovial cells. CD28 responsive complex was detected in nuclear extracts of freshly isolated lymphocytes from RA synovial tissues, but not those from osteoarthritis synovial tissues or normal peripheral blood, suggesting in vivo activation of T cells by the CD28/B7-1/B7-2 interactions. The irradiated autologous synovium infiltrating T cells notably enhanced IL-1beta, IL-6, and MMP-3 production by the synovial cells. The enhancement of proinflammatory cytokine and MMP-3 production by the synovial cells co-cultured with the T cells was abolished by the addition of CTLA4-Ig, anti-B7-1, and anti-B7-2 monoclonal antibodies. CONCLUSION: These results suggest that cellular interactions between synovium infiltrating T lymphocytes and synovial cells via B7/CD28 pathways are intimately associated with development and exacerbation of inflammation in RA synovial cells. PMID- 10529124 TI - Cyclosporine A in rheumatoid arthritis: randomized, placebo controlled dose finding study. AB - OBJECTIVE: To determine the lowest effective starting dose and residual benefit of cyclosporine A (CSA) in patients with rheumatoid arthritis (RA) refractory to other agents. METHODS: In a double blind (masked observer), controlled, multicenter study, patients with RA started CSA 0 (placebo; n = 61), 1.5 (n = 89), or 2.5 (n = 94) mg/kg/day in a 21 week study that permitted dose escalation after 8 weeks, 1 week tapering of dose at 16 weeks, and post-therapy observation for 4 weeks. RESULTS: Patients with RA taking CSA 2.5 mg/kg/day fared better than those in the placebo or CSA 1.5 mg/kg/day groups in Patient Global Assessment, Examiner Global Assessment, Pain/Tender Joint Count and Index, Swollen Joint Count, and the "Ability at this moment" part of a modified Health Assessment Questionnaire. There was no difference in response between CSA 1.5 mg/kg/day and placebo groups. In the CSA 2.5 mg/kg/day group: improvement occurred between 8 and 12 weeks of therapy; average CSA dose escalation resulted in CSA 2.85 mg/kg/day by Week 16; benefit was not maintained during post-therapy observation and 7 patients discontinued the study because of an adverse event. Adverse events were common in all groups and included gastrointestinal discomfort, hypertension, and increased creatinine. Adverse events remitted with adjustment of dose or after washout in most patients. CONCLUSION: In RA, treatment of patients with CSA 2.5 mg/kg/day, but not 1.5 mg/kg/day, resulted in improvement of 4 of 5 primary efficacy variables when compared to placebo. Adverse events were mostly manageable. CSA was an effective therapy for patients with RA who had failed at least one second line agent. PMID- 10529125 TI - The safety and efficacy of cyclosporine (Neoral) in rheumatoid arthritis. AB - OBJECTIVE: To assess the safety and efficacy of cyclosporine (Neoral), its steroid sparing effect, and its usefulness in combination therapy with methotrexate (MTX) in the treatment of refractory rheumatoid arthritis (RA). METHODS: All patients given cyclosporine for refractory RA over a 21 month period were included in an open, prospective study. Patients were reviewed initially fortnightly then monthly with clinical evaluation and serum creatinine. There were no restrictions on the use of other disease modifying agents, nonsteroidal antiinflammatory drugs, or corticosteroids. RESULTS: Forty-six patients with severe RA were included in the study, 33 (72%) female and 13 (28%) male, with a mean age of 54.8 years (range 20-74). At the completion of the study 30 (65%) were still taking cyclosporine at a mean dose of 2.94 mg/kg/day for a mean duration of 10.5 months. Thirteen patients discontinued cyclosporine due to side effects, most commonly gastrointestinal, and 3 due to inefficacy. Thirty-seven of the 46 patients were taking prednisolone at the start of the study at a mean dose of 10.36 mg/day, which decreased to 7.068 mg/day at the end of the study (p < 0.001). Thirty patients used cyclosporine in combination with MTX. The mean dose of MTX decreased from 15.08 to 13.67 mg/wk (p = 0.02). The mean serum creatinine increased by 13% from 74 to 83.7 micromol/l. Patients who continued therapy had a shorter duration of disease, with a mean of 9.93 years compared to 15.73 years in those who stopped therapy (p = 0.004). CONCLUSION: Cyclosporine (Neoral) was a safe and effective therapy in this population of patients with RA refractory to standard therapy. We observed a significant steroid sparing effect and have shown that combination therapy with MTX does not increase side effects and allows for a decrease in MTX dose. Renal function is not adversely affected if guidelines are followed. PMID- 10529126 TI - Compliance to drug treatment of patients with rheumatoid arthritis: a 3 year longitudinal study. AB - OBJECTIVE: Patient compliance is considered necessary for the success of drug treatment in chronic diseases. We document compliance with drug treatment and the factors affecting it in a cohort of patients with rheumatoid arthritis (RA). METHODS: A prospective cohort study of 556 patients with RA followed for 3 years in 4 counties: Oslo, Norway; Groningen, The Netherlands; and Nancy and Reims, France. Compliance to treatment was assessed annually by interview in terms of adherence to the dose and timing of the prescribed drug regimen. RESULTS: Of the 556 subjects, 429 (77.2%) were taking medication for RA throughout the observation period. Consistent behavior was recorded in 59.5% of cases: 35.7% were consistently compliant, and 23.8% consistently noncompliant. Factors significantly associated with good compliance were older age (p = 0.00), female sex (p = 0.03), decreased disability (p = 0.04), very satisfactory contacts with health care professionals (p = 0.03), and more personal knowledge about the disease and its treatment (p = 0.03). CONCLUSION: This longitudinal study identified compliance behavior as consistent over time in 60% of patients, determined by quality of contact with professionals and the amount of patient information available. PMID- 10529127 TI - Sulfasalazine treatment for rheumatoid arthritis: a metaanalysis of 15 randomized trials. AB - OBJECTIVE: To assess the efficacy and safety of sulfasalazine (SSZ) compared to placebo and other disease modifying drugs. METHODS: A metaanalysis was performed on 15 randomized clinical trials of rheumatoid arthritis (RA) that included SSZ (2 g/day average dose, 36 weeks average followup) as a treatment. Eight trials included a placebo group (PL), 2 hydroxychloroquine (HCQ) (350 mg/day average dose), 3 D-penicillamine (D-Pen) (667 mg/day average dose), and 4 gold sodium thiomalate or aurothioglucose (GST) (25 mg, 1 g/wk). RESULTS: Compared to PL, SSZ was superior for improvement in erythrocyte sedimentation rate (ESR) (SSZ 37%, PL 14%; p < 0.0001), morning stiffness duration (SSZ 61%, PL 33%; p = 0.008), pain visual analog scale (SSZ 42%, PL 15%; p < 0.0001), articular index (SSZ 46%, PL 20%; p < 0.0001), number of swollen joints (SSZ 51%, PL 26%; p < 0.0001), number of painful joints (SSZ 59%, PL 33%; p = 0.004), and patient global assessment (SSZ 26%, PL 14%; p = 0.02). Withdrawals from study because of adverse drug reactions were increased (SSZ 24%, PL 7%; p < 0.0001), but lack of efficacy dropouts were decreased (SSZ 8%, PL 21%; p < 0.0001). Compared to HCQ, SSZ tended to have fewer lack of efficacy dropouts (SSZ 5%, HCQ 15%; p = 0.055) and improved ESR (SSZ 43%, HCQ 26%; p = 0.10) and morning stiffness duration (SSZ 59%, HCQ 40%; p = 0.09). Compared to GST, adverse drug reaction dropouts were significantly fewer (SSZ 12%, GST 29%; p < 0.0001), while withdrawals due to lack of efficacy were greater (SSZ 13%, GST 4%; p = 0.006). More patients tended to complete treatment taking SSZ (SSZ 69%, GST 61%; p = 0.09). CONCLUSION: Over all, the metaanalysis provides data that support the effectiveness of SSZ as a treatment for RA. PMID- 10529128 TI - Classification of an intermediate group of patients with antiphospholipid syndrome and lupus-like disease: primary or secondary antiphospholipid syndrome? AB - OBJECTIVE: (1) To classify an intermediate group of patients (IntAPS) with antiphospholipid syndrome (APS) and lupus-like disease either as primary (PAPS) or secondary APS (SAPS) and to discuss 2 different classifications. (2) To compare patients of a division of rheumatology with either PAPS or SAPS. METHODS: Patients with APS and patients with systemic lupus erythematosus (SLE) followed at the Department of Rheumatology, University Hospital Bichat, Paris, from 1987 to 1996 were analyzed. A chart review and a standardized telephone interview in 1997 completed the data of this study. RESULTS: (1) We found a total of 108 patients with APS: 22 with PAPS, 69 with SAPS, and 17 with IntAPS. The group of IntAPS did not differ from PAPS in any clinical or laboratory signs with the exception of antibodies to dsDNA and to extractable nuclear antigen (ENA). Between IntAPS and SAPS, there were several significant differences in clinical signs of SLE (malar rash, discoid rash, arthralgia) and in laboratory values (leukocytopenia). (2) Comparison of PAPS and SAPS showed statistically significant differences for positive Coombs' test, leukocytopenia, lymphocytopenia, antinuclear antibodies, antibodies to dsDNA and to ENA, and hypocomplementemia. CONCLUSION: The mainstay of the diagnosis of APS is the clinical event of thrombosis or miscarriage in the presence of antiphospholipid antibodies. Less important are laboratory values, which may help to differentiate PAPS from SAPS in order to initiate adequate therapy (e.g., anticoagulation in the first and additional corticosteroids in the second). Patients with IntAPS are more likely to be integrated into the group of PAPS than in the group of SAPS; therefore, special exclusion criteria for PAPS are not appropriate. PMID- 10529129 TI - Natural history of hypercholesterolemia in systemic lupus erythematosus. AB - OBJECTIVE: To determine the natural history of hypercholesterolemia in the first 3 years of disease in an inception cohort of patients with systemic lupus erythematosus (SLE) followed at a single center and to determine the influence of hypercholesterolemia on the subsequent development of coronary artery disease (CAD) related events. METHODS: We identified patients who were seen at the University of Toronto lupus clinic within 1 year of diagnosis from January 1, 1974, to December 31, 1987, and who were seen at least once a year in the first 3 years. Patients were divided into 3 groups: Normal cholesterol: serum total cholesterol (TC) < 5.2 mmol/l throughout the 3 year period of study. Sustained hypercholesterolemia: at least one measurement of TC of > 5.2 mmol/l in each of the first 3 years at the clinic. Variable hypercholesterolemia: TC > 5.2 mmol/l in no more than 2 of the first 3 years of followup. Patients were followed from inception until the present day. The primary outcome was the time of the first CAD related event (myocardial infarction, angina, or sudden unexplained death). RESULTS: One hundred thirty-four patients (118 women, 16 men) were studied: 33 (24.6%) had normal cholesterol, 54 (40.3%) had sustained hypercholesterolemia, and 47 (35.1%) had variable hypercholesterolemia. Using multiple logistic regression the best predictors of sustained hypercholesterolemia were cumulative dose of steroids, no antimalarial therapy, and age of onset of SLE > 35 years old. CAD related events occurred in 1 (3%) of the normal TC group, 3 (6.4%) of the variable group, and in 15 (27.8%) of the sustained group (p = 0.003), 79% of all CAD events occurred in the sustained group. The best predictors of CAD were sustained hypercholesterolemia, lung involvement, and age at onset of SLE > 35 years. CONCLUSION: Within 3 years of diagnosis, 75.4% of patients with SLE had elevated TC, which was sustained in 40.3% of all patients. Older age at onset as well as increased cumulative dose of steroids and no antimalarial therapy are significant predictors of this group. It is this group that experiences the majority of CAD related events. Aggressive lipid lowering therapy should be targeted at such patients. PMID- 10529130 TI - C4A deficiency due to a 2 bp insertion is increased in patients with systemic lupus erythematosus. AB - OBJECTIVE: The association of C4A deficiency with systemic lupus erythematosus (SLE) is well documented. In Caucasian populations, the most common cause of C4A deficiency is a large gene deletion in linkage disequilibrium with a conserved MHC haplotype. Because of this linkage disequilibrium, it has been difficult to determine which of the genes constitutes the disease susceptibility allele. Evidence from non-caucasoid populations has supported a role for C4A deficiency in SLE. We investigated whether a specific genetic cause of C4A deficiency, not associated with A1, B8, DR3, is found with increased frequency in SLE compared to controls. METHODS: Polymerase chain reaction was used to identify carriers of a 2 base pair (bp) insertion in exon 29. In total, 188 patients with SLE from the Johns Hopkins lupus cohort and 222 controls were genotyped. RESULTS: The 2 bp insertion was found more frequently in patients with SLE compared to controls and was more common in Caucasian than in African American SLE patients. There were no clinical differences between patients that carried the mutation and those that did not. CONCLUSION: The association of this C4A null allele with SLE supports a role for C4A deficiency independent of other MHC associations in the etiopathogenesis of SLE. PMID- 10529131 TI - Genetic analysis of the contribution of IL10 to systemic lupus erythematosus. AB - OBJECTIVE: To study the contribution of the IL10 gene to the susceptibility to systemic lupus erythematosus (SLE). METHODS: Analysis by fluorescent semiautomated genotyping of a dinucleotide repeat located in the promoter region of the IL10 locus (microsatellite G). RESULTS: No significant difference was found in the frequencies of the microsatellite alleles of 330 Mexican patients with SLE compared to 368 controls from the same population. Two-point linkage analyses were carried out using 13 Mexican, 13 Swedish, and 8 Icelandic families with 2 or more cases with SLE. No linkage was revealed between IL10 and SLE, using a variety of parameter settings. CONCLUSION: Our results do not support that the IL10 gene contributes to the susceptibility to SLE in the populations we studied. PMID- 10529132 TI - Psychosocial correlates of morbidity in women with systemic lupus erythematosus. AB - OBJECTIVE: Modifiable psychosocial factors that are associated with health outcomes may provide new opportunities for treatment. We investigated the associations of various psychosocial factors with 3 measures of morbidity in women with systemic lupus erythematosus (SLE). METHODS: We collected information on 16 social, psychological, behavioral, and medical care factors in a cross sectional survey of 100 women with SLE, and related these to measures of physical disability (assessed by the Health Assessment Questionnaire Disability Index), SLE activity (assessed by the Systemic Lupus Activity Measure), and cumulative organ damage (assessed by the SLICC/ACR Damage Index). RESULTS: In multivariate analyses, greater physical disability was significantly associated with higher depression scale scores and higher body mass indexes. Greater SLE activity was associated with less adequate social support. Greater cumulative organ damage was associated with lower self-esteem and a time orientation that favored the present over the future. Financial barriers to medical care, knowledge about SLE, health locus of control, marital status, and health behaviors including compliance with medications, smoking, alcohol use, and exercise, were not significantly associated with any measure of morbidity. CONCLUSION: Selected psychosocial factors are associated with morbidity in SLE, but differ with the measure of morbidity examined. PMID- 10529133 TI - A disease severity scale for systemic sclerosis: development and testing. AB - OBJECTIVE: To develop and test a severity scale for individual organ involvements in systemic sclerosis (SSc, scleroderma). METHODS: An international study group completed the following tasks: (1) developed a glossary of terms including all pertinent variables for 9 potentially affected organ systems; (2) collected prospective data to determine the feasibility and practicality of each proposed variable; (3) revised the initial list of variables; (4) determined the association of each variable with mortality (a proxy for morbidity) using 579 patients in an existing comprehensive longitudinal scleroderma databank; (5) developed a severity grading scale for each organ system by discussion and consensus; and (6) externally validated the scale using an independent group of 680 patients from the same databank. RESULTS: Nine organ-specific severity scales were developed from 0 (no documented involvement) to 4 (endstage disease). The data required for scale completion are relatively easy and practical for all physicians to obtain. CONCLUSION: This preliminary severity scale will be useful for assessing disease severity status in individual patients both at one point in time and longitudinally. The severity scale will assist in the design and conduct of clinical trials and the comparison of study populations with one another. The scale will serve as a framework for developing a scleroderma disease activity index. PMID- 10529134 TI - Anti-annexin V antibodies and digital ischemia in patients with scleroderma. AB - OBJECTIVE: To elucidate the frequency and clinical significance of anti-annexin V antibodies in patients with scleroderma. METHODS: The study population consisted of 66 patients with scleroderma. Their sera were examined for IgG anti-annexin V antibodies by ELISA and immunoblotting. RESULTS: IgG anti-annexin V antibodies were detected by ELISA in 12 patients (18.2%) with scleroderma. Anti-annexin V antibodies were associated with digital ischemia in the patients with scleroderma. Only one patient of the 12 had anticardiolipin antibodies. Although 6 patients with anti-annexin V antibodies were also examined for activated partial thromboplastin time (APTT), none showed prolonged APTT. No IgG anti annexin V antibodies were detected by immunoblotting. CONCLUSION: Anti-annexin V antibodies in scleroderma are related to digital ischemia. These antibodies may be associated with the pathogenesis of vascular involvement in scleroderma. PMID- 10529135 TI - Preventive effect of an oral prostacyclin analog, beraprost sodium, on digital necrosis in systemic sclerosis. French Microcirculation Society Multicenter Group for the Study of Vascular Acrosyndromes. AB - OBJECTIVE: To compare the efficacy of 6 to 12 months of beraprost sodium (BPS) therapy with placebo in the prevention of digital ulceration in patients with systemic sclerosis (SSc). METHODS: One hundred seven patients with SSc were randomized in a multicenter double blind prospective trial. The primary endpoints were the percentage of patients with new digital ulceration, and median survival without recurrence of digital ulceration. Other secondary outcome measures included disability due to Raynaud's phenomenon, overall patient well being, the need for hospitalization, and endothelial damage, evaluated by variations in biological markers and nailfold microscopy. RESULTS: There was a trend towards fewer digital ulcerations in the BPS group than the placebo group (48.1 vs 58.8%, delta = 10.7%, p = 0.325), and median survival without recurrence of digital ulceration was longer in the BPS group (log-rank test, p = 0.233). Overall well being improved significantly more in the BPS group (p = 0.047), and von Willebrand factor decreased significantly more in the BPS group (p = 0.0001). The trend towards fewer digital ulcerations was more markedly in favor of BPS in the distal SSc group (delta for digital ulceration = 20.9%, p = 0.248) with a later onset (log-rank test, p = 0.057). Fewer patients were hospitalized in the BPS than the placebo group (4.0 vs 17.4%, p = 0.18). Side effects were mild. CONCLUSION: Our study suggests that oral beraprost sodium may benefit patients with SSc. Such patients, especially those with distal SSc, tend to have fewer recurrent ischemic digital ulcerations in winter, and the onset of their ulceration is delayed. PMID- 10529136 TI - Effects of non-mining occupational silica exposure on proportional mortality from silicosis and systemic sclerosis. AB - OBJECTIVE: Studies report an association between mining related silica exposure and systemic sclerosis (SSc). This study evaluated associations between non mining occupational silica exposure, silicosis, and SSc. METHODS: The US National Institute for Occupational Safety and Health (NIOSH) recently identified 36 non mining occupations in which workers face the "potential for substantial silica exposure." Data from death certificates in 25 US states during 1985-92 were analyzed to determine whether excess proportional mortality from silicosis or SSc occurred among those who worked in these occupations. RESULTS: Proportional mortality from silicosis in the occupations identified by NIOSH was significantly elevated among men [proportional mortality ratio (PMR) = 1.4; p < 0.0001] and women (PMR = 7.1; p = 0.002). The proportional mortality from SSc was not elevated among men (PMR = 1.0; p = 0.90) and was lower than expected among women (PMR = 0.8; p = 0.39). CONCLUSION: When cited on death certificates, the non mining occupational categories identified by NIOSH reflect prior silica exposure; however, no evidence of excess mortality from SSc was observed in these occupations. The reported increased frequency of SSc among miners may depend on a level or mode of silica exposure unique to the mining industry. PMID- 10529137 TI - Effects of hyaluronate on CD44 expression of infiltrating cells in exudate of rat air pouch, induced by sensitization with lipopolysaccharide. AB - OBJECTIVE: To investigate the effects of hyaluronate (HA) on CD44 expression of infiltrating cells in vivo. METHODS: Intra-air pouch injection of 10 microg lipopolysaccharide (LPS) with 0.4, 4.0, or 40 mg HA, 40 mg carboxymethylcellulose (CMC), or saline was performed on rats immunized with LPS. The percentage of CD44+ cells in the exudate of the air pouch was measured by flow cytometry, and the concentrations of interleukin 1beta (IL-1beta) and tumor necrosis factor alpha (TNF-alpha) in the air pouch exudate were measured by ELISA. IL-1beta and TNF-alpha in the air pouch lining layer were stained by immunohistochemistry. RESULTS: The percentage of CD44+ cells in air pouch exudate was greater in the presence of HA, with a dose dependent increase (0.4 mg, 9.4+/-2.6%, n = 4; 4.0 mg, 13.8+/-2.9%, n = 4; 40 mg, 24.9+/-6.3%, n = 3; p < 0.05), while it was 4.9+/ 1.2% (n = 4) in the presence of 40 mg CMC. The concentration of IL-1beta was lower in the presence of 40 mg HA (251.0+/-61.4 pg/ml, n = 4) or 40 mg CMC (168.2+/-43.5 pg/ml, n = 4; p < 0.05) than in saline (403.0+/-60.5 pg/ml, n = 4). The concentration of TNF-alpha was lower in the presence of 40 mg HA (14.0+/-6.7 pg/ml, n = 4) or 40 mg CMC (7.04+/-7.0 pg/ml, n = 4) than in saline (38.2+/-12.2 pg/ml, n = 4). Extensively stained lining cells in superficial layer of the air pouch with IL-1beta and TNF-alpha were observed in rats inoculated with 0.4 mg HA. CONCLUSION: These findings suggest that HA might affect the inflammatory process through modifying CD44 expression on infiltrating cells in air pouch exudate. PMID- 10529138 TI - Validity assessment of the disabilities of arm, shoulder, and hand questionnaire (DASH) for patients with psoriatic arthritis. AB - OBJECTIVE: To determine whether patients' perception of their functional ability, as measured by the disabilities of arm, shoulder. and hand (DASH) questionnaire, correlates with clinical measures of articular status in patients with psoriatic arthritis (PsA). METHODS: Patients attending the University of Toronto Psoriatic Arthritis Clinic between June and August 1997 were asked to complete a DASH questionnaire during their visits. Clinical assessments were performed according to a standard protocol including number of actively inflamed joints, total number of damaged joints, and grip strength. Spearman rank correlations were used to examine the relationship between clinical measures and the DASH questionnaire. RESULTS: Fifty consecutive patients, 28 men and 22 women, (mean age 49.2 yrs, mean disease duration 13 yrs) were included. DASH scores correlated with clinical measures of upper extremity function including right grip strength (r = -0.47, CI -0.67, -0.21) and number of active joints in the upper limbs (r = 0.65, CI 0.46, 0.79). As expected, the correlation between DASH scores and total number of active joints (r = 0.40, CI 0.14, 0.61) was lower than that between DASH scores and number of active joints in the upper limbs. The DASH was unrelated to clinical damage. CONCLUSION: DASH is a valid instrument for assessing upper extremity function and inflammatory disease activity in patients with PsA. PMID- 10529139 TI - Ethnicity and patterns of spondyloarthritis in South Africa--analysis of 100 patients. AB - OBJECTIVE: To determine the spectrum and ethnic differences of spondyloarthritis disease patterns in patients attending the Rheumatic Diseases Unit, University of Cape Town, South Africa. METHODS: A retrospective survey of case records of 100 patients with spondyloarthritis seen between January 1988 and January 1995. RESULTS: Of these 100 patients, 71 were male, 53 were Colored [mixed race descendants of Khoisan (Hottentot and Bushmen), Whites, Malays and Black Africans], 40 White, 5 Black and 2 Indian (descendants of immigrants from the Indian subcontinent). Our results show that the prevalence and disease patterns of spondyloarthritis in this South African cohort are comparable to those seen in Europe and North America with respect to clinical and radiological features, as well as therapeutic and orthopedic surgical requirements. No major ethnic differences in disease patterns were observed in White and Colored patients studied. CONCLUSION: The spectrum of spondyloarthritis in South Africa is similar to that seen elsewhere in the world. Our study confirmed the rarity of these conditions in Black South Africans. PMID- 10529140 TI - Biochemical markers of bone metabolism in mild ankylosing spondylitis and their relationship with bone mineral density and vertebral fractures. AB - OBJECTIVE: To determine the relationship of biochemical markers of bone metabolism in patients with mild ankylosing spondylitis (AS) with disease activity, bone mineral density (BMD), and vertebral fractures. METHODS: A total of 56 male patients with mild AS were studied. All patients had BMD measured by dual x-ray absorptiometry of the lumbar spine and hip and radiographs of the thoracic and lumbar spines. Radiographs of patients with AS were evaluated morphometrically and vertebral fractures were defined using established criteria. Serum osteocalcin, total and bone alkaline phosphatase (ALP, BALP, respectively), 25-hydroxyvitamin D (25-OHD), parathyroid hormone (PTH), urinary deoxypyridinoline (Dpyr), and pyridinoline (Pyr) were measured in the patients with AS and compared with 52 age and sex matched controls. Thirty-nine healthy male subjects aged 50-60 years, recruited from primary care registers, had spinal radiographs and served as controls for vertebral fractures. RESULTS: Patients with AS had reduced BMD of the lumbar spine, 0.98 (0.1) g/cm2 [T score = -1.14 (1.2) and Z score = -1.01 (1.2)], and femoral neck, 0.83 (0.1) g/cm2 [T score = 1.44 (1.2) and Z score = -0.73 (1.1)]. Of the 56 patients with AS, 11 (19.6%) had a vertebral fracture, whereas only one of 39 control subjects did (2.6%). Patients with AS had significantly lower mean serum osteocalcin compared with controls [9.03 (2.7) microg/l vs 11.05 (2.33) microg/l; p < 0.001], and significantly higher mean serum ALP [73.09 (19.5) U/l vs 53.02 (16.7) U/l; p < 0.001] and BALP [38.54 (9.9) U/l vs 30.35 (8.28) U/l; p < 0.001]. There was no significant difference in the excretion of Dpyr and Pyr between patients with AS and controls [4.90 (3.9) nmol/mmol vs 4.00 (3.6) nmol/mmol, and 15.66 (10.8) nmol/mmol vs 12.24 (10.3) nmol/mmol, respectively]. There were no significant differences in the mean 25-OHD and PTH levels in patients with AS compared to controls [20.03 (1.6) micromol/l vs 18.9 (2.9) micromol/l and 3.31 (1.5) pmol/l vs 2.63 (0.4) pmol/l, respectively]. The biochemical markers of bone formation and resorption correlated well with inflammatory indices of disease activity (acute phase reactants), but not with BMD of the lumbar spine or femoral neck. No significant difference was seen in any marker of bone turnover when AS patients with vertebral fractures were compared with those without. CONCLUSION: Bone turnover in patients with mild AS is characterized by low serum osteocalcin and ALP in the presence of normal calciotropic hormones. Biochemical markers of bone metabolism do not correlate with BMD or vertebral fractures. The disparity between osteocalcin and alkaline phosphatase in patients with AS needs further evaluation. PMID- 10529141 TI - Osteoporosis, body composition, and bone turnover in ankylosing spondylitis. AB - OBJECTIVE: To study the prevalence of osteoporosis (OP) and osteopenia in ankylosing spondylitis (AS) and to investigate the relationship between symptomatic and structural severity, the indices of bone turnover, and body composition. METHODS: Eighty patients with AS were enrolled prospectively: 52 men (65%) and 28 women, mean age 36.7 years +/- 11.5 (range 18-67); they were studied clinically, radiologically, and by dual energy x-ray absorptiometry. Sixty-three underwent biological assessment of bone turnover markers. RESULTS: OP and osteopenia as defined by the World Health Organization (T score < -2.5 SD and between -1 and -2.5 SD, respectively) were observed in 15 (18.7%) and 25 patients (31.2%) at the lumbar spine and in 11 (13.7%) and 33 patients (41.2%) at the femoral neck, respectively. Patients with OP had a lower body mass index (BMI) and fat mass percentage. There was a trend to a lower disease duration in patients with OP at the spine than in healthy subjects. Bone resorption markers (urinary D-pyridinoline or C-telopeptide concentrations) were increased in 34 patients (53.9%). Bone turnover markers were positively correlated with C reactive protein concentration and Larsen radiological hip score; they were negatively correlated with Schober index and fat mass percentage. CONCLUSION: (1) OP is frequent in AS and can be observed in early stages of the disease. (2) Patients with AS are more susceptible to develop OP when they have low BMI, low fat mass percentage, and active and severe disease. OP was observed in parallel with increased bone resorption. PMID- 10529142 TI - Significance of endogenous heat shock protein in adjuvant arthritis. AB - OBJECTIVE: To investigate the role of endogenous heat shock protein (HSP) (rat 60 kDa HSP, rHSP 60) in adjuvant arthritis (AA), the expression of rHSP 60 in AA susceptible Lewis rats and AA-resistant Fisher rats was studied. METHODS: The extent of arthritis was assessed by measuring footpad thickness. The proliferative response of mononuclear cells (MNC) to mycobacterial 65 kDa HSP (mHSP 65) was measured by [3H] thymidine incorporation. The messenger RNA for rHSP 60 was quantified by dot blot hybridization in peripheral blood mononuclear cells (PBMNC), splenic MNC (SMNC), and ankle joint synovial membrane (SM). RESULTS: In Lewis rats: AA developed on Day 14 after immunization with Freund's complete adjuvant. Proliferative responses of PBMNC and SMNC to mHSP 65 were observed on Day 14 and thereafter according to the development of AA. Messenger RNA levels of rHSP 60 in PBMNC, SMNC, and SM did not increase until Day 21. In Fisher rats: AA and proliferative response of MNC to mHSP 65 were not observed throughout the observation period. Messenger RNA levels of rHSP 60 were significantly higher in preimmune Fisher rats than those in preimmune Lewis rats. Messenger RNA levels of rHSP 60 in PBMNC and SMNC increased significantly on Day 7 and decreased to preimmune levels until Day 21. CONCLUSION: We propose that elevated levels and rapid elevation of endogenous HSP 60 levels relate to resistance to AA in Fisher rats; however, lower levels and delayed increase of endogenous HSP 60 levels relate to susceptibility to AA in Lewis rats. PMID- 10529143 TI - The influence of silicone implantation on murine lupus in MRL lpr/lpr mice. AB - OBJECTIVE: The use of silicone breast implants has been implicated in the development of autoimmune connective tissue diseases including systemic lupus erythematosus (SLE). We examined the influence of implanted silicones in MRL lpr/lpr and MRL +/+ mice, to determine whether silicone increases autoimmunity and exacerbates experimental lupus. METHODS: Mice were implanted with either silicone gel or silicone oil (polydimethylsiloxane; PDMS), while saline injected mice were used as controls. Proteinuria levels, palpation of lymphadenopathy, serum autoantibodies, circulating cytokines, and weight change were monitored for 18 weeks, when terminal glomerulonephritis was evaluated by histopathological techniques. Proteins were extracted from the surface of recovered implants, and the composition and immune reactive status of the silicone-binding proteins (SBP) were investigated. RESULTS: No adverse influence of silicone gel or silicone oil on the clinical aspects of lupus was observed. However, anti-DNA antibodies were significantly increased in MRL mice implanted with silicone gel compared to the control animals, and rheumatoid factor titers were modestly increased in implanted MRL lpr/lpr mice. Serum cytokine levels were influenced by silicone implantation in MRL lpr/lpr mice (but not MRL +/+ mice), with interleukin 1 (IL 1) levels increased in gel implanted animals and IL-2 levels elevated in PDMS (silicone oil) implanted mice. Different SBP were detected on implants recovered from MRL lpr/lpr mice compared with MRL +/+ mice, and Western blotting revealed the presence of strong autoantibodies to SBP in sera from MRL lpr/lpr mice, but not MRL +/+ mice. CONCLUSION: These findings suggest that silicone implantation may influence immunological responses during murine lupus, including the provocation or exacerbation of autoantibodies. However, these immune modifications did not appear to influence the clinical variables of this experimental lupus model. PMID- 10529145 TI - Muscle dysfunction in elderly individuals with hip fracture. AB - OBJECTIVE: To investigate muscle metabolism in elderly people with hip fracture. METHODS: Free carnitine, carnitine esters, and respiratory chain enzyme activity were measured in muscle tissue from 54 patients over 65 years who underwent surgery for hip fracture, and from 40 healthy controls. RESULTS: Eighty-five percent of patients older than 85 have either abnormal carnitine distribution or defects in the respiratory chain. CONCLUSION: Elderly patients with hip fracture have muscle metabolic alterations that may contribute to neuromuscular impairment and be amenable to therapy. PMID- 10529144 TI - Percutaneous vertebroplasty in the treatment of osteoporotic vertebral compression fractures: an open prospective study. AB - OBJECTIVE: To assess the efficacy and safety of percutaneous vertebroplasty in osteoporotic vertebral compression fractures responsible for severe and persistent pain. METHODS: Sixteen patients were included in this open prospective study. Inclusion criteria were: one or 2 vertebral fractures responsible for severe pain, i.e., higher than 50 mm on a visual analog scale (VAS: 0-100 mm), scores 3, 4 or 5 according to the McGill-Melzack scoring system, and evolving for more than 3 months. Assessment criteria were the changes over time (Days 3, 30, 90, 180) in VAS and McGill-Melzack scoring system. The changes over time in a generic health status instrument score [the Nottingham Health Profile (NHP)] were also assessed. Statistical comparisons were performed using the Wilcoxon T test. RESULTS: There were 9 women and 7 men: postmenopausal osteoporosis (n = 7), corticosteroid induced osteoporosis (n = 2), and male osteoporosis (n = 7). Vertebroplasty was performed in 20 vertebrae. A statistically significant decrease of both VAS (-53%, p < 0.0005) and McGill-Melzack scoring system (p < 0.005) was observed at Day 3. The results were also significant at Days 30, 90, and 180 for both scales (p < 0.005 and p < 0.01, respectively). A significant decrease over time for 5/6 dimensions of the NHP score was also noted: pain (p < 0.01), physical mobility (p < 0.05), emotional reactions (p < 0.05), social isolation (p < 0.05), and energy (p < 0.05). We observed no adverse event, and no vertebral fracture has occurred after 6 months of followup. CONCLUSION: Percutaneous vertebroplasty is a useful and safe procedure for treating persistent painful osteoporotic fractures. Controlled studies with longterm followup are required. PMID- 10529146 TI - Validity and reliability of the 6 minute walk in persons with fibromyalgia. AB - OBJECTIVE: To assess the reliability and construct validity of the 6 minute walk (6MW) in persons with fibromyalgia (FM) and to determine an equation for predicting peak oxygen consumption (pVO2) from the distance covered in 6 minutes. METHODS: Ninety-six women who met the American College of Rheumatology (ACR) criteria for FM were tested on the 6MW and the Fibromyalgia Impact Questionnaire (FIQ). A subset (n = 23) were tested on a separate day for pVO2 during a symptom limited, incremental treadmill test. Twelve subjects repeated the 6MW five times over 10 days. Heart rate and rating of perceived exertion (RPE) were recorded for each walk. Intraclass correlations were used to determine the reliability of the 6MW. Validity was examined by correlating the 6MW with pVO2 and the FIQ. Body mass index (BMI) and 6MW were independent variables in a stepwise regression to predict pVO2. RESULTS: A significant increase in distance occurred from Walk 1 to Walk 2 (p = 0.000) with the distance maintained on the remaining walks (p = 0.148) The correlations of the 6MW with the FIQ and pVO2 were -0.325 and 0.657, respectively. The regression equation to predict pVO2 from 6MW distance and BMI was: pVO2 (ml/kg/min) = 21.48 + (-0.4316 x BMI) + [0.0304 x distance(m)] (R = 0.76, R2 = 0.66). CONCLUSION: When using the 6MW it is necessary to conduct a practice walk, with the second walk taken as the baseline measure. It was determined from the correlations that the 6MW cannot replace the FIQ as a measure of function. The 6MW may be used as an indicator of aerobic fitness, although obtaining VO2 by means of a graded exercise test is preferable. PMID- 10529148 TI - Increased circulating vascular endothelial growth factor is correlated with disease activity in polyarticular juvenile rheumatoid arthritis. AB - OBJECTIVE: To investigate the relevance of vascular endothelial growth factor (VEGF) in the pathogenesis of juvenile rheumatoid arthritis (JRA). METHODS: Serum VEGF levels in 58 patients with JRA (systemic in 17, polyarticular in 29, pauciarticular in 12) were measured by ELISA and compared with those of 21 patients with infectious diseases and 50 healthy children. Correlations of VEGF levels with number of joints with active arthritis, erythrocyte sedimentation rate (ESR), and hyaluronic acid (HA) were examined. RESULTS: Serum levels of VEGF in patients with JRA were significantly higher than in healthy controls. Patients with systemic and polyarticular JRA showed statistically higher levels of VEGF than those with infectious diseases. VEGF levels correlated statistically with C reactive protein (CRP) in patients with both infectious diseases and polyarticular JRA, but the regression slope (VEGF/CRP) was much steeper in polyarticular JRA than in infectious diseases. Serum VEGF levels correlated with disease activity variables such as the number of joints with active arthritis, ESR, and serum HA levels in polyarticular JRA. CONCLUSION: The correlation of serum VEGF levels and disease activity in polyarticular JRA suggests that VEGF may take an active part in joint inflammation. PMID- 10529147 TI - Knee magnetic resonance imaging in childhood chronic monarthritis. AB - OBJECTIVE: To describe the usefulness of magnetic resonance imaging (MRI) of the knee in the evaluation of chronic monarthritis of uncertain cause in childhood. METHODS: We retrospectively reviewed 21 children referred to our clinic with a putative diagnosis of chronic inflammatory monarthritis of the knee who had MRI performed between May 1993 and June 1997. The median age was 13 years (range 2 17) and 11 were girls. RESULTS: The clinical diagnosis prior to MRI assessment was inflammatory arthritis in 16 patients, and a primary noninflammatory cause in 5. MRI was done in the patients with presumptive inflammatory arthritis when there were atypical symptoms, signs, or radiographs (n = 14), or when they failed to respond to therapy (n = 2). In the patients with a presumptive noninflammatory diagnosis, MRI was performed to clarify the diagnosis. Twelve children (57%) had MRI evidence of a noninflammatory diagnosis. In 4 children (19%) the MRI study indicated the presence of arthritis, and in 5 children (24%) the MRI studies were normal. The noninflammatory diagnoses included: lipoma arborescens (n = 1), vascular malformation [intraarticular (n = 1), extraarticular (n = 1)], synovial chondromatosis (n = 2), partial anterior cruciate ligament tear (n = 2), traumatic bone contusion (n = 2), possible meniscal tear (n = 1), osteochondritis dissecans (n = 1), and a soft tissue mass of uncertain significance in the suprapatellar pouch (n = 1). CONCLUSION: Inflammatory arthritis is usually diagnosed by clinical assessment alone. Uncommonly, when a single joint is involved, and atypical features are identified by a pediatric rheumatologist, other causes of chronic pain and swelling need to be excluded. In this selected patient population, MRI is a useful tool either to confirm the presence of inflammatory arthritis or to investigate a wide range of pathology that can mimic knee joint arthritis. PMID- 10529149 TI - Two cases of methotrexate induced lymphomas in rheumatoid arthritis: an association with increased serum IgE. AB - We describe 2 patients with rheumatoid arthritis (RA) in whom non-Hodgkin's lymphomas developed during low dose pulsed methotrexate (MTX) treatment. The tumors regressed after discontinuation of MTX with no additional treatment. Serum levels of IgE increased concomitantly with the development of lymphoma, and decreased along with the regression of the lymphoma in both patients. These 2 cases and a review of the literature suggest that measuring serum IgE may have a predictive value for monitoring lymphoma in patients with RA treated with MTX. PMID- 10529150 TI - Complex regional pain syndrome type-1: a rare complication of arteriovenous graft placement. AB - Complex regional pain syndrome (CRPS) type-1 (previously termed reflex sympathetic dystrophy syndrome) may be manifested as sympathetically mediated pain and swelling in an extremity. Among the numerous causes of reflex sympathetic dystrophy, the most common is trauma. We describe a 71-year-old diabetic man with endstage renal disease who presented with CRPS type-1 of the left hand one month after construction of a PTFE (polytetrafluroethylene) arteriovenous graft. The symptoms of CRPS improved greatly with stellate ganglion blocks and physical therapy. PMID- 10529151 TI - Dramatic improvement of left ventricular function after cytotoxic therapy in lupus patients with acute cardiomyopathy: report of 6 cases. AB - Although lupus cardiomyopathy is thought to be clinically uncommon, we encountered 6 patients with systemic lupus erythematosus (SLE) over a 10 year period who had severe left ventricular dysfunction and showed remarkable improvement in their cardiac function after cytotoxic therapy. All patients met the American College of Rheumatology criteria for classification of SLE and presented with signs of severe biventricular failure relatively early in their disease. Concurrent manifestations of SLE at the time of cardiomyopathy included rash, arthritis, myalgias, pleuritis, pericarditis, and nephritis. Four of the 6 patients were taking prednisone 20 mg/day at the time heart failure developed. In all cases the CPK were normal. Evaluation of cardiac function by echocardiogram and/or radionuclide gated blood pool scintigraphy revealed a severe depression of ventricular function with initial left ventricular ejection fraction (LVEF) ranging from 11 to 34% (mean 19%). Within 6 months of initiation of cytotoxic treatment all patients showed a dramatic response: the post-treatment LVEF ranged from 25 to 55%. This series of patients suggests that cardiomyopathy may be a more common complication of SLE than previously reported. Cardiomyopathy occurs relatively early in the course of SLE, may lead to severe cardiac dysfunction despite corticosteroid therapy, and appears to be responsive to cytotoxic therapy. PMID- 10529152 TI - Catastrophic manifestation of the antiphospholipid syndrome. AB - We describe a young woman who displayed the "malignant" variant of the antiphospholipid syndrome (APS), also known as the "catastrophic APS." Renal insufficiency, retinopathy, cerebral infarcts, bone marrow necrosis, skin ulcers, and nasal septum perforation were the result of widespread thrombotic microangiopathy. She recovered during high intensity anticoagulation. PMID- 10529153 TI - De novo systemic sclerosis after radiotherapy: a report of 3 cases. AB - We describe 3 patients in whom the onset of systemic scleroderma occurred shortly after ionizing irradiation for nasopharyngeal or breast carcinoma. This relationship has been described rarely as has the exacerbation of a preexisting scleroderma after irradiation. This gives indications for direction of studies. PMID- 10529154 TI - Colon stricture and volvulus in a patients with scleroderma. AB - Gastrointestinal involvement in scleroderma is almost universal. We describe a patient with a benign stricture and volvulus of transverse colon, a life threatening but treatable complication of scleroderma bowel disease. We review the literature on colon volvulus in scleroderma and discuss the importance of recognizing this rare complication. PMID- 10529155 TI - Isolated central nervous system vasculitis associated with hepatitis C infection. AB - Since its identification in 1989, hepatitis C has been implicated in the pathogenesis of an increasing number of diseases previously believed to be primary or idiopathic. We report 2 rarely seen cases of isolated central nervous system (CNS) vasculitis in patients with hepatitis C infection. Patient 1. A 43 year-old man with 4 day right temporal headache developed a left hemiparesis. Weakness was his only physical finding. Computed tomography (CT) scan demonstrated a large right frontotemporal hemorrhage, and angiography revealed focal dilatations and irregularities of multiple branches of the right middle and anterior cerebral arteries. Cerebral decompression was performed and leptomeningeal biopsies showed granulomatous angiitis. Laboratory results were normal except for elevated liver biochemical tests. Later testing for hepatitis C was positive. His neurological symptoms improved with corticosteroids and cyclophosphamide. Patient 2. A 39 yr old male developed 3 days of left sided weakness, slurred speech and difficulty swallowing fluids. Physical findings were limited to his weakness. Magnetic resonance imaging demonstrated a right superior pontine subacute infarct with a small left internal capsule lacunar infarct. Angiography revealed multiple areas of focal narrowing with no areas of abrupt vessel cut off. Cerebral spinal fluid showed 71 PMN, 29 RBC, normal glucose, elevated protein (64 mg/dl), no oligoclonal bands, and low myelin basic protein. Other laboratory analyses were normal including liver biochemical tests. However, hepatitis C serology was positive and mixed cryoglobulins were detected. CNS vasculitis was diagnosed and nearly full recovery was achieved with corticosteroids, cyclophosphamide and warfarin. PMID- 10529156 TI - Recurrent reactive arthritis associated with urinary tract infection by Escherichia coli. AB - We describe a patient with recurrent Escherichia coli urinary tract infection followed by recurrent reactive arthritis. During a 9 year period the patient developed 4 episodes of arthritis. During each attack, triggering infections were thoroughly investigated but no other causative infection was found. Although the urinary tract is not routinely targeted for triggering infections for reactive arthritis, we suggest that urinary tract infections should be included in the diagnostic investigations of patients with acute arthritis. PMID- 10529157 TI - Paucity of radiographic progression in rheumatoid arthritis treated with methotrexate as the first disease modifying antirheumatic drug. PMID- 10529158 TI - Circulating levels of matrix metalloproteinases in rheumatic diseases. PMID- 10529159 TI - Sonographic assessment of shoulder complaints in rheumatic diseases. PMID- 10529160 TI - Childhood lipoma arborescens. PMID- 10529161 TI - Treatment of rheumatoid arthritis with the dopamine agonist quinagolide. PMID- 10529162 TI - Treatment of acute attacks of gout with a small dose of intraarticular triamcinolone acetonide. PMID- 10529163 TI - Macrophagic myofasciitis: improvement with antibiotic therapy. PMID- 10529164 TI - Gadolinium contrast magnetic resonance imaging of the temporal artery in giant cell arteritis. PMID- 10529165 TI - Tuberculous rheumatism presenting as spondyloarthropathy. PMID- 10529167 TI - Orientation of chlorophyll transition moments in the higher-plant light harvesting complex CP29. AB - The Q(y) transition dipole moment vectors of all eight chlorophylls in the higher plant antenna protein CP29 were calculated by an original method on the basis of linear dichroism and absorption spectroscopy. The contribution of individual chromophores was determined from difference spectra between wild type and mutant proteins in which a single chlorophyll has been removed by mutating pigment binding residues. Recombinant proteins were constructed by overexpressing the apoprotein in bacteria and refolding of the pigment-protein complex in vitro [Bassi, R., Croce, R., Cugini, D., and Sandona, D. (1999) Proc. Natl. Acad. Sci. U.S.A. (in press)]. The spectroscopic data are interpreted on the basis of a protein structural model obtained via the homology with the major antenna complex LHCII [Kuhlbrandt, W., Wang, D. N., and Fujiyoshi, Y. (1994) Nature 367, 614 621]. The results allow us to determine the orientation of six chromophores within the protein structure. The orientations of the two remaining chromophores are inferred by considering the symmetry properties of CP29 and fitting steady state absorption and linear dichroism spectra by independent chlorophyll spectral forms. As a consequence, four "mixed" sites with different chlorophyll a and b binding affinities are identified in CP29. Geometrical data and the Forster mechanism for energy transfer suggest that excitation energy equilibrates rapidly among chlorophyll "pure" sites while energy preferentially flows outward from chlorophyll "mixed" sites. The orientation of the dipole moments of two chlorophyll molecules symmetrically located at the center of the protein and parallel to the carotenoid transition vectors suggests a role in energy transfer from xanthophyll to chlorophyll. PMID- 10529166 TI - Binding of exogenously added carbon monoxide at the active site of the iron-only hydrogenase (CpI) from Clostridium pasteurianum. AB - A site for the binding of exogenously added carbon monoxide has been identified at the active site of the Fe-only hydrogenase (CpI) from Clostridium pasteurianum. The binding and inhibition of carbon monoxide have been exploited in biochemical and spectroscopic studies to gain mechanistic insights. In the present study, we have taken advantage of the ability to generate an irreversibly carbon monoxide bound state of CpI. The crystallization and structural characterization of CpI inhibited in the presence of carbon monoxide indicates the addition of a single molecule of carbon monoxide. The ability to generate crystals of the carbon monoxide bound state of the hydrogenase that are isomorphous to those of the native enzyme has allowed for a direct comparison of the crystallographic data and an unambiguous identification of the site of carbon monoxide binding at the active site of CpI. Carbon monoxide binds to an Fe atom of the 2Fe subcluster at the site of a terminally bound water molecule in the as crystallized native state of CpI that has been previously suggested to be a potential site of reversible hydrogen oxidation. Binding of carbon monoxide at this site results in an active site that is coordinately saturated with strong ligands (S, CO, and CN), providing a rational potential mechanism for inhibition of reversible hydrogen oxidation at the active site of CpI. PMID- 10529168 TI - NMR structure of the complex between the zinc finger protein NCp10 of Moloney murine leukemia virus and the single-stranded pentanucleotide d(ACGCC): comparison with HIV-NCp7 complexes. AB - The structure of the 56 amino acid nucleocapsid protein NCp10 of retrovirus MoMuLV, which contains a single CX(2)CX(4)HX(4)C-type zinc finger, has been determined previously by NMR. The important role of NCp10 (or NCp7 for HIV-1) in the retroviral life cycle seems mainly related to their preferential binding to single-stranded nucleic acids. We report here the structure of the complex formed between the biologically active (14-53)NCp10 and the oligonucleotide d(ACGCC) in aqueous solution determined by 2D (1)H NMR based methods. The aromatic residue Trp(35) of NCp10 directs nucleic acid complexation as shown by its complete fluorescence quenching upon addition of d(ACGCC). (1)H and (31)P NMR studies support the insertion of Trp(35) between the G(3) and C(4) bases. A total of 577 NOE distance restraints, of which 40 were intermolecular, were used for the structure determination. The zinc finger provides a well-defined surface for the binding of d(ACGCC) through hydrophobic interactions and tryptophan stacking on the guanine. This latter interaction was also observed in the NMR-derived structures of the complexes between NCp7, which contains two successive zinc fingers, and single-stranded DNA and RNA, supporting the proposal for a major role played by aromatic residues of NCp proteins in nucleic acid recognition. Upon binding to the nucleotide a new loop in NCp10 that participates in the intermolecular interaction is formed. Additional interactions provided by positively charged residues surrounding the zinc finger appear necessary for tight binding. The structure of the complex NCp10-d(ACGCC) gives a structural explanation for the loss of virus infectivity following point mutations in the finger domain. PMID- 10529169 TI - Three-dimensional structure of adenosylcobinamide kinase/adenosylcobinamide phosphate guanylyltransferase (CobU) complexed with GMP: evidence for a substrate induced transferase active site. AB - The X-ray crystal structure of adenosylcobinamide kinase/adenosylcobinamide phosphate guanylyltransferase (CobU) from Salmonella typhimurium bound to GMP has been determined by molecular replacement to 2.2 A resolution. CobU is a bifunctional enzyme, which catalyzes the phosphorylation of the 1-amino-O-2 propanol side chain of the adenosylcobinamide ring and subsequently functions as a guanylyltransferase to form adenosylcobinamide.GDP. The transferase activity involves a covalent enzyme-guanylyl intermediate that is most likely a phosphoramidate linkage to His(46). Previous studies have shown that the enzyme is a homotrimer and adopts a pinwheel shape. Each subunit consists of a single domain of six parallel beta-strands and one antiparallel strand flanked on either side by a total of five alpha-helices and one helical turn. Interestingly, His(46) in the apoenzyme is located a considerable distance from the kinase active site or P-loop motif and is solvent-exposed [Thompson, T. B., et al. (1998) Biochemistry 37, 7686-7695]. To examine the structural relationship of the two active sites, CobU was cocrystallized with GTP and pyrophosphate. Crystals belong to space group P2(1)2(1)2(1) with the following unit cell dimensions: a = 58. 4 A, b = 87.8 A, and c = 101.6 A. The structure shows electron density for the hydrolysis product GMP rather than the expected covalent guanylyl intermediate which appears to have been hydrolyzed in the crystal lattice. Even so, CobU exhibits a substantial conformational rearrangement. The helix axis containing His(46), the site of guanylylation, rotates 30 degrees and translates 11 A relative to the apo structure and is accompanied by compensatory unwinding and rewinding at the helix ends to allow the induction of a guanosine binding pocket between beta-strand 2 and alpha-helix 2. This conformational change brings the C(alpha) of His(46) approximately 10 A closer to the P-loop motif such that a phosphate ion located in the P-loop is only 6 A from the alpha-phosphate of GMP. This suggests that the P-loop motif may be used to coordinate the terminal phosphates in both the transferase and kinase reactions and implies that the active sites for both reactions overlap. PMID- 10529170 TI - Formation of short-lived protein aggregates directly from the coil in two-state folding. AB - Recent results on the 102 residue protein U1A show that protein aggregation is not always slow and irreversible but may take place transiently in refolding studies on a millisecond time scale. In this study we observe a similar aggregation behavior with the classical two-state protein CI2. Since both U1A and CI2 appear to fold directly from the coil at low protein concentrations, it is likely that the aggregates also form directly from the coil. This is in contrast to the behavior of larger multistate proteins where aggregation occurs in connection to "sticky" intermediates. PMID- 10529171 TI - Identification of residues in the monocyte chemotactic protein-1 that contact the MCP-1 receptor, CCR2. AB - The CC chemokine, MCP-1, has been identified as a major chemoattractant for T cells and monocytes, and plays a significant role in the pathology of inflammatory diseases. To identify the regions of MCP-1 that contact its receptor, CCR2, we substituted all surface-exposed residues with alanine. Some residues were also mutated to other amino acids to identify the importance of charge, hydrophobicity, or aromaticity at specific positions. The binding affinity of each mutant for CCR2 was assayed with THP-1 and CCR2-transfected CHL cells. The majority of point mutations had no effect. Residues at the N-terminus of the protein, known to be crucial for signaling, contribute less than a factor of 10 to the binding affinity. However, two clusters of primarily basic residues (R24, K35, K38, K49, and Y13), separated by a 35 A hydrophobic groove, reduced the level of binding by 15-100-fold. A peptide fragment encompassing residues 13 35 recapitulated some of the mutational data derived from the intact protein. It exhibited modest binding as a linear peptide and dramatically improved affinity when the region which adopts a single turn of a 3(10)-helix in the protein, which includes R24, was constrained by a disulfide bond. Additional constraints at the ends of the peptide, corresponding to the disulfide between the first and third cysteines in MCP-1, yielded further improvements in affinity. Together, these data suggest a model in which a large surface area of MCP-1 contacts the receptor, and the accumulation of a number of weak interactions results in the 35 pM affinity observed for the wild-type (WT) protein. The receptor binding site of MCP-1 also is significantly different from the binding sites of RANTES and IL-8, providing insight into the issue of receptor specificity. It was previously shown that the N-terminus of CCR2 is critical for binding MCP-1 [Monteclaro, F. S., and Charo, I. F. (1996) J. Biol. Chem. 271, 19084-92; Monteclaro, F. S., and Charo, I. F. (1997) J. Biol. Chem. 272, 23186-90]. Point mutations of six acidic residues in this region of the receptor were made to test their role in ligand binding. This identified D25 and D27 of the DYDY motif as being important. On the basis of our data, we propose a model in which the receptor N-terminus lies along the hydrophobic groove in an extended fashion, placing the DYDY motif near the basic cluster involving R24 and K49 of MCP-1. This in turn orients the signaling residues (Y13 and the N-terminus) for productive interaction with the receptor. PMID- 10529172 TI - Intradomain distances in the regulatory domain of the myosin head in prepower and postpower stroke states: fluorescence energy transfer. AB - The relative movement of the catalytic and regulatory domains of the myosin head (S1) is likely to be the force generating conformational change in the energy transduction of muscle [Rayment, I., Holden, H. M., Whittaker, M., Yohn, C. B., Lorenz, M., Holmes, K. C., and Milligan, R. A. (1993) Science 261, 58-65]. To test this model we have measured, using frequency-modulated FRET, three distances between the catalytic domain and regulatory domains and within the regulatory domain of myosin. The donor/acceptor pairs included MHC cys707 and ELC cys177; ELC cys177 and RLC cys154; and ELC cys177 and gizzard RLC cys108. The IAEDANS (donor) or acceptor (DABMI or IAF) labeled light chains (ELC and RLC) were exchanged into monomeric myosin and the distances were measured in the putative prepower stroke states (in the presence of MgATP or ADP/AlF(4-)) and the postpower stroke states (ADP and the absence of nucleotides). For each of the three distances, the donor/acceptor pairs were reversed to minimize uncertainty in the distance measured, arising from probe orientational factors. The distances obtained from FRET were in close agreement with the distances in the crystal structure. Importantly, none of the measured distances varied by more than 2 A, putting a strong constraint on the extent of conformational changes within S1. The maximum axial movement of the distal part of myosin head was modeled using FRET distance changes within the myosin head reported here and previously. These models revealed an upper bound of 85 A for a swing of the regulatory domain with respect to the catalytic domain during the power stroke. Additionally, an upper bound of 22 A could be contributed to the power stroke by a reorientation of RLC with respect to the ELC during the power stroke. PMID- 10529173 TI - Mutagenesis of three residues, isoleucine-60, threonine-61, and aspartic acid-80, implicated in the GTPase activity of Escherichia coli elongation factor Tu. AB - The properties of variants of elongation factor (EF) Tu mutated at three positions implicated in its GTPase activity are presented. Mutation I60A, which reduces one wing of a "hydrophobic barrier" screening off the nucleophilic water molecule found at the GTP gamma-phosphate, causes a reduction of the intrinsic GTPase activity contrary to prediction and has practically no influence on other properties. Mutation D80N, which in the isolated G-domain of EF-Tu caused a strong stimulation of the intrinsic GTPase, reduces this activity in the intact molecule. However, whereas for wild-type EF-Tu complex formation with aa-tRNA reduces the GTPase, EF-Tu[D80N] shows a strongly increased activity when bound to Phe-tRNA. Moreover, ribosomes or kirromycin can stimulate its GTPase up to the same level as for wild-type. This indicates that a local destabilization of the magnesium binding network does not per se cause an increased GTPase but does affect its tight regulation. Interestingly, mutant D80N sequestrates EF-Ts by formation of a more stable complex. Substitutions T61A and T61N induce low intrinsic GTPase, and the stimulation by ribosome is less for T61A than for T61N but still detectable, while kirromycin stimulates the GTPase of both mutants equally. This provides more evidence that stimulation by kirromycin and ribosomes follows a different mechanism. The functional implications of these mutations are discussed in the context of a transition state mechanism for catalysis. An alternative structural explanation for the strong conservation of Ile-60 is proposed. PMID- 10529174 TI - The single amino acid changes in the yeast mitochondrial S4 ribosomal protein cause temperature-sensitive defect in the accumulation of mitochondrial 15S rRNA. AB - Four different mutant alleles of a nuclear gene (MNA6), which lose mt 15S rRNA at nonpermissive temperature (36 degrees C), were previously generated by EMS mutagenesis of Saccharomyces cerevisiae. To understand the biochemical basis for the loss of 15S rRNA in these mutants, the wild-type and mutant alleles of the MNA6 gene were isolated and characterized. The DNA sequencing of the cloned MNA6 gene revealed that it has an open reading frame specifying a 486 amino acid polypeptide, which appears to be a yeast mt homologue of the S4 r-protein family. The large size of this yeast S4 homologue is due to a nonhomologous long C terminal extension. The MNA6 gene also appeared to be identical to the previously isolated yeast NAM9 gene. The in vitro expression under coupled transcription translation reaction conditions followed by mt import demonstrated that MNA6 indeed encodes a approximately 56 kDa protein targeted to the mitochondria. We have also demonstrated by Western blot analysis using anti-Mna6p antibody that Mna6p is associated with the small subunit of mitoribosomes. The sequence analysis of the four mutant mna6 alleles revealed that Leu(109) --> Phe, Arg(111) --> Lys, Pro(424) --> Leu, or Pro(438) --> Leu amino acid substitution in Mna6p causes temperature-dependent loss of the 15S rRNA. These mutations do not affect the mitochondrial import or accumulation of Mna6p. Rather the evidence points to an inability of mutant Mna6p to be assembled into the mitoribosomes of cells grown at 36 degrees C. PMID- 10529176 TI - The peptide bond between E292-A293 of Escherichia coli leucyl-tRNA synthetase is essential for its activity. AB - Escherichia coli leucyl-tRNA synthetase (LeuRS) is a class I aminoacyl-tRNA synthetase that contains a large connecting polypeptide (CP1) inserted into its nucleotide binding fold, or active site. In this study, purified leucyl-tRNA synthetase was found to be cleaved between E292 and A293 in its CP1 domain. SDS PAGE analysis showed peptides of 63 and 34 kDa in addition to the native 97.3 kDa synthetase. By internal complementation, the two peptides could form a 97.3 kDa complex similar to the native LeuRS. This complex could support the ATP approximately PP(i) exchange activity of LeuRS, but could not complement for aminoacylation. To study the function of the region around the bond of E292 and A293, four pairs of peptides resulting from different cleavage sites in CP1 were reconstituted in vivo. With the exception of the enzyme assembled from the E292 A293 cleavage site, all the reassembled LeuRSs catalyzed the aminoacylation of tRNA(Leu). Although the E292-A293-cleaved LeuRS could not catalyze aminoacylation, fluorescence titration revealed that its tRNA binding ability was almost identical to that of wild-type LeuRS. These results suggest that the region around E292-A293 may be responsible for maintaining the proper conformation of LeuRS required for the tRNA charging activity. PMID- 10529175 TI - Genetic evidence that stress-activated p38 MAP kinase is necessary but not sufficient for UV activation of HIV gene expression. AB - We have examined the role of stress-activated p38 MAP kinase in regulating human immunodeficiency virus (HIV) gene expression in response to ultraviolet light (UV). We found that UV activated p38 in HeLa cells harboring stably integrated copies of an HIVcat plasmid to levels similar to those obtained by hyperosmotic shock. However, hyperosmotic shock resulted in one order of magnitude smaller increase in CAT activity than treatment with UV. The specific p38 inhibitor SB203580 significantly decreased (>80%) UV activation of HIV gene expression whereas PD98059, a specific MEK-1 inhibitor did not, suggesting that p38 is specifically involved in the HIV UV response and little to no contribution is provided by MEK-1 and the p42/p44 MAP kinase pathway. Whereas increased binding of NF-kappaB to an oligonucleotide spanning the HIV enhancer was observed after UV, as expected, this binding was not affected by SB203580. Furthermore, UV activation of HIV gene expression in cells having the cat reporter gene under control of an HIV promoter deleted of the enhancer (-69/+80) produced results indistinguishable from those using HIVcat/HeLa cells with an intact HIV promoter (-485/+80), suggesting that SB203580 acts through the basal transcription machinery. Northern blot analysis of steady-state RNA from HIVcat/HeLa cells revealed an almost complete inhibition of UV activation with SB203580 at the RNA level. Similarly, the UV response was almost completely obliterated at the CAT and RNA levels in HIVcat/HeLa cells stably transfected with a plasmid expressing a kinase-inactive mutant of p38 (isoform alpha), without affecting NF-kappaB activation, providing strong genetic evidence that p38, at least the alpha isoform, is necessary for UV activation of HIV gene expression and that NF-kappaB activation alone is insufficient. These results firmly establish p38 MAP kinase as a key modulator of HIV gene expression in response to UV that acts independently of NF-kappaB. PMID- 10529177 TI - Inhibitors of DNA strand transfer reactions catalyzed by HIV-1 reverse transcriptase. AB - The discovery and characterization of new inhibitors of HIV-1 reverse transcriptase (RT) is an important step toward understanding the mechanism of this multifunctional polymerase. We describe the identification of novel inhibitors of HIV-1 RT-catalyzed reactions utilizing a nucleic acid model system designed to mimic the essential features of DNA strand transfer reactions catalyzed by HIV-1 RT. This reaction requires the DNA polymerase and RNase H activities of RT, as well as the translocation of DNA from one template strand to another. In addition to the discovery of new inhibitors of DNA polymerase activity, two classes of inhibitors were identified that inhibit different steps of the DNA strand transfer reaction. One class of these, exemplified by actinomycin D, inhibits DNA strand transfer by interfering with the transfer of the DNA intermediate onto the acceptor template. The second class of strand transfer inhibitor, exemplified by the chlorophenylhydrazone of mesoxalic acid, was found to inhibit the ribonuclease H (RNase H) activity of HIV-1 RT under strand transfer conditions. This inhibitor is a potent and specific inhibitor of RNase H activity, which displays no inhibition of either DNA-dependent or RNA dependent DNA polymerase activity. Together, these three inhibitors block different steps reverse transcription and will be valuable in studying the mechanism of multistep reactions such as DNA strand transfer. In addition, these new inhibitors of in vitro reverse transcription point to new strategies for the intervention of retroviral DNA replication and could be useful in the development of new HIV-1 therapeutic strategies. PMID- 10529178 TI - Dimerization of the Escherichia coli biotin repressor: corepressor function in protein assembly. AB - The repressor of biotin biosynthesis binds to the biotin operator sequence to repress transcription initiation at the biotin biosynthetic operon. Site-specific binding of BirA to the biotin operator is allosterically regulated by binding of the small molecule, biotinyl-5'-adenylate (bio-5'-AMP). The operator is a 40 base pair imperfect inverted palindrome and two holorepressor monomers bind cooperatively to the two operator half-sites. Results of previous detailed analyses of binding of holoBirA to bioO indicate that site-specific DNA binding and protein dimerization are obligatorily linked in the system. In the present work equilibrium sedimentation measurements have been used to examine the assembly properties of the aporepressor and its complexes with small ligands biotin and bio-5'-AMP. Results of these measurements indicate that while the free protein and the biotin complex exhibit no tendency to self-associate, the adenylate-bound protein assembles into dimers with an equilibrium constant of 11 microM. The results suggest that one mechanism by which the adenylate promotes binding of BirA to the biotin operator is by promoting repressor dimerization. PMID- 10529179 TI - Purification and properties of the Xenopus Hat1 acetyltransferase: association with the 14-3-3 proteins in the oocyte nucleus. AB - We have purified the Xenopus histone acetyltransferase Hat1 holoenzyme from oocytes. The holoenzyme contains the catalytic subunit Hat1, the retinoblastoma associated protein RbAp48, and members of the phosphoserine binding family of 14 3-3 proteins. We have determined that the Hat1 holoenzyme specifically acetylates free histone H4 but not nucleosomal histones. RbAp48 is a phosphoprotein that contains a consensus recognition motif for the 14-3-3 proteins. The 14-3-3 proteins provide a regulatory function for the activity of many phosphoproteins. We find that the hugely abundant Hat1 holoenzyme is present in 10 000-fold excess over somatic cell levels. The holoenzyme is localized in the oocyte nucleus where acetylated histones are stored. The oocyte form of the Xenopus Hat1 holoenzyme may represent a specialized storage form of histone acetyltransferase. Following oocyte maturation and subsequent embryogenesis, the Hat1 enzyme is redistributed to the cytoplasm, where new histones are synthesized. PMID- 10529180 TI - tRNA glycylation system from Thermus thermophilus. tRNAGly identity and functional interrelation with the glycylation systems from other phylae. AB - The systems of tRNA glycylation belong to the most complex aminoacylation systems since neither the oligomeric structure of glycyl-tRNA synthetases (GlyRS) nor the discriminator bases in tRNAGly are conserved in the phylae. To better understand the structure-function relationship in glycylation systems of various origins and the functional peculiarities related to their structural divergences, the elements in tRNA conferring its glycine identity in Thermus thermophilus were characterized and compared to those of other systems. Thermophilic identity is conferred by the G1-C72, C2-G71, G3-C70, and C50-G64 pairs together with the G10, U16, C35, and C36 single residues. In contrast to most other aminoacylation systems, the discriminator base is not directly involved in identity. Transplantation of these elements in tRNAAsp and tRNAPhe converts specificity toward glycine albeit conservation of nucleotide 73. Analysis of the functional interrelation of the identity elements shows coupling in synthetase recognition of the elements from anticodon and G10 whereas those from acceptor arm are recognized independently. Despite nondirect implication in identity, the discriminator base contributes cooperatively with C36 in specificity of glycylation. The link between the structural heterogeneity and the functional divergence of the glycylation systems and the phylogenic interrelation of these systems were approached by comparing the ability of GlyRSs of various phylae to glycylate heterologous tRNAGly. Dimeric GlyRSs from mammalian and archaebacteria acylate efficiently only eukaryotic and archaebacterial tRNAGly with a discriminatory A73, whereas tetrameric Escherichia coli GlyRS acylates only eubacterial tRNAGly with a discriminatory U73. In contrast, dimeric yeast GlyRS acylates efficiently both eukaryotic and archaebacterial tRNAGly as well as peculiar prokaryotic isoacceptors. Species specificity is lost with the dimeric GlyRS from Thermus thermophilus that acylates efficiently eubacterial, archaebacterial, and eukaryotic tRNAGly. These features are discussed in the context of the evolution of the glycylation systems and the phylogenic interrelation of the organisms. PMID- 10529181 TI - Role of cysteine residues in pseudouridine synthases of different families. AB - The pseudouridine synthases catalyze the isomerization of uridine to pseudouridine in RNA molecules. An attractive mechanism was proposed based on that of thymidylate synthase, in which the thiol(ate) group of a cysteine side chain serves as the nucleophile in a Michael addition to C6 of the isomerized uridine. Such a role for cysteine in the pseudouridine synthase TruA (also named Psi synthase I) has been discredited by site-directed mutagenesis, but sequence alignments have led to the conclusion that there are four distinct "families" of pseudouridine synthases that share no statistically significant global sequence similarity. It was, therefore, necessary to probe the role of cysteine residues in pseudouridine synthases of the families that do not include TruA. We examined the enzymes RluA and TruB, which are members of different families than TruA and each other. Substitution of cysteine for amino acids with nonnucleophilic side chains did not significantly alter the catalytic activity of either pseudouridine synthase. We conclude, therefore, that neither TruB nor RluA require thiol(ate) groups to effect catalysis, excluding their participation in a Michael addition to C6 of uridine, although not eliminating that mechanism (with an alternate nucleophile) from future consideration. PMID- 10529182 TI - Sequence preference of mouse H1(0) and H1t. AB - Histone H1 proteins bind to DNA and are important in formation and maintenance of chromatin structure. Little is known about differences among variant H1 histones in their interactions with DNA. We examined the effects of histones H1(0) and H1t on thermal denaturation of several DNA species. One of the DNA molecules was a 214-base-pair fragment from the plasmid pBR322, which contains an AT-rich and a GC-rich region. Both H1(0) and H1t bound preferentially to one region of the DNA fragment, a region that is relatively GC-rich. This result indicates that histones H1(0) and H1t are not totally nonspecific but rather bind with some sequence preference to DNA. This conclusion was supported by studies of other DNA species, including two 92-base-pair fragments derived from the two regions of the 214-mer, and several synthetic homocopolymers of DNA. Data obtained with the homocopolymers suggested that the binding preference was not simple preference for GC base pairs. The binding of the two H1 variants was not identical: there appear to be differences in binding site sizes, affinities, and sequence selectivities between H1t and H1(0). PMID- 10529183 TI - Kinetic and spectroscopic characterization of the gamma-carbonic anhydrase from the methanoarchaeon Methanosarcina thermophila. AB - The zinc and cobalt forms of the prototypic gamma-carbonic anhydrase from Methanosarcina thermophila were characterized by extended X-ray absorption fine structure (EXAFS) and the kinetics were investigated using steady-state spectrophotometric and (18)O exchange equilibrium assays. EXAFS results indicate that cobalt isomorphously replaces zinc and that the metals coordinate three histidines and two or three water molecules. The efficiency of either Zn-Cam or Co-Cam for CO(2) hydration (k(cat)/K(m)) was severalfold greater than HCO(3-) dehydration at physiological pH values, a result consistent with the proposed physiological function for Cam during growth on acetate. For both Zn- and Co-Cam, the steady-state parameter k(cat) for CO(2) hydration was pH-dependent with a pK(a) of 6.5-6.8, whereas k(cat)/K(m) was dependent on two ionizations with pK(a) values of 6.7-6.9 and 8.2-8.4. The (18)O exchange assay also identified two ionizable groups in the pH profile of k(cat)/K(m) with apparent pK(a) values of 6.0 and 8.1. The steady-state parameter k(cat) (CO(2) hydration) is buffer dependent in a saturable manner at pH 8. 2, and the kinetic analysis suggested a ping-pong mechanism in which buffer is the second substrate. The calculated rate constant for intermolecular proton transfer is 3 x 10(7) M(-1) s(-1). At saturating buffer concentrations and pH 8.5, k(cat) is 2.6-fold higher in H(2)O than in D(2)O, suggesting that an intramolecular proton transfer step is at least partially rate-determining. At high pH (pH > 8), k(cat)/K(m) is not dependent on buffer and no solvent hydrogen isotope effect was observed, consistent with a zinc hydroxide mechanism. Therefore, at high pH the catalytic mechanism of Cam appears to resemble that of human CAII, despite significant structural differences in the active sites of these two unrelated enzymes. PMID- 10529184 TI - Dihydroorotate dehydrogenase B of Enterococcus faecalis. Characterization and insights into chemical mechanism. AB - Enterococcus faecalis dihydroorotate dehydrogenase B is a heterodimer of 28 and 33 kDa encoded by the pyrK and pyrDb genes. Both subunits copurify during all chromatographic steps, and, as determined by HPLC, one FMN and one FAD are bound per heterodimer. The enzyme catalyzes efficient oxidation of 4-S-NADH by orotate. Isotope effect and pH data suggest that reduction of flavin by NADH at the PyrK site is only partially rate limiting with no kinetically significant proton transfer occurring in the reductive half-reaction; therefore, a group exhibiting a pK of 5.7 +/- 0.2 represents a residue involved in binding of NADH rather than in catalysis. The reducing equivalents are shuttled between the NADH-oxidizing flavin in PyrK and the orotate-reacting flavin in PyrDb, by iron-sulfur centers through flavin semiquinones as intermediates. A solvent kinetic isotope effect of 2.5 +/- 0.2 on V is indicative of rate-limiting protonation in the oxidative half reaction and most likely reflects the interaction between the isoalloxazine N1 of the orotate-reducing flavin and Lys 168 (by analogy with L. lactis DHODase A). The oxidative half-reaction is facilitated by deprotonation of the group(s) with pK(s) of 5.8-6.3 and reflects either deprotonation of the reduced flavin or binding of orotate; this step is followed by hydride transfer to C6 and general acid-assisted protonation (pK of 9.1 +/- 0.2) at C5 of the product. PMID- 10529185 TI - Yeast protein farnesyltransferase. pKas of peptide substrates bound as zinc thiolates. AB - Protein farnesyltransferase (PFTase) is a zinc metalloenzyme that catalyzes the posttranslational alkylation of the cysteine in C-terminal -Ca(1)a(2)X sequences by a 15-carbon farnesyl residue, where C is cysteine, a(1) and a(2) are normally aliphatic amino acids, and X is an amino acid that specifies selectivity for the farnesyl moiety. Formation of a Zn(2+) thiolate in the PFTase. peptide complex was detected by the appearance of an absorbance at 236 nm (epsilon = 15 000 M(-1) cm(-1)), which was dependent on the concentration of peptide, in a UV difference spectrum in a solution of PFTase and the peptide substrate RTRCVIA. We developed a fluorescence anisotropy binding assay to measure the dissociation constants as a function of pH for peptide analogues by appending a 2',7'-difluorofluorescein to their N-terminus. The electron-withdrawing fluorine atoms allowed us to measure peptide binding down to pH 5.5 without having to correct for the changes in fluorescence intensity that accompany protonation of the fluorophore. Measurements of the pK(a)s for thiol groups in free and bound peptide indicate that peptide binding is accompanied by formation of a zinc thiolate and that binding to PFTase lowers the pK of the peptide thiol by 3 units. In similar studies with the betaY310F mutant, the pK(a) of the thiol moiety was lowered by 2 units upon binding, indicating that the hydroxyl group in the conserved tyrosine helps stabilize the bound thiolate. PMID- 10529186 TI - Iminoribitol transition state analogue inhibitors of protozoan nucleoside hydrolases. AB - Nucleoside N-ribohydrolases from protozoan parasites are targets for inhibitor design in these purine-auxotrophic organisms. Purine-specific and purine/pyrimidine-nonspecific nucleoside hydrolases have been reported. Iminoribitols that are 1-substituted with meta- and para-derivatized phenyl groups [(1S)-substituted 1, 4-dideoxy-1,4-imino-D-ribitols] are powerful inhibitors for the nonspecific nucleoside N-ribohydrolases, but are weak inhibitiors for purine-specific isozymes [Parkin, D. W., Limberg, G., Tyler, P. C., Furneaux, R. H., Chen, X.-Y., and Schramm, V. L. (1997) Biochemisty 36, 3528 3534]. Binding of these inhibitors to nonspecific nucleoside hydrolase occurs primarily via interaction with the iminoribitol, a ribooxocarbenium ion analogue of the transition state. Weaker interactions arise from hydrophobic interactions between the phenyl group and the purine/pyrimidine site. In contrast, the purine specific enzymes obtain equal catalytic potential from leaving group activation and ribooxocarbenium ion formation. Knowledge of the reaction mechanisms and transition states for these enzymes has guided the design of isozyme-specific transition state analogue inhibitors. New synthetic efforts have produced novel inhibitors that incorporate features of the leaving group hydrogen-bonding sites while retaining the iminoribitol group. These compounds provide the first transition state analogue inhibitors for purine-specific nucleoside hydrolase. The most inhibitory 1-substituted iminoribitol heterocycle is a sub-nanomolar inhibitor for the purine-specific nucleoside hydrolase from Trypanosoma brucei brucei. Novel nanomolar inhibitors are also described for the nonspecific nucleoside hydrolase from Crithidia fasciculata. The compounds reported here are the most powerful iminoribitol inhibitors yet described for the nucleoside hydrolases. PMID- 10529187 TI - Assignment of enzymatic functions to specific regions of the PLP-dependent heme protein cystathionine beta-synthase. AB - Cystathionine beta-synthase is a unique heme protein that catalyzes a pyridoxal phosphate (or PLP)-dependent beta-replacement reaction. The reaction involves the condensation of serine and homocysteine and constitutes one of the two major avenues for detoxification of homocysteine in mammals. The enzyme is allosterically regulated by S-adenosylmethionine (AdoMet). In this study, we have characterized the kinetic, spectroscopic, and ligand binding properties of a truncated catalytic core of cystathionine beta-synthase extending from residues 1 through 408 in which the C-terminal 143 residues have been deleted. This is similar to a natural variant of the protein that has been described in a homocystinuric patient in which the predicted peptide is 419 amino acids in length. Truncation leads to the formation of a dimeric enzyme in contrast to the tetrameric organization of the native enzyme. Some of the kinetic properties of the truncated enzyme are different from the full-length form, most notably, significantly higher K(m)s for the two substrates, and loss of activation by AdoMet. This is paralleled by the absence of AdoMet binding to the truncated form, whereas four AdoMet molecules bind cooperatively to the full-length tetrameric enzyme with a K(d) of 7. 4 microM. Steady-state kinetic analysis indicates that the order of substrate addition is important. Thus, preincubation of the enzyme with homocysteine leads to a 2-fold increase in V(max) relative to preincubation of the enzyme with serine. Since the intracellular concentration of serine is significantly greater than that of homocysteine, the physiological significance of this phenomenon needs to be considered. Based on ligand binding studies and homology searches with protein sequences in the database, we assign residues 68-209 as being important for PLP binding, residues 241-341 for heme binding, and residues 421-469 for AdoMet binding. PMID- 10529188 TI - Lysine 22 in UDP-N-acetylglucosamine enolpyruvyl transferase from Enterobacter cloacae is crucial for enzymatic activity and the formation of covalent adducts with the substrate phosphoenolpyruvate and the antibiotic fosfomycin. AB - UDP-N-acetylglucosamine enolpyruvyl transferase (MurA) catalyzes the first committed step in the biosynthesis of the bacterial cell wall component peptidoglycan. The enzyme is the target of the antibiotic fosfomycin. A lysine residue (K22), strictly conserved in MurAs and the structurally and mechanistically related 5-enolpyruvylshikimate 3-phosphate synthases (EPSPS), is located near the active center of the enzyme. This residue is thought to be involved directly in the binding of the substrate phosphoenolpyruvate (PEP) and also to participate in the conformational change leading to the formation of the catalytically competent enzyme complex. Using site-directed mutagenesis, we have replaced this lysine with arginine (K22R), valine (K22V), and glutamate (K22E). These mutant proteins were expressed, purified, and characterized in comparison to wild-type MurA and a previously described inactive C115S mutant protein. It was found that all three K22 mutant proteins had less than 0.5% of the wild-type activity. Using isothermal titration calorimetry, it could be shown that the binding parameters for the UDP-sugar nucleotide substrate are not affected by the mutations, except for the K22E mutant protein. Similarly, binding of PEP was found to be unaffected in the K22 mutant proteins as demonstrated by tryptophan fluorescence quench titrations. On the other hand, the level of formation of a covalent adduct with either PEP or fosfomycin with the thiol group of cysteine 115 was diminished. The propensity to form an adduct with PEP decreased in the following order: wild type >> K22R > K22V > K22E. A comparable effect was found on the formation of the inhibitory covalent adduct of MurA and the antibiotic fosfomycin. These results are discussed in terms of an involvement of lysine 22 in a conformational change of MurA. PMID- 10529190 TI - In Rhodobacter sphaeroides reaction centers, mutation of proline L209 to aromatic residues in the vicinity of a water channel alters the dynamic coupling between electron and proton transfer processes. AB - The X-ray crystallographic structure of the photosynthetic reaction center from Rhodobacter sphaeroides obtained at high resolution has revealed a number of internal water molecules (Ermler, U., Fritzsch, G., Buchanan, S. K., and Michel, H. (1994) Structure 2, 925-936; Stowell, M. H. B., McPhillips, T. M., Rees, D. C., Soltis, S. M., Abresch, E., and Feher, G. (1997) Science 276, 812-816). Some of them are organized into distinct hydrogen-bonded water chains that connect Q(B) (the terminal quinone electron acceptor of the reaction center) to the aqueous phase. To investigate the role of the water chains in the proton conduction process, proline L209, located immediately adjacent to a water chain, was mutated to the following residues: F, Y, W, E, and T. We have first analyzed the effects of the mutations on the kinetic and thermodynamic properties of the rate constants of the second electron transfer (k(AB)(2)) and of the coupled proton uptake (k(H)+) at the second flash. In all aromatic mutants, k(AB)(2) and k(H)+ are notably and concomitantly decreased compared to the wild-type, while no effect is observed in the other mutants. The temperature dependence of these rates shows activation energy values (DeltaH) similar for the proton and electron transfer processes in the wild-type and in most of the mutants, except for the L209PW and L209PF mutants. The analysis of the enthalpy factors related to the electron and proton-transfer processes in the L209PF and the L209PW mutants allows to distinguish the respective effects of the mutations for both transfer reactions. It is noteworthy that in the aromatic mutants a substantial increase of the free energies of activation is observed (DeltaG(L209PY) < DeltaG(L209PF) < DeltaG(L209PW)) for both proton and electron-transfer reactions, while in the other mutants, DeltaG is not affected. The salt concentration dependence of k(AB)(2) shows, in the L209PF and L209PW mutants, a higher screening of the protein surface potential experienced by Q(B). Our data suggest that residues F and W in position L209 increase the polarizability of the internal water molecules and polar residues by altering the organization of the hydrogen-bond network. We have also analyzed the rates of the first electron-transfer reaction (k(AB)(1)), in the 100 micros time domain. These kinetics have previously been shown to reflect protein relaxation events possibly including proton uptake events (Tiede, D. M., Vazquez, J., Cordova, J., and Marone, P. M. (1996) Biochemistry 35, 10763-10775). Interestingly, in the L209PF and L209PW mutants, k(AB)(1) is notably decreased in comparison to the wild type and the other mutants, in a similar way as k(AB)(2) and k(H)+. Our data imply that the dynamic organization of this web is tightly coupled to the electron transfer process that is kinetically limited by protonation events and/or conformational rearrangements within the protein. PMID- 10529189 TI - Mechanism of reactivation of coenzyme B12-dependent diol dehydratase by a molecular chaperone-like reactivating factor. AB - The mechanism of reactivation of diol dehydratase by its reactivating factor was investigated in vitro by using enzyme. cyanocobalamin complex as a model for inactivated holoenzyme. The factor mediated the exchange of the enzyme-bound, adenine-lacking cobalamins for free, adenine-containing cobalamins through intermediate formation of apoenzyme. The factor showed extremely low but distinct ATP-hydrolyzing activity. It formed a tight complex with apoenzyme in the presence of ADP but not at all in the presence of ATP. Incubation of the enzyme.cyanocobalamin complex with the reactivating factor in the presence of ADP brought about release of the enzyme-bound cobalamin, leaving the tight apoenzyme reactivating factor complex. Although the resulting complex was inactive even in the presence of added adenosylcobalamin, it dissociated by incubation with ATP, forming the apoenzyme, which was reconstitutable into active holoenzyme with added coenzyme. Thus, it was established that the reactivation of the inactivated holoenzyme by the factor in the presence of ATP and Mg2+ takes place in two steps: ADP-dependent cobalamin release and ATP-dependent dissociation of the apoenzyme.factor complex. ATP plays dual roles as a precursor of ADP in the first step and as an effector to change the factor into the low-affinity form for diol dehydratase. The enzyme-bound adenosylcobalamin was also susceptible to exchange with free adeninylpentylcobalamin, although to a much lesser degree. The mechanism for discrimination of adenine-containing cobalamins from adenine lacking cobalamins was explained in terms of formation equilibrium constants of the cobalamin.enzyme.reactivating factor ternary complexes. We propose that the reactivating factor is a new type of molecular chaperone that participates in reactivation of the inactivated enzymes. PMID- 10529191 TI - Electrochemical and spectroscopic investigations of the cytochrome bc1 complex from Rhodobacter capsulatus. AB - The cytochrome bc(1) complex from Rhodobacter capsulatus was investigated by protein electrochemistry and visible/IR spectroscopy. Infrared difference spectra, which represent redox-induced conformational changes of cofactors and their protein environments, show signals of the hemes, the quinone Q(i), and small conformational changes of the protein backbone. Furthermore, band features were tentatively assigned to protonated aspartic or glutamic acids involved in the redox transition of each of the b hemes, a proline in that of the [2Fe-2S] protein, and an arginine in that of cytochrome b(H). The midpoint potential of the [2Fe-2S] protein was determined for the first time at physiological temperature to be +290 mV at pH 8.7. The reduced minus oxidized difference extinction coefficients of the alpha-bands of the cytochromes were calculated as 11.5, 19, and 6.7 mM(-1) cm(-1) for cytochromes c(1), b(H), and b(L), respectively. A novel method has been developed to quantify protonation reactions of the complex during the redox reactions of its cofactors by evaluation of the buffer signals in the midinfrared region. Values will be discussed in relation to the pH dependence of the midpoint potentials. PMID- 10529192 TI - Probing intramolecular orientations in rhodopsin and metarhodopsin II by polarized infrared difference spectroscopy. AB - The light-induced conformational changes of rhodopsin, which lead to the formation of the G-protein activating metarhodopsin II intermediate, are studied by polarized attenuated total reflectance infrared difference spectroscopy. Orientations of protein groups as well as the retinylidene chromophore were calculated from the linear dichroism of infrared difference bands. These bands correspond to changes in the vibrational modes of individual molecular groups that are structurally active during receptor activation, i.e., during the rhodopsin to metarhodopsin II transition. The orientation of the transition dipole moments of bands previously assigned to the carboxyl (C=O) groups of Asp83 and Glu113 has been determined. The orientation of specific groups in the retinylidene chromophore has been inferred from the dichroism of the bands associated with the polyene C-C, C=C, and hydrogen-out-of-plane vibrations. Interestingly, the use of polarized infrared light reveals several difference bands in the rhodopsin to metarhodopsin II difference spectrum which were previously undetected, e.g., at 1736 and 939 cm(-1). The latter is tentatively assigned to the hydrogen-out-of-plane mode of the HC(11)=C(12)H segment of the chromophore. Our data suggest a significant change in orientation of this group in the late phase of rhodopsin activation. On the basis of available site directed mutagenesis data, bands at 1406, 1583, and 1736 cm(-1) are tentatively assigned to Glu134. The main features in the amide regions in the dichroic difference spectrum are discussed in terms of a slight reorientation of helical segments upon receptor activation. PMID- 10529193 TI - Location of the iron-sulfur clusters FA and FB in photosystem I: an electron paramagnetic resonance study of spin relaxation enhancement of P700+. AB - Photosystem I (PS I) mediates electron-transfer from plastocyanin to ferredoxin via a photochemically active chlorophyll dimer (P700), a monomeric chlorophyll electron acceptor (A0), a phylloquinone (A1), and three [4Fe-4S] clusters (FX/A/B). The sequence of electron-transfer events between the iron-sulfur cluster, FX, and ferredoxin is presently unclear. Owing to the presence of a 2 fold symmetry in the PsaC protein to which the iron-sulfur clusters F(A) and F(B) are bound, the spatial arrangement of these cofactors with respect to the C2-axis of symmetry in PS I is uncertain as well. An unequivocal determination of the spatial arrangement of the iron-sulfur clusters FA and FB within the protein is necessary to unravel the complete electron-transport chain in PS I. In the present study, we generate EPR signals from charge-separated spin pairs (P700+ FredX/A/B) in PS I and characterize them by progressive microwave power saturation measurements to determine the arrangement of the iron-sulfur clusters FX/A/B relative to P700. The microwave power at half saturation (P1/2) of P700+ is greater when both FA and FB are reduced in untreated PS I than when only FA is reduced in mercury-treated PS I. The experimental P1/2 values are compared to values calculated by using P700-FA/B crystallographic distances and assuming that either FA or FB is closer to P700+. On the basis of this comparison of experimental and theoretical values of spin relaxation enhancement effects on P700+ in P700+ [4Fe-4S]- charge-separated pairs, we find that iron-sulfur cluster FA is in closer proximity to P700 than the FB cluster. PMID- 10529194 TI - Dynamics of energy conversion in reaction center core complexes of the green sulfur bacterium Prosthecochloris aestuarii at low temperature. AB - Excited-state and electron-transfer dynamics at cryogenic temperature in reaction center core (RCC) complexes of the photosynthetic green sulfur bacterium Prosthecochloris aestuarii were studied by means of time-resolved absorption spectroscopy, using selective excitaton of bacteriochlorophyll (BChl) a and of chlorophyll (Chl) a 670. The results indicate that the BChls a of the RCC complex form an excitonically coupled system. Relaxation of the excitation energy within the ensemble of BChl a molecules occurred within 2 ps. A time constant of about 25 ps was ascribed to charge separation. Absorption changes in the 670 nm region, where Chl a 670 absorbs, were fairly complicated. They showed various time constants and were dependent on the wavelength of excitation and they did not lead to a simple picture of the electron acceptor reaction. Energy transfer from Chl a 670 to BChl a occurred with a time constant of 1.5 ps. However, upon excitation of Chl a 670 the amount of oxidized primary electron donor, P840(+), formed relative to that of excited BChl a was considerably larger than upon direct excitation of BChl a. This indicates the existence of an alternative pathway for charge separation which does not involve excited BChl a. PMID- 10529195 TI - Site-directed mutagenesis of firefly luciferase active site amino acids: a proposed model for bioluminescence color. AB - Under physiological conditions firefly luciferase catalyzes the highly efficient emission of yellow-green light from the substrates luciferin, Mg-ATP, and oxygen. In nature, bioluminescence emission by beetle luciferases is observed in colors ranging from green (approximately 530 nm) to red (approximately 635 nm), yet all known luciferases use the same luciferin substrate. In an earlier report [Branchini, B. R., Magyar, R. M., Murtiashaw, M. H., Anderson, S. M., and Zimmer, M. (1998) Biochemistry 37, 15311-15319], we described the effects of mutations at His245 on luciferase activity. In the context of molecular modeling results, we proposed that His245 is located at the luciferase active site. We noted too that the H245 mutants displayed red-shifted bioluminescent emission spectra. We report here the construction and purification of additional His245 mutants, as well as mutants at residues Lys529 and Thr343, all of which are stringently conserved in the beetle luciferase sequences. Analysis of specific activity and steady-state kinetic constants suggested that these residues are involved in luciferase catalysis and the productive binding of substrates. Bioluminescence emission spectroscopy studies indicated that point mutations at His245 and Thr343 produced luciferases that emitted light over the color range from green to red. The results of mutational and biochemical studies with luciferase reported here have enabled us to propose speculative mechanisms for color determination in firefly bioluminescence. An essential role for Thr343, the participation of His245 and Arg218, and the involvement of bound AMP are indicated. PMID- 10529196 TI - The sterol carrier protein-2 amino terminus: a membrane interaction domain. AB - Sterol carrier protein-2 (SCP2) is a small, 123 amino acid, protein postulated to play a role in intracellular transport and metabolism of lipids such as cholesterol, phospholipids, and branched chain fatty acids. While it is thought that interaction of SCP2 with membranes is necessary for lipid transfer, evidence for this possibility and identification of a membrane interaction domain within SCP2 has remained elusive. As shown herein with circular dichroism and a direct binding assay, SCP2 bound to small unilamellar vesicle (SUV) membranes to undergo significant alteration in secondary structure. The SCP2 amphipathic N-terminal 32 amino acids, comprised of two alpha-helical segments, were postulated to represent a putative phospholipid interaction site. This hypothesis was tested with a series of SCP2 N-terminal peptides, circular dichroism, and direct binding studies. The SCP2 N-terminal peptide (1-32)SCP2, primarily random coil in aqueous buffer, adopted alpha-helical structure upon interaction with membranes. The induction of alpha-helical structure in the peptide was maximal when the membranes contained a high mole percent of negatively charged phospholipid and of cholesterol. While deletion of the second alpha-helical segment within this peptide had no effect on formation of the first alpha-helix, it significantly weakened the peptide interaction with membranes. Substitution of Leu(20) with Glu(20) in the N-terminal peptide disrupted the alpha-helix structure and greatly weakened the peptide interaction with membranes. Finally, deletion of the first nine nonhelical amino acids had no effect either on formation of alpha-helix or on peptide binding to membranes. N-Terminal peptide (1-32)SCP2 competed with SCP2 for binding to SUV. These data were consistent with the N-terminus of SCP2 providing a membrane interaction domain that preferentially bound to membranes rich in anionic phospholipid and cholesterol. PMID- 10529197 TI - Resveratrol preferentially inhibits protein kinase C-catalyzed phosphorylation of a cofactor-independent, arginine-rich protein substrate by a novel mechanism. AB - Resveratrol, a polyphenolic natural product abundantly present in grape skins, is a candidate cancer chemopreventive agent that antagonizes each stage of carcinogenesis and inhibits protein kinase C (PKC), a key mediator of tumor promotion. While resveratrol has been shown to antagonize both isolated and cellular forms of PKC, the weak inhibitory potency observed against isolated PKC cannot account for the reported efficacy of the polyphenol against PKC in cells. In this report, we analyze the mechanism of PKC inhibition by resveratrol. Our results indicate that resveratrol has a broad range of inhibitory potencies against purified PKC that depend on the nature of the substrate and the cofactor dependence of the phosphotransferase reaction. Resveratrol weakly inhibited the Ca2+/phosphatidylserine-stimulated activity of a purified rat brain PKC isozyme mixture (IC(50) = 90 microM) by competition with ATP (K(i) = 55 microM). Consistent with the kinetic evidence for a catalytic domain-directed mechanism, resveratrol inhibited the lipid-dependent activity of PKC isozymes with divergent regulatory domains similarly, and it was even more effective in inhibiting a cofactor-independent catalytic domain fragment (CDF) of PKC generated by limited proteolysis. This suggested that regulatory features of PKC might impede resveratrol inhibition of the enzyme. To explore this, we examined the effects of resveratrol on PKC-catalyzed phosphorylation of the cofactor-independent substrate protamine sulfate, which is a polybasic protein that activates PKC by a novel mechanism. Resveratrol potently inhibited protamine sulfate phosphorylation (IC(50) = 10 microM) by a mechanism that entailed antagonism of the activation of PKC by protamine sulfate and did not involve competition with either substrate. On the basis of the presence of PKC isozymes at subcellular sites rich in polybasic proteins, it has been proposed that certain endogenous polybasic PKC substrates may activate PKC in cells by the same mechanism as protamine sulfate. Our results suggest that antagonism by resveratrol of the phosphorylation of cellular PKC substrates that resemble protamine sulfate in their interactions with PKC may contribute to the efficacy of resveratrol against PKC in cells. PMID- 10529198 TI - The N-terminal half of Cdc25 is essential for processing glucose signaling in Saccharomyces cerevisiae. AB - Saccharomyces cerevisiae Cdc25 is the prototype Ras GDP/GTP exchange protein. Its C-terminal catalytic domain was found to be highly conserved in the homologues p140(ras-GRF) and Sos. The regulatory domains in each Ras exchanger mediate the signals arriving from upstream elements such as tyrosine kinases for Sos, or Ca2+ and G proteins for p140.(Ras-GRF) In this study, we show that the N-terminal half (NTH) of S. cerevisiae Cdc25, as well as the C-terminal 37 amino acids, is essential for processing the elevation of cAMP in response to glucose. The mammalian p140(ras-GRF) catalytic domain (CGRF) restores glucose signaling in S. cerevisiae only if tethered between the N-terminal half (NTH) of S. cerevisiae Cdc25 and the C-terminal 37 amino acids. The glucose-induced transient elevation in cAMP is nullified or severely hampered by the deletion of domains within the NTH of Cdc25. These deletions, however, do not modify the intrinsic GDP/GTP exchange activity of mutant proteins as compared to native Cdc25. We also show that 7 Ser to Ala mutations at the cAMP-dependent protein kinase putative phosphorylation sites within the NTH of Cdc25 eliminate the descending portion of the glucose response curve, responsible for signal termination. These findings support a dual role of the NTH of Cdc25 in both enabling the glucose signal and being responsible for its attenuation. PMID- 10529199 TI - Heterodimerization of a functional GABAB receptor is mediated by parallel coiled coil alpha-helices. AB - A detailed understanding of GABAB receptor assembly is an important issue in view of its role as attractive target for treatment of epilepsy, anxiety, depression, cognitive defects, and nociceptive disorders. Heteromerization of GABAB-R1 and GABAB-R2 subunits is a prerequisite for the formation of a functional GABAB receptor. Each individual subunit contains one stretch of approximately 30 amino acid residues within its intracellular C-terminal domain that mediates heteromer formation. To investigate the mechanism of the GABAB-R1/GABAB-R2 interaction and to assess the subunit stoichiometry of the complex, recombinant polypeptide chain fragments containing the heteromerization site were produced by heterologous gene expression in Escherichia coli. When mixed in equimolar amounts, these peptides preferentially formed parallel coiled-coil heterodimers under physiological buffer conditions. This demonstrates that the short C-terminal regions are sufficient to determine the specificity of interaction between GABAB receptor subunits. In contrast, isolated GABAB-R1 peptides folded into relatively unstable homodimers, whereas GABAB-R2 peptides were largely unstructured. Together with the data reported in the literature, the results presented here indicate that the functional GABAB receptor is a heterodimer assembled by parallel coiled-coil alpha-helices. PMID- 10529200 TI - Transducer-binding and transducer-mutations modulate photoactive-site deprotonation in sensory rhodopsin I. AB - Sensory rhodopsin I (SRI) is a seven-transmembrane helix retinylidene protein that mediates color-sensitive phototaxis responses through its bound transducer HtrI in the archaeon Halobacterium salinarum. Deprotonation of the Schiff base attachment site of the chromophore accompanies formation of the SRI signaling state, S(373). We measured the rate of laser flash-induced S(373) formation in the presence and absence of HtrI, and the effects of mutations in SRI or HtrI on the kinetics of this process. In the absence of HtrI, deprotonation occurs rapidly (halftime 10 micros) if the proton acceptor Asp76 is ionized (pK(a) = approximately 7), and only very slowly (halftime > 10 ms) when Asp76 is protonated. Transducer-binding, although it increases the pK(a) of Asp76 so that it is protonated throughout the range of pH studied, results in a first order, pH independent rate of S(373) formation of approximately 300 micros. Therefore, the complexation of HtrI facilitates the proton-transfer reaction, increasing the rate approximately 50-fold at pH6. Arrhenius analysis shows that HtrI-binding accelerates the reaction primarily by an entropic effect, suggesting HtrI constrains the SRI molecule in the complex. Function-perturbing mutations in SRI and HtrI also alter the rate of S(373) formation and the lambda(max) of the parent state as assessed by laser flash-induced kinetic difference spectroscopy, and shifts to longer wavelength are correlated with slower deprotonation. The data indicate that HtrI affects electrostatic interactions of the protonated Schiff base and not only receives the signal from SRI but also optimizes the photochemical reaction process for SRI signaling. PMID- 10529201 TI - A singular state of membrane lipids at cell growth temperatures. AB - Cells adjust their membrane lipid composition when they adapt to grow at different temperatures. The consequences of this adjustment for membrane properties and functions are not well understood. Our report shows that the temperature dependence of the diffusion of a probe molecule in multilayers formed from total lipid extracts of E. coli has an anomalous maximum at a temperature corresponding to the growth temperature of each bacterial preparation (25, 29, and 32 degrees C). This increase in the lateral diffusion coefficient, D, is characteristic of membrane lipids in a critical state, for which large fluctuations of molecular area in the plane of the bilayer are expected. Therefore, changes in lipid composition may be due to a requirement that cells maintain their membranes in a state where molecular interactions and reaction rates are readily modulated by small, local perturbations of membrane organization. PMID- 10529202 TI - Ca2+-induced increased lipid packing and domain formation in submitochondrial particles. A possible early step in the mechanism of Ca2+-stimulated generation of reactive oxygen species by the respiratory chain. AB - Ca2+ and P(i) accumulation by mitochondria triggers a number of alterations leading to nonspecific increase in inner membrane permeability [Kowaltowski, A. J., et al. (1996) J. Biol. Chem. 271, 2929-2934]. The molecular nature of the membrane perturbation that precedes oxidative damage is still unknown. EPR spectra of spin probes incorporated in submitochondrial particles (SMP) and in model membranes suggest that Ca(2+)-cardiolipin (CL) complexation plays an important role. Ca(2+)-induced lipid domain formation was detected in SMP but not in mitoplasts, in SMP extracted lipids, or in CL-containing liposomes. The results were interpreted in terms of Ca2+ sequestration of CL tightly bound to membrane proteins, in particular the ADP-ATP carrier, and formation of CL enriched strongly immobilized clusters in lipid shells next to boundary lipid. The in-plane lipid and protein rearrangement is suggested to cause increased reactive oxygen species production in succinate-supplemented, antimycin A poisoned SMP, favoring the formation of carbon-centered radicals, detected by EPR spin trapping. Removal of tightly bound CL is also proposed to cause protein aggregation, facilitating intermolecular thiol oxidation. Lipid peroxidation was also monitored by the disappearance of the nitroxide EPR spectrum. The decay was faster for nitroxides in a more hydrophobic environment, and was inhibited by butylated hydroxytoluene, by EGTA, or by substituting Mg2+ for Ca2+. In addition, Ca2+ caused an increase in permeability, evidenced by the release of carboxyfluorescein from respiring SMP. The results strongly support Ca2+ binding to CL as one of the early steps in the molecular mechanism of Ca(2+)-induced nonspecific inner mitochondrial membrane permeabilization. PMID- 10529203 TI - Differential dynamic behavior of actin filaments containing tightly-bound Ca2+ or Mg2+ in the presence of myosin heads actively hydrolyzing ATP. AB - We have investigated how Ca2+ or Mg2+ bound at the high-affinity cation binding site in F-actin modulates the dynamic response of these filaments to ATP hydrolysis by attached myosin head fragments (S1). Rotational motions of the filaments were monitored using steady-state phosphorescence emission anisotropy of the triplet probe erythrosin-5-iodoacetamide covalently attached to cysteine 374 of actin. The anisotropy of filaments containing only Ca2+ increased from 0.080 to 0.137 upon binding S1 in a rigor complex and decreased to 0.065 in the presence of ATP, indicating that S1 induced additional rotational motions in the filament during ATP hydrolysis. The comparable anisotropy values for Mg(2+) containing filaments were 0.067, 0.137, and 0.065, indicating that S1 hydrolysis did not induce measurable rotational motions in these filaments. Phalloidin, a fungal toxin which stabilizes F-actin and increases its rigidity, increased the anisotropy of F-actin containing either Ca2+ or Mg2+ but not the anisotropy of the 1:1 S1-actin complexes of these filaments. Mg(2+)-containing filaments with phalloidin bound also displayed increased rotational motions during S1 ATP hydrolysis. A strong positive correlation between the phosphorescence anisotropy of F-actin under specific conditions and the extent of the rotational motions induced by S1 during ATP hydrolysis suggested that the long axis torsional rigidity of F-actin plays a crucial role in modulating the dynamic response of the filaments to ATP hydrolysis by S1. Cooperative responses of F-actin to dynamic perturbations induced by S1 during ATP hydrolysis may thus be physically mediated by the torsional rigidity of the filament. PMID- 10529204 TI - Altered regulatory function of two familial hypertrophic cardiomyopathy troponin T mutants. AB - Mutations in the gene encoding human cardiac troponin T can cause familial hypertrophic cardiomyopathy, a disease that is characterized by ventricular hypertrophy and sudden, premature death. Troponin T is the tropomyosin-binding subunit of troponin required for thin filament regulation of contraction. One mutation, a change in the intron 15 splice donor site, results in two truncated forms of troponin T [Thierfelder et al. (1994) Cell 77, 701-712]. In one form, the mRNA skips exon 16 that encodes the C-terminal 14 amino acids; in the other, seven novel residues replace the exon 15- and 16-encoded C-terminal 28 amino acids. The two troponin T cDNAs were expressed in Escherichia coli for functional analysis. Both C-terminal deletion mutants formed a complex with cardiac troponin C and troponin I that exhibited the same concentration dependence as wild-type for regulation of the actomyosin MgATPase. However, both mutants showed severely reduced activation of the regulated actomyosin in the presence of Ca2+, though the inhibition in the absence of Ca2+ and the Ca(2+)-dependence of activation were not altered. The C-terminal deletions reduce the effectiveness of Ca(2+) troponin to switch the thin filament from the "off" to the "on" state. Both mutant troponin Ts have reduced affinity for troponin I; the shorter mutant is at least 6-fold weaker than wild-type. The low level of activation of the ATPase would be consistent with reduced contractile performance, and the results suggest reduced troponin I affinity may be the molecular basis for the disease. PMID- 10529205 TI - Structural and functional characterization of EMF10, a heterodimeric disintegrin from Eristocophis macmahoni venom that selectively inhibits alpha 5 beta 1 integrin. AB - Alpha5beta1, a major fibronectin receptor, is a widely distributed integrin that is essential for cell growth and organ development. Here, we describe a novel heterodimeric disintegrin named EMF10, isolated from the Eristocophis macmahoni venom, that is an extremely potent and selective inhibitor of alpha5beta1. EMF10 inhibited adhesion of cells expressing alpha5beta1 to fibronectin (IC(50) = 1-4 nM) and caused expression of a ligand-induced binding site (LIBS) on the beta1 subunit of alpha5beta1 integrin. It partially inhibited adhesion of cells expressing alphaIIbbeta3, alphavbeta3, and alpha4beta1 to appropriate ligands only at concentration higher than 500 nM. Guinea pig megakaryocytes expressing alpha5beta1 adhered to immobilized EMF10 and showed extensive spreading and cytoskeletal mobilization. As determined by electrospray mass spectrometry, EMF10 is composed of two species with molecular masses of 14 575 and 14 949 Da, respectively. EMF10 is a heterodimer containing two subunits: EMF10A (Mr 7544 Da) and EMF10B (Mr 7405 and 7032 Da) linked covalently by S-S bonds. Subunit B showed heterogeneity and may be present as EMF10B1 (Mr 7032) and EMF10B2 (Mr 7405). In putative hairpin loops, EMF10A and EMF10B contained CKKGRGDNLNDYC and CWPAMGDWNDDYC motifs, respectively. The reduced and alkylated subunit B of EMF10 inhibited adhesion of K562 cells to fibronectin in a dose-dependent, saturable manner with IC(50) of 3 microM. The synthetic, cyclic CKKGRGDNLNDYC and CWPAMGDWNDDYC peptides expressed their inhibitory activity in the same system with IC(50) of 100 microM. We propose that alpha5beta1 recognition of EMF10 is associated with the MGDW motif located in a putative hairpin loop of the B subunit and that the expression of activity may also depend on the RGDN motif in the subunit A and on the C-termini of both subunits. PMID- 10529206 TI - Critical residues influence the affinity and selectivity of alpha-conotoxin MI for nicotinic acetylcholine receptors. AB - The mammalian skeletal muscle acetylcholine receptor contains two nonequivalent acetylcholine binding sites, one each at the alpha/delta and alpha/gamma subunit interfaces. Alpha-Conotoxin MI, a 14-amino acid competitive antagonist, binds at both interfaces but has approximately 10(4) higher affinity for the alpha/delta site. We performed an "alanine walk" to identify the residues in alpha-MI that contribute to this selective interaction with the alpha/delta site. Electrophysiological measurements with Xenopus oocytes expressing normal receptors or receptors lacking either the gamma or delta subunit were made to assay toxin-receptor interaction. Alanine substitutions in most amino acid positions had only modest effects on toxin potency at either binding site. However, substitutions in two positions, proline-6 and tyrosine-12, dramatically reduced toxin potency at the high-affinity alpha/delta site while having comparatively little effect on low-affinity alpha/gamma binding. When tyrosine-12 was replaced by alanine, the toxin's selectivity for the high-affinity site (relative to that for the low-affinity site) was reduced from 45,000- to 30-fold. A series of additional amino acid substitutions in this position showed that increasing side chain size/hydrophobicity increases toxin potency at the alpha/delta site without affecting alpha/gamma binding. In contrast, when tyrosine-12 is diiodinated, toxin binding is nearly irreversible at the alpha/delta site but also increases by approximately 500-fold at the alpha/gamma site. The effects of position 12 substitutions are accounted for almost entirely by changes in the rate of toxin dissociation from the high-affinity alpha/delta binding site. PMID- 10529207 TI - Rapid in vitro conformational changes of the catalytic site of PKC alpha assessed by FIM-1 fluorescence. AB - To study the activation process of protein kinase C (PKCalpha), we used a fluorescent probe, FIM-1, a bis-indolylmaleimide derivative, which binds to the ATP-binding site on the catalytic domain [Chen, C. S., and Poenie, M. (1993) J. Biol. Chem. 268, 15812]. This enabled us to directly observe the microenvironment of the ATP-binding site in vitro during the activation process. The FIM-1 binding affinity for PKCalpha (EC(50) between 6 and 10 nM) was affected neither by PKCalpha activating conditions nor by enzyme proteolysis. The fluorescence yield of the PKCalpha-FIM-1 complex depended on the PKCalpha activation state. This fluorescence yield was decreased upon proteolysis, which allowed us to study the rate of PKC proteolysis by mu-calpain and its modification by cofactors. Two binding sites were also observed for Ca2+ on the partially activated PKCalpha. After phorbol ester (TPA) application, PKC activation was characterized by biexponential kinetics, including a rapid phase completed within 5 min and a slow phase lasting at least 30 min, which reflected several activation steps. Two different binding sites for TPA were revealed on membrane-associated PKCalpha (EC(50) = 31 +/- 12 and 580 +/- 170 nM), and their modulation by phosphatidylserine and Ca2+ was characterized. The high-affinity TPA binding site was highly conserved, even on the soluble enzyme. Our study shows that binding of low concentrations of TPA triggers conformational changes in the soluble PKCalpha, which affect the microenvironment of its catalytic domain. PMID- 10529208 TI - Facilitation of the cellular uptake of a triplex-forming oligonucleotide by novel polyamine analogues: structure-activity relationships. AB - The inefficient uptake of oligodeoxynucleotides, including that of TFO, through the cell membrane is a limiting factor in developing gene therapy approaches for cancer and other diseases. To develop a new strategy for oligonucleotide delivery into the nucleus, we synthesized a series of novel polyamine analogues and examined their effects on the uptake of a 37-mer [32P]-labeled TFO, targeted to the promoter region of c-myc oncogene. We used MCF-7 breast cancer cells to investigate the efficacy of polyamines on the internalization of the TFO. The uptake of TFO was enhanced by complexing it with several unsubstituted polyamine analogues at 0. 1-5 microM concentrations, with up to 6-fold increase in TFO uptake in the presence of a hexamine, 1,21-diamino-4,9,13, 18-tetraazahenicosane (H2N(CH2)(3)NH(CH2)(4)NH(CH2)(3)NH(CH2)(4)NH(CH2)(3)NH(2) or 3-4-3-4-3). TFO uptake increased with the cationicity of the polyamines; however, bis(ethyl) substitution and structural features of the methylene bridging region had significant effects on TFO uptake. The majority of labeled TFO was recovered from the nuclear fraction containing genomic DNA. Electrophoretic mobility shift assay revealed enhanced binding of TFO to a target duplex containing promoter region sequence of c-myc oncogene. Treatment of MCF-7 cells with the TFO complexed with 0.5 microM 3-4-3-4-3 suppressed c-myc mRNA level by 65%, as determined by Northern blot analysis. These data indicate a novel approach to deliver oligodeoxynucleotides to the cell nucleus, and suppress the expression of target genes, and provide new insights into the mechanism of oligonucleotide transport in living cells. PMID- 10529209 TI - A Watson-Crick base-pair-disrupting methyl group (m1A9) is sufficient for cloverleaf folding of human mitochondrial tRNALys. AB - We have previously shown by chemical and enzymatic structure probing that, opposite to the native human mitochondrial tRNA(Lys), the corresponding in vitro transcript does not fold into the expected tRNA-specific cloverleaf structure. This RNA folds into a bulged hairpin, including an extended amino acid acceptor stem, an extra large loop instead of the T-stem and loop, and an anticodon-like domain. Hence, one or several of the six modified nucleotides present in the native tRNA are required and responsible for its cloverleaf structure. Phylogenetic comparisons as well as structural analysis of variant transcripts had pointed to m(1)A9 as the most likely important modified nucleotide in the folding process. Here we describe the synthesis of a chimeric tRNA(Lys) with m(1)A9 as the sole modified base and its structural analysis by chemical and enzymatic probing. Comparison of this structure to that of the unmodified RNA, the fully modified native tRNA, and a variant designed to mimic the effect of m(1)A9 demonstrates that the chimeric RNA folds indeed into a cloverleaf structure that resembles that of the native tRNA. Thus, due to Watson-Crick base pair disruption, a single methyl group is sufficient to induce the cloverleaf folding of this unusual tRNA. This is the first direct evidence of the role of a modified nucleotide in RNA folding. PMID- 10529211 TI - Energetics of assembling an artificial heterodimer with an alpha/beta motif: cleaved versus uncleaved Escherichia coli thioredoxin. AB - We have studied the folding/binding process between the N- and C-fragments (1-73, 74-108) of oxidized Escherichia coli thioredoxin (Trx) to compare the energetics between the cleaved and uncleaved Trx. Sedimentation equilibrium analysis in 0.1 M potassium phosphate, pH 5.7, shows (i) the strong and weak self-association of the N- and C-fragments, respectively, (ii) a heterodimer with a small dissociation constant (K(d)) ca. 100 nM, and (iii) monomeric Trx. To avoid self association, measurements were carried out in 10 mM potassium phosphate, pH 5.7. Far-UV CD spectra of the fragments at variable temperature show an isodichroic point at 208 nm and a non-cooperative cold induced disordering transition without concentration dependence. Deconvolution of these spectra indicates the presence of residual structure. Titration of the N-fragment with an excess of C-fragment indicates a 1:1 stoichiometric complex with an apparent K(d) ca. 49 nM. Analysis of this complex by CD and hydrogen exchange/2D-NMR (Tasayco and Chao (1995) Proteins: Struct., Funct., Genet. 22, 41-44) spectroscopy indicates the reassembly of the alpha/beta motif of Trx. GnHCl induced unfolding measurements give DeltaG(0) values of 9.5 +/- 0.2 and 10.0 +/- 0.4 kcal/mol at 20 degrees C for the uncleaved and cleaved Trx, respectively. The far-UV CD melting curve of uncleaved Trx indicates an intriguing non-cooperative upward baseline trend. CCA analysis of these spectra indicates the presence of a native-like folded intermediate. A three-state thermodynamic analysis of the thermal transition curves gives a total DeltaH(0) of unfolding of 121 +/- 4 kcal/mol at the T(m) (88 degrees C), while the two-state analysis for cleaved Trx gives 122 +/- 6 kcal/mol at 88 degrees C. Analysis of the chemical and thermal unfolding of both proteins indicates a value of ca. 1 M for the apparent effective concentration (C(eff)) of cleaved Trx. PMID- 10529210 TI - Effect of flap modifications on human FEN1 cleavage. AB - The flap endonuclease, FEN1, plays a critical role in DNA replication and repair. Human FEN1 exhibits both a 5' to 3' exonucleolytic and a structure-specific endonucleolytic activity. On primer-template substrates containing an unannealed 5'-tail, or flap structure, FEN1 employs a unique mechanism to cleave at the point of annealing, releasing the 5'-tail intact. FEN1 appears to track along the full length of the flap from the 5'-end to the point of cleavage. Substrates containing structural modifications to the flap have been used to explore the mechanism of tracking. To determine whether the nuclease must recognize a succession of nucleotides on the flap, chemical linkers were used to replace an interior nucleotide. The nuclease could readily traverse this site. The footprint of the nuclease at the time of cleavage does not extend beyond 25 nucleotides on the flap. Eleven-nucleotide branches attached to the flap beyond the footprinted region do not prevent cleavage. Single- or double-thymine dimers also allow cleavage. cis-Platinum adducts outside the protected region are moderately inhibitory. Platinum-modified branch structures are completely inert to cleavage. These results show that some flap modifications can prevent or inhibit tracking, but the tracking mechanism tolerates a variety of flap modifications. FEN1 has a flexible loop structure through which the flap has been proposed to thread. However, efficient cleavage of branched structures is inconsistent with threading the flap through a hole in the protein. PMID- 10529212 TI - Apparent radii of the native, stable intermediates and unfolded conformers of the alpha-subunit of tryptophan synthase from E. coli, a TIM barrel protein. AB - The urea-induced equilibrium unfolding of the alpha-subunit of tryptophan synthase (alphaTS) from Escherichia coli can be described by a four-state model, N right harpoon over left harpoon I1 right harpoon over left harpoon I2 right harpoon over left harpoon U, involving two highly populated intermediates, I1 and I2 [Gualfetti, P. J., Bilsel, O., and Matthews, C. R. (1999) Protein Sci. 8, 1623 1635]. To extend the physical characterization of these stable forms, the apparent radius was measured by several techniques. Size-exclusion chromatography (SEC), analytical ultracentrifugation (UC), and dynamic light scattering (DLS) experiments yield an apparent Stokes radius, R(s), of approximately 24 A for the native state of alphaTS. The small-angle X-ray scattering (SAXS) experiment yields a radius of gyration, R(g), of 19.1 A, consistent with the value predicted from the X-ray structure and the Stokes radius. As the equilibrium is shifted to favor I1 at approximately 3.2 M and I2 at 5.0 M urea, SEC and UC show that R(s) increases from approximately 38 to approximately 52 A. Measurements of the radius by DLS and SAXS between 2 and 4.5 M urea were complicated by the self-association of the I1 species at the relatively high concentrations required by those techniques. Above 6 M urea, SEC and UC reveal that R(s) increases linearly with increasing urea concentration to approximately 54 A at 8 M urea. The measurements of R(s) by DLS and R(g) by SAXS are sufficiently imprecise that both values appear to be identical for the I2 and U states and, considering the errors, are in good agreement with the results from SEC and UC. Thermodynamic parameters extracted from the SEC data for the N right harpoon over left harpoon I1 and I1 right harpoon over left harpoon I2 transitions agree with those from the optical data, showing that this technique accurately monitors a part of the equilibrium model. The lack of sensitivity to the I2 right harpoon over left harpoon U transition, beyond a simple swelling of both species with increasing urea concentration, implies that the Stokes radii for the I2 and U states are not distinguishable. Surprisingly, the hydrophobic core known to stabilize I2 at 5.0 M urea [Saab-Rincon, G., Gualfetti, P. J., and Matthews, C. R. (1996) Biochemistry 35, 1988-1994] develops without a significant contraction of the polypeptide, i.e., beyond that experienced by the unfolded form at decreasing urea concentrations. Kratky plots of the SAXS data, however, reveal that I2, similar to N and I1, has a globular structure while U has a more random coil-like form. By contrast, the formation of substantial secondary structure and the burial of aromatic side chains in I1 and, eventually, N are accompanied by substantial decreases in their Stokes radii and, presumably, the size of their respective conformational ensembles. PMID- 10529214 TI - Phosphorylation of ribosomal protein L18 is required for its folding and binding to 5S rRNA. AB - Ribosomal protein L18 from Bacillus stearothermophilus (bL18) includes a previously unreported phosphoserine residue. The folded conformation of the protein is stabilized by the dianionic form of the phosphate group of that residue. In the absence of Mg2+, the pK(a) of the phosphate group is so high that the protein is not fully folded at pH 7. In the presence of Mg2+, its pK(a) drops significantly, and consequently the native conformation of bL18 becomes stable at pH 7 and the protein is able to bind to 5S rRNA. Dephosphorylated bL18 does not bind to 5S rRNA at neutral pH. PMID- 10529213 TI - Buried, charged, non-ion-paired aspartic acid 76 contributes favorably to the conformational stability of ribonuclease T1. AB - The side-chain carboxyl of Asp 76 in ribonuclease T1 (RNase T1) is buried, charged, non-ion-paired, and forms three good intramolecular hydrogen bonds (2.63, 2.69, and 2.89 A) and a 2.66 A hydrogen bond to a buried, conserved water molecule. When Asp 76 was replaced by Asn, Ser, and Ala, the conformational stability of the protein decreased by 3.1, 3.2, and 3.7 kcal/mol, respectively. The stability was measured as a function of pH for wild-type RNase T1 and the D76N mutant and showed that the pH dependence below pH 3 was almost entirely due to Asp 76. The pK of Asp 76 is 0.5 in the native state and 3.7 in the denatured state. Thus, the hydrogen bonding of the carboxyl group of Asp 76 contributes more than half of the net stability of RNase T1 at pH 7. In addition, the charged carboxyl of Asp 76 stabilizes structure in the denatured states of RNase T1 that is not present in D76N, D76S, and D76A. PMID- 10529215 TI - Toward tailoring the specificity of the S1 pocket of subtilisin B. lentus: chemical modification of mutant enzymes as a strategy for removing specificity limitations. AB - In both protein chemistry studies and organic synthesis applications, it is desirable to have available a toolbox of inexpensive proteases with high selectivity and diverse substrate preferences. Toward this goal, we have generated a series of chemically modified mutant enzymes (CMMs) of subtilisin B. lentus (SBL) possessing expanded S(1) pocket specificity. Wild-type SBL shows a marked preference for substrates with large hydrophobic P(1) residues, such as the large Phe P(1) residue of the standard suc-AAPF-pNA substrate. To confer more universal P(1) specificity on S(1), a strategy of chemical modification in combination with site-directed mutagenesis was applied. For example, WT-SBL does not readily accept small uncharged P(1) residues such as the -CH(3) side chain of alanine. Accordingly, with a view to creating a S(1) pocket that would be of reduced volume providing a better fit for small P(1) side chains, a large cyclohexyl group was introduced by the CMM approach at position S166C with the aim of partially filling up the S(1) pocket. The S166C-S-CH(2)-c-C(6)H(11) CMM thus created showed a 2-fold improvement in k(cat)/K(M) with the suc-AAPA-pNA substrate and a 51-fold improvement in suc-AAPA-pNA/suc-AAPF-pNA selectivity relative to WT-SBL. Furthermore, WT-SBL does not readily accept positively or negatively charged P(1) residues. Therefore, to improve SBL's specificity toward positively and negatively charged P(1) residues, we applied the CMM methodology to introduce complementary negatively and positively charged groups, respectively, at position S166C in S(1). A series of mono-, di-, and trinegatively charged CMMs were generated and all showed improved k(cat)/K(M)s with the positively charged P(1) residue containing substrate, suc-AAPR-pNA. Furthermore, virtually arithmetic improvements in k(cat)/K(M) were exhibited with increasing number of negative charges on the S166C-R side chain. These increases culminated in a 9-fold improvement in k(cat)/K(M) for the suc-AAPR-pNA substrate and a 61-fold improvement in suc-AAPR-pNA/suc-AAPF-pNA selectivity compared to WT SBL for the trinegatively charged S166C-S-CH(2)CH(2)C(COO(-))(3) CMM. Conversely, the positively charged S166C-S-CH(2)CH(2)NH(3)(+) CMM generated showed a 19-fold improvement in k(cat)/K(M) for the suc-AAPE-pNA substrate and a 54-fold improvement in suc-AAPE-pNA/suc-AAPF-pNA selectivity relative to WT-SBL. PMID- 10529216 TI - Significance of metal ions in galactose-1-phosphate uridylyltransferase: an essential structural zinc and a nonessential structural iron. AB - Galactose-1-phosphate uridylyltransferase (GalT) catalyzes the reversible transformation of UDP-glucose and galactose-1-phosphate (Gal-1-P) into UDP galactose and glucose-1-phosphate (Glc-1-P) by a double displacement mechanism, with the intermediate formation of a covalent uridylyl-enzyme (UMP-enzyme). GalT is a metalloenzyme containing 1.2 mol of zinc and 0.7 mol of iron/mol of subunits [Ruzicka, F. J., Wedekind, J. E., Kim, J., Rayment, I., and Frey, P. A. (1995) Biochemistry 34, 5610-5617]. The zinc site lies 8 A from His 166 in active site, and the iron site lies 30 A from the active site [Wedekind,J. E., Frey, P. A., & Rayment, I. (1995) Biochemistry 34, 11049-11061]. Zinc is coordinated in tetrahedral geometry by Cys 52, Cys 55, His 115, and His 164. His 164 is part of the highly conserved active-site triad His 164-Pro 165-His 166, in which His 166 is the nucleophilic catalyst. Iron is coordinated in square pyramidal geometry with His 296, His 298, and Glu 182 in bidentate coordination providing the base ligands and His 281 providing the axial ligand. In the present study, site directed mutagenesis, kinetic, and metal analysis studies show that C52S-, C55S-, and H164N-GalT are 3000-, 600-, and 10000-fold less active than wild-type. None of the variants formed the UMP-enzyme in detectable amounts upon reaction with UDP-Glc in the absence of Gal-1-P. Their zinc content was very low, and the zinc + iron content was about 50% of that for wild-type GalT. Mutation of His 115 to Asn 115 resulted in decreased activity to 2.9% of wild-type, with retention of zinc and iron. In contrast to the zinc-binding site, Glu 182 in the iron site is not important for enzymatic activity. The variant E182A-GalT displayed about half the activity of wild-type GalT, and all of the active sites underwent uridylylation to the UMP-enzyme, similar to wild-type GalT, upon reaction with UDP-Glc. Metal analysis showed that while E182A-GalT contained 0.9 equiv of zinc/subunit, it contained no iron. The residual zinc can be removed by dialysis with 1,10-phenanthroline, with the loss in activity being proportional to the amount of residual zinc. It is concluded that the presence of zinc is essential for maintaining GalT function, whereas the presence of iron is not essential. PMID- 10529217 TI - Reversible S-nitrosation and inhibition of HIV-1 protease. AB - Nitric oxide (*NO) is a short-lived free radical with many functions including vasoregulation, synaptic plasticity, and immune modulation and has recently been associated with AIDS pathology. Various pathophysiological conditions, such as viral infection, trigger inducible nitric oxide synthase (iNOS) to synthesize NO in the cell. NO-derived species can react with thiols of proteins and form nitrosothiol adducts. HIV-1 protease (HIV-PR) contains two cysteine residues, Cys67 and Cys95, which are believed to serve a regulatory function. We have found that HIV-PR is inactivated by nitric oxide produced in vitro by NO donors and by iNOS. Sodium nitroprusside inhibited HIV-PR by 70%, and S-nitroso-N acetylpenicillamine completely inhibited the enzyme. Furthermore, iNOS generated sufficient NO to inhibit HIV-PR activity by almost 90%. This inactivation was reversed by the addition of reducing agents. Treatment of HIV-PR with NO donors and ritonavir (a competitive peptide inhibitor) indicates that NO exerts its effect through a site independent of the active site of HIV-PR. Using electrospray ionization mass spectrometry, we found that NO forms S-nitrosothiols on Cys67 and Cys95 of HIV-PR which directly correlate with a loss of activity. These data indicate that NO may suppress HIV-1 replication by directly inhibiting HIV-PR. PMID- 10529218 TI - Conserved aromatic residues of the C-terminus of human butyrylcholinesterase mediate the association of tetramers. AB - Human butyrylcholinesterase (BChE) in serum is composed predominantly of tetramers. The tetramerization domain of each subunit is contained within 40 C terminal residues. To identify key residues within this domain participating in tetramer stabilization, the interaction between C-terminal 46 residue peptides was quantitated in the yeast two-hybrid system. The wild-type peptide interacted strongly with another wild-type peptide in the yeast two-hybrid system. The C571A mutant peptides interacted to a similar degree as the wild-type. However, the mutant in which seven conserved aromatic residues (Trp 543, Phe 547, Trp 550, Tyr 553, Trp 557, Phe 561, and Tyr 564) and C571 were altered to alanines showed only 12% of the interaction seen with the wild-type peptide. The seven mutations (aromatics-off) were incorporated into the complete BChE molecule, with or without the C571A mutation, and expressed in 293T and CHO-K1 cells. Expression of wild-type BChE in these cell lines yielded 10% tetramers. The aromatics-off mutant formed dimers and monomers but no tetramers. The aromatics-off/C571A mutant yielded only monomers. Addition of poly-L-proline to culture medium, or coexpression with the N-terminus of COLQ including the proline-rich attachment domain (Q(N)PRAD), increased the amount of tetrameric wild-type BChE from 10 to 70%, but had no effect on the G534stop (lacking 41 C-terminal residues) and the aromatics-off mutants. Recombinant BChE produced by coexpression with Q(N)PRAD was purified by column chromatography. The purified tetramers contained the FLAG tagged Q(N)PRAD peptide. These observations suggest that the stabilization of BChE tetramers is mediated through the interaction of the seven conserved aromatic residues and that poly-L-proline and PRAD act through these aromatic residues to induce tetramerization. PMID- 10529219 TI - Assessment of roles of surface histidyl residues in the molecular basis of the Bohr effect and of beta 143 histidine in the binding of 2,3-bisphosphoglycerate in human normal adult hemoglobin. AB - Site-directed mutagenesis has been used to construct two mutant recombinant hemoglobins (rHbs), rHb(betaH116Q) and rHb(betaH143S). Purified rHbs were used to assign the C2 proton resonances of beta116His and beta143His and to resolve the ambiguous assignments made over the past years. In the present work, we have identified the C2 proton resonances of two surface histidyl residues of the beta chain, beta116His and beta143His, in both the carbonmonoxy and deoxy forms, by comparing the proton nuclear magnetic resonance (NMR) spectra of human normal adult hemoglobin (Hb A) with those of rHbs. Current assignments plus other previous assignments complete the assignments for all 24 surface histidyl residues of human normal adult hemoglobin. The individual pK values of 24 histidyl residues of Hb A were also measured in deuterium oxide (D(2)O) in 0.1 M N-(2-hydroxyethyl)piperazine-N'-2-ethanesulfonic acid (HEPES) buffer in the presence of 0.1 M chloride at 29 degrees C by monitoring the shifts of the C2 proton resonances of the histidyl residues as a function of pH. Among those surface histidyl residues, beta146His has the biggest contribution to the alkaline Bohr effect (63% at pH 7.4), and beta143His has the biggest contribution to the acid Bohr effect (71% at pH 5.1). alpha20His, alpha112His, and beta117His have essentially no contribution; alpha50His, alpha72His, alpha89His, beta97His, and beta116His have moderate positive contributions; and beta2His and beta77His have a moderate negative contribution to the Bohr effect. The sum of the contributions from 24 surface histidyl residues accounted for 86% of the alkaline Bohr effect at pH 7.4 and about 55% of the acid Bohr effect at pH 5.1. Although beta143His is located in the binding site for 2,3-bisphosphoglycerate (2,3-BPG) according to the crystal structure of deoxy-Hb A complexed with 2, 3-BPG, beta143His is not essential for the binding of 2,3-BPG in the neutral pH range according to the proton NMR and oxygen affinity studies presented here. With the accurately measured and assigned individual pK values for all surface histidyl residues, it is now possible to evaluate the Bohr effect microscopically for novel recombinant Hbs with important functional properties, such as low oxygen affinity and high cooperativity. The present study further confirms the importance of a global electrostatic network in regulating the Bohr effect of the hemoglobin molecule. PMID- 10529221 TI - Protein folding as a diffusional process. AB - A protein chain must move relative to the solvent molecules and explore many conformations when it folds from the extended unfolded state to the compact native state. Experimental and theoretical approaches suggest that diffusional processes in fact contribute to the kinetics of protein folding. We describe here how variations of the solvent viscosity can be employed to uncover the diffusional contributions to a folding reaction and assess the use of transition state theory and Kramers' rate theory for the analysis of protein folding reactions. PMID- 10529220 TI - Recombinant hemoglobin(alpha 29leucine --> phenylalanine, alpha 96valine --> tryptophan, beta 108asparagine --> lysine) exhibits low oxygen affinity and high cooperativity combined with resistance to autoxidation. AB - Using our hemoglobin expression system in Escherichia coli, we have constructed three recombinant hemoglobins (rHbs) with amino acid substitutions located in the alpha(1)beta(1) and alpha(1)beta(2) subunit interfaces and in the distal heme pocket of the alpha-chain: rHb(alphaV96W, betaN108K), rHb(alphaL29F, alphaV96W, betaN108K), and rHb(alphaL29F). rHb(alphaV96W, betaN108K) exhibits low oxygen affinity and high cooperativity and also ease of autoxidation of the heme iron atoms from the Fe2+ state to the Fe3+ state. It has been reported by Olson and co workers [Carver et al., (1992) J. Biol. Chem. 267, 14443-14450; Brantley et al. (1993) J. Biol. Chem. 268, 6995-7010] that a mutation at position 29 (B10, helix notation), e.g. , Leu --> Phe, can inhibit the autoxidation of the heme iron of myoglobin. We have introduced such a mutation into our rHb having low oxygen affinity and high cooperativity. This triply mutated rHb(alphaL29F, alphaV96W, betaN108K) is stabilized against autoxidation and azide-induced oxidation compared to the double mutant, rHb(alphaV96W, betaN108K), but still exhibits low oxygen affinity and good cooperativity. According to electron paramagnetic resonance results, the oxidized form of the triple mutant shows a high ratio of an anionic form of bishistidine hemichrome. Previous reports have suggested that this form does not have water present at the distal heme pocket. (1)H nuclear magnetic resonance spectra of the triple mutant in the ferric state also exhibit spectral features characteristic of hemichrome-type signals. We have carried out a series of biochemical measurements to characterize these three interesting rHbs and to compare them to human normal adult hemoglobin. These results provide new insights into the structure-function relationship of hemoglobin with amino acid substitutions in the alpha(1)beta(1) and alpha(1)beta(2) interfaces and in the heme pockets. PMID- 10529222 TI - A low-redox potential heme in the dinuclear center of bacterial nitric oxide reductase: implications for the evolution of energy-conserving heme-copper oxidases. AB - Bacterial nitric oxide reductase (NOR) catalyzes the two-electron reduction of nitric oxide to nitrous oxide. It is a highly diverged member of the superfamily of heme-copper oxidases. The main feature by which NOR is distinguished from the heme-copper oxidases is the elemental composition of the active site, a dinuclear center comprised of heme b(3) and non-heme iron (Fe(B)). The visible region electronic absorption spectrum of reduced NOR exhibits a maximum at 551 nm with a distinct shoulder at 560 nm; these are attributed to Fe(II) heme c (E(m) = 310 mV) and Fe(II) heme b (E(m) = 345 mV), respectively. The electronic absorption spectrum of oxidized NOR exhibits a characteristic shoulder around 595 nm that exhibits complex behavior in equilibrium redox titrations. The first phase of reduction is characterized by an apparent shift of the shoulder to 604 nm and a decrease in intensity. This is due to reduction of Fe(B) (E(m) = 320 mV), while the subsequent bleaching of the 604 nm band represents reduction of heme b(3) (E(m) = 60 mV). This separation of redox potentials (>200 mV) allows the enzyme to be poised in the three-electron reduced state for detailed spectroscopic examination of the Fe(III) heme b(3) center. The low midpoint potential of heme b(3) represents a thermodynamic barrier to the complete (two-electron) reduction of the dinuclear center. This may avoid formation of a stable Fe(II) heme b(3)-NO species during turnover, which may be an inhibited state of the enzyme. It would also appear that the evolution of significant oxygen reducing activity by heme copper oxidases was not simply a matter of the substitution of copper for non heme iron in the dinuclear center. Changes in the protein environment that modulate the midpoint redox potential of heme b(3) to facilitate both complete reduction of the dinuclear center (a prerequisite for oxygen binding) and rapid heme-heme electron transfer were also necessary. PMID- 10529223 TI - RACK1, a protein kinase C anchoring protein, coordinates the binding of activated protein kinase C and select pleckstrin homology domains in vitro. AB - The pleckstrin homology (PH) domain, identified in numerous signaling proteins including the beta-adrenergic receptor kinase (betaARK), was found to bind to various phospholipids as well as the beta subunit of heterotrimeric G proteins (Gbeta) [Touhara, K., et al. (1994) J. Biol. Chem. 269, 10217-10220]. Several PH domain-containing proteins are also substrates of protein kinase C (PKC). Because RACK1, an anchoring protein for activated PKC, is homologous to Gbeta (both contain seven repeats of the WD-40 motif), we determined (i) whether a direct interaction between various PH domains and RACK1 occurs and (ii) the effect of PKC on this interaction. We found that recombinant PH domains of several proteins exhibited differential binding to RACK1. Activated PKC and the PH domain of beta spectrin or dynamin-1 concomitantly bound to RACK1. Although PH domains bind acidic phospholipids, the interaction between various PH domains and RACK1 was not dependent on the phospholipid activators of PKC, phosphatidylserine and 1, 2 diacylglycerol. Binding of these PH domains to RACK1 was also not affected by either inositol 1,4,5-triphosphate (IP(3)) or phosphatidylinositol 4,5 bisphosphate (PIP(2)). Our in vitro data suggest that RACK1 binds selective PH domains, and that PKC regulates this interaction. We propose that, in vivo, RACK1 may colocalize the kinase with its PH domain-containing substrates. PMID- 10529224 TI - Interdomain interactions regulate GDP release from heterotrimeric G proteins. AB - The role of interdomain contact sites in basal GDP release from heterotrimeric G proteins is unknown. G(alpha)(o) and G(alpha)(i1) display a 5-fold difference in the rate of GDP dissociation with half-times of 2.3 +/- 0.2 and 10.4 +/- 1.3 min, respectively. To identify molecular determinants of the GDP release rate, we evaluated the rate of binding of the fluorescent guanine nucleotide 2'(3')-O-(N methyl-3'-anthraniloyl)guanosine 5'-O-(3-thiotriphosphate) (mGTPgammaS) to chimers of G(alpha)(o) and G(alpha)(i1). Although no one region of the G protein determined the GDP dissociation rate, when the C-terminal 123 amino acids in G(alpha)(i1) were replaced with those of G(alpha)(o), the GDP release rate increased 3.3-fold compared to that of wild-type G(alpha)(i1). Within the C terminal portion, modification of four amino acids in a coil between beta4 and the alpha3 helix resulted in GDP release kinetics identical to those of wild-type G(alpha)(o). Based on the G(alpha)(i1)-GDP crystal structure of this region, Leu(232) appeared to form a hydrophobic contact with Arg(144) of the helical domain. The role of this interaction was confirmed by G(alpha)(i1) L232Q and G(alpha)(i1) R144A which displayed 2-5-fold faster GDP release rates compared to wild-type G(alpha)(i1) (t(1/2) 4.7 and 1.5 min, respectively), suggesting that interdomain bridging contacts partially determine the basal rate of GDP release from heterotrimeric G proteins. PMID- 10529226 TI - Roles of active site aromatic residues in catalysis by ketosteroid isomerase from Pseudomonas putida biotype B. AB - The aromatic residues Phe-54, Phe-82, and Trp-116 in the hydrophobic substrate binding pocket of Delta(5)-3-ketosteroid isomerase from Pseudomonas putida biotype B have been characterized in their roles in steroid binding and catalysis. Kinetic and equilibrium binding analyses were carried out for the mutant enzymes with the substitutions Phe-54 --> Ala or Leu, Phe-82 --> Ala or Leu, and Trp-116 --> Ala, Phe, or Tyr. The removal of their bulky, aromatic side chains at any of these three positions results in reduced k(cat), particularly when Phe-82 or Trp-116 is replaced by Ala. The results are consistent with the binding interactions of the aromatic residues with the bound steroid contributing to catalysis. All the mutations except the F82A mutation increase K(m); the F82A mutation decreases K(m) by ca. 3-fold, suggesting a possibility that the phenyl ring at position 82 might be unfavorable for substrate binding. The K(D) values for d-equilenin, an intermediate analogue, suggest that a space-filling hydrophobic side chain at position 54, a phenyl ring at position 82, and a nonpolar aromatic or small side chain at position 116 might be favorable for binding the reaction intermediate. In contrast to the increased K(D) for equilenin, the enzymes with any substitutions at positions 54 and 116 display a decreased K(D) for 19-nortestosterone, a product analogue, indicating that Phe-54 and Trp-116 might be unfavorable for product binding. The crystal structure of F82A determined to 2.1-A resolution reveals that Phe-82 is important for maintaining the active site geometry. Taken together, our results demonstrate that Phe-54, Phe-82, and Trp-116 contribute differentially to the stabilization of steroid species including substrate, intermediate, and product. PMID- 10529225 TI - Restricting the mobility of Gs alpha: impact on receptor and effector coupling. AB - The alpha-subunit of the stimulatory G protein, Gs, has been shown to dissociate from the plasma membrane into the cytosol following activation by G protein coupled receptors (GPCR) in some experimental systems. This dissociation may involve depalmitoylation of an amino-terminal cysteine residue. However, the functional significance of this dissociation is not known. To investigate the functional consequence of Gs alpha dissociation, we constructed a membrane tethered Gs alpha (tetGs alpha), expressed it in Sf9 insect cells, and examined its ability to couple with the beta(2) adrenoceptor and to activate adenylyl cyclase. Compared to wild-type Gs alpha, tetGs alpha coupled much more efficiently to the beta 2 adrenoceptor and the D1 dopamine receptor as determined by agonist-stimulated GTP gamma S binding and GTPase activity. The high coupling efficiency was abolished when Gs )alpha was proteolytically cleaved from the membrane tether. The membrane tether did not prevent the coupling of tetGS alpha to adenylyl cyclase. These results demonstrate that regulating the mobility of Gs alpha relative to the plasma membrane, through fatty acylation or perhaps interactions with cytoskeletal proteins, could have a significant impact on receptor-G protein coupling. Furthermore, by enabling the use of more direct measures of receptor-G protein coupling (GTPase activity, GTP gamma S binding), tetGS alpha can facilitate the study for receptor-G protein interactions. PMID- 10529227 TI - Control of luminescence decay and flavin binding by the LuxA carboxyl-terminal regions in chimeric bacterial luciferases. AB - Bacterial luciferases (LuxAB) can be readily classed as slow or fast decay luciferases based on their rates of luminescence decay in a single turnover assay. Luciferases from Vibrio harveyi and Xenorhabdus (Photorhabdus) luminescens have slow decay rates, and those from the Photobacterium genus, such as P. (Vibrio) fischeri, P. phosphoreum, and P. leiognathi, have rapid decay rates. By generation of an X. luminescens-based chimeric luciferase with a 67 amino acid substitution from P. phosphoreum LuxA in the central region of the LuxA subunit, the "slow" X. luminescens luciferase was converted into a chimeric luciferase, LuxA(1)B, with a significantly more rapid decay rate. Two other chimeras with P. phosphoreum sequences substituted closer to the carboxyl terminal of LuxA, LuxA(2)B and LuxA(3)B, retained the characteristic slow decay rates of X. luminescens luciferase but had weaker interactions with both reduced and oxidized flavins, implicating the carboxyl-terminal regions in flavin binding. The dependence of the luminescence decay on concentration and type of fatty aldehyde indicated that the decay rate of "fast" luciferases arose due to a high dissociation constant (K(a)) for aldehyde (A) coupled with the rapid decay of the resultant aldehyde-free complex via a dark pathway. The decay rate of luminescence (k(T)) was related to the decanal concentration by the equation: k(T) = (k(L)A + k(D)K(a))/(K(a) + A), showing that the rate constant for luminescence decay is equal to the decay rate via the dark- (k(D)) and light emitting (k(L)) pathways at low and high aldehyde concentrations, respectively. These results strongly implicate the central region in LuxA(1)B as critical in differentiating between "slow" and "fast" luciferases and show that this distinction is primarily due to differences in aldehyde affinity and in the decomposition of the luciferase-flavin-oxygen intermediate. PMID- 10529228 TI - The structural mechanism for half-the-sites reactivity in an enzyme, thymidylate synthase, involves a relay of changes between subunits. AB - Thymidylate synthase (TS), a half-the-sites reactive enzyme, catalyzes the final step in the de novo biosynthesis of deoxythymidine monophosphate, dTMP, required for DNA replication. The cocrystal structure of TS from Pneumocystis carinii (PcTS), a new drug target for an important pathogen, with its substrate, deoxyuridine monophosphate (dUMP), and a cofactor mimic, CB3717, was determined. The structure, solved at 2.6 A resolution, shows an asymmetric dimer with two molecules of the substrate dUMP bound yet only one molecule of cofactor analogue bound. The structural evidence reveals that upon binding cofactor analogue and forming a covalent bond from the nucleophilic cysteine to the substrate, dUMP, at one active site, PcTS undergoes a conformational change that renders the opposite monomer incapable of forming a covalent bond or binding a molecule of cofactor analogue. The communication pathway between the two active sites is evident, allowing a structural definition of the basis of half-the-sites reactivity for thymidylate synthase and providing an example of such a mechanism for other half the-sites reactive enzymes. PMID- 10529229 TI - Selective recognition of mannose by the human eosinophil Charcot-Leyden crystal protein (galectin-10): a crystallographic study at 1.8 A resolution. AB - The role(s) of the eosinophil Charcot-Leyden crystal (CLC) protein in eosinophil or basophil function or associated inflammatory processes is yet to be established. Although the CLC protein has been reported to exhibit weak lysophospholipase activity, it shows virtually no sequence homology to any known member of this family of enzymes. The X-ray crystal structure of the CLC protein is very similar to the structure of the galectins, members of a beta-galactoside specific animal lectin family, including a partially conserved galectin carbohydrate recognition domain (CRD). In the absence of any known natural carbohydrate ligand for this protein, the functional role of the CLC protein (galectin-10) has remained speculative. Here we describe structural studies on the carbohydrate binding properties of the CLC protein and report the first structure of a carbohydrate in complex with the protein. Interestingly, the CLC protein demonstrates no affinity for beta-galactosides and binds mannose in a manner very different from those of other related galectins that have been shown to bind lactosamine. The partial conservation of residues involved in carbohydrate binding led to significant changes in the topology and chemical nature of the CRD, and has implications for carbohydrate recognition by the CLC protein in vivo and its functional role in the biology of inflammation. PMID- 10529230 TI - Conformational changes in human hepatitis C virus NS3 protease upon binding of product-based inhibitors. AB - One of the most promising approaches to anti-hepatitis C virus drug discovery is the development of inhibitors of the virally encoded protease NS3. This chymotrypsin-like serine protease is essential for the maturation of the viral polyprotein, and processing requires complex formation between NS3 and its cofactor NS4A. Recently, we reported on the discovery of potent cleavage product derived inhibitors [Ingallinella et al. (1998) Biochemistry 37, 8906-8914]. Here we study the interaction of these inhibitors with NS3 and the NS3/cofactor complex. Inhibitors bind NS3 according to an induced-fit mechanism. In the absence of cofactor different binding modes are apparent, while in the presence of cofactor all inhibitors show the same binding mode with a small rearrangement in the NS3 structure, as suggested by circular dichroism spectroscopy. These data are consistent with the hypothesis that NS4A complexation induces an NS3 structure that is already (but not entirely) preorganized for substrate binding not only for what concerns the S' site, as already suggested, but also for the S site. Inhibitor binding to the NS3/cofactor complex induces the stabilization of the enzyme structure as highlighted by limited proteolysis experiments. We envisage that this may occur through stabilization of the individual N-terminal and C-terminal domains where the cofactor and inhibitor, respectively, bind and subsequent tightening of the interdomain interaction in the ternary complex. PMID- 10529231 TI - Structure comparison between oxidized and reduced plastocyanin from a fern, Dryopteris crassirhizoma. AB - The X-ray crystal structures of oxidized and reduced plastocyanin obtained from the fern Dryopteris crassirhizoma have been determined at 1.7 and 1.8 A resolution, respectively. The fern plastocyanin is unique in the longer main chain composed of 102 amino acid residues and in the unusual pH dependence due to the pi-pi stacking interaction around the copper site [Kohzuma, T., et al. (1999) J. Biol. Chem. 274, 11817-11823]. Here we report the structural comparison between the fern plastocyanin and other plastocyanins from cyanobacteria, green algae, and other higher plants, together with the structural changes of fern plastocyanin upon reduction. Glu59 hydrogen bonds to the OH of Tyr83, which is thought to be a possible conduit for electrons, in the oxidized state. However, it moves away from Tyr83 upon reduction like poplar plastocyanin. PMID- 10529232 TI - Molecular dynamics simulations of human prion protein: importance of correct treatment of electrostatic interactions. AB - Molecular dynamics simulations have been used to investigate the dynamical and structural behavior of a homology model of human prion protein HuPrP(90-230) generated from the NMR structure of the Syrian hamster prion protein ShPrP(90 231) and of ShPrP(<90-231) itself. These PrPs have a large number of charged residues on the protein surface. At the simulation pH 7, HuPrP(90-230) has a net charge of -1 eu from 15 positively and 14 negatively charged residues. Simulations for both PrPs, using the AMBER94 force field in a periodic box model with explicit water molecules, showed high sensitivity to the correct treatment of the electrostatic interactions. Highly unstable behavior of the structured region of the PrPs (127-230) was found using the truncation method, and stable trajectories could be achieved only by including all the long-range electrostatic interactions using the particle mesh Ewald (PME) method. The instability using the truncation method could not be reduced by adding sodium and chloride ions nor by replacing some of the sodium ions with calcium ions. The PME simulations showed, in accordance with NMR experiments with ShPrP and mouse PrP, a flexibly disordered N-terminal part, PrP(90-126), and a structured C-terminal part, PrP(127-230), which includes three alpha-helices and a short antiparallel beta strand. The simulations showed some tendency for the highly conserved hydrophobic segment PrP(112-131) to adopt an alpha-helical conformation and for helix C to split at residues 212-213, a known disease-associated mutation site (Q212P). Three highly occupied salt bridges could be identified (E146/D144<-->R208, R164<- >D178, and R156<-->E196) which appear to be important for the stability of PrP by linking the stable main structured core (helices B and C) with the more flexible structured part (helix A and strands A and B). Two of these salt bridges involve disease-associated mutations (R208H and D178N). Decreased PrP stability shown by protein unfolding experiments on mutants of these residues and guanidinium chloride or temperature-induced unfolding studies indicating reduced stability at low pH are consistent with stabilization by salt bridges. The fact that electrostatic interactions, in general, and salt bridges, in particular, appear to play an important role in PrP stability has implications for PrP structure and stability at different pHs it may encounter physiologically during normal or abnormal recycling from the pH neutral membrane surface into endosomes or lysomes (acidic pHs) or in NMR experiments (5.2 for ShPrP and 4.5 for mouse PrP). PMID- 10529233 TI - Photoaffinity labeling of diphtheria toxin fragment A with 8-azidoadenosyl nicotinamide adenine dinucleotide. AB - Diphtheria toxin fragment A (DT-A) is an important enzyme in the class of mono(ADP-ribosyl)transferases. To identify peptides and amino acid residues which form the NAD(+) binding site of DT-A using a photoaffinity approach, the photoprobes nicotinamide 8-azidoadenine dinucleotide (8-N(3)-NAD) and nicotinamide 2-azidoadenine dinucleotide (2-N(3)-NAD) were synthesized. Binding studies gave an IC(50) of 2.5 microM for 8-N(3)-NAD and 5.0 microM for 2-N(3) NAD. Irradiation of DT-A and low concentrations of [alpha-(32)P]-8-N(3)-NAD with short-wavelength UV light resulted in rapid covalent incorporation of the photoprobe into the protein. The photoincorporation was shown to be specific for the active site with a stoichiometry of photoincorporation of 75-80%. After proteolytic digestion of photolabeled DT-A, derivatized peptides were isolated using immobilized boronate affinity chromatography followed by reversed phase HPLC. Radiolabeled peptides originating from two regions of the protein were identified. Chymotryptic digestion produced labeled peptides corresponding to His(21)-Gln(32) and Lys(33)-Phe(53). Lys-C digestion gave overlapping peptides Ser(11)-Lys(33) and Ser(40)-Lys(59). Tyr(27) was identified as the site of photoinsertion within the peptide His(21)-Gln(32) on the basis of the absence of PTH-Tyr at the predicted cycle during sequence analysis and by the lack of predicted chymotryptic cleavage at Tyr(27). Within the second modified peptide Ser(40)-Lys(59), Trp(50) is the most probable site of modification. Identification of Tyr(27) as a site of photoinsertion is in agreement with its placement in the NAD binding site of the X-ray structure of the proenzyme DT-NAD complex [Bell, C. E., and Eisenberg, D. (1996) Biochemistry 35, 1137]. Trp(50) is far from the adenine ring in the crystallographic model; however, site-directed mutagenesis studies suggest that Trp(50) is a major determinant of NAD binding affinity [Wilson, B. A., Blanke, S. R., Reich, K. A., and Collier, R. J. (1994) J. Biol. Chem. 269, 23296-23301]. PMID- 10529234 TI - Both ATP sites of human P-glycoprotein are essential but not symmetric. AB - Human P-glycoprotein (P-gp) is a cell surface drug efflux pump that contains two nucleotide binding domains (NBDs). Mutations were made in each of the Walker B consensus motifs of the NBDs at positions D555N and D1200N, thought to be involved in Mg(2+) binding. Although the mutant and wild-type P-gps were expressed equivalently at the cell surface and bound the drug analogue [(125)I]iodoarylazidoprazosin ([(125)I]IAAP) comparably, neither of the mutant proteins was able to transport fluorescent substrates nor had detectable basal nor drug-stimulated ATPase activities. The wild-type and D1200N P-gps were labeled comparably with [alpha-(32)P]-8-azido-ATP at a subsaturating concentration of 2.5 microM, whereas labeling of the D555N mutant was severely impaired. Mild trypsin digestion, to cleave the protein into two halves, demonstrated that the N-half of the wild-type and D1200N proteins was labeled preferentially with [alpha-(32)P]-8-azido-ATP. [alpha-(32)P]-8-Azido-ATP labeling at 4 degrees C was inhibited in a concentration-dependent manner by ATP with half maximal inhibition at approximately 10-20 microM for the P-gp-D1200N mutant and wild-type P-gp. A chimeric protein containing two N-half NBDs was found to be functional for transport and was also asymmetric with respect to [alpha-(32)P]-8 azido-ATP labeling, suggesting that the context of the ATP site rather than its exact sequence is an important determinant for ATP binding. By use of [alpha (32)P]-8-azido-ATP and vanadate trapping, it was determined that the C-half of wild-type P-gp was labeled preferentially under hydrolysis conditions; however, the N-half was still capable of being labeled with [alpha-(32)P]-8-azido-ATP. Neither mutant was labeled under vanadate trapping conditions, indicating loss of ATP hydrolysis activity in the mutants. In confirmation of the lack of ATP hydrolysis, no inhibition of [(125)I]IAAP labeling was observed in the mutants in the presence of vanadate. Taken together, these data suggest that the two NBDs are asymmetric and intimately linked and that a conformational change in the protein may occur upon ATP hydrolysis. Furthermore, these data are consistent with a model in which binding of ATP to one site affects ATP hydrolysis at the second site. PMID- 10529235 TI - The secondary multidrug transporter LmrP contains multiple drug interaction sites. AB - The secondary multidrug transporter LmrP of Lactococcus lactis mediates the efflux of Hoechst 33342 from the cytoplasmic leaflet of the membrane. Kinetic analysis of Hoechst 33342 transport in inside-out membrane vesicles of L. lactis showed that the LmrP-mediated H(+)/Hoechst 33342 antiport reaction obeyed Michaelis-Menten kinetics, with a low apparent affinity constant of 0.63 microM Hoechst 33342 (= 0.5 mmol Hoechst 33342/mol phospholipid). Several drugs significantly inhibited LmrP-mediated Hoechst 33342 transport through a direct interaction with the protein rather than through dissipation of the proton motive force or reduction of the membrane partitioning of Hoechst 33342. The characterization of the mechanism of inhibition of LmrP-mediated Hoechst 33342 transport indicated competitive inhibition by quinine and verapamil, noncompetitive inhibition by nicardipin and vinblastin, and uncompetitive inhibition by TPP(+). The three types of inhibition of LmrP-mediated Hoechst 33342 transport in inside-out membrane vesicles indicate for the first time the presence of multiple drug interaction sites in a secondary multidrug transporter. PMID- 10529236 TI - Rhodamine 110-linked amino acids and peptides as substrates to measure caspase activity upon apoptosis induction in intact cells. AB - Caspases (cysteine aspartate-specific proteases) are a structurally related group of cysteine proteases that cleave peptide bonds following specific recognition sequences. They play a central role in activating apoptosis of vertebrate cells. To measure apoptosis induced by various stimuli and at an early apoptotic stage, caspases are an ideal target. This is especially the case when apoptotic cells have to be analyzed ex vivo before phagocytes remove them. A new and sensitive caspase assay is based on a substrate that contains only aspartate residues linked to rhodamine 110. With this and similar substrates, we are able to detect intracellular caspase activation by flow cytometry after apoptosis induction in intact hematopoetic cell lines. PMID- 10529237 TI - Mutations in the rod outer segment membrane guanylate cyclase in a cone-rod dystrophy cause defects in calcium signaling. AB - Rod outer segment guanylate cyclase 1 (ROS-GC1) is a member of the subfamily of Ca(2+)-regulated membrane guanylate cyclases; and it is pivotal for vertebrate phototransduction. Two opposing regulatory modes control the activity of ROS-GC1. At nanomolar concentrations of Ca(2+), ROS-GC1 is activated by Ca(2+)-binding proteins named guanylate cyclase activating proteins (GCAPs). However, at micromolar concentrations of Ca(2+), ROS-GC1 is stimulated by S100beta [also named calcium-dependent (CD) GCAP]. This mode is not linked with phototransduction; instead, it is predicted to be involved in retinal synaptic activity. Two point mutations, E786D and R787C, in ROS-GC1 have been connected with cone-rod dystrophy (CORD6), with only one type of point mutation occurring in each family. The present study shows that the E786D mutation has no effect on the basal catalytic activity of ROS-GC1 and on its activation by GCAP1 and S100beta; however, the mutated cyclase becomes more activated by GCAP2. The R787C mutation has three consequences: (1) it causes major damage to the basal cyclase activity, (2) it makes the cyclase 5-fold more sensitive to activation by GCAP1; and 3) converts the cyclase into a form that is less sensitive to activation by GCAP2 and S100beta. Thus, the two CORD6-linked mutations in ROS-GC1, which occur at adjacent positions, result in vastly different biochemical phenotypes, and they are connected with very specific molecular defects in the Ca(2+) switching components of the cyclase. These defects, in turn, are proposed to have a profound effect on both the machinery of phototransduction and the retinal synapse. The study for the first time defines the biochemistry of CORD6 pathology in precise molecular terms. PMID- 10529239 TI - Characterization of stromelysin 1 (MMP-3), matrilysin (MMP-7), and membrane type 1 matrix metalloproteinase (MT1-MMP) derived fibrin(ogen) fragments D-dimer and D like monomer: NH2-terminal sequences of late-stage digest fragments. AB - Matrix metalloproteinases (MMPs) participate in physiological remodeling of the extracellular matrix. Recently we determined that both fibrinogen (Fg) and cross linked fibrin (XL-Fb) are substrates for selected MMPs. Specifically, XL-Fb clots were solubilized by MMP-3 (stromelysin 1) by cleavage at gamma Gly 404-Ala 405, resulting in a D-like monomer fragment. Similarly, MMP-7 (matrilysin) and MT1-MMP (membrane type 1 matrix metalloproteinase) solubilized XL-Fb clots. However, the molecular mass of fragment D-dimer, obtained after MMP-7 and MT1-MMP degradation of XL-Fb, is similar to that of fragment D-dimer from plasmin degradation ( approximately 186 kDa). In contrast, fragment D-like monomer, from MMP-3 degradation of both fibrinogen (Fg) and XL-Fb, is similar to fragment D from plasmin degradation of Fg ( approximately 94 kDa). Reduced chains from MMP-3, MMP 7, and MT1-MMP digests of Fg and XL-Fb were subjected to direct sequence analyses and D/D-dimer alpha-chain showed cleavage at both alpha Asp 97-Phe 98 and alpha Asn 102-Asn 103. Degradation of the beta-chain resulted in microheterogeneity of cleavage sites at beta Asp 123-Leu 124, beta Asn 137-Val 138, and beta Glu 141 Tyr 142, whereas all three enzymes cleaved the gamma-chain at gamma Thr 83-Leu 84. In both Fg and XL-Fb, several cleavage sites obtained by proteolysis with MMP 3, MMP-7, and MT1-MMP were found to be in very close proximity to those obtained by plasmin on these same substrates. That does not occur with other MMPs such as MMP-1, -2, and -9 and MT2-MMP. The degradation of XL-Fb by MMPs suggests both plasmin-dependent and independent mechanisms of fibrinolysis that might be relevant in inflammation, angiogenesis, arthritis, and atherosclerosis. PMID- 10529238 TI - Change in the positional specificity of lipoxygenase 1 due to insertion of fatty acids into phosphatidylcholine deoxycholate mixed micelles. AB - Linoleic and arachidonic acids were inserted into phosphatidylcholine deoxycholate mixed micelles (PDM-micelles) with their tail groups buried inside and carboxylic groups exposed outside. The fatty acid hydrophobic tail had a high affinity for the hydrophobic region of phosphatidylcholine micelles. The fatty acids inserted into phosphatidylcholine micelles were better substrates for soybean lipoxygenase 1 (LOX1) with two distinct pH optima at 7.0 and 10.0. With Tween 20-solubilized linoleic acid, the enzyme had a pH optimum at 9.0, exclusively forming 13-hydroperoxides. However, with linoleic and arachidonic acids inserted into PDM-micelles, LOX1 synthesized exclusively 9- and 5 hydroperoxides, respectively. The enzyme brought about the transformation of the substrate either at pH 7.4 or at 10.0, less efficiently at pH 10.0. However, the regioselectivity of the enzyme was not altered by increasing the pH from 7.4 to 10.0. Thus, LOX1 could utilize fatty acids bound to membranes as physiological substrates. The enzyme utilized the carboxylic group of linoleic and arachidonic acids inserted into the PDM-micelles as a recognition site to convert the compounds into 9- and 5-hydroperoxides, respectively. This was confirmed by activity measurements using methyl linoleate as the substrate. Circular dichroism measurement of LOX1 with PDM-micelles suggested that while there was a small change in the tertiary structure of LOX1, the secondary structure was unaffected. Soybean LOX1, which is arachidonate 15-LOX, acted as "5-LOX", thus making it possible to change the regiospecificity of the LOX1-catalyzed reaction by altering the physical state of the substrate. PMID- 10529240 TI - Glycosylation of natural human neutrophil gelatinase B and neutrophil gelatinase B-associated lipocalin. AB - Gelatinase B is a matrix metalloproteinase (MMP-9) involved in tissue remodeling, development, cancer, and inflammation. Neutrophils produce three major forms of (pro)gelatinase B: 92 kDa monomers, homodimers, and complexes of gelatinase B covalently bound to neutrophil gelatinase B-associated lipocalin (NGAL). In contrast to the case for other proteinases, little information about the glycosylation of any natural human MMP is available. Here, both gelatinase B and NGAL were purified from human peripheral blood neutrophils, and the entire contents of the released N- and O-glycan pools were analyzed simultaneously using recently developed high-performance liquid chromatography-based technology. The results are discussed within the context of the domain structure of gelatinase B and a molecular model of NGAL based on data from this study and the three dimensional nuclear magnetic resonance (NMR) structure of the protein. More than 95% of the N-linked glycans attached to both gelatinase B and NGAL were partially sialylated, core-fucosylated biantennary structures with and without outer arm fucose. The O-linked glycans, which were estimated to comprise approximately 85% of the total sugars on gelatinase B, mainly consisted of type 2 cores with Galbeta1,4GlcNAc (lactosamine) extensions, with or without sialic acid or outer arm fucose. This paper also contains the first report of O-linked glycans attached to NGAL. Although both proteins were isolated from neutrophils and contained O-linked glycans mainly with type 2 cores, the glycans attached to individual serine/threonine residue(s) in NGAL were significantly smaller than those on gelatinase B. In contrast to NGAL, gelatinase B contains a region rich in Ser, Thr, and Pro typical of O-glycosylated mucin-like domains. PMID- 10529241 TI - Substitutions in a flexible loop of horse liver alcohol dehydrogenase hinder the conformational change and unmask hydrogen transfer. AB - When horse liver alcohol dehydrogenase binds coenzyme, a rotation of about 10 degrees brings the catalytic domain closer to the coenzyme binding domain and closes the active site cleft. The conformational change requires that a flexible loop containing residues 293-298 in the coenzyme binding domain rearranges so that the coenzyme and some amino acid residues from the catalytic domain can be accommodated. The change appears to control the rate of dissociation of the coenzyme and to be necessary for installation of the proton relay system. In this study, directed mutagenesis produced the activated Gly293Ala/Pro295Thr enzyme. X ray crystallography shows that the conformations of both free and complexed forms of the mutated enzyme and wild-type apoenzyme are very similar. Binding of NAD(+) and 2,2, 2-trifluoroethanol do not cause the conformational change, but the nicotinamide ribose moiety and alcohol are not in a fixed position. Although the Gly293Ala and Pro295Thr substitutions do not disturb the apoenzyme structure, molecular modeling shows that the new side chains cannot be accommodated in the closed native holoenzyme complex without steric alterations. The mutated enzyme may be active in the "open" conformation. The turnover numbers with ethanol and acetaldehyde increase 1.5- and 5.5-fold, respectively, and dissociation constants for coenzymes and other kinetic constants increase 40-2,000-fold compared to those of the native enzyme. Substrate deuterium isotope effects on the steady state V or V/K(m) parameters of 4-6 with ethanol or benzyl alcohol indicate that hydrogen transfer is a major rate-limiting step in catalysis. Steady state oxidation of benzyl alcohol is most rapid above a pK of about 9 for V and V/K(m) and is 2-fold faster in D(2)O than in H(2)O. The results are consistent with hydride transfer from a ground state zinc alkoxide that forms a low-barrier hydrogen bond with the hydroxyl group of Ser48. PMID- 10529242 TI - Kinetics and thermodynamics of dimer formation and dissociation for a recombinant humanized monoclonal antibody to vascular endothelial growth factor. AB - The recombinant humanized antibody (rhuMAb) VEGF has a high affinity for vascular endothelial growth factor and is currently being evaluated in clinical trials as a cancer therapeutic. Under acidic pH and low ionic strength conditions, the antibody was predominantly present as monomer. Under physiological conditions, the appearance of significant amounts of a noncovalent, reversible dimer were observed by size-exclusion chromatography. The kinetics and thermodynamics of the reversible self-association for rhuMAb VEGF monomer were investigated as a function of pH, temperature, and ionic strength by size-exclusion chromatography using the concentration jump method. The rate constant for dimer formation ranged 23-112 M(-)(1) min(-)(1) under the conditions studied, values that are significantly lower than those reported in the literature for other proteins that self-associate. The rate constant for dissociation ranged 0.0039-0.021 min(-)(1). Gibbs' free energies, enthalpies, entropies, and activation energies were determined and revealed that dimer formation is optimal at pH 7.5-8.0, which may be reflective of charge shielding occurring near the pI of the protein. There was a negative change in entropy for dissociation (values from -18.1 to -12.8 cal/mol K). In the presence of D(2)O or 1 M NaCl, dimerization was enhanced. The results of the kinetic and thermodynamic analysis of this study indicate that rhuMAb VEGF dimerization occurs primarily through hydrophobic interactions. PMID- 10529244 TI - Pseudomonas aeruginosa contains a novel type V porphobilinogen synthase with no required catalytic metal ions. AB - Porphobilinogen synthases (PBGS) are metalloenzymes that catalyze the first common step in tetrapyrrole biosynthesis. The PBGS enzymes have previously been categorized into four types (I-IV) by the number of Zn(2+) and/or Mg(2+) utilized at three different metal binding sites termed A, B, and C. In this study Pseudomonas aeruginosa PBGS is found to bind only four Mg(2+) per octamer as determined by atomic absorption spectroscopy, in the presence or absence of substrate/product. This is the lowest number of bound metal ions yet found for PBGS where other enzymes bind 8-16 divalent ions. These four Mg(2+) allosterically stimulate a metal ion independent catalytic activity, in a fashion dependent upon both pH and K(+). The allosteric Mg(2+) of PBGS is located in metal binding site C, which is outside the active site. No evidence is found for metal binding to the potential high-affinity active site metal binding sites A and/or B. P. aeruginosa PBGS was investigated using Mn(2+) as an EPR probe for Mg(2+), and the active site was investigated using [3,5-(13)C]porphobilinogen as an NMR probe. The magnetic resonance data exclude the direct involvement of Mg(2+) in substrate binding and product formation. The combined data suggest that P. aeruginosa PBGS represents a new type V enzyme. Type V PBGS has the remarkable ability to synthesize porphobilinogen in a metal ion independent fashion. The total metal ion stoichiometry of only 4 per octamer suggests half-sites reactivity. PMID- 10529243 TI - Production, purification, and characterization of a Mg2+-responsive porphobilinogen synthase from Pseudomonas aeruginosa. AB - During tetrapyrrole biosynthesis the metalloenzyme porphobilinogen synthase (PBGS) catalyzes the condensation of two molecules of 5-aminolevulinic acid to form the pyrrole porphobilinogen. Pseudomonas aeruginosa PBGS was synthesized in Escherichia coli, and the enzyme was purified as a fusion protein with glutathione S-transferase (GST). After removal of GST, a molecular mass of 280 000 +/- 10 000 with a Stokes radius of 57 A was determined for native PBGS, indicating a homooctameric structure of the enzyme. Mg2+ stabilized the oligomeric state but was not essential for octamer formation. Alteration of N terminal amino acids changed the oligomeric state and reduced the activity of the enzyme, revealing the importance of this region for oligomerization and activity. EDTA treatment severely inhibited enzymatic activity which could be completely restored by the addition of Mg2+ or Mn2+. At concentrations in the micromolar range Co2+, Zn2+, and Ni2+ partially restored EDTA-inhibited enzymatic activity while higher concentrations of Zn2+ inhibited the enzyme. Pb2+, Cd2+, and Hg2+ did not restore activity. A stimulatory effect of monovalent ions was observed. A Km of 0.33 mM for ALA and a maximal specific activity of 60 micromol h-1 mg-1 at the pH optimum of 8.6 in the presence of Mg2+ and K+ were found. pH-dependent kinetic studies were combined with protein modifications to determine the structural basis of two observed pKa values of approximately 7.9 (pKa1) and 9.5 (pKa2). These are postulated respectively as ionization of an active site lysine residue and of free substrate during catalysis. Some PBGS inhibitors were characterized. Finally, we succeeded in obtaining well-ordered crystals of P. aeruginosa PBGS complexed with the substrate analogue levulinic acid. PMID- 10529245 TI - Mechanism of hypochlorite-mediated inactivation of proteinase inhibition by alpha 2-macroglobulin. AB - The proteinase-proteinase inhibitor balance plays an important role in mediating inflammation-associated tissue destruction. alpha 2-Macroglobulin (alpha 2M) is a high-affinity, broad-spectrum proteinase inhibitor found abundantly in plasma and interstitial fluids. Increased levels of alpha 2M and proteinase-alpha 2M complexes can be demonstrated in patients with sepsis, emphysema, peridontitis, rheumatoid arthritis, and other inflammatory diseases. Despite these increased levels, proteolysis remains a significant problem. We hypothesized that a mechanism for inactivating alpha 2M-mediated proteinase inhibition must exist and recently demonstrated that alpha 2M isolated from human rheumatoid arthritis synovial fluid is oxidized and has decreased functional activity. The oxidant responsible for alpha 2M inactivation and the mechanism of such destruction were not studied. We now report that while hypochlorite and hydroxyl radical both modify amino acid residues on alpha 2M, only hypochlorite can abolish the ability of alpha 2M to inhibit proteinases. Hydrogen peroxide, on the other hand, has no effect on alpha 2M structure or function. Protein unfolding with increased susceptibility to proteolytic cleavage appears to be involved in alpha 2M inactivation by oxidation. The in vivo relevance of this mechanism is supported by the presence of multiple cleavage fragments of alpha 2M in synovial fluid from patients with rheumatoid arthritis, where significant tissue destruction occurs, but not in patients with osteoarthritis. These results provide strong evidence that hypochlorite oxidation contributes to enhanced tissue destruction during inflammation by inactivating alpha 2M. PMID- 10529246 TI - Recombinant human liver betaine-homocysteine S-methyltransferase: identification of three cysteine residues critical for zinc binding. AB - Betaine-homocysteine S-methyltransferase (BHMT; EC 2.1.1.5) catalyzes the transfer of an N-methyl group from betaine to homocysteine to produce dimethylglycine and methionine, respectively. The enzyme is found in the pathway of choline oxidation and is abundantly expressed in liver and kidney. We have recently shown that human BHMT is a zinc metalloenzyme [Millian, N. S., and Garrow, T. A. (1998) Arch. Biochem. Biophys. 356, 93-98]. To facilitate the rapid purification of human BHMT for further physical and mechanistic studies, including characterizing its metal binding properties, we have overexpressed the enzyme in E. coli as a fusion construct which facilitated its subsequent purification by a self-cleavable affinity tag system (IMPACT T7). Using this expression and purification system in conjunction with site-directed mutagenesis, we have identified Cys217, Cys299, and Cys300 as zinc ligands. Mutating any of these Cys residues to Ala results in the complete loss of activity and a significant reduction in the ability of the protein to bind zinc. Comparing the regions of BHMT amino acid sequence surrounding these Cys residues with similar amino acid sequences retrievable from protein databases, we have identified the following motif: G[ILV]NCX(20,100)[ALV]X(2)[ILV]GGCCX(3)PX(2)I, which we propose to be a signature for a family of zinc-dependent methyltransferases that utilize thiols or selenols as methyl acceptors. Some of the members of this family include the vitamin B(12)-dependent methionine synthases, E. coli S methylmethionine-S-homocysteine methyltransferase, and A. bisulcatus S methylmethionine-selenocysteine methyltransferase. PMID- 10529247 TI - Rhodobacter sphaeroides phosphoribulokinase: identification of lysine-165 as a catalytic residue and evaluation of the contributions of invariant basic amino acids to ribulose 5-phosphate binding. AB - Rhodobacter sphaeroides phosphoribulokinase (PRK) is inactivated upon exposure to pyridoxal phosphate/sodium borohydride, suggesting a reactive lysine residue. Protection is afforded by a combination of the substrate ATP and the allosteric activator NADH, suggesting that the targeted lysine maps within the active site. PRK contains two invariant lysines, K53 and K165. PRK-K53M retains sensitivity to pyridoxal phosphate, implicating K165 as the target of this reagent. PRK-K165M retains wild-type structure, as judged by titration with effector NADH and the tight-binding alternative substrate trinitrophenyl-ATP. The catalytic activity of K165M and K165C mutants is depressed by >10(3)-fold. Residual activity of K165M is insensitive to pyridoxal phosphate, confirming K165 as the target of this reagent. The decreased catalytic efficiency of K165 mutants approaches the effect measured for a mutant of D169, which forms a salt-bridge to K165. K165M exhibits a 10-fold increase in S()1(/)()2 (ATP) and a 10(2)-fold increase in K(m) (Ru5P). To evaluate the contribution to Ru5P binding of K165 in comparison with this substrate's interaction with invariant H45, R49, R168, and R173, PRKs mutated at these positions have been used to determine relative K(i) values for 6 phosphogluconate, a competitive inhibitor with respect to Ru5P. Elimination of the basic side chain of K165, R49, and H45 results in increases in K(m) (Ru5P) which correlate well with the magnitude of increases in K(i) (phosphogluconate). In contrast, while mutations eliminating charge from R168 and R173 result in enzymes with substantial increases in K(m) (Ru5P), such mutant enzymes exhibit only small increases in K(i) (phosphogluconate). These observations suggest that K165, R49, and H45 are major contributors to Ru5P binding. PMID- 10529248 TI - Determinants for differential effects on D-Ala-D-lactate vs D-Ala-D-Ala formation by the VanA ligase from vancomycin-resistant enterococci. AB - Bacteria with either intrinsic or inducible resistance to vancomycin make peptidoglycan (PG) precursors of lowered affinity for the antibiotic by switching the PG-D-Ala-D-Ala termini that are the antibiotic-binding target to either PG-D Ala-D-lactate or PG-D-Ala-D-Ser as a consequence of altered specificity of the D Ala-D-X ligases in the cell wall biosynthetic pathway. The VanA ligase of vancomycin-resistant enterococci, a D-Ala-D-lactate depsipeptide ligase, has the ability to recognize and activate the weak nucleophile D-lactate selectively over D-Ala(2) to capture the D-Ala(1)-OPO(3)(2)(-) intermediate in the ligase active site. To ensure this selectivity in catalysis, VanA largely rejects the protonated (NH(3)(+)) form of D-Ala at subsite 2 (K(M2) of 210 mM at pH 7.5) but not at subsite 1. In contrast, the deprotonated (NH(2)) form of D-Ala (K(M2) of 0.66 mM, k(cat) of 550 min(-)(1)) is a 17-fold better substrate compared to D lactate (K(M) of 0.69 mM, k(cat) of 32 min(-)(1)). The low concentration of the free amine form of D-Ala at physiological conditions (i.e., 0.1% at pH 7.0) explains the inefficiency of VanA in dipeptide synthesis. Mutational analysis revealed a residue in the putative omega-loop region, Arg242, which is partially responsible for electrostatically repelling the protonated form of D-Ala(2). The VanA enzyme represents a subfamily of D-Ala-D-X ligases in which two key active site residues (Lys215 and Tyr216) in the active-site omega-loop of the Escherichia coli D-Ala-D-Ala ligase are absent. To look for functional complements in VanA, we have mutated 20 residues and evaluated effects on catalytic efficiency for both D-Ala-D-Ala dipeptide and D-Ala-D-lactate depsipeptide ligation. Mutation of Asp232 caused substantial defects in both dipeptide and depsipeptide ligase activity, suggesting a role in maintaining the loop position. In contrast, the H244A mutation caused an increase in K(M2) for D lactate but not D-Ala, indicating a differential role for His244 in the recognition of the weaker nucleophile D-lactate. Replacement of the VanA omega loop by that of VanC2, a D-Ala-D-Ser ligase, eliminated D-Ala-D-lactate activity while improving by 3-fold the catalytic efficacy of D-Ala-D-Ala and D-Ala-D-Ser activity. PMID- 10529249 TI - Tandem heterocyclization activity of the multidomain 230 kDa HMWP2 subunit of Yersinia pestis yersiniabactin synthetase: interaction of the 1-1382 and 1383 2035 fragments. AB - The six-domain, 2035-amino acid subunit high-molecular weight protein 2 (HMWP2) activates salicylate and two cysteines and loads them covalently on its three carrier protein domains during assembly of the iron-chelating virulence factor, yersiniabactin of the plague bacterium Yersinia pestis. The 1-1382 fragment of HMWP2 (ArCP-Cy1-A), overproduced in Escherichia coli, contains the first three domains: the aryl carrier protein (ArCP) domain, the cysteine specific adenylation domain (A), and the first condensation/cyclization domain (Cy1). The ArCP can be posttranslationally phosphopantetheinylated on Ser52 and then loaded with a salicyl group on the phosphopantetheine (Ppant) thiol by action of the YbtE, a salicyl-AMP ligase. The HMWP2 1-1382 fragment can activate L-cysteine as Cys-AMP. The HMWP2 1383-2035 fragment contains the remaining three domains: two peptidyl carrier proteins (PCP1 and PCP2) separated by a second condensation/cyclization domain (Cy2). Phosphopantetheinylation of the HMWP2 1383 2035 fragment at Ser1439 (PCP1) and Ser1977 (PCP2) facilitates cysteinylation of both thiols by HMWP2 1-1382. When the holo 1-1382 and bis-holo 1383-2035 protein fragments are mixed with ATP, salicylate, and cysteine, four products are slowly released [salicylcysteine (Sal-Cys), (hydroxyphenylthiazolinyl)cysteine (HPT Cys), HPT-Cys-Cys, and the bisheterocyclic HPTT-Cys], reflecting thiolytic rerouting by cysteine in solution of elongating acyl-S-enzyme intermediates tethered at ArCP, PCP1, and PCP2 carrier protein domains, respectively. Conducting the in trans reconstitution with the S1439A mutant of HMWP2 1383-2035 releases only Sal-Cys, while the S1977A mutant leads to HPT-Cys formation but not HPT-Cys-Cys or HPTT-Cys. These results suggest localization of particular acyl-S enzyme intermediates to each of the three carrier protein regions and also establish the sequential action of Cy1 and Cy2, with the latter producing the tandem 4,2-bisheterocyclic hydroxyphenylthiazolinylthiazolinyl (HPTT) moiety characteristic of this class of siderophores. PMID- 10529250 TI - The FinO repressor of bacterial conjugation contains two RNA binding regions. AB - Conjugative transfer of F-like plasmids in Escherichia coli is repressed by a plasmid-encoded protein, FinO. FinO blocks the translation of TraJ, a positive activator of transcription of genes required for conjugation. FinO binds a traJ antisense RNA, FinP, thereby protecting it from degradation, and catalyzes FinP traJ mRNA hybridization. Interactions between these two RNAs are predicted to block the traJ ribosomal binding site. In this paper, we use limited proteolysis, circular dichroism spectroscopy, and an electrophoretic mobility shift assay to map the regions within FinO that are required for interactions with RNA. Our results show that FinO is largely helical, binds to its highest affinity binding site within FinP as a monomer, and contains two distinct RNA binding regions, one of which is localized between residues 26 and 61, and a second which is localized between residues 62 and 186. PMID- 10529251 TI - Tallimustine lesions in cellular DNA are AT sequence-specific but not region specific. AB - Tallimustine (FCE 24517) is an AT-specific alkylating antitumor derivative of distamycin. This study examined levels of tallimustine lesions in intracellular DNA, their sequence- and region-specificity, and the long-range distribution of the drug binding motif. Tallimustine adducts in DNA converted to strand breaks by heating allowed the quantitation of drug lesions. In bulk DNA of intact human leukemia CEM cells, tallimustine formed 0.15 +/- 0.04 and 0.64 +/- 0.18 lesions/kbp at 5 and 50 microM, respectively. These lesions represent monoadducts as no interstrand cross-links or DNA-protein cross-links were detected. Tallimustine adducts in intracellularly treated DNA showed a general preference for sequences with T-tracts, suggesting a propensity for intrinsically bent motifs. Major drug-adducted sites identified by repetitive primer extension, included 5'-TTTTGPu-3' and 5'-TTTTGC-3' motif. Despite the high specificity at the nucleotide level, tallimustine did not differentiate among bulk DNA and three discrete AT-rich regions of genomic DNA examined by quantitative PCR stop assay with lesion frequencies ranging from 0.23 to 0.39 lesions/kbp at 25 microM drug. In comparisons of lesion frequencies and cytotoxicity, tallimustine adducts are approximately 50 times more lethal than relatively nonsequence specific cisplatin adducts but are >100 times less lethal than lesions by an unrelated AT-specific drug, bizelesin. However, the 5'-TTTTGPu-3' motifs targeted by tallimustine are relatively infrequent and scattered throughout the genome. In contrast, the motifs 5'-T(A/T)(4)A-3' motifs targeted by bizelesin, while also infrequent, cluster in defined AT-rich islands. The lack of region-specificity may be the reason tallimustine adducts, despite high AT-specificity at the nucleotide level, are less lethal than region-specific bizelesin adducts. PMID- 10529252 TI - C8-guanine adduct-induced stabilization of a -1 frame shift intermediate in a nonrepetitive DNA sequence. AB - The mechanism of frame shift mutagenesis induced by N-(deoxyguanosin-8-yl)-1 aminopyrene, the major DNA adduct formed by the carcinogen 1-nitropyrene, was investigated by thermal melting studies of a 13-mer in which the adduct was flanked by a 5' and a 3' C. Compared to the unmodified 13-mer, the adduct destabilized the duplex by 4-5 kcal/mol, and the DeltaDeltaG value remained approximately the same regardless of which base was placed opposite the adduct. In contrast, deletion of the base opposite the adduct stabilized the duplex by nearly 4 kcal/mol. The adduct in the same sequence context was inserted into a bacteriophage M13 DNA containing the simian virus 40 origin of replication. The constructed DNA template was replicated in vitro with extracts from normal human fibroblasts. The adduct was not removed from the progeny DNA following bidirectional semiconservative replication, which suggests that it had been bypassed, rather than repaired, by the cell extract. When newly replicated bacteriophage was evaluated for mutations in the region of the modified G, most contained a G at the adduct site, indicating error-free replication. A small number of mutants ( approximately 2 x 10(-3)) were detected, all of which contained a targeted G.C base pair deletion. This suggests a relationship between the thermodynamic stability of the adduct in DNA and the errors that occurred during replicative bypass by the human DNA polymerases. PMID- 10529253 TI - A new model for the K+-induced macromolecular structure of guanosine 5' monophosphate in solution. AB - The (31)P NMR spectra of (TMA)(2)(5'-GMP), where TMA is [(CH(3))(4)N](+) and 5' GMP is guanosine 5'-monophosphate, and K(2)(5'-GMP), containing various amounts of KCl or TMACl, have been obtained at 2 degrees C. Variable-temperature spectra have also been obtained for K(2)(5'-GMP). The TMA(+) ion serves to neutralize the charge on the dianionic 5'-GMP and permits the added K(+) to bond preferentially in structure-forming sites. (1)H NMR spectra (one- and two-dimensional) have been obtained for K(2)(5'-GMP) and used to assign the proton resonances in the self associated structures and determine that all residues have the anti glycosidic conformation. The (31)P and (1)H NMR spectra are very complex and indicate the presence of a large number of molecular environments and a structural variation dependent upon the mole ratio of 5'-GMP to K(+). A new model for the solution structure is proposed in which the 5'-GMP forms a pseudo-four-stranded helix with guanine-guanine hydrogen bonding forming a continuous helical strand, rather than the usual planar G-tetrad structure. The guanine-guanine hydrogen bonding sites are the same as that found in a G-tetrad. The K(+) ions would be located in the center of the helix and bonding to the carbonyl oxygens. They are interacting with the phosphates as well. Integration data from the largest sized species give an estimate of 14.3 +/- 1.1 residues in a helical structure. PMID- 10529254 TI - DNA toroids: stages in condensation. AB - The effects of polylysine (PLL) and PLL-asialoorosomucoid (AsOR) on DNA condensation have been analyzed by AFM. Different types of condensed DNA structures were observed, which show a sequence of conformational changes as circular plasmid DNA molecules condense progressively. The structures range from circular molecules with the length of the plasmid DNA to small toroids and short rods with approximately 1/6 to 1/8 the contour length of the uncondensed circular DNA. Single plasmid molecules of 6800 base pairs (bp) condense into single toroids of approximately 110 nm diameter, measured center-to-center. The results are consistent with a model for DNA condensation in which circular DNA molecules fold several times into progressively shorter rods. Structures intermediate between toroids and rods suggest that at least some toroids may form by the opening up of rods as proposed by Dunlap et al. [(1997) Nucleic Acids Res. 25, 3095]. Toroids and rods formed at lysine:nucleotide ratios of 5:1 and 6:1. This high lysine:nucleotide ratio is discussed in relation to entropic considerations and the overcharging of macroions. PLL-AsOR is much more effective than PLL alone for condensing DNA, because several PLL molecules are attached to a single AsOR molecule, resulting in an increased cation density. PMID- 10529255 TI - Kinetics and interactions of molybdenum and iron-sulfur centers in bacterial enzymes of the xanthine oxidase family: mechanistic implications. AB - For isoquinoline 1-oxidoreductase (IsoOr), the reaction mechanism under turnover conditions was studied by EPR spectroscopy using rapid-freeze methods. IsoOr displays several EPR-active Mo(V) species including the "very rapid" component found also in xanthine oxidase (XanOx). For IsoOr, unlike XanOx or quinoline 2 oxidoreductase (QuinOr), this species is stable for about 1 h in the absence of an oxidizing substrate [Canne, C., Stephan, I., Finsterbusch, J., Lingens, F., Kappl, R., Fetzner, S., and Huttermann, J. (1997) Biochemistry 36, 9780-9790]. Under rapid-freeze conditions in the presence of ferricyanide the very rapid species behaves as a kinetically competent intermediate present only during steady-state turnover. To explain the persistence of the very rapid species in IsoOr in the absence of an added oxidant, extremely slow product dissociation is required. This new finding that oxidative conditions facilitate decay of the very rapid signal for IsoOr supports the mechanism of substrate turnover proposed by Lowe, Richards, and Bray [Lowe, D. J., Richards, R. L., and Bray, R. C. (1997) Biochem. Soc. Trans. 25, 774-778]. Additional stopped-flow data reveal that alternative catalytic cycles occur in IsoOr and show that the product dissociates after transfer of a single oxidizing equivalent from ferricyanide. In rapid freeze measurements magnetic interactions of the very rapid Mo(V) species and the iron-sulfur center FeSI of IsoOr and QuinOr were observed, proving that FeSI is located close to the molybdopterin cofactor in the two proteins. This finding is used to relate the two different iron-sulfur centers of the aldehyde oxidoreductase structure with the EPR-detectable FeS species of the enzymes. PMID- 10529256 TI - Random mutations directed to transmembrane and loop domains of the light harvesting chlorophyll a/b protein: impact on pigment binding. AB - The major light-harvesting complex of photosystem II (LHCII) can be reconstituted in vitro by folding its bacterially expressed apoprotein, Lhcb, in detergent solution in the presence of chlorophylls and carotenoids. To compare the impact of alpha-helical transmembrane domains and hydrophilic loop domains of the apoprotein on complex formation and stability, we introduced random mutations into a segment of the protein comprising the stromal loop, the third (C-proximal) transmembrane helix, and part of the amphipathic helix in the C-terminal domain. The mutant versions of Lhcb were screened for the loss of their ability to form stable LHCII upon reconstitution in vitro. Most steps during the screening, including expression of the recombinant protein, its reconstitution with pigments, and the assay for complex formation by measuring energy transfer from chlorophyll b to chlorophyll a, were performed as one-vessel reactions on 96-well microtiter plates. This enabled us to screen several hundred mutant Lhcb versions. Mutants that had lost their ability to form stable LHCII carried between one and four amino acid exchanges. Among the single-point mutations, several were at positions in the C-proximal transmembrane helix, including an amino acid that is thought to be directly involved in chlorophyll binding. However, we also found four point mutations in the stromal loop domain that, in our assay, completely abolished the formation of stable LHCII. These data show that the stromal loop domain has a significant impact on LHCII formation and/or stability in vitro. PMID- 10529257 TI - Structural determinant of the vesicle aggregation activity of annexin I. AB - Some annexins, including annexins I, II, IV, and VII, can promote membrane aggregation. To identify amino acids involved in annexin I-mediated membrane aggregation, we generated truncated mutants of human annexin I lacking various parts of the amino terminus. The in vitro vesicle binding and aggregation activities of these mutants indicated that both the amino-terminal region of annexin I spanning residues 26-29 and the carboxy-terminal core are involved in membrane aggregation. This notion was further supported by the finding that a chimera composed of residues 24-35 of annexin I and the core of annexin V has vesicle aggregation activity that is significantly higher than that of annexin V but lower than that of annexin I. Further site-specific mutations in the amino terminal region of annexin I indicated that Lys-26 and Lys-29 are essential for its membrane aggregation activity. The comparison of tryptic digest patterns of free and vesicle-bound wild type and K29E mutant suggests that a primary role of Lys-26 and Lys-29 is to induce and stabilize an active conformation of annexin I for vesicle aggregation. PMID- 10529258 TI - Thermodynamic instability of human lambda 6 light chains: correlation with fibrillogenicity. AB - Certain types of human light chains have the propensity to deposit pathologically as amyloid fibrils as evidenced by the preferential association of monoclonal lambda 6 proteins with AL amyloidosis. However, the molecular features that render such proteins amyloidogenic have not been elucidated. Based upon the demonstrated relationship between the thermodynamic stability of light chains and their propensity to aggregate in vitro, we have initiated studies where the thermodynamic properties and fibrillogenic potential of two recombinant (r) V lambda 6 molecules were compared. The first protein was generated from cDNA cloned from marrow-derived plasma cells from a patient (Wil) who had AL amyloidosis and renal amyloid deposits; the second was from a patient (Jto) with multiple myeloma in whom the lambda 6 protein was deposited not as amyloid but in the form of renal tubular casts. The thermodynamic stabilities of rV lambda 6Wil and -Jto were determined from chaotropic and thermal denaturation studies. Based upon the Delta GH2O, Delta H, Delta G25 degrees C, Tm, and Cm values, the rV lambda 6Wil was less stable than its nonamyloidogenic counterpart, rV lambda 6Jto. Measurement of fibril formation using a novel in vitro fibril forming assay demonstrated that although both rV lambda 6 proteins formed fibrils in vitro, Wil had a shorter lag time and exhibited faster kinetics under physiologic conditions. Comparative amino acid sequence analyses of these two components and other lambda 6 amyloid-associated light chains revealed that the Jto protein had certain primary structural features that we posit contributed to its increased stability and thus rendered this protein nonamyloidogenic. Our studies provide the first evidence that stabilizing interactions within the V L domain can influence the kinetics of light chain fibrillogenicity. PMID- 10529260 TI - Analysis of the functional coupling between Calmodulin's calcium binding and peptide recognition properties PMID- 10529259 TI - Expression and mutagenesis of ZntA, a zinc-transporting P-type ATPase from Escherichia coli. AB - Cation-transporting P-type ATPases comprise a major membrane protein family, the members of which are found in eukaryotes, eubacteria, and archaea. A phylogenetically old branch of the P-type ATPase family is involved in the transport of heavy-metal ions such as copper, silver, cadmium, and zinc. In humans, two homologous P-type ATPases transport copper. Mutations in the human proteins cause disorders of copper metabolism known as Wilson and Menkes diseases. E. coli possesses two genes for heavy-metal translocating P-type ATPases. We have constructed an expression system for one of them, ZntA, which encodes a 732 amino acid residue protein capable of transporting Zn(2+). A vanadate-sensitive, Zn(2+)-dependent ATPase activity is present in the membrane fraction of our expression strain. In addition to Zn(2+), the heavy-metal ions Cd(2+), Pb(2+), and Ag(+) activate the ATPase. Incubation of membranes from the expression strain with [gamma-(33)P]ATP in the presence of Zn(2+), Cd(2+), or Pb(2+) brings about phosphorylation of two membrane proteins with molecular masses of approximately 90 and 190 kDa, most likely representing the ZntA monomer and dimer, respectively. Although Cu(2+) can stimulate phosphorylation by [gamma (33)P]ATP, it does not activate the ATPase. Cu(2+) also prevents the Zn(2+) activation of the ATPase when present in 2-fold excess over Zn(2+). Ag(+) and Cu(+) appear not to promote phosphorylation of the enzyme. To study the effects of Wilson disease mutations, we have constructed two site-directed mutants of ZntA, His475Gln and Glu470Ala, the human counterparts of which cause Wilson disease. Both mutants show a reduced metal ion stimulated ATPase activity (about 30-40% of the wild-type activity) and are phosphorylated much less efficiently by [gamma-(33)P]ATP than the wild type. In comparison to the wild type, the Glu470Ala mutant is phosphorylated more strongly by [(33)P]P(i), whereas the His475Gln mutant is phosphorylated more weakly. These results suggest that the mutation His475Gln affects the reaction with ATP and P(i) and stabilizes the enzyme in a dephosphorylated state. The Glu470Ala mutant seems to favor the E2 state. We conclude that His475 and Glu470 play important roles in the transport cycles of both the Wilson disease ATPase and ZntA. PMID- 10529261 TI - Long-term functional morbidity after mild hyperthermic isolated limb perfusion with melphalan. AB - AIMS: To assess long-term functional morbidity in patients entered in the prospective randomized EORTC trial investigating the role of adjuvant isolated limb perfusion (ILP) with melphalan for high-risk primary melanoma. METHODS: In 65 patients (ILP 36, wide excision only 29), limb circumference and joint mobility measurements were performed on the treated and the contralateral limb after a mean interval of 48 months after primary treatment. The two treatment groups were comparable regarding age, sex distribution, percentage of skin grafts or regional lymph-node dissections, and interval between primary treatment and physical measurements. RESULTS: None of the patients had severe complaints of the treated limb at the time of analysis. The ankle suffered most from ILP, with a statistical significant restricted extension in approximately 40% of the perfused patients. Abduction of the shoulder was minimally affected in treated upper limbs, probably as a result from the formation of scar tissue after axillary lymph-node dissection. Although no significant differences could be demonstrated in the circumference of upper or lower limbs, atrophy was seen in 24% of perfused lower limbs. Of the five perfused patients who developed oedema, four had also undergone a regional lymph-node dissection. CONCLUSION: This risk of long-term functional morbidity should be weighed against the possible advantages of ILP in patients with limb melanoma or sarcoma. PMID- 10529262 TI - Autoradiographic analysis of follicle-stimulating hormone and human chorionic gonadotropin receptors in the ovary of immature rats treated with equine chorionic gonadotropin. AB - The gonadotropin-primed immature rat has become the most common model for the study of follicular development and ovulation. In this study, prepubertal female rats, 23 and 24 days old, were injected s. c. with 5 IU eCG, and ovaries were collected for topical autoradiography of FSH and hCG receptors at 48 or 24 h post eCG, respectively (i.e., Day 25). In a baseline group, on Day 25 (before eCG), even the smallest preantral follicles with 1 layer of granulosa cells (GCs; primary follicles) possessed FSH receptors, but hCG receptors were found only on the theca of follicles with 2 or more layers of GCs. Human CG receptors were especially prominent in the interstitium that intimately surrounds preantral follicles without any distinction between theca and interstitial cells. There was a discrete theca surrounding antral follicles. Occasionally antral follicles had hCG receptors in the interstitium, but the adjacent theca was negative, suggesting that these follicles might be destined for atresia. By 24 h post-eCG, a now-discrete theca layer with hCG receptors surrounded all preantral follicles except for the primary follicles, which never responded to eCG. The interstitium was hypertrophied and epithelioid, as was the theca surrounding nonatretic preantral and antral follicles. Increased mitotic activity characterized the growing preantral follicle, and for the first time, FSH binding in GCs of antral follicles was greater than in the preantral population. By 48 h post-eCG, the primary follicles were still unresponsive to eCG. FSH receptors were even more pronounced in the GCs of large antral follicles, although hCG receptors were present in the GCs of only one third of the antral follicles, reflecting the small dose of eCG administered. By 48 h post-eCG, receptors in the interstitium were barely detectable. Using this model, the following study considers the functional in vitro changes in steroidogenesis in follicles from the smallest preantral follicles to the largest antral follicles. PMID- 10529263 TI - In vitro steroidogenesis by dissociated rat follicles, primary to antral, before and after injection of equine chorionic gonadotropin. AB - Prepubertal female rats were injected s.c. with 5.0 IU eCG, and ovaries were collected 24 and 48 h post-eCG, on Day 25, as well as from an untreated group also on Day 25. Large antral follicles were manually dissected, and the ovarian remnants were incubated with collagenase overnight to liberate preantral follicles from adhering stromal cells. The viability of the follicles was established by normal histology and lack of pyknotic granulosa cells (GCs) and by their ability to secrete steroids. After a 1-h baseline incubation, either 10 ng LH or 100 ng FSH was added for an additional hour, and the media-before and after gonadotropin administration-were used to measure progesterone, androstenedione, and estradiol by RIA. A distinct hierarchy existed in steroid synthesis, with the maximal production by the largest (700 microm) antral follicles. The major steroid that had accumulated after addition of LH at 48 h post-eCG was androstenedione (1099 pg/follicle per hour), followed by equal amounts of progesterone (155 pg/follicle per hour) and estradiol (191 pg/follicle per hour). There was a precipitous drop in steroid production by 550-microm and 400-microm antral follicles, especially in estradiol for the latter-sized follicles (0.08 pg/follicle per hour). Preantral follicles also produced progesterone and androstenedione after addition of LH. For example, follicles 222 microm in diameter with 4-5 layers of GCs and well-developed theca responded to LH at 48 h post-eCG by accumulating androstenedione (37 pg/follicle per hour) and progesterone (6 pg/follicle per hour) but negligible estradiol. The smallest follicles secreting steroids, 110-148 microm in diameter, had 2-4 layers of GCs. However, primary follicles (1 layer of GCs and no theca) did not synthesize appreciable amounts of any steroid. Although small preantral follicles were consistently stimulated by LH, FSH was ineffective. This result differs from findings in the hamster showing that intact preantral follicles with 1-4 layers of GCs and no theca respond to FSH by secreting progesterone in vitro (Roy and Greenwald, Biol Reprod 1987; 31:39-46). The technique developed to collect intact rat follicles should be useful for numerous investigations. PMID- 10529264 TI - FSP95, a testis-specific 95-kilodalton fibrous sheath antigen that undergoes tyrosine phosphorylation in capacitated human spermatozoa. AB - Protein tyrosine phosphorylation has been associated with both capacitation and motility of mammalian sperm. During capacitation, human spermatozoa undergo tyrosine phosphorylation of a characteristic set of proteins, only one of which has thus far been cloned and localized. We report here the sequence of a fibrous sheath protein of 95 kDa (FSP95) that undergoes tyrosine phosphorylation during capacitation of human spermatozoa and has similarity to sperm A-kinase anchor proteins (AKAPs). FSP95 is both auto- and iso-antigenic in humans as it is recognized by sera containing antisperm antibodies from infertile men and women. The 853-residue protein has a calculated molecular weight of 94.6 kDa and an isoelectric point (pI) of 6.0, and it contains multiple potential phosphorylation sites for protein kinase C and casein kinase II as well as one potential tyrosine kinase phosphorylation site at amino acid 435. The sequence has amino acid homology to mouse sperm fibrous sheath AKAP82 (pro-mAKAP82, 34% identity) and to human sperm fibrous sheath AKAP82 (pro-hAKAP82, 32% identity). The gene encoding FSP95 has 5 exons separated by 4 introns and is located on chromosome 12 at locus p13.3. Northern analysis detected a single transcript of approximately 3.0 kilobases, and Northern dot blot analysis of 50 human tissues revealed FSP95 mRNA expression only in testis. By employing sperm immobilization, indirect immunofluorescence, and immunoelectron microscopy with antisera to purified recombinant FSP95, the protein was localized to the ribs of the fibrous sheath in the principal piece of the sperm tail. FSP95 is the second fibrous sheath protein to be cloned, sequenced and localized in human spermatozoa. PMID- 10529265 TI - Control of gene expression at the onset of bovine embryonic development. AB - The objective of this study was to examine the timing and mechanisms involved in transcription initiation in bovine embryos. Transcriptional activity and its regulation were explored by labeling 1-cell zygotes and 2-cell embryos with [(3)H]uridine in the presence or absence of alpha-amanitin, aphidicolin, and tricostatin A (TSA) (inhibitors of mRNA synthesis, DNA replication, and histone deacetylases, respectively) followed by a total RNA isolation and determination of [(3)H]uridine incorporation. We also analyzed translation of zygotic/embryonic mRNAs by labeling zygotes and 2-cell embryos with [(35)S]methionine in the presence or absence of alpha-amanitin, aphidicolin, and TSA followed by two dimensional PAGE and autoradiography. We show that bovine 1-cell zygotes and 2 cell embryos are transcriptionally and translationally active. The first and second rounds of DNA replication are important regulators of early gene expression as the inhibition of DNA replication resulted in a dramatic decrease in both transcriptional and translational activity. Moreover, acetylation of histones plays an important role in this early gene activation at the onset of embryonic development in the cow. PMID- 10529266 TI - The proestrous prolactin surge is not the sole initiator of regressive changes in corpora lutea of normally cycling rats. AB - During the estrous cycle, secretion of prolactin is largely restricted to a surge on proestrus. We investigated whether this proestrous prolactin surge initiates regression of the corpora lutea of the preceding cycle. Adult rats were killed prior to the prolactin surge (Proestrus group), following the prolactin surge (Estrus group), after chemical blockade of the prolactin surge with bromocryptine (Estrus+BRC group), and after blockade of the prolactin surge and administration of prolactin (Estrus+BRC+PRL group). Corpora lutea of the current (proestrus) or preceding (estrus) cycle were dissected out, weighed, and sectioned for immunohistochemistry or cultured for examination of in vitro progestin production. Numbers of luteal monocytes/macrophages, differentiated macrophages, and apoptotic nuclei per high-power field were greater for Estrus and Estrus+BRC+PRL than for Estrus+BRC, which in turn had greater numbers than Proestrus (P< 0.05). In contrast, BRC completely reversed the decline in luteal weight observed between Proestrus and Estrus (P<0.05). Number of major histocompatibility complex II-positive cells was not different between groups (P>0.05). Finally, progestin production by corpora lutea in vitro was lower for Proestrus than for the other groups (P<0.05). The results indicate that the prolactin surge alone is not responsible for initiation of apoptosis or immune cell infiltration in regressing corpora lutea of the estrous cycle, although prolactin increases these markers of regression. Prolactin does cause a decline in luteal weight; however, the corpora lutea retain the capacity for steroidogenesis. We conclude that although prolactin has a role in luteal regression, it is not solely responsible for the initiation of this process. PMID- 10529267 TI - Analysis of expression of genes involved in apolipoprotein E-based lipoprotein metabolism in pregnant mice deficient in the receptor-associated protein, the low density lipoprotein receptor, or apolipoprotein E. AB - Mice deficient in receptor-associated protein (RAP) were phenotypically normal, but in contrast to results previously reported in RAP(-/-) mice, nearly 50% of the offspring died at or shortly after birth. To attempt to determine the reason for this, we analyzed the regulation of expression of genes involved in apolipoprotein E (apoE)-based mechanisms in RAP-deficient mice and compared this to results in mice deficient in low density lipoprotein receptor (LDLR) or apoE. The major finding concerned a large increase in hepatic lipoprotein receptor related protein (LRP) mRNA and LDLR mRNA levels in pregnant RAP knockout mice. This is in contrast to the down-regulation of LRP mRNA and LDLR mRNA, which is normally seen in wild-type mice. Also in LDLR knockout mice, a significant up regulation in expression of LRP mRNA was demonstrated. In apoE knockout mice, hepatic LRP mRNA did not change significantly, while hepatic LDLR mRNA expression was increased. In placenta and uterus, the deficiency of RAP did not markedly affect the expression of LRP and LDLR. Lipoprotein lipase mRNA and apoE mRNA increased during pregnancy in all mice, independent of their genetic status. The current study does not directly explain the increased mortality of RAP(-/-) pups. The data demonstrate, however, important relative changes in expression of the genes analyzed, an indication that LRP and LDLR play an important role in lipid metabolism during pregnancy. PMID- 10529268 TI - Intracellular Ca(2+)-Mg(2+)-ATPase regulates calcium influx and acrosomal exocytosis in bull and ram spermatozoa. AB - Calcium influx is required for the mammalian sperm acrosome reaction (AR), an exocytotic event occurring in the sperm head prior to fertilization. We show here that thapsigargin, a highly specific inhibitor of the microsomal Ca(2+)-Mg(2+) ATPase (Ca(2+) pump), can initiate acrosomal exocytosis in capacitated bovine and ram spermatozoa. Initiation of acrosomal exocytosis by thapsigargin requires an influx of Ca(2+), since incubation of cells in the absence of added Ca(2+) or in the presence of the calcium channel blocker, La(3+), completely inhibited thapsigargin-induced acrosomal exocytosis. ATP-Dependent calcium accumulation into nonmitochondrial stores was detected in permeabilized sperm in the presence of ATP and mitochondrial uncoupler. This activity was inhibited by thapsigargin. Thapsigargin elevated the intracellular Ca(2+) concentration ([Ca(2+)](i)), and this increase was inhibited when extracellular Ca(2+) was chelated by EGTA, indicating that this rise in Ca(2+) is derived from the external medium. This rise of [Ca(2+)](i) took place first in the head and later in the midpiece of the spermatozoon. However, immunostaining using a polyclonal antibody directed against the purified inositol 1,4,5-tris-phosphate receptor (IP(3)-R) identified specific staining in the acrosome region, in the postacrosome, and along the tail, but not in the midpiece region. No staining in the acrosome region was observed in sperm without acrosome, indicating that the acrosome cap was stained in intact sperm. The presence of IP(3)-R in the anterior acrosomal region as well as the induction, by thapsigargin, of intracellular Ca(2+) elevation in the acrosomal region and acrosomal exocytosis, implicates the acrosome as a potential cellular Ca(2+) store. We suggest here that the cytosolic Ca(2+) is actively transported into the acrosome by an ATP-dependent, thapsigargin-sensitive Ca(2+) pump and that the accumulated Ca(2+) is released from the acrosome via an IP(3) gated calcium channel. The ability of thapsigargin to increase [Ca(2+)](i) could be due to depletion of Ca(2+) in the acrosome, resulting in the opening of a capacitative calcium entry channel in the plasma membrane. The effect of thapsigargin on elevated [Ca(2+)](i) in capacitated cells was 2-fold higher than that in noncapacitated sperm, suggesting that the intracellular Ca pump is active during capacitation and that this pump may have a role in regulating [Ca(2+)](i) during capacitation and the AR. PMID- 10529269 TI - Detection of kallikrein gene expression and enzymatic activity in porcine endometrium during the estrous cycle and early pregnancy. AB - Porcine conceptuses rapidly elongate within the uterine horns prior to the period of placental attachment. During the time of elongation, secretion of estrogen by the developing conceptuses occurs for the establishment of pregnancy through maintenance of corpora lutea and facilitation of placental attachment. Factors associated with the uterine luminal epithelium accentuate embryo attachment by allowing close contact between the conceptus and the uterine epithelium. Kallikrein, a serine protease, may be involved with the timing of conceptus expansion and placental attachment to the uterine surface. The objective of this study was to evaluate kallikrein enzymatic activity, protein, and gene expression in the pig during the estrous cycle and early pregnancy. Enzymatic activity was first detected in uterine flushings (UTF) on Day 12 of the estrous cycle and pregnancy. Activity was enhanced on Day 12 of pregnancy compared to that in cyclic gilts, with a reversal of increased kallikrein activity in cyclic compared to pregnant flushings on Day 15. Western blot analysis with antiserum to human plasma kallikrein detected a 50-kDa product similar to human plasma kallikrein from Day 10 to Day 15 of the estrous cycle and pregnancy. Kallikrein enzymatic activity in UTF was associated with the presence of a 23-kDa reactive product. Gene expression of kallikrein as determined by reverse transcription-polymerase chain reaction indicated the presence of kallikrein mRNA in the porcine endometrium and conceptuses. Results indicate that an increase in uterine luminal kallikrein activity occurs during the estrous cycle at a period that corresponds to rapid conceptus elongation during pregnancy of the pig. The present information suggests that kallikrein may play a role in opening the window for establishment of pregnancy in the pig. PMID- 10529270 TI - Androgen and estrogen metabolism in the reproductive tract and accessory sex glands of the domestic boar (Sus scrofa). AB - Steroid metabolism in target tissues has relevance in assessing biological response. We have investigated the metabolism of testosterone and estrogens in the reproductive tract and accessory sex glands in the boar. Seminal vesicles were taken from four 6-mo-old animals; and seminal vesicles, prostate, vas deferens, and regions of the epididymis were taken from two mature boars (10 and 24 mo old). Tissues were incubated in 5 ml medium (TC-199) at 34 degrees C under 5% CO(2) and 95% air for 2 h with (3)H-labeled testosterone, estrone, and estradiol-17beta. Aliquots of spent media were taken to measure radioactivity before separation of unconjugated and conjugated steroids on Waters C(18) Sep-Pak cartridges. Sulfoconjugated steroids and glucuronidates were recovered in series from C(18) cartridges after solvolysis and enzyme hydrolysis, respectively. Profiles of metabolites for free and hydrolyzed fractions were obtained from gradient HPLC with acetonitrile:water on a reversed-phase C(18) column. No clear evidence of conjugation was seen for testosterone metabolites. 5alpha Dihydrotestosterone was the principal metabolite, but the amounts formed depended on the source, with little from the epididymal tissues and seminal vesicles, but greater quantities from the vas deferens (>25%) and prostate (>30%). The most noteworthy feature of estrogen metabolism was the extent of conjugation by all tissues. Almost all radioactivity in the conjugate fractions for the epididymis and vas was present as sulfates. Glucuronidates were seen for the prostate and were the dominant form of conjugation (about 60%) for the seminal vesicles. A striking parallel existed for the profiles of estrogen metabolites from all tissues for unconjugated and hydrolyzed fractions. Only in quantitative terms were some distinctions noted. These overall findings underscore a need to consider local metabolism of steroid hormones in target tissues of the male reproductive system. PMID- 10529271 TI - Cellular localization of estrogen receptor beta messenger ribonucleic acid in cynomolgus monkey reproductive organs. AB - There is now evidence that the recently identified estrogen receptor (ER) beta is more widely distributed in the body than is ER-alpha. In order to gain more information about the role of ER-beta in reproduction, we have investigated by in situ hybridization the localization of mRNA expression of this ER subtype in adult monkey reproductive organs. In the pituitary gland of animals of both sexes, in both the anterior and intermediate lobes, a large number of cells were positive. No specific signal was observed in the posterior lobe. In the ovary, granulosa cells in primary and growing follicles highly expressed ER-beta mRNA. The theca interna cells were also strongly labeled. In some corpora lutea, the luteal cells were strongly labeled, while in other ones, the signal was weak. A hybridization signal was also detected in the ovarian surface epithelium. In the uterus, ER-beta mRNA was found in high concentration in glandular epithelial cells and stromal cells of the endometrium, while weaker labeling was consistently observed in smooth muscle cells. In the mammary gland, labeling was detected in the epithelial cells of acini and interlobular ducts as well as stromal cells. In the testis, specific labeling was detected in the seminiferous epithelium whereas the interstitial Leydig cells were unlabeled. Although it was not possible to clearly identify all the positive cell types, it appears that Sertoli cells as well as the vast majority of germinal cells express ER-beta mRNA. In the prostate, the secretory epithelial cells exhibited a specific autoradiographic reaction while the stromal cells did not show mRNA expression. The epithelial cells of the prostatic urethra showed a strong labeling. No hybridization signal was detected in the seminal vesicles. It then appears quite clear that ER-beta is expressed in a cell-specific manner in all the monkey reproductive organs studied. In the female, the wide distribution of these receptors in the ovary and uterus suggests that ER-beta may play an important role in the mediation of the known effects of estrogen in reproduction functions. In the male testis and prostate, ER-beta has been found in cells that contain very little or no ER-alpha. The role of circulating or locally produced estrogens in the male reproductive system remains to be clarified. PMID- 10529273 TI - Cyclic modulation of integrin expression in bovine endometrium. AB - Integrins are heterodimeric glycoproteins involved in cell-cell and cell extracellular matrix adhesion. In this study, the spatial and temporal distribution of selected integrins and extracellular matrix proteins was determined in bovine endometrium from cycling and ovariectomized animals using indirect immunohistochemistry. The expression of integrins alpha(6) and alpha(v)beta(3) was estrous cycle-dependent. Strong immunostaining for alpha(v)beta(3) occurred in the basement membrane region of intercaruncular luminal epithelium except on Day 16 (P<0.05). Staining of subepithelial stromal cells declined in diestrous samples (P<0.05). In all samples, there was reduced alpha(v)beta(3) reactivity in the caruncles. Staining for alpha(6) decreased in the epithelial basement membrane at proestrus through estrus (Days 18-0). Expression of integrin subunits alpha(3) and alpha(4) was cycle-independent. Moderate staining for alpha(3) was detected on epithelium and alpha(4) was present on stromal cells. The distribution of beta(1) suggested dimerization with alpha(3), alpha(4), and alpha(6). Laminin was detected in the epithelial and vasculature basement membranes. Collagen IV was present in the glandular epithelium basement membrane and subepithelial stromal cells, whereas fibronectin was found only in the stroma. Estrous cycle-dependent distribution and expression of alpha(v)beta(3) and alpha(6) suggest their regulation by ovarian steroids, growth factors, and prostaglandins. PMID- 10529272 TI - Round spermatid-specific transcription of the mouse SP-10 gene is mediated by a 294-base pair proximal promoter. AB - Spermiogenesis is the terminal phase of male germ cell differentiation during which haploid spermatids engage in coordinate expression of a number of testis specific genes, including those specifying acrosomal proteins. To begin to understand the transcriptional regulation during acrosomal biogenesis, we initiated promoter analysis of the gene encoding the acrosomal protein SP-10. SP 10 was previously shown to be transcribed within Golgi-phase round spermatids in the human. The present study characterizes SP-10 gene expression during spermiogenesis in the mouse and identifies regions of the mouse SP-10 (mSP-10) promoter that are capable of driving round spermatid-specific transcription in vivo. Expression of mSP-10 mRNA was initiated in early round spermatids coincident with acrosomal biogenesis and was terminated prior to nuclear elongation. The core promoter of mSP-10 lacked a TATA box but contained a canonical initiator (Inr) element surrounding the transcription start site. Using transgenic mice, we showed that the -408 to +28-base pair (bp) or the -266 to +28 bp mSP-10 5' flanking region is sufficient to direct round spermatid-specific expression of a green fluorescent protein reporter gene. On the other hand, the 91 to +28-bp mSP-10 gene fragment lacked promoter activity in vivo. This is the first functional characterization of a testis-specific gene promoter active in early round spermatids. PMID- 10529274 TI - Incubation temperature influences sex-steroid levels in juvenile red-eared slider turtles, Trachemys scripta, a species with temperature-dependent sex determination. AB - Incubation temperature determines gonadal sex in the red-eared slider turtle, Trachemys scripta. However, little is known about the long-term effects of incubation temperature on traits other than gonadal sex in this species. To investigate the hypothesis that incubation temperature (independent of gonadal sex) influences sex steroid levels after hatching, we incubated eggs of the red eared slider turtle at three temperatures (26, 28.6, and 31 degrees C). We then measured plasma levels of dihydrotestosterone, estradiol, progesterone, and testosterone in 6-wk-old males from 26 degrees C and 28.6 degrees C eggs, and in 6-wk-old females from 28.6 degrees C and 31 degrees C eggs. We found that dihydrotestosterone levels were not influenced by incubation temperature or gonadal sex. However, progesterone levels were significantly higher in males from 26 degrees C eggs than in males from 28.6 degrees C eggs. In contrast, testosterone levels did not differ between males from 26 degrees C versus males from 28.6 degrees C eggs, but they were significantly higher in females from 28.6 degrees C than in females from 31 degrees C eggs. Progesterone and testosterone levels did not differ between males and females from 28.6 degrees C eggs. Temperature also influenced estradiol levels in both sexes, but the effects were enigmatic. We conclude that incubation temperature has lasting effects on sex steroid levels even after hatching. PMID- 10529275 TI - Differential regulation of prostaglandin EP and FP receptors in pregnant sheep myometrium and endometrium during spontaneous term labor. AB - In the present study, we characterized the mRNA abundance of prostaglandin E(2) receptor subtypes (EP1 and EP3, which stimulate excitatory responses; EP2 and EP4, which stimulate inhibitory responses) and the FP receptor in pregnant sheep myometrium and endometrium in relation to parturition. Myometrial and endometrial poly(A) RNA was extracted from control ewes at 143-147 days gestational age (dGA, n = 6) and from ewes in spontaneous term labor at 145-147 dGA (n = 6), and was subjected to Northern blot analysis for FP, EP1, EP2, EP3, and EP4 mRNA. Myometrial EP3, EP4, and FP mRNA abundance increased during labor (P<0.05); EP2 mRNA did not change. EP1 mRNA was not detectable in the myometrium. Endometrial EP2 and EP4 mRNA remained unchanged during labor. EP3 mRNA was expressed at a very low level, and EP1 and FP mRNA were not detected in endometrium in any animals studied. In conclusion, there is differential expression in myometrium and endometrium of EP subtypes and FP receptor in relation to labor. Increases in EP3 and FP, together with increased prostaglandin production from intrauterine tissues, may lead to the switch in the myometrial contraction pattern that occurs during labor. These differences within and between myometrium and endometrium may result from different anatomical location, such as longitudinal or circular layers of myometrium, or vascular location. PMID- 10529277 TI - Hormone secretion in the asian elephant (Elephas maximus): characterization of ovulatory and anovulatory luteinizing hormone surges. AB - In the elephant, two distinct LH surges occur 3 wk apart during the nonluteal phase of the estrous cycle, but only the second surge (ovLH) induces ovulation. The function of the first, anovulatory surge (anLH) is unknown, nor is it clear what regulates the timing of these two surges. To further study this observation in the Asian elephant, serum concentrations of LH, FSH, progesterone, inhibin, estradiol, and prolactin were quantified throughout the estrous cycle to establish temporal hormonal relationships. To examine long-term dynamics of hormone secretion, analyses were conducted in weekly blood samples collected from 3 Asian elephants for up to 3 yr. To determine whether differences existed in secretory patterns between the anLH and ovLH surges, daily blood samples were analyzed from 21 nonluteal-phase periods from 7 Asian elephants. During the nonluteal phase, serum LH was elevated for 1-2 days during anLH and ovLH surges with no differences in peak concentration between the two surges. The anLH surge occurred 19.9+/-1.2 days after the end of the luteal phase and was followed by the ovLH surge 20.8+/-0.5 days later. Serum FSH concentrations were highest at the beginning of the nonluteal phase and gradually declined to nadir concentrations within 4 days of the ovLH surge. FSH remained low until after the ovLH surge and then increased during the luteal phase. Serum inhibin concentrations were negatively correlated with FSH during the nonluteal phase (r = -0.53). Concentrations of estradiol and prolactin fluctuated throughout the estrous cycle with no discernible patterns evident. In sum, there were no clear differences in associated hormone secretory patterns between the anLH and ovLH surge. However, elevated FSH at the beginning of the nonluteal phase may be important for follicle recruitment, with the first anLH surge acting to complete the follicle selection process before ovulation. PMID- 10529276 TI - Variations of light and temperature regimes and resulting effects on reproductive parameters in medaka (Oryzias latipes). AB - In seasonally breeding fish species, altered fecundity, fertility, and spawning interval are associated with changes in environmental cues such as temperature and photoperiod. To determine quantitative impact of these cues on a suite of reproductive endpoints, groups of medaka (Oryzias latipes; two breeding pairs per group) were subjected to varying photoperiod and temperature regimes. Embryo production ceased after photoperiod reduction from 16L:8D to 8L:16D (at 25 degrees C). A severe decline in production was observed after a temperature decrease of 10 degrees C (25 degrees C to 15 degrees C [16L:8D]). Under reduced photoperiod, histologic analysis showed no mature ova and moderate oocyte atresia in all individuals. However, reduced temperature (15 degrees C) produced only mild oocyte atresia and fewer mature ova. Under both reduced photoperiod and reduced temperature regimes, mature spermatozoa were observed. Offspring viability, along with spawning interval, were not affected by photoperiod reduction. Temperature change had no effect on offspring viability but caused an increase in spawning interval. A shortened photoperiod profoundly affected medaka reproduction, whereas decreased temperature reduced, but did not arrest, fertility; reduced photoperiod decreased fecundity. These findings have important implications for culture of medaka as well as use of this teleost model for reproductive toxicology studies. PMID- 10529278 TI - Cellular and subcellular localization of six retinoid receptors in rat testis during postnatal development: identification of potential heterodimeric receptors. AB - Vitamin A is required in the testis for germ cell development. It acts through two families of retinoid receptors, retinoic acid receptors (RAR) and retinoid X receptors (RXR), each with three subtypes alpha, beta, and gamma. These receptors are postulated to dimerize and regulate the transcription of retinoid-responsive genes that are crucial for germ cell development. In this study, we determined the cellular and subcellular localization of six retinoid receptors in the developing rat testis to identify the specific cellular sites and times of receptor expression. Immunohistochemical results revealed the expression of RARalpha, RARbeta, RXRalpha, and RXRgamma proteins in somatic and germ cells throughout postnatal development. In contrast, the expression of RARgamma and RXRbeta did not increase until 30-35 days of age in somatic cells from the testis. Interestingly, RARalpha and RXRalpha had a similar subcellular localization pattern in Sertoli cells throughout postnatal testis development, while RARalpha and RXRgamma were both present in the nucleus of spermatocytes and elongating spermatids. These results suggest that RARalpha may potentially dimerize with RXRalpha in Sertoli cells and with RXRgamma in germ cells. In addition, we demonstrate that the only RAR in the nucleus of early meiotic germ cells is RARalpha. PMID- 10529279 TI - Collagenase and gelatinase messenger ribonucleic acid expression and activity during follicular development in the rat ovary. AB - Metalloproteinases are members of a family of proteinases that remodel the extracellular matrix throughout the body. To test the hypothesis that metalloproteinases are regulated by gonadotropin-induced changes during follicular growth, rats were injected with eCG (20 IU, s.c.), and ovaries and serum were collected at the time of eCG administration (0 h) and at 6, 12, 24, 36, or 48 h later for analysis of metalloproteinase mRNA expression, metalloproteinase activity, and steroidogenesis. Serum estradiol levels increased from 18.9 pg/ml at 0 h to 503.8 pg/ml at 48 h. Analysis of mRNA expression was performed for collagenase-3, 72-kDa gelatinase, and 92-kDa gelatinase (n = 3-4). For collagenase-3, eCG stimulated a 32-fold increase in collagenase-3 mRNA at 48 h after eCG injection as compared to that in ovaries collected at the time of eCG administration (i.e., 0-h control). The mRNA levels for 72-kDa gelatinase were 2.8-fold compared to 0 h at 36 h after eCG treatment and returned to control levels by 48 h after gonadotropin treatment. Levels of the 92-kDa mRNA expression peaked at 24 h (4. 2-fold compared to 0 h) and returned to control levels by 36 h. Gel zymography revealed 3 gelatinolytic bands corresponding to the gelatinases of approximately 72 kDa, 92 kDa, and 105 kDa. Analysis of metalloproteinase activity as the degradation of collagen or gelatin per ovary showed an increase in gelatinolytic and collagenolytic activity between 12 and 48 h after eCG treatment. In summary, these findings demonstrate that the gonadotropin induction of folliculogenesis results in changes in the metalloproteinases that may be responsible for extracellular matrix remodeling associated with follicular growth. PMID- 10529280 TI - Prostaglandin f(2alpha) concentrations, fatty acid profiles, and fertility in lipid-infused postpartum beef heifers. AB - Effects of lipid infusion into postpartum (PP) beef heifers on plasma concentrations of linoleic acid and prostaglandin (PG) F(2alpha) metabolite (PGFM), days to first estrus, and subsequent pregnancy rate were examined. Treatments (n = 5 per group) of 1 L intralipid (20% soybean oil; IL), 1 L 50% dextrose (DEXT; isocaloric to IL), 0.5 L intralipid (0.5 IL), and 1 L physiological saline (SAL) were infused i.v. over 4 h on each of Days 7 through 11 PP. Capacity of the uterus to produce PG was evaluated after i.v. injection of 150 IU of oxytocin (OT) to IL- and DEXT-treated heifers Day 12 PP. Change in plasma concentrations of PGFM from 0 to 4 h was greater for IL-treated heifers than for heifers given other treatments on Day 7 (P = 0.04) and on Day 11 (P = 0.01), but not on Day 9 (P>0.10). Plasma linoleic acid on Day 11 and OT-induced release of PGFM on Day 12 were greater in IL-treated heifers compared with DEXT treated heifers (P<0.06 and P = 0.01, respectively). There were no significant differences among treatments for mean days to first estrus or pregnancy rate. Infusion of lipid increased systemic concentrations of linoleic acid and increased the capacity of PP heifers to produce uterine PGF(2alpha) as indicated by plasma PGFM concentration after OT injection. PMID- 10529281 TI - Development and characterization of immortalized ovine endometrial cell lines. AB - The objective of this study was to generate immortalized endometrial epithelial and stromal cell lines from the ovine uterus. Luminal (LE) and glandular epithelial (GE) cells and stromal (ST) cells were enzymatically isolated from the uterus of a Day 5 cyclic ewe (estrus on Day 0), and primary cultures were immortalized by transduction with a retroviral vector (LXSN-16E6E7) packaged by the amphotropic fibroblast line PA-317. Cells having integrated the vector were selected by resistance to the neomycin analogue G418 (0.6-0.8 mg/ml). Surviving cells were maintained in complete culture medium containing G418 (0.1 mg/ml) and subcultured for more than 40 passages. Phase-contrast microscopy revealed that LE and GE cells exhibited a cobblestone morphology whereas immortalized ST cells were spindle shaped. The epithelial origin of LE and GE was confirmed by positive cytokeratin immunostaining, and ST cells were vimentin positive. All cell lines were negative for smooth muscle alpha-actin staining. Western blot analyses of cell extracts revealed the presence of signal transducers and activators of transcription (STAT) proteins 1, 2, and 3. In the LE cells, interferon tau (IFNtau) induced nuclear translocation of STAT proteins 1 and 2 and up-regulated several IFN-inducible genes, including STATs 1, 2, and 3 and ubiquitin cross reactive protein (UCRP/ISG17). In the LE cell line, IFN regulatory factor one was transiently up-regulated and then down-regulated by IFNtau. Immunostaining revealed the presence of nuclear estrogen receptor and progesterone receptor in all cell lines. These ovine endometrial cell lines provide useful in vitro model systems for the study of hormone and cytokine action, signal transduction pathways, cell-cell interactions, and gene expression in specific cell types of the ovine endometrium. PMID- 10529282 TI - Transplantation of germ cells from rabbits and dogs into mouse testes. AB - Spermatogonial stem cells of a fertile mouse transplanted into the seminiferous tubules of an infertile mouse can develop spermatogenesis and transmit the donor haplotype to progeny of the recipient mouse. When testis cells from rats or hamsters were transplanted to the testes of immunodeficient mice, complete rat or hamster spermatogenesis occurred in the recipient mouse testes, albeit with lower efficiency for the hamster. The objective of the present study was to investigate the effect of increasing phylogenetic distance between donor and recipient animals on the outcome of spermatogonial transplantation. Testis cells were collected from donor rabbits and dogs and transplanted into testes of immunodeficient recipient mice in which endogenous spermatogenesis had been destroyed. In separate experiments, rabbit or dog testis cells were frozen and stored in liquid nitrogen or cultured for 1 mo before transplantation to mice. Recipient testes were analyzed, using donor-specific polyclonal antibodies, from 1 to >12 mo after transplantation for the presence of donor germ cells. In addition, the presence of canine cells in recipient testes was demonstrated by polymerase chain reaction using primers specific for canine alpha-satellite DNA. Donor germ cells were present in the testes of all but one recipient. Donor germ cells predominantly formed chains and networks of round cells connected by intercellular bridges, but later stages of donor-derived spermatogenesis were not observed. The pattern of colonization after transplantation of cultured cells did not resemble spermatogonial proliferation. These results indicate that fresh and cryopreserved germ cells can colonize the mouse testis but do not differentiate beyond the stage of spermatogonial expansion. PMID- 10529283 TI - Pronuclear location before the first cell division determines ploidy of polyspermic pig embryos. AB - Polyspermy occurs frequently in the fertilization of mammalian eggs, but little is known about whether polyspermic eggs have developmental ability in vitro or in vivo. We previously reported that poly-pronuclear (PPN; 3 or more pronuclei) pig eggs developed normally to the blastocyst stage despite having fewer inner cell mass cell numbers as compared to blastocysts derived from two-pronuclear (2PN) eggs. Here it is shown that most PPN pig eggs have abnormal cleavage patterns (having 3 or more cells) in the first cell division and retarded development of pronuclei prior to syngamy as compared to 2PN eggs. Most blastocysts (14 of 18) that developed from PPN eggs showed abnormal ploidy (were haploid, triploid, and tetraploid) whereas 20 of 22 blastocysts derived from 2PN embryos were diploid. The size and morphology of most Day 40 fetuses that developed from PPN eggs appeared to be normal. Of 8 Day 40 fetuses analyzed, 1 was triploid (XXY) and another was a mosaic with both diploid (XX) and tetraploid cells (frequency of less than 10%, XXXX), and the others were diploid. Anomalies of chromosomal composition were not detected in these fetuses. Five live piglets and one dead piglet were born from two recipients of PPN eggs. It is proposed that not all pronuclei of PPN pig eggs participate in syngamy, resulting in diploid cells in the conceptus. Our data suggest that there are two types of pronuclei location in polyspermic pig eggs and that the resulting ploidy is determined at the zygote stage before the first cell division according to pronuclear location. PMID- 10529284 TI - Long-term effects of postovulatory aging of mouse oocytes on offspring: a two generational study. AB - Aims of this study were to analyze the long-term effects of postovulatory aging of mouse oocytes on 1) reproductive traits of parental (F(0)) and first (F(1)) generation females (pregnancy rate, gestation length, litter size, perinatal death, and sex ratio of offspring) and 2) developmental and behavioral variables of F(1) and second-generation (F(2)) offspring (birth weight and weight gain during preweaning development, postnatal day of attainment of immediate righting, spontaneous motor activity, and passive and active conditioned learning ability). Hybrid (C57BL/6JIco x CBA/JIco) females were artificially inseminated at 13 h (control group) or 22 h (oocyte-aged group) after GnRH injection. Experimental (oocyte-aged group) F(0) females exhibited lower pregnancy rate, shortened gestation length, decreased litter size, higher perinatal death of their pups, and increased percentage of male offspring compared to control F(0) females. Postovulatory aging of oocytes was also associated with increased number of growth-retarded pups, delayed development of the righting reflex, and higher spontaneous motor activity and emotionality of F(1) offspring. Postovulatory aging of F(0) oocytes did not affect birth weight, weight gain during preweaning development, passive and active conditioned learning ability of F(1) offspring, or reproductive traits of F(1) females or developmental and behavior variables of F(2) offspring. PMID- 10529285 TI - Induction of thumbtack sperm during coculture with oviduct epithelial cell monolayers in a marsupial, the brushtail possum (Trichosurus vulpecula). AB - A reorientation of the sperm head so that it is perpendicular to the sperm tail (i.e., T-shape or thumbtack) is considered an indicator of sperm capacitation in the Australian marsupial the brushtail possum (Trichosurus vulpecula). This study describes a method of oviduct epithelial cell monolayer and sperm coculture in the brushtail possum to obtain a high percentage of thumbtack sperm. The oviduct epithelial cell (OEC) monolayers were prepared in vitro from the isthmal and ampullary segments of eCG- and LH-primed brushtail possum oviducts. Coculture experiments demonstrated that cauda epididymidal sperm from the brushtail possum attached equally to the OEC monolayers derived from the isthmal and ampullary segments of the oviduct. After 2 h of coculture, a large number of sperm attached to OEC monolayers (ampulla, 60.1+/-4.7% and isthmus, 63.1+/-5.7%) as well as to controls (tracheal epithelial cell monolayer, 46.2+/-3.7%; Matrigel, 57.4+/-7.7%; plastic, 29.2+/-3.2%). After 6 h, fewer sperm were attached to tracheal epithelial cell monolayers (1.2+/-0.2%; P<0.01) and Matrigel (10.2+/-2.5%; P<0.01), compared to those attached to ampullary and isthmal OEC monolayers (37.9+/-7.2% and 44.6+/-2.2%, respectively), and none were attached to the plastic surface. Fewer sperm were released from the ampullary and isthmal OEC monolayers compared to those from controls (P<0.05). At 6 h of coculture with ampullary and isthmal OEC, the percentage motility of both attached and unattached spermatozoa was maintained at 40-50%, which was higher (P<0.05) than in controls. Progressive motility of unattached sperm was maintained at about 2 (on an arbitrary scale of 1-5) and was not different among treatments until 6 h. More than 60-70% sperm were viable at 6 h of coculture in all the treatments. Coculture of brushtail possum epididymal sperm with OEC monolayers transformed 60% of motile streamlined spermatozoa to thumbtack orientation at 2 h compared to approximately 25% in controls. No acrosomal modifications were induced in spermatozoa in any of the treatments. This study has demonstrated a role of the oviduct in transforming a large number of sperm from a streamlined to thumbtack orientation, which may have relevance in sperm capacitation and fertilization in this species. PMID- 10529286 TI - Nuclear and cytoplasmic maturation of mouse oocytes after treatment with synthetic meiosis-activating sterol in vitro. AB - Synthetically produced meiosis-activating sterol, a sterol originally derived from follicular fluid (FF-MAS), induces meiotic maturation of mouse oocytes in vitro. We therefore compared FF-MAS-induced maturation of naked mouse oocytes arrested in prophase I by either hypoxanthine (Hx) or forskolin (Fo) with spontaneous maturation of naked oocytes. FF-MAS-treated oocytes overcame the meiotic block by Hx or Fo, although germinal vesicle breakdown was delayed by 11 h and 7 h, respectively. We also investigated the influence of FF-MAS on chromosome, microtubule, and ultrastructural dynamics in Hx-cultured oocytes by immunocytochemistry and electron microscopy. Similarly to spontaneously matured oocytes, chromosomes became aligned, a barrel-shaped spindle formed, and overall organelle distribution was normal in FF-MAS-matured oocytes. The number of small cytoplasmic asters was elevated in FF-MAS-treated oocytes. Although the number of cortical granules (CGs) was similar to that in spontaneously matured oocytes, the overall distance between CGs and oolemma was increased in the FF-MAS group. These observations suggest that the initiation of meiotic maturation in FF-MAS-treated oocytes in the presence of high cAMP levels leads to a delayed but otherwise normal nuclear maturation. FF-MAS appears to improve oocyte quality by supporting microtubule assembly and by delaying CG release, which is known to contribute to reduced fertilization. PMID- 10529288 TI - The LASIK procedure. First the flap, then the zap. PMID- 10529287 TI - Laser eye surgery. Eyes wide open about LASIK. PMID- 10529289 TI - Fibromyalgia syndrome. Feeling more pain. PMID- 10529290 TI - Viagra. Why aren't more men taking it? PMID- 10529291 TI - Diet. The glycemic index. PMID- 10529292 TI - By the way, doctor... I'm 78 and have more and more difficulty urinating. I get up three times a night - sometimes more - to go to the bathroom and it takes me a while to start urinating and even longer to stop. I know that this is probably a prostate problem, but I don't want to have surgery. What can I do? PMID- 10529293 TI - High-tech dining: is America ready? PMID- 10529294 TI - Treating prostate cancer. Part III: surgery. PMID- 10529295 TI - Obesity: can a new drug help? PMID- 10529296 TI - Medical memo. Preoperative tests. PMID- 10529298 TI - Cloning and characterization of the 23S RNA pseudouridine 2633 synthase from bacillus subtilis PMID- 10529297 TI - By the way, doctor... I have diabetes, and my doctor does a blood test every now and then to see whether my blood sugar levels are under control. But I have always been a little confused by the results. The number I received most recently was 8, and the doctor said that 5 or less was normal. What does this mean? PMID- 10529299 TI - A comparison of logistic regression to decision tree induction in the diagnosis of carpal tunnel syndrome. AB - This paper aims to compare and contrast two types of model (logistic regression and decision tree induction) for the diagnosis of carpal tunnel syndrome using four ordered classification categories. Initially, we present the classification performance results based on more than two covariates (multivariate case). Our results suggest that there is no significant difference between the two methods. Further to this investigation, we present a detailed comparison of the structure of bivariate versions of the models. The first surprising result of this analysis is that the classification accuracy of the bivariate models is slightly higher than that of the multivariate ones. In addition, the bivariate models lend themselves to graphical analysis, where the corresponding decision regions can easily be represented in the two-dimensional covariate space. This analysis reveals important structural differences between the two models. PMID- 10529300 TI - Performances of hill-type and neural network muscle models-toward a myosignal based exoskeleton. AB - Muscle models are the essential components of any musculoskeletal simulation. In addition, muscle models which are incorporated in neural-based prosthetic and orthotic devices may significantly improve their performance. The aim of the study was to compare the performances of two types of muscle models in terms of predicting the moments developed at the human elbow joint complex based on joint kinematics and neuromuscular activity. The performance evaluation of the muscle models was required to implement them in a powered myosignal-driven exoskeleton (orthotic device). The experimental setup included a passive exoskeleton capable of measuring the joint kinematics and dynamics in addition to the muscle myosignal activity (EMG). Two types of models were developed and analyzed: (i) a Hill-based model and (ii) a neural network. The task, which was selected for evaluating the muscle models performance, was the flexion-extension movement of the forearm with a hand-held weight. For this task the muscle model inputs were the normalized neural activation levels of the four main flexor-extensor muscles of the elbow joint, and the elbow joint angle and angular velocity. Using this inputs, the muscle model predicted the moment applied on the elbow joint during the movement. Results indicated a good performance of the Hill model, although the neural network predictions appeared to be superior. Relative advantages and shortcomings of both approaches were presented and discussed. PMID- 10529301 TI - Classification of asthmatic breath sounds: preliminary results of the classifying capacity of human examiners versus artificial neural networks. AB - For continuous monitoring of the respiratory condition of patients, e.g., at the intensive care unit, computer assistance is required. Existing mechanical devices, such as the peak expiratory flow meter, provide only with incidental measurements. Moreover, such methods require cooperation of the patient, which at, e.g., the ICU is usually not possible. The evaluation of complicated phenomena such as asthmatic respiratory sounds may be accomplished by use of artificial neural networks. To investigate the merit of artificial neural networks, the capacities of neural networks and human examiners to classify breath sounds were compared in this study. Breath sounds were in vivo recorded from 50 school-age children with asthma and from 10 controls. Sound intervals with a duration of 20 seconds were randomly sampled from asthmatics during exacerbation, asthmatics in remission, and controls. The samples were digitized and related to peak expiratory flow. From each interval, two full breath cycles were selected. Of each selected breath cycle, a Fourier power spectrum was calculated. The so-obtained set of spectral vectors was classified by means of artificial neural networks. Humans evaluated graphic displays of the spectra. Human examiners could not clearly discriminate between the three groups by inspecting the spectrograms. Classification by self-classifying neural networks confirmed the existence of at least three classes; however, discrimination of 11 classes seemed more appropriate. Good results were obtained with supervised networks: as much as 95% of the training vectors could be classified correctly, and 43% of the test vectors. The three patient groups, as discriminated in advance, do not correspond with three sharply separated sets of spectrograms. More than three classes seem to be present. Humans cannot take up the spectral complexity and showed negative classification results. Artificial neural networks, however, are able to handle classification tasks and show positive results. PMID- 10529302 TI - Best parameters selection for wavelet packet-based compression of magnetic resonance images. AB - Transmission of compressed medical images is becoming a vital tool in telemedicine. Thus new methods are needed for efficient image compression. This study discovers the best design parameters for a data compression scheme applied to digital magnetic resonance (MR) images. The proposed technique aims at reducing the transmission cost while preserving the diagnostic information. By selecting the wavelet packet's filters, decomposition level, and subbands that are better adapted to the frequency characteristics of the image, one may achieve better image representation in the sense of lower entropy or minimal distortion. Experimental results show that the selection of the best parameters has a dramatic effect on the data compression rate of MR images. In all cases, decomposition at three or four levels with the Coiflet 5 wavelet (Coif 5) results in better compression performance than the other wavelets. Image resolution is found to have a remarkable effect on the compression rate. PMID- 10529303 TI - Identification of the occluded artery in patients with myocardial ischemia induced by prolonged percutaneous transluminal coronary angioplasty using traditional vs transformed ECG-based indexes. AB - We have studied the spatial properties of ischemic changes as induced by prolonged angioplasty and how the changes are related to different ECG indexes. Indexes based on measurements at specific points in time (ST level at J + 60 ms point, maximal T wave amplitude and position, QT interval, and QRS duration) and global indexes (based on the Karhunen-Loeve transform and applied to the QRS complex, ST-T complex, ST segment, and T wave), considering both repolarization and depolarization information, were analyzed. The changes during the occlusion period of the different indexes were used as variables in a multivariate discriminant analysis to determine which indexes showed the best discrimination of the three major occlusion sites (corresponding to LAD, RCA, and LCX coronary arteries). Occlusions in LCX artery were the most difficult to classify. With three local indexes (ST60 level measured in lead V3, T wave amplitude in I, and ST60 in III) it was possible to correctly classify 76% of patients by the occlusion site, and with three KLT-derived indexes (first-order KLT index for ST T complex in I and for QRS in leads V3 and I) 83% of correct classification was obtained. Using six indexes for local and KLT-derived indexes the correct classification was increased to 85 and 90% of patients, respectively. The use of different ECG indexes (from different intervals) on quasiorthogonal leads permitted the identification of the occluded artery in patients undergoing PTCA and may be extended to more general use. PMID- 10529304 TI - Freezing, drying, and/or vitrification of membrane- solute-water systems. AB - Membranes are often damaged by freezing and/or dehydration, and this damage may be reduced by solutes. In many cases, these phenomena can be explained by the physical behavior of membrane-solute-water systems. Both solutes and membranes reduce the freezing temperature of water, although their effects are not simply additive. The dehydration of membranes induces large mechanical stresses in the membranes. These stresses produce a range of physical deformations and changes in the phase behavior. These membrane stresses and strains are in general reduced by osmotic effects and possibly other effects of solutes-provided of course that the solutes can approach the membrane in question. Membrane stresses may also be affected by vitrification where this occurs between membranes. Many of the differences among the effects of different solutes can be explained by the differences in the crystallization, vitrification, volumetric, partitioning, and permeability properties of the solutes. PMID- 10529305 TI - Energy metabolism following prolonged hepatic cold preservation: benefits of interrupted hypoxia on the adenine nucleotide pool in rat liver. AB - The ability of brief hypothermic reperfusion (HtR) to restore hepatic energy metabolism following periods of cold hypoxic preservation was studied in isolated rat livers after storage times of 5, 10, and 24 h. In addition, investigations were performed on the effects of HtR used to restore liver oxidative metabolism in the middle of a prolonged (24 h) hypoxic preservation period. A histidine lactobionate-raffinose solution was used for the initial cold portal flush in all groups. Results showed that cold hypoxia for either 5 or 10 h yielded livers capable of similar recoveries of ATP, energy charge, and total adenine nucleotides, but that HtR after 24 h cold preservation resulted in reduced regeneration of ATP, a lower energy charge, and a fall in tissue adenine nucleotides. When livers were stored for 24 h but subjected to brief HtR after either 5 or 10 h before return to hypoxic storage, improved recoveries of the energy metabolites were seen over those recorded after 24 h hypoxia alone. The fact that these improvements were not due to an improved supply of adenine nucleotide precursors was demonstrated by studying groups which were given HtR with perfusate containing precursors of adenine nucleotides (adenosine, adenine, and inosine) after 24 h cold hypoxia. These data are consistent with the hypothesis that poor metabolic recovery after long-term hepatic cold preservation results more from decreased mitochondrial oxidative phosphorylation than from a lack of precursors for adenine nucleotide resynthesis. In addition, restoring oxidative metabolism at hypothermia for brief periods can to some extent protect final metabolic status after prolonged storage. PMID- 10529306 TI - Supercooling capacity of seeds and seedlings in Arabidopsis thaliana. AB - The influence of the water content of seeds and seedlings of Arabidopsis thaliana (Ecotype Columbia:2) on their supercooling capacity was investigated. Equilibration of the seeds to various air relative humidities resulted in final moisture contents ranging from 8 to 82% (dry weight basis). No supercooling point could be detected when the water content remained below 32.5%, and in seeds at just above this moisture level ice crystals started to form at -26 degrees C. However, cooling partly affected the germination of seeds down to a water content of 26.5%. Upon imbibition, the supercooling point of the seeds remained around 21.6 degrees C and rose sharply to -14.7 degrees C when visible germination started. It remained around -13 degrees C during the following 96 h while the water content of the seedlings increased from 155 to 870%. Hydrated seeds (above 32.5% water content), germinated seeds, and seedlings of Arabidopsis cannot survive being frozen. PMID- 10529307 TI - The effect of suboptimal growth temperature and growth phase on resistance of lactobacillus acidophilus to environmental stress AB - This paper describes the effect of growth temperature and the physiological age of Lactobacillus acidophilus CRL 639 on the development of cross resistance to different kinds of environmental stress. Exponential-phase cells growing at 25 degrees C and stationary-phase cells (8- and 11-h-old cultures at the optimal temperature of 37 degrees C) showed the highest survival and metabolic activity after challenge by freezing, heating, osmotic stress, and exposure to ethanol, peroxide, or acid. One-dimensional SDS-PAGE gels showed overexpression of three proteins (22.5, 39.6, and 49.2 kDa) at 25 degrees C, of two proteins (40.9 and 48.3 kDa) in 8-h-old cultures, and of one protein (30.3 kDa) after glucose depletion in 11-h-old cultures. These results strongly suggest the involvement of different sets of protein in the cellular processes of adaptation to environmental changes. Copyright 1999 Academic Press. PMID- 10529308 TI - Temperature regulation of glucose metabolism in red blood cells of the freeze tolerant wood frog. AB - The low-temperature metabolism of erythrocytes from the freeze-tolerant frog Rana sylvatica was investigated by (13)C and (31)P NMR spectroscopy. Erythrocytes readily took up high concentrations of the natural cryoprotectant, glucose, at both high (12 and 17 degrees C) and low (4 degrees C) temperatures but glucose was apparently not metabolized at 4 degrees C. Strong inhibition of glucose catabolism at low temperature would facilitate the maintenance of the very high concentrations of glucose (approximately 200 mM) that are accumulated to provide cryoprotection during freezing in wood frogs. Analysis of (13)C labeling of glycolytic intermediates at 4 degrees C showed mixing of label primarily in hexose (fructose) and hexose phosphate (glucose 6-phosphate, fructose 6 phosphate) pools but little label incorporation into triose phosphate intermediates. These data are consistent with a profound low-temperature-induced inhibition of phosphofructokinase (PFK). Investigations into potential PFK control mechanisms were undertaken. (31)P NMR analysis showed that the intracellular pH of erythrocytes increased from 7.0 to 7.3 as temperature decreased from 17 to 4 degrees C in a manner consistent with alphastat regulation. This change is exactly opposite to that expected if overall PFK activity was regulated by changes in cellular pH since PFK is less active at lower pH values in vitro. Other factors must, therefore, operate to regulate PFK at lower temperatures. PMID- 10529309 TI - Cryopreservation of keratinocytes in a monolayer. AB - The cryopreservation of cells in tissues is one of the major challenges in current cryobiology, especially with regard to the progressively increasing field of tissue engineering. It is very questionable whether protocols which were developed for the cryopreservation of isolated cells are also applicable for cells in more complex structures, such as tissues. As a starting point toward cryopreservation of these three-dimensional structures, the aim of this study was to find an optimum cryopreservation protocol for keratinocytes in a monolayer (two-dimensional structure). These epidermal cells can be transplanted as a monolayer grown on an appropriate matrix for the treatment of deep-dermal burns and leg ulcers. The successful cryopreservation of such transplants would offer the advantage of long-term storage and immediate availability of the transplant. In our study, the variables investigated were the cryoprotective solution and the cooling rate. In order to find a nontoxic cryoprotective agent (CPA) which could be transplanted without an additional washing step, we included hydroxyethyl starch (HES) as a possible CPA in our experimental protocol with the commonly used CPAs Me(2)SO, glycerol, and ethylene glycol. For the evaluation, the cell survival rate was determined by dye exclusion (trypan blue) and the cell metabolism was investigated by cell activity assay (alamarBlue). In conclusion, the cryopreservation protocol with 10 wt.-% HES resulted not only in the highest survival rate (72%) but also in the highest metabolic activity of the cells after thawing; comparable values for the other CPAs were: Me(2)SO, 48%; glycerol, 8%; and ethylene glycol, 10%. PMID- 10529310 TI - Temperature dependence of Kedem-Katchalsky membrane transport coefficients for mature mouse oocytes in the presence of ethylene glycol. AB - Ethylene glycol (EG) is the emerging cryoprotectant of choice for preservation of mammalian embryos but has not been widely used for oocyte preservation. Techniques for oocyte cryopreservation need to be improved before they can be incorporated into routine clinical practice. Hence the permeability characteristics of oocytes in the presence of EG have been determined in order to facilitate the design of cryopreservation protocols using this cryoprotectant. Individual mouse oocytes were held using negative pressure applied to the zona pellucida by means of a micropipet. Each oocyte was perfused with 1 ml 1.5 mol L( 1) EG at 30, 19, or 10 degrees C, a total of 10 oocytes being perfused at each temperature. The osmotic response of each oocyte before, during and after perfusion was recorded on videotape. Measurements of mean cell diameter across three axes were used to calculate oocyte volume, assuming them to be spherical, and, using mathematical modeling, values for hydraulic conductivity (L(p)) were found to be 0.91 +/- 0.05, 0.51 +/- 0.02, and 0.18 +/- 0.01 microm min(-1) atm( 1); cryoprotectant permeability (P(EG)) was 0.24 +/- 0.01, 0.09 +/- 0.005, and 0.03 +/- 0.004 microm s(-1); and reflection coefficient (sigma) was 0.98 +/- 0.005, 0.96 +/- 0.01, and 0.97 +/- 0.01 at 30, 19, and 10 degrees C, respectively. The activation energy (E(a)) of L(p) was 14. 0 kCal mol(-1) and of P(EG) was 16.4 kCal mol(-1). PMID- 10529311 TI - Membrane integrity, mitochondrial activity, ATP content, and motility of the European catfish (Silurus glanis) testicular spermatozoa after freezing with different cryoprotectants. AB - The extent of cellular damage was investigated after freeze-thawing of the European catfish testicular sperm with various cryoprotectants. The best protection was given by dimethylacetamide (10 and 15%) in a sucrose solution. Under these conditions, the percentage of cells with an intact membrane was high (90%), and the protection of the activity of the mitochondria was medium (47%). It was shown that the addition of dimethylacetamide largely increased the ATP content of the spermatozoa. It is suggested that this phenomenon is a decisive factor for the freezing resistance of European catfish testicular spermatozoa in the presence of dimethylacetamide (60% motility after thawing versus 90% before freezing). PMID- 10529312 TI - In vitro comparison of fowl sperm viability in ejaculates frozen by three different techniques and relationship with subsequent fertility in vivo. AB - A series of experiments was conducted to compare the viability of fresh fowl spermatozoa, samples suspended in three cryoprotectants (CPAs), frozen/thawed samples, and frozen/thawed samples maintained in vitro for up to 24 h. The CPAs used were glycerol (Glyc), dimethylacetamide (DMA), and dimethylformamide (DMF). Viability was assayed using two double stains, Eosin + Nigrosin or SYBR-14 + PI (propidium iodide). Semen samples examined with SYBR-14 + PI indicated significant differences in viability between fresh and ready-to-freeze preparations (fresh, 83%; Glyc, 73%; DMA, 74%; DMF, 72%; P < 0.05). In contrast, Eosin + Nigrosin did not detect any difference at this stage (fresh, 88%; Glyc, 86%; DMA, 87%; DMF, 88%; P > 0.05). The percentages of viable spermatozoa in frozen/thawed ejaculates stored in vitro for 0, 4, and 24 h were generally higher in samples treated with glycerol than in those treated with DMA or DMF, irrespective of the technique used to assess sperm viability (P < 0.05). Fertility in eggs obtained from hens inseminated with semen frozen in DMA reached levels comparable to those obtained from hens inseminated with fresh undiluted semen (88 and 93%, respectively; P > 0.05). In contrast, fertility of eggs from hens inseminated with semen frozen in DMF or glycerol was significantly lower, although still very good, than that observed in eggs from hens inseminated with semen frozen/thawed in DMA (79 and 76%, respectively; P < 0.05). Finally, the double stain SYBR-14 + PI was proven more effective than Eosin + Nigrosin to assess sperm viability in fresh, stored, and frozen fowl semen. However, additional tests (e.g., morphology, acrosomal status, motility) remain necessary to develop a working model of in vitro sperm analysis capable of revealing the fertilizing potential of fresh and frozen fowl spermatozoa. PMID- 10529313 TI - Factors affecting survival of cryopreserved oyster (Crassostrea gigas) embryos AB - A conventional two-step freezing procedure was developed and optimized in order to cryopreserve oyster embryos. The effects of cooling rate, choice of cryoprotectant, and seeding temperature on the survival of late-stage oyster embryos were examined. When these factors were optimized, improved survival rates of 78 +/- 8 and 83 +/- 7% were achieved using 2 M Me(2)SO or glycerol, respectively, as the cryoprotectant. The experimental results indicate that oyster embryos survive after freezing over a broad range of cooling rates ranging from -0.5 to -16 degrees C/min. Me(2)SO, glycerol, propylene glycol, and ethylene glycol may be used as cryoprotectants for the cryopreservation of oyster embryos. Copyright 1999 Academic Press. PMID- 10529314 TI - Unstable angina: the breakthrough. PMID- 10529315 TI - Specific antithrombins in the context of current treatment for acute coronary syndromes. PMID- 10529316 TI - New trials of LMW heparins - light and heavy weight as good on short but what about longer distances? PMID- 10529317 TI - Atrial fibrillation and mortality. PMID- 10529318 TI - Plant lipids that lower serum cholesterol. PMID- 10529319 TI - Depression and heart failure--not yet a target for therapy? PMID- 10529320 TI - Different benefits, different risks, equal cost. PMID- 10529321 TI - Improving outcome in acute coronary syndromes - as good as it gets? PMID- 10529322 TI - Evaluation of antiarrhythmic drug efficacy in patients with an ICD. Unlimited potential or replete with complexity and problems? AB - A report from a Study group, proposed by A. J. Camm, London, of the Working Groups on Arrhythmias and Cardiac Pacing of the European Society of Cardiology; co-sponsored by the North American Society of Pacing and Electrophysiology. The Study Group was convened on 29 August 1997 at Saltsjobaden, near Stockholm. The meeting was chaired by A. J. Camm, London, and C. M. Pratt, Houston. Based on the presentation and discussions, a first draft of the documents was prepared by C. Pratt and J. Camm which was then circulated to all members three times for their review. All members of the Study Group approved the final manuscript. This report represents the opinion of the members of this Study Group and does not necessarily reflect the official position of either society.The meeting of the Study Group was made possible by unrestricted educational grants from Medtronic, Guidant, Proctor & Gamble, Berlex and Sanofi.Also, presented, in part, at the Cardio-Renal Drugs Advisory Board meeting of the Food and Drug Administration, Bethesda, Maryland, on 30 April 1999. PMID- 10529323 TI - Comparison of two treatment durations (6 days and 14 days) of a low molecular weight heparin with a 6-day treatment of unfractionated heparin in the initial management of unstable angina or non-Q wave myocardial infarction: FRAX.I.S. (FRAxiparine in Ischaemic Syndrome). AB - AIM: To assess the benefit of short-term low molecular weight heparin nadroparin compared with unfractionated heparin in unstable angina or non-Q wave myocardial infarction patients and to determine whether a longer, 2-week low molecular weight heparin regimen would offer additional clinical benefit. PATIENTS, METHODS AND RESULTS: This was a multicentre, prospective, randomized, double-blind study in three parallel groups, involving 3468 patients. Patients received one of three treatment regimens: the unfractionated heparin group received an intravenous bolus of unfractionated heparin 5000 IU, followed by an activated partial thromboplastin time adjusted infusion of unfractionated heparin for 6+/-2 days; the nadroparin 6 group received an intravenous bolus of nadroparin 86 anti-Xa IU. kg(-1), followed by twice daily subcutaneous injections of nadroparin 86 anti-Xa IU. kg(-1)for 6+/-2 days, and the nadroparin 14 group received an intravenous bolus of nadroparin 86 anti-Xa IU. kg(-1), followed by twice daily subcutaneous injections of nadroparin 86 anti-Xa IU. kg(-1)for 14 days. No statistically significant differences were observed between the three treatment regimens with respect to the primary outcome (cardiac death, myocardial infarction, refractory angina, or recurrence of unstable angina at day 14). The absolute differences between the groups in the incidence of the primary outcome were: -0.3% (P=0.85) for the nadroparin 6 group vs the unfractionated heparin group and +1.9% (P=0.24) for the nadroparin 14 group vs the unfractionated heparin group. Furthermore, there were no significant intergroup differences regarding any of the secondary efficacy outcomes. However, there was an increased risk of major haemorrhages in the nadroparin 14 group compared with unfractionated heparin (3.5% vs 1.6%;P=0.0035). CONCLUSIONS: Treatment with nadroparin for 6+/-2 days provides similar efficacy and safety to treatment with unfractionated heparin, for the same period, in the therapeutic management of acute unstable angina or non-Q wave myocardial infarction, and may be easier to administer. A prolonged regimen of nadroparin (14 days) does not provide any additional clinical benefit. PMID- 10529324 TI - Comparison of the predictive value of ST segment elevation resolution at 90 and 180 min after start of streptokinase in acute myocardial infarction. A substudy of the hirudin for improvement of thrombolysis (HIT)-4 study. AB - AIMS: Previous studies revealed that >/=70% or <30% ST segment elevation resolution 180 min after the start of thrombolysis is a strong predictor of either favourable or poor outcome. The aim of this study was to compare the prognostic value of ST segment elevation resolution at 90 and 180 min after the start of streptokinase infusion. METHODS AND RESULTS: The Hirudin for Improvement of Thrombolysis (HIT)-4 study of 1208 patients compared streptokinase therapy in conjunction with either r-hirudin or heparin. Complete ST segment elevation resolution (>/=70%) at 90 and 180 min identified 25% and 50%, respectively, of all patients with a 30 day cardiac mortality of less than 2%. Forty-four percent of patients had no ST segment elevation resolution (<30%) at 90 min and the 30 day cardiac mortality was 7.3%. At 180 min, the no ST segment elevation resolution group decreased to 15% of all patients while the mortality risk increased to 13.6%. CONCLUSIONS: ST segment elevation resolution is a useful tool for early risk stratification and the strategy of rescue angioplasty. Complete ST segment elevation resolution within 180 min of the start of streptokinase therapy indicates excellent survival prospects in 50% of patients. A half of these low risk patients can be identified at 90 min. A high risk group appears to be best characterized by no ST segment elevation resolution at 180 min rather than at 90 min. PMID- 10529325 TI - Assessment of coronary angiograms prior to and after treatment with abciximab, and the outcome of angioplasty in refractory unstable angina patients. Angiographic results from the CAPTURE trial. AB - BACKGROUND: The CAPTURE study (c 7E3 A nti P latelet T herapy in U nstable Re fractory angina) was designed to assess outcome in patients with refractory angina undergoing angioplasty, receiving either abciximab or placebo. METHODS: One thousand two hundred and sixty-five patients with refractory unstable angina, defined as recurrent myocardial ischaemia despite medical treatment including heparin and nitrates were enrolled. After angiography, patients received an infusion of abciximab or placebo over 18-24 h preceding angioplasty, continuing until 1 h after the procedure. In 1197 patients undergoing angioplasty the angiographic committee centrally reviewed the baseline as well as the procedural angiograms. Coronary flow and lesion characteristics were assessed in the baseline angiogram as well as before intervention. Angiographic outcome, reason for failure as well as complications were assessed after angioplasty. RESULTS: At 30 days follow-up, patients receiving abciximab (n=595) compared with placebo (n=602) had a 30% reduction in the composite primary end-point death, myocardial infarction or urgent (re)intervention: 10.8% vs 15.4% (P=0.017). Baseline demographics were identical in the angiogram available group compared with the total study group. At 30 days, the non-angiogram available patients showed a higher incidence of events compared to those in whom the angiogram was reviewed: 19.4 vs 13.1% (P=ns). Lesion characteristics and coronary flow were not different at baseline between the placebo and abciximab groups. A primary end-point was reached in 9.6% of both placebo and abciximab patients with type A or B(1)lesions, in 17.0% vs 12.0% with type B(2)lesions, and in 19.1% vs 11.5% with type >B(2)or C lesions. Sixty-one percent of placebo and abciximab patients had TIMI 3 flow at baseline angiography. Pre-angioplasty TIMI 3 flow was observed in 69% and 72% respectively. The thrombus was resolved between the angiograms in 22% and 43% respectively, in the placebo and abciximab groups (P=0. 033). Angiographic success of the procedure was achieved in 88% and 94% in the placebo and abciximab patients, respectively (P<0.001). Stents were implanted in the ischaemia-related artery in 56 and 60 patients, respectively. However, failure of the stent procedure was more frequent in the placebo group than in the abciximab group, nine vs no patients (P=0.003). CONCLUSION: More frequent thrombus resolution was observed and a higher angiographic success rate was achieved in patients treated with abciximab before and during angioplasty compared with placebo. Patients with complex lesions as the underlying pathology reached fewer end-points if treated with abciximab before and during angioplasty. PMID- 10529326 TI - Relationship between psychological profile and cardiological variables in chronic heart failure. The role of patient subjectivity. AB - AIM: To analyse the relationships between the psychological profile, the satisfaction profile and cardiological variables in patients with chronic heart failure. MATERIAL AND METHODS: One hundred and fifty-two male patients with chronic heart failure in a stable clinical condition underwent cardiological evaluation and psychological assessment by means of two instruments: the Cognitive Behavioural Assessment 2.0 Battery and the Satisfaction Profile. RESULTS: Patients scored higher than healthy subjects in terms of psychophysiological disorders and depression. Patients in NYHA class III reported higher anxiety and depression scores and had more frequent problems in daily life than patients in NYHA classes I and II. Class III patients also reported lower satisfaction levels in many aspects of psychological and physical functioning. Pulmonary resistances >2.5 Wood units, pulmonary capillary wedge pressure >0. 18 mmHg and a diagnosis of ischaemic cardiomyopathy were associated with low satisfaction levels in the Satisfaction Profile 'physical functioning' factor. To be listed for heart transplantation and a history of more than three hospitalizations were related to low satisfaction levels in many items of the Satisfaction Profile. Finally, stepwise multiple regression showed that NYHA class, depression score and pulmonary capillary resistance accounted for 32% of the variance in the Satisfaction Profile physical functioning factor score. CONCLUSION: On the basis of chronic heart failure diagnosis only, a generic pattern of psychological distress can be predicted, common to many severe chronic diseases. Shifting from objective mental health measures towards the domain of subjective satisfaction, the only link which emerges is between objective cardiological data and satisfaction with physical functioning. Satisfaction in terms of other life aspects does not seem to be related to cardiological variables. These results support the importance of subjectivity in health related quality of life, as well as objective measures. PMID- 10529327 TI - Identification of a genetic risk factor for idiopathic dilated cardiomyopathy. Involvement of a polymorphism in the endothelin receptor type A gene. CARDIGENE group. AB - BACKGROUND: Idiopathic dilated cardiomyopathy is a frequent cause of heart failure, a major concern of public health. Although idiopathic dilated cardiomyopathy may be familial, most cases are sporadic and the disease is considered to be multifactorial, for which genetic factors may account for a significant part. METHODS AND RESULTS: We hypothesized that genetic abnormalities of the endothelin pathway may be involved in idiopathic dilated cardiomyopathy pathophysiology and therefore examined the possible association between idiopathic dilated cardiomyopathy and polymorphisms in genes encoding endothelin 1, endothelin type A and type B receptors, in a case-control study (433 patients and 400 age- and sex-matched control subjects). Analysis of the Exon 8 C/T polymorphism in the endothelin receptor type A gene indicated that individuals who are homozygote for the T allele were at significantly increased risk for the disease (odds ratio: 1.9; 95% confidence interval: 1.2 to 3. 01;P<0.006). Analysis of the other polymorphisms indicated that no significant difference was observed in genotype or allele frequencies between cases and controls. CONCLUSIONS: The variant in the Exon 8 of the endothelin receptor type A gene appears as a genetic risk factor for idiopathic forms of heart failure. These results provide a new approach to the pathophysiology of idiopathic dilated cardiomyopathy. PMID- 10529328 TI - Decrease in mortality in patients with a hospital diagnosis of atrial fibrillation in Denmark during the period 1980-1993. AB - BACKGROUND: Atrial fibrillation is associated with increased mortality. We hypothesized that the death rate in atrial fibrillation patients in Denmark has diminished during the period 1980-1993. METHODS: In a random sample of half of the Danish population, 30 330 patients were found to have a diagnosis of incident atrial fibrillation in the Danish National Hospital Discharge Register 1980-1993. Information on previous and concomitant cardiovascular and metabolic diseases during the period 1977-1993 was sought in the register. The temporal trend in total and cardiovascular mortality in the cohort of atrial fibrillation patients was analysed. RESULTS: A significant decrease in total and cardiovascular mortality was seen, 12-13% for total mortality and 17-18% for cardiovascular mortality. By adjusting for the decreasing cardiovascular mortality rate in the general population, a decrease in the relative risk of total mortality of 8-13% with time was seen for the atrial fibrillation cohort, compared with the population risk, while no reduction in the relative risk of cardiovascular death was seen. CONCLUSION: A significant decrease in mortality with calendar period occurred in the cohort of atrial fibrillation patients. PMID- 10529329 TI - Trial design for heart failure studies. PMID- 10529344 TI - Journal of Molecular Biology: a publishers perspective. PMID- 10529345 TI - The 100 most-cited articles from JMB. AB - Over the last 40 years, JMB has published many thousands of articles, all of which have been important in some way. Compiling a list of the 'most important' however, is an invidious task. Friendships can falter on such an undertaking, but the Institute of Scientific Information has provided us with an objective methodology for 'ranking' articles, according to the number of times any paper is cited in other publications. This evaluation can of course be criticised for its bias towards papers describing novel techniques or methods. Often, the true intellectual milestones may be found in the reference list of the most cited papers. With increasing age, each paper also has more time in which to have been cited, and so the group of highest scoring articles is also dominated by some of the oldest. On the other hand, with increasing time, papers have an increasing chance of being forgotten, and the citation rates of these are therefore also a measure of their persisting importance. On balance, it does represent a value in some way related to how often that paper has been used. With many caveats, we present the list of the 100 most cited papers in JMB over the past 40 years. Many of these papers have helped or influenced both a great many people, and a great many subsequent advances in molecular biology. PMID- 10529346 TI - From phage to chromosome biology: a personal account. AB - A personal account from phage morphogenesis to chromosome biology is described in this review. PMID- 10529347 TI - ATP-dependent chromatin remodelling: SWI/SNF and Co. are on the job. AB - SWI/SNF, RSC, NURF, CHRAC, ACF, RSF and NuRD are highly conserved multiprotein complexes that use the energy of ATP-hydrolysis to remodel chromatin. These complexes that have different subunit composition, all rely on helicase-like enzymes for ATPase activity and affect chromatin structure in similar ways. The specific function of the different complexes remains unclear, but many of them seem to be involved in transcriptional regulation. Although all cellular genes may not depend on chromatin remodelling for normal expression, recent data has shown that the complexes are required for both positive and negative control of a variety of cellular pathways. PMID- 10529348 TI - Zinc finger peptides for the regulation of gene expression. AB - Zinc fingers are small DNA-binding peptide motifs that were discovered in this laboratory. These motifs can be used as modular building blocks for the construction of larger protein domains that recognise and bind to specific DNA sequences. Phage display has been used to create a large library of different zinc fingers from which selections were made for binding to a given DNA sequence. From this database there have been elucidated elements of recognition rules that relate the amino acid sequence of a finger to its preferred DNA binding site. Control of gene expression using designed zinc finger peptides has been demonstrated by the specific inhibition of an oncogene mouse cell line and also by switching on genes in expression plasmids. These experiments demonstrate that zinc finger DNA-binding domains can be engineered de novo to recognise given DNA sequences. Five to six individual zinc fingers linked together would recognise a DNA sequence 15-18 bp in length, sufficiently long to constitute a rare address in the human genome. By adding functional groups to the engineered DNA-binding domains, e.g. silencing domains, novel transcription factors can be generated to up- or downregulate expression of a target gene. Among potential applications are the repression of oncogene expression and the disruption of the reproductive cycle of virus infection. PMID- 10529349 TI - Protein folds, functions and evolution. AB - The evolution of proteins and their functions is reviewed from a structural perspective in the light of the current database. Protein domain families segregate unequally between the three major classes, the 32 different architectures and almost 700 folds observed to date. We find that the number of new topologies is still increasing, although 25 new structures are now determined for each new topology. The corresponding analysis and classification of function is only just beginning, fuelled by the genome data. The structural data revealed unexpected conservations and divergence of function both within and between families. The next five years will see the compilation of a definitive dictionary of protein families and their related functions, based on structural data which reveals relationships hidden at the sequence level. Such information will provide the foundation to build a better understanding of the molecular basis of biological complexity and hopefully to facilitate rational molecular design. PMID- 10529350 TI - Roles for glycosylation of cell surface receptors involved in cellular immune recognition. AB - The majority of cell surface receptors involved in antigen recognition by T cells and in the orchestration of the subsequent cell signalling events are glycoproteins. The length of a typical N-linked sugar is comparable with that of an immunoglobulin domain (30 A). Thus, by virtue of their size alone, oligosaccharides may be expected to play a significant role in the functions and properties of the cell surface proteins to which they are attached. A databank of oligosaccharide structures has been constructed from NMR and crystallographic data to aid in the interpretation of crystal structures of glycoproteins. As unambiguous electron density can usually only be assigned to the glycan cores, the remainder of the sugar is then modelled into the crystal lattice by superimposing the appropriate oligosaccharide from the database. This approach provides insights into the roles that glycosylation might play in cell surface receptors, by providing models that delineate potential close packing interactions on the cell surface. It has been proposed that the specific recognition of antigen by T cells results in the formation of an immunological synapse between the T cell and the antigen-presenting cell. The cell adhesion glycoproteins, such as CD2 and CD48, help to form a cell junction, providing a molecular spacer between opposing cells. The oligosaccharides located on the membrane proximal domains of CD2 and CD48 provide a scaffold to orient the binding faces, which leads to increased affinity. In the next step, recruitment of the peptide major histocompatibility complex (pMHC) by the T-cell receptors (TCRs) requires mobility on the membrane surface. The TCR sugars are located such that they could prevent non-specific aggregation. Importantly, the sugars limit the possible geometry and spacing of TCR/MHC clusters which precede cell signalling. We postulate that, in the final stage, the sugars could play a general role in controlling the assembly and stabilisation of the complexes in the synapse and in protecting them from proteolysis during prolonged T-cell engagement. PMID- 10529351 TI - A Day in the Life of Dr K. or How I Learned to Stop Worrying and Love Lysozyme: a tragedy in six acts. AB - About the play: In modern drama, the agonizing nature of membrane protein work has not been adequately acknowledged. It is perhaps significant that the first attempt to bring this darker aspect of human existence into focus comes from a Scandinavian author, writing in the tradition of Ibsen and Strindberg but with a distinctly turn-of-the-millenium approach to the inner life of his characters: the despairing Dr K; the cynical Dr R with his post-modernistic life credo; the ambitious but unfeeling Dr C; the modern Ubermensch, Dr B. , with his almost Nietzschean view of human nature. This is a play that is brutally honest, yet full of empathy for the poor souls that get caught between the Scylla of unreachable scientific glory and the Charybdis of helpless mediocrity.James Glib Burdock, drama critic for The Stratford Observer. PMID- 10529352 TI - ABC-ATPases, adaptable energy generators fuelling transmembrane movement of a variety of molecules in organisms from bacteria to humans. AB - The approximately 27 kDa ABC-ATPase, an extraordinarily conserved, unique type of ATPase, acts as a machine to fuel the movement across membranes of almost any type of molecule, from large polypeptides to small ions, via many different membrane-spanning proteins. A particular ABC-ATPase must therefore be tailor-made to function in a complex with its cognate membrane protein, forming a transport pathway appropriate for a specific type of molecule, or in the case of some ABC transporters, several types of molecule. Molecules to be transported recognise their own transporter, bind and switch on the ATPase, which in turn activates or opens the transport pathway. ABC-dependent transport can be inwards across the membrane, or outwards to the cell exterior, and the ABC-ATPase can fuel transport through pathways which may involve a classical channel (CFTR), a "gateway" mechanism through a proteinacious chamber spanning the bilayer, or conceivably via a pathway at the protein-lipid interface of the outside of the membrane domain. This may be the case for drugs transported by Pgp, a multidrug resistance transporter. In this review, we try to identify the common fundamental principles which unite all ABC-transporters, including the basis of specificity for different transported compounds (allocrites), the interactions between the ATPase and membrane domains, activation of the ATPase and the coupling of consequent conformational changes, to the final movement of an allocrite through a given transport pathway. We discuss the so far limited structural information for the intact ABC-transporter complex and the exciting information from the first crystal structure of an ABC-ATPase. Finally, the action of specific transporters, CFTR (Cl- transport), Pgp, MRP and LmrA, all transporting many different drug molecules and HlyB transporting a large protein toxin are discussed. PMID- 10529353 TI - Geometry of phage head construction. AB - The process of phage capsid assembly is reviewed, with particular attention to the probable role of curvature in helping to determine head size and shape. Both measures of curvature (mean curvature and Gaussian curvature, explained in Appendix I), should act best when the assembling shell is spherical, which could account for procapsids having this shape. Procapsids are also relatively thick, which should help head size determination by the mean curvature. The accessory role of inner and outer scaffolds in size determination and head nucleation is also reviewed. Nucleation failure generates various malformations, including non closure, but the most common is the tube or polyhead, where the subunits' inherent curvature is expressed as a constant mean curvature. This induces lattice distortions that only partly understood. An extra tubular section in normal heads leads to the prolate shape, with a more complex and variable geometry. Newly assembled procapsids are both enlarged and toughened by the head transformation. In the procapsid the Gaussian curvature is uniformly distributed. But toughening tends to equalize bond lengths, so all the Gaussian curvature gets concentrated in the vertices, being zero elsewhere. This explains head angularization. Because of this change in Gaussian curvature, the regular subunit packing in the polyhedral head cannot be mapped onto the procapsid. This explains part of the hexon distortions found in this region. The implications of translocase-induced DNA twist, end rotation and the coiling of packaged DNA, are discussed. The symmetry mismatches between the head, connector and tail are discussed in relation to the possible alpha-helical structures of their DNA channels. PMID- 10529354 TI - Cloning and partial characterization of the proteasome S4 ATPase from Plasmodium falciparum. AB - Certad, G., Abrahem, A., and Georges, E. 1999. Cloning and Partial characterization of the proteasome S4 ATPase from Plasmodium falciparum. Experimental Parasitology 93, 123-131. The ATP-ubiquitin-proteasome pathway mediates the nonlysosomal degradation of cytosolic proteins in eukaryotic cells. The activities of this pathway have been shown to regulate cell growth and differentiation through modulation of regulatory proteins. The proteasome is a large complex consisting of two multisubunit structures, the 20S and 19S(PA700) or P28 complexes, that combine to form the 26S particles. In this study, we describe the cloning of a cDNA encoding the proteasome subunit 4 ATPase homologue from Plasmodium falciparum (PFS4). Analysis of the PFS4 cDNA sequence shows an open reading frame encoding a deduced protein of 455 amino acids. Moreover, comparison of PFS4 cDNA sequence to that of genomic fragments encoding PFS4 showed identical sequences with no detectable introns. Database searches revealed a high sequence identity to those of rice, yeast, mouse, Drosophila, and human S4 ATPases. However, PFS4 contains two unique inserts of nine and seven amino acid residues in the N-terminal domain. Interestingly, only the rice S4 contains the latter (seven amino acids) insert with four identical amino acids. In vitro expression of the full-length cDNA encoding the PFS4, using a transcription translation-coupled reticulocyte lysate, shows a 50-kDa [(35)S]methionine-labeled protein which was immunoprecipitated with PFS4 anti-peptide antiserum. Southern blot analysis of genomic DNA digests shows a single gene copy of PFS4 in P. falciparum. Of interest was the effect of the proteasome-specific natural product, lactacystin, on the growth of the parasite, with IC(50) values of 0.6 0.92 microM. The latter IC(50) values of lactacystin for different clones of P. falciparum are comparable to those obtained for mammalian cell lines (0.65 microM), suggesting the presence of a conserved proteasome complex. Moreover, lactacystin was equally toxic to drug-sensitive and resistant parasites. PMID- 10529355 TI - Schistosoma mansoni: continuous variation in susceptibility of the vector snail of schistosomiasis, Biomphalaria tenagophila I. Self-fertilization-lineage. AB - Mascara, D., Kawano, T., Magnanelli, A. C., Silva, R. P. S., Sant' Anna, O. A., and Morgante, J. S. 1999. Schistosoma mansoni: continuous variation in susceptibility of the vector snail of schistosomiasis, Biomphalaria tenagophila I. Self-Fertilization-Lineage. Experimental Parasitology 93, 133-141. Artificial selection of Biomphalaria tenagophila snails for susceptibility to infection by Schistosoma mansoni (Brazilian SJ strain) was carried out from natural populations. After five self-fertilization generations, two lineages were isolated and were designated as SUSC (highly susceptible 93-100%) and RES (nonsusceptible 5-0%). Length of the prepatent period, cercarial production, and mortality of the hosts in postexposure were determined in all generations (F(1) F(8)) and were analyzed as quantitative traits related to host susceptibility. Distribution patterns of frequencies were observed within snail families (samples derived from one F(0) snail), these traits showing a significant influence by selection applied to susceptibility. The multiple quantitative classes were described in terms of continuous variation. During the selection of SUSC lineage, classes with higher values of prepatent length and lower cercarial production were eliminated, and the heritability calculated for these two traits was 0.811 and 0.709, respectively. Experimental results were correlated with an increase in the level of susceptibility in the generations selected and are discussed in relation to inheritance patterns as well as the quantitative variation of susceptibility. PMID- 10529356 TI - Trypanosoma cruzi: suppression of tuzin gene expression by its 5'-UTR and spliced leader addition site. AB - Teixeira, S. M. R., Kirchhoff, L. V., and Donelson, J. E. 1999. Trypanosoma cruzi: Suppression of tuzin gene expression by its 5'-UTR and spliced leader addition site. Experimental Parasitology 93, 143-151. The genome of the protozoan parasite Trypanosoma cruzi contains a tandemly repeated array of two alternating genes, one encoding amastin and the other encoding tuzin. Amastin is an abundant amastigote surface protein, whereas tuzin is thought to be a rare protein whose location and function are unknown. The 137-nucleotide 5' untranslated region (5' UTR) of the tuzin mRNA has a 22-codon open translation reading frame containing 3 methionine codons followed by a stop codon that overlaps the methionine start codon of the tuzin coding region. A fragment containing the tuzin 5'-UTR and upstream intergenic region was placed in front of a luciferase reporter gene in a plasmid for transient transfection assays of luciferase activity. By mutating the three upstream ATGs in the tuzin 5'-UTR and replacing the tuzin spliced leader (SL) acceptor site with that of the amastin gene, we found that the 22-codon reading frame and the tuzin SL acceptor site combine to substantially reduce expression of the luciferase gene. These results indicate that expression of the multicopy tuzin gene is posttranscriptionally suppressed by both inefficient RNA processing and poor translation initiation, resulting in a low level of tuzin. PMID- 10529357 TI - Trichinella spp.: differential expression of two genes in the muscle larva of encapsulating and nonencapsulating species. AB - Kuratli, S., Lindh, J. G., Gottstein, B., Smith, D. F., and Connolly, B. 1999. Trichinella spp.: Differential expression of two genes in the muscle larva of encapsulating and nonencapsulating species. Experimental Parasitology 93, 153 159. The expression of the two genes tsmyd-1 and tsJ5 was studied in the muscle stage larva of three different species of Trichinella. T. spiralis and T. britovi are both encapsulating species, while T. pseudospiralis is a nonencapsulating species. Expression of tsJ5 is developmentally regulated in T. spiralis and has been shown in this study to be down-regulated in the T. pseudospiralis muscle larva compared with the other two species. Immunoblot analysis has also revealed that the relative abundance of the protein product of this gene, TSJ5, is lower in T. pseudospiralis muscle larvae. It has previously been shown that expression of tsmyd-1 is not developmentally regulated in T. spiralis (Connolly et al. 1996). In contrast, expression of this gene is slightly increased in the muscle larvae of T. pseudospiralis. Southern analysis of genomic DNA from the three Trichinella species shows that both genes are highly conserved. PMID- 10529358 TI - Leishmania: naive human T cells sensitized with promastigote antigen and IL-12 develop into potent Th1 and CD8(+) cytotoxic effectors. AB - Russo, D. M., Chakrabarti, P., and Higgins, A. Y. 1999. Leishmania: Naive human T cells sensitized with promastigote antigen and IL-12 develop into potent Th1 and CD8(+) cytotoxic effectors. Experimental Parasitology 93, 161-170. The differentiation of naive human T cells into Leishmania-specific Th1 or cytotoxic effector cells was examined by sensitizing T cells in vitro with dead Leishmania antigen in the presence or absence of IFN-gamma or IL-12. These Leishmania specific T cell lines proliferated and produced cytokines in response to challenge with autologous Leishmania-infected macrophages. Sensitization in the presence of IL-12 or IFN-gamma induced Leishmania-specific human Th1 responses, with IL-12 inducing more potent Th1 responses. However, IL-12-induced Th1 responses were IFN-gamma dependent. T cell lines exhibited Th2 or Th0 phenotypes when primed in the absence of cytokines. Only T cell lines primed in the presence of IL-12 contained high percentages of CD8(+) cells. These cells lysed autologous Leishmania-infected but not uninfected macrophages in an MHC-dependent manner. Thus, this in vitro sensitization system can be used to delineate the conditions for optimally priming human Leishmania-specific effector cells. PMID- 10529359 TI - Echinococcus granulosus: in vitro effects of ivermectin and praziquantel on hsp60 and hsp70 levels. AB - Martinez, J., Perez-Serrano, J., Bernadina, W. E., Rodriguez-Caabeiro, F. 1999 Echinococcus granulosus: In vitro effects of ivermectin and praziquantel on hsp60 and hsp70 levels. Experimental Parasitology93, 171-180. Organisms or cells exposed to injurious stresses such as heat shock or chemicals respond by increased (or altered) expression of heat-shock proteins (HSPs). Conversely, an earlier exposure to stress can prepare cells to cope with a subsequent more severe stress. In the present study, protoscolices of Echinococcus granulosus were subjected to several anthelmintic treatments, involving storage of the protoscolices for 18, 30, and 50 h with 0.1 mg/ml of ivermectin (IV), praziquantel (PZ), and a combination of each with albendazole (ALB). The organisms were analyzed for the effects of drug treatment on cell integrity and on levels of hsp60 and hsp70 production. Drug efficacy was evaluated by microscopy and by protein content measurement. Hsp60 and hsp70 were detected by Western blotting and incubation with anti-hsp60 and anti-hsp70 antibody, respectively, and quantitation of these proteins was obtained using image analysis. Incubation with IV alone produced the most damage to the protoscolices as indicated by viability loss, decreased protein content, and altered hsp60 and hsp70 levels; incubation with IV + ALB produced less damage as manifested by fewer changes in the aforementioned damage parameters but PZ and PZ + ALB, in this context, were poor anthelmintics. Exposure of protoscolices to thermal stress prior to anthelmintic treatment, in most cases, increased drug efficacy. It is concluded that in the E. granulosus model system drug efficacy is associated with decreased levels of hsp70 expression and increased levels of hsp60 expression. PMID- 10529360 TI - PPARalpha activation by Wy 14643 induces transactivation of the rat UCP-1 promoter without increasing UCP-1 mRNA levels and attenuates PPARgamma-mediated increases in UCP-1 mRNA levels induced by rosiglitazone in fetal rat brown adipocytes. AB - Rodent brown adipocytes express peroxisome proliferator activated receptor-alpha (PPARalpha) and PPARgamma and while the rodent uncoupling protein-1 (UCP-1) gene contains a putative peroxisome proliferator response element (PPRE), only PPARgamma activation by thiazolidinediones increase UCP-1 mRNA levels. We have investigated this phenomenon in foetal rat brown adipocytes (FBA) and show that although transactivation occurs in FBA containing a plasmid encoding 4.5kb of the 5'-flanking region of the rat UCP-1 promoter ((-4551)-UCP-1-CAT) treated with either the selective PPARgamma agonist rosiglitazone (1.0 microM) or the selective PPARalpha agonist Wy 14643 (10.0 microM), only rosiglitazone induced transcription of UCP-1 mRNA. Furthermore, Wy 14643 (10 and 100.0 microM) abolished rosiglitazone-induced UCP-1 mRNA induction in spite of a transactivation event occurring with the combination treatment. Thus in FBA PPARalpha-activation with Wy 14643 elicits a "blind" transactivation of the UCP-1 promoter which can prevent PPARgamma-mediated UCP-1 mRNA transcription either by competition for the PPRE or by an unidentified post-transcriptional event. PMID- 10529361 TI - Identification of two serine residues involved in catalysis by fatty acid amide hydrolase. AB - Fatty acid amide hydrolase is an integral membrane protein that hydrolyzes a novel and growing class of neuromodulatory fatty acid molecules, including anandamide, 2-arachidonyl glycerol, and oleamide. This activity is inhibited by serine and cysteine reactive agents, suggesting that the active site contains a serine or cysteine residue. Therefore serine and cysteine residues were mutated to alanine and the effects on activity were determined. Mutants were prepared using site-directed mutagenesis methods and expressed in COS-7 cells. Serine mutations S217A and S241A completely abolished enzymatic activity. Mutants S152A and C249A had no effect on activity, while S218A showed a slight decrease in activity. To confirm these results biochemically, the mutant enzymes were reacted with the irreversible inhibitor [(14)C]-diisopropyl fluorophosphate. All of the mutants except S217A and S241A were labeled. We therefore confirm that fatty acid amide hydrolase is a serine hydrolase and propose that both Ser-217 and Ser-241 are essential for enzyme activity. PMID- 10529362 TI - Immunosuppressive effect of leukotriene B(4) receptor antagonist in vitro. AB - Leukotriene B(4) (LTB(4)), which is an arachidonic acid metabolite produced by the 5-lipoxygenase pathway and a well-characterized chemical mediator of inflammation, has been proposed to be an immune response modulator. Here we showed the constitutive expression of the LTB(4) receptor (LTB(4)R) in resting and activated T cells. We found that the LTB(4)R antagonist inhibited T cell proliferation induced by Con A, immobilized anti-CD3 mAb, or IL-2. This inhibitory effect was abolished by addition of LTB(4)R agonist. The LTB(4)R antagonist inhibited IL-2, IFN-gamma, and IL-4 production by anti-CD3-stimulated T cells and also inhibited IL-12-induced IFN-gamma production. Moreover, the LTB(4)R antagonist exerted an additive inhibitory effect to FK506 on T cell proliferation. These results suggest that LTB(4) is intrinsically involved in T cell activation to upregulate cytokine production and proliferation, and thus the LTB(4)R antagonist might be useful as an immunosuppressive agent. PMID- 10529363 TI - Intramolecular quadruplex formation of the G-rich strand of the mouse hypervariable minisatellite Pc-1. AB - The minisatellite Pc-1, isolated from the mouse genome consisting of a tandem repeat of d(GGCAG), is hypervariable with a mutation rate of 0.15/generation. Here we describe a structural characterization of the G-rich strand of Pc-1 by biochemical and physicochemical methods. It was found to be comparatively resistant to both single-stranded DNA-binding protein binding and digestion by single-stranded DNA-specific nuclease and to cause arrest of DNA synthesis. The guanine imino proton NMR signals observed on the Pc-1 G-rich strand and their slow (1)H/(2)H exchange profiles pointed to a quadruplex structure with guanine quartets. The melting temperature of the quadruplex determined by CD was not dependent on DNA concentration. These results indicate that the G-rich strand of Pc-1 forms an intramolecular folded-back quadruplex structure under physiological conditions. Possible mechanisms of the Pc-1 mutations implicated with the formation of the quadruplex structure are discussed. PMID- 10529364 TI - A novel DIDS-sensitive, anion-dependent Mg(2+) efflux pathway in rat ventricular myocytes. AB - The aim of this study was to identify the existence of anion-dependent Mg transport systems in cardiac muscle. DIDS-sensitive and anion-dependent (either Cl(-)(o) or NO(-)(3o)) increases in [Mg(2+)](i) occurred during Mg(2+) loading conditions. Much larger elevations of [Mg(2+)](i) occurred under Cl(-)(o)-free conditions with 0.1 mmol l(-1) DIDS, compared to Cl(-)(o) replacement alone. All these effects were abolished in Mg(2+)(o)-free medium. These data suggest a novel Mg(2+)-anion symport for Mg(2+) efflux against the electrochemical gradient that is fueled mostly by the efflux of an endogenous anion (HCO(-)(3)?), but with a small contribution from intracellular Cl(-) probably supplied via the Cl(-)-HCO( )(3) exchanger. PMID- 10529365 TI - Characterization of multiple nuclear localization signals in herpes simplex virus type 1 thymidine kinase. AB - We have reported previously that the herpes simplex virus type 1 (HSV-1) thymidine kinase (TK) fused with green fluorescent protein (GFP) is localized in the nucleus of HSV-1 TK-GFP gene-transfected cells (Degreve et al. (1998) J. Virol. 72, 9535-9543). Deletion of the N-terminal 34 amino acids or selective mutation of the nonapeptide (25)RRTALRPRR(33), located in the N-terminal region of HSV-1 TK, resulted in the loss of the specific nuclear localization of HSV-1 TK. Utilizing information on the crystallographic structure of HSV-1 TK, we have now identified three additional putative nuclear localization signals and evaluated their potential role in the nuclear trafficking of HSV-1 TK by site directed mutagenesis. We found that the sites containing the amino acids R236 R237 and K317-R318 are absolutely required for specific nuclear targeting of HSV 1 TK. The K317-R318 region, located at the interface between the two monomers in the dimeric HSV-1 TK structure, could act as a nuclear localization signal for monomeric HSV-1 TK. Alternatively, crystallographic data indicate that R318 might be essential for the formation of the TK dimer, and therefore it is required if HSV-1 TK is transported as a dimer. PMID- 10529366 TI - Loss of molecular interaction between cytochrome c and cardiolipin due to lipid peroxidation. AB - To explore the molecular mechanism underlying the translocation of cytochrome c from the mitochondrial inner membrane to the cytosol during apoptosis, we analyzed the molecular interaction between cytochrome c and cardiolipin (CL) by (1)H NMR spectroscopy. Bovine heart CL induced a drastic broadening of the linewidth of the downfield signals at 31.4 and 34.2 ppm assigned to the heme methyl group-3 and -8, respectively, of horse heart cytochrome c. In contrast, CL mono- and dihydroperoxides were less active in broadening the signals than CL, and CL trihydroperoxides induced almost no broadening of their linewidth. This finding suggests that the peroxidation of CL induces a release of cytochrome c from mitochondria into the cytosol, which release induces apoptosis in the cells. PMID- 10529367 TI - Cell wall-affecting antibiotics induce expression of a novel gene, drp35, in Staphylococcus aureus. AB - A novel gene, drp35, of Staphylococcus aureus, which was inducible especially with cell wall-affecting antibiotics, has been cloned. Analysis of differential hybridization with mRNAs enhanced in the presence of beta-lactams resulted in two positive clones that harbored a new gene encoding a 35,845-Da protein (Drp35) and the penicillin-binding protein 2 (PBP2). Immunoblot analysis revealed that the Drp35 protein band was evidently enhanced after 30 min in the presence of beta lactams. The Drp35 expression was also enhanced with not only beta-lactams, but also vancomycin, bacitracin, and fosfomycin. Homology search revealed that Drp35 was a new protein. Our results revealed that it was specific in S. aureus and respondent to these agents in both methicillin-resistant and -sensitive strains of S. aureus. PMID- 10529368 TI - Proteasome inhibitors lactacystin and MG132 inhibit the dephosphorylation of HSF1 after heat shock and suppress thermal induction of heat shock gene expression. AB - Recently, we have shown that two proteasome inhibitors, MG132 and lactacystin, induce hyperphosphorylation and trimerization of HSF1, and transactivate heat shock genes at 37 degrees C. Here, we examined the effects of these proteasome inhibitors and, in addition, a phosphatase inhibitor calyculin A (CCA) on the activation of HSF1 upon heat shock and during post-heat-shock recovery, with emphasis on HSF1 hyperphosphorylation and the ability of HSF1 to transactivate heat shock genes. When lactacystin, MG132, or CCA was present after heat shock, HSF1 remained hyperphosphorylated during post-heat-shock recovery at 37 degrees C. Failure of HSF1 to recover to its preheated dephosphorylated state correlated well with the suppression of the heat-induced hsp70 expression. In vitro, HSF1 from heat-shocked cells, when dephosphorylated, showed an increase in HSE-binding affinity. Taken together, these data suggest that phosphorylation of HSF1 plays an important role in the negative regulation of heat-shock response. Specifically, during post-heat-shock recovery phase, prolonged hyperphosphorylation of HSF1 suppresses heat-induced expression of heat shock genes. PMID- 10529369 TI - p53 down-regulates ER-responsive genes by interfering with the binding of ER to ERE. AB - Overexpression of the tumor suppressor p53 in HeLa cells leads to loss of the estradiol- and genistein-induced human estrogen receptor (ERalpha) transactivity. The coactivator p300, which binds to both ERalpha and p53, does not prevent this loss of hERalpha function. In this report we demonstrate that p53 physically binds to multiple domains of the hERalpha. This binding did not interfere with either the ERalpha dimerization or the interaction between hERalpha and its coactivator SRC-1. However, p53 did interfere with the hERalpha-ERE binding. These results may explain how p53 down-regulates the expression of some estrogen responsive genes such as c-fos, c-jun, TPA, and bcl-2. This study supports the cross-talk between the p53 and the ERalpha signaling pathways. PMID- 10529370 TI - A novel compound heterozygote (FAP ATTR Arg104His/ATTR Val30Met) with high serum transthyretin (TTR) and retinol binding protein (RBP) levels. AB - A 64-year-old Japanese male suffering from very slowly progressive amyloidosis was studied by immunohistopathologic, mass spectrometric, and molecular genetic methods. After confirming the immunoreactivity of transthyretin (TTR) in the amyloid deposits using an anti-TTR polyclonal antibody, matrix-assisted laser desorption ionization/time-of-flight-mass spectrometry (MALDI/TOF-MS) was employed to look for the presence of variant TTR(s) in the serum. Two variant forms of TTR, one with a molecular weight 32 Da greater and another with a molecular weight 19 Da less than that of normal TTR encoded by the two respective alleles, were detected in this patient. Direct sequence analysis confirmed the presence of a double substitution: one at codon 30 from GTG (Val) to ATG (Met) and the other at codon 104 from CGC (Arg) to CAC (His) in the two alleles. MALDI/TOF-MS of the parents of the proband revealed that his father was a heterozygote of ATTR Arg104His and his mother was a heterozygote of ATTR Val30Met. The total TTR and retinol binding protein (RBP) concentrations in the serum samples of the proband were very high compared with those of FAP ATTR Val30Met patients and control subjects. We report here a new compound heterozygote in the TTR gene with familial amyloidotic polyneuropathy (FAP). PMID- 10529371 TI - Contribution of mitogen-activated protein kinase to stimulation of phospholipase D by the chemotactic peptide fMet-Leu-Phe in human neutrophils. AB - Phospholipase D (PLD) plays an important role in signaling through phosphatidylcholine (PC) and in the production of superoxide (respiratory burst) by polymorphonuclear leukocytes (PMN) stimulated by the chemoattractant fMet-Leu Phe (fMLP). However, the regulation of PLD activity by protein kinases is not fully understood. In the present study, we have used a mitogen-activated protein (MAP) kinase inhibitor (PD 98059) to investigate a possible connection between extracellular signal-regulated kinase (ERK) and PLD activity and respiratory burst. Using a range of concentrations (3-20 microM) which inhibit ERK activity, PD 98059 inhibited PLD activity induced by fMLP in cytochalasin B-primed PMN, as assessed by production-tritiated phosphatidylethanol (PEt), phosphatidic acid (PA), and hydrolysis of PC. However, the inhibition was partial (approximately 50%), while inhibition of PC hydrolysis was almost complete, suggesting a concomitant inhibition of PLA2 activity. In addition, PD 98059 reduced fMLP induced respiratory burst by 50%, an effect which was correlated with PLD inhibition of PLD (r = 0.981, P < 0.01), and neither did PD 98059 inhibit the PLD activity and respiratory burst induced by PKC upon its direct activation by phorbol myristate acetate. These data provide the first evidence for implication of the ERK cascade in the stimulation of PLD through Gi signaling. They further indicate that PLD stimulation by fMLP receptors occurs through two pathways, dependent and independent on MAP kinase, the former pathway being linked to superoxide production. PMID- 10529372 TI - 2,3,7,8-Tetrachlorodibenzo-p-dioxin-mediated oxidative stress in CYP1A2 knockout (CYP1A2-/-) mice. AB - The objective of the study was to compare alterations in indicators of oxidative stress following 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure in cytochrome P4501A2 (CYP1A2) knockout mice and their parental lineage strains (C57BL/6N and 129/Sv). This study will aid in determining the role, if any, of CYP1A2 in TCDD-mediated oxidative stress. Formation of thiobarbituric acid reactive substances (TBARS) as a measurement of lipid peroxidation, production of reactive oxygen species (ROS) via the in vitro reduction of cytochrome c in tissue homogenate, and changes in the biochemical antioxidant glutathione were monitored to determine oxidative stress 7 days following a single oral dose of 25 microg TCDD/kg. TBARS, reduction of cytochrome c, and changes in glutathione demonstrated a similar response in CYP1A2 knockout and parental strains. These data suggest that CYP1A2 does not play a critical role in the acute oxidative stress response following TCDD exposure. PMID- 10529373 TI - Cytochrome c-553 from the alkalophilic bacterium Bacillus pasteurii has the primary structure characteristics of a lipoprotein. AB - The complete sequence of Bacillus pasteurii cytochrome c-553 was determined by standard methods of Edman degradation of overlapping peptides combined with mass spectrometry. The protein contains 92 residues and a single heme-binding site. It is most similar to Bacillus licheniformis, Bacillus PS3, and Bacillus subtilis cytochromes c-551, which are lipoproteins that are partially solubilized through proteolytic cleavage of the N-terminal diacyl-glyceryl-cysteine membrane anchor. The high yield of the B. pasteurii cytochrome c-553, together with evidence that shorter forms of the cytochrome occur in the mixture of otherwise pure protein, suggests that the membrane anchor is very susceptible to proteolysis and that the soluble form of the cytochrome is therefore released from the membrane upon cell breakage. A sequence-based calculation of the protein secondary structure suggests the presence of a typical cytochrome helical fold with a random-coil N terminus tail. PMID- 10529374 TI - Single-strand breaks in agarose-embedded chromatin of nonapoptotic cells. AB - Loop-size chromatin fragmentation frequently observed upon apoptotic cell death is thought to be initiated by ss nicks. Here we show that the agarose-embedded, deproteinized chromatin of normal, non-apoptotic murine and human cells, as well as yeast protoplasts, falls apart to 50-300 kb ss fragments upon heat denaturation, as revealed by urea-TAE field-inversion agarose gel electrophoresis resolving ss and ds fragments alike. These data were in line with S1digestion experiments. The nicks (gaps) observed are best explained either by enzymatic cleavages occurring upon cell lysis instantaneously or by preexisting discontinuities becoming manifest upon heat denaturation. These discontinuities go unnoticed in the usual nondenaturaing circumstances but seem to be inevitably present in any DNA preparation. The loop-size ds DNA fragmentation in apoptosis may be based on these pre-existing or "ready-to-go" (upon cell lysis) ss discontinuities of the normal cellular chromatin. PMID- 10529375 TI - An upstream repressor element that contributes to hepatocyte-specific expression of the rat serum amyloid A1 gene. AB - Serum amyloid A (SAA) is a major acute-phase protein whose expression can be dramatically induced in response to tissue damage, infection, and inflammation. Its expression is highly tissue-specific, restricted almost exclusively to liver hepatocytes. We have shown that a 320-bp fragment of the rat SAA1 promoter could confer liver-cell-specific expression on a reporter gene when transfected into cultured cells. Here we report the identification of a 29-bp regulatory element that possesses inhibitory activities on SAA1 promoter in HeLa cells but has no such effects in liver cells. Moreover, this regulatory element has properties of a transcriptional repressor; in that, it can function with a heterologous promoter and is independent of orientation and distance from the transcription initiation site. Protein binding studies showed that this regulatory element can form specific protein-DNA complexes with nuclear proteins from several nonliver cell lines (HeLa, 10T(1/2), and C2) and placenta. However, the same DNA-protein complex was not detected in extracts from liver or liver-derived cell lines (HepG2 and Hep3B). Taken together, our results demonstrate the presence of a DNA binding protein, termed tissue-specific repressor, found only in nonhepatocytes which may function to repress SAA1 gene expression by interacting with a repressor element. Thus, liver-specific expression of the SAA1 gene is apparently regulated by both positive and negative regulatory elements and their interacting transcription factors to ensure that it is expressed only in suitable cell types. PMID- 10529376 TI - Characterization of the MEN1 ortholog in zebrafish. AB - Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant syndrome predisposing to multiple tumors. The responsible gene, MEN1, has been identified and inactivating mutations reported. It encodes a protein named menin, which lacks homology to any known proteins. Comparative genomics is used to ascertain important functional domains via the identification of evolutionary conserved regions. Here we report the sequencing and characterization of the MEN1 gene in zebrafish (Danio rerio) at the cDNA level. Zebrafish menin is a 617 amino acid protein and, when compared with human and rodent proteins, shows 75% and 76% similarity, respectively. The most conserved region is amino acid residues 41-322 which shows a human/zebrafish similarity of 83%. Amino acids affected by inactivating missense mutations in MEN1 patients in this region are completely conserved between human and zebrafish. Such high correlation between conservation throughout evolution and mutation position strongly emphasizes the importance of this region. PMID- 10529377 TI - Rapamycin inhibits activation of ryanodine receptors from skeletal muscle by the fatty acyl CoA-acyl CoA binding protein complex. AB - We previously showed (Fulceri et al., Biochem. J. 325, 423, 1997) that the fatty acyl CoA ester palmitoyl CoA (PCoA) complexed with a molar excess of its cytosolic binding protein (ACBP) causes a discrete Ca(2+) efflux or allows Ca(2+) release by suboptimal caffeine concentrations, in the Ca(2+)-preloaded terminal cisternae fraction (TC) from rabbit skeletal muscle, by activating ryanodine receptor Ca(2+) release channels (RyRC). We show here that both effects were abolished by pretreating TC with the FKBP12 ligand rapamycin (20 microM). Moreover, rapamycin reversed the Ca(2+) release induced by combined treatment with 3 mM caffeine and the PCoA-ACBP complex. Rapamycin also reduced the Ca(2+) releasing activity by PCoA alone. Under the above experimental conditions, rapamycin removed FKBP12 from the TC membranes, as revealed by Western blot analysis. We conclude that FKBP12 associated with RyRC in the TC membrane participates in the activation of the Ca(2+) channel by fatty acyl CoA esters. PMID- 10529378 TI - Cloning of a novel cytochrome P450 (CYP4C15) differentially expressed in the steroidogenic glands of an arthropod. AB - Biosynthesis of ecdysteroids, arthropod steroid molting hormones, proceeds from dietary cholesterol through a complex and still incompletely elucidated pathway. Most of the known steps are catalyzed by cytochrome P450 enzymes (CYPs) but none of their genes has yet been identified. We have established a cDNA library of crayfish steroidogenic glands (Y organs). A full length CYP-cDNA was characterized containing a 1539 bp open reading frame encoding a predicted protein of 513 amino acid residues. This novel CYP was assigned to the CYP4 family and designated CYP4C15. Northern blots demonstrated predominant expression of this gene in the active molting glands, suggesting a role in ecdysteroid biosynthesis rather than detoxification. PMID- 10529379 TI - Characterization and cDNA cloning of androgenic gland hormone of the terrestrial isopod Armadillidium vulgare. AB - The sex differentiation in crustaceans is known to be controlled by a peptide hormone called androgenic gland hormone (AGH). AGH was extracted and purified from the androgenic glands (AGs) of the male isopod Armadillidium vulgare by high performance liquid chromatography. AGH consisted of two peptide chains and their N-terminal amino acid sequences were determined. A cDNA encoding AGH was cloned by PCR and sequenced. The cDNA had an open reading frame of 432 bp, which encoded a preproAGH consisting of a signal peptide (21 residues), B chain (44 residues), C peptide (46 residues), and A chain (29 residues). Through processing, the A and B chains might form a heterodimer interlinked by disulfide bonds. The A chain possessed a putative N-linked glycosylation site. A Northern blot analysis using the cDNA as a probe detected a hybridization signal with 0.8 kb in the RNA preparation only from the AGs. PMID- 10529380 TI - Human serum immunoglobulin G3 subclass bound preferentially to asialo-, agalactoimmunoglobulin A1/Sepharose. AB - As we reported before, exoglycosidase treatment of human serum IgA1 changed it to a sticky molecule. In order to examine the presence of the specific binding protein to the sticky IgA1 in human serum, IgA1, asialo-IgA1 (IgA1-S) and asialo , agalacto-IgA1 (IgA1-SG)/Sepharose column chromatography of normal human serum was carried out. Purified hinge glycopeptide (HGP33) prepared from IgA1 was used for the preparation of HGP/Sepharose. A portion of the serum protein was bound to those columns and eluted with the buffer containing 1.0 M NaCl. About four times the amount of protein was eluted from the IgA1-SG/Sepharose column than that from IgA1/Sepharose. Most of the eluted protein was IgG, and the IgG1 and IgG3 subclasses but neither IgG2 nor IgG4 was dominant. Under the lower salt concentration, a portion of IgG was also bound to the HGP-SG/Sepharose column. The obtained results coincide well with the previous report of the co-present of IgG1 and IgG3 with deposited IgA1 in IgA nephropathy patients (Aucouturier, P., et al. (1989) Clin. Immunol. Immunopathol. 51, 338-347). Thus, the results solved the question of why the IgG3 was co-present with deposited IgA1 in IgA nephropathy patients. PMID- 10529381 TI - Identification of autoantibodies associated with rippling muscles and myasthenia gravis that recognize skeletal muscle proteins: possible relationship of antigens and stretch-activated ion channels. AB - The role of mechanosensitive calcium channels in skeletal muscle physiology is not understood. This study takes advantage of an autoimmune neuromuscular disorder (myasthenia gravis associated with rippling muscles) to identify components in the skeletal muscle myocyte that may play a role in mechanosensitive calcium channel activity. Rippling muscles are characterized by stretch or percussion activated wave-like muscle contractions that do not require motor unit action potentials for propagation. Autoantibodies from the sera of patients with autoimmune rippling muscles (associated with myasthenia gravis) are directed against high molecular weight muscle proteins. Some of these proteins are uniquely recognized by antisera from patients with autoimmune rippling muscles. This suggests these autoantigens are distinct from those normally associated with myasthenia gravis, and may play a role in the mechanosensitive activation of muscle contraction. PMID- 10529382 TI - Hypoxic stress induces cardiotrophin-1 expression in cardiac myocytes. AB - Cardiotrophin-1 (CT-1), a novel cytokine that belongs to the interleukin-6 cytokine family, activates gp130 dependent signaling pathway to transduce hypertrophic and cytoprotective signals in cardiac myocytes. To investigate the pathophysiological significance of CT-1 in myocardial disease, the expression of CT-1 was examined after hypoxic stimulation in cardiac myocytes. Highly expressed CT-1 mRNA was observed in embryonic and adult hearts by RNase protection assay. Cardiac myocytes subjected to hypoxic stimulation augmented CT-1 mRNA expression. Although CT-1 mRNA was expressed to a higher extent in non-myocardial cells, the expression was not affected with the stimulation. Conditioned medium from cultured cardiac myocytes presented the ability to tyrosine phosphorylate STAT3 through gp130 and that was further augmented with hypoxic conditioned medium. These results demonstrated for the first time that CT-1 expression is augmented after hypoxic stimulation and hypoxic conditioned medium presented enhanced ability to activate STAT3 in cardiac myocytes. CT-1 might play an important role in the pathogenesis of ischemic heart disease. PMID- 10529383 TI - Advanced glycation end product-induced peroxisome proliferator-activated receptor gamma gene expression in the cultured mesangial cells. AB - We identified the AGEs-induced expression of peroxisome proliferator-activated gamma (PPAR gamma) in the cultured mesangial cells using reverse transcription polymerase chain reaction, electrophoretic mobility shift assay (EMSA), and Western immunoblotting. Administration of AGEs-BSA into the cultured mesangial cells resulted in an increase in the levels of mRNA and proteins for PPAR gamma in a dose-dependent manner. Specific bands which indicate the protein binding to PPAR gamma responsive element (PPRE) in the nuclear extracts were also detected in AGEs-BSA-treated mesangial cells, but not found in BSA-treated cells by EMSA. Antioxidants, NAC, PDTC, and aminoguanidine, attenuated the gene expression and activity of PPAR gamma induced by AGEs. These results indicate that PPAR gamma was induced and activated by the oxidative signal(s) evoked by AGEs-ligand receptor interactions. AGEs-induced gene expression of PPAR gamma and the signal intensity of PPAR gamma and PPRE complex were attenuated furthermore by protein kinase C inhibitors, calphostin C and staurospolin, but not abolished completely, indicating that both signal transduction pathways through the induction of PKC activation and independent of PKC activation were involved in the AGEs-mediated expression and activation process of PPAR gamma. AGEs also increased the gene expression of smooth muscle alpha-actin, which is a marker for phenotypic change in mesangial cells. It is suggested therefore that AGEs-induced transcription factor as the oxidative stress may have a role in the differentiation of mesangial cells. PMID- 10529384 TI - Activity and substrate specificity of the murine STK2 Serine/Threonine kinase that is structurally related to the mitotic regulator protein NIMA of Aspergillus nidulans. AB - We isolated a murine STK2 (mSTK2) cDNA that is homologous to murine Nek1 serine/threonine kinase, a family member related to the cell cycle regulator kinase NIMA of Aspergillus nidulans. Structural comparison demonstrated that the kinase domain of mSTK2 is highly similar to NIMA/Nek family but the C-terminal region is not similar to any proteins except for human STK2 (hSTK2). Similarly to Nek1, mSTK2 is expressed ubiquitously among various organs and is upregulated in the testis. The expression and localization of mSTK2 are not associated with the cell cycle progression of mitogen-activated lymphocyte and DNA-transfected fibroblast. The substrate specificity of mSTK2 is similar to NIMA, but the phosphorylation is observed exclusively upon threonine residues rather than serine. The mSTK2 is shown to be a new member of the NIMA/Nek family with similar substrate specificity, which might participate in a different role from NIMA kinase involved in the cell cycle regulation. PMID- 10529385 TI - Cloning and characterization of a novel p21(Cip1/Waf1)-interacting zinc finger protein, ciz1. AB - p21(Cip1/Waf1) inhibits cell-cycle progression by binding to G1 cyclin/CDK complexes and proliferating cell nuclear antigen (PCNA) through its N- and C terminal domains, respectively. Here, we report a novel p21(Cip1/Waf1) interacting protein, Ciz1 (for Cip1 interacting zinc finger protein), which contains polyglutamine repeats and glutamine-rich region in the N-terminus as well as three zinc-finger motifs and one MH3 (matrin 3-homologous domain 3) in the C-terminal region. Ciz1 bound to the N-terminal, the CDK2-interacting part of p21(Cip1/Waf1), and the interaction was disrupted by the overexpression of CDK2. A region of about 150 amino acids containing the first zinc-finger motif in Ciz1 was the binding site for p21(Cip1/Waf1). When Ciz1 and p21(Cip1/Waf1) were individually overexpressed in U2-OS cells, they mostly localized in the nucleus. However, coexpression of Ciz1 induced cytoplasmic distribution of p21(Cip1/Waf1). These data indicate that Ciz1 is a unique nuclear protein that regulates the cellular localization of p21(Cip1/Waf1). PMID- 10529386 TI - Reconstituted HDL containing human apolipoprotein A-1 reduces VCAM-1 expression and neointima formation following periadventitial cuff-induced carotid injury in apoE null mice. AB - Arterial injury triggers an inflammatory response in part mediated by induction of adhesion molecules such as vascular cell adhesion molecule-1 (VCAM-1) and is implicated in neointimal thickening. Since HDL is known to reduce cytokine activated VCAM-1 expression, we tested the hypothesis that VCAM-1 expression and neontimal thickening following arterial injury are inhibited by reconstituted human HDL containing plasma-derived apoA-1 (rHDL). We used the carotid cuff injury in apoE (-/-) mice fed high cholesterol. Mice received rHDL (40 mg/kg) intravenously every other day for 3 weeks. Compared to control, rHDL treatment inhibited neointima formation (0. 008 +/- 0.004 mm(2) vs. 0.037 +/- 0.019 mm(2); P < 0.01) 21 days after injury, reduced VCAM-1 expression, and decreased monocyte/macrophage infiltration as assessed by histomorphometric analysis within the first week after injury. These changes occurred without any effect on plasma total and HDL cholesterol levels as well as the arterial tissue cholesterol levels. rHDL treatment also reduced the formation of modified lipoprotein in the arterial wall compared to control within the first week after injury. This finding suggests an antioxidant effect of rHDL associated with reduced VCAM-1 expression and neointimal formation after arterial injury. PMID- 10529387 TI - The tumor-suppressing activity of angiostatin protein resides within kringles 1 to 3. AB - Angiostatin protein, which comprises the first four kringle domains of plasminogen, is an endogenous inhibitor of angiogenesis that inhibits the growth of experimental primary and metastatic tumors. Truncation of Angiostatin K1-4 to K1-3 retained the activity of Angiostatin. We recombinantly expressed full-length human Angiostatin protein corresponding to the first four kringle domains of human plasminogen and a truncated form of the Angiostatin protein, kringles 1-3. Purified recombinant Angiostatin K1-3 and K1-4 proteins inhibited the formation of experimental B16-BL6 lung metastases by greater than 80% when administered at 30 nmol/kg/day. We demonstrate for the first time that Angiostatin protein, consisting of the first three kringle domains of human plasminogen, has in vivo biological activity in this assay indistinguishable from that of the full-length Angiostatin K1-4 protein and that the fourth kringle of plasminogen, when linked in sequence to K1-3, plays no direct role in the antitumor activity of Angiostatin. PMID- 10529388 TI - The importance of the putative helices 4 and 5 of human vitamin D(3) receptor for conformation and ligand binding. AB - The 3-D structure of the human vitamin D(3) receptor has not been solved to date. To study the conformation of the ligand binding pocket and the amino acid residues important for binding of calcitriol and its synthetic 20-epi analog MC1288, we have introduced several point mutations into putative helices 4 and 5 of human vitamin D(3) receptor by site-directed mutagenesis. The amino acid residues Ser256, Glu257, Asp258, Gln259, Lys264, Ser265, Ser266, Glu269, Arg274, Ser278, and Phe279 were substituted by alanine. Our results suggest that Arg274 is important for the binding of calcitriol and probably also for the binding of the synthetic vitamin D analog MC1288, whereas Asp258, Gln259, Glu269, and Phe279 may have an important role in stabilizing the conformation of hVDR after ligand binding. PMID- 10529389 TI - Identification of two novel mutations in the hypoglycemic phenotype of very long chain acyl-CoA dehydrogenase deficiency. AB - Very long chain acyl-CoA dehydrogenase (VLCAD) catalyzes the initial step of long chain fatty acid oxidation in the mitochondria. Patients with VLCAD deficiency have recently been observed with two clinical phenotypes. The cardiac form presents with an early onset cardiomyopathy and a high incidence of infant death, while the hypoglycemic form resembles medium chain acyl-CoA dehydrogenase (MCAD) manifesting with hypoketotic hypoglycemia. In our investigation on the molecular basis for these phenotypes, we identified two novel mutations in one VLCAD patient with the hypoglycemic form, a C953T (Pro318Leu) mutation in exon 10 resulting in a substitution of proline 318 by leucine on one allele, and a C1194A (Tyr398Stop) mutation in exon 12 which created a premature stop codon TAA on another allele. The Tyr398Stop mutation may result in a truncated protein or instable messenger RNA. PMID- 10529390 TI - Decreased contraction of glycated collagen lattices coincides with impaired matrix metalloproteinase production. AB - Nonenzymatic glycation of extracellular matrix (ECM) proteins is increased in diabetes mellitus and aging and triggers cellular events leading to an imbalance in ECM homeostasis. We studied the influence of collagen glycation on matrix metalloproteinase production by dermal fibroblasts using the model of lattice cultures. Contraction of glycated collagen lattices was strongly reduced when compared to controls. Meanwhile, fibroblasts synthesized lower amounts of interstitial collagenase (MMP-1). Gelatinase A (MMP-2) production was not modified, but its activation was strongly inhibited. These effects were independent from the intensity of lattice contraction and from any simultaneous modification of tissue inhibitors of metalloproteinase (TIMP-1 and 2) production. These results demonstrate that the impaired ability of fibroblasts to remodel and contract a glycated extracellular matrix coincides with a decrease in MMP production. PMID- 10529391 TI - Photoaffinity labeling of the herpes simplex virus type-1 single-strand DNA binding protein (ICP8) with oligodeoxyribonucleotides. AB - The herpes simplex virus type-1 single-strand DNA-binding protein ICP8 is a 128 kDa zinc metalloprotein. In this communication we have shown that unsubstituted and bromodeoxyuridine-substituted oligonucleotides can be specifically crosslinked to ICP8 by UV irradiation. We have used this approach to show that the single-strand DNA-binding site of ICP8 resides within a 53.5-kDa tryptic polypeptide. This polypeptide initiates at alanine 368 and was estimated to extend through arginine 902. A polypeptide encompassing residues 368-902 synthesized in vitro exhibited single-strand DNA-binding activity. We conclude that the region encompassing residues 368-902 contains the single-strand DNA binding site of ICP8. Moreover, photoaffinity labeling of ICP8 with oligonucleotides provides a means of specifically modifying its single-strand DNA binding site, thereby facilitating future studies on the importance of its single strand DNA-binding activity in its interaction with other DNA replication enzymes. PMID- 10529392 TI - Biophysical characterization of betabellin 16D: a beta-sandwich protein that forms narrow fibrils which associate into broad ribbons. AB - The betabellin structure is a de novo designed beta-sandwich protein consisting of two 32-residue beta sheets packed against one another by hydrophobic interactions. Betabellin 16S (B16S), a 32-residue peptide chain (HSLTAKIakLTFSIAahTYTCAVakYTAKVSH, where a is DAla, h is DHis, and k is DLys), did not have beta structure in water at pH 6.5. Air oxidation of B16S furnished betabellin 16D (B16D), a 64-residue disulfide-bridged two-chain protein, which also did not fold in water at pH 6.5. However, the extent of beta structure observed for B16D increased with pH and ionic strength of the solution and the B16D concentration as observed by circular dichroism spectropolarimetry. Transmission electron microscopy showed that B16D formed narrow fibrils that associated into broad ribbons in 5.0 mM Mops and 0.25 M NaCl at pH 6.9. PMID- 10529393 TI - Trypsinogen stabilization by mutation Arg117-->His: a unifying pathomechanism for hereditary pancreatitis? AB - Mutations Arg117-->His and Asn21-->Ile of the human cationic trypsinogen have been recently identified in patients affected by hereditary pancreatitis (HP). The Arg117-->His substitution is believed to cause pancreatitis by eliminating an essential autolytic cleavage site in trypsin, thereby rendering the protease resistant to inactivation through autolysis. Here we demonstrate that the Arg117- >His mutation also significantly inhibits autocatalytic trypsinogen breakdown under Ca(2+)-free conditions and stabilizes the zymogen form of rat trypsin. Taken together with recent findings demonstrating that the Asn21-->Ile mutation stabilizes rat trypsinogen against autoactivation and consequent autocatalytic degradation, the observations suggest a unifying molecular pathomechanism for HP in which zymogen stabilization plays a central role. PMID- 10529394 TI - Increased susceptibility to apoptosis of human hepatocarcinoma cells transfected with antisense N-acetylglucosaminyltransferase V cDNA. AB - The antisense cDNA of N-acetylglucosaminyltransferase V (GnT-V, EC 2. 4.1.155) was constructed as pcDNA3/GnT-V-AS plasmid and transfected into 7721 cells, a human hepatocarcinoma cell line. The transfection was confirmed with Northern blot. By using HPLC and HRP-lectin staining, it was found that the cells transfected with pcDNA3/GnT-V-AS (GnT-V-AS/7721) expressed less GnT-V activity and beta-1,6-GlcNAc branching in the cell glycoproteins compared with the cells mock-transfected with the vector pcDNA3 (pcDNA3/7721). The growth rate of GnT-V AS/7721 was decreased in serum-containing medium, while the cell death was accelerated in serum-free medium. The GnT-V-AS/7721 cells were more susceptible to the apoptosis induced by ATRA than the mock-transfected cells. This was evidenced by the obvious appearance of a hypoploid sub-G(1) fraction in the DNA histogram using FCM analysis, the more condensed new moon-type nuclei under morphological observation, and the more intensive TUNEL reaction for assaying the fragmented DNA. At the same time as GnT-V down-regulation by GnT-V-AS, an increase of another N-aceylglusaminyltransferase, GnT-III (EC 2.4.1.144), was observed, and the biological significance of this finding was discussed. PMID- 10529395 TI - Chromosomal instability of fanconi anemia cells is not the consequence of a defective repair activity of the ribosomal protein S3. AB - Fanconi anemia (FA) is an autosomal recessive chromosomal breakage disorder characterized by developmental defects, hypersensitivity toward oxygen and DNA crosslinking agents, and susceptibility to cancer. An increased level of reactive oxygen intermediates and an increased level of 8-oxoguanine in FA cells point to a defective oxygen metabolism. Recent investigations showed that FA cells from several complementation groups have a reduced capacity to repair oxidatively damaged DNA. One major enzyme involved in the repair of oxidative DNA lesions is the ribosomal protein S3. Previous reports implied a role for the ribosomal protein S3 in DNA repair in FA cells. However, a more detailed analysis of the ribosomal protein S3 in FA cells from complementation groups A-E could not confirm this. DNA analysis and Western blot analysis did not show significant differences in ribosomal protein S3 between FA cells and cells from healthy individuals. Furthermore, even the overexpression of the ribosomal protein S3 did not reduce the chromosomal instability of FA cells. PMID- 10529396 TI - Overproduction of the outer-membrane proteins FepA and FhuE responsible for iron transport in Escherichia coli hfq::cat mutant. AB - We found that 82- and 76-kDa proteins in the outer-membrane fractions were overproduced in the hfq::cat mutant cells when grown in synthetic media. Expression of these proteins was repressed by addition of FeCl(3) in the mutant as well as in the wild type. It was revealed that these are FepA and FhuE proteins involved in iron transport. The hfq::cat mutant was more susceptible to killing by hydrogen peroxide, probably due to the excess incorporation of iron, which potentially generates hydroxyl radicals. Increased incorporation of iron in the hfq::cat mutant was also confirmed by the suppressive effect on the ftsH1 mutation. These results suggest that the hfq gene product is involved in the defense mechanism against oxidative stress. PMID- 10529397 TI - Phosphorylation/Dephosphorylation steps are crucial for the induction of CYP2B1 and CYP2B2 gene expression by phenobarbital. AB - The effects of several protein kinase activators and protein phosphatase inhibitors on the phenobarbital (PB)-induced gene expression of CYP2B1 and CYP2B2 (CYP2B1/2B2) in adult rat hepatocytes were investigated. Insulin, epidermal growth factor, interleukin 6, cAMP, phorbol 12-myristate 13-acetate, tumor necrosis factor alpha, vanadate, and okadaic acid were found to suppress the induction of CYP2B1/2B2 at mRNA and protein levels in hepatocytes. cAMP and vanadate completely suppressed the induction of CYP2B1/2B2 gene expression in both rat hepatocytes and liver. The addition of genistein to vanadate-treated hepatocytes partially recovered the induction of CYP2B1/2B1 gene expression by PB. These results of the present study demonstrate that phosphorylation/dephosphorylation steps are crucial for the induction of CYP2B1/2B2 gene expression by PB. PMID- 10529398 TI - In vitro transcriptional and translational block of the bcl-2 gene operated by peptide nucleic acid. AB - The antisense and antigene activity of peptide nucleic acid (PNA) targeted to the human B-cell lymphoma (bcl)-2 gene was evaluated in vitro. Several PNAs complementary to different sequences of bcl-2, including the start codon and the 5'-untranslated region (5'-UTR), were tested. One PNA directed against the AUG start codon and another recognizing the 5'-UTR were able to specifically reduce Bcl-2 protein synthesis in a cell-free system; however, only partial inhibition (80 and 54%, respectively) was obtained when they were used singularly. Complete translation block was obtained with the simultaneous presence of both PNAs. A triplex-forming bis-PNA was targeted to a homopurine sequence on the coding strand of the bcl-2 cDNA. In an in vitro transcription assay this PNA specifically inhibited the transcription of bcl-2 at concentrations as low as 300 nM, with the concomitant appearance of a truncated 200-base-long product. These results demonstrate the ability of PNA to selectively modulate both translation and transcription of bcl-2 in vitro and suggest its potential use as an antisense and an antigene agent in order to downregulate bcl-2 expression in tumors. PMID- 10529399 TI - N(2)-(1-Carboxyethyl)deoxyguanosine, a nonenzymatic glycation adduct of DNA, induces single-strand breaks and increases mutation frequencies. AB - N(2)-(1-Carboxyethyl)deoxyguanosine (CEdG) is a major nonenzymatic glycation product of DNA. The effect of CEdG modification, which was specifically prepared by incubation with dihydroxyacetone, on plasmid DNA topology was evaluated by gel electrophoresis. A time-dependent decrease of supercoiled plasmid-DNA was observed in parallel to the increase of CEdG adducts; the half-life time of the supercoiled plasmid-DNA was estimated to be approximately 16-18 h. CEdG-modified plasmid DNA showed a 25-fold reduced transformation efficiency. When modified DNA was used to transform Escherichia coli cells, a 6-fold increase in mutation frequency was determined by measuring loss of alpha-complementation. For the mutator strain BMH71-18mutS, an 8-fold increase in mutation frequency was observed. Although the exact mechanism of DNA damage is unclear, the occurrence of spontaneous depurination is likely. These findings suggest that a defined DNA glycation reaction can lead to DNA damage in vivo. PMID- 10529401 TI - A novel mutant of green fluorescent protein with enhanced sensitivity for microanalysis at 488 nm excitation. AB - Green fluorescent protein (GFP) has been utilized as a powerful reporter of gene expression and protein localization in cells. We discovered a mutant carrying point mutation S208L from a UV-excitable GFP (F99S/M153T/V163A). It had the enhanced fluorescence intensity. Introduction of the red-shifted mutations (F64L/S65T) to this mutant led to the GFP having the brightest mutants reported which were expressed in Escherichia coli and excited at 488 nm. The relative fluorescence intensities to that of wild-type GFP and GFPuv were increased about 120- and 10-fold, respectively. It was shown that the S208L mutation contributes to both a higher intrinsic brightness of GFP and a higher expression level in E. coli. PMID- 10529400 TI - Akt phosphorylation site found in human caspase-9 is absent in mouse caspase-9. AB - Caspase-9 is one caspase upstream of caspase-3 and its activation is stimulated by Apaf-1/cytochrome c and inhibited by Akt signals. BAD phosphorylation by Akt is an essential step for growth factor-mediated inhibition of caspase activation. Recently, it was shown that human caspase-9 is phosphorylated by Akt and that its protease activity is reduced. To clarify the molecular mechanism of regulation of caspase-9 activation in neuronal apoptosis, we isolated two alternative splicing products of mouse caspase-9, caspase-9L and caspase-9S, from a P19 embryonal carcinoma cell cDNA library. Curiously, the Akt phosphorylation sites and motifs found in human caspase-9 were absent in both mouse caspase-9L and -9S. Mouse caspase-9 was not phosphorylated by activated Akt in vitro. Reverse transcription polymerase chain reaction analysis showed that the absent Akt motif is not limited to caspase-9 expressed in P19 embryonal carcinoma cells but also occurs in caspase-9 expressed in mouse, rat, and monkey. These results suggest that inhibition of caspase-9 activation by Akt-dependent phosphorylation is not generalized across species. PMID- 10529402 TI - Nuclear factor kappaB activity is essential for matrix metalloproteinase-1 and -3 upregulation in rabbit dermal fibroblasts. AB - Expression of matrix metalloproteinases (MMPs)-1 and -3 in fibroblasts is upregulated by pro-inflammatory cytokines and growth factors during proliferative inflammatory processes, including wound healing and rheumatoid arthritis. The Activator Protein-1 (AP-1) transcription factor is essential but, we show here, not sufficient for upregulation because platelet derived growth factor (PDGF) and basic fibroblast growth factor (bFGF), which strongly activate AP-1, poorly induce MMP-1 and -3. Interleukin-1alpha, which activates nuclear factor-kappaB (NF-kappaB), synergistically upregulates MMP-1 and -3 expression in the presence of bFGF or PDGF. Adenovirus mediated overexpression of IkappaBalpha, the inhibitor of NF-kappaB, completely suppresses MMP-1 and -3 protein and mRNA expression. Hence, we show for the first time that (NF-kappaB) activity is also essential for MMP-1 and -3 upregulation. PMID- 10529403 TI - Molecular cloning of macrophin, a human homologue of Drosophila kakapo with a close structural similarity to plectin and dystrophin. AB - We have determined the complete cDNA coding sequence of a novel cytoskeletal protein by the degenerative primer-mediated PCR strategy. This novel gene, named as macrophin (microfilament and actin filament cross-linker protein related to plectin and dystrophin, Accession No. AB029290), appears to be a human homologue of a Drosophila gene, kakapo, and shows close similarity to plectin and dystrophin on the search of BLAST homology-computed database. Comparison of the deduced protein sequences for macrophin and kakapo revealed that they were 66% similar, and both of them have an NH2-terminal actin-binding domain, a central rod region composed of spectrin-like repeats, and COOH-terminal Gas2-related region. The predicted sequences for macrophin are 5430 amino acids in length with a calculated molecular mass of 620 kDa, which is one of the largest size idenfied in human cytoskeletal proteins. High expression of macrophin was observed in brain, heart, lung, placenta, liver, kidney, and pancreas. In pancreas, we found that macrophin was specifically expressed in acinar cells than islet cells by in situ hybridization. By using radiation hybrid panel, we have mapped the macrophin gene to the chromosome 1p31-32. PMID- 10529404 TI - Secretory IgA-mediated basophil activation. II. Roles of GTP-binding regulatory proteins and phosphatidylinositol 3-kinase. AB - Secretory IgA (sIgA) is the most abundant Ig isotype in mucous secretions in the upper and lower airways, where basophils exert effector functions during allergic inflammation. We recently demonstrated that immobilized sIgA on Sepharose beads is capable of inducing basophil degranulation ( approximately 15% of total histamine). To investigate the detailed mechanisms of this degranulation, we established in this study a new assay system for sIgA-mediated basophil activation. Immobilized sIgA on a plastic surface induced strong histamine release ( approximately 50% of total histamine) comparable to anti-IgE, and we analyzed the nature of basophil signal transduction by sIgA using various inhibitors. sIgA-induced basophil histamine release was inhibited completely by pertussis toxin, but anti-IgE-induced release was not affected. sIgA-mediated release was also inhibited by phosphatidylinositol 3-kinase (PI 3-kinase) inhibitor, wortmannin. These results strongly suggest that sIgA activates basophils via an IgE-independent novel mechanism involving both Gi protein and PI 3-kinase. PMID- 10529405 TI - Vitellogenin-6 is a major carbonylated protein in aged nematode, Caenorhabditis elegans. AB - Recently we found that protein carbonyl content increases with age in wild-type as well as long- and short-lived Caenorhabditis elegans nematodes in inverse correlation with life span (Adachi et al., J. Gerontol. 53A, B240-B244, 1998; Yasuda et al., J. Gerontol. 54A, B47-B51, 1999). In the present study, we investigated carbonyl modification of individual proteins in young and old wild type nematodes by two-dimensional immunoblot using antibodies against 2, 4 dinitrophenylhydrazones. A protein with apparent molecular weight of 110 kDa was found to be a major carbonylated protein in aged animals. Amino acid sequence of peptide fragments of the protein was identical to that of vitellogenin-6, a yolk protein synthesized in and secreted from the intestine during egg-laying stage. Although the function(s) of the protein in aged nematodes is unclear, we suggest that the protein may have a role to protect other cellular components from oxidation because of its metal binding capacity. PMID- 10529406 TI - The expression of Escherichia coli SOS genes recA and uvrA is inducible by polyamines. AB - Polyamines are polycationic compounds that have been implicated in a variety of cellular processes. We first report that the expression of recA and uvrA genes in Escherichia coli is inducible by polyamines. The recA and uvrA genes were effectively inducible by spermine (tetra-amine) and less effectively inducible by spermidine (tri-amine). On the other hand, both genes were not significantly inducible by putrescine (diamine) and divalent cation Mg(2+). The expression of both genes was dependent on the charge of the polyamine in the order of spermine (+4), spermidine (+3), and putrescine (+2). The induction of recA and uvrA genes by polyamines showed a dose-dependent relationship and no synergistic effects. Introduction of gyrA mutation conferring DNA relaxation increased the basal expression of recA gene about 2.5-fold compared to the wild type, but did not significantly affect the polyamine-dependent induction ratios of the recA gene. The basal and the polyamine-dependent expression of uvrA gene are not dependent on gyrA mutation. These results suggest that the polyamine-dependent expression of recA and uvrA genes may be regulated in a different way. Our results indicated that polyamines are involved in the SOS induction of recA and uvrA genes at transcriptional levels in E. coli. PMID- 10529407 TI - 1,25-Dihydroxyvitamin D(3) and prostaglandin E(2) act directly on circulating human osteoclast precursors. AB - 1,25-Dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) and prostaglandin E(2) (PGE(2)) are known to influence osteoclast formation indirectly through their effects on osteoblasts. To determine whether 1, 25(OH)(2)D(3) and PGE(2) also have a direct effect on circulating osteoclast precursors, these factors were added to long term cultures of human peripheral blood mononuclear cells (PBMCs) in the presence of osteoprotegerin ligand and macrophage colony-stimulating factor (M-CSF) (+/ dexamethasone). The number of TRAP(+) and VNR(+) multinucleated cells and the area of lacunar resorption were decreased when 1,25(OH)(2)D(3) alone was added. A marked increase in resorption pit formation was noted when the combination of 1, 25(OH)(2)D(3) and dexamethasone was added to PBMC cultures. Dose-dependent inhibition of osteoclast formation and lacunar resorption was seen when PGE(2) was added to PBMC cultures in both the presence and the absence of dexamethasone. Thus, 1,25(OH)(2)D(3) and PGE(2) not only influence osteoclast formation in the presence of bone stromal cells but also act directly on circulating osteoclast precursors to influence osteoclast differentiation. PMID- 10529408 TI - Recovery of catalytic activity from an inactive aggregated mutant of l-aspartase. AB - Two highly conserved lysyl residues have been replaced with an arginine to examine their role in the mechanism of l-aspartase from Escherichia coli. Replacement of an active-site lysine results in a significant loss of catalytic efficiency [A. S. Saribas, J. F. Schindler, and R. E. Viola (1994) J. Biol. Chem. 269, 6313-6319], while replacement of the second lysine leads to a completely inactive and insoluble protein. Fluorescence spectral evidence has suggested that the loss of activity is due to the misfolding of this aspartase mutant. Some catalytic activity is recovered when the mutant is treated with varying levels of denaturants, and extended treatment with high levels of guanidine.HCl results in the recovery of a substantial fraction of the wild-type activity from this inactive mutant. However, upon removal of the denaturant this mutant enzyme slowly reverts to its inactive and insoluble form. Treatment with an artificial chaperone system in which solubilization by detergent is followed by its removal with beta-cyclodextrin leads to a stable enzyme under nondenaturing conditions with about half the catalytic activity of the wild-type enzyme. These results confirm a structural role for lysine-55 in l-aspartase and demonstrate that additional characterization is required before conclusions can be drawn from the production of an inactive mutant. PMID- 10529409 TI - Protection by protein A of apoptotic cell death caused by anti-AIDS drug zidovudine. AB - Zidovudine, the anti-AIDS drug, caused inhibition of mitogen-induced proliferation and perturbation of cell-cycle progression of cultured bone marrow cells of mice. There was significant hypoploidy observed in flow cytometric analysis of AZT-treated bone marrow cells. In apo-direct analysis, cells showed apoptosis in G0/G1 phase. In DNA gel analysis, characteristic laddering of apoptosis was observed in AZT-treated bone marrow cells. We demonstrated that, when the animals were pretreated with protein A (PA) of Staphylococcus aureus, the apoptotic changes could be prevented in bone marrow cells of AZT-treated animals. There is a significant (p < 0.05) increase in proliferation of bone marrow cells subjected to mitogen treatment in PA+AZT-treated animals, compared to only AZT-treated animals. However, cell-cycle phase distribution was not hampered and no laddering in DNA gel analysis was also observed in this group. In apo-direct analysis, PA treatment showed significant (p < 0.001) inhibition of AZT-induced apoptosis. These observations indicate that by using a suitable agent such as protein A the toxic side effects of AZT could be minimized. PMID- 10529410 TI - Combined enhancement of microtubule assembly and glucose metabolism in neuronal systems in vitro: decreased sensitivity to copper toxicity. AB - Brain cell-free extract greatly stimulates the polymerization rate of purified tubulin with a reduction of the nucleation period and without a significant alteration of the final assembly state. This effect is mimicked by neuroblastoma extract at 10-fold lower extract concentration, but not by excess muscle extract. Copper inhibits microtubule assembly in vitro but in the presence of brain extract the copper effect is suspended. Electron microscopic images showed that intact microtubules are formed and decorated by cytosolic proteins in the absence and presence of copper, while the copper alone induces the formation of S-shaped sheets and oligomeric threads. The flux of triosephosphate formation from glucose is enhanced by microtubules in brain extract, but not in muscle extract. Copper inhibits the glycolytic flux; however, the presence of microtubules not only suspends the inhibition by copper but the activation of glycolysis by microtubules is also preserved. We conclude that the organization of neuronal proteins modifies both the rates of microtubule assembly and glycolysis, and reduces their sensitivities against the inhibition caused by copper. PMID- 10529411 TI - Lim1 is required in both primitive streak-derived tissues and visceral endoderm for head formation in the mouse. AB - Lim1 is a homeobox gene expressed in the extraembryonic anterior visceral endoderm and in primitive streak-derived tissues of early mouse embryos. Mice homozygous for a targeted mutation of Lim1 lack head structures anterior to rhombomere 3 in the hindbrain. To determine in which tissues Lim1 is required for head formation and its mode of action, we have generated chimeric mouse embryos and performed tissue layer recombination explant assays. In chimeric embryos in which the visceral endoderm was composed of predominantly wild-type cells, we found that Lim1(-)(/)(-) cells were able to contribute to the anterior mesendoderm of embryonic day 7.5 chimeric embryos but that embryonic day 9.5 chimeric embryos displayed a range of head defects. In addition, early somite stage chimeras generated by injecting Lim1(-)(/)(-) embryonic stem cells into wild-type tetraploid blastocysts lacked forebrain and midbrain neural tissue. Furthermore, in explant recombination assays, anterior mesendoderm from Lim1( )(/)(-) embryos was unable to maintain the expression of the anterior neural marker gene Otx2 in wild-type ectoderm. In complementary experiments, embryonic day 9.5 chimeric embryos in which the visceral endoderm was composed of predominantly Lim1(-)(/)(-) cells and the embryo proper of largely wild-type cells, also phenocopied the Lim1(-)(/)(-) headless phenotype. These results indicate that Lim1 is required in both primitive streak-derived tissues and visceral endoderm for head formation and that its inactivation in these tissues produces cell non-autonomous defects. We discuss a double assurance model in which Lim1 regulates sequential signaling events required for head formation in the mouse. PMID- 10529412 TI - Compartments and organising boundaries in the Drosophila eye: the role of the homeodomain Iroquois proteins. AB - The Drosophila eye is patterned by a dorsal-ventral organising centre mechanistically similar to those in the fly wing and the vertebrate limb bud. Here we show how this organising centre in the eye is initiated - the first event in retinal patterning. Early in development the eye primordium is divided into dorsal and ventral compartments. The dorsally expressed homeodomain Iroquois genes are true selector genes for the dorsal compartment; their expression is regulated by Hedgehog and Wingless. The organising centre is then induced at the interface between the Iroquois-expressing and non-expressing cells at the eye midline. It was previously thought that the eye develops by a mechanism distinct from that operating in other imaginal discs, but our work establishes the importance of lineage compartments in the eye and thus supports their global role as fundamental units of patterning. PMID- 10529413 TI - fatvg encodes a new localized RNA that uses a 25-nucleotide element (FVLE1) to localize to the vegetal cortex of Xenopus oocytes. AB - Vegetally localized transcripts have been implicated in a number of important biological functions, including cell fate determination and embryonic patterning. We have isolated a cDNA, fatvg, which encodes a localized maternal transcript that exhibits a localization pattern reminiscent of Vg1 mRNA. fatvg is the homologue of a mammalian gene expressed in adipose tissues. The fatvg transcript, unlike Vg1 which localizes strictly through the Late pathway, also associates with the mitochondrial cloud that is characteristic of the METRO or Early pathway. This suggests that fatvg mRNA may utilize both the METRO and Late pathways to localize to the vegetal cortex during oogenesis. We have dissected the cis-acting localization elements of fatvg mRNA and compared these elements with Vg1 mRNA. Our results indicate that, like most localized RNAs, in a variety of systems, transcripts of fatvg contain localization elements in the 3'UTR. The 3'UTR of fatvg mRNA contains multiple elements that are able to function independently; however, it functions most efficiently when all of the elements are present. We have defined a short 25-nucleotide element that can direct vegetal localization as a single copy. This element differs in sequence from previously described Vg1 localization elements, suggesting that different localization elements are involved in the localization of RNAs through the Late pathway. PMID- 10529414 TI - Anteroposterior patterning of the epidermis by inductive influences from the vegetal hemisphere cells in the ascidian embryo. AB - Patterning along the anteroposterior axis is a critical step during animal embryogenesis. Although mechanisms of anteroposterior patterning in the neural tube have been studied in various chordates, little is known about those of the epidermis. To approach this issue, we investigated patterning mechanisms of the epidermis in the ascidian embryo. First we examined expression of homeobox genes (Hrdll-1, Hroth, HrHox-1 and Hrcad) in the epidermis. Hrdll-1 is expressed in the anterior tip of the epidermis that later forms the adhesive papillae, while Hroth is expressed in the anterior part of the trunk epidermis. HrHox-1 and Hrcad are expressed in middle and posterior parts of the epidermis, respectively. These data suggested that the epidermis of the ascidian embryo is patterned anteroposteriorly. In ascidian embryogenesis, the epidermis is exclusively derived from animal hemisphere cells. To investigate regulation of expression of the four homeobox genes in the epidermis by vegetal hemisphere cells, we next performed hemisphere isolation and cell ablation experiments. We showed that removal of the vegetal cells before the late 16-cell stage results in loss of expression of these homeobox genes in the animal hemisphere cells. Expression of Hrdll-1 and Hroth depends on contact with the anterior-vegetal (the A-line) cells, while expression of HrHox-1 and Hrcad requires contact with the posterior vegetal (the B-line) cells. We also demonstrated that contact with the vegetal cells until the late 32-cell stage is sufficient for animal cells to express Hrdll-1, Hroth and Hrcad, while longer contact is necessary for HrHox-1 expression. Contact with the A-line cells until the late 32-cell stage is also sufficient for formation of the adhesive papillae. Our data indicate that the epidermis of the ascidian embryo is patterned along the anteroposterior axis by multiple inductive influences from the vegetal hemisphere cells and provide the first insight into mechanisms of epidermis patterning in the chordate embryos. PMID- 10529415 TI - Msx1 antagonizes the myogenic activity of Pax3 in migrating limb muscle precursors. AB - The migration of myogenic precursors to the vertebrate limb exemplifies a common problem in development - namely, how migratory cells that are committed to a specific lineage postpone terminal differentiation until they reach their destination. Here we show that in chicken embryos, expression of the Msx1 homeobox gene overlaps with Pax3 in migrating limb muscle precursors, which are committed myoblasts that do not express myogenic differentiation genes such as MyoD. We find that ectopic expression of Msx1 in the forelimb and somites of chicken embryos inhibits MyoD expression as well as muscle differentiation. Conversely, ectopic expression of Pax3 activates MyoD expression, while co ectopic expression of Msx1 and Pax3 neutralizes their effects on MyoD. Moreover, we find that Msx1 represses and Pax3 activates MyoD regulatory elements in cell culture, while in combination, Msx1 and Pax3 oppose each other's trancriptional actions on MyoD. Finally, we show that the Msx1 protein interacts with Pax3 in vitro, thereby inhibiting DNA binding by Pax3. Thus, we propose that Msx1 antagonizes the myogenic activity of Pax3 in migrating limb muscle precursors via direct protein-protein interaction. Our results implicate functional antagonism through competitive protein-protein interactions as a mechanism for regulating the differentiation state of migrating cells. PMID- 10529416 TI - Bmp activity establishes a gradient of positional information throughout the entire neural plate. AB - Bone morphogenetic proteins (Bmps) are key regulators of dorsoventral (DV) patterning. Within the ectoderm, Bmp activity has been shown to inhibit neural development, promote epidermal differentiation and influence the specification of dorsal neurons and neural crest. In this study, we examine the patterning of neural tissue in mutant zebrafish embryos with compromised Bmp signalling activity. We find that although Bmp activity does not influence anteroposterior (AP) patterning, it does affect DV patterning at all AP levels of the neural plate. Thus, we show that Bmp activity is required for specification of cell fates around the margin of the entire neural plate, including forebrain regions that do not form neural crest. Surprisingly, we find that Bmp activity is also required for patterning neurons at all DV levels of the CNS. In swirl/bmp2b(-) (swr(-)) embryos, laterally positioned sensory neurons are absent whereas more medial interneuron populations are hugely expanded. However, in somitabun(-) (sbn(-)) embryos, which probably retain higher residual Bmp activity, it is the sensory neurons and not the interneurons that are expanded. Conversely, in severely Bmp depleted embryos, both interneurons and sensory neurons are absent and it is the most medial neurons that are expanded. These results are consistent with there being a gradient of Bmp-dependent positional information extending throughout the entire neural and non-neural ectoderm. PMID- 10529417 TI - Translational regulation of oskar mRNA occurs independent of the cap and poly(A) tail in Drosophila ovarian extracts. AB - Translational regulation plays a prominent role in Drosophila body patterning. Progress in elucidating the underlying mechanisms has been limited by the lack of a homologous in vitro system that supports regulation. Here we show that extracts prepared from Drosophila tissues are competent for translation. Ovarian extracts, but not embryonic extracts, support the Bruno response element- and Bruno dependent repression of oskar mRNA translation, which acts in vivo to prevent protein synthesis from transcripts not localized to the posterior pole of the oocyte. Consistent with suggestive evidence from in vivo experiments, regulation in vitro does not involve changes in poly(A) tail length. Moreover, inhibition studies strongly suggest that repression does not interfere with the process of 5' cap recognition. Translational regulation mediated through the Bruno response elements is thus likely to occur via a novel mechanism. PMID- 10529418 TI - Self-organization of periodic patterns by dissociated feather mesenchymal cells and the regulation of size, number and spacing of primordia. AB - Periodic patterning is a fundamental organizing process in biology. Using a feather reconstitution assay, we traced back to the initial stage of the patterning process. Cells started from an equivalent state and self-organized into a periodic pattern without previous cues or sequential propagation. When different numbers of dissociated mesenchymal cells were confronted with a piece of same-sized epithelium, the size of feather primordia remained constant, not the number or interbud spacing, suggesting size determination is intrinsic to dissociated cells. Increasing bone morphogenetic protein (BMP) receptor expression in mesenchymal cells decreased the size of primordia while antagonizing the BMP pathway with Noggin increased the size of primordia. A threshold number of mesenchymal cells with a basal level of adhesion molecules such as NCAM were sufficient to trigger the patterning process. The process is best visualized by the progressive restriction of beta-catenin transcripts in the epidermis. Therefore, feather size, number and spacing are modulated through the available morphogen ligands and receptors in the system. PMID- 10529419 TI - Compensatory defects associated with mutations in Hoxa1 restore normal palatogenesis to Hoxa2 mutants. AB - The rhombencephalic neural crest play several roles in craniofacial development. They give rise to the cranial sensory ganglia and much of the craniofacial skeleton, and are vital for patterning of the craniofacial muscles. The loss of Hoxa1 or Hoxa2 function affects the development of multiple neural crest-derived structures. To understand how these two genes function together in craniofacial development, an allele was generated that disrupts both of these linked genes. Some of the craniofacial defects observed in the double mutants were additive combinations of those that exist in each of the single mutants, indicating that each gene functions independently in the formation of these structures. Other defects were found only in the double mutants demonstrating overlapping or synergistic functions. We also uncovered multiple defects in the attachments and trajectories of the extrinsic tongue and hyoid muscles in Hoxa2 mutants. Interestingly, the abnormal trajectory of two of these muscles, the styloglossus and the stylohyoideus, blocked the attachment of the hyoglossus to the greater horn of the hyoid, which in turn correlated exactly with the presence of cleft palate in Hoxa2 mutants. We suggest that the hyoglossus, whose function is to depress the lateral edges of the tongue, when unable to make its proper attachment to the greater horn of the hyoid, forces the tongue to adopt an abnormal posture which blocks closure of the palatal shelves. Unexpectedly, in Hoxa1/Hoxa2 double mutants, the penetrance of cleft palate is dramatically reduced. We show that two compensatory defects, associated with the loss of Hoxa1 function, restore normal attachment of the hyoglossus to the greater horn thereby allowing the palatal shelves to lift and fuse above the flattened tongue. PMID- 10529420 TI - Mice mutant for both Hoxa1 and Hoxb1 show extensive remodeling of the hindbrain and defects in craniofacial development. AB - The analysis of mice mutant for both Hoxa1 and Hoxb1 suggests that these two genes function together to pattern the hindbrain. Separately, mutations in Hoxa1 and Hoxb1 have profoundly different effects on hindbrain development. Hoxa1 mutations disrupt the rhombomeric organization of the hindbrain, whereas Hoxb1 mutations do not alter the rhombomeric pattern, but instead influence the fate of cells originating in rhombomere 4. We suggest that these differences are not the consequences of different functional roles for these gene products, but rather reflect differences in the kinetics of Hoxa1 and Hoxb1 gene expression. In strong support of the idea that Hoxa1 and Hoxb1 have overlapping functions, Hoxa1/Hoxb1 double mutant homozygotes exhibit a plethora of defects either not seen, or seen only in a very mild form, in mice mutant for only Hoxa1 or Hoxb1. Examples include: the loss of both rhombomeres 4 and 5, the selective loss of the 2(nd) branchial arch, and the loss of most, but not all, 2(nd) branchial arch-derived tissues. We suggest that the early role for both of these genes in hindbrain development is specification of rhombomere identities and that the aberrant development of the hindbrain in Hoxa1/Hoxb1 double mutants proceeds through two phases, the misspecification of rhombomeres within the hindbrain, followed subsequently by size regulation of the misspecified hindbrain through induction of apoptosis. PMID- 10529421 TI - Development of the cardiac conduction system involves recruitment within a multipotent cardiomyogenic lineage. AB - The cardiac pacemaking and conduction system sets and maintains the rhythmic pumping action of the heart. Previously, we have shown that peripheral cells of the conduction network in chick (periarterial Purkinje fibers) are selected within a cardiomyogenic lineage and that this recruitment occurs as a result of paracrine cues from coronary arteries. At present, the cellular derivation of other elements of this specialized system (e.g. the nodes and bundles of the central conduction system) are controversial, with some proposing that the evidence supports a neurogenic and others a myogenic origin for these tissues. While such ontological questions remain, it is unlikely that progress can be made on the molecular mechanisms governing patterning and induction of the central conduction system. Here, we have undertaken lineage-tracing strategies based on the distinct properties of replication-incompetent adenoviral and retroviral lacZ expressing constructs. Using these complementary approaches, it is shown that cells constituting both peripheral and central conduction tissues originate from cardiomyogenic progenitors present in the looped, tubular heart with no detectable contribution by migratory neuroectoderm-derived populations. Moreover, clonal analyses of retrovirally infected cells incorporated within any part of the conduction system suggest that such cells share closer lineage relationships with nearby contractive myocytes than with other, more distal elements of the conduction system. Differentiation birthdating by label dilution using [(3)H]thymidine also demonstrates the occurrence of ongoing myocyte conscription to conductive specialization and provides a time course for this active and localized selection process in different parts of the system. Together, these data suggest that the cardiac conduction system does not develop by outgrowth from a prespecified pool of 'primary' myogenic progenitors. Rather, its assembly and elaboration occur via processes that include progressive and localized recruitment of multipotent cardiomyogenic cells to the developing network of specialized cardiac tissues. PMID- 10529422 TI - Key roles of retinoic acid receptors alpha and beta in the patterning of the caudal hindbrain, pharyngeal arches and otocyst in the mouse. AB - Mouse fetuses carrying targeted inactivations of both the RAR(&agr;) and the RARbeta genes display a variety of malformations in structures known to be partially derived from the mesenchymal neural crest originating from post-otic rhombomeres (e.g. thymus and great cephalic arteries) (Ghyselinck, N., Dupe, V., Dierich, A., Messaddeq, N., Garnier, J.M., Rochette-Egly, C., Chambon, P. and Mark M. (1997). Int. J. Dev. Biol. 41, 425-447). In a search for neural crest defects, we have analysed the rhombomeres, cranial nerves and pharyngeal arches of these double null mutants at early embryonic stages. The mutant post-otic cranial nerves are disorganized, indicating that RARs are involved in the patterning of structures derived from neurogenic neural crest, even though the lack of RARalpha and RARbeta has no detectable effect on the number and migration path of neural crest cells. Interestingly, the double null mutation impairs early developmental processes known to be independent of the neural crest e.g., the initial formation of the 3rd and 4th branchial pouches and of the 3rd, 4th and 6th arch arteries. The double mutation also results in an enlargement of rhombomere 5, which is likely to be responsible for the induction of supernumerary otic vesicles, in a disappearance of the rhombomere 5/6 boundary, and in profound alterations of rhombomere identities. In the mutant hindbrain, the expression domain of kreisler is twice its normal size and the caudal stripe of Krox-20 extends into the presumptive rhombomeres 6 and 7 region. In this region, Hoxb-1 is ectopically expressed, Hoxb-3 is ectopically up-regulated and Hoxd-4 expression is abolished. These data, which indicate that retinoic acid signaling through RARalpha and/or RARbeta is essential for the specification of rhombomere identities and for the control of caudal hindbrain segmentation by restricting the expression domains of kreisler and of Krox-20, also strongly suggest that this signaling plays a crucial role in the posteriorization of the hindbrain neurectoderm. PMID- 10529423 TI - Infertility associated with incomplete spermatogenic arrest and oligozoospermia in Egr4-deficient mice. AB - Male fertility is complex and depends upon endocrine/paracrine regulatory mechanisms and morphogenetic processes occurring during testicular development, spermatogenesis (mitosis and meiosis) and spermiogenesis (spermatid maturation). Egr4 (NGFI-C, pAT133), a member of the Egr family of zinc-finger transcription factors, is thought to be involved in cellular growth and differentiation, but its specific function has been previously unknown. We derived Egr4 null mice through targeted mutagenesis and found that they were phenotypically normal with the exception that males, but not females, were infertile. Egr4 is expressed at low levels within male germ cells during meiosis and is critical for germ cell maturation during the early-mid pachytene stage. While most Egr4 null male germ cells undergo apoptosis during early-mid pachytene, some are capable of maturing beyond an apparent Egr4-dependent developmental restriction point. Consequently, a limited degree of spermiogenesis occurs but this is accompanied by markedly abnormal spermatozoon morphology and severe oligozoospermia. Egr4 appears to regulate critical genes involved in early stages of meiosis and has a singularly important role in male murine fertility. These data raise the possibility that Egr4 may contribute to some forms of human idiopathic male infertility. PMID- 10529424 TI - Development of erythroid and myeloid progenitors in the yolk sac and embryo proper of the mouse. AB - In this study, we have mapped the onset of hematopoietic development in the mouse embryo using colony-forming progenitor assays and PCR-based gene expression analysis. With this approach, we demonstrate that commitment of embryonic cells to hematopoietic fates begins in proximal regions of the egg cylinder at the mid primitive streak stage (E7.0) with the simultaneous appearance of primitive erythroid and macrophage progenitors. Development of these progenitors was associated with the expression of SCL/tal-1 and GATA-1, genes known to be involved in the development and maturation of the hematopoietic system. Kinetic analysis revealed the transient nature of the primitive erythroid lineage, as progenitors increased in number in the developing yolk sac until early somite pair stages of development (E8.25) and then declined sharply to undetectable levels by 20 somite pairs (E9.0). Primitive erythroid progenitors were not detected in any other tissue at any stage of embryonic development. The early wave of primitive erythropoiesis was followed by the appearance of definitive erythroid progenitors (BFU-E) that were first detectable at 1-7 somite pairs (E8.25) exclusively within the yolk sac. The appearance of BFU-E was followed by the development of later stage definitive erythroid (CFU-E), mast cell and bipotential granulocyte/macrophage progenitors in the yolk sac. C-myb, a gene essential for definitive hematopoiesis, was expressed at low levels in the yolk sac just prior to and during the early development of these definitive erythroid progenitors. All hematopoietic activity was localized to the yolk sac until circulation was established (E8.5) at which time progenitors from all lineages were detected in the bloodstream and subsequently in the fetal liver following its development. This pattern of development suggests that definitive hematopoietic progenitors arise in the yolk sac, migrate through the bloodstream and seed the fetal liver to rapidly initiate the first phase of intraembryonic hematopoiesis. Together, these findings demonstrate that commitment to hematopoietic fates begins in early gastrulation, that the yolk sac is the only site of primitive erythropoiesis and that the yolk sac serves as the first source of definitive hematopoietic progenitors during embryonic development. PMID- 10529425 TI - oto is a homeotic locus with a role in anteroposterior development that is partially redundant with Lim1. AB - Genetic control of mammalian head development involves mechanisms that are shared with trunk development as well as mechanisms that are independent. For example, mutations in the nodal gene disrupt axis formation and head development while mutations in the Otx2 or Lim1 genes block head development without disrupting development of the trunk. We show here that the oto mutation on mouse chromosome 1 defines a locus with a critical role in anterior development. The oto mutation disrupts development of the telencephalic and optic vesicles, the pharyngeal endoderm and the first branchial arch. Also, oto embryos have dose-dependent, posterior homeotic transformations throughout the axial skeleton. To further dissect the role of the oto locus in head development, we crossed mice carrying oto and Lim1 mutations. Interactions between the two mutations indicate that the role of oto in the regulation of head development is partially redundant with that of Lim1. The phenotype of oto embryos points to an early and critical role for oto in the development of forebrain subregions. Transformations of the vertebrae in oto embryos reveal a Lim1-independent role in the establishment of positional information in the trunk. PMID- 10529426 TI - The subcellular localization and activity of Drosophila cubitus interruptus are regulated at multiple levels. AB - Cubitus interruptus (Ci), a Drosophila transcription factor, mediates Hedgehog (Hh) signaling during the patterning of embryonic epidermis and larval imaginal discs. In the absence of Hh signal, Ci is cleaved to generate a truncated nuclear form capable of transcriptional repression. Hh signaling stabilizes and activates the full-length Ci protein leading to strong activation of downstream target genes including patched and decapentaplegic. A number of molecules have been implicated in the regulation of Ci. Mutations in these molecules lead to changes in Ci protein level, the extent of Ci proteolysis and the expression of Ci target genes. This paper examines the regulation of Ci subcellular localization and activity. We first characterize a bipartite nuclear localization signal (NLS) within Ci. We propose that the subcellular distribution of Ci is affected by two opposing forces, the action of the NLS and that of at least two regions targeting Ci to the cytoplasm. Further our data show that loss of PKA or Costal-2 activity does not fully mimic Hh signaling, demonstrating that Ci proteolysis and Ci activation are two distinct events which are regulated through different paths. Finally, we propose that there are three levels of apparent Ci activity, corresponding to three zones along the AP axis with different sets of gene expression and different levels of Hh signaling. PMID- 10529427 TI - A novel fork head gene mediates early steps during Xenopus lens formation. AB - Xlens1 is a novel Xenopus member of the fork head gene family, named for its nearly restricted expression in the anterior ectodermal placode, presumptive lens ectoderm (PLE), and anterior epithelium of the differentiated lens. The temporal and spatial restriction of its expression suggests that: (1) Xlens1 is transcribed initially at neural plate stages in response to putative signals from the anterior neural plate that transform lens-competent ectoderm to lens-biased ectoderm; (2) further steps in the process of lens-forming bias restrict Xlens1 expression to the presumptive lens ectoderm (PLE) during later neural plate stages; (3) interactions with the optic vesicle maintain Xlens1 expression in the lens placode; and (4) Xlens1 expression is downregulated as committed lens cells undergo terminal differentiation. Induction assays demonstrate that pax6 induces Xlens1 expression, but unlike pax6, Xlens1 cannot induce the expression of the lens differentiation marker beta-crystallin. In the whole embryo, overexpression of Xlens1 in the lens ectoderm causes it to thicken and maintain gene expression characteristics of the PLE. Also, this overexpression suppresses differentiation in the lens ectoderm, suggesting that Xlens1 functions to maintain specified lens ectoderm in an undifferentiated state. Misexpression of Xlens1 in other regions causes hypertrophy of restricted tissues but only occasionally leads ectopic sites of gamma-crystallin protein expression in select anterior head regions. These results indicate that Xlens1 expression alone does not specify lens ectoderm. Lens specification and differentiation likely depends on a combination of other gene products and an appropriate level of Xlens1 activity. PMID- 10529428 TI - A petunia MADS box gene involved in the transition from vegetative to reproductive development. AB - We have identified a novel petunia MADS box gene, PETUNIA FLOWERING GENE (PFG), which is involved in the transition from vegetative to reproductive development. PFG is expressed in the entire plant except stamens, roots and seedlings. Highest expression levels of PFG are found in vegetative and inflorescence meristems. Inhibition of PFG expression in transgenic plants, using a cosuppression strategy, resulted in a unique nonflowering phenotype. Homozygous pfg cosuppression plants are blocked in the formation of inflorescences and maintain vegetative growth. In these mutants, the expression of both PFG and the MADS box gene FLORAL BINDING PROTEIN26 (FBP26), the putative petunia homolog of SQUAMOSA from Antirrhinum, are down-regulated. In hemizygous pfg cosuppression plants initially a few flowers are formed, after which the meristem reverts to the vegetative phase. This reverted phenotype suggests that PFG, besides being required for floral transition, is also required to maintain the reproductive identity after this transition. The position of PFG in the hierarchy of genes controlling floral meristem development was investigated using a double mutant of the floral meristem identity mutant aberrant leaf and flower (alf) and the pfg cosuppression mutant. This analysis revealed that the pfg cosuppression phenotype is epistatic to the alf mutant phenotype, indicating that PFG acts early in the transition to flowering. These results suggest that the petunia MADS box gene, PFG, functions as an inflorescence meristem identity gene required for the transition of the vegetative shoot apex to the reproductive phase and the maintenance of reproductive identity. PMID- 10529429 TI - Engrailed defines the position of dorsal di-mesencephalic boundary by repressing diencephalic fate. AB - Regionalization of a simple neural tube is a fundamental event during the development of central nervous system. To analyze in vivo the molecular mechanisms underlying the development of mesencephalon, we ectopically expressed Engrailed, which is expressed in developing mesencephalon, in the brain of chick embryos by in ovo electroporation. Misexpression of Engrailed caused a rostral shift of the di-mesencephalic boundary, and caused transformation of dorsal diencephalon into tectum, a derivative of dorsal mesencephalon. Ectopic Engrailed rapidly repressed Pax-6, a marker for diencephalon, which preceded the induction of mesencephalon-related genes such as Pax-2, Pax-5, Fgf8, Wnt-1 and EphrinA2. In contrast, a mutant Engrailed, En-2(F51rE), bearing mutation in EH1 domain, which has been shown to interact with a co-repressor, Groucho, did not show the phenotype induced by wild-type Engrailed. Furthermore, VP16-Engrailed chimeric protein, the dominant positive form of Engrailed, caused caudal shift of di mesencephalic boundary and ectopic Pax-6 expression in mesencephalon. These data suggest that (1) Engrailed defines the position of dorsal di-mesencephalic boundary by directly repressing diencephalic fate, and (2) Engrailed positively regulates the expression of mesencephalon-related genes by repressing the expression of their negative regulator(s). PMID- 10529430 TI - Regulation of Hox target genes by a DNA bound Homothorax/Hox/Extradenticle complex. AB - To regulate their target genes, the Hox proteins of Drosophila often bind to DNA as heterodimers with the homeodomain protein Extradenticle (EXD). For EXD to bind DNA, it must be in the nucleus, and its nuclear localization requires a third homeodomain protein, Homothorax (HTH). Here we show that a conserved N-terminal domain of HTH directly binds to EXD in vitro, and is sufficient to induce the nuclear localization of EXD in vivo. However, mutating a key DNA binding residue in the HTH homeodomain abolishes many of its in vivo functions. HTH binds to DNA as part of a HTH/Hox/EXD trimeric complex, and we show that this complex is essential for the activation of a natural Hox target enhancer. Using a dominant negative form of HTH we provide evidence that similar complexes are important for several Hox- and exd-mediated functions in vivo. These data suggest that Hox proteins often function as part of a multiprotein complex, composed of HTH, Hox, and EXD proteins, bound to DNA. PMID- 10529431 TI - Expression and functional analysis of Cititf1, an ascidian NK-2 class gene, suggest its role in endoderm development. AB - In solitary ascidians the fate of endoderm is determined at a very early stage of development and depends on cytoplasmic factors whose nature has not been determined. We have isolated a member of the NK-2 gene family, Cititf1, from the ascidian Ciona intestinalis, showing high sequence homology to mammalian TITF1. The Cititf1 gene was expressed in all endodermal precursors at the pregastrula and gastrula stages, and is thus the first specific regulatory endodermal marker to be isolated from an ascidian. Cititf1 expression was downregulated at the end of gastrulation to reappear at middle tailbud and larval stages in the most anterior and ventral parts of head endoderm, regions which give rise, after metamorphosis, to the adult endostyle, where Cititf1 mRNA was still present. Microinjection of Cititf1 mRNA into fertilized eggs resulted in tadpole larvae with abnormalities in head-trunk development consequent to the formation of excess endoderm, perhaps due to recruitment of notochord precursors to an endodermal fate. These data suggest that Cititf1 plays an important role in normal endoderm differentiation during ascidian embryogenesis. PMID- 10529432 TI - Migration of the Drosophila primordial midgut cells requires coordination of diverse PS integrin functions. AB - Cell migration during embryogenesis involves two populations of cells: the migrating cells and the underlying cells that provide the substratum for migration. The formation of the Drosophila larval midgut involves the migration of the primordial midgut cells along a visceral mesoderm substratum. We show that integrin adhesion receptors are required in both populations of cells for normal rates of migration. In the absence of the PS integrins, the visceral mesoderm is disorganised, the primordial midgut cells do not display their normal motile appearance and their migration is delayed by 2 hours. Removing PS integrin function from the visceral mesoderm alone results in visceral mesoderm disorganization, but only causes a modest delay in migration and does not affect the appearance of the migrating cells. Removing PS integrin function from the migrating cells causes as severe a delay in migration as the complete loss of PS integrin function. The functions of PS1 and PS2 are specific in the two tissues, endoderm and mesoderm, since they cannot substitute for each other. In addition there is a partial redundancy in the function of the two PS integrins expressed in the endoderm, PS1 (alphaPS1betaPS) and PS3 (alphaPS3betaPS), since loss of just one alpha subunit in the midgut results in either a modest delay (alphaPS1) or no effect (alphaPS3). We have also examined the roles of small GTPases in promoting migration of the primordial midgut cells. We find that dominant negative (N17) versions of Rac and Cdc42 cause a very similar defect in migration as loss of integrins, while those of Rho and Ras have no effect. Thus integrins are involved in mediating migration by creating an optimal substratum for adhesion, adhering to that substratum and possibly by activating Rac and Cdc42. PMID- 10529433 TI - Anterior patterning by synergistic activity of the early gastrula organizer and the anterior germ layer tissues of the mouse embryo. AB - Fragments of the germ layer tissues isolated from the early-primitive-streak (early-streak) stage mouse embryos were tested for axis induction activity by transplantation to late-gastrula (late-streak to early-bud) stage host embryos. The posterior epiblast fragment that contains the early gastrula organizer was able to recruit the host tissues to form an ectopic axis. However, the most anterior neural gene that was expressed in the ectopic axis was Krox20 that marks parts of the hindbrain, but markers of the mid- and forebrain (Otx2 and En1) were not expressed. Anterior visceral endoderm or the anterior epiblast alone did not induce any ectopic neural tissue. However, when these two anterior germ layer tissues were transplanted together, they can induce the formation of ectopic host derived neural tissues but these tissues rarely expressed anterior neural genes and did not show any organization of an ectopic axis. Therefore, although the anterior endoderm and epiblast together may display some inductive activity, they do not act like a classical organizer. Induction of the anterior neural genes in the ectopic axis was achieved only when a combination of the posterior epiblast fragment, anterior visceral endoderm and the anterior epiblast was transplanted to the host embryo. The formation of anterior neural structures therefore requires the synergistic interaction of the early gastrula organizer and anterior germ layer tissues. PMID- 10529434 TI - Defects in mouse mammary gland development caused by conditional haploinsufficiency of Patched-1. AB - In vertebrates, the hedgehog family of cell signaling proteins and associated downstream network components play an essential role in mediating tissue interactions during development and organogenesis. Loss-of-function or misexpression mutation of hedgehog network components can cause birth defects, skin cancer and other tumors. The mammary gland is a specialized skin derivative requiring epithelial-epithelial and epithelial-stromal tissue interactions similar to those required for development of other organs, where these interactions are often controlled by hedgehog signaling. We have investigated the role of the Patched-1 (Ptc1) hedgehog receptor gene in mammary development and neoplasia. Haploinsufficiency at the Ptc1 locus results in severe histological defects in ductal structure, and minor morphological changes in terminal end buds in heterozygous postpubescent virgin animals. Defects are mainly ductal hyperplasias and dysplasias characterized by multilayered ductal walls and dissociated cells impacting ductal lumens. This phenotype is 100% penetrant. Remarkably, defects are reverted during late pregnancy and lactation but return upon involution and gland remodeling. Whole mammary gland transplants into athymic mice demonstrates that the observed dysplasias reflect an intrisic developmental defect within the gland. However, Ptc1-induced epithelial dysplasias are not stable upon transplantation into a wild-type epithelium-free fat pad, suggesting stromal (or epithelial and stromal) function of Ptc1. Mammary expression of Ptc1 mRNA is both epithelial and stromal and is developmentally regulated. Phenotypic reversion correlates with developmentally regulated and enhanced expression of Indian hedgehog (Ihh) during pregnancy and lactation. Data demonstrate a critical mammary role for at least one component of the hedgehog signaling network and suggest that Ihh is the primary hedgehog gene active in the gland. PMID- 10529435 TI - Treating asthma with anti-IgE or anti-IL5. AB - In the last decades, several key mechanisms driving asthma pathophysiooogy have been discovered. These include the role of IgE in allergic disease, and the role of IL-5 in eosinophilic inflammation. In the last few years, tools to block each of these have been developed. At this time, early clinical studies with neutralizing antibodies against both IgE and IL-5 have been performed in asthma patients, with promising results, and larger studies are underway. The mechanisms of, and possible role of, both anti-IgE and anti-IL-5 treatment in asthma are discussed in this review article. PMID- 10529436 TI - Recent advances in matrix metalloproteinase inhibitors research. AB - Excess MMP proteolytic activity has been associated with a wide variety of pathological conditions such as arthritis, cancer and heart failure. The potential utility of MMP inhibitors as therapeutic interventions in these diverse and important disease states has led to an intense effort toward the development of such inhibitors. The first generation of compounds were peptide-like broad spectrum inhibitors, active against a broad range of MMPs. However, the induction of musculoskeletal side effects seen in clinical trials with these agents has emphasized the need for a better understanding of the role that each of the MMPs plays in normal tissue turnover and disease progression. Advances in our ability to engineer and synthesize selective inhibitors as well as the discovery of small molecule, non-peptidic inhibitors has spurred an intense effort to identify potent and bioavailable second generation compounds. There are now several such compounds targeted against various subsets of the MMPs in clinical development. This review will focus on the design and structure activity relationships of these second generation compounds. PMID- 10529437 TI - Melanotransferrin is produced by senile plaque-associated reactive microglia in Alzheimer's disease. AB - Melanotransferrin (MTf), also known as p97, has been localized in capillary endothelial cells of human brain. In Alzheimer's-diseased (AD) brain tissues, reactive microglial cells located in senile plaques exhibit elevated levels of MTf. The localization of the p97 protein may reflect its site of synthesis or could reflect a paracrine site of action. We examined the expression of MTf mRNA by in situ hybridization histochemistry using AD and healthy brain tissues. We also examined normal liver tissues by immunohistochemistry and in situ hybridization. In all the brain tissues examined, capillaries had positive signals for MTf mRNA. In AD tissues, expression of MTf mRNA appeared in reactive microglial cells in the grey matter specifically associated with dense plaques. In liver tissues, immunohistochemistry using anti-p97 antibody demonstrated that sinusoids were positively stained. In addition, in situ hybridization histochemistry revealed that hepatocytes had positive signals. These results suggest that p97 expression in reactive microglial cells are closely related to AD pathology. These results also support the notion that p97, which appears elevated in the cerebral spinal fluid and serum of AD patients, originates in the reactive microglia associated with dense senile plaques. Thus, p97 is a unique cellular hallmark of AD and further suggests that metal transport mechanisms play a role in this disease. PMID- 10529438 TI - Drosophila neurons respond differently to hypoxia and cyanide than rat neurons. AB - Compared with mammalian species, Drosophila melanogaster exhibits marked tolerance to hypoxia or anoxia. However, the underlying cellular mechanisms of tolerance are still largely unknown. In order to assess the electrophysiologic response to O(2) lack in Drosophila neurons and compare them to those in mammals, we used neurons from embryonic cultures of both Drosophila and rat. We studied the effects of hypoxia on membrane potential V(m), input resistance R(m), rheobase, and action potential characteristics before, during and after 3 to 5 min of hypoxia (measured PO(2)<20 Torr). In Drosophila neurons, on the one hand, V(m) reversibly hyperpolarized with hypoxia by an average of about 20 mV and input resistance decreased by 71% from control. In most cells studied, action potential (AP) amplitude decreased, its duration increased, and its threshold shifted in a hyperpolarized direction before AP generation was attenuated. On the other hand, V(m) in rat cortical neurons reversibly depolarized by an average of 10 mV with hypoxia. Input resistance was reversibly reduced by 58% and, in most cells studied, AP amplitude also decreased and its duration increased. In contrast to the effects of hypoxia on V(m), CN caused a depolarization by 22 mV and a slight increase in R(m) in Drosophila. In the rat, CN was similar to hypoxia in its effect on R(m). We conclude that (1) rat and Drosophila neurons decrease their excitability in hypoxia by activating different mechanisms; (2) the most likely explanation for the hyperpolarization and the decrease in R(m) in Drosophila neurons is the activation of a K(+) conductance; this activation, by itself, cannot explain the results in rat neurons and (3) hypoxia and cyanide have similar effects in rat neurons but are divergent in their effects in Drosophila neurons. PMID- 10529439 TI - Behavioral responses to stress are intact in CRF-deficient mice. AB - Corticotropin-releasing factor (CRF) has been implicated in endocrine and behavioral responses associated with stress. We have now studied the behavior of mice lacking the CRF gene (CRFko), comparing them to wild-type (WT) mice. Behaviors were observed in untreated mice, as well as following restraint or intraperitoneal administration of mouse interleukin-1beta (mIL-1beta). In the multicompartment chamber (MCC), the behaviors of CRFko and WT mice were very similar, and prior restraint and IL-1beta induced similar decreases in stimulus contact times in both genotypes. In the elevated plus maze (EPM), restraint decreased the number of open arm entries but the behavior of both genotypes was very similar. In the open field (OF), the changes in locomotor activity in response to restraint were similar in both genotypes, although CRFko mice displayed slightly increased locomotor activity compared to WT mice. In both the MCC and the EPM, grooming behavior was increased by restraint, and was higher in the CRFko than in the WT mice. Compared to WT mice, CRFko mice had lower basal plasma concentrations of corticosterone which did not increase significantly following footshock. Thus, CRFko mice showed a clear dichotomy; the stress related activation of the hypothalamo-pituitary-adrenal (HPA) axis was absent, whereas the stress-related behavioral responses thought to be mediated by brain CRF were unaffected. These results suggest that when mice develop in the absence of CRF, another factor (or factors) assumes the behavioral functions normally ascribed to brain CRF, but not activation of the HPA axis. Alternatively, the natural modulator of behavior may not be CRF, but some other molecule that can act on receptors sensitive to CRF. Thus, redundant CNS mechanisms appear to be involved in stress-related behaviors. PMID- 10529440 TI - GDNF partially protects grafted fetal dopaminergic neurons against 6 hydroxydopamine neurotoxicity. AB - Rats were given unilateral 6-hydroxydopamine (6-OHDA) lesions and subsequently received transplants of fetal ventral mesencephalic tissue into the denervated striatum. Four weeks later transplanted animals were tested for graft-mediated reduction of amphetamine-induced rotational behavior. Subsequently, transplanted animals received an intrastriatal injection of either GDNF (10 microg) or citrate buffer into a site lateral to the transplant, and then 6 h later received an injection of either 4.0 microg of 6-OHDA, 8.0 microg of 6-OHDA, or vehicle using the same stereotaxic coordinates that were used for the GDNF/citrate buffer injection. Animals were re-tested for amphetamine-induced rotational behavior 2 weeks later. Histological analysis revealed a significant reduction in the number of cell bodies immunostained for tyrosine hydroxylase (TH+) within the transplant for those animals pretreated with an intrastriatal injection of citrate buffer and subsequently given either dose of 6-OHDA. Transplanted animals pretreated with GDNF and subsequently administered 8.0 microg of 6-OHDA showed a significant reduction of TH+ neurons within the transplant compared to controls, however TH+ cell counts for this group remained significantly higher than the TH+ cell counts for the group of animals receiving the same dose of 6-OHDA but pretreated with citrate buffer. GDNF pretreatment completely protected TH+ cell bodies against 4.0 microg of 6-OHDA. Rotational scores indicated that GDNF provided only partial protection against 6-OHDA neurotoxicity in terms of transplant function. For both groups of transplanted animals receiving GDNF pretreatment and 6-OHDA injections, amphetamine-induced rotational scores dropped below the scores for animals pretreated with citrate buffer but remained significantly higher than the scores for transplanted animals that were not injected with 6-OHDA. Both histological and behavioral measures indicate GDNF partially protects integrated transplants against neurotoxic insult. PMID- 10529441 TI - Responses to hypoxia of petrosal ganglia in vitro. AB - NaCN is a classical stimulus used to elicit discharges from carotid body chemoreceptors. The effect is assumed to be mediated by glomus (type I) cells, which release an excitatory transmitter for the excitation of carotid nerve endings. Since the sensory perikarya of the glossopharyngeal nerve (from which the carotid nerve branches) are located in the petrosal ganglion, we tested whether application of this drug to the petrosal ganglion superfused in vitro elicits antidromic discharges in the carotid nerve. NaCN did indeed cause an intense and prolonged burst of nerve impulses in the carotid nerve, while provoking a less intense and much briefer burst of discharges in the glossopharyngeal branch. Carotid nerve responses to NaCN were reduced and shortened by prior or following application of dopamine to the ganglion. Sodium azide applied to the petrosal ganglion evoked a less intense and much briefer burst of impulses in the carotid nerve. Ganglionar application of 2,4 dinitrophenol did not induce discharges in the carotid nerve. Switching the superfusion of the ganglion from a normoxic to a hypoxic solution did not evoke discharges in the carotid nerve. Therefore, the perikarya of carotid nerve neurons are sensitive to NaCN, but are not excited by reducing the pO(2) of the superfusing solution. PMID- 10529443 TI - Effect of vitamin E intake on levels of vitamins E and C in the central nervous system and peripheral tissues: implications for health recommendations. AB - Vitamin E (alpha-gamma-tocopherol) is an important component in biological membranes. A decrease in its concentration imposes structural and functional damage to the cells. The object of this study was to assess the effect of a graded dietary vitamin E (E) intake on E concentration in specific regions of the brain, and its influence on vitamin C levels and neurological function. Following a 2-month period, rats supplemented with 5, 30, 60, 250 or 500 mg all-rac-alpha tocopherol-acetate/kg diet (mg E/kg diet) exhibited a significant increase of E concentration in brain and peripheral tissues. However, while blood and liver showed a dose response increase in E concentration which correlated well with the different levels of E in the diet, the central nervous system (CNS) followed the same pattern of increase of vitamin E in brain tissue only when the diet was supplemented with 5, 30, or 60 mg E/kg diet. No further increase in E concentration was observed when the diet was supplemented with 250 or 500 mg E/kg diet. Similarly, the heart tissue showed a significant increase in its E concentration when the was enriched with 5, 30, or 60 mg E/kg diet, with no further increases at 250 or 500 mg. Vitamin C concentration in brain cortex and cerebellum, plasma, liver, and heart was reduced in the groups receiving 250 or 500 mg E/kg diet. Compared to the low E group, rats supplemented with the 60, 250 or 500 mg E/kg diet showed a significant enhancement in striatal dopamine (DA) release, but no differences were observed among the latter three groups. PMID- 10529442 TI - Effects of L-arginine on the afferent resting activity in the cephalopod statocyst. AB - The effects of bath application of the nitric oxide (NO) precursor L-arginine (L ARG) on the resting activity (RA) of afferent crista fibers were studied in isolated statocysts of the cuttlefish Sepia officinalis under various experimental conditions. L-ARG (threshold 10(-7) M) had three different effects: inhibition, excitation, and excitation followed by an inhibition; only the inhibitory effect of L-ARG was dose-dependent. D-Arginine (D-ARG) had no effect. When the preparation was pre-treated with NO synthase inhibitors (N(G)-Nitric-L arginine methyl ester HCl (L-NAME), N(G)-Nitro-L-arginine (L-NOARG)), both the inhibitory and the excitatory effects of L-ARG significantly decreased at higher concentrations (10(-5 to -4) M), or were completely blocked at lower concentrations (10(-7 to -6) M), of L-ARG. When the preparation was pre-treated with guanylate cyclase inhibitors (1H-[1,2, 4]oxadiazolo[4,3,-a]quinoxalin-1-one (ODQ), methylene blue (M-BLU), cystamine (CYS)), L-ARG had only excitatory effects, whereas its effects were only inhibitory when the preparation was pre treated with adenylate cyclase inhibitors 2',3'-dideoxyadenosine (DDA), MDL 12330A (MDL), nicotinic acid (NIC-A)). L-ARG had no effects when the pre treatment was with a guanylate cyclase inhibitor and an adenylate cyclase inhibitor combined; in that situation, the RA of the afferent fibers remained. These data indicate that in cephalopod statocysts, a cGMP and a cAMP signal transduction pathway (presumably via the generation of NO) are responsible for the effects of L-ARG on the RA of crista afferent fibers. They also indicate that the L-ARG-cGMP pathway is the dominant pathway and is inhibitory, and that both pathways have only modulatory effects on, but are not essential for, the generation of the RA. PMID- 10529444 TI - Differential forebrain c-fos mRNA induction by ether inhalation and novelty: evidence for distinctive stress pathways. AB - Previous studies indicate the existence of subtypes of stressors invoking distinct patterns of neuronal integration. Pathways activated by stress appear to depend on whether the perceived threat is processive/neurogenic or systemic in nature. To test this hypothesis, the present study compares magnitude and extent of c-fos mRNA induction in response to novelty (open field (OF), representing a processive stressor) or ether exposure (representing a systemic stressor). Exposure to the OF or ether fumes both produced increases in plasma corticosterone (CORT) levels; notably, peak levels of secretion were elevated in the ether group, suggestive of augmented HPA secretory activity to this stressor. In situ hybridization analysis of c-fos mRNA induction reveals common and divergent activational properties in the two stress groups. The extent of c-fos mRNA expression was similar in the paraventricular nucleus (PVN), despite stress related differences in CORT secretion. Analysis of the dorsomedial hypothalamic nucleus, suprachiasmatic nucleus (SCN) and limbic sites, specifically, the lateral septum and medial amygdaloid nucleus, indicate greater c-fos mRNA induction in animals exposed to OF stress. The frontoparietal cortex was only forebrain region showing differential activation by ether. Differential c-fos induction was not observed in the medial preoptic area (ventrolateral quadrant), paraventricular thalamus, dorsolateral striatum or hippocampus. The results indicate that processive and systemic stressors differ not only in the patterning of neuronal activation in the CNS, but also in the extent to which selected stress-sensitive regions are induced. PMID- 10529445 TI - Behavior-related changes in the activity of substantia nigra pars reticulata neurons in freely moving rats. AB - As one of the primary targets of the striatum, the substantia nigra pars reticulata (SNr) has been hypothesized to play a role in normal motor behavior. Specifically, inhibition of usually high, tonic SNr output is predicted to correlate with motor activation. While support for this has come primarily from electrophysiological studies in primates performing goal-directed movements, we tested this hypothesis in rats behaving in an open-field arena. SNr single-unit activity was recorded during spontaneous bouts of open-field behavior (e.g., head and body movements, locomotion) and after rats were given D-amphetamine (1.0 mg/kg, s.c.), which reliably increases motor activity and elevates the firing of motor-related striatal neurons. Prior to drug administration, SNr neurons had either regular, slightly irregular or irregular firing patterns when animals rested quietly. During movement, some inhibitions were observed, but the majority ( approximately 79%) of analyzed units increased firing by as much as 38%. Regardless of the predrug behavioral response of the cell, amphetamine strongly inhibited firing rate ( approximately 90% below nonmovement baseline) and changed firing pattern such that all cells fired irregularly. Subsequent injection with the dopamine antagonist haloperidol (1.0 mg/kg, s.c.) reversed amphetamine induced inhibitions in all tested cells, which supports a role for dopamine in this effect. These results suggest that the pattern of striatal activity established by amphetamine, which may be critical for determining the drug induced behavioral pattern, is represented in the SNr regardless of the predrug behavioral response of the cell. PMID- 10529446 TI - Characterization of the central cell groups regulating the kidney in the rat. AB - Retrograde, transneuronal viral tracing technique combined with neurotransmitter immunohistochemistry was used to identify the type of neurons in spinal cord and brain that project to the rat's kidney. Pseudorabies virus (PRV) injections were made into the left kidney. After an incubation of 4 days postinjection, PRV infected neurons were located immunocytochemically in the ipsilateral intermediolateral (IML) cell column of the spinal cord and several brainstem cell groups: medullary raphe nuclei, ventromedial medulla (VMM), rostral ventrolateral medulla (RVLM), A5 cell group and the hypothalamic paraventricular nucleus (PVH). In the medulla, serotonin (5-HT)-immunoreactive neurons of the caudal raphe nuclei, substance P (SP)-immunoreactive neurons of the raphe obscurus (ROb) nuclei and tyrosine hydroxylase (TH)-immunoreactive neurons of A5 cells were infected. In the VMM and RVLM, immunoreactive phenylethanolamine-N methyltransferase (PNMT) neurons were infected. Some PRV-infected neurons in VMM contain 5-HT immunoreactivity. In the hypothalamus, immunoreactive vasopressin (VP) and oxytocin (OT) neurons were infected with PRV. This work indicates that sympathetic outflow to kidney is regulated by different types of neurons and the bulbospinal pathways regulating sympathetic outflow to the kidney are not obviously different from those regulating the other visceral, e.g., adrenal, heart, etc. PMID- 10529447 TI - Amygdala-kindled and pentylenetetrazole-induced seizures in glutamate transporter GLAST-deficient mice. AB - The glutamatergic system has been shown to be important for the induction of epileptiform activity and the development of epileptogenesis. To investigate the role of the astroglial glutamate transporter GLAST in epileptogenesis, we examined amygdala (AM)-kindled and pentylenetetrazole (PTZ)-induced seizures in GLAST-deficient mice (GLAST(-/-)) and compared them to those observed in wild type mice (GLAST(+/+)) and maternal C57Black6/J (C57) mice. AM-kindling resulted in no significant differences in afterdischarge threshold or in the seizure responses induced by first stimulation between these groups. In addition, although no significant differences were seen in kindled seizure development, the generalized seizure duration of AM-kindled seizures in GLAST(-/-) mice was significantly prolonged (approximately 35%) compared with that of C57 mice. Furthermore, GLAST(-/-) mice showed more severe stages of PTZ-induced seizures than GLAST(+/+) mice, and the latency to the onset of seizures was significantly shorter for the mutant mice. These results indicate that GLAST is one of factors determining seizure susceptibility. PMID- 10529448 TI - Protein kinase C inhibition blocks the early appearance of vestibular compensation. AB - This study tests the hypothesis that activation of protein kinase C (PKC) is a critical step for early recovery from spontaneous nystagmus after unilateral ablation of the vestibular periphery. Halothane-NO(2)-O(2)-anesthetized Long Evans rats received a 5-microl intracerebroventricular bolus of vehicle (distilled water, six rats), PKC inhibitor [Iso-H-7 (10 mM, four rats; 50 mM, five rats) or bisindolemaleimide I (Bis-I, 10 microM six rats)], PKG and PKA inhibitor (A-3, 1 mM, six rats), or the serine-threonine protein kinase inhibitor H-7 (1 mM, five rats; 10 mM, five rats). Surgical unilateral labyrinthectomy (UL) was completed within 15 min. Sham control groups showed no nystagmus. Bis-I and Iso-H-7 significantly retarded the disappearance of spontaneous nystagmus quick phases for 8 h after UL (p<0.05). The effects of Iso-H-7 were dose-dependent: more nystagmus quick phases (p<0.05) were present in the 50 mM than the 10 mM group at 7 and 8 h post-UL. The rats given A-3 showed a delayed retardation of nystagmus loss, which differed significantly (p<0.05) from controls at 4-8 h after labyrinthectomy. The number of nystagmus quick phases was significantly greater than controls (p<0. 05) in the 10 mM H-7 group at 4, 5, 6 and 48 h post UL, but only at 6 and 24 h post-UL in the 1 mM H-7 group. Thus, PKC activation is an important early requirement for vestibular compensation during the acute post labyrinthectomy period, while cyclic-nucleotide dependent kinases may be important in a later time frame. PMID- 10529449 TI - MK-801 administration blocks the effects of a 5-HT(2A/2C) agonist on melatonin rhythmicity and c-fos induction in the suprachiasmatic nucleus. AB - Both excitatory amino acids and serotonin have been implicated in the photic control of rhythms, but they have rarely been considered to interact. This study investigated the effects of the NMDA receptor antagonist, MK-801 on the phase shift of the melatonin rhythm and the induction of c-fos in the rat suprachiasmatic nucleus (SCN) provoked by the administration of the serotonin agonist DOI ((+/-)-1-(4-Iodo-2,5-dimethoxyphenyl)-2-aminopropane hydrochloride). The urinary excretion rate rhythm of the melatonin metabolite, 6 sulphatoxymelatonin was delayed by administration of DOI (0.5 mg/kg) at CT18 (6 h after subjective darkness onset) as previously reported by our group. Administration of MK-801 (3 mg/kg) 30 min before DOI blocked the shift in the onset of excretion of the melatonin metabolite on the following nights. Pre treatment with MK-801 also inhibited by approximately 90% the induction of c-fos in the SCN by DOI at ZT18 (6 h after actual darkness onset) as determined by immunohistochemistry. These results provide evidence for a role of excitatory amino acids in the photomimetic effects of serotonin 5-HT(2C) agonists in the rat. PMID- 10529450 TI - Neuroprotective effect of high-dose albumin therapy against global ischemic brain injury in rats. AB - The purpose of this study was to determine whether treatment with high-dose human serum albumin (HSA) would offer protection in a model of high-grade transient forebrain ischemia. Twenty-six fasted Wistar rats underwent bilateral common carotid artery occlusion and severe hypotension (50 mmHg) for 10 min. The agent (25% HSA) or vehicle (0.9% NaCl) was administered i.v. 5 min after termination of ischemia. HSA-treated rats showed significantly improved neurological deficits throughout a 7-day survival period. Histologically, HSA-treated rats showed 2.4- to 5.3-fold increases in numbers of surviving CA1 hippocampal pyramidal neurons compared to saline-treated animals. These results document that high-dose albumin therapy instituted 5 min after global ischemia significantly improves neurological score and reduces histological damage. PMID- 10529451 TI - Sensitization of methamphetamine-induced disorganization of daily locomotor activity rhythm in male rats. AB - Methamphetamine (MAP) was administered to rats through drinking water repeatedly (three sessions, one session:administration for 60 days followed by withdrawal of 30 days) in order to examine whether or not MAP-induced disorganization of daily activity rhythm is sensitized. Each session (60 days) was divided into six blocks of 10 days. In the 1st session, daily locomotor activity rhythm of rats became disorganized around at 40 days (4th block) after the start of MAP drinking. However, MAP-induced disorganization of daily activity rhythm appeared at 20 days (2nd block) in the 2nd session and at 10 days (1st block) in the 3rd session following re-start of MAP drinking. On the other hand, the amount of MAP intake was decreased on the 2nd and 3rd sessions as compared with the 1st session. These results indicate that the mechanism of MAP-induced disorganization of daily activity rhythm may involve sensitization. PMID- 10529452 TI - Comparison of behavioral responses to noxious cold and heat in mice. AB - We investigated behavioral responses to noxious cold and heat stimuli in mice. Similar to the hot-plate test, mice showed licking or jumping responses on a cold plate (0 degrees C). The sensitivity to noxious heat (55 degrees C) was not correlated to the sensitivity to noxious cold, indicating that nociceptive processing of cold and heat are different. Behavioral responses to noxious cold are inhibited by systemic morphine or intrathecal administration of morphine. Lesion of the medial frontal cortex, including the anterior cingulate cortex, or selective activation of two types of opioid receptors in the anterior cingulate cortex produces dose-dependent antinociceptive effects on behavioral responses to noxious cold stimuli. These results suggest that activation of opioid receptors in the anterior cingulate cortex can produce powerful antinociception. PMID- 10529453 TI - Expression of beta-galactosidase gene and endothelial nitric oxide synthase gene in rat vascular smooth muscle cells after in vitro lipotransfection. AB - The aim of this study was to optimize the conditions for in vitro lipotransfection of rat vascular smooth muscle cells (VSMC) with bacterial beta galactosidase gene and bovine endothelial nitric oxide synthase (ecNOS) gene. Transfection efficiency of four liposomes: Transfectam, Lipofectin, Unifectin-10, and Maxifectin was compared. The best results (efficiency 1-5%) were obtained with Maxifectin, when transfections were performed in VSMC cultures being at 50% confluency, with 1 microg DNA and 10 microl liposome per well, and when the liposome/DNA complexes were coincubated with the cells for 24 h. This method allowed detection of the transgene activity 12 h after the beginning of the transfection, with maximum values between the second and fourth days. The expression of the potentially therapeutic ecNOS gene was evidenced by confirmation of ecNOS mRNA generation, indirect detection of active ecNOS protein and by measurement of nitrite ion accumulation in the medium from the transfected cell cultures. PMID- 10529454 TI - Low density lipoprotein immunoapheresis does not increase plasma lipid peroxidation products in vivo. AB - Extracorporeal elimination of low density lipoprotein (LDL) is frequently used in drug-resistant hypercholesterolemia. LDL-immunoapheresis selectively removes LDL and lipoprotein(a) [Lp(a)] from plasma. Lipid peroxidation is one unwanted side effect, that occurs during extracorporeal plasma treatment. The purpose of this study was to investigate the effect of LDL immunoapheresis on lipid peroxidation. Before and after a single LDL-immunoapheresis treatment, plasma concentrations of lipid hydroperoxides, determined with two different spectophotometric assays, thiobarbituric acid-reacting substances (TBARS), determined spectrophotometrically and malondialdehyde (MDA), determined by an MDA-TBA/HPLC method, were measured in 13 hypercholesterolemic patients. In addition MDA was also determined in the eluate of the apheresis column. Before treatment, plasma cholesterol and LDL cholesterol concentrations were significantly higher in patients than in healthy control subjects, as were the lipid peroxidation products. LDL-immunoapheresis treatment of the patients led to significant decreases in total cholesterol (69+/-8%), LDL-cholesterol (79+/-7%), HDL cholesterol (35+/-17%), triglycerides (38+/-21%), apolipoprotein-B (77+/-6%), apolipoprotein-A1 (25+/-5%) and Lp(a) concentrations (76+/-10%). Changes in plasma lipid peroxide concentrations (17+/-8 nmol/l before vs. 14+/-5 nmol/l after treatment) were not significant, neither were those in TBARS (3. 0+/-2.6 micromol/l vs. 2.3+/-1.3 micromol/l) or MDA concentrations (1.03+/-0.17 micromol/l vs. 1.0+/-0.20 micromol/l). Patients with high baseline values showed a decrease, whereas others did not. MDA was present (0.57+/-0.13 micromol/l) in the eluate of the apheresis column, suggesting that, along with LDL, lipid peroxidation products are also removed. From these results we conclude that a single LDL-immunoapheresis treatment effectively reduces LDL and Lp(a) in the absence of increases in plasma lipid peroxidation products. PMID- 10529455 TI - Elevated beta-N-acetylhexosaminidase activity in focal dystonia fibroblasts. AB - Specific activities of beta-D-hexosaminidase, alpha-D-mannosidase, beta-D galactosidase and beta-D-glucuronidase were determined in fibroblasts of patients with writer's cramp and torticollis. These diseases show degenerative neurological disorders similar to those observed in lysosomal diseases. Hexosaminidase specific activities, determined using 4-methylumbelliferyl-beta-N acetylglucopyranoside and 4-methylumbelliferyl-beta-N-acetylglucopyranoside-6 sulphate as substrates, were significantly higher in the fibroblasts of patients than in controls. No significant differences were observed in the specific activities of the other lysosomal enzymes. The increased hexosaminidase specific activities in torticollis and writer's cramp may be additional markers for these diseases. PMID- 10529456 TI - Binding of Zn(2+) to the plasma protein inter-alpha-inhibitor. AB - Inter-alpha-inhibitor (IalphaI) is a serum protein consisting of a chondroitin sulfate-containing protein of 25 kDa (bikunin) and two other polypeptides of 75 80 kDa (heavy chains 1 and 2). The physiological function of IalphaI is unclear but recent results suggest that it is required for the formation of the extracellular matrix of certain cell types and that it has anti-inflammatory activity. It was previously reported that IalphaI isolated from serum contains bound Zn(2+), but details of this binding are lacking. Using equilibrium dialysis, we have found that when the free Zn(2+) concentration is raised from 0.3 to 50 micromol/L, the number of bound ions increases from 0.1 to 7. The concentration of free Zn(2+) in plasma is in the nanomolar range; our results therefore suggest that inter-alpha-inhibitor does not contain stoichiometric amounts of zinc ions under normal in vivo conditions. PMID- 10529457 TI - The value of analytical assays that are stability-indicating. AB - It is essential to validate an assay with regards to its precision, accuracy, reproducibility, selectivity and robustness. In pharmaceutical research, in addition to these requirements, an assay is often required to be proven beyond doubt to be stability-indicating. A stability-indicating assay is one that can accurately and selectively differentiate an intact drug from its potential decomposition products. An assay that fails to meet this requirement would produce stability data that are inaccurate and misleading. This concept could be applied to other fields of research, such as clinical laboratory chemistry where accurate and reliable clinical data are imperative in the diagnosis and following up of diseases. In this study, we employed an amperometric HPLC assay developed for the measurements of urinary catecholamines and metanephrines to illustrate the validation process of this assay with regards to its stability-indicating capacity. The implications of this study are also discussed. PMID- 10529458 TI - The effect of nitric oxide on ischemia-reperfusion injury in rat liver. AB - A dual role for nitric oxide (NO) in ischemia-reperfusion (I/R) injury is still controversial. This study aims to investigate the role of NO in rat hepatic reperfusion injury. Ischemia was induced by total occlusion of hepatic artery and portal vein for 30 min, then the tissue was reperfused for 30 min. The animals in the L-NAME group (n=10) received N(G)nitro-L-arginine methyl ester (L-NAME) (15 mg/kg) intraperitoneally 60 min before ischemia. The ischemia group (n=10) was given an equal volume of saline solution. The control group comprised eight healthy rats which were not exposed to ischemia or reperfusion. An indicator of hepatic injury, plasma alanine amino transferase (ALT) enzyme activities, were increased in the L-NAME group as compared with the ischemia group (p<0.001). The level of serum nitrite, an index of NO production, and hepatic reduced glutathione (GSH) concentration were lower in the L-NAME group than in the ischemia group (p<0.001, p<0.01, respectively). Hepatic levels of malondialdehyde (MDA) and conjugated dienes (CD) were significantly increased in the L-NAME group as compared to the ischemia group (p<0.05, p<0.001, respectively). Our results confirm that L-NAME, an inhibitor of the enzyme NO synthase, increased the lipid peroxidation and possibly tissue injury, due to the inhibition of cytoprotective effects of NO in a rat hepatic I/R model. PMID- 10529459 TI - Acute intermittent porphyria: biochemical and clinical analysis in the Argentinean population. AB - Acute intermittent porphyria (AIP) is the most common type of hepatic acute porphyria. In this work, we have analyzed the biochemical data of all Argentinean AIP families studied in the Porphyrins and Porphyrias Research Centre (CIPYP). We have shown that: (i) the prevalence for this population is about 1:125,000; (ii) the disease is more frequent in women than in men (7:3); (iii) about 60% are latent carriers; (iv) 15% of patients with symptomatic AIP died during an acute attack; (v) the most important precipitating factors of acute attacks in our population were the ingestion of therapeutic drugs (25%), anesthetics in surgical interventions (25%) and infections (20%); (vi) the initial symptom in Argentinean AIP individuals is severe abdominal pain (100%), and it is often accompanied by constipation (37%), anorexia (37%) and tachycardia (30%); and (vii) the percentage of recurrence of the acute attacks is high (81%). PMID- 10529460 TI - Improved determination of acetylcholinesterase activity in human whole blood. AB - Determination of erythrocyte acetylcholinesterase (AChE) activity is the appropriate tool for the diagnosis of organophosphate exposure and intoxication. The original colorimetric Ellman procedure is disturbed by a high hemoglobin absorption at 412 nm. In our modified method the wavelength was changed to 436 nm. This reduced the indicator absorption to 80% and the hemoglobin absorption to 25%. The signal-to-noise ratio was further enhanced by reduction of pH and substrate concentration, thus making it possible to measure 3% residual activity. AChE activity was determined in whole blood samples in the presence of the selective butyrylcholinesterase inhibitor ethopropazine. Dilution of blood samples (1:100) stops secondary reactions in the presence of inhibitor (organophosphate) and reactivator (oxime). Normalization of the AChE activity to the hemoglobin content, determined as cyanmethemoglobin, prevented dilution errors. This modified approach provides a simple way for sensitive and precise determination of AChE activity in whole blood in the presence of organophosphates even with low-tech equipment. PMID- 10529461 TI - A rapid test for the visual detection of anti-hepatitis C virus antibodies in whole blood. AB - A rapid test for the visual detection of anti-HCV antibodies in whole blood was evaluated in its accuracy when compared with EIA method. The rapid test was performed blind on 50 HCV EIA-positive (adsorbance greater than 3.0) and on 50 HCV EIA-negative samples. Each whole blood sample was 1:20, 1:50 and 1:100 diluted with saline solution for a total of 400 samples. Results showed a sensitivity of 100% when whole blood was tested, 96% when 1:20 diluted blood was tested, 30% when 1:50 diluted blood was tested and 4% when 1:100 diluted blood was tested. The specificity gave also better results and only one false-positive was found in all samples tested. The test took less than 3 min and only a mechanical pipette was required. In conclusion, the HCV Ab rapid test showed a very high accuracy and could be very useful for the detection of HCV-positive subjects in situations where rapid results are required or technical expertise is limited. PMID- 10529463 TI - Creatine kinase 2 mass measurement: methods comparison and study of the matrix effect. AB - Five different commercial immunoassays for the measurement of creatine kinase isoenzyme 2 mass concentration were compared using human plasma samples covering a wide range of creatine kinase 2 concentrations. The immunoassays studied differ in the detection systems, in the specificity of the antibodies and in the calibrators used. Intermethod comparison by regression analysis showed differences in the results of creatine kinase 2 mass concentration. The following ratios were deduced from the obtained equations: Elecsys=1.10xImmulite=1.20xIMx=1.26xACS:180= 1.33 x Stratus. The commutability of different materials prepared by diluting purified human creatine kinase 2 in biological and synthetic matrices was studied using the different immunoassays in comparison with human plasma specimens. Almost all the materials tested were not commutable. PMID- 10529462 TI - Reflex testing II: evaluation of an algorithm for use of cardiac markers in the assessment of emergency department patients with chest pain. AB - A reflex algorithm was developed and evaluated for the use of serum cardiac markers for the diagnosis and rule out of acute myocardial infarction (AMI), and risk stratification of unstable angina patients for those who present to emergency departments (ED) with chest pain. The process begins with testing of total CK and myoglobin at admission. Based on these results, the algorithm determines the need for subsequent testing for the CK-MB isoenzyme and cardiac troponin I (cTnI). The algorithm also directs the need for further blood collection and cardiac marker testing at 4, 8, and 12 h after presentation. A total of eleven stopping points were identified. For some of these stopping points, the algorithm concluded that further blood collections and testing was unnecessary and redundant. The algorithm was retrospectively evaluated on 101 non consecutive chest pain patients who presented to the EDs at three hospitals. For the AMI group (n=34), six of nine possible different stopping points were reached: 64.7% of cases were diagnosed with the first sample at admission, an additional 32.3% after 4 h, and 2.9% at 8 h. The 12-h sample was not necessary for any of the AMI patients. For the non-AMI group (n=67), most reached the stopping point of no cardiac injury or risk. There were five unstable angina patients who had minor myocardial damage on the basis of a marginally increased cTnI. Of these, one patient subsequently suffered AMI, and three others required angioplasty or bypass surgery. Compared to performing four tests on all patient samples, the reflex algorithm would have reduced the number of necessary tests from 442 to 130 (71% reduction) for AMI patients, and 871 to 469 (46% reduction) for non-AMI patients, if prospectively implemented. PMID- 10529464 TI - Comparison of biochemical markers of bone remodelling in the assessment of the effects of alendronate on bone in postmenopausal osteoporosis. AB - The effects of alendronate treatment on biochemical markers of bone remodelling and bone mineral density (BMD) were studied in 30 Caucasian women (postmenopausal for at least 3 years, age 42-76 years, with BMD of the lumbar spine at least 2 S.D. below the mean for mature, premenopausal women). The patients were randomly assigned to receive alendronate (10 mg/day) or placebo for 12 months (double blind). The study was subsequently extended to a second year of open alendronate treatment. The treatment with alendronate resulted in a significant and progressive increase in BMD of the lumbar spine and femoral neck. Under the treatment, the maximal decrease of biochemical markers of bone remodelling (osteocalcin in plasma, bone-specific alkaline phosphatase, N-terminal propeptide of type I procollagen and C-terminal telopeptide of type I collagen in serum, and cross-linked amino-terminal N-telopeptide and total hydroxyproline in urine) was observed at 6 months with no further change during the 2-year period. There were no significant differences in discriminating between patients treated for 1 year with alendronate or placebo using either the percentage change in spine BMD at month 12, or a single measurement of the marker at month 6, or log (percent of baseline at month 6 of value of the marker). In this respect, the power of all the biochemical markers were comparable. The markers are a valuable adjunct to the measurements of BMD, especially in the patients not showing an increase of 3% or more at the lumbar spine BMD after 1 year of treatment. PMID- 10529465 TI - Hypothalamic-pituitary-thyroid axis status of humans during development of ageing process. AB - We have investigated hypothalamic-pituitary-thyroid function in four groups of healthy elderly male humans. Group A (n=18, age range 20-45 years) served as healthy younger controls, group B (n=10, age range 50-60 years), group C (n=15, age range 60-70 years) and group D (n=16, age range 70-85 years) are the subjects of this study. Groups C and D showed significantly lower T3 and thyroid stimulating hormone (TSH), and higher T4 levels with respect to controls. Evidence for TSH circadian modulation was found in group A (control) and group B subjects. The TRH-stimulated TSH peak was reduced among all elderly subjects with respect to controls and appeared to be pronounced with the ageing process. The maximal prolactin response was also inhibited with increasing age. Our study suggest that a resetting of the pituitary threshold for the TSH feed-back suppression along with complex alterations in peripheral thyroid hormone concentrations may, in turn, develop in older people and that appeared to manifest prominently among the oldest population. Additionally, the TSH nocturnal response appeared to be impaired with increasing age indicating an alteration of hypothalamic function. PMID- 10529466 TI - Solid immunoassay and immunonephelemetric methods for serum C reactive protein (CRP) determination: discrepancy in CRP value for a patient presenting a monoclonal immunoglobulin G. PMID- 10529467 TI - Serum cytokeratin 19 fragment levels in non-small cell lung cancer patients according to T factor in the TNM classification. AB - We examined changes in cytokeratin 19 fragment (CYFRA 21-1) levels in relation to the T factor in 64 non-small cell N2M0 lung cancer patients. Although a correlation between the levels and T factor was found (rho=0.627, p<0.0001), there was no difference between the levels in T3 and T4. Serum CYFRA 21-1 levels increased in the order of the following groups: the limited tumor group (T1+T2: n=28), the group with tumor extending to the pleura or chest wall (T3: n=13), and the group with tumor invading into the mediastinum (T4: n=12). The level was lower in the group with malignant pleural effusion (T4: n=11) than in the group with tumor invading into the mediastinum (6.7+/-4.7 ng/ml vs. 12.2+/-8.1 ng/ml, p=0.0046). We suspect that the presence of malignant pleural effusion is not directly related to the three-dimensional expansion of the tumor and this is a reason why CYFRA 21-1 levels in T4 are not higher than those in T3. PMID- 10529468 TI - Genetic polymorphisms of orosomucoid on the Han population in Nanjing of China. AB - Human orosomucoid (ORM) is a major binding protein for various basic drugs. The genetic polymorphisms of ORM could be responsible for interindividual variation in the plasma binding of basic drugs, which might influence their effects or toxicities. The genetic polymorphisms of ORM on the Han population in Nanjing of China were analyzed by isoelectric focusing (IEF) on polyacrylamide gels following by immunoblotting. After desialylation of sera from 220 unrelated Chinese subjects, the band patterns of ORM showed that the polymorphism of the structural locus ORM1 is controlled by three codominant autosomal alleles, ORM1*F1, ORM1*F2 and ORM1*S, which presented five phenotypes, ORM1 F1, ORM1 S, ORM1 F1F2, ORM1 F1S, and ORM1 F2S. The allele frequencies were: ORM1*F1=0.7068, ORM1*F2=0. 0182, ORM1*S=0.2750. The results presented in this paper indicate the ORM1 locus is polymorphic and the ORM2 locus is monomorphic in sera from the Han population in Nanjing of China. PMID- 10529469 TI - Phosphorylation of mitogen-activated protein kinase is altered in neuroectodermal cells overexpressing the human amyloid precursor protein 751 isoform. AB - The aberrant expression or processing of the amyloid precursor protein (APP) is the only known genetic basis for presenile familial Alzheimer's disease, and the molecular connection between APP and tau has been perplexing. Attention has focused on proline-directed serine/threonine kinases as mediating the cytoskeletal modifications of Alzheimer's disease, and we show that overexpression of APP can influence the activation of a candidate kinase, the mitogen-activated protein kinase (MAPK). In murine embryonal carcinoma cells stably transfected with the human 751 isoform of APP, we observed steady-state hyperactivation of p42(MAPK) concomitant with APP overexpression 3 days after neuroectodermal differentiation. In more mature differentiated cells, immunocytochemical analysis revealed enhanced basal somatic and nuclear immunoreactivity for phosphorylated MAPK coupled with an attenuated phosphorylation response to growth factor stimulation. Our results suggest that APP can influence the MAPK signaling pathway in such a way that the absolute and time-dependent activation required for discrimination of the appropriate downstream response are compromised. Such an effect would have important consequences for the functioning of cells coincidentally expressing both proteins, a situation that occurs in neuronal populations vulnerable to Alzheimer's disease pathology. PMID- 10529470 TI - KDEL proteins are found on the surface of NG108-15 cells. AB - Although KDEL proteins are primarily localized to the endoplasmic reticulum (ER), we have employed surface biotinylation method to demonstrate that the KDEL proteins calreticulin (Crt), protein disulfide isomerase (PDI) and the 78-kDa glucose regulated protein (GRP78) are found on the surface of the NG108-15 cell line. In contrast, the 94-kDa glucose regulated protein (GRP94), another KDEL protein, is not found on the cell surface. Calnexin (Cnx), a type-1 integral transmembrane ER protein which is partially homologous to Crt but lacks the KDEL sequence, is not detected on the cell surface either. While only small amounts of the total GRP78, PDI and Crt molecules exist on the cell surface at steady state, a significant fraction of the newly synthesized molecules are transported to the cell surface and transport of these proteins is inhibited by treatment with brefeldin A. The surface GRP78 contains the KDEL sequence. On the cell surface, GRP78, PDI and Crt associate with other proteins and form complexes of different sizes. Surface Crt is found to be essential for the neurite formation when NG108 15 cells are induced to differentiate using dibutyryl cAMP. PMID- 10529471 TI - Attenuation of temporary focal cerebral ischemic injury in the mouse following transfection with interleukin-1 receptor antagonist. AB - The proinflammatory cytokine interleukin-1 beta (IL-1beta) is thought to play an important role in the stimulation of the inflammatory response following ischemia and reperfusion. This study investigated the inflammatory effect of IL-1beta during transient focal cerebral ischemia and reperfusion in the mouse transduced with the interleukin-1 receptor antagonist (IL-1ra) gene. An adenoviral vector encoding, either the human IL-1ra gene (AdRSVIL-1ra) or the LacZ gene (AdRSVlacZ) or normal saline, were injected into the right lateral ventricles of adult CD-1 mice (n=96). Five days later, the mice received 1 h temporary middle cerebral artery occlusion (tMACAO) followed by 23 h reperfusion. Cerebral blood flow (CBF), infarct volume, blood-brain barrier (BBB) permeability, and the number of intracellular adhesion molecule-1 positive vessels were measured to determine the effect of IL-1beta during postischemic reperfusion. Infarct volume in the AdRSVIL 1ra-transduced mice was markedly reduced compared to the AdRSVlacZ-transduced and saline-injected mice (36.0+/-5.3 mm(3) vs. 60.0+/-6.2 mm(3), 69. 5+/-6.3 mm(3), after 23 h of reperfusion, n=6-8 per group, p<0.05). BBB disruption and intracellular adhesion molecule-1 expression (135+/-23 vs. 311+/-40 and 357+/-51, n=6-8 per group, p<0.05) in the AdRSVIL-1ra-transduced mice were also less than that of the AdRSVlacZ-transduced and saline-injected mice. Our studies demonstrated that overexpression of IL-1ra in the mouse brain can downregulate intracellular adhesion molecule-1 expression both in the cortex and basal ganglia, which suggests that IL-1beta may play an important role in the activation of the inflammatory response during focal cerebral ischemia by promoting leukocyte adhesion to endothelial cells. The decrease of BBB disruption in AdRSVIL-1ra-transduced mice suggests that the endothelial cells may be a target for IL-1beta during postischemic reperfusion. PMID- 10529472 TI - Modulation of carbachol-stimulated AP-1 DNA binding activity by therapeutic agents for bipolar disorder in human neuroblastoma SH-SY5Y cells. AB - Lithium, carbamazepine and sodium valproate are mood stabilizers used in the treatment of bipolar disorder, and although their mechanisms of action remain unknown, signal transduction systems and the associated modulation of gene expression may constitute significant actions. We examined if acute or chronic treatments with these agents modulated the activation of the AP-1 transcription factor or the increased intracellular calcium levels in human neuroblastoma SH SY5Y cells caused by stimulation with carbachol. AP-1 activation stimulated by carbachol was reduced by pretreatment for 1 h, 24 h or 7 days with 1 mM lithium by 15%, 37%, and 60%, respectively, and with 0.05 mM carbamazepine by 3%, 21%, and 46%, respectively, but not by pretreatment with 0.5 mM sodium valproate. AP-1 DNA binding activity stimulated by carbachol or by phorbol ester-induced activation of protein kinase C was inhibited by the protein kinase C inhibitor Ro31-8220, but phorbol ester-stimulated AP-1 activation was unaltered by 7-day pretreatments with lithium or carbamazepine. Activation of AP-1 by carbachol was dependent on calcium, as it was inhibited by treatment with the extracellular calcium chelator EGTA, the intracellular calcium chelator BAPTA-AM, and the calcium/calmodulin kinase II inhibitor KN62. Pretreatment for 7 days with lithium or carbamazepine had no significant effect on carbachol-stimulated increases in intracellular calcium levels, but reduced the stimulation of AP-1 by the calcium ionophore ionomycin by 30% to 40%. Thus, chronic treatment with the antibipolar agents lithium and carbamazepine attenuates carbachol-stimulated AP-1 DNA binding activity, and these agents preferentially inhibit signaling cascades activated by the calcium rather than the protein kinase C arm of the phosphoinositide signaling pathway. PMID- 10529473 TI - Identification of mRNAs localizing in the postsynaptic region. AB - Local protein synthesis using mRNAs readily distributed in the dendrites is believed to play an important role in maintaining the already expressed synaptic plasticity. To find proteins translated in the postsynaptic region, such as neuronal dendrites, we tried to identify the mRNAs associated with the postsynaptic density (PSD) fraction prepared from a rat's forebrain. The PSD associated mRNAs were amplified by reverse transcriptase-based polymerase chain reaction (RT-PCR), separated by polyacrylamide gel electrophoresis, and sequenced. The database search revealed, among 130 mRNAs sequenced, 17 known and 108 unknown sequences, while five mRNAs were too short for the search. Of the mRNAs with unknown sequences, we selected 33 genes with a length longer than 150 bases, performed in situ hybridization, and found that at least 12 mRNA types were localized in the dendrites. These results suggest that a large number of mRNAs localize around the postsynaptic area of the neuronal cells in the central nervous system. In addition, our method proved efficient in identifying collectively the mRNAs localizing in the dendrites. PMID- 10529474 TI - Dual mechanism of Fas-induced cell death in neuroglioma cells: a role for reactive oxygen species. AB - ApoI/Fas belongs to the tumor necrosis factor receptor (TNFR) superfamily and mediates cell death in various cell types. A dual mode of Fas-triggered cell death has been reported depending on cell types used in the experiments. The present study was carried out to test the possible role of reactive oxygen species in this dual mechanism in neuroglioma cells. Anti-Fas antibody caused dose-dependent and time-dependent increase in cell death measured by lactate dehydrogenase (LDH) release in control neuroglioma cells and in cells that were transfected with catalase cDNA. However, cells transfected with copper/zinc superoxide dismutase (Cu/ZnSOD) cDNA showed marked attenuation of Fas-induced LDH release. Moreover, flow cytometry and confocal microscopy revealed that Fas induced cell death in control cells occur mostly through an apoptotic process. This process was also completely abrogated in cells overexpressing catalase or copper/zinc superoxide dismutase (Cu/ZnSOD). Further experiments revealed that Fas-induced cell death was associated with increased formation of superoxide anions in control neuroglioma cells and in cells overexpressing catalase. These increases were significantly suppressed by Cu/ZnSOD overexpression. These data indicate that Fas-mediated cell death in neuroglioma cells occur, in part, through the production of reactive oxygen species (ROS). These observations also suggest that Fas-induced cell death in these cells occur through apoptosis and necrosis. Thus overexpression of Cu/ZnSOD caused the suppression of both types of Fas-induced cell death whereas catalase prevented apoptotic but not necrotic cell death. These observations are discussed in terms of their support for a role for both peroxides and superoxide radicals in Fas-induced cell death. PMID- 10529476 TI - Expression of mt(1) melatonin receptor subtype mRNA in the entrained rat suprachiasmatic nucleus: a quantitative RT-PCR study across the diurnal cycle. AB - Melatonin acts on specific receptors in the suprachiasmatic nuclei (SCN) to phase dependently regulate the phase of the circadian clock. How the gating of melatonin's effect is restricted to particular times of day is not known, but may be related to temporal differences in receptor availability. In the present study, we used a competitive reverse transcription-polymerase chain reaction (RT PCR) method to determine if the expression of mt(1) melatonin receptor subtype mRNA in rat SCN varied across the 12:12 light-dark (LD) cycle. Measurement of core body temperature using radiotelemetry confirmed that the male Wistar rats used exhibited a robust diurnal rhythm. mt(1) receptor mRNA was readily detected in reduced SCN slices at all times of day. However, there was no significant variation in the amount of mt(1) mRNA with time of day. Expression of MT(2) melatonin receptor subtype mRNA in reduced SCN slices was confirmed by nested PCR. These results indicate that changes in the level of mt(1) mRNA do not underlie the diurnal and/or circadian variation in the response of the SCN circadian clock to the phase-resetting effects of melatonin. PMID- 10529475 TI - Effects of chronic alcohol consumption on the expression of different NR1 splice variants in the brain of AA and ANA lines of rats. AB - Following chronic alcohol treatment alterations in N-methyl-D-aspartate receptor subunit 1 and 2 (NR1 and NR2), mRNA and protein levels have been reported. The NR1 gene undergoes alternative RNA splicing, resulting in eight splice variants, which were shown to differ in their sensitivity to alcohol. Here, we studied mRNA and protein levels of NR1 splice variants in alcohol-preferring (AA) and alcohol nonpreferring (ANA) rat lines under basal conditions (alcohol-naive), and following chronic alcohol consumption. mRNA levels of three NR1 splice variants (NR1-1, NR1-2, NR1-4), and the protein levels of NR1 (NR1-1/NR1-2), and of NR1 alternative C-terminus (NR1-3/NR1-4) were determined in the hippocampus and nucleus accumbens by competitive RT-PCR and Western blot analysis, respectively. No significant differences in NR1 mRNA, or protein levels were found in the nucleus accumbens between the two rat lines under basal conditions, or following chronic alcohol consumption. In the hippocampus of alcohol-naive rats, the NR1-4 mRNA content was significantly higher in ANA compared to AA rats, however, no significant difference could be detected at the protein level. Following chronic alcohol consumption, the protein level of the NR1 alternative C-terminus (NR1 3/NR1-4) was significantly higher in AA rats compared to the corresponding control. Taken together, these results suggest: (i) brain site-specific alterations in NMDA receptor subunit composition occur following chronic alcohol consumption. (ii) In the hippocampus, NR1 splice variant mRNA levels differ between AA and ANA rats. (iii) The mRNA levels and protein levels of NR1 splice variants are differentially affected by chronic alcohol consumption. PMID- 10529477 TI - Lactoferrin is synthesized by mouse brain tissue and its expression is enhanced after MPTP treatment. AB - The presence of iron in brain tissue in increased concentrations in Parkinson's disease cases, where it might be responsible for oxidative stress, and the parallel observation that the iron transporter lactoferrin (Lf) was present in increased amounts in surviving neurons, led us to study the synthesis of Lf in a mouse model of Parkinson's disease. In this context, the origin and expression of brain Lf in normal, aged and MPTP (1-methyl-4-phenyl-1, 2,3,6-tetrahydropyridine) treated mice were investigated. Lf immunostaining was observed mainly on microvessels in the cerebral cortex of the adult mice and to a greater extent in older mice. Lf immunoreactivity was also present in the hippocampus only in the aged mouse brains, associated with structures which seemed to be pyramidal neurons and fibers. After RT-PCR (polymerase chain reaction), Lf transcripts were found in mouse brain tissue whatever the age of the animals studied but the level of their expression was very low. No up-regulation of Lf was detectable during aging. Lf distribution and expression in the MPTP-induced Parkinsonian mouse model were also investigated. A marked depletion of dopamine (DA) occurred in the high dose MPTP-treated mice. The level of Lf expression was found to be markedly increased in the same animals and this up-regulation occurred on the first day after MPTP administration. When the brain was stressed by the neurotoxin MPTP, Lf expression increased in line with antioxidant enzymes such as catalase and gamma glutamylcysteine synthetase, which may permit the protection of brain tissue from oxidative damage induced by the drug. PMID- 10529478 TI - Mutation of human mu opioid receptor extracellular "disulfide cysteine" residues alters ligand binding but does not prevent receptor targeting to the cell plasma membrane. AB - The mu opioid receptor, a primary site of action in the brain for opioid neuropeptides and opiate drugs of abuse, is a member of the seven transmembrane, G protein-coupled receptor (GPCR) superfamily. Two cysteine residues, one in each of the first two of three extracellular loops (ECLs), are highly conserved among GPCRs, and there is direct or circumstantial evidence that the residues form a disulfide bond in many of these receptors. Such a bond would dramatically govern the topology of the ECLs, and possibly affect the position of the membrane spanning domains. Recent findings from several laboratories indicate the importance of the ECLs for opioid ligand selectivity. These conserved cysteine residues in the mu opioid receptor were studied using site-directed mutagenesis. Little or no specific binding of radiolabled opiate alkaloid or opioid peptide agonists or antagonists was observed for receptors mutated at either "disulfide cysteine" residue. Each mutant mu opioid receptor was expressed in both transiently- and stably-transfected cells, in some cases at levels comparable to the wild type receptor. The two point mutants possessing serine-for-cysteine substitutions were also observed to successfully reach the cell plasma membrane, as evidenced by electron microscopy. Consistent with related work with other GPCRs, the mu opioid receptor apparently also employs the extracellular disulfide bond. This information now permits accurate molecular modeling of extracellular aspects of the receptor, including plausible scenarios of mu receptor docking of opioid ligands known to require specific extracellular loop features for high affinity binding. PMID- 10529479 TI - Enhanced BK-induced calcium responsiveness in PC12 cells expressing the C100 fragment of the amyloid precursor protein. AB - Several lines of evidence have implicated the amyloid precursor protein (APP) and its metabolic products as key players in Alzheimer's disease (AD) pathophysiology. The approximately 100 amino acid C-terminal fragment (C100) of APP has been shown to accumulate intracellularly in neurons expressing familial AD (FAD) mutants of APP and to cause neurodegeneration when expressed in transfected neuronal cells. Transgenic animals expressing this fragment in the brain also exhibit some neuropathological and behavioral AD-like deficits. Here, we present evidence that PC12 cells expressing the C100 fragment either via stable transfections or herpes simplex virus-mediated infections show alterations in calcium handling that are similar to those previously shown in fibroblasts from AD patients. This alteration in calcium homeostasis may contribute to the deleterious effects of C100 in PC12 cells. Our data also lend support for a pathophysiological role for C100 since it induces an alteration thought to play an important role in AD pathology. PMID- 10529480 TI - bcl-2 prolongs neuronal survival during hypoxia-induced apoptosis. AB - In vivo models of hypoxic-ischemic brain injury have shown altered expression of a number of genes that are important in regulating neuronal survival. However, it is not clear as to whether hypoxia alone can alter the expression of genes regulating neuronal survival. We hypothesized that (1) hypoxia alone alters the expression of bcl-2 in neurons, (2) the severity and duration of hypoxia influence bcl-2 expression, and (3) the alteration of bcl-2 expression has an important role in regulating neuronal survival during hypoxia. Embryonic rat neocortical neurons cultured for 7-10 days were exposed to 0.1, 1, or 3% oxygen for various durations and were removed for analyses at 24-h intervals. Under all hypoxic conditions, neurons exhibited morphologic changes, as assessed by electron microscopy and Annexin V staining, consistent with apoptosis. Immunoblot and immunofluorescence analyses revealed an increase in neuronal bcl-2 protein during hypoxic exposure. Quantitative immunofluorescence analyses of bcl-2 immunostained neurons indicated that expression of bcl-2 was altered by the duration and severity of hypoxia. Attenuation of bcl-2 expression by antisense oligonucleotides decreased the proportion of surviving neurons by approximately 50% after 48 h of exposure to 0.1% oxygen. We conclude that observed increase in bcl-2, in part, plays an important role in neuronal survival during exposure to hypoxia. PMID- 10529481 TI - Autoradiographic evidence that intrastriatal administration of adenosine A(1) receptor antisense oligodeoxynucleotide decreases adenosine A(1) receptors in the rat striatum and cortex. AB - In this study, the effect of a phosphorothioated A(1) adenosine receptor antisense oligodeoxynucleotide on A(1) receptor density and mRNA in the striatum and cortex of rats was determined. Receptor autoradiography and in situ hybridization revealed a reduction in striatal and cortical A(1) receptor density and cortical A(1) receptor mRNA, respectively, in antisense-treated brains but not in those treated with a mismatch oligonucleotide. There was no change in A(2) receptor binding. These data imply that the corticostriatal pathway synthesizes A(1) receptors and transports them to its terminals. PMID- 10529482 TI - Iron-independent neuronal expression of transferrin receptor mRNA in the rat. AB - Neuronal transferrin receptor protein expression is highly upregulated widely in CNS following iron deficiency. Using the medial habenular nucleus as a model of neuronal transferrin receptor mRNA expression, the present study examined 17-day old rats subjected to variations in dietary iron. Changing the iron availability resulted in alterations in plasma and cerebrospinal fluid (CSF) levels of transferrin and iron. The iron-binding capacity of transferrin in CSF was exceeded in normal and iron-overloaded rats. In spite of a lowering of the concentration of brain iron by approximately 22% in iron-deficient rats, neuronal transferrin receptor mRNA was not affected when measured by quantitative densitometry. Brain iron and neuronal transferrin receptor mRNA expression was unaltered in iron overloaded rats. The absence of a rise in transferrin receptor mRNA during iron deficiency suggests that neuronal transferrin receptor mRNA expression is regulated by another mechanism than the post-transcriptional regulation mechanism, which has been attributed to cells of non-neural tissue. PMID- 10529484 TI - Integrative physiology of coronary microcirculation. AB - Coronary microvessels play a crucial role for mechanoenergetic interaction between blood flow and myocardial function, which is not uniform transmurally. Thus, highly organized vascular regulations are required for matching local blood flow with myocardial energy requirement. Recently, new technologies to investigate in vivo coronary microcirculation with new knowledge of the signaling molecules for vascular regulation have revolutionized our abilities to understand the integrative regulation of coronary microcirculation. In this review, the mechanical aspects of the interaction between coronary blood flow and myocardium, coronary arte-rial tree and its roles in myocardial blood flow regulation, hierarchical and dynamic control of coronary flow, capillary network and function, function of venous drainage system, and molecular and cellular aspects of integrative coronary blood flow regulation are discussed, focusing on their integrational roles in maintaining coronary microvascular function and cell signaling. PMID- 10529483 TI - Roles of ion channels in carotid body chemotransmission of acute hypoxia. AB - In this review, we have highlighted the roles of ion channels in carotid body chemotransmission of acute hypoxia. With the application of new technologies, significant breakthroughs have been made in the last decade. The discovery of oxygen-sensitive K(+) channels in rabbit glomus cells has generated the membrane model of hypoxic chemotransmission: the inhibition of oxygen-sensitive K(+) channels by hypoxia initiates the depolarization of glomus cells and increases the firing frequency of glomus cells. The depolarization of glomus cells activates voltage-gated Ca(2+) channels, elevating intracellular Ca(2+) which triggers the release of neurotransmitters. The correlation of these events in rabbit glomus cells has been shown. However, a large corpus of data indicates that various mechanisms may be involved in different species. In rats, Ca(2+) activated K(+) channels are inhibited by hypoxia. The role of this inhibition on rat glomus cell function is controversial, and the contribution of leak-type K(+) channels to rat glomus cell depolarization has recently been proposed. On the other hand, in cats, nicotinic ACh receptors (ligand-gated cation channels) may play a key role in initiating the depolarization of glomus cells and increasing the cytosolic Ca(2+) of glomus cells in response to hypoxia. Hypoxic inhibition of oxygen-sensitive K(+) channels would participate to further depolarize cat glomus cells. Additionally, the activity of Cl(-) channels and the modulation of ion channels by neurotransmitters may influence the excitability of glomus cells. For generating action potentials in chemoreceptor afferent nerves, nicotinic ACh receptors appear to be involved in cats and rats. PMID- 10529485 TI - Two types of external Cl(-)-dependent Cl(-) channels and one type of stretch receptor cation channel contribute to the formation of isotonic blastocoel fluid in early medaka fish embryo. AB - Ionic channels in blastoderm cells dissociated from medaka fish embryos at the late blastula stage were studied by the patch-clamp technique in whole-cell, inside-out and cell-attached patch configurations. These cells were mechanically dissociated without using proteo-lytic enzymes. I have reported previously an external anion-dependent type I Cl(-) channel, which was observed at the early blastula stage, and its voltage dependency shifted toward the positive direction with a reduction in [Cl(-)](o). The type I Cl(-) channel was observed in about half of the blastoderm cells at the late blastula stage. I also observed another external anion-dependent Cl(-) channel (type II) whose voltage dependency shifted toward the negative direction with a reduction in [Cl(-)](o) and a cation selective stretchactivated channel. The$coexistence of the type II Cl(-) channel and stretch-activated cation channel would produce the efflux of both cations and anions in response to low ionic strength fluid. Further, the initial rise of blastocoel [Cl(-)] by this mechanism would trigger positive feedback between the type I Cl(-) channel conductance and blastocoel [Cl(-)]. At the early blastula stage, the blastocoel cavity fluid has a low ionic strength similar to that of pond water. I proposed that the cooperation of three types of ion channels at the late blastula stage generates isotonic blastocoel cavity fluid. PMID- 10529486 TI - Increased cardiovascular and metabolic tolerance to acute hypoxia in the rat with increased hemoglobin-O(2) affinity induced by Na-cyanate treatment. AB - Cyanate derivatives such as NaOCN have been known to increase the hypoxia tolerance of animals by increasing the affinity of hemoglobin (Hb) to O(2). To clarify the mechanism of this increase in hypoxia tolerance, we examined changes in metabolic rate and cardiovascular parameters during a hypoxia test in halothane-anesthetized, NaOCN-treated and spontaneously breathing rats (50 mg/kg/d S.C., 10 d). Control animals received saline. The capillary density in the skeletal muscle (sternocleidomastoid muscle), cardiac papillary muscle and medulla oblongata was also examined histologically. The Hb-O(2) affinity index, P(50), decreased from 38 (control rat) to 24 mmHg in NaOCN-treated rats. During hyperoxic gas breathing, the rat treated with NaOCN showed a significantly lower metabolic rate (V(.)O(2), V(.)CO(2)), higher cardiac stroke volume, slower heart rate, lower PvO(2), and lower O(2) extraction ratio than those in control rats. The NaOCN-treated rats exhibited well-maintained arterial blood pressure and a larger cardiac output response to reduction in FIO(2) to 0.10-0.08. The increase in O(2) extraction ratio with reduction in FIO(2) was larger in NaOCN-treated than in control rats. The circulatory and metabolic depressions at FIO(2) 0.05 were effectively attenuated in NaOCN-treated rats. The capillary density of the cardiac muscle and medulla oblongata but not the skeletal muscle was significantly higher in NaOCN-treated rats than in control rats. The greater hypoxia tolerance in NaOCN-treated rats is ascribed to the combined effects of left shift of Hb-O(2) dissociation curve, lower basal V(. )O(2), higher capillary density in the heart, and brain, and other adaptive mechanisms induced probably by prolonged tissue hypoxia. PMID- 10529487 TI - Inhibition of the endothelium-dependent relaxation by 18beta-glycyrrhetinic acid in the guinea-pig aorta. AB - The effect of 18beta-glycyrrhetinic acid (GA), an agent which interferes with gap junction conductivity, on endothelium-dependent relaxation produced by substance P was investigated in isolated aortic rings of the guinea-pig. In nor-adrenaline (NA)-contracted aortic rings, substance P (10(-7) M) induced an endothelium dependent, transient relaxation. The relaxation was only slightly reduced by the co-application of nitroarginine and diclofenac. When GA (2x10(-5) M) was applied first, it slightly reduced substance P-induced relaxation, and a subsequent co application of nitroarginine and diclofenac strongly reduced the relaxation. In aortic rings contracted with high-K solution ([K(+)](o) = 29.4 mM), substance P induced relaxation was reduced by the simultaneous application of GA, nitroarginine and diclofenac, but not by GA alone. In endothelium-denuded aortic rings, GA reduced the threshold concentration of NA required to produce contractions and increased the amplitude of NA-induced contractions. GA increased the amplitude of contraction produced by small increases of [K(+)](o) (<30 mM) but reduced those produced by higher concentrations of [K(+)](o) (>54 mM). In NA contracted aortic rings, Y-26763, a K(+)-channel opener, could relax muscles with reduced amplitude in the presence of GA. It is concluded that in guinea-pig aortic rings, GA inhibits mainly the EDHF-induced components of endothelium dependent relaxation. GA also modulated contractions produced by NA or high-K solutions. The possible effects of inhibition of gap junctions by GA on endothelium-dependent relaxation were discussed. PMID- 10529488 TI - Self-biofeedback control of heart rate during exercise. AB - To improve cardiac adjustment to exercise, we developed a new self-biofeedback heart rate (HR) controller. Using this device, we analyzed time courses of HR, running speed (RS), stride length (ST) and pitch of gait (PI) in response to various preset HR levels in 7 normal human subjects. When HR was preset at 80 bpm, HR increased rapidly in response to exercise and exceeded the preset level at 12. 1 s with overshoot. At the preset HRs of 100, 120, 140 and 160 bpm, the HRs increased to each preset level at 39.2, 64.5, 58.5 and 83.0 s after the onset of exercise, respectively, and the HRs were adjusted with a range of +/- 4%. For all preset HRs, RS, ST and PI increased more rapidly than the HR and reached the maximum values within 30 s. During exercise, RS, ST and PI remained constant within 1.5-5.5 min. HR, RS, ST and PI increased in proportion to the preset HR. The increases in PI against HR (DeltaPI/DeltaHR) decreased with the higher HR level, and at HRs of 160-170 bpm, HR and PI showed identical rhythm. The increases in RS were produced by 18-59% increases in PI and by 12-44% increases in ST. We concluded that, using our newly developed self-biofeedback HR control system, we could control HR to a given preset value by a change in RS due to PI and ST. PMID- 10529489 TI - The neural mechanism of rectal motility response induced by the epicardial application of lactic acid. AB - The epicardial application of lactic acid induced a biphasic rectal motility response in lightly anaesthetised, open-chested and artificially ventilated cats. This rectal biphasic response is reflexogenic in nature as epicardial lignocaine abolished such response. This rectal biphasic response is abolished by cardiac sympathectomy and reprecipitated by left inferior cardiac afferent nerve stimulation. Such response is also abolished by sacral ventral rhizotomy and reproduced by stimulation of the peripheral cut end of split sacral ventral roots. This indicates that the afferent and efferent pathways for such reflex are lying in the cardiac sympathetic and sacral pelvic nerves, respectively. The higher centers involved for such reflex are lying above the mid-collicular level of the brain as decerebration at the mid-collicular level completely abolished such type of rectal response. Furthermore, the relaxation phase and contraction phase of such rectal response are mediated through nitric oxide release and cholinergic neurones, respectively, as NG-nitro-L-arginine and atropine abolished relaxation and contraction phase of the rectal response, respectively. PMID- 10529490 TI - CBF change evoked by somatosensory activation measured by laser-Doppler flowmetry: independent evaluation of RBC velocity and RBC concentration. AB - The purpose of this study was to examine the timing and magnitude of cerebral blood flow (CBF) responses to neuronal activation. We measured the changes in local CBF (LCBF), red blood cell (RBC) velocity and RBC concentration by laser Doppler flowmetry (LDF) as well as field potential recordings during activation of the somatosensory cortex of the rat in response to electrical stimulation of the hind paw. Electrical stimuli, 0.1 ms pulses of 1-1.5 mA for 5 s, were applied at 0.2, 0.5, 5, 10 and 50 Hz under alpha-chloralose anesthesia. LCBF showed the maximum increase at 5 Hz, and rose approximately 0.5 s after the onset of stimulation regardless of the frequency. The maximum frequency of the field potentials was also obtained at 5 Hz. During activation of the somatosensory cortex, the onset of rise in RBC concentration did not precede that of RBC velocity, and the peak RBC concentration was noted earlier than that of both LCBF and RBC velocity, suggesting that both arteriolar diameter and active changes in the capillary contributed to the LCBF response. PMID- 10529491 TI - Thermoregulation as a switchboard of autonomic nervous and endocrine control. AB - Analyzing the multiple effectors of autonomic temperature regulation has, sometimes unexpectedly, provided insights into the characteristics according to which the non-thermoregulatory functions of these effectors are controlled. This article reviews the sympathetic control of cardiovascular and immune functions, hormonal control of energy balance and neurohormonal control of salt and fluid balance inasmuch as they are challenged by competing demands of thermoregulatory requirements. These interactions are taken as examples for the analytical power of the experimental conception to challenge non-thermoregulatory control systems with thermoregulatory activation, and vice versa. Animal models carrying spontaneous or intentionally produced gene defects and molecular and histochemical techniques of gene identification and neuronal tracing are becoming increasingly important. They are applied, in connection with physiological studies exploiting mutual interactions of autonomic control systems, with the aim to elucidate the cytoarchitecture of neuronal circuits by which specific autonomic regulatory activities are controlled. PMID- 10529492 TI - Molecular aspects of the excitation-contraction coupling in skeletal muscle. AB - Extremely rapid and massive release of Ca(2+) from the sarcoplasmic reticulum upon depolarization of the T-tubule regulates the contraction of skeletal muscle. The dihydropyridine receptor (voltage sensor) on the T-tubule faces the ryanodine receptor (Ca(2+) release channel) on the sarcoplasmic reticulum at the triadic junction and their intermolecular interaction is responsible for the swift release of Ca(2+) from the sarcoplasmic reticulum. We are now beginning to gain insight into the molecular structures that are important for the coupling mechanism. PMID- 10529494 TI - In vitro thermogenesis and phospholipid fatty acid composition of brown adipose tissue in fasted and refed rats. AB - Membrane phospholipids are known for their role in the regulation of membrane structures and functions. Membrane phospholipid fatty acid docosahexaenoic acid (DHA) has been recently indicated to be important for the regulation of cellular activities, including metabolic regulation. Our previous studies have indicated the involvement of DHA in the regulation of brown adipose tissue (BAT) thermogenesis. The objective of the present study is to examine the changes in BAT phospholipid fatty acid composition including DHA and thermogenic activity in fasted and refed rats. Phospholipid content per microgram DNA was decreased in rats fasted for 72 h and it was not restored to the control level by refeeding for 72 h. Phospholipid fatty acid composition of BAT, as expressed by mol%, was modified in the fasted rats. Most notably, DHA, which constituted about 89% of the n-3 polyunsaturated fatty acids, was decreased concomitant with the increase in the n-6 polyunsaturated fatty acid arachidonic acid. The monounsaturated to saturated fatty acid ratio, which is an index of Delta(9)-desaturase activity and membrane fluidity, was decreased. Thermogenesis, as measured by the in vitro oxygen consumption of BAT, was suppressed in the fasted rats. All of the above changes were restored to normal levels after refeeding the fasted rats for 72 h. In vitro oxygen consumption correlated with the level of DHA and monounsaturated to saturated fatty acid ratio. These results indicate that the modification of phospholipid fatty acid composition, especially the modification of n-3 polyunsaturated fatty acid DHA, and membrane fluidity are related to BAT thermoregulation in fasted and refed rats. PMID- 10529493 TI - Modulation of carbachol-induced Cl(-) currents and fluid secretion by isoproterenol in rat submandibular acinar cells. AB - The effects of muscarinic and beta-adrenergic agonists on Cl(-) currents in acinar cells were investigated to clarify their role in the regulation of fluid secretion in rat perfused submandibular glands. Additions of isoproterenol (IPR) at 10(-8) to 10(-6) M and 8-(4-chlorophenylthio)-cyclic AMP (CPT-cAMP) at 10(-3) M to the perfusate suppressed carbachol (CCh; 10(-6) M)-induced fluid secretion. IPR and CPT-cAMP also diminished CCh-induced oscillatory Cl(-) current and increased CCh-stimulated non-oscillatory Cl(-) current. Propranolol blocked the effect of IPR on fluid secretion. IPR did not modulate the CCh-induced increase in intracellular concentration of calcium ions and intracellular pH in isolated cells. Propranolol blocked IPR-induced changes in Cl(- )currents, while propranolol itself increased CCh-induced K(+) current and reduced CCh-induced oscillatory Cl(-) current. Increasing external osmolarity with 50 mM sucrose abolished IPR-enhanced non-oscillatory Cl(-) current. Neither CCh-induced oscillatory Cl(-) current nor IPR-induced suppression of the oscillatory Cl(-) current was influenced by the hypertonicity. Perfusion of the gland with the hypertonic solution did not affect the IPR-induced suppression of fluid secretion. These results suggest that IPR induces the suppression of CCh-induced oscillatory Cl(-) current and potentiation of the non-oscillatory Cl(-) current via an increase in cyclic AMP level, and that suppression of the oscillatory Cl( ) current by IPR may contribute to the inhibition of fluid secretion from submandibular glands. PMID- 10529495 TI - Simulation of spike-burst generation and Ca(2+) oscillation in pancreatic beta cells. AB - Based on the experimental evidence that Na(+)-Ca(2+) exchange participates in the regulation of intracellular Ca(2+) concentration in pancreatic beta-cells, we construct a mathematical model for the cyclic spike-bursts and oscillations of intracellular Ca(2+) concentration. In our model, an increase in ATP concentration by the stimulation of glucose metabolism leads to the closure of ATP-sensitive K(+) channels (K(ATP) channels) and gradual depolarization to the threshold of voltage-gated Ca(2+) channels. Spikes are generated by the alternate activation of voltage-gated Ca(2+) and K(+) channels, causing Ca(2+) entry. The accumulated Ca(2+) ions are extruded by Na(+)-Ca(2+) exchange and Ca(2+) active transport. An increase in Na(+) influx through Na(+)-Ca(2+) exchangers results in a rise in intracellular Na(+) concentration and the activation of Na(+)-K(+) active transport. The consumption of ATP during the process of Ca(2+) extrusion leads to the opening of K(ATP) channels and repolarization. The present model could reproduce the main experimental features of the spike-burst activity and Ca(2+) oscillations following changes in the extracellular glucose concentration. As the rate of ATP production increases, the spike-burst pattern changes from bursts with long silent phases to continuous spiking. Changes in the pattern of electrical activity produced by the alteration of extracellular Na(+) and K(+) concentrations and the addition of ouabain could be reproduced in the present model. PMID- 10529496 TI - Prostaglandin E(2)-activated housekeeping Cl(-) channels in the basolateral membrane of rat gastric parietal cells. AB - Cl(-) channels in the basolateral membrane of parietal cells within isolated rat gastric glands were studied by the whole-cell patch-clamp technique. The membrane potential (E(m)) of non-stimulated parietal cells changed as a function of the basolateral extracellular Cl(-) concentration (46 mV per decade), but E(m) did not change significantly as a function of the K(+) concentration. The extracellular addition of prostaglandin E(2) (PGE(2); 10 microM) increased the whole-cell Cl(-) current. A bifunctional prostaglandin EP3 agonist/EP1 antagonist, 5(Z)-7-[(1S, 2S,3S,5R)-3-(trans-beta-styren)sulfonamido-6,6 dimethylbi cyclo-(3.1. 1)hept-2-yl]-5-heptenoic acid (ONO-NT-012; 10 microM), also increased the Cl(-) current. Basal Cl(-) currents and the PGE(2)- and ONO-NT 012-increased Cl(-) currents were voltage-independent and inhibited by a Cl(-) channel blocker, 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB), at 500 microM. The single Cl(-) channel conductance was estimated to be 0.29 picosiemens (pS) by variance noise analysis. Both PGE(2) and ONO-NT-012 increased intracellular free Ca(2+) concentration in the fura-2-loaded parietal cell transiently. The present study has shown that housekeeping sub-pS Cl(-) channels are present in the basolateral membrane of rat parietal cell, and that the channels are regulated positively by PGE(2) via the EP3 receptor. PMID- 10529497 TI - Effect of renal denervation on the compensatory renal growth following nephrectomy in the cat. AB - The purpose of this study was to clarify the effect of denervation on the mass of the remaining kidney with or without unilateral nephrectomy using adult cats. The animals were divided into 4 groups: (1) control group, the weights of the right and left kidneys were measured intact in 5 cats; (2) nephrectomy group (Nx, n = 5 cats), the right kidney was removed and the left kidney was weighed 3-5 d after nephrectomy; (3) nephrectomy and denervation group (Nx+Dx, n = 7 cats), the left kidney was weighed on the 7th day after surgery in which the left kidney was denervated and the right kidney was removed; and (4) denervation group (Dx+Dx, n = 5 cats), both kidneys were weighed on the 7th day after denervation of the kidneys. In the control group, the left and right kidney weights per body weight (LKW and RKW) were the same (LKW, 0.74 +/- 0.06%; RKW, 0. 74 +/- 0.07%). In the Nx group, LKW increased to 0.90 +/- 0.03% 3-5 d after nephrectomy, although RKW of the removed kidney was 0.66 +/- 0.01%. In the Nx+Dx group, LKW increased to 0.97 +/- 0.15%, which was similar to that of the Nx group. In the Dx+Dx group, LKW (0.56 +/- 0.05%) and RKW (0.54 +/- 0.05%) were significantly less than those in the control group. We conclude that the renal nerves may contribute to maintaining the renal mass and that the neural effect on compensatory growth following nephrectomy may be covered by other growth factors. PMID- 10529498 TI - Automatic measurement of the red cell oxygen dissociation curve identical with the whole blood curve. AB - Automatic measurement of the entire oxygen dissociation curve (ODC) of blood and hemoglobin provides a useful means for evaluating their gas-transport function. The automatic oxygenation apparatus previously developed by Imai et al. (1970, 1981), which uses a polarographic determination of partial pressure of oxygen and a spectrophotometric determination of oxygen saturation of hemoglobin, has mostly been used for the measurement of accurate ODCs of hemoglobin solution. However, it was not suitable for red cell suspension because a significant noise was superimposed on the absorbance signal due to light-scattering by red cells. In the present study, we have overcome this problem by using an integrating sphere for the photometric system. Through extensive tests we found the optimal experimental conditions for obtaining the red cell oxygenation data that were identical with the whole blood data with respect to the position (oxygen affinity) and shape (sigmoid character) of the ODC and its pH-dependence (the Bohr effect). The accuracy was higher than that of commercially available automatic apparatuses such as the "Hemox-Analyzer" (Technical Consulting Service) and "Hem-O-Scan" (Aminco). Thus, our method provides an easy and convenient means for obtaining accurate ODCs mimicking the whole blood ODCs from one drop of whole blood. An application of our method to the effect of blood storage on ODC is presented, demonstrating the usefulness of our method. PMID- 10529499 TI - Biphasic nature of inotropic action of nitric oxide donor NOC7 in guinea-pig ventricular trabeculae. AB - The effects of nitric oxide (NO) donor on the contractility of guinea-pig ventricular trabeculae were explored to clarify whether NO affects the function of sarcoplasmic reticulum (SR) and the contractile elements. NO donor, 3-(2 hydroxy-1-methyl-2-nitroso-hydrazino)-N-methyl-1-propanamine (NOC7), increased monotonically the amplitude of the twitch tension induced by electrical stimulation at a concentration of 20 microM. A higher concentration of NOC7 (200 microM) caused a biphasic response: transiently increased the amplitude of twitch and then decreased it. On wash-off of the higher concentration of NOC7, a rebound increase of the twitch amplitude was observed. An inhibitor of NO-sensitive guanylyl cyclase, 1H-[1,2,4]oxadiazolo-[4, 3-a]quinoxalin-1-one (ODQ), abolished the mono-tonic increase and rebound increase in the amplitude of tension but did not affect the decrease in the amplitude of tension at the higher concentration of NOC7. Oscillatory contractions developed by beta-escin-skinned muscle fibers were not changed by NOC7 at either concentration. Caffeine-induced tension transients indicating the Ca(2+)-accumulating and -releasing functions of intracellular Ca(2+) stores were not affected by NOC7. NOC7 did not change the steady tension developed in 1.6 microM Ca(2+) containing solution with and without ODQ. These results suggest that the biphasic inotropic effects by NOC7 were not caused by modifying the function of SR and the Ca(2+) sensitivity of myofilaments of the guinea-pig ventricular trabecula, but at least the positive inotropic effect was mediated through cGMP-dependent mechanisms. PMID- 10529500 TI - Determination of the maximum steady state of lactate (MLSS) in saliva: an alternative to blood lactate determination. AB - Based on previous research which shows parallelism between the saliva and blood lactate response during incremental exercise, we hypothesized that a "maximum salivary lactate steady state" (saliva-MLSS) might exist. Thus, the aim of the present investigation was to establish 1) which lower limit for the increase in salivary lactate concentration during a constant workload (i.e., from the 10th to the 20th min) test could be used to determine the saliva-MLSS and 2) if the exercise intensity corresponding to the saliva-MLSS is identical to that evoking the (blood) MLSS. Twelve male amateur athletes of mean ( +/- SD) age 24 +/- 5 year were selected for the study. Based on the results of a previous maximal cycle ergometer test for lactate threshold (LT) determination, each subject performed consecutive constant workload tests of 20-min duration on separate days for MLSS determination. Blood and saliva (25 microl) samples were collected at 0, 10, and 20 min during the tests for lactate determination. A Student's t-test for paired data demonstrated that a salivary lactate increase of 0.8 mM corresponded to the saliva-MLSS. At this value, indeed, no significant differences were observed between the mean V(.)O(2) and W values corresponding to the MLSS and the saliva-MLSS. In conclusion, the present findings indicate that 0.8 mM is the lower limit for the increase in saliva lactate concentration during a constant load test and thus is that which might be used as a reference to determine saliva MLSS. Furthermore, saliva-MLSS might be used as an alternative to MLSS determination in blood samples. PMID- 10529501 TI - Natural goal-directed movements and the triphasic EMG. AB - Triphasic electromyographic (EMG) patterns have been described as characteristic of rapid, discrete, uniplanar, goal-directed movements. This experiment examined the effects of Response Type (experimenter- vs. subject-determined), Hand (preferred vs. nonpreferred), and Practice (early vs. late) on performance accuracy, and specific temporal EMG and kinematic measures during a dart throw. EMG was recorded from triceps (main agonist), brachioradialis, and biceps (main antagonist). The number of trials in which a triphasic EMG occurred varied systematically across conditions. The experimenter-determined, early practice condition resulted in greatest frequency (92%) of trials displaying a triphasic EMG and least accurate performance. In contrast, the lowest frequency (79%) of triphasic EMG and most accurate performance occurred in the subject-determined, late practice condition. The association among 14 temporal EMG, and kinematic measures for each trial of the dart throw was analyzed with multivariate factorial ANOVA. Four clusters of variables emerged: initial phase, braking phase, terminal phase, and movement speed and duration. Variables contributing to the initial-phase cluster were most strongly associated within the experimenter determined, early practice condition, and the strength of association was directly related to diminished performance accuracy. Paradoxically, best performance accuracy (subject-determined, late practice) was identified with a weaker association among variables representing the initial phase. PMID- 10529502 TI - Spontaneous and intentional pattern switching in a multisegmental bimanual coordination task. AB - Two experiments required right-handed subjects to trace circular trajectories while complying with either a symmetric or asymmetric pattern. In symmetric patterns, circles were traced in a mirror image either inward or outward. In asymmetric patterns, circles were traced in the same direction either clockwise or counterclockwise. Subjects were instructed to trace with spatial accuracy while maintaining a strict temporal relationship to a metronome that scaled movement rates from 1.25 to 3 Hz. The symmetric patterns were more stable than asymmetric patterns; the circularity of trajectories was greater for the dominant side; and there were spontaneous reversals in the direction of circling in the nondominant limb when performing asymmetric patterns. The second experiment examined the same subjects under the instruction of intentionally changing the pattern by reversing the left or right limb circling direction when cued to do so. The degree of interlimb interference was highly asymmetric and contingent on the direction of pattern change. Intentional direction reversals were more expedient and with less disruption to the contralateral limb when asymmetric to symmetric pattern changes were effected through a reversal in the direction of nondominant side. The results are interpreted with reference to evidence that the supplementary motor area mediates descending input to the upper limbs during disparate bimanual actions, but not during symmetric actions. PMID- 10529503 TI - Identification of time-varying stiffness, damping, and equilibrium position in human forearm movements. AB - Knowledge of how stiffness, damping, and the equilibrium position of specific limbs change during voluntary motion is important for understanding basic strategies of neuromotor control. Presented here is an algorithm for identifying time-dependent changes in joint stiffness, damping, and equilibrium position of the human forearm. The procedure requires data from only a single trial. The method relies neither on an analysis of the resonant frequency of the arm nor on the presence of an external bias force. Its validity was tested with a simulated forward model of the human forearm. Using the parameter estimations as forward model input, the angular kinematics (model output) were reconstructed and compared to the empirically measured data. Identification of mechanical impedance is based on a least-squares solution of the model equation. As a regularization technique and to improve the temporal resolution of the identification process, a moving temporal window with a variable width was imposed. The method's performance was tested by (a) identifying a priori known hypothetical time-series of stiffness, damping, and equilibrium position, and (b) determining impedance parameters from recorded single-joint forearm movements during a hold and a goal directed movement task. The method reliably reconstructed the original angular kinematics of the artificial and human data with an average positional error of less than 0.05 rad for movement amplitudes of up to 0.9 rad, and did not yield hypermetric trajectories like previous procedures not accounting for damping. PMID- 10529504 TI - Effects of direction and curvature on variable error pattern of reaching movements. AB - A number of studies have analyzed various indices of the final position variability in order to provide insight into different levels of neuromotor processing during reaching movements. Yet the possible effects of movement kinematics on variability have often been neglected. The present study was designed to test the effects of movement direction and curvature on the pattern of movement variable errors. Subjects performed series of reaching movements over the same distance and into the same target. However, due either to changes in starting position or to applied obstacles, the movements were performed in different directions or along the trajectories of different curvatures. The pattern of movement variable errors was assessed by means of the principal component analysis applied on the 2-D scatter of movement final positions. The orientation of these ellipses demonstrated changes associated with changes in both movement direction and curvature. However, neither movement direction nor movement curvature affected movement variable errors assessed by area of the ellipses. Therefore it was concluded that the end-point variability depends partly, but not exclusively, on movement kinematics. PMID- 10529505 TI - Kinematic characteristics of aiming movements as a function of temporal and spatial constraints. AB - This study investigated, in aiming movements, the conditions needed to produce a single movement and those needed to produce secondary submovements. Using a 2x2 (Temporal Constraint x Spatial Constraint) factorial design with repeated measures on both factors, subjects moved a stylus from a starting position to a target position 12 cm away. They participated in two testing sessions on consecutive days. The first session involved two nonrestrictive target (a set of crosshairs) conditions, moving to the target either within a goal of 400 ms (temporal-accuracy procedure) or within a minimum time (time-minimization procedure). In the second session the subjects performed two strict-target (circle) conditions, moving to the target either within a goal of 400 ms or within a minimum time. The results showed that the two strict-target conditions had a greater percentage of trials containing multiple-submovements than the two nonrestrictive target conditions, regardless of temporal requirements. Therefore, whether an aiming movement contains a single movement or multiple submovements may be a function of spatial constraints regardless of temporal constraints. It appears that with respect to the nature of the speed-accuracy tradeoff, spatial constraints are an important factor. PMID- 10529506 TI - Neuromuscular compartments of cat lateral gastrocnemius produce different torques about the ankle joint. AB - The primary purpose of this study was to establish whether the neuromuscular compartments of cat lateral gastrocnemius produce different mechanical actions on the skeletal system, by determining the contributions made by these compartments to the torques produced about the ankle joint. It was postulated that neuromuscular compartments might represent output elements of the spinal circuits. If so, they should produce unique mechanical actions. Isometric torques about the center of the ankle joint produced by the neuromuscular compartments of the cat lateral gastrocnemius were measured with a multiaxis force-moment sensor connected to the plantar surface of the foot. Individual compartment torques were elicited by activation of the primary compartment branches of the lateral gastrocnemius nerve. The magnitude of the individual torque components, and thus of the resultant torque, was significantly different between compartments. In three of the four lateral gastrocnemius compartments, significantly different torque trajectories were noted. The results, together with those from previous studies demonstrating that compartments can be activated in a task-dependent manner, suggest that neuromuscular compartments represent anatomical substrates that can be used by the nervous system for regulating movement. PMID- 10529507 TI - All-trans-retinoic-acid- and growth-factor- mediated induction of alkaline phosphatase activity in freshly isolated chronic myeloid leukemia cells. AB - Reduced or absent neutrophil alkaline phosphatase (NAP) activity is a common feature of neutrophilic granulocytes from patients with chronic myeloid leukemia (CML). In this study we examined whether NAP activity could be restored in vitro by stimulating CML cells with different promoters such as all-trans-retinoic acid (ATRA), granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF). The results obtained indicated that ATRA and G CSF, either alone or in combination, were effective in inducing NAP activity in CML cells, whereas GM-CSF was not. Further, NAP restoration in ATRA- and G-CSF treated cultures was accompanied by increased morphologic differentiation of the CML clone. It might be concluded that the CML clone could be driven in vitro by ATRA and G-CSF both to achieve granulocytic maturation and to correct functional NAP-related defects. PMID- 10529508 TI - The polysaccharide, PGG-glucan, enhances human myelopoiesis by direct action independent of and additive to early-acting cytokines. AB - beta-Glucans stimulate leukocyte anti-infective activity, enhance murine hematopoietic recovery following bone marrow injury and mobilize murine progenitor cells from bone marrow. This study evaluated the in vitro hematopoietic potential of the beta-glucan, PGG-glucan, on human bone marrow mononuclear cells (BMMC) and CD34+ BMMC compared with protein cytokines. In the presence of submaximal concentrations of recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF; 0.5 ng/ml), PGG-glucan significantly increased BMMC myeloid colony formation comparable to the increase observed with either interleukin-3 (rhIL-3) or stem cell factor (rhSCF). Moreover, the addition of PGG-glucan to cultures containing GM-CSF + IL-3 or GM-CSF + SCF significantly augmented granulocyte-macrophage colony production above baseline, demonstrating that PGG-glucan acts independently of those early-acting cytokines and can enhance their activity in an additive manner. Anti-PGG-glucan monoclonal antibody specifically abrogated the growth-enhancing effect of added PGG-glucan in a saturable manner and other control carbohydrate polymers failed to affect colony formation. Further, PGG-glucan was not associated with induction of IL-6, GM-CSF production and removal of accessory cells by CD34+ cell isolation did not alter the PGG-glucan effect. These data demonstrate that PGG-glucan acts on committed myeloid progenitors to enhance human hematopoietic activity by a mechanism of direct action independent of IL-3 or SCF and independent of secondary cytokine stimulation. PMID- 10529509 TI - wt1 gene expression in childhood leukemias. AB - The expression of the Wilms' tumor gene (wt1) was detected in various tissues during embryonic development. Mutations in the wt1 gene probably play an important role in certain tumors, e.g. the Wilms' tumor. Furthermore the expression of wt1 gene was found in some human leukemias. In the present study we investigated the expression of wt1 gene in several types of childhood leukemia by reverse transcriptase-polymerase chain reaction. Bone marrow or peripheral blood of 61 pediatric patients (48 at initial diagnosis, 13 at first or second relapse) were analyzed. wt1 gene expression was detected in 35/48 patients (73%) with newly diagnosed leukemias and in 12/13 cases (92%) who had suffered from relapse. The expression levels were higher for AML than for ALL. The frequency of wt1 expression in different subtypes of acute leukemia was compared with results found in adult patients. Our results show that the frequency of wt1 gene expression in acute childhood leukemias is similar to previous data reported for adults. PMID- 10529510 TI - Antigenic phenotyping of lymphoid cells and B cell gene rearrangement in type B gastritis and in gastritis not associated with Helicobacter pylori colonization. AB - Marginal-zone B cells of the mucosa-associated lymphoid tissue (MALT) are the normal counterpart of the neoplastic cells in MALT lymphoma. In both cases these lymphocytes express surface immunoglobulins, but are negative when stained for B cell associated antigens like CD10 and CD23. Furthermore, the B cell gene rearrangement has been found in Helicobacter pylori associated chronic gastritis and in extranodal type of marginal-zone lymphoma. The aim of this study was to quantify the number of IgM-, CD10-, and CD23-positive lymphocytes in patients with type B gastritis and to compare the results with the antigen profile of mononuclear cells in patients with gastritis not associated with H. pylori. Additionally, the immunoglobulin heavy-chain (IgH) gene rearrangement in H. pylori positive and H. pylori negative gastritis was studied. From 23 patients with a positive urease test and/or histologically proven H. pylori infection and chronic gastritis and from 22 patients with H. pylori negative chronic gastritis mucosa biopsy specimens were taken. Single-cell suspensions were obtained following enzymatic digestion. For immunocytochemistry, an alkaline phosphatase antialkaline phosphatase method was applied. IgH gene rearrangement in formalin fixed, paraffin-embedded specimens was determined by polymerase chain reaction in 11 patients with chronic gastritis. An increase in mu-positive plasma cells and B lymphocytes was detected in patients with H. pylori positive gastritis as compared with patients with H. pylori negative gastritis (10.0 vs. 3.9%, p < 0.001, and 4.3 vs. 1.6%, p < 0.01, respectively). In both groups, the proportion of CD10- and CD23-positive lymphocytes was <1%. IgH gene rearrangement was not restricted to type B gastritis; single bands were also present in 3 of 7 patients with H. pylori negative chronic gastritis. Our finding of IgH gene rearrangement in some of the patients with H. pylori negative chronic gastritis indicates that additional factors may be critical for these genotypical changes and for the pathogenesis of gastric MALT lymphoma. PMID- 10529511 TI - Splenomegaly at a university hospital compared to a nearby county hospital in 317 patients. AB - Splenomegaly and massive splenomegaly were diagnostically evaluated retrospectively at Stanford University Hospital in 147 patients over 8 years and compared to the nearby county hospital (Santa Clara Valley Medical Center; VMC) in 170 inpatients over 11 years. Hematologic diseases at Stanford (data for VMC in parentheses) occurred in 66% (35%; p < 0.001) of the patients with splenomegaly and in 84% (54%; p < 0.001) of those with massive splenomegaly. Hepatic diseases occurred in 9% (36%; p < 0.001) of the patients with splenomegaly and in 5% (29%; p < 0.001) of those with massive splenomegaly. Splenectomy was performed in 71% of the patients at Stanford and in 9% of those at VMC (p < 0.001). The combined Stanford-VMC series showed significant associations (p < 0.01): for hematologic diseases with massive splenomegaly, lymphadenopathy, and blood cytoses; for hepatic diseases with hepatomegaly, cytopenias as hypersplenism, and abnormal liver function tests, and for infectious diseases with fever. PMID- 10529512 TI - Flow-cytometric analysis of leukocyte alkaline phosphatase in myelodysplastic syndromes. AB - The expression of leukocyte alkaline phosphatase (LAP) in neutrophils is reduced in some patients with myelodysplastic syndrome (MDS). We quantitatively assayed for LAP in MDS leukocytes by a flow cytometry based method using a monoclonal antibody raised against human bone alkaline phosphatase. The LAP expression was assayed in blood samples from a group of 46 MDS patients, consisting of 39 patients with refractory anemia (RA), 3 with RA with excess blasts (RAEB), and 4 patients with RAEB in transformation. The percentage of LAP-positive cells was significantly higher in the MDS patients than in the normal subjects and also higher in RA than in RAEB and RAEB in transformation. To investigate the cause of the elevated LAP expression, we measured the serum concentrations of several cytokines. The granulocyte colony-stimulating factor (G-CSF) level was significantly elevated in MDS patients, and the serum G-CSF concentration clearly correlated with the percentage of LAP-positive cells. Thus, the LAP activity in RA is higher than in normal subjects, and G-CSF is thought to be one of the causes stimulating LAP expression in MDS neutrophils. PMID- 10529513 TI - Rituximab (anti-CD20 monoclonal antibody) administration in a young patient with resistant B-prolymphocytic leukemia. AB - Following the administration of the human anti-CD20 monoclonal antibody IDEC-C2B8 (rituximab), a 31-year-old woman with B-prolymphocytic leukemia, who had been resistant to CHOP, fludarabine, pentostatin and 2-CdA, achieved complete remission. Rituximab was administered intravenously once a week for 4 weeks. The patient only had mild but tolerable side effects during the first cycle of therapy. She remains in complete remission 8 months following the discontinuation of treatment. PMID- 10529514 TI - Cladribine treatment of a patient with hairy cell leukemia and concomitant multiple sclerosis. AB - This is the first report on a patient suffering from both multiple sclerosis (MS) and hairy cell leukemia. The patient was first treated with interferon-alpha. Due to disease progression two courses of cladribine were given resulting in an improvement of the clinical course of both diseases. Interestingly, it was possible to arrest and even ameliorate the progression of MS by administering as little as 30% of the dosage recently recommended for the treatment of this disease. PMID- 10529515 TI - Homozygous prothrombin gene mutation and ischemic cerebrovascular disease: a case report. AB - We report the case of a 31-year-old woman who, at the age of 26 suffered from an episode of superficial thrombophlebitis in the left leg, experienced two episodes of transient ischemic attacks at the age of 30 and had an ischemic stroke with left-sided hemiparesis at the age of 31 years. A cerebral CT scan showed an ischemic lesion in the right sylvian area involving the opercular and nucleocapsular regions. Her father had had an ischemic stroke at the age of 54 years and died at the age of 58; her mother had had a myocardial infarction at the age of 48 years and died at 51 years from breast cancer. Laboratory investigation of the patient demonstrated high levels of fibrinogen, F II, F VII, F 1 + 2, FPA and ACA-IgG with low levels of HDL cholesterol associated with homozygosity for the 20210 A genotype. There were no other genetic or acquired prothrombotic defects. In conclusion, this case strongly suggests a clinically significant role ot the prothrombin gene mutation in both arterial and venous thrombosis. PMID- 10529516 TI - Superior sagittal sinus thrombosis occurring at high altitude associated with protein C deficiency. AB - A 42-year-old male presented with right-sided weakness, dysphasia and seizures while climbing the French Alps at an approximate altitude of 3,000 m. Imaging studies were consistent with superior sagittal sinus thrombosis with hemorrhage. Laboratory testing for thrombophilic states, 18 days after presentation at our hospital, showed a low protein C level (0.32 U/ml, normal 0.80-1.60 U/ml). A family member was also found to have protein C deficiency without a history of thrombosis. The patient gradually improved and was discharged on warfarin and valproic acid. This is the first reported case of cerebral venous thrombosis in a patient with congenital protein C deficiency who ascended to high altitude. We postulate that the ascent to high altitude represented an additional prothrombotic risk factor to the congenital protein C deficiency leading to cerebral thrombosis. PMID- 10529517 TI - Successful treatment of chronic graft-versus-host disease with sulfasalazine in allogeneic bone marrow transplantation. PMID- 10529518 TI - Engraftment of allogeneic marrow following conditioning the donor with G-CSF. PMID- 10529519 TI - Macrolides and oral anticoagulants: a dangerous association. PMID- 10529520 TI - Synaptogenesis in retino-receptive layers of the superior colliculus of the opossum Didelphis marsupialis. AB - The maturation of the neuropil and synapse formation were examined in the retino receptive layers of the superior colliculus (SCr-r) in the opossum from a period prior to the onset of arborization of retinocollicular fibers (postnatal day 22 - P22), at 44% of the coecal period (CP), to the end of the fast phase of optic fiber myelination and weaning time (P81 - 118% CP). Development of the SCr-r neuropil follows a protracted time course and can be divided into three broad stages, which are characterized by (I) Large extracellular spaces, numerous growth cones that participate rarely in synaptic junctions, vesicles-poor immature synapses (P22-P30), (II) Synapses of varied morphology with abundant synaptic vesicles, and small terminals with dark mitochondria and round synaptic vesicles (RSD terminals) synapsing mostly onto dendritic shafts, flat-vesicles (F) terminals (P40-P56), (III) Sequential appearance of retinal (R) and pleomorphic-vesicles (P) terminals and of RSD terminals synapsing onto spine or spine-like processes, appearance of glomerulus-like synaptic arrays (synaptic islets) (P61-P81). The advancement of synaptogenesis in SCr-r from stage I to II and from stage II to III correlates closely with the differentiation of astrocytes and oligodendrocytes, respectively. PMID- 10529521 TI - Modality-specific segregation of input to ant mushroom bodies. AB - The mushroom bodies are central brain neuropils involved in the control of complex behavior. In ants, the mushroom bodies are relatively large compared to those of honey bees, whereas the optic lobes of ants are considerably smaller. The general morphology of ant mushroom bodies is similar to that of honey bees. As in other Hymenoptera, the main input region of the mushroom bodies, the calyx, is subdivided into three compartments: the lip, the collar, and the basal ring. In many ant species this compartmentalization is not obvious and can only be visualized using neuronal tracers. The lip region receives antennal input and is large in all ant species. It appears to be composed of at least two different regions that have not yet been characterized in detail. The collar is large in other Hymenoptera, yet in ant workers it varies in size and is always much smaller than the lip region. The collar receives visual input and is relatively larger in males, which generally are more dependant on vision than are workers. The basal ring receives input from both the optic and antennal lobes. In one ant tribe, the Ponerini, the collar region appears to have changed its position, but based on afferent input it appears to be homologous to the hymenopteran collar. Generally, the composition of the mushroom body calyx correlates with the living conditions of ants, reflecting the great importance of olfaction and the lesser and more variable significance of vision for workers of the observed ant species. PMID- 10529522 TI - The ocular morphology of the southern hemisphere lamprey geotria australis gray, with special reference to optical specialisations and the characterisation and phylogeny of photoreceptor types. AB - This paper describes the ocular morphology of young adults of the southern hemisphere lamprey Geotria australis, the sole representative of the Geotriidae, and makes comparisons with those of holarctic lampreys (Petromyzontidae). As previously reported for the holarctic lamprey Ichthyomyzon unicuspis [Collin and Fritzsch, 1993], the lens of G. australis is non-spherical and possesses a cone shaped posterior that may be capable of mediating variable focus. The avascular retina of G. australis is well differentiated, containing three retinal ganglion cell populations, three layers of horizontal cells and three photoreceptor types. In contrast to petromyzontids that contain only two photoreceptor types (short and long), G. australis possesses one rod-like (R1) and two cone-like (C1 and C2) photoreceptors. Although the rod-like receptor in G. australis may be homologous with the short receptors of holarctic lampreys, the two cone-like receptors have morphological characteristics that differ markedly from those of the long receptors of their holarctic counterparts. The features which distinguish the two cone-like receptors from those of the long receptor type in holarctic lampreys are the characteristics of the mitochondria and the presence of large amounts of two different types of stored secretory material in the endoplasmic reticulum of the myoid (refractile bodies). The endoplasmic reticulum of each receptor type has a different shape and staining profile and is polymorphic, each showing a continuum of distension. It is proposed that the presence of two cone-like photoreceptors with different characteristics would increase the spectral range of G. australis and thus be of value during the parasitic phase, when this lamprey lives in the surface marine waters. The irideal flap, present in G. australis but not petromyzontids, would assist in reducing intraocular flare during life in surface waters. The results of this study, which are discussed in the context of the proposed evolution of lampreys, emphasise that it is important to take into account the characteristics of the eyes of southern hemisphere lampreys when making generalizations about the eyes of lampreys as a whole. PMID- 10529523 TI - Encephalization, adaptation and evolution of chiroptera: A statistical analysis with further evidence for bat monophyly. AB - As part of a large-scale study on brain morphometrics and adaptations in mammals, we addressed the problem of chiropteran evolution. A specific statistical framework was designed to test which of two competing hypotheses (bat monophyly vs. diphyly) is more strongly supported by quantitative brain data. Our analyses, based on 120 species, revealed that megabats and microbats were more closely related to each other than to primates, and illustrated the convergent adaptations of the brain of bats to similar trophic (i.e. feeding related) niches. Ecologically-corrected characters were then used to derive a new phylogeny which also supports the chiropteran clade. The monophyletic origin of bats is the preferred hypothesis to explain brain quantitative evolution in chiropterans and primates. PMID- 10529524 TI - Dental caries, sugar-eating habits and toothbrushing in groups of 4-year-old children 1967-1997 in the city of Umea, Sweden. AB - Four-year-old children in the city of Umea, northern Sweden, have been the subjects for studies of dental caries at regular intervals between 1967 and 1997. Similar methods and criteria were used in all studies and the children were selected from the same catchment areas. The results of this study signified a shift in the trend towards a declining caries prevalence among 4-year-old children. There was a slight non-significant increase in the number of children with caries as well as in the mean dmft and dmfs values between 1992 and 1997. The mean dmfs value was 7.8 in 1967 and declined to 4.5 in 1971. It was 2.9 in 1976 and 2.0 in 1980 and 1987, 1.8 in 1992 and increased to 2.0 in 1997. Ten percent of the children were immigrant or refugee children in 1997 compared to 6% in 1992 and they had a significantly higher caries prevalence than children with a Swedish background (p<0.001). In 1997, 58% of the children had one or more daily intakes of sugary snacks such as buns, cakes sweets, soft drinks, etc. There was an increasing consumption of sweets and soft drinks between 1987 and 1997. PMID- 10529525 TI - Behaviour of approximal carious lesions assessed by clinical examination after tooth separation and radiography: a 2.5-year longitudinal study in young adults. AB - This study aimed to record and monitor over a 2.5-year period the occurrence of cavitation and lesion depth progression in approximal surfaces with radiographic caries at baseline. In total, 66 approximal sites (in 29 students), where at least one of the contacting surfaces had radiographic caries, were selected to take part in the study. A clinical examination undertaken before and after tooth separation in order to assess the presence/absence of cavitation was repeated every sixth month. To monitor lesion progression bite-wing radiographs were taken every sixth month, too. After each series of examinations, surfaces judged to be prone for disease progression were referred to operative caries treatment. In surfaces with radiographic dentinal caries at baseline the cavitation prevalence following tooth separation found at the various recall examinations ranged from 20 to 44%. In surfaces with radiographic enamel caries at baseline this prevalence ranged from 4 to 8% at the various recall examinations. In dentinal lesions found with an intact surface at baseline, the risk of cavitation development during the first 1.5-year period was assessed to be up to 22%. After this period no new cavitations were found in previously intact dentinal lesions. In intact enamel lesions the risk of cavitation formation was found to be 3% during the first 1-year period. After this period no new cavitations developed in previously intact enamel lesions. Three of 7 lesions, which showed radiographic caries progression from the outer one third to the inner two thirds of the dentine during the observation period, had intact surfaces at baseline. On the basis of these results it is recommended to re-examine carefully intact, dentinal lesions by repeated clinical examination after tooth separation and by radiography about 1-1.5 years after baseline. PMID- 10529526 TI - Caries-preventive effect of topical amine fluoride in children with high and low salivary levels of mutans streptococci. AB - The aim of the study was to assess the relationship between the salivary mutans streptococci (SMS) level and the effectiveness of a preventive intervention based on a biannual application of an amine fluoride solution (AmF). A total of 284 schoolchildren aged 6 years were recruited from eleven classes of a primary school in Milan and randomly assigned to an experimental (A) and a control group (B). SMS counts were obtained at baseline and caries incidence data (diseased, missing, filled teeth, DMFT) were recorded every 6 months for 5 years. The participants of the experimental group received application of an AmF 1% F(-) solution twice a year on the enamel surfaces of the first permanent molars for 5 years. Control group subjects received application of a placebo solution twice a year on the enamel surfaces of the first molars for the same period. The mean DMFT in the experimental and control groups were 0.56 and 0.22, respectively, at the beginning and 1.14 and 2.06 after 5 years. SMS data allowed children to be classified into low- (0-10(5) CFU/ml of saliva) and high- (>10(5) CFU/ml of saliva) SMS subjects. Survival analysis, performed on the first molar data split by SMS group, showed a significantly higher caries reduction in low-SMS experimental group subjects compared to low-SMS control group subjects after 5 years. No significant differences were found between the two high-SMS experimental and control groups. These findings indicate that the preventive effects of the treatment were significantly lower in subjects who had high SMS. It is concluded that the effectiveness of a simple and economical topical fluoride intervention applicable at a community level is significantly influenced by the SMS level of the subjects involved. Simple AmF preventive interventions, applied on low-SMS subjects, can give significant results in terms of caries reduction. PMID- 10529527 TI - Plaque pH and associated parameters in relation to caries. AB - Intensified plaque acidogenicity in caries-prone subjects was reported many years ago, but emerging evidence has suggested that the relationship may not be as strong as once thought. We have now determined a range of acidogenicity variables in subjects having both caries prevalence and incidence data, and have included plaque mineral data in the analysis. pH measurements were made in 20 randomly selected subjects from a high-caries group (mean DMFS = 8. 95) and 20 from a caries-free group of Beijing children aged 12 years participating in a caries prediction study. Subgroups with a 12-month DMFS increment >/=2 or = 0 were also formed from the two groups, respectively. Measurements were made with an iridium oxide electrode inserted between teeth 13/14, 23/24, 34/35 and 44/45, before and every 5 min for 30 min after rinsing with 10% sucrose, and the 4 resulting 'Stephan curves' averaged using a plaque pH analysis program. Supragingival plaque was collected from buccal and lingual smooth surfaces of posterior and upper anterior teeth and its acid extract analysed for Ca, P and F. Caries-free subjects (based on past experience) had a significantly higher maximum plaque pH and pH value after 30 min (reflecting a faster return to resting pH), a lower minimum enamel dissolution capacity of plaque and recorded less time below pH 7.0 than did high-caries subjects. No other differences were significant, including those of the principal acidogenic parameters 'minimum pH attained after a sugar rinse', 'curve area below the critical pH of 5.5' and 'time below the critical pH'. Selection of the caries groups on the basis of both experience and incidence did not reveal significant differences in more parameters. Upper arch plaque was significantly more acidogenic than lower arch plaque, and there was a consistently strong association between upper and lower arch values in individuals. Ca, P and F in the subjects' plaque had little or no influence on the principal acidogenic parameters. Our failure to find a relationship between caries prevalence or activity and these principal acidogenicity parameters may be related to differences between fissure and smooth surface plaque, temporal variations in acidogenicity and/or to use of F toothpaste during the 1-year observation period. These results support the view that factors such as the frequency of acidogenic episodes may be more important in caries progression than the degree of acidogenicity during any one episode. PMID- 10529528 TI - In vitro effectiveness of hand excavation of caries with the ART technique. Atraumatic restorative treatment. AB - The purpose of this study was to evaluate the effectiveness of the atraumatic restorative treatment (ART) hand-instruments and of round steel burs for removing caries at the enamel-dentine junction (EDJ) in occlusal cavities prepared in 50 extracted permanent molars. The teeth were divided randomly into two equal groups for the two treatments provided by 1 operator. Stained but hard dentine was not removed. The teeth were then sectioned vertically, buccolingually through the prepared cavities to give 200 sections, each 500 microm thick, which were photographed (x1 magnification) before and after staining with a caries detector dye. Assessments were made (x3 magnification) of (a) the amount of brown-stained residual dentine, and (b) the amount of red dye-stained dentine present at the EDJ, using two sets of standards devised and transparencies arranged as six category incremental rating scales. There was no statistically significant difference found between the two caries-removal methods for the amount of residual brown-stained dentine present (chi(2 )= 3. 394, p = 0.64), but there was for the amount of red dye-stained dentine present (chi(2) = 32.137, p<0.0001), although 57% of the ART and 80% of the steel bur sections had very little to mild red-dye staining present. Excavation of caries at the DJE appeared to be less effective with the ART technique, although the clinical implications of this may not be significant. PMID- 10529529 TI - Inhibitory effects of oolong tea extract on caries-inducing properties of mutans streptococci. AB - The inhibitory effects of oolong tea extract (OTE) on the caries-inducing properties of mutans streptococci were examined in vitro. OTE reduced the rate of acid production by mutans streptococci accompanied with the retardation of growth rate of mutans streptococci, while the action by chromatographically isolated oolong tea polyphenol (OTF6) was weak. On the other hand, both oolong tea products decreased cell surface hydrophobicity of almost all the oral streptococci examined in the present study, and also induced cellular aggregation of Streptococcus mutans, Streptococcus oralis, Streptococcus sanguis or Streptococcus gordonii. In these reactions, OTF6 showed a more pronounced activity than OTE. Furthermore, the oolong tea products inhibited the adherence of mutans streptococci to saliva-coated hydroxyapatite. These results suggest that OTF6 may inhibit bacterial adherence to the tooth surfaces by reducing the hydrophobicity of mutans streptococci, and OTE may inhibit caries-inducing activity of mutans streptococci by reducing the rate of acid production. PMID- 10529530 TI - Powdered milk micellar casein prevents oral colonization by Streptococcus sobrinus and dental caries in rats: a basis for the caries-protective effect of dairy products. AB - Three animal studies were performed to investigate the influence of the macromolecular structure of milk casein on caries incidence and the possible ecological changes of the oral microbiota by such casein fractions. Towards this end, rats were infected with mixed bacterial suspensions of Streptococcus sobrinus OMZ 176 and Actinomyces viscosus Ny1. Various milk protein fractions were incorporated into carefully balanced powdered cariogenic diets to constitute the sole major protein component. Diets containing micellar casein had a pronounced and highly significant effect on almost all clinical and microbiological parameters examined. Both the formation of advanced dentinal fissure (B) and smooth surface (E) caries lesions was inhibited by diets containing micellar casein; this caries-inhibiting effect appeared to be due mainly to modifications within the plaque microbiota. The proportion of S. sobrinus in the oral cavity of rats was reduced (73-80%) by micellar casein containing preparations, whereas the A. viscosus population was increased. Both these microbiological parameters were always negatively correlated. This appears to be the first example of a food component other than dietary sugars, selectively modifying the composition of the dental plaque microbiota of rats in such a way as to reduce its pathogenic potential. It also demonstrates the importance of establishing a molecular basis for the role of food components, which prove to be beneficial to oral health. PMID- 10529531 TI - Toothbrush abrasion of erosively altered enamel after intraoral exposure to saliva: an in situ study. AB - The aim of this in situ study was to test the effect of toothbrush abrasion on enamel previously exposed to a standardized artificial erosive agent. To generate moderate erosive lesions, slabs of the buccal surface of human premolars were immersed in a solution of citric acid for 3 min. Then they were attached to intraoral appliances and each one was exposed for 0 min (= toothbrushing immediately after intraoral exposure), 30 or 60 min to the oral milieu of 1 of 7 female subjects with a mean age of 22 years. Immediately thereafter the volunteers brushed the slabs for 30 s with toothpaste using their preferred brushing technique. For each test person the secretion rate of resting and paraffin-stimulated saliva, buffering capacity and pH were measured. The following mean losses of substance at the surface were registered: 0.258+/-0.141 microm (toothbrushing immediately after intraoral exposure), 0. 224+/-0.087 microm (toothbrushing after intraoral exposure of 30 min) and 0.195+/-0.075 microm (toothbrushing after intraoral exposure of 60 min). Toothbrush abrasion in situ was significantly lower after 60-min exposure to the oral environment than after 0-min (p<0.001). Also, the 30- and 60-min values were significantly different from each other (p<0.001). Multiple linear regression analyses revealed that in this model toothbrush abrasion was associated with the intraoral exposure to saliva (p = 0.026), the severity of the erosive attack (p<0.001) and the secretion rate of resting saliva (p = 0.029). If no other preventive measures are taken we suggest that individuals at risk for erosive tooth wear wait at least 1 h before brushing their teeth after consuming erosive foodstuffs or beverages. PMID- 10529532 TI - Fingernail fluoride: a method for monitoring fluoride exposure. AB - This work was based on the hypothesis that fingernail clippings can be used as a biomarker for the subchronic exposure to fluoride. The results provide data on factors that may affect the concentration of fluoride in fingernail clippings as determined with the electrode following HMDS-facilitated diffusion. The following variables had only minor or no effects on the concentrations: (1) the surface area of the clippings (intact, minced or filed into powder) that were placed into the diffusion dishes; (2) soaking in deionized water for up to 6 h; (3) soaking in fluoridated water (1.0 ppm) for 2 h, and (4) removal of the organic material of nails by dry ashing. Fingernail fluoride concentrations were approximately 50% higher than those in toenails. A 1-month period of increased fluoride intake by one of the authors resulted in significant increases in fingernail fluoride concentrations after a lag time of approximately 3.5 months. The fluoride concentrations in fingernail clippings obtained from three groups of Brazilian children were directly related to the concentrations in the drinking water (0.1, 1.6 or 2.3 ppm). The results indicate that: (1) HMDS-facilitated diffusion completely separates fluoride from intact nail clippings, so the need for ashing or other preparative methods is obviated; (2) fingernail fluoride is derived mainly from the systemic circulation, and (3) fluoride intake is reflected by the concentrations in fingernails. PMID- 10529533 TI - Pronase digestion of carious dentin. AB - The aim of this study was to investigate the individual capabilities of the proteolytic enzyme preparation Pronase, the enzyme collagenase and sodium hypochlorite to disintegrate and solubilize carious dentin. Samples of carious dentin, and samples of sound dentin for comparison, were extracted 4 times in succession for 24 h with buffered solutions of Pronase. Separate carious dentin samples were extracted in the same manner with buffered solutions of collagenase or with aqueous sodium hypochlorite. The extracts, the solid residues left over after the extractions and untreated samples of carious and sound dentin were digested with sulfuric acid-H(2)O(2) and then analyzed for nitrogen content by a special adaptation of the Berthelot color reaction. Although Pronase did not attack sound dentin, it solubilized more than 90% of the nitrogen present in carious dentin. Collagenase solubilized approximately 66% of the nitrogen, whereas sodium hypochlorite released only 12-20% of the nitrogen of carious dentin. In clinical dentistry, chemical disintegration of carious dentin may reduce the need for mechanical removal of sound tooth structure. PMID- 10529534 TI - Measurements of the wettability of protein-covered hydroxyapatite surfaces. AB - We developed a new method (dropping time method, DTM) to investigate the wettability of a surface of a protein layer adsorbed on glass plates in aqueous solution. However, the previous setup of DTM can only be utilized for optically transparent materials. In this study, we have extended the method to optically nontransparent materials such as hydroxyapatite plates. DTM is based on measuring the dropping time of a liquid film along a protein-covered surface when this surface is instantaneously vertically removed from the protein solution. The intensity of the reflected light beam depends on the presence of a liquid film on the surface. This allows to estimate the movement of the liquid film along the sorbent surface. Thus, the extended DTM can be used for determining the wettability of optically nontransparent solid plates. The adsorption behavior of four proteins (albumin, lysozyme, beta-lactoglobulin, ovalbumin) on a hydrophobic hydroxyapatite plate in water was studied by this method. When adsorbed from a protein solution of high concentration, the surfaces of adsorbed proteins, except ovalbumin, were fairly hydrophilic; this hydrophilicity was already attained at the initial stage of the adsorption process. The surface of ovalbumin on hydroxyapatite was more hydrophobic than those of the other proteins, and the hydrophilicity increased with the protein adsorption process. At low protein concentration, the hydrophilicity increased in the course of the adsorption process. The change in hydrophilicity with time depends on the kind of protein. Hen's egg lysozyme is more hydrophilic and the time to reach saturation is shorter than for the other proteins. The processes of increasing hydrophilicity of the surface of human serum albumin, beta-lactoglobulin and ovalbumin are similar. However, for beta-lactoglobulin hydrophobicity at adsorption saturation is stronger than for human serum albumin and ovalbumin. Thus, using DTM it is shown that the hydrophilicity of the surface of adsorbed protein on hydroxyapatite depends strongly on the kind of protein. PMID- 10529535 TI - Escalating immunotherapy of multiple sclerosis.Austrian-German- Swiss Multiple Sclerosis Therapy Consensus Group [MSTCG]. AB - The promising results of several multicenter studies during the last few years have improved the immunomodulatory treatment of multiple sclerosis (MS). The different compounds tested were shown to reduce the number of relapses and to modulate the course of disease to various extents. The transition of the results obtained in therapeutic trials into daily clinical practice is often delayed or even hampered by monetary restrictions or reluctance of the medical community to adjust their approach to new treatments. After an initial inquiry had shown that less than 50% of eligible patients received any active immunomodulating treatment, a consensus group of Austrian, German and Swiss MS societies was formed in order to prepare a report of the current treatment options in MS. The aim of this report is to present the consensus on a new concept of escalating immunotherapy in MS. Future updates of the report are planned on a yearly basis or whenever substantial new evidence becomes available. PMID- 10529536 TI - Lenticulocapsular hemorrhages presenting as pure sensory stroke. AB - Pure sensory stroke (PSS) syndrome is most often produced by a small infarct involving the lateral thalamus. Larger than lacune-sized putaminal hemorrhages have not been considered as a cause of this syndrome. The author describes 3 patients with hypertensive lenticulocapsular hemorrhage presenting with hemisensory symptoms without any other neurological deficits. In these patients, the sensory symptoms were more marked and persistent in the legs than in the other body parts. Neuroradiological data suggested that thalamocortical sensory pathways were exclusively involved. These patients highlight the heterogeneity of the vascular lesion producing PSS syndrome and illustrate that a putaminal hemorrhage should be included in the differential diagnosis of this clinical syndrome. PMID- 10529537 TI - Coeliac disease presenting with neurological disorders. AB - It is well known that coeliac disease may be associated with various neurological manifestations. We have had a high index of suspicion of coeliac disease during recent years in our neurological clinic. As a result 10 (7%) out of 144 of our new coeliac patients were detected because of neurological symptoms. The most common neurological manifestations were neuropathy, memory impairment and cerebellar ataxia. In these patient groups screening for coeliac disease with serological antibody tests helps to find patients who may suffer from this disease. PMID- 10529538 TI - Effect of bilateral subthalamic nucleus stimulation and dopatherapy on oral control in Parkinson's disease. AB - This study focuses on the speech organs of a parkinsonian patient who initially had been treated with levodopa for 13 years, and had become severely disabled by motor fluctuations. This patient has been treated with bilateral chronic stimulation of the subthalamic nucleus (STN) for the last 2 years. Upper lip, lower lip and tongue force production were examined before surgery under off and on medication conditions, and 2 years after surgery under off and on stimulation conditions. We compared the effect of stimulation and dopatherapy on the speech organs. L-Dopa had a poor effect whereas bilateral stimulation improved oral control and speech intelligibility. These results suggest that STN stimulation influences speech organs in a different way from the dopaminergic system and similarly affects oral and limb motor systems. PMID- 10529539 TI - Delayed movement disorders after carbon monoxide poisoning. AB - Of 242 patients with carbon monoxide (CO) poisoning examined between 1986 and 1996, delayed movement disorders were diagnosed in 32 (13. 2%). There were 15 men and 17 women. Ages at insult ranged from 9 to 69 years (mean 45.3 years). Of the 32 patients with delayed movement disorders, 23 (71.9%) had parkinsonism, 5 dystonia, 3 chorea and 1 myoclonus. All were associated with delayed CO encephalopathy. The median latency between CO poisoning and the onset of movement disorders was 4 weeks for parkinsonism, 51 weeks for dystonia, 4 weeks for chorea and 8 weeks for myoclonus. The latency of dystonia onset after CO poisoning was longer than that of other types of movement disorders. The CT findings in delayed movement disorders after CO poisoning were variable, and there was no correlation between the sites of imaging and the development of movement disorders. Abnormal dyskinesias disappeared within 8 weeks, and patients recovered from parkinsonism within 6 months. In conclusion, delayed movement disorders after CO poisoning are not rare, and usually appear as a part of delayed CO encephalopathy. The prognosis is good. PMID- 10529541 TI - Risk factors for stroke in different levels of cerebral arterial disease. AB - BACKGROUND AND PURPOSE: Despite recent studies on risk factors of stroke, it is still unclear whether certain risk factors differently affect different levels of cerebral arteries in stroke patients. We attempted to study risk factors and lifestyle factors in patients with cerebrovascular diseases affecting different levels of the cerebral arteries. METHODS: We studied 343 Korean patients (222 men and 121 women) with acute stroke. Strokes were subdivided into large vessel infarction (LVI, n = 156), small vessel infarction (SVI, n = 105), intracerebral hemorrhage (ICH, n = 82) according to modified TOAST criteria. Patients with cardiogenic embolic infarction, undetermined causes, and miscellaneous causes were excluded. Among the patients with LVI and SVI, 104 and 49 underwent angiographic studies, respectively, who were designated as having definite LVI and definite SVI. Definite LVI was further subdivided into proximal LVI when the internal carotid artery (n = 25), vertebral artery (n = 13) or basilar artery (n = 14) were primarily involved, and distal LVI when the middle cerebral artery (n = 35), anterior cerebral artery (n = 7) or posterior cerebral artery (n = 10) were involved. Using a structured interview, we assessed the risk factors and lifestyles: hypertension, diabetes mellitus (DM), cigarette smoking (CS), alcohol consumption, physical activity and exercise, and the amount of consumption of salt, vegetables, fat, fish and fruits. Serum cholesterol, body mass index and total body fat were also measured. RESULTS: Multivariate analysis showed that in men, DM was a discriminant factor favoring brain infarction (BI) over ICH, definite LVI or ICH and proximal LVI over ICH. Decreased fruit intake differentiated distal LVI from definite SVI, and distal LVI from ICH. In women, DM and abdominal obesity differentiated BI from ICH, while DM favored definite LVI over ICH, proximal LVI over ICH and definite SVI over ICH. Multivariate analysis including sex as a separate variate showed that DM was a factor favoring BI over ICH, definite LVI over ICH, proximal LVI over ICH, distal LVI over ICH and definite SVI over ICH. CS was a factor favoring BI over ICH, while heavy CS was a factor discriminating distal LVI from definite SVI. Male sex was a factor favoring definite LVI over ICH and proximal LVI over ICH. A lesser amount of fruit consumption favored definite SVI over distal LVI. CONCLUSION: Our results demonstrate that DM in all sexes and central obesity in women are factors discriminating BI from ICH. However, these factors as well as others were not found to discriminate the sizes of the vessels involved in the patients with ischemic stroke, except that being male may be a factor favoring proximal large vessel disease over ICH. Genetic factors or other not yet identified risk factors may be responsible for the differences in the subtypes of ischemic stroke among different ethnic populations. PMID- 10529540 TI - Clinical characteristics of aged Becker muscular dystrophy patients with onset after 30 years. AB - To elucidate the clinical characteristics of aged patients with Becker muscular dystrophy (BMD), 4 patients with this disease who were over 50 years were examined. The ages at onset in all patients were later than 30 years. All were proven to have a deletion around exons 45-55 of the Duchenne muscular dystrophy (DMD) gene. Two patients became wheelchair bound in their 40s or beyond, while the other 2 (aged 73 and 69, respectively) were still able to walk at the time of examination. Three of 4 patients had no obvious hypertrophy in their calves, which is known to be one of the characteristic clinical features in the juvenile BMD patients. Serum creatine kinase levels were elevated in all patients, but not markedly (mean 444.8 +/- 230.3 U/l; normal value < 180 U/l). Dilated cardiomyopathy was clinically apparent in 2 patients. We emphasize that some BMD patients are free of muscular symptoms until their 50s and are still self supporting in their 60s or 70s. PMID- 10529542 TI - A case of hereditary ceruloplasmin deficiency with iron deposition in the brain associated with chorea, dementia, diabetes mellitus and retinal pigmentation: administration of fresh-frozen human plasma. AB - We report a familial case of hereditary ceruloplasmin deficiency (HCD) showing an A-G transition in intron 6 of the ceruloplasmin gene. Clinical features consisted of chorea, cerebellar ataxia, dementia, diabetes mellitus, retinal pigmentation and iron deposition in the liver and brain without copper overload in those organs. The patient's children and siblings had similar laboratory results, but did not show any neurological abnormalities. She was medicated for diabetes mellitus at 43 years of age, and neurological signs appeared when she was 52 years old. The laboratory findings were anemia, low concentrations of iron and copper in serum and of copper in urine. Ceruloplasmin was not detected in the serum. The iron and copper contents in the liver were 3,580 and 10 microg/g wet tissue, respectively. MRI of the brain showed iron deposition in the basal ganglia, dentate nucleus and thalamus. This case did not show any abnormal increase in copper in the blood and urine following CuSO(4)5H(2)O oral overloading test. Following the intravenous administration of commercially available fresh-frozen human plasma (FFP) containing ceruloplasmin, the serum iron content increased for several hours due to ferroxidase activity of ceruloplasmin. In the liver, the iron content decreased more with the combined intravenous administration of FFP and deferoxamine than with FFP administration alone. Her neurological symptoms improved following repetitive FFP treatment. PMID- 10529543 TI - Decrease in the ciliary neurotrophic factor of the spinal cord in amyotrophic lateral sclerosis. AB - Ciliary neurotrophic factor (CNTF), a potent survival factor in spinal motoneurons of embryonic chick and rat, is currently being investigated in humans as a treatment for amyotrophic lateral sclerosis (ALS). However, its physiological and pathological activities in ALS remain unclear. We measured CNTF contents in the cervical enlargement of the spinal cord from 9 ALS patients and 12 age-matched control subjects using a sensitive enzyme-linked immunoassay. CNTF genotypes were determined by the polymerase chain reaction-restriction fragment length polymorphism method. In control subjects, there were 8 homozygotes and 4 heterozygotes, while in ALS patients there were 6 and 3, respectively. In both homozygotes and heterozygotes, CNTF expression in the spinal cord from ALS patients tended to decrease compared to control subjects. In homozygotes, the decrease was significant (p < 0.05). Concerning the regional concentrations of CNTF, in homozygotes, CNTF contents in the lateral corticospinal tract were markedly lower (p < 0.001) in ALS patients than in controls. The decrease in CNTF expression in the lateral corticospinal tract of the spinal cord from ALS patients may be a feature of ALS and could be related to motor neuron loss. PMID- 10529544 TI - Adrenoleukodystrophy associated with vitiligo and ulcerative colitis. AB - Adrenoleukodystrophy (ALD) is an X-linked inherited disorder of lipid metabolism usually presenting in childhood or early adolescence. It is a progressive disease with symptoms of adrenal insufficiency and central nervous system demyelination. The pathology results from the accumulation of very long-chain fatty acids and an inflammatory reaction in the brain white matter. We report a case of ALD associated with adrenal insufficiency and two autoimmune diseases: vitiligo and ulcerative colitis. PMID- 10529545 TI - Comparison of rizatriptan 10 mg vs. naratriptan 2.5 mg in migraine. AB - Rizatriptan (MAXALT(TM), Merck & Co., Inc.) is a selective 5-HT(1B/1D) receptor agonist with rapid oral absorption and early onset of action for the acute treatment of migraine. This randomized, double-masked, double-dummy, placebo controlled study compared rizatriptan 10 mg to naratriptan (NARAMIG(TM), AMERGE(TM), both Glaxo Wellcome plc) 2.5 mg in 522 patients treating a single migraine attack. Rizatriptan was more effective than naratriptan. Rizatriptan provided earlier headache relief than naratriptan (hazard ratio 1.62, p < 0.001), acting as early as 30 min. More patients were pain free at 2 h on rizatriptan than on naratriptan (44.8 vs. 20.7%, p < 0.001). Rizatriptan also provided earlier relief of associated migraine symptoms within 2 h than naratriptan and more patients had normal function at 2 h (39.3 vs. 22.6%, p < 0. 001). Both active treatments were effective compared to placebo. Both active treatments were well tolerated. The most common side effects with rizatriptan were dizziness, asthenia/fatigue, nausea and somnolence, while the most common side effects with naratriptan were dizziness and asthenia/fatigue. PMID- 10529546 TI - Postural tremor after thalamic infarction. PMID- 10529547 TI - Detection of HCV-specific sequences in chronic myopathy with hepatitis C: improvement with interferon-alpha 2A therapy. PMID- 10529548 TI - Apolipoprotein E epsilon4 allele is a risk factor for Alzheimer's disease: the central region of portugal (Coimbra) as a case study. PMID- 10529549 TI - Traditional African medicines complicate the management of febrile seizures. PMID- 10529551 TI - Hepatorenal interaction in glutathione redox state during partial hepatic ischemia-reperfusion in rats. AB - The hepatic glutathione metabolism during partial hepatic ischemia-reperfusion was studied in a rat model with special reference to the hepatorenal glutathione metabolism system. Bile samples, and in- and outflow blood and tissue samples of the rat liver, kidney and small intestine were collected during 60 min of 70% partial liver ischemia and reperfusion. Both reduced and oxidized glutathione levels were determined by HPLC. The tissue ratio of reduced to oxidized glutathione (GSH/GSSG ratio) decreased significantly in the ischemic hepatic lobe at 60 min after reperfusion. The GSH/GSSG ratio in bile from the ischemic hepatic lobes decreased significantly after 60 min of ischemia and gradually recovered after reperfusion. The net release of GSH from the nonischemic hepatic lobe increased at 60 min after reperfusion, since the calculated net release of GSH from the whole liver increased significantly whereas there was no change in the net release from the ischemic hepatic lobe. The tissue GSH level in the kidney increased significantly at 180 min after reperfusion. The calculated net uptake of GSH into the kidney, and the net release of total cysteine from the kidney, tended to decrease at the end of 60 min of ischemia, to increase at 60 min after reperfusion and then decrease at 180 min after reperfusion. We found that the hepatorenal glutathione metabolism was changed by partial hepatic ischemia reperfusion. These changes might reflect a hepatorenal interaction to maintain the glutathione redox state of the vital organs. PMID- 10529550 TI - Selective ablation of porcine and rabbit liver tissue using radiofrequency: preclinical study. AB - Temperature changes and their distribution induced by 13.56-MHz radiofrequency (RF) heating of agar phantom and porcine and rabbit liver were investigated. It was possible to produce selective local heating of approximately 50 degrees C in the RF field of 2 x 2 x 2 cm(3) of the pig or rabbit liver. Coagulation necrosis after heating became pronounced and the margin between the coagulated lesion and normal tissue became clearer with time. Within 1 week after RF heating at 50 degrees C for 20 min, the coagulated area was replaced selectively and totally by necrotic tissue. PMID- 10529552 TI - Prevention of reactive oxygen-induced endothelial cell injury by blocking its process. AB - Endothelial cell (EC) injury induced by reactive oxygen species (ROS) was investigated and effects of Ca(2+) channel blockers, agents which elevate intracellular cAMP levels ([cAMP](i)), and protein kinase inhibitors on H(2)O(2) induced EC injury were analyzed using human umbilical vein EC cultures. Exposure to H(2)O(2) increased intracellular Ca(2+) levels and decreased [cAMP](i). Ca(2+) channel blockers, [cAMP](i)-elevating agents, and protein kinase inhibitors significantly inhibited H(2)O(2)-induced EC injury. Data suggest that H(2)O(2) induced EC injury is mediated by extracellular Ca(2+) influx, intracellular cAMP efflux, and intracellular signaling, each of which is blocked by Ca(2+) channel blockers, [cAMP](i)-elevating agents, or protein kinase inhibitors. It is suggested that ischemia/reperfusion injury induced by ROS may be prevented by Ca(2+) channel blockers, [cAMP](i)-elevating agents, and protein kinase inhibitors. PMID- 10529553 TI - Augmentative effect of cyclosporin A on rat liver regeneration: influence on hepatocyte growth factor and transforming growth factor-beta(1). AB - We investigated the effect of cyclosporin A (CsA) on rat liver regeneration following partial hepatectomy with reference to cytokine production. Rats were divided into two groups: those without CsA pretreatment (group 1) and those with CsA pretreatment (group 2). Animals were given olive oil vehicle or CsA (10 mg/kg) dissolved in olive oil daily by gavage from 4 to 1 days before hepatectomy. The ratio of regenerating liver weight to initial body weight in group 2 was significantly higher than that in group 1 at 72 h. Although a peak 5 bromo-2-deoxyuridine labeling index was found at 24 h after hepatectomy in both groups, the peak value in the CsA-treated animals was significantly higher than in controls. In both groups, hepatocyte growth factor concentrations in both plasma and liver tissue showed maximal values at 12 h. Liver tissue values in group 2, however, were significantly higher from 1 to 12 h compared to group 1. Transforming growth factor-beta(1) (TGF-beta(1)) concentrations showed minimal serial changes in group 1, while those in liver tissue of group 2 rats were significantly lower than in group 1. Plasma TGF-beta(1) concentrations did not differ. These results suggest that upregulation of hepatic regeneration with CsA pretreatment might be attributed in part to changes in production of these mitogenic and mitoinhibitory cytokines. PMID- 10529554 TI - Inhibitory effect of a gastrin receptor antagonist, CR2945, on 1, 2 dimethylhydrazine-induced colorectal cancer in mice. AB - This study tested the effect of a new gastrin receptor antagonist, CR2945, on colorectal cancer induced by 1,2-dimethylhydrazine (DMH) in mice. 75 CD1 male mice were divided into 3 groups: group 1 received 1 weekly injection of 20 mg/kg of DMH and 2 daily intraperitoneal injections of 0.5 ml of NaCl 0.9% solution for 5 weeks; groups 2 and 3 received the same weekly dose of DMH and 2 daily injections of CR2945 at the respective doses of 2.5 and 7.5 mg/kg for 5 weeks. The animals were sacrificed 25 and 38 weeks after the first injection. No tumours were found at the 25th week. A lower cancer frequency (4%) was observed in treated animals compared to controls (37.4%) at the 38th week (p = 0.002). These data show that CR2945 could prevent chemically induced colon cancer development in mice. PMID- 10529555 TI - Pancreatic regeneration after subtotal distal resection in rats. Effects of bombesin and octreotide by the in vivo bromodeoxyuridine uptake technique. AB - Forty male Wistar rats were randomly allocated to four treatment groups after 90% distal pancreatectomy: group A (control) received saline (0.5 ml subcutaneously); group B received bombesin (BBS; 10 microg/kg intraperitoneally); group C received octreotide (2.5 microg/kg subcutaneously), and group D received BBS and octreotide. All substances were injected three times a day until sacrifice after 28 days. BBS increased pancreas weight (p = 0.003) and DNA synthesis (p < 0.001), as measured by a bromodeoxyuridine nuclear-labeling index (BrdU LI). The simultaneous administration of octreotide significantly decreases the remnant pancreas weight (p = 0.016) as compared to group B rats; however the BrdU LI is not significantly reduced in group D as compared to group B. BBS administration promotes regeneration of the remnant pancreas in terms of hypertrophy and hyperplasia. Although octreotide appears to significantly reduce the pancreatic weight increase induced by BBS, it does not reduce DNA synthesis and cell proliferation. PMID- 10529556 TI - Effect of experimental diabetes and growth hormone administration on the strength of colonic anastomoses in rats. AB - The influence of growth hormone (GH; 2 mg/kg/day) administration on the mechanical breaking strength of colonic anastomoses in diabetic rats has been investigated on the day of operation (suture binding capacity) and after 4 and 7 days of healing. In diabetic rats, the suture binding capacity was decreased by 26% in both ultimate load and relative failure energy. After 4 days of healing, no difference was observed between control and diabetic animals. After 7 days, relative failure energy in the diabetic animals was reduced by 33%. GH administration to diabetic animals did not alter strength during the first week of healing. We found an increased circumference (33%) and defatted dry weight (22%) of the colon in diabetic rats. In conclusion, diabetes impairs the suture binding capacity of the colon in rats, while there is only little influence on healing in the following week. GH administration could not influence the strength of colonic anastomoses in diabetic animals. PMID- 10529557 TI - Donor lung pretreatment with prostaglandin E(1) does not improve lung graft preservation. AB - Prostaglandin E(1) (PGE(1)) is widely used to improve early graft function after lung transplantation, but some studies have questioned its benefits. Therefore we evaluated the effect of donor pretreatment with PGE(1) in our porcine model of single lung transplantation. Donors received PGE(1) or placebo intravenously before flushing the pulmonary artery with modified Euro-Collins solution. After cold storage, the excised left lung was transplanted. Ischemic time was 4 h. We used our right side heart bypass model to measure standardized pulmonary vascular resistance and to study blood flow distribution between recipient's native and transplanted lung. Systemic and pulmonary hemodynamics and gas exchange were also measured. After transplantation, pulmonary vascular resistance was significantly higher in the transplanted lung, which received only one fourth of the total pulmonary blood flow. PGE(1) pretreatment did not improve pulmonary hemodynamic parameters, or gas exchange. PMID- 10529558 TI - 10th Walter Brendel Symposium on Applied Immunology and Microcirculation. Mauls, Sudtirol, Italy, January 30 to February 4, 1999. Abstracts. PMID- 10529559 TI - The natural history of early prostate cancer and the impact of endocrine treatment. AB - The best approximation of the natural course of early prostate cancer comes from studies on deferred treatment. In such studies, the 10-year disease-specific survival rate is 85-90% for patients with clinically localized tumours and 74% for those with tumours not confined to the prostate gland; 15-year data are sparse and diverging. Patients with poorly differentiated tumours have a worse prognosis than those with well or moderately differentiated tumours. Endocrine treatment of early prostate cancer is a revitalized concept, with the majority of the evidence for a beneficial effect coming from uncontrolled studies. Recently, however, data from two randomized studies have suggested that endocrine treatment may prolong survival in patients with non-metastatic prostate cancer. Also, in studies of radiotherapy for localized prostate cancer, a longer survival has been seen in patients randomized to neoadjuvant or adjuvant endocrine treatment compared with placebo. The current trend for earlier treatment means that the total time on endocrine therapy has increased and patients are younger when treatment is initiated. Therefore, randomized studies should not only substantiate a possible survival benefit of endocrine treatment, but also assess potential long-term side effects and their impact on quality of life. PMID- 10529560 TI - Early versus deferred hormone therapy. AB - Although hormone therapy is widely used in the management of prostate cancer, the optimal timing of its initiation remains a matter of debate. Immediate hormone treatment has been compared with deferred treatment in randomized studies conducted by the Veterans Administration Cooperative Urological Research Group, the South Sweden Prostate Cancer Study Group and the Medical Research Council. Despite criticism of the design of these studies, the results indicate that early treatment may be associated with advantages in time to progression and survival. It is anticipated that ongoing studies, such as the European Organization for Research and Treatment of Cancer protocols 30846 and 30891, will provide further information on the optimal timing of endocrine treatment. Prognostic and quality of life factors also have an impact on the treatment decision. On the basis of available evidence, early hormone therapy is recommended for younger men with poorly differentiated tumours or advanced disease and for those who are seen infrequently by their physician. Deferred treatment using a strategy of watchful waiting is probably the best option for older men with well differentiated, low volume prostate cancer. PMID- 10529561 TI - A review of studies of hormonal adjuvant therapy in prostate cancer. AB - There is increasing interest in the use of adjuvant hormonal therapies, which are given after the resection or destruction of all gross disease, in early-stage prostate cancer, as a significant proportion of patients experience progression and/or die from the disease despite undergoing therapy with curative intent. Several retrospective studies suggest that adjuvant hormonal therapy may improve long-term outcome after radical surgery in men with positive lymph nodes, although this approach has yet to be studied in a prospective setting. No studies of adjuvant therapy for patients with extracapsular extension at surgery have been completed, but in an interim analysis of an open controlled trial, adjuvant flutamide significantly improved progression-free survival at 4 years. Three prospective studies in the radiotherapy setting have shown that adjuvant luteinizing hormone-releasing hormone (LH-RH) agonist therapy significantly improves progression-free and/or overall survival. Future studies need to define patient subgroups who will benefit most from adjuvant therapy. The side effects of the different therapeutic options also need to be compared. It is hoped that many of the outstanding questions concerning adjuvant hormonal therapy will be answered by the ongoing Bicalutamide Early Prostate Cancer Programme. PMID- 10529562 TI - Quality of life issues relating to endocrine treatment options. AB - Recent interest has focused on the use of hormone therapy in prostate cancer for both the management of patients with non-metastatic disease and as a neoadjuvant or adjuvant to curative therapies. This has resulted in patients with fewer symptoms being treated for longer periods of time. Endocrine treatments for prostate cancer, such as castration, combined androgen blockade and non-steroidal antiandrogen monotherapy, have shown similar results in terms of time to progression and survival. The main difference between these treatments is their impact on patients' quality of life. Instruments for measuring health-related quality of life should assess both overall and disease-specific quality of life. Data from two large studies of bicalutamide monotherapy show that this non steroidal antiandrogen is associated with significant health-related quality of life advantages in the treatment of patients with locally advanced (M0) disease compared with castration, suggesting that this treatment may benefit patients with early disease. Bicalutamide was favoured in 8 out of 9 evaluable quality of life dimensions, and this was statistically significant for sexual interest and physical capacity. Endocrine treatments with minimal adverse effects on quality of life will be increasingly favoured for patients with non-metastatic disease who are being treated for longer periods of time. PMID- 10529563 TI - The future of endocrine treatment in early prostate cancer: concluding remarks. AB - Prostate cancer screening, involving markers such as prostate-specific antigen, has enabled the detection and diagnosis of prostate cancer much earlier in life and at an earlier stage of disease. This earlier diagnosis, with the potential for earlier and longer endocrine exposure, has implications on treatment options for early prostate cancer. Treatment options that need to be considered are immediate versus deferred treatment, adjuvant therapy, intermittent treatment and step-up treatment. Preliminary reports indicate advantages for immediate compared with deferred therapy in terms of time to progression and overall survival. There are studies in progress for the other treatment options that should resolve some of the uncertainties about the management of early prostate cancer. The need to balance long-term side effects against potential survival advantage is of particular importance in patients who may be receiving endocrine therapy for longer periods. As antiandrogens are associated with quality of life benefits when compared with castration, the non-steroidal antiandrogen bicalutamide, which is currently being investigated as monotherapy(1) and as adjuvant therapy, may prove to be a useful treatment option for patients with early prostate cancer. PMID- 10529565 TI - Extrasystoles in the fetal CTG during labour might be a first sign of fetal neonatal sepsis. AB - A case of extrasystoles in the fetal electrocardiogram during labour is presented as a possible early sign of fetal-neonatal sepsis. Colleagues are invited to respond similar experience. PMID- 10529564 TI - A rare cause of fetal ascites: A case report of Gunther's disease. AB - Despite an arsenal of ever-improving diagnostic tools, determining the precise etiology of fetal ascites is not always possible. We report a case history where moderately-severe fetal ascites was retrospectively determined to be due to Gunther's disease (congenital erythropoietic porphyria). The infant was found to carry the mutation associated with the most severe disease phenotype in which fetal hydrops has been described. PMID- 10529566 TI - Management of acute chylothorax with hydrops fetalis diagnosed in the third trimester of pregnancy. AB - A fetus with large pleural effusion and hydrops fetalis diagnosed in the third trimester was successfully treated with prompt vaginal delivery followed by drainage of the pleural cavity, after confirmation of congenital chylothorax and re-expansion of the lung with prenatal thoracentesis. PMID- 10529567 TI - Ontogeny of isolated ultrasound markers for fetal aneuploidy. AB - OBJECTIVE: To evaluate the natural history of isolated ultrasound markers for fetal aneuploidy observed at 14-16 weeks of affected gestations. STUDY DESIGN: 76 aneuploid gestations were diagnosed among a predominantly low-risk population undergoing targeted ultrasonography in the early second trimester. Indications for evaluation of fetal karyotype included fetal malformations or sonographic markers for aneuploidy, maternal age over 35 years and abnormal serum screening results. Markers were re-evaluated at the time of amniocentesis or at pregnancy termination for fetal anomalies. RESULTS: Sonographic markers for aneuploidy (SMA) were observed in 68 of 76 aneuploid gestations diagnosed in the study group. In 46 cases, SMA were associated with major fetal malformations and in 22 cases, markers were isolated. Only 2 of 22 isolated markers for aneuploidy were documented at the time of amniocentesis. CONCLUSION: Isolated ultrasound markers for aneuploidy are transient and may disappear later on in gestation. Transvaginal sonography at 14-16 weeks of gestation appears to provide the best time window for detection of markers for fetal abnormal karyotype. PMID- 10529568 TI - Recombinant erythropoietin in the prevention of late anaemia in intrauterine transfused neonates with Rh-haemolytic disease. AB - OBJECTIVE: To evaluate the efficacy of recombinant human erythropoietin (rHuEPO) in prevention of late anaemia due to Rh-haemolytic disease in neonates subjected to one or more intrauterine transfusions (IUTs). STUDY DESIGN: Six neonates (GA 28-38 weeks, BW 980-3,360 g), subjected to one or more IUTs for Rh-haemolytic disease, were treated for 3 weeks with rHuEPO (200 U/kg/day, s.c.) after the second week of life to prevent late anaemia and consequently reduce the need for blood transfusions. All treated neonates were supplemented weekly with iron, vitamin E and folinic acid, intramuscularly. RESULTS: Of the 6 patients studied, 4 preterm neonates, after commencement of rHuEPO treatment, showed a decrease in Hct values with persistent reticulocytopenia, and consequent need for one or more transfusions with packed and filtered red cells (PFRC). These 4 neonates had received a greater blood volume with IUTs than the 2 other term neonates, who, after starting rHuEPO treatment, showed an increase in Hct values and in reticulocyte count, with no transfusion requirements after birth (247 +/- 47 vs. 84 +/- 76 ml). CONCLUSIONS: Our results seem to correlate the efficacy of erythropoietin treatment in prevention of late anaemia resulting from Rh haemolytic disease to the severity of intrauterine anaemia and to gestational age. Erythropoietin, in fact, was less effective in cases of severe intrauterine anaemia requiring a high volume of PFRC; it was also less effective in the preterm babies, because of the simultaneous presence of anaemia of prematurity and other major diseases. PMID- 10529569 TI - Idiopathic intracranial haemorrhage in the fetus. AB - Intracranial haemorrhage in the fetus has been reported with associated mortality and morbidity. This case report describes idiopathic subdural haematomas diagnosed at 32 weeks of gestation, with delivery by caesarean section of a live male infant in good condition at 34 weeks. PMID- 10529570 TI - Amnioreduction: how much to drain? AB - OBJECTIVES: To assess the relationship between the volume of amniotic fluid removed and the change in amniotic fluid index (AFI) and calculate an equation describing this association. MATERIALS AND METHODS: A retrospective analysis of 19 amnioreduction procedures performed in our unit. Multiple regression analysis was used to assess the effect of gestational age and pre-procedure AFI on the change in AFI (DeltaAFI) after adjusting for the volume removed. RESULTS: As expected, a significant linear relationship was found between the change in AFI and the volume removed (r = 0.82, n = 19, p < 0.0001). DeltaAFI was not dependent on the gestational age or the pre-procedure AFI. The equation describing the association between the volume removed and DeltaAFI was: volume = (DeltaAFI - 2. 26)/0.008, which is close to 1 cm DeltaAFI for every 100 ml removed. DISCUSSION: Using the described equation, it is possible to predict the required volume to be removed in order to achieve a particular DeltaAFI, which may reduce the need to interrupt the procedure to measure the AFI. However, the limitation of AFI as a semiquantitative assessment of the liquor volume, together with its inter- and intra-observer variations mean this equation should be used only as a guide. PMID- 10529571 TI - Twenty-four cordocenteses in one woman. AB - We report on 2 consecutive pregnancies in a woman with a history of neonatal death secondary to Rhesus alloimmunization. Her first subsequent pregnancy was complicated by fetal hydrops at 20 weeks of gestation. The fetus received a total of 11 intrauterine transfusions, and was delivered at 38 weeks. In the patient's next pregnancy, the fetus developed hydrops at 18 weeks of gestation. Thirteen intrauterine transfusions were given to correct fetal anemia, and a healthy baby was delivered at 38 weeks of gestational age. Continuation of intravascular transfusion therapy may represent a reasonable alternative to selective premature delivery even in cases with highly aggressive maternal Rhesus alloimmunization. PMID- 10529572 TI - Unexplained elevated serum hCG is associated with raised amniotic fluid erythropoietin levels in second-trimester pregnancies. AB - OBJECTIVE: This study was designed to investigate the association between the concentrations of maternal serum hCG and amniotic fluid erythropoietin during the second trimester of pregnancy. METHODS: In a prospective case-control study, 42 consecutive singleton pregnancies showing unexplained elevated serum hCG concentrations (>2.0 multiples of the median, MoM) in Down's syndrome screening and 27 control pregnant women undergoing midtrimester amniocentesis because of a previous cytogenetic abnormality were studied. RESULTS: The mean amniotic fluid erythropoietin concentration in the study group was 1.8 (range 0.61-8.7) MoM, whereas it was 1.1 (range 0.71-3. 96) MoM in the controls (p = 0.035). A significantly increasing relationship (p < 0.05) was found between the concentrations of maternal serum hCG and amniotic fluid erythropoietin. CONCLUSIONS: The results of the current study revealed in vivo the association between elevated hCG and amniotic fluid erythropoietin levels which, in turn, supports the concept of early placental damage. The underlying pathology seems to be sufficient to cause an erythroblastic response. PMID- 10529573 TI - Improved specificity of NRBC detection in chorionic villus sample supernatant fluids using anti-zeta and anti-epsilon monoclonal antibodies. AB - OBJECTIVE: Fetal erythrocytes leak from fetal capillaries at the time of chorionic villus sampling (CVS). It has been reported that in approximately 60% of CVS cases fetal nucleated red blood cells (NRBC) can be isolated from the supernatant fluid by immunophenotyping with monoclonal antibody (Ab) against the gamma-chain of fetal hemoglobin and used as an additional source for confirmation of the fetal karyotype. However, the increased prevalence of beta-thalassemia mutations in countries such as Greece results in many pregnant women who produce gamma-positive cells. This makes it difficult to distinguish between the fetal and maternal origin of the NRBC. Use of Abs against embryonic hemoglobin chains zeta and epsilon may increase specificity for fetal NRBC detection. METHODS: Mouse monoclonal Abs against Hb-zeta and Hb-epsilon were used in order to examine if specificity for fetal NRBC detection in CVS supernatant fluids could be improved. 41 samples were studied using anti-zeta and 20 using anti-epsilon monoclonal Abs. RESULTS: Anti-zeta or anti-epsilon positive erythrocytes were, respectively, identified in 52 of 61 CVS samples and anti-zeta or anti-epsilon positive NRBC were present in all cases. The mean number of Hb-positive erythrocytes identified with the anti-zeta Ab was 58 and the mean number of NRBC 29. The mean number of anti-epsilon positive erythrocytes was 30 and of NRBC 23. FISH with X and Y chromosome specific probes was performed in 26 cases and the results were concordant with the CVS karyotype. Statistical analysis using the correlation test showed that anti-zeta and anti-epsilon were more specific for the detection of embryonic NRBCs. CONCLUSIONS: Since embryonic monoclonal Abs show increased specificity, they should be preferentially used for NRBC detection in CVS supernatant fluids. Furthermore, the increased specificity of anti-zeta and anti-epsilon Abs may considerably improve prenatal diagnosis from fetal cells isolated from maternal circulation. PMID- 10529574 TI - Lung development in diamniotic twins discordant for complete urinary tract obstruction. AB - OBJECTIVES: To investigate the effect of anhydramnios on the lung development of 1 twin in the presence of a normal amniotic fluid volume in its diamniotic co twin. METHODS: Three sets of diamniotic twins, discordant for complete urinary tract obstruction and anhydramnios, were followed prospectively with regular ultrasound scans and after delivery. RESULTS: All 3 twins with complete urinary tract obstruction and anhydramnios died within 2 days after birth, with confirmed severe pulmonary hypoplasia. In every case the twin with a normal amount of surrounding amniotic fluid had a normal postnatal outcome. CONCLUSIONS: The observation that a normal amniotic fluid volume in one sac does not protect the anhydramniotic twin from pulmonary hypoplasia has important implications for the aetiology of the condition and for the possibility of therapeutic septostomy. These results are discussed in relation to previous human and animal studies. PMID- 10529575 TI - Prenatal diagnosis of a congenital bladder diverticulum. Case report and benefits of prenatal diagnosis. AB - A case of congenital bladder diverticulum diagnosed at 37 weeks of gestation (measured from the first day after the last day of the last menstrual period) is reported. Delivery took place 24 h later. A postnatal urologic work-up confirmed the diagnosis of asymptomatic congenital bladder diverticulum. The infant underwent laparotomic surgery at the age of 6 months, with an extravesical diverticulectomy and ureteral reimplantation. There were no complications. This is the first case reported in the literature of a prenatal diagnosis of a congenital bladder diverticulum. This new aspect allows early management and avoidance of the diagnostic meanders to which the discovery of a pelvic mass might lead, as well as the complications that can follow bladder diverticula. PMID- 10529576 TI - Diagnosis of arachnoid cysts on prenatal ultrasound. AB - The aetiology and physiology of congenital arachnoid cysts are a source of controversy. We report a case where fetal cerebral ultrasonography shows an extraventricular sonolucent cystic formation after 20 weeks of pregnancy. Ultrasonography provides its topographic relations with adjacent brain structures and is also used to diagnose possible associated malformations. MRI confirms the ultrasonographic findings by investigating cerebral gyri. The rest of the examination involves detection of extracerebral anomalies and a karyotype study. Other differential diagnoses will be considered as a function of the embryological origin and topography of arachnoid cysts. The outcome of these arachnoid cysts depends on the age at the time of diagnosis, their size and their topography. The problem is that hydrocephalus, due to compression of the cerebrospinal fluid drainage pathways, may develop. Treatment, if necessary, is nearly always surgical. PMID- 10529577 TI - Prenatal ultrasound diagnosis of a femur-fibula-ulna complex during the first half of pregnancy. AB - A case of fetal femur-fibula-ulna (FFU) complex diagnosed by ultrasound is presented. Ultrasonographic features of a fetus displaying bilateral femoral hypoplasia, aplasia of the right forearm and the right hand, ray defects of the left hand are described. The importance of an early diagnosis of this malformation is emphasized with respect to parental counselling concerning prognosis and further prenatal management. PMID- 10529578 TI - [Mistletoe therapy from the pharmacologic perspective]. AB - Because of their cytostatic/apoptotic and immunomodulatory effects mistletoe extracts are often applied in tumour patients. Recent experimental data suggest that the mistletoe lectins Viscum album agglutinin (VAA)-I and -II are play an important role in the efficacy of mistletoe therapy. VAA-I and -II are members of the type-II ribosome-inactivating proteins. VAA-I has been shown to induce cytostatic effects in cultures of various eukaryotic cells in vitro. In 24-hour cultures of human peripheral lymphocytes, flow-cytometric investigations with propidium iodide (PI) in hypotonic buffer and quantitative assessment of DNA breaks with terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-digoxigenin nick end-labeling (TUNEL) assay were carried out; they revealed a dose-dependent VAA-I-induced apoptosis (lectin concentrations between 10 ng/ml and 1 microg/ml). In 24-hour cultures of peripheral blood mononuclear cells (PBMC), VAA-I in non cytotoxic concentrations (1+/-10 ng/ml) induced mRNA expression and enhanced the secretion of proinflammatory cytokines. The stimulation of NK cells by VAA-I in vitro was enhanced in additive manner by the combination of VAA-I with IL-2 and IL-12. In culture of PBMC and bone marrow CD34+ cells coincubation of VAA-I with other haematopoietic growth factors induced a dose-dependent increase in clonogenic growth. In cancer patients the mechanisms of natural immunity, believed to be essential for their survival, are often significantly decreased. VAA-I and standardized mistletoe extracts are able to stimulate the cellular parameters of natural immunity with a bell-shaped curve of efficacy. Studies in animal models suggest that application of 0.5-3 ng/kg VAA-I twice a week is effective in sustaining the elevation of the number and activity of peripheral blood NK cells. These parameters often exhibit high intrinsic fluctuations, in healthy persons, however, blind crossover studies reveal an optimal lectin dose of about 0.5 and 1 ng/kg bw, suggesting a potential use of mistletoe preparations as a modulator of the natural immune system. Selective apoptotic effects of VAA-I may represent a novel approach for pharmacological manipulation of the balance between cell growth and programmed cell death. Appropriate combination of immunomodulatory and cytotoxic doses may open new clinical perspectives in the mistletoe therapy. PMID- 10529579 TI - [Development of lymphocyte subsets in tumor patients after subcutaneous administration of mistletoe extracts]. AB - OBJECTIVE: In order to exclude the possibility that mistletoe therapy may result in immunosuppression, as indicated by a significant reduction of defined lymphocyte subsets, PATIENTS AND METHODS: peripheral blood cells of 23 tumour patients were treated subcutaneously with increasing concentrations of aqueous mistletoe extracts (Helixor(R)). RESULTS AND CONCLUSIONS: Within an observation period of 7 months, the relative amount of lymphocytes and the number of natural killer (NK) cells increased while the number of lymphocyte subsets (i. e. CD19+ B cells, CD4+ T helper cells, CD8+ CD28- suppressor cells, CD8+ CD28+ cytotoxic cells) and the proportion of CD25+ (activated) cells within T cells showed a statistically remarkable trend; due to the multiple test problem of statistical evaluation this trend is not allowed to be termed significant. The leucocytes decreased insignificantly within the observation period. However, we were unable to verify a suggested increase of defined lymphocyte subsets within 2-3 months after the onset of mistletoe treatment. Nevertheless, for the parameters CD19+ B cells, CD4+ T helper cells, CD8+ cells, CD8+ CD28+ cytotoxic cells and CD16+/CD56+ NK cells we observed statistically remarkable peaks within die 2nd and 3rd month of therapy, confirming the hypothesis. The responses to the extracts were obviously interindividually different; the immune responses especially of patients with a lower number of peripheral T cells were less significant as compared to those of patients with adequate T cell numbers. Surprisingly, even an increase of the drug concentration >3 ng mistletoe lectin (as determined within the whole plant extract) per kg body weight enhanced the number of CD4+ T helper cells. A decreased immunological reaction on mistletoe extracts was shown especially for patients with a reduced number of peripheral T cells, whereas patients with normal T-cell number were more reactive. PMID- 10529580 TI - [Effects of Kneippism, a standardized complex therapy, on pain, quality of life and use of medicines: cohort study with a one-year follow-up]. AB - BACKGROUND: Inpatient as well as outpatient cure in a spa environment with commonly 3- to 4-week duration features a combination of different treatments customized according to the needs of the individual patient. The physiological rationale and the mode of action are widely accepted. However, firm quantitative evidence of the clinical effectiveness is incomplete. OBJECTIVE: To document the effects of a well standardized complex therapeutic regimen (Kneippism) on pain, quality of life, and drug consumption during therapy and with 1-year follow up. STUDY DESIGN: Prospective cohort study with assessments at the beginning, during, and at the end of treatment and with follow-up investigations 3, 6, and 12 months thereafter. SETTING: Four spa clinics in Bad Worishofen, Southern Bavaria. PATIENTS: 363 patients (248 outpatients, mean duration of therapy 23.3 days, and 115 inpatients, mean duration of therapy 27.4 days), one half between 40 and 60 years old above 60 years of age, predominantly suffering from musculoskeletal and/or cardiovascular diseases. INTERVENTION: Custom-tailored combination of therapies comprising of hydro-, kinesi-, and phytotherapy, dietetics, 'ordnungstherapie', and continued disease-specific standard treatment, if necessary. MAIN OUTCOME MEASURE: Pain, patients' self-rating, (IRES questionnaire), medication. RESULTS: The monitored dimensions of pain improved significantly during treatment and remained at that level essentially for the complete follow up interval. The same was true for various dimensions of reported subjective complaints as well as for drug consumption. CONCLUSION: When estimating the clinical relevance of a complex therapeutic regimen such as a cure of 3- to 4-week duration, the question of the impact of the specific effect of single components is secondary to the question of the overall relevance of that therapeutic concept. The findings of this study point at potential long-term effects of at least 1-year duration. PMID- 10529581 TI - [Changes of short-time heart rate variability during hyperthermia treatment with infrared A whole body irradiation]. AB - BACKGROUND: Measures of heart rate variability (HRV) are widely recognized as being an important indicator method for autonomical function and cardiovascular neural regulation. The low-frequency component in spectral analysis is increased during conditions of raised sympathetic activation, the high-frequency component is closely correlated with vagal activity. Whole-body infrared-A irradiation (WBIAI) according to von Ardenne is a newly developed version of hyperthermia. In clinical use peripheral vasodilatation with significant increases in heart rate and hypotension were noted as acute effects of WBIAI. In order to evaluate the effect of WBIAI on autonomic function, we measured HRV during serial hyperthermias. PATIENTS AND METHODS: Power spectral density analysis of HRV during the first and the last of 4 serial WBIAI treatments with 30-60 min endurance was performed in 10 patients with chronic pain disorders. Tympanal body temperature was recorded continuously and was supposed to increase by 1 degrees C during treatment. Electrocardiographic monitoring was performed using a Marquette HoIter ECG. RR data were manually edited and standard frequency domain measures of the first and the last 5 min of treatment computed. The subjective efficacy of the treatment was rated by prestandardized interviews. RESULTS: Body temperature increased by 0.9 degrees C during the first treatment and 1.2 degrees C during the last treatment. In all patients the rise in body temperature due to treatment was accompanied by a significant increase in heart rate and a decrease of arterial blood pressure. During hyperthermia a slight decrease of absolute low frequency and total power as well as a sharp decrease of high-frequency power and a significant increase of the low/high-frequency ratio could be noted. The rise in low/high-frequency ratio during WBIAI was less distinct at the last treatment compared to the initial treatment. Hyperthermia was regarded as a safe, well tolerable and effective treatment. CONCLUSION: The results suggest that the cardiovascular response during WBIAI is accompanied by significant changes in autonomic cardiac regulation: A significant decrease of low-frequency power corresponding to depressed vagal activity results in an increase of Iow/high frequency ratio. During serial hyperthermias the acute response is diminished suggesting an adaption of the autonomic response to hyperthermia. Further studies are necessary to investigate the long-standing autonomic effects of the treatment and include analysis of influencing factors such as the level of physical activity and constitutional individual conditions. PMID- 10529582 TI - [Quality aspects of the use of complementary medicine--experiences from the work of a quality circle]. AB - OBJECTIVES: This article reports the results of a quality circle work, carried out by physicians using complementary medicine. It illustrates the problems occurring in this subject and summarizes characteristics of the daily work in relation to quality aspects. Patient characteristics were investigated; examples of mistakes in daily care are reported. METHODS: The report is based on the two moderators' subjective impressions, on analysis of attendance lists, case reports, protocols, and surveys. RESULTS: The quality circle was started in October 1994 and consists of a group of about 16 physicians, mainly general practitioners. Several objectives mentioned in official guidelines for quality circles could not be achieved, for example, the evaluation of results, analysis and assessment of the own performance according to established criteria, the degree to which published guidelines were followed, the development and implementation of problem-solving strategies. The quality circle was mainly a platform for exchange and continuing medical education. It also was the basis for an empirical study. CONCLUSION: The continuing interest of patients in complementary medicine should motivate physicians to systematically evaluate these therapies. Participants of quality circles should prepare themselves extensively for the meetings. They should take measures to keep up-to-date with the current medical knowledge. Elements and objectives aiming at a higher quality standard can be examined and implemented in routine practice work. Research results should be integrated into the daily practice. Furthermore, academic or other independent institutions should carry out experimental research into traditional complementary therapies. PMID- 10529583 TI - Recent advances in mastocytosis research. Summary of the Vienna Mastocytosis Meeting 1998. AB - The term mastocytosis denotes a heterogenous group of disorders characterized by abnormal growth and accumulation of mast cells in one or more organs. Cutaneous and systemic variants of the disease have been described. Mast cell disorders have also been categorized according to other aspects, such as family history, age, course of disease, or presence of a concomitant myeloid neoplasm. However, so far, generally accepted disease criteria are missing. Recently, a number of diagnostic (disease-related) markers have been identified in mastocytosis research. These include the mast cell enzyme tryptase, CD2, and mast cell growth factor receptor c-kit (CD117). Several gain-of-function-mutations in the kinase domain of c-kit appear to occur in mastocytosis supporting the clonal (neoplastic) nature of the disease. Also, certain point mutations appear to be associated with distinct variants of mastocytosis, i.e. Asp-816-->Val with a subset of sporadic persistent (systemic) mastocytosis (mostly adults), and Gly 839-->Lys with (a subset of) typical pediatric (mostly cutaneous) mastocytosis. Another potential indicator of mast cell neoplasm is the T-/NK-cell-associated marker CD2. This antigen (LFA-2) is abnormally expressed on neoplastic mast cells in cases of systemic mastocytosis or mast cell leukemia, but not found on normal mast cells. The mast cell enzyme tryptase is increasingly used as a serum- and immunohistochemical marker to estimate the actual spread of disease (burden of neoplastic mast cells). The clinical significance of novel mastocytosis markers is currently under investigation. First results indicate that they may be useful to define reliable criteria for the delineation of the disease. PMID- 10529584 TI - Role of IL-5 in the development of allergen-induced airway hyperresponsiveness. AB - This review evaluates the role of IL-5 and IL-5-mediated eosinophil airway infiltration in the development of allergen-driven airway hyperresponsiveness. It discusses the structure and function of IL-5 and its receptor and the mechanisms of IL-5-triggered eosinophil accumulation and inflammation of the airways. New research data from murine models of airway inflammation and hyperresponsiveness utilizing different modes of sensitization to allergen and anti-IL-5 antibody or IL-5-deficient knock-out mice underscore the outstanding role of this cytokine in the pathogenesis of bronchial asthma. This review identifies possible directions for future treatment of airway hyperresponsiveness and concludes that targeting IL-5-driven inflammatory responses may be most beneficial for a novel therapy in bronchial asthma. PMID- 10529585 TI - Isoforms of the major allergen of birch pollen induce different immune responses after genetic immunization. AB - BACKGROUND: Recent publications indicate that immunization with plasmid DNA encoding allergens might represent a potential approach in allergen-specific immunotherapy. OBJECTIVE: In the present study we have compared the immune responses induced by plasmid DNA encoding for two isoforms of Bet v 1, the major allergen of birch pollen. METHODS: BALB/c mice were injected intradermally with plasmid DNA encoding for the genes of Bet v 1a (pCMV-Beta) and Bet v 1d (pCMV Betd). In addition, the effect of immunostimulatory DNA sequences was investigated by appending and/or coinjecting CpG motifs. Antibody responses and IFN-gamma and IL-4 levels were measured by ELISA. Allergen-specific proliferation was determined by incorporation of [(3)H]-thymidine. RESULTS: The two isoforms induced a similar humoral response. The lack of any IgE production and the ratio of IgG1 to IgG2a clearly indicated a Th-1-type response. The antisera against both isoforms were highly cross-reactive, which was supported by the energy plot indicating similar folding of the two protein isoforms. However, determination of IFN-gamma and IL-4 in the serum elicited a strikingly different cytokine profile during the course of the immune response. In contrast to pCMV-Beta, pCMV-Betd caused no significant allergen-specific proliferation and induced only marginal levels of the key cytokines. CONCLUSIONS: Based on the assumption that the induction of a strong Th-1 type response is a prerequisite for successful treatment of allergy, our results favor the use of isoform Bet v 1a in combination with CpG motifs for a novel type of allergen immunotherapy based on plasmid DNA immunization. Additionally, the data also confirm the assumption that the antigen itself can have a marked influence on the immune response after genetic immunization. PMID- 10529586 TI - Involvement of carbohydrate epitopes in the IgE response of celery-allergic patients. AB - BACKGROUND: This study was performed to get further insights into antibody responses to cross-reactive carbohydrate determinants (CCD), including initial experiments to prove the biological activity of anti-CCD IgE. Earlier studies have shown that IgE specific for CCD occurs in about 25% of celery-allergic patients. The clinical significance of these antibody specificities is doubtful. METHODS: Patient sera were selected on the basis of a positive case history of celery allergy and multiple binding to high molecular weight celery allergens on immunoblots. Specific IgE to native and heated celery tuber was determined by the enzyme allergosorbent test (EAST). N-glycans were purified after extensive digestion of specific glycoproteins, such as pineapple stem bromelain, bovine fibrin, and human IgG, and used as antigens in an IgE ELISA as well as in EAST and immunoblotting inhibition experiments. Dose-related histamine release was performed with BSA neoglycoproteins containing 3-4 units of the purified glycopeptides. RESULTS: Seven celery-allergic patients were identified who clearly presented IgE against the N-glycan purified from bromelain which is a common structure within the plant kingdom. Chemical defucosylation showed that alpha1, 3-fucose is a key structure for IgE binding. In patients with anti-CCD IgE, the maximal inhibition of celery EAST by the bromelain glycan ranged from 22 to 100%. Inhibition of celery immunoblots by preincubation of patient serum with this glycan led to a quenching of multiple bands at masses >40 kD. After linking the bromelain glycopeptide to BSA, a strong dose-related histamine release was obtained in a celery-allergic patient, occurring at lower concentrations than with the recombinant major protein allergen from celery, Api g 1. CONCLUSIONS: Our results demonstrate that IgE specific for CCD is common in celery-allergic patients, and can represent the major proportion of IgE against this food. alpha1, 3-fucose was confirmed to be an essential part of the IgE epitope. Immunoblotting inhibition indicated the presence of this carbohydrate determinant on multiple glycoproteins in celery extract. Although histamine release was only performed in 1 patient, our data show that proteins carrying multiple glycan units can be biologically active in patients sensitized to CCD. PMID- 10529587 TI - Evaluation of specific IgE to the recombinant group 2 mite allergens Lep d 2 and Tyr p 2 in the Pharmacia CAP system. AB - BACKGROUND: Several recombinant allergens have been shown to be potentially useful for diagnosis of IgE-mediated allergy, but only a few recombinant allergens are at present commercially available in serological assays for detection of specific IgE. The aim of this study was to evaluate the IgE binding to the recombinant major dust mite allergens rLep d 2 and rTyr p 2 and compare it with the IgE binding to the commercial mite extracts Lepidoglyphus destructor and Tyrophagus putrescentiae in the Pharmacia RAST CAP System. METHODS: The recombinant allergens rLep d 2 and rTyr p 2 were immobilised on ImmunoCAPs, and sera from 461 Swedish farmers who are frequently exposed to mites were analysed for specific IgE antibodies. Immunoblotting was performed to evaluate discrepancies between the results obtained with the recombinant and the commercial CAP assays. RESULTS: The IgE values of each recombinant assay significantly correlated with the IgE values of the corresponding commercial CAP assay. The sensitivity of the rLep d 2 assay was 73.3% and that of the rTyr p 2 assay, 60.5% of that provided by the commercial L. destructor and T. putrescentiae assays. Two subjects out of 416, who tested negative in the commercial L. destructor assay, were positive to rLep d 2. The corresponding figures for rTyr p 2 and the T. putrescentiae extract were 5/418. The possibility that these subjects were sensitised to L. destructor and T. putrescentiae could not be excluded. CONCLUSIONS: The data suggest that it may be possible to use rLep d 2 and rTyr p 2 on ImmunoCAPs to detect and quantify IgE antibodies to these, the major allergens of L. destructor and T. putrescentiae. It appears likely that the addition of just a few more recombinant L. destructor and T. putrescentiae allergens in the CAP assay will be sufficient for in vitro diagnosis of IgE mediated allergy to L. destructor and T. putrescentiae. PMID- 10529588 TI - A repeat polymorphism in interleukin-4 gene is highly associated with specific clinical phenotypes of asthma. AB - BACKGROUND: IL-4 is a determining factor in immunologic mechanisms to allergy and inflammation. The authors designed a case-controlled study to investigate the potential association of a repeat polymorphism in IL-4 gene with specific clinical phenotypes of asthma. METHODS: The authors used the polymerase chain reaction to characterize the variation of the IL-4 intron 2 region in 145 unrelated Tunisian patients with asthma and 160 healthy control subjects. In order to strengthen the case-controlled study, analysis of IL-4 polymorphism was performed in families of several asthmatic patients. Asthma scores were determined and correlated with this polymorphism. RESULTS: Analysis of IL-4 polymorphism in patients with allergic asthma and in control subjects demonstrated a significant association between the IL-4 A1 allele and asthma. Further evidence of the strong association found between IL-4 intron 2 polymorphism and asthma was provided by the finding that asthma is transmitted in association with the inheritance of the IL-4 A1 marker. When patients were stratified into two groups according to the degree of the severity of asthma, the IL-4 A1 allele was specifically not associated with mild asthma, but highly associated with the moderate and severe forms of the disease. The relative risk (RR) of severe asthma is especially high in patients carrying the A1/A3 genotype (RR = 3.94, p = 0.0001). Conversely, a major decrease in the frequency of the IL 4 A3/A3 genotype was observed in patients with severe asthma, resulting in a significantly negative RR of this clinical phenotype of asthma (RR = 0.165, p = 0.0001). CONCLUSIONS: Tunisian persons carrying the IL-4 A1/A3 genotype may have an increased risk of severe asthma. PMID- 10529589 TI - Fragrance and contact allergens in vitro modulate the HLA-DR and E-cadherin expression on human epidermal Langerhans cells. AB - BACKGROUND: Epidermal Langerhans cells (LCs) play a critical role in the induction of contact hypersensitivity. The LCs leave the skin, move to the regional lymph nodes and present the allergens embedded in the HLA-DR molecule to naive T-lymphocytes. To allow LC emigration from the epidermis, E-cadherin must be downregulated. In this study, we have examined the early events that occur in the human epidermis after exposure to three strong contact sensitizers and two commonly used fragrances by examining alterations of E-cadherin and HLA-DR expression. METHODS: To determine whether E-cadherin and HLA-DR levels were modulated by allergens, flow cytometry was utilized to evaluate E-cadherin and HLA-DR expression on human epidermal LCs exposed to the different chemicals for 4 h at 37 degrees C. RESULTS: In vitro stimulation with the contact sensitizers isoeugenol, cinnamaldehyde, 2,4, 6-trinitrobenzenesulfonic acid, Bandrowski'sbase, or p-phenylene diamine resulted in a dose-dependent decrease of HLA-DR expression on the surface of LCs without affecting the number of positive cells. These contact allergens induced a downregulation of E-cadherin expression as well as a significant decrease of the percentage of E-cadherin-positive cells. Incubation with an irritant, sodium lauryl sulfate, did not significantly change HLA-DR and E-cadherin expression. CONCLUSIONS: Based on the alteration of E cadherin and HLA-DR expression of human LCs under short-term exposure conditions, there was a clear difference between contact sensitizers and a well-characterized irritant. For the first time, the ability of fragrance allergens in dipropylene glycol, a widely used vehicle in fragrance and cosmetic industries, was demonstrated to induce human LC phenotypic alterations. In combination with a series of in vitro tests, this rapid and simple method should help to detect the sensitizing potential of a substance to be applied onto the human skin as an alternative to animal testing. PMID- 10529590 TI - HgCl(2)-induced human lymphocyte activation in vitro: a superantigenic mechanism? AB - Mercuric chloride (HgCl(2)) has been proposed to be a mitogen for human blood lymphocytes in vitro. In our previous study, we demonstrated that HgCl(2) preferentially stimulates the CD4+ T cell subset to blast transformation and DNA synthesis and that the reaction is dependent on CD14+ accessory cells. In order to characterise the responding cells further and to elucidate the mechanism of T cell activation, the T cell receptor (TCR) Vbeta chain expression of the blast transformed cells was analysed by monoclonal antibodies and flow cytometry. The 22 TCR-Vbeta-specific antibodies used were found to react with 55-80% of the naive CD4+ and CD8+ blood T cells from the different donors. Six to 18% of the lymphocytes, mainly CD4+ T cells, were blast transformed after addition of HgCl(2). The distribution of the lymphoblasts carrying certain TCR Vbeta chains were skewed, and 15-40% expressed the TCR Vbeta2 chain. Furthermore, if cells were pretreated for 5 days with HgCl(2), whereafter recombinant interleukin-2 in fresh medium was added, the TCR Vbeta7+ T cell subset was also stimulated to blast transformation. The superantigen staphyloccal enterotoxin B, as a control, induced blast transformation in 10-26% of the lymphocytes, mainly CD4+ T cells, which were, as expected, positive for Vbeta3, Vbeta12, Vbeta14 or Vbeta17. We conclude that HgCl(2) has characteristics of a superantigen, activating the human lymphocytes in a Vbeta-chain-selective manner in vitro. PMID- 10529591 TI - A novel assay for serum complement activity: C42 generation assay. AB - BACKGROUND: In most clinics, laboratory tests for serum complement are limited to immunochemical determinations of C3 and C4 and are occasionally extended to the hemolytic titration of total complement functional activity (CH50). However, these tests are often not sufficient for the analysis of low CH50 serum. METHODS: A novel assay for serum complement activity, the C42 generation assay, has been developed. The principle of this assay is based on the hemolysis of sensitized sheep erythrocytes (EA) by complement components in two sera: C42 (the classical pathway C3 convertase) is generated on EA by C1, C4 and C2 in the first serum, followed by a second reaction leading to hemolysis by C3-C9 supplied by the addition of the second serum in the presence of EDTA. RESULTS: This methodology permits the evaluation of two distinct serum complement activities of a test serum. The combined activity of C1, C4 and C2, as well as the combined activity of C3-C9, can be estimated from the observed degrees of hemolysis. Information obtained from this assay is helpful for the analysis of test serum determined to have decreased CH50. Several clinical cases are presented in which this assay was utilized. CONCLUSIONS: The C42 generation assay is another functional assay of serum complement which can provide information beyond that obtained from the typical serum CH50 assay. Since intermediate cells or isolated complement components are not necessary, this assay can be employed rapidly and economically in a clinical setting. PMID- 10529592 TI - Lactic dehydrogenase virus infection inhibits allergic eosinophil reaction and IL 5 gene expression in vivo. AB - The effects of lactic dehydrogenase virus (LDV) infection on allergic eosinophil reaction and IL-5 gene expression were studied. LDV infection suppressed antigen induced eosinophil recruitment into the peritoneal cavity in sensitized mice. The elevation of IL-5 gene expression in the spleen and mesenteric lymph nodes 6 h after ovalbumin challenge was significantly suppressed in LDV-infected mice compared with uninfected (control) mice. The expression of the interferon-gamma and IL-2 genes in the spleen, but not in mesenteric lymph nodes, was significantly suppressed in LDV-infected mice compared with control mice. The present results suggest, that suppression of IL-5 gene expression by LDV infection may not be mediated by a mutual inhibitory mechanism between Th1 and Th2 cells. PMID- 10529593 TI - Human umbilical vein endothelial cells support interleukin-3- and interleukin-5 induced eosinophil differentiation from cord blood CD34+ cells. AB - BACKGROUND: Human umbilical vein endothelial cells (HUVEC) are an important source of hematopoietic cytokines, and interleukin-3 (IL-3)- and IL-5-induced eosinophil differentiation from CD34+ cells has been observed. To show the supportive effects of endothelial cells on eosinophil differentiation, we examined the effects of cocultured HUVEC on IL-3 and IL-5-induced eosinophil differentiation from human umbilical cord blood CD34+ cells. METHODS: CD34+ cells were obtained from the heparinized umbilical vein blood of 10 volunteers using a CD34-conjugated magnetic bead positive direct selection procedure. With HUVEC in Transwell, CD34+ cells were then cultured for 14-28 days. In neutralizing experiments on HUVEC-derived cytokines, antibodies to both stem cell factor (SCF) and granulocyte/macrophage colony-stimulating factor (GM-CSF) were added to the cell cultures. RESULTS: Cocultured HUVEC upregulated IL-3 and IL-5-induced eosinophil differentiation from CD34+ cells on day 28 of culture by 75.0%. The eosinophilopoietic effect of HUVEC was significantly only when the cells were present in the culture from day 15 to day 28. Addition of anti-SCF antibody or anti-GM-CSF monoclonal antibody to the culture significantly suppressed HUVEC combined IL-3- and IL-5-induced eosinophil differentiation on day 28 of culture by 49.2 and 55.0%, respectively. CONCLUSIONS: These results indicate that several cytokines including GM-CSF and SCF from HUVEC promote IL-3- and IL-5-induced eosinophil differentiation from CD34+ cells. PMID- 10529594 TI - Ecalectin as a T cell-derived eosinophil chemoattractant. AB - In our previous papers, we have shown that human T cells produce a unique eosinophil chemotactic factor (ECF), termed Ecalectin, with a molecular weight of about 30-50 kD during interaction with BALL-1 (a B cell line) extracts, antigen or mitogen. A 1.6-kB cDNA was isolated from a human T cell-derived expression library that encodes Ecalectin. Ecalectin is a 36-kD protein consisting of 323 amino acids based on its deduced amino acid sequence. Ecalectin was found to be a variant of human galectin-9, a member of the growing family of animal lectins exhibiting affinity for beta-galactosides. Recombinant Ecalectin, expressed in COS cells, exhibited potent ECF activity in vitro and in vivo in a dose-dependent manner but not chemotactic activity for neutrophils, lymphocytes, or monocytes. The finding that the Ecalectin transcript is present in various lymphatic tissues and that its expression increases substantially in antigen-activated peripheral blood mononuclear cells suggests that Ecalectin is an important T cell-derived regulator of eosinophil recruitment in allergic reaction sites. PMID- 10529595 TI - Analysis of biological activities of recombinant rat interleukin-5. AB - We analyzed the biological activities of recombinant rat interleukin-5 (IL-5). Purified recombinant rat IL-5 promoted the proliferation of T88-M cells in a concentration-dependent manner, and its effect was inhibited by an anti-murine IL 5-neutralizing polyclonal antibody. When bone marrow cells from normal rats were incubated with recombinant rat IL-5 in medium containing methylcellulose, the colony formation by eosinophilic cells was induced. Furthermore, when rat peritoneal eosinophils were incubated with recombinant rat IL-5, the spontaneous decrease in eosinophil viability was inhibited in a time- and concentration dependent manner. In addition, the recombinant rat IL-5-induced eosinophil survival was inhibited by an anti-murine IL-5-neutralizing polyclonal antibody. These findings suggest that rat IL-5 acts as B cell growth factor II, eosinophil differentiation factor, and eosinophil survival-enhancing factor. PMID- 10529596 TI - Primary role of CD4+ T cells and supplemental role of mast cells in allergic pulmonary eosinophilia. AB - BACKGROUND: We have recently demonstrated that allergic eosinophilic inflammation is transferred to unprimed mice by infusing IL-5-producing CD4+ T cells. The contribution of mast cells to the development of eosinophilic inflammation is controversial. METHODS: To clarify the possible different roles of CD4+ T cells and mast cells in eosinophilic inflammation, we compared antigen-induced airway eosinophilia between mast-cell-deficient mice (WBB6F1-W/W(v)) and their congenic normal littermates (WBB6F1-+/+). RESULTS: The time course study indicated that equivalent numbers of eosinophils were recruited into the airway of both +/+ and W/W(v) mice 6, 24, 96, and 216 h after antigen challenge, whereas the number of eosinophils 48 h after antigen challenge was significantly lower in W/W(v) compared to +/+ mice. Administration of either anti-CD4 or anti-IL-5 monoclonal antibody almost completely inhibited antigen-induced eosinophil recruitment in W/W(v) mice 48 h after antigen challenge. In contrast, the inhibitory effect of these antibodies in +/+ mice were partial (approximately 50% inhibition). Anti CD4 and anti-IL-5 antibodies equally suppressed airway eosinophilia in both +/+ and W/W(v) mice 96 h after antigen challenge. CONCLUSION: Our study indicates that CD4+ T cells are crucially involved in the development of allergic eosinophilic inflammation, while mast cells may play a supplemental role depending on the kinetics of the response. PMID- 10529597 TI - IL-4 induces eotaxin in human dermal fibroblasts. AB - A common feature of Th2-mediated skin diseases is selective eosinophil (Eo) infiltration in the affected dermis. However, the mechanisms of selective Eo recruitment are not yet fully understood. Our recent findings indicated that dermal fibroblasts actively participate in the accumulation of Eo in Th2-mediated skin inflammation. Following stimulation with IL-4, dermal fibroblasts produce eotaxin, which is the only biologically active Eo attractant detected in the supernatants. Natural eotaxin is comprised of a mixture of N-terminal-truncated and O-glycosylated variants. The expression of eotaxin mRNA is upregulated within 1 h after stimulation. TNF-alpha markedly enhances eotaxin mRNA expression and release of the protein product from IL-4-stimulated fibroblasts. As mast cells and Th2 cells produce both cytokines, it is probable that Eo recruitment into sites of allergen-induced, Th2-mediated skin reactions is, at least in part, due to IL-4-stimulated induction of eotaxin in dermal fibroblasts. PMID- 10529598 TI - Regulatory mechanisms of eosinophil adhesion to and transmigration across endothelial cells by alpha4 and beta2 integrins. AB - To participate in allergic airway inflammation, it is necessary for blood eosinophils (Eos) to adhere to and migrate across pulmonary vascular endothelial cells (EC). Accumulating evidence has established that Eos adhesion molecules, such as alpha4 and beta2 integrins, play central roles in these processes. We have reported that Eos spontaneously adhere to recombinant human (rh)-VCAM-1, while adhesion to rh-ICAM-1 requires a second stimulus such as GM-CSF. Furthermore, our study employing human pulmonary microvascular endothelial cells (HPMEC) revealed that although the alpha4 integrin/VCAM-1 pathway is crucial for the firm adhesion of Eos, subsequent transendothelial migration occurred dependent on the beta2 integrin/ICAM-1 pathway. We also discuss secretagogues that would affect Eos recruitment to the airways via regulation of alpha4 and beta2 integrins. PMID- 10529599 TI - Whole-blood flow-cytometric analysis of induction of adhesion molecules expression on eosinophils stimulated by phorbol-12-myristate-13 acetate/ionomycin. AB - BACKGROUND: Activation of eosinophils is closely associated with the pathology of allergic inflammatory disease, especially bronchial asthma. We recently investigated the activation of eosinophils by applying whole blood to a flow cytometer. We measured here beta1 and beta2 integrin on eosinophils stimulated by phorbol-12-myristate-13-acetate (PMA)/ionomycin to evaluate eosinophil activation in vitro using whole blood. METHODS: Heparinized whole blood was diluted with the same volume of RPMI 1640, then cells were incubated in the presence or absence of PMA and ionomycin for 45 min at 37 degrees C. After hemolyzation with lysing solution, flow-cytometric findings for CR3, LFA1-alpha, LFA1-beta and VLA-4 expression on eosinophils were examined. RESULTS: Mean fluorescent intensity (MFI) of CR3 and LFA1-beta stimulated by PMA and ionomycin was significantly higher than that of the unstimulated control. MFI of LFA1-alpha showed no significant difference from the unstimulated control. On the other hand, MFI of VLA-4 tended to decrease. CONCLUSIONS: Our method to distinguish eosinophils from various cell groups in whole blood is simple and time-saving, similar to conditions in vivo and may allow intensive investigation of eosinophils in clinical laboratories as well as in research laboratories. We are currently investigating the influence of different kinds of stimulations, regulation factors or agents on eosinophils using this method. PMID- 10529600 TI - Interferon-gamma increases CD62L expression on human eosinophils. AB - BACKGROUND: L-selectin (CD62L) and Mac-1 (CD11b/CD18) play a crucial role in the infiltration of eosinophils. However, changes in CD62L and CD11b expression on eosinophils after stimulation with cytokines were little studied. METHODS: Eosinophils in peripheral blood of healthy volunteers were purified and cultured with interleukin-5 (IL-5), granulocyte/macrophage colony-stimulating factor (GM CSF) or interferon-gamma (IFN-gamma). After up to 24 h incubation, CD62L and CD11b expression were examined using flow cytometry. The effects of dexamethasone (Dex), cycloheximide (CHX) or theophylline on CD62L expression on IFN-gamma stimulated eosinophils were also studied. RESULTS: IL-5 or GM-CSF downregulated CD62L and upregulated CD11b expression on eosinophils after 30 min stimulation. Conversely, IFN-gamma upregulated CD62L after 12 h stimulation in a time- and dose-dependent manner, and had no effect on CD11b expression. Dex, CHX or theophylline dose-dependently decreased CD62L expression on IFN-gamma-stimulated eosinophils. CONCLUSIONS: IFN-gamma is a particular cytokine which can increase CD62L expression on circulating or infiltrated human eosinophils. It is suggested that protein synthesis and intracellular cAMP participate in the increase in L selectin expression on IFN-gamma-stimulated eosinophils. PMID- 10529601 TI - Adhesion to fibronectin regulates expression of intercellular adhesion molecule-1 on eosinophilic cells. AB - BACKGROUND: Adhesion molecules may play an important role in eosinophilic activation as well as adherence of extracellular matrix (ECM) including fibronectin (FN) in the inflamed focus. Tissue eosinophils expressed inter cellular adhesion molecule-1 (ICAM-1, CD54), whereas peripheral blood eosinophils did not. The molecular mechanisms of ICAM-1 expression on eosinophils are not clear. We immunologically examined the effect of adherence to FN on the expression of adhesion molecules in an eosinophilic cell line (EoL-1). METHODS: EoL-1 cells, suspended at a concentration of 2 x 10(6) cells/ml, were cultured at 37 degrees C in BSA or FN-coated 24-well plates. After 4 h, the cells were removed by pipetting, and the expression of adhesion molecules was evaluated by immunofluorescence. RESULTS: Adherence of EoL-1 to FN enhanced ICAM-1 (CD54) expression on EoL-1 (FN vs. BSA: 84.76 +/- 17.90 vs. 43.73 +/- 7.60, mean fluorescent intensity). Regarding other adhesion molecules on EoL-1 (CD11a, CD18, CD49d), the expression was not significantly augmented by adherence to FN. CONCLUSIONS: We concluded that the signal via adhesion of FN regulates the expression of ICAM-1 on eosinophilic cells. This study suggests that eosinophilic adhesion to ECM via adhesion molecules plays an important role in the pathogenesis of allergic inflammation through eosinophilic functional changes, including regulation of expressive adhesion molecules such as ICAM-1. PMID- 10529602 TI - Functional expression of transmembrane 4 superfamily molecules on human eosinophils. AB - BACKGROUND: Transmembrane 4 superfamily (TM4SF) molecules are exclusively found on hematopoietic cells. Several members of the TM4SF are reported to be associated with other cell surface molecules, including integrins, and might participate in signal transduction, but little is known about their role on eosinophils. In the present study, we determined the expression and function of TM4SF molecules on human eosinophils. METHODS: Surface expression of TM4SF molecules on purified peripheral blood eosinophils was examined using indirect immunofluorescence and flow cytometry. Purified eosinophils were incubated with anti-TM4SF monoclonal antibodies (mAbs) for up to 24 h. Eosinophil activation was evaluated by measuring eosinophil homotypic aggregation as well as changes in surface expression of CD11b or CD62L by flow cytometry. RESULTS: Freshly isolated eosinophils expressed CD9, CD37, CD53, CD63 and CD81. Incubation with anti-CD9 mAb but not with anti-CD37, CD53, CD63 or CD81 mAb induced significant eosinophil homotypic aggregation. Incubation with any of the anti-TM4SF mAb for 30 min failed to alter the expression of either CD11b or CD62L on eosinophils. In contrast, the expression of CD11b was significantly enhanced after 24 h of incubation with anti-CD53 mAb, while the expression of CD62L was significantly reduced with anti-CD81 mAb. CONCLUSIONS: Cross-linking of some surface TM4SF molecules induced significant eosinophil homotypic aggregation, upregulation of CD11b expression, or CD62L shedding, consistent with activation of eosinophils. Our data suggest that several TM4SF molecules are functionally expressed on human eosinophils, and therefore might participate in allergic inflammation. PMID- 10529603 TI - CCR3 mRNA expression in bronchial epithelial cells and various cells in allergic inflammation. AB - BACKGROUND: RANTES and eotaxin are important chemokines involved in the activation and migration of eosinophils and are considered to play a major role in allergic inflammation. METHODS: In this study, we used RT-PCR to investigate the kinds of cells that express mRNA for CCR3, a common receptor of these chemokines, and eotaxin, a ligand for CCR3. RESULTS: CCR3 mRNA was expressed in eosinophils, peripheral mononuclear cells, an eosinophilic cell line (EoL-1), a bronchial epithelial cell line (NCI-H(292)), human endothelial cells and nasal washings from patients with allergic rhinitis. CONCLUSION: These results suggest that the CCR3-eotaxin system plays an important role in generating inflammation, since these substances are expressed not only in cells implicated in activation or migration of eosinophils but also in various other cells involved in allergic inflammation. PMID- 10529605 TI - Intracellular signaling pathways in human eosinophil activation: role of a beta2 integrin, alphaMbeta2. AB - Cellular adhesion is crucial roles for eosinophil effector functions such as degranulation. Here, we focused on the role of a beta2 integrin, alphaMbeta2, in intracellular signaling pathways of human eosinophil activation. We found that the ligation of alphaMbeta2 triggers the activation of an intracellular signaling cascade, including protein tyrosine phosphorylation and phosphoinositide turnover, and subsequent cellular degranulation in human eosinophils. These signaling events may play important roles in adhesion-dependent cellular functions of eosinophils. PMID- 10529604 TI - Effect of eotaxin on the generationof reactive oxygen species from eosinophil cell line, YY-1. AB - BACKGROUND: The CC chemokine eotaxin is a selective chemoattractant for eosinophils. Eosinophils have been considered to be the major effector cells in allergic inflammation, since not only eosinophil-specific granule proteins but also reactive oxygen species (ROS) from eosinophils may cause the damage to the cells or tissue of the mucosal epithelium. In this study, we examined the effect of eotaxin on ROS from an eosinophil cell line, YY-1. METHODS: ROS in luminol dependent reaction were examined. Calcium ionophore A23187 were added to the mixture of YY-1 cells with luminol, and then ROS were determined. RESULTS: Eotaxin primed the production of ROS from YY-1 cells. ROS from untreated YY-1 cells evoked with calcium ionophore A23187 in luminol-dependent chemiluminescence gave a maximal value of 1,928 +/- 223 intensity counts (IC; mean +/- SE, n = 4) and an integral value of 17.04 +/- 1. 51 IC (x10(-4)), while eosinophils that were treated with eotaxin gave a maximal value of 2,264 +/- 86 IC (10 nM), 2,691 +/- 124 IC (100 nM) and an integral value of 21.22 +/- 0.67 IC (x10(-4); 10 nM), 26.20 +/- 1.41 IC (x10(-4); 100 nM). CONCLUSION: Eotaxin might play important roles in the pathogenesis of allergic inflammation through eosinophil activation by priming of eosinophil oxidative metabolism as well as involvement in selective eosinophil chemotaxis. PMID- 10529606 TI - Tumor necrosis factor-alpha mediates antiapoptotic signals partially via p38 MAP kinase activation in human eosinophils. AB - Tumor necrosis factor-alpha (TNF-alpha) is a proinflammatory cytokine with many biological effects on a variety of cells. In particular, TNF-alpha has been shown to act as a death or survival factor which mediates apoptosis or antiapoptotic signals in various types of cells. In eosinophils, TNF-alpha has been reported to activate eosinophil functions. However, it is not clearly defined whether TNF alpha delivers antiapoptotic signals in eosinophils. In order to determine whether TNF-alpha prevents eosinophil apoptosis, we examined the effect of TNF alpha on eosinophil apoptosis by the survival assay and cell cycle analysis. We also determined whether intracellular MAP kinases (ERKs, Jun kinase/JNK, and p38 MAP kinase) are involved in the TNF-alpha-induced signaling for the prevention of eosinophil apoptosis. We showed that TNF-alpha mediated antiapoptotic signals in human eosinophils in part via activation of p38 MAP kinase, but not via activation of ERKs and JNK. Our data suggest that TNF-alpha/p38 MAP kinase pathways are involved in the regulation of eosinophil survival and, thus, would be important for the development of allergic eosinophil-rich inflammation. PMID- 10529607 TI - Mechanisms of eosinophil cationic protein release in the serum: role of adhesion molecules. AB - BACKGROUND: Measurement of eosinophil cationic protein (ECP) in serum has been utilized as a marker for allergic inflammation. The serum level of ECP represents the level found in vivo plus additional proteins released in vitro from peripheral blood eosinophils during the coagulation period. The mechanisms of release, however, are unclear. We investigated a possible involvement of adhesion molecules in the ECP release. MATERIALS AND METHODS: Venous blood was drawn in the presence of EDTA from allergic donors. The blood was incubated with neutralizing monoclonal antibodies to CD18, CD11a, CD11b, CD29, CD49d, CD54, alpha4beta7, or isotype-matched control antibodies, respectively, at 4 degrees C for 30 min. Calcium gluconate (calcium) was then added to induce coagulation. The blood was further incubated for 90 min and centrifuged to obtain the serum. ECP in the serum was measured with RIA. In some experiments, purified eosinophils were incubated with plasma and calcium, then ECP in the supernatants was assayed. RESULTS: ECP in the samples with calcium was significantly higher than in those without calcium. Purified eosinophils released ECP upon plasma coagulation. Anti CD18, CD49d, and alpha4beta7 antibodies significantly suppressed ECP levels in the serum. CONCLUSIONS: These results suggest that ECP release in the serum is calcium and plasma coagulation-dependent and that cell adhesion through alphaLbeta2, alphaMbeta2, alpha4beta1 and alpha4beta7 integrins is at least in part responsible for ECP release. PMID- 10529608 TI - Matrix metalloproteinase-9 in peripheral blood eosinophils. AB - BACKGROUND: Matrix metalloproteinases (MMPs) are major contributors to tumor invasion, remodeling of connective tissue and infiltration of inflammatory cells and may be important mediators in developing allergic inflammation. Overexpression of MMP-9 mRNA by eosinophils in the asthmatic airways has been reported. To clarify the relative significance of MMP as an inflammatory mediator from eosinophils, we determined the content of MMP-9 in the peripheral blood eosinophils and compared it with the other leukocyte fractions. METHODS: Peripheral blood eosinophils, neutrophils, and mononuclear cells were purified from normal and allergic donors with Percoll gradient centrifugation and CD16 negative selection. Cell lysate and culture supernatants stimulated with IL-5, PAF, and PMA were tested for MMP-9 with gelatin zymography and ELISA. RESULTS: The amount of MMP-9 in highly purified eosinophils, neutrophils, and mononuclear cells was 2.5 +/- 0.9, 4,073 +/- 581, and 7.6 +/- 1.4 ng/5 x 10(6) cells, respectively. There was no difference in MMP-9 content of eosinophils between normal donors and patients with asthma. Culture of peripheral blood eosinophils with IL-5 for 4 days did not induce MMP-9 production. The stimulation of eosinophils with PMA and other secretogogues caused only small amounts of MMP-9 secretion as compared with neutrophils. CONCLUSIONS: These findings suggest that circulating eosinophils normally have only small amounts of MMP-9 and that eosinophils may need complex activation signals to produce significant amounts of MMP as seen in tissues of allergic inflammation. PMID- 10529609 TI - NC/Nga mice: a mouse model for atopic dermatitis. AB - Atopic dermatitis (AD) is a common pruritic disease that occurs primarily in infancy and childhood. AD is characterized by itching and the patient having an individual or family history of atopic diseases. Although AD is also frequently associated with elevated serum IgE levels and with common environmental factors contributing to its pathogenesis, the etiology of AD is still unknown. We examined NC/Nga mice (NC mice) that showed AD-like skin lesions with aging as a possible mouse model for AD. NC mice were maintained under conventional (Conv) or specific pathogen-free (SPF) conditions. Clinical symptoms, serum IgE levels and histopathology of the skin were compared between these 2 groups, and we explored their application as a model of human AD. It was found that the skin lesions of inbred NC mice were clinically and histologically very similar to human AD when the mice were raised under Conv conditions, but not under SPF conditions, and we assumed that some kinds of environmental factors might trigger AD-like signs and symptoms in NC mice. To further investigate the pathophysiology and treatment of AD, a suitable animal model is absolutely required, and NC mice are very useful for this purpose. PMID- 10529610 TI - Atopic asthma is dominant in elderly onset asthmatics: possibility for an alteration of mast cell function by aging through Fc receptor expression. AB - The morbidity and mortality of elderly onset asthma have recently been on the increase. To evaluate the allergic status of the elderly onset asthmatics, we performed skin tests for allergens and estimated serum total IgE levels of the patients. The proportion of the skin-test-positive patients among the elderly onset asthmatics was significantly larger than that in the younger onset group, whereas no significant difference was observed in the frequency of patients with serum total IgE levels higher than the normal range (287.2 IU/ml). On the other hand, in the experiments with aged mice, isolated peritoneal mast cells (PMC) showed a considerable decrease in the FcgammaRIIB/III expression and higher degranulation triggered by 2.4G2, as compared to the PMC derived from young mice. Since FcgammaRIIB has been shown to act as a negative regulator, we speculate that in the aged mice Fc receptor-mediated mast cell function may be upregulated by a release from the negative regulation as a result of decreased FcgammaRIIB expression. Our results obtained from a mouse model system may help to understand pathophysiological mechanisms underlying elderly onset asthma. PMID- 10529611 TI - Correlation between respiratory function and airway inflammation in asthma. AB - BACKGROUND: Eosinophilic airway inflammation is thought to be associated with airflow limitation and airway hyperresponsiveness. METHODS: Spirometry, peak expiratory flow (PEF) measurement, histamine challenge test, and sputum induction with hypertonic saline inhalation were performed in 70 asymptomatic patients with asthma who were treated with bronchodilators or an inhaled corticosteroid (beclomethasone dipropionate, 400-800 microg/day) or both for more than 2 months. The relationships between the sputum eosinophils, respiratory function, and airway responsiveness to histamine were investigated. RESULTS: Induced sputum was obtained from 53 patients. Although significant correlations between sputum eosinophils and forced expiratory volume in 1 s/forced vital capacity x 100% (FEV1%) or maximum expiratory flow at 25% of the forced vital capacity (V25) divided by height (V25/Ht) were observed, no significant correlations were found between sputum eosinophils and percent predicted PEF (%PEF) or airway responsiveness to histamine. Furthermore, airway responsiveness to histamine was not correlated with FEV1%, V25/Ht or %PEF. CONCLUSION: Single-point measurements of FEV1% and V25/Ht but not PEF or airway responsiveness to histamine correlate with the degree of airway inflammation evaluated indirectly with induced sputum. PMID- 10529612 TI - Heterogeneity in expression of cytokine mRNA in freshly isolated peripheral blood eosinophils of patients with cutaneous-disease-associated eosinophilia. AB - BACKGROUND: It is thought that there are subpopulations of eosinophils concerning cytokine production: some of them predominantly produce Th1 cytokines, while others produce Th2. METHODS: By semiquantitative RT-PCR, the in vivo levels of the cytokine mRNA were measured in freshly isolated eosinophils and peripheral blood mononuclear cells (PBMCs) of 19 patients with eosinophilia, including 6 with atopic dermatitis, 4 with drug eruption, 1 with chronic dermatitis, 2 with erythroderma, 3 with bullous pemphigoid, 1 each with urticaria, adult T cell lymphoma, and hypereosinophilic syndrome, and 7 healthy individuals. Eosinophils and PBMCs were isolated by centrifugation over Ficoll-Paque and magnetic cell separation using anti-CD16 antibody. RESULTS: The IL-4/IFN-gamma ratio in eosinophils as well as PBMCs was increased and correlated well with IgE levels in atopic dermatitis. In other cytokines, such as IL-8, TNF-alpha and TGF-beta1, the levels of mRNA expression were variable among each sample. CONCLUSION: Not only helper T lymphocytes but also eosinophils may take part in the pathogenesis of cutaneous diseases by virtue of heterogeneity of Th1 and Th2 cytokine production. PMID- 10529613 TI - Difference in apoptotic function between eosinophils from peripheral blood and bronchoalveolar lavage in chronic eosinophilic pneumonia. AB - We compared apoptosis in eosinophils from bronchoalveolar lavage (BAL-Eos) with that in eosinophils from peripheral blood (PB-Eos) of 4 patients with chronic eosinophilic pneumonia (CEP). The survival rate of the BAL-Eos on the 3rd day of the culture was significantly higher than that of the PB-Eos (39.1 vs. 1.3%). The percentage of apoptotic cells in the PB-Eos after a 24-hour incubation was higher than that in the BAL-Eos (21.7 vs. 10.6%) according to an analysis with annexin V. We further found that ECF-PI9, an eosinophil chemotactic factor (ECF) derived from an established T cell line (STO-2), significantly suppressed the apoptosis of both PB-Eos and BAL-Eos and prolonged their survival. The expression of Fas on PB-Eos was significantly suppressed by ECF-PI9 (18.5 to 7.37%, p < 0. 05), whereas ECF-PI9 failed to suppress the Fas expression on BAL-Eos (3.3 to 3.6%). In addition, an ECF with similar physicochemical properties and biological functions was isolated from the BAL fluid of patients with CEP. These data demonstrate differences between PB-Eos and BAL-Eos, and indicate that ECF-PI9 is involved in the pathogenesis of CEP. PMID- 10529614 TI - Circadian variation in nasal reactivity in children with allergic rhinitis: correlation with the activity of eosinophils and basophilic cells. AB - BACKGROUND: In allergic rhinitis, the major symptoms of runny nose, sneezing, and stuffy nose tend to become worse upon waking up in the morning, and yet the mechanisms underlying this phenomenon are poorly understood. We investigated whether the worsening of allergic rhinitis in the morning is associated with changes in the activity of inflammatory cells. METHODS: Nasal reactivity to methacholine was assessed twice in 8 children with allergic rhinitis and 8 healthy control subjects at 6.00 a.m. and 3.00 p.m. The amounts of eosinophil cationic protein (ECP), histamine and tryptase in induced nasal secretions and peripheral blood were also measured. RESULTS: Nasal reactivity to methacholine was higher at 6.00 a.m. not only in patients but also in healthy controls. Serum ECP and plasma histamine levels showed no circadian patterns. On the other hand, significantly higher levels of inflammatory activation products were found in nasal secretions at 6.00 a.m., thus showing a direct association with nasal reactivity. CONCLUSION: These results suggest that the circadian variation in nasal reactivity is associated with changes in the activity of eosinophils and basophilic cells in the nasal mucosa. PMID- 10529615 TI - Overexpression of CD11b on eosinophils in atopic dermatitis: downregulation by cyclosporin A and upregulation by interleukin 5. AB - We examined the level of expression of CD11b on eosinophils in pripheral blood samples from patients with atopic dermatitis (AD) and non-AD volunteers. Eosinophils were defined using a new method employing CD14/CD45 and a backgate technique. Overexpression of CD11b was noted in eosinophils of AD patients. Treatment of AD with cyclosporin A resulted in clinical improvement as well as reduction in the expression of CD11b. Stimulation of eosinophils from patients with inactive AD by interleukin 5 upregulated the expression of CD11b on these cells. Our results suggest that the expression of CD11b surface molecule on eosinophils may play an important role in the activity of AD. PMID- 10529616 TI - Connexin 26 expression and extensive gap junctional coupling in cultures of GT1-7 cells secreting gonadotropin-releasing hormone. AB - Gap junctions (GJs) are transmembrane channels that permit rapid intercellular transit of various small molecules including ions, second messengers and metabolites. GJs promote communication and coordinated activity between coupled neurons, and may help facilitate the synchronous release and pulsatile secretion of neurohormones. A previous study using GnRH-secreting GT1-7 cells reported that connexin 26 was the major GJ subunit present, and that about 20% of the cultured cells engaged in GJ coupling as assayed by fluorescence recovery after photobleaching of 5,6-carboxyfluorescein diacetate (MW 460 D). To reassess GJ connectivity with a more permeant probe, we grew GT1-7 cells to 70% confluency on Matrigel-coated glass coverslips and microinjected Neurobiotin(TM) (MW 322 D) into single cells. Dye was allowed to diffuse for 30 min before cultures were fixed, and subsequently immunostained for Neurobiotin with 3,3'-diaminobenzidine HCl and examined by light microscopy. Dye coupling between 2 or more GT1-7 cells was observed after 75% of all microinjections. Connectivity involved the somata and neurites of an average of 6.6 +/- 2.0 adjoining cells, but in one instance was seen in a group of 32 GT1-7 neighbors. Western blotting and immunofluorescence staining confirmed that connexin 26 was the predominant GJ subunit expressed by GT1-7 cultures. Our results using Neurobiotin suggest these GJ channels may be smaller than anticipated. In addition, functional GJ connectivity between subconfluent GT1-7 cells is more extensive than previously reported, occurring with higher frequency and coupling significantly greater numbers of cultured cells. Since cAMP, IP3, and Ca(2+) are able to pass through GJs and can elicit secretion of GnRH by GT1 cell cultures, GJs may play an important role in the coordination and synchronization of GnRH release. PMID- 10529617 TI - Localization of estrogen-receptive neurons projecting to the GnRH neuron containing rostral preoptic area of the ewe. AB - Estrogen exerts important feedback effects upon the biosynthetic and secretory behavior of gonadotropin-releasing hormone (GnRH) neurons to control reproductive functioning. The mechanism of estrogen action upon these neurons is unclear and seems likely to involve the transsynaptic regulation of GnRH neurons. The objective of the present study was to identify the estrogen-receptive neural populations which project to the general vicinity of the GnRH perikarya in the rostral preoptic area and diagonal band of Broca (rPOA/DBB) of the ewe. Intact breeding-season ewes received an injection of the retrograde tracer fluorogold (FG) into the rPOA/DBB, and their hypothalami and brainstems examined for the presence of FG and estrogen receptor alpha (ERalpha) immunocytochemistry. Retrogradely labeled neurons were identified principally within the lateral septum (LS), lamina terminalis, bed nucleus of the stria terminalis, POA, arcuate nucleus (ARN), ventromedial nucleus (VMN) and median eminence. Smaller numbers of FG-immonoreactive cells were found in the caudal brainstem where they resided mostly in the ventrolateral medulla (VLM). Dual-labeled cells exhibiting both FG and ERalpha staining were prominent in the POA, LS and at all rostrocaudal levels of the VMN and ARN. Small numbers of dual-labeled cells were found in the VLM. These observations indicate that a number of distinct ERalpha-expressing neural populations project to the rPOA/DBB where the majority of the GnRH perikarya are found in the ewe. Although it is not possible to determine the direct connectivity of these projections with GnRH neurons, the findings provide an initial neuroanatomical framework through which the transsynaptic actions of estrogen on ovine GnRH neurons may be tested. PMID- 10529618 TI - Suppression of tonic luteinizing hormone secretion and norepinephrine release near the GnRH neurons by estradiol in ovariectomized rats. AB - One of the major neurotransmitters that controls pulsatile luteinizing hormone (LH) secretion is norepinephrine (NE). NE pulses detected in the median eminence of ovariectomized rhesus monkeys are highly correlated with both GnRH and LH pulses. In contrast, previous reports suggest that this is not the case in rats, thus it remains to be determined whether NE stimulates LH release on a pulse-by pulse basis in that species. Further, a variety of indirect evidence supports the hypothesis that in rats, estradiol exerts its negative feedback action on LH secretion in part by inhibiting noradrenergic neurotransmission that is stimulatory to LH release, but there is no direct evidence to support this hypothesis. Therefore the following study was designed to test the hypothesis that estradiol suppresses NE release in the vicinity of the GnRH neurons after ovariectomy. In addition, we examined whether episodes of NE release are correlated with LH pulses in ovariectomized rats. Blood samples and microdialysates of the diagonal band of Broca/medial preoptic area (DBB/MPOA) were collected every 5 min from 09:00 to 14:00 h from untreated or estradiol treated (4-5 days), long-term ovariectomized (1-4 months) rats for determination of plasma LH by RIA and NE release by HPLC. The results indicate that in both untreated and estradiol-treated ovariectomized rats, LH pulses are not correlated with episodes of NE. Thus, NE may play a permissive role in the control of pulsatile LH secretion in rats. Further, estradiol treatment leads to a suppression of both plasma LH levels and NE release in the DBB/MPOA, supporting the hypothesis that a decrease in NE neurotransmission that is stimulatory to LH release mediates the negative feedback action of estradiol on tonic LH secretion. PMID- 10529619 TI - Activin A regulation of gonadotropin-releasing hormone synthesis and release in vitro. AB - Activin is essential for the regulation of normal mammalian reproductive function at both the pituitary and gonadal levels. However, its central actions in the control of the hypothalamic-pituitary-gonadal axis remain largely unexplored. The present study aims to determine whether activin could regulate the reproductive axis at the level of the hypothalamus, through control of the GnRH neuroendocrine system. Using the GnRH-secreting GT1-7 neuronal cell line as a model system, we demonstrate expression of mRNAs encoding activin receptor types I, IB, and II. We examined the effects of activin A on GnRH protein secretion and mRNA levels in GT1-7 cells. Treatment with rh-activin A regulated both GnRH protein secretion and GnRH mRNA expression in the GT1-7 cells in a time-dependent fashion. Using transient transfection assays, we explored a potential transcriptional basis for these changes. Activin A increased reporter gene activity driven by minimal GnRH enhancer and promoter elements, suggesting that activin may regulate GnRH gene expression at the level of transcription. Lastly, activin A treatment of male rat hypothalami, in vitro, increased GnRH protein secretion. Collectively, molecular and physiological evidence support the presence of an activin system which might act at a hypothalamic site to regulate mammalian reproduction via activation of GnRH synthesis and release. PMID- 10529620 TI - Androgen regulation of immunological and biological activities of pituitary follicle-stimulating hormone isoforms in male rats. AB - Follicle-stimulating hormone (FSH) is involved in the regulation and maintenance of gametogenesis. It exists in multiple molecular forms with different oligosaccharide structures which in turn are influenced by the hormonal milieu. Previous studies from our laboratory demonstrated that antiandrogen administration to immature male rats altered the biological activity and the distribution profile of pituitary FSH isoforms. The aim of this study was to examine possible modifications in pituitary FSH polymorphism throughout sexual development (10-, 32- and 75-day-old rats). In addition, the effect of androgen deprivation by castration (32-day-old rats) and its replacement with a nonaromatizable androgen - dihydrotestosterone - on pituitary FSH polymorphism was determined. Concanavalin A affinity chromatography was used to isolate groups of FSH isoforms according to their carbohydrate inner structure. Radioimmunoassay and Sertoli cell bioassay were used to evaluate FSH immuno- and bioactivities. Androgen rise in serum was accompanied by a marked increase in pituitary bio- and immuno-FSH content in 32- and 75-day-old rats. However, FSH pituitary content did not vary despite the significant increment observed in serum FSH levels after castration and decrease to control levels after androgen replacement. The distribution profile of immuno- and bioactive FSH changed throughout sexual maturation. The proportion of pituitary FSH isoforms bearing complex oligosaccharide structures (triantennary, bisecting, complete and truncated biantennary) increased with age, with a concomitant decrease in the proportion of isoforms bearing incomplete carbohydrate chains. The distribution profile observed in castrated 32-day-old rats was similar to that determined in 10-day old animals. Androgen replacement restored the distribution profile to normal. These results suggest that androgens regulate the incorporation of sugar residues to the carbohydrate chains of pituitary FSH favoring the biosynthesis of complex type oligosaccharide structures. PMID- 10529621 TI - Effects of androgens and antiandrogens on the quantitative immunohistochemistry of gonadotrope cells in prepubertal male rats. AB - In the male rat, androgens are involved in the feedback regulation of gonadotropin synthesis and secretion. Specific androgen-receptor blockade by the nonsteroidal antiandrogens, flutamide and Casodex, has proven to be a valid tool for studying androgen effects in vivo. The aim of the present study was to investigate the effect of antiandrogen administration at the pituitary level by evaluating the changes in gonadotropes through quantitative immunohistochemistry, and by comparing these alterations with the effect of androgen deprivation by castration either with or without subsequent androgen replacement. Male Sprague Dawley rats (23 days old) were randomly divided into 5 groups for the following treatments: (a) controls; (b) flutamide-injected (10 mg/rat/day in a gelatin vehicle); (c) Casodex-injected (10 mg/rat/day in an oil vehicle); (d) castrated, and (e) castrated and dihydrotestosterone propionate-replaced (40 microg/rat/day in an oil vehicle). Groups were then sacrificed after 10 days of maintenance under each condition. Pituitaries were fixed in Bouin's fluid and embedded in paraffin. Serial sections (4 micrometer) were obtained at different levels and immunostained by means of the primary murine monoclonal antibodies anti-FSH and anti-LH and a peroxidase-mediated EnVision System (Dako). Measurements of volume density (VD) and individual mean cell area were made by means of an image analysis system (Imaging Technology, Optimas). Serum FSH and LH levels were determined by radioimmunoassay (RIA). Serum gonadotropin levels, VD, and mean cell area increased significantly in the flutamide-treated, Casodex-treated, and castrated groups (p < 0.05). Androgen replacement in the castrated rats, however, reduced VD, mean cell area, and serum gonadotropins to levels comparable to those of controls. We conclude that either androgen blockade by antiandrogens or castration produce an enhancement in the gonadotrope cell population in prepubertal rats, as shown by an increase in both VD and mean cell area, as well as an elevation in FSH- and LH-immunoreactive cells. These observations correlate well with the changes found in the levels of circulating gonadotropins as measured by RIA. PMID- 10529622 TI - Modulation of prohormone convertase 2 in spinal cord during gestation and hormone simulated pregnancy. AB - Gestation as well as its hormonal simulation (HSP) is characterized by an enhanced spinal dynorphin/kappa-opioid antinociception. This antinociception is accompanied by decreased content of dynorphin precursor intermediates and increased content of mature dynorphin peptides (1-17 and 1-8) in the lumbar spinal region. This suggests that augmented processing of spinal dynorphin precursor intermediates is an adaptive mechanism used by dynorphin neurons to meet increased synthetic demands necessitated by increased dynorphin neurotransmission. Prohormone convertase (PC) 1 and 2 represent major secretory granule proteolytic processing activities capable of converting neuroendocrine and neurotransmitter peptide (dynorphin) precursor intermediates to their mature, biologically active products. Accordingly, the current investigation was undertaken to assess their potential relevance to peptidergic (dynorphin) neuronal functional plasticity in vivo. In order to evaluate a molecular biological parameter of PC2 synthesis, a solution hybridization assay was developed with which to quantify changes in the spinal lumbar content of its mRNA. This study demonstrates that during gestation and HSP, lumbar PC2 protein content, but not that of PC1, is augmented. The increase in lumbar PC2 during HSP indicates that the pregnancy blood concentration profile of 17beta-estradiol and progesterone is a predominant facet of the pregnant condition responsible for its modulation during this condition. In contrast to the elevated content of lumbar PC2 protein, levels of PC2 mRNA in the lumbar cord of pregnant or HSP rats were essentially unchanged. This indicates that increased transcriptional activity is not, necessarily, a prerequisite for increased PC2 protein content to be manifest. These observations suggest positive modulation of PC2 to be a critical component of the mechanism(s) by which spinal dynorphin neurons adapt to the demand-induced increased production of mature dynorphin peptides. PMID- 10529623 TI - On the gender differences in sleep-endocrine regulation in young normal humans. AB - Sleep-endocrine regulation in humans involves high activity of the somatotropic axis at the beginning of the night and an increase in the hypothalamic-pituitary adrenocortical (HPA) system during the night. Gender differences were examined with regard to sleep-endocrine regulation in young healthy controls (10 men, 9 women). The sleep EEG was recorded (23:00-07:00 h) and plasma samples were collected and analyzed for GH, cortisol and ACTH at 20-min intervals. Cortisol secretion was significantly higher in females during the first half of the night (F = 9.9, p < 0.05), while ACTH was not different. In women, sleep-EEG analysis showed less slow wave sleep (SWS) during the second half of the night (F = 4.5, p < 0.05) and a significantly greater decrease in SWS and delta activity from the first to the second half of the night (F = 3.7 and 7.4, respectively, p < 0.05). Sigma activity increased during the night in women only (F = 3.7, p < 0.05). Our data are compatible with the hypothesis that in women compared to men activity of hypothalamic CRH neurons and central CRH release is greater, but is not reflected by greater HPA activity. PMID- 10529625 TI - Lipoprotein glomerulopathy: renal lipidosis induced by novel apolipoprotein E variants. PMID- 10529624 TI - Natriuretic peptide receptors of type A in human neuroblastomas. AB - Functional natriuretic peptide receptors of type A (NPR-A) were detected in the human neuroblastoma NB-OK-1, SK-N-SH and SK-N-BE, but not the SH-SY5Y, cell lines. Also, NPR-A mRNA was detected in 19 of the 25 tumor neuroblastoma samples tested in this study. Five of the eight tumor neuroblastoma samples that were assayed for atrial natriuretic peptide (ANP) binding revealed the presence of ANP binding sites. In the human neuroblastoma NB-OK-1 cell line, [(3)H] thymidine incorporation was increased in response to ANP, decreased in response to pituitary adenylate cyclase-activating polypeptide (PACAP-27), and the stimulatory effect of ANP was inhibited by PACAP-27. Tissue transglutaminase activity was decreased by ANP and PACAP-27, and their effects were additive. However, neither cell cycle phases, cell growth, or cell apoptosis were modified by ANP or PACAP-27 treatments. PMID- 10529626 TI - Antihypertensive therapy in renal patients - benefits and difficulties. AB - High blood pressure values, diastolic and systolic, are associated with decreased renal function. This is particularly true when the diastolic blood pressure is higher than 90 mm Hg. Several studies showed that lowering of the blood pressure within the range of normotension according to the WHO causes a reduction in the rate of progression to terminal renal failure. These studies have led to recommendations to aim at a target blood pressure of approximately 125/75 mm Hg in the treatment of patients with glomerular diseases and particularly diabetic nephropathy with proteinuria >1 g/day. In contrast to these results, blood pressure values corresponding to the recommendation ( tamsulosin > amosulalol > HV-723 > labetalol > ketanserin > clonidine > propranolol. Thus, although the affinity of tamsulosin to the alpha(1B)-AR subtype in the canine aorta was as high as that in the bovine prostate reported in our previous study, the affinity (pKi 7. 87) of this drug to alpha(1L)-AR in the canine aorta was lower than that (pKi 8.99) in the bovine prostate. These observations suggested that the pharmacological potencies of tamsulosin in the aorta and prostate may be different. PMID- 10529657 TI - Effects of organic anions and vinblastine on biliary excretion of erythromycin in rats. AB - Since little is known about the mechanism of biliary excretion of cationic drugs, biliary excretion of erythromycin was studied in rats. Infusion of sulfobromophthalein and taurocholate significantly decreased biliary erythromycin excretion, whereas infusion of dibromosulfophthalein, cefpiramide, ursodeoxycholate-3-O-glucuronide and taurolithocholate-3-sulfate had no effect on biliary excretion of erythromycin. Vinblastine significantly inhibited biliary erythromycin excretion. Phenothiazine treatment significantly increased biliary erythromycin excretion. However, erythromycin infusion did not affect biliary vinblastine excretion. These findings indicate a multiplicity of biliary excretory pathways for organic cations; at least one additonal pathway may exist for organic cations apart from P-glycoprotein. PMID- 10529658 TI - Effect of ozagrel on locomotor and motor coordination after transient cerebral ischemia in experimental animal models. AB - The effect of ozagrel, a selective thromboxane A(2) (TXA(2)) synthetase inhibitor, on the obstruction after cerebral ischemia-reperfusion was studied in experimental animal models. The reduced spontaneously locomotor activity and the obstruction of motor coordination were improved by the administration of ozagrel in the conscious cerebral ischemia-reperfusion mouse model. Ozagrel suppressed the decrease in specific gravity of the brain tissue induced by the occlusion reperfusion in the conscious cerebral ischemia-reperfusion SHR model, and recovered the postischemic decrease in cortical PO(2) after middle cerebral artery occlusion-reperfusion in cats. The level of TXB(2), a metabolite of TXA(2), in the brain increased after the cerebral ischemia-reperfusion, and ozagrel prevented this increase. Additionally, ozagrel also increased the level of 6-keto-PGF(1alpha), a metabolite of prostaglandin I(2) (PGI(2)), in the brain tissue after cerebral ischemia-reperfusion, and the administration of PGI(2) improved the reduced spontaneous locomotor activity in the conscious cerebral ischemia-reperfusion mouse model. Our data suggest that ozagrel suppressed the obstruction following cerebral ischemia-reperfusion by preserving the cerebral blood flow via preventing the increase in TXA(2) and causing an increase in the PGI(2) level. PMID- 10529659 TI - Effect on growth of two different dexamethasone courses for preterm infants at risk of chronic lung disease. A randomized trial. AB - A randomized study was designed to evaluate the effects of two different dexamethasone courses on the growth of preterm infants. The first phase included 30 preterm infants at high risk for chronic lung disease (CLD). 15 babies (moderately early dexamethasone group) were treated with dexamethasone for 14 days, from the 10th day of life, and received a total dose of 4.75 mg/kg; 15 babies were assigned to the control group. The second phase included 30 preterm infants at high risk for CLD. 15 babies (early dexamethasone group) were treated with dexamethasone for 7 days, from the 4th day of life, and received a total dose of 2.38 mg/kg; 15 babies were assigned to the control group. All the main clinical baseline characteristics were similar between the groups both in the first and in the second phase. Infants given the two dexamethasone courses showed significantly reduced weight gain during the period of treatment when compared to the respective control group, but they had a weight catch-up soon after the end of treatment. At 30 days of life the weight and length gain of each treated group were similar to those of control infants, but the moderately early dexamethasone group showed a significantly poorer head growth. No differences between the groups were observed at discharge. Dexamethasone treatment induces a slower weight gain which is time-limited to the period of treatment and is followed by a body weight catch-up. However, the poorer head growth detected at 30 days of life in the infants who received a higher dose of dexamethasone could indicate important adverse effects, possibly dose-related, on postnatal brain growth and development. PMID- 10529660 TI - Changes of superoxide dismutase in cultured rat aortic smooth muscle cells (A7r5) by an incubation of vitamin E. AB - Supplementation of antioxidants such as vitamin E and vitamin C as health promotion food is popular recently. Epidemiological studies supported the beneficial effect of these antioxidants because oxygen free radicals have been linked to the process of diseases and aging. The present study evaluated the effect of alpha-tocopherol (vitamin E) on the changes of superoxide dismutase (SOD) in cultured rat aortic smooth muscle cells (A7r5) after a short-term (2 days) or long-term (7 days) incubation. Incubation of A7r5 cells with vitamin E at a concentration of 50 micromol/l for 2 days caused an increase of both the activity and mRNA level of SOD. At higher concentrations, such as 100 or 200 micromol/l, vitamin E failed to enhance SOD more effectively. However, after incubation for 7 days, vitamin E caused a decrease in both the activity and mRNA level of SOD in a concentration-dependent manner. Otherwise, the protein amount of SOD remained the same in these samples regardless of the concentration of vitamin E or the duration of incubation. The obtained results suggest that vitamin E can increase the effect of SOD to result in the beneficial influence of this antioxidant only at low concentration under a short-term supplementation because a down-regulation of SOD was observed in cells receiving a long-term incubation. PMID- 10529661 TI - Neuroendocrine differentiation in prostatic carcinomas: histogenesis, biology, clinical relevance, and future therapeutical perspectives. AB - Since the introduction of immunohistochemistry, there is an increasing interest in neuroendocrine (NE) differentiation in prostatic carcinomas. Focal NE differentiation in prostatic adenocarcinomas is a very frequent finding. It is subject of numerous studies, since a negative impact on prognosis and an important role in antiandrogen therapy failure are suspected. NE-differentiated small-cell carcinoma is a very rare tumor comprising 0.5-2% of all prostatic carcinomas. Nevertheless, although very rare, it is of clinical importance because it is one of the most aggressive tumors of the prostate with a very poor prognosis. This review is focused on actual concepts of histogenesis, cell biology, clinical implications, and possible future therapeutic perspectives of these two tumor entities. PMID- 10529662 TI - Detection of prostate-specific antigen immunoreactivity in amniotic fluid. AB - OBJECTIVE: To examine whether prostate-specific antigen (PSA) is present in amniotic fluid, whether the amniotic fluid PSA concentration changes with gestational age, and whether there is an association between amniotic fluid PSA and fetal sex. METHODS: The PSA concentration was measured in the amniotic fluid of 48 pregnant women. Thirty-four samples were obtained during routine amniotic fluid analyses performed during gestational weeks 16-18, whereas 14 samples were obtained during cesarean section performed after gestational week 36. RESULTS: PSA was detected in all amniotic fluid samples. The median amniotic fluid PSA was 0.193 ng/ml during gestational weeks 16-18 and 0.39 ng/ml after gestational week 36 (p = 0.1). Furthermore, no significant association was seen between amniotic fluid PSA and fetal sex. The median amniotic fluid PSA level was 0.233 ng/ml for the 21 boys and 0.222 ng/ml for the 27 girls investigated (p = 0.72). CONCLUSIONS: These results confirm recent literature reports that PSA may serve as a growth regulator during normal fetal development. However, further studies are necessary to elucidate the exact role of PSA during fetal development. PMID- 10529663 TI - Flow-cytometric measurement of cellular changes in urine: a simple and rapid method for perioperatively monitoring patients after kidney transplantation. AB - OBJECTIVE: In renal transplant patients having graft dysfunction, it is usually difficult to obtain the accurate diagnosis, such as acute rejection, acute tubular necrosis, infection, or cyclosporin nephrotoxicity. An accurate diagnosis can provide the proper treatment of these patients, thereby lessening the chance of kidney loss. METHODS: A total of 42 patients were enrolled. By using the flow cytometric technique, the white cell populations of urine in these patients were analyzed and linked to their clinical course. All patients underwent sonography guided biopsy of the transplanted kidney with a definitive diagnosis. RESULTS: When 10% lymphocytes and 15% granulocytes in urine were set as the cutoff point of a normal ratio threshold, the flow-cytometric analysis presented the highest sensitivity and the highest negative predictive rate for acute tubular necrosis. However, a lower sensitivity and positive predictive rate was found in acute rejection cases. CONCLUSIONS: Our results suggest that flow-cytometric analysis of the urinary cell population can be used as an adjunct in patient follow-up after kidney transplantation. PMID- 10529664 TI - Nephroureterectomy through a single lumbar incision combined with endoscopic incision of a bladder cuff. AB - Nephroureterectomy is the accepted approach in treating upper urinary tract carcinoma. We present a modification of transurethral resection of the intramural ureter, using endoscopic incision of a bladder cuff around the ureteral ostium as the first step in performing nephroureterectomy through a single lumbar incision. PMID- 10529665 TI - Effect of fleroxacin on mouse bladder carcinogenesis with N-butyl-N- (4 hydroxybutyl) nitrosamine. AB - We recently reported fleroxacin significantly affected cell proliferation in a dose-dependent manner in transitional cell carcinoma cell lines. In this study, we investigated the effect of fleroxacin on mouse bladder carcinogenesis with N butyl-N-(4-hydroxybutyl) nitrosamine (BHBN). Five-week-old C57BL/6 female mice were divided into three groups. The forced oral administrations of fleroxacin (10 or 50 mg/kg/day) were done for 1 week, then the 0.05% BHBN was given for 8 or 12 weeks. Fleroxacin treatments were continued until sacrifice. The mice were sacrificed at 4 weeks or 8 weeks latent period after BHBN administration, and the histological changes in the bladder were examined. Our study suggested that fleroxacin tended to suppress the development of bladder carcinoma or the malignant changes induced by a shorter period of BHBN administration (8 weeks) in mice. However, in the group of the BHBN administration for 12 weeks, there were no significant effects on the prevention of bladder carcinoma in both of the treatment groups with fleroxacin (10 or 50 mg/kg/day). This study did not make it clear that oral administration of fleroxacin suppressed the development of bladder carcinoma induced by BHBN in mice. However, it is very preliminary data and further experiments need to be done with a much higher dose of the fleroxacin or with other kinds of fluoroquinolone antibiotics. PMID- 10529666 TI - Osteopontin expression in prostate cancer and benign prostatic hyperplasia. AB - OBJECTIVES: Osteopontin (OPN), a secreted adhesive glycoprotein, has been shown to be produced in excessive amounts in a variety of experimental models of malignancy. Increased levels of OPN exist in blood from the lungs, breasts, and gastrointestinal tracts of cancer patients with metastases. However, there have been no reports on the expression of OPN in human urological malignancies. The present study investigates the presence of OPN in adenocarcinoma of the human prostate and in benign prostatic hyperplasia (BPH). PATIENTS AND METHODS: Using prostate tissue from 34 patients with primary prostate cancer, 13 patients with prostate cancer after having undergone hormonal therapy, and 12 patients with BPH, formalin-fixed paraffin sections were prepared. Specimens were obtained by needle biopsy or radical prostatectomy. Immunohistochemical staining (ABC method) was then performed. Staining was divided into either negative or positive categories. RESULTS: Positive staining of OPN was observed on cancer cells and macrophages in 52.9% of the primary prostate cancers and 14.5% of the prostate cancers after hormonal therapy. In BPH specimens, 66.7% of the cases displayed positive staining of OPN. The staining level of OPN showed no correlation with serum prostate-specific antigen, but did correlate with stage, differentiation, and Gleason's score. CONCLUSIONS: The result postulates that the expression of OPN is an indicator of cell differentiation; however, it cannot be used as a marker of malignancy in prostate cancer. PMID- 10529667 TI - Unilateral testicular torsion: protective effect of verapamil on contralateral testicular histology. AB - In this experimental study, it was our aim to reduce the effects of ischemic insults to the contralateral testicle after unilateral testicular torsion. The protective effect of a calcium channel blocking agent (verapamil) on the histology and the tubular diameter of contralateral testicle was evaluated. Following a definite period of unilateral testicular torsion (i.e., 4 h), the protective effect of this specific medication was evaluated both after detorsion and orchiectomy procedures. The results of our study demonstrated the protective effect of verapamil on both parameters, especially in animals undergoing orchiectomy. The majority of the specimens demonstrated normal histologic findings together with preserved tubular structures after a 1-week period under verapamil medication. PMID- 10529668 TI - Locally recurrent malignant fibrous histiocytoma: a rare and aggressive genitourinary malignancy. AB - OBJECTIVE: In this study, 22 cases of locally recurrent urological malignant fibrous histiocytoma were reviewed considering therapeutic options, follow-up and prognosis. PATIENTS AND METHODS: In the available literature on this topic we identified 19 cases of locally recurrent genitourinary malignant fibrous histiocytoma. Three additional cases are discussed, primarily arising from the kidney, the bladder and the paratesticular region. RESULTS: The prognosis of locally recurrent urological malignant fibrous histiocytoma was found to be extraordinarily poor. Only 2 of 22 patients have survived for longer than 3.5 years. One of them reported herein is still alive 10 years after extensive lymphatic spread accompanying the first local recurrence. In this case, late local recurrence occurred after an 8-year interval free of disease. CONCLUSION: Malignant fibrous histiocytoma is an unusual urological malignancy with a high rate of local recurrence. The latter is frequently accompanied by metastatic disease and unrelenting progression. Despite the poor prognosis early detection of local failure and aggressive salvage therapy might offer the chance of long term survival in selected cases. Close and life-long follow-up is advisable for patients once treated for recurrent urological malignant fibrous histiocytoma. PMID- 10529669 TI - Renal metastasis from prostatic adenocarcinoma: a potential diagnostic pitfall. AB - We report a case of renal metastases from prostate cancer to show that the possibility of tumor metastasis, although rare, should always be considered in the differential diagnosis of renal mass. PMID- 10529670 TI - Idiopathic clitoral hypertrophy. AB - We report a case of clitoral hypertrophy of unknown origin. A 3-year-old girl showed clitoral hypertrophy at birth. Biochemical analysis of peripheral blood and endocrinological examinations were normal. Other virilizing symptoms were not recognized. Clitoroplasty was carried out, and the clitoris has not enlarged 20 months after the operation. Her mother was administered cefotiam and terbutaline (beta-stimulant) during pregnancy because of threatened abortion. No virilizing agents such as progesterone were used during pregnancy. PMID- 10529671 TI - Infected congenital urethral diverticulum in an adult male. AB - Congenital anterior urethral diverticulum is a rare anomaly. The majority present in infancy with urinary obstruction. Those who present beyond infancy do so on account of recurrent urinary tract infection or dribbling. We present a case of congenital anterior urethral diverticulum manifesting in adulthood with suppuration in the diverticulum. PMID- 10529672 TI - Vesicocutaneous fistula 23 years after hip arthroplasty. A case report. AB - Vesicocutaneous fistula after total hip replacement is a very rare but severe complication, which can appear months or years after operation. Intrapelvic cement (methylmethacrilate) spilling, loosening and dislocation of the prosthesis and infection are believed to be the cause of fistula formation. Only 4 cases of this kind of fistula have been reported in the literature. A new case of vesicocutaneous fistula is presented. The fistula developed 23 years after arthroplasty mainly because of hip-joint infection. Urinary tract symptoms caused by urinary infection appeared only few months earlier. PMID- 10529673 TI - Spontaneous closure of vesicouterine fistula. Account for effective hormonal treatment. AB - OBJECTIVES: To analyze the incidence of spontaneous closure, or non-surgical resolution, of vesicouterine fistula and discuss the resultant implications for the management. METHODS: Review of the literature supplemented by case report of a young woman with spontaneous healing of vesicouterine fistula. RESULTS: This is the 41st patient with spontaneous closure of vesicouterine fistula reported to date. Her clinical course was suggestive of endocrine involvement in the lesion's formation. Spontaneous healing was observed in 5% of 796 vesicouterine fistula cases. Induction of amenorrhea was effective in 8 (89%) of the 9 patients treated, a rate significantly higher (p < 0.001) than that observed without hormonal manipulation (4%). CONCLUSIONS: Conservative management by means of hormonal treatment should be considered before surgical repair. We suggest the role of estrogens and the endometrium in the formation of vesicouterine fistulas. PMID- 10529674 TI - Traumatic dislocation of the testis. AB - We report on a patient having testis dislocated into the superficial crus caused by an accident on a fishing boat. On physical examination, the left hemiscrotum was empty, and a painful mass was palpable inside the left thigh. On sonography, the testis appeared to be normal. Magnetic resonance imaging showed that the mass was located between subcutaneous tissue and muscle layer of the thigh. Surgical intervention was performed, and the left testis was found within the subcutaneous tissue inside the thigh, with slight torsion of the spermatic cord. The testis was fixed in the left hemiscrotum. To our knowledge, traumatic dislocation of the testis into the superficial crus is rare, and only 2 cases have been reported. PMID- 10529675 TI - Penetrating trauma to the scrotum and the corpora cavernosa caused by gunshot. AB - The authors describe a case of gunshot wound of the male genitalia by two low velocity bullets. The first bullet caused a lesion of the right testicle and came out of the right hemiscrotum; the second one had penetrated the left gluteal region with no exit wound. The penile ultrasound confirmed the presence of the bullet at the root of the right corpus cavernosum. The patient underwent exploratory surgery, drainage of the voluminous bilateral scrotal hematoma, and suture of a laceration of the right-testicle cranial portion. Due to the absence of active bleeding, voluminous hematoma and serious injuries in the corpus cavernosum, no surgical removal of the bullet in the right corpus cavernosum was required. The patient regained a normal sexual function 1 month after the operation. PMID- 10529677 TI - Inhibitor development in prospective clinical trials with recombinant factor VIII preparations in previously untreated patients. PMID- 10529676 TI - Epidemiology of inhibitor development in haemophilia A patients treated with virus-inactivated plasma-derived clotting factor concentrates. PMID- 10529678 TI - Other ongoing rFVIII PUP studies. PMID- 10529679 TI - Molecular biological aspects of inhibitor development. AB - Mutation genotype studies have led to a number of interesting observations about the role of the different types of mutations in inhibitor development. Searchable information on mutations, phenotype data, models, etc., are available on the internet at http://europium.mrc.rpms.ac.uk. A relatively high incidence of inhibitors occurs in patients with deletion mutations. Stop mutations are also associated with a high incidence of inhibitors, although there is anomalous distribution of the inhibitors associated with different stop codons. Inhibitors have been associated with a small number of missense mutations. Missense mutations are of great interest from the structure/function viewpoint; in many instances, a missense mutation which results in reduced function of a circulating protein can be mapped onto a model structure and inferences can be made about the effects on the functionality of the protein. A model of the A domains of FVIII is available, but as yet, no structure is available on which to model the C domain of FVIII. Stop codons appear to have a different incidence of inhibitor formation compared to other types of mutations. It is concluded that while genotype mutation studies have led to a number of interesting observations about the role of the different types of mutations in inhibitor development, though more questions have been raised than answers provided to date. PMID- 10529680 TI - Epitope specificity and inactivation mechanisms of factor VIII inhibitor antibodies. AB - The domain specificity of anti-factor VIII (FVIII) inhibitor antibodies was determined in assays using FVIII domains generated by thrombin cleavage or expressed as recombinant polypeptides to neutralise the inhibitor. The results revealed the existence of three major types of inhibitors, and various combinations of these antibodies were found in haemophilic and autoantibody patients. Anti-A2 domain inhibitors prevent normal function of the FVIII/factor IXa (FIXa)/phospholipid complex in an unknown manner. Binding of FVIII to phospholipid and to von Willebrand factor is blocked by anti-C2 domain antibodies, and the binding of FVIII to FIXa is prevented by anti-A3 domain antibodies. A rare type of inhibitor prevents release of activated FVIII from von Willebrand factor (vWf), and another probably interferes with FVIII binding to factor X (FX) because it shares the epitope of a monoclonal antibody with this property. PMID- 10529681 TI - Anti-idiotypic antibodies: from regulation to therapy of factor VIII inhibitors. AB - Evidence has recently accumulated showing that anti-idiotypic antibodies specific to anti-FVIII antibodies are present in the plasma of healthy individuals and of haemophilia A patients with or without inhibitors, where they can neutralise the FVIII inhibitory activity. Additionally, patients successfully desensitised towards FVIII have an increased production of anti-idiotypic antibodies with no significant reduction in anti-FVIII antibodies. We review here possible strategies for modulating the anti-FVIII immune response by idiotypic interactions. PMID- 10529682 TI - International immune tolerance registry, 1997 update. PMID- 10529683 TI - The German National Immune Tolerance Registry, 1997 update. Study Group of German Haemophilia Centres. PMID- 10529684 TI - Analysis of the North American Immune Tolerance Registry (NAITR) 1993-1997: current practice implications. ISTH Factor VIII/IX Subcommittee Members. PMID- 10529685 TI - Comparison of the international immune tolerance registry and the North American immune tolerance registry. AB - Two immune tolerance registries--the International Immune Tolerance Study Group (ITSG) and North American Immune Tolerance Study (NAITS) - are compared and findings from combined data reported. The registries differed with respect to data collection tools, location, host and environmental factors, start date distribution and treatment products. The success and failure rates were similar in the two studies. There was a highly significant association between maximum historical titre and immune tolerance success; the success rate decreased as the historical titre increased. There was a significant association between inhibitor titre immediately prior to treatment and the probability for treatment success, and between outcome and time from diagnosis to treatment in the ITSG (of borderline significance in the NAITS). There was a significant association between outcome and dose, though the direction of the associations was not the same. In the ITSG, success was associated with doses greater than or equal to 200 IU/kg/day, while in the NAITS, greater success was observed with doses of less than 50 IU/kg/day. There was no association between outcome and treatment product. Data from the two registries were combined to produce a table for calculating the chance of successful treatment by historical titre, pretreatment titre, and dose. PMID- 10529686 TI - The use of agents that by-pass factor VIII inhibitors in patients with haemophilia. PMID- 10529687 TI - Treatment of acute bleeding episodes with rFVIIa. PMID- 10529688 TI - Human factor VIII for bleeding in patients with inhibitors. PMID- 10529689 TI - Induction of immune tolerance in haemophilia A inhibitor patients by the 'Bonn Protocol': predictive parameter for therapy duration and outcome. AB - The treatment of inhibitors is one of the most challenging fields in haemophilia care. The present study reports the results of 60 haemophilia A inhibitor patients treated according to the 'Bonn Protocol' and evaluates predictors for the duration and outcome of therapy. Successful immune tolerance could be achieved in 52 patients (86.7%) while the therapy failed in eight patients (13.3%). The immune tolerance achieved was longlasting in all 52 patients, with no inhibitor relapse in up to 20-years follow-up. The course of ITT was influenced by several factors. Interruptions of treatment during the ITT course led to a substantial prolongation of ITT duration (median 39.9 months vs 14.1 months in continuously treated patients). Infections of intravenous central lines appeared to be frequently coincided with ITT prolongation and sometimes even ITT failure. Further negative predictors towards the ITT duration were high inhibitor titres at enrollment or during ITT. There was also a tendency towards longer ITT duration in patients exhibiting the prevalent intron 22 inversion. As a consequence of our data treatment interruptions and infections of intravenous central lines should be avoided during the course of ITT. Furthermore our data suggest, that ITT should be started at low inhibitor titres preferably with a high factor VIII dosage protocol. PMID- 10529690 TI - Immune tolerance protocols using low-dose factor VIII: a review of the literature. AB - Low-dose immune tolerance protocols use doses of factor VIII of less than 50 IU/kg/day to induce immune tolerance. Such protocols are chosen for a variety of reasons including cost, patient acceptability and ease of administration. Early published reports of low-dose protocols appeared in 1981, described small numbers of patients and moderate success. It was proposed that such protocols might modify inhibitor responses from high to low. Later reports are sparse and clinical success is again moderate. Useful treatment and expectation guidelines have emerged from experience with these protocols, and improvements for patients with inhibitors can be achieved. Consideration of immunological success, however, raises doubts about whether real immune tolerance is achieved with such protocols. PMID- 10529691 TI - Extracorporeal immunoadsorption for the treatment of haemophilic patients with inhibitors to factor VIII or IX. AB - BACKGROUND AND OBJECTIVES: This article reviews the relevance of immunoadsorption in the treatment of haemophilic patients with inhibitors. MATERIALS AND METHODS: Immunosorba (sepharose-bound staphylococcal protein A) and Ig-Therasorb (sepharose-bound polyclonal sheep antibodies to human immunoglobulin) columns are suitable for the clinical use of immunoadsorption. They allow the processing of large plasma volumes (>7,000 ml) without relevant side-effects. RESULTS: A haemophilic patient was treated with the Malmo protocol, another was admitted with intracerebral bleeding. Immunoadsorption reduced the inhibitor titer by 70 90%. CONCLUSIONS: Immunoadsorption can be used in cases of acute bleeding, before surgery, acquired factor VIII (FVIII) antibodies, and before the start of immune tolerance therapy. We suggest the inclusion of this method in immune tolerance protocols in order to improve levels, recovery, and half-life of FVIII and to save concentrates. PMID- 10529692 TI - Successful immune tolerance treatment with monoclonal or recombinant factor VIII concentrates in high responding inhibitor patients. AB - Very high purity factor VIII (FVIII) concentrates (plasma-derived or produced by recombinant-DNA technology) were used to achieve immune tolerance in five patients with high-responding inhibitors to FVIII. The mean time required for inhibitor disappearance was 5 months. Four out of five patients showed normalisation of the half-life of infused FVIII after 8-18 months of treatment with 100 IU FVIII/kg body weight administered once daily. Highly purified FVIII products thus appear to be suitable for achieving immune tolerance without negative effects on endogenous von Willebrand factor levels and activity. PMID- 10529693 TI - Factor IX antibody and immune tolerance. AB - Prevalence of FIX inhibitor is ten times lower than factor VIII inhibitor. FIX inhibitor patients pose major challenges for treatment because of the simultaneous occurrence of severe allergy to FIX at the time of inhibitor development. Immune tolerance induction in haemophilia B inhibitor patients with allergy to FIX is complicated due to the development of nephrotic syndrome. Moreover, response to ITI usually is poor. PMID- 10529694 TI - Factor Xa and prothrombin: mechanism of action of FEIBA. AB - A complex consisting of activated factor X (FX) (enzyme) and prothrombin (substrate), both highly purified from human plasma and virus inactivated, was formulated, characterised biochemically as well as in animal studies, and given the name Partial Prothrombinase (PPT). In vitro, PPT shortened the clotting time of a high-titre human factor VIII (FVIII) inhibitor plasma in a manner similar to that of the activated prothrombin complex concentrate FEIBA and triggered coagulation in plasma samples in which factor V (FV) is present. In vivo, the ability of PPT to activate coagulation in both chimpanzees and baboons was equivalent to that of FEIBA. PPT also triggered coagulation in a von Willebrand factor(vWF)-deficient dog and controlled bleeding in rabbits with antibody induced haemophilia A. Thus, studying the mechanism of action of PPT also explains the therapeutic principle of FEIBA. PMID- 10529695 TI - Malmo International Brother Study (MIBS): an international survey of brother pairs with haemophilia. AB - Malmo International Brother Study (MIBS) was initiated in 1996 in order to set up an international registry of twins and non-twin brothers with haemophilia and to search for genetic and compound factors predisposing for inhibitor development. As of July, 1997, 178 brother pairs are registered (143 haemophilia A and 35 haemophilia B patients). Sixteen of these pairs are twins. In 48 of the brother pairs (27%) there is a history of inhibitors, in 25 of them involving only one of the brothers. Immune tolerance induction has been attempted in 13 brother pairs (27%) and in four pairs the inhibitor has been eradicated. Additional demographic data need to be collected and, if possible plasma, IgG and DNA samples will be taken from inhibitor patients to serve as a tool for basic inhibitor experiments. PMID- 10529696 TI - Measuring of factor VIII inhibitors according to the Bethesda method in patients with product-related inhibitors in comparison to non-product related inhibitors. PMID- 10529697 TI - Factor VIII inhibitor-tests could be less sensitive than supposed. PMID- 10529700 TI - Conflict of interest.com. PMID- 10529698 TI - Degradation products of factor VIII which can lead to increased immunogenicity. AB - The biochemical and immunochemical aspects of the development of inhibitors with a plasma-derived, double-virus inactivated factor VIII (FVIII) concentrate (marketed as Octavi SDPlus in Germany and Bisinact in Belgium) are described. A total of 12 cases of inhibitor formation (predominantly type II) were reported in Germany, 8 in Belgium but none in Portugal. Initially, the only difference between the non-pasteurised, SD virus-inactivated product Octavi and the pasteurised product Octavi SDPlus appeared to be pasteurisation, though subsequently, the quality of source material for the product was found to differ in different countries. Separation studies revealed the presence of a 40 kDa peptide fragment in some batches. It was subsequently shown that there was a strong correlation between inhibitor development and batches containing the 40 kDa marker, and a relationship between elevated markers of coagulation activation (FPA in particular) and the occurrence of the 40 kDa marker. Further work revealed that analytical methods commonly used for quality control were not suitable to highlight batch-to-batch differences. It was concluded that inhibitor potential (neoantigenicity) in Octavi SDPlus arose due to two effects; degradation of FVIII already present in source material; and heating of unstable FVIII degradation products. In this case, inhibitors were not caused by the overall production process, nor by GMP failures. The problem of inhibitor potential can be avoided if appropriate preventive measures are taken. Further work is needed to prove non-neoantigenicity and to reinforce the scientific findings, and to characterise pilot batches. PMID- 10529701 TI - Is bladder outlet obstruction normal in elderly men without lower urinary tract symptoms? AB - The aim of the present study was to correlate basic voiding parameters, including uroflowmetry, symptom score, and residual urine volume with the results of pressure-flow studies applying the Abrams/Griffith nomogram, in a series of urologically asymptomatic elderly men. Twenty-nine consecutive male volunteers (median age, 66 years) without past or present urological complaints participated. Fifteen (52%) of the 29 subjectively normal men proved to have bladder outlet obstruction (BOO). Qmax <10 mL/s had a positive predictive value of 100% in diagnosing obstruction, whereas the predictive information of higher flow rates proved very modest. No significant difference existed between obstructed and unobstructed persons at any cutoff value concerning symptom score. The sensitivity as well as the positive predictive value of a residual urine volume >50 mL was zero. It is concluded that a surprisingly high prevalence of BOO in asymptomatic elderly men was demonstrated and that the correlation between pressure flow investigations and alternative diagnostic tests, i.e., flow rate, symptom score, and residual volume was weak in this group of men. It is suggested that a possible explanation for the high frequency of BOO observed in the evaluated asymptomatic men could be that the values defining obstruction have been set too low. Neurourol. Urodynam. 18:545-552, 1999. PMID- 10529702 TI - Editorial comment PMID- 10529703 TI - Variation in urinary flow according to voiding position in normal males. AB - Our objective was to study whether the urinary flow rate would vary according to voiding position. Twenty-one normal healthy male volunteers aged 24 to 40 years (mean, 29 years) were studied. The bed used was designed so that a hole could be opened at its center for voiding, and the bed could be bent at two points so that the subject could void in various positions. Urinary flow was measured with a portable uroflowmeter (P-Flow), which permits measuring urinary flow rate. Each subject assumed five voiding positions (standing, sitting, lateral, supine, and prone) in random order. Urinary flow was measured at least twice in each position to record a stable voiding. For voiding in the lateral position, subjects were instructed to void while bending the upper leg to keep an open angle between the legs. All subjects were also instructed to void without increasing abdominal pressure. Maximum flow rate was 20.7 +/- 6.59 mL/sec with voided volume of 262 +/ 77.8 mL in the lateral, 22.1 +/- 7.05 mLl/sec with voided volume of 309 +/- 130 mL in the supine, 25.0 +/- 8.25 mL/sec with voided volume of 287 +/- 122 mL in sitting, 27.1 +/- 8.89 mL/sec with voided volume of 263 +/- 102 mL in the standing, and 28.7 +/- 10.6 mL/sec with voided volume of 303 +/- 98 mL (mean +/- SD) in the prone positions. The maximum and mean urinary flow rates were greatest in the prone position. With regard to these parameters, significant differences were noted between the prone and lateral positions and between the prone and supine positions. In conclusions, the maximum urinary flow rate was highest in the prone position, followed by the standing, sitting, supine, and finally the lateral positions in normal males. Neurourol. Urodynam. 18:553-557, 1999. PMID- 10529704 TI - Evaluation of the etiology of nocturia in men: the nocturia and nocturnal bladder capacity indices. AB - To determine and quantify the cause of nocturia in men, we describe and evaluate the relative contribution of two complementary indices of nocturia: the nocturia index (Ni), a measure of nocturnal urine overproduction, and the nocturnal bladder capacity index (NBCi), reflective of nocturnal bladder capacity. The records of 100 consecutive men with lower urinary tract symptoms (LUTS), having undergone video-urodynamic studies (VUDS), were prospectively studied. Evaluation included American Urological Association symptom score (AUASS), micturition diary (day, night, and 24-hr voided volume), and VUDS. Voiding diary analysis was carried out as previously described by us, determining the Ni, NBCi, and nocturnal polyuria index (NPi) (nocturnal urine volume/24-hr urine volume). In the case of AUASS question #7 (degree of nocturia), the odds of having a severe AUA question #7 response was found to be 4.09 times higher for patients with NBCi > 2.0 compared with patients whose NBCi was 2 as highly significant in defining diminished NBC as a factor in the etiology of nocturia. In addition, we propose Ni of 1.5 as a threshold greater than which nocturia may be attributed to nocturnal urine overproduction in excess of maximum bladder capacity. Together, these indices describe in quantitative fashion the relative contributions of nocturnal urine overproduction and diminished NBC in identifying the etiology of nocturia in male patients. Neurourol. Urodynam. 18:559-565, 1999. PMID- 10529705 TI - Leaking urine: prevalence and associated factors in Australian women. AB - The Women's Health Australia project provided the opportunity to examine the prevalence of leaking urine and associated variables in three large cohorts of Australian women 18-23 years of age ("young" N = 14,761), 45-50 ("mid-age" N = 14,070), and 70-75 ("older" N = 12, 893). The proportion of women reporting leaking urine was 12.8% (95% CI: 12.2-13.3), 36.1% (35.2- 37.0), and 35% (34.1- 35.9) in each of the three cohorts, respectively. Logistic regression analysis showed significant associations between leaking urine and parity in the young and mid-age women, and between leaking urine and constipation, other bowel symptoms, body mass index, and urine that burns or stings in all three groups. In the mid age and older cohorts, women who reported having both hysterectomy and prolapse repair, or prolapse repair alone, were also more likely to report leaking urine. Lower scores on the physical and mental component summary scores of the medical outcomes survey short form (36 items) questionnaire suggest lower quality of life among women who report leaking urine, compared with those who do not. Neurourol. Urodynam. 18:567-577, 1999. PMID- 10529706 TI - Prevalence of anal incontinence in 409 patients investigated for stress urinary incontinence. AB - A clinical questionnaire concerning anorectal symptoms and urodynamic tests was used to investigate 409 women consulting for stress urinary incontinence. To compare urodynamic data, patients were divided into three groups of women who had either stress urinary incontinence associated with incontinence for formed and/or liquid stools or with gas incontinence, or isolated stress urinary incontinence. To take in account the patients'age for data interpretation, a Mantel-Haenszel test or covariate analysis was performed. Anal incontinence was reported in 114 (28%) of the 409 women investigated. The prevalence of incontinence for gas only, for liquid, or for solid stools was 18.3, 9.3, and 1%, respectively. The duration of gas incontinence was longer than that of fecal incontinence or stress urinary incontinence. Difficult defecation was more frequently observed in patients with double incontinence than in patients with only stress urinary incontinence, and the difference was significant between patients with gas incontinence and patients with stress urinary incontinence (53% versus 37%, P = 0. 03). There was no difference in the number of bowel movements per week among the three groups of patients. The number of vaginal deliveries was surprisingly lower in patients with fecal incontinence associated with urinary incontinence than in others. There was no urodynamic feature that could distinguish patients with urinary incontinence and patients with double incontinence. This study confirmed the close relationship between anal and stress urinary incontinence. Neurourol. Urodynam. 18:579-590, 1999. PMID- 10529707 TI - Validation of cough-induced leak point pressure measurement in the evaluation of pharmacological treatment of stress incontinence. AB - To improve routines in clinical practice and research, it is important that new tests are thoroughly evaluated before they gain widespread application. This includes establishing the reliability and validity of the new test. The purpose of this study was to establish the construct and criterion validity of cough induced leak point pressure (CILPP) measurement. Data on CILPP, maximum urethral pressure (MUP), and a short-term pad test from a phase-I trial of a new pharmacological agent (LS 4416), developed for the treatment of stress incontinence, was used to test the validity of CILPP. Fifteen post-menopausal women with stress incontinence were studied. Phenylpropanolamine (PPA) was used as a positive control. Administration of PPA produced a statistically significant increase in MUP and CILPP. There was a significantly better effect of treatment, expressed as an increase in MUP at 1.5 hr, when PPA was used than with placebo or LS 4416. When CILPP was used to detect change after therapy, PPA produced a significantly greater increase in CILPP than did placebo (least square mean of difference 17.25, P = 0.0202). There was a moderate but statistically significant correlation between CILPP and the short-term Pad Test. Construct validity was demonstrated by the ability of CILPP to detect limited improvement in patients with stress incontinence. Criterion validity was established by the correlation of CILPP to a short-term Pad Test. We propose that, thanks to its greater methodological qualities, leak point pressure measurement should be adopted as a standard method to ascertain the effect of treatment in patients with stress incontinence. Neurourol. Urodynam. 18:591-602, 1999. PMID- 10529708 TI - Simple test of pelvic muscle contraction during pelvic examination: correlation to surface electromyography. AB - The objective of this work was to evaluate the utility of a "Kegel" contraction test in a primary care setting. Fifty-seven adult women completed a questionnaire, underwent pelvic examination, "Kegel" assessment, and measurement of same by vaginal sensor electromyography. Thirty-seven underwent repeat evaluations within 4 weeks. Inter- and intra-rater reliability of digital scale, intra-rater reliability for sEMG measurement, correlation between raters and sEMG, and correlation between scale scores and sEMG with history and pelvic exam were determined. Fifty-six percent were pre-menopausal, 44% post-menopausal. Urinary (62%) and rectal (37%) dysfunction were reported. Inter-rater reliability, intra-observer reproducibility for both raters and sEMG measurements, and correlation between raters and sEMG were significant (P< 0.05). Comparison of continence status and digital scores showed scores /= 90% (P = .00002). Of those with excessive obstruction (PTR > 110%), 32% had detrusor instability (DI) and 47% had emptying phase dysfunction (EPD) compared to 6% and 24%, respectively, of those with PTR /= 90% but 300 mL and maximum bladder capacity >500 mL were significantly higher in patients with abnormal SCV than those with normal SCV (P < 0.03 and 0.001, respectively). Eleven of 16 patients with abnormal MCV showed voiding dysfunction, whereas all patients with normal MCV showed normal voiding (P < 0.001). These results suggest that lower urinary tract symptoms alone cannot predict diabetic vesicourethral dysfunction and that diabetic vesicourethral dysfunction is highly correlated with abnormal nerve conduction velocity. Neurourol. Urodynam. 18:639-645, 1999. PMID- 10529713 TI - Evaluation of morbidity of multi-channel pressure-flow studies. AB - This prospective study was carried out to evaluate the morbidity and complication rate of invasive urodynamics of the lower urinary tract after receiving oral antibiotic prophylactic treatment. A total of 105 patients, 55 men and 50 women, were included in the study and underwent pressure flow study (PFS) as part of the diagnostic assessment. Clinical diagnosis was prostatic obstruction from benign prostatic hyperplasia (BPH) in men and stress urinary incontinence or voiding dysfunction in women. Urine was screened for infection both before and after testing, and the incidence of urinary tract infections (UTI), dysuria, and other complications were assessed at 1-week follow-up to evaluate post-investigation morbidity. Dysuria of mild degree was experienced by 33% of patients, with no significant difference between male and female patients. Post-investigational UTI and fever were reported in 3.6% of men and 4% of women. Six patients had macroscopic hematuria of mild degree. No patient had urinary retention or severe complaints after the investigation and no patient required hospitalization. Post void residual volume was higher in men with BPH obstruction compared to women; a significant difference between post-investigational UTI and residual volume could not be demonstrated (P = 0.8). We conclude that the objective morbidity rate of invasive urodynamic investigation is low. Mild dysuria is common, while severe complications, fever, and hematuria are seldom reported, and the risk of developing UTIs is low with antibiotic prophylaxis, with no significant difference between men and women. Neurourol. Urodynam. 18:647-652, 1999. PMID- 10529714 TI - Collagen injection in the management of post-radical prostatectomy intrinsic sphincteric deficiency. AB - Transurethral injection of collagen is a minimally invasive option for the treatment of urinary incontinence secondary to intrinsic sphincteric deficiency (ISD). We report on the results of transurethral injection in 21 men with urinary incontinence secondary to ISD. Twenty-one consecutive men with a mean age of 69.5 years (range, 51-84), with ISD documented by demonstrating urinary leakage with Valsalva maneuver on physical examination and by video-urodynamic studies were treated with transurethral collagen injection. The etiologies of the incontinence were radical retropubic prostatectomy (RRP) in seven (33.3%), RRP followed by external radiation therapy in seven (33.3%), and transurethral resection of the prostate (TURP) with subsequent RRP in seven (33. 3%). The mean total volume of collagen injected per patient was 18.4 mL (range, 1-44.5). The average number of injections was 2.9 (range, 1-5). The mean follow-up was 12.5 months (range, 1 39). One (5%) patient became dry, 12 (57%) had significant improvement, and eight (38%) had no change. Overall pad use decreased from 2.5 pads/day to 1.68 pads/day, before and after collagen injection (P = 0.014). No difference in outcomes was demonstrated in African American men versus Caucasian American men (P = 0.38), age (<65 and >65 years, P = 0.88), presence of erectile dysfunction, or duration of incontinence (<20 or >20 months, P = 0.71). There were no reported complications. Collagen injection has minimal morbidity and is a viable option for improving incontinence status in men. Neither age, race, erectile function, nor duration of incontinence appears to affect treatment outcome. Neurourol. Urodynam. 18:653-658, 1999. PMID- 10529715 TI - Filling mechanics of obstructed and de-obstructed rat urinary bladders. AB - Chronic bladder distension occurs after partial outlet obstruction and can lead to decompensation and impaired function. To quantify the degree of chronic bladder distension, we previously defined the zero pressure volume (ZPV), the largest contained volume at zero transmural pressure. In the current study, we investigated the short- and long-term effects of outlet obstruction and de obstruction on chronic distension and passive bladder filling mechanics. Voiding patterns were measured 10 days (short term) or 6 weeks (long term) after partial bladder outlet obstruction and the bladders were tested in vitro at that time. De obstructed bladders were obstructed for 6 weeks, and voiding patterns were measured 10 days or 6 weeks after de-obstruction, followed by in vitro testing. Mean voided volume was increased in de-obstructed bladders but not obstructed bladders. The volume of urine in the bladder at euthanasia was greater than mean voided volume in obstructed bladders and less than mean voided volume in de obstructed bladders, indicating large residual urine in the obstructed bladders. ZPV was significantly increased only after long-term obstruction or de obstruction. Similarly, intravesical pressure and mean bladder wall stress were increased only after long-term obstruction or de-obstruction. We conclude that tissue remodeling occurs in the bladder wall after long-term obstruction, possibly both as a result of and leading to chronic overdistension and high residual urine. Tissue remodeling occurs in the bladder wall after long-term de obstruction, possibly due to large voided volumes. Neurourol. Urodynam. 18:659 671, 1999. PMID- 10529716 TI - The nitric oxide pathway in pig isolated calyceal smooth muscle. AB - In pig and humans, whose kidneys have a multi-calyceal collecting system, the initiation of ureteral peristalsis takes place in the renal calyces. In the pig and human ureter, recent evidence suggests that nitric oxide (NO) is an inhibitory mediator that may be involved in the regulation of peristalsis. This study was designed to assess whether the NO synthase/NO/cyclic GMP pathway modulates the motility of pig isolated calyceal smooth muscle. Immunohistochemistry revealed a moderate overall innervation of the smooth muscle layer, and no neuronal or inducible NO synthase (NOS) immunoreactivities. Endothelial NOS immunoreactivities were observed in the urothelium and vascular endothelium, and numerous cyclic GMP-immunoreactive (-IR) calyceal smooth muscle cells were found. As measured by monitoring the conversion of L-arginine to L citrulline, Ca(2+)-dependent NOS activity was moderate. Assessment of functional effects was performed in tissue baths and showed that NO and SIN-1 decreased spontaneous and induced contractions of isolated preparations in a concentration dependent manner. In strips exposed to NO, there was a 10-fold increase of the cyclic GMP levels compared with control preparations (P < 0.01). It is concluded that a non-neuronal NOS/NO/cyclic GMP pathway is present in pig calyces, where it may influence motility. The demonstration of cyclic GMP-IR smooth muscle cells suggests that NO acts directly on these cells. This NOS/NO/cyclic GMP pathway may be a target for drugs inhibiting peristalsis of mammalian upper urinary tract. Neurourol. Urodynam. 18:673-685, 1999. PMID- 10529717 TI - Recording the evoked canine detrusor electromyogram. AB - With increasing interest in detrusor disorders and possible detrusor myopathies, a method for recording the detrusor electromyogram (detrusor-EMG) would probably greatly assist the diagnosis of various bladder dysfunctions. Therefore, we investigated the electromyographic activity of the detrusor during sacral root stimulation and during spontaneous bladder contractions in six anesthetized dogs. In all experiments, a high correlation of detrusor-EMG recordings with bladder contraction was observed. Analysis in the time domain and power spectrum analysis revealed the most clear correlation of detrusor-EMG with intravesical pressure rise in a frequency band above 3 Hz. The spike duration was 100 to 250 ms with an amplitude of 100 to 500 microV. Low-frequency activity below 1 Hz was mainly presumed to be artifacts due to fluid movement under the electrode. Our trials indicate that smooth muscle EMG recordings from the detrusor smooth musculature are possible. The exact physiological relevance of the signal in the sub-Hertz domain (<1 Hz) is still uncertain. The presented animal model allows the pathophysiologic investigation of various pathologies of bladder dysfunction and detrusor myopathies. Neurourol. Urodynam. 18:687-695, 1999. PMID- 10529718 TI - Fatty acid profiles in normal and obstructed rabbit bladder smooth muscle and mucosa. AB - Partial bladder outlet obstruction results in progressive loss in contractile and specific cellular and subcellular membrane functions. There is evidence that ischemic activation of proteolytic and lipolytic enzymes play a major role in the etiology of bladder dysfunction secondary to partial outlet obstruction. The specific aims of the current study were to determine the fatty acid profiles in normal rabbit bladder smooth muscle and mucosa and to determine the effect of partial outlet obstruction on the distribution and content of free and total fatty acids. Fatty acids were isolated by extraction from obstructed and normal bladder smooth muscle and mucosal homogenates, and samples were analyzed by gas chromatography. All samples contained palmitic, stearic, oleic, linoleic, and arachidonic acids. A 100% increase in total fatty acid concentration was observed in the obstructed bladder muscle tissue relative to normal bladders, although the concentration of total arachidonic acid remained constant in the two groups. Significantly higher levels of free arachidonic acid were observed in the obstructed bladder muscle group compared to the normal group. No changes were observed in fatty acid concentrations or distributions in bladder mucosa. These data show that fatty acid composition is altered as a result of bladder obstruction and support the idea that obstruction increases the activity of lipase activity and/or decreases acyl transferase activity. Neurourol. Urodynam. 18:697-711, 1999. PMID- 10529719 TI - Medial prefrontal cortical output neurons to the ventral tegmental area (VTA) and their responses to burst-patterned stimulation of the VTA: neuroanatomical and in vivo electrophysiological analyses. AB - During a delayed period in a delayed-response task, prefrontal cortical neurons show a change in neuronal firing rate that is dependent on a functional mesocortical dopamine input. This change in firing rate has been attributed to be part of the cellular processes underlying working memory. However, it is unclear what neural mechanisms activate mesocortical dopamine neurons to provide an optimal level of dopamine to modulate the firing of the medial prefrontal cortical (mPFC) neurons. This study examined the possibility of whether mPFC neurons that project to the ventral tegmental area (VTA) might activate the ascending mesocortical dopamine neurons. To determine the locations of the mPFC- >VTA neurons, cholera toxin subunit B was microinjected into the VTA. Retrogradely labeled mPFC neurons mainly reside in the deep lamina V and VI. In vivo single unit recording in urethane-anesthetized rats were also used to determine the responses of some of these neurons to burst-patterned stimulation of the VTA. Single-pulse stimulation (1 Hz) of the VTA antidromically activated burst firing mPFC-->VTA neurons. In response to burst-patterned stimulation of the VTA, which mimicked burst firing of VTA dopamine neurons (4-10 pulses at 10 15 Hz cycled at 0.5-3 Hz), the temporal structure of spontaneous burst firing patterns of these neurons but not their mean firing rate were changed. However, the mean firing rate of the non-VTA projecting neurons (i.e., no antidromic response to VTA stimulations) was either increased or decreased by similar burst patterned stimulation of the VTA. These data suggest that burst-patterned stimulation of the ascending VTA-->mPFC or putative mesocortical dopamine neurons might have released dopamine and/or other neuromodulators to modulate the temporal code, rather than the rate code, of mPFC-->VTA neurons. Medial PFC neurons that project elsewhere (e.g., nucleus accumbens or mediodorsal thalamus) may mediate the sustained firing rate changes during, e.g., short-term working memory. PMID- 10529720 TI - Opioid peptide receptor studies. 10. Nor-BNI differentially inhibits kappa receptor agonist-induced G-protein activation in the guinea pig caudate: further evidence of kappa receptor heterogeneity. AB - There is strong evidence supporting the existence of multiple kappa receptors. Previous studies proposed that U69,593 and (+)-tifluadom act on different kappa receptor subtypes, kappa(1) (kappa(1)) and kappa(2) (kappa(2)), respectively. In this study, we investigated the effects of the kappa selective antagonist nor binaltorphimine (Nor-BNI) on U69,593- and (+)-tifluadom-induced receptor-mediated stimulation of [(35)S]-GTP-gamma-S binding in the guinea pig caudate. The IC(50) value of Nor-BNI in the presence of a stimulating concentration of U69,593 (1 microM) was 0.19+/-0.02; while the IC(50) for Nor-BNI in the presence of (+) tifluadom (1 microM) was 13.9+/- 1.62 nM. The mu-opioid receptor antagonist CTAP (10,000 nM) significantly reduced (+)-tifluadom-stimulated [(35)S]-GTP-gamma-S binding in rat brain sections and guinea pig brain membranes, indicating that (+) tifluadom has mu agonist activity. Under conditions in which the mu agonist activity of (+)-tifluadom was blocked by 1000 nM CTAP the Ki value for Nor-BNI for inhibition of U69,593-stimulated [(35)S]-GTP-gamma-S binding was 0.036+/-.004 nM, whereas, its Ki value for the (+)-tifluadom-stimulated [(35)S]-GTP-gamma-S binding was 0.27+/-.015 nM. These results suggest that (+)-tifluadom and U69,593 activate pharmacologically different receptors. This study provides functional evidence in support of kappa receptor heterogeneity. PMID- 10529721 TI - Differential effects of haloperidol and clozapine on ionotropic glutamate receptors in rats. AB - Despite multiple lines of investigation the effect of neuroleptics on glutamate mediated neurotransmission remains controversial. To study the effects of typical and atypical neuroleptics on selected parameters of glutamate-mediated neurotransmission, male Sprague-Dawley rats were randomly assigned to a 21-day oral treatment course with vehicle, haloperidol (HDL), or clozapine (CLZ). Coronal slices of rat brain were then incubated with tritiated ligands to measure NMDA, AMPA, and kainate receptor, and glutamate reuptake site density. Regions of interest included the frontal cortex, anterior cingulate cortex, dorsal striatum, ventral striatum, and the nucleus accumbens. CLZ increased the density of AMPA receptors significantly in the frontal and anterior cingulate cortices compared with normal controls. In the dorsal and ventral striatum, and nucleus accumbens as a whole, CLZ-treated rats had a higher AMPA receptor density compared with both the HDL- and vehicle-treated controls. Additionally, within the nucleus accumbens, CLZ-treated rats had a higher density of AMPA receptors compared with the HDL group in the core, and at trend level in the shell. There was a group by region interaction for NMDA receptor density, primarily reflecting the tendency of HDL treated rats to have high receptor densities in the frontal and anterior cingulate cortices. Kainate receptors and glutamate reuptake site densities did not differ significantly across groups. These results suggest a critical role for glutamate in the mediation of atypical antipsychotic drug action in anatomically specific regions, and further encourage the investigation of glutamate neurotransmitter systems in schizophrenia. PMID- 10529722 TI - Assessment of the serotonin reuptake blocking property of YM992: electrophysiological studies in the rat hippocampus and dorsal raphe. AB - YM992 is a selective serotonin (5-HT) reuptake inhibitor and a 5-HT(2A) antagonist with potential antidepressant activity. As expected from a 5-HT reuptake inhibitor, which induces an accumulation of 5-HT in the dorsal raphe, YM992 inhibited the firing activity of these 5-HT neurons (ED50: 2.0+/-0.2 mg/kg, i.v.). This effect was reversed by the 5-HT(1A) antagonist WAY 100635. YM992 also dose-dependently prolonged the time for CA3 neurons to recover 50% of their firing rate following microiontophoretic applications of 5-HT, a reliable index of the function of the 5-HT reuptake carrier. In a second series of experiments, the adaptative properties of 5-HT neurons were examined during sustained administration of YM992 (20 mg/kg/day, s.c., delivered by osmotic minipumps) after 2 days of treatment. YM992 decreased by more than 60% the firing activity of the 5-HT neurons. There was a partial recovery of firing after 7 days and a complete one after 14 days of treatment in the presence of the minipump still delivering the drug. In a third series of experiments, the sensitivity of pre- and postsynaptic 5-HT(1A) receptors in the dorsal raphe and the dorsal hippocampus were assessed. The results showed that YM992 attenuated the inhibitory effect of intravenous administration of LSD and the 5-HT(1A) agonist 8 OH-DPAT on the firing activity of 5-HT neurons. As did the selective 5-HT reuptake inhibitor fluvoxamine, YM992 markedly increased the effectiveness of the electrical stimulation of ascending 5-HT fibres on firing activity of the postsynaptic hippocampus pyramidal neurons. This enhancement of 5-HT neurotransmission by YM992 was attributable to a desensitization of the terminal 5-HT(1B) autoreceptors since the postsynaptic 5-HT(1A) receptors in the hippocampus remained normosensitive. PMID- 10529723 TI - Kinetic and equilibrium analyses of [(123)I]epidepride binding to striatal and extrastriatal dopamine D(2) receptors. AB - Quantitative SPECT measures of dopamine D(2) like receptors with [(123)I]epidepride is complicated by its high affinity and lipophilic metabolites. The purpose of this study was to use both parent (P) and lipophilic metabolites (M) as input functions in a kinetic paradigm and in comparison to the results of equilibrium studies. Kinetic studies on eleven healthy human subjects, ages 32+/- 10 were performed following i.v. injection of approximately 370 MBq of [(123)I]epidepride. Images were acquired for 13.5+/-1.0 hours. Equilibrium studies were done on seven of eleven subjects with a bolus injection of approximately 140 MBq, bolus/infusion ratio of 10 hours, and infusion for 30-32 hours. High (striatum) and low (temporal cortex) density regions were studied. Two (P and M) and one (P) input function models were applied in the kinetic studies. In receptor-rich regions, the distribution volumes in nondisplaceable compartments were fixed to those in cerebellum. In addition, in the two input function model, K(1)(P)/K(1)(M) was fixed to the values in the cerebellum. The one input function model provided V'(3) values (=f(1)*B'(max)/K(D)) which were consistent with those obtained in equilibrium studies in both receptor-rich regions, while the two input function model provided consistent values only in striatum. Poor identifiability of the rate constants of metabolites seemed to be the source of errors in the two input function model. These results suggest that correct V'(3) values can be obtained with the one input function model both in high- and low-density regions. PMID- 10529724 TI - Methamphetamine- and 1-methyl-4-phenyl- 1,2,3, 6-tetrahydropyridine-induced dopaminergic neurotoxicity in inducible nitric oxide synthase-deficient mice. AB - Previous studies have suggested a role for the retrograde messenger, nitric oxide (NO), in methamphetamine (METH)- and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)- induced dopaminergic neurotoxicity. Since evidence supported the involvement of the neuronal nitric oxide synthase (nNOS) isoform in the dopaminergic neurotoxicity, the present study was undertaken to investigate whether the inducible nitric oxide synthase (iNOS) isoform is also associated with METH- and MPTP-induced neurotoxicity. The administration of METH (5mg/kg x 3) to iNOS deficient mice [homozygote iNOS(-/-)] and wild type mice (C57BL/6) resulted in significantly smaller depletion of striatal dopaminergic markers in the iNOS(-/-) mice compared with the wild-type mice. METH-induced hyperthermia was also significantly lower in the iNOS(-/-) mice than in wild-type mice. In contrast to the outcome of METH administration, MPTP injections (20 mg/kg x 3) resulted in a similar decrease in striatal dopaminergic markers in iNOS(-/-) and wild-type mice. In the set of behavioral experiments, METH-induced locomotor sensitization was investigated. The acute administration of METH (1.0 mg/kg) resulted in the same intensity of locomotor activity in iNOS(-/-) and wild-type mice. Moreover, 68 to 72 h after the exposure to the high-dose METH regimen (5 mg/kg x 3), a marked sensitized response to a challenge injection of METH (1.0 mg/kg) was observed in both the iNOS(-/-) and wild-type mice. The finding that iNOS(-/-) mice were unprotected from MPTP-induced neurotoxicity suggests that the partial protection against METH-induced neurotoxicity observed was primarily associated with the diminished hyperthermic effect of METH seen in the iNOS(-/-) mice. Moreover, in contrast to nNOS deficiency, iNOS deficiency did not affect METH-induced behavioral sensitization. PMID- 10529725 TI - Stimulation of dopa decarboxylase activity in striatum of healthy human brain secondary to NMDA receptor antagonism with a low dose of amantadine. AB - The efficacy of amantadine in alleviating motor symptoms of Parkinson's disease may be mediated in part by stimulation of cerebral dopa decarboxylase (DDC) activity, secondary to antagonism of N-methyl-D-aspartate (NMDA) type glutamate receptors. We tested the specific hypothesis that amantadine increases the decarboxylation rate of 6-[(18)F]fluoro-L-DOPA (FDOPA), an exogenous substrate for DDC, in healthy human brain. Radioactivity concentrations in brain tissue of neurologically normal volunteers (n = 5) injected intravenously with FDOPA ( approximately 4.5 mCi) were recorded by positron emission tomography (PET) for 120 min, first in a baseline condition, and again following three consecutive days of treatment with amantadine (100 mg/day, p.o.). Data from four telencephalic regions of interest containing appreciable DDC activity were analyzed with the tissue slope-intercept plot, using cerebellar cortex as the reference tissue, to estimate a coefficient of in situ FDOPA decarboxylation (k(3)(r), min(-1)). Mean estimates of k(3)(r) were increased following amantadine treatment in caudate nucleus (+12%), putamen (+28%), ventral striatum (+27%), and frontal cortex (+9%). For an initial confidence level of 95%, paired one-sided Student's t-tests with Bonferroni correction for multiple comparisons revealed a statistically significant drug effect in ventral striatum. Present results are consistent with stimulation of DDC activity in striatum of healthy human brain secondary to NMDA receptor antagonism with a low dose of amantadine, and suggest that this response is an important mechanism underlying the anti-parkinsonian properties of amantadine. Nonetheless, PET studies in parkinsonian patients using higher, clinically effective doses of amantadine may reveal more pronounced enhancements of cerebral DDC activity. PMID- 10529726 TI - An Editor's thank You PMID- 10529727 TI - Preface PMID- 10529728 TI - UVB-induced gpt mutations in the skin of gpt delta transgenic mice. AB - Ultraviolet light B (UVB)-induced mutagenesis was studied in gpt delta transgenic mice, which contain the lambdaEG10 shuttle vector as a transgene. The mice were exposed to UVB at single doses of 0.3, 0.5, 1.0, 1.5, and 2.0 kJ/m(2). At 4 weeks after irradiation, the mutant frequencies (MF) of the gpt gene were determined in the epidermis and the dermis, and the gpt mutations in the epidermis were identified by DNA sequencing. The epidermis exhibited a higher sensitivity to UVB than the dermis at doses of 0.3 and 0.5 kJ/m(2) UVB: the MF of the epidermis were more than nine times higher than those of the nonirradiated mice, whereas the MF of the dermis were only two to three times higher than the nonirradiated level at the doses used. The UVB-induced mutation spectrum in the epidermis was dominated by G:C to A:T transitions at dipyrimidine sites, such as 5'-TC-3', 5'-CC-3', and 5'-T/C-CG-3'. Tandem transitions such as CC to TT were also observed. Interestingly, a remarkable bias towards the template strand of the gpt gene was observed in the single transitions at 5'-TC-3' and 5'-CC-3' sites, but not at 5' T/C-CG-3' site. In contrast, G:C to A:T transitions at CpG sites and deletions were observed in nonirradiated mice. Hot spots of transitions were observed at different sites in UVB-irradiated and nonirradiated mice. These results indicate that gpt delta transgenic mouse is a suitable model for studying in vivo UVB induced mutations at the molecular level. PMID- 10529730 TI - Heterozygous Aprt mouse model: detection and study of a broad range of autosomal somatic mutations in vivo. AB - During the development of cancer a series of specific genetic alterations have to occur in a stepwise fashion to transform a normal somatic cell into a malignant tumor cell. These genetic changes can be roughly divided in two groups: mutations in proto-oncogenes that result in a constantly activated gene product and mutations in tumor-suppressor genes that result in loss of function. While oncogenic mutations often have a dominant phenotype and mutation of one allele is sufficient for activation, in general both alleles of a tumor suppressor gene have to be disrupted to abolish its function. The requested specificity for activating mutations in proto-oncogenes is high, since only a limited number of mutations at specific sites result in an activated protein. In contrast, disruption of a tumor suppressor gene can be accomplished via various mechanisms. Familial cancers often contain a germline mutation in one allele of a tumor suppressor gene. In tumors, the second allele is then frequently lost by genetic alterations that also affect the heterozygous state of multiple loci adjacent to the tumor suppressor gene. Genetic events especially, such as mitotic recombination, chromosome loss and deletion, are frequently responsible for the loss of the functional allele of heterozygous mutant tumor suppressor genes. We generated an Aprt(+/-) mouse model that allows us to study in detail the nature of the alterations that lead to loss of the wild-type Aprt allele in somatic cells. These genetic changes are thought to be analogous to those occurring at autosomal tumour suppressor genes, where they may contribute to the development of cancer. Furthermore, this mouse model allows determination of the extent and mechanisms by which chemical carcinogens induce loss of heterozygosity and identification of the nature of the DNA adducts responsible. PMID- 10529731 TI - Characterization of new transgenic Big Blue(R) mouse and rat primary fibroblast cell strains for use in molecular toxicology studies. AB - We have established and characterized primary mouse and rat cell strains for studies designed to complement in vivo gene mutation assays using the Big Blue(R) mouse or rat. Primary fibroblast cell strains, designated BBM1 and BBR1, were derived from a transgenic male Big Blue(R) B6C3F1 mouse and from a male Big Blue(R) Fischer-344 rat, respectively. Both BBM1 and BBR1 are genetically stable and mostly diploid. Both cell strains have low spontaneous frequencies of mutation at the lacI and cII loci as well as low frequencies of sister chromatid exchange and micronuclei formation. In addition, N-ethyl-N-nitrosourea (ENU) induces mutations at the cII locus in both BBM1 and BBR1 cells. These new primary Big Blue(R) mouse (BBM1) and rat (BBR1) fibroblast cell strains represent useful new models for molecular toxicology studies. Environ. Mol. Mutagen. 34:90-96, 1999 Published 1999 Wiley-Liss, Inc. PMID- 10529729 TI - Mutational spectrum of dimethylnitrosamine in the liver of 3- and 6-week-old lacI transgenic mice. AB - We determined the spectrum of mutations in the lacI gene in the liver of Big Blue(R) transgenic mice after exposure to five daily doses of 2 mg/kg dimethylnitrosamine (DMN) at 3 and 6 weeks of age. This dose has been reported to increase the mutant frequency 9-fold when the animals are 3 weeks old. The lacI mutations recovered when treated at 3 weeks consist of mainly G:C --> A:T transitions, predominantly at non-CpG sites, and thus are consistent with mutagenesis by DMN. No increase in mutant frequency was reported when the mice were treated at 6 weeks of age. As we have previously shown that changes in mutational spectrum can be detected even when no statistically significant increase in mutant frequency is seen, we also examined the spectrum after treatment at 6 weeks. No changes from the spontaneous spectrum were detected. The comparison of the outcome of DMN treatment at 3 and 6 weeks confirms a change in metabolic activation, adduct removal, or mutation fixation between 3 and 6 weeks of age. PMID- 10529732 TI - Molecular nature of mutations induced by a high dose of x-rays in spleen, liver, and brain of the lacZ-transgenic mouse. AB - DNA sequences of 103 spontaneous mutants and 102 X-ray-induced mutants of the lacZ transgene from spleen, liver, and brain of the MutaMouse were examined and compared to elucidate characteristics of radiation-induced mutations in vivo. The radiation-induced mutants were isolated from genomic DNA of each tissue collected at 3.5 days after 200 Gy of whole body irradiation. Base substitution was predominant (80% or more) in nonirradiated tissues, while deletion was prevalent (about 55%) in irradiated tissues. The other types of mutation appeared at similar frequencies in both control and irradiated tissues. The size of the deletions was smaller than 438 nucleotides, with a predominance of one basepair deletions in both control and irradiated tissues. A close look at the nucleotides at the deletion endpoints revealed that many of the radiation-induced deletions did not have repeated sequences at the break point termini, whereas all deletions found in unirradiated tissues showed one or more bases of repeated sequences at the termini. Further, eight complex-type deletion mutations were found only in irradiated tissues. Comparison among the three types irradiated tissue did not reveal any tissue-specificity. The data indicate that the molecular nature of mutations induced in tissues with ionizing radiation is different from that of spontaneous mutations. PMID- 10529733 TI - Efficient detection of deletions induced by a single treatment of mitomycin C in transgenic mouse gpt delta using the Spi(-) selection. AB - Transgenic rodent mutation assays permit the detection and molecular analysis of various types of gene mutations, such as base changes and frameshifts, in a number of tissues. It is reported, however, that deletion mutations are not efficiently detected by the assays, in particular those using lambda phage shuttle vectors. Recently, a new transgenic mouse model, i.e., gpt delta, has been developed to selectively detect some types of deletions by Spi(-) selection. Spi(-) selection has an advantage over the other selections to preferentially identify deletions because only lambda phages deficient in both the red and gam gene functions are allowed to form phage plaques. In this study, we examined whether in vivo deletions induced by the treatment of mitomycin C (MMC) are detectable by the Spi(-) assay in the mouse model. The mice were treated with MMC (0.5, 1.0, 2.0, and 4.0 mg/kg, single intraperitoneal injection) and sacrificed 14 days after the dosing. The treatment at 4.0 mg/kg approximately doubled the mutant frequency of Spi(-) in the bone marrow, i.e., 2.52 x 10(-6) vs. 1.31 x 10( 6). The molecular analyses using polymerase chain reaction (PCR) and DNA sequencing indicated that seven Spi(-) mutants at 4.0 mg/kg group had deletions with molecular sizes from 0.8 kilo basepairs (kb) to 8.5 kb, whereas no such deletions were observed in the Spi(-) mutants in the control group. The results suggest that deletions induced by MMC in the bone marrow are efficiently detectable by Spi(-) selection and also that the molecular analyses are useful to evaluate the significance of a marginal increase in mutant frequency in the transgenic rodent mutation assays. PMID- 10529734 TI - Background mutations and polymorphisms in lacZ-plasmid transgenic mice. AB - Transgenic animal models harboring chromosomally integrated shuttle vectors with bacterial reporter genes are now widely used to measure in vivo mutant frequencies. The lacZ-plasmid transgenic mouse model has a unique sensitivity to large rearrangements compared to systems using bacteriophage lambda vectors, which mainly detect point mutations and small deletions or insertions. In this study, the background mutant frequencies and spectra in the lacZ-plasmid transgenic mouse model were investigated. While the majority of the recovered lacZ-mutants appeared to have originated in the mouse, a subset of mutants are likely to represent artifacts, and occur with a frequency of about 1.3 x 10(-5), irrespective of the total mutant frequency. Galactose-insensitive host cells, due to galE back mutations or galK or galT forward mutations, grow through the positive selection system and cause a small subset of the background. When using HindIII to excise the plasmids from genomic DNA, the largest contribution to the background, (1.1 +/- 0.3) x 10(-5), appeared to be caused by star activity, i.e., cleavage at nucleotide sequences other than the HindIII restriction enzyme recognition sequence, during the recovery procedure. Finally, a total of 10 polymorphic sites in different copies of the lacZ-plasmid cluster in founder line 60 were discovered. PMID- 10529735 TI - Partial hepatectomy strongly increased the mutagenicity of N-ethyl-N-nitrosourea in MutaMouse liver. AB - The relationship between cell proliferation and mutagenesis can be investigated in vivo due to the advent of transgenic animal mutation assays. In these assays, slowly proliferating tissues, such as mammary gland and liver, that are exposed to mutagens generally have longer manifestation times for mutations and lower mutant frequencies. This may be because the cells have enough time prior to cell division to repair DNA damage. We carried out this study using the MutaMouse positive selection system to investigate the effect of a high rate of cell proliferation induced by partial hepatectomy (PH) on mutation induction. We used a 2 x 2 experimental design for PH (or no PH) and 50 mg/kg N-ethyl-N-nitrosourea (ENU) (or phosphate buffer), with the chemical injected 16-19 hr after PH. In the non-ENU groups, the mean MF was slightly but not significantly higher in the PH group than in the non-PH group. In the ENU non-PH group, the MF was also slightly but not significantly increased. In the ENU PH group, in contrast, the mean MF was 7 times the mean MF of the group that received either treatment alone. These results strongly support the hypothesis that ENU induced pre-mutational DNA lesions in liver are completely repaired prior to cell division, and PH increases the mutagen-induced MF by reducing the amount of time available for such repair. PMID- 10529736 TI - Effect of heterozygous loss of p53 on benzo[a]pyrene-induced mutations and tumors in DNA repair-deficient XPA mice. AB - XPA-deficient mice have a complete deficiency in nucleotide excision repair, and as such they display a cancer predisposition after exposure to several carcinogens. Besides being sensitive to genotoxic agents applied to the skin, they are also susceptible to human carcinogens given orally, like benzo[a]pyrene (B[a]P). To study the role of the tumor suppressor gene p53 in DNA repair, gene mutation, and tumor induction, we crossed XPA-deficient mice with p53 knockout mice and lacZ (pUR288) gene marker mice. When treated orally (by gavage) with B[a]P, the XPA(-/-)/p53(+/-) double transgenic mice developed tumors much earlier and with higher frequency compared to their single transgenic counterparts. The major tumor type found in all genotypes was generalized lymphoma mainly residing in the spleen; several sarcomas were observed in p53(+/-) and XPA(-/-)/p53(+/-) mice. Next, we determined lacZ mutation frequencies in several (non)target tissues. It appeared that in the spleen (the major tumor target tissue) of XPA(-/ ) and XPA(-/-)/p53(+/-) mice the lacZ mutation frequency was significantly elevated (80-100 x 10(-5)), and was two times higher as found in spleens of B[a]P treated WT and p53(+/-) mice (P = 0.003). In nontumor target tissues like liver and lung, we found a moderate increase in the lacZ gene mutation frequency (30-40 x 10(-5)), which was independent of the genotype. The results obtained with the DNA-repair deficient XPA mice indicate that a significantly increased lacZ mutation frequency in a particular organ/tissue is an early marker for tumor development at later stages at the same site. However, the synergistic effect of a XPA(-/-)- and a p53(+/-)-deficiency in tumor development is not reflected by an absolute increase in the lacZ mutation frequency in the major tumor target tissue of XPA(-/-)/p53(+/-) or p53(+/-) mice compared to that of XPA(-/-) and WT mice, respectively. PMID- 10529737 TI - Analysis of gene mutations and clastogenicity following short-term treatment with azathioprine in MutaMouse. AB - The mutagenicity and clastogenicity of the immunosuppressive drug azathioprine (AZA), a multitissue rodent carcinogen and IARC-classified human carcinogen, was investigated using transgenic lacZ mice (MutaMouse). Male animals (n = 5 per group) were dosed with AZA (10, 50, 100 mg/kg p.o. daily for 5 days), vehicle (n = 10), or the positive control, chlorambucil (15 mg/kg i.p., n = 3), and killed 24 hr or 25 days after the last treatment. Micronucleus assays were performed with bone marrow (24-hr samples) or peripheral blood (24-hr and 25-day samples) and DNA was extracted from bone marrow and liver for gene mutation analysis at the transgenic lacZ locus. AZA induced 5.3-111.3-fold increases in %MNPCE (P < 0.01) in bone marrow compared with vehicle control, accompanied by 4.4-5. 6-fold increases in %MNRETs (P < 0.01) in peripheral blood. Chlorambucil caused a 14.5 fold increase in %MNRET and there was evidence of significant stem cell toxicity in both positive control and AZA treatment groups. By day 25, however, there was evidence of substantial recovery of the bone marrow as determined by the frequency of RET, and the %MNRET in all treatment groups was the same as the vehicle control. Analysis of lacZ MF showed 1.4-1.6-fold increases in AZA 24-hr bone marrow samples, increasing to approximately 2.0-fold above concurrent controls by day 25 (medium dose P < 0.05, high dose P < 0.01). For liver, there was a 2-fold increase in MF (P < 0.05) in the 24-hr sample at the highest dose only, and increases of 1.3-1.5-fold by day 25 in the medium (P < 0. 05) and high (P = 0.055) dose groups, respectively. The positive control, chlorambucil, induced 2-3-fold increases (P < 0.01) in mean MF in both bone marrow (25-day sample) and liver (24-hr and 25-day samples). These data confirm the clastogenicity of AZA in the mouse, and show that this compound induces gene mutations in bone marrow and liver, in vivo, at the highest dose and supports the view that AZA is a genotoxic carcinogen. PMID- 10529738 TI - Treatment of lacZ plasmid-based transgenic mice with benzo[a]pyrene: measurement of DNA adduct levels, mutant frequencies, and mutant spectra. AB - The evaluation of the relationship between the dose to DNA of a genotoxic carcinogen and in vivo somatic cell mutagenesis may provide important information on the mechanisms leading to induced mutational events. The induction of DNA adducts and DNA mutations in different tissues of lacZ plasmid-based transgenic mice has been investigated following a single intraperitoneal administration of the polycyclic aromatic hydrocarbon benzo[a]pyrene (B[a]P). DNA adducts were measured by (32)P-postlabeling at times between 1 and 28 days following injection and DNA mutations in the lacZ reporter gene were quantified using a highly efficient immunomagnetic purification of the lacZ-containing pUR288 plasmid, followed by electrotransformation of circularized plasmids into a host restriction negative E. coli C (DeltalacZ, galE (-)) host. The major DNA adduct formed by B[a]P, N (2)-(10beta-[7beta,8alpha, 9alpha-trihydroxy-7,8,9, 10 tetrahydrobenzo(a)pyrene]yl-deoxyguanosine, reached maximum levels between 5 and 7 days followed by a gradual decrease. The induced mutant frequency reached a maximum between 7 and 14 days, after the DNA adduct level in a particular tissue had reached its apparent maximum between 5 and 7 days. The mutant spectrum, defined as the percentage of no-change and size-change mutations in a particular tissue, changed from predominantly size-change mutations in untreated tissues to predominantly no-change mutations in tissues displaying the highest B[a]P-induced mutant frequencies. Contrary to the assumption that there are no factors that can eliminate mutations once they exist, it was observed that mutant frequencies in the organs studied declined to background levels, which was accompanied by a change in the mutant spectrum from predominantly no-change mutations to predominantly size-change mutations. PMID- 10529739 TI - Hepatic gene mutations induced in Big Blue rats by both the potent rat liver azo carcinogen 6BT and its reported noncarcinogenic analogue 5BT. AB - The potent rat liver carcinogen 6-p-dimethylaminophenylazobenzthiazole (6BT) and its reported noncarcinogenic analogue 5-p-dimethylaminophenylazobenzthiazole (5BT; evaluated for carcinogenicity under the similar limited bioassay conditions used for 6BT) have been studied in order to seek an explanation for their different carcinogenic activities. Both compounds act as DNA-damaging agents to the rat liver, and both have now been shown to induce lacI (-) gene mutations in the liver of Big Blue(trade mark) transgenic rats. Both compounds were mutagenic following ten daily gavage doses or following administration in diet for 10 days. Neither chemical induced cell proliferation in the liver following repeat gavage administrations. In contrast, dietary administration of 6BT, and to a lesser extent of 5BT, induced hepatic cell proliferation. The carcinogen 6BT, but not the noncarcinogen 5BT, caused proliferation of oval stem cells in the livers by both routes of administration. It is possible that mutations induced in oval cells by 6BT are responsible for its potent carcinogenicity, and that the comparative absence of these cells in 5BT-treated livers may account for the carcinogenic inactivity of 5BT. Equally, the proliferation of the oval cells may reflect changes in liver homeostasis associated with the liver toxicity observed at the dose level of 6BT used (which was, nonetheless, the dose level used in the positive cancer bioassays). It is concluded that the new data presented cannot explain the differing carcinogenic activities of 5BT and 6BT, and that the reported noncarcinogen 5BT may also be carcinogenic when adequately assessed for this activity. PMID- 10529740 TI - Tissue-specific mutant frequencies and mutational spectra in cyclophosphamide treated lacI transgenic mice. AB - The induction and nature of mutations in the lacI transgene were evaluated in multiple tissues after exposure of adult male B6C3F1 lacI transgenic mice to cyclophosphamide (CP). Mice were given a single i.p. injection of 25 mg CP/kg, 100 mg CP/kg, or vehicle (PBS) and then necropsied 6 weeks after treatment to allow DNA extraction and lacI mutant recovery. Tissues evaluated included target tissues for tumorigenesis (lung, urinary bladder) and sites not susceptible to tumor formation in B6C3F1 mice (kidney, bone marrow, splenic T-lymphocytes). After exposure to the high dose of CP, a significant increase in the mutant frequency (Mf) was detected in the lungs and urinary bladders, compared to the respective tissues from vehicle-treated controls. In contrast, the Mfs in kidney, bone marrow, and splenic T cells from CP-treated mice were not significantly different from controls. The spectra of mutations in lacI from lung and urinary bladder were significantly changed after high-dose CP treatment, with a significant increase in the frequency of A. T --> T. A transversions found in both tissues and a significantly elevated frequency of deletions in the lungs. Conversely, in vehicle-treated mice, the two predominant classes of lacI mutations recovered in lung and urinary bladder were G. C --> A. T transitions at CpG sites and G. C --> T. A transversions. These CP exposures were also genotoxic as measured by the significant induction of micronuclei in peripheral blood 48 hr after exposure. These data indicate that under these study conditions, CP-induced mutations are detectable in the lacI transgene in the target tissues, but not in nontarget tissues for CP-induced cancer. With the lacI assay it is possible to study mutagenicity in a variety of critical tissues to provide mechanistic information related to genotoxicity and carcinogenicity in vivo. PMID- 10529741 TI - Detection of cyclophosphamide-induced mutations at the Hprt but not the lacI locus in splenic lymphocytes of exposed mice. AB - The relative sensitivities and specificities of the endogenous Hprt gene and the lacI transgene as mutational targets were evaluated in splenic lymphocytes from male standard B6C3F1 mice (only Hprt assayed) and from lacI transgenic B6C3F1 mice treated at 6-7 weeks- of-age with the indirect-acting agent, cyclophosphamide (CP). To define the effects of the time elapsed since CP treatment on Hprt mutant frequencies (Mfs), nontransgenic mice were given single i.p. injections of 25 mg CP/kg or vehicle (PBS) alone and then necropsied 2, 4, 6, 8, or 10 weeks after treatment. Peak Mfs were found at 6 weeks postexposure, with mean Mf values ranging from 2.27 to 3.27 x 10(-5) using two different lots of CP in standard packaging (compared with mean control Mf values of 0.14 to 0.26 x 10(-5) in various experiments). To determine the dose response for Hprt Mfs, nontransgenic mice were given single doses of 0, 12.5, 25, 50, or 100 mg CP/kg and necropsied 4 weeks postexposure. These treatments produced a supralinear dose response curve for CP-induced Hprt Mfs. Based on these experiments, CP mutagenicities at Hprt and lacI were compared in transgenic mice treated with 0, 25, or 100 mg CP/kg (using another lot of CP in ISOPAC((R)) bottles; Sigma) and necropsied 6 weeks later. There was a significant increase in Hprt Mfs in treated transgenic mice (100 mg CP/kg: 0.75 +/- 0.09 x 10(-5); 25 mg CP/kg: 0.39 +/- 0.05 x 10(-5)) versus controls (0.10 +/- 0.01 x 10(-5)); however, the Mfs in lacI of lymphocytes from the same CP-treated animals were not significantly different from controls (100 mg CP/kg: 9.4 +/- 1.1 x 10(-5); 25 mg CP/kg: 6.7 +/- 0. 8 x 10(-5); control: 7.7 +/- 0.7 x 10(-5)). Hprt mutational spectra data in CP treated transgenic and nontransgenic mice were different from those of control mice, whereas the spectra of mutations in lacI of lymphocytes from Big Blue((R)) transgenic mice were not significantly changed after CP treatment. These data indicate that, under these treatment conditions, CP-induced mutations in splenic lymphocytes were detectable in the Hprt gene but not the lacI transgene of this nontarget tissue for CP-induced cancer. PMID- 10529742 TI - Kinetics of induction of DNA damage and lacZ gene mutations in stomach mucosa of mice treated with beta-propiolactone and N-methyl-N'-nitro-N-nitrosoguanidine, using single-cell gel electrophoresis and MutaMouse models. AB - beta-Propiolactone (BPL) and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) are two direct alkylating agents that induce multiple genetic lesions and tumors in the rodent stomach. We measured the kinetics of the induction of DNA damage by using the single-cell gel electrophoresis assay (SCGE) and the induction of gene mutations by using the MutaMouse model in the glandular stomach mucosa of mice exposed to a single oral administration of BPL or MNNG. The aims were to determine the optimal sampling time and to investigate the cause-effect relationship between DNA damage and gene mutations. The induction of comets, evaluated in individual cells with the tail moment, was analyzed 1, 2, 4, 24, and 72 hr after a single oral administration of 25 mg/kg BPL or 20 mg/kg MNNG. The effects of both compounds were most intense at the earlier sampling times (1-2 hr), tailing off 4 hr after treatment and becoming undetectable at 72 hr. The lacZ mutant frequency (MF) was measured 3, 7, 14, 28, and 50 days after a single oral administration of 150 mg/kg BPL or 100 mg/kg MNNG, and 3 and 14 days after a single administration of 25 mg/kg BPL or 20 mg/kg MNNG. The MF was strongly enhanced at the highest doses and all sampling times, the most marked effects being observed 14 days (11.1-fold) and 28 days (19.0-fold) after BPL and MNNG administration, respectively. At the lowest doses, only a small increase in MF ( approximately 2.5- to 3.5-fold) was found at both sampling times. Primary DNA damage detected with SCGE shortly after treatment (1-2 hr) was rapidly (3 days) transformed into stable gene mutations that remained detectable for 50 days. These results illustrate the ability and complementarity of the SCGE and MutaMouse models to assess the genotoxicity of direct alkylating agents in the mouse gastric mucosa in vivo. PMID- 10529743 TI - Evidence for the lack of base-change and small-deletion mutation induction by trichloroethylene in lacZ transgenic mice. AB - Trichloroethylene (TCE) is a widely used industrial solvent employed mainly for degreasing and cold-cleaning metal parts. It is also used for dry cleaning, and in the production of a number of chemical products. It has been shown to induce liver and lung tumors in rodents, and have a variety of positive and negative results using in vitro and in vivo mutagenicity tests. In order to assist in the interpretation of the mechanism of carcinogenicity, TCE was tested for the ability to induce gene mutations and small deletions using the lacZ transgenic mouse model (MutaMouse). Male and female animals were exposed by inhalation to 0, 203, 1153, and 3141 ppm TCE, 6 h per day for 12 days. 14 and 60 days following the last exposure, animals were sacrificed and the mutation frequency in bone marrow, kidney, spleen, liver, lung, and testicular germ cells determined. The results of this study indicate that TCE did not induce base-change or small deletion mutations as detected in this assay in any of the tissues examined. Environ. Mol. Mutagen. 34: 190-194, 1999. Published 1999 Wiley-Liss, Inc. PMID- 10529744 TI - Spontaneous mutation frequency and pattern in Big Blue mice fed a vitamin E supplemented diet. AB - Endogenous oxidative DNA damage caused by normal cellular processes may play a vital role in carcinogenesis. To directly test the hypothesis that antioxidants will protect DNA from oxidative damage in vivo, Big Blue((R)) mice were fed either a control diet (66 IU vitamin E/kg diet) or a high-dose vitamin E diet containing 1000 IU vitamin E/kg diet of racemic d,l-alpha-tocopherol acetate from conception until 3 months of age. Using the standard Big Blue((R)) protocol, 15.5 million plaque forming units (pfu) were examined from five tissues (heart, liver, adipose tissue, thymus, and testis) of three control and three high-dose vitamin E supplemented male mice generating 433 mutants, which represented 373 independent mutations upon sequencing the lacI transgene. The alpha-tocopherol tissue concentration increased with high-dose vitamin E supplementation. In four of the tissues, individually or combined, mutation frequency changed little if any with vitamin E supplementation. In adipose tissue, which accumulated the highest levels of vitamin E, mutation frequency was significantly reduced with high-dose vitamin E supplementation (P = 0.047). Within the constraints of sample size, the pattern of mutation in adipose tissue was not altered significantly (P = 0.40). When data from all tissues were combined, a reduction in G:C --> T:A transversions was observed (P = 0.044). These results may have implications for cancer chemoprevention and provide insight into the efficacy of vitamin E supplementation in reducing spontaneous oxidative DNA damage in vivo. More dramatic alterations of mutation frequency and pattern may be observed with higher doses of vitamin E and substitution of the racemic supplement with d-alpha tocopherol acetate. PMID- 10529745 TI - The cII locus in the MutaMouse system. AB - Here, we report the first application and characterization of the cII locus as a mutational target for use with the Muta(trade mark)Mouse system for quantifying somatic mutations in vivo. This locus can be analyzed for mutations using positive selection and is identical in sequence to the cII in the Big Blue((R)) Mouse. The cII displays similar spontaneous (5.5 x 10(-5)) and induced mutation frequencies when compared to the lacZ gene in the small intestine of MutaMice treated with ENU (N-ethyl-N-nitrosourea). After acute treatment with 250 mg/kg ENU (ip) the mutant frequencies were 127 x 10(-5) at the cII and 147 x 10(-5) at the lacZ loci, reaching a maximal mutant frequency 10 days posttreatment and remaining constant thereafter. These data prove that this transgene is genetically neutral, conferring neither selective advantage nor disadvantage on the host cells. The cII dose response curve was linear (R(2) = 0.93) comparable to the lacZ after treatments with 0, 50, 150, or 250 mg/kg ENU. Use of the cII locus (0.3 kb) addresses the single most significant drawback associated with the MutaMouse system, namely the inability to obtain sequence spectra efficiently, due to the large size of the lacZ gene (3.0 kb). Moreover, a less obvious application, but nevertheless of considerable importance, is the easy identification of jackpot mutations, without sequencing. The cII, identical in both sequence and origin on the transgenic constructs used in producing the Big Blue and MutaMouse systems, provides the first transgenic locus common to the two widely used in vivo mutagenesis assays. PMID- 10529746 TI - Effects of extended chronic exposures on endogenous and transgenic loci: implications for low-dose extrapolations. AB - Although transgenic and endogenous loci generally respond alike to acute mutagenic exposures, those loci that have been tested respond differently to daily or continuous exposures. During chronic exposures, the transgenes accumulate mutations linearly, whereas the endogenous loci are less mutable initially but later accumulate mutations at an accelerating rate. The result is a reverse dose rate effect in which the same total dose is more mutagenic for the endogenous locus when spread over a longer time. This makes extrapolations to still lower chronic exposures uncertain. Here we report extension of a chronic exposure to N-ethyl-N-nitrosourea (ENU) in drinking water to longer times and to lower doses. The F(1) of MutaMouse males x SWR that were used permit detection of mutations at both lacZ and Dlb-1. Both of these mutations were found to be genetically neutral over this period. Extension of the exposure from 30 to 90 days at 94 microg/ml/d showed a continuation of the curves found previously for 10 to 30 days, namely, linear for mutations of the lacZ transgene and concave upward for the Dlb-1 endogenous gene. A simple model for these data is presented. Of the extended exposures, only the highest dose produced a significant increase in Dlb-1 mutant frequency, an increase consistent with the model. The time (at 2.8 microg/ml/d), concentration (after 480 days exposure), and dose (concentration x duration of exposure) response curves were not significantly different from linearity. The data for the transgene are not as convincing, due to the high spontaneous mutant frequency, obscuring the induced response. PMID- 10529747 TI - Quantitative correlation between radiation-induced mutagenesis in endogenous genes and transgenes of mouse spermatogonial stem cells. AB - In order to evaluate the pUR288-plasmid transgenic mouse model, utilizing the bacterial lacZ gene as the mutational target, radiation-induced mutagenesis was primarily analyzed in spermatogonial stem cells. A combined hydroxyurea (HU)-X ray treatment protocol was used, known to sensitize dramatically the induction of mutations in endogenous genes. In the testes of untreated animals, a mutant frequency of 6.7 +/- 4.4 x 10(-5) was found. In animals treated with HU or X ray alone, moderate elevations were seen (factors of about 4 and 2 over untreated animal values). In testes of mice having received the HU + X-ray combination treatment, a mutant frequency of 63.0 +/- 36.1 x 10(-5) was found. The results obtained showed a good quantitative correlation between endogenous genes and the transgene, indicating the suitability of pUR288 transgenic mice for also efficiently recording radiation-induced genetic damage. Radiosensitization, seen in spermatogonial stem cells, was not observed in other studied organs such as spleen, brain, or lung. PMID- 10529748 TI - Effects of O (6)-alkylguanine-DNA alkyltransferase deficiency in Escherichia coli as the host for the detection of mutations in lacI transgenic mice. AB - Transgenic mice are widely used to detect gene mutations in vivo induced by a variety of chemicals. It is known, however, that no mutagenicity of methyl methanesulfonate (MMS) is detected in epididymal sperm in various transgenic mice assays, although MMS induces the dominant lethal and specific locus mutations in male mice. To investigate the issue of whether unrepaired lesions in DNA of mature sperm can be transformed into mutations during replication of the lambda phage in Escherichia coli cells, we developed an E. coli strain YG5152, which is a derivative of strain SCS-8 but is deficient in the genes encoding O (6) alkylguanine-DNA alkyltransferases. When lambda LIZalpha phages were treated with MMS or N-ethyl-N-nitrosourea (ENU) in vitro and infected to the E. coli strains, the mutant frequencies of lacI were markedly higher in strain YG5152 than in strain SCS-8. When Big Blue(trade mark) mice were treated with MMS (160 mg/kg) or ENU (125 or 250 mg/kg) and the phages rescued from mature sperm were infected to the strains, the mutation frequency (MF) of phages from ENU-treated mice at a dose of 250 mg/kg in strain YG5152 was about two times higher than that in strain SCS-8. However, no increase in the MF was observed in the MMS-treated mice even in strain YG5152. These results suggest that, although strain YG5152 efficiently detects ex vivo mutations caused by mutagenic alkyl adducts formed by MMS in lambda phage DNA, no detectable levels of mutagenic methyl adducts are present in mature sperm of MMS-treated mice. Possible reasons for this lack of mutagenicity of MMS in mature sperm using transgenic mice assays are discussed. PMID- 10529749 TI - Review neuroimaging in amyotrophic lateral sclerosis. AB - Amyotrophic lateral sclerosis (ALS) is a chronic degenerative disorder of unknown etiology affecting the motor system. Conventional and non-conventional neuroimaging techniques can provide essential help both to increase the confidence in ALS diagnosis and to assess the disease evolution. Signal abnormalities at the level of the motor cortex and the corticospinal tract on conventional T2-weighted magnetic resonance (MR) images are a potentially useful marker of ALS pathology. However, the prognostic value of these conventional MR abnormalities is still hampered by their low pathological specificity. Non conventional MR techniques with a higher pathological specificity, such as MR spectroscopy, magnetization transfer imaging and diffusion-weighted imaging, seem to have some potential not only for ALS diagnosis, but also for monitoring disease evolution either naturally or when modified by experimental treatments. PMID- 10529750 TI - Manganese-containing superoxide dismutase signal sequence polymorphism associated with sporadic motor neuron disease. AB - An alanin-9valin (Ala-9Val) polymorphism in the mitochondrial targeting sequence of manganese-containing superoxide dismutase (Mn-SOD) has recently been described. We studied this polymorphism in 72 Swedish patients with sporadic motor neuron diseases (MND) and controls using an oligonucleotide ligation assay. There were significant differences in genotype between MND patients and controls (P = 0.025), and between male and female MND patients (P = 0.009). Individuals homozygous for the Ala allele had a higher risk for MND [odds ratio, 2.9; 95% confidence interval (CI), 1.3-6.6], which was increased when including only females in the analysis (odds ratio, 5.0; 95% CI, 1.8-14.0). In classical amyotrophic lateral sclerosis, the odds ratio was 3.8 (95% CI, 1.3-10.0), and 5. 5 (95% CI, 1.5-19.9) when including only females. The results suggest that mutations influencing the cellular allocation of Mn-SOD may be a risk factor in MND, especially in females, and that MND may be a disease of misdistribution of the superoxide dismutase enzymes. PMID- 10529752 TI - Executive functions and speed of mental processing in elderly patients with frontal or nonfrontal ischemic stroke. AB - Impairments in executive functions have been related to aging and frontal lobe lesions. Aging also causes slowing of mental processing. We examined whether ischemic stroke in the frontal brain area results in dysexecutive syndrome, or whether the frontal stroke causes increased slowing of mental processing. Neurological, radiological and neuropsychological examinations were carried out 3 months post-stroke on 250 ischemic stroke patients (55-85 years) and on 39 healthy control subjects. Of the patients, 62 had frontal and 188 had nonfrontal lesions. The neuropsychological examination comprised several cognitive domains, including tests considered to measure executive functions. The frontal group was slower than the nonfrontal group in tasks measuring speed of mental processing which were time-limited (Trail Making A, Stroop dots and fluency). They were also inferior in the Digit Span backwards task. There were no differences between the groups in other cognitive domains, nor in some tests which are considered to be measures of executive functions (e.g. WCST). Impairments in executive functions were evident in both the frontal and the nonfrontal groups compared with the controls, but no dysexecutive syndrome specifically related to frontal lesions was found. Frontal stroke related mainly to the slowing of mental processing. PMID- 10529751 TI - Heterogeneity of cognitive profiles in aging: successful aging, normal aging, and individuals at risk for cognitive decline. AB - Neuropsychological clinical decision-making is complicated by the fact that variability in test performance increases with advancing age. This research explores the presence of homogeneous subgroups in 120 neurologically healthy individuals, from 55 to 85 years of age. Subjects at risk for dementing diseases were diagnosed as Aging-Associated Cognitive Decline (AACD) and Mild Cognitive Impairment (MCI). Cluster analysis was applied on 11 neuropsychological variables assessing logical memory immediate recall and retention percentage, visual memory immediate recall and retention, conceptual thinking, naming, verbal fluency, constructional functions, motor speed, flexibility and finger tapping. Five clusters were extracted, one representing cognitively successfully aged, and two consisting of individuals with normal or average level of performance. One cluster was characterized by older subjects with difficulties in visual memory, visuoconstructional functions, and speed and attention, most of the younger subjects in the same cluster had a diagnosis of AACD or MCI. The fifth cluster represented individuals at risk for dementing diseases; most of them were diagnosed having AACD and more than half had a diagnosis of MCI. Age, activity and intellectual levels, and to a lesser degree education, were significantly related to the cluster solution. The present findings caution against treating samples of elderly individuals as homogeneous. PMID- 10529753 TI - Changes in short-term memory processes in patients with multiple sclerosis. AB - In this study we compared the performance of 39 multiple sclerosis (MS) patients with 28 age-, sex- and education-matched controls on both the Mini-Mental State Examination, a global cognitive assessment tool, and the Sternberg Short-Term memory scanning task, a standardized test of short-term memory (STM) processes. While the STM span of our MS patients did not differ from that of our controls, STM scanning time of the MS group was reliably slower than that of the controls and a significant correlation was observed between STM scanning time and duration but not severity of illness. Our results suggest that processing stages other than the manipulation of data within the STM buffer are also affected by MS. PMID- 10529754 TI - Plasma and cerebrospinal fluid human immunodeficiency virus type-1 (HIV-1) RNA levels in HIV-related cognitive impairment. AB - Cerebrospinal fluid (CSF) and plasma HIV-1 RNA levels were prospectively measured by the Roche Amplicor Monitor polymerase chain reaction assay in 30 HIV-1 infected patients without central nervous system opportunistic infections. All participants completed a global neuropsychological battery consisting of Mattis Dementia Rating Scale. Additional tests were used to better characterize the type of cognitive changes with a specific reference to frontal lobe function. The neuropsychological evaluation confirmed the subcortical pattern of cognitive dysfunction. CSF and plasma HIV-1 RNA levels were significantly correlated. No correlation was detected with either blood or CSF RNA levels and the global cognitive status, but when stratified in three cognitive subgroups, higher CSF HIV-1 RNA levels were observed in the more cognitively impaired subjects. Our results provide further evidence that plasma and CSF HIV-1 RNA level cannot be used as a reliable diagnostic marker for HIV-1 associated cognitive disorders. Only longitudinal studies will determine whether a high CSF HIV-1 level could be a risk factor for HIV-1 dementia. PMID- 10529755 TI - Early differentiation of cardioembolic from atherothrombotic cerebral infarction: a multivariate analysis. AB - The aim of this study was to determine factors predictive of cerebral infarction subtype from clinical data collected within 48 h of neurologic deficit. All cardioembolic (n = 231) and atherothrombotic infarctions (n = 369) included in prospective stroke registry of the Sagrat Cor-Alianza Hospital of Barcelona were analysed. Demographic characteristics, anamnestic findings, cerebrovascular risk factors and clinical data of patients with embolic stroke and patients with thrombotic infarction were compared. Predictors of stroke subtype were assessed by means of a logistic regression model based on 16 clinical variables. After multivariate analysis, atrial dysrhythmia and sudden onset to maximal deficit were significant predictors of embolic stroke, whereas hypertension, chronic obstructive pulmonary disease, diabetes, hypercholesterolemia and/or hypertriglyceridemia and age were independent predictive factors of atherothrombotic stroke. Setting a cut-off point of 0.50 for predicting mechanism of stroke on admission resulted in a sensitivity of 76%, specificity of 87% and total correct classification of 83%. Clinical features alone that are observed at stroke onset can help to distinguish cardioembolic from atherothrombotic infarctions. PMID- 10529756 TI - Supression of cardiac parasympathetic functions in patients with right hemispheric stroke. AB - OBJECTIVES: The aim of this study was to investigate the relationship between sympathetic and cardiac parasympathetic function and the side of the lesion in stroke patients. METHODS: Thirty-two patients with stroke and 29 healthy age matched control subjects were studied. Sympathetic skin responses (SSR) and RR interval variations (RRIV) during rest and deep breathing were recorded for the assessment of sympathetic and vagal parasympathetic function, respectively. RESULTS: The mean SSR amplitude values in patients compared with controls were 337 +/- 244 versus 1897 +/- 848 (P < 0.0001) for right hemispheric lesions and 466 +/- 398 versus 1873 +/- 843 (P < 0.0001) for left hemispheric lesions. The mean SSR latencies in patients compared with controls were 1526 +/- 163 versus 1395 +/- 109 (P < 0.05) for right hemispheric lesions and 1490 +/- 125 versus 1423 +/- 112 (P < 0.05) for left hemispheric lesions. RRIV (during deep breathing)/RRIV (at rest) ratios in patients compared with controls were 1.20 +/- 0.25 versus 1.84 +/- 0. 52 (P < 0.0001), and 1.55 +/- 0.88 versus 1.84 +/- 0.52 (P < 0.05) for right and left hemispheric lesions, respectively. CONCLUSION: Supression of vagal parasympathetic activity was more apparent in stroke patients with right hemispheric lesions in our series. Therefore, the right hemisphere seems to have a greater effect upon parasympathetic activity. PMID- 10529757 TI - LDL receptor mutations and ApoB mutations are not risk factors for ischemic cerebrovascular disease of the young, but lipids and lipoproteins are. AB - BACKGROUND: The genetic background for ischemic cerebrovascular disease of the young and the role of lipids and lipoproteins as risk factors are not clear. METHODS: We determined five LDL receptor mutations (Trp23Stop, Trp66Gly, Trp556Ser, 313+1G --> A, 1846-1G --> A) and three apolipoprotein B mutations (Arg3500Gln, Arg3500Trp, Arg3531Cys), and other risk factors for ischemic cerebrovascular disease in 80 patients (36 women, 44 men) with onset of disease before the age of 50 years compared with 3366 individuals from a general population sample within the same age range. RESULTS: None of the patients were carriers of mutations in the LDL receptor (Trp23Stop, Trp66Gly, Trp556Ser, 313+1G --> A, 1846 - 1G --> A) or the apolipoprotein B gene (Arg3500Gln, Arg3500Trp, Arg3531Cys) associated with hypercholesterolemia. However, on univariate analysis as well as on logistic regression analysis allowing for age and gender, plasma cholesterol (OR 1.4; P < 0.0005), HDL-cholesterol (OR 0.4; P < 0.005), diabetes (OR 5.8; P < 0.0001), and hypertension (OR 3.9; P < 0.001) were significant predictors of ischemic cerebrovascular disease. CONCLUSIONS: The five most common LDL receptor mutations in Danish patients with familial hypercholesterolemia and three mutations in the apolipoprotein B gene did not predispose to ischemic cerebrovascular disease of the young. However, cholesterol and HDL-cholesterol are important risk factors for ischemic cerebrovascular disease of the young in the present study. The elevation in cholesterol could in some patients be due to rare LDL receptor mutations not tested for, and could in other patients be multifactorial in origin. PMID- 10529758 TI - The diversity of seizures related to alcohol use. A study of consecutive patients. AB - The aim of this study was to investigate the influence of hazardous alcohol drinking on the occurrence of epileptic seizures, the semiology of such seizures, and the extent of the problem. A consecutive sample of 142 acute seizure patients (78 male and 64 female, mean age 46 (16-79) years) was studied. Control groups were 185 consecutive sciatica patients and 254 healthy individuals. Subjects with a hazardous alcohol drinking level were identified by a score >8 in the Alcohol Use Disorders Identification Test (AUDIT). Seizures in AUDIT-positive individuals occurring within 72 h of the last drink were considered to be related to alcohol withdrawal. Generalized or partial onset seizures were classified on the basis of history, electroencephalographic (EEG) and neuroradiological findings. Thirty five percent of seizure patients were AUDIT-positive, whereas conversely 27% were abstainers. Two-thirds of AUDIT-positive seizure patients met the criteria for withdrawal seizures. Indications of partial onset seizures were found in 25 (51%) of AUDIT-positive patients, all secondarily generalized seizures. Sixty percent of generalized onset seizure patients were AUDIT-positive. In conclusion, seizure patients included significantly more AUDIT-positive subjects, as well as abstainers, than healthy Norwegian controls and consecutive sciatica patients from our hospital. Partial onset seizures are more frequent among hazardous drinkers than hitherto recognized. A generalized onset seizure in adults warrants a high suspicion of alcohol as a provoking factor. Routine screening of acute seizure admissions with the Alcohol Use Disorders Identification Test is recommended. PMID- 10529760 TI - Colour vision abnormalities and movement time in Parkinson's disease. AB - Visual system dysfunction has been reported in Parkinson's disease (PD). The objective of the present study was to evaluate a putative association of distorted colour vision and delayed initiation and execution of movement in PD. We performed the Farnsworth-Munsell 100-hue test and estimated the total error score in 30 previously untreated parkinsonian patients and 30 age- and sex matched controls. We then determined slowness in motor readiness and motor programming in the parkinsonian subjects on the same day. Subjects were asked to press a start button and release it after the randomized appearance of a visual stimulus and to move their right index finger to a reaction button as quickly as possible. Reaction time was considered as elapsed time between onset of the stimulus light and release of the start button, movement time was the time period between release of the start button and the pressing of the reaction button. Significant differences appeared between parkinsonian patients' and controls' reaction times (P = 0.007), movement time (P = 0.001) and total error score (P = 2.23E-08). A significant relation (Spearman R = 0.473, P = 0.008) was found between movement time and total error score, but not between reaction time and total error score (Spearman R = 0.259, P = 0.166). We conclude, that visual dysfunction and execution of movement are more influenced by altered dopaminergic neurotransmission in PD in comparison to the initiation of movement. PMID- 10529759 TI - Serum and hair trace element levels in patients with epilepsy and healthy subjects: does the antiepileptic therapy affect the element concentrations of hair? AB - In this study, hair magnesium (Mg), zinc (Zn), copper (Cu), and manganese (Mn) levels, and serum Zn and Mg levels were measured by atomic absorption spectrophotometer in patients with epilepsy (n = 33) and healthy subjects (n = 21), and results obtained were statistically compared. The mean hair Cu, Mg, and Zn levels of epileptic patients were significantly lower than the levels of control subjects. There was no significant difference between epileptic patients and control subjects in respect to the mean Mn levels. Mean serum Mg levels in epileptic patients showed significant difference, but serum Zn levels were similar among both groups. When the effects of anticonvulsant therapy on Cu, Zn, Mn, and Mg in the hair, and Mg and Zn in the serum were analyzed in epileptics, there was no significant difference between the patients with or without therapy. Likewise, the mean trace element levels in epileptics showed no significant difference according to the type of antiepileptic drug and seizure, and gender. We suggest that the changed element status (Zn, Mg, and Cu) in hair may play an indicator role in the diagnosis of epileptic patients. PMID- 10529761 TI - Bilateral thalamic pain secondary to bilateral thalamic infarcts relieved by a further unilateral ischaemic episode. AB - This study reports a patient with severe, debilitating bilateral thalamic pain caused by bilateral thalamic infarcts. The authors consider it to be a unique case as a further clinically unilateral lesion led to pain relief bilaterally. PMID- 10529762 TI - Bilateral symmetrical enhancing brainstem lesions: an unusual presentation of primary CNS lymphoma. AB - We report a patient with a progressive brainstem syndrome, who on magnetic resonance imaging had large bilateral, symmetrical, contrast-enhancing, infratentorial space-occupying lesions. Biopsy of one of the lesions revealed this unusual appearance to be due to a primary central nervous system (CNS) lymphoma of B-cell type. Symmetry of lesions may be a clue to the diagnosis, perhaps reflecting the mechanism by which CNS lymphomas spread. PMID- 10529763 TI - Single cerebral Toxoplasma abscess: first manifestation of HIV infection. PMID- 10529764 TI - Industrial-academic collaboration: a bridge too far? PMID- 10529766 TI - Enhancing information sharing. PMID- 10529765 TI - Nicotine: not just for cigarettes anymore. PMID- 10529767 TI - Growth hormone secretagogues: recent advances and applications. AB - The discovery of a new class of compounds that stimulate the release of growth hormone (GH) in a manner distinctly different from growth hormone-releasing hormone (GHRH) is advancing the understanding of the mechanisms that control GH secretion. These compounds, the GH secretagogues, act at both pituitary and hypothalamic levels, and might even elicit effects in the CNS and peripheral systems. A receptor with high affinity for the GH secretagogues has been identified and several observations suggest the presence of additional receptors. The existence of these specific endogenous receptors could indicate that the mechanism of GH release is not yet fully understood. Several potential indications have been explored clinically and, as some of these compounds are orally active, they could offer attractive alternatives to recombinant human growth hormone (hGH) in treating GH disorders such as growth hormone deficiency (GHD), age-related conditions, obesity and catabolic conditions. PMID- 10529768 TI - Photodynamic therapeutics: basic principles and clinical applications. AB - Photodynamic therapy (PDT) is a promising new treatment for cancer that has been recently accepted in the clinic. PDT involves the localization of a light sensitive drug (photosensitizer) in the target tissue prior to illumination using an appropriate wavelength. Cytotoxic agents generated upon illumination trigger a cascade of biochemical responses that inactivate cancer cells either directly or through the induction of vascular stasis. These treatments are better tolerated as they destroy diseased tissue while leaving normal tissue intact. The haematoporphyrin derivative, Photofrin(R), has been approved in a number of European and Asian countries, as well as in North America. To enhance the potential of PDT and explore its application for other conditions, second generation photosensitizers are being rigorously investigated. PMID- 10529769 TI - Hepatitis C virus: an overview of current approaches and progress. AB - Hepatitis C virus (HCV) was unambiguously identified in the year 1989 as the agent responsible for most cases of non-A, non-B hepatitis, a chronic disease that often leads to cirrhosis and hepatocellular carcinoma. Having developed the means to detect the virus in the general population, it is now apparent that HCV infection is widespread and is likely to remain a health threat unless effective treatments are developed. The inability to propagate the virus in tissue culture and the scarcity of convenient animal models have proved to be major obstacles in drug discovery. Despite these limitations, several opportunities exist for targeted drug development based on the viral enzymes that have been characterized so far. These targets and inhibitors reported to be active against them are discussed in the following review. PMID- 10529771 TI - Combinatorial chemistry. PMID- 10529770 TI - Monitor: molecules and profiles. AB - Monitor provides an insight into the latest developments in drug discovery through brief synopses of recent presentations and publications together with expert commentaries on the latest technologies. There are two sections: Molecules summarizes the chemistry and the pharmacological significance and biological relevance of new molecules reported in the literature and on the conference scene; Profiles offers commentary on promising lines of research, emerging molecular targets, novel technology, advances in synthetic and separation techniques and legislative issues. PMID- 10529772 TI - Is there a balance between microglia and astrocytes in regulating Th1/Th2-cell responses and neuropathologies? PMID- 10529773 TI - Immunology of ocular tumours. PMID- 10529774 TI - Can eradication of helminthic infections change the face of AIDS and tuberculosis? PMID- 10529775 TI - What is the real role of CD40 in cancer immunotherapy? PMID- 10529776 TI - Structure and mucosal adjuvanticity of cholera and Escherichia coli heat-labile enterotoxins. PMID- 10529777 TI - Eotaxin: from an eosinophilic chemokine to a major regulator of allergic reactions. PMID- 10529778 TI - T cells orchestrate intestinal mucosal shape and integrity. PMID- 10529779 TI - Is rapeseed really an allergenic plant? Popular myths versus scientific realities. PMID- 10529780 TI - Conserved lipid and peptide presentation functions of nonclassical class I molecules. PMID- 10529781 TI - Mismatch repair and immunoglobulin gene hypermutation: did we learn something? PMID- 10529782 TI - The immunopathology of extrinsic (atopic) and intrinsic (non-atopic) asthma: more similarities than differences. PMID- 10529783 TI - Th2-deviation of fetus-specific T cells. PMID- 10529785 TI - Rare disorder could provide target for CF therapy. PMID- 10529784 TI - Duchenne muscular dystrophy improved by gentamicin. PMID- 10529786 TI - A role for interleukin 13 in Hodgkin's disease. PMID- 10529787 TI - 'Axogenic' and 'somagenic' neurodegenerative diseases: definitions and therapeutic implications. AB - Neurodegenerative diseases are characterized by a relentless loss of specific groups of neuronal subtypes. Many of these diseases share similar molecular mechanisms and extracellular mediators of neuronal loss. We now suggest that neurodegeneration originating in the neuronal cell bodies (e.g. in Alzheimer's disease, Parkinson's disease and amyotrophic lateral sclerosis) should be distinguished from that originating in the axons (e.g. in glaucoma, certain peripheral neuropathies and spinal stenosis). We propose that the former group of diseases be defined as 'somagenic' and the latter as 'axogenic'. Although axogenic disorders may share common symptoms and mediators of toxicity with somagenic disorders, they have distinct temporal, subcellular and signal transduction features. We further suggest that, by adopting this classification of disorders based on pathophysiological processes, we will come to recognize additional diseases (in particular, those defined as axogenic) as being neurodegenerative and therefore possibly amenable to neuroprotective therapy. PMID- 10529788 TI - The search for neural progenitor cells: prospects for the therapy of neurodegenerative disease. AB - The etiology of many neurodegenerative diseases has been identified in recent years. Treatment of central nervous system (CNS) disease could focus on one or more steps that lead to cell loss. In the past decade, cell therapy and/or ex vivo gene therapy have emerged as possible strategies for the treatment of neurodegenerative diseases. The ability to grow CNS-derived neural progenitor cells using growth factors has been extremely useful to study diverse phenomena including lineage choice, commitment and differentiation. By virtue of their biological properties and their presence in the adult CNS, neural progenitors represent good candidates for multiple cell-based therapies for neural diseases. Further identification of the molecules that direct the differentiation of adult neural progenitors may allow their activation in vivo to induce self-repair. This review addresses the nature, distribution and regulation of neural stem cells and the potential for applying these cells to both structural CNS repair and gene therapy. PMID- 10529789 TI - New perspectives on the molecular basis of hereditary bone tumours. AB - Bone development is a highly regulated process sensitive to a wide variety of hormones, inflammatory mediators and growth factors. One of the most common hereditary skeletal dysplasias, hereditary multiple exostoses (HME), is an autosomal dominant disorder characterized by skeletal malformations that manifest as bony, benign tumours near the end of long bones. HME is usually caused by defects in either one of two genes, EXT1 and EXT2, which encode enzymes that catalyse the biosynthesis of heparan sulphate, an important component of the extracellular matrix. Thus, HME-linked bone tumours, like many other skeletal dysplasias, probably result from disruptions in cell surface architecture. However, despite the recent success in unravelling functions for several members of the EXT gene family, significant challenges remain before this knowledge can be used to develop new approaches for the diagnosis and treatment of disease. PMID- 10529790 TI - Beyond BCG: the potential for a more effective TB vaccine. AB - The 'Bacille Calmette-Guerin' (BCG) vaccine has been used throughout most parts of the world for the majority of the century. It is safe to use and cheap to produce, but there have been increasing doubts about its effectiveness. These doubts could not come at a worse time, as tuberculosis (TB) rates continue to rise, compounded by the AIDS epidemic, and outbreaks of tuberculosis caused by multidrug-resistant strains are more common even in advanced countries. As a result, there is now a concerted research effort to produce new TB vaccine candidates. These include DNA vaccines, recombinant and auxotrophic vaccines and subunit vaccines, all of which hold promise. The real difficulty will probably arise at the clinical trial level, when a decision must be made either to replace BCG or to boost existing BCG-induced immunity using these new-generation vaccines. PMID- 10529792 TI - Theta activity, virtual navigation and the human hippocampus. PMID- 10529793 TI - Using intracranial recordings to study thetaResponse to J. O'Keefe and N. Burgess (1999). PMID- 10529791 TI - Molecular understanding of chronic pancreatitis: a perspective on the future. AB - Despite the recent development of medical imaging technology, chronic pancreatitis can only be diagnosed when the disease is fully established. This is due to the lack of specific and sensitive markers for this disease. The discovery of mutations in the cationic trypsinogen gene in patients with hereditary pancreatitis and a high incidence of mutations in the cystic fibrosis transmembrane conductance regulator gene in patients with chronic pancreatitis might be important clues to understanding the molecular mechanisms of this disease. The interaction between ethanol and ion channels might be the missing link between alcohol ingestion and chronic pancreatitis. PMID- 10529794 TI - Selective attention in reaching: when is an object not a distractor? PMID- 10529795 TI - Reply to Tresilian. PMID- 10529796 TI - Cognitive approaches to the development of short-term memory. AB - The capacity to retain information for brief periods of time increases dramatically during the childhood years. The increases in temporary storage of speech-based material that take place in the period spanning the pre-school years and adolescence reflect complex changes in many of the different component processes, including perceptual analysis, construction and maintenance of a memory trace, retention of order information, rehearsal, retrieval and redintegration. Another crucial capacity that undergoes a similar striking development is complex working memory, the ability to manipulate and store material simultaneously. Possible sources of age-related changes in working memory include increases in processing efficiency and attentional capacity, and task-switching. These two short-term memory systems might play significant but distinct roles in supporting the acquisition of knowledge and skills during childhood. Whereas phonological short-term memory is linked specifically with the learning of the phonological structures of new words, complex working memory appears to support processing and learning in a wide range of contexts, in both childhood and adulthood. PMID- 10529797 TI - The role of gesture in communication and thinking. AB - People move their hands as they talk - they gesture. Gesturing is a robust phenomenon, found across cultures, ages, and tasks. Gesture is even found in individuals blind from birth. But what purpose, if any, does gesture serve? In this review, I begin by examining gesture when it stands on its own, substituting for speech and clearly serving a communicative function. When called upon to carry the full burden of communication, gesture assumes a language-like form, with structure at word and sentence levels. However, when produced along with speech, gesture assumes a different form - it becomes imagistic and analog. Despite its form, the gesture that accompanies speech also communicates. Trained coders can glean substantive information from gesture - information that is not always identical to that gleaned from speech. Gesture can thus serve as a research tool, shedding light on speakers' unspoken thoughts. The controversial question is whether gesture conveys information to listeners not trained to read them. Do spontaneous gestures communicate to ordinary listeners? Or might they be produced only for speakers themselves? I suggest these are not mutually exclusive functions - gesture serves as both a tool for communication for listeners, and a tool for thinking for speakers. PMID- 10529798 TI - Strategic development. AB - Strategic development is more diverse, multifaceted and eventful than previously realized. Individual children know and use multiple strategies for solving a given kind of problem; they choose adaptively among available alternatives; and they frequently discover new strategies that enhance their problem-solving abilities. A recently formulated computer simulation (SCADS) indicates how associative and metacognitive processes work together to produce adaptive choices among existing strategies and discovery of useful new approaches. PMID- 10529799 TI - Successful intelligence: finding a balance. AB - Human intelligence has long been on the borderline between a scientific and a quasi-scientific field within the scope of psychological science. This is partially because its study and measurement have been particularly susceptible to socio-political agendas, but also because empirical tests of theories of intelligence have too often ranged from inadequate to nonexistent. In this article it is argued that two extremes have prevailed in the study of intelligence. At one extreme are general-ability (g) theorists, who have collected large amounts of data to test the theory of general intelligence, but often using restricted ranges of participants, materials or situational contexts. They also show a tendency to limit their methods of data analysis (e.g. to exploratory factor analysis). At another extreme are theorists arguing for new, multiple intelligences, whose theories have been subjected to few or no empirical tests. I argue that a middle ground is needed that recognizes the multifarious nature of intelligence and of people's conceptions of it, but that also is subjected to rigorous empirical tests. PMID- 10529800 TI - The mahogany mouse mutation: further links between pigmentation, obesity and the immune system. AB - Completely different lines of experimentation have identified attractin, a protein that seems to have multiple roles in regulating physiological processes. It affects the balance between agonist and antagonist at receptors on melanocytes, modifies behaviour and basal metabolic rate, and mediates an interaction between activated T cells and macrophages. It may well be a target for development of drugs to treat obesity. PMID- 10529801 TI - Genomic imprinting in mammals: an interplay between chromatin and DNA methylation? AB - Most imprinted loci have key regulatory elements that are methylated on only one of the parental chromosomes. For several of these 'differentially methylated regions', recent studies establish that the unmethylated chromosome has a specialized chromatin organization that is characterized by nuclease hypersensitivity. The novel data raise the question of whether specific proteins and associated chromatin features regulate the allele-specificity of DNA methylation at these imprinting control elements. PMID- 10529802 TI - Copying out our ABCs: the role of gene redundancy in interpreting genetic hierarchies. AB - The complete sequence of the Arabidopsis genome is scheduled to be determined by the end of the year 2000. While this goal could prove to be something of a moving target (the estimated size of the genome has grown from 120 Mb to 130 Mb over the last year1), it is clear that the majority of genes required for higher plant growth, reproduction and development will have been described within this time frame. Some of the implications of this landmark achievement are already becoming clear, even though less than a half of the genome has been sequenced. PMID- 10529803 TI - Small introns tend to occur in GC-rich regions in some but not all vertebrates. PMID- 10529804 TI - A conserved RNA structure element involved in the regulation of bacterial riboflavin synthesis genes. PMID- 10529805 TI - Orthology: another terminology muddle. PMID- 10529807 TI - Singling out Drosophila tendon cells: a dialogue between two distinct cell types. AB - The precise match between somatic muscles and their epidermal attachment cells is achieved through a continuous dialogue between these two cell types. Whereas tendon cells direct myotube migration and final patterning, the muscles are essential for the maintenance of the fate of tendon cells. The Drosophila neuregulin-like ligand, Vein, and its receptor, the epidermal growth factor receptor (Egfr), are critical components in the inductive signaling process that takes place between muscles and tendon cells. Additional gene products that relay the Vein-Egfr effect in Drosophila are conserved in the vertebrate neuregulin mediated cascade. This review describes genetic and molecular aspects of the muscle-tendon inductive processes in Drosophila, and compares them with the relevant mechanisms in the vertebrate embryo. PMID- 10529808 TI - Guilt by association: non-coding RNAs, chromosome-specific proteins and dosage compensation in Drosophila. AB - Dosage compensation is a striking example of the interplay between gene-specific regulation and chromosomal architecture. This process has evolved to make X linked gene expression equivalent in males with one X chromosome and females with two. Examining species at the molecular level has shown that dosage compensation is mediated by sex-specific factors that decorate the X chromosomes to regulate chromatin structure and gene expression. In Drosophila, dosage compensation is achieved, at least in part, through site-specific histone H4 acetylation, which is modulated by a male- and X-specific protein complex. The discovery of non coding RNAs that 'paint' dosage-compensated X chromosomes in mammals and in Drosophila suggests that RNAs play an intriguing, unexpected role in the regulation of chromatin structure and gene expression. PMID- 10529809 TI - Ci: a complex transducer of the hedgehog signal. AB - Recent progress has unveiled Cubitus interruptus (Ci) as a complex transcription factor whose diverse activities as an activator and repressor are regulated by its proteolysis, localization and concentration. The principal role of Ci is to elaborate the developmental program directed by the morphogen Hedgehog (Hh), and it uses its various activities to target the expression of key downstream genes to different spatial domains. Here, we highlight recent advances in the Ci story, and discuss remaining questions whose resolution promise to help explain how morphogens like Hh signal their distant targets. PMID- 10529810 TI - Genetic approaches to memory storage. AB - The ability to remember is perhaps the most significant and distinctive feature of our mental life. We are who we are largely because of what we have learned and what we remember. In turn, impairments in learning and memory can lead to disorders that range from the moderately inconvenient benign senescent forgetfulness associated with normal aging to the devastating memory losses associated with Alzheimer disease. Of the various, higher-cognitive abilities that human beings possess, such as reasoning and language, memory is the only one that can be studied effectively in simple experimental organisms that are accessible to genetic manipulation, such as snails, flies and mice. In these organisms, the effectiveness of genetic approaches in the study of memory has improved significantly in the past five years. Below we review these advances. PMID- 10529811 TI - What's new in the library? What's new in GenBank? let PubCrawler tell you. PMID- 10529812 TI - Curing amyloidosis: will it work in humans? PMID- 10529813 TI - VENT cells: a fresh breeze in a stuffy field? PMID- 10529814 TI - New Ca2+-releasing messengers: are they important in the nervous system? AB - In the nervous system, Ca2+ signalling is determined primarily by voltage-gated Ca2+-selective channels in the plasma membrane, but there is increasing evidence for involvement of intracellular Ca2+ stores in such signalling. It is generally assumed that neurotransmitter-elicited release of Ca2+ from internal stores is primarily mediated by Ins(1,4,5)P3, as originally discovered in pancreatic acinar cells. The more-recently discovered Ca2+-releasing messenger, cyclic ADP-ribose (cADPR), which activates ryanodine receptors, has so far only been implicated in a few cases, and the possible importance of another Ca2+-releasing molecule, nicotinic acid adenine dinucleotide phosphate (NAADP), has been ignored. Recent investigations of the action of the brain-gut peptide cholecystokinin on pancreatic acinar cells have indicated that NAADP and cADPR receptors are essential for Ca2+ release. Tools are available for testing the possible involvement of NAADP and cADPR in neurotransmitter-elicited intracellular Ca2+ release, and such studies could reveal complex mechanisms that control this release in the nervous system. PMID- 10529815 TI - MAPK and memory. PMID- 10529816 TI - Contralateral effects of peripheral nerve injury. PMID- 10529817 TI - Rod pathways: the importance of seeing nothing. AB - Anatomical and physiological studies of the mammalian retina have revealed two primary pathways available for the transmission of rod signals to the ganglion cells: one via ON rod bipolars, amacrine II cells, and ON and OFF cone bipolars, which is exquisitely designed for the transmission of single-photon absorption events; and a second via rod-cone gap junctions, and ON and OFF cone bipolars, which is designed for the transmission of multiple photon-absorption events at higher light levels. Psychophysical and electroretinographic (ERG) studies in normal observers and in two rare types of observer, who are devoid of either rod or cone function, support an analogous duality in the human visual system, the clearest signature of which is a loss of flicker visibility and ERG amplitude at frequencies near 15 Hz that results from destructive interference between sensitive 'slow' and insensitive 'fast' rod signals. The slow rod signal is most probably derived from the ON rod bipolar pathway and the fast signal from the rod cone gap junction and cone pathways. Evidence has emerged recently for a third, insensitive rod pathway between rods and OFF cone bipolars, but it has so far only been observed clearly in rodents. PMID- 10529818 TI - Chemokines in the CNS: plurifunctional mediators in diverse states. AB - The past decade has witnessed the remarkable ascendance of chemokines as pivotal regulatory molecules in cellular communication and trafficking. Evidence increasingly implicates chemokines and chemokine receptors as plurifunctional molecules that have a significant impact on the CNS. Initially, these molecules were found to be involved in the pathogenesis of many important neuroinflammatory diseases that range from multiple sclerosis and stroke to HIV encephalopathy. However, more-recent studies have fuelled the realization that, in addition to their role in pathological states, chemokines and their receptors have an important role in cellular communication in the developing and the normal adult CNS. For example, stromal-cell-derived factor 1, which is synthesized constitutively in the developing brain, has an obligate role in neurone migration during the formation of the granule-cell layer of the cerebellum. Many chemokines are capable of directly regulating signal-transduction pathways that are involved in a variety of cellular functions, which range from synaptic transmission to growth. Clearly, the potential use of chemokines and their receptors as targets for therapeutic intervention in CNS disease might now have to be considered in the context of the broader physiological functions of these molecules. PMID- 10529819 TI - Dynamics of neuronal processing in rat somatosensory cortex. AB - Recently, the study of sensory cortex has focused on the context-dependent evolution of receptive fields and cortical maps over millisecond to second time scales. This article reviews advances in our understanding of these processes in the rat primary somatosensory cortex (SI). Subthreshold input to individual rat SI neurons is extensive, spanning several vibrissae from the center of the receptive field, and arrives within 25 ms of vibrissa deflection. These large subthreshold receptive fields provide a broad substrate for rapid excitatory and inhibitory multi-vibrissa interactions. The 'whisking' behavior, an approximately 8 Hz ellipsoid movement of the vibrissae, introduces a context-dependent change in the pattern of vibrissa movement during tactile exploration. Stimulation of vibrissae over this frequency range modulates the pattern of activity in thalamic and cortical neurons, and, at the level of the cortical map, focuses the extent of the vibrissa representation relative to lower frequency stimulation (1 Hz). These findings suggest that one function of whisking is to reset cortical organization to improve tactile discrimination. Recent discoveries in primary visual cortex (VI) demonstrate parallel non-linearities in center-surround interactions in rat SI and VI, and provide a model for the rapid integration of multi-vibrissa input. The studies discussed in this article suggest that, despite its original conception as a uniquely segregated cortex, rat SI has a wide array of dynamic interactions, and that the study of this region will provide insight into the general mechanisms of cortical dynamics engaged by sensory systems. PMID- 10529820 TI - The dopamine hypothesis of reward: past and current status. AB - Mesolimbic dopaminergic neurons are thought to serve as a final common neural pathway for mediating reinforcement processes. However, several recent findings have challenged the view that mesolimbic dopamine has a crucial role in the maintenance of reinforcement processes, or the subjective rewarding actions of natural rewards and drugs of abuse. Instead, there is growing evidence that dopamine is involved in the formation of associations between salient contextual stimuli and internal rewarding or aversive events. This evidence suggests that dopaminergic-neuron activation aids the organism in learning to recognize stimuli associated with such events. Thus, mesolimbic dopaminergic neurons have an important function in the acquisition of behavior reinforced by natural reward and drug stimuli. Furthermore, long-lasting neuroadaptive changes in mesolimbic dopamine-mediated transmission that develop during chronic drug use might contribute to compulsive drug-seeking behavior and relapse. PMID- 10529821 TI - BAXing in the hypersensitive response. PMID- 10529822 TI - Improving oil functionality by tuning catalysis of thioesterase. PMID- 10529823 TI - Gibberellin signal transduction presents ellipsisthe SPY who O-GlcNAc'd me. AB - Although the molecular mechanisms by which plants respond to gibberellins are largely unknown, several components of the signal transduction pathway have been identified and a broad outline of how these components act in signal transduction is emerging. One component of the pathway, SPINDLY, is believed to be an O-GlcNAc transferase that post-translationally modifies cytosolic and nuclear proteins by the addition of O-linked N-acetylglucosamine. Although analysis of the properties of O-GlcNAc-modified proteins from animals has led to the hypothesis that this modification is regulatory, it has not been linked to specific signal transduction pathways. PMID- 10529824 TI - Current perspectives on mRNA stability in plants: multiple levels and mechanisms of control. AB - The control of mRNA stability plays a fundamental role in the regulation of gene expression in plants and other eukaryotes. This control can be influenced by the basal mRNA decay machinery, sequence-specific decay components, and regulatory factors that respond to various stimuli. Important progress has been made towards the identification of some of these elements over the past several years. This is true particularly with respect to cis-acting sequences that control mRNA stability, the identification of which has been the focus of much of the initial work in the field. Characterization of mRNA fragments associated with post transcriptional gene silencing and two plant transcripts that give rise to detectable decay intermediates have provided insight into the mRNA decay pathways. These, and other studies, are indicative of similarities, as well as of interesting differences between mRNA decay mechanisms in plants and yeast - the system that has been used for most of the pioneering work. Future studies in this area, particularly when enhanced by emerging genetic and genomic approaches, have tremendous potential to provide additional knowledge that is unique to plants or of broad significance. PMID- 10529825 TI - Lipid modifications of proteins - slipping in and out of membranes. AB - Protein lipid modification, once thought to act as a stable membrane anchor for soluble proteins, is now attracting more widespread attention for its emerging role in diverse signaling pathways and regulatory mechanisms. Most multicellular organisms have recruited specific types of lipids and a suite of unique enzymes to catalyze the modification of a select number of proteins, many of which are evolutionarily conserved in plants, animals and fungi. Each of the three known types of lipid modification - palmitoylation, myristylation and prenylation - allows cells to target proteins to the plasma membrane, as well as to other subcellular compartments. Among the lipid modifications, protein prenylation might also function as a relay between cytoplasmic isoprene biosynthesis and regulatory pathways that control cell cycle and growth. Molecular and genetic studies of an Arabidopsis mutant that lacks farnesyl transferase suggest that the enzyme has a role in abscisic acid signaling during seed germination and in the stomata. It is becoming clear that lipid modifications are not just fat for the protein, but part of a highly conserved intricate network that plays a role in coordinating complex cellular functions. PMID- 10529826 TI - Metabolism and functions of gamma-aminobutyric acid. AB - Gamma-aminobutyric acid (GABA), a four-carbon non-protein amino acid, is a significant component of the free amino acid pool in most prokaryotic and eukaryotic organisms. In plants, stress initiates a signal-transduction pathway, in which increased cytosolic Ca2+ activates Ca2+/calmodulin-dependent glutamate decarboxylase activity and GABA synthesis. Elevated H+ and substrate levels can also stimulate glutamate decarboxylase activity. GABA accumulation probably is mediated primarily by glutamate decarboxylase. However, more information is needed concerning the control of the catabolic mitochondrial enzymes (GABA transaminase and succinic semialdehyde dehydrogenase) and the intracellular and intercellular transport of GABA. Experimental evidence supports the involvement of GABA synthesis in pH regulation, nitrogen storage, plant development and defence, as well as a compatible osmolyte and an alternative pathway for glutamate utilization. There is a need to identify the genes of enzymes involved in GABA metabolism, and to generate mutants with which to elucidate the physiological function(s) of GABA in plants. PMID- 10529827 TI - Virus resistance and gene silencing: killing the messenger. AB - On occassion, virus-derived transgenes in plants can be poorly expressed and yet provide excellent virus resistance, and transgene constructs designed to supplement the expression of endogenous genes can have the effect of co suppressing themselves and the endogenous genes. These two phenomena appear to result from the same post-transcriptional silencing mechanism, which operates by targeted-RNA degradation. Recent research into RNA-mediated virus resistance and co-suppression has provided insights into the interactions between plant viruses and their hosts, and spawned several models to explain the phenomenon. PMID- 10529828 TI - The role of the ER and cytoskeleton in plant viral trafficking. AB - Plant viruses spread from cell to cell via plasmodesmata, which bridge the rigid cell wall and connect adjacent cells. The spread of infection is aided by interactions between the virus and the host components. These interactions have been intensively studied to understand the crosstalk between pathogen and host. The use of green fluorescent protein has shed new light on the close association between viral movement proteins and elements of the cytoskeleton and the endomembrane system. PMID- 10529830 TI - Immunoglobulin Fc receptors in clinical strains of staphylococcus aureus do not confer resistance to phagocytosis in an in vitro assay AB - Staphylococcus aureus binds Immunoglobulin G (IgG) on its external surface due to the presence of specific receptors for the Fc domain of this immunoglobulin. This mechanism represents a kind of camouflage against phagocytic cells. In order to confirm that possibility an in vitro evaluation of the phagocytic activity of leukocytes polymorpho-nuclear (PMN) against strains of Staphylococcus aureus was done, comparing 18 strains isolated from clinical samples and 16 from healthy individuals. The presence of Fc receptors was evaluated by haemagglutination (HA) with erythrocytes group A after incubation of the strains with IgG anti blood group A. Phagocytosis of S. aureus was carried out by mixing live bacteria with a suspension of human PMN and incubating at 37 degrees C for 1 h; survivors were counted as colony forming units by plating. The strains from clinical specimens showed higher HA than those from healthy individuals (p = 0.01); but the former were killed more efficiently than the latter (80-90% and 40%, respectively). It is may be possible that S. aureus showed different behavior in vivo, where could express other virulence factors to prevent the action of phagocytes. PMID- 10529831 TI - A ten-year survey of onychomycosis in the central region of the rio grande do sul, brazil AB - Onychomycosis is a common infection of the nail plate caused by fungal microrganisms, and represents approximately 50% of nails disorders and 30% of all superficial mycotic infections. We present a study of the frequency, epidemiology and clinical aspects of onychomycosis in the central region of Rio Grande do Sul during the period 1988-1997. PMID- 10529832 TI - Detection of a giardia lamblia coproantigen by using a commercially available immunoenzymatic assay, in belo horizonte, brazil AB - It is known that fecal examination to detect Giardia lamblia cysts or trophozoites produces a high percentage of false-negative results. A commercially available immunoenzymatic assay (ProSpecT Giardia Microplate Assay, Alexon, Inc., BIOBRAS) to detect G. lamblia specific coproantigen was evaluated for the first time in Brazil. A total of 90 specimens were tested. Each specimen was first tested as unpreserved stool, and then it was preserved in 10% Formalin to be tested 2 months later. The assay was able to identify all the 30 positive patients (sensitivity = 100.0%) by visual or spectrophotometric examination in the unpreserved specimens and was negative in 57 of the 60 patients without G. lamblia (specificity = 95.0%). The assay identified 27 of the 30 positive patients (sensitivity = 90.0%) and was negative in 59 of the 60 negatives (specificity = 98.3%) in the preserved stools according to both readings. A marked difference was observed in the optical densities in both groups, preserved and unpreserved stools, when the G. lamblia-positive specimens were compared to the negative or positive for other intestinal parasites than G. lamblia. The assay seems a good alternative for giardiasis diagnosis, especially when the fecal examination was repeatedly negative and the patient presents giardiasislike symptoms. PMID- 10529833 TI - HIV-(1/2) indeterminate western blot results: follow-up of asymptomatic blood donors in belo horizonte, minas gerais, brazil AB - The clinical and public health importance of indeterminate results in HIV-(1/2) testing is still difficult to evaluate in volunteer blood donors. At Fundacao Hemominas, HIV-(1/2) ELISA is used as the screening test and, if reactive, is followed by Western blot (WB). We have evaluated 84 blood donors who had repeatedly reactive ELISA tests for HIV-(1/2), but indeterminate WB results. Sixteen of the 84 donors (19.0%) had history of sexually transmitted diseases; 18/84 (21.4%) informed receiving or paying for sex; 3/84 (3.6%) had homosexual contact; 2/26 women (7.6%) had past history of multiple illegal abortions and 3/84 (3.6%) had been previously transfused. Four out of 62 donors (6.5%) had positive anti-nuclear factor (Hep2), with titles up to 1:640. Parasitological examination of the stool revealed eggs of S. mansoni in 4/62 (6.4%) donors and other parasites in 8/62 (12.9%). Five (5.9%) of the subjects presented overt seroconversion for HIV-(1/2), 43/84 (51.2%) had negative results on the last visit, while 36/84 (42.9%) remained WB indeterminate. Although some conditions could be found associated with the HIV-(1/2) indeterminate WB results and many donors had past of risky behavior, the significance of the majority of the results remains to be determined. PMID- 10529834 TI - Efficiency of indirect immunofluorescence assay as a confirmatory test for the diagnosis of human retrovirus infection (HIV-1 and HTLV-I/II) in different at risk populations AB - We compared the indirect immunofluorescence assay (IFA) with Western blot (Wb) as a confirmatory method to detect antibodies anti retrovirus (HIV-1 and HTLV-I/II). Positive and negative HIV-1 and HTLV-I/II serum samples from different risk populations were studied. Sensitivity, specificity, positive, negative predictive and kappa index values were assayed, to assess the IFA efficiency versus Wb. The following cell lines were used as a source of viral antigens: H9 ( HTLV-III b); MT-2 and MT-4 (persistently infected with HTLV-I) and MO-T (persistently infected with HTLV-II). Sensitivity and specificity rates for HIV-1 were 96.80% and 98.60% respectively, while predictive positive and negative values were 99.50% and 92.00% respectively. No differences were found in HIV IFA performance between the various populations studied. As for IFA HTLV system, the sensitivity and specificity values were 97.91% and 100% respectively with positive and negative predictive values of 100% and 97.92%. Moreover, the sensitivity of the IFA for HTLV-I/II proved to be higher when the samples were tested simultaneously against both antigens (HTLV-I-MT-2 and HTLV-II-MO-T). The overall IFA efficiency for HIV 1 and HTLV-I/II-MT-2 antibody detection probed to be very satisfactory with an excellent correlation with Wb (Kappa indexes 0. 93 and 0.98 respectively). These results confirmed that the IFA is a sensitive and specific alternative method for the confirmatory diagnosis of HIV-1 and HTLV-I/II infection in populations at different levels of risk to acquire the infection and suggest that IFA could be included in the serologic diagnostic algorithm. PMID- 10529835 TI - Characterization of rotavirus P genotypes circulating among paediatric inpatients in northern brazil AB - Between November 1992 and August 1993, twenty-eight rotavirus-positive stool samples obtained from paediatric inpatients in Belem, Brazil, aged less than four years, were tested by RT-PCR to determine the P genotype specificities. With the exception of 7 non-diarrhoeic children, all patients were either diarrhoeic at admission or developed diarrhoea while in hospital. Rotavirus strains with the gene 4 alleles corresponding to P1B[4] and P1A[8] types (both of which bearing G2 specificity) predominated, accounting for 78.6% of the strains. While only one P2A[6] type strain - with (mixed) G1 and 4 type specificities - was detected, the gene 4 allele could not be identified in 4 (14.3%) of the strains. Most (81%) of the specimens were obtained from children during their first 18 months of life. Rotavirus strains bearing single P1B[4] type-specificity were identified in both diarrhoeic (either nosocomial, 28.6% or community-acquired diarrhoea, 28.6%) and non-diarrhoeic (42.8%) children. P1A[8] gene 4 allele, on the other hand, was detected only among diarrhoeic children, at rates of 57.1% and 42.9% for nosocomial- and- community acquired diarrhoea, respectively. Mixed P1A[8],1B[4] type infection was identified in only one case of community-acquired diarrhoea. PMID- 10529836 TI - Seroprevalence of toxoplasmosis in fortaleza, Ceara, brazil AB - A serological survey of Toxoplasma gondii infection in population groups in Fortaleza, Brazil, was conducted, in order to identify seroprevalence rates according to age and risk factors associated with acquired infection. A Toxoplasma IgG-antibody enzyme immunoassay (Sanofi Pasteur Diagnostics, Marnes la Coquette, France) was employed to assess the immunity. Public day-care centers and schools were randomly selected, while three large antenatal clinics and maternity units were choosen by its importance. Population groups and results of 997 blood tests were: 227 children (mean age 3. 8 years), 22.8% seropositives; 584 students (mean 11.4 years), 58.4%, and pregnant and postpartum women (mean 24 years), 71.5% seropositives (p < 0.001). Of 256 participants reporting close contact with cats, 59.8% were seropositive, in contrast with 51% seropositives without contact (relative risk 1.17; 95% confidence interval 1.04-1.33; p = 0.01). Having three or more siblings at home was related to higher seroprevalence in children and students (relative risk 1.39; 95% confidence interval 1.21-1.60; p < 0.01). In conclusion, toxoplasmosis seroprevalence showed a rapid increase during the first ten years of life, in association with close contact with cats and larger households, probably related to inappropriate hygiene and child-care practices. PMID- 10529837 TI - Serological diagnosis of toxoplasmosis: usefulness of IgA detection and IgG avidity determination in a patient with a persistent IgM antibody response to toxoplasma gondii AB - We report the detection of specific IgA antibodies and the determination of IgG avidity in sequential serum samples from a patient exhibiting significant levels of Toxoplasma-specific IgM antibodies for seven years after the onset of the clinical symptoms of toxoplasmosis. IgM antibodies were detected by an indirect immunofluorescence test and by three commercial enzyme-linked immunosorbent assays (ELISA). Anti-T. gondii IgA was quantified by the alpha-capture ELISA technique using a commercial kit. As defined by the manufacturer of the IgA ELISA test used, most patients with acute toxoplasmosis have antibody levels > 40 arbitrary units per ml (AU/mL). At this cut-off level, the patient still had a positive ELISA result (45 AU/mL) in a serum sample taken one year after the beginning of clinical manifestations. The IgG avidity-ELISA test was performed with the Falcon assay screening test (F.A.S.T.(R)) - ELISA system. Avidity indices compatible with a recent Toxoplasma infection were found only in serum samples taken during the first 5 months after the onset of the clinical symptoms of toxoplasmosis. These results show that the interpretation of positive IgM results as indicative of recently acquired toxoplasmosis requires additional laboratory confirmation either by other tests or by the demonstration of a significant rise in the antibody titers in sequential serum samples. PMID- 10529838 TI - Production of TNF-alpha by primary cultures of human keratinocytes challenged with loxosceles gaucho venom AB - Primary cultures of human keratinocytes were challenged with increasing doses from 10 ng/mL to 2 &mgr;g/mL of Loxosceles gaucho venom, responsible for dermonecrotic lesion in humans. TNF-alpha was investigated by bioassay and ELISA in the supernatant of the cultures challenged with 100 ng/mL, 500 ng/mL, 1 and 2 &mgr;g/mL of venom. TNF-alpha was detected by bioassay in the supernatant of cultures challenged with 100 ng/mL, after 6 h. The cytokine was detected by ELISA in the supernatant of the cells challenged with doses of l &mgr;g/mL, after 6 and 12 h. The results point out the capacity of this venom to activate the keratinocytes in primary cultures to produce TNF-alpha. The production of cytokines could contribute to the local inflammatory process in patients bitten by Loxosceles sp. PMID- 10529839 TI - Chronic hepatitis C virus infections in brazilian patients: association with genotypes, clinical parameters and response to long term alpha interferon therapy AB - The present study assessed the clinical significance of hepatitis C virus (HCV) genotypes and their influence on response to long term recombinant-interferon alpha (r-IFN-alpha) therapy in Brazilian patients. One hundred and thirty samples from patients previously genotyped for the HCV and with histologically confirmed chronic hepatitis C (CH-C) were evaluated for clinical and epidemiological parameters (sex, age, time of HCV infection and transmission routes). No difference in disease activity, sex, age or mode and time of transmission were seen among patients infected with HCV types 1, 2 or 3. One hundred and thirteen of them were treated with 3 million units of r-IFN-alpha, 3 times a week for 12 months. Initial response (IR) was significantly better in patients with genotype 2 (100%) and 3 (46%) infections than in patients with genotype 1 (29%) (p < 0. 005). Among subtypes, difference in IR was observed between 1b and 2 (p < 0.005), and between 1b and 3a (p < 0.05). Sustained response (SR) was observed in 12% for (sub)type 1a, 13% for 1b, 19% for 3a, and 40% for type 2; significant differences were found between 1b and 2 (p < 0.001), and between 1b and 3a (p < 0.05). Moreover, presence of cirrhosis was significantly associated with non response and response with relapse (p < 0.05). In conclusion, non-1 HCV genotype and lack of histological diagnosis of cirrhosis were the only baseline features associated with sustained response to treatment. These data indicate that HCV genotyping may have prognostic relevance in the responsiveness to r-IFN-alpha therapy in Brazilian patients with chronic HCV infection, as seen in other reports worldwide. PMID- 10529840 TI - Safety and immunogenicity of hepatitis B vaccine ButaNG in adults AB - Recombinant yeast-derived hepatitis B vaccine manufactured by Instituto Butantan was administered in two groups of adult volunteers (I, II) following two different schedules of immunization. In the first trial (10 &mgr;g doses and 0, 1, 3 months vaccination schedule) 106 individuals completed the full immunization program. The results of seroconversion by age group varied from 70 to 100% and the GMT from 46.5 to 124.9 mIU mL-1. In the second trial with 68 individuals (for dosage comparison and 0, 1, 6 months vaccination schedule) indicated that the vaccine formulated in 20 &mgr;g was more effective than in 10 &mgr;g. The adverse reactions observed in the vaccinees were less frequent than the ones previously found since the introduction of similar vaccines. PMID- 10529841 TI - Atypical disseminated cutaneous histoplasmosis in an immunocompetent child, caused by an "aberrant" variant of histoplasma capsulatum var. capsulatum AB - A case of atypical disseminated cutaneous histoplasmosis in a five-year old, otherwise healthy child, native and resident in Sao Paulo metropolitan area is reported. Cutaneous lesions were clinically atypical. Histologic examination disclosed a granulomatous reaction but no fungal structures could be demonstrated by specific staining nor by immunohistochemical reaction. The fungus was isolated from biopsy material on two different occasions, confirming diagnosis of an unusual fungal infection. The fungus, originally thought to be a Sepedonium sp. due to the large sized, hyaline or brownish colored tuberculated macroconidia and to lack of dimorphism (yeast form at 37 degrees C) produce H and M antigens, visualized by the immunodiffusion with rabbit anti-Histoplasma capsulatum hyperimmune serum. Patient's serum sample was non reactive with H. capsulatum antigen by immunodiffusion, counterimmunoelectrophoresis and complement fixation tests, and immunoenzymatic assay failed to detect the specific circulating antigen. This serum was tested negative by double immunodiffusion when antigen obtained from one of the isolated samples was used. Both cultures were sent to Dr. Leo Kaufman, Ph.D. (Mycoses Immunodiagnostic Laboratory, CDC-Atlanta/USA), who identified them as H. capsulatum by the exoantigen and gen-probe tests. Both clinic and mycologic characteristics of the present case were atypical, suggesting the fungus isolated is an "aberrant variant" of H. capsulatum var. capsulatum, as described by SUTTON et al. in 1997. Treatment with itraconazole 100 mg/day led to cure within 90 days PMID- 10529842 TI - A case report of vascular catheter-associated bacteremia caused by mycobacterium tuberculosis in a non-immunosuppressed patient AB - Mycobacterium tuberculosis was isolated from a central venous catheter in a non immunosuppressed patient with systemic tuberculosis. This case report represents a very uncommon form of isolation of Mycobacterium tuberculosis. A total improvement was obtained after treatment. PMID- 10529843 TI - Cryptic infections in mice with the trypanosoma cruzi CL-14 clone PMID- 10529844 TI - Detection and identification of dengue-2 virus from santa cruz-bolivia by a single tube RT-PCR method PMID- 10529845 TI - Re: "Constitutive melanin in the cell wall of the etiologic agent of Lobo's disease" PMID- 10529846 TI - The authors reply: re: "Constitutive melanin in cell wall of etilogic agent of Lobo's disease" PMID- 10529847 TI - Role of the nitric oxide-cGMP pathway in the cardiovascular effects of anesthetic agents: a review. AB - Over the past 10 years, there has been an explosion of new information on the physiological and pathophysiological roles of nitric oxide (NO) within the cardiovascular system. With the increasing knowledge on the importance of NO in the regulation of cardiovascular function, possible involvement of the NO-cyclic guanosine-monmphosphate (cGMP) pathway in myocardial effects of anesthetics has attracted wide attention. This paper reviews the literature on the role of the nitric oxide-cGMP pathway in cardiovascular effects of volatile and intravenous anesthetics. PMID- 10529848 TI - Ventilatory adequacy and respiratory mechanics with laryngeal mask versus tracheal intubation during positive pressure ventilation. AB - The ventilatory adequacy and respiratory mechanics during positive pressure ventilation (PPV) via the laryngeal mask airway (LMA) are compared with the respiratory mechanics via the tracheal tube (TT). Thirty patients undergoing breast surgery were studied. After induction of anesthesia and muscle relaxation an LMA was inserted. Data were collected every 5 min for a 15 min period and included inspired (VTinsp) and expired (VTexp) tidal volumes, I:E ratio, peak airway pressure (Ppeak), plateau pressure (Pplat), total dynamic compliance (C), and the percentage of VT exhaled passively in the first second of expiration (V1.0%). Then the trachea was intubated and measurements were repeated as previously. Gas leak was calculated as the fraction (VTinsp- VTexp)/Vtinsp. VTinsp and VTexp did not differ significantly between the LMA or TT anesthesia at any time (P = 0.9318, P = 0.7071 for VTinsp and VTexp respectively), neither the Ppeak (P = 0.1382). Significant differences were found for Pplat (P = 0.000) and C (P = 0.0001). Individual comparisons showed a significant difference between the LMA Pplat at 5 min when compared with all the Pplat mean values recorded with the LMA or the TT (P < 0.05-0.01). The C mean value with the LMA at 5 min was significantly lower when compared with all the C mean values via the TT anesthesia (P < 0.05-0.01). Significant differences were found among the V1.0% measurements (P = 0.030) but not between individual comparisons. Leak was similar with the LMA or TT airway management. It is concluded that, in patients with normal airway pressure and compliance, PPV using the LMA is comparatively effective with the use of TT. PMID- 10529849 TI - Influence of methane on infrared gas analysis of volatile anesthetics. AB - Contemporary multigas analyzers determine anesthetic gas concentrations using (near) infrared analysis at either 3.3 or 8-9 microns. Methane also absorbs infrared light at 3.3 microns, but not at 8-9 microns. Consequently, erroneous anesthetic agent readings may result when methane is present in the circuit (e.g. during closed circuit anesthesia), potentially compromising patient safety. We have analyzed in laboratory conditions the influence of different known methane concentrations (100, 500 and 1000 ppm) on the gas-analysis readings provided by some clinical monitoring devices that use infrared absorption for the measurement of inhalation anesthetic concentration. At 3.3 microns wavelength the influence on the measurement of halothane was important, whereas the influence on that of enflurane and isoflurane was less pronounced. For desflurane and sevoflurane measurements, the influence of methane at 3.3 microns wavelength proved to be minimal. At higher wavelengths (8-9 microns) no influence of methane could be demonstrated. PMID- 10529850 TI - Transition time: a new parameter coinciding with fair intubating conditions. AB - With rocuronium optimal intubating conditions are earlier achieved than the adductor pollicis muscle onset time. Using the transition time we defined a better parameter for clinical relaxation. The onset of relaxation was determined in 20 patients. After a stable response was achieved with a 0.1 Hz single twitch stimulation 0.60 mg/kg rocuronium was injected. The three different stades during the onset of relaxation were determined. These are the lag time, transition time (transition between second and third phase) and onset time. Whether the transiton time corresponds with optimal intubating conditions was evaluated in 40 other patients. The median transition time was 67.4 (P25:52.5, P75:76.3) seconds with a corresponding relaxation of 76.2 (P25:81.4, P75:70.7)%. The intubating conditions were significantly better at a relaxation level corresponding with the transition time. We conclude that the transition time approximates the intubating time and corresponds with fair intubating conditions. This parameter can be preferred to define the moment with optimal intubating conditions. PMID- 10529851 TI - Prospective, randomized comparison of epidural and combined spinal epidural analgesia during labor. AB - The analgesic efficacy and incidence of maternal, fetal and neonatal side-effects of combined spinal epidural (CSE) and epidural (EPI) analgesia, using a mixture of bupivacaine 0.125%, epinephrine (1.25 micrograms.ml-1) and sufentanil (0.75 microgram.ml-1) for the relief of labor pain, were randomly and prospectively compared in 110 parturients. A 29 gauge Whitacre tip spinal needle was used to perforate the dura in CSE patients. Compared to EPI, CSE resulted in rapid (326 +/- 22 vs 766 +/- 79 sec, p < 0.05), excellent analgesia, using less bupivacaine (23.5 +/- 2.3 vs 33.9 +/- 2.9 mg, p < 0.05) and sufentanil (12.5 +/- 1.0 vs 16.5 +/- .7 micrograms, p < 0.05). A tendency to improved patient satisfaction in the CSE group was observed. The incidence of maternal or neonatal side effects was similar in both groups. No PDPH was observed. We conclude that CSE analgesia results in excellent pain relief during labor with immediate gratification as compared to epidural analgesia. PMID- 10529852 TI - [The atrioverter: an atrial defibrillator for the treatment of atrial fibrillation]. PMID- 10529853 TI - [Pulmonary vascular reactivity and the development of edema in the presence of prostaglandin inhibitors in the isolated canine lobe]. AB - Alveolar hypoxia is the most powerful pulmonary vasoconstrictor. In a previous work, we did not demonstrate significant changes in vascular reactivity and edema formation in an isolated canine lobe model during alveolar hypoxia. The purpose of this study is to define vascular pulmonary reactivity and edema formation after induction of pulmonary vasoconstriction using a prostaglandin inhibitor like tiaprofenic acid and alveolar hypoxia. Six isolated canine pulmonary lobules were instrumented and studied, all of them under two conditions (normoxia FIO2 21% and hypoxia FIO2 5%) four starting in normoxia condition and 2 starting in hypoxia condition. RESULTS: No significant changes in filtration rate were found, normoxia 0.42 +/- 0.41, hypoxia 0.37 +/- 0.51 ml/min/100 g pulmonary tissue P = NS. The arterial pressure in basal conditions was 25.1 +/- 6.21, and during hypoxia increased to 37 +/- 7.19 cm H2O (Delta 12.0 +/- 1.2 cm H2O). P < 0.001. CONCLUSION: Hypoxia vascular reactivity was significantly increased in tiaprofenic acid pretreated isolated canine lobes, no changes in pulmonary permeability was found nor increased rate in edema formation. PMID- 10529854 TI - [Surgical complications and mortality in octogenarian patients undergoing revascularization surgery]. AB - Our objective was to identify preoperative, operative and postoperative factors associated with complications and mortality in patients equal to or greater than 70 years of age with coronary artery disease treated with coronary bypass surgery. From january 1990 to june 1994 of those that underwent 37 coronary artery bypass surgery. 32 were men (86.5%) and five women (13.5%). History of cardiovascular disease, diabetes mellitus, systemic arterial hypertension, pulmonary disease, hypercholesterolemia, renal function, and severity of coronary artery disease were considered. Also analysed were aortic clamp and cardiopulmonary bypass time, number and type of grafts. Use of intraaortic balloon counterpulsation, inotropic drugs, ventilatory support, hemorrhage, infection, renal and liver failure, neurological, rhythm and conduction abnormalities and myocardial ischemia were also considered. Identified risk factors: diabetes mellitus, (p = 0.028), ejection fraction < 30% (p = 0.023), ventricular wall motion abnormalities (p < 0.05), aortic clamp > 60 minutes (p = 0.026), cardiopulmonary bypass < 120 minutes (p = 0.022), reverse saphenous vein grafts (p = 0.014), prolonged ventilatory support, inotropic drugs and intraaortic balloon counterpulsation. CONCLUSIONS: Surgery should be reserved for patients with at least three vessel or left main coronary artery disease or proximal lesion of the left anterior descending artery with severe ischemia, deteriorated myocardial function and angina with no response to medical treatment; age of the patient is not a contraindication. PMID- 10529855 TI - [Low-molecular-weight heparin in unstable angina pectoris]. AB - We studied the therapeutic effect of standard heparin (HS) compared with low molecular-weight (HBPM) in two homogeneous groups of 14 patients heparin selected at random, with clinical history and electrocardiographic signs of unstable angina pectoris. Patients received the conventional treatment with platelets' inhibitors, nitrates, adrenergic beta-blockers or calcium antagonists. Both heparins, separately, showed statistical therapeutic effect on the symptoms and signs of unstable angina pectoris. They decreased to zero the number and duration of symptomatic myocardial ischemic events observed by ambulatory electrocardiogram (EKG-Holter). The symptoms of the angina pectoris disappeared at the same elapsed time: in 51.9 +/- 20.2 min. for the HS, and in 48.14 +/- 20.7 min. for the HBPM. They decreased the frequency of the silent myocardial ischemia observed at the EKG-Holter: 9 events decreased to 4 with the HS, and 8 events decreased to 3 with the HBPM. They decreased the total elapsed time of the silent ischemia from 52 min. to 15 min., and the mean elapsed time of the silent ischemia decreased from 3.71 +/- 3.29 min. to 1.07 +/- 1.81 min. with the HS (P < 0.001). With HBPM it decreased the total elapsed time of the silent ischemia from 60 min to 10 min, and the mean elapsed time of the silent ischemia decreased from 4.28 +/- 4.49 min. to 0.71 +/- 1.43 min. (P < 0.02). Both heparins considerably decreased the frequency of the lethal arrhythmias. Although in this study we did not find statistical differences in the therapeutic action of either heparins, HBPM reduced rapidly angina symptoms and the events associated to angina pectoris, cardiac arrhythmias, specially lethal extrasystolia, conduction defects and atrial paroxysmal tachycardia. Compared to HS, HBPM is easily applied, does not produce side effects on coagulation or bleeding time. PMID- 10529857 TI - [Embolism in the right heart chambers: the diagnostic and therapeutic aspects]. AB - Deep venus thrombosis may result in pulmonary embolism. In rare instances, embolization has occurred, not directly to the pulmonary arterial tree, but to the right heart chambers. Although the value of echocardiography in the diagnosis is well recognised, their is no consensus for the appropriate treatment. We report herein six cases of floating right atrial thrombi, diagnosed by echocardiography, in patients with pulmonary embolism, or unexplained shock or syncope. Surgical embolectomy was carried out in 4 patients, and thrombolytic therapy in 2, without in-hospital mortality. The high mortality associated to this entity may be improved by rapid echocardiographic recognition and emergency treatment with thrombolysis or surgery. Our data suggest the possible use of thrombolysis as a first-choice therapy in selected patients. PMID- 10529856 TI - [The familial incidence of accessory atrioventricular pathways (the pre excitation syndrome)]. AB - Cases of familial preexcitation syndrome represent a specific subgroup of patients that may result from diverse mechanisms: failure in development and genetic predisposition are the main mechanism involved. We determined the prevalence of this syndrome in first degree relatives of patients with proved accessory pathways by electrophysiologic study and compared such prevalence with the general population (0.15%). In five out of 469 patients (1.06%) we found an accessory pathway in one or more member of their family. Only 6 out of 3752 had preexcitation (0.15%); this prevalence was similar to the general population (P = NS). The identification of family members with this syndrome may be incomplete because we only chose for the study symptomatic patients. We did not observed multiple pathways and in one case we found atrial septal defect. Our data demonstrated familial preexcitation in five families suggesting hereditary predisposition. PMID- 10529858 TI - [Complete congenital heart block. Its natural history and evolution]. AB - This study describes the natural history and evolution of 67 patients with congenital auriculoventricular heart block admitted in the Instituto Nacional de Cardiologia "Ignacio Chavez", Mexico, D.F. from 1944 to 1998. There were 35 (52%) females and 32 (47%) males, with mean follow up period of 93.7 +/- 104 months. Most of the patients were without structural cardiovascular disease (90%). The most frequent symptoms were dyspnea and syncope. Electrocardiograms showed a ventricular heart rate of 42.2 +/- 9 beats/minute. 85.7% of patients had a supra Hisian complete heart block. In 31% of patients a pacemaker was implanted because syncope. Overall mortality was 4.4% and malignant ventricular arrhythmias were the principal contributors. Risk factors for mortality identified in this study were junctional escape rhythm lower than 50 beats/minute, inappropriate chronotropic response during exercise, R-R interval prolongation at night, enlargement of cardiac chambers, depressed left ventricular ejection fraction and prolonged QT interval. In all of these conditions we recommend permanent pacemaker implantation. PMID- 10529860 TI - [Constrictive pericarditis and restrictive myocardiopathy]. AB - The purpose of this study was to assess the clinical and echocardiographic characteristics of constrictive pericarditis (CP) and restrictive cardiomyopathy (RC) and to compare them with the results obtained with cardiac catheterization. Clinical history, electrocardiogram and X-ray were taken in all patients, and transthoracic and transesophageal echocardiography were performed. Cardiac catheterization with transmyocardial biopsy was performed on only 5 patients. Wall thickness and left ventricular dimensions were normal in all patients with CP. Wall thickness was increased in those with RC. No patients demonstrated alterations in segmental wall movement. The pericardium was thickened and abnormally bright in the 3 patients with CP. In patients with CP the percentage of atrioventricular, semilunar, pulmonary and hepatic flow changes with respiration were more than 10%. In patients with RC this flow variation was less notable. However, the percentage of systolic and diastolic flow velocity increase of hepatic veins during expiration was greater than in CP. We can conclude that M mode, two dimensional and Doppler echocardiography is extremely useful noninvasive method to differentiate CP and RC with good correlation with cardiac catheterization. PMID- 10529859 TI - [A complete atrioventricular block during exertion]. AB - Exercise-induced atrioventricular (AV) block in patients with normal electrocardiogram at rest is uncommon. We report the clinical features of two patients with AV block during treadmill test. The first patient was a woman of 54 years of age with presyncope on exercise. She developed complete AV block during exercise testing without evidence of ischemic myocardial disease. Electrophysiologic study documented distal AV block. The second patient was a man 31 years old who developed angina and third-degree AV block with depression of ST segment during treadmill test. Myocardial perfusion study suggested ischemic heart disease. He received anti-ischemic drugs with improvement of symptoms. A control exercise testing demonstrated normal AV conduction and electrophysiologic study was normal. In the first case, exercise AV block was probably due to abnormal His Purkinje conduction system refractoriness to autonomic modulation, while in second case AV block was secondary to ischemic heart disease. Third degree AV block at exercise can be present in patients without conduction system abnormalities at rest. Exercise-induced infra-Hisian AV block must be treated with pacing until ischemic heart disease has been ruled out. PMID- 10529861 TI - [The pharmacological inhibition of the intracellular signals involved in cellular proliferation: a cardiovascular approach]. PMID- 10529862 TI - [Glycoprotein IIb/IIIa blockers. Their usefulness in current cardiovascular therapeutics]. PMID- 10529863 TI - The vagaries of personality development. PMID- 10529864 TI - Resin-based composite reinforced sealant. AB - PURPOSE: To describe and picture a technique for using a resin-bonded sealant reinforced by inclusion of resin-based composite, before polymerization. METHOD: Using an extracted mandibular molar, embedded in plaster and isolated with a rubber dam, a step-by-step depiction of the technique is shown. RESULTS: A picture of a six-year-old sealant in a first permanent molar, still in perfect condition, is shown. CONCLUSION: This technique offers a reliable method for treating pits and fissures. PMID- 10529865 TI - Fracture strength of adhesively restored pulpotomized primary molars. AB - PURPOSE: To evaluate the cuspal fracture resistance of primary teeth restored with different adhesive techniques. METHODS: Eighty primary molars were selected for pulpotomy preparations. The molars were matched according to molar type (upper or lower, 1st or 2nd), and divided into four groups: Group 1: Pulpotomy cavity restored with IRM, Dispersalloy amalgam (Control); Group 2: IRM, Hydroxyline, Amalgambond Plus and Dispersalloy; Group 3: IRM, Hydroxyline, Prime & Bond 2.1 and Dyract; Group 4: IRM, Hydroxyline, Single Bond and Z100 resin composite. In Groups 2,3 & 4, Hydroxyline was placed over the IRM to avoid direct contact of the adhesives and resin with the IRM. All specimens were thermocycled (1000 cycles, 5-55 degrees C), and mounted in acrylic 2 mm apical to the CEJ. The molars were then tested for cuspal fracture strength using an Instron machine with tapered steel balls made to match the occlusal area of the primary molars tested. Repeated measures ANOVA and paired t-test were used to test the statistical significance of the results. RESULTS: Mean fracture loads in Newtons + S.D.) were as follows: G1: 1087 + 284; G2: 1371 + 434; G3: 1336 + 320; G4: 1404 + 312. The three bonded procedures significantly (P < 0.001) increased the resistance of the primary molars to fracture compared to the control group. There was no significant difference between the three bonded procedures. CONCLUSIONS: Bonded restorations of pulpotomized primary molars may be an alternative restorative procedure to stainless steel crowns. PMID- 10529866 TI - Treating children with disabilities and their families. AB - Increased numbers of dental students and practitioners are necessary to provide services for children with disabilities. In many instances, the technical aspects of treatment are complicated by the interaction between the dentists, patient and the family. The stresses and strains in a family with a child with disabilities can and do impact on both the office visit and in the home setting. A review of the literature is provided in an effort to familiarize practitioners with these realities. PMID- 10529867 TI - Behavior and toothbrushing of young Israeli adolescents. AB - The purpose of the present study was to assess the toothbrushing behavior of Israeli adolescents attending a pediatric dental clinic, with regard to the maxillary anterior region. One hundred twenty-three adolescents (57 boys, 66 girls, mean age 12.45 +/- 1.76), participated in this study. The amount of plaque extending from the gingival line incisally was measured. Brushing characteristics and daily habits were recorded. In 87 patients, 4 mm or more of plaque were measured on the labial surfaces of the maxillary central incisors, while thirty six patients demonstrated less than 4 mm plaque. Patients who brushed the maxillary and mandibular incisors separately showed significantly less plaque than patients who brushed them simultaneously. Brushing the maxillary and the mandibular incisors separately was significantly more prevalent in the thirteen to-seventeen-year-olds, and among girls. Also, significantly more girls brushed twice per day, and for more than three seconds on each incisor. PMID- 10529868 TI - Evaluation of the experiences of fearful children at a Special Dental Care Centre. AB - A representative sample of parents was interviewed one to two years after their children had been treated at a Dutch Special Dental Care Centre (SBT). Information was sought about the children's current behavior during visits to family dentists. Within one to two years after the experiences at SBT more than 90 percent of the children visited a family dentist, 60 percent of them required restorative treatment and 80 percent of this treatment was performed, using local anesthesia. Retrospectively, the parents report a decrease of their children's dental anxiety, when leaving the SBT. This level of anxiety is unchanged after one to two years visiting a family dentist. For the children there was a significant relationship between dental anxiety as reported by the parents and not visiting a family dentist. Results suggest that referring dentists arrange the initial treatment at the Special Dental Care Centre and that the children in this group referred for dental anxiety belong to a caries-risk group. PMID- 10529869 TI - Practical tips managing children's behavior. PMID- 10529870 TI - Children with mental retardation grow older. AB - In an effort to encourage dental practitioners to provide needed services for children with mental retardation as they grow older, a review is provided of the ongoing difficulties faced by families and their children--including attitudes, residence, sibling and older parent dilemmas, cost of care and the realities of sexuality. PMID- 10529871 TI - The use of anorganic bovine bone to correct a residual osseous defect after tooth extrusion: report of case. AB - This report deals with the clinical treatment of an osseous defect resulting from the extrusion of an impacted canine in a twelve-year-old girl. An apical horizontal root resorption of the adjacent lateral incisor was a related condition to this impaction, influencing the choice of treatment. The bone defect, caused by a guided canine extrusion, was restored by an anorganic bovine bone mineral (Bio-Oss). After a one-year follow-up, the lateral incisor responded vital and showed no discoloration; no progress of the resorption was found. Furthermore, by improving the conduction of new bone formation from the defect walls, the osteoconductive potential of this anorganic bovine bone became obvious. This case report illustrates a new indication area for its use in pediatric dentistry. PMID- 10529873 TI - Bilateral fusion of the mandibular primary incisors: report of case. AB - Fusion is a common dental finding. However, bilateral mandibular fusion of the primary incisors is a rare event, occurring with a prevalence of less than 0.02 percent. When all four permanent successors are present, this event becomes rarer still. Once fusion has been diagnosed, careful monitoring is required, since problems with exfoliation can occur, along with caries formation in the groove of the incompletely fused teeth. PMID- 10529872 TI - A case of dental mutilation. AB - A case of ritual mutilation in a fourteen-year-old Ethiopian girl is described. When the girl was three years old she had frequent stomach problems. According to tribal tradition her illness was thought to arise from her mandibular primary canines and these teeth were removed by a medicine man. The extraction damaged the tooth germs of the succedaneous teeth and resulted in deformed permanent canines. This is the first report of a case of dental mutilation from Ethiopia. PMID- 10529874 TI - Vancomycin-resistant enterococci (VRE) colonization of high-risk patients in tertiary care Canadian hospitals. Canadian VRE Surveillance Group. AB - We isolated 1487 Enterococcus species from 1200 stool specimens collected from high-risk patients in 12 Canadian tertiary care hospitals between October 1995 and November 1996. The composition of the 1487 isolates was 601 vancomycin sensitive Enterococcus faecalis (40.4%), 667 vancomycin-sensitive Enterococcus faecium (44.9%), 18 vancomycin-resistant (nine isolates MIC 8-16 micrograms/mL; nine isolates MIC > or = 32 micrograms/mL) E. faecium (VREF) (1.2%), 95 vancomycin-sensitive Enterococcus gallinarum (6.4%), 29 vancomycin-resistant (all MICs 8-16 micrograms/mL) E. gallinarum (2.0%), and 77 vancomycin-sensitive Enterococcus casseliflavus (5.2%). Nine of the 18 VREF isolates collected possessed the vanA genotype and were from three patients at one hospital. Two other VREF isolates, of the vanB genotype, were from a single patient at a second hospital, and the remaining seven isolates, also all of the vanB genotype, were from five patients at a third hospital. All VREF were ampicillin resistant (MIC > or = 16 micrograms/mL), streptomycin resistant (MIC > 1000 micrograms/mL), and ciprofloxacin resistant (MIC > or = 4 micrograms/mL). Ten of the 18 VREF were also resistant to gentamicin (MIC > 500 micrograms/mL), while all 18 isolates had quinupristin/dalfopristin MICs < or = 0.5 microgram/mL. In conclusion, high-risk patients in tertiary care Canadian hospitals are rarely colonized (9/1200 patients, 0.75%) with VREF in their lower gastrointestinal tract. These findings correlate well with the lack of reported VREF infection in high-risk patients in Canadian hospitals. Quinupristin/dalfopristin demonstrated excellent in vitro activity against VREF and other non-faecalis species of Enterococcus, many of which also possessed high-level ampicillin, and/or high-level aminoglycoside, and/or ciprofloxacin resistance. PMID- 10529875 TI - Identification of hippurate-negative thermophilic campylobacters. AB - Twenty-eight thermophilic campylobacter isolates showing negative hippurate hydrolysis were further characterized. Using Campylobacter jejuni and coli specific primers for ceuE gene, five of the isolates with repeatedly negative results in rapid hippurate hydrolysis were positive in C. jejuni-specific polymerase chain reaction (PCR) and 13 isolates were shown to be C. coli. All except one isolate with positive results in C. jejuni PCR were negative in C. coli PCR including those with repeatedly negative hippurate hydrolysis results. One isolate was positive in both PCRs due to a mixed culture. In comparison with PCR, API Campy gave concordant results in only 20 of the 28 isolates tested. Hybridization with PCR probes for ceuE gene of known C. jejuni and coli strains confirmed PCR results in all 27 isolates tested. In contrast to hippurate hydrolysis, PCR seemed to be a more reliable method to identify C. coli. PMID- 10529877 TI - International surveillance of blood stream infections due to Candida species in the European SENTRY Program: species distribution and antifungal susceptibility including the investigational triazole and echinocandin agents. SENTRY Participant Group (Europe). AB - The SENTRY Antimicrobial Surveillance Program, an international study of blood stream infections (BSIs), detected 170 episodes of candidemia in 20 European medical centers (13 nations) between January and December, 1997. Twenty-three percent of the candidal BSI occurred in patients hospitalized in an intensive care unit, 21% in patients in an internal medicine service, 13% in patients in a surgical service, and 9% in patients in an oncology service. Overall, 53% of the BSI were attributable to Candida albicans followed in prevalence by C. parapsilosis (21%), C. glabrata (12%), C. tropicalis (6%), C. famata (2%), C. krusei (1%), and C. inconspicua (1%). As observed previously in Canada and Latin America, C. parapsilosis and not C. glabrata, was the most common non-albicans species causing yeast BSI in Europe. The proportion of these candidemias attributable to C. albicans varied widely from 0-100% among the 20 European centers. Among the different species of Candida, resistance to fluconazole (MIC, > or = 64 micrograms/mL) and itraconazole (MIC, > or = 1.0 microgram/mL) was observed with C. glabrata and C. krusei and was observed more rarely among other species (e.g., C. inconspicua). Isolates of C. albicans, C. parapsilosis, C. tropicalis, and C. guilliermondii were all highly susceptible to both fluconazole and itraconazole. Furthermore, the investigational triazoles (BMS-207147, Sch 56592, and voriconazole) and an echinocandin (MK-0991) all demonstrated potent in vitro activity (MIC90s, 0.5, 0.5, 1.0, and 2.0 micrograms/mL, respectively) against these isolates. Continued surveillance at an international level will be important to monitor trends in species distribution and antifungal susceptibility among invasive strains of Candida. PMID- 10529876 TI - Detection of methicillin-resistant Staphylococcus aureus (MRSA) from growth on mannitol salt oxacillin agar using PCR for nosocomial surveillance. AB - This study evaluated a polymerase chain reaction (PCR) method for detection of methicillin-resistant Staphylococcus aureus (MRSA) in specimens referred for nosocomial surveillance. PCR was used to detect the mecA and nuc gene targets using yellow growth on mannitol salt agar containing 6 mg/liter oxacillin (MSO-6) as a source of DNA (N = 645). The diagnostic values for PCR compared with culture methods were 97% specificity, 100% sensitivity, 96% positive predictive value, and 100% negative predictive value. Total cost for PCR per test is $3.62 compared to $4.77 for culture. However, the total cost per specimen is significantly lower due to only 20% of all surveillance specimens producing yellow colonies on MSO-6. The average turnaround time for the PCR method is 48 h compared with 82 h for culture. PCR amplification of mecA and nuc genes using yellow colonies on MSO-6 is a simple, fast, accurate and cost-effective method for routine use in clinical laboratories for detecting MRSA in surveillance specimens. PMID- 10529878 TI - Development of a PCR assay for diagnosis of Pneumocystis carinii pneumonia based on amplification of the multicopy major surface glycoprotein gene family. AB - We have evaluated a PCR technique using primers based on Pneumocystis carinii major surface glycoprotein (MSG) genes, a multicopy gene family, for utility in detection of P. carinii in BAL and oropharyngeal samples obtained from immunosuppressed patients. These primers were able to detect P. carinii DNA in as little as 16 fg of genomic DNA. PCR using MSG primers detected P. carinii DNA in 7 smear-positive BAL samples (100% sensitivity), and found no P. carinii DNA in 12 smear-negative BAL samples (100% specificity). Mitochondrial ribosomal RNA (mrRNA) primers, commonly used in PCR studies of PCP, detected P. carinii in six of seven positive samples (85.7% sensitivity) and none of 12 were negative samples (100% specificity). Diagnosis of PCP by amplification of 81 oropharyngeal samples using MSG primers had a 50% sensitivity (4/8) and 96% specificity (70/73). PCR with mrRNA primers was 37.5% sensitive (3/8) and 100% specific (73/73). All three false-positive MSG results showed a very low intensity on Southern hybridization. PCR using MSG gene primers should prove valuable in the diagnosis of PCP. PMID- 10529879 TI - Comparative evaluation of two methods for testing metronidazole susceptibility of Helicobacter pylori in routine practice. AB - Applicabilities of modified disk diffusion method (MDDM) and screening agar method (SAM) were evaluated in order to detect metronidazole resistance in vitro among 96 Helicobacter pylori strains. SAM was based on 8-mg/liter metronidazole as breakpoint, and MDDM was performed using disks containing 5 micrograms of the drug. Of 96 strains tested, 48 (50%) exhibited inhibitory zones of 26 mm or less by MDDM, and 44 (45.8%) were found resistant to 8 mg/liter metronidazole. Colonies growing within the inhibitory zones by MDDM were detected in six (6.2%) of all strains tested, and the subcultures revealed resistance by both methods. Overall agreement between the results of both tests was found in 90 (93.8%) of the 96 isolates. There is a possibility of great discrepancies between the two methods for testing H. pylori against metronidazole, so the results must be interpreted with caution. PMID- 10529880 TI - In vitro activity of the novel ketolide HMR 3647 and comparative oral antibiotics against Canadian respiratory tract isolates of Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis. AB - The in vitro activities of HMR 3647, erythromycin A, clarithromycin, azithromycin, roxithromycin, penicillin G, ampicillin, cefuroxime, trimethoprim/sulfamethoxazole, tetracycline, ciprofloxacin, and levofloxacin were determined for 1179 Streptococcus pneumoniae, 1438 Haemophilus influenzae, and 428 Moraxella catarrhalis isolated from respiratory tract specimens by 18 medical centers across Canada during 1997-1998. On a per weight basis, HMR 3647 was the most active agent tested against S. pneumoniae with MIC90s of < or = 0.12 microgram/mL for both penicillin susceptible and penicillin intermediate isolates and 0.25 microgram/mL for penicillin-resistant isolates. HMR 3647 was also highly active against M. catarrhalis (MIC90, < or = 0.12 microgram/mL), but less active against H. influenzae (MIC90, 4 micrograms/mL). PMID- 10529881 TI - In vitro susceptibility to pexiganan of bacteria isolated from infected diabetic foot ulcers. AB - During two clinical trials involving the treatment of 835 outpatients with infected diabetic foot ulcers, 2515 bacterial isolates, including 2337 aerobes and 178 anaerobes, were grown from cultures of the ulcers. The in vitro susceptibility of these isolates was determined to pexiganan, a peptide anti infective evaluated in these clinical trials, and to other classes of antibiotics. Pexiganan demonstrated broad spectrum antimicrobial activity against Gram-positive and Gram-negative aerobes and anaerobes. The MIC90 values for the most common species among 1735 Gram-positive aerobes isolated, such as Staphylococcus aureus, coagulase-negative staphylococci, Group A streptococci, and Group B streptococci, were 16 micrograms/mL or less. Of 602 Gram-negative aerobes tested, the MIC90 values for pexiganan were 16 micrograms/mL or less for Acinetobacter, Pseudomonas, Stenotrophomonas, Citrobacter, Enterobacter, Escherichia, Klebsiella, and Flavobacterium species. Pexiganan had a MIC90 of 4 to 16 micrograms/mL against the anaerobic isolates of Bacteroides, Peptostreptococcus, Clostridium, and Prevotella species. Importantly, pexiganan did not exhibit cross-resistance with other commonly used antibiotics, including beta-lactams, quinolones, macrolides, and lincosamides. The broad spectrum in vitro antimicrobial activity of pexiganan against clinical isolates from infected diabetic foot ulcers supports its potential as a local therapy for infected diabetic foot ulcers. PMID- 10529882 TI - Characteristics of pathogens causing urinary tract infections in hospitals in North America: results from the SENTRY Antimicrobial Surveillance Program, 1997. AB - Urinary tract infection (UTI) is common and involves pathogens with changing susceptibility patterns. The SENTRY Antimicrobial Surveillance Program evaluates international pathogen incidence patterns to detect and manage the emergence of resistant strains. We describe the antimicrobial resistance patterns among 1617 pathogens recovered from UTIs during the third-quarter of 1997 in North America (United States and Canada), as part of this worldwide program. The isolates were tested against more than 50 antimicrobial agents (20 reported) by reference broth microdilution methods, and selected isolates were characterized by pulsed-field gel electrophoresis (PFGE) and automated ribotyping. The five most frequently isolated species were Escherichia coli (48.6%), Enterococcus spp. (13.7%), Klebsiella spp. (12.0%), Pseudomonas aeruginosa (6.2%), and Enterobacter spp. or Proteus mirabilis (3.8% each). For each nation, imipenem and cefepime produced the widest spectrum of coverage among the beta-lactams and amikacin was best among the aminoglycosides. For Gram-negative species, high resistance among beta lactam antimicrobial agents was noted especially for various penicillins against E. coli (37.9% to 42.8%) and for the cephalosporins tested against enterococci (99.4% and 100%). Approximately 7.0% of enterococci in the USA were vancomycin resistant (88% with Van A). P. aeruginosa provided the most consistent levels of resistance, but the following agents were most active against these organisms: amikacin (96.6 to 97.4% susceptible), tobramycin (89.5 to 100.0%), piperacillin/tazobactam (89.5 to 100.0%), piperacillin (89.5 to 96.6%), imipenem (89.7 to 92.1%), cefepime (77.6 to 89.7%), and ceftazidime (82.9 to 86.2%). E. coli (2.2 to 2.7%), K. pneumoniae (6.2 to 6.4%), and a single Enterobacter cloacae strain produced extended-spectrum beta-lactamases; and five other Enterobacter spp. were likely to have expressed chromosomally mediated (Amp C) Stably derepressed cephalosporinases with associated resistance to ceftazidime (16.7 to 21.2% resistance). These data demonstrated that several UTI isolates in SENTRY hospitals have high levels of resistance to various classes of antimicrobial agents with little evidence of clonal dissemination. PMID- 10529883 TI - Multicenter evaluation of the antimicrobial activity for seven broad-spectrum beta-lactams in Turkey using the Etest method. Turkish Antimicrobial Resistance Study Group. AB - From March through July 1997, a nine laboratory surveillance project was initiated in Turkey to monitor the potency and spectrum of seven broad-spectrum antimicrobial agents (cefepime, ceftazidime, cefotaxime, imipenem, aztreonam, cefoperazone/sulbactam, and ticarcillin/clavulanate) tested against approximately 100 organisms (average 82; range 70 to 95 isolates) per participant center (736 strains). Eleven groups of organisms were tested by the Etest method (AB BIODISK, Solna, Sweden) with results validated by concurrent quality control strain analysis. Results from all centers were tabulated and 91.1% of quality assurance tests were within ranges recommended by the National Committee for Clinical Laboratory Standards. Among the seven beta-lactam-class drugs tested, imipenem and cefepime were the most active beta-lactams tested against all isolates. Overall, the rank order of susceptibility of the seven agents was imipenem > cefepime > cefoperazone/sulbactam > ceftazidime > cefotaxime > aztreonam > ticarcillin/clavulanate. Both cefepime and imipenem were active against ceftazidime-resistant strains of Enterobacteriaceae as well as against Streptococcus spp. and oxacillin-susceptible Staphylococcus aureus. Resistance phenotypes consistent with extended spectrum beta-lactamases were documented among Escherichia coli and Klebsiella spp., and profiles consistent with stably derepressed Bush-Jacoby-Mederios group 1 (Amp C) cephalosporinases were common among Enterobacter spp., Citrobacter spp., and Serratia spp. These data should be used to guide empiric therapy with beta-lactams in Turkey, and additionally will provide a reference statistical baseline to which future national studies of drugs in this class can be compared. PMID- 10529885 TI - Comparative activity of clinafloxacin and nine other compounds tested against 2000 contemporary clinical isolates from patients in United States hospitals. AB - The in vitro activity of clinafloxacin (formerly CI-960, AM-1091, PD-127391) was compared with other fluoroquinolones, cephalosporins, gentamicin, vancomycin, imipenem, piperacillin/tazobactam, clindamycin, and metronidazole against 2000 recent clinical strains from a large number of hospitals in the United States. Overall, clinafloxacin was the most active compound tested. Against Pseudomonas aeruginosa, clinafloxacin and ciprofloxacin demonstrated comparable activity (88% and 80% susceptible, respectively), and were four- to 16-fold more potent than levofloxacin (MIC90, 16 micrograms/ml) or trovafloxacin (MIC90, 32 micrograms/ml). Among anaerobic bacteria, clinafloxacin (MIC50s, 0.25-0.5 microgram/ml) and trovafloxacin (MIC50s, 0.5-2.0 micrograms/ml) were the most active quinolones, whereas metronidazole, imipenem and piperacillin/tazobactam were the most potent comparators. Clinafloxacin demonstrated sustained activity when compared to several available peer drugs against contemporary clinical isolates. The clinafloxacin spectrum against the 15 important pathogens monitored ranged from nil or 4.0% (vancomycin-resistant enterococci) to 100.0% (four different species) susceptible with an average percent susceptibility of 94.0%. This degree of potency and spectrum for clinafloxacin provides a wide potential for use against many species with established resistance to other anti-microbial classes. PMID- 10529884 TI - Ciprofloxacin in treatment of nosocomial meningitis in neonates and in infants: report of 12 cases and review. AB - Twelve cases of neonatal and infant nosocomial meningitis treated with intravenous ciprofloxacin in doses of 10 to 60 mg/kg/day are described. Four neonates were 21 to 28 days old and eight infants were 2 to 6 months old. Six presented with Gram-negative meningitis: Escherichia coli (2), Salmonella enteritidis (1), Acinetobacter calcoaceticus (1), two with two organisms, and (H. influenzae plus Staphylococcus epidermidis, Acinetobacter spp. plus S. epidermidis), and six were attributable to Gram-positive cocci (four S. aureus and two Enterococcus faecalis). Ten cases were cured. In two cases, reversible hydrocephalus appeared that responded to intraventricular punctures. In seven children, no neurologic sequellae appeared after a 2- to 4-year follow-up. One neonate had relapse of meningitis 3 months later and was ultimately cured, but developed a sequellae of psychomotoric retardation. Follow-up varied from 27 months to 10 years. Current published case reports from Medline on quinolone use in meningitis in neonates and infants are reviewed. PMID- 10529886 TI - Colony morphology of Candida spp. as a guide to species identification. AB - The presence of colony projections, often referred to as "feet," have typically been considered a characteristic of Candida albicans. In the current study that examined the colony morphology of numerous different species of Candida, several clinical isolates of Candida tropicalis and Candida krusei were also noted to produce "feet." The medium and growth conditions under which colony projections were produced by these species were characterized. PMID- 10529887 TI - Improved dissolution behavior of fenbufen by spherical crystallization. AB - Fenbufen is an analgesic, antipyretic and anti-inflammatory drug that is characterized by poor water solubility, a defect increased by very low wettability. Poor water solubility, particularly at low pH, could decrease absorption in the upper part of the gastrointestinal tract, which would be inconvenient for good bioavailability. Different spherical crystallization processes have been considered as methods to improve fenbufen dissolution behavior. A two-solvent system, in the presence of a bridging liquid, is the only method capable of producing spherical fenbufen crystals. In a first step, fenbufen solubility was considered in different solvents. The drug crystals formed were typically needle shaped. This characteristic was considered as a favorable parameter to obtain spherical crystals. After the selection of the best fenbufen solvent, several ratios of solvent (S)-nonsolvent (NS) (tetrahydrofuran [THF]-demineralized water) were studied. The addition of a bridging liquid (isopropyl acetate) improved spherical crystallization. The results from this method were reproducible batch to batch. The spherical crystals obtained showed a clear improvement in dissolution capacity, probably due to better wettability. Dissolution studies were then carried out on these spherical crystals stored for 1 month at different relative humidities (RHs). The dissolution profiles remained unchanged. PMID- 10529888 TI - Calculating the risk of batch failure in the manufacture of drug products. AB - A statistical tool has been developed to estimate the risk of batch failure in commercial production from the results of the development scale-up campaigns. This tool serves to indicate the robustness of the manufacturing procedure and its ability to meet the specifications set. Other uses include assistance in the resetting of specifications in the light of actual scale-up batch manufacturing experience and assessing the impact of changes in specifications proposed by the registration authorities. A template is given that simplifies the calculations required. PMID- 10529889 TI - Drug release from spray layered and coated drug-containing beads: effects of pH and comparison of different dissolution methods. AB - Based on dissolution profiles of three model drugs on spray layered beads with the same percentage of Aquacoat coating, it was concluded that in vitro dissolution of oral controlled-release formulations should be performed in both gastric and intestinal media for ionizable drugs. Ketoprofen (weak acid, pKa 4.8), nicardipine HCl (salt of weak organic base, pKa 8.6), and acetaminophen (very weak organic acid, pKa 9.7, not ionized at physiologic pH) provided different dissolution characteristics in enzyme-free simulated gastric fluid (pH 1.4) and enzyme-free simulated intestinal fluid (pH 7.4), indicating that the rate of drug release was pH dependent and related to drug ionization even though the solubility of the coating (ethylcellulose) is pH independent. In acidic media, ketoprofen release from the beads containing low-level coating (3%) was slower than that of nicardipine HCl, with the opposite holding true in basic media. Acetaminophen was released at approximately the same rate in both acidic and basic media. A comparison of drug release profiles for nicardipine HCl nude beads was also investigated among three different dissolution methods: USP dissolution apparatus I (basket method, 50 rpm), USP dissolution apparatus II (paddle method, 50 rpm), and USP dissolution apparatus III (Bio-Dis, Van-Kel Industries, 5 and 10 dpm). Release profiles obtained from all methods were similar, indicating that the three dissolution methods were comparable. PMID- 10529890 TI - The relationship between the rigidity of the liposomal membrane and the absorption of insulin after nasal administration of liposomes modified with an enhancer containing insulin in rabbits. AB - The relationship between the rigidity of the liposomal membrane and the absorption of insulin after nasal administration of liposomes modified with an enhancer containing insulin was investigated for the nasal delivery of peptide drugs in rabbits. The rigid liposomal membrane makes liposomes stable, protecting insulin from enzymatic degradation. Soybean-derived sterol (SS) or its sterylglucoside (SG) was used as an enhancer. Dipalmitoylphosphatidylcholine (DPPC) liposomes modified with SG had increased fluidity of the hydrophobic group of the liposome bilayer compared with the liposomes modified with cholesterol (Ch) or SS, as shown by measurements of the steady-state fluorescence anisotropy of 1,6-diphenyl-1,3,5,-hexatriene (DPH); however, the fluidity of the polar group of the liposome bilayer was decreased according to measurements of steady-state fluorescence anisotropy of dansylhexadecylamine (DSHA) at 37 degrees C. These findings suggest that the fluidity of the hydrophobic group of the liposome bilayer is responsible for the increase of liposomal leakage and instability of the liposomes. When insulin was administered nasally to rabbits as a solution, no hypoglycemic effect was observed. The administration of insulin contained in DPPC/SG (7/4, mole) liposomes with high fluidity caused a high glucose reduction of long duration (8 hr). DPPC/SS and DPPC/Ch (7/4) liposomes with low fluidity caused low glucose reductions. These results demonstrated that liposomes modified with SG can be useful as carriers of insulin administered nasally. PMID- 10529891 TI - Effect of divalent cations on the membrane properties of the lipid A analog E5531. AB - To obtain information on the effects of Mg2+ on the membrane properties of the lipid A analog E5531, we determined the size, structure, zeta potential, membrane fluidity, and micropolarity of the aggregates and the permeability of the E5531 membrane after the addition of Mg2+. E5531 forms a vesicle structure and within the molar ratio of [E5531]:[Mg2+] = 1:3, Mg2+ increased the zeta potential of the E5531 membrane, but did not change the size of the aggregates (approximately 20 nm). Within that molar ratio, Mg2+ decreased the membrane fluidity and micropolarity of E5531 and increased the phase transition temperature. Above the molar ratio of [E5531]:[Mg2+] = 1:5, the size of the aggregates was increased, but at [E5531]:[Mg2+] = 1:3, the size of the aggregates was similar to that in the absence of Mg2+ (approximately 20 nm), and we could stabilize the aggregates in rat plasma. PMID- 10529892 TI - Water determination in drugs containing ascorbic acid. AB - A new rapid analytical method was applied for water determination in tablets of vitamin C, Ce De Calcium Veterinary, and C-Tamin-500 containing ascorbic acid and therefore is not amenable for direct Karl Fischer (KF) titration. The method is based on the consecutive titration first of ascorbic acid by a novel reagent and then of water by a conventional KF reagent (KFR) in the same sample and cell with the electrometric "dead-stop" location of the end point in both titrations. The new reagent consists of iodine, potassium iodide, and sodium acetate in nonaqueous medium. Estimated repeatability and accuracy of both water and ascorbic acid determination are satisfactory. PMID- 10529893 TI - The evaluation of fine-particle hydroxypropylcellulose as a roller compaction binder in pharmaceutical applications. AB - In solid dosage manufacturing, roller compaction technology plays an important role in providing cost control and a quality product. The objective of this study was to evaluate the effectiveness of fine-particle hydroxypropylcellulose (HPC) as a dry binder in roller compaction processing. The formula included acetaminophen (APAP), microcrystalline cellulose, fine-particle HPC, croscarmellose sodium, and magnesium stearate. The fine-particle HPC was incorporated into the formula at 4%, 6%, and 8% w/w levels. Three compaction pressures (30, 40, and 50 bars) were used for each formulation. The roller compaction equipment used in this study had a processing capacity of 40 to 80 kg/hr. A tablet compression profile was generated on a rotary tablet press, and compression forces used were 5, 10, 15, 20, and 25 kN. The significant criteria for tablet evaluation were capping, hardness, friability, ejection force, and drug dissolution. As the binder concentration of HPC increased, tablet capping decreased, and tablet friability improved. As the concentration of HPC increased, only slight differences were noted in tablet hardness. All the formulations pass the USP requirement of 80% APAP dissolved within 30 min. Using 8% HPC could eliminate the formula capping problem. The friability results were less than 1% at all compression forces. The minimum tablet ejection forces were found in the formulations prepared under 40 bars compaction pressure. The utility of fine particle HPC as a roller compaction binder was established. The applicable binder concentrations and roller compaction pressures were found. Using HPC at these binder levels and operating parameters could overcome capping and friability problems and achieve the optimal tablet dosage forms. PMID- 10529894 TI - Investigating the fundamental effects of binders on pharmaceutical tablet performance. AB - For solid dosage forms, a better understanding of the fundamental properties of the binders helps in developing better formulations and products. The objective of this study was to determine the effects of binder toughness and plastic flow on tablet hardness, friability, and capping. The characteristic of binder toughness was determined, and the correlation between the ejection force of the tablet and the toughness of the binder was established. Evaluation was conducted using acetaminophen tablets with different kinds of binders (i.e., hydroxypropylcellulose, methylcellulose [MC], povidone [PVP], starch, etc.). A rotary tablet press was used for tableting at three different speeds. The properties of binders and acetaminophen tablets were determined using a diametral compression test. The toughness was measured as the curve of the area under the load versus deflection. The microbehavior of these binders was also studied. The acetaminophen tablets with the binders were subjected to predetermined loads and then examined under a scanning electron microscope. The tablets that contained hydroxypropylcellulose as a binder showed the highest toughness and had the lowest ejection force. The ejection force of tablets decreased with increasing concentrations of hydroxypropylcellulose in the dosage forms. The tablets that contained other binders failed by capping and random cracking in the middle. These results show that hydroxypropylcellulose, a thermoplastic polymer, provides the best physical characteristics for the tablets. This effect could help in improving tablet manufacturing conditions (e.g., compression force and speed). PMID- 10529895 TI - Preformulation: effect of moisture content on microcrystalline cellulose (Avicel PH-302) and its consequences on packing performances. AB - This study evaluates the influence of moisture content on the packing performances of a new grade of microcrystalline cellulose (MCC) (Avicel PH-302) either by classical method or by an unconventional compression technique (constant volume reduction of powder bed). An increase in moisture content decreases the apparent density of the powder bed, resulting from interparticulate friction enhancement. This modification of apparent density seems to be the main effect caused by the presence of humidity, which explains the variations of compression properties, like an increase of powder plasticity generally observed in the experimental conditions. PMID- 10529896 TI - Simultaneous determination of metronidazole benzoate, methylparaben, and propylparaben by high-performance liquid chromatography. AB - The simultaneous determination of metronidazole benzoate (MB), methylparaben (MP), and propylparaben (PP) in an oral suspension formulation was developed using high-performance liquid chromatography (HPLC). The method was developed using a Novapak C18 (3.9 x 150 mm, 4 microns) column, methanol-water (50:50, v/v) as the mobile phase and an ultraviolet (UV) detector at 254 nm. The peak area response versus concentration was linear in a concentration range from 40 to 400 micrograms/ml of MB, 0.8 to 8.0 micrograms/ml of MP, and 0.2 to 2.0 micrograms of PP. The correlation coefficients were 0.9997 for MB, 0.9987 for MP, and 0.9983 for PP, with relative standard errors of 1.12%, 1.28%, and 1.67%, respectively. PMID- 10529897 TI - The biology of pro-thyrotropin-releasing hormone-derived peptides. PMID- 10529898 TI - Peroxisome proliferator-activated receptors: nuclear control of metabolism. PMID- 10529899 TI - Orphan nuclear receptors: from gene to function. PMID- 10529900 TI - The role of androgens in male gender role behavior. PMID- 10529901 TI - Hyperhomocysteinemia and the endocrine system: implications for atherosclerosis and thrombosis. PMID- 10529902 TI - Studies on the treatment of tuberculosis undertaken by the British Medical Research Council tuberculosis units, 1946-1986, with relevant subsequent publications. PMID- 10529903 TI - The genetics of hearing loss. PMID- 10529904 TI - Development of inner ear (cochlear and vestibular) function in the fetus-neonate. AB - The development of function in the various receptors in the inner ear was studied in the neonatal rat, which is altricious with respect to hearing, using short latency evoked potentials, both auditory (ABR) and vestibular (VsEP). It was found that VsEPs could be recorded from all the vestibular end-organs by post natal day (PND) 8, whilst ABR could only be recorded from all animals on PND 14, showing the earlier onset of vestibular function in the inner ear. These results are discussed with relation to onset of inner ear function in the human fetus. PMID- 10529905 TI - Otological evaluation of newborns who failed otoacoustic emission screening. AB - Early identification of congenital hearing loss and early rehabilitation is extremely important. Otoacoustic emissions (OAE) are an efficient tool for hearing screening. Previous studies using click evoked otoacoustic emissions (CEOAEs) for newborn hearing screening resulted in approximately 70% pass rate, reflecting intact hearing. The aim of our study was to perform a detailed otological evaluation of newborns who failed OAE screening, using otoscopy, tympanometry and ABR. CEOAEs were recorded from 257 newborns prior to their release from the hospital. Those babies who did not pass the CEOAE were examined by DPOAE, otoscopy, tympanometry and ABR, if needed. 73% of all the newborns had CEOAE in both ears. 20% had CEOAE in only one ear. When the test was administered again three days postpartum, the CEOAE pass rate increased; 98% passed in at least one ear. Most of the newborns (84%) who failed had an obstruction of the external ear canal (collapsed ear canal or debris). There was a good correlation between the otoscopy and the tympanometry. Based on the above results, a newborn hearing screening protocol was introduced. PMID- 10529906 TI - Subclinical audiological findings in carriers of recessive genes for deafness. AB - The existence of subclinical signs in the hearing of carriers of recessive mutations for deafness has aroused much controversy in the literature. The present study comprised 30 carriers of recessive mutations for deafness, and a control group of 30 healthy volunteers, matched for gender and age. All participants underwent a series of hearing tests, including pure-tone audiometry, speech tests, Bekesy audiometry and notch noise tests. The main results were: hearing loss in high frequencies (3000 and 4000 Hz), an elevation of the acoustic reflex threshold, as well as an elevation of the identification of 2000 Hz pure tone in the presence of white noise and notch noise. A notch in the Bekesy audiogram was also identified in several carriers. An interaction was found between gender and the carrier trait in the hearing threshold at 4000 Hz, and in the ipsi- and contralateral acoustic reflex at 500 Hz and 1000 Hz. These subclinical signs may be complementary to DNA research in the investigation of genetic deafness of unknown origin. PMID- 10529907 TI - The perception of phonologically significant contrasts using speech envelope cues. AB - The observation that many cochlear implantees demonstrate high levels of speech recognition, despite limited or distorted spectral information, has motivated research on the importance of temporal information for the perception of speech. The purpose of this study was to measure the recognition of speech contrasts via only the speech envelope before and after training. Test stimuli consisted of eight segmental and two suprasegmental contrasts of the Hebrew Speech Pattern Contrast test using a binary forced-choice paradigm. Multiplying the speech waveform with white noise eliminated spectral information. Results show that stress, intonation and manner of articulation were very well perceived using only temporal information, whereas voicing and place of articulation were perceived above chance levels. Results also show that vowels were more susceptible to the removal of spectral information than consonants. These findings help to better understand speech perception performance of hearing-impaired individuals, including cochlear implant users. They may also have practical implications for aural rehabilitation and sensory aids design for the Hebrew speaking population. PMID- 10529908 TI - Auditory brain-stem evoked potentials in patients with thyroid and parathyroid dysfunction: adaptation to chronic hormonal dysequilibrium. AB - Auditory Brainstem Evoked Potentials (ABEPs) and pure tone audiograms were obtained from 24 patients with parathyroid dysfunction (17 hypercalcemia and 7 hypocalcemia) and 12 patients with thyroid dysfunction (6 hyperthyroid and 6 hypothyroid) and from 10 control subjects. ABEPs were characterized by I-V interpeak latency difference at 10/sec click rate and by the effect of increasing stimulus rate to 55/sec. None of the ABEP measures were significantly affected by levels of serum calcium, thyroid hormones or their interactions. Moreover no correlation was found between biochemical and electrophysiological measures. This stability of ABEP measures contrasts with earlier reports on acute effects of calcium and thyroid hormonal levels on auditory brainstem evoked potentials. We propose that chronic calcium or thyroid hormonal homeostatic changes are associated with adaptive mechanisms resulting in normal function of the auditory brainstem. PMID- 10529910 TI - Strain distribution in the layered wall of the esophagus. AB - The function of the esophagus is to move food by peristaltic motion, which is the result of the interaction of the tissue forces in the esophageal wall and the hydrodynamic forces in the food bolus. To understand the tissue forces in the esophagus, it is necessary to know the zero-stress state of the esophagus, and the stress-strain relationships of the tissues. This article is addressed to the first topic: the representation of zero-stress state of the esophagus by the states of zero stress-resultant and zero bending moment of the mucosa-submucosa and the muscle layers. It is shown that at the states of zero stress-resultant and zero bending moment, these two layers are not tubes of smaller radii but are open sectors whose shapes are approximately cylindrical and more or less circular. When the sectors are approximated by circular sectors, we measured their radii, opening angles, and average thickness around the circumference. Data on the radii, thickness-to-radius ratios, and the opening angles of these sectors are presented. Knowing the zero-stress state of these two layers, we can compute the strain distribution in the wall at any in vivo state, as well as the residual strain in the esophageal wall at the no-load state. The results of the in vivo states are compared to those obtained by a conventional approach, which treats the esophageal wall as a homogeneous material, and to another popular simplification, which ignores the residual strains completely. It is shown that the errors caused by the homogeneous wall assumption are relatively minor, but those caused by ignoring the residual strains completely are severe. PMID- 10529909 TI - Effect of impact load on articular cartilage: cell metabolism and viability, and matrix water content. AB - Significant evidence exists that trauma to a joint produced by a single impact load below that which causes subchondral bone fracture can result in permanent damage to the cartilage matrix, including surface fissures, loss of proteoglycan, and cell death. Limited information exists, however, on the effect of a varying impact stress on chondrocyte biophysiology and matrix integrity. Based on our previous work, we hypothesized that a stress-dependent response exists for both the chondrocyte's metabolic activity and viability and the matrix's hydration. This hypothesis was tested by impacting bovine cartilage explants with nominal stresses ranging from 0.5 to 65 MPa and measuring proteoglycan biosynthesis, cell viability, and water content immediately after impaction and 24 hours later. We found that proteoglycan biosynthesis decreased and water content increased with increasing impact stress. However, there appeared to be a critical threshold stress (15-20 MPa) that caused cell death and apparent rupture of the collagen fiber matrix at the time of impaction. We concluded that the cell death and collagen rupture are responsible for the observed alterations in the tissue's metabolism and water content, respectively, although the exact mechanism causing this damage could not be determined. PMID- 10529911 TI - A poroelastic continuum model of the cupula partition and the response dynamics of the vestibular semicircular canal. AB - Using mixture theory, an axisymmetric continuum model is presented describing the response dynamics of the vestibular semicircular canals to canal-centered head rotation in which the cupula partition is modeled as a poroelastic mixture of interpenetrating solid and fluid constituents. The solid matrix of the cupula is assumed to behave as a linear elastic material, whereas the fluid constituent is assumed to be Newtonian. A regular perturbation analysis of the fluid dynamics in the canal provides a dynamic boundary condition, which acts across the cupula partition. Numerical solution of the coupled system of momentum equations provides the spatio-temporal displacement fields for both the fluid and solid constituents of the cupula. Results indicate that at frequencies above 1 Hz, the fluid constituent is dynamically entrained by the solid matrix such that their motions are bound as if to exist as a single component. The resulting high frequency response is consistent with the macromechanical response predicted by single-component viscoelastic models of the cupula. Below 1 Hz, the dynamic coupling between the fluid and solid constituents weakens and the transcupular differential pressure is sufficient to force fluid through the mixture with little deformation of the solid matrix. Results are sensitive to the precise value of the cupular permeability. One of the most important distinctions between the present analysis and previous impermeable models of the cupula arises at the micromechanical level in terms of the local fluid flow that is predicted to occur within the cupula and around the ciliary bundles and sensory hair cells. Another important result reveals that the permeation dynamics predicted below 1 Hz gives rise to the same low-frequency macromechanical response as would occur with an impermeable viscoelastic structure having a much greater stiffness. Current estimates of the mechanical stiffness of the cupula, based solely on afferent nerve data, may therefore overestimate the true value intrinsic to the solid matrix by as much as an order of magnitude. PMID- 10529912 TI - Analysis of indentation: implications for measuring mechanical properties with atomic force microscopy. AB - Indentation using the atomic force microscope (AFM) has potential to measure detailed micromechanical properties of soft biological samples. However, interpretation of the results is complicated by the tapered shape of the AFM probe tip, and its small size relative to the depth of indentation. Finite element models (FEMs) were used to examine effects of indentation depth, tip geometry, and material nonlinearity and heterogeneity on the finite indentation response. Widely applied infinitesimal strain models agreed with FEM results for linear elastic materials, but yielded substantial errors in the estimated properties for nonlinear elastic materials. By accounting for the indenter geometry to compute an apparent elastic modulus as a function of indentation depth, nonlinearity and heterogeneity of material properties may be identified. Furthermore, combined finite indentation and biaxial stretch may reveal the specific functional form of the constitutive law--a requirement for quantitative estimates of material constants to be extracted from AFM indentation data. PMID- 10529913 TI - Chained vesicles in vascular endothelial cells. AB - There is extensive ultrastructural evidence in endothelium for the presence of chained vesicles or clusters of attached vesicles, and they are considered to be involved in specific transport mechanisms, such as the formation of trans endothelial channels. However, few details are known about their mechanical characteristics. In this study, the formation mechanism and mechanical aspects of vascular endothelial chained vesicles are investigated theoretically, based on membrane bending strain energy analysis. The shape of the axisymmetric vesicles was computed on the assumption that the cytoplasmic side of the vesicle has a molecular layer or cytoskeleton attached to the lipid bilayer, which induces a spontaneous curvature in the resting state. The bending strain energy is the only elasticity involved, while the shear elasticity is assumed to be negligible. The surface area of the membrane is assumed to be constant due to constant lipid bilayer thickness. Mechanically stable shapes of chained vesicles are revealed, in addition to a cylindrical tube shape. Unfolding of vesicles into a more flattened shape is associated with increase in bending energy without a significant increase in membrane tension. These results provide insights into the formation mechanism and mechanics of the chained vesicle. PMID- 10529914 TI - Dynamic contrast-enhanced MRI and fractal characteristics of percolation clusters in two-dimensional tumor blood perfusion. AB - Dynamic contrast-enhanced magnetic resonance imaging (DE-MRI) of the tumor blood pool is used to study tumor tissue perfusion. The results are then analyzed using percolation models. Percolation cluster geometry is depicted using the wash-in component of MRI contrast signal intensity. Fractal characteristics are determined for each two-dimensional cluster. The invasion percolation model is used to describe the evolution of the tumor perfusion front. Although tumor perfusion can be depicted rigorously only in three dimensions, two-dimensional cases are used to validate the methodology. It is concluded that the blood perfusion in a two-dimensional tumor vessel network has a fractal structure and that the evolution of the perfusion front can be characterized using invasion percolation. For all the cases studied, the front starts to grow from the periphery of the tumor (where the feeding vessel was assumed to lie) and continues to grow toward the center of the tumor, accounting for the well documented perfused periphery and necrotic core of the tumor tissue. PMID- 10529915 TI - Airway closure: occluding liquid bridges in strongly buckled elastic tubes. AB - This paper is concerned with the airway closure problem and investigates the quasi-steady deformation characteristics of strongly collapsed (buckled) airways occluded by liquid bridges of high surface tension. The airway wall is modeled as a thin-walled elastic shell, which deforms in response to an external pressure and to the compression due to the surface tension of the liquid bridge. The governing equations are solved numerically using physiological parameter values. It is shown that axisymmetric configurations are statically unstable, as are buckled tubes whose opposite walls are not in contact. The quasi-steady deformation characteristics of strongly collapsed airways whose walls are in opposite wall contact show a pronounced hysteresis during the collapse/reopening cycle. Buckling is shown to occur over a short axial length with moderate circumferential wavenumbers. Finally, further implications of the results for the airway collapse/reopening problem are discussed. PMID- 10529916 TI - A nonlinear axisymmetric model with fluid-wall interactions for steady viscous flow in stenotic elastic tubes. AB - Arteries with high-grade stenoses may compress under physiologic conditions due to negative transmural pressure caused by high-velocity flow passing through the stenoses. To quantify the compressive conditions near the stenosis, a nonlinear axisymmetric model with fluid-wall interactions is introduced to simulate the viscous flow in a compliant stenotic tube. The nonlinear elastic properties of the tube (tube law) are measured experimentally and used in the model. The model is solved using ADINA (Automatic Dynamic Incremental Nonlinear Analysis), which is a finite element package capable of solving problems with fluid-structure interactions. Our results indicate that severe stenoses cause critical flow conditions such as negative pressure and high and low shear stresses, which may be related to artery compression, plaque cap rupture, platelet activation, and thrombus formation. The pressure filed near a stenosis has a complex pattern not seen in one-dimensional models. Negative transmural pressure as low as -24 mmHg for a 78 percent stenosis by diameter is observed at the throat of the stenosis for a downstream pressure of 30 mmHg. Maximum shear stress as a high as 1860 dyn/cm2 occurs at the throat of the stenoses, while low shear stress with reversed direction is observed right distal to the stenosis. Compressive stresses are observed inside the tube wall. The maximal principal stress and hoop stress in the 78 percent stenosis are 80 percent higher than that from the 50 percent stenosis used in our simulation. Flow rates under different pressure drop conditions are calculated and compared with experimental measurements and reasonable agreement is found for the prebuckling stage. PMID- 10529917 TI - A perturbation model for the oscillatory flow of a Bingham plastic in rigid and periodically displaced tubes. AB - An approximate analytical model for the pulsatile flow of an ideal Bingham plastic fluid in both a rigid and a periodically displaced tube has been developed using regular perturbation methods. Relationships are derived for the velocity field and dimensionless flow rate. The solution compares adequately with available experimentally measured oscillatory non-Newtonian fluid flow data. These solutions provide useful analytical models supporting experimental and computation studies of arterial blood flow. PMID- 10529918 TI - Mathematical model of gas bubble evolution in a straight tube. AB - Deep sea divers suffer from decompression sickness (DCS) when their rate of ascent to the surface is too rapid. When the ambient pressure drops, inert gas bubbles may form in blood vessels and tissues. The evolution of a gas bubble in a rigid tube filled with slowly moving fluid, intended to simulate a bubble in a blood vessel, is studied by solving a coupled system of fluid-flow and gas transport equations. The governing equations for the fluid motion are solved using two techniques: an analytical method appropriate for small nondeformable spherical bubbles, and the boundary element method for deformable bubbles of arbitrary size, given an applied steady flow rate. A steady convection-diffusion equation is then solved numerically to determine the concentration of gas. The bubble volume, or equivalently the gas mass inside the bubble for a constant bubble pressure, is adjusted over time according to the mass flux at the bubble surface. Using a quasi-steady approximation, the evolution of a gas bubble in a tube is obtained. Results show that convection increases the gas pressure gradient at the bubble surface, hence increasing the rate of bubble evolution. Comparing with the result for a single gas bubble in an infinite tissue, the rate of evolution in a tube is approximately twice as fast. Surface tension is also shown to have a significant effect. These findings may have important implications for our understanding of the mechanisms of inert gas bubbles in the circulation underlying decompression sickness. PMID- 10529919 TI - Conditions for equivalency of countercurrent vessel heat transfer formulations. AB - Previous models of countercurrent blood vessel heat transfer have used one of two, different, equally valid but previously unreconciled formulations, based either on: (1) the difference between the arterial and venous vessels' average wall temperatures, or (2) the difference between those vessels' blood bulk fluid temperatures. This paper shows that these two formulations are only equivalent when the four, previously undefined, "convective heat transfer coefficients" that are used in the bulk temperature difference formulation (two coefficients each for the artery and vein) have very specific, problem-dependent relationships to the standard convective heat transfer coefficients. (The average wall temperature formulation uses those standard coefficients correctly.) The correct values of these bulk temperature difference formulation "convective heat transfer coefficients" are shown to be either: (1) specific functions of (a) the tissue conduction resistances, (b) the standard convective heat transfer coefficients, and (c) the independently specified bulk arterial, bulk venous and tissue temperatures, or (2) arbitrary, user defined values. Thus, they are generally not equivalent to the standard convective heat transfer coefficients that are regularly used, and must change values depending on the blood and tissue temperatures. This dependence can significantly limit the convenience and usefulness of the bulk temperature difference formulations. PMID- 10529920 TI - Significance of nonsagittal power terms in analysis of a dynamic elastic response prosthetic foot. AB - Dynamic elastic response prosthetic feet generally utilize a solid ankle, limiting dominant motion to the sagittal plane. However, researchers often use total rotational ankle joint power in the analysis of these feet. This investigation measured joint power terms in each plane for the Carbon Copy High Performance prosthetic foot. The significance of the frontal and transverse plane terms was assessed. Addition of these terms to the dominant sagittal power term revealed only slight differences, indicating that the sagittal power term is likely sufficient. PMID- 10529921 TI - Joint surface modeling with thin-plate splines. AB - Mathematical joint surface models based on experimentally determined data points can be used to investigate joint characteristics such as curvature, congruency, cartilage thickness, joint contact areas, as well as to provide geometric information well suited for finite element analysis. Commonly, surface modeling methods are based on B-splines, which involve tensor products. These methods have had success; however, they are limited due to the complex organizational aspect of working with surface patches, and modeling unordered, scattered experimental data points. An alternative method for mathematical joint surface modeling is presented based on the thin-plate spline (TPS). It has the advantage that it does not involve surface patches, and can model scattered data points without experimental data preparation. An analytical surface was developed and modeled with the TPS to quantify its interpolating and smoothing characteristics. Some limitations of the TPS include discontinuity of curvature at exactly the experimental surface data points, and numerical problems dealing with data sets in excess of 2000 points. However, suggestions for overcoming these limitations are presented. Testing the TPS with real experimental data, the patellofemoral joint of a cat was measured with multistation digital photogrammetry and modeled using the TPS to determine cartilage thicknesses and surface curvature. The cartilage thickness distribution ranged between 100 to 550 microns on the patella, and 100 to 300 microns on the femur. It was found that the TPS was an effective tool for modeling joint surfaces because no preparation of the experimental data points was necessary, and the resulting unique function representing the entire surface does not involve surface patches. A detailed algorithm is presented for implementation of the TPS. PMID- 10529922 TI - A damage model for nonlinear tensile behavior of cortical bone. AB - To describe the time-dependent nonlinear tensile behavior observed in experimental studies of cortical bone, a damage model was developed using two internal state variables (ISV's). One ISV is a damage parameter that represents the loss of stiffness. A rule for the evolution of this ISV was defined based on previously observed creep behavior. The second ISV represents the inelastic strain due to viscosity and internal friction. The model was tested by simulating experiments in tensile and bending loading. Using average values from previous creep studies for parameters in the damage evolution rule, the model tended to underestimate the maximum nonlinear strains and to overestimate the nonlinear strain accumulated after load reversal in the tensile test simulations. Varying the parameters for the individual tests produced excellent fits to the experimental data. Similarly, the model simulations of the bending tests could produce excellent fits to the experimental data. The results demonstrate that the 2-ISV model combining damage (stiffness loss) with slip and viscous behavior could capture the nonlinear tensile behavior of cortical bone in axial and bending loading. PMID- 10529923 TI - Finite element analysis of vertebral body mechanics with a nonlinear microstructural model for the trabecular core. AB - In this study, a finite element model of a vertebral body was used to study the load-bearing role of the two components (shell and core) under compression. The model of the vertebral body has the characteristic kidney shape transverse cross section with concave lateral surfaces and flat superior and inferior surfaces. A nonlinear unit cell based foam model was used for the trabecular core, where nonlinearity was introduced as coupled elastoplastic beam behavior of individual trabeculae. The advantage of the foam model is that architecture and material properties are separated, thus facilitating studies of the effects of architecture on the apparent behavior. Age-related changes in the trabecular architecture were considered in order to address the effects of osteoporosis on the load-sharing behavior. Stiffness changes with age (architecture and porosity changes) for the trabecular bone model were shown to follow trends in published experimental results. Elastic analyses showed that the relative contribution of the shell to the load-bearing ability of the vertebra decreases with increasing age and lateral wall curvature. Elasto-plastic (non-linear) analyses showed that failure regions were concentrated in the upper posterior region of the vertebra in both the shell and core components. The ultimate load of the vertebral body model varied from 2800 N to 5600 N, depending on age (architecture and porosity of the trabecular core) and shell thickness. The model predictions lie within the range of experimental results. The results provide an understanding of the relative role of the core and shell in vertebral body mechanics and shed light on the yield and post-yield behavior of the vertebral body. PMID- 10529924 TI - A method for planar biaxial mechanical testing that includes in-plane shear. AB - A limitation in virtually all planar biaxial studies of soft tissues has been the inability to include the effects of in-plane shear. This is due to the inability of current mechanical testing devices to induce a state of in-plane shear, due to the added cost and complexity. In the current study, a straightforward method is presented for planar biaxial testing that induces a combined state of in-plane shear and normal strains. The method relies on rotation of the test specimen's material axes with respect to the device axes and on rotating carriages to allow the specimen to undergo in-plane shear freely. To demonstrate the method, five glutaraldehyde treated bovine pericardium specimens were prepared with their preferred fiber directions (defining the material axes) oriented at 45 deg to the device axes to induce a maximum shear state. The test protocol included a wide range of biaxial strain states, and the resulting biaxial data re-expressed in material axes coordinate system. The resulting biaxial data was then fit to the following strain energy function W: [equation: see text] where E'ij is the Green's strain tensor in the material axes coordinate system and c and Ai are constants. While W was able to fit the data very well, the constants A5 and A6 were found not to contribute significantly to the fit and were considered unnecessary to model the shear strain response. In conclusion, while not able to control the amount of shear strain independently or induce a state of pure shear, the method presented readily produces a state of simultaneous in-plane shear and normal strains. Further, the method is very general and can be applied to any anisotropic planar tissue that has identifiable material axes. PMID- 10529925 TI - How new is new, and is it better? PMID- 10529926 TI - Spatial relationship of motion automated perimetry and optic disc topography in patients with glaucomatous optic neuropathy. AB - PURPOSE: To compare the spatial relationship of focal motion automated perimetry (MAP) visual field defect with focal defect in optic disc topography. METHODS: Patients (n = 12) with focal MAP visual field loss and focal change in optic disc topography were studied. The MAP visual field was divided into 12 field zones representing retinal nerve fiber layer arcuate bundles. Zones of MAP loss were related to rim area ratio (RAR), which was obtained by dividing the rim area, measured by the Heidelberg Retina Tomograph (HRT; Heidelberg Engineering, Heidelberg, Germany), into 36 10 degrees sectors and then dividing the area of each sector by the total rim area for each subject. Rim area ratio was compared to a normative database (n = 76) to quantify change in optic disc topography. In these same patients, the spatial relationship between standard automated perimetry (SAP) and short-wavelength perimetry (SWAP) and optic disc topography was also assessed. RESULTS: Motion automated perimetry superior visual field zones 14 through 19 were most often associated with a reduction in RAR for inferior sectors 24 through 29, and inferior visual field zones 4 through 7 were most often associated with a reduction in RAR for superior temporal sectors 11 through 16. Similar spatial relationships were found between SWAP and SAP and the RAR. CONCLUSION: Focal MAP visual field loss and focal changes in optic disc topography are spatially related. This relationship is similar to that found between SWAP and SAP with optic disc topography. Focal thinning or notching detected by RAR analysis might be independent of the specific functional test employed. PMID- 10529927 TI - Kinetic and static fixation methods in automated threshold perimetry. AB - PURPOSE: Static fixation is the standard method for stabilizing the eye during automated perimetry. Kinetic fixation is an alternative for fixation control in which the eye follows a moving target. This study was conducted to evaluate the fixation accuracy of static and kinetic fixation perimetry and to determine their ability to detect the absolute scotoma of the physiologic blind spot. METHODS: The 71 patients with early glaucomatous field loss (mean age 65 years) and 45 control subjects (mean age 57 years) recruited from five clinical sites underwent threshold testing on the Dicon perimeter (kinetic fixation; Vismed, San Diego, CA) and Humphrey Field Analyzer (static fixation). The frequency of Heijl-Krakau fixation catch-trial errors was used as an indicator of fixation accuracy, and the measured sensitivity at the physiologic blind spot was used as an indicator of perimetric accuracy. RESULTS: In patients with glaucoma, the frequency of fixation errors was significantly greater for kinetic fixation (17.2%) than for static fixation (10.2%). In the control group, the frequency of fixation errors also was significantly greater for kinetic fixation (27.5%) than for static fixation (12.6%). The threshold at the presumed location of the blind spot (15 degrees temporal, 3 degrees inferior from fixation) was 14.8 dB using kinetic fixation versus 4.0 dB with static fixation in patients with glaucoma, and 18.5 dB using kinetic fixation versus 2.5 dB using static fixation in the control group. CONCLUSION: Relative to static fixation, kinetic fixation was associated with fixation inaccuracy and underestimation of the absolute scotoma at the physiologic blind spot. PMID- 10529928 TI - Evaluation of a portable fundus camera for use in the teleophthalmologic diagnosis of glaucoma. AB - PURPOSE: The digital images of the optic disk from a portable fundus camera were evaluated for suitability in teleophthalmologic screening for glaucoma. METHODS: Fifty-one eyes of 27 consecutive patients from our glaucoma clinic were dilated and photographed with a Zeiss FF retinal camera (Carl Zeiss, Oberkochen, Germany) and a portable Nidek NM-100 (Nidek, Tokyo, Japan) fundus camera. Digital images from the portable fundus camera were digitized, compressed and stored in a Fujix DF-10M (Fuji, Tokyo, Japan) digitizer. Lossy compressed digital images and photographs from the Zeiss camera were presented separately in random order to three ophthalmologists for estimation of vertical cup:disk ratios (VCDR) and to evaluate image quality as good, acceptable, or unacceptable for screening glaucoma. Gold standard VCDRs were measured from monoscopic photographic slides obtained using the Zeiss camera by a fourth ophthalmologist. RESULTS: Measurement of agreement (Kappa values) between estimated VCDR of digital images and photographs by the three ophthalmologists were 0.52, 0.38, and 0.50 respectively. Agreement between gold standard and estimated VCDR from photographs were 0.87, 0.45, and 0.84 respectively (specificity between 79% and 97%, sensitivity between 70% and 95%). Kappa values obtained between gold standard and estimated VCDR from digital images were 0.52, 0.49, and 0.49, respectively (specificity between 68% and 79%, sensitivity between 67% and 87%. CONCLUSION: Moderate to good agreement indicates that the digital images from the portable fundus camera may be suitable for optic disk assessment in the current configuration. This easy to use Nidek hand-held camera could be a viable instrument for teleophthalmology if a better digitizing system is incorporated to improve the quality of the images. PMID- 10529930 TI - Autologous patch graft in tube shunt surgery. AB - PURPOSE: To evaluate an alternate method of covering the subconjunctival portion of the tube in aqueous shunt surgery. Evidence of tube erosion, graft-related infection, graft melting, or other associated intraocular complications were evaluated. METHODS: A retrospective study of 16 patients (17 eyes) who underwent tube shunt surgery at Wills Eye Hospital between July 1991 and October 1996 was conducted. An autologous either "free" or "rotating" scleral lamellar graft was created to cover the subconjunctival portion of the tube shunt. RESULTS: All patients were evaluated for at least 6 months, with a mean follow-up of 14.8 months (range 6-62 months). All eyes tolerated the autologous graft well, with no clinical evidence of tube erosion, or graft-related or intraocular complications. CONCLUSION: Autologous patch graft in tube shunt surgery appears--in selected cases--to be an effective, safe and inexpensive surgical alternative to allogenic graft materials. It also offers ease of availability, and eliminates the risk of transmitting infectious disease. PMID- 10529929 TI - The effect of mitomycin C after long-term storage on human Tenon's fibroblast proliferation. AB - PURPOSE: To investigate the effect of mitomycin C (MMC) after long-term storage on proliferation of human Tenon's fibroblasts in vitro. METHODS: Human Tenon's fibroblasts in tissue culture were exposed for 5 minutes to MMC (0.4 mg/mL) that was either freshly prepared or had been stored for as long as 18 months at either 4 degrees C or -20 degrees C. The MTT colorimetric assay was used to determine the inhibition of proliferation as measured indirectly by mitochondrial activity. RESULTS: The inhibition rate was 88% using fresh MMC, and declined to a mean of 73% when using MMC that had been stored for as long as 18 months at 4 degrees C; this decrease was not statistically significant. The mean inhibition for MMC stored at -20 degrees C was 68%, and this was significantly less than inhibition with fresh MMC. Inhibition did not vary significantly with MMC after different storage times. CONCLUSION: Mitomycin C continues to have strong in vitro antiproliferative effects when stored for as long as 18 months at 4 degrees C or 20 degrees C. A significant decline in potency compared with fresh MMC occurs when MMC is stored at -20 degrees C. PMID- 10529931 TI - Results of another modality for extending glaucoma drainage tubes. AB - PURPOSE: For a variety of reasons, on rare occasions a surgeon needs to reposition the intraocular portion of a glaucoma drainage implant tube, but finds the tube "too short" to do so. This study describes results of a different technique for "extending" the tube (rather than replacing the entire apparatus). METHODS: Four eyes of four patients required a tube "extender," for either tube tip blockage associated with uncontrolled intraocular pressure (IOP; n = 3), or to avoid total seton replacement after tube "cheesewiring" (n = 1). An extender was fashioned from common angiocatheter material. Postoperative complications, IOP, and need for further surgery were reviewed. RESULTS: None of the four patients required further, more invasive surgery or experienced any related postoperative complications. Final IOP averaged 11.5 +/- 4.2 mmHg (range 6-16 mmHg). CONCLUSION: Tube extension using angiocatheter material is a viable, cost effective option in these difficult cases, saving the surgeon from having to explant and replace the entire implant. PMID- 10529932 TI - The dorzolamide/timolol combination versus timolol plus pilocarpine: patient preference and impact on daily life. United States Patient Preference Study Group. International Patient Preference Study Group. AB - PURPOSE: To compare the 2.0% dorzolamide/0.5% timolol fixed combination (COSOPT; Merck & Co., Whitehouse Station, NJ) to 0.5% timolol plus 2.0% pilocarpine given concomitantly, and to determine patient preference, tolerability, and impact on daily life in patients with elevated intraocular pressure (IOP). METHODS: Two multi-center, randomized, cross-over, observer masked studies were conducted, one in the United States (97 patients) and one in Europe (93 patients). The Comparison of Ophthalmic Medications for Tolerability questionnaire was used to assess patient preference and perception of side effects and activity limitations resulting from study medications. Intraocular pressure was measured before and 2 hours after the morning dose of study medication (hour 0 and hour 2). RESULTS: In both studies, among patients with a preference, the combination was preterred to timolol plus pilocarpine by a ratio of 4 to 1. The most commonly cited reason for this preference was side effects. Patients in both studies also reported that the combination interfered significantly less with daily life in terms of side effects and activity limitations. They also reported missing significantly fewer doses of study medication while taking the combination and being significantly more satisfied with it. The efficacy of these two treatments was not significantly different, based on IOP measurements at hour 0 and 2 hours after administration. Patients reported significantly more adverse events while receiving timolol plus pilocarpine in both studies, and in the U.S. study, significantly more patients discontinued therapy while receiving timolol plus pilocarpine than while receiving the combination. CONCLUSION: Compared with timolol plus pilocarpine, patients preferred the combination of 2% dorzolamide/0.5% timolol, and reported less interference in daily activities, better tolerability, and better compliance with therapy. PMID- 10529933 TI - Management of chronic choroidal effusions. PMID- 10529934 TI - How does the trabecular meshwork regulate outflow? Clues from the vascular endothelium. AB - Although it is well known that the trabecular meshwork plays a central role in regulating the outflow of aqueous humor from the eye, the cellular mechanisms and events responsible for this function are poorly understood. In contrast, the mechanisms by which vascular endothelial cells modulate the permeability of blood vessels have been more thoroughly investigated. It is hypothesized that the cells of the TM employ mechanisms similar to those observed in the vascular endothelium to modulate aqueous humor permeability. Specifically, it is hypothesized that the cells of the TM employ Na-K-Cl cotransport to modulate their intracellular volume and thus the volume of the paracellular pathways through which aqueous humor may travel. The current knowledge about the role of Na-K-Cl cotransport and volume regulation in the regulation of vascular permeability, and evidence that similar physiologic events occur in the TM are reviewed. In addition, the implications for further study of aqueous humor outflow physiology are discussed. PMID- 10529935 TI - Transpupillary argon laser cyclophotocoagulation in the treatment of traumatic glaucoma. AB - PURPOSE: A patient with traumatic glaucoma who underwent transpupillary argon laser cyclophotocoagulation for management of uncontrolled intraocular pressure (IOP) despite maximally tolerated medical therapy is discussed. METHODS: In this patient, pars plana vitrectomy, lensectomy, and removal of 180 degrees of necrotic iris had been performed after a blunt trauma with a bungee cord. Six weeks after surgery, the patient presented with an IOP of 40 mmHg despite therapy with three aqueous suppressants. The patient refused further surgical intervention and opted for transpupillary argon laser cyclophotocoagulation (TALC). The laser setting was 1,000 mW, with a 50-micron spot size for 0.1 second. A total of 293 laser exposures through a Goldmann contact lens was administered to all visible ciliary processes over 180 degrees where iris structures were absent. RESULTS: Ten weeks after TALC, the patient's IOP remained controlled with medications at 16 mmHg, and visual acuity had improved to 20/25 with an aphakic contact lens. CONCLUSION: In selected patients whose ciliary processes are visible with indirect gonioscopy due to the defect in the iris, TALC may be an effective alternative cyclodestructive procedure to lower IOP when conventional medical or laser treatments are not successful. PMID- 10529936 TI - "Parapapillary" versus "peripapillary". PMID- 10529937 TI - WHACS your patients. PMID- 10529939 TI - An outcomes study of an occupational medicine intervention program for the reduction of musculoskeletal disorders and cumulative trauma disorders in the workplace. AB - Upper-extremity musculoskeletal pains or disorders (MSDs) account for a significant number of work-related illnesses in the US workforce. Although the concept of MSD prevention is appealing, little has been done to demonstrate the successful application and benefit of these programs. In 1995, an aircraft manufacturer established a unique risk-management program based on the individual risk assessment (CtdMAP) for new hires. The MSD intervention program was designed to prospectively evaluate each new employee for their individual risk of developing MSDs in the workplace. Before job placement, individuals at higher risk were assigned to a period of transitional work. Workers' compensation costs decreases per year were 16%, 3%, 24%, and 12%, while work hours increased by 56%. Employer-estimated savings in direct workers' compensation costs per year were $469,990, $678,337, $1,936,105, and $1,995,759. PMID- 10529940 TI - Health effects costs of particulate air pollution. AB - We conducted a cross-sectional study in December 1994 in three metropolitan areas of the Rhone-Alpes region in France (Lyon, Grenoble, and Chambery; total number of inhabitants = 970,000) to assess the medical costs resulting from exposure to particulate air pollution. Probability samples of the general population (508 families, 1265 subjects) and of the physicians (395) and 13 hospital respiratory care and emergency units in the study area provided data on the prevalence of respiratory disorders and on medical care usage. Measurements from air-quality monitoring networks were used to ascribe a fraction of the respiratory morbidity to the ambient air particle concentrations present during the study period, on the basis of attributable risk estimates drawn from recent meta-analyses. The medical care usage and absenteeism related to respiratory disorders were converted into direct and indirect medical and social costs by use of a "cost of illness" approach. These costs were extrapolated to annual costs of disease attributable to particulate pollution in 1994, using daily values of air pollution. The average particulate concentrations during the study period were moderate (39, 41, and 10 micrograms/m3 in the three cities), yielding attributable fractions that ranged between 0.6% and 13.8% according to the health condition and to the city. Three hundred ninety-five subjects reported respiratory symptoms (prevalence, 31.2%) during the study period; 1182 patients visited a doctor and 158 used hospital services. The extrapolated annual estimates of the attributable cost of respiratory diseases for a population of 1 million range between 79 and 135 million French francs (FF) (20th and 80th percentiles of the cost distribution, after a Monte Carlo simulation, respectively; 50th percentile, 107 x 10(6) FF [16.3 x 10(6) Euros]). Over-the counter drug consumption represents the largest cost item (approximately 44% of total costs), followed by wage losses (38%). Hospital expenditures amount to a low percentage of total costs (about 5%) because most respiratory disorders do not require hospital care. Mortality was not considered in this study. Most of these costs occur at relatively low levels of air pollution (67% of the total annual costs are incurred during days with particle concentrations lower than 50 micrograms/m3). Such substantial figures are useful for assessing the social impacts of air pollution and for evaluating the cost efficiency of abatement policies. PMID- 10529938 TI - Call for an international ban on asbestos. AB - To eliminate the burden of disease and death that is caused worldwide by exposure to asbestos, the Collegium Ramazzini calls for an immediate ban on all mining and use of asbestos. To be effective, the ban must be international in scope and must be enforced in every country in the world. PMID- 10529941 TI - Occupational health and safety management systems in the Canadian pulp and paper industry: methods of auditing. AB - The Canadian Pulp and Paper Association has developed eight "Guiding Principles for Management of Occupational Health and Safety" (OH&S) for its member companies. As part of a study to assess member companies' OH&S activities, an on site audit was performed for 11 sites. The audit assessed five key components of a management system for a number of OH&S activities. Management-system components more likely to be in place included system ownership and goals and procedures, whereas measures of performance, a review of measures, and corrective action were less likely to be present. Environmental surveillance and injury reduction were most actively monitored, as indicated by the number of measures of performance relating to these activities. The auditing process demonstrated leadership and communicated the OH&S priorities of the Canadian Pulp and Paper Association to the sites. PMID- 10529942 TI - The role of health risk factors and disease on worker productivity. AB - The costs attributed to employee health problems are usually measured by employers in terms of direct health care costs, such as medical plan claims. Although it has been understood that employee health problems also produce indirect costs for employers, their measurement has been far less frequent. At best, studies have recorded one component of indirect health costs: the time lost to employee absenteeism and disability. The study presented here includes a measure of the actual decrease in the productivity of employees while they are on the job, in addition to measures of absenteeism and disability. These three measurements were combined to produce a Worker Productivity Index (WPI). The WPIs of 564 telephone customer-service agents were correlated with the employees' number and type of health risks, as measured by a Health Risk Appraisal. Additionally, the WPI was also examined across different disease states in the same population of employees. As the number of health risks increased, an employee's productivity decreased. The nature of the health risk may also differentially affect the pattern of the decrease. Finally, disease states were also associated with different patterns of productivity reduction. PMID- 10529943 TI - Occupational ingestion of P-32: the value of monitoring techniques to determine dose. A case report. AB - The purpose of this article is to described the analytical methods used to assess the internal dose from a P-32-labeled compound that was inadvertently ingested. Bioassay data, using the International Commission on Radiation Protection (ICRP) 30 model, enabled the calculation of internal dose. Whole body counting (WBC) and urinary measurement with liquid scintillation counting were utilized to estimate the amount of radioactive material deposited in body organs. This metabolic model assumes that 80% of the material ingested is absorbed through the gastrointestinal tract because P-32 is soluble. The time of the intake, a critical variable in this method, was estimated on the basis of urine contamination of clothing. Twenty-four-hour urine sampling over a 6-week period, coupled with daily WBC over the same period, was performed. Because P-32 does not emit photons, WBC relied on measuring the bremsstrahlung radiation produced as a result of interaction of beta radiation with the body's tissues. A P-32-spiked phantom was used as a control. Over the 6-week monitoring period, urinary results indicated an ingestion of 560 microCi of P-32, whereas WBC estimated on intake of 580 microCi. An assessment of the laboratory where the accident occurred indicated that approximately 600 microCi of radioactive phosphorous was missing. The total effective dose equivalent was estimated at 4.8 rem (48 mSv). On the basis of this study, the ICRP model appears to fit the data obtained from urine measurements and WBC. No symptoms were noted from the ingestion of 580 microCi. The committed organ doses were well within the occupational nonstochastic limits of 50 (0.5 Sv) permitted by the Nuclear Regulatory Commission. These results were confirmed by NUREG/CR-4884 and commercial software (CINDY). This report confirms the value of using the ICRP-30 model with urinary measurements and WBC to estimate the dose received as a result of ingestion of radioactive P-32. PMID- 10529944 TI - Work-related upper-extremity disorders: prospective evaluation of clinical and functional outcomes. AB - The purpose of this study was to describe the demographic, vocational, medical, workplace, and psychosocial characteristics of patients treated for work-related upper-extremity disorders, to document treatment patterns in a community-practice setting, and to determine which of these factors predicts subsequent employment and functional status outcomes. A questionnaire was administered by mail or telephone to 112 patients seen at the University of Massachusetts Occupational Upper Extremities Disorders Clinic and included measures of disease-specific functional status, pain, reactions to pain, employer-employee relations, and number and type of interventions used to treat the disorder. Results were compared with baseline data obtained, on average, 16 months prior to follow-up. Of the original cohort (n = 124), 112 participated in the prospective study. Although most patients reported improvement in pain severity, fear of pain, life situation, and functional status, there was little change in employment status. Patients' self-reported intentions of return to work at baseline did not predict work status at follow-up. In general, those who were employed at baseline remained employed, had a greater reduction in symptom severity over time, and were significantly more likely to report improvement in their problem than those who were unemployed. The efficacy of various interventions was examined by type, mix, and intensity (number of different interventions undergone by the patient). No positive relationship was found between these measures and employment status, self-reported change in the problem, or self-reported improvement in functional status. Significant negative relationships were found between surgery, psychotherapeutic interventions, and outcomes. This was likely to have occurred because of a selection bias toward the more chronic and severely disabled patients for these treatments. However, the relative ineffectiveness of such intensive interventions as surgery in improving the work and health status of chronically symptomatic work-related upper-extremity patients cannot be overlooked. The findings suggest that more emphasis be placed on interventions aimed at resolving differences between employers and injured employees. More careful selection of patients for expensive and invasive procedures is recommended. PMID- 10529945 TI - Job characteristics as predictors of neck pain. A 4-year prospective study. AB - Data from a community-based 4-year prospective study were used to investigate job characteristics as predictors of neck pain. Of 1791 working responders who completed a questionnaire in 1990, 1429 (79.8%) returned a second questionnaire 4 years later (1994). In responders without neck pain during the previous 12 months in 1990, the "little influence on own work situation" factor predicted neck pain during the previous 12 months (odds ratio = 2.21; 95% confidence interval, 1.18 to 4.14) and previous 7 days in 1994 (OR = 2.85; 95% confidence interval, 1.21 to 6.73) after adjustment for a series of potential confounders. In responders with neck pain in 1990, the little influence on own work situation factor was associated with persistent neck pain 4 years later. The study indicates that having little influence on one's own work situation is a predictor of neck pain. PMID- 10529946 TI - Chemical exposures of rocket-engine test-stand personnel and cancer mortality in a cohort of aerospace workers. AB - We conducted a retrospective cohort study of 6107 aerospace workers to examine whether exposure to chemicals--primarily hydrazine fuels--during rocket-engine fueling and testing affects cancer mortality. When conditional logistic regression analysis was applied and adjusted for confounding variables, the estimated rate ratio for lung cancer mortality, comparing exposed to unexposed workers from the same facility, ranged from 1.68 (95% confidence interval, 1.12 to 2.52) to 2.10 (95% confidence interval, 1.36 to 3.25), depending on job duration threshold (6 or 24 months) and lag (0 to 15 years). Similar results were obtained for hemato- and lymphopoietic cancer and for bladder and kidney cancer mortality, but estimates for these cancers were imprecise. We concluded that occupational exposure to hydrazine or other chemicals associated with rocket engine testing jobs increased the risk of dying from lung cancer, and possibly other cancers, in this population of aerospace workers; however, our results need to be replicated in other populations. PMID- 10529947 TI - State licensing laws and the interstate practice of occupational medicine. AB - In 1996, the Occupational Health Law and Policy Section of the American College of Occupational and Environmental Medicine (ACOEM) was asked to undertake a study of state licensing regulations after the prosecution of an ACOEM member for practicing medicine without a license. In response to that member's experience, the ACOEM House of Delegates passed a resolution asking the College to lobby individual states for an exclusion to their licensing acts for occupational and environmental physicians. Recognizing the tremendous obstacles to this task, the ACOEM Board of Directors then referred the issue to the Occupational Health Law and Policy Section for further study and analysis. What follows is a report of that study, including the results of a survey mailed to the licensing authorities of the 50 US states and four US territories. The results of this study are not meant to offer advice to College members regarding compliance with specific state licensing regulations, nor does it define the official position of the states that responded. States that responded were careful to disclaim their responses as the official position of their states' agencies and were assured that the responses were provided for informational purposes only. The purpose of the survey was not to provide information for reference but simply to identify general trends and document the various positions that states may take on certain licensing issues. PMID- 10529948 TI - Are salivary immunoglobulin A and lysozyme biomarkers of stress among nurses? AB - Salivary immunoglobulin A (IgA) and lysozyme have been studied as possible biomarkers of stress. This study examined the stress levels among female nurses in various units and the relationship between these stress levels and salivary IgA and lysozyme secretion. One hundred ninety-five (43%) of 457 eligible female nurses from surgical wards/operating theaters (SURG), medical wards (MED), and outpatient clinics/day-surgery theaters (OPD) completed a self-administered questionnaire. From this group of 195 nurses, 124 provided a salivary sample accumulated over 5 minutes. Stress levels were assessed with a ten-point Stress Assessment Score (SAS) for Asians and a direct question on perceived life stress. Enzyme-linked immunosorbent assay and lyso-plate methods were used to determine salivary IgA and lysozyme levels. Forty-five percent of SURG, 35% of MED, and 17% of OPD nurses scored at least four points on the SAS. SURG nurses had the lowest IgA secretion (geometric mean; 95% confidence interval [CI]) rates (43 micrograms/min; 36 to 51 micrograms/min). The other groups had significantly higher salivary IgA secretion rates: MED (96 micrograms/min; 80 to 116 micrograms/min) and OPD (77 micrograms/min; 60 to 98 micrograms/min) Findings for salivary lysozyme (microgram/min) were similar; SURG (9 micrograms/min; 6 to 13 micrograms/min) MED (19 micrograms/min; 12 to 28 micrograms/min) and OPD (16 micrograms/min; 9 to 28 micrograms/min). The salivary IgA (Spearman's r = -0.22, P = 0.01) but not the lysozyme (Spearman's r = -0.01, P = 0.9) secretion rate correlated negatively with SAS. Nurses working in various units under different conditions experienced dissimilar levels of stress. Salivary IgA, but not lysozyme, correlated inversely with self-reported levels of stress. It may thus be a potential biomarker in future studies on stress. PMID- 10529949 TI - Quality of life and health-services utilization in a population-based sample of military personnel reporting multiple chemical sensitivities. AB - We sought to assess quality of life and health-services utilization variables in persons with symptoms suggestive of multiple chemical sensitivity/idiopathic environmental intolerance (MCS/IEI) among military personnel. We conducted a cross-sectional telephone survey of a population-based sample of Persian Gulf War (PGW) veterans from Iowa and a comparison group of PGW-era military personnel. A complex sample survey design was used, selecting subjects from four domains: PGW active duly, PGW National Guard/Reserve, non-PGW active duty, and non-PGW National Guard/Reserve. Each domain was substratified by age, gender, race, rank, and military branch. The criteria for MCS/IEI were developed by expert consensus and from the medical literature. In the total sample, 169 subjects (4.6%) of the 3695 who participated (76% of those eligible) met our criteria for MCS/IEI. Persons who met the criteria for MCS/IEI more often reported the following than did other subjects: more than 12 days in bed due to disability, Veteran's Affairs disability status, Veteran's Affairs disability compensation, medical disability, and unemployment. MCS/IEI cases also had higher outpatient rates of physician visits, emergency department visits, and inpatient hospital stays. Subjects who met the criteria for MCS/IEI more often reported impaired functioning on each Medical Outcomes Study 36-Item Short Form subscale, compared with those who did not meet the criteria. We concluded that although the diagnosis of MCS/IEI remains controversial, the persons who met our criteria for the disorder are functionally impaired. PMID- 10529950 TI - Accreditation of methadone programs proposed. PMID- 10529952 TI - How to COPE with serious mental illness. PMID- 10529951 TI - The high cost of anxiety. PMID- 10529953 TI - Sense of purpose enhanced by participation. PMID- 10529954 TI - Dr. Smoyak speaks on stalking. PMID- 10529955 TI - 'Women think, men drink'. PMID- 10529956 TI - The sins of the father... PMID- 10529957 TI - Measuring the psychological aspect of pain. PMID- 10529958 TI - Cognitive behavioral group therapy for residents in assisted-living facilities. AB - Geriatric depression is a disabling illness that is associated with increased morbidity and mortality and deserves aggressive and early intervention. Cognitive behavioral group therapy is an effective intervention for the treatment of geriatric depression and can be used with residents in an assisted living facility to manage and prevent depression. Cognitive-behavioral group therapy protocol modifications for residents in assisted-living facilities aids their participation and learning. PMID- 10529959 TI - Managed care. The effect of case management on state psychiatric clients. AB - This study examined whether case management services, mandated under the managed care contract for adult clients in a medium-sized state psychiatric hospital in Tennessee between July 1996 and June 1997, were offered as specified, and the impact these services had on recidivism for individuals who were identified as having a severe or persistent mental illness. Although all of the clients were offered case management, 47% refused the service. Of the 14 who had one or more readmissions, six (43%) had case management. These findings demonstrate that health care providers must offer sufficient information to their clients so that they can use the managed care system more effectively. PMID- 10529960 TI - Psychiatric nursing for the 21st century. The PACED model. AB - Health care reform has had a profound effect on the way psychiatric treatment occurs in this country. Decreasing length of stay, increasing acuity, and reducing staff levels are making traditional approaches to mental health care infeasible. Resources already in place may still be focusing on long-term treatment issues and do not facilitate rapid stabilization and discharge planning that includes continued care within an integrated system. Research supports the feasibility of quality mental health care, which can be accomplished in shortened lengths of stay, as long as clinical managers plan inpatient programs focused on short-term goals followed by appropriate aftercare. In addition to recommendations for clinical managers, this article provides a proposal for executives redesigning a mental health care delivery system, which includes the goals of rapid assessment and stabilization, as well as discharge planning and appropriate follow up within an integrated system. PMID- 10529961 TI - Identifying depression as a symptom of sleep apnea. AB - Sleep apnea syndrome is a common, life-threatening disorder, causing significant physical, emotional, and social problems. The symptoms of obstructive sleep apnea syndrome can lead to an erroneous diagnosis of dysthymia. For patients with sleep apnea syndrome, standard antidepressant treatments may exacerbate the condition. DSM-IV's diagnostic model needs only a small change to allow consideration of sleep apnea syndrome as a cause of a mood disorder. PMID- 10529962 TI - Bridging gaps between mind, body, & spirit. Healing the whole person. AB - Fifty percent of visits of primary care providers are for psychiatric problems making it desirable to screen for mental, addictive, or behavioral disorders at the level of primary care. Psychiatric/mental health nurses prepared at the master's level to practice in the blended clinical specialist/nurse practitioner role are well placed to treat or collaborate in the treatment of people who present with symptoms of physical or psychological problems. The role of the clinical specialist/nurse practitioner is evolving in response to changes in health demographics, epidemiology, scientific and technological advances, and changes in managed care. Advanced practice nursing education must continue to anticipate and meet on-going changes and challenges. PMID- 10529963 TI - Voices from the field--a qualitative analysis of classroom, school, district, and state health education policies and programs. AB - This study provides a qualitative analysis of responses from classroom-, school-, district-, and state-level educators and administrators to open-ended questions about school health education. These questions were posed as part of the School Health Policies and Programs Study (SHPPS), conducted by the Centers for Disease Control and Prevention in 1994, and elicited a range of responses about the status of school health education programs and factors that facilitated and hindered the delivery of such programs. To improve school health education in the United States, respondents cited the need to increase the value and priority of health education in the school curriculum and advocated for 1) professional preparation in health education for persons teaching health-related courses, 2) health education course curricula to address important and timely issues, 3) student testing in health education, 4) improved resources and support for health education, and 5) increased communication and collaboration within their schools and communities related to health education. PMID- 10529964 TI - School-related violence among high school students in the United States, 1993 1995. AB - Using logistic regression to calculate odds ratios and confidence intervals, this secondary analysis determined statistically significant predictors and prevalence rates for high school students in the United States who carried a weapon, were threatened or injured with a weapon, or were involved in physical fighting on school property from 1993-1995. Geographic region, grade in school, race/ethnicity, and gender were the independent variables. Data regarding school related violence among adolescents were collected in 1993 and 1995 using the Youth Risk Behavior Survey developed and implemented by the Centers for Disease Control and Prevention. Results indicated gender, race, and grade in school were predictors of school-related violence for students in grades 9-12 from 1993 to 1995. Males were involved in violent acts more often than females. Minority students, especially Hispanics and Blacks, and students in lower grades at school participate in and are victims of violence more often than other students. PMID- 10529966 TI - Relationship between self-esteem and smoking behavior among Japanese early adolescents: initial results from a three-year study. AB - Researchers examined the relationship between self-esteem and smoking behavior among Japanese elementary and junior high school students. Students (2,090) in fourth to ninth grade from three elementary schools and two junior high schools in the Hyogo and Niigata prefectures completed an anonymous questionnaire. Self esteem was measured using the Harter Perceived Competence Scale, the Pope Self Esteem Scale, and the Rosenberg Self-Esteem Scale. Results indicated that never smokers had higher cognitive, family, and global self-esteem, but lower physical self-esteem than ever smokers. Grade and gender were significantly associated with self-esteem, showing a decrease of self-esteem with increases in grade and a higher level of self-esteem among boys than girls. The results suggest that effective smoking prevention programs for Japanese early adolescents should be integrated into more comprehensive health education or health promotion programs including self-esteem enhancement training. PMID- 10529965 TI - The Florida School Violence Policies and Programs Study. AB - The Florida School Violence Policies and Programs Study (FSVPPS) assessed characteristics of violence prevention and control policies and education programs in Florida's 67 public school districts. Data were collected using the Florida School Violence Policies and Programs Questionnaire (FSVPPQ). Instruments used in the national School Health Policies and Programs Study (SHPPS) served as the framework for item development. A questionnaire designed to solicit information about violence policies and education programs was mailed to all 67 school districts in Florida during spring 1996. Sixty-seven key informants were selected by asking district office personnel to identify the person with primary responsibility for violence prevention activities in their district. Fifty-five school districts returned completed questionnaires, yielding a usable response rate of 82%. Respondents included personnel of superintendent rank, district administrators and directors, instructional specialists, principals, and "Safe & Drug Free Schools" coordinators. Participating school districts were evenly distributed geographically across Florida. The FSVPPS data reported in this paper include administrative and programmatic information about violence prevention education. PMID- 10529967 TI - Physical activity and stages of change in fifth and sixth graders. AB - The Stage of Change (SC) paradigm was adapted to assess readiness to be or stay physically active among fifth and sixth graders. Students completed a four-item SC survey, the Past Year Leisure Time Physical Activity Questionnaire, and the Modifiable Physical Activity Questionnaire for Adolescents. Precontemplation, contemplation, and preparation stages were grouped as "pre-action" (PRE), and action and maintenance as "post-action" (AX) stages. Nearly 40% of all students were in PRE, compared to 60% of students in AX stages. Twenty-two percent of all students were in the sedentary precontemplation or contemplation stages. Significantly more boys were in maintenance than girls, and more girls than boys were in contemplation. Students averaged 14-21 hours/week of television, video, or computer work, and 1.6 hours/week of physical activity outside of school. Interventions may be targeted at a specific SC to enable a child to move forward along the SC continuum toward an active lifestyle. PMID- 10529968 TI - Food industry marketing in elementary schools: implications for school health professionals. PMID- 10529969 TI - Anatomic review and topographic diagnosis. AB - Neuro-ophthalmologic disorders affecting the afferent and efferent visual systems are frequently encountered by neurosurgeons in clinical practice. This article focuses on aspects of neuroanatomy that are most important to the diagnosis of afferent and efferent visual pathway lesions. The use of optic disc and fundus appearance, visual fields, and ocular motility and pupillary findings are emphasized in the discussions of topographic diagnosis. PMID- 10529970 TI - The neuro-ophthalmologic examination. AB - Neurosurgical practice requires a solid foundation in the principles and interpretation of the neuro-ophthalmologic examination. In this article, the techniques used in the neuro-ophthalmologic examination to assess visual acuity, ocular motility, visual fields, the pupils, the eyelids, and the fundus are reviewed. An emphasis is placed on those techniques most relevant to neurosurgical practice. PMID- 10529971 TI - Coma. AB - Comatose patients are unresponsive to all external stimuli, noxious or otherwise. Neuro-ophthalmic techniques are paramount in the neurologic assessment of comatose patients, especially with regard to brain stem localization and diagnosis. Examination of the pupils, fundi, and corneal reflexes are all helpful; however, the ocular motility examination is especially important because the pathways governing ocular motility traverse the entire brain stem, so pathology in this region often produces recognizable eye movement abnormalities. Conversely, if the eye movements are all normal, it is likely that the entire brain stem is normal. This article details the pupillary, eye movement, and funduscopic abnormalities in coma. PMID- 10529972 TI - Neuro-ophthalmic complications of raised intracranial pressure, hydrocephalus, and shunt malfunction. AB - 1. Ophthalmic signs are important for the diagnosis and management of elevated intracranial pressure. 2. Visual loss, visual field loss, dorsal midbrain syndrome, and acute papilledema may occur well in advance of ventricular dilation. 3. For younger patients with hydrocephalus, amblyopia should be checked for, and the absence of papilledema does not ensure normal intracranial pressure. 4. Treatment should be delivered to control intracranial pressure and preserve vision in a timely fashion. PMID- 10529973 TI - Pseudotumor cerebri. AB - Pseudotumor cerebri is an unusual syndrome of increased intracranial pressure without a space-occupying mass. Many associations are known, but the pathogenesis remains a mystery. The diagnosis and treatment of pseudotumor cerebri are often challenging. Because it is not rare, neurosurgeons, neurologists, and ophthalmologists frequently work in concert to manage these patients. This article reviews the diagnostic criteria and differential diagnosis of pseudotumor cerebri. The medical and surgical treatments currently employed in this disorder are discussed. PMID- 10529974 TI - Neuro-ophthalmologic diagnosis and therapy of central nervous system trauma. AB - Careful examination and judicious use of neuroimaging will lead to the initiation of appropriate management for injuries of the afferent and efferent visual systems. This article concentrates on those aspects of neuro-ophthalmologic trauma that are most relevant to the neurosurgeon, particularly those disorders that are commonly faced when caring for acutely injured patients. Although direct injury to the orbit and globes can easily result in neuro-ophthalmologic disorders, it is less appreciated that distant head injury can similarly result in injury to the retrobulbar afferent visual pathways or cranial nerves. It is therefore necessary that the neurosurgeon be suspicious of both latent and manifest injuries to the neuro-ophthalmologic system after head trauma, and maintain awareness of the need for timely intervention in a subset of these disorders. Therefore, even patients with injuries removed from the eyes should be checked for visual problems, and, if any are detected, an ophthalmologist or neuro-ophthalmologist should be consulted. PMID- 10529975 TI - Aneurysms and subarachnoid hemorrhage. AB - Most unruptured intracranial aneurysms that produce neuro-ophthalmologic signs arise from the junction of the internal carotid and posterior communicating arteries. These aneurysms typically compress the third nerve in the subarachnoid space. Compression of cranial nerves within the cavernous sinus is less common, resulting in single or multiple and often painful ocular motor nerve pareses. Unruptured aneurysms of the proximal and distal segments of the intracranial portion of the internal carotid artery can compress the anterior visual pathways and cause visual loss. Ocular symptoms and signs may be the presenting manifestations of intracranial aneurysms. Prompt recognition of an aneurysm prior to rupture can prevent devastating intracerebral or subarachnoid hemorrhage. Moreover, visual complications are a not infrequent source of morbidity in those patients surviving acute intracranial bleeding. Recent advances in noninvasive neuroimaging and endovascular therapies facilitate early diagnosis and treatment and therefore may limit such complications. PMID- 10529976 TI - Carotid-cavernous fistulas. AB - Carotid-cavernous fistulas often cause ophthalmologic manifestations that may result in loss of vision. Knowledge of these ocular signs is essential for prompt diagnosis and treatment. Appropriate management frequently involves multiple specialists including neuro-ophthalmologists, interventional radiologists, and neurosurgeons. This article reviews current concepts essential to all specialists who treat patients with carotid-cavernous fistulas. PMID- 10529977 TI - Neuro-ophthalmology of arteriovenous malformations. AB - Intracranial arteriovenous malformations may result in a number of neuro ophthalmologic signs and symptoms. In some patients, the neuro-ophthalmic findings are the presenting or only manifestations. A multi-disciplinary team approach including neurologists, ophthalmologists, neurosurgeons, interventional radiologists, and radiation therapists is ideal. PMID- 10529978 TI - Gliomas of the anterior visual pathways. AB - Gliomas of the anterior visual pathways include the more commonly encountered benign tumors of children and the rare and highly aggressive malignant tumors of adults. Diagnostic biopsy is not required in most cases of childhood gliomas, especially those unassociated with visual loss or other neurologic symptoms, because the clinical and neuroimaging features are often sufficiently characteristic. Because of the infiltrative nature of these tumors, the neurosurgeon has little to offer symptomatic patients beyond debulking large exophytic masses and shunting if hydrocephalus is present. PMID- 10529979 TI - Optic nerve and orbital tumors. AB - The close proximity of the orbit to the brain and the high frequency of ocular symptoms in patients with neurosurgical disease make it mandatory for neurosurgeons to be familiar with manifestations of orbital disease. Tumors of the optic nerve and orbit are important causes of vision loss and eye movement abnormalities. Similarly, intracranial tumors frequently present with eye movement abnormalities, vision loss, and optic nerve swelling. This overlap in the clinical characteristics of patients with orbital tumors and patients with neurosurgical problems makes familiarity with the types of clinical presentation of various orbital tumors important to the neurosurgeon. The history and examination of patients with orbital tumors are discussed and the clinical presentation of various orbital tumors is presented. PMID- 10529980 TI - Neuro-ophthalmic manifestations of pituitary tumors. AB - Pituitary tumors manifest with a variety of unique localizing signs and symptoms that neurosurgeons and neuro-ophthalmologists must readily identify. Visual disturbances may be the presenting complaint. Loss of central visual acuity, visual field deficits, and mechanisms of diplopia are discussed with case examples used to highlight each. PMID- 10529981 TI - The cavernous sinus. AB - The parasellar region, in particular the cavernous sinus, represents the confluence of critical structures involved in both the afferent and efferent visual pathways. It is not surprising that lesions affecting the area of the cavernous sinus most frequently present with neuro-ophthalmic complaints, which include double vision, decreased vision, pain, and numbness. Neuro-surgical intervention in the area of hte cavernous sinus often produces neuro-ophthalmic complications. A rehabilitative effort directed to the neuro-ophthalmic manifestations is essential for patients with skull-base pathology. PMID- 10529982 TI - Other intracranial mass lesions. AB - This article outlines the various signs and symptoms associated with intracranial gliomas, meningiomas, and hypertensive hemorrhages. It emphasizes the neurologic and neuro-ophthalmologic presentations of these intracranial mass lesions. Important pathways that subserve horizontal eye movements are examined and the ocular motility disorders associated with lesions of these fibers are outlined. PMID- 10529983 TI - The phacomatoses. AB - The phacomatoses are disorders characterized by multiple hamartomas of the central and peripheral nervous system, eye, skin, and viscera. Many of these diseases have well-defined Mendelian patterns of inheritance because of a mutation of a single gene. In other instances, no clear patterns of inheritance or genetic susceptibility have been identified. In some cases, patients are at increased risk of malignancy. The combination of ocular and CNS manifestations seen in patients with the phacomatoses makes neuro-ophthalmologic evaluation particularly important in the diagnosis and management of these patients. PMID- 10529984 TI - Applications of genetic algorithms on the structure-activity relationship analysis of some cinnamamides. AB - Quantitative structure-activity relationships (QSARs) for 35 cinnamamides were studied. By using a genetic algorithm (GA), a group of multiple regression models with high fitness scores was generated. From the statistical analyses of the descriptors used in the evolution procedure, the principal features affecting the anticonvulsant activity were found. The significant descriptors include the partition coefficient, the molar refraction, the Hammet sigma constant of the substituents on the benzene ring, and the formation energy of the molecules. It could be found that the steric complementarity and the hydrophobic interaction between the inhibitors and the receptor were very important to the biological activity, while the contribution of the electronic effect was not so obvious. Moreover, by construction of the spline models for these four principal descriptors, the effective range for each descriptor was identified. PMID- 10529985 TI - Comparative spectra analysis (CoSA): spectra as three-dimensional molecular descriptors for the prediction of biological activities. AB - A novel 3D QSAR approach, comparative spectra analysis (CoSA), in which molecular spectra are used as three-dimensional molecular descriptors for the prediction of biological activities, is presented and discussed. To this purpose, experimentally determined 1H NMR, mass, and IR spectra, as well as simulated IR and 13C NMR spectra, for a set of 45 diverse progestagens are converted by a program, SpecMat, into matrixes, which are subsequently employed in a multivariate regression analysis (PLS). The results are compared with those resulting from a comparative molecular field analysis (CoMFA). When used individually, spectral descriptors yield better correlations and predictions than molecular field descriptors. A combination of spectral descriptors with other descriptors, either spectral or molecular field in nature, leads in most cases to models that are statistically superior to the ones obtained by their corresponding individual spectral or molecular field descriptors. PMID- 10529986 TI - Database searching for compounds with similar biological activity using short binary bit string representations of molecules. AB - In an effort to identify biologically active molecules in compound databases, we have investigated similarity searching using short binary bit strings with a maximum of 54 bit positions. These "minifingerprints" (MFPs) were designed to account for the presence or absence of structural fragments and/or aromatic character, flexibility, and hydrogen-bonding capacity of molecules. MFP design was based on an analysis of distributions of molecular descriptors and structural fragments in two large compound collections. The performance of different MFPs and a reference fingerprint was tested by systematic "one-against-all" similarity searches of molecules in a database containing 364 compounds with different biological activities. For each fingerprint, the most effective similarity cutoff value was determined. An MFP accounting for only 32 structural fragments showed less than 2% false positive similarity matches and correctly assigned on average approximately 40% of the compounds with the same biological activity to a query molecule. Inclusion of three numerical two-dimensional (2D) molecular descriptors increased the performance by 15%. This MFP performed better than a complex 2D fingerprint. At a similarity cutoff value of 0.85, the 2D fingerprint totally eliminated false positives but recognized less than 10% of the compounds within the same activity class. PMID- 10529987 TI - Automated pharmacophore identification for large chemical data sets. AB - The identification of three-dimensional pharmacophores from large, heterogeneous data sets is still an unsolved problem. We developed a novel program, SCAMPI (statistical classification of activities of molecules for pharmacophore identification), for this purpose by combining a fast conformation search with recursive partitioning, a data-mining technique, which can easily handle large data sets. The pharmacophore identification process is designed to run recursively, and the conformation spaces are resampled under the constraints of the evolving pharmacophore model. This program is capable of deriving pharmacophores from a data set of 1000-2000 compounds, with thousands of conformations generated for each compound and in less than 1 day of computational time. For two test data sets, the identified pharmacophores are consistent with the known results from the literature. PMID- 10529988 TI - Strategic pooling of compounds for high-throughput screening. AB - Bringing new medicines to the market depends on the rapid discovery of new and effective drugs, often initiated through the biological testing of many thousands of compounds in high-throughput screening (HTS). Mixing compounds together into pools for screening is one way to accelerate this process and reduce costs. This paper contains both theoretical and experimental data which suggest that careful selection of compounds to be pooled together is necessary in order to reduce the risk of reactivity between compounds within the pools. PMID- 10529989 TI - CONCAWE report. PMID- 10529990 TI - Evaluation of activated carbon filters for removal of ozone at the PPB level. AB - Performance of filters for the removal of ozone at ambient concentration is characterized. The removal efficiency and pressure drop of 10 commercial filters- including 8 made of granule or powdered activated carbon, 1 activated carbon fiber filter, and 1 packed bed made of an ozone catalyst--were measured for an influent ozone concentration of 120 ppb at 50% relative humidity and 2.54 m/sec face velocity. Activated carbon filters can be very effective at ozone removal, although not indefinitely because chemical reactions of ozone and carbon change the carbon. Initial efficiencies of the 1.27-cm thick flat samples varied from 4.6 to 98.3%. Analysis of the structure and composition of the filters with scanning electron microscopy and X-ray photoemission spectrometry showed chemical reactions permanently changed the composition of the carbon and decreased the surface area. Consequently, removal efficiency decreased with use. Moreover, it was not feasible to regenerate the filters by simply removing them from ozone laden air. Changes in relative humidity, from 20 to 80%, had no measurable effect on the performance of a granule activated carbon filter. However, because the rate of adsorption of water is faster and the pores are smaller in activated carbon fiber, efficiency of the fiber filter decreased when relative humidity was raised from 20 to 50%. A quality factor, equal to the ratio of a threshold breakthrough time and pressure drop, is used to compare filters. In general, those with higher carbon surface area per unit volume had higher efficiencies and greater pressure drops. Future work should address the removal of ozone in the presence of other gases. PMID- 10529991 TI - Simulated workplace performance of N95 respirators. AB - During July 1995 the National Institute for Occupational Safety and Health (NIOSH) began to certify nine new classes of particulate respirators. To determine the level of performance of these respirators, NIOSH researchers conducted a study to (1) measure the simulated workplace performance of 21 N95 respirator models, (2) determine whether fit-testing affected the performance, and (3) investigate the effect of varying fit-test pass/fail criteria on respirator performance. The performance of each respirator model was measured by conducting 100 total penetration tests. The performance of each respirator model was then estimated by determining the 95th percentile of the total penetration through the respirator (i.e., 95% of wearers of that respirator can expect to have a total penetration value below the 95th percentile penetration value). The 95th percentile of total penetrations for each respirator without fit-testing ranged from 6 to 88%. The 95th percentile of total penetrations for all the respirators combined was 33%, which exceeds the amount of total penetration (10%) normally expected of a half-mask respirator. When a surrogate fit test (1% criterion) was applied to the data, the 95th percentile of total penetrations for each respirator decreased to 1 to 16%. The 95th percentile of total penetrations for all the respirators combined was only 4%. Therefore, fit-testing of N95 respirators is necessary to ensure that the user receives the expected level of protection. The study also found that respirator performance was dependent on the value of the pass/fail criterion used in the surrogate fit-test. PMID- 10529993 TI - A protocol system for testing biohazardous materials in an impact biomechanics research facility. AB - This article presents a protocol system, comprised of a review process and a series of checklists, that was developed for testing cadaveric tissue in an impact biomechanics research facility. The use of cadaveric tissue may expose personnel to bloodborne pathogens including HIV and hepatitis B, which have been shown to remain virulent in a cadaver for several weeks after death. To minimize exposure risks, the protocol system presented emphasizes initial blood screening to keep infectious tissue from entering the laboratory, and adopts universal precautions to prevent exposure by treating all tissue as though it were infected. All lab employees must read, sign, and demonstrate proficiency in the protocol. Well-developed test procedures for the handling of biohazardous materials along with an annual individual protocol review have proven effective for the past 6 years in minimizing exposure risks. PMID- 10529992 TI - Evaluation of exposure to methyl methacrylate among dental laboratory technicians. AB - Following the diagnosis of two cases of occupational asthma among dental technicians, an industrial hygiene survey was conducted in two dental laboratories to determine time-weighted average and peak concentrations of methyl methacrylate vapor and time-weighted average concentration of acrylic dust. The time-weighted average concentrations of methyl methacrylate vapor were 0.7 ppm and 1.6 ppm and average peak concentrations were 9.3 ppm and 9.7 ppm for the first and second laboratory, respectively. The use of a local exhaust ventilation system was significant in reducing the peak concentration of methyl methacrylate vapor in the breathing zone of dental technicians. However, the local exhaust ventilation was not efficient in reducing the concentration of airborne acrylic dusts. Occupational exposure of dental technicians to dental materials, in particular to methyl methacrylate, requires further investigation. Local exhaust ventilation systems can reduce the concentration of methyl methacrylate in the dental laboratories to a significant extent if installed and used properly. PMID- 10529994 TI - Employee exposure to diesel exhaust in the electric utility industry. AB - The purpose of this study was to assess diesel exhaust exposures in the electric utility industry and to compare these findings with worker exposures reported in other industries and to proposed and established occupational exposure limits. Two sampling approaches were used: particulates were analyzed for elemental (EC) and organic (OC) carbon via the thermal-optical method; and gaseous components (NO2, SO2, NO, and CO) were determined using a direct reading instrument, the Metrosonics pm-7400. Concentrations of the gases were all well within established occupational exposure levels. The EC percentage of the total carbon was generally lower than results reported from other studies resulting in OC levels representing a higher percentage of the total carbon concentrations. Smokers had higher average OC exposure (79 micrograms/m3) than nonsmokers (57 micrograms/m3), but cigarette smoke did not contribute to EC levels in this study (smokers and nonsmokers = 3 micrograms/m3). Two of 120 individual personal exposure levels were found to exceed the proposed threshold limit value of 150 micrograms/m3 for total particulate, but geometric mean levels were found to be significantly less than the proposed value. Questions are raised concerning the use of EC as the sole surrogate for estimating diesel content for comparison with an exposure standard. PMID- 10529995 TI - Dust exposures in the wood processing industry. AB - Workers at four different woodworking processes--two logging sites, four sawmills, one major woodchipping operation, and five joineries situated in the state of New South Wales in Australia--were studied for personal inhalable dust exposures (N = 182). The geometric mean exposure at logging sites was 0.6 mg/m3 (N = 7), sawmills 1.6 mg/m3 (N = 93), woodchipping 1.9 mg/m3 (N = 9), and joineries 3.7 mg/m3 (N = 66). Overall, 62% of the exposures exceeded the current standards. Among joineries, 95% of the hardwood exposures and 35% of the softwood exposures were above the relevant standards. A majority of workers (approximately 90%) did not wear appropriate respirators approved for wood dust, while the ones who did wear them, used them on average less than 50% of the time. The significant determinants of personal wood dust exposures (n = 163) were found to be local exhaust ventilation, job title, use of handheld tools, cleaning method used, use of compressed air, and green or dry wood processed. Type of wood processed (softwood or hardwood) was not found to be statistically significant. PMID- 10529996 TI - Comparison of personal exposure meter placement for the determination of office worker ELF magnetic field exposures. AB - This article compares extremely low frequency (ELF) magnetic field exposures measured by placing EMDEX Lite personal exposure meters (PEMs) at the head, chest, and waist level for a group of office workers. Twenty-three volunteers were solicited to wear three PEMs simultaneously; one was attached to a baseball cap worn on the head, one was attached to a band and worn around the neck (positioned on the chest), and one was worn in a belted pouch around the waist (positioned on the right side of the hip). The effect of PEM placement was evaluated by comparing full-shift average exposures and daily maximum or peak exposure. The results of this investigation indicate that time-weighted average magnetic field exposures determined at the hip provide the highest mean exposure estimates. Averages of the full-shift mean magnetic field measurements taken at hip and head levels were statistically greater than measurements taken at the chest level by 33 and 22%, respectively. Comparisons of the maximum or peak magnetic field exposures by body position indicate that the hip position produced an average exposure estimate that was 136% greater than the average head-level measurement. Results suggest that for office workers PEM meter placement on the body does not produce large differences in full-shift average ELF magnetic flux density exposures. However, the hip position produced the largest daily maximum or peak exposures. It is recommended that PEMs be placed on the hip for exposure assessments in office environments, because this placement is the most commonly used, the most convenient, and resulted in the highest magnetic field exposures. PMID- 10529997 TI - Issues concerning the measurement of borate in occupational environments. AB - Borates are susceptible to weight change due to uptake or loss of water and this hydration instability can lead to gravimetric and interpretation errors in occupational hygiene field sampling of dust. The hydration stability for inhalable borate dust particles (mean diameter 7-22 microns) was characterized over a range of ambient temperature and relative humidity conditions simulating field sampling. Borax 10 mol (Na2O.2B2O3.10H2O), a fully hydrated borate, has a relatively high vapor pressure to water that led to rapid dehydration with significant weight change. Low hydrate borates, Neobor borax 5 mol (Na2O.2B2O3.5H2O), anhydrous boric acid (B2O3), and anhydrous borax (Na2O.2B2O3) were found to hydrate rapidly with an increase in weight. In contrast, boric acid (B[OH]3) and borax 5 mol were found to be stable to dehydration under all conditions. Boron can be measured with high analytical accuracy, but because the specific borate species or borate compounds collected in a 37-mm dust sampler cannot be accurately identified, it is argued that occupational exposure values should be revised to reflect exposure to boron and exposure values for these borates should be the same based on equivalent boron content. PMID- 10529998 TI - Heat stress and strain in an aluminum smelter. AB - Studies of worker heat stress and strain in aluminum smelters have found that heat exposure likely to exceed the American Conference of Governmental Industrial Hygienists' threshold limit value (TLV) and that the dose-response relationship between heat stress and strain was weak. A heat stress model based on climatic data and a task analysis indicated exposures to heat stress in excess of the TLV during the July/August study period. To study the impact of working above the TLV, heat strain data (i.e., oral temperature, recovery heart rate, average heart rate) were collected. Recovery heart rates indicated high strain most of the time, and oral temperatures after peak demands were above the no-strain threshold of 37.5 degrees C about a quarter of the time, indicating that heat stress had an effect. About 95% of the readings were below 38.0 degrees C, the acute oral temperature threshold for a safe exposure. Average heart rates over 6- and 12 hour intervals were generally below acceptable limits of 120 and 110 bpm, respectively. Oral temperature and average heart rates indicated good control of heat stress exposures. Because recovery heart rates were high, some employees were working near their individual limits. The dose-response relationship for recovery heart rate and oral temperature were examined against the level of heat stress above the TLV. There was no relationship between oral temperature and heat stress level. There was an apparent trend toward higher recovery heart rates with heat stress. The lack of a dose-response relationship may be explained by brief periods of very high wet bulb globe temperatures that drove the time-weighted average up out of proportion to the physiological response. PMID- 10529999 TI - Incident trends for a hazardous waste cleanup company. AB - Published reports to assess incidents in hazardous waste operations are scarce. This study was designed to evaluate incident trends in a relatively large hazardous waste cleanup company. The data for 6.5 years, winter 1990 through spring 1996, provided 1848 incident reports with 87% involving injury/illness cases. Over 75% of injury/illness incidents were due to mechanical agents, 10% occurred because of chemical exposure, 5% involved poisonous plants and insect bites, 2% resulted from temperature extremes, 1% were from cumulative injuries/illnesses, and in 7% the agent was not recorded. Almost 31% of injuries were related to the upper extremities, with the fingers most often injured, followed by the hands. Lower back strain cases constituted 11% of injuries, ankle/foot/toe cases 9%, and knee cases 5%. Recovery technicians (laborers) had the highest frequency of injury/illness incidents (52%), followed by supervisors (15%) and heavy machinery operators (10%). The incidence rates (IRs) for all recordable incidents ranged from 11.9 for the second quarter of 1990 down to 1.2 for the fourth quarter of 1995 with a mean (SD) and median of 6.3 (3.0) and 6.1, respectively. For the time period studied, IRs decreased significantly (p < 0.01). It was concluded that hands-on experience in the field and improvements in the health and safety program of the company--including expanding its focus (originally the prevention of chemical exposure) to include construction safety- reduced the incidents considerably. Introduction of new regulations has also contributed to this trend. PMID- 10530000 TI - Differences in the association between psychosocial work conditions and physical work load in female- and male-dominated occupations. MUSIC-Norrtalje Study Group. AB - This study investigated whether there is a relationship between high physical work load and adverse psychosocial work factors, and whether this relationship is different for women and men. Separate analyses for female registered nurses and assistant nurses were made because these are common occupations involving high physical and psychological demands. This study was part of the MUSIC-Norrtalje study, a population study with the overall aim of identifying risk factors for musculoskeletal disorders. The respondents, 1423 gainfully employed men and women, were randomly selected from the study population. The exposure assessments referred to a typical workday during the previous 12 months. Physical exposure was investigated by interview, psychosocial work factors by interview and questionnaire. For the women, but not the men, mainly routine work and a job strain situation, according to the model of Karasek and Theorell, increased the probability of having a high physical work load, assessed as a time-weighted average of energy expenditure in multiples of the resting metabolic rate. Results indicated that in female-dominated occupations, high physical work load might also imply adverse psychosocial conditions. A higher frequency of high physical work load and job strain was observed among assistant nurses compared with registered nurses. Covariance between physical and psychosocial demands makes it difficult to determine the relative influence of each in health problems. Results of the present study imply that this is a larger problem in studies of women than men. PMID- 10530001 TI - Worker exposures to particulates, endotoxins, and bioaerosols in two refuse derived fuel plants. AB - Exposures of refuse-derived fuel (RDF) production workers to total particulates, endotoxin, and total (viable plus nonviable) bioaerosols were characterized at two RDF production plants. Full-shift personal air monitoring for 35 workers was conducted for total particulates, analyzed by gravimetric analysis; endotoxin, analyzed by chromogenic endpoint assay; and total bioaerosols, analyzed by fluorescent microscopy (FM). Geometric mean values of personal air samples were 0.50 mg/m3 for total dust, 29.0 EU/m3 (2.9 ng/m3) for endotoxin, and 6.8 x 10(5) organisms/m3 for bioaerosols. Significant differences were observed between the two plants only for total endotoxin exposures. The mean concentrations for total particulates, total FM bioaerosols, and endotoxins did not differ among the day, evening, or night shifts. Interjob differences were found for exposures to total dust, total endotoxin, and FM bioaerosols. Individual comparisons for total particulates and endotoxin exposures were significant for comparisons between job categories as a result of the greater exposures for personnel cleaning the plants. Significant correlations were detected between total particulates and total endotoxin measurements and between inhalable and total particulates. PMID- 10530002 TI - Effect of the tetrazole cis-amide bond surrogate on the complexing ability of some enkephalin analogues toward Cu(II) ions. AB - A study of the effect of the tetrazole moiety, a cis-amide bond surrogate, on the Cu(II) coordinating properties of oligopeptides is reported. The insertion of the tetrazole moiety psi (CN4) into the peptide sequence of [Leu5]enkephalin considerably changes the coordination ability of the ligand. Potentiometric and spectroscopic results indicate that if the tetrazole moiety is in a suitable position in the peptide chain, i.e. if it follows the third residue, an unusual stable CuH-1L species involving 4N coordination is formed in the physiological pH region. The tetrazole psi (CN4) ring provides one of these nitrogens. The data indicate that Cu(II) ions are strongly trapped inside a bent peptide backbone. However, the coordination mode involving the tetrazole ring nitrogen does not prevent the hydrolysis process under strongly basic conditions. PMID- 10530003 TI - Copper(II) complexes of a nonsteroidal anti-inflammatory drug niflumic acid. Synthesis, crystal structure of tetrakis-mu-(2-[3-(trifluoromethyl) phenyl]aminonicotinato)bis(dimethylsulfoxide)-dicopper(II) complex at 190 K. Anti inflammatory properties. AB - The synthesis and characterization of three complexes with a potent nonsteroidal anti-inflammatory drug niflumic acid {2-[3-(trifluoromethyl)phenyl]aminonicotinic acid} with formula [Cu(niflumato)2L] (L = H2O, DMSO = dimethylsulfoxide, DMF = N,N-dimethylformamide) were investigated. The crystal and molecular structure of the {Cu(niflumato)2(DMSO)}2 was reported. Crystallographic data are as follows: monoclinic system, space group P2(1)/n, Z = 2, a = 11.1318(8), b = 17.513(2), c = 15.336(1) A, beta = 103.316(8) degrees, V = 2909.4(4) A3. The structure was refined to R = 0.030 and wR = 0.037 for 3702 reflections with I > sigma (I). It consists of centrosymmetric binuclear units with the Cu-Cui (symmetry code i: 1 x, -y, 1-z) distance between two centrosymmetrically related ions of 2.6272(5) A. Each Cu(II) ion in [Cu2(DMSO)2(mu-niflumato)4] is coordinated to an apical dimethylsulfoxide O atom on the one hand and to the equatorial carbonyl and carboxylic O atoms of two crystallographically independent niflumate moieties and their centrosymmetric counterparts on the other hand. In spite of the low temperature (190 K) crystal measurements, one L-CF3 grouping exhibits some disorder. The biological activities of these complexes were compared to that of niflumic acid. Niflumic acid and its various copper complexes significantly inhibited polymorphonuclear leukocyte (PMNL) oxidative metabolism, as assessed by chemiluminescence and O2- generation measurement. This effect was dose-dependent. All copper complexes exerted a similar inhibiting effect which was always significantly higher than that exerted by the parent drug. PMID- 10530005 TI - Generation and biomimetic chemistry of tungsten-dithiolene complexes containing the hydrotris(3,5-dimethylpyrazol-1-yl)borate ligand. AB - Reactions of bis(thio)-W(VI) complexes, LWS2X (L = hydrotris (3,5-dimethylpyrazol 1-yl)borate, X = monoanion), with alkynes produce dithiolene complexes, LWX(dithiolene). The synthesis and characterization of orange LW(OPh){S2C2(CO2Me)2} (1) and burgundyred LW(SePh) {S2C2(Ph)(2-quinoxalinyl)} (2) and the X-ray crystal structure of 1.0.5CH2Cl2, are described in detail. Crystals of 1.0.5CH2Cl2 are orthorhombic, space group Pbcn, with a = 29.826(6), b = 13.291(4), c = 16.078(4) A, V = 6373(5) A3, and Z = 8. The six-coordinate, distorted-octahedral complex features a tridentate L ligand, a monodentate phenoxide ligand, and a bidentate dithiolene ligand. The short W-S bonds (2.267(4) and 2.279(4) A) and the parameters associated with the phenoxide ligand (W-O = 1.850(8) A, W-O-C = 146(1) degree), point to a considerable degree of W ligand multiple bonding in the [W(OPh)(dithiolene)]+ unit. For 2, W-Se and average W-S distances of 2.49(2) A and 2.30(2) A, respectively, have been obtained from EXAFS studies. The formation of yellow 3,3'-dithiobis[2 (phenyl)thieno[2,3-b]quinoxaline] (3) upon oxidation of 2 supports the likely generation of urothione upon oxidative degradation of molybdopterin-containing tungsten enzymes from hyperthermophilic organisms. PMID- 10530004 TI - Synthesis and characterization of novel palladium(II) complexes of bis(thiosemicarbazone). Structure, cytotoxic activity and DNA binding of Pd(II) benzyl bis(thiosemicarbazonate). AB - The preparation of palladium(II) complexes of 3,5-diacyl-1,2,4-triazole bis(thiosemicarbazone) (H2L2), 2,6-diacylpyridine bis(thiosemicarbazone) (H2L3) and benzyl bis(thiosemicarbazone) (H2L4) is described. The new complexes [PdCl2(H2L2)] (1), [PdCl2(H2L3)] (2) and [PdL4].DMF (3) have been characterized by elemental analyses and spectroscopic studies (IR, 1H NMR and UV-Vis). The crystal and molecular structure of PdL4.DMF (L = bideprotonated form of benzyl bis(thiosemicarbazone)) has been determined by single-crystal X-ray diffraction: green triclinic crystal, a = 10.258(5), b = 10.595(5), c = 11.189(5) A, alpha = 97.820(5), beta = 108.140(5), gamma = 105.283(5) degrees, space group P1, Z = 1. The palladium atom is tetracoordinated by four donor atoms (SNNS) from L4 to form a planar tricyclic ligating system. The testing of the cytotoxic activity of compound 3 against several human, monkey and murine cell lines sensitive (HeLa, Vero and Pam 212) and resistant to cis-DDP (Pam-ras) suggests that compound 3 might be endowed with important antitumor properties since it shows IC50 values in a microM range similar to those of cis-DDP [cis-diamminedichloroplatinum(II)]. Moreover, compound 3 displays notable cytotoxic activity in Pam-ras cells resistant to cis-DDP (IC50 values of 78 microM versus 156 microM, respectively). On the other hand, the analysis of the interaction of this novel Pd thiosemicarbazone compound with DNA secondary structure by means of circular dichroism spectroscopy indicates that it induces on the double helix conformational changes different from those induced by cis-DDP. PMID- 10530006 TI - Synthesis, characterization and the effect of ligand planarity of [Ru(bpy)2L]2+ on DNA binding affinity. AB - Two structurally related ligands (L) 4,5,9,18-tetraazaphenanthreno[9,10-b] triphenylene (taptp) and 2,3-diphenyl-1,4,8,9-tetraazatriphenylene (dptatp), and their related complexes of [Ru(bpy)2L]2+ have been synthesized and characterized by elemental analyses, 1H NMR and mass spectra. Their electrochemical properties were also examined. Both complexes emit intense luminescence in organic solvent but are quenched in water to different extents. The interactions of the complexes with calf thymus DNA have been investigated by viscosity, absorption, emission and circular dichroism spectra. The intrinsic binding constants of [Ru(bpy)2(taptp)]2+ and [Ru(bpy)2(dptatp)]2+ are 1.7 x 10(5) and 3.8 x 10(4) M-1, respectively. All data indicate that both complexes bind enantioselectively to double-stranded calf thymus DNA via the intercalative mode, with stronger affinity for the fully planar ligand complex of [Ru(bpy)2(taptp)]2+. PMID- 10530007 TI - Copper(II) complexation by pituitary adenylate cyclase activating polypeptide fragments. AB - Results are reported from potentiometric and spectroscopic (UV-Vis, CD, and ESR) studies of the protonation constants and Cu2+ complex stability constants of pituitary adenylate cyclase activating polypeptide fragments (HSDGI-NH2, TDSYS NH2, RKQMAVKKYLAAVL-NH2). With HSDGI-NH2, the formation of a dimeric complex Cu2H 2L2 was found in the pH range 5-8, in which the coordination of copper(II) is glycylglycine-like, while the fourth coordination site is occupied by the imidazole N3 nitrogen atom, forming a bridge between two copper(II) ions. The formation of dimeric species does not prevent the deprotonation and coordination of the amide nitrogen, and in pH above 8 the CuH-2L complex is formed. Aspartic acid in the third position of peptide sequence stabilizes the CuH-2L species and prevents the coordination of a fourth nitrogen donor. Aspartic acid residue in the second position of TDSYS-NH2 stabilizes the CuL (2N) complex but does not prevent deprotonation and binding of the second and third peptide nitrogens to give 3N and 4N complexes at higher pH. The tetradecapeptide amide forms with copper(II) ions unusually stable 3N and 4N complexes compared to pentaalanine amide. PMID- 10530008 TI - ESR characterization and metallokinetic analysis of Cr(V) in the blood of rats given carcinogen chromate(VI) compounds. AB - It has been shown that bio-trace metal elements are related to many diseases and the aging process. For many years, carcinogen hexavalent chromium (VI) has been known to be toxic to animals, but its dynamic toxicological mechanism is not sufficiently elucidated. Bioinorganic chemistry in terms of metallokinetic analysis of beneficial or toxic metal ions and their complexes is an important investigation for understanding their biochemical and physiological roles. We have tried to examine the real-time behavior of paramagnetic metal ions and complexes in animals, in which electron spin resonance (ESR) was capable of measuring paramagnetic species in chemical and biological systems. On the basis of our previous results on stable nitroxide spin probes, we have developed the in vivo blood circulation monitoring-electron spin resonance (BCM-ESR) method to analyze time-dependent ESR signal changes due to paramagnetic metal ions and their complexes in real time. When K2Cr2O7 or Na2Cr2O7 in saline was intravenously administered to rats, two ESR signals due to pentavalent chromium(V) were detectable in the circulating blood of rats. Cr(V) detected in the blood was indicated to be in the CrO(O4) and CrO(S2O2) coordination modes after the study on model complexes. From the changes of ESR signal intensities due to Cr(V) in the blood, the metallokinetic parameters were obtained using the pharmacokinetic analysis and the curve-fitting methods. The obtained results are important for understanding carcinogen chromate in terms of the formation of Cr(V) in animals. In addition, we propose the BCM-ESR method, which is useful to analyze the disposition of paramagnetic metal species in the blood of living animals. PMID- 10530010 TI - On the combined effects of signal-to-noise ratio and room acoustics on speech intelligibility. AB - Speech intelligibility in rooms is influenced by room acoustics effects and by the signal-to-noise ratio (S/N) of the speech and ambient noise. Several measures such as useful-to-detrimental sound ratios and the speech transmission index predict the combined effects of both types of factors. These measures were evaluated relative to speech intelligibility test results obtained in simulated sound fields. The use of simulated sound fields made it possible to create the full range of combinations of room acoustics and S/N effects likely to be found in rooms for speech. The S/N aspect is shown to be much more important than room acoustics effects and new broadband useful-to-detrimental ratios were validated. Useful-to-detrimental ratios, speech transmission index measures, and values of the articulation loss for consonants were all reasonably accurate predictors of speech intelligibility. Further improvements to these combined measures are suggested. PMID- 10530009 TI - The precedence effect. AB - In a reverberant environment, sounds reach the ears through several paths. Although the direct sound is followed by multiple reflections, which would be audible in isolation, the first-arriving wavefront dominates many aspects of perception. The "precedence effect" refers to a group of phenomena that are thought to be involved in resolving competition for perception and localization between a direct sound and a reflection. This article is divided into five major sections. First, it begins with a review of recent work on psychoacoustics, which divides the phenomena into measurements of fusion, localization dominance, and discrimination suppression. Second, buildup of precedence and breakdown of precedence are discussed. Third measurements in several animal species, developmental changes in humans, and animal studies are described. Fourth, recent physiological measurements that might be helpful in providing a fuller understanding of precedence effects are reviewed. Fifth, a number of psychophysical models are described which illustrate fundamentally different approaches and have distinct advantages and disadvantages. The purpose of this review is to provide a framework within which to describe the effects of precedence and to help in the integration of data from both psychophysical and physiological experiments. It is probably only through the combined efforts of these fields that a full theory of precedence will evolve and useful models will be developed. PMID- 10530011 TI - A composite model of the auditory periphery for simulating responses to complex sounds. AB - This paper presents a phenomenological model of the cochlea. It consists of a bank of nonlinear time-varying parallel filters and an active distributed feedback. Realistic filter shapes are obtained with the all-pole gamma-tone filter (APGF), which provides both a good approximation of the far more complex wave propagation or cochlear mechanics models and a very simple implementation. Special care has been taken in modeling nonlinear properties in order to mimic the responses of the cochlea to complex stimuli. As a result, the model reproduces several observed phenomena including compression, two-tone suppression, and suppression of tones by noise. The distributed feedback, based on physiological evidence from outer hair cell (OHC) functioning, controls the damping parameter of the APGF and provides good modeling of both low-side and high-side suppression. Responses to more complex stimuli as well as a study of the model's parameters are also presented. Areas of application of this type of model include understanding of signal coding in the cochlea and auditory nerve, development of hearing aids, speech analysis, as well as input to neural models of higher auditory centers. PMID- 10530012 TI - The representation of pure tones and noise in a model of cochlear nucleus neurons. AB - The limited dynamic range of the majority of auditory-nerve fibers represents a difficulty in accounting for normal hearing capabilities over the known psychoacoustic intensity range. The presence of noise is an additional complication because it will tend to saturate these fibers, thereby considerably reducing their dynamic range, i.e., the range of mean firing rates. In this study, simulations involving a model of auditory nerve and cochlear nucleus neurons were conducted using pure-tone stimuli in the presence of noise. The main focus is on the role of inhibition in regulating the activity of cells, improving their capability to represent signals in background noise. This concerns in particular those inhibitory neurons that receive input from a wide range of auditory-nerve fibers and respond with an onset chopper pattern. A detailed model of stellate cells is used. It allows several parameters such as the number, location, and strength of inputs to be manipulated. The fist part of this paper presents the model and its responses to pure-tone and noise stimuli presented separately. The model's capacity to generalize to tone/noise combinations is then tested. Responses to these stimuli are found to be qualitatively similar to neurophysiological findings. Model neurons exhibit appropriate shifts in their rate-level functions and their responses are inhibited or suppressed by tones outside their characteristic frequency. The model stellate cell is also found to display many of the temporal patterns reported in electrophysiological studies as a result of appropriate settings of certain parameters. Therefore, the model is sufficient to account for a larger number of findings and should serve as a basis for predicting responses to novel stimuli, or as a building block for modeling larger networks. PMID- 10530013 TI - Solution of the inverse problem for a linear cochlear model: a tonotopic cochlear amplifier. AB - The extraordinary fine-tuning characteristic of normal mammalian hearing is attributed to physiological mechanisms collectively known as the cochlear amplifier (CA), which amplifies and sharpens the basilar membrane (BM) vibration response to incoming acoustic pressure oscillations. Electromechanical properties of outer hair cells (OHCs) are believed to be the critical component of the CA, but its "circuitry" as yet remains unknown. Here, the required frequency-space response characteristics of the CA are computationally determined when typical in vivo tuning data are introduced as input to a linear hydroelastic cochlear model whose cross-sectional dynamics are represented by two coupled vibrational degrees of freedom. It is assumed that the CA senses motion at the tectorial membrane (TM) reticular lamina (RL) and applies proportional, equal, and opposite forces to the BM and the RL. The results show the CA to be tonotopically tuned, meaning it conforms to a space-frequency similarity principle like other cochlear dynamical responses. This requires that the active mechanism use information distributed along the cochlear partition. The physiological mechanism responsible for such behavior remains unknown, but here the computed CA characteristics can be qualitatively reproduced by a circuit spanning the length of the cochlea. This does not preclude other mechanisms, but is intended to motivate closer experimental investigation of extracellular and intercellular ionic flow pathways. PMID- 10530014 TI - Modeling otoacoustic emissions by active nonlinear oscillators. AB - The phenomenology of spontaneous otoacoustic emissions (OAEs) is compared to theoretical predictions given by models in which they are produced by active nonlinear oscillators. Along with the well-known Van der Pol oscillator, a new active oscillator model is proposed and analyzed here. Numerical simulations and multi-scale analytical computation results are compared to experimental data of neonatal spontaneous and evoked OAEs. A simple analysis technique is proposed, in which the time evolution after a click stimulus of the amplitude of each spectral line corresponding to a spontaneous OAE is studied. Apart from a few stationary lines, an approximately exponential decay law, with characteristic damping coefficients in the 20-200 Hz range, was found to fit the data. These results are not compatible with a Van der Pol oscillator model, and show that some important aspects of the OAE phenomenology can be better explained by the proposed oscillator. Other interesting features of the spontaneous end evoked OAE phenomenology, such as spontaneous OAE suppression by external tones and the following recovery, as well as stimulus/response curves in the linear and nonlinear mode of acquisition, are also well reproduced by the proposed model. PMID- 10530015 TI - Detection and intensity discrimination of Gaussian-shaped tone pulses as a function of duration. AB - Van Schijndel et al. [J. Acoust. Soc. Am. 105, 3425-3435 (1999)] proposed that the auditory system partitions the spectro-temporal domain into frequency-time (f t) windows and that the characteristics of these windows could be explored by measuring intensity discrimination for Gaussian-shaped tone pulses presented just above their detection threshold in noise. They reasoned that for a long-duration tone pulse, the auditory representation would be maximally compact in the frequency domain, but would spread across several f-t windows in the time domain. For a very-short-duration tone pulse, the auditory representation would be maximally compact in the time domain, but would spread across several f-t windows in the frequency domain. There should be some intermediate duration at which the auditory representation is compact in both the time and frequency domains and for which intensity-discrimination performance should worsen, due to the limited opportunity for multiple looks. Their data for signal frequencies of 1 and 4 kHz were consistent with this expectation; intensity discrimination was poorest at a duration of about 3-5 ms at 1 kHz and 1 ms at 4 kHz (durations are specified between 6.8-dB-down points on the envelope). This experiment attempted to replicate those results and to extend them to a wider range of frequencies and levels. Intensity discrimination of Gaussian-shaped tone pulses was measured at three levels: 10 dB above absolute threshold or above masked threshold in a pink noise with a spectrum level of either 15 or 40 dB at 1 kHz. The signal frequency was 0.25 kHz (durations from 2 to 320 ms), 1 kHz (durations from 0.5 to 80 ms), or 4 kHz (durations from 0.1 to 20 ms). Three normally hearing subjects were tested. At 1 and 4 kHz, performance was poorest overall for the 15-dB pink noise level, and thresholds showed a peak at intermediate durations (about 3-5 ms at 1 kHz and 1 ms at 4 kHz). Such peaks were still apparent, but smaller in the no noise condition and were almost absent at the higher noise level. For the 0.25 kHz signal frequency, peaks were not observed consistently at any level, although two subjects showed small peaks for durations around 10 ms. An explanation is offered for the results in terms of the level and frequency dependence of basilar membrane input-output functions. PMID- 10530016 TI - Intensity discrimination and detection of amplitude modulation. AB - Thresholds for detection of low-rate sinusoidal amplitude modulation and for detection of intensity increments were measured over a wide range of levels in an examination of the relationship between these fundamental aspects of intensity processing. As expected, thresholds measured with a continuous 1-kHz tone decrease with increasing carrier/pedestal level. For levels between 6 and 85 dB SPL the data are well described by 10 log delta I/I = 0.44.(20 log m) + D(fm), where delta I/I is the Weber fraction for increment detection, m is the modulation index at threshold, and D(fm) depends on modulation rate (fm). The relationship between the psychometric functions for modulation and increment detection is also consistent with this equation. The data indicate a clear relationship between modulation and increment detection and thus provide an important additional consideration for models of modulation processing. No existing models provide an adequate account of this relationship. PMID- 10530017 TI - Gap detection in single- and multiple-channel stimuli by LAURA cochlear implantees. AB - Gap-detection thresholds were determined for different complex patterns of electrical stimulation in four postlingually deafened LAURA cochlear implantees, to examine the nature of within- and across-channel auditory processes in more detail. Gap detectability was examined as a function of stimulus complexity (one, two, or three channels), channel distance within and across multichannel pre- and post-gap markers, stimulus asymmetry, and pulse rate. All markers roved in duration from 200 to 500 ms to ensure that subjects were not using overall stimulus duration as a cue. Gap-detection thresholds for all subjects were short (< 5 ms) when the pre- and post-gap markers stimulated the same single or multiple channels, even when the distance between simultaneously stimulated channels was large (exp. 1). For some subjects, gap detectability was more difficult in the across-channel condition, when the pre- and post-gap markers each stimulated different channels, although performance improved substantially in most subjects after extensive training (exp. 2). Additional tests with random maskers also suggest that neural interaction only affects performance mildly, and that the magnitude of the gap-detection threshold probably depends more on the subject's cognitive (in)ability to attend to the temporal gap than on the temporal acuity of their auditory system. Other stimulus conditions showed a difference in performance related to the order of the markers: gap thresholds were longer when the pre-gap marker stimulated one channel and the post-gap marker stimulated two or more channels, than vice versa (exp. 3). In addition, gap thresholds of three of the subjects increased with decreasing pulse rate from 1250 to 400 pps, a finding which may be related to the rate of the speech processing strategies used by each individual implantee (exp. 4). PMID- 10530018 TI - Dependence of binaural masking level differences on center frequency, masker bandwidth, and interaural parameters. AB - Thresholds for sinusoidal signals masked by noise of various bandwidths were obtained for three binaural configurations: N0S0 (both masker and signal interaurally in phase), N0S pi (masker interaurally in phase and signal interaurally phase-reversed), and N pi S0 (masker interaurally phase-reversed and signal interaurally in phase). Signal frequencies of 125, 250, 500, 1000, 2000, and 4000 Hz were combined with masker bandwidths of 5, 10, 25, 50, 100, 250, 500, 1000, 2000, 4000, and 8000 Hz, with the restriction that masker bandwidths never exceeded twice the signal frequency. The overall noise power was kept constant at 70 dB SPL for all bandwidths. Results, expressed as signal-to-total-noise power ratios, show that N0S0 thresholds generally decrease with increasing bandwidth, even for subcritical bandwidths. Only at frequencies of 2 and 4 kHz do thresholds appear to remain constant for bandwidths around the critical bandwidth. N0S pi thresholds are generally less dependent on bandwidth up to two or three times the (monaural) critical bandwidth. Beyond this bandwidth, thresholds decrease with a similar slope as for the N0S0 condition. N pi S0 conditions show about the same bandwidth dependence as N0S pi, but thresholds in the former condition are generally higher. This threshold difference is largest at low frequencies and disappears above 2 kHz. An explanation for wider operational binaural critical bandwidth is given which assumes that binaural disparities are combined across frequency in an optimally weighted way. PMID- 10530019 TI - Azimuthal tuning of human perceptual channels for sound location. AB - Human sound localization is acute for frontal locations, but relatively poor in the lateral hemifields. Previous studies in man have not, however, provided evidence on the tuning of the perceptual channels for auditory space that subserve this pattern of acuity. The spatial tuning of perceptual channels used in human azimuthal sound localization was determined using a between-channel auditory temporal gap detection paradigm. In this paradigm, gap thresholds are low when the markers bounding the silent period (gap) activate the same perceptual channel but are elevated when the two markers activate different channels. To determine the tuning of spatial channels, gap thresholds were obtained in an anechoic room with white noise markers coming from each combination of 12 leading marker locations and 18 trailing marker locations throughout the full 360 degrees of azimuth in the horizontal plane through the interaural axis. Gap thresholds remained low (2-4 ms) for all combinations of leading and trailing markers between 30 degrees and 150 degrees in both lateral hemifields. When the leading marker was located deep in one hemifield, and the trailing marker was in the opposite hemifield, gap thresholds rose to 8-16 ms. For leading marker locations at 30 degrees from the midline, gap thresholds were low for all trailing marker locations in the ipsilateral hemifield and locations near the midline in the contralateral hemifield, and were elevated (6-8 ms) only near the contralateral pole. Finally, for leading marker locations at 0 degree or 180 degrees, gap thresholds were low for any trailing location within 30 degrees of the midline at the front or back, and thresholds were elevated for trailing locations at the lateral poles. These data are accountable in terms of two broadly tuned perceptual channels, occupying the left and right auditory hemifields, respectively, each extending 30 degrees across the midline. These channels have widths and locations similar to the spatial receptive fields previously described for central auditory neurons in animals. The data suggest a model of spatial acuity based on the rates of activation of two spatially overlapping channels, rather than the selective activation of members of a large population of finely tuned channels. PMID- 10530020 TI - Auditory localization of nearby sources. II. Localization of a broadband source. AB - Although many researchers have examined auditory localization for relatively distant sound sources, little is known about the spatial perception of nearby sources. In the region within 1 m of a listener's head, defined as the "proximal region," the interaural level difference increases dramatically as the source approaches the head, while the interaural time delay is roughly independent of distance. An experiment has been performed to evaluate proximal-region localization performance. An auditory point source was moved to a random position within 1 m of the subject's head, and the subject responded by pointing to the perceived location of the sound with an electromagnetic position sensor. The overall angular error (17 degrees) was roughly comparable to previously measured results in distal-region experiments. Azimuth error increased slightly as the sound source approached the head, but elevation performance was essentially independent of source distance. Distance localization performance was generally better than has been reported in distal-region experiments and was strongly dependent on azimuth, with the stimulus-response correlation ranging from 0.85 to the side of the head to less than 0.4 in the median plane. The results suggest that the enlarged binaural difference cues found in the head-related transfer function (HRTF) for nearby sources are important to auditory distance perception in the proximal region. PMID- 10530021 TI - Determination of optimal data placement for psychometric function estimation: a computer simulation. AB - Psychometric functions are used to relate the responses of a subject to physical stimuli in a variety of psychophysical tasks. However, it is time consuming to obtain data to determine a psychometric function if many stimulus levels and many trials are required. A computer simulation was conducted to determine the minimum number of data points needed for such a determination. The computer simulation also determined the optimal placements of the stimuli and the number of trials per datum point for psychometric function determinations. Results indicate that a 2-point sampling method with 30-50 trials per point at optimal locations can produce a psychometric function with accurate spread and threshold estimates in a yes-no paradigm. However, the 4-point sampling method yields statistically smaller variances of the estimates. For the 2-alternative forced-choice paradigm, at least 120 trials per point are needed for the 2-point sampling method's estimated parameters to differ from the known parameter values by less than 5%. The simulation results suggest that 3-alternative or 4-alternative forced-choice is preferable to 2-alternative. Furthermore, when a criterion-free paradigm is not required, the yes-no paradigm is a better procedure than m-alternative forced choice for obtaining the corresponding psychometric function because of smaller standard deviation of the estimates and smaller number of trials/point required. PMID- 10530022 TI - Methods for estimating the sound pressure at the eardrum. AB - The problem of estimating the sound pressure generated at individual eardrums is systematically investigated. In audiometry, the reference to the pressure at the eardrum is usually realized by using a coupler such as the IEC 711 ear simulator, which is intended to approximate an average ear. The errors caused by individually shaped ear canals are calculated for a typical audiometric earphone (Beyer DT 48) in combination with the IEC 711 ear simulator and with an "ideal" coupler. These errors can reach 15 dB and are clearly more important than deviations of the ear simulator from an average ear. In order to obtain correct estimations, the chain matrices of individual ear canals have to be determined. Best estimates are obtained using the "reflectance phase method," but the "pressure minima method" also provides surprisingly good results, except in narrow frequency ranges. The reflectance phase method is checked using a physical model of the ear canal and the middle ear. The resulting errors of estimation remain within a limit of 3 dB up to more than 10 kHz. PMID- 10530023 TI - Geometry, kinematics, and acoustics of Tamil liquid consonants. AB - Tamil is unusual among the world's languages in that some of its dialects have five contrasting liquids. This paper focuses on the characterization of these sounds in terms of articulatory geometry and kinematics, as well as their articulatory-acoustic relations. This study illustrates the use of multiple techniques--static palatography, magnetic resonance imaging (MRI), and magnetometry (EMMA)--for investigating both static and dynamic articulatory characteristics using a single native speaker of Tamil. Dialectal merger and neutralization phenomena exhibited by the liquids of Tamil are discussed. Comparisons of English /[symbol: see text]/ and /l/ with Tamil provide evidence for generality in underlying mechanisms of rhotic and lateral production. The articulatory data justify the postulation of a class of rhotics and a class of laterals in Tamil, but do not provide evidence in favor of a larger class of liquids. Such a superclass appears to have largely an acoustic basis. PMID- 10530024 TI - Viscoelastic shear properties of human vocal fold mucosa: measurement methodology and empirical results. AB - A standard method for the empirical rheological characterization of viscoelastic materials was adopted to measure the viscoelastic shear properties of human vocal fold mucosal tissues (the superficial layer of lamina propria). A parallel-plate rotational rheometer was employed to measure shear deformation of viscoelastic tissue samples, which were deformed between two rigid circular plates rotating in small-amplitude sinusoidal oscillations. Elastic and viscous shear moduli of the samples were then quantified as a function of oscillation frequency (0.01 to 15 Hz) based on shear stresses and strains recorded by the rheometer. Data were obtained from 15 excised human larynges (10 male and 5 female). Results showed that the elastic shear modulus mu and the damping ratio zeta of human vocal-fold mucosa were relatively constant across the range of frequencies observed, while the dynamic viscosity eta decreased monotonically with frequency (i.e., shear thinning). Intersubject differences in mu and eta as large as an order of magnitude were observed, part of which may reflect age-related and gender-related differences. Some molecular interpretations of the findings are discussed. PMID- 10530025 TI - Interarticulator phasing, locus equations, and degree of coarticulation. AB - A locus equation plots the frequency of the second formant at vowel onset against the target frequency of the same formant for the vowel in a consonant-vowel sequence, across different vowel contexts. It has generally been assumed that the slope of the locus equation reflects the degree of coarticulation between the consonant and the vowel, with a steeper slope showing more coarticulation. This study examined the articulatory basis for this assumption. Four subjects participated and produced VCV sequences of the consonants /b, d, g/ and the vowels /i, a, u/. The movements of the tongue and the lips were recorded using a magnetometer system. One articulatory measure was the temporal phasing between the onset of the lip closing movement for the bilabial consonant and the onset of the tongue movement from the first to the second vowel in a VCV sequence. A second measure was the magnitude of the tongue movement during the oral stop closure, averaged across four receivers on the tongue. A third measure was the magnitude of the tongue movement from the onset of the second vowel to the tongue position for that vowel. When compared with the corresponding locus equations, no measure showed any support for the assumption that the slope serves as an index of the degree of coarticulation between the consonant and the vowel. PMID- 10530026 TI - Effects of syllable-initial voicing and speaking rate on the temporal characteristics of monosyllabic words. AB - Two speech production experiments tested the validity of the traditional method of creating voice-onset-time (VOT) continua for perceptual studies in which the systematic increase in VOT across the continuum is accompanied by a concomitant decrease in the duration of the following vowel. In experiment 1, segmental durations were measured for matched monosyllabic words beginning with either a voiced stop (e.g., big, duck, gap) or a voiceless stop (e.g., pig, tuck, cap). Results from four talkers showed that the change from voiced to voiceless stop produced not only an increase in VOT, but also a decrease in vowel duration. However, the decrease in vowel duration was consistently less than the increase in VOT. In experiment 2, results from four new talkers replicated these findings at two rates of speech, as well as highlighted the contrasting temporal effects on vowel duration of an increase in VOT due to a change in syllable-initial voicing versus a change in speaking rate. It was concluded that the traditional method of creating VOT continua for perceptual experiments, although not perfect, approximates natural speech by capturing the basic trade-off between VOT and vowel duration in syllable-initial voiced versus voiceless stop consonants. PMID- 10530027 TI - A model of auditory perception as front end for automatic speech recognition. AB - A front end for automatic speech recognizers is proposed and evaluated which is based on a quantitative model of the "effective" peripheral auditory processing. The model simulates both spectral and temporal properties of sound processing in the auditory system which were found in psychoacoustical and physiological experiments. The robustness of the auditory-based representation of speech was evaluated in speaker-independent, isolated word recognition experiments in different types of additive noise. The results show a higher robustness of the auditory front end in noise, compared to common mel-scale cepstral feature extraction. In a second set of experiments, different processing stages of the auditory front end were modified to study their contribution to robust speech signal representation in detail. The adaptive compression stage which enhances temporal changes of the input signal appeared to be the most important processing stage towards robust speech representation in noise. Low-pass filtering of the fast fluctuating envelope in each frequency band further reduces the influence of noise in the auditory-based representation of speech. PMID- 10530028 TI - Acoustical and perceptual study of gemination in Italian stops. AB - On the basis of theoretical considerations and results from acoustic and perceptual analyses, it is hypothesized that closure duration is the primary cue for gemination in Italian. Results of an acoustic analysis of a large number of single and geminate Italian utterances show two acoustic correlates of gemination: the length of the closure and the length of the vowel preceding the consonant. Other acoustic parameters were not systematically related to gemination. These results were validated perceptually. At the perceptual level, the above cues were used by the listeners in the geminate/nongeminate discrimination; however, closure duration played a major role. Moreover, it was found that the significant lengthening of consonant was only partially compensated by the shortening of the previous vowel and by a small lengthening of the geminate utterance with respect to the nongeminate one. This result suggests that speakers follow a sort of timing (rhythm) which is fixed in duration and depends on the number of syllables in the word: words with equal numbers of syllables do not change in utterance length, an elongated segment being partly compensated by the shortening of another. This process seems to be applied also perceptually suggesting that the timing (rhythm) of a language is also an auditory attitude. PMID- 10530029 TI - Contributions of temporal encodings of voicing, voicelessness, fundamental frequency, and amplitude variation to audio-visual and auditory speech perception. AB - Auditory and audio-visual speech perception was investigated using auditory signals of invariant spectral envelope that temporally encoded the presence of voiced and voiceless excitation, variations in amplitude envelope and F0. In experiment 1, the contribution of the timing of voicing was compared in consonant identification to the additional effects of variations in F0 and the amplitude of voiced speech. In audio-visual conditions only, amplitude variation slightly increased accuracy globally and for manner features. F0 variation slightly increased overall accuracy and manner perception in auditory and audio-visual conditions. Experiment 2 examined consonant information derived from the presence and amplitude variation of voiceless speech in addition to that from voicing, F0, and voiced speech amplitude. Binary indication of voiceless excitation improved accuracy overall and for voicing and manner. The amplitude variation of voiceless speech produced only a small increment in place of articulation scores. A final experiment examined audio-visual sentence perception using encodings of voiceless excitation and amplitude variation added to a signal representing voicing and F0. There was a contribution of amplitude variation to sentence perception, but not of voiceless excitation. The timing of voiced and voiceless excitation appears to be the major temporal cues to consonant identity. PMID- 10530030 TI - Recognition of spoken words by native and non-native listeners: talker-, listener , and item-related factors. AB - In order to gain insight into the interplay between the talker-, listener-, and item-related factors that influence speech perception, a large multi-talker database of digitally recorded spoken words was developed, and was then submitted to intelligibility tests with multiple listeners. Ten talkers produced two lists of words at three speaking rates. One list contained lexically "easy" words (words with few phonetically similar sounding "neighbors" with which they could be confused), and the other list contained lexically "hard" words (words with many phonetically similar sounding "neighbors"). An analysis of the intelligibility data obtained with native speakers of English (experiment 1) showed a strong effect of lexical similarity. Easy words had higher intelligibility scores than hard words. A strong effect of speaking rate was also found whereby slow and medium rate words had higher intelligibility scores than fast rate words. Finally, a relationship was also observed between the various stimulus factors whereby the perceptual difficulties imposed by one factor, such as a hard word spoken at a fast rate, could be overcome by the advantage gained through the listener's experience and familiarity with the speech of a particular talker. In experiment 2, the investigation was extended to another listener population, namely, non-native listeners. Results showed that the ability to take advantage of surface phonetic information, such as a consistent talker across items, is a perceptual skill that transfers easily from first to second language perception. However, non-native listeners had particular difficulty with lexically hard words even when familiarity with the items was controlled, suggesting that non-native word recognition may be compromised when fine phonetic discrimination at the segmental level is required. Taken together, the results of this study provide insight into the signal-dependent and signal-independent factors that influence spoken language processing in native and non-native listeners. PMID- 10530031 TI - Effects of lengthened formant transition duration on discrimination and neural representation of synthetic CV syllables by normal and learning-disabled children. AB - In order to investigate the precise acoustic features of stop consonants that pose perceptual difficulties for some children with learning problems, discrimination thresholds along two separate synthetic /da-ga/ continua were compared in a group of children with learning problems (LP) and a group of normal children. The continua differed only in the duration of the formant transitions. Results showed that simply lengthening the formant transition duration from 40 to 80 ms did not result in improved discrimination thresholds for the LP group relative to the normal group. Consistent with previous findings, an electrophysiologic response that is known to reflect the brain's representation of a change from one auditory stimulus to another--the mismatch negativity (MMN)- indicated diminished responses in the LP group relative to the normal group to /da/ versus /ga/ when the transition duration was 40 ms. In the lengthened transition duration condition the MMN responses from the LP group were more similar to those from the normal group, and were enhanced relative to the short transition duration condition. These data suggest that extending the duration of the critical portion of the acoustic stimulus can result in enhanced encoding at a preattentive neural level; however, this stimulus manipulation on its own is not a sufficient acoustic enhancement to facilitate increased perceptual discrimination of this place-of-articulation contrast. PMID- 10530032 TI - On the number of channels needed to understand speech. AB - Recent studies have shown that high levels of speech understanding could be achieved when the speech spectrum was divided into four channels and then reconstructed as a sum of four noise bands or sine waves with frequencies equal to the center frequencies of the channels. In these studies speech understanding was assessed using sentences produced by a single male talker. The aim of experiment 1 was to assess the number of channels necessary for a high level of speech understanding when sentences were produced by multiple talkers. In experiment 1, sentences produced by 135 different talkers were processed through n (2 < or = n < or = 16) number of channels, synthesized as a sum of n sine waves with frequencies equal to the center frequencies of the filters, and presented to normal-hearing listeners for identification. A minimum of five channels was needed to achieve a high level (90%) of speech understanding. Asymptotic performance was achieved with eight channels, at least for the speech material used in this study. The outcome of experiment 1 demonstrated that the number of channels needed to reach asymptotic performance varies as a function of the recognition task and/or need for listeners to attend to fine phonetic detail. In experiment 2, sentences were processed through 6 and 16 channels and quantized into a small number of steps. The purpose of this experiment was to investigate whether listeners use across-channel differences in amplitude to code frequency information, particularly when speech is processed through a small number of channels. For sentences processed through six channels there was a significant reduction in speech understanding when the spectral amplitudes were quantized into a small number (< 8) of steps. High levels (92%) of speech understanding were maintained for sentences processed through 16 channels and quantized into only 2 steps. The findings of experiment 2 suggest an inverse relationship between the importance of spectral amplitude resolution (number of steps) and spectral resolution (number of channels). PMID- 10530033 TI - Subharmonic backscattering from ultrasound contrast agents. AB - The ultrasonic contrast of blood in tissue, which is needed for ultrasonic estimation of tissue perfusion, can be increased by injecting the blood with bubbles or hollow microspheres. It has been shown that an even greater improvement in contrast can be obtained by using the subharmonic generated by irradiated microspheres. By obtaining analytical solutions to the modified RPNNP equation for a coated microbubble, the relationship between the physical parameters of the encapsulated bubble and the threshold pressure is established. The observed increase in the resonance frequency of a coated microsphere is explained by introducing the concept of "acoustic radius" of the encapsulated bubble. It is predicted that subharmonic generation in contrast agents requires a threshold insonifying pressure, and should be a minimum when microspheres are insonated at twice their resonance frequency. Experiments confirm the existence of this optimum incident frequency and of a reasonably low threshold pressure for the generation of the subharmonic. The existence of the low threshold pressures for subharmonic generation in contrast agents may prove to be very valuable in ultrasonic contrast imaging. PMID- 10530034 TI - Full-field mapping of ultrasonic field by light-source-synchronized projection. AB - A simple method for imaging ultrasonic fields in clear media is introduced. A modulated laser source is used to project the ultrasonic field onto a CCD camera. By use of the source-synchronized lock-in detection scheme, 2D images of the amplitude and phase distributions can be determined simultaneously. This technique is experimentally demonstrated with a 1-MHz and a 3.5-MHz ultrasonic transducer operated in continuous-wave mode. This method is very straightforward to implement and can be combined with the traditional tomographic reconstruction technique to obtain the 3D distribution of an ultrasonic field. PMID- 10530035 TI - Ecological approach of macrolide-lincosamides-streptogramin producing actinomyces from Cuban soils. AB - We report in this study the frequency of Streptomyces strains to produce macrolide-lincosamide-streptogramin (MLS) antibiotics isolated from Cuban soils. The screening assay is based on the induction of MLS-resistance phenotype in a clinical isolated strain of Staphylococcus aureus S-18. Our results suggest that of 800 Streptomyces strains isolated from different soil samples, 6% were positives in the screening test used. The ferralitic red soil from Pinar del Rio (north) provided the major percentage (3.6%) of MLS producing strains. The other soil samples tested belonging to Guira de Melena and Bauta in Havana, Matanzas City, Topes De Collantes (Villa Clara), and Soroa Mountains (Pinar del Rio) hill reached very low percentages. PMID- 10530036 TI - Molecular fingerprinting of Salmonella serotype Glostrup. AB - Thirteen isolates of Salmonella serotype Glostrup (antigenic formula, 6.8:z10:e,n,z15) from various sources and countries were analysed by ribotyping and IS200 fingerprinting. Both methods provided a high index of strain discrimination by allowing detection of three ribotypes and eight IS200 fingerprints which, though generally related, were readily distinguishable. The findings of this analysis confirm the usefulness of ribotyping and IS200 fingerprinting for studying the epidemiology of rarely isolated salmonellae of serogroup C. PMID- 10530037 TI - In situ analysis of the bacterial communities associated to farmed eel by whole cell hybridization. AB - Bacterial communities in water samples and eel slime were investigated by fluorescence in situ hybridization of whole bacterial cells in an eel intensive culture system over 1 year. A newly developed probe, matching 27 Vibrio spp., and a specific probe for Vibrio vulnificus were used. Phylogenetic probes complementary to selected regions of the 16S and 23S ribosomal RNA revealed that Proteobacteria of the alpha and beta subclass were predominant in water and eel slime. Members of the gamma subclass (e.g. vibrios and aeromonads) were more abundant in eel slime, although no V. vulnificus was detected. PMID- 10530038 TI - Combined action of nisin and carvacrol on Bacillus cereus and Listeria monocytogenes. AB - Nisin, a small antimicrobial protein, was tested for its bactericidal action against Listeria monocytogenes and Bacillus cereus and a typical biphasic reduction of the viable count was observed. The reduction was most fast during the first 10 min of exposure, while the viable count remained stable in the last part of the exposure period. Bacillus cereus was more sensitive towards nisin than L. monocytogenes and the inhibitory effect of nisin was stronger towards cells cultivated and exposed at 8 degrees C than towards cells cultivated and exposed at 20 degrees C. Combining nisin with sublethal doses of carvacrol resulted in an increased reduction in the viable count of both organisms, indicating synergy between nisin and carvacrol. Addition of lysozyme as a third preservative factor increased the synergistic effect between nisin and carvone, especially in the last part of the exposure period. PMID- 10530039 TI - Molecular characterization of Pseudomonas aeruginosa 2NR degrading naphthalene. AB - Three naphthalene-degrading strains were isolated from compost, characterized by morphological and physiological properties and differentiated by 16S rDNA RFLP. During growth on naphthalene, Pseudomonas aeruginosa 2NR produced ortho catechol pathway intermediates and gentisic acid. The ability to accumulate and degrade gentisic acid shows that Ps. aeruginosa 2NR has a different salicylate pathway to that of the intensely studied Ps. putida NCIB 9816. Molecular analysis showed the presence both of genes of the upper naphthalene pathway and genes of the ortho and meta catechol pathways. The insertion of nagH and nagG, coding for salicylate 5-hydroxylase in Pseudomonas sp. U2, was absent in Ps. aeruginosa 2NR, as in Ps. putida NCIMB 9816. PMID- 10530040 TI - Hydrogenation of polyunsaturated fatty acids by human colonic bacteria. AB - Emulsions of the fatty acids linoleic (C18:2 n-6), alpha-linolenic (C18:3 n-3) and arachidonic acid (C20:4 n-6) were incubated for 4 h under anaerobic conditions with human faecal suspensions. Linoleic acid was significantly decreased (P < 0.001) and there was a significant rise (P < 0.05) in its hydrogenation product, stearic acid. Linolenic acid was also significantly decreased (P < 0.01), and significant increases in C18:3 cis-trans isomers (P < 0.01) and linoleic acid (P < 0.05) were seen. With each acid, there were non significant increases in acids considered to be intermediates in biohydrogenation. The study provides evidence that bacteria from the human colon can hydrogenate C18 essential polyunsaturated fatty acids. However, with arachidonic acid there was no evidence of hydrogenation. PMID- 10530041 TI - Resistance to gentamicin and related aminoglycosides in Staphylococcus aureus isolated in Brazil. AB - Isolates of Staphylococcus aureus obtained from a Brazilian university hospital were characterized in relation to resistance to gentamicin and related aminoglycosides. Thirty-six isolates were susceptible to methicillin (MSSA) and 14 were resistant (MRSA). All isolates were sensitive to nucleic acid-binding compounds. All MRSA isolates and one MSSA isolate were demonstrated to be resistant to gentamicin and were coincidentally resistant to amikacin, kanamycin, neomycin and tobramycin. Among the gentamicin sensitive MSSA isolates, five isolates were found to be resistant only to kanamycin/neomycin. The resistance to gentamicin (and related aminoglycosides: kanamycin and tobramycin) must be due to AAC(6')-APH(2") activity. As these isolates also showed resistance to neomycin, they must carry an additional genetic element, probably the one responsible for APH(3')III activity, which accounts for the high level of resistance to kanamycin and to amikacin. The resistance to kanamycin/neomycin in the gentamicin sensitive isolates could not be attributed to the AAD(4')(4") activity because of the tobramycin sensitivity, and so could be ascribed to the APH(3')III activity. Curing and transfer experiments, as well as electrophoresis procedures, indicate that gentamicin resistance in Staph. aureus strains here studied has, characteristically, chromosomal localization. PMID- 10530042 TI - Inactivation of aflatoxigenic aspergilli by treatment with ozone. AB - The D-values of conidia of aflatoxigenic Aspergillus flavus and Aspergillus parasiticus exposed to 1.74 ppm. ozone in 1 mM potassium phosphate buffer (pH 7.0 and 5.5) at 25 degrees C were determined. D-values of A. flavus conidia were 1.72 and 1.54 min at pH 5.5 and 7.0, respectively; D-values of A. parasiticus were 2.08 and 1.71 min, respectively. None of these D-values was significantly (P < or = 0.05) different from each other. PMID- 10530044 TI - The waiting-list group. AB - This report describes a pilot study of a waiting-list group (preliminary process group [PPG]) that provided treatment for applicants to a university affiliated, urban mental health center. All individuals on the treatment waiting list were informed of the PPG. This semistructured group, meeting weekly, began with members presenting their problems, followed by free discussion, and ending with goal setting for the next week. Approximately one seventh (35 out of 262) of the clinic's applicants during a 4 1/2-month period chose to enter the PPG. They differed from those who chose not to participate (wait list) by being older and less educated. Approximately 80% of both wait-list and PPG participants subsequently entered therapy. Significantly more PPG patients than those on the wait list entered group treatment. The PPG served clinic needs by providing prompt service for self selected individuals and by supporting the group therapy program. PMID- 10530043 TI - Reduction of Escherichia coli O157:H7 by stimulated Pediococcus acidilactici. AB - This study was conducted to determine if stimulating the growth of meat starter culture (Pediococcus acidilactici) in a laboratory medium (Brain Heart Infusion broth +2.3% NaCl + 1.5% sucrose; LBHI) and during meat fermentation would control Escherichia coli O157:H7. In LBHI medium without P. acidilactici, the numbers of E. coli O157:H7 increased from 4.00 to 8.34 log10 cfu ml-1, whereas in the presence of P. acidilactici (approximately 6.0 log10 cfu ml-1) in LBHI, LBHIM (LBHI + 0.005% MnSO4), LBHIO (LBHI + 0.3 unit ml-1 Oxyrase), and LBHIMO (LBHI + M + O), the numbers of E. coli O157:H7 increased from 4.00 to 8.05, 7.50, 7.99, and 6.50 log10 cfu ml-1, respectively, after incubation at 40 degrees C for 15 h. During salami fermentation, the numbers of E. coli O157:H7 changed from 7.00 to 6.40 and 5.10 log10 cfu g-1 without and with P. acidilactici (approximately 7.0 log10 cfu g-1), respectively. Stimulated P. acidilactici by M, O, and MO further reduced the number of E. coli O157:H7 from 7.00 to 4.00, 4.80, and 3.65 log10 cfu g-1, respectively. The combination of MO was a better growth stimulator for P. acidilactici, which controlled E. coli O157:H7 in both systems (P < 0.05). PMID- 10530045 TI - Ethnocultural considerations in group psychotherapy. AB - The increasingly heterogeneous populations of the industrialized countries necessitate a reappraisal of the sociocultural norms and group-therapeutic approaches that were based primarily on White Judeo-Christian values. This author reviews the literature on the treatment of ethnically different patients and discusses socio-cultural phenomena relevant to group therapy that differ from the mainstream culture. The culturally determined contrasts in perception, attitude, communication, and behavior, which minority members may exhibit in the group, are examined. The specific demands on the therapist working with ethnic group members are highlighted along with the modifications in therapeutic technique. Recommendations for a culturally sensitive and broader training of group therapists are offered. PMID- 10530046 TI - Attachment style, traumatic bonding, and developing relational capacities in a long-term trauma group for women. AB - Adults with histories of severe childhood abuse often experience considerable difficulties with interpersonal trust. At the same time, they may strongly desire to be less alone with the painful aftereffects of their traumatic pasts. Psychotherapy groups have often been recommended as important components of treatment for reducing survivors' feelings of isolation and shame. We propose that an understanding of attachment styles and of traumatic bonding helps to clarify the specific manifestations of interpersonal distrust as they may emerge in a survivors' group. In addition, we suggest guidelines for determining what kind of group may be appropriate for a given individual at a particular point in treatment. PMID- 10530047 TI - A multiple group psychotherapy approach to adolescents with psychiatric and substance abuse comorbidity. AB - Multiple group psychotherapy was employed as the primary treatment modality in a day-treatment program as an innovative multifaceted approach to treating adolescents comorbid for psychiatric and substance abuse diagnoses. The concurrent educational program included a high school on site. The groups included Substance Abuse Group, which promoted the 12-step model; Health Group; Psychotherapy Group; Leisure Time Group; Self-Awareness Group; and Multiple Family Group. The effect of the multiple groups was to provide a variety of experiences focusing on varied aspects of normal and dysfunctional adolescent development. Together the combination of groups served to strengthen the participants' cohesiveness, communicating skills, and hopefulness. PMID- 10530048 TI - Joining group psychotherapy: developmental considerations. AB - This article examines the experience of new members joining an ongoing psychotherapy group. The group's stage of development and the new member's personality development are suggested to be important variables in this significant event. The experience of joining is found to share some things in common with the beginning phase of group for the group as a whole, but also noted are some unique elements. The process of joining is viewed as a highly anxious event for the new member, with antecedents in the individual's life experience. The new member is compromised by not knowing the group members, nor their shared history, norms, and dynamics. Concurrently, the group may experience a range of fears, wishes, and anxieties about the new member. Developmental considerations for the new member and the stage of group help to inform intervention strategies. PMID- 10530049 TI - Trauma-focused group therapy for patients with post-traumatic stress. PMID- 10530050 TI - Structuring provider-sponsored organizations. The legal and regulatory hurdles. PMID- 10530051 TI - Off-label prescribing. Legal implications. PMID- 10530052 TI - Physicians, malpractice, and state lines. A guide to personal jurisdiction in medical malpractice lawsuits. PMID- 10530053 TI - Equitable allocation of human organs. An examination of the new federal regulation. PMID- 10530054 TI - Health reform for children in Japan's aging society: a paradigm shift for pediatricians. AB - Japan confronts a dilemma in reshaping its health care for children: the low birth rate is reducing the proportion of children in the overall population, while the health issues for children are becoming increasingly complex and connected to social behavior. The present paper reviews Japan's changing demographics and the declining proportion of children in the population structure. This change has important implications for how the government sets priorities in the health sector. Next, the paper considers how 'problems' are defined for health policy in society and how the agenda for health reform is determined. Attention is directed to the political dimensions of these two processes of problem definition and agenda setting. Two examples of health reform for children, in other countries, are discussed to show how these two processes have worked in practice. In the next section, the paper explores new health issues for children in Japan and the challenges to Japanese pediatricians in shaping the agenda for health reform. PMID- 10530055 TI - Isolation of a Kawasaki disease-associated bacterial sequence from peripheral blood leukocytes. AB - BACKGROUND: The clinical and epidemiologic features of Kawasaki disease (KD) suggest an infectious etiology, but the agent(s) remains unknown. We aimed to isolate the causative bacterial gene from peripheral blood leukocytes of patients with acute KD. METHODS: Nested polymerase chain reaction (PCR) assay was used to amplify the bacterial 16S ribosomal RNA gene (rDNA). The amplified DNA were cloned into a plasmid vector and sequenced. Phylogenetic analysis was performed with clustal W program and the neighbour-joining method. RESULTS: First, the PCR reagents were examined by the PCR assay using conservative primers and we found more than 10 16S rDNA sequences contaminating the reagents. We then examined five KD patients using the PCR assay, excluding the contaminated sequences, and obtained five 16S rDNA sequences as possible KD-associated sequences. The primers specific to each 16S rDNA sequence were synthesized and used for specific PCR assays. Only the PCR assay specific to the 16S rDNA sequence termed 16S71-33 did not show any false positives with the control DNA from non-KD patients. The 16S71 33 sequence was positive in three of 20 patients with acute KD before gamma globulin therapy, but it became negative after therapy. The phylogenetic analysis showed a new species of the genus Corynebacterium as the origin of the 16S71-33 sequence. CONCLUSIONS: These data show that an infectious KD agent is traced in peripheral leukocytes and that a new Corynebacterium species may be responsible for KD in some cases. The true frequency and the role of the new Corynebacterium in KD would be clarified by measuring specific antibodies to it. PMID- 10530056 TI - Serum hepatocyte growth factor levels in Henoch-Schonlein purpura. AB - BACKGROUND: Henoch-Schonlein purpura (HSP) is the most common form of vasculitis in children. The potential role of hepatocyte growth factor (HGF) in acute immune mediated vasculitis has not been elucidated. METHODS: Serum HGF levels were determined in patients with HSP. RESULTS: In patients with acute-phase HSP, mean (+/- SD) serum HGF levels were 0.32 +/- 0.14 ng/mL and were significantly higher than those in the control group (0.11 +/- 0.10 ng/mL). This elevation of serum HGF levels recovered to control levels in parallel with improvement of the clinical symptoms. CONCLUSIONS: It is suggested that elevation of serum HGF levels in patients with acute-phase HSP may reflect endothelial cell damage or dysfunction in HSP. PMID- 10530057 TI - Clinical and biological aspects of acute lymphoblastic leukemia in 62 infants: retrospective analysis of the Tokyo Children's Cancer Study Group. AB - BACKGROUND: As a first step to formulate a new treatment strategy for refractory acute lymphoblastic leukemia (ALL) in infants, clinical results and immunophenotypic and cytogenetic data were analyzed and compared with those from overseas. METHODS: There were 62 infants with ALL who were treated between 1977 and 1995 at 30 institutions affiliated with the Tokyo Children's Cancer Study Group. Clinical and laboratory data obtained from these infants (all under 1 year of age) were retrospectively studied. RESULTS: The morphological diagnoses were FAB-L1 for 51 patients (82.2%) and FAB-L2 for 11 patients (17.8%). Hepatomegaly and splenomegaly were found in 40 (70.0%) and 40 patients (68.3%), respectively. The mean (+/- SEM) leukocyte count at diagnosis was 205,900 +/- 35,700/microL. The involvement of the central nervous system was evident in nine of 36 patients who were subjected to lumbar puncture, while three of these nine patients were free of neurological symptoms at diagnosis. Thirty-one patients (55.4%) were CD10 negative and 14 (25.0%) were CD10 positive. Thirty-one of 47 patients (65.9%) exhibited chromosomal abnormalities, including 28 patients (59.6%) with 11q23 abnormalities. Rearrangements in the MLL gene were found in nine of 13 infants (69.2%) examined. Translocation of 11q23 and/or MLL gene rearrangement (11q23/MLL) was significantly associated with the absence of the CD10 antigen. Hyperleukocytosis of more than 50,000/microL and 11q23/MLL gene rearrangements were related to a poor prognosis. The probability of an event-free survival in 62 infants was 13.1 +/- 4.8% at 48 months. CONCLUSIONS: New therapeutic strategies and large-scale cooperative prospective trials are needed to improve the prognosis of ALL in infants. PMID- 10530058 TI - Strong response of T cells in infants with dual infection by Epstein-Barr virus and cytomegalovirus. AB - BACKGROUND: Although a reversed CD4/CD8 ratio and increased proportion of CD8+ HLA-DR+ T cells are well known as the characteristic immune response in infectious mononucleosis (IM), it has not been elucidated whether these immune responses are affected by patient age and pathogenetic viruses. METHODS: T cell subsets were analyzed by two-color flow cytometry using fluorescein isothiocyanate- and phycoerythrin-conjugated monoclonal antibodies in 115 infants and children aged from 4 months to 10 years with IM due to Epstein-Barr virus (EBV), cytomegalovirus (CMV) and dual infection with both viruses. RESULTS: A reversed CD4/CD8 ratio and increased proportions of CD4+/HLA-DR+ T cells, CD8+ T cells and CD8+/HLA-DR+ T cells became more prominent as the age of the patients became older. No differences were observed in proportions of T cell subsets between EBV- and CMV-infection among patients aged from 6 to 17 months. Although the responses of these T cells were weak in infants with single virus infection by EBV and CMV, markedly strong T cell responses comparable with those in older children were observed in infants with EBV/CMV dual infection. Clinical symptoms were more severe in patients with EBV/CMV dual infection than those with EBV or CMV alone. CONCLUSION: The manner of these T cell responses in the acute phase of IM was considered to be age dependent, although strong T cell responses and severe disease were observed in EBV/CMV dual infection irrespective of patient age. PMID- 10530059 TI - High prevalence of Epstein-Barr virus type A strain with the 30 b.p. deletion of the latent membrane protein-1 gene in a Japanese population. AB - BACKGROUND: The pathogenic activity of Epstein-Barr virus (EBV) with a characteristic 30 b.p. deletion of the latent membrane protein-1 (LMP-1) gene is controversial. We analyzed the LMP-1 gene and two major strains of EBV, type A and type B, in Japanese patients with EBV-associated disease. METHODS: We directly sequenced the carboxy terminal part of the LMP-1 gene from 15 EBV infected patients; 10 patients with infectious mononucleosis (IM) and one patient each with Hodgkin's disease, B cell lymphoma, Wiskott-Aldrich syndrome (WAS), AIDS and atypical EBV infection (atEBV). The EBV subtype was studied by determining the 3' divergence of Epstein-Barr virus nuclear antigen (EBNA)-2 using polymerase chain reaction primers. RESULTS: Twelve of 15 patients had EBV with the 30 b.p. deletion and numerous point mutations of the LMP-1 gene, regardless of the disease. Two patients, one with IM and one with WAS, had EBV without the 30 b.p. deletion. One patient with atEBV had two types of LMP-1 gene, one with and one without the 30 b.p. deletion. Thirteen patients had EBV type A, the WAS patient had the type B strain and the atEBV patient had both types A and B. In the patient with atEBV, the two types of LMP-1 gene and two EBV subtypes were detected simultaneously. CONCLUSIONS: The characteristic 30 b.p. deletion of the LMP-1 gene is not an important factor in the pathogenesis of EBV-associated diseases. The EBV type A strain with the 30 b.p. deletion of the LMP-1 gene is prevalent in the Japanese population. PMID- 10530060 TI - Negative C-reactive protein in children with bacterial infection. AB - PURPOSE: To evaluate the practical value of initial C-reactive protein (CRP) in the diagnosis of bacterial infection in children. METHODS: The subjects comprised 11 children, six boys and five girls, aged 3 months through to 3 years (median age 16 months), whose initial CRP levels were < 1.0 mg/dL despite bacterial infection. C-reactive protein was quantitated at the first medical examination by nephelometry. RESULTS: The diagnosis was urinary tract infection (n = 4), bacterial meningitis (n = 2), sepsis (n = 2), pneumonia (n = 2) and arthritis of the hip joint (n = 1). The CRP levels were significantly elevated during the course of infection, ranging from 7.6 to 28.5 mg/dL. The bacterial etiology was non-specific. Eight patients were examined within 12 h of onset, three exhibited negative CRP values despite the duration of the insult over 12 h. Six patients were tentatively diagnosed as having a bacterial infection, but the other five were not. Each patient was treated, leading to a favorable outcome without any serious complications. CONCLUSIONS: Low levels of CRP do not rule out the possibility of bacterial infection in children. The initial value of CRP may be negative, even in patients with severe bacterial infection or even after 12 h from onset. The data suggest that pediatricians should consistently be aware of the possibility of bacterial infection even if the initial CRP test result is negative and that serial CRP measurements appear to be practical. PMID- 10530061 TI - Lymphocyte subpopulations in full-term septic neonates. AB - PURPOSE: To estimate the absolute leukocyte and lymphocyte counts and relative and absolute sizes of CD19+ B lymphocytes, CD3+, CD4+, CD8+ and CD3+/HLA-DR+ T lymphocytes in full-term septic neonates and the influence of some perinatal risk factors on these lymphocyte subsets. METHODS: Twenty-one septic and mechanically ventilated full-term neonates (13 boys and eight girls) and 15 healthy full-term neonates born vaginally with an Apgar score > 9 and without hyperbilirubinemia were investigated. Two-color flow cytometric immunophenotyping with appropriate antibody panels using lysed whole vein blood was performed. RESULTS: The mean relative and absolute sizes of CD19+ B lymphocytes, CD3+/CD8+ and CD3+/HLA-DR+ T lymphocytes in septic neonates did not differ significantly from control. In contrast, the mean relative sizes of CD3+ and CD3+/CD4+ T lymphocytes and the CD4+/CD8+ ratio in septic neonates were significantly higher than in healthy neonates. With regard to the absolute size in septic neonates, only CD4+ T cells were significantly higher compared with the control group. Perinatal risk factors (birth asphyxia, gestation and delivery complications) had no significant effect on the relative and absolute counts of all estimated lymphocyte subpopulations in septic neonates. CONCLUSIONS: Increases in the relative sizes of CD3+ and CD3+/CD4+ T lymphocytes and the CD4+/CD8+ ratio in full-term septic neonates provides important information about changes in cell-mediated immunity during the early neonatal period. PMID- 10530062 TI - Seroprevalence of cytomegalovirus infection among children and females in Ankara, Turkey, 1995. AB - BACKGROUND: Cytomegalovirus (CMV) is the most frequent known cause of congenital viral infections in humans. Cytomegalovirus is endemic throughout the world, affecting most of the population where the seroprevalence of CMV IgG is known to vary among countries. METHODS: The present study was designed to show the prevalence of CMV antibodies among children aged 1 day to 15 years and women of child-bearing age in Ankara, Turkey. Antibodies to CMV were analyzed in serum samples of 318 children and 745 women using a passive particle-agglutination test. RESULTS: The overall prevalence of CMV antibodies was 90.6% among children and 99% among women aged 15-49 years. The difference between stratified age groups was not statistically significant (chi 2 = 4.92, P = 0.177) in either children or females. CONCLUSIONS: Our findings confirm that CMV is very prevalent in Turkey and is at the higher end of worldwide ranges. Using the results of the present study, the transmission mode of CMV infection and the risk for congenital CMV infection are discussed. We have come to the conclusion that the risk of fetal infection in Turkey cannot be predicted; however, most CMV infections in the first year of life are transmitted from mother to infant and this is the main source of infection in Turkey. PMID- 10530064 TI - Neutrophil chemotaxis in acutely infected and clinically stable cystic fibrosis patients. AB - PURPOSE: The aim of the present study was to evaluate the role of neutrophil chemotaxis in cystic fibrosis (CF) and to also determine whether an acute bacterial infection and the nutritional status of a child can affect neutrophil chemotaxis. METHODS: Twelve acutely infected and 12 clinically stable CF patients and 10 healthy age-matched controls were studied. Neutrophil chemotaxis and random migration were investigated in vitro in the peripheral blood of subjects by the Boyden chamber method and the results were expressed as chemotactic index (CI). The nutritional status of the cases was evaluated as body mass index (BMI). RESULTS: The CI values in the acutely infected group were found to be significantly lower than the clinically stable and healthy control groups (P < 0.05 and P < 0.005, respectively). There was no significant difference between the clinically stable CF group and the healthy control group (P > 0.1). No significant correlation was detected between the CI and BMI of the two groups of CF patients (P > 0.05). CONCLUSIONS: The present study confirms that neutrophil chemotaxis and random migration are normal in clinically stable CF patients. The decreased CI in the acutely infected patients indicates the possible role of infection itself on neutrophil chemotaxis. PMID- 10530063 TI - Tuberculous peritonitis in 11 children: clinical features and diagnostic approach. AB - BACKGROUND: Tuberculous peritonitis (TBP) is a rare manifestation of childhood tuberculosis characterized by long-lasting abdominal symptoms and exudate and lymphocytes in the ascitic fluid. The diagnosis of TBP is rarely established unless a high index of suspicion is maintained. METHODS: The diagnostic features of 11 cases who were hospitalized with TBP in the Pediatric Infectious Diseases Ward of Dicle University Hospital, Turkey, were evaluated retrospectively. RESULTS: Seven cases were male and the ages of all cases ranged between 1 and 11 years. The onset of symptoms was 1-12 months (mean +/- SD 3.1 +/- 2.7 months) prior to the admission time. Nine patients gave a history of familial tuberculosis. Three cases had Bacillus Calmette-Guerin (BCG) scars and the results of five tuberculin unit (TU) tests in cases without and with BCG were over 10 and 15 mm, respectively. The most common presenting clinical symptoms and signs at admission were abdominal distention and ascites (100%), fever (27%) and loss of weight (18%). One case had accompanying tuberculous meningitis and two cases had concomitant pulmonary tuberculosis. Only one of 11 samples of ascitic fluid yielded Mycobacterium tuberculosis by the polymerase chain reaction method and no other microbiologic evidence was obtained in culture specimens. Ultrasonographic and computed tomographic imagings revealed high-density ascites that contributed well to the diagnosis. The diagnosis in two patients was proven histopathologically via peritoneoscopy and laparoscopy. All cases were treated with isoniazide, rifampisin for 9 months and pyrazinamide for the first 2 months. CONCLUSIONS: Radiologic diagnostic techniques, positive skin tests and a history of exposure to tuberculosis may contribute to the diagnosis of TBP, helped by clinical symptoms and findings, particularly when invasive diagnostic methods via peritoneoscopy and laparoscopy are not available in developing countries. PMID- 10530065 TI - Acute lower respiratory infections in hospitalized children over a 6 year period in Tokyo. AB - BACKGROUND: Acute lower respiratory infections are major causes of hospitalization in children and are mainly caused by respiratory viruses. In the present study, we investigated the etiologic agents responsible for acute lower respiratory infections from the period November 1986 to October 1992 in order to determine the seasonal pattern and different characteristics of age distribution of respiratory infectious agents, mainly virus infections. METHODS: A total of 1521 patients with lower respiratory infections was hospitalized in Saiseikai Central Hospital, Tokyo, Japan. Nasopharyngeal secretions were obtained for virus isolation and paired sera in the acute and convalescent phases were obtained for serological examination. RESULTS: Etiological agents were identified in 668 of 1521 patients (43.9%) by serological antibody responses, virus isolation and/or detection of virus antigen: 240 (15.8%) with respiratory syncytial (RS) virus; 62 (4.1%) with influenza virus type A; 26 (1.7%) with influenza virus type B; 86 (5.7%) with adenovirus; 81 (5.3%) with parainfluenza virus; 32 (2.1%) with measles virus; 20 (1.3%) with enteroviruses or Herpes virus other than respiratory viruses; 75 (4.9%) with Mycoplasma pneumoniae; 10 (0.7%) with pertussis; and 36 (2.4%) with mixed infections. In the remaining 853 patients (56.1%), etiologic agents were not identified. Respiratory syncytial (RS) virus was a main causative agent of respiratory infections in patients younger than 3 years of age. Influenza virus and M. pneumoniae were two main causative agents in patients with acute respiratory illness over 5 years of age. Parainfluenza virus type 3 was frequently observed in infants from 9 to 12 months of age. A distinct seasonal pattern of viral infections was consistently observed in each year during the study period; RS and influenza viruses were prevalent in winter, parainfluenza virus was prevalent in spring and M. pneumoniae was prevalent in summer and autumn. However, adenovirus infections were observed in all seasons. Serological responses were poor in patients younger than 1 year of age and they were mainly diagnosed by virus isolation or detection of virus antigen. CONCLUSIONS: Virological epidemiology provides useful information in daily clinical practice for the prediction of etiological agents based on patient age and the seasonal distribution of agents. We should examine virus isolation and the detection of virus antigen, along with serological examinations in patients with respiratory infections, especially in infants younger than 1 year of age because of poor serological responses. PMID- 10530067 TI - Autonomic nervous system function in childhood migraine. AB - BACKGROUND: Although the pathogenesis of migraine is controversial, autonomic nervous system (ANS) dysfunction has been reported in patients with adult migraine in recent years. The present study was planned to investigate ANS function in childhood migraine. METHODS: The migraine and control groups consisted of 25 migraineur and 30 healthy children, respectively. Orthostatic test, sustained handgrip, Valsalva ratio, 30/15 ratio and heart rate responses to deep breathing were used as non-invasive ANS function tests in both groups. RESULTS: In the orthostatic test, systolic (SBP) and diastolic blood pressures (DBP) were higher in the upright than the supine position in the migraine group, but were higher in the supine than upright position in the control group. In the sustained handgrip test, the mean difference in SBP was higher in the migraine than the control group (P = 0.0278), but there was no significant difference in DBP between migraine and control groups (P = 0.107). The Valsalva ratio was higher in the migraine than the control group (P = 0.0002), as was the 30/15 ratio (P = 0.0108). Heart rate responses to deep breathing were not different between the migraine and control groups (P = 0.749). CONCLUSIONS: Our results demonstrate ANS dysfunction, with hyperactivity of both the sympathetic and parasympathetic nervous system, in children with migraine. PMID- 10530066 TI - Reduction in blood glucose values following indomethacin therapy for patent ductus arteriosus. AB - PURPOSE: To evaluate the effects of indomethacin on blood glucose values in premature infants with patent ductus arteriosus (PDA). METHODS: Twenty-five very low birthweight infants with PDA were given 0.2 mg/kg, i.v., indomethacin for up to three doses. We examined the relationship between blood glucose values and glucose infusion rate before and after indomethacin therapy. RESULTS: There was a significant reduction in blood glucose values between 12 and 96 h following i.v. indomethacin therapy. Eleven of 25 infants (44%) had blood glucose values below 40 mg/dL between 12 and 60 h (mean 32.7 h) after the initial dose. Although the glucose infusion rate during the first 12 h was constant (3.56 +/- 0.98 mg/kg per min), the blood glucose values decreased from 96 +/- 32 mg/dL at the starting point to 75 +/- 29 mg/dL at 12 h (P < 0.05). The maximum blood glucose reduction was 51.6 +/- 34.7 mg/dL and the maximum blood glucose reduction rate was 50.4 +/- 20.2%. CONCLUSIONS: The results suggest that blood glucose values should be measured at least every 6 h for 72 h until they stabilize in order to prevent unexpected hypoglycemia. PMID- 10530068 TI - Histopathologic aspects of endomyocardial biopsy in pediatric patients with idiopathic ventricular tachycardia. AB - PURPOSE: The present study aimed to investigate the clinicopathologic findings and histopathologic characteristics of endomyocardial biopsy in pediatric patients with idiopathic ventricular tachycardia. METHODS: Histopathological findings of endomyocardial biopsy from 17 patients aged 7-15 years with idiopathic ventricular tachycardia (VT) but no organic heart disease were examined. Patients considered to have cardiomyopathy of the dilated, hypertrophic or specific form or arrhythmogenic right ventricular cardiomyopathy were excluded from this study. RESULTS: Advanced histopathologic findings, including myocyte hypertrophy, degeneration, interstitial fibrosis and disarrangement of muscle bundles, were disclosed in three cases (17.6%). One of these cases exhibited sustained VT with left bundle branch block configuration and showed increased frequency of VT during exercise testing. The remaining two cases had non sustained VT with multifocal origin and had syncope episodes. Another 14 cases showed mild or no significant findings in the biopsy. CONCLUSIONS: These results indicate that advanced histopathology in endomyocardial biopsy is occasionally disclosed in cases of idiopathic VT, especially those of exercise-related VT or multifocal VT, and that these patients may be considered as having heart muscle disease. PMID- 10530069 TI - Efficacy and safety of rectal thiopental: sedation for children undergoing computed tomography and magnetic resonance imaging. AB - PURPOSE: We evaluated the clinical safety, effectiveness, efficiency and potential side effects of rectally administered thiopental in 30 children undergoing computed tomography (CT) and magnetic resonance imaging (MRI). METHODS: The doses of thiopental used were 50 mg/kg for infants under 6 months of age, 35 mg/kg for infants between 6 and 12 months of age and 25 mg/kg for older children. After administration of the sedative, oxygen saturation was continuously monitored and vital signs were recorded every 20 min during the imaging procedure and then every 20 min until discharge. RESULTS: Successful sedation and adequate imaging were obtained in 29 of 30 (96.7%) patients. Respiratory depression was not observed in any patient. However, oxygen saturation dropped below 90% transiently (to 88%) in three patients (10.0%) and this was immediately corrected by repositioning the child's neck to open the upper airway. All successfully sedated patients were asleep within 15 min (mean +/- SD 7.3 +/- 2.7 min) and sedation was sufficient for at least 30 min. Prolonged sedation was observed in two patients. CONCLUSIONS: We believe that rectal thiopental is a safe, effective and efficient form of sedation for pediatric imaging. PMID- 10530070 TI - Infant feeding and growth: a study on Turkish infants from birth to 6 months. AB - BACKGROUND: To evaluate the impact of various feeding patterns on the physical growth and mental development of infants, particularly during the first 6 months of life, and to compare growth patterns of Turkish infants with those of infants living in various countries. METHODS: One hundred and seventy-two healthy newborn infants were included in the study and were divided into three feeding groups: (i) 62 infants were exclusively breast-fed (BF); (ii) 58 infants were mixed-fed (MF) with both breast milk and formula; and (iii) 52 infants were formula-fed (FF). Infants were assessed at birth and at 1, 2, 3, 4, 5 and 6 months of age. Anthropometry was repeated on each occasion. The weight and length of the infants was also recorded. Analysis of variance and modified t-test were used for statistical evaluation of the results. RESULTS: Values in the BF group were the closest to the tabular norms for weight. Infants in the FF group tended towards a lower weight during the first 3 months (P < 0.05). During the second 3 months, weight gain observed in the FF group was significantly higher than that of BF infants. In comparison with MF infants, a significant progressive weight gain was detected in BF infants (P < 0.05). The values obtained for length increments were consistent with those for weight (P < 0.05 for BF vs FF). No significant difference was found between the length increments detected for BF and MF infants from birth to 6 months. CONCLUSIONS: These results suggest that exclusive breast feeding is the most appropriate feeding pattern for newborn infants in Turkey and is sufficient during the first 6 months, the most important fraction of life. PMID- 10530071 TI - Prevalence and correlates of malnutrition among children in rural minority areas of China. AB - BACKGROUND: Child growth retardation and malnutrition remain a matter of uttermost public concern in economically disadvantaged areas of China. The present study aimed to estimate the prevalence of protein-energy malnutrition with various anthropometric indices and examine its correlates in a large sample of poor rural minority children. METHODS: A total of 2019 children under 7 years of age belonging to the Hani, Yi, Hui, Miao ethnic minority groups and the Han major group were drawn from four poor rural minority counties in the Yunnan Province of China. Well-trained investigators completed child physical measurements and maternal interviews. Protein-energy malnutrition was defined as being underweight (weight for age), wasting (weight for height) and stunting (height for age) on the basis of reference data from the National Center of Health Statistics (NCHS)/World Health Organization (WHO). RESULTS: The respective prevalence of moderate and severe protein-energy malnutrition was 15.8 and 3.1% for underweight children, 31.8 and 19.2% for stunting and 0.9 and 0.5% for wasting. Stunting was most common in children aged 2 years. Boys were more likely to suffer from malnutrition. Logistic regression analyses showed that lower family income, lower parental height, belonging to the Miao, Yi and Hani ethnic groups compared with Han and poorer maternal child-rearing behavior significantly increased the risk for stunting of children. CONCLUSIONS: Protein-energy malnutrition is relatively high in the rural minority children of China. Chronic socioeconomic underdevelopment and genetic effects, rather than a severe or immediate lack of food, may lead to protein-energy malnutrition. PMID- 10530072 TI - Maternal preeclampsia and jitteriness in preterm infants. AB - AIM: To clarify the role of maternal preeclampsia in jitteriness in preterm infants. METHODS: Sixteen premature infants of preeclamptic mothers were observed for occurrence of jitteriness and were compared with 32 premature infants born to normotensive women. RESULTS: Jitteriness was present in a significantly higher percentage (75 vs 6%) and persisted longer (4.5 +/- 5.6 vs 1.5 +/- 0.7 days) in the preterm infants of preeclamptic mothers. CONCLUSIONS: Maternal preeclampsia could be included among the pathological factors that cause jitteriness in preterm babies. PMID- 10530073 TI - Pancytopenia with hemophagocytosis secondary to parvovirus B19 infection in a family with hereditary spherocytosis. PMID- 10530074 TI - Terminal deletion of chromosome 10q: clinical features and literature review. PMID- 10530075 TI - Cervical esophageal web in a 13-year-old boy with growth failure. PMID- 10530076 TI - Lethal junctional epidermolysis bullosa showing mild blister at birth. PMID- 10530077 TI - An infant with severe atopic dermatitis and progressive hepatomegaly due to fatty liver. PMID- 10530078 TI - Diaphragmatic hernia in an infant of a diabetic mother: an unusual association in diabetic embryopathy. PMID- 10530079 TI - Effective treatment of cyclic thrombocytopenia with cepharanthin. PMID- 10530080 TI - 'Hypertrophic cardiomyopathy' in a girl with adult-type myotonic dystrophy. PMID- 10530081 TI - Chronic granulomatous disease in Japan: incidence and natural history. The Study Group of Phagocyte Disorders of Japan. AB - PURPOSE: To analyze the incidence and causes of fatality of patients with chronic granulomatous disease (CGD) in Japan and to provide an opportunity for comparison with reports from other countries. METHODS: The Study Group of Phagocytic Disorders in Japan conducted a questionnaire survey on CGD patients in Japan, results of which formed the basis of the study. RESULTS: Clinical details of 221 patients were analyzed: 194 male and 27 female (ratio: 7.2/1), 152 living, 51 dead and 18 unknown. The prevalence of CGD was estimated to be 1/287,709 live births. The fatality rate was 23.1%. The mean age of the surviving patients increased from 8 years 4 months in 1985 to 16 years 0 months in 1998. Although the mean age of death was advanced by 4 years 11 months during the same period of time, the fatality rate has remained practically unchanged during the past 13 years. The number of living adult patients has tended to increase (32.1%). At the time of study, approximately 90% of patients had been placed on sulfamethoxazole trimethoprim prophylactically or therapeutically, either singly or in combination with other modalities, including interferon-gamma, antifungal agents and various antibiotics. Pulmonary infections were responsible for 58.3% of fatalities. CONCLUSIONS: With early diagnosis and prompt institution of appropriate therapy, the mean age of CGD patients in Japan has been increasing, but the fatality rate has remained practically unchanged during the last 13 years, mostly due to the fungal infections. PMID- 10530082 TI - Analysis and evaluation of the Trajectory Theory of Chronic Illness Management. AB - The development and testing of theories for use in nursing research and practice is essential for advancement of the profession. The Trajectory Theory of Chronic Illness Management is a middle-range nursing theory that has been proposed by Corbin and Strauss (1991a). Analysis and evaluation of this theory was performed using Fawcett and Downs's (1992) guidelines. Theory analysis and evaluation are important first steps before using a theory for practical purposes. Theory analysis makes a theory more understandable and helps to identify the strengths and weaknesses of the theory. Evaluation extends the analysis process by making judgments about the potential contribution of the theory based on published data. Results of the theory analysis and evaluation suggest that the Trajectory Theory has theoretical and social significance but that further theoretical work is necessary to enhance the internal consistency and parsimony of the theory. Although several authors have suggested that the theory has pragmatic adequacy, evidence for empirical adequacy of the theory is needed. Before more empirical studies are conducted, further theoretical work is recommended. PMID- 10530084 TI - Reflections on clinical judgment of nurse practitioners. PMID- 10530083 TI - Patterns of resistance: African American mothers and adult children with HIV illness. AB - Although the research on caregiving and caregivers has been extensive, there have been few studies on the cultural context and meaning of African American caregiving in relation to HIV illness. Many Black feminists have argued that African American women experience a world different from those who are not Black and that failure to take account of race, class, and gender is paramount in an attempt to authentically portray the lives of African American women. This study argues that rural African American culture and experiences of racism and discrimination in the rural South shaped the responses of mothers when their adult children developed HIV illness. The study employed the ethnographic techniques of participant observation and in-depth interviews with 14 rural, poor, African American mothers who cared for adult children with HIV illness. Analysis of the data identified patterns of resistance that mothers employed throughout the caregiving experience. Mothers resisted labels and other controlling images that they believed marginalized them and negated what was happening to their children. Mothers used culturally patterned behaviors to protect their families and resist the stigma of HIV/AIDS. PMID- 10530085 TI - Recommended composition of influenza virus vaccines for use in 2000. PMID- 10530086 TI - A new slant on memory loss. PMID- 10530087 TI - Understanding your triglyceride levels. PMID- 10530088 TI - Cosmetic surgery: is it for you? PMID- 10530089 TI - Reaping the benefits of designer estrogens. PMID- 10530091 TI - Which decongestants are safe to use if you have high blood pressure? PMID- 10530090 TI - Are the new plastic corneal ring implants a safe alternative to eye-glasses and laser eye surgery? PMID- 10530092 TI - Electric toothbrush use. Attitudes and experience among dental practitioners in Germany. AB - PURPOSE: To evaluate the attitudes and experience of electric toothbrush use among dental practitioners in Germany. MATERIALS AND METHODS: A telephone survey of 399 dentists was conducted. RESULTS: Many dentists in Germany (41%) thought that between half and 70% of their patients do not clean their teeth correctly, and that this is the result of either poor brushing technique or insufficient brushing time coupled with insufficient visits to the dentist. Most dentists surveyed (61%) would recommend an electric toothbrush to their patients in order to improve oral hygiene control, and of these, 82% would recommend the Braun Oral B Plaque Remover. When these dentists were asked if there had been any change in tooth and gum condition among those patients who switched from using a manual toothbrush to the Braun Oral-B Plaque Remover, 73% said that they had observed an improvement. None of the dentists interviewed had noted any deterioration. These views of German dentists are in agreement with laboratory and clinical studies that have shown an advantage for the Braun Oral-B Plaque Remover over a manual toothbrush. The results indicate that clinical trial results are a relevant way of assessing toothbrush performance, and that results may be extrapolated to normal home use. PMID- 10530093 TI - Cleaning efficacy of a new electric toothbrush. AB - PURPOSE: To evaluate the cleaning efficacy of a new electric toothbrush and new brush head design, using a robot system to simulate normal clinical toothbrush use. MATERIALS AND METHODS: The study compared the new oscillating/rotating/pulsating electric toothbrush (Braun Oral-B 3D Plaque Remover) comprised of the D15 handle and new EB15 brush head, with the clinically established oscillating/rotating electric toothbrush (Braun Oral-B Ultra Plaque Remover-D9) and brush head (EB9). Plaque substitute was applied to the artificial teeth of typodonts, which were cleaned by the robot system for 2 minutes. The remaining plaque substitute was measured for buccal + lingual/palatal and occlusal surfaces, as well as gumline and interproximal sites using a computerized vision system. RESULTS: The new D15/EB15 combination was found to be significantly more effective than the D9/EB9 combination in removing plaque substitute at all surfaces combined and all specific sites (P < 0.001). These results show that the new toothbrush which features an additional pulsating motion (D15), combined with the new brush head design (EB15), offer improved efficacy in comparison with the established oscillating/rotating combination (D9/EB9). PMID- 10530094 TI - Clinical plaque removing efficacy of a new power toothbrush. AB - PURPOSE: To evaluate the effect of adding a pulsating bristle action to the established oscillating/rotating action of the Braun Oral-B Ultra Plaque Remover (D9) on plaque removal. MATERIALS AND METHODS: Plaque removal was evaluated using the modified Quigley-Hein Plaque Index in a double blind randomized, crossover study involving 32 healthy volunteers without any dental training. After 2 weeks use of the D9 during which time subjects received training in its use, subjects abstained from oral hygiene for 48 hours. They were then assessed for plaque after which they brushed their teeth using an experimental toothbrush randomly set to either the D9 oscillating/rotating action or to the new 3D action with an additional pulsating movement of the brush head in the direction of the long axis of the bristles. After brushing, plaque was again evaluated. Following a further 2 weeks of normal home use of the D9, subjects returned and the procedure was repeated using the brush set in the second mode. RESULTS: Both toothbrush actions were found to be effective at removing plaque from all sites and surfaces in the mouth. The 3D action was consistently more effective than that of the D9, the difference being statistically significant for the whole mouth, the upper jaw, the lingual surfaces and for all interproximal sites, in particular in the upper jaw. PMID- 10530095 TI - A 3-month clinical investigation comparing the safety and efficacy of a novel electric toothbrush (Braun Oral-B 3D Plaque Remover) with a manual toothbrush. AB - PURPOSE: To compare the efficacy and safety of a novel electric toothbrush (Braun Oral-B 3D Plaque Remover) with a standard reference ADA manual toothbrush. MATERIALS AND METHODS: 114 subjects were included in a 3-month randomized, parallel group, examiner-blind study and divided into two groups: 3D users and manual toothbrush users. Subjects were instructed to brush twice daily for 2 minutes. Evaluation of oral soft and hard tissue for safety, plaque, gingivitis and bleeding was conducted prior to the start of product use (at baseline), at days 14 and 35, and at 3 months. RESULTS: 105 subjects (55 3D users and 50 manual toothbrush users) completed the study. At days 14, 35 and at 3 months both groups showed reductions from baseline in whole mouth plaque, gingivitis and bleeding that were statistically significant (P < 0.005), except in the case of plaque at day 35 in manual toothbrush users. At 3 months, reductions for whole mouth plaque, gingivitis and bleeding were 15%, 16% and 65%, for 3D users and 8%, 12% and 56%, for manual toothbrush users, respectively. Group differences were significant (P < 0.05) in favor of the 3D with respect to plaque reduction for the whole mouth and for interproximal and anterior lingual sites at all three time periods. With respect to gingivitis, reductions for the whole mouth and interproximal and posterior lingual sites at 3 months were significantly greater in the 3D group. There was no clinically significant soft or hard tissue abrasion in either group. In conclusion, the 3D electric toothbrush was found to be safe and had increased efficacy with respect to reduction of plaque and gingivitis compared to a manual toothbrush. PMID- 10530096 TI - A comparison of the efficacy of a novel electric toothbrush and a manual toothbrush in the treatment of gingivitis. AB - PURPOSE: To compare the efficiency of a new electric toothbrush featuring a novel three-dimensional brush head action, with a manual toothbrush, in resolving gingivitis which had been allowed to develop in a group of subjects prior to the treatment phase of the study. MATERIALS AND METHODS: This was a randomized split mouth study. A total of 35 healthy non-dental students refrained from any oral hygiene on the lower jaw for a period of 21 days in order to develop gingivitis. They then brushed one quadrant of the lower jaw with the Braun Oral-B 3D Plaque Remover and the other with a manual toothbrush for a period of 4 weeks. Plaque and gingivitis were evaluated at the start of the study, after the 21 days of no oral hygiene, and after 1, 2, 3 and 4 weeks of brushing twice a day. RESULTS: At the end of the study, the 3D was found to be significantly more effective at reducing bleeding on probing (P < 0.05) for all sites combined and all individual sites. Plaque removal was also significantly more effective with the 3D. Subjects in the study reported that they preferred the 3D to the manual toothbrush and said that it would encourage them to brush for longer. It is concluded that the new Braun Oral-B 3D Plaque Remover offers advantages over a manual toothbrush in terms of plaque control and improvement of gingival condition. PMID- 10530097 TI - A comparison of two electric toothbrushes with respect to plaque removal and subject preference. AB - PURPOSE: To compare the safety and plaque removal efficacy, and subject preference of two electric toothbrushes in a single-blind, randomized, split mouth study. MATERIALS AND METHODS: The devices studied were the Braun Oral-B 3D Plaque Remover and the sonicare electric toothbrush. The 3D toothbrush combines the clinically proven oscillating/rotating action of the D9 with a sonic frequency pulsating action in the direction of the long axis of the bristles, giving a three-dimensional cleaning action. At an initial visit, the subjects (n = 44) were given a baseline examination of the oral soft tissues and a full mouth prophylaxis and instructed in the use of the two products, which was followed by a 4-week training period. After 2 weeks of using each toothbrush on alternate days, brushing technique was checked. After a further 2 weeks, having abstained from oral hygiene for 48 hours, two contralateral quadrants of the mouth were randomly assigned to be cleaned by the subjects with each toothbrush. Plaque assessments and soft tissue examinations were made before and after brushing. Plaque was evaluated according to a refinement of the Modified Navy Plaque Index. At the end of the study, subjects completed a product evaluation questionnaire. RESULTS: Plaque levels were significantly reduced by both toothbrushes (P = 0.001), but the efficacy of the 3D electric toothbrush was significantly greater than that of the sonicare toothbrush (P = 0.001) for all comparisons. Plaque reduction was particularly marked in the interproximal areas, reaching 87% with the 3D product, compared with 68% for sonicare (P = 0.001). In the responses to the questionnaire, the majority of subjects stated that they preferred the 3D toothbrush (88% vs 12%), mainly because of the smaller size of the brush, ease of control and overall maneuverability. PMID- 10530098 TI - A comparison of electric toothbrushes in their potential to cause gingival abrasion of oral soft tissues. AB - PURPOSE: It has previously been established that gingival abrasion associated with the Braun Oral-B Ultra Plaque Remover (D9) is minimal and comparable to that observed with a manual toothbrush. The purpose of this single-use study was to compare gingival abrasion with the D9 and a novel electric toothbrush, the Braun Oral-B 3D Plaque Remover (3D). MATERIALS AND METHODS: 49 subjects (non-dental students) with at least 24 natural teeth and an absence of periodontal probing depths of 5 mm or greater were included in this study. Following a familiarization phase, all subjects received a single prophylaxis and were asked not to brush their teeth for 24 hours prior to their appointment. At this visit, the gums were disclosed by Mira-2-Tone solution and all gingival abrasion on the soft tissues was assessed and recorded. Subjects then brushed their teeth in a random split-mouth order with the two electric toothbrushes, using brush heads of the same design (EB9). The gums were then re-disclosed and gingival abrasions were recorded. They were recorded as either small sites of abrasion (< or = 5 mm) or large sites of abrasion (> 5 mm). RESULTS: The mean number of small traumas increased from 2.57 at baseline to 4.04 after brushing with the D9 and from 1.98 to 4.14 after brushing with the 3D. There was no statistically significant difference between the two groups. For both toothbrushes, more small traumas were found in the upper jaw compared with the lower jaw. No increase in number of large traumas was observed. PMID- 10530099 TI - How does diabetes alter treatment in the dental office? AB - Diabetes mellitus affects approximately seven percent of the American population; thus, patients with diabetes are seen in every dental practice. Advances in medical management of diabetes have resulted in intensification of treatment regimens, with a resulting decrease in long-term complications of the eyes, kidneys, and nervous system. This intensified treatment may place the diabetic patient at increased risk of hypoglycemic emergencies during dental appointments. Dental practitioners should understand methods of preventing hypoglycemia and must be able to recognize and treat hypoglycemia, should it occur in the office. PMID- 10530100 TI - Form follows function: occlusion based rationale for esthetic dentistry. AB - Esthetic dentistry is one of the prime essential areas in dentistry today. The purpose of this article is to address the dependent relationship between excellent esthetics and optimum occlusion. Occlusal objectives must be addressed and achieved. If one is to expect to recreate ideal esthetics, one must first thoroughly investigate, diagnose, and establish an ideal occlusal scheme. The case study within this article revisits the most imperative principle with regards to esthetics and that is: form always follows function. PMID- 10530101 TI - Planning for the future when restoring a hemisected molar: a clinical report. AB - An intracoronal attachment was incorporated into the distal surface of a surveyed complete crown of a tooth anterior to a hemisected molar. This is a recommended procedure when the distal section of a fixed partial denture is supported by a questionable long-term abutment and may require eventual replacement by a removable partial denture when an implant supported prosthesis is not indicated. PMID- 10530102 TI - Myofascial pain syndrome: characteristics, diagnosis, and treatment. AB - Myofascial Pain Syndrome (MPS) is defined as a chronic muscle pain disorder associated with focal areas of exquisite tenderness called trigger points. It is a common cause of chronic orofacial pain and is commonly seen in patients with temporomandibular disorders. When stimulated, trigger points refer pain to surrounding areas, and the pain may resemble other conditions. Diagnosis of MPS is made by identifying trigger points and their associated pain referral pattern. Physical therapy maneuvers and injection are the most common and successful treatments. Eradication of trigger points can diminish or eliminate the pain associated with this syndrome. PMID- 10530103 TI - A resin-modified glass ionomer restorative: three-year clinical results. AB - Resin modified glass ionomer (RMGI) restorative materials have gained popularity in recent years. Their use is most often indicated in Class III and Class V cavities in adults and in numerous applications in children. This popularity in use has taken place in the absence of scientific knowledge of the RMGI materials. Lacking are adequate clinical trials to validate the proposed indications. The purpose of this study was to examine a representative RMGI in cervical abrasions and root caries in adults. Patients were recalled up to three years to evaluate standard clinical criteria. Results found the RMGI to be inferior to conventional composite resin in similar applications. The most noted deficiencies were in color stability and anatomic form, or wear of the RMGI. The results of this prospective clinical trial would suggest a limited longevity for RMGI compared to traditional restorative materials. This study also reinforces the need for evidence of clinical performance prior to making decisions regarding material selections. PMID- 10530104 TI - Dentifrice whitening after professional bleaching. AB - Thirty patients whose teeth had been bleached were given one of two toothpastes. One half of the group brushed with a toothpaste containing 10% carbamide peroxide (DW) and the other half with a toothpaste containing 3% hydrogen peroxide (HP) for three months. Color change from baseline was evaluated after 4 weeks and again after 12 weeks, using a shade guide and measured with a colorimeter at baseline. The DW toothpaste was able to stabilize the tooth-lightening effect of the bleaching gel better than the HP toothpaste. The HP product had a lower tooth sensitivity rating. PMID- 10530105 TI - Indirect implant provisionalization tests esthetics, comfort. PMID- 10530106 TI - Techniques of past served well by new prosthodontic technology, materials. AB - During the past 15 years, advances in laboratory technology, prosthetic teeth, and materials have provided the practicing clinical dentist with many new options for the treatment of the removable prosthodontic patient. Some of these advances which allow improved quality of care for the patient as well as increased convenience for the dentist and technician will be presented. Additionally, some of the research available on these products will be discussed in an attempt to aid the practitioner in exploring the option of incorporating these advances into their clinical practice. PMID- 10530107 TI - Ceramic materials more durable, suitable for fixed prosthetics. AB - The practice of dentistry progressed with the recent developments and innovations in dental materials. The clinicians need to be familiar with these materials to successfully incorporate them in their practices. Fixed prosthodontics has assumed a growing role in the practice of restorative dentistry since the application of the lost wax casting technique. With the increased life expectancy, a greater demand of fixed prosthodontics is expected in the future. Recent material and technical and clinical advances have made treatment planing and decision-making more complex; most have attempted to improve current techniques to achieve the ideal pleasant smile. Intelligently planned, well documented, evidence-based research has provided a reliable foundation to many of these advances affecting the practice of fixed prosthodontics. This article will attempt to focus on some of these clinically relevant and applicable developments and briefly review the current literature and encourage pratictioners to review subsequent literature. PMID- 10530108 TI - Instrumentation enhances today's endodontic care. AB - The field of endodontics has virtually exploded in the last several years with technological advances and improvements in many areas. The largest change has occurred with the introduction of nickel-titanium rotary instrumentation, warm gutta-percha obturation, microscopes, and digital imaging. Prominent new devices and instruments are presented with a brief overview of items listed with a source. PMID- 10530109 TI - Compomers, reattachment method expand restoration capabilities. AB - The purpose of this article is to review one new material and one new technique being used in restorative dentistry today. Compomers, new fluoride-releasing resin restorative materials, are compared to conventional glass ionomers in terms of classification, physical properties, and clinical usage. Compomers are not true glass ionomer materials since the acid/base setting reaction, charactheristic of conventional glass ionomers, does not occur. As a consequence, their physical properties of translucency, coefficient of thermal expansion, and strength more closely resemble composite resins than conventional glass ionomers. These differences in physical properties have clinical implications in their usage. In terms of new techniques, clinical and laboratory data now exist to support the method of reattachment of fractured tooth fragments using only dentin bonding agents, in cases where the tooth fragment is available. This method can restore up to 50 percent of the original strength of intact teeth. The technique advocates the use of acid etching and enamel and dentin bonding, without any tooth preparation. In vitro studies have achieved total (100 percent) restoration of intact teeth by bonding a porcelain veneer to the tooth after the reattachment. PMID- 10530111 TI - The etiology of altered sensation in the inferior alveolar, lingual, and mental nerves as a result of dental treatment. AB - In a review of 163 consecutive patients referred with trigeminal nerve (inferior alveolar or lingual nerve) involvement following dental treatment, the most common etiology was third-molar removal (87 patients). The second most common cause was an inferior alveolar nerve block injection (34 patients), with a smaller number of endodontic and periodontal complications. Female patients outnumbered male 3.3 to 1. Twenty-seven patients were offered surgical exploration and possible nerve repair surgery; of them, 14 underwent surgery. Forty percent of the patients admitted to being involved in litigation during the time they were undergoing treatment. PMID- 10530110 TI - Safety needles. New requirements of the Occupational Safety and Health Administration bloodborne pathogens rule. AB - In September 1998, a California Assembly bill was signed into law that requires significant changes to the Cal/OSHA Bloodborne Pathogens Standard. As of July 1, 1999, all health care employers in the state must begin providing sharps safety devices. For dentistry, this means a shift from the traditional needles to safety needles with engineered built-in safety mechanisms. Some exceptions are provided in this new regulatory change. There is no reliable data on the safety and efficacy of the available devices. This article explores the regulatory changes and begins to provide information on the devices available. Design features, usability by the practitioner, and safety to the patient are important issues to consider when deciding whether these devices are appropriate for dental anesthesia. Most practitioners will find it difficult to conduct an independent evaluation and must rely on information in the professional literature to help guide their decisions. PMID- 10530112 TI - Cost-effectiveness model for prevention of early childhood caries. AB - This study presents and illustrates a model that determines the cost effectiveness of three successively more complete levels of preventive intervention (minimal, intermediate, and comprehensive) in treating dental caries in disadvantaged children up to 6 years of age. Using existing data on the costs of early childhood caries (ECC), the authors estimated the probable cost effectiveness of each of the three preventive intervention levels by comparing treatment costs to prevention costs as applied to a typical low-income California child for five years. They found that, in general, prevention becomes cost-saving if at least 59 percent of carious lesions receive restorative treatment. Assuming an average restoration cost of $112 per surface, the model predicts cost savings of $66 to $73 in preventing a one-surface, carious lesion. Thus, all three levels of preventive intervention should be relatively cost-effective. Comprehensive intervention would provide the greatest oral health benefit; however, because more children would receive reparative care, overall program costs would rise even as per-child treatment costs decline. PMID- 10530113 TI - Periodontal plastic surgery. AB - As the demand for esthetic dentistry has increased, dentistry has developed techniques to meet this demand. Periodontal plastic surgery has been part of this effort. This article outlines the scope of periodontal plastic surgery procedures to aid the dental team in diagnosis and treatment of esthetic dental cases. PMID- 10530114 TI - Routine prophylactic antibiotic use in diabetic dental patients. AB - There is no scientific evidence in the literature to support the premise that well-controlled, or even moderately well-controlled, nonketotic diabetic patients are prone to infection when undergoing uncomplicated dentoalveolar surgery. Routine administration of prophylactic antibiotics should be considered only in situations where prophylactic antimicrobials would be used for a nondiabetic patient. Poorly controlled diabetics (whether Type I or II), with fasting glucose levels above 250 mg/dL, should be referred for improved control of their blood sugar before non-emergency surgery is performed. If emergency surgery is needed for a poorly controlled patient, then prophylactic antibiotics are prudent, using the accepted principles of such use. Infections in diabetic patients, regardless of their control levels, should be managed aggressively, including possible early referral to oral and maxillofacial surgeons. PMID- 10530115 TI - Two approaches to the diagnosis of lesions of the oral mucosa. AB - This article describes two approaches to the classification of oral mucosal lesions. One is based on the etiopathogenesis of the lesion and the second on the clinical appearance. These two approaches are compared and contrasted, and their integration is described. Combining these two classification schemas allows an excellent understanding of the various lesions so than an expeditious and correct diagnosis can result. Appropriate management and treatment can then follow. PMID- 10530116 TI - Using risk assessment to customize periodontal treatment. AB - In recent years, understanding of the multifactorial nature of periodontal disease has taken great strides. Periodontal disease is initiated and sustained by the presence of bacteria, but disease progression is significantly modified by the body's response to the bacteria. This article highlights the emerging evidence regarding which risk factors are predominant in influencing the disease process and how the incorporation of prognostic risk factors in overall diagnosis can help facilitate treatment planning. These factors appear to be smoking, genetic susceptibility, compliance, and diabetes. The first three factors mentioned are the focus of this article. Each is discussed with regard to their role in amplifying the disease process and how this information can be used in clinical practice. By acknowledging the importance of these factors, dentists can consider their patients' risk to allow for more cost-effective planning and treatment. The opportunity to identify high-risk patients and treat them more proactively is significant; the challenge rests with dentists' willingness and ability to embrace the change before them. PMID- 10530117 TI - Periodontal disease and preterm low birth weight deliveries. PMID- 10530118 TI - Silicoating in resin-metal bonding: restorative implications for osseointegrated implant-bone unison. AB - Detailed background information leading to the development of a silicoating technique is presented with its advantages and disadvantages and its role in implant dentistry. The rationale is shown for coining a new acronym "osseointegrated Implant-Bone Unison" with an interest in biomechanical considerations for a prosthodontic restorative scenario. Several clinical cases are presented that involve silicoating and composite restorations. PMID- 10530119 TI - Fluoride in tea--its dental significance: a review. AB - It has long been accepted that fluoride accumulates in the leaves of the tea plant. Camellia sinensis. In addition it is known that some of this fluoride is released into the infusion which is drunk as tea. The exact concentration of fluoride in a cup of tea and the effects of this fluoride have been the subject of many international studies. This review summarizes the main points of such studies which have been carried out in an attempt to establish the dental significance of fluoride in tea. The most popular teas in Ireland are not readily available in any other country and therefore, Irish data may not be assumed to be similar to those in the studies reviewed here. By identifying potential sources of high fluoride ingestion, recommendations can be made to reduce consumption from these sources in patients who may be at risk of dental fluorosis. In conclusion, it is recommended that a research project be carried out to analyse the fluoride levels released and the rates of the releases from teas available on the Irish market. Ireland has the highest per capita consumption of tea in the world. PMID- 10530120 TI - Compounding inequalities in the oral health of older women living outside Dublin. AB - BACKGROUND: The Irish population is ageing with an increase in the absolute number of older people. However, there is a deficiency of information on the oral health status of older people in the Republic of Ireland. Traditionally, the primary measure of oral health status in older populations has been the prevalence of edentulousness. The aim of this retrospective study was to obtain information on the edentulous status of older people and to investigate the compounding effects of age, gender and area-of-residency on the prevalence of edentulism, using as a sample those over 65 year-olds who registered at dental hospitals in the Republic of Ireland during 1995. DESIGN: All patient records from both dental hospitals in the Republic were identified on electronic databases, and information was collected over a period of six months. Of the 566 patients who registered at the dental hospitals in 1995, it was possible to access 524 (93 per cent) records. RESULTS: The edentulous rate in the sample was 42 per cent, but significant variations were apparent with age, gender and geographical residency. In addition, there was evidence of compounding inequalities in the prevalence of edentulism when the combined effects of age, gender and area of residency were studied. Considerable inequalities were observed; at age 65 women resident outside Dublin were 1.65 times more likely (or 65 per cent more likely) to be edentulous than men resident in Dublin of the same age. At age 75, women were 1.46 times more likely to be edentulous than men. PMID- 10530121 TI - Making the right moves. A strategic look at office ethics. PMID- 10530122 TI - Survey examines patients' fear of dental treatment. AB - From September through November 1996, 158 of 844 patients at the general dentistry clinic of Tufts University School of Dental Medicine completed surveys concerning their fear of dental treatment. High levels of dental fear affected 65 percent of the respondents, with patients under the age of 45 reporting higher levels of fear than patients ages 45 years and older. Results showed that the four most common causes of fear in patients occur when the dentist seems rushed (65 percent), when the patient feels uninformed (50 percent), when the patient worries if the local anesthetic will be effective (43 percent), and when the patient's feelings are neglected (40 percent). PMID- 10530123 TI - Tooth extraction as a reconstructive event. PMID- 10530124 TI - Improve dental care at home with a powered toothbrush. PMID- 10530125 TI - Water fluoridation in Massachusetts: a thirty-year review. AB - By the end of 1997, only 121 communities (59 percent) of the population in Massachusetts were receiving fluoridated water serving 3,523,615 people. Eleven communities, three of which are naturally fluoridated, are partially fluoridated communities. According to the 1992 Fluoridation Census. Massachusetts was ranked 35th in the nation by percentage of its population living in fluoridated communities. From 1968 to 1997, there were 135 fluoridation orders by 112 communities, of which 67 (49.6 percent) had binding referenda, with 30 (45 percent) winning and 37 (55 percent) losing the vote (one community had one order and two referenda due to a court decision). Eventually, 78 (58 percent) of the 135 orders resulted in fluoridation being implemented. The average length of time from the order of fluoridation to its implementation was seven years, with a range of less than one year to 29 years. From 1968 to 1977, there were 91 fluoridation orders as compared to only eight from 1988 to 1997. Fluoridation is still the most cost-effective preventive measure for dental disease and needs to be promoted once again in Massachusetts. PMID- 10530126 TI - Improving clinical dentistry with six low-cost devices. PMID- 10530127 TI - A clinico-pathologic presentation. Metastatic melanoma. PMID- 10530128 TI - Evaluation of filling materials in membrane--protected bone defects. A comparative histomorphometric study in the mandible of miniature pigs. AB - In recent years, bone grafts and bone substitutes have been increasingly utilized underneath barrier membranes to optimize the treatment outcome of bone reconstructive therapy for defects in the alveolar process. In the present study, 4 different filling materials were evaluated in bone defects of similar dimensions in the mandible of miniature pigs. Blood clots and autografts were used as controls. The defects were covered with barrier membranes and allowed to heal for 4, 12 or 24 weeks. Histologic examination demonstrated that bone repair progressed through a programmed sequence of maturation steps closely resembling the pattern of bone development and growth regardless of whether bone grafts or substitutes were present or not. Histomorphometric analysis showed that autologous bone grafts (autografts) had the best osteoconductive properties during the initial healing period, with 39% of newly formed bone inside the membrane-covered defects at 4 weeks of healing. In addition, 87% of the graft surfaces were already covered by bone at this time. Both values were significantly higher for autografts than for the 4 alternative bone fillers (P < or = 0.05). At 12 weeks, these differences were no longer apparent, with all 5 filling materials showing similar values. Among the tested bone substitutes, tricalcium phosphate (TCP) showed a significantly higher percentage of bone fill at 24 weeks of healing. It can be concluded that sites filled with autografts clearly demonstrated the best results underneath barrier membranes in the early phase of healing. As far as degradation and substitution are concerned, TCP showed the most promising results. This filler, however, needs to be tested further in a more demanding animal model. Less favorable results were obtained for coral-derived hydroxyapatite granules and for demineralized freeze-dried bone allografts. PMID- 10530129 TI - The effect of a deproteinized bovine bone mineral on bone regeneration around titanium dental implants. AB - The aim of the present experiment was to test the effect of a deproteinized bovine bone mineral (Bio-Oss) on guided bone regeneration (GBR) in dehiscence defects around implants. The first 2 molars and all premolars were extracted on both sides of the mandibles of 3 monkeys (Macaca fascicularis). Three months later, 2 titanium plasma-coated cylindrical implants were placed in all quadrants of each monkey. During the surgical procedure, standardized dehiscence defects were produced buccally and lingually, measuring 2.5 mm in width and 3 mm in height. Four different experimental situations were created: 2 sites in each monkey were covered with an ePTFE membrane (M), 2 were filled with the graft material (DBBM), 2 were filled with the graft material and also covered with a membrane (M + DBBM), and 2 control sites were neither grafted nor covered (C). The flaps were sutured to allow for primary healing. Linear measurements of bone height and width were calculated on histological specimens obtained 6 months following surgery. In addition, values for bone density and for surface fraction of graft to new bone contact were measured. Vertical bone growth along the implant surface of 100% (SD 0%) for M + DBBM, 91% (SD 9%) for M, 52% (SD 24%) for DBBM, and 42% (SD 35%) for C was measured. The width of the regenerated bone 1.5 mm above the bottom of the original defect, i.e. at the 50% mark of the vertical extension of the defect, in relation to the width at the bottom of the defect amounted to 97% (SD 2%) for M + DBBM, 85% (SD 9%) for M, 42% (SD 41%) for DBBM, and 23% (SD 31%) for C. Assessment of bone density within the confinement of the regenerated bone resulted in an increase of 30% (SD 11%) for M + DBBM, 45% (SD 20%) for M, 33% (SD 20%) for DBBM, and 22% (SD 23%) for C. The values for graft to new bone contact within this compartment amounted to 80% (SD 15%) for M + DBBM and 89% (SD 14%) for DBBM. In conclusion, Bio-Oss exhibited osteoconductive properties and hence can be recommended for GBR procedures in dehiscence defects with respect to vertical and horizontal growth of bone. PMID- 10530130 TI - Unbiased stereological methods used for the quantitative evaluation of guided bone regeneration. AB - The present study describes the use of unbiased stereological methods for the quantitative evaluation of the amount of regenerated bone. Using the principle of guided bone regeneration the amount of regenerated bone after placement of degradable or non-degradable membranes covering defects in rabbit calvaria was compared. Forty rabbits were divided into 5 groups. A titanium microplate was placed over the defect to prevent collapse of the membrane. The non-degradable expanded polytetrafluoroethylene membrane and the degradable Polyglactin 910 material were both placed unicortically and bicortically. Undecalcified sections were prepared for stereologic evaluation after an observation period of 8 weeks. Complete bone healing of the defects was not observed in any of the specimens. Unbiased stereologic estimates revealed 48% bone regeneration in defects covered by 2 ePTFE membranes, and 12% in defects covered by 2 Polyglactin 910 membranes. Defects covered by 1 ePTFE or 1 Polyglactin 910 membranes revealed 10% or 18% bone regeneration, respectively. The control group regenerated 14%. The major difference of the estimates was caused by real difference between specimens, i.e. biologic variation, whereas only minimal variance was added by the stereologic estimation procedure. PMID- 10530131 TI - A 5-year randomized clinical trial on the influence of splinted and unsplinted oral implants in the mandibular overdenture therapy. Part I: Peri-implant outcome. AB - Thirty-six completely edentulous patients were enrolled for a 5-year prospective study testing the treatment outcome between splinted and unsplinted implants retaining a mandibular hinging overdenture. The patients were randomized into 3 groups of equal size depending on the attachment system used such as: magnets, ball attachments or bars (reference group). Only 1 implant out of the 72 had failed at the abutment stage. Not a single implant failed during the 5-year loading period. The accumulation of plaque was significantly higher for the Magnet than for the Ball group. Bleeding on probing, as well as marginal bone level, attachment level and Periotest values did not statistically differ among the groups, neither at year 1 nor at year 5. However, the Periotest values were significantly lower at year 5 compared to year 1 for all groups, which indicates a higher rigidity at the bone-implant interface. No correlation was found between bleeding on probing and marginal bone loss. We conclude that the connection state of 2 implants retaining a hinging overdenture did not influence the peri-implant outcome. PMID- 10530132 TI - The EsthetiCone abutment: three-year results of a prospective multicenter investigation. AB - The EsthetiCone abutment, designed for subgingival restorative margins, makes it possible to achieve ideal esthetics, function, and predictability. This study included 200 EsthetiCone abutments placed in 50 partial or complete edentulous patients participating in a multicenter study performed in Europe and the United States. The patients were followed for 3 years and the marginal bone level of the implants was determined from intra-oral radiographs. During the follow-up period, a total of 9 of patients were withdrawn from the study. Based on the remaining patients, a total of 164 EsthetiCone abutments (82%) were evaluated through the 3 year follow-up visit. Only 1 EsthetiCone abutment failed during the study period, resulting in a cumulative success rate of 99.2% in the maxilla and 100% in the mandible after 3 years. No bone loss due to the usage of short abutment collars (1 mm) was seen in this study. The marginal bone resorption during the 3-year period did not exceed 1 mm as a mean neither for the 1 mm nor the higher abutments when analyzed separately. During the follow-up period, few complications were reported and they were all easily rectified without compromising the treatment success. The outcome of this study indicates that safe long-term and esthetically good results can be achieved when the Branemark implants are loaded with EsthetiCone abutments. The reported need of a 3 mm peri implant sulcus with an animal model was not confirmed with patients in this study. PMID- 10530133 TI - An XPS and SEM evaluation of six chemical and physical techniques for cleaning of contaminated titanium implants. AB - The purpose of the present study was to analyse clinically failed and retrieved implants prior to and after cleaning by means of scanning electron microscopy (SEM) and X-ray induced photoelectron spectroscopy (XPS) as compared to unused controls. Six different chemical and physical techniques for cleaning of contaminated titanium implants were evaluated: 1) rinsing in absolute ethanol for 10 min, 2) cleaning in ultrasonic baths containing trichloroethylene (TRI) and absolute ethanol, 10 min in each solution, 3) abrasive cleaning for 30 s, 4) cleaning in supersaturated citric acid for 30 s, 5) cleaning with continuous CO2 laser in dry conditions at 5 W for 10 s, 6) cleaning with continuous CO2-laser in wet conditions (saline) at 5 W for 10 s. SEM of failed implants showed the presence of contaminants of varying sizes and XPS showed almost no titanium but high carbon signals. XPS of unused titanium implants showed lower levels of titanium as previously reported, probably due to contamination of carbon which increased with time in room air. Cleaning of used implants in citric acid followed by rinsing with deionized water for 5 min followed by cleaning in ultrasonic baths with TRI and absolute ethanol gave the best results with regard to macroscopical appearance and surface composition. However, as compared to the unused implants the results from an element composition point of view were still unsatisfactory. It is concluded that further development and testing of techniques for cleaning of organically contaminated titanium is needed. PMID- 10530134 TI - Attachment and proliferation of human oral fibroblasts to titanium surfaces blasted with TiO2 particles. A scanning electron microscopic and histomorphometric analysis. AB - The purpose of this study was to determine the effect of c.p. titanium surfaces blasted with TiO2 particles on the biological responses of human gingival fibroblasts (HGF). Fibroblast morphology and attachment were investigated on turned (control) titanium surfaces and those blasted with 45 microns (standard), 45-63 microns, and 63-90 microns TiO2 particles. The specimens were analyzed using a confocal laser scanner and SEM. The cell profile areas were measured using a semiautomatic interactive image analyser. The figures were expressed as percent of attachment. The turned samples had the smoothest surfaces and the roughest were those blasted with 63-90 microns. All TiO2 blasted specimens had homogeneous surfaces. Cells appeared to flatten, spread and form cellular bridges with the adjacent cells. Fibroblasts on the turned titanium surfaces appeared to follow the direction of the fine irregularities on the surface but tended to spread haphazardly on the blasted surfaces. The attachment assays showed no significant difference in the percentage of fibroblast cell attachment on the standard surfaces compared to the turned surfaces. Both surfaces blasted with 45 63 microns or 63-90 microns had significantly (P < 0.05) lower percentages of cell attachment than the control. The surfaces blasted with 63-90 microns particles had the lowest rate of cell attachment. A significant correlation (P < 0.01) was found between the degree of particle size and attachment of fibroblasts after 1-72 h. It is concluded that surface micro-texture influences the attachment and growth of HGF: surfaces blasted with 45 microns TiO2 do not inhibit fibroblast attachment and smooth or finely grooved surfaces could be conducive to cellular attachment. PMID- 10530135 TI - HA-coated root-form implants--is there cause for concern? PMID- 10530136 TI - A new implant impression technique for prosthodontic accuracy. PMID- 10530137 TI - Implant system based on the quality of bone. PMID- 10530138 TI - Use of an acellular dermal allograft for soft-tissue augmentation. PMID- 10530139 TI - Bioactive ceramics in implant, grafting, and periodontal treatment. PMID- 10530140 TI - Combination procedure for augmenting the maxillary ridge for implant placement. PMID- 10530142 TI - Laboratory principles of implant rehabilitation for fixed prosthesis. PMID- 10530141 TI - The use of a bone harvest system for autogenous bone grafts during implant procedures. PMID- 10530143 TI - Immediate placement of wide-diameter implants in the premaxilla. PMID- 10530144 TI - Treatment planning and management of implant position for reconstructive success. PMID- 10530145 TI - The angled implant as a single-stage implant alternative. PMID- 10530146 TI - Resorbable or non-resorbable membranes: where, why, and how. PMID- 10530147 TI - Subtleties in the uses of a non-resorbable membrane with select implant surgeries. PMID- 10530149 TI - Review of membranes used for guided tissue regeneration. PMID- 10530148 TI - Surface roughness of intraoral dental implant fixtures. PMID- 10530150 TI - Brain abscesses caused by oral infection. AB - Brain abscesses are rare but can be life-threatening infections. Recent progress in microbiological classification and identification has indicated that they are sometimes caused by oral infection and dental treatment. It has been postulated that oral microorganisms may enter the cranium by several pathways: 1) by direct extension, 2) by hematogenous spread, 3) by local lymphatics, and 4) indirectly, by extraoral odontogenic infection. In the direct extension, oral infections spread along the fascial planes. Hematogenous spreading occurs along the facial, angular, ophthalmic, or other veins which lack valves, through the cavernous sinus and into the cranium. Another hematogenous pathway is through the general circulation. Oral bacteria may cause systemic infections, e.g., endocarditis, and then indirectly initiate brain abscess. Microbiota, complications, and the prevention and management of odontogenic brain abscesses are also discussed in this review. PMID- 10530151 TI - beta-Hemolytic streptococci and other beta-hemolytic organisms in apical periodontitis and severe marginal periodontitis. AB - Thirteen teeth with necrotic pulps and apical periodontitis and nine severe periodontal pockets were cultured for presence of beta-hemolytic streptococci and other beta-hemolytic organisms. Samples were dispersed and plated on two non selective and one selective growth media and incubated anaerobically and in 10% CO2 in air. A total of 59 beta-hemolytic colonies were purified and identified. Eight beta-hemolytic streptococcal isolates were obtained from three of the severe marginal periodontitis sites. All were identified as belonging to the Streptococcus sanguis group. No beta-hemolytic streptococci were detected in apical periodontitis samples. Twenty obligately anaerobic isolates were detected, all of which were known periodontal and endodontic pathogens. Isolates from apical periodontitis sites were identified as Propionibacterium acnes, Actinomyces naeslundii, Actionomyces odontolyticus and Peptostreptococcus micros, while severe marginal periodontal sites contained the same species with the addition of Actinomyces viscosus and Actinomyces meyeri. Of 19 staphylococci and micrococci, Staphylococcus epidermidis was the predominant isolate in both apical periodontitis and severe marginal periodontitis sites. However, less commonly known organisms such as Staphylococcus cohnii and Micrococcus sp. were identified in severe marginal periodontitis sites. The isolation of Bacillus sp. (12 isolates) in one severe marginal periodontitis and two apical periodontitis subjects was especially interesting, warranting consideration of this organism as a legitimate isolate and potential pathogen in oral disease. PMID- 10530152 TI - Endotoxin activity measured by limulus assay. AB - The binding activity of endotoxin (Escherichia coli 0111) to dentin powder pre treated with 10% sodium hypochlorite and with 3% hydrogen peroxide was investigated. This was carried out for periods of 30, 10 and 1 min. The endotoxin was diluted into 100 ng/ml solutions. Dentin powder suspended in a 34 micrograms/ml protein concentration was used in the study. The limulus amoebocyte lysate test was used to determine the amount of endotoxin binding, which was measured by the microtiter plate reader. The binding activity level of that concentration of endotoxin was significantly lower in dentin powder treated with 10% sodium hypochlorite than in that treated with 3% hydrogen peroxide at 1 and 10 min. Endotoxin binding activity measured the lowest in the 30 min test period. PMID- 10530153 TI - Drying and rewetting anterior crown fragments prior to bonding. AB - Fractured anterior teeth can be restored by adhesive bonding of the fragment to the remaining tooth structure. This in vitro study describes the effect on fracture strength of fragments dried and rewetted for various periods of time prior to bonding. Seventy central incisors from sheep were fractured. The resulting incisal crown fragments were then stored in air at room temperature at ambient humidity (70 +/- 16%) for 5 s, 30 min, 1 h, 3 h, 6 h, 12 h, or 24 h. The apical parts of the fractured teeth were stored in water. After storage in air each fragment was then bonded to the matching apical tooth structure with a bonding agent and a low-viscosity composite resin. After water storage for 2 days, mean fracture strength was measured. Another group of teeth comprising 40 sheep central incisors was fractured and the fragments were stored in air at room temperature for 24 h as above. The fragments were then immersed in water for 10 min, 1 h, 1 day, or 7 days, prior to bonding and measurement as described above. Statistical analysis revealed that the fracture strength of the fragment-bonded teeth was unaffected by air storage of the fragment for up to 1 h prior to bonding, after which additional drying resulted in decreased fracture strength. Fragments dried for 24 h in air and rewetted by immersion in water for at least 1 day were fragment-bonded without loss of fracture strength. PMID- 10530154 TI - Epidemiology of traumatic injuries to the permanent incisors of 9-12-year-old schoolchildren in Damascus, Syria. AB - This cross-sectional survey was carried out to assess epidemiological data concerning dental injuries to the permanent incisors of Syrian children. It included 1087 children aged 9 to 12 years, of both sexes, randomly selected from public and private primary schools in Damascus. The response rate was 100%. The prevalence of traumatic injuries to the permanent incisors rose from 5.2% at the age of 9 years to 11.7% at the age of 12 years (P = 0.007). The difference in prevalence between boys and girls was not statistically significant (P > 0.05). The majority (59.8%) of children who had experienced injuries to the permanent incisors reported that they were not taken to the dentist for evaluation or treatment of the damage. Among those children who had experienced traumatic injuries to the teeth 93.1% presented with untreated damage. Because some injuries were minor, such as small enamel fractures, the proportion of children who needed treatment was 63.2%. There was a tendency for children with an incisal overjet greater than 5 mm to have experienced dental injuries (P = 0.06). Children with inadequate lip coverage were more likely to have experienced dental injuries than those with adequate lip coverage (P = 0.000). The most common reported cause of injuries to the permanent incisors was violence (42.5%), followed by traffic accidents (24.1%), collisions with people or inanimate objects (16.0%) and falls (9.1%). In conclusion, traumatic dental injury may pose a serious dental public health problem. PMID- 10530155 TI - Coronal leakage along apical root fillings after immediate and delayed post space preparation. AB - The seal provided by a full-length root canal filling may be compromised by post space preparation. Eighty human mandibular premolars each with a single canal were obturated with laterally condensed gutta-percha cones and a sealer. Immediate post space preparation was carried out on half the number of teeth and delayed post space preparation on the remaining 40 teeth. Leakage along the apical root fillings was determined using a fluid transport device under a head space pressure of 30 kPa (0.3 atm). More leakage was found after delayed preparation than after immediate preparation (P = 0.0059). PMID- 10530156 TI - Evaluation of the ability of thermal and electrical tests to register pulp vitality. AB - The aim of the present study was to evaluate the ability of thermal and electrical tests to register pulp vitality. Sensitivity, specificity, negative predictive value and positive predictive value were calculated by comparing the test results with a "gold standard". The thermal tests studied were a cold test (ethyl chloride) and a heat test (hot gutta-percha). For the electrical test, the Analytic Technology Pulp Tester was used. The examined teeth were 59 teeth with unknown pulpal status in need of endodontic treatment and 16 intact teeth, all with radiographically normal periapical bone structures. In total 46 teeth with vital pulps and 29 teeth with necrotic pulps were tested. This gave a disease prevalence of 39%. The gold standard was established by direct pulp inspection of the 59 teeth in need of endodontic treatment. In the 16 intact teeth the pulp was judged as vital. The number of true positive (TP), false positive (FP), true negative (TN) and false negative (FN) test results was calculated for each method as compared to the gold standard. Based on this, the sensitivity, specificity, positive predictive value and negative predictive value were calculated for each method. The sensitivity was 0.83 for the cold test, 0.86 for the heat test and 0.72 for the electrical test. The specificity was 0.93 for the cold test, 0.41 for the heat test and 0.93 for the electrical test. The positive predictive value was 0.89 for the cold test, 0.48 for the heat test and 0.88 for the electrical test, and the negative predictive value was 0.90 for the cold test, 0.83 for the heat test and 0.84 for the electrical test. This indicated that the probability of a non-sensitive reaction representing a necrotic pulp was 89% with the cold test, 48% with the heat test and 88% with the electrical test. It also indicated that the probability of a sensitive reaction representing a vital pulp was 90% with the cold test, 83% with the heat test and 84% with the electrical test. PMID- 10530158 TI - Replantation of avulsed central incisor with advanced periodontal disease: a case report. AB - This paper describes the case of a 31-year-old woman with advanced periodontal disease who lost a tooth due to trauma. The avulsed tooth had minimal bony support of only 4-5 mm. The patient described was under good periodontal maintenance. The tooth was kept moist, and replantation occurred within an hour of avulsion. The tooth was returned to its position, splinted, and later endodontically treated. After 2 years the tooth appears and functions normally as it did before avulsion. PMID- 10530157 TI - Treatment of root perforation by intentional reimplantation: a case report. AB - Intentional reimplantation is defined as a procedure in which an intentional tooth extraction is performed followed by reinsertion of the extracted tooth into its own alveolus. In this paper, intentional reimplantation is described and discussed as a treatment approach to root canal instrument separation in conjunction with root perforation. An 8-year follow-up case report is presented. The reimplanted tooth is now a fixed bridge abutment. Although successful in this case, the intentional reimplantation procedure should be considered a treatment of last resort, that is, when another treatment option is not viable for the treatment of root perforation/instrument retrieval. PMID- 10530159 TI - Management of an avulsed primary incisor. AB - The case describes the management of an avulsed maxillary central primary incisor of a 3 1/2-year-old girl. The tooth was retained in the oral cavity for 30 min. After replantation it was splinted for 17 days. At day 11 the root canal was completely instrumented and obturated with a calcium hydroxide paste. The 1-year follow-up documented no pathologic clinical or radiographic findings. One and a half years after the trauma the tooth was extracted since a fistula and extensive external inflammatory resorption had developed. The permanent successor erupted along with its neighboring central incisor without any complications 6 months later. Conventional approaches for treating avulsed permanent teeth could also be applied to avulsed primary incisors to preserve them for a certain period without the additional risk of damaging their developing permanent successors. PMID- 10530160 TI - The need to introduce gene therapy to the dental curriculum. AB - Recombinant DNA technology is finding its way into many aspects of clinical medicine. No application currently is more dramatic than gene therapy. Proofs of principle have already been established for gene therapy targeted to oral tissues, and more are likely to be demonstrated in the near future. The dental curriculum must begin to include the biological basis of DNA-based therapies and other related biomedical science progress. PMID- 10530161 TI - The effect of surgical technique on lingual nerve damage during lower 3rd molar removal by dental students. AB - We have previously shown that avoidance of lingual flap retraction with a Howarth periosteal elevator during lower 3rd molar removal, reduces the incidence of lingual nerve damage. In that study, the surgery was undertaken by qualified staff and we have now assessed the effect of revising the method taught to our junior undergraduate dental students. We evaluated the outcome of surgery undertaken by 2 consecutive years of students, each group being taught 1 of the 2 methods. A total of 200 patients requiring lower 3rd molar removal under local anaesthesia were included in the study. In year 1, the surgery included elevation of a lingual flap and insertion of a Howarth elevator adjacent to the lingual plate; in year 2 this part of the procedure was avoided by using a purely buccal approach. There were no significant differences between the levels of tooth eruption and types of impaction of the teeth removed in each year. Lingual sensory disturbance occurred in 3 patients in the 'flap' group (3.3%) and in 1 patient (0.9%) in the 'no flap' group. As this incidence is not significantly different in the 2 groups (P < 0.4), we conclude that avoidance of lingual retraction by students undertaking lower 3rd molar removal does not appear to place the lingual nerve at greater risk. In view of the results of our previous study, we therefore advocate this method for use in undergraduate dental education. PMID- 10530162 TI - A survey of the computer literacy of undergraduate dental students at a University Dental School in Ireland during the academic year 1997-98. AB - As dental practice management becomes more computer-based, the efficient functioning of the dentist will become dependent on adequate computer literacy. A survey has been carried out into the computer literacy of a cohort of 140 undergraduate dental students at a University Dental School in Ireland (years 1 5), in the academic year 1997-98. Aspects investigated by anonymous questionnaire were: (1) keyboard skills; (2) computer skills; (3) access to computer facilities; (4) software competencies and (5) use of medical library computer facilities. The students are relatively unfamiliar with basic computer hardware and software: 51.1% considered their expertise with computers as "poor"; 34.3% had taken a formal typewriting or computer keyboarding course; 7.9% had taken a formal computer course at university level and 67.2% were without access to computer facilities at their term-time residences. A majority of students had never used either word-processing, spreadsheet, or graphics programs. Programs relating to "informatics" were more popular, such as literature searching, accessing the Internet and the use of e-mail which represent the major use of the computers in the medical library. The lack of experience with computers may be addressed by including suitable computing courses at the secondary level (age 13 18 years) and/or tertiary level (FE/HE) education programmes. Such training may promote greater use of generic softwares, particularly in the library, with a more electronic-based approach to data handling. PMID- 10530163 TI - A survey of the IT skills and attitudes of final year dental students at Bristol University in 1996 and 1997. AB - Surveys of final year dental students were conducted in 1996 and 1997 to see if there were any detectable differences in students' perception of their own information technology IT skills and attitudes towards information technology following the opening of a dedicated computer-assisted-learning (CAL) room. An increase was seen in students' confidence levels, with fewer assessing themselves as IT "beginners" (1996 = 36%; 1997 = 14%), and more assessing themselves as competent in some basic skills (52%; 41%). Although more students were found to be using basic computer facilities (word processing, email, the World Wide Web), there was little difference in attitudes towards these packages between the 2 years. There was a significant increase in the number of students agreeing or strongly agreeing with the statement "the use of IT had added value to this course for me" (1996 = 39%; 1997 = 67%). The main obstacle which was identified by students as a barrier to using IT was the lack of adequate training. Over half the students in both years felt that insufficient training had been provided to enable them to cope with the course without difficulty. PMID- 10530164 TI - Education in periodontology. A need for a new teaching model. AB - The objectives of the present work were to elucidate faculty perceptions and effects on traditional teaching and the supplement of a problem-based learning model in periodontal education. Students and faculty members at one university were asked to respond to a battery of relevant questions in periodontology that had been previously discussed at two European academic workshops. Differences in responses were noticed both between faculty, students, and their responses as compared to responses given by participants at the European workshops. On several topics, faculty within the selected university held significantly different opinions. Thus lecture content and concepts may vary dependent on who gives the lecture and may not be consistent with the common interpretation of the scientific evidence. Responses given by those participating in the workshops indicated consensus. The study results suggest that students were able to extract appropriate conclusions from the scientific literature in the they were able to concede with an expert panel on specific issues studied. Students were however unable to convince other students, who had not participated in the specific activity, that their conclusions were consistent with the scientific evidence and different than what had been taught in class. In many cases, these conclusions differed from their previous perceptions obtained during traditional lectures. In conclusion, the present study suggests that problem-based learning activities should be introduced early in the curriculum to avoid bias in understanding which may occur when students have been previously exposed to information presented in standard lectures. The use of interactive teaching via internet is discussed. PMID- 10530165 TI - International Association for Dental Research. Symposium: Ageing Changes in Human Muscle and Bone in Relation to Oral Function and General Health. Nice, 26 June 1998. PMID- 10530166 TI - Oro-facial muscles: internal structure, function and ageing. AB - Structure and function are reviewed in the masticatory muscles and in the muscles of the lower face and tongue. The enormous strength of jaw closure is in large part due to the pinnated arrangement of the muscle fibres in the masseter. This muscle, like other masticatory muscles, is unusual in that the cell bodies of the muscle spindle afferents lie in the brain stem rather than in an external ganglion; spindles are absent in the lower facial muscles. Although few data are available, the numbers of motor units in the masticatory muscles, and probably in the lower facial muscles also, appear to be much greater than in limb muscles. The motor units in the facial and tongue muscles are largely composed of histochemical type II ('fast-twitch') fibres, but in the masticatory muscles there are substantial numbers of fibres intermediate between type I ('slow twitch') and type II, and fibre type grouping is present. In comparison with limb muscles, there is little information on ageing changes in oro-facial muscles. The masticatory muscles do, however, show some atrophy and loss of X-ray density, while motor unit twitches are prolonged. Strength is reduced in the tongue and masticatory muscles. It is known that limb muscle properties are largely governed by their innervation, both through the pattern and amount of impulse activity, and the delivery of trophic messengers; the situation for oro-facial muscles is unclear. The structural and functional differences between the two types of muscle indicate the need for conducting ageing studies on the oro-facial muscles, rather than relying on extrapolations from limb muscles. PMID- 10530167 TI - Exercise and nutritional needs of elderly people: effects on muscle and bone. AB - Advancing age is associated with a remarkable number of changes in body composition. Reductions in lean body mass have been well characterized. This decreased lean body mass occurs primarily as a result of losses in skeletal muscle mass. This age-related loss in muscle mass has been termed sarcopenia. Loss in muscle mass accounts for the age-associated decreases in basal metabolic rate, muscle strength, and activity levels, which, in turn is the cause of the decreased energy requirements of the elderly. In sedentary individuals, the main determinant of energy expenditure is fat-free mass, which declines by about 15% between the third and eighth decade of life. It also appears that declining caloric needs are not matched by an appropriate decline in caloric intake, with the ultimate result an increased body fat content with advancing age. Increased body fatness along with increased abdominal obesity are thought to be directly linked to the greatly increased incidence of Type II diabetes among the elderly. This review will discuss the extent to which regularly performed exercise can effect nutritional needs and functional capacity in the elderly. In addition, some basic guidelines for beginning an exercise program for older men and women, and establishing community-based programs are provided. PMID- 10530168 TI - Age changes in bone. AB - Changes in bone structure as a function of age have been studied by simple inspection, x-ray imaging, stereo-photography, deep field optical microscopy, circularly polarised light microscopy, and scanning electron microscopy (SEM), including both topographic and compositional backscattered electron (BSE) imaging modes. The study of bone as a three-dimensional object, rather than in thin sections, enables us to envisage modelling and remodelling processes in context. The study of ultra-flat block surfaces permits the acquisition of data from an effectively very thin layer in the block face, and to examine bone as a spectrum of tissue types varying in the degree of mineralisation. Particular attention has been paid in our earlier studies to the iliac crest, lumbar vertebral bodies, femoral mid-shaft, neck and head and parietal and frontal skull bones. Recently, we have compared findings from these sites with observations on the mandible. We conclude, from our new imaging data, that common generalisations about the changes in bone in ageing and osteoporosis are too simplified, and that the mandible differs sufficiently from post-cranial skeletal sites that it would be unwise to extrapolate from findings in the jaw to the circumstances elsewhere. PMID- 10530169 TI - Cellular and molecular aspects of muscle growth, adaptation and ageing. AB - Although major advances have been made over the past few decades in prosthetic dentistry, deterioration in oral function and altered facial appearance are still common accompaniments of ageing. Molecular biology methods now allow us to understand these age-related changes at the level of gene expression. Muscle loss as well as bone loss still present major problems, the magnitude of which increases as the age profile of our society changes. Both muscle and bone tissue respond to mechanical signals for which bone depends on muscle and for muscle, stretch has been shown to be important as it induces protein synthesis and an increase in girth as well as length of the muscle fibres. The latter involves the production of more sarcomeres in series so that the jaw muscles adapt to a new functional length following changes in vertical dimension of occlusion. It also determines the postural position of the lower jaw. In our investigations into the control of muscle mass we have recently cloned a growth factor which is expressed in exercised and/or overloaded muscles. This comes in two forms: an autocrine or local form and a paracrine or systemic form. Both growth factors influence muscle growth markedly and it is probable that the systemic type is also involved in maintenance of bone. The discovery of these growth factors provides the mechanisms by which mechanical signals are transduced into chemical signals that in turn regulate gene expression and protein synthesis. PMID- 10530170 TI - Partial dentures as an independent indicator of root caries risk in a group of older adults. AB - OBJECTIVES: To estimate the independent association between the wearing of removable partial dentures (RPD) and the presence of root caries in a population of older adults. DESIGN: Multivariate logistic regression modeling of root caries prevalence using different measures of root caries as dependent variables. The model included measures of disease history as indicators of historical risk. SETTING: Data collected in the field from three areas of England. SUBJECTS: Random sample of adults aged 60 years and over, drawn from lists of patients registered with general medical practitioners. INTERVENTION: Field measurements of a range of oral health variables including oral disease, disease history, oral status and various social and demographic measures. MAIN OUTCOME MEASURES: The presence of root caries, unsound and sound root restorations. RESULTS: Of the five different models of root caries prevalence which were used, RPDs featured as an independent risk indicator for root surface caries in the three which were related to the presence of untreated disease. The odds ratios for the contribution made by RPDs were all over 1.6, and when considered alone was in excess of 2 in one model. These models were generally well fitting. RPDs did not feature as a risk indicator in the two models which related only to the presence of root surface restorations. CONCLUSIONS: In this study, where RPDs were present, the odds of untreated disease being present increased substantially. PMID- 10530171 TI - The normative need for tooth extractions in older adults in Ontario, Canada. AB - OBJECTIVE: To determine the reasons for clinically defined need for tooth extractions were examined. DESIGN: Descriptive survey; interview and clinical data. SETTING: The City of North York, Canada. SUBJECTS: 1,531 dentate adults aged 65 and over, 69% being nursing home residents. MEASUREMENTS: Age, sex, type of residence and dental attendance pattern. OUTCOME MEASURE: Normative need for tooth extraction. RESULTS: One or more extractions were required by 38% of nursing home residents and 21% of independently-living subjects. The mean number of teeth indicated for extraction were 1.4 and 0.6, respectively. Among nursing home residents, caries was more often the reason for extraction for almost all tooth types, but for independently-living subjects periodontal reasons were more common. Overall, a significantly higher proportion of nursing home residents needed extractions due to both caries and periodontal reasons (25% and 16%) compared to subjects who lived independently (10% and 11%). For nursing home residents in all age groups, more subjects required caries-related extractions, but for independently-living subjects about equal proportions required extractions due to caries and periodontal diseases. Although the percentage of subjects requiring extraction due to caries varied significantly by age and residence type, the percentage needing extractions due to periodontal diseases showed slight variation and remained under 20%. CONCLUSION: This study indicates that among high risk groups of older adults caries continues to be the major reason for tooth extraction. However, problems with sampling limit the external validity of these results. PMID- 10530172 TI - The reasons for tooth loss in geriatric patients attending two surgical clinics in Jerusalem, Israel. AB - OBJECTIVES: Numerous studies around the world have been conducted in order to understand the reasons for tooth extractions in various age groups. Most studies have dealt with the general adult population but little attention has been paid to the elderly population. In Israel, as in most of the western countries, the elderly population is growing rapidly and thus demands for its dental needs are also increasing. In order to meet the dental requirements of the geriatric population, data on the main reasons of tooth mortality have been collected. DESIGN: Retrospective analysis of reasons for extraction divided into three categories: caries, periodontal disease and "other". SUBJECTS: The files of 302 consecutive elderly patients aged 65-95 years attending for extraction. The cause for tooth extraction was gathered from the written diagnosis described by the operator as well as from radiographs. Setting Two surgical clinics in Jerusalem serving low income residents. Results Results indicated that 30% of the extractions were due to caries, 65% were due to periodontal disease and only 6.4% related to "other" reasons. In both, males and females, periodontal disease was the major cause for tooth loss yet, females exhibited more extractions due to caries than males (35% vs 23% respectively). A relatively high incidence of tooth loss was documented for the 85+ age group. Of the teeth that were extracted, incisors and molars were equally the most frequent (29%) followed by premolars (26%) and canines (17%). Premolars were the teeth most frequently extracted out of the teeth removed due to caries (32%) whereas incisors were the most frequently removed within the group of teeth extracted due to periodontal disease (31%). CONCLUSIONS: The results of this study point to the importance of prevention and treatment of dental diseases, particularly periodontal disease, in adults aged 0 years and above in order to prevent tooth loss in their later years. PMID- 10530173 TI - Oral health status and related behaviours of U.S. nursing home residents, 1995. AB - SETTING: The 1995 U.S. National Nursing Home Survey. AIMS: A descriptive overview of reported oral health status and related behaviours for residents. DESIGN: Cross-sectional survey. SUBJECTS: 8,056 residents. INTERVENTION: Interviews by knowledgeable staff, examination of charts and/or personal knowledge. MAIN OUTCOME MEASURES: Information on the overall condition of oral health, edentulous status, presence/absence of dentures, difficulty chewing or biting, dental care received in past month, and method of payment for dental services in addition to personal characteristics and medical information. RESULTS: Fifteen percent of the residents were described as having excellent or very good oral health. Forty-one percent of the residents had difficulty chewing and biting. Almost one half of nursing home residents (47%) were totally edentulous. Twenty-six percent of residents had dental services paid for during the past month. Nursing home residents have multiple known barriers to oral health: ability to pay, extended periods of time without direct access to the dental care delivery system, and serious chronic medical conditions. CONCLUSIONS: There are serious deficiencies in achieving one of the US national health objectives: ensuring adequate oral health care for institutionalized individuals. PMID- 10530174 TI - Cephalometric evaluation of the changes in mandibular symphysis after 7 years of denture wearing. AB - OBJECTIVE: To evaluate changes in mandibular symphysis during 7 years of complete denture wearing following extraction of natural anterior teeth. DESIGN: Comparison among measurements taken at four different occasions in a prospective cephalometric study. SETTING: The study was conducted at the Dental School of Athens University. SUBJECTS: 10 complete denture wearers (5 women, 5 men) with average age of 53.2 years, at the beginning of the study. MEASURES: Linear and area measurements of the mandibular symphysis. RESULTS: The overall reduction in the anterior mandibular height was 7.87 mm and in the mandibular symphysis area 54.8 mm2 (p < 0.05). Females presented higher total average reduction in both variables tested, and more rapid bone loss during the first two years of denture wear, compared to men. Superimposition of the tracings revealed considerable individual variation in mandibular symphysis changes. CONCLUSION: Results are in line with the findings of other authors indicating continuous reduction and dramatic inter-subject variation in the mandibular alveolar bone, following extraction of natural teeth and wearing of complete dentures. PMID- 10530175 TI - Oral health knowledge, beliefs and practices of a sample of Chinese elders in Inner London, UK, aged 54 years and over: a pilot investigation. AB - OBJECTIVES: To explore the self-reported oral health and health behaviours of a sample of Inner London Chinese elders and the impact of their self-reported oral health on their social functioning and eating ability. DESIGN: Cross sectional. SETTING: Luncheon clubs in Central and East London, UK. SUBJECTS: 54 Chinese elders aged 54-81 years. INTERVENTION: A structured questionnaire, administered by two interviewers in Cantonese. MAIN OUTCOME MEASURES: Knowledge and beliefs about the causes and prevention of tooth decay and gum disease, the oral conditions experienced in the previous twelve months and the impact of these conditions. RESULTS: Overall health was rated more positively than oral health, although those who reported below average oral health were more likely to report below average overall health. Whilst over half thought that sugar and sweet food could cause tooth decay, only 19% thought that poor oral hygiene could cause gum disease. Over half thought that "hot air" caused gum disease. Three quarters brushed their teeth at least twice a day. Two thirds had experienced at least one oral condition in the previous twelve months, with the more elderly being more likely to report this. Social impacts affected 41% of the sample whilst 44% suffered at least one dietary impact. CONCLUSIONS: The burden of oral conditions is substantial, especially on the more elderly members, impacting on the performance of social functions. These burdens indicate a need for oral health services. Beliefs in the traditional Chinese explanation of "hot air" as a cause of gum disease were common. Health promotion effort should consider these when developing health messages. PMID- 10530176 TI - The teaching of prosthodontic care for older people: a non-rote philosophy. AB - This essay complements that de Baat et al in the last issue with emphasis on the importance of the variability between individual older people. The consequent need for an open minded approach towards planning Prosthodontics is discussed, based on each patient's motivation for aesthetics, function, comfort and self esteem. Both functional expectations and motivation to learn effective health behaviour vary widely, and evaluation of both is essential for realistic planning because further tooth loss and the need for partial dentures occur so frequently. The consequent variation in plans raises the question--which are the strategic teeth to maintain a stable dental occlusion or a future tooth stabilised denture? For undergraduates this demands a non-rote approach to learning. PMID- 10530177 TI - A comparison of four home-care fluoride programs on the caries incidence in the elderly. AB - OBJECTIVE: To estimate the caries preventive effect of 4 fluoride programs over 2 years in the elderly. SETTING: The Public Dental Clinics of Balsta and Knivsta and the Faculty of Odontology in Goteborg, Sweden. SUBJECTS: One hundred and sixty-four individuals, aged 60 years and older (mean age 71.5 years) who were considered to be at risk from caries. DESIGN: The participants were randomly assigned either to: 1) rinse twice a day with a 0.05% NaF solution (n = 49; rinsing group), 2) suck twice a day on a 1.66 mg NaF tablet (n = 51; tablet group), 3) brush their teeth three times a day using a toothpaste slurry rinsing technique (n = 32; slurry group), or 4) brush their teeth in their usual manner (n = 32; control group). The participants in all 4 groups used a fluoride toothpaste (containing 0.32% NaF) at least twice daily. RESULTS: No new carious lesions were found in 67% of the participants in the rinsing, 43% in the tablet, 25% in the slurry and 16% in the control group over the 2 years. The mean (+/- SD) 2-year caries increment was 0.8 +/- 1.4, 1.4 +/- 1.7, 1.9 +/- 1.9 and 2.3 +/- 2.1 DFS in the rinsing, tablet, slurry and control groups, respectively; it was significantly lower in the rinsing than in the control group (p < 0.01). A lower incidence of DFS was also found in the tablet group than in the slurry group, but only for the lingual surfaces (p < 0.05). CONCLUSION: The type of fluoride program may be of importance in the reduction of new caries lesions in an older population. PMID- 10530178 TI - Attitudes to the importance of retaining natural teeth in an adult Swedish population. AB - OBJECTIVES: To evaluate the attitudes to retaining natural teeth in an adult Swedish population, and to correlate the attitude to retaining natural teeth with some presumed influencing background factors. DESIGN: Cross-sectional study using a newly developed questionnaire. SUBJECTS: From the national census register of four municipalities in the southern part of the province of Holland, Sweden, with a total population of 126,878 adult (> or = 20 years) inhabitants, 4,200 persons were selected at random. The sample was randomised by age and sex, and 300 men and 300 women from the age groups 20, 30, 40, 50, 60, 70, and 80 years were included. INTERVENTION AND MAIN OUTCOME MEASURES: The questionnaire aimed to evaluate the number of remaining natural teeth, the dental care habits, the self estimated quality of natural teeth, and the attitude to retaining natural teeth in the studied population, and also to evaluate the possible correlation between those factors, in particular, the attitude to retaining natural teeth versus the other factors. RESULTS: It was found that the attitude to the importance of retaining natural teeth was strongly correlated with the number of remaining natural teeth, the dental care habits, and the self-estimated quality of natural teeth. Also sex had an influence on this attitude but not age. CONCLUSIONS: The attitude to the importance of retaining natural teeth in an adult Swedish population is correlated with the number of remaining natural teeth, the dental care habits, the self-estimated quality of natural teeth, and sex, but not with age. PMID- 10530179 TI - Reasons for referral of very elderly patients to the community dental service in rural England and the implications for developing oral health care services. AB - OBJECTIVE: To analyse the medical conditions and dental treatment requirements. DESIGN: Retrospective analysis. SETTING: Rural South West Surrey, England. SUBJECTS: 100 patients aged 75 or over who were referred for dental treatment to the Community Dental Services (CDS) because of various medical disabilities. MEASUREMENTS: Age, sex, medical history including drug intake, dental treatment provided, time taken for treatment. RESULTS: Two-thirds of the patients required domiciliary care; two-thirds had problems of mental confusion. In addition, 89% were dependent on carers. Treatment requirements indicate that the majority of dental care, although time consuming, were simple in nature and within the technical scope of a competent general dental practitioner (GDP). CONCLUSION: This study implies that the reasons for referral are other than lack of dental skills and may be due to perceived difficulties in managing patients. Further research is recommended to establish whether aspects of patient management are problematic so that resources, including appropriate training programmes, can be developed to enable a quality service to be provided. In the evolving health care system in the United Kingdom (UK) these issues will need to be considered when developing contracts at a local level. PMID- 10530180 TI - Burning mouth in Parkinson's disease sufferers. AB - OBJECTIVE: The purpose of the study was to determine the prevalence of burning mouth (BM) in a population of Parkinson's Disease (PD) sufferers and also to assess the use of pain profiles in identifying the type of burning sensation experienced. DESIGN: Subjects were surveyed by means of a one shot postal questionnaire for which ethical approval had previously been granted. Anonymity was guaranteed and therefore no attempt was made to follow up non-respondents. MAIN OUTCOME: BM was reported by 24% of respondents. The pain profiles were completed by 17 BM sufferers. CONCLUSION: Burning mouth is reported to occur in 24% of PD sufferers which is 5 times greater than that of the general population. The reason for this is uncertain but the result has implications for the future care of PD patients and indicates the need for increased dental input at PD outpatient clinics. PMID- 10530181 TI - Maryland veterans' knowledge of risk factors for and signs of oral cancers and their use of dental services. AB - OBJECTIVES: The purpose of this study was to evaluate outpatient veteran'i knowledge about risk factors for and signs of oral cancers, and their utilization of dental services. DESIGN: Patients receiving primary health care services were surveyed during August 1997. SETTING: Primary health care services at three medical centres within the VA Maryland Health Care System (VAMHCS). SUBJECTS: A total of 135 outpatient veterans were interviewed. INTERVENTION: Questionnaire administered by trained interviewers. MAIN OUTCOME MEASURES: Fifteen percent of the sample were eligible for dental care at the VA, while over 40% of those veterans participating in the study were unaware of their VA eligibility for dental services. Fifty six percent of the total sample received dental services from a private dentist, while 13% reported they had no provider of dental care. Of those not eligible for dental care at the VA (n = 115), the majority (67%) received dental care from a private dentist. Current use of tobacco and alcohol was reported by 27% of the sample. Nonsmokers were more likely to visit the dentist in the previous year than smokers (OR = 2.39, 95% C.I. 1.11,5.12). Although 84% correctly identified tobacco use as a risk factor, only 39% correctly identified regular alcohol use as a risk factor. CONCLUSIONS: Veterans at higher risk for oral cancers were less likely to have visited the dentist in the previous year, and, overall, were ill informed and misinformed about these cancers. PMID- 10530182 TI - Affective state and acceptance of dentures in elderly patients. AB - OBJECTIVE: To examine the relationship between the affective state and acceptance of dentures by elderly patients. SETTING: Dental Outpatient clinics and a residential home for the elderly. Bialystok, Poland. DESIGN: Cross sectional survey of denture wearers. SUBJECTS: One hundred and forty one patients between the ages of 60-94. INTERVENTION: A semi-structured interview, which focused on the patient's self-evaluation of their affective state, and a clinical examination. RESULTS: There was a trend for the negative affects (irritability, boredom, anger, loneliness, helplessness) to relate to denture intolerance and for the positive affects (joy, peace, usefulness) to relate to denture acceptance. CONCLUSIONS: Anger related significantly to intolerance of upper and lower dentures. The relationship of age and gender to the acceptance of dentures in the present sample was not significant. PMID- 10530183 TI - A study of the impact of oral health on the quality of life of older people in the UK--findings from a national survey. AB - OBJECTIVES: The aim of this study was to was to determine whether older adults perceive oral health as affecting their life quality and to identify variations in impacts in relation to socio-demographic factors, dental service utilisation and method of payment. DESIGN: This study formed part of the Office for National Statistics Omnibus Survey, which utilised a random probability sample of addresses from the British Postcode Address File (PAF). SETTING: 3,000 homes were selected from one hundred post sectors across Britain. Respondents were interviewed in their homes about how oral health affects their quality of life. Subjects 454 older people (aged 65 and over) took part in the survey. MAIN OUTCOME MEASURES: The study group perceived oral health as impacting on their quality of life in general (negative and/or positive impact) (70%, 318), enhancing (53%, 241) and detracting (44%, 199) from their life quality. The most frequently perceived way in which oral health affects life quality was its effect on eating and comfort. Older people from higher socio-economic groups reported that oral health had a greater impact on their quality of life in general (positive and/or negative impacts), (OR = 1.77, 95% CI = 1.22,2.78) and specifically, enhancing their quality of life (OR = 1.52, 95% CI = 1.01,2.30). Those who reported attending the dentist within the last year perceived that their oral health enhanced their life quality (OR = 1.55, 95% CI = 1.01,2.38). CONCLUSIONS: Socio-economic background and dental attendance pattern are associated with how older people perceived oral health as affecting quality of life. These findings may have implications for promoting regular dental attendance and auditing dental services for older people. PMID- 10530184 TI - Utilisation of domiciliary dental services. AB - OBJECTIVES: To develop undergraduate dental student understanding of the attitudes of elderly people towards dentistry and of the barriers which prevent them from seeking treatment. DESIGN: Each student interviewed a) an elderly person already known to them and b) an elderly person at a Day Rehabilitation Unit using a questionnaire. Students were not trained nor standardised. SETTING: a) in the student's home locality, b) in a Rehabilitation Unit in Sheffield. SUBJECTS: 161 people were interviewed with mean ages a) 78 years and b) 85 years. MAIN OUTCOME MEASURES: Perceptions of treatment need and domiciliary treatment by these elderly people. The results are summarised but not analysed because of the inherent limitations of data derived by unsupervised students. CONCLUSIONS: This educational exercise successfully engaged the studentsi minds and, for some, generated enthusiasm. They achieved valuable insight into the topic and realised that many people who inevitably have dental problems either feel that this is acceptable or do not know that they can obtain domiciliary dental care. PMID- 10530185 TI - The effect of variation of the lingual shape of mandibular complete dentures on lingual resistance to lifting forces. AB - Increasing life expectancy, age related reduction in adaptability and progressive severe mandibular resorption all add to the importance of any factor improving the prosthetic success. OBJECTIVE: To investigate the effect of two different lingual shapes of lower dentures on patients' ability to resist lifting forces. DESIGN: Tongue pressures on the lingual surface of complete mandibular experimental dentures were recorded from mid-line, premolar and molar transducers. Two experimental prostheses were fabricated for each subject: one conventionally contoured, the other formed by piezography. SETTING: A clinical research laboratory. SUBJECTS: Five experienced complete denture wearers between age 64 and 82 years. INTERVENTION: Lifting forces were applied at the midline, left and right premolar sites in random order. MAIN OUTCOME MEASURES: Peak resistance to lifting forces and lingual pressures used during these tests. RESULTS: Lingual pressures exerted anteriorly were dramatically higher than those on premolar and molar surfaces. Significantly higher pressures were used to resist lifting forces applied to piezographically than conventionally formed contours; correspondingly, significantly higher lifting peak forces were, on average, resisted. CONCLUSIONS: Providing a lower denture with a piezographically produced lingual surface was shown, in this preliminary study, to enhance tongue retentive ability over a conventional design. It seems reasonable to maximise retentive potential with oblique sublingual polished surfaces and minimise the adaptive demand, particularly for older patients, by using a piezographic technique which "customises" the contour and precludes over-extension. PMID- 10530186 TI - In vitro comparison of the shear bond strength of amalgam to tooth structure using two bonding agents--lutting glass ionomer and 4-META. AB - Bonding dental amalgam to tooth structure using 4-META has become an accepted clinical procedure. Glass ionomer cements possess the ability to bind to tooth structure as well as to the components of dental amalgam. The present in vitro study evaluates the shear bond strength of amalgam to tooth structure using luting glass ionomer as a bond mediating agent, and compares with that obtained using 4-META. Results indicate that it is possible to bond amalgam to tooth structure using a thin layer of glass ionomer cement. The shear bond strength of glass ionomer cement mediated bond is significant and may be adequate for clinical application. PMID- 10530187 TI - Effect of re-using nickel-chromium alloy on its ultimate tensile strength, yield strength and modulus of elasticity. AB - In order to bring down the cost of the castings used in fixed partial denture prosthesis, a study was conducted by reusing the nickel chromium alloy without adding any new material. We studied the effect on the ultimate tensile strength, yield strength and modulus of elasticity after reusing the nickel-chromium alloys. Results showed degenerative changes in the properties evaluated after recasting. These changes were statistically significant, suggesting that it can not be reused. PMID- 10530189 TI - Metastasis to maxillary gingiva from carcinoma of breast. A case report. AB - A rare case of metastatic infiltrating duct carcinoma involving gingiva in relation to maxillary left canine-premolar of a 40 year old female is presented. The unilateral gingival enlargement in canine-premolar region was quite unusual. Pertinent history and histological examination revealed that the tumor was a metastatic carcinoma, the primary lesion of which was in the breast. The differential diagnosis of gingival enlargements is discussed in this paper. The resemblance of this mass to an inflammatory hyperplastic condition reflects the need for a detailed case history and examination. PMID- 10530188 TI - Intraoral botryomycosis masquerading as a pyogenic granuloma. AB - Bacterial pseudomycosis also known as Botryomycosis is a rare, indolent infection that has been described in patients with immunodeficiency and tissue with decreased healing ability. It affects the visceral organs and infection in the head and neck has been described as affecting the tongue and jaw bones. Histologically, the disease is characterized by the presence of 'Bollinger granules', surrounded by neutrophils in a fibrocellular stroma. A case of gingival Botryomycosis is presented which was diagnosed as a routine pyogenic granuloma in a healthy male. PMID- 10530190 TI - CCTV study of cemental annulations in determining the age from single tooth. AB - Cementum in human tooth is a hard tissue in its root deposited around dentin in layers throughout the life. Microscopically each layer is seen as a set of alternating dark and light bands and called as Cemental Annulations. Intact teeth obtained from subjects of either sex and of known ages processed by ground sectioning manually and mounted on a glass slide. The cemental annulations were then counted by light microscope using CCTV Screen, at the junction of cervical with middle third of root. Age was then determined by adding the eruption age in years of tooth in study to the annulations counted. This was found to be matching with actual age almost to an accuracy of +/- 1-2 years. PMID- 10530191 TI - Comparison and classification of dental arch forms of Indian and Chinese subjects with normal occlusions. AB - To compare the dental arch forms of Indian and Chinese subjects, 30 untreated Indian and 30 untreated Chinese adults with normal occlusion and symmetrical arches were examined. The arches were classified as narrow, wide, mid, pointed and flat, according to the method developed by Monique Raberin etal., from Lyon, France. For the sample examined the Chinese population was found to have significantly wider arches compared to the Indian population. PMID- 10530192 TI - Efficacy of Chitra granule and powder (hydroxyapatite) in alveolar bone regeneration in rabbits. A histological evaluation. AB - The reconstruction or restoration of osseous defects caused by inflammatory periodontal disease is a continuing challenge in periodontal therapy. Great strides are being made to this effect using alloplasts such as hydroxyapatite. The present study was designed in Newzealand dwarf rabbits to observe the biologic response of periodontal tissues to synthetically prepared hydroxyapatite in both powder and granule forms by Srichitra Tirunal Institute for Medical Sciences & Technology, Trivandrum and to compare it with that of a commercially available, pure resorbable hydroxyapatite, OsteoGen (HA Resorb). The test materials were implanted in the artificially created bonydefects in the mandible via an intraoral approach. The wounds were allowed to heal upto 26 weeks postimplantation. The clinical evaluation at 12 weeks and 26 weeks postimplantation revealed neither any evidence of inflammation, infection or abscess formation nor any exposure or exfoliation of test materials. The histological examination of the implant sites at 12 and 26 weeks postimplantation revealed varying extent of formation of new osseous tissue and periodontal fibers. A comparison between test materials and control suggested that Chitra granules exhibit a relatively greater potential for newbone and periodontal fibre formation. PMID- 10530193 TI - Occurrence of midline diastema and frenum attachments amongst school children in Nairobi, Kenya. AB - The aim of this study was to determine the prevalence of midline diastema, tongue tie and frenum attachments amongst school children in Nairobi. A total of 1802 children aged between 4 and 16 years were selected randomly using multistage sampling technique. To avoid oversampling in either sex, a proportionate sampling procedure was used. Thereafter, a thorough intra-oral examination was carried out using a mouth mirror under artificial or natural light with the children lying on a supine position. Presence or absence of midline interdental spaces unusually bigger than other interdental spaces were noted and recorded on a prepared dateacollection form. Accurate location of the origin of the frenum was done using Placek et al Morphological-functional classification of the labial frenum attachments. Data was analyzed manually by tally method. Results showed that 35% had upper and lower midline diastema. 55% were females and 45% were males. Their mean age was 7.6 years. 0.2% had a high lingual frenum. The commonest location of frenum attachment amongst children with lower midline diastema was the mucogingival junction (86%) whereas amongst those with upper midline diastema it was attached gingiva (50%). None of the children had frenum attachment on the interdental papilla. It was concluded that the maxilla had a higher prevalence of midline diastema than the mandible and that papillary penetrating frenum attachments amongst these patients were higher in the maxilla than the mandible. PMID- 10530194 TI - Expression of lectin binding in oral submucous fibrosis. AB - Lectins are a group of specific glycoproteins present in cells, particularly cell membrane. Recently, lectin binding studies have been used as a diagnostic as well as prognostic indicator of neoplasm's. Oral submucous fibrosis (OSMF) is a potential premalignant condition predominantly seen in Indian subcontinent. A comparison of expression of lectin binding was studied in normal tissue, OSMF cases and oral squamous cell carcinoma. The OSMF cases were grouped into early and advanced conditions as per the histopathologic criteria. Patterns of lectin binding observed with advanced OSMF cases were comparable with that of Oral squamous cell carcinoma. The role of lectin binding studies in assessing the malignant potential of a pre-malignant condition is discussed. PMID- 10530195 TI - Salivary counts of mutans streptococci and lactobacilli in children ageing 6-8 year old having a socioeconomic background in Brazil. AB - Saliva samples from students aged 6 to 8 year-old were analysed in order to determine the incidence of Streptococcus group mutans and Lactobacillus. Two hundred children were examined, distributed in five socioeconomic categories (A to E). Stimulated saliva samples were collected and inoculated into the SB20 and Rogsa agar culture medium for the Streptococcus and Lactobacillus cultivation. After growth, the number of these microorganisms (CUF/mL) was determined after identification of the representative colonies by biochemical methods on the basis of carbohydrate fermentation. A significative part of the population, particularly among the lower socioeconomic categories (D/E) was considered a high risk group in developing dental caries because of the high number of Streptococcus group mutans and Lactobacillus. PMID- 10530196 TI - Crystal growth technique for composite resin bonding. A SEM study. AB - This study aimed to compare the surface structure of the exposed enamel and dentine treated with acid etching and various concentrations of crystal growth agents (CGA) under Scanning Electron Microscope (SEM). Fifteen maxillary central incisors which fulfilled the inclusion and exclusion criteria were used as samples. The acid used for etching the enamel was 37% orthophosphoric acid and CGA's were 2%, 5% and 10% for micacid saturated with ammonium sulfate. The samples were divided into 5 groups and expect control group others were subjected to various surface treatments. They were observed under SEM at X 1000 magnification. The acid etched enamel surface structure was similar to the previously published reports. Maximum crystal growth on enamel and dentin were observed with 5% CGA. The surface irregularity produced by crystals were similar to acid etched surface. PMID- 10530197 TI - Crystal growth bonded composite restorations. A bond strength evaluation. AB - Sixty maxillary central incisors whose incisal angle were restored with composite resin after acid etching surface treatment and various crystal growth surface treatment were subjected to bond-strength evaluation in universal testing machine, after 24 hours. The acid etching was done with 37% orthophosphoric acid. The crystal growth agents (CGA) were 2%, 5% and 10% formic acid saturated with ammonium sulfate. The mean bond strength of composite restoration treated with 5% CGA were not statistically from acid etched restorations. PMID- 10530198 TI - Current research in diagnostic methods for assessing periodontal disease. AB - The management of chronic periodontal disease is beset with a numbers of diagnostic as well as therapeutic problems. Assessment of periodontal disease progression has been made mainly based on the conventional methods such as clinical parameters and radiographic interpretation. A high level interest in the development of diagnostic tests capable of detecting factors associated with progressing periodontal disease has continued in the past years. Recent research has shown evidence that certain microbial species and specific genetic and related factors are some strong indicators of susceptibility to severe periodontitis. Here an attempt is made to review the current trends and diagnostic tests used to assess the disease activity and predict its progression. PMID- 10530199 TI - Local antimicrobial delivery in periodontal therapy. AB - Treatment strategies towards periodontal diseases have evolved to eliminate specific pathogens or suppress destructive host response. The inherent activity of the antibiotic against the target microorganism and various Pharmacokinetic parameters such as potency, permeability, intrinsic efficacy, and substantivity of the drug dictate the success of the therapeutic outcome. However selection of an appropriate delivery system is an important factor. Rapid advances in molecular biology have helped to overcome the disadvantages of systemic and topical applications, by direct placement of antimicrobial agent (s) into subgingival sites, thus minimizing antimicrobial impact on non-oral body sites. As periodontitis is a 'localized' disease condition it is amenable to localized drug treatments. By means of controlled local delivery from within the periodontal pocket, a single administration of a few milligram of an antibacterial agent can maintain therapeutic concentrations within the crevicular fluid for a longer period of time than any other mode of delivery. This paper shall review the different local delivery systems along with the commonly employed drugs through these delivery systems. PMID- 10530200 TI - Effect of X-ray beam vertical angulation on radiographic assessment of alveolar crest level. AB - Periodontal diseases are diagnosed and monitored by various methods. Probing pocket depth measurements and dental radiographs are two of the most commonly used methods. The aim of this study was to assess the effect of x-ray beam vertical angulation on radiographic assessment of alveolar crest level in five human mandibles. A standardized technique was used to take bitewing radiographs with -10 degrees, 0 degree and +10 degrees angulation of X-Ray beam. The range of the mean differences at individual sites was from 1.84 mm (0.58 +/- SD) to 3.70 mm (1.01 +/- SD). It was found that there was a wide range of over or underestimation of the alveolar crest level due to a change in beam angulation. It was concluded that, to monitor patients with periodontal disease or treatment outcomes, it is important to have reproducible images and bitewing film holders should be used to minimize the X-Ray beam angulation error in general dental practice. PMID- 10530201 TI - Sturge Weber syndrome with intraoral manifestations. A case report. AB - A 15 Year old female patient with Sturge Weber Syndrome is presented. This neurocutaneous syndrome consists of angiomatosis of the skin and mucosa as well as the leptomeninges. This case report describes the classic presentation of the syndrome, emphasizing the oral manifestations. The radiographic and CT scan show the typical "tram line" intracranial calcifications. This case report presents a typical case of Sturge Weber Syndrome. It gives the radiological and CT scan findings and the important role played by them in the diagnosis of this syndrome. Emphasis is given to the differentiation of diphenylhydantoin induced gingival hyperplasia from the angiomatous enlargement of the gingiva before any treatment is planned. PMID- 10530202 TI - Shaping ability of .04 and .06 taper ProFile rotary nickel-titanium instruments in simulated root canals. AB - AIM: The aim of this study was to determine the shaping ability of ProFile .04 and .06 taper rotary nickel-titanium instruments in simulated canals. METHODOLOGY: A total of 40 simulated root canals made up of four different shapes in terms of angle and position of curvature were prepared using the 'crowndown' approach recommended by the manufacturer. Pre-operative pictures of each canal were recorded on optical discs using an image analysis package. The simulated canals were prepared and postoperative pictures superimposed on the original images. RESULTS: No instrument fractures occurred and none deformed; none of the canals became blocked with debris. Change in working distance was, on average, 0.063 mm with 33 canals retaining the correct length. Overall, five zips (12.5%) were created and 24 (60%) canals demonstrated a widened area on the outer aspect of the canal between the end-point and the curve. Two danger zones (5%) were created and two perforations but no ledges were found. Between canal shapes there were highly significant differences (P < 0.0001) for the incidence of zips and elbows but not for the other aberrations. There were highly significant differences (P < 0.0001) for the total width of the canals between the various canal shapes at the apex of the curve, the beginning of the curve and half way to the orifice, and a significant difference (P < 0.05) at the end-point. There were highly significant differences (P < 0.0001) for the amount of resin removed from the outer aspect of the curve at the end-point and at the beginning of the curve, and significant differences (P < 0.05) at the apex of the curve and half way to the orifice. There were highly significant differences (P < 0.0001) for the amount of resin removed from the inner aspect of the curve at the beginning of the curve and half way to the orifice. Overall, transportation was towards the outer aspect of the canal except at the beginning of the curve. CONCLUSIONS: Under the conditions of this study the combined use of .04 and .06 taper ProFile instruments was rapid, effective and produced good canal shapes except in those specimens with short curves that began near the end-point. PMID- 10530203 TI - Micro-computed tomography: a new tool for experimental endodontology. AB - AIM: Micro-computed tomography (MCT) using conebeam geometry is a method of producing true 3D images of the structure of small samples. A prototype MCT unit was adapted for imaging teeth to examine whether it could be used to quantify the instrumentation of root canals. METHODOLOGY: Ten mandibular first molar teeth that had intact crowns and fully formed roots were scanned using MCT at a resolution of 0.081 mm and 3D-rendered images created; root canals were segmented from this. Reproducibility of MCT was verified for root canal shape and size. Access cavities were prepared into the pulp space and root canals enlarged to a continously tapering preparation using a crowndown technique. Each tooth was scanned again to allow comparison of pre- and post-instrumentation images. The roots were then sectioned at five predetermined horizontal levels for video digitized measurement of dimensions of roots and root canals. The video images had a resolution of 0.025 mm. Video-digitized images of the physical cut surfaces were compared with equivalent MCT reconstructed images. The total area of the root canals (internal) and root (external) at each level were calculated from both MCT reconstructions and video-digitized images, and compared. RESULTS: There was a highly significant correlation between MCT and video images for both external and internal areas (r = 0.94). Rendered 3D images were constructed to show the root canal systems of teeth. The total volumes of the apical 7.5 mm of root canals were calculated from rendered images of nine teeth before and after instrumentation. The mean amount of dentine removed by instrumentation was 3.725 mm3, which was 28% of the original canal volume. CONCLUSIONS: Micro-computed tomography was shown to be accurate for experimental endodontology. PMID- 10530205 TI - Clinical problems associated with unusual cases of talon cusp. AB - CASE REPORT: Talon cusp is an uncommon dental anomaly manifested as an accessory cusp-like structure on the crown of anterior teeth. This report describes two unusual cases of talon cusp. Case 1 showed bilateral anomalous cusps on the palatal aspects of maxillary supernumerary teeth, causing premature contact and tooth impaction. In case 2, a double-fused talon cusp was projected from the palatal surface of a large geminated central incisor. A talon cusp is not an innocuous defect, as it may provide a substantial diagnostic, treatment planning and procedural challenge. Early diagnosis and management are important to avoid complications. PMID- 10530204 TI - Response of periradicular tissues to growth factors introduced into the surgical site in the root-end filling material. AB - AIM: The objective of this study was to evaluate the healing of the periradicular tissues when exogenous growth factors were delivered to the respected root-end. The healing response was compared with that when Diaket was used as a control. METHODOLOGY: Non-surgical root canal treatment was performed on mandibular teeth in mongrel dogs. Surgical treatment followed and included root-end resection and root-end cavity preparation. Insulin-like growth factor in combination with platelet-derived growth factor, or fibroblast growth factor alone, were then placed in the root-end preparations on a polylactic acid carrier (Atrisorb) with or without the incorporation of the carrier tetracalcium phosphate. The healing was evaluated at 60 days with regard to presence of inflammatory response, bone regeneration, periodontal ligament formation and cementum formation. RESULTS: Osseous regeneration in the excisional would and periodontal formation were significantly greater when Diaket was used as the root-end filling material. Likewise, cementum deposition occurred significantly more frequently in the Diaket group (P < 0.05). The polylactic carrier Atrisorb remained in the surgical sites for the duration of the study. CONCLUSIONS: The use of specific growth factors, FGF and a combination of IGF/PDGF, delivered to the prepared root end in a collagen carrier did not initiate the desired periradicular tissue response of regeneration. Diaket, as used in this study, did stimulate a periradicular tissue response compatible with regeneration. PMID- 10530206 TI - Cure behaviour of visible light-activated pattern materials. AB - AIM: This study tested the hypothesis that the cure behaviour (depth of cure and polymerization contraction) of light-activated pattern materials was no worse than that of light-activated composite resins, allowing them to be handled in a similar fashion. METHODOLOGY: Depth of cure was measured by a penetrometer method. RESULTS: Cure depths were comparable to those of composite resins, ranging from 3.52 mm (Lumin-X paste) to 6.76 mm (Visioform) after visible light activation for 30 s. There were significant differences in the depth of cure of the three materials tested (P < 0.001). Polymerization contraction was assessed by a minimal load transducer method. Values ranged from 0.45% (Lumin-X paste) to 1.89% (Visioform), lower than that of composite resins. There were significant differences in the polymerization contraction values for each of the three materials (P < 0.001). CONCLUSIONS: It was concluded that light-activated pattern materials cure in a manner comparable to composite resins, and may be built up incrementally in a similar fashion. PMID- 10530208 TI - Black-pigmented anaerobic rods in closed periapical lesions. AB - AIM: This study determined the frequency of Porphyromonas endodontalis, Porphyromonas gingivalis, Prevotella intermedia and Prevotella nigrescens in 20 closed periapical lesions associated with symptomatic and asymptomatic refractory endodontic disease. METHODOLOGY: To deliniate possible oral sources of P. endodontalis, the presence of the organism was assessed in selected subgingival sites and saliva in the same study patients. Periapical samples were obtained by paper points during surgical endodontic procedures using methods designed to minimize contamination by non-endodontic microorganisms. Subgingival plaque samples were obtained by paper points from three periodontal pockets and from the pocket of the tooth associated with the closed periapical lesion. Unstimulated saliva was collected from the surface of the soft palate. Bacterial identification was performed using a species-specific polymerase chain reaction (PCR) detection method. RESULTS: P. endodontalis was not identified in any periapical lesion, even though subgingival samples from eight patients (40%) revealed the P. endodontalis-specific amplicon. P. gingivalis occurred in one periapical lesion that was associated with moderate pain. P. nigrescens, P. endodontalis and P. intermedia were not detected in any periapical lesion studied. CONCLUSIONS: Black-pigmented anaerobic rods appear to be infrequent inhabitants of the closed periapical lesion. PMID- 10530207 TI - A comparison of MTA, Super-EBA, composite and amalgam as root-end filling materials using a bacterial microleakage model. AB - AIM: The aim of this study was to compare traditional and newly developed root end filling materials for resistance to bacterial microleakage. METHODOLOGY: Sixty extracted single-rooted teeth were randomly divided into five groups for root-end filling with mineral trioxide aggregate, Super-EBA, TPH composite resin with ProBond dentine bonding agent, Dispersalloy amalgam with and without ProBond, and positive and negative control groups. Root canals were instrumented using the step-back technique and simulated root-end resections performed. Root end filling materials were placed in 3 mm ultrasonic retropreparations. Nail varnish was applied to all external root surfaces to the level of the resected root ends to prevent lateral microleakage. Samples were sterilized in an ethylene oxide sterilizer for 12 h. Using a newly designed model system, the apical 3-4 mm of the roots were immersed in BHI culture medium with phenol red indicator within culture chambers. The coronal access of each specimen was inoculated every 48 h with a suspension of Streptococcus salivarius. Culture media were observed every 24 h for colour change indicating bacterial contamination. Media demonstrating colour change were plated for S. salivarius. Samples were observed for 12 weeks. RESULTS: At 4 weeks 10% of specimens from each experimental group had evidence of leakage. At 8 weeks 20% of specimens filled with amalgam without dentine bonding agent, Super-EBA and MTA had evidence of leakage. At 12 weeks minor differences between materials were noted. CONCLUSIONS: Under the conditions of the study, despite some variations, there were no statistically significant differences in rate of microleakage among the five groups tested at either 4, 8 or 12 weeks. PMID- 10530209 TI - Root canal anatomy of anterior and premolar teeth in Down's syndrome. AB - AIM: The aim of this study was to examine tooth length and root canal anatomy of 281 anterior and premolar teeth from 66 DS individuals using a standard protocol. RESULTS: The results indicated that root canals in DS are relatively simple and that there is a significant reduction in root and crown length. CONCLUSIONS: We propose that these observations are commensurate with the suggestion that trisomy 21 exerts its effect by slowing the mitotic cycle and rate of cell proliferation, resulting in generalized retardation of growth. PMID- 10530210 TI - The removal of the smear layer using the Quantec system. A study using the scanning electron microscope. AB - AIM: The aim of this study was to determine the ability of the Quantec Series 2000 rotary nickel-titanium endodontic system to remove dentinal debris and smear layer produced during canal preparation. METHODOLOGY: A first group (control) of 12 curved root canals was prepared using conventional manual instruments and the step-back technique. A second group of 12 curved root canals was instrumented using the complete Quantec sequence according to the manufacturer's instructions. In both groups, irrigation was performed using a 3% NaOCl solution. The canal walls were observed under a scanning electron microscope and the coronal, middle and apical thirds of each canal photographed at a magnification of 500. The views were divided into 10 subareas by overlaying a grid, and the absence or presence of a smear layer was rated and scored on three appearances using the scale described by Ciucchi et al. (1989). RESULTS: The scores were higher (i.e. less debris was present) in the middle third (P < 0.0001) and in the apical third (P < 0.0001) of canals prepared with the Quantec system when compared with those prepared with K-files. Nevertheless, in canals prepared with Quantec instruments, the scores were significantly higher in the coronal third compared with the apical third (P < 0.005). CONCLUSIONS: The Quantec rotary system produced cleaner canal walls than conventional manual instrumentation, particularly in the middle and apical thirds. This finding may imply that stresses applied to the cutting regions of Quantec instruments by accumulation and compression of the smear layer are minimized. PMID- 10530211 TI - The control of haemorrhage at the operative site during periradicular surgery. AB - CLINICAL TECHNIQUE: Effective apical sealing in endodontic surgery requires a dry root-end cavity to insert the filling material. A number of procedures for controlling haemorrhage have been described in the literature. An improvement of these techniques is proposed in this paper: by using a mixture of surgical wax and fibres of calcium alginate. This device, easy to place, sterile and non toxic, permits placement of a root-end filling under more favourable conditions. PMID- 10530212 TI - Endodontic management of a mandibular third molar fused with a fourth molar. AB - CASE REPORT: Developmental anomalies in permanent molars frequently require surgical intervention. A case of a mandibular third molar fused with a fourth molar which was successfully treated with conservative endodontic therapy is reported. PMID- 10530213 TI - Deposition of calcified tissue around an overextended gutta-percha cone: case report. AB - CASE REPORT: Root canal treatment was performed in a mandibular right second premolar with a periapical lesion and apical resorption. The root canal was prepared with K-files using the step-back technique and 3% NaOCl as an irrigant; during obturation gross overfilling of gutta-percha occurred. The tooth was permanently restored with a post and core along with a crown. Although healing of the periapical lesion occurred and the patient reported that he was symptom-free, the tooth was extracted after 4 years because of a subgingival root fracture. Following extraction the tooth was examined with SEM. The examination revealed the presence of newly formed calcified tissue at resorption sites on the root apex. This newly formed tissue extended from the surface of the root around the apex to the extruded gutta-percha cone to which it was well adapted, forming a bridge between the cone and the root. PMID- 10530214 TI - A case of unusual anatomy: a maxillary lateral incisor with three canals. AB - CASE REPORT: A case of an unusual maxillary lateral incisor with three root canals is presented. The patient presented with localized swelling and a sinus tract. Because of its abnormal anatomical configuration, the tooth presented diagnostic and procedural difficulties. Conventional root canal treatment was performed successfully. PMID- 10530215 TI - A survey of cases of cancrum oris seen in Ile-Ife, Nigeria. AB - AIM: To review the records of child patients with cancrum oris who presented at a teaching hospital in Nigeria. SETTING: Obafemi Awolowo University Teaching Hospital, Ile-Ife, Nigeria. SAMPLE AND METHODS: The study was carried out through review of records of child patients diagnosed as having cancrum oris or post cancrum oris defects between 1982 and 1996. Age, gender, site distribution, treatment and its outcome were recorded. RESULTS: One hundred and forty-two cases were diagnosed during the study period. Mean age was 4.65 years (range 2-16 years). The maxillary quadrants were affected more often than the mandibular. Seventeen patients completed treatment satisfactorily, but 55 failed to do so. All patients had evidence of malnutrition. CONCLUSIONS: The findings of this and other studies demonstrate the need for major initiatives to address the underlying causes of cancrum oris and to promote the utilization of health care. PMID- 10530216 TI - Preventive oral health care and health promotion provided for children and adolescents by the Municipal Dental Health Service in Denmark. AB - OBJECTIVE: To describe the current organization of health promoting and preventive activities within the Danish Municipal Dental Health Service and to assess how the service has chosen to comply with the directives as formulated by the National Board of Health. DESIGN: A cross-sectional survey of the municipal dental health services was carried out on a national scale. Postal questionnaires were used to collect information on active and passive preventive care activities and community-orientated health promotion. SETTING: The survey was conducted to aid the reorientation and adjustment of the Municipal Dental Health Services in Denmark. SUBJECTS: All municipal dental health services in Denmark were considered relevant for the survey and 141 services (71%) responded to the questionnaire. OUTCOME MEASURES: Quantitative methods were used to measure recall intervals for children and adolescents, passive and active prevention, identification of and care for individuals at risk, and health education. Qualitative methods were applied to record the organization of community health activities. RESULTS AND CONCLUSIONS: The majority of dental services stated that preschool children are called at regular intervals (every 3, 6 or 8 months); school-children and adolescents are most often recalled according to individual needs. Chairside assistants, dentists or dental hygienists give oral hygiene instructions systematically to children of grades 0 through to 3. Fluoride is frequently administered through topical application by dentists; fluoride tables are not used. Permanent molars are sealed when this is indicated. Clinical and socio-behavioural criteria are used to identify children at risk. Half of the services reported school-based health education, and in one-quarter of the municipalities community health activities took place. Adjustment of the services should consider population-directed activities and greater use of ancillary personnel. PMID- 10530217 TI - Pre- and post-treatment levels of salivary mutans streptococci and lactobacilli in pre-school children. AB - OBJECTIVES: To examine the effect of operative and restorative treatment of dental caries on the levels of caries associated microorganisms in saliva and to relate alterations to the type and extent of treatment. DESIGN: Longitudinal. SETTING: Paediatric Dentistry Department at a central hospital in Sweden. SUBJECTS AND METHODS: One hundred and eight pre-school children with severe dental caries scheduled for treatment under general anaesthesia. Chair-side tests were used to estimate the levels of salivary mutans streptococci, lactobacilli and buffer capacity before the surgery and at recall appointments 1 and 6 months after treatment. Caries were assessed according to WHO guidelines and the number of extracted teeth and filled surfaces during surgery were recorded. RESULTS: The results demonstrate that the post-treatment levels of mutans streptococci and lactobacilli were significantly reduced (P < 0.001) compared to pretreatment levels. Lactobacilli levels were more dramatically reduced than mutans streptococci. The reduction of mutans streptococci was positively correlated to the number of extracted teeth (P < 0.01), but not to the number of restored or ground surfaces. Lactobacilli reduction was not significantly related to the type of treatment. CONCLUSION: The findings suggest that extensive operative and restorative dental care effectively reduces the levels of caries associated with microorganisms during a period of at least 6 months. PMID- 10530218 TI - Evaluation of dentition status and oral hygiene in Polish children and adolescents with congenital haemorrhagic diatheses. AB - AIM: To investigate the dental status and oral cleanliness of a group of patients with haemorrhagic diatheses and to compare these to values for a group of healthy controls. DESIGN: Cross-sectional study of patients and controls. METHODS: The group of patients was made up of 80 children aged 4-18 years. The total included 77 patients with type A or type B haemophilia and three with von Willebrand's disease. The control group consisted of 80 healthy children of the same age range. A clinical examination was carried out to determine dental status and oral hygiene in the two groups. RESULTS AND CONCLUSIONS: No significant differences were found between the dental status of sick and healthy children. However, worse dental status was seen in children with severe forms of haemophilia A and von Willebrand's disease compared to other sick children. Oral hygiene status was worse in the sick children than in healthy children. PMID- 10530219 TI - Shifts in tooth maturation patterns in non-French Canadian boys. AB - AIMS: This study was designed to evaluate the predictability of dental maturation patterns in non-French Canadian boys using the maturation curves developed by Demirjian & Goldstein [1]. SAMPLE: Archival cephalometric radiographs taken during a longitudinal study (1930-1960) of Caucasian Americans were re-evaluated. METHODS: Maturation of the permanent teeth on the left side of the mandible was determined according to the methods described by Demirjian & Goldstein. RESULTS: Of the 79 boys studied, 12 (15.2%) started below Demirjian's median and continued as such during the maturation process. Seven (8.9%) started within the second standard deviation above Demirjian's median and continued as such during the maturation process. The remaining 60 boys (75.9%) started below the median of Demirjian's curves at an early age. They matured at a rate that placed them above the median value at the end of the study period. The shift took place before the age of 6 years in 46 (76.6%) cases and between the ages of 6 and 8 years in another nine (15%) cases. In five of the cases which started below Demirjian's median (8.3%) the shift only took place after the age of 9 years. CONCLUSION: The shift from below median to above median value was considered an important factor in treatment planning. The data indicate that there is considerable risk for treatment planning prior to the age of 8 years. The risk is highest when the children are less than 6 years of age due to growth prediction uncertainties. PMID- 10530220 TI - Dental maturation of 18th and 19th century British children using Demirjian's method. AB - AIM: To compare dental age with chronological age in a group of children born approximately 200 years ago and a group of modern children. METHODS: Dental maturation of 15 skeletal remains (range 3.0-15.1 years) of London children of known age-at-death was compared to an age and sex matched control group of contemporary children (n = 30). The method of Demirjian, Goldstein and Tanner (1973, 1976, 1978) was used to assess maturity. RESULTS: The difference between dental age (DA) and chronological age (CA) for both groups was not significant, suggesting similar maturation over 200 years, however, many of the younger children from Spitalfields were dentally delayed. Several of the younger individuals from both groups had a dental age less than the lowest limit of this scale (2.5 years), highlighting one pitfall of this method. CONCLUSION: These results suggest that this method is not entirely suitable for younger children. PMID- 10530221 TI - Desmoplastic fibroma of the mandible: report of a case. AB - This paper describes a case of desmoplastic fibroma in a 4-year-old patient with a history of a small slowly growing swelling at the right angle of the mandible over a 3-month period. Desmoplastic fibroma was diagnosed on histological and immunohistochemical bases. The lesion responded well to thorough curettage and has not shown signs of recurrence 3 years after the surgical intervention. The clinical picture, the pathology and the management of the case are described, and the differential diagnosis and treatment are discussed. PMID- 10530222 TI - A nasopalatine cyst in an 8-year-old child. AB - The nasopalatine cyst (NPC) was first described in 1914 and it is considered the most common non-odontogenic cyst. Most studies show a higher incidence of NPC among males than females, with a male/female ratio of 1.7:1. The majority of the cases described in people in their fifth decade involve Afro-Caribbeans, while those in their sixth decade are mainly caucasians. These cysts are normally asymptomatic, unless they are infected. The most commonly reported clinical symptom is swelling in the anterior part of the palate. The treatment of choice is enucleation. Even though it has been stated that NPCs may occur at any age no reports have been made on children less than 9 years old. Some reports support a predisposition in young Afro-Caribbeans, where NPCs appear to be more aggressive, larger and symptomatic. We present a case of a NPC in an 8-year-old caucasian female. PMID- 10530223 TI - Leiomyomatous hamartoma of the anterior tongue: report of a case and review of the literature. AB - A case of leiomyomatous hamartoma located on the anterior third of the tongue in an otherwise healthy 6-year-old boy is reported. The lesion had been present since birth as an asymptomatic, pedunculated tubular tumour. The lesion was surgically excised and histopathological examination disclosed a non-neoplastic mass composed of irregularly arranged bundles of smooth muscle embedded in a fibrovascular stroma. No recurrence was observed after 1 year of follow-up. PMID- 10530224 TI - The safety and efficacy of treatment with air abrasion technology. AB - AIMS: To evaluate patient and operator exposure to respirable particulates following the use of air abrasion in tooth preparation, and to compare the microleakage of pit and fissure sealants after conventional, bur and air abrasion preparation of the pits and fissures. METHODS: To examine air abrasion safety, sampling data were collected using a physical model of the upper torso of a patient. Previously extracted bovine incisors were prepared using an air abrasion instrument. Patient and operator exposure samples were collected. The variables examined included the size of the alumina oxide particles, the speed of particle delivery and the method of dust collection. To assess the efficacy of air abrasion, 36 extracted human molars were divided into three groups. The groups were prepared by conventional acid etching, opening the pits and fissures with a round bur, or by air abrasion. To simulate oral conditions, sealed teeth were immersed in artificial saliva and thermocycled. Teeth were immersed in a 1% solution of methylene blue and sectioned to assess the microleakage associated with each sealant. CONCLUSIONS: (1) Dust from the KCP 1000 is insufficient to be a health hazard to patients or operators, (2) chair-side suction can be used as an alternative to the KCP 1000 suction, (3) superior sealants were obtained when tooth surfaces were prepared by a bur, compared to air abrasion and conventionally prepared surfaces, and (4) air abrasion tooth surfaces demonstrated less microleakage than conventionally prepared tooth surfaces. PMID- 10530225 TI - Endochondral vs. intramembranous demineralized bone matrices as implants for osseous defects. AB - This study focuses on the difference in regenerative capacity between endochondral and intramembranous demineralized bone matrices (DBMs) when implanted into bony defects. It also focuses on the possible influence of the type of skeletal recipient site (orthotopic or heterotopic). Of 34 Wistar rats, 10 served as a source of DBM, and 24 were divided into two groups of 12 animals. In group A identical defects were produced in the parietal bones, whereas in group B the defects were produced in each radius. The right defects were implanted with endochondral DBM and the left defects were implanted with intramembranous DBM. Descriptive and/or histomorphometric analyses were performed by means of light and polarized microscopy, and radiography (group B). Right and left data were compared to disclose differences in bone-healing capacity. The quantitative results demonstrated that endochondral DBM displays a greater regenerative capacity than intramembranous DBM when implanted heterotopically. The different clinical performances of endochondral and intramembranous bone grafts might be explained on the basis of the mechanical rather than the osteoinductive principle. The qualitative results suggest that the type of bone deposition induced by the DBMs is not related to the type of implanted DBM. Recipient site characteristics and/or environmental factors seem decisive in the occurrence of either types of ossification. PMID- 10530226 TI - What is the best possible design of temporary anterior mandibular osteotomy? A histomorphological animal study. AB - When squamous cell carcinomas in the oral cavity have advanced to the size of T2 to T3 or when they start to encroach on the middle, occasionally it is necessary to divide the mandible at the parasymphysis and swing the hemimandible away from the midline for intraoral exposure of tumor resection. The osteotomy is usually performed in either a straight vertical cut, in a step-cut fashion, or in a dove tail geometric configuration. The design of the geometric pattern of the osteotomy determines whether or not the postoperative period is prolonged. The authors analyzed the best possible design of temporary anterior mandibular osteotomy. In an animal study of 12 Goettingen minipigs (GMPs) the authors investigated the best possible design of geometric pattern of mandibular osteotomy and the fragment healing process. Their primary interest was directed at the histomorphological bone-healing process after straight vertical bone cut, step-cut fashion, and dove-tail osteotomy of the mandible, and rigid fixation with an osteosynthesis plate. From this study it can be concluded that the dove tail osteotomy of the mandible is the best possible design of temporary anterior mandibular osteotomy in preventing pseudoarthrosis due to the mainly primary bone contact and gap healing process. PMID- 10530227 TI - Single lag screw fixation for malar fracture (type B) fixation: reduction of hardware treatment costs. AB - In an attempt to define the most simple and inexpensive method of achieving postreduction stability in zygomatic fractures, the authors compared two different methods of internal rigid fixation of the frontozygomatic suture line in one group with miniplates and in another group with Panfix lag screws. The randomized study protocol consisted of a control group of 20 patients and 40 patients with laterally displaced body fractures of the zygoma. Statistical differences were not seen between the control subjects and the patients who required open reduction and internal fixation either with one osteosynthesis plate at the frontozygomatic suture line or a lag screw osteosynthesis. Lag screw fixation in malar type B fractures could lower hardware treatment costs and is an alternative method that provides sufficient stability in indicated patients. PMID- 10530228 TI - A modified fixation method in supraorbital bar advancement. AB - In almost all congenital craniofacial deformity reconstructions there is a need to advance the supraorbital bar. This bar, which is fixed by several techniques, should be firm enough to minimize a relapse. In this paper a new modification during osteotomy of the supraorbital bar is presented that provides firmness and prevents relapse even without grafts. The last 15 patients with craniofacial anomalies were operated with this modification. At the stage of the supraorbital bar osteotomy, bilateral small triangles are created at the end of the bar. Then, on the lateral orbital rim, two small notches are created in which to place these triangles. By fixating these triangles to the notches, sliding of the bar and subsequent relapse is prevented, and also the fixation provided is much more rigid. PMID- 10530229 TI - Mandibular distraction osteogenesis with multidirectional extraoral distraction device in hemifacial microsomia patients: three-dimensional treatment planning, prediction tracings, and case outcomes. AB - Distraction osteogenesis of the craniofacial skeleton with the use of several different types of distraction devices (i.e., extraoral, intraoral, unidirectional, multidirectional, and customized) have been documented. However, the details of treatment planning and the method of predicting the distraction of the mandible in patients with hemifacial microsomia have not been published previously. This paper presents a technique for (1) three-dimensional treatment planning for mandibular distraction, (2) three-dimensional prediction tracings with conventional radiographs (panoramic, lateral, and posterior-anterior cephalometric), and (3) correlating the treatment planning and clinical applications. Lastly, 2 patients with hemifacial microsomia planned and treated with this approach are reported. PMID- 10530230 TI - One-stage correction of complex facial disproportion. AB - Patients with facial disharmony frequently have abnormal nasal form and disordered jaw relationships. Both orthognathic surgery and rhinoplasty are required to correct such facial disproportions. During a 10-year period 100 patients with a spectrum of indications have had concomitant orthognathic and rhinoplasty surgery by a team consisting of the same plastic surgeon, oral and maxillofacial surgeon, and orthodontist. Of this group 51 patients were operated on solely for aesthetic reasons, the majority being long face syndrome. All had a rhinoplasty usually with septal surgery. Depending on the deformity, the jaw surgery varied: 5 patients had mandibular surgery only, 12 had maxillary surgery alone, and the remaining 34 patients had both mandibular and maxillary procedures. Patients were followed for between 1 and 62 months by the plastic surgeon and for at least 2 years by the orthodontist. There was no orthognathic relapse or other major complications, but 4 patients required secondary minor nasal tip surgery under local anesthesia and 2 patients had persisting unilateral inferior alveolar nerve damage. Orthognathic surgery and rhinoplasty are not routinely performed concomitantly due to the difficulty in predicting the outcome of the soft-tissue relationships and increased morbidity. In this series, a one stage approach was used to provide facial harmony. This reduces the overall surgical and anesthetic morbidity, inconvenience, and expense, and has resulted in good cosmetic and functional results. Therefore, it is suggested that with a competent team, orthognathic surgery and rhinoplasty can be performed concomitantly with dependable results and without significant complications. PMID- 10530231 TI - Commentary on a management strategy for palatal fractures: a 12-year review. PMID- 10530232 TI - Cranial vault deformity and intracranial hypertension secondary to cephalic molding at delivery: a case and its management. AB - Cephalic molding at birth has been traditionally felt to be benign, resulting in only a transient and self-correcting cranial deformity. However, we report a 6 month-old infant who presented with extensive cephalic molding at birth in combination with persistent brachyturricephaly from unilateral coronal synostosis and occipital deformation. Helmet therapy over a 3-month period failed despite patient compliance and numerous adjustments. Intracranial hypertension developed, as documented by multiple occipital bony erosions on computed tomographic scan and by an elevated direct intracranial pressure reading. The cranial vault asymmetry was corrected in two surgical stages: (1) occipital bar advancement, temporoparietal bone remodeling, and midline sagittal strip compression to reduce vertical height, followed in 3 months by (2) fronto-orbital advancement and remodeling. PMID- 10530233 TI - Resorbable PLLA-PGA screw fixation of mandibular sagittal split osteotomies. AB - The purpose of this study was to evaluate the intraoperative placement and clinical effectiveness of resorbable copolymeric screws for mandibular sagittal split ramus osteotomies. Thirty-seven patients who underwent bilateral sagittal split osteotomies of the mandible were fixated with three 2.5-mm copolymeric poly L-lactic-polyglycolic (PLLA-PGA) screws on each side. No postoperative maxillomandibular fixation was applied. Twenty-five patients experienced mandibular advancement and 12 patients had setbacks. The average advancement was 6.5 mm (range, 3-17 mm) and the average set-back was 5.2 mm (range, 3-8 mm). Intraoperative placement was uncomplicated and no screws were stripped during placement. No problems in immediate postoperative stability were encountered and relapse was not evident in any patient. Follow-up ranged from 3 to 17 months. The screw holes remained evident radiographically after 1 year. Two and one-half millimeter copolymeric PLLA-PGA resorbable screws for mandibular ramus osteotomies appear to offer clinical results comparable with metallic screw fixation. PMID- 10530235 TI - Monobloc and facial bipartition distraction with internal devices. AB - Distraction osteogenesis (DO) permits gradual lengthening of the craniofacial skeleton. With the advent of new internal devices, monobloc (M) and facial bipartition (FB) DO are feasible. The rationale behind M and FB distraction is (1) gradual advancement of the M segment is not associated with a substantial retrofrontal dead space; (2) because 5 to 7 days elapse prior to distraction, the nasofrontal opening, in theory, is allowed to remucosalize; (3) gradual expansion of the soft tissues takes advantage of skin creep, potentially limiting relapse; (4) the procedure appears to be less invasive with decreased blood loss and operative time, enabling its use in infants; (5) overdistraction may eliminate or reduce the frequency of subsequent procedures; and (6) the procedure may be combined with FB and skull vault remodeling to provide excellent results in more complex craniofacial dysostosis problems. Five children underwent M advancement (N = 3) and M with FB (N = 2) at 9 months to 5 years of age to correct functional abnormalities such as corneal exposure, increased intracranial pressure, and apnea, as well as severe craniofacial disfigurement. Each patient underwent from 22 to 30 mm of distraction with the Modular Internal Distraction (MID) system, developed by the first author (SRC). There was one infection late in the series along the DO cable track. There were no cases of epidural abscess. In conclusion, MDO, with and without FB, appears to be a safe and effective technique for transcranial frontofacial advancement. The morbidity of the procedure appears to be less than that of conventional M advancement. PMID- 10530234 TI - A biomechanical analysis of the orbitozygomatic complex in human cadavers: examination of load sharing and failure patterns following fixation with titanium and bioresorbable plating systems. AB - The orbitozygomatic complex (OZC) and zygomatic arch act as key buttresses in the restoration of midfacial projection and width in the treatment of panfacial fractures, yet little is known about the biomechanical and deformational forces placed on this region under applied load conditions. The aims of this project were (1) to study the stress-force relationships and load sharing of the intact human OZC under subfailure loads, (2) to assess load sharing and breaking strength of the OZC when intact and after four-point miniplate fixation with either titanium (1.2 and 1.7 mm) or bioresorbable (1.5 and 2.0 mm) systems, and (3) to analyze failure patterns. Using the MTS machine, fresh frozen human skulls stripped of soft tissue underwent loading with subfailure and failure forces directed in a standard fashion. Electrical resistance gauges applied directly to local and remote bony buttresses demonstrated temporary deformation at local (zygomatic arch, lateral and inferior orbital rim) and remote (supraorbital rim) buttresses under subfailure loading conditions. Breaking strength of the OZC (N = 10) measured before and after four-point application of 1.2- or 1.7-mm titanium or 1.5- or 2.0-mm bioresorbable miniplates demonstrated a significant (p < 0.05) decrease compared with intact controls. Surprisingly, the 2.0-mm bioresorbable miniplate construct provided only 13% of the intact breaking strength of the OZC compared with 39% for the 1.7-mm titanium system (p < 0.05). Plate bending or breakage was responsible for failure of the OZC following rigid fixation. Biomechanical testing of the OZC demonstrates (1) load sharing at regional and remote bony buttresses, (2) significant decreases in breaking strength following miniplate fixation, and (3) deformation of miniplates as a primary cause of failure under load conditions. Data generated from this project may be useful with regard to optimizing fixation of the OZC in the context and treatment of panfacial fractures. PMID- 10530236 TI - Cephalometric study of posterior airway space in patients affected by Class II occlusion and treated with orthognathic surgery. AB - The posterior airway space (PAS) is delimited by hard and soft tissues with anomalies that may produce alterations in volume. In patients with severe hypoplasia of the middle and lower third of the face, a decrease in volume of the PAS is present, producing a polysyndromic condition that ranges from snoring to obstructive sleep apnea syndrome (OSAS). The aim of this report is to define PAS variations via a cephalometric study in patients affected by class II occlusion and treated with orthognathic surgery. The authors studied 44 patients affected by class II occlusion who underwent surgery to correct the maxillomandibular malformation. The patient cohort was classified according to the type of surgery performed: Le Fort I osteotomy with or without a sagittal split osteotomy. To evaluate PAS variation, cephalometric analyses were performed by pre- and postoperative lateral teleradiography. This study showed an increase in PAS volume, especially at the hypopharynx and the lower part of the oropharynx, when the sagittal split osteotomy was performed and/or the maxilla was moved anteriorly and/or superiorly. A decrease of PAS can be seen in downward and/or backward maxillary movements. In conclusion, cephalometric studies of hard and soft tissues (such as tongue, pharynx, soft palate, etc.) should be performed in all patients affected by maxillomandibular malformation. This approach may provide data for the diagnosis of respiratory pathologies that vary from snoring to OSAS. PMID- 10530237 TI - The value of transcranial Doppler ultrasonography in craniosynostosis. AB - Some children with craniosynostosis demonstrate raised intracranial pressure (ICP), requiring surgical decompression. Conventional methods of measuring ICP in such children are invasive, expensive, and require expertise. Transcranial Doppler ultrasonography (TCD) is an alternative, useful means of assessing ICP qualitatively, and is noninvasive, inexpensive, and safe. We evaluated the use of TCD prospectively in 16 children with craniosynostosis and correlated TCD findings with intraoperative ICP measurements by lumbar puncture (LP) and with computed tomographic (CT) findings. TCD evaluations were performed before and after surgery to determine the pulsatility index (PI), which is known to show close correlation with ICP. The three modalities--TCD, ICP, and CT--showed poor correlation with each other. However, the fall in the PI value after surgery, as determined by TCD, was shown to be clinically useful, with a rise in the PI value after surgery being an ominous sign. PMID- 10530238 TI - Case 8. Periapical infection. PMID- 10530239 TI - Cytotoxic evaluation of root-end filling materials in cultures of human osteoblast-like cells and periodontal ligament cells. AB - The cytotoxicity of three root-end filling materials (amalgam, IRM, and Super EBA) was evaluated in cultures of human periodontal ligament cells and human osteoblast-like cells. Ten-millimeter-long plastic test tubes were filled with 3 mm of freshly mixed root-end filling materials at one end (1.5 mm diameter). The opposite end was sealed and attached by heat to a 35-mm cell culture dish. Human periodontal ligament cells and human osteoblast-like cells were seeded in the dishes. The size of cell-free zones around the root-end filling materials and the total cell number per dish were calculated after 3 and 7 days. Empty test tubes used as controls did not influence the growth and distribution of the cultured cells. Cell density increased in all groups in the test period. Amalgam had a larger cell-free zone, compared with IRM and Super-EBA and showed a reduction in total cell number per dish for both tested cell types. IRM and Super-EBA also had a cell-free inhibition zone for both cell types, but no significant reduction in total cell number per dish. This study showed that amalgam had a higher cell toxicity to human periodontal ligament cells and human osteoblast-like cells than IRM and Super-EBA. PMID- 10530240 TI - Association of black-pigmented bacteria with endodontic infections. AB - Black-pigmented bacteria (BPB) have been associated with endodontic infections. The purpose of this study was to evaluate further the presence of BPB with the clinical signs and symptoms associated with endodontic infections. Microbial samples were collected from the root canals of 40 intact teeth with necrotic pulps and apical periodontitis. Conventional laboratory methods were used for identification of the strains of BPB isolated in pure culture. In addition, the polymerase chain reaction and specific primers for 16S r-RNA genes were used to differentiate Prevotella nigrescens from Prevotella intermedia. Twenty-two (55%) samples were positive for the growth of BPB. Of those, 11 of 22 (50%) were identified as P. nigrescens, 8 of 22 (36%) were P. intermedia, 2 of 22 (9%) were Porphyromonas gingivalis, and 1 of 22 (5%) was Prevotella melaninogenica. Sixteen of the 22 root canals positive for the growth of BPB were associated with purulent drainage either from the root canal or an associated sinus tract. Statistical analysis did not show a significant relationship for the presence of BPB with clinical signs and symptoms. PMID- 10530241 TI - Antimicrobial evaluation of calcium hydroxide in infected dentinal tubules. AB - The objective of this study was to evaluate the antimicrobial activity of calcium hydroxide in infected dentinal tubules. Four microorganisms, strains of ATCC (Streptococcus faecalis (ATCC-29212), Staphylococcus aureus (ATCC-6538), Bacillus subtilis (ATCC-6633), and Pseudomonas aeruginosa (ATCC-27853)) and one mixture of these were used. These strains were inoculated in brain heart infusion (BHI) and incubated at 37 degrees C for 24 h. Sixty-three human maxillary central incisors were prepared and sterilized by autoclaving. Five groups of 12 teeth each were contaminated for 28 days using new 24-h cultures every 72 h, prepared and adjusted to tube 2 of the MacFarland scale (6 x 10(8) cells/ml). Root canals were then irrigated with 5 ml of saline, dried, and completely filled with calcium hydroxide paste. At intervals of 0, 48, and 72 h, and 7 days, dressings were removed and teeth were immersed in 5 ml of BHI and incubated at 37 degrees C for 48 h to observe the growth and multiplication of the microorganisms. Three uninoculated teeth were maintained in a humid environment as an aseptic control. These teeth were immersed in BHI and maintained at 37 degrees C for 7 days to determine microbial growth. Bacterial growth was shown by turbidity of the culture medium and confirmed by seeding these broths on BHI agar at 37 degrees C for 24 h. The positive BHI tubes were selected, and inoculum was spread on the surface of BHI agar, followed by the same incubation conditions. Gram stain was conducted from BHI growth and from colonies growing on solid medium. Calcium hydroxide in infected dentinal tubules showed no antimicrobial effect on S. faecalis, S. aureus, B. subtilis, P. aeruginosa, or on the bacterial mixture used throughout the experiment. PMID- 10530242 TI - Evaluation of the cytocompatibility of three endodontic materials. AB - The goal of this in vitro study was to evaluate the relative cytocompatibility of three endodontic materials: calcium hydroxide, a calcium oxide-based compound, and a zinc oxide-eugenol-based sealer. The evaluation was conducted 24, 72, and 168 h after contact with the compounds and involved three complementary techniques: a colorimetric cytotoxicity test, scanning electron microscopy, and flow cytometry. The results we obtained confirmed the initial cytotoxicity of the zinc oxide-eugenol-based sealer and showed that the calcium oxide-based compound had the same relative cytocompatibility as calcium hydroxide. PMID- 10530243 TI - The combined occluding effect of sodium fluoride varnish and Nd:YAG laser irradiation on human dentinal tubules. AB - Various methods and materials used in the treatment of dentin hypersensitivity are thought to achieve a therapeutic benefit by tubule occlusion. The aim of the present study was to evaluate the combined occluding effect of sodium fluoride varnish and Nd:YAG laser irradiation on human dentinal tubules. Thirty-six dentin specimens with exposed dentinal tubule orifices were used in this study. The samples were randomly divided into four groups. Groups A, B, and C were varnished by sodium fluoride, whereas group D served as a control. Then, group C was lased by 30 mJ of Nd:YAG laser, 10 pulses/s for 2 min by light painting. Three hours later, groups B and C were brushed by an electrical toothbrush for 30 min. Under SEM observation, the control group showed numerous exposed dentinal tubule orifices, and the sodium fluoride varnished specimens showed closure of exposed dentinal tubule orifices. After electrical toothbrushing, most of the sodium fluoride varnish was brushed away, except in the specimens that were irradiated by Nd:YAG laser. Over 90% of the dentinal tubule orifices were occluded by sodium fluoride varnish combined with Nd:YAG laser irradiation. PMID- 10530244 TI - Comparison of cutting efficiency and instrumentation of curved canals with nickel titanium and stainless-steel instruments. AB - The cutting efficiency and the effects of instrumentation on curved canal shape of both stainless-steel and nickel-titanium nonstandardized ProFile Series 29 hand instruments and stainless-steel Flexoreamer were investigated under standardized conditions. Concerning cutting efficiency in rotary motion, the Flexoreamer had significantly (p < 0.01) greater cutting efficiency than stainless-steel ProFiles and nickel-titanium ProFiles. Changes in the canal shape differed significantly between the different instruments at all measuring points (p < 0.001). After instrumentation with Flexoreamers with inclusion of two half sizes (#17 and #22), there were fewer undesirable changes in canal shape compared with both stainless-steel and nickel-titanium ProFile Series 29 instruments. In this study, it seemed that flexible stainless-steel instruments with noncutting tips were superior to the nonstandardized ProFile Series 29 instruments with regard to cutting efficiency and instrumentation of curved canals. PMID- 10530245 TI - Dentinal tubule penetration of root canal sealers after root canal dressing with calcium hydroxide. AB - The purpose of this study was to evaluate the dentinal tubule penetration of root canal sealers after root canal dressing with calcium hydroxide (Ca(OH)2). Forty two single-rooted teeth were instrumented to size 60. Six teeth served as the control group, and the remaining teeth were assigned to two groups. Root canals of the first group were filled with the Ca(OH)2 paste; the second group was filled with TempCanal, and all were incubated for 7 days. The samples were either irrigated with only NaOCl or with EDTA, followed by NaOCl to remove Ca(OH)2. All of the teeth were obturated with CRCS, AH26, and Ketac Endo by a lateral condensation technique. The specimens were then kept at the same conditions for another 7 days, and then all of the roots were prepared for scanning electron microscopic evaluation. Scanning electron microscopic examination revealed that Ca(OH)2 was not completely removed from the root canal surfaces, and root canal sealers did not penetrate into the dentinal tubules when only NaOCl was used. EDTA followed by NaOCl irrigation resulted in complete removal of Ca(OH)2 and root canal sealers penetrated into the dentinal tubules. PMID- 10530247 TI - An analysis of canal centering using mechanical instrumentation techniques. AB - The purpose of this study was to compare canal transportation in moderately curved canals using mechanical instrumentation systems. Mesial roots of mandibular first or second molars were mounted in resin using a modified Bramante muffle system and divided into four groups. The roots were cross-sectioned 2 mm from the working length and at the height of root curvature. Tracings of the canal were made from preinstrumentation slides of the cross-sections. The canals were prepared using ProFile Series 29 rotary instruments, Quantec 2000 rotary instruments, Flex-R files in the Endo Gripper contra-angle handpiece, and Shaping Hedstrom files in the M4 contra-angle handpiece. Tracings of the prepared canals were made onto the originals from postinstrumentation slides. A canal centering ratio was calculated along the line of maximum transportation. Quantec 2000 rotary instruments yielded significantly greater transportation at the apical level when compared with the Profile Series 29 system. There were no other significant differences in transportation at either level. There were no differences in the direction of canal transportation between instrument systems, and the direction of canal transportation was not related to the direction of canal curvature. Canal preparation time was shortest with Profile Series 29 system followed by Flex-R files in the Endo Gripper, Quantec 2000, and Shaping Hedstrom files in the M4. PMID- 10530246 TI - Dynamic and cyclic fatigue of engine-driven rotary nickel-titanium endodontic instruments. AB - The absence of adequate testing standards for engine-driven nickel-titanium (NiTi) instruments necessitates further study of these instruments in all areas. This study examined three groups of engine-driven rotary NiTi endodontic instruments (Profile, Hero, and Quantec) and assessed the times for dynamic fracture in relation to the radius of curvature to which the instruments were subjected during preparation, with the instrument diameter determined by size and taper and the mode by which the fracture occurred. Ten instruments were randomly selected representing each size and taper for each group and for each radius of curvature: 600 in total. The instruments were rotated at 350 rpm and introduced into a tempered steel curve that simulated a canal. Two radii of curvature of canals were used: 5 and 10 mm. Time at fracture was noted for all files, and the fracture faces of each file were analyzed with scanning electron microscopy. Radius of curvature was found to be the most significant factor in determining the fatigue resistance of the files. As radius of curvature decreased, fracture time decreased. Taper of files was found to be significant in determining fracture time. As diameter increased, fracture time decreased. In all cases, fracture was found to be of a ductile nature, thus implicating cyclic fatigue as a major cause of failure and necessitating further analyses and setting of standards in this area. PMID- 10530248 TI - Canal morphology of maxillary molars: clinical observations of canal configurations. AB - An examination of 1732 conventionally treated maxillary molars was made in an attempt to determine the percentage of MB2 canals that could be located routinely. The teeth examined were 1096 first molars, 611 second molars, and 25 third molars. The results were recorded on a modified computer program over an 8 yr period of time. An interesting trend was noted. The MB2 canal was found in 802 (73.2%) first molars, 310 (50.7%) second molars, and 5 (20.0%) third molars. It occurred as a separate canal in 54.9% of first molars, 45.6% of second molars, and joined in all third molars. However, as the operator became more experienced, scheduled sufficient clinical time, routinely employed the dental operating microscope, and used specific instruments adapted for microendodontics, MB2 canals were located in 93.0% of first molars and 60.4% in second molars. PMID- 10530249 TI - The endodontic autopsy: a valid learning tool. AB - Clinical success rates in endodontics can be improved if the causes of failures can be determined, prevented, or corrected. The endodontic autopsy is recommended as a method for determining preoperative, operative, and postoperative causes of failures in endodontics. PMID- 10530251 TI - A comparison of instrumentation using Naviflex and Profile nickel-titanium engine driven rotary instruments. AB - This study was designed to compare the changes in canal configuration resulting from instrumentation by either Profile or Naviflex instruments. Forty mesial canals in extracted human molar teeth were embedded and sectioned at two root levels. Reassembled teeth were instrumented with a modified crown-down technique as described in the Profile training video for Profile files and in a similar manner for Naviflex instruments. Superimposed pre- and postinstrumented cross sectional root images were projected, traced, and scanned into a computer for analysis. Canal movement, in relation to the furca, and canal area change were recorded. The results showed no significant difference in canal center movement or canal area change between the Profile or Naviflex groups. The degree of canal curvature had no effect on canal center movement or canal area change. PMID- 10530250 TI - Microleakage between endodontic temporary restorative materials placed at different times. AB - Occlusal endodontic access preparations are occasionally made in teeth without removing the original restoration. However, microleakage between restorative materials that are placed at different times has not been extensively studied. Therefore, our objective was to compare microleakage at three areas: between an access opening restorative material and the cavity wall; between an additional material placed later to patch a secondary opening in the first restorative material and the original restorative material itself; and between the secondarily placed material and the cavity wall. Standard endodontic access preparations were made in 120 noncarious, nonrestored crowns of extracted human molars. These teeth were divided into six experimental groups. Another four molars were controls. The endodontic access cavities were restored with either IRM or amalgam as the primary restorative material. After 14 days, half of the primary restorations was removed, and this defect was filled with a secondary restorative material: IRM, Caviton, or a double seal of Caviton and IRM. Microleakage was measured linearly as the extent of basic fuchsin dye penetration under a stereomicroscope after thermal cycling (5 degrees and 55 degrees C for 100 cycles) and tooth sectioning. Wilcoxon signed-rank test was used for statistical analysis. Results indicated significantly less microleakage between primary and secondary restorative materials placed at different times than microleakage between primary temporary restorative materials and the access cavity wall, regardless of the type of primary restorative material used (IRM or amalgam). PMID- 10530252 TI - Quality of obturation in student cases instructed by endodontic versus general dentistry faculty. AB - Endodontic literature suggests that only about 60% of endodontic therapy meets current technical standards and that general dentists may be making a significant contribution to this compromised care. If so, where in the continuum of dental education does this begin. This study evaluated the quality of obturation in mandibular molars provided on the one hand by 3rd year dental students instructed by endodontic faculty, and on the other hand by 4th year students instructed by general dentistry faculty, versus the quality of obturation achieved by endodontic residents who served as a control for both groups. Final radiographs were chosen from students in all three groups so that there were 22 samples per group. Three evaluators rank-ordered the radiographs. In order of excellence, the results were: (a) residents, (b) 3rd year students, and (c) 4th year students. There was no significant difference between the 3rd year students or the residents, only between residents and 4th year students (p < 0.05). The reasons for this outcome may range from dental school objectives to the private practice procedures of the general dentists who instructed the 4th year students. PMID- 10530253 TI - Concerning ask a friend: Case #5, November 1998: pulpal versus periodontal pain? PMID- 10530254 TI - Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and polymerase chain reaction for differentiation of Prevotella intermedia and Prevotella nigrescens. AB - Isolates previously thought to be Prevotella intermedia have been shown to be a closely related species now known as Prevotella nigrescens. The purpose of this study was to determine the efficacy of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and polymerase chain reaction (PCR) to differentiate endodontic isolates of P. nigrescens from P. intermedia. Fifty-six strains of black-pigmented bacteria isolated from endodontic infections and conventionally identified as P. intermedia were used in this study. Using SDS-PAGE, novel polypeptide bands were used to differentiate P. nigrescens from P. intermedia. PCR was accomplished with specific primers for the 16S ribosomal RNA gene of both strains. Of 56 endodontic isolates, 41 (73%) strains were identified by SDS-PAGE as P. nigrescens and 15 (27%) strains as P. intermedia. Of the 41 strains of P. nigrescens identified by SDS-PAGE, PCR identified 37 strains as P. nigrescens. Restriction endonuclease digestion of amplified 16S ribosomal RNA genes indicated that the remaining four strains originally identified by SDS-PAGE as P. nigrescens were actually strains of Prevotella distinct from P. nigrescens and P. intermedia. Of 15 strains of P. intermedia identified by SDS-PAGE, PCR identified 14 strains as P. intermedia; but, one strain was identified as P. nigrescens. The results indicated that PCR was a more precise method than SDS-PAGE to differentiate P. intermedia from P. nigrescens. This study confirms that P. nigrescens is more commonly isolated in pure culture from endodontic infections than P. intermedia. PMID- 10530255 TI - pH changes and calcium ion diffusion from calcium hydroxide dressing materials through root dentin. AB - The purpose of this study was to evaluate Ca2+ and OH- diffusion properties through root dentin by using different calcium hydroxide (CH) dressing materials. Twenty-eight single-rooted teeth were instrumented and external defects were created on the root surface. 17% EDTA was used to eliminate the smear layer. All surfaces except the external defects were sealed, and the teeth were placed in normal saline. Ca2+ concentrations and the pH in the saline were determined for 3 days as the control period. After removing the teeth from normal saline, they were filled with: (i) DT Temporary Dressing CH; (ii) CH powder and normal saline; (iii) TempCanal; and (iv) CH points. The teeth were then placed in normal saline, and Ca2+ concentrations and pH values were measured at 1, 3, 7, 14, and 28 days. Nonsetting CH pastes gradually released Ca2+, whereas this increase was absent from CH points. None of the test materials induced a pH increase in the media during the observation period. This study demonstrated that, when nonsetting CH pastes are applied to the root canal, diffusion of Ca2+ without an increase in pH to the surrounding media occurs. This implies that these type of material are more suitable than CH points for treatment of external root resorption. PMID- 10530256 TI - Mechanical reduction of the bacterial population in the root canal by three instrumentation techniques. AB - The in vitro reduction of the bacterial population in the root canal by the mechanical action of instrumentation and irrigation was evaluated. Root canals inoculated with a Enterococcus faecalis suspension were instrumented using hand Nitiflex files, Greater Taper (GT) files, and Profile 0.06 taper Series 29 rotary instruments. Irrigation was performed using sterile saline solution. Root canals were sampled before and after instrumentation. In the group of the Nitiflex files, samples were also taken after each file size. After serial dilution, samples were plated onto Mitis-Salivarius agar, and the colony forming units grown were counted. All techniques and instruments tested were able to reduce significantly the number of bacterial cells in the root canal. Instrumentation to a Nitiflex #30 was significantly more effective than GT files. There were no significant differences when comparing the effects of the Profile instrument #5 with either the GT files or the Nitiflex #30. Enlargement to a Nitiflex #40 was significantly more effective in eliminating bacteria when compared with the other techniques and instruments tested (p < 0.05). The results of this study showed that the instrumentation and irrigation can mechanically remove more than 90% of bacterial cells from the root canal. PMID- 10530257 TI - Molecular detection of Bacteroides forsythus in infected root canals. AB - The purpose of the present study was to investigate the occurrence of Bacteroides forsythus in infections of dental root canals. Eleven samples from infected root canals were analyzed by four different molecular methods. The prevalence of the monitored species varied as a function of the detection method. The polymerase chain reaction-DNA probe method after immunocapture yielded the highest prevalence value (6/11), whereas the lowest value was observed with the slot-blot (3/11). Of the 11 canal samples, 5 were positive by ELISA and 4 were positive by immunofluorescence. The presence of B. forsythus was detected by all four methods in 3/11 canals, whereas 4/11 appeared to be free of B. forsythus. Our data indicate that B. forsythus can be part of the endodontic microflora. The procedure consisting of immunomagnetic capture and a polymerase chain reaction DNA probe assay can be useful as an alternative to culture for clinical studies of the species infecting human dental pulp. PMID- 10530258 TI - In vitro effect of the resin component bisphenol A on substrate adherence capacity of macrophages. AB - This study was design to investigate the "in vitro" effect of bisphenol A (BPA), a component of resin used in dentistry, on viability, and substrate adherence capacity of macrophages. Peritoneal macrophages were obtained from Wistar rats and resuspended in RPMI-1640 medium. Viability was determined by trypan blue exclusion. As a test of macrophage adhesion, the adherence capacity of macrophages to a plastic surface was determined and the adherence index was calculated. Assays were conducted in Eppendorf tubes for 60 min of incubation at 37 degrees C in a humidified atmosphere of 5% CO2 in air. BPA did not alter significantly macrophage viability at concentrations as high as 10(-5) M, but BPA decreased in a dose-dependent manner the adherence index of rat peritoneal macrophages. Control peritoneal macrophages showed an adherence index = 81.5 +/- 7.9%. In the presence of 10(-8) M BPA, the Al of macrophages decreased to 41.4 +/ 12.2% (p < 0.05). Higher BPA concentrations (10(-7) to 10(-5) M) also caused a significant inhibition of the adherence index. Half-maximal inhibition (IC50) was obtained at 4.92 +/- 0.39 x 10(-6) M BPA. The in vitro study shows that the resin component BPA can alter macrophage adhesion. Taking into account that adhesion is the first step in the phagocytic process of macrophages and in antigen presentation, BPA could inhibit macrophage function and modulate immune and inflammatory responses in dental pulp and periapical tissues. PMID- 10530259 TI - Endodontic treatment of teeth with apical periodontitis: single vs. multivisit treatment. AB - This study was performed to evaluate radiographic healing of teeth with apical periodontitis, treated in one visit or in two visits (a) with or (b) without calcium hydroxide as an intracanal disinfecting medicament. The patients were assigned one of the three treatment groups by the throwing of a die. The Periapical Index (PAI) Scoring Method was used to compare differences in periapical status from the beginning of treatment to a 52-wk follow-up evaluation. Overall, the periapical status of the treated teeth improved significantly after 52 wk (p < 0.0001). A PAI score of 1 or 2 was considered as representing a "good" periapical status while 3, 4, or 5 was a "bad" status. When base line PAI scores were controlled for, the calcium hydroxide group showed the most improvement in PAI score (3, 4, or 5 to 1 or 2), followed by the one-step group (74% vs. 64%). The teeth that were left empty between visits had clearly inferior healing results. Power statistics were conducted to determine the numbers required for significant differences between the groups, and it was shown that large experimental groups on the order of hundreds of patients would be required to show significant differences. PMID- 10530260 TI - An evaluation of the action of different root canal irrigants on facultative aerobic-anaerobic, obligate anaerobic, and microaerophilic bacteria. AB - The aim of this study was to test the effect of different concentrations of sodium hypochlorite, chlorhexidine, and cetrimide on the following bacterial strains: Facultative aerobes-anaerobes: Candida albicans ATCC 10231; Enterococcus faecalis ATCC 29212; Escherichia coli ATCC 25,922; Pseudomonas aeruginosa ATCC 27,853; Streptococcus mitis ATCC 9811; Streptococcus mutans ATCC 35668; Streptococcus salivarius ATCC 13419; and Streptococcus sanguis ATCC 10556. Microaerophiles: Actinobacillus actinomycetemcomitans ATCC 29522. Obligate anaerobes: Actinomyces odontolyticus ATCC 17929; Fusobacterium nucleatum ATCC 25,586; Porphyromonas gingivalis ATCC 33277; and Prevotella melaninogenica ATCC 25845. Each irrigant was kept in contact with the bacterial species used for the experiment for 10, 20, or 30 min. Results showed that all irrigants had a bactericidal effect on both facultative aerobes-anaerobes and on microaerophilic and obligate anaerobic strains, in all concentrations and even after short periods of contact. PMID- 10530261 TI - Morphological alterations associated with the cytotoxic and cytostatic effects of citric acid on cultured human dental pulp cells. AB - Citric acid exerts potential harmful effects on the pulp when used for root surface demineralization, smear layer removal, and dentin etching. Using 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay, we found that incubation of cultured human dental pulp cells in medium containing 0.5% (pH 4.74) or 1.0% (pH 3.42) of citric acid for 2 h lead to 25% and 48% of cell death, respectively. Cytotoxicity of citric acid was associated with its acidity. Exposure of cells to pure 1% citric acid (pH 2.26) for 60 s lead to immediate cell death. Cytotoxicity was usually preceded by cell retraction, cell surface blebbing, and finally uptake of trypan blue, implicating the presence of cell membrane damage. A medium containing 0.05% citric acid can retard the growth of pulp cells. These results indicate that adequate protection of the pulp is important, especially when the remaining dentin is thin in deep carious lesions or in the presence of accessory canals on the root surface. PMID- 10530263 TI - Histological periapical repair after obturation of infected root canals in dogs. AB - Histological periapical healing of infected roots obturated in one-step or with prior calcium hydroxide (Ca(OH)2) disinfection was compared. Seventy-two roots of vital dog teeth were instrumented to ISO size 45. Sixty roots were infected with dental plaque and closed. Six weeks later, apical periodontitis was radiographically confirmed in the infected roots. The teeth were divided into the following groups: group 1, one-step (n = 24)-roots were irrigated with 10 ml of saline, obturated, and permanently restored; group 2, Ca(OH)2 (n = 24)-roots were treated as in group 1, except that after saline irrigation, Ca(OH)2 dressing was placed in the canal for 1 wk before obturation; group 3, positive control (n = 12)--the roots were irrigated with saline, but the canals were not obturated; and an additional group, group 4, served as a negative control (n = 12)--these teeth that were not infected with plaque were aseptically obturated. The dogs were sacrificed after 6 months. The roots and surrounding apical tissues were prepared and histologically examined by two independent evaluators blinded to the treatment groups. A two-way ANOVA test demonstrated that the four treatment groups were significantly different from one another. The positive control showed the most inflammation, the negative control the least, and the Ca(OH)2 group had significantly less inflammation than the one-step group (p < 0.05). It is concluded that Ca(OH)2 disinfection before obturation of infected root canals results in significantly less periapical inflammation than obturation alone. PMID- 10530262 TI - Cytotoxic and mutagenic potencies of various root canal filling materials in eukaryotic and prokaryotic cells in vitro. AB - Cytotoxic and mutagenic effects of root canal filling cements of various chemical composition were determined in vitro. Materials set for 24 h and 1 wk were eluted for 24 h in cell culture medium (cytotoxicity testing) and dimethyl sulfoxide or physiological saline (mutagenicity testing). The differences between cytotoxic potencies of eluates of the endodontic materials on L-929 cells were quantified colorimetrically (MTT test). Eluates of Traitement SPAD were about 5- to 30-fold more toxic than silver-free AH26, Tubli-Seal, CRCS, and Endomethsone N. The rank order of the toxic effects depended on the setting time of mixed materials. Dimethyl sulfoxide and saline eluates of Traitement SPAD, Tubli-Seal, Endomethasone N, CRCS, and Ketac-Endo were not mutagenic in the Ames test. Both eluates of silver-free AH26 set for 24 h were weakly mutagenic in Salmonella typhimurium TA100. Weak mutagenicity of saline eluates of the material was also observed in TA97a and TA102. These results point to the possibility that mixed silver-free AH26 might contain small amounts of two mutagenic substances: bisphenol A diglycidyl ether and formaldehyde. PMID- 10530264 TI - Recommendations for endodontic referral among practitioners in a dental HMO. AB - This study assessed the effect of patients' presenting conditions on general practitioners' (GPs') self-reported endodontic referral patterns, and compared GPs' perceived indications for referral with those of endodontists. The study was based on a self-administered, confidential survey distributed to 79 GPs and 7 endodontists who provide care to members of one Dental HMO in the Pacific Northwest. GPs were most likely to recommend referral for teeth they felt needed surgical retreatment, but GPs and endodontists did not always agree on indications for referral. Compared with GPs, endodontists were more likely to recommend referral for patients with complex problems, but not necessarily technically difficult teeth. Compared with those with less experience, GPs with more than 10 yr both in dentistry and at this HMO were more likely to recommend (a) referring difficult cases rather than performing endodontic therapy themselves and (b) extracting perforated or root-fractured teeth prior to obturation rather than continuing treatment. Indications for referral that maximize favorable dental outcomes need to be identified. PMID- 10530265 TI - Comparison of the obturation of lateral canals by six techniques. AB - The following obturation techniques were compared on their ability to obturate lateral canals in vitro, lateral condensation (LC), continuous wave of condensation (CW), warm vertical condensation (WV), carrier-based thermoplasticized gutta-percha (CB), warm lateral condensation (WL), and vertically condensed high-temperature gutta-percha (HT). A root canal system with lateral canals in the coronal, middle, and apical thirds was prepared in resin blocks. Each block was obturated using each technique (n = 30, 15 each, with and without sealer). The length of gutta-percha and sealer in each of the lateral canals was measured with a measuring microscope and statistically compared. All techniques obturated all three levels of lateral canals with sealer. WV, CB, and CW were able to fill the lateral canals with gutta-percha significantly better when root canal sealer was used. WV, CB, CW, and HT filled the coronal and middle lateral canals significantly better with gutta-percha than LC or WL condensation. CB and CW filled the apical lateral canal significantly better with gutta-percha than HT, WV, WL, or LC. PMID- 10530266 TI - Comparison of effect of intracanal use of ketorolac tromethamine and dexamethasone with oral ibuprofen on post treatment endodontic pain. AB - The purpose of this study was to compare the pain-reducing efficacy of dexamethasone and ketorolac tromethamine when used as an intracanal medication, with oral ibuprofen and a placebo. An additional objective was to establish if any relationship exists between the incidence and severity of pretreatment pain and the incidence and severity of postinstrumentation pain. A total of 48 patients who presented to the University of Illinois postgraduate endodontic clinic were invited to participate. Patients were randomly assigned to 1 of 4 groups: oral ibuprofen, placebo, dexamethasone, or ketorolac tromethamine. Patients were asked to evaluate their pretreatment pain when they presented to the clinic with a Visual Analog Scale. The root canal treatment was performed in two appointments. The first appointment consisted of cleansing and shaping of the canal/s and placement of an intracanal medication. All teeth were closed with a sterile cotton pellet and IRM. Each patient was sent home with a Visual Analog Scale to fill out at 6, 12, 24 and 48 h after initiation of therapy. At the 12-h period, both dexamethasone and ketorolac provided statistically significant better pain relief than placebo. At the 24-h period, only ketorolac demonstrated better pain relief than the placebo. There were no statistically significant differences among the groups at 6 and 48 h. Although ibuprofen pain ratings were less than the placebo at all time points, the reduction was not significant. In addition, no significant differences were demonstrated between ibuprofen and either dexamethasone or ketorolac. PMID- 10530267 TI - An evaluation of plaster of Paris barriers used under various materials to repair furcation perforations (in vitro study). AB - The present study was undertaken to evaluate the effect of plaster of Paris barriers on the sealing ability of various materials when used to repair furcation perforations. The study was conducted on 90 human permanent molars with well developed, nonfused roots. Access openings and furcation perforations were prepared in the pulp chamber floor. The teeth were divided into one control and five experimental groups of 15 teeth each. In the experimental group, plaster of Paris barriers were created in the perforations and then sealed with silver amalgam, glass ionomer (GI), self-cured composite resin, IRM, or AH26. Access openings were filled with composite resin. The teeth were submerged in a solution of 2% methylene blue dye for 2 wk. Finally, the samples were sectioned and evaluated for linear dye leakage and analyzed statistically. Maximum dye penetration was observed in amalgam followed in a decreasing order by GI, composite, IRM, and AH26. AH26 showed the best sealing ability in the presence of plaster of Paris barriers followed by IRM, composite resin, and GI. Amalgam showed the poorest sealing ability when used to repair furcation perforations. PMID- 10530268 TI - Endo-Antral syndrome and various endodontic complications. AB - The purpose of this paper was to examine the varied impact of the pathological spread of dental sepsis into the adjacent maxillary sinus. This complex of tissue destruction is called Endo-Antral Syndrome; the usual radiographic diagnostic features are identified in the paper. The four different cases presented serve to illuminate a few of the many diagnostic and treatment challenges involved. Emphasis is placed on the utilization of a keen sense of wariness when endodontically treating maxillary posterior teeth whose apexes are close to the sinus. Dental examination should include an appraisal of antral health prior to root canal therapy to best plan treatment and to establish a base line against which to judge subsequent developments. PMID- 10530269 TI - P-ASA block injection: a new palatal technique to anesthetize maxillary anterior teeth. AB - This technique article presents a new local anesthetic injection that is reported to produce anesthesia of the six maxillary anterior teeth, the anterior third of the palate, and the facial gingiva from a single-site injection. The injection is referred to as the palatal approach anterior superior alveolar (P-ASA) nerve block. The 0.9 to 1.4 mL dosage recommendation for this block injection is significantly less than for a traditional supraperiosteal approach. The primary advantage of this injection is that it allows the dentist to anesthetize the teeth and gingiva without collateral anesthesia to the lips, face, or muscles of facial expression. Therefore, the smile line is not distorted during the operative phase of an appointment, and the patient is more comfortable postoperatively. CLINICAL SIGNIFICANCE: The P-ASA is a new block injection technique that provides anesthesia of the maxillary anterior teeth from a single injection without numbness of the face, lips, and muscles of facial expression. This injection technique prevents distortion of the smile line and enhances restorative procedures that use the lip line as an esthetic reference element. PMID- 10530270 TI - Ceramic restorations for posterior teeth: guidelines for the clinician. AB - Metal-free ceramic restorations are increasingly popular for restoring posterior teeth. These restorations are generated through a variety of techniques (e.g., CAD-CAM, copy-milling, heat-pressing, and firing). When appropriately indicated and made, ceramic inlays or onlays can be reliable and provide a highly serviceable restoration. The aim of this article is to review and present updated information regarding indications, restorative technique, and maintenance for this class of restorations, with emphasis to fired ceramic inlays or onlays. The information presented is based on 15 years of controlled clinical experience with this category of restorations. Clinical cases that represent some applications also are presented. CLINICAL SIGNIFICANCE: The fired ceramic inlay/onlay technique is presented as a viable option for the esthetic and adhesive restoration of posterior teeth. PMID- 10530271 TI - Colorimetric assessment of laser and home bleaching techniques. AB - PURPOSE: This study recorded in vitro color change of three tooth bleaching techniques that included laser-activated hydrogen peroxide and two concentrations of carbamide peroxide. MATERIALS AND METHODS: Forty extracted human central incisors were exposed to argon laser-activated 35% H2O2, 10% carbamide peroxide, or 20% carbamide peroxide. A fourth group (control) did not receive any bleach treatment (n = 10/group). Commission International de l'Eclariage (CIE) L*a*b* coordinates were recorded prior to bleaching (baseline), at 1 week, and at 2 weeks. The color difference (delta E*ab) between baseline and subsequent measurements was calculated. RESULTS: The control group did not demonstrate significant color difference over time (p > .05). The laser group was not statistically different from the control group (p > .01). The color difference of the 10% and 20% carbamide peroxide groups was statistically different from the control group (p < .01). CLINICAL SIGNIFICANCE: Exposure to 20% carbamide peroxide produced the greatest perceivable change in color. The recommended one time application of laser-activated hydrogen peroxide did not demonstrate any perceivable color change. The clinician should be aware that additional or longer applications may be required. PMID- 10530272 TI - Efficacy of nightguard vital bleaching technique beyond the borders of a shortened tray. AB - PURPOSE: The purpose of this study was to determine if bleaching extends beyond the borders of a shortened tray or if a demarcation line is found. MATERIALS AND METHODS: Fifteen extracted teeth were selected darker than B2 on a Vita Lumin Shade guide (Vita Lumin, Bad Sackingen, Germany). The teeth were mounted in arch like fashion in dental stone. Alginate impressions were made, and a stone replica of the four arches of teeth was generated. Vacuum-formed bleaching trays were fabricated for each arch, without and with reservoirs, as per the product to be tested. The nonreservoir trays were trimmed to one half the clinical crown length, and the reservoir trays were trimmed 1 mm beyond the border of the half length reservoir. Measurements were taken from the cementoenamel junction (CEJ) to the tray border for each tooth. The trays averaged 5.1 mm short of the CEJ. The reservoir group was treated with viscous glycerin-based 10% carbamide peroxide (Opalescence, Ultradent Products Inc., South Jordan, Utah); the nonreservoir group with creamy dentifrice-based 10% carbamide peroxide (Platinum, Colgate Oral Pharmaceuticals, Canton, Massachusetts). The trays were loaded and fully seated on the teeth. Excess bleaching material was removed with a toothbrush and water rinse. Each assembly was placed in a humidor with incisal edges down to simulate oral conditions. The process was repeated for fourteen 6- to 8-hour bleaching sessions. Blinded and nonblinded operators determined post bleaching shades, with consensus reached on differing shades. RESULTS: All teeth demonstrated lightening of 1 to 10 (mean 5.2) increments on the value-oriented shade guide. The bleaching effect extended beyond the tray and no demarcation lines were noted on any of the teeth. CLINICAL SIGNIFICANCE: This in vitro study suggests that successful bleaching occurs beyond the borders of inadvertently shortened trays. The clinician does not necessarily need to remake the tray if the tray does not cover all portions of the tooth. PMID- 10530273 TI - Shear bond strengths of one-bottle adhesives to moist enamel. AB - PURPOSE: This study evaluated bond strengths of six one-bottle bonding agents and a control (primer plus unfilled resin) to moist enamel. MATERIALS AND METHODS: One-hundred and five bovine teeth were randomly assigned to seven groups of 15. Enamel was etched for 15 seconds with 35% phosphoric acid. Etched enamel was rinsed, and excess water was blotted with tissue paper. Following application of the adhesive, composite resin was bonded using a gelatin capsule technique. Shear bond strengths to enamel were determined using a universal testing machine (Instron Corp., Canton, Massachusetts). RESULTS: Mean bond strengths ranged from 21.9 MPa for OptiBond Solo (Kerr Corp., Orange, California) to 29.6 MPa for Prime & Bond 2.1 (Dentsply/Caulk, Milford, Delaware). Prime & Bond 2.1 had a significantly higher mean bond strength than the other adhesives. CLINICAL SIGNIFICANCE: The results of this study suggest that all of the one-bottle systems tested should provide clinically acceptable bonding to moist enamel. PMID- 10530274 TI - Effect of cutting instruments on permeability and morphology of the dentin surface. AB - There are major differences in morphological detail after cutting the dentin surface among the methods commonly used to prepare dental cavities. The purpose of this study was to compare dentin permeability and the morphology of the dentin surfaces prepared with diamond and carbide steel burs after etching with 6% citric acid. Twenty-four freshly extracted human third molars were sectioned, mounted on plexiglass, and connected to the dentin-permeability measuring apparatus. The permeability of dentin was measured by fluid filtration and expressed as hydraulic conductance. There were two study groups of 12 teeth. Each tooth had one occlusal cavity preparation prepared but utilized three depths: the original was prepared just into the dentin, the second 0.5 mm deeper than the first, and the third 0.5 mm deeper than the second. One group had the first cavity prepared with a diamond, the second deepened using a steel bur, then the third depth was made by use of the diamond. The other group had the first cavity preparation prepared with a steel bur, deepened 0.5 mm again using a diamond, then deepened again using a steel bur. Dentin permeability was measured after cavity preparation, then after 2 minutes of acid etching. Analysis of variance and Duncan's multiple range test were used to establish whether differences were significant at the 0.05 confidence level. Prepared and acid-etched surfaces were characterized using a scanning electron microscope to identify any differences between the two groups. After acid etching with 6% citric acid, the permeability of dentin cavities prepared with diamond burs was significantly less than the permeability of cavities prepared with carbide steel burs. After etching, there were differences in the appearance of the dentin surfaces prepared with diamonds and steel burs. Dentin bonding agents may have their effectiveness reduced when placed following cavity preparation by use of a diamond. PMID- 10530275 TI - Marginal adaptation of heat-pressed glass-ceramic veneers to dentin in vitro. AB - The purpose of the present study was to examine the marginal adaptation of ceramic veneers to dentin at the cervical margins and to enamel at the palatoincisal margins using four dual-curing composite resin cements of different viscosity with their corresponding dentin bonding systems. Thirty-six caries-free human maxillary incisors were prepared for facial ceramic veneers with cervical cavity margins located in dentin. Heat-pressed glass-ceramic veneers (IPS Empress) were inserted adhesively using one of the following luting systems: Sono Cem (SC) with EBS; Variolink Ultra (VU), Variolink High Viscosity (VHV), and Variolink Low Viscosity (VLV) with Syntac. Both the cervical and the palatoincisal margins of the veneers (tooth/composite resin cement interface and ceramic/composite resin cement interface) were evaluated before and after thermocycling and mechanical loading (TCML) by quantitative margin analysis under a scanning electron microscope (SEM) using an image analysis system. Microleakage was assessed by dye penetration after TCML. Before TCML, SC and VU showed statistically significantly fewer marginal gaps than VHV and VLV. After TCML, SC, VU, and VHV revealed significantly fewer marginal gaps than VLV. TCML had a statistically significant influence on marginal gap formation at both the dentin and enamel margins. After TCML, the percentage of marginal gaps was not significantly different at the cervical dentin than at the palatoincisal enamel margins. Cervical dye penetration after TCML showed no statistically significant differences in microleakage among the four luting systems. In conclusion, this in vitro study showed that similarly favorable marginal adaptations of ceramic veneers to dentin and enamel can be achieved using Sono-Cem, Variolink Ultra, or Variolink High Viscosity with their corresponding dentin bonding systems. PMID- 10530276 TI - Dentin bond strength and marginal adaptation: direct composite resins vs ceramic inlays. AB - The aim of this in vitro study was to compare the dentin bond strength and marginal adaptation of directly and indirectly inserted restorations. A conically modified push-out test was designed to consider polymerization shrinkage and facilitate inlay placement. A total of 260 cavities were prepared into disks of freshly extracted human third molars and filled with direct composite resins or with adhesively luted ceramic inlays. Dentin adhesives of the third--(with self etching primer: ART Bond, Syntac Classic), fourth--(with total etching: Scotchbond Multi-Purpose Plus), and fifth-generation (one-bottle adhesives: Syntac Single Component, Prime & Bond 2.1) were used in combination with one hybrid composite (Tetric) or luting composite (Variolink Low). Control groups did not use an adhesive. Polymerization of the bonding agent was carried out prior to insertion of the filling/inlay or afterwards simultaneously with the composite/luting composite. The thickness of the adhesive layer and luting composite was recorded, and after 7 days of storage and 24 hours of thermocycling (1150 cycles) replicas were made and extrusion testing performed. Fracture modes were determined and replicas were examined regarding marginal adaptation using SEM (X200 magnification). Precuring of the bonding resin increased dentin bond strength independent of the material combination or insertion mode (P < 0.05). In general, third- and fourth-generation dentin adhesives produced better results in bond strength and marginal adaptation than one-bottle systems (P < 0.05). In the third generation, ART Bond achieved significantly higher push-out values than Syntac (P < 0.05), but no better marginal adaptation. Cohesive fractures within the dentin were only observed in the inlay groups with precured resin. Precuring of the bonding resin is an important factor for both direct and indirect restorations. Nevertheless, precuring of the bonding resin prior to insertion of adhesive inlays cannot be recommended clinically, because the 120-micron luting spaces were too large. In simulated cavities, direct composite fillings with precuring achieved bond strengths similar to inlays without precuring. One-bottle adhesive systems performed poorly compared with multi-step adhesives of the third and fourth generation. PMID- 10530277 TI - Shear bond strength of repaired composite resins using a hybrid composite resin. AB - The shear bond strength of different types of composite resins repaired with a hybrid composite was evaluated. The hybrid composite resin was repaired with itself as a control. The results of this study showed that the shear bond strength of the repair with most types of composite resins was minimally adequate. PMID- 10530278 TI - Microleakage of a consolidated silver direct filling material. AB - Microleakage of an experimental direct filling material comprised of a chemically precipitated silver powder that had been surface treated with a dilute acid to promote cold welding upon consolidation was evaluated. Microleakage was compared to both dispersed-phase and spherical amalgam by use of an in vitro gas-diffusion method and in class 5 restorations placed in extracted human teeth. The effect of two cavity varnishes and two dentin adhesives as cavity liners on microleakage was also evaluated using extracted teeth. Microleakage of silver powder consolidated with dental instruments was less than that found with dental amalgam. The use of copal or polyamide cavity varnish resulted in the lowest combination of microleakage on dentin and enamel margins. PMID- 10530279 TI - Self-etching primer vs phosphoric acid: an alternative concept for composite-to enamel bonding. AB - The purpose of this in vitro study was (1) to investigate the composite-to-enamel bond strength and (2) to analyze the marginal adaptation of resin composite restorations in class 2 cavities using three self-etching priming agents in comparison to conventional phosphoric acid etching and bonding application. In the first part of the study 24 extracted bovine incisors were embedded in acrylic resin and ground flat with 800-grit paper. The following three self-etching priming agents/composite resins were applied to the enamel surfaces of six teeth each: Clearfil Liner Bond 2/Clearfil AP-X (Group I), Etch & Prime 3.0/Degufill mineral (Group II), Resulcin AquaPrime + MonoBond/Ecusit (Group III). In Group IV Ecusit-Mono/Ecusit was used after enamel etching with phosphoric acid (37%). Shear bond strength values measured on a T22 K testing machine at a crosshead speed of 1 mm/min were: 24.2 +/- 3.0 MPa (Group I), 21.9 +/- 1.4 MPa (II), 34.0 +/- 3.6 MPa (III), and 26.3 +/- 1.8 MPa (IV). ANOVA revealed significant (P < 0.05) differences in shear bond strength between groups, except comparison of Group I and II, and Group I and IV. In the second part of the study 24 standardized class 2 cavity preparations with the approximal box extending 1 mm above the CEJ were prepared in extracted human molars. Enamel margins were beveled and the teeth were divided into four groups of six teeth each. Cavities were restored using the self-etching priming agents Clearfil Liner Bond 2 (Group I), Etch & Prime 3.0 (Group II), and Resulcin AquaPrime + MonoBond (Group III). In Group IV composite resin restorations were placed after 37% phosphoric acid etching and bonding application (Ecusit-Mono). Quantitative SEM analysis of the marginal adaptation of the restorations after thermocycling (5-55 degrees C, 2500 cycles) and mechanical loading (100 N, 500,000 cycles) revealed excellent, gap free margins in 91.2% (Group I), 93.0% (Group II), 92.0% (Group III), and 92.5% (Group IV) of the restorations' approximal area. There were no statistically significant differences between the four groups (P < 0.05). In conclusion, results of the present in vitro study indicate that use of self-etching primers may be an alternative to conventional phosphoric acid pre-treatment in composite to-enamel bonding restorative techniques. PMID- 10530281 TI - Epidemiology for the practicing orthodontist. AB - Epidemiological principles and biostatistical techniques are essential when performing research in human populations. This article describes the methods used for designing valid and reliable research and the statistical concepts necessary to analyze data from such studies. Specifically, the overview includes the different epidemiological study designs, sampling techniques, sources of bias encountered in research studies, and statistical tests to assess significance. It is hoped that clinical orthodontists will benefit from this review. PMID- 10530280 TI - Nanoleakage at the dentin adhesive interface vs microtensile bond strength. AB - Excessive etching of the dentin may decrease bond strength because the adhesive may fail to completely infiltrate to the base of the over-etched demineralized collagen network. The purpose of the present study was to evaluate the influence of increasing etching times on the microtensile bond strength of Single Bond and the leakage of silver ions within the hybrid layer. After etching occlusal dentin for 15, 30, or 60 seconds with 35% phosphoric acid gel, Single Bond was applied and cured for 10 seconds. Z100 was added and cured for 60 seconds. After 24 hours of water immersion, the teeth were sectioned into slices 0.7 mm thick, and hour glass-shaped specimens were prepared. Alternate slices were either dried for 30 minutes in air, kept wet, or they were coated with fingernail varnish except for 0.5 mm around the bonded area. Only the varnished samples were then stained with 50% AgNO3. Microtensile bond strength was tested using a Vitrodyne V-1000 universal tester. The samples of the stained group were embedded in self-curing PMMA and polished. All samples were observed with an SEM. Nanoleakage of silver ions was measured by exposure to laser ablation with an inductively connected plasma mass spectrometer and by electron dispersive elemental analysis. Increasing etching times seemed to have a negligible effect on bond strength of Single Bond, producing an average value of ca 38 MPa. However, the silver uptake increased upon prolonged etching times. Short-term results suggest that overetching has no detrimental effect on bond strength values of Single Bond. However, increased silver uptake, depending on the etching time, raises concern about the long-term stability of the bond. PMID- 10530282 TI - Developing outcome measures in orthodontics that reflect patient and provider values. AB - During the past decade, emphasis in orthodontics has been directed toward the development of outcome measures from both the patient and clinician perspectives. New methodological standards of rigor have been introduced into research design to eliminate bias and test well-defined questions. Sample size calculations and established exclusion and inclusion criteria define sample populations and the ability to statistically accept or reject hypothesis-driven clinical studies. Although advances in our understanding of evidence-based medicine and dentistry from the provider perspective have been productive, the emerging value placed on patient perspective has not been as forthcoming. The emphasis placed on patient oriented clinical research has resulted in new constructs of surveys and questionnaires in which the items are derived and tested from the patient's point of view. Because orthodontics is a condition without the natural history of a disease process for which no intervention has predictable consequences, new strategies have been developed to estimate need and demand for orthodontic treatment. Studies to measure seekers and nonseekers of orthodontic treatment are reported, as well as sex and cross-cultural issues in the use of established process and outcome measures. The design of clinical studies is discussed in the context of future directions for clinical research, and the usefulness of the information generated will directly relate to providing patients with the necessary information to make decisions and hence knowledgeably give informed consent for treatment interventions. PMID- 10530283 TI - Some comments on clinical studies in orthodontics and their applications to orthodontic treatment. AB - This article indicates the origins and background of the current series of National Institute of Dental and Craniofacial Research-funded, university-based clinical studies of orthodontic treatment. It suggests that future studies should be less focused on refining our estimates of mean changes during treatment and concentrate research on the systematic analysis of individual differences among patients' responses to treatment, and study how skilled clinicians make in-course corrections in response to unexpected changes in treatment conditions. Finally, some suggestions are made concerning optimization of decision making in the presence of uncertainty. PMID- 10530284 TI - Evidence of correction of open bite malocclusion using active vertical corrector treatment. AB - This study used a cephalometric analysis that isolated tipping and bodily tooth movements of the upper and the lower incisors and measured vertical skeletal changes in the anterior region of the maxilla and mandible to evaluate the outcome of two-phase nonextraction treatment for open bite malocclusion. Twenty nine subjects treated with an active vertical corrector (AVC) followed by fixed 022 orthodontic appliances were selected by one orthodontist from his private practice. All subjects lacked vertical incisor contact at the start of treatment and had adequate initial, end of phase 1, and deband lateral cephalograms. Each subject in the treated group was matched by age and sex with an untreated subject from the Broadbent Bolton Collection, Cleveland, OH. Data were collected for three time intervals; phase 1 treatment with the AVC (T1 to T2), phase 2 fixed appliance treatment (T2 to T3), and over the total treatment period (T1 to T3). Analysis of the data showed no statistically significant (P < or = .002) changes between treated subjects and controls for any variables during the phase 1 (T1 to T2) or phase 2 (T2 to T3) treatment intervals. However, overbite was significantly improved compared with controls (P < or = .002) during the T1 to T3 time interval. It was concluded that two-phase treatment with the AVC followed by fixed orthodontic appliance treatment results in a statistically significant increase in incisor overbite. The observed increase in overbite was the result of small but clinically significant changes in relative mandibular vertical growth, bodily incisor movement toward the occlusal plane, and lingual tipping of the lower incisors. PMID- 10530285 TI - Expansion with vestibular shields: an experimental test of the periosteal-pull hypothesis. AB - Two mechanisms are said to be responsible for the expansion commonly produced by buccal shields: (1) unbalanced tongue pressure and (2) periosteal pull; that is, periosteal traction on the bone overlying the molar roots. The present study used 44 young albino rats to examine these two alternatives. Half the rats wore buccal shields to produce molar expansion, and half had their maxillary molar crowns ground down to the gingiva to eliminate the effect of the tongue. In the resulting 2 x 2 design, four experimental groups were formed: (1) SM, shields and intact molars; (2) Sm, shields and reduced molars; (3) sM, no shields and intact molars; and (4) sm, no shields and reduced molars. It was hypothesized that if vestibular shields produce expansion through periosteal traction, the presence or absence of molar crowns should make no difference. Conversely, if the expansion is caused by unbalanced tongue pressure, shields should have an effect only in conjunction with intact maxillary molars. Palatal amalgam implants and dorsoventral cephalograms were used to measure the maxillary basal and dental expansion that occurred during the 6 weeks of the experiment. Analysis of variance showed the presence of highly significant interaction between shields and molars: the shields produced an increase in posterior dental expansion, but only when molar crowns were present. In contrast, basal expansion was unaffected by any combination of treatments. At least for the rat, it may be concluded that unbalanced tongue pressure, rather than periosteal traction, is probably responsible for the expansion produced by buccal shields. PMID- 10530286 TI - Root resorption caused by orthodontic treatment: an evidence-based review of literature. AB - Literature on apical root shortening published in the 1990s is reviewed. Three categories were assigned: sample-based clinical studies, clinical case reports, and animal model studies. Great variability for root shortening was reported, including resorption experienced by individuals who had never undergone orthodontic treatment. Some studies attempted to predict severe resorption whereas others compared different types of treatment. Included are several histological studies of teeth that were moved in humans. Very few of the sample based clinical studies were prospective, randomized clinical trials. Data from a study that the author was involved in was used to estimate the percentage of patients who would experience different amounts of apical root shortening. It was estimated that 5% of the patients treated would experience more than 5 mm of root shortening. PMID- 10530287 TI - Osler: of reputation and the man. PMID- 10530288 TI - Ottawa signals new interest in population health. PMID- 10530289 TI - Winnipeg becoming leader in MRI-based research. PMID- 10530290 TI - Blood-donor ban for UK visitors stems from Krever report. PMID- 10530291 TI - Screening for type 2 diabetes. PMID- 10530292 TI - Difficult decisions for long-term tube-feeding. PMID- 10530293 TI - The long-term view on refractive surgery. PMID- 10530294 TI - Doubts about the college. PMID- 10530295 TI - The walnut manoeuvre. PMID- 10530296 TI - Slapping and spanking in childhood and its association with lifetime prevalence of psychiatric disorders in a general population sample. AB - BACKGROUND: Little information is available in Canada about the prevalence of and outcomes associated with a history of slapping and spanking in childhood. The objectives of this study were to estimate the prevalence of a history of slapping or spanking in a general population sample and to assess the relation between such a history and the lifetime prevalence of psychiatric disorders. METHODS: In this general population survey, a probability sample of 9953 residents of Ontario aged 15 years and older who participated in the Ontario Health Supplement was used to examine the prevalence of a history of slapping and spanking. A subgroup of this sample (n = 4888), which comprised people aged 15 to 64 years who did not report a history of physical or sexual abuse during childhood, was used to assess the relation between a history of slapping or spanking and the lifetime prevalence of 4 categories of psychiatric disorder. The measures included a self administered questionnaire with a question about frequency of slapping and spanking during childhood, as well as an interviewer-administered questionnaire to measure psychiatric disorder. RESULTS: The majority of respondents indicated that they had been slapped or spanked, or both, by an adult during childhood "sometimes" (33.4%) or "rarely" (40.9%); 5.5% reported that this occurred "often." The remainder (20.2%) reported "never" experiencing these behaviours. Among the respondents without a history of physical or sexual abuse during childhood, those who reported being slapped or spanked "often" or "sometimes" had significantly higher lifetime rates of anxiety disorders (adjusted odds ratio [OR] 1.43, 95% confidence interval [CI] 1.04-1.96), alcohol abuse or dependence (adjusted OR 2.02, 95% CI 1.27-3.21) and one or more externalizing problems (adjusted OR 2.08, 95% CI 1.36-3.16), compared with those who reported "never" being slapped or spanked. There was also an association between a history of slapping or spanking and major depression, but it was not statistically significant (adjusted OR 1.64, 95% CI 0.96-2.80). INTERPRETATION: There appears to be a linear association between the frequency of slapping and spanking during childhood and a lifetime prevalence of anxiety disorder, alcohol abuse or dependence and externalizing problems. PMID- 10530297 TI - Fairness in the coronary angiography queue. AB - BACKGROUND: Since waiting lists for coronary angiography are generally managed without explicit queuing criteria, patients may not receive priority on the basis of clinical acuity. The objective of this study was to examine clinical and nonclinical determinants of the length of time patients wait for coronary angiography. METHODS: In this single-centre prospective cohort study conducted in the autumn of 1997, 357 consecutive patients were followed from initial triage until a coronary angiography was performed or an adverse cardiac event occurred. The referring physicians' hospital affiliation (physicians at Sunnybrook & Women's College Health Sciences Centre, those who practice at another centre but perform angiography at Sunnybrook and those with no previous association with Sunnybrook) was used to compare processes of care. A clinical urgency rating scale was used to assign a recommended maximum waiting time (RMWT) to each patient retrospectively, but this was not used in the queuing process. RMWTs and actual waiting times for patients in the 3 referral groups were compared; the influence clinical and nonclinical variables had on the actual length of time patients waited for coronary angiography was assessed; and possible predictors of adverse events were examined. RESULTS: Of 357 patients referred to Sunnybrook, 22 (6.2%) experienced adverse events while in the queue. Among those who remained, 308 (91.9%) were in need of coronary angiography; 201 (60.0%) of those patients received one within the RMWT. The length of time to angiography was influenced by clinical characteristics similar to those specified on the urgency rating scale, leading to a moderate agreement between actual waiting times and RMWTs (kappa = 0.53). However, physician affiliation was a highly significant (p < 0.001) and independent predictor of waiting time. Whereas 45.6% of the variation in waiting time was explained by all clinical factors combined, 9.3% of the variation was explained by physician affiliation alone. INTERPRETATION: Informal queuing practices for coronary angiography do reflect clinical acuity, but they are also influenced by nonclinical factors, such as the nature of the physicians' association with the catheterization facility. PMID- 10530298 TI - Is it time to ban corporal punishment of children? PMID- 10530299 TI - Waiting for medical care: is it who you know that counts? PMID- 10530300 TI - Tuition fees for residents: one physician's perspective. AB - Although the education, expertise and guidance of Canada's academic physicians cannot be overlooked, individual universities appear to see tuition fees for residents as an easy source of much needed revenue. If tuition should "rise to market levels," perhaps residents' wages should similarly rise to reflect the amount of training received, skills required, responsibilities discharged and time expended. Unfortunately, tuition fees will be an area of contention for some time. Support of provincial resident associations and medical societies may lend both moral and, possibly, financial support to future members of the profession. PMID- 10530301 TI - Reputation unrevised: celebrating the Osler sesquicentennial. PMID- 10530302 TI - William Osler at 150. PMID- 10530303 TI - Tea with Sir William Osler. PMID- 10530304 TI - Did Osler suffer from "paranoia antitherapeuticum baltimorensis"? A comparative content analysis of The Principles and Practice of Medicine and Harrison's Principles of Internal Medicine, 11th edition. AB - One of the most important legacies of Sir William Osler was his textbook The Principles and Practice of Medicine. A common criticism of the book when it was first published was its deficiency in the area of therapeutics. In this article, the 1st edition of The Principles and Practice of Medicine is compared with the 11th edition of Harrison's Principles of Internal Medicine. The analysis focuses on the treatment recommendations for 4 conditions that were covered in both books (diabetes mellitus, ischemic heart disease, pneumonia and typhoid fever). Osler's textbook dealt with typhoid fever and pneumonia at greater length, whereas Harrison's placed more emphasis on diabetes mellitus and ischemic heart disease. Notwithstanding Osler's reputation as a therapeutic nihilist, the 2 books devoted equivalent space to treatment (in terms of proportion of total sentences for the conditions). For all conditions except ischemic heart disease, Osler concentrated on general measures and symptomatic care. Throughout Osler's textbook numerous negative comments are made about the medicinal treatment of various conditions. A more accurate statement about Osler's therapeutic approach was that he was a "medicinal nihilist." His demand for proof of efficacy before use of a medication remains relevant. PMID- 10530305 TI - Osler's unusual case--was it Churg-Strauss syndrome? PMID- 10530306 TI - Osler usque ad mare: the SS William Osler. AB - William Osler's connections with the sea included a strong family history of seafaring, his own transatlantic crossings (of which there were at least 32) and the occasional use of nautical imagery in his inspirational writings. An unusual Oslerian connection with the sea emerged after his death in the form of a World War II Liberty ship. Through the SS William Osler and its sister ships, Osler was symbolically reunited with colleagues associated with the early days of the Johns Hopkins Hospital. The William Osler circumnavigated the globe in 1943 without engaging the enemy. She was then converted into an army hospital ship and renamed the USHS Wisteria. PMID- 10530307 TI - The virtual Dr. Osler. PMID- 10530308 TI - With teen pregnancies skyrocketing, ob/gyns seek support for nonprescription "morning-after pill". PMID- 10530309 TI - Huge declines in price as competition heats up in Vancouver's booming laser surgery market. PMID- 10530310 TI - Preferential treatment for WCB patients angers some MDs. PMID- 10530311 TI - Is fee-for-service on the way out for Ontario FPs? PMID- 10530312 TI - Risking it all to bring sight to Nepal. PMID- 10530313 TI - Number of roles for female MDs mushrooming as century draws to close. PMID- 10530314 TI - Texas pledges an end to "lawsuit abuse". PMID- 10530315 TI - Artist brings a touch of humanity to the ICU. PMID- 10530316 TI - Serious hematologic reactions associated with ticlopidine--update. PMID- 10530318 TI - Six new cases of canine STSS in Florida PMID- 10530317 TI - Leukotriene receptor antagonists: suspected adverse reactions. PMID- 10530319 TI - Extralabel drug use in aquaculture. PMID- 10530320 TI - Is there a place for unionization in the veterinary profession? PMID- 10530321 TI - What is your diagnosis? Femoral and tibial subchondral bone cysts in a horse. PMID- 10530322 TI - Comparison of ketoprofen and carprofen administered prior to orthopedic surgery for control of postoperative pain in dogs. AB - OBJECTIVE: To compare analgesic and adverse effects of ketoprofen and carprofen when used to control pain associated with elective orthopedic surgeries in dogs. DESIGN: Prospective randomized clinical trial. ANIMALS: 93 client-owned dogs: 46 undergoing reconstruction of the cranial cruciate ligament, 47 undergoing femoral head and neck excision, and 15 control dogs anesthetized for radiographic procedures. PROCEDURE: Dogs undergoing surgery were randomly given ketoprofen, carprofen, or saline (0.9% NaCl) solution, SC, prior to surgery. Pain score and serum cortisol concentration were recorded for 12 hours after surgery for all dogs. When pain score was > or = 7, oxymorphone was administered i.m. Bleeding time was measured prior to and during surgery. RESULTS: The proportion of dogs that required oxymorphone was significantly higher for the carprofen and placebo groups than for the ketoprofen group. Pain score for the placebo group was significantly higher than for the ketoprofen and carprofen groups, 2, 8, and 9 hours after surgery. Cortisol concentration was significantly higher for the placebo group than for the carprofen group at 4 and 6 hours after surgery. Significant differences were not detected between ketoprofen and carprofen groups with respect to pain score and cortisol concentration. Bleeding time was significantly longer for the ketoprofen group than for the other groups during surgery. One dog treated with ketoprofen developed a hematoma at the surgical site. CONCLUSIONS AND CLINICAL RELEVANCE: Ketoprofen and carprofen given prior to surgery were effective for postoperative pain relief in dogs. However, ketoprofen should not be used when noncompressible bleeding may be a problem. PMID- 10530323 TI - Effect of oral melatonin administration on sex hormone, prolactin, and thyroid hormone concentrations in adult dogs. AB - OBJECTIVE: To determine the effect of oral melatonin (MT) administration on serum concentrations of sex hormones, prolactin, and thyroxine in dogs. DESIGN: Prospective study. ANIMALS: 8 male and 8 female adult sexually intact dogs. PROCEDURE: 5 male and 5 female dogs were treated with MT (1.0 to 1.3 mg/kg [0.45 to 0.59 mg/lb] of body weight), PO, every 12 hours for 28 days; the other 6 dogs were used as controls. Blood samples were collected on days 0, 14, and 28, and serum concentrations of estradiol-17 beta, progesterone, testosterone, androstenedione, 17-hydroxyprogesterone (17-HP), dihydroepiandrostenedione sulfate (DHEAS), prolactin, and thyroxine were determined. On day 5, serum MT concentrations were measured before and periodically for up to 8 hours after MT administration in 4 treated dogs. RESULTS: Female dogs treated with MT had significant decreases in serum estradiol, testosterone, and DHEAS concentrations between days 0 and 28. Male dogs treated with MT had significant decreases in serum estradiol and 17-HP concentrations between days 0 and 28. Serum MT concentrations increased significantly after MT administration and remained high for at least 8 hours. Prolactin and thyroxine concentrations were unaffected by treatment. CONCLUSIONS AND CLINICAL RELEVANCE: Melatonin is well absorbed following oral administration and may alter serum sex hormone concentrations. PMID- 10530325 TI - Leukopenia in six healthy Belgian Tervuren. AB - A healthy 6.5-year-old sexually intact female Belgian Tervuren was found to be persistently leukopenic during preoperative evaluation for routine ovariohysterectomy. Abnormalities of the erythroid or myeloid series were not detected during bone marrow analysis. Blood samples for CBC were collected from 8 additional healthy Belgian Tervuren of both sexes and of various ages. Six of the 9 dogs were leukopenic, with WBC counts between 2.38 and 5.42 x 10(3) WBC/microl (mean +/- SD, 4.13 +/- 1.04 x 10(3) WBC/microl). Leukopenia was a persistent finding in the 3 dogs from which multiple blood samples were collected. All dogs were otherwise clinically normal. Leukopenia, as defined by a WBC count < 6.00 x 10(3) WBC/microl, may be a common finding in the Belgian Tervuren breed. PMID- 10530324 TI - Effects of intramuscular administration of low doses of medetomidine and medetomidine-butorphanol in middle-aged and old dogs. AB - OBJECTIVE: To determine effects of low doses of medetomidine administered with and without butorphanol and glycopyrrolate to middle-aged and old dogs. DESIGN: Prospective randomized clinical trial. ANIMALS: 88 healthy dogs > or = 5 years old. PROCEDURE: Dogs were assigned randomly to receive medetomidine (2, 5, or 10 micrograms/kg [0.9, 2.3, or 4.6 micrograms/lb] of body weight, i.m.) alone or with glycopyrrolate (0.01 mg/kg [0.005 mg/lb], s.c.), medetomidine (10 micrograms/kg) and butorphanol (0.2 mg/kg [0.1 mg/lb], i.m.), or medetomidine (10 micrograms/kg), butorphanol (0.2 mg/kg), and glycopyrrolate (0.01 mg/kg). Anesthesia was induced with thiopental sodium and maintained with isoflurane. Degree of sedation and analgesia were determined before and after medetomidine administration. Respiratory rate, heart rate, and mean arterial blood pressure were determined 10 and 30 minutes after medetomidine administration. Adverse effects and amounts of thiopental and isoflurane used were recorded. RESULTS: Sedation increased after medetomidine administration in 79 of 88 dogs, but decreased in 7 dogs that received 2 or 5 micrograms of medetomidine/kg. Mean postsedation analgesia score and amounts of thiopental and isoflurane used were less in dogs that received medetomidine and butorphanol, compared with other groups. Respiratory rate, heart rate, and blood pressure were not different among groups. Significantly more adverse effects developed in dogs that did not receive glycopyrrolate. CONCLUSIONS AND CLINICAL RELEVANCE: Administration of medetomidine (10 micrograms/kg, i.m.) and butorphanol (0.2 mg/kg, i.m.) induced sedation and analgesia and reduced amounts of thiopental and isoflurane required for anesthesia in middle-aged and old dogs. Glycopyrrolate decreased frequency of medetomidine-associated adverse effects. PMID- 10530326 TI - Acute toxoplasmosis following renal transplantation in three cats and a dog. AB - Three cats and 1 dog that had undergone renal transplantation because of end stage renal disease were examined because of complications 3 to 6 weeks after surgery. One cat died prior to treatment of the complications; Toxoplasma cysts were found in sections of the renal allograft, and Toxoplasma tachyzoites were found in other organs. The other 2 cats and the dog died despite treatment, and protozoal cysts, as well as tachyzoites, were identified in other organs but not within the allografts, suggesting that reactivation of latent infection following immunosuppression was the most likely cause of disseminated toxoplasmosis. These cases illustrate that toxoplasmosis can be a fatal complication in renal transplant recipients. We currently recommend that feline and canine donors and recipients undergo serologic testing for toxoplasmosis prior to surgery. In addition, we suggest that seropositive donors not be used for seronegative recipients and that seropositive recipients and that seropositive recipients be monitored closely after surgery for clinical signs of toxoplasmosis. PMID- 10530327 TI - Preparturient hypocalcemia in four cats. AB - Preparturient hypocalcemia was identified in 4 cats in a specific pathogen-free colony between 1995 and 1996. All cats had an acute onset of clinical signs, 3 to 17 days prior to parturition. Signs of depression, weakness, tachypnea, and mild muscle tremors were the most common clinical signs, following by vomiting and anorexia. Additional abnormalities included hypothermia, third eyelid prolapse, dehydration, pallor, lethargy, flaccid paralysis, and hyperexcitability. Hematologic abnormalities included leukocytosis with neutrophilia and lymphopenia. Hypocalcemia was documented in each queen. Common serum biochemical abnormalities included high aspartate aminotransferase and creatine kinase activities. All cats responded to IV or SC administration of 10% calcium gluconate. Queens were then given calcium orally prior to and following parturition. The queens did not have additional complications for the duration of the gestational or lactational periods. PMID- 10530328 TI - Prevalence of gastric ulcers in show horses. AB - OBJECTIVE: To determine prevalence and risk factors for gastric ulcers in show horses. DESIGN: Field survey. ANIMALS: 50 horses in active training that had been transported to at least 1 event in the 30 days prior to endoscopy. PROCEDURE: Interview of owner, physical examination, serum biochemical analysis, CBC, and gastric endoscopy were performed. RESULTS: Gastric ulceration was detected in 58% of the horses. Horses with a nervous disposition were more likely to have ulceration than quiet or behaviorally normal horses. Horses with gastric ulceration had significantly lower RBC counts and hemoglobin concentrations than those without ulceration. CONCLUSIONS AND CLINICAL RELEVANCE: Show horses have a high prevalence of gastric ulceration. Lower RBC counts and hemoglobin concentrations may be the result of chronic gastric ulceration. PMID- 10530329 TI - Presumptive Sry-negative XX sex reversal in a llama with multiple congenital anomalies. AB - Multiple congenital abnormalities of the external genitalia consistent with XX sex reversal were detected in a juvenile llama. The llama had a typical female karyotype (74, XX) and did not have a Y chromosome, but a minute chromosome was detected. To determine whether a piece of Y chromosome containing the Sry gene might be located in a small translocation, DNA analysis by polymerase chain reaction was performed; the Sry gene was not detected. Histologic examination revealed ovarian tissue, whereas testicular tissue was not found. External genitalia were partially masculinized, indicating that the urogenital sinus, genital tubercle, and genital swellings had been exposed to androgens during development, although the dam had not received exogenous androgens. Testicular tissue in the ovaries may have been undetected or had regressed prior to birth, as has been reported in sex reversal in mice. PMID- 10530330 TI - XX/XY chimerism and freemartinism in a female llama co-twin to a male. AB - A 20-month-old female llama was examined because at the time of mating, the male llama was apparently unable to achieve intromission. The female llama had been born co-twin to a male. On physical examination, the vaginal vestibule appeared to end in a blind pouch, and the uterus, cervix, and ovaries could not be identified during transrectal palpation or ultrasonography. Karyotyping was done, and 43% of blood lymphocytes had 2 X chromosomes, and 57% had 1 X and 1 Y chromosome. All skin fibroblasts had 2 X chromosomes. A diagnosis of freemartinism and XX/XY chimerism was made. Because conception of twins may be more common in llamas than birth of twins, it is possible that freemartinism could develop in singleton females, if, for instance, a male twin was conceived and died after the placentas had anastomosed. More widespread use of karyotyping in llamas with congenital defects of the reproductive tract will help to define the incidence of freemartinism. PMID- 10530331 TI - Ophthalmic examination and conjunctival bacteriologic culture results from a herd of North American bison. AB - OBJECTIVE: To establish ocular characteristics, determine nature and prevalence of ocular lesions, and identify representative bacterial flora from the conjunctiva of North American bison (Bison bison). DESIGN: Prospective study. ANIMALS: 63 bison; 45 males and 18 females. PROCEDURE: Ophthalmic examinations were performed on 1 group of 38 bison in December 1997 and on a second group of 25 in March 1998. Eyes were examined with a penlight, magnification loop, and indirect ophthalmoscope. Two culture swabs were used to obtain samples from the inferior conjunctival sac. One swab was submitted for isolation of bacteria and the second was submitted for isolation of Mycoplasma organisms. RESULTS: 15 ocular abnormalities were observed in 13 of the 63 bison. These included minor ocular discharge in 5 animals, 1 eyelid laceration, 1 periocular Demodex spp infection, 6 corneal abnormalities, 1 anterior synechia, and 1 cataract. Seventeen species of bacteria were isolated from the 63 swabs submitted for culture. The most prevalent bacteria were of the genus Bacillus (74.6%). Mycoplasma organisms were not observed. CONCLUSIONS AND CLINICAL RELEVANCE: Corneal abnormalities were the most frequently identified ocular lesions in bison. Bacterial flora of the conjunctiva and ocular characteristics were similar to those reported for cattle. PMID- 10530332 TI - Karyotypic and histopathologic findings in an accessory breast. AB - Accessory breasts are a clearly hereditary anomaly. They enlarge during pregnancy and lactation as a consequence of high blood levels of estrogen and prolactin, and are subject to all the diseases that occur in normal breasts. Cytogenetic analysis was performed on one accessory breast. The monossomy of chromosome 16 was the main alteration found in this material. Nonscheduled cell proliferations may produce chromosome alterations, most of them with no clinical meaning. When relevant genes are altered, major proliferations or progression to malignancy may occur. PMID- 10530333 TI - Genotoxic effects of mercury on in vitro cultures of human cells. AB - Mercury is a highly deleterious environmental pollutant, with recognized mutagenic and teratogenic effects. Given this, we evaluated the changes induced in vitro by two mercury compounds (mercury chloride--MC--and methyl mercuric chloride--MMC) on the genetic) material of a human lymphoblastoid cell line (TK6) on the basis of both the frequency of mutations at the hprt locus, and the number of chromosomic anomalies. The frequencies of HPRT- mutants in the TK6 cell line following exposure to the mercury compounds are inconclusive with regard to a mutagenic effect. However, both mercury compounds exhibit a clear cytotoxic effect, which increases with dosage. There was also no statistically significant increase in the frequency of chromosomic bleakage or gaps, nor in the number of cells with chromosomic alterations in the lymphoblastoid line. Nevertheless, MMC did provoke a marked reduction in the frequency of mitosis, both on its own and in combination with MC. PMID- 10530334 TI - Evaluating the microbial diversity of soil samples: methodological innovations. AB - This manuscript is a review of the innovative methodologies that enable more precise evaluations of soil microbial diversity. Highlighting the molecular approach, which does not require the isolation of microorganisms and allows the inclusion of non-culturable genotypes in the analyses, the described methodologies revolutionised the environmental microbiology and opened gateways for an accurate understanding of the ecology and diversity of microorganisms. The application of techniques based on soil total DNA extraction, PCR amplification of genes or gene fragments, and sequence analysis revealed that the microbial universe is far more complex than ever imagined. Examples of applications of the molecular approach to study the diversity of soil diazotrophic bacteria are given. PMID- 10530335 TI - Adaptation of the differential display RT-PCR technique to isolate sugarcane genes induced by plant association with endophytic nitrogen-fixing bacteria. AB - Differential Display RT-PCR is a powerful technique that has been used to isolate differentially expressed genes. This technique was first described by Liang & Pardee, in 1992. Afterwards, several modifications were introduced in the original version, including a simplification described by Sokolov & Prockop, in 1994. In this work, we describe an adaptation of the Sokolov & Prockop technique, in order to isolate sugarcane genes induced by plant association with endophytic nitrogen-fixing bacteria. Several modifications were introduced: the use of oligo dT primer in the first strand cDNA synthesis, replicates of the PCR reactions, analysis of the amplified fragments on silver stained polyacrilamide gel and confirmation of cloning the differentially amplified selected band prior to its molecular characterisation. The methodology established was successfully used to identify a large number of differentially amplified sugarcane cDNAs. So far, one of these cDNAs was already isolated and characterized. PMID- 10530336 TI - Identification, sequencing and structural analysis of a nifA-like gene of Acetobacter diazotrophicus. AB - A recombinant plasmid, pAD101, containing a DNA fragment of Acetobacter diazotrophicus strain PAL5 was isolated by its ability to restore Nif+ phenotype to a nifA- ntrC- double mutant of Azotobacter vinelandii. Hybridization with the nifA genes of Azospirillum brasilense located the nifA gene more precisely to specific fragments of pAD101. DNA sequencing of appropriate subclones of pAD101 revealed that the nifA gene was adjacent to the nifB gene in A. diazotrophicus, and the 5' end of the nifB gene was located downstream of the nitrogenase MoFe subunit gene, nifK. The deduced aminoacid sequence of A. diazotrophicus nifA and nifB gene were most similar to the NifA and NifB proteins of Azorhizobium caulinodans and Rhodobacter capsulatus, respectively. In addition, nucleotide sequences upstream of the A. diazotrophicus nifA-encoding region indicate features similar to those in the A. caulinodans nifA promoter region involved in O2 and fixed N regulation of nifA expression. PMID- 10530337 TI - Genetic variability of Colombian populations of Trypanosoma cruzi and Trypanosoma rangeli. AB - This paper presents our study of genetic variability of Trypanosoma cruzi and Trypanosoma rangeli strains isolated from different Colombian biological hosts, using multilocus enzyme electrophoresis for 15 enzyme systems and electrophoretic analysis of kinetoplast DNA (kDNA) digested with EcoRI and MspI endonucleases. Isoenzyme profiles were used to determine genotypes for each of the strains. Populations of T. cruzi and T. rangeli were found to have a polymorphic average of 86.7% and 26.7%, respectively. Schyzodeme analysis showed high variability for T. cruzi, since its genetic distance values were found to be greater than 50%, considerably higher than those previously reported for several T. cruzi strains from other countries. These results suggest that Colombian strains should be considered as genetically independent entities and are worth studying independently from each other to clearly establish their biological and clinical characteristics. PMID- 10530338 TI - Pressure oscillations in anesthetized dogs and its conversion into quasi-periodic orbits. AB - Using a basic representation of dynamic systems, arterial blood pressure pulsation is converted into quasi-periodic orbits with the purpose of transforming a periodic phenomena into a cyclical one by plotting the pressure p(t) versus its first derivative dp/dt. This elementary mathematical procedure made it possible to evaluate the variability of the systemic arterial pressure pulsations, both systolic and diastolic, as well as the slope variability of the anachrotic and catachrotic phases. Two periods, which can be used to estimate different sources of variability, can be distinguished in the catachrotic phase. One corresponds to the open aortic valves, and the other is associated with the closed valves. Furthermore, through the first derivative of pressure oscillations we were able to identify small changes in arterial pressure, which appeared when the sampling rate was at least 150 samples per second. Since the time variable was converted into a parameter, the result was a synoptic or holistic approach, which is a considerable improvement for the visual analysis of cardiovascular phenomena. This simplified mathematical procedure can be easily implemented on a personal computer in real time and applied to all rhythmic phenomena in Physiology and Pathology. PMID- 10530339 TI - Effect of cholinergic agonists on muscular tonus of the lizard small intestine and esophagus. AB - The underlying mechanisms of acetylcholine-induced intestinal relaxation in the lizard Liolaemus tenuis tenuis are still unknown. By using a classical model of intestinal recording of isometric contraction and relaxation in conjunction with specific pharmacological tools, this article studies the possible influence of EDRF/NO and nicotinic ganglionar receptors on the Ach-induced relaxation in an effort to elucidate the probable mechanisms involved in ACh effect. It was observed that the relaxation of the lizard intestine elicited by ACh (10(-7) - 4 x 10(-4) M) was not affected by hexametonium (5 x 10(-4) M) or tetrodotoxin (10( 6) M). Nicotine (10(-7) to 10(-4) M) induced relaxation was significantly antagonized by hexametonium; however, it was not influenced by tetrodotoxin. These results allow us to discard a neuronal pathway in cholinergic-induced relaxation, suggesting a more direct cholinergic effect on the smooth muscle, perhaps mediated by an unknown substance released by some specialized tissue. N nitro-L-arginine, used to block NO-synthase and NO production, induced no changes in ACh-induced relaxation. Methylene blue, a soluble guanylate cyclase inhibitor, induced no changes in ACh-induced relaxation. These results allow us to discard a probable role of EDRF/nitric oxide in the ACh-induced relaxation of lizard small intestine, providing evidence that this mechanism could be different from that reported in other species. PMID- 10530340 TI - Signal transduction networks and the biology of plant cells. AB - The development of plant transformation in the mid-1980s and of many new tools for cell biology, molecular genetics, and biochemistry has resulted in enormous progress in plant biology in the past decade. With the completion of the genome sequence of Arabidopsis thaliana just around the corner, we can expect even faster progress in the next decade. The interface between cell biology and signal transduction is emerging as a new and important field of research. In the past we thought of cell biology strictly in terms of organelles and their biogenesis and function, and researchers focused on questions such as, how do proteins enter chloroplasts? or, what is the structure of the macromolecules of the cell wall and how are these molecules secreted? Signal transduction dealt primarily with the perception of light (photomorphogenesis) or hormones and with the effect such signals have on enhancing the activity of specific genes. Now we see that the fields of cell biology and signal transduction are merging because signals pass between organelles and a single signal transduction pathway usually involves multiple organelles or cellular structures. Here are some examples to illustrate this new paradigm. How does abscisic acid (ABA) regulate stomatal closure? This pathway involves not only ABA receptors whose location is not yet known, but cation and anion channels in the plasma membrane, changes in the cytoskeleton, movement of water through water channels in the tonoplast and the plasma membrane, proteins with a farnesyl tail that can be located either in the cytosol or attached to a membrane, and probably unidentified ion channels in the tonoplast. In addition there are highly localized calcium oscillations in the cytoplasm resulting from the release of calcium stored in various compartments. The activities of all these cellular structures need to be coordinated during ABA induced stomatal closure. For another example of the interplay between the proteins of signal transduction pathways and cytoplasmic structures, consider how plants mount defense responses against pathogens. Elicitors produced by pathogens bind to receptors on the plant plasma membrane or in the cytosol and eventually activate a large number of genes. This results in the coordination of activities at the plasma membrane (production of reactive oxygen species), in the cytoskeleton, localized calcium oscillations, and the modulation of protein kinases and protein phosphatases whose locations remain to be determined. The movement of transcription factors into the nucleus to activate the defense genes requires their release from cytosolic anchors and passage through the nuclear pore complexes of the nuclear envelope. This review does not cover all the recent progress in plant signal transduction and cell biology; it is confined to the topics that were discussed at a recent (November 1998) workshop held in Santiago at which lecturers from Chile, the USA and the UK presented recent results from their laboratories. PMID- 10530341 TI - [Dermato-venereal diseases in Africa: a real public health problem]. PMID- 10530342 TI - [Dermatology and extracorporeal photochemotherapy 10 years later]. PMID- 10530343 TI - [Muir-Torre syndrome]. PMID- 10530344 TI - [Muir-Torre syndrome and familial colorectal cancer: 2 families with molecular genetic analysis]. AB - INTRODUCTION: The Muir-Torre Syndrome is a rare genodermatosis, defined by the occurrence of cutaneous tumors (such as sebaceous adenomas, epitheliomas, or carcinomas, and/or keratoacanthomas), and internal malignancies. Recently, molecular analysis in hereditary non polyposis colorectal cancer demonstrated a common genetic basis, linking these two disorders, with the observation of germline mutations in the hMSH2 gene (one of the DNA mismatch repair system genes) in both syndromes. Such molecular demonstration of a single nosological entity should be clinically used to improve the indications of molecular testings in oncogenetics, still restricted to highly stringent criteria for hereditary non polyposis colorectal cancer. CLINICAL CASES: We identified three patients from two different families, who fulfilled the criteria for both Muir-Torre Syndrome and hereditary non polyposis colorectal cancer. The search of a germline mutation of the hMSH2 gene was performed on an affected member from each family, and their relatives with their informed consent. Within each family, all individuals with colorectal cancer were carriers of the same mutation. In the first family, this mutation was a pathogenic microinsertion, usable for predictive testing. In the second family, a missense mutation was identified, requesting further demonstration of its pathogenicity before its use in a predictive purpose. CONCLUSIONS: The diagnosis of cutaneous tumors compatible with Muir-Torre syndrome should lead the dermatologist to suspect an hereditary non polyposis colorectal cancer that should bring to an oncogenetic approach: personnal and familial history of colorectal cancer, molecular analysis, recommendations for colonoscopic screening in at-risk relatives. In the case of a colorectal cancer at a young age, or in the case of familial cases, the gastroenterologist should screen for cutaneous lesions of Muir-Torre syndrome, which could add a criteria for an hereditary syndrome, and lead to molecular oncogenetic analysis. PMID- 10530345 TI - [Oral ulcers induced by nicorandil: prevalence and clinicopathological aspects]. AB - INTRODUCTION: The first observations of "giant buccal aphthosis" induced by nicorandil were published in 1996. Nicorandil is a potassium channel activator used in the treatment of angina pectoris, which seems to induce specific buccal ulcerations. The purpose of this study was to analyze the clinicopathologic data of patients with aphthosis induced by nicorandil and to study the prevalence of this side effect. PATIENTS AND METHODS: We have seen 3 patients who spontaneously consulted, and 5 patients who were addressed to us after a telephone survey. We have then examined 100 consecutive patients treated by nicorandil for at least 1 month, who were hospitalized in 3 departments of cardiology in Strasbourg, and 100 age- and sex-matched controls who were treated by other antianginal drugs. RESULTS: Our 8 patients suffered from large, chronic and painful ulcerations of a 4-week duration, located on the tongue, the gingiva and the cheeks despite various symptomatic treatments. In one case, histopathologic data were consistent with an eosinophilic ulcer. Prospective study: among 100 patients treated by nicorandil, 5 had unusual chronic buccal ulcerations, whereas none of the 100 controls had aphthosis (p = 0.03). The confidence interval (99 p. 100) of this side effect prevalence was therefore 1 p. 100 to 14 p. 100. DISCUSSION: Nicorandil can induce large and painful buccal ulcerations with severe dysphagia, weight loss, and depression. Dermatologists should be aware of this particular side-effect, since our study showed a high prevalence, and because lesions heal rapidly after withdrawal of nicorandil. Why nicorandil may be associated with mouth ulcers remains unanswered. A past history of aphthae could be a cofactor of this side-effect. PMID- 10530346 TI - [Erysipelas in the young population of a military hospital]. AB - INTRODUCTION: Erysipelas usually affects either elderly patients or patients with predisposing factors. We studied erysipelas in a young healthy military population. SUBJECTS AND METHODS: A retrospective study of patients, less than 30 years old, admitted to the Hopital d'Instruction des Armees Robert Picque between 1991 and 1997 was performed. RESULTS: Eighty-one patients were studied, 80 were men, mean age was 21.4 years. Localization of erysipelas was: face: 2; leg: 74. Facilitating factors were: portal of entry in 100 p. 100; venous insufficiency in 1 case; alcohol abuse in 1 case. Anti-inflammatory agents had been given at the beginning of treatment in 32.1 p. 100 cases. 7.1 p. 100 patients had a recurrence.. Complications were: abscess in 8 cases and bursitis in 1 case. No facilitating factor was detected. The course was not related with bacterial findings. No thrombo-embolic complication was observed. DISCUSSION: In a young healthy population, erysipelas is not a rare infection. Face is an exceptional localization of erysipelas in relation to older population. The main risk factor for developing this infection is local factor found in 100 p. 100 cases but general factors as repeated hikes, care access, long standing could also be incriminated. Use of anti-inflammatory agents is frequent but role of anti inflammatory agents in developing complications is not clear. Complications are rare and benign except for frequence of recurrences while the good health. PMID- 10530347 TI - [Mutation in the MSH2 gene in Muir-Torre syndrome]. AB - BACKGROUND: The Muir-Torre syndrome is an autosomal dominant hereditary condition predisposing to cancer. It is characterized by skin tumors associated with adenocarcinoma of the colon or other neoplasias observed in the context of hereditary non-polyposis colorectal cancer (HNPCC). The Muir-Torre syndrome is also characterized by the frequent presence of multiple colonic polyps and the relatively moderate aggressivity of the tumors. CASE REPORT: We studied a family with Muir-Torre syndrome. We sequenced the exons of the hMSH2 gene in this family and identified heterozygous germinal mutation by G insert at position 2427 (2427insG). This mutation changes the lecture phase producing a premature codon stop. DISCUSSION: Our study confirms the predominant responsibility of the hMSH2 gene in Tuir-Torre syndrome. This clinical case and data reported in the literature demonstrate the importance of searching for a history of non-polyposis colorectal cancer in patients and relatives and the unstable genome characteristic of these tumors found in sebaceous tumors or keratoacanthomas. Sequencing the hMSH2 gene should be a priority when clinical features, history and laboratory tests suggest Muir-Torre syndrome. PMID- 10530348 TI - [Lichen pemphigoid associated with developing hepatitis B in a child]. AB - INTRODUCTION: Lichen planus pemphigoides is a rare acquired auto-immune bullous dermatosis which usually affects adults. Only four cases have been reported in children. We describe a new case of lichen planus pemphigoides in a child unusual by its association with an evolutive hepatitis B and by the occurrence of a lichen planus relapse. CASE REPORT: A 10-year-old African boy has been seen for a pruritic dermatosis with papular lichenoid lesions on the trunk and the limbs and blisters on the lower limbs, both arise on lichen planus lesions and normal skin. The diagnosis of lichen planus pemphigoides was confirmed by histology which showed the features of lichen planus on a papule and of a sub-epidermal split on a bulla and by direct and indirect immunofluorescent studies which revealed an IgG and C3 linear deposit at the dermo-epidermal junction and the presence of circulating IgG anti-basement membrane zone antibodies. Laboratory investigations showed an evolutive hepatitis B (HBsAg +). Healing was obtained by dapsone and topical steroid therapy. Eight months after withdrawal of treatment the patient presented a non-bullous relapse of lichen planus. The histology showed a typical aspect of lichen planus and the immunofluorescence studies were negative. The hepatitis B serology was unchanged. The lesions rapidly improved with topical steroid and coaltar. One year later the patient exhibited few slight lichen planus lesions on the limbs and the hepatitis B serology showed the onset of sero conversion. DISCUSSION: Lichen planus and lichen planus pemphigoides are probably variants of the same disease. Their successive occurrence in our case report favours this hypothesis as does the association with an hepatitis B. PMID- 10530349 TI - [Cutaneous granulomatous lesions disclosing ataxia-telangiectasia]. AB - INTRODUCTION: Idiopathic cutaneous granulomatous lesions are exceptionally described in the course of congenital immunodeficiency, including ataxia telangiectasia. CASE REPORT: We describe a new case of a 28-month girl who presented granulomatous skin lesions revealing a previously unknown ataxia telangiectasia in the absence of typical neurologic signs, telangiectasia and infectious complications. The clinical aspect showed infiltrated erythemato squamous plaques and nodules predominating on the face and limbs. These lesions increased in number without remission. Histological examination revealed a nodular, lymphohistiocytic infiltration with granulomatous tendency in the deep dermis and the hypodermis. Before the onset of skin treatment, the child developed an Epstein-Barr-virus related lymphoproliferation. Immunoglobulins and oral corticosteroids associated with chemotherapy permitted the regression of the granulomatous lesions but not of the fatal spread of the lymphoproliferative syndrome. DISCUSSION: These rare cutaneous manifestations are important to know because they can be the initial sign of an immunodeficiency. Clinical and histological aspects are characteristic. They are eventually associated with visceral granulomatous lesions. Physiopathology remains hypothetical. An abnormal immune response to an undetermined antigenic stimulation could be suspected in this particular context. The question of a correlation between these lesions and a proliferative syndrome remains open. PMID- 10530350 TI - [Resistant acquired bullous epidermolysis with severe ocular involvement: the success of extracorporeal photochemotherapy]. AB - BACKGROUND: Epidermolysis bullosa aquisista can leave several functional sequelae. The lesions sometimes resist treatment. CASE REPORT: We report a case of a 25-year-old young man presenting with a severe epidermolysis bullosa acquisita confirmed by the electronic immunomicroscopy. He had a major ocular involvement with symblepharon and cicatricial synechial lesions. He was almost blind because of corneal scars. All immunosuppressive treatments had failed: systemic corticoids, cyclosporin, azathioprine. The introduction of extracorporeal photochemotherapy resulted in the healing of the lesions, after a total of 32 procedures. All treatment are now stopped, and the lesions are purely cicatricial, without any relapse of the disease since 9 months. Corneal grafts are now under process, to try to recover a part of the lost visual acuity. DISCUSSION: This case demonstrates the efficacy of extracorporeal chemotherapy to be tried in case other treatments failed. PMID- 10530351 TI - [Pigmented pemphigoid]. AB - BACKGROUND: Pigmented bullous pemphigoid has many clinical manifestations. We report a pigmented erythematous form with disseminated bullae. CASE REPORT: A 70 year-old woman with a history of breast adenocarcinoma developed an eruption of pigmented macules on the trunk and members of 72 hour duration. At five days, there was an eruption of tight bullae. The diagnosis of bullous pemphigoid was retained because of the association of infraepidermic bullae, linear deposits of C3 along the basal membrane and the presence of the 180 kDa minor bullous pemphigoid antigen on immunoblotting. DISCUSSION: The initial pigmented aspect of this bullous pemphigoid suggested disseminated pigmented bullous erythema, but histology data with immunotransfer corrected the diagnosis. This very atypical presentation led us to look for a particular etiology. There was no argument in favor of a malignancy in this patient in complete remission after treatment of her breast adenocarcinoma. The fact that the patient was phototype IV would probably explain the pigmented nature of the initial lesions. PMID- 10530352 TI - [Psychological impact of neurofibromatosis type 1: the analysis of interviews with 12 patients with regard to an evaluation of the quality of life]. PMID- 10530353 TI - [Diagnostic case. Cutaneous B-cell pseudolymphoma of the nose]. PMID- 10530354 TI - [Diagnostic case. Sclerous cutaneous fibroma]. PMID- 10530355 TI - [Syphilis (except congenital syphilis)]. PMID- 10530356 TI - [Pruritus without skin lesions]. PMID- 10530357 TI - [Expectant management of pruritus in pregnant women]. PMID- 10530358 TI - [Cicatricial pemphigoid]. PMID- 10530359 TI - [Langerhans cells generated in vitro: advances and application]. PMID- 10530360 TI - [Multiple herpes simplex type 1 whitlow lesions]. PMID- 10530362 TI - [Internet Grateful Med V2.6.2]. PMID- 10530361 TI - [Overdose of nortriptyline during treatment with terbinafine (1st reported case)]. PMID- 10530363 TI - [What are the indications for non-steroidal anti-inflammatory agents in dermatology?]. PMID- 10530364 TI - [The injection of insulin through clothing: a safe practice?]. PMID- 10530365 TI - Harnessing energy to overcome conflict. PMID- 10530366 TI - President's message. PMID- 10530367 TI - International education--a kaleidoscopic view. AB - International travel presents opportunities for perioperative nurses to share their knowledge about surgical patient care and preventing the spread of infection. This article outlines the author's personal experiences with international travel, educational pursuits, and conferring with nurses, physicians, epidemiologists, hospital administrators, and medical product company representatives. It also discusses culture and values, teaching and learning, and accomplishments in settings where both financial and human resources are sparse, but outcomes are positive and meaningful. PMID- 10530368 TI - Perioperative care coordinator nurse competency statements. Nursing Practice Committee. Association of periOperative Registered Nurses. PMID- 10530369 TI - A collaborative approach to isolated limb perfusion. AB - Isolated limb perfusion is used to treat unresectable sarcoma, melanoma, and other select tumors. It is performed in the OR and requires collaboration by surgeons, perioperative nurses, anesthesia care providers, pharmacists, perfusion technologists, and nuclear medicine personnel. The procedure involves complete isolation of the vascular supply to a limb before an infusion of high dose chemotherapeutic medications. PMID- 10530370 TI - Harvesting and implanting allograft bone. AB - Allograft bone (i.e., bone that is obtained from a donor) implantation is used for bone replacement and for reconstructing serious bone defects during total joint revision surgery. It is also used for bone replacement after large excisions of bone tumors. An extensive variety of laboratory tests must be completed, appropriate storage facilities for allograft bone have to be available, and protocols must be developed before a facility can set up a bone bank. Many ethical issues and nursing responsibilities need to be considered before the decision is made to use allograft bone. PMID- 10530372 TI - Orthopedic surgery policy; reuse of tray wrappers; rapid-read biological indicators; needle safety PMID- 10530371 TI - Sterile supply storage--finding a place for everything. PMID- 10530373 TI - Internet resources about allografts in total joint replacement. PMID- 10530374 TI - Making grassroots action effective. PMID- 10530375 TI - When an adverse sentinel event is the cause for action. PMID- 10530376 TI - [Cytogenetic effect in lymphocytes in astronauts after 2 lengthy flights on board MIR orbital station]. AB - The cytogenetic analysis of peripheric blood of cosmonauts who participated in the 196- and 194-day Mir missions evidenced increases in the number of aberrant cells and the rate of chromosomal aberrations. However, in most cases these were not statistically significant as compared to control values. Nonetheless, the lesions in cell chromosomal apparatus can amplify the risk of delayed consequences of exposure to ionizing radiation. PMID- 10530377 TI - [Tuberculin-dependent and nonspecific migration activity of leukocytes in astronauts]. AB - There have been presented the results of studying the reactivity of leucocytes with respect to tuberculin and monspecific migration activity of leukocytes in the cosmonatus pre flight and after completion of orbital expeditions of various duration are presented. In some cosmonauts tuberculin rearranged migration of leukocytes. This may bear witness to activisation of the immunity system in response to the antigens which had been already familiar to test-subjects. Experimental data pointed to significant changes in leukocyte mobility in 12 out of 28 cosmonauts examined on days 1 and 7-14 of recovery after the first or repeated long-duration space missions indicating some functional disorders in the phagocytic link which may lead to a typical decline in human resistance to infections. PMID- 10530378 TI - [The relationship between astronauts erroneous actions, their psychophysiological state and work and rest regimen]. AB - The correlation analysis of the data obtained in the MIR 18, and 21 through to 23 main missions was supportive of the strong dependence of erroneous actions by cosmonauts, their psychophysiologic state and the work/rest schedule. This conclusion guidelined the development of a prototype of technique to perform an integrated analysis of erroneous crew actions which can also help to identification of impacts of psychophysiologic state and deviations from the requisite rest/work schedule in order to set the origin of erroneous actions, and to work out recommendations towards the enhancement of ground-based crew training and other matters. With these findings, the predictive virtues of the technique has been demonstrated. PMID- 10530379 TI - [Radiation risk for astronauts during space flights on board the space station "MIR"]. AB - Described is the algorithm for calculating the radiation risk to cosmonauts on orbital missions during different solar phases. The algorithm and the Fortran calculation program lie in the basis of presented radiation risk estimations for orbital manned missions of varying duration. The dependence of in-flight radiation risk on mission length, solar phase, and cosmonaut's age was analyzed. Magnitudes of radiation risk to cosmonauts were compared with the national demographic risk of male lethality over a similar period of time. PMID- 10530380 TI - [Mechanisms of early changes in water-electrolyte metabolism in man in various ground-based models of the effects of microgravity]. AB - Analysed was an array of data amassed in 4 series of experiments with simulated microgravity effects (HDT (-6 degrees); dry and suited immersions in horizontal and vertical position). Water-electrolyte turnover in blood and urine and hormonal controls have been under study. It was demonstrated that irrespective of the body axis position relative to the gravity vector in the immersion models, reactions of the active regulation of the water--electrolyte metabolism were noted already within the initial hours, i.e. more rapidly than in HDT. During the early phase of adaptation to the immersion models, some ostensibly similar responses were governed by different hormonal controls. Unlike the dry immersion, the suited immersion tests with various body positioning modified not only the strength but the dynamics of the water-electrolyte regulation, too. Findings also included differences in the dynamics osmo- and volumoregulation in the vertical and horizontal suited immersion tests which are, possibly, reflective of recruitment of different stimulating mechanisms and efficiency of organs-targets. PMID- 10530381 TI - [Results of studies of carbohydrate metabolism and ultrasonography of the pancreas in man after continuous anti-orthostatic hypokinesia]. AB - Data on the hypokinesia-induced transformation of the glycemic profile and ultrasonic changes in the pancreas structure are presented. The AOH study gave further evidence of transforming glycemic curves. Moreover, increased sizes of tail and head of the pancreas and a decrease in its echogeneity were observed in all test-subjects. Structural changes in the pancreas were confirmed by biochemical investigations which revealed increased levels of blood enzymes and activation of insulin secretion. Increases in the liver size, thickness of the wall of the stomach, diameter of the splenic vein were indicative of progressing venous plethora in the portal vein system. It was shown that venous plethora are the main cause for changes in the upper GI, and in the pancreas state in particular, which can be qualified as dysfunctional. These structural changes in the pancreas could suppress its functional activity manifested by increases in blood enzymes and hormones and transformation of the glycemic profile during the glucose load. PMID- 10530383 TI - [Experimental study of pilot-assisted detection of changes in the information field of the on-board view indicator]. AB - The paper reports experimental data about the effects of operative memory restrictions on the indices of reliability of the visual identification of changes in external objects represented on the onboard visual indicator display. It was demonstrated that parallel piloting and visual tracking reduced the probability of identification of changes in two external objects up to 0.89 and deteriorated the ability to sustain the designated mode of piloting in six and more times. Two types of erroneous responses were delineated: overlooking changes in symbol-targets and false anxieties. Changes in the background symbols appeared to provoke false anxieties. The highest levels of reliability of eye tracking could be reached by changing color and/or contours of symbols-targets on the display. PMID- 10530382 TI - [The syndrome of reduced colonization of periodontium tissues during long-term anti-orthostatic hypokinesia]. AB - Effects of antiorthostatic hypokinesia (AOH, -6 degrees) during 60 and 120 days on the parodontium in 10 human subjects aged 24-33 yr. have been studied. Clinical, functional, immunologic, and microbiological analyses were performed 30 days prior to, on days 7 and 30, and on day 7 since the experiment. The parodontium was assessed by relevant indices, immunoglobulines in the oral fluid, and bacteriological parameters determined with the anaerobic cultivation technique. AOH was found to reduce the colonisation resistance of the parodontium due to obviously, the immunologic inertia and consequent changes in the qualitative composition of microflora. It is concluded that the parodontium responds to extreme conditions by replacement of the commensal microflora by opportunistic and obligate parodontosis pathogens. PMID- 10530384 TI - [Human resistance to decompression sickness and nonspecific methods of its elevation]. AB - The object of this study was the dependence of decompression sickness (DCS) tolerance determined by the intensity of venous gas embolism on functioning of the cardiovascular and respiratory systems, and micro-circulation which predetermine wash-out of an indifferent gas. Efficiency of nonspecific methods, e.g. hypercapnic training, hyperbaric oxygenation, exposures to pulse current to enhance human tolerance to DCS was experimentally substantiated. PMID- 10530385 TI - [The aspects of adenylate cyclase activity regulation in myocardium cell membranes during hypokinesia]. AB - Nonstimulated and isoproterenol, GTF, GITF, NaF stimulated activities of the adenylate cyclase in sarcolemma in white rats' myocardium was studied after two weeks of hypokinesia. As was established, in restrained animals the sensitivity of adenylate cyclase to the specified agents was increased and transition to the bimodal GTF regulation took place. It is hypothesised that involvement of membrane-bound Gi-proteins in the adrenergic effects on cardiomyocytes is one of mechanisms of the cardiotropic effects of restraint and heart distresses. PMID- 10530386 TI - [The effects of space flights on the lipid composition of blood, adrenal glands and liver in rats]. AB - Numerous postflight biochemical and morphological investigations of rats evidenced some shifts in lipid metabolism due to weightlessness and/or gravitation stresses. In the "Spacelab-2" experiment with rats, lipid spectra of blood serum (plasma), liver and adrenal glands were explored with the thin layer silicagel chromatography with the aim of evaluating stress effects of space flight on lipid metabolism. For the first time tissues were gathered and analyzed on day 13 in microgravity. Lipid composition of the liver remained unchanged after 13 days of flight. Those were cholesterol ethers of blood serum that were significantly increased while no changes had occurred in the relative quantities of each class of lipids in their totality. The adrenal glands considerably reduced concentrations of total lipids, free cholesterols, triglycerides, and free fatty acids. Within four hours into landing, free fatty acids were found to increase in blood plasma whereas the relative content of triglycerides decreased. Hepatic lipids remained unchanged and the adrenal ones did not differ from the control. On day 14 of recovery, the total content of blood plasma lipids was significantly increased on the score of triglycerides, and free and esterified cholesterols. The liver displayed decreases in all classes of fats; however, similar changes were in the control rats, too. The lipid composition of the adrenal did not differ from that of control. According to the total balance of lipids in rats' blood serum and liver, no signs of acute or chronic stress had been developed by the concluding stage of the 140-day space mission. Yet, one cannot exclude an acute stress early in mission that might have been the cause for the shifts in lipid metabolism in adrenals resulting in their reversible delipoidization. PMID- 10530387 TI - [The use of pharmacological agents for the increase of +Gz tolerance in macaca mulatta]. AB - Tolerance of rhesus monkeys to increased +Gz loads was studied in relation to the conditions of the blood circulation functioning. In four anaesthetised monkeys, the external carotid arteries were ligated and a biomedical cuff was implanted on the common carotid arteries to measure blood pressure, linear blood velocity, and to register EKG. Blood flow in the carotid artery nearing the zero served as the criterion for termination of rotation with a gradient of 0.01 units/s Each animal was centrifuged 4 times at a week interval: first when intact and then after injection of m-cholinolytic metacine, adrenomimetic metazone and or reopolyglucin which increases the plasma volume. These studies demonstrated that the reflex effect on the heart (elevated CO and increased arterial pressure as a result of moderate tachicardia conditioned by metacine) or vessels (activation of alfa receptors by metazone) as well as an increase in the volume of intravascular fluids, are almost equally favourable to the maintenance of the central blood volume, and the +Gz-tolerance, i.e. about 35% (delta = 1.62-1.77 units). PMID- 10530388 TI - [The role of intravertebral disk protrusion in pathogenesis of lumbosacral radiculopathies in flying personnel]. AB - The significance of x-ray computer tomography in determining the mechanisms of pathogenesis of lumbar-sacral radiculopathy in patients with protrusions of intervertebral disks of the lumbar-sacral backbone is demonstrated. Pilots with the protrusions (n = 135) and the intervertebral disk herinas (79) have been examined. The clinical course of the sickness was reviewed in both groups of patients. The computer-tomographic semiotics of various protrusions was elaborated and expanded and their clinical implications demonstrated. The fresh paradigm of pathogenesis of radiculopathy at various forms of protrusions is presented. The roentgenomorphologic classification of arthrosis of archiprossesed joints was studied and proposed. PMID- 10530389 TI - [Contemporary approaches to the treatment of the irritable bowel syndrome in pilots]. AB - Reported are results of treating 53 pilots with the irritated large intestine syndrome verified by endoscopic and morphologic investigations. Salozinal proved to be highly helpful to young patients. One of the therapeutic mechanisms is blocking of the cycloxygenase path circumstantially confirmed by normalization of lipid peroxidation and AOS owing to salozinal. PMID- 10530390 TI - [Radiation diagnosis of spleen changes in flying personnel of fighter and helicopter aviation]. AB - Some specific alterations of the spleen in flying personnel of fighter and helicopter aircraft have been elucidated. Regular patterns of splenomegaly appear to be linked with the adverse effects of air flight. The ultrasonic investigation was proved to be one of the most significant elements of the diagnostic algorithm used to determine the size and structure of the glandular parenchyma. The X-ray computer tomography and methods of computational analysis are no more than ancillary tools to get better insight into syntopy of the organ and surrounding tissues. The radionuclide investigation provides tentative data concerning the functional state of the organ. PMID- 10530391 TI - [The model of the rate of radiation-induced mammalian death based on the determination of delayed consequences of different doses of radiation]. AB - Analyzed were model descriptions of the probability of mammalian lethality and a possible life span reduction consequent to acute and chronic exposures to different dose rates. The proposed model of radiation rate of mammalian death links variations of the coefficient of mammalian death due to acute and prolonged radiation exposures with age. Application of the model to relevant experimental data yielded model coefficients for these periods of exposure. The radiation modified dependence of lethality coefficients on age makes it possible to obtain fairly simple analytical expressions describing the survival probability long after exposure as a function of dose and dose rate, estimating radiation risk at any time point following exposure, and shortening of mean life span associated with exposure to different doses. PMID- 10530392 TI - [A method of preservation of urine for the determination of creatinine]. PMID- 10530393 TI - A systematic review of the role of human papillomavirus testing within a cervical screening programme. PMID- 10530394 TI - Making the best use of human embryos. PMID- 10530395 TI - Strategies for embryo utilization in assisted reproduction--how do we assess their relative effectiveness? PMID- 10530396 TI - What is the best strategy for presenting results in assisted reproductive technology? PMID- 10530397 TI - What is the best strategy for presenting ART results? A controversial comment. PMID- 10530398 TI - Are the Australian ART results as poor as they appear? PMID- 10530399 TI - What is the best strategy for using embryos and presenting results in assisted reproductive technology? A strategy for valid comparisons of results. PMID- 10530400 TI - How to present ART results: do we need a gold standard? PMID- 10530402 TI - Relationship of the human cumulus-free oocyte maturational profile with in vitro outcome parameters after intracytoplasmic sperm injection. AB - PURPOSE: We investigated whether the human oocyte maturational profile at the removal of cumulus/corona cells affects the fertilization rate and subsequent embryo quality after intracytoplasmic sperm injection. METHODS: A total of 1011 oocytes from 150 cycles was included in this retrospective analysis. Cumulus-free oocytes that were in prophase or metaphase I of meiosis at the removal of cumulus/corona cells were incubated in vitro until they reached metaphase II (in vitro-matured oocytes) and were then immediately injected with a single spermatozoa. Oocytes that were in metaphase II at the removal of cumulus/corona cells (MII oocytes) received sperm injection after 3-4 hr of preinjection incubation. RESULTS: The fertilization rate of the MII oocytes was significantly higher than that of in vitro-matured oocytes (81 vs 62%; P < 0.001). The cleavage rates were similar in the two groups (MII oocytes, 94%; in vitro-matured oocytes, 91%). However, MII oocytes had significantly higher percentages of good-quality embryos (grade 1-3 embryos, 87 vs 58%, P < 0.001) and embryos with high cumulative embryo scores (score 10-32 embryos, 62 vs 33%, P < 0.001). The mean cumulative embryo score of MII oocytes after fertilization was also higher than that of in vitro-matured oocytes (12.1 +/- 3.8 vs 8.8 +/- 3.4; P = 0.014). CONCLUSIONS: MII oocytes that extruded the first polar body at the removal of cumulus/corona cells had better fertilization rates and embryo morphology than in vitro-matured oocytes that extruded the first polar body following the removal of cumulus/corona cells and in vitro culture. PMID- 10530401 TI - Relationship of total motile sperm count and percentage motile sperm to successful pregnancy rates following intrauterine insemination. AB - PURPOSE: This study sought (i) to investigate the relationship between postwash total motile sperm count and postwash percentage motile sperm in predicting successful intrauterine insemination and (ii) to determine the minimal postwash total motile sperm count required to achieve pregnancy with intrauterine insemination. METHODS: Five hundred four women, who underwent 1636 intrauterine insemination cycles with their partner's sperm for infertility treatment from 1993 through 1995, were included in this retrospective study. All patient charts were reviewed for age, infertility etiology, ovarian stimulation regimens, semen characteristics, and treatment outcome. To determine the relationship between total motile sperm count and intrauterine insemination outcome, patients were grouped as (1) less than 0.5 million, (2) 0.5 to 1 million, (3) 1 to 5 million, (4) greater than 5 million, and (5) greater than 20 million. RESULTS: Similar live birth rates (per cycle) were seen among the postwash total motile sperm count groups: group 1, 3.5%; group 2, 2.4%; group 3, 7.0%; group 4, 6.9%; and group 5, 7.0% (P = 0.37). However, regardless of the postwash total motile sperm count, the postwash motility predicted intrauterine insemination success at a cutoff value of 40%. CONCLUSIONS: The percentage of postwash sperm motility, and not the postwash total motile sperm count, can predict successful intrauterine insemination outcome. Such information can be useful in counseling patients regarding their chance of success with intrauterine insemination and in determining when alternate methods of assisted reproduction may be a better approach. PMID- 10530403 TI - Subpopulations of human granulosa-luteal cells obtained from gonadotropin- or gonadotropin-releasing hormone agonist/gonadotropin-treated follicles in in vitro fertilization-embryo transfer cycles. AB - PURPOSE: Our purpose was to find the differences in granulosa-luteal cells obtained from gonadotropin-versus gonadotropin-releasing hormone (GnRH) agonist/gonadotropin-treated follicles in in vitro fertilization-embryo transfer (IVF-ET) cycles. METHODS: Granulosa-luteal cells were obtained from 45 follicles of women undergoing IVF-ET with gonadotropin releasing hormone (GnRH) agonist and human menopausal gonadotropin (hMG) and from 45 follicles of women with hMG IVF ET cycles. Subpopulations of granulosa-luteal cells were observed by computerized image analysis in which human chorionic gonadotropin (hCG) was localized using immunoperoxidase staining. RESULTS: The luteinized granulosa-luteal cells from hMG-treated follicles were larger than those from GnRH agonist/hMG-treated follicles. The hMG-treated follicles contained more hCG-stained cells, particularly those with cytoplasmic hCG localization. CONCLUSIONS: We found differences in morphometric characteristics and hCG localization in granulosa luteal cells obtained from hMG-versus GnRH agonist/hMG-treated follicles. We presume that the results indicate the influence and importance of luteal-phase support on the clinical pregnancy rate in GnRH agonist/hMG-treated IVF-ET cycles. PMID- 10530404 TI - The ratio of X- and Y-bearing sperm in ejaculates of men with three or more children of the same sex. AB - PURPOSE: The present study evaluated the proportions of X-bearing and Y-bearing sperm within the semen of donors who were the declared fathers of three or more sons or daughters. METHODS: The proportions of sperm were determined using dual color fluorescence in situ hybridization to identify the X and Y chromosomes. RESULTS: The only difference observed was in semen volume. There was no increase in the proportion of Y-bearing sperm for men with only sons (49.7 +/- 1.3%) or of X-bearing sperm for men with only daughters (44.8 +/- 2.6%). CONCLUSIONS: A preponderance of either sons or daughters in a family cannot be explained simply by an altered ratio of X-bearing and Y-bearing sperm in the father's semen. PMID- 10530405 TI - The optimum time for exogenous human chorionic gonadotropin to rescue the corpus luteum. AB - PURPOSE: Our purpose was to study the optimum time to administer exogenous human chorionic gonadotropin (hCG) to rescue the human corpus luteum during the luteal phase of normal menstrual cycles. METHODS: Groups of normally cycling women were given 4-day regimes of exogenous hCG by daily injection beginning 4 (Group A), 8 (Group B), and 12 (Group C) days after the midcycle luteinizing hormone surge. The hCG regime used was designed to mimic hCG levels following a spontaneous implantation. All subjects acted as their own controls in a preceding normal menstrual cycle. RESULTS: Group A subjects exhibited patterns and levels of salivary progesterone concentration similar to those seen in the control cycles throughout the normal luteal phase. In contrast, subjects in both Group B and Group C demonstrated a rapid and sustained increase in progesterone production following the hCG injections. Furthermore, subjects in Group B achieved the highest mean peak progesterone concentrations and the total amount of salivary progesterone secreted was significantly higher than in the control cycles (P < 0.05). Although the mean luteal-phase length was greatest in Group C, the response of the corpus luteum was suboptimal, with a delayed rise in salivary progesterone. CONCLUSIONS: These data show that the qualitative and quantitative response of corpus luteum to an early pregnancy-type hCG signal is maximal around the midluteal phase, coincident with the time of implantation. PMID- 10530406 TI - Familial risk among patients with endometriosis. AB - PURPOSE: The objective of the present study was to determine the prevalence of endometriosis among the relatives of patients with confirmed endometriosis. METHODS: We analyzed the prevalence of endometriosis among first-, second-, and third-degree relatives in a group of 101 patients with varying symptoms related to endometriosis seen at two public hospitals and submitted to laparoscopy and/or laparotomy. The control group consisted of 43 women submitted to laparoscopy without a diagnosis of endometriosis. RESULTS: Among the patients with endometriosis, we detected nine families with a positive history of endometriosis, comprising one mother, six sisters, three aunts, and two cousins, as opposed to no case among the controls. CONCLUSIONS: These data confirm a familial tendency for endometriosis and suggest that this disorder has a genetic basis. PMID- 10530407 TI - The effect of colchicine treatment on spermatozoa: a cytogenetic approach. PMID- 10530408 TI - Clonal rearrangements in childhood and adult precursor B acute lymphoblastic leukemia: a comparative polymerase chain reaction study using multiple sets of primers. AB - Ig heavy chain (IgH) and T-cell receptor (TCR) gene rearrangements were investigated by polymerase chain reaction (PCR) amplification of diagnostic tumour samples from 91 patients (57 children and 34 adults, with cut-off at age 16) with precursor B acute lymphoblastic leukemia (ALL). Using primers directed to the framework regions (FR) 1, 2 and 3 of the IgH gene, clonal IgH rearrangements were observed in 82, 58 and 58%, respectively, whereas clonality was presented in 45 and 27% using primers hybridising to the TCR delta and gamma genes. A combination of all five primer sets used resulted in 96% positive cases (children 100%, adults 88%). The frequency of clonal IgH rearrangements correlated to patient age with a significantly lower fraction of positive cases in the adult group. The concomitant usage of more than one V(H) family gene was similar for childhood and adult ALL, and an over-representation of V(H)6 rearrangements was found in childhood ALL. Twenty-five out of 91 cases (27%) displayed an oligoclonal pattern for either IgH or TCR gene rearrangements (children 37%, adults 12%). A comparative analysis of samples from different compartments was performed in 23 patients, and differences between two or three compartments were observed in seven cases. Unexpectedly large, clonally appearing PCR products of 540-715 bp were found in three leukemias and sequence analysis verified their clonal nature. In summary, using multiple sets of primers clonal rearrangements of IgH and TCR genes can be detected in a very high frequency, including previously neglected large size PCR products. A common heterogeneity was demonstrated in different compartments reflecting ongoing clonal evolution, which can make detection of minimal residual disease (MRD) in ALL troublesome. Therefore, we suggest that a minimum of three targets should be used to minimise false-negative results. PMID- 10530409 TI - Leukaemia cell drug resistance and prognostic factors in AML. AB - In 93 cases of acute myeloid leukaemia (AML) the extent to which prognostic factors mirrored the in vitro cellular chemotherapy resistance (to anthracyclines aclarubicin (Acla) and daunorubicin (Dau) as well as nucleoside analogue cytarabine (Ara-C)) was investigated using a 4-d MTT (3-[4,5-dimethylthiazol-2 yl]-2,5-diphenyl tetrazolium bromide) assay. We found that age at presentation and presence of secondary AML were significantly correlated to leukaemia cell Ara C resistance. Thus, analysis of in vitro drug resistance data revealed that age at presentation and presence of secondary leukaemia were both independently correlated to cellular drug resistance, with older age being associated with higher Ara-C resistance in vitro (p=0.02 and 0.01 in univariate and multivariate analyses, respectively) and with secondary leukaemia being associated with higher Ara-C resistance (p=0.04 and 0.059 in univariate and multivariate analysis, respectively). Median LC-50 values (Ara-C) were: 178 ng/ml in paediatric cases, 356 ng/ml in younger adult cases, and 584 ng/ml in elderly (age > or = 60 yr) cases giving a resistance ratio between these age subgroups of 1:2.0:3.3. Median LC-50 values (Ara-C) was 381 ng/ml in de novo cases as opposed to 891 ng/ml (resistance ratio 1:2.3) in secondary cases. By contrast, cytogenetic findings, presenting leucocyte count, FAB-subtype, and gender were not consistently correlated to in vitro drug resistance to any of the three drugs. We conclude that at least two major adverse prognostic factors in AML (advanced age at presentation and presence of secondary leukaemia) are associated with increased leukaemia cell Ara-C resistance. High leucocyte count is not associated with increased cellular drug resistance towards Acla, Ara-C or Dau. PMID- 10530411 TI - A longitudinal study of serum ferritin in 319 adolescent Danish boys and girls examined in 1986 and 1992. AB - This study examined trends in iron status in adolescents. Serum ferritin was measured in 1986 and 1992 in 319 Danes (161 males) stratified into 5 groups: I. median age 9 yr in 1986 vs. 15 yr in 1992; II. 11 vs. 17 yr; III. 13 vs. 19 yr; IV. 15 vs. 21 yr; V. 17 vs. 23 yr. Males in group I demonstrated no change in ferritin or estimated iron stores in mg/kg; groups II-V displayed an increase in iron status parameters. All groups showed an increase in estimated total iron stores. Changes in iron status parameters were inversely correlated with height velocity in group III, and positively correlated with height velocity in group V. Females in age groups I and II demonstrated a fall in ferritin and estimated iron stores in mg/kg in association with menarche; values were unchanged in groups III and IV, and increased in group V. All groups showed an increase in estimated total iron stores. Changes in iron status parameters were inversely correlated with height velocity in groups I and II. In conclusion, ferritin levels in adolescents display great variation during growth spurt and at menarche. Changes in ferritin showed no consistent association with growth velocity. In both genders, estimated total iron stores increased with age. PMID- 10530410 TI - Response rate and survival after conventional chemotherapy for multiple myeloma by hospitals with different inclusion rates of patients to the trials. A Finnish Leukemia Group study. AB - The purpose of this study was to analyse the influence of the hospital size, measured as the number of annual patient enrolments in the Finnish Leukaemia Group trials in the period 1979-85, on response rate and survival after conventional chemotherapy for multiple myeloma. The 432 myeloma patients for this study were derived from 4 prospective multicentre trials of the Finnish Leukaemia Group. The comparisons of the response rate to primary chemotherapy, progression free survival time (PFS), response rate after first relapse and overall survival time (OS) were made between patients treated in 3 hospital categories according to annual enrolment rate. 273 of the patients had been treated in "large" hospitals, 120 in "intermediate" and 38 in "small" ones. The median OS of patients aged up to 70 yr was 49 months and that of patients older than 70 yr was 31 months. There were no significant differences in response rate, PFS, response rate after first relapse and OS between the hospital categories. The most important endpoints in the treatment of multiple myeloma, response rate, PFS and OS are independent of the size of the hospital, measured as the number of annual patient entries in the trial. This allows the decentralisation of the conventional chemotherapy of multiple myeloma, within the framework of properly organised clinical trials. PMID- 10530412 TI - The proportion of reticulated platelets is higher in bone marrow than in peripheral blood in haematological patients. AB - Since the detection that platelets originate from megakaryocytes (MK), the site of megakaryocyte fragmentation has been disputed. Some authors have even postulated that platelets are solely produced in the lungs. Thus, we have directly measured platelet generation in the bone marrow (BM) by comparing the relative number of young RNA-containing, so-called reticulated platelets (%RP) in the BM and in the peripheral blood (PB). Two separate prospective, cross sectional trials have been conducted in patients routinely undergoing BM biopsies for diagnostic purposes. In the first part of the study 30 patients with stem cell or bone marrow transplantation were examined. The second part of the study was performed in 62 haematological patients visiting the outpatient's clinic. Median %RP were higher in BM than in PB (p <0.001). In the second part of the study the difference averaged 133% (interquartile range: 30-383%). There was a moderate correlation between %RP in BM and in PB (r = 0.67; p <0.001). The absolute number of RP in PB correlated weakly with the number of megakaryocytes (0.42; p = 0.001), which was due to a correlation between the platelet counts and the megakaryocyte counts (r = 0.55; p <0.001 in biopsies). Two patients with autoimmune antibodies against GPIIb/IIIa exhibited 10% and 16% RP in PB, and had 29% and 59% RP in BM, respectively. It is concluded that the relative number of RP is significantly higher in BM than in blood. This supports the notion that platelets are at least in part released from MK in the bone marrow, particularly in patients suffering from immune thrombocytopenia. PMID- 10530413 TI - Serum thrombopoietin levels in thrombocytopenic and non-thrombocytopenic patients with human immunodeficiency virus (HIV-1) infection. AB - HIV-1 seropositive patients often exhibit thrombocytopenia, considered of multifactorial aetiology. Thrombopoietin (TPO), a recently isolated cytokine, is the main regulator of megakaryocyte and platelet production. The objective of this study was to analyse serum TPO levels in thrombocytopenic and non thrombocytopenic HIV-1 infected patients. Serum TPO levels were measured by ELISA in 43 healthy individuals and in 88 HIV-1 infected patients: 68 thrombocytopenics and 20 non-thrombocytopenics. Thrombocytopenic HIV-1 infected patients showed higher TPO concentrations (263 +/- 342 pg/ml) than non-thrombocytopenics (191 +/- 86 pg/ml); levels in both groups were significantly higher than those of healthy controls (121 +/- 58 pg/ml). Two subgroups of thrombocytopenic patients, the autoimmune thrombocytopenic purpura (AITP) group and the mild thrombocytopenic group, presented TPO levels similar to those of non-thrombocytopenics. Patients exhibiting pancytopenia showed the highest TPO concentrations. However, there was no correlation between TPO levels and platelet counts in any group of HIV-1 infected patients. TPO levels in HIV-1 seropositive patients were slightly increased and the differences in TPO levels between thrombocytopenic and non thrombocytopenic patients were generally small. The finding of mildly increased TPO levels along with the recently described recovery of thrombocytopenia following recombinant TPO administration confirms the implication of ineffective platelet production in the origin of HIV-associated thrombocytopenia. PMID- 10530414 TI - Natural effector cells in patients with acute myeloid leukemia treated with the immunomodulator Linomide after autologous bone marrow transplantation. AB - Roquinimex, Linomide, is a quinoline derivative with pleiotropic immunomodulatory activities which has been shown to enhance NK function. As part of a phase III placebo-controlled multicenter study patients were randomized to receive Roquinimex, 0.2 mg/kg body weight, or a placebo twice weekly for a duration of 2 yr following autologous bone marrow transplantation for acute myeloid leukemia in remission. At Arhus University Hospital 7 patients were randomized to receive the active drug and 6 to receive the placebo. Surviving patients were followed for 2 yr with immunological monitoring of their natural immune effector cells (NK- and LAK cell activity). Peripheral heparinized blood samples were obtained twice before the onset of conditioning therapy and at several time points after ABMT, and whole blood samples were analyzed by flow cytometry for the detection of leukocyte differentiation antigens as well as by 4 h 51Cr release assays for cytotoxicity. In contrast to previous experience with Linomide, in the present study we found that at 36 wk or later time points Linomide patients exhibited a significant suppression of circulating natural effector cell number and activity when compared with the control group. These observations underline the need for further exploration into novel and manageable immunostimulators. PMID- 10530415 TI - Clinical immunology of chronic cold agglutinin disease. AB - We studied clinical and immunological characteristics of 15 patients with chronic cold agglutinin disease (CAD). Mean age at disease debut was 68 years for female and 67 years for male patients. The patients had no signs of other autoimmune diseases. All patients had V(H)4-34 encoded IgM kappa cold agglutinins (CA) in high titre. In five patients IgM increased significantly with advancing disease. Seven patients had reduced concentrations of lymphocytes, largely of CD4 and CD8 T cells. Percentages of NK cells (CD56) and B cells (CD19) were increased in seven and three patients, respectively. In six out of nine patients a clonal expansion of kappa positive B cells was found. Serum C3 was decreased in nine patients and C4 was decreased in 11 patients, six of whom had reduced CH50. Such data indicate that patients with CAD experience a continuous low-grade complement consumption. Five patients had experienced increased haemolysis during infections. After addition of active complement to patient sera in vitro, six sera showed increased haemolytic activity. Our results indicate that some patients with CAD have a relative deficit of complement in their serum and that an increase of complement production occurs during an acute phase reaction which enhances haemolysis. Our data also indicate that both CA titre and thermal amplitude are important characteristics when predicting complement activation and clinical course in CAD. PMID- 10530416 TI - Deferiprone (L1) associated neutropenia in beta thalassemia major: an Indian experience. PMID- 10530417 TI - Severe aplastic anemia evolving into T cell acute lymphoblastic leukemia. PMID- 10530418 TI - Chemiluminescence: a test for predicting hematopoietic recovery after autologous or allogeneic bone marrow transplantation. PMID- 10530419 TI - A patient with acute lymphoblastic leukemia who presented with osteoporosis and vertebral fractures. PMID- 10530420 TI - Parvovirus B19 infection does not contribute significantly to severe anaemia in children with malaria in Malawi. PMID- 10530421 TI - A successful combination of plasma exchange and intravenous cyclophosphamide in a patient with a refractory thrombotic thrombocytopenic purpura. PMID- 10530422 TI - BCR-negative chronic myeloid leukemia in relapse after bone marrow transplantation is susceptible to remission induction by donor lymphocyte infusions. PMID- 10530423 TI - Histiocytic, necrotizing lymphadenitis as rare cause of cervical lymphadenopathy and fever of unknown origin--a case of biopsy proven recurrence over 19 years. PMID- 10530424 TI - Prevalence of herds with young sows seropositive to pseudorabies (Aujeszky's disease) in northern Belgium. AB - In Belgium, pseudorabies in swine has been the subject of a mandatory eradication programme since 1993. From December 1995 to February 1996, a survey was conducted in the five provinces of northern Belgium to estimate the provincial pseudorabies virus (PRV) herd seroprevalence. Seven hundred and twenty randomly selected herds were included in this survey. To detect recently infected animals, only young sows were sampled. The results show that 44% of these herds had an important number of PRV-seropositive young sows. The highest herd seroprevalence was observed in West Flanders (68%), followed by Antwerp (60%), East Flanders (43%), Limburg (18%), and Flemish Brabant (8%). Assuming a diagnostic test sensitivity and specificity of 95% and 99%, respectively, and a true PRV within-herd prevalence of 43%, the overall true PRV herd prevalence was estimated to be 35%. A logistic multiple-regression revealed that the presence of finishing pigs was associated with a two-fold increase in odds of a herd being seropositive (odds ratio (OR)=2.07, 95% confidence interval (CI) = 1.31-3.26); a breeding herd size > or =70 sows was associated with a four-fold increase in odds of a herd being seropositive (OR = 4.09, 95% CI = 2.18-7.67); a pig density in the municipality of >455 pigs/km2 was associated with a 10-fold increase in odds of a herd being seropositive (OR = 9.68, 95% CI = 5.17-18.12). No association was detected between the PRV herd seroprevalence and purchase policy of breeding pigs (purchased gilts, or use of homebred gilts only). PMID- 10530425 TI - Public preferences regarding rabies-prevention policies in the UK. AB - The current 6-month quarantine system for all cats and dogs entering the UK has kept the UK rabies-free since 1922. However, pressure is mounting for a change to a system of vaccination, microchip identification and serological testing. In response to the increasing controversy surrounding the quarantine system, the UK government recently set up an independent review panel to assess the alternatives. This paper quantifies public preferences for the current policy and three alternative rabies-prevention measures. A survey was used not only to assess the overall preferences for rabies-prevention policies but also to assess the importance of policy attributes and socio-economic characteristics in determining policy preferences. We interviewed a sample of pet-owners in North Yorkshire. The results showed that the existing system was the single most preferred policy option. However, a large proportion of the sample preferred the vaccination-based policies. A logistic-regression model and ordered probit models were used to find that safety and animal welfare were the most-important factors determining policy preferences. The respondents' awareness of the rabies-policy review, a desire to take a pet abroad, the amount of foreign travel, occupation and previous experience of quarantine were all important factors in policy choice. Socio-economic characteristics such as income, pets owned and the number of children were not significant determinants of policy preference. PMID- 10530427 TI - Culling of dairy cows. Part II. Effects of diseases and reproductive performance on culling in Finnish Ayrshire cows. AB - The effects of 15 diseases and reproductive performance on culling were studied in 39727 Finnish Ayrshire cows that calved in 1993 and were followed until culling or next calving. Survival analysis, using the Cox proportional hazards model, was performed with diseases and pregnancy status as time-dependent covariates. Parity, calving season and herd were included as covariates in every model. The effect of the number of inseminations was also studied. The farmer's knowledge of the cow's pregnancy status had a significant effect on culling. It varied according to the stage of lactation a cow was in; the earlier the farmer knew a cow was pregnant, the smaller was the risk of culling. If a cow had not been inseminated at all, her risk of culling was 10 times higher than if she was inseminated once. If a cow was inseminated more than once, she had a slightly lower risk of being culled than a cow inseminated only once. The effect of parity decreased when pregnancy status and number of inseminations were added to the model, indicating that part of the parity effect was accounted for by reproductive performance. Including diseases in the model with pregnancy status and the number of inseminations did not change the effects of reproductive performance on culling. Mastitis, teat injuries and lameness had the greatest effect on culling (whether adjusted for reproductive performance or not), increasing the risk of culling, followed by anestrus, ovarian cysts and milk fever. In general, the effects of diseases decreased when reproductive performance was also accounted for in the model. When pregnancy status was included in the model, the effects of anestrus and ovarian cysts became slightly more protective, but when the number of inseminations was also considered, they became non-significant at the beginning of lactation and they increased the risk of culling at the end of lactation. Sensitivity analysis, which was run to evaluate the effects of our censoring mechanism on the results, indicated that the censoring times (i.e., the time of next calving) were not fully independent of the event (culling) times; the effects of the diseases and pregnancy status at the very end of the lactation changed slightly from the original model. PMID- 10530426 TI - Seroprevalence to bovine virus diarrhoea virus and other viruses of the bovine respiratory complex in Venezuela (Apure State). AB - Six hundred and fifteen serum samples obtained from cows in five districts of Apure State, Venezuela, were tested by ELISA for antibodies to bovine virus diarrhoea virus (BVDV). The same samples were also ELISA-tested for antibodies to bovine herpesvirus type 1 (BHV-1) and bovine respiratory syncytial virus (BRSV). Additionally, the haemagglutination-inhibition (HI) test was used for detecting antibodies to parainfluenza virus type 3 (PIV-3). Overall, seroprevalence to BVDV was 36+/-7% (SE); seroprevalence varied by district (19-42%). BHV-1 seroprevalence was 67+/-4%; variation by district was similar to that of BVDV. However, the first 80 serum samples tested by BHV-1 ELISA all had a strong background reaction with the control antigen. Therefore, these sera were adsorbed to a homogenate of non-infected bovine kidney cell line (MDBK) and retested by ELISA. The non-specific reactivity was significantly reduced (p<0.001 by Wilcoxon's signed-rank test). Compared to the virus-neutralisation (VN) test, the adsorbed BHV-1 ELISA showed 94% agreement and gave a kappa value of 0.84, indicating that the adsorption did not interfere with test accuracy. Seroprevalence against BRSV was 85+/-3%, and showed differences across districts. Most of the cows (94+/-2%) were seropositive to PIV-3, and there were no significant differences among districts. PMID- 10530428 TI - Culling of dairy cows. Part III. Effects of diseases, pregnancy status and milk yield on culling in Finnish Ayrshire cows. AB - The effects of 15 diseases, pregnancy status and milk yield on culling were studied in 39727 Finnish Ayrshire cows that calved in 1993 and were followed until culling or next calving. Survival analysis, using the Cox proportional hazards model, was performed with diseases, pregnancy status and milk yield as time-dependent covariates. Effects of parity, calving season and herd were also accounted for. Pregnancy status was the single most influential factor affecting culling decisions, followed by milk yield. Several diseases also had a significant effect on culling, the most influential ones being mastitis, lameness, teat injuries, and milk fever. The effects of all of these factors varied according to the stage of lactation. Milk yield had a significant effect on culling decisions, depending on the stage of lactation. At the beginning of lactation, milk production did not have any effect on culling decisions, but later on, the highest producers were at the lowest risk of being culled and the lowest producers had the highest risk. Adjusting for milk yield modified the effects of parity, most diseases and also pregnancy status on culling. Effects of parity increased after including milk yield in the model, indicating that milk yield and parity are interrelated in their effects on culling. The effects of pregnancy status also increased towards the end of lactation when milk yield was accounted for in the model. The effects of mastitis, teat injuries and lameness decreased after adjusting for milk production. These diseases lower milk yield and thus, part of their effect on culling was mediated through milk production. The effects of anestrus and ovarian cysts were mainly modified by pregnancy status, but not by milk yield. The effects of milk fever on culling increased at the beginning of lactation after including milk yield in the model. This suggests that even though cows with milk fever tend to be higher producers, it is the disease as such that triggers the culling decision early in the lactation. The changes in the effects of other diseases after adjusting for milk yield varied, depending on the disease and the stage of lactation. PMID- 10530429 TI - Immunopotentiating and immunotherapeutic effects of thymic hormones and factors with special emphasis on thymic humoral factor THF-gamma2. AB - The essential role played by the thymus in the development of the immune response was well documented in many publications. These findings prompted a long series of studies devised to define the factors produced and secreted by thymus cells, which are involved in the development and nature of immunological responsiveness. First experiments done with crude thymus extracts were followed by isolation of purified products and finally by chemical characterization and synthesis of immunologically active thymus-derived peptides. In this article we review the various thymic hormones and factors described, that is, thymosin fractions 5, the thymosins, prothymosin alpha, thymulin (FTS-Zn), thymopoietin, thymostimulin (TP 1), Thymic humoral factor (THF), and THF-gamma2. Studies demonstrating the activity of the various thymic factors in increasing the immunocompetence potential in both in vitro and in vivo conditions are discussed. The immunostimulatory potential of thymic factors was also investigated in experimental models where beneficial therapeutic effects were sought in a situation of immunological malfunction. The last part of the review is dedicated to clinical trials with thymic factors that revealed improvement in the immunocompetence potential in cases of immunodeficiencies, viral infections, and cancer and its correlation with therapeutic effectiveness. It seems that more research is required in order to better define conditions for the use of thymic factors in immunotherapy. PMID- 10530430 TI - Immune dysfunction as a factor in heat illness. AB - The influence of stress on immune function is well recognized. Indeed for some authors, the changes induced by combinations of vigorous exercise and heat exposure are merely examples of a more generalized stress response. However, there has been surprisingly little consideration of how far disturbances of immune function contribute to heat illness and heat fatalities. The physiology of exercise in hot environments has recently been reviewed. This article provides a brief outline of the main features of heat illness. It then summarizes current knowledge of general immune responses to stress and specific reactions to heat exposure and heavy exercise, and discusses the potential impact of such changes on the outcome of heat illness. PMID- 10530431 TI - Novel strategies using DNA for the induction of mucosal immunity. AB - The mucosal surfaces are the primary sites for transmission of most infectious diseases. However, most conventional vaccines are administered parenterally [e.g., by intramuscular (IM) or intradermal (ID) injection] and induce systemic but rarely mucosal immunity. Novel vaccination strategies capable of inducing both systemic and mucosal immune responses could greatly reduce infection and morbidity worldwide. One of the most exciting advances in vaccine technology in recent years has been the development of DNA vaccines, through which the antigen is synthesized in vivo after direct introduction of its encoding sequences. The vast majority of DNA vaccines have been delivered parenterally; however, in recent years a number of studies have reported successful mucosal immunization with DNA vaccines. The induction of strong immune responses following the introduction of DNA appears to be partly due to the potent adjuvant effect of unmethylated immunostimulatory CpG motifs present in the DNA backbone. Synthetic oligodeoxynucleotides (ODN) containing such immunostimulatory CpG motifs are potent adjuvants systemically and mucosally in mice, and have synergistic action with other adjuvants, such as alum and cholera toxin (CT). This article highlights the recent advances in vaccination strategies using DNA delivered to mucosal surfaces either as an antigen-encoding plasmid or as an adjuvant. PMID- 10530432 TI - Structure and function of the CD7 molecule. AB - CD7 is a single-domain Ig superfamily molecule expressed on human T and NK cells, as well as on cells in the early stages of T, B, and myeloid cell differentiation. CD7 is highly expressed on malignant immature T cells and is generally absent on malignant mature T cells, such as CD4+ Sezary leukemia and HTLV-1+ adult T-cell leukemia cells. Because of lack of identification of a natural ligand and lack of a monoclonal antibody against murine CD7, the in vivo functions of CD7 have until recently remained obscure. Recent studies in CD7 deficient mice have provided new insights into CD7 function, and demonstrated key roles for CD7 in regulating peripheral T and NK cell cytokine production and sensitivity to LPS-induced shock syndromes. This article reviews recent work on the expression, structure, and function of CD7, and discusses roles the CD7 molecule might play in T and NK cell development and function. PMID- 10530433 TI - Application of polymerase chain reaction to the diagnosis of infectious diseases. PMID- 10530434 TI - Changing epidemiology of infections in patients with neutropenia and cancer: emphasis on gram-positive and resistant bacteria. AB - Over the past 3 decades, considerable changes have occurred in the types of bacteria causing infection in febrile patients with neutropenia and cancer. Twenty years ago, gram-negative bacteria caused approximately 70% of bloodstream infections. As a probable consequence of long-dwelling intravascular devices, fluoroquinolone prophylaxis, and high-dose chemotherapy-induced mucositis, there has been a shift toward gram-positive coccal bacteremia. In most centers today, approximately 70% of bacteremic isolates are gram-positive cocci. Of potential concern is that antimicrobial-resistant gram-positive organisms are becoming increasingly frequent in patients with neutropenia. Fluoroquinolone-resistant Escherichia coli are being isolated from several cancer centers. Several "new" organisms, such as Stomatococcus mucilaginosus, Bacillus cereus, Leuconostoc species, Corynebacterium jeikeium, Rhodococcus species, Stenotrophomonas maltophilia, Moraxella catarrhalis, Burkholderia cepacia, and Bartonella species, now cause infections in these patients. Careful application of infection-control principles, judicious prophylaxis, appropriate evaluation of new antibiotics, and prompt effective therapy will maximize benefits for these patients. PMID- 10530436 TI - Vancomycin as part of initial empirical antibiotic therapy for febrile neutropenia in patients with cancer: pros and cons. AB - Gram-positive organisms predominate as the bacterial pathogens identified in episodes of febrile neutropenia. This has led to increased use of antibiotics with efficacy against gram-positive organisms (often vancomycin) as part of empirical antibiotic regimens for treating febrile neutropenia. Among 101 children randomized to receive amikacin, ticarcillin, and vancomycin or ticarcillin/clavulanate and amikacin along with vancomycin placebo, treatment success in those treated with vancomycin was higher (85% vs. 62%). In 1990, the European Organization for Research and Treatment of Cancer-National Cancer Institute of Canada Clinical Trials Group compared amikacin and ceftazidime with and without vancomycin and concluded that there was no need to include vancomycin in initial empirical antibiotic therapy. Results from another study and a retrospective review of a large clinical trial also support the previous conclusion. In 1999, most experts in the field recommend vancomycin not be part of the initial empirical therapy regimen for treating febrile neutropenia in patients with cancer. PMID- 10530435 TI - Contemporary antimicrobial susceptibility patterns of bacterial pathogens commonly associated with febrile patients with neutropenia. AB - One of the most challenging problems in antimicrobial chemotherapy is the effective empirical treatment of infection in patients with neutropenia. The rates of occurrence for pathogens have significantly changed (from predominance of gram-negative to gram-positive organisms) under selective pressure of broad spectrum antimicrobial therapy or prophylaxis, and novel resistance mechanisms have emerged. To address the need for appropriate monotherapy or combination regimens for patients with neutropenia, physicians must prescribe agents with a spectrum of antimicrobial activity to inhibit the major, prevalent pathogens encountered in bloodstream infection and pneumonia; in addition, these selected agents must be active against recently described resistant organisms. Data from the SENTRY Antimicrobial Surveillance Program indicate that several broad spectrum agents remain highly active and can be used alone or in combinations. In most cases, the newer compounds with increased activity and spectrum against gram positive cocci (i.e., carbapenems, cefepime, levofloxacin, and trovafloxacin) offer a greater inhibitory potential for empirical therapy among patients with neutropenia and severe infections. PMID- 10530437 TI - Is monotherapy for febrile neutropenia still a viable alternative? AB - Monotherapy for empirical treatment of febrile neutropenia is effective and often less costly than combination therapy but remains controversial. The controversy results from observations that combination therapy for Pseudomonas aeruginosa improved outcomes, and this approach became a standard. Many subsequent publications, including the Infectious Diseases Society of America guidelines for febrile neutropenia, now support monotherapy. However, changes in the pathogens involved in febrile neutropenia and in their resistance prompt a reevaluation. In the evaluation of new antibiotics, recent trials comparing either cefepime or meropenem with combination therapy or with ceftazidime confirm that monotherapy remains a viable therapeutic approach, with infectious mortality in the 5% range in all arms. The choice of monotherapy should, however, be made on the basis of resistance patterns seen in an institution. The agent selected should be very active against the organisms that are likely to cause rapidly fatal infections, and clinicians must be prepared to modify monotherapy as appropriate. PMID- 10530438 TI - New trends in patient management: risk-based therapy for febrile patients with neutropenia. AB - Standard management of febrile neutropenia includes the prompt administration of empirical, broad-spectrum, parenteral antibiotic therapy. This is generally done in a hospital-based setting. Although effective (overall survival of >90%), such therapy leads to prolonged hospitalization, excessive resource utilization, and increased costs. Recently, risk-assessment models have been developed that reliably differentiate febrile patients with neutropenia that are at low risk for morbidity and/or mortality. This has enabled clinicians to administer risk-based treatment to such patients. High-risk patients still receive standard, hospital based, parenteral treatment. Many patients, however, defervesce promptly and can be discharged home with parenteral or oral antibiotics. Low-risk patients need not be hospitalized at all and can be safely treated with parenteral or oral antibiotics in the outpatient or home setting. Careful risk assessment and patient selection, appropriate antimicrobial regimen(s), and meticulous monitoring for response or the development of complications or toxicity are essential for the success of risk-based therapy. PMID- 10530439 TI - Study on microbial persistence in end-stage idiopathic dilated cardiomyopathy. AB - Microbial persistence may be involved in the pathogenesis of idiopathic dilated cardiomyopathy (IDC). Therefore, we evaluated the role of various cardiopathogenic microorganisms in patients with end-stage IDC. In a previous study, we did not find evidence for the persistence of enterovirus RNA in end stage IDC. In the present study, we looked for other microorganisms that are frequently associated with heart disease, including cytomegalovirus, hepatitis B virus, hepatitis C virus, Borrelia burgdorferi, Chlamydia species, mycoplasmata, and Toxoplasma gondii. Serology, polymerase chain reaction (PCR) analysis specific for detection of microbial genomic sequences, or both investigations were performed on myocardial samples from 37 patients with end-stage IDC. PCR analysis was performed on multiple myocardial samples per patient. Thirty-nine patients with end-stage heart disease of known cause were included as controls. On the basis of our serological data and PCR analyses, we did not find any evidence that microbial persistence in the heart is involved in the end-stage disease process of IDC. PMID- 10530440 TI - Editorial response: microbial persistence and idiopathic dilated cardiomyopathy. PMID- 10530441 TI - Rhinovirus infections in myelosuppressed adult blood and marrow transplant recipients. AB - Scant data are available on the clinical significance of rhinovirus infections in immunocompromised patients. We reviewed the clinical courses of and outcomes for 22 myelosuppressed adult blood and marrow transplant recipients with rhinovirus infections who were hospitalized at the M.D. Anderson Cancer Center (Houston) from January 1992 to January 1997. In 15 patients (68%), illnesses remained confined to the upper respiratory tract. Seven patients (32%) developed fatal pneumonia. These patients had profound respiratory failure a mean of 12 days (range, 3-21 days) after the onset of symptoms. In six of these seven cases, rhinovirus was isolated before death from a bronchoalveolar lavage fluid specimen and/or an endotracheal aspirate. Five patients underwent autopsies, one of which revealed disseminated aspergillosis and four of which revealed interstitial pneumonitis and/or acute respiratory distress syndrome and no other organisms. In conclusion, rhinovirus infections may be associated with considerable pulmonary related morbidity and mortality in severely myelosuppressed immunocompromised patients. PMID- 10530442 TI - Editorial response: rhinovirus pneumonia--a clinical entity? PMID- 10530443 TI - Impact of bacterial pneumonia and Pneumocystis carinii pneumonia on human immunodeficiency virus disease progression. Pulmonary Complications of HIV Study Group. AB - The course of pneumonia caused by pyogenic bacteria and Pneumocystis carinii was examined in a multicity cohort study of HIV infection. The median duration of survival among 150 individuals following initial bacterial pneumonia was 24 months, compared with 37 months among 299 human immunodeficiency virus (HIV) infected control subjects matched by study site and CD4 lymphocyte count (P<.001). For 152 subjects with P. carinii pneumonia, median survival was 23 months, compared with 30 months for 280 matched control subjects (P = .002). Median durations of survival associated with the two types of pneumonia differed by only 47 days, despite a higher median CD4 lymphocyte count associated with bacterial pneumonia. These results suggest that both P. carinii pneumonia and bacterial pneumonia are associated with a significantly worse subsequent HIV disease course. The similarity of prognosis after one episode of bacterial pneumonia vs. an AIDS-defining opportunistic infection and the proportion of cases occurring in association with a CD4 lymphocyte count of >200 suggest that measures to prevent bacterial pneumonia should be emphasized. PMID- 10530444 TI - Editorial response: do bacterial pneumonia and Pneumocystis carinii pneumonia accelerate progression of human immunodeficiency virus disease? PMID- 10530445 TI - Photo quiz. Retropharyngeal abscess secondary to cervical tuberculous osteomyelitis. PMID- 10530446 TI - Escherichia coli: epidemiology and analysis of risk factors for infections caused by resistant strains. AB - This study analyzes the epidemiology of hospital and community-acquired infections caused by Escherichia coli. The antimicrobial resistance pattern was used to characterize the isolates, and a prospective observational study was performed to assess the relationship between antimicrobial use and bacterial resistance. The study was conducted during a 3-month period in a 1,200-bed tertiary care hospital in Nantes, France. An E. coli infection was diagnosed in 3.8% of the patients (507 of 13,384) admitted to the hospital between 1 January and 31 March 1996. Of the 507 isolates, 205 (40.4%) were resistant to at least one antimicrobial; 40% were resistant to amoxicillin, 30% to amoxicillin/clavulanate, 38% to ticarcillin, and 16% to trimethoprim sulfamethoxazole, while resistance to other antimicrobials was low. Prior receipt of antimicrobial and/or immunosuppressive therapy was significantly associated with infection caused by a resistant organism. PMID- 10530447 TI - Outcome of multidrug-resistant tuberculosis in human immunodeficiency virus infected patients. AB - Among 324 cases of culture-proven tuberculosis from 1988 to 1996 in a hospital in Milan, Italy, 90 (27.8%) were due to Mycobacterium tuberculosis strains resistant to isoniazid and rifampin. Sixty-one of 69 isolates tested had identical restriction fragment length polymorphism patterns. The prevalent strain tested susceptible only to ethionamide and was also resistant to ethambutol, streptomycin, cycloserine, amikacin, kanamycin, terizodone, ofloxacin, rifabutin, rifapentin, and KRM 1648. The median survival time was 94 days. Multivariate analysis showed a trend toward better outcome in the period 1994-1996 (hazard ratio, 4.16; P<.001), and extrapulmonary localization of tuberculosis was the only other independent predictor of a negative outcome (hazard ratio, 2.1; P = .019). The delay from symptoms to beginning of therapy did not seem to be a determining factor in survival time. Standard antituberculosis therapy with four drugs (isoniazid, rifampin, ethambutol, and pyrazinamide) had a higher efficacy than did other regimens with fewer drugs but without a statistically significant difference. PMID- 10530448 TI - Chagas' disease in patients with kidney transplants: 7 years of experience 1989 1996. AB - Chagas' disease was present in 17.22% of persons undergoing kidney transplantation in an Argentine Hospital. The criterion for attributing reactivation of chronic Chagas' disease and transmission of Trypanosoma cruzi to grafts was detection of parasites in blood (patent parasitemia) or tissues. Reactivation was diagnosed in 5 (21.7%) of 23 recipients. Ten (43.4%) of 23 chagasic recipients without reactivation of chronic Chagas' disease had abrogation of serological reactivity. T. cruzi infection was transmitted to 3 (18.7%) of 16 non-chagasic recipients. Reactivation and infection were diagnosed by patent parasitemia or cutaneous panniculitis. For diagnosis, detection of parasites in blood and tissues had more relevance than serology. Sequential monitoring detected early reactivation and infection, permitting application of preemptive or therapeutic therapy with benznidazole, thus inhibiting, in all patients, severe clinical disease produced by a progressive and systemic replication of the parasite. PMID- 10530449 TI - Bacteremic pneumococcal pneumonia in children. AB - We carried out a nationwide retrospective study on bacteremic pneumococcal pneumonia diagnosed from 1985 to 1994 in Finland. The records of 85 children were reviewed for symptoms, signs, laboratory data, and response to antibiotic therapy. The chest radiographs were reevaluated. Bacteremic pneumococcal pneumonia was characterized by high fever (> or =39.0 degrees C in 93%), leukocytosis (WBC count on admission of > or =15x10(9)/L in 84%), and ill appearance (in 79%). Lobar or segmental consolidation was found in 79% of the chest radiographs. Of the patients, 28% had no respiratory symptoms, 6% presented with only gastrointestinal symptoms in addition to fever, and 4% had fever only. Tachypnea was recorded in 19% and rales in 14% of the patients. After onset of antimicrobial treatment, children became afebrile within an average of 22 hours. One patient developed pleural empyema, and none of the patients died. PMID- 10530450 TI - A polyclonal outbreak of predominantly VanB vancomycin-resistant enterococci in northeast Ohio. Northeast Ohio Vancomycin-Resistant Enterococcus Surveillance Program. AB - We studied the molecular epidemiology of vancomycin-resistant enterococci (VRE) isolated in northeast Ohio during 1996 and examined the association between isolation of VRE from samples other than stool and antimicrobial purchases for five Cleveland hospitals. Susceptibility testing and pulsed-field gel electrophoresis were used to analyze 363 isolates from individual patients from 13 hospitals. Susceptibility testing indicated that 287 strains (79%) expressed the VanB phenotype and 76 (21%) expressed the VanA phenotype. The outbreak was polyclonal, with 30 total genotypes. Both VanA and VanB VRE demonstrated multiple genotypes. One genotype was present in all hospitals, suggesting spread between hospitals. For five teaching hospitals, rates of isolation from non-stool sources and from blood correlated positively with purchases of ticarcillin/clavulanic acid (P = .005). In summary, this outbreak demonstrates transmission of VRE between several hospitals in a geographic region and suggests that use of certain beta-lactam antibiotics may be associated with an increased prevalence of VRE. PMID- 10530451 TI - Penicillin-nonsusceptible Streptococcus pneumoniae at San Francisco General Hospital. AB - Positive pneumococcal cultures of specimens from adult inpatients at San Francisco General Hospital (SFGH) during the period of 11 August 1994 through 31 December 1996 were identified retrospectively. Of the isolates recovered, 15.5% were not penicillin-susceptible (MIC, > or =.1 microg/mL). A case-control study was performed to evaluate risk factors for colonization or infection with penicillin-nonsusceptible Streptococcus pneumoniae (PNSP) and outcomes. Cases (n = 65) were adult inpatients with a positive culture for PNSP, and controls (n = 411) were adult inpatients with a positive culture for penicillin-susceptible pneumococci (PSSP) and no evidence of PNSP. Cases were less likely to have pneumococcal bacteremia (15.4% versus 39.4%; P<.001) and less likely to have pneumonia (50.8% versus 68.9%; P = .006). In a multiple logistic regression model, recent hospital admission and absence of bacteremia were independent predictors of penicillin-nonsusceptibility. Human immunodeficiency virus infection, mortality, and length of hospitalization were not significantly different among cases and controls. These data suggest that PNSP may be less virulent (cause less pulmonary infection) and/or less invasive (cause fewer bloodstream infections) than PSSP at SFGH. PMID- 10530452 TI - Kawasaki syndrome-like illness associated with infection caused by enterotoxin B secreting Staphylococcus aureus. AB - Two children had symptoms and clinical signs that were characteristic of the diagnostic criteria for Kawasaki syndrome, temporally associated with Staphylococcus aureus bacteremia. One child initially had focal osteomyelitis that was evident clinically and radiographically, and radiographic evidence of multifocal osteomyelitis was noted at follow-up. The blood-borne S. aureus isolates from these two patients secreted staphylococcal enterotoxin B and were negative for toxic shock syndrome toxin. Staphylococcal and streptococcal superantigens may play a role in the pathogenesis of some cases of Kawasaki syndrome or Kawasaki syndrome-like illness. PMID- 10530453 TI - Prevalence of enterotoxigenic Bacteroides fragilis in adult patients with diarrhea and healthy controls. AB - Enterotoxigenic strains of Bacteroides fragilis (ETBF) have been associated with diarrheal diseases in animals and humans. The enterotoxin of ETBF induces fluid changes in ligated intestinal segments and a cytotoxic response in HT29/C1 cells. An assay based on immunomagnetic-beads separation in combination with PCR was used to detect ETBF in fecal samples from patients with diarrhea and healthy Swedish adults. A total of 922 fecal samples were analyzed in this study, including 728 samples from patients with diarrhea and 194 samples from controls. ETBF was detected in 195 of 728 patients (26.8%) and 24 of 194 healthy controls (12.4%). The difference between the two groups was statistically significant (P<.01). ETBF was the only potential diarrheal agent in 91 (12.5%) of 728 patients. All ETBF-positive samples from patients and controls were also positive in the HT29/C1 assay. The data show high carriage of ETBF in Swedish adults, which might be associated with diarrheal disease. PMID- 10530454 TI - Survey of bloodstream infections due to gram-negative bacilli: frequency of occurrence and antimicrobial susceptibility of isolates collected in the United States, Canada, and Latin America for the SENTRY Antimicrobial Surveillance Program, 1997. AB - During 1997, a total of 4,267 nosocomial and community-acquired bloodstream infections due to gram-negative organisms were reported from SENTRY hospitals in Canada (8 sites), the United States (30 sites), and Latin America (10 sites). Escherichia coli was the most common isolate (41% of all gram-negative isolates), followed by Klebsiella species (17.9%), Pseudomonas aeruginosa (10.6%), and Enterobacter species (9.4%). For all gram-negative isolates combined, the most active antimicrobials tested were meropenem, imipenem, and cefepime. The quinolones levofloxacin (MIC90, 2 microg/mL), ciprofloxacin (MIC90, 1 microg/mL), gatifloxacin (MIC90, 2 microg/mL), sparfloxacin (MIC90, 2 microg/mL), and trovafloxacin (MIC90, 2 microg/mL) were also active against most isolates. Bloodstream infection isolates from Latin America were uniformly more resistant to all classes of antimicrobial agents tested than were isolates from Canada or the United States. Resistance phenotypes consistent with extended-spectrum beta lactamase production were also most common among E. coli and Klebsiella species from Latin America. Further investigation of the reasons for regional differences in resistance patterns is needed, as is ongoing surveillance to detect resistance trends and to guide antimicrobial use. PMID- 10530455 TI - Self-reported bacterial infections among women with or at risk for human immunodeficiency virus infection. AB - Bacterial infections are a major cause of morbidity and mortality in persons with human immunodeficiency virus (HIV) infection, particularly women. We performed a cross-sectional analysis of a history of bacterial infections among 1,310 women with or at risk for HIV infection. HIV-seropositive women were significantly more likely than seronegative women to report recent and lifetime histories of bacterial infection, even after history of injection drug use since 1977 was adjusted for; this included recent pneumonia (odds ratio [OR], 3.2; 95% confidence interval [CI], 1.5-6.6), sinusitis (OR, 1.4; 95% CI, 1.0-2.0), and urinary tract infection (OR, 1.5; 95% CI, 1.1-2.1). Compared with HIV-negative women, women with CD4 cell counts of <200 were about eight times more likely to report recent pneumonia (OR, 7.8; 95% CI, 3.4-17.7); those with CD4 cell counts of 200-500 were almost three times more likely to do so (OR, 2.6; CI, 1.2-5.7). Logistic regression analysis revealed that only CD4 cell category and a recent history of smoking had a significant relationship to self-reported pneumonia. PMID- 10530456 TI - Persistent high risk of diarrhea among foreigners in Nepal during the first 2 years of residence. AB - Although numerous studies have shown that diarrhea is the most common illness occurring during the first few weeks of travel, systematic studies of the incidence of diarrhea during long-term residence in developing countries have not been performed. We conducted a cohort study of the incidence and etiology of diarrhea among 77 expatriate adults who had lived in Nepal for <2 years. Persons were followed prospectively for up to 1 year (mean, 9 months). The incidence of diarrhea during the surveillance period was 3.3 episodes of diarrhea per person per year, or 0.27 episodes per person per month. The annual attack rate of specific pathogens was 42% for enterotoxigenic Escherichia coli, 32% for Cyclospora species, 16% for Giardia lamblia, 16% for Shigella species, 10% for Campylobacter species, > or =10% for rotavirus, and 6% for Entamoeba histolytica. This study suggests that adult persons from developed countries who move to developing countries such as Nepal remain at high risk for diarrhea during their first 2 years of residence. PMID- 10530457 TI - Pigeon pneumonia in provence: a bird-borne Q fever outbreak. AB - Q fever is a widespread zoonosis caused by Coxiella burnetii, an obligate intracellular bacterium, which humans usually acquire through the inhalation of infected dust from subclinically infected mammals. Human infection commonly takes place when an infected mammal gives birth, since high concentrations of the organism are found in the products of conception. Worldwide, cattle, sheep, and goats are the most common reservoirs for C. burnetii. A few investigators have also reported parturient cats and dogs as the sources of human outbreaks of Q fever. During a 10-day period in May 1996, all five members of one family living on a farm in Provence, in the south of France, became ill with fever, general malaise, and cough. All of them had acute Q fever. An epidemiological investigation suggested that this outbreak resulted from exposure to contaminated pigeon feces and ticks. PMID- 10530458 TI - Small-colony variants of Pseudomonas aeruginosa in cystic fibrosis. AB - In the context of chronic lung infection due to Pseudomonas aeruginosa in cystic fibrosis (CF), attention has been focused on the presence of the most common mucoid phenotype. In this study, the presence of small-colony variants (SCVs) of P. aeruginosa in respiratory tract specimens from patients with CF was investigated, and the clinical conditions predisposing to SCVs were analyzed. P. aeruginosa SCVs were isolated from 33 of 86 P. aeruginosa-positive CF patients over a 2-year period. Fast-growing revertants with larger surface colonies could be isolated from SCV populations. Electron microscopy revealed no significant difference in cell size or morphology. MICs of a broad range of antipseudomonas agents for SCVs were two- to eightfold higher than values for revertants. Recovery of SCVs was correlated with parameters revealing poor lung function and was significantly associated with daily inhalation of tobramycin or colistin. PMID- 10530459 TI - A case of severe chronic active infection with Epstein-Barr virus: immunologic deficiencies associated with a lytic virus strain. AB - Infectious mononucleosis (IM) is a self-limiting, lymphoproliferative disease induced by primary infection with the Epstein-Barr virus (EBV). Infection with EBV leads in general to lifelong asymptomatic persistence of the virus. We report the case of a woman who acquired IM at the age of 15 years and then suffered from recurrent high fever, fatigue, and signs of immunologic disorder for more than 12 years until she died of liver failure. In an attempt to describe and to define the course of chronic active infection with EBV, we performed immunologic and molecular assays that demonstrated lytic replication of EBV in the B and T cells of the peripheral blood. In addition to signs of humoral and cellular immune deficiency, we detected an EBV strain with an impaired capability to immortalize B cells and a tendency to lytic replication, thus contributing to the pathogenesis of this chronic active infection. PMID- 10530460 TI - Novelties in the field of anti-infectives in 1998. AB - In 1998, about 30 new medicinal chemical entities in the field of antibacterials and antifungals were presented during the 38th Interscience Conference on Antimicrobial Agents and Chemotherapy, held in San Diego. Among these compounds, only a few of them seem to be candidates for preclinical and clinical development: GAR 936, RWJ 54,418, ME 1209, L-084, FK 483, and DC 756. Other compounds seem to be in the preselection stage or potential lead compounds. PMID- 10530462 TI - Acute infection with Sin Nombre hantavirus without pulmonary edema. AB - Acute infection with Sin Nombre virus has been associated with development of hantavirus cardiopulmonary syndrome (HCPS), a severe cardiopulmonary illness with respiratory failure and shock. We present two cases of Sin Nombre hantavirus infections that did not lead to marked pulmonary complications in two otherwise healthy young adults from Utah and California. Sin Nombre virus causes a wider spectrum of disease severity than has been previously reported. PMID- 10530463 TI - Community-acquired Pseudomonas stutzeri vertebral osteomyelitis in a previously healthy patient: case report and review. AB - Pseudomonas stutzeri is a rare pathogen, and its recovery is often associated with colonization and contamination. We report a case that, to our knowledge, is the first of community-acquired P. stutzeri vertebral osteomyelitis in a previously healthy patient, and we review the literature regarding infections with this uncommon organism. Of the 29 previously reported cases of P. stutzeri infection cited in the literature, only two resulted in death, reflecting the relatively low degree of virulence of this organism. Predisposing risk factors for P. stutzeri infection can be categorized as follows: (1) previous surgery or procedure (implying probable nosocomial acquisition), with or without a foreign body; (2) immunocompromise (an underlying predisposition to infection by an organism with low virulence); (3) immunocompromise and a previous procedure; and (4) previous trauma or superficial infection, with or without possible nosocomial contamination. Our patient lacked any known risk factors for either pyogenic vertebral osteomyelitis or P. stutzeri infection. PMID- 10530461 TI - Persistent Bacillus licheniformis bacteremia associated with an international injection of organic drain cleaner. AB - In recent years manufacturers have developed several products containing saprophytic bacteria, previously believed to be of minimal pathogenicity. We describe the first case of persistent Bacillus licheniformis bacteremia occurring after intentional injection of a consumer product that includes B. licheniformis spores. We postulate that these spores remained in the tissue, unaffected by antimicrobials, ultimately necessitating soft-tissue debridement of the area surrounding the injection site. On the basis of this case and a review of the literature, we submit that some consumer products contain bacteria with demonstrated pathogenicity. Manufacturers should study these bacteria in detail in order to rapidly provide information such as bacteriologic data and antimicrobial susceptibility data to clinicians. PMID- 10530464 TI - Corticosteroids as adjunctive therapy for severe Pneumocystis carinii pneumonia in non-human immunodeficiency virus-infected patients: retrospective study of 31 patients. AB - The aim of this retrospective study was to assess whether corticosteroid adjunctive therapy (CAT) could prevent death in immunocompromised patients with severe Pneumocystis carinii pneumonia (PCP) who do not have human immunodeficiency virus (HIV) infection, similarly to what has been demonstrated for HIV-infected patients. The charts of all non-HIV-infected patients who were admitted to two medical intensive care units between 1988 and 1996 because of severe PCP, defined by an arterial oxygen pressure (determined while the patient was breathing room air) of <70 mm Hg, and who were treated with trimethoprim sulfamethoxazole were analyzed retrospectively. Thirty-one patients met the study criteria, of whom 23 received CAT (within 72 hours of antibiotic therapy) and eight did not receive CAT. The need for mechanical ventilation (10 [43%] of 23 vs. 4 [50%] of 8) and the mortality rate (9 [39%] of 23 vs. 4 [50%] of 8) were similar for the two groups. Although this small study does not have a statistical power high enough to rule out the possibility of a difference, the results suggest that CAT does not improve the survival of non-HIV-infected patients as has been described for HIV-infected patients with severe PCP. PMID- 10530465 TI - Muscle infections caused by Salmonella species: case report and review. AB - We describe a patient with salmonella pyomyositis and review 30 other cases reported during the past 4 decades. Men outnumbered women by 2.9 to 1, and the median age of the patients was 51 years. Approximately one-half the cases were caused by Salmonella enteritidis. Infected vascular aneurysms were observed in seven patients. Prior salmonella infections and local trauma or lesions were common. Diverse underlying conditions, mainly diabetes and human immunodeficiency virus infection, were present in 81% of the patients, and the psoas muscle was involved in 55% of the cases. One-third of the patients died, and relapses were common after a median time of 5 weeks (range, 4.5-27 weeks) in those who survived. Most patients had anemia, and pathogens were recovered from blood samples from two-thirds of the patients. Salmonella should be considered as a causative agent of muscle infections in the appropriate clinical setting, particularly in patients with underlying diseases or preexisting vascular aneurysms. PMID- 10530466 TI - Visceral leishmaniasis in Costa Rica: first case report. AB - We describe a 15-month-old eutrophic immunocompetent male who presented with fever, hepatosplenomegaly, pancytopenia, and hypergammaglobulinemia. Leishmania amastigotes were identified in spleen and bone marrow specimens. In addition, tissue culture, animal inoculation, and isoenzyme analysis identified the parasite as Leishmania donovani infantum or Leishmania donovani chagasi. The infant was successfully treated with an antimonial drug. These findings represent the first case of visceral leishmaniasis reported in Costa Rica. PMID- 10530467 TI - Albendazole therapy for loiasis refractory to diethylcarbamazine treatment. AB - Although diethylcarbamazine is curative in approximately 60% of patients who acquire loiasis as long-term visitors to an endemic area, some individuals continue to have signs and symptoms of infection despite multiple courses of diethylcarbamazine. On the basis of a study of albendazole treatment of loiasis in microfilaremic patients that suggested a macrofilaricidal effect of the drug, we treated three patients who had symptomatic loiasis refractory to more than four courses of diethylcarbamazine with albendazole. At the time of treatment, all patients had persistent symptoms despite decreasing titers of antifilarial antibodies and normal eosinophil counts. Symptoms resolved in all three patients following albendazole therapy. In one patient, nonspecific symptoms recurred 2 years later, but unlike her symptoms before albendazole therapy, they were not accompanied by the appearance of subcutaneous nodules containing adult worms. The other two patients have been symptom-free in the 8 years after albendazole treatment. In summary, albendazole may be useful for the treatment of loiasis when diethylcarbamazine is ineffective or cannot be used. PMID- 10530468 TI - Cutaneous cryptococcosis resembling molluscum contagiosum in a patient with non Hodgkin's lymphoma. PMID- 10530469 TI - Management of progressive outer retinal necrosis with cidofovir in a human immunodeficiency virus-infected patient. PMID- 10530470 TI - Staphylococcus saprophyticus as an unusual agent of nosocomial pneumonia. PMID- 10530471 TI - Failure of a lipid amphotericin B preparation to eradicate candiduria: preliminary findings based on three cases. PMID- 10530472 TI - Serum levels of nitrite and nitrate in patients with systemic inflammatory response syndrome. PMID- 10530473 TI - Cutaneous phaeohyphomycosis caused by Veronaea bothryosa in a liver transplant recipient successfully treated with itraconazole. PMID- 10530474 TI - Cutaneous aspergillus invasion from sinusitis. PMID- 10530475 TI - Early Microascus cinereus endocarditis of a prosthetic valve implanted after Staphylococcus aureus endocarditis of the native valve. PMID- 10530476 TI - Hydroxyurea toxicity in human immunodeficiency virus-positive patients. PMID- 10530477 TI - Tuberculosis and porphyria. PMID- 10530478 TI - Antibiotic therapy for Lyme disease in a population-based cohort. PMID- 10530479 TI - Bacteremia following placement of intracervical laminaria tents. PMID- 10530480 TI - Acute pulmonary edema complicating ovale malaria. PMID- 10530481 TI - First case of Pasteurella gallinarum isolation from blood of a patient with symptoms of acute gastroenteritis in Japan. PMID- 10530482 TI - Salmonella wernigerode infection--report of the first human cases in the United States. PMID- 10530483 TI - Orbital apex syndrome and cavernous sinus thrombosis due to infection with Staphylococcus aureus and Pseudomonas aeruginosa. PMID- 10530484 TI - Activity of the combination of nelfinavir and saquinavir against human immunodeficiency virus after failure of prior protease inhibitor therapy. PMID- 10530485 TI - Diagnosis of Kingella kingae arthritis by polymerase chain reaction analysis. PMID- 10530486 TI - Disseminated toxoplasmosis after liver transplantation. PMID- 10530487 TI - Dual nucleoside therapy in resource-poor and medium-income countries. PMID- 10530488 TI - Powerful radiotherapy for hepatocellular carcinoma. PMID- 10530489 TI - Inflammation-induced cholestasis. AB - Inflammatory cytokines produced in response to various infectious and non infectious stimuli are potent inducers of intrahepatic cholestasis (inflammation induced cholestasis). The cholestatic effect of cytokines results mainly from inhibition of expression and function of hepatocellular transport systems which normally mediate hepatic uptake and biliary excretion of bile salts and various non-bile salt organic anions (e.g. bilirubin). These cytokine effects are reversible and bile secretory function is restored upon disappearance of the inflammatory injury. This review summarizes the clinical, pathophysiological and molecular aspects of inflammation-induced cholestasis. PMID- 10530490 TI - Molecular genetic basis of Gilbert's syndrome. AB - Gilbert's syndrome, an hereditary, chronic, mild, unconjugated hyperbilirubinaemia resulting from impaired hepatic bilirubin clearance and otherwise normal liver function, is arguably the most common syndrome known in humans. Recent molecular genetic studies have determined that the clinical phenotype can be described by a dinucleotide polymorphism in the TATA box promoter of the bilirubin uridine diphosphate-glucuronosyltransferase (UGT-1A1) gene, most frequently (TA)7TAA, affecting up to 36% of Africans, but only 3% of Asians. However, a second common heterozygous mutation in the coding exon 1 of the UGT-1A1 gene (G71R) can also cause the Gilbert's phenotype in Japanese and Asians. The clinical phenotype may not be apparent as frequently as the determined genotype, due to environmental factors such as alcohol-induced hepatic bilirubin glucuronidation, reducing serum bilirubin levels and causing a latent condition. Gilbert's disease is a contributory factor of prolonged neonatal jaundice in breast-fed infants and may precipitate jaundice when coinherited with other disorders of haem metabolism. The genetic variation described as Gilbert's syndrome may lead to pharmacological variation in drug glucuronidation and unexpected toxicity from therapeutic agents. PMID- 10530491 TI - Role of complement regulatory membrane proteins in ischaemia-reperfusion injury of rat gastric mucosa. AB - BACKGROUND: The role of complement in ischaemia-reperfusion injury has not been well investigated. 5I2 is a monoclonal antibody (mAb) directed against a rat membrane inhibitor of the C3 convertase step, which is the rat counterpart of mouse Crry/p65. 6D1 is a mAb against rat CD59 which inhibits the formation of membrane attack complexes. METHODS: We visualized the tissue distribution of these membrane inhibitors in rat gastrointestinal tract by immunohistochemical staining with the appropriate mAb. Then, we tested the hypothesis that complement regulatory proteins protect rat gastric mucosa against ischaemia-reperfusion stress by using these mAbs. Gastric mucosal integrity was continuously monitored by measuring the blood-to-lumen clearance of [51Cr]-labelled ethylenediaminetetraacetic acid (EDTA) under control conditions, during ischaemia and after reperfusion. RESULTS: Rat 6D1 and 5I2 antigens were both widely distributed and predominantly expressed on smooth muscle and endothelial cells in gastrointestinal tracts. Blockade of complement regulatory proteins with 5I2 and 6D1 mAbs resulted in a significant increase in [51Cr]-EDTA clearance after reperfusion. CONCLUSIONS: These findings support the hypothesis that endogenous complement regulatory proteins may act as important protective factors against ischaemia-reperfusion stress in rat gastric mucosa. PMID- 10530492 TI - Cytomegalovirus infection of the human gastrointestinal tract. AB - BACKGROUND: Current interest in cytomegalovirus (CMV) is largely due to an increase in the number of cases of acquired immunodeficiency syndrome and organ transplantation in recent years. The proper recognition of CMV-infected cells in gastrointestinal mucosal biopsies is critical for effective treatment of this condition. METHODS: A total of 6580 endoscopic mucosal biopsies from 6323 patients in the 8-year period (1989-1996) were examined for CMV inclusion bodies. The endoscopic appearance and particularly the presence of ulcers were also analysed. RESULTS AND CONCLUSIONS: The prevalence of cytomegalovirus (CMV) inclusions was 9 per thousand in the gastrointestinal mucosal biopsies from an unselected group of patients. Of the 54 patients with CMV infection, 37 were immunocompromised and 17 apparently immunocompetent. Typical Cowdry inclusions and atypical inclusions were present, the latter more frequently in immunocompromised patients. The maximum prevalence of inclusions was in the oesophageal mucosa in immunocompromised individuals. PMID- 10530493 TI - Assessment of decisions in the treatment of Helicobacter pylori-related duodenal ulcer: a cost-effectiveness study. AB - AIMS: Many treatment trials for Helicobacter pylori have been reported but few have evaluated treatment in terms of both cost and effectiveness. It is important to find a therapy with a high eradication rate and low cost, especially in China. The aim of the study is to assess the efficiency of therapy for duodenal ulcers, including ulcer healing, H. pylori eradication and ulcer recurrence. METHODS: Ninety-six consecutive patients with duodenal ulcers and H. pylori infection were randomly allocated into two groups: AMT group (amoxycillin + metronidazole + tagamet); OA group (omeprazole + amoxycillin). Side-effects were recorded during the treatment period. Endoscopic examinations were repeated at the 7th or 8th week to assess ulcer healing. Patients were followed up for 6 months and repeat endoscopy was performed. Ulcer healing rate, H. pylori eradication rate and ulcer recurrence rate were compared. All costs were recorded and a cost-effectiveness analysis was conducted. RESULTS: In the AMT and OA groups, the ulcer healing rate was 83.7 and 93.5%, respectively (P = 0.27). The eradication rate of H. pylori was 65.1 and 69.6%, respectively and was significantly higher in patients with an ulcer diameter < or = 1 cm compared with those with an ulcer diameter > 1 cm, irrespective of treatment group. There was no difference in recurrence rate, duration of pain or the time lost because of the disease. Moderate or severe side effects were found in 8.9% in AMT group and 6.5% in OA group. The cost of treatment for ulcer healing, H. pylori eradication and reduction in ulcer recurrence were all lower in the AMT group than in the OA group. Sensitivity analysis supported the result that AMT was more cost effective than OA. CONCLUSIONS: The AMT therapy was more effective and less costly than the OA therapy, especially in patients with H. pylori-related duodenal ulcers < 1 cm diameter. PMID- 10530494 TI - Effect of immersion of biopsy forceps in formalin on tissue urease activity. AB - BACKGROUND AND AIMS: It is routine practice to wash biopsy forceps that have been immersed in formalin solution before taking gastric biopsies to test for urease activity as formalin is thought to inactivate the urease enzyme. The aim of this study was to assess the effect of pre-immersion of biopsy forceps in formalin solution on the ability to detect Helicobacter pylori urease activity in biopsies obtained with the same forceps. METHODS: Two hundred consecutive patients undergoing gastroscopy who had macroscopic evidence of possible H. pylori infection had an initial antral biopsy taken using sterile forceps for determining biopsy urease activity. The same forceps were then used to obtain an antral biopsy for histological examination. The forceps were then used, without washing off any adherent formalin solution, to obtain a further antral biopsy for urease testing. RESULTS: The concordance rate for urease tests, with or without formalin exposure, was 100% (95% confidence interval (CI) 98.2-100%). Fifty-six of 200 patients (28%) were found to have urease-positive biopsies. Of these, 52/56 (92.9%) had identifiable H. pylori on histopathology. One hundred and forty four of 200 patients (72%) were found to have urease-negative biopsies. Of these, seven (4.9%) had identifiable H. pylori on histopathology. Six of seven (85.7%) had only a small number of organisms identified. The sensitivity and specificity for the urease test compared with the histopathology as a reference standard was 88.1% (95% CI 79.9-96.4%) and 97.2% (95% CI 94.4-99.9%), respectively. CONCLUSION: Immersion of biopsy forceps in formalin did not reduce the ability to detect urease activity in gastric biopsies taken subsequently. PMID- 10530495 TI - Interleukin-6 and soluble interleukin-6 receptor in the colonic mucosa of inflammatory bowel disease. AB - BACKGROUND: Interleukin-6 (IL-6) has multiple immunological effects on a wide variety of cells and tissues. The expression of IL-6 and IL-6 receptor (IL-6R) may be important to the pathogenesis of inflammatory bowel disease (IBD). METHODS: In the present study, we examined whether mucosal IL-6 and soluble IL-6R were associated with the pathophysiology of IBD using the colonic mucosal specimens obtained from patients with IBD. Enzyme-linked immunosorbent assay was used to measure the levels of IL-6 and sIL-6R in organ cultures of mucosal tissues and in cell cultures of fractionated mucosal cells as well as in the serum. Expression of IL-6 and IL-6R was analysed by reverse transcription polymerase chain reaction analysis using freshly isolated lamina propria mononuclear cells (LPMC). RESULTS: The levels of IL-6 and sIL-6R in organ cultures were substantially elevated in patients with IBD, especially in those with histologically active inflammation. In contrast, considerably higher levels of sIL-6R were detected in patients with other types of colonic inflammation who were included as inflammatory controls, but elevation of IL-6 was less prominent in such patients. The positivity for expression of IL-6 and IL-6R mRNA in LPMC was in parallel with the results obtained in organ cultures. In cell cultures, mucosal macrophages were the main cell type producing both IL-6 and sIL-6R on a per cell basis and other cell fractions including colonic epithelial cells and lymphocytes produced substantially lower amounts of these molecules. The levels of IL-6 and sIL-6R in organ cultures, but not those in the serum, showed a significantly positive correlation with the degree of clinical disease activity in patients with IBD. CONCLUSIONS: Enhanced IL-6/sIL-6R-mediated immune and inflammatory responses may be implicated, at least partly, in the continuation of intestinal inflammation in patients with IBD. PMID- 10530496 TI - Effect of B7.1-transfected human colon cancer cells on the induction of autologous tumour-specific cytotoxic T cells. AB - BACKGROUND: The induction of tumour-specific immunity is important for advanced cancer therapy. There are many molecules, including costimulatory molecules, that have been identified as the activator for tumour-specific T cells. METHODS: To induce autologous tumour-specific cytotoxic T lymphocytes (CTL) more effectively, we studied whether the expression of the B7 gene may render human colon cancer cells able to stimulate autologous peripheral blood mononuclear cells (PBMC) to become tumour-specific cytotoxic T cells. After the establishment of a B7.1 gene transfected tumour cell line, Cw2/B7.1, we first examined its stimulatory effect on autologous PBMC and subsequently, its effect on the induction of parental cell (Cw2)-specific CTL. RESULTS: The results showed that Cw2/B7.1 had a more potent stimulatory effect on PBMC for the induction of both proliferation and cytotoxicity than Cw2. By adding a low dose of interleukin-2, Cw2/B7.1-activated killer cell activity was significantly increased. The specificity of Cw2/B7.1 activated killer cells was demonstrated by the absence of their cytotoxicity to either human lymphocyte antigen (HLA)-A33 identical (ORF) or HLA-non-identical (MT) allogenic colon cancer cell lines. Furthermore, such Cw2-specific cytotoxic activity was significantly reduced by the deletion of CD8+ cells but not CD4+ cells, indicating that these killer cells were mainly CD8+ T cells. CONCLUSIONS: Thus, our results demonstrate that, by using B7.1 gene-transfected tumour cell lines, we effectively induced autologous tumour-specific CTL. These results will provide us with new tools for adoptive immunotherapy for colon cancer patients. PMID- 10530497 TI - Expression of adhesion molecules in hepatic metastases of colorectal carcinoma: relationship to primary tumours and prognosis after hepatic resection. AB - BACKGROUND: Adhesion molecules are closely involved in the development and growth of metastatic tumours. METHODS: We examined the expression of two adhesion molecules in liver metastatic tumours originating from colorectal carcinomas and correlated the expression of E-cadherin (EC) and CD44 variant exon 6 (v6) in these tumours with prognosis after hepatic resection. We examined 39 primary colorectal and 44 liver metastatic tumours obtained from 39 patients and 30 non metastatic colorectal carcinomas as controls. The expression of EC in primary colorectal carcinomas of the metastasis group was significantly lower than in the non-metastasis group (P < 0.05). The expression of EC was low in metastatic liver tumours. RESULTS: The expression of CD44v6 in primary colorectal carcinomas of the metastasis group was significantly higher than in the non-metastasis group (P < 0.01). Expression of CD44v6 was high in metastatic liver tumours. However, there was no correlation between the expression of EC and CD44v6 or between each of these molecules and clinicopathological features of primary and metastatic tumours. Negative expression of EC and CD44v6 was a poor prognostic factor for survival after hepatic resection. CONCLUSIONS: Our results indicate that the lack of expression of EC and CD44v6 in liver metastases of colorectal cancer is associated with poor survival after surgery. PMID- 10530498 TI - Protection of regenerating liver after partial hepatectomy from carbon tetrachloride hepatotoxicity in rats: role of hepatic stimulator substance. AB - BACKGROUND: In the present study, we examined the effect of hepatic stimulator substance (HSS) on modulating hepatotoxicity induced by carbon tetrachloride (CCl4) in the regenerating rat liver. METHODS: Hepatotoxicity was induced in vivo by administering CCl4 to rats that had undergone a 68% partial hepatectomy (PH). In vitro studies were also performed in hepatocytes isolated from PH rats. RESULTS: Hepatic stimulator substance was extracted from regenerating rat liver 96 h after PH and its activity, as determined according to the method of LaBrecque, reached its maximum 96 h after PH. At this time, the mortality induced by CCl4 was significantly decreased in PH rats compared with sham-operated rats (18 vs 59%, P < 0.01). Likewise, changes in serum alanine aminotransferase (ALT) or bilirubin induced by CCl4 were less in rats after 96 h PH. The resistance of regenerating hepatocytes to CCl4 was retained in in vitro samples. Thus, leakage of intracellular ALT or aspartate aminotransferase induced by CCl4 in hepatocytes from 96 h hepatectomised rats was less than in control hepatocytes. HSS demonstrated a protective effect on hepatocytes against CCl4 both in vivo and in vitro. In additional studies, regenerating liver showed increased mitochondrial respiratory activity and enhanced plasma membrane fluidity. The HSS was also shown to increase hepatic mitochondrial respiratory activity and enhance plasma membrane fluidity. Further, the protective effect induced by HSS was correlated with the restoration of mitochondrial respiratory activity and plasma membrane fluidity induced by CCl4. CONCLUSIONS: Regenerating rat liver exhibits resistance to CCl4-induced hepatotoxicity, and the protection afforded by the regenerating state can be attributed, at least in part, to HSS-induced increases in mitochondrial respiratory activity and plasma membrane fluidity. PMID- 10530499 TI - Magnitude of activity in chronic hepatitis C is influenced by apoptosis of T cells responsible for hepatitis C virus. AB - BACKGROUND: The mechanisms of hepatitis C virus (HCV) persistence are unknown, but down-regulation of immune response in a host is likely to play a major role in it. METHODS: To investigate whether T cell apoptosis contributes to such down regulation, we compared peripheral T cell apoptosis in patients with chronic hepatitis C (CHC) with the serum titre of HCV-RNA, serum alanine aminotransferase (sALT) levels and its change, or peripheral T cell proliferation to the recombinant core antigen of HCV, JCC-1. RESULTS: The percentage of apoptosis in T cells was 0.30 +/- 0.31% (mean +/- SD) in 44 patients with CHC and 0.10 +/- 0.05% in 10 normal volunteers (P < 0.05). In patients with CHC there was no statistical correlation between apoptosis in T cells and sALT levels, titre of HCV-RNA or T cell proliferation to JCC-1 antigen. But, in patients showing relatively more apoptosis in T cells (more than mean + 2SD of apoptosis in T cells from normal volunteers), sALT levels decreased. CONCLUSIONS: Thus, T cell apoptosis in patients with CHC is considered to cause a reduction in sALT, contributing to HCV persistence in patients with CHC. PMID- 10530500 TI - A pilot study of three-dimensional conformal radiotherapy in unresectable hepatocellular carcinoma. AB - BACKGROUND: The purpose of this study was to determine the potential role of three-dimensional (3-D) conformal radiotherapy (RT) in treatment of unresectable hepatocellular carcinoma (HCC). METHODS: Thirteen patients were included in this study, which was conducted between 1993 and 1996. Nine patients (group A) were treated with 3-D conformal RT alone because of main portal vein thrombosis, inferior vena cava thrombosis, obstructive jaundice and failure of previous transcatheter arterial chemoembolization (TACE) to control the disease. The remaining four patients (group B) were treated with a combination of TACE and 3-D conformal RT. RESULTS: The greatest dimension of the main tumour in the whole group of patients ranged from 6 to 25 cm (median 15 cm). The radiation dose ranged from 40 to 60 Gy. The tumour response was evaluated by computed tomography scans of the liver 6-8 weeks after completion of radiotherapy. Partial response was observed in 58% of the patients (seven of 12) and minimal response in another 25% of patients (three of 12). One patient could not be evaluated because of the development of hepatic failure 1 month after completion of RT. All patients in group B lived for more than 1 year (range 16-40 months). In group A, one patient who had a large tumour (11 x 10 x 21 cm) with portal vein thrombosis was converted to become resectable after 45 Gy of radiation. The resection specimen revealed no residual cancer cells. This patient is alive longer than 15 months after treatment without the evidence of disease. CONCLUSIONS: Our experience indicates that HCC is more radiosensitive than it was traditionally expected. Three-dimensional reconstruction of tumour and surrounding organs helps to avoid excessive exposure of the liver and adjacent organs to RT and makes it a safer treatment modality for unresectable HCC. Our preliminary data show promise and are worthy of further study to explore the potential role of radiotherapy in the treatment strategy for HCC at various stages of involvement. PMID- 10530501 TI - Case report: oral antioxidant therapy for the treatment of primary biliary cirrhosis: a pilot study. AB - BACKGROUND: The symptoms of the chronic cholestatic liver disease primary biliary cirrhosis (PBC), in particular fatigue and chronic pruritus, adversely affect quality of life and respond only poorly to treatment. Recent studies have suggested that oxidative stress may play a role in tissue damage in cholestatic liver disease and may contribute to symptoms, such as fatigue. We have, therefore, examined, in an open-label pilot study, the therapeutic effects of antioxidant medication on the biochemistry and symptomatology of PBC. METHODS: Patients were randomized to 3 months treatment with a compound antioxidant vitamin preparation (Bio-Antox), four tablets daily (n = 11, group 1), or the combination of Bio-Quinone Q10 (100 mg) with Bio-Antox (n = 13, group 2). Biochemical and symptomatic responses were assessed at 3 months. RESULTS: Significant improvement in both pruritus and fatigue was seen in the patients in group 2. Mean itch visual analogue score improved from 2.4 +/- 3.0 to 0.4 +/- 0.7 post therapy (P < 0.05) while mean night itch severity score improved from 2.6 +/ 1.9 to 1.3 +/- 0.7 (P < 0.05). Nine of 13 of these patients reported less fatigue, while 10/13 showed an improvement in at least one domain of their Fisk Fatigue Severity Score. No significant improvement in itch and only limited improvement in fatigue were seen in the patients in group 1. No change in biochemical parameters was seen in either group. CONCLUSIONS: Antioxidant therapy, as a combination of Bio-Antox and Bio-Quinone Q10, may improve the pruritus and fatigue of PBC. This combination of therapy should be investigated further in a double-blind, placebo-controlled trial. PMID- 10530502 TI - Case report: spontaneous peritonitis caused by Candida albicans. AB - We report a 40-year-old man with decompensated alcoholic liver cirrhosis, who developed spontaneous peritonitis caused by Candida albicans after complete recovery from a recent episode of acute pancreatitis. The patient was successfully treated with amphotericin B. A search of the literature showed that this is the fourth reported case of spontaneous peritonitis caused by Candida albicans. PMID- 10530503 TI - Hepatobiliary and pancreatic: an enhanced lesion in the liver. PMID- 10530504 TI - Gastrointestinal: melanosis coli. PMID- 10530505 TI - Morphologic alteration of the olfactory bulb after acute ozone exposure in rats. AB - The interaction of ozone with some molecules results in an increased production of free radicals. The objective of this study was to identify whether acute ozone exposure to 1-1.5 ppm for 4 h, produced cytological and ultrastructural modifications in the olfactory bulb cells. The results showed that in rats exposed to ozone there was a significant loss of dendritic spines on primary and secondary dendrites of granule cells, whereas the control rats did not present such changes. Besides these exposed cells showed vacuolation of neuronal cytoplasm, swelling of Golgi apparatus and mitochondrion, dilation cisterns of the rough endoplasmic reticulum. These findings suggest that oxidative stress produced by ozone induces alterations in the granule layer of the olfactory bulb, which may be related to functional modifications. PMID- 10530506 TI - Subchronic toluene exposure in low concentrations produces signs of reduced dysfunction in the 6-hydroxydopamine lesioned nigrostriatal dopaminergic system of the rat. AB - The effect of a subchronic (4-week) exposure to low concentrations of toluene (40 or 80 parts per million, ppm) on the brain dopaminergic system has been examined in a rat model of Parkinson's disease. A unilateral lesion of the substantia nigra (SN) dopamine (DA) nerve cells was performed by injection of a low dose of 6-hydroxydopamine (6-OH DA). The peak activity of contralateral rotational behavior induced by apomorphine was significantly decreased after exposure to 80 ppm toluene. Analysis of the neostriatum and SN ipsilateral to the lesion revealed that toluene (80 ppm, but not 40 ppm) counteracted the 6-OH DA-induced reductions of DA tissue levels both within the SN and the neostriatum. Also the lesion-induced reduction of immunoreactivity for tyrosine hydroxylase (TH IR) in the neostriatum was partly counteracted by the toluene exposure (80 ppm). In conclusion, a subchronic exposure to low doses of toluene (80 ppm) leads to signs of reduced dysfunction of the nigrostriatal dopaminergic system after the neurotoxic treatment. PMID- 10530507 TI - Suppression of serotonin hyperinnervation does not alter the dysregulatory influences of dopamine depletion on striatal neuropeptide gene expression in rodent neonates. AB - Sixty days following neonatal dopamine depletion (>98%) with 6-hydroxydopamine, preprotachykinin and preprodynorphin mRNA levels were significantly reduced (67 and 78% of vehicle controls, respectively) in the anterior striatum as determined by in situ hybridization while preproenkephalin mRNA expression was elevated (133% of vehicle controls). Suppression of the serotonin hyperinnervation phenomenon in the dopamine-depleted rat with 5,7-dihydroxytryptamine yielded no significant alterations in reduced striatal preprotachykinin (66%) or preprodynorphin (64%) mRNA levels, while preproenkephalin mRNA expression remained significantly elevated (140%). These data suggest that striatal serotonin hyperinnervation does not contribute to the development of dysregulated striatal neuropeptide transmission in either direct or indirect striatal output pathways following neonatal dopamine depletion. PMID- 10530508 TI - Parathyroid hormone-related protein protects against kainic acid excitotoxicity in rat cerebellar granule cells by regulating L-type channel calcium flux. AB - The parathyroid hormone-related peptide (PTHrP) and PTH/PTHrP receptor genes are widely expressed in the CNS and both are highly expressed in the cerebellar granule cell. We have shown previously that PTHrP gene expression in granule cells is depolarization-dependent in vitro and is regulated specifically by Ca2+ influx via L-type voltage-sensitive calcium channels (L-VSCCs). Kainic acid induces long-latency excitotoxicity in granule cells via L-VSCC-mediated Ca2+ influx. Here, we show that PTHrP is just as effective as the L-VSCC blocker, nitrendipine (NTR), in preventing kainate excitotoxicity. A competitive inhibitor of PTHrP binding abrogates its neuroprotective effect. Both NTR and PTHrP decrease 45Ca2+ influx to the same degree. These findings suggest that PTHrP functions in an autocrine/paracrine neuroprotective feedback loop that can combat L-VSCC-mediated excitotoxcity. PMID- 10530509 TI - Detection of alpha-L fucose containing carbohydrates in mouse immature olfactory neurons. AB - The cellular location of alpha-L fucose was studied in mice olfactory epithelium (OE) using the Ulex europaeus agglutinin-I (UEA-I) and Tetragonolobus purpureas agglutinin (TPA). In adult mice, UEA-I and TPA revealed a layer of cells that mostly correspond to immature olfactory neurons. Moreover, autoradiographic studies after 3H-T incorporation showed that UEA-I cell labelling occurred during the week following the division of neural cell precursors. In newborn animals, active neurogenesis process is associated with a higher number of UEA-I labelled cells. Olfactory neurogenesis may thus involve a transient event of protein fucosylation, preceding axonal growth. PMID- 10530510 TI - Mutations in the gene encoding human persyn are not associated with amyotrophic lateral sclerosis or familial Parkinson's disease. AB - The synucleins are a family of small proteins expressed in nervous tissue, which have been implicated in neurodegeneration. Using single strand conformation polymorphism analysis we screened for polymorphisms and mutations in the gene encoding human persyn, a recently discovered member of the synuclein family, in controls, patients with sporadic or familial amyotrophic lateral sclerosis (ALS) or familial Parkinson's disease (PD). Six polymorphisms in the genomic sequence of persyn were detected; A590C (5' untranslated region), G1943C (exon 3), G2049A (intron 3), T4502C (intron 3), T4552A (exon 4) and C5019T (3' untranslated region). However no associations with disease state were found in our sample group. PMID- 10530511 TI - Buprenorphine potentiates L-DOPA-induced contralateral rotation in 6 hydroxydopamine-treated rats. AB - Animals are commonly given opioid analgetics such as buprenorphine for post operative pain management. In this study, the effect of the analgetic buprenorphine, a partial mu receptor agonist and kappa receptor antagonist, on L DOPA-induced contralateral rotation was measured in 6-hydroxydopamine (6-OHDA) treated rats. Male Sprague-Dawley rats received dopamine-depleting brain lesions by infusion of 6-OHDA into the medial forebrain bundle. After the procedure, buprenorphine was administered (430 microg/kg, s.c.) to 17 of 54 animals. Three weeks after 6-OHDA treatment, animals were given benserazide HCI (25 mg/kg, i.p.) and L-DOPA (4 mg/kg, i.p.). Contralateral rotations were monitored for 2 h. Animals receiving buprenorphine had significantly higher rates of rotation as compared with non-buprenorphine-treated animals (P = 0.023). The results suggest that buprenorphine sensitizes animals to the effects of L-DOPA. PMID- 10530512 TI - Frontal midline theta rhythms reflect alternative activation of prefrontal cortex and anterior cingulate cortex in humans. AB - Frontal midline theta rhythm (Fm theta) often appears on electroencephalogram (EEG) during consecutive mental tasks. To clarify the source of rhythmic activity, magnetoencephalogram (MEG) and EEG were simultaneously measured in six healthy volunteers during different mental tasks using whole head MEG system. MEG records were averaged every one cycle of Fm theta rhythms using individual positive peaks of Fm theta waves in Fz EEG as a trigger. Averaged theta components of MEG signals were analyzed with a multi-dipole model. Two sources were estimated to the regions both of the prefrontal-medial superficial cortex and anterior cingulate cortex (ACC). These regions were alternatively activated in about 40 to 120 degrees phase shift during one Fm theta cycle. From above results, we hypothesize that appearance of Fm theta during consecutive mental tasks reflects alternative activities of the medial prefrontal cortex and ACC. PMID- 10530513 TI - Biochemical and conformational variability of human prion strains in sporadic Creutzfeldt-Jakob disease. AB - The pathogenesis of prion (PrP) diseases is thought to be related to conformational changes of a normal cellular protein, PrPC, into a protease resistant protein called PrPSc, which is infectious by itself. A difficulty with this 'protein only' hypothesis is the existence of numerous PrP strains, that require PrPSc to have multiple conformations. Sporadic Creutzfeldt-Jakob disease (CJD), which accounts for nearly 80% of human prionoses, was reported to include at least two 'strains' termed types 1 and 2 which differ by electrophoretic patterns of their proteinase K (PK)-resistant fragments (PrP27-30). We have analyzed the biochemical and structural properties of PrPSc and PrP27-30 isolates from six sporadic CJD patients. Fourier transform-infra-red spectroscopy, PrP27 30 glycosylation patterns and studies of PK sensitivity revealed a striking heterogeneity. Furthermore, one isolate yielded a PrP27-30 fragment with a lower mobility clearly different from previously described sporadic CJD types. Although the average beta-sheet content was higher among type 1 isolates, there was overlap between the two types. Our study suggests that human sporadic CJD-related prions display a significant heterogeneity. PMID- 10530514 TI - Polymorphisms of the human apolipoprotein E promoter and bleomycin hydrolase gene: risk factors for Alzheimer's dementia? AB - In addition to the apolipoprotein E (ApoE) tri-allele polymorphism, genetic variants of the apolipoprotein E promoter (-491A/T) and the bleomycin hydrolase (BH-PEN) gene have also been proposed as genetic risk factors for Alzheimer's dementia (AD). Since reports about the relevance of these polymorphisms for the pathogenesis of AD have been contradictory, we performed an association study with some modifications. First, the control group in this study was made up of non-demented psychiatric inpatients, rather than healthy subjects. This procedure allows the specificity of the relationship between a given genotype and AD (as opposed to other psychiatric disorders) to be determined. Second, as an alternative to preexisting relatively time consuming techniques, BH-PEN polymorphism was determined using a simplified method based on PCR genotyping. We found a significant linkage disequilibrium between the -491A/T and ApoE polymorphisms. However, no direct association was observed between the -491A/T or BH-PEN polymorphism and AD. PMID- 10530515 TI - Decline in spontaneous activity of group Aalphabeta sensory afferents after sciatic nerve axotomy in rat. AB - Changes are observed in the strength of central synaptic transmission and the firing behavior of primary afferents damaged by peripheral nerve injury. To clarify the relationship between synaptic strength and amount of spontaneous activity, firing behavior was studied in adult, male Sprague-Dawley rats in which sciatic nerve afferents were axotomized. Intra-axonal recordings were taken from Aalphabeta afferents within 7 h (acute, n = 309), at 3 days (n = 228), or at 10 days (n = 230) after sciatic nerve cut. The proportion of spontaneously discharging afferents fell from 22% in the acute group to < or = 13% in chronic groups. Thus, neither the progressive decline in the strength of central synaptic transmission from cut primary afferents nor the altered sensation observed after nerve cut can be explained by chronic changes in spontaneous activity of cut Aalpha/Abeta afferents. PMID- 10530516 TI - Immunoreactive Akt, PI3-K and ERK protein kinase expression in ischemic rat brain. AB - In order to clarify the role of protein kinases in ischemic brain injury, the spatiotemporal expression of immunoreactive serine-threonine kinase Akt, phosphatidylinositol 3-kinase (PI3-K) and extracellular signal-regulated kinase (ERK) were examined at 3, 8, or 24 h after permanent middle cerebral artery occlusion (MCAO) in rats. Weak staining for these protein kinases was found in both cortical and caudate neurons in sham controls. The staining for Akt-1 and PI3-K was increased at 3-8 h in the ischemic penumbral region and declined at 24 h. A slight induction of these kinases was observed in the ischemic core region. Robust expression of ERK was noted at 3-8 h in most neurons in the area of ischemia. At 24 h, ERK continued to be expressed in the ischemic penumbra, but decreased in the ischemic core. These findings suggest that the signaling for Akt and PI3-K are different from the ERK dependent signal transduction during ischemic brain injury. PMID- 10530517 TI - Some physiological and pharmacological properties of slow depolarization of substantia gelatinosa neurons by repetitive stimulation of C-fibers of dorsal root in adult rat spinal cord slices with dorsal root attached. AB - Transverse slices of the spinal cord with dorsal root attached were prepared from adult rats and used to record the response of the spinal substantia gelatinosa neurons evoked by repetitive stimulation of the dorsal root with a whole cell patch clamp method. The repetitive stimulation of 10 trains (20 pulses at 100 Hz/train) with an intensity necessary to activate C-fibers, but not A-fibers alone, evoked slow depolarization (SD) during it and the SD disappeared within 1 min after the termination of it. The SD was completely inhibited by 500 nM of tetrodotoxin (TTX), but not significantly changed by 50 nM of TTX, nor 20 microM CNQX + 50 microM AP5, nor 1 microM CP-99994. DAMGO inhibited the SD in a concentration dependent manner (10 nM-1 microM), but 1 microM DPDPE and 1 microM U-50488H did not. The inhibitory effect of DAMGO (1 microM) was reversed by naloxone (1 microM). These results suggest that the SD of substantia gelatinosa neurons evoked by repetitive stimulation of C-fibers of the dorsal root is an event relevant to nociception in the spinal dorsal horn. PMID- 10530518 TI - Localization of peptidylarginine deiminase type II in a stage-specific immature oligodendrocyte from rat cerebral hemisphere. AB - Myelin basic protein (MBP) is composed of multiple charged isomers as the products of various posttranslational modifications. The least cationic component contains six citrulline residues converted from arginine residues by peptidylarginine deiminase (PAD). The modified MBP differs markedly from unmodified MBP in the ability to aggregate acidic lipid vesicles. However, the localization of PAD in brain has remained rather elusive. We performed Western blotting and immunocytochemical analyses of PAD type II and found that it was present in stage-specific immature oligodendrocytes but not in either type-1 astrocytes or neurons. We also confirmed that only the oligodendrocyte homogenate contained the PAD activity utilizing a sensitive method to detect citrulline containing proteins. These data suggest that PAD type II localized in oligodendrocytes is responsible for deiminating MBP. PMID- 10530519 TI - Characterization of molecular forms of acetyl- and butyrylcholinesterase in human acoustic neurinomas. AB - Acoustic neurinomas were sequentially extracted with saline and saline-Triton X 100 buffers. Detergent was required to detach the bulk of acetylcholinesterase (AChE), but butyrylcholinesterase (BuChE) was mostly released with saline buffer. Sedimentation analysis and hydrophobic chromatography revealed that neurinomas contain principally amphiphilic AChE tetramers, dimers and monomers, and hydrophilic BuChE tetramers. The AChE dimers and monomers remained amphiphilic after incubation with phosphatidylinositol-specific phospholipase C (PIPLC), after or without prior treatment with alkaline hydroxylamine, which shows that, in contrast to the meningioma AChE dimers and monomers, the neurinoma isoforms are devoid of glycolipid. Neurinoma AChE reacted with the antibodies HR2 and AE1 raised against AChE from human brain or erythrocyte, whereas BuChE bound to a sheep antiserum. PMID- 10530520 TI - The tau gene in progressive supranuclear palsy: exclusion of mutations in coding exons and exon 10 splice sites, and identification of a new intronic variant of the disease-associated H1 haplotype in Italian cases. AB - Mutations in coding exons or exon 10 5'-splice-site of the gene for microtubule associated protein tau can cause chromosome 17-linked frontotemporal dementia and parkinsonism (FTDP-17). We sequenced the 11 coding exons plus exon-intron boundaries of the tau gene in 15 cases of progressive supranuclear palsy (PSP), and found no mutations in coding exons or exon ten 5'-splice sites. These data indicate that typical PSP is not associated with tau gene mutations similar to those causing FTDP-17. We also observed a +39deltaG base change in the intron following exon 4 in three out of 69 PSP cases (all three Italians), whereas it was not found in 150 Dutch controls and once in 112 Italian controls. The +39deltaG variant arose in the context of the PSP-associated tau H1 haplotype. Although a pathogenic role cannot be entirely excluded, +39deltaG is likely to be a rare polymorphism that may be in linkage disequilibrium with a biologically relevant locus inside or near to the tau gene. PMID- 10530521 TI - Cessation of human motor unit discharge during sustained maximal voluntary contraction. AB - The purpose of this study was to determine whether cessation of motor unit discharge contributes to fatigue in human subjects. Multiple fine-wire and tungsten microelectrodes were inserted into the extensor digitorum or extensor indicis muscles of the forearm in an attempt to record the activity of the same motor unit from different locations within either muscle while subjects maintained a maximal voluntary contraction of the finger extensors until force dropped by approximately 50%. The activities of 13 motor units were followed for extended periods during the fatigue task. Of these, six appeared to cease discharging prior to the end of the task, which could not be attributed to electrode movement. These findings suggest that some motor neurons may not be able to discharge continuously in the presence of sustained volitional synaptic drive or that excitatory drive may diminish during maximal voluntary effort. PMID- 10530522 TI - Injury surveillance in a pediatric emergency department. AB - In this study we have tried to determine physician success in the collection of injury data during the emergency department visit. Prospective data were collected from all children between the ages of 0 to 18 treated for an injury. Data were collected at the time of the visit and by chart review the next day. At an urban, university-affiliated, children's hospital, data were collected on 2,156 injured children. Fifty-one percent of the data forms were completed by the treating physician. Physician completion rate was lower on the weekends (46%) than on weekdays (52%, P = .02). The most common mechanisms of injury were falls (34%), motor-vehicle crashes or pedestrians struck (13%), and nonintentional struck by blunt object (12%). The most common mechanism of injury in all age groups was falls. Our results demonstrate that emergency physicians are not successful data collectors. However, when physician data collection is combined with next-day review of patient records, virtually 100% of patients are captured. Active emergency department data collection is important because in contrast to studies which use hospital discharge and mortality data, we found that overall falls account for more injuries presenting to the ED than transportation-related causes. An active surveillance system in emergency departments that does not require extra work on the part of the treating physician would be ideal and may give a more comprehensive description of the scope of the injury problem. PMID- 10530523 TI - ED use of flexion-extension cervical spine radiography in the evaluation of blunt trauma. AB - Dynamic cervical spine radiography (CSR) is used to detect ligamentous instability. We investigated the ED use of dynamic CSR through a retrospective descriptive review using a convenience sample study design at a university emergency department. Adult blunt trauma patients with static (lateral, AP, odontoid) and dynamic (flex, extend) CSR participated. 451 patients (52% male with mean age of 33.6 years) met entry criteria. Injury mechanisms were 74% MVA, 12% fall, 8% direct trauma, and 6% other. Indications for dynamic CSR were 100% traumatic mechanism, 86% neck pain, 70% midline neck tenderness, and 18% abnormal static CSR. Static CSR were normal in 372, 5 of which had abnormal dynamic CSR (5 cervical contour line disruption [CCLD], 2 posterior element abnormality [PEA]); of these 5 patients, none required invasive stabilization. Static CSR were abnormal in 79 patients (38 CCLD, 30 lordotic curve reversal, 17 PEA, 4 soft tissue swelling) of which 16 had abnormal dynamic CSR (9 increased CCLD, 4 PEA, and 4 fracture); of these 16 patients, 4 required invasive stabilization. Final diagnoses were 428 cervical soft tissue injury, 11 subluxation, 8 fracture, 2 fracture-subluxation, and 2 spinal cord injury without radiographic abnormality. Spine consultation was made in 12%. Stabilization therapy was required: 21 soft collar, 4 surgical, 3 halo-device, and 5 other. No complications of dynamic CSR were noted. The blunt trauma patient with neck complaints and an abnormal static CSR was more likely to have an abnormal dynamic CSR demonstrating a cervical injury requiring stabilization compared to patients with normal static CSR. PMID- 10530524 TI - Anxiety levels in EMS providers: effects of violence and shifts schedules. AB - We tried to measure anxiety levels in emergency medical service (EMS) providers to determine the effects of (1) having had a violent encounter during a shift and (2) different shift schedules, conducting a prospective observational study over 3 months in an urban EMS system setting. A convenience sample of 23 EMTs and 40 EMT-Ps was observed. Anxiety levels were measured using the Spielberger State Trait Anxiety Inventory. A total of 99 inventories were completed by 63 EMS providers. The mean state (32.6+/-8) and trait (31.7+/-7.1) scores were less than normative scores (35.7+/-10.4 and 34.9+/-9.2 respectively) for working adult males (P = .004 and .007, respectively). Paramedics had lower anxiety scores than basic EMTs (P = .015 and .039) and years of experience also decreased anxiety scores (P < .0001). There was no significant difference in state scores between those EMS providers who had encountered violence during the preceding 12 hours and those providers who had not. Comparisons of state scores of providers assessed at hour 12 of a 12 hour shift, hour 12 of a 24 hour shift, and hour 24 of a 24 hour shift failed to show any significant differences. Although the working environment of the EMS provider contains numerous stressors and uncertainties, this population of providers were no more anxious than the general working public. Advanced training and years of experience decreased anxiety. Violent encounters during a shift did not appear to affect anxiety levels. There was no difference in anxiety levels in providers who worked 12 and 24 hour shifts. PMID- 10530525 TI - Is hospitalization necessary for treatment of SVT? Predictive variables for recurrence and negative outcome. AB - In this article we attempt to identify variables that may predict which pediatric patients presenting with supraventricular tachycardia to the emergency department are at risk for immediate recurrence or negative outcome. We studied an 11-year chart review and follow-up of pediatric patients presenting to the pediatric ED with a diagnosis of SVT. Recurrences of SVT during the first 24 hours after initial episode and occurrences of negative outcome (ie, respiratory distress, cardiac arrest) during hospital stay were measured. 51 patients met our inclusion criteria: 74.5% were treated as inpatients. 18% experienced a recurrence during 24 hours; 3 patients were less than 3 months of age. Recurrence within 90 minutes of conversion occurred in 4/7 patients. All negative outcomes (3.9%) occurred in the less than 3-month age group. Patients less than 3 months or with immediate supraventricular tachycardia recurrence may have a higher probability of recurrence and negative outcome and therefore may require hospitalization. PMID- 10530526 TI - Emergency department presentation of idiopathic intracranial hypertension. AB - Idiopathic intracranial hypertension (IIH), or pseudotumor cerebri, is a syndrome characterized by an elevated intracranial pressure in the absence of a focal lesion, infective process, or hydrocephalus. New onset IIH may present to the emergency department in a variety of ways. To describe the etiologic associations and clinical features in this disorder, we performed a retrospective analysis of consecutive emergency department patients with new onset IIH during the calendar years 1987-1996. A total of 52 patients met all study criteria. The mean patient age was 27+/-8.9 years; the female-to-male ratio was 7:1. An etiologic association could be identified in 85% of cases and included obesity, hypertension, drugs, endocrine, and systemic disorders. Headache was a dominant complaint in most patients (48/52) and associated with dizziness, nausea, and/or visual complaints. Fourteen patients (27%) were not diagnosed on their initial ED visit and were more likely to have atypical clinical features (71% vs. 24%; P = .004). Atypical features included paraesthesias, neck/back pain, unilateral headache, vertigo, and nystagmus. Papilledema, the ophthalmoscopic hallmark of IIH, was not detected initially in 11 patients (21%). These results suggest that IIH is a relatively uncommon neurological illness that may have a variety of causes. The emergency department diagnosis may be complicated by atypical clinical features and a lack of detectable papilledema. PMID- 10530527 TI - Utility of clinical characteristics in identifying depression in geriatric ED patients. AB - We will determine if clinical characteristics can be useful in identifying depression in geriatric Emergency Department (ED) patients. We have provided a cross-sectional observational study of geriatric patients presenting to an urban university-affiliated public hospital. A brief self-rated depression scale (SRDS) was used to identify depression. Clinical characteristics, examined retrospectively, included chief complaint, chronic illnesses, mode and time of arrival and discharge disposition. Relative prevalence of depression was calculated for these clinical characteristics. 70 (27%; 95% CI, 22% to 32%) of 259 patients were found to be depressed by the SRDS. Patients with nonspecific chief complaints were more commonly depressed than patients with system-specific chief complaints, but not significantly (relative prevalence 1.6; 95% CI, 1.0 to 2.4; p = 0.19). The relative prevalence of depression also did not vary significantly when analyzed by specific chronic illness (P = 0.42) except cardiac disease (1.6; 95% CI, 1.1 to 2.4), PM or night arrival (1.3; 95% CI, 0.8 to 2.3; p = 0.17), ambulance use (1.1; 95% CI, 0.7 to 1.7; p = 0.88), or need for medical admission (1.0; 95% CI, 0.7 to 1.5; p = 0.97). Depression is common in geriatric ED patients. Clinical characteristics fail to identify elderly ED patients who are likely to be depressed. Use of a brief SRDS can aid in recognition of depression in this group. PMID- 10530528 TI - Changes in asthma claims in a Medicaid population from 1991-1994. AB - Previous reports have found an increase in asthma prevalence and severity during the 1980s. The purpose of this study was to evaluate changes in asthma claims in a cohort of Medicaid enrollees from 1991 through 1994. A historical study used Ohio Medicaid claims data for fiscal years 1989 through 1994. Adult and pediatric enrollees with at least one claim for asthma care during fiscal year 1989 who subsequently were continuously eligible through fiscal year 1994 were divided into 2 cohorts consisting of those with (Cohort 1) or without (Cohort 2) an asthma claim in 1990. Claims were then analyzed forward from 1991-1994. Patients consisted of 3,027 enrollees, including 2,206 children and 821 adults. Overall emergency department visits increased in both cohorts, primarily due to an increase in adult visits. Visits by children who were preschool-aged in 1989 decreased. Patients in the cohort with an asthma visit in both 1989 and 1990 continued to have an increased frequency of emergency department (ED) visits compared to cohort 2. Outpatient visits decreased in both cohorts but the decrease was greater in cohort 1 in spite of the assumption that these patients should have greater attendance at outpatient clinics. The percentage of patients with an outpatient clinic visit within 3 days of their ED visit also decreased. Admissions decreased in both cohorts indicating that there was not a marked increase in asthma severity. Patients in cohort 1 with ED visits in 2 successive years represent those under relatively poorer control. In spite of a continued higher frequency of ED use, these patients had decreased use of outpatient facilities. Severity to the extent reflected by admissions did not increase over this period. Further research may demonstrate that improved outpatient management and followup of higher frequency ED users may lead to greater asthma control. PMID- 10530529 TI - Exertional heat illness and hyponatremia in hikers. AB - We compared clinical presentation and course of exercise-associated hyponatremia with heat exhaustion among summertime hikers in Grand Canyon National Park. Cases were selected from among hikers who requested medical help from the National Park Service Emergency Medical Service (EMS) or who presented to the medical clinic on the rim of the canyon with complaints related to exercise in the heat. Of 44 patients who had serum samples analyzed, 7 had hyponatremia with clinically significant symptoms and serum sodium levels <130 mmol/L: 3 had grand mal seizures, 2 had other major central nervous system disorders, and 2 had minor neurological symptoms. Seizures and change of mental status distinguished hyponatremia, (P = 0.0002). Indirect evidence suggests that hyponatremic patients were hyperhydrated. Other common symptoms included nausea, vomiting, headache, and dizziness, but these symptoms did not predict the level of serum sodium. When exercise in the heat is prolonged, hyponatremia is suggested either by altered mental status or by seizures without hyperpyrexia or hypoglycemia. No mortality or long-term morbidity occurred in any of these cases of hyponatremia. PMID- 10530530 TI - Evolution of abstracts presented at the annual scientific meetings of academic emergency medicine. AB - There has been a general trend in medicine toward greater sophistication in research design. To assess this trend in emergency medicine, we compared the characteristics of abstracts presented at the 1974, 1983, 1989, and 1997 annual scientific meetings of Academic Emergency Medicine. All 870 abstracts were reviewed by 1 of 3 investigators who determined research design attributes using a standardized classification scheme that has good interrater reliability. Over the last 25 years, the following trends were noted: more surveys (0% v1% v3% v8%, P=.002), more randomized studies (0% v10% v12% v15%, P=.05), and more blinded studies (0% v7% v5% v11%, P=.01). Tests of statistical significance were reported with increasing frequency (8% v26% v59% v 69%, P < .001), as were power calculations (0% v0% v1% v3%, P=.02). During the study period, there were also increases in the median number of authors, proportion of foreign lead authors, and the proportion of studies involving human subjects. These results reflect considerable improvement in the degree of research design sophistication reported in selected abstracts of academic emergency medicine over the study period. Further strategies to assure continued enhancement of emergency medicine research should be explored. PMID- 10530531 TI - Paramedic decisions with placement of out-of-hospital intravenous lines. AB - To determine the incidence of unused out-of-hospital intravenous line (IV) placements, we prospectively studied IV placement in emergency medical services (EMS) patients. Unused IV placement was defined as any patient having an EMS initiated IV that was not used for fluid bolus or medication administration in the field or in the emergency department (ED). Data were analyzed on placement and use of IV lines in the field and in the ED, transport time, years of paramedic practice, and paramedic student presence. Of 290 patients, 165 had an IV initiated (147) or attempted (18). Twenty-nine percent (84 of 290) of the patients received an unused EMS IV. One hundred twenty-five patients had no IV initiated by EMS. Seven subsequently had an IV started and used in the ED, for an undertreatment rate of 2.4% (7 of 290). The presence of a paramedic student increased the odds of an unused IV 1.4 (95% CI, 1.1 to 2.0). IVs are frequently started and not used. PMID- 10530532 TI - The ratio of interleukin-6 to interleukin-10 correlates with severity in patients with chest and abdominal trauma. AB - To evaluate the relation between proinflammatory cytokines and antiinflammatory cytokines after major trauma, we measured pro-inflammatory cytokine (interleukin [IL]-6) and antiinflammatory cytokine (IL-10) concentrations after trauma, and evaluated the relationship between the ratio of IL-6 to IL-10 and injury severity. In 20 patients who sustained chest and abdominal trauma, IL-10, IL-6, and lactate concentrations were measured at 0, 1, 2, and 4 days. The Injury Severity Score (ISS), Acute Physiology and Chronic Health Evaluation (APACHE) II score, and the ratio of IL-6 to IL-10 were calculated. IL-6, IL-10, and lactate concentrations were 197.2+/-28.4 (mean +/- SEM), 71.1+/-10.1 pg/mL, and 46.7+/ 9.4 mg/dL at entry. These concentrations were significantly decreased at day 4. The ratio of IL-6 to IL-10 was 3.11+/-0.47 at entry and was significantly correlated with ISS (r=.872, P<.01), APACHE II score (r=.887, P<.01). The IL-10, IL-6, and lactate concentrations were elevated immediately after major trauma, and the ratio of IL-6 to IL-10 was correlated with injury severity. This suggests that the ratio of IL-6 to IL-10 may be used to predict the injury severity after trauma. PMID- 10530533 TI - Summary statistics for acute cardiac ischemia and chest pain visits to United States EDs, 1995-1996. AB - In this article we describe characteristics of emergency encounters for patients with a diagnosis of acute cardiac ischemia (ACI) and for patients with chest pain complaints. Data are from the National Hospital Ambulatory Medical Care Survey, which includes abstracts from the medical records of a national probability sample of visits to emergency departments (ED). Analysis was limited to records of patients 25 years of age and older with a diagnosis of either confirmed or suspected acute myocardial infarction (AMI) or unstable angina pectoris and records with a nontraumatic chest pain complaint. There was an estimated annual average of 1.2 million visits to EDs by patients 25 years and over with a diagnosis of ACI in 1995-1996, an average annual rate of 7.2 visits per 1,000 persons. Visit rates varied by patient's age, race, and gender. Chest pain was a complaint in three-fourths of all ACI visits. There were an estimated 4.6 million annual ED visits where in patients aged 25 years and older had complaints of nontraumatic chest pain, an average annual rate of 27.7 visits per 1,000 persons. ACI accounted for 11% of all chest pain visits, but the probability of the chest pain visit having an ACI diagnosis varied by patient's age and race. There remains a large amount of variation in treatment for suspected and confirmed AMI, and for patients presenting with chest pain to EDs. PMID- 10530534 TI - Utilization of the 911 system as an identifier of domestic violence. AB - This cross-sectional study was performed to determine (1) whether female victims of domestic violence (DV) are more likely to use the 911 system than nonvictims (NVs) and (2) whether DV and NVs call 911 for different reasons so that 911 may be used as a screening tool for abuse. The study was performed in an academic adult urban emergency department (ED). Ambulatory female patients presenting to the ED were studied. Eligible patients were administered a brief survey by trained research assistants. Questions included (1) history of DV, (2) relationship of assailant to victim, (3) chief complaint, and (4) use of the 911 system. Records of 911 calls were obtained by patient's address. Four hundred sixty-one women were enrolled in the study. One hundred seven (23%) reported a history of DV. Intimate partners accounted for 67.2% of assailants. DV victims were more likely to be single and younger (P < .05). Of DV victims, 77% reported calling 911 for any reason in the past 2 years compared with 47% of nonvictims (difference = 30%; 95% CI, 19%, 40%). DV victims were more likely to call 911 than nonvictims for definite and possible cases of domestic dispute (1.4 v0.5 calls, P = .007; 11.7 v6.1 calls, P = .0003). Victims and nonvictims did not differ in the number of nondomestic dispute calls (8.4 v6.2 calls; P = .15). DV victims were more likely to access the 911 system and call for domestic disturbances compared with nonvictims. 911 calls may serve as an indicator of ongoing abuse and may identify women at risk, providing a potential opportunity for intervention. PMID- 10530535 TI - Anesthetic methods for reduction of acute shoulder dislocations: a prospective randomized study comparing intraarticular lidocaine with intravenous analgesia and sedation. AB - A prospective, randomized, nonblinded clinical trial was undertaken to evaluate whether local intraarticular lidocaine injection (IAL) is equally effective in facilitating reduction of acute anterior shoulder dislocations (AASD) as intravenous analgesia/sedation (IVAS). The setting was an urban, Level 1, trauma center. Patients enrolled presented to the emergency department (ED) with radiographically confirmed AASD and were randomized either to the IVAS group or the IAL group. Ease of reduction and pain associated with reduction were measured subjectively using a 10-point scale. There were 49 patients entered into the study, 20 in the IVAS group and 29 in the IAL group. There was no statistically significant difference between mean +/- SD pain scores of 3.32+/-2.39 in the IVAS group and 4.90+/-2.34 in the IAL group (P = .18), or mean +/- SD ease of reduction scores of 3.32+/-2.36 in the IVAS group and 4.45+/-2.46 in the IAL group (P = .12). Although IVAS tended to have a higher success rate (20 of 20) than IAL (25 of 29) in this study, Kaplan-Meier estimates for delayed time-events curves applying the log-rank test showed that this difference was not statistically significant overall (P = .16). However, with reduction rate evaluated as a function of time delay in seeking treatment, patients presenting 5.5 hours after dislocation were more likely to fail treatment with IAL (P = .00001). Additionally, half of the patients in the IAL group who had experience with IVAS did not favor IAL. Emergency physicians should be aware of IAL as an alternative technique that may be considered in patients when there are reasons to avoid systemic analgesia or sedation. PMID- 10530536 TI - Effect of ED management on ICU use in acute pulmonary edema. AB - Acute pulmonary edema (APE) is a common Emergency Department (ED) presentation requiring admission to an intensive care unit (ICU). This study was undertaken to examine the effect of ED management on the need for ICU admission in patients with APE. ED records of APE patients were abstracted for patient age, prehospital and ED pharmacological treatment, diagnoses, airway interventions, and ICU length of stay (LOS). Statistical analysis was through multiple regression, logistic regression, chi-square, and ANOVA. One hundred eighty-one patients composed the study group. Pharmacological treatment included nitroglycerin (NTG), 147 patients (81%); morphine sulfate (MS), 88 (49%); loop diuretics (LD), 133 (73%); and captopril sublingual (CSL), 47 (26%). Use of CSL and MS were associated with opposing needs for ICU admission. MS use was associated with increased ICU admissions (odds ratio, 3.08; P = .002), whereas CSL use was associated with decreased ICU admissions (odds ratio, 0.29; P = .002). Morphine sulfate use also demonstrated an increased need for endotracheal intubation (ETI) (odds ratio, 5.04; P = .001), whereas CSL demonstrated a decreased need for ETI (odds ratio, 0.16; P = .008). Ninety-three patients required some form of respiratory support. Forty received noninvasive pressure support ventilation (NPSV) from a bilevel positive airway pressure system (BiPAP), and 60 received endotracheal intubation. Some patients received more than 1 form of respiratory support; all other patients received supplemental oxygen only. The ICU-LOS associated with different airway interventions were supplemental oxygen, 0.72 days; BiPAP, 1.48 days; and ETI, 3.70 days (P < .001). Specific ED pharmacological interventions are associated with a decreased need for ICU admission and endotracheal intubation in acute pulmonary edema patients, whereas use of noninvasive pressure support ventilation correlates with a reduction in the ICU length of stay for patients who do require critical care admission. PMID- 10530537 TI - Injury patterns with snowboarding. AB - Snowboarding is a winter sport that has shown a considerable increase in popularity during the last 2 decades. As a result, there has been a continued rise in the number of visits to the emergency department (ED) for injuries sustained while snowboarding. Previous studies have concluded that those injured tend to be male, younger, and more inexperienced than their alpine skiing counterparts. This study examines the injury patterns seen in one ED during peak winter sport recreational months over a 5-year period. This retrospective review describes 71 patients with a broad spectrum of injury patterns, but reports a higher incidence of head and spinal injuries than previously documented. Furthermore, recommendations to prevent future injuries as well as education for first responders and physicians regarding the high likelihood of serious injury is discussed. PMID- 10530538 TI - ED echocardiography for peripartum cardiomyopathy. AB - Although peripartum cardiomyopathy is uncommon, emergency physicians should be knowledgeable of it because of its high morbidity and mortality. Emergency physicians should be alert to the fact that the clinical presentation of peripartum cardiomyopathy is nonspecific. Its clinical manifestations are found in other medical conditions that can present in the late prepartum or postpartum patient. We present a case of peripartum cardiomyopathy that illustrates how its nonspecific respiratory signs and symptoms led to an initial diagnosis of pulmonary embolism. The case also highlights the need for echocardiography in the evaluation of peripartum cardiomyopathy. We discuss the clinical presentation, diagnosis, and treatment of peripartum cardiomyopathy. PMID- 10530539 TI - Perionychial infections associated with sculptured nails. AB - Two cases of perionychial infections associated with the use of sculptured fingernails are presented. Both patients developed paroncyhia necessitating incision and drainage. One patient, a diabetic, had a concomitant subungual abscess and felon which required repeat drainage and debridement as well as intravenous antibiotics over an extended period for complete resolution. Sculptured fingernails may be risk factors for the development of digit infections through various mechanisms, and users of these cosmetic devices, especially diabetics and immunocompromised people, should be made aware of their potential for infectious complications. PMID- 10530540 TI - Methylprednisolone anaphylaxis. AB - The exacerbation of asthma is a problem frequently encountered by emergency physicians. In addition to oxygen and beta adrenergic agonists, oral and intravenous corticosteroids are increasingly being used to alleviate bronchospasm and to prevent recurrence of dyspnea. Methylprednisolone sodium succinate has been advocated as an intravenous adjunct in the treatment of asthma. We present the case of a steroid-dependent, 17-year-old male asthmatic, who experienced anaphylaxis, with respiratory arrest, within minutes of receiving intravenous methylprednisolone. Our patient rapidly responded to respiratory support and epinephrine. Methylprednisolone-induced anaphylaxis is reviewed. PMID- 10530541 TI - Patient and physician agreement on abdominal pain severity and need for opioid analgesia. AB - Whereas controversy surrounds emergency department (ED) analgesia administration to patients with undifferentiated abdominal pain, few studies have addressed the level of patient-physician agreement on abdominal pain severity and need for opioid analgesia. This prospective study was undertaken to assess concordance between emergency physicians and patients on abdominal pain severity. Study subjects were a convenience sample of 30 adults seen in an urban university affiliated tertiary care ED (annual census 65,000) who had undifferentiated abdominal pain meeting an initial severity threshold of 5 on a 10 cm visual analog scale (VAS) marked by the patient. Patients' and physicians' VAS scores, obtained in blinded fashion at presentation (t0) and at one (t1) and two (t2) hours into the ED stay, were compared with t test (VAS scores) and sign-rank (percent change in VAS scores) analyses. In addition, patients and physicians were asked at each assessment time, in blinded fashion, "Is the pain severe enough to warrant morphine?" The kappa statistic was used to characterize the degree of agreement between physician and patient assessments as to whether opioids were indicated. At t0, t1, and t2, patients' mean VAS scores (7.5, 6.7, and 5.1) were significantly (P < .05) higher than the corresponding physicians' VAS scores (5.3, 4.7, and 3.9). Though VAS scores for physicians started lower than those of patients, the percentage changes in scores from one assessment to the next were similar by Wilcoxon sign-rank testing (P > .50 for time intervals t0 - t1 and t1 - t2). Overall, patients and physicians agreed on the question of whether pain was sufficient to warrant opioids in 71 of 90 (78.9%) assessments; the corresponding kappa statistic of .57 indicated moderate agreement (P < .0001). These results, indicating that patients and physicians usually agree on whether opioids are warranted for abdominal pain, have important implications for further research on ED analgesia in this population. PMID- 10530542 TI - The spectrum of emergency admissions for thyroidectomy. AB - Emergency physicians frequently encounter patients with thyroid disease. However, it is unusual for these thyroid disorders to create acute, life-threatening situations. Critical airway compression attributable to benign or malignant thyroid enlargement may occur suddenly. Similarly, venous obstruction from thyroid tumors or severe physiological compromise from thyrotoxicosis may require urgent treatment. We reviewed a group of patients who were evaluated by emergency physicians for acute thyroid pathology and subsequently admitted for urgent thyroidectomy. Over a 7-year period, 13 patients had acute airway compressive symptoms, and 1 had acute venous compressive symptoms. Six patients had thyrotoxicosis with physiological compromise. Common airway management techniques were successfully used. Nineteen patients underwent thyroidectomy. One patient suffered a cardiopulmonary arrest before thyroidectomy could be performed. Surgical morbidity may be increased for patients undergoing thyroidectomy for urgent indications. PMID- 10530543 TI - Necrotizing fasciitis due to appendicitis. AB - Necrotizing fasciitis, although rare, is one of the more serious, life threatening complications of missed acute appendicitis. Patients who are predisposed to developing necrotizing fasciitis, regardless of the cause, are typically immunocompromised. We present a case of a 49-year-old immunocompetent female whose diagnosis of acute appendicitis was missed and who subsequently developed necrotizing fasciitis of the abdominal wall and flank. She recovered 1 month after admission due to aggressive surgical and medical therapy. PMID- 10530544 TI - Trigeminal neuralgia: a diagnostic challenge. AB - A 38-year-old white woman came to the emergency department complaining of severe, unilateral jaw pain. She had consulted her primary care physician and dentist without achieving the correct diagnosis or significant relief of her symptoms. The emergency physician made the diagnosis of trigeminal neuralgia by obtaining a history of severe paroxysmal ipsilateral facial pain activated by numerous facial stimuli. A light stimulation of the trigger point precipitated the pain. Her pain relief from carbamazepine lent further credence to the diagnosis of trigeminal neuralgia and appropriate referral to a neurosurgeon. Pain relief was ultimately achieved for the last 8 months by a neurectomy of the right infraorbital nerve. PMID- 10530545 TI - A young man with recurrent syncopes, right bundle branch block and ST segment elevation. AB - We report on the case of a 33-year-old man with recurrent syncopes appearing suddenly due to sustained monomorphic ventricular tachycardias. The electrocardiogram (ECG) showed a right bundle branch block pattern and ST segment elevation in the precordial leads V1 to V2, not explained by ischemia, electrolyte disturbances, toxic ingestion, or structural heart disease (coronary and right ventricle angiograms as well as biopsies of the right ventricle were normal). ECG image was compatible with the so-called Brugada syndrome, first described in 1992. This entity is very rare. Missed diagnosis can be disastrous because life-threatening ventricular arrhythmias often develop in patients. PMID- 10530546 TI - Prehospital ECG monitoring of chest pain patients. AB - Prehospital electrocardiograms (ECGs) have been shown to decrease the time from onset of pain to onset of treatment. They are obtained prior to treatment while the patient is likely to have his/her most intense pain. With paramedics initiating care in the field, patient assessments may be clinically different by the time the patient reaches the hospital. Thus, obtaining an ECG as early as possible during treatment could aid in the access to treatment for the few patients whose ECGs improve with prehospital care. We present a case in which the prehospital presentation was indicative of an acute myocardial infarction (MI), whereas the presentation to the hospital was not as clear-cut. The patient was taken immediately to the catheterization laboratory based on the prehospital findings and was found to have an acute MI that was treated. Laboratory findings indicated that there was a significant improvement in patient outcome based on this early treatment. This case further illustrates the role of a prehospital ECG. PMID- 10530547 TI - Cases in electrocardiography. PMID- 10530548 TI - The inadequacies of contemporary oropharyngeal suction. AB - Airway management is the highest priority in any resuscitation. Suction equipment capable of rapidly clearing the oropharynx is mandatory for airway management. Inadequate oropharyngeal suction with standard equipment may be associated with major complications in emergency airway management. We report cases that illustrate the inadequacies of standard suction equipment. Available oropharyngeal aspirators and their limitations are discussed. Recent advances in the field of oropharyngeal suction also are described. PMID- 10530549 TI - ED presentation of abdominal pain misdiagnosed as appendicitis. PMID- 10530550 TI - Solitary osseus plasmacytoma as a cause of back pain in a young patient. PMID- 10530551 TI - A case of severe crush syndrome with marked hyperkalemia: special consideration for the prevention of acute renal failure. PMID- 10530552 TI - Neonatal group B beta-hemolytic streptococcus osteomyelitis. PMID- 10530553 TI - Congenital cervical cystic hygroma causing an airway emergency. PMID- 10530554 TI - Intravenous catheterization in the ED: is there a role for topical anesthesia? PMID- 10530555 TI - Complete senorineural hearing loss as an initial presentation of leukemia. PMID- 10530556 TI - Sertoliform endometrioid carcinomas of the ovary: a clinicopathologic and immunohistochemical study of 13 cases. AB - Ovarian endometrioid carcinomas with sertoliform features (SECs) are infrequent and often misinterpreted as sex cord-stromal tumors. The clinicopathologic features and immunohistochemical expression of keratin, epithelial membrane antigen (EMA), inhibin, and estrogen and progesterone receptors were evaluated in 13 cases of SEC. The women were 41 to 89 years of age (mean, 60 yr) with abdominal enlargement secondary to a unilateral ovarian mass as the most frequent clinical presentation. One patient displayed virilization. At presentation, 10 patients were Stage I, one was Stage II and two were Stage III. The tumors were composed of compact anastomosing cords and small tubules embedded within a fibrous stroma. Nuclear features were Grade 1 or 2 in all but one tumor. Areas of conventional endometrioid carcinoma were observed in 12 cases. An adenofibromatous component comprising 5 to 60% of the lesion was present in seven cases. All 12 cases examined immunohistochemically were positive for keratin and EMA and negative for inhibin with focal, luteinized stromal cells positive for inhibin in 10 cases. Estrogen and progesterone receptors were positive in 10 and 11 cases, respectively. Follow-up on 6 of 10 patients with Stage I and the one patient with Stage II disease displayed no evidence of disease 10 to 120 months (mean, 57 mo). Progressive disease and death occurred at 12 and 72 months only in the two women with Stage III disease, one of which had an associated serous carcinoma in the contralateral ovary. Adequate sampling, a careful search for areas of conventional endometrioid carcinoma, and immunohistochemical studies (including EMA, keratin, and inhibin) are helpful in the evaluation of ovarian tumors with sex cord-stromal features. SEC should be considered a well differentiated endometrioid carcinoma despite the presence of a solid, sex cord like proliferation, with a good prognosis when confined to the ovary. PMID- 10530557 TI - Assessment of pathologic prognostic factors in breast core needle biopsies. AB - BACKGROUND: Core needle biopsies (CNB) are being used increasingly as the initial diagnostic procedure in women with breast cancer. Many clinicians are interested in obtaining as much prognostic information as possible from these limited specimens. However, the accuracy of assessing pathologic prognostic factors in core biopsy material has not been studied in detail. DESIGN: We studied CNB and subsequent excision specimens from 79 women with invasive breast cancer. Slides from CNB and excision specimens were reviewed in a blinded fashion and each was assessed for histologic type, tumor size, histologic grade, lymphatic vessel invasion (LVI), and the presence of an extensive intraductal component (EIC). RESULTS: Among the 79 cancers, there were 58 invasive ductal carcinomas, six invasive lobular carcinomas, 13 invasive carcinomas with ductal and lobular features and two tubular carcinomas, based on examination of the excision specimens. Histologic type on CNB correlated with that on excision in 64 cases (81%). Although there was a significant correlation between tumor size on CNB and excision specimens (r2 = 0.30, P = 0.01), the pathologic T stage was underestimated on CNB in 79% of cases. Furthermore, T substage was underestimated on CNB in 71% of T1 lesions. There was concordance in histologic grade between CNB and excisions in 75% of cases. Among the 20 discordant cases, the grade was higher in the excision than in the CNB in 13 cases and lower in seven. However, all discrepancies were within one grade. None of the 17 cancers with LVI in the excision specimen showed LVI on the CNB. Among 14 cases with an EIC on the excision specimen, only four (29%) were scored as having an EIC on CNB. CONCLUSION: Histologic type can be accurately determined on CNB in most cases. While there was concordance in histologic grade between CNB and excision in the majority of cases, grade was discordant in a substantial minority (25%). The ability to accurately determine tumor size (pathologic T-stage), LVI, and EIC on CNB is severely limited. PMID- 10530558 TI - Pigmented thymic carcinoids: a clinicopathological and immunohistochemical study of two cases. AB - Two cases of pigmented thymic carcinoids are presented. The patients were two men, 32 and 47 years of age. The two patients were asymptomatic and the tumor was discovered on routine chest radiographic evaluation. The tumors were treated by surgical excision in both patients. Grossly, they presented as tan-white tumors without evidence of necrosis or hemorrhage or any visible pigmentation. Histologically, the tumors were characterized by a monotonous proliferation of tumor cells arranged in a trabecular or nesting pattern. The tumor cells showed moderate amounts of pale eosinophilic cytoplasm, round to oval nuclei, and inconspicuous nucleoli. Mitotic activity varied from three to eight per 10 high power fields. In addition, both tumors showed prominent areas of intra- and extracellular dark pigment deposition. The pigment reacted positively with the Fontana-Masson stain and was negative for iron stain. Immunohistochemical studies performed in one case showed immunoreactivity of the tumor cells for chromogranin, Leu 7, and keratin. Synaptophysin and P53 immunostains were negative. Clinical follow-up was obtained in one patient who was alive and well 10 years after surgical resection. The presence of abundant melanin pigment in thymic carcinoids highlights an important pitfall for diagnosis in these tumors that should be taken into consideration in the evaluation of anterior mediastinal lesions. PMID- 10530559 TI - Histopathology of typhoid enteritis: morphologic and immunophenotypic findings. AB - Enteric fever is a systemic illness caused by Salmonella infection, with S. typhi, S. paratyphi, and S. enteritidis being the most common serotypes. Humans are the only reservoir for S. typhi, and its predilection for the ileum is due to the fact that organisms enter the body by translocation across specialized Peyer's patch epithelium and then proliferate in the mucosal macrophages. The lesions in bowel and mesenteric lymph nodes are distinctive and mimic Kikuchi Fujimoto disease and Rosai-Dorfman disease as well as infections caused by some non-salmonella bacteria. The four cases presented in this report, two culture confirmed, all exhibited ileal mucosal hypertrophy caused by a neutrophil-poor monocyte/macrophage-rich hyperplasia. Though diffuse areas were present, much of the lesional proliferation was nodular, representing macrophage infiltration and colonization by the monocytes and macrophages. Immunophenotypic studies, which showed a CD68+, lysozyme+, UCHL-1+, OPD4-, CD4-, s100- profile, were helpful in distinguishing these lesions from other processes, including Kikuchi-Fujimoto disease and Rosai-Dorfman disease. Although rare in developed countries, enteric fever should be considered in any patient with recent travel to endemic areas and in the context of illness thought to be related to contaminated foods. PMID- 10530560 TI - KIT protein expression and analysis of c-kit gene mutation in adenoid cystic carcinoma. AB - The c-kit proto-oncogene encodes a transmembrane receptor tyrosine kinase (KIT), which is expressed in several normal human tissues, especially mast cells and interstitial cells of Cajal. Expression of KIT has been noted in several types of neoplasms and gene mutation has been shown as a mechanism of c-kit oncogene activation in some tumors. Recently, a single adnexal adenoid cystic carcinoma (ACC) was reported to demonstrate KIT expression, however, examination of KIT expression or c-kit mutation in ACC of salivary glands has not been performed. We examined archival tissue samples from 30 ACC of major and minor salivary glands for KIT protein expression by immunohistochemistry with a polyclonal antibody and c-kit gene mutation by polymerase chain reaction amplification and DNA sequencing. KIT protein expression was noted in 90% of ACCs. An association between the presence of at least 50% KIT positive neoplastic cells and Grade 3 ACC or a solid growth pattern was observed (P < .05). KIT expression in normal or nonneoplastic salivary gland tissue was absent. No c-kit juxtamembrane domain (exon 11) or phosphotransferase domain (exon 17) mutations were found in any of the tumors examined. In conclusion, KIT protein expression is correlated with tumor grade of salivary ACC. However, gene mutation of exon 11 or exon 17 is not a mechanism of c-kit activation in these neoplasms. PMID- 10530561 TI - Pathologic prognostic factors in esophageal squamous cell carcinoma: a follow-up study of 74 patients with or without preoperative chemoradiation therapy. AB - One of the primary goals of pathologic examination of esophageal squamous cell carcinoma resection specimens is to provide information regarding morphologic features which can help prognosticate and guide management of affected patients. The purpose of this study was to determine the prognostic utility of a variety of histopathologic prognostic factors in patients with esophageal squamous cell carcinoma with and without preoperative chemotherapy and radiotherapy (chemrad). Multiple clinical and histologic features such as peri-tumoral lymphocytic infiltrate, Crohn's-like lymphoid reaction, degree of residual tumor, mitosis per 1000 cells, tumor differentiation, lymphatic/vascular invasion, perineural invasion, desmoplastic reaction, and tumor growth pattern were evaluated in patients with (53) and without (21) preoperative chemrad and correlated with survival (mean follow-up, 25 mo). Data were analyzed for the entire cohort and for each separate treatment group by univariate and multivariate analysis. Patients who received chemrad showed no significant survival benefit (hazard ratio = 2.5, P = .10). In the whole cohort of patients, higher pathologic stage (P = .04), poor tumor differentiation (P = .003), increased mitotic count (P = .005), perineural invasion (P = .01), lymphatic/vascular invasion (P = .002), tumor size (P = .05), and absence of a Crohn's-like lymphoid reaction (P = .05) were significantly associated with poor survival by univariate analysis. In multivariate analysis, poor tumor differentiation (P = .005), high mitotic count (P = .01), and vascular invasion (P = .03) were important prognostic features, independent of pathologic stage, for the entire cohort. In the chemrad group only, tumor size (in patients with macroscopic residual tumor) (P = .05), lymph node metastasis (P = .03), mitotic count (P = .01), and lymphatic/vascular invasion (P = .02) were significant prognostic indicators by univariate analysis. Upon multivariate analysis, only lymphatic/vascular invasion (P = .02) and mitotic rate (P = .01) were independent predictors of survival. In the nonchemrad group, only tumor differentiation was significant by both univariate (P = .008) and multivariate analysis (P = .03). The differences in pathologic prognostic factors between chemrad and nonchemrad treated cases suggests that chemrad has a significant effect on the biologic properties of these tumors. PMID- 10530562 TI - Lipid-rich follicular carcinoma of the thyroid in a patient with McCune-Albright syndrome. AB - A 41-year-old woman with McCune-Albright syndrome and a 2-cm thyroid nodule of ten years' duration presented for fine-needle aspiration, which yielded vacuolated clear cells with granular chromatin in pseudopapillary arrangement. The resected tumor showed 90% clear cells and 10% nonclear cells with capsular and vascular invasion. The cytoplasmic vacuoles in the clear cells were 3+ for oil red O stain in touch imprint cytology. Immunohistochemistry demonstrated thyroglobulin positivity in the nonclear neoplastic cells, whereas most of the clear cells were negative. Ultrastructural study demonstrated the gradual transition from protein synthesis to lipid synthesis as the neoplastic cells progressed from nonclear to clear. The study suggested that the lipid accumulation resulted from the uncontrolled fatty acid synthesis in the neoplastic cells rather than metaplasia. The karyotype of the tumor cells was normal, 46XX. Literature of lipid-rich thyroid neoplasms were reviewed. PMID- 10530563 TI - MAGE-1 and MAGE-3 tumor rejection antigens in human germ cell tumors. AB - The MAGE-1 and MAGE-3 genes are members of the melanoma antigen-encoding gene family. These genes encode for HLA-restricted tumor-associated rejection antigens recognized by cytotoxic T lymphocytes. MAGE-1 and MAGE-3 gene expression has been identified in a number of human malignancies, and MAGE-1 and MAGE-3 antigenic peptides are potential targets for tumor-specific immunotherapy. The only normal tissues known to express these genes are testicular germ cells and placental tissue. The objective of this study was to examine MAGE-1 and MAGE-3 antigens immunohistochemically in testicular germ cell tumors, including seminoma, a germ cell tumor frequently associated with a lymphoid infiltrate. Forty-three germ cell tumors (24 seminomas, six embryonal carcinomas and 13 mixed germ cell tumors), and 10 Leydig cell tumors were selected for study, and standard immunohistochemical techniques were used on formalin-fixed paraffin-embedded tissues using mouse monoclonal antibodies to MAGE-1 (clone M454) and MAGE-3 (clone 57B) antigens. MAGE-1 antigen was identified in 16.6% of seminomas. No embryonal carcinomas, yolk sac tumors, or teratomas contained MAGE-1 protein. MAGE-3 antigen was identified in 41.8% of seminomas, and this protein was not identified in embryonal carcinomas, yolk sac tumors, or teratoma. Spermatogonia and primary spermatocytes contained MAGE-1 and MAGE-3 antigen, and more mature forms, including spermatids, were weakly positive to negative. Leydig cell tumors were negative for MAGE-1 and MAGE-3. In seminoma, the presence of MAGE-1 and MAGE 3 antigens did not correlate with tumor size, tumor stage, the presence of a lymphoid infiltrate, or patient outcome. The low frequency of MAGE-specific HLA alleles in the population, the loss of the HLA class I antigens in neoplastic germ cells, and the finding that the majority of seminomas and all non seminomatous germ cell tumors lacked MAGE-1 and MAGE-3 antigenic peptides indicate that immunotherapy directed towards MAGE-1 and MAGE-3 antigen is not a likely treatment option for seminoma and nonseminomatous germ cell tumors. The significance of MAGE-1 and MAGE-3 proteins in normal spermatogonia and primary spermatocytes will require additional study. PMID- 10530564 TI - Different immunohistochemical patterns of Fhit protein expression in renal neoplasms. AB - BACKGROUND: The FHIT gene on human chromosome 3p14.2 is deleted in a variety of malignant tumors, including clear cell renal carcinomas (RCCs) resulting in a loss of expression of Fhit protein. The Fhit expression in specific subtypes of renal carcinomas has not been characterized. We have investigated the association of Fhit expression with particular subtypes of renal tumors to determine the role and specificity of this putative tumor suppressor gene in renal neoplasia. MATERIAL AND METHODS: The immunohistochemical expression of Fhit was tested in normal kidneys and in 109 renal neoplasms consisting of 51 clear cell RCCs, 26 papillary RCCs, two chromophobe carcinomas, six oncocytomas, four pelvic transitional cell carcinomas and 20 Wilms' tumors from formalin fixed and routinely processed tissue. RESULTS: Normal renal tubules expressed Fhit strongly and consistently. The majority (78%) of clear cell RCCs showed reduced or absent expression of Fhit, whereas the majority (74%) of papillary carcinomas, all chromophobe renal cell carcinomas, and oncocytomas were strongly positive. Sixty eight percent of low-grade (G1 plus G2) but only 9% of high-grade (G3 plus G4) clear cell carcinomas were Fhit negative. Wilms' tumors demonstrated focal staining in the epithelial component in 8 of 20 cases (40%). CONCLUSIONS: The loss of Fhit expression in a high percentage of clear cell RCCs with conservation of Fhit in other types of tumors supports the proposed role of FHIT alterations in the genesis of clear cell carcinomas in contrast to other types of renal epithelial tumors. FHIT expression may play a role in epithelial differentiation of nephroblastomas (Wilms' tumors). PMID- 10530566 TI - Foucar K: chronic lymphoid leukemias and lymphoproliferative disorders. Mod Pathol 12:141, 1999. PMID- 10530565 TI - Epstein-Barr virus-associated adult respiratory distress syndrome in a patient with AIDS: a case report and review. AB - BACKGROUND: Epstein-Barr virus (EBV) infection has been associated with fatal pneumonitis in immunocompetent patients. We present a case of fatal adult respiratory distress syndrome caused by EBV infection in a patient with acquired immunodeficiency syndrome (AIDS), to our knowledge the first such reported case, along with a survey of archival autopsy cases to assess baseline expression of EBV in AIDS patients. DESIGN: The case patient's autopsy material was studied exhaustively for infectious agents by culture, histochemistry, and immunohistochemistry, with negative results. Formalin-fixed paraffin-embedded lung, spleen, lymph node, and liver tissue were further studied by in situ hybridization using a probe for EBV early RNA (EBER, Kreatech). The same method was applied to lymphoid tissues from eight other archival AIDS autopsy cases. Case patient tissues were also examined by electron microscopy. RESULTS: Strikingly numerous lymphocytes were positive for EBV early RNA in the case patient's spleen, lymph nodes, and hepatic portal areas. In addition to positive lymphocytes in the lung, EBV-infected pneumocytes were also present. Electron microscopy also demonstrated viral material in lymphocytes and pneumocytes. Of the archival cases studied, only one spleen was found to have rare positive lymphocytes. CONCLUSION: Primary or reactivation EBV infection may represent a previously underreported cause of morbidity and mortality in AIDS patients. Autopsy tissues from AIDS patients do not routinely show overexpression of EBV early RNA by in situ hybridization, making this technique ideal for assessing the contribution of EBV to terminal events in these patients. PMID- 10530567 TI - Serious infections in young infants in developing countries: rationale for a multicenter study. The WHO Young Infants Study Group. PMID- 10530568 TI - Methodology for a multicenter study of serious infections in young infants in developing countries. The WHO Young Infants Study Group. PMID- 10530569 TI - Bacterial etiology of serious infections in young infants in developing countries: results of a multicenter study. The WHO Young Infants Study Group. PMID- 10530570 TI - Clinical prediction of serious bacterial infections in young infants in developing countries. The WHO Young Infants Study Group. PMID- 10530571 TI - Conclusions from the WHO multicenter study of serious infections in young infants. The WHO Young Infants Study Group. PMID- 10530572 TI - Etiology of serious infections in young Gambian infants. AB - BACKGROUND: Despite improvements in infant mortality rates in many developing countries including The Gambia, neonatal mortality remains high and many neonatal deaths are caused by infection. The study described in this paper was conducted to determine the bacterial and viral etiology of serious infections in Gambian infants younger than 91 days old. METHODS: At a first level health facility 497 infants with symptoms that could indicate serious infection were enrolled, of whom 239 with 1 or more signs of serious infection and 55 with no signs were investigated, yielding 17 cases with positive bacterial cultures of blood and/or cerebrospinal fluid. At a nearby pediatric referral hospital 198 infants were seen and 182 were investigated, yielding 35 positive bacterial cultures. RESULTS: There were 15 culture positive cases of meningitis caused by Streptococcus pneumoniae (7), Streptococcus pyogenes (2), Enterobacter cloacae (2), Escherichia coli (1), Haemophilus influenzae type b (1), Streptococcus agalactiae (1) and Salmonella spp. (1). Six of these children died. Thirty-three infants without meningitis had positive blood cultures for Staphylococcus aureus (17), S. pneumoniae (3), Salmonella spp. (5), E. coli (3), other enterobacteria (4) and S. agalactiae (1), of whom 14 died. Nasopharyngeal aspirates from 438 children were investigated for common respiratory viruses. Respiratory syncytial virus was found in 51, influenza A in 46, influenza B in 22, parainfluenza in 26 and adenovirus in 16. Respiratory syncytial virus and influenza A isolates were found most frequently toward the end of the wet season. Nasopharyngeal carriage of S. pneumoniae and H. influenzae was studied in 320 infants recruited during the first year. Of these 184 (58%) were positive for S. pneumoniae and 141 (44%) were positive for H. influenzae, 18 of which were type b. Infants with a bacterial isolate from blood or cerebrospinal fluid were more likely than the rest to die, whereas those with a viral isolate were less likely to die. CONCLUSIONS: The most important causes of serious infections in young Gambian infants are Staphylococcus aureus, S. pneumoniae and Salmonella spp. PMID- 10530573 TI - Bacterial and viral etiology of severe infection in children less than three months old in the highlands of Papua New Guinea. AB - OBJECTIVE: Determine the bacterial and viral etiology of severe infection in young Papua New Guinean infants as part of a multicenter study in four developing countries aimed at improving case management guidelines. METHODS: Between March, 1991, and April, 1993, children aged <3 months were recruited at the outpatient department of Goroka Base Hospital, Papua New Guinea (PNG). Children with pre defined inclusion criteria were enrolled, a history was taken and clinical examination was performed. Blood and urine were collected from children with signs suggestive of severe disease together with eye, umbilical and pernasal swabs as appropriate. Nasopharyngeal aspirates (NPAs) were collected from children with and without signs of severe disease for identification of viruses and Chlamydia trachomatis by direct fluorescent antibody staining. RESULTS: 3280 infants were triaged and 2168 enrolled, among whom 968 had signs suggestive of severe disease. Group A Streptococcus (Streptococcus pyogenes) and Staphylococcus aureus were the most important bacterial pathogens isolated from children < 1 month old with severe infections, and Streptococcus pneumoniae, S. pyogenes and Staphylococcus aureus were most important in older children. Of 292 eye swabs 19 (7%) grew Neisseria gonorrhoeae. Of 116 umbilical swabs 51 (44%) grew S. pyogenes and 45 (39%) grew Staphylococcus aureus. Respiratory syncytial virus was the most important viral cause of acute lower respiratory infection. CONCLUSIONS: S. pyogenes, S. pneumoniae and Staphylococcus aureus are important causes of severe infection in young children in the PNG highlands. It is necessary to improve access to clean water, promote hand-washing in the hospital and at home and investigate further the use of maternal immunization for the prevention of severe disease in young infants. PMID- 10530574 TI - Bacterial and viral etiology of serious infections in very young Filipino infants. AB - OBJECTIVE: Pneumonia, meningitis and other serious infections are leading causes of death in developing countries. As part of a multicenter study we aimed to determine the etiology of pneumonia, meningitis and other serious infections in a cohort of Filipino infants ages 90 days or younger. METHOD: During a 2-year period, 2053 infants age 90 days or younger presenting to 1 of 3 Manila community hospitals were screened; 873 had signs or symptoms suggestive of an infectious illness, and 608 were judged to have clinical features suggestive of severe infection and had laboratory workup including blood for culture and white blood cell count, nasopharyngeal aspirate for virology, cerebrospinal fluid culture when indicated and chest radiograph. Chest radiographs were read independently by 3 radiologists without knowledge of clinical findings. RESULTS: Of the 873 enrolled infants, 81 died (91%). After exclusion of presumed contaminants, positive bacterial culture from blood and/or cerebrospinal fluid was obtained in 35 infants (5.8%; 95% confidence interval 4%, 8%), 9 of whom died. The organisms responsible for meningitis were Acinetobacter spp. (4), Streptococcus pneumoniae (2), Escherichia coli (2), Enterobacter spp. (1), Pseudomonas aeruginosa (1), Haemophilus influenzae (1) and Staphylococcus aureus (1); those responsible for the other clinical diagnoses were Salmonella spp. (6), Enterobacter spp. (3), Streptococcus pyogenes (3), other Gram-negative organisms (8), S. pneumoniae (1) and Staphylococcus aureus (2). In 685 infants examined for viral causes of their illness, 223 viruses were isolated from 219 infants (32%; 95% confidence interval 28%, 36%). Enteroviruses were the most common potential pathogens identified (22% of infants studied), followed by respiratory syncytial virus (17%), rhinovirus (10%) and adenovirus (4%). Concomitant virus identification occurred in 10 of those with positive bacterial culture (29%; 95% confidence interval, 15%, 46%), with enterovirus being found in 7 of these cases. CONCLUSION: Many young Filipino infants with life-threatening illness were evaluated in this study. Thirty-five had infections attributable to bacteria, with Salmonella spp. being the most common, followed by Gram-negative organisms. Pneumococcus was an unusual cause. PMID- 10530575 TI - Etiology of pneumonia, sepsis and meningitis in infants younger than three months of age in Ethiopia. AB - METHODS: Within a multicenter study coordinated by WHO, an investigation of the etiologic agents of pneumonia, sepsis and meningitis was performed among infants younger than 3 months of age seen at the Ethio-Swedish Children's Hospital in Addis Ababa for a period of 2 years. Of the 816 infants enrolled 405 had clinical indications for investigation. RESULTS: There were a total of 41 isolates from blood cultures from 40 infants. The study showed that the traditionally known acute respiratory infection pathogen Streptococcus pneumoniae was most common in this extended neonatal age group, found in 10 of 41 blood isolates. Streptococcus pyogenes was a common pathogen in this setting (9 of 41 blood isolates), whereas Salmonella group B was found in 5 of 41 isolates. Streptococcus agalactiae, which is a common pathogen in developed countries, was absent. A study of the susceptibility pattern of these organisms suggests that a combination of ampicillin with an aminoglycoside is adequate for initial treatment of these serious bacterial infections, but the combination is not optimal for the treatment of Salmonella infections. Among 202 infants on whom immunofluorescent antibody studies for viruses were performed based on nasopharyngeal aspirates, respiratory syncytial virus was found in 57 (28%) infants, and Chlamydia trachomatis was isolated in 32 (15.8%) of 203 infants. PMID- 10530576 TI - High rates of Chlamydia trachomatis infections in young Papua New Guinean infants. AB - OBJECTIVE: Determine the importance of Chlamydia trachomatis in the etiology of severe infection in young Papua New Guinean infants. METHODS: Between March, 1991, and April, 1993, children <3 months old were recruited as outpatients at Goroka Base Hospital, Papua New Guinea, as part of a multicenter study in four developing countries. Children with predefined inclusion criteria were enrolled. C. trachomatis was identified by direct fluorescent antibody staining in nasopharyngeal aspirates (NPAs) collected from children with and without signs of severe disease and eye swabs from children with and without conjunctivitis. Two to three radiologists read chest radiographs without knowledge of clinical and laboratory findings. RESULTS: Of 3280 outpatients seen 2168 enrolled, 955 NPAs were tested for C. trachomatis and 549 chest radiographs were read. Of 210 eye swabs from children with conjunctivitis 57% were positive for C. trachomatis compared with 8% from 167 children with no conjunctivitis. The prevalence of C. trachomatis in NPAs was 9% in asymptomatic children and 18 and 33% in children with nonsevere or severe pneumonia, respectively. C. trachomatis in NPAs was strongly associated with clinically severe pneumonia [odds ratio (OR), 2.91], reduced arterial oxygen saturation (OR 2.58) and radiographic evidence of pneumonia (OR 5.84) and was also associated with pneumococcal bacteremia (OR 3.48). CONCLUSIONS: In Papua New Guinea Chlamydia must be considered as a cause when treating pneumonia in infants, and effective treatment and prevention of sexually transmitted diseases are urgently needed for a number of reasons, including the need to curb high rates of chlamydial infection in women and infants. PMID- 10530577 TI - Only the pneumococcus... PMID- 10530578 TI - Development of local guidelines for prevention of respiratory syncytial viral infections. PMID- 10530579 TI - Short course therapy with cefuroxime axetil for acute otitis media: results of a randomized multicenter comparison with amoxicillin/clavulanate. AB - BACKGROUND: Otitis media is a common infection of childhood. Increasing antibiotic resistance rates among the principal causative pathogens, Streptococcus pneumoniae and Haemophilus influenzae, are associated with failure of first line agents. OBJECTIVE: This open, randomized, multicenter study compared the clinical efficacy of a short 5-day course of cefuroxime axetil (CAE) suspension with that of amoxicillin/clavulanate (A/CA) suspension for 8 or 10 days. METHODS: Children age 6 to 36 months with acute otitis media with effusion, diagnosed by tympanocentesis and microbiologic culture, were randomized to receive CAE (30 mg/kg/day in two divided doses for 5 days) or A/CA 40 mg/kg/day in three divided doses for 10 days (A/CA-10). In French centers A/CA was given at 80 mg/kg/day in three divided doses for 8 days (A/CA-8). Patients were assessed 1 to 4 days after completing the course (posttreatment) and followed up at 21 to 28 days after completing the course. RESULTS: Of the 716 patients randomized, 252 were treated with CAE, 255 with A/CA-10 and 209 with A/CA-8. In the clinically evaluable population, the proportions of patients with clinical cure at posttreatment were 175 of 203 (86%), 181 of 205 (88%) and 145 of 164 (88%) in the CAE, A/CA-10 and A/CA-8 groups, respectively, demonstrating equivalence among the three treatments. For patients <18 months old, clinical cures were 111 of 134 (83%), 116 of 131 (89%) and 83 of 99 (84%) in the CAE, A/CA-10 and A/CA-8 groups, respectively; equivalence was also demonstrated. At follow-up, 130 of 175 (74%) CAE, 121 of 172 (70%) A/CA-10, and 112 of 142 (79%) A/CA-8 had maintained cure. A total of 837 pretreatment pathogens were isolated from middle ear fluid in 73% (522 of 716) patients, the majority of isolates were S. pneumoniae (30%) and H. influenzae (27%). The most common adverse events were gastrointestinal, the incidence of drug-related diarrhea being higher in the A/CA-10 group (18%) than in either the CAE or A/CA-8 groups (10%). CONCLUSIONS: A 5-day course of CAE, given twice daily, was shown to be equivalent to the two regimens of A/CA for treatment of acute otitis media with effusion in children. PMID- 10530580 TI - High incidence of Alloiococcus otitis in otitis media with effusion. AB - BACKGROUND: The etiology of otitis media with effusion (OME) is unclear. Although the majority of effusions show inflammation, culture methods yield positive results for bacteria in only 20 to 30% of cases. METHODS: The polymerase chain reaction was used for detection of three upper respiratory tract pathogens, Haemophilus influenzae, Moraxella catarrhalis and Streptococcus pneumoniae, and a fairly recently described bacterium, Alloiococcus otitis (A. otitidis), that is solely found in OME. The study included 67 middle ear effusions that were collected from 48 pediatric OME patients during ventilation tube placement. RESULTS: PCR tested positive for 57 (85.1%) of the middle ear effusions. Thirty one (46.3%) A. otitis-, 12 (17.9%) H. influenzae-, 25 (37.3%) M. catarrhalis- and 14 (20.9%) S. pneumoniae-positive effusions were obtained. All four study organisms showed similar distribution in effusions of various duration (P = 0.72) and in different effusion types (P = 0.59). Only the proportion of M. catarrhalis positive effusions was lowered by recent antimicrobial therapy (P < 0.05). Although the study organisms had equal distributions among singly and multiply positive specimens (P = 0.90), A. otitis was detected significantly more often with one of the three other species (15 of 19, 78.9%) than the other species with each other (4 of 19, 21.1%, P < 0.001). CONCLUSIONS: The findings suggest a bacterial etiology for OME. Association of A. otitis with the three other species implies that this organism might have the capability of augmenting bacterial colonization in the middle ear. PMID- 10530581 TI - Variable morbidity of respiratory syncytial virus infection in patients with underlying lung disease: a review of the PICNIC RSV database. Pediatric Investigators Collaborative Network on Infections in Canada. AB - OBJECTIVE: We wished to compare outcomes of respiratory syncytial virus (RSV) infection in children with bronchopulmonary dysplasia (BPD) with those with other pulmonary disorders: cystic fibrosis, recurrent aspiration pneumonitis, pulmonary malformation, neurogenic disorders interfering with pulmonary toilet, and tracheoesophageal fistula. METHODS: Children with RSV infection hospitalized at seven Canadian pediatric tertiary care hospitals in 1993 through 1994 and 9 hospitals in 1994 through 1995 were enrolled and prospectively followed. This study is a secondary analysis of data from this prospective cohort. RESULTS: Of the 1516 patients enrolled the outcomes of 159 with preexisting lung disorders before RSV lower respiratory tract infection constitute this report. There were no significant differences among the 7 groups (BPD, cystic fibrosis, recurrent aspiration pneumonitis, pulmonary malformation, neurogenic disorders interfering with pulmonary toilet, tracheoesophageal fistula, other) for the morbidity measures: duration of hospitalization, intensive care unit (ICU) admission, duration of ICU stay, mechanical ventilation and duration of mechanical ventilation. Patients using home oxygen were more likely to be admitted to the ICU than those who had never or previously used home oxygen (current 57.1%, past 23.8%, never 33.3%, P = 0.03). CONCLUSIONS: Children with other underlying diseases have morbidity similar to those with BPD. Prophylactic interventions against RSV should also be studied in these groups. PMID- 10530582 TI - Clinical significance of enteroviruses in serious summer febrile illnesses of children. AB - BACKGROUND: Enteroviruses are common causes of aseptic meningitis and nonspecific febrile illnesses in young children. During the summer-fall months, enterovirus infected children are frequently evaluated in emergency room settings to rule out bacterial sepsis and/or meningitis. OBJECTIVES: We sought to determine the clinical significance of enterovirus infections in children evaluated for serious febrile illnesses in pediatric emergency rooms during the summer-fall season. METHODS: Children admitted to emergency rooms at four university teaching hospitals during a single summer-fall season who required blood culture and/or lumbar puncture to rule out bacterial sepsis/meningitis were prospectively studied. An extensive questionnaire was administered, and specimens of cerebrospinal fluid, serum, urine and throat were tested for enteroviruses by viral culture and PCR. Patients were followed to determine the duration, management and outcome of their illnesses. RESULTS: Of 203 patients studied 173 had no apparent explanation for their illness (e.g. bacterial sepsis, bacterial urinary tract infection, etc.). Of those 173 patients 79 (46%) were infected with enteroviruses, including 33 of 47 (70%) patients with aseptic meningitis, 13 of 25 (52%) patients with nonspecific febrile episodes and 33 of 101 (33%) patients with fever and focal findings (P < 0.0001 for aseptic meningitis vs. fever and focal findings; P = 0.0001 for aseptic meningitis vs. combined nonspecific febrile episodes and fever/focal patients). Among 119 hospitalized patients 65 (55%) were enterovirus-infected. Children < or =90 days of age were more likely to be enterovirus-infected (66 of 122; 54%) than children older than 90 days (13 of 51; 25%) (P = 0.0001). Enterovirus-infected children were more likely to be hospitalized as a result of the current emergency room visit (65 of 79 vs. 54 of 94; P = 0.0005) and were more likely to have had an additional hospitalization for the same illness (10 of 79 vs. 1 of 94; P = 0.003). Enterovirus-infected patients also had a shorter period from illness onset to presentation. Enterovirus-infected children were indistinguishable from those without enterovirus infection in their symptoms at onset, signs at presentation and total duration of illness (>7 days in both groups). Enterovirus-infected children were almost all treated with antibiotics (78 of 79; 99%), with 74 of 79 (94%) receiving parenteral antibiotics for a mean of 3.6 days. CONCLUSIONS: During the summer-fall months, 39% (79 of 203) of children for whom blood cultures and/or lumbar punctures were performed for suspected bacterial infection had enterovirus infection identified as the only explanation for their illness. Of those patients with no alternative diagnosis, enterovirus infection was confirmed in 46% (79 of 179). The majority of those patients requiring hospitalization were infected with enteroviruses. The use of PCR increases the number of children for whom a specific etiology of illness can be determined and may in the future reduce the hospitalization and use of unnecessary antibiotics in patients with enterovirus infections. PMID- 10530583 TI - Comparison of procalcitonin with C-reactive protein, interleukin 6 and interferon alpha for differentiation of bacterial vs. viral infections. AB - BACKGROUND: Procalcitonin (PCT) concentration increases in bacterial infections but remains low in viral infections and inflammatory diseases. The change is rapid and the molecule is stable, making it a potentially useful marker for distinguishing between bacterial and viral infections. METHODS: PCT concentration was determined with an immunoluminometric assay on plasma collected at admission in 360 infants and children hospitalized for bacterial or viral infection. It was compared with C-reactive protein (CRP), interleukin 6 and interferon-alpha measured on the same sample. RESULTS: The mean PCT concentration was 46 microg/l (median, 17.8) in 46 children with septicemia or bacterial meningitis. PCT concentration was > 1 microg/l in 44 of 46 in this group and in 59 of 78 children with a localized bacterial infection who had a negative blood culture (sensitivity, 83%). PCT concentration was > 1 microg/l in 16 of 236 children with a viral infection (specificity, 93%). PCT concentration was low in 9 of 10 patients with inflammatory disease and fever. A CRP value > or =20 mg/l was observed in 61 of 236 patients (26%) with viral infection and in 105 of 124 patients (86%) with bacterial infection. IL-6 was > 100 pg/ml in 14% of patients infected with virus and in 53% with bacteria. A secretion of interferon-alpha was found in serum in 77% of viral infected patients and in 8.6% of bacterial infected patients. CONCLUSIONS: In this study a PCT value of 1 microg/l or greater had better specificity, sensitivity and predictive value than CRP, interleukin 6 and interferon-alpha in children for distinguishing between viral and bacterial infections. PCT values are higher in invasive bacterial infections, but the cutoff value of 1 microg/l indicates the severity of the disease in localized bacterial infection and helps to decide antibiotic treatment in emergency room. PCT may be useful in an emergency room for differentiation of bacterial vs. viral infections in children and for making decisions about antibiotic treatments. PMID- 10530584 TI - Rifapentine pharmacokinetics in adolescents. AB - OBJECTIVE: Determination of rifapentine pharmacokinetics in healthy adolescent children. DESIGN: Prospective Phase II clinical trial. SETTING: Clinical research center within a university children's hospital. PATIENTS: Twelve subjects ranging in age from 12 to 15 years, male and female. INTERVENTIONS: A single oral dose of rifapentine was administered to healthy adolescent volunteers, 450 mg if <45 kg or 600 mg if > or =45 kg. Blood was collected at serial intervals (0, 2, 3, 4, 5, 6, 8, 12, 18, 24, 48 and 72 h postdose). Subjects were observed for adverse effects during the period of study. MEASUREMENTS: High pressure liquid chromatography was used to measure the plasma concentration of rifapentine and 25 desacetyl rifapentine in each blood sample. For each subject a plot of mean plasma concentration vs. time data for rifapentine and its metabolite (i.e. 25 desacetyl rifapentine) were created. Subsequently model-independent methods were used to determine the pharmacokinetic profiles for each subject. RESULTS: All subjects tolerated rifapentine without adverse effects. The 2-h postdose plasma concentrations of rifapentine (6.59 to 9.05 microg/ml) and 25-desacetyl rifapentine (0.57 to 2.64 microg/ml) far exceeded the MIC of Mycobacterium tuberculosis to rifapentine (approximately 0.12 microg/ml). The combination of a high Cmax (rifapentine, 9.95 to 18.63 microg/ml; 25-desacetyl rifapentine, 3.73 to 7.46 microg/ml) and lengthy terminal elimination phase t1/2 (rifapentine, 10 to 23 h; 25-desacetyl rifapentine, 14 to 35 h) resulted in potentially effective plasma concentrations of both compounds that persisted for at least 48 h in most subjects. CONCLUSIONS: A well-tolerated oral rifapentine dose produced rapid and sustained plasma drug concentrations in adolescents that should effectively treat infections caused by M. tuberculosis. Rifapentine pharmacokinetics appears to be similar in adolescent and adult populations. PMID- 10530585 TI - Citrobacter urinary tract infections in children. AB - BACKGROUND: Citrobacter species have been described as the etiologic agents in cases of bacteremia, meningitis, diarrhea and brain abscess, but little is known of their role as a cause of urinary tract infections in children. The purpose of this study was to define the role of Citrobacter species in pediatric urinary tract infections. METHODS: The project consisted of a retrospective chart review of microbiologic and medical records of patients younger than 18 years of age with urine cultures positive for Citrobacter species during a 3-year period. RESULTS: Thirty-four patients with 37 infections were included in the review. The average patient age was 6.9 years (range, 1 month to 18 years) and 71% were female. Fifty-six percent of the patients had urinary tract/renal anomalies or neurologic impairment and 26% represented nosocomial infections. Thirty-seven percent of patients were asymptomatic at the time of diagnosis, whereas 63% complained of at least one of the following findings: gastrointestinal symptoms; dysuria; fever; incontinence; penile/vaginal discharge; frequency; flank pain; and hematuria. Twenty-six of the isolates were Citrobacter freundii and 11 were Citrobacter koseri. Blood cultures were obtained in 9 patients and all were negative for Citrobacter isolates. CONCLUSIONS: Although it is uncommon Citrobacter can cause urinary tract infections in the pediatric population, which occur more frequently in children with underlying medical conditions. It appears that treatment similar to that of other gram-negative enteric organisms is the most prudent approach to these children until more information can be gathered. PMID- 10530586 TI - Effect of the Factor V Leiden mutation on the severity of meningococcal disease. AB - BACKGROUND: One of the most serious complications of meningococcal disease is the syndrome of purpura fulminans, which is characterized by intravascular thrombosis and hemorrhagic infarction of skin, limbs and digits. The reasons why some patients with meningococcal disease develop purpura fulminans while others have minimal thrombotic and skin involvement despite having profound septic shock are not yet understood. The Factor V Leiden mutation (FV(L)) is associated with thrombotic events, and we hypothesized that children carrying FV(L) who develop meningococcal disease may be at increased risk of purpura fulminans. METHODS: We determined the FV(L) genotype by PCR and restriction enzyme digestion (Mnl1) in 259 children with meningococcal disease and 80 healthy controls. In addition 79 parents of children with fatal meningococcal disease were studied. RESULTS: There was no significant increase in the frequency of FV(L) in patients with meningococcal disease (10%) as compared with healthy controls (9%) or with the parents of children who died of meningococcal disease (12%). Although the mortality was not increased in patients heterozygous for FV(L), they had increased complications of purpura fulminans, as assessed by requirement for skin grafting, referral to plastic surgeon and/or amputation. Among survivors 5 of 24 (21%) of those heterozygous for FV(L) had complications, compared with 14 of 233 (7%) who were wild type [P < 0.03; relative risk, 3.1 (95% confidence intervals, 1.2 to 7.9)]. CONCLUSIONS: FV(L) exacerbates purpura fulminans in meningococcal disease but does not have a significant effect on mortality. PMID- 10530587 TI - Chloramphenicol pharmacokinetics in infants less than three months of age in the Philippines and The Gambia. AB - BACKGROUND: The broad antimicrobial spectrum and affordable price of chloramphenicol make it an attractive first line treatment option for children with severe illnesses in developing countries. Little is known, however, about its pharmacokinetics in young infants in these settings. METHODS: We studied infants younger than 3 months of age hospitalized in Manila, Philippines and The Gambia with possible severe bacterial infections likely to benefit from treatment with chloramphenicol. Infants in the first week of life received intramuscular doses of 25 mg/kg chloramphenicol once daily, twice daily in the second through fourth week of life and three times daily from 5 to 12 weeks of age. Blood samples were taken at 0.5, 1, 2 and 3 h after the first dose, 1 h before the second dose and before the repetition doses on subsequent days. In the Philippines a second group of infants was treated with oral chloramphenicol according to the same dosage schedule. RESULTS: Thirty-eight infants received intramuscular chloramphenicol, and 20 received oral drug. Intramuscular administration resulted in therapeutic concentrations (10 to 25 microg/ml) in 73 to 86% of children in each of the three age groups in the first 6 h and in 50 to 80% on Days 2 and 3. Between 33 and 38% of children had potentially toxic values on Days 2 and 3. In contrast, after oral administration, only about one-half of the children reached therapeutic values in serum at any time up to Day 3 after start of treatment. CONCLUSIONS: Intramuscular chloramphenicol can be used as a second line drug for the treatment of severe infections in infants younger than 90 days of age, where third generation cephalosporins are not available. It quickly achieves therapeutic values in a high proportion of children. However, severe infections should not be treated with oral chloramphenicol in this age group, because therapeutic serum concentrations were inconsistently achieved. PMID- 10530588 TI - Meningitis in pediatric cancer patients: a review of forty cases from a single institution. AB - BACKGROUND: Although the clinical features of bacterial meningitis in adult cancer patients and in healthy children have been described, no previous large series has described the clinical features of meningitis in pediatric cancer patients. We performed a retrospective review of bacterial or fungal meningitis in pediatric cancer patients to determine its clinical presentation, microbiology and outcome. METHOD: We reviewed the medical records of all patients younger than 18 years old with a diagnoses of any malignancy and bacterial or fungal meningitis at Children's Hospital and Regional Medical Center in Seattle, WA, from January, 1981, to June, 1998. RESULTS: During the study period there were 40 cases of bacterial or fungal meningitis in 36 pediatric cancer patients. Most patients (65%) had recent neurosurgery, a central nervous system device or cerebrospinal fluid leak. Neutropenia was present in 30% of patients. Fever and altered mental status were the most consistent signs at presentation. In addition at least one additional symptom or sign of meningitis (headache, neck pain or rigidity, seizures or photophobia) was present in 77% of cases. Staphylococcus aureus and Streptococcus pneumoniae were the most common microbiologic isolates. The five patients with fatal outcome were neutropenic. Neutropenia and seizures within 2 days of presentation were associated with long neurologic sequelae. CONCLUSIONS: Meningitis in pediatric cancer patients was associated with significant morbidity and mortality. Pediatric cancer patients with meningitis had clinical features and microbiology distinctly different from those of adult cancer patients and normal children with meningitis. PMID- 10530589 TI - Foodborne diseases: Shiga toxin producing E. coli (STEC). PMID- 10530590 TI - Foodborne diseases: seafood. PMID- 10530591 TI - Foodborne diseases: fruits and vegetables. PMID- 10530592 TI - Lyme disease. PMID- 10530593 TI - The promise of immunoprophylaxis for prevention of acute otitis media. PMID- 10530594 TI - Ulcerative lesion of the nasal bridge in a five-month-old infant. PMID- 10530595 TI - Measles antibody in pregnant Argentinian women relative to vaccine-induced immunity and natural infection. PMID- 10530596 TI - Polymerase chain reaction assay for detecting Chlamydia pneumoniae in middle ear fluid of children with otitis media with effusion. PMID- 10530597 TI - Primary sternal osteomyelitis in children with sickle cell disease. PMID- 10530598 TI - Streptococcus pneumoniae causes otitis media with higher fever and more redness of tympanic membranes than Haemophilus influenzae or Moraxella catarrhalis. PMID- 10530599 TI - Observations on the impact of breast-feeding and of intestinal helminthiasis on a rapid agglutination assay for fecal lactoferrin in Nicaraguan children with diarrhea. PMID- 10530600 TI - Occupational magnetic field exposure and site-specific cancer incidence: a Swedish cohort study. AB - OBJECTIVE: Based on 1,596,959 men and 806,278 women, site-specific cancer incidence during 1971 through 1984 was analyzed in relation to occupational magnetic field exposure. The objective was to explore potential associations for cancer diseases beyond those extensively studied before (leukemia and brain tumors). METHODS: Exposure was assessed from Census information on occupations that were linked to a job exposure matrix based on measurements. In a basic analysis, three levels of exposure were used. In addition, subjects with a more definite low exposure were compared with an aggregate of occupations with more definite exposures. RESULTS: Observed associations were weak and there were no evident exposure-response relationships. For all cancer, an approximate 10% increase in risk was seen in the medium and high exposure groups. Several types of cancer were associated with exposure among men, including cancer of the colon, biliary passages and liver, larynx and lung, testis, kidney, urinary organs, malignant melanoma, non-melanoma skin cancer, astrocytoma III-IV. For women, associations were seen for cancer of the lung, breast, corpus uteri, malignant melanoma and chronic lymphocytic leukemia. CONCLUSIONS: In the analysis of occupations with a more definite exposure, the most notable finding for men was an increased risk of testicular cancer in young workers, and for women a clear association emerged for cancer of the corpus uteri. The outcome suggests an interaction with the endocrine/immune system. PMID- 10530601 TI - Evaluation of breast cancer incidence: is the increase due entirely to mammographic screening? AB - OBJECTIVES: To examine the trends in the incidence rates of breast cancer in a population with mammographic screening and in the unscreened women within that population. METHODS: Data consisted of incident cases of breast cancer notified to the Victorian Cancer Registry in Victoria, Australia, between 1988 and 1996 and cases detected in the population-based BreastScreen Program. These data were grouped by age (25-39, 40-49, 50-59, 60-69 and > or = 70 years of age) and size of tumor (< or =10 mm, > 10-< or =15 mm, and > 15 mm). Poisson regression modeling was used to examine trends by age, tumor size, calendar year and availability of screening. RESULTS: The incidence rate of breast cancer in the total population increased between 1988 and 1996. The greatest increase was seen after 1993 when population-based screening became available. In unscreened women, modeling demonstrated a statistically significant (p < 0.01) 1.5% annual increase in the incidence rate. The annual increase in this rate differed by size of tumor and was approximately 8% (p < 0.01) for small tumors (< or = 10 mm) but not significant for tumors > 10 mm. The greatest increase was in small tumors for women > or = 50 years of age. CONCLUSION: The incidence of breast cancer has increased since population-based mammographic screening was introduced in 1994. The rate in unscreened women also showed a significant increase. This was greatest in small tumors for women > or = 50 years of age. Whether this will translate into an increase in mortality is uncertain and long-term monitoring is required to determine if cohort and period effects impact on the underlying incidence of breast cancer in Victoria. PMID- 10530602 TI - The association between cigarette smoking and low-grade cervical abnormalities in reproductive-age women. AB - OBJECTIVE: To evaluate the association between smoking and the occurrence of low grade cervical cytological abnormalities. METHODS: We conducted a population based cross-sectional study of smoking and other exposures in reproductive-age women with normal and abnormal cytology results (Class 1-4 Pap tests). Participants (n = 2,448) were enrollees of the Group Health Cooperative of Puget Sound, a health maintenance organization (HMO) in Washington state, USA. Non pregnant women were selected monthly from the HMO's cytology database during 1995 6, with over-sampling of women with low-grade abnormalities. All participants completed a structured telephone-administered survey. RESULTS: Of the 2,448 participants, 19% (n = 465) had Class 2 Pap results, and 5% (n = 117) had Class 3 4 results. Forty percent of the sample (n = 975) reported ever smoking. Women reporting current/recent smoking (n = 514, 21%) had an increased likelihood of cervical abnormalities (adjusted odds ratio (OR) 1.4, 95% confidence interval (CI 1.1-1.8). Women who had never smoked but who reported recent passive smoking exposure also had a greater likelihood of abnormal test results (OR 1.4, 95% CI 1.0-2.0). Prior smoking was not associated with cytology status. CONCLUSIONS: Our results, examining low-grade cervical abnormalities, are compatible with those from studies of more severe cervical lesions, lending added support to the hypothesis that smoking predisposes to development of a spectrum of cervical abnormalities. Thus, even cytologic screening visits represent an opportunity to counsel women smokers about their health risks, particularly the more proximal risks of cervical abnormalities and cancer. PMID- 10530603 TI - Cancer mortality rates in Menofeia, Egypt: comparison with US mortality rates. PMID- 10530604 TI - Cancer mortality in Menofeia, Egypt: comparison with US mortality rates. AB - OBJECTIVES: In developing countries where cancer registries are unavailable, mortality statistics from death certification may be a practical source of cancer statistics. We aimed at describing the cancer mortality in Egypt and comparing it to that in the US. METHODS: We used the mandatory and routinely available mortality records of Menofeia province in the Nile Delta region of Egypt, which is typical of the rest of Egypt. We determined cancer mortality rates, and compared them with the Surveillance, Epidemiology, and End Results (SEER) mortality rates of the US. RESULTS: Bladder and liver cancers were the two most common causes of cancer mortality in Menofeia, Egypt. When adjusted for age the Egyptian rates were much higher than the US rates (9.5/100,000 and 8.4/100,000 for bladder and liver cancer, respectively, compared with 2.3/100,000 and 2.5/100,000 for the same cancers from SEER data). We also observed that age specific rates for early-onset colorectal cancer under age 40 and premenopausal breast cancer were higher in Egypt than in the US. CONCLUSION: This study confirms our earlier observations about the higher proportion of early-onset colorectal cancer in Egypt, and opens the door for future studies to investigate familial clustering of cancer in Egypt. PMID- 10530605 TI - Risk factors for prostate cancer: results from the Canadian National Enhanced Cancer Surveillance System. The Canadian Cancer Registries Epidemiology Research Group. AB - OBJECTIVES: To evaluate the relationship between prostate cancer and several potential lifestyle risk factors. METHODS: We analyzed data obtained from a population-based case-control study conducted in eight Canadian provinces. Risk estimates were generated by applying multivariate logistic regression methods to 1623 histologically confirmed prostate cancer cases and 1623 male controls aged 50-74. RESULTS: Cases were more likely to have a first-degree relative with a history of cancer, particularly prostate cancer (OR = 3.1, 95% CI = 1.8-5.4). Reduced risks of prostate cancer were observed among those of Indian descent (OR = 0.2, 95% CI = 0.1-0.5) or any Asian descent (OR = 0.3, 95% CI = 0.2-0.6) relative to those of western European descent. Total fat consumption, tomato and energy intake, were not associated with prostate cancer. The risk of prostate cancer was inversely related to the number of cigarettes smoked daily (p = 0.06) and cigarette pack-years (p < 0.01), while no association was observed between the total number of smoking years or the number of years since smoking cessation. Anthropometric measures and moderate and strenuous levels of leisure time physical activity were not strongly related to prostate cancer. In contrast, strenuous occupational activities at younger ages appeared protective. CONCLUSIONS: Our analyses are limited by the absence of data related to tumor severity and screening history. Further studies are needed to investigate the relationship between behavioral risk factors and prostate cancer screening practices. PMID- 10530606 TI - Alcohol dehydrogenase 3 genotype modification of the association of alcohol consumption with breast cancer risk. AB - OBJECTIVES: Because alcohol dehydrogenase 3 (ADH3) is rate-limiting in alcohol oxidation and is polymorphic, we examined ADH3 genotype in relation to alcohol intake and breast cancer risk. METHODS: We conducted a case-control study among Caucasian women aged 40-85 with incident, pathologically confirmed breast cancer and controls, frequency-matched on age and county. Queries included alcohol intake in the past 20 years. Genomic DNA was genotyped for the exon VIII ADH polymorphism by PCR followed by restriction enzyme digestion. Computation of odds ratios (OR) and 95% confidence intervals (CI) was by unconditional logistic regression. RESULTS: We found increased risk among pre- (OR 2.3, 95%, CI 1.2-4.3) but not postmenopausal women (OR 1.1, 95% CI 0.7-1.7) associated with ADH3(1-1) compared to ADH3(1-2) and ADH3(2-2) genotypes. Risk was increased for premenopausal women with the ADH3(1-1) genotype and alcohol intake above the median (OR 3.6, 95% CI 1.5-8.8) compared to lighter drinkers with the ADH3(2-2) or ADH3(1-2) genotypes. ORs were close to null for premenopausal women in other drinking and genotype groups and for postmenopausal women categorized by genotype and alcohol consumption. CONCLUSION: Among premenopausal women there may be a group more genetically susceptible to an alcohol consumption effect on breast cancer risk. PMID- 10530608 TI - Diet and risk of colorectal cancer in a cohort of Finnish men. AB - OBJECTIVES: Based on previous epidemiological studies, high fat and meat consumption may increase and fiber, calcium, and vegetable consumption may decrease the risk of colorectal cancer. We sought to address these hypotheses in a male Finnish cohort. METHODS: We analyzed data from the Alpha-Tocopherol, Beta Carotene Cancer Prevention Study (ATBC Study) where 27, 111 male smokers completed a validated dietary questionnaire at baseline. After an average of 8 years of follow-up, we documented 185 cases of colorectal cancer. The analyses were carried out using the Cox proportional hazards model. RESULTS: The relative risk (RR) for men in the highest quartile of calcium intake compared with men in the lowest quartile was 0.6 (95% CI 0.4-0.9, p for trend 0.04). Likewise, the intake of milk protein and the consumption of milk products was inversely associated with risk of colorectal cancer. However, intake of dietary fiber was not associated with risk, nor was fat intake. Consumption of meat or different types of meat, and fried meat, fruits or vegetables were not associated with risk. CONCLUSIONS: In this cohort of men consuming a diet high in fat, meat, and fiber and low in vegetables, high calcium intake was associated with lowered risk of colorectal cancer. PMID- 10530607 TI - Glycosylated hemoglobin and risk of colorectal cancer and adenoma (United States). AB - OBJECTIVES: The consistently observed epidemiologic associations of obesity and physical activity with colorectal cancer and precursor adenoma risk suggest that insulin and glucose control may be contributory. We evaluated the association of glycosylated hemoglobin (HbA1c), a clinical indicator of average glycemia over the previous 2 months, and possibly, indirectly, a marker of average blood insulin level, with colorectal carcinogenesis. METHODS: Among women in the Nurses' Health Study, who provided blood in 1989-90 and were diagnosed subsequently in 1989-94, we included 79 colorectal cancer cases and 156 matched controls, and 201 distal colorectal adenoma cases and 201 matched controls. HbA1c concentrations in red blood cells were determined blindly by turbidometric immunoinhibition. Odds ratios (OR) and 95% confidence intervals (CI) were estimated from conditional logistic regression models. RESULTS: HbA1c level did not significantly differ between colorectal cancer cases (median 5.5%) and controls (5.5%, p = 0.5), although a small difference between adenoma cases (5.6%) and controls (5.5%, p = 0.06) was noted. Compared to the lowest tertile of HbA1c (median 5.2%), women in the middle (median 5.5%, OR = 1.2, CI = 0.6-2.5) and upper (5.8%, OR = 1.2, CI = 0.6-2.7) tertiles were not at an increased risk for colorectal cancer. A modestly elevated risk of distal colorectal adenoma in the upper (median 5.8%, OR = 1.4, CI = 0.9-2.3) versus lower (median 5.3%) tertile could not be excluded. These associations were not appreciably altered after adjusting for known and suspected colorectal cancer risk factors. CONCLUSION: Over the range of levels observed in this relatively small sample of middle-aged women, prediagnostic HbA1c does not clearly predict colorectal cancer and adenoma risk. PMID- 10530609 TI - Pregnancy recency and risk of ovarian cancer. AB - OBJECTIVE: A recent analysis suggested that ovarian cancer risk increased with time since last birth, possibly because of some aspect of pregnancy that affects the clearance of cells that have undergone malignant transformation. We analyzed data from four case-control studies pertaining to ovarian cancer risk in relation to age at first pregnancy, age at last pregnancy, and years since last pregnancy: 628 cases and 3432 neighborhood or population controls, ages 18-79, were included. METHODS: We used logistic regression to analyze associations between ovarian cancer risk, controlling for study, age (at diagnosis or corresponding reference age for controls), race, parity, oral contraceptive use, tubal ligation, family history of ovarian or breast cancer, and excluding women with a history of infertility. RESULTS: An early age at first pregnancy was associated with an increased risk of ovarian cancer (odds ratio 1.4, 95% confidence interval (1.1-1.8) for ages < or =19 compared to > or =25). Years since last pregnancy was also associated with increased ovarian cancer risk, with odds ratios of 1.4, 1.4, 1.8, and 2.1 for 10-14, 15-19, 20-24, and > or =25 years compared to 0-9 years (trend test p = 0.004), respectively. CONCLUSION: These observations support the results from the previous study, and raise additional questions about the role of pregnancy in the etiology of ovarian cancer. PMID- 10530610 TI - Favorable trends in melanoma incidence: can we claim credit? PMID- 10530611 TI - Trends by anatomic site in the incidence of cutaneous malignant melanoma in Canada, 1969-93. AB - Trends in melanoma incidence by anatomic site were examined in Canada where ascertainment of cancer has been of a high standard. The analysis included 41,239 malignant melanomas registered between 1969 and 1993 and used an age, period and cohort modeling approach. The estimated annual increase was 4.8% for males and 3.1% for females but slowed appreciably in the later years. The lifetime risk of melanoma appeared to have peaked with women born about 1934 and men born about 1944. The age-standardized rates have now stabilized for women and are expected to plateau for men in the near future. The largest relative increases occurred for the upper limbs followed by the trunk for both sexes. Comparable generation effects were observed for intermittently exposed sites but the patterns of trend differed between sites for men and women. This supports effects due to sex- and site-specific pattern of sun exposure. The evidence, for the first time, of more favorable trends among post-World War II generations is thought to reflect reduced exposure to UV rays. This may possibly be the earliest signs of the impact of primary prevention programs, particularly if UV radiation also acts as a tumor promoter. PMID- 10530612 TI - A case-control study on family history of liver cancer as a risk factor for hepatocellular carcinoma in North Italy. Brescia HCC Study. AB - OBJECTIVES: We carried out a case-control study to investigate the role of history of liver cancer in a first-degree relative as a risk factor for hepatocellular carcinoma (HCC). METHODS: Two hundred eighty-seven HCC incident cases and 450 subjects unaffected by liver disease (controls) were enrolled in the study. Family history of liver cancer and other malignancies and history of alcohol intake were collected by face-to-face interview. Blood samples were analyzed for HBsAg, anti-HCV and HCV RNA positivity. RESULTS: Family history of liver cancer was associated with HCC (odds ratio [OR] = 2.4; 95% confidence interval [CI] = 1.2-4.7), whereas family history of other malignancies was not (OR = 1.0; 95% CI = 0.61.5). An increased OR for family history of liver cancer was found among subjects negative for the other risk factors (OR = 2.0; 95% CI = 0.6-6.9). A synergism of family history of liver cancer was also evident with hepatitis B and hepatitis C virus infection and with heavy alcohol intake. CONCLUSIONS: This study suggests a role of family history independent from and interacting with known risk factors for hepatocellular carcinoma. PMID- 10530613 TI - Occupational physical activity and risk for breast cancer in a nationwide cohort study in Sweden. AB - OBJECTIVE: Our purpose was to investigate effects of physical activity on risk for breast cancer. METHODS: From the Swedish nationwide censuses in 1960 and 1970 we defined three partly overlapping cohorts of women whose occupational titles allowed reproducible classification of physical demands at work in 1960 (n = 704,904), in 1970 (n = 982,270), or with the same demands in both 1960 and 1970 (n = 253,336). The incidence of breast cancer during 1971-89 was ascertained through record linkage to the Swedish Cancer Register. We used Poisson regression to estimate relative risks (RR). RESULTS: A total of 20,419, 22,840, and 8261 breast cancers, respectively, were detected in the three cohorts. In all three cohorts the risk for breast cancer increased monotonically with decreasing level of occupational physical activity and with increasing socioeconomic status. Among women with the same estimated physical activity level in 1960 and 1970 the RR was 1.3 for sedentary as compared with high/very high activity level (95% CI 1.2-1.4; p for trend < 0.001). Adjustment for socioeconomic status virtually eliminated this association (RR 1.1; 95% CI 0.9-1.2; p for trend 0.12) leaving a statistically significant 30% gradient only among women aged 50-59 years at follow-up. The association between socioeconomic status and breast cancer risk was largely unchanged after adjustment for occupational physical activity. CONCLUSION: The protective effect of occupational physical activity on breast cancer risk, if any, appears to be confined to certain age groups. PMID- 10530614 TI - The shape of age-incidence curves of female breast cancer by hormone-receptor status. AB - OBJECTIVES: Substantial decline of ovarian hormones at menopause plays an important role in breast cancer etiology. Hormones must bind to specific receptors to elicit biological responses, however. We therefore hypothesized and examined whether the age-specific risk of breast cancer, especially its change at menopause, differs by estrogen and progesterone receptor (ER/PR) status. METHODS: Age-specific incidence rates, stratified by ER/PR status, were estimated by multiplying the age-specific ER/PR distribution among 3359 cases in the Danish Breast Cancer Cooperative Group by Danish national age-specific incidence rates. International variations in the age-incidence curve were also reviewed in relation to the hypothesis. RESULTS: The incidence of ER +/PR + subtype (62.9% of all cases) increased with age continually, with a sudden decrease in the rate of increase around age 44. The incidence of ER-/PR- subtype (17.6%) increased with age prior to about age 50 but remained unchanged subsequently. The incidence of ER+ /PR- subtype (13.9%) increased rapidly during the menopausal period but only slightly afterwards. The incidence of ER-/PR+ subtype (5.6%) increased until about age 43 and decreased subsequently. The international comparison revealed Western women, particularly the elderly, might be at substantially higher risk for ER+ /PR+ subtype compared to Japanese women. CONCLUSION: Age-specific risk of breast cancer differs by ER/PR status. The large international variation of breast cancer incidence rates may be explained largely by the risk difference for ER+ /PR+ subtype. PMID- 10530615 TI - Energy intake and dietary pattern in cancer of the oral cavity and pharynx. AB - OBJECTIVE: To explore the role of energy and macronutrients in cancers of the oral cavity and pharynx. METHODS: Case-control study: 754 individuals with first incident cancer of the oral cavity and pharynx and 1775 controls in hospital for acute, non-neoplastic diseases were interviewed in two Italian areas and in the Swiss Canton of Vaud between 1992 and 1997. RESULTS: Cases reported higher total energy intake, due to higher intake of alcohol energy. Non-alcohol energy intake was lower in cases than controls. The composition of diet also differed: proteins (OR for an addition of 100 kcal/day = 0.8) and monounsaturated fatty acids (OR = 0.8) were inversely associated, while saturated fatty acids (OR = 1.4) were directly associated with oral cancer risk. Vegetable intake, which was positively correlated with oil intake, was lower in cases than controls, but accounted only partly for the observed difference in fat intake pattern. CONCLUSIONS: Alcohol energy may not be used efficiently after some threshold. Protein deficiency may enhance cancer risk in heavy drinkers. An opposite influence of different types of fat is emerging for cancer of the oral cavity and pharynx as well as other sites of the upper aero-digestive tract. PMID- 10530616 TI - Elevated risk of cancer of the urinary tract for alcohol drinkers: a meta analysis. AB - OBJECTIVES: Recent narrative reviews have concluded that there is no support for an association between alcohol consumption and urinary tract cancer. Many individual studies, however, have reported positive associations, although rarely statistically significant. The purpose of this meta-analysis is to summarize and quantify this relationship with more statistical power and to perform a sensitivity analysis on the study characteristics. METHODS: We included 16 epidemiological studies published up to April 1999 and calculated summary odds ratios (SORs), both upgraded and adjusted for age, sex and smoking by meta regression analyses. The age- and smoking-adjusted SORs (current alcohol drinking vs. non-drinking) were 1.3 (95% CI 0.9-2.0) for six studies with men and 1.0 (95% CI 0.4-2.6) for four studies with women. RESULTS: The age-, sex- and smoking adjusted SOR was 1.2 (95% CI 0.9-1.7) for seven studies with men and women combined. CONCLUSION: Even though studies differed in methodology, the results were rather consistent. Subgroup analyses by type or amount of alcohol were not possible due to sparse data. We conclude that the available data suggest a slightly increased risk of urinary tract cancer from alcohol consumption for men. The risk related to alcohol consumption for women and the influence of the amount and type of alcohol remain unclear. PMID- 10530617 TI - Cancer in asbestos-exposed occupational cohorts: a meta-analysis. AB - OBJECTIVE: To examine existing asbestos-exposed occupational cohorts and apply a meta-analytic technique to determine the magnitude of association between exposure and lung cancer and to investigate other cancer sites that may be related to such an exposure. METHODS: We summarized the data from 69 asbestos exposed occupational cohorts reporting on cancer morbidity and mortality. Data were extracted regarding numbers of deaths for each cancer, numbers of mesotheliomas, occupations and latency for respiratory, gastrointestinal, urinary and lymphohematopoietic cancers. For each cancer, we calculated a meta-SMR and examined heterogeneity of results using a chi-square test and by calculating a Z statistic for each study. To examine the dose-response effect, we divided the studies into tertiles according to the percentage of mesothelioma deaths that served as a proxy estimation of asbestos exposure. RESULTS: Lung cancer data demonstrated meta-SMRs of 163 and 148 with and without latency, respectively, with significant heterogeneity of results even after stratification according to occupational groups. Stratification of lung cancer studies according to percentage of mesothelioma deaths showed a dose-response effect. Z-scores ranged from -12.21 to + 29.49. Analysis for laryngeal cancer yielded meta-SMRs of 157 and 133 with and without latency, respectively, demonstrating homogeneous results across studies but accompanied by no evidence of a dose-response effect. Data for gastrointestinal cancers showed no evidence of a significant association and no dose-response effect. Kidney cancer demonstrated statistically non-significant meta-SMRs of 120 (95% CI 88-160) and 111 (95% CI 94-131) with and without latency respectively. CONCLUSIONS: This meta-analysis demonstrates a wide variability of the association between occupational asbestos and lung cancer. There was a suggestion of an association between asbestos and laryngeal carcinoma and no clear association with other cancers. PMID- 10530618 TI - Hormone replacement therapy and improved survival among postmenopausal women diagnosed with colon cancer (USA). AB - OBJECTIVES: Hormone replacement therapy (HRT) has been inversely associated with colon cancer incidence in several epidemiologic studies. In this study we used data from a population-based incident case-control study of colon cancer to evaluate the role of HRT use in survival after diagnosis with colon cancer. METHODS: Data from 815 postmenopausal women living in Utah, California, and Minnesota diagnosed between 1 September 1991 and 30 September 1994 were used to examine associations between HRT and survival. RESULTS: After adjusting for age at time of diagnosis, stage of disease at time of diagnosis, study center, and body mass index (BMI), we observed that women who had ever used HRT had a 30% lesser probability of dying of any cause and a 40% lower probability of dying from colon cancer specifically during the follow-up period. Further evaluation by years of HRT use showed that those who had used HRT for 4 or more years had the lowest risk of dying of colon cancer (hazard rate ratio 0.5, 95% confidence interval 0.3-0.9). Evaluation of other lifestyle variables with HRT use did not show significant confounding or effect modification. CONCLUSIONS: These findings suggest that HRT use may improve short-term survival after diagnosis with colon cancer; there is no suggestion that HRT use is detrimental to survival. PMID- 10530619 TI - Prospective study of smoking, antioxidant intake, and lung cancer in middle-aged women (USA). AB - BACKGROUND: Although substantial evidence suggests that higher intake of fruits and vegetables can reduce the adverse impact of smoking on lung cancer risk, great uncertainty exists regarding the specific foods and their constituents that are protective. We therefore examine prospectively the relation between cigarette smoking and lung cancer incidence among women, and quantify the associations between dietary antioxidants, other nutrients, and lung cancer risk. METHODS: In a 16-year prospective cohort study (the Nurses' Health Study), 593 cases of lung cancer were confirmed during 1,793,327 person-years of follow-up. Dietary data, including vitamin supplement use and food intake, were collected in 1980 using a validated semiquantitative food frequency questionnaire. RESULTS: The risk of lung cancer increased with the number of cigarettes smoked and with early onset of cigarette smoking. The risk decreased rapidly with the discontinuation of smoking but took 15 years to fall to about the level of risk for women who had never smoked. Dietary intake of fat was not related to the risk of lung cancer. Although beta-carotene intake was not related to risk, intake of carrots showed a strong inverse relation: women who reported consuming five or more carrots per week had a relative risk of 0.4 (95% CI = 0.2-0.8) compared with the risk for women who never ate carrots. CONCLUSIONS: Smoking is the most important risk factor for lung cancer in women, as it is in men. Higher vegetable consumption, particularly of carrots, may significantly reduce the risk of lung cancer, but dietary modification cannot be considered a substitute for smoking prevention and cessation. PMID- 10530620 TI - Antioxidative action of flavonoids, quercetin and catechin, mediated by the activation of glutathione peroxidase. AB - Antioxidative action of flavonoids have been attracted attention of many investigators and a good deal of studies on it were reported. While their interests were mostly centered to the direct scavenging action of flavonoids against free radicals and active oxygen species, we expected that the interaction of flavonoids and intracellularly occurring antioxidative agents such as glutathione peroxidase (GSH-PO) could synergistically enhance their antioxidative activities. For this purpose, cultured rat hepatocytes (BL-9), which are highly expressing GSH-PO, were employed. One group of the cells were cultured with Se deficient media (Se(-) cells) to diminish the activity and the expression of GSH PO protein and mRNA, and the other group was cultured with Se supplemented media (Se(+) cells). The oxidative cell damage was induced by the addition of H2O2 and two representative antioxidative flavonoids, quercetin and catechin, were added to the media to test their cytoprotective action. In Se(+) cells, the remarkable cytoprotective activity of those flavonoids were confirmed, whereas none of such activity was evidenced in Se(-) cells. It was proved that the intracellular antioxidative function of flavonoids requires the interaction with GSH-PO, at least in the cells expressing the enzyme. Interestingly, the flavonoid activated GSH-PO clearly, and its mechanism is discussed. PMID- 10530621 TI - Immunohistochemical expressions of prohormone convertase (PC)1/3 and PC2 in carcinoids of various organs. AB - In order to clarify the expression of prohormone convertase (PC) 1/3 and PC2 in various carcinoids and non-carcinoid endocrine tumors, we performed indirect immunoperoxidase staining on total of 19 cases of carcinoids (9 cases of bronchial carcinoids, 4 cases of rectal carcinoids, 4 cases of gastric carcinoids and 2 cases of bile duct carcinoids). Our study also included 7 non-carcinoid endocrine tumors. Seventy-nine% and 26% of carcinoids highly or strongly expressed positive staining for PC1/3 and PC2, respectively. High and strong expressions (3+ or 4+) of both PC1/3 and PC2 were noted in only bronchial carcinoids. Strong expressions for only PC1/3 were noted in rectal carcinoids. Bile duct carcinoids also demonstrated higher expressions of PC1/3 than those of PC2. These results suggested that high expressions of both PC1/3 and PC2 in bronchial carcinoids might reflect their diverse and frequent peptide production. The expressions of PC1/3 mRNA and PC2 mRNA detected by in situ hybridization in the bronchial carcinoids and rectal carcinoids were correlated with immunoexpressions of both of the antigens. The granular immunoexpression pattern of PC1/3 and PC2 visualized by confocal laser scanning microscopy would suggest the site of post-translational processing in the secretory granules. Non carcinoid endocrine tumors showed low expressions (+ or 2+) of PC1/3 and PC2, except for thyroid medullary carcinoma showing high immunoexpression of PC1/3. Other non-carcinoid endocrine tumors (parathyroid adenomas and adrenal pheochromocytomas) revealed low immunoexpressions for both PC1/3 and PC2. PMID- 10530622 TI - Vaginal applicators (ovoids) for local control and alleviation of rectal complications of cervical cancers treated by brachytherapy. AB - PURPOSE: To assess the effects of vaginal applicators (ovoids) on dose distribution, local control and rectal complications in cervical cancers treated by brachytherapy. METHODS: From 1984 to 1992, 41 patients (15 stages I+II; 26 stages III+IV) were treated by high-dose-rate brachytherapy (20 Gy in 4 fractions) after external beam irradiation (36-56 Gy). Twenty-three patients were treated by standard application of both intrauterine tandem and ovoids, and 18 patients were treated by non-standard application. RESULTS: The five-year, local relapse-free rates by standard application and tandem alone were 69 and 65% overall; 83 and 100% for stages I+II; and 62 and 49% for stages III+IV. Local control was related to tumor response following the external beam irradiation and initial tumor size by multivariate analysis. Rectal complications seen in patients followed for more than 1 year were 33% after standard application and 22% for tandem use alone. The rectal doses given for standard application (24 Gy) and tandem use (16 Gy) were significantly different. CONCLUSIONS: The ovoid sources did not always contribute to local control, and occasionally led to rectal complications. The optimization of brachytherapy was dependent on patients' anatomy, tumor size, and tumor response. PMID- 10530623 TI - Nurses' perspectives concerning do-not-resuscitate (DNR) orders. AB - The purpose of this study was to investigate the views of the nursing staff concerning do-not-resuscitate (DNR) orders at the Tokai University Hospital where a controversial incident occurred several years ago. A 'Questionnaire on DNR Orders' was circulated and the anonymous answers were collected two weeks later. The questionnaire was returned by 706 of 780 (90.5%) nurses from every ward/specialty, which revealed that 87% of the nurses felt that DNRs were occasionally necessary, with more than 40% of the nurses answering that they took part in DNR. Further, 36% of the nurses stated that patient consent was indispensable, and 64% thought that the patient's family and physician could decide DNR in the event the patient was physically unable to give consent. Moreover, 66% of the nurses expected the establishment of a DNR order sheet to be formulated as a matter of hospital policy; only 5% of the nurses thought that such an order sheet would not be necessary. Comparing these results with a previous study polling physicians at the Tokai University Hospital, nurses are more likely than physicians to think that patient consent is indispensable, and want the establishment of a standardized DNR order sheet as hospital policy. There is, in fact, a "tacit understanding" between physicians and patients' families in medical practice in Japan. However, DNR is definitely a medical decision. Therefore it should be clearly stated in a standardized format, although such a procedure presently seems unlikely, in view of the Japanese traditional value system. PMID- 10530624 TI - A structured intervention for family caregivers of dementia patients: a pilot study. AB - There has been an increasing number of patients with dementia in Japan. Although such patients were hospitalized longer than in other countries, the length of the hospital stay is becoming shorter due to changes in insurance systems. Therefore, the families of such patients are experiencing greater stress. In order to investigate the efficacy of a group structured intervention, 20 family caregivers participated in a series of five weekly sessions, each of which consisted of an educational approach, problem-solving techniques, psychological support, and relaxation. All family caregivers were women whose ages ranged from 47-66 years (mean= 54.7 +/- 4.4). The period of care at home ranged from 1-12 years (mean= 5.8 +/- 2.7). Concerning the original disease of patients, 10 had vascular dementia and 8 had senile dementia of Alzheimer type (Alzheimer disease). Two psychometries, i. e., Profile of Mood States (POMS) and General Health Questionnaire-30 (GHQ-30), were administered pre- and post-intervention. The results indicated that there was significant improvement (p<0.05) in the scores of depression, anger-hostility, fatigue, confusion in the POMS, and physical symptoms, anxiety-mood disorder, and suicidality-depression in the GHQ-30. This preliminary study suggests that this kind of intervention appears quite effective for relieving the emotional and physical discomfort suffered by family caregivers. PMID- 10530625 TI - Sexual orientation and suicidality: a co-twin control study in adult men. AB - BACKGROUND: Several recent studies have found a higher lifetime prevalence of suicide attempts in homosexual males compared with heterosexual control subjects or population rates. These studies used either convenience samples, most without controls, or population-based samples in which confounding factors such as depression and substance abuse were not measured. METHODS: This study used twins from the population-based Vietnam Era Twin Registry, Hines, Ill. An analytic sample of 103 middle-aged male-male twin pairs from the registry was identified in which one member of the pair reported male sex partners after age 18 years while the other did not. Four lifetime symptoms of suicidality as measured by the Diagnostic Interview Schedule were analyzed: thoughts about death, wanting to die, thoughts about committing suicide, and attempted suicide. A composite measure of reporting at least one suicidality symptom was also assessed. RESULTS: Same-gender sexual orientation is significantly associated with each of the suicidality measures. Unadjusted matched-pair odds ratios follow: 2.4 (95% confidence interval [CI], 1.2 - 4.6) for thoughts about death; 4.4 (95% CI, 1.7 - 11.6) for wanted to die; 4.1 (95% CI, 2.1 - 8.2) for suicidal ideation; 6.5 (95% CI, 1.5 - 28.8) for attempted suicide; and 5.1 (95% CI, 2.4 - 10.9) for any of the suicidal symptoms. After adjustment for substance abuse and depressive symptoms (other than suicidality), all of the suicidality measures remain significantly associated with same-gender sexual orientation except for wanting to die (odds ratio, 2.5 [95% CI, 0.7 - 8.81). CONCLUSIONS: The substantially increased lifetime risk of suicidal behaviors in homosexual men is unlikely to be due solely to substance abuse or other psychiatric comorbidity. While the underlying causes of the suicidal behaviors remain unclear, future research needs to address the inadequacies in the measurement of both sexual orientation and suicidality in population-based samples. PMID- 10530626 TI - Is sexual orientation related to mental health problems and suicidality in young people? AB - BACKGROUND: This study examines the extent to which gay, lesbian, and bisexual young people are at increased risk of psychiatric disorder and suicidal behaviors using data gathered on a New Zealand birth cohort studied to age 21 years. METHODS: Data were gathered during the course of the Christchurch Health and Development Study, a 21-year longitudinal study of a birth cohort of 1265 children born in Christchurch, New Zealand. At 21 years of age, 1007 sample members were questioned about their sexual orientation and relationships with same-sex partners since the age of 16 years. Twenty-eight subjects (2.8%) were classified as being of gay, lesbian, or bisexual sexual orientation. Over the period from age 14 to 21 years, data were gathered on a range of psychiatric disorders that included major depression, generalized anxiety disorder, conduct disorder, and substance use disorders. Data were also gathered on suicidal ideation and suicide attempts. RESULTS: Gay, lesbian, and bisexual young people were at increased risks of major depression (odds ratio [OR], 4.0; 95% confidence interval [CI], 1.8-9.3), generalized anxiety disorder (OR, 2.8; 95% CI, 1.2-6.5), conduct disorder (OR, 3.8; 95% CI, 1.7-8.7), nicotine dependence (OR, 5.0; 95%, CI, 2.3-10.9), other substance abuse and/or dependence (OR, 1.9; 95% CI, 0.9 4.2), multiple disorders (OR, 5.9; 95% CI, 2.4-14.8), suicidal ideation (OR, 5.4; 95% CI, 2.4-12.2), and suicide attempts (OR, 6.2; 95% CI, 2.7-14.3). CONCLUSIONS: Findings support recent evidence suggesting that gay, lesbian, and bisexual young people are at increased risk of mental health problems, with these associations being particularly evident for measures of suicidal behavior and multiple disorder. PMID- 10530627 TI - Homosexuality and mental illness. PMID- 10530628 TI - Suicide and sexual orientation: nearing the end of controversy? PMID- 10530629 TI - Homosexuality, psychopathology, and suicidality. PMID- 10530630 TI - Minor and major depression and the risk of death in older persons. AB - BACKGROUND: The association between depression and mortality in older community dwelling populations is still unresolved. This study determined the effect of both minor and major depression on mortality and examined the role of confounding and explanatory variables on this relationship. METHODS: A cohort of 3056 men and women from the Netherlands aged 55 to 85 years were followed up for 4 years. Major depression was defined according to DSM-III criteria by means of the Diagnostic Interview Schedule. Minor depression was defined as clinically relevant depression (defined by a Center for Epidemiologic Studies Depression score > or = 16) not fulfilling diagnostic criteria for major depression. RESULTS: After adjustment for confounding variables (sociodemographics, health status), men with minor depression had a 1.80-fold higher risk of death (95% confidence interval, 1.35-2.39) during follow-up than nondepressed men. In women, minor depression did not significantly increase the mortality risk. Irrespective of sex, major depression was associated with a 1.83-fold higher mortality risk (95% confidence interval, 1.09-3.10) after adjustment for sociodemographics and health status. Health behaviors such as smoking and physical inactivity explained only a small part of the excess mortality risk associated with depression. CONCLUSION: Even after adjustment for sociodemographics, health status, and health behaviors, minor depression in older men and major depression in both older men and women increase the risk of dying. PMID- 10530631 TI - Functioning and utility for current health of patients with depression or chronic medical conditions in managed, primary care practices. AB - BACKGROUND: Health utility is the recommended outcome metric for medical cost effectiveness studies. We compared health utility and quality of life for primary care patients with depression or chronic medical conditions. METHODS: Respondents were outpatients (N = 17 558) of primary care clinicians (N = 181) in 7 managed care organizations. Utility was assessed by time tradeoff, or the years of life that patients would exchange for perfect health, and standard gamble, or the required chance of success to accept a treatment that can cause immediate death or survival in perfect health. Probable 12-month depressive disorder and affective syndromes were assessed through self-report items from a diagnostic interview. Medical conditions were assessed with self-report. Quality of life was assessed by the 12-Item Short-Form Health Survey. Regression models were used to compare quality of life and utility for patients with depression vs chronic medical conditions. RESULTS: Patients with probable 12-month depressive disorder had worse mental health and role-emotional and social functioning and lower utility for their current health than patients with each chronic medical condition (for most comparisons, P<.001). Depressed patients had worse physical functioning than patients with 4 common chronic conditions but better physical functioning than patients with 4 other conditions (each P<.001). Patients with lifetime bipolar illness and 12-month double depression had the poorest quality of life and lowest utility. CONCLUSIONS: Primary care patients with depressive conditions have poorer mental, role-emotional, and social functioning than patients with common chronic medical conditions, and physical functioning in the midrange. The low utility of depressed patients relative to patients with chronic medical conditions suggests that recovery from depression should be a high practice priority. PMID- 10530632 TI - Reduced gray matter volume in schizophrenia. AB - BACKGROUND: There is emerging evidence that gray matter (GM) is reduced in patients with schizophrenia. Information on the extent of global differences in the 3 principal supertentorial compartments is necessary for interpretation of regional effects. The relation of GM reduction to clinical status and neurocognition also requires examination. METHODS: Magnetic resonance imaging, neurocognitive measures, and clinical assessment of symptoms and functioning were obtained for 130 patients (51 neuroleptic naive, 79 previously treated) and 130 healthy controls (75 men, 55 women in each group). RESULTS: Overall GM volume was reduced in patients compared with controls. This was evident in men (6% reduction) and women (2% reduction) and was already evident at the first presentation of neuroleptic-naive patients. The reduction sustained correction for age and total intracranial volume. Compartmental volumes did not correlate with the severity of positive (r, -0.08 to 0.23) or negative (r, -0.01 to -0.07) symptoms, but GM volume was associated with better premorbid functioning in women (r, 0.36-0.51). Small but significant correlations (r, 0.19-0.44) were observed between GM volume and performance in 6 neurocognitive domains. These correlations varied by diagnosis, most higher in patients, and were moderated by sex. CONCLUSIONS: Gray matter volume reduction in schizophrenia is already evident in men and women at first presentation. While this reduction is not correlated with symptom severity, it is associated with cognitive performance. Since GM development accelerates in the later part of gestation, while white matter growth is primarily postnatal, the results may support the hypothesis that neurodevelopmental processes relate to GM deficit. PMID- 10530633 TI - Orbital frontal and amygdala volume reductions in obsessive-compulsive disorder. AB - BACKGROUND: Functional neuroimaging studies have implicated the frontal lobes and the hippocampus-amygdala complex in the pathophysiology of obsessive-compulsive disorder (OCD). These brain regions have not been well investigated in patients with OCD, however, using magnetic resonance imaging. METHODS: Volumes of the superior frontal gyrus, anterior cingulate gyrus, orbital frontal region, hippocampus, and amygdala were computed from contiguous magnetic resonance images in a sample of 26 patients with OCD and 26 healthy comparison subjects. RESULTS: Patients with OCD had significantly reduced bilateral orbital frontal and amygdala volumes compared with healthy comparison subjects and lacked the normal hemispheric asymmetry of the hippocampus-amygdala complex. Neither brain structure volumes nor asymmetry indices were significantly correlated with total illness duration or length of current OCD episode. CONCLUSIONS: Findings of reduced orbital frontal and amygdala volumes in patients implicate a structural abnormality of these brain regions in the pathophysiology of OCD. Absence of the normal hemispheric asymmetry of the hippocampus-amygdala complex in patients is consistent with an anomalous neurodevelopmental process. PMID- 10530635 TI - Toward the identification of core psychopathological processes? PMID- 10530634 TI - The structure of common mental disorders. AB - BACKGROUND: This report presents the results of confirmatory factor analyses of patterns of comorbidity among 10 common mental disorders in the National Comorbidity Survey, a national probability sample of US civilians who completed structured diagnostic interviews. METHODS: Patterns of comorbidity among DSM-III R mental disorders were analyzed via confirmatory factor analyses for the entire National Comorbidity Survey sample (N = 8098; age range, 15-54 years), for random halves of the sample, for men and women separately, and for a subsample of participants who were seeing a professional about their mental health problems. Four models were compared: a 1-factor model, a 2-factor model in which some disorders represented internalizing problems and others represented externalizing problems, a 3-factor variant of the 2-factor model in which internalizing was modeled as having 2 subfactors (anxious-misery and fear), and a 4-factor model in which the disorders represented separate affective, anxiety, substance dependence, and antisocial factors. RESULTS: The 3-factor model provided the best fit in the entire sample. This result was replicated across random halves of the sample as well as across women and men. The substantial empirical intercorrelation between anxious-misery and fear (0.73) suggested that these factors were most appropriately conceived as subfactors of a higher-order internalizing factor. In the treatment sample, the 2-factor model fit best. CONCLUSIONS: The results offer a novel perspective on comorbidity, suggesting that comorbidity results from common, underlying core psychopathological processes. The results thereby argue for focusing research on these core processes themselves, rather than on their varied manifestations as separate disorders. PMID- 10530637 TI - High rates of schizophrenia in adults with velo-cardio-facial syndrome. AB - BACKGROUND: Velo-cardio-facial syndrome (VCFS), a syndrome characterized by an increased frequency of schizophrenia and bipolar disorder, is associated with small interstitial deletions of chromosome 22q11. METHODS: We evaluated 50 adults with VCFS using a structured clinical interview (Schedules for Clinical Assessment in Neuropsychiatry or Psychiatric Assessment Schedule for Adults With Developmental Disability if IQ <50) to establish a DSM-IV diagnosis. The schizophrenia phenotype in individuals with VCFS and schizophrenia was compared with a matched series of individuals with schizophrenia and without VCFS (n = 12). The King's Schizotypy Questionnaire was administered to individuals with VCFS (n = 41), their first-degree relatives (n = 68), and a series of unrelated normal controls (n = 316). All individuals with VCFS deleted for the N25 probe (n = 48) were genotyped for a genetic polymorphism in the COMT gene that results in variations in enzymatic activity. RESULTS: Fifteen individuals with VCFS (30%) had a psychotic disorder, with 24% (n = 12) fulfilling DSM-IV criteria for schizophrenia. In addition, 6 (12%) had major depression without psychotic features. The individuals with schizophrenia had fewer negative symptoms and a relatively later age of onset compared with those with schizophrenia and without VCFS. We found no evidence that possession of the low-activity COMT allele was associated with schizophrenia in our sample of individuals with VCFS. CONCLUSIONS: The high prevalence of schizophrenia in this group suggests that chromosome 22q11 might harbor a gene or genes relevant to the etiology of schizophrenia in the wider population. PMID- 10530638 TI - Clozapine plus lamotrigine in treatment-resistant schizophrenia. PMID- 10530636 TI - Differential response to antidepressants in women with premenstrual syndrome/premenstrual dysphoric disorder: a randomized controlled trial. AB - BACKGROUND: Studies show that selective serotonin reuptake inhibitors are effective for severe premenstrual syndrome and premenstrual dysphoric disorder. This study compares the efficacy of a selective serotonin reuptake inhibitor with that of a tricyclic antidepressant to determine whether efficacy for premenstrual syndrome/premenstrual dysphoric disorder is a general or more serotonergic effect of antidepressants. METHODS: After 3 screening months, 189 subjects were randomized to sertraline hydrochloride, desipramine hydrochloride, or placebo for 3 months of double-blind treatment. The flexible dosage range was 50 to 150 mg/d. The outcome measures included the Penn Daily Symptom Report (DSR), the Hamilton Depression Rating Scale, the Clinical Global Impressions-Severity Scale, the Quality of Life Scale, and Patient Global Ratings of Functioning and Improvement. Analyses included all subjects with treatment data, with the last observation carried forward. RESULTS: Sertraline was significantly more effective than placebo or desipramine; desipramine was not better than placebo (F2,163 = 12.47, P<.001). All DSR factors were more improved with sertraline compared with desipramine and placebo; the factors for mood (P<.001) and pain (P = .05) were significant, and the results of all outcome measures were consistent. A history of depression, postmenstrual symptom levels, and other diagnostic variables added individually as covariates did not alter the treatment results. At end point analysis, DSR symptoms had decreased by more than 50% in 40 subjects (65%) in the sertraline group, 18 subjects (36%) in the desipramine group, and 16 subjects (29%) in the placebo group (P<.001). CONCLUSIONS: The comparison of 2 classes of antidepressants strongly favored the serotonergic drug, which effectively reduced symptoms and improved functioning and was well tolerated by women with severe premenstrual syndrome. A history of depression did not alter the treatment results. PMID- 10530639 TI - Critical differences between binge eating and overeating. PMID- 10530640 TI - The epidemiology and control of VRE: still struggling to come of age. PMID- 10530641 TI - Molecular epidemiology of vancomycin-resistant enterococci: a 2-year perspective. AB - OBJECTIVE: To determine the molecular epidemiology of vancomycin-resistant enterococci (VRE) at our medical center in order to identify the extent of strain clonality and possible transmission patterns of this pathogen. DESIGN: An important facet of our infection control program includes molecular typing of all clinical and surveillance isolates of VRE to determine transmission patterns in the hospital. Molecular strain typing is performed by restriction endonuclease analysis (REA) of genomic DNA. REA patterns are visually compared to categorize VRE strains into type and subtype designations. SETTING: A 588-bed, university affiliated, tertiary-care hospital and a neighboring 155-bed rehabilitation facility. RESULTS: From January 1995 through December 1996, 379 VRE isolates were collected from 197 patients. Thirty-three genotypes were determined by REA typing; 15 genotypes were implicated in 29 instances of potential nosocomial transmission. Three major clusters of VRE involving patients on multiple nursing units and two adjacent hospitals were identified. The remaining instances of nosocomial transmission occurred in small patient clusters. CONCLUSIONS: In conclusion, the VRE epidemic at this medical center is polyclonal. VRE transmission patterns are complex, and, while large clusters do occur, the usual pattern of nosocomial acquisition of this pathogen occurs in the setting of "mini clusters". PMID- 10530642 TI - Association between mucositis severity and vancomycin-resistant enterococcal bloodstream infection in hospitalized cancer patients. AB - OBJECTIVE: To determine the role of mucositis severity in the development of vancomycin-resistant enterococcal (VRE) bloodstream infection (BSI). SETTING: A tertiary-care university medical center. PARTICIPANTS: Hematology-oncology-unit inpatients. DESIGN: Patients with VRE BSI (case-patients) were compared with VRE colonized (control) patients from September 1994 through August 1997. Oral mucositis severity was recorded on the day of VRE BSI for case-patients and on hospital day 22 (median day of hospitalization of case-patient VRE BSI) for controls. There were 19 case-patients and 31 controls. RESULTS: In univariate analysis, case-patients were significantly more likely than controls to have a higher mucositis severity score, diarrhea, or a higher severity of illness score. In multivariate analysis, only mucositis remained as an independent risk factor, and increasing mucositis score was significantly associated with VRE BSI. CONCLUSIONS: Mucositis severity was independently associated with an increasing risk for VRE BSI. Interventions to alter mucositis severity may help to prevent VRE BSI in hospitalized cancer patients. PMID- 10530643 TI - Enteric carriage of vancomycin-resistant Enterococcus faecium in patients tested for Clostridium difficile. AB - OBJECTIVE: To identify independent risk factors for enteric carriage of vancomycin-resistant Enterococcus faecium (VREF) in hospitalized patients tested for Clostridium difficile toxin. DESIGN: Retrospective case-cohort study. SETTING: Tertiary-care teaching hospital. PATIENTS: Convenience sample of 215 adult inpatients who had stool tested for C difficile between January 29 and February 25, 1996. RESULTS: 41 (19%) of 215 patients had enteric carriage of VREE Five independent risk factors for enteric VREF were identified: history of prior C difficile (odds ratio [OR], 15.21; 95% confidence interval [CI95], 3.30-70.10; P < .001), parenteral treatment with vancomycin for > or = 5 days (OR, 4.06; CI95, 1.54-10.73; P = .005), treatment with antimicrobials effective against gram negative organisms (OR, 3.44; CI95, 1.20-9.87; P = .021), admission from another institution (OR, 2.95; CI95, 1.21-7.18; P =.017), and age > 60 years (OR 2.57; CI95, 1.13-5.82; P = .024). These risk factors for enteric VREF were independent of the patient's current C difficile status. CONCLUSIONS: Antimicrobial exposures are the most important modifiable independent risk factors for enteric carriage of VREF in hospitalized patients tested for C difficile. PMID- 10530644 TI - Reporting of vancomycin-resistant enterococci in Connecticut: implementation and validation of a state-based surveillance system. AB - OBJECTIVE: To assess state-based surveillance for isolation from a sterile site of vancomycin-resistant enterococci (VRE) in Connecticut. DESIGN: Clinical laboratory reporting (passive surveillance) of VRE isolates to the Connecticut Department of Public Health (CDPH) was followed by state-initiated validation, laboratory proficiency testing, and review of hospital demographic characteristics. SETTINGS: All 45 clinical laboratories and all 37 (36 for 1995 and 1996) acute-care hospitals in Connecticut were included in the study. MAIN OUTCOME MEASURES: The outcome measures included determination of the statewide incidence of VRE and the accuracy of passive reporting, determination of clinical laboratory proficiency in detecting VRE, and analysis of hospital characteristics that might be associated with an increased incidence of VRE. RESULTS: During 1994 through 1996, 29 (78%) of 37 hospital-affiliated clinical laboratories and 1 (11%) of 9 commercial or other laboratories in Connecticut reported to the CDPH the isolation of VRE from sterile sites; 158 isolates were reported for these 3 years. Based on verification, we discovered that these laboratories actually detected 58 VRE isolates in 1994, 104 in 1995, and 104 in 1996 (total, 266). The age-standardized incidence rate of VRE was 14.1 cases per million population in 1994 and 26.8 cases per million population for both 1995 and 1996. Laboratory proficiency testing revealed that high-level vancomycin resistance was identified accurately and that low- and moderate-level resistance was not detected. The incidence of VRE isolates was three times greater in hospitals with over 300 beds compared with categories of hospitals with fewer beds. Increases in the number of VRE isolates were at least twice as likely in hospitals located in areas with a higher population density, or with a residency program or trauma center in the hospital. CONCLUSIONS: Passive reporting of VRE isolates from sterile sites markedly underestimated the actual number of iso lates, as determined in a statewide reporting system. Statewide passive surveillance systems for routine or emerging pathogens must be validated and laboratory proficiency ensured if results are to be accurate and substantial underreporting is to be corrected. PMID- 10530645 TI - Comparative in vitro activity of antiseptics and disinfectants versus clinical isolates of Candida species. AB - OBJECTIVE: To evaluate the in vitro activity of antiseptics and detergents against Candida. DESIGN: One strain each of Candida albicans, Candida tropicalis, Candida lusitaniae, Candida parapsilosis, Candida kefyr, Candida glabrata, and an American Type Culture Collection strain of Escherichia coli (control) were studied. Clinical isolates were obtained from patients in a bone marrow unit of a large tertiary hospital. Antiseptic and disinfectant agents studied were used in the hospital where isolates were identified for cleaning of inanimate surfaces or hand washing. In vitro susceptibility was determined using a broth macrodilution method with exposure times to antiseptic or disinfectant agent of 15 seconds to 4 minutes and concentrations of agents that ranged from undiluted to 1:10,000 dilution. SETTING: A 900-bed teaching hospital. RESULTS: Of disinfectants tested, Vestal and Sparquat inhibited growth of all species at dilutions of < or = 1:100 at all contact times for all species. Clorox showed inhibition of growth at 1:100 dilution after 30 seconds of contact time for all isolates. Of antiseptics studied, Hibiclens inhibited growth of all species except C tropicalis at dilutions of < or = 1:100 at all contact times and for C tropicalis after 60 seconds. Clinidine inhibited growth of all species at dilutions of < or = 1:100 at all contact times for all species with the exception of Cglabrata and C tropicalis, which grew at the undiluted concentration. Ultradex failed to demonstrate killing of any species for any dilutions tested. CONCLUSIONS: The results of this study show varying degrees of in vitro inhibition of growth by a variety of antiseptics and disinfectants against clinical isolates of Candida species from hospitalized patients. PMID- 10530646 TI - Undetected vancomycin-resistant Enterococcus in surgical intensive care unit patients. AB - The rates of vancomycin-resistant Enterococcus (VRE) in a high-risk population were investigated prospectively using an active surveillance method. The costs of conducting active surveillance were calculated. Among the 10 patients found to have VRE, routine cultures identified 3 (30%); thus, 70% of the VRE-colonized patients would have gone undetected in the absence of active surveillance. The total cost for 5 weeks of active surveillance was $2,234. Although active surveillance identified a high rate of VRE-colonized patients who otherwise may not have been identified, it remains to be determined if the additional costs are justified and result in reduced transmission. PMID- 10530647 TI - Recovery of high-level streptomycin-resistant enterococci from hemodialysis water and dialysate in 85 Greek renal units. AB - In the 85 renal units of Greece, enterococci were recovered from 10 samples of tap water, 6 of treated hemodialysis water, and 21 of dialysate. Eleven isolates were Enterococcus faecium, and 8 were Enterococcus raffinosus; 6 other additional enterococcal species were found. Twenty-two strains exhibited high-level resistance to streptomycin, 16 were resistant to rifampicin, and one to erythromycin. In our hemodialysis units, treated water and dialysate raise concern regarding transfer to patients of uncommon enterococcal species exhibiting high-level streptomycin resistance. PMID- 10530648 TI - Methicillin-resistant Staphylococcus aureus in the community: a hospital-based study. AB - To determine the proportion of methicillin-resistant Staphylococcus aureus (MRSA) among patients presenting for hospitalization and to assess risk factors for MRSA carriage, we conducted a study for 13 months at five Pittsburgh-area hospitals. Of 504 S aureus identified, 125 (25%) were MRSA. Independent risk factors for MRSA included organ transplantation, employment in a healthcare facility, pressure sores, tube feeding, and hospitalization within the preceding year. PMID- 10530649 TI - Do influenza epidemics affect patterns of sickness absence among British hospital staff? AB - Influenza vaccination for healthcare workers is not recommended in Britain, but some hospitals offer vaccine to reduce sickness absence. However, in Nottingham, the influenza epidemics of 1993-94 and 1996-97 made no impact on staff absence. Annual vaccination of healthcare workers against influenza is unlikely to reduce absence most winters, but there may be gains in terms of preventing nosocomial infection. PMID- 10530650 TI - Requirements for infrastructure and essential activities of infection control and epidemiology in out-of-hospital settings: a consensus panel report. Association for Professionals in Infection Control and Epidemiology and Society for Healthcare Epidemiology of America. AB - In 1997 the Association for Professionals in Infection Control and Epidemiology and the Society for Healthcare Epidemiology of America established a consensus panel to develop recommendations for optimal infrastructure and essential activities of infection control and epidemiology programs in out-of-hospital settings. The following report represents the Consensus Panel's best assessment of requirements for a healthy and effective out-of-hospital-based infection control and epidemiology program. The recommendations fall into 5 categories: managing critical data and information; developing and recommending policies and procedures; intervening directly to prevent infections; educating and training of health care workers, patients, and nonmedical caregivers; and resources. The Consensus Panel used an evidence-based approach and categorized recommendations according to modifications of the scheme developed by the Clinical Affairs Committee of the Infectious Diseases Society of America and the Centers for Disease Control and Prevention's Healthcare Infection Control Practices Advisory Committee. PMID- 10530651 TI - Who is at risk of what? AB - If you have calculated the sample size required for an employee survey or an observational study of departmental practices but found that the number of observations required is larger than the number of employees, chances are the error is due to use of approximation formulae. Many of us unknowingly were taught to use approximations that fail to include the finite population correction factor. Depending on the objective of a study and the proportion of a population sampled, it may be necessary to consider this correction factor in order to estimate standard error and sample size accurately. PMID- 10530652 TI - Propofol for emergency department procedural sedation--not yet ready for prime time. PMID- 10530653 TI - The continuing search to identify the very-low-risk chest pain patient. PMID- 10530654 TI - Misguided residency applicant questions? PMID- 10530655 TI - A comparison of the bioavailabilities of oral and intravenous ethanol in healthy male volunteers. AB - OBJECTIVES: Ethanol (EtOH), the antidote for methanol and ethylene glycol, is administered by the oral (PO) and intravenous (IV) routes. Serum concentrations (SCs) of 100 mg/dL or more are targeted for clinical effect. This study was completed to validate the assumption that there are minimal differences in SC achieved between these two routes. METHODS: Twenty healthy male volunteers were randomized to receive either PO or IV EtOH. Subjects abstained from EtOH for 48 hours before each phase. After a seven-day washout period, the subjects crossed over to the other group. Inclusion criteria were no history of medical problems, age between 21 and 40 years, and actual body weight within 10% of ideal weight. Baseline EtOH SCs were obtained before participation in each phase. Two hours after a standard breakfast, the subjects received 700 mg/kg of PO or IV EtOH. PO EtOH was administered as a 20% solution in juice over 10 minutes. IV EtOH, controlled by an infusion pump, was administered as a 10% solution over 30 minutes. Blood was drawn for EtOH SCs at 45, 75, 105, 135, 165, 225, 285, and 345 minutes after start of the dose. RESULTS: All initial EtOH SCs were 0. EtOH SCs were higher after IV administration. Mean peak SC was 103.6 mg/dL after IV administration and 71.3 mg/dL after PO administration (p<0.0001). Mean time to peak was 46.5 minutes after IV administration and 103.5 minutes after PO administration (p<0.0001). Total area under the curve was 17,440 min-mg/dL after IV administration and 13,875 min-mg/dL after PO administration (p<0.003). The order of treatments did not affect results (p>0.1). CONCLUSION: Significant differences exist between the SCs of EtOH as well as the times to peak SC after PO and IV administrations. PMID- 10530656 TI - A clinical trial of propofol vs midazolam for procedural sedation in a pediatric emergency department. AB - OBJECTIVE: To compare the effectiveness, recovery time from sedation, and complication rate of propofol with those of midazolam when used for procedural sedation in the pediatric emergency department (PED). METHODS: A prospective, blinded, randomized, clinical trial comparing propofol and midazolam was conducted in the PED of a tertiary pediatric center. Eligible patients were aged 2-18 years with isolated extremity injuries necessitating closed reduction. All patients received morphine for pain, then were randomized to receive propofol or midazolam for sedation. Vital signs, pulse oximetry, and sedation scores were recorded prior to sedation and every 5 minutes thereafter until recovery. Recovery time, time from cast completion to discharge, and other time intervals during the PED course and all sedation-related complications were also recorded. RESULTS: Between August 1996 and October 1997, 91 patients were enrolled. Demographic data, morphine doses, and sedation scores were similar between the propofol and midazolam groups. Mean +/- SD recovery time for the propofol group was 14.9+/-11.1 minutes, compared with 76.4+/-47.5 minutes for the midazolam group, p<0.001. Mild transient hypoxemia was the most significant complication, occurring in 5 of 43 (11.6%) patients given propofol and 5 of 46 (10.9%) patients given midazolam (odds ratio 1.08, 95% CI = 0.24 to 4.76). CONCLUSION: In this study, propofol induced sedation as effectively as midazolam but with a shorter recovery time. Complication rates for propofol and midazolam were comparable, though the small study population limits the power of this comparison. Propofol may be an appropriate agent for sedation in the PED; however, further study is necessary before routine use can be recommended. PMID- 10530657 TI - Resting sestamibi imaging for the prognosis of low-risk chest pain. AB - OBJECTIVE: To assess the prognostic value of resting Tc-99m sestamibi scanning for adverse cardiac events (ACEs) in ED chest pain patients with a low probability of acute cardiac ischemia (ACI). METHODS: Sixty-nine consenting, hemodynamically stable patients with chest pain and a nondiagnostic electrocardiogram received an injection of 25 mCi of sestamibi during or within two hours of active pain. Scans were interpreted locally by a nuclear cardiologist or radiologist. Interrater reliability was assessed. ACEs of myocardial infarction (MI), death, or revascularization were assessed during the index hospitalization and over a one-year follow-up period. RESULTS: For ACEs, rest scanning with sestamibi had a sensitivity of 71% (95% CI = 0.33 to 0.97), a specificity of 92% (95% CI = 0.82 to 0.97), and an accuracy of 90% (95% CI = 0.87 to 0.99). The positive predictive value was 50% (95% CI = 0.19 to 0.82) and the negative predictive value was 97% (95% CI = 0.87 to 0.98). Sestamibi scanning was highly discriminating, with 62% of patients with positive scans but only 3% with negative scans having ACEs (p<0.001, log rank test). CONCLUSION: In patients with low-risk chest pain, sestamibi scanning has good specificity and moderate sensitivity for ACEs over a 12-month period. PMID- 10530658 TI - Validation of the Ottawa Ankle Rules in children with ankle injuries. AB - OBJECTIVES: The Ottawa Ankle Rules (OAR) have been found to be 100% sensitive in adult patients with ankle injuries, and application of the OAR has resulted in a 28% reduction in the number of x-rays ordered. The objectives of this study were to determine the sensitivity and specificity of the OAR in children and to determine the potential change in x-ray utilization. METHODS: Children, aged 2-16 years, presenting to the EDs of two children's hospitals, with an ankle injury in the previous 48 hours, were enrolled. All patients were assessed by either staff physicians or fellows. X-rays were ordered according to standard clinical practice. Prior to reviewing x-rays, the physical examination was recorded on a standardized form. Positive outcomes (clinically significant) were defined as fractures with fragments > or =3 mm. Patients not x-rayed and asymptomatic at five to seven days postinjury were considered to have no significant fracture. RESULTS: Six hundred seventy patients were enrolled. The OAR were 100% sensitive (95% CI = 95% to 100%) for significant ankle fractures, with a specificity of 24% (95% CI = 20% to 28%). The OAR were 100% sensitive (95% CI = 82% to 100%) for the midfoot, with a specificity of 36% (95% CI = 29% to 43%). If the OAR had been followed, there would have been a reduction of ankle x-rays by 16% and foot x rays by 29% without missing any clinically significant fracture. However, analysis of the two hospitals showed that if the rules had been applied, one would have a reduction in x-rays, while the other center would have an increase. CONCLUSIONS: This study demonstrates the OAR to be sensitive for detecting clinically significant (> or =3 mm) ankle and midfoot fractures in children. The application of these rules may reduce the number of x-rays ordered. A further study is required to determine the effect of using the OAR in clinical practice. PMID- 10530659 TI - A decision guideline for emergency department utilization of noncontrast head computed tomography in HIV-infected patients. AB - OBJECTIVE: To determine which neurologic signs or symptoms are predictive of new focal lesions on head CT in HIV-infected patients. METHODS: Prospective study with convenience sample enrollment of HIV-infected patients who presented to a large inner-city university-based ED over an 11-month period. Patients were assessed using a standardized neurologic evaluation to ascertain whether they had developed new or changed neurologic signs or symptoms. Patients with any new or changed neurologic findings had a head CT scan in the ED. The association between individual complaints or findings and new focal lesions on head CT was assessed by univariate analysis, and sensitivity, specificity, and positive predictive values were calculated. Stepwise logistic regression analysis was then carried out to estimate the relative risk for those variables independently associated with new focal lesions on CT scans. A decision guideline was developed incorporating those variables. RESULTS: One hundred ten patients were identified as having new or changed neurologic signs or symptoms and had a head CT done in the ED. Twenty-seven patients (24%) had focal lesions on head CT, of which 19 (18%) were identified as new focal lesions; eight of these (7%) demonstrated a mass effect. Clinical findings most strongly associated with new focal findings on head CT were: 1) new seizure, relative risk (RR) = 73.5, 95% CI = 6.2 to 873.0; 2) depressed or altered orientation, RR = 39.1, 95% CI = 4.6 to 330.0; and 3) headache, different in quality, RR = 27.0, 95% CI = 3.2 to 230.1. Use of these three findings as a screen for ordering head CT in the ED would have identified 95% (18/19) of the patients with new focal intracranial lesions, and resulted in a 53% reduction in the number of head CTs ordered in the ED. Inclusion of one additional parameter (prolonged headache, > or =3 days), would have resulted in identification of 100% of all new focal lesions, with a 37% reduction in the number of head CTs ordered. Among those patients with new focal findings, 74% required emergent management (i.e., seizure control, IV antibiotics, IV steroids or surgery). The most common intracranial lesion among patients with CD4 counts less than 200 cells/microL was toxoplasmosis, while cerebrovascular accidents (ischemic or hemorrhagic) were most common in those with CD4 counts greater than 200 cells/microL. CONCLUSION: Specific clinical signs and symptoms were associated with the presence of new intracranial lesions in a group of HIV infected patients who presented to the ED with neurologic complaints. These clinical findings can be incorporated into guidelines for determining the need for emergent head CT. Validation and widespread application of these guidelines could result in limiting the use of emergent neuroimaging to a more well-defined HIV-infected patient population. PMID- 10530660 TI - Decreasing length of stay with emergency ultrasound examination of the gallbladder. AB - OBJECTIVE: To determine whether patients who received emergency screening ultrasound examinations (ESUEs) of the gallbladder by emergency physicians (EPs) have a shorter ED length of stay (LOS) than do those receiving ultrasound studies from radiology. METHODS: A retrospective chart review from July 1995 to August 1998 identified 1,242 patients who received gallbladder ultrasound examinations. Seven hundred fifty-three patients received ESUEs by EPs of varying levels of ultrasound experience. Four hundred eighty-nine patients received gallbladder ultrasound examinations from radiology, and were not scanned by EPs. The LOSs of the two groups were compared. Significance was evaluated using a two-tailed t test. RESULTS: When patients received an ESUE by an EP, the median LOS was 7% (22 min) less than that for those who received an ultrasound examination by radiology (p = 0.017; 95% CI = 4 min to 41 min). When evaluated by disposition, patients discharged home and scanned by EPs had their median LOSs shortened by 11% or 32 minutes (p = 0.02; 95% CI = 5 min to 55 min). When evaluated by time of day, patients who presented after hours (6 PM-6 AM) and were scanned by EPs spent 15% (52 min) less time in the ED (p = 0.0002; 95% CI = 26 min to 89 min). Those who were seen after hours and discharged home had their LOSs shortened by 20% (1 hr, 13 min, p = 0.001; 95% CI = 28 min to 1 hr, 56 min). CONCLUSIONS: In a teaching hospital with a residency program, ESUEs decrease ED LOS for these patients. The difference was most apparent for patients presenting after hours. PMID- 10530661 TI - The utility of a dilatation and evacuation procedure in patients with symptoms suggestive of ectopic pregnancy and indeterminate transvaginal ultrasonography. AB - OBJECTIVES: To ascertain the overall frequency of obtaining chorionic villi (CV) in patients with indeterminate transvaginal ultrasound (US) examinations who have had a dilatation and evacuation (D+E) procedure performed, to determine whether the frequency of obtaining CV is dependent on whether the endometrial cavity is empty at US, and to determine the likelihood of ectopic pregnancy in patients without CV after D+E and with or without an empty endometrial cavity at US. METHODS: A retrospective review was made of consecutive ED patients presenting to an urban teaching hospital from August 1991 through August 1997 with abdominal pain or vaginal bleeding and a positive beta-human chorionic gonatropin (beta hCG) test. Patients who had a transvaginal US that was read as indeterminate (no extrauterine findings of ectopic pregnancy, and no intrauterine fetal pole or yolk sac) and who had a D+E performed within 48 hours of the ED visit were eligible. US exams were subdivided into two groups, those with empty endometrial cavities and those with endometrial cavities that contained fluid, echogenic material, or sac-like structures. The presence or absence of CV was based on the official pathology report. Patients were excluded if pathology results were not available. RESULTS: A total of 255 patients met eligibility criteria. Of these, pathology results were not available for five patients. Of the remaining patients, 177 of 250 (70.8%: 95% CI = 64.7% to 76.3%) had CV identified in the pathology specimen. The difference in the frequencies of obtaining CV in those with empty endometrial cavities (35/78; 44.9%: 95% CI = 34% to 56%) vs. those without empty endometrial cavities (142/172; 82.6%: 95% CI = 76% to 88%) was significant (p<0.001). Ectopic pregnancy was diagnosed in 17 of 42 (40.0%) with empty uteri at US and no CV at pathology vs 5 of 26 (19.2%) in whom the uterus was not empty and no CV were obtained (p = 0.07). CONCLUSION: In symptomatic patients with indeterminate transvaginal ultrasound exams, CV will be identified after D+E in approximately 70% of cases. Although CV were found with increased frequency when the endometrial cavity was not empty, still almost half of the patients with empty uteri had villi identified. Finally, although the frequency of ectopic pregnancy was higher in the patients with empty uteri and no CV at D+E, vs. those without an empty uterus and no CV, this difference did not reach statistical significance. PMID- 10530662 TI - Trends in emergency department utilization, 1988-1997. AB - OBJECTIVES: To study changes in ED utilization over a ten-year period; and to try to identify factors that affect utilization. METHODS: This study was conducted in a university-affiliated rural tertiary referral center in a stage 1 managed care market, providing primary emergency services to a county of 120,000 and tertiary services to a 29-county area with 1.2 million people. The year of visit, time of visit, level of care required, length of stay (LOS), and admission status were entered into a computer database for each ED visit. RESULTS: Over the period from 1988 to 1997, the population grew by 18.7%. Over the same time period, the number of ED visits grew 27%. By regression analysis, the number of ED visits was directly related to the size of the service population (correlation coefficient 0.97). During the study period, patient acuity increased, with urgent visits increasing from 45% to 52% while nonurgent visits declined from 55% to 48%. Percentage of patients admitted increased from 14% in 1989 to 20% in 1997. Percentage of patients with LOS exceeding six hours also increased, from 8% in 1989 to 16% in 1997. CONCLUSIONS: For the study hospital there was a direct relationship between the ED utilization and population size as well as a historical trend toward increased patient acuity. These trends quantified at one hospital may reflect trends occurring throughout the United States that would affect ED staffing, space, and resource needs. PMID- 10530663 TI - Occupational exposures to blood among emergency medicine residents. AB - OBJECTIVES: To investigate the epidemiologic characteristics of potentially infectious occupational exposures to blood among emergency medicine (EM) residents. METHODS: A SAEM-sponsored multiple-choice survey was administered anonymously to all EM residents participating in the 1998 American Board of Emergency Medicine in-service examination. Survey questions included resident demographics, use of universal precautions, frequency and types of exposures to blood, and exposure reporting. Residents who experienced at least one exposure were then asked to complete an additional set of questions referring only to their latest exposure. Mean values were calculated for each variable and differences between groups were compared by chi-square analysis. RESULTS: Three thousand one hundred sixty-two surveys were distributed to the resident participants, and 2,985 surveys (94.4%) were returned. Of the participants, 56.1% reported at least one exposure to blood during their EM training. The frequency of this self-reported exposure increased with advancing EM level of training (43% EM-1, 58% EM-2, 64% EM-3, 76% EM-4, p<0.001). Of these residents, 36.6% always followed universal precautions, 54% frequently, and 9.4% sometimes, rarely, or never. Those individuals who "always" followed universal precautions reported significantly fewer exposures than those who did not (p<0.005). The latest exposures were most commonly caused by a solid needle or sharp object (39.4%), by a hollow-bore needle (30.6%), or by eye splashes (17.2%). Of these exposures, 71.7% occurred in the ED setting, and only 46.7% of these exposures were reported to health care providers. CONCLUSION: Emergency medicine residents are frequently exposed to blood, most commonly due to puncture injuries by sharp objects. The rate of exposure reporting is low, which may compromise appropriate postexposure counseling and prophylaxis. PMID- 10530664 TI - Tuberculosis exposure risk in emergency medicine residents. AB - OBJECTIVES: To assess purified protein derivative (PPD) test surveillance and respiratory protection practices of emergency medicine (EM) residents, along with the prevalence of PPD test conversion and the development of active tuberculosis (TB) in EM residents. METHODS: The study instrument was an anonymous, self reporting, multiple-choice survey administered to U.S. and Canadian EM residents. It was distributed for voluntary completion in conjunction with the American Board of Emergency Medicine's annual in-service examination, which was administered February 25, 1998. RESULTS: A total of 89.3% (n = 2,985) of residents eligible to complete the survey completed at least part of it. The majority of residents are PPD-tested once a year. The prevalence of PPD test conversions in EM residents was between 1.4% (36/2,575) and 2.0% (52/2,575). Of the residents who PPD test-converted, the ED was most often the perceived area of TB source exposure (n = 15). Two residents (0.08%) reported having developed active TB, including chest radiographic findings or clinical infection, which equals a 0.14% (95% CI = 0.005 to 0.31) risk of developing active TB over a three year residency. Half of all the residents do not routinely wear National Institute for Occupational Safety and Health (NIOSH)-approved particulate filtration respirator (PFR) masks in patient encounters at risk for TB exposure. While more than a third of EM residents have not undergone fit testing for a NIOSH-approved PFR mask, the lack of routine easy availability of such masks is the most common reason they are not routinely worn by EM residents during at-risk encounters for TB transmission. CONCLUSIONS: Most surveillance PPD testing of EM residents is performed at intervals recommended by the CDC. TB control programs at institutions sponsoring EM residencies need to improve both compliance with PFR mask fit testing by EM residents and availability of approved PFR masks in appropriate areas of the ED. Despite poor compliance with personal respiratory protection in ED patient encounters at risk for TB transmission, the risk of an EM resident's developing active TB over a three-year residency is low. PMID- 10530665 TI - The occupational risk of motor vehicle collisions for emergency medicine residents. AB - OBJECTIVE: To determine the prevalence and risk factors associated with motor vehicle collisions (MVCs) and near-crashes as reported by emergency medicine (EM) residents following various ED shifts. METHODS: A survey was sent to all allopathic EM-2-EM-4 residents in May 1996 asking whether they had ever been involved in an MVC or near-crash while driving home after an ED shift. The residents' night shift schedules, self-reported tolerance of night work, ability to overcome drowsiness, sleep flexibility, and morningness/eveningness tendencies also were collected. RESULTS: Seventy-eight programs participated and 62% of 1,554 eligible residents returned usable surveys. Seventy-six (8%, 95% CI = 6% to 10%) residents reported having 96 crashes and 553 (58%, 95% CI = 55% to 61%) residents reported being involved in 1,446 near-crashes. Nearly three fourths of the MVCs and 80% of the near-crashes followed the night shift. Stepwise logistic regression of all variables demonstrated a cumulative association (R = 0.19, p = 0.0004) that accounted for 4% of the observed variability in MVCs and near crashes. Univariate analysis showed that MVCs and near-crashes were inversely related to residents' shiftwork tolerance (p = 0.019) and positively related to the number of night shifts worked per month (p = 0.035). CONCLUSIONS: Residents reported being involved in a higher number of MVCs and near-crashes while driving home after a night shift compared with other shifts. Driving home after a night shift appears to be a significant occupational risk for EM residents. PMID- 10530666 TI - Why don't emergency department patients have advance directives? AB - OBJECTIVES: In 1997 the authors determined that only 27% of their adult ED patients had advance directives (ADs). The purpose of this follow-up study was to determine the reasons why their adult ED patients do not have ADs. METHODS: This prospective study enrolled patients from a convenience sample of representative shifts in the ED selected over a three-month period. Survey questions included demographic information, whether the patients had a life-threatening medical problem, whether they had an AD, with whom they had discussed their ADs, and the reasons why they did not have an AD. We excluded those who refused participation or who were incapacitated (i.e., any patient with a condition that precluded him or her from answering the questionnaire himself or herself, such as an altered level of consciousness, dementia, mental retardation, or inability to understand English). RESULTS: Four hundred seventy-six subjects were enrolled during the study period from an ED census of 816 adult patients. Three hundred forty patients were not included in the study for the following reasons: inability to complete the survey, refusal to participate, or not being approached by the interviewers. Of those enrolled, 77% of the patients did not have an AD (females, 73%; males, 80%). The most frequent reasons given for not having an AD were: 40% never thought about it, 24% preferred family to make the decision, and 23% were procrastinating. Factors jointly predictive of having an AD were older age, having a specialist, having a life-threatening medical problem, and not being Catholic. Patients who had ADs were discussing their ADs with their primary care physicians (PCPs) only 5% of the time. CONCLUSION: Many patients, even when they have life-threatening medical problems, do not have an AD, and several reasons for this have been identified. Few of these ED patients who had ADs had discussed them with their physicians. Further studies should assess whether more physician intervention would increase the percentage of patients who have ADs. PMID- 10530667 TI - Perceptions of pediatric emergency medicine fellows and program directors about research education. AB - Pediatric emergency medicine (PEM) fellows who entered training after January 1995 are required to complete three years of fellowship training. Additionally, they are required to receive instruction in related basic sciences and to demonstrate research competence. OBJECTIVES: To determine: 1) whether PEM fellows and program directors perceive their programs as providing adequate training in research principles, 2) the manner in which these principles are taught, and 3) the commitment of fellows and program directors to research and research training. METHODS: Pediatric emergency medicine fellows who participated in the Fourth Annual PEM Fellows Conference (Miami, Florida, March 1997) were surveyed. The survey was then extended via mail to all PEM fellows and program directors in the United States and Canada. RESULTS: A total of 159 of the 220 fellows (72%) in the United States and Canada returned completed questionnaires. Fifty-three of 70 PEM fellows (76%) who attended the conference completed questionnaires, and 106 of 167 fellows (63%) who did not attend the conference responded by mail. Fifty three of the 63 program directors (84%) returned completed questionnaires. Of 159 responding fellows, 86 (54%), and of 53 responding program directors, 29 (58%) reported that their programs lacked adequate training in one or more of the surveyed research areas. Thirteen program directors (25%) reported no formal research training in their curricula. Programs that included formal research training were perceived to have higher overall quality than programs that failed to offer such formal training. Sixty-six of 158 responding fellows (42%) anticipated an ongoing commitment to research in their careers. One hundred fourteen of 153 responding fellows (75%) indicated that, if given the option, they would have pursued a two-year "clinical track" PEM board certification that did not include a research requirement. CONCLUSIONS: More than half of surveyed PEM fellows and program directors perceived important deficiencies in research education within their training programs. Further research is necessary in order to evaluate the validity of these perceptions. PMID- 10530668 TI - SAEM ethical guidelines for academic emergency physicians who provide medical malpractice consultation. SAEM Ethics Committee. Society for Academic Emergency Medicine. PMID- 10530669 TI - SAEM emergency medicine research fellowship guidelines. SAEM Research Committee. Society for Academic Emergency Medicine. PMID- 10530670 TI - SAEM emergency medical services fellowship guidelines. SAEM EMS Task Force. Society for Academic Emergency Medicine. PMID- 10530671 TI - Temazepam overdose associated with bullous eruptions. PMID- 10530672 TI - Perimortem cesarean section of twin pregnancy: case report and review of the literature. PMID- 10530673 TI - Metabolic acidosis in restraint-associated cardiac arrest. PMID- 10530674 TI - Cardiorespiratory consequences of the hobble restraint. PMID- 10530675 TI - Clinical experience with new antiepileptic drugs: antiepileptic drugs in Europe. AB - The use of antiepileptic drugs (AEDs), singly and in combination, has been marked by variation among European countries and by a slow progress toward a standard of care that is still far from uniform. Phenobarbital, phenytoin, trimethadione, and primidone, given in various combinations, were the predominant agents used in the first half of this century. Prescribing habits differed among the Latin countries, the United Kingdom, and Scandinavia, based on local trends, divergent teaching philosophies of medical schools, and the medical specialty of the prescribing physician. The advent of carbamazepine and valproate, in the early 1960s, changed European prescribing habits. Despite early fears regarding bone marrow toxicity, carbamazepine was found to be superior for treatment of complex partial seizures. Valproate, when the proper therapeutic dosage was belatedly realized, was seen as a superior treatment for generalized and partial epilepsies. Both agents are now considered first-line treatments for these seizure types. The role of the benzodiazepines as adjunctive anticonvulsant therapy remains controversial because of concerns about neurotoxicity and patient tolerance. The number of AEDs marketed in Europe has grown dramatically in the past decade, with agents such as felbamate, gabapentin, lamotrigine, and tiagabine having been approved as either adjunctive or sole therapy. However, not all new agents are available in each European country, and some variation in prescribing persists. PMID- 10530676 TI - Options after the first antiepileptic drug has failed. AB - Long-term antiepileptic drug (AED) treatment is the standard therapy for epilepsies. In about 60% of patients, the first AED tried usually leads to seizure control. After failure of the first AED, it is important to achieve seizure control rapidly and without side effects. Combining the first drug with an add-on drug appears to be more effective than a second monotherapy. However, no scientifically based data are available that favor any particular drug combination. Along with pharmacokinetic considerations, clinical experience is an important determinant in choosing a second AED for use as an add-on. The time needed to introduce a particular AED is a further consideration. The simultaneous introduction of two add-on drugs, one requiring slow titration and one permitting rapid introduction, may be a useful strategy. If the quickly introduced drug is effective as an add-on, introduction of a slowly titrated second add-on can be obviated. If the quickly introduced drug reduces seizure frequency, the patient's quality of life is improved during titration of the second add-on. If the second, slowly titrated drug is more effective than the quickly introduced one, the less effective drug can be withdrawn. This strategy also allows a direct comparison between two add-on drugs at the same time. PMID- 10530677 TI - Monotherapy trials with gabapentin for partial epilepsy. AB - The efficacy and safety of gabapentin as monotherapy for treatment of partial onset seizures were evaluated in three large multicenter, double-blind, parallel group, dose-controlled trials. In the first trial, 275 outpatients with refractory partial epilepsy maintained on stable doses of one or two antiepileptic drugs (AEDs) were switched to gabapentin (GBP) monotherapy at 600 mg, 1200 mg, or 2400 mg daily. Patients were required to exit the 26-week double blind phase of the study if they experienced worsening of seizure frequency. With respect to time to exit, there was no statistically significant difference among the three groups; only 3% of patients withdrew from the trial because of adverse events. In the second study, 82 hospitalized patients with medically refractory epilepsy were tapered off baseline AEDs and randomly assigned to GBP monotherapy at 300 mg/day or 3600 mg/day. Patients remained in the trial for a maximum of 8 days but had to exit the trial if they experienced one or more exit events. Time to exit was significantly longer in patients in the 3600-mg group (151 h) compared with those in the 300-mg group (85 h) (p = 0.0001). None of the patients withdrew from the trial because of side effects. In the third study, 292 patients with newly diagnosed partial seizures were randomized to GBP 300, 900, or 1800 mg/day or to carbamazepine (CBZ) 600 mg/day. Patients remained in the trial for up to 6 months or until they experienced an exit event. Mean time to exit was significantly longer for patients who received GBP 900 mg/day (p = 0.02) or 1800 mg/day (p = 0.04) compared with those who received 300 mg/day. The completion rate for the CBZ group (37%) was similar to that of the GBP 900-mg (39%) and 1800 mg (38%) groups. Patients receiving CBZ had a higher withdrawal rate because of adverse events compared with the GBP 900-mg and 1800-mg groups. The results of these trials provide good evidence of the efficacy and safety of GBP as monotherapy for the treatment of partial-onset seizures. PMID- 10530678 TI - Monostars: an aid to choosing an antiepileptic drug as monotherapy. AB - This article describes a flexible, dynamic system for comparing antiepileptic drugs (AEDs) as monotherapy, taking into account the needs of the patient and the characteristics of the treatment. Because differences in efficacy between AEDs cannot readily be demonstrated in regulatory clinical trials, safety is of paramount importance. Each drug has been judged across 11 criteria. These include knowledge of mechanism of action, suitable pharmacokinetics, drug interactions, delineated range of efficacy, ease of titration, idiosyncratic reactions, sedative burden, neuropsychiatric profile, teratogenic potential, and the likelihood of producing long-term side effects. The final consideration relates to how "comfortable" the doctor is with prescribing the drug as monotherapy. Scores of -1 (drawback), 0 (neutral/unknown), or +1 (advantage) have been allocated under each category, depending on current knowledge and clinical experience. The sum of the individual scores determines the awarding of "stars." In addition, the positive and negative features of each AED, when used as monotherapy, are highlighted. A range of established and new AEDs has been examined using the "monostars" method, including phenobarbital, phenytoin, carbamazepine, sodium valproate, lamotrigine, gabapentin, oxcarbazepine, and vigabatrin. Scores can be adjusted as new information comes to light. Other agents can be added when suitable monotherapy data become available. This analysis supports the contention that choice of treatment in newly diagnosed epilepsy should take into consideration the patient's age, sex, general health, coexisting disabilities, concomitant medication, and life style. Seizure type, syndrome, and the AED's pharmacology, efficacy, and safety profile should also be considered. Because dosing is often modest, cost should rarely be the overriding factor in choosing a drug for a patient with newly diagnosed epilepsy in the developed world. With these criteria, some of the newer AEDs have potentially more to offer the patient as monotherapy than do the established agents. PMID- 10530679 TI - Dosing to efficacy with neurontin: the STEPS trial. Study of Titration to Effect Profile of Safety. AB - The STEPS (Study of Titration to Effect Profile of Safety) trial, a 16-week, open label, postmarketing multicenter study, assessed the efficacy of gabapentin as adjunctive therapy in patients with inadequately controlled partial seizures. Inclusion criteria were less restrictive than for Phase III studies, to include a population more representative of the patient population treated in clinical practice. Gabapentin was titrated up to a maximal dosage of 3600 mg/day to achieve seizure control or to tolerability. The efficacy analysis included compliant patients who had completed approximately 16 weeks of therapy (n = 1055); 573 received < or = 1800 mg/day and 482 received > or = 1800 mg/day. The average decrease in seizure frequency was 61%, the percentage of seizure-free patients was 46.35%, and the percentage of patients with a > or = 50% decrease in seizure frequency was 76.05%. The cumulative percentage of responders and of patients who were seizure free increased with each dosage increase. The results confirm that titration to efficacy is appropriate for adjunctive therapy with gabapentin in patients with partial epilepsy treated in clinical practice. PMID- 10530680 TI - Gabapentin as adjunctive therapy for partial seizures. AB - The safety, tolerability, and efficacy of gabapentin as adjunctive therapy were assessed in epileptic patients who had experienced up to four complex partial seizures per month while receiving phenytoin and/or carbamazepine. This was a multicenter, open-label prospective study, with the treatment period lasting 20 weeks. The gabapentin dosage was titrated to effective tolerated dose up to 2400 mg/day. Quality of life was evaluated with the QOLIE-10 questionnaire. A total of 141 patients were enrolled; 114 patients were evaluated for efficacy analysis. The mean maintenance dose of gabapentin was 1600 mg/day. Seventy-one percent of patients (81 patients) experienced a 50% or greater reduction in seizure frequency and 46% (52 patients) became seizure free. The most frequent adverse effects included drowsiness (16%), dizziness (9%), and asthenia (6%). Sixteen patients (11%) discontinued the study prematurely because of adverse events. A significant improvement was observed in five of the 10 questions of the QOLIE-10. PMID- 10530681 TI - Use of new antiepileptic drugs in the treatment of childhood epilepsy. AB - The management of epilepsy in children requires careful evaluation, classification, and pharmacologic treatment. With classic antiepileptic drugs (AEDs), at least 25% of children remain refractory to appropriate therapy. The past decade has allowed the introduction of a number of newer AEDs for treatment of both adults and children with epilepsy. These include felbamate, gabapentin, lamotrigine, topiramate, tiagabine, and vigabatrin. Emerging information regarding the efficacy of these AEDs in treating childhood epilepsy syndromes suggests advantages for many patients. Limited data are available that define the optimal use of new AEDs in pediatric patients. Further research must be completed to validate the positive effects described in existing clinical trials of the new AEDs in the treatment of childhood epilepsy. PMID- 10530682 TI - Gabapentin in the management of convulsive disorders. AB - Gabapentin, in clinical use since 1993, is indicated as an adjunctive antiepileptic drug (AED) for treatment of complex partial seizures, with or without secondary generalization, in patients over 12 years of age. Although several cellular actions have been described in the literature, the molecular mechanism(s) of action responsible for the anticonvulsant effect of gabapentin has not been conclusively determined. It is likely that gabapentin has multiple concentration-dependent actions that combine in a unique manner to produce antiepileptic efficacy. The pharmacokinetic properties of this water-soluble, amino-acid AED are generally favorable. Absorption appears to be dependent on transport by the L-system amino acid transporter. Elimination of unmetabolized drug occurs by the renal route. Although its therapeutic range is not well characterized, gabapentin has a broad therapeutic index. This implies that a wide range of doses can be used, based on individual patient needs, without significant limitation due to dose-dependent side effects. Gabapentin has few drug-drug interactions, none of which is clinically limiting. Several studies have demonstrated the long-term efficacy of gabapentin with no systematic evidence of tachyphylaxis. In addition, there is increasing evidence to support the use of gabapentin as monotherapy. Gabapentin is safe and is generally well tolerated. To date, nearly 3 million patients have been treated in studies and in open use without causal relationship to a specific life-threatening organ toxicity. Seizure control superior to that observed in well-controlled trials has been reported at higher doses used in clinical practice and in studies. Therefore, gabapentin dosing must be optimized on an individual basis to achieve an adequate trial of the drug and obtain the best seizure control. PMID- 10530683 TI - Postherpetic neuralgia: role of gabapentin and other treatment modalities. AB - Postherpetic neuralgia (PHN) is a chronic and painful condition that may occur after a herpes zoster infection. The frequency of PHN after untreated zoster varies widely. Age is the most important risk factor for development of PHN. The condition occurs in an estimated 50% of patients older than 50 years. The pain of PHN can be severe and debilitating and is frequently associated with allodynia. Although in most patients pain remits within the first year, it may persist for a lifetime. Tricyclic antidepressants (TCAs), topical agents, opioids, and gabapentin, a structural gamma-amino butyric acid (GABA) analogue, are the only agents that have demonstrated efficacy in randomized clinical trials for treatment of both the shooting and the burning form of pain associated with PHN. TCAs are among the most commonly used classes of agents for treating PHN and are effective in a significant proportion of patients. However, various adverse events can limit treatment. These side effects tend to be more acute in the elderly, the population most likely to suffer from PHN. Topical agents have led to mild to moderate improvement in patients with PHN but are usually ineffective as monotherapy for this condition. Until recently, carbamazepine was the only antiepileptic drug evaluated for the treatment of PHN. Over the past few years, however, gabapentin has received increasing attention as a useful treatment for neuropathic pain. Gabapentin lacks significant drug-drug interactions and has a favorable safety profile, which makes it particularly useful for treatment of PHN. PMID- 10530684 TI - Gabapentin monotherapy for the symptomatic treatment of painful neuropathy: a multicenter, double-blind, placebo-controlled trial in patients with diabetes mellitus. AB - Pain is the most disturbing symptom of diabetic neuropathy. Traditionally this type of pain was treated with tricyclic antidepressants which frequently have many side effects. In the study reported here, gabapentin was administered in escalating doses up to 3600 mg per day to eligible patients with moderate to severe diabetic neuropathy pain in a double blind placebo controlled fashion. Gabapentin provided superior and significant pain relief over placebo. In addition, patients taking gabapentin had improvement of sleep scores and a number of items on mood and quality of life questionnaires. Gabapentin was tolerated well with mild and tolerable side effects. PMID- 10530685 TI - Social phobia: issues in assessment and management. AB - Social phobia was initially classified with phobic anxiety states and was believed to be quite rare, but it is now gaining due recognition as a widespread and often crippling disorder. The boundaries of social phobia merge into traits of shyness and universal performance anxiety, with symptoms commonly appearing in the teenage years. If left untreated, social phobia is a remarkably persistent condition, leading to potentially lifelong impairment in social development and occupational functioning. It may also give rise to other co-morbid disorders, particularly dysthymia, depression, obsessive-compulsive disorder, other phobic disorders, and substance abuse. Over the years, social phobia has been all too frequently viewed as a somewhat trivial, minor form of psychiatric illness and has received little clinical attention. This erroneous perception is now giving way under the mounting evidence in support of the extensive morbidity and disability associated with social phobia and the probable role of genetic and environmental influences. Furthermore, data from multiple controlled clinical trials reveal that this is a treatable condition, responding to both psychosocial and pharmacologic interventions. Here we examine issues to consider in the differential diagnosis of social phobia, review the goals of treatment, and summarize evidence in support of the effectiveness of individual pharmacologic treatments. PMID- 10530687 TI - New antiepileptic drugs: basic science and clinical use in children and adults PMID- 10530686 TI - Nonepileptic uses of gabapentin. AB - For decades, antiepileptic drugs (AEDs) have been used to treat a variety of nonepileptic conditions such as chronic pain, psychiatric disorders, and movement disorders. As indicated by recent published reports, gabapentin, a relatively new AED, is useful for treating a wide range of neurologic and psychiatric conditions. Although its exact mechanism of action has yet to be determined, gabapentin is likely to have multiple effects. Unlike conventional AEDs used to treat nonepileptic disorders (e.g., carbamazepine, phenytoin, valproate) gabapentin offers the advantages of low toxicity and a favorable side-effect profile. The largest area of nonepileptic use of gabapentin is neuropathic pain, in which it has demonstrated efficacy in treatment of postherpetic neuralgia, diabetic neuropathy, and trigeminal neuralgia. It has also been reported effective as therapy for several psychiatric disorders, most notably bipolar disorder. In addition, review of the published literature reveals the usefulness of gabapentin in movement disorders, migraine prophylaxis, and cocaine dependence. Future clinical studies will provide further insight into the range of conditions for which gabapentin is effective. PMID- 10530688 TI - Comparative anticonvulsant and mechanistic profile of the established and newer antiepileptic drugs. AB - Since 1993, several new antiepileptic drugs (AEDs) have been introduced for management of partial seizures. Like the established AEDs, the new drugs are believed to exert their anticonvulsant action through enhancement of inhibitory mediated neurotransmission, or reduction of excitatory-mediated neurotransmission, or by a combination of both. Among the new drugs, vigabatrin (VGB) and tiagabine (TGB) are unique in that they were derived from mechanistic based drug discovery programs designed to identify effective AEDs that inhibit the metabolism and reuptake of the inhibitory neurotransmitter GABA, respectively. For many of the newer AEDs, several molecular mechanisms of action have been identified. For example, felbamate (FBM), lamotrigine (LTG), zonisamide (ZNS), topiramate (TPM), oxcarbazepine (OCBZ), and possibly gabapentin (GBP) share a similar mechanism with that defined for phenytoin (PHT) and carbamazepine (CBZ), i.e., a voltage- and use-dependent block of voltage-sensitive sodium (Na+) channels. In addition to their effects on Na+ currents, TPM, ZNS, and FBM also appear to act as allosteric modulators of the GABA(A) receptor, whereas GBP appears to increase brain GABA levels. GBP, ZNS, FBM, LTG, and OCBZ attenuate voltage-sensitive calcium (Ca2+) channels, albeit through different mechanisms and with different classes of Ca2+ channels. FBM and TPM differ from both the established and newer AEDs in their ability to modulate NMDA- and AMPA/kainate mediated excitatory neurotransmission, respectively. The multiple mechanisms of action associated with FBM, TPM, ZNS, GBP, and perhaps LTG, and the unique modulation of GABA levels by VGB and TGB, are likely to account for the anticonvulsant efficacy of these newer AEDs in patients with epilepsy. For each of the new drugs, their proposed mechanisms of action are discussed in relationship to their preclinical and clinical anticonvulsant profiles. PMID- 10530689 TI - Vigabatrin. AB - Vigabatrin (VGB) is a structural analogue of the inhibitory neurotransmitter gamma-amino butyric acid (GABA), which produces its antiepileptic effect by irreversibly inhibiting the degradative enzyme GABA-transaminase. This produces an increase in central nervous system (CNS) GABA levels. VGB is among the few antiepileptic drugs (AEDs) that was synthesized with a specific targeted mechanism in mind and was subsequently demonstrated to function by that mechanism. Tiagabine, a GABA reuptake blocker, is the only other "designer drug" among the currently available AEDs. Therefore, VGB is among the few AEDs for which the mechanism of action is well understood. Recently, safety issues have been raised with regard to the use of vigabatrin. This article reviews the mechanism of action, pharmacokinetics, safety, and efficacy of VGB. PMID- 10530690 TI - Tiagabine. AB - Tiagabine (TGB) is a recently approved antiepileptic drug (AED) that inhibits y aminobutyric acid (GABA) reuptake into neurons and glia, a mechanism of action that is specific and unique among the AEDs. TGB is potent and has linear and predictable pharmacokinetics. It has no clinically relevant effects on hepatic metabolism or serum concentrations of other AEDs, effects on laboratory values, or interactions with common non-AEDs. TGB is effective as add-on therapy for partial seizures in patients with medically refractory epilepsy in doses ranging from 30 to 56 mg daily. Conversion to TGB monotherapy can be achieved in patients with medically refractory epilepsy, although additional controlled studies are needed to confirm the efficacy of TGB as monotherapy and to establish the effective dosage range. In controlled studies, the most common adverse events of TGB are dizziness, asthenia, somnolence, accidental injury, infection, headache, nausea, and nervousness. These are usually mild to moderate in severity and almost always resolve without medical intervention. PMID- 10530691 TI - Zonisamide. AB - Zonisamide (ZNS) is a broad-spectrum antiepileptic drug in both animal models of epilepsy and patients with epilepsy. It is effective for both localization related and generalized epilepsies and appears to be particularly potent in progressive myoclonic epilepsy syndromes. Its pharmokinetic profile is favorable, with a long half-life and low protein binding. However, its insolubility may make the development of a parenteral formulation difficult. Its safety profile is good, although teratogenicity in animal models is of concern. Adult doses of 400 600 mg per day in two doses, with blood levels from 20 to 30 mg/ml, appear to be effective. PMID- 10530692 TI - Lamotrigine. AB - Clinical use of the antiepileptic drug (AED) lamotrigine (LTG) has dramatically increased since its introduction in Europe in 1991 and in the United States in 1994. This article surveys the English-language literature of LTG published before 1998. This literature is concerned with the molecular mechanisms of LTG's antiepileptic action, evaluation of its clinical antiepileptic efficacy, adverse experiences associated with its clinical use, and current guidelines for its initiation. LTG's efficacy has been extensively confirmed in multiple postmarketing studies, and its applications are broad. The most serious adverse experiences have involved skin rash. Valproic acid affects LTG metabolism, and a specific set of guidelines for the concurrent use of valproic acid and LTG has been developed. Unique issues are also associated with its pediatric use. LTG has a significant place in clinical management of a wide range of epilepsy syndromes, and the scope of its use is expanding. Accumulating clinical data enable the clinician to maximize its efficacy and minimize adverse experiences. Guidelines for its pediatric use must be followed diligently. PMID- 10530693 TI - Oxcarbazepine. AB - The success of carbamazepine (CBZ) as a broad-spectrum antiepileptic drug (AED) has led to its use as first-line therapy in children and adults for partial and generalized tonic-clonic seizures. The limitations of CBZ include toxicity in sensitive individuals, autoinduction, which requires dose adjustment when therapy is initiated, and chronic hepatic induction, producing drug interactions when CBZ is used with AEDs and other drugs that undergo hepatic metabolism. One of two main products of CBZ microsomal metabolism, CBZ-10,11-epoxide (formed by oxidation of the double bond between C-10 and C-11), appears to provide antiepileptic efficacy but contributes significantly to clinical toxicity. The most common adverse effects of CBZ are central nervous system (CNS) symptoms, followed by gastrointestinal, hepatic, endocrine disturbances, and teratogenic effects. Oxcarbazepine (OXC) was developed to provide a compound chemically similar enough to CBZ to mimic its efficacy and overall safety while improving its side-effect profile. Biotransformation of OXC does not involve formation of an epoxide metabolite. Compared with the parent compound, hepatic microsomal enzyme induction and autoinduction are greatly reduced. The clinical efficacy of OXC compares favorably with CBZ in clinical trials. Clinical development of OXC began in Europe. Results of Phase I trials started to appear in the early 1980s. Controlled clinical trials, reported in the mid- to late 1980s, led to approval of OXC in many European countries, and now in over 50 nations around the world. United States multicenter clinical trials have recently been completed, and at this writing the drug is awaiting approval by the FDA. This article reviews the pharmacology, animal data, outcomes of published controlled clinical trials, postmarketing data, adverse experiences, and current recommendations for clinical use of OXC. PMID- 10530694 TI - Evidence-based medicine and antiepileptic drugs. AB - Evidence based health care uses systematic literature reviews with statistical strategies like meta-analysis to aid decision-making. This information can help clinicians by organizing data and providing up-to-date quantitative summaries of efficacy and adverse effects of treatments. Limitations of meta-analysis include problems inherent in combining data from trials of somewhat different design, choice of appropriate dosages, and summarizing complex questions as a single odds ratios. I summarize the results of a meta-analysis of the following antiepileptic treatments for partial seizures in adults: gabapentin, lamotrigine, topiramate, tiagabine, valproate and the vagal nerve stimulator. Each treatment was significantly more efficacious than placebo, and there were nonsignificant trends toward differences among the treatments in efficacy and tolerability. Quantitative analysis of adverse effects is presented. Absent the availability of a comprehensive randomized controlled trial for comparison, a rigorously conducted meta-analysis provides some useful information. PMID- 10530695 TI - Felbamate. AB - Felbamate (FBM) was the first of the new antiepileptic drugs (AEDs) approved in the United States in 1993 with broad-spectrum efficacy against partial and generalized seizures of various types, and indicated for use as adjunctive and monotherapy. The identification of idiosyncratic aplastic anemia and hepatotoxicity, however, drastically curtailed its use. To update information concerning FBM and its idiosyncratic effects, case studies and literature reviews were undertaken. Thirty-four FBM-associated aplastic anemia patients have been reported, with 13 known fatalities. The overall FBM aplastic anemia risk is estimated at between 27 and 209 per million vs. 2 to 2.5 per million in the general population. Prior AED hypersensitivity, cytopenia, and immune disease significantly increase risk. FBM aplastic anemia has not been reported in children below the age of 13 years. Hepatic failure is much less common, occurring with an overall risk similar to that associated with valproate, but children below the age of 5 years have been affected. The recent identification of a reactive metabolite, atropaldehyde, and HLA studies suggest that high-risk patients can be identified. The efficacy profile of FBM should encourage further investigations to allow its better use, but at present FBM is not a first-line AED. PMID- 10530696 TI - Gabapentin. AB - Gabapentin (GBP) is a antiepileptic drug (AED) indicated as adjunct therapy for treatment of partial seizures, with and without secondary generalization, in patients 12 and older with epilepsy. GBP (1-(aminomethyl) cyclohexaneacetic acid) is structurally related to gamma-aminobutyric acid (GABA), which readily crosses the blood-brain barrier. Radiolabeled GBP binds throughout the central nervous system in anatomic areas important in treatment of seizures. Its precise mechanism of action is unknown. An open-label, dose-ranging study of doses up to 1,800 mg produced > or =50% seizure reductions [responder rate (RR)] in 29% of patients with partial seizures. Three double-blind, placebo-controlled, parallel add-on trials at doses of 300-1,800 mg have produced RR of up to 28%, with a placebo RR of 8-10%. An active controlled, parallel group comparison of 600 mg to 2,400 mg in monotherapy conversion design showed no significant difference among the 600 mg, 1,200 mg, and 2,400 mg groups compared to a placebo group. An inpatient, active-controlled comparison of 300 mg and 3,600 mg in a parallel design monotherapy trial showed that time to exit from the study was significantly longer for the 3,600-mg group and the completion rate significantly higher (53% vs. 17%) for patients receiving 3,600 mg/day vs. 300 mg/day of GBP. Successful double-blind, placebo-controlled trials in refractory childhood partial seizures and benign childhood epilepsy with centrotemporal spikes have been recently concluded. Absence was not successfully treated in one small double blind trial. Open-label reports emphasize adjustments of patients to higher doses than those indicated in the package labeling. An open-label trial of GBP therapy in patients with partial seizures (n = 2,216) produced progressively greater seizure freedom rates as patients were titrated from > or =900 mg daily to > or = 1,800 mg daily (15.1% vs. 33.4%), with a similar effect on RR (18.1% vs. 44.9%). An add-on, open-label study treating partial seizures (n = 141) reported an RR of 71%, with 46% seizure-free in the last 8 weeks of treatment and doses up to 2,400 mg daily. A comparison trial of three doses of GBP to 600 mg of carbamazepine showed similar retention rates for 1,800 mg of GBP and 600 mg of CBZ. Another study reported 48% of patients experiencing 50% reduction, nine of whom had doses greater than 2,400 mg. Treatment in children has reported a 34.4% RR in 32 children with refractory partial seizures. A French open-label adjunctive trial documented a 33.9% RR; 13.4% were seizure-free during the evaluation period. Adverse experiences most commonly noted included somnolence, dizziness, and ataxia. Weight gain was sometimes reported with higher doses of GBP, and pediatric reports cite prominent behavioral changes, including hyperactivity, irritability, and agitation. GBP appears best used at doses at and potentially above those suggested in its package labeling. Although efficacy occurs at lower levels, increased GBP doses are associated with additional efficacy. Reports suggest that initiation at 2,400 mg or 3,600 mg may not be associated with increased adverse experiences. Titration to 900 or 1,200 mg on the first day of GBP therapy appear to be well tolerated. PMID- 10530697 TI - Topiramate. AB - Six studies are cited to demonstrate that topiramate is effective as adjunctive therapy for refractory partial-onset seizures in adults. Subsequent studies indicate that topiramate is also effective as monotherapy in adults and as adjunctive therapy for partial-onset seizures in children, tonic-clonic seizures of nonfocal origin in children and adults, and drop attacks in Lennox-Gastaut syndrome. Adverse effects for adults and children included dizziness, fatigue, ataxia, confusion, somnolence, nephrolithiasis, paresthesia, and weight loss. More adverse effects were observed at higher doses. Topiramate exhibits rapid absorption, long duration of action, and minimal interaction with other antiepileptic drugs. PMID- 10530698 TI - Pharmacology of antiepileptic drug polypharmacy. AB - Therapeutic options for epilepsy are increasing, and all drugs can produce adverse side effects. Physicians should first use monotherapy when treating an epileptic patient. If one drug proves insufficient to control all seizures without intolerable side effects, then careful combination of the fewest drugs possible must be used. Combine drugs with different mechanisms of action rather than drugs with similar mechanisms of action. Closely monitor the patient for adverse side effects, which increase when multiple drugs are taken, and measure antiepileptic blood levels more frequently than with patients who are on monotherapy. PMID- 10530699 TI - Daily cost of newer glaucoma agents. AB - The purpose of this study was to evaluate the drop characteristics of newer glaucoma medicines compared to timolol solution and timolol gel forming solution (Timoptic-XE, Merck). We evaluated latanoprost 0.005% (2.5 ml bottle), brimonidine 0.2%, apraclonidine 0.5%, dorzolamide 2%, timolol solution 0.5% (5 and 10 ml bottles), and timolol gel forming solution 0.5% (5 ml bottle) in 14 patients with primary open-angle glaucoma or ocular hypertension. Each patient placed 10 drops onto an analytical scale (one drop every 10 seconds) for all ten preparations. Patients then attempted to instill 10 drops of a tear replacement solution into their ocular cul-de-sac. Medication bottles were weighed before and after patients dispensed from the bottle and then after the bottle was emptied. Weights were converted to volume using the density of the medicine. A statistical difference existed between groups for mean drop volume with latanoprost having the smallest drop volume (.0273 +/- .004 ml) (P<0.005). All manufacturers filled correctly or overfilled their bottles with product and had <10% of medicine wasted. Patients instilled 77.9% of the tear solution correctly. When dosed according to labeling, latanoprost had the lowest cost of therapy at $0.87 daily compared to the other newly released medications (range $1.05 to $1.40). Latanoprost was more expensive, however, than timolol maleate solution or gel (range $0.45 to $0.54 per day). Latanoprost therapy is less expensive per day than dorzolamide, brimonidine or apraclonidine, but more expensive than timolol maleate. Cost per day could be further reduced by limiting medicine wastage upon instillation, however. PMID- 10530700 TI - Pituitary adenylate cyclase-activating polypeptide- and VIP-induced activation of adenylate cyclase in the porcine non-pigmented ciliary epithelium: effects of antagonists. AB - Vasoactive intestinal polypeptide (VIP) and two pituitary adenylate cyclase activating polypeptides (PACAP-38 and PACAP-27) were investigated for their ability to activate the adenylate cyclase system in membrane preparations from the porcine non-pigmented ciliary epithelium (NPE). The NPE was dissociated from the adjacent pigmented ciliary epithelium of the iris-ciliary body (ICB) by incubation in low Ca2+ Ringer's solution. All three peptides caused a dose dependent increase in cAMP formation in the NPE and the remaining part of the ICB. A VIP antagonist had a small effect on the dose-response curve for VIP, while the two PACAP fragments, PACAP(6-38) and PACAP(6-3 1), caused a rightward shift of the concentration response curves for PACAP-38, PACAP-27 and VIP. The results of the present study indicate that the non-pigmented ciliary epithelium of the porcine eye contain receptors for PACAP- and VIP-coupled to adenylate cyclase activation. PMID- 10530701 TI - Toxic effects of mitomycin-C on cultured corneal keratocytes and endothelial cells. AB - Improper use of mitomycin-C in ocular medication may result in damage to corneal cells. In this study, the toxic effects of mitomycin-C on cultured porcine keratocytes and endothelial cells were estimated by MTT, 3H-thymidine uptake and cellular counting assay methods. It was found that mitomycin-C caused a dose dependent toxic effect to keratocytes and endothelial cells. Both cells were treated with mitomycin-C at the concentration ranging from 100, 10, 1, 0.1 to 0.01 microg/ml for 3 min, 5 min or 100 min. The 50% inhibitory dose (ID50) of mitomycin-C to keratocytes and endothelial cells as measured by MTT assay was 0.40, 0.18, 0.16 mg/ml and 0.27, 0.15, 0.14 mg/ml, respectively, after 3, 5 and 100 minutes drug treatment. The ID50 for keratocytes and endothelial cells as measured by 3H-thymidine uptake immediately, 1 day and 7 days after 100 minutes mitomycin-C treatment was 0.3, 0.0002, 143.2 microg/ml and 45.1, 101.1, 450.2 microg/ml, respectively. The ID50 for keratocytes and endothelial cells as measured by cellular counting 1 day and 7 days after mitomycin-C treatment was 232.5, 109.7 microg/ml and 239.9, 367.5 microg/ml, respectively. It is concluded that mitomycin-C is more toxic to cellular proliferation in cultured corneal keratocytes than in endothelial cells. PMID- 10530702 TI - Angiostatic activity of steroids in the chick embryo CAM and rabbit cornea models of neovascularization. AB - Ocular neovascular diseases represent a major cause of blindness in the world. Angiostatic steroids are a unique class of compounds which inhibit the formation of new blood vessels in various models, including ocular models of angiogenesis. In search of potent new anti-angiogenic agents for the treatment of ocular neovascular disease, a large group of steroids were evaluated for angiostatic activity in the chick embryo CAM model. Angiostatic activity was found among all steroid classes included in the study. There was a good correlation between the angiostatic efficacies of 15 diverse steroids tested in the chick CAM and in the rabbit LPS-induced corneal pocket models of neovascularization (r=0.76, p=0.01). These studies show that potent angiostatic steroids inhibit neovascularization in two different animal models, suggesting a common mechanism of action. Glucocorticoid therapy is sometimes associated with ocular side effects. Two of the most potent angiostatic steroids, AL-3789 and AL-4940, were evaluated for glucocorticoid-mediated antiinflammatory activity in the in vitro U937 cell model of LPS-induced IL-1 induction and found to be devoid of glucocorticoid activity. Angiostatic steroids which lack glucocorticoid activity should be attractive drug candidates for treating ocular neovascular disease. PMID- 10530703 TI - Corticosteroids as an antiangiogenic agent for histoplasmosis-related subfoveal choroidal neovascularization. AB - The purpose of this study was to evaluate the role of corticosteroids in managing subfoveal choroidal neovascularization (CNV) secondary to the presumed ocular histoplasmosis syndrome. The cases of eighteen patients with histoplasmosis related subfoveal CNV treated with corticosteroids were reviewed. Ten patients received oral prednisone for 4 to 6 weeks, and eight received a single sub Tenon's injection of triamcinalone. Visual acuity outcomes were analyzed along with side effect profiles. At two-week follow-up, the prednisone group showed a median improvement in Snellen visual acuity of +2.0 lines, while the triamcinalone group remained essentially stable with a 0.5 line median loss. At treatment end (4 to 6 weeks), both groups showed no significant change in median acuity at 0.0 and -1.0 lines, respectively. Median final vision at 3 months also remained essentially stable at -0.5 lines for each group. Three patients reported anxiety, all of whom were taking prednisone 80 mg daily. Two patients reported increased appetite and weight gain on regimens of prednisone 80 and 100 mg daily. There were no adverse effects reported in the other patients receiving oral prednisone or in any patient receiving sub-Tenon's triamcinalone. The results suggest a beneficial effect of corticosteroids in stabilizing subfoveal CNV secondary to ocular histoplasmosis. In this small series, oral prednisone resulted in a short-term improvement in visual acuity, which stabilized over longer follow-up. The sub-Tenon's triamcinalone group achieved similar final stabilization without the initial improvement. Corticosteroids may be particularly valuable in managing neovascularization in patients who are awaiting interventions currently under development, in preventing recurrence after subfoveal surgery, or in treating non-surgical candidates. Further study is warranted to define the precise role of corticosteroids in this condition. PMID- 10530704 TI - Influence of cyclodextrins on the in vitro corneal permeability and in vivo ocular distribution of thalidomide. AB - The aim of the present study has been to develop aqueous Thalidomide (THA) eye drops in order to minimize systemic side effects and to improve bioavailability following topical application. Cyclodextrins (CDs), suitable vehicles to improve aqueous solubility of THA, were evaluated with regard to their ability to influence in vitro corneal permeability of THA. Additionally, rabbit eyes received either THA-suspension (0.04%) (THA-SP) or THA (0.04%)/hydroxypropyl-beta cyclodextrin (HP-beta-CD) (12.5%) solution (THA-CD). In vitro corneal permeation studies demonstrated that the absolute amount of THA permeated could not be increased by means of CDs. The percentile release of THA was extensively decreased using saturated THA/CD solutions. Following loading doses of either THA CD or THA-SP onto the rabbit eye, significantly increased aqueous humor levels were obtained for THA-CD 30 min (THA-CD:THA-SP=4.6:1) and 60 min (THA-CD:THA SP=3.1:1) post instillation (p<0.05). In the iris-ciliary body, significantly increased THA levels were obtained using THA suspension (THA-CD(60 min):THA-SP(60 min)=1:6.1) (p<0.05). In the cornea, conjunctiva, vitreous and sclera, differences between the THA tissue levels were not statistically significant. Cyclodextrins might be a useful tool to formulate aqueous THA eye drop solutions and modify intraocular drug bioavailability. PMID- 10530705 TI - Penetration and decay of meropenem into the human aqueous humor and vitreous. AB - The aim of this study was to determine the penetration of intravenously administered meropenem into the human aqueous humor and vitreous. Thirty patients about to undergo cataract surgery and fourteen patients about to undergo vitrectomy received a 2 g dose of meropenem before surgery. Specimens of aqueous humor or vitreous and blood were obtained intraoperatively and analyzed by high performance liquid chromatography (HPLC). The study was designed as a non randomized prospective trial. Thirty min to 12 hr after administration, mean aqueous humor levels of 13.4 and 1.1 mg/l and vitreous levels between 8.94 and 1.08 mg/l were found, respectively. The peak concentrations are distinctly above the in vitro measured minimum inhibitory concentration of meropenem for 90% (MIC90) of almost all relevant gram-positive and gram-negative organisms, including Pseudomonas aeruginosa and Enterobacteriaceae. With regard to its broad spectrum, high antibacterial activity, and good penetration into ocular fluids, meropenem seems to be an alternative to currently used systemic drugs. Its usefulness in perioperative prophylaxis, as initial therapy after perforating or penetrating injuries, or in the therapy of bacterial endophthalmitis has yet to be proved. PMID- 10530706 TI - Involvement of dopamine D1-like receptors in mediating increases in protein secretion from rabbit lacrimal gland. AB - To identify and localize the dopamine receptor subtypes in rabbit lacrimal gland which mediate protein secretion, the effects were determined of different dopamine receptor subtype agonists, antagonists, and a beta adrenergic antagonist on this process. Protein secretion into the medium was quantified with the Bradford assay. Dopamine increased protein secretion between 10(-7) and 10(-4)M, and it could be maintained for a subsequent 80 min. The relatively selective D1 like receptor agonist, SKF-38393 (10(-4)M) had a similar effect which was suppressed by the D1-like receptor antagonist, SCH-23390. However, neither the D2 like receptor agonist, quinpirole (10(-4)M), nor the selective D2-like receptor antagonist, sulpiride (10(-4)M) altered either the basal level or the stimulated response to dopamine. The dopamine (10(-4)M)-elicited increases in protein secretion were completely suppressed in the presence of either 10(-4)M propranolol or 10(-4)M bretylium. Protein secretion in rabbit lacrimal gland is mediated by dopaminergic nerves through stimulation of the presynaptic D1-like receptor. PMID- 10530707 TI - Microdialysis for pharmacokinetic studies of ceftazidime in rabbit vitreous. AB - The method of microdialysis was used for collecting series of samples from the rabbit vitreous after systemic and intravitreous administration of ceftazidime. The purpose of the study was to compare the method with traditional pharmacokinetic sampling. Ceftazidime was injected intramuscularly (1 mg/kg) or intravitreally (1 mg) in rabbits, with a previously implanted microdialysis probe in the vitreous. The membrane was perfused with a buffer, and the dialysate was collected in samples where the concentration of the drug was analyzed by HPLC. After intramuscular administration, blood samples were taken to calculate systemic pharmacokinetics. The same procedures were repeated with rabbits with mild intraocular inflammation induced by the injection of 400 EU of endotoxin into the vitreous, 12-15 hr before drug administration. Pharmacokinetic parameters, such as half-life and AUC, were calculated. The penetration into the vitreous after intramuscular injection was higher (42%) in inflamed than in non inflamed eyes (20%), suggesting an interference with the blood retinal barrier. Other kinetic parameters did not differ significantly between the groups. The advantage of the method is that fewer experimental animals can be used to obtain the necessary data compared to traditional pharmacokinetic methods. In conclusion, intraocular dialysis with chronically implanted probes is a technique well suited for pharmacokinetic studies of systemically administered ceftazidime or other drugs that will pass a dialysis membrane. PMID- 10530709 TI - Introduction: emerging therapies for hematologic malignancies: antibodies, antisense, and cytokine approaches PMID- 10530708 TI - Reducing chorioretinal viral counts with intravitreal foscarnet injections in a rabbit model of Herpes simplex virus type-1 retinitis. AB - The efficacy of intravitreal foscarnet injections was evaluated in a rabbit model of Herpes simplex virus type-1 (HSV-1) retinitis. In untreated infected animals, viral titration revealed that the optic chiasm, vitreous and chorioretina were positive for HSV-1. On the other hand, foscarnet treatment significantly decreased the viral count in the chorioretina when compared to the untreated group. Immunolocalization of HSV in untreated infected animals clearly showed infected cells in the outer and inner layers of the retina and also in the ciliary body of the eye. Clinical examination by indirect ophthalmoscopy indicated an absence of optic nerve congestion and a lower level of vitritis in foscarnet treated animals compared to the untreated group. It is concluded that intravitreal injections of foscarnet reduced the viral titer in the chorioretina in a rabbit model of HSV-1 retinitis. This route of administration might be valuable for the treatment of CMV retinitis in AIDS patients with sight threatening lesions or intolerance to intravenous anti-CMV drugs. PMID- 10530710 TI - Antibody-targeted therapy for myeloid leukemia. AB - The availability of antibodies reactive with antigens expressed only by hematopoietic cells has provided clinical investigators with new tools for use in developing therapies for acute myeloid leukemia (AML). Studies performed to date have investigated the use of such antibodies in an unmodified state, combined with potent chemicals to form immunotoxins or combined with various radionuclides. Encouraging results have been obtained in all three settings. The CD33 antigen is expressed by most early myeloid cells and by more than 90% of cases of AML but is not present on the hematopoietic stem cell. Initial in vivo studies with an unmodified murine anti-CD33 antibody in patients with AML demonstrated that the antibody quickly bound to leukemia cells and that the antigen-antibody complex rapidly internalized following cell binding. However, when administered to patients with overt leukemia, unmodified antibody resulted in only brief decreases in peripheral blast counts, not in sustained response. A number of CD33-based immunotoxins have also been studied, including a calicheamicin conjugate, CMA-676. In a phase I dose-escalation study of patients with refractory AML, CMA-676 was well tolerated with the only consistent toxicities being the development of fevers and chills several hours after administration and the subsequent development of temporary pancytopenia. A phase III study has been performed involving patients with AML in first relapse. An interim analysis of the first 23 patients found that in 10, treatment with CMA 676 resulted in elimination of blasts from peripheral blood and marrow. This was achieved with far less toxicity than seen with standard chemotherapy. Radiolabeled antibodies have been explored as a stand-alone treatment or in the context of bone marrow transplantation. In an effort to avoid toxicities to normal stem cells residing alongside leukemic cells in the marrow, studies have been performed to explore the use of 231Bi conjugated to an anti-CD33 monoclonal antibody. The short path length of this alpha-emitter could theoretically allow killing of the targeted leukemic cell without damage to normal neighbors. Of 12 patients with recurrent AML who received this drug, eight had reductions in marrow and peripheral blast counts. Complete remissions (CRs) have not been observed to date. Another set of studies focused on the use of radiolabeled antibodies to deliver radiation specifically to sites of leukemia as part of a transplant preparative regimen. In a phase I clinical trial, 131I-labeled anti CD45 antibody delivered at least threefold more radiotherapy to spleen and marrow than any other organ. In a phase II trial, among 25 AML patients in first remission, 22 are alive and in remission for periods up to 6 years. PMID- 10530711 TI - Antisense therapy of hematologic malignancies. AB - Many tumor cells are inherently resistant to curative treatment due to an altered pattern of gene expression. It is an attractive and logical proposition to use this difference within the lymphoma cell to eradicate the malignant process. One such new therapeutic approach based on the "silencing" of genes involved in the prevention of apoptosis is Bcl-2 antisense oligonucleotide (AO) therapy. In the field of lymphoma, obvious targets included follicular lymphoma with the t(15;18) translocation, which results in deregulated expression of the Bcl-2 gene, chemoresistance, and subsequent protection against lymphoma cell death. Targeting the initiating codon of the Bcl-2 gene decreases both cell viability and Bcl-2 protein expression in lymphoma and leukemia cell lines that overexpress Bcl-2. Preclinical toxicity studies using a Bcl-2 AO G3139 (Genta, San Diego, CA) show good tolerance at a dose of 10 mg/kg, which is considerably higher than the dose required for good antilymphoma efficacy. In a phase I clinical study, G3139 was well tolerated with minimal toxicity in a dose escalation up to 147.2 mg/m2/d. Evidence of efficacy includes a responder with stage IVB follicular lymphoma who achieved complete clinical and radiologic response that has lasted more than 2 years. The main dose-limiting toxicity has been reversible thrombocytopenia related to the thioate backbone. Other antisense reagents are also in development to combat non-Hodgkin's lymphoma (NHL). These include oligonucleotides that target the messages of the Bcl-X(L) and protein kinase-Calpha (PKCalpha) genes. AOs may also have an application in tumors expressing mutant p53. AOs against MDM2 genes have shown the ability to restore wild-type p53 expression, suggesting that as oncogenic pathways are unraveled, normal cell growth and death patterns may be restored by molecular manipulation. Downregulation of antiapoptosis by AOs in the human setting has low toxicity and antilymphoma activity in cases in which conventional chemotherapy has failed. In the future, antisense therapy followed by chemotherapy may overcome chemoresistance to provide effective therapy for a range of malignancies. PMID- 10530712 TI - Antibody-targeted therapy for low-grade lymphoma. AB - Monoclonal antibodies (MoAbs) have now become a successful treatment for selected patients with non-Hodgkin's lymphoma (NHL). Antibody targets most commonly used for the treatment of B-cell NHL include CD20, CD19, and CD22. Unconjugated MoAbs are cytotoxic by several mechanisms, including complement-dependent cytotoxicity (CDC), antibody-dependent cell-mediated cytotoxicity (ADCC), and signal transduction leading to apoptosis. In an attempt to augment the effectiveness of naked antibody preparations, various radioconjugates, immunotoxins, chemotherapeutic agents, or immune-modifiers have been attached to the antibodies. The immunotoxin tested most extensively in clinical trials is B4 blocked ricin (anti-CD19 with a partially blocked ricin toxin). The use of radioimmunoconjugates to augment the effectiveness of unlabeled antibodies has been one of the most popular strategies. Antibodies against these targets have now been chelated with radioconjugates such as 131I or 90Y and tested in recent clinical trials. Radioimmunotherapy has the theoretical advantage over naked antibody therapy or immunotoxin therapy in that the MoAb conjugated with a radioisotope can have a "cross-fire" effect such that antigen-negative tumor cells adjacent to those expressing the target antigen may also be killed. This may enhance the likelihood of tumor sterilization even in fairly bulky disease. Future studies will focus on testing these antibodies in larger patient populations, sequentially or in combination, and on combining MoAb therapy with standard- or high-dose chemotherapy and hematopoietic stem-cell transplantation. PMID- 10530713 TI - Advances in supportive care of myelodysplastic syndromes. AB - The myelodysplastic syndromes (MDS) are a heterogenous family of hematologic disorders characterized by ineffective hematopoiesis. Because of the variability between patients regarding prognosis and morbidity related to the disease, consensus regarding the management of these patients has been difficult. Over the past several years, new prognostic scoring systems such as the International Prognostic Scoring System (IPSS) have attempted to provide a projection for long term stability of the percentage of patients who have "low-grade" or indolent MDS. Unfortunately, its lack of prospective use in clinical trials and other settings has thus far failed to validate it as a functional decision-making tool. Thus, investigators have hypothesized that separating patients based on more simplistic treatment-oriented guidelines may be more efficient. For the majority of patients with MDS, no curative option exists. Patients who are young enough and have an available matched sibling or matched unrelated donor may undergo an allogeneic bone marrow transplant (BMT) with a potential cure rate of 30% to 50%. The major issue regarding this approach is the relatively high morbidity and the risk that the patient's lives may be shortened, that their quality of life will be worsened, or that no overall benefit will occur (relapse). Compounding the issue of selection and timing for BMT is the fact that the best results in terms of relapse-free survival appear to be in the subset of patients with early or low grade MDS, characterized by refractory anemia with or without ringed sideroblasts. For these patients, lacking a donor for BMT, the major issue has become the consideration of induction chemotherapy. While dose-intensive chemotherapy may improve outcome in a small percentage of patients, the majority of elderly patients with MDS are not optimal candidates for such an approach. As a result, supportive care has a major role for patients with MDS and depending on the French-American-British (FAB) presentation and comorbid illnesses may be the preferred approach. Erythropoietin, a growth factor, is perhaps the most commonly used supportive care after transfusion. The use of colony-stimulating growth factors to support leukopenia is currently under investigation. The use of thrombopoietic agents has lagged behind in the management of MDS patients. Investigation of interleukin-6 (IL-6), a thrombopoietic cytokine, showed some ability to increase platelets through significant toxicity. Investigation of IL 11, an approved thrombopoietic growth factor, is preparing to start and should aid in determining its role in this setting. PMID- 10530714 TI - Introduction: molecular diagnostics in hematology. AB - Advances in molecular biology of the last 30 tears have transformed the field of hematology. Molecular analysis has clarified the pathogenesis of a host of hematologic disorders, including hemoglobinopathies, coagulation disorders, hypercoaguable states, and hematologic malignancies. This volume is aimed at bringing these compelling advances in molecular biology research to the bedside. Through progress in the molecular understanding of hematology, great strides have been made in our ability to diagnose and treat patients with hematologic disease. These advances in turn have given rise to more questions that will guide future studies of the biology of hematopoietic abnormalities and continue the exciting interaction of basic science and clinical medicine. PMID- 10530715 TI - Molecular diagnosis of hemoglobinopathies and other red blood cell disorders. AB - This article provides an overview of the techniques currently available for the molecular diagnosis of hemoglobinopathies and other inherited erythrocyte disorders. Advances in both clinical practice and molecular biology have permitted rapid genetic diagnosis of many of these disorders. In turn, rapid diagnosis has led to improvements in prenatal diagnosis and in early detection of at-risk individuals, permitting appropriate prenatal counseling to at-risk couples, allowing for appropriate patient education, and improving clinical outcome. PMID- 10530716 TI - Molecular diagnosis of inherited bleeding disorders and thrombophilia. AB - Molecular genetic characterization of the hemostatic system began more than 15 years ago, and, as a result, our knowledge of the genetic pathology of inherited bleeding disorders is well advanced. However, molecular testing for hemophilia and von Willebrand disease (vWD) has been impeded by the large size and complex genomic organization of the genes involved, and by the heterogeneity of the mutations underlying these disorders. Such limitations have significantly reduced the ability to provide diagnostic testing for these conditions through direct mutation detection. Many diagnostic laboratories continue to utilize linkage analysis with highly informative intragenic polymorphisms for the investigation of hemophilia. The one important exception to this trend has been the factor VIII inversion mutation, which provides a definitive test for the mutant allele in approximately 45% of severe hemophilia A patients. In contrast to patients with bleeding disorders, characterization of the factor V Leiden and prothrombin 20,210 variants has dramatically improved the diagnostic yield for patients with inherited thrombophilia. One of these two genotypes is now found in greater than 60% of patients with a clinical history of familial thrombophilia. Finally, recent studies indicate that molecular genetic analysis is beginning to permit preliminary progress in the identification of arterial thrombotic risk. Further advances in characterizing the multigenic basis of thrombotic disease and the application of new technologies to aid in the assessment of genetic variability predict an increasingly important role for molecular diagnostic approaches to the evaluation of disorders of hemostasis. PMID- 10530717 TI - Molecular diagnosis of viral disease in hematology patients. AB - Viruses, through their direct and indirect effects on hematopoietic cells, are not only etiological agents of hematologic disease, but they both complicate transfusion therapy and are major causes of morbidity and mortality in immunosuppressed patients. The development of molecular techniques, especially DNA hybridization and amplification, has allowed new viruses to be discovered, has aided in the early diagnosis of infection before the development of an antibody response, and has revolutionized the detection of viruses in blood transfusions. PMID- 10530718 TI - Evaluation of clonality in myeloid stem-cell disorders. AB - Clonality in myeloid stem-cell disorders can be determined using either indirect methods such as analysis of X-chromosome inactivation patterns (XCIPs), or detection of specific abnormalities such as the chromosomal translocations characteristic of myeloid leukemias. XCIPs are particularly useful for disorders lacking evidence of a specific marker. Most females can be studied using polymerase chain reaction (PCR) analysis of differential DNA methylation patterns in the human androgen receptor (HUMARA) or phosphoglycerate kinase (PGK) genes, and approximately 68% can be studied using transcription assays of three polymorphic genes, glucose-6-phosphate dehydrogenase (G6PD), iduronate-2 sulfatase (IDS), and p55. Studies are limited by the incidence of constitutive and acquired (age-related) skewing and results must be carefully interpreted with reference to appropriate control samples. These techniques have been applied to clonality status of hematological disorders, lineage involvement in a clonal process, and detection of clonal evolution. PMID- 10530719 TI - Molecular approaches to the diagnosis and evaluation of lymphoid malignancies. AB - The processes of somatic immunoglobulin (Ig) and T-cell receptor (TCR) gene rearrangement that occur in lymphoid precursors provide insights into the pathogenesis and molecular analysis of lymphoid malignancies, in addition to the more universal molecular oncogenic mechanisms. Detection of lymphoid clonality can help distinguish polyclonal reactive disorders from clonal, predominantly, but not exclusively, malignant proliferations. Ig/TCR V-(D)-J polymerase chain reaction (PCR) amplification has largely replaced Southern blotting, and the techniques of PCR product analysis are evolving rapidly. V-(D)-J errors are often involved in genetic abnormalities leading to lymphoid malignancies, with consequent deregulated expression of the associated proto-oncogenes. Genetic abnormalities producing fusion transcripts and chimeric proteins are also frequent, particularly in acute lymphoblastic leukemia (ALL). A variety of molecular techniques, including reverse-transcriptase (RT)-PCR, Southern blotting, and fluorescence in situ hybridization (FISH) are finding an increasingly established place in the diagnosis, prognostic evaluation, therapeutic stratification, and follow-up of lymphoblastic leukemias, and it is likely that the same methods will be applied to non-Hodgkin's lymphoma (NHL) and to chronic leukemias. PMID- 10530720 TI - Molecular evaluation of acute myeloid leukemias. AB - Traditionally, acute myeloid leukemia (AML) has been diagnosed and classified based on the morphologic and cytochemical criteria of the French-American-British (FAB) classification system. However, more recent studies have demonstrated that the cytogenetic and molecular genetic abnormalities consistently associated with distinct forms of AML confer the most important prognostic information. Each broad category of de novo AML that arises in infants and children, young adults, and the elderly, or, AML arising secondary to antecedent myelodysplasia (MDS) or prior exposure to chemotherapy, radiotherapy, or environmental agents is characterized by distinct cytogenetic and molecular genetic abnormalities. Over the past 15 years, a number of these recurrent cytogenetic abnormalities have been cloned and characterized, yielding valuable insights into the mechanisms of leukemogenesis and providing powerful molecular tools for precise diagnosis and monitoring of minimal residual disease. As cytogenetic and molecular genetic characterization of AML cells is now considered crucial for both diagnostic and therapeutic decision-making, it is essential to have a working knowledge of the molecular basis of AML. This chapter reviews the molecular genetic features of different forms of AML and the new automated molecular technologies for residual disease assessment; the emerging molecular paradigms in this disease that are being exploited for therapeutic decision-making are presented. PMID- 10530721 TI - Molecular diagnosis and clinical decisions in adult acute leukemia. AB - A number of relatively new molecularly based diagnostic tests are now available to the physician involved in the treatment of patients with acute leukemia. These assays have considerable potential to improve prognostic accuracy, to aid in the selection of therapy, and to substantially enhance the ability to monitor results of treatment. Cytogenetic analysis now is the most important diagnostic test for determining prognosis and selecting therapy in acute myeloid leukemia (AML), and arguably, in acute lymphoblastic leukemia (ALL) as well. The accuracy of cytogenetics in many cases can be enhanced through the use of polymerase chain reaction (PCR)-based assays. Molecularly based assays of the leukemia cell's sensitivity or resistance to particular classes of drugs are beginning to emerge as potentially important tools for selecting therapies. Studies using molecular markers of disease to monitor response to therapy are yielding important clues about the biology of leukemia and are increasingly being used to guide therapy. The best uses of these new molecularly based assays are still uncertain and are the subjects of a large number of current clinic trials. While questions abound about their full potential, almost without question they will change the way we think about, diagnose, and treat leukemia. PMID- 10530722 TI - Hemispheric preponderance in categorical and coordinate visual processes. AB - Two experiments were conducted to address whether a left hemispheric bias would be observed for categorical processing in both 'what' and 'where' systems (experiment 1) while a reverse bias would characterize coordinate processing whatever the systems (experiment 2). Young normal subjects were tested using divided visual field tasks. The results of experiment 1 indicated that subjects made categorical judgments in both what and where systems faster when stimuli are presented to the left hemisphere. The results of experiment 2 showed a significant interaction between visual field and difficulty of processing coordinate relationships. Indeed, a left-hemisphere advantage was observed when the task required easy processing whereas a right-hemisphere advantage was noted for difficult distinctions either in location (where system) or in lightness (what system). The left-hemisphere advantage we observed for categorization in both systems confirms the Kosslyn's hypothesis (1989) for the where system and suggests that the same left-hemisphere advantage also exists for the what system. Concerning coordinate processing, our findings highlight the influence of processing difficulty on the hemispheric lateralization and evidence a right hemispheric advantage for difficult coordinate processing and a left hemispheric advantage for easy coordinate processing. The results are discussed in terms of possible link between on the one hand difficulty and coordinate processing, and easiness and categorization on the other hand. PMID- 10530723 TI - Frontal lobe lesions and electrodermal activity: effects of significance. AB - Several studies have shown that cortical damage, especially to the right hemisphere and to frontal lobes, may attenuate skin conductance responses selectively to psychologically significant stimuli. We tested this hypothesis in 32 patients with frontal lesions, verified by computer assisted tomography and magnetic resonance imaging, and 45 healthy controls. Patients and controls were given a protocol which included a rest period, a series of innocuous tones, and a reaction time task. Patients were given a second protocol in which they viewed slides with positive and negative emotional content and neutral slides. Results showed attenuated electrodermal activity (EDA) during task instructions and smaller skin conductance responses to reaction-time stimuli in patients compared to controls but few differences under passive conditions or in orienting responses to simple tones. Patients with lateral prefrontal and paraventricular lesions were especially low in EDA in the reaction time task, and those with right and bilateral lesions in the cingulate gyrus and/or frontal operculum had attenuated EDA in both protocols. We conclude that the effects of certain frontal lesions are on the psychological response to significance which is indexed by EDA rather than directly on EDA per se. PMID- 10530724 TI - Visual temporal asymmetries are related to asymmetries in linguistic perception. AB - There are numerous recent reports of low-level temporal asymmetries favouring the left hemisphere, and increasing speculation that the left hemisphere's relative superiority at linguistic processing may be related to these asymmetries. The present study sought to test this claim by assessing linguistic lateralization with the Fused Dichotic Words Test and visual temporal asymmetries with a lateralized version of an inspection-time test in a sample of 40 participants balanced for sex and handedness. We found evidence for a significant right-visual field (left-hemisphere) advantage for accuracy on the inspection-time task, F(1,36)=4.38, P = 0.043, and this asymmetry was significantly correlated with laterality scores on the linguistic dichotic-listening task, r = 0.306, P < 0.028 before disattenuation, and r = 0.486 after disattenuation. This result supports the position that low-level temporal asymmetries are related to asymmetries in linguistic processing. PMID- 10530725 TI - Cerebral blood flow patterns underlying the differential impairment in category vs letter fluency in Alzheimer's disease. AB - Verbal fluency tasks are used to assess language functioning in Alzheimer's disease (AD), and performance typically declines as the disease progresses. However, several studies have shown that Category Fluency performance (produce words from a category) declines faster than Letter Fluency performance (produce words beginning with a certain letter), which is not the case for other dementias. The purpose of this study was to determine if each of these two types of fluency tasks was associated with different patterns of cerebral blood flow abnormality in AD. A resting, Xenon-inhalation regional cerebral blood flow measurement (133Xe rCBF) and neuropsychological evaluation was administered to 25 patients with probable AD and 24 healthy elderly controls. Stepwise regression using rCBF measures as predictor variables was used to predict Category and Letter Fluency performance, in both a combined group of patients and controls, and in the patient group alone. Correlations were also computed between rCBF variables and the difference between normatively corrected scores on each task for each subject, which characterized the extent of the discrepancy between them. In full sample regressions, both Category and Letter Fluency were predicted by education and the decline in left inferior parietal flow, a focal AD-related deficit. Additional variance in Category fluency, however, was predicted by global mean flow, while additional variance in Letter Fluency was predicted by frontal flow. Within the patient sample, in turn, the primary predictor of Category Fluency was mean flow; the primary predictor of Letter Fluency was left sided frontal flow. Analysis of the fluency difference score revealed that relatively greater impairment of Category Fluency was associated with more typical, AD-related deficits in posterior temporal and parietal perfusion. When the two were equivalently impaired, typical AD-related deficits were accompanied by marked deficits in frontal perfusion. These findings are consistent with the underlying neuropsychology of these tasks, and suggest that Category Fluency's stronger association to the most typical CBF deficits of AD account for its greater sensitivity to this disease. Letter Fluency deficits, on the other hand, carry significant information about the degree to which perfusion deficits have spread to frontal cortex. PMID- 10530726 TI - A normal' category-specific advantage for naming living things. AB - 'Artefactual' accounts of category-specific disorders for living things have highlighted that compared to nonliving things, living things have lower name frequency, lower concept familiarity and greater visual complexity and greater within-category structural similarity or 'visual crowding' [7]. These hypotheses imply that deficits for living things are an exaggeration of some 'normal tendency'. Contrary to these notions, we found that normal subjects were consistently worse at naming nonliving than living things in a speeded presentation paradigm. Moreover, their naming was not predicted by concept familiarity, name frequency or visual complexity; however, a novel measure of visual familiarity (i.e. for the appearance of things) did significantly predict naming. We propose that under speeded conditions, normal subjects find nonliving things harder to name because their representations are less visually predictable than for living things (i.e. nonliving things show greater within-item structural variability). Finally, because nonliving things have multiple representations in the real world, this may lower the probability of finding impaired naming and recognition in this category. PMID- 10530727 TI - Interarticulator co-ordination in deaf signers with Parkinson's disease. AB - Motor control deficits in signers with Parkinson's disease (PD) were examined through analysis of their production of American Sign Language (ASL) fingerspelling, which is sequential and rapid motor behavior that has theoretical models of its underlying structure. Free conversation of two Deaf signers with PD and two Deaf control signers was analysed. In addition, scripted productions of one control signer were also analysed and directly compared to the same productions by the signers with PD. A featural analysis of ASL fingerspelling and a frame-by-frame analysis of multiple articulator movements were used to examine the fingerspelled productions. On the basis of the featural analysis, the signers with PD showed a variety of error patterns, all of which reflected attempts to reduce the motoric demands of coarticulation and thereby facilitate ease of articulation. Signers with PD either held individual segments in a fingerspelling sequence for a long time (segmentation), blended adjacent segments into a single segment (sequential blending), or broke handshapes down sequentially into their component features (featural unraveling). The results of both the featural analysis and the frame-by-frame analysis show that the PD signers have difficulty co-ordinating the movements of independent articulators in complex sequences. For example, the movements of independent articulators for fingerspelling (the thumb, fingers, and wrist) were markedly farther apart in time and more variable for the signers with PD. In addition, the signers with PD used fewer wrist movements while fingerspelling. Such deficits are consistent with claims that patients with PD are impaired in their ability to use ongoing sensorimotor information to program multi-articulator movements. PMID- 10530728 TI - Intact recognition of emotional prosody following amygdala damage. AB - Bilateral damage to the amygdala in a variety of animal species can impair emotional reactions to stimuli in several sensory modalities. Such damage in humans impairs visual recognition of emotion in facial expressions, but possible impairments in modalities other than vision have not been sufficiently explored. We examined two subjects with complete bilateral amygdala damage, and seven with unilateral amygdala damage, on a standardized task of emotional prosody recognition. The data were compared to those from 15 brain-damaged and from 14 normal control subjects. One of the bilateral amygdala subjects, whose lesions were restricted to the amygdala, was entirely normal in recognizing emotion in prosody on all tasks, the other, whose damage included substantial lesions also in extra-amygdalar structures, especially in right hemisphere, was normal on most, albeit not all, measures of emotional prosody recognition. We suggest that the human amygdala's role in recognizing emotion in prosody may not be as critical as it is for facial expressions, and that extra-amygdalar structures in right hemisphere may be more important for recognizing emotional prosody. It remains possible that recognition of emotion in classes of auditory stimuli other than prosody will require the amygdala. PMID- 10530729 TI - Attentional demands of perception of passive self-motion in darkness. AB - The purpose of this study was to determine whether significant attentional resources are required to accurately monitor changes in bodily orientation, using vestibular information. This question was addressed firstly using a dual-task paradigm in which orientation perception tasks and a speeded auditory tone discrimination task were carried out either singly or in combination. For the active orientation perception task, subjects were seated in darkness on a motorised chair which could be rotated about an earth-vertical axis. Following passive angular displacements, subjects were required to return the chair to their perceived starting position, using a joy-stick which controlled chair motion. For the speeded auditory task, subjects pushed a hand-held button as fast as possible when a tone was presented over headphones. When the two tasks were combined, reaction times on the auditory task increased. Reaction time also increased when subjects were simply asked to fixate during rotation. A second experiment demonstrated that if attention was occupied by performance of a demanding mental arithmetic task during the passive rotation, accuracy of subsequently repositioning the chair to the origin declined, implying that change in orientation had been less accurately registered when performing the concurrent mental task. In combination, these findings indicate that a small but significant degree of attention or cognitive effort is necessary to monitor accurately the direction and amplitude of a brief angular rotation, and to suppress vestibulo ocular reflex eye movement. PMID- 10530730 TI - The dependence of span and delayed-response performance on prefrontal cortex. AB - Theoretical and empirical research on the cognitive functions of the prefrontal cortex have established that this region mediates what have been called 'executive' processes that can influence working and long-term memory. Despite the accumulation of such empirical evidence, the dependence of purely mnemonic portions of memory tasks on PFC remains unresolved. To address this issue, we performed an analysis of reports of performance on tests of working memory of patients with lesions of the dorsolateral prefrontal cortex, focusing on published reports in the literature of simple span and delayed-response tasks. We found that none of the eleven studies of forward verbal and spatial span in patients with prefrontal cortical lesions that we reviewed (reflecting the performance of 166 individual patients) demonstrated a statistically significant deficit relative to normal controls. In contrast, our review of the delayed response literature indicated that there are conditions under which PFC lesions disrupt delayed-response performance. Based on the results of our review of the literature, we present testable hypotheses about the working memory functions of the PFC. PMID- 10530731 TI - Abnormalities of imaged motor sequences in children with developmental coordination disorder. AB - The chronometry of real and imagined movements was investigated in a group of children with developmental coordination disorder (DCD) and a group of matched controls. The visually-guided pointing task was used to investigate the speed for accuracy trade-offs that occur as target size is varied for both real and imagined performance. In the control group, the speed for accuracy trade-off for both real and imagined performance conformed to Fitts' law. In the DCD group only real movements conformed to Fitts' law. This pattern of performance suggests that children with DCD have an impairment in the ability to generate internal representations of volitional movements. This may reflect part of a general impairment in the processing of efference copy in DCD. PMID- 10530732 TI - Cochlear implant in patients with residual hearing. AB - OBJECTIVE: The postoperative speech perception abilities of severely hearing impaired patients with multi-channel cochlear implant were compared with preoperative speech perception performance with conventional hearing aids. METHODS: Cochlear implantation was performed in six severely to profoundly hearing-impaired patients. They had unaided pure-tone thresholds of 70-100-dB HL and aided thresholds of 35-90-dB HL in the better ear, but were not able to perceive speech sounds well with hearing aids. RESULTS: Postoperatively, all the patients had significantly improved speech perception performance, exceeded the average skills of profoundly deaf cochlear implant users, and were able to communicate without writing. CONCLUSION: These results imply that cochlear implant may be indicated for severely to profoundly deaf subjects, if they receive little or no benefit from conventional hearing aids. PMID- 10530733 TI - The protective effect of the sympathetic nervous system against acoustic trauma. AB - OBJECTIVE: the cochlea is innervated by the sympathetic nerve fibers. However, the functions of those fibers in the cochlea are still controversial. The present study was designed to determine whether the sympathetic nervous system (SNS) exerts a protective or enhancing effect on acoustic trauma. METHODS: acoustic overstimulation (either of 110, 115, or 130 dB SPL for 10 min) was performed in guinea pigs during electrical stimulation of the ipsilateral cervical SNS, after its surgical elimination or in the non-treated condition. The threshold shift of the compound action potential (CAP) from the pre-exposure value was measured at 1 h and at 1 week after acoustic overstimulation. Two-way analyses of variance (ANOVAs) were completed for the SNS conditions and the frequencies. RESULTS: although no significant difference was found at 1 h after overstimulation among these three groups, the CAP threshold shift at 1 week (110 and 115 dB SPL) was significantly smaller in the SNS stimulation group than in the other two groups. CONCLUSION: a protective effect was observed in the SNS stimulation group 1 week after the exposure to acoustic overstimulation of moderate intensity (from 110 to 115 dB SPL for 10 min). PMID- 10530734 TI - Mechanics of the middle ear system: computerized measurements of its pressure volume relationship. AB - A new method is described measuring the pressure-volume relationship of the middle ear system (MES). These measurements express the dynamic mechanical properties of the MES. Ear canal pressure changes are measured in response to tympanic membrane (TM) volume displacements in a material of 39 younger normal adults. During one recording procedure several displacements curves are obtained from which one curve is isolated representing the neutral position of the TM. From this curve the following variables are determined: hysteresis (microJ) describing the viscoelastic properties of the MES, compliance (mm3/kPa) describing its elasticity, Prange (kPa) describing the pressure range of the curve, and Pec0 (kPa) describing the ear canal pressure for the neutral position of the TM. Normative data are presented and compared with tympanometric measurements. Compliance correlates significantly to static admittance (P<0.001), while Pec0 correlates significantly to middle ear pressure (P<0.001). Further, data on repeatability and sources of measurement errors are reported, which support a high reliability of the method. Compared with tympanometry the method is more detailed and has several advantages, which are discussed, and it has been found valuable for future mechanical studies of the MES. These studies include possibilities for diagnostics of middle ear disorders and derivation of pressure volume equations useful in modeling of the MES. PMID- 10530735 TI - Hearing impairment in patients with acoustic neuroma--analysis by electrocochleography. AB - OBJECTIVE: In patients with acoustic neuroma, the site and severity of hearing impairment are important in discussing surgical approaches. Since the effectiveness of conventional auditory psychological testing is limited, we studied objectively hearing impairment of the cochlea and the cochlear nerve due to the tumor. METHODS: Electrocochleography (ECochG) was carried out in 21 patients with acoustic neuroma. Cochlear microphonic potential (CM) and action potential (AP) in ECochG evoked with clicks and short tone bursts were recorded through a transtympanic needle electrode technique. Cochlear function was studied using the detection thresholds of CM, and cochlear nerve involvement was analyzed by differences between AP and CM detection thresholds. RESULTS: The 1 kHz CM detection threshold was elevated in 17 (81.0%) of 21 patients indicating cochlear impairment. Of seven patients with normal hearing or mild sensorineural hearing loss in pure tone audiometry, three had a slightly elevated CM detection threshold. Of five patients with pronounced pure tone levels, four showed a CM response and were thought to have mild cochlear dysfunction. Cochlear nerve impairment was confirmed in three of four patients with well-developed CM based on elevated AP detection thresholds. Three patients had CM response but no AP response, suggesting severe cochlear nerve impairment. CONCLUSION: Disorders of the cochlea and the cochlear nerve can be evaluated with ECochG AP and CM measurement. The findings of ECochG are thought to be important information to judge hearing prognosis, thereby enhancing its clinical utility. PMID- 10530737 TI - Autonomic nervous function in patients with Meniere's disease evaluated by power spectral analysis of heart rate variability. AB - OBJECTIVE: A hypothesis has been advanced that the autonomic nervous dysfunction (AND) relates to the development of vertigo in Meniere's disease (MD). We also studied the causal relationship between AND and vertigo in MD. METHODS: We evaluated autonomic nervous function in 17 patients with MD (five men and 12 women ranging in age from 16 to 70 years) by classifying them by their stages of attack and interval of vertigo and with power spectral analysis (PSA) of heart rate variability. Fourteen healthy volunteers were also tested as controls. RESULTS: At the interval stage, parasympathetic nervous hypofunction and significant depression of sympathetic response due to postural changes from the supine to the standing position were observed in many of those patients. At the attack stage, sympathetic nervous hypofunction was observed in some of the patients. CONCLUSION: These findings lead us to the conclusion that AND relates to vertigo in MD as a predisposing factor. However, the question of whether AND relates as a trigger or as a consequence of vertigo in MD has not been adequately solved in this study. We will make further studies on circadian variation of autonomic nervous function. PMID- 10530736 TI - Lidocaine test in patients with tinnitus: rationale of accomplishment and relation to the treatment with carbamazepine. AB - OBJECTIVES: There is strong evidence in the literature about the effect of local anesthetics such as lidocaine in controlling tinnitus; these agents act by stabilizing hair cell membrane and cochlear nerve fibers. However, the effect of intravenous lidocaine is transient, and its oral analog (tocainide) does not have the same efficacy for long-term treatment in patients with tinnitus. Some oral anti-epileptic drugs (carbamazepine, for instance) have been used alternatively in several studies. The aim of this work is to evaluate the response to intravenous lidocaine in patients with intractable tinnitus and the effect of oral carbamazepine in long-term maintenance of tinnitus relief. PATIENTS AND METHODS: We studied prospectively 50 patients (28 females and 22 males; mean age 50.9 years) who underwent the lidocaine test, performed by a 3-min intravenous infusion of 2% lidocaine chloridrate. The patients who experienced any relief after the test started treatment with oral carbamazepine in ascending dosages (50 600 mg/day). RESULTS: The results were classified as tinnitus abolition (18%), marked relief (32%), partial relief (26%), unchanged (22%), or worsening (2%). The lidocaine test showed favorable results in 76% of patients, especially those with bilateral tinnitus (P < 0.001). Afterwards, 50% of patients treated with carbamazepine maintained the improvement of tinnitus (P = 0.0034). CONCLUSION: The authors conclude that intravenous lidocaine is effective in reducing intractable tinnitus and that there is a close association between lidocaine and oral carbamazepine effects. Therefore, carbamazepine can be used for the treatment of tinnitus when the patient achieves improvement of symptom after the lidocaine test. PMID- 10530738 TI - Headache in Meniere's disease. AB - OBJECTIVE: We examined headache in 86 patients with severe or moderately severe Meniere's disease (MD) or with Meniere's syndrome (MS) chosen for intratympanic gentamicin treatment. Forty-five patients with vestibular neuronitis (VN) served as a control group. METHODS: In addition to a clinical examination, the patients filled out a questionnaire concerning their headache. RESULTS: Altogether 60 MD patients (70%) and 26 VN patients (58%) reported headache. Headache was severe in 35 MD patients (58%), moderate in 16 patients (27%) and slight in nine MD patients (15%), and as a whole more severe than that of the VN patients (P < 0.01). The MD patients exhibited significantly more occipital (P < 0.005) and neck headache (P < 0.005) than the VN patients. Analgesics had been used by 82%, antidepressants by 35%, sumatriptan by 13% and carbamazepine and/or amitriptyline by 12% of the MD patients suffering from headache. Pain relief was reported as good by 27%, satisfactory by 60%, poor by 5% of the MD patients and 8% could not rate the pain relief. In this study migraine was diagnosed in 5 MD patients. CONCLUSION: It is concluded that MD is associated with headache that can be handicapping, and tricyclic antidepressants with pain alleviating medication is often needed to treat the headache in MD whereas sumatriptan did not alleviate the headache of the non-migraine patients. PMID- 10530740 TI - Endoscopic marsupialization of bilateral lacrimal sac mucoceles with nasolacrimal duct cysts. AB - A lacrimal sac mucocele is an uncommon disease usually treated by ophthalmologists. In rare cases, it is sometimes associated with a nasolacrimal duct cyst presenting as an intranasal cystic mass, which needs the involvement of an otolaryngologist in diagnosis and management. Two cases of lacrimal sac mucoceles with nasolacrimal duct cysts are presented with a brief literature review. Both cases presented with intranasal cystic masses that caused nasal obstruction and were cured with endoscopic marsupialization of the cysts. PMID- 10530739 TI - Unilateral examination of olfactory threshold using the Jet Stream Olfactometer. AB - OBJECTIVE: The Jet Stream Olfactometer, a modification by the T&T olfactometer, has been recently developed and is now commercially available. This Jet Stream Olfactometer can routinely measure the unilateral sense of smell. Clinical usefulness of the Jet Stream Olfactometer was evaluated. METHODS: Twenty-three patients with sinus-related symptoms were examined. Unilateral olfactory acuity was examined using the Jet Stream Olfactometer and compared it with the anatomy of the olfactory cleft by computed tomographic (CT) scans. RESULTS: In 13 of the 23 patients examined the right olfactory threshold was similar to that of the left. Of these 13, in seven patients there was transport damage of the odorants and in the other six there was sensorineural damage. Ten patients showed an apparent difference between the right and left smell thresholds. Anosmia in seven of these ten patients was due to a conductive olfactory disturbance resulting from rhinosinusitis, whereas sensorineural damage was recognized in the other three patients. The difference in the detection threshold between the right and left nasal cavities was well correlated to that of the opacity between the right and left olfactory clefts. CONCLUSION: Jet Stream Olfactometer provides a convenient and reliable means for assessing the ability of unilateral smell. PMID- 10530741 TI - Advantage and disadvantage of KTP-532 laser tonsillectomy compared with conventional method. AB - OBJECTIVE: The purpose of this prospective study is to define the advantages and disadvantages of KTP laser tonsillectomy compared with those of the conventional method. METHODS: Eighteen adult patients (ten male and eight female, ranging in age from 14 to 44 years) underwent KTP-532 laser tonsillectomy on one side and standard dissection surgery on the other side under general anesthesia. RESULTS: By KTP laser tonsillectomy, there was a reduction in intraoperative blood loss and average time for removing one tonsil. On the second day of tonsillectomy, subjective pain on the KTP laser surgery side was less than that on the conventional surgery side. By the days 5-8, however, this effect disappeared and many patients indicated the laser side was more painful. There was no postoperative bleeding after KTP laser tonsillectomy. Laser surgery appeared to lead to slow wound healing during the whole post-operative course with significant difference compared with the conventional method. Disadvantages of postoperative pain and the possibility of secondary infection due to slow wound healing could be prevented by application of antibiotics and an anodyne. CONCLUSION: Considering safety and reliability during surgery, KTP laser was considered useful for adult tonsillectomy. PMID- 10530742 TI - Noninvasive scintigraphic method to quantify unstimulated secretions from individual salivary glands. AB - PURPOSE: Historically salivary gland function has been associated with maintenance of oral hygiene The difference in secretory behavior of parotid and submandibular glands has previously been shown. The purpose of this study was to establish a noninvasive technique for quantification of unstimulated (resting state) secretion of saliva based on the tracer output theory. PROCEDURES: A total of 14 99mTc-pertechnetate salivary function studies were performed under Gamma camera. The time activity curves were subjected to a two step background subtraction protocol (area normalised background subtraction, followed by a graphical method for background correction). Individual salivary glands were modeled as Organ Curve = Input - Output. From these Uptake rates and unstimulated salivary gland fractional output rates (FOR) were calculated. MAIN RESULTS: Parotid as well as submandibular glands have identical Uptake rates for the tracer. A distinct pattern was noted in submandibular glands as against parotid glands. Submandibular glands showed a steady rise in total quantity of unstimulated secretion. The FOR for submandibular glands was about three times higher than parotid glands (P< 0.000001). The observed distribution of FOR for parotid and submandibular glands FOR showed that parotid-FOR was normally on the lower side whereas submandibular-FOR showed a wide range of distribution which was multimodal in nature. PRINCIPAL CONCLUSION: A unique approach has been presented for quantification of unstimulated salivary secretion. The method is simple and noninvasive. PMID- 10530743 TI - Otorhinolaryngological aspects of Xeroderma pigmentosum. AB - OBJECTIVE: to evaluate the probable presence of otorhinolaryngological pathology accompanied by head and neck region skin findings in patients with Xeroderma pigmentosum. METHODS: a total of 19 patients with Xeroderma pigmentosum were investigated for otorhinolaryngological findings. The patients gave their anamnesis and underwent physical examination, audiological tests and endoscopic examination. RESULTS: various malignancies developed in 14 patients on the sun exposed areas of the head and neck region. Multiple malignancies were found in six of them. There was no other pathological condition secondary to this rare clinical entity. CONCLUSION: Xeroderma pigmentosum causes skin lesions. Some otolaryngological findings such as rhinitis, sinusitis etc. were thought to be coincidental. PMID- 10530744 TI - Developmental changes in histochemical properties of intrinsic laryngeal muscles in rats. AB - OBJECTIVE: Using neonatal rats, the developmental changes in muscle fiber type of the intrinsic laryngeal muscles were analyzed. The potential influence of two factors were also studied, that were predicted would influence developmental changes in muscle fiber type, denervation and hypothyroidism. METHODS: Using the histochemical technique of myosin ATPase staining, postnatal changes in the ratio of muscle fiber types of each intrinsic laryngeal muscle were determined. In addition, to clarify factors influencing the development of the intrinsic laryngeal muscles, the same technique was employed to study recurrent laryngeal nerve-denervated rats and rats with experimentally-induced hypothyroidism. RESULTS: In normal pups, type 2C fibers had almost disappeared by postnatal day (PND) 14. In denervated pups, differentiation to type 1 and 2A muscle fibers was not observed. In contrast, differentiation to type 2B muscle fibers was impaired in the hypothyroid pups. CONCLUSION: The differentiation of intrinsic laryngeal muscles occur earlier than that of hindlimb muscles. Each intrinsic laryngeal muscle exhibits a particular pattern of developmental changes in normal pups. The developmental changes in the intrinsic laryngeal muscles are affected by recurrent laryngeal nerve innervation and thyroid hormonal control. The findings suggest that both recurrent laryngeal nerve innervation and thyroid hormone play important roles in the differentiation of the intrinsic laryngeal muscles. PMID- 10530745 TI - Middle ear inflation for diagnosis and treatment of otitis media with effusion. AB - An adult (18 years), healthy, male subject with persistent bilateral middle ear (ME) underpressure and a history of recurrent otitis media into his teen years was identified. The response of his MEs to air inflation was evaluated and showed an immediate pressure increase after a Valsalva maneuver followed by a rapid pressure drop to approach the pre-inflation levels. That response is consistent with the presence of ME effusion, which was not diagnosed by otoendoscopy or tympanometry, but was visualized bilaterally within the mastoid regions using magnetic resonance imaging (MRI). The patient was treated for 25 days with ME inflation (3/day) and then re-examined. On each treatment day, he recorded his ME pressure using tympanometry before and after one inflation maneuver. The patient's compliance with the treatment protocol was high, and successful gas transfers were documented on most days. Over the course of treatment, pre inflation ME pressure became more normal bilaterally. When compared to the pre treatment test, the post-treatment inflation test showed a similar rate of ME pressure decrease, but significantly higher terminal pressures. On follow-up but not during the pre-treatment period, discrete changes in ME pressure attributable to ET openings were noted during test sessions. MRI documented lesser amounts of effusion in the mastoid, but not complete disease resolution. The significance of these observations to the design of a well controlled clinical trail of ME inflation as a treatment for otitis media is discussed. PMID- 10530746 TI - Malignant paraganglioma of frontoethmoidal region. AB - Nonchromaffin paragangliomas are unusual tumours arising from widely distributed paraganglionic tissues probably of neural crest origin. In the head and neck region they are usually seen as carotid body or jugulotympanic tumours. Other rarely reported sites in the head and neck region are the orbit, nose and larynx. This report deals with a case of sinonasal paraganglioma which was initially treated with surgery and radiotherapy. Twenty two years later the tumour recurred and showed a rapid growth due to malignant transformation which we believe is late effect of radiotherapy. The clinical features, histopathology and role of radiotherapy in sinonasal paragangliomas together with a review of the medical literature have been discussed. PMID- 10530747 TI - The operation of upper esophageal web in Plummer-Vinson syndrome: a case report. AB - Most cases of dysphagia associated with Plummer-Vinson syndrome are expected to improve with the oral administration of ferrous agents. When a web is the cause of the symptoms, surgical management is rarely necessary. However the surgical indication and technique for the web have been controversial. The patient was a 56-year-old woman who complained of restricted dietary habit because of an upper esophageal circumferential web associated with Plummer-Vinson syndrome. The circumferential and membranous web was resected with a surgical knife and scissors through the inner lumen of esophagus and the raw surface was sutured at five places with 4-0 proline thread under microlaryngosurgery. This surgical treatment resulted in diminished dysphagia and no recurrence of the web after the surgery. PMID- 10530748 TI - Two cases of piriform sinus fistula which required a long time for diagnosis. AB - Two cases of piriform sinus fistula which had contradistinctive occurrences, were reported. In case 1, a 58-year-old man suffered from the disease without having any symptom for a long time and in case 2, a 39-year-old woman had been troubled with repeated cervical abscess from 3 years old. In both cases, indirect laryngoscopy and laryngofiberscopy showed saliva pooling in the bilateral piriform sinus and barium fluoroscopy with the Valsalva maneuver revealed the fistula originating from the apex of left piriform sinus. In case 2, computed tomography (CT) and magnetic resonance imaging (MRI) demonstrated an abscess in and around the left lobe of the thyroid. In both cases the fistulectomy was performed and their postoperative conditions have been uneventful for more than 2 years without recurrence. The etiology, cause, clinical features, diagnosis and therapy of piriform sinus fistula were reviewed. PMID- 10530749 TI - Mechanisms of photoreceptor death and survival in mammalian retina. AB - The mammalian retina, like the rest of the central nervous system, is highly stable and can maintain its structure and function for the full life of the individual, in humans for many decades. Photoreceptor dystrophies are instances of retinal instability. Many are precipitated by genetic mutations and scores of photoreceptor-lethal mutations have now been identified at the codon level. This review explores the factors which make the photoreceptor more vulnerable to small mutations of its proteins than any other cell of the body, and more vulnerable to environmental factors than any other retinal neurone. These factors include the highly specialised structure and function of the photoreceptors, their high appetite for energy, their self-protective mechanisms and the architecture of their energy supply from the choroidal circulation. Particularly important are the properties of the choroidal circulation, especially its fast flow of near arterial blood and its inability to autoregulate. Mechanisms which make the retina stable and unstable are then reviewed in three different models of retinal degeneration, retinal detachment, photoreceptor dystrophy and light damage. A two stage model of the genesis of photoreceptor dystrophies is proposed, comprising an initial "depletion" stage caused by genetic or environmental insult and a second "late" stage during which oxygen toxicity damages and eventually destroys any photoreceptors which survive the initial depletion. It is a feature of the model that the second "late" stage of retinal dystrophies is driven by oxygen toxicity. The implications of these ideas for therapy of retinal dystrophies are discussed. PMID- 10530750 TI - Primate retina: cell types, circuits and color opponency. AB - The link between morphology and physiology for some of the cell types of the macaque monkey retina is reviewed with emphasis on understanding the neural mechanism for spectral opponency in the light response of ganglion cells. An in vitro preparation of the retina is used in which morphologically identified cell types are selectively targeted for intracellular recording and staining under microscopic control. The goal is to trace the physiological signals from the long (L), middle (M) and short-wavelength sensitive (S) cones to identified cell types that participate in opponent and non-opponent signal pathways. Heterochromatic modulation photometry and silent substitution are used to characterize L-, M- or S-cone inputs to the receptive fields of distinct horizontal cell, bipolar cell, ganglion cell and amacrine cell types. The majority of the retinal cell types await detailed analysis, and knowledge of the mechanisms of opponency remains incomplete. However results thus far have established: (1) Horizontal cell interneurons make preferential connections with the three cone types, but cannot provide a basis for spectral opponency in the circuitry of the outer retina. (2) A morphologically distinctive bistratified ganglion cell type transmits a blue-ON yellow-OFF spectral opponent signal to the parvocellular division of lateral geniculate nucleus. The morphology of this ganglion cell type suggests a simple synaptic mechanism for blue yellow opponency via converging input from an S-cone connecting ON-bipolar cell and an L - M cone connecting OFF-bipolar cell. (3) Midget ganglion cells, whose axons project to the parvocellular layers of the lateral geniculate nucleus and are assumed to be the origin of red/green opponent signals, show a non-opponent, achromatic physiology when recorded in the retinal periphery the underlying circuitry for red green opponency thus remains controversial, and (4) recent recordings from identified bipolar and amacrine cells in macaque suggest that a more complete accounting of opponent circuitry is a realistic goal. PMID- 10530751 TI - Glutamate receptors and circuits in the vertebrate retina. AB - We survey the evidence for L-glutamate's role as the primary excitatory neurotransmitter of vertebrate retinas. The physiological and molecular properties of glutamate receptors in the retina are reviewed in relation to what has been learned from studies of glutamate function in other brain areas and in expression systems. We have focused on (a) the evidence for the presence of L glutamate in retinal neurons, (b) the processes by which glutamate is released, (c) the presence and function of ionotropic receptors for L-glutamate in retinal neurons, (d) the presence and function of metabotropic receptors for L-glutamate in retinal neurons, and (e) the variety and distribution of glutamate transporters in the vertebrate retina. Modulatory pathways which influence glutamate release and the behavior of its receptors are described. Emphasis has been placed on the cellular mechanisms of glutamate-mediated neurotransmission in relation to the encoding of visual information by retinal circuits. PMID- 10530752 TI - Amino acid neurochemistry of the vertebrate retina. AB - The dominant neurochemicals involved in encoding sensory information are the amino acid neurotransmitters, glutamate, gamma-aminobutyrate (GABA) and glycine, which mediate fast point-to-point synaptic transmission in the retina and other parts of the central nervous system. The relative abundance of these neurochemicals and the existence of neuronal and glial uptake mechanisms as well as a plethora of receptors support the key role these neurochemicals play in shaping neural information. However, in addition to subserving neurotransmitter roles, amino acids subserve normal metabolic,cellular functions, may be precursors for other amino acids, and may also be associated with protein synthesis. Post-embedding immunocytochemistry of small molecules has allowed the characterization of multiple amino acid profiles within subpopulations of neurons in the vertebrate retina. The general theme emerging from these studies is that the retinal through pathway uses glutamate as its neurotransmitter, and the lateral elements, GABA and/or glycine. Co-localization studies using quantitative immunocytochemistry have shown that virtually all neuronal space can be accounted for by the three dominant amino acids. In addition, co-localization studies have demonstrated that there are no purely aspartate, glutamine, alanine. leucine or ornithine immunoreactive neurons and thus these amino acids are likely to act as metabolites and may sustain glutamate production through a multitude of enzymatic pathways. The mapping of multiple cellular metabolic profiles during development or in degenerating retinas has shown that amino acid neurochemistry is a sensitive marker for metabolic activity. In the degenerating retina, (RCS retina), neurochemical anomalies were evident early in development (from birth), even before photoreceptors mature at PND6-8 implying a generalized metabolic dysfunction. Identification of metabolic anomalies within subpopulation of neurons is now possible and can be used to investigate a multitude of retinal functions including amino acid metabolic and neurochemical changes secondary to external insult as well as to expand our understanding of the intricate interrelationship between neurons and glia. PMID- 10530753 TI - Synchronous and asynchronous updating in cellular automata. AB - We analyze the properties of a synchronous and of various asynchronous methods to iterate cellular automata. Asynchronous methods in which the time variable is not explicitly defined, operate by specifying an updating order of the cells. The statistical properties of this order have significant consequences for the dynamics and the patterns generated by the cellular automata. Stronger correlations between consecutive steps in the updating order result in more, artificial structure in the patterns. Among these step-driven methods, using random choice with replacement to pick the next cell for updating, yields results that are least influenced by the updating method. We also analyse a time-driven method in which the state transitions of single cells are governed by a probability per unit time that determines an exponential distribution of the waiting time until the next transition. The statistical properties of this method are completely independent of the size of the grid. Consecutive updating steps therefore show no correlation at all. The stationary states of a cellular automaton do not depend on whether a synchronous or asynchronous updating method is used. Their basins of attraction might, however, be vastly different under synchronous and asynchronous iteration. Cyclic dynamics occur only with synchronous updating. PMID- 10530754 TI - Embryonic electronics. AB - Within the general domain of bio-inspired computing, a particular trend over the past few years has been that of constructing actual hardware devices that are inspired by nature. This paper describes one such project-Embryonics (embryonic electronics)-inspired in particular by the process of embryogenesis. Our ultimate objective is the construction of large-scale integrated circuits, exhibiting the properties of self-repair (healing) and self-replication, found until now only in living beings. We present the silicon-based artificial cell, followed by a description of mechanisms operating at the cellular level: cellular differentiation, cellular division, regeneration, and replication. We then present the cell's composition as an ensemble of lower-level elements, known as 'molecules'. As electronic chips grow evermore complex, the need for self-repair capabilities will become increasingly crucial. The Embryonics approach represents one possible way of confronting this pivotal problem. PMID- 10530755 TI - Evolution of consciousness. AB - The evolution of codification within the increasing complexity and diversity of relations is examined within a triadic architectural frame of filiated hierarchical levels of both material organization and consciousness. Inertia and entropy are understood as basic forces within codal organization, and evolution is examined as a force of mediation between these two forces, operating to move energy to more complex and diverse codal relations. PMID- 10530756 TI - Kinetic analysis of enzyme reactions with slow-binding inhibition. AB - In this paper we present a general kinetic study of slow-binding inhibition processes, i.e. enzyme reactions that do not respond instantly to the presence of a competitive inhibitor. The analysis that we present is based on the equation that describes the formation of products with time in each case on the experimental progress curve. It is carried out under the condition of limiting enzyme concentration and allows the discrimination between the different cases of slow-binding inhibition. The mechanism in which the formation of complex enzyme inhibitor is a single or two slow steps or follow a rapid equilibrium, has been considered. The corresponding explicit equations of each case have been obtained and checked by numerical integration. A kinetic data analysis to evaluate the corresponding kinetic parameters is suggested. We illustrate the method, numerically by computer simulation, of the reaction and present some numerical examples that demonstrate the applicability of our procedure. PMID- 10530757 TI - Implications of relaxation dynamics in the synaptic control of olfactory cortex activity. AB - In a previous work (Ballain et al., 1998. Biol. Cyber. 79, 323-336) we reported the analysis of a model for the piriform cortex activity in rats based on experimental data. In this paper, we study an extension of this model by supplementing it with equations for the post-synaptic conductance and/or the pre synaptic activation threshold. We use the present model's outputs to account for experimental data based on paired stimulation in the opossum or the rat, obtained either through electrical recording or optical mapping of the cortex activity. The model exhibits great robustness when it comes to large variation in synaptic characteristics. Model outputs mimic satisfactorily the three kind of responses to paired stimuli (Litaudon and Cattarelli, 1996. Eur. J. Neurosci. 8, 21-29) and the recovery of the excitable capacities as demonstrated by Haberly (1973. J. Neurophysiol. 36 (4), 789-802) and Ferreyra-Moyano et al. (1985. Brain Res. Bull. 15, 237 248). PMID- 10530758 TI - Menstruation: induction by matrix metalloproteinases and inflammatory cells. AB - Menstruation occurs at the end of a normal reproductive cycle in the human female, following the fall in progesterone resulting from the demise of the corpus luteum. Current data support a central role for the matrix metalloproteinases in menstruation but their focal pattern of expression within peri-menstrual and menstrual endometrium suggests local rather than hormonal regulation. This review emphasizes the similarities between menstruation and an inflammatory process and examines the relationship between cells of hemopoietic lineage, particularly mast cells, eosinophils, neutrophils and macrophages, and the local production and activation of matrix metalloproteinases within the endometrium. It proposes a complex of critical regulatory circuits, initially activated by the withdrawal of progesterone, which provide interactions between the migratory cells that produce a myriad of important regulatory molecules and endometrial stromal and epithelial cells which produce both chemokines and matrix metalloproteinases. These mechanisms could account for the focal nature of the tissue degradation at menstruation. PMID- 10530759 TI - Influences of surgical castration on the thymus of male rats. AB - In previous studies, we have shown that castration of Sprague-Dawley rats enhances thymic weight through puberty whilst sex steroids reduce the castration induced hypertrophy. In the current study, we have confirmed that castration enhances thymic growth compared to age-matched intact controls. In addition, immunoassays were used to measure thymosin alpha1 and thymosin beta4 levels in sera from intact and castrate rats. Castrate animals displayed greater sera levels of thymosins compared to sera from intact animals. To test whether the enhanced thymic weight and increased levels of thymosins observed post-castration were able to influence immune function of castrate animals, concanavalin A was used in the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide assay to examine lymphocyte and thymocyte responses from both intact and castrate male rats. Responses of cells isolated from castrate rats demonstrated that lymphocytes and thymocytes were stimulated at low levels of concanavalin A (1.56 3.13 microg/ml for lymphocytes and 1.56 microg/ml for thymocytes) compared to the same cell types isolated from intact rats. Concentrations of concanavalin A ranging from 6.25 to 200 microg/ml produced no significant differences in response from intact and castrate animals. PMID- 10530760 TI - Microchimerism and blocking activity in women with recurrent spontaneous abortion (RSA) after alloimmunization with the partner's lymphocytes. AB - Alloimmunization therapy using the partner's leukocytes has been reported to be effective in preventing the failure of pregnancy in patients who have suffered RSA of unknown cause. After alloimmunization, several investigators have reported the presence of blocking factors (BF) in women with successful pregnancies in in vitro assays of lymphocyte response. The recent discovery of small numbers of ubiquitous donor cells (microchimerism) in human transplants up to 29 years post transplantation has raised questions about the migration of the chimeric cells and their role in the induction and perpetuation of tolerance. We have investigated the production of BF in the mixed lymphocyte reaction (MLR) before and in some patients after alloimmunization and its possible relation with the development of microchimerism (M). Before the treatment we studied 14 couples with three or more abortions who were evaluated clinically to rule-out genetic, structural, endocrine, infectious and autoimmune causes. The M study was done by nested PCR-SSP technique with HLA-DR alleles, before and after 30 days of the last immunization. Before the treatment only one patient was M positive and none were BF positive with inhibition effect (IE) > 50. Only eight underwent treatment. The patients had between three and nine alloimmunizations (x = 4.7). After treatment, all patients were M positive with IE > 50. Six months after the last immunization, four patients are M positive with IE > 50. In conclusion, the hypothesis proposes that alloimmunization establishes a state of microchimerism that would be the necessary allogeneic stimulus for T-cell activation, and the induction or maintenance of tolerance to the fetus during pregnancy. reserved. PMID- 10530762 TI - Infection in pregnancy. Report on the workshop held by the Materno-Fetal Immunology Group (MFIG) of the British Society for Immunology at the 6th Annual Congress of the BSI, December 4, 1998, in Harrogate, UK. PMID- 10530761 TI - Microscopic evidence against HIV-1 infection of germ cells or attachment to sperm. AB - For a number of years we have intensively investigated the localization of HIV-1 in male genital tract tissues and secretions using a variety of microscopy techniques including immunocytochemistry, in situ hybridization, in situ PCR and electron microscopy. Our studies have failed to demonstrate an association between HIV-1 and either testicular germ cells or spermatozoa. In this article we present our results in the context of other related studies, and discuss the strengths and weaknesses of the techniques that have been used to address this important research question. PMID- 10530763 TI - Inhibitory effect of mimosine on proliferation of human lung cancer cells is mediated by multiple mechanisms. AB - The plant amino acid mimosine has been reported to block cell cycle progression in the late G1 phase. A recent study showed that mimosine might induce growth arrest by activating the expression of p21CIP1, a cyclin-dependent kinase inhibitor (CDKI), and by inhibiting the activity of cyclin E-associated kinases in human breast cancer cells. However, mimosine at higher concentrations also blocked proliferation of p21-/- cells by unknown mechanisms. In this study, we investigated the effect of mimosine on the expression of cyclins and CDKIs in human lung cancer cells. We found that mimosine specifically inhibited cyclin D1 expression in H226 cells. The expression of another G1 cyclin, cyclin E, was not regulated by mimosine in all lung cancer cell lines examined. Moreover, mimosine induced p21CIP1 expression in H226 and H358 cells, while it activated p27KIP1 expression in H322 cells. However, mimosine does not affect transcription of these genes directly because significant changes in cyclin D1 or CDKI expression were observed at 12-24 h after drug addition. Our results indicate that mimosine may block cell proliferation by multiple mechanisms and this amino acid is a useful agent for the study of cell cycle control. PMID- 10530764 TI - Morphological and functional effects of antisense RNA to the deleted in colorectal carcinoma (DCC) gene in a pancreatic carcinoma cell line. AB - To investigate the role of the deleted in colorectal carcinoma gene (DCC) in cells of pancreatic origin (MiaPaCa-2) we established cell lines stably expressing DCC antisense RNA. Expression of DCC antisense RNA led to striking alterations in the MiaPaCa-2 cell line. Antisense transfectants had nearly lost adherence and had acquired a spherical morphology. The ordered structure of actin bundles in the parental cell line had been lost largely in DCC antisense RNA expressing cell clones. Moreover, the antisense DCC transfected cells displayed a decreased growth rate, a decrease of cells in G1 phase and an accumulation in S phase of the cell cycle. These heavily altered characteristics of MiaPaCa-2 cells expressing DCC antisense RNA point to a yet unknown role for DCC in an important intracellular pathway. PMID- 10530765 TI - Induction of apoptosis in HL-60 cells by eicosapentaenoic acid (EPA) is associated with downregulation of bcl-2 expression. AB - Dietary polyunsaturated fatty acids (PUFAs) have been reported as a potential group of natural products which modulate tumor cell growth. In present study, eicosapentaenoic acid (EPA) was found to inhibit proliferation of human leukemic HL-60 and K-562 cells in vitro. EPA arrested cell cycle progression at G0/G1 phase, and induced necrosis in both HL-60 and K-562 cells. However, EPA induced apoptosis only in HL-60 but not K-562 cells. Also, bcl-2 protein expression was downregulated in much greater extent than that of bax showing that depression of bcl-2 might be an important step during the EPA-induced apoptosis in HL-60 cells. PMID- 10530767 TI - Effect of liquorice and glycyrrhizin on rat liver carcinogen metabolizing enzymes. AB - We investigated the effect of single or repeated intake of conspicuous amounts of licorice root extract (LE, 3138 or 6276 mg/kg body weight (bw) per os) or its natural constituent glycyrrhizin (G, 240 or 480 mg/kg bw per os) on Sprague Dawley rat liver monooxygenases. Whereas a single LE or G dose was unable to affect CYP superfamily, four daily doses induced CYP3A, CYP1A2 and to varying extents CYP2B1-linked monooxygenases. A boosting effect on testosterone 6beta- (CYP3A1/2, CYP1A1/2), 7alpha- (CYP1A1/2, CYP2A1), 16alpha- (CYP2B1, CYP2C11), 2alpha- (CYP2C11) and 2beta- (CYP3A1, CYP1A1) -dependent oxidases as well as on androst-4-ene-3,17-dione- (CYP3A1/2) -supported monooxygenases were also achieved. Harmful outcomes associated to CYP changes (e.g. cotoxicity, cocarcinogenicity and promotion) may be of concern. PMID- 10530766 TI - Combination of paclitaxel and radiation: genotoxicity in vitro in four mammalian cell lines. AB - Paclitaxel is an antimicrotubular agent that blocks the cells in the G2/M phase of the cell cycle. Due to this action, it is presumed that this drug could function as a radiation sensitizer. We studied the genotoxic effects of a combination of paclitaxel and radiation in four mammalian cell lines in the micronucleus assay. The results do not show a clear radiation-sensitizing effect. In the three cell lines, L5178Y, V79 and HeLa, the micronucleus frequencies varied around a theoretical additive effect of the single treatments (paclitaxel or radiation alone). Only the human breast cancer cell line MCF-7 showed consistently lower than additive micronucleus frequencies, although the deviation was not significant. Overall, it remains inconclusive whether paclitaxel exerts a radiosensitizing effect and, if so, whether this effect depends on the cell type or other characteristics of tumor biology. PMID- 10530768 TI - Features of HPV infection among the healthy attendants of gynecological practice in St. Petersburg, Russia. AB - Prevalence of human papillomavirus (HPV) infection was estimated in women from St. Petersburg, Russia. The study included 309 attendants of gynecological practice, who met the following criteria: (1) history of sexual activity; (2) reproductive age; (3) lack of evidence for a specific disease of the genital tract or a current pregnancy; and (4) no cervical abnormalities revealed by cytological examination. Papillomavirus detection was carried out by PCR using MY09/11 primers. Ninety (29%) females turned out to be HPV-positive. HPV presence did not correlate with the current age, age at the sexual debut, or time interval since the first intercourse. However, women with the history of more than two contraceptive abortions had a higher prevalence of papillomavirus infection as compared to the remaining group (30/66 (45%) vs. 56/207 (27%); P = 0.005; OR = 2.25 (1.27-3.97)). HPV genotyping procedure involved reverse dot-blot hybridization and restriction endonuclease analysis. High-risk, low-risk and non identified viruses were detected in 58, 26, and 16% of the positive samples, respectively. HPV16 was the most prevalent type, being present alone in 21% of the infected women, and in combination with other HPVs in 5% of the virus positive females. No other papillomavirus types showed exceptionally prominent prevalence. The data suggest that HPV occurrence among Russian women is within the range of world-wide variations. PMID- 10530770 TI - Inhibitory effects of chlorophyllin on 7,12-dimethylbenz[a]anthracene-induced bacterial mutagenesis and mouse skin carcinogenesis. AB - Chlorophyllin (CHL), a water-soluble derivative of chlorophyll, has been used for the treatment of several abnormal human conditions without apparent toxicity. Recent studies have revealed that CHL has the excellent chemopreventive potential. In the present investigation, we have found the inhibitory activities of CHL against 7,12-dimethylbenz[a]anthracene (DMBA)-induced mutagenesis in Salmonella typhimurium TA100 and also on DMBA-initiated and 12-O-tetradecanoyl phorbol-13-acetate (TPA)-promoted mouse skin tumor formation. The incidence and the multiplicity of skin tumors were not significantly decreased in mice by a single topical application of CHL prior to the DMBA treatment, but there was a marked suppression of papillomagenesis in mice treated with CHL during the promotional stage. Furthermore, the formation of DMBA-induced papillomagenesis was reduced in all mice that had received CHL for 6 weeks following treatment with TPA for 6, 18 and 24 weeks. These results indicate that CHL can inhibit both tumor promotion and the progression of papillomagenesis in the two-stage mouse skin carcinogenesis induced by DMBA and TPA. PMID- 10530769 TI - Interindividual variability in the expression and NNK carbonyl reductase activity of 11beta-hydroxysteroid dehydrogenase 1 in human lung. AB - The balance between metabolic activation and detoxification is critical in determining the susceptibility to lung cancer upon exposure to the tobacco specific nitrosamine, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). Carbonyl reduction of NNK, followed by glucuronidation, is the main detoxification pathway of this lung carcinogen in humans. Recently, we have identified 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD 1) as microsomal NNK carbonyl reductase in liver and lung. In the present study, the interindividual variability of 11beta-HSD 1 expression and NNK-carbonyl reductase activity was examined in human lung by RT-PCR, Western blot analysis and enzyme activity. Levels of 11beta-HSD 1 mRNA varied over an almost 20-fold range among different subjects. Levels of NNK carbonyl reductase activity in lung microsomes closely resembled the relative amounts of immunoreactive protein as determined by Western blot analysis. In view of the large interindividual differences in the susceptibility of tobacco smoke related lung cancer, we present the first data on the variability of 11beta-HSD 1 expression and NNK carbonyl reduction in human lung. PMID- 10530771 TI - Phenotype modulation of cellular UV-sensitivity. AB - Mammalian cell shape is critically important to cell differentiation, apoptosis, cell division, and growth arrest. In the present study we examined the relationship among cell density, cell phenotype (which include shape and coupling) and cell survival using the human A549, H596 and H520 non-small cell lung carcinoma lines. Thus, cells from monolayers, aggregated and suspended cultures at different densities were exposed to UV-radiation and both the density and the phenotype of the cells induce shifts in cellular growth rate. Except in suspended cultures, we observed a UV-sensitivity closely related to the proliferative status of the cells. The variability of the cellular response to UV were investigated taking into account the shape and the coupling potential of the cell lines, suggesting that an intercellular-contact mechanism provides further protection against UV-radiation damage. PMID- 10530772 TI - Serum homocysteine levels in postmenopausal breast cancer patients treated with tamoxifen. AB - Adjuvant treatment of breast cancer with tamoxifen may be associated with reduced risk of cardiovascular disease. Serum homocysteine level has been suggested to be a risk factor for cardiovascular disease influenced by estrogenic hormones. We evaluated a subset of postmenopausal women who had participated in a longitudinal, double-blind, randomized, placebo-controlled toxicity study of tamoxifen 10 mg orally, twice daily. Twenty-seven treated subjects and 37 placebo subjects had measurements of serum homocysteine levels made on previously frozen samples obtained at baseline and after 12 months. After treatment with tamoxifen, we found lower levels of serum homocysteine of borderline statistical significance. PMID- 10530774 TI - Biological differences between reflux stimulated proliferative stomal lesions and N-methyl-N'-nitro-N-nitrosoguanidine induced carcinomas in Wistar rats. AB - The morphology and evolution of epithelial lesions that developed at a gastrojejunal stoma due to reflux of duodenal contents were compared with MNNG induced carcinomas in the pyloric mucosa of rats in a long term experiment. Random bred male Wistar rats were given MNNG in drinking water (100 mg/l) for 12 weeks and then one group was submitted to a gastrojejunal anastomosis at the greater curvature in the oxyntic mucosa. Untreated rats underwent either gastrojejunostomy or gastrotomy. The animals were killed at the 24th and 66th weeks of the experiment. The lesions obtained in the pyloric mucosa and in the mucosa of the gastrojejunal stoma were analyzed histologically using hematoxylin and eosin staining and immunohistochemistry for pepsinogen isoenzyme 1. Duodenal reflux induced proliferative lesions at the gastrojejunal junction that increased in incidence and size with time. Histologically they consisted of benign epithelial proliferation of gastric type. No evidence of malignant transformation within the gastric components of the proliferative lesions at the gastrojejunal stoma was observed even at the 66th week. Adenocarcinomas induced by MNNG in the pyloric mucosa increased in size during the experiment and were morphologically and histochemically distinct from the proliferative lesions at the gastrojejunal junction. In conclusion, proliferative lesions at the gastrojejunal stoma stimulated by duodenal reflux are biologically distinct from adenocarcinomas induced by MNNG in the pyloric mucosa. They do not seem to be precursor lesions of gastric carcinogenesis, as they do not undergo malignant transformation even after long-term, up to 66 weeks, follow-up. PMID- 10530773 TI - Effects of gadolinium ions upon rat prostatic cancer cell lines of markedly different metastatic potential. AB - The effects of gadolinium chloride, a non-specific blocker of mechanosensitive ion channels (MSICs), upon the motility and proliferation of two Dunning rat prostatic tumour cell lines of markedly different metastatic potential were investigated. Gadolinium (2-10 microM) caused a dose-dependent increase in the distance moved in 'wound' assays over a 48-h testing period. The highly metastatic MAT-LyLu cell line was significantly more sensitive to Gd3+, the weakly metastatic AT-2 cells responding only at the highest concentration (10 microM) used. There was no effect on the cells' proliferative rates. These data suggest that mechanosensitive channels could play a role in metastasis by modulating cell migration. PMID- 10530775 TI - Inhibitory activity of nm23-H1 on invasion and colonization of human prostate carcinoma cells is not mediated by its NDP kinase activity. AB - The human nm23-H1 gene product, a putative metastasis suppressor, was identified as nucleoside diphosphate kinase (NDPK) A isoform. To investigate the functional effect of nm23-H1's NDPK activity on its suppression of the components of metastatic phenotype, we transfected a human prostate carcinoma cell line, DU145, with the cDNA encoding nm23-H1 mutant protein lacking NDPK activity. The mutant nm23-H1 transfected cell lines displayed decreased invasiveness and colonization in soft agar as the wild-type nm23-H1 transfectants did when compared with the control transfected line. The results suggest that the metastasis suppressing function of nm23-H1 is independent of the NDPK enzymatic activity. PMID- 10530776 TI - Susceptibility to urethane carcinogenesis of transgenic mice carrying a human prototype c-Ha-ras gene (rasH2 mice) and its modification by butylhydroxytoluene. AB - The susceptibility of rasH2 mice to urethane lung carcinogenesis and the modifying effects of butylhydroxytoluene (BHT) on development of pulmonary lesions were examined. Single i.p. injections of urethane at 250 mg/kg in males or 500 mg/kg in females induced alveolar/bronchiolar adenomas within 6 weeks. At 4 weeks after the injection with a dose of 1000 mg/kg, adenomas occurred in both sexes. BHT administration increased the multiplicity of hyperplasias observed 3 weeks after the urethane injection and additionally caused adenomas which did not occur in the urethane alone-treated animals. The overall data suggest the possibility of rapid assays for lung carcinogens using rasH2 mice. PMID- 10530778 TI - Elimination of Na+-dependent bile acid transporter from small intestine by ileum resection increases [correction of increase] colonic tumorigenesis in the rat fed deoxycholic acid. AB - Ileal Na+-dependent bile acid transporter (ISBT) constituting a gateway to enterohepatic circulation of bile acids occurs exclusively at the distal site of the small intestine. In the present study, we examined colonic tumorigenesis promoted by deoxycholic acid in relation to the expression of the ISBT. For this purpose, the small intestine of a Fischer-344 rat was resected a length of 20 cm above the ileo-cecal valve (ileal resection) or below the duodenum (jejunal resection). Then, rats were treated with an intraperitoneal injection of azoxymethane (15 mg/kg body wt.) once a week for 3 weeks and fed a 20% casein diet supplemented with 0.2% deoxycholate for 39 weeks. Northern blot analysis demonstrated that the ISBT mRNA was hardly detectable in ileum-resected rats. The excretion of fecal bile acids was 1.5-fold higher in the ileum-resected group than in the jejunum-resected group (P < 0.05). On the contrary, the serum bile acids concentration of ileal-resected rats was about one-half of that of jejunum resected animals (P < 0.05). The tumor incidence and the total tumor number were significantly higher in the ileum-resected group than in the jejunum-resected one (P < 0.05). Interestingly, no tumor was found at the proximal colon in the jejunum-resected group while tumors developed frequently at the proximal site as well as mid and distal colon in the ileum-resected group. These observations demonstrate that malabsorption of bile acids owing to the lack of ISBT enhanced colon tumorigenesis. PMID- 10530777 TI - Anti-angiogenic activity of a novel synthetic agent, 9alpha fluoromedroxyprogesterone acetate. AB - 9Alpha-fluoromedroxyprogesterone acetate (FMPA) is a novel synthetic analog of medroxyprogesterone acetate (MPA), widely used as therapeutic agent for breast and endometrium cancers. FMPA showed almost the same binding affinities to the progesterone and glucocorticoid receptors as MPA. In the rabbit corneal assay, FMPA, MPA and fumagillin significantly inhibited the angiogenic response induced by rat mammary tumor at doses of 0. 1, 1 and 50 microg/pellet, respectively, so FMPA showed greater anti-angiogenic activity than MPA and fumagillin. In the mouse dorsal air sac method, FMPA inhibited the mouse sarcoma 180 cell-induced angiogenesis by oral administration at a dose of 200 mg/kg. FMPA inhibited the activity of plasminogen activator (PA) in bovine endothelial cells. These results suggest that FMPA may be useful for diseases associated with angiogenesis by oral administration. PMID- 10530779 TI - Augmentation of antitumor activity of 5'-deoxy-5-fluorouridine by thymosin fraction 5 in mouse bladder cancer cells in vitro and in vivo. AB - 5'-Deoxy-5-fluorouridine (5'-dFUrd) is a prodrug of 5-fluorouracil (5-FUra) activated by pyrimidine nucleoside phosphorylase (PyN Pase), mainly by uridine phosphorylase (Urd Pase) in rodents and by thymidine phosphorylase (TdR Pase) in humans, which is preferentially located in tumor tissues compared to normal tissues. It has been reported that PyN Pase is induced by cytokines such as tumor necrosis factor (TNF), interleukin-1alpha (IL-1alpha) and interferon (IFN). Thymosin is a glycoprotein extract obtained from the calf thymus and is a potent immunopotentiating preparation. In this study, the antiproliferative activity of 5'-dFUrd used in combination with thymosin fraction 5 (TF5) was investigated in mouse bladder cancer cell line MBT-2 in vitro and in vivo. In vitro TF5 enhanced the activity of 5'-dFUrd by up to 4.11-fold, whereas the activity of other cytostatics such as 5-FUra, mitomycin C, adriamycin, cis-platinum, etoposide, vinblastine and methotrexate was not changed. In vivo when the effects of combination therapy with 5'-dFUrd and TF5 in C3H/HeN mice implanted with MBT2 were studied, tumor growth was not suppressed by TF5 alone while tumor growth was suppressed to some degree by 5'-dFUrd alone. However, tumor growth suppression was enhanced when 5'-dFUrd was used in combination with TF5. In order to investigate this mechanism, Urd Pase in MB2 was measured, and it was found that TF5 increased enzyme activity by up to 1.8-fold in MBT2. This increased susceptibility might be a result of the induction of Urd Pase, which is the essential enzyme for the conversion of 5'-dFUrd to 5-FUra. These results suggested that the therapeutic benefit of 5'-dFUrd would be improved by its use in combination with TF5 and the modulation of converting enzymes for antitumor prodrugs could be a novel therapeutic strategy for treating human cancers. PMID- 10530781 TI - Expression of mucins and cytokeratins in ovarian cancer cell lines. AB - The expression pattern of the epithelial cell markers MUC1 (CA15-3, EMA), CA125 (OC125), human epithelial antigen HEA (Ber-EP4) and cytokeratins (Ck7, Ck8, Ck7/8, Ck8/18/19) was studied in seven human ovarian cancer cell lines. We analyzed the cell lines by immunofluorescence to determine the surface as well as cytoplasmic expression. Furthermore, we evaluated the mRNA expression of MUC1, Ck18 and Ck19 by reverse transcriptase-polymerase chain reaction (RT-PCR). All cell lines were positive for MUC1. However, expression patterns and staining intensity depended on the different epitope-specific antibodies. CA125, a typical serum marker for ovarian carcinomas, was positive only in two cell lines. HEA was strongly positive in three cell lines, whereas the others expressed the antigen only weakly in the cytoplasm. Ck7 was not expressed in three of the seven cell lines. Ck7/8 was detectable in all cell lines and was strongly expressed in four of them. MUC1 mRNA was expressed by all cell lines as detected by RT-PCR. These findings permit selection of a suitable marker for the detection of disseminated ovarian cancer cells. PMID- 10530780 TI - Peroxiredoxin I expression in human thyroid tumors. AB - Peroxiredoxin I (Prx I) is newly discovered oxidative stress inducible protein, having a thioredoxin peroxidase activity. The Prx I expression level in 107 samples out of 60 thyroid lesions, including normal thyroid, tumors and thyroiditis including Graves' disease were examined using immunoblotting. Prx I expression levels in follicular neoplasm (P = 0.00005) and thyroiditis group (P = 0.0037) were significantly higher than that of the control group, while papillary carcinoma group did not show statistical significance. Immunohistochemistry indicated that Prx I was in epithelial cells of thyroid follicles. These results suggest that Prx I is expected to be a candidate for novel tumor markers to discriminate tissue types of tumors. PMID- 10530784 TI - Genetics of aging in Drosophila. AB - The genetics of aging in Drosophila are reviewed under the separate headings of population genetics, physiological genetics, and molecular genetics. However, connections between these sub-fields are brought forward for discussion. PMID- 10530782 TI - Carcinogenic effects of N-ethyl-N-hydroxyethylnitrosamine and its metabolites in rats and mice. AB - N-ethyl-N-hydroxyethylnitrosamine (EHEN), a member of the nitrosamine class of carcinogens induces renal cancer. However, since very little is known about the metabolic products of EHEN and their effects, these were investigated in rats and mice. EHEN, N-ethyl-N-formylmethylnitrosamine (EFMN) and N-ethyl-N-carboxymethyl nitrosamine (ECMN) were administered in the drinking water for 2 weeks and the animals were then maintained until sacrifice at week 32. The urine of the rats was collected over the 2-week exposure period and analyzed by HPLC. The results showed that EHEN but not EFMN or ECMN induces tumors in the kidneys of rats. In mice the lungs were targeted not only by the parent compound but also by both metabolites. The findings suggest that the kidney is the most susceptible organ to EHEN effects in the rat while the lung is the most susceptible organ in mice. These results are consistent with inter-species variation in the metabolism of xenobiotics. PMID- 10530783 TI - Inducible nitric oxide synthase inhibitor of the Chinese herb I. Saposhnikovia divaricata (Turcz.) Schischk. AB - L-Arginine derived nitric oxide (NO) and its derivatives, such as nitrogen dioxide and peroxynitrite, play a role in inflammation and also possibly in the multistage process of carcinogenesis. Four furanocoumarins and eight chromones isolated from the dried root of Saposhnikovia divaricata (Fang Feng in Chinese) and evaluated for their effects on the synthesis of NO induced by lipopolysaccharide (LPS) in macrophage cell line RAW 264.7. The inhibition of nitrite production, as an index for NO released from the macrophage cells, was quantitatively analyzed by Griess reaction. The results showed that imperatorin and deltoin are potential NO production inhibitor, and their IC50 values for inhibition of nitrite production were 17.3 and 11.6 microg/ml, respectively. Western-blot analysis demonstrated that iNOS enzyme activity was not inhibited by treatment with imperatorin or deltoin, but revealed that both compounds inhibited the expression of the iNOS protein. PMID- 10530786 TI - The mitochondrial theory of aging: do damaged mitochondria accumulate by delayed degradation? AB - The mitochondrial theory of aging states that the slow accumulation of impaired mitochondria is the driving force of the aging process. In recent years, this theory has gained new support with the discovery of age-related mitochondrial DNA deletions. However, the underlying mechanism of the accumulation of defective mitochondria remained unclear. This has changed recently with the proposal of de Grey that damaged mitochondria have a decreased degradation rate. The resulting increase in biological half-life would be a strong selection advantage leading to the accumulation of defective mitochondria. In this article, I summarize current ideas on how damaged organelles can build up in a cell as well as the shortcomings of these ideas. Then the new hypothesis and its justification are described. It appears that de Grey's hypothesis is a very promising concept that elegantly solves inconsistencies of current models and is in accordance with experimental findings. PMID- 10530787 TI - The mitochondrial theory of aging: is the culprit a faulty disposal system rather than indigenous mitochondrial alterations? AB - Mitochondrial damage and the proportion of effete mitochondria in cells increase with age. According to the mitochondrial theory of aging, this phenomenon is mostly due to oxidative damage and is a major (and, some argue, the main) determinant of aging. It will be argued briefly that this phenomenon plays a role that is not exclusively crucial in aging. It will also be contended, essentially on theoretical grounds (for lack of sufficient current information), that there is low probability that the accumulation of reduced degradation of affected mitochondria is due to diminished production of hydroxyl radicals, as suggested by Aubrey and de Grey (1997) and expanded by Kowald (in this issue). What seems more likely is that the phagolysosomal disposal system of effete mitochondria is considerably altered in cells of aging organisms. Also, in view of the significant role of damaged mitochondria in the initial steps in apoptosis and the lack of evidence of massive apoptosis of cells in senescent individuals, the damage that exists may be milder than anticipated by the mitochondrial theory of aging. A brief fundamental summary on the biology of mitochondria is included for the sake of better understanding the arguments presented in this article. Also, suggestions are made for experimental testing of the hypotheses presented by Aubrey and de Grey (1997) and Kowald (1999). PMID- 10530785 TI - Theoretical basis for the benefit of postmenopausal estrogen substitution. AB - Women are being presented with an increasing number of choices for health care management as they move through the aging process. Estrogen has positive effects on mood, sexual function, target end organs and cognitive function, and may play an important role in the etiology of Alzheimer's Disease by acting to prevent amyloid plaque formation, oxidative stress, or deterioration of the cholinergic neurotransmitter system. The benefits of estrogen therapy for osteoporosis, the cardiovascular system, and lipid metabolism are far reaching, but the possibility of developing breast cancer later in life is also relevant. Understanding the mechanisms for the action of the estrogens, anti-estrogens, and the selective estrogen receptor modulators, and possible alternative routes of symptom management for some menopausal events is important to make appropriate decisions on choice of therapy. This review discusses the theoretical basis for estrogen's actions in the management of the postmenopausal stage of the life cycle. PMID- 10530788 TI - Differences in locomotor activity across the lifespan of Drosophila melanogaster. AB - The identification of differential patterns of change across the lifespan in quantitative traits is of abiding interest in the biological and gerontological research communities. These differential phenotypic patterns in complex systems illuminate developmental processes and form the foundation for the identification of putative biomarkers of aging. The goal of the present study was to explore changes in locomotor activity through the lifespan of Drosophila melanogaster. A replicated serial cross-sectional sampling design was used to test activity in five genetically independent inbred strains at 7, 14, 21, 28, 35, and 42 days of age. Differences were observed in activity level across ages and strains, suggesting that patterns of activity throughout the lifespan of D. melanogaster are influenced by genetic factors. Observed sex differences in change in activity level indicate that the aging processes may proceed differently in males and females. PMID- 10530789 TI - Resistance to apoptosis in human CD8+ T cells that reach replicative senescence after multiple rounds of antigen-specific proliferation. AB - We have established an in vitro culture model of cellular aging in which antigen specific T cells are stimulated repeatedly to divide until they reach the irreversible state of growth arrest known as "replicative senescence." T lymphocytes that reach replicative senescence in culture show complete loss of CD28 expression, shortened telomeres, undetectable telomerase, and reduced ability to produce heat shock proteins. We now document that in response to treatment with apoptotic stimuli, senescent CD8+ T-cell cultures show reduced apoptosis and diminished caspase 3 activity compared with quiescent early passage cultures from the same donor. Our results suggest that the progressive accumulation of T cells showing many of the hallmarks of replicative senescence during aging, chronic infection, and autoimmune disease may, in part, reflect the diminished capacity of such cells to undergo normal programmed cell death. PMID- 10530792 TI - Effects in vitro of several antioxidants on the natural killer function of aging mice. AB - The aim of the present work is to study the change with aging in the effect in vitro of several antioxidants: thiazolidine-4-carboxylic acid or thioproline, N acetylcysteine (NAC), ascorbic acid (AA), and alpha-tocopherol (vitamin E, VE) on the natural killer (NK) activity in mononuclear cells from axillary nodes, spleen, thymus and peritoneal leukocytes from BALB/c male mice. Young (8+/-2 weeks), adult (24+/-2 weeks). mature (48+/-2 weeks), and old (72+/-2 weeks) animals were studied. A nonradioactive cytotoxic assay with cells from the murine lymphoma YAC-1 as target cells and a relation effector cells/target cells of 10/1 were used. The concentrations of the different antioxidants were: 1 mM for thioproline and N-acetylcysteine and 5 microM for ascorbic acid and alpha tocopherol, which induced a maximum effect in our previous dose-response experiments. The results show that, in general, the above antioxidants cause an enhancement of the NK activity at all ages studied, this stimulation being higher with thioproline and N-acetylcysteine than with ascorbic acid and alpha tocopherol. The effects were similar for the three lymphoid organs and the peritoneum. This stimulation of the NK activity by antioxidants is an important favorable response, especially in old mice, in which age results in a decrease in NK function and, therefore, in a higher incidence of neoplasia. PMID- 10530793 TI - Effect of aging on plasma membrane fluidity of rat aortic endothelial cells. AB - To assess age-related changes in the physical properties of vascular endothelial cell (EC) plasma membranes, we measured membrane fluidity with 1,6-diphenyl-1,3,5 hexatriene (DPH), 1-(4-trimethylammonium-phenyl)-6-phenyl-1,3,5-hexatriene, and 10-(1-pyrene)dodecanoic acid, and investigated the parameters affecting membrane fluidity of endothelial cells (ECs) cultured from the thoracic aortas of young (5 week-old) and aged (100-week-old) rats. Plasma membrane fluidity of aged rat ECs was significantly lower than that of young rat ECs, as assessed by increased 1,6 diphenyl-1,3,5-hexatriene fluorescence polarization and by decreased pyrene excimer formation, although 1-(4-trimethylammonium-phenyl)-6-phenyl-1,3,5 hexatriene did not demonstrate a change in membrane fluidity with aging. Compared with those in young rat ECs, cholesterol concentrations in aged rat ECs were significantly higher, whereas phospholipid concentrations were unchanged; consequently, the cholesterol/phospholipid molar ratio was significantly higher in aged rat ECs. Lipid peroxide levels measured with thiobarbituric acid reactive substances were significantly higher in EC plasma membranes of aged rats. These results indicate that age-related increases in cholesterol and lipid peroxide in vascular EC plasma membranes reduce membrane fluidity. PMID- 10530791 TI - Age-related changes in the expression of CD95 (APO1/FAS) on blood lymphocytes. AB - Aging is associated with alterations of the immune system, thought to be related to an increased susceptibility to infectious diseases, and possibly to cancer and autoimmunity in the elderly. In the present paper we report data obtained on freshly collected blood from 148 healthy subjects of different ages (from cord blood to 102 years old). The subjects were divided into seven age classes (cord blood, 3-11 years, 15-39 years, 41-60 years, 61-74 years, 75-84 years, 85-102 years) and their lymphocyte subsets and the expression of the apoptosis-related molecule CD95 were evaluated. In respect of lymphocyte subsets, the major differences were found in the cord-blood samples compared with the oldest old groups. In the cord-blood group, the absolute number of all the lymphocyte subsets was enhanced, but in the oldest group, an increase of CD16+ lymphocytes was observed, whereas CD19+ lymphocytes, which progressively decrease with age, continue to decrease further in the very old. The data show that the expression of CD95 increases until age 74 years, whereas in the oldest old it tends to decrease again. The trend of CD95 expression seems to be related to the change of expression of CD95 on CD4+ lymphocytes, because the CD8+/CD95+ population rose steadily throughout the entire age range. The evaluation of CD95+/CD45R0+ lymphocytes shows similar results to those observed analyzing CD95 on total lymphocytes. Furthermore, a constant increase of CD95+/CD28+ and a related decline of CD28+ lymphocytes was observed in all age groups. These data suggest that the expression of CD95 on the different subsets of lymphocytes can be considered a good marker for studies of immunosenescence, because it may be predictive of successful aging, and can partially explain the change in lymphocytes subsets in elderly. PMID- 10530790 TI - Effect of age on DNA binding of the ku protein in irradiated human peripheral blood mononuclear cells (PBMC). AB - DNA binding of the ku protein was investigated in peripheral blood mononuclear cells (PBMC) from 24 subjects of different ages (20-89 years old) displaying age related changes in DNA repair, mitotic responsiveness, and cytokine production. Ku is an heterodimeric protein composed of two subunits of 70 and 80 kDa, which is involved in the earliest steps of DNA damage recognition. DNA binding of ku 70/80 was found unchanged in normal PBMC from aging subjects but progressively declined in x-ray-irradiated PBMC from young to adult, and elderly subjects. This finding was concomitant with the age-related fall of DNA repair in the whole population. PMID- 10530794 TI - Ischemia-reperfusion in the adult mouse heart influence of age. AB - Age-related changes in the mouse heart after ischemia-reperfusion have not been well characterized. To test the hypothesis that advanced age was associated with increased susceptibility to myocardial injury after ischemia/reperfusion, we studied the hearts of young adult and old mice. In young adult (6-8 months) and aged (22-24 months) C57 BL/6 mice, we performed left anterior descending coronary artery ligation and subjected the hearts to 45 min of ischemia followed by varying periods of reperfusion of 15 min, 1 h, 4 h, and 24 h. We found that there was a significant age difference in the size of the infarct between the young adult and old hearts. There was also greater damage in the old hearts in terms of contraction band necrosis, myofiber tears, DNA fragmentation, and mitochondrial disruption. Thus, the old heart is more susceptible to injury after ischemia reperfusion. This may be partly due to an age-associated decrease in coronary circulation and collateral flow, as well as other factors. PMID- 10530795 TI - Occurrence and expansion of trisomy 7 in a fibroblast strain from a centenarian individual. AB - We describe the presence of metaphases with non-random gain of one or two chromosomes in a skin fibroblast strain derived from a centenarian individual. The extra elements were chromosomes 7, X, and 18, and, among these, the most frequent was a 7. During in vitro propagation +7 cells seemed to be stable and overrode the diploid ones. After prolonged growth in culture, the cell population displayed the typical senescence signs. Our findings confirm the proneness to aneuploidy in cells from aged individuals and indicate that, while the presence of a trisomic 7 may confer a selective advantage to cells grown in vitro, it does not seem to prevent cellular senescence. PMID- 10530796 TI - A model for persistent infection with Epstein-Barr virus: the stealth virus of human B cells. AB - Most adult humans are infected benignly and for life with the herpesvirus Epstein Barr virus. EBV has been a focus of research because of its status as a candidate tumor virus for a number of lymphomas and carcinomas. In vitro EBV has the ability to establish a latent infection in proliferating B lymphoblasts. This is the only system available for studying human herpesvirus latency in culture and has been extremely useful for elucidating how EBV promotes cellular growth. However, to understand how EBV survives in the healthy host and what goes awry, leading to disease, it is essential to know how EBV establishes and maintains a persistent infection in vivo. Early studies on the mechanism of EBV persistence produced inconclusive and often contradictory results because the techniques available were crude and insensitive. Recent advances in PCR technology and the application of sophisticated cell fractionation techniques have now provided new insights into the behavior of the virus. Most dramatically it has been shown that EBV in vivo does not establish latency in a proliferating lymphoblast, but in a resting memory B cell. The contrasting behaviors of being able to establish a latent infection in proliferating B blasts and resting memory B cells can be resolved in terms of a model where EBV performs its complete life cycle in B lymphocytes. The virus achieves this not by disrupting normal B cell biology but by using it. PMID- 10530797 TI - Paradoxical effect of neuroleptic drugs on prolactin secretion by rat pituitary cell cultures. AB - Several antipsychotic drugs reverse the dopamine-induced inhibition of prolactin release by rat pituitary cell cultures. Paradoxically, at high doses and without dopamine, antipsychotic drugs can also inhibit prolactin secretion. The mechanism underlying this phenomenon is unclear. Some evidence suggests that these drugs have an agonistic action. We sought to verify whether clozapine and fluphenazine, at doses higher than those reversing dopamine-induced inhibition of prolactin secretion in vitro, show this paradoxical effect and eventually a partial agonistic action. Both antipsychotics inhibited prolactin secretion, clozapine at doses starting from 10(-6) M and fluphenazine from 10(-7) M. Haloperidol reversed clozapine-induced prolactin inhibition but left fluphenazine-induced inhibition unchanged. These in vitro findings suggest that clozapine has a partial agonistic action on dopaminergic receptors but fluphenazine does not. PMID- 10530798 TI - Platelet activation and modulation of the induction of nitric oxide synthase in the conscious rat. AB - Injection of lipopolysaccharide (LPS) (Salmonella W. Typhosa i.v. bolus) into conscious rats, induced a rapid drop of circulating platelets analogous to that induced by ADP. The animals showed a small fall in mean arterial blood pressure (MABP), an increase in heart rate and a significant increase in plasma nitrite and nitrate level. This result is consistent with the stimulation of an inducible NO synthase (i-NOS). The administration of the stable prostacyclin analogue, iloprost plus ADP or LPS, significantly protected against the decrease in free platelet number induced by ADP or LPS. The plasma nitrite and nitrate level stimulated by LPS was significantly reduced by iloprost and also by prostacyclin. These results are consistent with an inhibition of i-NOS by agents that increase the intracellular level of cAMP. The administration of the NO donor S-Nitroso-N acyl-D-penicillamine (SNAP) plus ADP or LPS, significantly prevented thrombocytopenia induced by ADP and by LPS. SNAP did not decrease the plasma nitrite and nitrate level stimulated by LPS; furthermore it induced a significant increase of heart rate, without affecting MABP, suggesting a direct accelerating effect of NO on the sino-atrial node. The administration of S-nitroso-glutathione (GSNO), a stable nitrosothiol, plus ADP or LPS, significantly prevented thrombocytopenia induced by ADP but not by LPS. GSNO significantly reduced the plasma nitrite and nitrate level stimulated by LPS. These data demonstrate that the L-Arginine: NO pathway in vivo may be modulated by prostanoids and that compounds which increase cAMP, such as iloprost, are able to protect against LPS induced early thrombocytopenia. PMID- 10530799 TI - Neurotrophic effect of isoquinoline derivatives in primary cortical culture. AB - Recent studies indicate that the N-methyl-D-aspartate (NMDA) antagonist, (+)-1 methyl-1-phenyl-1,2,3,4-tetrahydroisoquinoline hydrochloride (FR 115427), enhanced neuronal survival in primary culture of cortical neurons from mouse embryos. In the present study isoquinoline derivatives were examined for the neurotrophic activity in primary culture of cortical neurons and were also examined for anti-NMDA activity. In spite of varying level of anti-NMDA activity, isoquinoline derivatives enhanced neuronal survival at the concentration of 10 microM. To elucidate of the mechanisms of neurotrophic activity in primary cortical culture, nicardipine and flunarizine, known calcium channel blockers, were also tested. Neither nicardipine nor flunarizine showed neurotrophic activity up to the doses causing toxicity in cultured neurons. NBQX, an AMPA receptor antagonist, was also tested for neurotrophic activity. However no enhancement of neuronal survival was observed. These data suggest that one of the mechanisms to promote neuronal survival may depend on the structure of isoquinoline ring. Moreover neurotrophic activity observed in our culture systems might not relate on anti-NMDA activity, blockade of voltage dependent L-type calcium channels and antagonization of AMPA receptor. PMID- 10530800 TI - Preparation of highly purified momordin II without ribonuclease activity. AB - Momordin II, a ribosome-inactivating protein from Momordica charantia seeds, was purified by a procedure involving a series of chromatographies on S-Sepharose, Sephadex G-50, CM-Sepharose, and Red Sepharose columns. Highly purified momordin II inhibited cell-free protein synthesis, released adenine from rat liver ribosomes and from DNA, and had no RNase activity. PMID- 10530802 TI - Effects of a novel Mac-1 inhibitor, NPC 15669, on hemostatic parameters during preconditioned myocardial infarction. AB - NPC 15669, a member of the leumedins family, inhibits leukocyte adhesion to the endothelium by blockage of upregulation of a member of beta2 integrin family Mac 1 (CD11b/CD18). Inhibition of neutrophil-endothelial interactions may alter the course of myocardial reperfusion injury. However, the effects of NPC 15669 supplementation on the hemostatic profile during ischemia-reperfusion are unknown. The aim of the present study was to define changes in the certain hemostatic factors in the natural course of preconditioned myocardial infarction. Twelve consecutive Yorkshire swine underwent myocardial stunning (8 min. left anterior descending artery occlusion followed by 90 min. of reperfusion) and then preconditioned myocardial infarction (50 min. occlusion followed by 3 hours of reperfusion) experiments. NPC 15669 (10 mg/kg loading dose followed by constant infusion at 6 mg x kg(-1) x h(-1)) was administered in 6 animals; another 6 swine received saline and served as controls. Blood samples were obtained at baseline, twice during occlusion; and three times during reperfusion. The levels of antithrombin-III, Protein C, total Protein S, fibronectin, endothelin-1, as well as the stable metabolites of thromboxane (TxB2) and prostacyclin (6-keto-PGF1a), were determined. NPC 15669 treatment was associated with diminished endothelin-1, TxB2 levels and increased fibronectin, 6-keto-PGF1a, Protein C and total Protein S concentrations in the setting of preconditioned myocardial infarction. There were no changes in the plasma concentrations of antithrombin-III in NPC 15669 group when compared with controls. The increase in Protein C, total Protein S, and 6-keto-PGF1a (favoring antithrombosis), and decrease in endothelin-1 and TxB2 levels (favoring vasodilatation), following NPC 15669 may explain the reduction in infarct size previously reported with this agent. PMID- 10530801 TI - Effect of NK-104, a new synthetic HMG-CoA reductase inhibitor, on triglyceride secretion and fatty acid oxidation in rat liver. AB - For the investigation of the mechanism responsible for the hypotriglyceridemic effect of NK-104, a new synthetic inhibitor of HMG-CoA reductase, the rate limiting enzyme for cholesterol synthesis, isolated rat liver was perfused with or without NK-104 in the presence of exogenous [1-(14)C]oleic acid substrate. Addition of NK-104 tended to increase the ketone body production while it caused a significant decrease in the secretion rate of triglyceride by the perfused liver without affecting uptake of exogenous [1-(14)C]oleic acid. The inhibitor also significantly decreased hepatic triglyceride concentration. The altered triglyceride secretion was accompanied by a concomitant decreased incorporation of exogenous [1-(14)C]oleate into triglyceride. The conversion of exogenous [1 (14)C]oleic acid substrate indicated an inverse relationship between the pathways of oxidation and esterification. No effect of NK-104 on hepatic secretion of cholesterol was observed. These results suggest that NK-104 exerts its hypotriglyceridemic action, primarily by diverting the exogenous free fatty acid to the pathways of oxidation at the expense of esterification. PMID- 10530803 TI - Nitric oxide-cyclic GMP system potentiates glucose-induced rise in cytosolic Ca2+ concentration in rat pancreatic beta-cells. AB - Involvement of nitric oxide (NO) in the regulation of insulin secretion from pancreatic beta-cells was investigated by measuring cytosolic Ca2+ concentration ([Ca2+]i) in isolated rat pancreatic beta-cells. At 7.0 mM glucose, L-arginine (0.1 mM) elevated [Ca2+]i in about 50% of the beta-cells examined. The response was partially inhibited by an NO synthase inhibitor, N(G)-monomethyl-L-arginine (L-NMA; 0.1 mM), suggesting that part of the response was mediated by the production of NO from L-arginine. D-Arginine at higher concentrations (3 or 10 mM) also increased [Ca2+]i at 7.0 mM glucose; however, the response was not affected by L-NMA (0.1 mM). Similar [Ca2+]i elevation was produced by NO (10 nM) and sodium nitroprusside (SNP; 10 microM) at 7.0 mM glucose. The SNP-induced increase in [Ca2+]i was abolished by nicardipine (1 microM), suggesting that the [Ca2+]i response is mediated by Ca2+ influx through L-type voltage-operated Ca2+ channels. In the presence of oxyhemoglobin (1 microM), the [Ca2+]i elevation induced by NO (10 nM) was abolished. Neither degradation products of NO, NO2- nor NO3-, caused any changes in [Ca2+]i. 8-Bromo-cyclic GMP (8-Br-cGMP; 3 mM) and atrial natriuretic peptide (0.1 microM) elevated [Ca2+]i at 7.0 mM glucose. We conclude that NO, which is produced from L-arginine in pancreatic islets, facilitates glucose-induced [Ca2+]i increase via the elevation of cGMP in rat pancreatic beta-cells. NO-cGMP system may physiologically regulate insulin secretion from pancreatic beta-cells. PMID- 10530804 TI - Reduced uptake and enhanced release of cadmium in cadmium-resistant metallothionein null fibroblasts. AB - Metallothionein (MT) is known to play a predominant role in the protection of cells from cadmium (Cd) toxicity. To investigate other factors involved in Cd resistance, we established Cd-resistant cell lines from simian virus 40 transformed MT null fibroblasts. Cd-resistant MT null cells, Cd-rA7 and Cd-rB5, developed approximately 10-fold resistance to Cd compared to parental cells, but showed no cross-resistance to Zn, Cu, Hg, Ni, As, cisplatin or H2O2. Accumulation of Cd in the resistant cells was 13-18% of that of parental cells after treatment with Cd for 24 h. A short-term experiment revealed that the rate of Cd incorporation into the Cd-resistant cells was suppressed, and the rate of Cd release was enhanced in the resistant cells compared with that of parental cells. These results indicate that the altered transport of Cd, slow uptake and rapid release, may confer resistance to Cd on the Cd-resistant cells established from MT null fibroblasts. PMID- 10530805 TI - In human and rat lung membranes [35S]GTPgammaS binding is a tool for pharmacological characterization of G protein-coupled dinucleotide receptors. AB - The P2Y receptor family is activated by extracellular nucleotides such as ATP and UTP. P2Y receptors regulate physiological functions in numerous cell types. In lung, the P2Y2 receptor subtype plays a role in controlling Cl- and fluid transport. Besides ATP or UTP, also diadenosine tetraphosphate (Ap4A), a stable nucleotide, seems to be of physiological importance. In membrane preparations from human and rat lung we applied several diadenosine polyphosphates to investigate whether they act as agonists for G protein-coupled receptors. We assessed this by determining the stimulation of [35S]GTPgammaS binding. Stimulation of [35S]GTPgammaS binding to G proteins has already been successfully applied to elucidate agonist binding to various G protein-coupled receptors. Ap(n)A (n = 2 to 6) enhanced [35S]GTPgammaS binding similarly in human and rat lung membranes, an indication of the existence of G protein-coupled receptor binding sites specific for diadenosine polyphosphates. Moreover, in both human and rat lung membranes comparable pharmacological properties were found for a diadenosine polyphosphate ([3H]Ap4A) binding site. The affinity for Ap2A, Ap3A, Ap4A, Ap5A, and Ap6A was also comparable. 8-Diazido-Ap4A and ATP were less potent, whereas the pyrimidine nucleotide UTP showed hardly any affinity. Thus, we present evidence that different diadenosine polyphosphates bind to a common G protein-coupled receptor binding site in membranes derived either from human or rat lung. PMID- 10530806 TI - Long-term administration of GHB does not affect muscular mass in alcoholics. AB - Gamma-hydroxybutyric acid (GHB) is a drug recently utilized for alcoholism management. It has been shown that GHB has anabolic effects since it can increase growth hormone (GH) release in healthy subjects. At present, there are no studies investigating body composition in alcoholics during long-term GHB treatment. In this study body composition and GH secretion in alcohol dependent subjects was evaluated during addiction and at different time of GHB administration and alcohol abstinence. A total of 45 male alcohol dependent patients (mean age 39.7+/-9.8 yrs, mean height: 171+/-6.8 cm, body mass index--BMI--22.1+/-1.6 kg/m( 2)) were consecutively enrolled. Body composition was assessed by anthropometry, bioimpedance analysis and tritiated water method. A 7-day food diary was collected. Plasma GH levels were determined by radioimmunoassay. A 6-month total abstinence was obtained in 22 patients, by means of psychological support counseling and self-help groups in 9 subjects and also by 50 mg/kg/day of GHB in 13 subjects. At 1, 2, 3 and 6 months of abstinence, the biochemical assessment and metabolic variables were re-examined. Fat-free mass (FFM) and basal GH secretion were similar at the different times of follow-up in both groups of patients. GHB treated patients and those receiving psychological support alone showed similar values in FFM and GH. Both groups of patients did not differ in FFM and plasma GH level from healthy controls at any of the times evaluated. Waist-to-hip ratio did not differ between patient groups, while higher values were shown in alcoholics with respect to control subjects. The present study shows that long-term administration of GHB did not affect muscular mass and did not determine an increase of GH release in chronic alcoholics. This findings could be due to an impairment of the hypothalamic-limbic system and of GABAergic neurotransmission in alcoholics' brain. PMID- 10530807 TI - Sustained QT prolongation induced by tacrolimus in guinea pigs. AB - Recently, clinical cases have been reported of QT prolongation and torsades de pointes associated with the use of tacrolimus (FK506). We examined the relationship between QTc prolongation and the pharmacokinetics of FK506 in guinea pigs in order to evaluate the arrhythmogenicity of FK506 in comparison with quinidine (QND). FK506 (0.1 or 0.01 mg/hr/kg) or QND (30 mg/hr/kg) was intravenously infused to guinea pigs and time profiles of drug concentration in blood and QTc interval were examined during and after infusion. Both FK506 and QND evoked a significant QTc prolongation, and the dose-response relationship showed an anti-clockwise hysteresis, FK506-induced QTc prolongation persisted throughout the duration of the experiment despite a decline in the plasma FK506 concentration, whilst QND-induced QTc prolongation disappeared as plasma concentrations decreased. FK506 induced a sustained QTc prolongation in guinea pigs at drug concentrations in blood that correspond to its therapeutic range in human, suggesting that it might be of clinical significance to monitor the electrocardiogram, especially when patients have congenital or acquired QT prolonging risk factors. PMID- 10530808 TI - Pharmacological agents acting at subtypes of metabotropic glutamate receptors. AB - Metabotropic (G-protein-coupled) glutamate (mGlu) receptors have now emerged as a recognized, but still relatively new area of excitatory amino acid research. Current understanding of the roles and involvement of mGlu receptor subtypes in physiological/pathophysiological functions of the central nervous system has been recently propelled by the emergence of various structurally novel, potent, and mGlu receptor selective pharmacological agents. This article reviews the evolution of pharmacological agents that have been reported to target mGlu receptors, with a focus on the known receptor subtype selectivities of current agents. PMID- 10530810 TI - Inhibition of N-linked glycosylation of the human type 1alpha metabotropic glutamate receptor by tunicamycin: effects on cell-surface receptor expression and function. AB - The potential role of N-linked glycosylation of the human type 1alpha metabotropic glutamate (mGlu1alpha) receptor was studied in a recombinant, inducible expression system, where receptor expression was induced in the absence and presence of tunicamycin. In the absence of tunicamycin the mGlu1alpha receptor appeared to be expressed, at least in part, as a dimer consisting of monomers of approx. 145 and 160 KDa relative molecular mass (Mr). In the presence of tunicamycin only a single monomeric protein could be detected approximating the Mr predicted for the human mGlu1alpha receptor based on its primary amino acid sequence (130 KDa). Exposure to tunicamycin during receptor induction did not appear to affect the cell surface expression of the mGlu1alpha receptor as determined immunocytochemically or using a cell-surface biotinylation strategy, but reduced agonist-stimulated phosphoinositide hydrolysis by approximately 50% compared to control cell populations. Our data suggest that non-N-glycosylated human mGlu1alpha receptors can traffic to the cell surface and activate phospholipase C. PMID- 10530809 TI - Group-I metabotropic glutamate receptors: hypotheses to explain their dual role in neurotoxicity and neuroprotection. AB - The role of group-I metabotropic glutamate receptors (mGlu1 and 5) in neurodegeneration is still controversial. While antagonists of these receptors are consistently neuroprotective, agonists have been found to either amplify or attenuate excitotoxic neuronal death. At least three variables affect responses to agonists: (i) the presence of the NR2C subunit in the NMDA receptor complex; (ii) the existence of an activity-dependent functional switch of group-I mGlu receptors, similar to that described for the regulation of glutamate release; and (iii) the presence of astrocytes expressing mGlu5 receptors. Thus, a number of factors, including the heteromeric composition of NMDA receptors, the exposure time to drugs or to ambient glutamate, and the function of astrocytes clearing extracellular glutamate and producing neurotoxic or neuroprotective factors, must be taken into account when examining the role of group-I mGlu receptors in neurodegeneration/neuroprotection. PMID- 10530811 TI - 2-Methyl-6-(phenylethynyl)-pyridine (MPEP), a potent, selective and systemically active mGlu5 receptor antagonist. AB - In the present paper we describe 2-methyl-6-(phenylethynyl)-pyridine (MPEP) as a potent, selective and systemically active antagonist for the metabotropic glutamate receptor subtype 5 (mGlu5). At the human mGlu5a receptor expressed in recombinant cells, MPEP completely inhibited quisqualate-stimulated phosphoinositide (PI) hydrolysis with an IC50 value of 36 nM while having no agonist or antagonist activities at cells expressing the human mGlu1b receptor at concentrations up to 30 microM. When tested at group II and III receptors, MPEP did not show agonist or antagonist activity at 100 microM on human mGlu2, -3, 4a, -7b, and -8a receptors nor at 10 microM on the human mGlu6 receptor. Electrophysiological recordings in Xenopus laevis oocytes demonstrated no significant effect at 100 microM on human NMDA (NMDA1A/2A), rat AMPA (Glu3 (flop)) and human kainate (Glu6-(IYQ)) receptor subtypes nor at 10 microM on the human NMDA1A/2B receptor. In rat neonatal brain slices, MPEP inhibited DHPG stimulated PI hydrolysis with a potency and selectivity similar to that observed on human mGlu receptors. Furthermore, in extracellular recordings in the CA1 area of the hippocampus in anesthetized rats, the microiontophoretic application of DHPG induced neuronal firing that was blocked when MPEP was administered by iontophoretic or intravenous routes. Excitations induced by microiontophoretic application of AMPA were not affected. PMID- 10530814 TI - [3H]-LY341495 as a novel antagonist radioligand for group II metabotropic glutamate (mGlu) receptors: characterization of binding to membranes of mGlu receptor subtype expressing cells. AB - Metabotropic glutamate (mGlu) receptors are a family of eight known subtypes termed mGlu1-8. Currently, few ligands are available to study the pharmacology of mGlu receptor subtypes. In functional assays, we previously described LY341495 as a highly potent and selective mGlu2 and mGlu3 receptor antagonist. In this study, radiolabeled [3H]-LY341495 was used to investigate the characteristics of receptor binding to membranes from cells expressing human mGlu receptor subtypes. Using membranes from cells expressing human mGlu2 and mGlu3 receptors, [3H] LY341495 (1 nM) specific binding was > 90% of total binding. At an approximate K(D) concentration for [3H]-LY341495 binding to human mGlu2 and mGlu3 receptors (1 nM), no appreciable specific binding of [3H-]LY341495 was found in membranes of cells expressing human mGlu1a, mGlu5a, mGlu4a, mGlu6, or mGlu7a receptors. However, modest (approximately 20% of mGlu2/3) specific [3H]-LY341495 (1 nM) binding was observed in human mGlu8 expressing cells. [3H]-LY341495 bound to membranes expressing human mGlu2 and mGlu3 receptors in a reversible and saturable manner with relatively high affinities (Bmax 20.5 +/- 5.4 and 32.0 +/- 7.0 pmol/mg protein; and K(D) = 1.67 +/- 0.20 and 0.75 +/- 0.43 nM, respectively). The pharmacology of [3H]-LY341495 binding in mGlu2 and mGlu3 expressing cells was consistent with that previously described for LY341495 in functional assays. [3H]-LY341495 binding provides a useful way to further investigate regulation of receptor expression and pharmacological properties of mGlu2 and mGlu3 receptor subtypes in recombinant systems. PMID- 10530812 TI - Evaluation of agonists and antagonists acting at Group I metabotropic glutamate receptors in the thalamus in vivo. AB - Recordings were made from single neurones in the ventrobasal thalamus of anaesthetised rats in order to evaluate the properties of several agonists and antagonists of Group I mGlu receptors. The selective mGlu1 receptor antagonist LY367385 was found to reduce excitatory responses to iontophoretically applied ACPD and DHPG whereas the mGlu5 agonist CHPG was resistant to antagonism. The antagonists LY367366 and LY393053 reduced responses to all three agonists, but without reducing responses to NMDA or AMPA. Although AIDA was also found to reduce mGlu agonist-evoked responses, this antagonist also produced significant reductions in responses to NMDA and AMPA. These data suggest that there are functional mGlu1 and mGlu5 receptors in the thalamus. Furthermore, LY367385 is a useful tool for investigating mGlu1 functions whereas LY367366 and LY393053 have a broader spectrum of action. The usefulness of AIDA as an antagonist in physiological experiments would appear to be limited by its effects against NMDA and AMPA. PMID- 10530813 TI - Group 1 mGlu receptors elevate [Ca2+]i in rat cultured cortical type 2 astrocytes: [Ca2+]i synergy with adenosine A1 receptors. AB - Brain macroglia are known to express a diverse array of neurotransmitter receptors whose signal transduction pathways may be subject to heteroreceptor 'cross-talk'. In the current study we have examined group 1 mGlu receptor-evoked [Ca2+]i signalling, and possible heteroreceptor cross-talk, in cultured type 2 astrocytes. The selective group 1 metabotropic glutamate (mGlu) receptor agonist (S)-3,5-dihydroxyphenylglycine (DHPG) elevated [Ca2+]i (EC50 = 1.7 +/- 0.6 microM); an effect reversed by the selective mGlu receptor antagonist (S)-alpha methyl-4-carboxyphenylglycine (IC50 = 52.7 +/- 8.7 microM). DHPG-evoked [Ca2+]i responses were abolished by (1) thapsigargin (100 nM), implicating the involvement of internal Ca2+ stores in group 1 mGlu [Ca2+]i responses and (2) the removal of extracellular Ca2+. When applied alone, the selective adenosine A1 receptor agonist, N6-cyclopentyladenosine (CPA, 100 nM) failed to influence [Ca2+]i. However, in the presence of 1 microM DHPG, CPA potently (EC50 = 12.3 +/- 1.9 nM) increased [Ca2+]i responses. In the presence of 100 nM CPA, the efficacy of DHPG was doubled without any significant change in the DHPG EC50 value. This effect was reversed by either the selective adenosine A1 receptor antagonist, 8 cyclopentyltheophylline (IC50 = 50.3 +/- 19.9 nM) or overnight incubation with Pertussis toxin (100 ng/ml). We conclude that (1) type 2 astrocytes contain group 1 mGlu receptors coupled to [Ca2+]i signalling and (2) co-activation of adenosine A1 receptors enhances group 1 mGlu-evoked [Ca2+]i responses in these cells via a Gi/o G protein-mediated mechanism. PMID- 10530815 TI - Inhibition of uptake and release of a novel mGluR agonist (L-F2CCG-I) by anion transport blockers in the rat spinal cord. AB - A new metabotropic glutamate receptor (mGluR) agonist, (2S,1'S,2'S)-2-(2-carboxy 3,3-difluorocyclopropyl)glycine (L-F2CCG-I), induces a priming effect on (RS) alpha-aminopimelate in the isolated spinal cord of newborn rats. Similar to (RS) alpha-aminopimelate, L-glutamate (30-100 microM) neither affected spinal reflexes nor the resting membrane potentials of motoneurones, but preferentially potentiated the depression of monosynaptic excitation caused by L-F2CCG-I (0.4 microM). Following L-F2CCG-I treatment (1-2 microM), L-glutamate decreased the monosynaptic spinal reflexes in a concentration dependent manner, indicating a priming' effect of L-F2CCG-I. Thus L-glutamate is completely compatible with (RS) alpha-aminopimelate in revealing the priming effect. An anion transport blocker, 4,4'-dinitrostilbene-2,2'-disulphonic acid (DNDS) (100 microM), markedly inhibited both the response to (RS)-alpha-aminopimelate and the induction of the L-F2CCG-I priming effect. The data suggest that L-F2CCG-I is Cl- -dependently incorporated into certain stores, and that (RS)-alpha-aminopimelate or L glutamate must stimulate the release of L-F2CCG-I from the storage site. There were pharmacological similarities between the quisqualate and L-F2CCG-I priming effect. The physiological significance of the quisqualate or L-F2CCG-I priming is not yet established. L-F2CCG-I would be expected to be a useful pharmacological probe for elucidating the mechanism of the priming. PMID- 10530816 TI - Extended glutamate activates metabotropic receptor types 1, 2 and 4: selective features at mGluR4 binding site. AB - To get an insight into the bioactive conformation of glutamic acid and its topological environment at the mGluR4 binding site, a pharmacophore model was constructed using molecular modeling. Agonists of known activities were used to run the Apex-3D program or to validate the resulting model. An extended glutamate conformer, two selective hydrophilic sites and bulk tolerance regions are disclosed. Selective features of mGluR1, mGluR2 and mGluR4 are discussed. PMID- 10530817 TI - Modulation of synaptic transmission and differential localisation of mGlus in cultured hippocampal autapses. AB - Metabotropic glutamate receptors (mGlus) are known to modulate synaptic transmission in various pathways of the central nervous system, but the exact mechanisms by which this modulation occurs remain unclear. Here we utilise electrophysiological and immunocytochemical techniques on cultured autaptic hippocampal neurones to investigate the mechanism of action and distribution of mGlus. Agonists at all three groups of mGlus depressed glutamatergic transmission, whereas only agonists at group I mGlus depressed GABAergic transmission. Agonists at all mGlus failed to modulate Ca2+ and K+ channels in glutamatergic autapses whereas an agonist at group III mGlus did depress the frequency of miniature excitatory postsynaptic currents (mEPSCs). Agonists failed to modulate Ca2+ or K+ channels and miniature inhibitory postsynaptic currents (mIPSCs) in GABAergic autapses. Distribution studies using selective antibodies revealed punctate staining for group III mGlus that co-localised with the synaptic marker, synaptophysin. Staining for the remaining mGlus was more diffuse throughout the soma and processes with little co-localisation with synaptophysin. The distribution of the group III receptors is consistent with the direct 'downstream' modulation of mEPSCs, although the exact mechanism of action for the remaining receptors remains unclear. PMID- 10530818 TI - Potentiation of NMDA and AMPA responses by the specific mGluR5 agonist CHPG in spinal cord motoneurons. AB - The specific metabotropic glutamate receptor (mGluR)5 agonist (RS)-2-chloro-5 hydroxyphenylglycine (CHPG) is able to potentiate NMDA and AMPA responses recorded from ventral roots of the isolated hemisected baby rat spinal cord. Previously we have demonstrated that activation of group I mGluRs (mGluR1 and mGluR5) with the broad spectrum mGluR agonist 1S,3R-1-amino-1,3 cyclopentanedicarboxylate (ACPD) produced potentiation of ionotropic glutamate responses. In contrast to ACPD-induced potentiation, however, no evidence for an involvement of protein kinase C (PKC) is found in the CHPG-induced potentiation of both NMDA and AMPA depolarization because the PKC blockers chelerythrine chloride or calphostin C did not antagonize this effect. Moreover, in the absence of Ca2+ in the perfusing medium or depleting intracellular Ca2+ stores with thapsigargin or dantrolene did not modify the CHPG-induced enhancement of NMDA depolarizations. Phorbol-12,13-diacetate (PDA), on the other hand, was able to attenuate this effect, which was reversed by chelerythrine chloride. These results suggest that both mGluR5 and mGluR1 may act to enhance ionotropic glutamate responses but the two types of mGluRs may have different intracellular mechanisms of action. PMID- 10530819 TI - DHPG-induced LTD in area CA1 of juvenile rat hippocampus; characterisation and sensitivity to novel mGlu receptor antagonists. AB - We have used extracellular microelectrode recording to characterise a form of long-term depression (LTD) of synaptic transmission that can be induced by metabotropic glutamate (mGlu) receptor activation in the CA1 region of the young (12-18 day old) rat hippocampus. Activation of group I mGlu receptors by the specific agonist 3,5-dihydroxyphenylglyine (DHPG) induced LTD of field excitatory postsynaptic potentials (fEPSPs). The mGlu5 selective agonist 2-chloro-5 hydroxyphenylglycine was also capable of inducing LTD. In contrast, the group II specific agonist DCG-IV had no effect on synaptic transmission, whilst the group III receptor agonist (S)-2-amino-4-phosphonobutyrate elicited a depression that reversed fully upon agonist washout. DHPG-induced LTD could still be generated after prior saturation of electrically-induced NMDA receptor-dependent LTD. DHPG induced LTD was reversed by tetanic stimulation comprising 100 shocks delivered at 100 Hz. A novel mGlu receptor antagonist, (RS)-2-amino-2-(3-cis and trans carboxycyclobutyl-3-(9-thioxanthyl)propionic acid) (LY393053) that potently inhibits mGlu1 and mGlu5 receptors, prevented the induction of DHPG-induced LTD. Like other mGlu receptor antagonists, LY393053 also reversed pre-established DHPG induced LTD. In contrast, a potent mGlu1 selective antagonist (S)-2-methyl-4 carboxyphenylglycine (LY367385) did not prevent the induction of DHPG-induced LTD. In conclusion, DHPG, probably via activation of mGlu5 receptors, is able to induce a robust form of LTD in the CA1 region of the young rat hippocampus that is mechanistically distinct from NMDA receptor-dependent homosynaptic LTD. PMID- 10530820 TI - An investigation into signal transduction mechanisms involved in DHPG-induced LTD in the CA1 region of the hippocampus. AB - Previously, we have found that activation of mGlu receptors using a group I specific mGlu receptor agonist, (RS)-3,5-DHPG, can induce long-term depression (LTD) in the CA1 region of the hippocampus and that, once established, this synaptic depression can be reversed by application of the mGlu receptor antagonist, (S)-MCPG [Palmer et al., 1997. Neuropharmacology 36, 1517-1532]. We have started to investigate the signal transduction mechanisms involved in these effects. Group I mGlu receptors couple to phospholipase C and therefore can activate protein kinase C and mobilise Ca2+ from intracellular stores. However, neither protein kinase C inhibitors (chelerythrine or Ro 31-8220) nor agents which deplete intracellular Ca2+ stores (thapsigargin or cyclopiazonic acid) were able to prevent DHPG-induced LTD. Furthermore, the ability of MCPG to reverse DHPG-induced LTD was not prevented by these compounds. These results suggest that it is unlikely that DHPG-induced LTD, or its reversal by MCPG, is produced via activation of either protein kinase C or by release of Ca2+ from intracellular stores. PMID- 10530822 TI - Protection with metabotropic glutamate 1 receptor antagonists in models of ischemic neuronal death: time-course and mechanisms. AB - In order to study the role of metabotropic glutamate 1 (mGlu1) receptors in ischemic neuronal death, we examined the effects of the recently characterized and relatively selective mGlu1 receptor antagonists 1-aminoindan-1,5-dicarboxylic acid (AIDA) and (S)-(+)-2-(3'-carboxybicyclo[1.1.1]pentyl)-glycine (CBPG) in murine cortical cell cultures and rat organotypic hippocampal slices exposed to oxygen glucose deprivation (OGD) and in vivo, following transient global ischemia in gerbils. AIDA and CBPG significantly reduced neuronal death when added to the incubation medium during the OGD insult and the subsequent recovery period. Neuroprotection was observed even when these compounds were added up to 60 min (in cortical neurons) or 30 min (in hippocampal slices) after OGD. In vivo, i.c.v. administration of AIDA and CBPG reduced hippocampal CA1 pyramidal cell injury following transient global ischemia. Neuroprotection was also observed when AIDA was added to the hippocampal perfusion fluid in microdialysis experiments, and this effect was associated with an increase in the basal output of GABA. These findings demonstrate that AIDA and CBPG are neuroprotective when administered during the maturation of ischemic damage and that different mechanisms are likely to be involved in mediating their effects following blockade of mGlu1 receptors in cortical and hippocampal neurons. PMID- 10530821 TI - Induction of LTD by activation of group I mGluR in the dentate gyrus in vitro. AB - The ability of activation of group I metabotropic glutamate receptors (mGluR) to induce long-term depression (LTD) was investigated in the medial perforant path of the dentate gyrus in vitro. Application of the group I agonists (RS)-3,5 dihydroxyphenylglycine (DHPG) and (RS)-2-chloro-5-hydroxyphenylglycine (CHPG), and also the partial agonist (S)-(+)-2-(3'-Carboxybicyclo[1.1.1]pentyl)-glycine (UPF 596), induced LTD of the field EPSP. The induction of LTD is likely to be mediated via mGluR5 since CHPG and UPF 596 are selective agonists/partial agonists at that receptor. Further evidence for the involvement of group I mGluR in LTD induction was the finding, that the DHPG and low frequency stimulation induced LTD were inhibited by the group I mGluR antagonist [CRS]-1-aminoindan-1,5 dicarboxylic acid (AIDA). Investigation of the intracellular mechanisms underlying the induction of the group I mGluR-mediated LTD showed an inhibition of the LTD by the protein kinase C (PKC) inhibitor bisindolylmaleimide I and the protein tyrosine kinase inhibitor lavendustin A, but not the PKA inhibitor H89. These studies demonstrate that DHPG-induced LTD can be induced by the activation of mGluR5 followed by intracellular stimulation of PKC and tyrosine kinase. PMID- 10530823 TI - Selective activation of group II mGluRs with LY354740 does not prevent neuronal excitotoxicity. AB - Recent reports have suggested a role for group II metabotropic glutamate receptors (mGluRs) in the attenuation of excitotoxicity. Here we examined the effects of the recently available group II agonist (+)-2 Aminobicyclo[3.1.0]hexane-2-6-dicarboxylic acid (LY354740) on N-methyl-D aspartate (NMDA)-induced excitotoxic neuronal death, as well as on hypoxic ischemic neuronal death both in vitro and in vivo. At concentrations shown to be selective for group II mGluRs expressed in cell lines (0.1-100 nM), LY354740 did not attenuate NMDA-mediated neuronal death in vitro or in vivo. Furthermore, LY354740 did not attenuate oxygen-glucose deprivation-induced neuronal death in vitro or ischemic infarction after transient middle cerebral artery occlusion in rats. In addition, the neuroprotective effect of another group II agonist, (S)-4 carboxy-3-phenylglycine (4C3HPG), which has shown injury attenuating effects both in vitro and in vivo, was not blocked by the group II antagonists (2 S)-alpha ethylglutamic acid (EGLU), (RS)-alpha-methyl-4-sulphonophenylglycine (MSPG), or the group III antagonist (S)-alpha-methyl-3-carboxyphenylalanine (MCPA), suggesting that this neuroprotection may be mediated by other effects such as upon group I mGluRs. PMID- 10530824 TI - mGluR7-like receptor and GABA(B) receptor activation enhance neurotoxic effects of N-methyl-D-aspartate in cultured mouse striatal GABAergic neurones. AB - Presynaptic metabotropic glutamate receptors (mGluRs) of group III constitute possible targets for putative neuroprotective drugs acting against glutamate excitotoxic insults. Indeed, in glutamatergic cerebellar granule neurones in culture, high concentrations of L-2-amino-4-phosphonobutyrate (L-AP4, above 0.3 mM, thus activating mGluR7) inhibit NMDA-induced cell death. In contrast, in striatal cultures which are enriched in GABAergic neurones, we show that high concentrations of L-AP4 increased neuronal death in control as well as in NMDA stimulated cultures. Moreover, similar results were obtained with the GABA(B)R agonist. baclofen. Both the neuroprotective effects in cerebellar granule cells and the neurotoxic effects in striatal neurones were mediated via Gi-Go-coupled mGluRs, suggesting that these effects were probably mediated by mGluR7a or b and GABA(B)R expressed in these neurones. In striatal neurones, we found that L-AP4 and baclofen inhibited both basal and NMDA-stimulated GABA release. These inhibitions of GABA release may be responsible for the increase in basal and NMDA stimulated neuronal death. Indeed, blockade of GABA(A) receptors with bicuculline increased neuronal death of control and NMDA-treated striatal cultures. Taken together, these results suggest that L-AP4 and baclofen, via mGluR7 and GABA(B)R, reduced the neuroprotective effect of GABA present in striatal cultures acting via GABA(A) receptors. Although caution must be taken when extrapolating from in vitro to in vivo situations, the present experiments and the recent observations that mGluR7 and GABA(B)R are expressed in heterologous synapses, should be taken into consideration when evaluating the neuroprotective action of future mGluR7 specific agonists or GABA(B)R specific antagonists. PMID- 10530825 TI - Patient care information systems and health care work: a sociotechnical approach. AB - Those who face the difficulties of developing useful patient care information systems (PCISs) often stress the importance of 'organizational issues'. Building upon recent sociological insights in the construction and use of information technologies for (health care) work, this paper underscores the importance of these insights for the development and evaluation of these systems. A sociotechnical approach to PCISs in health care is outlined, and two implications of this empirically grounded approach for the practices of developing and evaluating IT applications in health care practices are discussed. First, getting such technologies to work in concrete health care practices appears to be a politically textured process of organizational change, in which users have to be put at center-stage. This requires an iterative approach, in which the distinctions between 'analysis', 'design', 'implementation' and 'evaluation' blur. Second, a sociotechnical approach sheds new light on the potential roles of IT applications in health care practices. It is critical of approaches that denounce the 'messy' and 'ad hoc' nature of health care work, and that attempt to structure this work through the formal, standardized and 'rational' nature of IT systems. Optimal utilization of IT applications, it is argued, is dependent on the meticulous interrelation of the system's functioning with the skilled and pragmatically oriented work of health care professionals. PMID- 10530826 TI - A search engine for virtual patient records. AB - Virtual patient records provide a means for integrated access to patient information that may be scattered around different healthcare settings. Within the boundaries of a health district providing all levels of care, this concept can be implemented in an Intranet environment to support longitudinal patient care activities across the participating healthcare providers. Since medical information is stored on multiple Intranet sites in various forms (e.g. codified data, transcribed documents, and images), a suite of appropriate tools is needed to enable access to such information in combined form. In most cases, however, access to medical information should be restricted to authorized users. To serve this purpose, a prototype search engine incorporating an authorization and access control functionality has been developed and presented in this paper. The system is based on the signature file access method and an experimental implementation written in JAVA is also described. PMID- 10530827 TI - Cascaded MRI-SPAMM for LV motion analysis during a whole cardiac cycle. AB - We present a new paradigm which incorporates multiple sets of tagged MRI data (MRI-SPAMM) acquired in a cascaded fashion in order to estimate the full 3-D motion of the left ventricle (LV) during its entire cardiac cycle. Our technique is based on an extension of our volumetric physics-based deformable models, whose parameters are functions. Using these parameters, we can characterize the local shape variation of an object with a small number of intuitive parameters. By integrating a cascaded sequence of SPAMM data sets into our modeling technique, we have extended the capability of the MRI-SPAMM technique and have provided an accurate representation of the LV motion during the full cardiac cycle (from end diastole to end-diastole) to better understand cardiac mechanics. PMID- 10530828 TI - A computer-based information system for epilepsy and electroencephalography. AB - This paper describes a standardised computer-based information system for electroencephalography (EEG) focusing on epilepsy. The system was developed using a prototyping approach. It is based on international recommendations for EEG examination, interpretation and terminology, international guidelines for epidemiological studies on epilepsy and classification of epileptic seizures and syndromes and international classification of diseases. It is divided into: (1) clinical information and epilepsy relevant data; and (2) EEG data, which is hierarchically structured including description and interpretation of EEG. Data is coded but is supplemented with unrestricted text. The resulting patient database can be integrated with other clinical databases and with the patient record system and may facilitate clinical and epidemiological research and development of standards and guidelines for EEG description and interpretation. The system is currently used for teleconsultation between Gentofte and Lisbon. PMID- 10530829 TI - Fuzzy logic algorithm for quantitative tissue characterization of diffuse liver diseases from ultrasound images. AB - Computerized ultrasound tissue characterization has become an objective means for diagnosis of liver diseases. It is difficult to differentiate diffuse liver diseases, namely cirrhotic and fatty liver by visual inspection from the ultrasound images. The visual criteria for differentiating diffused diseases are rather confusing and highly dependent upon the sonographer's experience. This often causes a bias effects in the diagnostic procedure and limits its objectivity and reproducibility. Computerized tissue characterization to assist quantitatively the sonographer for the accurate differentiation and to minimize the degree of risk is thus justified. Fuzzy logic has emerged as one of the most active area in classification. In this paper, we present an approach that employs Fuzzy reasoning techniques to automatically differentiate diffuse liver diseases using numerical quantitative features measured from the ultrasound images. Fuzzy rules were generated from over 140 cases consisting of normal, fatty, and cirrhotic livers. The input to the fuzzy system is an eight dimensional vector of feature values: the mean gray level (MGL), the percentile 10%, the contrast (CON), the angular second moment (ASM), the entropy (ENT), the correlation (COR), the attenuation (ATTEN) and the speckle separation. The output of the fuzzy system is one of the three categories: cirrhosis, fatty or normal. The steps done for differentiating the pathologies are data acquisition and feature extraction, dividing the input spaces of the measured quantitative data into fuzzy sets. Based on the expert knowledge, the fuzzy rules are generated and applied using the fuzzy inference procedures to determine the pathology. Different membership functions are developed for the input spaces. This approach has resulted in very good sensitivities and specificity for classifying diffused liver pathologies. This classification technique can be used in the diagnostic process, together with the history information, laboratory, clinical and pathological examinations. PMID- 10530830 TI - A computer-based outpatient clinical referral system. AB - The process of generating a clinical referral for a patient, and the resulting transfer of information from the primary care physician to the specialist and back again, are key components in the struggle to deliver less costly and more effective clinical care. We have created a computer-based, outpatient clinical referral application that facilitates: (1) identifying an appropriate specialist; (2) collecting the clinical, demographic, and financial data required to generate a referral; and (3) transferring the information between the specialist and the primary care physician (PCP). This article describes the development of the application itself and several of the knowledge bases that were created to facilitate this process. Preliminary results indicate that the new computer-based referral process is faster to use than conventional methods. PMID- 10530831 TI - Infection in the operating room. PMID- 10530832 TI - The diagnosis and treatment of injury to the tarsometatarsal joint complex. PMID- 10530833 TI - Detailed analysis of proprioception in normal and ACL-deficient knees. AB - We assessed proprioception using threshold levels for the perception of knee movement at slow angular velocities (0.1 degrees/s to 0.85 degrees/s) in 20 patients with unilateral tears of the anterior cruciate ligament (ACL) and 15 age related control subjects. Failure to detect movement was also analysed. The threshold levels of detection did not differ between the damaged and undamaged knees in the patients or between the patients and the control group. Failure to appreciate movement, however, was significantly greater in knees with ACL loss compared with the undamaged knees of patients and the control group. Our findings show a proprioceptive deficit in the absence of the ACL. Measurements of threshold levels of detection of passive movement alone are not suitable for the evaluation of proprioceptive loss in ACL deficiency; assessment of failure to appreciate movement is essential. PMID- 10530835 TI - Arthroscopic reconstruction of the anterior cruciate ligament with patellar tendon autograft and interference screw fixation. The results at seven years. AB - Deficiency of the anterior cruciate ligament (ACL) is a common disorder which can lead to changes in lifestyle. We followed 59 patients who had had arthroscopic reconstruction of the ACL using a central-third patellar-tendon autograft for seven years to assess the long-term effectiveness of recent advances in reconstruction of the ACL. The standard criteria for evaluation of the International Knee Documentation Committee, the Lysholm knee score and measurements using the KT 1000 arthrometer all showed satisfactory results. Deterioration in the clinical performance after seven years was associated with osteoarthritic changes and correlated with chronic ligament injuries and meniscectomy. There were three traumatic and three spontaneous ruptures. We believe that the procedure can be successful, but remain concerned about failure of the graft and osteoarthritis. The results raise questions about the best time to operate and suggest that early surgery may reduce the risk of osteoarthritis. PMID- 10530836 TI - Osteotomy of the tibia for correction of complex deformity. AB - Twenty complex tibial deformities due to anterior poliomyelitis in 18 patients were corrected by a modified O'Donoghue osteotomy. This technique allowed correction of the deformity in three planes. This was achieved by widening the rectangular window distally to correct both rotation and valgus and by trimming the anterior edges of the step cuts to correct flexion deformity. An above-knee cast was applied for eight to 13 weeks and the patients followed up for a mean of 3.2 years. One of the 18 patients developed delayed union because of fracture of the medial limb of the step cut. The results showed excellent correction of the three-plane deformity and there was no recurrence. This method of osteotomy is a safe and simple procedure which does not require internal fixation and allows correction of torsional and angular deformity. PMID- 10530834 TI - Correction of genu recurvatum by the Ilizarov method. AB - The Ilizarov apparatus was used to carry out opening-wedge callotasis of the proximal tibia in ten patients who had suffered premature asymmetrical closure of the proximal tibial physis and subsequent genu recurvatum. In four knees, the genu recurvatum was entirely due to osseous deformity, whereas in six it was associated with capsuloligamentous abnormality. Preoperatively, the angle of recurvatum averaged 19.6 degrees (15 to 26), the angle of tilt of the tibial plateau, 76.6 degrees (62 to 90), and the ipsilateral limb shortening, 2.7 cm (0.5 to 8.7). The average time for correction was 49 days (23 to 85). The average duration of external fixation was 150 days (88 to 210). Three patients suffered complications including patella infera, pin-track infection and transient peroneal nerve palsy. At a mean follow-up of 4.4 years, all patients, except one, had achieved an excellent or good radiological and functional outcome. PMID- 10530838 TI - The length of the patellar tendon after unicompartmental and total knee replacement. AB - Patella infera may occur after reconstruction of the anterior cruciate ligament (ACL), high tibial osteotomy and total knee replacement (TKR). Restriction of movement of the knee and pain may result. Our aim was to compare the incidence and to assess the effects of patella infera after TKR and unicompartmental knee replacement (UKR). We reviewed radiographs of the knees of 84 patients who had had either TKR or UKR as part of a randomised, controlled trial The length of the patellar tendon was measured on serial radiographs taken before, at eight months and at five years after operation. There was no significant change in the length of the patellar tendon after UKR, but a significant reduction was observed after TKR. Five years after the operation, the shortening of the tendon had increased to a mean of 3.5 mm. Of the knees with TKR reviewed at five years, 34% developed patella infera, defined as 10% or more of shortening, compared with 5% of those with UKR. Shortening was greatest in those knees which had required a lateral release; in this subgroup the mean shortening was 7.2 mm. Shortening correlated with restriction of movement and pain in the knee. Our study has shown that patella infera develops in most patients after TKR with lateral release, and in approximately 25% of patients after TKR without this additional procedure. Patella infera rarely occurs after UKR. It is associated with restriction of movement and pain in the knee. It may be an effect of the more extensive exposure required to perform TKR and may, in part, explain the better clinical results of UKR. PMID- 10530837 TI - Does arthritis progress in the retained compartments after 'Oxford' medial unicompartmental arthroplasty? A clinical and radiological study with a minimum ten-year follow-up. AB - We determined the outcome of 56 'Oxford' unicompartmental replacements performed for anteromedial osteoarthritis of the knee between 1982 and 1987. Of these, 24 were in patients who had died without revision, one was lost to follow-up and two had been revised. Of the remaining 29 knees, 26 were examined clinically and radiologically, two were only examined clinically and one patient was contacted by telephone. The mean age of the patients was 80.3 years. At a mean follow-up of 11.4 years (10 to 14) the measurements of the knee score, range of movement and degree of deformity were not significantly different from those made one to two years after operation, except that the range of flexion had improved. Comparison of fluoroscopically-controlled radiographs at a similar interval of time showed no change in the appearance of the lateral compartments. The retained articular cartilage continued to function for ten or more years which suggests that anteromedial osteoarthritis may be considered as a focal disorder of the knee. This justifies continued efforts to develop methods of treatment which preserve intact joint structures. PMID- 10530839 TI - Osteosarcoma of the pelvis. AB - Over a 25-year period we have treated 36 patients with osteosarcoma of the pelvis. Of the tumours, 24 (67%) were primary osteosarcomas and 12 (33%) arose either after irradiation or in association with Paget's disease. Six patients had a hindquarter amputation and 12 were treated by a limb-salvage procedure with intrapelvic excision. The five-year survival rate of all the patients with pelvic osteosarcoma was 18%, while for 17 treated by chemotherapy and surgery it was 41%. The prognosis for patients presenting with metastases or with secondary osteosarcoma was appalling and none survived after 29 months. No patient over the age of 50 years when seen initially survived for a year. Youth and a good response to chemotherapy along with complete surgical excision offer the best chance of cure. PMID- 10530840 TI - Prosthetic reconstruction for tumours of the distal tibia and fibula. AB - We have carried out prosthetic reconstruction in six patients with malignant or aggressively benign bone tumours of the distal tibia or fibula. The diagnoses were osteosarcoma in four patients, parosteal osteosarcoma in one and recurrent giant-cell tumour in one. Five tumours were in the distal tibia and one in the distal fibula. The mean duration of follow-up was 5.3 years (2.0 to 7.1). Reconstruction was achieved using custom-made, hinged prostheses which replaced the distal tibia and the ankle. The mean range of ankle movement after operation was 31 degrees and the joints were stable. The average functional score according to the system of the International Society of Limb Salvage was 24.2 and five of the patients had a good outcome. Complications occurred in two with wound infection and talar collapse. All patients were free from neoplastic disease at the latest follow-up. Prosthetic reconstruction may be used for the treatment of malignant tumours of the distal tibia and fibula in selected patients. PMID- 10530841 TI - Reconstruction and limb salvage after resection for malignant bone tumour of the proximal humerus. A sling procedure using a free vascularised fibular graft. AB - We assessed the intermediate functional results of eight patients after wide resection of the proximal humerus for malignant bone tumour. We used a free vascularised fibular graft as a functional spacer and a sling procedure to preserve passive scapulohumeral movement. Scapulohumeral arthrodesis was not carried out. Five patients had osteosarcoma, two achondrosarcoma and one a malignant fibrous histiocytoma of the bone. The mean duration of follow-up was 70 months (median, 76) for the seven patients who were still alive at the time of the latest follow-up. One patient died from the disease 12 months after surgery. There were no local recurrences. The functional results were described and graded quantitatively according to the rating system of the Musculoskeletal Tumour Society. Our results were satisfactory with regard to pain, emotional acceptance and manual dexterity. Function and lifting ability were unsatisfactory in two patients. One patient had delayed union between host and graft, but this united after six months without further surgery. Radiographs of the shoulder showed absorption or collapse of the head of the fibula in four of the eight patients and a fracture in another. No functional problems related to absorption or fracture of the head of the fibula were noted. There was no infection or subluxation of the head. We conclude that this is a reasonably effective technique of limb salvage after resection of the proximal humerus. PMID- 10530842 TI - Osteoid osteoma. Direct visual identification and intralesional excision of the nidus with minimal removal of bone. AB - We describe 100 consecutive patients with osteoid osteoma. Of the 97 who had operations, 89 were treated by intralesional excision and eight by wide resection. The three remaining patients were not operated on because the osteoid osteoma was almost painless, or was found in the pedicle of the 12th thoracic vertebra at the site of entrance of the artery of Adamkjewicz. The diagnosis was confirmed histologically in all specimens. No local recurrences were observed at a minimum follow-up of one year. All except one patient were mobilised two to four days after surgery. A precise preoperative diagnosis of the lesion is mandatory, based on clinical findings, standard radiographs, thin-section CT and a bone scan. We compared our operative technique with 247 cases in which the percutaneous technique of removal or coagulation of the nidus had been performed. The latter procedure has a less constant rate of primary cure (83% v 100%). Its principal indication appears to be for osteoid osteomas in the proximal femur and the pelvis. PMID- 10530843 TI - Pelvic obliquity after fusion of the spine in Duchenne muscular dystrophy. AB - Spinal fusion, ending caudally at L5 rather than at the sacrum, is recommended for selected patients with scoliosis due to Duchenne muscular dystrophy. We present a retrospective review of 48 patients operated on for this condition. Patients having spinal curvature with a Cobb angle of less than 40 degrees and with less than 10 degrees between a line tangential to the superior margins of both iliac crests and a line perpendicular to the spinous processes of L4 and L5, were fused to L5 (38 patients); patients not meeting these criteria were fused to the sacrum (10 patients). Spinal and sitting obliquity increased in patients fused to L5, rather than to the sacrum, but the severity of the worsening obliquity was significantly greater in patients in whom the apex of the curve was below L1. Two of the ten latter patients required revision procedures for worsening obliquity when their pulmonary function deteriorated to less than 25% of predicted values. We recommend fusion to the sacrum for scoliosis in Duchenne muscular dystrophy, especially for patients with an apex to their curve below L1. PMID- 10530844 TI - The placement of lumbar pedicle screws using computerised stereotactic guidance. AB - Computer-assisted frameless stereotactic image guidance allows precise preoperative planning and intraoperative localisation of the image. It has been developed and tested in the laboratory. We evaluated the efficacy, clinical results and complications of placement of a pedicle screw in the lumbar spine using this technique. A total of 62 patients (28 men, 34 women) had lumbar decompression and spinal fusion with segmental pedicle screws. Postoperative CT scans were taken of 35 patients to investigate the placement of 330 screws. None showed penetration of the medial or inferior wall of a pedicle. Registration was carried out 66 times. The number of fiducial points used on each registration averaged 5.8 (4 to 7) The mean registration error was 0.75 mm (0.32 to 1.72). This technique provides a safe and reliable guide for placement of transpedicular screws in the lumbar spine. PMID- 10530845 TI - The appearance on MRI of vertebrae in acute compression of the spinal cord due to metastases. AB - We studied MR images of the spine in a consecutive series of 100 patients with acute compression of the spinal cord due to metastases. All patients had documented neurological deficit and histologically proven carcinoma. MRI was used to localise bony metastatic involvement and soft-tissue impingement of the cord. A systematic method of documenting metastatic involvement is described. A total of 43 patients had compression at multiple levels; 160 vertebral levels were studied. In 120 vertebrae (75%), anterior, lateral and posterior bony elements were involved. Soft-tissue impingement of the spinal cord often involved more than one quadrant of its circumference. In 69 vertebrae (43%) there was concomitant anterior and posterior compression. Isolated involvement of a vertebral body was observed in only six vertebrae (3.8%). We have shown that in most cases of acute compression of the spinal cord due to metastases there is coexisting involvement of both anterior and posterior structures. PMID- 10530846 TI - Poor eight-year survival of cemented zirconia-polyethylene total hip replacements. AB - Between January 1988 and January 1991 we performed 100 consecutive cemented total hip replacements using a zirconia head, a titanium alloy stem and a polyethylene cup. We reviewed 78 of these hips in 61 patients in detail at a mean of 5.8 years (1 to 9). Aseptic loosening was seen in 11 hips (14%). Eight needed revision. In total, 37 cups (47.5%) showed radiolucent lines, all at the cement-bone interface, with 18 (23%) involving all the interface. Of the 78 femoral implants, 17 (21.7%) showed radiolucent lines, and two, which had a complete line of more than 1 mm thick, definite endocortical osteolyses. There was also an abnormally high incidence of osteolysis of more than 2 mm at the calcar. Survivorship analysis showed that only 63% were in situ at eight years. These worrying results led us to abandon the use of zirconia heads, since at the same hospital, using the same femoral stem, cement and polyethylene cup, but with alumina femoral heads, the survival rate was 93% at nine years. We discuss the possible reasons for the poor performance of zirconia ceramic. PMID- 10530847 TI - Nerve palsy after leg lengthening in total replacement arthroplasty for developmental dysplasia of the hip. AB - We reviewed 508 consecutive total hip replacements in 370 patients with old developmental dysplasia of the hip, to relate the amount of leg lengthening to the incidence of nerve palsies after operation. There were eight nerve palsies (two femoral, six sciatic), two complete and six incomplete. We found no statistical correlation between the amount of lengthening and the incidence of nerve damage (p = 0.47), but in seven of the eight hips, the surgeon had rated the intervention as difficult because of previous surgery, severe deformity, a defect of the acetabular roof, or considerable flexion deformity. The correlation between difficulty and nerve palsy was significant (p = 0.041). We conclude that nerve injury is most commonly caused by direct or indirect mechanical trauma and not by limb lengthening on its own. PMID- 10530848 TI - The use of ultrasound in determining the initiation of treatment in instability of the hip in neonates. AB - We have evaluated the effect of the use of ultrasound in determining the initiation of treatment in neonatal instability of the hip. A total of 99 newborn infants (1.5% of all live births) with neonatal hip instability did not have treatment from birth, but were re-examined at eight to 15 days. In the 31 who had persisting clinical instability and ultrasound abnormality, treatment was then started with a Frejka pillow. The hips in the remaining 68 infants showed spontaneous clinical stabilisation and improvement of the ultrasound findings. Treatment was therefore withheld. There was a marked trend towards normal development in mildly unstable hips, whereas no hips with severe instability did so spontaneously. Further follow-up showed normal development in all the hips which had been treated, and in all except five of the 68 untreated infants. These five infants showed persistent hip dysplasia on both ultrasound and radiological examination at four to five months of age. Treatment with an abduction splint was then started and their hips developed normally. Ultrasound is very useful in deciding on treatment if the examiners have adequate experience with the method. Its use substantially reduces the rate of treatment. Spontaneous resolution occurred in more than half of the unstable hips. Since five of the untreated infants developed hip dysplasia a strict follow-up is essential to identify and treat these cases. PMID- 10530849 TI - Financial justification for routine ultrasound screening of the neonatal hip. AB - We have analysed the patterns of management of developmental dysplasia of the hip (DDH) in Coventry over a period of 20 years during which three different screening policies were used. From 1976 to the end of 1985 we relied on clinical examination alone. The mean surgical cost for the treatment of DDH during this period was Pound Sterling 5110 per 1000 live births. This was reduced to Pound Sterling 3811 after the introduction of ultrasound for infants with known risk factors. Since June 1989 we have routinely scanned all infants at birth with a mean surgical cost of Pound Sterling 468 per 1000 live births. This reduction in cost is a result of the earlier detection of DDH with fewer children requiring surgery. In those who do, fewer and less invasive procedures are needed. The overall rate of treatment has not increased and regular review of patients managed in a Pavlik harness has allowed us to avoid the complication of avascular necrosis. When we add the cost of running the screening programme to the expense of treating the condition, the overall cost for the management of DDH is comparable for the different screening policies. PMID- 10530850 TI - The treatment of congenital club foot by operation to correct deformity and achieve dynamic muscle balance. AB - We operated on 111 patients with 159 congenital club feet with the aim of correcting the deformity and achieving dynamic muscle balance. Clinical and biomechanical assessment was undertaken at least six years after operation when the patient was more than 13 years of age. The mean follow-up was for 11 years 10 months (6 to 36 years). Good and excellent results were obtained in 91.8%. Patients with normal function of the calf had a better outcome than those with weak calf muscles. The radiological changes were assessed in relation to the clinical outcome. The distribution of pressure under the foot was measured for biomechanical assessment. Our results support the view that muscle imbalance is an aetiological factor in club foot. Early surgery seems to be preferable. It is suggested that operation should be undertaken as soon as possible after the age of six months, although it may be carried out up to the age of five years. The establishment of dynamic muscle balance appears to be an effective method of maintaining correction. Satisfactory long-term results can be achieved with adequate appearance and function. PMID- 10530851 TI - Shock-wave therapy is effective for chronic calcifying tendinitis of the shoulder. AB - We report a prospective study of the effects of extracorporeal shock-wave therapy in 195 patients with chronic calcifying tendinitis. In part A 80 patients with chronic symptoms were randomly assigned to a control and three subgroups which had different treatment by low-energy and high-energy shock waves. In part B 115 patients had either one or two high-energy sessions. We recorded subjective, functional and radiological findings at six months after treatment. The results showed energy-dependent success, with relief of pain ranging from 5% in our control group up to 58% after two high-energy sessions. The Constant scores and the radiological disintegration of calcification were also dose-dependent. Shockwave therapy should be considered for chronic pain due to calcific tendinitis which is resistant to conservative treatment. PMID- 10530852 TI - Work practices and histopathological changes in the tenosynovium and flexor retinaculum in carpal tunnel syndrome in women. AB - We studied 58 women of employable age with the carpal tunnel syndrome in order to determine whether the histological appearances of the carpal tunnel, tenosynovium and flexor retinaculum are influenced by work practices. Age, body mass index and the duration of symptoms did not correlate with the extent of oedema or fibrosis within the tenosynovium. The incidence of abnormality on histological examination of the tenosynovium was the same in employed and unemployed patients (p = 1.0), and was not influenced by the level of repetition (p = 0.89) or force (p = 0.29) of work. Myxoid degeneration within the flexor retinaculum was, however, more common in women undertaking 'high-force' work. Apart from this finding, the results suggest that work practices do not affect tenosynovial thickening, fibrosis or oedema in patients with carpal tunnel syndrome. PMID- 10530853 TI - The relationship between the site of nonunion of the scaphoid and scaphoid nonunion advanced collapse (SNAC). AB - We studied retrospectively the radiographs of 33 patients with late symptoms after scaphoid nonunion in an attempt to relate the incidence of scaphoid nonunion advanced collapse (SNAC) to the level of the original fracture. We found differing patterns for nonunion at the proximal, middle and distal thirds. The mean intervals between fracture and complaint were 20.9, 6.7 and 12.6 years and obvious degenerative changes occurred in 85.7%, 40.0% and 33.3%, for the six proximal-, eight middle- and two distal-third nonunions, respectively. Nonunion at the proximal and middle thirds showed the first degenerative changes at the radioscaphoid joint, and this was followed by narrowing of the scaphocapitate and then the lunocapitate joints. In our two nonunions of the distal third degenerative changes were seen only at the lunocapitate joint. Most patients with SNAC and nonunion of the middle or distal third showed dorsal intercalated instability; few patients with nonunion of the proximal third developed this deformity. We discuss the initial management of nonunion of the scaphoid at different levels in the light of our findings, and make recommendations. PMID- 10530854 TI - Percutaneous repair of the ruptured tendo Achillis. AB - Percutaneous repair of the ruptured tendo Achillis has a low rate of failure and negligible complications with the wound, but the sural nerve may be damaged. We describe a new technique which minimises the risk of injury to this nerve. The repair is carried out using three midline stab incisions over the posterior aspect of the tendon. A No. 1 nylon suture on a 90 mm cutting needle approximates the tendon with two box stitches. The procedure can be carried out under local anaesthesia. We reviewed 27 patients who had a percutaneous repair at a median interval of 35 months after the injury. They returned to work at four weeks and to sport at 16. One developed a minor wound infection and another complex regional pain syndrome type II. There were no injuries to the sural nerve or late reruptures. This technique is simple to undertake and has a low rate of complications. PMID- 10530855 TI - Total dislocations of the navicular: are they ever isolated injuries? AB - Isolated dislocations of the navicular are rare injuries; we present our experience of six cases in which the navicular was dislocated without fracture. All patients had complex injuries, with considerable disruption of the midfoot. Five patients had open reduction and stabilisation with Kirschner wires. One developed subluxation and deformity of the midfoot because of inadequate stabilisation of the lateral column, and there was one patient with ischaemic necrosis. We believe that the navicular cannot dislocate in isolation because of the rigid bony supports around it; there has to be significant disruption of both longitudinal columns of the foot. Most commonly, an abduction/pronation injury causes a midtarsal dislocation, and on spontaneous reduction the navicular may dislocate medially. This mechanism is similar to a perilunate dislocation. Stabilisation of both medial and lateral columns of the foot may sometimes be essential for isolated dislocations. In spite of our low incidence of ischaemic necrosis, there is always a likelihood of this complication. PMID- 10530857 TI - Vascular complications of osteotomies in limb reconstruction. AB - Osteotomies are commonly carried out in orthopaedic surgery, particularly in limb reconstruction. Complications are uncommon provided that sufficient care is taken and a sound technique used. We describe three cases of formation of false aneurysm after osteotomy, with acute, delayed and asymptomatic onset. The diagnosis was supported by ultrasound investigation, and confirmed by angiography. Embolisation with coils was a successful method of treatment. We recommend a safe method of osteotomy with good bone exposure and adequate soft tissue protection. PMID- 10530858 TI - Pyoderma gangrenosum. A diagnosis not to be missed. AB - We describe a case of pyoderma gangrenosum which presented with severe wound breakdown after elective hip replacement. The patient was treated successfully with minimal wound debridement and steroids. This diagnosis should always be considered when confronted with an enlarging painful skin lesion which does not grow organisms when cultured and fails to respond to antibiotic therapy, especially if there are similar lesions in other sites. In patients who have a past history of pyoderma gangrenosum, prophylactic steroids may be indicated at the time of surgery or may be required early in the postoperative period. PMID- 10530856 TI - Intraoperative bacterial contamination in operations for joint replacement. AB - All surgical operations have the potential for contamination, and the equipment used can harbour bacteria. We collected samples from 100 elective primary hip and knee arthroplasties. These showed rates of contamination of 11.4% for the sucker tips, 14.5% for light handles, 9.4% for skin blades and 3.2% for the inside blades used during surgery; 28.7% of gloves used for preparation were also contaminated. Of the samples taken from the collection bags used during hip arthroplasty, 20% grew bacteria, which represents a significant microbial reservoir. Also, 17% of theatre gowns were contaminated at the end of the operation. Contamination was found in 10% of the needles used during closure of the fascia. Overall, 76% of the organisms grown were coagulase-negative staphylococcus. A total of 63% of operations showed contamination in the field of operation. Some changes in practice are suggested. Follow-up for a minimum of two years revealed one deep infection but the organism was not identified as a contaminant. These data provide a baseline for studying the bacteriology of the surgery of revision arthroplasty. PMID- 10530859 TI - Muscle recovery after immobilisation by external fixation. AB - We immobilised the right hindlimbs of six-month-old female Wistar rats for four weeks using a biplanar external fixation bridging the knee. The untreated left limbs served as a control group. An additional group of rats was allowed to recover for four weeks after removal of the frame. Immobilisation caused reduction in the wet weights of approximately 50% in the gastrocnemius, quadriceps, soleus and plantaris muscles; this was not restored completely after remobilisation. There was an increase in the activity of acid phosphatase of approximately 85% in the gastrocnemius and quadriceps muscles whereas that of creatine phosphokinase was reduced by about 40%. These values returned to nearly normal after remobilisation. Histological and ultrastructural examination showed a marked myopathy of the gastrocnemius muscle after immobilisation although the morphology was largely restored after remobilisation. We conclude that after four weeks of remobilisation, hind-limb muscles do not return to preimmobilisation weights, although biochemical activities and ultrastructural appearance are largely restored. PMID- 10530860 TI - The regeneration of sensory neurones in the reconstruction of the anterior cruciate ligament. AB - We examined whether somatosensory evoked potentials (SEPs) were detectable after direct electrical stimulation of injured, reconstructed and normal anterior cruciate ligaments (ACL) during arthroscopy under general anaesthesia. We investigated the position sense of the knee before and after reconstruction and the correlation between the SEP and instability. We found detectable SEPs in all ligaments which had been reconstructed with autogenous semitendinosus and gracilis tendons over the past 18 months as well as in all cases of the normal group. The SEP was detectable in only 15 out of 32 cases in the injured group, although the voltages in the injured group were significantly lower than those of the controls. This was not the case in the reconstructed group. The postoperative position sense in 17 knees improved significantly, but there was no correlation between it and the voltage. The voltage of stable knees was significantly higher than that of the unstable joints. Our findings showed that sensory reinnervation occurred in the reconstructed human ACL and was closely related to the function of the knee. PMID- 10530862 TI - The connective-tissue envelope in revascularisation of patellar tendon grafts. AB - Free patellar tendon grafts used for the intra-articular replacement of ruptured anterior cruciate ligaments (ACL) lack perfusion at the time of implantation. The central core of the graft undergoes a process of ischaemic necrosis which may result in failure. Early reperfusion of the graft may diminish the extent of this process. We assessed the role of peritendinous connective tissue in the revascularisation of the patellar tendon graft from the day of implantation up to 24 days in a murine model using intravital microscopy. The peritendinous connective-tissue envelope of the graft was either completely removed, partially removed or not stripped before implantation into dorsal skinfold chambers of recipient mice. Initial revascularisation of the grafts with preserved peritendinous connective tissues began after two days. The process was delayed by five to six times in completely stripped patellar tendons (p < 0.05). Only grafts with preserved connective tissues showed high viability whereas those which were completely stripped appeared to be subvital. The presence of peritendinous connective tissues accelerates the revascularisation of free patellar tendon grafts. PMID- 10530861 TI - Characteristics of bone ingrowth and interface mechanics of a new porous tantalum biomaterial. AB - We have studied the characteristics of bone ingrowth of a new porous tantalum biomaterial in a simple transcortical canine model using cylindrical implants 5 x 10 mm in size. The material was 75% to 80% porous by volume and had a repeating arrangement of slender interconnecting struts which formed a regular array of dodecahedron-shaped pores. We performed histological studies on two types of material, one with a smaller pore size averaging 430 microm at 4, 16 and 52 weeks and the other with a larger pore size averaging 650 microm at 2, 3, 4, 16 and 52 weeks. Mechanical push-out tests at 4 and 16 weeks were used to assess the shear strength of the bone-implant interface on implants of the smaller pore size. The extent of filling of the pores of the tantalum material with new bone increased from 13% at two weeks to between 42% and 53% at four weeks. By 16 and 52 weeks the average extent of bone ingrowth ranged from 63% to 80%. The tissue response to the small and large pore sizes was similar, with regions of contact between bone and implant increasing with time and with evidence of Haversian remodelling within the pores at later periods. Mechanical tests at four weeks indicated a minimum shear fixation strength of 18.5 MPa, substantially higher than has been obtained with other porous materials with less volumetric porosity. This porous tantalum biomaterial has desirable characteristics for bone ingrowth; further studies are warranted to ascertain its potential for clinical reconstructive orthopaedics. PMID- 10530864 TI - Extensor carpi radialis brevis. An anatomical analysis of its origin. AB - We studied the origin of extensor carpi radialis brevis using 40 fresh frozen human cadaver specimens. Ten were stained with haematoxylin and eosin and trichrome which showed the collagenous structure of the extensor tendons at their origin. Gross anatomical observation showed that there was no definitive separation between brevis and communis at the osseotendinous junction. The histological findings confirmed the lack of separation between the two tendons. The extensor tendons were in close proximity to the joint capsule but trichrome staining showed no interdigitation of the tendon with the capsule. The validity of ascribing the pain of lateral epicondylitis to extensor carpi radialis brevis must be questioned. It appears to arise more from the 'common extensor' origin. PMID- 10530863 TI - c-Myc protein in the rabbit growth plate. Changes in immunolocalisation with age and possible roles from proliferation to apoptosis. AB - Growth plates taken from five- to 20-week-old Japanese white rabbits were immunostained for c-Myc protein. This was localised both in the proliferating zone and upper hypertrophic zone at five weeks, whereas after ten weeks it was found mostly in the lower hypertrophic zone. The proliferating chondrocytes tended to show nuclear staining and the hypertrophic cells cytoplasmic staining, although the terminal hypertrophic chondrocytes sometimes expressed the protein in their nuclei. In the younger rabbits, c-Myc co-localised with proliferating cell nuclear antigen, whereas in the hypertrophic zone of older rabbits, it was present in some chondrocytes the nuclei of which also contained DNA breaks. Our study suggests that, in the rabbit growth plate, c-Myc is associated with different cellular processes, depending on the age and the developmental stage of the chondrocytes. PMID- 10530865 TI - The tourniquet in total knee arthroplasty. PMID- 10530866 TI - The tourniquet in total knee arthroplasty. PMID- 10530867 TI - The tourniquet in total knee arthroplasty. PMID- 10530868 TI - Acute fractures of the scaphoid. PMID- 10530869 TI - Salvage of the head of the radius after fracture-dislocation of the elbow. PMID- 10530870 TI - The pathology of bone allograft. PMID- 10530872 TI - The next wave: protein tyrosine phosphatases enter T cell antigen receptor signalling. AB - Recent years have seen an exponentially increasing interest in the molecular mechanisms of signal transduction. Much of the focus has been on protein tyrosine kinase-mediated signalling, while the study of protein tyrosine phosphatases has lagged behind. We predict that the phosphatases will become a "hot topic" in the field within the next few years. This review summarizes the current state-of-the art in our understanding of the structure, regulation and role of protein tyrosine phosphatases in T lymphocyte activation. PMID- 10530871 TI - Integrin signalling in neutrophils and macrophages. AB - Integrins have been characterized extensively as adhesion receptors capable of transducing signals inside the cell. In myelomonocytic cells, integrin-mediated adhesive interactions regulate different selective cell responses, such as transmigration into the inflammatory site, cytokine secretion, production or reactive oxygen intermediates, degranulation and phagocytosis. In the last few years, great progress has been made in elucidating mechanisms of signal transduction by integrins in neutrophils and macrophages. This review summarises the current information on the role of integrins in regulating myelomonocytic cell functions and highlights the signalling pathways activated by integrin engagement in these cells. Also, exploiting the current knowledge of mechanisms of integrin signal transduction in other cell types, we propose a model to explain how integrins transduce signals inside neutrophils and macrophages, and how signaling pathways leading to regulation of selective cell functions may be coordinated. PMID- 10530873 TI - Functional properties of Ca2+-inhibitable type 5 and type 6 adenylyl cyclases and role of Ca2+ increase in the inhibition of intracellular cAMP content. AB - Among the different adenylyl cyclase (AC) isoforms, type 5 and type 6 constitute a subfamily which has the remarkable property of being inhibited by submicromolar Ca2+ concentrations in addition to Galphai-mediated processes. These independent and cumulative negative regulations are associated to a low basal enzymatic activity which can be strongly activated by Galphas-mediated interactions or forskolin. These properties ensure possible wide changes of cAMP synthesis. Regulation of cAMP synthesis by Ca2+ was studied in cultured or native cells which express naturally type 5 and/or type 6 AC, including well-defined renal epithelial cells. The results underline two characteristics of the inhibition due to agonist-elicited increase of intracellular Ca2+: i) Ca2+ rises achieved through capacitive Ca2+ entry or intracellular Ca2+ release can inhibit AC to a similar extent; and ii) in a same cell type, different agonists inducing similar overall Ca2+ rises elicit a variable inhibition of AC activity. The results suggest that a high efficiency of AC regulation by Ca2+ is linked to a requisite close localization of AC enzyme and Ca2+ rises. PMID- 10530874 TI - Dexamethasone differentially regulates cytokine transcription and translation in macrophages responding to bacteria or okadaic acid. AB - Many microorganisms and microbial products induce expression of pro-inflammatory cytokines such as interleukin-1 (IL-1alpha/beta) and tumour necrosis factor-alpha (TNF-alpha) in macrophages, primarily by transcriptional activation. We show here, by using mouse macrophages in primary culture, that pre-treatment with dexamethasone inhibits bacteria-induced IL-1beta expression as mRNA and cellular pro-IL-1beta in parallel, consistent with an effect primarily on transcriptional activation. In contrast, the expression of TNF-alpha mRNA was only partly inhibited despite virtually complete inhibition of TNF-alpha protein formation. Furthermore, the selective induction of primarily cell-associated 26,000 M, pro TNF-alpha by the protein phosphatase inhibitor okadaic acid also was partly inhibited at the mRNA level by dexamethasone, whereas additional translational inhibition appeared to be lacking. This latter finding is reminiscent of earlier findings regarding signalling to activation of cytosolic phospholipase A2, which is sensitive to dexamethasone when elicited by bacteria, but not when elicited by okadaic acid. The present results raise the possibility that the inhibitory effect of dexamethasone on TNF-alpha translation, but not on transcriptional activation, is mediated by one or more okadaic acid-sensitive protein phosphatases. PMID- 10530875 TI - Effects of the phosphorylation of myosin phosphatase by cyclic GMP-dependent protein kinase. AB - Cyclic GMP-dependent protein kinase (PKG) phosphorylated, in vitro, the large (MYPT1) and small (M20) regulatory subunits of myosin phosphatase (MP) with maximum stoichiometries of 1.8 and 0.6 mol of phosphate/mol subunit, respectively. The phosphorylation of these subunits by PKG did not affect the phosphatase activity towards the 20 kDa myosin light chain. However, phosphorylation of the MP holoenzyme decreased the binding of MP to phospholipid. The phosphorylation of the serine residue of the C-terminal part of MYPT1 was crucial for these interactions. These results suggest that the phosphorylation of MP by PKG is not a direct mechanism in activating MP activity, and that other indirect mechanisms, including the interaction between MP and phospholipids, might be candidates for Ca2+ desensitization via cGMP in smooth muscle. PMID- 10530876 TI - Roles of RhoA and phospholipase A2 in the elevation of intracellular H2O2 by transforming growth factor-beta in Swiss 3T3 fibroblasts. AB - We have investigated the mechanisms by which transforming growth factor-beta (TGF beta) increased intracellular H2O2 in Swiss 3T3 fibroblasts. Increase of intracellular H2O2 by TGF-beta was maximal at 30 min and blocked by catalase from Aspergillus niger. Scrape-loading of C3 transferase, which down-regulated RhoA, inhibited the production of H2O2 in response to TGF-beta. TGF-beta stimulated release of arachidonic acid, which was completely inhibited by mepacrine, a phospholipase A2 inhibitor. Mepacrine also blocked the increase of H2O2 by TGF beta. In addition, arachidonic acid increased intracellular H2O2. Furthermore, TGF-beta stimulated stress fibre formation, which was blocked by catalase, without membrane ruffling. Catalase also inhibited stimulation of thymidine incorporation by TGF-beta. These results suggested that TGF-beta increased intracellular H2O2 through RhoA and phospholipase A2, and also suggested that intracellular H2O2 was required for the stimulation of stress fibre formation and DNA synthesis in response to TGF-beta. PMID- 10530877 TI - Effects of dexamethosone and glucagon after long-term exposure on cyclic AMP phosphodiesterase 4 in cultured rat hepatocytes. AB - 67% of total cAMP phosphodiesterase activity (PDE) in cultured rat hepatocytes could be detected in the cytosol, 15% in plasma membrane, 15% in 'dense vesicle,' and 3% in endoplasmatic reticulum fractions. Up to 84% of the PDE activity of the cytosol is represented by the rolipram-sensitive PDE 4. ICI 118233-inhibited PDE 3 was found predominantly in membranes. We were able to show that dexamethasone acts on the PDE 4 in cytosolic and plasma membrane fractions whereas glucagon effected the PDE 4 of the cytosol and the PDE 3 in 'dense vesicle' membranes. Primary culture of hepatocytes was used to study long-term effects of dexamethasone and glucagon on PDE 4 activity. Addition of dexamethasone (0.1 microM) at the beginning of cultivation leads to a decrease of total PDE 4 activity whereas after 24 h precultivation no dexamethasone effect could be observed. Glucagon effects on PDE 4 were investigated in 20 h precultured hepatocytes. Maximal stimulation was achieved after 2 h of exposure. PDE 4 subtypes A, B , D and, to a lesser degree, subtype C could be detected by RT-PCR analysis. The results of semiquantitative RT-PCR show that the presence of dexamethasone during the first 24 h of cultivation reduced selectively the transcription of PDE 4D, whereas glucagon was without any effect. Also the translation of PDE 4D was reduced as shown in the Western blot. We would like to discuss the way that dexamethasone influences PDE 4D expression-most likely in combination with other factors such as cytokines--during the time of cell plating, whereas glucagon actions are part of metabolic regulations via phosphorylation reactions. PMID- 10530878 TI - Evidence for multiple rat VPAC1 receptor states with different affinities for agonists. AB - We compare the binding properties of [125I-VIP] and [125I]-Ro 25 1553 to VPAC1 receptors, expressed in stably transfected CHO cells. [125I]-VIP labelled two VPAC1 receptor states, while [125I]-Ro 25 1553 labelled selectively a limited number of high-affinity receptors. This high-affinity state probably corresponds to an agonist-receptor-Gs ternary complex as its properties (guanyl nucleotides, EC50 values and maximal effect) were affected by cholera toxin pre-treatment. Both high- and low-affinity receptors participated in the adenylate cyclase activation. This suggested that agonists activate not only low-affinity uncoupled receptors by facilitating the ternary complex formation, but also activated the high-affinity ternary complex by accelerating the GTP binding to emptied, receptor-bound G proteins. PMID- 10530880 TI - Rationalisation continues at the VLA. Veterinary Laboratories Agency. PMID- 10530879 TI - Forskolin inhibits 5-hydroxytryptamine-induced phosphoinositide hydrolysis and Ca+2 Mobilisation in canine cultured aorta smooth muscle cells. AB - The effect of forskolin on 5-hydroxytryptamine (5-HT)-induced inositol phosphate (IP) and Ca2+ mobilisation was investigated in canine cultured aorta smooth muscle cells (ASMCs). Pretreatment of ASMCs with forskolin attenuated 5-HT induced IP accumulation and Ca2+ mobilisation in a time- and concentration dependent manner. The half-maximal effects (pEC50) of forskolin to attenuate IP and Ca2+ responses to 5-HT occurred at concentrations of 6.28 and 6.64, respectively. Pretreatment of ASMCs with cholera toxin caused a similar inhibition on 5-HT-induced responses. Even after treatment with forskolin for 24 h, the 5-HT-induced responses were still inhibited. The inhibitory effect of forskolin resulted from both a depression of the maximal response and a shift to the right of the concentration-effect curves of 5-HT in these responses. The water-soluble forskolin analogue L-858051 [7-deacetyl-7beta-(gamma-N methylpiperazino)-butyryl forskolin] significantly inhibited the 5-HT-stimulated IP accumulation. In contrast, the addition of 1,9-dideoxy forskolin, an inactive forskolin analogue, had little effect on IP response. Moreover, SQ-22536 [9 (tetrahydro-2-furanyl)-9-H-purin-6-amine], an inhibitor of adenylate cyclase, and both H-89 [N-(2-aminoethyl)-5-iosquinolinesulphonamide] and HA-1004 [N-(2 guanidinoethyl)-5-iosquinolinesulphonamide], inhibitors of cAMP-dependent protein kinase (PKA), attenuated the ability of forskolin to inhibit the 5-HT-stimulated accumulation of IP in ASMCs. These results indicate that activation of cAMP/PKA might inhibit the 5-HT-stimulated IP accumulation and consequently reduce Ca2+ mobilisation, or inhibit both responses independently. PMID- 10530881 TI - Biological and migrational characteristics of transponders implanted into beagle dogs. AB - Ninety transponders (microchips), 45 made of bioglass, 30 made of acid-etched bioglass, and 15 made of bioglass and provided with a polypropylene cap, were inserted into 15 dogs in six different locations: on the left and right side of the head, and on the left and right shoulders, both cranial and caudal to the dorsum. The transponders were left in place for 16 weeks, during which their position was determined by means of an electronic reader and radiographs, and they were then retrieved and examined histologically. A clinical evaluation revealed that about half of the transponders inserted in all the shoulder locations had migrated to some extent, whereas the transponders in the head location had hardly moved. There were no differences in the extent of migration between the different types of transponders. Histological analysis showed that almost all the transponders were surrounded by a thin fibrous capsule with no sign of any gross inflammatory reaction. PMID- 10530882 TI - Monitoring follicular development in cattle by real-time ultrasonography: a review. AB - The application of real-time ultrasonography to monitoring ovarian function in mammals has advanced the understanding of follicular dynamics and its regulation. Follicular development is a wave-like sequence of organised events. The waves consist of the synchronous growth of small (4 to 5 mm) antral follicles, followed by the selection and growth of one dominant follicle which achieves the largest diameter and suppresses the growth of the subordinate follicles. In the absence of luteal regression, the dominant follicle eventually regresses (becomes atretic) and a new follicular wave begins. The dominant follicle regulates the growth of the subordinate follicles, because the appearance of the next wave is accelerated if the dominant follicle is ablated, and delayed if the lifespan of the dominant follicle is prolonged. During bovine oestrous cycles, two or three successive waves emerge, on average, on the day of ovulation (day 0) and day 10 for two-wave cycles, and on days 0, 9 and 16 for three-wave cycles. During the oestrous cycle there are thus two or three successive dominant follicles, and the last of these ovulates. Ovarian folliculogenesis is a complex process involving interactions between pituitary gonadotrophins, ovarian steroids and non-steroidal factors. Subtle changes in the hormonal milieu regulate folliculogenesis and the emergence of a follicular wave is preceded by a small increase in the concentration of plasma follicle-stimulating hormone. The mechanisms that promote the selection of a dominant follicle have not been elucidated, but considerable progress has been made in understanding follicular development and its regulation. Most treatments designed to control the development of follicular waves have been based on the physical or hormonal removal of the suppressive effect of the dominant follicle, and the consequent controlled induction of the emergence of a new follicular wave. The studies reviewed here describe current methods for regulating the bovine ovarian cycle, interesting models for future studies, and information that may be used for improving reproductive efficiency. PMID- 10530884 TI - Seroprevalence of Theileria ovis in small ruminants in north-east Spain determined by the indirect fluorescent antibody test. PMID- 10530886 TI - Slaughter of infant calves. PMID- 10530883 TI - Acid-base disorders in milk-fed calves with chronic indigestion. AB - Acid-base disorders were investigated in 50 calves with chronic indigestion and metabolic acidosis. In the calves that were unable to stand up, the acidosis was significantly more severe than in the calves that could stand up. The anion gap and four different components of the base excess were calculated by the method described by Fencl. The anion gap was high in more than half of the calves, and it was significantly associated with the base excess due to unidentified anions. However, in seven of the calves, the excess of unidentified anions would not have been detected without the calculations, which made it possible to measure the effect of sodium, chloride, plasma protein and unidentified anions on the acid base balance. Twenty-four of the calves had a combination of hyperchloraemic and high anion gap metabolic acidosis. Changes in sodium and plasma protein concentrations had a minor impact on the calves' acid-base status. PMID- 10530885 TI - Comparative study of serological tests for the diagnosis of equine aspergillosis. PMID- 10530887 TI - Use of unlicensed medicines. PMID- 10530888 TI - Uterine torsion in a cow. PMID- 10530889 TI - Study of lead and zinc toxicity in swans PMID- 10530890 TI - Effects of seasonality on xenobiotic and antioxidant defence mechanisms of bivalve molluscs. AB - Levels of chemical pollutants in the environment often display wide seasonal variation in response to climatic and other factors. Use of bioindicators such as enzyme activities in biomonitoring studies is complicated by this variation. Many such enzyme activities themselves show considerable seasonal fluctuation and there is known to be seasonality also in natural exposure to oxidative stress. This review attempts to explore some consequences of seasonal variation for biomonitoring studies with bivalve molluscs. It is suggested that independence of seasonal variation should be seen as a desirable feature of a bioindicator molecule. Where such molecules show seasonal variation, however, this should be incorporated into interpretation of biomonitoring studies by the use of appropriate controls. PMID- 10530891 TI - Treatment with 6-hydroxydopamine in planaria (Dugesia gonocephala s.l.): morphological and behavioral study. AB - Morpho-functional and behavioral effects of exposure to 6-hydroxydopamine (OHDA) HCI (24 microg/ml per day for 24 h and 7 days) were studied in planarias (Dugesia gonocephala s.l.). Exposure to 6-OHDA-HC1 for 24 h produced hypokinesia of the specimens. These behavioral changes were more pronounced, leading to complete immobility, after 7 days of exposure to the neurotoxin. Moreover, specimens exposed to 6-OHDA-HCI for 24 h and 7 days failed to show any behavioral response to nomifensine, thus furnishing evidence of the damage of presynaptic dopamine terminals. Exposure to 6-OHDA-HCl for 24 h significantly reduced cathecolamine content in neuropil region, as demonstrated by histochemistry, and electron-dense presynaptic vesicles, as observed on electron microscopy examination. All these alterations were significantly more pronounced and were accompanied by swelling and strong increase of electron-density in cytoplasm of numerous neurons after exposure to the neurotoxin for 7 days. This appears to be the first demonstration of the neurotoxic effects of 6-OHDA-HCI in flatworms. PMID- 10530892 TI - Effect of zinc on carp (Cyprinus carpio L.) erythrocytes. AB - The effect of zinc exposure on some properties of the carp erythrocyte membrane was studied in vitro. Red blood cells plasma membranes were separated from other cellular membranes using a combination of differential and density gradient centrifugation. The purity of obtained plasma membrane preparations was determined by measuring the activity of the marker enzymes. Electrophoretic patterns of the main erythrocyte membrane proteins excluded their degradation during the isolation and purification procedure. Carp erythrocyte membranes, obtained from cells previously incubated with increasing ZnSO4 concentrations, were used to elucidate the effect of zinc ions on their physical and biochemical properties. Using fluorescent probes: 12-AS and TMA-DPH, we found that zinc ions reduced the fluidity of the lipid bilayer, both in the middle and near the aqueous interface. Moreover, it was observed that zinc had no significant influence neither on the Na,K-ATPase activity nor on the thiol groups content in the erythrocyte membrane. We also detected that incubation of erythrocytes with zinc lead to the marked decrease of hemolytic resistance of the cells. Our studies demonstrate that zinc at higher concentrations may be toxic to carp erythrocytes causing changes in the membrane fluidity and hemolytic resistance. PMID- 10530893 TI - Eicosanoids mediate nodulation reactions to bacterial infections in larvae of the butterfly, Colias eurytheme. AB - Nodulation is the first, and qualitatively predominant, cellular defense reaction to bacterial infections in insects. Treating larvae of the butterfly Colias eurytheme with the eicosanoid biosynthesis inhibitor dexamethasone, strongly impaired nodulation reactions to bacterial infections. The influence of dexamethasone was reversed by treating infected insects with arachidonic acid, an eicosanoid precursor. An eicosanoid biosynthesis system in C. eurytheme larvae is documented. Specifically, the presence of eicosanoid-precursor polyunsaturated fatty acids in tissue phospholipids was determined, an intracellular phospholipase A2 that can release arachidonic acid from tissue phospholipids was recorded, and eicosanoid biosynthesis, registered as conversion of exogenous radioactive 20:4n-6 into eicosanoids, was observed. These findings support the hypothesis that eicosanoids mediate cellular immune responses to bacterial infections in these butterfly larvae, and more broadly, in most, if not all, insects. PMID- 10530894 TI - Regional variation of white adipocyte lipolysis during the annual cycle of the alpine marmot. AB - During winter, hibernating animals rely on their lipid stores for survival. In vitro lipolytic activity of isolated adipocytes from gonadal and subcutaneous white adipose tissue (WAT) was studied in captive alpine marmots (Marmota marmota) at two different times of their yearly cycle. During the summer, when marmots were eating, adipocyte responsiveness and sensitivity to isoprenaline and noradrenaline were higher in gonadal than in subcutaneous WAT. During hibernation, when marmots were spontaneously fasting. both the response and sensitivity to catecholamines decreased in gonadal WAT to the level of subcutaneous WAT. A similar pattern of response was also observed when lipolysis was stimulated with glucagon but the lipolytic rate was three times lower than with catecholamines. Adenosine deaminase (ADA) had a marked stimulatory effect on lipolysis, especially during the 'feeding' period, suggesting that adenosine may be a potent lipolytic modulator in marmot adipocytes. It is concluded that in marmots, lipolysis could be differentially regulated between fat depots during the annual cycle possibly to optimize either the building-up or the use of fat reserves. PMID- 10530895 TI - Sex-hormone binding globulin from sheep serum: purification and effects of pregnancy and treatment with exogenous estradiol. AB - Sex-hormone binding globulin (SHBG) is a protein that binds sex steroids in the serum of many species. SHBG binds androgens and estrogens in humans and primates with high affinity, but behaves as an androgen binding protein in other species. Here we purified SHBG from ewe and ram sera to homogeneity, by a simple and rapid method. The K(D) of the purified protein was found to be 3.63 nM for testosterone and around 600 nM for estradiol. We also studied the effect of pregnancy on SHBG levels in ewes and the effect of exogenous estradiol administration either orally or parenterally on SHBG levels in rams. Basal levels of SHBG in sheep are not affected by pregnancy or exposure to exogenous estradiol. It is concluded that SHBG regulation of expression in ewes and rams differs from that in humans in that it is not affected by estrogen and possibly is species specific. PMID- 10530896 TI - Quantitative structure-activity relationships for phosphoramidothioate toxicity in housefly. AB - Quantitative structure activity relationships were formulated for a series of phosphoramidothioate (Ace) analogs. Ace II- and Ace IV-induced inhibition of fly head AChE was influenced by the spatial configuration of the inhibitor. The 3D structure of a potent phosphoramidothioate such as methamidophos was such that its P-O(-)group interacted with the enzyme's 'oxy-anion hole', NH2+ group formed an H-bond with the enzyme's H-bonding site, and leaving group (-S) oriented toward the AChE 'gorge' opening. An alteration in its 3D structure also altered its toxicity. The k(i) for Ace II-induced inhibition of fly-AChE correlated with sterimol indices L1 (the substituent's length), B1 (minimum axis perpendicular to this length), and B3 (-90 degrees to B1) and Ebend. The k(i) for Ace IV-induced inhibition of fly-AChE correlated with dispersion, H-bond donor, and E(dihedral). Thus, the 3D characteristics of a substituent and molecular charge play a key role in the inhibition of fly-head AChE by phosphoramidothioate. LD50 for housefly exposed to Ace I, II, III, and IV analogs was governed by their electronic, topological, steric, and sterimol (steric effects of substituents) properties. Hydrophobic interaction either played a minor role or adversely affected their toxicity in housefly. PMID- 10530897 TI - Antioxidant processes of the wild tambaqui, Colossoma macropomum (Osteichthyes, Serrasalmidae) from the Amazon. AB - Colossoma macropomum, locally called tambaqui, is a freshwater migratory teleost that shows good tolerance to oxygen and pH changes in water, and both chemical physical parameters change markedly during the day time and seasonal water level oscillations in the Amazon Basin. In order to obtain a general view about the basal levels of antioxidants in different tissues of wild tambaqui, enzymatic (superoxide dismutase and catalase) and non-enzymatic (alpha-tocopherol, beta carotene, and glutathione) antioxidants and lipid peroxidation levels were assessed in the liver, blood and plasma of ten specimens collected during the dry season (September) in a pond near Manaus-AM, Brazil. Superoxide dismutase, catalase, and lipid peroxidation levels were high in the liver and low in the blood and plasma. Confirming previous results on tambaqui, catalase was detected in the blood of one specimen only. beta-Carotene was not found in any analyzed tissue, while alpha-tocopherol was found only in the liver (7.8 +/- 7.0 nmol g( 1), mean +/- S.E.M.) and plasma (4.3 +/- 0.9 nmol ml(-1)). Blood glutathione concentrations (2.4 +/- 0.17 mmol l(-1)) of tambaqui were comparable with those found in other Amazonian teleosts. Antioxidant defenses and lipid peroxidation contents from liver. blood and plasma exhibited interesting correlations. These relationships suggest that antioxidant defenses located in different tissues and in different sub-cellular compartments act in concert. PMID- 10530898 TI - Possibility of inhibin as a regulator of androstenedione production by the ovary during the period of delayed ovulation in a vespertilionid bat, Scotophilus heathi. AB - The aim of the present study was to evaluate the role of inhibin in regulating androstenedione production by the ovary of Scotophilus heathi during the period of delayed ovulation. Inhibin alone increased the androstenedione synthesis in vitro by the ovary during all reproductive phases, but augmented the hCG induced androstenedione production only during November. The follicles produced significantly higher androstenedione as compared to the stromal or granulosa cell in response to both hCG and inhibin. However, inhibin augmented hCG stimulated androstenedione in the stromal cell only. Immunoreactivity of inhibin subtype alpha, betaA and betaB were mainly localized in the thecal and interstitial cell from September to November and then declined during the preovulatory period. However, in granulosa cells weak to moderate immunoreactivity of all inhibin subtype was observed during these phases. These results indicate the possible role of inhibin in regulating androstenedione synthesis by the ovary and thus. may be indirectly responsible for causing delayed ovulation in S. heathi. PMID- 10530899 TI - Effects of starvation on liver microsomal P450 activity in juvenile Pleuronectes americanus. AB - Recent work has produced evidence to support the existence of a cytochrome P450 CYP2E1-like isoform in the marine fish, Pleuronectes americanus (winter flounder) (Wall K, Crivello JF. Toxicol Appl Pharmacol 1998;151:98-104). Starvation has been previously demonstrated to induce CYP2E1 activity (assayed as chlorzozazone 6-hydroxylase activity) in mammals and this study was undertaken to determine the effects of starvation on liver chlorzozaxone-6-hydroxylase and ethoxy-resorufin-O deethylase activity (a CYP1A1 activity) in juvenile winter flounder liver microsomes. A 2-week starvation period resulted in a statistically significant increase in liver microsomal protein, and a decrease in liver lipid and liver glycogen. Ethoxy-resorufin-O-deethylase activity (pmol/min/mg microsomal protein) was reduced with starvation, chlorzoxazone 6-hydroxylase activity (pmol/min per mg microsomal protein) initially decreased but then increased over controls. When these activities were expressed per gm/liver (to account for the starvation induced changes in liver microsomal protein), chlorzoxazone 6-hydroxylase activity doubled over control during starvation but ethoxy-resorufin-O-deethylase was not significantly changed. The effects of starvation on liver microsomal chlorzoxazone 6-hydroxylase and ethoxy-resorufin-O-deethylase activities are discussed in the context of the impact of physiological states on the ability of fish to detoxify marine xenobiotics. PMID- 10530900 TI - Pharmacokinetics of intravenously administered ketamine in southern elephant seals (Mirounga leonina). AB - Southern elephant seals (Mirounga leonina) are large, potentially dangerous animals which must be restrained before study or treatment. However, chemical restraint is unpredictable, possibly because of differences in body composition during fasting ashore, and circulatory adaptations to enable diving. The pharmacokinetics of ketamine (1.1 mg/kg i.v.) was studied in 15 southern elephant seals which had come ashore on Macquarie Island. The animals were first sedated with pethidine (5 mg/kg i.m.) to allow intravenous access. There was great variability in the calculated pharmacokinetic parameters, possibly due to circulatory changes associated with periods of apnoea which characterise this animal's response to anaesthetics. The median values were similar to those reported for other species: distribution t1/2 = 2.5 min (range 1-11 min); elimination t1/2 = 43 min (range 17-108 min); apparent volume of distribution, 1474 ml/kg (range 830-9301 ml/kg); clearance, 33 ml/min/kg (range 12-57 ml/min/kg). This was the first investigation of drug disposition in any seal species. and the first in any free-ranging wild animal. It is important to obtain pharmacological data in animals in whom drugs are used, but pre-catheterised captive animals may provide less variable data. PMID- 10530901 TI - Heparin-binding molecules with growth factor activities in regenerating-tissues of the starfish Asterias rubens. AB - Regenerating-tissues of the starfish Asterias rubens were studied for the presence of growth factors liable to stimulate the proliferation of fibroblast and epithelial cells (3T3, BHK21 and Hela cells). As a first attempt to isolate growth factors, the extracts were fixed on heparin-affinity column and were eluted by 1-1.2 M NaCl. After separation on a Vydac C18 HPLC column. a fraction that stimulates the proliferation of fibroblast cells was isolated. It contained four different peptides, separated by electrophoresis, and for which the amino acid composition and molecular mass were determined. All the peptides were lysine rich and one presented an amino-acid composition comparable to basic-fibroblast growth factor (b-FGF) while its molecular weight was higher. PMID- 10530903 TI - The relationship in neonates between clinically measured head circumference and brain volume estimated from head CT-scans. AB - OBJECTIVE: To determine if the frontal-occipital head circumference correlates with brain volume on CT and to investigate correlations between the volumes of different brain subdivisions in live neonates. METHODS: Records were studied from 27 neonates with anatomically normal head CT-scans which were ordered for clinical reasons, and which were performed at Johns Hopkins Hospital. Clinical data were abstracted from medical records. Brain volumes were estimated by digitizing the structures of interest on each slice of head CT-scans. RESULTS: In this sample of 27 infants with a mean birth weight of 3000.4 +/- 668 g, mean head circumference of 33.5 +/- 1.8 cm, and mean gestational age of 37 weeks and 4 days +/- 24 days, the mean of total brain volume was 333.0 +/- 78.3 ml. The correlation between clinically measured head circumference and total brain volume was 0.55 (P < 0.003). Regression of total brain volume on head circumference and its second and third powers accounted for 43% of the variation in total brain volume. Other predictor variables, namely infant race, sex, gestational age, and maternal age, were not significant in this regression once head circumference was included. The slope of the cubic function of head circumference as a predictor of brain volume was greatest below the mean head circumference of 33.5 cm. Brain volume leveled off at head circumferences greater than the mean. CONCLUSION: Head circumference is a powerful predictor of total brain volume in the neonate: below the approximate head-circumference mean of 33.5 cm, smaller head circumference indicates smaller total brain volume. PMID- 10530902 TI - A risk factor for early-onset infection in premature newborns: invasion of chorioamniotic tissues by leukocytes. AB - The authors report a prospective study of correlation between histopathological alterations of the placenta, risk factors and early-onset bacterial infections in 224 premature newborns. They used a mathematical model for evaluation and prediction of neonatal bacterial infection according to the localization in chorioamniotic tissues (chorioamniotic plate, amniotic membranes and umbilical cord) invaded by leukocytes. Septicemia, pneumonia or omphalitis were documented in 45 (20%) infected premature newborns and inflammatory lesions in the placenta were observed in all of them. In order of statistical significance, the most important variables for early-onset bacterial neonatal infection were invasion of the chorioamniotic plate, amniotic membranes and umbilical cord tissues by PMNL (P < 0.0000), premature rupture of membranes (P < 0.0000), birthweight lower than 1500 g (P < 0.0000), gestational age under 34 weeks (P < 0.0001), foul smell (P < 0.0038), no antibiotics before delivery (P < 0.0066) and intrapartum fever (P < 0.0087). By logistic stepwise multiple regression analysis, invasion of fetal chorioamniotic plate and of amniotic membranes by leukocytes were the only statistically significant variables. The probability of neonatal infection in premature newborns, when polymorphonuclear neutrophils were present in chorioamniotic plate and in amniotic membranes, was 62.5%, while the probability was 0.5% when these tissues were normal. These data suggest that histological chorioamnionitis has to be considered as an important risk factor for early-onset infection in premature newborns. PMID- 10530904 TI - Patterns of inflammatory cytokine serum concentrations during the perinatal period. AB - Inflammatory cytokines interleukin-1beta (IL-1beta), interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha) were measured in the serum of healthy, term neonates on the first (N1), fifth (N5) and 40th (N40) day after birth, compared with those in maternal serum (MS), umbilical cord (UC) and in adult controls. All three cytokines were significantly elevated in N1 and N5, compared with those in UC and adults (P < 0.0001). IL-1beta and IL-6 declined significantly from N1 to N40 (P < 0.0001), while TNF-alpha increased significantly from N1 to N5 and declined thereafter. TNF-alpha values in UC were significantly higher than in adults, but lower than in N40 (P < 0.0001), while IL 1beta and IL-6 values in UC did not differ from those in N40 and in adults. IL 1beta and IL-6, but not TNF-alpha values in MS were significantly higher than those in controls (P < 0.0001). IL-1beta values in MS were significantly higher than those in N1 (P < 0.0001), while those of IL-6 and TNF-alpha were significantly lower (P < 0.0001). Moreover, IL- 1beta values were dependent on the mode of delivery in N1 (P < 0.001), in MS (P < 0.02) and in UC (0.03), while IL-1beta and TNF-alpha values in N1 were strongly interrelated (r = 0.7; P < 0.01). In conclusion, the increased values of IL-1beta, IL-6 and TNF-alpha during the perinatal period might reflect a newborn immune response to the stress of delivery and to environmental changes after birth. PMID- 10530905 TI - Cord blood concentrations of vitamin A in preterm infants. AB - Plasma vitamin A concentrations were measured in cord blood samples from 56 infants of gestational ages < 33 weeks. Outcome was followed prospectively. Mothers' dietary habits and use of multivitamins during pregnancy were evaluated by means of a questionnaire. Vitamin A concentrations less than 1.05 micromol/l (low) were measured in 22, but levels below 0.7 micromol/l (deficient) only in two cases. The concentrations were not correlated with the infants' gestational ages. Infants with low concentrations were significantly more often multiplets compared to those with normal levels and the vitamin A concentrations of the multiplets were significantly lower than those of the singletons. The outcome measures used and the mothers' dietary habits and multivitamin use were similar in cases with low and normal vitamin A concentrations. Multiple gestation seems to be correlated with low plasma vitamin A concentrations in preterm infants at birth, and a complete assessment of vitamin A status to detect possible deficiency might be indicated in these cases. PMID- 10530906 TI - Blood pressure rhythms during the perinatal period in very immature, extremely low birthweight neonates. AB - The aim of this study was to investigate if blood pressure (BP) rhythms were present in the perinatal period in very immature infants. Twenty-two infants, median gestational age 24-28 weeks, who had indwelling arterial lines with undamped arterial BP waveforms, were studied. The infants were all receiving intensive care under constant conditions. The hourly mean, systolic and diastolic BPs on days 2 and 7 were examined. A cosinor analysis of the mean BP was performed examining period lengths of 4, 8, 12, 16, 20, and 24 h to determine whether ultradian and/or circadian rhythms existed. On day 2, but not day 7, the mean and systolic BPs showed significant variation and circadian and ultradian rhythms were demonstrated. We suggest that maternal influences may be responsible for the BP rhythms noted in very immature infants on day 2. PMID- 10530907 TI - Quantitative analysis of fetal general movements: methodological considerations. AB - OBJECTIVES: We studied the effects of gestational age and various smoothing procedures on four incidence parameters of fetal general movement, to evaluate reported variation in previous studies and to establish the optimal way of smoothing. SUBJECTS AND METHODS: General movements were studied longitudinally between 24 and 40 weeks of gestation in 29 healthy fetuses. The number of movement bursts per hour, burst duration, onset-onset interval between successive bursts (OOI) and the percentage incidence were analysed in detail. RESULTS: Advancing gestation was characterised by a proportional increase in OOI's lasting > 60 s and a decreased number of bursts, whereas burst duration remained relatively stable (unsmoothed data). Smoothing resulted in an exaggerated decrease in the number of bursts and in increases in burst duration, OOI and percentage incidence. These changes occurred in a gestational age specific manner and could largely explain the variation in results between previous studies. CONCLUSIONS: The temporal patterning of fetal general movements undergoes developmental change, as shown by differential effects of smoothing between mid and late pregnancy. A smoothing procedure is to be preferred which includes short intervals (1-3 s) between the elements composing a burst, since small changes in movement generation can still be recognised this way. PMID- 10530908 TI - Energy intake, appetite and body mass in infancy. AB - Energy intake in infancy depends on the infant's appetite, which, in turn, depends to a considerable extent on the infant's size, as size is an important determinant of energy expenditure. Previous work on six-week old breast-fed infants has suggested that, at this age, milk intake in infants is proportional to the square root of body weight (wt.(0.5)). In this paper, the form of the relationship between body weight and energy intake is examined in infants of different ages, using data from two longitudinal studies, one of initially breast fed and one of initially bottle-fed infants. In the first data set, energy intake is proportional to body weight raised to powers ranging from 0.63 to 1.23 at different ages and, in the second, to body weight raised to powers ranging from 0.50 to 1.07 at different ages. No single value is consistent with all the data at all ages. In general, the powers decrease up to six months of age, and then increase again, a pattern that may be due to the pattern of changes in the adiposity of the infants, as reflected in their body mass indexes (BMIs). PMID- 10530909 TI - Type I diabetes, also known as insulin dependent diabetes mellitus or juvenile diabetes, is one of the disorders with a complex genetic basis. PMID- 10530910 TI - Fluoride in whole saliva, parotid ductal saliva and plasma in children. AB - The purpose was to investigate the relation between fluoride concentrations in whole saliva, parotid ductal saliva, and plasma in 5- to 10-year-old children (n = 17). Two stimulated whole-saliva samples were obtained from each child. Before the second sample was obtained, each child rinsed several times with a total of 100 ml of deionized water. Parotid saliva samples were obtained by use of a Lashley cup. Fluoride concentrations were determined by fluoride ion-specific electrode after diffusion with hexamethyldisiloxane. Rinsing with deionized water did not significantly reduce the fluoride concentration in whole saliva. The whole-saliva fluoride concentrations were not significantly related to those in plasma or parotid ductal saliva. Parotid fluoride concentrations, however, were significantly related to plasma fluoride concentrations (p < 0.0001) by a proportionality constant of 0.80. It was concluded that parotid salivary fluoride concentrations can be used to estimate plasma fluoride concentrations in 5- to 10 year-old children. PMID- 10530911 TI - Isolation of bacteria from cervical lymph nodes in patients with oral cancer. AB - Thirty patients with oral mucosal cancer were studied in relation to oral mucosal damage and bacterial translocation to the regional lymph nodes in the neck. All 30 patients (gingiva 11, tongue 13, cheek mucosa four, oral floor two) underwent extensive, clean-contaminated, head-and-neck surgery (including neck dissection) with free flap reconstruction. A total of 153 lymph nodes was harvested for microbial and histological examination. Viable bacteria were isolated from 70 lymph nodes (45.8%) from 25 patients (83.3%). Bacterial cells in the nodes were detected by scanning electron microscopy. Bacterial translocation was found more often in metastatic nodes (75.0%) than in uninvolved nodes (40.3%) (p = 0.015; chi2 test). Gingival carcinoma yielded 56.4% of bacterial growth in the regional lymph nodes compared to tongue (40.3%), oral floor (40.0%) and cheek mucosa (37.5%). As the gingival carcinoma group includes more T4 cases (11/11; 100%) than the other three groups (7/19; 36.8%), bacterial translocation in uninvolved nodes could be caused by the size and invasion of the primary oral tumor. Oral streptococci (Streptococcus intermedius, Strep. constellatus, Strep. oralis, Strep. mitis, Strep. sanguis, Strep. salivarius) were the most common isolates. Aerobic enteric bacteria (Enterococcus, Escherichia, Klebsiella etc.) were also found in the lymph nodes. Among the anaerobic bacteria, Peptostreptococcus spp. were isolated from 12 patients. Damaged oral mucosa in patients with oral cancer might allow the new bacterial colonization on the surface and subsequently drain the bacteria into the regional lymph nodes as well as the general circulation. PMID- 10530912 TI - A statistical analysis of the overexpression of the msx2 RNA in Xenopus laevis. AB - The msx family of genes are important during development, and implicated in the development of various craniofacial structures. Here the frog, Xenopus laevis, was used to perform gain-of-function experiments to obtain further insight into the role of the msx2 gene. mRNAs of wild-type and mutated forms of the msx2 gene were injected into developing Xenopus embryos. Phenotypic changes in these embryos were noted, scored, and subjected to statistical analyses. Overexpression of the wild-type form of the msx2 gene resulted in embryos that were ventralized, i.e. with loss of anterior structures including head and eyes. A surprising finding was the statistically significant difference in phenotypic changes (p < 0.001 when compared to a buffer-injected group) of embryos microinjected with the mRNA of a mutated form of the msx2 gene (without the homeobox region). It is proposed that the msx2 overexpression system can be used as a consistent and reliable bioassay to map and study the functions of the msx2 gene during development, especially of the craniofacial region. PMID- 10530913 TI - Haemodynamic responses in chronically painful, human trapezius muscle to cold pressor stimulation. AB - The aim was to compare haemodynamic responses in trapezius muscles to cold pressor stimulation in individuals with localized trapezius myalgia and asymptomatic controls. Nine males with chronic localized pain in the trapezius (mean age, 23.2 years) and nine male controls (mean age, 24.6 years) who had no medical history of migraine, hypertension or sustained pain in the trapezius region were investigated. Two experimental (cold pressor and mock) trials were performed in a randomly assigned sequence. In the cold pressor trial the participant's left foot and ankle were immersed in 4 degrees C cold water for 2 min; the mock trial was done without that stimulus. Blood volume was continuously recorded 1 min before, 2 min during, and 5 min after cold pressor stimulation using near-infrared spectroscopy. Each participant's blood-volume data were baseline-corrected and submitted to statistical analysis. Results showed that the individuals with muscle pain exhibited a significantly lower mean blood volume than the controls during cold pressor stimulation (p = 0.0367). Upon withdrawal of that stimulation, the mean blood volume in both groups fell below the baseline. These results suggest that individuals with chronic regional trapezius myalgia have less capacity to vasodilate this muscle during cold pressor stimulation than those without such myalgia. It is not yet known if this difference in the haemodynamic response is a cause or an effect of the myalgia. PMID- 10530914 TI - A micromechanics model of the elastic properties of human dentine. AB - A generalized, self-consistent model of cylindrical inclusions in a homogeneous and isotropic matrix phase was used to study the effects of tubule orientation on the elastic properties of dentine. Closed-form expressions for the five independent elastic constants of dentine were derived in terms of tubule concentration, and the Young's moduli and Poisson ratios of peri- and intertubular dentine. An atomic-force microscope indentation technique determined the Young's moduli of the peri- and intertubular dentine as approx. 30 and 15 GPa, respectively. Over the natural variation in tubule density found in dentine, there was only a slight variation in the axial and transverse shear moduli with position in the tooth, and there was no measurable effect of tubule orientation. It was concluded that tubule orientation has no appreciable effect on the elastic behaviour of normal dentine, and that the elastic properties of healthy dentine can be modelled as an isotropic continuum with a Young's modulus of approx. 16 GPa and a shear modulus of 6.2 GPa. PMID- 10530915 TI - Retroviral transduction of human periodontal cells with a temperature-sensitive SV40 large T antigen. AB - The periodontal ligament (PDL) is considered to contain subpopulations of cells responsible for the development, repair and regeneration of the periodontium. Cell cultures have been used as model systems in order to understand the complex cellular and biochemical events underlying these processes. In order to obtain long-term cultures of these cells that can be cloned and characterized, primary cultures of PDL and gingival cells were infected with an amphotropic retroviral construct encoding a temperature-sensitive SV40 large T antigen (tsT). After selection for drug resistance, the cells expressed the T antigen and proliferated at 34 degrees C for more than 40 passages. However, when the T antigen was inactivated by incubation at 39 degrees C, the cultures became growth-arrested and the granularity of the cells increased, possibly as a result of differentiation. Reverse transcribed-polymerase chain reaction and flow cytometry showed that the tsT-transduced cells expressed a number of soft and hard connective-tissue antigens, including osteocalcin, osteonectin, osteopontin, collagen type I and alkaline phosphatase. Moreover, incubation of the transduced PDL cells at 39 degrees C was found to upregulate the expression of osteocalcin, osteopontin and collagen type I, but downregulate osteonectin. At this temperature, the presence of the dexamethasone downregulated type I collagen, while vitamin D3 had no effect on the expression of any of the antigens examined. Under all culture conditions, antigen expression was far higher in the transduced PDL cells than the gingival cells. The findings thus show that growth of the tsT transduced PDL and gingival cells is temperature-dependent and that the presence of the T antigen increases their lifespan but does not ablate the expression of certain of their characteristic phenotypic and functional features. PMID- 10530916 TI - Cardiovascular responses in humans to experimental chewing of gums of different consistencies. AB - Although the cardiovascular effects of exercise have been extensively investigated in man, little attention has been paid to such responses to jaw muscle activity. The aim here was to investigate the general cardiovascular effects of chewing activity in a single-blind, cross-over design. Ten healthy individuals performed one of the following chewing tasks in four separate sessions: chewing a very hard gum, chewing a moderately hard gum, chewing a soft gum, and "empty chewing" without a bolus. Unilateral chewing of gum or empty chewing was performed for 20 min on the participant's most convenient chewing side at a constant rate of 80 cycles/min. In each session, heart rate and arterial blood pressure were recorded together with electromyographic activity in the masseter and anterior temporalis muscles on the chewing side. Ratings of perceived masticatory fatigue were recorded with visual analogue scales. The heart rate and blood pressure were significantly increased (ANOVA; p < or= 0.01) during the chewing tasks and the increases were, in parallel with the muscle activity, more pronounced the harder the gum. With the very hard gum, heart rate increased by up to 11 beats/min, the systolic blood pressure was 14 mmHg (1.9kPa) higher, and the diastolic blood pressure was 11 mmHg (1.5kPa) higher. The perceived fatigue was proportional to the level of muscle activity. After 10 min of recovery from exercise, heart rate and arterial blood pressures were slightly but still significantly elevated. The results demonstrate that chewing is associated with general circulatory effects proportional to the bolus resistance. PMID- 10530917 TI - Nicotine- and arecoline-induced interleukin-1 secretion and intercellular adhesion molecular-1 expression in human oral epidermoid carcinoma cells in vitro. AB - The purpose was to examine interleukin (IL)-1 concentrations and intercellular adhesion molecule (ICAM)-1 expression in nicotine/arecoline-exposed oral KB CCL17 cultures. Enzyme-linked immunosorbent assay was used to quantify IL-1 concentrations in culture supernatants. A repeated-measures analysis of variance was used to identify differences among the groups. IL-1 beta concentrations increased by 2.6, 2.7 and 7.5 times those of the control in groups treated with 1 microM nicotine, arecoline or with both, respectively. IL-1 beta concentrations were more dramatically increased when the agents tested were at 100 microM concentration. Similar, although less dramatic, alterations in IL-1 alpha concentrations were observed. The fluorescence intensity of ICAM-1 (CD54) analysed by flow cytometry was also significantly increased in a dose-dependent manner when the cells were treated with nicotine and/or arecoline. Nicotine and arecoline therefore significantly increased IL-1 alpha and -1 beta secretions and the surface expression of ICAM-1 in KB CCL17 cells. PMID- 10530918 TI - Distribution of parathyroid hormone-related protein (PTHrP) and type I parathyroid hormone (PTH) PTHrP receptor in developing mouse mandibular condylar cartilage. AB - The mandibular condylar cartilage undergoes endochondral bone formation and is an important growth site in the mandible. Parathyroid hormone-related protein (PTHrP) has received attention as a physiological regulator attenuating chondrocytic differentiation and preventing apoptotic cell death. In order to examine the localization of PTHrP and its receptor during fetal development of the condylar cartilage, an immunohistochemical study of PTHrP and the type I PTH/PTHrP receptor was carried out. At day 15 of gestation, the condylar cartilage was evident and some chondrocytes showed positive staining for PTHrP. At day 16, the cartilage was increasing in length and width, and PTHrP was localized in the flattened and hypertrophic cell layers. After day 17, when endochondral bone formation had already started, PTHrP was mainly observed in the flattened cell layer and in a few layers of the hypertrophic chondrocytes. The localization of the type I PTH/PTHrP receptor was similar to that of PTHrP on days 15 and 16, and was broadly distributed at day 18. Apoptotic chondrocytes were scarcely observed on days 15 and 16, and only a few cells were present in the erosion front at day 18. This temporal and spatial localization of PTHrP and the type I PTH/PTHrP receptor suggests that PTHrP is a possible autocrine/ paracrine factor regulating condylar chondrocytic differentiation during development. PMID- 10530920 TI - Characterization of the mucosal immune response in breast milk after peroral immunization of chimpanzees (Pan troglodytes) with Streptococcus mutans. AB - The characteristics of the mucosal immune response to Streptococcus mutans cells, antigen A, antigen B, glucosyltransferases and glucan-binding proteins were examined in four pregnant chimpanzees that had been immunized perorally with Strep. mutans. Six pregnant chimpanzees served as non-immunized controls. None of the chimpanzees harbored S. mutans. Samples of milk were collected from all animals throughout the experiment. Peroral immunization resulted in an overall 17 fold median increase in SIgA in milk. Although SIgA1 comprised almost two-thirds of milk SIgA, Strep. mutans whole-cell antibody activity was contained predominantly in the SIgA2 subclass. The difference between the specific activities of anti-Strep. mutans SIgA1 and SIgA2 antibodies compared over time reached the borderline of statistical significance (p = 0.08). The avidity of anti-Strep. mutans antibodies was low in three of four chimpanzees and there was no evidence of affinity maturation. SIgA antibodies from the milk of all four immunized chimpanzees recognized antigen A. In three animals these antibodies were restricted to the SIgA1 subclass and, in one animal, anti-A antibodies were confined to SIgA2. Antibodies from all of the immunized chimpanzees recognized degradation products of antigen B in both the SIgA1 and the SIgA2 subclasses. Only two of four immunized chimpanzees responded to glucosyltransferases and these antibodies were restricted to the SIgA1 subclass. None of the chimpanzees responded to the 74-kDa glucan-binding protein. However, three animals produced SIgA1 antibodies against the 59-kDa glucan-binding protein and two of these also produced SIgA2 antibodies against this protein. PMID- 10530919 TI - Tissue-non-specific alkaline phosphatase mRNA expression and alkaline phosphatase activity following application of retinoic acid in cultured human dental pulp cells. AB - Retinoic acid is a potent inducer of tissue-non-specific alkaline phosphatase (TNSALP) expression in various osteoblastic and fibroblastic cells, and may be involved in morphogenesis, cellular growth and differentiation. This study investigates the effects of retinoic acid on alkaline phosphatase activity and TNSALP gene expression in human dental pulp cells. Cultured cells were treated with various concentrations of retinoic acid (0, 10(-7), 10(- 6), 10 (-5) M) in 0.5% bovine serum albumin without serum. Alkaline phosphatase activity was determined by the rate of p-nitrophenyl phosphate hydrolysis and was also assayed in the presence of various inhibitors and under thermal inactivation. A set of specific oligonucleotide primers was selected, based on the nucleotide sequences of two human TNSALP mRNA (bone and liver) types, and reverse transcription polymerase chain reaction (RT-PCR) performed. Inhibitory and thermal inactivation experiments revealed that the elevated alkaline phosphatase activity had properties of the TNSALP type. RT-PCR showed that retinoic acid enhanced the expression of bone-type TNSALP mRNA in pulp cells. However, the liver-type TNSALP mRNA was not detected. These findings suggest that the high alkaline phosphatase activity of retinoic acid-treated dental pulp cells is associated with increased transcription of the bone-type mRNA of the TNSALP gene and not with liver-type. PMID- 10530921 TI - Experimental studies of human dentine wear. AB - Previous in-vitro studies have described the relation between rates of enamel wear and variables such as applied load and lubricant pH. The aim here was to extend understanding of tooth-wear processes by considering the rate of wear in human dentine. Enamel was removed from extracted third molar teeth that had been sectioned mesiodistally. Moisture fluctuation within dentine was minimized by conducting all procedures under copious irrigation or in sealed containers of lubricant at pH 7.0. Specimens were subjected to wear using a purpose-built apparatus at loads of 6.2, 9.95 and 13.2 kg. All experiments were done with a unidirectional wear stroke of 3 mm at a rate of 80 cycles/min for 75 min and repeated for 75 min. Dentine wear was assessed by specimen weight loss. At pH 7.0, wear rates ranged from 0.50 mg/10(3) cycles at a load of 6.2 kg to 0.77 mg/10(3) cycles when a load of 13.2 kg was applied. At higher loads, dentine wear rates were similar to those of enamel. Increasing load is thus associated with a progressive increase in the rate of dentine wear. This relation differs significantly from that for enamel, reflecting fundamental differences in the composition and structure of these tissues. PMID- 10530922 TI - Evaluation of [18F]VUF 5000 as a potential PET ligand for brain imaging of the histamine H3 receptor. AB - [18F]VUF 5000 was evaluated as a potential PET ligand for the histamine H3 receptor. In the rat a high uptake of [18F]VUF 5000 was observed in liver, lung and kidney and a low uptake in the brain. In order to explain these findings we determined the LogD(oct,7.2) of [18F]VUF 5000, studied the biodistribution in the presence of carrier VUF 5000, modified [18F]VUF 5000 chemically and studied the binding of [18F]VUF 5000 to human serum albumin. From the results of these experiments it was concluded that [18F]VUF 5000 is not suitable as a PET ligand for brain imaging of the histamine H3 receptor, since [18F]VUF 5000 hardly penetrates into the brain. PMID- 10530923 TI - Stereoelectronic features of the cinchona alkaloids determine their differential antimalarial activity. AB - For most potent antimalarial activity, the cinchona alkaloids appear to require certain electronic features, particularly a sufficiently acidic hydroxyl proton and an electric field direction pointing from the aliphatic nitrogen atom towards the quinoline ring. These observations are the result of an analysis of molecular electronic properties of eight cinchona alkaloids and an in vivo metabolite calculated using ab initio 3-21G quantum chemical methods in relation to their in vitro IC50 values against chloroquine-sensitive and chloroquine-resistant Plasmodium falciparum parasites. The purpose is to provide a profile of the electronic characteristics necessary for potent antimalarial activity for use in the design of new antimalarial agents and to gain insight into the mechanistic path for antimalarial activity. Distinguishing features of the weakly active epiquinine and epiquinidine include a higher dipole moment, a different direction of the electric field, a greater intrinsic nucleophilicity, lower acidity of the hydroxyl proton, a lesser electron affinity of the lowest unoccupied molecular orbitals, and a higher proton affinity than the active cinchona alkaloids. A moderately potent quinine metabolite possesses some, but not all, of the same electronic features as the most potent cinchona alkaloids. Both the positioning of the hydroxyl and aliphatic amine groups and their electronic features appear to play a crucial role for antimalarial potency of the cinchona alkaloids, most likely by controlling the ability of these groups to form effective intermolecular hydrogen bonds. PMID- 10530924 TI - Melanocyte-directed enzyme prodrug therapy (MDEPT): development of a targeted treatment for malignant melanoma. AB - A novel prodrug rationally designed to function as a tyrosinase substrate has been synthesised to allow targeted treatment of malignant melanoma. This agent has been evaluated for tyrosinase-mediated drug release, and has been shown to act in the desired manner. Furthermore, differential cytotoxicity has been demonstrated in cell lines which express tyrosinase and those which do not. PMID- 10530925 TI - Combinatorial discovery process yields antimicrobial peptoids. AB - N-alkylated glycine trimers are generically referred to as peptoids. The identification of antimicrobial peptoids from a statistically unbiased diverse combinatorial chemistry library led to the design of the optimization peptoid library that we describe in this manuscript. This optimization library was designed using structural information from the most active peptoids in the unbiased library. Screening of the optimization library for antimicrobial activity identified a single pool of peptoids with activity against both Staphylococcus aureus and Escherichia coli. The active peptoids from this pool were active against drug sensitive and drug resistant organisms and represent novel antibacterial compounds. PMID- 10530927 TI - Direct solid-phase synthesis of octreotide conjugates: precursors for use as tumor-targeted radiopharmaceuticals. AB - Somatostatin analogues, such as octreotide, are useful for the visualization and treatment of tumors. Unfortunately, these compounds were produced synthetically using complex and inefficient procedures. Here, we describe a novel approach for the synthesis of octreotide and its analogues using p-carboxybenzaldehyde to anchor Fmoc-threoninol to solid phase resins. The reaction of the two hydroxyl groups of Fmoc-threoninol with p-carboxybenzaldehyde was catalyzed with p toluenesulphonic acid in chloroform using a Dean-Stark apparatus to form Fmoc threoninol p-carboxybenzacetal in 91% yield. The Fmoc-threoninol p carboxybenzacetal acted as an Fmoc-amino acid derivative and the carboxyl group of Fmoc-threoninol p-carboxybenzacetal was coupled to an amine-resin via a DCC coupling reaction. The synthesis of protected octreotide and its conjugates were carried out in their entirety using a conventional Fmoc protocol and an autosynthesizer. The acetal was stable during the stepwise elongation of each Fmoc-amino acid as shown by the averaged coupling yield (> 95%). Octreotide (74 to 78% yield) and five conjugated derivatives were synthesized with high yields using this procedure, including three radiotherapy octreotides (62 to 75% yield) and two cellular markers (72 to 76% yield). This novel approach provides a strategy for the rapid and efficient large-scale synthesis of octreotide and its analogues for radiopharmaceutical and tagged conjugates. PMID- 10530926 TI - Heterocyclic analogues of L-citrulline as inhibitors of the isoforms of nitric oxide synthase (NOS) and identification of N(delta)-(4,5-dihydrothiazol-2 yl)ornithine as a potent inhibitor. AB - L-Thiocitrulline is a known potent inhibitor of several isoforms of nitric oxide synthase (NOS). To explore the structure-activity relationships (SARs) for this molecule in more depth than has previously been reported, three analogues substituted at the sulphur of the isothioureas have been synthesised. In two of these, the S-substituent was 'tied back' sterically by cyclisation to the nitrogen remote from the amino-acid unit. N(delta)-(4,5-Dihydrothiazol-2 yl)ornithine was identified as an inhibitor of rat inducible and constitutive isoforms of NOS and of a constitutive NOS derived from a human tumour xenograft. Analogous N(delta)-(thiazol-2-yl)ornithines were less active, whereas the corresponding N(delta)-(oxazol-2-yl)ornithine and N(delta)-(pyrimidin-2 yl)ornithine failed completely to inhibit NOS. A new efficient preparation of the critical synthetic intermediate, N(alpha)-Boc-thiocitrulline t-butyl ester, has been developed. Further exploration of the SAR with 2-amino-5 (heterocyclylthio)pentanoic acids (synthesised from 2-(Boc-amino)-5 bromopentanoic acid t-butyl ester), with N-(4-aminobutyl)thiourea and with 2-(4 aminobutylamino)-4,5-dihydrothiazole enabled refinement of our previous model for binding of the substrate, L-arginine, and the inhibitors to NOS. PMID- 10530928 TI - Molecular modeling of the dopamine D2 and serotonin 5-HT1A receptor binding modes of the enantiomers of 5-OMe-BPAT. AB - Molecular modeling studies were undertaken in order to elucidate the possible dopamine D2 and serotonin 5-HT1A receptor binding modes of the enantiomers of 5 methoxy-2-[N-(2-benzamidoethyl)-N-n-propylamino]tetralin (5-OMe-BPAT, 1). For this purpose, a combination of indirect molecular modeling and direct construction of the seven transmembrane (7TM) domains of the receptors was employed in a stepwise, objective manner. Pharmacophore models and corresponding receptor maps were identified by superimposing selected sets of receptor agonists in their presumed pharmacologically active conformations, while taking the conformational freedom of the ligands into account. The 7TM models were then constructed around the agonist pharmacophore models, by adding the TM domains one by-one. Initially, the relative positions of TM3, TM4, and TM5 were determined using the three-dimensional structure of bacteriorhodopsin, but subsequently the orientations of all TM domains were adjusted in order to mimic the topology of the TM domains of rhodopsin. The presumed dopamine D2 receptor binding conformations of (S)- and (R)-1 were determined by using the semirigid dopamine D2 receptor antagonist N-benzylpiquindone as a template for superposition. Similarly, the selective serotonin 5-HT1A receptor agonist flesinoxan was employed for identifying the serotonin 5-HT1A receptor binding conformations of the enantiomers of 1. After docking of the presumed pharmacologically active conformations in the 7TM models and subsequent optimization of the binding sites, specific interactions between the ligands and the surrounding amino acid residues, consistent with the structure-activity relationships, were observed. Thus, both enantiomers of 1 bound to the dopamine D2 receptor model in a similar fashion: a reinforced electrostatic interaction was present between the protonated nitrogen atoms and Asp114 in TM3; their carbonyl groups accepted a H bond from Ser121 in TM3; their amide NH groups acted as H-bond donor to Tyr416 in TM7; and their benzamide phenyl rings were involved in a hydrophobic edge-to-face interaction with Trp386 in TM6. Differences were observed in the orientations of the 2-aminotetralin moieties, which occupied the agonist binding site. Whereas the (S)-enantiomer could form a H-bond between its 5-methoxy substituent and Ser193 in TM5, the (R)-enantiomer could not, which may account for the differences in their intrinsic efficacies at the dopamine D2 receptor. In the serotonin 5-HT1A receptor model, the benzamide phenyl rings of both enantiomers were involved in hydrophobic face-to-face interactions with Phe112 in TM3, while their protonated nitrogen atoms formed a reinforced electrostatic interaction with Asp116 in TM3. Consistent with the structure-affinity relationships of 1, the amide moieties were not involved in specific interactions. Both enantiomers of 1 could form a hydrogen bond between their 5-methoxy substituent and Thr200 in TM5, which may account for their full serotonin 5-HT1A receptor agonist properties. PMID- 10530930 TI - Synthesis of novel progestin-rhenium conjugates as potential ligands for the progesterone receptor. AB - To assist in the development of technetium-based radiopharmaceuticals that are useful for the diagnostic imaging of steroid receptor-positive breast tumors, we have synthesized a series of small-sized metal chelates according to 'n + 1' mixed-ligand, thioether-carbonyl and organometallic designs. In these preliminary investigations, rhenium was used as a model for the radioactive technetium. The metal chelates contain the rhenium metal in several oxidation states, being + 5, + 3, and + 1, and they were attached to 21-substituted progesterone derivatives. A competitive receptor-binding assay (rat uterine cytosol, 0 degrees C) was used to determine the binding affinity of these conjugates for the progesterone receptor. The highest affinity of 9% (RU5020 = 100%) was obtained with a '3 + 1' mixed-ligand complex, containing a NMe group as the central donor atom in the tridentate ligand part. This value reflects a relative binding affinity of 75% compared with the parent steroid progesterone. PMID- 10530929 TI - Semisynthesis and cytotoxicity of amino acetogenins and derivatives. AB - Semisynthetic derivatives were prepared from two natural annonaceous acetogenins, rolliniastatin-1 and squamocin, and their cytotoxicity was evaluated. Amino derivatives show decreased bioactivity. Isorolliniastatin-1 was found to be much less toxic than rolliniastatin-1 after intraperitoneal administration to mice, although the in vitro cytotoxicity of both compounds was comparable. PMID- 10530931 TI - Synthesis and antagonistic activity at muscarinic receptor subtypes of some 2 carbonyl derivatives of diphenidol. AB - A series of 2-carbonyl analogues of the muscarinic antagonist diphenidol bearing 1-substituents of different lipophilic, electronic, and steric properties was synthesized and their affinity for the M2 and M3 muscarinic receptor subtypes was evaluated by functional tests. Two derivatives (2g and 2d) showed an M2-selective profile which was confirmed by functional tests on the M1 and M4 receptors. A possible relationship between M2 selectivity and lipophilicity of the 1 substituent was suggested by structure-activity analysis. This work showed that appropriate structural modification of diphenidol can lead to M2-selective muscarinic antagonists of possible interest in the field of Alzheimer's disease. PMID- 10530932 TI - Analogues of 1-(3,10-dibromo-8-chloro-6,11-dihydro-5H-benzo-[5,6]-cyclohepta [1,2 b]pyridin-11-yl)piperidine as inhibitors of farnesyl protein transferase. AB - The synthesis of several 4-pyridylacetyl N-oxide derivatives of 4-(3-bromo-6,11 dihydro-5H-benzo[5,6]-cyclohepta[1,2-b]-pyridin-11-yl)pi perazine/piperidine 3 is described. This study was aimed at identifying fomesyl protein transferase (FPT) inhibitors in these two series of tricycles containing different phenyl ring substituents. The in vitro activity profile of the initial group of compounds 7a 7g led to the synthesis of the 8-methyl-10-methoxy and 8-methyl-10-bromo analogues 7i, 13i, and 13j. The 11R(-) enantiomers of these compounds were found to exhibit potent in vitro FPT inhibition activity. PMID- 10530933 TI - Synthesis and investigation of glycosylated mono- and diarylporphyrins for photodynamic therapy. AB - The synthesis of a diaryl substituted porphyrin bearing a galactosyl and a cholesteryloxy substituent and of a galactosyl substituted monoaryl porphyrin is described. The spectroscopic and aggregation properties of both compounds were investigated. The galactosyl substituted monoaryl porphyrin (12) was efficiently incorporated into liposomes and lipoproteins whereas the diaryl porphyrin showed no interaction with these lipids. Furthermore the binding constants of compound 12 to HDL, LDL, VLDL, and PE and DMPC liposomes were estimated. PMID- 10530934 TI - Chemoenzymatic synthesis and antitumor promoting activity of 6'- and 3-esters of 2-O-beta-D-glucosylglycerol. AB - Through a simple chemoenzymatic approach 6'- and 3-esters of 2-O-beta-D glucosylglycerol 1, with short-medium length fatty acid acyl chains, were prepared. The study of their in vitro antitumor promoting effect on Epstein-Barr virus early antigen (EBV-EA) activation, in comparison with that of the 1-esters previously described, confirms the significant activity of such monoacylated glycoglycerolipid analogues and establishes for the glucose series that the 1 substitution and the hexanoyl chain are the proper structural features for the maximum activity. PMID- 10530935 TI - Synthesis and binding properties of photoactivable biotin-conjugated verapamil derivatives for the study of P-170 glycoprotein. AB - The design and synthesis of two photoactivable biotin-labeled analogues of verapamil (6 and 7) is reported. Preliminary evaluation of the biological profile of 6 (EDP 137) and 7 (EDP 141) shows that they have comparable affinities to that of verapamil for P-170, the protein responsible for multidrug resistance (MDR). Since both appear to bind irreversibly to the protein and the presence of biotin in their structure makes them easily detectable by avidin, they promise to be of great help in studying the protein and its mechanism of action. PMID- 10530936 TI - Synthesis of heparin-like antithrombotics having perphosphorylated thrombin binding domains. AB - The synthesis of three heparin analogues (i.e. compounds VI-VIII) having perphosphorylated thrombin binding domains (TBDs) is reported. These compounds were tested in vitro for their antithrombin III (ATIII)-mediated anti-Xa and antithrombin activities. Conjugates VI and VIII show a remarkable increase in antithrombin activity compared to the structurally related conjugates with persulfated TBDs (i.e. compounds IV and V), whereas compound VII displays a diminished activity. PMID- 10530937 TI - Novel cobalt complex inhibitors of mitochondrial calcium uptake. AB - Reperfusion of the ischaemic myocardium leads to intracellular calcium overload followed by mitochondrial dysfunction, resulting in insufficient energy supply and ultimately myocardial necrosis. Ruthenium red (RR), a potent mitochondrial calcium uptake inhibitor, prevents this disruption to mitochondrial metabolism and improves post reperfusion recovery. This therefore suggested that mitochondrial calcium influx is an attractive target for the treatment of reperfusion injury. However, RR is unsuitable for therapeutic use, so we undertook a search for novel compounds which inhibit mitochondrial calcium uptake. The most potent compounds discovered were simple tris(ethylenediamine) transition metal complexes and dinuclear Co complexes. The structure-activity relationship (SAR) of these small molecules has helped to define the structural requirements for inhibition of calcium transport by outlining the size and charge dependency of the interactive site on the mitochondrial calcium uniporter. PMID- 10530938 TI - Biomimetic degradation of lignin and lignin model compounds by synthetic anionic and cationic water soluble manganese and iron porphyrins. AB - The biomimetic oxidation of 5-5' condensed and diphenylmethane lignin model compounds with several water soluble anionic and cationic iron and manganese porphyrins in the presence of hydrogen peroxide is reported. The oxidative efficiency of manganese and iron meso-tetra(2,6-dichloro-3-sulphonatophenyl) porphyrin chloride (TDCSPPMnCl and TDCSPPFeCl, respectively), meso-tetra-3 sulphonatophenyl porphyrin chloride (TSPPMnCl) and manganese meso-tetra(N methylpyridinio)porphyrin pentaacetate (TPyMePMn(CH3COO)5) was compared on the basis of the oxidation extent of the models tested. Manganese porphyrins were found more effective in degrading lignin substructures than iron ones. Among them the cationic TPyMePMn (CH3COO)5, never used before in lignin oxidation, showed to be the best catalyst. The catalytic activity of porphyrins in hydrogen peroxide oxidation of residual kraft lignin was also investigated. The use of quantitative 31P NMR allowed the focusing on the occurrence of different degradative pathways depending on the catalyst used. TPyMePMn(CH3COO)5 was able to perform the most extensive degradation of the lignin structure, as demonstrated by the decrease of aliphatic hydroxyl groups and carboxylic acids. Noteworthy, no significant condensation reactions occurred during manganese porphyrins catalyzed oxidations of residual kraft lignin, while in the presence of iron porphyrins a substantial increase of condensed substructures was detected. PMID- 10530939 TI - Anti-AIDS agents. Part 36: 17-carboxylated steroids as potential anti-HIV agents. AB - In our search for novel anti-HIV agents, seven 17-carboxylated steroid derivatives were synthesized and evaluated as potential anti-HIV agents. Compound 13 exhibited potent anti-HIV activity in acutely infected H9 lymphocytes with EC50 and therapeutic index values of 0.8 microM and 300, respectively. PMID- 10530940 TI - Imidazole substituted biphenyls: a new class of highly potent and in vivo active inhibitors of P450 17 as potential therapeutics for treatment of prostate cancer. AB - 3- And 4-imidazol-1-yl-methyl substituted biphenyl compounds (named as meta- and para-substituted compounds) were synthesized bearing additional substituents in 3'-/4'-position as inhibitors of P450 17 (17alpha-hydroxylase-C17,20-lyase). P450 17 is the key enzyme of androgen biosynthesis. Its inhibition is a novel therapeutic strategy for treatment of prostate cancer (PC). Twenty-nine compounds were synthesized by Ar-Mg-Br, Negishi or Suzuki aryl-aryl cross coupling and tested toward human and rat enzyme. Most of the compounds showed moderate to excellent activity against one of the enzymes (0.087 microM < or = IC50 < or = 7.7 microM (ketoconazole: 0.74 microM) for the human enzyme, 0.63 microM < or = IC50 < or = 32 microM (ketoconazole: 67 microM) for the rat enzyme). Interestingly, strong species differences were observed. In addition compounds were tested for inhibition toward P450 arom. The 3-imidazol-1-yl-methyl substituted compounds showed good inhibitory activity of P450 arom, while for the 4-substituted compounds negligible inhibition was found. For the most active group of P450 17 inhibitors, (i.e. the 4-imidazol-1-yl-methyl substituted compounds) a QSAR study was performed for inhibition of the human enzyme leading to the result that a hydrophilic substituent in 3'-/4'-position is very important. The most promising compounds (with respect to activity toward both enzymes) were tested in vivo using SD-rats for reduction of plasma testosterone concentrations 2 and 6 h after single i.p. application. The fluorine substituted compound 8c decreased the testosterone plasma concentration to castration level (after 2 h; 5 mg/kg) showing a biological half live of about 6 h. PMID- 10530941 TI - Synthesis and biological evaluation of 2-methoxy- and 2-methylthio-6-[2' alkylamino)ethyl]-4(3H)-pyrimidinones with anti-rubella virus activity. AB - The synthesis of a new family of antiviral compounds, 2-methoxy-, and 2 methylthio-6-[(2'-alkylamino)ethyl]-4(3H)-pyrimidinones, has been accomplished. The activity of these agents against positive strand (rubella virus and sindbis virus) and negative strand (vesicular stomatitis virus) RNA viruses is reported. Some of these compounds are efficient and selective inhibitors of rubella virus. PMID- 10530942 TI - Synthesis and antifungal activity of coumarins and angular furanocoumarins. AB - Angelicin, a naturally occurring furanocoumarin, that showed antifungal activity, was considered as a lead structure for a group of synthetic coumarins. Antifungal activities of the synthesized coumarins and angelicin derivatives were reported against Candida albicans, Cryptococcus neoformans, Saccharomyces cerevisiae and Aspergillus niger. Human cell line cytotoxicity of several coumarins was evaluated against KB cells. Angelicin and several potent antifungals showed to be non-toxic in this assay. PMID- 10530943 TI - Imidazole-containing amino acids as selective inhibitors of nitric oxide synthases. AB - Two series of imidazole-containing amino acids (1a-e and 2a-c), all larger homologues and analogues of L-histidine, were prepared. Since imidazole and phenyl substituted imidazoles have been reported to be inhibitors of NOS and the mode of action of these compounds as heme ligands is a potential mechanism of inhibitory action, we designed imidazole-containing amino acids as combined inhibitors at both the amino acid as well as heme binding sites. To study the influence of the distance between the amino acid moiety and the imidazole moiety on inhibitory potency, the number of carbons between these two functional groups was varied from two to six. The structure-activity relationships of this class of inhibitors can be correlated with the distance between the heme and the amino acid binding sites of the enzyme. Two of the compounds (1b and 1d) with three and five methylenes between the imidazole and amino acid functional groups, respectively, were found to be potent and selective inhibitors for nNOS and iNOS over eNOS. When phenyl was substituted on the nitrogen of the imidazole, both the potency and isoform selectivity diminished. PMID- 10530944 TI - The synthesis and antibacterial activity of totarol derivatives. Part 1: modifications of ring-C and pro-drugs. AB - A series of analogues of, and potential pro-drugs derived from, the potent antibacterial diterpene totarol (1) were synthesized in order to elucidate the minimum structural requirements for antibacterial activity and to seek compounds with good bioavailability in vivo. These analogues varied in the structural features of their aromatic rings and the prodrugs were O-glycosylated derivatives. They were tested in vitro against three gram-positive bacteria: beta lactamase-positive and high level gentamycin-resistant Enterococcus faecalis, penicillin-resistant Streptococcus pneumoniae, and methicillin-resistant Staphylococcus aureus (MRSA); and against the gram-negative multi-drug-resistant Klebsiella pneumoniae. None of the analogues was more potent than totarol itself, which is effective against these gram-positive bacteria at MIC values of 7 microM. The results were evaluated in terms of a structure-activity relationship and this showed that a phenolic moiety was essential for potent antibacterial activity. Amongst the pro-drugs, totaryl alpha-D-mannopyranoside (22) proved the most active in vitro (MIC 18 microM). The in vivo antibacterial activities of compounds 1, 22 and totarol beta-lactoside (23) were assessed in a mouse model of infection, but they were found to be ineffective. Compounds 1 and 22 were shown to be cytotoxic towards proliferating human cell cultures, CH 2983, HeLa, and MG 63, but only at concentrations of > 30 microM. PMID- 10530945 TI - Synthesis and deconvolution of the first combinatorial library of glycosidase inhibitors. AB - A combinatorial library of 125 compounds with a structure consisting of 1 azafagomine linked at N-1 via an acetic acid linker to a variable tripeptide was synthesised. The library was synthesised by Merrifield split and mix synthesis of the peptide, followed by capping with chloroacetate, regioselective nucleophilic substitution with 1-azafagomine and cleavage from the polymeric support. The library was screened for inhibition of beta-glucosidase, alpha-glucosidase and glycogen phosphorylase and found to display beta-glucosidase inhibition. Deconvolution of the library revealed that some inhibition was caused by all library members but the strongest inhibitor was clearly a compound having three hydroxyproline residues in the peptide fragment. This compound was a weaker but more selective inhibitor than 1-azafagomine itself. PMID- 10530946 TI - Lipase-catalysed regio- and enantioselective deacetylation of 2,4-diacetoxyphenyl alkyl ketones. AB - Porcine pancreatic lipase in tetrahydrofuran catalyses the deacetylation of 2,4 diacetoxyphenyl alkyl ketones in a highly regioselective fashion. The strategy of regioselective deacetylation of diacetoxyphenyl alkyl ketones has also resulted in the enantiomeric resolution of a racemic diacetoxyphenyl alkyl ketone, i.e. (+/-)-2,4-diacetoxyphenyl (1-ethyl)pentyl ketone, a precursor for the synthesis of an antifungal coumarin, 7-acetoxy-4-(1-ethyl)pentyl-3-phenyl-2H-1-benzopyran-2 one. PMID- 10530947 TI - tRNA(Phe) binds aminoglycoside antibiotics. AB - Aminoglycoside antibiotics have recently been found to bind to a variety of unrelated RNA molecules, including sequences that are important for retroviral replication. We report the binding of neomycin B, kanamycin A, and Neo-Neo (a synthetic neomycin-neomycin dimer) to tRNA(Phe). Using thermal denaturation studies, fluorescence spectroscopy, Pb2+-mediated tRNA(Phe) cleavage, and gel mobility shift assays, we have established that aminoglycosides interact with yeast tRNA(Phe) and are likely to induce a conformational change. Thermal denaturation studies revealed that aminoglycosides have a substantial stabilizing effect on tRNA(Phe) secondary and tertiary structures, much greater than the stabilization effect of spermine, an unstructured polyamine. Aminoglycoside induced inhibition of Pb2+-mediated tRNA(Phe) cleavage yielded IC50 values of: 5 microM for Neo-Neo, 100 microM for neomycin B, > 1 mM for kanamycin A, and > 10 mM for spermine. Enzymatic and chemical footprinting indicate that the anticodon stem as well as the junction of the TpsiC and D loops are preferred aminoglycoside binding sites. PMID- 10530948 TI - Modulating pyridoxamine-mediated transamination through a betabeta alpha motif peptide scaffold. AB - A pyridoxamine coenzyme amino acid chimera (Pam) was incorporated into a designed betabeta alpha motif peptide to explore the ability of a small synthetic peptide scaffold to influence coenzyme mediated transamination. Structural characterization of this peptide by CD and NMR spectroscopy suggested that the pyridoxamine containing residue was accommodated into the sheet region of the motif without gross structural perturbations. To investigate the ability of the peptide architecture to influence the amount and distribution of transamination product in the conversion of pyruvic acid to alanine, a family of 18 related peptides, CBP01-CBP18, was rapidly synthesized and purified in parallel. These peptides were designed to generate different peptide environments for the pyridoxamine functionality within the context of the structured betabeta alpha peptide motif. Studies of peptide-mediated transamination revealed clear trends in stereospecific production of L-alanine as a function of substitutions at positions five and seven of the motif. Furthermore, new trends favoring the other enantiomeric product resulted from the addition of copper(II) ion, a known chelator of the transamination reaction intermediates. In the presence of copper(II) ion the amount of alanine product generated was increased by up to 32 fold relative to a pyridoxamine model compound in the presence of copper(II) ion. These functional results, accompanied by further CD and NMR spectroscopic analysis of CBP14, one of the CBP family of peptides, suggest that small synthetic betabeta alpha motif peptides can be used to influence the functional properties of coenzymes. PMID- 10530949 TI - Oligosaccharides corresponding to the regular sequence of heparin: chemical synthesis and interaction with FGF-2. AB - It has been proposed that oligosaccharides corresponding to the so-called regular region of heparin/heparan sulfate (HS) bind to fibroblast growth factor-2 (FGF 2). In order to explore the molecular basis of FGF/HS interaction, we describe here the chemical synthesis of a tetra and a hexasaccharide, prepared as methyl glycosides, corresponding to the regular sequence of heparin. The strategy relies on the efficient preparation of three building blocks: a seeding block, an elongating block and a capping block. The hexasaccharide inhibited the binding of FGF-2 on its receptor on human aorta vascular smooth muscle cells with an IC50 value (16+/-1.2 microg/mL) close to that of standard heparin (14.8+/-0.5 microg/mL) whereas the tetrasaccharide was much less potent (IC50 = 127+/-10.5 microg/mL). The hexasaccharide and heparin, inhibited in a dose-dependent manner FGF-2 (30 nM) induced proliferation (IC50 = 23.7+/-1.6 and 30.1+/-1.3 microg/mL, respectively). Under the same experimental conditions, the tetrasaccharide only slightly inhibited the mitogenic effect of FGF-2 (IC50 > 100 microg/mL). PMID- 10530950 TI - In vitro cytotoxicity of 5-aminosubstituted 20(S)-camptothecins. Part 1. AB - A number of 5-aminosubstituted 20(S)-camptothecin analogues were prepared via semi-synthesis starting from 20(S)-camptothecin and 9-methoxy 20(S)-camptothecin. In vitro anti-cancer activity of these analogues was determined using 60 human tumor cell line assay. Although water solubility of most of these compounds was improved compared to 20(S)-camptothecin, their anti-cancer activity was considerably diminished. However, only smaller substituents such as methylamine or hydroxylamine as present in 8s and 8t, respectively, showed good activity with improved water solubility. PMID- 10530951 TI - CH/pi interaction in the conformation of peptides. A database study. AB - A study was carried out, with use of the Cambridge Structural Database, to examine the role of the CH/pi interaction in the conformation of peptides. A number of short intramolecular CH/pi distances have been shown in the crystal structure of peptides bearing at least an aromatic residue in the sequence. The molecular structure in the crystal was inspected individually to know whether the conformation is merely a consequence of the so-called packing forces, or the CH/pi interaction plays a role. It has been demonstrated that the CH/pi interaction constitutes one of the key factors in controlling the conformation of peptides. PMID- 10530952 TI - Chemical modification of apomorphine to discover sigma ligands: 6H dibenzo[b,d]pyran and carbazole analogues. AB - It seems that many sigma ligands have been designed from known sigma ligands. We focused on a difference in structural flexibility between haloperidol and apomorphine, and studied chemical modification of apomorphine, a compound with high affinity for dopamine D2 receptors but not for sigma receptors, for discovery of sigma ligands. The first modification yielded good results with 6H dibenzo[b,d]pyran analogues with weak affinity for sigma receptors but not D2 receptors. Furthermore, carbazole analogues, compounds designed from 6H dibenzo[b,d]pyran analogues, potentially acted at sigma receptors with high selectivity. This paper describes the design, synthesis and sigma/D2 selectivity of 6H-dibenzo[b,d]pyran and carbazole analogues. PMID- 10530953 TI - Synthesis and evaluation of homofarnesoyl-substituted CAAX-peptidomimetics as farnesyltransferase inhibitors and antiproliferative agents. AB - Several CAAX-peptidomimetics were linked to homofarnesoic acid via a beta-alanyl spacer with the intention to obtain a novel type of bisubstrate analogue farnesyltransferase inhibitors. However, the compounds were found to be only weakly active in the farnesyltransferase inhibition assay. Nevertheless, they displayed antiproliferative activity against different tumor cell lines in the low micromolar range. Replacement of the beta-alanine moiety by aspartic acid-1 methyl ester resulted in a compound which inhibited the farnesyltransferase with an IC50 of 860 nM. The corresponding free acid showed a eightfold loss in activity (IC50 = 6.9 microM). PMID- 10530954 TI - Lucidenic acid O and lactone, new terpene inhibitors of eukaryotic DNA polymerases from a basidiomycete, Ganoderma lucidum. AB - Terpenoids, 1, 2 and 3, which selectively inhibit eukaryotic DNA polymerase activities, were isolated from the fruiting body of a basidiomycete, Ganoderma lucidum, and their structures were determined by spectroscopic analyses. New terpenes, lucidenic acid O (1) and lucidenic lactone (2), prevented not only the activities of calf DNA polymerase alpha and rat DNA polymerase beta, but also these of human immunodeficiency virus type 1 reverse transcriptase. Cerevisterol (3), which was reported to be a cytotoxic steroid, inhibited only the activity of DNA polymerase alpha. Although these compounds did not influence the activities of prokaryotic DNA polymerases and other DNA metabolic enzymes such as T7 RNA polymerase and deoxyribonuclease I. PMID- 10530955 TI - Synthesis of an artificial glycoconjugate polymer carrying Pk-antigenic trisaccharide and its potent neutralization activity against Shiga-like toxin. AB - Fluorescence-labeled glycoconjugate polymers carrying carbohydrate segments of a globotriaosyl ceramide (Gb3) were synthesized and subjected to biological assays using Escherichia coli O-157 strains and Shiga-like toxins (Stx-I and Stx-II). For the fluorescence labeling, a new polymerizable fluorescent monomer with a TBMB carbonyl chromophore (Ex. 325 nm, Em. 410 nm) was designed. A glycosyl monomer of the trisaccharide segment of Gb3 was prepared from p-nitrophenyl beta lactoside and copolymerized with acrylamide and the fluorescent monomer to prepare a fluorescence-labeled glycoconjugate copolymer carrying [alpha-D galactopyranosyl-(1-->4)-beta-D-galactopyranosyl]-(1-->4)-beta- D glucopyranoside. The polymer showed potent neutralization activity against Stx-I and also binding activity onto E. coli O-157 strains. PMID- 10530956 TI - Structure-activity relationship of HIV-1 protease inhibitors containing alpha hydroxy-beta-amino acids. Detailed study of P1 site. AB - The structure-activity relationship of HIV-1 protease (HIV-1 PR) inhibitors containing alpha-hydroxy-beta-amino acids is discussed. We demonstrated that substituent groups on the P1 aromatic rings of the inhibitors exert significant influence on their biological activity. Inhibitors bearing an alkyl or a fluorine atom at the meta and para position on their P1 benzene ring were found to be good inhibitors. We also discovered that the substitution positions of the P2 benzamides were crucial for good antiviral potency. In this study, inhibitor 48 was the most potent [IC90 (CEM/HIV-1 IIIB) 27 nM] and showed good pharmacokinetics in rats. PMID- 10530957 TI - Mechanism of hemolysis and erythrocyte transformation caused by lipogrammistin-A, a lipophilic and acylated cyclic polyamine from the skin secretion of soapfishes (Grammistidae). AB - The mechanism of hemolysis and erythrocyte transformation caused by lipogrammistin-A (LGA), a lipophilic and acylated cyclic polyamine from the skin secretion of soapfishes (Grammistidae), was investigated. The dependency of hemolysis on the erythrocyte concentration indicated that the amount of membrane bound LGA required for 50% hemolysis is about 13% of the total phospholipids in erythrocytes on a molar basis. A synthetic analogue which lacked a long alkyl chain exhibited much less activity, suggesting that the alkyl chain is important for membrane-binding. In addition, microscopic observations showed that LGA elicited the invagination of erythrocytes at sublytic concentrations, which makes LGA one of the most potent agents with this transforming activity known to date. Its protonated secondary amino group is responsible for the unequal distribution of LGA in the inner leaflet of the lipid bilayer, which leads to invagination, since acetylation at the amino group markedly reduced the invagination activity. Furthermore, the size of LGA-induced lesions on erythrocyte membrane was estimated to be 7-29 A based on osmotic protection experiments, where the external addition of isotonic molecules in this size range gradually increased the effective dose of LGA. Based on these lines of evidence, the mode of LGA action on erythrocytes is deduced to be as follows. First, LGA molecules bind to erythrocyte membrane by lipophilicity. Second, the molecules accumulate in the inner leaflet of the lipid bilayer by interaction of their cationic ammonium groups with acidic residues of membrane lipid in the inner surface. This uneven distribution of LGA distorts the bilayer structure, and results in a change in cell shape and consequent small lesions. Third, small solutes permeate through the lesions, which induces an osmotic change across the membrane, which leads to colloid-osmotic rupture. This mode of action of LGA on erythrocytes accompanied by cell invagination is the first reported example for natural defense substances. PMID- 10530958 TI - Resolution, molecular structure and biological activities of the D- and L enantiomers of potent anti-implantation agent, DL-2-[4-(2 piperidinoethoxy)phenyl]-3-phenyl-2H-1-benzopyran. AB - Compound 1 (DL-2-[4-(2-piperidinoethoxy)phenyl]-3-phenyl-2H-1-benzopyran, CDRI 85/287) a potent anti-estrogen and anti-implantation agent has been successfully resolved into its pure D- and L-enantiomers. Biological studies showed L enantiomer to be the active form, exhibiting a fivefold higher receptor affinity for the rat uterine cytosolic estrogen receptor, 100% contraceptive efficacy at 1.3 mg/kg dose in single day schedule and 89% inhibition of estradiol induced increase of uterine weight at its contraceptive dose. The absolute stereochemistry determined by X-ray crystallographic analysis showed that the L enantiomer has 2R configuration at its asymmetric centre. PMID- 10530959 TI - Mechanism of biochemical action of substituted 4-methylbenzopyran-2-ones. Part 5: Pulse radiolysis studies on the antioxidant action of 7,8-diacetoxy-4 methylcoumarin. AB - 7,8-Dihydroxy-4-methylcoumarin (1, DHMC) and 7,8-diacetoxy-4-methylcoumarin (2, DAMC) were shown to possess radical scavenging property and strongly inhibit membrane lipid peroxidation. Although free polyphenolic compounds are known to be antioxidants, the antioxidant action of the acetoxy compound DAMC was intriguing. Hence, pulse radiolysis studies were undertaken to explain the antioxidant action of DAMC. Accordingly, DAMC and DHMC were separately reacted with the system generating azide radicals and the resulting transient spectra were recorded. The spectra so obtained in both the cases demonstrated peak at 410 nm, characteristic of phenoxyl radical. The rate constants for the formation of phenoxyl radical from DHMC and DAMC were 34 x 10(8) M(-1) s(-1) and 6.2 x 10(8) M(-1) s(-1), respectively. We propose that the free radical mediated oxidation of DAMC initially produces a radical cation that loses an acetyl carbocation to yield the phenoxyl radical. It is possible to conclude that the mechanism of the antioxidant action of DAMC follows the pathway similar to that of DHMC involving the formation of a stable phenoxyl radical. PMID- 10530961 TI - Design and synthesis of a novel series of HIV-1 protease inhibitors. AB - The synthesis and the SAR study of novel pseudo symmetric inhibitors of HIV-1 protease are described. Michael addition of amino acid derivatives to vinyl ketones was utilized to derive a potent (nM) series of HIV-1 protease inhibitors. PMID- 10530960 TI - Synthesis and antitumor activity of goniofufurone analogues. AB - Synthesis and antitumor activity of goniofufurone analogues 15, 16, 17, 33, and 46 is reported. Key step in the synthesis is Pd (II) mediated oxidative cyclisation of vinyl-(hydroxy) furans 18, 19 to the corresponding lactols 32, 43. Cytotoxicities of 15, 16, 17, 33, and 46 tested against six human cancer cell lines are reported. Change of stereochemistry at C-5, C-6 and C-7 position of goniofufurone (1) did not enhance the cytotoxicities significantly. PMID- 10530963 TI - Dolls and shaken baby syndrome. PMID- 10530962 TI - Synthesis and biology of 3'-N-acyl-N-debenzoylpaclitaxel analogues. AB - The, 3'-N-acyl-N-debenzoylpaclitaxel analogues 1a-d were synthesized and evaluated on biological systems. Some of the analogues 1a-d exhibited higher cytotoxicities (up to 20-fold) and stronger abilities to induce apoptosis than paclitaxel. In an in vivo experiment against i.p. implanted B16 melanoma, the most cytotoxic compound 1b in vitro caused tumor growth inhibition more than paclitaxel. PMID- 10530964 TI - Retinopathy of prematurity survey results. PMID- 10530965 TI - Funduscopic lesions associated with mortality in shaken baby syndrome. AB - PURPOSE: The shaken baby syndrome (SBS) has been defined as a syndrome of intraocular and intracranial hemorrhage in young children, thought to be caused by violent shaking inflicted by an adult. In many cases SBS is fatal as a result of intracranial injury. Intraocular findings include hemorrhage, which may be accompanied by characteristic retinal folds or retinoschisis lesions. This study was performed to determine whether acute ophthalmologic findings might predict a fatal outcome. METHODS: A consecutive series of 10 patients meeting a strict definition of SBS was reviewed for ophthalmic findings at presentation and outcome. RESULTS: Seven patients survived, and three died. Of the six funduscopic characteristics identified in these patients, two were significantly associated with a fatal outcome: circular perimacular retinal folds found in four patients (p = 0.048) and peripheral retinoschisis lesions seen in three patients (p = 0.012). Lack of visual response at initial examination was also significantly associated with a fatal outcome (p = 0.033). CONCLUSIONS: Ophthalmic examination of children with suspected SBS is important for prognostic as well as diagnostic purposes. Circular perimacular retinal folds, peripheral retinoschisis lesions, and lack of visual response correlated with fatal neurologic trauma and may be useful in predicting severity of central nervous system injury in shaken baby syndrome. PMID- 10530966 TI - Dissociative phenomena in congenital monocular elevation deficiency. AB - INTRODUCTION: Monocular elevation deficiency is characterized by unilateral limitation of elevation in both adduction and abduction and is usually present at birth. Dissociative phenomena such as dissociated vertical deviation are well recognized in association with conditions such as congenital esotropia but much less so in association with conditions such as congenital monocular elevation deficiency. METHODS: All 129 patients given the diagnosis of monocular elevation deficiency or double elevator palsy in the Pediatric Ophthalmology and Strabismus Clinic at the University of Iowa Hospitals and Clinics between 1971 and 1995 were reviewed. After those with history of trauma, myasthenia gravis, thyroid eye disease, orbital lesions, Brown syndrome, or monocular elevation deficiency with acquired onset were excluded, 31 patients with congenital monocular elevation deficiency remained for retrospective study. RESULTS: First diagnosed at median age 2.6 years (although all were noted by parents at less than 6 months of age) with mean follow-up of 5.0 years (up to 15.5 years), 9 of 31 (29%) developed dissociated vertical deviation in the eye with monocular elevation deficiency, all of whom had undergone strabismus surgery 0 to 9.7 years previously (mean 3.5 years). Those who developed dissociated vertical deviation were generally younger, were followed up longer, and had more accompanying horizontal strabismus than did those who did not develop dissociated vertical deviation. The results did not reach significance. CONCLUSION: The current study demonstrates that dissociated vertical deviation occurs in association with monocular elevation deficiency. PMID- 10530968 TI - Assessment of stereopsis in college baseball pitchers and batters. AB - BACKGROUND/PURPOSE: In baseball pitching is a much less visually demanding task than hitting is. We sought to find a stronger relationship between visual function (as measured by stereopsis) and hitting a baseball than between stereopsis and pitching skill. METHODS: Multiple parameters including near and distance stereopsis (as a measure of visual function) were measured in 23 returning college baseball players (with a prior year's record). Their level of stereopsis (measured at distance with use of the B-VAT II BVS Binocular Vision Testing System) was compared with their batting statistics (in the case of the 14 position players) or their pitching statistics (for the 9 pitchers). Batting average and slugging percentage were used as measures of hitting skill, and earned run average, "out percentage" (1.000--Opposing players' batting average) and "strike-out percentage" (frequency of batters struck out) were used as pitching parameters. RESULTS: As a group, visual parameters were excellent in these college athletes. Although the newer test of distance stereopsis correlated with the standard near stereo test (Spearman coefficients), there was no correlation between distance stereopsis and any of the pitching or hitting performance parameters. CONCLUSIONS: The nature of the visual demands required of successful baseball hitters may have yet to be determined or are difficult to identify among a population homogeneous for excellent visual function, true for most groups of competitive athletes. In addition, simpler parameters such as plain visual acuity may be the most important factor(s). PMID- 10530967 TI - Levodopa-carbidopa and childhood retinal disease. AB - PURPOSE: Our purpose was to determine the influence of levodopa-carbidopa on visual function in children with retinal disease. METHOD: Two studies were undertaken, a single-dose study and a longitudinal dosing study. A double-masked, placebo controlled single-dose study was undertaken of levodopa-carbidopa (2.08 mg/kg of body weight levodopa with 25% carbidopa) on monocular visual acuity in 14 children with retinal disease. Subjects received two capsules approximately 2.5 hours apart, and monocular visual acuity was measured 2 hours after each capsule ingestion. The second study was a double-masked, placebo-controlled 12 week longitudinal dosing (0.62 mg/kg of body weight) crossover study in which subjects received levodopa-carbidopa for 6 weeks and placebo for 6 weeks. RESULTS: The single-dose study revealed a small but statistically significant improvement in monocular visual acuity after levodopa-carbidopa ingestion. The longitudinal study revealed a small but statistically significant improvement in binocular visual acuity after levodopa ingestion. In both studies placebo had no significant effect on visual acuity. Six subjects participated in both studies and demonstrated a significant correlation (r = 0.76, p < 0.05) between change in visual acuity in the single-dose study and the longitudinal dosing study. CONCLUSION: The results are consistent with the hypothesis that dopamine influences the receptive field characteristics of retinal cells. The results also suggest that there may be low retinal dopamine levels in some types of retinal disease, which may be amenable to treatment. PMID- 10530969 TI - The use of "diagnostic V codes" in pediatric ophthalmology. AB - BACKGROUND: Children requiring comprehensive eye examination for signs of familial eye disease or suspected systemic condition are referred to pediatric ophthalmologists. Such examinations may be critical to diagnosis and patient management, yet medically necessary screening examinations are not reimbursed by insurers when the patient is "normal." Faced with this dilemma regularly in a children's hospital practice, we began tracking insurer acceptance of "diagnostic V codes," which are diagnostic codes generally related to symptoms or status conditions rather than to medical states. The authors felt that "diagnostic V codes" might represent a more appropriate and correct code in the circumstance of a medically necessary screening examination found to be normal. METHODS: In July 1995 the use of 11 primary diagnostic V codes and 7 secondary diagnostic V codes was discussed in our faculty meeting and a list was displayed in each examination lane. Acceptance of V codes was then tracked for an 8-month period and analyzed 1 year later to provide a set of closed accounts. RESULTS: The code V71.8, or "observation for other specified, suspected condition" had a 73% acceptance rate by insurers on the basis of 207 examinations. The acceptance rate was not dependent on the use of a secondary code (76% for V71.8 alone vs 71% for secondary code used). Other V codes were infrequently used. CONCLUSIONS: The code V71.8 was an accepted code by insurers in our locality for medically necessary screening examinations found to be normal. The authors feel this high acceptance rate by insurers confirms our impression that V codes are the appropriate and correct codes to use for the medically necessary screening examination found to be normal. PMID- 10530970 TI - Motion perimetry in anisometropic amblyopia: elevated size thresholds extend into the midperiphery. AB - PURPOSE: Our purpose was to determine whether motion detection abnormalities in patients with anisometropic amblyopia exist and to determine the extent of these abnormalities in the central and midperipheral visual field. METHODS: We used of motion perimetry to evaluated 10 anisometropic subjects with no manifest strabismus. Each of 44 locations in the visual field corresponding to the test sites of the Humphrey 24-2 program was tested with circular patches of motion (random dot cinematograms) displayed on a computer screen. Stimulus patch size was reduced in a 2/1 staircase manner to determine the smallest patch of motion detectable at each test location (threshold). Data from 15 age-matched normal subjects were used as controls. RESULTS: Vision in the amblyopic eye ranged from 20/25 to count fingers. The overall mean size threshold for amblyopic eyes was elevated (61% +/- 73%) compared with fellow eyes and age-matched normal eyes (p < 0.03) (i.e., the moving patch of dots in the field had to be larger for it to be detected when viewing with the amblyopic eye). The increase in size threshold was consistent across the visual field and was not greater for central locations. CONCLUSION: The amblyopia caused by anisometropia is associated with an abnormality in motion detection that extends into the midperiphery of the visual field. PMID- 10530971 TI - Clinical, cytogenetic, and molecular testing of Argentine patients with retinoblastoma. AB - PURPOSE: The purpose of this study is to determine the clinical, chromosomal, and molecular characteristics of Argentine patients with unilateral and bilateral retinoblastoma. STUDY DESIGN: Eighty-six patients belonging to 82 families were studied; 59% of them were examined during the first year of life. Leukocoria was the most common reason for consultation. Other presenting signs were strabismus and glaucoma. Enucleation of the affected eye was performed in 85% of the cases and the complication rate was 13%. RESULTS: An appropriate therapy allowed the survival of 84 of the 86 patients. Two children with malformations and growth retardation had an abnormal karyotype with a deletion in 13q14. Segregation analysis of polymorphic sites within the retinoblastoma gene and the parental origin of the allele lost in the tumor were analyzed in 30 of the 82 families. Five mutant alleles transmitted through the germline and six de novo germline mutant alleles were identified in 12 patients with hereditary retinoblastoma. Most de novo germline mutant alleles were paternally derived. Molecular analysis of nonhereditary retinoblastoma showed loss of heterozygosity in three of eight cases. From these, two maternal alleles and one paternal allele were lost, thus not indicating a significant difference in the parental origin for the lost allele. CONCLUSIONS: These data are useful for deoxyribonucleic acid diagnosis of susceptibility to retinoblastoma in relatives of hereditary patients, even if mutations have not been identified. PMID- 10530972 TI - The efficacy of SimulVue and Unilens RGP aspheric bifocal contact lenses in the treatment of esotropia associated with a high accommodative convergence/accommodation ratio. AB - PURPOSE: This was a prospective study assessing the efficacy of the SimulVue bifocal contact lens and the Unilens RGP aspheric multifocal contact lens (Unilens, Largo, Fla.) in the treatment of high accommodative convergence/accommodation (AC/A) esotropia in an adolescent and postadolescent population. METHODS: Those patients meeting the inclusion criteria were fit with contact lenses with use of full cycloplegic refraction and later retested by an examiner masked to the previous binocular status. Particular attention was given to the sensory status and the motor fusion of each patient in their bifocal spectacles and then in their bifocal contact lenses. All patients were followed up for at least 6 months after the contact lenses were fitted. RESULTS: Five of the six patients demonstrated larger angles of esophoria or tropia at near with the contact lenses than with bifocal spectacles. The only patient who maintained excellent stereopsis in bifocal contact lenses was the one who normalized her AC/A ratio during this study and no longer required a bifocal in her spectacle correction for fusion. The two patients who initially had no stereopsis but good alignment in spectacle correction had a large esotropia at near fixation with bifocal contact lenses. CONCLUSIONS: The SimulVue and Unilens RGP aspheric bifocal contact lenses did not adequately treat adolescent patients who had esotropia associated with a high AC/A ratio. PMID- 10530973 TI - The anesthetic management of the pediatric strabismus patient. AB - Postoperative nausea and vomiting continues to be a common perioperative complication for pediatric strabismus patients. Postoperative pain management and the choice of general anesthetic can increase the incidence of perioperative nausea. Current techniques for induction of general anesthesia and selection of agents, prevention and treatment of postoperative pain, and options for antiemetic therapy will be reviewed. PMID- 10530974 TI - Bilateral eyelid ecchymosis and subconjunctival hemorrhage associated with coughing paroxysms in pertussis infection. PMID- 10530975 TI - Pseudotumor of the orbit in early childhood. AB - Orbital pseudotumor, also known as idiopathic orbital inflammation, is defined as a nonspecific, nonneoplastic inflammatory process of the orbit without identifiable local or systemic causes. The disorder, first described by Birch Hirschfield in 1905, is more prevalent in the adult population than in the pediatric population. In our study we discuss two cases of pseudotumor of the orbit in children less than 18 months old. This report will highlight the evaluation and management of pediatric orbital pseudotumor and the importance of its inclusion in the differential diagnosis of orbital disorders in young children. PMID- 10530976 TI - Histologic study of a torn inferior oblique muscle. PMID- 10530977 TI - Vertical strabismus resulting from an anomalous extraocular muscle. AB - Unusual ocular motility abnormalities have been rarely reported to result from anomalous extraocular structures. These structures, which may be either muscular or fibrotic, attach to the globe and produce a mechanical restriction resulting in incomitant strabismus. To our knowledge, we report the first patient with an anomalous extraocular muscle in whom the clinical, radiologic, surgical, and histopathologic findings are described. PMID- 10530978 TI - Apparent contralateral inferior oblique muscle overaction after unilateral inferior oblique muscle weakening procedures. AB - BACKGROUND: Unilateral inferior oblique muscle weakening surgical procedures often lead to the appearance of inferior oblique muscle overaction in the contralateral eye. The purpose of this study was to determine how different types of unilateral inferior oblique muscle procedures affect the apparent function of the inferior oblique muscle in the contralateral eye. METHODS: A computer search was performed to locate all patients on the pediatric ophthalmology service at the Wilmer Ophthalmological Institute who underwent a unilateral inferior oblique muscle weakening procedure from 1980 to 1994. Only patients with a diagnosis of primary inferior oblique muscle overaction were included in the study. RESULTS: Fourteen patients met the inclusion criteria. One patient had undergone an anterior transposition of the inferior oblique muscle, seven patients had undergone a 10 mm recession of the inferior oblique muscle, and six patients had undergone a myectomy of the inferior oblique muscle. Before the operation,there was no difference in the inferior oblique muscle function of the contralateral eye among the three groups. However, after the operation apparent inferior oblique muscle overaction developed more frequently and to a greater degree in the contralateral eye among patients in the anterior transposition and 10 mm recession groups than among patients in the myectomy group. CONCLUSION: Either anterior transposition or 10 mm recession of the inferior oblique muscle may limit elevation in abduction in the eye on which inferior oblique muscle surgery was performed. The limitation of elevation in abduction may create apparent inferior oblique muscle overaction in the contralateral eye. PMID- 10530979 TI - Grave complications after superior oblique tenotomy or tenectomy for Brown syndrome. AB - INTRODUCTION: Superior oblique tenotomy or tenectomy was the preferred procedure for Brown syndrome for decades. Superior oblique palsy was reported as a complication, but the complex nature of this palsy and the difficulty in treating it was not emphasized. We documented cases with distressing symptoms of unilateral and bilateral superior oblique palsy after a free tenotomy or tenectomy of the superior oblique tendon for Brown syndrome. METHODS: Four cases referred for management of complex strabismus after superior oblique surgery for Brown syndrome were identified. Case histories, complications, and corresponding management are described in detail. RESULTS: All cases presented with bothersome symptomatic superior oblique palsy-incomitant vertical deviation with significant torsion, diplopia worse in functional down gaze, and anomalous head postures. Although reanastomoses of the superior oblique tendon was attempted in all cases,the procedure was modestly successful in only one case. Superior oblique palsy could not be reversed. After three procedures, Case 1 was orthotropic in primary, right, and left gaze but had a small intermittent tropia in down gaze. Case 2 underwent bilateral superior oblique operations. Despite a unilateral reanastomosed superior oblique tendon, hypertropia in down and left gaze and 15 degrees chin down position persisted. Case 3 showed correction of head tilt and a negative cover test 1 week after the second operation for iatrogenic superior oblique palsy, but stability of the procedure could not be ascertained. Case 4 had a successful attempt at reanastomosis and regained control of the vertical deviation as a hyperphoria. Both anomalous head posture and torsion improved. CONCLUSION: Superior oblique surgery for Brown syndrome may cause irreversible serious strabismus problems. Patients are left with distressing cyclovertical deviation worse in the functional position of gaze with significant torsional component that was resistant to therapy. Although adaptive mechanisms, such as an anomalous head posture, may develop, patients are left with a permanent disability that could not be reversed. One should scrupulously adhere to the indications for surgery in Brown syndrome. Preoperative assessment of superior oblique function is stressed. An alternative surgical procedure that is potentially reversible should be considered. This should include a reliable method to recover both ends of the tenotomized superior oblique tendon in case the procedure needs to be modified at a later date. PMID- 10530980 TI - The prevalence of strabismus in congenital nystagmus: the influence of anterior visual pathway disease. AB - PURPOSE: To determine the influence of underlying visual system disorders on the risk of developing strabismus in children with congenital nystagmus. METHODS: We retrospectively reviewed 82 cases of congenital nystagmus from a pediatric ophthalmology referral practice. RESULTS: Strabismus was found in 50% of children with congenital nystagmus. The prevalence of strabismus was 82% in children with bilateral optic nerve hypoplasia, 53% in patients with albinism, 36% in children with congenital retinal dystrophies, and 17% in children with idiopathic congenital nystagmus. CONCLUSION: The risk that a child with congenital nystagmus will have strabismus develop can be predicted from the nature of the underlying visual disorder. PMID- 10530981 TI - Vertical muscle transposition augmented with lateral fixation. AB - INTRODUCTION: Full vertical rectus muscle transpositions have been shown to be an effective treatment for lateral rectus palsies and type I Duane syndrome. This operation is usually accompanied by mechanical or botulinum toxin treatment of one or both medial rectus muscles. This series evaluates the effect of augmenting the transposed muscles with lateral fixation sutures. METHODS: Transposition of the vertical rectus muscles to the lateral rectus muscle was performed in 23 eyes of 21 patients; transposition to the medial rectus muscle was performed in one eye of one of these 21 patients. A lateral fixation suture of 5-0 Dacron polyester filament was placed in the sclera 16 mm posterior to the limbus and adjacent to the lateral rectus muscle, incorporating one fourth of the transposed vertical rectus muscle. Of the 21 patients, five had type I Duane syndrome with a face turn and esotropia in the primary position, seven had a unilateral lateral rectus palsy, two had bilateral lateral rectus palsy, four had an ipsilateral lateral rectus palsy combined with a contralateral lateral rectus paresis (a recess resect procedure was performed on the paretic eye along with the augmented transposition on the paralyzed eye), two had gaze palsies, and one had a unilateral lateral rectus palsy with recurrent esotropia after a transposition procedure performed 16 years previously. Lateral fixation sutures alone were used in the last case listed. Postoperative diplopia-free fields were measured when possible (10 cases). RESULTS: In most cases (19/23 eyes), alignment was achieved in the primary position with the use of the augmented transposition procedure alone. On average,20 degrees of binocular fusion into the abducted field was obtained. No postoperative limitation of adduction in the transposed eye was noted. Among the patients with Duane syndrome, 80% had elimination of the face turn; one patient had 5 degrees of residual face turn. The one patient with previous transposition surgery alone had an 80% (16 PD) reduction of the recurrent esotropia after placement of lateral fixation sutures. After augmented transpositions, induced vertical deviations in the primary position were uncommon (4/20 patients) and not greater than 2 PD. Significant lid fissure changes were not seen. CONCLUSIONS: The addition of lateral fixation sutures to full vertical rectus muscle transpositions improves the tonic abducting force of the procedure for patients with lateral rectus palsy and type I Duane syndrome without compromising adduction. PMID- 10530982 TI - Recession of both horizontal rectus muscles in Duane syndrome with globe retraction in primary position. AB - INTRODUCTION: Duane syndrome is characterized by abduction deficiency, narrowing of the palpebral fissure on adduction, and globe retraction,which can be the most prominent aspect of the motility disorder. Recession of both horizontal rectus muscles was investigated for treatment of severe globe retraction. METHODS: Three patients with Duane syndrome were operated on for severe globe retraction. The medial rectus muscles were recessed from between 5.5 to 6.5 mm and the lateral rectus muscles 7.0 to 9.0 mm simultaneously. The recessions were asymmetric, as evidenced by amount of esotropia and face turn. Preoperative Hertel measurements were made in primary gaze, 30-degree left gaze, and 30-degree right gaze. The measurements were repeated at 6 months and 1 year after the operation. RESULTS: All three patients had improvement in globe retraction. The 6-month Hertel readings in primary position were improved by a mean of 3.0 mm (range 2.5 to 3.5 mm) measured in primary gaze. Hertel measurements were stable at 1 year after the operation. No complications were encountered. CONCLUSIONS: Recession of both horizontal rectus muscles is effective in the treatment of significant globe retraction in Duane syndrome. PMID- 10530983 TI - Ultrasonographically guided injection of corticosteroids for the treatment of retroseptal capillary hemangiomas in infants. AB - PURPOSE: Injection of corticosteroids is a well-documented and successful mode of treatment for periorbital capillary hemangiomas. Because of the greater potential risk involved with retrobulbar injections, no prior study has described this treatment for tumors located behind the orbital septum. Although retroseptal intraorbital capillary hemangiomas comprise only 7% of all adnexal capillary hemangiomas, complications such as optic nerve compression or astigmatism may necessitate treatment. METHODS: Three patients with deep orbital hemangiomas that caused vision-threatening complications were treated with intralesional injections of triamcinolone and betamethasone. Orbital injection was performed with use of real-time ultrasonographic guidance of the needle. This technique was valuable in providing continuous, accurate, and safe advancement of the needletip in the orbit to avoid the globe and orbital walls. Ultrasonography also permitted precise placement of the needle tip within the tumor and visualization of the injected material. RESULTS: Significant improvement was demonstrated in all cases on the basis of both ultrasonographic measurements and regression of clinical manifestations such as astigmatism, chemosis, proptosis, and optic nerve pallor. No complications were noted. CONCLUSION: Intralesional injection of corticosteroids to treat retroseptal and retrobulbar capillary hemangiomas was found to be a safe and effective treatment modality in our patients. Positioning of the injecting needle was guided by ultrasonography. PMID- 10530984 TI - Baerveldt implant surgery in the treatment of advanced childhood glaucoma. AB - BACKGROUND: The efficacy of Baerveldt implant (Pharmacia & Upjohn, Inc., Kalamazoo, Mich.) surgery in the treatment of advanced childhood glaucoma is unknown. METHODS: We reviewed the results of 23 consecutive 350 mm Baerveldt implants in 20 eyes of 17 children. Results were classified as follows: (1) success; no further reoperation, no decrease in vision, and intraocular pressure at last follow-up less than 21 mm Hg with no medications; (2) qualified success; medication necessary to bring intraocular pressure to less than 21 mm Hg or complication not associated with tube failure; and (3) failure; intraocular pressure >20 mm Hg, tube failure complication or reoperation causing tube removal, phthisis, or enucleation. RESULTS: Original glaucoma types were bilateral aphakic (five), unilateral aphakic or persistent hyperplastic primary vitreous (four), primary infantile (four), juvenile (three), secondary(two), Peter syndrome (one), and Lowe syndrome (one). Patients had undergone a mean of 2.8 previous intraocular procedures. Mean preoperative intraocular pressure was 33.6 mm Hg; average number of preoperative glaucoma medications was 3.0. Mean follow-up was 19 months (range, 6 to 32 months). Eight procedures were considered successful (mean intraocular pressure 15.5 mm Hg), six were qualified successes (mean number of medications 0.8; mean intraocular pressure 16 mm Hg), and nine failed. Two eyes in the qualified success group do not have useful vision as a result of complications. Complications included retinal detachment (five), corneal decompensation (five), corneal graft rejection in five of six grafts; dislocated tubes (three), and recurrent uveitis (two). One of these eyes is phthisic and one has been enucleated. Only two of nine procedures in eyes with a history of one or no previous intraocular operations failed,whereas seven of 13 procedures in eyes with a history of three or more previous procedures failed. Only seven of 13 procedures in aphakic eyes were successes or qualified successes, whereas seven of 10 procedures in phakic eyes had good results. CONCLUSION: Baerveldt implants can produce good short-term results, especially in phakic eyes. Aphakic eyes and eyes that have undergone multiple procedures are at a much higher risk for devastating complications. PMID- 10530985 TI - Outcome-based management of retinopathy of prematurity. Multicenter Trial of Cryotherapy for Retinopathy of prematurity Cooperative Group. AB - PURPOSE: A system is presented for sequentially computing the risk of progression of retinopathy of prematurity (ROP) for infants born weighing not more than 1250 gm. A personal computer program is used to monitor infants' risk of threshold ROP from first appearance of ROP, and the progression in severity is tracked with multiple logistic risk models developed from data in the Multicenter Trial of Cryotherapy for Retinopathy of Prematurity. METHODS: After entry of the infant's birth weight, gestational age, ethnicity, birth in the current hospital or elsewhere, single or multiple birth, and maturity zone of retinal vessels, risk of progression to threshold severity is calculated. New estimates of risk are computed at onset of ROP and prethreshold ROP (any zone I ROP, zone II stage 2+ or 3) according to the extent of retinal vascularization when ROP first appears, how rapidly ROP progresses, and how severe it is. When threshold ROP (8 total or 5 contiguous clock hours of stage 3+ in zone I or II) is reached,the system provides separate estimates of risk that the eye will have an unfavorable 3-month outcome if treated or not. RESULTS: Estimates of risk of progression to threshold disease among the 4099 patients in the Multicenter Trial of Cryotherapy for Retinopathy of Prematurity natural history study varied from less than 1% to more than 70%. For eyes with threshold disease, the risk of an unfavorable outcome at 3 months without treatment varied from less than 10% to more than 90%. CONCLUSION: This method of tracking identifies infants at high risk for severe ROP and poor structural outcome. It provides information about prognosis with a specificity heretofore impossible. PMID- 10530986 TI - Restriction of elevation in abduction after inferior oblique anteriorization. AB - PURPOSE: Inferior oblique anteriorization is gaining popularity for the treatment of dissociated vertical divergence associated with inferior oblique overaction. This procedure is based on the theory that moving the insertion of the inferior oblique muscle anterior to the equator changes its vector of force from one of elevation to one that opposes elevation. The purpose of this investigation is to describe, investigate the cause, and outline treatment for a complication I observed after inferior oblique anteriorization. This postoperative syndrome consists of a motility pattern that resembles marked residual inferior oblique overaction associated with a Y or V pattern. It is probably caused by a restriction of elevation of the abducting eye causing fixation duress, with a resultant upshoot of the contralateral adducting eye. METHODS: A retrospective chart review was conducted for all patients on whom I performed bilateral inferior oblique anteriorization for inferior oblique overaction associated with dissociated vertical divergence. Patients in whom this postoperative syndrome developed were compared with those in whom it did not with respect to type and extent of surgery. In addition, cases of patients I treated or examined for this complication but whose inferior oblique anteriorization had been performed by other ophthalmologists were also analyzed. RESULTS: I performed bilateral inferior oblique anteriorization in 77 patients. In 29 patients the inferior oblique muscles were placed level with the insertions of the inferior rectus muscles, in 31 patients they were placed 1 mm anterior to the insertions of the inferior rectus muscles, and in 17 patients they were placed 2 mm anterior. The postoperative syndrome described here developed in two of the 77 patients; both had the inferior oblique muscles placed 2 mm anterior to the insertions of the inferior rectus muscle. These were also the only two patients in this series in whom the new insertion of the inferior oblique muscle was spread out laterally at the time of anteriorization. I have seen an additional six patients in whom this syndrome developed after undergoing operations by other ophthalmologists. In four, the inferior oblique muscles were placed 2 mm anterior to the insertions of the inferior rectus muscles, and in two they were placed 3 mm anterior. Of the eight patients I have observed with this complication, I reoperated on six. The surgical procedure consisted of denervation or extirpation of both inferior oblique muscles in four patients and conversion to standard recessions of the inferior oblique muscles in two patients. In all six patients,the versions were markedly improved and the Y orV pattern was eliminated after reoperation. CONCLUSIONS: Anteriorization of the inferior oblique muscles more than 1 mm anterior to the insertions of the inferior rectus muscle may cause a limitation of elevation in abduction, resulting in a Y or V pattern that mimics inferior oblique overaction. This may be more likely to occur if the new insertions of the inferior oblique muscles are spread out laterally at the time of anteriorization. PMID- 10530987 TI - Unilateral retraction of eyelid on vomiting. PMID- 10530988 TI - Does a sex bias really exist in the management of women with coronary heart disease? PMID- 10530989 TI - Authors' reply PMID- 10530990 TI - Metalloproteinase inhibitors in snakebite envenomations. PMID- 10530992 TI - Rapid update PMID- 10530991 TI - New hepatitis virus discovered. PMID- 10530993 TI - Unilateral arthritis: contralateral effects. PMID- 10530994 TI - A glimpse of the machinery. AB - Asymmetric mRNA localization within cells plays an important part in both development and physiology. Recent studies have provided a glimpse of the conserved molecular machinery that directs the localization of specific mRNAs. PMID- 10530995 TI - Putting the parts together. AB - The structures of several different domains and subunits of potassium channels have recently been solved. Reassembling these fragments into a working model of an intact voltage-gated channel will be a major challenge. PMID- 10530996 TI - SWItched-on mobility. AB - Recent studies have shown that two nucleosome-remodeling complexes, NURF and CHRAC, open chromatin for transcription and replication by using their common catalytic subunit, the nucleosomal ATPase ISWI, to increase the mobility of nucleosomes relative to DNA sequence. PMID- 10530997 TI - A new deadly Syn? AB - Insulin-mediated regulation of the exocytosis of vesicles containing the glucose transporter GLUT4 has similarities to regulated synaptic transmission. A recent study has now identified a key regulated component of the fusion step in the exocytosis of these GLUT4-containing vesicles. PMID- 10530998 TI - How are moving images segmented? AB - Visual images are segmented perceptually by a variety of cues, including color and motion. Recent experiments, using perceptual and neurophysiological approaches, have explored the complex interaction between these attributes. A full account will certainly include the effects of directed attention. PMID- 10530999 TI - Cashing in on crystals. AB - Structures of the ribosome and its two subunits have been determined by X-ray crystallography at resolutions sufficient to reveal interactions between RNA and protein in the subunits and orientations of substrates at the subunit interface in the intact ribosome. PMID- 10531000 TI - Who chaperones nascent chains in bacteria? AB - The physiological roles of the molecular chaperones trigger factor and DnaK in de novo protein folding have been unclear, but two new studies have shown that they perform essential, yet partially redundant, functions in chaperoning nascent protein chains in bacteria. PMID- 10531001 TI - Thermotoga heats up lateral gene transfer. AB - The complete sequence of the bacterium Thermotoga maritima genome has revealed a large fraction of genes most closely related to those of archaeal species. This adds to the accumulating evidence that lateral gene transfer is a potent evolutionary force in prokaryotes, though questions of its magnitude remain. PMID- 10531002 TI - The yeast CDC9 gene encodes both a nuclear and a mitochondrial form of DNA ligase I. AB - BACKGROUND: The yeast CDC9 gene encodes a DNA ligase I activity required during nuclear DNA replication to ligate the Okazaki fragments formed when the lagging DNA strand is synthesised. The only other DNA ligase predicted from the yeast genome sequence, DNL4/LIG4, is specifically involved in a non-homologous DNA end joining reaction. What then is the source of the DNA ligase activity required for replication of the yeast mitochondrial genome? RESULTS: We report that CDC9 encodes two distinct polypeptides expressed from consecutive in-frame AUG codons. Translational initiation at these two sites gives rise to polypeptides differing by a 23 residue amino-terminal extension, which corresponds to a functional mitochondrial pre-sequence sufficient to direct import into yeast mitochondria. Initiation at the first AUG codon results in a 755 amino-acid polypeptide that is imported into mitochondria, whereupon the pre-sequence is proteolytically removed to yield the mature mitochondrial form of Cdc9p. Initiation at the second AUG codon produces a 732 amino-acid polypeptide, which is localised to the nucleus. Cells expressing only the nuclear isoform were found to be specifically defective in the maintenance of the mitochondrial genome. CONCLUSIONS: CDC9 encodes two distinct forms of DNA ligase I. The first is targeted to the mitochondrion and is required for propagation and maintenance of mitochondrial DNA, the second localises to the nucleus and is sufficient for the essential cell-division function associated with this gene. PMID- 10531003 TI - Newly assembled snRNPs associate with coiled bodies before speckles, suggesting a nuclear snRNP maturation pathway. AB - BACKGROUND: Small nuclear ribonucleoproteins (snRNPs), which are essential components of the mRNA splicing machinery, comprise small nuclear RNAs, each complexed with a set of proteins. An early event in the maturation of snRNPs is the binding of the core proteins - the Sm proteins - to snRNAs in the cytoplasm followed by nuclear import. Immunolabelling with antibodies against Sm proteins shows that splicing snRNPs have a complex steady-state localisation within the nucleus, the result of the association of snRNPs with several distinct subnuclear structures. These include speckles, coiled bodies and nucleoli, in addition to a diffuse nucleoplasmic compartment. The reasons for snRNP accumulation in these different structures are unclear. RESULTS: When mammalian cells were microinjected with plasmids encoding the Sm proteins B, D1 and E, each tagged with either the green fluorescent protein (GFP) or yellow-shifted GFP (YFP), a pulse of expression of the tagged proteins was observed. In each case, the newly synthesised GFP/YFP-labelled snRNPs accumulated first in coiled bodies and nucleoli, and later in nuclear speckles. Mature snRNPs localised immediately to speckles upon entering the nucleus after cell division. CONCLUSIONS: The complex nuclear localisation of splicing snRNPs results, at least in part, from a specific pathway for newly assembled snRNPs. The data demonstrate that the distribution of snRNPs between coiled bodies and speckles is directed and not random. PMID- 10531004 TI - Role of actin-filament disassembly in lamellipodium protrusion in motile cells revealed using the drug jasplakinolide. AB - BACKGROUND: In motile cells, protrusion of the lamellipodium (a type of cell margin) requires assembly of actin monomers into actin filaments at the tip of the lamellipodium. The importance of actin-filament disassembly in this process is less well understood, and is assessed here using the actin drug jasplakinolide, which has two known activities - inhibition of filament disassembly and induction of an increase in actin polymer. RESULTS: In cells the two activities of jasplakinolide were found to be separable; 1 microM jasplakinolide could permeate cells, bind cellular filamentous actin (F-actin) and inhibit filament disassembly within 3.5 minutes, but significant increase in actin polymer was not detected until 60 minutes of treatment. In live, permeabilised cells, jasplakinolide did not inhibit filament assembly from supplied, purified actin monomers. In migrating chick fibroblasts, lamellipodium protrusion was blocked within 1-5 minutes of treatment with 1 microM jasplakinolide, without any perturbation of actin organisation. In non-migrating chick fibroblasts, there was a delay in the onset of jasplakinolide-induced inhibition of lamellipodium protrusion, during which lamellipodium length increased linearly with no increase in protrusion rate. Motility of the bacterium Listeria in infected PtK2 cells was reduced 2.3-fold within 3 minutes of treatment with 1 microM jasplakinolide. CONCLUSIONS: Actin-filament disassembly is tightly coupled to lamellipodium protrusion in migrating chick fibroblasts and motility of Listeria in PtK2 cells. One simple interpretation of these data is a situation whereby ongoing actin-filament assembly uses free actin monomer derived from filament disassembly, in preference to stored monomer. PMID- 10531005 TI - Cyclin D1 production in cycling cells depends on ras in a cell-cycle-specific manner. AB - BACKGROUND: Cellular Ras and cyclin D1 are required at similar times of the cell cycle in quiescent NIH3T3 cells that have been induced to proliferate, but not in the case of cycling NIH3T3 cells. In asynchronous cultures, Ras activity has been found to be required only during G2 phase to promote passage through the entire upcoming cell cycle, whereas cyclin D1 is required through G1 phase until DNA synthesis begins. To explain these results in molecular terms, we propose a model whereby continuous cell cycle progression in NIH3T3 cells requires cellular Ras activity to promote the synthesis of cyclin D1 during G2 phase. Cyclin D1 expression then continues through G1 phase independently of Ras activity, and drives the G1-S phase transition. RESULTS: We found high levels of cyclin D1 expression during the G2, M and G1 phases of the cell cycle in cycling NIH3T3 cells, using quantitative fluorescent antibody measurements of individual cells. By microinjecting anti-Ras antibody, we found that the induction of cyclin D1 expression beginning in G2 phase was dependent on Ras activity. Consistent with our model, cyclin D1 expression during G1 phase was particularly stable following neutralization of cellular Ras. Finally, ectopic expression of cyclin D1 largely overcame the requirement for cellular Ras activity during the continuous proliferation of cycling NIH3T3 cells. CONCLUSIONS: Ras-dependent induction of cyclin D1 expression beginning in G2 phase is critical for continuous cell cycle progression in NIH3T3 cells. PMID- 10531006 TI - A retention mechanism for distribution of mitochondria during cell division in budding yeast. AB - Mitochondria are indispensable for normal eukaryotic cell function. As they cannot be synthesized de novo and are self-replicating, mitochondria must be transferred from mother to daughter cells. Studies in the budding yeast Saccharomyces cerevisiae indicate that mitochondria enter the bud immediately after bud emergence, interact with the actin cytoskeleton for linear, polarized movement of mitochondria from mother to bud, but are equally distributed among mother and daughter cells [1] [2] [3]. It is not clear how the mother cell maintains its own supply of mitochondria. Here, we found that mother cells retain mitochondria by immobilization of some mitochondria in the 'retention zone', the base of the mother cell distal to the bud. Retention requires the actin cytoskeleton as mitochondria colocalized with actin cables in the retention zone, and mutations that perturb actin dynamics or actin-mitochondrial interactions produced retention defects. Our results support the model that equal distribution of mitochondria during cell division is a consequence of two actin-dependent processes: movement of some mitochondria into the daughter bud and immobilization of others in the mother cell. PMID- 10531007 TI - Absence of Brca2 causes genome instability by chromosome breakage and loss associated with centrosome amplification. AB - Women heterozygous for mutations in the breast-cancer susceptibility genes BRCA1 and BRCA2 have a highly elevated risk of developing breast cancer [1]. BRCA1 and BRCA2 encode large proteins with no sequence similarity to one another. Although involvement in DNA repair and transcription has been suggested, it is still not understood how loss of function of these genes leads to breast cancer [2]. Embryonic fibroblasts (MEFs) derived from mice homozygous for a hypomorphic mutation (Brca2(Tr2014)) within the 3' region of exon 11 in Brca2 [3], or a similar mutation (Brca2(Tr)) [4], proliferate poorly in culture and overexpress the tumour suppressor p53 and the cyclin-dependent kinase inhibitor p21(Waf1/Cip1). These MEFs have intact p53-dependent DNA damage G(1)-S [3] [4] and G(2)-M checkpoints [4], but are impaired in DNA double-strand break repair [3] and develop chromosome aberrations [4]. Here, we report that Brca2(Tr2014/Tr2014) MEFs frequently develop micronuclei. These abnormal DNA containing bodies were formed through both loss of acentric chromosome fragments and by chromosome missegregation, which resulted in aneuploidy. Absence of Brca2 also led to centrosome amplification, which we found associated with the formation of micronuclei. These data suggest a potential mechanism whereby loss of BRCA2 may, within subclones, drive the loss of cell-cycle regulation genes, enabling proliferation and tumourigenesis. PMID- 10531009 TI - Lateral inhibition of inositol 1,4,5-trisphosphate receptors by cytosolic Ca(2+). AB - Ryanodine and inositol 1,4,5-trisphosphate (IP(3)) receptors - two related families of Ca(2+) channels responsible for release of Ca(2+) from intracellular stores [1] - are biphasically regulated by cytosolic Ca(2+) [2] [3] [4]. It is thought that the resulting positive feedback allows localised Ca(2+)-release events to propagate regeneratively, and that the negative feedback limits the amplitude of individual events [5] [6]. Stimulation of IP(3) receptors by Ca(2+) occurs through a Ca(2+)-binding site that becomes exposed only after IP(3) has bound to its receptor [7] [8]. Here, we report that rapid inhibition of IP(3) receptors by Ca(2+) occurs only if the receptor has not bound IP(3). The IP(3) therefore switches its receptor from a state in which only an inhibitory Ca(2+) binding site is accessible to one in which only a stimulatory site is available. This regulation ensures that Ca(2+) released by an active IP(3) receptor may rapidly inhibit its unliganded neighbours, but it cannot terminate the activity of a receptor with IP(3) bound. Such lateral inhibition, which is a universal feature of sensory systems where it improves contrast and dynamic range, may fulfil similar roles in intracellular Ca(2+) signalling by providing increased sensitivity to IP(3) and allowing rapid graded recruitment of IP(3) receptors. PMID- 10531008 TI - Yeast Ku protein plays a direct role in telomeric silencing and counteracts inhibition by rif proteins. AB - Yku70p/Yku80p, the yeast Ku protein homologue, is a DNA end-binding heterodimer involved in non-homologous end joining. It also binds to telomeres, where it plays an important role in the maintenance of telomeric DNA structure [1] [2] [3] [4] [5]. Ku protein, together with Rap1p, a telomeric DNA (TG(1-3) repeat) binding protein, is also required to initiate transcriptional silencing, or telomere-position effect (TPE). Here, we provide evidence for a direct role of Ku in TPE, which is most likely to be in either the recruitment or activation of Sir4 protein at the telomere. Surprisingly, however, the essential role of Ku in TPE is to overcome the inhibitory effect of two Rap1p-interacting proteins, Rif1p and Rif2p, both of which also play an important role in telomere length regulation [6] [7]. Previous studies showed that Rif and Sir proteins compete for binding to the carboxyl terminus of Rap1p [7] [8] [9]. In the absence of this competition, for example, when RIF genes are mutated, Ku is no longer necessary for TPE, whereas the Rap1p carboxyl terminus is still absolutely required. We show that Rif1p is localized to telomeres, indicating that its inhibitory effect on TPE is direct. Our data implicate a role for Ku in the competition between Sir and Rif proteins for access to the telomeric array of Rap1p molecules, which results in a balance between telomeric silencing and telomere length control. PMID- 10531010 TI - A role for the extraembryonic yolk syncytial layer in patterning the zebrafish embryo suggested by properties of the hex gene. AB - Recent studies in mouse suggest that the extraembryonic endoderm has an important role in early embryonic patterning [1]. To analyze whether similar mechanisms operate in other vertebrates, we cloned the zebrafish homologue of Hex, a homeobox gene that is expressed asymmetrically in the mouse visceral endoderm [2]. Early expression of zebrafish hex is restricted to the dorsal portion of the yolk syncytial layer (YSL), an extraembryonic tissue. By the onset of gastrulation, hex is expressed in the entire dorsal half of the YSL, which directly underlies the cells fated to form the neural plate. We show that hex expression is initially regulated by the maternal Wnt pathway and later by a Bmp mediated pathway. Overexpression experiments of wild-type and chimeric Hex constructs indicate that Hex functions as a transcriptional repressor and its overexpression led to the downregulation of bmp2b and wnt8 expression and the expansion of chordin expression. These findings provide further evidence that the zebrafish YSL is the functional equivalent of the mouse visceral endoderm and that extraembryonic structures may regulate early embryonic patterning in many vertebrates. PMID- 10531011 TI - Mouse suppressor of fused is a negative regulator of sonic hedgehog signaling and alters the subcellular distribution of Gli1. AB - The Hedgehog (Hh) signaling pathway has critical functions during embryogenesis of both invertebrate and vertebrate species [1]; defects in this pathway in humans can cause developmental disorders as well as neoplasia [2]. Although the Gli1, Gli2, and Gli3 zinc finger proteins are known to be effectors of Hh signaling in vertebrates, the mechanisms regulating activity of these transcription factors remain poorly understood [3] [4]. In Drosophila, activity of the Gli homolog Cubitus interruptus (Ci) is likely to be modulated by its interaction with a cytoplasmic complex containing several other proteins [5] [6], including Costal2, Fused (Fu), and Suppressor of fused (Su(fu)), the last of which has been shown to interact directly with Ci [7]. We have cloned mouse Suppressor of fused (mSu(fu)) and detected its 4.5 kb transcript throughout embryogenesis and in several adult tissues. In cultured cells, mSu(fu) overexpression inhibited transcriptional activation mediated by Sonic hedgehog (Shh), Gli1 and Gli2. Co-immunoprecipitation of epitope-tagged proteins indicated that mSu(fu) interacts with Gli1, Gli2, and Gli3, and that the inhibitory effects of mSu(fu) on Gli1's transcriptional activity were mediated through interactions with both amino- and carboxy-terminal regions of Gli1. Gli1 was localized primarily to the nucleus of both HeLa cells and the Shh-responsive cell line MNS 70; co-expression with mSu(fu) resulted in a striking increase in cytoplasmic Gli1 immunostaining. Our findings indicate that mSu(fu) can function as a negative regulator of Shh signaling and suggest that this effect is mediated by interaction with Gli transcription factors. PMID- 10531012 TI - Fluorescence lifetime imaging of receptor tyrosine kinase activity in cells. AB - We report a highly specific fluorescence lifetime imaging microscopy (FLIM) method for monitoring epidermal growth factor receptor (EGFR) phosphorylation in cells based on fluorescence resonance energy transfer (FRET). EGFR phosphorylation was monitored using a green fluorescent protein (GFP)-tagged EGFR and Cy3-conjugated anti-phosphotyrosine antibodies. In this FRET-based imaging method, the information about phosphorylation is contained only in the (donor) GFP fluorescence lifetime and is independent of the antibody-derived (acceptor) fluorescence signal. A pixel-by-pixel reference lifetime of the donor GFP in the absence of FRET was acquired from the same cell after photobleaching of the acceptor. We show that this calibration, by acceptor photobleaching, works for the GFP-Cy3 donor-acceptor pair and allows the full quantitation of FRET efficiencies, and therefore the degree of exposed phosphotyrosines, at each pixel. The hallmark of EGFR stimulation is receptor dimerisation [1] [2] [3] [4] and concomitant activation of its intracellular tyrosine kinase domain [5] [6] [7]. Trans-autophosphorylation of the receptor [8] [9] on specific tyrosine residues couples the activated dimer to the intracellular signal transduction machinery as these phosphorylated residues serve as docking sites for adaptor and effector molecules containing Src homology 2 (SH2; reviewed in [10]) and phosphotyrosine-binding (PTB) [11] domains. The time-course and extent of EGFR phosphorylation are therefore important determinants of the underlying pathway and resulting cellular response. Our results strongly suggest that secondary proteins are recruited by activated receptors in endosomes, indicating that these are active compartments in signal transduction. PMID- 10531013 TI - Caffeine inhibits the checkpoint kinase ATM. AB - The basis of many anti-cancer therapies is the use of genotoxic agents that damage DNA and thus kill dividing cells. Agents that cause cells to override the DNA-damage checkpoint are predicted to sensitize cells to killing by genotoxic agents. They have therefore been sought as adjuncts in radiation therapy and chemotherapy. One such compound, caffeine, uncouples cell-cycle progression from the replication and repair of DNA [1] [2]. Caffeine therefore servers as a model compound in establishing the principle that agents that override DNA-damage checkpoints can be used to sensitize cells to the killing effects of genotoxic drugs [3]. But despite more than 20 years of use, the molecular mechanisms by which caffeine affects the cell cycle and checkpoint responses have not been identified. We investigated the effects of caffeine on the G2/M DNA-damage checkpoint in human cells. We report that the radiation-induced activation of the kinase Cds1 [4] (also known as Chk2 [5]) is inhibited by caffeine in vivo and that ATM kinase activity is directly inhibited by caffeine in vitro. Inhibition of ATM provides a molecular explanation of the attenuation of DNA-damage checkpoint responses and for the increased radiosensitivity of caffeine-treated cells [6] [7] [8]. PMID- 10531015 TI - The shower PMID- 10531014 TI - A lipid-binding domain in Wnt: a case of mistaken identity? PMID- 10531016 TI - Networks of neurons. PMID- 10531017 TI - Natural killer cells. PMID- 10531018 TI - Biology in pictures. How the snake lost its legs. PMID- 10531020 TI - Sperm competition: defining the rules of engagement. AB - Genetic and cell biological analyses of sperm behavior in the female reproductive tract are providing important clues to the mechanisms of sperm competition, a form of sexual selection that is an important force that shapes reproductive behavior, physiology and morphology in a wide range of species. PMID- 10531019 TI - Adaptive thermogenesis: orchestrating mitochondrial biogenesis. AB - The biogenesis of mitochondria requires products of the nuclear and mitochondrial genomes. Recent studies of adaptive thermogenesis have shown how mitochondrial proliferation and respiratory activity in brown fat and skeletal muscle are directed by the transcriptional coactivator PGC-1. PMID- 10531021 TI - Myosin structure: does the tail wag the dog? AB - Previous crystal structures of the myosin head have shown two different conformations, postulated to be the beginning and the end of the actomyosin power stroke. A new crystal structure reveals a dramatically different conformation; but how does this conformation fit into the force-generating cycle of actomyosin interactions? PMID- 10531022 TI - Cytoskeleton: centrosom-in absentia. AB - Recent results challenge long-held assumptions that centrosomes are essential organizers of mitotic spindles, but suggest that they couple spindle behavior with developmental and cellular events, perhaps by nucleating astral microtubules which mediate interactions with other cytoskeletal components. PMID- 10531023 TI - Axis development: the mouse becomes a dachshund. AB - Targeted deletion of the gene for GDF11, a novel member of the TGFbeta family, has been found to cause an increase in the number of thoracic and lumbar vertebrae in the mouse. This is the first hint that a secreted factor may influence the specification of segment identity. PMID- 10531025 TI - Cancer biology: extracellular proteinases in malignancy. AB - Secreted, matrix-degrading proteinases have been viewed as contributing to tumor metastasis. A recent study indicates that the gene for one of these enzymes, the matrix metalloproteinase stromelysin-1, can actually cause cancer when expressed in transgenic mice. PMID- 10531024 TI - Protein translocation: is Hsp70 pulling my chain? AB - Hsp70 proteins in the lumen of the endoplasmic reticulum and in the mitochondrial matrix are thought to drive the translocation of proteins into each organelle. Recent experiments aimed at distinguishing between two models for Hsp70 function appear to reach opposite conclusions. PMID- 10531026 TI - Cocaine addiction: clues from Drosophila on drugs. AB - Recent studies have shown that the fruitfly Drosophila exhibits behavioral sensitization in response to repeated exposure to cocaine; the exploitation of this genetically tractable model system for studying cocaine addiction is already providing new clues that may help understand the process of drug addiction in man. PMID- 10531027 TI - Rapid epithelial-sheet sealing in the Caenorhabditis elegans embryo requires cadherin-dependent filopodial priming. AB - BACKGROUND: During embryonic development, epithelia with free edges must join together to create continuous tissues that seal the interior of the organism from the outside environment; failure of epithelial sealing underlies several common human birth defects. Sealing of epithelial sheets in embryos can be extremely rapid, dramatically exceeding the rate of adherens junction formation by epithelial cells in culture or during healing of epithelial wounds. Little is known about the dynamic redistribution of cellular junctional components during such events in living embryos. RESULTS: We have used time-lapse, multiphoton laser-scanning microscopy and green fluorescent protein fusion proteins to analyze the sealing of the Caenorhabditis elegans epidermis in living embryos. Rapid recruitment of alpha-catenin to sites of filopodial contact between contralateral migrating epithelial cells, concomitant with clearing of cytoplasmic alpha-catenin, resulted in formation of nascent junctions; this preceded the formation of mature junctions. Surprisingly, upon inactivation of the entire cadherin-catenin complex, only adhesive strengthening between filopodia was reproducibly affected. Other ventral epidermal cells, which did not extend filopodia and appeared to seal along the ventral midline by coordinated changes in cell shape, successfully adhered in the absence of these proteins. CONCLUSIONS: We propose that 'filopodial priming' - prealignment of bundled actin in filopodia combined with the rapid recruitment of alpha-catenin from cytoplasmic reserves at sites of filopodial contact - accounts for the rapid rate of sealing of the embryonic epidermis of C. elegans. Filopodial priming may provide a general mechanism for rapid creation of adherens junctions during epithelial-sheet sealing in embryos. PMID- 10531028 TI - The solution structure of VAT-N reveals a 'missing link' in the evolution of complex enzymes from a simple betaalphabetabeta element. AB - BACKGROUND: The VAT protein of the archaebacterium Thermoplasma acidophilum, like all other members of the Cdc48/p97 family of AAA ATPases, has two ATPase domains and a 185-residue amino-terminal substrate-recognition domain, VAT-N. VAT shows activity in protein folding and unfolding and thus shares the common function of these ATPases in disassembly and/or degradation of protein complexes. RESULTS: Using nuclear magnetic resonance (NMR) spectroscopy, we found that VAT-N is composed of two equally sized subdomains. The amino-terminal subdomain VAT-Nn (comprising residues Met1-Thr92) forms a double-psi beta-barrel whose pseudo twofold symmetry is mirrored by an internal sequence repeat of 42 residues. The carboxy-terminal subdomain VAT-Nc (comprising residues Glu93-Gly185) forms a novel six-stranded beta-clam fold. Together, VAT-Nn and VAT-Nc form a kidney shaped structure, in close agreement with results from electron microscopy. Sequence and structure analyses showed that VAT-Nn is related to numerous proteins including prokaryotic transcription factors, metabolic enzymes, the protease cofactors UFD1 and PrlF, and aspartic proteinases. These proteins map out an evolutionary path from simple homodimeric transcription factors containing a single copy of the VAT-Nn repeat to complex enzymes containing four copies. CONCLUSIONS: Our results suggest that VAT-N is a precursor of the aspartic proteinases that has acquired peptide-binding activity while remaining proteolytically incompetent. We propose that the binding site of the protein is similar to that of aspartic proteinases, in that it lies between the psi-loops of the amino-terminal beta-barrel and that it coincides with a crescent-shaped band of positive charge extending across the upper face of the molecule. PMID- 10531029 TI - A molecular pathway leading to endoderm formation in zebrafish. AB - BACKGROUND: Several potentially important regulators of vertebrate endoderm development have been identified, including Activin-related growth factors and their receptors; transcriptional regulators encoded by the genes Mixer, Xsox17, and HNF3beta; zebrafish One-eyed pinhead (Oep), a member of the Cripto/FRL 1/Cryptic family of epidermal growth factor related proteins (EGF-CFC); and the product of the zebrafish locus casanova, which plays an essential cell-autonomous role in endoderm formation. RESULTS: Using overexpression studies and the analysis of different zebrafish mutants, we have assembled a molecular pathway that leads to endoderm formation. We report that a zebrafish Sox17 homologue is expressed during gastrulation exclusively in the endoderm and that casanova mutants lack all sox17 expression. Overexpression of mixer induces ectopic sox17 expressing cells in wild-type embryos and promotes endoderm formation in oep mutants, but does not rescue sox17 expression or endoderm formation in casanova mutants. Overexpression of a constitutively active form of the type I transforming growth factor beta (TGF-beta) receptor TARAM-A also promotes sox17 expression in wild-type and oep mutant embryos, but not in casanova mutants. We also show that the Nodal-related molecules Cyclops and Squint and the transmembrane protein Oep are essential for normal mixer expression. CONCLUSIONS: The data indicate that the following pathway leads to zebrafish endoderm formation: Cyclops and Squint activate receptors such as TARAM-A; Oep also appears to act upstream of such receptors; signals transduced by these receptors lead to the expression of mixer, Mixer then acts through casanova to promote the expression of sox17 and differentiation of the endoderm. PMID- 10531030 TI - N-linked glycans containing linear poly-N-acetyllactosamine as sorting signals in endocytosis in Trypanosoma brucei. AB - African trypanosomes, such as Trypanosoma brucei, are protozoan parasites that are transmitted by the tsetse fly and cause sleeping sickness in humans and Nagana in cattle. Trypanosomes evade the immune responses of their hosts by varying their surface coat protein (VSG) and restricting exocytosis and endocytosis to an invagination of the plasma membrane called the flagellar pocket (FP). The FP represents only 0.5% of the cellular surface but membrane turnover here occurs at high rates [1] [2] [3]. No model has yet been proposed to account for the sequestration of membrane proteins and the rate of membrane turnover that occur in the FP. Recent data have suggested that glycans are involved in the sorting of membrane proteins in polarized cells [4] [5] [6] [7]. Here, we show that N-linked glycans containing linear poly-N-acetyllactosamine (pNAL) are only associated with proteins of the FP/endocytic pathway in T. brucei and are present only in bloodstream forms of the parasite. These glycoproteins bind to tomato lectin (TL), a property that allowed their single-step isolation. Chito oligosaccharides that compete specifically for pNAL binding to TL also inhibited receptor-mediated uptake of several ligands. These results suggest a model in which N-linked linear pNAL acts as a sorting signal for endocytosis in trypanosomes. PMID- 10531031 TI - Biallelic transcription of Igf2 and H19 in individual cells suggests a post transcriptional contribution to genomic imprinting. AB - The H19 and insulin-like growth factor 2 (Igf2) genes in the mouse are models for genomic imprinting during development. The genes are located only 90 kb apart in the same transcriptional orientation [1], but are reciprocally imprinted: Igf2 is paternally expressed while H19 is maternally expressed. It has been suggested that expression of H19 and repression of Igf2 (or the converse) on a given chromosome are mechanistically linked and that the parental imprint operates at the level of transcription [2]. Although expression of Igf2 and H19 is thought to be monoallelic, the data have so far been obtained exclusively by looking at steady-state RNA levels using techniques that reflect the average activity of the genes in a cell population [3] [4]. Here, we have adapted a fluorescent in situ hybridisation (FISH) method to detect nascent RNA molecules of Igf2 and H19 at the initial transcription sites in the nuclei of wild-type mouse embryonic liver cells. Nine different transcription patterns were observed, reflecting a high heterogeneity of transcription at the single-cell level. Our observations suggest that regulation of Igf2 and H19 by parental imprinting is much more complex than previously proposed and acts at both transcriptional and post-transcriptional levels. PMID- 10531032 TI - The guanine-nucleotide-exchange factor Cdc24p is targeted to the nucleus and polarized growth sites. AB - Generation of cellular asymmetry or cell polarity plays a critical role in cell cycle-regulated morphogenetic processes involving the actin cytoskeleton. The GTPase Cdc42 regulates actin rearrangements and signal transduction pathways in all eukaryotic cells [1], and the temporal and spatial regulation of Cdc42p depends on the activity and targeting of its guanine-nucleotide exchange factor (GEF). Cdc24p, the Saccharomyces cerevisiae GEF for Cdc42p, is found in a particulate fraction and localizes to the plasma membrane [2] [3] at sites of polarized growth [4]. We show that Cdc24p labeled with green fluorescent protein (GFP-Cdc24p) was targeted to pre-bud sites, the tips and sides of enlarging buds, and mating projections in pheromone-treated cells. Unexpectedly, GFP-Cdc24p also localized to the nucleus and GFP-Cdc24p levels diminished before nuclear division followed by its reappearance in divided nuclei and mother-bud necks during cytokinesis. The Cdc24p amino-terminal 283 amino acids were necessary and sufficient for nuclear localization, which depended on the cyclin-dependent kinase inhibitor Far1p. The Cdc24p carboxy-terminal 289 amino acids were necessary and sufficient for targeting to the pre-bud site, bud, mother-bud neck, and mating projection. Targeting was independent of the Cdc24p-binding proteins Far1p, the GTPase Rsr1p/Bud1p, the scaffold protein Bem1p, and the G(beta) subunit Ste4p. These data are consistent with a temporal and spatial regulation of Cdc24p-dependent activation of Cdc42p during the cell cycle. PMID- 10531033 TI - A gene trap approach in Xenopus. AB - The frog transgenesis technique ultimately promises to make mutagenesis possible through random insertion of plasmid DNA into the genome. This study was undertaken to evaluate whether a gene trap approach combined with transgenesis would be appropriate for performing insertional mutagenesis in Xenopus embryos. Firstly, we confirmed that the transgenic technique results in stable integration into the genome and that transmission through the germline occurs in the expected Mendelian fashion. Secondly, we developed several gene trap vectors, using the green fluorescent protein (GFP) as a marker. Using these vectors, we trapped several genes in Xenopus laevis that are expressed in a spatially restricted manner, including expression in the epiphysis, the olfactory bulb and placodes, the eyes, ear, brain, muscles, tail and intestine. Finally, we cloned one of the trapped genes using 5' rapid amplification of cDNA ends polymerase chain reaction (RACE PCR). These results suggest that the transgenic technique combined with a gene trap approach might provide a powerful method for generating mutations in endogenous genes in Xenopus. PMID- 10531034 TI - Saccadic suppression precedes visual motion analysis. AB - There is now good evidence that perception of motion is strongly suppressed during saccades (rapid shifts of gaze), presumably to blunt the disturbing sense of motion that saccades would otherwise elicit. Other aspects of vision, such as contrast detection of high-frequency or equiluminant gratings, are virtually unaffected by saccades [1] [2] [3] [4] [5]. This has led to the suggestion that saccades may suppress selectively the magnocellular pathway (which is strongly implicated in motion perception), leaving the parvocellular pathway unaffected [5] [6]. Here, we investigate the neural level at which perception of motion is suppressed. We used a simple technique in which an impression of motion is generated from only two frames, allowing precise control over the stimulus [7] [8]. One frame has a certain fixed contrast, whereas the contrast of the other (the test frame) is varied to determine the threshold for motion discrimination (that is, the lowest test-frame contrast level at which the direction of motion can be correctly guessed). Contrast thresholds of the test depended strongly and non-monotonically on the contrast of the fixed-contrast frame, with a minimum at medium contrast. To study the effect of saccadic suppression, we triggered the two-frame sequence by a voluntary saccade. Thresholds during saccades increased in a way that suggested that saccadic suppression precedes motion analysis: when the test frame was first in the motion sequence there was a general depression of sensitivity, whereas when it was second, the contrast response curve was shifted to a higher contrast range, sometimes even resulting in higher sensitivity than without a saccade. The dependence on presentation order suggests that saccadic suppression occurs at an early stage of visual processing, on the single frames themselves rather than on the combined motion signal. As motion detection itself is thought to occur at an early stage, saccadic suppression must take place at a very early phenomenon. PMID- 10531035 TI - Identification of a family of human F-box proteins. AB - F-box proteins are an expanding family of eukaryotic proteins characterized by an approximately 40 aminoacid motif, the F box (so named because cyclin F was one of the first proteins in which this motif was identified) [1]. Some F-box proteins have been shown to be critical for the controlled degradation of cellular regulatory proteins [2] [3]. In fact, F-box proteins are one of the four subunits of ubiquitin protein ligases called SCFs. The other three subunits are the Skp1 protein; one of the cullin proteins (Cul1 in metazoans and Cdc53 or Cul A in the yeast Saccharomyces cerevisiae); and the recently identified Roc1 protein (also called Rbx1 or Hrt1). SCF ligases bring ubiquitin conjugating enzymes (either Ubc3 or Ubc4) to substrates that are specifically recruited by the different F box proteins. The need for high substrate specificity and the large number of known F-box proteins in yeast and worms [2] [4] suggest the existence of a large family of mammalian F-box proteins. Using Skp1 as a bait in a yeast two-hybrid screen and by searching DNA databases, we identified a family of 26 human F-box proteins, 25 of which were novel. Some of these proteins contained WD-40 domains or leucine-rich repeats; others contained either different protein-protein interaction modules or no recognizable motifs. We have named the F-box proteins that contain WD-40 domains Fbws, those containing leucine-rich repeats, Fbls, and the remaining ones Fbxs. We have further characterized representative members of these three classes of F-box proteins. PMID- 10531036 TI - Regulation of ARNO nucleotide exchange by a PH domain electrostatic switch. AB - ARNO is a member of a family of guanine nucleotide exchange factors that activate small GTPases called ADP-ribosylation factors (ARFs) [1] [2] [3], which regulate vesicular trafficking and, in one case (ARF6), also regulate cortical actin structure [4]. ARNO is located at the plasma membrane, and in the presence of activated protein kinase C (PKC) can induce cortical actin rearrangements reminiscent of those produced by active ARF6 [5] [6] [7] [8]. High-affinity binding of ARNO to membranes, which is required for exchange activity, is mediated cooperatively by a pleckstrin homology (PH) domain and an adjacent carboxy-terminal polybasic domain [3] [9]. ARNO is phosphorylated in vivo by PKC on a single serine residue, S392, located within the carboxy-terminal polybasic domain. Mutation of S392 to alanine does not prevent ARNO-mediated actin rearrangements, suggesting that phosphorylation does not lead to ARNO activation [6]. Here, we report that phosphorylation negatively regulates ARNO exchange activity through a 'PH domain electrostatic switch'. Introduction of a negatively charged phosphate into the polybasic domain reduced interaction of ARNO with membranes both in vitro and in vivo, and inhibited exchange in vitro. This regulated membrane association is similar to the myristoyl electrostatic switch that controls membrane binding of the myristoylated alanine-rich C kinase substrate (MARCKS) [10], but to our knowledge is the first demonstration of an electrostatic switch regulating the membrane interaction of a protein containing a PH domain. This mechanism allows regulation of ARNO lipid binding and exchange activity at two levels, phosphoinositide-dependent recruitment and PKC-dependent displacement from the membrane. PMID- 10531037 TI - A family of mammalian F-box proteins. AB - Ubiquitin-mediated destruction of regulatory proteins is a frequent means of controlling progression through signaling pathways [1]. F-box proteins [2] are components of modular E3 ubiquitin protein ligases called SCFs, which function in phosphorylation-dependent ubiquitination ([3] [4] [5], reviewed in [6] [7]). F box proteins contain a carboxy-terminal domain that interacts with substrates and a 42-48 amino-acid F-box motif which binds to the protein Skp1 [2] [3] [4]. Skp1 binding links the F-box protein with a core ubiquitin ligase composed of the proteins Cdc53/Cul1, Rbx1 (also called Hrt1 and Roc1) and the E2 ubiquitin conjugating enzyme Cdc34 [8] [9] [10] [11]. The genomes of the budding yeast Saccharomyces cerevisiae and the nematode worm Caenorhabditis elegans contain, respectively, 16 and more than 60 F-box proteins [2] [7]; in S. cerevisiae, the F box proteins Cdc4, Grr1 and Met30 target cyclin-dependent kinase inhibitors, G1 cyclins and transcriptional regulators for ubiquitination ([3] [4] [5] [8] [10], reviewed in [6] [7]). Only four mammalian F-box proteins (Cyclin F, Skp1, beta TRCP and NFB42) have been identified so far [2] [12]. Here, we report the identification of a family of 33 novel mammalian F-box proteins. The large number of these proteins in mammals suggests that the SCF system controls a correspondingly large number of regulatory pathways in vertebrates. Four of these proteins contain a novel conserved motif, the F-box-associated (FBA) domain, which may represent a new protein-protein interaction motif. The identification of these genes will help uncover pathways controlled by ubiquitin-mediated proteolysis in mammals. PMID- 10531038 TI - The COP9 signalosome is essential for development of Drosophila melanogaster. AB - The COP9 signalosome (originally described as the COP9 complex) is an essential multi-subunit repressor of light-regulated development in plants [1] [2]. It has also been identified in mammals, though its role remains obscure [3] [4] [5]. This complex is similar to the regulatory lid of the proteasome and eIF3 [5] [9] [10] [11] [12] and several of its subunits are known to be involved in kinase signaling pathways [4] [6] [7] [8]. No proteins homologous to COP9 signalosome components were identified in the Saccharomyces cerevisiae genome, suggesting that the COP9 signalosome is specific for multi-cellular differentiation [13]. In order to reveal the developmental function of the COP9 signalosome in animals, we have isolated Drosophila melanogaster genes encoding eight subunits of the COP9 signalosome, and have shown by co-immunoprecipitation and gel-filtration analysis that these proteins are components of the Drosophila COP9 signalosome. Yeast two hybrid assays indicated that several of these proteins interact, some through the PCI domain. Disruption of one of the subunits by either a P-element insertion or deletion of the gene caused lethality at the late larval or pupal stages. This lethality is probably a result of numerous pleiotropic effects. Our results indicate that the COP9 signalosome is conserved in invertebrates and that it has an essential role in animal development. PMID- 10531039 TI - Deletion of the Cul1 gene in mice causes arrest in early embryogenesis and accumulation of cyclin E. AB - The stability of many proteins is controlled by the ubiquitin proteolytic system, which recognizes specific substrates through the action of E3 ubiquitin ligases [1]. The SCFs are a recently described class of ubiquitin ligase that target a number of cell cycle regulators and other proteins for degradation in both yeast and mammalian cells [2] [3] [4] [5] [6]. Each SCF complex is composed of the core protein subunits Skp1, Rbx1 and Cul1 (known as Cdc53 in yeast), and substrate specific adaptor subunits called F-box proteins [2] [3] [4]. To understand the physiological role of SCF complexes in mammalian cells, we generated mice carrying a deletion in the Cul1 gene. Cul1(-/-) embryos arrested around embryonic day 6.5 (E6.5) before the onset of gastrulation. In all cells of the mutant embryos, cyclin E protein, but not mRNA, was highly elevated. Outgrowths of Cul1( /-) blastocysts had limited proliferative capacity in vitro and accumulated cyclin E in all cells. Within Cul1(-/-) blastocyst cultures, trophoblast giant cells continued to endocycle despite the elevated cyclin E levels. These results suggest that cyclin E abundance is controlled by SCF activity, possibly through SCF-dependent degradation of cyclin E. PMID- 10531040 TI - MEF2 is upregulated during cardiac hypertrophy and is required for normal post natal growth of the myocardium. AB - In mammals, growth of the fetal heart is regulated by proliferation of cardiac muscle cells. At later stages of pre-natal life, this proliferation diminishes profoundly [1] [2] and the dramatic expansion in heart size during the transition to adulthood is due exclusively to hypertrophy of individual cardiomyocytes [3] [4] [5]. Cardiomyocyte hypertrophy also contributes to the pathology of most post natal heart disease [6] [7] [8] [9] [10]. Within this context, numerous signal transduction pathways have been implicated as the link between the effector(s) and altered cardiac gene expression [11] [12] [13] [14] [15] [16]. A common pathway has yet to be discovered, however. Here, we found that the activity of the stress-activated kinase p38 was enhanced in both types of cardiomyocyte hypertrophy. We also found that a target of the activated p38 kinase is the cardiac transcription factor MEF2. Transgenic mice expressing a dominant-negative form of MEF2C displayed attenuated post-natal growth of the myocardium. These results provide the first evidence for a single pathway regulating both normal and pathologic cardiomyocyte hypertrophy. PMID- 10531042 TI - Moron peer review. PMID- 10531041 TI - Identification of novel F-box proteins in Xenopus laevis. PMID- 10531044 TI - Rhone-Poulenc S.A PMID- 10531043 TI - DNA double-strand break repair. PMID- 10531045 TI - Synaptic roulette. PMID- 10531046 TI - Mickey mozart? PMID- 10531047 TI - Seven-transmembrane proteins as odorant and chemosensory receptors. AB - The olfactory systems of various species solve the challenging problem of general molecular recognition in widely differing ways. Despite this variety, the molecular receptors are invariably G protein-coupled seven-transmembrane proteins, and are encoded by the largest gene families known to exist in a given animal genome. Receptor gene families have been identified in vertebrates and two invertebrate species, the nematode Caenorhabditis elegans and the fruit fly Drosophila melanogaster. The complexity of the odorant receptor repertoire is estimated in mouse and rat at 1000 genes, or 1 percent of the genome, surpassing that of the immunoglobulin and T cell receptor genes combined. Two distinct seven transmembrane gene families may encode in rodents the chemosensory receptors of the vomeronasal organ, which is specialized in the detection of pheromones. Remarkably, these five receptor families have practically no sequence homology among them. Genetic manipulation experiments in mice imply that vertebrate odorant receptors may fulfill a dual role, also serving as address molecules that guide axons of olfactory sensory neurons to their precise target in the brain. PMID- 10531048 TI - The olfactory bulb: coding and processing of odor molecule information. AB - Olfactory sensory neurons detect a large variety of odor molecules and send information through their axons to the olfactory bulb, the first site for the processing of olfactory information in the brain. The axonal connection is precisely organized so that signals from 1000 different types of odorant receptors are sorted out in 1800 glomeruli in the mouse olfactory bulb. Individual glomerular modules presumably represent a single type of receptor and are thus tuned to specific molecular features of odorants. Local neuronal circuits in the bulb mediate lateral inhibition among glomerular modules to sharpen the tuning specificity of output neurons. They also mediate synchronized oscillatory discharges among specific combinations of output neurons and may contribute to the integration of signals from distinct odorant receptors in the olfactory cortex. PMID- 10531049 TI - The vomeronasal organ. AB - The vomeronasal organ (VNO) is a chemoreceptor organ enclosed in a cartilaginous capsule and separated from the main olfactory epithelium. The vomeronasal neurons have two distinct types of receptor that differ from each other and from the large family of odorant receptors. The VNO receptors are seven-transmembrane receptors coupled to GTP-binding protein, but appear to activate inositol 1,4,5 trisphosphate signaling as opposed to cyclic adenosine monophosphate. The nature of stimulus access suggests that the VNO responds to nonvolatile cues, leading to activation of the hypothalamus by way of the accessory olfactory bulb and amygdala. The areas of hypothalamus innervated regulate reproductive, defensive, and ingestive behavior as well as neuroendocrine secretion. PMID- 10531050 TI - Olfactory reception in invertebrates. AB - Recent progress in understanding the principles and mechanisms in olfaction is the result of multidisciplinary research efforts that explored chemosensation by using a variety of model organisms. Studies on invertebrates, notably nematodes, insects, and crustaceans, to which diverse experimental approaches can be applied, have greatly helped elucidate various aspects of olfactory signaling. From the converging results of genetic, molecular, and physiological studies, a common set of chemosensory mechanisms emerges. Recognition and discrimination of odorants as well as chemo-electrical transduction and processing of olfactory signals appear to be mediated by fundamentally similar mechanisms in phylogenetically diverse animals. The common challenge of organisms to decipher the world of odors was apparently met by a phylogenetically conserved strategy. Thus, comparative studies should continue to provide important contributions toward an understanding of the sense of smell. PMID- 10531051 TI - A systems perspective on early olfactory coding. AB - This review critically examines neuronal coding strategies and how they might apply to olfactory processing. Basic notions such as identity, spatial, temporal, and correlation codes are defined and different perspectives are brought to the study of neural codes. Odors as physical stimuli and their processing by the early olfactory system, one or two synapses away from the receptors, are discussed. Finally, the concept of lateral inhibition, as usually understood and applied to odor coding by mitral (or equivalent) cells, is challenged and extended to a broader context, possibly more appropriate for olfactory processing. PMID- 10531053 TI - Contact-dependent inhibition of cortical neurite growth mediated by notch signaling. AB - The exuberant growth of neurites during development becomes markedly reduced as cortical neurons mature. In vitro studies of neurons from mouse cerebral cortex revealed that contact-mediated Notch signaling regulates the capacity of neurons to extend and elaborate neurites. Up-regulation of Notch activity was concomitant with an increase in the number of interneuronal contacts and cessation of neurite growth. In neurons with low Notch activity, which readily extend neurites, up regulation of Notch activity either inhibited extension or caused retraction of neurites. Conversely, in more mature neurons that had ceased their growth after establishing numerous connections and displayed high Notch activity, inhibition of Notch signaling promoted neurite extension. Thus, the formation of neuronal contacts results in activation of Notch receptors, leading to restriction of neuronal growth and a subsequent arrest in maturity. PMID- 10531052 TI - Beta-secretase cleavage of Alzheimer's amyloid precursor protein by the transmembrane aspartic protease BACE. AB - Cerebral deposition of amyloid beta peptide (Abeta) is an early and critical feature of Alzheimer's disease. Abeta generation depends on proteolytic cleavage of the amyloid precursor protein (APP) by two unknown proteases: beta-secretase and gamma-secretase. These proteases are prime therapeutic targets. A transmembrane aspartic protease with all the known characteristics of beta secretase was cloned and characterized. Overexpression of this protease, termed BACE (for beta-site APP-cleaving enzyme) increased the amount of beta-secretase cleavage products, and these were cleaved exactly and only at known beta secretase positions. Antisense inhibition of endogenous BACE messenger RNA decreased the amount of beta-secretase cleavage products, and purified BACE protein cleaved APP-derived substrates with the same sequence specificity as beta secretase. Finally, the expression pattern and subcellular localization of BACE were consistent with that expected for beta-secretase. Future development of BACE inhibitors may prove beneficial for the treatment of Alzheimer's disease. PMID- 10531054 TI - All-Inorganic Field Effect Transistors Fabricated by Printing. AB - A solution of cadmium selenide nanocrystals was used to print inorganic thin-film transistors with field effect mobilities up to 1 square centimeter per volt second. This mobility is an order of magnitude larger than those reported for printed organic transistors. A field effect was achieved by developing a synthesis that yielded discretely sized nanocrystals less than 2 nanometers in size, which were free of intimately bound organic capping groups. The resulting nanocrystal solution exhibited low-temperature grain growth, which formed single crystal areas encompassing hundreds of nanocrystals. This process suggests a route to inexpensive, all-printed, high-quality inorganic logic on plastic substrates. PMID- 10531055 TI - Bidirectional Semiconductor Laser. AB - A semiconductor laser capable of operating under both positive and negative bias voltage is reported. Its active region behaves functionally as two different laser materials, emitting different wavelengths, depending on the design, when biased with opposite polarities. This concept was used for the generation of two wavelengths (6.3 and 6.5 micrometers) in the midinfrared region of the spectrum from a single quantum cascade laser structure. The two wavelengths are excited independently of each other and separated in time. This may have considerable impact on various semiconductor laser applications including trace gas analysis in remote sensing applications with differential absorption spectroscopy. PMID- 10531056 TI - Asymmetric Electrical Structure in the Mantle Beneath the East Pacific Rise at 17 degrees S. AB - The magnetotelluric component of the Mantle Electromagnetic and Tomography (MELT) Experiment measured the electrical resistivity structure of the mantle beneath the fast-spreading southern East Pacific Rise (EPR). The data reveal an asymmetric resistivity structure, with lower resistivity to the west of the ridge. The uppermost 100 kilometers of mantle immediately to the east of the ridge is consistent with a dry olivine resistivity structure indicating a mantle depleted of melt and volatiles. Mantle resistivities to the west of the ridge are consistent with a low-melt fraction (about 1 to 2 percent interconnected melt) distributed over a broad region and extending to depths of about 150 kilometers. The asymmetry in resistivity structure may be the result of asymmetric spreading rates and a westward migration of the ridge axis and suggests distinct styles of melt formation and delivery in the mantle beneath the two plates. PMID- 10531057 TI - Subtropical North Atlantic Temperatures 60,000 to 30,000 Years Ago. AB - A reconstruction of sea surface temperature based on alkenone unsaturation ratios in sediments of the Bermuda Rise provides a detailed record of subtropical climate from 60,000 to 30,000 years ago. Northern Sargasso Sea temperatures changed repeatedly by 2 degrees to 5 degrees C, covarying with high-latitude temperatures that were previously inferred from Greenland ice cores. The largest temperature increases were comparable in magnitude to the full glacial-Holocene warming at the site. Abrupt cold reversals of 3 degrees to 5 degrees C, lasting less than 250 years, occurred during the onset of two such events (Greenland interstadials 8 and 12), suggesting that the largest, most rapid warmings were especially unstable. PMID- 10531058 TI - Evolution of Complexity in Paleozoic Ammonoid Sutures. AB - The septal sutures of 588 genera of Paleozoic ammonoids showed a 1600 percent increase in mean complexity over 140 million years. Within 475 ancestor/descendant pairs, descendants were more than twice as likely to be more complex than their ancestors. Twelve subclades (373 genera) averaged 34 percent increased complexity. These patterns are compatible with an active or driven system of long-term bias for increased complexity. Mass extinctions acted in opposition to this long-term trend, tending to eliminate more-complex forms and resetting the trend with each extinction event. PMID- 10531059 TI - A Triassic Fauna from Madagascar, Including Early Dinosaurs. AB - The discovery of a Middle to Late Triassic ( approximately 225 to 230 million years old) terrestrial vertebrate fauna from Madagascar is reported. This fauna documents a temporal interval not well represented by continental vertebrate assemblages elsewhere in the world. It contains two new prosauropod dinosaurs, representing some of the earliest dinosaur occurrences known globally. This assemblage provides information about the poorly understood transition to the dinosaur-dominated faunas of the latest Triassic. PMID- 10531060 TI - Interlocked feedback loops within the Drosophila circadian oscillator. AB - Drosophila Clock (dClk) is rhythmically expressed, with peaks in mRNA and protein (dCLK) abundance early in the morning. dClk mRNA cycling is shown here to be regulated by PERIOD-TIMELESS (PER-TIM)-mediated release of dCLK- and CYCLE (CYC) dependent repression. Lack of both PER-TIM derepression and dCLK-CYC repression results in high levels of dClk mRNA, which implies that a separate dClk activator is present. These results demonstrate that the Drosophila circadian feedback loop is composed of two interlocked negative feedback loops: a per-tim loop, which is activated by dCLK-CYC and repressed by PER-TIM, and a dClk loop, which is repressed by dCLK-CYC and derepressed by PER-TIM. PMID- 10531061 TI - Light-independent role of CRY1 and CRY2 in the mammalian circadian clock. AB - Cryptochrome (CRY), a photoreceptor for the circadian clock in Drosophila, binds to the clock component TIM in a light-dependent fashion and blocks its function. In mammals, genetic evidence suggests a role for CRYs within the clock, distinct from hypothetical photoreceptor functions. Mammalian CRY1 and CRY2 are here shown to act as light-independent inhibitors of CLOCK-BMAL1, the activator driving Per1 transcription. CRY1 or CRY2 (or both) showed light-independent interactions with CLOCK and BMAL1, as well as with PER1, PER2, and TIM. Thus, mammalian CRYs act as light-independent components of the circadian clock and probably regulate Per1 transcriptional cycling by contacting both the activator and its feedback inhibitors. PMID- 10531062 TI - Negative feedback regulation of TGF-beta signaling by the SnoN oncoprotein. AB - Smad proteins mediate transforming growth factor-beta (TGF-beta) signaling to regulate cell growth and differentiation. The SnoN oncoprotein was found to interact with Smad2 and Smad4 and to repress their abilities to activate transcription through recruitment of the transcriptional corepressor N-CoR. Immediately after TGF-beta stimulation, SnoN is rapidly degraded by the nuclear accumulation of Smad3, allowing the activation of TGF-beta target genes. By 2 hours, TGF-beta induces a marked increase in SnoN expression, resulting in termination of Smad-mediated transactivation. Thus, SnoN maintains the repressed state of TGF-beta-responsive genes in the absence of ligand and participates in negative feedback regulation of TGF-beta signaling. PMID- 10531063 TI - Aging-dependent large accumulation of point mutations in the human mtDNA control region for replication. AB - Progressive damage to mitochondrial DNA (mtDNA) during life is thought to contribute to aging processes. However, this idea has been difficult to reconcile with the small fraction of mtDNA so far found to be altered. Here, examination of mtDNA revealed high copy point mutations at specific positions in the control region for replication of human fibroblast mtDNA from normal old, but not young, individuals. Furthermore, in longitudinal studies, one or more mutations appeared in an individual only at an advanced age. Some mutations appeared in more than one individual. Most strikingly, a T414G transversion was found, in a generally high proportion (up to 50 percent) of mtDNA molecules, in 8 of 14 individuals above 65 years of age (57 percent) but was absent in 13 younger individuals. PMID- 10531064 TI - Crystal structure of the ectodomain of human transferrin receptor. AB - The transferrin receptor (TfR) undergoes multiple rounds of clathrin-mediated endocytosis and reemergence at the cell surface, importing iron-loaded transferrin (Tf) and recycling apotransferrin after discharge of iron in the endosome. The crystal structure of the dimeric ectodomain of the human TfR, determined here to 3.2 angstroms resolution, reveals a three-domain subunit. One domain closely resembles carboxy- and aminopeptidases, and features of membrane glutamate carboxypeptidase can be deduced from the TfR structure. A model is proposed for Tf binding to the receptor. PMID- 10531065 TI - Microtubule disassembly by ATP-dependent oligomerization of the AAA enzyme katanin. AB - Katanin, a member of the AAA adenosine triphosphatase (ATPase) superfamily, uses nucleotide hydrolysis energy to sever and disassemble microtubules. Many AAA enzymes disassemble stable protein-protein complexes, but their mechanisms are not well understood. A fluorescence resonance energy transfer assay demonstrated that the p60 subunit of katanin oligomerized in an adenosine triphosphate (ATP)- and microtubule-dependent manner. Oligomerization increased the affinity of katanin for microtubules and stimulated its ATPase activity. After hydrolysis of ATP, microtubule-bound katanin oligomers disassembled microtubules and then dissociated into free katanin monomers. Coupling a nucleotide-dependent oligomerization cycle to the disassembly of a target protein complex may be a general feature of ATP-hydrolyzing AAA domains. PMID- 10531066 TI - Neuronal activity-dependent cell survival mediated by transcription factor MEF2. AB - During mammalian development, electrical activity promotes the calcium-dependent survival of neurons that have made appropriate synaptic connections. However, the mechanisms by which calcium mediates neuronal survival during development are not well characterized. A transcription-dependent mechanism was identified by which calcium influx into neurons promoted cell survival. The transcription factor MEF2 was selectively expressed in newly generated postmitotic neurons and was required for the survival of these neurons. Calcium influx into cerebellar granule neurons led to activation of p38 mitogen-activated protein kinase-dependent phosphorylation and activation of MEF2. Once activated, MEF2 regulated neuronal survival by stimulating MEF2-dependent gene transcription. These findings demonstrate that MEF2 is a calcium-regulated transcription factor and define a function for MEF2 during nervous system development that is distinct from previously well-characterized functions of MEF2 during muscle differentiation. PMID- 10531067 TI - Apoptosis of T cells mediated by Ca2+-induced release of the transcription factor MEF2. AB - T cell receptor (TCR)-induced apoptosis of thymocytes is mediated by calcium dependent expression of the steroid receptors Nur77 and Nor1. Nur77 expression is controlled by the transcription factor myocyte enhancer factor 2 (MEF2), but how MEF2 is activated by calcium signaling is still obscure. Cabin1, a calcineurin inhibitor, was found to regulate MEF2. MEF2 was normally sequestered by Cabin1 in a transcriptionally inactive state. TCR engagement led to an increase in intracellular calcium concentration and the dissociation of MEF2 from Cabin1, as a result of competitive binding of activated calmodulin to Cabin1. The interplay between Cabin1, MEF2, and calmodulin defines a distinct signaling pathway from the TCR to the Nur77 promoter during T cell apoptosis. PMID- 10531068 TI - Reactive or infectious arthritis. PMID- 10531069 TI - Chronic wrist pain: diagnosis and management. Development and use of a new algorithm. AB - OBJECTIVE: Chronic wrist pain can be difficult to manage and the differential diagnosis is extensive. To provide guidelines for assessment of the painful wrist an algorithm was developed to encourage a structured approach to the diagnosis and management of these patients. METHODS: A review of the literature on causes of chronic wrist pain was undertaken; history taking, physical examination and imaging studies were evaluated systematically to determine which of the many potential conditions was the cause of the wrist pain. Chronic wrist pain was subdivided into pain of probable intra-articular or extra-articular origin. By means of this classification a clinical algorithm was developed to establish a diagnosis and its clinical usefulness was tested in a prospective study of 84 patients presenting to our outpatient clinic. RESULTS: A definite diagnosis could be established in 59% (49 of 84) of the cases by careful history taking, extensive physical examination, plain radiographs, ultrasound examination and bone scintigraphy. In 19% of the cases (16 of 84) a probable diagnosis was made resulting in a total figure 78% (65 of 84). Additional imaging studies (arthrography, magnetic resonance imaging and computed tomography) increased the definite diagnoses to 70% (59 of 84). CONCLUSION: The algorithm proved easy to use and by the use of careful history taking, thorough physical examination and simple imaging techniques (ultrasonography and scintigraphy) a diagnosis was made in 78% of cases. PMID- 10531070 TI - Heberden's and Bouchard's nodes. PMID- 10531071 TI - An unusual cause of atlanto-axial subluxation. PMID- 10531073 TI - Analysis of 16 different matrix metalloproteinases (MMP-1 to MMP-20) in the synovial membrane: different profiles in trauma and rheumatoid arthritis. AB - OBJECTIVE: To define the pattern of mRNA expression of all human matrix metalloproteinases (MMPs) described to date in rheumatoid arthritis (RA) and traumatic synovial membrane, in order to differentiate between a physiological tissue remodelling pattern and that associated with inflammatory tissue destruction. METHODS: Analysis of SwissProt protein and EMBL/GenBank nucleotide sequence banks, protein sequence alignment, reverse transcriptase-polymerase chain reaction and nucleotide sequencing were used. RESULTS: MMP-2 (gelatinase A), MMP-3 (stromelysin-1), MMP-11 (stromelysin-3) and MMP-19 were constitutively expressed. MMP-1 (fibroblast type collagenase), MMP-9 (gelatinase B) and MMP-14 (MT1-MMP) were expressed in all RA, but only in 55-80% of trauma samples. MMP-13 (collagenase-3) and MMP-15 (MT2-MMP) were expressed exclusively in RA (80-90% of the samples). MMP-20 (enamelysin) was absent and MMP-8 (collagenase-2) was rarely found in RA or trauma. All other MMPs (-7, -10, -12, -16, -17) had an intermediate pattern of expression. CONCLUSIONS: Some MMPs without interstitial collagenase activity seem to have a constitutive pattern of expression and probably participate in physiological synovial tissue remodelling. Some MMPs are exclusively associated to RA synovitis, for example, MMP-13, which preferentially degrades type II collagen and aggrecan, and MMP-15, which activates proMMP-2 and proMMP-13 and is involved in tumour necrosis factor alpha processing. This clear cut rheumatoid/inflammatory MMP profile, more complex than has been previously appreciated, may facilitate inflammatory tissue destruction in RA. PMID- 10531072 TI - Expression of laminins and their integrin receptors in different conditions of synovial membrane and synovial membrane-like interface tissue. AB - OBJECTIVE: To demonstrate the expression of laminins (Lns) and their integrin (Int) receptors in different synovial samples and synovial membrane-like interface tissues from well fixed and aseptically loosened total hip replacement (THR), and the potential role of Ln-Int interaction in the production of collagenases and cytokines. METHODS: Immunohistochemical staining was done to detect the distribution of EHS Ln, Ln alpha2, alpha3, alpha5, beta1, beta2 chains and Int alpha1, alpha2, alpha3, alpha6, beta1, beta4 subunits in different samples. Double immunofluorescence labelling was used to find colocalisation of Int alpha6 subunit and collagenase-1/collagenase-3/TNFalpha/IL6. RESULTS: General Ln immunoreactivity was detected in all specimens. Ln alpha5, beta1 and beta2, but not alpha2 and alpha3 chains were seen in the synovial lining and the basement membrane of blood vessels with the intensity/extent of labelling in the following rank order: rheumatoid arthritis (RA) loosened prostheses, osteoarthritis, well fixed prostheses, traumatic knees. Among Int subunits, staining for beta1 was usually the strongest, followed by staining for Int alpha6, alpha1, alpha3, and alpha2 subunits, with the same rank order for overall expression of Lns. Int beta4 subunit was not detectable in most of the specimens. Double labelling focused on Int alpha6 subunit disclosed its frequent colocalisation with collagenases 1 and 3 and with tumour necrosis factor alpha and interleukin 6 in synovial lining. CONCLUSION: Synovial lining contains Ln-10, Ln-11, and Int alpha6beta1 and alpha1beta1 receptors. In aseptic loosening of THR, interface tissue has a similar Ln subtype and Int receptor composition as RA synovium, which confirms its "lining-like" phenotype. Synovial lining does not contain Ln-5 (alpha3beta3gamma2) or Int alpha6beta4, which are components of epithelial hemidesmosomes. The expression of Lns and their Int receptors is upregulated in inflammation. The close spatial relation between Ln and its Int receptors in synovial lining cells containing proteinases and cytokines suggests a potential role in joint destruction and prosthetic loosening. PMID- 10531074 TI - Prevalence and clinical features of cryoglobulinaemia in multitransfused beta thalassaemia patients. AB - OBJECTIVE: The aim of the study was to determine the prevalence of cryoglobulinaemia and its clinical features among beta-thalassaemia patients. METHODS: Eighty eight multitransfused beta-thalassaemia patients were studied. They were physically examined and asked about the presence of cryoglobulinaemia related symptoms. Hepatitis C virus (HCV) serology, HCV-RNA, HCV subtypes, viraemia, serum ferritin, liver and kidney function tests, rheumatoid factor (RF), circulating immune complexes (CIC), complement levels and autoantibodies were all evaluated. The patients were divided into four groups: HCV-RNA positive patients with and without cryoglobulinaemia (groups A and B), HCV-Ab positive/HCV RNA negative patients (group C), HCV-Ab negative patients (group D). RESULTS: Cryoglobulinaemia was present in 35 of 53 (66.0%) patients with chronic HCV infection. They had higher viraemia than non-cryoglobulinaemic viral carriers, but no statistical difference relating to sex or HCV subtypes was found. In comparison with the other groups, group A patients were older, had undergone transfusion therapy for a longer period, had received a higher number of transfusions, and had increased levels of RF and CIC, as well as consumption of C4; in addition, they had a higher prevalence of cirrhosis. Cutaneous lesions (purpura, Raynaud's phenomenon, nodules and leg rash), peripheral neuropathy and sicca syndrome symptoms were present only in group A. Musculoskeletal symptoms (bone pain, arthralgia and myalgia), weakness, splenomegaly, lymphadenopathy, skin ulcers and proteinuria were also commoner in group A, but the difference did not reach statistical significance, possibly because of partial overlap between cryoglobulinaemia and beta-thalassaemia syndromes. CONCLUSION: Because of its high prevalence in multitransfused beta-thalassaemia patients, cryoglobulinaemia needs to be systematically studied and considered in the differential diagnosis of various beta-thalassaemia manifestations. PMID- 10531075 TI - A measure of limited joint motion and deformity correlates with HLA-DRB1 and DQB1 alleles in patients with rheumatoid arthritis. AB - OBJECTIVE: To assess factors associated with a poor outcome in rheumatoid arthritis (RA), a measure was developed of limited joint motion and deformity, a deformity index (DI), and correlated biochemical and genetic variables with the magnitude of the DI. METHODS: Forty patients were evaluated in a cross sectional study. Clinical measures included the DI and Health Assessment Questionnaire, and disease variables included the erythrocyte sedimentation rate, C reactive protein, rheumatoid factor, and HLA-DRB1 and DQB1 alleles. RESULTS: Significant correlations were noted between increasing DI and duration of RA and concentration of C reactive protein. Patients with a DQB1*301 allele or DR4 allele had a higher DI than those without, and a positive trend was noted between increasing DI and dose of DRB1 RA susceptibility alleles. The trend was lost when a non-linear regression technique was used to remove the effect attributable to C reactive protein, suggesting an interrelation between persistent inflammation and genetics in determining total joint damage. CONCLUSIONS: The DI may be useful to study interactions between genetic and inflammatory processes in rheumatoid disease progression. PMID- 10531076 TI - Expression of adhesion molecules on synovial fluid and peripheral blood monocytes in patients with inflammatory joint disease and osteoarthritis. AB - OBJECTIVE: To determine the presence of adhesion molecules on monocytes/macrophages (Mphi) from peripheral blood (PB) and synovial fluid (SF) in patients with osteoarthritis (OA) and inflammatory joint diseases (rheumatoid (RA) and reactive arthritis (ReA)) in order to improve our understanding of the possible mechanisms underlying the inflammatory process. METHODS: Whole blood and SF cells were stained with monoclonal antibodies against CD11a (LFA-1), CD15 s (sialyl-Lewis X), CD44, CD54, VLA-4, and HLA-DR counterstained with anti-CD14 antibodies as a Mphi marker for dual fluorescence analysis by flowcytometry. RESULTS: On PB-Mphi, CD15s was markedly increased in both RA as well as ReA compared with OA. Furthermore, in the PB LFA-1, CD44, and HLA-DR showed a higher surface density on Mphi in ReA than in OA. Comparison between SF and PB showed significantly higher CD44 and CD54 expression on SF-Mphi. These molecules play an important part in lymphocyte-Mphi interaction. CONCLUSION: In PB from patients with inflammatory joint diseases, Mphi are activated, allowing recruitment into the synovial compartment. These disorders, in contrast with OA seem to be "systemic" in nature. Within the SF, different adhesion molecules are expressed on CD14(+) Mphi as compared with PB. PMID- 10531077 TI - A randomised trial of differentiated prednisolone treatment in active rheumatoid arthritis. Clinical benefits and skeletal side effects. AB - OBJECTIVES: To study benefits and skeletal side effects of carefully monitored prednisolone treatment in patients with active rheumatoid arthritis. METHODS: One hundred and two patients with active rheumatoid arthritis were randomly allocated to treatment with disease modifying anti-inflammatory drug (DMARD) alone or DMARD and prednisolone in a one year follow up study. Prednisolone was given in a dose regimen adapted to the disease activity of the individual patient. The mean dose was 6 mg and the mean cumulated dose was 2160 mg. Patients were followed up with disease activity parameters, radiograph of the hands (Larsen score), and bone mineral density (BMD) of the lumbar spine, distal forearm and hand. At one year 26 patients had withdrawn from the investigation leaving 76 patients for evaluation. RESULTS: The results showed that disease activity in the prednisolone treated group was reduced within two weeks. In the DMARD alone group disease activity was gradually reduced over months. At six months there was no difference between the groups as evaluated by an improvement score using a number of ACR criteria. Prednisolone in the present set up was not able to protect significantly against radiological disease progression, although there was a trend towards less progression in Larsen score in the prednisolone group, a matter that was further underlined in an intention to treat analysis. BMD data revealed a significant reduction in spinal BMD in the prednisolone group, whereas prednisolone seemed to have a protective effect against bone loss in the hand and distal forearm. CONCLUSIONS: This study does not allow any firm conclusions for or against the treatment of rheumatoid arthritis with prednisolone. The data suggest that the beneficial effects of prednisolone are not as clear cut in established rheumatoid arthritis as in early disease. Furthermore the data indicate that treatment in the chosen relatively low dose does not provide sufficient control of disease. On the other hand the spinal bone loss observed in the prednisolone group does invite considerations about using higher doses. PMID- 10531078 TI - Antidepressants and upper gastrointestinal bleeding. PMID- 10531079 TI - Carbon monoxide poisoning. PMID- 10531080 TI - Hyperbaric oxygen in carbon monoxide poisoning. PMID- 10531081 TI - Preventing the cardiotoxicity of anthracyclines by dexrazoxane. PMID- 10531082 TI - HIV and tuberculosis in the commonwealth. PMID- 10531084 TI - Health secretary will target heart disease PMID- 10531083 TI - Paediatricians propose plan to insure every US child. PMID- 10531087 TI - In brief PMID- 10531086 TI - Doctors not obliged to carry out treatment they think "futile". PMID- 10531085 TI - Professor tried to get surgery stopped. PMID- 10531088 TI - Sertraline approved for post-traumatic stress disorder PMID- 10531089 TI - WMA to tackle adverse health conditions in prisons. PMID- 10531090 TI - WHO celebrates triumph over river blindness. PMID- 10531091 TI - Antismoking campaigns should "target 4 year olds". PMID- 10531092 TI - Scottish secretary of the BMA resigns PMID- 10531093 TI - Medical abortion still not available in most countries. PMID- 10531094 TI - Study into medical errors planned for the UK. PMID- 10531096 TI - MRI found suitable for detecting coronary heart disease PMID- 10531095 TI - Young less tolerant of mentally ill than the old. PMID- 10531097 TI - Prediction of survival for preterm births by weight and gestational age: retrospective population based study. AB - OBJECTIVE: To produce current data on survival of preterm infants. DESIGN: Retrospective population based study. SETTING: Trent health region. SUBJECTS: All European and Asian live births, stillbirths, and late fetal losses from 22 to 32 weeks' gestation, excluding those with major congenital malformations, in women resident in the Trent health region between 1 January 1994 and 31 December 1997. MAIN OUTCOME MEASURES: Birth weight and gestational age specific survival for both European and Asian infants (a) known to be alive at the onset of labour, and (b) admitted for neonatal care. RESULTS: 738 deaths occurred in 3760 infants born between 22 and 32 weeks' gestation during the study period, giving an overall survival rate of 80.4%. The survival rate for the 3489 (92.8%) infants admitted for neonatal care was 86.6%. For European infants known to be alive at the onset of labour, significant variations in gestation specific survival by birth weight emerged from 24 weeks' gestation: survival ranged from 9% (95% confidence interval 7% to 13%) for infants of birth weight 250-499 g to 21% (16% to 28%) for those of 1000-1249 g. At 27 weeks' gestation, survival ranged from 55% (49% to 61%) for infants of birth weight 500-749 g (below the 10th centile) to 80% (76% to 85%) for those of 1250-1499 g. Infants who were large for dates (>/=27 weeks' gestation) had a slightly reduced, but not significant, predicted survival. Similar survival rates were observed for Asian infants. The odds ratio for the survival of infants from a multiple birth compared with singleton infants was 1.4 (1.1 to 1.8). Survival graphs for infants admitted for neonatal care are presented by sex. CONCLUSION: Easy to use birth weight and gestational age specific predicted survival graphs for preterm infants facilitate decision making for clinicians and parents. It is important that these graphs are representative, are produced for a geographically defined population, and are not biased towards the outcomes of particular centres. Such graphs, produced in two stages, allow for the changing pattern of survival of infants from the start of the intrapartum period to immediately after admission for neonatal care. PMID- 10531099 TI - Identifying very fat and very thin children: test of criterion standards for screening test. PMID- 10531098 TI - Two view mammography at incident screens: cost effectiveness analysis of policy options. AB - OBJECTIVE: To determine the cost effectiveness of two view mammography at incident screens. DESIGN: Incremental cost effectiveness analyses recognising differences in current reading policy, based on effectiveness data from an observational study. SETTING: Breast screening programmes in England and Wales. MAIN OUTCOME MEASURES: Health service costs, cancers detected, incremental cost effectiveness ratios per cancer detected, whole time equivalent staff. RESULTS: For programmes currently using one view with some form of double reading, the incremental cost effectiveness ratio of two view mammography at incident screens ranged between 6589 pounds and 6716 pounds, depending on the reading policy. For programmes currently using one view with single reading, two policy options were found to be more efficient than two view single reading: one view with double reading (arbitration; incremental cost effectiveness ratio of 210 pounds) and two view double reading (arbitration). If programmes using one view with single reading changed to double reading (arbitration) and then subsequently to two views double reading (arbitration), additional cancers could be detected with an incremental cost effectiveness ratio of 7983. The implementation cost of two view mammography at incident screens in programmes in England and Wales would be 2.9 million pounds and would require 13.4 whole time equivalent radiologists. CONCLUSIONS: The cost effectiveness of two view mammography at incident screens depends on the film reading policy. A policy of two view mammography at incident screens in England and Wales would be efficient only if programmes using single reading moved to double reading. Given limited resources, priority should be given to introducing double reading in the subset of programmes currently using single reading as this requires fewer additional radiologists and is more cost effective. PMID- 10531100 TI - Maternal mortality in the former east Germany before and after reunification: changes in risk by marital status. PMID- 10531101 TI - Knowledge PMID- 10531102 TI - A word out of place PMID- 10531104 TI - Women in the operating theatre PMID- 10531103 TI - Association between selective serotonin reuptake inhibitors and upper gastrointestinal bleeding: population based case-control study. AB - OBJECTIVE: To examine the association between selective serotonin reuptake inhibitors and risk of upper gastrointestinal bleeding. DESIGN: Population based case-control study. SETTING: General practices included in the UK general practice research database. SUBJECTS: 1651 incident cases of upper gastrointestinal bleeding and 248 cases of ulcer perforation among patients aged 40 to 79 years between April 1993 and September 1997, and 10 000 controls matched for age, sex, and year that the case was identified. INTERVENTIONS: Review of computer profiles for all potential cases, and an internal validation study to confirm the accuracy of the diagnosis on the basis of the computerised information. MAIN OUTCOME MEASURES: Current use of selective serotonin reuptake inhibitors or other antidepressants within 30 days before the index date. RESULTS: Current exposure to selective serotonin reuptake inhibitors was identified in 3.1% (52 of 1651) of patients with upper gastrointestinal bleeding but only 1.0% (95 of 10 000) of controls, giving an adjusted rate ratio of 3.0 (95% confidence interval 2.1 to 4.4). This effect measure was not modified by sex, age, dose, or treatment duration. A crude incidence of 1 case per 8000 prescriptions was estimated. A small association was found with non-selective serotonin reuptake inhibitors (relative risk 1.4, 1.1 to 1.9) but not with antidepressants lacking this inhibitory effect. None of the groups of antidepressants was associated with ulcer perforation. The concurrent use of selective serotonin reuptake inhibitors with non-steroidal anti-inflammatory drugs increased the risk of upper gastrointestinal bleeding beyond the sum of their independent effects (15.6, 6.6 to 36.6). A smaller interaction was also found between selective serotonin reuptake inhibitors and low dose aspirin (7.2, 3.1 to 17.1). CONCLUSIONS: Selective serotonin reuptake inhibitors increase the risk of upper gastrointestinal bleeding. The absolute effect is, however, moderate and about equivalent to low dose ibuprofen. The concurrent use of non steroidal anti-inflammatory drugs or aspirin with selective serotonin reuptake inhibitors greatly increases the risk of upper gastrointestinal bleeding. PMID- 10531105 TI - Recent advances: otolaryngology. PMID- 10531106 TI - Lesson of the week: digital examination for oral cancer. PMID- 10531107 TI - The baby and the bathwater PMID- 10531109 TI - London operating theatres PMID- 10531108 TI - ABC of complementary medicine. Homoeopathy. PMID- 10531110 TI - Geriatric care in the United Kingdom: aligning services to needs. PMID- 10531111 TI - The NHS in Dumfries and Galloway: straining but optimistic. PMID- 10531113 TI - A doctor's reputation PMID- 10531112 TI - The hospital of the future. Better out than in? Alternatives to acute hospital care. PMID- 10531114 TI - Effects of drug overdose in television drama on presentations for self poisoning. Antifreeze poisonings give more insight into copycat behaviour. PMID- 10531115 TI - Medical fiction could be misleading. PMID- 10531116 TI - Association between type 1 diabetes and hib vaccine. Causal relation is likely. PMID- 10531117 TI - Radioiodine and thyroid eye disease. Routine steroid prophylaxis is not yet justified. PMID- 10531118 TI - Non-attendance at outpatients departments. More information was needed for non-UK readers. PMID- 10531119 TI - Anaesthetists need consent, but not written consent. PMID- 10531120 TI - Risks of medicine and air travel. PMID- 10531121 TI - Immunosuppression in renal transplantation. Meta-analysis should not have included one of the studies. PMID- 10531123 TI - Cecil thompson buchanan adams PMID- 10531122 TI - Australia has considerable experience of transporting critically ill patients. PMID- 10531124 TI - Juniors' pay offer will not mean a pay cut PMID- 10531125 TI - Parkinson's disease: the treatment options PMID- 10531126 TI - Difficult asthma PMID- 10531127 TI - Learning to be you PMID- 10531128 TI - Carbon monoxide PMID- 10531130 TI - Words failed me PMID- 10531129 TI - In praise of hunch backing PMID- 10531132 TI - Survival graphs for preterm infants would help counselling PMID- 10531131 TI - Lancet editor defends decision to publish GM research paper PMID- 10531133 TI - Two view mammography at subsequent screens is cost effective only with double reading PMID- 10531134 TI - Body mass index in children varies with height PMID- 10531135 TI - Maternal mortality among east germans worsened after reunification PMID- 10531136 TI - Selective serotonin reuptake inhibitors increase risk of upper gastrointestinal bleeding PMID- 10531137 TI - Care of the elderly in the UK needs restructuring PMID- 10531138 TI - Lower extremity venography : still the gold standard PMID- 10531139 TI - Lower extremity venography : still the gold standard. PMID- 10531140 TI - The dull-edged sword of inhaled corticosteroids. PMID- 10531141 TI - "Won't get fooled again" (by tuberculosis) PMID- 10531142 TI - Mechanical ventilation in hematopoietic stem cell transplant patients: is there need for reevaluation? PMID- 10531143 TI - Nosocomial pneumonia : blood cultures remain useful. PMID- 10531144 TI - Is blood pressure response to the Valsalva maneuver related to neurohormones, exercise capacity, and clinical findings in heart failure? AB - OBJECTIVES: To investigate the relationship of the BP response to the Valsalva maneuver (VM) to parameters of congestive heart failure (CHF) other than hemodynamic measures. DESIGN: Comparison of neurohormones (atrial natriuretic peptide [ANP], brain natriuretic peptide [BNP], norepinephrine [NE]), parameters of spiroergometry, and clinical parameters with BP response to the VM. SETTING: Tertiary care center. PATIENTS: Forty-five patients with stable CHF (ejection fraction, 28 +/- 7%). MEASUREMENTS: Pulse amplitude ratio (PAR) calculated between the end and the beginning of the VM using the last two and the first three beats of the straining phase. Failure of the systolic BP to fall below the resting level during the VM. RESULTS: Patients in the New York Heart Association class III (n = 15) had a higher PAR than those in class II (0.82 +/- 0.21 vs 0.63 +/- 0.20; p < 0.01). There was a close correlation between PAR and ANP (r = 0.76) and BNP (r = 0.62), whereas other parameters were less well correlated (eg, for peak f1.gif" BORDER="0">O(2), r = -0.35; p < 0.05). Patients with failure of the systolic BP to fall below the resting level (n = 24) had higher neurohormones (mean ANP, 246 +/- 158 vs 84 +/- 43 pg/mL; mean BNP, 282 +/- 289 vs 81 +/- 85 pg/mL; p < 0.001; mean NE, 3.9 +/- 1.7 vs 3.4 +/- 1.5 nmol/L; nanosecond), lower exercise capacity (19.8 +/- 5.2 vs 23.0 +/- 3.7 mL/kg/min; p < 0.05), and their quality of life (Minnesota questionnaire) was more compromised (31 +/- 19 vs 18 +/- 15; p < 0. 05). CONCLUSIONS: The BP response to the VM is related to a broad range of clinical and neurohumoral parameters of CHF. Whether or not it is also related to prognosis remains to be determined. Nevertheless, this easily applicable test should be part of the assessment of patients with CHF. PMID- 10531145 TI - Mechanism of overshoot in cardiac function during recovery from submaximal exercise in man. AB - BACKGROUND: A sudden increase (overshoot) in the left ventricular ejection fraction during the recovery from maximal exercise has been reported in patients with coronary artery disease, but its mechanism has not been fully clarified. We investigated whether this phenomenon may occur in normal subjects, and whether it depends on the intensity of exercise. METHODS: Thirteen normal subjects (mean [+/ SD] age, 59 +/- 8 years old) performed two levels (25 W and 50 W) of mild intensity, constant-work-rate exercise for 6 min on a cycle ergometer. Left ventricular function was monitored continuously during the recovery from exercise using a computerized cadmium telluride detector. RESULTS: An overshoot was observed in the ejection fraction during the first minute of recovery compared with the end-exercise value. The overshoot in the ejection fraction during recovery after the 50-W exercise was greater than that seen after the 25-W exercise. An overshoot phenomenon in stroke volume was also observed during the recovery from 50-W exercise. CONCLUSIONS: The overshoot in cardiac function observed during the early phase of recovery, which was caused mainly by an immediate decrease in end-systolic volume, occurred even after exercise of mild intensity. This phenomenon appears to suggest the existence of a transient mismatch between cardiac contractility and afterload reduction during the recovery from mild-intensity exercise, even in normal subjects. PMID- 10531146 TI - Coronary artery disease in potential lung transplant recipients > 50 years old: the role of coronary intervention. AB - STUDY OBJECTIVES: To review the experience of the Alfred Hospital in the systematic assessment of coronary artery disease (CAD) using coronary angiography (CA), and the subsequent management of CAD, in potential lung transplant recipients. DESIGN: Retrospective study. SETTING: The Alfred Hospital Lung and Heart Transplant Service. PATIENTS: CAD risk factors were sought in potential recipients of lung transplantation (LTx) who were > 50 years old, including a history of smoking, male gender, hypertension, diabetes, hypercholesterolemia, angina, and a family history of CAD. When feasible, and in the presence of more than one cardiac risk factor, CA was performed. RESULTS: From 243 referral patients who were > 50 years old, 97 were listed for LTx, and 77 underwent LTx. Four patients were refused LTx on the basis of CAD. Of 101 patients undergoing a detailed cardiac assessment for LTx, 83 had smoked, 56 were male, 48 had hypercholesterolemia, 22 had hypertension, 4 had diabetes, and 3 had a history consistent with angina. Eighty-five patients underwent CA. In 32 cases, CA revealed CAD, and half of these cases were significant stenoses. Eight patients who were assessed required intervention. Five patients of this group have been transplanted; of these, three patients underwent coronary artery grafting at the time of LTx, and two patients underwent preoperative angioplasty or stenting. Only one of these five patients died < 90 days postoperatively. CONCLUSION: Significant CAD is a common finding in older patients who are presenting for LTx. Coronary revascularization for severe large vessel stenoses can allow safe LTx. CAD risk factors may predict who should undergo CA, but further studies of clinical and noninvasive assessments of CAD are needed if CA is to be safely avoided in patients at low risk of CAD. PMID- 10531147 TI - The relationship between polymorphisms in the endothelial cell nitric oxide synthase gene and the platelet GPIIIa gene with myocardial infarction and venous thromboembolism in African Americans. AB - STUDY OBJECTIVES: To determine whether the polymorphic dinucleotide repeats found in intron 4 of the endothelial cell nitric oxide synthase (ecNOS) gene and the platelet GPIIIa PLA(1)/A(2) polymorphism are associated with myocardial infarction (MI) and venous thromboembolism (VTE) in African Americans. Because these two genes may interact physiologically, the third objective was to determine if there was a relationship between the polymorphisms with respect to MI and VTE. DESIGN: A hospital-based case-control study. After informed consent was obtained, blood used for DNA extraction was drawn from the subjects. SETTING: The study was conducted in the Anticoagulant Clinic and the Cardiology Clinic at Grady Memorial Hospital in Atlanta Georgia. PATIENTS: Subjects were recruited from African-American patients with a reported history of MI (n = 110) or VTE (n = 91). Control subjects (n = 185) without a history of cardiovascular or venous disease were recruited from an outpatient clinic. MEASUREMENTS AND RESULTS: The 393 ecNOS allele was more common among MI cases (36%; p = 0.01) and VTE cases (35%; p = 0.04) than among control subjects (26%). There was no association between the GPIIIa genotypes and either MI or VTE. However, among the MI subjects, there was a strong association between the ecNOS 393/393 genotype and the Pl(A2) allele. It was also found that the frequency of the 393 allele was higher in African-American persons (0.26) compared with what has been reported for Australian Caucasians (0. 14) and Japanese (0.10). CONCLUSIONS: The 393 allele but not the Pl(A2) allele was significantly associated with both MI and VTE in African Americans. Homozygosity for the 393 allele was significantly associated to the diagnosis of MI prior to the age of 45. The combination of the 393 allele and a Pl(A2) allele was also highly associated with MI. The frequency of the 393 allele was significantly higher in African Americans than what has been reported for other populations. This study furthers not only extends the association of the 393 allele to VTE but has demonstrated an interaction with the Pl(A2) allele with respect to MI. PMID- 10531148 TI - Retrograde cerebral perfusion as an adjunct to prolonged hypothermic circulatory arrest. AB - STUDY OBJECTIVE: This study was designed to evaluate the use of retrograde cerebral perfusion (RCP) combined with deep hypothermic circulatory arrest (DHCA) in the treatment of complex congenital and adult cardiac disease. DESIGN: Retrospective chart review of 52 cardiac surgery patients (34 male and 18 female; age range, 3 weeks to 89 years old; mean age, 60 years old) who received RCP in conjunction with DHCA from July 1991 through August 1998. RESULTS: Surgical procedures consisted of the following: (1) repair of ascending aortic aneurysms (n = 16); (2) repair of type A aortic dissection (n = 16); (3) repair of arch aneurysms (n = 10); (4) renal cell carcinoma with tumor extension to the inferior vena cava (IVC) and right atrium (n = 5); (6) coronary artery bypass grafting and concomitant aortic valve replacement with calcified aorta (n = 2); (7) Norwood procedure and take down of a Pott's shunt (n = 2); and (8) massive air embolism treatment (n = 1). Mean RCP time was 39 min (range, 3 to 88 min). Thirteen patients had RCP times > 60 min. Mean core temperature (rectal or bladder) was 19 degrees C (range, 15 degrees to 28 degrees C). There were six early deaths, four of which were related to persistent low-output cardiac failure, and two resulted from perioperative stroke. All remaining patients recovered fully without neurologic deficits. CONCLUSION: RCP is a reliable and technically appealing tool that does the following: (1) it improves DHCA safety and is applicable in a variety of clinical settings with relative ease; (2) it potentially provides oxygen and nutritional support to the brain during DHCA; (3) it helps remove air and other debris from the cerebral vessels; and (4) it is useful in dealing with congenital heart disease and tumor extension into the IVC. PMID- 10531149 TI - Complement activation in coronary artery bypass grafting patients without cardiopulmonary bypass: the role of tissue injury by surgical incision. AB - STUDY OBJECTIVES: Complement activation is a trigger in inducing inflammation in patients who undergo coronary artery bypass grafting (CABG) and is usually thought to be induced by the use of cardiopulmonary bypass (CPB). In this study, we examined whether tissue injury caused by chest surgical incision per se contributes to complement activation in CABG patients. DESIGN: Prospective study. SETTING: Thorax center in university hospital. PATIENTS: Twenty-two patients undergoing CABG without CPB were prospectively divided into two groups: a small chest incision via an anterolateral thoracotomy representing a minimized tissue injury (lateral group, n = 8), and a conventional median sternotomy representing a large tissue injury (median group, n = 14). Biochemical markers indicating complement activation as well as systemic inflammatory response were determined before, during, and after the operation. MEASUREMENTS AND RESULTS: Plasma concentrations of complement 3a increased in both the lateral and median groups right after chest incision (p < 0.01 and p < 0.05, respectively) and by the end of operation increased only in the median group (p < 0.01). The terminal complement complex 5b-9 did not increase in the lateral group, but it did increase in the median group both after incision and by the end of the operation (p < 0.05 and p < 0.05, respectively). During surgery, complement 4a did not increase, suggesting that it is the alternative rather than the classic pathway that is involved in complement activation by tissue injury. Postoperatively, interleukin-6 production was greater in the median group (p < 0.01) than the lateral group (p < 0.05), suggesting a more pronounced inflammatory response to a larger chest incision. CONCLUSIONS: Tissue injury caused by surgical incision contributes to complement activation in CABG patients who are operated on without CPB. A small anterolateral thoracotomy is associated with reduced complement activation in comparison with a median sternotomy. PMID- 10531150 TI - Physical development of surgically treated patients with primary spontaneous pneumothorax. AB - STUDY OBJECTIVES: There have been many studies on the physical characteristics at the time of contraction of a primary spontaneous pneumothorax (PSP), but it has not been shown when and how such physical characteristics develop. These issues were investigated. PATIENTS AND DESIGN: Physical development of 27 male patients with PSP were examined. Their physical records were collected with the patients' permission, and standard curves, estimated from the Japanese nationwide records in the year corresponding to the ages of the patients, were plotted as control values. RESULTS: The height of patients was already greater at 6 years of age. It showed a marked increase from 11 to 14 years. The body weight was more than the standard until 9 years, but it became less after age 11, and this difference increased after age 15. Rohrer's index was significantly lower than the standard at all ages, and the difference was particularly large from 11 to 15 years. In the standard group, there was a balance between the annual height and weight gain. In the patient group, annual weight gain was similar to that in the standard group whereas height began to increase 2 years earlier, and as a result, ectomorphy, which was also observed before this age, became marked at this age. CONCLUSIONS: The rapid increase in the vertical dimension of the thorax compared with the horizontal dimension during the period of rapid physical development is considered to affect intrathoracic pressure at the apex of lung, which would have some influence on enhancing cyst formation. PMID- 10531151 TI - High prevalence of detectable deep venous thrombosis in patients with acute pulmonary embolism. AB - STUDY OBJECTIVES: Because specific studies are unavailable, the exact prevalence of detectable "residual" deep venous thrombosis (DVT) in patients with acute pulmonary embolism (PE) is unknown. DESIGN: Review of clinical records and radiologic documents of consecutive patients. SETTING: Pulmonary diseases and radiology departments at a university hospital. PATIENTS: All patients hospitalized in the Department of Pulmonary Diseases with a diagnosis of acute PE during a 5-year period (1984 to 1988). During this period, the diagnosis of PE was based exclusively on pulmonary angiography, and bilateral lower limb venography was routine in patients with proven acute PE. MEASUREMENTS AND RESULTS: Among 228 consecutive patients with angiography-proven PE, 213 underwent bilateral lower limb venography within 48 h of the diagnosis. Venography demonstrated DVT in 174 patients (81.7%; 95% confidence interval, 76.5 to 86.9%), including 128 patients (60%) with proximal DVT. Signs or symptoms of DVT were present in only 72 patients (42%) with DVT. The prevalence of detectable DVT was significantly lower in patients with recent pelvic surgery or delivery (6 of 12, 50%) than in the other patients, whatever their individual risk factors (p < 0.05). The mean pulmonary vascular obstruction was significantly lower in patients with normal venography than in patients with detectable DVT (37.6 +/- 20.9% vs 48.4 +/- 21.7%; p = 0.007). CONCLUSIONS: Lower limb venography demonstrates a high prevalence (82%) of residual DVT in patients with angiography proven PE. These data should be taken into account in the diagnostic and therapeutic management of patients with suspected or proven PE. PMID- 10531152 TI - Incidence of acute pulmonary embolism in a general hospital: relation to age, sex, and race. AB - PURPOSE: The purpose of this investigation is to determine the incidence of acute pulmonary embolism (PE) according to age, sex, and race in a tertiary care general hospital. BACKGROUND: Population-based investigations and autopsy studies have shown that acute PE occurs predominantly in middle-aged and elderly people. The incidence of PE according to age, race, and sex in a general hospital has been only sparsely studied. METHODS: Patients with PE diagnosed by a high probability ventilation/perfusion lung scan or pulmonary angiography were identified in a tertiary care general hospital. The incidence of PE was determined according to age, sex, and race. RESULTS: The incidence of PE was 400 of 175,730 (0.23%; 95% CI, 0.21 to 0.25%). The incidence was linearly related to age (r = 0.94). Among patients >/= 50 years of age, the incidence of PE was higher among women (0.40% vs 0.29%; p < 0.01). The incidence was comparable among patients < 50 years of age. African Americans showed an incidence of 0.26%, and whites showed an incidence of 0. 21% (p < 0.05). CONCLUSION: Acute PE in a tertiary care hospital is more frequent than previously reported among short-term hospitals. Occasionally, young adults and adolescents had PE, although PE occurred primarily among middle-aged and elderly patients. Among patients >/= 50 years of age, the incidence of PE was higher among women. The incidence was not higher among women < 50 years of age, suggesting that childbirth and birth control pills had little impact. Only a trivial difference of the incidence of PE was observed among African Americans compared to whites. PMID- 10531153 TI - Effects of long-term infusion of prostacyclin on exercise performance in patients with primary pulmonary hypertension. AB - STUDY OBJECTIVES: To determine whether long-term IV prostacyclin (PGI(2)) use improves exercise capacity in patients with primary pulmonary hypertension (PPH). DESIGN: Cycle ergometry and the 6-min walk was used to evaluate the exercise performance of patients with PPH. The patients underwent serial exercise testing after starting continuous IV PGI(2) and were followed up for 19.5 +/- 7.5 months. Peak work, peak oxygen consumption (f1.gif" BORDER="0">O(2)), peak O(2) pulse, and distance walked in 6 min were used to evaluate performance. BACKGROUND: PPH is characterized by medial hypertrophy and intimal proliferation of the pulmonary arterioles, leading to elevation of pulmonary artery pressure, right ventricular failure, and death. Palliative treatment consists of vasodilators, anticoagulants, cardiac glycosides, diuretics, and transplantation. PGI(2), a potent vasodilator and inhibitor of platelet aggregation, has been used for long term treatment when conventional therapy has been unsuccessful. PATIENTS: Sixteen patients with PPH (10 women, 6 men; mean age, 24 years). RESULTS: At the initiation of PGI(2), peak work (+/- SD) was 35.5 +/- 11% of predicted; peak f1.gif" BORDER="0">O(2), 39 +/- 10.4%; peak O(2) pulse, 5.0 +/- 1.7 mL/min; and distance on the 6-min walk, 428 +/- 78 feet. At 18 to 27 months, peak work increased to 58.8 +/- 23% of predicted (p = 0.001), peak f1.gif" BORDER="0">O(2) increased to 52 +/- 15% of predicted (p = 0. 02), peak O(2) pulse increased to 7.1 +/- 3.0 mL/beat (p = 0.004), and performance on the 6-min walk increased to 526 +/- 62 feet (p = 0.001). There was a positive correlation between peak f1.gif" BORDER="0">O(2) and peak 6-min walk of 0.6 (p < 0.005) and between peak work and peak 6-min walk of 0.6 (p < 0.005). CONCLUSIONS: Exercise capacity improved in our patients at up to 27 months of follow-up. Exercise testing is helpful in assessing the functional capacity of patients with PPH and may be useful in guiding therapy. Patients who deteriorate while receiving optimal conventional therapy should be considered for IV PGI(2) therapy. PMID- 10531154 TI - The acute effects of dexfenfluramine on human and porcine pulmonary vascular tone and resistance. AB - STUDY OBJECTIVES: Treatment with anorectics has become an important aspect of care for the severely obese. One such anorectic, the phenylethylamine dexfenfluramine (dFen), has been associated with the development of pulmonary hypertension. It works by reducing the neuronal uptake of 5-hydroxytryptamine (5 HT; serotonin) through inhibition of the 5-HT transporter. In this study we investigated whether dFen has a direct vasoconstrictor action on human and porcine pulmonary vasculature. DESIGN: For the human study, tissue was obtained from patients who had undergone lung and heart-lung transplantation. The effect of dFen was studied in seven isolated colloid perfused human lungs and in rings of human pulmonary artery (PA) dissected from the lungs of a further 19 patients. For the porcine study, regional pulmonary vascular resistances (PVRs) were measured in isolated perfused porcine lungs. Vasoconstriction was assessed following dFen alone and in combination with hypoxia, cyclo-oxygenase blockade (indomethacin, 10(-5) mol/L), or nitric oxide synthase (NOS) blockade (N(G)-nitro L-arginine, 10(-5) mol/L). RESULTS: In the human study, 5-HT and dFen caused only limited increases in tension of isolated rings of PA. The concentration of dFen, 10(-4) mol/L, that was needed to increase tension was higher than that found normally in treated patients where peak levels are 3. 3 x 10(-7) mol/L. Other vasoconstrictors such as prostaglandin F(2)alpha, 10(-5) mol/L, and the thromboxane analog U46619, 10(-6) mol/L, produced far greater increases in tension. Ketanserin, 10(-4) mol/L, attenuated the constrictor response to 5-HT but had no effect on the constrictor response to dFen. Removal of the endothelium did not influence the response to dFen. In the isolated ventilated and perfused lungs, dFen caused an increase in PVR again only at a comparatively high concentration, 10(-4) mol/L. In the porcine study, dFen, 10(-4) mol/L, did not increase any PVR during normoxia or following NOS blockade. Small insignificant increases in PVR occurred during hypoxia and after cyclo-oxygenase blockade. CONCLUSION: These results do not support the view that dFen would act as a direct vasoconstrictor when given in the usual doses. However, delayed elimination of dFen could raise tissue concentrations to high levels and give rise to vasoconstriction and pulmonary hypertension. PMID- 10531155 TI - Nocturnal cortisol secretion in asthmatic patients after inhalation of fluticasone propionate. AB - OBJECTIVES: This study was designed to assess the relationship between the degree of airflow obstruction and the suppression of the hypothalamic-pituitary-adrenal axis after inhalation of fluticasone propionate (FP) in asthmatic patients with varying degrees of airway obstruction. STUDY DESIGN: The nocturnal cortisol production (from 10:00 PM to 6:00 AM), defined as the integrated area under the curve of nocturnal plasma cortisol, was measured following inhalation of a placebo or a single dose of 500 microg FP at 8:00 PM in 28 patients with mild to moderate asthma, in a single, blind, 2-night study. RESULTS: The mean morning rise of cortisol decreased significantly following a single dose of inhaled FP. When the total nocturnal cortisol production after the second night (when the FP was inhaled) was compared to that after the first night (when the placebo was administered), it was found to have decreased by 29.4%. There was a statistically significant correlation between the FEV(1) and the fall in cortisol production just before the inhalation of FP (p < 0. 001). There was no correlation between baseline cortisol production and the fall in cortisol production. CONCLUSIONS: Our findings suggest that the degree of airway obstruction affects the systemic bioavailability of FP. FP is likely to induce a more severe decrease in nocturnal cortisol secretion in less obstructed patients. In order to reduce the risk for systemic side effects, the patient's degree of airway obstruction should be considered when planning inhaled FP treatment. PMID- 10531156 TI - Evaluation of the effect of a large volume spacer on the systemic bioactivity of fluticasone propionate metered-dose inhaler. AB - BACKGROUND: Inhaled corticosteroids such as fluticasone propionate (FP) have dose related systemic effects, including adrenal suppression. We have therefore investigated the effect of adding a large volume spacer on the systemic bioactivity of FP given via a pressurized metered-dose inhaler (pMDI). METHODS: Fourteen healthy volunteers (mean age, 29.9 years old) were studied using an open, randomized, placebo-controlled, three-way crossover design. Single doses of the following were given at 5:00 PM in a randomized sequence: (1) eight puffs of FP by pMDI, 1.76 mg (250 microg ex-valve, 220 microg ex-actuator); (2) eight puffs of FP by pMDI, 250 microg, with 750-mL spacer (Volumatic; Allen & Hanburys; Uxbridge, UK); and (3) eight puffs of placebo by pMDI. Measurements were made after each dose, including overnight and early morning urinary cortisol/creatinine ratios and 8:00 AM serum cortisol. RESULTS: Significant (p < 0.05) suppression of all three end points occurred with each active treatment compared to treatment with placebo. Furthermore, significant (p < 0.05) additional suppression occurred when comparing FP by pMDI alone to FP by pMDI with spacer. Geometric mean fold differences (95% confidence interval for fold difference) between FP by pMDI alone and FP by pMDI with spacer were 1.94-fold (1.00-3.78) for overnight urinary cortisol/creatinine ratio and 1.98-fold (1.26 3.10) for 8:00 AM serum cortisol. This was mirrored by a twofold rise in the number of values for uncorrected overnight urinary cortisol < 10 nmol/10 h: placebo treatment (none of 14 subjects); FP by pMDI (6 of 14 subjects; 43%); and FP by pMDI with spacer (12 of 14 subjects; 86%). CONCLUSIONS: The use of a large volume spacer with FP by pMDI results in a twofold increase in the systemic bioavailability as assessed by sensitive measures of adrenal suppression. This, in turn, reflects a twofold improvement in respirable dose delivery with the spacer device. PMID- 10531157 TI - Effect of sputum induction on spirometric measurements and arterial oxygen saturation in asthmatic patients, smokers, and healthy subjects. AB - BACKGROUND: Sputum production induced by inhalation of hypertonic saline solution has been proposed as a technique to collect secretions and inflammatory cells from the airways of subjects with bronchial asthma or with a history of smoking. The aim of this study was to determine the effect of a sputum induction procedure on spirometric results and arterial oxygen saturation (SaO(2)) in asthmatic patients, smokers, and healthy subjects. METHODS: We recruited 14 subjects suffering from asthma (11 men and 3 women; age range, 18 to 49 years), 14 subjects with a history of smoking (5 men and 9 women; age range, 23 to 64 years), and 9 healthy volunteers (7 men and 2 women; age range, 28 to 54 years). To obtain a sample of induced sputum, all subjects inhaled a mist of 3% hypertonic saline solution nebulized for 5 min and repeated the cycle no more than four times. Asthmatic patients were pretreated with 200 microg salbutamol (inhaled). During sputum induction, the transcutaneous SaO(2) was continuously measured and baseline, fall, and the differences between baseline and fall SaO(2) were recorded. Additionally, we measured the duration of mild desaturation (change in SaO(2), < 4%) and of marked desaturation (change in SaO(2), > 5%) in each subject. Finally, baseline FEV(1) and changes in FEV(1) as a percentage of baseline values were recorded in all subjects. RESULTS: We found that baseline and fall SaO(2) values for the three groups were similar. However, in each group a significant mean change in SaO(2) was evident during sputum production (asthmatic patients, 6.0%; smokers, 5.3%; healthy subjects, 6.0%). Moreover, the mean durations of mild desaturation were 7 min, 21 s in asthma patients; 8 min, 24 s in smokers; and 7 min, 16 s in healthy subjects. Similarly, the durations of marked desaturation were 1 min, 25 s in asthmatic patients, 1 min, 19 s in smokers, and 1 min, 21 s in healthy subjects. The mean (+/- SD) fall in FEV(1) was not statistically different among the three groups (asthmatic patients, 1.36 +/- 5.6%; smokers, 7.58 +/- 11.76%; and healthy subjects, 0.05 +/- 9.6%). However, one smoker did experience excessive bronchoconstriction (fall in FEV(1), > 20%). CONCLUSIONS: This study demonstrated a significant and comparable fall in SaO(2) during sputum induction by inhalation of hypertonic saline solution in asthmatic patients, smokers, and healthy subjects. The results suggest that subjects who are hypoxemic before sputum induction require SaO(2) monitoring during the procedure. PMID- 10531158 TI - Specific antibody response against the 23-valent pneumococcal vaccine in patients with alpha(1)-antitrypsin deficiency with and without bronchiectasis. AB - OBJECTIVE: To assess the specific antibody response against polyvalent pneumococcal vaccine in patients with alpha(1)-antitrypsin deficiency (AATD) and respiratory infections. DESIGN AND PARTICIPANTS: We investigated specific IgG, IgG1, and IgG2 antibody responses against the 23-valent antipneumococcal vaccine in 18 patients with AATD phenotype PiZZ, 9 of whom had bronchiectasis and 4 a history of recurrent pneumonia, and compared them with a control group of 40 healthy volunteers. INTERVENTIONS: Blood samples were drawn just prior to and 3 weeks after immunization. MEASUREMENTS AND RESULTS: Quantification of specific IgG and its subclasses was performed by an enzyme-linked immunosorbent assay. For patients with AATD, mean increases in specific antipneumococcal titers were 4.7 fold (25 to 75% quartiles, 2.5- to 6.8-fold) for total IgG, 3.2-fold (1.2- to 4.9 fold) for IgG1, and 2.1-fold (1.8- to 3.7-fold) for IgG2. For the control group, the values were 3.3-fold (1.8- to 5.8-fold) for total IgG, 2. 5-fold (1.9- to 3.4 fold) for IgG1, and 3.1-fold (1.9- to 4.5-fold) for IgG2; differences were not significant. Patients with bronchiectasis showed a tendency toward higher levels of IgG subclasses than both control subjects and patients without bronchiectasis; however, there was a tendency toward lower postvaccination serum levels of specific antipneumococcal IgG, IgG1, and IgG2 in patients with bronchiectasis compared with patients without bronchiectasis, but this trend did not reach statistical significance. Three of the four patients with recurrent pneumonia did not show an appropriate IgG2 response. CONCLUSIONS: These results suggest that, as a group, patients with AATD have a preserved antibody response against pneumococcal polysaccharides. Patients with bronchiectasis show a tendency toward a decreased antibody response, even with increased serum levels of most Ig types. Individuals with an impaired IgG2 response seem to be at increased risk of recurrent pneumonia. Considering the pernicious effect of pulmonary infections on these patients and the preserved antibody response in a majority of them, pneumococcal vaccination should be recommended to patients with AATD. PMID- 10531159 TI - The relationships among pulmonary function, aerobic fitness, and cognitive functioning in older COPD patients. AB - STUDY OBJECTIVES: To study the predictive relationships among age, pulmonary function, aerobic fitness, and cognition in people with COPD. DESIGN: Observational study conducted during baseline testing with COPD patients who volunteered to participate in an exercise intervention. PARTICIPANTS: Older adults (age, 56 to 80 years) with COPD. MEASUREMENTS AND RESULTS: Age, depression, education level, aerobic fitness, blood oxygen saturation levels, and pulmonary function were assessed. Participants were randomly assigned to take cognitive tests of (1) fluid intelligence, (2) processing speed and working memory span, or (3) processing speed and inhibition. After controlling for education and depression (F(2,57) = 7.43; r(2) = 0. 21), performance on the 6-min walk (F(1,56) = 15.27; r(2) = 0.17) and age (F(1,55) = 7.52; r(2) = 0.08) were significant predictors of fluid intelligence. On the speed-of-processing task, performance on the 6-min walk (F(1,30) = 8.17; r(2) = 0.20), maximum voluntary ventilation (F(1,29) = 5.81; r(2) = 0.16), and age (F(1,28) = 5.26; r(2) = 0.10) were significant predictors. FVC was a significant predictor (F(1,25) = 6.37; r(2) = 0.18) of working memory span. The ability to inhibit a response was not significantly predicted by any of the variables assessed. CONCLUSIONS: In an older COPD sample, age, aerobic fitness, and pulmonary function are predictive of cognitive performance on various tasks. In particular, age and aerobic fitness are predictive of speed of processing, which is a cognitive variable that may itself underlie performance on a majority of cognitive tasks. PMID- 10531160 TI - Does aging modify pulmonary tuberculosis?: A meta-analytical review. AB - STUDY OBJECTIVES: To evaluate the differences in the clinical, radiologic, and laboratory features of pulmonary tuberculosis (TB) in older patients, as compared to younger patients. DESIGN: A meta-analysis (the Schmidt-Hunter method) of published works found in MEDLINE and other sources was performed. A total of 12 studies were collected, and each variable was submitted to meta-analysis. RESULTS: No differences were found between older (>/= 60 years old) and younger TB patients with respect to male predominance, evolution time before diagnosis, prevalence of cough, sputum production, weight loss, fatigue/malaise, radiographic upper lobes lesions, positive acid-fast bacilli in sputum, anemia or hemoglobin level, and serum aminotransferases. A lower prevalence of fever, sweating, hemoptysis, cavitary disease, and positive purified protein derivative, as well as lower levels of serum albumin and blood leukocytes were noticed among older patients. In addition, the older population had a greater prevalence of dyspnea and some concomitant conditions, such as cardiovascular disorders, COPD, diabetes, gastrectomy history, and malignancies. CONCLUSIONS: This meta analytical review identified the main differences of older TB patients, as compared to younger TB patients, that should be considered during the diagnostic evaluation. Most of these differences are explained by the already known physiologic changes that occur during aging. PMID- 10531161 TI - Predicting active pulmonary tuberculosis using an artificial neural network. AB - BACKGROUND: Nosocomial outbreaks of tuberculosis (TB) have been attributed to unrecognized pulmonary TB. Accurate assessment in identifying index cases of active TB is essential in preventing transmission of the disease. OBJECTIVES: To develop an artificial neural network using clinical and radiographic information to predict active pulmonary TB at the time of presentation at a health-care facility that is superior to physicians' opinion. DESIGN: Nonconcurrent prospective study. SETTING: University-affiliated hospital. PARTICIPANTS: A derivation group of 563 isolation episodes and a validation group of 119 isolation episodes. INTERVENTIONS: A general regression neural network (GRNN) was used to develop the predictive model. MEASUREMENTS: Predictive accuracy of the neural network compared with clinicians' assessment. RESULTS: Predictive accuracy was assessed by the c-index, which is equivalent to the area under the receiver operating characteristic curve. The GRNN significantly outperformed the physicians' prediction, with calculated c-indices (+/- SEM) of 0.947 +/- 0.028 and 0.61 +/- 0.045, respectively (p < 0.001). When the GRNN was applied to the validation group, the corresponding c-indices were 0. 923 +/- 0.056 and 0.716 +/- 0.095, respectively. CONCLUSION: An artificial neural network can identify patients with active pulmonary TB more accurately than physicians' clinical assessment. PMID- 10531162 TI - A multicenter study of grepafloxacin and clarithromycin in the treatment of patients with community-acquired pneumonia. AB - STUDY OBJECTIVES: To compare the efficacies of 10-day regimens of grepafloxacin (GFX) (Raxar or Vaxar; Glaxo Wellcome; Greenford, UK), 600 qd, and clarithromycin (CLA) (Klacid, Biaxin, or Klaracid; Abbott Laboratories; Chicago, IL), 500 mg bid, in patients with community-acquired pneumonia (CAP), on the basis of clinical response, including radiographic evidence, and bacteriologic efficacy. DESIGN: Phase IIIb, double-blind, double-dummy, randomized, prospective, parallel group, comparative study conducted at 58 centers in 11 countries. PATIENTS AND SETTING: Adult patients with signs and symptoms of CAP that was confirmed by radiographic evidence and who did not require parenteral therapy were included in the study. ASSESSMENTS: Patients were assessed before treatment, during treatment, after treatment, and at follow-up (28 to 35 days after treatment completion). Clinical response was evaluated. Blood and sputum samples were cultured for bacterial pathogens, and serology testing was performed to detect atypical pneumonia. RESULTS: A total of 504 patients were enrolled into the trial: 251 were randomly assigned to receive GFX and 253 to receive CLA. In patients able to be clinically evaluated, clinical success rates at follow-up were 147 of 163 patients (90%) in the GFX group and 148 of 167 patients (89%) in the CLA group (95% confidence interval, -6% to 9%). Pretreatment pathogens were confirmed in 131 of 504 patients (26%), either by culture or serology testing, the primary pathogens being Streptococcus pneumoniae (22%), Haemophilus influenzae (17%), Mycoplasma pneumoniae (25%), and Chlamydia pneumoniae (11%). For patients able to be evaluated who had typical pathogens, bacteriologic success was achieved in 92% of the patients in each treatment group. For patients able to be evaluated who had atypical pathogens, 18 of 18 patients (100%) in the GFX and 23 of 26 patients (88%) in the CLA group had a successful clinical outcome. There were similar rates of adverse events in each group, resulting in /= 30) and in the less obese group (BMI < 30). Similarly, in the severe (RDI >/= 40) and less severe groups (RDI < 40), as well as in both age groups (< and > 60 years of age), the op-nCPAP was significantly higher in the Sup posture than it was in the Lat posture. Irrespective of the four parameters mentioned, the actual differences in op-nCPAP between the two body postures were almost identical, ranging between 2.31 and 2.66 cm H(2)O. CONCLUSIONS: For most OSA patients, the op-nCPAP level is significantly higher in the Sup position than it is in the Lat position. This is true for REM and NREM sleep, for obese and nonobese patients, for patients with different degrees of severity, and for young and old OSA patients. Since the op-nCPAP was highest in the Sup posture during REM sleep, no nCPAP titration should be considered complete without the patient having slept in the Sup posture during REM sleep. PMID- 10531166 TI - Exhaled carbon monoxide levels elevated in diabetes and correlated with glucose concentration in blood: a new test for monitoring the disease? AB - PURPOSE: In diabetes, the interaction of glycated proteins with their cell surface binding sites leads to oxidative stress and induction of the stress protein heme oxygenase (HO)-1. Considering that carbon monoxide (CO) is a product of HO activity, we studied the level of exhaled CO as a marker of oxidative stress in diabetes. METHODS: Eight patients with insulin-dependent diabetes mellitus (type 1) (4 men, 4 women; [mean +/- SEM] age, 50 +/- 8 years) were studied, of whom 2 had peripheral neuropathy and 1 had renal failure. Sixteen patients with non-insulin-dependent diabetes mellitus (type 2) (5 men, 11 women; age 63 +/- 8 years) were studied, of whom 2 had peripheral neuropathy. Glycosylated hemoglobin (HbA(1)c) levels were higher (7.4 +/- 0.3%) in patients with type 1 (mean duration of the disease, 20 +/- 5 years) than in type 2 (4.9 +/ 0.4%; p < 0.05; mean duration of the disease, 11 +/- 2 years). All of the patients were lifelong nonsmokers. RESULTS: Levels of exhaled CO were higher in patients with diabetes (type 1, 4.0 +/- 0.7 ppm; type 2, 5.0 +/- 0.4 ppm) when compared to 37 nonsmoking healthy subjects (20 men, 17 women; age, 33 +/- 3 years) (2.9 +/- 0.2 ppm; p < 0.05). There was a positive correlation between exhaled CO levels and the incidence of glycemia in all subjects (r = 0.52, p < 0.05) and the duration of diabetes (r = 0.48, p < 0.05), but there was not a strong correlation with concentrations of HbA(1)c (r = 0.06, p = 0.8). In addition, an oral glucose tolerance test was performed in five healthy nonsmoking volunteers (three men; age, 33 +/- 4 years). The maximal glucose increase (from 3.9 +/- 0.2 to 5.5 +/- 0.1 mmol/L at 15 min; p < 0.05) was associated with a significant increase in exhaled CO concentration (from 3.0 +/- 0.5 to 6.3 +/- 1.0 ppm; p < 0. 05). Both parameters returned to the baseline at 40 min after glucose administration. CONCLUSIONS: Elevated levels of exhaled CO in diabetes may reflect HO-1 induction and oxidative stress. The measurement of CO may be a new tool for disease monitoring. PMID- 10531167 TI - Mechanical ventilation in hematopoietic stem cell transplantation: can We effectively predict outcomes? AB - BACKGROUND: Survival rates from mechanical ventilation (MV) in allogeneic bone marrow transplantation are poor, but little is known about the need for and outcomes from MV in patients who undergo autologous hematopoietic stem cell transplantation (AHSCT). STUDY OBJECTIVE: To determine the frequency of and risk factors for the use of MV in recipients of AHSCT and to identify predictors of survival in mechanically ventilated AHSCT patients. DESIGN: Retrospective, cohort analysis SETTING: Tertiary-care, university-affiliated medical center. PATIENTS: One hundred fifty-nine consecutive patients who underwent AHSCT. INTERVENTIONS: Patient surveillance and data collection. MEASUREMENTS AND RESULTS: The primary outcome measure was the need for MV, and the secondary end point was survival after MV. Of 159 patients, 17 required MV (10. 7%). Three variables were associated with the need for MV: increasing age, use of total body irradiation in the conditioning regimen, and treatment with amphotericin B. As a screening test to predict the need for MV, no risk factor had a sensitivity or specificity > 82%. Three of the 17 mechanically ventilated patients (17.6%) survived to discharge. Only the mean APACHE (acute physiology and chronic health evaluation) II score separated survivors from nonsurvivors (21.7 vs 31.4; p = 0.029). Both the duration of MV and the length of stay in the ICU were similar in survivors and nonsurvivors. CONCLUSIONS: We conclude that MV is infrequently needed following AHSCT. Although survival after MV in these patients is limited, clinical variables do not reliably allow clinicians to prospectively identify patients destined to die. PMID- 10531168 TI - Differences in the response times of pages originating from the ICU. AB - STUDY OBJECTIVES: To determine whether the type of paging system causes significant differences in the response time by physicians to their pages in an ICU setting. DESIGN AND SETTING: Prospective cohort study performed in the ICU of two university-affiliated hospitals. All pages were classified by several different variables, including the type of paging system: direct paging if a nurse or hospital operator could directly place the page, or indirect paging if a nurse or hospital operator was required to contact the physician's office or a private answering service who would then independently contact the physician. The main outcome measure was physicians' response time, in minutes, to pages originating from the ICU. RESULTS: During a 100-day period, 402 pages were sent and answered by 166 different physicians (87 attending physicians and 79 housestaff/physician assistants). The median response time for all pages was 3 min with a 25 to 75% quartile of 1 to 8 min. Twenty-five percent of the pages placed through an indirect system were associated with a response time of >/= 29 min. In a multivariate model with the response time dichotomized at >/= 15 min ("slow") or < 15 min ("adequate"), pages placed through an indirect system were answered significantly more slowly than pages placed through a direct system (p < 0.001; odds ratio, 4.36; 95% confidence interval, 2.05 to 9.29). Pages answered in an adequate amount of time were also associated with a significantly higher degree of overall nursing satisfaction with the care delivered by the physician in response to the specific page when compared with pages answered in a "slow" manner (p < 0.001). CONCLUSIONS: Physicians who use an indirect paging system are significantly slower in their response to ICU pages when compared with physicians who utilize a direct paging system. These results may lead to improvements in paging systems used by physicians who care for patients in an ICU setting. PMID- 10531169 TI - Reengineering respiratory support following extubation: avoidance of critical care unit costs. AB - STUDY OBJECTIVE: We prospectively investigated alternative clinical practice strategies for critically ill trauma patients following extubation to evaluate the cost-effectiveness of these maneuvers. The primary change was elimination of the routine use of postextubation supplemental oxygen, with concurrent utilization of noninvasive positive pressure ventilatory support (NPPV) to manage occurrences of postextubation hypoxemia. DESIGN: Prospective, consecutive accrual of patients undergoing extubation. SETTING: Trauma ICU in a university hospital. INTERVENTIONS AND MEASUREMENTS: All patients received mechanical ventilation using pressure support ventilation (PSV) with continuous positive airway pressure (CPAP) as the primary mode. The patients were extubated to room air following a 20-min preextubation trial of 5 cm H(2)O CPAP at FIO(2) of 0.21, and demonstrating a spontaneous respiratory rate /= 7.30, PaCO(2) /= 50 mm Hg. The subgroup of patients who became hypoxemic (pulse oximetric saturation < 88%) within 24 h of extubation were treated with NPPV for up to 48 h duration. Patients who failed NPPV were reintubated. Four hundred fifty-one (84%) patients were successfully extubated to room air. Seventy-two patients (13%) became hypoxemic within 24 h, and NPPV was administered. Fifty-two patients (72% of those who were hypoxemic) responded to NPPV, while 20 patients failed to respond to therapy, were reintubated, and received mechanical ventilation for a mean of 4 days. Thirteen additional patients (2%) were reintubated for reasons other than hypoxemia. The overall reintubation rate for the group (n = 536) was 6.2%; for the postextubation hypoxemic group who failed NPPV, the reintubation rate was 3.7%. The elimination of routine supplemental oxygen via nasal cannula following extubation resulted in a potential direct cost avoidance of $50,006.88 for 451 patient days. Moreover, the 52 patients who were spared reintubation and mechanical ventilation provided an additional potential cost avoidance of $19,740.24 in unused ventilator days per patient. CONCLUSION: Eliminating the routine use of supplemental oxygen and employing NPPV as a method to prevent reintubation can facilitate a more aggressive, cost-effective strategy for the management of the trauma ICU patient who has been extubated. PMID- 10531170 TI - Evaluation of the cuff-leak test in a cardiac surgery population. AB - OBJECTIVE: The purpose of this study is to determine the accuracy of the cuff leak test in a cardiovascular ICU. METHODS: Five hundred twenty-four patients were tested immediately before being extubated 531 times. The cuff-leak test was performed with the ventilator set in assist-control mode at a tidal volume (VT) of 10 to 12 mL/kg. The leak was taken to be the difference between the preset inspiratory VT and the average of the three lowest of the subsequent six expiratory VTs. A leak 72 h who had new or progressive lung infiltrate plus at least two of three clinical criteria for VAP. INTERVENTIONS: BAL and blood culture performed within 24 h of establishing a clinical diagnosis of VAP. MEASUREMENTS AND RESULTS: Ninety patients were BAL positive (BAL+), satisfying a microbiological definition of VAP (>/= 10(4) cfu/mL), 72 patients were BAL negative (BAL-). Bacteremia was diagnosed when at least two sets of blood cultures yielded a microorganism or when only one set was positive, but the same bacteria was present at a concentration >/= 10(4) cfu/mL in the BAL fluid. Bacteremia was significantly more frequent in the BAL+ than in the BAL- group (22/90 patients vs 5/72 patients; p = 0.006). In 6 of 22 BAL+ patients with bacteremia, an extrapulmonary site of infection was the source of bacteremia. Sensitivity of blood culture for disclosing the pathogenic microorganism in BAL+ patients was 26%, and the positive predictive value to detect the pathogen was 73%. Factors associated with mortality were age > 50 years, simplified acute physiology score > 14, prior inadequate antibiotic therapy, PaO(2)/fraction of inspired oxygen < 205, and use of H(2) blockers. By multivariate analysis, only the use of prior inadequate antimicrobial therapy (odds ratio [OR], 6.47) and age > 50 years (OR, 5.12) were independently associated with higher mortality. The rate of complications was not different in patients with bacteremia. CONCLUSIONS: Blood cultures have a low sensitivity for detecting the same pathogenic microorganism as BAL culture in patients with VAP. The presence of bacteremia does not predict complications, it is not related to the length of stay, and it does not identify patients with more severe illness. Inadequacy of prior antimicrobial therapy and age > 50 years were the only factors associated with mortality in a multivariate analysis. Blood cultures in patients with VAP are clearly useful if there is suspicion of another probable infectious condition, but the isolation of a microorganism in the blood does not confirm that microorganism as the pathogen causing VAP. PMID- 10531179 TI - A 51-year-old man with fever, ulnar neuropathy, and bilateral pleural effusions. Lupus pleuritis. PMID- 10531178 TI - Dyspnea differentiation index: A new method for the rapid separation of cardiac vs pulmonary dyspnea. AB - STUDY OBJECTIVE: To assess the utility of a new parameter in the differentiation of dyspnea of cardiac origin from dyspnea of pulmonary origin. METHODS: The peak expiratory flow (PEF) rate and the partial pressure of oxygen in arterial blood (PaO(2)) were measured in 71 patients with the chief complaint of dyspnea. The patients were treated in the hospital, and the final diagnosis (cardiac or pulmonary) of the cause of dyspnea was made at discharge. We defined a new measure, the dyspnea differentiation index (DDI), as (PEF x PaO(2))/1,000. We performed a receiver operating characteristic (ROC) curve analysis of the data to define the measure that best distinguished cardiac from pulmonary dyspnea. The curves also allowed us to establish an optimal cut-off point to distinguish between cardiac and pulmonary dyspnea. RESULTS: Patients with pulmonary dyspnea had a significantly lower mean PEF than patients with cardiac dyspnea (144 +/- 66 vs 267 +/- 97 L/min, respectively; p < 0.001). They also had a lower DDI than patients with cardiac dyspnea (8.4 +/- 4.0 vs 18.4 +/- 7.9 L-mm/min, respectively; p < 0.001). These two measures, PEF and DDI, also best distinguished pulmonary from cardiac dyspnea. PEF was able to diagnose the correct cause of dyspnea in 72% of patients, and DDI was correct in 79% of patients. This compares favorably to the performance of the emergency department physicians, who were able to predict the correct diagnosis in only 69% of patients. CONCLUSION: These results demonstrate that the PEF by itself is useful in differentiating between cardiac and pulmonary causes of dyspnea, but that the calculation of DDI is superior in this regard. PMID- 10531180 TI - Nd-YAG laser vs bronchoscopic electrocautery for palliation of symptomatic airway obstruction: a cost-effectiveness study. AB - STUDY OBJECTIVE: To evaluate the cost effectiveness of the Nd-YAG laser and bronchoscopic electrocautery for palliation in patients with symptomatic tumor obstruction. DESIGN: A retrospective study. SETTING: Bronchoscopy unit of a university hospital. PATIENTS AND INTERVENTION: Thirty-one consecutive patients with inoperable non-small cell lung cancer and symptomatic intraluminal tumor underwent bronchoscopic treatment. Dyspnea relief was the primary goal of treatment. Fourteen patients were treated with the Nd-YAG laser and 17 patients with electrocautery. MEASUREMENTS AND RESULTS: Improvement of symptoms was achieved in 70% of patients treated by either Nd-YAG laser or electrocautery. Mean +/- SD survival was 8.0 +/- 2.5 months after Nd-YAG laser treatment and 11.5 +/- 3.5 months after electrocautery. The number of treatment sessions per patient was comparable: Nd-YAG laser, 1.1; electrocautery, 1.2. Duration of hospital stay was longer in patients treated with the Nd-YAG laser (8.4 vs 6.7 days). Average treatment costs, including admission charges, were $5,321 for the Nd-YAG laser and $4,290 for electrocautery. Higher costs in the group treated with the Nd-YAG laser were caused by a longer hospital stay before bronchoscopic treatment. Costs of equipment (electrocautery $6,701 and Nd-YAG laser $208,333), write-offs, maintenance, and repair were not included in this calculation. CONCLUSION: Bronchoscopic electrocautery is equally effective but is a less expensive and, in our hospital, a more accessible modality than the Nd-YAG laser for symptomatic palliation of patients with intraluminal airway obstruction. PMID- 10531181 TI - An HIV-infected patient with pleural effusion. PMID- 10531182 TI - Multiple coronary aneurysms in a case of systemic lupus erythematosus. AB - A 29-year-old woman with active systemic lupus erythematosus (SLE) sustained an anterior myocardial infarction and demonstrated angiographic evidence of multiple, diffuse coronary aneurysms. Coronary artery aneurysms have been reported in 11 prior cases of patients with SLE. A Medline search of the literature revealed no prior reports of extensive aneurysmal dilatations involving all three main coronary arteries (left anterior descending, left circumflex, and right coronary arteries). PMID- 10531183 TI - Video-assisted thoracic surgical non-rib spreading simultaneously stapled lobectomy: a more patient-friendly oncologic resection. AB - OBJECTIVE: To evaluate the outcomes from a new surgical technique for lobectomy. PATIENTS: Two hundred fifty consecutive patients with an average age of 67.3 years underwent simultaneously stapled lobectomy. METHODS: Video-assisted thoracic surgical non-rib spreading lobectomy (VNSSL) is a new technique that has been evolving for approximately 6.5 years. During 1990, we began using video assisted thoracic surgery (VATS) for simple, benign diseases. Throughout 1991, VATS was applied to malignant problems, ie, mediastinal masses, staging of lymph nodes, malignant effusions, and coin lesions. As experience was acquired, more complex procedures were attempted, such as lobectomy. On September 9, 1991, our first VATS lobectomy, using anatomic hilar dissection and lymph node sampling, was performed for primary carcinoma of the lung. One year later, we performed our first VNSSL using simultaneous stapling. RESULTS: Currently, 400 VNSSLs have been performed. In this entire series, there have been no surgical mortality, bronchopleural fistulas, port implantations, or transfusions. Bronchial stumps have averaged 4 mm in length, and all have been microscopically negative for neoplasm. In order to evaluate long-term survival for primary carcinoma of the lung in patients with an adequate duration of follow-up, the first 250 consecutive VNSSLs have been reviewed. There were 120 male and 130 female patients ranging in age from 20 to 92 years old who had 62 right upper lobe, 20 right middle lobe, 58 right lower lobe, 63 left upper lobe, and 33 left lower lobe lobectomies, and 14 bilobectomies. The lesions consisted of 214 primary carcinomas, 8 metastatic lesions, and 28 benign problems. Seven to 18 lymph nodes could be resected during staging of the primary neoplasms. The tumors ranged in size from 1 to 9 cm, and operating times averaged 78.6 min. Hospitalization averaged 2.83 days. No patient was admitted to the ICU. Intensive monitoring or narcotic analgesia were not needed. No epidural or intercostal anesthesia was used. Complications were infrequent and minor. Most patients returned to preoperative levels of physical activity within 7 to 10 days. Overall survival at a mean of 34 months, when all stages of neoplasms were combined, is 83%. For stage I, overall survival is 92%. The cost of VNSSL is approximately 50% less than the traditional open thoracotomy. CONCLUSIONS: VNSSL is an oncologic technique that has been clinically rewarding and economically beneficial for patients with malignant lesions. Long-term survival for primary carcinoma currently exceeds reports being published for the traditional open thoracotomy. Scientific reasons for this extraordinary survival are emerging. Complications, surgical mortality, pain, and length of stay have all been reduced. Patient recovery, comfort, and satisfaction have been extraordinary. PMID- 10531184 TI - Tracheal bronchus associated with lung cancer: a case report. AB - Tracheal bronchus is a rarely found congenital bronchial anomaly. It usually originates from the right lateral wall of the trachea at the level < 2 cm above the tracheal bifurcation. The patients usually are asymptomatic, but some may experience recurrent pneumonia, chronic bronchitis, or bronchiectasis. It is very rare for a malignant tumor to grow from this aberrant bronchus. There are only four cases of lung cancer developing from the tracheal bronchus reported in the world literature, and we present a fifth case. PMID- 10531185 TI - Continuous intravenous epoprostenol therapy for pulmonary hypertension in Gaucher's disease. AB - Gaucher's disease is a rare disorder characterized by a deficiency of lysosomal beta-glucosidase. Pulmonary hypertension, the etiology of which is unclear, has been reported to occur in association with Gaucher's disease. We report the use of continuous intravenous epoprostenol (prostacyclin), which has been used to treat other forms of pulmonary hypertension, in a patient with pulmonary hypertension associated with Gaucher's disease. Although its mechanism of action remains unknown, epoprostenol may be an effective form of therapy for chronic pulmonary hypertension due to a variety of conditions, one of which is Gaucher's disease. PMID- 10531186 TI - Serial scintigraphic assessment of iodine-123 metaiodobenzylguanidine lung uptake in a patient with high-altitude pulmonary edema. AB - Iodine-123 metaiodobenzylguanidine ((123)I-MIBG) can be considered an indicator of pulmonary endothelial cell function. Serial (123)I-MIBG images of the chest were acquired in a patient with high altitude pulmonary edema (HAPE). The initial evaluation was performed 7 days after admission. The lung to upper mediastinum ratios (LMRs) of (123)I-MIBG uptake were 1.33 (for the right lung) and 1.12 (for the left lung). The second examination of (123)I-MIBG lung uptake, which was performed 2 months later, showed LMRs of 1.39 (right lung) and 1.33 (left lung). We speculated that the decreased lung uptake of (123)I-MIBG at the early recovery stage could reflect an impairment in pulmonary endothelial cell metabolic function in the development of HAPE. PMID- 10531187 TI - Contralateral tension pneumothorax following unilateral chest tube drainage of bilateral pneumothoraces in a heart-lung transplant patient. AB - Bilateral pneumothoraces can result from unilateral air leak after heart-lung transplantation. The recommended initial management of such patients is insertion of a unilateral chest tube. We report a patient who developed bilateral pneumothoraces after undergoing transbronchial biopsies 2 years after a heart lung transplant. The unilateral chest tube failed to drain the contralateral pneumothorax and a tension pneumothorax developed. The advocated approach should be used with caution. PMID- 10531188 TI - Right pneumothorax resulting from an endocardial screw-in atrial lead. AB - Right pneumothorax complicated by an endocardial atrial lead has never been reported. Herein, we report on a small-build 79-year-old Taiwanese woman who suffered from complete AV block and underwent dual-chamber permanent pacemaker implantation. An active fixation screw-in atrial lead was chosen. The procedure was complicated by right pneumothorax associated with atrial perforation. Since simple measurements of the implantation parameters could not be used to predict the occurrence of perforation, great caution should be taken in to avoid overscrewing the atrial lead, and in scrutinizing the penetration depth of the helix of the lead under fluoroscopy. PMID- 10531189 TI - Advanced heart block as a manifestation of a paraneoplastic syndrome from malignant thymoma. AB - Malignant thymoma is a rare tumor that is associated with paraneoplastic syndrome. Myocarditis as a paraneoplastic syndrome has been rarely described. Reported herein is a young male patient with malignant thymoma and myocarditis as part of a paraneoplastic syndrome. This resulted in high-degree heart block and an asystolic cardiac arrest despite placement of a permanent pacemaker. PMID- 10531190 TI - Apples and oranges : flaws and guffaws. PMID- 10531191 TI - Percutaneous tracheostomy is really better-if done correctly. PMID- 10531192 TI - Pneumonitis and herbicide exposure. PMID- 10531193 TI - Bias and precision in noninvasive monitoring. PMID- 10531194 TI - Exercise limitation testing. PMID- 10531195 TI - Tuberculous mycotic aneurysms. PMID- 10531196 TI - Air sampling for tuberculosis- homage to the lowly guinea pig. PMID- 10531197 TI - Oxygen delivery in septic shock. PMID- 10531198 TI - Carrier properties of a protein derived from outer membrane protein A of Klebsiella pneumoniae. AB - We have recently cloned a new protein, recombinant P40 (rP40). When tested in vivo after conjugation to a B-cell epitope, rP40 induces an important antibody response without the need for adjuvant. To characterize its potency, this carrier protein was coupled to a peptide derived from respiratory syncytial virus attachment G protein (G1'). After immunization of mice with the rP40-G1' conjugate, strong antipeptide antibodies were detected, whereas peptide alone was not immunogenic. To emphasize the carrier properties of rP40, a polysaccharide derived from Haemophilus influenzae type b (Hib) was coupled to it. Immunoglobulin G responses against the Hib polysaccharide were observed after coupling to rP40. Interestingly, an antipeptide antibody response was observed despite preexisting anti-rP40 antibodies generated by preimmunization with rP40. In addition, rP40 compares well with the reference carrier protein, tetanus toxoid (TT), since antibody responses of equal intensity were observed when a peptide or a polysaccharide was coupled to TT and rP40. Moreover, rP40 had advantages compared to TT; e.g., it induced a mixed Th1/Th2 response, whereas TT induced only a Th2 profile. Together, the results indicate that rP40 is a novel carrier protein with potential for use as an alternative carrier for human vaccination. PMID- 10531199 TI - Differential T-cell recognition of native and recombinant Mycobacterium tuberculosis GroES. AB - Mycobacterium tuberculosis GroES was purified from culture filtrate, and its identity was confirmed by immunoblot analysis and N-terminal sequencing. Comparing the immunological recognition of native and recombinant GroES, we found that whereas native GroES elicited a strong proliferative response and release of gamma interferon-gamma by peripheral blood mononuclear cells from healthy tuberculin reactors, the recombinant protein failed to do so. The same difference in immunological recognition was observed in a mouse model of TB infection. Both the native and recombinant preparations were recognized by mice immunized with the recombinant protein. Biochemical characterization including sodium dodecyl sulfate-polyacrylamide gel electrophoresis, two-dimensional electrophoresis, and mass spectrometry analysis of both proteins demonstrated no differences between the native and recombinant forms of GroES except for the eight additional N terminal amino acids derived from the fusion partner in recombinant GroES. The recombinant fusion protein, still tagged with the maltose binding protein, was recognized by T cells isolated from TB-infected mice if mixed with culture filtrate before affinity purification on an amylose column. The maltose binding protein treated in the same manner as a control preparation was not recognized. Based on the data presented, we suggest that the association of biologically active molecules from culture filtrate with the chaperone GroES may be responsible for the observed T-cell recognition of the native preparation. PMID- 10531200 TI - Control of Leishmania infantum infection is associated with CD8(+) and gamma interferon- and interleukin-5-producing CD4(+) antigen-specific T cells. AB - Visceral leishmaniasis is a severe and lethal disease caused by the protozoan parasites of the genus Leishmania. In areas where leishmaniasis is endemic, most infected individuals control the infection and remain asymptomatic; chemotherapy of visceral leishmaniasis restores some immunity which protects against relapses. In the present study, Leishmania-specific T-cell clones were established from six asymptomatic and five cured patients. Cytokines production by these clones was analyzed. A large fraction of the parasite-specific T-cell clones from asymptomatic patients were CD8(+) and produced high amounts of gamma interferon (IFN-gamma). Most CD4(+) T-cell clones from two asymptomatic subjects exhibited an unusual phenotype: production of high levels of IFN-gamma low levels of interleukin-4, (IL-4), but high levels of IL-5. In contrast, only few parasite specific CD8(+) T-cell clones were obtained from cured patients after chemotherapy; moreover, CD4(+) T-cell clones from these patients exhibited an heterogeneous profile of cytokines from Th1-like to Th2-like phenotypes. These results point to CD8(+) T cells and to IL-5- and IFN-gamma-producing CD4(+) T cells as possible contributors to human resistance to Leishmania infection. They should stimulate new immunological approaches in the control of this disease. PMID- 10531201 TI - Deglycosylation of the 45/47-kilodalton antigen complex of Mycobacterium tuberculosis decreases its capacity to elicit in vivo or in vitro cellular immune responses. AB - A protection against a challenge with Mycobacterium tuberculosis is induced by previous immunization with living attenuated mycobacteria, usually bacillus Calmette-Guerin (BCG). The 45/47-kDa antigen complex (Apa) present in culture filtrates of BCG of M. tuberculosis has been identified and isolated based on its ability to interact mainly with T lymphocytes and/or antibodies induced by immunization with living bacteria. The protein is glycosylated. A large batch of Apa was purified from M. tuberculosis culture filtrate to determine the extent of glycosylation and its role on the expression of the immune responses. Mass spectrometry revealed a spectrum of glycosylated molecules, with the majority of species bearing six, seven, or eight mannose residues (22, 24, and 17%, respectively), while others three, four, or five mannoses (5, 9, and 14%, respectively). Molecules with one, two, or nine mannoses were rare (1.5, 3, and 3%, respectively), as were unglycosylated species (in the range of 1%). To eliminate the mannose residues linked to the protein, the glycosylated Apa molecules were chemically or enzymatically treated. The deglycosylated antigen was 10-fold less active than native molecules in eliciting delayed-type hypersensitivity reactions in guinea pigs immunized with BCG. It was 30-fold less active than native molecules when assayed in vitro for its capacity to stimulate T lymphocytes primed in vivo. The presence of the mannose residues on the Apa protein was essential for the antigenicity of the molecules in T-cell-dependent immune responses in vitro and in vivo. PMID- 10531202 TI - Differentiation of monocytes to macrophages primes cells for lipopolysaccharide stimulation via accumulation of cytoplasmic nuclear factor kappaB. AB - During infection, circulating blood monocytes migrate from the vasculature to the extravascular compartments where they mature into tissue macrophages. The maturation process prepares the cell to actively participate in the inflammatory and the immune responses, and many transcription factors have been found to be involved. Here we report on a novel role for nuclear factor kappaB (NF-kappaB) in this process. Its accumulation in the cytoplasm of differentiated macrophages is responsible for the enhanced ability of the cell to respond to lipopolysaccharide (LPS) stimulation, as determined by tumor necrosis factor alpha (TNF-alpha) secretion. Differentiation of the human monocytic cell line THP-1 into macrophage like cells was induced by exposure of the cells to phorbol myristate acetate. DNA bindable NF-kappaB was not detected in the cytoplasm of undifferentiated THP-1 cells but accumulated in the cytoplasm of the cells following differentiation. No TNF-alpha was detected in the media of resting differentiated and nondifferentiated THP-1 cells. Stimulation with LPS of differentiated cells induced the production of higher levels of TNF-alpha than stimulation of nondifferentiated cells. This hyperresponsiveness to LPS was found in the mRNA and secreted TNF-alpha levels. Furthermore, stimulation with LPS induced the translocation of NF-kappaB from the cytoplasm into the nucleus. This translocation process was more rapid in the differentiated cells than in the nondifferentiated cells, and the resultant accumulated levels of NF-kappaB in the nucleus were higher. The DNA-bindable NF-kappaB was identified as a heterodimer of p65 and p50. The results suggest that NF-kappaB accumulation in the cytoplasm during maturation of monocytes to macrophages primes the cells for enhanced responsiveness to LPS and results in the rapid secretion of inflammatory mediators, such as TNF-alpha, by mature macrophages following LPS challenge. PMID- 10531203 TI - The endogenous balance of soluble tumor necrosis factor receptors and tumor necrosis factor modulates cachexia and mortality in mice acutely infected with Trypanosoma cruzi. AB - To better understand the role of tumor necrosis factor (TNF) during Trypanosoma cruzi infection in BALB/c mice, we have investigated the kinetics of circulating tumor necrosis factor (TNF), soluble TNF receptor 1 (sTNR1), and sTNFR2 levels, as well as the interactions between such factors, in relation to parasitemia, cachexia, and mortality of acutely infected animals. Our data show that the parasitemic phase of T. cruzi infection in mice is associated with high levels of circulating TNF and sTNFR2, resulting in the formation of cytokine-receptor complexes and some degree of neutralization of TNF bioactivity. Although sTNR2 levels always exceeded TNF levels, low sTNFR/TNF circulating ratios were associated with cachexia in all infected mice, whereas the lowest ratios were observed in dying animals harboring the highest parasitemia. We also studied the modulation of sTNFR/TNF ratios induced by anti-TNF antibodies administered to infected animals and their consequences on the outcome of the infection. The injection of anti-TNF monoclonal antibody (MAb) TN3 into infected mice resulted in a paradoxical overproduction of TNF (associated with a higher parasitemia), lowered the sTNFR/TNF circulating ratios, and considerably worsened cachexia and mortality of animals. Another anti-TNF MAb (1F3F3) decreased the in vivo availability of TNF as well as parasite levels and reduced cachexia. Altogether, such results highlight that, besides playing a beneficial role early in infection, TNF also triggers harmful effects in the parasitemic phase, which are limited by the in vivo simultaneous endogenous production of soluble receptors. PMID- 10531205 TI - Dichotomy of cytokine profiles in patients and high-risk healthy subjects exposed to tuberculosis. AB - To better understand the role of cytokines in susceptible and resistant subjects exposed to Mycobacterium tuberculosis infection, intracellular gamma interferon (IFN-gamma) and interleukin-4 (IL-4) in ex vivo peripheral blood-derived CD4(+) T cells were examined by flow cytometry. Of the 37 individuals examined, 20 had clinical evidence of pulmonary tuberculosis and showed acid-fast bacilli in the sputum. Other individuals in close contact with these patients showed no evidence of disease. Patients had a higher number of CD4(+) T cells expressing IFN-gamma and IL-4 in unstimulated cultures compared to healthy subjects. Despite this, the ratio of IFN-gamma(+) to IL-4(+) CD4(+) T cells was similar in both groups. The Th1 response seen in CD4(+) T cells in patients was also observed in the overall pattern of IFN-gamma and IL-4 detected in control culture supernatants by enzyme linked immunosorbent assay (ELISA). However, after in vitro stimulation of PBMC with heat-killed M. tuberculosis there was a significant reduction in the percentage of IFN-gamma(+) CD4(+) T cells (P < 0.001) in patients. This trend was reflected in the IFN-gamma ELISA assay with supernatants derived from stimulated cultures. However, the accumulated levels of IFN-gamma were higher than those for IL-4. The reduction of IFN-gamma(+) CD4(+) T cells resulted in the dominance of IL-4(+) CD4(+) T cells in 13 patients (P < 0.05). The elevated levels of IL-4(+) CD4(+) T cells seen in patients may contribute to the downregulation of IFN-gamma expression and the crucial effector function of CD4 T cells, leading to the persistence of disease and the immunopathology characteristically seen in patients. Preliminary data on the indicators of apoptosis in antigen-stimulated cultures in PBMC derived from patients are presented. Of the 17 high-risk healthy individuals examined, 11 differed in that, after mycobacterial-antigen stimulation, there was an enhancement in IFN-gamma(+) CD4(+) T cells. PMID- 10531204 TI - Characterization of pic, a secreted protease of Shigella flexneri and enteroaggregative Escherichia coli. AB - We have identified and characterized a secreted protein, designated Pic, which is encoded on the chromosomes of enteroaggregative Escherichia coli (EAEC) 042 and Shigella flexneri 2457T. The product of the pic gene is synthesized as a 146.5 kDa precursor molecule which is processed at the N and C termini during secretion, allowing the release of a mature protein (109.8 kDa) into the culture supernatant. The deduced amino acid sequence of Pic shows high homology to autotransporter proteins, particularly a subgroup termed the SPATEs (serine protease autotransporters of the Enterobacteriaceae). Present in all members of this subgroup is a motif similar to the active sites of certain serine proteases. Pic catalyzes gelatin degradation, which can be abolished by disruption of the predicted proteolytic active site. Functional analysis of the Pic protein implicates this factor in mucinase activity, serum resistance, and hemagglutination. Our data suggest that Pic may be a multifunctional protein involved in enteric pathogenesis. PMID- 10531206 TI - CD4(+) T-cell- and gamma interferon-dependent protection against murine malaria by immunization with linear synthetic peptides from a Plasmodium yoelii 17 kilodalton hepatocyte erythrocyte protein. AB - Most work on protective immunity against the pre-erythrocytic stages of malaria has focused on induction of antibodies that prevent sporozoite invasion of hepatocytes, and CD8(+) T-cell responses that eliminate infected hepatocytes. We recently reported that immunization of A/J mice with an 18-amino-acid synthetic linear peptide from Plasmodium yoelii sporozoite surface protein 2 (SSP2) in TiterMax adjuvant induces sterile protection that is dependent on CD4(+) T cells and gamma interferon (IFN-gamma). We now report that immunization of inbred A/J mice and outbred CD1 mice with each of two linear synthetic peptides from the 17 kDa P. yoelii hepatocyte erythrocyte protein (HEP17) in the same adjuvant also induces protection against sporozoite challenge that is dependent on CD4(+) T cells and IFN-gamma. The SSP2 peptide and the two HEP17 peptides are recognized by B cells as well as T cells, and the protection induced by these peptides appears to be directed against the infected hepatocytes. In contrast to the peptide-induced protection, immunization of eight different strains of mice with radiation-attenuated sporozoites induces protection that is absolutely dependent on CD8(+) T cells. Data represented here demonstrate that CD4(+) T-cell-dependent protection can be induced by immunization with linear synthetic peptides. These studies therefore provide the foundation for an approach to pre-erythrocytic stage malaria vaccine development, based on the induction of protective CD4(+) T cell responses, which will complement efforts to induce protective antibody and CD8(+) T-cell responses. PMID- 10531207 TI - Interleukin-1 inhibits gamma interferon-induced bacteriostasis in human uroepithelial cells. AB - The most prominent gamma interferon (IFN-gamma)-induced antimicrobial effector mechanisms are the induction of nitric oxide (NO) synthase (NOS) and of indoleamine 2,3-dioxygenase (IDO) activity. We have recently found that human glioblastoma cells and human macrophages inhibit the growth of group B streptococci after stimulation with IFN-gamma. In this report, we show that in addition, human RT4 (uroepithelial) cells can inhibit the growth of enterococci. Murine macrophages (RAW cells) are unable to inhibit bacterial growth after IFN gamma stimulation. Stimulation of human glioblastoma cells, macrophages, and RT4 cells with human IFN-gamma results in a strong expression of IDO activity; however, NO production remains undetectable. In strong contrast, murine RAW cells produce large amounts of NO when stimulated with murine IFN-gamma and IDO activity is not detectable. Interleukin-1 (IL-1) induces NO synthase in human RT4 cells when the cells are costimulated with IFN-gamma. We found that IL-1 inhibits IFN-gamma-stimulated IDO activity and antimicrobial effects in RT4 cells, while in human glioblastoma cells, which lack detectable NO synthase activity, neither of these effects was altered by costimulation with IFN-gamma and IL-1. The IL-1 mediated inhibition of IDO activity and of subsequent antibacterial effect is due to the production of NO. This conclusion was supported by evidence that N(G) monomethyl-L-arginine, a competitive inhibitor of inducible NOS activity, is able to block the inhibitory action of IL-1 on IFN-gamma-induced bacteriostasis. We therefore conclude that NO production does not inhibit the growth of enterococci but might be involved in the regulation of IDO activity in some human cells. PMID- 10531208 TI - Identification by subtractive hybridization of a novel insertion sequence specific for virulent strains of Porphyromonas gingivalis. AB - Subtractive hybridization was employed to isolate specific genes from virulent Porphyromonas gingivalis strains that are possibly related to abscess formation. The genomic DNA from the virulent strain P. gingivalis W83 was subtracted with DNA from the avirulent strain ATCC 33277. Three clones unique to strain W83 were isolated and sequenced. The cloned DNA fragments were 885, 369, and 132 bp and had slight homology with only Bacillus stearothermophilus IS5377, which is a putative transposase. The regions flanking the cloned DNA fragments were isolated and sequenced, and the gene structure around the clones was revealed. These three clones were located side-by-side in a gene reported as an outer membrane protein. The three clones interrupt the open reading frame of the outer membrane protein gene. This inserted DNA, consisting of three isolated clones, was designated IS1598, which was 1,396 bp (i.e., a 1,158-bp open reading frame) in length and was flanked by 16-bp terminal inverted repeats and a 9-bp duplicated target sequence. IS1598 was detected in P. gingivalis W83, W50, and FDC 381 by Southern hybridization. All three P. gingivalis strains have been shown to possess abscess forming ability in animal models. However, IS1598 was not detected in avirulent strains of P. gingivalis, including ATCC 33277. The IS1598 may interrupt the synthesis of the outer membrane protein, resulting in changes in the structure of the bacterial outer membrane. The IS1598 isolated in this study is a novel insertion element which might be a specific marker for virulent P. gingivalis strains. PMID- 10531210 TI - Identification of a Clostridium perfringens enterotoxin region required for large complex formation and cytotoxicity by random mutagenesis. AB - Clostridium perfringens enterotoxin (CPE), a single polypeptide of 319 amino acids, has a unique multistep mechanism of action. In the first step, CPE binds to claudin proteins and/or a 50-kDa eukaryotic membrane protein receptor, forming a small ( approximately 90-kDa) complex. This small complex apparently then associates with a 70-kDa eukaryotic membrane protein, resulting in formation of a large complex that induces the onset of membrane permeability alterations. To better define the boundaries of CPE functional regions and to identify specific amino acid residues involved in various steps of CPE action, in this study we subjected the cloned cpe gene to random mutagenesis in XL-1 Red strains of Escherichia coli. Seven CPE random mutants with reduced cytotoxicity for Vero cells were phenotypically characterized for the ability to complete each step in CPE action. Five of these seven recombinant CPE (rCPE) random mutants (G49D, S59L, R116S, R137G, and S167P) exhibited binding characteristics similar to those of rCPE or native CPE, while the Y310C and W226Stop mutants showed reduced binding and no binding, respectively, to brush border membranes. Interestingly, two completely nontoxic mutants (G49D and S59L) were able to bind and form small complex but they did not mediate any detectable large complex formation. Another strongly attenuated mutant, R116S, formed reduced amounts of an anomalously migrating large complex. Collectively, these results provide further support for large complex formation being an essential step in CPE action and also identify the CPE region ranging from residues approximately 45 to 116 as important for large complex formation. Finally, we also report that limited removal of extreme N-terminal CPE sequences, which may occur in vivo during disease, stimulates cytotoxic activity by enhancing large complex formation. PMID- 10531209 TI - Endothelial adhesion molecule expression and its inhibition by recombinant bactericidal/permeability-increasing protein are influenced by the capsulation and lipooligosaccharide structure of Neisseria meningitidis. AB - Vascular endothelial injury is responsible for many of the clinical manifestations of severe meningococcal disease. Binding and migration of activated host inflammatory cells is a central process in vascular damage. The expression and function of adhesion molecules regulate interactions between leukocytes and endothelial cells. Little is known about how meningococci directly influence these receptors. In this study we have explored the effect of Neisseria meningitidis on endothelial adhesion molecule expression and found this organism to be a potent inducer of the adhesion molecules CD62E, ICAM-1, and VCAM-1. Exposure of endothelium to a serogroup B strain of Neisseria meningitidis, B1940, and a range of isogenic mutants revealed that lipooligosaccharide (LOS) structure and capsulation influence the expression of adhesion molecules. Following only a brief exposure (15 min) to the bacteria, there were large differences in the capacity of the different mutants to induce vascular cell adhesion molecules, with the unencapsulated and truncated LOS strains being most potent (P < 0.05). Furthermore, the pattern of cell adhesion molecule expression was different with purified endotoxin from that with intact bacteria. Meningococci were more potent stimuli of CD62E expression than was endotoxin, whereas endotoxin was at least as effective as meningococci in inducing ICAM-1 and VCAM-1. The effect of bactericidal/permeability increasing protein (rBPI(21)), an antibacterial molecule with antiendotoxin properties, was also dependent on LOS structure. The strains which possessed a truncated or nonsialylated LOS, whether capsulated or not, were more sensitive to the inhibitory effects of rBPI(21). These findings could have important implications for the use of antiendotoxin therapy in meningococcal disease. PMID- 10531211 TI - Temporal sequence of pulmonary and systemic inflammatory responses to graded polymicrobial peritonitis in mice. AB - The lungs are the remote organ most commonly affected in human peritonitis. The major goals of this study were to define the dose- and time-dependent relationship between graded septic peritonitis and systemic and pulmonary inflammatory responses in mice. BALB/c mice were treated with intraperitoneal polymicrobial inoculi and sacrificed at 3, 12, and 24 h. The treatment protocol resulted in distinct groups of animals with respect to mortality rate, kinetics, and concentrations of a broad spectrum of pro- and anti-inflammatory endogenous mediators, intrapulmonary bacterial accumulation, and static lung compliance. In sublethally infected mice, pulmonary bacterial proliferation was controlled. Levels of monocyte chemoattractant protein-1 (MCP-1), interleukin-10, interleukin 6, granulocyte colony-stimulating factor (G-CSF), and tumor necrosis factor (TNF) in plasma were elevated 3 h after infection exclusively. At 3 h, MCP-1, gamma interferon, and TNF were detected in extracts of pulmonary tissue or in bronchoalveolar lavage (BAL) fluid. Static lung compliance (C(st)) was transiently decreased at 12 h. In contrast, in lethally infected mice pulmonary bacterial proliferation was not contained. Concentrations of MCP-1, G-CSF, and TNF in plasma were maximal at 24 h, as were pulmonary MCP-1 levels. Lung myeloperoxidase activity was increased at 3, 12, and 24 h. C(st) was reduced after 3 h and did not reach control values at 24 h. Pulmonary cyclooxygenase-2 mRNA and eicosanoids in BAL fluid and plasma were elevated at 3 and 24 h. This study shows that polymicrobial peritonitis in mice leads to dose-dependent systemic and pulmonary inflammation accompanied by a decrease in lung compliance. PMID- 10531212 TI - Analysis of virulence of clinical isolates of Salmonella enteritidis in vivo and in vitro. AB - Salmonella enterica serotype Enteritidis (S. enteritidis) is a major food-borne pathogen, and its incidence among all Salmonella serotypes has increased dramatically in the last two decades. To study the virulence characteristics of clinical isolates of S. enteritidis, we determined the 50% lethal doses (LD(50)) in mice of isolates of two major phage types (4 and 8). Isolates of both phage types showed a wide range of LD(50) after oral inoculation, varying from under 10(2) organisms to over 10(8) organisms. No significant difference in LD(50) was observed between the phage types. These observations indicated that clinical isolates of S. enteritidis are highly heterogeneous in their ability to cause death in mice. We compared the LD(50)s of these isolates to the results observed from in vitro pathogenicity assays. We also analyzed these isolates for recognized Salmonella virulence loci (spv, sodCI, sopE, and sef). The in vitro phenotypes of the isolates showed no obvious correlation with their LD(50) in any given assay, and the virulence genes tested were present in all isolates. However, the isolate with the lowest LD(50) (isolate 97A 2472) was resistant to acidified sodium nitrite (ASN). Moreover, the most acid-susceptible, macrophage susceptible, and ASN-susceptible isolates were attenuated for virulence in mice. These results, based on extensive analysis of clinical isolates of S. enteritidis, demonstrate the complex nature of Salmonella pathogenesis in mice. Our results also indicate the limitation of in vitro assays in predicting in vivo virulence. PMID- 10531213 TI - Repeated administration of synthetic oligodeoxynucleotides expressing CpG motifs provides long-term protection against bacterial infection. AB - Synthetic oligodeoxynucleotides (ODN) expressing unmethylated CpG motifs stimulate an innate immune response characterized by the production of polyreactive immunoglobulin M antibodies and immunomodulatory cytokines. This immune response has been shown to protect mice from challenge by Listeria monocytogenes and Francisella tularensis for up to 2 weeks. By repeatedly administering CpG ODN two to four times/month, we found that this protection could be maintained indefinitely. Protection was associated with a significant increase in the number of spleen cells that could be triggered by subsequent pathogen exposure to secrete gamma interferon and interleukin-6 in vivo (P < 0.01). ODN-treated animals remained healthy and developed neither macroscopic nor microscopic evidence of tissue damage or inflammation. Thus, repeated administration of CpG ODN may provide a safe means of conferring long-term protection against infectious pathogens. PMID- 10531214 TI - Differences in surface expression of NspA among Neisseria meningitidis group B strains. AB - NspA is a highly conserved membrane protein that is reported to elicit protective antibody responses against Neisseria meningitidis serogroups A, B and C in mice (D. Martin, N. Cadieux, J. Hanel, and B. R. Brodeur, J. Exp. Med. 185:1173-1183, 1997). To investigate the vaccine potential of NspA, we produced mouse anti recombinant NspA (rNspA) antisera, which were used to evaluate the accessibility of NspA epitopes on the surface of different serogroup B strains by an immunofluorescence flow cytometric assay and by susceptibility to antibody dependent, complement-mediated bacteriolysis. Among 17 genetically diverse strains tested, 11 (65%) were positive for NspA cell surface epitopes and 6 (35%) were negative. All six negative strains also were resistant to bactericidal activity induced by the anti-rNspA antiserum. In contrast, of the 11 NspA surface positive strains, 8 (73%; P < 0.05) were killed by the antiserum and complement. In infant rats challenged with one of these eight strains, the anti-rNspA antiserum conferred protection against bacteremia, whereas the antiserum failed to protect rats challenged by one of the six NspA cell surface-negative strains. Neither NspA expression nor protein sequence accounted for differences in NspA surface accessibility, since all six negative strains expressed NspA in outer membrane preparations and since their predicted NspA amino acid sequences were 99 to 100% identical to those of three representative positive strains. However, the six NspA cell surface-negative strains produced, on average, larger amounts of group B polysaccharide than did the 11 positive strains (reciprocal geometric mean titers, 676 and 224, respectively; P < 0.05), which suggests that the capsule may limit the accessibility of NspA surface epitopes. Given these strain differences in NspA surface accessibility, an rNspA-based meningococcal B vaccine may have to be supplemented by additional antigens. PMID- 10531215 TI - Expression and characterization of the Mycobacterium tuberculosis serine/threonine protein kinase PknB. AB - PknB is a member of the newly discovered eukaryotic-like protein serine/threonine kinase (PSTK) family of proteins. The pknB gene was cloned and expressed in Escherichia coli. The active recombinant protein was purified and shown to be reactive with antiphosphoserine antibodies, as well as with antibodies to the phosphorylated eukaryotic Ser/Thr kinases mitogen-activated protein kinase kinase 3 and 6, P38, and Creb. In vitro kinase assays demonstrated that PknB is a functional kinase that is autophosphorylated on serine/threonine residues and is also able to phosphorylate the peptide substrate myelin basic protein. Analysis of pknB expression in Mycobacterium tuberculosis indicates the presence of pknB mRNA in (i) organisms grown in vitro in bacteriological media, (ii) a murine macrophage in vitro infection model, and (iii) in vivo alveolar macrophages from a patient with tuberculosis. PMID- 10531216 TI - Identification of promiscuous epitopes from the Mycobacterial 65-kilodalton heat shock protein recognized by human CD4(+) T cells of the Mycobacterium leprae memory repertoire. AB - By using a synthetic peptide approach, we mapped epitopes from the mycobacterial 65-kDa heat shock protein (HSP65) recognized by human T cells belonging to the Mycobacterium leprae memory repertoire. A panel of HSP65 reactive CD4(+) T-cell lines and clones were established from healthy donors 8 years after immunization with heat-killed M. leprae and then tested for proliferative reactivity against overlapping peptides comprising both the M. leprae and Mycobacterium tuberculosis HSP65 sequences. The results showed that the antigen-specific T-cell lines and clones established responded to 12 mycobacterial HSP65 peptides, of which 9 peptides represented epitopes crossreactive between the M. tuberculosis and M. leprae HSP65 (amino acids [aa] 61 to 75, 141 to 155, 151 to 165, 331 to 345, 371 to 385, 411 to 425, 431 to 445, 441 to 455, and 501 to 515) and 3 peptides (aa 343 to 355, 417 to 429, and 522 to 534) represented M. leprae HSP65-specific epitopes. Major histocompatibility complex restriction analysis showed that presentation of 9 of the 12 peptides to T cells were restricted by one of the 2 HLA-DR molecules expressed from self HLA-DRB1 genes, whereas 3 peptides with sequences completely identical between the M. leprae and M. tuberculosis HSP65 were presented to T cells by multiple HLA-DR molecules: peptide (aa 61 to 75) was presented by HLA-DR1, -DR2, and -DR7, peptide (aa 141 to 155) was presented by HLA-DR2, -DR7, and -DR53, whereas both HLA-DR2 and -DR4 (Dw4 and Dw14) were able to present peptide (aa 501 to 515) to T cells. In addition, the T-cell lines responding to these peptides in proliferation assays showed cytotoxic activity against autologous monocytes/macrophages pulsed with the same HSP65 peptides. In conclusion, we demonstrated that promiscuous peptide epitopes from the mycobacterial HSP65 antigen can serve as targets for cytotoxic CD4(+) T cells which belong to the human memory T-cell repertoire against M. leprae. The results suggest that such epitopes might be used in the peptide-based design of subunit vaccines against mycobacterial diseases. PMID- 10531217 TI - Salmonella typhimurium virulence genes are induced upon bacterial invasion into phagocytic and nonphagocytic cells. AB - Survival and growth of salmonellae within host cells are important aspects of bacterial virulence. We have developed an assay to identify Salmonella typhimurium genes that are induced inside Salmonella-containing vacuoles within macrophage and epithelial cells. A promoterless luciferase gene cassette was inserted randomly into the Salmonella chromosome, and the resulting mutants were screened for genes upregulated in intracellular bacteria compared to extracellular bacteria. We identified four genes in S. typhimurium that were upregulated upon bacterial invasion of both phagocytic and nonphagocytic cells. Expression of these genes was not induced by factors secreted by host cells or media alone. All four genes were induced at early time points (2 to 4 h) postinvasion and continued to be upregulated within host cells at later times (5 to 7 h). One mutant contained an insertion in the ssaR gene, within Salmonella pathogenicity island 2 (SPI-2), which abolished bacterial virulence in a murine typhoid model. Two other mutants contained insertions within SPI-5, one in the sopB/sigD gene and the other in a downstream gene, pipB. The insertions within SPI-5 resulted in the attenuation of S. typhimurium in the mouse model. The fourth mutant contained an insertion within a previously undescribed region of the S. typhimurium chromosome, iicA (induced intracellularly A). We detected no effect on virulence as a result of this insertion. In conclusion, all but one of the genes identified in this study were virulence factors within pathogenicity islands, illustrating the requirement for specific gene expression inside mammalian cells and indicating the key role that virulence factor regulation plays in Salmonella pathogenesis. PMID- 10531218 TI - Experimental gonococcal genital tract infection and opacity protein expression in estradiol-treated mice. AB - The development of effective prophylactic agents against gonorrhea and the study of adaptation by Neisseria gonorrhoeae to the urogenital mucosa are hindered by the lack of a well-established animal model of gonococcal genital tract infection. Here, a murine model of long-term gonococcal genital tract infection is described. Female BALB/c mice were treated with 17-beta-estradiol and inoculated intravaginally with wild-type gonococcal strain FA1090 or MS11. N. gonorrhoeae was recovered from vaginal swabs for an average of 12 to 13 days following inoculation with 10(6) CFU of either strain. Inflammation occurred in over 80% of infected mice, and diplococci were associated with epithelial cells and neutrophils in stained vaginal smears. Ascended infection occurred in 17 to 20% of mice inoculated with strain FA1090. An outbred mouse strain (SLC:ddY) previously reported to be naturally susceptible to N. gonorrhoeae was also tested; however, as with BALB/c mice, estradiol was required for prolonged infection. Although piliation was not maintained during experimental murine infection, 46 to 100% of vaginal isolates from four of eight BALB/c mice and three of four SLC:ddY mice expressed one or more opacity (Opa) proteins within 4 days after inoculation with an Opa-negative variant of strain FA1090. The observed selection for and/or induction of gonococcal Opa protein expression during murine infection appears to parallel events that occur during experimental urethritis in volunteers. PMID- 10531220 TI - Monoclonal antibody to Shiga toxin 2 which blocks receptor binding and neutralizes cytotoxicity. AB - A monoclonal antibody (MAb) was raised against Shiga toxin 2 (Stx2) of Escherichia coli O157:H7. MAb VTm1.1 belonged to the immunoglobulin G1 subclass and had a kappa light chain, and it could neutralize the cytotoxic activity of Stx2 and variants derived from patient strains but not that of variants derived from animals. MAb VTm1.1 was shown to bind to the B subunit of these neutralized Stx2s by Western blotting. Comparison of B-subunit amino acid sequences and reactivities to these Stxs suggested six amino acids (Ser30, Ser53, Glu56, Gln65, Asn68, and Asp69) that were candidates for the MAb VTm1.1 epitope. Consequently, five Stx2 mutants (S30N, S53N, E56H, Q65K, and N68Ter) were prepared by site directed mutagenesis to determine which residue is essential for the epitope. All of these mutants showed cytotoxicity almost equal to that of the wild-type Stx2. Of the five Stx2 mutants, only E56H could not be neutralized by MAb VTm1.1. Western blot analysis also showed that MAb VTm1.1 could not bind to the E56H B subunit. These results indicated that Glu56 is an important residue recognized by MAb VTm1. 1. Immunofluorescence analysis further indicated that MAb VTm1.1 inhibits the binding of Stx2 to its receptors. MAb VTm1.1 could be a useful therapeutic agent for Shiga toxin-producing E. coli infection. PMID- 10531219 TI - Population dynamics of a naturally occurring heterogeneous mixture of Borrelia burgdorferi clones. AB - Two unique isolates of Borrelia burgdorferi, differing in plasmid content and outer surface protein C expression, were cultured on sequential captures of a single free-living Peromyscus leucopus mouse and were examined for differences in transmissibility. Both isolates were transmissible from inoculated C.B-17 mice to larval Ixodes scapularis ticks and, subsequently, from infected nymphal ticks to C3H/HeJ mice. Plasmid and protein analyses suggested that the original isolates were a mixed population of B. burgdorferi, and cloning by limiting dilution resulted in the identification of two clonal groups. In addition to being heterogeneous in plasmid and genomic macrorestriction analyses, the clones varied with respect to the electrophoretic mobilities and antigenicity of their OspC proteins, as shown by their reactivity to a panel of monoclonal antibodies. Plasmid analysis of sequential isolates from C3H mice experimentally infected with the primary isolate or various mixtures of its subclones showed an apparently random fluctuation in clonal dominance in the majority of mice. Surprisingly, mice infected with each subclone were permissive to superinfection with the heterologous subclone, despite the presence of anti-B. burgdorferi antibodies at the time of the secondary challenge. These results show conclusively that mice captured at Lyme disease enzootic sites may be infected by mixed populations of genetically and antigenically distinct B. burgdorferi clones and that these infections can be acquired by coinfection or by sequential infection. The lack of cross-immunization between clones existing within a naturally occurring population may play a role in the maintenance of the genetic heterogeneity of B. burgdorferi in nature. PMID- 10531221 TI - Lysogenic conversion of environmental Vibrio mimicus strains by CTXPhi. AB - The filamentous bacteriophage CTXPhi, which encodes cholera toxin (CT) in toxigenic Vibrio cholerae, is known to propagate by infecting susceptible strains of V. cholerae by using the toxin coregulated pilus (TCP) as its receptor and thereby causing the origination of new strains of toxigenic V. cholerae from nontoxigenic progenitors. Besides V. cholerae, Vibrio mimicus strains which are normally TCP negative have also been shown to occasionally produce CT and cause diarrhea in humans. We analyzed nontoxigenic V. mimicus strains isolated from surface waters in Bangladesh for susceptibility and lysogenic conversion by CTXPhi and studied the expression of CT in the lysogens by using genetically marked derivatives of the phage. Of 27 V. mimicus strains analyzed, which were all negative for genes encoding TCP but positive for the regulatory gene toxR, 2 strains (7.4%) were infected by CTX-KmPhi, derived from strain SM44(P27459 ctx::km), and the phage genome integrated into the host chromosome, forming stable lysogens. The lysogens spontaneously produced infectious phage particles in the supernatant fluids of the culture, and high titers of the phage could be achieved when the lysogens were induced with mitomycin C. This is the first demonstration of lysogenic conversion of V. mimicus strains by CTXPhi. When a genetically marked derivative of the replicative form of the CTXPhi genome carrying a functional ctxAB operon, pMSF9.2, was introduced into nontoxigenic V. mimicus strains, the plasmid integrated into the host genome and the strains produced CT both in vitro and inside the intestines of adult rabbits and caused mild-to-severe diarrhea in rabbits. This suggested that in the natural habitat infection of nontoxigenic V. mimicus strains by wild-type CTXPhi may lead to the origination of toxigenic V. mimicus strains which are capable of producing biologically active CT. The results of this study also supported the existence of a TCP-independent mechanism for infection by CTXPhi and showed that at least one species of Vibrio other than V. cholerae may contribute to the propagation of the phage. PMID- 10531222 TI - Interaction of Mycobacterium tuberculosis-induced transforming growth factor beta1 and interleukin-10. AB - Mycobacterium tuberculosis is associated with the activation of cytokine circuits both at sites of active tuberculosis in vivo and in cultures of mononuclear cells stimulated by M. tuberculosis or its components in vitro. Interactive stimulatory and/or inhibitory pathways are established between cytokines, which may result in potentiation or attenuation of the effects of each molecule on T-cell responses. Here we examined the interaction of transforming growth factor beta1 (TGF-beta1) and interleukin-10 (IL-10) in purified protein derivative (PPD)-stimulated human mononuclear cell cultures in vitro. TGF-beta1 induced monocyte IL-10 (but not tumor necrosis factor alpha) production (by 70-fold, P < 0.02) and mRNA expression in the absence but not in the presence of PPD. Both exogenous recombinant (r) IL-10 and rTGF-beta1 independently suppressed the production of PPD-induced gamma interferon (IFN-gamma) in mononuclear cells from PPD skin test positive individuals. Synergistic suppression of IFN-gamma in cultures containing both rTGF-beta1 and rIL-10 was only seen when the responder cell population were peripheral blood mononuclear cells (PBMC) and not monocyte-depleted mononuclear cells and when PBMC were pretreated with rTGF-beta1 but not with rIL-10. Suppression of PPD-induced IFN-gamma in PBMC containing both rTGF-beta1 (1 ng/ml) and rIL-10 (100 pg/ml) was 1.5-fold higher (P < 0.05) than cultures containing TGF-beta1 alone and 5.7-fold higher (P < 0.004) than cultures containing IL-10 alone. Also, neutralization of endogenous TGF-beta1 and IL-10 together enhanced PPD-induced IFN-gamma in PBMC in a synergistic manner. Thus, TGF-beta1 and IL-10 together potentiate the downmodulatory effect on M. tuberculosis-induced T-cell production of IFN-gamma, and TGF-beta1 alone enhances IL-10 production. At sites of active M. tuberculosis infection, these interactions may be conducive to the suppression of mononuclear cell functions. PMID- 10531223 TI - Lack of humoral immune protection against Treponema denticola virulence in a murine model. AB - This study investigated the characteristics of humoral immune responses to Treponema denticola following primary infection, reinfection, and active immunization, as well as immune protection in mice. Primary infection with T. denticola induced a significant (400-fold) serum immunoglobulin G (IgG) response compared to that in control uninfected mice. The IgG response to reinfection was 20, 000-fold higher than that for control mice and 10-fold higher than that for primary infection. Mice actively immunized with formalin-killed treponemes developed serum antibody levels seven- to eightfold greater than those in animals after primary infection. Nevertheless, mice with this acquired antibody following primary infection or active immunization demonstrated no significant alterations of lesion induction or decreased size of the abscesses following a challenge infection. Mice with primary infection developed increased levels of IgG3, IgG2b, and IgG2a antibodies, with IgG1 being lower than the other subclasses. Reinfected mice developed enhanced IgG2b, IgG2a, and IgG3 and less IgG1. In contrast, immunized mice developed higher IgG1 and lower IgG3 antibody responses to infection. These IgG subclass distributions indicate a stimulation of both Th1 and Th2 activities in development of the humoral immune response to infection and immunization. Our findings also demonstrated a broad antigen reactivity of the serum antibody, which was significantly increased with reinfection and active immunization. Furthermore, serum antibody was effective in vitro in immobilizing and clumping the bacteria but did not inhibit growth or passively prevent the treponemal infection. These observations suggest that humoral immune responses, as manifested by antibody levels, isotype, and antigenic specificity, were not capable of resolving a T. denticola infection. PMID- 10531224 TI - Interleukin-6 and interleukin-12 participate in induction of a type 1 protective T-cell response during vaccination with a tuberculosis subunit vaccine. AB - We examined the role of cytokines in the development of gamma interferon (IFN gamma)-secreting protective T cells following immunization with a culture filtrate subunit vaccine against Mycobacterium tuberculosis containing the adjuvant dimethyldioctadecylammonium bromide (DDA). Depletion of either interleukin-6 (IL-6) or IL-12 with specific neutralizing antibodies during vaccination reduced the priming of T cells for antigen-specific proliferation and IFN-gamma secretion. Such reduction was also observed in IL-6 gene-disrupted mice as compared to wild-type animals. IL-6 was found to play a role in the initial differentiation of Th1 cells but not in their expansion. The defect found after IL-6 depletion or in IL-6-knockout mice was compensated by the inclusion of recombinant mouse IL-12 in the vaccine. The induction of protective immunity against an intravenous or an aerosol challenge with live, virulent M. tuberculosis was markedly reduced by neutralizing either IL-6 or IL-12 during immunization with the vaccine. Likewise, the effects of IL-6 neutralization were partially reversed by including IL-12 in the vaccine. Our data point to an important role of IL-6 and IL-12 in the generation of cell-mediated immunity to tuberculosis. PMID- 10531225 TI - Lysine residue 117 of the FasG adhesin of enterotoxigenic Escherichia coli is essential for binding of 987P fimbriae to sulfatide. AB - The FasG subunit of the 987P fimbriae of enterotoxigenic strains of Escherichia coli was previously shown to mediate fimbrial binding to a glycoprotein and a sulfatide receptor on intestinal brush borders of piglets. Moreover, the 987P adhesin FasG is required for fimbrial expression, since fasG null mutants are nonfimbriated. In this study, fasG was modified by site-directed mutagenesis to study its sulfatide binding properties. Twenty single mutants were generated by replacing positively charged lysine (K) or arginine (R) residues with small, nonpolar alanine (A) residues. Reduced levels of binding to sulfatide-containing liposomes correlated with reduced fimbriation and FasG surface display in four fasG mutants (R27A, R286A, R226A, and R368). Among the 16 remaining normally fimbriated mutants with wild-type levels of surface-exposed FasG, only one mutant (K117A) did not interact at all with sulfatide-containing liposomes. Four mutants (K117A, R116A, K118A, and R200A) demonstrated reduced binding to such liposomes. Since complete phenotypic dissociation between the structure and specific function of 987P was observed only with mutant K117A, this residue is proposed to play an essential role in the FasG-sulfatide interaction, possibly communicating with the sulfate group of sulfatide by hydrogen bonding and/or salt bridge formation. Residues K17, R116, K118, and R200 may stabilize this interaction. PMID- 10531226 TI - Upregulation of p75 tumor necrosis factor alpha receptor in Mycobacterium avium infected mice: evidence for a functional role. AB - The bacterial growth and the production of tumor necrosis factor alpha (TNF alpha) and TNF receptors (TNF-Rs) in the spleen and blood of BALB/c mice challenged with Mycobacterium avium complex (MAC) were monitored. Infection developed in two phases: the first, up to day 21, was associated with rapid MAC multiplication in the spleen and a drop in the mycobacteremia, and the second was associated with control of the infection in both compartments. In the spleen, TNF alpha and TNF-RII mRNA levels peaked on day 21 and then slowly decreased; however, no increase in the level of TNF-RI mRNA was observed throughout these experiments. The level of circulating soluble TNF-RII (sTNF-RII) was transiently increased after day 21. In a model in which overproduction of bioactive TNF-alpha was triggered in response to a second infection with MAC, an increased production of sTNF-RII by cultured splenocytes was also observed. Administration of an antagonist anti-TNF-RII monoclonal antibody (MAb 6G1) to infected mice inhibited the bacterial growth in the spleen, suggesting that the TNF-RII and/or sTNF-RII was functionally involved in the mechanisms that control the infection. Overall, these observations suggest that upregulation of TNF-RII or sTNF-RII contributes to modulation of the TNF-alpha antibacterial activity in MAC infections. PMID- 10531227 TI - Genomic analysis reveals variation between Mycobacterium tuberculosis H37Rv and the attenuated M. tuberculosis H37Ra strain. AB - Mycobacterium tuberculosis H37Ra is an attenuated tubercle bacillus closely related to the virulent type strain M. tuberculosis H37Rv. Despite extensive study, the reason for the decreased virulence of M. tuberculosis H37Ra has not been determined. A genomic approach was therefore initiated to identify genetic differences between M. tuberculosis H37Rv and M. tuberculosis H37Ra as a means of pinpointing the attenuating mutation(s). Digestion with the rare-cutting restriction endonuclease DraI revealed two polymorphisms between the strains: a 480-kb fragment in M. tuberculosis H37Rv was replaced by two fragments of 220 and 260 kb in M. tuberculosis H37Ra, while there was a approximately 7.9-kb DraI fragment in M. tuberculosis H37Ra that had no counterpart in M. tuberculosis H37Rv. As the M. tuberculosis insertion sequence IS6110 contains a single DraI restriction site, it was considered possible that these polymorphisms were the result of IS6110 transposition events in M. tuberculosis H37Ra, events that may have inactivated virulence genes. The 7.9-kb polymorphism was found to be due to the presence of the previously described H37Rv RvD2 deletion in M. tuberculosis H37Ra, with sequence analysis suggesting an IS6110-mediated deletion mechanism for loss of RvD2. Three other IS6110-catalyzed deletions from the M. tuberculosis H37Rv chromosome (RvD3 to RvD5) were also identified, suggesting that this mechanism plays an important role in genome plasticity in the tubercle bacilli. Comparative mapping and sequencing revealed that the 480-kb polymorphism was due to an IS6110 insertion in M. tuberculosis H37Ra near oriC. Complementation of M. tuberculosis H37Ra with a 2.9-kb restriction fragment from M. tuberculosis H37Rv that encompassed the IS6110 insertion did not increase the survival of recombinant M. tuberculosis H37Ra in mice. In conclusion, this study describes the presence and mechanisms of genomic variation between M. tuberculosis H37Ra and M. tuberculosis H37Rv, although the role that they play in the attenuation of M. tuberculosis H37Ra is unclear. PMID- 10531228 TI - Outer membrane protein A-promoted actin condensation of brain microvascular endothelial cells is required for Escherichia coli invasion. AB - Escherichia coli is the most common gram-negative bacterium that causes meningitis during the neonatal period. We have previously shown that the entry of circulating E. coli organisms into the central nervous system is due to their ability to invade the blood-brain barrier, which is composed of a layer of brain microvascular endothelial cells (BMEC). In this report, we show by transmission electron microscopy that E. coli transmigrates through BMEC in an enclosed vacuole without intracellular multiplication. The microfilament-disrupting agents cytochalasin D and latrunculin A completely blocked E. coli invasion of BMEC. Cells treated with the microtubule inhibitors nocodazole, colchicine, vincristin, and vinblastine and the microtubule-stabilizing agent taxol also exhibited 50 to 60% inhibition of E. coli invasion. Confocal laser scanning fluorescence microscopy showed F-actin condensation associated with the invasive E. coli but no alterations in microtubule distribution. These results suggest that E. coli uses a microfilament-dependent phagocytosis-like endocytic mechanism for invasion of BMEC. Previously we showed that OmpA expression significantly enhances the E. coli invasion of BMEC. We therefore examined whether OmpA expression is related to the recruitment of F-actin. OmpA(+) E. coli induced the accumulation of actin in BMEC to a level similar to that induced by the parental strain, whereas OmpA( ) E. coli did not. Despite the presence of OmpA, a noninvasive E. coli isolate, however, did not show F-actin condensation. OmpA(+)-E. coli-associated condensation of F-actin was blocked by synthetic peptides corresponding to the N terminal extracellular domains of OmpA as well as BMEC receptor analogues for OmpA, chitooligomers (GlcNAcbeta1-4GlcNAc oligomers). These findings suggest that OmpA interaction is critical for the expression or modulation of other bacterial proteins that will subsequently cause actin accumulation for the uptake of bacteria. PMID- 10531229 TI - Plasmodium falciparum field isolates commonly use erythrocyte invasion pathways that are independent of sialic acid residues of glycophorin A. AB - Erythrocyte invasion by malaria parasites is mediated by specific molecular interactions. Sialic acid residues of glycophorin A are used as invasion receptors by Plasmodium falciparum. In vitro invasion studies have demonstrated that some cloned P. falciparum lines can use alternate receptors independent of sialic acid residues of glycophorin A. It is not known if invasion by alternate pathways occurs commonly in the field. In this study, we used in vitro growth assays and erythrocyte invasion assays to determine the invasion phenotypes of 15 P. falciparum field isolates. Of the 15 field isolates tested, 5 multiply in both neuraminidase and trypsin-treated erythrocytes, 3 multiply in neuraminidase treated but not trypsin-treated erythrocytes, and 4 multiply in trypsin-treated but not neuraminidase-treated erythrocytes; 12 of the 15 field isolates tested use alternate invasion pathways that are not dependent on sialic acid residues of glycophorin A. Alternate invasion pathways are thus commonly used by P. falciparum field isolates. Typing based on two polymorphic markers, MSP-1 and MSP 2, and two microsatellite markers suggests that only 1 of the 15 field isolates tested contains multiple parasite genotypes. Individual P. falciparum lines can thus use multiple invasion pathways in the field. These observations have important implications for malaria vaccine development efforts based on EBA-175, the P. falciparum protein that binds sialic acid residues of glycophorin A during invasion. It may be necessary to target parasite ligands responsible for the alternate invasion pathways in addition to EBA-175 to effectively block erythrocyte invasion by P. falciparum. PMID- 10531230 TI - Invasion of human coronary artery cells by periodontal pathogens. AB - There is an emerging paradigm shift from coronary heart disease having a purely hereditary and nutritional causation to possibly having an infectious etiology. Recent epidemiological studies have shown a correlation between periodontal disease and coronary heart disease. However, to date, there is minimal information as to the possible disease mechanisms of this association. It is our hypothesis that invasion of the coronary artery cells by oral bacteria may start and/or exacerbate the inflammatory response in atherosclerosis. Since a few periodontal pathogens have been reported to invade oral epithelial tissues, we tested the ability of three putative periodontal pathogens-Eikenella corrodens, Porphyromonas gingivalis, and Prevotella intermedia-to invade human coronary artery endothelial cells and coronary artery smooth muscle cells. In this study we demonstrate by an antibiotic protection assay and electron microscopy that specific species and strains invade coronary artery cells at a significant level. Actin polymerization and eukaryotic protein synthesis in metabolically active cells were required since the corresponding inhibitors nearly abrogated invasion. Many intracellular P. gingivalis organisms were seen to be present in multimembranous vacuoles resembling autophagosomes by morphological analysis. This is the first report of oral microorganisms invading human primary cell cultures of the vasculature. PMID- 10531231 TI - Evaluation of a truncated recombinant flagellin subunit vaccine against Campylobacter jejuni. AB - A recombinant protein comprising the maltose-binding protein (MBP) of Escherichia coli fused to amino acids 5 to 337 of the FlaA flagellin of Campylobacter coli VC167 was evaluated for immunogenicity and protective efficacy against challenge by a heterologous strain of campylobacter, Campylobacter jejuni 81-176, in two murine models. The sequence of the flaA gene of strain 81-176 revealed a predicted protein which was 98.1% similar to that of VC167 FlaA over the region expressed in the fusion protein. Mice were immunized intranasally with two doses of 3 to 50 microgram of MBP-FlaA, given 8 days apart, with or without 5 microgram of the mutant E. coli heat-labile enterotoxin (LT(R192G)) as a mucosal adjuvant. The full range of MBP-FlaA doses were effective in eliciting antigen-specific serum immunoglobulin G (IgG) responses, and these responses were enhanced by adjuvant use, except in the highest dosing group. Stimulation of FlaA-specific intestinal secretory IgA (sIgA) responses required immunization with higher doses of MBP-FlaA (>/=25 microgram) or coadministration of lower doses with the adjuvant. When vaccinated mice were challenged intranasally 26 days after immunization, the best protection was seen in animals given 50 microgram of MBP FlaA plus LT(R192G). The protective efficacies of this dose against disease symptoms and intestinal colonization were 81.1 and 84%, respectively. When mice which had been immunized with 50 microgram of MBP-FlaA plus LT(R192G) intranasally were challenged orally with 8 x 10(10), 8 x 10(9), or 8 x 10(8) cells of strain 81-176, the protective efficacies against intestinal colonization at 7 days postinfection were 71.4, 71.4, and 100%, respectively. PMID- 10531232 TI - Safety and immunogenicity of Vi conjugate vaccines for typhoid fever in adults, teenagers, and 2- to 4-year-old children in Vietnam. AB - The capsular polysaccharide of Salmonella typhi, Vi, is an essential virulence factor and a protective vaccine for people older than 5 years. The safety and immunogenicity of two investigational Vi conjugate vaccines were evaluated in adults, 5- to 14-year-old children, and 2- to 4-year-old children in Vietnam. The conjugates were prepared with Pseudomonas aeruginosa recombinant exoprotein A (rEPA) as the carrier, using either N-succinimidyl-3-(2-pyridyldithio)-propionate (SPDP; Vi-rEPA(1)) or adipic acid dihydrazide (ADH; Vi-rEPA(2)) as linkers. None of the recipients experienced a temperature of >38.5 degrees C or significant local reactions. One injection of Vi-rEPA(2) into adults elicited a geometric mean (GM) increase in anti-Vi immunoglobulin G (IgG) from 9.62 enzyme-linked immunosorbent assay units/ml (EU) to 465 EU at 6 weeks; this level fell to 119 EU after 26 weeks. In the 5- to 14-year-old children, anti-Vi IgG levels at 6 weeks elicited by Vi-rEPA(2), Vi-rEPA(1), and Vi were 169, 22.8, and 18.9 EU, respectively (P = 0.0001 for Vi-rEPA(1) and Vi with respect to Vi-rEPA(2)). At 26 weeks, the anti-Vi IgG levels for recipients of Vi-rEPA(2), Vi-rEPA(1), and Vi were 30.0, 10.8, and 13.4 EU, respectively (P < 0.001 for Vi-rEPA(1) and Vi with respect to Vi-rEPA(2)); all were higher than the preinjection levels (P = 0. 0001). Vi-rEPA(2) also elicited the highest anti-Vi IgM and IgA levels of the three vaccines. In the 2- to 4-year-old children at 6 weeks following the first injection, Vi-rEPA(2) elicited an anti-Vi IgG level of 69.9 EU compared to 28.9 EU for Vi-rEPA(1) (P = 0.0001). Reinjection increased Vi antibody levels from 69.9 to 95.4 EU for Vi-rEPA(2) and from 28.9 to 83.0 EU for Vi-rEPA(1). At 26 weeks, anti-Vi IgG levels remained higher than those at preinjection (30.6 versus 0.18 for Vi-rEPA(2) and 12.8 versus 0.33 for Vi-rEPA(1); P = 0.0001 for both). Vi vaccine is recommended for individuals of 5 years of age or older. In the present study, the GM level of anti-Vi IgG elicited by two injections of Vi-rEPA(2) in the 2- to 4-year-old children was higher than that elicited by Vi in the 5- to 14 year-old children (30.6 versus 13.4; P = 0.0001). The safety and immunogenicity of the Vi-rEPA(2) conjugate warrant further investigation. PMID- 10531233 TI - Consequence of Nramp1 deletion to Mycobacterium tuberculosis infection in mice. AB - 129sv mice functionally deleted of the antimicrobial resistance gene, Nramp1, were found to be as resistant as wild-type mice to infection with the virulent H37Rv strain of Mycobacterium tuberculosis, as determined by monitoring bacterial growth in major organs and recording host survival times. Death of infected mice of both types was associated with extensive infection-induced pathology in the lungs but not in other major organs. These findings are in keeping with the view that Nramp1 is of limited importance in resistance to tuberculosis in mice. PMID- 10531234 TI - Construction and characterization of Moraxella catarrhalis mutants defective in expression of transferrin receptors. AB - We have previously reported the construction of an isogenic mutant defective in expression of OmpB1, the TbpB homologue, in Moraxella catarrhalis 7169. In this report, we have extended these studies by constructing and characterizing two new isogenic mutants in this clinical isolate. One mutant is defective in expression of TbpA, and the other mutant is defective in expression of both TbpA and TbpB. These isogenic mutants were confirmed by using PCR analysis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and sequencing. In vitro growth studies, comparing all three mutants, demonstrated that the tbpA mutant and the tbpAB mutant were severely limited in their ability to grow with human holotransferrin as the sole source of iron. In contrast, the ompB1 (tbpB) mutant was capable of utilizing iron from human transferrin, although not to the extent of the parental strain. While affinity chromatography with human holotransferrin showed that each Tbp was capable of binding independently to transferrin, solid phase transferrin binding studies using whole cells demonstrated that the tbpA mutant exhibited binding characteristics similar to those seen with the wild-type bacteria. However, the ompB1 (tbpB) mutant exhibited a diminished capacity for binding transferrin, and no binding was detected with the double mutant. These data suggest that the M. catarrhalis TbpA is necessary for the acquisition of iron from transferrin. In contrast, TbpB is not essential but may serve as a facilitory protein that functions to optimize this process. Together these mutants are essential to provide a more thorough understanding of iron acquisition mechanisms utilized by M. catarrhalis. PMID- 10531235 TI - Local production of chemokines during experimental vaginal candidiasis. AB - Recurrent vulvovaginal candidiasis, caused by Candida albicans, is a significant problem in women of childbearing age. Although cell-mediated immunity (CMI) due to T cells and cytokines is the predominant host defense mechanism against C. albicans at mucosal tissue sites, host defense mechanisms against C. albicans at the vaginal mucosa are poorly understood. Based on an estrogen-dependent murine model of vaginal candidiasis, our data suggest that systemic CMI is ineffective against C. albicans vaginal infections. Thus, we have postulated that local immune mechanisms are critical for protection against infection. In the present study, the kinetic production of chemokines normally associated with the chemotaxis of T cells, macrophages (RANTES, MIP-1alpha, MCP-1), and polymorphonuclear neutrophils (MIP-2) was examined following intravaginal inoculation of C. albicans in estrogen-treated or untreated mice. Results showed significant increases in MCP-1 protein and mRNA in vaginal tissue of infected mice as early as 2 and 4 days postinoculation, respectively, that continued through a 21-day observation period, irrespective of estrogen status. No significant changes were observed with RANTES, MIP-1alpha, or MIP-2, although relatively high constitutive levels of RANTES mRNA and MIP-2 protein were observed. Furthermore, intravaginal immunoneutralization of MCP-1 with anti-MCP-1 antibodies resulted in a significant increase in vaginal fungal burden early during infection, suggesting that MCP-1 plays some role in reducing the fungal burden during vaginal infection. However, the lack of changes in leukocyte profiles in vaginal lavage fluids collected from infected versus uninfected mice suggests that MCP-1 functions to control vaginal C. albicans titers in a manner independent of cellular chemotactic activity. PMID- 10531236 TI - Enhanced macrophage resistance to Pseudomonas exotoxin A is correlated with decreased expression of the low-density lipoprotein receptor-related protein. AB - Cellular intoxification by exotoxin A of Pseudomonas aeruginosa (PEA) begins when PEA binds to its cellular receptor, the low-density lipoprotein receptor-related protein (LRP). This receptor is particularly abundant on macrophages. We hypothesize here that inducible changes in cellular expression levels of the LRP represent an important mechanism by which macrophage susceptibility to PEA is regulated by the host. We have examined the effect of lipopolysaccharide (LPS) on LRP expression and PEA sensitivity in the macrophage-like cell line HS-P. Using a [(3)H]leucine incorporation assay to measure inhibition of protein synthesis, we have demonstrated that HS-P macrophages are highly sensitive to PEA and that PEA toxicity is decreased by the LRP antagonist receptor-associated protein. LPS pretreatment decreases HS-P PEA sensitivity in a time- and dose-dependent manner. The dose of toxin required to inhibit protein synthesis by 50% increased from 11.3 +/- 1.2 ng/ml in untreated cells to 25.7 +/- 2.0 ng/ml in cells treated with LPS. In pulse experiments, involving brief exposure to saturating concentrations of PEA, [(3)H]leucine incorporation was more than threefold higher in cells pretreated with LPS than in untreated macrophages. These changes in HS-P PEA sensitivity following LPS treatment were consistently associated with a fivefold decrease in HS-P LRP mRNA expression as measured by Northern blot analysis and a three-and-a-half-fold decrease in HS-P LRP-specific ligand internalization as determined by activated alpha(2)-macroglobulin internalization studies. These data demonstrate for the first time that modulation of LRP levels by extracellular signaling molecules can alter cellular PEA sensitivity. PMID- 10531237 TI - Emergence of Anaplasma marginale antigenic variants during persistent rickettsemia. AB - Anaplasma marginale is an ehrlichial pathogen of cattle, in the order Rickettsiales, that establishes persistent cyclic rickettsemia in the infected host. Within each rickettsemic cycle, A. marginale expressing antigenically variant major surface protein 2 (MSP2) emerge. By cloning 17 full-length msp2 transcripts expressed during cyclic rickettsemia, we determined that emergent variants have a single, central hypervariable region encoding variant B-cell epitopes. The N- and C-terminal regions are highly conserved among the expressed A. marginale variants, and similar sequences define the MSP2 homologues in the agent of human granulocytic ehrlichiosis (HGE). This is in contrast to the MSP2 homologues in ehrlichial genogroup I pathogens, Ehrlichia chaffeensis, Ehrlichia canis, and Cowdria ruminantium, that have multiple hypervariable regions. By defining the variable and conserved regions, we were able to show that the single hypervariable region of A. marginale MSP2 encodes epitopes that are immunogenic and induce variant-specific antibody responses during persistent infection. These findings demonstrate that the MSP2 structural variants that emerge during each cycle of persistent rickettsemia are true antigenic variants, consistent with MSP2 antigenic variation as a mechanism of A. marginale persistence. PMID- 10531238 TI - Native and mutant forms of cholera toxin and heat-labile enterotoxin effectively enhance protective efficacy of live attenuated and heat-killed Shigella vaccines. AB - Both native and mutant forms of cholera toxin (CT) and heat-labile enterotoxin (LT) are effective adjuvants for antigens and killed whole-cell preparations. To determine whether these toxin molecules could also boost the immunogenicity and efficacy of live attenuated vaccines directed against shigellosis, the guinea pig keratoconjunctivitis model was used to evaluate the adjuvant effect of these toxin molecules on EcSf2a-3, a DeltavirG DeltaaroD Escherichia coli-Shigella flexneri 2a hybrid vaccine strain that was previously found to be less protective than its parent strain in the guinea pig model. Experiments using native and mutant toxin molecules showed that both CT and LT and mutant derivatives were effective as an adjuvant for EcSf2a-3 and that the mutant toxin molecules, which were developed to retain adjuvanticity without the toxicity associated with the native molecules, were as effective as the native toxin molecules as adjuvants. Protective efficacy was enhanced for both the oral and intranasal routes of immunization. Serum antibody response to the S. flexneri 2a O antigen, the primary antigen for protective immunity, was not dependent on the addition of an adjuvant. However, enumeration of the O-antigen-specific immunoglobulin G (IgG) and IgA antibody-secreting cells in the spleen and draining lymph nodes following intranasal immunization suggested that enhancement of the local immune response by the toxin molecules may contribute to the observed increase in protective efficacy. The efficacy of heat-killed S. flexneri 2a was enhanced only by mutant LT molecules. These results suggest that the best candidates for enhancing the efficacy of both live attenuated and heat-killed Shigella vaccines with minimal reactogenicity are the mutant toxin molecules. PMID- 10531239 TI - Coadministration of interleukin 12 expression vector with antigen 2 cDNA enhances induction of protective immunity against Coccidioides immitis. AB - Interleukin 12 (IL-12) plays an important role in the induction of protective immunity against cancer and infectious diseases. In this study we asked whether IL-12 cDNA could increase the protective capacity of the antigen 2 (Ag2) gene vaccine in experimental coccidioidomycosis. Coimmunization of BALB/c mice with a single-chain IL-12 cDNA (p40-L-p35) and Ag2 cDNA, both subcloned into the pVR1012 plasmid, significantly enhanced protection against systemic challenge with 2,500 arthroconidia, as evidenced by a greater-than-1.3-log-unit reduction in the fungal load in the lungs and spleens compared to mice receiving the pVR1012 vector alone, Ag2 cDNA alone, or IL-12 cDNA alone. The enhanced protection was associated with increased gamma interferon secretion; production of immunoglobulin G2a (IgG2a), IgG2b, and IgG3 antibodies to Coccidioides immitis antigen; and the influx of CD4(+) and CD8(+) T cells in lungs and spleens. When challenged by the pulmonary route, mice covaccinated with Ag2 cDNA and IL-12 cDNA were not protected at the lung level but did show a significant reduction in the fungal load in their livers and spleens compared to mice vaccinated with Ag2 cDNA or IL-12 cDNA alone. These results suggest that IL-12 acts as a therapeutic adjuvant to enhance Ag2 cDNA-induced protective immunity against experimental coccidioidomycosis through the induction of Th1-associated immune responses. PMID- 10531240 TI - Contribution of quorum sensing to the virulence of Pseudomonas aeruginosa in burn wound infections. AB - The Pseudomonas aeruginosa quorum-sensing systems, las and rhl, control the production of numerous virulence factors. In this study, we have used the burned mouse model to examine the contribution of quorum-sensing systems to the pathogenesis of P. aeruginosa infections in burn wounds. Different quorum-sensing mutants of P. aeruginosa PAO1 that were defective in the lasR, lasI, or rhlI gene or both the lasI and rhlI genes were utilized. The following parameters of the P. aeruginosa infection were examined: (i) lethality to the burned mouse, (ii) dissemination of the P. aeruginosa strain within the body of the infected mouse (by determining the numbers of CFU of P. aeruginosa within the liver and spleen), and (iii) spread of the P. aeruginosa strain within the burned skin (by determining the numbers of CFU of P. aeruginosa at the inoculation site and at a site about 15 mm from the inoculation site [distant site]). In comparison with that of PAO1, the in vivo virulence of lasI, lasR, and rhlI mutants was significantly reduced. However, the most significant reduction in in vivo virulence was seen with the lasI rhlI mutant. The numbers of CFU that were recovered from the livers, spleens, and skin of mice infected with different mutants were significantly lower than those of PAO1. At 8 and 16 h post burn infection, comparable numbers of CFU of PAO1 and lasI and rhlI mutants were obtained from both the inoculation and distant sites of the burned skin of infected mice. In contrast, CFU of the lasR mutant and the lasI rhlI double mutant were recovered only from the inoculation site of infected mice at 8 and 16 h post burn infection. The ability of a plasmid carrying either the lasI or rhlI gene or the lasI and rhlI genes to complement the defect of the lasI rhlI double mutant was also examined. The presence of any of these plasmids within the lasI rhlI double mutant significantly enhanced its in vivo virulence, as well as its ability to spread within the burned skin. These results suggest that the quorum sensing systems play an important role in the horizontal spread of P. aeruginosa within burned skin and in the dissemination of P. aeruginosa within the bodies of burned-and-infected mice and contributed to the overall virulence of P. aeruginosa in this animal model. PMID- 10531241 TI - Targeted salivary gland immunization with plasmid DNA elicits specific salivary immunoglobulin A and G antibodies and serum immunoglobulin G antibodies in mice. AB - For the development of vaccines against oral and pharyngeal pathogens invading the mucosal epithelia, both secretory and serum immunoglobulin A (IgA) and IgG antibodies and cytotoxic T lymphocytes (CTL) have been induced. We used a novel approach, targeted salivary gland (TSG) immunization, using plasmid pcDNA3/fimA, coding for Porphyromonas gingivalis fimbriae. Expression of subunit protein, fimbrillin, was observed in eukaryotic cells growing in vitro following transfection with pcDNA3/fimA. In this study, we obtained good humoral and cell mediated immune responses in BALB/c mice by TSG administration using the above mentioned DNA vaccine. The production of fimbria-specific IgA and IgG antibodies in saliva and serum IgG antibody was significantly stimulated by TSG immunization. Injection of DNA vaccine into salivary gland elicited high-level production of antigen-specific IgG antibody, similar to that induced following intramuscular immunization. The major IgG subclass that recognized fimbriae was IgG2a in serum from pcDNA3/fimA-immunized mice. Reverse transcription-PCR analysis of mononuclear cells from salivary glands showed that levels of Th2 cytokine-specific mRNA were higher in the immunized group than in the nonimmunized group. In addition, TSG DNA immunization resulted in the generation of antigen-specific CTL in spleen. These results indicate that TSG immunization with plasmid DNA may represent a genetic immunization strategy against infection by oral and pharyngeal pathogens that may invade local, mucosal surfaces. PMID- 10531242 TI - Immune CD8(+) T cells prevent reactivation of Toxoplasma gondii infection in the immunocompromised host. AB - Toxoplasma gondii remains a serious cause of morbidity and mortality in individuals that are immunosuppressed, patients with AIDS in particular. The cellular immune response, especially by gamma interferon (IFN-gamma)-producing CD8(+) T cells, is an essential component of protective immunity against the parasite. In the present study the role of CD8(+) T cells during the reactivation of Toxoplasma infection in an immunocompromised murine model was evaluated. Chronically infected mice were challenged with LP-BM5 virus, and the kinetics of CD8(+) T-cell function was studied. At 10 weeks after viral infection, mice showed obvious signs of systemic illness and began to die. At this stage, CD8(+) T cells were unresponsive to antigenic stimulation and unable to kill Toxoplasma infected targets. IFN-gamma production by the CD8(+) T cells from dual-infected animals reached background levels, and a dramatic fall in the frequency of precursor cytotoxic T lymphocytes was observed. Histopathological analysis of the tissues demonstrated signs of disseminated toxoplasmosis as a result of reactivation of infection. However, treatment of the dual-infected animals with immune CD8(+) T cells at 5 weeks post-LP-BM5 challenge prevented the reactivation of toxoplasmosis, and mice continued to live. Our study for the first time demonstrates a therapeutic role for CD8(+) T cells against an opportunistic infection in an immunocompromised state. PMID- 10531243 TI - Protection of mice against brucellosis by vaccination with Brucella melitensis WR201(16MDeltapurEK). AB - Human brucellosis can be acquired from infected animal tissues by ingestion, inhalation, or contamination of the conjunctiva or traumatized skin by infected animal products. A vaccine to protect humans from occupational exposure or from zoonotic infection in areas where the disease is endemic would reduce an important cause of morbidity worldwide. Vaccines currently used in animals are unsuitable for human use. We tested a live, attenuated, purine-auxotrophic mutant strain of Brucella melitensis, WR201, for its ability to elicit cellular and humoral immune responses and to protect mice against intranasal challenge with B. melitensis 16M. Mice inoculated intraperitoneally with WR201 made serum antibody to lipopolysaccharide and non-O-polysaccharide antigens. Splenocytes from immunized animals released interleukin-2 (IL-2), gamma interferon, and IL-10 when cultured with Brucella antigens. Immunization led to protection from disseminated infection but had only a slight effect on clearance of the challenge inoculum from the lungs. These studies suggest that WR201 should be further investigated as a vaccine to prevent human brucellosis. PMID- 10531245 TI - Intranasal immunization with pneumococcal polysaccharide conjugate vaccines with nontoxic mutants of Escherichia coli heat-labile enterotoxins as adjuvants protects mice against invasive pneumococcal infections. AB - Host defenses against Streptococcus pneumoniae depend largely on phagocytosis following opsonization by polysaccharide-specific immunoglobulin G (IgG) antibodies and complement. Since colonization of the respiratory mucosa is the first step in pneumococcal pathogenesis, mucosal immune responses may play a significant role. In addition to inducing systemic immune responses, mucosal vaccination with an effective adjuvant has the advantage of inducing mucosal IgA antibodies. The heat-labile enterotoxin (LT) of Escherichia coli is a well studied mucosal adjuvant, and adjuvant activity of nontoxic LT mutants has been demonstrated for several protein antigens. We investigated the immunogenicity of pneumococcal polysaccharide conjugate vaccines (PNC) of serotypes 1 and 3 in mice after intranasal (i.n.) immunization by using as an adjuvant the nontoxic LT mutant LT-K63 or LT-R72, which has minimal residual toxicity. Pneumococcal serotype-specific antibodies were measured in serum (IgM, IgG, and IgA) and saliva (IgA), and vaccine-induced protection was evaluated by i.n. challenge with virulent pneumococci of the homologous serotype. When administered with LT mutants, i.n. immunization with both conjugates induced systemic and mucosal immune responses, and serum IgG antibody levels were significantly higher than after subcutaneous immunization. All mice immunized i.n. with PNC-1 and LT mutants were protected against bacteremia and cleared the pneumococci from the lung 24 h after i.n. challenge; pneumococcal density correlated significantly with serum IgG antibody levels. Similarly, the survival of mice immunized i.n. with PNC-3 and LT mutants was significantly prolonged. These results demonstrate that i.n. vaccination with PNC and potent adjuvants can protect mice against invasive and lethal pneumococcal infections, indicating that mucosal vaccination with PNC may be an alternative vaccination strategy for humans. PMID- 10531244 TI - Reactive nitrogen and oxygen species ameliorate experimental cryptosporidiosis in the neonatal BALB/c mouse model. AB - Four-day-old BALB/c mice were infected by the oral administration of 50,000 Cryptosporidium parvum oocysts, and the resulting infection was scored histologically and by counting colonic oocysts. Infection occurred in the ileum and proximal colon (but not duodenum and jejunum), peaked on days 14 to 18, and was cleared between days 24 and 30. Nitric oxide (NO) appeared to play a protective role in this model as evidenced by the facts that plasma nitrite and nitrate levels increased during the period of peak parasitosis; immunohistochemically detected inducible nitric oxide synthase (iNOS) was increased in the ileum and colon enterocytes of infected animals; the NOS inhibitor L-N-iminoethyl lysine or N-nitro-L-arginine methyl ester (L-NAME) decreased the elevated plasma nitrite and nitrate levels while exacerbating the infection and increasing oocyst shedding; administration of a NO donor, S-nitroso N-penicillamine, reduced oocyst and infection scores; and neonatal iNOS knockout mice exhibited a slightly longer infection than control animals. The oral administration of oocysts to L-NAME-treated BALB/c mice, but not control animals, between 24 and 40 days old resulted in the fecal excretion of oocysts 1 week later. Administration of the antioxidant ascorbic acid also exacerbated the C. parvum infection, suggesting a protective role for reactive nitrogen and/or reactive oxygen compounds, while administration of the superoxide scavenger superoxide dismutase exacerbated the infection. Taken together these data suggest that both reactive nitrogen and reactive oxygen species play protective roles in experimental cryptosporidiosis. PMID- 10531246 TI - Alternative mechanism of cholera toxin acquisition by Vibrio cholerae: generalized transduction of CTXPhi by bacteriophage CP-T1. AB - Horizontal transfer of genes encoding virulence factors has played a central role in the evolution of many pathogenic bacteria. The unexpected discovery that the genes encoding cholera toxin (ctxAB), the main cause of the profuse secretory diarrhea characteristic of cholera, are encoded on a novel filamentous phage named CTXPhi, has resulted in a renewed interest in the potential mechanisms of transfer of virulence genes among Vibrio cholerae. We describe here an alternative mechanism of cholera toxin gene transfer into nontoxigenic V. cholerae isolates, including strains that lack both the CTXPhi receptor, the toxin coregulated pilus (TCP), and attRS, the chromosomal attachment site for CTXPhi integration. A temperature-sensitive mutant of the V. cholerae generalized transducing bacteriophage CP-T1 (CP-T1ts) was used to transfer a genetically marked derivative of the CTX prophage into four nontoxigenic V. cholerae strains, including two V. cholerae vaccine strains. We demonstrate that CTXPhi transduced by CP-T1ts can replicate and integrate into these nontoxigenic V. cholerae strains with high efficiency. In fact, CP-T1ts transduces the CTX prophage preferentially when compared with other chromosomal markers. These results reveal a potential mechanism by which CTXPhi(+) V. cholerae strains that lack the TCP receptor may have arisen. Finally, these findings indicate an additional pathway for reversion of live-attenuated V. cholerae vaccine strains. PMID- 10531248 TI - Effect of multiple antigenic exposures in the gut on oral tolerance and induction of antibacterial systemic immunity. AB - We have analyzed oral tolerance of microbial antigens in an experimental model in which mice are treated orally with a single small dose of soluble antigen and challenged systemically with the antigen in complete Freund's adjuvant. We found that, while oral administration of sonicated extracts of either Leishmania major, Leishmania donovani, or Staphylococcus aureus was tolerogenic, as was administration of the nominal antigen ovalbumin or conalbumin, oral administration of Escherichia coli or Salmonella typhimurium sonicated extract was not. Since E. coli is an enteric commensal that colonizes the intestine soon after birth, these data suggested that lack of demonstrable oral tolerance may be related to the frequency of oral exposure to an antigen. In support of this, we found that multiple oral doses of ovalbumin or S. aureus or L. donovani antigens did not increase systemic hyporesponsiveness beyond that achieved with a single oral dose. We have also tested the ability of mice fed with sonicates of the tolerogenic S. aureus or the nontolerogenic S. typhimurium to clear a subsequent systemic infection with the homologous bacteria and found that, while clearance of S. aureus was unaffected by prior feeding, clearance of S. typhimurium was actually enhanced. The data suggest that frequent oral antigenic exposure may eventually lead to induction of systemic immunity in tolerant mice. PMID- 10531247 TI - Allelic diversity and antibody recognition of Plasmodium falciparum merozoite surface protein 1 during hypoendemic malaria transmission in the Brazilian amazon region. AB - The polymorphic merozoite surface protein (MSP-1) of Plasmodium falciparum is a major asexual blood-stage malaria vaccine candidate. The impact of allelic diversity on recognition of MSP-1 during the immune response remains to be investigated in areas of hypoendemicity such as the Brazilian Amazon region. In this study, PCR was used to type variable regions, blocks 2, 4, and 10, of the msp-1 gene and to characterize major gene types (unique combinations of allelic types in variable blocks) in P. falciparum isolates collected across the Amazon basin over a period of 12 years. Twelve of the 24 possible gene types were found among 181 isolates, and 68 (38%) of them had more than one gene type. Temporal, but not spatial, variation was found in the distribution of MSP-1 gene types in the Amazon. Interestingly, some gene types occurred more frequently than expected from random assortment of allelic types in different blocks, as previously found in other areas of endemicity. We also compared the antibody recognition of polymorphic (block 2), dimorphic (block 6), and conserved (block 3) regions of MSP-1 in Amazonian malaria patients and clinically immune Africans, using a panel of recombinant peptides. Results were summarized as follows. (i) All blocks were targeted by naturally acquired cytophilic antibodies of the subclasses IgG1 and IgG3, but the balance between IgG1 and IgG3 depended on the subjects' cumulative exposure to malaria. (ii) The balance between IgG1 and IgG3 subclasses and the duration of antibody responses differed in relation to distinct MSP-1 peptides. (iii) Antibody responses to variable blocks 2 and 6 were predominantly type specific, but variant-specific antibodies that target isolate-specific repetitive motifs within block 2 were more frequent in Amazonian patients than in previously studied African populations. PMID- 10531249 TI - A DNA sequence corresponding to the gene encoding cysteine proteinase 5 in Entamoeba histolytica is present and positionally conserved but highly degenerated in Entamoeba dispar. AB - Cysteine proteinases of Entamoeba histolytica are considered to be one of the most important classes of molecules responsible for the parasite's ability to destroy human tissues. Interestingly, one particular cysteine proteinase, located on the surface of E. histolytica trophozoites and designated cysteine proteinase 5 (CP5), is not expressed in the closely related but nonpathogenic species Entamoeba dispar. By comparing the E. histolytica and E. dispar genomic loci containing the gene for CP5 (cp5), it was found that the position of cp5 within the genomic context is conserved between the two organisms, but that the gene is highly degenerated in E. dispar, as it contains numerous nucleotide exchanges, insertions, and deletions, resulting in multiple stop codons within the cp5 reading frame. An alignment of all available orthologous E. histolytica and E. dispar DNA sequences suggested that cp5 started to degenerate in E. dispar coincidently when the two organisms began to diverge from a common ancestor. PMID- 10531250 TI - Molecular characterization of the locus encoding biosynthesis of the lipopolysaccharide O antigen of Escherichia coli serotype O113. AB - Shiga toxigenic Escherichia coli (STEC) strains are a diverse group of organisms capable of causing severe gastrointestinal disease in humans. Within the STEC family, eae-positive STEC strains, particularly those belonging to serogroups O157 and O111, appear to have greater virulence for humans. However, in spite of being eae negative, STEC strains belonging to serogroup O113 have frequently been associated with cases of severe STEC disease, including hemolytic-uremic syndrome (HUS). Western blot analysis with convalescent-phase serum from a patient with HUS caused by an O113:H21 STEC strain indicated that human immune responses were directed principally against lipopolysaccharide O antigen. Accordingly, the serum was used to isolate a clone expressing O113 O antigen from a cosmid library of O113:H21 DNA constructed in E. coli K-12. Sequence analysis indicated that the O113 O-antigen biosynthesis (rfb) locus contains a cluster of nine genes which may be cotranscribed. Comparison with sequence databases identified candidate genes for four glycosyl transferases, an O-acetyl transferase, an O-unit flippase, and an O-antigen polymerase, as well as copies of galE and gnd. Two additional, separately transcribed genes downstream of the O113 rfb region were predicted to encode enzymes involved in synthesis of activated sugar precursors, one of which (designated wbnF) was essential for O113 O-antigen synthesis, and so is clearly a part of the O113 rfb locus. Interestingly, expression of O113 O antigen by E. coli K-12 significantly increased in vitro adherence to both HEp-2 and Henle 407 cells. PMID- 10531251 TI - Shiga toxin-producing Escherichia coli can impair T84 cell structure and function without inducing attaching/effacing lesions. AB - Enteropathogenic Escherichia coli (EPEC) intimately adhere to epithelial cells producing cytoskeletal rearrangement with typical attaching and effacing lesions and altered epithelial barrier and transport function. Since EPEC and Shiga toxin producing E. coli (STEC) share similar genes in the "locus for enterocyte effacement" (LEE) thought to cause these changes, it has been assumed that STEC shares similar pathogenic mechanisms with EPEC. The aims of this study were to compare the effects of EPEC and STEC on bacterial-epithelial interactions and to examine changes in epithelial function. T84 monolayers were infected with STEC O157:H7 (wild strain EDL 933 or non-toxin-producing strain 85/170), EPEC (strain E2348/69), or HB101 (nonpathogenic) and studied at various times after infection. EPEC bound more avidly than EDL 933, but both strains exhibited greater binding than HB101. Attaching and effacing lesions and severe disruption to the actin cytoskeleton were observed in EPEC by 3 h postinfection but not in EDL 933 or HB101 at any time point. EPEC and EDL 933 increased monolayer permeability to [(3)H]mannitol 5- to 10-fold. In contrast to EPEC, EDL 933 completely abolished secretagogue-stimulated anion secretion as assessed under voltage clamp conditions in Ussing chambers. Several other STEC strains induced changes similar to those of EDL 933. In conclusion, STEC impairs epithelial barrier function and ion transport without causing major disruption to the actin cytoskeleton. Pathogenic factors other than products of LEE may be operant in STEC. PMID- 10531252 TI - Recognition of schistosome glycolipids by immunoglobulin E: possible role in immunity. AB - To investigate the role of antibody responses to (glyco)lipids in immunity to schistosome infection, lipids extracted from Schistosoma mansoni eggs and adult worms were fractionated, and the antibody isotype profile reactive to the fractionated lipids in a well-characterized S. haematobium-infected population was investigated. In tests of 10 plasma samples it was found that immunoglobulin G (IgG) reactivity was highest to the fraction containing ceramidepolyhexosides, whereas IgE reactivity was most prominent to both cerebroside- and ceramidepolyhexoside-containing fractions. The fraction containing ceramidepolyhexosides was then tested for reactivity with IgG subclasses and IgE in plasma samples from 66 S. haematobium-infected patients. Considering IgG4 and IgE, isotypes of particular interest in helminth infections, we found that both isotypes recognized egg (glyco)proteins in more than 90% of the infected subjects. However, in the case of glycolipids, IgE reactivity was much more prominent than IgG4 reactivity (found in 80 and 41% of the subjects, respectively). Furthermore, worm glycolipid-specific IgE prior to treatment of the subjects with praziquantel was negatively correlated with egg counts at 2 years posttreatment, indicating that IgE directed towards glycolipids could play an important role in resistance to reinfection. PMID- 10531253 TI - Human onchocerciasis and tetanus vaccination: impact on the postvaccination antitetanus antibody response. AB - To investigate whether helminth infections may affect the efficacy of vaccines by impairing the immune response to nonparasite vaccine antigens, we compared the antibody responses to tetanus toxoid (TT) after tetanus vaccination in 193 subjects with Onchocerca volvulus infection with 85 comparable noninfected controls. After vaccination, the proportions of subjects in each group attaining protective levels of antitetanus antibodies were similar (96.9% infected versus 97.6% noninfected). Postvaccination increases in antitetanus immunoglobulin G (IgG) and the predominant IgG isotype, IgG1, were equivalent in both groups, as were increases in specific IgG4 and IgE; however, significantly greater increases in specific IgG2 (P < 0.05) and IgG3 (P < 0.001) were observed in the noninfected group. Stratification of the O. volvulus-infected group into two groups representing light and heavy infections revealed a significantly impaired antitetanus IgG response in those with heavy infections compared to those with light infections (P < 0.01) or no infection (P < 0.05). The impact of concurrent intestinal helminth infections on the antitetanus response was also examined; an increased IgG4/IgE ratio was seen in those infected with Strongyloides stercoralis (P < 0.05) and when all helminth infections were combined as a single group (P < 0.05). These findings indicate that concurrent infection with O. volvulus does not prevent the development of a protective antitetanus response, although heavier O. volvulus infections are able to alter the magnitude of this response, and concurrent helminth infections (O. volvulus and intestinal helminths) may alter TT-specific antibody isotype responses. PMID- 10531254 TI - Experimental gestational pyelonephritis induces preterm births and low birth weights in C3H/HeJ mice. AB - Urinary tract infections (UTIs) are associated with approximately 27% of premature births. Escherichia coli is the most frequent causal agent of UTIs and expresses virulence factors, including surface adhesins that recognize specific host tissue receptors. We have reported that E. coli Dr adhesin recognizes decay accelerating factor as the host tissue receptor and that these receptors are increased during pregnancy. Induction of pathogenesis is a cumulative effect of the host-pathogen relationship involving specific host factors and virulence characteristics of the invading organism. Recently, an experimental model of chronic pyelonephritis has been developed with E. coli bearing Dr adhesin (E. coli Dr(+)) in nonpregnant lipopolysaccharide hyporesponder C3H/HeJ mice. In this study, we investigated the role of E. coli Dr(+) on the outcome of pregnancy in C3H/HeJ mice. Groups of pregnant mice were infected with E. coli Dr(+) or its isogenic mutant which does not bear the Dr adhesin (E. coli Dr(-)) by urethral catheterization. Nearly 90% of pregnant mice infected with E. coli Dr(+) delivered preterm (before 90% gestation) compared to 10% of mice infected with E. coli Dr(-) and none of the mice treated with phosphate-buffered saline (PBS). Also, there was a significant reduction in fetal birth weight in the E. coli Dr(+)-infected group compared to the E. coli Dr(-)- and PBS-treated groups (P = 0.003). This experimental model of E. coli Dr(+)-induced preterm delivery in mice may help in understanding the molecular mechanisms involved in UTI-induced preterm labor involving bacterial adhesins. PMID- 10531255 TI - T-cell recognition of Mycobacterium tuberculosis culture filtrate fractions in tuberculosis patients and their household contacts. AB - We examined the immune responses of patients with active pulmonary tuberculosis (TB) and their healthy household contacts to short-term culture filtrate (ST-CF) of Mycobacterium tuberculosis or molecular mass fractions derived from it. Our goal was to identify fractions strongly recognized by donors and differences among the donor groups of possible relevance for vaccine development. The study population consisted of 65 human immunodeficiency virus-negative donors from the Hossana Regional Hospital, Hossana, Ethiopia. Peripheral blood leukocytes from the donors were stimulated with different antigens and immune responses were determined. Household contacts produced significantly higher levels of gamma interferon (IFN-gamma) than the TB patients in response to antigens present in ST CF and the 10 narrow-molecular-mass fractions. A similar difference in leukocyte proliferative responses to the antigens between the two groups was also found. In general, while all fractions stimulated immune responses, the highest activity was seen with the low-molecular-mass fractions, which include well-defined TB antigens such as ESAT-6. Leukocytes from contacts of TB patients with severe disease produced higher levels of antigen-specific IFN-gamma than those from contacts of patients with minimal disease. Both groups of contacts exhibited higher cell-mediated responses than the patients themselves. The enhanced immune response of healthy contacts, especially those of patients with severe disease, to secreted mycobacterial antigens is suggestive of an early stage of infection by M. tuberculosis, which could in time result in overt disease or containment of the infection. This possibility is currently being investigated by follow-up studies of the household contacts. PMID- 10531257 TI - Determination of antibody responses of elderly adults to all 23 capsular polysaccharides after pneumococcal vaccination. AB - The 23-valent pneumococcal polysaccharide vaccine was formulated to prevent invasive infection in the elderly and other high-risk populations from the most prevalent Streptococcus pneumoniae serotypes. However, the immunogenicity of all 23 vaccine polysaccharides has not been fully characterized in elderly adults. We previously reported that whereas the majority of elderly subjects had vigorous immune responses to selected pneumococcal vaccine polysaccharides, a subset of elderly individuals responded to fewer than two of seven vaccine serotypes after immunization. To determine whether these elderly low responders have a general inability to respond to pneumococcal vaccine and to determine whether elderly low responders might be identified by their responses to a few polysaccharides, we measured antibody responses of elderly adults to all 23 vaccine polysaccharides after pneumococcal immunization. As a group, elderly subjects showed a significant rise after immunization in geometric mean antibody levels to all 23 vaccine serotypes. However, when individual rather than group immune responses were assessed, the 23-valent vaccine did not appear to be uniformly immunogenic in these elderly subjects. Eleven elderly subjects (20%) had twofold increases in specific antibody after vaccination to only 5 or fewer of the 23 vaccine polysaccharides, and they did not respond to the most prevalent serotypes causing invasive disease. Antibody responses to serotype 9N were found to reliably distinguish low vaccine responders from other elderly subjects. However, no particular group of vaccine polysaccharides could be used as a marker for adequate immune responses if only postvaccination sera were analyzed. PMID- 10531256 TI - Bordetella bronchiseptica-mediated cytotoxicity to macrophages is dependent on bvg-regulated factors, including pertactin. AB - The effect of Bordetella bronchiseptica infection on the viability of murine macrophage-like cells and on primary porcine alveolar macrophages was investigated. The bacterium was shown to be cytotoxic for both cell types, particularly where tight cell-to-cell contacts were established. In addition, bvg mutants were poorly cytotoxic for the eukaryotic cells, while a prn mutant was significantly less toxic than wild-type bacteria. B. bronchiseptica-mediated cytotoxicity was inhibited in the presence of cytochalasin D or cycloheximide, an inhibitor of microfilament-dependent phagocytosis or de novo eukaryotic protein synthesis, respectively. The mechanism of eukaryotic cell death was examined, and cell death was found to occur primarily through a necrotic pathway, although a small proportion of the population underwent apoptosis. PMID- 10531258 TI - Shiga toxins stimulate secretion of interleukin-8 from intestinal epithelial cells. AB - In the 1980s, Shiga toxin (Stx)-producing Escherichia coli O157:H7 (STEC) was identified as a cause of hemorrhagic colitis in the United States and was found to be associated with hemolytic uremic syndrome (HUS), a microangiopathic hemolytic anemia characterized by thrombocytopenia and renal failure. The precise way that Stxs cause hemorrhagic colitis and HUS is unclear. Stxs have been thought to cause disease by killing or irreversibly harming sensitive cells through a nonspecific blockade of mRNA translation, eventually resulting in cytotoxicity by preventing synthesis of critical molecules needed to maintain cell integrity. Because STEC is noninvasive, we have been exploring the host toxin response at the level of the gastrointestinal mucosa, where STEC infection begins. We have found that Stx is capable of interleukin-8 (IL-8) superinduction in a human colonic epithelial cell line. Despite a general blockade of mRNA translation, Stx treatment results in increased IL-8 mRNA as well as increased synthesis and secretion of IL-8 protein. Our data suggest that an active Stx A subunit is required for this activity. Ricin, which has the same enzymatic activity and trafficking pathway as Stx, has similar effects. Exploration of the effects of other protein synthesis inhibitors (cycloheximide, anisomycin) suggests a mechanism of gene regulation that is distinct from a general translational blockade. Use of the specific p38/RK inhibitor SB202190 showed that blocking of this pathway results in decreased Stx-mediated IL-8 secretion. Furthermore, Stxs induced mRNA of the primary response gene c-jun, which was subsequently partially blocked by SB202190. These data suggest a novel model of how Stxs contribute to disease, namely that Stxs may alter regulation of host cell processes in sensitive cells via activation of at least one member of the mitogen-activated protein kinase family in the p38/RK cascade and induction of c jun mRNA. Stx-induced increases in chemokine synthesis from intestinal epithelial cells could be important in augmenting the host mucosal inflammatory response to STEC infection. PMID- 10531259 TI - A genomic island, termed high-pathogenicity island, is present in certain non O157 Shiga toxin-producing Escherichia coli clonal lineages. AB - Shiga toxin-producing Escherichia coli (STEC) strains cause a wide spectrum of diseases in humans. In this study, we tested 206 STEC strains isolated from patients for potential virulence genes including stx, eae, and enterohemorrhagic E. coli hly. In addition, all strains were examined for the presence of another genetic element, the high-pathogenicity island (HPI). The HPI was first described in pathogenic Yersinia species and encodes the pesticin receptor FyuA and the siderophore yersiniabactin. The HPI was found in the genome of distinct clonal lineages of STEC, including all 31 eae-positive O26:H11/H(-) strains and 7 of 12 eae-negative O128:H2/H(-) strains. In total, the HPI was found in 56 (27.2%) of 206 STEC strains. However, it was absent from the genome of all 37 O157:H7/H(-), 14 O111:H(-), 13 O103:H2, and 13 O145:H(-) STEC isolates, all of which were positive for eae. Polypeptides encoded by the fyuA gene located on the HPI could be detected by using immunoblot analysis in most of the HPI-positive STEC strains, suggesting the presence of a functional yersiniabactin system. The HPI in STEC was located next to the tRNA gene asnT. In contrast to the HPI of other pathogenic enterobacteria, the HPI of O26 STEC strains shows a deletion at its left junction, leading to a truncated integrase gene int. We conclude from this study that the Yersinia HPI is disseminated among certain clonal subgroups of STEC strains. The hypothesis that the HPI in STEC contributes to the fitness of the strains in certain ecological niches rather than to their pathogenic potential is discussed. PMID- 10531260 TI - Importance of B cells, but not specific antibodies, in primary and secondary protective immunity to the intracellular bacterium Francisella tularensis live vaccine strain. AB - Although there appears to be little if any role for specific antibodies in protection against intracellular bacteria, such as the model pathogen F. tularensis live vaccine strain (LVS), the role of B cells themselves in primary and secondary infection with such bacteria has not been examined directly. We show here that mice deficient in mature B cells and antibodies (B-cell knockout mice) are marginally compromised in controlling primary sublethal infection but are 100-fold less well protected against secondary lethal challenge than are their normal counterparts. This defect in optimal specific protective immunity was readily reconstituted by the transfer of primed, and to a lesser degree, unprimed B cells, but not by the transfer of specific antibodies. The results indicate a previously unappreciated role for B cells in secondary immunity to intracellular pathogens through a function other than antibody production. PMID- 10531262 TI - Citrobacter rodentium espB is necessary for signal transduction and for infection of laboratory mice. AB - Citrobacter rodentium is the causative agent of transmissible murine colonic hyperplasia and contains a locus of enterocyte effacement (LEE) similar to that found in enteropathogenic Escherichia coli (EPEC). EPEC espB is necessary for intimate attachment and signal transduction between EPEC and cultured cell monolayers. Mice challenged with wild-type C. rodentium develop a mucosal immunoglobulin A response to EspB. In this study, C. rodentium espB has been cloned and its nucleotide sequence has been determined. C. rodentium espB was found to have 90% identity to EPEC espB. A nonpolar insertion mutation in C. rodentium espB was constructed and used to replace the chromosomal wild-type allele. The C. rodentium espB mutant exhibited reduced cell association and had no detectable fluorescent actin staining activity on cultured cell monolayers. The C. rodentium espB mutant also failed to colonize laboratory mice following experimental inoculation. The espB mutation could be complemented with a plasmid encoded copy of the gene, which restored both cell association and fluorescent actin staining activity, as well as the ability to colonize laboratory mice. These studies indicate that espB is necessary for signal transduction and for colonization of laboratory mice by C. rodentium. PMID- 10531261 TI - Identification, characterization, and expression of three new members of the Borrelia burgdorferi Mlp (2.9) lipoprotein gene family. AB - We previously reported on the existence of a family of lipoprotein genes, designated 2.9 lipoprotein genes, encoded in at least seven versions on the circular (supercoiled) cp32 and cp18 plasmids of Borrelia burgdorferi 297. A distinguishing feature of the 2.9 lipoproteins were highly similar signal sequences but variable mature polypeptides that segregated into two antigenic classes. Further screenings of B. burgdorferi 297 genomic libraries led to the identification of three additional 2.9 lipoprotein genes, renamed herein mlp, for multicopy lipoprotein genes. Computer analyses and immunoblotting revealed that Mlp-9 segregated with the antigenic class I lipoproteins, whereas Mlp-8 and Mlp 10 were members of class II. Northern blotting showed that all three of the mlp genes were expressed when B. burgdorferi was cultivated in vitro at 34 degrees C, although mlp-9 and mlp-10 transcripts were expressed at very low levels. Additional combined immunoblotting and comparative reverse transcription-PCR analyses performed on borreliae cultivated in vitro at 23, 34, or 37 degrees C indicated that although Mlp-8 was substantially more abundant than Mlp-9 or Mlp 10, all three of the mlp genes were upregulated during B. burgdorferi replication at 37 degrees C. Expression of the same three lipoproteins was further enhanced upon growth of the spirochetes within dialysis membrane chambers (DMCs) implanted intraperitoneally in rats (i.e., spirochetes in a mammalian host-adapted state), suggesting that temperature alone did not account for maximal upregulation of the mlp genes. That certain mlp genes are likely expressed during the growth of B. burgdorferi in mammalian tissues was supported by findings of antibodies against all three Mlp lipoproteins in mice after challenge with Ixodes scapularis nymphs harboring B. burgdorferi 297. The combined data suggest that as opposed to being differentially expressed in any reciprocal fashion (e.g., OspA/OspC), at least three mlp genes are simultaneously upregulated by temperature (37 degrees C) and some other mammalian host factor(s). The findings have importance not only for understanding alternative modes of differential antigen expression by B. burgdorferi but also for assessing whether one or more of the Mlp lipoproteins represent new candidate vaccinogens for Lyme disease. PMID- 10531263 TI - Differential expression of translational elements by life cycle variants of Coxiella burnetii. AB - Coxiella burnetii replicates as distinct morphological forms, which may allow potential life cycle variants to survive the harsh environment of the phagolysosome. Monoclonal antibodies (MAbs) were compared by Western blotting for reactivity with large cell variant (LCV) and small cell variant (SCV) antigens to characterize proteins differentially expressed by C. burnetii. MAb NM7.3 reacted with a approximately 32-kDa LCV-upregulated antigen, and MAb NM183 reacted with a approximately 45-kDa LCV-specific antigen. MAb NM7.3 was used to screen a lambdaZapII C. burnetii DNA expression library, and an immunoreactive clone was identified with sequence similarity to the Escherichia coli tsf gene, which encodes elongation factor Ts (EF-Ts). Since a similar screen with MAb NM183 did not identify immunoreactive clones, an alternate strategy was devised to clone the reactive antigen based on observations of cross-reactivity with the 45-kDa elongation factor Tu (EF-Tu) protein from Chlamydia trachomatis. The highly conserved nature of EF-Tu among eubacteria allowed PCR amplification of a tuf gene fragment (encoding approximately 95% of the predicted EF-Tu open reading frame) from C. burnetii using degenerate primers. The product of the cloned tuf gene fragment reacted with MAb NM183 in Western blot analysis, confirming the identity of the 45-kDa LCV-specific antigen. Identification of two proteins differentially expressed by C. burnetii, EF-Tu and EF-Ts, both essential components of the translational machinery of the cell, supports the hypothesis that LCVs are metabolically more active than SCVs. PMID- 10531265 TI - Overexpression of the Candida albicans ALA1 gene in Saccharomyces cerevisiae results in aggregation following attachment of yeast cells to extracellular matrix proteins, adherence properties similar to those of Candida albicans. AB - Candida albicans maintains a commensal relationship with human hosts, probably by adhering to mucosal tissue in a variety of physiological conditions. We show that adherence due to the C. albicans gene ALA1 when transformed into Saccharomyces cerevisiae, is comprised of two sequential steps. Initially, C. albicans rapidly attaches to extracellular matrix (ECM) protein-coated magnetic beads in small numbers (the attachment phase). This is followed by a relatively slower step in which cell-to-cell interactions predominate (the aggregation phase). Neither of these phases is observed in S. cerevisiae. However, expression of the C. albicans ALA1 gene from a low-copy vector causes S. cerevisiae transformants to attach to ECM-coated magnetic beads without appreciable aggregation. Expression of ALA1 from a high-copy vector results in both attachment and aggregation. Moreover, transcriptional fusion of ALA1 with the galactose-inducible promoters GALS, GALL, and GAL1, allowing for low, moderate, and high levels of inducible transcription, respectively, causes attachment and aggregation that correlates with the strength of the GAL promoter. The adherence of C. albicans and S. cerevisiae overexpressing ALA1 to a number of protein ligands occurs over a broad pH range, is resistant to shear forces generated by vortexing, and is unaffected by the presence of sugars, high salt levels, free ligands, or detergents. Adherence is, however, inhibited by agents that disrupt hydrogen bonds. The similarities in the adherence and aggregation properties of C. albicans and S. cerevisiae overexpressing ALA1 suggest a role in adherence and aggregation for ALA1 and ALA1 like genes in C. albicans. PMID- 10531264 TI - Laccase protects Cryptococcus neoformans from antifungal activity of alveolar macrophages. AB - While laccase of Cryptococcus neoformans is implicated in the virulence of the organism, our recent studies showing absence of melanin in the infected mouse brain has led us to a search for alternative roles for laccase in cryptococcosis. We investigated the role of laccase in protection of C. neoformans against murine alveolar macrophage (AM)-mediated antifungal activity by using a pair of congenic laccase-positive (2E-TUC) and laccase-deficient (2E-TU) strains. The laccase positive cells with laccase derepression were more resistant to the antifungal activity of AM than a laccase-deficient strain ([28.9 +/- 1.2]% versus [40.2 +/- 2.6]% killing). Addition of L-dopa to Cryptococcus to produce melanin in a laccase-positive strain resulted in a slight increase in protection of C. neoformans from the antifungal activity of macrophages ([25.4 +/- 3.4]% versus [28.9 +/- 1.2]% killing). Recombinant cryptococcal laccase exhibited iron oxidase activity in converting Fe(II) to Fe(III). Moreover, recombinant laccase inhibited killing of C. neoformans by hydroxyl radicals catalyzed by iron in a cell-free system. Addition of the hydroxyl radical scavenger mannitol or dimethyl sulfoxide to AMs prior to the introduction of cryptococcal cells decreased killing of both strains and reduced the difference in susceptibility between the laccase-positive and laccase-deficient strains. Furthermore, laccase-mediated protection from AM killing was inhibited by the addition of Fe(II), presumably by overcoming the effects of the iron oxidase activity of cryptococcal laccase. These results suggest that the iron oxidase activity of laccase may protect C. neoformans from macrophages by oxidation of phagosomal iron to Fe(III) with a resultant decrease in hydroxyl radical formation. PMID- 10531266 TI - De Novo induction of atherosclerosis by Chlamydia pneumoniae in a rabbit model. AB - Chlamydia pneumoniae, a bacterial respiratory tract pathogen, has been associated with atherosclerosis in humans. C. pneumoniae infection of the respiratory tracts of rabbits fed a noncholesterol diet induced changes of atherosclerosis of the aorta in 6 (26.1%) of 23 animals after a single inoculum at 3 months. Multiple inocula given three times within 6 weeks resulted in grade III atherosclerosis in 8 (34.8%) of 23 rabbits, with an additional 5 (21. 7%) showing increased myxoid changes in the intima-media junction and exhibiting 8 (34.8%) focal periaortitis. Control animals inoculated with carrier broth (n = 24), HEp-2 cells (n = 12), or another respiratory pathogen, Mycoplasma pneumoniae (n = 32), produced no changes of atherosclerosis after 3 months. The histological changes were dissimilar (fewer foam cells) from those of rabbits fed a 0.5% cholesterol diet but were highly similar to or indistinguishable from changes in rabbits fed a 0.15% cholesterol diet (similar to that of humans). Proinflammatory cytokines and tissue growth factors were more consistently detected in cholesterol-induced aortic lesions than those induced by C. pneumoniae. These data are compatible with de novo induction of atherogenesis by C. pneumoniae in rabbits and suggest that C. pneumoniae may be important in the pathogenesis of atherosclerosis in humans. PMID- 10531267 TI - Increased interleukin-1 (IL-1) and imbalance between IL-1 and IL-1 receptor antagonist during acute inflammation in experimental Shigellosis. AB - Infection by the enteric bacterial pathogen Shigella results in intense mucosal inflammation and destruction of the colonic and rectal epithelium in infected humans. Initial bacterial translocation occurs through the follicle-associated epithelium. Previous experiments suggest that interleukin-1 (IL-1) is crucial to trigger inflammation, particularly in the follicular zones. During the first 4 hours of infection in a rabbit ligated-loop model of intestinal invasion, there are two salient characteristics: (i) a high concentration of IL-1alpha and IL 1beta, both in infected Peyer's patch tissue and in the corresponding efferent mesenteric blood, and (ii) a very low level of expression of IL-1 receptor antagonist (IL-1ra). These may reflect a combination of regulation of expression and secretion of IL-1alpha, IL-1beta, and IL-1ra by both resident and recruited phagocytes and the induction of mononuclear phagocyte apoptosis by Shigella. This low IL-1ra/IL-1 ratio likely accounts for the rapid, uncontrolled inflammation characteristic of shigellosis. PMID- 10531268 TI - Enterococcus faecalis bearing aggregation substance is resistant to killing by human neutrophils despite phagocytosis and neutrophil activation. AB - Enterococcus faecalis aggregation substance (AS) mediates efficient bacterium bacterium contact to facilitate plasmid exchange as part of a bacterial sex pheromone system. We have previously determined that AS promotes direct, opsonin independent binding of E. faecalis to human neutrophils (PMNs) via complement receptor type 3 and other receptors on the PMN surface. We have now examined the functional consequences of this bacterium-host cell interaction. AS-bearing E. faecalis was phagocytosed and internalized by PMNs, as determined by deconvolution fluorescence microscopy. However, these bacteria were not killed by PMNs, and internalized bacteria excluded propidium iodide, indicating intact bacterial membranes. Resistance to killing occurred despite activation of PMNs, as indicated by an increase in both functional and total surface Mac-1 expression, shedding of L-selectin, and an increase in PMN extracellular superoxide and phagosomal oxidant production. Deconvolution fluorescence microscopy also revealed that phagosomes containing AS-bearing bacteria were markedly larger than phagosomes containing opsonized E. faecalis, suggesting that some modification of phagosomal maturation may be involved in AS-induced resistance to killing. PMN phagosomal pH was significantly higher after ingestion of nonopsonized AS-bearing E. faecalis than after that of opsonized bacteria. The novel ability of AS to promote intracellular survival of E. faecalis inside PMNs suggests that AS may be a virulence factor used by strains of E. faecalis. PMID- 10531270 TI - Augmentation of innate host defense by expression of a cathelicidin antimicrobial peptide. AB - Antimicrobial peptides, such as defensins or cathelicidins, are effector substances of the innate immune system and are thought to have antimicrobial properties that contribute to host defense. The evidence that vertebrate antimicrobial peptides contribute to innate immunity in vivo is based on their expression pattern and in vitro activity against microorganisms. The goal of this study was to investigate whether the overexpression of an antimicrobial peptide results in augmented protection against bacterial infection. C57BL/6 mice were given an adenovirus vector containing the cDNA for LL-37/hCAP-18, a human cathelicidin antimicrobial peptide. Mice treated with intratracheal LL-37/hCAP-18 vector had a lower bacterial load and a smaller inflammatory response than did untreated mice following pulmonary challenge with Pseudomonas aeruginosa PAO1. Systemic expression of LL-37/hCAP-18 after intravenous injection of recombinant adenovirus resulted in improved survival rates following intravenous injection of lipopolysaccharide with galactosamine or Escherichia coli CP9. In conclusion, the data demonstrate that expression of an antimicrobial peptide by gene transfer results in augmentation of the innate immune response, providing support for the hypothesis that vertebrate antimicrobial peptides protect against microorganisms in vivo. PMID- 10531269 TI - Phenotypic switching in Cryptococcus neoformans results in changes in cellular morphology and glucuronoxylomannan structure. AB - Cryptococcus neoformans strains exhibit variability in their capsular polysaccharide, cell morphology, karyotype, and virulence, but the relationship between these variables is poorly understood. A hypovirulent C. neoformans 24067A isolate, which usually produces smooth (SM) colony types, was found to undergo phenotypic switching and to produce wrinkled (WR) and pseudohyphal (PH) colony types at frequencies of approximately 10(-4) to 10(-5) when plated on Sabouraud agar. Cells from these colony types had large polysaccharide capsules and PH morphology, respectively. Scanning electron microscopy showed that different colony types were the result of altered cellular packing in the colony. Phenotypic switching was associated with quantitative and qualitative changes in capsular polysaccharide. Specifically, the glucuronoxylomannan (GXM) of the WR polysaccharide differed in the proportion of structural reporter groups and in increased xylose residue content linked at the 4 to 0 position. The relative virulence of the colony types was WR > PH > SM, as measured by CFU in rat lungs after intratracheal infection. Karyotype instability was observed in strain 24067A and involved primarily two chromosomes. Colonies with an alternative colony type exhibited more karyotype changes, which did not revert to the original karyotype in reverted colonies. In summary, this study revealed that phenotypic switching in C. neoformans (i) can produce WR colonies consisting of cells with either large capsule or PH morphology, (ii) is associated with production of structurally different GXM, (iii) is commonly associated with karyotype changes, (iv) can produce cells of PH morphology, and (v) can increase the virulence of a strain. Hence, phenotypic switching is an adaptive mechanism linked to virulence that can generate cell types with very different biological characteristics. PMID- 10531271 TI - CD4 T cells and major histocompatibility complex class II expression influence worm expulsion and increased intestinal muscle contraction during Trichinella spiralis infection. AB - Expulsion of intestinal nematode parasites and the associated increased contraction by intestinal muscle are T cell dependent, since both are attenuated in athymic rodents. The CD4 T-cell subset has been strongly associated with worm expulsion; however, the relationship between these cells, antigen presentation, and worm expulsion is not definitive and the role of these factors in intestinal muscle hypercontractility has not been defined. We infected C57BL/6, athymic, CD4 deficient, CD8alpha-deficient, and major histocompatibility complex class II (MHC II)-deficient (C2d) mice with Trichinella spiralis larvae. We examined intestinal worm numbers, longitudinal muscle contraction, and MHC II expression. Numerous MHC II-positive cells were identified within the muscularis externa of infected but not uninfected C57BL/6 mice. C57BL/6 and CD8alpha-deficient mice developed large increases in muscle contraction, expelling the parasite by day 21. Athymic and C2d mice exhibited much smaller increases in muscle contraction and delayed parasite expulsion. CD4-deficient mice exhibited intermediate levels of muscle contraction and delayed parasite expulsion. To further examine the role of MHC II and CD4 T cells, we irradiated C2d mice and reconstituted them with C57BL/6 bone marrow alone or with C57BL/6 CD4 T cells. C57BL/6 bone marrow alone did not affect muscle function or worm expulsion in recipient C2d mice. Partial CD4 T cell reconstitution was sufficient to restore increased muscle contraction but not worm expulsion. Thus, hematopoietic MHC II expression alone is insufficient for the development of muscle hypercontractility and worm expulsion, but the addition of even small numbers of CD4 T cells was sufficient to induce intestinal muscle pathophysiology. PMID- 10531272 TI - A new rat model of otitis media caused by Streptococcus pneumoniae: conditions and application in immunization protocols. AB - Streptococcus pneumoniae (pneumococcus [Pn]) can be cultured from up to 50% of acute otitis media (AOM) effusions, and these bacteria are the most common cause of AOM-related complications. With the recent advent of antibiotic-resistant Pn strains, treatment of Pn infections may meet with serious difficulties. Prevention through vaccination, notably for the four most common occurring Pn serotypes in humans (i.e., Pn 6B, Pn 14, Pn 19F, and Pn 23F), is a helpful alternative. Testing of vaccine efficacy should occur in an appropriate animal AOM model, which is presented here. The four involved Pn serotypes are not pathogenic to the rat, which was chosen as the experimental animal for practical reasons. To induce a natural infection (i.e., ascending through the eustachian tube), the mucociliary clearance of the eustachian tube was impaired by infusing histamine into the tympanic cavity on 2 consecutive days before intranasal inoculation of the bacteria. With this simple protocol, high and reproducible infection rates, as determined with bacterial cultures, of Pn-induced AOM (approximately 70%) with the two major Pn serotypes 14 and 19F (Pn 14 and Pn 19F) were obtained, whereas lower infection rates (25 to 50%) with Pn 6B and Pn 23F were obtained. In this model, intranasal priming with pneumococci, as well as subcutaneous vaccination with Pn 14 tetanus toxoid-conjugated polysaccharide, induced a protective effect against the induction of otitis media with these bacteria. This shows that immunity to Pn 14 AOM can be induced by both mucosal and systemic presentations of antigen. In conclusion, we have developed an animal model for Pn-induced AOM, which is suitable for the evaluation of the protecting effect of immunization. PMID- 10531273 TI - Identification and molecular analysis of the gene encoding Rickettsia typhi hemolysin. AB - Rickettsia typhi, the causative agent of murine typhus, grows directly within the host cell cytoplasm, accumulating a large number of progeny, and eventually lyses the cells. Typhus group rickettsiae (R. typhi and R. prowazekii) adhere to and lyse human and sheep erythrocytes. However, the molecular mechanism underlying erythrocyte lysis by R. typhi has not been defined. Here we describe the cloning and nucleotide sequence analysis of the gene (tlyC) encoding a hemolysin from R. typhi. DNA sequence analysis of R. typhi tlyC revealed an open reading frame of 912 bp, which encodes a protein of 304 amino acids with a predicted molecular mass of 38 kDa. To associate the R. typhi tlyC gene product with hemolytic activity, we performed complementation studies with hemolysin-negative Proteus mirabilis WPM111 (a HpmA(-) mutant of BA6163) transformed with R. typhi tlyC or R. typhi GFPuv-tlyC constructs. We demonstrated that the cloned tlyC gene conferred a hemolytic phenotype on an otherwise nonhemolytic mutant of P. mirabilis. The availability of the cloned R. typhi tlyC will permit further characterization and definition of its role in rickettsial virulence. PMID- 10531275 TI - Identification of regions of the chromosome of Neisseria meningitidis and Neisseria gonorrhoeae which are specific to the pathogenic Neisseria species. AB - Neisseria meningitidis and Neisseria gonorrhoeae give rise to dramatically different diseases. Their interactions with the host, however, do share common characteristics: they are both human pathogens which do not survive in the environment and which colonize and invade mucosa at their port of entry. It is therefore likely that they have common properties that might not be found in nonpathogenic bacteria belonging to the same genetically related group, such as Neisseria lactamica. Their common properties may be determined by chromosomal regions found only in the pathogenic Neisseria species. To address this issue, we used a previously described technique (C. R. Tinsley and X. Nassif, Proc. Natl. Acad. Sci. USA 93:11109-11114, 1996) to identify sequences of DNA specific for pathogenic neisseriae and not found in N. lactamica. Sequences present in N. lactamica were physically subtracted from the N. meningitidis Z2491 sequence and also from the N. gonorrhoeae FA1090 sequence. The clones obtained from each subtraction were tested by Southern blotting for their reactivity with the three species, and only those which reacted with both N. meningitidis and N. gonorrhoeae (i.e., not specific to either one of the pathogens) were further investigated. In a first step, these clones were mapped onto the chromosomes of both N. meningitidis and N. gonorrhoeae. The majority of the clones were arranged in clusters extending up to 10 kb, suggesting the presence of chromosomal regions common to N. meningitidis and N. gonorrhoeae which distinguish these pathogens from the commensal N. lactamica. The sequences surrounding these clones were determined from the N. meningitidis genome-sequencing project. Several clones corresponded to previously described factors required for colonization and survival at the port of entry, such as immunoglobulin A protease and PilC. Others were homologous to virulence-associated proteins in other bacteria, demonstrating that the subtractive clones are capable of pinpointing chromosomal regions shared by N. meningitidis and N. gonorrhoeae which are involved in common aspects of the host interaction of both pathogens. PMID- 10531274 TI - Pregenomic comparative analysis between bordetella bronchiseptica RB50 and Bordetella pertussis tohama I in murine models of respiratory tract infection. AB - We describe here a side-by-side comparison of murine respiratory infection by Bordetella pertussis and Bordetella bronchiseptica strains whose genomes are currently being sequenced (Tohama I and RB50, respectively). B. pertussis and B. bronchiseptica are most appropriately classified as subspecies. Their high degree of genotypic and phenotypic relatedness facilitates comparative studies of pathogenesis. RB50 and Tohama I differ in their abilities to grow in the nose, trachea, and lungs of BALB/c mice and to induce apoptosis, lung pathology, and an antibody response. To focus on the interactions between the bacteria and particular aspects of the host immune response, we used mice with specific immune defects. Mice lacking B cells and T cells were highly susceptible to B. bronchiseptica and were killed by intranasal inoculation with doses as low as 500 CFU. These mice were not killed by B. pertussis, even when doses as high as 10(5) CFU were delivered to the lungs. B. bronchiseptica, which was highly resistant to naive serum in vitro, caused bacteremia in these immunodeficient mice, while B. pertussis, which was highly sensitive to naive serum, did not cause bacteremia. B. bronchiseptica was, however, killed by immune serum in vitro, and adoptive transfer of anti-Bordetella antibodies protected SCID-beige mice from B. bronchiseptica lethal infection. Neutropenic mice were similarly killed by B. bronchiseptica but not B. pertussis infection, suggesting neutrophils are critical to the early inflammatory response to the former but not the latter. B. bronchiseptica was dramatically more active than B. pertussis in mediating the lysis of J774 cells in vitro and in inducing apoptosis of inflammatory cells in mouse lungs. This side-by-side comparison describes phenotypic differences that may be correlated with genetic differences in the comparative analysis of the genomes of these two highly related organisms. PMID- 10531277 TI - Isolation, characterization, cDNA cloning, and antimicrobial properties of two distinct subfamilies of alpha-defensins from rhesus macaque leukocytes. AB - Experiments to isolate and characterize rhesus macaque myeloid alpha-defensins (RMADs) were conducted. Seven RMAD peptides were isolated and sequenced, and the cDNAs encoding six of these peptides and one other alpha-defensin from bone marrow were also characterized. Four of the RMADs were found to be highly similar to human neutrophil alpha-defensins HNP-1 to HNP-3, while the remaining four peptides were much more similar to human enteric alpha-defensin HD-5. Two alpha defensin pairs differed only by the presence or absence of an additional arginine at the amino termini of their mature peptides, indicative of alternate posttranslational processing. The primary translation products of RMAD-1 to -8 are 94- and 96-amino-acid prepropeptides that are highly similar to those of human alpha-defensins. Immunolocalization experiments revealed a granular cytoplasmic pattern in the cytoplasms of neutrophils, indistinguishable from the pattern observed after immunostaining of human myeloid alpha-defensins in polymorphonuclear leukocytes. Each of the purified peptides was tested for its in vitro activities against Staphylococcus aureus 502a, Listeria monocytogenes EGD, Escherichia coli ML35, and Cryptococcus neoformans 271A. Several of the peptides were microbicidal for the gram-positive bacteria and C. neoformans at defensin concentrations in the range of 2 to 5 microM. All of the peptides were bacteriostatic against E. coli, but none were bactericidal for this organism. This study is the first to characterize the sequences and activities of alpha defensins from nonhuman primates, data that should aid in delineating the role of these peptides in rhesus macaque host defense. PMID- 10531276 TI - Bacterial species- and strain-dependent induction of tissue factor in human vascular endothelial cells. AB - A cardinal process in bacterial endocarditis (BE) is the activation of the clotting system and the formation of a fibrin clot on the inner surface of the heart, the so-called endocardial vegetation. The processes that lead to the activation of the clotting system on endothelial surfaces upon exposure to bacteria are largely unknown. In the present study, we investigated in an in vitro model whether infection of human endothelial cells (EC) with bacteria that are relevant to BE, such as Staphylococcus aureus, Streptococcus sanguis, and Staphylococcus epidermidis, leads to induction of tissue factor (TF)-dependent procoagulant activity (TFA) and whether this process is influenced by host factors, such as interleukin-1 (IL-1), that are produced in response to the bacteremia in vivo. The results show that S. aureus binds to and is internalized by EC, resulting in expression of TF mRNA and TF surface protein as well as generation of TFA within 4 to 8 h after infection. No TFA was found when EC were exposed to UV-irradiated S. aureus or bacterial cell wall fragments. S. sanguis and S. epidermidis, although also binding to EC, did not induce endothelial TFA. This indicates a species and strain dependency. EC also expressed TFA after exposure to IL-1. The enhanced TFA of EC after exposure to S. aureus was not prevented by IL-1 receptor antagonist, arguing against an auto- or paracrine contribution of endogenous IL-1. When IL-1 was applied together with bacteria, this had a synergistic effect on the induction of EC TFA. This was found in particular with S. aureus but also, although to a lesser degree, with S. sanguis and S. epidermidis. This influence of IL-1 on the species- and strain-dependent induction of EC TFA suggests that bacterial factors as well as host factors orchestrate the induction of coagulation in an early stage in the pathogenesis of endovascular disease, such as BE. PMID- 10531278 TI - Chlamydia trachomatis (mouse pneumonitis strain) induces cardiovascular pathology following respiratory tract infection. AB - Chlamydia, especially Chlamydia pneumoniae, infection is closely associated with human cardiovascular diseases. Thus far, however, few experimental studies have been carried out to investigate whether natural C. trachomatis infection can induce cardiovascular pathological changes. In this article, we report that pulmonary infection with C. trachomatis mouse pneumonitis strain (MoPn) can induce myocardial and perivascular inflammation and fibrosis in C57BL/6 mice. The pulmonary MoPn infection appeared to be disseminated systemically, because chlamydial antigens were readily detectable in multiple organs including the cardiovascular tissues. In addition, gamma interferon gene knockout mice with a C57BL/6 genetic background showed significant endocarditis and pancarditis characterized by vegetation in aortic valves, interstitial and pericardial inflammatory cellular infiltration, and growth of the organisms in the heart following respiratory tract MoPn infection. The results indicate that C. trachomatis can induce cardiovascular diseases following respiratory tract infection and suggest that murine MoPn respiratory tract infection may be a useful experimental model for investigating cardiovascular diseases caused by chlamydial infection. PMID- 10531279 TI - Klebsiella pneumoniae capsule expression is necessary for colonization of large intestines of streptomycin-treated mice. AB - The role of the Klebsiella pneumoniae capsular polysaccharide (K antigen) during colonization of the mouse large intestine was assessed with wild-type K. pneumoniae LM21 and its isogenic capsule-defective mutant. When bacterial strains were fed alone to mice, the capsulated bacteria persisted in the intestinal tract at levels of 10(8) CFU/g of feces while the capsule-defective strain colonized at low levels, 10(4) CFU/g of feces. In mixed-infection experiments, the mutant was rapidly outcompeted by the wild type. In situ hybridization on colonic sections revealed that bacterial cells of both strains were evenly distributed in the mucus layer at day 1 after infection, while at day 20 the wild type remained dispersed and the capsule-defective strain was seen in clusters in the mucus layer. These results suggest that capsular polysaccharide plays an important role in the gut colonization ability of K. pneumoniae. PMID- 10531280 TI - Analysis of Pneumocystis carinii introns. AB - Pneumocystis carinii is an ascomycete phylogenetically related to Schizosaccharomyces pombe. Little is known about gene regulation in P. carinii. The removal of introns from pre-mRNA requires spliceosomal recognition of the intron-exon boundary. In S. pombe and higher eukaryotes, this boundary and a branch site within the intron are conserved. We recently demonstrated that P. carinii cdc2 cDNA can complement S. pombe containing conditional mutations of cdc2, an essential gene involved in cell cycle regulation. We next tested whether P. carinii genomic cdc2 (with six introns) could also complement S. pombe cdc2 mutants and found genomic sequences incapable of this activity. Reverse transcriptase PCR confirmed the inability of the S. pombe cdc2 mutants to splice the P. carinii genomic cdc2. Analysis of 83 introns from 19 P. carinii protein encoding genes demonstrated that the sequence GTWWDW functions as a donor consensus in P. carinii, whereas YAG serves as an acceptor consensus. These sequences are similar in S. pombe; however, a branch site sequence was not found in the P. carinii genes studied. PMID- 10531281 TI - Binding of pili from uropathogenic Escherichia coli to membranes secreted by human colonocytes and enterocytes. AB - PapG adhesins mediate the binding of uropathogenic Escherichia coli. Although receptors for these adhesins have not been demonstrated in intestinal epithelia, the colonic microflora includes strains of uropathogenic E. coli. We now report that surfactant-like particles secreted by the human intestine contain receptors for PapG adhesins and may provide an intestinal habitat for uropathogenic bacteria. PMID- 10531282 TI - Cell death of human polymorphonuclear neutrophils induced by a Pseudomonas aeruginosa cystic fibrosis isolate requires a functional type III secretion system. AB - With a coincubation model incorporating Pseudomonas aeruginosa and human polymorphonuclear neutrophils (PMNs), a cystic fibrosis (CF) P. aeruginosa isolate has been shown to resist the bactericidal action of PMNs and to induce their cellular death. An isogenic mutant of this CF isolate in which the type III secretion system was rendered nonfunctional was unable to induce cellular death of PMNs. PMID- 10531283 TI - Effect of mutS and recD mutations on Salmonella virulence. AB - Hybrid derivatives of closely related bacteria may be used to dissect strain specific functions that contribute to virulence within a host. However, mismatches between DNA sequences are a potent barrier to recombination. Recipients with mutS and recD mutations overcome this barrier, allowing construction of genetic hybrids. To determine whether Salmonella hybrids constructed in a mutS recD host can be used to study virulence, we assayed the effect of mutS and recD mutations on the virulence of Salmonella typhimurium 14028s in mice. Mutants defective in either mutS or recD do not affect the time course or the 50% lethal dose (LD(50)) of the infection. In contrast, the inactivation of both mutS and recD results in a synthetic phenotype which substantially increases the time required to cause a lethal infection without changing the LD(50). This phenotype results from an inability of mutS recD double mutants to rapidly adapt to purine-limiting conditions present within macrophages. Although the disease progression is slower, S. typhimurium mutS recD mutants retain the ability to cause lethal infections, and, thus, hybrids constructed in mutS recD hosts may permit the analysis of virulence factors in a surrogate animal model. PMID- 10531284 TI - Role of the amino-terminal region of Porphyromonas gingivalis fimbriae in adherence to epithelial cells. AB - Porphyromonas gingivalis fimbriae elicit many responses in eukaryotic cells, including mitogenicity, cytokine production, epithelial cell invasion, and cellular immune response. Specific domains of the major fimbrial protein (FimA) have been shown to be important in triggering some of these functions. The goal of the present study was to identify the domain(s) of P. gingivalis FimA responsible for specific interaction with human mucosal epithelial cells. Fimbriated P. gingivalis strains have been shown to bind to buccal epithelial cells, whereas nonfimbriated strains bind at low levels or not at all. This and other studies provide evidence that FimA mediates the adherence of P. gingivalis to oral epithelial cells. To determine the specific region(s) of P. gingivalis FimA involved in epithelial cell binding, specific antipeptide antibodies were used to inhibit the binding of iodinated purified fimbriae as well as the binding of P. gingivalis cells to epithelial cells. Antibodies directed against peptides 49 to 68 (VVMANTAGAMELVGKTLAEVK) and 69 to 90 (ALTTELTAENQEAAGLIMTAEP) were found to highly inhibit both the binding of fimbriae and the binding of P. gingivalis cells to epithelial cells. The antibody against FimA peptides 69 to 90 also reacted with P. gingivalis fimbriae in immunogold labeling and immunoblot analysis, thereby indicating that this peptide domain is exposed on the surface of fimbriae. Our results suggest that the amino-terminal domain corresponding to amino acid residues 49 to 90 of the fimbrillin protein is a major epithelial cell binding domain of P. gingivalis fimbriae. PMID- 10531285 TI - Modulation of B-lymphocyte and NK cell activities by glycoinositolphospholipid purified from Trypanosoma cruzi. AB - Glycoinositolphospholipids (GIPLs) are some of the major glycolipids of the Trypanosoma cruzi surface that were previously shown to activate B cells. In the present study, we investigated whether (i) T. cruzi GIPLs could induce immunoglobulin secretion from B cells in the absence of T cells and NK cells and whether (ii) NK cells are also stimulated by the GIPLs. B cells purified from mice deficient in both T and NK cells (CD3epsilon transgenic mice) secreted immunoglobulin in response to the GIPL. This response was increased by coculture with a murine NK cell line. The T. cruzi GIPL also increased the NK cell (interleukin-2 induced) proliferative response. Our data indicate that the T. cruzi GIPL has a direct stimulatory effect on NK cells and induces immunoglobulin secretion in the absence of T lymphocytes and NK cells. These findings suggest that this T. cruzi-derived molecule may be one of the stimulators that lead to NK cell activation during T. cruzi infection. PMID- 10531286 TI - The outer membrane of Brucella ovis shows increased permeability to hydrophobic probes and is more susceptible to cationic peptides than are the outer membranes of mutant rough Brucella abortus strains. AB - The permeability of the outer membrane (OM) to hydrophobic probes and its susceptibility to bactericidal cationic peptides were investigated for natural rough Brucella ovis and for mutant rough Brucella abortus strains. The OM of B. ovis displayed an abrupt and faster kinetic profile than rough B. abortus during the uptake of the hydrophobic probe N-phenyl-naphthylamine. B. ovis was more sensitive than rough B. abortus to the action of cationic peptides. Bactenecins 5 and 7 induced morphological alterations on the OMs of both rough Brucella strains. B. ovis lipopolysaccharide (LPS) captured considerably more polymyxin B than LPSs from both rough and smooth B. abortus strains. Polymyxin B, poly-L lysine, and poly-L-ornithine produced a thick coating on the surfaces of both strains, which was more evident in B. ovis than in rough B. abortus. The distinct functional properties of the OMs of these two rough strains correlate with some structural differences of their OMs and with their different biological behaviors in animals and culture cells. PMID- 10531287 TI - Purification and immunoreactivity of three components from the 30/32-kilodalton antigen 85 complex in Mycobacterium tuberculosis. AB - The three proteins of the antigen 85 complex (85A, 85B, and 85C), which are major secretory products of Mycobacterium tuberculosis, were purified to homogeneity in large amounts by a combination of chromatography on hydroxylapatite, DEAE Sepharose, and DEAE-Sephacel and gel filtration from M. tuberculosis culture filtrate. Then we examined the immunological reactivity of the three proteins in tuberculosis patients and healthy controls. Antibody responses to the 85B and 85A proteins in patients were significantly greater than responses to the 85C protein. In contrast, all three antigens induced significant lymphoproliferation and gamma interferon production in peripheral blood mononuclear cells from healthy tuberculin reactors. PMID- 10531288 TI - Syntheses and immunologic properties of Escherichia coli O157 O-specific polysaccharide and Shiga toxin 1 B subunit conjugates in mice. AB - Escherichia coli O157 is the major cause of diarrhea-associated hemolytic uremic syndrome (HUS). Strains causing HUS contain either Shiga toxin 1 (Stx1) or Stx2, or both. In adult volunteers, conjugate vaccines of detoxified lipopolysaccharide (LPS) elicited bactericidal antibodies to E. coli O157. Here, the detoxified LPS was conjugated with improved schemes to the nontoxic B subunit of Stx1. Mice injected with these bivalent conjugates elicited both bactericidal antibodies to E. coli O157 and neutralization antibodies to Stx1. PMID- 10531289 TI - Homotypic and heterotypic antibody responses to a 56-kilodalton protein of Orientia tsutsugamushi. AB - We analyzed homotypic and heterotypic antibody responses to a type-specific antigen (Tsa), a 56-kDa protein of Orientia tsutsugamushi, by using sera from mice immunized with strains Gilliam, Karp, Kato, and Boryong. We generated a series of deletion constructs of the tsa gene and expressed them as MalE fusion proteins. Variable domain I (VD I) showed strong responses to homotypic antibodies. Antigenic domain II (AD II) from Boryong and Karp showed cross reactivities to each other. VD III showed no responses to any of the antibodies. Sera from Kato-immunized mice showed only homotypic responses to AD III. On the other hand, sera of the mice immunized with Gilliam, Karp, or Boryong showed homotypic as well as heterotypic responses to this region. VD IV showed the strongest heterotypic antibody responses among the fragments tested. These data suggest that VD I is important in homotypic antibody responses and that AD II, AD III, and VD IV are important in heterotypic antibody responses of the mice to Tsa. PMID- 10531290 TI - Superantigen immune stimulation evokes epithelial monocyte chemoattractant protein 1 and RANTES production. AB - Bacterial superantigens (SAgs) have been implicated in inflammatory disease, and SAg-treated mice have increased jejunal T cells. Here we show that T84 cells (a human epithelial cell line) display increased MCP-1 and RANTES mRNA expression and protein production in response to conditioned medium from Staphylococcus aureus enterotoxin B (SEB; a model SAg)-activated immune cells. Also, MCP-1 and RANTES mRNAs were increased in jejunal enterocytes isolated from SEB-treated mice. We suggest that T-cell recruitment to the gut following SAg immune activation could be partially due to epithelium-derived chemokines. PMID- 10531291 TI - Inhibition of neutrophil apoptosis by verotoxin 2 derived from Escherichia coli O157:H7. AB - In order to evaluate the pathological role of verotoxin 2 (VT2), we investigated the effects of VT2 on neutrophil apoptosis in vitro. The results showed that VT2 caused a significant delay in spontaneous neutrophil apoptosis and that the effect was abrogated by a protein kinase C inhibitor. These data indicate that longer survival of neutrophils may aggravate neutrophil-mediated tissue damage in VT2-associated diseases. PMID- 10531292 TI - Urease-based mucosal immunization against Helicobacter heilmannii infection induces corpus atrophy in mice. AB - Mucosal immunization with Helicobacter heilmannii urease B or Helicobacter pylori urease, given nasally with cholera toxin, protects BALB/c mice against H. heilmannii infection and significantly reduces a preexisting infection. However, immunization aggravates gastric corpus atrophy. Our results underline the necessity of defining immunization regimens that do not enhance mucosal damage. PMID- 10531293 TI - Anatomic segmentation of the intestinal immune response in nonhuman primates: differential distribution of B cells after oral and rectal immunizations to sites defined by their source of vascularization. AB - We show that the distribution of specific antibodies and antibody-secreting cells in the intestine after oral and rectal immunizations corresponds to the vascularization and lymph drainage patterns of the gut. Oral immunizations induce antibody responses along the parts of the intestine connected to the superior mesenteric vessels and lymph ducts, whereas rectal immunizations induce antibody responses along the parts of the intestine associated with the inferior mesenteric vessels and ducts. PMID- 10531294 TI - P48 major surface antigen of Mycoplasma agalactiae is homologous to a malp product of Mycoplasma fermentans and belongs to a selected family of bacterial lipoproteins. AB - A major surface antigenic lipoprotein of Mycoplasma agalactiae, promptly recognized by the host's immune system, was characterized. The mature product, P48, showed significant similarity and shared conserved amino acid motifs with lipoproteins or predicted lipoproteins from Mycoplasma fermentans, Mycoplasma hyorhinis, relapsing fever Borrelia spp., Bacillus subtilis, and Treponema pallidum. PMID- 10531295 TI - Identification of genes coding for exported proteins of Actinobacillus actinomycetemcomitans. AB - Random fusions of genomic DNA fragments to a partial gene encoding a signal sequence-deficient bacterial alkaline phosphatase were utilized to screen for exported proteins of Actinobacillus actinomycetemcomitans in Escherichia coli. Twenty-four PhoA(+) clones were isolated and sequenced. Membrane localization signals in the form of signal sequences were deduced from most of these sequences. Several of the deduced amino acid sequences were found to be homologous to known exported or membrane-associated proteins. The complete genes corresponding to two of these sequences were isolated from an A. actinomycetemcomitans lambda phage library. One gene was found to be homologous to the outer membrane lipoprotein LolB. The second gene product had homology with a Haemophilus influenzae protein and was localized to the inner membrane of A. actinomycetemcomitans. PMID- 10531296 TI - Differential expression of caprine beta-defensins in digestive and respiratory tissues. AB - We identified two novel beta-defensin precursors, preproGBD-1 and preproGBD-2, in the tissues of a goat. Although the precursors were identical in 96.8% of their bases and 88.2% (60 of 68) of their amino acids, preproGBD-1 was expressed principally in the tongue and respiratory tract, whereas preproGBD-2 expression predominated throughout the intestine. These findings exemplify the phenomenon of tissue-specific expression in a family of host defense peptides that arose before the avian and mammalian lineages diverged. PMID- 10531297 TI - Histone H1 and evolution of sperm nuclear basic proteins. PMID- 10531298 TI - p21-activated protein kinase gamma-PAK is activated by ionizing radiation and other DNA-damaging agents. Similarities and differences to alpha-PAK. AB - The p21-activated protein kinase gamma-PAK is activated 2-5-fold in response to ionizing radiation (IR) in 3T3-L1 fibroblasts and U937 leukemia cells. gamma-PAK is activated in a dose- and time-dependent manner. Doses from 1 to 100 Gy result in significant stimulation of activity at 30 min, whereas maximal stimulation is observed at 120 min after irradiation. UV (80 J/m(2)) and the DNA-damaging drugs cytosine beta-D-arabinofuranoside (AraC) and cis-platinum(II)diammine dichloride (cisplatin) also induce gamma-PAK activation. The activation of gamma-PAK in response to IR or AraC is dependent on tyrosine kinase and phosphoinositide 3 kinase activity, as demonstrated by use of the inhibitors genistein and wortmannin; in contrast activation of gamma-PAK by cisplatin and UV is not affected significantly by these inhibitors, suggesting that gamma-PAK can be activated by more than one pathway in response to different types of DNA damage. In contrast to gamma-PAK, alpha-PAK and JNK are activated only by cisplatin and UV in 3T3-L1 cells, suggesting differential regulation of the protein kinases. This is the first time that members of the Ste20/PAK family of protein kinases have been shown to be involved in the cellular response to IR and other DNA damaging agents. PMID- 10531299 TI - Correction of defective protein trafficking of a mutant HERG potassium channel in human long QT syndrome. Pharmacological and temperature effects. AB - The chromosome 7-linked form of congenital long QT syndrome (LQT2) is caused by mutations in the human ether-a-go-go-related gene (HERG) that encodes the rapidly activating delayed rectifier potassium channel. One mechanism for the loss of normal channel function in LQT2 is defective protein trafficking, which results in the failure of the channel protein to reach the plasma membrane. Here we show that the N470D LQT2 mutant protein is trafficking-deficient when expressed at 37 degrees C in HEK293 cells, whereas at 27 degrees C its trafficking to the plasma membrane and channel function are markedly improved. We further show that the antiarrhythmic drug E-4031, which selectively blocks HERG channels, also corrects defective protein trafficking of the N470D mutant and can restore the generation of HERG current. Similar findings were obtained with the drugs astemizole and cisapride, as well as with high concentrations of glycerol. The effect of E-4031 on HERG protein trafficking was concentration-dependent and required low drug concentrations (saturation present at 5 microM), developed rapidly with drug exposure, and occurred post-translationally. These findings suggest that protein misfolding leading to defective trafficking of some HERG LQT mutations may be corrected by specific pharmacological strategies. PMID- 10531300 TI - Sensing of ionizing radiation-induced DNA damage by ATM through interaction with histone deacetylase. AB - The ATM gene is mutated in individuals with ataxia telangiectasia, a human genetic disease characterized by extreme sensitivity to radiation. The ATM protein acts as a sensor of radiation-induced cellular damage and contributes to cell cycle regulation, signal transduction, and DNA repair; however, the mechanisms underlying these functions of ATM remain largely unknown. Binding and immunoprecipitation assays have now shown that ATM interacts with the histone deacetylase HDAC1 both in vitro and in vivo, and that the extent of this association is increased after exposure of MRC5CV1 human fibroblasts to ionizing radiation. Histone deacetylase activity was also detected in immunoprecipitates prepared from these cells with antibodies to ATM, and this activity was blocked by the histone deacetylase inhibitor trichostatin A. These results suggest a previously unanticipated role for ATM in the modification of chromatin components in response to ionizing radiation. PMID- 10531301 TI - A new 30-kDa ubiquitin-related SUMO-1 hydrolase from bovine brain. AB - SUMO-1 is a ubiquitin-like protein functioning as an important reversible protein modifier. To date there is no report on a SUMO-1 hydrolase/isopeptidase catalyzing the release of SUMO-1 from its precursor or SUMO-1-ligated proteins in mammalian tissues. Here we found multiple activities that cleave the SUMO-1 moiety from two model substrates, (125)I-SUMO-1-alphaNH-HSTVGSMHISPPEPESEEEEEHYC and/or GST-SUMO-1-(35)S-RanGAP1 conjugate, in bovine brain extracts. Of them, a major SUMO-1 C-terminal hydrolase had been partially purified by successive chromatographic operations. The enzyme had the ability to cleave SUMO-1 not only from its precursor but also from a SUMO-1-ligated RanGAP1 but did not exhibit any significant cleavage of the ubiquitin- and NEDD8-precursor. The activity of SUMO 1 hydrolase was almost completely inhibited by N-ethylmaleimide, but not by phenylmethanesulfonyl fluoride, EDTA, and ubiquitin-aldehyde known as a potent inhibitor of deubiquitinylating enzymes. Intriguingly, the apparent molecular mass of the isolated SUMO-1 hydrolase was approximately 30 kDa, which is significantly smaller than the recently identified yeast Smt3/SUMO-1 specific protease Ulp1. These results indicate that there are multiple SUMO-1 hydrolase/isopeptidases in mammalian cells and that the 30-kDa small SUMO-1 hydrolase plays a central role in processing of the SUMO-1-precursor. PMID- 10531302 TI - The cytoplasmic coatomer protein COPI. A potential translational regulator. AB - Expression of the asialoglycoprotein receptor (ASGR) by the human hepatocellular carcinoma cell lines HepG2 and HuH-7 in response to intracellular cGMP concentrations was previously shown to be regulated at the translational level (1). Stable transfection of COS-7 cells with deletion constructs encoding the asialoglycoprotein receptor H2b subunit localized the cGMP-responsive cis-acting element to the mRNA 5'-untranslated region. Resolution by anion exchange chromatography of an S-100 isolated from human liver resulted in the partial purification of an RNA-binding protein specific to this cis-acting element. Northwestern analysis using the 5'-untranslated region as probe indicated that a 140-kDa protein was the potential RNA-binding protein. Sequence of tryptic peptides suggested that the 140-kDa protein was the alpha-COP subunit of coatomer protein COPI, usually associated with trans-Golgi network membrane traffic. Immunoblot analysis confirmed the presence of alpha-COP in the Mono-Q fraction as well as that of a second coatomer subunit, beta-COP. Antibody induced gel retardation supershift confirmed the identification of the RNA-binding proteins as alpha- and beta-COP. Although the RNA recognition motif appears to reside solely in alpha-COP, antibody-induced supershift strongly indicated that the entire coatomer complex was the trans-acting factor. Depletion of S-100 with the antibody to beta-COP confirmed that the coatomer was the sole protein binding to the ASGR mRNA 5'-untranslated region in liver cytosol and responsible for inhibition of in vitro translation of the asialoglycoprotein receptor. PMID- 10531303 TI - Activation of the unfolded protein response pathway induces human asparagine synthetase gene expression. AB - The gene for the amino acid biosynthetic activity asparagine synthetase (AS) is induced by both amino acid and glucose deprivation of cells. The data reported here document that the human AS gene is induced following activation of the Unfolded Response Pathway (UPR), also known as the Endoplasmic Reticulum Stress Response (ERSR) in mammals. Increased AS transcription occurs in response to glucose deprivation, tunicamycin, or azetidine-2-carboxylate, all known to activate the UPR/ERSR pathway. Previously identified ERSR target genes contain multiple copies of a single highly conserved cis-element. In contrast, the human AS gene does not contain the ERSR element, as it has been described for other responsive genes. Instead, AS induction requires an Sp1-like sequence, a sequence previously shown to be associated with amino acid control of transcription, and possibly, a third region containing no consensus sequences for known transcription factors. Oligonucleotides covering each of these regions form DNA protein complexes in vitro, and for some the amount of these complexes is greater when nuclear extracts from glucose-starved cells are tested. These results document that a wider range of metabolic activities are activated by the UPR/ERSR pathway than previously recognized and that genomic elements other than those already described can serve to enhance transcription of specific target genes. PMID- 10531304 TI - The N terminus of the cardiac L-type Ca(2+) channel alpha(1C) subunit. The initial segment is ubiquitous and crucial for protein kinase C modulation, but is not directly phosphorylated. AB - The first 46 amino acids (aa) of the N terminus of the rabbit heart (RH) L-type cardiac Ca(2+) channel alpha(1C) subunit are crucial for the stimulating action of protein kinase C (PKC) and also hinder channel gating (Shistik, E., Ivanina, T., Blumenstein, Y., and Dascal, N. (1998) J. Biol. Chem. 273, 17901-17909). The mechanism of PKC action and the location of the PKC target site are not known. Moreover, uncertainties in the genomic sequence of the N-terminal region of alpha(1C) leave open the question of the presence of RH-type N terminus in L-type channels in mammalian tissues. Here, we demonstrate the presence of alpha(1C) protein containing an RH-type initial N-terminal segment in rat heart and brain by using a newly prepared polyclonal antibody. Using deletion mutants of alpha(1C) expressed in Xenopus oocytes, we further narrowed down the part of the N terminus crucial for both inhibitory gating and for PKC effect to the first 20 amino acid residues, and we identify the first 5 aa as an important determinant of PKC action and of N-terminal effect on gating. The absence of serines and threonines in the first 5 aa and the absence of phosphorylation by PKC of a glutathione S-transferase-fusion protein containing the initial segment suggest that the effect of PKC does not arise through a direct phosphorylation of this segment. We propose that PKC acts by attenuating the inhibitory action of the N terminus via phosphorylation of a remote site, in the channel or in an auxiliary protein, that interacts with the initial segment of the N terminus. PMID- 10531305 TI - Quinone reductase inhibitors block SAPK/JNK and NFkappaB pathways and potentiate apoptosis. AB - A variety of environmental stresses stimulate the mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) kinase (MEKK) > stress activated protein kinase (SAPK)-ERK kinase (SEK) > SAPK/c-Jun NH(2)-terminal kinase (JNK) stress-activated protein kinase cascade and coordinately activate the transcription factor NFkappaB. Mechanisms of stress activation upstream of MEKK1 have not been precisely determined. Redox mechanisms involving sulfhydryls are likely because N-acetyl-cysteine at millimolar concentrations blocks stress signals. Because intracellular sulfhydryl concentrations can be regulated through redox cycling involving reactive quinones (1), we tested the ability of quinone reductase inhibitors to alter stress signaling. Several quinone reductases are inhibited by dicoumarol, a coumarin derivative. Dicoumarol prevented SAPK activation in vivo by chemical cell stressors and also prevented SAPK activation induced by expression of the tumor necrosis factor alpha (TNFalpha) receptor associated protein TRAF2 but not by expression of truncated active MEKK1. Other coumarin derivatives failed to block SAPK activation, but other inhibitors of quinone reductases, particularly menadione, similarly blocked SAPK activation. Cells deficient in a major quinone reductase, NQO1, displayed hypersensitivity to dicoumarol stress inhibition, whereas SAPK in cells reconstituted with the NQO1 gene displayed relative dicoumarol resistance. Consistent with the proposed role of overlapping upstream signaling cascades in activation of NFkappaB, dicoumarol also blocked NFkappaB activation in primary macrophages stimulated with either lipopolysaccharide or TNFalpha. In addition, dicoumarol strongly potentiated TNFalpha-induced apoptosis in HeLa cells, probably by blocking the anti-apoptotic effect of NFkappaB. The ability of dicoumarol to simultaneously inhibit SAPK and NFkappaB activation and to potentiate apoptotic cell death suggests that SAPK is not an obligate participant in apoptosis. Dicoumarol, currently in clinical use as an oral anticoagulant, represents a potential therapeutic inhibitor of the SAPK and NFkappaB response. PMID- 10531306 TI - Dual mechanisms for glucose 6-phosphate inhibition of human brain hexokinase. AB - Brain hexokinase (HKI) is inhibited potently by its product glucose 6-phosphate (G6P); however, the mechanism of inhibition is unsettled. Two hypotheses have been proposed to account for product inhibition of HKI. In one, G6P binds to the active site (the C-terminal half of HKI) and competes directly with ATP, whereas in the alternative suggestion the inhibitor binds to an allosteric site (the N terminal half of HKI), which indirectly displaces ATP from the active site. Single mutations within G6P binding pockets, as defined by crystal structures, at either the N- or C-terminal half of HKI have no significant effect on G6P inhibition. On the other hand, the corresponding mutations eliminate product inhibition in a truncated form of HKI, consisting only of the C-terminal half of the enzyme. Only through combined mutations at the active and allosteric sites, using residues for which single mutations had little effect, was product inhibition eliminated in HKI. Evidently, potent inhibition of HKI by G6P can occur from both active and allosteric binding sites. Furthermore, kinetic data reported here, in conjunction with published equilibrium binding data, are consistent with inhibitory sites of comparable affinity linked by a mechanism of negative cooperativity. PMID- 10531307 TI - Decay accelerating activity of complement receptor type 1 (CD35). Two active sites are required for dissociating C5 convertases. AB - The goal of this study was to identify the site(s) in CR1 that mediate the dissociation of the C3 and C5 convertases. To that end, truncated derivatives of CR1 whose extracellular part is composed of 30 tandem repeating modules, termed complement control protein repeats (CCPs), were generated. Site 1 (CCPs 1-3) alone mediated the decay acceleration of the classical and alternative pathway C3 convertases. Site 2 (CCPs 8-10 or the nearly identical CCPs 15-17) had one-fifth the activity of site 1. In contrast, for the C5 convertase, site 1 had only 0.5% of the decay accelerating activity, while site 2 had no detectable activity. Efficient C5 decay accelerating activity was detected in recombinants that carried both site 1 and site 2. The activity was reduced if the intervening repeats between site 1 and site 2 were deleted. The results indicate that, for the C5 convertases, decay accelerating activity is mediated primarily by site 1. A properly spaced site 2 has an important auxiliary role, which may involve its C3b binding capacity. Moreover, using homologous substitution mutagenesis, residues important in site 1 for dissociating activity were identified. Based on these results, we generated proteins one-fourth the size of CR1 but with enhanced decay accelerating activity for the C3 convertases. PMID- 10531308 TI - Identification of a prothoracicostatic peptide in the larval brain of the silkworm, Bombyx mori. AB - Prothoracicotropic hormone (PTTH) stimulates ecdysteroid biosynthesis in the prothoracic gland (PG) of insects. A peptide inhibiting ecdysteroid biosynthesis in the PG was isolated from the extracts of 2,000 larval brains of the silkworm, Bombyx mori, using a protocol that included four reversed-phase high performance liquid chromatography procedures. The primary structure of this prothoracicostatic peptide (Bom-PTSP) was determined to be H-Ala-Trp-Gln-Asp-Leu Asn-Ser-Ala-Trp-NH(2). This neuropeptide has the same sequence as Mas-MIP-I, a myoinhibitory peptide previously isolated from the ventral nerve cord of the tobacco hornworm, Manduca sexta, and is highly homologous with the N-terminal portion of vertebrate peptides of the galanin family. This peptide inhibited PTTH stimulated ecdysteroidogenesis in the PG at both the spinning and feeding stages, which indicates that Bom-PTSP interferes with PTTH-stimulated ecdysteroidogenesis. PMID- 10531309 TI - Discriminating between capacitative and arachidonate-activated Ca(2+) entry pathways in HEK293 cells. AB - We have recently questioned whether the capacitative or store-operated model for receptor-activated Ca(2+) entry can account for the influx of Ca(2+) seen at low agonist concentrations, such a those typically producing [Ca(2+)](i) oscillations. Instead, we have identified an arachidonic acid-regulated, noncapacitative Ca(2+) entry mechanism that appears to be specifically responsible for the receptor-activated entry of Ca(2+) under these conditions. However, it is unclear whether these two systems reflect the activity of distinct entry pathways or simply different mechanisms of regulating a common pathway. We therefore used the known selectivity of the Ca(2+)-stimulated type VIII adenylyl cyclase for Ca(2+) entry occurring via the capacitative pathway (Fagan, K. A., Mahey, R., and Cooper, D. M. F. (1996) J. Biol. Chem. 271, 12438-12444) to attempt to discriminate between these two entry mechanisms in HEK293 cells. Consistent with the earlier reports, we found that thapsigargin induced an approximate 3-fold increase in adenylyl cyclase activity that was unrelated to global changes in [Ca(2+)](i) or to the release of Ca(2+) from internal stores but was specifically dependent on the induced capacitative entry of Ca(2+). In marked contrast, the arachidonate-induced entry of Ca(2+) completely failed to affect adenylyl cyclase activity despite producing a substantially greater rate of entry than that induced by thapsigargin. These data demonstrate that the arachidonate-activated entry of Ca(2+) occurs via an entirely distinct influx pathway. PMID- 10531310 TI - Insulin receptor substrate-4 enhances insulin-like growth factor-I-induced cell proliferation. AB - The insulin receptor substrates (IRSs)-1-4 play important roles in signal transduction emanating from the insulin and insulin-like growth factor (IGF)-I receptors. IRS-4 is the most recently characterized member, which has been found primarily in human cells and tissues. It interacts with SH2-containing proteins such as phosphatidylinositol 3'-kinase (PI3K), Grb2, Crk-II, and CrkL. In this study, we transfected IRS-4 in mouse NIH-3T3 cells that overexpress IGF-I receptors. Clones expressing IRS-4 showed enhanced cellular proliferation when cells were cultured in 1% fetal bovine serum without added IGF-I. Addition of IGF I enhanced cellular proliferation in cells overexpressing the IGF-I receptor alone but had an even greater proliferative effect in cells overexpressing both the IGF-I receptors and IRS-4. When etoposide and methylmethane sulfonate (MMS), both DNA damaging agents, were added to the cells, they uniformly induced cell cycle arrest. Fluorescence-activated cell sorter analysis demonstrated that the arrest of the cell cycle occurred at the G(1) checkpoint, and furthermore no significant degree of apoptosis was demonstrated with the use of either agent. In cells, overexpressing IGF-I receptors alone, IGF-I addition enhanced cellular proliferation, even in the presence of etoposide and MMS. In cells overexpressing IGF-I receptors and IRS-4, the effect of IGF-I in overcoming the cell cycle arrest was even more pronounced. These results suggest that IRS-4 is implicated in the IGF-I receptor mitogenic signaling pathway. PMID- 10531311 TI - Mitochondrial nitric-oxide synthase stimulation causes cytochrome c release from isolated mitochondria. Evidence for intramitochondrial peroxynitrite formation. AB - Nitric oxide (NO) is synthesized by members of the NO synthase (NOS) family. Recently the existence of a mitochondrial NOS (mtNOS), its Ca(2+) dependence, and its relevance for mitochondrial bioenergetics was reported (Ghafourifar, P., and Richter, C. (1997) FEBS Lett. 418, 291-296; Giulivi, C., Poderoso, J. J., and Boveris, A. (1998) J. Biol. Chem. 273, 11038-11043). Here we report on the possible involvement of mtNOS in apoptosis. We show that uptake of Ca(2+) by mitochondria triggers mtNOS activity and causes the release of cytochrome c from isolated mitochondria in a Bcl-2-sensitive manner. mtNOS-induced cytochrome c release was paralleled by increased lipid peroxidation. The release of cytochrome c as well as increase in lipid peroxidation were prevented by NOS inhibitors, a superoxide dismutase mimic, and a peroxynitrite scavenger. We show that mtNOS induced cytochrome c release is not mediated via the mitochondrial permeability transition pore because the release was aggravated by cyclosporin A and abolished by blockade of mitochondrial calcium uptake by ruthenium red. We conclude that, upon Ca(2+)-induced mtNOS activation, peroxynitrite is formed within mitochondria, which causes the release of cytochrome c from isolated mitochondria, and we propose a mechanism by which elevated Ca(2+) levels induce apoptosis. PMID- 10531312 TI - Catalytic mechanism of the tryptophan synthase alpha(2)beta(2) complex. Effects of pH, isotopic substitution, and allosteric ligands. AB - The mechanism of the tryptophan synthase alpha(2)beta(2) complex from Salmonella typhimurium is explored by determining the effects of pH, of temperature, and of isotopic substitution on the pyridoxal phosphate-dependent reaction of L-serine with indole to form L-tryptophan. The pH dependence of the kinetic parameters indicates that three ionizing groups are involved in substrate binding and catalysis with pK(a)1 = 6.5, pK(a)2 = 7.3, and pK(a)3 = 8.2-9. A significant primary isotope effect (approximately 3.5) on V and V/K is observed at low pH (pH 7), but not at high pH (pH 9), indicating that the base that accepts the alpha proton (betaLys-87) is protonated at low pH, slowing the abstraction of the alpha proton and making this step at least partially rate-limiting. pK(a)2 is assigned to betaLys-87 on the basis of the kinetic isotope effect results and of the observation that the competitive inhibitors glycine and oxindolyl-L-alanine display single pK(i) values of 7.3. The residue with this pK(a) (betaLys-87) must be unprotonated for binding glycine or oxindolyl-L-alanine, and, by inference, L serine. Investigations of the temperature dependence of the pK(a) values support the assignment of pK(a)2 to betaLys-87 and suggest that the ionizing residue with pK(a)1 could be a carboxylate, possibly betaAsp-305, and that the residue associated with a conformational change at pK(a)3 may be betaLys-167. The occurrence of a closed to open conformational conversion at high pH is supported by investigations of the effects of pH on reaction specificity and on the equilibrium distribution of enzyme-substrate intermediates. PMID- 10531313 TI - On the diversity of secreted phospholipases A(2). Cloning, tissue distribution, and functional expression of two novel mouse group II enzymes. AB - Over the last decade, an expanding diversity of secreted phospholipases A(2) (sPLA(2)s) has been identified in mammals. Here, we report the cloning in mice of three additional sPLA(2)s called mouse group IIE (mGIIE), IIF (mGIIF), and X (mGX) sPLA(2)s, thus giving rise to eight distinct sPLA(2)s in this species. Both mGIIE and mGIIF sPLA(2)s contain the typical cysteines of group II sPLA(2)s, but have relatively low levels of identity (less than 51%) with other mouse sPLA(2)s, indicating that these enzymes are novel group II sPLA(2)s. However, a unique feature of mGIIF sPLA(2) is the presence of a C-terminal extension of 23 amino acids containing a single cysteine. mGX sPLA(2) has 72% identity with the previously cloned human group X (hGX) sPLA(2) and displays similar structural features, making it likely that mGX sPLA(2) is the ortholog of hGX sPLA(2). Genes for mGIIE and mGIIF sPLA(2)s are located on chromosome 4, and that of mGX sPLA(2) on chromosome 16. Northern and dot blot experiments with 22 tissues indicate that all eight mouse sPLA(2)s have different tissue distributions, suggesting specific functions for each. mGIIE sPLA(2) is highly expressed in uterus, and at lower levels in various other tissues. mGIIF sPLA(2) is strongly expressed during embryogenesis and in adult testis. mGX sPLA(2) is mostly expressed in adult testis and stomach. When the cDNAs for the eight mouse sPLA(2)s were transiently transfected in COS cells, sPLA(2) activity was found to accumulate in cell medium, indicating that each enzyme is secreted and catalytically active. Using COS cell medium as a source of enzymes, pH rate profile and phospholipid headgroup specificity of the novel sPLA(2)s were analyzed and compared with the other mouse sPLA(2)s. PMID- 10531314 TI - Conversion of aspartate aminotransferase into an L-aspartate beta-decarboxylase by a triple active-site mutation. AB - The conjoint substitution of three active-site residues in aspartate aminotransferase (AspAT) of Escherichia coli (Y225R/R292K/R386A) increases the ratio of L-aspartate beta-decarboxylase activity to transaminase activity >25 million-fold. This result was achieved by combining an arginine shift mutation (Y225R/R386A) with a conservative substitution of a substrate-binding residue (R292K). In the wild-type enzyme, Arg(386) interacts with the alpha-carboxylate group of the substrate and is one of the four residues that are invariant in all aminotransferases; Tyr(225) is in its vicinity, forming a hydrogen bond with O-3' of the cofactor; and Arg(292) interacts with the distal carboxylate group of the substrate. In the triple-mutant enzyme, k(cat)' for beta-decarboxylation of L aspartate was 0.08 s(-1), whereas k(cat)' for transamination was decreased to 0.01 s(-1). AspAT was thus converted into an L-aspartate beta-decarboxylase that catalyzes transamination as a side reaction. The major pathway of beta decarboxylation directly produces L-alanine without intermediary formation of pyruvate. The various single- or double-mutant AspATs corresponding to the triple mutant enzyme showed, with the exception of AspAT Y225R/R386A, no measurable or only very low beta-decarboxylase activity. The arginine shift mutation Y225R/R386A elicits beta-decarboxylase activity, whereas the R292K substitution suppresses transaminase activity. The reaction specificity of the triple-mutant enzyme is thus achieved in the same way as that of wild-type pyridoxal 5' phosphate-dependent enzymes in general and possibly of many other enzymes, i.e. by accelerating the specific reaction and suppressing potential side reactions. PMID- 10531315 TI - Ubiquinone at center N is responsible for triphasic reduction of cytochrome b in the cytochrome bc(1) complex. AB - We have examined the pre-steady state reduction kinetics of the Saccharomyces cerevisiae cytochrome bc(1) complex by menaquinol in the presence and absence of endogenous ubiquinone to elucidate the mechanism of triphasic cytochrome b reduction. With cytochrome bc(1) complex from wild type yeast, cytochrome b reduction was triphasic, consisting of a rapid partial reduction phase, an apparent partial reoxidation phase, and a slow rereduction phase. Absorbance spectra taken by rapid scanning spectroscopy at 1-ms intervals before, during, and after the apparent reoxidation phase showed that this was caused by a bona fide reoxidation of cytochrome b and not by any negative spectral contribution from cytochrome c(1). With cytochrome bc(1) complex from a yeast mutant that cannot synthesize ubiquinone, cytochrome b reduction by either menaquinol or ubiquinol was rapid and monophasic. Addition of ubiquinone restored triphasic cytochrome b reduction, and the duration of the reoxidation phase increased as the ubiquinone concentration increased. When reduction of the cytochrome bc(1) complex through center P was blocked, cytochrome b reduction through center N was biphasic and was slowed by the addition of exogenous ubiquinone. These results show that ubiquinone residing at center N in the oxidized cytochrome bc(1) complex is responsible for the triphasic reduction of cytochrome b. PMID- 10531316 TI - Hypoxic enhancement of quantal catecholamine secretion. Evidence for the involvement of amyloid beta-peptides. AB - Prolonged exposure to hypoxia (10% O(2)) enhanced quantal catecholamine release evoked from O(2)-sensing pheochromocytoma (PC12) cells, as monitored using single cell amperometric recordings. The enhancement of exocytosis was apparent after 12 h of hypoxia and was maximal at 24 h. Elevated levels of secretion were due to the emergence of a Ca(2+) influx pathway that persisted during complete blockade of known voltage-gated Ca(2+) channels. Secretion triggered by this Ca(2+) influx was severely reduced by known inhibitors of Alzheimer's amyloid beta-peptides (AbetaPs), including an N terminus-directed monoclonal antibody. The enhancing effect on secretion of chronic hypoxia was mimicked closely by direct application of AbetaP to cells under normoxic conditions, although the effects of AbetaP were more rapid at onset, being maximal after only 6 h. The present results suggest that prolonged hypoxia can induce formation of Ca(2+)-permeable AbetaP channels and that such induction can lead directly to excessive neurosecretion. This is a potential contributory factor to AbetaP pathophysiology following cerebral ischemia. PMID- 10531317 TI - Anchorage-dependent regulation of the mitogen-activated protein kinase cascade by growth factors is supported by a variety of integrin alpha chains. AB - Integrin cooperation with growth factor receptors to enable permissive signaling to the mitogen-activated protein (MAP) kinase pathway has important implications for cell proliferation, differentiation, and survival. Here we have sought to determine whether anchorage regulation of the MAP kinase pathway is specific to the alpha chain subunit of the integrins employed during adhesion. Human umbilical vein endothelial cells (HUVECs) anchored via endogenous alpha(2), alpha(3), or alpha(5) integrin subunits or NIH3T3 fibroblast cells lines anchored via ectopically expressed human integrin alpha(2) or alpha(5) subunits displayed comparable MAP kinase activation upon growth factor stimulation, regardless of the integrin alpha chain employed. In contrast, when either cell type was maintained in suspension, growth factor treatment inefficiently activated the MAP kinase pathway. The integrin-mediated enhancement of MAP kinase activation by growth factor correlated with the tyrosine phosphorylation of focal adhesion kinase but was independent of Shc. These data indicate that integrin modulation of the MAP kinase pathway is supported by a variety of integrin complexes and imply that other pathways may be required for the previously reported alpha chain specific effects on cell cycle regulation and cell differentiation. PMID- 10531318 TI - Smad3 inhibits transforming growth factor-beta and activin signaling by competing with Smad4 for FAST-2 binding. AB - Transcriptional regulation by transforming growth factor-beta and activin is mediated by interaction of Smad2 and Smad3 with specific transcription factors and/or DNA elements. However, Smad3 behaves differently from Smad2 in regulating transcription by a winged-helix transcription factor, FAST-2, on an activin responsive element (ARE) in the Xenopus Mix.2 promoter. Smad3 alone was able to stimulate the ARE through FAST-2, but inhibited the ARE transactivation mediated by Smad2/Smad4 following receptor activation. We characterized the functional domains that are involved in these two activities of Smad3. Deletion of the MH1 domain as well as mutations of four lysine residues in the MH1 domain abrogated the inhibitory activity of Smad3, but did not compromise the self-stimulatory function. In contrast, deletion of the MH2 domain or a point mutation of glycine 379 within this domain obliterated the self-stimulatory activity of Smad3, but not the inhibitory activity. In an electrophoretic mobility shift assay, we found that Smad3 was able to associate with the FAST-2.ARE complex and that this association was dependent on FAST-2. In addition, Smad3 was not able to directly bind the ARE in a DNase I protection assay, in which FAST-2 binds the ARE around a motif (TGTGTATT) previously characterized to associate with the human FAST-1 protein. Interestingly, Smad4 was also able to directly associate with the FAST 2.ARE complex through binding with FAST-2. In a gel shift assay, the association of FAST-2 with Smad4 was mutually exclusive from the association with Smad3. Taken together, these data indicate that Smad3 exerts the inhibitory activity by competitive association with FAST-2. PMID- 10531319 TI - Highly specific recognition of primer RNA structures for 2'-OH priming reaction by bacterial reverse transcriptases. AB - A minor population of Escherichia coli contains retro-elements called retrons, which encode reverse transcriptases (RT) to synthesize peculiar satellite DNAs called multicopy single-stranded DNA (msDNA). These RTs recognize specific RNA structures in their individual primer-template RNAs to initiate cDNA synthesis from the 2'-OH group of a specific internal G residue (branching G residue). The resulting products (msDNA) consist of RNA and single-stranded DNA, sharing hardly any sequence homology. Here, we investigated how RT-Ec86 recognizes the specific RNA structure in its primer-template RNA. On the basis of structural comparison with HIV-1 RT, domain exchanges were carried out between two E. coli RTs, RT-Ec86 and RT-Ec73. RT-Ec86 (320 residues) and RT-Ec73 (316 residues) share only 71 identical residues (22%). From the analysis of 10 such constructs, the C-terminal 91-residue sequence of RT-Ec86 was found to be essential for the recognition of the unique stem-loop structure and the branching G residue in the primer-template RNA for retron-Ec86. Using the SELEX (systematic evolution of ligands by exponential enrichment) method with RT-Ec86 and primer RNAs containing random sequences, the identical stem-loop structure (including the 3-U loop) to that found in the retron-Ec86 primer-template RNA was enriched. In addition, the highly conserved 4-base sequence (UAGC), including the branching G residue, was also enriched. These results indicate that the highly diverse C-terminal region recognizes specific stem-loop structures and the branching G residue located upstream of the stem-loop structure. The present results with seemingly primitive RNA-dependent DNA polymerases provide insight into the mechanisms for specific protein RNA recognition. PMID- 10531320 TI - Prostaglandin E(2) mediates inhibition of insulin secretion by interleukin-1beta. AB - Interleukin-1beta (IL-1beta) and prostaglandin E(2) (PGE(2)), frequently co participants in inflammatory states, are two well recognized inhibitors of glucose-induced insulin secretion. Previous reports have concluded that the inhibitory effects of these two autacoids on pancreatic beta cell function are not related because indomethacin, a potent prostaglandin synthesis inhibitor, does not prevent IL-1beta effects. However, indomethacin is not a specific cyclooxygenase inhibitor, and its other pharmacologic effects are likely to inhibit insulin secretion independently. Since we recently observed that IL-1beta induces cyclooxygenase-2 (COX-2) gene expression and PGE(2) synthesis in islet beta cells, we have reassessed the possibility that PGE(2) mediates IL-1beta effects on beta function. By using two cell lines (HIT-T15 and betaHC13) as well as Wistar rat isolated pancreatic islets, we examined the ability of two COX-2 specific antagonists, NS-398 and SC-236, to prevent IL-1beta inhibition of insulin secretion. Both drugs prevented IL-1beta from inducing PGE(2) synthesis and inhibiting insulin secretion; adding back exogenous PGE(2) re-established inhibition of insulin secretion in the presence of IL-1beta. We also found that EP3, the PGE(2) receptor subtype whose post-receptor effect is to decrease adenylyl cyclase activity and, thereby, insulin secretion, is the dominant mRNA subtype expressed. We conclude that endogenous PGE(2) mediates the inhibitory effects of exogenous IL-1beta on beta cell function. PMID- 10531321 TI - Stepwise transfer of tRNA through the double membrane of Leishmania mitochondria. AB - Import of tRNA into Leishmania mitochondria involves transfer through a double membrane barrier. To examine whether specific sorting mechanisms for individual tRNAs direct them to different mitochondrial compartments, the distribution of tRNA transcripts, internalized in vitro, was examined by suborganellar fractionation. Significant amounts of tRNA(Tyr) were localized in the matrix and on the outer face of the inner mitochondrial membrane. With time, the matrix:membrane ratio increased. Translocation through the inner membrane apparently required the presence of a specific signal in the D arm of tRNA(Tyr), and tRNA(Gln)(CUG), lacking this sequence, was excluded. Hydrolysis of ATP was necessary at both the outer and inner membranes. However, the protonophores carbonylcyanide m-chlorophenylhydrazone and nigericin, the K(+) ionophore valinomycin, and the F(1)F(0) ATPase inhibitor oligomycin had only marginal effects on uptake through the outer membrane but severely inhibited inner membrane translocation, indicating the unusual requirement of both the electrical and chemical components of the electromotive force generated across the inner membrane. The results are consistent with a mechanism involving stepwise transfer of tRNA through distinct outer and inner membrane channels. PMID- 10531322 TI - Deletion of the regulatory domain in the pyridoxal phosphate-dependent heme protein cystathionine beta-synthase alleviates the defect observed in a catalytic site mutant. AB - The most common cause of severely elevated homocysteine or homocystinuria is inherited disorders in cystathionine beta-synthase. The latter enzyme is a unique hemeprotein that catalyzes pyridoxal phosphate (PLP)-dependent condensation of serine and homocysteine to give cystathionine, thus committing homocysteine to catabolism. A point mutation, V168M, has been described in a homocystinuric cell line and is associated with a B(6)-responsive phenotype. In this study, we have examined the kinetic properties of this mutant and demonstrate that the mutation affects the PLP but not the heme content. The approximately 13-fold diminution in activity because of the mutation corresponds to an approximately 7-fold decrease in the level of bound PLP. This may be explained by half of the sites activity associated with cystathionine beta-synthase. The addition of PLP results in partial but not full restoration of activity to wild type levels. Elimination of the C-terminal quarter of the mutant protein results in alleviation of the catalytic penalty imposed by the V168M mutation. The resulting truncated protein is very similar to the corresponding truncated enzyme with wild type sequence and is now able to bind the full complement of both heme and PLP cofactors. These results indicate that the V168M mutation per se does not affect binding of PLP directly and that interactions between the regulatory C terminus and the catalytic N terminus are important in modulating the cofactor content and therefore the activity of the full-length enzyme. These studies provide the first biochemical explanation for the B(6)-responsive phenotype associated with a cystathionine beta-synthase-impaired homocystinuric genotype. PMID- 10531324 TI - Thermodynamic analyses reveal role of water release in epitope recognition by a monoclonal antibody against the human guanylyl cyclase C receptor. AB - The thermodynamics of a monoclonal antibody (mAb)-peptide interaction have been characterized by isothermal titration microcalorimetry. GCC:B10 mAb, generated against human guanylyl cyclase C, a membrane-associated receptor and a potential marker for metastatic colon cancer, recognizes the cognate peptide epitope HIPPENIFPLE and its two contiguous mimotopes, HIPPEN and ENIFPLE, specifically and reversibly. The exothermic binding reactions between 6.4 and 42 degrees C are driven by dominant favorable enthalpic contributions between 20 and 42 degrees C, with a large negative heat capacity (DeltaC(p)) of -421 +/- 27 cal mol(-1) K(-1). The unfavorable negative value of entropy (DeltaS(b)(0)) at 25 degrees C, an unusual feature among protein-protein interactions, becomes a positive one below an inversion temperature of 20.5 degrees C. Enthalpy-entropy compensation due to solvent reorganization accounts for an essentially unchanged free energy of interaction (DeltaDeltaG(b)(0) congruent with 0). The role of water molecules in the recognition process was tested by coupling an osmotic stress technique with isothermal titration microcalorimetry. The results provide direct and compelling evidence that GCC:B10 mAb recognizes the peptides HIPPENIFPLE, HIPPEN, and ENIFPLE differentially, with a concomitant release of variable and nonadditive numbers of water molecules (15, 7, and 3, respectively) from the vicinity of the binding site. PMID- 10531323 TI - Cholecystokinin activates PYK2/CAKbeta by a phospholipase C-dependent mechanism and its association with the mitogen-activated protein kinase signaling pathway in pancreatic acinar cells. AB - PYK2/CAKbeta is a recently described cytoplasmic tyrosine kinase related to p125 focal adhesion kinase (p125(FAK)) that can be activated by a number of stimuli including growth factors, lipids, and some G protein-coupled receptors. Studies suggest PYK2/CAKbeta may be important for coupling various G protein-coupled receptors to the mitogen-activated protein kinase (MAPK) cascade. The hormone neurotransmitter cholecystokinin (CCK) is known to activate both phospholipase C dependent cascades and MAPK signaling pathways; however, the relationship between these remain unclear. In rat pancreatic acini, CCK-8 (10 nM) rapidly stimulated tyrosine phosphorylation and activation of PYK2/CAKbeta by both activation of high affinity and low affinity CCK(A) receptor states. Blockage of CCK-stimulated increases in protein kinase C activity or CCK-stimulated increases in [Ca(2+)](i), inhibited by 40-50% PYK2/CAKbeta but not p125(FAK) tyrosine phosphorylation. Simultaneous blockage of both phospholipase C cascades inhibited PYK2/CAKbeta tyrosine phosphorylation completely and p125(FAK) tyrosine phosphorylation by 50%. CCK-8 stimulated a rapid increase in PYK2/CAKbeta kinase activity assessed by both an in vitro kinase assay and autophosphorylation. Total PYK2/CAKbeta under basal conditions was largely localized (77 +/- 7%) in the membrane fraction, whereas total p125(FAK) was largely localized (86 +/- 3%) in the cytosolic fraction. With CCK stimulation, both p125(FAK) and PYK2/CAKbeta translocated to the plasma membrane. Moreover CCK stimulation causes a rapid formation of both PYK2/CAKbeta-Grb2 and PYK2/CAKbeta-Crk complexes. These results demonstrate that PYK2/CAKbeta and p125(FAK) are regulated differently by CCK(A) receptor stimulation and that PYK2/CAKbeta is probably an important mediator of downstream signals by CCK-8, especially in its ability to activate the MAPK signaling pathway, which possibly mediates CCK growth effects in normal and neoplastic tissues. PMID- 10531325 TI - Effects of tropomyosin internal deletions on thin filament function. AB - Striated muscle tropomyosin spans seven actin monomers and contains seven quasi repeating regions with loose sequence similarity. Each region contains a hypothesized actin binding motif. To examine the functions of these regions, full length tropomyosin was compared with tropomyosin internal deletion mutants spanning either five or four actins. Actin-troponin-tropomyosin filaments lacking tropomyosin regions 2-3 exhibited calcium-sensitive regulation in in vitro motility and myosin S1 ATP hydrolysis experiments, similar to filaments with full length tropomyosin. In contrast, filaments lacking tropomyosin regions 3-4 were inhibitory to these myosin functions. Deletion of regions 2-4, 3-5, or 4-6 had little effect on tropomyosin binding to actin in the presence of troponin or troponin-Ca(2+), or in the absence of troponin. However, all of these mutants inhibited myosin cycling. Deletion of the quasi-repeating regions diminished the prominent effect of myosin S1 on tropomyosin-actin binding. Interruption of this cooperative, myosin-tropomyosin interaction was least severe for the mutant lacking regions 2-3 and therefore correlated with inhibition of myosin cycling. Regions 3, 4, and 5 each contributed about 1.5 kcal/mol to this process, whereas regions 2 and 6 contributed much less. We suggest that a myosin-induced conformational change in actin facilitates the azimuthal repositioning of tropomyosin which is an essential part of regulation. PMID- 10531327 TI - The anaerobic ribonucleotide reductase from Escherichia coli. The small protein is an activating enzyme containing a [4fe-4s](2+) center. AB - For deoxyribonucleotide synthesis during anaerobic growth, Escherichia coli cells depend on an oxygen-sensitive class III ribonucleotide reductase. The enzyme system consists of two proteins: protein alpha, on which ribonucleotides bind and are reduced, and protein beta, of which the function is to introduce a catalytically essential glycyl radical on protein alpha. Protein beta can assemble one [4Fe-4S] center per polypeptide enjoying both the [4Fe-4S](2+) and [4Fe-4S](1+) redox state, as shown by iron and sulfide analysis, Mossbauer spectroscopy (delta = 0.43 mm.s(-1), DeltaE(Q) = 1.0 mm.s(-1), [4Fe-4S](2+)), and EPR spectroscopy (g = 2. 03 and 1.93, [4Fe-4S](1+)). This iron center is sensitive to oxygen and can decompose into stable [2Fe-2S](2+) centers during exposure to air. This degraded form is nevertheless active, albeit to a lesser extent because of the conversion of the cluster into [4Fe-4S] forms during the strongly reductive conditions of the assay. Furthermore, protein beta has the potential to activate several molecules of protein alpha, suggesting that protein beta is an activating enzyme rather than a component of an alpha(2)beta(2) complex as previously claimed. PMID- 10531326 TI - A novel human apolipoprotein (apoM). AB - A novel human apolipoprotein designated apolipoprotein M (apoM) is described. The unique N-terminal amino acid sequence of apoM was found in an approximately 26 kDa protein present in a protein extract of triglyceride-rich lipoproteins (TGRLP). The isolated apoM cDNA (734 base pairs) encoded a 188-amino acid residue long protein, distantly related to the lipocalin family. The mRNA of apoM was detected in the liver and kidney. Western blotting demonstrated apoM to be present in high density lipoprotein (HDL) and to a lesser extent in TGRLP and low density lipoproteins (LDL). The first 20 amino acid residues of apoM constituted a hydrophobic segment with characteristic features of a signal peptide. This was retained in the mature protein because of the lack of a signal peptidase cleavage site. In vitro translation in the presence of microsomes demonstrated translocation of apoM over the membrane and glycosylation but no signal peptide cleavage. The in vitro translated product remained associated with the microsomes after treatment with carbonate at pH 11, demonstrating that apoM is an integral protein. This finding suggests that apoM is linked to the single phospholipid layer of lipoproteins with a hydrophobic signal anchor. In conclusion, a novel human apolipoprotein, the function of which remains to be determined, is described. PMID- 10531328 TI - Transcription of BRCA1 is dependent on the formation of a specific protein-DNA complex on the minimal BRCA1 Bi-directional promoter. AB - BRCA1 is the first tumor suppressor gene linked to hereditary breast and ovarian cancers. Its involvement in sporadic breast cancer, however, remains unclear. Recent studies showed that a loss or lowered expression of BRCA1 is not uncommon in nonfamilial breast cancers. In addition, there have been cases of inherited BRCA1-linked breast cancer with as yet unidentified mutation. Misregulation of BRCA1 at the transcription level is a possible mechanism for loss of BRCA1 expression. To understand transcriptional regulation of the BRCA1 gene, we cloned and examined the BRCA1 promoter, by both functional reporter gene analyses and protein-DNA complex formation electrophorectic mobility shift assays. A bi directional promoter could be located within a 229-base pair (bp) intergenic region between BRCA1 and its neighboring gene, NBR2. Deletion analyses further delineated a minimal 56-bp EcoRI-HaeIII fragment, which could drive transcription in the NBR2 gene direction 2-4-fold higher than in the BRCA1 direction in all cell lines tested. Furthermore, transcriptional activity in the BRCA1 direction was undetectable in the muscle cell line C2C12, whereas activity in the NBR2 direction was maintained. These results were consistent with the expression pattern of the respective genes. A specific protein-DNA complex was detected when nuclear extracts from HeLa cells and Caco2, a colon cell line, were incubated with the 56-bp minimal promoter. This protein binding activity was further localized to an 18-bp fragment and might involve a tissue-specific factor, because binding was not detected in the C2C12 cell line. The correlation of the detection of this protein-DNA complex only in those cell lines that expressed the chloramphenicol acetyltransferase reporter gene in the BRCA1 direction suggests a significant positive role of this complex in the transcription of the BRCA1 gene. PMID- 10531329 TI - The second and fourth cluster of class A cysteine-rich repeats of the low density lipoprotein receptor-related protein share ligand-binding properties. AB - The low density lipoprotein receptor-related protein (LRP) is a multifunctional endocytic cell-surface receptor that binds and internalizes a diverse array of ligands. The receptor contains four putative ligand-binding domains, generally referred to as clusters I, II, III, and IV. In this study, soluble recombinant receptor fragments, representing each of the four individual clusters, were used to map the binding sites of a set of structurally and functionally distinct ligands. Using surface plasmon resonance, we studied the binding of these fragments to methylamine-activated alpha(2)-macroglobulin, pro-urokinase-type plasminogen activator, tissue-type plasminogen activator (t-PA), plasminogen activator inhibitor-1, t-PA.plasminogen activator inhibitor-1 complexes, lipoprotein lipase, apolipoprotein E, tissue factor pathway inhibitor, lactoferrin, the light chain of blood coagulation factor VIII, and the intracellular chaperone receptor-associated protein (RAP). No binding of the cluster I fragment to any of the tested ligands was observed. The cluster III fragment only bound to the anti-LRP monoclonal antibody alpha(2)MRalpha3 and weakly to RAP. Except for t-PA, we found that each of the ligands tested binds both to cluster II and to cluster IV. The affinity rate constants of ligand binding to clusters II and IV and to LRP were measured, showing that clusters II and IV display only minor differences in ligand-binding kinetics. Furthermore, we demonstrate that the subdomains C3-C7 of cluster II are essential for binding of ligands and that this segment partially overlaps with a RAP-binding site on cluster II. Finally, we show that one RAP molecule can bind to different clusters simultaneously, supporting a model in which RAP binding to LRP induces a conformational change in the receptor that is incompatible with ligand binding. PMID- 10531330 TI - Discordant effects of glucosamine on insulin-stimulated glucose metabolism and phosphatidylinositol 3-kinase activity. AB - The impact of increased GlcN availability on insulin-stimulated p85/p110 phosphatidylinositol 3-kinase (PI3K) activity in skeletal muscle was examined in relation to GlcN-induced defects in peripheral insulin action. Primed continuous GlcN infusion (750 micromol/kg bolus; 30 micromol/kg.min) in conscious rats limited both maximal stimulation of muscle PI3K by acute insulin (I) (1 unit/kg) bolus (I + GlcN = 1.9-fold versus saline = 3.3-fold above fasting levels; p < 0.01) and chronic activation of PI3K following 3-h euglycemic, hyperinsulinemic (18 milliunits/kg.min) clamp studies (I + GlcN = 1.2-fold versus saline = 2.6 fold stimulation; p < 0.01). To determine the time course of GlcN-induced defects in insulin-stimulated PI3K activity and peripheral insulin action, GlcN was administered for 30, 60, 90, or 120 min during 2-h euglycemic, hyperinsulinemic clamp studies. Activation of muscle PI3K by insulin was attenuated following only 30 min of GlcN infusion (GlcN 30 min = 1.5-fold versus saline = 2.5-fold stimulation; p < 0.05). In contrast, the first impairment in insulin-mediated glucose uptake (Rd) developed following 110 min of GlcN infusion (110 min = 39.9 +/- 1.8 versus 30 min = 42.8 +/- 1.4 mg/kg.min, p < 0.05). However, the ability of insulin to stimulate phosphatidylinositol 3,4, 5-trisphosphate production and to activate glycogen synthase in skeletal muscle was preserved following up to 180 min of GlcN infusion. Thus, increased GlcN availability induced (a) profound and early inhibition of proximal insulin signaling at the level of PI3K and (b) delayed effects on insulin-mediated glucose uptake, yet (c) complete sparing of insulin-mediated glycogen synthase activation. The pattern and time sequence of GlcN-induced defects suggest that the etiology of peripheral insulin resistance may be distinct from the rapid and marked impairment in insulin signaling. PMID- 10531331 TI - Characterization of receptor-interacting protein 140 in retinoid receptor activities. AB - Receptor-interacting protein 140 (RIP140) contains multiple receptor interaction domains and interacts with retinoic acid receptors in a ligand-dependent manner. Nine LXXLL receptor-interacting motifs are organized into two clusters within this molecule, each differentially interacting with retinoic acid receptor (RAR) and retinoid X receptor (RXR). RAR interacts with the 5' cluster, whereas RXR interacts with both clusters. Additionally, a third ligand-dependent receptor interacting domain is assigned to the very C terminus of this molecule, which contains no LXXLL motif. In mammalian cells, receptor heterodimerization is required for efficient interaction of RAR/RXR with RIP140. Furthermore, the heterodimeric, holoreceptors cooperatively interact with RIP140, which requires the activation function 2 domains of both receptors. By using different retinoic acid reporter systems, it is demonstrated that RIP140 strongly suppresses retinoic acid induction of reporter activities, but coactivator SRC-1 enhances it. Furthermore, an intrinsic repressive activity of RIP140 is demonstrated in a GAL4 fusion system. Unlike receptor corepressor, which interacts with antagonist bound RAR/RXRs, RIP140 does not interact with antagonist-occupied RAR/RXR dimers. These data suggest that RIP140 represents a third coregulator category that is able to suppress the activation of certain agonist-bound hormone receptors. PMID- 10531332 TI - The N terminus of Kaposi's sarcoma-associated herpesvirus G protein-coupled receptor is necessary for high affinity chemokine binding but not for constitutive activity. AB - Kaposi's sarcoma-associated herpesvirus (KSHV) contains a gene encoding a G protein-coupled receptor (KSHV-GPCR) that is homologous to mammalian chemokine receptors. KSHV-GPCR signals constitutively (in an agonist-independent manner) via the phosphoinositide-inositol 1,4,5-trisphosphate pathway. Because it has been proposed that the N terminus (N-TERM) of other GPCRs may act as tethered agonists, we determined whether the N-TERM of KSHV-GPCR is necessary for constitutive signaling activity or ligand binding, or both. We show that replacement of the entire N-TERM of KSHV-GPCR with those of two other GPCRs, deletion of residues within the N-TERM, and disruption of a putative disulfide bond that may hold the N-TERM in close proximity to extracellular loop 3 do not affect constitutive signaling activity but decrease chemokine binding. There were differences in the effects of mutation of the N-TERM on binding of the chemokines growth-related oncogene alpha, which is an agonist, and interferon-gamma inducible protein-10, which is an inverse agonist. The effects on chemokine binding were accompanied by changes in chemokine regulation of KSHV-GPCR signaling. We conclude that the N-TERM is not necessary for constitutive KSHV GPCR signaling, i.e. the N-TERM is not a tethered agonist, but plays a crucial role in binding of chemokine ligands and of chemokine regulation of KSHV-GPCR signaling. PMID- 10531333 TI - Two structural states of complexes of peptide and class II major histocompatibility complex revealed by photoaffinity-labeled peptides. AB - The complex of the murine class II histocompatibility molecules I-A(k) with high affinity binding peptides were resistant to denaturation when examined by SDS polyacrylamide gel electrophoresis at various pH levels. In contrast, complexes made with low affinity binding peptides were highly sensitive to denaturation by SDS. This effect was more pronounced at low pH. Placing a photoactivatable probe at the amino terminus of the peptides resulted in their covalent linkage to soluble I-A(k) molecules. We found an inverse relationship between the capacity of peptides to form SDS-stable complexes with I-A(k) and their extent of covalent association with either the alpha or beta chain. The relationship held true for three different peptides in which the main anchor residues were changed so as to affect their binding affinity for I-A(k) molecules. Thus, high affinity peptides generate a complex in which the motion of their amino termini was restricted, whereas complexes of low affinity peptides are more flexible. In agreement with this observation, complexes of I-A(k) with high affinity peptides were highly resistant to proteolysis, in contrast to those formed with weakly binding peptides, which were more likely to be cleaved. Complexes with low affinity peptides generate a structure with enhanced flexibility as compared with complexes with high affinity peptides. PMID- 10531335 TI - Tapasin enhances assembly of transporters associated with antigen processing dependent and -independent peptides with HLA-A2 and HLA-B27 expressed in insect cells. AB - Assembly of HLA class I-peptide complexes is assisted by multiple proteins that associate with HLA molecules in loading complexes. These include the housekeeping chaperones calnexin and calreticulin and two essential proteins, the transporters associated with antigen processing (TAP) for peptide supply, and the protein tapasin which is thought to act as a specialized chaperone. We dissected functional effects of processing cofactors by co-expressing in insect cells various combinations of the human proteins HLA-A2, HLA-B27, beta(2) microglobulin, TAP, calnexin, calreticulin, and tapasin. Stability at 37 degrees C and surface expression of class I dimers correlated closely in baculovirus infected Sf9 cells, suggesting that these cells retain empty dimers in the endoplasmic reticulum. Both HLA molecules form substantial quantities of stable complexes with insect cell-produced peptide pools. These pools are TAP-selected cytosolic peptides for HLA-B27 but endoplasmic reticulum-derived, i.e. TAP independent peptides for HLA-A2. This discrepancy may be due to peptide selection by human TAP which is much better adapted to the HLA-B27 than to the HLA-A2 ligand preferences. HLA class I assembly with peptides from TAP-dependent and independent pools was enhanced strongly by tapasin. Thus, tapasin acts as a chaperone and/or peptide editor that facilitates assembly of peptides with HLA class I molecules independently of mediating their interaction with TAP and/or retention in the endoplasmic reticulum. PMID- 10531334 TI - Serine 19 of human 6-pyruvoyltetrahydropterin synthase is phosphorylated by cGMP protein kinase II. AB - 6-Pyruvoyltetrahydropterin synthase (PTPS) participates in tetrahydrobiopterin cofactor biosynthesis. We previously identified in a PTPS-deficient patient an inactive PTPS allele with an Arg(16) to Cys codon mutation. Arg(16) is located in the protein surface exposed phosphorylation motif Arg(16)-Arg-Ile-Ser, with Ser(19) as the putative phosphorylation site for serine-threonine protein kinases. Purification of recombinant PTPS-S19A from bacterial cells resulted in an active enzyme (k(cat)/K(m) = 6.4 x 10(3) M(-1) s(-1)), which was similar to wild-type PTPS (k(cat)/K(m) = 4.1 x 10(3) M(-1) s(-1)). In assays with purified enzymes, wild-type but not PTPS-S19A was a specific substrate for the cGMP dependent protein kinase (cGK) type I and II. Upon expression in COS-1 cells, PTPS-S19A was stable but not phosphorylated and had a reduced activity of approximately 33% in comparison to wild-type PTPS. Extracts from several human cell lines, including brain, contained a kinase that bound to and phosphorylated immobilized wild-type, but not mutant PTPS. Addition of cGMP stimulated phosphotransferase activity 2-fold. Extracts from transfected COS-1 cells overexpressing cGKII stimulated Ser(19) phosphorylation more than 100-fold, but only 4-fold from cGKI overexpressing cells. Moreover, fibroblast extracts from mice lacking cGKII exhibited significantly reduced phosphorylation of PTPS. These results suggest that Ser(19) of human PTPS may be a substrate for cGKII phosphorylation also in vivo, a modification that is essential for normal activity. PMID- 10531336 TI - Expression of hepatitis B virus polymerase in Ty1-his3AI retroelement of Saccharomyces cerevisiae. AB - Hepatitis B virus (HBV), although a DNA virus, replicates using reverse transcriptase encoded by the HBV polymerase (pol) gene. The biochemical dissection of HBV pol has been hampered by failure to liberate enzymatically active protein from nucleocapsids. Here, we have employed a yeast-based genetic approach to express the HBV reverse transcriptase. In this strategy, the reverse transcriptase of yeast retrotransposon Ty1 element is replaced with the HBV pol gene to produce the hybrid Ty1/HBV element. Additionally, the indicator gene his3AI is combined in an antisense orientation to the transcripts of the hybrid Ty1/HBVRT element. The splicing of his3AI, cDNA synthesis of the Ty1/HBVRT RNA and subsequent integration relies on the reverse transcriptase activity. The production of histidine prototrophs results from the successful reverse transcription of Ty1/HBVRThis3AI transcripts followed by either homologous recombination or integrase-mediated insertion and subsequent expression of HIS3 gene. Using this approach we successfully detected the reverse transcriptase activity of HBV in yeast strains defective in endogenous Ty1 expression. Consistent with the unique priming activity associated with HBV pol, the minus strand DNA synthesis was protein-primed. Deletion of HBV reverse transcriptase (RT) or RNase H domains resulted in a dramatic drop in histidine prototrophs. The addition of HBV encoded HBx protein in virus-like particles during in vitro RT reaction stimulated the RT reaction by severalfold. Furthermore, in the presence of 3TC, a known inhibitor of HBV reverse transcriptase, yeast His(+) growth of His protrophs was not observed. Thus, this approach, which is based on genetic selection in yeast, is safe, economic, and a reliable strategy with a potential for large scale screening of cofactors and inhibitors of HBV polymerase functions. PMID- 10531337 TI - Oxidation of the alpha(3)(betaD311C/R333C)(3)gamma subcomplex of the thermophilic Bacillus PS3 F(1)-ATPase indicates that only two beta subunits can exist in the closed conformation simultaneously. AB - In the crystal structure of the bovine heart mitochondrial F(1)-ATPase (Abrahams, J. P., Leslie, A. G. W., Lutter, R., and Walker, J. E. (1994) Nature 370, 621 628), the two liganded beta subunits, one with MgAMP-PNP bound to the catalytic site (beta(T)) and the other with MgADP bound (beta(D)) have closed conformations. The empty beta subunit (beta(E)) has an open conformation. In beta(T) and beta(D), the distance between the carboxylate of beta-Asp(315) and the guanidinium of beta-Arg(337) is 3.0-4.0 A. These side chains are at least 10 A apart in beta(E). The alpha(3)(betaD311C/R333C)(3)gamma subcomplex of TF(1) with the corresponding residues substituted with cysteine has very low ATPase activity unless it is reduced prior to assay or assayed in the presence of dithiothreitol. The reduced subcomplex hydrolyzes ATP at 50% the rate of wild type and is rapidly inactivated by oxidation by CuCl(2) with or without magnesium nucleotides bound to catalytic sites. Titration of the subcomplex with iodo[(14)C]acetamide after prolonged treatment with CuCl(2) in the presence or absence of 1 mM MgADP revealed nearly two free sulfhydryl groups/mol of enzyme. Therefore, one pair of introduced cysteines is located on a beta subunit that exists in the open or partially open conformation even when catalytic sites are saturated with MgADP. Since V(max) of ATP hydrolysis is attained when three catalytic sites of F(1) are saturated, the catalytic site that binds ATP must be closing as the catalytic site that releases products is opening. PMID- 10531338 TI - Engineering of the myosin-ibeta nucleotide-binding pocket to create selective sensitivity to N(6)-modified ADP analogs. AB - Distinguishing the cellular functions carried out by enzymes of highly similar structure would be simplified by the availability of isozyme-selective inhibitors. To determine roles played by individual members of the large myosin superfamily, we designed a mutation in myosin's nucleotide-binding pocket that permits binding of adenine nucleotides modified with bulky N(6) substituents. Introduction of this mutation, Y61G in rat myosin-Ibeta, did not alter the enzyme's affinity for ATP or actin and actually increased its ATPase activity and actin-translocation rate. We also synthesized several N(6)-modified ADP analogs that should bind to and inhibit mutant, but not wild-type, myosin molecules. Several of these N(6)-modified ADP analogs were more than 40-fold more potent at inhibiting ATP hydrolysis by Y61G than wild-type myosin-Ibeta; in doing so, these analogs locked Y61G myosin-Ibeta tightly to actin. N(6)-(2-methylbutyl) ADP abolished actin filament motility mediated by Y61G, but not wild-type, myosin Ibeta. Furthermore, a small fraction of inhibited Y61G molecules was sufficient to block filament motility mediated by mixtures of wild-type and Y61G myosin Ibeta. Introduction of Y61G myosin-Ibeta molecules into a cell should permit selective inhibition by N(6)-modified ADP analogs of cellular processes dependent on myosin-Ibeta. PMID- 10531339 TI - Conformation of the regulatory domain of cardiac muscle troponin C in its complex with cardiac troponin I. AB - Calcium activation of fast striated muscle results from an opening of the regulatory N-terminal domain of fast skeletal troponin C (fsTnC), and a substantial exposure of a hydrophobic patch, essential for Ca(2+)-dependent interaction with fast skeletal troponin I (fsTnI). This interaction is obligatory to relieve the inhibition of strong, force-generating actin-myosin interactions. We have determined intersite distances in the N-terminal domain of cardiac TnC (cTnC) by fluorescence resonance energy transfer measurements and found negligible increases in these distances when the single regulatory site is saturated with Ca(2+). However, in the presence of bound cardiac TnI (cTnI), activator Ca(2+) induces significant increases in the distances and a substantial opening of the N-domain. This open conformation within the cTnC.cTnI complex has properties favorable for the Ca(2+)-induced interaction with an additional segment of cTnI. Thus, the binding of cTnI to cTnC is a prerequisite to achieve a Ca(2+)-induced open N-domain similar to that previously observed in fsTnC with no bound fsTnI. This role of cardiac TnI has not been previously recognized. Our results also indicate that structural information derived from a single protein may not be sufficient for inference of a structure/function relationship. PMID- 10531340 TI - A Haemophilus influenzae gene that encodes a membrane bound 3-deoxy-D-manno octulosonic acid (Kdo) kinase. Possible involvement of kdo phosphorylation in bacterial virulence. AB - The lipopolysaccharide of Haemophilus influenzae contains a single 3-deoxy-D manno-octulosonic acid (Kdo) residue derivatized with either a phosphate or an ethanolamine pyrophosphate moiety at the 4-OH position. In previous studies, we identified a kinase unique to H. influenzae extracts that phosphorylates Kdo lipid IV(A), a key precursor of lipopolysaccharide in this organism. We have now identified the gene encoding the Kdo kinase by using an expression cloning approach. A cosmid library containing random DNA fragments from H. influenzae strain Rd was constructed in Escherichia coli. Extracts of 472 colonies containing individual hybrid cosmids were assayed for Kdo kinase activity. A single hybrid cosmid directing expression of the kinase was found. The kinase gene was identified by activity assays, sub-cloning, and DNA sequencing. When the putative kinase gene was expressed in E. coli behind a T7 promoter, massive overproduction of kinase activity was achieved ( approximately 8000-fold higher than in H. influenzae membranes). The catalytic properties and the product generated by the overexpressed kinase, assayed with Kdo-lipid IV(A) as the substrate, were the same as observed with H. influenzae membranes. Unexpectedly, the kinase gene was identical to a previously characterized open reading frame (orfZ), which had been shown to be important for establishing bacteremia in an infant rat model (Hood, D. W., Deadman, M. E., Allen, T., Masoud, H., Martin, A., Brisson, J. R., Fleischmann, R., Venter, J. C., Richards, J. C., and Moxon, E. R. (1996) Mol. Microbiol. 22, 951-965). However, based solely on the genome sequence of H. influenzae Rd, no biochemical function had been assigned to the product of orfZ, which we now designate kdkA ("Kdo kinase A"). Although Kdo phosphorylation may be critical for bacterial virulence of H. influenzae, it does not appear to be required for growth. PMID- 10531341 TI - An internal signal sequence mediates the targeting and retention of the human UDP glucuronosyltransferase 1A6 to the endoplasmic reticulum. AB - The human UDP-glucuronosyltransferase isoform UGT1A6 is predicted to be a type I transmembrane protein anchored in the endoplasmic reticulum by a single C terminal transmembrane domain, followed by a short cytoplasmic tail. This topology is thought to be established through the sequential action of a cleavable N-terminal signal peptide and of a C-terminal stop transfer/anchor sequence. We found that the deletion of the signal peptide did not prevent membrane targeting and insertion of this protein expressed in an in vitro transcription/translation system or in yeast Pichia pastoris. Interestingly, the same results were obtained when the protein was depleted of both the signal peptide and the C-terminal transmembrane domain/cytoplasmic tail sequences, suggesting the presence of an internal topogenic element able to translocate and retain UGT1A6 in the endoplasmic reticulum membrane in vitro and in yeast cells. To identify such a sequence, the insertion of several N-terminal deletion mutants of UGT1A6 into microsomal membranes was investigated in vitro. The data clearly showed that the deletion of the N-terminal end did not affect endoplasmic reticulum targeting and retention until residues 140-240 were deleted. The signal like activity of the 140-240 region was demonstrated by the ability of this segment to confer endoplasmic reticulum residency to the cytosolic green fluorescent protein expressed in mammalian cells. Finally, we show that this novel topogenic sequence can posttranslationally mediate the translocation of UGT1A6. This study provides the first evidence that the membrane assembly of the human UGT1A6 involves an internal signal retention sequence. PMID- 10531342 TI - Molecular cloning and characterization of a novel repeat-containing Leishmania major gene, ppg1, that encodes a membrane-associated form of proteophosphoglycan with a putative glycosylphosphatidylinositol anchor. AB - Leishmania parasites secrete a variety of proteins that are modified by phosphoglycan chains structurally similar to those of the cell surface glycolipid lipophosphoglycan. These proteins are collectively called proteophosphoglycans. We report here the cloning and sequencing of a novel Leishmania major proteophosphoglycan gene, ppg1. It encodes a large polypeptide of approximately 2300 amino acids. The N-terminal domain of approximately 70 kDa exhibits 11 imperfect amino acid repeats that show some homology to promastigote surface glycoproteins of the psa2/gp46 complex. The large central domain apparently consists exclusively of approximately 100 repetitive peptides of the sequence APSASSSSA(P/S)SSSSS(+/-S). Gene fusion experiments demonstrate that these peptide repeats are the targets of phosphoglycosylation in Leishmania and that they form extended filamentous structures reminiscent of mammalian mucins. The C-terminal domain contains a functional glycosylphosphatidylinositol anchor addition signal sequence, which confers cell surface localization to a normally secreted Leishmania acid phosphatase, when fused to its C terminus. Antibody binding studies show that the ppg1 gene product is phosphoglycosylated by phosphoglycan repeats and cap oligosaccharides. In contrast to previously characterized proteophosphoglycans, the ppg1 gene product is predominantly membrane-associated and it is expressed on the promastigote cell surface. Therefore this membrane bound proteophosphoglycan may be important for direct host-parasite interactions. PMID- 10531343 TI - Conserved N-terminal cysteine motif is essential for homo- and heterodimer formation of synaptotagmins III, V, VI, and X. AB - The synaptotagmins now constitute a large family of membrane proteins characterized by one transmembrane region and two C2 domains. Dimerization of synaptotagmin (Syt) I, a putative low affinity Ca(2+) sensor for neurotransmitter release, is thought to be important for expression of function during exocytosis of synaptic vesicles. However, little is known about the self-dimerization properties of other isoforms. In this study, we demonstrate that a subclass of synaptotagmins (III, V, VI, and X) (Ibata, K., Fukuda, M., and Mikoshiba, K. (1998) J. Biol. Chem. 273, 12267-12273) forms beta-mercaptoethanol-sensitive homodimers and identify three evolutionarily conserved cysteine residues at the N terminus (N-terminal cysteine motif, at amino acids 10, 21, and 33 of mouse Syt III) that are not conserved in other isoforms. Site-directed mutagenesis of these cysteine residues and co-immunoprecipitation experiments clearly indicate that the first cysteine residue is essential for the stable homodimer formation of Syt III, V, or VI, and heterodimer formation between Syts III, V, VI, and X. We also show that native Syt III from mouse brain forms a beta-mercaptoethanol-sensitive homodimer. Our results suggest that the cysteine-based heterodimerization between Syt III and Syt V, VI, or X, which have different biochemical properties, may modulate the proposed function of Syt III as a putative high affinity Ca(2+) sensor for neurotransmitter release. PMID- 10531344 TI - A novel alternatively spliced variant of synaptotagmin VI lacking a transmembrane domain. Implications for distinct functions of the two isoforms. AB - Synaptotagmins are a family of membrane proteins that are characterized by a single transmembrane region and tandem C2 domains and that are likely to regulate constitutive and/or regulated vesicle traffic. We have shown that a subclass of synaptotagmins (III, V, VI, and X) forms homo- and heterodimers through an evolutionarily conserved cysteine motif at their N termini (Fukuda, M., Kanno, E., and Mikoshiba, K. (1999) J. Biol. Chem. 274, 31421-31427). In this study, we identified a novel alternatively spliced variant of synaptotagmin (Syt) VI that lacks the N-terminal 85 amino acids including the transmembrane region (thus designated as Syt VIDeltaTM). Because it lacks the cysteine motif responsible for self-dimerization, Syt VIDeltaTM could not associate with Syt VI even in the presence of Ca(2+). Despite lacking the transmembrane region, Syt VIDeltaTM can associate with the plasma membrane through the C-terminal 29 amino acids. In adult mouse brain, two closely comigrating bands at M(r) approximately 50,000, which closely corresponded to the molecular weight of recombinant Syt VIDeltaTM, were detected by anti-Syt VI antibody. These immunoreactive bands were found in both soluble and membrane fractions of mouse brain, indicating that they are membrane-associated proteins (Syt VIDeltaTM), but not transmembrane proteins (Syt VI). Expression of Syt VI and Syt VIDeltaTM in PC12 or COS-7 cells indicated that the two molecules have a distinct subcellular distribution: Syt VIDeltaTM is present in the cytosol or is associated with the plasma membrane or internal membrane structures, whereas Syt VI is localized to the endoplasmic reticulum and/or Golgi-like perinuclear compartment. These results suggest that Syt VI and Syt VIDeltaTM may play distinct roles in vesicular trafficking. PMID- 10531346 TI - Covalent homodimers of murine secretory component induced by epitope substitution unravel the capacity of the polymeric Ig receptor to dimerize noncovalently in the absence of IgA ligand. AB - Recombinant secretory immunoglobulin A containing a bacterial epitope in domain I of the secretory component (SC) moiety can serve as a mucosal delivery vehicle triggering both mucosal and systemic responses (Corthesy, B., Kaufmann, M., Phalipon, A., Peitsch, M., Neutra, M. R., and Kraehenbuhl, J.-P. (1996) J. Biol. Chem. 271, 33670-33677). To load recombinant secretory IgA with multiple B and T epitopes and extend its biological functions, we selected, based on molecular modeling, five surface-exposed sites in domains II and III of murine SC. Loops predicted to be exposed at the surface of SC domains were replaced with the DYKDDDDK octapeptide (FLAG). Another two mutants were obtained with the FLAG inserted in between domains II and III or at the carboxyl terminus of SC. As shown by mass spectrometry, internal substitution of the FLAG into four of the mutants induced the formation of disulfide-linked homodimers. Three of the dimers and two of the monomers from SC mutants could be affinity-purified using an antibody to the FLAG, mapping them as candidates for insertion. FLAG-induced dimerization also occurred with the polymeric immunoglobulin receptor (pIgR) and might reflect the so-far nondemonstrated capacity of the receptor to oligomerize. By co-expressing in COS-7 cells and epithelial Caco-2 cells two pIgR constructs tagged at the carboxyl terminus with hexahistidine or FLAG, we provide the strongest evidence reported to date that the pIgR dimerizes noncovalently in the plasma membrane in the absence of polymeric IgA ligand. The implication of this finding is discussed in terms of IgA transport and specific antibody response at mucosal surfaces. PMID- 10531345 TI - Different functional aspects of the group II subfamily (Types IIA and V) and type X secretory phospholipase A(2)s in regulating arachidonic acid release and prostaglandin generation. Implications of cyclooxygenase-2 induction and phospholipid scramblase-mediated cellular membrane perturbation. AB - We have recently reported that members of the heparin-binding group II subfamily of secretory PLA(2)s (sPLA(2)s) (types IIA and V), when transfected into 293 cells, released [(3)H]arachidonic acid (AA) preferentially in response to interleukin-1 (IL-1) and acted as "signaling" PLA(2)s that were functionally coupled with prostaglandin biosynthesis. Here we show that these group II subfamily sPLA(2)s and the type X sPLA(2) behave in a different manner, the former being more efficiently coupled with the prostaglandin-biosynthetic pathway than the latter, in 293 transfectants. Type X sPLA(2), which bound only minimally to cell surface proteoglycans, augmented the release of both [(3)H]AA and [(3)H]oleic acid in the presence of serum but not IL-1. Both types IIA and V sPLA(2), the AA released by which was efficiently converted to prostaglandin E(2), markedly augmented IL-1-induced expression of cyclooxygenase (COX)-2 in a heparin-sensitive fashion, whereas type X sPLA(2) lacked the ability to augment COX-2 expression, thereby exhibiting the poor prostaglandin E(2)-biosynthetic response unless either of the COX isozymes was forcibly introduced into type X sPLA(2)-expressing cells. Implication of phospholipid scramblase, an enzyme responsible for the perturbation of plasma membrane asymmetry, revealed that the scramblase-transfected cells became more sensitive to types IIA and V, but not X, sPLA(2), releasing both [(3)H]AA and [(3)H]oleic acid in an IL-1-independent manner. Thus, although phospholipid scramblase-mediated alteration in plasma membrane asymmetry actually led to the increased cellular susceptibility to the group II subfamily of sPLA(2)s, several lines of evidence suggest that it does not entirely mimic their actions on cells after IL-1 signaling. Interestingly, coexpression of type IIA or V, but not X, sPLA(2) and phospholipid scramblase resulted in a marked reduction in cell growth, revealing an unexplored antiproliferative aspect of particular classes of sPLA(2). PMID- 10531347 TI - Mapping the interaction between murine IgA and murine secretory component carrying epitope substitutions reveals a role of domains II and III in covalent binding to IgA. AB - We have identified sites for epitope insertion in the murine secretory component (SC) by replacing individual surface-exposed loops in domains I, II, and III with the FLAG sequence (Crottet, P., Peitsch, M. C., Servis, C., and Corthesy, B. (1999) J. Biol. Chem. 274, 31445-31455). We had previously shown that epitope carrying SC reassociated with dimeric IgA (IgA(d)) can serve as a mucosal delivery vehicle. When analyzing the capacity of SC mutants to associate with IgA(d), we found that all domain II and III mutants bound specifically with immobilized IgA(d), and their affinity for IgA(d) was comparable to that of the wild type protein (IC(50) approximately 1 nM). We conclude that domains II and III in SC are permissive to local mutation and represent convenient sites to antigenize the SC molecule. No mutant bound to monomeric IgA. SC mutants exposing the FLAG at their surface maintained this property once bound to IgA(d), thereby defining regions not required for high affinity binding to IgA(d). Association of IgA(d) with SC mutants carrying a buried FLAG did not expose de novo the epitope, consistent with limited, local changes in the SC structure upon binding. Only wild type and two mutant SCs bound covalently to IgA(d), thus implicating domains II and III in the correct positioning of the reactive cysteine in SC. This establishes that the integrity of murine SC domains II and III is not essential to preserve specific IgA(d) binding but is necessary for covalency to take place. Finally, SC mutants existing in the monomeric and dimeric forms exhibited the same IgA(d) binding capacity as monomeric wild type SC known to bind with a 1:1 stoichiometry. PMID- 10531348 TI - Role of human Cds1 (Chk2) kinase in DNA damage checkpoint and its regulation by p53. AB - In response to DNA damage, mammalian cells adopt checkpoint regulation, by phosphorylation and stabilization of p53, to delay cell cycle progression. However, most cancer cells that lack functional p53 retain an unknown checkpoint mechanism(s) by which cells are arrested at the G(2)/M phase. Here we demonstrate that a human homolog of Cds1/Rad53 kinase (hCds1) is rapidly phosphorylated and activated in response to DNA damage not only in normal cells but in cancer cells lacking functional p53. A survey of various cancer cell lines revealed that the expression level of hCds1 mRNA is inversely related to the presence of functional p53. In addition, transfection of normal human fibroblasts with SV40 T antigen or human papilloma viruses E6 or E7 causes a marked induction of hCds1 mRNA, and the introduction of functional p53 into SV40 T antigen- and E6-, but not E7-, transfected cells decreases the hCds1 level, suggesting that p53 negatively regulates the expression of hCds1. In cells without functional ataxia telangiectasia mutated (ATM) protein, phosphorylation and activation of hCds1 were observed in response to DNA damage induced by UV but not by ionizing irradiation. These results suggest that hCds1 is activated through an ATM dependent as well as -independent pathway and that it may complement the function of p53 in DNA damage checkpoints in mammalian cells. PMID- 10531349 TI - Acidic pH modulates the interaction between human granulocyte-macrophage colony stimulating factor and glycosaminoglycans. AB - Granulocyte-macrophage colony-stimulating factor (GM-CSF) controls growth and differentiation of hematopoietic cells. Previous reports have indicated that the mitogenic activity of GM-CSF may be modulated by the glycosidic moiety of proteoglycans associated with the membrane of stromal cells. In this work, we have performed in vitro studies of the interaction between GM-CSF and glycosaminoglycans. The addition of heparin promoted a marked blue shift in the fluorescence emission spectrum of GM-CSF as well as a 30-fold increase in the intensity of light scattering, which indicates formation of large molecular weight complexes between the two molecules. Interestingly, heparin-induced changes in the spectral properties of GM-CSF were only observed at acidic pH. The dependence on acidic pH, together with a strict dependence on glycosaminoglycan sulfation and the fact that high ionic strength destabilized the interaction, indicates that the association between GM-CSF and glycosaminoglycans is mediated by electrostatic interactions. These interactions probably involve sulfate groups in the glycosaminoglycans and positively charged histidine residues in GM-CSF. We propose that negatively charged glycolipids present on the plasma membrane of the hematopoietic and/or the stromal cell could promote an acidic microenvironment capable of triggering interaction between GM-CSF and membrane-bound proteoglycans in vivo. PMID- 10531350 TI - Low molecular weight group IIA and group V phospholipase A(2) enzymes have different intracellular locations in mouse bone marrow-derived mast cells. AB - The subcellular location of the enzymes of eicosanoid biosynthesis is critical for their co-ordinate action in the generation of leukotrienes and prostaglandins. This activity is thought to occur predominantly at a perinuclear location. Whereas the subcellular locations of cytosolic phospholipase (PL) A(2) and each of the pathway enzymes of eicosanoid generation have been defined, the distribution of the low molecular weight species of PLA(2) has remained elusive because of the lack of antibodies that distinguish among homologous family members. We have prepared affinity-purified rabbit antipeptide IgG antibodies that distinguish mouse group IIA PLA(2) and group V PLA(2). Immunofluorescence staining and immunogold electron microscopy reveal different subcellular locations for the enzymes. Group IIA(2) PLA(2) is present in the secretory granules of mouse bone marrow-derived mast cells, consistent with its putative role in facilitating secretory granule exocytosis and its consequent extracellular action. In contrast, group V PLA(2) is associated with various membranous organelles including the Golgi apparatus, nuclear envelope, and plasma membrane. The perinuclear location of group V PLA(2) is consistent with a putative interaction with translocated cytosolic PLA(2) in supplying arachidonic acid for generation of eicosanoid products, while the location in Golgi cisternae may also reflect its action as a secreted enzyme. The spatial segregation of group IIA PLA(2) and group V PLA(2) implies that these enzymes are not functionally redundant. PMID- 10531351 TI - Functional characterization of ABC10alpha, an essential polypeptide shared by all three forms of eukaryotic DNA-dependent RNA polymerases. AB - ABC10alpha is a small polypeptide shared by the three yeast RNA polymerases. Homologous polypeptides in higher eukaryotes have a zinc-binding CX(2)C. CX(2)C motif and a conserved basic C-terminal end. These features are also found in archaeal gene products that may encode an RNA polymerase subunit. The CX(2)C. CX(2)C motif is partly dispensable, since only its first cysteine is essential for growth, whereas the basic C-terminal end is critical in vivo. A mutant in the latter domain has an RNA polymerase III-specific defect and, in vitro, impairs RNA polymerase III assembly. Polymerase activity was, however, not affected in various faithful transcription assays. The mutant is suppressed by a high gene dosage of the second largest subunit of RNA polymerase III, whereas the homologous subunits of RNA polymerase I and II have aggravating effects. In a two hybrid assay, ABC10alpha binds to the C-terminal half of the second largest subunit of RNA polymerase I, in a way that requires the integrity of the CX(2)C. CX(2)C motif. Thus, ABC10alpha appears to interact directly with the second largest subunit during polymerase assembly. This interaction is presumably a major rate-limiting step in assembly, since diploid cells containing only one functional gene copy for ABC10alpha have a partial growth defect. PMID- 10531352 TI - "RKKH" peptides from the snake venom metalloproteinase of Bothrops jararaca bind near the metal ion-dependent adhesion site of the human integrin alpha(2) I domain. AB - Integrin alpha(1)beta(1) and alpha(2)beta(1) are the major cellular receptors for collagen, and collagens bind to these integrins at the inserted I-domain in their alpha subunit. We have previously shown that a cyclic peptide derived from the metalloproteinase domain of the snake venom protein jararhagin blocks the collagen-binding function of the alpha(2) I-domain. Here, we have optimized the structure of the peptide and identified the site where the peptide binds to the alpha(2) I-domain. The peptide sequence Arg-Lys-Lys-His is critical for recognition by the I-domain, and five negatively charged residues surrounding the "metal ion-dependent adhesion site" (MIDAS) of the I-domain, when mutated, show significantly impaired binding of the peptide. Removal of helix alphaC, located along one side of the MIDAS and suggested to be involved in collagen-binding in these I-domains, does not affect peptide binding. This study supports the notion that the metalloproteinase initially binds to the alpha(2) I-domain at a location distant from the active site of the protease, thus blocking collagen binding to the adhesion molecule in the vicinity of the MIDAS, while at the same time leaving the active site free to degrade nearby proteins, the closest being the beta(1) subunit of the alpha(2)beta(1) cell-surface integrin itself. PMID- 10531353 TI - Lethal kinesin mutations reveal amino acids important for ATPase activation and structural coupling. AB - To study the relationship between conventional kinesin's structure and function, we identified 13 lethal mutations in the Drosophila kinesin heavy chain motor domain and tested a subset for effects on mechanochemistry. S246F is a moderate mutation that occurs in loop 11 between the ATP- and microtubule-binding sites. While ATP and microtubule binding appear normal, there is a 3-fold decrease in the rate of ATP turnover. This is consistent with the hypothesis that loop 11 provides a structural link that is important for the activation of ATP turnover by microtubule binding. T291M is a severe mutation that occurs in alpha-helix 5 near the center of the microtubule-binding surface. It impairs the microtubule kinesin interaction and directly effects the ATP-binding pocket, allowing an increase in ATP turnover in the absence of microtubules. The T291M mutation may mimic the structure of a microtubule-bound, partially activated state. E164K is a moderate mutation that occurs at the beta-sheet 5a/loop 8b junction, remote from the ATP pocket. Surprisingly, it causes both tighter ATP-binding and a 2-fold decrease in ATP turnover. We propose that E164 forms an ionic bridge with alpha helix 5 and speculate that it helps coordinate the alternating site catalysis of dimerized kinesin heavy chain motor domains. PMID- 10531354 TI - Properties of secretin receptor internalization differ from those of the beta(2) adrenergic receptor. AB - The endocytic pathway of the secretin receptor, a class II GPCR, is unknown. Some class I G protein-coupled receptors (GPCRs), such as the beta(2)-adrenergic receptor (beta(2)-AR), internalize in clathrin-coated vesicles and this process is mediated by G protein-coupled receptor kinases (GRKs), beta-arrestin, and dynamin. However, other class I GPCRs, for example, the angiotensin II type 1A receptor (AT(1A)R), exhibit different internalization properties than the beta(2) AR. The secretin receptor, a class II GPCR, is a GRK substrate, suggesting that like the beta(2)-AR, it may internalize via a beta-arrestin and dynamin directed process. In this paper we characterize the internalization of a wild-type and carboxyl-terminal (COOH-terminal) truncated secretin receptor using flow cytometry and fluorescence imaging, and compare the properties of secretin receptor internalization to that of the beta(2)-AR. In HEK 293 cells, sequestration of both the wild-type and COOH-terminal truncated secretin receptors was unaffected by GRK phosphorylation, whereas inhibition of cAMP dependent protein kinase mediated phosphorylation markedly decreased sequestration. Addition of secretin to cells resulted in a rapid translocation of beta-arrestin to plasma membrane localized receptors; however, secretin receptor internalization was not reduced by expression of dominant negative beta-arrestin. Thus, like the AT(1A)R, secretin receptor internalization is not inhibited by reagents that interfere with clathrin-coated vesicle-mediated internalization and in accordance with these results, we show that secretin and AT(1A) receptors colocalize in endocytic vesicles. This study demonstrates that the ability of secretin receptor to undergo GRK phosphorylation and beta-arrestin binding is not sufficient to facilitate or mediate its internalization. These results suggest that other receptors may undergo endocytosis by mechanisms used by the secretin and AT(1A) receptors and that kinases other than GRKs may play a greater role in GPCR endocytosis than previously appreciated. PMID- 10531355 TI - Ethylene modulates gene expression in cells of the marine sponge Suberites domuncula and reduces the degree of apoptosis. AB - Sponges (phylum Porifera) live in an aqueous milieu that contains dissolved organic carbon. This is degraded photochemically by ultraviolet radiation to alkenes, particularly to ethylene. This study demonstrates that sponge cells (here the demosponge Suberites domuncula has been used), which have assembled to primmorphs, react to 5 microM ethylene with a significant up-regulation of intracellular Ca(2+) concentration and with a reduction of starvation-induced apoptosis. In primmorphs from S. domuncula the expression of two genes is up regulated after exposure to ethylene. The cDNA of the first gene (SDERR) isolated from S. domuncula encodes a potential ethylene-responsive protein, termed ERR_SUBDO; its putative M(r) is 32,704. Data bank search revealed that the sponge polypeptide shares high similarity (82% on amino acid level) with the corresponding plant molecule, the ethylene-inducible protein from Hevea brasiliensis. Until now no other metazoan ethylene-responsive proteins have been identified. The second gene, whose expression is up-regulated in response to ethylene is a Ca(2+)/calmodulin-dependent protein kinase II. Its cDNA, SDCCdPK, encodes a M(r) 54,863 putative kinase that shares 69% similarity with the corresponding enzyme from Drosophila melanogaster. The expression of both genes in primmorphs from S. domuncula is increased by approximately 5-fold after a 3 day incubation period with ethylene. It is concluded that also metazoan cells, with sponge cells as a model, may react to ethylene with an activation of cell metabolism including gene induction. PMID- 10531356 TI - The human homologue of the yeast proteins Skb1 and Hsl7p interacts with Jak kinases and contains protein methyltransferase activity. AB - To expand our understanding of the role of Jak2 in cellular signaling, we used the yeast two-hybrid system to identify Jak2-interacting proteins. One of the clones identified represents a human homologue of the Schizosaccaromyces pombe Shk1 kinase-binding protein 1, Skb1, and the protein encoded by the Saccharomyces cerevisiae HSL7 (histone synthetic lethal 7) gene. Since no functional motifs or biochemical activities for this protein or its homologues had been reported, we sought to determine a biochemical function for this human protein. We demonstrate that this protein is a protein methyltransferase. This protein, designated JBP1 (Jak-binding protein 1), and its homologues contain motifs conserved among protein methyltransferases. JBP1 can be cross-linked to radiolabeled S adenosylmethionine (AdoMet) and methylates histones (H2A and H4) and myelin basic protein. Mutants containing substitutions within a conserved region likely to be involved in AdoMet binding exhibit little or no activity. We mapped the JBP1 gene to chromosome 14q11.2-21. In addition, JBP1 co-immunoprecipitates with several other proteins, which serve as methyl group acceptors and which may represent physiological targets of this methyltransferase. Messenger RNA for JBP1 is widely expressed in human tissues. We have also identified and sequenced a homologue of JBP1 in Drosophila melanogaster. This report provides a clue to the biochemical function for this conserved protein and suggests that protein methyltransferases may have a role in cellular signaling. PMID- 10531357 TI - The homeodomain transcription factor NK-4 acts as either a transcriptional activator or repressor and interacts with the p300 coactivator and the Groucho corepressor. AB - NK-4 (tinman) encodes an NK-2 class homeodomain transcription factor that is required for development of the Drosophila dorsal mesoderm, including heart. Genetic evidence suggests its important role in mesoderm subdivision, yet the properties of NK-4 as a transcriptional regulator and the mechanism of gene transcription by NK-4 are not completely understood. Here, we describe its properties as a transcription factor and its interaction with the p300 coactivator and the Groucho corepressor. We demonstrate that NK-4 can activate or repress target genes in cultured cells, depending on functional domains that are conserved between Drosophila melanogaster and Drosophila virilis NK-4 genes. Using GAL4-NK-4 fusion constructs, we have mapped a transcriptional activation domain (amino acids 1-110) and repression domains (amino acids 111-188 and the homeodomain) and found an inhibitory function for the homeodomain in transactivation by NK-4. Furthermore, we demonstrate that NK-4-dependent transactivation is augmented by the p300 coactivator and show that NK-4 physically interacts with p300 via the activation domain. In addition, cotransfection experiments indicate that the repressor activity of NK-4 is strongly enhanced by the Groucho corepressor. Using immunoprecipitation and in vitro pull-down assays, we show that NK-4 directly interacts with the Groucho corepressor, for which the homeodomain is required. Together, our results indicate that NK-4 can act as either a transcriptional activator or repressor and provide the first evidence of NK-4 interactions with the p300 coactivator and the Groucho corepressor. PMID- 10531358 TI - Cloning and characterization of a novel human phosphatidylinositol transfer protein, rdgBbeta. AB - The various PITP, retinal degeneration B (rdgB), and amino-terminal domain interacting receptor (Nir) phosphatidylinositol transfer proteins can be divided into two structural families. The small, soluble PITP isoforms contain only a phosphatidylinositol transfer domain and have been implicated in phosphoinositide signaling and vesicle trafficking. In contrast, the rdgB proteins, which include Nir2 and Nir3, contain an amino-terminal PITP-like domain, an acidic, Ca(2+) binding domain, six putative transmembrane domains, and a conserved carboxyl terminal domain. However, the biological function of rdgB proteins is unclear. Here, we report the isolation of a cDNA encoding a novel rdgB protein, mammalian rdgBbeta (MrdgBbeta). The 38-kDa MrdgBbeta protein contains an amino-terminal PITP-like domain and a short carboxyl-terminal domain. In contrast to other rdgB like proteins, MrdgBbeta contains no transmembrane motifs or the conserved carboxyl-terminal domain. Using Northern and reverse transcription-polymerase chain reaction analysis, we demonstrate that MrdgBbeta mRNA is ubiquitously expressed. Immunofluorescence analysis of ectopic MrdgBbeta showed cytoplasmic staining, and the ability of recombinant MrdgBbeta to transfer phosphatidylinositol in vitro was similar to other PITP-like domains. Although early reports found functional degeneracy in vitro, the identification of a fifth mammalian PITP-like protein with a unique domain organization and widespread expression supports more recent results that suggest that different PITP-like domains have distinct functions in vivo. PMID- 10531359 TI - Role of the cyclic AMP-protein kinase A pathway in lipopolysaccharide-induced nitric oxide synthase expression in RAW 264.7 macrophages. Involvement of cyclooxygenase-2. AB - The signaling pathway for lipopolysaccharide (LPS)-induced nitric oxide (NO) release in RAW 264.7 macrophages involves the protein kinase C and p38 activation pathways (Chen, C. C., Wang, J. K., and Lin, S. B. (1998) J. Immunol. 161, 6206 6214; Chen, C. C., and Wang, J. K. (1999) Mol. Pharmacol. 55, 481-488). In this study, the role of the cAMP-dependent protein kinase A (PKA) pathway was investigated. The PKA inhibitors, KT-5720 and H8, reduced LPS-induced NO release and inducible nitric oxide synthase (iNOS) expression. The direct PKA activator, Bt(2)cAMP, caused concentration-dependent NO release and iNOS expression, as confirmed by immunofluorescence studies. The intracellular cAMP concentration did not increase until after 6 h of LPS treatment. Two cAMP-elevating agents, forskolin and cholera toxin, potentiated the LPS-induced NO release and iNOS expression. Stimulation of cells with LPS or Bt(2)cAMP for periods of 10 min to 24 h caused nuclear factor-kappaB (NF-kappaB) activation in the nuclei, as shown by detection of NF-kappaB-specific DNA-protein binding. The PKA inhibitor, H8, inhibited the NF-kappaB activation induced by 6- or 12-h treatment with LPS but not that induced after 1, 3, or 24 h. The cyclooxygenase-2 (COX-2) inhibitors, NS 398 and indomethacin, attenuated LPS-induced NO release, iNOS expression, and NF kappaB DNA-protein complex formation. LPS induced COX-2 expression in a time dependent manner, and prostaglandin E(2) production was induced in parallel. These results suggest that 6 h of treatment with LPS increases intracellular cAMP levels via COX-2 induction and prostaglandin E(2) production, resulting in PKA activation, NF-kappaB activation, iNOS expression, and NO production. PMID- 10531360 TI - Generation of a dominant-negative mutant of endothelial PAS domain protein 1 by deletion of a potent C-terminal transactivation domain. AB - Endothelial PAS domain protein 1 (EPAS1) is a basic helix-loop-helix/PAS domain transcription factor that is preferentially expressed in vascular endothelial cells. EPAS1 shares high homology with hypoxia-inducible factor-1alpha (HIF 1alpha) and, like HIF-1alpha, has been shown to bind to the HIF-1-binding site and to activate its downstream genes such as vascular endothelial growth factor (VEGF) and erythropoietin. In this report, we show that EPAS1 increased VEGF gene expression through the HIF-1-binding site. This transactivation was enhanced further by cotransfection of an aryl hydrocarbon receptor nuclear translocator expression plasmid. Deletion analysis of EPAS1 revealed a potent activation domain (amino acids 486-639) essential for EPAS1 to transactivate the VEGF promoter. We confirmed the ability of this domain to activate transcription using a Gal4 fusion protein system. Because a truncated EPAS1 protein lacking the transactivation domain at amino acids 486-639 eliminated induction of the VEGF promoter by wild-type EPAS1, the truncated protein functions as a dominant negative mutant. Most important, infection of the cells with an adenoviral construct expressing this mutant inhibited the induction of VEGF mRNA under conditions that mimic hypoxia. Our results suggest that EPAS1 is an important regulator of VEGF gene expression. Since VEGF plays a crucial role in angiogenesis, the ability of dominant-negative EPAS1 to inhibit VEGF promoter activity raises the possibility of a novel approach to inhibiting pathological angiogenesis. PMID- 10531361 TI - Visinin-like protein (VILIP) is a neuron-specific calcium-dependent double stranded RNA-binding protein. AB - Double-stranded RNA-binding proteins function in regulating the stability, translation, and localization of specific mRNAs. In this study, we have demonstrated that the neuron-specific, calcium-binding protein, visinin-like protein (VILIP) contains one double-stranded RNA-binding domain, a protein motif conserved among many double-stranded RNA-binding proteins. We showed that VILIP can specifically bind double-stranded RNA, and this interaction specifically requires the presence of calcium. Mobility shift studies indicated that VILIP binds double-stranded RNA as a single protein-RNA complex with an apparent equilibrium dissociation constant of 9.0 x 10(-6) M. To our knowledge, VILIP is the first double-stranded RNA-binding protein shown to be calcium-dependent. Furthermore, VILIP specifically binds the 3'-untranslated region of the neurotrophin receptor, trkB, an mRNA localized to hippocampal dendrites in an activity-dependent manner. Given that VILIP is also expressed in the hippocampus, these data suggest that VILIP may employ a novel, calcium-dependent mechanism to regulate its binding to important localized mRNAs in the central nervous system. PMID- 10531362 TI - Interaction and functional cooperation of PEBP2/CBF with Smads. Synergistic induction of the immunoglobulin germline Calpha promoter. AB - Smads are signal transducers for members of the transforming growth factor-beta (TGF-beta) superfamily. Upon ligand stimulation, receptor-regulated Smads (R Smads) are phosphorylated by serine/threonine kinase receptors, form complexes with common-partner Smad, and translocate into the nucleus, where they regulate the transcription of target genes together with other transcription factors. Polyomavirus enhancer binding protein 2/core binding factor (PEBP2/CBF) is a transcription factor complex composed of alpha and beta subunits. The alpha subunits of PEBP2/CBF, which contain the highly conserved Runt domain, play essential roles in hematopoiesis and osteogenesis. Here we show that three mammalian alpha subunits of PEBP2/CBF form complexes with R-Smads that act in TGF beta/activin pathways as well as those acting in bone morphogenetic protein (BMP) pathways. Among them, PEBP2alphaC/CBFA3/AML2 forms a complex with Smad3 and stimulates transcription of the germline Ig Calpha promoter in a cooperative manner, for which binding of both factors to their specific binding sites is essential. PEBP2 may thus be a nuclear target of TGF-beta/BMP signaling. PMID- 10531363 TI - Phospho-carboxyl-terminal domain binding and the role of a prolyl isomerase in pre-mRNA 3'-End formation. AB - A phospho-carboxyl-terminal domain (CTD) affinity column created with yeast CTD kinase I and the CTD of RNA polymerase II was used to identify Ess1/Pin1 as a phospho-CTD-binding protein. Ess1/Pin1 is a peptidyl prolyl isomerase involved in both mitotic regulation and pre-mRNA 3'-end formation. Like native Ess1, a GSTEss1 fusion protein associates specifically with the phosphorylated but not with the unphosphorylated CTD. Further, hyperphosphorylated RNA polymerase II appears to be the dominant Ess1 binding protein in total yeast extracts. We demonstrate that phospho-CTD binding is mediated by the small WW domain of Ess1 rather than the isomerase domain. These findings suggest a mechanism in which the WW domain binds the phosphorylated CTD of elongating RNA polymerase II and the isomerase domain reconfigures the CTD though isomerization of proline residues perhaps by a processive mechanism. This process may be linked to a variety of pre mRNA maturation events that use the phosphorylated CTD, including the coupled processes of pre-mRNA 3'-end formation and transcription termination. PMID- 10531364 TI - Contribution of the ERK5/MEK5 pathway to Ras/Raf signaling and growth control. AB - The activity of the catalytic domain of the orphan MAP kinase ERK5 is increased by Ras but not Raf-1 in cells, which suggests that ERK5 might mediate Raf independent signaling by Ras. We found that Raf-1 does contribute to Ras activation of ERK5 but in a manner that does not correlate with Raf-1 catalytic activity. A clue to the mechanism of action of Raf-1 on ERK5 comes from the observation that endogenous Raf-1 binds to endogenous ERK5, suggesting the involvement of regulatory protein-protein interactions. This interaction is specific because Raf-1 binds only to ERK5 and not ERK2 or SAPK. Finally, we demonstrate the ERK5/MEK5 pathway is required for Raf-dependent cellular transformation and that a constitutively active form of MEK5, MEK5DD, synergizes with Raf to transform NIH 3T3 cells. These observations suggest that ERK5 plays a large role in Raf-1-mediated signal transduction. PMID- 10531365 TI - Ca(2+)-dependent association of S100A6 (Calcyclin) with the plasma membrane and the nuclear envelope. AB - Expression of S100A6 (Calcyclin), a member of the S100 family and of Zn(2+) binding proteins is elevated in a number of malignant tumors. In vitro the protein associates with several actin-binding proteins and annexins in a Ca(2+) dependent manner. We have now studied the subcellular localization of S100A6 using a new, specific monoclonal antibody. Immunofluorescence microscopy of unfixed, ultrathin, frozen sections demonstrated a dual localization of S100A6 at the nuclear envelope and the plasma membrane of porcine smooth muscle only in the presence of Ca(2+). The same localization was found by immunofluorescence and immunogold electron microscopy as well as by confocal laser scanning microscopy with cultured, fixed, human CaKi-2 and porcine ST interphase cells. Upon cell division, however, S100A6 was found exclusively in the cytoplasm. Cell fractionation studies showed that S100A6 was present in the microsomal fraction in the presence of Ca(2+) and was released from this fraction by the addition of EGTA/EDTA but not by Triton X-100. The data demonstrate that S100A6 is localized both at the plasma membrane and the nuclear envelope in vivo and suggest a Ca(2+) dependent interaction with annexins or other components of the nuclear envelope. PMID- 10531366 TI - CCAAT/enhancer-binding protein beta (C/EBPbeta) and C/EBPdelta contribute to growth hormone-regulated transcription of c-fos. AB - Using the c-fos enhancer as a model to analyze growth hormone (GH)-promoted gene expression, the roles of CCAAT/enhancer-binding proteins (C/EBPs) in GH-regulated transcription were investigated. In 3T3-F442A fibroblasts stably expressing the c fos promoter mutated at the C/EBP binding site upstream of luciferase, c-fos promoter activity is stimulated by GH 6-7-fold; wild type c-fos promoter shows only a 2-fold induction by GH. This suggests that C/EBP restrains GH-stimulated expression of c-fos. Electrophoretic mobility shift assays with nuclear extracts from 3T3-F442A cells indicate that GH rapidly (2-5 min) increases binding of C/EBPbeta and C/EBPdelta, to the c-fos C/EBP binding site. Both liver activating protein (LAP) and liver inhibitory protein (LIP), forms of C/EBPbeta, are detected in 3T3-F442A cells by immunoblotting. GH increases the binding of LAP/LAP and LAP/LIP dimers. Overexpression of LIP interferes with GH-promoted reporter expression in CHO cells expressing GH receptors, consistent with the possibility that LIP restrains GH-stimulated c-fos expression. Overexpression of LAP elevates basal luciferase activity but does not influence promoter activation by GH, while overexpressed C/EBPdelta elevates basal promoter activity and enhances the stimulation by GH. GH stimulates the expression of mRNA for C/EBPbeta and -delta and increases levels of C/EBPdelta. Although C/EBPbeta is not detectably altered, GH induces a shift to more rapidly migrating forms of LIP and LAP upon immunoblotting. Treatment of extracts from GH-treated cells with alkaline phosphatase causes a shift of the slower migrating form to the rapidly migrating form, consistent with GH promoting dephosphorylation of LIP and LAP. These studies implicate C/EBPbeta and -delta in GH-regulated gene expression. They also indicate that GH stimulates the binding of C/EBPbeta and -delta to the c-fos promoter and promotes the dephosphorylation of LIP and LAP. These events may contribute to the ability of C/EBPbeta and -delta to regulate GH-stimulated expression of c-fos. PMID- 10531367 TI - Binding of pigment epithelium-derived factor (PEDF) to retinoblastoma cells and cerebellar granule neurons. Evidence for a PEDF receptor. AB - Pigment epithelium-derived factor (PEDF) has neuronal differentiation and survival activity on retinoblastoma and cerebellar granule (CG) cells. Here, we investigated the presence of PEDF receptors on retinoblastoma Y-79 and CG cells. PEDF radiolabeled with (l25)I remained biologically active and was used for radioligand binding analysis. The binding was saturable and specific to a single class of receptors on both cells and with similar affinities (K(d) = 1.7-3.6 nM, B(max) = 0.5-2.7 x 10(5) sites/Y-79 cell; and K(d) = 3.2 nM, B(max) = 1.1 x 10(3) sites/CG cell). A polyclonal antiserum to PEDF, previously shown to block the PEDF neurotrophic activity, prevented the (125)I-PEDF binding. We designed two peptides from a region previously shown to confer the neurotrophic property to human PEDF, synthetic peptides 34-mer (positions 44-77) and 44-mer (positions 78 121). Only peptide 44-mer competed for the binding to Y-79 cell receptors (EC(50) = 5 nM) and exhibited neuronal differentiating activity. PEDF affinity column chromatography of membrane proteins from both cell types revealed a PEDF-binding protein of approximately 80 kDa. These results are the first demonstration of a PEDF-binding protein with characteristics of a PEDF receptor and suggest that the region comprising amino acid positions 78-121 of PEDF might be involved in ligand receptor interactions. PMID- 10531368 TI - Direct evidence for immiscible cholesterol domains in human ocular lens fiber cell plasma membranes. AB - The molecular structure of human ocular lens fiber cell plasma membranes was examined directly using small angle x-ray diffraction approaches. A distinct biochemical feature of these membranes is their high relative levels of free cholesterol; the mole ratio of cholesterol to phospholipid (C/P) measured in these membranes ranges from 1 to 4. The organization of cholesterol in this membrane system is not well understood, however. In this study, the structure of plasma membrane samples isolated from nuclear (3.3 C/P) and cortical (2.4 C/P) regions of human lenses was evaluated with x-ray diffraction approaches. Meridional diffraction patterns obtained from the oriented membrane samples demonstrated the presence of an immiscible cholesterol domain with a unit cell periodicity of 34.0 A, consistent with a cholesterol monohydrate bilayer. The dimensions of the sterol-rich domains remained constant over a broad range of temperatures (5-20 degrees C) and relative humidity levels (31-97%). In contrast, dimensions of the surrounding sterol-poor phase were significantly affected by experimental conditions. Similar structural features were observed in membranes reconstituted from fiber cell plasma membrane lipid extracts. The results of this study indicate that the lens fiber cell plasma membrane is a complex structure consisting of separate sterol-rich and -poor domains. Maintenance of these separate domains may be required for the normal function of lens fiber cell plasma membrane and may interfere with the cataractogenic aggregation of soluble lens proteins at the membrane surface. PMID- 10531369 TI - Binding of the human papillomavirus type 16 p97 promoter by the adeno-associated virus Rep78 major regulatory protein correlates with inhibition. AB - Human papillomavirus type 16 (HPV-16) infection is positively associated with cervical cancer, whereas adeno-associated virus (AAV) infection is negatively associated with this same cancer. In earlier studies these two virus types have been shown to directly interact, with AAV inhibiting or enhancing papillomavirus functions depending upon the specific circumstances. One defined interaction between these two viruses is the ability of the AAV Rep78 major regulatory protein to inhibit gene expression of the E6 promoter of BPV-1 (bovine papillomavirus type 1) and HPV types 16 and 18. As Rep78 is a DNA binding transcription factor, we considered whether Rep78 might bind HPV-16 DNA. Here, Rep78 is demonstrated to bind a 44-base pair region (nucleotides 14-56) within the HPV-16 p97 promoter using the electrophoretic mobility shift assay. This region is important for HPV-16 because it includes functional Sp1 and E2 protein binding motifs as well as part of the origin of replication. Furthermore, two Rep78 amino acid substitution mutants, at positions 77 or 64-65, were identified that did not recognize p97 DNA. Both of these Rep78 mutants were found to be defective for inhibition of p97 promoter activity in HeLa and T-47D nuclear extracts in vitro, in a transient chloramphenicol acetyltransferase assay, as well as defective for full inhibition of HPV-16-directed focus formation. These data, taken together, strongly suggest that the Rep78-p97 promoter interaction is at least partially responsible for Rep78-mediated inhibition of HPV-16. Finally, the finding that Rep78 specifically recognizes p97 DNA is surprising because the p97 promoter region contains no GAGC motifs, the core motif for Rep78 recognition. These data suggest that the p97 promoter may represent a new prototypical DNA target type for Rep78. PMID- 10531370 TI - The pimB gene of Mycobacterium tuberculosis encodes a mannosyltransferase involved in lipoarabinomannan biosynthesis. AB - The biosynthesis of lipoarabinomannan (LAM), a key mycobacterial lipoglycan that has been implicated in numerous immunoregulatory functions, was examined utilizing D-mannosamine (ManN) as a tool to identify mannosyltransferase genes involved in LAM synthesis. Cell-free reactions utilizing cellular membranes of mycobacteria as the enzyme source indicated that ManN inhibited the synthesis of phosphatidylinositol mannosides, early precursors to LAM. A selection strategy was devised to screen a Mycobacterium tuberculosis genomic library in Mycobacterium smegmatis for clones conferring conditional resistance to ManN, with the rationale that overexpression of the gene(s) encoding a target of ManN would impart a ManN-resistant phenotype under these conditions. This strategy led to the identification of pimB, whose deduced amino acid sequence shows similarity to mannosyltransferases and other glycosyltransferases. Partially purified recombinant PimB protein from Escherichia coli or membranes from M. smegmatis overexpressing the pimB gene were used in cell-free assays to show that PimB catalyzes the formation of triacylphosphatidylinositol dimannoside from GDP mannose and triacylphosphatidylinositol monomannoside. PMID- 10531371 TI - Carboxypeptidase M, a glycosylphosphatidylinositol-anchored protein, is localized on both the apical and basolateral domains of polarized Madin-Darby canine kidney cells. AB - Carboxypeptidase M, a glycosylphosphatidylinositol-anchored membrane glycoprotein, is highly expressed in Madin-Darby canine kidney (MDCK) cells, where it was previously shown that the glycosylphosphatidylinositol anchor and N linked carbohydrate are apical targeting signals. Here, we show that carboxypeptidase M has an unusual, non-polarized distribution, with up to 44% on the basolateral domain of polarized MDCK cells grown on semipermeable inserts. Alkaline phosphatase, as well as five other glycosylphosphatidylinositol-anchored proteins, and transmembrane gamma-glutamyl transpeptidase exhibited the expected apical localization. Basolateral carboxypeptidase M was readily released by exogenous phosphatidylinositol-specific phospholipase C, showing it is glycosylphosphatidylinositol-anchored, whereas apical carboxypeptidase M was more resistant to release. In contrast, the spontaneous release of carboxypeptidase M into the medium was much higher on the apical than the basolateral domain. In pulse-chase studies, newly synthesized carboxypeptidase M arrived in equal amounts within 30 min on both domains, indicating direct sorting. After 4-8 h of chase, the steady-state distribution was attained, possibly due to transcytosis from the basolateral to the apical domain. These data suggest the presence of a unique basolateral targeting signal in carboxypeptidase M that competes with its apical targeting signals, resulting in a non-polarized distribution in MDCK cells. PMID- 10531372 TI - Reciprocal signaling between heterotrimeric G proteins and the p21-stimulated protein kinase. AB - p21-activated protein kinase (PAK)-1 phosphorylated Galpha(z), a member of the Galpha(i) family that is found in the brain, platelets, and adrenal medulla. Phosphorylation approached 1 mol of phosphate/mol of Galpha(z) in vitro. In transfected cells, Galpha(z) was phosphorylated both by wild-type PAK1 when stimulated by the GTP-binding protein Rac1 and by constitutively active PAK1 mutants. In vitro, phosphorylation occurred only at Ser(16), one of two Ser residues that are the major substrate sites for protein kinase C (PKC). PAK1 did not phosphorylate other Galpha subunits (i1, i2, i3, o, s, or q). PAK1 phosphorylated Galpha(z) was resistant both to RGSZ1, a G(z)-selective GTPase activating protein (GAP), and to RGS4, a relatively nonselective GAP for the G(i) and G(q) families of G proteins. Phosphorylation of Ser(27) by PKC did not alter sensitivity to either GAP. The previously described inhibition of G(z) GAPs by PKC is therefore mediated by phosphorylation of Ser(16). Phosphorylation of either Ser(16) by PAK1 or Ser(27) by PKC decreased the affinity of Galpha(z) for Gbetagamma; phosphorylation of both residues by PKC caused no further effect. PAK1 thus regulates Galpha(z) function by attenuating the inhibitory effects of both GAPs and Gbetagamma. In this context, the kinase activity of PAK1 toward several protein substrates was directly inhibited by Gbetagamma, suggesting that PAK1 acts as a Gbetagamma-regulated effector protein. This inhibition of mammalian PAK1 by Gbetagamma contrasts with the stimulation of the PAK homolog Ste20p in Saccharomyces cerevisiae by the Gbetagamma homolog Ste4p/Ste18p. PMID- 10531373 TI - Inhibition of extracellular signal-regulated protein kinase or c-Jun N-terminal protein kinase cascade, differentially activated by cisplatin, sensitizes human ovarian cancer cell line. AB - We have studied the roles of c-Jun N-terminal protein kinase (JNK) and extracellular signal-regulated protein kinase (ERK) cascade in both the cisplatin resistant Caov-3 and the cisplatin-sensitive A2780 human ovarian cancer cell lines. Treatment of both cells with cisplatin but not transplatin isomer activates JNK and ERK. Activation of JNK by cisplatin occurred at 30 min, reached a plateau at 3 h, and declined thereafter, whereas activation of ERK by cisplatin showed a biphasic pattern, indicating the different time frame. Activation of JNK by cisplatin was maximal at 1000 microM, whereas activation of ERK was maximal at 100 microM and was less at higher concentrations, indicating the different dose dependence. Cisplatin-induced JNK activation was neither extracellular and intracellular Ca(2+)- nor protein kinase C-dependent, whereas cisplatin-induced ERK activation was extracellular and intracellular Ca(2+)- dependent and protein kinase C-dependent. A mitogen-activated protein kinase/extracellular signal regulated kinase kinase inhibitor, PD98059, had no effect on the cisplatin induced JNK activity, suggesting an absence of cross-talk between the ERK and JNK cascades. We further examined the effect of each cascade on the viability following cisplatin treatment. Either exogenous expression of dominant negative c Jun or the treatment by PD98059 induced sensitivity to cisplatin in both cells. Our findings suggest that cisplatin-induced DNA damage differentially activates JNK and ERK cascades and that inhibition of either of these cascades sensitizes ovarian cancer cells to cisplatin. PMID- 10531374 TI - Activation of activator protein 1 and stress response kinases in epithelial cells colonized by Helicobacter pylori encoding the cag pathogenicity island. AB - Helicobacter pylori interacts with the apical membrane of the gastric epithelium and induces a number of proinflammatory cytokines/chemokines. The subsequent infiltration of macrophages and granulocytes into the mucosa leads to gastric inflammation accompanied by epithelial degeneration. Gastric diseases, e.g. peptic ulcer or gastric adenocarcinoma, are more common among people infected with H. pylori strains producing VacA (vacuolating cytotoxin A) and possessing a cag (cytotoxin-associated antigen A) pathogenicity island. For the induction of the cytokine/chemokine genes in response to H. pylori, we studied the signaling leading to the nuclear activation of the early response transcription factor activator protein 1 (AP-1). We found that H. pylori strains carrying the pathogenicity island induce activation of AP-1 and nuclear factor kappaB. In contrast to the wild type or an isogenic strain without the vacA gene, isogenic H. pylori strains with mutations in certain cag genes revealed only weak AP-1 and nuclear factor kappaB activation. In respect to the molecular components that direct AP-1 activity, our results indicate a cascade of the cellular stress response kinases c-Jun N-terminal kinase, MAP kinase kinase 4, and p21-activated kinase, and small Rho-GTPases including Rac1 and Cdc42, which contributes to the activation of proinflammatory cytokines/chemokines induced by H. pylori encoding the cag pathogenicity island. PMID- 10531375 TI - Cadmium induces conformational modifications of wild-type p53 and suppresses p53 response to DNA damage in cultured cells. AB - The p53 tumor suppressor protein is a transcription factor that binds DNA in a sequence-specific manner through a protein domain stabilized by the coordination of zinc within a tetrahedral cluster of three cysteine residues and one histidine residue. We show that cadmium, a metal that binds thiols with high affinity and substitutes for zinc in the cysteinyl clusters of many proteins, inhibits the binding of recombinant, purified murine p53 to DNA. In human breast cancer MCF7 cells (expressing wild-type p53), exposure to cadmium (5-40 microM) disrupts native (wild-type) p53 conformation, inhibits DNA binding, and down-regulates transcriptional activation of a reporter gene. Cadmium at 10-30 microM impairs the p53 induction in response to DNA-damaging agents such as actinomycin D, methylmethane sulfonate, and hydrogen peroxide. Exposure to cadmium at 20 microM also suppresses the p53-dependent cell cycle arrest in G(1) and G(2)/M phases induced by gamma-irradiation. These observations indicate that cadmium at subtoxic levels impairs p53 function by inducing conformational changes in the wild-type protein. There is evidence that cadmium is carcinogenic to humans, in particular for lung and prostate, and cadmium is known to accumulate in several organs. This inhibition of p53 function could play a role in cadmium carcinogenicity. PMID- 10531376 TI - A highly conserved signal controls degradation of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase in eukaryotes. AB - Sterol synthesis by the mevalonate pathway is modulated, in part, through feedback-regulated degradation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR). In both mammals and yeast, a non-sterol isoprenoid signal positively regulates the rate of HMGR degradation. To define more precisely the molecule that serves as the source of this signal, we have conducted both pharmacological and genetic manipulations of the mevalonate pathway in yeast. We now demonstrate that farnesyl diphosphate (FPP) is the source of the positive signal for Hmg2p degradation in yeast. This FPP-derived signal does not act by altering the endoplasmic reticulum degradation machinery in general. Rather, the FPP-derived signal specifically modulates Hmg2p stability. In mammalian cells, an FPP-derived molecule also serves as a positive signal for HMGR degradation. Thus, both yeast and mammalian cells employ the same strategy for regulation of HMGR degradation, perhaps by conserved molecular processes. PMID- 10531377 TI - Cloning and sequencing of the coenzyme B(12)-binding domain of isobutyryl-CoA mutase from Streptomyces cinnamonensis, reconstitution of mutase activity, and characterization of the recombinant enzyme produced in Escherichia coli. AB - Isobutyryl-CoA mutase (ICM) catalyzes the reversible, coenzyme B(12)-dependent rearrangement of isobutyryl-CoA to n-butyryl-CoA, which is similar to, but distinct from, that catalyzed by methylmalonyl-CoA mutase. ICM has been detected so far in a variety of aerobic and anaerobic bacteria, where it appears to play a key role in valine and fatty acid catabolism. ICM from Streptomyces cinnamonensis is composed of a large subunit (IcmA) of 62.5 kDa and a small subunit (IcmB) of 14.3 kDa. icmB encodes a protein of 136 residues with high sequence similarity to the cobalamin-binding domains of methylmalonyl-CoA mutase, glutamate mutase, methyleneglutarate mutase, and cobalamin-dependent methionine synthase, including a conserved DXHXXG cobalamin-binding motif. Using IcmA and IcmB produced separately in Escherichia coli, we show that IcmB is necessary and sufficient with IcmA and coenzyme B(12) to afford the active ICM holoenzyme. The large subunit (IcmA) forms a tightly associated homodimer, whereas IcmB alone exists as a monomer. In the absence of coenzyme B(12), the association between IcmA and IcmB is weak. The ICM holoenzyme appears to comprise an alpha(2)beta(2) heterotetramer with up to two molecules of bound coenzyme B(12). The equilibrium constant for the ICM reaction at 30 degrees C is 1.7 in favor of isobutyryl-CoA, and the pH optimum is near 7.4. The K(m) values for isobutyryl-CoA, n-butyryl CoA, and coenzyme B(12) determined with an equimolar ratio of IcmA and IcmB are 57 +/- 13, 54 +/- 12, and 12 +/- 2 microM, respectively. A V(max) of 38 +/- 3 units/mg IcmA and a k(cat) of 39 +/- 3 s(-1) were determined under saturating molar ratios of IcmB to IcmA. PMID- 10531378 TI - Activation of the AT(2) receptor of angiotensin II induces neurite outgrowth and cell migration in microexplant cultures of the cerebellum. AB - Microexplant cultures from three-day-old rats were used to investigate whether angiotensin II (Ang II), through its AT(1) and AT(2) receptors, could be involved in the morphological differentiation of cerebellar cells. Specific activation of the AT(2) receptor during 4-day treatment induced two major morphological changes. The first was characterized by increased elongation of neurites. The second change was cell migration from the edge of the microexplant toward the periphery. Western blot analyses and indirect immunofluorescence studies revealed an increase in the expression of neuron-specific betaIII-tubulin, as well as an increase in expression of the microtubule-associated proteins tau and MAP2. These effects were demonstrated by co-incubation of Ang II with 1 microM DUP 753 (AT(1) receptor antagonist) or with 10 nM CGP 42112 (AT(2) receptor agonist) but abolished when Ang II was co-incubated with 1 microM PD 123319 (AT(2) receptor antagonist), indicating that differentiation occurs through AT(2) receptor activation and that the AT(1) receptor inhibits the AT(2) effect. Taken together, these results demonstrate that Ang II is involved in cerebellum development for both neurite outgrowth and cell migration, two important processes in the organization of the various layers of the cerebellum. PMID- 10531379 TI - Interaction of the metalloprotease disintegrins MDC9 and MDC15 with two SH3 domain-containing proteins, endophilin I and SH3PX1. AB - Metalloprotease disintegrins (a disintegrin and metalloprotease (ADAM) and metalloprotease, disintegrin, cysteine-rich proteins (MDC)) are a family of membrane-anchored glycoproteins that function in diverse biological processes, including fertilization, neurogenesis, myogenesis, and ectodomain processing of cytokines and other proteins. The cytoplasmic domains of ADAMs often include putative signaling motifs, such as proline-rich SH3 ligand domains, suggesting that interactions with cytoplasmic proteins may affect metalloprotease disintegrin function. Here we report that two SH3 domain-containing proteins, endophilin I (SH3GL2, SH3p4) and a novel SH3 domain- and phox homology (PX) domain-containing protein, termed SH3PX1, can interact with the cytoplasmic domains of the metalloprotease disintegrins MDC9 and MDC15. These interactions were initially identified in a yeast two-hybrid screen and then confirmed using bacterial fusion proteins and co-immunoprecipitations from eukaryotic cells expressing both binding partners. SH3PX1 and endophilin I both preferentially bind the precursor but not the processed form of MDC9 and MDC15 in COS-7 cells. Since rat endophilin I is thought to play a role in synaptic vesicle endocytosis and SH3PX1 has sequence similarity to sorting nexins in yeast, we propose that endophilin I and SH3PX1 may have a role in regulating the function of MDC9 and MDC15 by influencing their intracellular processing, transport, or final subcellular localization. PMID- 10531380 TI - Characterization of the rat type III hexokinase gene promoter. A functional octamer 1 motif is critical for basal promoter activity. AB - A 1532-base pair 5'-flanking region of the gene encoding rat type III hexokinase has been cloned and sequenced. The total sequence includes positions -1548 to -17 (A of the translational start ATG as position +1). Using luciferase reporter constructs transfected into PC12 (rat pheochromocytoma) and L2 (rat lung) cells, basal promoter activity has been associated with sequence between -182 and -89. This includes a single transcriptional start site, an adenine at position -134 identified by primer extension. Together with previously cloned cDNA sequence, this accounts for an mRNA of approximately 3.9 kilobases, found by Northern blotting of RNA from rat lung and kidney. Sequence upstream of the transcriptional start site was devoid of canonical TATA and CAAT elements. An octamer 1 (Oct-1) binding site, located between positions -166 and -159 was shown by deletion analysis and site-directed mutation to be critical for promoter activity. Nuclear extracts from PC12 cells contained a protein (or proteins) specifically binding the octamer sequence, and supershift experiments with anti Oct-1 indicated involvement of this ubiquitously expressed transcription factor in the complex. Sequence including the Oct-1 site and immediately adjacent regions was protected from DNase I digestion in footprinting experiments with nuclear extracts from PC12 cells. Reverse transcription polymerase chain reaction indicated that levels of type III hexokinase mRNA in rat tissues increased in the order brain < liver < lung approximately kidney; immunoblotting indicated that type III hexokinase protein in these tissues increased in a similar manner. PMID- 10531381 TI - Ligand-induced ubiquitination of the epidermal growth factor receptor involves the interaction of the c-Cbl RING finger and UbcH7. AB - c-Cbl plays a negative regulatory role in tyrosine kinase signaling by an as yet undefined mechanism. We demonstrate here, using the yeast two-hybrid system and an in vitro binding assay, that the c-Cbl RING finger domain interacts with UbcH7, a ubiquitin-conjugating enzyme (E2). UbcH7 interacted with the wild-type c Cbl RING finger domain but not with a RING finger domain that lacks the amino acids that are deleted in 70Z-Cbl, an oncogenic mutant of c-Cbl. The in vitro interaction was enhanced by sequences on both the N- and C-terminal sides of the RING finger. In vivo and in vitro experiments revealed that c-Cbl and UbcH7 synergistically promote the ligand-induced ubiquitination of the epidermal growth factor receptor (EGFR). In contrast, 70Z-Cbl markedly reduced the ligand-induced, UbcH7-mediated ubiquitination of the EGFR. MG132, a proteasome inhibitor, significantly prolonged the ligand-induced phosphorylation of both the EGFR and c Cbl. Thus, c-Cbl plays an essential role in the ligand-induced ubiquitination of the EGFR by a mechanism that involves an interaction of the RING finger domain with UbcH7. This mechanism participates in the down-regulation of tyrosine kinase receptors and loss of this function, as occurs in the naturally occurring 70Z-Cbl isoform, probably contributes to oncogenic transformation. PMID- 10531382 TI - Determination of the binding site on the extracellular domain of guanylyl cyclase C to heat-stable enterotoxin. AB - Guanylyl cyclase C, one of the family of membrane-bound guanylyl cyclases, consists of an extracellular domain and an intracellular domain, which are connected by a single transmembrane polypeptide. The extracellular domain binds unique small polypeptides with high specificity, which include the endogenous peptide hormones, guanylin and uroguanylin, as well as an exogenous enterotoxigenic peptide, heat-stable enterotoxin, secreted by pathogenic Escherichia coli. Information on this specific binding is propagated into the intracellular domain, followed by the synthesis of cGMP, a second messenger that regulates a variety of intracellular physiological processes. This study reports the design of a photoaffinity labeled analog of heat-stable enterotoxin (biotinyl (AC(5))(2)-[Gly(4), Pap(11)]STp(4-17)), which incorporates a Pap residue (p azidophenylalanine) at position 11 and a biotin moiety at the N terminus, and the use of this analog to determine the ligand-binding region of the extracellular domain of guanylyl cyclase C. The endoproteinase Lys-C digestion of the extracellular domain, which was covalently labeled by this ligand, and mass spectrometric analyses of the digest revealed that the ligand specifically binds to the region (residue 387 to residue 393) of guanylyl cyclase C. This region is localized close to the transmembrane portion of guanylyl cyclase C on the external cellular surface. This result was further confirmed by characterization of site-directed mutants of guanylyl cyclase C in which each amino acid residue was substituted by an Ala residue instead of residues normally located in the region. This experiment provides the first direct demonstration of the ligand binding site of guanylyl cyclase C and will contribute toward an understanding of the receptor recognition of a ligand and the modeling of the interaction of the receptor and its ligand at the molecular level. PMID- 10531383 TI - A conserved inositol phospholipid binding site within the pleckstrin homology domain of the Gab1 docking protein is required for epithelial morphogenesis. AB - Stimulation of the hepatocyte growth factor receptor tyrosine kinase, Met, induces the inherent morphogenic program of epithelial cells. The multisubstrate binding protein Gab1 (Grb2-associated binder-1) is the major phosphorylated protein in epithelial cells following activation of Met. Gab1 contains a pleckstrin homology domain and multiple tyrosine residues that act to couple Met with multiple signaling proteins. Met receptor mutants that are impaired in their association with Gab1 fail to induce a morphogenic program in epithelial cells, which is rescued by overexpression of Gab1. The Gab1 pleckstrin homology domain binds to phosphatidylinositol 3,4, 5-trisphosphate and contains conserved residues, shown from studies of other pleckstrin homology domains to be crucial for phospholipid binding. Mutation of conserved phospholipid binding residues tryptophan 26 and arginine 29, generates Gab1 proteins with decreased phosphatidylinositol 3,4,5-trisphosphate binding, decreased localization at sites of cell-cell contact, and reduced ability to rescue Met-dependent morphogenesis. We conclude that the ability of the Gab1 pleckstrin homology domain to bind phosphatidylinositol 3,4,5-trisphosphate is critical for subcellular localization of Gab1 and for efficient morphogenesis downstream from the Met receptor. PMID- 10531384 TI - Interaction of a kinesin-like protein with calmodulin isoforms from Arabidopsis. AB - In Arabidopsis and other plants there are multiple calmodulin isoforms. However, the role of these isoforms in regulating the activity of target proteins is obscure. Here, we analyzed the interaction between a kinesin-like calmodulin binding motor protein (Reddy, A. S. N., Safadi, F., Narasimhulu, S. B., Golovkin, M., and Hu, X. (1996) J. Biol. Chem. 271, 7052-7060) and three calmodulin isoforms (calmodulin-2, -4, and -6) from Arabidopsis using different approaches. Gel mobility and fluorescence shift assays revealed that the motor binds to all calmodulin isoforms in a calcium-dependent manner. Furthermore, all calmodulin isoforms were able to activate bovine calcium/calmodulin-dependent phosphodiesterase. However, the concentration of calmodulin-2 required for half maximal activation of phosphodiesterase is 2- and 6-fold lower compared with calmodulin-4 and -6, respectively. The dissociation constants of the motor to calmodulin-2, -4, and -6 are 12.8, 27.0, and 27.8 nM, respectively, indicating that calmodulin-2 has 2-fold higher affinity for the motor than calmodulin-4 and 6. Similar results were obtained using another assay that involves the binding of (35)S-labeled calmodulin isoforms to the motor. The binding saturation curves of the motor with calmodulin isoforms have confirmed that calmodulin-2 has 2-fold higher affinity to the motor. However, the affinity of calmodulin-4 and -6 isoforms for the motor was about the same. Based on these studies, we conclude that all calmodulin isoforms bind to the motor protein but with different affinities. PMID- 10531385 TI - Functional consequences of the sustained or transient activation by Bax of the mitochondrial permeability transition pore. AB - The overexpression of Bax kills cells by a mechanism that depends on induction of the mitochondrial permeability transition (MPT) (Pastorino, J. G., Chen, S.-T., Tafani, M., Snyder, J. W., and Farber, J. L. (1998) J. Biol. Chem. 273, 7770 7775). In the present study, purified, recombinant Bax opened the mitochondrial permeability transition pore (PTP). Depending on its concentration, Bax had two distinct effects. At a concentration of 125 nM, Bax caused the release of the intermembranous proteins cytochrome c and adenylate kinase and the release from the matrix of sequestered calcein, effects prevented by the inhibitor of the PTP cyclosporin A (CSA). At this concentration of Bax, there was no detectable mitochondrial swelling or depolarization. These effects of low Bax concentrations are interpreted as the consequence of transient, non-synchronous activation of the PTP followed by a prompt recovery of mitochondrial integrity. By contrast, Bax concentrations between 250 nM and 1 microM caused a sustained opening of the PTP with consequent persistent mitochondrial swelling and deenergization (the MPT). CSA prevented the MPT induced by Bax. Increasing concentrations of calcium caused a greater proportion of the mitochondria to undergo the MPT in the presence of Bax. Importantly, two known mediators of apoptosis, ceramide and GD3 ganglioside, potentiated the induction by Bax of the MPT. The data imply that Bax mediates the opening of the mitochondrial PTP with the resultant release of cytochrome c from the intermembranous space. PMID- 10531386 TI - N(epsilon)-(carboxymethyl)lysine adducts of proteins are ligands for receptor for advanced glycation end products that activate cell signaling pathways and modulate gene expression. AB - Recent studies suggested that interruption of the interaction of advanced glycation end products (AGEs), with the signal-transducing receptor receptor for AGE (RAGE), by administration of the soluble, extracellular ligand-binding domain of RAGE, reversed vascular hyperpermeability and suppressed accelerated atherosclerosis in diabetic rodents. Since the precise molecular target of soluble RAGE in those settings was not elucidated, we tested the hypothesis that predominant specific AGEs within the tissues in disorders such as diabetes and renal failure, N(epsilon)-(carboxymethyl)lysine (CML) adducts, are ligands of RAGE. We demonstrate here that physiologically relevant CML modifications of proteins engage cellular RAGE, thereby activating key cell signaling pathways such as NF-kappaB and modulating gene expression. Thus, CML-RAGE interaction triggers processes intimately linked to accelerated vascular and inflammatory complications that typify disorders in which inflammation is an established component. PMID- 10531387 TI - Molecular mechanisms of cytochrome P-450 induction by xenobiotics: An expanded role for nuclear hormone receptors. PMID- 10531388 TI - Inverse agonism and neutral antagonism at alpha(1a)- and alpha(1b)-adrenergic receptor subtypes. AB - We have characterized the pharmacological antagonism, i.e., neutral antagonism or inverse agonism, displayed by a number of alpha-blockers at two alpha1-adrenergic receptor (AR) subtypes, alpha(1a)- and alpha(1b)-AR. Constitutively activating mutations were introduced into the alpha(1a)-AR at the position homologous to A293 of the alpha(1b)-AR where activating mutations were previously described. Twenty-four alpha-blockers differing in their chemical structures were initially tested for their effect on the agonist-independent inositol phosphate response mediated by the constitutively active A271E and A293E mutants expressed in COS-7 cells. A selected number of drugs also were tested for their effect on the small, but measurable spontaneous activity of the wild-type alpha(1a)- and alpha(1b)-AR expressed in COS-7 cells. The results of our study demonstrate that a large number of structurally different alpha-blockers display profound negative efficacy at both the alpha(1a)- and alpha(1b)-AR subtypes. For other drugs, the negative efficacy varied at the different constitutively active mutants. The most striking difference concerns a group of N-arylpiperazines, including 8-[2-[4-(5 chloro-2-methoxyphenyl)-1-piperazinyl]ethyl]-8-azaspiro [4, 5] decane-7,9-dione (REC 15/3039), REC 15/2739, and REC 15/3011, which are inverse agonists with profound negative efficacy at the wild-type alpha(1b)-AR, but not at the alpha(1a)-AR. PMID- 10531389 TI - Implication of radical oxygen species in ceramide generation, c-Jun N-terminal kinase activation and apoptosis induced by daunorubicin. AB - Anthracyclines such as daunorubicin (DNR) generate radical oxygen species (ROS), which account, at least in part, for their cytotoxic effect. We observed that early ceramide generation (within 6-10 min) through neutral sphingomyelinase stimulation was inhibitable by the antioxidants N-acetylcysteine and pyrrolidine dithiocarbamate, which led to a decrease in apoptosis (>95% decrease in DNA fragmentation after 6 h). Furthermore, we observed that DNR triggers the c-Jun N terminal kinase (JNK) and the transcription factor activated protein-1 through an antioxidant-inhibitable mechanism. Treatment of U937 cells with cell-permeant ceramides induced both an increase in ROS generation and JNK activation, and apoptosis, all of which were antioxidant-sensitive. In conclusion, DNR-triggered apoptosis implicates a ceramide-mediated, ROS-dependent JNK and activated protein 1 activation. PMID- 10531390 TI - Binding pockets of the beta(1)- and beta(2)-adrenergic receptors for subtype selective agonists. AB - We examined the subtype-selective binding site of the beta-adrenergic receptors (betaARs). The beta(1)/beta(2)-chimeric receptors showed the importance of the second and seventh transmembrane domains (TM2 and TM7) of the beta(2)AR for the binding of the beta(2)-selective agonists such as formoterol and procaterol. Alanine-substituted mutants of TM7 of the beta(2)AR showed that Tyr(308,) located at the top of TM7, mainly contributed to beta(2) selectivity. However, Tyr(308) interacted with formoterol and procaterol in two different ways. The results of Ala- and Phe-substituted mutants indicated that the phenyl group of Tyr(308) interacted with the phenyl group in the N-substituent of formoterol (hydrophobic interaction), and the hydroxyl group of Tyr(308) interacted with the protonated amine of procaterol (hydrophilic interaction). In contrast to beta(2)AR, TM2 is a major determinant that beta(1)-selective agonists such as denopamine and T-0509 bound the beta(1)AR with high affinity. Three amino acids (Leu(110), Thr(117), and Val(120)) in TM2 of the beta(1)AR were identified as major determinants for beta(1)-selective binding of these agonists. Three-dimensional models built on the basis of the predicted structure of rhodopsin showed that Tyr(308) of the beta(2)AR covered the binding pocket formed by TM2 and TM7 from the upper side, and Thr(117) of the beta(1)AR located in the middle of the binding pocket to provide a hydrogen bonding for the beta(1)-selective agonists. These data indicate that TM2 and TM7 of the betaAR formed the binding pocket that binds the betaAR subtype-selective agonists with high affinity. PMID- 10531391 TI - Calcineurin enhances acetylcholinesterase mRNA stability during C2-C12 muscle cell differentiation. AB - Treatment of C2-C12 mouse myoblasts with the immunosuppressant drug cyclosporin A (CsA) enhances the increase in acetylcholinesterase (AChE) expression observed during skeletal muscle differentiation. The enhanced AChE expression is due primarily to increased mRNA stability because CsA treatment increases the half life of AChE mRNA, but not the apparent transcriptional rate of the gene. Neither tacrolimus (FK506), an immunosuppressive agent with a distinct structure, nor cyclosporine H, an inactive congener of CsA, alters AChE expression. The enhanced AChE expression is associated with the muscle differentiation process, but cannot be triggered by CsA exposure before differentiation. Myoblasts and myotubes of C2 C12 cells express similar amounts of cyclophilin A and FKBP12, immunophilins known to be intracellular-binding targets for CsA and tacrolimus, respectively. However, cellular levels of calcineurin, a calcium/calmodulin-dependent phosphatase known to be the cellular target of ligand-immunophilin complexes, increase 3-fold during myogenesis. Overexpression of constitutively active calcineurin in differentiating cells reduces AChE mRNA levels and CsA antagonizes such an inhibition. Conversely, overexpression of a dominant negative calcineurin construct increases AChE mRNA levels, which are further enhanced by CsA. Thus, a CsA sensitive, calcineurin mediated pathway appears linked to differentiation induced stabilization of AChE mRNA during myogenesis. PMID- 10531392 TI - Activating mutation of adenylyl cyclase reverses its inhibition by G proteins. AB - We have implemented a yeast genetic selection developed previously by our laboratory to identify mutant mammalian type V adenylyl cyclases insensitive to inhibition by G(ialpha.) One mutation isolated was localized to the first cytoplasmic domain at a Phe residue (position 400), which is conserved in all nine isoforms of membrane-bound mammalian adenylyl cyclase. Biochemical characterization of the F400Y mutant revealed a dramatic conversion of the G(ialpha) response from inhibitory to stimulatory. This mutation results in additional activating effects. The mutant exhibits an enhanced sensitivity toward activation by either G(salpha) or forskolin. Synergism between G(salpha) and forskolin is not observed for the F400Y mutant, presumably because the mutant already is in the sensitized state. Additionally, an enhancement of the basal unstimulated activity was observed. This mutation, which is the first demonstration of an activating point in a mammalian adenylyl cyclase, mimics a sensitized conformation of the wild-type enzyme that underlies the synergism between stimulatory inputs, and additionally, removes the inhibitory regulatory input provided by G(ialpha). Because sensitizing adenylyl cyclase toward its stimulators can have profound biological implications, this raises the possibility that naturally occurring mutations resembling those at the Phe400 residue may be associated with human disease states. PMID- 10531393 TI - Delta-opioid-induced liberation of Gbetagamma mobilizes Ca2+ stores in NG108-15 cells. AB - Activation of delta-opioid receptors in NG108-15 cells releases Ca2+ from an intracellular store through activation of a pertussis toxin-sensitive G protein. We tested the hypothesis that activation of delta-opioid receptors mobilizes inositol 1,4,5-trisphosphate (IP(3))-sensitive Ca2+ stores via liberation of Gbetagamma. Fura-2-based digital imaging was used to study the mechanism of opioid-induced increases in [Ca2+](i) in NG108-15 cells. Exposure to D-Ala(2)-D Leu(5) enkephalin (100 nM) for 90 s induced increases in [Ca2+](i) that were blocked by microinjection of the IP(3) receptor antagonist heparin (pipette concentration = 100 mg/ml) but not by sham injection. Microinjection of a peptide that binds Gbetagamma (QEHA, 1 mM) decreased the D-Ala(2)-D-Leu(5) enkephalin evoked response. Microinjection of an inactive peptide (SKEE, 1 mM) that does not bind to Gbetagamma failed to inhibit the opioid-induced increase in [Ca2+](i). Microinjection of a peptide (QLKK, 15 mM) that binds to free Galpha(q) blocked the increase evoked by 3 nM bradykinin, but microinjection of an inactive peptide (ADRK, 15 mM) did not. Microinjection of QLKK did not significantly affect the opioid-induced increase in [Ca2+](i). Collectively, these data demonstrate that activation of delta-opioid receptors induces the release of Ca2+ from IP(3) sensitive stores in NG108-15 cells through activation of the betagamma subunits of inhibitory G proteins. PMID- 10531394 TI - Study of interaction between agonists and asn293 in helix VI of human beta(2) adrenergic receptor. AB - Previously, we demonstrated the involvement of Asn293 in helix VI of the human beta(2)-adrenergic receptor in stereoselective agonist recognition and activation. In the present study, we have further explored the role of this residue by synthesizing derivatives of isoproterenol and clenbuterol, two beta adrenergic receptor agonists. We analyzed their efficacy and affinity on the wild type and a mutant receptor (Asn293Leu). Each compound had similar efficacy (tau values) on both the wild-type and mutant receptor, although tau values varied considerably among the eight compounds studied. It appeared that one derivative of isoproterenol, but not of clenbuterol, showed a gain in affinity from the wild type to the mutant receptor. This derivative had a methyl substituent instead of the usual beta-OH group in the aliphatic side chain of isoproterenol, compatible with the more lipophilic nature of the leucine side chain. Such a "gain of function" approach through a combination of synthetic chemistry with molecular biology, may be useful to enhance our insight into the precise atomic events that govern ligand-receptor interactions. PMID- 10531395 TI - Identification and functional analysis of novel cAMP response element binding protein splice variants lacking the basic/leucine zipper domain. AB - Two novel cAMP response element binding protein (CREB) splice variants were found by reverse transcription-polymerase chain reaction cloning by using mouse brain RNA as a template. One splice variant, named Delta-14, lacks 14 nucleotides at the beginning of exon 9 of the CREBDelta isoform. The other, named Delta-35, lacks 35 nucleotides at the beginning of exon 8 of CREBDelta. These nucleotide deletions cause frame shifts for codon usage, producing proteins which conserve the major phosphorylation site (Ser(133)) but lack the basic/leucine zipper domain, which is essential for binding to DNA and to other transcription factors. Both variants are widely expressed in peripheral tissues, but are enriched in brain, thymus, and testis. CREBDelta-14 and Delta-35 variant proteins were expressed by using an in vitro translation system and by transfecting into human embryonic kidney 293 cells. Both variants were detected by a CREB antibody that recognizes the CREBDelta amino terminus, but not by an antibody which recognizes the CREBDelta carboxy terminus, as would be predicted based on the frame shift. Activation of the cAMP pathway increased phospho-CREB immunoreactivity, indicating that these variants are substrates of cAMP-dependent protein kinase. In addition, immunocytochemical analysis demonstrated that CREBDelta-14 and Delta 35 are primarily cytosolic, whereas CREBalpha is predominantly in the nucleus. Finally, expression of CREBDelta-14 or Delta-35 decreased cAMP responsive element chloramphenicol acetyltransferase reporter activity, demonstrating that both can function as repressors of endogenous CREB. PMID- 10531396 TI - Neutrophil inhibitory factor abrogates neutrophil adhesion by blockade of CD11a and CD11b beta(2) integrins. AB - We studied the basis of inhibition of polymorphonuclear leukocyte (PMN) adhesion induced by neutrophil inhibitory factor (NIF), a 41-kDa CD11/CD18 beta(2) integrin-binding protein isolated from the canine hookworm (Ancylostoma caninum). NIF blocked PMN adhesion in a concentration-dependent manner with complete blockade occurring at approximately 10 nM NIF. Because CD11a and CD11b beta(2) integrins are functionally active on stimulated PMNs, and yet NIF is postulated to inhibit only CD11b integrin by binding to its I domain, we evaluated the contributions of CD11a and CD11b beta(2) integrins in the mechanism of inhibition of PMN adhesion to endothelial cells. We observed an additive inhibitory effect (>90% inhibition) of PMN adhesion to endothelial cells when NIF was used in combination with anti-CD11b monoclonal antibodies, which alone at saturating concentrations reduced PMN adhesion by only 50%. NIF also prevented aggregation of phorbol ester-stimulated JY lymphoblastoid cells that expressed only the functionally active CD11a, suggesting that NIF also can inhibit CD11a-dependent response. We transduced the NIF cDNA into human dermal microvessel endothelial cells in which NIF synthesis and release prevented PMN adhesion to the transduced human dermal microvessel endothelial cells. These data indicated that the potent antiadhesive effect of NIF may be the result of inhibition of CD11a and CD11b beta(2) integrins on PMNs. Moreover, the strategy of NIF release from transduced endothelial cells suggests the feasibility of blocking the CD11a- and CD11b beta(2) integrin-dependent PMN adhesion and PMN migration responses specifically at sites of endothelial cell activation. PMID- 10531397 TI - m2-toxin: A selective ligand for M2 muscarinic receptors. AB - Selective ligands are needed for distinguishing the functional roles of M2 receptors in tissues containing several muscarinic receptor subtypes. Because the venom of the green mamba Dendroaspis angusticeps contains the most specific antagonists known for M1 and M4 receptors (m1-toxin and m4-toxin), it was screened for toxins that inhibit the binding of [(3)H]N-methylscopolamine ([(3)H]NMS) to cloned M2 receptors. Desalted venom had as much anti-M2 as anti-M4 activity. The most active anti-M2 toxin in the venom was isolated by gel filtration, cation-exchange chromatography, and reversed-phase HPLC, and called m2-toxin. Spectrometry yielded a mass of 7095 Da, and N-terminal sequencing of 53 amino acids showed RICHSQMSSQPPTTTFCRVNSCYRRTLRDPHDPRGT-IIVRGCGCPRMKPGTKL. This sequence is more homologous to antinicotinic than antimuscarinic toxins, but it lacks three almost invariant residues of antinicotinic toxins required for their activity. m2-Toxin fully blocked the binding of [(3)H]NMS and [(3)H]oxotremorine M to M2 receptors with Hill coefficients near 1, and blocked 77% of the binding sites for 0.1 nM [(3)H]NMS in the rat brainstem (K(i) = 11 nM). Concentrations that fully blocked cloned M2 receptors had no effect on M4 receptors, but slightly increased [(3)H]NMS binding to M1 receptors, an allosteric effect. In accord with these results, light microscopic autoradiography of the rat brain showed that m2-toxin decreased [(3)H]NMS binding in regions rich in M2 receptors and increased binding in regions rich in M1 receptors. Thus m2-toxin is a novel M2-selective, short-chain neurotoxin that may prove useful for binding and functional studies. PMID- 10531398 TI - Tumor suppressor p53 but not cGMP mediates NO-induced expression of p21(Waf1/Cip1/Sdi1) in vascular smooth muscle cells. AB - Cyclin-dependent kinase inhibitor p21(Waf1/Cip1/Sdi1) has been suggested to be involved in the antiproliferative effect of nitric oxide (NO) in vascular smooth muscle cells (VSMCs). To elucidate the mechanism underlying NO-induced p21 expression, we investigated the roles of tumor suppressor p53 and the guanylate cyclase-cGMP pathway. The induction of p21 by the NO donor S-nitroso-N acetylpenicillamine (SNAP) seemed to be due to transactivation because SNAP elevated the activity of p21 promoter but did not stabilize p21 mRNA and protein. Because SNAP did not stimulate the deletion mutant of p21 promoter that lacked p53 binding sites, we tested the involvement of p53. The expression level of p53 was down-regulated after mitogenic stimulation, whereas it was sustained in the presence of SNAP. SNAP markedly stimulated DNA binding activity of p53. Furthermore, SNAP failed to induce p21 in VSMCs obtained from p53-knock out mice and in A431 cells that contained mutated p53. The antiproliferative effect of SNAP also was attenuated in these cells. NO stimulates guanylate cyclase and its product cGMP has been shown to inhibit VSMC proliferation. However, 1H [1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, a guanylate cyclase inhibitor, did not prevent SNAP-induced p21 expression. 8-Bromo-cGMP, 3-isobutyl-1-methylxanthine, and their combination did not induce p21. Although 8-bromo-cGMP had a small antiproliferative effect, the elevation of cGMP concentration induced by SNAP was little throughout the G(1) phase. The antiproliferative effect of SNAP was not attenuated by Rp-8-bromoguanosine-3',5'-monophosphorothioate, an inhibitor of cGMP-dependent protein kinase. These results suggested that NO induces p21 through a p53-dependent but cGMP-independent pathway. PMID- 10531399 TI - A role of p75 in NGF-mediated down-regulation of the A(2A) adenosine receptors in PC12 cells. AB - Nerve growth factor (NGF) induces differentiation of the rat pheochromocytoma clone (PC12) by activating the high affinity receptor, p140(trkA), linked to mitogen-activated protein kinase. While the physiological role of the low affinity NGF receptor (p75) has not been clearly defined, this receptor promotes activation of nuclear factor (NF) kappaB in Schwann cells. PC12 cells express the A(2A) adenosine receptor (AR), whose expression is significantly decreased by NGF treatment. In this study, we determined whether TrkA or p75 is involved in NGF mediated regulation of A(2A)AR expression. NGF treatment decreased A(2A)AR in a time-dependent manner, with maximal effects observed by 1 day, and continued down regulation of the receptor for up to 3 days in the presence of NGF. The decrease in A(2A)AR was associated with a more delayed decrease in the steady-state levels of the A(2A)AR mRNA. Down-regulation of the A(2A)AR at 1 day was mimicked by activators of NFkappaB, such as H(2)O(2), and ceramide, and was attenuated by the inhibitor pyrrolidine dithiocarbamate or following transient transfection of PC12 cells with a dominant negative IkappaBalpha mutant. Moreover, NGF stimulated nuclear accumulation of p65 subunits of NFkappaB (but not p50 subunits) in PC12 cells, as determined by electrophoretic mobility shift assays and by Western blotting. In contrast, inhibition of TrkA by AG879 or of TrkA-dependent mitogen activated protein kinase mitogen-activated protein kinase kinase with PD98059 blocked PC12 cell differentiation without affecting A(2A)AR down-regulation, suggesting dissociation between these two phenomena. Taken together, these data provide strong support for the involvement of the p75/NFkappaB pathway in NGF mediated down-regulation of A(2A)AR in PC12 cells. PMID- 10531400 TI - Coexpression with the inward rectifier K(+) channel Kir6.1 increases the affinity of the vascular sulfonylurea receptor SUR2B for glibenclamide. AB - ATP-sensitive K(+) channels are closed by the hypoglycemic sulfonylureas like glibenclamide (GBC) and activated by a class of vasorelaxant compounds, the K(+) channel openers. These channels are octamers of Kir6.x and sulfonylurea receptor (SUR) subunits with 4:4 stoichiometry. The properties of the opener-sensitive K(+) channel in the vasculature are well matched by the SUR2B/Kir6.1 channel; however, the GBC sensitivity of the recombinant channel is unknown. In binding experiments we have determined the affinity of GBC for SUR2B and the SUR2B/Kir6.1 channel and compared the results with the channel blocking potency of GBC. All experiments were performed in whole transfected human embryonic kidney cells at 37 degrees C. The equilibrium dissociation constants (K(D)) of GBC binding to SUR2B and to the SUR2B/Kir6.1 complex were determined to be 32 and 6 nM, respectively; the K(D) value of the opener P1075 (N-cyano-N'-(1, 1 dimethylpropyl)-N"-3-pyridylguanidine) ( approximately 5 nM) was, however, not affected by cotransfection. In whole cell voltage-clamp experiments, GBC inhibited the SUR2B/Kir6.1 channel with IC(50) approximately 43 nM. The data show that, in the intact cell: 1) SUR2B, previously considered to be a low-affinity SUR, has a rather high affinity for GBC; 2) coexpression with the inward rectifier Kir6.1 increases the affinity of SUR2B for GBC; 3) the recombinant channel exhibits the same GBC affinity as the opener-sensitive K(+) channel in vascular tissue; and 4) the K(D) value of GBC binding to the octameric channel is 7 times lower than the IC(50) value for channel inhibition. The latter finding suggests that occupation of all four GBC sites per channel is required for channel closure. PMID- 10531401 TI - Using a radioalloster to test predictions of the cooperativity model for gallamine binding to the allosteric site of muscarinic acetylcholine M(2) receptors. AB - The muscarinic M(2) receptor contains an orthosteric and an allosteric site. Binding of an allosteric agent may induce a shift alpha of the equilibrium dissociation constant K(D) of a radioligand for the orthosteric site. According to the cooperativity model, the K(A) of alloster binding is expected to be shifted to an identical extent depending on whether the orthosteric site is occupied by the orthoster or not. Here, the novel radioalloster [(3)H]dimethyl W84 (N,N'-bis[3-(1,3-dihydro-1, 3-dioxo-4-methyl-2H-isoindol-2-yl)propyl] N,N,N',N'-tetramethyl-1, 6-hexanediaminium diiodide) was applied to directly measure the K(A) shift induced for the prototype allosteric modulator gallamine by binding of N-methylscopolamine (NMS) to the orthosteric site of porcine heart M(2) receptors (4 mM Na(2)HPO(4), 1 mM KH(2)PO(4), pH 7.4; 23 degrees C; data are means +/- S.E.). First, in the common way, the concentration-dependent inhibition by gallamine of [(3)H]NMS equilibrium binding was measured and analyzed using the cooperativity model, which yielded for the affinity of gallamine binding at free receptors a pK(A)= 8.35 +/- 0.09 and a cooperativity factor alpha = 46 (n = 5). The dissociation constant for gallamine binding at NMS-occupied receptors was predicted as p(alpha. K(A)) = 6.69. Labeling of the allosteric site by [(3)H]dimethyl-W84 allowed the measure of competitive displacement curves for gallamine. The K(i) for gallamine at free receptors amounted to pK(i,-NMS) = 8.27 +/- 0.39 (n = 5), which is in line with the prediction of the cooperativtiy model. In the presence of 1 microM NMS, to occupy the orthosteric site, gallamine displaced [(3)H]dimethyl-W84 with pK(i, +NMS) = 6.60 +/- 0.19 (n = 3). Thus, the NMS-induced pK(i) shift amounted to 47, which matches the predicted value of alpha = 46. These results validate the cooperativity model. PMID- 10531402 TI - Modulation of cisplatinum cytotoxicity by p53: effect of p53-mediated apoptosis and DNA repair. AB - A stable transfectant (S2SN7) of p53-null SaOS-2 (human osteosarcoma) cells expressing wild-type p53 under the tight control of a tetracycline-responsive promoter was used to study the functional roles of p53 in cellular response to cisplatinum (CP). When cells were grown in media containing normal concentrations (10%) of serum, induction of p53 by tetracycline withdrawal resulted in an 8-fold decrease in sensitivity to CP. In contrast, when cells were grown in lower serum (1%) media, induction of p53 led to a 10-fold increase in sensitivity to CP. The p53-mediated sensitivity to CP under lower serum conditions was attributed, at least in part, to increased susceptibility of p53-mediated apoptosis. Under lower serum (0.1-1%) but not normal serum conditions, p53 induction correlated with selective down-regulation of bcl-2, an inhibitor of apoptosis. In addition, a host-cell reactivation assay showed that induction of p53 caused a significant increase in repair of CP-induced DNA damage under normal serum but not low serum conditions. These data suggest that growth conditions may modulate and possibly reverse p53-mediated CP sensitivity by altering p53-mediated gene regulation and, as a result, susceptibility to apoptosis. They also suggest that a combined effect of p53-mediated apoptosis and DNA repair may be the ultimate determinant in p53-mediated cellular resistance or sensitivity to chemotherapeutic drugs. PMID- 10531403 TI - Contribution of individual subunits to the multimeric P2X(2) receptor: estimates based on methanethiosulfonate block at T336C. AB - P2X receptors are membrane proteins that incorporate a cation-selective ion channel that can be opened by the binding of extracellular ATP. They associate as hetero- and homo-multimers of currently unknown stoichiometry. In this study, we have used Xenopus laevis oocytes to express rat P2X(2) receptor subunits, which carry a cysteine mutation at position 336. ATP-induced currents at this mutant receptor subunit were blocked by more than 90% when exposed to [2 (trimethylammonium) ethyl] methanethiosulfonate (MTSET), whereas currents from wild-type subunits were not affected. To compare mutant and wild-type channel expression, we introduced an epitope in their extracellular domains and found for both channels a similar linear relationship between antibody binding and currents induced by ATP. To study the contribution of the individual subunits to the block by MTSET, we coinjected different mixtures of wild-type and mutant-encoding mRNAs. We found that the inhibition by MTSET depended linearly on the proportion of mutant subunits, which was clearly contrary to the hypothesis that a single mutant subunit could act in a dominant fashion. Subsequent concatenation of wild type and mutant-encoding cDNAs resulted in an inhibition by MTSET that also depended linearly on the number of mutant subunits and was independent of the position of the mutant subunit, as long as only two or three P2X(2) subunits were joined. With four or six subunits joined, however, the inhibition by MTSET became strongly position-dependent. The present results show that a "per-subunit" channel block causes the blocking effects of MTSET and they suggest that not four but maximally three subunits actively participate in the channel formation. PMID- 10531404 TI - Lead inhibition of DNA-binding mechanism of Cys(2)His(2) zinc finger proteins. AB - The association of lead with chromatin in cells suggests that deleterious metal effects may in part be mediated through alterations in gene function. To elucidate if and how lead may alter DNA binding of cysteine-rich zinc finger proteins, lead ions were analyzed for their ability to alter the DNA binding mechanism of the Cys(2)His(2) zinc finger protein transcription factor IIIA (TFIIIA). As assayed by DNase I protection, the interaction of TFIIIA with the 50 bp internal control region of the 5S ribosomal gene was partially inhibited by 5 microM lead ions and completely inhibited by 10 to 20 microM lead ions. Preincubation of free TFIIIA with lead resulted in DNA-binding inhibition, whereas preincubation of a TFIIIA/5S RNA complex with lead did not result in DNA binding inhibition. Because 5S RNA binds TFIIIA zinc fingers, this result is consistent with an inhibition mechanism via lead binding to zinc fingers. The complete loss of DNase I protection on the 5S gene indicates the mechanism of inhibition minimally involves the N-terminal fingers of TFIIIA. Inhibition was not readily reversible and occurred in the presence of an excess of beta mercaptoethanol. Inhibition kinetics were fast, progressing to completion in approximately 5 min. Millimolar concentrations of sulfhydryl-specific arsenic ions were not inhibitory for TFIIIA binding. Micromolar concentrations of lead inhibited DNA binding by Sp1, another Cys(2)His(2) finger protein, but not by the nonfinger protein AP2. Inhibition of Cys(2)His(2) zinc finger transcription factors by lead ions at concentrations near those known to have deleterious physiological effects points to new molecular mechanisms for lead toxicity in promoting disease. PMID- 10531406 TI - Domain exchangeability between the multidrug transporter (MDR1) and phosphatidylcholine flippase (MDR2). AB - Multidrug resistance (MDR) mediated by P-glycoprotein (MDR1) is clinically significant. Understanding how MDR1 substrate specificity is determined will help to overcome MDR to improve cancer treatment. One potential approach to achieve this goal is to study chimeras of MDR1 and its homolog MDR2 (also called MDR3), which has been identified as a phosphatidylcholine flippase. With an approach involving exchanging homologous segments of MDR1 and MDR2 and site-directed mutagenesis, we previously demonstrated MDR1 residues Q330, V331, and L332 in transmembrane domain 6 (TM6) are essential for multidrug transport activity; substituting these residues allows the N-terminal transmembrane region of MDR2 to support MDR1 activity. To further determine the exchangeability between MDR1 and MDR2, we constructed additional MDR1/MDR2 chimeras. We found that the N-terminal half of MDR1 and MDR2 was mostly exchangeable except for a few residues in TM6. However, this degree of exchangeability was not found in the C-terminal half of MDR1 and MDR2. In addition, with substitution of MDR1 residues 318-332 (TM6) and 937-994 (TM11-12), MDR2 had relatively normal affinity for MDR1 substrates, but it did not have multidrug transporter activity. These results suggest that the inability of MDR2 to transport most MDR1 drugs efficiently may be due to failure of those drugs to stimulate ATPase and activate transport as well as to decreased drug binding. PMID- 10531405 TI - Inverse agonist properties of dopaminergic antagonists at the D(1A) dopamine receptor: uncoupling of the D(1A) dopamine receptor from G(s) protein. AB - The interaction of dopaminergic antagonists with the D(1A) dopamine receptor was assessed in PC2 cells that transiently express this receptor. The maximal binding and dissociation constants for the D(1A) dopamine receptor, using the ligand [(125)I]SCH23982 were 0.38 +/- 0.09 nM and 1 to 4 pmol/mg, respectively, when assessed 48 h after transfection with cDNA encoding the rat D(1A) receptor. Basal adenylyl cyclase activity increased 50 to 60% in membranes of transfected PC2 cells compared with control membranes. The dopaminergic antagonists clozapine, cis-flupenthixol, (+)-butaclamol, haloperidol, chlorpromazine, and fluphenazine inhibited constitutive adenylyl cyclase activity in membranes of cells expressing the D(1A) receptor. SCH23390, a selective D(1) dopamine receptor antagonist, and (-)-butaclamol did not alter basal cyclase activity, whereas dopamine increased enzyme activity in membranes expressing the D(1A) dopamine receptor. The coupling of D(1A) receptors with G(s) proteins was examined by immunoprecipitation of membrane G(salpha) followed by immunoblotting with a D(1A) dopamine receptor monoclonal antibody. Clozapine, cis-flupenthixol, (+)-butaclamol, haloperidol, and fluphenazine but not SCH23390 or (-)-butaclamol decreased D(1A) receptor G(salpha) coupling by 70 to 80%, and SCH23390 was able to prevent the receptor G(salpha) uncoupling induced by haloperidol or clozapine. These results indicate that some dopaminergic antagonists suppress basal signal transduction and behave as inverse agonists at the D(1A) dopamine receptor. This action of the dopamine receptor antagonists may contribute to their antidopaminergic properties that seem to underlie their clinical actions as antipsychotic drugs. PMID- 10531407 TI - G(i) activator region of alpha(2A)-adrenergic receptors: distinct basic residues mediate G(i) versus G(s) activation. AB - The structural determinants of G protein coupling versus activation by G protein coupled receptors are not well understood. We examine the role of two distinct basic regions in the carboxyl terminal portion of the third intracellular loop of the alpha(2A)-adrenergic receptor to dissect these aspects of function. Changing three arginines to alanines by mutagenesis and stable expression in Chinese hamster ovary-K1 cells impaired the alpha(2)-adrenergic receptor G(s)-mediated stimulation of cyclic AMP (cAMP) accumulation, whereas G(i)-mediated inhibition was normal. When two (B2) or three (B3) basic residues closer to transmembrane span 6 were mutated to alanine, normal ligand binding was observed, but G(i) mediated inhibition of cAMP accumulation showed 20-fold and 50-fold decreases in agonist potency for the B2 and B3 mutants, respectively. Surprisingly, a normal G(s) response was seen for the B2 mutant, and the B3 mutant showed only a 6-fold decrease in agonist potency. Mutation of both the three alanines and B3 residues to alanines showed a 200-fold decrease in agonist potency for G(i)-mediated inhibition of cAMP accumulation, whereas the G(s) response was nearly completely eliminated. The three basic residues (which include the BB of the BBXXF motif) play a role as G(i) activators rather than in receptor-G protein coupling, because high-affinity agonist binding is intact. Thus, we have identified three basic residues required for activation of G(i) but not required for receptor-G protein coupling. Also, distinct basic residues are required for optimal G(i) and G(s) responses, defining a microspecificity determinant within the carboxyl terminal portion of the third intracellular loop of the alpha(2a) adrenergic receptor. PMID- 10531409 TI - Human dopamine D(3) receptors mediate mitogen-activated protein kinase activation via a phosphatidylinositol 3-kinase and an atypical protein kinase C-dependent mechanism. AB - The mitogen-activated protein kinase (MAPK) cascade is stimulated by both receptor tyrosine kinases and G protein-coupled receptors. We show that recombinant human dopamine D(3) receptors expressed in Chinese hamster ovary cells transiently activate MAPK via pertussis toxin-sensitive Gi and/or Go proteins. The involvement of D(3) receptors was confirmed by use of the D(3) agonists PD 128,907 and (+)-7-hydroxy-2-dipropylaminotetralin, which mimicked the response to dopamine (DA). Furthermore, haloperidol and the selective D(3) receptor antagonists S 14297 and GR 218,231 attenuated DA-induced MAPK activation; however, when tested alone, S 14297 weakly stimulated MAPK activity, suggesting partial agonist activity. The transduction mechanisms by which hD(3) receptors activate MAPK were explored with specific kinase inhibitors. Genistein and lavendustin A, inhibitors of tyrosine kinase activity, did not reduce DA induced MAPK activation. In contrast, PD 98059, an inhibitor of MAPK kinase, and Ro 31-8220 and Go 6983, inhibitors of protein kinase C (PKC), blocked DA-induced MAPK activation. However, MAPK activation was insensitive to PKC down-regulation by phorbol esters, indicating the involvement of an "atypical" PKC. Furthermore, MAPK activation involved phosphatidylinositol 3-kinase inasmuch as its inhibition by LY 294002 and wortmannin reduced DA-induced MAPK activation. In conclusion, this study demonstrates that stimulation of hD(3) receptors activates MAPK. This action is mediated via an atypical isoform of PKC, possibly involving cross-talk with products of phosphatidylinositol 3-kinase activation. PMID- 10531408 TI - Tight association of the human Mel(1a)-melatonin receptor and G(i): precoupling and constitutive activity. AB - If stably expressed in human embryonic kidney (HEK)293 cells, the human Mel(1a) melatonin receptor activates G(i)-dependent, pertussis toxin-sensitive signaling pathways, i.e., inhibition of adenylyl cyclase and stimulation of phospholipase Cbeta; the latter on condition that G(q) is coactivated. The antagonist luzindole blocks the effects of melatonin and acts as an inverse agonist at the Mel(1a) receptor in both intact cells and isolated membranes. This suggests that the Mel(1a) receptor is endowed with constitutive activity, a finding confirmed on reconstitution of the Mel(1a) receptor with G(i). Because the receptor density is in the physiological range, constitutive activity is not an artifact arising from overexpression of the receptor. In addition, the following findings indicate that the Mel(1a) receptor forms a very tight complex with G(i) which can be observed both in the presence and absence of an agonist. 1) In intact cells and in membranes, high-affinity agonist binding is resistant to the destabilizing effect of guanine nucleotides. 2) The ability to bind an agonist with high affinity is preserved even after exposure of the cells to pertussis toxin, because a fraction of G(i) is inaccessible to the toxin in cells expressing Mel(1a) receptors (but not the A(1)-adenosine receptor, another G(i)-coupled receptor). 3) An antiserum directed against the Mel(1a) receptor coprecipitates G(i) even in the absence of an agonist. We therefore conclude that the Mel(1a) receptor is tightly precoupled and that its constitutive activity may play a role in pacing the biological clock, an action known to involve the melatonin receptors in the suprachiasmatic nucleus. PMID- 10531411 TI - Slow sequential conformational changes in Escherichia coli ribosomes induced by lincomycin: kinetic evidence. AB - In a cell-free system derived from Escherichia coli, lincomycin produces biphasic logarithmic time plots for inhibition of peptide-bond formation when puromycin is used as an acceptor substrate and AcPhe-tRNA as a donor substrate. In a previous study, initial slope analysis of the logarithmic time plots revealed that the encounter complex CI between the initiator ribosomal complex (C) and lincomycin (I) undergoes a slow isomerization to C*I. During this change, the bound AcPhe tRNA and lincomycin are rearranged to also accommodate puromycin, and this may account for the mixed noncompetitive inhibition (K(i)* = 70 microM) established at higher concentrations of the drug. The above-mentioned effect was further investigated by analyzing the late phase of the logarithmic time plots. It was found that C*I complex reacts with a second molecule of I, giving C*I(2) complex. However, the logarithmic time plots remain biphasic even at high concentrations of lincomycin, making possible the identification of another inhibition constant K(i)*', which is equal to 18 microM. The simplest explanation of this finding is to assume the existence of a second isomerization step C*I(2) <--> C*I(2'), slowly equilibrated. The determination of K(i)*' enables us to calculate the isomerization constant (K(isom) = 2.9) with the formula K(i)*' = K(i)*/(1 + K(isom)). Our results suggest that whenever a fast and reversible interaction of lincomycin with the elongating ribosomal complex C occurs, the latter undergoes a slow isomerization, which may be the result of conformational changes induced by the drug. PMID- 10531410 TI - Alanine-scanning mutagenesis of transmembrane domain 6 of the M(1) muscarinic acetylcholine receptor suggests that Tyr381 plays key roles in receptor function. AB - Transmembrane domain 6 of the muscarinic acetylcholine (ACh) receptors is important in ligand binding and in the conformational transitions of the receptor but the roles of individual residues are poorly understood. We have carried out a systematic alanine-scanning mutagenesis study on residues Tyr381 to Val387 within the binding domain of the M(1) muscarinic ACh receptor. The seven mutations were then analyzed to define the effects on receptor expression, agonist and antagonist binding, and signaling efficacy. Tyr381Ala produced a 40-fold reduction in ACh affinity and a 50-fold reduction in ACh-signaling efficacy. Leu386Ala had similar but smaller effects. Asn382Ala caused the largest inhibition of antagonist binding. The roles of the hydroxyl group and benzene ring of Tyr381 were probed further by comparative analysis of the Tyr381Phe and Tyr381Ala mutants using three series of ligands: ACh analogs, azanorbornane- and quinuclidine-based ligands, and atropine analogs. These data suggested that the hydroxyl group of Tyr381 is primarily involved in forming hydrogen bond interactions with the oxygen atoms present in the side chain of ACh. We propose that this interaction is established in the ground state and preserved in the activated state of the receptor. In contrast, the Tyr381 benzene ring may form a cation-pi interaction with the positively charged head group of ACh that contributes to the activated state of the receptor but not the ground state. However, the hydroxyl group and benzene ring of Tyr381 both participate in interactions with azanorbornane- and quinuclidine-based ligands and atropine analogs in the ground state as well as the activated state of the receptor. PMID- 10531412 TI - Ion dependence of carrier-mediated release in dopamine or norepinephrine transporter-transfected cells questions the hypothesis of facilitated exchange diffusion. AB - The mechanism of release mediated by the human dopamine and norepinephrine transporter (DAT and NET, respectively) was studied by a superfusion technique in human embryonic kidney 293 cells stably transfected with the respective transporter cDNA and loaded with the metabolically inert substrate [(3)H]1-methyl 4-phenylpyridinium. Release was induced by amphetamine, dopamine, and norepinephrine or by lowering the sodium or chloride concentration in the superfusion buffer (iso-osmotic replacement by lithium and isethionate, respectively). Efflux of [(3)H]1-methyl-4-phenylpyridinium was analyzed at 30-s time resolution. In both transporters, release induced by the substrates amphetamine, dopamine, and norepinephrine followed the same time course as release induced by the removal of chloride and was faster than that caused by the removal of sodium. In the presence of low sodium (DAT: 10 mM; NET: 5 mM) none of the substrates was able to induce release from either type of cell, but adding back sodium to control conditions promptly restored the releasing action. In the presence of low chloride (DAT: 3 mM; NET: 2 mM), however, amphetamine as well as the catecholamines stimulated release from both types of cell. In contrast with the ion dependence of release observed in superfusion experiments, uptake initial rates of substrates at concentrations used in release experiments were the same or even higher at low sodium than at low chloride. The results indicate a decisive role of extracellular sodium for carrier-mediated release unrelated to the sodium-dependent uptake of the releasing substrate, and suggest a release mechanism different from simple exchange diffusion considering only the amines as substrates. PMID- 10531413 TI - Activation of leukotriene synthesis in human neutrophils by exogenous arachidonic acid: inhibition by adenosine A(2a) receptor agonists and crucial role of autocrine activation by leukotriene B(4). AB - We report here that the apparent inability of isolated human polymorphonuclear leukocytes (PMNs) to efficiently transform arachidonic acid (AA) is the consequence of A(2a) receptor engagement by endogenous adenosine accumulating in incubation media. Indeed, when adenosine is eliminated from PMN suspensions by the addition of adenosine deaminase, or when cells are incubated with adenosine A(2a) receptor antagonists, important quantities (40-80 pmol/10(6) cells) of 5 lipoxygenase products are synthesized by PMN incubated with 1 to 5 microM exogenous AA. The selective A(2a) receptor agonist CGS21680 was a very potent inhibitor of the AA-induced leukotriene (LT) synthesis, showing an IC(50) of approximately 1 nM. The mechanism of AA-induced stimulation of LT synthesis observed in the absence of extracellular adenosine was investigated. In adenosine deaminase-treated PMN, exogenous AA induced Ca(2+) mobilization and the translocation of 5-lipoxygenase to nuclear structures. A time lag of 20 to 60 s (variable between PMN preparations) was observed consistently between the addition of AA and the elevation of intracellular Ca(2+) concentration (and LT synthesis), indicating that AA itself did not trigger the Ca(2+) mobilization in PMN. This AA-induced Ca(2+) mobilization, as well as the corresponding 5 lipoxygenase translocation and stimulation of LT synthesis, was blocked efficiently by the LT synthesis inhibitor MK0591, the LTB(4) receptor antagonists CP105696 and LY223982, and the LTA(4) hydrolase inhibitor SC57461A. These data demonstrate that AA is a highly potent and effective activator of LT synthesis and acts through a mechanism that requires an autocrine stimulatory loop by LTB(4). PMID- 10531414 TI - Mechanisms of acquired resistance to thymidylate synthase inhibitors: the role of enzyme stability. AB - Inhibitors of the enzyme thymidylate synthase (TS), such as the fluoropyrimidines 5-fluorouracil and 5'-fluoro-2'-deoxyuridine (FdUrd) or the antifolates AG337, ZD1694, and BW1843U89, are widely used in the chemotherapy of cancer, particularly cancer of the colon and rectum. Numerous studies have shown that TS gene amplification, leading to mRNA and enzyme overproduction, is a major mechanism of resistance to these inhibitors. In the present work, we have isolated and characterized FdUrd-resistant derivatives of several human colon tumor cell lines. Although gene amplification was commonly observed, the increases in mRNA and enzyme were strikingly discordant. In one drug-resistant line, a deficiency of enzyme relative to mRNA was shown to be caused by expression of a metabolically unstable TS molecule. The reduced half-life of TS in this line was caused by a Pro-to-Leu substitution at residue 303 of the TS polypeptide. The mutant enzyme conferred resistance to FdUrd as well as antifolates in transfected cells. In another FdUrd-resistant line, which had an excess of enzyme relative to mRNA, the TS molecule was more stable than in the parent line. However, no amino acid substitutions were detected in the TS polypeptide from this line, which suggests that the stabilization must be caused by changes in one or more cellular factors that regulate TS degradation. The results indicate that changes in the stability of the TS polypeptide accompany, and even contribute to, acquired resistance to TS inhibitors in colon tumor cells. PMID- 10531415 TI - N-linked glycosylation is required for plasma membrane localization of D5, but not D1, dopamine receptors in transfected mammalian cells. AB - We have analyzed the role of N-linked glycosylation in functional cell surface expression of the D1 and D5 dopamine receptor subtypes. Treatment of transfected HEK 293 cells with tunicamycin, an inhibitor of N-linked oligosaccharide addition, was found to prevent localization of D5 receptors in the plasma membrane. In contrast, tunicamycin treatment had no effect on the plasma membrane localization of the D1 receptor. Polymerase chain reaction mutagenesis was used to generate a panel of D5 receptors containing mutations in the three predicted sites of N-linked glycosylation. Expression of mutant receptors indicated that glycosylation of residue N7 was the major determinant of D5 receptor plasma membrane localization. Mutation of a comparable site in the D1 receptor at position N5 had no effect on the delivery of the D1 receptor to the cell surface. Tunicamycin treatment during receptor biosynthesis, but not N-glycosidase F digestion of mature receptors, abrogated binding of the D5 receptor antagonist [(3)H]SCH23390, suggesting that while oligosaccharide moieties play a key role in the cell surface expression of D5 receptors, they do not appear to contribute to the receptor's ligand binding properties. Together, our data indicate a differential requirement for N-linked glycosylation in functional cell surface expression of D1 and D5 dopamine receptors. PMID- 10531416 TI - Beta2-adrenergic receptor agonists and cAMP arrest human cultured airway smooth muscle cells in the G(1) phase of the cell cycle: role of proteasome degradation of cyclin D1. AB - Hyperplasia of airway smooth muscle (ASM) contributes to the airway hyperresponsiveness that characterizes asthma. We have investigated the relationship between cAMP-induced growth arrest of ASM cells and thrombin stimulated, extracellular-regulated protein kinase (ERK) activity, cyclin D1, and the restriction protein retinoblastoma. The beta(2)-adrenergic receptor agonist albuterol (100 nM) inhibited DNA synthesis after incubation with ASM for periods as brief as 1 h when these coincided with the timing of the restriction point. Inhibition of thrombin-stimulated DNA synthesis by albuterol (1-100 nM), 8-bromo cAMP (300 microM), or prostaglandin E(2) (1 microM) was accompanied by a reduction in cyclin D1 protein levels. The ERK kinase inhibitor PD98059 (3-30 microM) attenuated thrombin-stimulated ERK phosphorylation and activity and the increase in cyclin D1 protein levels, as did albuterol (1-100 nM) or 8-bromo-cAMP (300 microM). In contrast, neither albuterol (100 nM) nor PD98059 (30 microM) reduced cyclin D1 mRNA levels between 4 and 20 h after thrombin addition, which suggests that elevation of cAMP regulates cyclin D1 by a post transcriptional mechanism. The proteasome inhibitor MG132 (30 and 100 nM) and the calpain I inhibitor N-acetyl-Leu-Leu-leucinal (10 microM) attenuated the reduction in thrombin-stimulated cyclin D1 levels in ASM exposed to albuterol (100 nM), 8 bromo-cAMP (300 microM), or the phosphodiesterase inhibitor isobutylmethylxanthine (100 microM). Thus, the cAMP-induced arrest of ASM in the G(1) phase of the cell cycle is associated with a proteasomal degradation of cyclin D1 protein and a reduced protein retinoblastoma phosphorylation that prevents passage through the restriction point. PMID- 10531417 TI - Agonist gating and isoflurane potentiation in the human gamma-aminobutyric acid type A receptor determined by the volume of a second transmembrane domain residue. AB - Gamma-aminobutyric acid type A (GABA(A) )receptors are targets for allosteric modulation by general anesthetics. Mutation of Ser270 within the second transmembrane domain of the GABA(A) receptor alpha subunit can ablate the modulation of the receptor by the anesthetic ether isoflurane. To investigate further the function of this critical amino acid residue, we made multiple amino acid substitutions at Ser270 and analyzed the concentration-dependent gating by GABA and regulation by isoflurane in each mutant receptor. There is a strong negative correlation between the EC(50) for GABA and the molecular volume of the amino acid residue at position 270. Replacement of Ser by large residues such as His and Trp produced a shift of the GABA concentration-response curve to the left, whereas replacement of Ser with Gly had the opposite effect. There also was a strong negative association between the molecular volume of the amino acid residue at 270 and the degree of enhancement of submaximal GABA responses by isoflurane. These results indicate the significance of the amino acid at position alpha270 in gating of the GABA(A) receptor. In addition, the data on isoflurane are consistent with the existence of a cavity of finite size in the region of alpha270 that may be filled by the anesthetic molecule or by the side chain of a larger residue at alpha270. The introduction of isoflurane, or of a large residue, into this cavity may stabilize the open state of the GABA(A) receptor relative to the closed state. PMID- 10531418 TI - Ca(2+) permeation of AMPA receptors in cerebellar neurons expressing glu receptor 2. AB - AMPA receptors in cultured cerebellar neurons were characterized by whole-cell electrophysiological studies and single cell PCR-based quantitation of subunit mRNA expression. Purkinje neurons consistently expressed high levels of Glu receptor 2 (GluR2) mRNA and AMPA receptors with low but nonzero Ca(2+) permeability. Other cerebellar neurons expressed AMPA receptors with a wide range of Ca(2+) permeability and of fractional GluR2. These properties correlated on a cell-by-cell basis. Their relationship was well fit by a model that assumed stochastic assembly of subunits and GluR2 dominance in controlling divalent cation permeation, suggesting that AMPA receptor properties in individual neurons may be determined primarily by relative levels of subunit transcription. A fraction of receptors, lacking GluR2, can contribute a highly Ca(2+)-permeable component to AMPA receptor responses, even in cells expressing GluR2. PMID- 10531419 TI - Receptors of the glial cell line-derived neurotrophic factor family of neurotrophic factors signal cell survival through the phosphatidylinositol 3 kinase pathway in spinal cord motoneurons. AB - The members of the glial cell line-derived neurotrophic factor (GDNF) family of neurotrophic factors (GDNF, neurturin, persephin, and artemin) are able to promote in vivo and in vitro survival of different neuronal populations, including spinal cord motoneurons. These factors signal via multicomponent receptors that consist of the Ret receptor tyrosine kinase plus a member of the GDNF family receptor alpha (GRFalpha) family of glycosylphosphatidylinositol linked coreceptors. Activation of the receptor induces Ret phosphorylation that leads the survival-promoting effects. Ret phosphorylation causes the activation of several intracellular pathways, but the biological effects caused by the activation of each of these pathways are still unknown. In the present work, we describe the ability of the GDNF family members to promote chicken motoneuron survival in culture. We show the presence of Ret and GFRalpha-1, GFRalpha-2, and GFRalpha-4 in chicken motoneurons using in situ hybridization and reverse transcription-PCR techniques. By Western blot analysis and kinase assays, we demonstrate the ability of these factors to induce the phosphatidylinositol 3 kinase (PI 3-kinase) and the extracellular regulated kinase (ERK)-mitogen activated protein (MAP) kinase pathways activation. To characterize the involvement of these pathways in the survival effect, we used the PI 3-kinase inhibitor LY 294002 and the MAP kinase and ERK kinase (MEK) inhibitor PD 98059. We demonstrate that LY 294002, but not PD 98059, prevents GDNF-, neurturin-, and persephin-induced motoneuron survival, suggesting that PI 3-kinase intracellular pathway is responsible in mediating the neurotrophic effect. PMID- 10531420 TI - The Alzheimer's disease amyloid precursor protein modulates copper-induced toxicity and oxidative stress in primary neuronal cultures. AB - The amyloid precursor protein (APP) of Alzheimer's disease can reduce copper (II) to copper (I) in a cell-free system potentially leading to increased oxidative stress in neurons. We used neuronal cultures derived from APP knock-out (APP(-/ )) and wild-type (WT) mice to examine the role of APP in copper neurotoxicity. WT cortical, cerebellar, and hippocampal neurons were significantly more susceptible than their respective APP(-/-) neurons to toxicity induced by physiological concentrations of copper but not by zinc or iron. There was no difference in copper toxicity between APLP2(-/-) and WT neurons, demonstrating specificity for APP-associated copper toxicity. Copper uptake was the same in WT and APP(-/-) neurons, suggesting APP may interact with copper to induce a localized increase in oxidative stress through copper (I) production. This was supported by significantly higher levels of copper-induced lipid peroxidation in WT neurons. Treatment of neuronal cultures with a peptide corresponding to the human APP copper-binding domain (APP142-166) potentiated copper but not iron or zinc toxicity. Incubation of APP142-166 with low-density lipoprotein (LDL) and copper resulted in significantly increased lipid peroxidation compared to copper and LDL alone. Substitution of the copper coordinating histidine residues with asparagines (APP142-166(H147N, H149N, H151N)) abrogated the toxic effects. A peptide corresponding to the zinc-binding domain (APP181-208) failed to induce copper or zinc toxicity in neuronal cultures. These data support a role for the APP copper-binding domain in APP-mediated copper (I) generation and toxicity in primary neurons, a process that has important implications for Alzheimer's disease and other neurodegenerative disorders. PMID- 10531421 TI - Multiple and opposing roles of cholinergic transmission in the main olfactory bulb. AB - The main olfactory bulb is a critical relay step between the olfactory epithelium and the olfactory cortex. A marked feature of the bulb is its massive innervation by cholinergic inputs from the basal forebrain. In this study, we addressed the functional interaction between cholinergic inputs and intrinsic bulbar circuitry. Determining the roles of acetylcholine (ACh) requires the characterization of cholinergic effects on both neural excitability and synaptic transmission. For this purpose, we used electrophysiological techniques to localize and characterize the diverse roles of ACh in mouse olfactory bulb slices. We found that cholinergic inputs have a surprising number of target receptor populations that are expressed on three different neuronal types in the bulb. Specifically, nicotinic acetylcholine receptors excite both the output neurons of the bulb, i.e., the mitral cells, as well as interneurons located in the periglomerular regions. These nicotine-induced responses in interneurons are short lasting, whereas responses in mitral cells are long lasting. In contrast, muscarinic receptors have an inhibitory effect on the firing rate of interneurons from a deeper layer, granule cells, while at the same time they increase the degree of activity-independent transmitter release from these cells onto mitral cells. Cholinergic signaling thus was found to have multiple and opposing roles in the olfactory bulb. These dual cholinergic effects on mitral cells and interneurons may be important in modulating olfactory bulb output to central structures required for driven behaviors and may be relevant to understanding mechanisms underlying the perturbations of cholinergic inputs to cortex that occur in Alzheimer's disease. PMID- 10531422 TI - A(2A) adenosine receptor deficiency attenuates brain injury induced by transient focal ischemia in mice. AB - Extracellular adenosine critically modulates ischemic brain injury, at least in part through activation of the A(1) adenosine receptor. However, the role played by the A(2A) receptor has been obscured by intrinsic limitations of A(2A) adenosinergic agents. To overcome these pharmacological limitations, we explored the consequences of deleting the A(2A) adenosine receptor on brain damage after transient focal ischemia. Cerebral morphology, as well as vascular and physiological measures (before, during, and after ischemia) did not differ between A(2A) receptor knock-out and wild-type littermates. The volume of cerebral infarction, as well as the associated neurological deficit induced by transient filament occlusion of the middle cerebral artery, were significantly attenuated in A(2A) receptor knock-out mice. This neuroprotective phenotype of A(2A) receptor-deficient mice was observed in different genetic backgrounds, confirming A(2A) receptor disruption as its cause. Together with complimentary pharmacological studies, these data suggest that A(2A) receptors play a prominent role in the development of ischemic injury within brain and demonstrate the potential for anatomical and functional neuroprotection against stroke by A(2A) receptor antagonists. PMID- 10531423 TI - Neuronal nitric oxide synthase mRNA upregulation in rat sensory neurons after spinal nerve ligation: lack of a role in allodynia development. AB - Pharmacological evidence suggests a functional role for spinal nitric oxide (NO) in the modulation of thermal and/or inflammatory hyperalgesia. To assess the role of NO in nerve injury-induced tactile allodynia, we examined neuronal NO synthase (nNOS) expression in the spinal cord and dorsal root ganglia (DRG) of rats with tactile allodynia because of either tight ligation of the left fifth and sixth lumbar spinal nerves or streptozotocin-induced diabetic neuropathy. RNase protection assays indicated that nNOS mRNA (1) was upregulated in DRG, but not spinal cord, neurons on the injury side beginning 1 d after nerve ligation, (2) peaked (approximately 10-fold increase) at 2 d, and (3) remained elevated for at least 13 weeks. A corresponding increase in DRG nNOS protein was also observed and localized principally to small and occasionally medium-size sensory neurons. In rats with diabetic neuropathy, there was no significant change in DRG nNOS mRNA. However, similar increases in DRG nNOS mRNA were observed in rats that did not develop allodynia after nerve ligation and in rats fully recovered from allodynia 3 months after the nerve ligation. Systemic treatment with a specific pharmacological inhibitor of nNOS failed to prevent or reverse allodynia in nerve injured rats. Thus, regulation of nNOS may contribute to the development of neuronal plasticity after specific types of peripheral nerve injury. However, upregulation of nNOS is not responsible for the development and/or maintenance of allodynia after nerve injury. PMID- 10531424 TI - Optical imaging reveals elevated intracellular chloride in hippocampal pyramidal neurons after oxidative stress. AB - The accumulation of reactive oxygen species (ROS) in the brain is associated with several neurodegenerative conditions. ROS can affect ionic homeostasis leading to impaired neurotransmission. Here, we determined the ability of H(2)O(2), a membrane permeant ROS, to alter intraneuronal Cl(-), an important regulator of neuronal excitability. Real-time alterations in intracellular chloride, [Cl(-)]i, were measured with UV laser scanning confocal microscopy in hippocampal slices loaded with the cell-permeant form of 6-methoxy-N-ethylquinolium iodide (MEQ), a Cl(-)-sensitive fluorescent probe. In slices superfused with H(2)O(2) for 10 min, there was a significant decrease in MEQ fluorescence (elevation in [Cl(-)]i) in area CA1 pyramidal cell soma but not in interneurons located in stratum radiatum. Alterations in [Cl(-)]i induced by H(2)O(2) were prevented by the iron chelator deferoxamine and the vitamin E analog Trolox, suggesting the involvement of free radicals. The influx of Cl(-) probably occurred through the GABA-gated Cl(-) channel because the effects of H(2)O(2) were blocked by picrotoxin. In addition, HPLC analysis of the superfusates indicated that GABA and glutamate accumulated extracellularly after H(2)O(2) exposure. Excitatory amino acid receptor antagonists 2-amino-5-phoshopentanoic acid and 1,2,3,4-tetrahydro-6-nitro-2, 3 dioxo-benzo[f]quinoxaline-7-sulfonamide also attenuated the effect of H(2)O(2) on MEQ fluorescence. The changes in [Cl(-)]i induced by H(2)O(2) were Ca(2+) dependent and Na(+)-independent. After exposure of slices to H(2)O(2), the ability of the GABA agonist muscimol to increase [Cl(-)]i was attenuated. Thus, ROS, like H(2)O(2), may impair transmembrane Cl(-) gradients and reduce inhibitory neurotransmission, further promoting neuronal damage in oxidative stress-related disease and in aging. PMID- 10531425 TI - Knockdown of AMPA receptor GluR2 expression causes delayed neurodegeneration and increases damage by sublethal ischemia in hippocampal CA1 and CA3 neurons. AB - Considerable evidence suggests that Ca(2+)-permeable AMPA receptors are critical mediators of the delayed, selective neuronal death associated with transient global ischemia and sustained seizures. Global ischemia suppresses mRNA and protein expression of the glutamate receptor subunit GluR2 and increases AMPA receptor-mediated Ca(2+) influx into vulnerable neurons of the hippocampal CA1 before the onset of neurodegeneration. Status epilepticus suppresses GluR2 mRNA and protein in CA3 before neurodegeneration in this region. To examine whether acute downregulation of the GluR2 subunit, even in the absence of a neurological insult, can cause neuronal cell death, we performed GluR2 "knockdown" experiments. Intracerebral injection of antisense oligodeoxynucleotides targeted to GluR2 mRNA induced delayed death of pyramidal neurons in CA1 and CA3. Antisense-induced neurodegeneration was preceded by a reduction in GluR2 mRNA, as indicated by in situ hybridization, and in GluR2 protein, as indicated by Western blot analysis. GluR2 antisense suppressed GluR2 mRNA in the dentate gyrus but did not cause cell death. The AMPA receptor antagonist 6-cyano-7-nitroquinoxiline-2,3 dione (CNQX) and the Ca(2+)-permeable AMPA receptor channel blocker 1-naphthyl acetyl spermine protected against antisense-induced cell death. This result indicates that antisense-induced cell death is mediated by Ca(2+)-permeable AMPA receptors. GluR2 antisense and brief sublethal global ischemia acted synergistically to cause degeneration of pyramidal neurons, consistent with action by a common mechanism. These findings demonstrate that downregulation of GluR2 is sufficient to induce delayed death of specific neuronal populations. PMID- 10531426 TI - Subunit-specific association of protein kinase C and the receptor for activated C kinase with GABA type A receptors. AB - GABA receptors (GABA(A)) are the major sites of fast synaptic inhibition in the brain and can be assembled from five subunit classes: alpha, beta, gamma, delta, and epsilon. Receptor function can be regulated by direct phosphorylation of beta and gamma2 subunits, but how kinases are targeted to GABA(A) receptors is unknown. Here we show that protein kinase C-betaII (PKC-betaII) is capable of directly binding to the intracellular domain of the receptor beta1 and beta3 subunits, but not to those of the alpha1 or gamma2 subunits. Moreover, associating PKC-betaII is capable of specifically phosphorylating serine 409 in beta1 subunit and serines 408/409 within the beta3 subunit, key residues for modulating GABA(A) receptor function. The receptor for activated C kinase (RACK 1) was found also to bind to the beta1 subunit intracellular domain, but PKC binding appeared to be independent of this protein. Using immunoprecipitation, the association of PKC isoforms and RACK-1 with neuronal GABA(A) receptors was seen. Furthermore, PKC isoforms associating with neuronal receptors were capable of phosphorylating the receptor beta3 subunit. Together, these observations suggest GABA(A) receptors are intimately associated with PKC isoforms via a direct interaction with receptor beta subunits. This interaction may serve to localize PKC activity to GABA(A) receptors in neurons allowing the rapid regulation of receptor activity by cell-signaling pathways that modify PKC activity. PMID- 10531427 TI - An R-type Ca(2+) current in neurohypophysial terminals preferentially regulates oxytocin secretion. AB - Multiple types of voltage-dependent Ca(2+) channels are involved in the regulation of neurotransmitter release (Tsien et al., 1991; Dunlap et al., 1995). In the nerve terminals of the neurohypophysis, the roles of L-, N-, and P/Q-type Ca(2+) channels in neuropeptide release have been identified previously (Wang et al., 1997a). Although the L- and N-type Ca(2+) currents play equivalent roles in both vasopressin and oxytocin release, the P/Q-type Ca(2+) current only regulates vasopressin release. An oxytocin-release and Ca(2+) current component is resistant to the L-, N-, and P/Q-type Ca(2+) channel blockers but is inhibited by Ni(2+). A new polypeptide toxin, SNX-482, which is a specific alpha(1E)-type Ca(2+) channel blocker (Newcomb et al., 1998), was used to characterize the biophysical properties of this resistant Ca(2+) current component and its role in neuropeptide release. This resistant component was dose dependently inhibited by SNX-482, with an IC(50) of 4.1 nM. Furthermore, SNX-482 did not affect the other Ca(2+) current types in these CNS terminals. Like the N- and P/Q-type Ca(2+) currents, this SNX-482-sensitive transient Ca(2+) current is high-threshold activated and shows moderate steady-state inactivation. At the same concentrations, SNX-482 blocked the component of oxytocin, but not of vasopressin, release that was resistant to the other channel blockers, indicating a preferential role for this type of Ca(2+) current in oxytocin release from neurohypophysial terminals. Our results suggest that an alpha(1E) or "R"-type Ca(2+) channel exists in oxytocinergic nerve terminals and, thus, functions in controlling only oxytocin release from the rat neurohypophysis. PMID- 10531428 TI - Substrate turnover by transporters curtails synaptic glutamate transients. AB - Although inhibitors of glutamate transport prolong synaptic currents at many glutamate synapses, the cause of the current prolongation is unclear. Transport inhibitors may prolong synaptic currents by simply interfering with synaptic glutamate binding to transporters, by inhibiting substrate translocation, or by promoting accumulation of ambient glutamate, which may act cooperatively at receptors with synaptic glutamate. We show that reversal of the membrane potential of astrocytes surrounding the synapse prolongs synaptic currents but does not decrease the apparent affinity of transporters or significantly alter glutamate-dependent kinetics of macroscopic transporter currents in excised membrane patches. Positive membrane potentials do not affect binding of a nontransported glutamate analog, nor do positive membrane potentials alter the number of transporters available to bind analog. We also test the hypothesis that glutamate accumulation during uptake inhibition by transporter substrates is the direct cause of synaptic current prolongations. Transporter substrates elevate ambient glutamate near synapses by fostering reverse transport of endogenous glutamate. However, increases in ambient glutamate cannot account for the prolongations of synaptic currents, because a nonsubstrate transport inhibitor does not foster reverse uptake yet it prolongs synaptic currents. Moreover, exogenous glutamate does not mimic synaptic current prolongations induced by substrate inhibitors. These results provide strong support for a major role of substrate translocation in determining the time course of the glutamate concentration transient at excitatory synapses. PMID- 10531429 TI - Pharmacological isolation of the synaptic and nonsynaptic components of the GABA mediated biphasic response in rat CA1 hippocampal pyramidal cells. AB - High-frequency stimulation (HFS) applied to stratum radiatum of a rat hippocampal slice in the presence of ionotropic glutamate receptor antagonists evokes a biphasic GABA(A) receptor-dependent response in CA1 pyramidal neurons, with a brief hyperpolarizing IPSP (hIPSP) followed by a long-lasting depolarization. We show now that it is possible to pharmacologically separate the hIPSP and late depolarization from one another. In neurons intracellularly perfused for 1-2 hr with F(-) as the major anion and no ATP, the hIPSP (and the corresponding current, hIPSC) evoked by HFS was blocked, whereas neither the late depolarization nor its underlying current was attenuated. In contrast, internal perfusion with a high concentration (5 mM) of the impermeant lidocaine derivative QX-314 selectively abolished the depolarizing component of the biphasic response and also strongly reduced depolarizations evoked by extracellular microinjection of K(+). Bath application of quinine (0. 2-0.5 mM) or quinidine (0.1 mM) resulted in a pronounced inhibition of the HFS-induced extracellular K(+) concentration ([K(+)](o)) transient but not of the bicarbonate-dependent alkaline shift in extracellular pH. The attenuation of the [K(+)](o) transient was closely paralleled by a suppression of the HFS-evoked depolarization but not of the hIPSP. Quini(di)ne did not affect depolarizations induced by exogenous K(+) either. These data provide direct pharmacological evidence for the view that the HFS-induced biphasic response of the pyramidal neuron is composed of mechanistically distinct components: a direct GABA(A) receptor-mediated phase, which is followed by a slow, nonsynaptic [K(+)](o)-mediated depolarization. The bicarbonate-dependent, activity-induced [K(+)](o) transient can be blocked by quini(di)ne, whereas its depolarizing action in the pyramidal neuron is inhibited by internal QX-314. The presence of fundamentally distinct components in GABA(A) receptor-mediated actions evoked by HFS calls for further investigations of their functional role(s) in standard experimental maneuvers, such as those used in studies of synaptic plasticity and induction of gamma oscillations. PMID- 10531430 TI - Mitochondria regulate the Ca(2+)-exocytosis relationship of bovine adrenal chromaffin cells. AB - The present study expands the contemporary view of mitochondria as important participants in cellular Ca(2+) dynamics and provides evidence that mitochondria regulate the supply of release-competent secretory granules. Using pharmacological probes to inhibit mitochondrial Ca(2+) import, the ability of mitochondria to modulate secretory activity in single, patch-clamped bovine chromaffin cells was examined by simultaneously monitoring rapid changes in membrane surface area (DeltaC(m)) and cytosolic Ca(2+) levels ([Ca(2+)](c)). Repetitive step depolarizations or action potential waveforms were found to raise the [Ca(2+)](c) of chromaffin cells into the 1 microM to tens of micromolar range. Inhibiting mitochondria by treatment with carbonyl cyanide p-(trifuoro methoxy)phenylhydrazone, antimycin-oligomycin, or ruthenium red revealed that mitochondria are a prominent component for the clearance of Ca(2+) that entered via voltage-activated Ca(2+) channels. Disruption of cellular Ca(2+) homeostasis by poisoning mitochondria enhanced the secretory responsiveness of chromaffin cells by increasing the amplitude of the transient rise and the time course of recovery to baseline of the evoked Delta[Ca(2+)](c). The enhancement of the secretory response was represented by significant deviation of the Ca(2+) exocytosis relationship from a standard relationship that equates Ca(2+) influx and DeltaC(m). Thus, mitochondria would play a critical role in the control of secretory activity in chromaffin cells that undergo tonic or repetitive depolarizing activity, likely by limiting the Ca(2+)-dependent activation of specific proteins that recruit or prime secretory granules for exocytosis. PMID- 10531431 TI - The CB1 cannabinoid receptor can sequester G-proteins, making them unavailable to couple to other receptors. AB - We tested the hypothesis that human CB1 cannabinoid receptors (hCB1) can sequester G(i/o)-proteins from a common pool and prevent other receptors from signaling. Human CB1 cannabinoid receptors were expressed in superior cervical ganglion (SCG) neurons by microinjection of hCB1 cDNA. Expression of hCB1 cannabinoid receptors abolished the Ca(2+) current inhibition by endogenous pertussis toxin-sensitive G(i/o)-coupled receptors for norepinephrine (NE) and somatostatin (SOM) but not by endogenous pertussis toxin-insensitive G(s)-coupled receptors for vasoactive intestinal polypeptide. Signaling by NE was rescued by expression of Galpha(oB), Gbeta(1), and Ggamma(3). Expression of mGluR2 metabotropic glutamate receptors, another pertussis toxin-sensitive G-protein coupled receptor, had no effect on the signaling by NE or SOM. Some hCB1 receptors were constitutively active because the cannabinoid receptor inverse agonist SR 141617A enhanced the Ca(2+) current. Some hCB1 receptors also appear to be precoupled to G(i/o)-proteins because the cannabinoid agonist WIN 55,212-2 decreased the Ca(2+) current at a time when no G-proteins were available to couple to alpha(2)-adrenergic and somatostatin receptors. In SCG neurons microinjected with a lower concentration of hCB1 cDNA, the effect of SR 141716A was reduced, and the response to NE and SOM was partially restored. Subsequent to the application of SR 141716A, the Ca(2+) current inhibition by NE and SOM was abolished. These results suggest that both the active and inactive states of the hCB1 receptor can sequester G(i/o)-proteins from a common pool. Cannabinoid receptors thus have the potential to prevent other G(i/o)-coupled receptors from transducing their biological signals. PMID- 10531432 TI - Receptor subtype-induced targeting and subtype-specific internalization of human alpha(2)-adrenoceptors in PC12 cells. AB - The three alpha(2)-adrenergic receptor subtypes have distinct tissue distributions, desensitization properties, and, in some cell types, subtype specific subcellular localization and trafficking properties. The subtypes also differ in their neuronal physiology. Therefore, we have investigated the localization and targeting of human alpha(2)-adrenoceptors (alpha(2)-AR) in PC12 cells, which were transfected to express the alpha(2)-AR subtypes A, B, and C. Inspection of the receptors by indirect immunofluorescence and confocal microscopy showed that alpha(2A)-AR were mainly targeted to the tips of the neurites, alpha(2B)-AR were evenly distributed in the plasma membrane, and alpha(2C)-AR were mostly located in an intracellular perinuclear compartment. After agonist treatment, alpha(2A)- and alpha(2B)-AR were internalized into partly overlapping populations of intracellular vesicles. Receptor subtype specific changes in PC12 cell morphology were also discovered: expression of alpha(2A)-AR, but not of alpha(2B)- or alpha(2C)-AR, induced differentiation-like changes in cells not treated with NGF. Also alpha(2B)-AR were targeted to the tips of neurites when they were coexpressed in the same cells with alpha(2A)-AR, indicating that the targeting of receptors to the tips of neurites is a consequence of a change in PC12 cell membrane protein trafficking that the alpha(2A)-subtype induces. The marked agonist-induced internalization of alpha(2A)-AR observed in both nondifferentiated and differentiated PC12 cells contrasts with earlier results from non-neuronal cells and points out the importance of the cellular environment for receptor endocytosis and trafficking. The targeting of alpha(2A)-AR to nerve terminals in PC12 cells is in line with the putative physiological role of this receptor subtype as a presynaptic autoreceptor. PMID- 10531433 TI - Loss of postsynaptic GABA(A) receptor clustering in gephyrin-deficient mice. AB - The tubulin-binding protein gephyrin, which anchors the inhibitory glycine receptor (GlyR) at postsynaptic sites, decorates GABAergic postsynaptic membranes in various brain regions, and postsynaptic gephyrin clusters are absent from cortical cultures of mice deficient for the GABA(A) receptor gamma2 subunit. Here, we investigated the postsynaptic clustering of GABA(A) receptors in gephyrin knock-out (geph -/-) mice. Both in brain sections and cultured hippocampal neurons derived from geph -/- mice, synaptic GABA(A) receptor clusters containing either the gamma2 or the alpha2 subunit were absent, whereas glutamate receptor subunits were normally localized at postsynaptic sites. Western blot analysis and electrophysiological recording revealed that normal levels of functional GABA(A) receptors are expressed in geph -/- neurons, however the pool size of intracellular GABA(A) receptors appeared increased in the mutant cells. Thus, gephyrin is required for the synaptic localization of GlyRs and GABA(A) receptors containing the gamma2 and/or alpha2 subunits but not for the targeting of these receptors to the neuronal plasma membrane. In addition, gephyrin may be important for efficient membrane insertion and/or metabolic stabilization of inhibitory receptors at developing postsynaptic sites. PMID- 10531434 TI - Multiorgan autonomic dysfunction in mice lacking the beta2 and the beta4 subunits of neuronal nicotinic acetylcholine receptors. AB - Transcripts for the beta2 and the beta4 nicotinic acetylcholine receptor (nAChR) subunits are found throughout the CNS and the peripheral nervous system. These two beta subunits can form heteromultimeric channels with any of the alpha2, alpha3, alpha4, or alpha5 subunits in heterologous expression systems. Nonetheless, the subunit composition of native nAChRs and the role of different nAChR subtypes in vivo remain unclear. We prepared null mutations for the beta2 and the beta4 genes and bred beta2-/-beta4-/- mice by mating mice of identical beta2-/-beta4+/- or beta2+/-beta4-/- genotype. The beta2-/- and the beta4-/- single-mutant mice grow to adulthood with no visible phenotypic abnormalities. The beta2-/-beta4-/- double mutants survive to birth but have impaired growth and increased perinatal mortality. They also present enlarged bladders with dribbling urination and develop urinary infection and bladder stones. The ocular pupils are widely dilated and do not constrict in response to light. Histological studies revealed no significant abnormalities of brain or peripheral tissues except for hyperplasia in the bladder mucosa of beta4-/- and beta2-/-beta4-/- mutants. Bladder strips from beta2-/-beta4-/- mice did not respond to nicotine but contracted when stimulated with a muscarinic agonist or electric field stimulation. Bladder strips from beta4 mutants did not respond to nicotine despite the absence of major bladder dysfunction in vivo. Acetylcholine-activated whole-cell currents were absent in superior cervical ganglion neurons from beta2 /-beta4-/- mice and reduced in neurons from beta4-/- mice. Although there is apparent redundancy and a superficially normal phenotype in beta2-/- and beta4-/- mice, physiological studies indicate major deficits in the beta4-/- mice. Our previous description of a similar phenotype in alpha3-/- mice and the current data suggest that the alpha3 and the beta4 subunits are major components in autonomic nAChRs. The phenotype of the beta2-/-beta4-/- and alpha3-/- mice resembles the autosomal recessive megacystis-microcolon-hypoperistalsis syndrome in humans. PMID- 10531435 TI - Excitatory synaptogenesis between identified Lymnaea neurons requires extrinsic trophic factors and is mediated by receptor tyrosine kinases. AB - Neurotrophic factors have well established roles in neuronal development and adult synaptic plasticity, but their precise role in synapse formation has yet to be determined. This paper provides the first direct evidence that neurotrophic factors in brain conditioned medium (CM) differentially regulate excitatory and inhibitory synapse formation. Somata of identified presynaptic and postsynaptic neurons were isolated from the CNS of Lymnaea and were cultured in a soma-soma configuration in the presence (CM) or absence [defined medium (DM)] of trophic factors. In DM, excitatory synapses did not form. When they were paired in CM or in DM containing Lymnaea epidermal growth factor (EGF); however, all presynaptic neurons reestablished their specific excitatory synapses, which had electrical properties similar to those seen in vivo. CM-induced formation of excitatory synapses required transcription and de novo protein synthesis, as indicated by the observations that synapse formation was blocked by the protein synthesis inhibitor anisomycin and the protein transcription blocker actinomycin D; the CM factor was inactivated by boiling. They were also blocked by receptor tyrosine kinase inhibitors (lavendustin A, genistein, K252a, and KT5926) but not by inactive analogs (genistin and lavendustin B), suggesting that the effect was mediated by receptor tyrosine kinases. These results, together with our previously published data, demonstrate that trophic factors are required for excitatory, but not inhibitory, synapse formation and extends the role of EGF from cell proliferation, neurite outgrowth, and survival to excitatory synapse formation. PMID- 10531436 TI - Targeted deletion of a cyclic nucleotide-gated channel subunit (OCNC1): biochemical and morphological consequences in adult mice. AB - The olfactory cyclic nucleotide-gated channel subunit 1 (OCNC1) is required for signal transduction in olfactory receptor cells. To further investigate the role of this channel in the olfactory system, the biochemical and morphological consequences of targeted disruption of OCNC1 were investigated in adult mice. Null as compared to wild-type mice had smaller olfactory bulbs, suggesting compromised development of the central target of the receptor cells. Ectopic olfactory marker protein (OMP)-stained fibers localized to the external plexiform layer reflected the relative immaturity of the olfactory bulb in the null mice. The olfactory epithelium of the knock-out mouse was thinner and showed lower expression of olfactory marker protein and growth-associated protein 43, indicating decreases in both generation and maturation of receptor cells. Tyrosine hydroxylase (TH) expression in the olfactory bulb, examined as a reflection of afferent activity, was reduced in the majority of periglomerular neurons but retained in atypical or "necklace" glomeruli localized to posterior aspects of the olfactory bulb. Double label studies demonstrated that the remaining TH-immunostained neurons received their innervation from a subset of receptor cells previously shown to express a phosphodiesterase that differs from that found in most receptor cells. These data indicate that expression of OCNC1 is required for normal development of the olfactory epithelium and olfactory bulb. The robust expression of TH in some periglomerular cells in the OCNC1-null mice suggests that receptor cells innervating these glomeruli may use an alternate signal transduction pathway. PMID- 10531437 TI - Analysis of the retrograde transport of glial cell line-derived neurotrophic factor (GDNF), neurturin, and persephin suggests that in vivo signaling for the GDNF family is GFRalpha coreceptor-specific. AB - Neurturin (NRTN) and glial cell line-derived neurotrophic factor (GDNF) are members of a family of trophic factors with similar actions in vitro on certain neuronal classes. Retrograde transport of GDNF and NRTN was compared in peripheral sensory, sympathetic, and motor neurons to determine whether in vivo these factors are transported selectively by different neuronal populations. After sciatic nerve injections, NRTN was transported by sensory neurons of the dorsal root ganglion (DRG). Competition studies demonstrated only limited cross competition between NRTN and GDNF, indicating selective receptor-mediated transport of these factors. By using immunohistochemistry, we identified two populations of NRTN-transporting DRG neurons: a major population of small, RET positive, IB4-positive, non-TrkA-expressing neurons that also show the ability to transport GDNF and a minor population of calretinin-expressing neurons that fail to transport GDNF. Spinal motor neurons in the adult showed relatively less ability to transport NRTN than to transport GDNF, although NRTN prevented the cell death of neonatal motor neurons in a manner very similar to GDNF (Yan et al., 1995) and persephin (PSPN) (Milbrandt et al., 1998). Last, NRTN, like GDNF, was not transported to sympathetic neurons of the adult superior cervical ganglion (SCG) after injection into the anterior eye chamber. These data reveal a high degree of functional selectivity of GDNF family receptor-alpha (GFRalpha) coreceptor subtypes for NRTN and GDNF in vivo. PMID- 10531438 TI - K(+) channel expression distinguishes subpopulations of parvalbumin- and somatostatin-containing neocortical interneurons. AB - Kv3.1 and Kv3.2 K(+) channel proteins form similar voltage-gated K(+) channels with unusual properties, including fast activation at voltages positive to -10 mV and very fast deactivation rates. These properties are thought to facilitate sustained high-frequency firing. Kv3.1 subunits are specifically found in fast spiking, parvalbumin (PV)-containing cortical interneurons, and recent studies have provided support for a crucial role in the generation of the fast-spiking phenotype. Kv3.2 mRNAs are also found in a small subset of neocortical neurons, although the distribution of these neurons is different. We raised antibodies directed against Kv3.2 proteins and used dual-labeling methods to identify the neocortical neurons expressing Kv3.2 proteins and to determine their subcellular localization. Kv3.2 proteins are prominently expressed in patches in somatic and proximal dendritic membrane as well as in axons and presynaptic terminals of GABAergic interneurons. Kv3.2 subunits are found in all PV-containing neurons in deep cortical layers where they probably form heteromultimeric channels with Kv3.1 subunits. In contrast, in superficial layer PV-positive neurons Kv3.2 immunoreactivity is low, but Kv3.1 is still prominently expressed. Because Kv3.1 and Kv3.2 channels are differentially modulated by protein kinases, these results raise the possibility that the fast-spiking properties of superficial- and deep layer PV neurons are differentially regulated by neuromodulators. Interestingly, Kv3. 2 but not Kv3.1 proteins are also prominent in a subset of seemingly non fast-spiking, somatostatin- and calbindin-containing interneurons, suggesting that the Kv3.1-Kv3.2 current type can have functions other than facilitating high frequency firing. PMID- 10531439 TI - Loss of synaptic depression in mammalian anterior cingulate cortex after amputation. AB - Two forms of activity-dependent long-term depression (LTD) in the CNS, as defined by their sensitivity to the blockade of NMDA receptors, are thought to be important in learning, memory, and development. Here, we report that NMDA receptor-independent LTD is the major form of long-term plasticity in the anterior cingulate cortex (ACC). Both L-type voltage-gated calcium channels and metabotropic glutamate receptors are required for inducing LTD. Amputation of a third hindpaw digit in an adult rat induced rapid expression of immediate early genes in the ACC bilaterally and caused a loss of LTD that persisted for at least 2 weeks. Our results suggest that synaptic LTD in the ACC may contribute to enhanced neuronal responses to subsequent somatosensory stimuli after amputation. PMID- 10531440 TI - Glucocorticoid receptor expression in the spinal cord after traumatic injury in adult rats. AB - Methylprednisolone (MP), a glucocorticoid, is the only effective therapeutic agent used in the clinical treatment of acute spinal cord injury (SCI). MP given within 8 hr after SCI significantly improves neurological function. Although the glucocorticoid receptor (GR) is suggested to mediate MP actions, limited knowledge is available on its expression and possible function after SCI. Presently, the expression of GR was studied in a weight-drop SCI model in adult rats. Immunohistochemistry and Western blot analysis revealed an increase in GR protein expression as early as 15 min after injury. GR expression sharply increased at 4 hr (22-fold), peaked at 8 hr (56-fold), rapidly declined at 1 d, and returned to the baseline level at and after 3 d. During its peak expression, GR was localized in neural somata and dendrites but not in axons and their terminals. GR immunoreactivity was also found in oligodendrocytes and astrocytes. Interestingly, other cell types, such as endothelial cells, were GR-negative. An increase in the binding activity of nuclear proteins to the glucocorticoid responsive element was also observed after SCI, demonstrating a functional element of GR activation. Finally, colocalization of GR and tumor necrosis factor alpha (TNF-alpha), an inflammatory cytokine, was observed in neurons and glial cells, consistent with MP regulation of TNF-alpha in this model. Thus, the transient expression of high levels of GR after SCI may provide new insights into the anti-inflammatory action of MP. PMID- 10531441 TI - Neuralization of the Xenopus embryo by inhibition of p300/ CREB-binding protein function. AB - p300/ CREB-binding protein (CBP) is a transcriptional coactivator for a plethora of transcription factors and plays critical roles in signal transduction pathways. We report that the inhibition of p300/CBP function in the Xenopus embryo abolishes non-neural tissue formation and, strikingly, initiates neural induction and primary neurogenesis in the entire embryo. The observed neuralization is achieved in the absence of anterior or posterior gene expression, suggesting that neural fate activation and anterior patterning may represent distinct molecular events. We further demonstrate that the neuralizing and anteriorizing activities of chordin and noggin are separable properties of these neural inducers. This study reveals that all embryonic cells possess intrinsic neuralizing capability and that p300/CBP function is essential for embryonic germ layer formation and neural fate suppression during vertebrate embryogenesis. PMID- 10531442 TI - Targeted expression of truncated glued disrupts giant fiber synapse formation in Drosophila. AB - Glued(1) (Gl(1)) mutants produce a truncated protein that acts as a poison subunit and disables the cytoplasmic retrograde motor dynein. Heterozygous mutants have axonal defects in the adult eye and the nervous system. Here we show that selective expression of the poison subunit in neurons of the giant fiber (GF) system disrupts synaptogenesis between the GF and one of its targets, the tergotrochanteral motorneuron (TTMn). Growth and pathfinding by the GF axon and the TTMn dendrite are normal, but the terminal of the GF axon fails to develop normally and becomes swollen with large vesicles. This is a presynaptic defect because expression of truncated Glued restricted to the GF results in the same defect. When tested electrophysiologically, the flies with abnormal axons show a weakened or absent GF-TTMn connection. In Glued(1) heterozygotes, GF-TTMn synapse formation appears morphologically normal, but adult flies show abnormal responses to repetitive stimuli. This physiological effect is also observed when tetanus toxin is expressed in the GFs. Because the GF-TTMn is thought to be a mixed electrochemical synapse, the results show that Glued has a role in assembling both the chemical and electrical components. We speculate that disrupting transport of a retrograde signal disrupts synapse formation and maturation. PMID- 10531443 TI - Lesions of an avian forebrain nucleus that disrupt song development alter synaptic connectivity and transmission in the vocal premotor pathway. AB - The avian forebrain nucleus, the lateral magnocellular nucleus of the anterior neostriatum (LMAN), is necessary for normal song development because LMAN lesions made in juvenile birds disrupt song production but do not disrupt song when made in adults. Although these age-limited behavioral effects implicate LMAN in song learning, a potential confound is that LMAN lesions could disrupt normal vocal motor function independent of any learning role by altering LMAN's premotor target, the song nucleus, the robust nucleus of the archistriatum (RA). To date, however, no studies have examined directly the effects of LMAN lesions on the circuitry of the RA. We report here that juvenile LMAN lesions rapidly and profoundly affect RA, altering synaptic connectivity within this nucleus, including descending inputs from the song nucleus HVc. Specifically, morphological assays of the dendritic spines of RA projection neurons and axon terminal boutons arising from HVc show a numerical decline in the density of connections in RA in LMAN-lesioned juveniles compared with controls. Concurrently, LMAN lesions alter excitatory transmission within the juvenile RA: after LMAN lesions, the stimulus-response relationship between HVc fibers and RA neurons steepens, and the amplitude of spontaneous monophasic EPSCs increases. Rather than arresting RA in a juvenile state, LMAN lesions transform the structure and function of RA and its connections, such that it is distinct from that of the normal juvenile. In many ways, RA circuitry in LMAN-lesioned juveniles resembles that of normal adults, suggesting that LMAN lesions induce a premature maturation of the vocal motor pathway, which may lead to a loss of behavioral plasticity and abnormal song development. PMID- 10531444 TI - Disruption of laminin beta2 chain production causes alterations in morphology and function in the CNS. AB - From the elegant studies of Ramon y Cajal (1909) to the current advances in molecular cloning (e.g., Farber and Danciger, 1997), the retina has served as an ideal model for the entire CNS. We have taken advantage of the well described anatomy, physiology, and molecular biology of the retina to begin to examine the role of the laminins, one component of the extracellular matrix, on the processes of neuronal differentiation and synapse formation in the CNS. We have examined the effect of the deletion of one laminin chain, the beta2 chain, on retinal development. The gross development of retinas from laminin beta2 chain-deficient animals appears normal, and photoreceptors are formed. However, these retinas exhibit several pathologies: laminin beta2 chain-deficient mice display abnormal outer segment elongation, abnormal electroretinograms, and abnormal rod photoreceptor synapses. Morphologically, the outer segments are reduced by 50% in length; the outer plexiform layer of mutant animals is disrupted specifically, because only 7% of observed rod invaginating synapses appear normal, whereas the inner plexiform layer is undisturbed; finally, the rate of apoptosis in the mutant photoreceptor layer is twice that of control mice. Physiologically, the electroretinogram is altered; the amplitude of the b-wave and the slope of the b wave intensity-response function are both decreased, consistent with synaptic disruption in the outer retina. Together, these results emphasize the prominence of the extracellular matrix and, in particular, the laminins in the development and maintenance of synaptic function and morphogenesis in the CNS. PMID- 10531445 TI - Unilateral GluR2(B) hippocampal knockdown: a novel partial seizure model in the developing rat. AB - Kainic acid (KA) induces status epilepticus in both adult and young rats but with different consequences on pathology and gene expression. In adults, GluR2(B) AMPA subunit expression is markedly reduced in CA3 neurons before neurodegeneration. In pups, the GluR2(B) subunit is sustained, possibly contributing to neuronal survival. Mechanisms underlying the reduced vulnerability of developing neurons to seizures was investigated by examining the effects of unilateral microinfusions of GluR2(B) antisense oligodeoxynucleotides (AS-ODNs) into the hippocampus of young rats in the presence or absence of a subconvulsive dose of KA. GluR2(B) AS-ODN infusions resulted in spontaneous seizure-like behavior, high stimulus intensity population spikes in the absence of long-term potentiation, and neurodegeneration of CA3 neurons lateral to the infusion site. Electroencephalography revealed paroxysmal activity and high-frequency high amplitude discharges associated with vigorous and continuous scratching, wild running, or bilateral jerking movements. Pups lacking phenotypic behavior exhibited high-rhythmic oscillations and status epilepticus by the dose of KA used. Radiolabeled AS-ODNs accumulated throughout the ipsilateral dorsal hippocampus. GluR2(B) but not GluR1(A) receptor protein was markedly reduced after GluR2(B) knockdown. In contrast, GluR1(A) knockdown reduced GluR1(A) but not GluR2(B) protein without change in behavior or morphology. Therefore, unilateral downregulation of hippocampal GluR2(B) but not GluR1(A) protein reduces the seizure threshold and survival of CA3 neurons in the immature hippocampus, possibly providing a novel partial seizure model in the developing rat. PMID- 10531446 TI - Regulation of neuregulin-mediated acetylcholine receptor synthesis by protein tyrosine phosphatase SHP2. AB - Synapse-specific expression of the nicotinic acetylcholine receptor (AChR) is believed to be mediated by neuregulin, an epidermal growth factor-like trophic factor released by somatic motoneurons at the neuromuscular junction (NMJ). Neuregulin stimulates ErbB2, ErbB3, and ErbB4, members of the ErbB family of receptor tyrosine kinases. SHP2 is a cytoplasmic protein tyrosine phosphatase containing two Src homology 2 domains near its N terminus, and has been shown to be a positive mediator of mitogenic responses to various growth factors. We found that SHP2 interacted with ErbB2 and ErbB3 after neuregulin stimulation of muscle cells. Expression of SHP2 in C2C12 mouse muscle cells attenuated the neuregulin induced expression of an AChR epsilon-promoter reporter gene, whereas a catalytically inactive SHP2 mutant or a mutant lacking the N-terminal Src homology 2 (SH2) domain enhanced reporter expression, suggesting that SHP2 negatively regulates the neuregulin signaling pathway. In fibroblast cells that express a mutant SHP2 with a targeted deletion of the N-terminal SH2 domain, neuregulin-mediated activation of the Ras/Raf/extracellular signal-regulated kinase cascade was enhanced. Furthermore, we found that SHP2 immunoreactivity colocalized with the staining of alpha-bungarotoxin, a marker of the NMJ. These results demonstrate a negative role of SHP2 in the neuregulin signal that leads to AChR gene expression at the NMJ. PMID- 10531447 TI - Membrane recycling in the neuronal growth cone revealed by FM1-43 labeling. AB - Membrane dynamics within the chick ciliary neuronal growth cone were investigated by using the membrane-impermeant dye FM1-43. A depolarization-evoked endocytosis was observed that shared many properties with the synaptic vesicle recycling previously described at the presynaptic terminal. In addition, in the absence of depolarization a basal level of constitutive endocytotic activity was observed at approximately 30% of the rate of evoked endocytosis. This constitutive endocytosis accounted for large amounts of membrane retrieval: the equivalent of the entire growth cone surface area could be internalized within a 30 min period. Endosomes generated via constitutive and evoked processes were highly mobile and could move considerable distances both within the growth cone and out to the neurite. In addition to their different requirements for formation, evoked and constitutive endosomes displayed a significant difference in release properties. After a subsequent depolarization of labeled growth cones, evoked endosomes were released although constitutive endosomes were not released. Furthermore, treatment with latrotoxin released evoked endosomes, but not constitutive endosomes. Although the properties of evoked endosomes are highly reminiscent of synaptic vesicles, constitutive endosomes appear to be a separate pool resulting from a distinct and highly active process within the neuronal growth cone. PMID- 10531449 TI - Growth factors and taurine protect against excitotoxicity by stabilizing calcium homeostasis and energy metabolism. AB - Taurine, brain derived neurotrophic factor (BDNF), and basic fibroblast growth factor (bFGF) are known to control the development of early postnatal cerebellar granule cells. This study attempted to investigate possible mechanisms of this control by determining neuronal survival, calcium homeostasis, and related calcium-mediated functions, as well as the site of action during glutamate induced excitotoxicity in cultures of cerebellar granule cells. We report that stimulation of glutamate receptors induced a rapid increase in intracellular calcium concentrations ([Ca(2+)](i)) and a decrease in mitochondrial energy metabolism. These effects of glutamate were time- and concentration-dependent and could be specifically blocked by glutamate receptor antagonists. Taurine and bFGF but not BDNF differently regulated [Ca(2+)](i), and preserved the mitochondrial energy metabolism in the presence of glutamate. The regulation of [Ca(2+)](i) by bFGF and taurine required pretreatment of cells with these factors. Confocal microscope analysis of [Ca(2+)](i) and (45)Ca(2+) uptake studies showed that bFGF reduced the magnitude of glutamate-induced calcium uptake with no apparent regulation thereafter. Taurine, on the other hand, did not affect the level of calcium uptake induced by glutamate but rather the duration of the maximal response; this maximal response was transient and returned to basal levels approximately 10 min after glutamate receptor stimulation. We conclude from these data that bFGF and taurine prevent glutamate excitotoxicity through regulation of [Ca(2+)](i) and mitochondrial energy metabolism. Furthermore, the neuroprotective role of taurine and bFGF was enhanced by their collaboration. PMID- 10531448 TI - Differentiation of mammalian vestibular hair cells from conditionally immortal, postnatal supporting cells. AB - We provide evidence from a newly established, conditionally immortal cell line (UB/UE-1) that vestibular supporting cells from the mammalian inner ear can differentiate postnatally into more than one variant of hair cell. A clonal supporting cell line was established from pure utricular sensory epithelia of H2k(b)tsA58 transgenic mice 2 d after birth. Cell proliferation was dependent on conditional expression of the immortalizing gene, the "T" antigen from the SV40 virus. Proliferating cells expressed cytokeratins, and patch-clamp recordings revealed that they all expressed small membrane currents with little time dependence. They stopped dividing within 2 d of being transferred to differentiating conditions, and within a week they formed three defined populations expressing membrane currents characteristic of supporting cells and two kinds of neonatal hair cell. The cells expressed several characteristic features of normal hair cells, including the transcription factor Brn3.1, a functional acetylcholine receptor composed of alpha9 subunits, and the cytoskeletal proteins myosin VI, myosin VIIa, and fimbrin. Immunofluorescence labeling and electron microscopy showed that the cells formed complex cytoskeletal arrays on their upper surfaces with structural features resembling those at the apices of normal hair cells. The cell line UB/UE-1 provides a valuable in vitro preparation in which the expression of numerous structural and physiological components can be initiated or upregulated during early stages of mammalian hair cell commitment and differentiation. PMID- 10531450 TI - The neural cell adhesion molecules L1 and NCAM-180 act in different steps of neurite outgrowth. AB - The formation of neurocircuitry depends on the control of neurite outgrowth that, in turn, can be divided into two processes: nerve growth cone protrusion and neurite extension. It has long been known that the neural cell adhesion molecules L1 and NCAM-180 promote neurite outgrowth, but how they function in growth cones is unclear. We addressed the roles of L1 and NCAM-180 in neurite outgrowth by using microscale chromophore-assisted laser inactivation (micro-CALI) of these proteins to perturb their functions at precise times in single growth cones of embryonic chick dorsal root ganglion neurons grown in culture. Micro-CALI of L1 causes neurite retraction after a 10 min lag period but does not affect growth cone protrusion. In contrast, micro-CALI of NCAM-180 causes rapid growth cone retraction but does not affect neurite extension. The simultaneous inactivation of both these molecules resulted in both distinct effects that were segregated in time. The behavior of growth cones after these micro-CALI treatments resemble the drug-induced perturbation of microtubules for L1 and F-actin for NCAM-180. These findings suggest distinct roles in the growth cone for L1 and NCAM-180 in different steps of neurite outgrowth: L1 functions in neurite extension,whereas NCAM-180 functions in growth cone protrusion. PMID- 10531451 TI - Areas involved in encoding and applying directional expectations to moving objects. AB - Two experiments used functional magnetic resonance imaging (fMRI) to examine the cortical areas involved in establishing an expectation about the direction of motion of an upcoming object and applying that expectation to the analysis of the object. In Experiment 1, subjects saw a stationary cue that either indicated the direction of motion of a subsequent test stimulus (directional cue) or provided no directional information (neutral cue). Their task was to detect the presence of coherent motion in the test stimulus. The stationary directional cue produced larger modulations than the neutral cue, with respect to a passive viewing baseline, both in motion-sensitive areas such as left MT+ and the anterior intraparietal sulcus, as well as motion-insensitive areas such as the posterior intraparietal sulcus and the junction of the left medial precentral sulcus and superior frontal sulcus. Experiment 2 used an event-related fMRI technique to separate signals during the cue period, in which the expectation was encoded and maintained, from signals during the subsequent test period, in which the expectation was applied to the test object. Cue period activations from a stationary, directional cue included many of the same motion-sensitive and insensitive areas from Experiment 1 that produced directionally specific modulations. Prefrontal activations were not observed during the cue period, even though the stationary cue information had to be translated into a format appropriate for influencing motion detection, and this format was maintained for the duration of the cue period (approximately 5 sec). PMID- 10531452 TI - Replay and time compression of recurring spike sequences in the hippocampus. AB - Information in neuronal networks may be represented by the spatiotemporal patterns of spikes. Here we examined the temporal coordination of pyramidal cell spikes in the rat hippocampus during slow-wave sleep. In addition, rats were trained to run in a defined position in space (running wheel) to activate a selected group of pyramidal cells. A template-matching method and a joint probability map method were used for sequence search. Repeating spike sequences in excess of chance occurrence were examined by comparing the number of repeating sequences in the original spike trains and in surrogate trains after Monte Carlo shuffling of the spikes. Four different shuffling procedures were used to control for the population dynamics of hippocampal neurons. Repeating spike sequences in the recorded cell assemblies were present in both the awake and sleeping animal in excess of what might be predicted by random variations. Spike sequences observed during wheel running were "replayed" at a faster timescale during single sharp-wave bursts of slow-wave sleep. We hypothesize that the endogenously expressed spike sequences during sleep reflect reactivation of the circuitry modified by previous experience. Reactivation of acquired sequences may serve to consolidate information. PMID- 10531453 TI - c-fos expression in brainstem premotor interneurons during cholinergically induced active sleep in the cat. AB - The present study was undertaken to identify trigeminal premotor interneurons that become activated during carbachol-induced active sleep (c-AS). Their identification is a critical step in determining the neural circuits responsible for the atonia of active sleep. Accordingly, the retrograde tracer cholera toxin subunit B (CTb) was injected into the trigeminal motor nuclei complex to label trigeminal interneurons. To identify retrograde-labeled activated neurons, immunocytochemical techniques, designed to label the Fos protein, were used. Double-labeled (i.e., CTb(+), Fos(+)) neurons were found exclusively in the ventral portion of the medullary reticular formation, medial to the facial motor nucleus and lateral to the inferior olive. This region, which encompasses the ventral portion of the nucleus reticularis gigantocellularis and the nucleus magnocellularis, corresponds to the rostral portion of the classic inhibitory region of. This region contained a mean of 606 +/- 41.5 ipsilateral and 90 +/- 32.0 contralateral, CTb-labeled neurons. These cells were of medium-size with an average soma diameter of 20-35 micrometer. Approximately 55% of the retrogradely labeled cells expressed c-fos during a prolonged episode of c-AS. We propose that these neurons are the interneurons responsible for the nonreciprocal postsynaptic inhibition of trigeminal motoneurons that occurs during active sleep. PMID- 10531454 TI - Highly specific neuron loss preserves lateral inhibitory circuits in the dentate gyrus of kainate-induced epileptic rats. AB - Patients with temporal lobe epilepsy display neuron loss in the hilus of the dentate gyrus. This has been proposed to be epileptogenic by a variety of different mechanisms. The present study examines the specificity and extent of neuron loss in the dentate gyrus of kainate-treated rats, a model of temporal lobe epilepsy. Kainate-treated rats lose an average of 52% of their GAD-negative hilar neurons (putative mossy cells) and 13% of their GAD-positive cells (GABAergic interneurons) in the dentate gyrus. Interneuron loss is remarkably specific; 83% of the missing GAD-positive neurons are somatostatin immunoreactive. Of the total neuron loss in the hilus, 97% is attributed to two cell types-mossy cells and somatostatinergic interneurons. The retrograde tracer wheat germ agglutinin (WGA)-apoHRP-gold was used to identify neurons with appropriate axon projections for generating lateral inhibition. Previously, it was shown that lateral inhibition between regions separated by 1 mm persists in the dentate gyrus of kainate-treated rats with hilar neuron loss. Retrogradely labeled GABAergic interneurons are found consistently in sections extending 1 mm septotemporally from the tracer injection site in control and kainate-treated rats. Retrogradely labeled putative mossy cells are found up to 4 mm from the injection site, but kainate-treated rats have fewer than controls, and in several kainate-treated rats virtually all of these cells are missing. These findings support hypotheses of temporal lobe epileptogenesis that involve mossy cell and somatostatinergic neuron loss and suggest that lateral inhibition in the dentate gyrus does not require mossy cells but, instead, may be generated directly by GABAergic interneurons. PMID- 10531455 TI - Learning induces a CDC2-related protein kinase, KKIAMRE. AB - To elucidate molecular mechanisms in learning and memory, we analyzed expression of mRNAs in brains of rabbits undergoing eyeblink conditioning. Infusion of the transcription inhibitor actinomycin D into the cerebellar interpositus nucleus reversibly blocked learning but not performance of the conditioned response. Differential display PCR analysis of cerebellar interpositus RNAs from trained and pseudotrained rabbits identified a 207 bp band that was induced with learning. The fragment was used to isolate a cDNA from a lambdagt11 rabbit brain library containing a 1698 bp open reading frame. The deduced amino acid sequence contains the KKIAMRE motif, which is conserved among cell division cycle 2 (cdc2) related kinases. These results suggest that there is a new category of cdc2 related kinases in the brain whose function may be important in learning and memory. PMID- 10531456 TI - Regulation of learning by EphA receptors: a protein targeting study. AB - EphA family receptor tyrosine kinases and their ephrin-A ligands are involved in patterning axonal connections during brain development, but until now a role for these molecules in the mature brain had not been elucidated. Here, we show that both the EphA5 receptor and its ephrin-A ligands (2 and 5) are expressed in the adult mouse hippocampus, and the EphA5 protein is present in a phosphorylated form. Because there are no pharmacological agents available for EphA receptors, we designed recombinant immunoadhesins that specifically bind to the receptor binding site of the ephrin-A ligand (antagonist) or the ligand binding site of the EphA receptor (agonist) and thus target EphA function. We demonstrate that intrahippocampal infusion of an EphA antagonist immunoadhesin leads to impaired performance in two behavioral paradigms, T-maze spontaneous alternation and context-dependent fear conditioning, sensitive to hippocampal function, whereas activation of EphA by infusion of an agonist immunoadhesin results in enhanced performance on these tasks. Because the two behavioral tasks have different motivational, perceptual, and motor requirements, we infer the changes were not caused by these performance factors but rather to cognitive alterations. We also find bidirectional changes in gene expression and in electrophysiological measures of synaptic efficacy that correlate with the behavioral results. Thus, EphA receptors and their ligands are implicated as mediators of plasticity in the adult mammalian brain. PMID- 10531458 TI - The fundamental role of pirouettes in Caenorhabditis elegans chemotaxis. AB - To investigate the behavioral mechanism of chemotaxis in Caenorhabditis elegans, we recorded the instantaneous position, speed, and turning rate of single worms as a function of time during chemotaxis in gradients of the attractants ammonium chloride or biotin. Analysis of turning rate showed that each worm track could be divided into periods of smooth swimming (runs) and periods of frequent turning (pirouettes). The initiation of pirouettes was correlated with the rate of change of concentration (dC/dt) but not with absolute concentration. Pirouettes were most likely to occur when a worm was heading down the gradient (dC/dt < 0) and least likely to occur when a worm was heading up the gradient (dC/dt > 0). Further analysis revealed that the average direction of movement after a pirouette was up the gradient. These observations suggest that chemotaxis is produced by a series of pirouettes that reorient the animal to the gradient. We tested this idea by imposing the correlation between pirouettes and dC/dt on a stochastic point model of worm motion. The model exhibited chemotaxis behavior in a radial gradient and also in a novel planar gradient. Thus, the pirouette model of C. elegans chemotaxis is sufficient and general. PMID- 10531457 TI - Dopamine D4 receptor-knock-out mice exhibit reduced exploration of novel stimuli. AB - The involvement of dopamine neurotransmission in behavioral responses to novelty is suggested by reports that reward is related to increased dopamine activity, that dopamine modulates exploratory behavior in animals, and that Parkinson's disease patients report diminished responses to novelty. Some studies have reported that polymorphisms of the human dopamine D4 receptor (D4R) gene are associated with personality inventory measures of the trait called "novelty seeking". To explore a potential role for the D4R in behavioral responses to novelty, we evaluated D4R-knock-out (D4R-/-) and wild-type (D4R+/+) mice in three approach-avoidance paradigms: the open field, emergence, and novel object tests. These three paradigms differ in the degree to which they elicit approach, or exploratory behavior, and avoidance, or anxiety-related behavior. Thus, we used these three tests to determine whether the D4R primarily influences the exploratory or the anxious component of responses to approach-avoidance conflicts. D4R-/- mice were significantly less behaviorally responsive to novelty than D4R+/+ mice in all three tests. The largest phenotypic differences were observed in the novel object test, which maximizes approach behavior, and the smallest phenotypic differences were found in the open field test, which maximizes avoidance behavior. Hence, D4R-/- mice exhibit reductions in behavioral responses to novelty, reflecting a decrease in novelty-related exploration. PMID- 10531459 TI - Auditory thalamus, dorsal hippocampus, basolateral amygdala, and perirhinal cortex role in the consolidation of conditioned freezing to context and to acoustic conditioned stimulus in the rat. AB - On the basis of previous experimental evidence, it is known that the auditory thalamus (AT), the dorsal hippocampus (DH), the basolateral amygdala (BLA), and the perirhinal cortex (PC) are involved in the mnemonic processing of conditioned freezing. In particular, BLA and PC appear to be involved both in conditioned stimulus (CS) and context conditioned freezing. Through AT, the auditory CS is sent to other sites, whereas DH is involved in context conditioning. Nevertheless, the existing evidence does not make it possible to assess AT, DH, BLA, and PC involvement during the consolidation phase of conditioned freezing. To address this question, fully reversible tetrodotoxin (TTX) inactivation was performed on adult male Wistar rats having undergone CS and context fear training. Anesthetized animals were injected stereotaxically with TTX (either 5 or 10 ng in 0.5 or 1.0 microliter of saline, according to site dimensions) at increasing post-acquisition delays. Context and CS freezing durations were measured during retention testing, always performed 48 and 72 hr after TTX administration. The results showed that AT inactivation does not disrupt consolidation of either contextual or auditory fear memories. In contrast, inactivation of the other three structures disrupted consolidation. For the DH, this disruption was specific to contextual cues and only occurred when inactivation was performed early (up to 1.5 hr) after training. The BLA and PC were shown to be involved in the consolidation of both contextual and auditory fear. Their involvement persisted for longer periods of time (2d for BLA and 8 d for PC). These findings provide information to build a temporal profile for the post-training processing of fear memories in structures known to be important for this form of learning. The results are discussed in relation to previous studies on conditioned freezing and other aversive conditioned response neural correlates. PMID- 10531460 TI - A single exposure to amphetamine is sufficient to induce long-term behavioral, neuroendocrine, and neurochemical sensitization in rats. AB - Repeated treatment with psychostimulant drugs causes long-lasting behavioral sensitization and associated neuroadaptations. Although sensitization induced by a single psychostimulant exposure has also been reported, information on the behavioral and neurochemical consequences of a single psychostimulant exposure is sparse. Therefore, to evaluate whether behavioral sensitization evoked by single and repeated psychostimulant pretreatment regimens represent the same neurobiological phenomenon, the time-dependent expression of behavioral, neurochemical, and neuroendocrine sensitization after a single exposure to amphetamine was investigated in rats. A single exposure to amphetamine (5 mg/kg, i.p.) caused context-independent sensitization of the locomotor effects of amphetamine, which intensified over time. Thus, sensitization to amphetamine was marginal at 3 d after treatment and more evident after 1 week, whereas 3 weeks after treatment, profound sensitization, as well as cross-sensitization, to cocaine was observed. Amphetamine pretreatment caused an increase in the electrically evoked release of [(3)H]dopamine from nucleus accumbens, caudate putamen, and medial prefrontal cortex slices and of [(14)C]acetylcholine from accumbens and caudate slices. The hyperreactivity of dopaminergic nerve terminals appeared to parallel the development of locomotor sensitization, i.e., whereas hyperreactivity of accumbens dopaminergic terminals increased between 3 d and 3 weeks after treatment, the hyperreactivity of medial prefrontal dopaminergic terminals decreased. Pre-exposure to amphetamine also sensitized the hypothalamus pituitary-adrenal axis response to amphetamine at 1 and 3 weeks, but not at 3 d after treatment. Because these data closely resemble those reported previously for repeated amphetamine pretreatment, it is concluded that a single exposure to amphetamine is sufficient to induce long-term behavioral, neurochemical, and neuroendocrine sensitization in rats. PMID- 10531461 TI - Synaptic basis of cortical persistent activity: the importance of NMDA receptors to working memory. AB - Delay-period activity of prefrontal cortical cells, the neural hallmark of working memory, is generally assumed to be sustained by reverberating synaptic excitation in the prefrontal cortical circuit. Previous model studies of working memory emphasized the high efficacy of recurrent synapses, but did not investigate the role of temporal synaptic dynamics. In this theoretical work, I show that biophysical properties of cortical synaptic transmission are important to the generation and stabilization of a network persistent state. This is especially the case when negative feedback mechanisms (such as spike-frequency adaptation, feedback shunting inhibition, and short-term depression of recurrent excitatory synapses) are included so that the neural firing rates are controlled within a physiological range (10-50 Hz), in spite of the exuberant recurrent excitation. Moreover, it is found that, to achieve a stable persistent state, recurrent excitatory synapses must be dominated by a slow component. If neuronal firings are asynchronous, the synaptic decay time constant needs to be comparable to that of the negative feedback; whereas in the case of partially synchronous dynamics, it needs to be comparable to a typical interspike interval (or oscillation period). Slow synaptic current kinetics also leads to the saturation of synaptic drive at high firing frequencies that contributes to rate control in a persistent state. For these reasons the slow NMDA receptor-mediated synaptic transmission is likely required for sustaining persistent network activity at low firing rates. This result suggests a critical role of the NMDA receptor channels in normal working memory function of the prefrontal cortex. PMID- 10531462 TI - Metabotropic glutamate receptor-mediated hippocampal phosphoinositide turnover is blunted in spatial learning-impaired aged rats. AB - Maximal phosphoinositide (PI) turnover was examined in the hippocampus of young and aged Long-Evans rats that were behaviorally characterized for spatial learning in the Morris water maze. The type 1 metabotropic glutamate receptor (mGluR) agonist 1S,3R ACPD was used to stimulate PI turnover and to determine the E(MAX) for each rat. Protein levels in hippocampus for type 1 mGluRs, Galphaq11, and phospholipase Cbeta-1 (PLCbeta-1) were also measured by quantitative Western blotting. The results show that PI turnover mediated by the mGluRs was blunted in the aged rats. The magnitude of the decrement in PI turnover was also significantly correlated with age-related spatial memory decline. The decrease in mGluR-mediated PI turnover occurred without changes in the protein level of either the mGluRs or the G-protein coupled to those receptors, Galphaq11. A significant decrease in the immunoreactivity of PLCbeta-1, however, was observed in the hippocampus of aged rats; PLCbeta-1 immunoreactivity was significantly correlated with spatial learning only when the young and aged rats were considered together. The decrement in mGluR-mediated signal transduction in the hippocampus that is related to cognitive impairment in aging may be attributable, at least in part, to a deficiency in the enzyme PLCbeta-1. That deficiency may also contribute to a blunted response in muscarinic stimulation of hippocampal PI turnover that we previously found in this same study population. An age-related alteration in this signal transduction system may provide a functional basis for cognitive decline independent of any loss of neurons in the hippocampus. PMID- 10531463 TI - Anterograde and retrograde amnesia after lesions to frontal cortex in rats. AB - A socially acquired food-preference test was used to assess effects of lesions to the frontal cortex on anterograde and retrograde memory in rats. In Experiment 1, there was no effect of lesion when rats were administered a two-choice test in which the target food was to be selected in the presence of a single distractor. In Experiment 2, a three-choice memory test was administered in which the target food was presented along with two equally palatable alternatives. In the latter test, lesioned groups displayed anterograde amnesia that increased with the length of the interval between postoperative acquisition and test, and a severe retrograde amnesia that extended equally over the entire range of intervals between preoperative acquisition and test. This outcome, which contrasted with the pattern of memory loss previously observed in rats with hippocampal lesions on this test, was interpreted as evidence for the strategic role of the frontal lobes in directing response selection and retrieval processes in memory. PMID- 10531464 TI - The Aplysia mytilus inhibitory peptide-related peptides: identification, cloning, processing, distribution, and action. AB - Neuropeptides are a ubiquitous class of signaling molecules. In our attempt to understand the generation of feeding behavior in Aplysia, we have sought to identify and fully characterize the neuropeptides operating in this system. Preliminary evidence indicated that Mytilus inhibitory peptide (MIP)-like peptides are present and operating in the circuitry that generates feeding in Aplysia. MIPs were originally isolated from the bivalve mollusc Mytilus edulis, and related peptides have been identified in other invertebrate species, but no precursor has been identified. In this study, we describe the isolation and characterization of novel Aplysia MIP-related peptides (AMRPs) and their precursor. Several AMRPs appear to have some structural and functional features similar to vertebrate opioid peptides. We use matrix-assisted laser desorption/ionization time-of-flight mass spectrometry to confirm that all 14 AMRPs predicted by the precursor are processed in isolated neurons. Northern analysis, whole-mount in situ hybridization, and immunohistochemistry are used to map the abundant expression of these peptides in the CNS and peripheral tissues such as the digestive tract, vasculature, and the reproductive organs. Physiological studies demonstrate that the rank order of the inhibitory actions of these peptides is different for three target muscles. These results underscore the importance of using a multidisciplinary approach to identifying and characterizing the actions of neuropeptides in an effort to gain understanding of their role in systems of interest. The widespread distribution of the AMRPs indicates that they may be operating in many different systems of Aplysia. PMID- 10531465 TI - Amygdala neurons mediate acquisition but not maintenance of instrumental avoidance behavior in rabbits. AB - Whereas the amygdala is generally understood to be involved in aversively motivated learning, the specific associative function of the amygdala remains controversial. This study addressed the amygdalar role in mediation of discriminative instrumental avoidance learning of rabbits. Bilateral microinjection of the GABA receptor agonist muscimol centered in the basolateral nucleus of the amygdala was given to inactivate amygdalar neurons at each of three stages of acquisition. The absence of behavioral learning in rabbits trained immediately after amygdalar inactivation confirmed previous results with electrolytic lesions. The absence of savings during training after muscimol had become ineffective indicated an amygdalar role in the establishment of acquisition-relevant neural plasticity, not simply in the expression of the learned response. A time-limited role of the amygdala in instrumental avoidance learning was indicated by the finding that intra-amygdalar muscimol failed to disrupt performance of the well-established avoidance response. The passage of time alone (with no training trials) was sufficient to reduce amygdalar involvement in response performance. These results and demonstrations that other limbic system areas make time-limited contributions to learning indicate that the amygdala is part of a larger intermediate memory system that supports learning and performance before habit consolidation. PMID- 10531466 TI - Spinal opioid analgesia: how critical is the regulation of substance P signaling? AB - Although opioids can reduce stimulus-evoked efflux of Substance P (SP) from nociceptive primary afferents, the consequences of this reduction on spinal cord nociceptive processing has not been studied. Rather than assaying SP release, in the present study we examined the effect of opioids on two postsynaptic measures of SP release, Fos expression and neurokinin-1 (NK-1) receptor internalization, in the rat. The functional significance of the latter was first established in in vitro studies that showed that SP-induced Ca(2+) mobilization is highly correlated with the magnitude of SP-induced NK-1 receptor internalization in dorsal horn neurons. Using an in vivo analysis, we found that morphine had little effect on noxious stimulus-evoked internalization of the NK-1 receptor in lamina I neurons. However, internalization was reduced when we coadministered morphine with a dose of an NK-1 receptor antagonist that by itself was without effect. Thus, although opioids may modulate SP release, the residual release is sufficient to exert maximal effects on the target NK-1 receptors. Morphine significantly reduced noxious stimulus-induced Fos expression in lamina I, but the Fos inhibition was less pronounced in neurons that expressed the NK-1 receptor. Taken together, these results suggest that opioid analgesia predominantly involves postsynaptic inhibitory mechanisms and/or presynaptic control of non-SP-containing primary afferent nociceptors. PMID- 10531467 TI - Involvement of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and p53 in neuronal apoptosis: evidence that GAPDH is upregulated by p53. AB - We recently reported that cytosine arabinoside (AraC)-induced apoptosis of cerebellar neurons involves the overexpression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The present study was undertaken to investigate whether p53 and/or Bax overexpression participates in the AraC-induced apoptosis of cerebellar granule cells and, if so, the relationship between p53 induction and GAPDH overexpression in these cells. AraC-induced apoptosis of cerebellar granule cells was preceded by an increase in levels of p53 mRNA and protein detected between 1 and 8 hr after treatment. The mRNA level for a p53 target gene, Bax, was also increased. The increase in GAPDH mRNA lasted longer than that of either p53 or Bax, and the level of GAPDH protein in the particulate fraction increased after induction of GAPDH mRNA. The antisense oligonucleotide to p53 protected granule cells from AraC-induced chromatin condensation, internucleosomal cleavage, and apoptotic death. The inhibition of p53 expression by the p53 antisense oligonucleotide not only blocked the expression of Bax but also partially suppressed the increased GAPDH mRNA and protein levels. Conversely, the suppression of GAPDH expression and subsequent attenuation of apoptosis of granule cells by GAPDH antisense oligonucleotide did not influence the expression of p53 or Bax. Cerebellar granule cells prepared from p53 knock-out mice were resistant to AraC toxicity, and the p53 gene knock-out suppressed AraC upregulated GAPDH expression. Moreover, infection of PC12 cells with an adenoviral vector containing p53 gene dramatically increased GAPDH expression and triggered cell apoptosis. These results suggest that AraC-induced apoptosis of cerebellar granule cells involves the expression of both GAPDH and p53 and that, similar to Bax, GAPDH is upregulated by p53 after exposure to the apoptotic insult. PMID- 10531468 TI - Glutamate transporters contribute to the time course of synaptic transmission in cerebellar granule cells. AB - Transporters are thought to assist in the termination of synaptic transmission at some synapses by removing neurotransmitter from the synapse. To investigate the role of glutamate transport in shaping the time course of excitatory transmission at the mossy fiber-granule cell synapse, the effects of transport impairment were studied using whole-cell voltage- and current-clamp recordings in slices of rat cerebellum. Impairment of transport by L-trans-pyrrolidine-2,4-dicarboxylate (PDC) produced a prolongation of the decay of the AMPA receptor-mediated current after a repetitive stimulus, as well as prolongation of single stimulus-evoked EPSCs when AMPA receptor desensitization was blocked. PDC also produced a prolongation of both single and repetitive-evoked NMDA receptor-mediated EPSCs. Enzymatic degradation of extracellular glutamate did not reverse the PDC-induced prolongation of AMPA receptor-mediated current after a repetitive stimulus, suggesting that transporter binding sites participate in limiting glutamate spillover. In current-clamp recordings, PDC dramatically increased the total area of the EPSP and the burst duration evoked by single and repetitive stimuli. These data indicate that glutamate transporters play a significant role in sculpting the time course of synaptic transmission at granule cell synapses, most likely by limiting the extent of glutamate spillover. The contribution of transporters is particularly striking during repetitive stimulus trains at physiologically relevant frequencies. Hence, the structural arrangement of the glomerulus may enhance the contribution of transporters to information processing by limiting the extent of glutamate spillover between adjacent synapses. PMID- 10531469 TI - Coding of sound pressure level in the barn owl's auditory nerve. AB - Rate-intensity functions, i.e., the relation between discharge rate and sound pressure level, were recorded from single auditory nerve fibers in the barn owl. Differences in sound pressure level between the owl's two ears are known to be an important cue in sound localization. One objective was therefore to quantify the discharge rates of auditory nerve fibers, as a basis for higher-order processing of sound pressure level. The second aim was to investigate the rate-intensity functions for cues to the underlying cochlear mechanisms, using a model developed in mammals. Rate-intensity functions at the most sensitive frequency mostly showed a well-defined breakpoint between an initial steep segment and a progressively flattening segment. This shape has, in mammals, been convincingly traced to a compressive nonlinearity in the cochlear mechanics, which in turn is a reflection of the cochlear amplifier enhancing low-level stimuli. The similarity of the rate-intensity functions of the barn owl is thus further evidence for a similar mechanism in birds. An interesting difference from mammalian data was that this compressive nonlinearity was not shared among fibers of similar characteristic frequency, suggesting a different mechanism with a more locally differentiated operation than in mammals. In all fibers, the steepest change in discharge rate with rising sound pressure level occurred within 10-20 dB of their respective thresholds. Because the range of neural thresholds at any one characteristic frequency is small in the owl, auditory nerve fibers were collectively most sensitive for changes in sound pressure level within approximately 30 dB of the best thresholds. Fibers most sensitive to high frequencies (>6-7 kHz) showed a smaller increase of rate above spontaneous discharge rate than did lower-frequency fibers. PMID- 10531470 TI - Molecular mechanisms of long-term potentiation in the insular cortex in vivo. AB - We have investigated molecular mechanisms of synaptic plasticity in the pathway between two forebrain structures important for taste learning, the basolateral amygdala (BLA) and the insular cortex. We report here that in vivo long-term potentiation (LTP) induced by BLA stimulation requires functional NMDA receptors and is modulated by muscarinic acetylcholine receptors. In addition, LTP results in the activation of cortical extracellular regulated kinase 1/2 (ERK1/2) and is blocked by inhibitors of ERK1/2 activation. Previous findings demonstrated the involvement of the same molecular mechanisms in the same cortical area during novel taste learning. The results demonstrate that both synaptic and behavioral plasticity share common molecular mechanisms in the insular cortex. PMID- 10531471 TI - Src potentiation of NMDA receptors in hippocampal and spinal neurons is not mediated by reducing zinc inhibition. AB - The protein-tyrosine kinase Src is known to potentiate the function of NMDA receptors, which is necessary for the induction of long-term potentiation in the hippocampus. With recombinant receptors composed of NR1-1a/NR2A or NR1-1a/2B subunits, Src reduces voltage-independent inhibition by the divalent cation Zn2+. Thereby the function of recombinant NMDA receptors is potentiated by Src only when the Zn2+ level is sufficient to cause tonic inhibition. Here we investigated whether the Src-induced potentiation of NMDA receptor function in neurons is caused by reducing voltage-independent Zn2+ inhibition. Whereas chelating extracellular Zn2+ blocked the Src-induced potentiation of NR1-1a/2A receptors, we found that Zn2+ chelation did not affect the potentiation of NMDA receptor (NMDAR) currents by Src applied into hippocampal CA1 or CA3 neurons. Moreover, Src did not alter the Zn2+ concentration-inhibition relationship for NMDAR currents in CA1 or CA3 neurons. Also, chelating extracellular Zn2+ did not prevent the upregulation of NMDA single-channel activity by endogenous Src in membrane patches from spinal dorsal horn neurons. Taking these results together we conclude that Src-induced potentiation of NMDAR currents is not mediated by reducing Zn2+ inhibition in hippocampal and dorsal horn neurons. PMID- 10531472 TI - Ab initio solution and refinement of two high-potential iron protein structures at atomic resolution. AB - The crystal structure of the reduced high-potential iron protein (HiPIP) from Chromatium vinosum has been redetermined in a new orthorhombic crystal modification, and the structure of its H42Q mutant has been determined in orthorhombic (H42Q-1) and cubic (H42Q-2) modifications. The first two were solved by ab initio direct methods using data collected to atomic resolution (1.20 and 0. 93 A, respectively). The recombinant wild type (rc-WT) with two HiPIP molecules in the asymmetric unit has 1264 protein atoms and 335 solvent sites, and is the second largest structure reported so far that has been solved by pure direct methods. The solutions were obtained in a fully automated way and included more than 80% of the protein atoms. Restrained anisotropic refinement for rc-WT and H42Q-1 converged to R(1) = summation operator||F(o)| - |F(c)|| / summation operator|F(o)| of 12.0 and 13.6%, respectively [data with I > 2sigma(I)], and 12.8 and 15.5% (all data). H42Q-2 contains two molecules in the asymmetric unit and diffracted only to 2.6 A. In both molecules of rc-WT and in the single unique molecule of H42Q-1 the [Fe(4)S(4)](2+) cluster dimensions are very similar and show a characteristic tetragonal distortion with four short Fe-S bonds along four approximately parallel cube edges, and eight long Fe-S bonds. The unique protein molecules in H42Q-2 and rc-WT are also very similar in other respects, except for the hydrogen bonding around the mutated residue that is at the surface of the protein, supporting the hypothesis that the difference in redox potentials at lower pH values is caused primarily by differences in the charge distribution near the surface of the protein rather than by structural differences in the cluster region. PMID- 10531473 TI - Structure of the RWJ-51084-bovine pancreatic beta-trypsin complex at 1.8 A. AB - The three-dimensional structure of bovine pancreatic trypsin complexed with the inhibitor RWJ-51084 has been determined at 1.8 A resolution. These crystals belong to the trigonal space group P3(1)21, with unit-cell parameters a = b = 53.43, c = 107.76 A. The refined R and R(free) values are 0.175 and 0.237, respectively. The carbonyl group bonded to the benzothiazole group of the inhibitor is covalently linked to the hydroxyl O atom of Ser195, forming a tetrahedral intermediate hemiketal structure. The other carbonyl O atom of the inhibitor forms a hydrogen bond with the Gln192 side-chain amide group. The benzothiazole group is oriented with the aromatic N atom of RWJ-51084 accepting a hydrogen bond from His57 NE2. The arginine side chain of the inhibitor extends into the deep and narrow pocket of the S1 specificity site of trypsin, forming a network of hydrogen bonds. PMID- 10531474 TI - Structure of oxalate-substituted diferric mare lactoferrin at 2.7 A resolution. AB - Lactoferrin binds two Fe(3+) and two CO(2-)(3) ions with high affinity. It can also bind other metal ions and anions. In order to determine the perturbations in the environments of the binding sites in the N and C lobes and elsewhere in the protein, the crystal structure of oxalate-substituted diferric mare lactoferrin has been determined at 2.7 A resolution. The final model has a crystallographic R factor of 21.3% for all data in the resolution range 17.0-2.7 A. The substitution of an oxalate anion does not perturb the overall structure of the protein, but produces several significant changes at the metal-binding and anion-binding sites. The binding of the oxalate anion is symmetrical in both the N and C lobes, unlike in diferric dioxalate human lactoferrin, where the oxalate anion binds the metal ion symmetrically in the C lobe and asymmetrically in the N lobe. PMID- 10531475 TI - Lactoferrin-metal interactions: first crystal structure of a complex of lactoferrin with a lanthanide ion (Sm3+) at 3.4 A resolution. AB - Lactoferrin is an important member of the transferrin family. A characteristic property of transferrins is their ability to bind very tightly (K(app) approximately/= 10(20)) but reversibly two Fe(3+) ions. The structural consequences of binding a metal other than Fe(3+) have been examined by crystallographic analysis at 3.4 A resolution of mare samarium-lactoferrin (Sm(2)Lf). The structure was refined to an R factor of 0.219 for 8776 reflections in the resolution range 17.0-3.4 A. The samarium geometry (distorted octahedral coordination) is similar in both lobes. However, the anion interactions are quite different in the two lobes. In the N lobe, the anion is able to form only two hydrogen bonds instead of the four observed in the C lobe of Sm(2)Lf and the six observed in Fe(2)Lf. This is because Arg121, Thr117 and Gly124 have moved away from the anion as a consequence of the binding of the Sm(3+) ion. The protein ligands in the binding cleft of Sm(2)Lf show large displacements, but the overall protein structure remains the same. The binding of Sm(3+) by lactoferrin shows that the protein is capable of sequestering ions of different sizes and charges, though with reduced affinity. This conclusion should be true of other transferrins also. PMID- 10531476 TI - Structure of buffalo lactoferrin at 2.5 A resolution using crystals grown at 303 K shows different orientations of the N and C lobes. AB - The structure of buffalo lactoferrin has been determined at 303 K. The crystals belong to orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 77.5, b = 91.0, c = 131.5 A and Z = 4. The structure has been refined to an R factor of 0.187. The overall structure of the protein is similar to its structure determined at 277 K in a different crystal form. However, the lobe orientations in the two structures differ by 9.0 degrees, suggesting significant inter-lobe flexibility in this family of proteins. The inter-lobe interactions are predominantly hydrophobic and could act as a cushion for a change in orientation under the influence of external conditions. On the other hand, the domain arrangements are found to be similar in 277 and 303 K crystal structures, with orientations differing by 1.5 and 1.0 degrees in the N and C lobes, respectively. The results of these investigations suggest that the increase in temperature helps in the production of better quality crystals. PMID- 10531477 TI - Cryocrystallography of a Kunitz-type serine protease inhibitor: the 90 K structure of winged bean chymotrypsin inhibitor (WCI) at 2.13 A resolution. AB - The crystal structure of a Kunitz-type double-headed alpha--chymotrypsin inhibitor from winged bean seeds has been refined at 2.13 A resolution using data collected from cryo-cooled (90 K) crystals which belong to the hexagonal space group P6(1)22 with unit-cell parameters a = b = 60.84, c = 207.91 A. The volume of the unit cell is reduced by 5.3% on cooling. The refinement converged to an R value of 20.0% (R(free) = 25.8%) for 11100 unique reflections and the model shows good stereochemistry, with r.m.s. deviations from ideal values for bond lengths and bond angles of 0.011 A and 1.4 degrees, respectively. The structural architecture of the protein consists of 12 antiparallel beta-strands joined in the form of a characteristic beta-trefoil fold, with the two reactive-site regions, Asn38-Leu43 and Gln63-Phe68, situated on two external loops. Although the overall protein fold is the same as that of the room-temperature model, some conformational changes are observed in the loop regions and in the side chains of a few surface residues. A total of 176 ordered water molecules and five sulfate ions are included in the model. PMID- 10531478 TI - Structure of a phosphoglycerate mutase:3-phosphoglyceric acid complex at 1.7 A. AB - The crystal structure of the tetrameric glycolytic enzyme phosphoglycerate mutase from the yeast Saccharomyces cerevisiae has been determined to 1.7 A resolution in complex with the sugar substrate. The difference map indicates that 3 phosphoglycerate is bound at the base of a 12 A cleft, positioning C2 of the substrate within 3.5 A of the primary catalytic residue, histidine 8. PMID- 10531479 TI - Structure of ribosomal protein L30 from Thermus thermophilus at 1.9 A resolution: conformational flexibility of the molecule. AB - The crystal structure of ribosomal protein L30 from the extreme thermophilic bacterium Thermus thermophilus has been determined at 1. 9 A resolution. The crystals are trigonal and belong to space group P3(2)21, with unit-cell parameters a = b = 63.5, c = 77.8 A, alpha = beta = 90, gamma = 120 degrees and two molecules per asymmetric unit. The structure was solved by the molecular replacement method with AMoRe and refined with X-PLOR to an R value of 20.3% and an R(free) of 25.3% in the resolution range 8-1.9 A. Detailed analyses of the structures of the two molecules in the asymmetric unit and comparison of T. thermophilus L30 structure with the structure of homologous L30 from Bacillus stearothermophilus reveal two flexible regions at opposite ends of the rather elongated molecule. Such flexibility could be important for the protein fitting in the ribosome. A comparison with B. stearothermophilus L30 shows a higher number of salt bridges and unbound positively charged residues and an increased accessible hydrophobic area on the surface of T. thermophilus L30. This could contribute to the stability of both the extreme thermophile protein and the ribosome as a whole. PMID- 10531480 TI - Structure of acutolysin-C, a haemorrhagic toxin from the venom of Agkistrodon acutus, providing further evidence for the mechanism of the pH-dependent proteolytic reaction of zinc metalloproteinases. AB - The structure of acutolysin-C, a haemorrhagic zinc metalloproteinase from the venom of Agkistrodon acutus, has been analyzed and refined at 2.2 A resolution. The space group of the crystal is P2(1)2(1)2(1), with unit-cell dimensions a = 46.84, b = 49.52, c = 95.34 A. One molecule was found in each asymmetric unit. The phasing problem was solved by the molecular-replacement program AMoRe. Crystallographic refinement was performed using X-PLOR, leading to final R and free R factors of 0.176 and 0.272, respectively. The residue sequence of acutolysin-C was determined mainly by electron density. No density was found for the first residue at the N--terminus and the last two residues at the C-terminus, which was also the case for most other P-I class snake-venom metalloproteinases (SVMPs). Acutolysin-C has two highly conserved characteristic sequences His142 Glu143-X-X-His146-X-X-Gly149-X-X-His15 2 and Cys162-Ile163-Met164. The enzyme has three disulfide bridges: Cys117-Cys195, Cys157-Cys179 and Cys159-Cys162. The entire structure shows good agreement with that of other reported P-I class SVMPs and has two subdomains with a cleft in which one catalytic zinc ion is localized. However, the local conformation (especially the disulfide configurations), the coordination of the catalytic water molecules and some residue side chains differ compared with other P-I class SVMPs. The proteolytic activities of SVMPs are sensitive to the pH value. The molecular superpositions around the proteolytic active sites of all the P-I class SVMP crystal structures show that the distances between the zinc ion and its ligands are not correlated with the crystallization pH values, although the contact distances between the catalytic water molecule and the O atoms of the Glu143 carboxylate group in the neutral and weakly alkaline structures are shorter than those in weakly acidic structures, and the closer the crystallization pH value of one enzyme is to its optimal activity pH value, the shorter the contact distances. Overall, all P-I class SVMPs have similar conformations in the active-site cleft. The size of the active site is not correlated with the crystallization pH values or the proteolytic activities. The disulfide bridge Cys117-Cys195 is conserved in all crystal structures of P-I class SVMPs, whereas the conformation and number of disulfide bridges in the C terminal subdomain differ. Acutolysin-C has no structural calcium ion, which may not affect the proteolytic activity or haemorrhagic activity directly. PMID- 10531481 TI - Refined structure of the histidine-containing phosphotransfer (HPt) domain of the anaerobic sensor kinase ArcB from Escherichia coli at 1.57 A resolution. AB - The crystal structure of the histidine-containing phosphotransfer (HPt) domain of the anaerobic sensor kinase ArcB from Escherichia coli has been refined to 1.57 A resolution, using the coordinates of the earlier 2.06 A structure as a starting model. The final model contained 956 protein atoms, one zinc ion and 156 water molecules, with an R factor of 19.0%. The high-resolution electron-density maps clearly revealed additional solvent molecules and seven discrete rotamers in the protein side chains. One residue, Met755, was fully buried but was able to occupy the space in the hydrophobic core by means of the two-state conformation of its side chain. One water molecule was buried in the protein core and contributed to the rigidity of the HPt domain, cooperating in the coordination of the zinc ion. PMID- 10531482 TI - Structures of cellular retinoic acid binding proteins I and II in complex with synthetic retinoids. AB - Retinoids play important roles in diverse cellular processes including growth, cell differentiation and vision. Many natural and synthetic retinoids are used as drugs in dermatology and oncology. A large amount of data has been accumulated on the cellular activity of different synthetic retinoids. They are stabilized and transported inside the cell cytoplasm by binding and transport proteins, such as cellular retinol-binding proteins and cellular retinoic acid binding proteins (CRABPs). The structures of human CRABP II in complex with two different synthetic retinoids, Ro13-6307 and Ro12--7310 (at 2.1 and 2.0 A resolution, respectively) and of bovine CRABP I in complex with a retinobenzoic acid, Am80 (at 2.8 A resolution) are described. The binding affinities of human CRABP I and II for the retinoids studied here have been determined. All these compounds have comparable binding affinities (nanomolar range) for both CRABPs. Apart from the particular interactions of the carboxylate group of the retinoids with specific protein groups, each structure reveals characteristic interactions. Studying the atomic details of the interaction of retinoids with retinoid-binding proteins facilitates the understanding of the kinetics of retinoid trafficking inside the cytoplasm. PMID- 10531483 TI - Dehydration leads to a phase transition in monoclinic factor XIII crystals. AB - Monoclinic factor XIII crystals have been transferred to a solution containing increasing amounts of the precipitant PEG 6000. At a concentration of about 36%(w/v) PEG 6000, a phase transition was observed. The space group of the crystals was preserved on the transition, but half of the 2(1) screw axes were lost, which meant that the unit-cell volume and the content of the asymmetric unit were doubled. The structure of factor XIII in the new crystal form was solved by molecular replacement. About 80% of the changes accompanying the transition can be explained by a rigid-body rotation of half of the factor XIII dimers in the lattice by about 5 degrees. The remaining changes are mostly small interdomain movements of the four domains which constitute one factor XIII monomer. PMID- 10531484 TI - Reciprocal-space solvent flattening. AB - Solvent flattening is a powerful tool for improving crystallographic phases for macromolecular structures obtained at moderate resolution, but uncertainties in the optimal weighting of experimental phases and modified phases make it difficult to extract all the phase information possible. Solvent flattening is essentially an iterative method for maximizing a likelihood function which consists of (i) experimental phase information and (ii) information on the likelihood of various arrangements of electron density in a map, but the likelihood function is generally not explicitly defined. In this work, a procedure is described for reciprocal-space maximization of a likelihood function based on experimental phases and characteristics of the electron-density map. The procedure can readily be applied to phase improvement based on solvent flattening and can potentially incorporate information on a wide variety of other characteristics of the electron-density map. PMID- 10531485 TI - Evaluation of macromolecular electron-density map quality using the correlation of local r.m.s. density. AB - It has recently been shown that the standard deviation of local r.m. s. electron density is a good indicator of the presence of distinct regions of solvent and protein in macromolecular electron-density maps [Terwilliger & Berendzen (1999). Acta Cryst. D55, 501-505]. Here, it is demonstrated that a complementary measure, the correlation of local r.m.s. density in adjacent regions on the unit cell, is also a good measure of the presence of distinct solvent and protein regions. The correlation of local r.m.s. density is essentially a measure of how contiguous the solvent (and protein) regions are in the electron-density map. This statistic can be calculated in real space or in reciprocal space and has potential uses in evaluation of heavy-atom solutions in the MIR and MAD methods as well as for evaluation of trial phase sets in ab initio phasing procedures. PMID- 10531486 TI - Experimental assessment of differences between related protein crystal structures. AB - Prior to attaching any biological significance to differences between two related protein crystal structures, it must be established that such differences are genuine, rather than artefacts of the structure-determination protocol. This will be all the more important as more and more related protein structures are solved and comparative structural biology attempts to correlate structural differences with variations in biological function, activity or affinity. A method has been developed which enables unbiased assessment of differences between the structures of related biomacromolecules using experimental crystallographic information alone. It is based on the use of local density-correlation maps, which contain information regarding the similarity of the experimental electron density for corresponding parts of different copies of a molecule. The method can be used to assess a priori which parts of two or more molecules are likely to be structurally similar; this information can then be employed during structure refinement. Alternatively, the method can be used a posteriori to verify that differences observed in two or more models are supported by the experimental information. Several examples are discussed which validate the notion that local conformational variability is highly correlated to differences in the local experimental electron density. PMID- 10531487 TI - Crystallization of the 10-23 DNA enzyme using a combinatorial screen of paired oligonucleotides. AB - One of the most difficult steps in the X-ray crystallography of nucleic acids is obtaining crystals that diffract to high resolution. The choice of the nucleotide sequence has proven to be more important in producing high-quality crystals than the composition of the crystallization solution. This manuscript describes a systematic procedure for identifying the optimal sizes of a multi-stranded nucleic acid complex which provide high-quality crystals. This approach was used to crystallize the in vitro evolved 10-23 DNA enzyme complexed with its RNA substrate. In less than two months, 81 different enzyme-substrate complexes were generated by combinatorial mixing and annealing of complementary oligonucleotides which differed in length, resulting in duplexes of varying length, with or without nucleotide overhangs. Each of these complexes was screened against a standard set of 48 crystallization conditions and evaluated for crystal formation. The screen resulted in over 40 crystal forms, the best of which diffracted to 2.8 A resolution when exposed to a synchrotron X-ray source. PMID- 10531488 TI - A two-dimensional histogram-matching method for protein phase refinement and extension. AB - A new method has been developed for protein phase improvement using the joint distribution of the electron density and its gradient (two-dimensional histogram) as a constraint in a density-modification procedure. Matching the two-dimensional (2D) histogram of a given map to that of an ideal 2D histogram was achieved through alternating applications of one-dimensional (1D) histogram matching on electron density and on density gradient. The 2D histogram-matching method was compared with the 1D density histogram-matching method for phase refinement and extension starting from either medium-resolution or high-resolution data on three different types of phases. These included phase refinement and extension using MIR phases of T(6) insulin and phases with randomly generated errors. The test results demonstrated significant improvement of the phases and the overall map quality using the 2D histogram-matching method compared with the 1D density histogram-matching method in each of the three test cases. PMID- 10531489 TI - Crystallization and preliminary crystallographic analysis of major nitroreductase from Escherichia coli. AB - NADPH:nitrocompound oxidoreductase from Escherichia coli, NfsA, has been crystallized in the presence of FMN by the vapor-diffusion method using polyethylene glycol 6000 as a precipitant. The crystals belonged to the triclinic space group P1 with cell dimensions, a = 52.2, b = 52.7, c = 53.3 A, alpha = 75.1, beta = 60.1, gamma = 60.5 degrees. The crystals are expected to contain two NfsA molecules per asymmetric unit. The crystals diffracted X-rays to at least 2.3 A resolution and are appropriate for structural analysis at high resolution. PMID- 10531490 TI - Crystallization of Helix pomatia agglutinin (HPA), a protein from the edible snail. AB - Crystals of Helix pomatia agglutinin (HPA) have been grown by the hanging-drop technique using polyethylene glycol as the precipitant at 293 K. Over a period of one to two weeks the crystals grew to maximum dimensions of 0.10 x 0.05 x 0.02 mm. The crystals belong to space group P6(3)22, with unit-cell dimensions a = b = 63.3, c = 105. 2 A and Z = 12 identical monomers of M(r) = 13 kDa, aggregating into two 78 kDa hexameric protein molecules per unit cell, each with symmetry 32 (D(3)). The diffraction pattern extends to 3.6 A at 293 K. PMID- 10531491 TI - Purification, solution properties and crystallization of SIV integrase containing a continuous core and C-terminal domain. AB - The C-terminal two-thirds segment of integrase derived from the simian immunodeficiency virus has been cloned, expressed in Escherichia coli, and purified to greater than 95% homogeneity. The protein encompasses amino-acid residues 50-293 and contains a F185H substitution to enhance solubility. In dilute solutions at concentrations below 1 mg ml(-1), the enzyme is predominantly dimeric. At the higher concentrations (>10 mg ml(-1)) required to enable crystallization, the enzyme self-associates to form species with molecular weights greater than 200 kDa. Despite the apparent high aggregation in solution, the enzyme crystallizes from a 8%(v/v) polyethylene glycol (molecular weight 6000) solution in a form suitable for X-ray diffraction studies. The resulting single crystals belong to the space group P2(1)2(1)2(1), with unit-cell parameters a = 79.76, b = 99.98, c = 150.2 A, alpha = beta = gamma = 90 degrees and Z = 4. Under X-ray irradiation generated with a rotating-anode generator, the crystals diffract to 2.8 A resolution and allow collection of a native 3 A resolution diffraction data set. PMID- 10531492 TI - Crystallization and preliminary x-ray studies of flavocetin-A, a platelet glycoprotein Ib-binding protein from the habu snake venom. AB - Flavocetin-A (FL-A) is a platelet glycoprotein Ib-binding protein, a high molecular mass oligomer (149 kDa) of C-type lectin-like subunits alpha and beta isolated from the habu snake venom. Purified FL-A crystallized in the tetragonal space group I4 with unit-cell dimensions a = b = 121.0, c = 63.2 A. The crystals diffract to at least 2.4 A resolution. The structure has been solved by molecular replacement using the crystal structure of factors IX/X-binding protein (PDB code 1ixx) as a search model. The asymmetric unit contains one heterodimer, showing that FL-A is a novel tetradimer (alphabeta)(4) composed of four heterodimers related by a crystallographic fourfold axis. PMID- 10531493 TI - Crystallization and x-ray diffraction data analysis of human deoxyhaemoglobin A(0) fully stripped of any anions. AB - In this work, initial crystallographic studies of human haemoglobin (Hb) crystallized in isoionic and oxygen-free PEG solution are presented. Under these conditions, functional measurements of the O(2)-linked binding of water molecules and release of protons have evidenced that Hb assumes an unforeseen new allosteric conformation. The determination of the high-resolution structure of the crystal of human deoxy-Hb fully stripped of anions may provide a structural explanation for the role of anions in the allosteric properties of Hb and, particularly, for the influence of chloride on the Bohr effect, the mechanism by which Hb oxygen affinity is regulated by pH. X-ray diffraction data were collected to 1.87 A resolution using a synchrotron-radiation source. Crystals belong to the space group P2(1)2(1)2 and preliminary analysis revealed the presence of one tetramer in the asymmetric unit. The structure is currently being refined using maximum-likelihood protocols. PMID- 10531494 TI - Crystallization and preliminary x-ray diffraction studies of hyperthermostable glutamate dehydrogenase from Thermococcus profundus. AB - Recombinant glutamate dehydrogenase from a hyperthermophilic archaeon, Thermococcus profundus, was crystallized in the presence of both polyethylene glycol 8000 and lithium sulfate. Four types of crystals having different morphologies appeared in the crystallization trials; however, only one type was suitable for X-ray crystal structure analysis. The crystal belonged to the monoclinic space group P2(1) and the unit-cell parameters were a = 112.99, b = 163.70, c = 133.07 A, beta = 113.46 degrees at 110 K. The calculated V(M) value of 3.42 A(3) Da(-1) was acceptable when one hexamer of the enzyme, which was the physiological functional unit, occupied a crystallographic asymmetric unit. X-ray diffraction intensity data were collected to a resolution of 2.25 A with good statistics at the BL44B2 beamline of SPring-8. PMID- 10531495 TI - Crystallization and preliminary x-ray diffraction studies of a Bowman-Birk inhibitor from Vigna unguiculata seeds. AB - A Bowman-Birk type trypsin/chymotrypsin inhibitor isolated from Vigna unguiculata seeds has been crystallized. Crystals were grown using the vapour-diffusion method at pH 4.0 using citrate/phosphate as a buffer and 30% saturated ammonium sulfate as precipitant. The crystals belonged to the monoclinic space group P2(1), with unit-cell parameters a = 32.4, b = 61.8, c = 32.9 A, beta = 114.5 degrees. The Matthews coefficient calculated assuming two molecules in the asymmetric unit was 1.95 A(3) Da(-1), which corresponds to a 37% solvent content. X-ray data were collected to 2.5 A resolution from a flash-frozen crystal. The structure was solved using the molecular-replacement method using tracy soybean inhibitor structure (PDB entry 1pi2) as a model. PMID- 10531496 TI - Crystallization of the N-terminal domain of the Escherichia coli regulatory protein TyrR. AB - The N-terminal domain of the regulatory protein TyrR from Escherichia coli forms a dimer in solution and has been purified and crystallized. The crystals belong to space group C2 with unit-cell parameters a = 134.5, b = 72.1, c = 96.7 A, beta = 98.5 degrees. The crystals diffract to 2.8 A. Assuming a molecular weight of 23219 Da, a V(m) of 2.5 A(3) Da(-1) is obtained for two dimers in the asymmetric unit. PMID- 10531497 TI - Crystallization and preliminary x-ray structure determination of Lupinus luteus PR10 protein. AB - The pathogenesis-related protein of the PR10 class from Lupinus luteus (yellow lupin), LlPR10.1A, is constitutively expressed in roots. It is also accumulated in leaves treated with a suspension of pathogenic bacteria as a response to stress. Recombinant yellow-lupin LlPR10.1A protein has been overexpressed in Escherichia coli as a fusion product with maltose-binding protein. LlPR10.1A crystallizes in the orthorhombic P2(1)2(1)2(1) space group and the crystals diffract to 2.45 A resolution. The structure has been solved by molecular replacement, using the structure of a birch-pollen allergen protein as a model. PMID- 10531498 TI - Crystallization and preliminary x-ray diffraction studies of guanidinoacetate methyltransferase from rat liver. AB - Guanidinoacetate methyltransferase is the enzyme which catalyzes the last step of creatine biosynthesis. The enzyme is found ubiquitously and in abundance in the livers of all vertebrates. Recombinant rat-liver guanidinoacetate methyltransferase has been crystallized with guanidinoacetate and S adenosylhomocysteine. The crystals belong to the monoclinic space group P2(1), with unit-cell parameters a = 54.8, b = 162.5, c = 56.1 A, beta = 96.8 (1) degrees at 93 K, and typically diffract beyond 2.8 A. PMID- 10531499 TI - Crystallization and preliminary x-ray diffraction analysis of malic enzyme from pigeon liver. AB - Recombinant pigeon-liver malic enzyme was expressed in Escherichia coli and purified to homogeneity. Two different crystal forms were grown by the hanging drop vapour-diffusion method. Both types of crystals belong to the tetragonal space group P4(2)22, with unit-cell dimensions a = b = 163.8, c = 174.3 A for the octahedral crystals and a = b = 124.5, c = 179.2 A for the rod-like crystals. X ray diffraction data were collected at 100 K using a synchrotron-radiation X-ray source. The Matthews parameter suggests that there are four and two molecules per asymmetric unit for the larger and the smaller tetragonal unit cells, respectively. PMID- 10531500 TI - Crystallization and preliminary diffraction studies of two quinoprotein alcohol dehydrogenases (ADHs): a soluble monomeric ADH from Pseudomonas putida HK5 (ADH IIB) and a heterotrimeric membrane-bound ADH from Gluconobacter suboxydans (ADH GS). AB - Crystals of a soluble monomeric quinocytochrome alcohol dehydrogenase (ADH-IIB) and of a trimeric membrane-associated quinocytochrome alcohol dehydrogenase (ADH GS) have been obtained. The ADH-IIB crystals are triclinic, with one monomer in the unit cell, and were obtained in the presence of PEG 8000, sodium citrate, HEPES buffer and 2-propanol. X-ray data were collected at 110 K to 1. 9 A resolution (R(merge) = 6.4%) and the orientation of a methanol dehydrogenase search molecule (from Methylophilus methylotrophus W3A1) was obtained by molecular replacement. Preliminary refinement of this model (10.0-3.0 A resolution, R = 0.37, R(free) = 0.40) led to tentative identification of the two highest peaks in a native anomalous difference Fourier map as the Fe atom of the heme and a calcium ion interacting with the PQQ prosthetic group. The ADH-GS crystals are tetragonal, displaying six similar lattices, both primitive and centered, and were grown by the sitting-drop method after replacement of Triton X 100 by dodecylmaltoside or octaethylene glycol monododecyl ether in the presence of ammonium sulfate and sodium acetate buffer, with and without PEG 3500 and calcium ion. The best diffraction is obtained at 110 K where the resolution extends to about 4 A in the a and b directions and about 3 A in the c direction. PMID- 10531501 TI - Crystallization and preliminary x-ray diffraction studies of C4-form phosphoenolpyruvate carboxylase from maize. AB - Phosphoenolpyruvate carboxylase is a key enzyme in the fixation of atmospheric CO(2) in C(4) and crassulacean acid metabolism (CAM) plants. The enzyme catalyzes the irreversible carboxylation of phosphoenolpyruvate to form oxaloacetate and inorganic phosphate, the first committed step in the fixation of external CO(2) in these plants. The enzyme has been isolated from maize leaves and crystallized using the hanging-drop vapour-diffusion method with PEG 8000 as a precipitant at pH 7.5. The crystals belong to space group C222(1), with unit-cell dimensions a = 160.2, b = 175.6, c = 255.5 A, and diffract to 3.2 A resolution. PMID- 10531502 TI - Expression of a selenomethionyl derivative and preliminary crystallographic studies of human cystatin C. AB - Human cystatin C, a protein with amyloidogenic properties and a potent inhibitor of papain-like mammalian proteases, has been produced in its full-length form by recombinant techniques and crystallized in two polymorphic forms: cubic and tetragonal. A selenomethionyl derivative of the protein, obtained by Escherichia coli expression and with complete Met-->Se-Met substitution confirmed by mass spectrometry, amino-acid analysis and X-ray absorption spectra, was crystallized in the cubic form. A truncated variant of the protein, lacking ten N-terminal residues, has also been crystallized. The crystals of this variant are tetragonal and, like the two polymorphs of the full-length protein, contain multiple copies of the molecule in the asymmetric unit, suggesting oligomerization of the protein. PMID- 10531503 TI - Crystallization and preliminary diffraction analysis of the HincII restriction endonuclease-DNA complex. AB - Crystals of the 60 kDa dimeric HincII restriction enzyme bound to a 12 base-pair dyad-symmetric duplex DNA carrying the specific 5'-GTCGAC recognition site have been obtained. Crystals grew by hanging-drop vapor diffusion from solutions containing polyethylene glycol 4000 as precipitating agent. The rod-shaped crystals belong to space group I222 (or I2(1)2(1)2(1)), with unit-cell dimensions a = 66.9, b = 176.7, c = 256.0 A. There are most likely to be two dimeric complexes in the asymmetric unit. A complete native data set has been collected from a high-energy synchrotron source to a resolution of 2.5 A at 100 K, with an R(merge) of 4.8%. PMID- 10531504 TI - Initiating a structural study of 2-keto-3-deoxy-6-phosphogluconate aldolase from Escherichia coli. AB - 2-Keto-3-deoxy-6-phosphogluconate aldolase (KDPG aldolase, E.C. 4.1. 2.14) is a member of the pyruvate/phosphoenolpyruvate aldolase family. It is also a synthetically useful enzyme, capable of catalyzing the stereoselective aldol addition of pyruvate to a range of unnatural electrophilic substrates. The recombinant protein was purified by a two-step HPLC protocol involving anion exchange and hydrophobic chromatography. Dynamic light-scattering experiments indicated the protein to be monodisperse. Crystals were obtained using the sitting-drop vapour-diffusion method, with PEG 6K as precipitant. Diffraction data were collected on a frozen crystal to a resolution of 2.26 A on station PX9.6 at the Daresbury synchrotron. The crystal belongs to space group P2(1)2(1)2(1), with unit-cell parameters a = 53.2, b = 77.9, c = 146.8 A. PMID- 10531505 TI - Crystallization and preliminary x-ray diffraction studies on the conserved GTPase domain of the signal recognition particle from Acidianus ambivalens. AB - The signal recognition particle (SRP) of bacteria consists of only one protein, known as Ffh or the SRP54 homologue, which forms a complex with 4.5S RNA. It also binds to signal peptides and contains a GTPase which displays interesting differences to Ras GTPases. The conserved NG-domain of Ffh from the archaebacterium Acidianus ambivalens was cloned and overexpressed with a C terminal His tag in Escherichia coli. Crystallization experiments of the native protein as well as of the Thr112Ala mutant, which is deficient in GTP hydrolysis, resulted in crystals suitable for X-ray diffraction. The crystals belong to the orthorhombic space group C222(1), with unit-cell parameters a = 64.5, b = 128.3, c = 72.0 A. At cryogenic temperatures, the crystals diffracted to a resolution limit of 2.8 A using a rotating-anode generator and contain one molecule per asymmetric unit. A native data set has been collected using synchrotron radiation to around 2.0 A resolution. Selenomethionine protein was produced; its crystals diffract in-house to about 2.8 A resolution. PMID- 10531506 TI - Crystallization and preliminary X-ray diffraction analysis of the homodimeric form alpha2 of the HU protein from Escherichia coli. AB - The homodimeric form alpha(2) of the Escherichia coli DNA-binding protein HU was crystallized by the hanging-drop vapour-diffusion method using PEG 4000 as a precipitant. The crystals belong to space group I222, with unit-cell parameters a = 31.09, b = 55.34, c = 117. 63 A, and contain one monomer per asymmetric unit. A full diffraction data set was collected to 2.3 A resolution on a conventional X ray source. The molecular-replacement method, using the HU crystallographic model from Bacillus stearothermophilus as a starting point, gave a reliable solution for the rotation and translation functions. PMID- 10531507 TI - Expression, purification, crystallization and preliminary x-ray data of Escherichia coli galactoside acetyltransferase. AB - Crystals of galactoside acetyltransferase from Escherichia coli have been prepared from solutions of ammonium sulfate containing acetyl-CoA. These crystals diffract to at least 2.7 A resolution, belong to space group C222(1) and contain one copy of the trimeric enzyme in the asymmetric unit. PMID- 10531508 TI - Purification, crystallization and preliminary crystallographic studies on the N terminal fragment of human protein disulfide isomerase. AB - A fragment of human protein disulfide isomerase composed of the thioredoxin-like a and b domains (ab) has been expressed in Escherichia coli as a fusion protein with glutathione-S-transferase and purified after thrombin cleavage. Two forms of ab crystal were obtained with polyethylene glycol as precipitant and different additives at pH 7.5. The space group of form I is P4(1)2(1)2 or P4(3)2(1)2, with unit-cell dimensions a = 81.5, c = 259.7 A. The space group of form II is P4(1)22 or P4(3)22, with unit-cell dimensions a = 82.7, c = 86.5 A. PMID- 10531509 TI - Purification and crystallization of precursors and autoprocessed enzymes of Flavobacterium glycosylasparaginase: an N-terminal nucleophile hydrolase. AB - Glycosylasparaginase (GA) represents a novel group of proteins that are activated by self-catalyzed peptide-bond cleavage from a single-chain precursor to yield the two subunits required for hydrolase activity. The wild-type GA precursor autoproteolyzes spontaneously into alpha and beta subunits. Strategies are reported here for purification to homogeneity of GA from Flavobacterium meningosepticum in both single-chain precursor and mature (autoprocessed) forms. The recombinant proteins crystallize in different space groups: P1 and P2(1) for the precursor and mature enzymes, respectively. The precursor crystals diffract to 1.9 A resolution with laboratory X-ray radiation. PMID- 10531510 TI - Preparation and preliminary study of crystals of the recombinant calcium regulated photoprotein obelin from the bioluminescent hydroid Obelia longissima. AB - Crystals of recombinant obelin, the Ca(2+)-regulated photoprotein from the marine hydroid Obelia longissima, have been grown from sodium citrate solutions. Crystals grow as hexagonal light-yellow rods (0.1 x 0.1 x 1.0 mm) which diffract to beyond 1.8 A with synchrotron radiation of 1.0 A wavelength. The crystals have a primitive hexagonal lattice with unit-cell parameters a = 81.55, c = 86.95 A. The asymmetric unit contains two molecules. This represents the successful preparation of single crystals of a photoprotein obelin which have promising diffraction properties. PMID- 10531511 TI - Data processing. AB - X-ray diffraction data processing proceeds through indexing, pre-refinement of camera parameters and crystal orientation, intensity integration, post-refinement and scaling. The DENZO program has set new standards for autoindexing, but no publication has appeared which describes the algorithm. In the development of the new Data Processing Suite (DPS), one of the first aims has been the development of an autoindexing procedure at least as powerful as that used by DENZO. The resultant algorithm will be described. Another major problem which has arisen in recent years is scaling and post-refinement of data from different images when there are few, if any, full reflections. This occurs when the mosaic spread approaches or exceeds the angle of oscillation, as is usually the case for frozen crystals. A procedure which is able to obtain satisfactory results for such a situation will be described. PMID- 10531512 TI - Cool data: quantity AND quality. AB - The use of cryo-techniques in macromolecular crystallography has increased enormously over the last eight years and has become a vital part of modern X-ray data-collection methods. This paper presents some reasons for the rise in popularity of cryo-techniques and a brief outline of the basic methods, followed by a detailed discussion of factors to be considered when trying to optimize both the quantity and quality of the data collected. As more experimenters at synchrotrons observe significant radiation damage to crystals held near 100 K, the available options for further prolonging crystal lifetime and extending the techniques become worth investigating. Some possibilities and parameters to be considered are presented, although these must remain speculative until more experimental data are available. PMID- 10531514 TI - Matching X-ray source, optics and detectors to protein crystallography requirements. AB - A review of the requirements for collecting X-ray diffraction data from protein crystals is given, with an emphasis on the properties of the crystal and its diffraction pattern. The size, unit-cell dimensions and perfection of the crystals can all be related to the required size and divergence of the incident X ray beam, together with the size and spatial resolution of the detector. The X ray beam causes primary radiation damage, even in frozen crystals. If the incident beam is very intense, temperature rises and gradients could occur in the crystal. The extent to which these problems can be overcome is also discussed. PMID- 10531513 TI - Macromolecular crystallography with a third-generation synchrotron source. AB - The European Synchrotron Radiation Facility (ESRF) at Grenoble, France, is a 6 GeV machine producing hard X-radiation that can be used for pure and applied research in a wide range of disciplines including physics, chemistry, structural biology, materials science, the earth sciences, engineering and medicine. The overall nature of the machine will be described, including the features that give rise to the notation 'third-generation source'. The ESRF is equipped with a number of beamlines which can be used for macromolecular crystallography. Applications include the use of very small crystals, large unit cells, data collection at high resolution, anomalous dispersion measurements for phase determination and time-resolved studies. Key features of these applications will be described. PMID- 10531515 TI - Experiences with CCD detectors on a home X-ray source. AB - Charged-coupled device (CCD) detectors have been widely accepted as detectors for collecting X-ray diffraction images. The CCD detector offers a sensitive detection system well suited for diffraction analysis and, compared with other detectors on the market, a relatively rapid system for read-out of the collected image. The two predominant markets for the CCD detector have been those in which relatively short exposure times are used, i.e. small-molecule X-ray diffraction and large-molecule crystallography at high-intensity synchrotron sources. CCD detectors have not been commonly used on rotating-anode X-ray sources for large molecule crystallography. Comparison of the performance of the CCD detectors with commercially available image-plate detectors shows that the CCD detectors function in a similar fashion to image-plate-based detectors. PMID- 10531516 TI - Experiences and expectations of a novel X-ray microsource with focusing mirror. I. AB - Data are presented from a novel microfocus X-ray generator installed with a choice of ellipsoidal specularly reflecting mirrors. Diffraction data from proteins show the useful flux from this low-power device to be approaching equivalence with that from many far more powerful generators. Intensity measurements show that for small crystals the brilliance is now restricted by the performance of the mirror, which appears to be limited by imperfections in the figure of its surface rather than by a low reflectivity. Suitable choices of ellipsoidal mirror enable the size and divergence of the X-ray beam to be altered readily to match the different requirements of successive samples and appropriate designs are proposed. Alternative types of mirror are expected to be advantageous, especially for the smallest crystals. For crystals of sizes 300 microm or less, which need a small well collimated beam with low divergence, the output from this X-ray tube running at 24 W provides a usable flux similar to that available from rotating-anode generators. The relative performance of this tube and mirror combination becomes increasingly advantageous with the study of ever-smaller crystals. PMID- 10531517 TI - Optics systems for the home laboratory: caveat emptor. AB - A careful and detailed evaluation of different multilayer optics (Osmic Cross coupled Max-Flux Optics and Osmic Confocal Max-Flux Optics) compared with MSC/Yale Total-Reflection Mirrors has been completed. This report provides a detailed comparison of usable flux, spectral purity, divergence, beam profile and data quality for these systems. The most striking results have been obtained using either the Osmic #4 or #7 Confocal Max-Flux Optic, which were designed for 0. l and 0.2 mm focal spots, respectively, in conjunction with a 0.3 mm focal spot. These optic configurations provide a 5.8-fold and 8.2-fold increase in flux through a 0.2 mm aperture, respectively, compared with the MSC/Yale Mirrors. PMID- 10531518 TI - The Rossmann Fourier autoindexing algorithm in MOSFLM. AB - The fast Fourier transform (FFT) autoindexing routines written by the Rossmann group at Purdue University have been incorporated in MOSFLM, providing a rapid and reliable method of indexing oscillation images. This is a procedure which extracts direct-space information about the unit cell from the FFT. The method and its implementation in MOSFLM are discussed. PMID- 10531519 TI - Integration of macromolecular diffraction data. AB - Diffraction intensities can be evaluated by two distinct procedures: summation integration and profile fitting. Equations are derived for evaluating the intensities and their standard errors for both cases, based on Poisson statistics. These equations highlight the importance of the contribution of the X ray background to the standard error and give an estimate of the improvement which can be achieved by profile fitting. Profile fitting offers additional advantages in allowing estimation of saturated reflections and in dealing with incompletely resolved diffraction spots. PMID- 10531520 TI - Data-collection strategies. AB - The optimal strategy for collecting X-ray diffraction data from macromolecular crystals is discussed. Two kinds of factors influencing the completeness of data are considered. The first are geometric, arising from the symmetry of the reciprocal lattice and from the experimental setup; they affect quantitatively the completeness of the measured set of reflections. The second concern the quality, or information content, of the recorded intensities of these measured reflections. PMID- 10531521 TI - The finer things in X-ray diffraction data collection. AB - X-ray diffraction images from two-dimensional position-sensitive detectors can be characterized as thick or thin, depending on whether the rotation-angle increment per image is greater than or less than the crystal mosaicity, respectively. The expectations and consequences of the processing of thick and thin images in terms of spatial overlap, saturated pixels, X-ray background and I/sigma(I) are discussed. The d*TREK software suite for processing diffraction images is briefly introduced, and results from d*TREK are compared with those from another popular package. PMID- 10531522 TI - MAD data collection - current trends. AB - The multiwavelength anomalous dispersion (MAD) method of protein structure determination is becoming a routine technique in protein crystallography. The increased number of wavelength-tuneable synchrotron beamlines capable of performing challenging MAD experiments, coupled with the widespread availability of charge-coupled device (CCD) based X-ray detectors with fast read-out times have brought MAD structure determination to a new exciting level. Ultrafast MAD data collection is now possible and, with the widespread use of selenium in the form of selenomethionine for phase determination, the method is growing in popularity. Recent developments in crystallographic software are complementing the above advances, paving the way for rapid protein structure determination. An overview of a typical MAD experiment is described, with emphasis on the rates and quality of data acquisition now achievable at third-generation synchrotron sources. PMID- 10531523 TI - New processing tools for weak and/or spatially overlapped macromolecular diffraction patterns. AB - Tools originally developed for the treatment of weak and/or spatially overlapped time-resolved Laue patterns were extended to improve the processing of difficult monochromatic data sets. The integration program PrOW allows deconvolution of spatially overlapped spots which are usually rejected by standard packages. By using dynamically adjusted profile-fitting areas, a carefully built library of reference spots and interpolation of reference profiles, this program also provides a more accurate evaluation of weak spots. In addition, by using Wilson statistics, it allows rejection of non-redundant strong outliers such as zingers, which otherwise may badly corrupt the data. A weighting method for optimizing structure-factor amplitude differences, based on Bayesian statistics and originally applied to low signal-to-noise ratio time-resolved Laue data, is also shown to significantly improve other types of subtle amplitude differences, such as anomalous differences. PMID- 10531524 TI - Complementing crystallography: the role of cryo-electron microscopy in structural biology. AB - Dramatic improvements in experimental methods and computational techniques have revolutionized three-dimensional image reconstruction from electron micrographs (EM) of vitrified samples. Recent results include the first determination of a protein fold (for the core protein of the hepatitis B virus) by non-crystalline imaging techniques. These developments have generated interest within the crystallographic community and have led to a re-evaluation of the technique, particularly amongst those working in the field of virus structure or struggling with the phasing of large macromolecular assemblies. A simple discussion of the techniques of EM image reconstruction and its advantages and problems in terms familiar to crystallographers will hopefully allow an appreciation of the essential complementarity of the two techniques and the practical potentials for phasing applications. PMID- 10531525 TI - Analysis and characterization of data from twinned crystals. AB - It is difficult but not impossible to determine a macromolecular structure using X-ray data obtained from twinned crystals, providing it is noticed and corrected. For perfectly twinned crystals, the structure can probably only be solved by molecular replacement. It is possible to detect and characterize twinning from an analysis of the intensity statistics and crystal packing density. Tables of likely twinning operators and some examples are discussed here. PMID- 10531526 TI - Detecting outliers in non-redundant diffraction data. AB - Outliers are observations which are very unlikely to be correct, as judged by independent observations or other prior information. Such unexpected observations are treated, effectively, as being more informative about possible models, so they can seriously impede the course of structure determination and refinement. The best way to detect and eliminate outliers is to collect highly redundant data, but it is not always possible to make multiple measurements of every reflection. For non-redundant data, the prior expectation given either by a Wilson distribution of intensities or model-based structure-factor probability distributions can be used to detect outliers. This captures mostly the excessively strong reflections, which dominate the features of electron-density maps or, even more so, Patterson maps. The outlier rejection tests have been implemented in a program, Outliar. PMID- 10531527 TI - Protein microcrystals and the design of a microdiffractometer: current experience and plans at EMBL and ESRF/ID13. AB - There is a growing demand for the examination of protein microcrystals at third generation synchrotron sources. After successful pilot experiments at EMBL/ESRF, which proved that protein microcrystals are often suitable for data collection, operation of the microfocus beamline ID13 was made more user-friendly and suitable for macromolecular crystallography experiments. Given the excellent quality of the beamline microfocusing optics, the key element for successful experiments becomes the handling and visualization of microcrystals. To address this, a microdiffractometer has been designed to allow maximum precision combined with ease of usage and is currently under construction. PMID- 10531528 TI - Some notes on choices in data collection. AB - Collecting optimum X-ray diffraction data involves a number of choices and compromises, including choice of crystal, source, rotation range, exposure time and programs for integration and scaling. This paper presents a series of questions which should be considered in planning a data-collection experiment. PMID- 10531529 TI - Retroactive interference in 3-month-old infants. AB - Three experiments assessed the effect of the similarity and timing of interpolated information on retroactive interference in human infants. After 3 month-olds learned to move a mobile (the cue) in a distinctive context by kicking, they were exposed to novel stimuli and were tested 24 hr later for recognition of their training stimuli. In Experiment 1, the interpolated cue, context, or both were novel. Retroactive interference occurred unless the interpolated and training stimuli shared no components. In Experiments 2 and 3, the timing of exposure to the interpolated context or cue, respectively, varied. A novel context impaired recognition after exposure delays up to 2 hr, whereas a novel cue impaired recognition after exposure delays up to 40 min. The finding that the cue is less vulnerable to retroactive interference than the context suggests that it is processed more rapidly. These experiments reveal that retroactive interference in infants depends on both the similarity and timing of the interpolated information. PMID- 10531530 TI - Motor overflow and children's tracking performance: is there a link? AB - The aim of this study was to investigate the interaction of skill performance, motor overflow, and hand linkage in the form of mirror movements in a visual manual tracking paradigm across practice trials. We hypothesized that both the amount of motor overflow and the degree of hand linkage would be linked in an inverse way to the quality of task performance. Furthermore, we expected a short term decrease in both of these factors as children practiced and improved their task performance. Sixteen right-handed, 6-year-old children tracked a visual target with their right hand by pinching two parallel steel bars instrumented with strain gauges. The left hand was also positioned by similar instrumented steel bars to measure overflow/mirroring. At both the beginning and end of practice trials, a cluster analysis was used to determine relationships among performance, overflow, and hand linkage variables. In general, the results support the main hypothesis that the amount of motor overflow and the degree of hand linkage are linked to the quality of task performance, but the relationships between these variables across short-term learning are nonlinear. PMID- 10531531 TI - The start of a new school year: individual differences in salivary cortisol response in relation to child temperament. AB - Noon and evening salivary cortisol levels were examined in 70 elementary school children during the 1st week of a new school year. Samples were obtained on the 1st and 5th days of school and on weekend days. Delta cortisol scores were created to measure the change in children's levels on initial school days relative to weekend days. Temperament was assessed using Rothbart's Child Behavior Questionnaire, a parent report instrument. The three dimensions of surgency or extroversion, negative affectivity, and effortful control were examined. Positive correlations were obtained with Day 1 delta cortisol for negative affectivity and Day 5 delta cortisol for surgency. Contrary to the expectation that internalizing aspects of temperament (shyness, fearfulness) would be associated with larger increases in cortisol to the novelty and challenge of a new school year, these data indicate that larger increases in cortisol were observed in more extroverted children. PMID- 10531532 TI - Experience with a flavor in mother's milk modifies the infant's acceptance of flavored cereal. AB - The present series of studies aimed to investigate whether experience with a flavor in mothers' milk modifies the infants' acceptance of similarly flavored foods at weaning. First, we established, using methods developed in our laboratory, that the ingestion of carrot juice by lactating women produced a sensory change in their milk approximately 2 to 3 hr after the ingestion of the beverage. Second, we randomly formed two groups of breast-fed infants who had been fed cereal for a few weeks but had only experienced cereal prepared with water. Their mothers were asked to consume one of two types of beverages (i.e., carrot juice, water) during the exposure period. Each mother was observed feeding her infant cereal during four test sessions. The first two sessions occurred during the 2 days before the exposure period; in counterbalanced order, infants were fed cereal prepared with water on 1 testing day and cereal prepared with carrot juice on the other. These two test sessions were then repeated following the exposure period. The results demonstrated that the infants who had exposure to the flavor of carrots in their mothers' milk during the exposure period consumed less of the carrot-flavored cereal and spent less time feeding when compared to the control infants whose mothers consumed the water. This may be a form of sensory-specific satiety such that the infants become less responsive to a flavor that they have been extensively exposed to in the very recent past. PMID- 10531533 TI - Prenatal experience and neonatal responsiveness to vocal expressions of emotion. AB - Newborn differentiation of emotion and the relevance of prenatal experience in influencing responsiveness to emotion was tested by examining newborn responses to the presentation of a range of vocal expressions. Differential responding was observed, as indicated by an increase in eye opening behavior in response to the presentation of happy speech patterns. More importantly, differential responding was observed only when the infants listened to emotional speech as spoken by speakers of their maternal language. No evidence of discrimination was found in the groups of infants listening to the same vocal expressions in a novel language. The results suggest that as a consequence of prenatal exposure to the distinctive prosodic maternal speech patterns that specify different emotions and to the temporally related stimuli created by distinctive maternal physiological concomitants of emotion, the fetus learns to differentiate those emotional speech patterns typical of the infant's maternal language. PMID- 10531534 TI - Augmented prenatal tactile and vestibular stimulation alters postnatal auditory and visual responsiveness in bobwhite quail chicks. AB - The fact that the sensory systems do not become functional at the same time during early development raises the question of how sensory systems and their respective stimulative histories might influence one another. Previous studies have shown that unusually early visual experience can alter subsequent responsiveness of both the visual system and the earlier developing olfactory and auditory systems. The question remains as to the extent which modified stimulation to an earlier developing system can also result in changes in responsiveness in later developing sensory systems. This study examined the effects of augmented prenatal tactile and vestibular stimulation on bobwhite quail chicks' postnatal visual and auditory responsiveness to maternal cues. Results indicate that augmented prenatal tactile and vestibular stimulation can alter postnatal perceptual responsivensss in the later developing auditory and visual sensory systems. Chicks exposed to augmented prenatal proximal stimulation continued to respond to maternal auditory cues into later stages of postnatal development and failed to demonstrate responsiveness to maternal visual cues in the days following hatching. However, augmented tactile and vestibular stimulation did not appear to affect prenatal auditory learning of an individual maternal call. These findings indicate a strong but selective pattern of influence between the sensory modalities during the prenatal period and support the view that substantially increased amounts of prenatal sensory stimulation can interfere with the emergence of species-typical perceptual functioning. PMID- 10531535 TI - The effects of neonatal choline dietary supplementation on adult spatial and configural learning and memory in rats. AB - The facilitative effects of pre- and early postnatal choline chloride dietary supplementation on adult rat spatial and nonspatial learning and memory were examined using a delayed match-to-place and a transverse-patterning discrimination task. Animals were exposed to the choline supplementation both prenatally (through the diet of pregnant rats) and postnatally (subcutaneous injection) for 24 days. In the first experiment, 90-day-old rats were given pairs of trials in which they first found a hidden platform in a Morris water maze in a particular location (acquisition trials), and then were required to remember that position 10 min later (test trials). Those animals given neonatal choline pretreatment found the platform on test trials significantly faster than did animals in a saline-treated control group. All animals were subsequently tested in the same paradigm following atropine sulfate injections. The atropine eliminated the difference between experimental and control animals on test trials. In a second experiment, neonatally treated choline rats performed significantly better than controls in acquiring a visual transverse patterning discrimination task previously found to be sensitive to hippocampal and/or frontal damage. The present study extends the description of long-term functional enhancement produced by perinatal choline supplementation to include the ability to use and remember visual configural associations, working spatial memory, and to relate these effects to modifications in cholinergic basal forebrain systems. PMID- 10531536 TI - The effect of early handling is dependent upon the state of the rabbit (Oryctolagus cuniculus) pups around nursing. AB - We investigated the behavior toward humans in 4-week-old pups and adult rabbits handled daily at different times around the nursing visits during their 1st week of life. The timing of handling significantly influenced its efficiency in altering the subsequent behavior of rabbits. Animals handled around nursing readily approached a human hand when tested at weaning. Other pups, handled either 6, 12, or 18 hr after nursing, avoided the human hand. Our results show that there is a narrow sensitive period for successful stimulation, because only those rabbits that were handled within the interval starting 15 min before and ending 30 min after nursing became tame. The effect of early handling proved to be long-lasting because nonhandled rabbits tested as adults were afraid of humans and showed behavioral elements of avoidance, while the handled ones behaved fearlessly in the open field. The effect of handling proved to be specific toward humans because both handled and nonhandled animals showed avoidance toward a stuffed fox. PMID- 10531537 TI - Development of motoneurons and primary sensory afferents in the thoracic and lumbar spinal cord of the South American opossum Monodelphis domestica. AB - The postnatal development of the primary sensory afferent projection to the thoracic (T4) and lumbar (L4) spinal cord of the marsupial species Monodelphis domestica was studied by using anterograde and retrograde neuronal tracers. Large numbers of primary afferents and motoneurons were labelled by application of the carbocyanine dye DiI into individual dorsal root ganglia (DRG) afferents in short term organ cultures. Dorsal root axons had entered the cord at birth, but most primary afferent innervation of the grey matter and the establishment of cytoarchitectural lamination occurs postnatally. In addition to ipsilateral projections, some primary afferents that projected to the dorsal horn extended across the midline into the equivalent contralateral regions of the grey matter. Similarly, motoneuron dendrites occasionally extended across midline and into the contralateral grey matter. The first fibres innervating the spinal cord project to the ventral horn and formed increasingly complex terminal arbours in the motor columns between P1 and P7. After P5 many afferents were seen projecting to the dorsal horn, with the superficial dorsal horn being the last region of the spinal grey to be innervated. Histochemical labelling with the lectin Griffonia simplicifolia indicated that C fibre primary afferents had arborised in the superficial dorsal horn by P14. The sequence of primary afferent innervation is thus similar to that described in the rat, but this sequence occurs over a period of several weeks in Monodelphis, compared with several days in the rat. PMID- 10531538 TI - Cellular pathology of hilar neurons in Ammon's horn sclerosis. AB - In addition to functionally affected neuronal signaling pathways, altered axonal, dendritic, and synaptic morphology may contribute to hippocampal hyperexcitability in chronic mesial temporal lobe epilepsies (MTLE). The sclerotic hippocampus in Ammon's horn sclerosis (AHS)-associated MTLE, which shows segmental neuronal cell loss, axonal reorganization, and astrogliosis, would appear particularly susceptible to such changes. To characterize the cellular hippocampal pathology in MTLE, we have analyzed hilar neurons in surgical hippocampus specimens from patients with MTLE. Anatomically well preserved hippocampal specimens from patients with AHS (n = 44) and from patients with focal temporal lesions (non-AHS; n = 20) were studied using confocal laser scanning microscopy (CFLSM) and electron microscopy (EM). Hippocampal samples from three tumor patients without chronic epilepsies and autopsy samples were used as controls. Using intracellular Lucifer Yellow injection and CFLSM, spiny pyramidal, multipolar, and mossy cells as well as non-spiny multipolar neurons have been identified as major hilar cell types in controls and lesion-associated MTLE specimens. In contrast, none of the hilar neurons from AHS specimens displayed a morphology reminiscent of mossy cells. In AHS, a major portion of the pyramidal and multipolar neurons showed extensive dendritic ramification and periodic nodular swellings of dendritic shafts. EM analysis confirmed the altered cellular morphology, with an accumulation of cytoskeletal filaments and increased numbers of mitochondria as the most prominent findings. To characterize cytoskeletal alterations in hilar neurons further, immunohistochemical reactions for neurofilament proteins (NFP), microtubule-associated proteins, and tau were performed. This analysis specifically identified large and atypical hilar neurons with an accumulation of low weight NFP. Our data demonstrate striking structural alterations in hilar neurons of patients with AHS compared with controls and non sclerotic MTLE specimens. Such changes may develop during cellular reorganization in the epileptogenic hippocampus and are likely to contribute to the pathogenesis or maintenance of temporal lobe epilepsy. PMID- 10531539 TI - Distribution pattern of inhibitory and excitatory synapses in the dendritic tree of single masseter alpha-motoneurons in the cat. AB - Little is known about the differences in the distributions of inhibitory and excitatory synapses in the dendritic tree of single motoneurons in the brainstem and spinal cord. In this study, the distribution of gamma-aminobutyric acid (GABA)-, glycine-, and glutamate-like immunoreactivity in axon terminals on dendrites of cat masseter alpha-motoneurons, stained intracellularly with horseradish peroxidase, was examined by using postembedding immunogold histochemistry in serial ultrathin sections. The dendritic tree was divided into three segments: primary (Pd) and distal (Dd) dendrites and intermediate (Id) dendrites between the two segments. Quantitative analysis of 175, 279, and 105 boutons synapsing on 13 Pd, 54 Id, and 81 Dd, respectively, was performed. Fifty percent of the total number of studied boutons were immunopositive for GABA and/or glycine and 48% for glutamate. Among the former, 27% showed glycine immunoreactivity only and 14% were immunoreactive to both glycine and GABA. The remainder (9%) showed immunoreactivity for GABA only. As few as 3% of the boutons were immunonegative for the three amino acids. Most boutons immunoreactive to inhibitory amino acid(s) contained a mixture of spherical, oval, and flattened synaptic vesicles. Most boutons immunoreactive to excitatory amino acid contained clear, spherical, synaptic vesicles with a few dense-cored vesicles. When comparisons of the inhibitory and excitatory boutons were made between the three dendritic segments, the proportion of the inhibitory to the excitatory boutons was high in the Pd (60% vs. 37%) but somewhat low in the Id (46% vs. 52%) and Dd (44% vs. 53%). The percentage of synaptic covering and packing density of the inhibitory synaptic boutons decreased in the order Pd, Id, and Dd, but this trend was not applicable to the excitatory boutons. The present study provides possible evidence that the spatial distribution patterns of inhibitory and excitatory synapses are different in the dendritic tree of jaw-closing alpha-motoneurons. PMID- 10531540 TI - Retinal afferents to the dorsal raphe nucleus in rats and Mongolian gerbils. AB - A direct pathway from the retina to the dorsal raphe nucleus (DRN) has been demonstrated in both albino rats and Mongolian gerbils. Following intraocular injection of cholera toxin subunit B (CTB), a diffuse stream of CTB-positive, fine-caliber optic axons emerged from the optic tract at the level of the pretectum/anterior mesencephalon. In gerbils, CTB-positive axons descended ventromedially into the periaqueductal gray, moving caudally and arborizing extensively throughout the DRN. In rats, the retinal-DRN projection comprised fewer, but larger caliber, axons, which arborized in a relatively restricted region of the lateral and ventral DRN. Following injection of CTB into the lateral DRN, retrogradely labeled ganglion cells (GCs) were observed in whole mount retinas of both species. In gerbils, CTB-positive GCs were distributed over the entire retina, and a nearest-neighbor analysis of CTB-positive GCs showed significant regularity (nonrandomness) in their distribution. The overall distribution of gerbil GC soma diameters ranged from 8 to 22 micrometer and was skewed slightly towards the larger soma diameters. Based on an adaptive mixtures model statistical analysis, two Gaussian distributions appeared to comprise the total GC distribution, with mean soma diameters of 13 (SEM +/-1.7) micrometer, and 17 (SEM +/-1.5) micrometer, respectively. In rats, many fewer CTB-positive GCs were labeled following CTB injections into the lateral DRN, and nearly all occurred in the inferior retina. The total distribution of rat GC soma diameters was similar to that in gerbils and also was skewed towards the larger soma diameters. Major differences observed in the extent and configuration of the retinal-DRN pathway may be related to the diurnal/crepuscular vs. nocturnal habits of these two species. PMID- 10531541 TI - Stereological assessment of the glial reaction to chronic deafferentation of the cochlear nuclei in the macaque monkey (Macaca fascicularis). AB - Neurectomy of the auditory nerve produces a massive deafferentation of the cochlear nuclei (CN) in the brainstem. Degenerating primary afferents are removed in the acute phase, and this is followed by a synaptic reorganization in the CN. As part of an ongoing study on the effect and applicability of auditory brain implants in the CN of Macaca fascicularis monkeys, we studied the chronic response of astrocytes in the CN to bilateral deafferentation of the VIIIth cranial nerve. Four control and five deafferentated animals were employed. The treated animals had a bilateral extradural section of the VIIIth cranial nerve and a survival of 3 months. Animals were euthanized and perfused, and the brainstem was serially sectioned. The astrocyte population of the CN was studied by glial fibrillary acidic protein (GFAP) immunohistochemistry and quantified by unbiased stereological methods. The total length of astrocyte processes, L(proc), was estimated as the product of nuclear volume V(nuc), which was estimated by the Cavalieri method, times the ratio L(V)(proc, nuc) of process length to nuclear volume. Mean nuclear volume was significantly lower in deafferented animals, whereas the mean ratio L(V)(proc, nuc) was higher (albeit no statistical significance was reached). However, the mean total astrocytic process length was virtually the same in both groups. The absence of a length increase in the glial processes indicates a decrease of the astrocytic reaction after the acute phase. No glial scar is present in the CN of the monkey after long-term deafferentation, so the usefulness of auditory brain implants to stimulate CN neurons directly as a means to overcome deafness resulting from direct damage to the VIIIth cranial nerve (i.e., acoustic neuromas) is plausible. PMID- 10531542 TI - Influence of the axotomy to cell body distance in rat rubrospinal and spinal motoneurons: differential regulation of GAP-43, tubulins, and neurofilament-M. AB - Axotomized motoneurons regenerate their axons regardless of whether axotomy occurs proximally or distally from their cell bodies. In contrast, regeneration of rubrospinal axons into peripheral nerve grafts has been detected after cervical but not after thoracic injury of the rubrospinal tract. By using in situ hybridization (ISH) combined with reliable retrograde tracing methods, we compared regeneration-associated gene expression after proximal and distal axotomy in spinal motoneurons versus rubrospinal neurons. Regardless of whether they were axotomized at the iliac crest (proximal) or popliteal fossa (distal), sciatic motoneurons underwent highly pronounced changes in ISH signals for Growth Associated Protein 43 (GAP-43) (10-20x increase) and neurofilament M (60-85% decrease). In contrast, tubulin ISH signals substantially increased only after proximal axotomy (3-5x increase). To compare these changes in gene expression with those of axotomized rubrospinal neurons, the rubrospinal tract was transected at the cervical (proximal) or thoracic (distal) levels of the spinal cord. Cervically axotomized rubrospinal neurons showed three- to fivefold increases in ISH signals for GAP-43 and tubulins (only transient) and a 75% decrease for neurofilament-M. In sharp contrast, thoracic axotomy had only marginal effects. After implantation of peripheral nerve transplants into the spinal cord injury sites, retrograde labeling with the sensitive retrograde tracer Fluoro-Gold identified regenerating rubrospinal neurons only after cervical axotomy. Furthermore, rubrospinal neurons specifically regenerating into the transplants were hypertrophied and expressed high levels of GAP-43 and tubulins. Taken together, these data support the concept that, even if central nervous system (CNS) axons are presented with a permissive/supportive environment, appropriate cell body responses to injury are a prerequisite for CNS axonal regeneration. PMID- 10531543 TI - Immunocytochemical localization of FLRFamide-, proctolin-, and CCAP-like peptides in the stomatogastric nervous system and neurohemal structures of the crayfish, Cherax destructor. AB - To compare the stomatogastric nervous system of the crayfish Cherax destructor with those of other decapod species, the distribution of FLRF (Phe-Leu-Arg-Phe) amide-, proctolin- and crustacean cardioactive peptide (CCAP)-like immunoreactivities was studied in the stomatogastric nervous system and in neurosecretory structures by using wholemount immunocytochemical techniques and confocal microscopy. In addition, the number of cells in the stomatogastric ganglion (19-24) and axon profiles in the stomatogastric nerve (157-165) were counted. FLRFamide-like immunoreactivity was present within numerous cell bodies and neuropil of the commissural ganglia, in the neuropil of the stomatogastric ganglion, and in one cell body of the esophageal ganglion. FLRFamide-like immunoreactivity was also found in two cell bodies at the junction of the stomatogastric nerve with the superior esophageal nerve and in two cell bodies in the inferior ventricular nerve. Proctolin-like immunoreactivity was present in numerous cell bodies and neuropil of the paired commissural ganglia and in the neuropil of the stomatogastric ganglion. CCAP-like immunoreactivity was found in the neuropil and in one to four cell bodies in the commissural ganglia. Both proctolin- and CCAP-immunoreactive varicosities occurred on the surface of the circumesophageal connectives and on the postesophageal commissure, indicating a neurohemal source within the stomatogastric nervous system, which was verified by electron microscopy. The pericardial organs showed FLRFamide-, proctolin-, and CCAP-like immunoreactivity. This staining pattern suggests that FLRFamide-like and proctolin-like peptides are used as neurohormones and as neuromodulators in the stomatogastric nervous system of the crayfish C. destructor, whereas CCAP like peptides may only affect the stomatogastric ganglion as a neurohormone. PMID- 10531544 TI - Reelin expression during embryonic brain development in lacertilian lizards. AB - The expression of reelin mRNA and protein was studied during embryonic brain development in the lacertilian lizards L. viridis and L. galloti, by using radioactive in situ hybridization and immunohistochemistry. At all stages studied, high reelin expression was consistently found in the olfactory bulb, in the lateral cortex, and in neurons of the marginal zone and subplate of medial and dorsal cortical sectors. In the dorsal ventricular ridge (DVR), reelin expression was confined to deeply located, large cells which were more abundant in the caudal than the rostral part of the DVR. In the diencephalon, the ventral lateral geniculate complex and the perirotundal were strongly positive, whereas other nuclei were mostly negative. High reelin signal was associated with some layers in the tectum, with the torus semicircularis, cerebellar granule cell layers, and the ventral horn of the spinal cord. A more moderate signal was detected in the septal nuclei, striatum, retina, habenular nuclei, preoptic and periventricular hypothalamic components, and in reticular nuclei of the mid- and hindbrain. The medial and dorsal cortical plate and Purkinje cells were reelin negative but expressed disabled-1 (Dab1) mRNA. When they are compared with reelin expression during mammalian brain development, our data reveal an evolutionarily conserved canvas of reelin expression, as well as significant differences, particularly in developing cortical fields. The developing lizard cortex differs from that of turtles, birds, crocodiles, and mammals in that it displays heavy reelin expression not only in neurons of the marginal zone that might be homologous to mammalian Cajal-Retzius cells, but also in subplate neurons. This difference in the pattern of reelin expression suggests that the elaborate radial organization of the lacertilian cortical plate, somewhat reminiscent of its mammalian counterpart, results from evolutionary convergence. Our data lend support to the hypothesis that the reelin signaling pathway played a significant role during cortical evolution. PMID- 10531545 TI - Synaptogenesis in the brachial and lumbosacral enlargements of the spinal cord in the postnatal opossum, Monodelphis domestica. AB - Synaptic proteins were localized in light microscopy on sections of the brachial and lumbosacral enlargements of the spinal cord of postnatal opossums, Monodelphis domestica, to determine whether their expression correlates with the development of major motor pathways and simple motor behaviors. The tissues were fixed, cryoprotected, frozen, cut in 15-micrometer sections, and processed immunohistochemically using antibodies against synaptophysin, synaptotagmin-I, or SNAP-25. Immunolabeling was observed in the presumptive white matter before the presumptive gray matter, suggesting that the proteins are evidenced in growing axons before the onset of synaptogenesis, and it was observed in presumed propriospinal axons before most presumed descending axons of supraspinal origin. In the newborn opossum, the immunolabeling was scant in the gray matter and was limited to the periphery of the ventral horn, and indeed few synapses were seen in electron microscopy in nonexperimental material. Labeling increased in intensity and spread throughout the gray matter until 5-7 weeks, when it was no longer found in the white matter and resembled the adult pattern of labeling. Considering the location and relative intensity of the immunolabeling for the three proteins over time in the two enlargements, synaptogenesis occurs according to three general gradients: rostrocaudal, ventrodorsal, and lateromedial. These gradients match those of spinal cord and limb development, and of the growth of descending axons into the cord. Synaptogenesis is most intense when the spinal sensorimotor reflexes begin to be expressed. PMID- 10531546 TI - Trimethylamine oxide transport across plasma membranes of elasmobranch erythrocytes. AB - The aim of this study was to characterize the erythrocyte cell membrane transport of trimethylamine oxide (TMAO) in the little skate, Raja erincea. Uptake of TMAO occurs by two processes, Na(+)-dependent and Na(+)-independent. 2,4 dinitrophenol (2,4 DNP), a known ATP synthesis inhibitor, inhibited TMAO uptake, suggesting the involvement of the Na(+)/K(+)-ATP pump in Na(+)-dependent TMAO transport. Na(+) independent TMAO uptake was stimulated by cell swelling when erythrocytes were incubated in hypotonic elasmobranch incubation medium. Swelling-activated, Na(+) independent TMAO uptake was inhibited by the anion transport inhibitors quinine and 4, 4'-diisthiocyanostilbene-2,2'-disulfonic acid (DIDS), two blockers of the swelling-activated osmolyte channel in skate erythrocytes. TMAO efflux was stimulated by hypotonic stress in the erythrocytes of the spiny dogfish, Squalus acanthias. DIDS also inhibited this efflux, indicating that TMAO is transported by the organic osmolyte channel in the erythrocytes of this elasmobranch as well. J. Exp. Zool. 284:605-609, 1999. PMID- 10531547 TI - Protection against oxidative stress in liver of four different vertebrates. AB - The possible relation between respiratory capacity and antioxidant capacity and susceptibility to oxidative stress of the liver has been investigated in Rattus norvegicus, Gallus gallus domesticus, Lacerta s. sicula, and Rana esculenta. Accordingly, we measured oxygen consumption and cytochrome oxidase activity, glutathione peroxidase and glutathione reductase activity and overall antioxidant capacity, and lipid peroxidation and response to oxidative stress in vitro in liver. The order of liver oxygen consumption and cytochrome oxidase activity among the different species was rat > chick > lizard > frog. The antioxidant defenses supplied by the combined action of glutathione peroxidase and glutathione reductase were not adapted to the respiratory capacities. In particular, there was no correlation either between the activities of two enzymes or between their activities and oxygen consumption. In contrast, the overall antioxidant capacity of the liver appeared to be related to its oxidative capacity, and the malondialdehyde formation, an indirect measure of lipid peroxidation, was inversely related to antioxidant capacity. The response to oxidative stress in vitro indicated that the liver susceptibility to oxidative challenge is higher in ectothermic than in endothermic species. Such higher susceptibility appeared to depend on both lower antioxidant capacity and higher levels of free radical producing species. This finding is apparently in contrast with a higher content of cytochromes in endotherms, which are able to determine both respiratory characteristics and sensitivity to pro-oxidants. However, it could indicate the existence of species-related differences in the tissue content of either preventive antioxidants or hemoproteins able to trap the radicals produced at their active center. J. Exp. Zool. 284:610-616, 1999. PMID- 10531548 TI - Ultracytochemical location of Na(+)/K(+)-atpase activity and effect of high salinity acclimation in gill and renal epithelia of the freshwater shrimp Macrobrachium olfersii (Crustacea, Decapoda). AB - Accumulation sites of lead phosphate reaction product consequent to Na(+)/K(+) ATPase activity in gill and renal epithelia of the freshwater shrimp Macrobrachium olfersii were located ultracytochemically by para-nitrophenyl phosphate hydrolysis and lead precipitation, and quantified per unit membrane area and cytoplasmic volume. In shrimps in freshwater (<0.5 per thousand S, 20 mOsm/kg H(2)O, 0.7 mEq Na(+)/liter), numerous sites of electron-dense, Na(+)/K(+) ATPase reaction product accumulation were demonstrated in the membrane invaginations of the mitochondria-rich, intralamellar septal cells (12.5 +/- 1.7 sites/microm(2) membrane, 179 +/- 22 sites/microm(3) cytoplasm, mean+/- SEM, N 0.01). The blastocyst development was decreased from 49.8% in the control group to 20.3, 7.8, 3.4, and 2.3% with sperm exposed to doses of 5, 10, 50, and 100 GY, respectively. Of the transferred blastocyst in the control group, 69.8% were implanted and 33.9% developed into live fetuses. These rates were 57.1 and 21. 4%, 20 and 0% when sperm were exposed to doses of 5 and 10 GY with a significant difference (P < 0.01). The present study clearly shows that DNA damaged sperm (regardless of degree of damage) have the ability to fertilize the oocyte, but that embryonic development is very much related to the degree of DNA damage. However, the oocyte has the capacity to repair DNA damage of sperm when it is damaged less than 8%. Damage beyond this level will result in low rate of embryonic development and high early pregnancy loss. J. Exp. Zool. 284:696-704, 1999. PMID- 10531555 TI - Developmental fate of the yolk protein lipovitellin in embryos and larvae of winter flounder, Pleuronectes americanus. AB - The developmental fate of the vitellogenin-derived yolk protein, lipovitellin (Lv), was investigated in winter flounder embryos and yolk-sac larvae. Since Lv is present as only one major polypeptide in ovulated winter flounder eggs, unlike the multiple yolk polypeptides found in the mature eggs of most teleosts, this system is presented as a simpler model of yolk protein structure and utilization during teleostean development. Winter flounder Lv is cleaved during embryogenesis from a 94 kD polypeptide at fertilization to 67 kD and 26 kD polypeptides at hatching. The rate of this proteolytic processing is slow during early embryonic development, but enters a more rapid phase between days 8 and 12 post fertilization in embryos reared at 4-5 degrees C, and approaches 50% completion at day 10. Lv processing is essentially complete 3 days before hatching; nevertheless, major degradation of the Lv peptide by the developing winter flounder does not occur until after hatching. The Stokes radius of Lv changes only moderately following processing, from 4.50 nm in unfertilized eggs to 4.19 nm in late embryos and newly hatched larvae, whereas the processed Lv retains its heat stability relative to other yolk polypeptides. Nearly 50% of its lipid content, however, is released from the Lv particle during embryogenesis, concomitant with cleavage of the Lv 94 kD polypeptide. Lv processing may thus render a portion of the yolk protein-associated lipid more accessible to the developing embryo, whereas other yolk components are retained for later use by the winter flounder larva. Alternately, removal of lipid may lead to proteolytic vulnerability of the Lv polypeptide. In either case, only a portion of the lipid moiety of the Lv particle appears to play a significant nutritive role for the embryo, whereas its protein component is reserved for larval use. J. Exp. Zool. 284:686-695, 1999. PMID- 10531557 TI - Differential expression of SOX9 in gonads of the sea turtle Lepidochelys olivacea at male- or female-promoting temperatures. AB - In mouse and chick embryos, the SOX9 gene is down-regulated in genetic females whereas in genetic males it remains in the Sertoli cells. We studied the distribution of SOX9 protein in developing genital ridges of embryos of the sea turtle Lepidochelys olivacea incubated at male- or female-promoting temperatures, using the antibody for detection. At stages 22-24, cells in medullary cords show SOX9 positive nuclei, while coelomic epithelial cells appear negative. At stage 25 however, most medullary cells are SOX9 negative and at the female-promoting temperature, and from stage 26 onwards, SOX9 protein is not detected. At the male promoting temperature, medullary cords remain SOX9-positive at all stages. These results suggest that SOX9 is up-regulated in Sertoli cells irrespective of primary sex-determining switch. Sex is irreversibly determined at stage 24 or 26 at the male- or female-promoting temperature, respectively (Merchant-Larios et al.,'97). The present results suggest that there is a correlation between SOX9 expression and sex determination in the olive ridley. At the male-promoting temperature, Sertoli cells expressing SOX9 become committed at stage 24 and male sex is determined, whereas at the female-promoting temperature, SOX9 is down regulated at stage 26 and female sex is determined. J. Exp. Zool. 284:705-710, 1999. PMID- 10531559 TI - Introduction to the special issue: the 1898 cambridge anthropological expedition to the Torres strait. PMID- 10531558 TI - Possible role of non-classical chromatophorotropins on the regulation of the crustacean erythrophore. AB - Two neuropeptides, the pigment dispersing hormone (PDH) and the pigment concentrating hormone (PCH), are well known to respectively promote centrifugal and centripetal granule translocation in the freshwater shrimp Macrobrachium potiuna erythrophores. Herein, we demonstrate for the first time the effects of crustacean non-classical chromatophorotropins on the pigment migration in M. potiuna erythrophores. Although proctolin, 20-hydroxyecdisone (20HE), and melatonin were ineffective, the crustacean cardioactive peptide (CCAP) was a full agonist, inducing pigment dispersion in a dose-dependent manner with EC(50) of 9.5. 10(-7) M. In addition, concentrations of CCAP lower than the minimal effective dose (10(-8) and 10(-7) M) decreased the PCH-induced aggregation, shifting rightward the dose-response curve (DRC) to PCH 2.2- and 29-fold, respectively. Surprisingly, melatonin (10(-7) and 10(-6) M) also shifted to the right 8.7- and 46.5-fold, respectively, the DRC to PCH. In conclusion, our data demonstrate that besides PCH and PDH, CCAP and melatonin also regulate the pigment migration within the crustacean erythrophore. J. Exp. Zool. 284:711-716, 1999. PMID- 10531560 TI - Dire straits: the divisive legacy of the 1898 Cambridge anthropological expedition. AB - Bartlett's claim that the Cambridge anthropological expedition of 1898 to the Torres Strait "put a social and ethnological stamp upon Cambridge psychology" does not bear close examination. Rivers pursued his interests in both anthropology and psychology but came to regard them as largely independent pursuits. Myers, through the influence of Rivers, came to identify himself primarily as a psychologist. McDougall was very quickly marginalized. There were two occasions when the promise of the Torres Strait began to be fulfilled: first, the reunion of Rivers, Myers, McDougall, and Seligman, all medically trained, at Maghull Hospital to help in the treatment of shell-shocked soldiers; second, Bartlett's attempt, early in his career, to establish a sociocultural psychology. However, the remarkable "academy" at Maghull was disbanded at the end of the war, and Bartlett, in his attempt to promote the "upstart subject" of psychology at Cambridge, increasingly came to distance his department from social and ethnological concerns. There is a neglected legacy of the Torres Strait expedition, the curious belief that the methods of experimental psychology, and indeed psychophysics, could (somehow) be foundational to the human sciences. This legacy has served both to suggest that psychology must have something to do with anthropology, while perpetually deferring any actual integration between the two disciplines. PMID- 10531561 TI - W. H. R. Rivers and the war neuroses. AB - W. H. R. Rivers was the most famous member of the Cambridge Expedition to the Torres Strait. At the time, he was a physician and had an international reputation as a researcher in physiological psychology. The expedition signaled the beginning of his career in social anthropology, but also a long hiatus in his activities in medicine. His clinical interests revived during World War I. As an officer in the Royal Army Medical Corps (RAMC), Rivers became a leading proponent of "psychological medicine." Today, his war-time psychiatry is remembered mainly in association with his patient, Siegfried Sassoon. This article focuses on his wartime activities, his clinical practices, and his theories concerning the war neuroses and the unconscious. The currently popular view of Rivers as a quasi Freudian humanist is challenged. PMID- 10531562 TI - Gregory Bateson's lost world: the anthropology of Haddon and Rivers continued and deflected. AB - Gregory Bateson was one of the last and most distinguished products of the school of anthropology that Haddon and Rivers created in Cambridge after the Torres Strait Expedition. Beginning his career shortly after Rivers' death, Bateson used the interwar years to create a theoretical approach that continued and deflected that of Haddon and Rivers. His major ethnography from this period, Naven, evidenced his complex academic positioning between the legacy of Rivers and the new paradigm emerging around Malinowski and Radcliffe-Brown. After the Second World War, Bateson's intellectual project emerged as even closer to Rivers' in both psychological and evolutionary dimensions. PMID- 10531563 TI - John Dewey's unknown critique of marginal utility doctrine: instrumentalism, motivation, and values. AB - During the late nineteenth century, marginal utility theory became the dominant ideology of the new academic discipline of economics. This article explicates and discusses previously unexamined lecture notes prepared in 1913 by John Dewey on marginal utility doctrine. After briefly characterizing the state of marginal utility doctrine, Dewey's criticisms are shown to derive from his instrumentalism and general theory of value. In Dewey's view, marginalism privatized value and by doing so induced moral agnosticism, a condition of permanently suspended judgment regarding individual and social needs that was likely to undermine the foundations of a democratic community since it immunizes value against collective appraisal by public bodies. Moreover, while marginal utility theory represented a serious concerted attempt to deal with substantive problems of value in economics and the economy, to Dewey it was ultimately a form of apologetics for the existing mode of social relations. PMID- 10531564 TI - Briefly noted PMID- 10531565 TI - News and notes PMID- 10531566 TI - Investigation of leukemia cells from children with common acute lymphoblastic leukemia for genomic sequences of the primate polyomaviruses JC virus, BK virus, and simian virus 40. AB - BACKGROUND: An infectious etiology for childhood acute lymphoblastic leukemia (ALL) has long been suspected, although the characteristics of the putative childhood ALL-inducing agent(s) remain a mystery. We describe the testing of ALL leukemia cells for the presence of DNA sequences of the polyomavirus family: JC virus, BK virus, and simian virus 40 (SV40). PROCEDURE: Cryopreserved leukemia cells from 25 children between 2 and 5 years of age at the time of diagnosis and classified as having "common" ALL (B-precursor ALL expressing the CD19 and CD10 surface antigens) were tested for the presence of polyomavirus sequences using standard PCR methods. RESULTS: Human beta-globin gene sequences were detected in 22 of 25 leukemia specimens. However, polyomavirus sequences were not detected in any of the 22 specimens with amplifiable DNA. CONCLUSIONS: The inability to detect JC virus, BK virus, and SV40 virus DNA sequences in any of the 22 specimens with amplifiable DNA suggests that that these members of the polyomavirus family are unlikely to be causally associated with most childhood ALL. Published 1999 Wiley-Liss, Inc. PMID- 10531567 TI - Nonmetastatic pelvic Ewing sarcoma: report of the French society of pediatric oncology. AB - BACKGROUND: Since January, 1984, 59 children with histologically confirmed Ewing sarcoma of the pelvic bone have been treated with three successive chemotherapy protocols recommended by the French Society of Pediatric Oncology. The purpose of the current study was to evaluate the role of surgery and/or radiotherapy in local progression-free, disease-free, and overall survivals (LPFS, DFS, and OS, respectively). PROCEDURE: We retrospectively examined 59 children treated for nonmetastatic, pelvic Ewing sarcoma over the last 12 years. All were first treated with chemotherapy according to the current French protocol. Six patients developed progressive disease before local treatment and were excluded for local control and survival analysis. Local treatment was surgery alone in 17 cases, radiation therapy in 27 cases, and surgery plus radiation therapy in 9 cases. RESULTS: With a median of follow-up of 6.5 years, no significant differences in local control or survival were observed with the three chemotherapeutic protocols. Of the 53 patients evaluable for local control, 6 relapsed locally only, 8 had local and distant relapses, and 9 had distant metastases only. The 5 year OS rate was worst for patients with radiotherapy alone compared to those with surgery or combined modality treatment (44 % vs. 72 %, P = 0.043). The 5 year LPFS and DFS rates were worst in the radiotherapy-alone group but not significantly (63% vs. 79%, P = 0. 22 and 42% vs 71%, P =0.07, respectively). The importance of surgery to OS and DFS was confirmed by multivariate analysis (P = 0.026 and P = 0.048, respectively). One surviving patient was diagnosed with in field fibrosarcoma, which was presumably radiation induced. CONCLUSIONS: Despite intensive, multiagent chemotherapy, survival from pelvic Ewing sarcoma has not improved over the past decade; however, the survival rate does not seem to be worse than that from Ewing sarcoma at other locations, insofar as at least 50% of the patients were cured. Surgery or a combination of surgery and radiation therapy are the best local treatment; exclusive radiation therapy should be reserved for patients with inoperable lesions or partially or nonchemosensitive tumors or when surgery would be an amputation. PMID- 10531568 TI - Phase I study of high-dose thiotepa with busulfan, etoposide, and autologous stem cell support in children with disseminated solid tumors. AB - BACKGROUND: The aim of this phase I study was to define the maximum tolerated dose (MTD) of thiotepa (TT), administered with busulfan (BU) 480 mg/m(2) and etoposide 2,400 mg/m(2), followed by autologous bone marrow transplantation (ABMT) or peripheral blood stem cell transplantation (APBSCT) support in children with solid tumors either disseminated at diagnosis or after relapse. PROCEDURE: Nineteen patients, between 2 and 16 years of age, received a high-dose chemotherapy regimen including escalating doses of TT starting from 150 mg/m(2). Subsequent dose escalation was determined by a modified Fibonacci scheme. Whenever one patient at one dosage level showed a grade III or grade IV reversible toxicity, additional patients were admitted (one by one) up to a maximum number of 6. Upon observing grade III or IV reversible toxicity in two or more systems, in 3 of the 6 patients, no further escalation was performed, and the corresponding dosage was taken as the MTD. WHO criteria were adopted to assess grade of toxicity. RESULTS: All patients had hematological recovery; and neutrophils and platelet engraftment were observed after median times of 12 and 29 days from stem cell infusion, respectively. The MTD of TT was determined to be 750 mg/ m(2). At this level, 3 of 6 patients experienced grade III mucositis and/or grade III gastrointestinal toxicity. No patient died of treatment-related toxicity. CONCLUSIONS: A dose of 750 mg/m(2) TT is the MTD when it is associated with BU 480 mg/m(2) and etoposide 2, 400 mg/m(2). This ablative regimen represents a feasible and tolerable combination for high-dose chemotherapy followed by hematopoietic stem cell rescue (HSCR). Phase II studies in children with poor-prognosis solid tumors are required to evaluate the effectiveness of this treatment. PMID- 10531569 TI - Disturbance in bone turnover in children with a malignancy at completion of chemotherapy. AB - BACKGROUND: Osteoporosis and pathological fractures have been observed in children with a malignancy. The mechanisms of osteopenia in childhood malignancies have not been well established. The purpose of the present study was to evaluate changes in bone turnover and in bone hormonal metabolism in children with a malignancy at completion of their chemotherapy. PROCEDURE: Serum levels of human intact osteocalcin, type I collagen carboxyterminal propeptide (PICP), type I collagen carboxyterminal telopeptide (ICTP), 25-hydroxyvitamin D [25-(OH)-D], 1,25-dihydroxyvitamin D [1, 25-(OH)(2)-D], intact parathyroid hormone, insulin like growth factor I (IGF-I), IGF binding protein 3 (IGFBP-3), alkaline phosphatase, calcium, and phosphate were analyzed in 22 children with acute lymphoblastic leukemia and in 26 children with other malignancies. Results were expressed as Z-scores [mean (95% confidence intervals)] relative to healthy Caucasian-children. RESULTS: The marker of collagen degradation (ICTP) was significantly increased [1.43 (1.10-1.76), P < 0.0001] compared to reference values, whereas the markers of bone formation (PICP, osteocalcin) were not changed [0.07 (-0.55 to 0.49), 0.35 (-0.05 to 0.74), respectively, NS]. Serum 25 (OH)-D, 1,25-(OH)(2)-D, and calcium were significantly reduced [-0.65 (-0.87 to 0.42), -0.68 (-0.92 to -0. 42), -1.42 (-1.80 to -1.04), P < 0.0001, respectively]. CONCLUSIONS: Disturbance in bone turnover with low serum 25-(OH) D, 1, 25-(OH)(2)-D, and calcium was observed in children with a malignancy at completion of their chemotherapy. A controlled study determining the possible benefits of vitamin D and calcium supplementation on bone turnover could be considered in these patients. PMID- 10531570 TI - Hepatotoxicity in patients treated according to the nephroblastoma trial and study SIOP-9/GPOH. AB - BACKGROUND: A major problem for children receiving Wilms tumor (WT) chemotherapy is hepatotoxicity, which may even be life-threatening. Dactinomycin (AMD) has been shown to be an important factor, as has abdominal irradiation. PROCEDURE: In the nephroblastoma trial and study SIOP-9 (SIOP-9) two different regimens for the application of AMD were used (standard dose over 3-5 days vs. double dose on a single day). In children at increased risk for local relapse, postoperative abdominal irradiation was given. We analyzed the influence of AMD and radiotherapy on the development of hepatotoxicity in 481 children treated in centers of the German Paediatric Oncology and Haematology Society (GPOH). A special questionaire was sent out for all patients with reduced treatment or delay of more than 1 week because of hepatotoxicity. Because SIOP and the National Wilms Tumor Study (NWTS) used different criteria to asses hepatotoxicity,we applied both definitions. RESULTS: All 72 cases of mild or severe hepatotoxicity occurred during treatment with AMD over 3-5 days with the standard dose (9.4-22.5 microgram/kg/week) compared to none in the group receiving a double dose on 1 day (3.75-8 microgram/kg/week; P < 0.001). Irradiation of the right abdomen, including parts of the liver, enhanced liver toxicity significantly, with a relative risk (RR) of 2.6 (P < 0.003). Preoperative liver toxicity was more frequent in smaller children (P = 0.02) and especially if no dose reduction was done in children with body weight of less than 12 kg (RR 5.3, P = 0.01). If severe liver toxicity was defined according to NWTS criteria, 10% of all treated patients were affected compared to 4.8% if McDonald's criteria for hepatic veno-occlusive disease (VOD) were applied. CONCLUSIONS: To diminish the hepatotoxicity of WT treatment, AMD dose intensity should be reduced (below 10 microgram/kg per week), especially in smaller children or when the liver is irradiated. PMID- 10531571 TI - Family history of cancer in children and young adults with colorectal cancer. AB - BACKGROUND: Family history of colorectal cancer among adult patients has been reported in the literature. Although extremely rare in children, colorectal cancer in this population may represent a unique group in whom genetic factors play a significant etiologic role. The aim of the present study was to assess genetic contribution, as measured by family history, to the development of colorectal cancer in probands under 21 years of age at diagnosis. PROCEDURE: Detailed family histories were obtained from surviving patients or their parents. The risk [standardized incidence ratio (SIR)] of cancer in the relatives was calculated by comparing the observed and the expected incidence based on rates in the general population and person-years at risk. RESULTS: Twenty-five patients (median age at diagnosis 15 years) diagnosed with colorectal cancer at St. Jude Children's Research Center since 1964 or their surviving next of kin were available for interview. The 461 relatives contributed 18,908 person-years of follow-up. Statistically significant increased risk of colorectal cancer was present among all relatives (SIR = 6.0, 95% CI, 2.7-10.6), and the increased risk of colorectal cancer was confined to relatives of probands who were under 15 years of age at diagnosis (SIR = 10.0, 95% CI, 4.5-17.6). In addition, there was an excess of uterine/cervical cancer among all female relatives (SIR = 6.5, 95% CI, 3.2-10.9). CONCLUSIONS: The observed excess of colorectal cancer, in relatives of younger probands, suggests the need to examine these kindreds for genetic instability resulting from defects in mismatch repair genes to characterize further the patterns of risk observed. PMID- 10531573 TI - Cyclosporine A therapy for multisystem langerhans cell histiocytosis. AB - BACKGROUND: Treatment of multisystem Langerhans cell histiocytosis (LCH) remains difficult. Various regimens of single and multiagent chemotherapy have been used, but a significant proportion of patients fail to respond to treatment. PROCEDURE: We have evaluated the use of cyclosporine A (CSA) in a controlled group of patients, who had received a systematic primary therapy (LCH-I). Patients received CSA either as a single agent (10 patients) or in combination with vinblastine, etoposide, prednisolone, and/or antithymocyte globulin (16 patients). RESULTS: Among the total of 26 patients treated, a single patient developed a complete response and three a partial response, whereas 85% (22 patients) had no response to CSA. CONCLUSIONS: CSA is at best of limited value in the treatment of patients with multisystem LCH, particularly those who had progressive disease while receiving chemotherapy. PMID- 10531572 TI - Nucleoside analogues in the therapy of Langerhans cell histiocytosis: a survey of members of the histiocyte society and review of the literature. AB - BACKGROUND: Previous reports have suggested activity of the nucleoside analogues 2-chlorodeoxyadenosine (2-CdA) and 2'-deoxycoformycin (2'-DCF) in Langerhans cell histiocytosis (LCH). PROCEDURE: To assess the efficacy of 2-CdA and 2'-DCF as salvage therapy for LCH, a survey of members of the Histiocyte Society and a literature review were undertaken. Twenty-three patients treated with 2-CdA and 4 treated with 2'-DCF were found, age range 2 months to 49 years. RESULTS: All 15 survey patients had multiorgan involvement, and 14 were heavily pretreated. Doses of 2-CdA ranged from 0.1 mg/kg/day continuous infusion for 5-7 days (majority of patients) to 13 mg/m(2)/day for 5 days, for 1-6 courses. One of the 15 patients had an early death, 5 had no response (NR), 3 had partial response (PR), and 6 achieved complete response (CR). Among 8 published patients, 7 achieved stable CR and 1 NR. Among 4 patients treated with 2'-DCF (4 mg/m(2)/week for 8 weeks then q 2 weekly), 2 achieved CR for 16+ and 18+ months and 2 PR for 2 and 5 months. Toxicity consisted mainly of combined myelo- and immunosuppression but no significant infections occurred and there were no toxic deaths. A cumulative thrombocytopenia was noted, which in 1 case took up to 6 months to resolve. Transient gastrointestinal toxicity and elevation of liver enzymes was seen, and 2 patients developed renal tubular acidosis. The peripheral neuropathy reported in adult patients receiving high doses was not seen. CONCLUSIONS: 2-CdA and 2' DCF appear to have a useful role in LCH and are worthy of prospective trial in patients unresponsive to routine therapy. PMID- 10531574 TI - Neuropsychologic deficits in children with Langerhans cell histiocytosis. AB - BACKGROUND: Manifestations of Langerhans cell histiocytosis (LCH) in children range from only a rash, to bony lesions accompanied by pain, to major organ disease. When the central nervous system (CNS) is affected, the LCH patient may exhibit signs and symptoms of hypothalamic and pituitary dysfunction (most often resulting in diabetes insipidus or other endocrinopathies) or more global neurologic and neuropsychologic sequelae. Surprisingly, researchers have only recently begun to examine the neuropsychologic manifestations of the disease, but early findings suggest that they may, in fact, be significant in a small percentage of children with LCH. PROCEDURE: We evaluated two CNS-positive patients with LCH and long-term intermittent treatments, using extensive neuropsychologic assessments, including intellectual functioning, memory, visual motor functioning, attention and concentration, sensory and motor performance, and gross academic achievement. Objective measures of behavior were obtained through parental report. Neuroradiologic imaging was obtained concurrently with the neuropsychologic evaluations. RESULTS: The neuropsychologic assessments indicated significant deficits in a number of the measured areas of functioning. Global cognitive deficiencies in full-scale IQ were identified, as were deficits in memory, attention/concentration, and perceptual-organizational capabilities. Similarities were noted in the patterns of deficits obtained with both patients, despite differences in the pathophysiology of their disease. Behavioral functioning in both children had suffered, presumably in relation to the neuropsychologic deficits. There were radiologic findings of gross cerebellar white matter damage in one patient, in addition to focal (e.g., hypothalamic) lesions in the other. CONCLUSIONS: LCH has an adverse impact on cognitive functions in some children with evidence of CNS involvement, and further study into the etiology, incidence, and means of remedial intervention is needed. PMID- 10531575 TI - Concurrent chemotherapy and radiotherapy: an affordable important improvement in the treatment of advanced cervical cancer. PMID- 10531576 TI - Nephroblastoma in the adult. PMID- 10531577 TI - Successful clinical response to irinotecan in desmoplastic round blue cell tumor. PMID- 10531578 TI - Fulminant fatal Strongyloides stercoralis infection in a postchemotherapy immunosuppressed cancer patient. PMID- 10531579 TI - Severe dactinomycin overdose in an 18-month-old child. PMID- 10531580 TI - Solid-pseudopapillary tumor of the pancreas. PMID- 10531581 TI - The histiocytoses come of age PMID- 10531582 TI - Music Educators' Perceptions Regarding the Inclusion of Students with Severe Disabilities in Music Classrooms. AB - The purpose of the present study was to: (a) examine music educators' perceptions regarding the practice of full inclusion, (b) conduct a descriptive analysis of their perceptions, and (c) compare and contrast choral, instrumental, and general music educators' perceptions regarding the practice of inclusion. The data collection technique used in the study was personal interviewing. Participants were instrumental, choral, and general music educators (N = 35) in a midwestern school district that supports the practice of full inclusion. Written transcripts of the 35 interviews were coded and analyzed for recurring themes and patterns using content analysis. Music educators identified 13 critical issues related to the inclusion of students with disabilities. The need for collaboration or consultation with special educators, music therapists, or others knowledgeable about students with disabilities was identified as a critical issue by nearly all of the participants. Many participants also identified as critical issues: the need for more information about the students included in their music classroom, the amount of time required to successfully include students with disabilities, and the range of abilities often found in the inclusive classroom. Most music educators felt that inclusion has had a positive impact on students both with and without disabilities, though reservations were also expressed by some of the music educators. Subject responses were also analyzed for frequency of: disabilities mentioned, positive and negative statements made regarding inclusion, personal anecdotes, and references to music therapy. Suggestions are given for the role music therapists can play in facilitating the inclusion of students with disabilities. PMID- 10531583 TI - Music Therapy in School Settings: Current Practice. AB - The practice of music therapy in school settings was the focus of this study. Survey forms were mailed to 244 NAMT members who indicated school setting as their place of employment. A total of 190 forms were received, 138 of which fit the qualifications for inclusion and were included in the data summaries. Greater numbers of respondents lived in Texas (21), New York (17), and Michigan (11), and were employed full-time (60&percent;). Employers were more typically school systems (53&percent;) for the highest percentage of full-time respondents (80&percent;), and self-employers (25&percent;) for the highest percentage of part-time respondents (80&percent;). A considerably higher percentage of time was spent each week in direct service delivery (62&percent;) than in consultation (13&percent;), travel (18&percent;), documentation (11&percent;), or preparation (14&percent;). Over 40&percent; of the respondents had been music therapists for more than 8 years, but not necessarily in their current positions. Almost 40&percent; needed a valid teaching certificate for employment, while over 50&percent; currently held one. Respondents most frequently worked with persons who were developmentally disabled (80&percent;). The impact of employer and the inclusion movement on professional practice issues was discussed, as were possible trends in the practice of music therapy in school settings. PMID- 10531584 TI - The Effect of Music on the Self-Injurious Behavior of an Adult Female with Severe Developmental Disabilities. AB - This study compared the effects of contingent blocking, music listening, water play, and instrument playing on self-injurious behavior. The subject was a 23 year-old female with severe developmental disabilities, including communication deficits, motor deficits, and visual impairment. A single subject reversal design with four conditions (ABACADA) was used. Subject was videotaped 10 minutes before and 10 minutes after the interventions to study changes in rate and variability of three behaviors: teeth-grinding, mouth-scratching, and head-hitting. Visual analysis showed a strong downward trend after music listening for teeth-grinding, and no change over all conditions for mouth-scratching. Subject did not exhibit any preintervention head-hitting during both music conditions, and therefore showed no change in behavior. Subject responded to water play and contingent blocking by reducing head-hitting. PMID- 10531585 TI - Marian Erdman: Contributions of an American Red Cross Hospital Recreation Worker. AB - The purpose of this historical study was to investigate the life experiences of Marian Erdman, specifically her use of music in military hospitals. The work of Marian Erdman, as a hospital recreation worker for the American Red Cross from 1945 through 1948, provides personal insights into the role of musicians and music in military hospitals at the close of World War II. Questions addressed by this biographical study include (1) What were the professional experiences of Marian Erdman? (2) What were her views regarding the purpose and function of music in hospitals during the post-war era? (3) Did the Red Cross provide guidelines regarding the purpose and use of music with patients?, and (4) How did the events of the mid-1940s contribute to the formation of music therapy as an organized profession? PMID- 10531586 TI - Diagnosing heart failure by the Valsalva maneuver : Isn't It finally time? PMID- 10531587 TI - Xinafoic acid is not a solvent for salmeterol. PMID- 10531588 TI - Propofol containing sulfite-potential for injury. PMID- 10531590 TI - Aging lung and possible animal models. PMID- 10531589 TI - Carcinoid-a diagnostic and therapeutic dilemma. PMID- 10531591 TI - Fatty acid profiles needed for dairy foods from the United States. PMID- 10531592 TI - Autolysis of the proteinase from Pseudomonas fluorescens. AB - The gene encoding the proteinase from Pseudomonas fluorescens was cloned and sequenced in an effort to identify the cleavage sites involved in its autolysis at 50 degrees C. A single open reading frame consisting of 1449 nucleotides, encoding a protein of 482 amino acids, was found. Analysis of the N-terminal amino acid sequence of the purified proteinase indicated the presence of a prosequence consisting of 13 amino acid residues. The molecular weight of the mature protein was calculated as 48,900 based on the deduced amino acid sequence, which was consistent with that of the purified proteinase as determined by sodium dodecylsulfate-PAGE. Greater than 90% loss of proteolytic activity was observed upon heating at 50 degrees C for 2 min compared with the unheated enzyme. Incubation of the proteinase at 50 degrees C led to disappearance of the intact enzyme, as shown by sodium dodecyl sulfate-PAGE, whereas it was stable in the presence of the protease inhibitor o-phenanthroline. Autolytic fragments were fractionated by reverse-phase HPLC and subjected to N-terminal amino acid sequence analysis in an effort to determine the cleavage sites. The cleavage profile was not definitive; however, amino acid residues with small side chain groups, such as glycine or alanine, were frequently found adjacent to the cleavage sites. PMID- 10531593 TI - Alternative splicing of lactophorin mRNA from lactating mammary gland of the camel (Camelus dromedarius). AB - The objective of this study was to determine the corrected structure of lactophorin, a major whey protein in camel milk. The protein had 60.4% amino acid sequence identity to a proteose peptone component 3 protein from bovine whey and 30.3% identity to the glycosylation-dependent cell adhesion molecule 1 in mice. The N-terminal heterogeneity of the protein was a result of alternative mRNA splicing. About 75% of the protein was expressed as a long variant A with 137 amino acid residues and a molecular mass of 15.7 kDa; about 25% was as a short variant B with 122 amino acid residues and a molecular mass of 13.8 kDa. Both proteins are probably threefold phosphorylated. In contrast to the related proteins, no glycosylation was found in camel lactophorin. Because of this difference, specific interaction with carbohydrate binding proteins, as reported for the murine protein, can be excluded, and a function of the protein other than cell recognition or rotaviral inhibition is proposed. The concentration of lactophorin in camel milk was found to be about three times higher than the concentration of the bovine homologue in bovine milk. Pronounced similarities existed between the primary and secondary structures of bovine and camel proteins. We speculated that camel lactophorin has a similar function to that of bovine protein in milk, which is supposed to be the prevention of fat globule aggregation and the inhibition of spontaneous lipolysis by lipoprotein lipase. PMID- 10531594 TI - Efficacy of a biological response modifier in preventing Staphylococcus aureus intramammary infections after calving. AB - A change in the epidemiology of mastitis in recent years has emphasized the role of the udder immune system in the pathogenesis of Staphylococcus aureus. Therefore, if the bovine or udder immune capability could be enhanced, susceptibility to Staph. aureus could be reduced and antibiotic efficacy could be increased. Immune system defense mechanisms could be enhanced by vaccination and by biological response modifiers. Within this latter group, a biological response modifier obtained from Parapox ovis that was attenuated over 200 tissue culture passages was recently developed and commercialized in some European countries. This study reports the results of a field trial on the efficacy of this biological response modifier in reducing Staph. aureus intramammary infection (IMI) after calving in primiparous and pluriparous cows. The trial included 106 cows sampled six times (55 cows from herd A and 51 from herd B) for a total of 2544 quarter milk samples. The analysis of IMI prevalence showed that 25.09% of samples were bacteriologically positive in the placebo group, and 23.17% of the positive samples were observed in the biological response modifier group. Staphylococcus aureus IMI had a frequency of 11.44% in the placebo group and 6.00% in the biological response modifier group. The dynamic of the hazards showed significantly lower rates in the biological response modifier group than in the placebo group (risk ratio = 0.47). Treatment with the parapox-containing biological response modifier showed significant reduction of Staph. aureus IMI around calving, and this reduction was attributed to an increase in immune defenses. PMID- 10531595 TI - The natural food grade inhibitor, lacticin 3147, reduced the incidence of mastitis after experimental challenge with Streptococcus dysgalactiae in nonlactating dairy cows. AB - Lacticin 3147 is a broad-spectrum bacteriocin produced by the food-grade organism Lactococcus lactis. Lacticin 3147 is active at a neutral pH and has been shown to be bactericidal to streptococci and staphylococci in vitro. The effectiveness of an intramammary teat seal formulation, and a teat seal containing lacticin 3147 was evaluated at drying off in 68 uninfected quarters of 18 cows. Following infusion of either teat seal or lacticin 3147 combined with teat seal, a deliberate infection challenge of Streptococcus dysgalactiae (approximately equal to 1.5 x 10(4) cfu per teat) was administered by direct inoculation into the teat sinus. During an 8-d experimental period following inoculation, 61% of control quarters and 6% of the treatment quarters either developed clinical mastitis or were shedding the challenge organism. Randomly amplified polymorphic DNA polymerase chain reaction genetic typing was used to confirm that both the new infections and the bacteria surviving in the teats at the end of the experiment were the challenge strain. The combination of teat seal and lacticin 3147 was well tolerated within the udder and elicited only a temporary increase in somatic cell count to 5.7 x 10(5)/ml (88 h after infusion) in a previously uninfected lactating udder quarter. Therefore, we concluded that this nonantibiotic approach to mastitis prevention may contribute to a reduction in the routine application of antibiotics at drying off in the future. PMID- 10531596 TI - Evaluation of ruminally protected methionine and lysine or blood meal and fish meal as protein sources for lactating Holsteins. AB - Forty lactating Holstein cows averaging 55 days in milk were used in a randomized block designed experiment to evaluate the effectiveness of ruminally protected Met and Lys compared with that of ruminally undegradable protein for supporting lactation. Cows were fed total mixed diets for 15 wk. Diets were formulated to be isonitrogenous with the same base ingredients resulting in base crude protein percentage of 15.5. Supplemental crude protein supplied by urea, soybean meal, or a 50:50 (wt/wt) mixture of fish and blood meal increased total dietary nitrogen to 18.0% of diet DM. Two additional diets consisted of the basal diets soybean meal and urea, which were supplemented with ruminally protected DL-Met and Lys HCL at 10 and 25 g/d, respectively (soybean meal + amino acids (AA), urea + AA). Mean measures of dry matter intake, milk yield, milk protein percentage, and milk fat percentage were not affected by protein supplement. Milk protein yield, milk fat yield, casein yield, and casein percentage also were not affected by source of supplemental protein. Results indicate that at the level of crude protein intake relative to milk production in this experiment, the source of protein did not affect lactational performance. PMID- 10531597 TI - Fibrolytic enzyme supplements for dairy cows in early lactation. AB - Twenty multiparous lactating Holstein cows in early lactation were used to investigate effects of exogenous fibrolytic enzyme supplementation on dry matter intake, milk production, and digestibility. Cows were blocked according to parity, expected calving date, and milk yield in the previous lactation, and then randomly assigned after calving to two treatments: control or enzyme. The enzyme mixture, which contained mainly xylanase and cellulase activities (Pro-Mote, Biovance Technol. Inc., Omaha, NE), was added to the concentrate to supply 1.3 g/kg of total mixed ration (dry matter basis). The total mixed rations contained 24% corn silage, 15% alfalfa hay, and 61% barley concentrate (dry matter basis) and were offered for ad libitum intake. Enzyme addition did not affect dry matter intake. However, total digestibility of nutrients, determined using Cr2O3, was dramatically increased by enzyme treatment (dry matter, 61.7 vs. 69.1%; neutral detergent fiber, 42.5 vs. 51.0%; acid detergent fiber, 31.7 vs. 41.9%; crude protein, 61.7 vs. 69.8%). Consequently, milk yield tended to increase (35.9 vs. 39.5 kg/d). Percentage of milk fat was lower, and percentages of milk protein tended to be lower for cows fed a diet supplemented with enzymes, such that component yields were similar for cows fed either diet. Energy deficiency was numerically lower for cows fed a diet supplemented with enzymes than for cows fed the control diet (-3.62 vs. -3.33 Mcal/d). Supplementing dairy cow diets with a fibrolytic enzyme mixture has the potential to enhance milk yield and nutrient digestibility of cows in early lactation without changing feed intake. PMID- 10531598 TI - Brown midrib sorghum in diets for lactating dairy cows. AB - In Experiment 1, 16 Holstein cows were assigned to one of four diets in replicated 4 x 4 Latin squares with 4-wk periods to measure dietary effect on short-term lactational performance. Additionally, 3 fistulated cows were assigned to the same diets in a 3 x 4 Youden square design with 4-wk periods to measure ruminal rate and extent of fiber digestion, fractional passage rate of fiber, ruminal pH, and concentration of volatile fatty acids. Diets comprised 65% of brown midrib (BMR) forage sorghum, standard forage sorghum, alfalfa or corn silages and 35% concentrate. Experiment 2 was conducted with 30 Holstein cows in early lactation to evaluate the same BMR sorghum hybrid in a 10-wk study with 35.3% standard sorghum, BMR sorghum, or corn silages as dietary treatments. Milk production was significantly higher for brown midrib than for standard sorghum in Experiment 1. Ruminal pH and acetate to propionate ratio did not differ among diets. The fractional passage rate of silage was not significantly different among the forages. In situ extent of ruminal fiber digestion was significantly higher for BMR than for standard sorghum, but rate of fiber digestion was not different. Similarly, in Experiment 2, in vitro extent of fiber digestion was significantly higher for BMR sorghum than for standard sorghum. Dry matter intake and body condition score were not significantly different between cows fed BMR and standard sorghum, but cows fed BMR sorghum resulted in long-term milk production greater than cows fed standard sorghum and similar to cows fed corn silage. PMID- 10531599 TI - Evaluation of tropical grasses for milk production by dual-purpose cows in tropical Mexico. AB - Two experiments using the Cornell Net Carbohydrate and Protein System were conducted to characterize the carbohydrate and protein fractions and corresponding rates of digestion of 15 tropical pasture grasses and to evaluate their ability to support milk production by dual-purpose cows. In the first experiment, ranges in carbohydrate and protein fractions of 15 grasses at 35 to 42 d of regrowth were: neutral detergent fiber (NDF) 63.5 to 74.9% of DM; permanganate lignin 4.7 to 7.8% of NDF; CP 5.5 to 11.9% of DM; and soluble protein 15.1 to 44.1% of crude protein (CP). The ranges of rates of digestion expressed as percent per hour were neutral detergent solubles (7.5 to 27.4); NDF (3.8 to 8.4); and neutral detergent insoluble protein (2.9 to 9.5). Predictions of the amount of milk that could be produced based on the amount of metabolizable energy supplied by the diet decreased 35% when NDF increased from 60 to 80%, and increased 88% when the rate of digestion of NDF increased from 3 to 6%/h. The milk production that could be sustained based on metabolizable protein in the diet doubled as CP increased from 4 to 12%. In the second experiment, nitrogen fertilization reduced NDF 7.3% and increased CP 84% without changing protein solubility, resulting in increased rumen nitrogen and metabolizable protein balances. With all forages, the Cornell Net Carbohydrate and Protein System predicted that availability of metabolizable protein would limit milk production. Predicted microbial growth was limited by ruminally available protein rather than by available carbohydrate. PMID- 10531600 TI - Conjugated linoleic acid content of milk from cows fed different diets. AB - Conjugated linoleic acid in milk was determined from cows fed different diets. In Experiment 1, cows were fed either normal or high oil corn and corn silage. Conjugated linoleic acid was 3.8 and 3.9 mg/g of milk fatty acids in normal and high oil treatments, respectively. In Experiment 2, cows consumed one-third, two thirds, or their entire feed from a permanent pasture. Alfalfa hay and concentrates supplied the balance of feed for the one-third and two-third pasture treatments. Conjugated linoleic acid was 8.9, 14.3, and 22.1 mg/g of milk fatty acids in the one-third, two-third, and all pasture treatments, respectively. Cows grazing pasture and receiving no supplemental feed had 500% more conjugated linoleic acid in milk fat than cows fed typical dairy diets (Experiment 1). In Experiment 3, cows were fed either a control diet containing 55% alfalfa silage and 45% grain, or similar diets supplemented with 3% fish meal, or 250 g of monensin/cow/per day, or fish meal and monensin together. Conjugated linoleic acid was 5.3, 8.6, 6.8, and 8.9 mg/g of milk fatty acids in the control, fish meal, monensin, and fish meal plus monensin treatments, respectively. In Experiment 4, cows were fed either finely chopped alfalfa hay (Treatment 1), or coarsely chopped alfalfa hay (Treatment 2) in a 50% forage and 50% grain diet, or 66.6% grass hay and 33.4% grain (Treatment 3), or 98.2% grass hay (Treatment 4). Conjugated linoleic acid was 7.3, 8.3, 9.0, and 7.9 mg/g of milk fatty acids in treatments 1 through 4, respectively. PMID- 10531601 TI - Influence of long-term feeding of limited amounts of phosphorus on dry matter intake, milk production, and body weight of dairy cows. AB - For almost two lactations, 24 high-yielding, multiparous dairy cows were fed a basal diet and concentrate mixtures with three different P concentrations. The basal diet consisted of grass (silage or artificially dried), corn silage, wet beet pulp, straw, and concentrates. The concentrate mixtures differed only in P content by varying the amount of monosodium phosphate. The number of cows and the amount of dietary P, expressed as a percentage of current recommendations in the Netherlands were: 6 cows, 100% (P100); 9 cows, 80% (P80); and 9 cows, 67% (P67). This resulted in dietary P concentrations of 3.3, 2.8, and 2.4 g/kg of dietary DM for the P100, P80, and P67 treatments, respectively. The trial lasted for 21 mo, including two lactations and two dry periods. Feed intake of the P67 group was reduced significantly during the first dry period. Dry matter intake, milk yield, and body weight were all reduced with the low P treatment during the second lactation. Phosphorus had no effect on reproductive performance. Between P100 and P80, no effect on any of the variables in this trial was observed. Results suggests that the diet with 2.8 g of P/kg of dietary DM proved to be sufficient to meet the P requirement of dairy cows producing approximately 9000 kg of milk per lactation. PMID- 10531602 TI - Impact of dairy farming on well water nitrate level and soil content of phosphorus and potassium. AB - The Cornell Teaching and Research Dairy Farm was used to study the historical influence of dairy farming on water quality and soil chemical properties. The farm has milked approximately 360 cows for the past 20 yr and is situated on 526 ha of cropland (390 ha utilized for dairy production) near Harford, New York. Mass nutrient balances (N, P, K) were constructed with historical data from 1979 and 1994 for the 390 ha used for dairy production. The amount of imported N increased more than 40% from 1979 to 1994, although there were year-to-year variations, depending on crop yields. Although nutrient balance (imported minus exported nutrients) as a percentage of imported nutrients on the farm remained relatively unchanged during this period, balance of N increased from 43.1 metric tonnes in 1979 to 66.0 metric tonnes in 1994. However, P and K remained about the same because of the reduced use of fertilizers in the 1990s. During the 15-yr period, total milk production increased more than 40% (2502 to 3604 metric tonnes from 1979 to 1994). Analysis of well water suggested that increasing amount of N balance on the farm resulted in increased well NO3-N concentration. The mean of five wells located in the corn fields increased from 3.3 to 7.0 mg/kg in NO3-N concentration, 70% of the EPA upper limit. Soil P increased from 6.0 to 24.0 (kg/ha) during the same period. Soil K did not change. Mass nutrient balances are important in determining the amount of nutrients remaining on farm. This study suggests N, P, and K balance can be used as an indicator of potential for increased NO3-N concentrations in wells and soil P and K levels, respectively. PMID- 10531603 TI - Genetic analysis of cow survival in the Israeli dairy cattle population. AB - The linear model method of VanRaden and Klaaskate for analyzing herd life was expanded. Information on conception and protein yield was included in the estimation of predicted herd life of Israeli Holsteins. Variance components were estimated by a multitrait animal model. Heritability was slightly higher for herd life than for number of parities, but genetic correlations were close to unity. Animal model heritability estimates of herd life were higher than were sire model estimates. The expected herd life of pregnant cows was 420 d greater than for open cows. Each kilogram of increase in protein yield increased expected herd life by 9.5 d. Heritability of expected herd life increased from 0.11 for cows 6 mo after first calving to 0.14 for cows 3 yr from first calving. The genetic correlation of expected and actual herd life increased from 0.87 for records cut after 6 mo to 0.99 for records cut 3 yr after first calving. Phenotypic correlations increased from 0.61 to 0.94. Sire genetic evaluations based on predicted herd life of live cows were strongly biased if all records were weighted equally, and evaluations derived by weighting incomplete records to account for the effects of current herd life on variance components were nearly unbiased. PMID- 10531604 TI - Genetic evaluation for length of productive life with censored records. AB - This study was conducted to investigate the impact of censoring on the accuracy of sire evaluation for the length of productive life estimated by means of survival analysis using simulated and real dairy cattle data from the Swiss Braunvieh population. Data were simulated under a Weibull model with two fixed effects and a random sire effect with a sire variance of 0.04. Two different family structures investigated were 1000 sires with 10 daughters each and 200 sires with 50 daughters each. Sires were assumed to be related through their sires. The reference data were generated assuming no censoring. Sire effects were estimated from the reference data with and without considering the relationships among sires and referred to as the estimated transmitting abilities (ETA) of sires. The impact of censoring on accuracy of ETA and ranking of sires was investigated by computing rank correlations among true and estimated sire effects and among estimated sire effects from the reference data and from several different data files with increased proportion of censored records. Estimated transmitting abilities were generally more accurate with a large number of daughters. The rank correlations among the ETA of sires from the data with censored records and the ETA from the reference data decreased with an increased proportion of censored records. Considering relationships among sires resulted in higher rank correlations when the proportion of censored records was large. With 50 daughters per sire, accuracy of 70% can be achieved approximately 2 yr after first calving of the daughters with about 50% censored records. With the real data, a rank correlation with the ETA of sires from the reference data of 0.70 to 0.80 can be achieved with about 65% of records censored and about 2.5 yr after the first calving of the youngest daughters of the sires. PMID- 10531605 TI - Prediction of parental dominance combinations for planned matings, methodology, and simulation results. AB - Optimal use of dominance information requires a mating system and predictions of specific combining abilities for each set of prospective parents. Current evaluation procedures provide such predictions only for a limited number of parents. A procedure is described that predicts the specific combining ability for any parents. In this procedure, for each set of parents and their ancestors, the additive relationship matrix is created as a dense matrix. This matrix is then used to create a parental dominance matrix in a sparse matrix form, in which the rows of the matrix correspond to all parental combinations for which predictions are already available. Each new prediction requires a solution of the system of equations with the parental dominance matrix as the left-hand side. The efficiency of the mating system that accounts for dominance was evaluated in a simulation study. The simulated data files varied with respect to proportion of males and females selected, proportion of cattle born through embryo transfer, and additive and dominance variance. Sires and dams were preselected based on the additive merits only, but specific matings were arranged based on the combined additive plus dominance merit. The response to selection with consideration of dominance increased from 3.8 to 16.6% of the response from one generation of additive selection. The response was greater when the additive variance was smaller, the dominance variance was larger, the intensity of additive selection was lower, and the proportion of full sibs was greater. Use of dominance in the mating system is feasible and results in an additional genetic response to selection. PMID- 10531606 TI - Persistency of lactation yield: a novel approach. AB - The objectives of our study were to propose a new, simple, and easy-to-understand definition for persistency of lactation yield and to develop a new mathematical model to describe a lactation curve, one that includes a measure for persistency of lactation yield according to the proposed definition. Our definition of persistency is the number of days during which the level of constant yield is maintained. No lactation model exists that includes a measure of persistency in terms of duration of time or that allows this measure of persistency to be derived from model parameters. It was necessary, therefore, to develop a new model to describe the lactation curve: [formula: see text] where yt = yield at time t, t1 = time at transition from increased yield to constant yield, yP = level of constant yield, b3 = rate of decline in yield from the end of constant yield to the end of lactation, and P = persistency of constant yield. These four parameters measured directly the important biological characteristics of a lactation curve. Two test day data sets, one for individual records from a high producing cow and one for average records from 17,607 cows, were used to illustrate the model and to estimate persistency. The proposed measure of persistency should be important for genetic selection because it might be desirable to select for increased persistency (e.g., for extended lactations) without increasing peak yield and, hence, subjecting the cow to undesirable stress. PMID- 10531607 TI - Genetic parameters for clinical mastitis, somatic cell score, production, udder type traits, and milking ease in first lactation Holsteins. AB - Genetic parameters were estimated by restricted maximum likelihood with an animal model on first lactation data of 29,284 French Holstein cows for clinical mastitis, lactation somatic cell score, milking ease, production, and nine udder type traits. The heritability was low for clinical mastitis (0.024), moderate for lactation somatic cell score (0.17) and milking ease (0.17), and ranged from 0.17 to 0.30 for type traits. A high (0.72) but lower than unity genetic correlation was found between clinical mastitis and lactation somatic cell score and indicated that both traits were genetically favorably associated. The antagonism with production was stronger for clinical mastitis than for lactation somatic cell score (genetic correlations 0.45 and 0.15, respectively). Udder depth, fore udder attachment, and udder balance were favorably associated with lactation somatic cell score and clinical mastitis with genetic correlations ranging from 0.29 to -0.46, whereas low correlations were found with teat length. Milking ease was found to be unfavorably correlated with lactation somatic cell score (genetic correlation 0.44) but not with clinical mastitis. PMID- 10531608 TI - Mathematical representations of correlations among yield traits and somatic cell score on test day. AB - Prediction of lactation yields and accuracies of yields for use in genetic evaluation can be improved by including information from test day correlations, especially for milk recording plans that vary in the numbers of milk weights recorded and component samples taken. Daily milk weights for 658 lactations of Canadian cows and monthly test records of milk, fat, and protein yields and somatic cell scores for 500,000 lactations of US cows were used to estimate phenotypic correlations between test days within herd-year. Correlations between daily yields for a designated interval between test days generally were highest for midlactation and were lowest for early and late lactation. Regression (two linear, two quadratic, and interaction effects) on mean DIM and interval between test days predicted correlations with a squared correlation of 0.94 for daily milk yields. Similar relationships were found for US monthly data. Variation in sampling was reduced, computer memory was minimized, and positive definiteness was guaranteed by fitting regressions on simply defined sources of correlation. An autoregressive matrix represented the within-trait correlations very well. The equations developed could be used to derive covariances and, subsequently, to estimate lactation yields and accuracies from combinations of individual daily milk, fat, and protein yields and somatic cell score. PMID- 10531609 TI - Analysis of environmental effects on test day milk yields of Sarda dairy ewes. AB - Temporal evolution of Sarda ewes milk production was analyzed with mixed linear models and factor analysis to investigate environmental effects on the shape of lactation curves and on milk yields during different lactation stages. Parity, year of production, level of altitude, and flock within altitude affected milk yield, whereas the effect of type of lambing was not significant. Lactation curves pertaining to the three altitudes were clearly different. Factor analysis suggested an independence among yields recorded during different phases of lactation and a specific behavior of (co)variances structure among test day yields obtained in mountains compared with hills and plains. PMID- 10531610 TI - Interaction between milk yield of Holstein cows in Mexico and the United States. AB - Genotype by environment interaction for milk yield was investigated by analyzing 55,162 mature equivalent, first lactation records of daughters from 1339 Holstein sires in Mexico and 499,401 daughters from 663 Holstein sires in the northeastern US. There were 474 US sires in common. Herd-year standard deviation was used to define non-overlapping high (> or = 1600 kg) and low (< or = 1300 kg) Mexican environments and a low (< or = 1025 kg) US environment. Variance components across Mexican environments were about 40% less than those of the US environment. Genetic correlation coefficients between milk yield in various Mexican environments and all US environments ranged from 0.60 to 0.71 and were different from unity (P < 0.001). Genetic correlation coefficients with low environment in the US ranged between 0.69 and 0.93; the largest correlation was between the low US and high Mexico environments. Both reductions in the size of genetic variance in Mexican environments relative to the US and genetic correlation coefficients less than unity were indicative of genotype by environment interaction. A significant rank change in estimated breeding values (EBV) of sires in Mexican environments relative to the US was another indicator of genotype of EBV of a sire estimated from daughters performances in low and high environments in Mexico were 0.46 and 0.62 against EBV of sires estimated from all data in the US. Against EBV estimated from the low environment in the US they were 0.57 and 0.83. The US low environment was a better predictor of performance in Mexican environments. PMID- 10531611 TI - On-farm quality assurance programs: a survey of producer and industry leader opinions. AB - To assess interest in implementing a California dairy quality assurance program, practices and opinions of dairy producers and industry leaders were surveyed by a mailed questionnaire and by focus groups. The majority of the 55 participants in the focus group were dairy producers; processor marketing executives, extension dairy advisors, packinghouse executives, and dairy veterinarians were represented. The consensus among the focus groups was that a quality assurance program should be voluntary, be managed by creameries, and confer an economic advantage to participants. Focus group members listed chemical and microbial food safety (in both meat and milk), environmental health, and animal welfare as issues that should be addressed. Of the 1440 questionnaires mailed with producers' milk checks, 413 were returned. Information was collected regarding opinions and practices pertaining to administration of drugs to animals, medical records and animal identification, culling practices, manure management, cow welfare, and feeding of animal protein. An overwhelming 99% of producers believed they were responsible for the safety of meat and milk leaving the farm. Sixty percent of producers said that they would consider joining a California-specific quality assurance program, whereas 9% indicated that they would not. Producers would be more likely to join if their processor believed it would impart a market advantage and if the program standards were controlled by producers. PMID- 10531612 TI - Method R estimates of heritability for milk, fat, and protein yields of United States dairy cattle. AB - Heritabilities for milk, fat, and protein yields were estimated from first lactation data used for USDA-Dairy Herd Improvement Association (DHIA) genetic evaluations. Contemporary group assignments and standard deviations within herd year were determined with the procedure used for national evaluations. Pedigree data were included for animals born since 1970; yield data were included for cows born since 1980. Lactation records were divided into four mutually exclusive data sets based on standard deviations. Ranges for standard deviations were chosen so that data sets were approximately equal in size. Method R was used to estimate heritability with 25 different random samples of half of the data for each data set. Because of the large number of Holstein observations, estimates of heritability for Holsteins were based on random subsets of the complete data file; each subset included approximately 5% of the data. Mean heritability estimates increased with standard deviations, and estimates ranged from 0.18 to 0.51 across breeds. Repeatability estimates for milk yield of Holsteins were approximately 0.50 and did not change with standard deviation. These heritability estimates were higher than those previously used in the USDA-DHIA genetic evaluation. Heritability used in the USDA-DHIA genetic evaluation have been increased based on these results. PMID- 10531613 TI - Maximizing the value of milk through separation technologies. AB - Milk is the source of a wide range of proteins that deliver nutrition to the most promising new food products today. Isolated milk proteins are natural, trusted food ingredients with excellent functionality. Separation technologies provide the basis for adding value to milk through the production of proteins that provide the food industry with ingredients to meet specific needs, not possible with milk itself or with other ingredients. The major milk proteins, casein and whey protein, can be isolated by manipulating their compositional and physical properties and then by using various separation technologies to recover the proteins. Additionally, they can be processed in various ways to create a wide range of ingredients with diverse functional characteristics. These ingredients include milk protein concentrate, milk protein isolate, casein, caseinate, whey protein concentrate, whey protein isolate, hydrolysates, and various milk fractions. Within each of these ingredient categories, there is further differentiation according to the functional and nutritional requirements of the finished food. Adding value to milk by expanding from consumer products to ingredients often requires different technologies, marketing structure and distribution channels. The worldwide market for both consumer products and ingredients from milk continues to grow. Technology often precedes market demand. Methods for the commercial production of individual milk components now exist, and in the future as clinical evidence develops, the opportunity for adding value to dairy products as functional foods with health benefits may be achieved. The research and development of today will be the basis of those value-added milk products for tomorrow. PMID- 10531614 TI - Enhancing market value of milk by adding cultures. AB - Fluid milk and several dairy products are an excellent medium to generate an array of products that fit into the current consumer demand for health-driven foods. Several technologies associated with culture addition, fermentation, or both are available for creating an assortment of flavors and textures in milk products. It appears that accentuating the positive attributes of inherent milk constituents, incorporating health-promoting cultures, and offering a variety of flavors and textures to the consumer could enhance milk consumption. Recent advances in probiotic research show much promise in new product development of functional foods based on milk. Several scientifically sound clinical studies have verified some of the anecdotal reports of the past. Among the reported beneficial effects of consuming certain strains of cultures, or their metabolites, or both are enhanced immune response, balancing of colonic microbiota, vaccine adjuvant effect, reduction of fecal enzymes implicated in cancer initiation, treatment of diarrhea associated with travel, antibiotic therapy, control of rotavirus and Clostridium difficile, control of ulcers related to Helicobacter pylori, reduction of serum cholesterol, antagonism against food-borne pathogens and tooth decay organisms, and amelioration of lactose malabsorption symptoms. The mode of action in most cases seems to involve modulation of ecosystem of the gastrointestinal tract of the host. Several strains belonging to genera Enterococcus, Lactobacillus, and Bifidobacterium, which have desirable clinical benefits, are now available. They are being incorporated in yogurts, dairy snacks, breakfast foods, drinks, refrigerated desserts, cheeses, spreads, frozen desserts, and unfermented sweet cultured milk. PMID- 10531615 TI - Unphosphorylated crossbridges and latch: smooth muscle regulation revisited. PMID- 10531616 TI - Opposite changes in myosin heavy chain composition of human masseter and biceps brachii muscles during aging. AB - The myosin heavy chain (MyHC) content in functionally different parts of the human masseter muscle of six elderly and five young adult subjects (mean age 74 and 22 years, respectively) was determined, using gel electrophoresis. The MyHC composition of the old masseter was also studied by enzyme- and immunohistochemical methods and compared with previous data for young adults. For comparison, the biceps brachii muscle of the same subjects was also analysed. The old masseter contained smaller amounts of slow and larger amounts of fast and fetal MyHCs. These differences were region-dependent and were more pronounced in the superficial portion. There was also a larger proportion of "hybrid" fibres, containing two to four MyHC isoforms (42%), compared with the young adult masseter (23%). No such differences were observed between old and young biceps. In contrast to the masseter, the old biceps contained more slow MyHC and less fast MyHC. This investigation demonstrates that the aging process in human skeletal muscle is accompanied by a modification in the muscle phenotype which is both muscle and region specific; a transformation towards a fast and fetal phenotype concomitant with an increased number of fibres with a mixture of different MyHC isoforms in the masseter; and an opposite shift towards a slower phenotype in the biceps brachii. The results might reflect differences between jaw and limb muscles in genetic programs and adaptive responses to changed functional demands following aging. PMID- 10531617 TI - Thin-filament linked regulation of smooth muscle myosin. AB - Phosphorylation of the regulatory light chain subunit of smooth muscle myosin is sufficient, but not necessary for muscle contraction. It has been suggested that thin-filament regulation may also contribute to the regulation of contraction. A hallmark feature of regulated thin filaments, previously described for vertebrate skeletal muscle, is the capacity of strong-binding or rigor-like cross bridges to "turn-on" the actin filament. Turned-on thin filaments stimulate cross-bridge attachment even in the absence of calcium. The present study utilized an in vitro sliding-filament motility assay to test for thin-filament regulation of both unphosphorylated and phosphorylated smooth muscle myosins. Regulated thin filaments were reconstituted from skeletal muscle actin and chicken gizzard smooth muscle tropomyosin (TmCG), and then turned-on either (1) by rigor cross bridges at low concentrations of MgATP, or (2) by adding N-ethyl-maleimide modified skeletal subfragment S1(NEM-S1), which forms rigor-like bonds in the presence of MgATP. For control actin.TmCG filaments, force production by unphosphorylated myosin was 0.5% of that produced by thiophosphorylated myosin. The force exerted on actin.Tm filaments by both unphosphorylated and phosphorylated myosins was increased by reducing the [MgATP] to 10-100 microM MgATP (rigor-dependent activation). Force was also increased by actin.TmCG filaments that had been turned-on by NEM-S1 binding, with force production by unphosphorylated myosin increased 80-fold vs. 2.3-fold for thiophosphorylated myosin. TmCG was required for increased force production with both low MgATP and NEM-S1. Unloaded filament velocity for NEM-S1-activated thin filaments was 0.72 micron/sec with unphosphorylated myosin compared to 1.24 microns/sec with thiophosphorylated myosin. Taken together, these results suggest that thin filament regulation may play a role in the activation of both unphosphorylated and phosphorylated smooth muscle myosins and suggest a possible mechanism for activation of slowly cycling unphosphorylated cross bridges (i.e. latch-state) during tonic contractions of smooth muscle. PMID- 10531618 TI - Z-line structural diversity in frog single muscle fiber in the passive state. AB - The structural changes of the Z-line between small square net (ss) and basket weave (bw) cross-sectional patterns were examined using intact single fibers and mechanically skinned fibers in the passive state to determine if the pattern is related to the sarcomere length (SL) and if the pattern undergoes a reversible transition in low- and high-osmotic medium. Frog single fibers were isolated from the anterior tibial muscle in Ringer's solution. Entirely or partially skinned single fibers were prepared in relaxing solution (also called low-osmotic medium). The high osmotic medium contained 10% polyvinylpyrrolidone (PVP) in relaxing solution. The sarcomere length (SL) of each fiber was measured directly by use of a laser beam or indirectly from electron micrographs with use of a correction factor. The ss and bw forms in cross sections were quantified by analysis of electron micrographs. The results show that the structural change of Z-line occurs around bw << 2.3-2.4 microns << ss (n = 25) and bw << 3.1-3.2 microns << ss (n = 13) in intact single fibers and skinned fibers, respectively. With the quick freeze-freeze substitution method, an intact single fiber with a SL of 2.35 microns showed almost 100% of ss form. The structural transition in cross section was also confirmed in four partially skinned fibers, where patterns went from mostly ss form (intact portion) to mostly bw form (skinned portion) at the SL between 2.40 to 3.20 microns. The reversibility of the change between ss and bw was proved by using low- and high-osmotic medium. The transition and reversion of cross-sectional patterns both occur in the passive state. PMID- 10531619 TI - Differential distribution of dystrophin and beta-spectrin at the sarcolemma of fast twitch skeletal muscle fibers. AB - We used double label immunofluorescence and confocal microscopy to examine the organization of beta-spectrin and dystrophin at the sarcolemma of fast twitch myofibers in the Extensor Digitorum Longus (EDL) of the rat. Both beta-spectrin and dystrophin are concentrated in costameres, a rectilinear sarcolemmal array composed of longitudinal strands and transverse elements overlying Z and M lines. In contrast, intercostameric regions, lying between these linear structures, contain significant levels of dystrophin but little detectable beta-spectrin. The dystrophin-associated proteins, syntrophin and beta-dystroglycan, are also concentrated at costameres but, like dystrophin, are present in intercostameric regions as well. Dystrophin is present at costameres and intercostameric regions in fast twitch muscles of the mouse but is absent from all regions of the sarcolemma in the mdx mouse, which lacks dystrophin. Areas of the sarcolemma near myonuclei also contain dystrophin without beta-spectrin, consistent with the idea that the distribution of dystrophin at the sarcolemma is not dependent on beta spectrin. We conclude that dystrophin is present under all areas of the sarcolemma. The increased fragility of the sarcolemma in patients with Duchennes muscular dystrophy may be explained in part by the absence of dystrophin not only from costameres, but also from intercostameric regions. PMID- 10531620 TI - Prolonged passive stretch of rat soleus muscle provokes an increase in the mRNA levels of the muscle regulatory factors distributed along the entire length of the fibers. AB - The mRNA levels of the adult and the neonatal sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPases (SERCA1a and SERCA1b, respectively) and those of the muscle regulatory factors (MRFs: myoD, myf-5, myogenin, MRF4) have been assessed by RT PCR in rat soleus muscles immobilized for 3 days in an extended position (passive stretch). The transcript level of the fast type SERCA1a Ca(2+)-transport ATPase decreased to half of its normal value, whereas that of neonatal SERCA1b isoform increased 5-fold above control in stretched muscles. Immunostaining of muscle cross sections showed that the fraction of fibers expressing the SERCA1a protein was decreased evenly along the length of the stretched muscles indicating that a transformation occurred of fast fibers to slow ones. The mRNA levels of MRFs were elevated 3- to 6-fold above the normal level and were distributed evenly along the length of the stretched muscles. However in the controls these transcripts were more abundant at both ends of the muscle. The stretch increased the level of myoD and immunocytochemistry showed the expression of myoD protein in a number of nuclei of the stretched muscles whereas it was practically undetectable by this method in the control muscles. Western blotting did not indicate a significant stretch-induced increase in the level of the myogenin protein, in spite of the fact that immunocytochemistry tended to show more myogenin-positive nuclei in stretched muscles as compared to the controls. These data indicate that after 3 days of passive stretch the central and the terminal parts of the soleus muscle adapt similarly by increasing the levels of the MRFs, by decreasing the overall levels of the fast SERCA1-type of ATPase and by partially re-establishing a neonatal mode of alternative SERCA1 transcript splicing resulting in an increased SERCA1b/1a ratio. PMID- 10531621 TI - Interaction of triadin with histidine-rich Ca(2+)-binding protein at the triadic junction in skeletal muscle fibers. AB - The present study documents the binding interaction of skeletal muscle sarcoplasmic reticulum (SR) transmembrane protein triadin with peripheral histidine-rich, Ca(2+)-binding protein (HCP). In addition to providing further evidence that HCP coenriches with RyR1, FKBP-12, triadin and calsequestrin (CS) in sucrose-density-purified TC vesicles, using specific polyclonal antibody, we show it to be expressed as a single protein species, both in fast-twitch and slow twitch fibers, and to identically localize to the I-band. Colocalization of HCP and triadin at junctional triads is supported by the overlapping staining pattern using monoclonal antibodies to triadin. We show a specific binding interaction between digoxigenin-HCP and triadin, using ligand blot techniques. The importance of this finding is strengthened by the similarities in binding affinity and in Ca2+ dependence, (0.1-1 mM Ca2+) of the interaction of digoxigenin-HCP with immobilized TC vesicles. Suggesting that triadin dually interacts with HCP and with CS, at distinct sites, we have found that triadin-CS interaction in overlays does not require the presence of Ca2+. Consistent with the binding of CS to triadin luminal domain (Guo and Campbell, 1995), we show that binding sites for digoxigenin-CS, although not binding sites for digoxigenin-HCP, can be recovered in the 92 kDa triadin fragment, after chymotryptic cleavage of the NH2-terminal end of the folded molecule in intact TC vesicles. These differential effects form the basis for the hypothesis that HCP anchors to the junctional membrane domain of the SR, through binding to triadin short cytoplasmic domain at the NH2 terminus. Although the function of this interaction, as such, is not well understood, it seems of potential biological interest within the more general context of the structural-functional role of triadin at the triadic junction in skeletal muscle. PMID- 10531622 TI - Fibre type-specific expression of p94, a skeletal muscle-specific calpain. AB - Members of the calpain proteinase family are present in all mammalian cells, although a novel calpain 94 kDa isoform is found almost exclusively in skeletal muscle. p94 is difficult to purify from muscle and recombinant p94 autolyses rapidly when expressed in COS cells. However, in vivo the enzyme may be stabilised by interaction with titin, which has two well-characterised binding sites for p94 at the N2- and M-lines. Both these titin subdomains are subject to muscle-specific alternative splicing, which could be related to p94 expression level or stability in muscles of different fibre type. In this study, porcine longissimus dorsi (LD), trapezius (TZ) and adductor longus (AL) were characterised as fast, intermediate and slow using commercially available specific anti-human fast- and slow-myosin heavy chain mAbs and also by conventional histochemistry. p94 was quantified both in whole muscle preparations and single fibres by western blotting using an anti-p94 antiserum generated by expressing a recombinant p94 sequence as a GST fusion protein antigen. SDS PAGE and immunoblotting revealed a single band of approximately 94 kDa with identical mobility in all muscle and fibre preparations. The intensity of the 94 kDa band was greater in LD (22 +/- 1.7 densitometric units mean +/- SEM, n = 3) than TZ and AL (10 +/- 2.3 and 6 +/- 0.9 units, respectively). Expressed as a ratio relative to actin immunoreactivity, p94 is present in all types of single fibres isolated from TZ, but at a significantly lower level (P < 0.01) in slow type I (0.08 +/- 0.01, n = 9), compared to fast IIA/IIB fibres (0.22 +/- 0.02, n = 26). No evidence was seen for rapid or variable rate of p94 degradation in either type of fibre. These data suggest a positive correlation between p94 expression level and fast glycolytic characteristics in porcine muscle. PMID- 10531624 TI - A group motivational treatment for chemical dependency. AB - Patient "motivation" has been implicated as a critical component in addiction treatment outcomes. To date, treatments utilizing motivational elements have been conducted as individual interventions. We describe the development of a Group Motivational Intervention (GMI), a four-session, manual-driven group approach that employs key hypothesized motivational elements. These include the six motivational elements derived by Miller and Sanchez (1994) from successful alcoholism treatments, described with the acronym, FRAMES (feedback, responsibility, advice, menu of options, empathy, and self-efficacy). GMI is additionally informed by concepts derived from "self-determination theory" (Deci & Ryan, 1985), concerned with understanding motivation as either internal/autonomous or external/controlled. Evidence indicates that people will value and persist longer in behaviors that they perceive as autonomously motivated. GMI techniques utilize the interpersonal factors found to be autonomy supportive in self-determination theory. Preliminary results from a randomized clinical trial suggest that key motivational processes are affected by GMI: patients perceive the GMI environment and group leader as significantly more "autonomy supportive" than treatment "as usual." PMID- 10531623 TI - Contractile properties of rabbit psoas muscle fibres inhibited by beryllium fluoride. AB - The structure of truncated, recombinant Dictyostelium myosin motor domain complexed with Mg.ADP and slowly dissociating analogues of Pi has previously been characterized as two main states (S1-MgADP plus BeFx vs. A1F4- or Vi). The BeFx bound state is thought to mimic the weak actin-binding M.ATP complex, while the states with A1F4- or Vi bound mimic the M.ADP.Pi state. While the effects of A1F4 and Vi on fibre mechanics have been previously described (Chase et al., 1994, 1993), the effects of BeFx have not been characterized in detail. At pCa 4.5 (12 degrees C), we measured (i) steady-state isometric tension, (ii) stiffness (KS; 1 kHz sinusoids), and (iii) unloaded shortening velocity (Vu; slack test) in single skinned muscle fibres from rabbit psoas. Results were compared when tension was inhibited with either BeFx or 2,3-butanedione-monoxime (BDM) or modulated by altering myoplasmic [Ca2+]. With 3 mM total fluoride, 1 mM BeFx inhibited both tension and KS by approximately 50% (compared to 7-10 mM BDM and 50-100 microM A1F4-). Increasing [BeFx] to 10 mM further reduced tension to approximately 15% P0, but had little further effect on KS; with BDM and altered [Ca2+], KS scaled more proportionately with tension. Inhibition of tension and KS by BeFx was more rapidly reversible, compared with slow recovery from tension inhibition with A1F4 or Vi. Vu exhibited a complex dependence on [BeFx], being relatively unaffected by concentrations < or = 1 mM, and becoming inhibited steeply for [BeFx] above this level. With BDM, Vu co-varied more directly with force. Our results suggest that BeFx may induce a different cross-bridge state in fibres than do A1F4- or Vi, but all three analogues of Pi form complexes that mimic crossbridge states that follow ATP hydrolysis. PMID- 10531625 TI - A new method for detoxifying opioid-dependent patients. AB - The purpose of the study was to investigate a new method for detoxifying opioid dependent patients. Butorphanol is an opiod with mixed agonist-antagonist properties, and is marketed as a nasal spray. Undiluted, it will cause significant physical withdrawal symptoms in a population dependent on opioids. Forty patients dependent on opioids were detoxified using dilute butorphanol spray. The initial concentration was 40% and gradually reduced. This technique was highly successful in keeping this difficult group of patients engaged in the treatment process for a longer period. This time increased the probability of getting these patients involved with postdetoxification services. There was a subset of patients who had chronic pain and were opioid dependent, who had adequate control of pain with butorphanol spray at 10 to 20% concentration. This unique and unusual approach is worthy of open discussion and further scientific studies. PMID- 10531626 TI - Client engagement in drug treatment. AB - Client engagement in drug abuse treatment is associated with favorable treatment outcomes, but it is not completely understood why some clients are more likely to engage in treatment. This study examines a wide array of client characteristics and treatment experiences potentially associated with engagement. Findings from the Los Angeles Target Cities Project, funded by the Center for Substance Abuse Treatment, indicate that the predictors of treatment engagement are generally confined to current treatment experiences. For both women and men, the perceived utility of treatment, ancillary services, and the client-counselor relationship are the strongest predictors of client engagement in treatment. Client characteristics are generally not strong predictors of treatment engagement. Concerning the client-counselor relationship, the findings suggest that women may respond more favorably to an empathic counseling style, whereas men may respond to a more utilitarian style. The findings contradict popular stereotypes about the treatment-"receptive" client, identify possible directions for treatment improvement, and highlight the need for more research examining the treatment experience of the client. Other research, clinical, and policy implications are discussed. PMID- 10531627 TI - Comparison of substance abuse treatment outcomes for inpatients and outpatients. AB - This study was conducted to determine whether inpatient substance abuse treatment was associated with higher posttreatment abstinence rates than outpatient treatment. The follow-up sample of 2,476 adults represented 183 Minnesota treatment programs. Composite measures were constructed based on psychometric analyses of a modified version of the Addiction Severity Index and additional variables. A series of analyses was conducted, including hierarchical logistic regression and a contingency table analysis addressing multiple problem severity. For the total sample, setting was not significantly associated with abstinence once other outcome predictors and differences between inpatients and outpatients were controlled for. However, when the logistic regression analysis was extended to include all possible two-way interactions of setting with clinically related severity variables, recent suicidal behavior was found to be a moderator of the association between setting and outcome. Inpatient treatment significantly predicted a higher posttreatment abstinence rate than outpatient treatment for the small subset of patients (16% of the sample) who reported recent suicidal ideation or attempt. PMID- 10531628 TI - Methods of changing patterns of substance use among individuals with co-occurring schizophrenia and substance use disorder. AB - Individuals with a severe mental illness and substance use disorder tend to have medical and social problems and to make slower progress in treatment than those who have either disorder alone. Nevertheless, little attention has been paid to the discovery of effective methods of modifying substance use in the severely mentally ill (SMI). The purpose of this study was to collect qualitative data as a way to help identify techniques that might help to change patterns of substance use in the SMI. The participants were 21 men and women who were psychiatric clinic outpatients and who had a current schizophrenia spectrum diagnosis. A total of 18 participants had a lifetime diagnosis of alcohol abuse or dependence, and 21 lifetime other drug diagnoses were recorded for the sample. These individuals participated in focus group discussions about topics related to substance use and people's experiences with trying to quit. The results showed that participants identified several therapeutic and extratherapeutic factors that helped them to initiate and maintain changes in their substance use, as well as factors that hindered change. The findings are related to knowledge about the effectiveness of substance use disorder treatment techniques in general, and implications of the data are discussed for the conduct of integrated treatment of individuals with severe mental illness and a substance use disorder. PMID- 10531629 TI - Use of monetary reinforcers by cocaine-dependent outpatients. AB - Monetary reinforcers have not been widely used as contingent reinforcers in the treatment of drug abuse, despite their demonstrated effectiveness. This is primarily due to concern that drug abusers will use monetary reinforcers to procure drugs. The present study addressed this concern by examining 48 cocaine dependent outpatients' biweekly self-reports of how they used their earned reinforcers. For each subject, their reinforcement usage was classified into 12 higher-order categories and 34 subcategories. Usage proportions were calculated for each. Results indicated that monetary reinforcers were used very infrequently to acquire drugs or alcohol (2%). Reinforcers were used primarily for daily life activities (25%) (e.g., food and gas), money-related uses (18%) (e.g., savings and repaying debts), personal use (15%) (e.g., cosmetics and clothes), and household items (11%) (e.g., rent and bills). These findings challenge the concern that drug abusers use monetary reinforcers to purchase drugs and have important implications for the use of contingent monetary reinforcers in treatment settings. PMID- 10531630 TI - Three oral formulations of methadone. A clinical and pharmacodynamic comparison. AB - This study was done to determine whether there were any differences in subjective symptoms of opiate withdrawal or methadone pharmacodynamics among patients as they were switched between three different oral formulations of methadone. Patients enrolled in a three-way double-blind crossover trial of three methadone formulations. Subjective symptoms and pharmacodynamic measures were assessed throughout the study period. Eighteen patients were enrolled the study. No statistically significant differences in any of the pharmacodynamic parameters studied were found among the three methadone preparations. There was no significant difference among preparations in the rate and extent of rise and fall in plasma methadone levels during a 24-hour intensive sampling period. Subjective symptoms also did not correlate with methadone formulation. Intolerance to changes in methadone formulation, often observed clinically, do not appear to have a pharmacodynamic basis. Our findings support the notion that such change intolerance reflects factors other than the pharmacologic properties of the different formulations of methadone. PMID- 10531631 TI - The substance abuse subtle screening inventory minimizes the need for toxicology screening of prenatal patients. AB - Multiple authors have reported attempts to effectively address the discovery of substance abuse in pregnancy using various mechanisms to encourage positive self reports and urine toxicology to augment identification. In this study, we evaluated 1,251 patients with (a) self-report, (b) the Substance Abuse Subtle Screening Inventory (SASSI), and (c) urine toxicology screening to determine which modality or combination would yield the most cost-effective discovery. Combining the SASSI with the self-report was the most clinically effective and cost effective mode of discovery. This led to the development of a clinical protocol using the SASSI and self-report with limited use of urine toxicology for specific patient subgroups. Alcohol abuse, which is missed by toxicology and self report, is detected by the SASSI. PMID- 10531632 TI - Making residential treatment available to methadone clients. AB - This article addresses challenges to integrating clients on methadone into residential treatment, with the goal of promoting greater access for this population. It describes the basic administrative conditions needed for success, and discusses barriers and problems within the methadone program and the residential program. Staff communication, procedures for coordination, client and staff attitudes and understanding, and ongoing education are seen as the key to creating an environment conducive to success for the client. PMID- 10531633 TI - Clinicians' self-assessment. Questions and answers in substance abuse treatment. PMID- 10531634 TI - Young male violent death trends in the general population during the Vietnam era. AB - Suicide and homicide rates significantly increased throughout the Vietnam War among young American civilian males who constituted the principal manpower pool for the war. Ironically, men who reached military age after the war were at greatest risk. Years of high combat intensity were not associated with higher suicide or homicide rates than years of low combat intensity. Suicide and homicide rates were correlated at .95, suggesting a common source of pathogenesis. No similar trend was found for motor vehicle death. Broad social forces, not the Vietnam War itself, were responsible for the changes in violent mortality trends that we observed among the birth cohorts of men in our study. PMID- 10531635 TI - Young female violent death trends in the general population during the Vietnam era. AB - Five birth cohorts of young women who became age 20 before, during, and after the Vietnam War were selected for this study of motor vehicle accident, suicide, and homicide between ages 13 and 30. Suicide and homicide increased strongly in succeeding years (1953-1986). Motor vehicle accident rates were affected primarily by age. But age, birth cohort membership, and age x cohort interaction significantly affected suicide and homicide rates, with the 19-24-year-old age range most greatly affected. The correlation between suicide and homicide risk (r = .92) was similar to that for young males (r = .95) and suggested a common source of pathogenesis for both genders. PMID- 10531636 TI - Suicide ideation and its relationship to depressed mood in a community sample of adolescents in Hong Kong. AB - A sample of 996 Chinese adolescents in Hong Kong provided information about their suicide ideation. Their depressive symptoms were measured by the Chinese Beck Depression Inventory (C-BDI), and a broad range of stressors were also assessed as potential predictors to level of suicide ideation. The stressors and C-BDI scores predicted 33% of the variance in suicide ideation. In boys, most of the prediction was attributable to C-BDI scores, suggesting that depression mediated the effects of stressors. In girls, C-BDI scores, perceptions of low parental caring, and high conflict with parents had additive effects in predicting level of suicide ideation. This study contributes needed information about a non Western population and highlights gender differences. PMID- 10531637 TI - A model for the description and interpretation of suicide prevention. AB - Views about suicide prevention are based on underlying beliefs about the origins of problems and basic concepts about humankind. These implicit theories have an effect on prevention practices. Making these views explicit is one of the keys for the further development of suicide prevention. In this study a paradigm for analyzing suicide prevention of means of a coding frame and interpreting findings by means of theoretical models of prevention was elaborated. The analysis was based on empirical data consisting of definitions of prevention given by psychologists (N = 34) participating in the national suicide prevention project in Finland. The study demonstrates that suicide prevention can be differentiated at the operational level by means of the analysis method generated. Moreover, the findings can be interpreted according to theoretical criteria. Views expressed by the psychologists seemed to correspond largely to central features of current prevention models. Furthermore, the data can be seen to serve as an empirical validation of these models. Suicide prevention proved to be a multifactorial concept manifesting mainly process theory and interactional explanations of suicidality, and prevention practices fell into a simple typology of four categories. PMID- 10531638 TI - Survivors of suicide do grieve differently: empirical support for a common sense proposition. AB - Previous empirical investigations have produced mixed results on the question of whether mode of death differentially affects grief. To further investigate the influence of suicide on grief, 350 previously bereaved university students completed a questionnaire package consisting of several standardized measures. Participants were separated into four groups based on the mode of death experienced as either survivors of suicide (n = 34), accident (n = 57), unanticipated natural (n = 102), or anticipated natural (n = 157) deaths. Hierarchical multiple regression analyses indicated that suicide survivors, compared against the other groups, experienced more frequent feelings of rejection, responsibility, "unique" reactions, and more total grief reactions. Trends indicating increased levels of shame and perceived stigmatization were also evident. Aggregate factors of death "naturalness" and "expectedness" showed less influence than mode of death in influencing grief. Overall, results support previous clinical and research findings and intuitive logic in demonstrating that the grief experienced by suicide survivors includes elements that are less frequently seen in the case of nonsuicidal deaths. PMID- 10531639 TI - Postponed suicide death? Suicides around birthdays and major public holidays. AB - The relationship between suicide and birthdays, and suicide and public holidays has been studied from data on 32,291 Danish suicides by persons ages 15 years and older in the 25-year period 1970-1994. Evidence was found to support the theory of the "broken-promise effect" for major public holidays in that there appears to be a postponement of a significant number of suicides from before a holiday until after. The division of holidays into nonworking and (half-time) working days showed that a "holiday effect" could only be found around major public holidays, particularly Christmas, Easter, and Whitsun. The postponing or transpositioning effect is relevant to prevention, especially because of the availability and accessibility of help at the end of and after major public (nonworking) holidays. PMID- 10531640 TI - Microbial production of 1,3-propanediol. AB - 1,3-Propanediol (1,3-PD) production by fermentation of glycerol was described in 1881 but little attention was paid to this microbial route for over a century. Glycerol conversion to 1,3-PD can be carried out by Clostridia as well as Enterobacteriaceae. The main intermediate of the oxidative pathway is pyruvate, the further utilization of which produces CO2, H2, acetate, butyrate, ethanol, butanol and 2,3-butanediol. In addition, lactate and succinate are generated. The yield of 1,3-PD per glycerol is determined by the availability of NADH2, which is mainly affected by the product distribution (of the oxidative pathway) and depends first of all on the microorganism used but also on the process conditions (type of fermentation, substrate excess, various inhibitions). In the past decade, research to produce 1,3-PD microbially was considerably expanded as the diol can be used for various polycondensates. In particular, polyesters with useful properties can be manufactured. A prerequisite for making a "green" polyester is a most cost-effective production of 1,3-PD, which, in practical terms, can only be achieved by using an alternative substrate, such as glucose instead of glycerol. Therefore, great efforts are now being made to combine the pathway from glucose to glycerol successfully with the bacterial route from glycerol to 1,3-PD. Thus, 1,3-PD may become the first bulk chemical produced by a genetically engineered microorganism. PMID- 10531642 TI - Yeast cells as tools for target-oriented screening. AB - Information about biomolecular interaction networks is crucial for understanding cellular functions and the development of disease processes. Many diseases are known to be based on aberrations of DNA sequences encoding proteins with key functions in the cellular metabolism. Alterations in the respective proteins often lead to disturbances in biomolecular interactions caused by unbalanced stoichiometries, and thus result in alterations of molecule fluxes, cell architecture and signalling pathways. Drug discovery programmes have been designed to find promising chemical lead structures with the help of target oriented bioassay systems. These are, in most cases, based upon the interaction of small molecules to specific macromolecular targets in vivo or in vitro, as exemplified by enzyme assays or small-ligand-based receptor systems. In addition, interactions between large biomolecules, such as proteins or nucleic acids, offer a huge arsenal of potential drug targets that can be addressed by small chemical compounds. This latter approach is gaining considerable attention because many potential target structures are becoming available through genomic research. Funnelling these new targets into high-throughput screening programs represents a major challenge for today's pharmaceutical research. An important outcome of the ongoing genome projects is the fact that the basic cellular structures, pathways and signalling principles show a high degree of conservation. Model organisms that are easily approachable by genetic, biochemical and physiological means can thus play an important role in the design of target-oriented screening systems. They offer the possibility to express individual proteins, nucleic acids or even more complex aggregates of biomolecules such as protein-interaction networks or transcription-initiation complexes, which can be addressed by small effector molecules in vivo. Combining these targets with biological signalling systems is an attractive way of creating robust cellular assay systems. PMID- 10531641 TI - Biosynthesis of gibberellins in Gibberella fujikuroi: biomolecular aspects. AB - Gibberellins (GAs) are a large family of isoprenoid plant hormones hormones, some of which are bioactive growth regulators, controlling seed germination, stem elongation, and flowering. The rice pathogen Gibberella fujikuroi (mating population C) is able to produce large amounts of GAs, especially the bioactive compounds gibberellic acid (GA3) and its precursors, GA4 and GA7. The main steps of the biosynthetic pathway have long been established from the identification of intermediates in wild-type G. fujikuroi and mutant strains. However, the genetics of the fungus have been rather under-developed, and molecular genetic studies of the GA pathway started just recently. The progress in researching GA biosynthesis in the last 2 years resulted primarily from development of the molecular tools, e.g. transformation systems for the fungus, and cloning the genes encoding GA biosynthesis enzymes, such as the bifunctional ent-copalyl diphosphate/kaurene synthase and several cytochrome P450 monooxygenases. The availability of these genes opened new horizons both for detailed study of the pathway and the regulation mechanisms at the molecular level, and for modern strain improvement programs. This review gives a short overview of the well-known physiological and biochemical studies and concentrates mainly on the new molecular genetic data from GA research, including new information on the regulation of GA biosynthesis. PMID- 10531643 TI - Production of 2,3-butanediol by newly isolated Enterobacter cloacae. AB - Enterobacter cloacae NRRL B-23289 was isolated from local decaying wood/corn soil samples while screening for microorganisms for conversion of L-arabinose to fuel ethanol. The major product of fermentation by the bacterium was meso-2,3 butanediol (2,3-BD). In a typical fermentation, a BD yield of 0.4 g/g arabinose was obtained with a corresponding productivity of 0.63 g/l per hour at an initial arabinose concentration of 50 g/l. The effects of initial arabinose concentration, temperature, pH, agitation, various monosaccharides, and multiple sugar mixtures on 2,3-BD production were investigated. BD productivity, yield, and byproduct formation were influenced significantly within these parameters. The bacterium utilized sugars from acid plus enzyme saccharified corn fiber and produced BD (0.35 g/g available sugars). It also produced BD from dilute acid pretreated corn fiber by simultaneous saccharification and fermentation (0.34 g/g theoretical sugars). PMID- 10531644 TI - Production of (R)-3-pentyn-2-ol through stereoinversion of racemic 3-pentyn-2-ol by Nocardia fusca AKU 2123. AB - Wet cells of Nocardia fusca AKU 2123 are good catalysts for the production of (R) 3-pentyn-2-ol (PYOH) from (RS)-PYOH through a stereoinversion reaction. Under optimal conditions (350 mM potassium phosphate buffer, pH 8.0, 30% (w/v) wet cells, 0.12% NADPH, 10% glucose, and 30 U/ml glucose dehydrogenase) (R)-PYOH of high optical purity (98.7% e.e.) was produced from 2% (v/v) (RS)-PYOH with a yield of 70.4% by 140 h incubation. PMID- 10531645 TI - Succinoglycan production by solid-state fermentation with Agrobacterium tumefaciens. AB - Succinoglycan was produced by cultivating Agrobacterium tumefaciens on various solid substrates, including agar medium, spent malt grains, ivory nut shavings, and grated carrots, impregnated with a nutrient+ solution. Fermentations were performed on a laboratory scale, both under static conditions and with agitation, using bottles and a prototype horizontal bioreactor. Several fermentation parameters were examined and optimized, including carbon and nitrogen composition, water content and layer thickness of the substrate. The yields and rheological properties of the polymers obtained under different fermentation conditions were compared. The highest succinoglycan yield was achieved in static cultivation, reaching 42 g/l of impregnating solution, corresponding to 30 g/kg of wet substrate. The polymer production in the horizontal bioreactor was faster, but the final yield was lower (29 g/l of impregnating solution). PMID- 10531646 TI - Improved process for production of recombinant yeast-derived monomeric human G CSF. AB - The human granulocyte colony-stimulating factor (hG-CSF) was efficiently secreted at high levels in fed-batch cultures of recombinant Saccharomyces cerevisiae. However, the secreted recombinant hG-CSF (rhG-CSF) was shown to exist as large multimers in the culture broth due to strong hydrophobic interaction. It was hardly monomerized even by urea at high concentration. This multimer has been reported to diminish specific receptor-binding activity of hG-CSF and causes undesirable problems in the downstream process. When the rhG-CSF was secreted to extracellular broth in the presence of a non-ionic surfactant (Tween 80) in the culture media, the multimerization of the secreted rhG-CSF was efficiently prevented in the fed-batch cultures. Also, the monomer fraction and secreted efficiency of rhG-CSF were significantly increased at the higher culture pH (6.5). Without using any denaturing agents, the secreted rhG-CSF monomer was easily purified with high recovery yield and purity via a simple purification process under acidic conditions, consisting of diafiltration, cation exchange, and gel filtration chromatography. A lyophilization process devoid of intermonomer aggregation was also designed using effective stabilizing agents. PMID- 10531648 TI - Benzene/toluene/p-xylene degradation. Part I. Solvent selection and toluene degradation in a two-phase partitioning bioreactor. AB - A two-phase organic/aqueous reactor configuration was developed for use in the biodegradation of benzene, toluene and p-xylene, and tested with toluene. An immiscible organic phase was systematically selected on the basis of predicted and experimentally determined properties, such as high boiling points, low solubilities in the aqueous phase, good phase stability, biocompatibility, and good predicted partition coefficients for benzene, toluene and p-xylene. An industrial grade of oleyl alcohol was ultimately selected for use in the two phase partitioning bioreactor. In order to examine the behavior of the system, a single-component fermentation of toluene was conducted with Pseudomonas sp. ATCC 55595. A 0.5-1 sample of Adol 85 NF was loaded with 10.4 g toluene, which partitioned into the cell containing 1 l aqueous medium at a concentration of approximately 50 mg/l. In consuming the toluene to completion, the organisms were able to achieve a volumetric degradation rate of 0.115 g l-1 h-1. This system is self-regulating with respect to toluene delivery to the aqueous phase, and requires only feedback control of temperature and pH. PMID- 10531647 TI - An integrated strategy for the process development of a recombinant antibody cytokine fusion protein expressed in BHK cells. AB - Recombinant fusion proteins offer important new therapeutic approaches for the future. This report describes the use of three different genetic strategies (i.e. "mono-", "bi-" and "tri-cistronic" vectors) to achieve stable secretion from BHK cells of a glycosylated antibody-cytokine fusion protein designed for use in antitumour therapy. It describes selection of a robust and effective production cell line based on stability of secretion of the product, quality of mRNA and protein products and performance in in vitro bioassays for potency. The data obtained at this stage were utilised in the selection of a suitable candidate production cell line. The relative productivity and general performance of the cells in stirred tank and fixed bed culture systems indicated that a variety of cell culture technologies provided robust tools for production of a highly selected cell clone. Consistency of the product glycosylation was determined by analysis of released oligosaccharides using matrix-assisted laser desorption ionisation-time of flight mass spectrometry and high-performance anion exchange chromatography. These investigations showed consistent expression of three glycoforms of the fusion protein which varied in their relative proportions in different culture systems and at different time points in a fixed bed reactor with continuous perfusion. In conclusion, this study dealt with a range of important scientific and technical issues which are essential for regulatory approval and commercial success of a recombinant protein and elucidates some useful markers for process development for similar recombinant biologicals. PMID- 10531649 TI - Benzene/toluene/p-xylene degradation. Part II. Effect of substrate interactions and feeding strategies in toluene/benzene and toluene/p-xylene fermentations in a partitioning bioreactor. AB - A two-phase aqueous/organic partitioning bioreactor scheme was used to degrade mixtures of toluene and benzene, and toluene and p-xylene, using simultaneous and sequential feeding strategies. The aqueous phase of the partitioning bioreactor contained Pseudomonas sp. ATCC 55595, an organism able to degrade benzene, toluene and p-xylene simultaneously. An industrial grade of oleyl alcohol served as the organic phase. In each experiment, the organic phase of the bioreactor was loaded with 10.15 g toluene, and either 2.0 g benzene or 2.1 g p-xylene. The resulting aqueous phase concentrations were 50 mg/l, 25 mg/l and 8 mg/l toluene, benzene and p-xylene respectively. The simultaneous fermentation of benzene and toluene consumed these compounds at volumetric rates of 0.024 g l-1 h-1 and 0.067 g l-1 h-1, respectively. The simultaneous fermentation of toluene and p-xylene consumed these xenobiotics at volumetric rates of 0.066 g l-1 h-1 and 0.018 g l-1 h-1, respectively. A sequential feeding strategy was employed in which toluene was added initially, but the benzene or p-xylene aliquot was added only after the cells had consumed half of the initial toluene concentration. This strategy was shown to improve overall degradation rates, and to reduce the stress on the microorganisms. In the sequential fermentation of benzene and toluene, the volumetric degradation rates were 0.056 g l-1 h-1 and 0.079 g l-1 h-1, respectively. In the toluene/p-xylene sequential fermentation, the initial toluene load was consumed before the p-xylene aliquot was consumed. After 12 h in which no p-xylene degradation was observed, a 4.0-g toluene aliquot was added, and p-xylene degradation resumed. Excluding that 12-h period, the microbes consumed toluene and p-xylene at volumetric rates of 0.074 g l-1 h-1 and 0.025 g l-1 h-1, respectively. Oxygen limitation occurred in all fermentations during the rapid growth phase. PMID- 10531650 TI - Immunoaffinity layering of enzymes. Stabilization and use in flow injection analysis of glucose and hydrogen peroxide. AB - A general procedure for the high yield immobilization of enzymes with the help of specific anti-enzyme antibodies is described. Polyclonal antibodies were raised against Aspergillus niger glucose oxidase and horseradish peroxidase in rabbits and the gamma globulin (IgG) fraction from the immune sera isolated by ammonium sulphate fractionation followed by ion-exchange chromatography. Immobilization of glucose oxidase and horseradish peroxidase was achieved by initially binding the enzymes to a Sepharose matrix coupled with IgG isolated from anti-(glucose oxidase) and anti-(horseradish peroxidase) sera, respectively. This was followed by alternate incubation with the IgG and the enzyme to assemble layers of enzyme and antibody on the support. The immunoaffinity-layered preparations obtained thus were highly active and, after six binding cycles, the amount of enzyme immobilized could be raised about 25 times over that bound initially. It was also possible to assemble layers of glucose oxidase using unfractionated antiserum in place of the IgG. The bioaffinity-layered preparations of glucose oxidase and horseradish peroxidase exhibited good enzyme activities and improved resistance to heat-induced inactivation. The sensitivity of a flow injection analysis system for measuring glucose and hydrogen peroxide could be remarkably improved using immunoaffinity-layered glucose oxidase and horseradish peroxidase. For the detection of glucose, a Clark-type oxygen electrode, constructed as a small flow through cell integrated with a cartridge bearing immunoaffinity-layered glucose oxidase was employed. The hydrogen peroxide concentration was analysed spectrophotometrically using a flow-through cell and the layered horseradish peroxidase packed into a cartridge. The immunoaffinity-layered enzymes could be conveniently solubilized at acid pH and fresh enzyme loaded onto the support. Immunoaffinity-layered glucose oxidase was successfully used for the on-line monitoring of the glucose concentration during the cultivation of Streptomyces cerevisiae. PMID- 10531651 TI - Cloning, sequence analysis, and expression in Escherichia coli of the gene encoding phenylacetaldehyde reductase from styrene-assimilating Corynebacterium sp. strain ST-10. AB - The gene encoding phenylacetaldehyde reductase (PAR), a useful biocatalyst for producing chiral alcohols, was cloned from the genomic DNA of the styrene assimilating Corynebacterium sp. strain ST-10. The gene contained an opening reading frame consisting of 1,158 nucleotides corresponding to 385 amino acid residues. The subunit molecular weight was calculated to be 40,299, which was in agreement with that determined by polyacrylamide gel electrophoresis. The enzyme was sufficiently expressed in recombinant Escherichia coli cells for practical use and purified to homogeneity by three-column chromatography steps. The predicted amino acid sequence displayed only 20-29% identity with zinc containing, NAD(+)-dependent, long-chain alcohol dehydrogenases. Nevertheless, the probable NAD(+)- and zinc-binding sites are conserved although one of the three catalytic zinc-binding residues of the zinc-containing, long-chain alcohol dehydrogenases was substituted by Asp in PAR. The protein contains 7.6 mol zinc/mol tetramer. Therefore, the enzyme was considered as a new member of zinc containing, long-chain alcohol dehydrogenases with a particular and broad substrate specificity. PMID- 10531652 TI - Characterization of a gene encoding Trametes versicolor laccase A and improved heterologous expression in Saccharomyces cerevisiae by decreased cultivation temperature. AB - Laccase can be used for enzymatic detoxification of lignocellulosic hydrolysates. A Saccharomyces cerevisiae strain with enhanced resistance to phenolic inhibitors and thereby improved ability to ferment lignocellulosic hydrolysates would presumably be obtained by heterologous expression of laccase. Sequencing of the cDNA for the novel laccase gene lcc2 from the lignin-degrading basidiomycete Trametes versicolor showed that it encodes an isoenzyme of 499 amino-acid residues preceded by a 21-residue signal peptide. By comparison with Edman degradation data, it was concluded that lcc2 encodes an isoenzyme corresponding to laccase A. The gene product of lcc2 displays 71% identity with the previously characterized T. versicolor lcc1 gene product. An alignment of laccase sequences revealed that the T. versicolor isoenzymes in general are more closely related to corresponding isoenzymes from other white-rot fungi than to the other T. versicolor isoenzymes. The multiplicity of laccase is thus a conserved feature of T. versicolor and related species of white-rot fungi. When the T. versicolor lcc2 cDNA was expressed in S. cerevisiae, the production of active enzyme was strongly dependent on the temperature. After 3 days of incubation, a 16-fold higher laccase activity was found when a positive transformant was kept at 19 degrees C instead of 28 degrees C. Similar experiments with Pichia pastoris expressing the T. versicolor laccase gene lcc1 also showed that the expression level was favoured considerably by lower cultivation temperature, indicating that the observation made for the S. cerevisiae expression system is of general significance. PMID- 10531653 TI - Genetic transformation of a Rhizomucor pusillus mutant defective in asparagine linked glycosylation: production of a milk-clotting enzyme in a less-glycosylated form. AB - Rhizomucor pusillus 1116R3 has a defect in alg2 encoding a mannosyltransferase in the asparagine (N)-linked oligosaccharide biosynthetic pathway and produces proteins in less-glycosylated forms. For development of a genetic transformation system for this zygomycete, an uracil auxotroph (mutant 1116U17) as the host strain was derived by ultraviolet (UV) mutagenesis as 5-fluoroorotic acid resistant colonies and the orotidine-5'-monophosphate (OMP) decarboxylase (pyr4) gene as a selection marker was cloned from the wild-type strain R. pusillus F27 by the polymerase chain reaction with primers designed on the basis of the pyr4 sequences from other fungi. The amino acid sequence of R. pusillus Pyr4 deduced from the nucleotide sequence showed high homology with the OMP decarboxylases from various fungi. The pyr4 gene on pUC19 (plasmid pRPPyr4) was introduced into protoplasts of R. pusillus 1116U17 by polyethylene glycol-assisted transformation. Transformation under optimized conditions yielded 5 Ura+ transformants with 1 microgram pRPPyr4 DNA and 1 x 10(7) viable protoplasts. Southern blot analysis of the genomic DNA from the transformants showed that multiple copies of the pRPPyr4 sequence were integrated into the genome by homologous recombination at the pyr4 locus. For the purpose of production of a milk-clotting aspartic proteinase (MPP) in a less-glycosylated form, mpp from the wild-type strain was cloned in pRPPyr4 and introduced into protoplasts of R. pusillus 1116U17. Transformants obtained in this way contained multiple copies of mpp at the chromosomal mpp locus and produced MPP as a mixture of molecules having no sugar chains and Man0-1GlcNAc2 at the two N-linked glycosylation sites in an amount about 12 times larger than the parent strain. The transformation system for R. pusillus 1116U17 would be useful for production of proteins with truncated N-linked oligosaccharide chains. PMID- 10531654 TI - Expression and secretion of a biologically active mouse sonic hedgehog protein by the methylotrophic yeast Pichia pastoris. AB - We have successfully secreted the amino-terminal functional domain of mouse sonic hedgehog protein (SHH) into culture fluid using a yeast Pichia pastoris expression system. A cDNA fragment encoding the amino-terminal domain of mouse SHH was inserted downstream of the Saccharomyces cerevisiae alpha-mating factor secretion signal. The DNA fragment was introduced into the host genome by the spheroplast transformation method. Transformants were selected based on their resistance to G418: His+ transformants which showed resistance to over 8 mg G418/ml were selected and analyzed for determination of the plasmid copy number. One His+ clone which has eight copies of the expression cassette per genome was cultured in minimal medium deficient for histidine, and further cultured in buffered medium supplemented with methanol which activates the AOX1 promoter. SDS PAGE analysis indicated efficient expression and secretion of mouse SHH into culture fluid. The yield of secreted SHH was estimated to be 50 micrograms/ml. Purified protein was assayed for biological activity and found to activate the transcription of the Patched genes (Ptc-1 and Ptc-2) encoding receptors for SHH. PMID- 10531655 TI - Amylose-like polysaccharide accumulation and hyphal cell-surface structure in relation to citric acid production by Aspergillus niger in shake culture. AB - When 120 mg glucose/ml was used as a carbon source, in shake culture Aspergillus niger Yang no. 2 maximally produced only 15.4 mg citric acid/ml but accumulated 3.0 mg extracellular polysaccharide/ml. The polysaccharide secreted by mycelia of Yang no. 2 in shake culture was confirmed to be an amylose-like alpha-1,4-glucan by hydrolysis analysis with acid, amylase and glucoamylase. However, in static cultures, such as semisolid and surface cultures free from physical stresses caused by shaking damage, Yang no. 2 produced more citric acid but did not accumulate the polysaccharide. With cultivation time in shake culture, the amount of extracellular polysaccharide and the viscosity of the culture broth increased. The increase of shaking speed caused a remarkable increase in the accumulation of extracellular polysaccharide, e.g. 11.2 mg extracellular polysaccharide/ml was accumulated in the medium at a shaking speed of 200 rpm. The addition of 2.0 mg carboxymethylcellulose (CMC)/ml as a viscous additive to the medium reduced drastically the amount of extracellular polysaccharide accumulated to 1.5 mg/ml, but increased the citric acid produced to 52.0 mg/ml. However, intracellular polysaccharide accumulation kept up a steady rate of 0.26 microgram/mg dried mycelium through the entire period of cultivation. The addition of 3.0 mg polysaccharide/ml purified from the culture broth to the medium at the start of a culture resulted in a decrease of extracellular polysaccharide accumulation but an increase of citric acid accumulation. From electronmicroscopic observation, cell surfaces of hyphae cultivated with CMC were smooth, while hyphae cultivated without CMC had fibrous and granular polysaccharide on the cell surface. These results suggested that Yang no. 2 secreted the polysaccharide on the cell surface as a viscous substance and/or a shock absorber to protect itself from physical stresses caused by shaking damage in shake culture. PMID- 10531656 TI - Autolysis of Escherichia coli and Bacillus subtilis cells in low gravity. AB - The role of gravity in the autolysis of Bacillus subtilis and Escherichia coli was studied by growing cells on Earth and in microgravity on Space Station Mir. Autolysis analysis was completed by examining the death phase or exponential decay of cells for approximately 4 months following the stationary phase. Consistent with published findings, the stationary-phase cell population was 170% and 90% higher in flight B. subtilis and E. coli cultures, respectively, than in ground cultures. Although both flight autolysis curves began at higher cell densities than control curves, the rate of autolysis in flight cultures was identical to that of their respective ground control rates. PMID- 10531657 TI - Bioavailability of water-polluting sulfonoaromatic compounds. AB - Highly substituted arenesulfonates are chemically stable compounds with a range of industrial applications, and they are widely regarded as being poorly degradable. We did enrichment cultures for bacteria able to utilise the sulfonate moiety of 14 compounds, and we obtained mixed cultures that were able to desulfonate each compound. The products formed were usually identified as the corresponding phenol, but because we could not obtain pure cultures, we followed up these findings with quantitative work in pure cultures of, e.g., Pseudomonas putida S-313, which generated the same phenols from the compounds studied. Many of these phenols are known to be biodegradable, or to be subject to binding to soil components. We thus presume that the capacity to degrade aromatic sulfonates extensively is widespread in the environment, even though the degradative capacity is spread over several organisms and conditions. PMID- 10531658 TI - Superordinate category formation in pigeons: association with a common delay or probability of food reinforcement makes perceptually dissimilar stimuli functionally equivalent. AB - Training associated pairs of perceptually dissimilar stimulus classes with a common delay or probability of food reinforcement in pigeons. Then, different choice responses were trained to 1 component class in each pair. In a choice test, the untrained class in each pair occasioned the same response as did the choice-trained class. In a 3rd experiment, 2 classes had reinforcement delays of 1 s and 15 s, respectively, and 2 other classes had reinforcement probabilities of 0.1 and 0.9. Then, 1 choice response was reinforced to a class previously associated with a better condition of reinforcement (e.g., 1-s delay or 1.0 probability), and a different response was reinforced to a class previously associated with a worse condition of reinforcement (0.1 probability or 0-s delay). Testing with all classes suggested that categorization was based on the relative reinforcement or hedonic value and not on the parametric details of reinforcement. PMID- 10531659 TI - The pigeon's variability discrimination with lists of successively presented visual stimuli. AB - Pigeons previously trained to peck 1 button (same) after the successive presentation of 16 identical pictures and to peck a 2nd button (different) after the successive presentation of 16 nonidentical pictures were tested on lists involving different degrees of variability, different list lengths, and different temporal organizations of list items. The pigeons' performances on this successive same-different task revealed a strong sensitivity to list entropy; but, their discrimination was also affected by their memory for list items and by the accumulated evidence for a same versus a different response. Statistical models confirmed and quantified the importance of these additional factors. PMID- 10531661 TI - Decision processes in discrimination: fundamental misrepresentations of signal detection theory. AB - A new approach to studying decision making in discrimination tasks is described that does not depend on the technical assumptions of signal detection theory (e.g., normality of the encoding distributions). In 3 different experiments, results of these new distribution-free tests converge on a single, surprising conclusion: response biases had substantial effects on the encoding distributions but no effect on the decision rule, which was uniformly unbiased in equal and unequal base rate conditions and in symmetric and asymmetric payoff conditions. This seemingly paradoxical result is fundamentally inconsistent with the entire family of signal detection theory models, raising some important questions about the significance of many published results in the human performance literature. PMID- 10531660 TI - Drug-onset cues as signals: intraadministration associations and tolerance. AB - On the basis of a conditioning analysis of drug tolerance, drug-associated cues become associated with the drug effect. These cues elicit conditional compensatory responses and modulate the expression of tolerance. Although there are many findings consistent with the conditioning analysis of tolerance, there also are contrary findings. The results of these experiments suggest that some of the apparently contradictory findings result because interoceptive pharmacological cues, as well as exteroceptive environmental cues, are paired with a drug effect. That is, within each administration, early drug-onset cues may become associated with the later, larger drug effect, and these pharmacological cues may overshadow simultaneously present environmental cues. We demonstrate the contribution of such intraadministration associations to tolerance to the analgesic effect of morphine and to the expression of conditional compensatory hyperalgesia. PMID- 10531662 TI - Intentional switching between behavioral patterns of homologous and nonhomologous effector combinations. AB - Intentional switching between preferred coordination modes (Experiment 1) and between isofrequency and multifrequency conditions (Experiments 2 and 3) was compared across different effector combinations. Experiment 1 showed that homologous limbs switched faster toward the in-phase and anti-phase mode than nonhomologous limbs, supporting their distinct degree of coordinative stability during 1:1 synchronization. Experiments 2 and 3 revealed that switching time between isofrequency and multifrequency conditions depended on the attractiveness of both coordination dynamics associated with the combination of segments involved. These results are consistent with the unique prediction derived from dynamic pattern approach in which the differential stability of the coordination modes determines the switching time. PMID- 10531663 TI - Can practice eliminate the psychological refractory period effect? AB - Can people learn to perform two tasks at the same time without interference? To answer this question, the authors trained 6 participants for 36 sessions in a Psychological Refractory Period (PRP) experiment, where Task 1 required a speeded vocal response to an auditory stimulus and Task 2 required a speeded manual response to a visual stimulus. The large PRP effect found initially (353 ms in Session 1) shrank to only about 40 ms over the course of practice, disappearing entirely for 1 of the 6 participants. This reduction in the PRP effect with practice is considerably larger than has been previously reported. The obtained pattern of factor interactions between stimulus onset asynchrony and each of three task difficulty manipulations (Task 1 judgment difficulty, Task 2 stimulus contrast, and Task 2 mapping compatibility) supports a postponement (bottleneck) account of dual-task interference, both before and after practice. PMID- 10531664 TI - Postural coordination modes considered as emergent phenomena. AB - The coordination of multiple body segments (torso and legs) in the control of standing posture during a suprapostural task was studied. The analysis was motivated by dynamical theories of motor coordination. In 2 experiments it was found that multisegment postural coordination could be described by the relative phase of rotations around the hip and ankle joints. The effective length of the feet, the height of the center of mass, and the amplitude of head motions in a visual tracking task were varied. Across these variations, 2 modes of hip-ankle coordination were observed: in-phase and anti-phase. The emergence of these modes was influenced by constraints imposed by the suprapostural tracking task, supporting the idea that such tasks influence postural control in an adaptive manner. Results are interpreted in terms of a dynamical approach to coordination in which postural coordination modes can be viewed as emergent phenomena. PMID- 10531665 TI - Correlations for timing consistency among tapping and drawing tasks: evidence against a single timing process for motor control. AB - Three experiments were conducted to examine whether timing processes can be shared by continuous tapping and drawing tasks. In all 3 experiments, temporal precision in tapping was not related to temporal precision in continuous drawing. There were modest correlations among the tapping tasks, and there were significant correlations among the drawing tasks. In Experiment 3, the function relating timing variance to the square of the observed movement duration for tapping was different from that for drawing. The conclusions drawn were that timing is not an ability to be shared by a variety of tasks but instead that the temporal qualities of skilled movement are the result of the specific processes necessary to produce a trajectory. These results are consistent with the idea that timing is an emergent property of movement. PMID- 10531666 TI - Parallel and competitive processes in hierarchical analysis: perceptual grouping and encoding of closure. AB - The role of perceptual grouping and the encoding of closure of local elements in the processing of hierarchical patterns was studied. Experiments 1 and 2 showed a global advantage over the local level for 2 tasks involving the discrimination of orientation and closure, but there was a local advantage for the closure discrimination task relative to the orientation discrimination task. Experiment 3 showed a local precedence effect for the closure discrimination task when local element grouping was weakened by embedding the stimuli from Experiment 1 in a background made up of cross patterns. Experiments 4A and 4B found that dissimilarity of closure between the local elements of hierarchical stimuli and the background figures could facilitate the grouping of closed local elements and enhanced the perception of global structure. Experiment 5 showed that the advantage for detecting the closure of local elements in hierarchical analysis also held under divided- and selective-attention conditions. Results are consistent with the idea that grouping between local elements takes place in parallel and competes with the computation of closure of local elements in determining the selection between global and local levels of hierarchical patterns for response. PMID- 10531667 TI - The maintenance of apparent luminance of an object. AB - Results from luminance discriminations with objects defined by apparent motion suggest an object-specific temporal integration of luminance. Further experiments suggested that this integration is weighted to favor the initial display of an object and involves the percept of surface reflectance (lightness). These results are consistent with the object-file metaphor suggested by D. Kahneman, A. Treisman, and B. Gibbs (1992), in which an object's perceived initial surface reflectance is assigned and maintained in an object file. A strategy is proposed in which the intrinsic properties of an object are assumed not to change over time. As intrinsic properties are generally invariant and possibly difficult to compute, this strategy would have the advantage of relatively high accuracy at relatively low computational cost. PMID- 10531668 TI - When fair is foul and foul is fair: reverse priming in automatic evaluation. AB - Responses to information were facilitated by the rapid prior presentation of evaluatively congruent material. This fundamental discovery (R. H. Fazio, D. M. Sanbonmatsu, M. C. Powell, & F. R. Kardes, 1986) marked a breakthrough in research on automatic information processing by demonstrating that evaluative meaning is grasped without conscious control. Experiments employing a word naming task provided stringent tests of the automaticity of evaluation and found support for it. More strikingly, a previously unobserved reversal of these effects (i.e., slower responses to evaluatively matched rather than mismatched items) was found when primes were evaluatively extreme. Procedural variances across 6 experiments revealed that the reverse priming effect was highly robust. This discovery is analogous to demonstrations of contrast effects in controlled judgments. It is theorized that the reverse priming effect reflects an automatic correction for the biasing influence of the prime. PMID- 10531669 TI - Self-observation of social behavior and metaperception. AB - In 3 experiments (Ns = 68, 72, and 101) the authors tested the hypothesis that the opportunity to observe oneself in social interaction increases the accuracy of metaperception (prediction of others' social judgments of oneself). Small groups were videotaped during a decision-making task, after which group members judged each other's social anxiety. Participants watched either the videotape of their group's interaction or a videotape of another group's interaction. After watching the videotape, participants predicted how they were judged by each member of the group. Results from the 3 experiments confirmed the hypothesis that self-observation increases the accuracy of metaperception. Presumably, self observation provides objective information about one's behavior, which increases the ability to determine how one is judged by others, assuming self and others share meaning systems. PMID- 10531670 TI - Abortion as stigma: cognitive and emotional implications of concealment. AB - This study examined the stigma of abortion and psychological implications of concealment among 442 women followed for 2 years from the day of their abortion. As predicted, women who felt stigmatized by abortion were more likely to feel a need to keep it a secret from family and friends. Secrecy was related positively to suppressing thoughts of the abortion and negatively to disclosing abortion related emotions to others. Greater thought suppression was associated with experiencing more intrusive thoughts of the abortion. Both suppression and intrusive thoughts, in turn, were positively related to increases in psychological distress over time. Emotional disclosure moderated the association between intrusive thoughts and distress. Disclosure was associated with decreases in distress among women experiencing intrusive thoughts of their abortion, but was unrelated to distress among women not experiencing intrusive thoughts. PMID- 10531671 TI - Less pain and more to gain: why high-status group members blame their failure on discrimination. AB - Members of high-status groups are more likely than members of low-status groups to blame their failure on discrimination and are less likely to blame it on themselves. This tendency was demonstrated in 3 experiments comparing men and women, White and Black students, and members of experimentally created high- and low-status groups. Results also showed that when making an attribution to discrimination, high-status group members were less likely to experience a threat to their social state self-esteem, performance perceived control, and social perceived control and were more likely to protect their performance state self esteem. These findings help to explain why high-status group members are more willing to blame their failure on discrimination by showing that it is less harmful for them than for low-status group members. PMID- 10531672 TI - "Thanks for sharing that": ruminators and their social support networks. AB - Receiving positive social support after a trauma generally is related to better adjustment to the trauma. The personality of trauma survivors may affect the extent to which they seek social support, their perceived receipt of social support, and the extent to which they benefit from social support. The authors hypothesized that people with a ruminative coping style, who tended to focus excessively on their own emotional reactions to a trauma, compared to those without a ruminative coping style, would seek more social support, and would benefit more from social support, but would report receiving less social support. These hypotheses were confirmed in a longitudinal study of people who lost a loved one to a terminal illness. PMID- 10531673 TI - Resilient, overcontrolled, and undercontrolled personality prototypes in childhood: replicability, predictive power, and the trait-type issue. AB - In a longitudinal study, Q-sort patterns of German preschool children were analyzed for personality prototypes and related to developmental outcomes up to age 12. Q-factor analyses confirmed 3 prototypic patterns that showed a high continuity and cross-judge consistency; were similar to those found for North American, Dutch, and Icelandic children; and can be interpreted as resilient, overcontrolled, and undercontrolled. Relations reported by R. W. Robins, O. P. John, A. Caspi, T. E. Moffitt, & M. Stouthamer-Loeber (1996) between these 3 patterns and the Big Five were fully replicated. Growth curve analyses showed that the 3 patterns predicted important developmental outcomes in both the social and the cognitive domains. Evidence was found for both traits and types: A continuous dimension of resiliency bifurcates in its lower part into two relatively discrete personality types, overcontrollers and undercontrollers. PMID- 10531674 TI - Uncontrollability, depression, and the construction of mental models. AB - Three studies examined mental model generation after preexposure to uncontrollability and in a depressive state. The purpose of the experiments was to test the implications of the cognitive exhaustion model, applying an explicit conceptualization of social mental models and a process-tracing method developed by U. von Hecker (1997). An experimental situation was created for observation of consecutive, rule-based construction steps as a function of input diagnosticity, and for the quality assessment of constructed mental models. The findings show that participants preexposed to uncontrollability, as well as depressed students, were able, as were controls, to identify rule-relevant information needed for model construction. However, they were less able than control participants to engage in a more cognitively demanding and generative step of processing (i.e., in integrating the pieces of input information into a coherent mental model of sentiment relations). PMID- 10531675 TI - A longitudinal study of negative affect and self-perceived competence in young adolescents. AB - In a 4-wave, 2-year longitudinal design, the authors obtained measures of negative affect (NA) and self-perceived competence from 220 boys and 216 girls who were 7th graders at the beginning of this study. NA was operationalized as the common dimension underlying self-reports of depressive symptoms, anxiety symptoms, and negative emotions. Self-perceived competence consisted of 2 higher order constructs: a well-behaved/good-student factor and an attractive/athletic/popular factor. Structural equation modeling revealed very high stability estimates for all constructs. Nevertheless, self-perceived competence in the attractive/athletic/popular domain predicted changes in NA. Conversely, NA predicted changes in self-perceived competence in the well behaved/good-student domain. The primacy of NA versus self-cognitions depends, in part, on the type of self-cognitions being examined. PMID- 10531676 TI - Differential association of traits of fear and anxiety with norepinephrine- and dark-induced pupil reactivity. AB - The relation between fear and anxiety remains unclear, though psychometric data strongly suggest they are independent emotional systems. Because central norepinephrine (NE) projection systems are at the core of models of both fear and anxiety, the present experiment explored whether this independence extends to NE functioning. Two different aspects of NE functioning were assessed in a healthy young adult sample (N = 18): pupillary reactivity to (a) a specific NE alpha-1 agonist challenge to assess receptor reactivity and (b) a darkness challenge to assess contributions of central NE. Pupillary reactivity to the former was strongly and specifically related to A. Tellegen's (1982) Multidimensional Personality Questionnaire (MPQ) Harm Avoidance scale (i.e., trait fear), whereas the latter was strongly and specifically related to MPQ Negative Emotionality (i.e., trait anxiety). Implications for conceptualizing fear and anxiety as emotional systems are discussed. PMID- 10531677 TI - Relationship between thigmotropism and Candida biofilm formation in vitro. AB - The biofilm formation of the oral fungal pathogen Candida on denture acrylic strips coated with saliva or serum was examined in relation to the ability to induce hyphae by thigmotropic reaction, using C. albicans (4 isolates), C. glabrata (3 isolates) and C. tropicalis (3 isolates). Both the degree of biofilm formation and the amount of hyphae exhibiting thigmotropism varied depending upon both the species and strains of Candida. Although there was no significant correlation between the amount of hyphae induced by thigmotropic reaction of fungal isolates and biofilm formation on uncoated control specimens (r = 0.577; p < 0.05), the ability of hyphae induced by thigmotropic reaction significantly correlated with the amount of both saliva- and serum-admixed biofilms (r = 0.734; p < 0.05 and r = 0.793; p < 0.01, respectively). Taken together our in vitro data suggested that the hyphal induction by thigmotropic reaction is of importance in candidal biofilm formation on saliva- or serum-coated acrylic surfaces. PMID- 10531678 TI - Role of the armadillo Dasypus novemcinctus in the epidemiology of paracoccidioidomycosis. AB - A study conducted in a rural area of Ibia, State of Minas Gerais, Brazil, where Paracoccidioides brasiliensis was recently isolated from soil, sought to determine if the armadillo Dasypus novemcinctus developed paracoccidioidomycosis. Out of 21 armadillos captured in the area, one had a lung granuloma containing fungal cells attributable to those of P. brasiliensis. The present report presents the first histopathological evidence for the presence of this pathogen in the lungs of an armadillo. This finding permitted us to establish a linkage between the two species and to suggest that fungal infections of humans and of armadillos occur in the same agricultural and forest microenvironment that they share. PMID- 10531679 TI - Candida biotypes isolated from clinical specimens in Malaysia. AB - The distribution of Candida species was examined using 1114 yeasts isolated from various clinical specimens. The isolates were identified by germ tube test, hyphal/pseudohyphae and chlamydoconidia production and carbohydrate assimilation test using ten carbohydrates (glucose, sucrose, trehalose, cellobiose, arabinose, galactose, mannitol, raffinose, lactose and maltose). Among the 1114 isolates studied, 9 species of Candida were identified and the relative frequency of isolation was C. albicans (44.2%), C. parapsilosis (26.0%), C. tropicalis (17.7%), C. glabrata (9.6%), C. krusei (1.2%), C. rugosa (0.6%), C. guilliermondii (0.2%), C. lusitaniae (0.08%) and C. kefyr (0.08%). Non-C. albicans was the most common Candida species isolated from blood, respiratory system, urine and skin. The isolate from vaginal swabs was predominantly C. albicans. 82.2% of C. glabrata and 64.2% of C. krusei isolated in this study were from vaginal swabs. PMID- 10531680 TI - A murine model of zygomycosis by Cunninghamella bertholletiae. AB - Infections by Cunninghamella bertholletiae have been on the increase in recent years. However, little is known about this fungus and its infection. To clarify the pathogenicity of C. bertholletiae, we made a murine model, and to our knowledge, the first infectious model of this fungus. ICR mice pretreated with cortisone acetate and cyclophosphamide were inoculated intratracheally with 5 x 10(5) spores of C. bertholletiae. About half of the mice died by day 4 and 90% died by day 9. C. bertholletiae was cultured from the lungs, and the pathological analysis disclosed diffuse hyphal growth in the lungs, resulting in necrosis in the later stage. Angioinvasion with alveolar hemorrhage was extremely pronounced from the early stage, and this was the most characteristic feature of this infection. Treatment with amphotericin B showed only minimal improvement of survival, comparable to the poor result of this treatment in actual human cases. In fact, our model has many similarities to the actual human infection by C. bertholletiae, and will be useful for further investigations of this infection. PMID- 10531682 TI - Dimorphic fungus characteristic of fumonisin-producing strains of Fusarium moniliforme from Zhejiang. AB - Fusarium moniliforme and its fumonisins have been shown to be carcinogenic in lab animals and have been linked to high incidences of human esophageal cancer. In this study we report the dimorphic fungus characteristic of fumonisin-producing strains of F. moniliforme from foodstuffs in Zhejiang, China. All of the twenty strains of F. moniliforme shown produce fumonisin B1 475.9-6322.2 micrograms/g in corn medium. These strains of F. moniliforme form yeast-like colonies in Sabouraud's agar plates contained 9% NaCl at 37 degrees C incubator and shows mostly budding reproduction. In blood agar plates these strains of F. moniliforme appear grass-green haemolytic reactions. This is the first report that yeast-like growth, dimorphic pathogenic fungus feature is found in F. moniliforme. These results suggest that it is also important to program epidemiological surveys of F. moniliforme as a primary pathogenic fungus, while proceeding to produce mycotoxins of F. moniliforme in food hygiene. PMID- 10531681 TI - Expression of the complement-binding protein (MP60) of Candida albicans in experimental vaginitis. AB - The expression of the Candida albicans complement-binding C3d protein (MP60) was investigated both in vitro and in vivo by immunogold labelling and electron microscopy. In vivo expression was determined in a rat vaginitis model. Reactivity of in vitro-grown cells to an anti-MP60 rabbit serum was associated with both cytoplasmic and cell wall sites. Immunostaining in the cell wall of both yeast and hyphae was most concentrated in the inner, electron-lucid layer. Immunogold stained preparations of C. albicans from vaginal smears of infected animals also showed intense localization of the MP60 in the inner cell wall, plasma membrane. However, immunogold label was also intense at the cell surface in these samples, mostly in the area of close adherence with the keratinocytes of the vaginal epithelia. These observations indicate that MP60 is expressed both in vitro and in vivo, but to a different degree in the different cell wall layers. PMID- 10531683 TI - Properties of Aspergillus tamarii, A. caelatus and related species from acidic tea field soils in Japan. AB - Fungi in Aspergillus section Flavi include both aflatoxin producers and non producers. Aspergillus caelatus is a recently described non-aflatoxigenic species in this section, which has some common characteristics with A. tamarii, such as yellowish brown color and double walled spores. In contrast to the morphological similarities, all of the A. caelatus isolates tested produced no cyclopiazonic acid whereas most isolates of A. tamarii produce this compound. There are six nucleotide differences that distinguish the DNA sequences of these two species in the regions of ITS1, ITS2, 5.8S rDNA and 28S rDNA and this is a consistent difference. Both species were isolated from acidified field soils, but A. tamarii isolates were more common than A. caelatus in highly acidic soils. PMID- 10531684 TI - [Brazilian Consensus about cardiopathy and pregnancy: guidelines of the Brazilian Cardiology Society to pregnancy and family planning of the women with heart disease]. PMID- 10531685 TI - Pregnancy does not cause structural bioprosthesis alteration. PMID- 10531686 TI - Cardiogenic shock. PMID- 10531687 TI - Systematic overviews and meta-analyses in cardiology. Evidence-based cardiology. VIII. PMID- 10531688 TI - Effects of protein-calorie restriction on mechanical function of hypertrophied cardiac muscle. AB - OBJECTIVE: To assess the effect of food restriction (FR) on hypertrophied cardiac muscle in spontaneously hypertensive rats (SHR). METHODS: Isolated papillary muscle preparations of the left ventricle (LV) of 60-day-old SHR and of normotensive Wistar-Kyoto (WKY) rats were studied. The rats were fed either an unrestricted diet or FR diet (50% of the intake of the control diet) for 30 days. The mechanical function of the muscles was evaluated through monitoring isometric and isotonic contractions. RESULTS: FR caused: 1) reduction in the body weight and LV weight of SHR and WKY rats; 2) increase in the time to peak shortening and the time to peak developed tension (DT) in the hypertrophied myocardium of the SHR; 3) diverging changes in the mechanical function of the normal cardiac muscles of WKY rats with reduction in maximum velocity of isotonic shortening and of the time for DT to decrease 50% of its maximum value, and increase of the resting tension and of the rate of tension decline. CONCLUSION: Short-term FR causes prolongation of the contraction time of hypertrophied muscles and paradoxal changes in mechanical performance of normal cardiac fibers, with worsening of the shortening indices and of the resting tension, and improvement of the isometric relaxation. PMID- 10531689 TI - Transmyocardial laser revascularization. Early clinical experience. AB - OBJECTIVE: To analyze the initial clinical experience of transmyocardial laser revascularization (TMLR) in patients with severe diffuse coronary artery disease. METHODS: Between February, 1998 and February, 1999, 20 patients were submitted to TMLR at the Heart Institute (InCor), University of Sao Paulo Medical School, Brazil, isolated or in association with conventional coronary artery bypass graft (CABG). All patients had severe diffuse coronary artery disease, with angina functional class III/IV (Canadian Cardiovascular Society score) unresponsive to medical therapy. Fourteen patients were submitted to TMLR as the sole therapy, whereas 6 underwent concomitant CABG. Fifty per cent of the patients had either been previously submitted to a CABG or to a percutaneous transluminal coronary angioplasty (PTCA). Mean age was 60 years, ranging from 45 to 74 years. RESULTS: All patients had three-vessel disease, with normal or mildly impaired left ventricular global function. Follow-up ranged from 1 to 13 months (mean 6.6 months), with no postoperative short or long term mortality. There was significant symptom improvement after the procedure, with 85% of the patients free of angina, and the remaining 15% of the patients showing improvement in functional class, as well as in exercise tolerance. CONCLUSION: This novel technique can be considered a low risk alternative for a highly selected group of patients not suitable for conventional revascularization procedures. PMID- 10531690 TI - Characteristics and identification of sites of chagasic ventricular tachycardia by endocardial mapping. AB - OBJECTIVE: To study electrophysiological characteristics that enable the identification and ablation of sites of chagasic tachycardia. METHODS: Thirty-one patients with chronic Chagas' heart disease and sustained ventricular tachycardia (SVT) underwent electrophysiological study to map and ablate that arrhythmia. Fifteen patients had hemodynamically stable SVT reproducible by programmed ventricular stimulation, 9 men and 6 women with ages ranging from 37 to 67 years and ejection fraction varying from 0.17 to 0.64. Endocardial mapping was performed during SVT in all patients. Radiofrequency (RF) current was applied to sites of presystolic activity of at least 30 ms. Entrainment was used to identify reentrant circuits. In both successful and unsuccessful sites of RF current application, electrogram and entrainment were analyzed. RESULTS: Entrainment was obtained during all mapped SVT. In 70.5% of the sites we observed concealed entrainment and ventricular tachycardia termination in the first 15 seconds of RF current application. In the unsuccessful sites, significantly earlier electrical activity was seen than in the successful ones. Concealed entrainment was significantly associated with ventricular tachycardia termination. Bystander areas were not observed. CONCLUSION: The reentrant mechanism was responsible for the genesis of all tachycardias. In 70.5% of the studied sites, the endocardial participation of the slow conducting zone of reentrant circuits was shown. Concealed entrainment was the main electrophysiological parameter associated with successful RF current application. There was no electrophysiological evidence of bystander regions in the mapped circuits of SVT. PMID- 10531691 TI - Aortic stenosis. Gender influence on left ventricular geometry and function in patients under 70 years of age. AB - OBJECTIVE: To verify if adaptive left ventricle (LV) characteristics are also present in individuals under 70 years of age with severe aortic stenosis (AS). METHODS: The study comprised 40 consecutive patients under 70 years of age with AS and no associated coronary artery disease, referred for valve surgery. Out of the 40 patients, 22 were men and 18 women, and the mean age was 49.8 +/- 14.3 years. Cardiac symptoms, presence of systemic hypertension (SH), functional class according to the New York Heart Association (NYHA), and valve lesion etiology were considered. LV cavity dimensions, ejection fraction (EF), fractional shortening (FS), mass (MS), and relative diastolic thickness (RDT) were examined by Doppler echocardiography. RESULTS: Fourteen (63.6%) men and 11 (61.6%) women were classified as NYHA class III/IV (p = 0.70). There was no difference in the frequency of angina, syncope or dyspnea between genders. The incidence of SH was greater in women than in men (10 versus 2, p = 0.0044). Women had a smaller LV end-diastolic diameter index (32.1 +/- 6.5 x 36.5 +/- 5.3 mm/m2, p = 0.027), LV end-systolic diameter index (19.9 +/- 5.9 x 26.5 +/- 6.4 mm/m2, p = 0.0022) and LV mass index (MS) (211.4 +/- 71.1 x 270.9 +/- 74.9 g/m2, p = 0.017) when compared with men. EF (66.2 +/- 13.4 x 52.0 +/- 14.6%, p = 0.0032), FS (37.6 +/- 10.7 x 27.9 +/- 9.6%, p = 0.0046) and RDT (0.58 +/- 0.22 x 0.44 +/- 0.09, p = 0.0095) were significantly greater in women than in men. CONCLUSION: It is the patient gender rather than age that influences left ventricular adaptive response to AS. PMID- 10531692 TI - Mitral restenosis in the early postoperative period of a patient with systemic lupus erythematosus. AB - A forty-eight year old woman, who had undergone mitral comissurotomy and subsequently developed early restenosis, presented with major comissural fusion and verrucous lesions on the cuspid edges of the mitral valve, with normal subvalvar apparatus. Patient did well for the first six months after surgery when she began to present dyspnea on light exertion. A clinical diagnosis of restenosis was made, which was confirmed by an echocardiogram and cardiac catheterization. She underwent surgery, and a stenotic mitral valve with verrucous lesions suggesting Libman-Sacks' endocarditis was found. Because the diagnosis of systemic lupus erythematosus (SLE) had not been confirmed at that time, a bovine pericardium bioprosthesis (FISICS-INCOR) was implanted. The patient did well in the late follow-up and is now in NYHA Class I. PMID- 10531693 TI - Paraplegia following intraaortic balloon circulatory assistance. AB - Intraaortic balloon counterpulsation is frequently used in patients experiencing severe ventricular dysfunction following maximal drug therapy. However, even with the improvement of percutaneous insertion techniques, the procedure has always been followed by vascular; infectious, and neurological complications. This article describes a case of paraplegia due to intraaortic balloon counterpulsation in the postoperative period of cardiac surgery. PMID- 10531694 TI - Clinicopathologic session. Case 2/99: a 40-year-old woman with a pericardial effusion. Instituto do Coracao do Hospital das Clinicas--FMUSP. PMID- 10531695 TI - Cellular and biomolecular mechanisms in dilated cardiomyopathy. PMID- 10531696 TI - Familial hypertrophic cardiomyopathy. Genetic characterization. PMID- 10531698 TI - Outcomes improvement: the true mark of quality in managed care. AB - The definition of the term "quality" continues to be centered on health care providers and not the health of the community. The shift to managed care financing provides a unique opportunity to raise the importance of health outcomes as the true mark of quality in managed care. A widespread fear that managed care organizations are too ready to reduce quality for increased profits has lead to a current national backlash against managed care. Instead of only viewing health plan members as recipients of medical services, health plans should also view members as a population group with subpopulations within them, needing both medical and nonmedical services to improve their health. We introduce the Outcomes Improvement System, an outcomes-driven method for managed care plans and other health systems that incorporates both medical care and population-based health services in a managed care setting. PMID- 10531697 TI - The cost and efficiency of hospital care provided by primary care physicians and medical subspecialists. AB - There is a perceived excess of subspecialists compared with primary care doctors, but there are few severity-adjusted data that characterize the care provided by these physician groups. In a nationwide hospital network, we studied outcomes of 17,185 patients who were hospitalized for 1 of 9 common internal medicine illnesses. For 4 of 9 conditions, the subspecialists treated more severely ill (P < .001) patients. The raw total charges for their care were higher (P < .002) for 4 of 9 conditions and longer stays were required for 2 conditions. After adjusting for severity of illness, differences between the physician groups became minimal. In nine-severity adjusted medical illnesses, subspecialists and primary care physicians provide care that produces similar results for length of stay, charge, and mortality. Health care manpower projections should be re evaluated in light of this information. PMID- 10531699 TI - A quality management project in Israeli navy primary care clinics. AB - The objective of this project was to establish a measurable process of continuous quality improvement of health care services in the Israeli naval primary care clinics. All navy clinics were surveyed at 6-month intervals. The quality of medical recording was evaluated, and instructive workshops were given on the matter. Real-time physician-patient interactions were evaluated, and immediate feedback was given to the examining physician. Complementary medical services were evaluated and steps toward improvement were taken. A total of 1043 medical records were examined. A general improvement in medical-record documentation (from a score of 6.0 +/- 2.5 to a score of 7.4 +/- 1.9, P < .001) was demonstrated during the first 3 years of the project. No significant change was noticed in the physician-patient interaction score. Complementary medical services improved from a score of 4.9 +/- 1.5 in 1994 to a score of 7.4 +/- 0.9 3 years later (P < .02). This project achieved a significant improvement in the quality of medical recording and of complementary medical services. PMID- 10531700 TI - Using artificial neural networks and the Gutenberg-Richter power law to "rightsize" a behavioral health care system. AB - The authors propose a new paradigm for designing and managing behavioral health care systems by using artificial neural networks to measure quality of care (Q), using length-of-stay (LOS) prediction and the variation in LOS prediction, and subsequently using the variation of Q to obtain a measure of uncertainty in treatment. The paper proposes that mental illness is fractal in nature (self similar at all scales) and conforms to power laws like the Gutenberg-Richter (G R) law, whereby there is a log-log relationship between frequency of episodes (i.e., admissions) and the severity of those episodes. The paper also hypothesizes that 28% is the average uncertainty (residual or excess entropy) in the treatment of mental illness. The authors use the G-R paradigm to calculate the severity of admission and, subsequently, the minimum number of beds for different behavioral health care facilities and propose the optimal partition of beds between community and state services, thereby "balancing" the delivery system. The data presented support the notion that mental illness manifests complexity and "self-organized criticality." The authors hypothesize that correcting deviations from the theoretical G-R curve for each level of care will allow optimum resource distribution, improve quality of care, and reduce costs. PMID- 10531701 TI - Attention-deficit/hyperactivity disorder in children and adolescents: rethinking diagnosis and treatment implications for complicated cases. AB - Children and adolescents who currently present for treatment frequently carry a principal diagnosis of attention-deficit/hyperactivity disorder in addition to other diagnoses. Many of these cases have long histories, are chronic, and are significantly complicated. Recognition of complicated cases requires rigorous diagnostic differentiation. Effective treatment and sustainable responses require individualized protocols that include integrated psychosocial and psychopharmacological interventions to minimize adverse reactions and enhance quality of life. PMID- 10531702 TI - Testing pregnant women for HIV: a survey of obstetricians and review of patient prenatal/obstetric medical records--Connecticut 1996-1997. AB - High rates of prenatal testing are needed to identify all HIV-infected pregnant women and prevent transmission to their offspring. To evaluate HIV testing of pregnant women in Connecticut, a survey was conducted in 1998 of licensed obstetricians and a review was performed of 992 randomly sampled prenatal and obstetric medical records for births occurring in 1996. Results of the survey indicated that 78.8% of respondents routinely offered HIV counseling and 76.4% routinely offered HIV testing to pregnant patients in 1997. However, only 44% reported that greater than half of their patients were being tested. Providers who had an HIV testing policy that included HIV-testing unless the patient refused had the highest rate (80.8%; P < 0.05). The review of medical records revealed that only 28.8% of women who delivered in 1996 had an HIV test performed during prenatal care. Maternal characteristics associated (P < 0.05) with testing included Hispanic ethnicity (51.8% tested), younger age (53.6%, < 20 years old), having Medicaid (48.0%), hospital clinic services (44.6%), and history of intravenous drug use (IDU) (76.9%). Testing rate varied by hospital from 0% to 82.8%. These results indicate that considerable modification of provider and hospital practices and policies is needed to raise HIV screening rates. PMID- 10531703 TI - Pancreatic tuberculosis. AB - Tuberculosis of the pancreas is a rare gastrointestinal tract entity. This unusual manifestation of tuberculosis should be included in the differential diagnosis of high-risk patients presenting with a pancreatic mass. PMID- 10531704 TI - The double-crush phenomenon--an unusual presentation and literature review. AB - The double-crush syndrome was initially described by Upton and McComas in 1973. They postulated that nonsymptomatic impairment of axoplasmic flow at more than one site along a nerve might summate to cause a symptomatic neuropathy. This was suggested by their clinical observation that the majority of their patients had a median or ulnar neuropathy associated with evidence of cervicothoracic root lesions. They also hypothesized that one of the constraints on axoplasmic flow could be a metabolic neuropathy, and this is supported by the high association of diabetes and carpal tunnel syndrome. Other researchers have since reported series of patients supporting the frequent association of a proximal and distal nerve compression syndrome, including carpal tunnel syndrome associated with cervical radiculopathy, brachial plexus compression, and diabetic neuropathy. Subsequently, MacKinnon and Dellon have expanded the description of this syndrome to include a) multiple anatomic regions along a peripheral nerve, b) multiple anatomic structures across a peripheral nerve within an anatomic region, c) superimposed on a neuropathy, and d) combinations of the above. We present an unusual case of symptomatic nerve compression caused by two nonanatomic structures within an anatomic region. PMID- 10531705 TI - Hypopharyngeal perforation from a swallowed fork: a brief report and comment. AB - Hypopharyngeal perforations are usually seen as a complication of endotracheal intubation by less experienced physicians in emergency situations. The site most commonly perforated is the pharynx, posterior to the cricopharyngeal muscle; the second most common site is the pyriform sinus. We report here an unusual cause of hypopharyngeal perforation from a swallowed plastic fork in a psychiatric patient. PMID- 10531707 TI - Breast cancer--real progress. PMID- 10531706 TI - Listening outside the box: the case of Lyme disease and other tick-borne diseases. AB - To sum up, get out of the box and look at Lyme disease from the point of view of those who believe they suffer from chronic Lyme disease. Meanwhile, spread the word about how to prevent the disease and pass out information about the facts of the disease. PMID- 10531708 TI - Documentation or hyper-documentation? PMID- 10531709 TI - Women's health in perspective. A real lady killer. PMID- 10531710 TI - A review of male violence against women in Hawaii. AB - This review attempts to emphasize the urgency in addressing issues of violence against women in Hawaii. It demonstrates that violence against women is a significant, challenging, and often overwhelming and overlooked public health problem. While attention to this problem has dramatically increased, more needs to be done to end violence against women and improve the well-being of women and our society as a whole. PMID- 10531711 TI - Local and gay: addressing the health needs of Asian and Pacific Islander American (A/PIA) lesbians and gay men in Hawaii. AB - Asian and Pacific Islander American lesbians and gay men, who are "local" born and raised in Hawaii face conflicting personal and social expectations due to factors including prejudicial attitudes about homosexuality, A/PIA racial/ethnic traditions, and the unique cultural milieu of Hawaii. Based on anecdotal and research reports of this Hawaii population, health and social needs are discussed with implications for professional health practice. PMID- 10531712 TI - Arsenic: an endothelial cell toxin in the liver? PMID- 10531714 TI - Percutaneous sonography-guided fine needle aspiration biopsy of colonoscopic biopsy-negative colonic lesions. AB - BACKGROUND: Pre-operative tissue diagnosis with colonoscopy is not always possible in patients with colonic lesions. OBJECTIVE: To study the usefulness and efficacy of percutaneous ultrasound-guided aspiration biopsy of colonic lesions. METHODS: Fifty consecutive patients with colonic lesions in whom colonoscopic brush cytology and biopsy were either negative or the lesion was not accessible on colonoscopy on two attempts, underwent percutaneous ultrasound-guided fine needle aspiration biopsy. The results were compared with surgical findings. RESULTS: Fine-needle aspiration biopsy revealed adenocarcinoma in 40 patients; one had lymphoma, 2 had tuberculosis, 2 had abscess and 5 patients had negative aspiration. Forty-eight patients (excluding 2 with tuberculosis) underwent laparotomy and the diagnoses on aspiration biopsy were confirmed. Of the 5 negative aspirations, 3 had adenocarcinoma, one had tuberculosis and one intussusception. Thus, we had sensitivity of 91.8%, specificity of 100%, predictive value of positive results 100%, predictive value of negative results 20%, and percentage of false negative results 8.1%. Two patients developed complications--hemorrhage into the peritoneum and sepsis due to perforation at the site of aspiration; both survived after surgery. CONCLUSION: Percutaneous ultrasound-guided aspiration biopsy may be attempted for diagnosis of colonic lesions in situations where it may be the only means of obtaining a cytological diagnosis before surgery. PMID- 10531715 TI - Transabdominal gastroesophageal devascularization without esophageal transection for emergency treatment of variceal hemorrhage. AB - BACKGROUND: The Suguira procedure is an effective non-shunting operation to treat life-threatening hemorrhage from esophageal or gastric varices. The goal of esophageal transection is interruption of submucosal varices, but this leads to high morbidity and mortality rates from esophageal fistulization. AIM: To evaluate a variant of this procedure in which the esophagus is not transected, but the varices are underrun from outside the lumen. METHODS: During the last four and a half years, we performed this modified gastroesophageal devascularization with or without splenectomy in 18 patients as emergency treatment of bleeding esophageal and gastric varices. The data were analyzed retrospectively. RESULTS: Bleeding was controlled in all patients. Three patients with Child's class C disease undergoing emergency surgery died during the early postoperative period. Rebleeding rate was 17% (3 patients). The overall survival was 72.2% (13 of 18). No patient had encephalopathy over a mean follow up of 30 months. CONCLUSION: Gastroesophageal devascularization with variceal under running without esophageal transection is an effective and safe alternative to shunt surgery in the emergency situation. PMID- 10531716 TI - Hepatic manifestations in chronic arsenic toxicity. AB - OBJECTIVE: The hepatotoxic action of arsenic, when used as a therapeutic agent, has long been recognized. Data on liver involvement following chronic exposure to arsenic-contaminated water are scanty. We report the nature and degree of liver involvement on the basis of hospital-based and cohort follow-up studies in patients who consumed arsenic-contaminated drinking water for 1 to 15 years. METHODS: 248 patients with evidence of chronic arsenic toxicity underwent clinical and laboratory examinations including liver function tests and HBsAg status. Liver biopsy was done in 69 cases; in 29 patients, liver arsenic content was estimated by neutron activation analysis. A cohort follow up of 23 patients who took arsenic-free water for 2-12 years was also carried out. RESULTS: Hepatomegaly was present in 190 of 248 patients (76.6%). Noncirrhotic portal fibrosis (91.3%) was the predominant lesion in liver histology. The maximum arsenic content in liver was 6 mg/Kg (mean 1.46 [0.42], control value 0.16 [0.04]; p < 0.001); it was undetected in 6 of 29 samples studied. Cohort follow up studies showed elevation of globulin in four cases and development of esophageal varices in one case. CONCLUSION: We report the largest number of patients with liver disease due to chronic arsenicosis from drinking arsenic contaminated water. Noncirrhotic portal fibrosis is the predominant lesion in this population. PMID- 10531717 TI - Evidence for oxidant stress in chronic pancreatitis. AB - BACKGROUND: Oxidant stress leading to lipid peroxidation is reported to be the common link in the pathogenesis of chronic pancreatitis irrespective of etiology. AIM: To look for evidence of lipid peroxidation in duodenal juice in patients with chronic pancreatitis. METHODS: 19 patients with chronic pancreatitis (14 tropical, 5 alcoholic) and 19 age- and sex-matched subjects with abdominal pain without any cause were studied. Contents were aspirated from the second part of the duodenum during gastroduodenoscopy. Malonyl dialdehyde (MDA) levels were measured in duodenal juice by the thiobarbituric acid method. RESULTS: MDA levels were higher in patients than in the control group (mean [SD] 42.6 [17.0] vs 29.2 [11.7] nmol/mL; p < 0.05). On linear and multiple regression analysis, none of the disease factors correlated with duodenal juice MDA levels. CONCLUSIONS: Lipid peroxidation products are increased in patients with chronic tropical and alcoholic pancreatitis. PMID- 10531718 TI - Primary pulmonary hypertension in cirrhosis of liver. AB - BACKGROUND: Primary pulmonary hypertension (PPH) is a grave association of portal hypertension, and is potentially fatal in liver transplant candidates. AIM: To investigate the prevalence of PPH among cirrhotics with portal hypertension. METHODS: 43 cirrhotics with portal hypertension (Child B 22, C 14), after screening for cardiopulmonary diseases, were evaluated by hemodynamic study. RESULTS: PPH was detected in 2 cases (4.7%), both in Child B, hepatitis B and C viruses being the etiologies. Neither had portal axis thrombosis. Two other cases also had pulmonary hypertension, but with high pulmonary capillary wedge pressure (PCWP). The 41 cases without and 2 cases with PPH had, respectively, mean pulmonary artery pressure (MPAP) 16.3 (5.9) mmHg, 26 mmHg and 33 mmHg; PCWP 11.5 (6.7) mmHg, 12 mmHg and 11 mmHg; transpulmonary pressure gradient 4.8 (2.6) mmHg (n = 27), 14 mmHg and 22 mmHg; and pulmonary vascular resistance 80.2 (55.8) dyne.sec.cm-5 (n = 27), 155.6 dyne.sec.cm-5 and 366.7 dyne.sec.cm-5. No correlation of MPAP was found with either Child-Pugh scoring (r2 = 0.0347) or with hepatic venous pressure gradient (r2 = 0.0021). CONCLUSION: PPH has a prevalence of 4.7% among cirrhotics with portal hypertension; it bears no relation with severity of liver disease. PMID- 10531719 TI - Molecular mechanisms of pathogenesis in amebiasis. AB - Though both Entamoeba histolytica and E. dispar colonize the human gut, only the former is capable of invading tissues and causing disease. Although the biology of the parasite and the mechanism of pathogenesis have been intensively studied, there is a lack of consensus about the molecules of E. histolytica that actively participate in pathogenesis. This article reviews some key molecules involved. Ga1NAc-inhibitable adhesin is a membrane-associated glycoprotein nature, consisting of heavy and light subunits; each of these is encoded by multiple genes. The heavy subunit is useful in differentiating E. histolytica from E. dispar. Three structurally similar isoforms of amebapore, A, B and C, have been identified in E. histolytica but C is absent in E. dispar. Proteolytic enzymes such as collagenase and cysteine proteinases and cytolytic enzymes like phospholipase A are important. Collagenase activity is mainly accumulated in electron-dense granules. Cysteine proteinase is encoded by six genes, of which EhCP5 is exclusively present in E. histolytica. PMID- 10531720 TI - Gastrointestinal endoscopy training in India. AB - A structured endoscopy training program with clear goals for proper teaching and evaluation serves to alleviate apprehensions in the minds of trainees regarding this crucial area. It also ensures that training is acquired not in isolation but in the setting of ongoing patient care, so that the emphasis is on how the procedure fits into the overall management plan for the patient. By specifying the details of the endoscopy unit set-up, the qualifications of the trainer and the number of procedures to be performed by the trainee, it is hoped that uniformity will be produced in the quality of training imparted, whether it be in a teaching or a non-teaching hospital. The end-product of such training, through the DM/MCh or the DNB stream, is a gastroenterologist who is also a certified endoscopist, capable of performing all standard diagnostic and therapeutic procedures. A further period of focused training for 1 to 2 years is required to achieve the level of competence expected of an advanced therapeutic endoscopist. There is little room for short-term training courses in endoscopy for the basic training of an endoscopist, although such courses are useful as CME activities, for the maintenance and renewal of skills of the trained endoscopist, as well as providing him with exposure to new and evolving therapeutic techniques. Efforts at improving and standardizing the training and practice of GI endoscopy in India are likely to remain exercises in futility without the active and dynamic involvement of all the leading professional societies in the country. The need of the hour is the establishment of technical committees for laying down standards in training and practice of GI endoscopy that should be voluntarily approved by all these societies so that they may then be implemented by the State medical councils and the MCI. A move in this direction from within the profession is far more appropriate and is also likely to find greater acceptance than such moves imposed from above, at the behest of judicial authorities. A system of hospital accreditation committees for large public and private sector hospitals offering endoscopy services, supervised by the accreditation committee of the State medical council, needs to be established. Clinics and nursing homes offering these services also need to be approved by the same committee after meeting standards similar to those laid down for larger hospitals. Mechanisms for audits of performance and outcome of endoscopic procedures as well as periodic participation in CME activities for maintenance of skills and expertise need to be established and linked to periodic renewal of credentials for practising GI endoscopy. Procedures for credentialing for new endoscopic techniques need to be established. The path ahead is long and arduous but we must tread it for it will only become more difficult if we procrastinate. PMID- 10531721 TI - Ulcerative colitis and immune thrombocytopenia: a report of two cases. AB - Ulcerative colitis is known to be associated with autoimmune diseases. We report two patients with coexistent ulcerative colitis and immune thrombocytopenia, a rare association. PMID- 10531722 TI - Teratocarcinoma presenting with duodenal metastases diagnosed on endoscopic fine needle aspiration. AB - Upper gastrointestinal bleed as the first symptom of metastatic testicular tumors is rare. We describe a 17-year-old man who presented with upper gastrointestinal bleed; endoscopic fine needle aspiration cytology from a duodenal mass suggested germ cell tumor, which was later confirmed on histology of the testis. PMID- 10531723 TI - Primary melanocarcinoma of stomach. AB - Melanomas of the gastrointestinal tract are infrequent. Among primary melanomas, melanoma of the stomach is a rarity. We report a patient with primary melanocarcinoma of the stomach. PMID- 10531724 TI - Hepatic mesothelioma. AB - We report a 70-year-old woman with mesothelioma in the liver. The tumor was excised, and the patient is well two years later. Light microscopy of the tumor showed papillary and tubular pattern, in addition to areas of densely packed cells arranged in fascicles. These features and the results of histochemistry and immunohistochemistry indicate a probable histogenesis from submesothelial connective tissue. PMID- 10531725 TI - Hepatic sarcoidosis responding to chloroquine as steroid-sparing drug. AB - We report a 59-year-old lady who presented with exertional dyspnea and was diagnosed to have sarcoidosis. She responded to steroids, but one year later developed abdominal symptoms and was found to have hepatosplenomegaly. Liver biopsy showed non caseating granulomas. As she had developed steroid-induced diabetes she was started on chloroquine and responded well with regression of the liver and spleen during one year of treatment. PMID- 10531726 TI - Double gall bladder with two disease processes. AB - A double gall bladder is a rare congenital anomaly which is usually diagnosed by preoperative ultrasonography. Either one, or both lobes, of the double gall bladder, may be diseased. We report a patient in whom the two lobes were affected by different disease processes, namely, cholesterosis, and cholelithiasis with mucocele. PMID- 10531728 TI - Asymptomatic T-tube remnant in common bile duct. AB - A 46-year-old lady presented with itching, five years after a primary common bile duct repair following cholecystectomy. Prior to this she underwent an interno external biliary drainage. At laparotomy the horizontal limb of a T-tube was found in the common hepatic duct. Eleven months after a Roux loop hepatico jejunostomy she is asymptomatic. PMID- 10531727 TI - Coexisting carcinoma and tuberculosis of stomach. AB - We report a rare association of carcinoma and tuberculosis of the stomach. It is difficult to explain the simultaneous occurrence or a causal relationship of the two diseases. PMID- 10531729 TI - Trend of hepatitis B virus infection in southern Indian blood donors. PMID- 10531730 TI - Tc-99m sucralfate scintigraphy in ulcerative colitis. PMID- 10531731 TI - In-house rapid urease test kit and commercial kit: which is better? PMID- 10531732 TI - Mucocutaneous exposure to body fluids during endoscopy. PMID- 10531733 TI - The single-tooth implant as a standard of care. PMID- 10531734 TI - Evaluation of bone-implant integration: efficiency and precision of 3 methods. AB - Computer-assisted planimetry, computer-assisted lineal analysis, and point counting stereology have been compared with respect to their reproducibility and the time required to analyze bone-implant integration. Sections of 6 threaded dental implants selected from a bone augmentation experiment for their wide range of new bone formation were analyzed by each method 3 times. The bone density and percentage of osseous integration were evaluated at 4 sites around each implant section. It was found that computer-assisted planimetry demonstrated a modest but significantly greater variance (P < .05) in bone density estimates when compared to the computer-assisted lineal analysis and point-counting methods. Computer assisted planimetry requires a different method of measuring each parameter and separate fields of view to evaluate fields distant from the implant. However, this can all be accomplished with line probes, as in computer-assisted lineal analysis, which extend from the implant surface into the surrounding alveolar bone. Whereas computer-assisted planimetry requires a separate identification of the perimeter of each field to be analyzed (next to and distant from the implant), computer-assisted lineal analysis allows expansion of the field to be evaluated without creating a new field of view. Also, following a limited learning curve, both point-counting and computer-assisted lineal analysis required less time to complete than did computer-assisted planimetry. PMID- 10531735 TI - Survival of the Branemark implant in partially edentulous jaws: a 10-year prospective multicenter study. AB - A total of 127 partially edentulous patients, treated according to the Branemark protocol, was followed for 10 years after completion of prosthetic treatment. The patients ranged in age from 18 to 70 years, and 57% were female. Four hundred sixty-one implants were placed in 56 maxillae and 71 mandibles. In 125 patients, 163 fixed partial prostheses were attached to the implants; a majority of the prostheses (83%) were located in posterior regions. At the end of the 10-year period, 73% of the implants could be traced either as failed or in function, providing cumulative implant survival rates of 90.2% and 93.7% for the maxilla and mandible, respectively. Of the original fixed prostheses, 63% (cumulatively 86.5%) were still in use, whereas the level of continuous cumulative prosthesis function, including primary and remade restorations, was 94.3% at the end of the evaluation period. Marginal bone resorption at the implants was low (mean = 0.7 mm), and mucosal health was good. No severe complications apart from the above mentioned implant and prosthetic failures were reported. The Branemark Implant System is a safe and predictable method for restoring partially edentulous patients, as demonstrated by this 10-year follow-up investigation. PMID- 10531736 TI - Treatment of edentulism using Astra Tech implants and ball abutments to retain mandibular overdentures. AB - The goal of this study was to provide evidence to support simplified treatment of mandibular edentulism using denture fabrication and implant placement to circumvent the need for second-stage surgeries or prosthodontic superstructures. A 5-year prospective clinical trial is reported, which involved treatment of mandibular edentulism using the single-stage surgical placement of a TiOblast microthreaded titanium screw implant with immediate replacement of a relieved mandibular overdenture and eventual retention of the overdenture with reduced ball abutments. Fifty-eight patients were treated; 116 implants were placed using a single-stage surgical approach, with a duplicate mandibular denture as the tomographic/surgical template. Mandibular dentures were relieved and relined with a tissue conditioning material and placed immediately after implant surgery. After 3 months, Conical Seal Design ball abutments were placed and attachments were secured in the overdentures by heat-polymerizing laboratory reline methods. Five of the 116 consecutively placed implants failed at 2 to 4 months, providing an immediate implant survival rate of 95.69% at the time of attachment connection. Pain and inflammation were not common to all failures, and infection was not reported in any of the 5 failures. The immediate placement of implants by a single-stage surgical procedure in the parasymphyseal region of the mandible, followed by placement of a relined mandibular denture, results in predictable and asymptomatic healing of implants that display the clinical and radiographic features of osseointegration. Encouraging results at the immediate observation period (attachment connection) must be tempered by the need for prudent and detailed clinical and radiologic evaluation over the 5-year trial period. PMID- 10531737 TI - Prevention of bacterial leakage into and from prefabricated screw-retained crowns on implants in vitro. AB - Previous in vitro studies have shown that a mean gap of less than 4 microns between prefabricated crowns and implants of the Ha-Ti implant system is not a barrier to infiltration by Staphylococcus aureus. These studies confirmed earlier in vivo work showing that a multitude of oral microorganisms could colonize and infiltrate these gaps. In the present investigation, 30 Ha-Ti implant-crown assemblies were tested for bacterial leakage after the gaps were sealed with the chlorhexidine-containing varnish Cervitec. S. aureus leakage into the totally submerged test specimens was detected in 1 of 5 samples incubated for 4 weeks, while no leakage was detected in specimens incubated for 3, 5, 6, 7, and 8 weeks. When the sealed test specimens were partially submerged (that is, excluding the screw hole of the crown) and incubated for 3 to 11 weeks, none of the internal surfaces of the 30 test specimens manifested contamination. The clinical relevance of gap sealing in maintaining inflammation-free marginal mucosa and in achieving clinically successful treatment of peri-implantitis has yet to be determined. PMID- 10531738 TI - Crystallographic characteristics of plasma-sprayed calcium phosphate coatings on Ti-6Al-4V. AB - The purpose of this study was to investigate the crystallographic characteristics of 3 sets of plasma-sprayed hydroxyapatite (HA) coatings prepared with different degrees of crystallinity on Ti-6Al-4V substrates. X-ray diffraction analyses were performed on the coatings to determine mean percent crystallinity, calcium phosphate phases present, average crystallite size, and residual strain. The mean percent crystallinity for the 3 sets of coatings ranged from 49 to 60%. The coatings that achieved the highest crystallinity consisted almost entirely of HA. As the coating crystallinity decreased, increasing amounts of alpha- and beta tricalcium phosphate and tetracalcium phosphate were detected. The mean HA crystallite size for the 3 sets of coatings ranged from 0.02 to 0.05 micron. Differences in mean interplanar spacing for selected crystallographic planes of HA, compared with the pure ICDD (International Center for Diffraction Data) powder standards, implied that the coatings were in an uneven state of tensile strain. PMID- 10531739 TI - Life table analysis and clinical evaluation of oral implants supporting prostheses after resection of malignant tumors. AB - Seventeen mostly elderly patients, 13 men and 4 women, were consecutively admitted for implant-prosthodontic treatment after they had undergone resection of malignant tumors in the oral cavity. A total of 53 dental implants (ITI Straumann) was placed, 12 in the maxilla, 41 in the mandible. The prosthetic rehabilitation consisted of overdenture therapy in 15 patients, and 2 patients were treated with fixed partial prostheses. Thirty-three implants were prescribed for patients who received radiotherapy either before or after implant placement. The average dose varied between 50 and 74 Gy. Eighteen implants were located in grafted bone from the fibula, scapula, or hip. For 2 patients, hyperbaric oxygen therapy was also prescribed after osteoradionecrosis had developed. One implant was lost before prosthetic loading. During an observation period of up to 7 years after loading, 3 more implants were removed. All implant losses occurred in the mandibles of patients who had received radiotherapy. A life table analysis was performed, and the cumulative survival rates, calculated for 2, 3, and 5 years, were 93%, 90%, and 90% respectively. No failures or complications were observed with technical components of the implants or prostheses. All prostheses could be maintained during the entire observation time. Although in the present investigation the survival rate of implants was slightly lower than under standard conditions, the treatment with implant-supported prostheses seemed to be advantageous for patients who had undergone intraoral resections. PMID- 10531740 TI - Measuring abutment/implant joint integrity with the Periotest instrument. AB - Maintenance of the integrity of the abutment/implant interface is essential and is dependent on the abutment screw retaining a preload. Evaluation of this joint is usually done by manual assessment. The purpose of the current study was to determine whether the Periotest instrument could be used to evaluate abutment screw loosening. A custom-designed apparatus was constructed to measure abutment screw loosening. Abutment screws were torqued to 10, 20, 32, and 45 Ncm and then loosened. Objective assessment of screw loosening was carried out with the Periotest device. Subjective evaluation was done by 3 experienced clinicians. The Periotest was found to be more sensitive than manual detection of abutment screw loosening. With a change of 2 in the Periotest value, it was found that the tensile preload in the joint was lost. While the Periotest was more sensitive than manual evaluation, the instrument was not sensitive enough to indicate deterioration of abutment screw loosening prior to loss of tensile preload. PMID- 10531741 TI - A histometric comparison of smooth and rough titanium implants in human low density jawbone. AB - The aim of this investigation was to conduct a comparative histometric analysis of bone-implant interface between a rough titanium surface and smooth implants in low-density human jawbone after 3, 6, and 12 months of submerged, undisturbed healing. Six adult volunteer patients undergoing standard implant placement were enrolled in this project. Each patient received 1 smooth and 1 rough implant. After 3, 6, and 12 months, the implants were harvested for histometric analysis. The values of bone-implant contact were the following: 3 months smooth 6.2%, 3 months rough 58.9%, 6 months smooth 3.55%, 6 months rough 72.9%, 12 months smooth 6.7%, and 12 months rough 76.75%. The results showed that in low-density bone the rough surface dramatically enhanced the amount of bone-to-implant contact. Because of the small number of implants examined, definite conclusions cannot be drawn, even though the statistical analysis showed significant differences between the smooth and rough groups (P = .0129; F = 76.065). Nevertheless, a trend was evident in these observations: while a rough implant surface may enhance the rate of osseointegration, it is not able to significantly change the bone density, and an implant placed low-density bone is at a higher risk of failure when occlusal loading begins. PMID- 10531742 TI - A histomorphometric study of bone reactions to titanium implants in irradiated bone and the effect of hyperbaric oxygen treatment. AB - The present study was undertaken to histomorphometrically analyze early peri implant bone tissue reactions that occur after radiotherapy and to determine whether hyperbaric oxygen therapy (HBO) affects bone tissue at the microscopic level by altering bone morphology. Twelve rabbits received a single dose (15 Gy) of cobalt60 radiation to one hind leg and the other hind leg served as a control. Titanium screws were placed into the femur and tibia directly after irradiation. Six animals received HBO during the first 4 postoperative weeks. After 8 weeks of follow-up, bone specimens containing the screws were prepared for histomorphometry. Bone-metal contact and the amount of bone in the thread areas and in the mirror areas were measured in a blinded manner. Periosteal bone formation and bone remodeling decreased after irradiation; also after HBO treatment. Hyperbaric oxygen therapy improved bone formation in nonirradiated bone and to some extent also in the irradiated bone. Bone maturation was improved in the HBO animals after irradiation. It was concluded that irradiation reduces the capacity for osseointegration of titanium implants. Hyperbaric oxygen treatment may improve bone formation and especially has positive effects on bone maturation after irradiation. PMID- 10531743 TI - Maxillary antral-nasal inlay autogenous bone graft reconstruction of compromised maxilla: a 12-year retrospective study. AB - During a 12-year period (1984-1996), 118 maxillary inlay autogenous bone grafts and 248 commercially pure titanium threaded root-form endosseous implants were placed in 54 consecutively treated patients with compromised maxillary bone. In this retrospective clinical study, 3 groups of patients were reviewed, group selection being based on anatomic location and surgical access to the recipient site. Group 1 included patients with bone grafts placed in the antrum floor via an intraoral antrostomy exposure, group 2 included patients with bone grafts placed in the nasal floor via an anterior intraoral nasotomy exposure, and group 3 included patients with bone grafts placed in the antral and nasal floor via an intraoral Le Fort I osteotomy downfracture exposure. Each patient received an implant-supported dental prosthesis. For the combined 3 groups, survival rates were 87% for endosseous implants and 100% for autogenous bone grafts. The success rate for the dental prostheses in the 3 groups was 95%. Sixty-nine dental prostheses functioned a mean of 57.1 months, whereas 3 prostheses required remaking because of implant loss. Of the medical and mechanical risk factors tabulated in this study, current use of nicotine, history of sinusitis, molar site implant placement, and shorter implant lengths had the most influence on implant failure. PMID- 10531744 TI - Rate of pull-out strength gain of dual-etched titanium implants: a comparative study in rabbits. AB - The purpose of this study was to investigate the rate of pull-out strength gain of an etched titanium implant surface. Rabbit tibiae were used to compare machined titanium and proprietary dual-etched titanium implants. Two custom cylindric implants (3 mm in diameter and 4 mm in length) were placed in each right anteromedial tibia in 31 rabbits. At weeks 1, 2, 3, 4, 5, and 8, the implants in 5 rabbits were subjected to failure shear loading in a pull-out test. For shear failure testing, each tibial segment was mounted in a precision alignment jig, and an Instron pull-out test was performed on each implant. Beginning at week 3, there was a statistically significant difference (P < .01) between the dual-etched and the machined implants. There was a significant increase in strength for dual-etched implants between week 5 and week 8, while the machined implants did not show an increase during this time interval. The etched implants maintained a significantly greater pull-out strength for the remainder of the study, with a 3.2-fold greater mean strength at 8 weeks, equivalent to 6 months in humans. At 3 weeks, the etched implant's strength exceeded the strength that the machined implant had achieved at 8 weeks. In short term healing in the rabbit tibia, the dual-etched surface demonstrated a more rapid rate of pull-out strength gain than the machined surface and remained significantly stronger throughout the 8 weeks of the study. PMID- 10531745 TI - Histomorphometric analysis of a half hydroxyapatite-coated implant in humans: a pilot study. AB - The aim of this study was to compare the characteristics of the bone-to-implant interface of hydroxyapatite-coated and non-coated commercially pure titanium threaded implants after different periods of healing in humans. To eliminate possible variations of the results from differences in bone quality and in surgical techniques used in the different test and control sites, only one half of each implant was coated with hydroxyapatite. The coated portions of the implants showed a tendency toward a higher percentage of direct bone-to-implant contact at each period of healing that was observed, although the small number of specimens does not allow definitive conclusions to be made. PMID- 10531746 TI - Outcome of treatment with implant-retained dental prostheses in patients with Sjogren syndrome. AB - The purpose of this investigation was to evaluate the outcome of treatment with implant-retained prostheses in patients suffering from Sjogren syndrome. Eight women were included in the study; all had suffered oral symptoms of Sjogren syndrome for many years. Seven patients were edentulous in both arches, and 1 patient was edentulous in the maxilla only. All patients reported poor or very poor comfort levels with their conventional dentures. It was the intention to treat each arch that showed subjective and objective denture problems with a complete fixed prosthesis after placement of 6 implants. In all, 54 Branemark dental implants were placed in these patients. No implants were lost, but 7 implants in 4 patients were clinically not osseointegrated at the time of the abutment connection procedure. Because of nonosseointegrated implants and lack of jawbone, 3 arches were treated with an implant-retained overdenture. Fixed prostheses were made with a titanium framework of premachined components welded together (Procera) and acrylic resin teeth and flanges. Patients answered a questionnaire regarding their oral function before the onset of treatment and 1 month and 2 years after treatment. An average radiographic bone loss of 0.7 mm from the time of implant placement to 1 year after treatment was observed; additional bone loss of less than 0.6 mm was recorded 4 years after treatment. During the first year of function 2 implants lost osseointegration. No prostheses were lost or remade. Treatment with implant-retained prostheses considerably increased the prosthetic comfort and function of the patients. Two years after prosthetic treatment, only 1 patient indicated poor comfort of the prostheses, while the remaining patients reported good or very good comfort levels. PMID- 10531747 TI - Retrospective review of grafting techniques utilized in conjunction with endosseous implant placement. AB - Bone resorptive patterns may prevent the ideal placement of endosseous implants. Numerous techniques have been described to create a more favorable surgical site for implant placement. This retrospective review was conducted to determine the frequency of need for implant site preparation in an outpatient clinical setting. In addition, different techniques of surgical site preparation were evaluated to determine their frequency of use and surgical outcome. A history review was conducted of all consecutively treated partially edentulous patients between January 1993 and December 1997. This review evaluated the number of implants placed, the age and gender of patient, the type of graft used, and the status of the implant. In all, 542 patients were seen in this time interval, with a total of 1,313 implants placed. Implant site preparation was needed in 4.4% of the patients, with the requirement for grafts occurring more frequently in the maxilla. Implant site preparation is a relatively infrequent requirement in the general population. Grafts are required more frequently in the maxilla than in the mandible. Complications following grafting were relatively infrequent and were not severe. PMID- 10531749 TI - [Mycotoxins and Cuban encephaloneuromyelopathy]. PMID- 10531748 TI - Long-term results after placement of dental implants: longitudinal study of 1,964 implants over 16 years. AB - In a retrospective study, Kaplan-Meier implant survival analyses were conducted on 883 patients with 1,964 implants of various systems placed, followed up, documented, and statistically evaluated at an oral surgery and dentistry practice between January 1981 and January 1997. The goal of this study was to evaluate the success of osseointegrated implants of the Branemark, Frialit-1 (Tubinger Implant), Frialit-2, and IMZ systems and Linkow blade implants. For all systems, mandibular implants were generally more successful than maxillary implants. The preprosthetic loss rate was 1.9%, and 4.3% of implants were lost after prosthetic treatment. The lowest loss rates were seen with implants in intermediate and distal extension spaces and with single-tooth replacements using IMZ, Frialit-2, and Branemark implants. In edentulous arches, implants of the IMZ and Branemark systems had the lowest failure rates. PMID- 10531750 TI - [Ultrastructural identification of Ehrlichia sp in an experimentally infected dog in Venezuela]. AB - This study is the first report made in Venezuela concerning the ultrastructural characteristics of Ehrlichia sp in mononuclear blood cells from an experimentally infected dog. The animal developed clinical manifestations characteristic of the infection, and typical intracitoplasmic inclusion bodies were clearly seen in blood smears stained with modified Giemsa examined by light microscopy. Microorganisms were visualized by transmission electron microscopy. The cytoplasmic inclusions, consisted of membrane-lined vacuole-containing elementary bodies. The organisms were extremely pleomorphic. Elementary bodies were surrounded by two distinct membranes and each was constituted by electro-dense granules. These findings corresponded to the described electron microscopy morphology which characterizes the Ehrlichia genus. PMID- 10531751 TI - [Cytogenetic findings in ductal carcinoma of the breast]. AB - Breast cancer in women is an important medical problem with public health and social implications. In spite of its great clinical importance, little is known about the cytogenetic features of breast carcinomas. Chromosomal abnormalities in some malignant tumors have been used for diagnosis and prognosis or for localizing genes involved in the pathology malignancies. In this report, we present the chromosomal abnormalities found in 32 primary breast ductal carcinomas. The tumor samples were studied using the technique for short-term culturing and cytogenetic analysis with G-banding. Only one tumor with normal karyotype was observed. Thirty one (99%) of the tumors had chromosomal abnormalities including 21 (65.6%) in which chromosome 1 was involved (trisomy, monosomy or structural abnormalities of the type t(1q;2p) and del(1q42). Other recurrent anomalies such as del(12p); del 4(p); +7; +8; -7; -3; were observed. The significance of these findings and their role in tumorogenesis will become more evident with close follow-up of women who have tumors with an abnormal karyotype. PMID- 10531752 TI - [Anemia in indigenous population of the West of Venezuela]. AB - The purpose of the study was to investigate the frequency of nutritional anemia among western venezuelan indians. Three hundred and ninety nine Yucpa indians from the communities of Aroy, Marewa and Peraya were studied. The concentrations of hemoglobin, serum iron, total iron binding capacity, serum ferritin, serum folate and serum vitamin B12 and the frequency of anemia and nutrient deficiency were determined. Anemia was found in 71.7% of people from Aroy, 52.25 from Marewa and in 74.4% from Peraya. No nutrient deficiencies were found in 48.1% of cases with anemia, while iron deficiency anemia was present in 39% of the population studied, and folate and or vitamin B12 deficiency were associated with anemia in only 12.9% of cases. The high frequency of anemia, unrelated to nutrient deficiency, among the Yucpa indians, is attributed to the prevalence of chronic infectious diseases such as hepatitis and parasitic infections, as well as skin and respiratory infectious processes. PMID- 10531753 TI - [Detection of beta thalassemia by the technique of refractory amplification of mutation systems (ARMS-PCR)]. AB - beta Thalassemia (Thal) mutations were studied in DNA from 80/159 patients with hemolytic anemia and high levels of Hb A2 by the amplification refractory mutation system technique (ARMS-PCR). This method detects point mutations and insertions or deletions of just a few nucleotides in the beta globin gene by the polymerase chain reaction of allele-specific priming. In 43/80 patients with different clinical presentations of beta Thalassemia and 37/80 compound heterozygous for hemoglobinopathies and beta Thalassemia the most frequent mutation found was the -29 (of African origin), followed by the CD39 (of Mediterranean origin) and in a lower frequency also was found the -88, the IVSI-6 and the IVSI-110. We conclude that this technique is an useful approach in determining the beta thalassemia mutations in population surveys, because it allows to make a differential diagnosis between beta Thalassemia minor and individuals with high levels of Hb A2. It helps to clarify the diagnosis of patients with structural hemoglobinopathies that also presents high levels of Hb A2. PMID- 10531754 TI - [Pulmonary atresia with intact interventricular septum. Report of a case diagnosed by fetal echocardiography]. AB - Pulmonary atresia with intact ventricular septum (PA/IVS) is a rare cardiac congenital malformation involving the right ventricle (RV) in which the communication through pulmonary valve, is absent. A case of a congenital heart disease (CHD) consisting of pulmonary atresia with intact ventricular septum (PA/IVS) and small right ventricle (RV) or type I of Greenwold, coming from a twin pregnancy in which the other was an inutero dead fetus, is reported. Although the case was referred after the death of one of the fetuses, the prenatal diagnosis was made by the use of echocardiographic studies and confirmed by anatomopathology of the still-born fetus with the CHD. The very useful echocardiographic prenatal diagnosis and surgical therapeutical options are emphasized and discussed. PMID- 10531756 TI - Legal issues in the delivery of alternative medicine. AB - As use of alternative therapies grows, there appears to be heightened concern among health care professionals about the liability implications of delivering these therapies. Little is known about malpractice law in this area. We begin by reviewing the type and frequency of claims brought against alternative medicine practitioners and by analyzing the standard of care to which these practitioners are held when sued. Next we turn to the standard of care question as it relates to physicians (MDs/DOs) who incorporate alternative therapies into their practices. Few cases have addressed this question to date. We argue, however, that when courts decide cases at the intersection between conventional and alternative medicine, they may judge conduct according to standards enunciated by: 1) alternative medicine practitioners who regularly deliver the treatment at issue, 2) physicians who have established similar practices, or 3) conventional practitioners. This latter possibility should be taken seriously; it raises troubling questions for physicians at the outset of the negligence inquiry. Available case law highlights the importance of ensuring that patients are fully informed about any alternative therapies they elect to receive, as well as any conventional treatments they may be foregoing, and that patients expressly consent to treatment in light of this information, preferably in writing. PMID- 10531755 TI - Toward integrated medicine. PMID- 10531757 TI - Research on complementary and alternative therapies for cancer: issues and methodological considerations. AB - Recent evidence suggests that at least one cancer patient in three uses some form of complementary and alternative medicine (CAM). We conducted a review of the published research on the efficacy of these treatments for breast cancer, which resulted in some observations about the current state of research and guidelines for future research. Although many of the papers reported encouraging results, the preponderance of phase I and II trials and other limitations precluded definitive conclusions about the efficacy of the treatments reported in these studies. A growing institutional base in this country has begun to facilitate improved research on CAM for cancer, yet many gaps remain. For example, there are no published reports of clinical trials or observational studies with survival endpoints for treatment agents used by many cancer patients. Clinical trials of a few CAM treatments are now in progress, but the results will not be available for several years. More complex and customized treatments may require innovative study designs and practitioner-investigator collaborations. Given the mounting evidence that CAM treatments are biologically active as well as widely used, CAM research may affect cancer outcomes. PMID- 10531758 TI - Hormone-modulating herbs: implications for women's health. AB - Women in the United States are increasingly turning to botanical medicines to treat conditions throughout their life cycles. Many herbs traditionally used for women's health conditions have been found to contain phytoestrogens. Phytoestrogens and their metabolites can bind estrogen receptors and can have both estrogenic and anti-estrogenic effects. Many women are attracted to the idea of using phytomedicine as an alternative to hormone replacement therapy. It is unclear, however, whether these herbs are safe for women at risk for breast cancer or its recurrence. This paper considers the estrogenicity of herbs such as black cohosh (Cimicifuga racemosa) and the implications for women's health. PMID- 10531759 TI - Botanical medicines with gynecological anticancer activity: a literature review. AB - This article reviews the existing scientific literature reporting effects of botanical substances on the prevention and treatment of gynecological cancers. Anticancer effects were reported for 14 of 27 herbal substances searched for, 8 of which had reported effects specifically in gynecological cancer models. Research reviewed included in vitro studies in gynecological cancer cell lines, animal studies, an ex vivo study, and an epidemiological study. No clinical trials on gynecological cancer prevention or treatment were found for any of the 8 identified agents. Despite the increasing use of botanical medicines in the prevention and treatment of cancer in general, there is a paucity of studies describing their efficacy or safety in gynecological cancer. Given the prevalent use of botanical medicine in complementary and alternative cancer therapy, the need for research to evaluate safety and efficacy using both in vitro and in vivo methods is pressing. PMID- 10531760 TI - Dietary supplement-drug interactions. AB - Recent surveys show that 18% of adults in the United States use prescription drugs concurrently with herbal or vitamin products, placing an estimated 15 million patients at risk of potential drug-supplement interactions. Despite this widespread concurrent use of conventional and alternative medicines, documented drug-herb interactions are sparse. This review focuses on possible interactions between drugs and herbal medicines used for phytoestrogen-hormone and antiplatelet-oral anticoagulant therapy. Interactions with phytoestrogens are purely speculative, based on competitive estrogen-receptor binding or antiestrogenic effects. In contrast, several case reports document bleeding complications with Ginkgo biloba, with or without concomitant drug therapy. Case reports are also suggestive of interaction between warfarin and dong quai or Panax ginseng. Recommendations for counseling patients at highest risk of adverse interactions are given. PMID- 10531761 TI - Use of complementary and alternative medicine among African-American and Hispanic women in New York City: a pilot study. AB - OBJECTIVE: To explore the use of complementary and alternative medicine (CAM) among African-American and Hispanic women residing in New York City, including use of specific treatments and practitioners, perceived effectiveness of CAM, and culturally specific words or expressions for CAM. METHODS: Focus groups were conducted with two groups of African-American and two groups of Hispanic women (age 18-40 and 41-80) as preparation for the development of a quantitative instrument to assess the prevalence and determinants of CAM use among women of various ethnic backgrounds. Participants were recruited using a standard random digit dial procedure. RESULTS: The most commonly used CAM remedies were teas and herbs, vitamins and nutritional supplements, prayer and spiritual healing, meditation and relaxation techniques. Practitioners most frequently seen were chiropractors, herbalists, and acupuncturists. Use of alternative remedies and practitioners, particularly the latter, was most common among older women in both groups. Younger Hispanic women reported the most skepticism toward CAM, especially when it was used by relatives as a substitute for conventional medical care. Overall, these African-American and Hispanic women used CAM for a wide range of health conditions and for prevention. Few racial and ethnic differences emerged in patterns of CAM use for either self-care or treatment by practitioners, but there was a distinct age variation, especially in attitudes toward CAM. PMID- 10531762 TI - Distress and conception in infertile women: a complementary approach. AB - OBJECTIVES: To examine the relationship of pretreatment psychological distress and demographics to conception in infertile women attending a group cognitive behavioral treatment program. METHODS: Pre- and postprogram psychological measures and live birth rates were collected for 132 infertile women attending a ten-session group cognitive-behavioral treatment program. Subjects completed the Beck Depression Inventory (BDI) and the Symptom Checklist-90 (Revised) (SCL-90R). Conceptions that resulted in live births within six months of completing the program were noted. RESULTS: Women who conceived viable pregnancies within six months of the program had higher levels of psychological distress at program entry. Using logistic regression analysis, the best predictors of viable birth were younger age and a higher score on the global severity index of the SCL-90R. Significant pre- to postprogram psychological improvement was demonstrated by the SCL-90R and the BDI. Forty-two percent of the sample conceived viable pregnancies within six months of completing the program. CONCLUSIONS: Preprogram psychological distress and younger age were associated with significantly higher viable pregnancy rates. PMID- 10531763 TI - Dietary supplements: current FDA activities. PMID- 10531764 TI - AMWA physicians' views of and experiences with complementary and alternative medicine. PMID- 10531765 TI - Family planning training in Maryland family practice and obstetrics/gynecology residency programs. AB - OBJECTIVE: To examine the extent of family planning and abortion training in Maryland family practice (FP) and obstetrics/gynecology (OB) residency training programs. METHODS: All final-year residents in every FP and OB residency training program in Maryland were asked in 1998 how many cases (0, 1-10, > 10) of ten methods of contraception and abortion they had managed. RESULTS: Seventy-five percent (55) of the 73 residents responded. Fifty percent of FP residents had never inserted an intrauterine device (IUD), 43% had never inserted an implant (Nor-plant), 37% had never prescribed emergency contraceptive pills, and 30% had never fitted a diaphragm. Ninety-seven percent of FP residents had no experience with elective termination of pregnancy, and 83% had no experience with sterilization. Twenty percent of OB residents had never inserted an IUD, 16% had never inserted implants or prescribed emergency contraceptive pills, 20% had never fitted a diaphragm, and 36% had no experience in elective termination of pregnancy. Not one FP resident had inserted or fitted more than ten IUDs, implants, diaphragms, or cervical caps. Except for oral and injectable contraception, the majority of OB residents had not managed more than ten cases of any other reversible contraceptive method: 80% had not inserted more than ten IUDs, 72% had not inserted more than ten implants, 88% had not fitted more than ten diaphragms, and 100% had not fitted more than ten cervical caps. CONCLUSION: These survey results indicate a need for more formal instruction in most contraceptive methods for OB and FP residency programs in Maryland. This study concurs with previous national studies showing deficits in family planning training. PMID- 10531767 TI - Early detection of prostate cancer. PMID- 10531766 TI - Physician use of diagnostic codes for child and adult abuse. AB - OBJECTIVE: To report on physician use of diagnostic codes for child and adult abuse according to national medical care utilization data. METHODS: Secondary data analysis was performed on the National Ambulatory Medical Care Survey (NAMCS) of office-based physician visits and the National Hospital Ambulatory Medical Care Survey (NHAMCS) of visits to hospital emergency and outpatient departments for 1993 to 1996. Both databases describe physician, patient, and visit characteristics, and variables include up to three diagnoses per visit. RESULTS: Only 93 diagnoses of child or adult abuse were coded for 351,359 patient visits during the four years. As we would expect, child abuse was diagnosed more often than adult abuse (67 v 26), and the majority of cases (n = 57) were visits to emergency departments. CONCLUSION: Diagnostic codes for abuse are not often used. Because these codes represent an important tool for reporting the prevalence and incidence of abuse, such documentation could lead to greater support for health care policies and resource allocation for victims of abuse. Lack of awareness about the diagnostic codes for abuse may be one explanation for underuse, but other barriers are also discussed. PMID- 10531768 TI - Evaluation of prostate specific antigen as a tumor marker in cancer prostate. AB - The present study was undertaken to evaluate the prostate specific antigen (PSA) alongwith other diagnostic methods as an application for a screening test, tumor marker and its relation to post surgical situation. The PSA has shown a sensitivity of 73.3% and specificity of 77.2%. The predictive value for positive PSA was 57% and for negative test was 66.6%. Local standards for PSA values in Pakistani community need to be established. The PSA test, inspite of its low specificity holds good promise for its contributory role as a tumor marker in prostate cancer. PMID- 10531769 TI - The spectrum of bacterial infections in febrile neutropenic patients: effect on empiric antibiotic therapy. AB - The aim of this retrospective analysis was to look at the spectrum of bacterial isolates and their resistance patterns to the commonly used antibiotics in the setting of febrile neutropenia. A total of 127 bacteria were isolated from patients with acute leukemias, lymphoproliferative disorders, aplastic anaemia and various solid tumours. Fifty-four percent organisms were gram negative; while the rest were gram positive. E. coli, pseudomonas aeruginosa, staphylococcus aureus, enterococcus and streptococci were the commonly isolated organisms. Forty eight percent organisms were isolated from blood, 16% from urine, 13% from wounds and superficial abscesses and 11% from respiratory tract. E. coli exhibited a great degree of resistance to the commonly used antibiotics, such as pipericillin (70%), ofloxacin (50%) and aztreonam (50%). Pseudomonas and klebsiella also showed varying degree of resistance against the antibiotics. Staphylococcus aureus and staphylococcus epidermidis were almost universally resistant to penicillin and showed a variable degree of resistance to other antibiotics too. Compared to the previous reports, the pattern of bacterial isolates and their resistance to antibiotics has changed over the past years. Aminoglycosides and third generation cephalosporins seem to be the choice of antibiotics for the upfront management of febrile neutropenic patients. PMID- 10531770 TI - Awareness of health care personnel about preventive aspects of HIV infection/AIDS and their practices and attitudes concerning such patients. AB - The study was carried out at a tertiary care hospital in Lahore to assess the level of awareness of health care personnel regarding various preventive aspects of HIV/AIDS and determine their practices and attitudes while dealing with such patients. Study population consisted of 147 (55.68%) physicians, 82 (31.06%) nurses and 35 (13.26%) paramedics. Most of the participants had awareness regarding prevention of sexual transmission, however there were gaps in their knowledge as to how spread could be checked through the use of screened blood and sterilized syringes and instruments and perinatal transmission. One hundred and twenty three (46.5%) respondents were unaware of precautionary measures to be observed by health care providers while looking after these patients and 140 (53.03%) made improper responses when inquired about their responsibility and practices concerning HIV seropositive individuals and AIDS patients. As a result of misconceptions and ignorance, 108 (40.90%) participants' responses regarding their attitudes towards HIV/AIDS cases were incorrect and improper. PMID- 10531771 TI - Intravenous tenoxicam to treat acute renal colic: comparison with buscopan compositum. AB - Forty-seven patients with acute renal colic were treated with either tenoxicam 20 mg i.v. or buscopan compositum (hyoscine butylbromide 20 mg and dipyrone 2.5 g) i.v. in a double blind study. Renal colic was diagnosed with use of a general urine examination, intravenous urogram, ultrasonography or voiding of calculus. The severity of symptoms were assessed by a verbal six point scale. Results demonstrated that 80% of patients treated with tenoxicam and 72.7% of patients treated with buscopan compositum showed significant improvement after 1 hour. Sixty-two percent of the patients who showed initial response to buscopan compositum had pain relapse during next 24 hours and required rescue treatment with pethidine 100 mg i.m. None of the patients treated with tenoxicam i.v. had pain relapse. No side effects were reported with use of tenoxicam. It is concluded that tenoxicam i.v. was more effective than antispasmodics and has rapid onset of analgesia and prolonged action in the treatment of acute renal colic. PMID- 10531772 TI - Study of efficacy and tolerance of ketoprofen and diclofenac sodium in the treatment of acute rheumatic and traumatic conditions. AB - A comparative, multi-centre study, was conducted during June to December, 1996 to evaluate the efficacy and tolerance of Ketoprofen 100 mg Enteric Coated (EC) tablet and 100 mg intra-muscular injection; with that of Diclofenac Sodium 50 mg tablet and 75 mg intra-muscular injection in acute rheumatic and traumatic disorders. Total of 180 patients (90 per drug), were studied, 82 men and 98 women, between the ages of 18 and 75 years. The symptoms and the number of patients were backache 50, arthritis 64, frozen shoulder 32 and sprains 34. Pain was qualitatively assessed by visual analogue scale (VAS), XY pain index, pain at mobilization and the level of pain handicap. For pain (VAS 75-100) the treatment was initiated with an injectable bid, followed by tablets bid or tid. If the pain score on VAS was less than 75, tablets were given in a bid dosage. The duration of treatment was 15 days in each case. The overall complete relief of symptoms occurred in 25% (23/90) patients with Ketoprofen and in 10% (9/90) diclofenac sodium. Moderate to mild relief was found in 75% (67/90) cases with Ketoprofen and 87% (78/90) with diclofenac sodium. No pain relief was seen in 3% (3/90) with diclofenac sodium, as against no failure in pain relief in the ketoprofen group. Tolerance was found as excellent-good for ketoprofen in 72% (65/90) with diclofenac sodium in 50%, moderate to poor for ketoprofen in 28% (35/90) and with diclofenac sodium in 50% (45/90). Our results indicate that ketoprofen compared to diclofenac sodium is efficacious in acute rheumatic and traumatic injuries. Ketoprofen injection, compared to diclofenac sodium was found to be more effective in providing analgesia. PMID- 10531773 TI - Efficacy of low dose intra-dermal hepatitis B vaccination schedule. AB - Efficacy of low-dose, intra-dermal hepatitis B vaccination was assessed among sixty-one doctors and paramedical staff of Chandka Medical College Hospital, Larkana. Subjects were randomly divided into two groups. Group-A (n = 25) received 20 micrograms of purified Hepatitis B Surface Antigen (Euvax B, LG Chemicals) by i.m. injection in the deltoid by 0, 1 and 6 schedule while group B (n = 36) received 5 micrograms intra-dermally by same schedule. The mean value of HBsAb titers checked 4 weeks after the last dose in group A were 158.6 +/- 51.8 mlU/ml and that in Group B were 68.2 +/- 26.6 mlU/ml (p < 0.0001; 95% C.I. 69.2 and 111.5). Number of subjects attaining the protective titers of 10 mlU/ml in group A was 95.7% and that in group B was 90.9% (p = 0.5). We conclude that 5 micrograms intradermal hepatitis B vaccination is effective. PMID- 10531774 TI - Praziquantel for treatment of malaria. AB - The possible effect of oral praziquantel on malaria parasites was studied. Nine patients with P. falciparum and one patient with P. vivax were treated with 30 mg/kg/day of praziquantel in three divided doses for a maximum of 8 days. The results showed that eight patients had complete cure within 4-6 days of using praziquantel. Two patients with P. falciparum complicated by jaundice and severe anemia showed no response and required antimalarial drugs. One patient had bloody diarrhoea. It could be concluded that praziquantel might represent a new line for treatment of malaria. PMID- 10531775 TI - Aplastic anaemia evolving into myelodysplastic syndrome. PMID- 10531776 TI - Protocol for management of hypertension by family practitioners. PMID- 10531778 TI - International normalized ratio. PMID- 10531777 TI - "A shadow apart": a symposium on biomedical imaging. PMID- 10531779 TI - Cancer chemo-prevention--pragmatic or over-optimistic. PMID- 10531780 TI - Presenting features in Pakistani patients suffering from the antineutrophil cytoplasmic antibody--classical subtype (c-ANCA) associated vasculitis. AB - OBJECTIVES: To study the clinicopathological features in c-ANCA positive patients suffering from vasculitis with a view to find out the most common mode of presentation. STUDY DESIGN: Retrospective. SETTINGS: Department of Immunology, AFIP, Rawalpindi, MH Rawalpindi, CMH Rawalpindi, Department of Rheumatology, PIMS, Islamabad, RGH Rawalpindi, FFH, Rawalpindi. SUBJECTS: Seventeen patients suffering from vasculitis and found to be positive for c-ANCA. MAIN OUTCOME MEASURES: Clinico-pathological features at presentation. RESULTS: There were 9 males in age range 11-60 years (mean age 32.5 years) and 8 females in age range 26-42 years (mean age 32.3 years). Common presenting features were a combination of cough, blocked nose and post nasal drip 14/17 (82%) followed by nose bleed and haematuria 11/17 (65%). Six patients were demonstrated to be suffering from Wegener's granulomatosis after biopsy. C-ANCA was detected by indirect immunofluorescence in the titre range of 8-640. The auto antibody levels related to disease activity. CONCLUSION: c-ANCA associated vasculitis is a rare (17 patients in five years) but aggressive form of vasculitis. It must be suspected in patients with persistent respiratory tract related symptoms associated with fever and joint pains which fail to respond to adequate treatment for infections. The c-ANCA estimations can be utilised as sensitive and specific diagnostic and prognostic marker in this form of vasculitis. PMID- 10531781 TI - Determinants of neonatal mortality. AB - OBJECTIVES: To identify the common causes and contributory factors for high neonatal mortality in Pakistan and propose effective strategies to safeguard against it. METHODS: This retrospective study was carried out in Neonatal Unit (NNU) of Rawalpindi General Hospital (RGH) from January 1995 to December 1996. Relevant prenatal information obtained from referral forms, admission files or attendants was recorded on a proforma at the time of death of a newborn. This included obstetric and medical management of the mother, neonatal resuscitation and care, birth weight (wt) and gestational age. Cause of death was based on available clinical and laboratory data. RESULTS: During the study period, there were 3005 admissions and 268 deaths, resulting in 9% neonatal mortality. Eighty eight percent of this mortality was due to early neonatal deaths. More than 50% of deaths were unavoidable due to their critical condition at admission and occurred within first 24 hours of hospitalization. Neonatal infections and birth asphyxia were two major causes of neonatal mortality (37% and 31% respectively), followed by idiopathic respiratory distress syndromes (IRDS), Meconium aspiration syndrome (MAS) and congenital malformations. Sixty-eight percent of mortality was contributed by low birth weight (LBW), 74% of them being preterm suggesting high mortality among LBW-preterm infants. Other less common contributory factors were maternal medical disease, complicated deliveries and multiple gestation. CONCLUSION: These causes and contributory factors of neonatal mortality reflect poor prenatal health services in this country. Hence there is a need for coordinated efforts to organise and regionalize MCH and prenatal health services with the help of a neonatal task force. PMID- 10531782 TI - Down's syndrome: prospects for prevention by antenatal diagnosis. AB - The results of a prospective study on cytogenetic analysis of Chorionic Villus Samples (CVS) taken in early pregnancy (after 10 weeks) from pregnant ladies aged between 22 and 50 years are being presented. OBJECTIVE: To find out the prevalence of chromosomal defects with advancing age of the mother. SETTING: Department of Medical Genetics, Armed Forces Institute of Pathology, Rawalpindi. METHODS: A total of 48 samples have been studied. Ten patients were above the age of 35 years and 38 were below the age of 35 years. Chorionic villus samples were obtained after 10th week of pregnancy through transabdominal approach. Cytogenetic cultures were set up both by the direct and routine methods. RESULTS: Three out of the seven samples taken from expecting mothers aged above 35 and one culture from a patient aged less than 35, showed trisomy 21. CONCLUSION: This study highlights the fact that incidence of chromosomal aberrations and the Down's syndrome in particular, increases with the advancing maternal age. Prenatal studies can therefore be utilized to decrease the incidence of various chromosomal abnormalities. PMID- 10531783 TI - Clinico-morphological pattern of intracranial tumors in children. AB - OBJECTIVE: The objective of present study was to observe the histopathological pattern of intracranial tumors in children (< 15 yrs) and to correlate the site of lesion along with the histological diagnosis. SETTING: The study included consecutive cases of intracranial tumors diagnosed in children (< 15 yrs.) in the section of histopathology at the Aga Khan University Hospital, Karachi during the period of three years. METHODS: The initial histological evaluation of these lesions was performed on H and E stained section of paraffin embedded tissue. Special stains and immunohistochemical analysis was done whenever indicated. RESULTS: During the study period, fifty-four cases of intracranial tumors were diagnosed in children. The age ranged from 1-1/2 years to 4 years with male to female ratio of 1.1:1. Astrocytoma comprised 39% of all intracranial tumors of childhood. Medulloblastoma (18.6%) ranked the second most prevalent brain tumor followed by empendymoma (13%), oligodendroglioma 7.5% while non-Hodgkin's lymphoma, primitive neuroblastoma 3.7% and ganglioglioma 3.7% while non-Hodgkin's lymphoma, primitive neuroectodermal tumors, mixed germ cell tumor, pineoblastoma, choroid plexus carcinoma and malignant meningioma constituted 1.8% each. CONCLUSION: Astrocytoma was the most common pediatric brain tumor. Medulloblastoma was more common in males while pilocytic astrocytoma was more frequent in females. Posterior cranial fossa was the most common site (43.5%) of pediatric brain tumors. Low grade astrocytoma was more prevalent in posterior cranial fossa as compared to high grade astrocytoma which was more frequent in the supratentorial region. PMID- 10531784 TI - CT and MR imaging in young stroke patients. AB - BACKGROUND: This study investigates the role of CT and MR imaging in the diagnosis and management of young stroke patients. METHODS: CT scan findings of 108 patients and MR findings of 30 patients between 15-45 years of age were reviewed retrospectively. The variables included the territory of infarct on CT and MR imaging, the cortical distribution and size of infarct. RESULTS: About 80% of the patients had infarcts of the carotid territory and 20% the vertebro basilar distribution. More than half of the infarcts were cortical (56%). The yield of MR imaging was much higher for deeper structures such as basal ganglia, thalamus and brainstem. In half the cases, the infarct size was more than 3 cm. CONCLUSION: The ratio of carotid to vertebro-basilar infarcts was similar to that reported previously. A large proportion of the carotid territory infarcts were cortical. Deeper infarcts were better imaged with MR scan. There was a high proportion of large infarcts. PMID- 10531785 TI - Bell's palsy--revisited. PMID- 10531786 TI - Persistent mullerian duct syndrome: report of two boys with associated transverse testicular ectopia. PMID- 10531787 TI - View box--case 3. Pulmonary lymphangioleiomyomatosis. PMID- 10531788 TI - Dealing with irritable bowel syndrome. PMID- 10531789 TI - Intrauterine growth restriction: a perspective for Pakistan. PMID- 10531790 TI - [Possibilities of non-hormonal drug treatment of osteoporosis]. AB - In many countries osteoporosis is the most common metabolic bone diseases. A great deal is known about the pathophysiology and the treatment of the disease. There is a lot of treatment possibilities and many new treatments are being tested. Therapeutic agents for osteoporosis are correspondingly classified as substances primarily inhibiting bone turnover (most of them are inhibitors of bone resorption as well) and agents that appear capable of restoring bone mass previously lost (stimulators of bone formation). From a didactic point of view the distinction of these to groups is generally accepted, but pharmacologically there is a considerable overlap between two. In this review the authors evaluate non-hormonal drugs, which are being used widely in the treatment of osteoporosis. The key points of the evaluation are the mechanism of action, the effects on bone mass, bone strength and fracture. PMID- 10531791 TI - [Comparative study of pancreatic head resection in chronic pancreatitis]. AB - Between 1991 and 1998 the authors performed 21 pylorus-preserving pancreatoduodenectomies (PPPD), 32 Beger and 13 Frey procedures in chronic pancreatitis with inflammatory head enlargement. The pre- and intraoperative data, as well as the postoperative early and late results were also compared. The preoperative clinical features and the intraoperative morphology were similar in the three groups. Considering the operative and late mortality and the reoperation there was no statistical difference between the procedures. The postoperative hospitalization time was the shortest after the duodenum-preserving pancreatic head resections (Beger and Frey). While the rate of early complications was significantly higher after PPPD, there was no difference in the rate of late complications between the groups. Although each operation gave similarly good late results (freedom from pain, weight gain, exocrine function, quality of life), the condition of the patients was better and the development rate of diabetes was lower (1/27), after Beger procedure, that after PPPD (6/14). Consequently the duodenum-preserving pancreatic head resections seem to be more advantageous, than the PPPD. Nevertheless the latter procedure is the only possibility in some cases. PMID- 10531792 TI - [Result of Panzytrat 25000 therapy following total gastrectomy]. AB - Thirteen patients who had undergone total gastrectomy because of gastric cancer in 11 cases and gastric lymphoma in 2 cases (6 female, 7 male) at the 3rd Department of Surgery, Semmelweis University, Medical School have been followed up. The length of follow up period varied between 7.5 months and six years. Vitamin B12 substitution was applied in each case (300 micrograms/month). In 6 cases early temporary iron substitution was necessary. Regular pancreatic enzyme substitution (pancreatin) was used during meals in these patients. At the beginning Kreon (Chinoin) or Neo-Panpur (Egis) treatment was applied. Since April 1996 the patients have been treated by Panzytrat 25,000 (Knoll) which has higher enzyme content comparing with the previous ones. Following gastrectomy the digestion and absorption improved due to pancreatic enzyme substitution and the body weight increased. The serum albumin and cholesterol levels elevated significantly, while the serum uric acid levels did not changed. The iron absorption improved, patients did not require iron substitution later, except two cases. One of them needed transitoric and the other continuous iron substitution. Side effects were recorded in six cases. One of the 13 patients stopped application of Panzytrat 25,000 because of epigastric pain among other side effects (2 epigastric pain, 2 hyperuric state, 2 frequent discharge of stool) and returned to well tolerated Neo-Panpur. PMID- 10531793 TI - [Non-invasive management of multiple brain abscesses. Case report and review of the literature]. AB - The authors report a case and treatment of multiple brain abscesses located in the cerebrum and cerebellum combined with subdural empyema. In conjunction with the case report, the authors review the literature on the pathogenesis of brain abscesses and discuss therapeutic strategies concerning the topic. In the case presented, the primary infection persisted in the lung causing subclinical bronchitis. The hemoculture showed evidence of Streptococcus mitis infection. Although the etiological role of this bacterium in meningitis is known, it rarely causes bacterial meningitis without underlying predisposing factors. In their case, the patient was free of the most common predisposing factors such as congenital heart disease or immunodeficiency. Following the 2 month period of latency, a rapid onset of the symptoms of intracranial inflammation could be observed: fever, headache, meningeal symptoms, focal neurological symptoms and coma. They were not able to identify any bacteria in the cerebrospinal fluid; the Streptocossus mitis could be cultivated only from the haemoculture. The cytological analysis of the cerebrospinal fluid showed typical signs of bacterial infection and the cranial Computed Tomography revealed multiple cerebral abscesses. Neurosurgical intervention was not recommended because of the number, localization and size of the focal lesions. The therapy consisted of intravenous administration of 24 x 10(6) IU/die Penicillin and 4 g/die ceftriaxon. For supportive therapy, Mannitol B, 3 mg/die clonazepam and 300 mg/die phenytoin were administered. Corticosteroids were not used during the course of therapy. Two years later the 55 year old female is symptom free and doing well. PMID- 10531794 TI - [In memoriam Gyula Jaki, M.D]. PMID- 10531795 TI - ["I shall create you here". Fetus and newborn infant in ancient Egypt]. PMID- 10531796 TI - [Life, death and the beyond]. PMID- 10531798 TI - Alcohol not all bad. PMID- 10531797 TI - [Hairy cell leukemia]. PMID- 10531799 TI - Physicians do counsel patients. PMID- 10531800 TI - Blue Cross and Blue Shield warning. PMID- 10531801 TI - Tired of hearing that physicians are to blame for not resisting the corporatization of medicine. PMID- 10531802 TI - MedBytes. PMID- 10531803 TI - Symposium on environmental medicine: into the next millennium. AB - Physicians and other health care providers are witnessing a growth in the number of patients who present with concerns related to environmental exposures. Environmental medicine emphasizes evaluation and prevention of exposure-related illness. Both patients and their communities view physicians as credible sources of information about environmental health matters; however, the knowledge and training that physicians have in this area are generally limited. Recognizing this shortcoming, the Institute of Medicine (IOM) has recommended that competency based training be integrated into all levels of medical education for physicians. This symposium issue includes articles written by Texas physicians from various forms of practice, and complements the IOM learning approach by serving as a resource of information. The goal is to increase knowledge and awareness of environmental issues among physicians who play a special stewardship role for our planet and the health of its human inhabitants. PMID- 10531804 TI - Environmental awareness among physicians: what are environmental health and environmental medicine? AB - Increased awareness of environmental health issues by the biomedical community and the general public places higher demands on all health and medical practitioners to understand the key terms and concepts related to environmental health and medicine. We all exist in 3 different environments (home, workplace, and community), each of which has its own array of hazards and means of exposure. Environmental medicine is a discipline that addresses preventive measures and provides assessment, diagnosis, and treatment of persons who experience adverse health effects of environmental exposures. In addition to basic clinical skills, environmental medicine uses specialized tools including biostatistics and epidemiologic studies, the science of toxicology and risk assessment, the discipline of industrial hygiene, and administrative and public policy skills. Examples of environmental issues that are faced by society today include asbestos exposure in public buildings as well as pesticide and other agricultural chemical residues present in various environmental media. PMID- 10531805 TI - Physicians' role in environmental issues. AB - Our health and the health of our patients are linked closely to the quality of our environment. As physicians, we ourselves must learn and then we must educate our patients about the importance of preventing health-endangering pollution of our air, water, food, and soil. As busy physicians, we need access to the most current, important new scientific information regarding environmental degradation where our patients work and live so we can appreciate the clinical problems that some patients present to us today. The National Association of Physicians for the Environment (NAPE) is an organization of physicians and 46 medical professional societies serving to promote protection of the environment as a fundamental responsibility of physicians and other health professionals. NAPE sponsors national conferences, promotes the publication of current "best practices" information in various general and specialty areas, and serves as a national forum for the discussion of issues and dissemination of scientifically valid information needed for physician and patient education. PMID- 10531806 TI - The role of research in environmental science and health. AB - Research plays a crucial central role between physicians and regulatory agencies in assessing the potential risks posed by an ever-increasing variety of environmental pollutants. The explosion in our understanding of biology and the development of the powerful tools of molecular biology during the last 50 years have provided us with a unique opportunity to apply this knowledge to predict the dangers of these pollutants and to act to protect the public where appropriate. To realize this vision requires multidisciplinary cooperation, continued research funding, and improved scientific literacy among the public. In this article, we do not attempt to review all the literature relating the role that research has played in environmental health and science, but rather we hope to provide the reader with a broad overview of the role that research has played in the past, emphasizing our own area of expertise (the respiratory system) and the role that research should play as our increasingly complex society moves into the 21st century. PMID- 10531807 TI - The alphabet soup of environmental regulations. AB - As we approach the beginning of the 21st century, our desire to protect and preserve our environment is stronger than ever. Many of the developed and developing countries of the world have enacted legislation to aid the protection of their people and the environment by regulating the manufacture, import, and use of industrial chemicals. In the United States, government regulations at the national, state, and local levels typically complement the legal system's common law remedies. The most significant development in modern environmental law in the second half of the 20th century has been the growth in environmental statutes that take on the anticipatory, proactive regulatory function that the common law does not. PMID- 10531808 TI - Environmental health issues along the United States-Mexico border: an airshed in evolution. AB - The El Paso area represents a unique ecosystem with a transboundary airshed shared by 3 cities, 3 states, and 2 countries. That toxic air pollutants respect no borders is exemplified clearly in this federally designated nonattainment area. However, a combination of grassroots involvement and studies under way through the scientific community has led to evolving approaches directed at improving air quality and identifying its impact on human health. The involvement of physicians and cooperative efforts among governments and communities has and will be key in this continued effort. PMID- 10531809 TI - Occupational asthma. AB - The seriousness of asthma in the general population has been recognized by increased prevalence, morbidity, and mortality rates in the past 20 years. The effects of occupational asthma on health and productivity in the workplace have been so deleterious that the Occupational Safety and Health Administration targeted 1995 as a crisis year for effective remediation. Several risk factors have been identified, but all asthma is multifactorial. Inhaled chemical, physical, and microbiological agents in the form of dust, fumes, gases, and vapors may cause workplace asthma, which is mediated through pharmacologic, immunologic, or irritant mechanisms. Because of the complexity of these mechanisms after exposure to the offending agents, the clinical manifestations may be classed as immediate, delayed, or dual responses. Evaluating causation and relationship to work requires a thorough history (including detailed job description), physical examination, and definitive studies to determine the presence of bronchospasm, bronchial hyperreactivity, atopy, work-relatedness, and presence of specific sensitization. Goals of treatment for occupational asthma are to maintain pulmonary function as close to normal as possible, to maintain a normal lifestyle, and to prevent exacerbation. In occupational asthma, particularly, the patient (or the inciting cause, if known) should be removed from the offending environment as soon as possible. Specific treatment depends upon the specific offending agent, and antiasthma therapy may be needed following the guidelines of the National Heart, Lung, and Blood Institute. PMID- 10531810 TI - The Texarkana mercury incident. AB - In November 1997, 2 teenagers allegedly removed a large amount of metallic mercury from an abandoned sign plant and distributed the material among friends. One teenager developed symptoms and admitted playing with mercury to his physician. His blood mercury was elevated. In February 1998, faculty from the University of Texas Health Center at Tyler conducted an investigation that included in-depth evaluations on 10 patients with urine mercury concentrations up to 100 micrograms/L. Exposure pathways and timelines were reconstructed from records assembled by the Arkansas State Health Department epidemiologist. Mercury contamination was found among teenagers, children, and adults who came in contact with the metal. Biomarkers of exposure documented reduction in mercury concentrations after these persons were removed from their homes and sources of mercury. Neurobehavioral assessment, including assessment of tremor, failed to establish a relationship between mercury exposure and performance. PMID- 10531811 TI - The silent voice of reason. AB - In the arena of environmental health science, we need more data, and we must better interpret the information we have already. Perhaps our greatest deficit, however, lies in the need to communicate more effectively with the public on matters of environmental health science. At the heart of better environmental health, we believe, is a critical need for scientists to become more active in communicating environmental risks. PMID- 10531812 TI - Being a midwife or doing midwifery? AB - As a profession, we need to be more conscious of the way in which our discipline has been subsumed into techno-rational science and away from our 'with women' focus. The ACMI Philosophy says that we value 'being with' women as the foundation for midwifery practice, therefore the concept 'being with' should be a competency which is reflected in the ACMI Professional Standards and Competencies. Currently, there is no Professional Standard for "Being With Women": all midwifery care has been subsumed under the rubric of problem-solving. The problem-solving approach is part of the modernist, techno-rational approach to human life experiences. Problem-solving, I am claiming, has been over generalised and imposed upon human experiences where it is not appropriate. This paper argues against adopting problem-solving as the framework for practice because this means that every aspect of midwifery care has to be problematised. This paper concludes by suggesting one way in which our philosophy could be honoured in our professional standards and competencies. PMID- 10531813 TI - Postnatal depression: a study of mothers in the metropolitan area of Perth, Western Australia. AB - This paper sets out the extent to which postnatal depression (PND) existed within the Perth metropolitan area of Western Australia in 1993/94. More importantly, the phenomenon of PND, from the mothers' perspective, is described. The paper is based on a two-stage research carried out on PND from 1993 to 1994. In the first stage, the Edinburgh Postnatal Depression Scale (EPDS) was used to screen mothers for PND. A random sample of 399 women showed 72 (18 percent) were found to be suffering from a depressive illness of varying severity. The second stage of the study used a grounded theory approach. Fifteen mothers with PND from the group screened were interviewed in their own homes. A schema, developed from the data, revealed that the main problems of the mothers was vulnerability. Support played a pivotal role in the transition from PND to stable motherhood. There was a long and short circuit followed, according to the availability and non-availability of support. PMID- 10531814 TI - Multiculturalism and the midwife. AB - This article explores the concept of multiculturalism and the relationship between the midwife and clients from non-English speaking backgrounds. There is evidence to suggest that cultural biases and stereotyping inhibits equity in care provision. To transcend these cultural barriers, there are important implications for health administrators, educators and health professionals to consider when caring for a multicultural clientele. PMID- 10531815 TI - An overview of lactation education in Australia for health professionals. PMID- 10531816 TI - "A survey of antenatal clinic staff: some perceived barriers to the promotion of smoking cessation in pregnancy". AB - Antenatal clinic staff were surveyed for their attitudes to smoking in pregnancy in 1993 and again in 1996 to monitor the effect of a randomised controlled trial of a smoking intervention conducted in the clinic over the period. Descriptive analysis showed that staff believe smoking in pregnancy is an important health risk for both mother and baby, quitting smoking is difficult, counselling is only moderately successful, they lack the skill to counsel smokers and there is little time to do so. The lack of structural support within clinic administration, the lack of a comprehensive hospital policy on smoking and unclear public health messages, were also identified as barriers to reducing the prevalence of smoking. PMID- 10531817 TI - The art of midwifery: lost to technology? AB - This paper looks at how the art of midwifery is affected by the increasing availability and use of sophisticated technology. The use of the cardiotocograph is an example of how overuse of such technology can have detrimental affects, not only for the midwife but also for the woman in labour. While this technology has made a great impact in obstetric nursing, the effects on the low-risk pregnancy need to be evaluated. Midwives need to be research-based in their clinical practice and question the overuse of technology, such as the cardiotocograph, in cases where it is not warranted. PMID- 10531818 TI - Nurses must protect patients' nutritional needs. PMID- 10531819 TI - Clinical negligence legal advice to NHS trusts. PMID- 10531820 TI - Professional misconduct case studies. Case 2: Altering medication dosages. Increase in dose of diamorphine without proper authority. PMID- 10531821 TI - Infection control. 1: A practical guide to glove usage. AB - With increased demands from the general public for healthcare professionals to be accountable for their actions, many are becoming familiar with clinical governance and other initiatives to improve clinical practice. Good infection control is central to nursing practice. To achieve higher standards of clinical practice, especially when thinking about how to reduce the risk of cross infection, it is necessary to not only do the right thing, but also do the thing right. Safe practice should be uppermost in the minds of healthcare professionals when caring for patients. This new series of articles attempts to look at the practical aspects of infection control, highlighting the requirements for risk assessment and applying the principles of infection control to a variety of patient care situations. This article investigates the use of protective clothing and gloves. It looks at the types of gloves available for use, the importance of choosing the correct glove for the task to be undertaken, and the modern day problems of allergies to latex. PMID- 10531822 TI - A practical guide to venepuncture and management of complications. AB - For years many nurses have felt that if they had the ability to perform skills such as venepuncture they would be able to provide a more holistic and efficient service. The culture in which nurses and doctors have traditionally worked has often made it difficult for nurses to become competent at such skills. However, the boundaries of medical and nursing roles have started to change and a culture of shared roles is emerging which has many benefits for patients, medical staff and nursing staff. This article provides a practical guide to venepuncture. It highlights the structure of a vein and the veins that are suitable for venepuncture. It also addresses the prevention and management of potential complications. PMID- 10531823 TI - The DIAL-log study. 2: Support in the early stages of dementia. AB - In the previous article (Vol 8(6): 387-93) the background information and study aims/limitations of the Dementia Information and Advice Lines (DIAL-log) project were outlined. This article introduces the main findings of the project and suggests that the transition into dementia is experienced through rising degrees of uncertainty and anxiety. People with the early experience of dementia also requested strategies for coping with memory loss and uncertainty over the meaning and purpose of memory testing. Building on the main findings of the project, the article concludes with a five-stage framework to help shape future research and service support. PMID- 10531824 TI - Is the sexuality of people with a learning disability being denied? AB - This article explores how the nursing profession has become more liberal in its attitudes towards sexuality as a consequence of and a response to HIV/AIDS. This liberal attitude has, however, failed to be generalized towards people with a learning disability. The continued use of labelling terms for people with a learning disability serves as an excuse to either justify the control of people's sexuality and fertility, or a rationale to argue that people with a learning disability do not have a sexual identity. Developments in the self-advocacy and advocacy movement provide opportunities for people with a learning disability to have a say in the development of sexuality policies to guide the practice and philosophy within an organization. PMID- 10531825 TI - The use of music and colour theory as a behaviour modifier. AB - For many centuries various aspects of healing have been linked to the use of the arts, in particular music and colour because of their innate ability to bring about a mental, emotional and physical calmness. Although much has been written on the use of colour and music as relaxants specifically within a nursing/medical context, there appears to be little information available as to why music and colour have this calming effect. This article examines music and colour as relaxants by briefly describing the neurological and physical mechanisms that bring about the effect of relaxation. This brief exploration is placed within the context of learning disability care. The aim is to provide ideas for a more peaceful and relaxing environment for an adult with learning disabilities who also has autism and exhibits severe challenging behaviour. The results of a small case study and implications for other areas of nursing are discussed. PMID- 10531826 TI - Clinical governance: the new NHS, new responsibilities? AB - The declared aim of the White Paper 'The New NHS: Modern, Dependable' is to restructure the NHS so that it is based on 'partnership and driven by performance' (Department of Health (DoH), 1997). The key organizing principles are those of efficiency and excellence. Establishing a framework for clinical governance is seen as central to establishing 'an environment in which excellence in clinical care can flourish' (DoH, 1998). These new arrangements will apply to the the practice of all clinical professionals, including nurses. Clinical governance offers a new opportunity for the nursing profession to raise standards of practice without being hamstrung by issues of patient throughput and price, as was the case under the last government. However, it also presents a challenge to the frameworks within which nursing has traditionally operated. In addition, clinical governance brings with it the potential for more direct involvement of non-clinicians in professional practice. PMID- 10531827 TI - Pressure ulcer prevention and treatment: the Transair range. AB - The Transair range has been updated as a result of clinical and technical advances. The Transair 500 cushion system utilizes foam and air technology to minimize disturbance to the user. Clinical trials suggest that this cushion is of benefit in both the prevention and treatment of pressure ulcers. The Transair 1001 alternating pressure air overlay and the Transair 2002 mattress replacement have both been revised to provide improved performance and both have been subjected to clinical trials. The results of these trials suggest that both have a role in the prevention and treatment of pressure sores in the individual in the high/very high risk group. This article examines these innovations in the Transair mattress and seating systems. PMID- 10531828 TI - The Sims Portex Catheter Valve: an alternative to the leg bag. AB - Any patients who would normally use a catheter and leg bag could benefit from exchanging their leg bag for a catheter valve, as long as they have the manual dexterity to open their valve at regular intervals and remember to do so. The Sims Portex Catheter Valve is a lightweight and compact catheter valve that is discrete and offers more comfort than a leg bag system, especially to the more mobile patient. This article examines this type of catheter valve and assesses its relevance for use in both the community and the acute sector. PMID- 10531829 TI - The nature of health promotion in acute and community settings. AB - Since the late 1980s, there has been a consistent call from nursing's advisory and legislative bodies to incorporate the discipline of health promotion into the nursing profession. On reflection, however, evidence indicates that despite these calls there has not been a universal uptake of health promotional activity into the profession. What is evident is that where health promotional activity does take place, it occurs more in the community than in the acute setting. This article sets out to explore the reasons for this. It suggests that certain current and future activities may help to further promote the universal adoption of health promotion within nursing practice. PMID- 10531830 TI - Clinical governance and self-regulation in nursing. PMID- 10531831 TI - How much proof do we need? PMID- 10531832 TI - Care efficient strategies? PMID- 10531833 TI - An experience in telenursing. AB - The purpose of this article is to describe how a telemedicine system was used to complete a family and home assessment and to discuss issues facing advanced practice nurses (APNs) when they use such systems in practice. Incorporating discharge care into advanced practice is an increasingly important component of the nursing care given during acute illness. Telemedicine systems offer a mechanism for assessing the ways in which home situations impact on patient recovery. Telemedicine provides a method for early intervention that can ameliorate or prevent developing problems relatively inexpensively. The advantages and disadvantages of one such system, the Picasso, are described. PMID- 10531834 TI - Implementation of a patient-family pathway: effects on patients and families. AB - The purpose of this study was to discover whether implementation of a patient family pathway with patients and families undergoing coronary artery bypass graft (CABG) surgery impacted anxiety, information with care planning, and patient length of stay. Using an experimental design, a sample of 60 patients and family members was studied. Each patient and his or her designated family member received either the patient-family pathway or the hospital's standard care planning. Findings indicated no statistically significant differences in state anxiety or information with care planning between patients and family members receiving the patient-family pathway and those receiving standard care planning. There was no statistically significant difference in length of stay between the two patient groups. The results indicate that the CABG patient-family pathway has limited value to patients and families as measured in this study. Resources can be real-located to other uses that may have a more positive impact on the patient and family experience. PMID- 10531835 TI - Casting a cold eye on reality: Orwell's The road to Wigan Pier. PMID- 10531836 TI - Documenting the effectiveness of nursing practice PMID- 10531837 TI - CNS facilitation of a cardiac surgery clinical pathway program. AB - In this collaborative project, the Clinical Nurse Specialist (CNS) worked with various members of the healthcare team using a clinical pathway group work process to implement changes in the nursing, medical, and respiratory care of cardiac surgery patients. The patient population (N = 598) comprised cardiac surgery patients undergoing coronary artery bypass graft, mitral valve replacement, or aortic valve replacement. The practice changes implemented were earlier extubation, earlier ambulation, the administration of fentanyl and propofol, and the administration of gastrointestinal (GI) prophylactic medications. The overall outcomes were decreased incidence of pneumonia, earlier increase in level of consciousness, improved ambulation abilities, and improved nausea levels. Pneumonia decreased significantly, from 2.49% to 1.67% (p = 0.05). For patients who met early extubation criteria, mean time on the ventilator decreased from 17 hours to 8 hours, and length of stay decreased from 8 days to 7 days in a subgroup of patients (diagnosis-related group (DRG) 105). The overall annual charge savings was approximately $201,000. These results add to the belief that CNS-guided patient care in collaboration with the healthcare team has positive benefits. PMID- 10531838 TI - Enhancing psychiatric nursing practice: role of an advanced practice nurse. AB - In response to demands for alternative health models that deliver cost-effective quality care, one large Midwestern medical center implemented a change in their nursing practice model. The change involved the introduction of unit-based nursing leadership teams that included advanced practice nurses (APNs). This article reports the findings from an investigation that employed a case study design to evaluate the process and outcomes of integrating an APN on a psychiatric unit with experienced nurses. Data collection methods included a nursing survey (n = 34), interviews with nine randomly selected nurses, and two multidisciplinary open forums. Consistent with the staff nurses' survey ratings of important advanced practice role functions, responses from the nursing interviews and open forums suggested nurses' professional development to be the most positive outcome. Role confusion was identified as a negative outcome. Recommendations for improved integration and use of APNs in today's psychiatric health environments were identified. PMID- 10531839 TI - Update on clinical nurse specialist recognition and prescriptive authority. PMID- 10531840 TI - Who prepares the nurse for preventing codes? PMID- 10531842 TI - The importance of evidence-based practice. PMID- 10531841 TI - Inhospital cardiac arrest: pre-event variables and nursing response. AB - This retrospective study examined the medical records of 100 patients who experienced an in-hospital cardiopulmonary arrest. The purposes of this study were to identify pre-arrest physiologic changes that may have occurred in the patient and to determine whether physician notification time, physiologic variables, patient location, and the presence of an electrocardiogram (ECG) monitor before the arrest affected the resuscitation outcome. The results showed that assessment variances were present in most patients before the arrest and also were recognized by the nursing staff. Implications for practice include formation of quality improvement screening tools to assess the patient's pre arrest status, development of competency tests that include scenarios involving changes in a patient's physiologic parameters, staff education, and evaluation of current nursing policies for obtaining vital signs and assessments. PMID- 10531843 TI - Architects of the diabetes team. PMID- 10531844 TI - Communicating through stories: experience of the Native American Diabetes Project. AB - Stories appear to provide an indirect way of confronting the inherent conflict between the concepts of disease and wellness and assisting in the transition to a new concept of living well with the disease. This new concept may engender feelings of acceptance and hope that can facilitate application of knowledge and behavior change. In addition, culturally appropriate stories allow people to draw from their own personal beliefs and values to interpret and apply new information to their own lives. A good story takes listeners on a collective journey with many paths; each path is uniquely suited to the needs of the individual, with wisdom gained that is uniquely suited to their own life. PMID- 10531845 TI - Developing a computerized diabetes self-management education module for documenting outcomes. AB - PURPOSE: This paper describes a process used to develop a computerized diabetes self-management education record that complies with the National Standards for Diabetes Self-Management Education Programs. METHODS: A working prototype was developed to computerize 1 of the 15 content modules outlined in the National Standards for use on the World Wide Web. During program development, three consultants reviewed the content and proposed structure. For the subsequent prototype, five diabetes educators served as users and evaluated the content, design, and flow of the system. RESULTS: The module was found to be thorough in terms of curriculum content and proposed structure for subsequent teaching. Overall, users were satisfied with the graphic interface screens and provided important feedback for determining specific modifications for future development. CONCLUSIONS: The World Wide Web format provides a universal platform for documenting diabetes education outcomes and allows a broad range of access and networking capabilities. PMID- 10531846 TI - Involvement in health behaviors among youth with diabetes. AB - PURPOSE: The purpose of this study was to examine involvement in a broad range of health behaviors among adolescents and young adults with diabetes. METHODS: The sample consisted of 107 adolescents and young adults (12 to 24 years old) with Type 1 diabetes mellitus. Participants were asked to report involvement in health enhancing, health-compromising, and diabetes mismanagement behaviors. RESULTS: The participants reported low levels of health-compromising behaviors and high levels of health-enhancing behaviors. Females reported significantly higher levels of diabetes mismanagement than males. Males in late adolescence (18 to 24 years) reported significantly higher levels of health-compromising behaviors than males in early (12 to 14 years) and middle (15 to 17 years) adolescence. Females in late adolescence (18 to 24 years) reported significantly higher levels of health-compromising behaviors than females in early adolescence (12 to 14 years). CONCLUSIONS: Diabetes educators who work with youth may want to assess all of these health behaviors, keeping in mind age and gender differences. PMID- 10531847 TI - Culture counts: why current treatment models fail Hispanic women with type 2 diabetes. AB - PURPOSE: The purpose of this article is to evaluate whether current treatment models adequately address the cultural factors involved in treatment adherence in Hispanic females with Type 2 diabetes. METHODS: A review of relevant professional literature was conducted. RESULTS: Established health behavior models do not adequately address the unique needs of the female Hispanic population, especially those older women who hold traditional religious and cultural beliefs. CONCLUSIONS: To decrease the devastating effects of Type 2 diabetes among Hispanic women, interventions must be based on a comprehensive, culturally sensitive model that works with cultural values, not against them. PMID- 10531849 TI - Lispro insulin: adsorption and stability in selected intravenous devices. AB - PURPOSE: The adsorption characteristics and stability profile of an insulin analog, lispro insulin, were evaluated against a recombinant human regular insulin using intravenous infusion sets and syringes. METHODS: Studies were performed using either 0.9% NaCl or 5% dextrose intravenous injection solution. Effects of container type, infusion rate, product concentration, presence-absence of an in-line filter, and storage condition on release profiles of lispro and human regular insulin infusion solutions were determined. RESULTS: Lispro insulin and m-cresol were chemically stable. Release rates of insulin (both types) were steady after an initial lag time. The lag time was much longer with intravenous bag infusion than with intravenous syringe infusion. A higher product concentration, faster flow rate, and prewash of the infusion tubing were shown to substantially decrease the lag time. CONCLUSIONS: The adsorption profile of lispro insulin was the same as that of human regular insulin in both syringes and bags. Use of a load-and-sit prewash scheme may shorten or nearly eliminate the lag time, which in turn may be used to make a more accurate calculation of a patient's dose. PMID- 10531848 TI - Culturally competent diabetes education for Mexican Americans: the Starr County Study. AB - PURPOSE: Few culturally competent health programs have been designed for Mexican Americans, a group that bears a disproportionate burden of Type 2 diabetes. In Starr County, a Texas-Mexico border community, investigators designed and tested a culturally competent intervention aimed at improving the health of this target population. The purpose of this article is to describe the development process of this diabetes education and support group intervention. METHODS: The development stages were (1) community assessment, (2) intervention design, (3) selection or development of outcomes, (4) pilot testing, and (5) a randomized clinical investigation. RESULTS: Focus group participants identified knowledge deficits regarding diabetes and self-management strategies, and suggested characteristics of an effective intervention for Mexican Americans. Outcome measures included metabolic control indicators, a newly developed knowledge instrument, and an existing health belief instrument. Preliminary analyses indicated that the intervention was successful in significantly improving metabolic control in the target population. CONCLUSIONS: Developing successful diabetes interventions for minority groups requires a number of stages, careful planning, assessment of cultural characteristics of the target population, and a systematic approach to implementation. PMID- 10531850 TI - Using solution-focused therapy strategies in empowerment-based education. PMID- 10531851 TI - The heart of a multidisciplinary approach. PMID- 10531852 TI - Outcomes-driven diabetes education. PMID- 10531853 TI - A search for answers about foods with polyols (sugar alcohols). PMID- 10531854 TI - Depression in diabetes mellitus: a national survey of office-based encounters, 1990-1995. PMID- 10531855 TI - Participant satisfaction with a culturally appropriate diabetes education program: the Native American Diabetes Project. AB - PURPOSE: The purpose of this paper is to report on participant satisfaction with the Native American Diabetes Project diabetes education program. METHODS: A questionnaire was designed to measure satisfaction among participants in the diabetes education program, which consisted of five sessions designed according to the Transtheoretical Model of Change and Social Action Theory with input from community members. Eight pueblo communities participated in the program. Sessions were taught by community mentors in three sites in New Mexico. One site taught sessions in a one-on-one format, and two sites taught sessions in a group format. RESULTS: The results showed that participant satisfaction did not vary based on session delivery type or by session site. Overall, participants responded positively to sessions designed according to Social Action Theory and with cultural competency. Retention rates for the sessions were 81% for group sessions and 91% for one-one-one sessions. CONCLUSIONS: Using a strong theoretical framework and community input to design diabetes education sessions may be important factors in participant satisfaction and retention in diabetes lifestyle education sessions. PMID- 10531856 TI - The Diabetes Day Treatment experiment: a preliminary report on what we learned. AB - PURPOSE: This paper presents data on the efficacy of a diabetes day treatment program to modify the healthcare behavior of elderly African Americans with diabetes. METHODS: African American patients with Type 1 or Type 2 diabetes who were referred by their certified diabetes educator were eligible to participate in the day treatment program. The program was designed to serve eight patients for 4 hours 1 day a week over 9 months. Participants engaged in informal discussions, low-impact armchair exercises, and discussions of various diabetes issues. A flow sheet was initiated and maintained by the investigators to record information pertaining to each participant's blood pressure, blood sugar, and weight at each session. Attendance and reasons for not attending sessions were recorded. To obtain more in-depth information, the group leaders used a technique known as participant observation. RESULTS: Having CDEs administer a blood sugar test, take blood pressure, and weigh each patient at each clinic visit promotes patient adherence to the diabetes treatment regimen. Memory loss was observed to be especially prevalent among the subjects. CONCLUSIONS: The Diabetes Day Treatment Program may be used as a model for working with elderly persons with diabetes from different ethnic groups. PMID- 10531857 TI - Diabetes knowledge and its determinants in a Mexican population. AB - PURPOSE: The purpose of this study was to measure the level of diabetes knowledge in a representative group of Mexican individuals with diabetes and to identify the factors that influence it. METHODS: A validated questionnaire was administered to 570 outpatients; 11.2% had Type 1 diabetes, 36.4% had Type 2 diabetes treated with insulin, and 52.2% had Type 2 diabetes treated with oral agents. Samples for HbA1c determination also were obtained. RESULTS: The percentage of correct answers in each section of the questionnaire was low. Type 1 patients had the highest scores, followed by the insulin-treated Type 2 patients; those with chronic complications also had high scores. Educational background, attendance at diabetes courses, age, and HbA1c concentration were the main predictors of knowledge. Attendance at courses was influenced by the severity of the disease. CONCLUSIONS: The amount of patient knowledge about diabetes-related issues was low in this representative Mexican population. The educational efforts were focused on those with the worst metabolic control and/or with diabetes complications. PMID- 10531858 TI - Body size and body shape: perceptions of black women with diabetes. AB - PURPOSE: This qualitative study was conducted to explore perceptions of body size and shape in a group of black women with Type 2 diabetes. METHODS: Thirty-three black women with Type 2 diabetes participated in one of three focus groups to discuss perceptions about body size and body shape. Transcriptions of the discussion were analyzed for themes of participants' perceptions about their bodies, their ideas about body size and body shape, and personal and environmental influences on their preferences about size and shape. RESULTS: Participants preferred a middle-to-small body size but indicated that a middle-to large body size was healthier. They also said that a large body size did result in some untoward social consequences. Participants preferred a pear-shaped body (a figure without abdominal adiposity). The three major influences on body image perceptions were children, parents, and the media. CONCLUSIONS: With these findings in mind, diabetes education programs that are geared for black women may benefit from the inclusion of key family members. Additionally, the importance of body image perceptions should be recognized in the design and implementation of weight-related diabetes education programs. PMID- 10531860 TI - Malnutrition in nursing facilities: the responsibility of every health care provider. PMID- 10531859 TI - Clinical implications of amylin and amylin deficiency. AB - PURPOSE: This paper presents an overview of the physiology of glycemic control and the mechanisms of amylin deficiency in people with diabetes. Benefits of replacement therapy with both pramlintide and insulin are discussed. METHODS: The discovery of the pancreatic beta-cell hormone amylin, which is cosecreted with insulin in response to hyperglycemia, has prompted a reanalysis of the mechanisms underlying the control of glucose homeostasis. A review of the current literature on amylin and amylin deficiency provides the basis of this reanalysis, with a discussion of the clinical implications for people with diabetes. RESULTS: Amylin appears to work with insulin to regulate plasma glucose concentrations in the bloodstream, suppressing the postprandial secretion of glucagon and restraining the rate of gastric emptying. People with diabetes have a deficiency in the secretion of amylin that parallels the deficiency in insulin secretion, resulting in an excessive inflow of glucose into the bloodstream during the postprandial period. CONCLUSIONS: While insulin replacement therapy is a cornerstone of diabetes treatment, replacement of the function of both amylin and insulin may allow a more complete restoration of the normal physiology of glucose control. PMID- 10531861 TI - "They just stood there and did nothing!". PMID- 10531862 TI - Resident autonomy as impacted by the internal environment of an organization. PMID- 10531864 TI - The use of plush animals in long-term care facilities. PMID- 10531863 TI - Nurses: take time to speak up on SNF PPS proposed rules.... PMID- 10531865 TI - The mind-body connection. PMID- 10531866 TI - LTC nurses ... are we losing more opportunities. PMID- 10531867 TI - Martha M. Mohler, RN, MN, MHSA. Senior policy analyst for the National Committee to Preserve Social Security and Medicare. Interview by Pamela Squires. PMID- 10531868 TI - Long-term care resident case analysis: ethical issues. PMID- 10531869 TI - Therapeutic touch. PMID- 10531870 TI - A profile of nursing facility residents: vulnerable to malnutrition and nutrition related health problems. PMID- 10531871 TI - Evolution and development of a renal recipient transplant co-ordinator for the Asian community. AB - The aim of this paper is to share with colleagues the development of this innovative role and the unprecedented experience in addressing issues of transplantation for a large Asian population. The author is the named transplant co-ordinator for all Asian patients who account for 30% of the total waiting list. During the first six months of the post the author has undertaken home visits, nurse-led clinics and liaison with dialysis units. This continuous direct rapport is unique, and facilitates the living donor programme. The author has been involved in developing specific educational material relevant and appropriate to this patient population. Patients and families comments and responses have been favourable and positive. Audit and evaluation of the role is on-going and the author feels privileged to be central to this process of providing holistic care equitably to all patients in a multicultural society. PMID- 10531872 TI - Residual renal function and post dialysis urea rebound. AB - Kt/v is used to estimate haemodialysis prescription and treatment adequacy and usually does not include evaluation of urea rebound. In addition urea clearance provided by the presence of residual renal function (RRF) is additive to dialytic urea clearance. Rebound is a phenomenon involving other molecules such as creatinine and phosphorus. The aim of this study was to investigate the role of RRF on post dialysis urea, creatinine and phosphorus rebounds. We investigated 7 patients with RRF (group 1) and 7 uraemic anuric patients (group 2). Urea rebound was lower in group 1 than group 2: 9.5 +/- 4% vs 18 +/- 7% (p = 0.04). Creatinine rebound and phosphorus rebound were similar in both groups 15.2 +/- 9% group 1 vs 14.6 +/- 5% group 2 (p = NS) and 16.7 +/- 7% group 1 vs 20 +/- 7% group 2 (p = NS) respectively. Our data suggest Kt/v calculated without considering rebound overestimates haemodialysis efficacy. PMID- 10531873 TI - Blood pressure measurement in haemodialysis patients. AB - Several studies suggest that the 24 hour ambulatory blood pressure monitoring (ABPM) predicts left ventricular hypertrophy more accurately than conventional blood pressure measurement (CBPM) with mercury sphygmomanometer. We estimated the left ventricular mass by M-mode echocardiography in 58 patients on regular haemodialysis treatment during the midweek haemodialysis (HD) interval. ABPM was recorded during the 24 hours preceding the dialysis session and the average of values were compared with the average of the 13 pre HD CBPM recorded by nurses during the month preceding the echocardiography study. The two types of BP measurements correlated significantly with each other, (systolic BP r = 0.62; p < 0.001 and diastolic BP r = 0.74; p < 0.001). The correlation of left ventricular mass with pre-HD systolic BP was stronger (r = 0.54; p < 0.001) than with 24h systolic BP (r = 0.33; p < 0.01). The overall accuracy of prediction was also similar (68% for pre HD-CBPM; 67% for 24h-ABPM). Measurements of diastolic BP did not correlate significantly with LVM. Our data suggest that 24h-ABPM does not offer any advantage over pre HD-CBPM in predicting left ventricular hypertrophy in HD patients. PMID- 10531874 TI - Exercising to fitness on dialysis. AB - Exercise tolerance is a recognised consequence of chronic renal failure. Physical de-conditioning is an integral factor in preventing the achievement of an otherwise very acceptable quality of life which can be accomplished by good dialysis therapy. This paper reports on the implementation of an exercise programme in an ESRD setting with a patient population of over 1,100. Patients aged between 20 and 65 years were identified for phase one of the programme. All patients had an in-depth medical assessment to eliminate potential risks. The programme encompasses stretching, strengthening and aerobic exercises and a individualised exercise prescription was compiled for each participant. Twelve months after the initiation of this programme there is evidence of increased exercise tolerance, increased feelings of well-being and enhanced stability on dialysis. PMID- 10531876 TI - The influence of hygienic practices to the exit site/tunnel on peritoneal catheter infections. AB - In Peritoneal Dialysis (PD), correct handling of the equipment and especially of the catheter exit site are essential. The PD nurse is responsible for preparing the patient to be able to carry out self care. As there are no universally accepted procedures for catheter care, we evaluated the procedure recommended in our unit, to identify the level of achievement by our patients and compare it with the infection incidences of exit site/tunnel (ES/T). 31 patients were surveyed on aspects related with hygiene, care and support of the catheter. This was complemented with a direct observation of the way of fixing and protecting the catheter and exit site, relating these facts with the incidence of infection. The results show the incidence of the infection is similar to data from other centres. It seems that the non compliance with the recommendations is due to either a misunderstanding on the patients' part or difficulty to adapt these recommendations to their particular way of life. PMID- 10531875 TI - Intraperitoneal hydrostatic pressure and volume in peritoneal dialysis patients. AB - Dialysate Intraperitoneal Volume (IPV) represents one of the major determinants of Peritoneal Dialysis (PD) efficiency, but most adult patients are currently treated with the same standard IPV, regardless of body size. In order to evaluate the tolerability of different IPV, we adapted the current connection in use at our Institution to produce a simple method to directly measure Intraperitoneal Hydrostatic Pressure (IPP). We studied the relationship between IPV and IPP in 30 adult (age 19-77 years) patients (19 males) of various body sizes, on PD between 17 +/- 17 months. Mean end-inspiratory and end-expiratory IPP with different IPV were measured in each patient in the IPV range of clinical interest. A total of 210 individual measurements showed a statistically significant positive relationship between BSA-normalised IPV and IPP (r = 0.355, p < 0.001). Interpatient variability was high, thus suggesting that individualization of IPV according to body size is not accurate, IPP being often higher in larger body size. Direct IPP measurement with different IPV in the single patient is a simple, safe and reproducible procedure, allowing an individually tailored IPV prescription which should optimise PD efficiency while monitoring for IPP related complications. PMID- 10531877 TI - Peritoneal membrane transport for low molecular weight substances with the use of one bag dialysate collection. AB - The 24-hour collection of dialysate provides an accurate method for evaluation of both adequacy of dialysis and peritoneal membrane transport characteristics in patients on chronic ambulatory peritoneal dialysis. However, this test requires 24 hours to complete and therefore it is inconvenient for both patients and nurses in the every day practice. We determined the peritoneal membrane transport characteristics for low molecular weight substances of ten patients by using the dialysate collection of only one bag. Dialysate/plasma creatinine ratios were calculated for each of the 4 bags (DATT1, DATT2, DATT3, DATT4) as well as for the 24 hour dialysate (DATTo). We found a very good correlation between DATTo and the four DATTs. We therefore propose that the evaluation of the peritoneal membrane transport, at least for creatinine could be determined with the use of one bag dialysate collection. PMID- 10531878 TI - Communication as a teamwork tool in peritoneal dialysis. AB - The growing complexity of the problems which have to be faced in providing a peritoneal dialysis service, such as giving assistance over a territory, consideration of the needs of the individual and the greater emphasis on the prevention of complications, makes multidisciplinary teamwork necessary, with the interaction of various skills. In order to succeed, teamwork requires the identification of clear objectives which are shared by all the members of the team. The various skills can best be exploited by defining roles and agreeing common aims and objectives so that the team can provide the patient with holistic treatment, which recognises the needs of the individual rather than the needs of the illness. Active cohesion between the various components is only possible through a defined communication strategy and an on-going transfer of knowledge. PMID- 10531879 TI - Pre-dialysis education. A change in clinical practice. How effective is it? AB - The last decade has seen a great expansion in the provision of pre-dialysis education within the UK, with our programme in operation since 1996. In order to evaluate its impact on patient care, two methodologies were employed: the first used a satisfaction survey with which to examine patient perception of the programme; the second approach examined resource utilisation, in particular length of stay, as an outcome measure. Difficulties have arisen in discerning the impact of the programme amongst the other education interventions to which the patient is exposed. Comparative length of stay data was used as an alternative means of assessment. The results of the audit led us to question the effectiveness of the methodology that we employed and has led us to propose the development of a standard. It is hoped that the adoption of a more structured approach will not only aid the effectiveness of the programme itself but also facilitate the evaluation process. PMID- 10531880 TI - Focus charting in renal nursing: a quality issue. AB - The paper explores how to minimise charting time, yet ensure that relevant information is documented in a concise manner in locations that are easy to access. This type of charting lends itself to review and data collection which is useful for quality improvement incentives. Patient centred documentation leads to quality improvements which ultimately results on cost savings. PMID- 10531881 TI - Basic tools to integration in management and continuous quality improvement: protocols and procedures. AB - In order to support interested colleagues with the concepts of Continuous Quality Improvement (CQI) and Clinical Standards for Nephrology Practice, a package of education support has been set up. We enhance the use of procedures, protocols, working plans, guidelines and nursing care plans as instruments of organisational integration. For the nursing care of renal patients in HD and PD we believe that all the procedures and protocols (P&P) above could have an important impact in the renal nursing practice. We propose that group members run a programme which is presented and implemented as a modular package including, theoretical knowledge, setting up working groups involved in literature revision, evaluation of defined P&P after a period of implementation, final implementation & periodic evaluation. PMID- 10531882 TI - Do anaemia co-ordinators have to be nurses? AB - The role of the anaemia co-ordinator has developed subsequent to the introduction of erythropoietin therapy for renal anaemia, and posts have been established at an increasing number of hospitals in the UK. While co-ordinators have previously tended to come from a nursing background, the post at our hospital has been held jointly by a pharmacist and a clinical nurse specialist since July 1997. This paper presents an informal evaluation of our experience of joint working, and has drawn on diary entries to outline the components of the service provided. The main focus is on the boundary negotiated between pharmacist and nurse responsibilities and expertise, involving as it does areas of potential conflict and complementarity. Through a critical examination of the assumptions and expectations associated with 'generic' pharmacist and nurse roles, we begin to clarify the respective contributions which the disciplines make to anaemia management. PMID- 10531883 TI - Nursing implications of a vancomycin resistant enterococcus outbreak in a renal unit. AB - Vancomycin Resistant Enterococcus is a difficult organism to treat and is becoming more prevalent world-wide. When it was identified in a busy renal unit, radical and expensive measures were taken to try and control the outbreak. Although these had costs to both patients and staff, the organism was eradicated from the unit. PMID- 10531884 TI - Caring for renal patients during loss and bereavement. AB - Many of us are not prepared in assisting patients through the final phase of the life cycle. Recognising physical manifestations of stress, understanding why patients experience loss with kidney disease, and realising the supportive role that staff play for a patient undergoing this process, are key to assisting the patient to cope with loss and bereavement. The renal caregiver can assist the patient to view the ESRD life from another perspective as they journey through the life cycle to death. In so doing, the renal caregiver can recognise the needs of the family during this period, thus making them feel they fulfil a valuable role in the patient's final care. Being with a patient and family at the time of death is extremely rewarding as it helps the staff bring closure to the nurse patient relationship. PMID- 10531885 TI - A fable of paradox. PMID- 10531886 TI - Helping adolescents who have disabilities negotiate transitions to adulthood. AB - Adolescents who have disabilities face unique challenges as they progress through the transitions necessary to achieve optimum functioning in adulthood. These youths often need professional assistance to successfully negotiate these important transitions. Our article describes processes for collaborating with these adolescents, their families, and other professionals to facilitate successful transitions to a more healthy, productive, and satisfying adulthood. PMID- 10531887 TI - Renegotiating HIV/AIDS prevention for adolescents. AB - According to the Centers for Disease Control in 1997, approximately 25% of the estimated new human immunodeficiency virus (HIV) infections in the United States may occur in youths under 20 years. Based on the research regarding adolescents and HIV disease, it is clear that several issues need to be addressed. This article reviews adolescent HIV risks, behaviors, and knowledge and discusses the implications for prevention. PMID- 10531888 TI - The lived experience of childhood cancer: one sibling's perspective. AB - The demands of cancer on children and their parents have been studied and understood for many years now. However, very little focus has been placed on one other very important part of the family system--the siblings. In the health care profession today, there is a growing awareness that the psychosocial needs of siblings of children with cancer are less adequately met than those of other family members. Research suggests that siblings are particularly vulnerable to adjustment difficulties (depression, anger, anxiety, feelings of guilt, and social isolation), and they experience similar stress to that of the ill child Siblings have been identified as the most emotionally neglected and unhappy of all family members during serious childhood illnesses. The purpose of this study was to gain a better understanding of the lived experience of one 14-year-old sibling's experience with childhood cancer. Through the qualitative research process of phenomenology, the researcher gained a greater understanding of the participant's experience and how the childhood cancer experiences affected her and her family. Themes that emerged through the process of content analysis included emotional intensity, increased empathy for others, personal growth, need for support, and desire to help others. PMID- 10531889 TI - A study of self-esteem among well adolescents: seeking a new direction. AB - Over the past decade, nursing has identified the significance of self-esteem in maintaining wellness among adolescents. Low self-esteem has been linked to numerous adolescent risk behaviors such as smoking, drug use, and sexual activity. Adolescents engaging in these risk behaviors may have subsequent health problems, such as alcohol and drug addiction, as well as teen pregnancy. Present treatment modalities for low self-esteem have not been optimally effective. Nursing needs to examine adolescent self-esteem within the discipline of nursing and develop its own prevention and intervention strategies. Guided by the Roy Adaptation Model, our study used a descriptive, correlational design and examined the self-report of self-esteem on age group, gender, exercise participation, smoking, parental alcohol usage, depression, and anger in a nonclinical, community sample of adolescents aged 12-19. PMID- 10531890 TI - Children's responses to immunizations: lullabies as a distraction. AB - The purpose of this study was to investigate the effects of audiotaped lullabies on physiological and behavioral distress and perceived pain among children during routine immunization. An experimental design was used to study 99 healthy children ages 3 to 6 years. Half the children received the musical intervention during the immunizations, while the other half did not. Groups were assessed during five phases: baseline, preimmunizations, during the immunization, after Band-Aid application, and 2 min after phase 4. Physiological variables (heart rate, blood pressure) were obtained in phases 1, 4, and 5. Behavioral distress was measured using the Observational Scale of Behavioral Distress during phases 1, 2, 3, and 4. Pain perception was measured using the Oucher in phases 1 and 4. No significant differences were found between experimental and control groups for heart rate, blood pressure, or Oucher scores. However, total distress scores were significantly less for the experimental group. These results indicate that immunization is a stressful experience for children. Recommendations include further study incorporating pharmacological and nonpharmacological interventions. PMID- 10531891 TI - Crisis child care: an intervention for at-risk families. AB - Crisis child care programs provide parents in crisis with a break from the stresses of childrearing and provide at-risk children with a safe environment. This study describes parenting stress as the construct of measurement in a pilot project in a predominantly rural midwestern state. The Parenting Stress Index/Short Form (PSI/SF) was used to measure the amount of stress experienced by the parent as a result of the parenting role. Comparisons of the mothers' PSI/SF pretest and posttest scores indicated significant improvement in the construct areas of Total Stress. Parental Distress. Difficult Child, and Life Stress. Comparison of child maltreatment rates indicated that there was a significant decrease (X2 = 16.91, p < 0.0001) in the reported incidence of child maltreatment in the rural counties with a crisis child care program compared with counties that did not offer this intervention. Overall, these findings indicate that crisis child care is an effective preventive intervention for at-risk families. The investigator suggests that nurses enhance their collaborative relationships with these programs to provide a more seamless avenue for both receiving and providing referrals. PMID- 10531892 TI - Self-esteem in adolescents treated in an outpatient mental health setting. AB - Although self-esteem is an important concept, nursing has only begun to focus on the significance of self-esteem as a mechanism for achieving wellness among adolescents, and as a variable for targeted intervention. Nursing studies identifying self-esteem as the primary focus of their research in an adolescent population seeking treatment in mental health settings are scarce. The Roy Adaptation Model's Theory of a Person as an Adaptive System was used to guide this descriptive, correlational study. Research examined the self-report of self esteem on age, gender, smoking, exercise, depression, anger, and parental alcohol use in a sample of adolescents ages 12-19 years who were being treated in an outpatient mental health setting. PMID- 10531893 TI - Sensitive topics and adolescents: making research about risk behaviors happen. AB - This article discusses conducting research with adolescents as a positive experience, both from a clinical and scholarly perspective. However, topics involving risk-taking behaviors may be especially difficult for adolescents to discuss openly. To implement research protocols with the adolescent population, particularly when dealing with risk-taking behaviors, investigators need to be aware of developmental challenges that warrant specific methodological choices. In a pilot study that involved adolescents who had been hospitalized for traumatic injury, the researchers gained valuable experience in conducting a study on substance use. Experiences with the study provide direction for future research about investigating sensitive topics with adolescents. PMID- 10531894 TI - Conflict management: assessing educational needs. AB - In today's healthcare environment, the potential for conflict among healthcare providers exists as changes are occurring at a supersonic pace. The outcomes of conflict may affect patient care and are directly related to the effectiveness of the resolution. Clinical educators and staff development educators are essential in resolution if they assess strategies that are currently being used in managing conflict and then offer more effective resolution strategies. PMID- 10531895 TI - Role transition from graduate to staff nurse: a qualitative analysis. AB - The purpose of this study was to describe the initial steps in the role transition of graduate to staff nurse. During the first 3 weeks of an orientation to a clinical unit in an acute care hospital, graduate nurses and their preceptors used feedback sheets to document the learning activities of the graduate nurse, communicate the need for and evaluation of learning experiences, and plan activities to meet the continued needs of graduate nurses. Daily feedback sheets from 27 orientees and preceptors were analyzed using content analysis. A model representing the process and components of role transition was developed. The model was based on five themes which emerged from the data: Real Nurse Work, Guidance, Transitional Processes, Institutional Context, and Interpersonal Dynamics. Analysis of results revealed that the initial transition of a graduate nurse to the role of a staff nurse was a dynamic and interactive process occurring between the graduate nurse and the preceptor. Guided learning led to progress in balancing increasingly complex care within a specific institution. Interpersonal dynamics among staff, preceptors, and graduate nurses affected the process of role transition. PMID- 10531896 TI - A mock trial at nursing grand rounds. AB - In staff development, varying methods of presentation can often make or break the learning process. A mock trial can help to increase nurses' awareness of the legal system and decrease the likelihood of a nurse being named in a medical negligence case. The mock trial presented at Texas Children's Hospital demonstrated what should and should not be documented and the importance of practicing within nursing standards of practice and hospital policies. PMID- 10531897 TI - Courses without classrooms. AB - There is an alternative to classroom lecture that provides faster, more complete instruction and introduces the learner to clinical application of skills in a safe environment. This teaching style uses multiple media to present professional, published resources that provide excellent quality, topic-specific information. The benefits of this type of teaching/learning module include improved use of student and instructor time, scheduling advantages, increased learning, revenue generation potential, and student empowerment. With this approach, a strong, sound educational base is built, and each course includes some degree or form of clinical application as a key component. PMID- 10531898 TI - A descriptive study of current transcultural education programs for registered nurses in selected Pennsylvania home health agencies. AB - The purpose of this study was to examine the features that comprise transcultural education programs provided by home health agencies for registered nurse staff within selected Pennsylvania agencies. This descriptive research study encompassed a survey that was mailed to a representative sample of home health agency administrators with a 54% response rate. The survey solicited answers to questions regarding agency demographics, patient and staff composition, and current transcultural education programs. The study serves to expand basic research in transcultural education programs within home health care. PMID- 10531899 TI - The impact of education on nurses' beliefs regarding family presence in a resuscitation room. AB - Allowing families to be present in the resuscitation room is an issue in sudden death situations. This study examined whether or not a class given to critical care and emergency nurses could change nurses' beliefs regarding the presence of family members in the resuscitation room. A convenience sample of 46 nurses was given a class regarding the benefits of families in the resuscitation room, present law, hospital policy, and how to implement this nursing action. The study used a quantitative, quasi-experimental study with a pre- and posttest design. It was found that nurses' beliefs regarding families in the resuscitation room during sudden death situations changed to a statistically significant level after attending a class. The study shows that an educational class can make a difference in the beliefs of nurses caring for families of sudden death victims. PMID- 10531900 TI - Making the best of a bad situation. PMID- 10531901 TI - Assessing children for obstructive sleep apnea. AB - The incidence of obstructive sleep apnea (OSA) in children has only recently been appreciated. Early identification of this disorder is extremely important to minimize neurologic, cardiac, and/or developmental complications that may occur as a result of the syndrome. This article presents an overview of OSA syndrome, pediatric risk factors for sleep apnea, symptoms common in children with OSA, and assessment information to guide the health practitioner in identifying children whose sleep patterns require further evaluation. PMID- 10531902 TI - Pediatric practice guidelines: implications for nurse practitioners. AB - Practice guidelines are promoted as an important means of achieving high-quality, cost-effective health care. Nurse practitioners must understand what practice guidelines are and how they are developed and be willing to put them into practice. This discussion begins with a description of practice guidelines specific to pediatrics. The terminology used in reference to these "clinical tools" are differentiated and their historic and contemporary influences are summarized. The complexity of guideline development and attributes of a quality practice guideline are described. Finally, the pivotal roles nurse practitioners can play in putting guidelines into practice are suggested. PMID- 10531903 TI - Male child sexual abuse. AB - Up to 92,000 male children report sexual abuse each year, and as many as 31% of all male children under age 18 years experience sexual molestation. Male child sexual abuse is now believed to be a far more common occurrence than it once was. Pediatric nurse practitioners are in a key position to prevent and recognize the sexual exploitation of male children. This article addresses the incidence of male child sexual abuse, the psychological and physical ramifications for the child, and the roles and responsibilities of the clinician, including interview, physical and psychological assessment, legal aspects of reporting, and referral. Prevention techniques in a primary care setting are also discussed. PMID- 10531904 TI - Playing for time: adolescent perspectives of lung transplantation for cystic fibrosis. AB - A single-case study approach was used to provide an in-depth examination of the special events surrounding the decision of a 21-year-old adolescent to undergo lung transplantation for end-stage cystic fibrosis. The central theme "playing for time" characterized the interplay between the disease progression and adolescent development as illustrated by 3 subthemes: (a) a strange balance; (b) playing chicken; and (c) being listed. The adolescent's developmental needs provided the context for the struggle with the competing demands of physiologic and psychologic readiness for the transplant, quality-of-life issues, and a renewed hope for the future. Developmental needs were more important to the adolescent than the opportunity for increased length of survival provided by lung transplantation. Advanced practice nurses are in an excellent position to provide continuity of care for chronic illness management over time and across settings. PMID- 10531906 TI - Sun smarts: the essentials of sun protection. PMID- 10531905 TI - Risk taking in young Hispanic children. AB - INTRODUCTION: The purpose of this study was to examine risk taking and daring behavior in preschool-age Hispanic children. The study sought to describe aspects of children's personality, behavior, and culture that may inform us of their propensity to take risks that lead to injuries. METHOD: Forty-five children (4 to 5 years of age) and their parents, drawn from a primary care practice, participated in the study. Instruments completed by the families included the Acculturation Scale. Child Shyness Report, Injury Report, and the Injury Behavior Checklist. Children were interviewed about risk taking and daring behavior using a projective technique (Child Sensation Seeking Profile). RESULTS: These children, irrespective of gender and socioeconomic status, reported similar rates of daring and risk taking behavior. Injury behaviors were not predicted by personality profiles of shyness or the child's self-report of risk taking and daring behavior. Actual injuries increased with higher levels of acculturation, but children's injury behaviors reported by parents were low in comparison with other populations. Discrepancies were observed between parental perception and report of children's injury behavior and children's expressed preferences in some domains of daring and risky behavior. DISCUSSION: Primary care providers must consider ethnic differences in rates and causes of injury when developing interventions and injury prevention programs. PNPs can use these findings to better meet the health promotion goals of Healthy Children 2000. PMID- 10531907 TI - Extended families: social support systems for children. PMID- 10531908 TI - Identifying and documenting findings of physical child abuse and neglect. PMID- 10531909 TI - Chondromalacia patella. PMID- 10531910 TI - "Access to pediatric care" provisions should include PNPs. PMID- 10531911 TI - Enabling parents to "read" their baby. PMID- 10531912 TI - The role of pediatric professionals in early emotional development. PMID- 10531914 TI - Motherhood: the emotional awakening. AB - During her physical pregnancy, a mother also undergoes a psychological pregnancy in which she imagines her baby as having certain physical and intellectual attributes. Shortly after birth, a mother finds that her "imagined baby" encounters her "real baby"--a moment when her prepartum thoughts of an idealized baby may conflict with the reality of her newborn. This encounter contributes to the future of their relationship in many ways, some of which may have clinical implications. For example, in the case of a preterm or premature birth, the mother is also premature psychologically and may need professional support to effectively deal with her unexpected infant. The Brazelton Neonatal Behavioral Assessment Scale can be an effective therapeutic tool during these perinatal encounters. PMID- 10531913 TI - Research to practice: emotional development and maternal/infant attachment. AB - A mother's responsiveness to her infant's signals is important for developing their personal relationship and the child's social and cognitive competence. While interacting, both mother and infant emit signals to capture each other's attention and to indicate whether to join, sustain, or terminate their interaction. Maternal sensitivity to these signals is a central feature in the development of optimal or secure attachment. However, a mother's perceptions and expectations of her infant's behavior affects her sensitivity to infant signals. Because of the effect of these parental perceptions, modifying the parent's cognitive sets and coping strategies to help them better deal with the challenge of responding to infant distress can be beneficial. PMID- 10531915 TI - Using the language of the child's behavior in your work with families. AB - A child's behavior, and the way parents react to and understand that behavior, are important for professionals who work with families. Parental representations of a child's behavior provide an insight into their awareness of child development, as well as their thoughts, past experiences, values, and concerns. An appreciation of this parental viewpoint leads to a deeper and more effective professional relationship. In particular, when parents and professionals share their observations of a child's behavior and explore the meanings therein, they discover beneficial new ways of working together in support of the child and the family. PMID- 10531916 TI - Impact of a community-based program on early childhood development. AB - To determine the impact of an integrated community-based program in rural villages on early childhood development, a controlled trial was conducted in the Nakhon Sawan province of Thailand. The program involved the cooperation of governmental agencies, nongovernmental agencies, academic institutions, and community organizations. At baseline, 3 control and 3 program villages were similar in terms of nutritional status, developmental performance and parental care. After 2 years of intervention in the program villages, improvements were noted in nutritional status, developmental performance, and intelligence quotient scores, as well as overall utilization of health care resources and parental attitude and involvement. PMID- 10531917 TI - How do you manufacture a good cushion or mattress designed to reduce the incidence of pressure sores? PMID- 10531918 TI - Medical Editors Trial Amnesty (META) PMID- 10531919 TI - Observations on mattress covers: results of a pilot study. AB - Samples of covers from three commercially available mattresses were examined in the laboratory using test methods originally devised for testing surgical dressings. These revealed that although the covers shared many common features, there were differences in the conformability and tensile properties which may be of some clinical relevance. The study also confirmed that with some minor modifications, the experimental techniques used would be suitable for a future, more comprehensive review of mattress performance. In a separate investigation designed to examine the consequences of a failure of a mattress cover, the bioburden of a foam core removed from a damaged cover revealed the presence of very large numbers of microorganism, well in excess of 10(10) per gram of foam which could act as a recevoir of contamination and thus a source of cross infection. PMID- 10531921 TI - Testing the effectiveness of patient support systems: the importance of indentor geometry. PMID- 10531920 TI - Water vapour permeable materials for mattress coverings. PMID- 10531922 TI - Guide to flexible polyurethane foams. PMID- 10531923 TI - The role of the biomedical engineer in pressure sore prevention--a personal view. PMID- 10531924 TI - Paediatric pressure sore risk assessment. PMID- 10531925 TI - Better use made of research based evidence about clinical effectiveness. PMID- 10531926 TI - Removing the 'estimates and guesses' from practice--evidence based tissue viability. AB - The development of evidence based tissue viability is reviewed using the prevention and management of pressure sores as a specific example. Although data is limited, pressure sores may be more common at the end of the 1990's than they were fifteen years ago, despite changes in clinical practice. If evidence based tissue viability is to grow and mature, then two key obstacles must be overcome; changes in patient populations must be reflected within prevalence or incidence data (case-mix adjustment) so allowing interpretation of longitudinal changes in the occurrence of sores. Secondly, the efficacy and effectiveness of interventions such as pressure-redistributing beds and mattresses must be identified through rigorous and appropriate methodologies. Only through such steps can clinical skill be blended with research evidence to fully realise evidence based practice. PMID- 10531927 TI - Evidence-based medicine: a critique. PMID- 10531928 TI - Regional study days. Their role within the education strategy of the TVS. AB - The Tissue Viability Society (TVS) Regional Study Days (RSDs) aim to bring high quality education on pressure sore prevention and wound care to a large section of health care professionals who may well not attend conferences and who often do not find the time to read articles in journals and magazines. Since 1987 over 200 RSDs have been held providing education to over 20,000 delegates. The RSDs have also contributed in the region of 50,000 Pounds to the Society's funds. The first RSDs were held in the West Country, but since that time the area has extended to include the whole of the United Kingdom. The panel of approximately 12 speakers across the country are all recognised experts in the subjects of pressure sore prevention and wound care. Attendance at RSDs is free to all NHS staff. The historical background of the RSDs is presented and the manner in which they fit into the overall strategy of the TVS to provide an educational service to all levels of health care professionals. PMID- 10531929 TI - Wound management products--the evidence we need and the difficulties in obtaining it. AB - The management of complex wounds requires the expertise and co-ordinated effort of multiple disciplines in a variety of healthcare settings. Providers of wound care will be held increasingly responsible for the impact of clinical and economic outcomes for the patient, healthcare system and, ultimately, society as a whole. Collecting and analysing outcome data and comparing it to regional and national standards, will be the method which demonstrates the quality of service provided. This paper will review the design implications for conducting comparisons of therapies used in the management of wounds. Consideration will be given to overall design, sample size calculations, clinical setting, choice of comparator and mechanisms for exploring how efficacy and cost data can be linked. Evidence obtained from several prospective randomised clinical trials will be used as examples to support the theoretical considerations. PMID- 10531930 TI - Disturbance and resilience: an overview of evidence based practice. PMID- 10531931 TI - The role of the researcher in tissue viability--a personal view from a biomedical engineer. PMID- 10531932 TI - Pressure sore incidence at St George's Healthcare NHS Trust. PMID- 10531933 TI - The cost-effectiveness of high compression. PMID- 10531934 TI - Microbial involvement in chronic wound malodour. AB - The role of specific micro-organisms in producing chronic wound malodour was investigated by directly comparing odour severity and microbiology in infected and non-infected leg ulcers. Malodour was most frequently associated with infected wounds involving mixed aerobic and anaerobic, Gram-positive and Gram negative microbial populations. Infected ulcers that were not characterised by an offensive odour were rarely colonised with anaerobic bacteria. A reduced incidence of pigmented and non-pigmented Gram-negative anaerobes (Bacteriodes spp, Prevotella spp, Porphyromonas spp) was evident in non-infected, non malodorous leg ulcers. These observations emphasise the significance of specific anaerobic bacteria in the generation of wound malodour, and it is probable that their effect is potentiated by coexistence with mixed facultative micro organisms. PMID- 10531935 TI - Hospital monitoring of pressure ulcers in the UK. AB - Hospital trusts and health authorities are increasingly being expected to report on numbers of patients with pressure ulcers. Although pressure ulcer audits should be undertaken in a systematic way, there are no agreed standards and little is known about the current status of pressure ulcer monitoring. A postal survey was conducted to establish a national picture of both assessment and audit of pressure ulcers. Questionnaires were sent to 276 acute hospital trusts throughout the UK; 204 were returned, giving a response rate of 74%. Fewer than half of the trusts (40%) employed a tissue viability nurse, and 28.5% of respondents were another type of clinical nurse specialist. Most hospitals used single scales for grading pressure ulcers and risk assessment but the frequency and method of such surveys varied considerably. Relatively few hospitals (5%) monitored the number of pressure ulcers on a daily basis, 30% made some form of weekly return and 22% made monthly returns. The authors conclude that there are national variations with respect to the pressure ulcer assessment tools used to provide audit data, and to the methodology and documentation used to collate data. A national consensus is recommended on auditing of pressure ulcers in the UK. PMID- 10531936 TI - Using care pathways in pressure area management: a pilot study. AB - Care pathways are a method of managing, delivering and documenting care. A pilot study of a care pathway system in pressure area management was conducted in two clinical areas. Results from this pilot study indicate that the pathway approach to pressure area management is both useful and valid as a means to enhance clinical decision-making and to facilitate comprehensive pressure area management and the completeness of care documentation. Implementation of the pathway requires a facilitator to provide support and education and to monitor and maintain the change process. PMID- 10531937 TI - The management of fungating wounds. PMID- 10531938 TI - Wound debridement, Part 1: Non-sharp techniques. PMID- 10531939 TI - Comparative cost-effectiveness of four-layer bandaging in the treatment of venous leg ulceration. AB - This paper reports the results of an analysis designed to estimate the expected annual cost per patient of treating venous leg ulcers, and to evaluate the relative cost-effectiveness of a systematic treatment regimen using a four-layer compression bandaging system (Profore) compared with usual care. A Markov model has been developed which simulates the transition of patients between health states (healed and unhealed) over a 52-week period. Healing rates used in the model are derived from those reported in the literature. By running the model for a cohort of 100 patients over 52 weeks it is possible to estimate expected outcomes and annual budgetary costs for alternative treatment regimens. Results suggest that, when compared with usual care, a systematic treatment regimen using Profore is unambiguously more cost-effective. Patient outcomes are improved and annual treatment costs reduced. An important implication is that failure to co ordinate treatment policies and to use the most cost-effective treatments may result in substantial inefficiency in the use of NHS resources. This inefficiency could represent the equivalent of between 350,000 Pounds and 1.08 million Pounds annually for a typical health authority. PMID- 10531940 TI - Swab taking. PMID- 10531941 TI - The supply of dressings by hospital pharmacy departments in Wales. AB - A research project to examine the dressings and wound management materials supplied by pharmacy departments in Wales was carried out. Data were collected from the NHS computer database, which contains information on 17 hospitals, servicing 80-90% of the Welsh population. Over a 12-month period, hospital pharmacy departments throughout Wales issued dressings and related materials costing 2.1 m Pounds. As not all pharmacy departments supply the full range of dressings used in hospitals, and only one issues surgical sutures, this sum probably represents less than half of the total expenditure in Wales on this commodity group. Existing methods for recording the issue of dressings do not provide reliable information either on the range of products used or the total quantities issued throughout Wales. Despite expectations to the contrary, hospital use of dressings did not appear to have a major influence over prescribing patterns in the primary sector. The amount of cavity wound dressings that are supplied on an outpatient basis suggests a real need for these items to be available in the community. (Since completion of this study, cavity wound dressings have been added to the Drug Tariff.) PMID- 10531942 TI - Using information technology in wound management. PMID- 10531943 TI - [Two in one bed]. PMID- 10531944 TI - [Which factors are influencing nurses in the evaluation of pain?]. PMID- 10531946 TI - [Drawings by children]. PMID- 10531945 TI - [Travel vaccinations for children]. PMID- 10531947 TI - ["Wherever there are children ..."--viewpoints and work of an educator in the pediatric ward]. PMID- 10531948 TI - [How do you feel about mistakes? Thoughts of a pediatric nurse on the topic of mistakes and learning]. PMID- 10531949 TI - [Pediatric intensive care in Ulm]. PMID- 10531950 TI - [DTP-combinations: optimal protection with minimal stress]. PMID- 10531951 TI - [Malaria update 1999]. PMID- 10531952 TI - [Piercing is medical practice and is allowed only with a special permit]. PMID- 10531953 TI - Nurses and fight against tuberculosis. PMID- 10531954 TI - Myeloma. An anecdote. PMID- 10531955 TI - Oh! I have found my key! PMID- 10531956 TI - A psycho-educational programme for the primigravidae. PMID- 10531957 TI - The nursing service administrator. PMID- 10531958 TI - AIDS and the role of nurses. PMID- 10531959 TI - Self-awareness in breast cancer. PMID- 10531960 TI - The cyclonic disaster of Gujarat. PMID- 10531961 TI - The role of a nurse in early trauma management. AB - Optimal care of the patient with multiple trauma injuries requires an enormous amount of effort which begins with planning and preparation of system development in which personnel are educated and trained. Guidelines must include all aspects of the patient's care and the team must be drilled well. Protocols for management should be customized to suit the facilities available and the needs of the patient. The Primary Survey provides basic data essential when life is threatened. The Secondary Survey provides detailed assessment of the various organ systems. Inclusion of Psycho-Social aspects achieves a holistic approach to the victims of Trauma. PMID- 10531962 TI - Adaptability and cohesiveness among rural schizophrenic, epileptic and normal families. A comparative study. PMID- 10531963 TI - When a disaster strikes. Managing multi and mass casualties. PMID- 10531964 TI - Dr. (Ms.) Gro Harlem Brundtland. First woman Director-General of WHO. PMID- 10531965 TI - Mentally ill abandoned even in death. PMID- 10531967 TI - Focus on innovative approaches to nursing education. PMID- 10531966 TI - Nurses and nursing students must get their due PMID- 10531968 TI - Nurses' role in the prevention of suicide. PMID- 10531970 TI - Real boost for nursing. PMID- 10531969 TI - Negotiation. An art for nurses. PMID- 10531971 TI - Saving lives: a new crusade. PMID- 10531972 TI - The new Nye nurses. PMID- 10531973 TI - Sun, sea and sex. PMID- 10531974 TI - Giving refuge to those in need. Interview by Dina Leifer. PMID- 10531975 TI - Fighting spirit. PMID- 10531976 TI - Impulsive action. PMID- 10531977 TI - It's good to be back. PMID- 10531979 TI - See how you like it. PMID- 10531978 TI - England's new strategy for nursing is a beginning. PMID- 10531982 TI - The nurse practitioner role in New South Wales: development and policy. AB - The award of a Florence Nightingale Scholarship enabled research which looked at the role and development of nurse practitioners to be undertaken in three Australian states. Maxine Offredy relates the issues to current developments in the UK. PMID- 10531980 TI - ICN on tobacco. PMID- 10531981 TI - The expanded role of the registered nurse: studying nurses' perceptions. AB - Expanded roles for nurses have developed in the UK over the past 25 years and are now commonplace in most clinical settings. However, considerable confusion and difference of opinion exist about the role and its implications for practice. Shelagh Leonard describes a study undertaken to ascertain how the registered nurses who carry out expanded role activities perceive their role. PMID- 10531983 TI - Audiology and hearing impairment: improving the quality of care. AB - Hearing impairment can hinder nurses' and carers' ability to provide high quality care. In this article, the author provides basic information on audiology and hearing aids to help nurses and carers communicate more effectively with people with a hearing impairment. PMID- 10531984 TI - Cardiopulmonary resuscitation: the laryngeal mask airway. PMID- 10531985 TI - The risk of sun exposure. PMID- 10531986 TI - The National Board in Scotland. PMID- 10531987 TI - Mental health nursing: education. PMID- 10531988 TI - Bricks and mortification. PMID- 10531989 TI - Helping you care for patients. PMID- 10531990 TI - Closing the theory-practice gap. PMID- 10531991 TI - Praise is not enough. PMID- 10531992 TI - Life after brain injury. PMID- 10531993 TI - No way out. PMID- 10531994 TI - Learning what to say. PMID- 10531995 TI - Common sense caring. PMID- 10531996 TI - Star teachers. PMID- 10531997 TI - The joy of writing. PMID- 10531998 TI - Lest we forget. PMID- 10531999 TI - RCN R&D Co-ordinating Centre. PMID- 10532001 TI - Academic essay writing in the first person: a guide for undergraduates. AB - In this article, Conal Hamill aims to contribute to the on-going debate about the appropriate use of first person writing in academic nursing assignments and provide guidance for nursing undergraduates. PMID- 10532000 TI - Learning disability: promoting health equality in the community. AB - A project to discover if people with a learning disability would benefit from a pop-in health clinic has shown that a needs-led service, co-ordinated by a health professional, can reduce gaps and inequalities within healthcare provision. PMID- 10532003 TI - Cannabis: the evidence. AB - There has been a lot of publicity over the past year on the issue of whether the legal status of cannabis should be changed. Much of the debate has been emotional in nature. The aim of this article is to review the literature and to come to some conclusions based on sound scientific rationale and empirical studies. PMID- 10532002 TI - Reciprocal secondment. AB - Reciprocal secondment can improve relationships between organisations and be of great benefit to the individuals involved. Damien Black and Paula Martyn describe the planning and outcomes of their reciprocal secondment in the fields of health authority nursing and nurse education. PMID- 10532004 TI - Mechanical ventilation. AB - In this article Nigel Henderson discusses the nursing role in the mechanical ventilation of patients. He describes the different ventilator modes, and the physical and psychological care required by ventilated patients. PMID- 10532005 TI - The diet of people with Crohn's disease. PMID- 10532006 TI - The Welsh National Board. PMID- 10532007 TI - Sick children at home. PMID- 10532008 TI - Investing in our future professionals: Pediatric Nursing Fellowship Program. PMID- 10532009 TI - Health promotion and injury prevention behaviors of elementary school children. AB - This cross-sectional study examined health and lifestyle behaviors of 302 urban elementary school children. Over half of the children considered themselves very healthy, with asthma and allergies the most commonly reported illnesses. The majority of children reported high levels of injury prevention and health promotion behaviors. Over 75% of the students reported that they did not smoke, drink, use guns or drugs; they looked both ways before crossing a street; took medicine only with parental permission; had a working smoke detector at home; and knew how to safely exit their home during a fire. Boys reported more risk-taking behaviors than did girls; White youth had lower injury-prevention scores than Black youth; and younger children and children with behavior or emotional disorders in specialized classrooms reported fewer health promotion activities related to nutrition, exercise, and dental hygiene. Findings suggest the need for tailoring health education efforts for different subgroups of children. PMID- 10532010 TI - Pediatric occupant car safety: clinical implications based on recent literature. AB - Seat belts and child safety seats can provide effective protection against serious and fatal injuries, but motor vehicle crashes (MVCs) remain a leading cause of death and injury among infants and children. Recent fatalities associated with air bags have intensified awareness of pediatric occupant car safety issues. A review of pediatric literature from 1989 to 1997 summarizes studies on the correct use, incorrect use, and non-use of child safety restraint devices; injuries patterns to children as occupants in MVCs; transporting children with special needs; lethal air bag injuries; and injury prevention educational programs. PMID- 10532012 TI - Development of a breastfeeding support program at Naval Hospital Sigonella, Italy. AB - The Naval Hospital (NH) Sigonella, which contains a level one nursery, cares for prenatal patients in an isolated military community. Many of the traditional support systems available to breastfeeding mothers in the United States, such as Le Leche League, are unavailable in this setting. Some prenatal patients in this setting who have had the desire to breastfeed have failed to do so because they were not offered a structured educational support system that encouraged and monitored their breastfeeding efforts. This prompted the pediatric nurse, functioning as breastfeeding counselor nurse at NH Sigonella, to develop a breastfeeding support program. The goal of the program is to educate new mothers about breastfeeding and to encourage them to initiate and continue breastfeeding for at least 5 to 6 months. These goals are in line with the Healthy People 2000 (1990) objectives, which are to increase to at least 75% the proportion of mothers who breastfeed their babies in the early postpartum period and to at least 50% the proportion of those who continue breastfeeding until their babies are 5 to 6 months old. At delivery, the breastfeeding counselor meets individually with each new mother who expresses the desire to breastfeed. During this visit, she observes and evaluates each mother's breastfeeding technique. Discharge follow-up via telephone is conducted weekly for 4 weeks and at 3 and 6 months postpartum. Data collection after 6 months revealed that 86.7% of all new mothers initiated breastfeeding and 51.4% of those mothers continued breastfeeding for 5 to 6 months. The Breastfeeding Counseling Support Program will enhance NH Sigonella's efforts in continuing to meeting the objectives of Healthy People 2000 and the Breast Feeding Hospital Initiative while providing the benefits of breastfeeding to mother and infant. PMID- 10532011 TI - Are ball pits the playground for potentially harmful bacteria? AB - Ball pits, enclosed play areas with padded floors and pits of small plastic balls, have become popular features for children at fast food restaurants. This pilot study sought to identify and confirm bacterial organisms that place children at a potential health risk in three play pits within fast food restaurants. Data for this descriptive study were randomly collected from restaurants offering play pits with multicolored, round, hollow, plastic balls within urban communities of the Tidewater region of Virginia. Specimens were collected from entrances into the ball pits as well as various areas of the bottom lining to incur a representative sample. Results indicated an increased level of normal flora as well as nonhuman flora, demonstrating that bacteria are present within the ball pits. The results question the safety of these play pits for both health care providers and parents. Nurses play a vital role in public awareness through health education. Disinfection protocol and proper handwashing are the keys to making ball pit play areas safe for children. PMID- 10532013 TI - Guideline for i.v. infiltrations in pediatric patients. AB - A large Midwestern tertiary care center used a multidisciplinary approach to develop an intravenous infiltration/extravasation guideline for pediatric patients, ages 0-18 years old, using the Iowa Model for research utilization. This infiltration clinical practice guideline included a site appearance staging tool, decision algorithm, research-based antidotes, and standard of care. The goal of the guideline was to prevent or minimize adverse occurrences for pediatric patients at risk for intravenous infiltrations. Quality assessment and improvement tracking suggested that there was an increased consistency in use of practice guideline interventions for and reporting of the infiltration event, a reduction in adverse patient outcomes and potential cost savings. PMID- 10532014 TI - Telemedicine in the neonatal intensive care unit. AB - A program of telemedicine has been developed in the NICU at the Beth Israel Deaconess Medical Center (BIDMC), in Boston, Massachusetts. The telemedicine project, "Baby Care Link," allows families to have increased access to their infant, care team, and educational information during hospitalization and following discharge. The goal of this multidisciplinary project is to provide an approach to managing the care of the very low birth weight (VLBW) infant. Workstations in the families' homes provide education, care schedules, remote sensor links to clinical providers, and videoconferencing. The linkage of home, clinician, and community centers will allow for the episodic needs of high-risk infants while continuously monitoring progress and managing treatments systematically in the home rather than the NICU setting. It is believed that the use of this emerging technology might lead to shorter hospital stays as well as increased parental understanding and comfort, overall parental satisfaction with care, improvement in health status, improvement in clinician satisfaction, and lastly, a reduction in hospital associated costs. PMID- 10532015 TI - Infant nasal-pharyngeal suctioning: is it beneficial? AB - PURPOSE: To identify and validate the indicators registered nurses (RNs) use when deciding whether or not infants experiencing respiratory illnesses require nasal pharyngeal (NP) suctioning. METHOD: The first part of this study involved 43 RNs completing a questionnaire that examined the assessment parameters RNs use in determining when to perform NP suctioning. The second part of this study involved assessing infants' respiratory status before and after NP suctioning to determine what, if anything, changed post suctioning. FINDINGS: The three assessment parameters most frequently used by nurses in determining the need for suctioning were pulse oximeter readings, visible secretions, and audible secretions. The only three parameters to show statistically significant improvement post suctioning were pulse oximeter readings, visible secretions, and audible secretions. CONCLUSIONS: Nurses are using valid assessment parameters when determining the need for infant NP suctioning, and NP suctioning does improve certain aspects of infants' respiratory status. There are presumed benefits to NP suctioning. However, there are also potential risks. Nasal-pharyngeal suctioning can produce bradycardia, laryngospasm, cardiac dysrhythmias, and edema and trauma to mucous membranes (Oberc, 1991), tachycardia, emotional distress, bronchospasm, and cardiac arrest (Young, 1988). The procedure involves inserting a catheter into an infant's nose and advancing it to the back of the throat (pharynx) approximately 4-8 cm (Oberc, 1991). Once at the pharynx, suction is intermittently applied and the catheter is slowly removed. The purpose of NP suctioning is to clear the airway of an infant who is unable to do so independently. However, the effectiveness and outcomes of this procedure have not been supported by research. PMID- 10532016 TI - Ethics, the outpatient pediatric nurse and managed care. AB - The nurse in an ambulatory setting has a new role in participating in the managed care process. This article reviews new responsibilities and the ethical dilemmas that might be faced. PMID- 10532017 TI - Pediatric management problems. Bacterial vaginosis (BV). PMID- 10532018 TI - Rotavirus vaccine, live, oral, tetravalent (RotaShield). AB - Nearly every child will experience rotavirus infection before the age of 5. Rotavirus is transmitted via a fecal-oral route. Because the virus is shed in the stool, outbreaks of rotavirus infection can occur on the pediatric hospital wards and in day care centers. Ingestion of the rotavirus particles infects the cells in the villi of the small intestine. Copious amounts of watery diarrhea will occur after an incubation period of 1 to 2 days. If untreated, children less than 2 years of age can die from the resulting severe gastroenteritis dehydration. In the United States, rotavirus infection peaks during the winter months and is the cause of most cases of diarrhea in infants and young children. Rotavirus infection accounts for approximately 70,000 hospital admissions for diarrhea and as many as 100 deaths each year in the United States. World wide rotavirus infection accounts for approximately 1 million deaths each year (Bass, 1996). Although the number of deaths from rotaviral disease in the United States is low, parents frequently miss work, have to arrange for alternative care, travel to the doctor, give their child oral rehydrating solutions, and buy extra diapers. Implementation of mass vaccination with RotaShield will significantly reduce these indirect costs. PMID- 10532019 TI - Psychological comorbidity in children with chronic conditions. AB - The incidence and prevalence of chronic conditions in children are increasing along with a concomitant rise in the number of psychological problems they experience. When psychological comorbidity exists, the child is at greater risk for numerous problems. This article will define psychological comorbidity and discuss its incidence, presentation, associated factors, and interventions. PMID- 10532020 TI - Recommended immunization schedule for 1999: update on the changes. PMID- 10532021 TI - Exploring language about families. AB - When we work with families in health care settings, it is important to be aware of the way we communicate. Often overlooked is how the language we use to describe and understand families affects how we work with them. The language we use in thinking about a family can shape our perceptions of the family and may affect how we approach working with them. The language we use in describing a family to another health care provider can affect how that person will perceive and approach the family. The language families hear us use can affect families' perceptions of themselves, their perceptions of us, and, consequently, how they relate to us. In Project Copernicus' Family Centered Communication Skills: Facilitator's Guide (Edelman, Greenland, & Mills, 1993), an activity entitled "Watch Our Language" explores commonly used negative terminology about families and guidelines for better language. This exercise has been used with groups of nurses and other health care providers at several conferences and has generated thoughtful brainstorming about negative language related to families, its impact on families, and its impact on the nurses working with them. Those participating in the workshops explored better terminology about families and brainstormed a list of selected family strengths which are shared here. PMID- 10532023 TI - Helping people cope with epilepsy PMID- 10532022 TI - A role we can learn from PMID- 10532024 TI - Empowering alice PMID- 10532025 TI - Staff perceptions of night nurse clinicians. AB - Many night nurses receive insufficient support from senior nurses. Reducing junior doctors' hours increases pressure on night nurses to carry out a wider range of duties. Night nurse clinicians benefit junior doctors, nurses and patients. PMID- 10532026 TI - Qualitative research: data analysis techniques. AB - Qualitative data usually consist of the words or actions of participants. These data can be difficult to condense and organise without losing their meaning. Analysis of qualitative data requires considerable creativity on the part of the researcher. PMID- 10532027 TI - Weaning patients from mechanical ventilation. AB - Enhancing the nursing role can allow staff to provide more individualised care. Nurses in ICU can become involved in weaning patients from ventilators. Multidisciplinary team communication is vital in such advanced nursing practice. PMID- 10532028 TI - Managing workplace stress in outpatient nursing. AB - Nurses are subject to a variety of work-related stressors. Recent reorganisation in the NHS has led to job insecurity. Nurses have developed a variety of strategies for coping with stress. Meeting training needs, protecting time for discussion, and improvements in two-way communication help alleviate stress. PMID- 10532029 TI - Introducing and auditing a nurse-led leg ulcer service. AB - Leg ulcers have a significant impact on quality of life, and represent a major treatment cost for health services. Improved healing rates benefit patients and health service provision. Nurse-led clinics provide an appropriate setting for treatment. PMID- 10532030 TI - Anaphylactic reactions during chemotherapy. AB - Anaphylactic reactions are very frightening for patients. New chemotherapy drugs increase the possibility of anaphylaxis. Its rapid onset and severity make it vital that nurses can recognise anaphylaxis. PMID- 10532031 TI - Contraception. PMID- 10532032 TI - Epilepsy management. PMID- 10532033 TI - Home and laboratory pregnancy-testing kits. AB - Home pregnancy test kits are more sensitive than many pregnancy kits used in laboratories or health centres. Nurses should know the sensitivity of the kit used in their centre or laboratory. A positive home pregnancy test should always be treated as the correct result until sufficient tests have been carried out to prove otherwise. PMID- 10532034 TI - Dementia. Part 1: Person-centred care. AB - Those caring for people with dementia should promote 'personhood'. Humanistic philosophical theories offer nurses a positive basis for person-centred care. Humanistic values about personhood should be central to nursing practice, regardless of the care setting. PMID- 10532035 TI - Nursing children at home. PMID- 10532036 TI - [Nursing is a good profession]. PMID- 10532037 TI - [Complaining in order not to get lost]. PMID- 10532038 TI - [Beating isolation by talking]. PMID- 10532039 TI - [Enteral nutrition: from hospital to home]. PMID- 10532040 TI - [Plea for well executed enteral nutrition]. PMID- 10532041 TI - [Endoscopic gastrostomy for alimentation]. PMID- 10532042 TI - [Nursing care and enteral nutrition]. PMID- 10532043 TI - [Practical advice for enteral nutrition at home]. PMID- 10532044 TI - [Enteral nutrition from the dietitian's point of view]. PMID- 10532045 TI - [Home enteral nutrition]. PMID- 10532046 TI - [Nursing care at death]. PMID- 10532047 TI - [Facing the wasteland of accessible public transportation]. PMID- 10532048 TI - [A nurse in Naples, heat and chaos]. PMID- 10532049 TI - [How to improve patient admission]. PMID- 10532050 TI - [A battle in the air ... the fight against Aspergillus]. PMID- 10532051 TI - [Consistency of care in surgery for scoliosis]. PMID- 10532052 TI - [When a child's limp leads to hospitalization]. PMID- 10532053 TI - [Surgery for pyloric stenosis by laparoscopy]. PMID- 10532054 TI - [Educational role of the nurse before and after an intervention for urinary and fecal incontinence]. PMID- 10532055 TI - [Caring for a newborn after surgery for anorectal malformation]. PMID- 10532057 TI - [The nurse's responsibility for oral prescriptions]. PMID- 10532056 TI - [Ouch, it hurts...]. PMID- 10532058 TI - [The place of the nurse within the hospital]. PMID- 10532059 TI - [150 years Public Health]. PMID- 10532060 TI - [Interdisciplinary care of a patient with Huntington's disease]. PMID- 10532061 TI - [The newborn needing surgery, an interdisciplinary challenge]. PMID- 10532062 TI - [Prolonged fever]. PMID- 10532063 TI - [Fever after returning from a tropical country]. PMID- 10532064 TI - [Fever in children less than 3 months old]. PMID- 10532065 TI - [Severe hyperthermia in children]. PMID- 10532066 TI - [Measurement of temperature and fever in children]. PMID- 10532067 TI - [Thermometers: alternatives to mercury]. PMID- 10532068 TI - [Antipyretics]. PMID- 10532069 TI - [Fever: a symptom that should always be taken seriously]. PMID- 10532070 TI - [Small lexicon of molecular genetics]. PMID- 10532071 TI - [The incest taboo: fundamental indicators of identity]. PMID- 10532072 TI - [Nursing in Europe]. PMID- 10532073 TI - [Role and responsibility of nurses in the treatment of pain] [In Process Citation] PMID- 10532074 TI - [Solidarity against a virus]. PMID- 10532075 TI - [Evaluation of the introduction of a liaison notebook in the education nurses anesthetists]. PMID- 10532076 TI - [Cross infections]. PMID- 10532077 TI - [Importance of septic isolation to reduce the acquisition of multiply resistant bacteria in intensive care. Effects on the incidence of cross infections]. PMID- 10532078 TI - [Survey of cross infections in intensive care: search for a durable strategy]. PMID- 10532079 TI - [Action plan 1995-2000 against cross infections]. PMID- 10532080 TI - [Costs of cross infections]. PMID- 10532081 TI - [Standard hygiene. A method for decompartmentalization]. PMID- 10532082 TI - [Infection control nurse, soon a specialty all of its own]. PMID- 10532084 TI - [Talking about hygienic handwashing]. PMID- 10532083 TI - [Cleaning and disinfection of a room after a patient has left]. PMID- 10532085 TI - [Two new forms of Augmentin and Ciblor, frequently used in the fight against cross infections]. PMID- 10532086 TI - [What we have to remember about cross infections]. PMID- 10532087 TI - [The newborn needing surgery. Nursing diagnoses during the preoperative, perioperative and postoperative periods]. PMID- 10532088 TI - [The first years of ANFIIDE with Leonie Chaptal]. PMID- 10532089 TI - [Current chemo- and hormone therapy in disseminated malignant melanoma of the skin]. PMID- 10532090 TI - [Choice of first-line chemotherapy for advanced ovarian cancer]. PMID- 10532091 TI - [Estrogen-dependent peroxidase and hormone-dependent carcinogenesis]. PMID- 10532092 TI - [Letrozole (Femara), a new aromatase inhibitor for advanced breast cancer]. AB - The study compares letrozole (Femara and aminoglutethimide (AG), a standard therapy for postmenopausal women with advanced breast cancer, previously treated with anti-estrogens. 555 women were randomly assigned letrozole 2.5 mg once daily (n = 185), letrozole 0.5 mg once daily (n = 192) or aminoglutethimide 250 mg twice daily with corticosteroid support (n = 178) in an open-label, multicenter trial. The primary end-point was objective response rate (ORR), with time events as secondary. ORR was analysed nine months after enrollment of the last patient, while survival was analysed 15 months after the last patients was enrolled. We report the results of these analyses plus an extended period of observation (covering a total duration of approximately 45 months) to determine the duration of response and clinical benefit. Overall objective response rates (complete + partial) of 19.5%, 16.7% and 12.4% were seen for letrozole 2.5 mg, 0.5 mg and AG respectively. Median duration of response and stable disease was longest for letrozole 2.5 mg (21 months) compared with letrozole 0.5 mg (18 months) and AG (14 months). Letrozole 2.5 mg was superior to AG in time to progression, time to treatment failure and overall survival. Treatment-related adverse events occurred in fewer patients on letrozole (33%) than on AG (46%). Letrozole 2.5 mg offers longer disease control than aminoglutethimide and letrozole 0.5 mg in the treatment of postmenopausal women with advanced breast cancer, previously treated with anti-estrogens. PMID- 10532094 TI - [Detrimental effects of Daunorubicin and Doxorubicin on human erythrocytes in vitro]. AB - In isotonic medium, daunorubicin in the concentration range of 0.5-5.0 mg/ml of cells and doxorubicin in the concentration range of 0.2-1.0 mg/ml of cells caused hemoglobin (Hb) and K+ to efflux from red blood cells (RBC), to increase RBC size and to lower their deformability. Hb and K+ efflux rates were proportional to the antibiotic concentrations and remained stable for a few hours. Hb efflux did not change significantly in the 4-21 degrees C range (exempt at an experimental concentration of daunorubicin of 5.0 mg/ml of cells) but soared sharply at 37 degrees C. At the daunorubicin and doxorubicin concentration of 0.2 mg/ml of cells, Hb and K+ efflux virtually did not differ from control values. At the antibiotic concentration of 1.0 mg/ml of cells, 37 degrees C, Hb efflux rate was 0.34-5.6%, while that of K+(-) 1.0-8.2%, per hour, of possible maximum value. For the daunorubicin level of 5.0 mg/ml of cells, the respective values were 10 and 17%. At the daunorubicin concentration of 5.0 mg/ml, at 4 degrees C, Hb efflux from RBC was significantly higher than at 21 degrees C. RBC malleability, which was determined as their ability to pass through membrane filters having 3 mm dia pores, did not differ significantly from control values for a few hours antibiotic concentrations not exceeding 0.3 mg/ml of cells. At the antibiotic concentration of 1.0 mg/ml and higher RBC deformability dropped to zero within 10 min. ATP level in RBC practically remained identical to control values during incubation with the antibiotics for several hours. PMID- 10532093 TI - [Inhibin and ovarian cancer]. AB - Previous observations from our laboratory have demonstrated that the levels of immunoreactive inhibin (ir-inh) are elevated in almost all patients with granulosa cell tumors and in the majority of postmenopausal women with mucinous ovarian cancers. The present report confirms these findings in a larger group of post-menopausal women. Immunohistochemistry for the inhibin alpha. beta A and beta B sununits shows predominantly epithelial staining in granulosa cell tumors and in the majority of mucinous cancers. Serous cystadenocarcinomas also frequently show positive staining. Studies seeking to identify G alpha i-2 or FSH receptor mutations have provided negative results in contrast to other reports. Further studies of the roles of the inhibin-related family of peptides in ovarian cancer diagnosis and monitoring are clearly indicated. PMID- 10532095 TI - [Methods of autologous peripheral blood stem cell (PBSC) mobilization in oncological and hematological patients]. AB - The potential of peripheral blood stem cell (PBSC) mobilization with granulocyte (G-CSF) and granulocyte-macrophage (GM-CSF) colony-stimulating factors of chemotherapy has been assessed in patients with tumor or hematological disease. The study was intended to aid PBSC transplantation. It was shown that G-CSF administration increased the number of leukocytes in leukemia patients dramatically while mononuclear cell and colony-forming unit levels in G-CSF mobilised PBSC were significantly higher than those collected with GM-CSF or chemotherapy. PMID- 10532096 TI - [Increased apoptosis of peripheral blood leukocytes correlates with leukopenia after high-dose chemotherapy]. AB - Time-related changes in apoptotic leukocyte numbers were studied in peripheral blood sampled from 12 cancer patients before and after myeloablative chemotherapy (MC); six of them received hemopoietic cell grafts. Apoptotic leukocytes were counted in fresh and ex vivo incubated blood samples. Nuclear chromatin autolysis was registered in supravital Acridin Orange-stained granulocytes and lymphocytes. The levels of apoptosis in fresh peripheral blood were under 1.5%. Incubation for another 3 h revealed considerable numbers of apoptosis-prone leukocytes both in untreated patients and healthy donors. High-dose chemotherapy with busulphan, cyclophosphamide or alkeran was followed by a sharp increase in apoptotic cell counts which peaked on days 7-9, before leukopenia started on days 13-15 at the earliest. It is suggested that apoptosis of mature leukocytes is a major cause of leukopenia development induced by chemotherapy. The simple technique of apoptosis prone cell detection in whole blood may be effectively used in serial screening of hematological side-effects of chemotherapy. PMID- 10532097 TI - [Alpha-interferons in the treatment of patients with chronic myeloid leukemia]. AB - Chronic disease duration and survival have been investigated in three groups of patients suffering chronic myeloid leukemia (CML). The first group included 13 patients on alpha-interferons 6-9 mln MU/24 h (mean dose--48 mln MU/week). 31 patients received 2 mln MU/m2/24 h; mean weekly dose--15 x 10(6) MU. Standard chemotherapy was given to another 79 patients (group III). Actual survival and chronic disease duration were computed after Kaplan-Meyer: 4-year survival in group I--88%; group II--85.6% and group III--54%. Five-year survival in patients who had received standard or lower doses of alpha-interferon was 78.7%; chemotherapy alone--28.9%. Median survival in alpha-interferon-treated patients was 66 months; chemotherapy--48 months. After standard alpha-interferon, chronic disease three years after CML diagnosis was in 87% of those treated with standard alpha-interferon, 89% of those receiving lower doses of the drug and 53.4% of chemotherapy-treated patients. After 4 years, chronic disease was registered in 75.5% (alpha-interferon)--74.8% in group I and 72.9% in group II, and in 34.4% of patients treated with myelosan or hydroxyurea. Median chronic stage duration after interferon was 51 months and 39 months in group III, hence, both standard and lower doses of alpha-interferon prolong chronic disease and improve survival in CML patients. PMID- 10532098 TI - [Alpha-2a-interferon (Reaferon) in the treatment of patients with multiple myeloma]. AB - Sixty-six patients with multiple myeloma were divided into four groups: treatment with alpha-2a-interferon (reaferon) 3-5 mln MU/m2 alone (group); alpha-interferon + pulsed therapy with dexamethasone (group II); reaferon at high or low dose + different regemens of polychemotherapy (group III), and polychemotherapy followed by therapeutic support with melfalan and prednisone (MP) (control). Patients in group I untreated earlier with MP showed positive response in 80% while only 20% were in partial remission. Chemoresistance was broken in 86% (complete clinico hematological remission--18%) following administration of high doses of alpha interferon in conjunction with cytostatics. Reaferon + dexamethasone modality proved less effective. After chemoresistance was broken in 83%, tumor process reached a plateau. Low-dose reaferon was less effective, too, with complete remission never being reported. When administered in high doses as support therapy, it proved fairly effective, with median response being double that registered at low dosage or chemotherapy + MP. Whenever high-dose alpha interferon tolerability is poor, dosage can be decreased; the patients may continue on low dosage or switch to MP support. PMID- 10532099 TI - [Diagnostic and prognostic significance of blood-serum ceruloplasmin, acetylcholinesterase and total proteolytic activity in patients with multiple myeloma]. AB - The study included 82 patients with multiple myeloma (MM) to evaluate the diagnostic and prognostic significance of correlation between of levels of ceruloplasmin (CP), acetylcholinesterase (ACE) and total proteolytic activity (TPA) in blood serum and immunochemical pattern, tumor mass and response to chemotherapy. It was shown that CP, ACE and TPA determination may be used as additional markers to confirm therapeutic benefits, to timely detect relapse and to verify resistance to chemotherapy. It was also demonstrated that resultant decrease in CP and TPA and increase in ACE levels are reliable indicators of changes developing in MM course and remission fulfillment. High values for CP and low ones for ACE point to pathological activity. Both diagnostic and prognostic significance of the indices has been shown. The highest levels of CP and TPA in blood serum were identified in cases of A-myeloma and myeloma of Bence Jones while ACE concentrations in those patients were lower than in G-myeloma which correlated with median survival. Application of said assays might improve diagnosis, management and prognosis of MM. PMID- 10532100 TI - [Immunomodulator Cycloferon in the comprehensive treatment of patients with acute non-lymphoblastic leukemia]. AB - The therapeutic benefit and action of the immunomodulator Cycloferon were evaluated in patients with acute non-lymphoblastic leukemia. When administered in conjunction with chemotherapy during first complete remission, it reduced both early-onset relapse frequency and degree of cytostatic-induced immune deficiency. The drug's action is mediated by changes taking place in cytokine and interferon synthesis. PMID- 10532101 TI - [Treatment of metastatic pleural effusion with intrapleural administration of bleomycetin]. AB - The report presents data on conservative treatment of pleuritis induced by metastases from tumors of different localization by bleomycetin. After the pleural cavity was drained dry in 24 patients, bleomycetin 45 mg was introduced into it. The latter procedure was repeated in 6 cases because profuse exudation continued. The results were evaluated by clinical and roentgenological means. Bleomycetin treatment was painless. Mean drainage duration was 3.8 days, admission time--6.2 days. Stable remission was recorded in 16 patients, partial- 7. Pneumonia (2) and chronic empyema (1) were observed as complications. Bleomycetin-induced pleurodesis correlated with exudate pH: a pH value in excess of 7.3 made a patient good candidate for carrying out the procedure. According to Karnofsky index for long-term results, life quality of the patients improved. Eleven patients survived for 1 year after pleurodesis, and 2--more than two years. PMID- 10532102 TI - [Short-term and end-results of combined treatment of patients with stage-II cervical cancer]. AB - The short-term and end-results of combined therapy of 109 patients with stage II epidermal carcinoma of cervix uteri are presented. Preoperative intracavitary large-fraction contact gamma therapy from the AGAT-B installation was followed by Wertheim's or simple hysterectomy, lymphadenectomy and distant gamma therapy. Postoperative mortality rate was 0.8%; five-year survival--81.8 +/- 4.4%. Early- and late-onset postoperative complication frequency was largely determined by extent of surgery rather than preoperative intracavitary irradiation. PMID- 10532103 TI - [Long-term results of comprehensive treatment for cervical cancer with poor prognosis]. AB - The data on a randomized treatment of 100 patients with locally advanced cervical carcinoma are discussed. Half of the patients received complex therapy while the rest--radiation alone (control). The modality also included immunotherapy with epithalamin 100 mg, intramuscularly, neoadjuvant intraarterial polychemotherapy with 5-fluorouracil 2 g/m2 and cisplatin 100 mg/m2, concurrent radiotherapy with an interval and 4-6 courses of adjuvant polychemotherapy. As a consequence, short term results improved by 18.8% in cases tumor size under T3; end-results--by 14%. Improvement in survival was accounted for by causing tumor process to slow down considerably. The adjuvant polychemotherapy component was thought to be responsible for a 25.4% increase in one-year survival and 12.5%--in 3-year survival. PMID- 10532104 TI - [Evaluation of the anti-emetic effectiveness of two drug formulations of Ondansetron in combined chemotherapy for children with malignant tumors]. AB - A double-blind randomized comparison (protocol S3AB4003, Glaxo Wellcome, Great Britain) carried out in a group of 52 children showed that the 5-HT3-receptor antagonist ondansetron (in syrup) effectively prevented vomiting, nausea and loss of appetite caused in combination chemotherapy with highly- or moderately emetogenic cytostatic drugs in 92.3; 69.3 and 81.0%, respectively. Treatment was given to patients who were on dexamethasone. With intravenous injection of ondansetron plus dexamethasone, per os, said indices were 88.5; 73.2%; 73.2%, respectively, thus showing no significant differences. No side-effects were observed with either regimen. PMID- 10532105 TI - [Taxotere and methylnitrosourea: experimental appraisal of combination chemotherapy]. AB - The effectiveness of taxotere and methylnitrosourea (MNU) combined application against murine P388 leukemia has been studied. Antitumor drugs were administered separately or in combination in doses of 50, 60 or 70 mg/kg per day, i/p, beginning from day 1 after tumor transplantation. Combined administration was shown to potentiate antitumor effect, which also largely depended on schedule. Survival increase varied within 50-130% of control values. Optimal effect was observed with the following schedules: taxotere + MNU, 24-hour interval between injections, and MNU-taxotere, 4-day interval between injections. Synergism was in evidence since the therapeutic effect of combination treatment was significantly higher than that of either drug administered alone. PMID- 10532106 TI - [Experimental investigation of 2,2'-diamino-1,1'-dinaphthyl in rats and mice]. AB - Carcinogenicity of 2,2'-diamino-1,1'-dinaphthyl has been investigated in chronic experiments on LIO rats and CC57 mice. Single doses of 20 mg were given to rats and 1-5 mg--mice. Subcutaneous injections were followed by sarcoma development on injection site in 90% of male rats and 31%--females. Female rats (46%) also revealed multiple benign tumors of the mammary gland. Such tumors alone were detected in females (44%) receiving the agent with feed. All mouse males and 71 100% of females developed hepato- and cholangiocellular tumors as well as hepatic hemangiomas following subcutaneous injection, oral or skin administration. 2,2' diamino-1,1'-dinaphthyl proved a potent carcinogenic agent; it is suggested that it might metabolize to such carcinogen as 2-naphthylamine. PMID- 10532107 TI - [Evaluation of the CCNU-COP regimen for primary refractory and recurrent non Hodgkin's lymphoma]. AB - Thirty patients with non-Hodgkin's lymphoma with tumor progression after failure of standard anthracyclin and alkylating agent treatment (8), and early-onset relapse after the same therapy received 1-4 cycles of CCNU-COP regimen: (22). Complete and partial regression established in 66.6%: complete--4 (13.3%); partial (regression by more than 50%)--16 (53.3%), and stabilisation of disease- 5 (16.7%). Response frequency in patients with low- and moderate-grade tumor was 68%. The most frequent side-effects were leukopenia (60%) and anemia (36.7%) which required special correction in separate cases only. CCNU-COP regimen proved as effective as any other-cost "salvage" ones (ESHAP, MIME, etc.). PMID- 10532108 TI - [Administration of liposomal preparation of DaunoXome for breast cancer in patients with poor prognosis]. AB - A liposomal drug DaunoXome was used as a single agent for stage III-IV breast cancer in 9 patients. Response was recorded in 6; partial regression of tumor--3, and stabilization of disease--in 3 cases. Nausea and vomiting were among the most frequent side-effects; stage I leukopenia was observed in 4; stage II--3, stage III--2 patients. No cardiotoxicity was reported. PMID- 10532109 TI - [Effectiveness of current chemotherapy in metastatic breast cancer resistant to anthracycline antibiotics]. AB - The efficacy of different antitumor therapies was evaluated in a study involving 53 patients with metastatic breast cancer resistant to anthracyclin antibiotics. Complete and partial regression of tumor was recorded following combined administration of vinorelbin and doxorubicin (27.3%) and taxanes (21.3%). Median therapeutic benefit duration was 22 and 28 weeks, respectively. Response frequency for mitomicin C + mitoxantron + methotrexate regimen was 11.1%. However, no objective therapeutic effects were observed in patients on ifosfamide + mitoxantron and regular 5-fluorouracil infusins. PMID- 10532110 TI - [Experience with glyciphonic ointment for malignant and precancerous lesions of the skin]. AB - Skin was smeared with glyciphonic ointment containing 30% of methylphosphonic diglycidyl ether (Tatkhimfarmpreparaty Company) in 495 patients with histologically confirmed basalioma and 36 patients with senile keratosis. During daily application necrotic tissue was removed. Considerable decomposition of tumor occurred on day 6 or later. It took the wound 9-12, seldom, 15 days to heal, with scars forming on days 15-20. Stable cure was registered (5-7 yrs) in 492 patients with skin basalioma and all cases of senile keratosis. No changes in peripheral blood count or any skin-resorption side-effects were recorded. Several patients suffered hyperemia, edema, itching or local pain. PMID- 10532111 TI - [Glyciphonic ointment in the treatment of radiation-induced basal cell carcinoma of the skin]. AB - Skin basalcell carcinoma of the head and neck occurring within 1.5-23 yrs on the site of radiation fibrosis were smeared daily with glyciphonic ointment without dressing the wound. Tumor disintegrated after 2-6 applications followed by edema tumor tissue development on days 22-24. After a 2-3 week interval, perifocal edema subsided, and tumor deed shrank leaving a small scab. Basalioma cells were detected underneath. Complete cure was recorded after another 2-3 week therapy in all cases. Only one patient relapsed three years later. PMID- 10532112 TI - [Management of patients with seminoma, residual after induction chemotherapy, and disseminated in the retroperitoneal space]. PMID- 10532113 TI - [Role of radiotherapy in the combined treatment of uterine choriocarcinoma with brain metastasis]. AB - A retrospective analysis of data on 27 patients with uterine choriocarcinoma metastatic to the brain and other distant sites treated at the Center's Clinic (1980-1996) has been undertaken. Advantages offered by combined treatment (irradiation of the brain + combination chemotherapy) of 15 patients are discussed and practical advice is given. PMID- 10532114 TI - [The use of monoclonal antibodies in the treatment of solid tumors]. PMID- 10532115 TI - [Peptic ulcer with hemorrhage. An analysis of hospital discharges]. AB - BACKGROUND: The gastric and duodenal ulcer is the most common cause of gastrointestinal bleeding in 25% of patients, and accounting annually for approximately 50-100 admissions per 100,000 population. OBJECTIVE: To determine prevalence of gastrointestinal bleeding in hospital admission, according to age and sex and identifying the risk factors. METHOD: Determined the prevalence rates of number of hospital discharges of patients with gastrointestinal bleeding for GU and DU during period 1991 to 1997. Determined the prevalence rates of peptic ulcer occurrence for age-adjusted rates, sex and seasonal variation for 1000 hospital discharges. We investigated the features of hemorrhage of the upper gastrointestinal tract, and identification of risk factors as NSAID, alcohol and tobacco. STATISTICAL ANALYSIS: Chi square and t Student. RESULTS: The annual prevalence rates were 46.6/1000 hospital discharges. There were more frequent between 6th to 8th decades, female sex and during May, June and November. The average stay of patients were 4.2 days (range 1 to 18). A total of 275 patients were found to have GU or DU with gastrointestinal bleeding, 66% were male and mean age was 57 years. The risk factors found were tobacco 52%, alcohol 40% and NSAID 44% (P < 0.05). Melena and hematemesis was found in 64% y 36% respectively. The GU (41%) was more frequent than DU (40%) (P NS). CONCLUSIONS: The annual prevalence rates were 46.6/1000 hospital discharges more frequent in males (66%) and gastric ulcer was found more frequent (41%) than DU (P NS). PMID- 10532116 TI - [Benign esophageal strictures in toddlers and pre-school children. Results of endoscopic dilation]. AB - BACKGROUND: The most frequent causes of dysphagia in children are benign strictures and therefore require special consideration. OBJECTIVE: To evaluate safety and efficacy of endoscopic dilation in children with benign esophageal strictures. MATERIAL-METHODS-RESULTS: Twenty four consecutive children of 1.5 to 5.5 years (mean 3.5), with benign esophageal strictures were evaluated in a prospective manner over a 3-year period. The most frequent causes of esophageal stricture were caustic ingestion (Group A) and in Group B were included other benign strictures. Dilation was done on a weekly base using Savary-Gilliard bougies and was considered adequate, if the esophageal lumen could be dilated to 11 mm with complete relief of dysphagia. Of the 24 patients, 16 had corrosive strictures, 6 complications of gastroesophageal reflux and 2 post surgical strictures. Group A required a significantly higher number of session (14.3 +/- 10.84 vs 7.0 +/- 2.94 p: > 0.05), less free-time dysphagia (1.1 +/- 0.39 vs 2.6 +/- 0.95 months p: < 0.01) and a higher number of recurrences (3.12 +/- 1.12 vs 1.25 +/- 0.95 p: < 0.01). Two esophageal perforations occurred during a total of 292 dilation sessions (0.68%). There was one fatality. CONCLUSIONS: Benign esophageal strictures in children can be treated effectively and with acceptable safety by means of endoscopic dilation. PMID- 10532117 TI - [Esophageal manometry in gastroesophageal reflux disease. Lower esophageal sphincter incompetence or esophageal dismotility?]. AB - BACKGROUND DATA: Hipotensive lower esophageal sphincter (HLES) has been considered the most frequent finding in patients with gastroesophageal reflux disease (GERD). Recently it has been published that esophageal dismotility (ED) has a higher prevalence in GERD. OBJECTIVE: To compare the prevalence of HLES with ED in patients with GERD. MATERIAL AND METHODS: Consecutive patients with endoscopic esophagitis grade II or higher and abnormal esophageal acid exposure time in pH-monitoring were evaluated. Stationary esophageal manometry were performed in all patients. HLES was defined by: a) LES pressure < 10 mmHg, b) LES length < 2 cm and c) LES abdominal segment < 1 cm. ED was defined by: a) presence of more than 30% of peristaltic waves with an amplitude < 30 mmHg or b) more than 10% of simultaneous waves in distal esophagus. RESULTS: Thirty-seven patients, 27 women and 10 men were evaluated. Twelve patients (32.4%) had ED, 5 (13.5%) showed HLES. Four of these 17 patients had both abnormalities. Fifteen patients (40.5%) had normal findings and in 5 (13.5%) a high LES pressure was found. CONCLUSIONS: Esophageal dismotility is the most common manometric abnormality in patients with GERD. PMID- 10532118 TI - [Colonic polyps in children. Experience with polypectomy]. AB - BACKGROUND: Colonic polyp, the most common gastrointestinal tumor in children, is considered a cause of rectal bleeding in the pediatric population. Colonoscopy is the "gold standard" procedure in diagnosis and therapeutic. OBJECTIVE: To know the incidence and symptomatology of colonic polyps in children to remark on the usefulness of the endoscopic examination. PATIENTS-METHODS AND RESULTS: Between 1985 and 1996, over 1,000 colonoscopies were performed on 50 children between 8 months and 14 years old. The patients had colonic polyps and lower gastrointestinal bleeding. In 40 cases polyps were solitary, 82% were located in rectum sigmoid, and 80% of polyps were found to be juvenile (retention). There were no complications associated with diagnostic and therapeutic endoscopy. CONCLUSIONS: The endoscopic method was shown to be very useful for diagnosis as well as treatment of the colonic polyps in children. PMID- 10532119 TI - [Quality of the presentation of free research papers in the National Congress of Gastroenterology: Morelia-1997]. AB - BACKGROUND: The quality of the presentation of a free paper in a medical congress is not necessarily related to the quality of the methodology. OBJECTIVE: To analyze the quality of the presentation of the free papers in the National Congress of Gastroenterology in Mexico (Morelia-1997). METHODS: A prospective study was designed to evaluate the following aspects: Limitation to time assigned, adequate use and design of slides, and mentioning of the main methodologic characteristics. RESULTS: There was a high quality of presentation in the majority of papers. The most frequent problems identified, amenable to improvement, were non-limitation to assigned time (24%), as well as problems in the design of slides (too many lines/columns in 32% and excessive number in 23%). CONCLUSIONS: The knowledge of the results may help to improve the presentations of the free papers in the national congresses of gastroenterology. PMID- 10532120 TI - [Malignant stromal tumor of the transverse colon. Case report]. AB - OBJECTIVE: Case report of a malignant stromal tumor of the transverse colon. BACKGROUND: Colon sarcomas are rare; the most frequent presentation is the leiomyosarcoma. Forty five cases of malignant stromal tumor have been reported in the international literature. The histogenesis of these mesenchymatous neoplasms is determined by ultrastructural analysis and immunohistochemical stains, nevertheless when special techniques are negative and there is uncertainty related to the cellular differentiation line (smooth muscle, neural or undifferentiated) it is preferable to call them stromal tumors of gastrointestinal tract. METHOD: A 46 year old patient with the diagnosis of malignant stromal tumor of transverse colon and the prescribed treatment was reported. RESULTS: A case of a patient with the diagnosis of malignant stromal tumor in transverse colon is presented, who had as the main clinical features abdominal pain, transanal hemorrhage and finally intestinal occlusion. He was submitted to exploratory laparotomy finding a transverse colon intussusception, which was treated with an extended right hemicolectomy and a post surgical satisfactory recovery. Follow-up to three years hasn't found tumoral activity. CONCLUSIONS: Stromal tumors are rare in colon, treatment is a wide surgical resection with curative or palliative purposes. PMID- 10532121 TI - [Anal hamartoma]. PMID- 10532122 TI - [Opening presidential address of the Asociacion Mexicana de Gastroenterolgia]. PMID- 10532123 TI - [Closing presidential address of the Asociacion Mexicana de Gastroenterolgia]. PMID- 10532124 TI - [In Process Citation] PMID- 10532125 TI - [In Process Citation] PMID- 10532126 TI - [In Process Citation] PMID- 10532127 TI - [In Process Citation] PMID- 10532128 TI - [Laparascopic surgical treatment of non-parasitic hepatic cyst]. AB - The experience with seven cases of non-parasitic liver cyst, seen in a seven year period, (between 1992 to 1998) at the "Hospital Espanol" of Veracruz is presented. Clinical symptoms and signs are described, being the main one upper abdominal pain due to the size of the cyst and the compression over different organs. The diagnosis was made by images obtained from ultrasound and CT scan. The cyst average size was 14 cm, and in 28.57% of them, other small cysts were found. The surgical treatment included resection of the superficial capsule of the cyst and the electrocoagulation of the residual inner capsule. The procedure was performed using laparoscopic techniques. There were no surgical incidents, neither post-operatory complications or mortality. The clinical evolution of all patients was successful. We found some evidence from the international literature about the laparoscopic techniques in these patients. Our results were similar to other reports. There are no previous reports in the Mexican literature about this surgical technique in the management of the non-parasitic liver cyst disease. PMID- 10532129 TI - [Abdominal complications after cardiopulmonary procedures]. AB - OBJECTIVE: To know the frequency of intraabdominal complications and its impact on survival of patients submitted to cardiopulmonary bypass for common open-heart surgical procedures. BACKGROUND: The gastrointestinal complications after cardiac surgery with cardiopulmonary bypass (CPB) have an incidence of 0.3 to 3% but mortality can exceed 60%. Despite improvements in preoperative, operative and postoperative care it has been the general impression that abdominal complications remain a significant problem. TYPE OF STUDY: Retrospective case control study. MATERIAL AND METHODS: Consecutive patients submitted to cardiac surgery with CPB between March 1995 to March 1997 were included. Any gastrointestinal complication was identified as well as its diagnosis, medical or surgical management and mortality. RESULTS: One thousand and three hundred fifty two patients were studied of which 516 (38%) were operated for coronary revascularization, 502 (37%) valvular replacement, 68 (5.2%) a combination of valvular replacement and revascularization, 144 (10.6%) correction of congenital defects and 122 (9.6%) treated of diverse problems. Forty-four patients developed complications (3.3%) and they were, postoperative intestinal ileus in 14 cases (32%), half of them had concomitant hyperamylasemia. Hepatobiliary complications represented 29.5% (13 cases). Ten patients (22.7%) developed peptic ulcer disease complicated with perforation or hemorrhage. Severe acute pancreatitis was observed in two patients as well as two with bowel necrosis. Three patients had complications considered not related to CPB as grade I liver trauma, acute appendicitis and amebic colitis. The mortality was 11/44 (25%). As a control group, 73 patients operated upon over the same time period and on the same days as the study patients were analyzed. The mortality in this group was 5/73 (6.8%). The medical history of peptic ulcer disease (< 0.01) and postoperative hemodynamic unstability (< 0.05), the use of intra-aortic balloon pump (< 0.05) and respiratory failure with prolonged ventilatory support (< 0.05) were separate statistical significant determinants for the development of postoperative abdominal complications. CONCLUSIONS: Factors indicative of or contributing to periods of decreased end-organ perfusion appear to be significantly related to abdominal complications. Also, medical history of peptic ulcer disease represented an individual determinant of severe surgical complications as ulcer perforation and massive bleeding. PMID- 10532130 TI - [Microsporidiasis in AIDS patients with chronic diarrhea. Expieriences at the National Institute of Nutrition "Salvador Zubriran"]. AB - BACKGROUND: Microsporidium sp. has been considered as a rare cause of diarrhea in AIDS patients. However, the improvement of some histochemical stains in the analysis of small bowel biopsies has shown an increase in its prevalence. In Mexico there are no series reporting intestinal microsporidiasis. DESIGN: Small bowel biopsies of 98 patients with AIDS and chronic diarrhea stained with HE and Giemsa were reviewed (January 1987-December 1994). The clinical, demographic and laboratory information was obtained from the clinical charts. RESULTS: In 50 patients an opportunistic microorganism was identified in the small bowel biopsy (51%). Microsporidium sp. was identified in 30 patients (31%). The clinical charts were reviewed in all but six cases. Of the 24 patients with microsporidiasis as the cause of diarrhea, 17 were male and seven female with a median age, of 33 years, old. Homosexuality was the main risk factor in males (11/17), and blood transfusion in females (4/7). A low socioeconomical classification was found in 75% cases. The initial manifestation of AIDS was diarrhea in 16/24 (67%), CD4 count cell below 200 mm3 was identified in 13/24 patients and more than 200 mm3 in 2/24. The stool examination and the original histologic interpretations were negative for Microsporidium sp. Lymphoplasmocytic inflammatory infiltrate with eosinophils in the lamina propia and atrophy was frequently seen. A pale red and gray color was observed in spore and merogonial phases of Microsporidium stained with Giemsa. CONCLUSION: Microsporidium sp. was present as the only pathogen in 31% of the small bowel biopsies reviewed by light microscopy. Diarrhea due to Microsporidium sp. is frequently seen in advanced stages of AIDS with CD4 count cell below 200 mm3 Giemsa stain in the evaluation of small biopsies is a cheap and useful method to, identify Microsporidium sp. PMID- 10532131 TI - [Gastric carcinoma in patients under 35 years]. AB - OBJECTIVE: To address the frequency, type of clinical presentation, treatment modalities and survival of gastric carcinoma in young Mexicans. PLACE: Hospital de Especialidades, Centro Medico Nacional Siglo XXI, IMSS, Mexico City. METHODS: A retrospective review of all charts of patients 35 years old or younger, with a diagnosis of gastric carcinoma treated at the department of surgery, from July 1986 to January 1990 was performed. Follow up was conducted at the surgery clinic until death or up to January 1998. RESULTS: Eleven patients under 35 years of age were identified, they represent 13.7% of 80 patients treated during that time period. Women were affected more frequently (1.7:1). Ten patients presented with advanced disease, and only one patient had Stage II. Five gastric resections were performed, one patient had a bypass only, three underwent surgical explorations and two were not operated. Ten had diffuse type lesions and only one intestinal adenocarcinoma. Median survival was 15.3 months and only one patient is alive and well. CONCLUSIONS: In our series, gastric carcinoma is frequent, it is diagnosed in advanced stages, and this is the reason for a very poor prognosis. PMID- 10532132 TI - [Morbidity and mortality in surgery for gastric cancer]. AB - BACKGROUND DATA: Surgery stays as the only effective therapy against gastric cancer. Several factors have been postulated to influence morbidity and mortality risk in gastric cancer surgery. OBJECTIVE: Determine morbidity and mortality of gastric cancer surgery and establish risk factors. METHOD: We reviewed the charts of patients who underwent surgery for gastric adenocarcinoma. Morbidity and mortality is reported. Demographic factors, preoperative physical evaluation, biochemical parameters, surgical technique and tumor biology were analyzed as risk factors for morbidity and mortality. RESULTS: During a seven year period, 120 patients were operated for gastric cancer. Median age was 58.07 years. Subtotal gastrectomy was the most common surgical procedure in 51 patients (42.5%). Morbidity was 26.66% (n = 32). Medical most common complication was renal failure (n = 6, 14.63%) and surgical most common complication was wound infection (n = 7, 17.07%). Mortality was 13.33% (n = 16). Statistically significant risk factors for morbidity were age, ECOG status, Goldman Cardiac Risk Index and a total lymphocyte count. Statistically significant risk factors for mortality were Goldman Cardiac Risk Index, albumin, creatinine, and total lymphocyte count. CONCLUSIONS: Morbidity and mortality after gastric cancer surgery is influenced by preoperative conditions of patients. PMID- 10532133 TI - [Comparative study of the use of sennoside A and B vs polyethylene glycol and electrolytes in anterograde preparation of the colon]. AB - OBJECTIVE: Compare effectiveness of antegrade bowel preparation by liquid diet, polyethylenglycol (PEG) and electrolytes solution plus sennosides A and B, vs. liquid diet plus sennosides A and B only. BACKGROUND DATA: Preparation for colonoscopy with a balanced solution (PEG and electrolytes) has some physiological advantages. Nonetheless, drawbacks of such preparation include nasty flavor, large volumes and low availability in our country. METHOD: A randomized, comparative, prospective, transversal and blind trial that included 200 patients scheduled for colonoscopy were randomly assigned (one hundred each group) to receive: group 1: liquid diet, sennosides A and B and a two litter of solution with PEG and electrolytes; and group 2: liquid diet plus sennosides A and B. Compliance, tolerance and effectiveness of both preparations were evaluated blindly. The results were assessed by Student's T test. RESULTS: The effectiveness of group 2 preparation proved superior (p < 0.05) to group 1. Tolerance and side effects were similar for both groups with no related complications. PMID- 10532134 TI - [Angiomyolipoma of the liver not associated with tuberous sclerosis]. AB - Angiomyolipoma is a rare benign mesenchymal tumor of the liver. We present the case of a 32-year-old female patient that seeks medical consultation to confirm pregnancy. Ultrasound was performed and a hyperechoic lesion was detected in the left lobe of the liver. CT scan showed a heterogenic mass arising from the left lobe of the liver, fine needle aspiration biopsy was performed and diagnosis of liposarcoma was made. A laparotomy was done and a left lateral segmentectomy performed, postoperative course was uneventful. Histology and immunohistochemical analysis of the tumor revealed classical findings of primary angiomyolipoma of the liver. This case shows the difficulty often found when a preoperative diagnosis of fatty liver lesions is made. A literature review is presented and the diagnosis and management of these lesions is discussed. PMID- 10532135 TI - [Duodenal pseudomelanosis]. AB - Pseudomelanosis duodeni is a rare entity characterized by dark pigmentation of duodenal mucosa of uncertain etiology and clinical significance. We report a case with endoscopic and pathologic correlation. Some aspects about etiology, clinical and histopathologic characteristics are discussed. PMID- 10532137 TI - [In Process Citation] PMID- 10532136 TI - [Enteral feeding. Importance in clinical practice]. AB - The gut is the most important organ for the final digestion, absorption and metabolizing of the nutrients ingested. When the gut is available, it must be used as the ideal way to provide nutrients to the body tissues, either orally or by using a feeding tube to help delivering the nutrients. This type of nutrition has important advantages: One of them is the stimulation of the intestinal cells to preserve its integrity and the guts barrier function; it is economic, safe, efficient and it has been associated to improve the patient's outcome. Nowadays, there are many industrialized formulas available for tube feeding use, as well as a great variety of tubes designed for administrating this formulas, depending on the age, size, and time in which the patient will be using them. The future directions for some cases like its use in critically ill patients or the molecular effects of providing nutrients enterally is still under investigation, and there are still questions to be answered in this broad field. PMID- 10532138 TI - [Prognostic factors in 793 cases of gastric cancer in an oncologic referral center]. AB - OBJECTIVE: To describe the 12-year experience with Gastric Cancer (GC), with special emphasis in prognostic factors. BACKGROUND: GC is the most common gastrointestinal malignancy and is the second cause of cancer-related mortality in Mexico. Poor results have been reported, and new treatments have not improved the life expectancy. The available information regarding GC in our country is limited. METHODS: Retrospective cohort study of 793 patients with gastric adenocarcinoma treated in an oncologic referral center in Mexico City. Demographic and clinical data, and the results of surgical treatment are presented. Survival curves by TNM stage and other prognostic factors are described. RESULTS: Sixty two percent of the patients presented in stage IV, with a median survival of 8.6 months. Only 33% of the whole group underwent surgical resection. One hundred and sixty two subtotal, 86 total and 12 proximal gastrectomies were performed, 74% with curative intention and in 26% for palliation. Operative morbidity and mortality were 23.3% and 10.9%, respectively. The multivariate analysis showed that the independent prognostic factors were TNM stage (Risk ratio 1.49; 95% CI 1.26-1.76; p < 0.0001), operative morbidity (RR 6.05; 95% IC 3.74-9.7; p < 0.0001), seralbumin (RR 1.26; 95% CI 1.03-1.5; p < 0.03), age (RR 1.01; 95% CI 0.9-1.02; p < 0.057), type of lymphadenectomy (RR 1.59; 95% CI 0.97-2.59; p < 0.06) and gastrectomy performed (RR 1.9; IC 95% 0.9 4.2; p < 0.06). CONCLUSION: The TNM staging system was the most important prognostic factor. The high rate of GC in advanced stages affects directly the results. Better survival may be expected if the relative frequency of stages I and II increase. Endoscopy is warranted to patients with dispeptic symptoms who present no response to treatment or recurrence. Our experience reflects the importance of this health problem in Mexico. PMID- 10532139 TI - [Effectiveness and safety of mebendazole compared to nitazoxanide in the treatment of Giardia lamblia in children]. AB - OBJECTIVE: Compare the effectiveness and safety of mebendazole versus nitazoxanide in the treatment of Giardia lamblia in children. Giardiasis is an intestinal protozoan of worldwide distribution which most frequently affects the infantile population. In Mexico we have, found a frequency of three to sixty percent. We have, used different medications in it's treatment, but the experience with mebendazole and nitazoxanide is scarce. METHOD: An experimental study as a clinical assay. We included children from the ages of 4 to 12 years old, which had a positive Giardia lamblia cysts in their feces. The children were divided into to two groups: A, were a administered 100 mg of mebendazole every 12 hours, for three days; B, were administered 100 mg of nitazoxanide every 12 hours, for three days; A feces control study was performed at three, five and seven days post treatment. At the end of the treatment we asked the parents if the children had presented any adverse events during the administration of the medication. For the statistical analysis we used Student's t and Chi squared. RESULTS: We studied 82 children, 41 (50%) for each group. In group A, the control feces studies were negative 33 resulting in a 80.4% effectiveness; in group B, 32 were negative resulting in a 78.0% effectiveness, without being statistically significant with a p = 0.8. We found adverse, events in 9 (22%) of the children in group A and 16 (39%) in group B, there was, statistically significant difference with p = 0.09. However, we discovered that the children who received nitazoxanide suffered from abdominal pain more frequently. CONCLUSIONS: We can conclude that both mebendazole and nitazoxanide are efficient for use against infection due to Giardia lamblia, however, the secondary reactions with nitazoxanide were more frequent than with mebendazol. PMID- 10532140 TI - [Hartmann's procedure. Institutional experience with 92 consecutive cases]. AB - BACKGROUND: Since it's description in 1923, Hartmann's procedure is widely used for the surgical treatment of acute left colonic complications when preoperative bowel lavage is not feasible and/or there is high risk of anastomotic dehiscence. OBJECTIVE: Analyze the results of Hartmann's operation in the surgical treatment of consecutive patients at a single institution during a 30-month interval. TYPE OF STUDY: Prospective, non-randomized and longitudinal study. MATERIAL AND METHODS: Patients treated with the Hartmann procedure between March 1995 and September 1998. Surgical indication, intraoperative findings, morbidity and mortality were analyzed as well as the rate of reestablishment of bowel continuity and it's morbimortality. RESULTS: Ninety-two patients underwent a Hartmann procedure. The mean patient's age was 60 +/- 25 years (range of 21 to 88 years) and 60% were older than 65 years. An emergency operation was carried out in 91% of the cases. Most of the patients had intra-abdominal sepsis (56%) and benign colonic process (83%). The morbidity rate was 34% and mortality rate 19. During follow-up the bowel continuity was reestablished in 32% of the cases without fatalities. CONCLUSIONS: Hartmann's procedure is a good option for non elective surgical treatment complicated rectosigmoid pathology. The morbidity and mortality of the operation are highly dependent on the degree of preoperative sepsis and the patient's preexisting condition. The rate of reestablishment of bowel continuity was low probably because of short follow-up. PMID- 10532141 TI - [Percutaneous drainage of amebic hepatic abscess by guided ultraound. Preliminary results]. AB - Amebic hepatic abscess (AHA) is the most frequent extraintestinal complication of amebiasis. Over time, its treatment has gone through some changes and at present is based on amebicides and in some cases, percutaneous drainage. The objective of this work is to present our experience with percutaneous drainage by means of guided ultrasound in patients with AHA. In this work, we include 170 patients admitted to the Gastroenterology Unit of the Hospital General de Zona No. 1, (IMSS) in Mexico City during a period of eight years (1990-1997). These cases included the following criteria: Failure to medical therapy, AHA of liquid matter greater than 5 cm determined by ultrasound; risk of rupture, prolonged incapacity with no data of toxico-infection, accessible drainage route; availability of operating room before risk of complication, and normal coagulation tests. A modified Seldinger's technique was utilized. A single punction was carried out in 131 patients, who had only one abscess. Thirty-nine patients required a second evacuation on presenting two abscesses, and in four cases, a third evacuation was required due to the presence of three or more abscesses. Only one case required an urgent surgical procedure due to abscess rupture to pleura. Five patients suffered complications, including the latter. The remaining four patients had a spontaneous resolution. All patients were released during the 24 hours following surgery, and no patient required hospitalization. For this reason, this can be considered a procedure that in the expert hands of interventionist radiologists, has less morbidity. This work will be carried out to 10 years. PMID- 10532142 TI - [Gastrointestinal stroma tumor with gastric involvement. Presentation of a case and review of the literature]. AB - BACKGROUND DATA: Gastrointestinal stromal tumors (GIST) are considered the most common group of non-epithelial neoplasms of stomach and small bowel. OBJECTIVE: To present a case report and literature review. METHOD: Sixty-seven year-old white male with abdominal mass and gastrointestinal bleeding. Laboratory and X ray test were done resulting in gastric leiomyosarcoma suspect. RESULTS: Patient underwent to exploratory celiotomy with subtotal gastrectomy with splenectomy. Immunohistochemistry results confirms gastrointestinal stromal tumor diagnosis of the stomach. PMID- 10532143 TI - [Adrenal myelolipoma associated with lithiasic cholecystitis, hiatal hernia and esophagitis]. AB - The adrenal myelolipoma are rare tumors that are generally asymptomatic, unilateral and nonfunctional and in the majority of the cases they are found incidentally. OBJECTIVE: Is to present a clinical case of adrenal myelolipoma, associated with gallstones. CASE REPORT: A 30 year old obese female with chronic abdominal pain, which underwent a GI series, having found a hiatal hernia and esophagitis. An ultrasound, of the gallbladder and bile ducts showed gallstones and incidentally a tumor of 9.3 x 8 x 7 cm was found between the right kidney and the liver. In the CT of the abdomen of observed, a little vascularized tumor of the adipose composition of 9.2 x 6 x 5 cm over the top of the right kidney. A cholecystectomy was performed and the resection of the tumor, histopathological study of the tumor reported normal adrenal tissues, mature adiposis and hematopoietic with all of its series. CONCLUSIONS: The association of adrenal myelolipoma and gallstones is not common and it could be an incidental association. With the new tools available we can diagnose the adrenal myelolipoma with a greater degree of certainty. PMID- 10532144 TI - [Bile duct neuroma. A benign neoplasm resembling Klatskin's tumor. Report of a case]. PMID- 10532145 TI - Evolving guidelines for publication of qualitative research studies in psychology and related fields. AB - We present a set of evolving guidelines for reviewing qualitative research, to serve four functions: to contribute to the process of legitimizing qualitative research; to ensure more appropriate and valid scientific reviews of qualitative manuscripts, theses, and dissertations; to encourage better quality control in qualitative research through better self- and other-monitoring; and to encourage further developments in approach and method. Building on a review of existing principles of good practice in qualitative research, we used an iterative process of revision and feedback from colleagues who engage in qualitative research, resulting in a set of seven guidelines common to both qualitative and quantitative research and seven guidelines especially pertinent to qualitative investigations in psychology and related social sciences. The Evolving Guidelines are subject to continuing revision and should not be used in a rigid manner, in order to avoid stifling creativity in this rapidly evolving, rich research tradition. PMID- 10532146 TI - Dependency, self-criticism, interpersonal behaviour and affect: evolutionary perspectives. AB - Evolutionary accounts of vulnerability to depression have focussed either on the attachment system (Bowlby, 1980) or the social rank system (Gilbert, 1992; Price, 1972). According to a two-factor evolutionary model, depression-prone dependent and self-critical individuals suffer from insecurities regarding both attachment and social rank, but they differ in their strategies for dealing with those insecurities. Event-contingent recording was used to assess agentic (dominant submissive) and communal (agreeable-quarrelsome) interpersonal behaviour as well as affect in 119 employed adults over 20 days. Participants also completed questionnaire measures of agency and communion. Self-criticism predicted low levels of agency and low levels of communion. In the sample as a whole, agentic and communal behaviours were associated with pleasant affect, but highly self critical participants experienced relatively less pleasant affect when they acted communally or agentically. Individuals with high levels of immature dependency (neediness) were low in agency, whereas those with high levels of mature dependency (connectedness) were high in communion. Implications for evolutionary theories of vulnerability to depression were discussed, and interpersonal processes that may contribute to vulnerability were identified. PMID- 10532147 TI - PTSD symptoms, response to intrusive memories and coping in ambulance service workers. AB - OBJECTIVES: To examine the relationship of coping strategies and responses to intrusive memories with post-traumatic stress disorder (PTSD) and other psychiatric symptoms in ambulance service workers. METHOD: Fifty-six ambulance service workers describe the most distressing aspects of their work and completed questionnaires designed to measure their coping strategies in dealing with distressing incidents. They also described their intrusive memories of particularly distressing incidents and completed a questionnaire designed to measure their interpretation of these intrusions and their responses to them. In addition, they completed the Post-traumatic Stress Symptom Scale (PSS; Foa, Riggs, Dancu & Rothbaum, 1993) and the General Health Questionnaire (GHQ; Goldberg & Hiller, 1979). RESULTS: Of the participants, 21% met DSM-III-R criteria for PTSD, and 22% met GHQ screening criteria for psychiatric symptoms. Wishful thinking and mental disengagement when confronted with critical incidents at work, negative interpretations of intrusive memories, and maladaptive responses to these memories (rumination, suppression and dissociation) correlated with PTSD severity. CONCLUSION: The results are consistent with the hypothesis that coping strategies and responses to intrusive memories that prevent emotional processing of the distressing event maintain PTSD. They also support Ehlers & Steil's (1995) hypotheses about the role of negative interpretations of post traumatic intrusions in PTSD. A substantial subgroup of emergency service personnel may need support in processing distressing incidents at work and may benefit from information that normalizes post-traumatic symptoms such as intrusions. PMID- 10532148 TI - Attentional bias for emotional faces in generalized anxiety disorder. AB - OBJECTIVES: Recent cognitive theories propose that attentional biases cause or maintain anxiety disorders. This study had several aims: (i) to investigate such biases in generalized anxiety disorder (GAD) using naturalistic, ecologically valid stimuli, namely, emotional facial expressions; (ii) to test the emotionality hypothesis by examining biases for happy as well as threat faces; and (iii) to assess the time course of the attentional bias. DESIGN: The dependent variable was an index of attentional bias derived from manual RTs to probe stimuli. There were four independent variables: one between-subjects variable of group (2: GAD, control), and three within-subjects variables: Type of emotional face (2: threat, happy), Stimulus duration (2: 500 ms, 1250 ms) and Half of task (2: first, second). METHOD: Attentional bias was assessed with a dot probe task. The stimuli comprised photographs of threatening, happy and neutral faces, presented using two exposure durations: 500 ms and 1250 ms. RESULTS: Anxious patients showed greater vigilance for threatening faces relative to neutral faces, compared with normal controls. This effect did not significantly vary as a function of stimulus duration. Anxious patients also showed enhanced vigilance for happy faces, but this was only significant in the second half of the task. CONCLUSIONS: The study confirmed not only that GAD patients show a bias in selective attention to threat, relative to controls, but also that this bias operates for naturalistic, non-verbal stimuli. As the attentional biases for threat and happy faces appeared to develop over a different time frame, different underlying mechanisms may be responsible. PMID- 10532149 TI - Bulimia nervosa: mood changes do have an impact on body width estimation. AB - OBJECTIVES: This study was designed to investigate the impact of mood changes on body width estimation in women with bulimia nervosa. DESIGN: A pre-post controlled experimental design was chosen. METHOD: Mood changes were induced in 40 women with bulimia nervosa, 20 women with panic disorder and 40 women with no diagnosis of a psychological disorder. A combination of autobiographical memory method and music induction method was used to induce positive and negative mood, respectively. Before and after mood induction a video distorting technique was used for body width estimation. RESULTS: Induction of negative mood increased and induction of positive mood decreased the body width estimations of women with bulimia. Patients with panic disorder and 'healthy' controls did not show these changes after mood induction. CONCLUSION: The findings suggest that change in mood state rather than the more habitual mood quality are relevant for bulimic women's body perception. PMID- 10532150 TI - Factor structure of the Matson Evaluation of Social Skills for Individuals with Severe Retardation (MESSIER). AB - OBJECTIVE: A factor analysis of the Matson Evaluation of Social Skills for Individuals with Severe Retardation (MESSIER) was conducted to determine if there was an underlying factor structure which supported a distinction between positive and negative social skills. DESIGN: Principal Axis Factoring with oblimin rotation was used to determine if a two-factor solution was valid. This method was selected to account for shared variance between the items and for correlation between the factors. METHOD: The MESSIER was administered by trained staff to 805 individuals with severe and profound intellectual disability residing in a state residential facility. RESULTS: Results of the factor analysis yielded two categories (positive and negative behaviours) that corresponded to the general division of the clinically derived subscales. CONCLUSIONS: The psychometric research on the MESSIER was extended with an examination of the factor structure. The results of the factor analysis, corresponding with the general division of the clinically derived subscales, are promising. Future research should be conducted to determine if factor scores can be used to determine norms. PMID- 10532151 TI - Coping with anxiety and panic: a factor analytic study. AB - OBJECTIVES: This study aimed to explore the coping styles that agoraphobia sufferers adopt when attempting to cope with symptoms of anxiety and panic. It aimed to extend Watts's (1989) Coping with Anxiety Questionnaire (CAQ) by including items to assess self-vigilance. It was hypothesized that agoraphobia sufferers would adopt consistent coping styles that would be related to symptom severity. DESIGN: A factor analysis was performed on questionnaire data. METHOD: A postal questionnaire was completed by members (N = 112) of a self-help group for agoraphobia and panic sufferers. All participants completed the Beck Anxiety Inventory, an Agoraphobia Severity Scale and a slightly modified version of the CAQ. Coping styles were identified via factor analysis of the CAQ items. RESULTS: Three coping styles were identified, which were labelled Effective Coping, Avoidant Coping and Self-vigilance. The latter two coping styles were found to be correlated with increased levels of agoraphobic symptomatology and with higher levels of anxiety. CONCLUSIONS: The present results support the previous research on coping tactics in anxiety and are compatible with cognitive therapy accounts of the role of self-vigilance in anxiety disorders. PMID- 10532152 TI - The Doors and People Memory Test: validation of norms and some new correction formulae. AB - OBJECTIVES: To validate the normative data on the Doors and People Memory Test (D&P) using a new sample of normal participants, and to investigate the relationship between D&P performance and general intellectual level. DESIGN: 281 normal participants (16-75 years), subdivided into 10-year age bands, were tested on the D&P and the National Adult Reading Test (NART). METHOD: Each participant's raw scores on the D&P were converted into scaled scores, and scaled memory 'indices' were derived using the test manual. Stepwise multi-linear regression was used to predict the indices using age and NART error score as predictor variables. For each participant the discrepancy between the predicted and obtained values of each index was converted into a [symbol: see text] score using the SD of the discrepancies from the whole sample. RESULTS: The distributions of raw and scaled scores on the D&P were similar to those of the original standardization sample. The Visual-Verbal and Recall-Recognition Discrepancy indices had smaller dispersions in the present sample than in the original sample. None of the indices was significantly related to age. The Total Memory, Combined Visual Memory, Combined Verbal Memory, and Overall Forgetting indices were significantly correlated with NART error score. CONCLUSIONS: The present data constitute a cross-validation of the normative data presented in the D&P test manual. Two points of dissimilarity are noted: (i) cutting scores derived for the Visual-Verbal and Recall-Recognition indices based on the test manual norms may be unduly conservative; and (ii) the relationship between some of the D&P indices and NART error score may lead to systematic errors in interpreting the scaled scores derived from the manual. 'Correction formulae' based on the regression equations derived from the present sample are provided. PMID- 10532153 TI - Commentary on Garety & Freeman. I: 'Cognitive approaches to delusions--a critical review of theories and evidence'. PMID- 10532155 TI - Commentary on Garety & Freeman. III: Three psychological investigators and an elephant. PMID- 10532156 TI - Rapid palatal expansion in treatment of Class II malocclusions. AB - A technique which combines the use of rapid maxillary expansion and fixed appliance in growing patients, is presented. The treatment in three patients with Class II division 1 malocclusion and different skeletal patterns is described, and relative advantages highlighted. PMID- 10532157 TI - Breakage incidence with direct-bonded lingual retainers. AB - This study examined the effects of a number of patient and clinical variables on the breakage of bonded retainers, and consisted of a retrospective review of the survival of 200 bonded retainers. Data was collected from two clinical centres between November 1996 and February 1997. The subjects comprised 198 patients of both sexes divided into three age groups. Retainers at both centres were made in 018-inch co-axial wire with Relyabond and Helioprogress adhesives used at each respective centre. The effects on time to first breakage of adhesive, patient sex, and arch (upper/lower) were considered using Kaplan Meier survival graphs and in Log Rank Tests. Finally, a Cox Proportional Hazard Model was used to examine the joint effects of these factors and the patients' ages. Breakage over a 5-year period with Relyabond was 38.8 per cent upper, 22.1 per cent lower, and with Helioprogress 75 per cent upper and 23.2 per cent lower. Breakage appears to be unrelated to the materials used or to the age and sex of the patients. Upper retainers break more often than lowers (P = 0.016) and early breakage is more likely to occur at an adhesive pad than at a wire (9.6 versus 2.5 per cent within 6 months). PMID- 10532158 TI - The heritability of malocclusion: part 2. The influence of genetics in malocclusion. AB - The relative influence of genetics and environmental factors in the aetiology of malocclusion has been a matter for discussion, debate and controversy in the orthodontic literature. This paper reviews the literature and summarises the evidence for the influence of genetics in dental anomalies and malocclusion. Among the conclusions are that, while phenotype is inevitably the result of both genetic and environmental factors, there is irrefutable evidence for a significant genetic influence in many dental and occlusal variables. The influence of genetics however varies according to the trait under consideration and in general remains poorly understood. More precise research tools and methods are required to improve knowledge and understanding, which in turn is a prerequisite to the appreciation of the potential for genetic and/or environmental manipulation in orthodontic therapy. PMID- 10532159 TI - An investigation into the changes in airway dimension and the efficacy of mandibular advancement appliances in subjects with obstructive sleep apnoea. AB - This prospective clinical study evaluates a group of 37 male Caucasians with obstructive sleep apnoea for changes in airway dimension and the efficacy associated with the use of mandibular advancement splints. Lateral skull radiographs were obtained with the subjects--upright in occlusion, supine in occlusion, and supine in protrusion. Each radiograph was traced and digitized, and changes in mandibular position, airway dimensions, and hyoid were examined. Subjects were invited to complete pre- and post-treatment questionnaires, and interviewed following fitting of a removable Herbst mandibular advancement splint. Significant changes were recorded in the airway dimensions in response to both a change in position, from upright to supine, and in response to mandibular advancement. A compliance rate of 76 per cent was achieved with no reported serious complications associated with the use of mandibular advancement devices. PMID- 10532160 TI - A 5-year post-operative review of secondary alveolar bone grafting in the Yorkshire region. AB - The objective of this study was to determine the quality of secondary alveolar bone grafting in the Yorkshire region, and consisted of a retrospective review of patients case notes and radiographs at five surgical units within the Yorkshire region. The subjects were 109 patients who had secondary alveolar bone grafting between 1.9.91. and 31.8.96. The quality of outcome was assessed using a four point radiographic scale from occlusal radiographs taken at least 3 months post operatively: Grade 1 = > 75 per cent bony in-fill, Grade 2 = 50-75 per cent bony in-fill, Grade 3 = < 50 per cent bony in-fill, and Grade 4 = no bony bridge. The radiographic assessment scale was assessed for reliability: inter-examiner weighted kappa = 0.622-0.715 and intra-examiner = 0.818-0.943. Grade 1 results were achieved in 63.2 per cent patients receiving orthodontic expansion and in 40 per cent without expansion before grafting. The four-point radiographic scale described is a useful tool in assessing alveolar bone grafting, Orthodontic expansion. PMID- 10532161 TI - An ex-vivo investigation into the effect of bracket displacement on the resistance to sliding. AB - This ex-vivo study investigated the effect that repeated bracket displacement has on sliding friction and the magnitude of bracket displacement, and hence tooth movement, required to release bracket/archwire binding. The design consisted of an ex-vivo laboratory study. A jig was designed that allowed repeated displacement of a bracket to occur, while the resistance to sliding (friction) was measured using an Instron universal testing machine. One type of stainless steel bracket was used in conjunction with four archwire types (0.016-inch stainless steel, 0.019 x 0.025-inch stainless steel, 0.021 x 0.025-inch stainless steel, 0.019 x 0.025-inch beta-titanium) and four magnitudes of displacement. Repeated bracket displacement has a significant effect on the sliding resistance at the bracket/archwire interface (P < 0.001). The reduction in sliding resistance noted with displacement depended on the archwire. Over the range of displacements tested, there was an 85 and 80 per cent reduction associated with 0.021 x 0.025-inch and 0.019 x 0.025-inch stainless steel, respectively. For 0.019 x 0.025-inch beta-titanium and 0.016-inch stainless steel, these reductions were 27 and 19 per cent, respectively. The importance of true friction, given the likelihood of bracket and/or archwire displacements in vivo, may be lessened. PMID- 10532162 TI - The effectiveness and efficiency of hygienists in carrying out orthodontic auxiliary procedures. AB - The aim of this study was to compare the ability and efficiency of dental hygienists, after preliminary training as orthodontic auxiliaries, with post graduate orthodontists. The study was cross-sectional and prospective. The sample consisted of five second-year hygienists and five qualified orthodontists from Manchester University Dental Hospital. All subjects carried out a range of orthodontic exercises on phantom head typodonts. The ability and efficiency for each task was measured, and comparison made between hygienists and orthodontic groups. There was no statistically significant differences between hygienists and orthodontists in terms of their ability to carry out potential orthodontic auxiliary procedures. However, orthodontists were more efficient (P < 0.05). The ability of hygienists to carry out potential orthodontic auxiliary tasks after appropriate training is supported. Trained orthodontists are more efficient than newly trained hygienists in carrying out potential orthodontic auxiliary tasks. PMID- 10532163 TI - The controversy over how to present research findings. PMID- 10532164 TI - How do current Senior Registrar job profiles relate to proposed Specialist Registrar FTTA posts? Fixed-term training appointments. AB - The proposed United Kingdom training pathway for Orthodontic Specialist Registrars is now accepted to be of 3 years duration. In the final year, Specialist Registrars will take the Membership in Orthodontics, with the end point of training marked by the award of the Certificate of Completion on Specialist Training (CCST). There will be a predetermined number of fixed-term training appointments (FTTAs), available through competitive entry, which will provide 2 years of additional training and lead to eligibility to apply for a Consultant appointment. The end point of the Specialist Registrar (FTTA) will be marked by the Intercollegiate Specialty Examination (ISE). The current 3-year Senior Registrar orthodontic training will be reduced to 2 years as the transition to the Specialist Registrar FTTA grade occurs. In the light of these changes, a survey of full time NHS Senior Registrar posts was carried out to examine current job profiles with particular reference to their suitability for assimilation into the Specialist Registrar (FTTA) grade and preparation for the ISE. PMID- 10532165 TI - Fluoride mouthrinses. PMID- 10532166 TI - Re: Statistical significance testing. PMID- 10532167 TI - Rachel's story. PMID- 10532168 TI - Re: Modification of a bite registration device. PMID- 10532169 TI - New contracts for specialist orthodontic practitioners? AB - This paper discusses the possibility of new forms of contacting or commissioning emerging between UK Health Authorities (or other parties such as Primary Care Groups and Primary Care Trusts) and established providers of specialist orthodontic services. PMID- 10532170 TI - Expanding the phenotype of Filippi syndrome: a report of three cases. AB - This report is of two brothers and a male singleton with clinical characteristics of Filippi syndrome, born to young, healthy, non-consanguineous parents. Their features, which include borderline to milder developmental delay, particularly of speech and language, primary microdontia and previously unreported radiological findings are described to further delineate and expand the clinical spectrum of the condition. PMID- 10532171 TI - A second family with Micro syndrome. AB - We present the cases of two sisters, daughters of healthy, non-consanguineous parents, who have a clinical syndrome characterized by microcephaly, cortical dysplasia, ventriculomegaly, hypoplasia of the corpus callosum, hypogenesis of the cerebellar vermis, cataracts, microphthalmia, optic nerve atrophy, retinal coloboma, weight and height below 3rd centile, severe mental retardation, no speech, inability to sit, no sphincter control and a spastic tetraparesis. The facies are mildly dysmorphic, but not distinctive. No metabolic, nor chromosomal anomalies were found. The cases are very similar to, but not identical, to those described by Warburg et al [Am J Med Genet (1993) 147:1309-1312] as Micro syndrome. PMID- 10532172 TI - Familial association of camptodactyly, mental retardation, whistling face and Pierre Robin sequence. AB - Two sibs are reported with severe developmental retardation combined with the clinical triad of camptodactyly, whistling face and Pierre Robin sequence as clinical signs of fetal hypokinesia. In spite of tracheotomy, the first child of the family died 10 hours after birth. A sister of this child was born 1 year later. During pregnancy prenatal diagnosis of hydrocephaly was made by ultrasonographic examination. MRI scan showed holoprosencephaly. At 15 months of age psychomotor development is severely impaired, birth and length are delayed. PMID- 10532174 TI - Jeune syndrome (asphyxiating thoracic dystrophy) associated with Hirschsprung disease. AB - We describe two children with diagnostic features of Jeune syndrome who also had Hirschsprung disease. An association between the two conditions has not previously been described and has implications both for clinical management and for further study. PMID- 10532173 TI - Recurrence of the severe form of microgastria-limb reduction defect in a consanguineous family. AB - This report concerns two sibs from a consanguineous Sudanese family with microgastria-limb reduction defect associated with hydrocephalus and agenesis of corpus callosum. We suggest that these cases together with other previously reported cases of central nervous system (CNS) anomalies associated with microgastria-limb reduction defect could represent an autosomal recessive syndrome differing from the classical microgastria-limb reduction defect by its severity, presence of CNS anomalies and its pattern of inheritance. PMID- 10532175 TI - Non-ossifying fibromas and giant cell reparative granulomas in a child with ocular-ectodermal syndrome. AB - We report on a patient with ocular-ectodermal syndrome who was previously described in 1993 [Am J Med Genet (1993) 45:764-766]. This boy has now developed additional manifestations, including giant cell granulomas and non-ossifying fibromas. This adds to the list of phenotypic manifestations of this condition. PMID- 10532176 TI - Aortic dissection, patent ductus arteriosus, iris hypoplasia and brachytelephalangy in a male adolescent. AB - We describe a 14-year-old male with dissection of the descending aorta, bilateral iris hypoplasia, striae distensae and brachytelephalangy, the latter being most marked in the thumbs. Inguinal herniae and a patent ductus arteriosus were surgically repaired in infancy. The pattern of abnormalities may constitute a previously undescribed syndrome. The proband died suddenly at the age of 17 years. PMID- 10532177 TI - Disorganisation: a case with popliteal pterygia and placental-skin appendages. AB - We report a girl with congenital anomalies which include amniotic rings and scars, cleft lip and palate, thumb abnormalities, hexadactyly of feet, severe flexion deformities of legs and unusual finger-like appendages which were attached to the placenta. We suggest this patient represents another example of human homologue for the mouse mutant disorganisation (Ds). PMID- 10532178 TI - A child with hemimegalencephaly, hemihypertrophy, macrocephaly, cutaneous vascular malformation, psychomotor retardation and intestinal lymphangiectasia--a diagnostic dilemma. AB - Although the clinical delineation of the majority of overgrowth syndromes is straightforward, we believe there is a subset of patients with overlapping features from a number of overgrowth syndromes. We report a patient with hemimegalencephaly, hemihypertrophy, macrocephaly, vascular lesions, psychomotor retardation and intestinal lymphangiectasia. The clinical history and findings posed a diagnostic dilemma as the features overlapped between several conditions, namely macrocephaly-cutis marmorata telangiectatica congenita (M-CMTC), Klippel Trenaunay-Weber syndrome (KTWS), Proteus syndrome and a provisional unique syndrome described by Reardon et al. (1996, Am J Med Genet 66:144-149). We anticipate that only when the molecular basis is delineated will it become clear whether these disorders are separate entities or merely differing ends of the same spectrum. PMID- 10532179 TI - A girl with ectodermal dysplasia, choanal atresia and polysyndactyly. AB - We present a 3-year-old child with clinical features of ectodermal dysplasia comprising sparse hair, dystrophic and ridged nails and bilateral obstruction of the nasolacrimal ducts. Additional findings were unilateral choanal atresia, bilateral syndactyly of the feet and polydactyly. We discuss the differential diagnosis of these clinical findings. PMID- 10532180 TI - Bilateral hydronephrosis due to megacalicosis as a prenatal sonographic finding in a female with Schinzel-Giedion syndrome. AB - A case of Schinzel-Giedion syndrome, a rare malformation syndrome with a life expectancy of less than 2 years is described. Features present in this and previous cases are discussed. The association of agenesis of the corpus callosum with the Arnold-Chiari malformation found in this patient has not previously been described. PMID- 10532181 TI - Another case of achalasia-microcephaly syndrome. AB - The first German patient with achalasia-microcephaly syndrome is described. The mother was exposed to the anti-malarial drug Mefloquine during the first 8 weeks of pregnancy. PMID- 10532182 TI - On the expansion of the phenotype of the acro-renal-ocular syndrome. PMID- 10532183 TI - Syndrome of psychomotor retardation, bulbous nose, and epilepsy (Hernandez syndrome): a Brazilian case. AB - A new case of Hernandez syndrome is described in a 16-year-old Brazilian girl. The syndrome consists mainly of psychomotor retardation, epilepsy, a bulbous nose and obesity. PMID- 10532184 TI - Does the cranio-cerebello-cardiac syndrome (3C syndrome) include abdominal malformations? AB - We report two children of nonconsanguineous parents, with hypotonia, severe psychomotor retardation, short stature, a prominent forehead, ptosis, a wide flat nasal bridge, broad nasal tip, a high arched palate, bilateral small cerebellar hemispheres, vermis hypoplasia, a large cisterna magna, and an atrial septal defect and a duodenal stenosis in one case. These features are part of the 3C syndrome. The duodenal stenosis present in one of our sibs has not been reported before in this syndrome. PMID- 10532185 TI - Laparoscopic resection of colonic neoplasms: current status. PMID- 10532186 TI - Minimally invasive surgery in urology. PMID- 10532187 TI - Laparoscopy in gynaecologic oncology: a review. PMID- 10532188 TI - Pro and contra in minimally invasive oncological surgery. Minimally invasive surgery in lung cancer. PMID- 10532189 TI - Who is in control of the immune system in head and neck cancer? PMID- 10532190 TI - Issues in the management of nasopharyngeal carcinoma. PMID- 10532191 TI - Hepatitis B virus infection and bone marrow transplantation. PMID- 10532192 TI - Present achievements in the medical treatment of metastatic renal cell carcinoma. PMID- 10532193 TI - The biology and management of bone metastases. PMID- 10532194 TI - Suppression of p53-mediated growth factor withdrawal-induced apoptosis in the myeloid compartment by hematopoietic cytokines: an overview of hematopoiesis and apoptosis with a presentation of thrombopoietin and the M07E cell line as a model system. AB - The resistance of leukemic cells to apoptosis induction is one of the great obstacles in clinically alleviating the neoplastic state which they create. Even though p53 is the most commonly mutated protein found in tumors, there does exist the remaining body of cancers that possess wild-type p53. Within the hematopoietic compartment, the majority of acute myelogenous leukemia isolates fall into this latter category ([174, 175,375-377], for example). In particular, it is found that in most AML isolates p53 is genotypically wild-type but found to have an immunophenotype typical of mutant p53, or in the promoter conformation [170, 174,175,377]. The clinical significance of the research which aims to discern the molecular machinery involved in p53 conformational modulation must certainly include the notion that a therapeutic modality may be developed which interferes with the molecular process of shifting p53 from a viability compromising suppressor conformation to the survival-enhancing promoter conformation. Further studies that investigate the interface between cytokine signaling transduction cascades and the subsystem(s) regulating the redox state of p53 are critical to advancing this field. PMID- 10532195 TI - A critical review of the management of bladder cancer. PMID- 10532196 TI - Pathological prognostic factors in breast cancer. PMID- 10532197 TI - The crucial role of adjuvant irradiation after surgery for breast cancer. PMID- 10532198 TI - Intraperitoneal chemotherapy. PMID- 10532200 TI - Phase-transition polymers for drug delivery. AB - Phase-transition polymers show changes in response to external stimuli, such as pH, temperature, light, metabolite, and electric current. Based on the stimuli induced phase transition, many applications have been developed to improve drug delivery. This paper summarizes various phase-transition polymers and their applications relevant to modulated-drug delivery. PMID- 10532199 TI - Transdermal and transmucosal powdered drug delivery. AB - High-velocity powder injection is a promising new drug-delivery technique that provides needle- and pain-free delivery of traditional drugs, drugs from biotechnology such as proteins, peptides, and oligonucleotides as well as traditional and genetic vaccines. The energy of a transient helium gas jet accelerates fine drug particles of 20 microns-100 microns diameter to high velocities and delivers them into skin or mucosal sites. This review describes the configuration and operating principles of devices that accelerate the particles, the required properties of the particles, the characteristics of the target tissues, and features of the developmental test methods. Preclinical and clinical results that best characterize the technology and introduce its potential as a drug-delivery platform are presented. PMID- 10532201 TI - Incidence of postinfarction aneurysm within one month of infarct. Experiences with sixteen patients in Hawaii. AB - BACKGROUND: We report on sixteen patients with a left ventricular aneurysm presenting at less than a month following myocardial infarction. METHODS: All patients had significant left anterior descending coronary artery disease, and in eight cases (50%), this was the only significant pathology. Two patients who were treated conservatively, died within three months of infarction. RESULTS: Of the fourteen surgically treated patients, one died. There have been two late deaths, one at ten months and the other at four years postinfarction. Patients who present early after infarction, usually have a large anterior aneurysm, requiring early surgical repair with ventricular aneurysmectomy and revascularization. This group of patients showed a higher risk for major complications (such as thrombo embolism, arrhythmias) and/or death. Emergency coronary artery bypass surgery may prove beneficial in the prevention of aneurysm formation by revascularizing the viable but ischemic tissue in that area. PMID- 10532202 TI - Significance of the human immunodeficiency virus infection in patients submitted to cardiac surgery. AB - BACKGROUND: To realize if cardiac surgery could interfere with the evolution of HIV infected patients to the acquired immunodeficiency syndrome (AIDS). METHODS: The study group consisted of 30 HIV positive patients (0.21%) among 14,785 who underwent cardiac surgery at the Heart Institute of University of Sao Paulo Medical School (Incor-FMUSP) from November 1988 to December 1994. Patients were followed up until they were discharged from hospital and a new contact was kept at the end of the first semester of 1995. RESULTS: All patients were asymptomatic at the time they were operated. Two patients progressed to death during hospitalization due to non-infectious complications and other three patients could not be traced. After all 25 patients had their progression evaluated. Six patients (24%) died within a period ranging from 1 to 46 months (average=17 months): 2 due to bacterial pneumonia and 04 due to AIDS-related complications. The average follow-up period for the 19 surviving patients was 33.6 months (ranging from 13 to 74 months), and only one of them (5.3%) saw the infection progress to AIDS. In summary, 5/25 (20%) saw HIV infection progress to AIDS within a maximum period of 74 months. CONCLUSIONS: Data available up to now show no conclusive evidence of acceleration of HIV into AIDS associated with cardiac surgery. PMID- 10532203 TI - Effects of heat stress on metabolism of high-energy phosphates. Comparison of normothermic and hypothermic ischemia. AB - BACKGROUND: Alterations in metabolic pathways may contribute to the cardioprotective effects of heat stress (HS). We investigated the effects of HS on ATP and phosphocreatine (PCr) levels in the ischemic rat myocardium, after both normothermic and hypothermic ischemia. METHODS: Two protocols were used: (1) normothermic ischemia (20 min at 37 degrees C) with no myocardial protection (n=6 HS; n=6 control); (2) hypothermic ischemia (4 hrs at 4 degrees C) after cardioplegic arrest (n=6 HS; n=6 control). ATP and PCr levels in the heart were measured using 31P nuclear magnetic resonance spectroscopy. RESULTS: At the end of normothermic ischemia, ATP levels were better maintained in HS hearts (C vs HS: 4.51+/-0.66 vs 7.81+/-1.06 micromol/g dry wt+/-SEM, p=0.04). A trend for higher ATP content in HS hearts was observed after 40 min of reperfusion (C vs HS: 11.7+/-1.5 vs 16.9+/-2.0 micromol/g dry wt+/-SEM, p=0.09). PCr content was also higher at the end of 40 minutes of reperfusion in HS hearts (C vs HS: 46.4+/ 2.9 vs 56.9+/-3.0 micromol/g dry wt+/-SEM, p=0.03). After prolonged hypothermic ischemia under cardioplegic arrest, heat stress again led to better preservation of ATP levels at the end of ischemia (C vs HS: 5.71+/-0.88 vs 9.23+/-1.38 micromol/g dry wt+/-SEM, p=0.05) and after 40 minutes of reperfusion (C vs HS: 16.8+/-1.4 vs 24.6+/-2.8 micromol/g dry wt+/-SEM, p=0.03). PCr levels were also better maintained at the end of ischemia (C vs HS: 4.87+/-0.77 vs 12.4+/-3.0 micromol/g dry wt+/-SEM, p=0.03) and after 40 minutes of reperfusion in HS hearts (C vs HS: 55.1+/-7.0.vs 79.8+/-7.3 micromol/g dry wt+/-SEM, p=0.03). CONCLUSIONS: Heat stress induces changes in the energy profile of the heart which results in better preservation of ATP and phosphocreatine levels. These changes could be observed after brief normothermic ischemia and also after prolonged hypothermic ischemia under cardioplegic arrest, mimicking conditions of preservation for cardiac transplantation. PMID- 10532204 TI - Extracorporeal circulation does not induce intra-alveolar release of Endothelin 1, but only a modest overproduction in pulmonary circulation. AB - OBJECTIVE: To investigate whether ECC may produce regional liberation of inflammatory mediators capable of inducing vascular effects and organ damage. EXPERIMENTAL DESIGN: Comparative study [corrected]. SETTING: Cardiac surgery department in a University hospital. PATIENTS: Fifteen patients undergoing coronary artery bypass grafting (CABG, group A) and ten patients operated for infrarenal abdominal aortic aneurysm (controls, group B) have been studied. MEASURES: Levels of Interleukin 1beta (IL1), Tumor Necrosis Factor alpha (TNF), Interleukin 6 (IL6), and Endothelin 1 (ET1) were measured in pulmonary capillary, arterial, and venous blood and in bronchoalveolar lavages (BAL) before, during and after extracorporeal circulation (ECC) or surgical intervention. RESULTS: TNF alpha (never >35 pg/ml) and IL1beta (range 20-300 pg/ml) values did not change over time for both groups. IL6 concentrations in all samples of group A increased between five and twenty fold, during and after ECC (from 3-5 pg/ml up to 240 pg/ml, p<0.001). This trend was similar in controls after surgical stress. Endothelin 1 was always undetectable in the BAL fluid, with a modest, but significant increase in pulmonary capillary blood of group A, after ECC, (from 11+/-4 pg/ml to 18+/-5 pg/ml, p<0.001). This increment correlated well with the PVR increase, but was transient and after 24 hours, ET1 values returned to baseline levels. Mean values of ET1 increased also in controls, but not significantly. CONCLUSIONS: ECC may induce ET1 liberation in pulmonary circulation with transient pulmonary vasoconstriction, but wihout intra-alveolar release, or lung damage. Augmented concentrations of IL6 probably express a response to surgical procedure rather than an effect exclusively related to ECC. PMID- 10532205 TI - ATP-MgCl2 utilization for spinal cord protection during experimental thoracic aortic occlusion. AB - BACKGROUND: In this experimental study we investigate the effect of intravenous ATP-MgCl**2 administration for prevention of spinal cord injury occurring due to ischemia induced by aortic cross clamping. METHODS: Ten rabbits were studied. The abdominal aorta is ligated below the left renal artery. Five rabbits served as a control group and received no medication during 30 minutes of ischemic period. The other 5 rabbits received during 30 minutes of aortic occlusion ATP-MgCl2 solution (100 micromol/ml for each). Distal and proximal aortic pressures are measured during the procedure and incisions are closed. Rabbits are observed for 24 hours for their neurological status and scored accordingly. Specimens from the spinal cord are taken for electron microscopic investigations. RESULTS: All of the control group rabbits were paraplegic. One of the ATP-MgCl2 group rabbits was paraparesic and the others were normal Distal aortic pressure was 9+/-3 mmHg for the control group and was 17+/-4 mmHg for the ATP-MgCl2 group (p<0.05). Electron microscopic studies showed the preserved ultrastructure for ATP-MgCl2 group. CONCLUSIONS: ATP-MgCl2 administration during spinal cord ischemia reduces spinal cord injury. This may be an alternative modality for the protection of the spinal cord during aortic surgery. PMID- 10532206 TI - Diagnostic value of adenosine deaminase activity in pericardial fluids. AB - BACKGROUND: The activity of adenosine deaminase (ADA) was determined in serum and pericardial fluid of 70 patients (ages 21 to 71 years) with pericardial effusions of various etiologies and in 15 control subjects. METHODS: The patients were subdivided into five groups on the basis of definite diagnosis: 1) 24 patients with tuberculosis; 2) 22 with malignancies; 3) 12 with uremic pericarditis; 4) 12 with purulent pericarditis; 5) 15 control individuals without pericardial disease. The activity of ADA was determined at the same time in serum and cell free pericardial fluid according to the method of Karker with minor modification. RESULTS: Mean (+/-SD) ADA activity in pericardial fluid was 66.92+/-4.12 IU/L in group 1; 27.50+/-6.02 in group 2; 28.65+/-4.73 in group 3; 53.05+/-11.14 in group 4; and 5.67+/-1.99 in group 5. Comparing the level achieved in group 1 with all others, the difference is significant at the p<0.001 level. When the cut-off value of 50 IU/L is used the sensitivity of the test for diagnosis of tuberculous effusion is 1, and the specificity is 0.83. Statistical analysis showed that there was no correlation between serum ADA activity and ADA activity in pericardial fluid. CONCLUSIONS: We recommend that determinations of ADA activity in pathologic pericardial fluids seem to be of great value in the early diagnosis of tuberculous pericardial effusions. Levels above 50 IU/L in effusions indicate probable tuberculosis. PMID- 10532207 TI - Inhibitors of the platelet receptor glycoprotein IIb-IIIa and complications during percutaneous coronary revascularization. Management strategies for the cardiac surgeon. AB - Platelet-mediated thrombosis has a pivotal role in the pathophysiology of acute ischemic coronary syndromes (AICS) and the acute complications of percutaneous coronary interventions. Because cross-linking of the activated platelet receptor glycoprotein (GP) IIb-IIIa by plasma fibrinogen represents the final common pathway to coronary thrombus formation, several GP IIb-IIIa inhibitors have been developed as a potentially more effective antithrombotic therapy than agents currently used for this purpose, namely aspirin and heparin. However, use of GP IIb-IIIa inhibitors in patients with AICS and those scheduled for percutaneous coronary interventions may increase the risk of serious clinical and bleeding events among patients who require emergency or urgent bypass surgery. This review describes clinical experience with various GP IIb-IIIa inhibitors and suggests management strategies for patients undergoing emergency or urgent bypass surgery shortly after treatment with GP IIb-IIIa inhibitors. PMID- 10532208 TI - Midterm results of the neonatal arterial switch operation. A review. AB - Complete transposition of the great arteries is a relatively common anomaly, which comprises 5 to 7% of all instances of cardiac malformations. Given the decreasing mortality rates associated with a neonatal arterial switch operation and the unacceptable morbidity associated with atrial baffle operations, it is reasonable to accept this operation as the procedure of choice for the treatment of the complete transposition of the great arteries. It represents a major improvement since it does not introduce any additional intracardiac anomaly, restores the left ventricle to its natural systemic function, and maintains the sinus node function. Long-term survival exceeds 90%. Midterm general health status is encouraging. To date, there have been limited long-term follow-up studies after a neonatal arterial switch operation, although the intermediate data are promising. It will be necessary to determine whether the theorized benefits of the anatomic repair are realized, since experience is limited to the last 15 years. Translocation of the coronary arteries remains one of the most difficult aspects of the operation and late mortality appears to coincide with coronary artery events with sudden death secondary to acute myocardial infarction being reported in 1-2% of hospital survivors. Supravalvar pulmonary stenosis, neoaortic root dilation and valvar regurgitation, bronchopulmonary collateral arteries, coronary insufficiency, and myocardial perfusion abnormalities are other specific areas which require close observation and further investigation. PMID- 10532209 TI - Left internal mammary artery to innominate vein fistula complicating pacemaker insertion. Treatment with endovascular transarterial coil embolization. AB - Arteriovenous fistula (AVF) is rarely encountered as a complication of pacemaker insertion. Percutaneous angiographic therapy of such iatrogenic fistulas can be both safe and effective, leading to important reductions in costs. A 60-year-old woman was admitted to the hospital four weeks after left subclavian pacemaker insertion complaining of signs of congestive heart failure. A loud continuous machinery bruit was heard over the left upper chest. An arteriogram revealed a false aneurysm from the LIMA, 6 mm in-diameter, with formation of an AVF between the LIMA and the left innominate vein. Embolization of the LIMA was carried out using seven Platinum coils at the level of the AVF and the false aneurysm was embolized with 3 controlled-release IDC coils. The complete occlusion of the fistula was achieved and the distal LIMA persisted patent due to the opening of collateral vessels from the intercostal arteries. AVF between the subclavian artery or its branches and the subclavian or innominate veins have been reported to be congenital, traumatic and iatrogenic (associated to central venous access to hemodynamic monitoring, dialysis, and very infrequently to pacemaker insertion) but the internal mammary arteries are only rarely involved. The course of AVF is undefined, but generally, surgical or percutaneous embolization is warranted because of the potential appearance of a great number of complications. Surgical repair is associated with significant morbidity and mortality. Whenever possible, percutaneous nonsurgical occlusion of the AVF with coil embolization is the procedure of choice, because of its high success rate and low morbidity. PMID- 10532210 TI - Isolated tricuspid regurgitation due to atypical morphology of anterior-posterior leaflets in an adult: a case report and review of the literature. AB - A 73-year-old woman with congenital isolated organic tricuspid regurgitation was reported. She had neither the history of chest trauma nor rheumatic fever nor the evidence of infective endocarditis. The patient was successfully treated with a bioprosthetic valve replacement in tricuspid position. Operative findings revealed hypoplastic anterior leaflet and relatively large posterior leaflet. Structural anomaly of the valve, coaptation disorder due to the thickened valve leaflets, as well as enlarged valve ring and the occurrence of atrial fibrillation was thought to be the causes of massive regurgitation. PMID- 10532211 TI - Spontaneous dissection of coronary artery in a patient with ascending aortic aneurysm and aortic valve regurgitation. AB - Spontaneous coronary artery dissection is a rare cause of myocardial infarction associated with a significant high morbidity and mortality. It usually occurs in relatively young patients and it is frequently found at autopsy. We report a case of a 42-year-old woman, who underwent resection of subaortic diaphragm ten years earlier presenting with postero-lateral myocardial infarction. Coronary arteriography revealed a dissection of the left main stem extending distally to the left anterior descending artery (LAD) and circumflex artery (Cx); occlusion of the postero-lateral branch of the Cx; severe aortic valve regurgitation and ascending aortic aneurysm. She was successfully operated on in emergency and underwent myocardial revascularization and separate replacement of the aortic valve and the ascending aorta. In this specific case of coronary dissection and severe aortic regurgitation it is mandatory to perform surgery in emergency to limit infarction evolution and avert loss of life. PMID- 10532212 TI - Surgical management of arteriosclerotic coronary artery aneurysm. AB - A 60-year-old man suffered antero-septal myocardial infarction at the age of 56. Coronary angiography demonstrated total occlusion of the left anterior descending artery and a large saccular aneurysm of the right coronary artery. Diffuse coronary ectasia was also shown in the right coronary artery adjacent to the aneurysm. Despite anticoagulant therapy, the aneurysm formed a thrombus and developed coronary artery stenosis distal to the aneurysm. Ligation of the aneurysm and in situ gastroepiploic artery grafting were performed. Sudden heart failure was developed during skin closure. As this condition was considered to be graft hypoperfusion, supplemental saphenous vein grafting was placed. Ligation is a simple, reliable technique to prevent future complications for a large saccular right coronary artery aneurysm, however, gastroepiploic artery might be an inappropriate bypass conduit for the ligated coronary artery with diffuse ectasia. PMID- 10532213 TI - Posterior-septal pseudo-pseudoaneurysm with limited left-to-right shunt: an unexpected easy repair. AB - Cardiac rupture represents a fatal complication of acute myocardial infarction within the first two weeks. In exceptional cases, the postinfarction rupture of the myocardium is not transmural but remains circumscribed within the wall itself as a cavity joined to the left ventricle through a narrow neck. This finding is usually defined as pseudo-pseudoaneurysm. We report a rare case of postinfarction posterior pseudo-pseudoaneurysm of the left ventricle, perforated into the right ventricle. This unusual anatomy resulted, over a period of several years, by progressive intramural dissection of the surrounding necrotic myocardium with late formation of a large, partially fibrotic chamber, communicating either with left and right ventricles. Despite correct preoperative diagnosis was not achieved by 2D echocardiography, pulsed Doppler and contrast ventriculography, a successful surgical treatment was possible with a really good outcome. PMID- 10532214 TI - Extracardiac total cavopulmonary connection via an atrial and pericardial tunnel. AB - We performed extracardiac total cavopulmonary connection using only native tissue in two patients with complex heart disease. The extracardiac lateral tunnel was constructed from pedicled autologous pericardium and atrial wall During follow-up both patients remained in the New York Heart Association functional class I and demonstrated normal sinus rhythm. These modifications not only avoid the risk of potential postoperative complications due to artificial materials, but also allow growth of the conduit. PMID- 10532215 TI - Repair of intramural hematoma of the ascending aorta without graft interposition. AB - A 68-year-old woman was admitted to hospital with a one-hour history of chest pain and syncopal episode. Transesophageal echocardiography showed an intramural aortic hematoma with cardiac tamponade. The patient underwent repair of the ascending aorta without graft interposition (resection and end-to-end anastomosis). The patient had an uneventful postoperative course and the 38-month follow-up was event-free. This case report shows that end-to-end anastomosis in patients with intramural hematoma and absence of intimal tearing, may provide good long-term results. PMID- 10532216 TI - Redo cardiac surgery for atrial septal defect in a patient with idiopathic thrombocytopenic purpura. Preoperative management with high-dose intravenous gamma-globulin. AB - A 58-year-old woman with idiopathic thrombocytopenic purpura required redo cardiac surgery of atrial septal defect closure, mitral annuloplasty, and tricuspid annuloplasty. Preoperative high-dose intravenous gamma-globulin and platelet transfusion after termination of cardiopulmonary bypass allowed successful redo cardiac surgery. Management of a patient with idiopathic thrombocytopenic purpura undergoing open heart surgery are discussed. PMID- 10532217 TI - Internal mammary artery grafts and competitive flow. Controversies persist. AB - We report one case in which chronic native competitive flow from an almost normal target coronary artery did not influence IMA graft patency. This patient underwent control postoperative angiography 11 months after surgery and the mammary artery-left anterior descending graft was found to be normofunctioning despite the fact that the coronary artery showed no residual stenosis. PMID- 10532218 TI - Selective patching in carotid endarterectomy: is patching always necessary? AB - BACKGROUND: The value of carotid patching in carotid endarterectomy in achieving low perioperative morbidity and long-term freedom from restenosis is controversial. We hypothesized that if large internal carotid arteries were closed primarily and smaller arteries selectively patched, there would be no difference in early or long-term results between the two groups. METHODS: A retrospective analysis of 133 carotid endarterectomies performed by one surgeon in a community teaching hospital was performed to evaluate a selective approach to patching vs primary closure. Primary closure was performed if the arteriotomy could be closed without tension over a Javid shunt. Seventy-seven arteries underwent primary closure and 56 underwent patching (Vein-14, PTFE-17, Dacron 25). Postoperative (>6 month) duplex scans were available on 46/77 (60%) patients undergoing primary closure, and 33/56 (59%) of patients with patch repair. RESULTS: There were 2 perioperative neurologic deficits, both in the patch group. Restenosis of equal or greater than 50% at 11 months occurred in 5/46 (10.8%) of patients with primary closure and 2/34 patients (5.9%) with patch closure (p=ns). No patient in either group had a late neurologic event or required a redo operation. CONCLUSIONS: Selective primary closure is not associated with increased risk of perioperative neurologic events or statistically significant evidence of late postoperative stenosis if primary closure is performed in large internal carotid arteries. PMID- 10532219 TI - Long-term outcome in patients under 40 years after revascularization for chronic lower limb ischaemia. AB - BACKGROUND: In order to find out if surgical or endoluminal treatment changes the long-term results of atherosclerotic occlusive disease in patients of under 40 years of age we reviewed 17 consecutive patients. METHODS: Their mean age was 36.5. Patients with Buerger's disease or inflammatory arteriopathy were excluded. All patients were extremely heavy smokers. The indications for surgical procedures were disabling claudication (less than 100 meters) for 11 patients, rest pain for 4 patients and grangrene of a lower limb for 2 patients. The lesions were aorto-iliac in 12 cases and femoro-popliteal in 5. Ten surgical procedures were performed (5 aorto-femoral bypasses, 1 ilio-femoral bypass associated with an aorto-renal bypass, 2 femoropopliteal bypasses, 1 aorto-iliac endarteriectomy, 1 femoral endarteriectomy). On the other hand there were 7 endoluminal procedures (1 aortic, 4 iliac, 1 femoral and 1 popliteal). RESULTS: The mean follow-up was 97.3+/-50 months (range, from 34 to 216 months). Two patients died by 57 and 132 months respectively. At 5 years the survival rate was 94%; the primary patency rate was 59%; the secondary patency rate was 81% and the limb salvage rate was 94%. At 10 years these rates were respectively 94%, 44%, 54% and 75%. A total of 21 reoperations were performed. During follow-up 11 patients were better, 2 were stable and 4 were worse with 2 limbs lost. CONCLUSIONS: These bad results suggest keeping the surgical and endoluminal indications for patients younger than 40 years with threatened limbs. PMID- 10532220 TI - Venous hemodynamic changes after external banding valvuloplasty with varicosectomy in the treatment of primary varicose veins. AB - BACKGROUND: To evaluate venous hemodynamic changes after an external banding valvuloplasty in the treatment of primary varicose veins with saphenofemoral incompetence. METHODS: From June 1996 to December 1997, 79 limbs (10 male and 69 female, age 20-57 years) were treated for primary saphenofemoral incompetence by external banding valvuloplasty. Tightening of the banding was accomplished using a polyester-tailored mesh to narrow the terminal and/or subterminal valve areas of the dilated greater saphenous vein (GSV), same size as its minimum diameter during spasm. Evaluation was done through a pre- and postoperative color-flow duplex scanning and an air-plethysmography (APG). RESULTS: Sixty-three limbs (79.7%) remained patent and were competent. Fourteen limbs (17.7%) remained patent but showed reflux. Two limbs (2.5%) had thrombus within the GSV after surgery. The diameter of GSV of mid-thigh was 6.7+/-1.6 mm preoperatively and 4.1+/-0.9 mm postoperatively (p-value=7.04E-10). Reduction of the diameter was 61.4+/-12.3%. Venous volume was 136.1+/-59.8 ml preoperatively and 103.5+/-39.8 ml postoperatively (p-value=1.6E-20). Reduction of the venous volume was 12.9+/ 17.0%. Venous filling index (VFI) was 6.6+11.3 ml/sec preoperatively and 1.9+/ 3.3 ml/sec postoperatively (p-value=1.2E-10). Reduction of the VFI was 55.0+/ 29.1%. Ejection fraction (EF) was 48.9+/-13.8% preoperatively and 60.1+/-17.2% postoperatively (p-value=2.6E-17). Increase of EF was 29.4+/-43.5%. The residual volume fraction (RVF) was 42.1+/-13.9% preoperatively and 30.2+/-14.5% postoperatively (p-value=5.6E-19). Reduction of RVF was 17.6+/-43.6%. CONCLUSIONS: Early evaluation of saphenofemoral external banding valvuloplasty confirms the satisfactory patency and improvement in venous hemodynamics. Long term evaluation is clearly indicated but the early safety and efficacy of the procedure have been confirmed. PMID- 10532221 TI - Congestive heart failure due to adjuvant arteriovenous fistula in a femoroperoneal bypass. AB - The authors describe a patient who developed congestive heart failure one month after femoroperoneal bypass and adjuvant common ostium arteriovenous fistula procedure. The occlusion of the concomitant vein proximally to the arteriovenous fistula promptly resolved such a serious complication which, to our knowledge, has never been described previously. PMID- 10532222 TI - Experimental evaluation of retrograde cerebral perfusion by single photon emission computed tomography technique (SPECT). AB - BACKGROUND: Protection of the brain is of vital importance during aortic arch aneurysms. In this study efficiency of retrograde cerebral perfusion was evaluated with the use of single photon emission computed tomography technique (SPECT) by using 99mTc hexamethylpropylene amine oxime (HMPAO). METHODS: Four animals were used. The internal maxillary vein was the site of retrograde cerebral perfusion. The animals were studied after the heart rate and respiration were stopped with the use of the high dose drug administration and the brain was perfused with cold Ringer's lactated solution. After this procedure, 99mTc HMPAO SPECT study was performed. RESULTS: In one animal we did not get any cerebral image because of the competent venous valve in the internal maxillary vein. In the remaining animals, normal brain perfusion was achieved. CONCLUSIONS: 99mTc HMPAO-SPECT study documented that blood flow via the retrograde way meets the metabolic demand of the brain. Retrograde delivery of 99mTc HMPAO did not conclude any poorly perfused area in the brain when in given both sides and all parts of the brain can be effectively perfused by cerebral venous system in hypothermic conditions. PMID- 10532223 TI - Isolated true atherosclerotic aneurysm of the profunda femoris artery. Case report. AB - The authors report a case of true isolated atherosclerotic aneurysm of the profunda femoris artery. On the basis of a careful search of the literature some aspects of this rare disease are illuminated in terms of its low incidence, pathologic background and treatment; the last should always be aggressive due to the high possibility (about 50%) of major complications mainly represented by rupture. Simple aneurysmectomy without flow re-establishment may be allowed only if the femoropopliteal tract is normal PMID- 10532224 TI - Hydatid cyst of the abdominal aorta and bilateral common iliac arteries. A case report. AB - Hydatid disease, a parasitic infestation, is endemic in live-stock raising countries where the custom of feeding offal to dogs prevails. Though infestation of any part of the human body can occur, arterial involvement is a rare affliction. We report here the first case in which abdominal aorta and bilateral iliac arteries were totally occluded with intraluminal cysts. PMID- 10532225 TI - Simultaneous surgical intervention to coronary artery disease, peripheral arterial disease and superior mesenteric artery stenosis. AB - A patient, suffering from angina pectoris, claudicatio intermittens and postprandial abdominal pain underwent coronary and peripheral arteriographic examination; coronary arterial disease and aortoiliac occlusive disease was diagnosed. Color Doppler ultrasonography revealed superior mesenteric artery stenosis. CABG with MIDCAB (minimal invasive direct coronary artery bypass) technique was performed together with aortabifemoral graft interposition and graft bypass to superior mesenteric artery and considerable success was obtained. PMID- 10532226 TI - Successful tracheocarinal transplantation. AB - BACKGROUND: When extensive portions of the trachea and carina are resected, grafting is required. METHODS: Two experiments were performed in dogs to assess the feasibility of extensive tracheocarinal replacement using short-segment tracheocarinal autografts, only to avoid the immunologic complexity of allografts. To determine the effect of tension on graft survival, extensive tracheal defects (12 to 18 rings) were created in four animals. These were subsequently reconstructed using 6-ring autografts. In the second experiment, three animals underwent excision of a maximal length of trachea determined in experiment 1 including the carina. Long-term viability of each graft was assessed using bronchoscopy and histologic examination. RESULTS: The limit of tracheal resection successfully reconstructed using a 6-ring autograft was 14 rings (experiment 1). The tracheal grafts in which the tension was greater than 1.2 kg did not maintain their structural integrity. All of the autografts in experiment 2 were subjected to a tension of less than 1.0 kg at the anastomoses, and showed long-term viability. CONCLUSIONS: We conclude that extensive tracheal and carinal defects may be successfully reconstructed using short-segment tracheocarinal grafts if the anastomoses are subjected to less than 1.0 kg of tension. PMID- 10532227 TI - A standard muscle-sparing utility thoracotomy for VATS procedures. AB - BACKGROUND: Improvements in surgical equipment have rendered video-assisted thoracic surgery (VATS) an effective device for thoracic surgeons and nowadays several intrathoracic diseases can benefit from this approach. This development has expanded potential use and recently the technical feasibility of major lung resections by VATS has been demonstrated. The authors present their experience with a standard muscle-sparing utility thoracotomy (UT) utilized for all VATS procedures, including major lung resections. METHODS: From November 1996 to October 1997, 30 patients were operated on. There were 22 males and 8 females (medium age 58 years; range 24-78). There were 13 anatomical lung resections (i.e.: 11 lobectomies, 1 left pneumonectomy, 1 segmental resection), 8 wedge resections, 3 lung biopsies, 2 debridements of pleural empyema, 2 mediastinal nodes biopsies, 1 esophageal resection for leiomyoma, 1 excision of benign mediastinal cyst. RESULTS: No mortality or major morbidity were recorded, as well as no rib fractures due to the rib spreader. Two patients suffered from prolonged air-leaks after respectively left upper lobectomy and lung biopsy and required prolonged chest drainage. Concerning anatomic major lung resections the medium hospital stay was 7.9 days and medium chest tube time was 5.6 days. The utility thoracotomy through the auscultatory triangle proved to be a safe approach and confirmed the technical feasibility of various type of surgical procedures with results comparable to standard open thoracotomy. Our data shows that VATS approach did not seriously affect the duration of hospital stay, chest tube time, the overall morbidity or lung function. CONCLUSIONS: As the real benefit of this approach remains controversial, the majority of the studies comparing the VATS approach to conventional muscle-sparing thoracotomy neither nor prospective nor randomized, and several parameters are difficult to evaluate in the literature further study are mandatory. PMID- 10532228 TI - Vocal fold injection of collagen for unilateral vocal fold paralysis caused by chest diseases. AB - BACKGROUND: Patients having malignant chest diseases sometimes suffer from vocal fold paralysis. Treatment for vocal fold paralysis is important for such patients, because vocal fold paralysis causes lack of the versatility of the human voice which is essential for our communication. METHODS: Seventeen patients suffering from unilateral vocal fold paralysis were treated with vocal fold injections of collagen. Three patients received twice, and 20 treatments were conducted. A flexible bronchofiberscope was used under local anesthesia in order to observe the whole procedure of vocal fold injection. Using an injector and a long needle, collagen was injected with transcutaneous technique mainly through the cricothyroid membrane. The amount of collagen was determined with bronchoscopic findings. RESULTS: During and after treatment, no complication was observed. Of 20 treatments, a marked improvement was observed in 8, and moderate improvement was observed in 9 treatments. CONCLUSIONS: Vocal fold injection of collagen is a very useful and safe treatment for unilateral vocal fold paralysis caused by chest diseases. PMID- 10532230 TI - Comparison of anastomotic suturing techniques in the rat esophagus. AB - BACKGROUND: Long-gap esophageal atresia continues to be a challenging pediatric thoracic surgical problem. Despite the use of various tension relieving procedures, the esophageal anastomosis is often performed under considerable tension. Excessive tension can cause anastomotic sutures to pull through the esophageal tissue, with resultant early esophageal anastomotic dehiscence. To test the hypothesis that interrupted horizontal mattress sutures would withstand the forces of tension better than interrupted simple sutures, an experimental study of rat esophageal anastomoses was done. METHODS: Twenty rats were killed and their esophagi were excised. The esophagi were divided in the mid portion and end-to-end anastomoses were done using interrupted 6-0 polypropylene sutures. Ten rats had anastomoses done with interrupted simple sutures and ten had interrupted horizontal mattress suturing. Anastomotic breaking strength was tested in a tensiometer. RESULTS: Anastomotic breaking strength was 3.22+/-0.56 N for the interrupted simple sutured anastomoses and 3.51+/-0.61 N for the interrupted horizontal mattress group (p=0.30). The difference was not significant. CONCLUSIONS: In this animal study interrupted simple and horizontal mattress suturing withstood the disruptive forces of anastomotic tension equally well. PMID- 10532229 TI - Carcinoid tumors of the lung. An analysis of 65 operated cases. AB - BACKGROUND: The aim of this study was to analyse two groups of patients operated for bronchopulmonary neuroendocrine neoplasms (bronchial carcinoid and well differentiated neuroendocrine carcinoma) and to investigate their clinico pathological data and long-term survival. METHODS: From January 1978 to June 1996, 65 patients with bronchial carcinoids underwent operation at our Institution. There were 33 males and 32 females, whose mean age was 49.8 years. Forty-four neoplasms (67.7%) were considered to be central. Histology revealed 54 typical bronchial carcinoids (83%) and 11 well-differentiated neuroendocrine carcinomas (17%). Surgical resection of tumor and complete lymph node dissection was performed in all cases. RESULTS: All patients entered follow-up: 5-year survival was 91% for patients with bronchial carcinoid and 49% for those with well-differentiated neuroendocrine carcinoma (p<0.05). Univariate analysis found that there was a significant decrease in survival also for peripheral location of the tumor, advanced pathologic stage and histologically positive lymph nodes. CONCLUSIONS: These results point out that carcinoid tumors are malignant neoplasms, so they require a complete and radical surgical resection. Most tumors are only locally invasive and show a low aggressive behaviour; therefore, when possible, it is recommended to attempt a limited resection. Frozen sections of bronchial margins and complete lymphadenectomy should be routinely performed. The same criteria should apply to well differentiated neuroendocrine carcinomas, though their behaviour is more aggressive. PMID- 10532231 TI - Monofilament absorbable sutures in median sternotomy. AB - BACKGROUND: The most common material used for closure of median sternotomy incision is steel suture in open heart surgery. Some complications and disadvantages have been investigated recently. These complications are the breaking down of steel suture, erosion of sternum tabulae especially in osteoporotic patients, erosion of the dermis especially in patients with thin subdermic layer and cause of infection. Another disadventage of steel suture material is cosmetic problems or discomfort. For these reasons some suture materials such as silk, polyfilament polyester, monofilament material, polypropylene have been used recently. Silk and polyester have a risk of high infection, and polypropylene causes granulation tissue according to the number of knots. These facts encouraged the usage of an absorbable suture material. The available polyfilament absorbable sutures in the market a few years ago had a short absorption time, causing sternal infection and dehiscence. Polydiaxone, a monofilament suture material introduced recently has a considerably longer absorption time. METHODS: 153 sternal closures were performed with monofilament absorbable suture material in a period of seven months at the Kosuyolu Heart and Research Hospital. The mean age of the patients was 32.55, ranging from 8/12 to 71 years. The mean body weight is 48.37, ranging between 7 kg and 75 kg. RESULTS: Only two patients had sternal dehiscence. CONCLUSIONS: We conclude that monofilament absorbable suture is a safe alternative for all kinds of steel suture material for closure of sternotomy. PMID- 10532232 TI - Spectrophotometric monitoring in continuous-flow Boc-based solid-phase peptide synthesis. AB - It has been demonstrated that SPPS with Boc amino group protection can be monitored spectrophotometrically if it is performed in a continuous-flow reactor of variable volume. It is shown that this approach provides useful and adequate evidence on the beginning/end-point of most steps of the SPPS cycle. At the deprotection step the spectrophotometric monitoring enables real-time recording of the initial and final moments of the process. When synthesizing a 'difficult' polyalanine sequence, we were able to monitor variation in the deprotection dynamics associated with the aggregation phenomena. The time actually necessary for the Boc protecting group removal appeared to be significantly smaller than that usually preset in the available Boc-SPPS protocols. PMID- 10532233 TI - Synthesis of hydrazinopeptides using solid-phase N-electrophilic amination: extension to the Fmoc/tert-butyl strategy and chemistry of the N-N bond in strong acid media. AB - The synthesis of hydrazinopeptides using solid-phase N-electrophilic amination was extended to the Fmoc/tert-butyl strategy. Both Boc/benzyl and Fmoc/tert-butyl strategies led to the isolation of by-products arising from the partial instability of the N-N bond during the final cleavage and deprotection step. Two paths of decomposition have been shown: the cleavage of the N-N bond leading to the regeneration of the amine and a Hofmann-type elimination yielding original dianisyl adducts. Our data suggest that the Fmoc/tert-butyl strategy is better suited for the synthesis of hydrazinopeptides. PMID- 10532234 TI - Isolation and characterization of a novel superoxide dismutase from fungal strain Humicola lutea 110. AB - A novel thermostable MnSOD was purified to electrophoretic homogeneity from the fungal strain Humicola lutea 110. The preparation of the pure metalloenzyme was performed using treatment with acetone followed by ion exchange and gel permeation chromatography. We found that the activity of this enzyme comprises about 80% of the total superoxide dismutase activity in the crude extract, containing two proteins: MnSOD and Cu/ZnSOD. The MnSOD has a molecular mass of approximately 76 kDa and 7200 U/mg protein specific activity. It is a tetrameric enzyme with four identical subunits of 18 860 Da each as indicated by SDS-PAGE, amino acid analysis and mass spectrometry. N-terminal sequence analysis of MnSOD from the fungal strain revealed a high degree of structural homology with enzymes from other eukaryotic sources. Physicochemical properties were determined by absorption spectroscopy and circular dichroism measurements. The UV absorption spectrum was typical for an MnSOD enzyme, but displayed an increased absorption in the 280 nm region (epsilon280 = 10.4 mM(-1). cm(-1)), attributed to aromatic amino acid residues. The CD data show that MnSOD has two negative Cotton effects at 208 and 222 nm allowing the calculation of its helical content. The ellipticity at 222 nm is 6800 deg. x m(2) x dmol(-1) and thus similar to the values reported for other MnSODs. The MnSOD from H. lutea 110 is stable over a wide range of pH (4.5-8), even in the presence of EDTA. The enzyme is thermostable at 70-75 degrees C, and more stable than MnSODs from other sources. PMID- 10532235 TI - NMR analysis of human salivary mucin (MUC7) derived O-linked model glycopeptides: comparison of structural features and carbohydrate-peptide interactions. AB - Two series of glycopeptides with mono- and disaccharides, [GalNAc and Galbeta (1 3)GalNAc] O-linked to serine and threonine at one, two or three contiguous sites were synthesized and characterized by 1H NMR. The conformational effects governed by O-glycosylation were studied and compared with the corresponding non glycosylated counterparts using NMR, CD and molecular modelling. These model peptides encompassing the aa sequence, PAPPSSSAPPE (series I) and APPETTAAPPT (series II) were essentially derived from a 23-aa tandem repeat sequence of low molecular weight human salivary mucin (MUC7). NOEs, chemical shift perturbations and temperature coefficients of amide protons in aqueous and nonaqueous media suggest that carbohydrate moiety in threonine glycosylated peptides (series II) is in close proximity to the peptide backbone. An intramolecular hydrogen bonding between the amide proton of GalNAc or Galbeta (1-3)GalNAc and the carbonyl oxygen of the O-linked threonine residue is found to be the key structure stabilizing element. The carbohydrates in serine glycosylated peptides (series I), on the other hand, lack such intramolecular hydrogen bonding and assume a more apical position, thus allowing more rotational freedom around the O-glycosidic bond. The effect of O-glycosylation on peptide backbone is clearly reflected from the observed overall differences in sequential NOEs and CD band intensities among the various glycosylated and non-glycosylated analogues. Delineation of solution structure of these (glyco)peptides by NMR and CD revealed largely a poly L proline type II and/or random coil conformation for the peptide core. Typical peptide fragments of tandem repeat sequence of mucin (MUC7) showing profound glycosylation effects and distinct differences between serine and threonine glycosylation as observed in the present investigation could serve as template for further studies to understand the multifunctional role played by mucin glycoproteins. PMID- 10532236 TI - Comparative gene transfer efficiency of low molecular weight polylysine DNA condensing peptides. AB - In a previous report (M.S. Wadhwa et al. (1997) Bioconjugate Chem. 8, 81-88), we synthesized a panel of polylysine-containing peptides and determined that a minimal repeating lysine chain of 18 residues followed by a tryptophan and an alkylated cysteine residue (AlkCWK18) resulted in the formation of optimal size (78 nm diameter) plasmid DNA condensates that mediated efficient in vitro gene transfer. Shorter polylysine chains produced larger DNA condensates and mediated much lower gene expression while longer lysine chains were equivalent to AlkCWK18. Surprisingly, AlkCWK18 (molecular weight 2672) was a much better gene transfer agent than commercially available low molecular weight polylysine (molecular weight 1000-4000), despite its similar molecular weight. Possible explanations were that the cysteine or tryptophan residue in AlkCWK18 contributed to the DNA binding and the formation of small condensates or that the homogeneity of AlkCWK18 relative to low molecular weight polylysine facilitated optimal condensation. To test these hypotheses, the present study prepared AlkCYK18 and K20 and used these to form DNA condensates and conduct in vitro gene transfer. The results established that DNA condensates prepared with either AlkCYK18 or K20 possessed identical particle size and mediated in vitro gene transfer efficiencies that were indistinguishable from AlkCWK18 DNA condensates, eliminating the possibility of contributions from cysteine or tryptophan. However, a detailed chromatographic and electrospray mass spectrometry analysis of low molecular weight polylysine revealed it to possess a much lower than anticipated average chain length of dp 6. Thus, the short chain length of low molecular weight polylysine explains its inability to form small DNA condensates and mediate efficient gene transfer relative to AlkCWK18 DNA condensates. These experiments further emphasize the need to develop homogenous low molecular weight carrier molecules for nonviral gene delivery. PMID- 10532237 TI - Spot synthesis: observations and optimizations. AB - Positionally addressable syntheses of peptides on continuous cellulose membranes (spot synthesis) have often been reported in detail, but important questions dealing with synthesis quality, reproducibility and subsequent binding assays have largely been under-emphasized. In this report we have investigated some of these problems. The most important results were: (i) the signal intensity of ligate binding to cellulose-bound peptides and the affinity of the corresponding soluble peptides show good correlation, illustrated by three different ligate binding assays; (ii) reducing peptide density on the cellulose avoids the 'ring spot' effect, i.e. where less binding is observed in the spot-center compared to the rim. We recommend a peptide density of 10 nmol/cm2 as a reasonable starting point for further optimization; (iii) statistical analysis of binding assay reproducibility with more than 15000 peptides resulted in a mean standard signal deviation of 0.18; and (iv) optimization of side-chain deprotection revealed that a 30-min pretreatment of the cellulose with 90% trifluoroacetic acid followed by the standard deprotection protocol resulted in higher purity of the synthesized products. PMID- 10532238 TI - A convergent liquid-phase synthesis of salmon calcitonin. AB - Salmon calcitonin (sCT) was prepared in good yield and high purity by the condensation of Nalpha-Boc-cyclic decapeptide, Boc-C1SNLSTC7VLG-OH (1,7 disulfide), with protected docosapeptide (Psc)LSQE(OPse)LHK(Psc)LQTYPRTNTGSGTP NH2 x 3TFA, followed by deprotection of Boc with trifluoroacetic acid and Psc/Pse with piperidine. The 2-(phenylsulfonyl)ethoxycarbonyl (Psc) and 2 (phenylsulfonyl)ethyl (Pse) protecting groups were recently developed. The two peptides were built up by stepwise and fragment condensation using appropriate Nalpha-Boc-amino acids and subsequent deprotection in solution. The synthetic sCT exhibited hypocalcemic potency of more than 4000 IU/mg in rats. PMID- 10532239 TI - An evaluation of fMLP pocket dimensions and features using formyltetrapeptides. AB - The formyltetrapeptides for-Met-Leu-Leu-Phe-OMe 1, for-Met-Leu-Aib-Phe-OMe 2, for Met-Leu-Ac6c-Phe-OMe 3, for-Met-Leu-Pro-Phe-OMe 4, for-Met-Pro-Pro-Phe-OMe 5, for Met-Aib-Aib-Phe-OMe 6, for-Met-Pro-Aib-Phe-OMe 7 and for-Met-Aib-Pro-Phe-OMe 8 were synthesized and biologically tested on human neutrophils in an attempt to evaluate the specific receptor pocket dimensions and features. Our results indicate that the shift in the Phe residue to the fourth position in these compounds strongly reduces chemotactic response, but is efficacious in triggering superoxide anion production and lysozyme release (order of potency 3 > 2 > 1 > 4 > 6 > 8 > 5 > 7). The potency of the two latter responses correlates well with the affinity data obtained in binding experiments. PMID- 10532240 TI - Characterization of monomeric and dimeric B domain of Staphylococcal protein A. AB - Both monomeric and dimeric constructs of the B domain of protein A from Staphylococcus aureus have been characterized by NMR, CD and fluorescence spectroscopy. The monomeric form of the protein was synthesized using a novel method incorporating the use of a recombinant, folded, chimeric protein. A comparison of the recombinant monomeric form with the commercially available dimeric form indicates that, although the dimer retains the integrity of the three-helix bundle structure present in the monomer, there are interdomain contacts in the dimeric form. A single long-lived water molecule in the hydrophobic core of the three-helix bundle of monomeric protein A may represent an important stabilizing factor for the three-helix bundle topology. PMID- 10532241 TI - A spectroscopic and molecular modeling study on novel pseudopeptides exhibiting biological activity. AB - A series of pseudopeptides, containing two fluorophores, such as naphthalene (N) and indole (I), and exhibiting interesting biological activity as tachykinin receptor antagonists, were investigated by electronic absorption, CD and steady state fluorescence experiments. In polar solvents (e.g. methanol), bioactivity is coupled with a stacked, charge-separated complex between I and N, the amount of which depends on the stereochemical features and conformational mobility of the central scaffold in the molecules examined. This agrees with the idea that dipolar charged, spatially close, aromatic moieties are important topochemical elements in the mechanism of action of these receptor antagonists. Molecular mechanics calculations allowed us to build up hypothetical, low-energy conformations of a few representative pseudopeptides, whose structural features are consistent with the experimental findings. PMID- 10532242 TI - Influence of lipophilic groups in cationic alpha-helical peptides on their abilities to bind with DNA and deliver genes into cells. AB - For the purpose of achieving gene transfer into cells mediated by peptides with a short chain length, we employed two kinds of amphiphilic alpha-helix peptides, mastoparan (INLK-ALAA-LAKK-IL-NH2) obtained from wasp venom and an alpha-helix model peptide (LARL-LARL-LARL-NH2). Furthermore, to strengthen the hydrophobicity of the peptide required for the formation of the aggregates with the DNA, we modified these peptides using several lipophilic groups, i.e. acyl groups with a single chain, a dialkylcarbamoyl group and a cholesteryloxycarbonyl group. We examined the ability of the peptides and their derivatives to bind and aggregate with plasmid DNA, the structural change in the peptides caused by binding with the DNA and the in vitro gene transfer abilities into COS-7 cells. As a result, mastoparan was found to acquire the DNA binding ability by introduction of the lipophilic group. The conformational change in the peptides depended on the hydrophobicity of the introduced acyl group. The DNA complex of most lipophilic mastoparan derivatives could be incorporated into the cells via the endocytosis pathway. In the case of the helix model peptide, the acyl group with a moderate chain length was required for the formation of the aggregate which is competent for incorporation into the cells. In this study, we succeeded in giving such short peptides sufficient gene transfer ability by modifying them with some lipophilic groups. However, the influence of the modification by the lipophilic groups on the formation of aggregates with DNA and the gene transfer ability depended on the structure of the peptide portion. These results indicate that consideration of total hydrophobicity balance is needed for the design of an efficient gene carrier peptide. PMID- 10532243 TI - SERM: a new concept in the management of osteoporosis. Presentation. PMID- 10532244 TI - The burden of osteoporosis. PMID- 10532245 TI - Skeletal effects of estrogens. PMID- 10532247 TI - The effects of SERMs on the skeleton. PMID- 10532246 TI - Estrogen receptor and the SERM concept. PMID- 10532248 TI - Treatment induced changes of bone density and relative risk of vertebral fracture. PMID- 10532249 TI - Postmenopausal hormone therapy, SERMs, and coronary heart disease in women. PMID- 10532251 TI - Effect of SERMs on breast tissue. PMID- 10532250 TI - Effect of SERMs on the uterus and menopausal symptoms. PMID- 10532252 TI - Tolerability profile of SERMs. PMID- 10532253 TI - The role of SERMs in the management of postmenopausal osteoporosis. PMID- 10532254 TI - Choosing the patient for treatment. PMID- 10532256 TI - Intestinal physiology relevant to short-bowel syndrome. AB - In summary, with resection of the small bowel, the remaining intestine mounts an adaptive response to increase its absorptive surface area. The malabsorption of nutrients is predicated on the importance of the bowel resected to the absorption of a given nutrient. Motor activity is little effected by small-bowel resection. In general, resection of the upper small intestine is better tolerated than resection of the lower small intestine, particularly if the ileocecal junction is affected. PMID- 10532257 TI - Overview of intestinal adaptation and its stimulation. AB - Total parenteral nutrition (TPN) can be life-saving for many patients with short bowel syndrome (SBS). However, chronic TPN administration is associated with nutritional deficiencies, septic complications, high health care costs, and life threatening organ failure. In an effort to rehabilitate SBS patients so they may achieve enteral autonomy, investigators have attempted to stimulate the adaptive response following extensive small-bowel resection. Intestinal adaptation may include: 1) morphological changes of the residual bowel which increase the absorptive surface area; 2) functional changes that increase the absorptive capacity of individual enterocytes and colonocytes; and 3) changes in colonic production and absorption of short-chain fatty acids which improve intestinal vitality and maximize efficiency of energy and fluid absorption. Several peptides, nutrients, cytokines, and other factors promote intestinal adaptation in animals. These "growth" factors may predominantly affect one aspect of the adaptive response while having little or no effect on other physiologic or morphologic parameters. In addition, combined administration of stimulatory agents may be necessary to enhance adaptation. Dietary constituents may have profound positive and negative effects on adaptation and must be considered in developing an overall plan for treatment of the SBS patients. Only a few clinical studies have been performed to evaluate therapeutic regimens for SBS beyond standard supportive care and TPN administration. The combined administration of growth hormone, glutamine and a modified diet to over 225 adults has been shown to eliminate or decrease TPN dependence in 80% of patients receiving this therapy. Further study is required to optimize the treatment of humans with intestinal failure and to determine which patients are most likely to benefit from medical therapy. The authors conclude that the intestinal length to body weight index may be one predictive factor useful for determining which SBS patients will benefit from a trial of pharmacologic manipulation before attempting alternative, potentially more invasive therapies. PMID- 10532258 TI - Intestinal adaptation in pediatric patients with short-bowel syndrome. AB - The purpose of this study is to evaluate 12 pediatric short-bowel syndrome (SBS) patients experienced at Osaka University Hospital and its affiliated hospitals and to study the intestinal length for achieving intestinal adaptation and the metabolic characteristics. The length of the residual small intestine ranged from 0 to 75 cm with an average of 47 cm and an ileocecal valve had been resected in five cases. Total parenteral nutrition (TPN) was started immediately after operation and was gradually substituted by enteral nutrition. No patient died during the follow-up period. Eight of 12 patients could be weaned from TPN with residual intestinal length of 27 to 75 cm (mean 57 cm). Four patients with the residual intestine of 0 to 45 cm (mean 22 cm) were unable to achieve intestinal adaptation. The rate of catheter-related sepsis per 1000 catheter days was 0.63. Fatty liver was detected in two cases, but no patient developed progressive liver failure. Plasma arginine and citrulline were decreased in patients who were unable to achieve intestinal adaptation. Our nutritional support program provided excellent survival for pediatric SBS patients primarily due to the low incidence of catheter-related sepsis and no episode of severe liver disease. Patients with more than 16 cm of residual intestinal length can be expected to be weaned from TPN. PMID- 10532259 TI - Surgical stress, bacteria, and mucosal immune function. AB - Bacteria share a benign coexistence with host mucosal surfaces in the gastrointestinal tract during periods of health. Both host epithelial defense function and bacterial virulence phenotypes are significantly affected by stress. Via discreet and specific sensory input signals to bacteria, the molecular machinery of otherwise commensal strains of bacteria can shift the phenotypes of residential colonizers to more virulent and invasive strains. This occurs at a time when the host may be relatively immunosuppressed by the injury. This adaptive response demonstrates the duplicitous nature of bacteria residing on mucosal surfaces whose ability to shift their virulence characteristics may play an important role in infectious-related morbidity following surgical stress. PMID- 10532260 TI - Enteral and parenteral nutrition in patients with short-bowel syndrome. AB - Short-bowel syndrome is functionally defined as a state of malabsorption following loss of small bowel. Most cases occur in the neonatal period after extensive resection for necrotizing enterocolitis, or due to congenital anomalies of the gastrointestinal tract. A smaller percentage originate later in life from surgical treatment of Crohn's disease, neoplastic disorders, or vascular events. The physiological, morphological and functional intestinal gradient determines the clinical picture leading to better tolerance of jejunal than ileal resections. The subsequent adaptation process requires enteral feeding with a different impact of specific nutrients, and is also influenced by a number of humoral mediators such as enteroglucagon, gastrin, growth factors, prostaglandins and polyamines. Nutritional management starts parenterally via a central venous line covering basic demands, substituting current losses and restoring pre existing deficiencies. Continuous enteral tube feeding is added as soon as postoperative ileus resolves, beginning with an elemental diet, which is gradually increased first in concentration, then in quantity, and supplemented by small oral meals. Cycling of parenteral nutrition is the next step. As soon as sufficient stability is reached, the child should be discharged home under continued outpatient care. Main long-term problems comprise bacterial overgrowth, fluid and electrolyte disequilibration, nutritional deficiencies, parenteral nutrition-related liver disease, and central venous line complications such as sepsis and thrombosis. PMID- 10532261 TI - Bacterial translocation is favored by the preservation of the ileocecal valve in experimental short bowel with total parenteral nutrition. AB - Sepsis in short-bowel syndrome (SBS) is in part due to bacterial translocation (BT). Parenteral nutrition (PN) is often necessary in SBS and promotes BT. The aim of this study was to asses the effect of the presence or absence of ileocecal valve (ICV) on BT in parenterally-fed rats with massive intestinal resection. Sixty-five adult Wistar rats underwent central venous cannulations and were randomly assigned to one of five groups receiving for ten days five treatment regimes: Sham (n = 17) standard rat chow + i.v. saline. PN (n = 17) fasting + PN. Res-Sham (n = 10) standard rat chow + i.v. saline + 80% gut resection. Res-PN (n = 11) fasting, PN + 80% gut resection. Res-ICV-PN (n = 10) fasting, PN + 80% gut resection including ICV. At the end of the experiment they were euthanized and mesenteric lymph nodes (MLN), spleen and peripheral and portal blood specimens were recovered and cultured. BT was found in 47% of PN animals, 91% of Res-PN rats, 100% of Res-Sham group and 60% of Res-ICV-PN animals, but not in Sham ones. 97% of BT+ animals had positive cultures in MLN and/or portal blood, whereas germs beyond liver were detected in 30% of Res-Sham, 37% of PN, 50% of Res-PN and 0% of Res-ICV-PN rats. The present study confirms that both massive intestinal resection and PN promote BT. In addition, it shows that animals deprived of ICV have lower incidence of BT in this setting than those with it and that the germs do not reach in them peripheral blood in the same proportions as in ICV-intact animals. These results suggest that the presence of an intact ICV favor BT in parenterally-fed rats with massive intestinal resection. PMID- 10532262 TI - Bacterial translocation in short-bowel syndrome in rats. AB - Massive intestinal resection results in short-bowel syndrome (SBS) and is associated with an increased risk of infectious complications mainly caused by the egress of intestinal bacteria to distant organs, a process termed bacterial translocation (BT). The purpose of this experimental study in rats was to investigate in different models of SBS the impact of the type of intestinal resection on bacterial growth in the residual small bowel and on the occurrence of BT. SBS was created in 30 rats either by jejunal resection (JR), by ileal resection (IR) or by ileal resection including the ileocecal valve (IR+ICV). 10 animals underwent only a sham laparotomy (SL) and served as controls. Two weeks after the operative procedure, intestinal bacterial colonization and BT to the portal vein, vena cava, mesenteric lymph nodes, liver and spleen were determined. All resected animals showed a decreased weight gain and a significant bacterial overgrowth in the residual small bowel compared to the SL group. BT occurred after SL in 12%, after JR in 70%, after IR in 58%, and was significantly less frequent (35%) after IR+ICV, respectively. These experimental findings suggest that BT in SBS might be promoted by the intestinal bacterial overgrowth in the residual bowel, and the incidence of BT seems to be related to the presence or absence of the ileocecal valve. PMID- 10532263 TI - An analysis of the morbidity and mortality of short-bowel syndrome in the pediatric age group. AB - Twenty-two children with short-bowel syndrome (SBS) were treated at the C. S. Mott Children's Hospital in the University of Michigan Medical Center between June 1983 and May 1993. Definition of SBS was loss of 70% or more of the total small bowel. Seventeen of these children are currently alive, a 77% survival rate. Patients were followed for a mean of 1,148 days. The mean age of SBS development was 71 days of life. The only predictive indicator of patient survival was direct bilirubin levels. Sixty-seven percent of the children died if they had a direct bilirubin of > 4 mg/dl > or = 6 months duration. PMID- 10532264 TI - Morbidity and mortality of the short-bowel syndrome. AB - From 1976 to 1998 we have treated 17 neonates with short-bowel syndrome. Those 8 patients who had an intact ileocecal valve as well as the total colon preserved did significantly better than the 9 children without ileocecal valve and > 50% missing colon. In addition to the length of the intestinal remnants, motility had a major impact on the incidence of complications and final outcome. Four patients died (23.5%). All of them had an intestinal length of less than 30 cm, severe dysmotility, no ileocecal valve and an incomplete colon. The average duration of hospitalization of the children weaned from parenteral nutrition (n = 11) was 8.5 months. The majority of them still need supplementation of vitamins and/or trace elements. Two children suffer from recurrent d-lactic acidemia. Six children have a significant psychomotor developmental delay with three suffering from congenital cerebral abnormalities. PMID- 10532265 TI - Severe short-bowel syndrome in children. Clinical experience. AB - INTRODUCTION: Innovative surgical and pharmacological therapeutic measures in short-bowel syndrome (SBS) are constantly changing the prognosis of this devastating condition. The aim of this paper is to present our most recent experience in the treatment of this disease, with particular emphasis on the impact of home parenteral nutrition (HPN) and the use of growth hormone (GH). METHODS: A group of 8 patients with severe SBS have been studied for the past 4 years. Intestinal length of less than 25% normal at the time of bowel resection was the criterion for inclusion in this study. RESULTS: Mean age at the time of diagnosis was 2 years (ranging from 1 day to 9 years). The etiology of the SBS was Hirschsprung's disease (n = 3), midgut volvulus (n = 2), gastroschisis (n = 1), omphalocele with ileal atresia and necrotizing enterocolitis (n = 1) and Crohn's disease (n = 1). Length of the residual bowel was 8 and 50 cm with ileocecal valve (ICV) preservation and 23, 27, 30, 50, 70, 100 cm without ICV. Sixty percent of the patients survived. Two patients died due to fulminant gram negative sepsis and one due to cardiac malformation. Two patients are still on parenteral nutrition (PN) providing 30 and 60% of total calories. Human GH (0.3 U/kg/day) was used in two patients over a period of 28 days. In these patients, an increased tolerance to enteral feeding was observed. HPN was provided in 5 cases, allowing regular school attendance in 3 patients. In 3 cases, discontinuation of the PN was achieved at 24, 25 and 35 months respectively. CONCLUSIONS: Human GH can improve tolerance of enteral feeding. HPN has a beneficial effect on child behaviour. Intestinal transplantation must be considered when no other surgical or medical measures are available. PMID- 10532266 TI - Progressive perinatal bowel obstruction--a rare cause of short-bowel syndrome. AB - A girl was born after an uneventful pregnancy of 36 weeks. Prenatally, distended bowel loops had been seen on ultrasound. Multiple small-bowel atresia was diagnosed and treated surgically. In the course of the next eleven weeks, previously patent segments of small bowel became obstructed. In 4 separate operative sessions, several segments of jejunum and ileum were resected, leaving 23 cm of ileum with the ileocecal valve in place. On microscopic examination of all resected material, necrosis of the mucosa was found consistent with ischemia. The child survived and tolerated full enteral feeding at the age of 8.5 months. The origin of the progressive obliterating process remains unknown. PMID- 10532267 TI - Hepatopathy in two infants with short-bowel syndrome and cytomegalovirus infection. AB - In children with short-bowel syndrome and the need for long-term parenteral nutrition, hepatic dysfunction is a multifactorial phenomenon that has not been completely understood. Alterations in gut motility lead to intraluminal stasis which is thought to be a major etiologic factor for bacterial overgrowth and subsequent cholestasis, especially when the ileocecal valve is absent. We report on two infants with short-bowel syndrome caused by gastroschisis and intestinal atresia. The intestinal lengths after resection were 18 and 55 cm. Long-term parenteral nutrition (PN) was obligatory due to intestinal shortness in the first patient and dilatation of the preatretic bowel segment with ineffective peristalsis in the second patient. Despite multiple trials of enteral nutrition and medical therapy for gut decontamination and stimulation of bowel motility, hepatopathy developed in both patients in a similar period of time and to about the same degree. At the age of 4 and 6 weeks, respectively, increasing bilirubin values were measured. Deterioration of liver function and thrombocytopenia at the age of 3 to 4 months led to the diagnosis of acute cytomegalovirus (CMV) infection. Treatment with ganciclovir followed. Both patients died of acute liver failure at the age of 7 and 9 months, respectively. Additional hepatic injury secondary to CMV infection might have contributed to the rapid deterioration of liver disease. Screening for further hepatotoxic factors, especially infectious etiologies, is therefore recommended in children with short-bowel syndrome. Liver transplantation should be considered early in cases of progressive hepatic dysfunction. PMID- 10532268 TI - Congenital short-bowel; a case study and review of the literature. AB - A congenital short bowel (CSB) is a rare entity in pediatric surgery. We present the case of a newborn boy with a total small intestinal length of 47 cm, malrotation and gastroesophageal reflux, who is 19 months old at the time of this report. Main treatment steps were Ladd's procedure, a fundoplication and long term parenteral nutrition. We suggest that missing physiological herniation of the gut into the coelomic cavity may impair normal intestinal growth and rotation and lead to congenital short bowel. Review of all cases reported in the literature shows a considerable mortality of 88%. The limiting factor seems to be reduced motility of the short small bowel causing functional obstruction and liver failure. PMID- 10532269 TI - Short-bowel syndrome associated with subtotal necrosis of small intestine after rectal trauma. AB - We report on a 4-year-old girl who experienced rectal trauma during swimming, sitting on an uncovered draining valve in the swimming pool. This resulted in a powerful suction effect on her rectum, followed by rupture of the sigmoid colon and evisceration of the small intestine. Laparotomy showed a near complete necrosis of the small bowel because of thrombotic lesions and wall lacerations of the superior mesenteric artery (SMA). A subtotal bowel removal associated with a jejuno-ileostoma was carried out, a total length of about 35 cm of the small intestine could be left in situ. Parenteral nutrition was stopped after eight months. At the moment defecation takes place 2-3 times a day, growth and weight gain are quite normal. PMID- 10532270 TI - Near total intestinal aganglionosis with extreme short-bowel syndrome--a difficult surgical dilemma. AB - Forty cases of total or near-total intestinal agangliosis (NTIA) were described to date in the English literature. Most cases had a lethal outcome. We describe the 41st case--a Beduin male neonate--who had only 30 cm of proximal hypoganglionic jejunum. He is presently almost one-year-old and thriving on home TPN, receiving one quarter of his caloric requirements orally using pregestamil, an MCT formula. The initial intricate course, diagnosis and several operative procedures, are elaborated. A review of the scant literature is discussed. The elusiveness of the correct diagnosis is pointed out and means to overcome these errors are described. Various surgical procedures have been suggested, none of which offer the perfect solution to the severe basic problem of short bowel. Long term parenteral hyperalimentation is still the main modality of treatment. Based on our modest experience, we suggest saving every possible length of jejunum, even if hypoganglionic, since this bowel, following a few weeks of adaptation, starts to function fairly well, suggesting perhaps some neuro-muscular maturation. The best surgical approach is still pending. We present a report of a child with this disease and discuss the therapeutic dilemma. PMID- 10532271 TI - Experience with longitudinal intestinal lengthening and tailoring. AB - Over a 16-year period, 20 neonates and infants with short-bowel syndrome underwent longitudinal intestinal lengthening and tailoring because of a dysfunctional dilated jejunum. There was no operative mortality, and morbidity was limited to 2 hemiloop anastomotic stenoses and 1 spontaneously resolving air and bile leak. Long-term survival was 45%. Survivors had >40 cm residual jejunum and a greater number also retained their ileocaecal valve and a longer colonic length. They underwent bowel lengthening at a later time and had minimal hepatic dysfunction. 7 of 9 survivors established full enteral nutrition. These children could be regarded as self-selected survivors with residual bowel dysfunction who had come through the hazardous neonatal phase with minimal hepatic injury. Non survivors often had <40 cm jejunum and limited distal colon. Death was commonly due to end-stage liver failure. It is likely that the severely reduced gut associated lymphoid tissue contributed to increased bacterial translocation from the dilated bowel and early onset of progressive liver injury. It is possible to conclude that bowel lengthening should be offered only to self-selected survivors with residual bowel dysfunction and minimal liver injury. It seems, however, even more appropriate, to offer early bowel tailoring and lengthening with its recognized reduction in stasis and bacterial translocation, improved absorption and enhanced intestinal adaption, particularly to those high-risk neonates with <40 cm of dilated jejunum with a view to reducing the risk of infection and lethal hepatic injury, thereby improving their chances for quality survival. PMID- 10532272 TI - What do children look like after longitudinal intestinal lengthening. AB - The longitudinal intestinal lengthening, described by Bianchi in 1980, has been shown to be effective in improving intestinal function, absorption and transit time in patients with short-bowel syndrome. We report the long-term results of 18 survivors of a series of 25 intestinal lengthening procedures performed since 1984. Mean age of the patients was 18 months (range of 5 to 52 months), mean follow-up 6 years (0.9 to 12 years). Parenteral nutrition was progressively reduced in all patients and discontinued after 1 to 10 months (mean 5.1 months). Frequently encountered problems during long-term follow-up are hyperphagia, hyponatremia and hypochloremia, metabolic acidosis, including D-lactic acidosis, cholelithiasis and urolithiasis, gastro-esophageal reflux, dystrophy and symptoms caused by secondary dilatation of the lengthened bowel loops: a protruding abdomen, enteral stasis, leading to constipation or diarrhea with bacterial overgrowth. Overall performance has been acceptable in 13 out of 18 patients. Longitudinal intestinal lengthening is effective enabling patients with short bowel syndrome to be weaned from parenteral nutrition, allowing for long-term survival. However, it is only one step on a long and difficult way. Multiple problems have to be searched for and adequately dealt with to achieve an acceptable and future worth living. PMID- 10532273 TI - Surgical approach to the short-bowel syndrome: procedures to slow intestinal transit. AB - Procedures designed to slow intestinal transit should be applied cautiously in patients with near adequate remnant length and demonstrated rapid transit. They should be considered after maximum adaptation has occurred. A reversed intestinal segment appears to be efficacious in the short term. Colon interposition and intestinal valves might also have merit in selected patients but clinical experience is limited. Unfortunately, these procedures are applicable to only a small proportion of patients with SBS. PMID- 10532274 TI - Ultra-short-bowel syndrome is not an absolute indication to small-bowel transplantation in childhood. AB - Short-bowel syndrome (SBS) either in adults or in children is considered as an indication to small-bowel transplantation (SBTx), particularly in its most severe form with a residual bowel length below 20 cm. Among factors likely to worsen the prognosis, more recent reports also indicate the number of surgical interventions, early onset sepsis and early development of liver disease. We report six cases of ultra-short-bowel syndrome followed from birth to verify the importance of various prognostic factors. In our case series, the male sex is predominating (5:1). Intestinal resection was indicated in 3 patients for multiple intestinal atresias, in 2 for volvulus and in 1 for necrotizing enterocolitis. The length of intestine remaining was invariably less than 20 cm and 2 patients had a preserved ileocecal valve. In most cases, more than 50% of the colon remained. The number of abdominal operations ranged from 1 to 4. In almost all cases (5 of 6), sepsis and hepatopathy developed early. Our experience suggests that rather than depending on the length of intestine remaining or the presence of the ileocecal valve, the prognosis of patients with the extreme-short bowel syndrome depends on recurrent neonatal onset sepsis and early onset liver impairment. In addition, our case review shows that the extreme-short-bowel syndrome is not necessarily an indication for bowel transplantation. PMID- 10532276 TI - Charter on visitation of training centres: UEMS, October 1997. Union of European Medical Specialists PMID- 10532275 TI - Intestinal transplantation. Experience in the United States. AB - This review presents recent experience with intestinal transplantation in the United States and Canada. Indications are for combined liver and small intestinal transplantation irreversible intestinal failure and end-stage liver disease, and- for isolated intestinal transplantation--intestinal failure associated with severe progressive complications of parenteral nutrition. With a proportion of 80%, short-bowel syndrome represents the major disease complex thus treated. Long term graft and patient survival exceeds 50% in large series with better outcome for isolated intestinal grafts than for combined liver and small-bowel transplants. Limiting factors are infections (responsible for 60% of graft losses), frequently due to CMV or EBV, technical and management errors (22%), and rejection (14%). One of the main problems of immunosuppression is post-transplant lymphoproliferative disease with an incidence of 20% which seems especially linked with EBV infections, OKT3 and steroids. The first line immunosuppressive agent nowadays used is tacrolimus. Trials with unmodified donor bone-marrow infusions to promote graft acceptance have not proved successful so far. Hopes for the future include increasing use of living related donors in order to overcome the shortage of donor organs, and continuing progress in immunosuppression. PMID- 10532277 TI - Cancer therapy: new concepts on active immunization. AB - There is increasing evidence that tumors express putative target molecules for a therapeutic immune reaction. Yet, tumor cells lack the prerequisites for appropriate antigen presentation and--hence--the immune system does not respond. This difficulty can probably be circumvented when tumor antigens are processed by conventional antigen presenting cells. Thus, the identification of immunogenic tumor-associated antigens may allow new modes of vaccination with the hope of adding a fourth and hopefully powerful weapon to surgery, radiation and chemotherapy in the fight against cancer. PMID- 10532278 TI - Immune response against heat shock proteins in infectious diseases. AB - Heat shock proteins (hsp) are conserved molecules that play an important role in protein folding and assembly and in translocation of proteins between different compartments. Under stress, hsp synthesis is drastically increased, representing a mechanism essential for cell survival. During infection or inflammation, numerous hsp are overexpressed. Not surprisingly, hsp represent dominant antigens in many infectious and autoimmune diseases that induce strong humoral and cellular immune responses. There is substantial evidence that hsp are dominant immune targets in a number of diseases, to the benefit or detriment of man. Nevertheless, findings also exist which argue against a universal role for hsp as target antigens in disease situations. It is suggested that hsp mainly serve as 'early' targets in the immune response, thus providing support for anti infectious or autoaggressive immune responses directed against unique pathogen- or disease-associated antigens, respectively. PMID- 10532279 TI - Effect of electromagnetic fields on several CD markers and transcription and expression of CD4. AB - We carried out flow cytometric analysis for multiparametric evaluation of cell surface markers related to cellular functions. Specifically, we studied the expression of CD4, CD8, CD3, CD16, CD19, HLA-DR, and CD14 macrophage receptors expression and cell cycle progression on cells exposed to ELF-EMF. In addition, we tested the effects of ELF-EMF on CD4 mRNA protein transcription and translation and the cell-cycle progression using an immunofluorescence method. Our data show that same CD surface marker expression are weakly influenced by electromagnetic fields, with no differences between cells exposed or not exposed to ELF-EMFs. However, when the CD4 protein generation was studied, an indication of protein production was found in lymphocytes exposed to ELF-EMF, as evidenced by immunofluorescence, Western blotting and RT-PCR analysis. CD16 and CD14 expression were affected by EMF exposure at all times studied (24, 48, 72 h). The results obtained with cell cycle analysis show that after 48 h of exposure to ELF EMF, PHA-activated and not activated cells in S phase increase with respect to non-exposed cells. The findings from this study demonstrate that under our defined experimental conditions there is evidence that ELF-EMF has a slight effect on CD4, CD14 and CD16 receptor expression, while the other CD receptors are not affected. PMID- 10532280 TI - Involvement of CD28 cosignalling in the T cell-mediated suppression of the IgG antibody response against the TI-2 antigen alpha(1-->3) dextran. AB - The humoral immune response against alpha(1-->3) dextran (Dex) in BALB/c mice is characterized by the formation of predominantly IgM antibodies bearing the J558 idiotype. IgG antibodies do not appear in euthymic mice. In athymic animals, however, the response proceeds to a vigorous IgG production. In euthymic mice formation of IgG is suppressed by J558 idiotype specific regulatory T cells recognizing in association with I-Ed and in cognate T/B interaction the V(H) CDR3 derived peptide of the J558 idiotype. Only B-2 lymphocytes produce IgG whereas B 1 cells do not participate in the production of this Ig class. Using novel synthetic all alpha(1-->3)-D-gluco configured tetrasaccharide the Dex-specific B cells can for the first time be analyzed in FACS. In experiments using this newly designed low molecular Dex no signs of B cell apoptosis can be found. This demonstrates a true silencing of persisting Bgamma memory cells as previously suggested by adoptive transfer experiments. In this suppression a further involvement of CD28 and B7-1 interaction can be demonstrated which delivers a necessary costimulatory suppression signal in addition to the cognate TCR/peptide I-Ed interaction between J558 specific T cells and J558 idiotype bearing B cells. PMID- 10532281 TI - Initial characterization of the complement activating compounds in extracts of smokeless tobacco. AB - Aqueous extracts of smokeless tobacco (ST) have been shown to be potent activators of complement. However, the mechanisms by which smokeless tobacco activates complement are not well understood. This study was undertaken to identify the complement activating compounds in ST extracts. The approximate molecular size of the activating agent(s) in smokeless tobacco was determined by dialyzing aqueous extracts of loose leaf chewing tobacco (1S1), dry snuff (1S2), and moist snuff (1S3). Following dialysis (total dilution effect of 1:10(9)), using a membrane with a molecular weight retention limit of 12-14 kDa, all extracts retained full capacity to activate serum complement as determined by a hemolytic assay. Fractionation of the extracts by gel filtration chromatography revealed that the complement activating agents in ST were high molecular weight compounds that eluted between 400 kDa and the void volume (1500 kDa) of a Sephacryl S300 column. The high molecular weight complement-activating peak was isolated and found to be a more potent complement activator than the unfractionated extract. The chemical nature of the complement activating compounds was determined by subjecting the extracts to boiling for 30 min, an organic extraction with chloroform/methanol 2:1, or treatment with a DNAse/RNAse enzyme cocktail. None of these treatments destroyed the capacity of ST extracts to activate complement, suggesting that the activating agents may be carbohydrate like. Finally, an extraction protocol designed to remove polyphenols significantly diminished the complement activating capacity of the ST extracts. These results clearly demonstrate that the complement activating substances in smokeless tobacco extracts may be large (>400 kDa) polyphenol-containing compounds (i.e. tannins). Identification of this agent(s) will be important for distinguishing the mechanism of smokeless tobacco-induced complement activation. PMID- 10532282 TI - Quantification of B, T and null lymphocyte subpopulations in the blood and lymphoid organs of the pig. AB - Research on the pig's immune system is not only of general biological interest; the pig is also becoming more important as a large animal model in human biomedical research, e.g. as a donor for xeno-transplantation. With the increasing panel of monoclonal antibodies against porcine lymphocyte markers it is possible to gain more insight into the distribution and phenotype of lymphocyte subpopulations in the pig. In this study we investigated B cells (surface IgG: sIgG, sIgM and sIgA) and T cells (CD2, CD4, CD8, 8/1, MAC320) in the peripheral blood (pBL), thymus, spleen, tonsil, mesenteric and inguinal lymph nodes (mLN, iLN), jejunal and ileal Peyer's patches (jejPP, ilPP) in Gottingen minipigs. A flow cytometric technique was employed which enabled three color indirect immunofluorescence. B cell stained for surface IgG and surface IgA were found only in small percentages. Surface IgM positive cells were distributed at higher rates, with up to 24.9% in the iLN. Up to 64.2% of CD4+ and up to 73.1% of CD8+ cells were observed in the thymus. Most of the CD4+ cells were CD4/CD8 double positive cells. These cells were mostly triple positive in combination with CD2. A larger fraction of CD2- were CD8- which are taken to be NK cells. MAC320, a marker for a subtype of gamma/delta T cells, was predominantly found on cells in the pBL. The standardized flow cytometric technique produced comparable data on the distribution of major lymphocyte subpopulations in the blood and different lymphoid organs of the pig. The results provide a basis for future studies using the pig as animal model. PMID- 10532283 TI - Airway epithelial cell-induced activation of monocytes and eosinophils in respiratory syncytial viral infection. AB - The early inflammatory events in respiratory syncytial viral (RSV) infection are likely to be crucial in the development of clinical disease, which is characterized by bronchiolitis with mononuclear cell inflammation, some eosinophil involvement and airway hyperreactivity. Since RSV replication is restricted to airway epithelial cells, our working hypothesis is that inflammatory cell recruitment by the infected cells will set the stage for late immunopathology. We have identified the selective induction and release of mononuclear cell and eosinophil-attracting beta-chemokines MIP-1alpha and RANTES, but not eotaxin, by RSV-infected airway epithelial cells and herein demonstrated the recruitment of eosinophils and monocytes, but not neutrophils, in response to chemokines produced by infected epithelial cells during viral replication and dissemination. The chemotactic response of both eosinophils and monocytes was inhibited by antibodies to RANTES but not to MIP-1alpha. Interaction of eosinophils or monocytes with RSV-infected epithelial cells resulted in the production of additional beta-chemokines MCP-1 and MIP-1beta, and increased levels of MIP-1alpha. The monocyte containing cultures produced >10 fold the amount of these chemokines compared to eosinophil containing cultures. On the other hand, the levels of RANTES and the lack of eotaxin were not altered in the cocultures, RSV-infected monocytes appeared to be the main source of MIP-1alpha and MIP-1beta, while MCP-1 was derived from monocytes as well as epithelial cells following coculture. These data implicate RANTES as the primary chemokine responsible for selectively recruiting eosinophils and monocytes to the site of RSV infection. This inflammatory response results in the production of high levels of additional chemokines capable of setting up a full-fledged inflammatory response including lymphocytes. PMID- 10532284 TI - Colcemid but not taxol modulates the migratory behavior of human T lymphocytes within 3-D collagen lattices. AB - T cell migration within tissue requires engagement of the cytoskeleton, however, little is known about the functional role of both actin- and tubulin-based cytoskeleton in this process. We investigated the direct effect of microtubule disruption and stabilization using colcemid and taxol, respectively, on the locomotion of peripheral human T cells within three-dimensional (3-D) collagen lattices. Microtubules network disassembly very potently enhanced T cell migration, nearly doubling the fraction of locomoting cells. Both a recruitment of previously sessile cells as well as an increase in the mean duration of active locomotion contributed to the promigratory effect. The stimulatory effect was correlated with the loss of the integrity of the tubulin cytoskeleton. Reassembly of microtubules, subsequent to the removal of colcemid from the cells, resulted in the successive return of the migratory activity to baseline levels. On the contrary, taxol failed to modulate T cell migration in our in vitro assay despite its potency to assemble tubulin into compact clots. Our observations underscore the view that tubulin-dependent cellular deformability is not the rate-limiting factor for locomotion and provide evidence that the increase in migratory activity subsequent to colcemid-treatment is due to a secondary phenomenon, most likely the activation of the actin cytoskeleton. PMID- 10532285 TI - The role of intestinal bacterial flora in the tuning of the T cell repertoire. AB - The role of the intestinal bacterial flora on the Vbeta repertoire was examined using the gnotobiotic murine model. The ratio of Vbeta6-positive T cells in the periphery of DBA/2 mice under SPF conditions was only 2.2% (mean, n = 4), since the cells were eliminated by the endogenous superantigen Mls(a). However, the ratio in germ-free (GF) mice was 31.7%. Similarly, the contamination of the GF Mice with the intestinal flora from SPF mice reduced the ratio of Vbeta6 in GF mice from 22.9% to 13.7%. In contrast, in BALB/c mice (Mls(b)) in which Vbeta6 cells do not react with this endogenous superantigen, the ratio of Vbeta6 cells do not react with this endogenous superantigen, the ratio of Vbeta6 of SPF mice (15.4%, mean, n = 3) was found to be comparable to that of GF mice (15.6%, n = 3). These data suggested that the absence of intestinal flora deteriorated a part of the Mls(a) determinant, which reacted with the Vbeta6 T cells and thereby eliminated them, thus resulting in an increase of these cells in GF mice. Moreover, the alloantigenicity of minor histocompatible alloantigen(s) (mHAg) in SPF mice, which was detected in H-2 identical MLR experiments and a murine graft versus-host (GVH) model, was reduced in GF and decontaminated SPF mice, thus indicating that the intestinal flora upregulated the mHAg including a part of Mls determinant. These results therefore suggest that the intestinal flora plays a role in the upregulation of mHAg including a part of endogenous superantigen and the consequent tuning of the Vbeta repertoire. PMID- 10532286 TI - Dissimilar attenuation of Candida albicans virulence properties by human immunodeficiency virus type 1 protease inhibitors. AB - The secreted aspartyl proteinase (Sap) of Candida albicans, which is believed to represent an important virulence factor of this opportunistic yeast, and the human immunodeficiency virus type 1 (HIV-1) protease, which is obligatory for the production of infectious virions, both belong to the same family of aspartyl proteinases. We have previously shown that the HIV-1 protease inhibitor Indinavir directly inhibits secretion and proteinase activity of Sap in a dose-dependent manner. Furthermore, at very high concentrations, viability of C. albicans is markedly reduced by Indinavir, indicating that HIV-1 protease inhibitors may possess antifungal activity. We thus proposed that these drugs may add to the resolution of mucosal candidiasis in HIV-1 infected subjects. We have now compared three different HIV-1 protease inhibitors. The rank order of Sap inhibition, already significant at 0.1 mg/ml for all protease inhibitors, was Ritonavir > Indinavir > Saquinavir. However, the cross-reactivity of Ritonavir to pepsin was also more pronounced compared with the other two. Indinavir did not affect Candida viability at concentrations up to 1 mg/ml, in line with our previous study. In contrast, at this concentration Saquinavir was even fungicidal as assessed by three different viability assays (colony formation assay, MTT assay, propidium iodide staining) whereas Ritonavir significantly affected the mitochondrial activity only (MTT assay). No influence on Candida viability was observed for any of the three at concentrations of 0.1 mg/ml or lower. It remains to be examined whether HIV-1 protease inhibitors or derivatives thereof may be suitable for in vivo therapy of subjects suffering from mucosal candidiasis resistant to current antimycotics. PMID- 10532287 TI - Evaluation of immune memory of human lymphocytes engrafted in SCID mice. AB - Peripheral blood lymphocytes (PBL) isolated from 17 healthy subjects were engrafted i.p. each to an individual SCID mouse. After 48 hours the mice were tested for skin response to the common recall antigen PPD at various sites on the body periphery. In 8 of them PPD elicited a distinct positive skin reaction only at the abdomen. This procedure could provide a basis for detection of specific cellular immune response such as in autoimmune diseases. PMID- 10532288 TI - What are allergens? PMID- 10532289 TI - Genetic and ethnic factors in allergy and asthma. PMID- 10532290 TI - To E or not to E? PMID- 10532291 TI - Prognostic value of immunologic parameters in neonates. PMID- 10532292 TI - Allergy and the environment. PMID- 10532293 TI - Allergic inflammation: cellular aspects. PMID- 10532294 TI - Allergic inflammation in the nose: mediators and adhesion molecules. PMID- 10532295 TI - Allergic inflammation: skin. PMID- 10532296 TI - Allergic airway disease: is asthma inevitable? PMID- 10532297 TI - Watch what you eat. PMID- 10532298 TI - T cells in allergy and anergy. PMID- 10532299 TI - Immunologic mechanisms of specific immunotherapy. PMID- 10532300 TI - T-cell reactivity of modified allergens. PMID- 10532301 TI - Allergen-specific immunotherapy in the management of allergic rhinitis. PMID- 10532302 TI - Specific immunotherapy in asthma. PMID- 10532303 TI - How can animal models lead to improved specific immunotherapy (SIT)? PMID- 10532304 TI - Oral and sublingual specific immunotherapy (SIT). PMID- 10532305 TI - Recombinant allergens for specific immunotherapy. PMID- 10532306 TI - Peptide immunotherapy. PMID- 10532307 TI - DNA vaccines. PMID- 10532308 TI - The future of immunotherapy. PMID- 10532309 TI - Response of Djun and Dfos mRNA abundance to signal transduction pathways in cultured cells of Drosophila melanogaster. AB - The mammalian proto-oncogenes c-jun and c-fos are situated at the end of multiple signal transduction pathways and activation of their products Jun and Fos, components of the transcription factor AP-1, are able to regulate gene transcription in response to extracellular stimuli. Djun and Dfos, the products of the Drosophila proto-oncongenes Djun and Dfos, are similar in size and sequence to their mammalian counterparts c-Jun and c-Fos and are related to their mammalian counterparts by their antigenic properties. However, very little is known about how they are regulated through signal transduction pathways. This paper has investigated the response of their mRNA abundance levels to three signal transduction pathways in Drosophila cultured cells. Various agonists and antagonists that stimulate and inhibit specific enzymes in the pathways have been tested. The results suggest that Djun and Dfos mRNA are continuously expressed and their abundance levels are transiently regulated by multiple signaling pathways, the peak response coming at 1-2 hours after perturbation. Dfos is more highly regulated than Djun which is only modulated. The receptor tyrosine kinase pathways positively regulate Dfos and Djun. The cAMP-mediated pathway positively regulates Dfos but negatively regulates Djun. The protein kinase C-activated pathway does not affect Djun whereas it negatively regulates Dfos. PMID- 10532311 TI - Trace 5-methylaminomethyl-2-selenouridine in bovine tRNA and the selenouridine synthase activity in bovine liver. AB - We measured the amount of Se in bovine liver tRNA. tRNA was chromatographed on a BD-cellulose column and Se-rich tRNA was eluted from the column in front of a main tRNA peak. There was 0.3 mmol Se/mol of tRNA and this level is about one tenth that of Escherichia coli tRNA. This suggests the presence of an enzyme that modifies tRNA with Se in bovine liver. We isolated the activity of this enzyme (selenouridine synthase) by chromatography of bovine liver extracts on a DEAE cellulose column. ATP and selenophosphate synthetase, as well as selenouridine synthase and tRNA, were necessary for the reaction. 75Se was used to label the reaction products, which were analyzed by TLC after digestion with ribonuclease T2. The position of the 75Se-nucleotide on a TLC plate was identical to that of the Se-nucleotide, 5-methylaminomethyl-2-seleno-Up, prepared from 75Se-tRNA in E. coli. PMID- 10532310 TI - A hemizygous short tandem repeat polymorphism 3' to the human phosphoglycerate kinase gene. AB - The human phosphoglycerate kinase (PGK) gene is located within Xq11-Xq13, a region implicated in genitourinary diseases including: prostate cancer, androgen insensitivity, perineal hypospadias, and other genetic abnormalities. The PGK gene and the androgen receptor gene are in linkage disequilibrium. PGK has been mapped extensively for nuclease-sensitive sites, methylation sites, and flanking DNA sequences. A PGK-associated BstXI polymorphism has been used to determine clonality of neoplastic tissues. Using fluorescent PCR product analysis and DNA sequencing, we discovered that a short tandem repeat (STR) in the 3' flanking region of the PGK gene is polymorphic. Among 231 individuals, there were nine distinct alleles, including eight based on variations in the number of TATC repeats. The PGK STR demonstrated hemizygosity, consistent with its X-chromosomal location and with an absence of cross-hybridizing autosomal homologs. The polymorphic PGK STR shows promise for rapid investigation of neoplastic clonality, for personal identification, and for studies of inherited predisposition to urologic disorders. PMID- 10532312 TI - Interaction between Wiskott-Aldrich Syndrome protein (WASP) and the Fyn protein tyrosine kinase. AB - Wiskott-Aldrich Syndrome (WAS) is a severe X-linked disorder characterised by immune deficiency, thrombocytopenia and eczema, resulting from abnormalities in a range of haematopoietic cell types. The protein that is defective in WAS, named WASP, appears to be involved in regulating changes in the cytoskeletal organisation of haematopoietic cells in response to external stimuli. In support of this idea, WASP has been found to be physically associated in haematopoietic cells in vivo with a number of SH3 domain-containing proteins involved in signal transduction, including the cytoplasmic protein-tyrosine kinase Fyn. Here, we have used a baculovirus expression system to explore the biochemical consequences of the interaction between WASP and Fyn. We find that the kinase activity of Fyn is stimulated as a result of binding to WASP, and that a cellular protein, which may be WASP itself, becomes phosphorylated on tyrosine as a result of the binding of WASP to Fyn. PMID- 10532313 TI - Age-dependent degradation of amyloid precursor protein in the post-mortem mouse brain cortex. AB - We have examined the degradation of amyloid precursor protein (APP) in the brain cortex of adult (24 +/- 2) and old (58 +/- 2) mice at different post-mortem time intervals (0, 1.5, 3, 6, 12 and 24 h). The brain cortex extract was prepared and processed for immunoblotting using antibodies against N-terminal 47-62 amino acids (Asp29) and central 301-316 amino acids containing Kunitz protease inhibitor (KPI) domain (Asp45) of APP. Asp29 (N-terminal) recognizes two bands of 140 and 112 kDa. The amount of 140 kDa is relatively higher in adult than old. The level of 112 kDa is 1.6 times lower in adult than old. It shows no remarkable change with varying post-mortem time. On the other hand, Asp45 (KPI) detects two bands of 110 and 116 kDa. While 116 kDa disappears rapidly after death of the animal, 110 kDa shows no remarkable change with different post-mortem periods. Further incubation of the disrupted tissue at 4 degrees C for 24 h and immunoblot analysis with Asp29 (N-terminal) shows 112 kDa in both ages but 58.5 kDa in adult and 70 kDa in old only. Analysis with Asp45 (KPI) shows only 54 kDa which increases after 3 h in adult but decreases significantly after 1.5 h and becomes undetectable at 24 h in old. Thus the present findings indicate that APP is degraded in a precise pattern and it depends on cellular intactness, post-mortem period and age of the animal. PMID- 10532314 TI - The homeodomain transcription factor CDP/cut interacts with the cell cycle regulatory element of histone H4 genes packaged into nucleosomes. AB - The homeodomain transcription factor CDP/cut contains four separate DNA binding domains and interacts with large segments of DNA. Thus, CDP/cut has the potential to function as an architectural protein and perhaps to support modifications in chromatin structure and nucleosomal organization. To begin to examine the ability of CDP/cut to interact with chromatin, we analyzed binding of CDP/cut to the histone H4 gene promoter (-90 to +75) reconstituted into nucleosome cores. The 90 to +75 region encompasses the cell cycle regulatory element (Site II) that controls histone H4 gene transcription, a CDP/cut binding site and a nuclease hypersensitive region. Using electrophoretic mobility shift assays and DNase I footprinting experiments, we show that CDP/cut specifically interacts with its recognition motif in a nucleosomal context without displacing the nucleosome core. The competency of CDP/cut to interact with nucleosomes suggests that this transcription factor may facilitate chromatin remodeling in response to cell cycle regulatory and/or developmental cues. PMID- 10532315 TI - An unusual arrangement of pur and lpx genes in the photosynthetic purple sulfur bacterium Allochromatium vinosum. AB - The nucleotide sequence of a 1634 bp DNA fragment from the photosynthetic purple sulfur bacterium Allochromatium vinosum contains one complete and two partial open reading frames. Sequence comparisons to genes from other organisms suggest that this A. vinosum DNA fragment contains, starting from the 5' end, the following: (1) 234 bp at the 3' end of the A. vinosum purH gene, coding for 78 amino acids at the C-terminus of the bi-functional 5'-phosphoribosyl-5 aminoimidazole-4-carboxamide formyltransferase/IMP cyclohydrolase (EC 2.1.2.3), an enzyme involved in de novo purine biosynthesis; (2) 777 bp of the A. vinosum lpxA gene, coding for all 259 amino acids of the UDP-N-acetylglucosamine-O acyltransferase, an enzyme involved in lipid A biosynthesis; and (3) 567 bp at the 5' end of the A. vinosum purD gene, coding for 189 amino acids at the N terminus of 5'-phosphoribosyl glycinamide synthetase (EC 6.3.4.13), a second enzyme involved in de novo purine biosynthesis. The presence of a gene coding for an enzyme involved in lipid A biosynthesis between two genes coding for enzymes of the de novo purine biosynthesis pathway represents a unique arrangement of these genes. PMID- 10532316 TI - The short 5' untranslated region of the betaA3/A1-crystallin mRNA is responsible for leaky ribosomal scanning. AB - Leaky ribosomal scanning allows the expression of multiple proteins from a single mRNA by occasionally skipping the first start codon, and initiating translation at a subsequent one. BetaA3- and betaA1-crystallin, two members of the beta crystallin family of vertebrate eye lens proteins, are produced via this mechanism, of which, until now, only very few examples have been found in eukaryotic genes. Since the two start codons on the betaA3/A1 messenger lie in the same reading frame, the two translated proteins are identical, except for the 17 residues shorter N-terminal extension of betaA1-crystallin. It has been suggested that the very short leader (5-7 nucleotides) of the betaA3/A1 messenger might cause slippage at the first start codon, although the unfavorable context of this start codon might also be responsible. Using transient transfections, we now demonstrate that increasing the length of the leader sequence to 67 nucleotides indeed completely abolishes translation initiation at the second start codon, and thus expression of the betaA1-crystallin protein. Messengers having a leader of 5, 7 or 14 nucleotides all express both betaA3- and betaA1 crystallin at very similar relative levels. PMID- 10532317 TI - Site-specific recombination in mammalian cells expressing the Int recombinase of bacteriophage HK022. AB - The int gene of bacteriophage HK022, coding for the integrase protein, was cloned in a mammalian expression vector downstream of the human cytomegalovirus (CMV) promoter. Green monkey kidney cells (COS-1) and mouse embryo fibroblast cells (NIH3T3) transiently transfected with the recombinant plasmid express the integrase protein. Co-transfection of this plasmid with reporter plasmids for site-specific recombination and PCR analyses show that the integrase promotes site-specific integration as well as excision. These reactions occurred without the need to supply integration host factor and excisionase, the accessory proteins that are required for integrase-promoted site-specific recombination in vitro as well as in the natural host Escherichia coli. PMID- 10532318 TI - Spatial and temporal distribution of selected canine cancers in Michigan, USA, 1964-1994. AB - Although rates are commonly used to compare regional disease occurrence, rate independent methods might also be useful in circumstances where geographic occurrence of a disease is known, but calculation of disease rates is not feasible. This is frequently the case for diseases in companion animals, where accurate enumeration of populations-at-risk is often arduous. This study had two objectives: to demonstrate a rate-independent method for investigating disease aggregation in companion animals; and, to assess the spatial and temporal clustering of canine cases of four cancers that are biologically similar in dogs and humans. Geographic information systems and point-pattern analysis were used to assess the spatial and temporal clustering of incident cases of four types of canine cancer in three counties in Michigan between 1964 and 1994, and to generate hypotheses concerning disease aggregation. Significant (P < or = 0.01) spatial clustering was found that varied by county and cancer type. No definitive temporal patterns could be deduced from a temporal analysis of the cases of canine cancer in this study. These results demonstrate distance-based methods for assessing clustering of disease, and suggest that processes determining the aggregation of canine cancer cases do not act in a spatially uniform manner. PMID- 10532319 TI - Predicting the effect of vaccination on the transmission dynamics of heartwater (Cowdria ruminantium infection). AB - We used a mathematical description of the transmission dynamics of the tick-borne infection Cowdria ruminantium in commercial beef enterprises in Zimbabwe to consider the potential impact of a candidate vaccine to prevent heartwater. The important characteristics of the vaccine were (1) a delay in development of full protection, (2) prevention of clinical disease but not of infection and (3) a waning period of protection in the absence of challenge. We considered three different scenarios in which the vaccine might be used: prophylactically in susceptible cattle prior to the introduction of infection into a herd; in susceptible cattle in the face of an epidemic (i.e., when the infection is introduced and disease is first noticed); and at equilibrium (i.e., when parasite, vector and host have been co-existing for some time). The epidemic rise in infection was modelled assuming two different patterns (i.e., resulting from slow and fast increases in tick challenge). Vaccination (administered both in the face of an epidemic and prophylactically) reduced and delayed the peak of the epidemic. With insufficiently frequent revaccination, this can result in the epidemic occurring during a period of susceptibility, so that the benefit derived from a more-efficacious vaccine is lower than that from a less-efficacious vaccine. A vaccine of only 30% or 50% efficacy (if given to the whole herd) can have important effects on both morbidity and mortality if administered with sufficient frequency. However, a highly efficacious vaccine (e.g., 90%) can have only minimal effect if revaccination occurs too infrequently - especially if the epidemic of disease occurs when tick challenge is high and vaccination-related immunity has waned. There was a fairly consistent pattern of decreasing returns on increasing protection, although this was reversed in the situation of annual vaccination undertaken prophylactically combined with an epidemic of infection that occurred when the tick challenge was relatively low. Vaccination in equilibrium situations was most beneficial at low and intermediate tick challenges. There was very little effect of vaccination in high-transmission areas regardless of vaccine efficacy and/or frequency of revaccination because most animals were infected during periods of innate or maternally derived immunity (i.e., under endemic stability). Our results suggest that where relatively high tick challenge can be achieved and consistently maintained, vaccination may be used in susceptible herds to minimise losses in a policy of transition to endemic stability. PMID- 10532320 TI - Influence of Trypanosoma evansi infection on milk yield of dairy cattle in northeast Thailand. AB - Effect of subclinical Trypanosoma evansi infection on the milk yield of newly introduced Holstein Friesian dairy cattle were investigated. Five hundred pregnant heifers were introduced in Loei Province, northeast Thailand and a total of 168 blood samples were collected at 20 farms during 6 visits over 2 years. Trypanosomes were found in cattle in June and November 1996, after which the parasite was rarely seen. On the other hand, the infection prevalences by antigen detection ELISA (Ag-ELISA) were around 40% from the first sampling through October 1997; then, antigenemic cattle decreased to 20% by June 1998. Milk yields of the cattle with detectable parasitaemia in June and November 1996 were significantly lower than those of the non-infected cattle by Student's t-test. Similarly, the milk yields of Ag-ELISA positive cattle were lower than those of negative cattle at every sampling and significant differences were observed during the first year and in February, 1998 (tested by 2-way ANOVA; T. evansi status and herd as factors). This study suggested that subclinical trypanosomosis caused decrease in milk yield of newly introduced dairy. PMID- 10532322 TI - Change in lameness risk estimates in piglets due to the modelling of herd-level variation. AB - In a previous study (Christensen, 1996. Prev. Vet. Med. 26, 107-118), an effect parameter changed from positive to negative depending on the model used. The study considered lameness in suckling piglets and the dataset included 7632 litters from 35 herds from the Health and Production Surveillance (HEPS) system database. To further investigate the change in the effect parameter, we performed a simple test to demonstrate the presence of herd-level variation. Finding substantial herd-level variation, we have re-analysed the data with special attention to the herd-level variation, and how to account for it. When herd-level variation was not accounted for in the model, a detrimental effect of prior treatment of the sow was seen but this turned into a beneficial effect when we accounted for the herd-level variation. The treatment of sow refers to any therapeutic treatment given to the sow after farrowing but before lameness or weaning occurred. This is, therefore, an example where not only the variances but also the effect parameters changed when we accounted for herd-level variation. PMID- 10532321 TI - The usage of veterinary antibacterial drugs for mastitis in cattle in Norway and Sweden during 1990-1997. AB - The prescribing patterns and annual incidence of use of antibacterial drugs for the treatment of mastitis in cattle in Norway and Sweden during the period 1990 1997 were estimated from drug wholesaler statistics. Although the drugs included in this study are also used in other species and/or other indications, mastitis in cattle is by far the most-common indication for their use. We used these sales figures to evaluate trends in the use of antibacterial drugs and changes in the incidence of treatment in bovine mastitis in Norway and Sweden. To facilitate comparisons (correcting for differences in activity and dosage) between the relative proportions of antibacterial drugs used in bovine mastitis, we introduced defined daily dose cow (DDDcow) as unit of measurement. Tentative DDDcow for the various injectable drugs were derived from doses recommended in Norway and Sweden. For intramammary drugs administered in the form of single-dose applicators, one applicator was defined as the DDDcow. The prescribing patterns of antibacterial drugs in bovine mastitis in Norway and Sweden during the study period seemed to be influenced by treatment policies, substances and formulations approved and treatment cost; length of the withdrawal period also seemed to play a role. PMID- 10532323 TI - Chiral separations in capillary electrophoresis. AB - The marked increase in the number of communications on the utilization of electrophoresis for practical chiral separations within the last three years is the most evident, and the most important fact. It reveals that the basic period of intensive research in the field is finished. The search for chiral selectors discriminating racemates in a reasonably analytical manner and the study of both the mechanism and physicochemical aspects of the chiral discrimination process were the main features of that period. Here, we review the state of the art in the field and state the references of the related literature up to the end of 1998. PMID- 10532324 TI - Selector-selectand interactions in chiral capillary electrophoresis. AB - This review discusses selected aspects of selector-select and interactions in chiral capillary electrophoresis (CE). Studies performed using nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry (MS) and X-ray crystallography for a better understanding of chiral recognition mechanisms in CE are summarized. The theoretical background of chiral CE in general, mathematical models, method development and optimization strategies, etc., are not covered. A general overview on the most recent developments in chiral CE is presented in this volume in the review paper by Bocek [1]. PMID- 10532325 TI - Enantiomeric separation by capillary electrophoresis using a crown ether as chiral selector. AB - This paper reviews chiral separations of primary amines by capillary electrophoresis and crown ether as chiral selector. Two possible mechanisms of chiral recognition by host-guest complexation are discussed: (i) The substituents of the crown ether act as barriers for the guest compounds, and (ii) lateral electrostatic interactions between host and guest occur. Experimental conditions affecting the separation are discussed in detail. A literature overview of practical applications is presented as well. More than 80 different primary amines were analyzed, whereupon the majority could be resolved using a screening method. It is shown that a synergistic effect on the resolution of chiral amines is observed when the chiral crown ether and cyclodextrins are simultaneously used in the same buffer system. This approach opens interesting perspectives for further method optimization. PMID- 10532326 TI - Vinylpyrrolidine-beta-cyclodextrin copolymer: a novel chiral selector for capillary electrophoresis. AB - The synthesis and characterization of a novel polymer consisting of an alkyl backbone and pendant beta-cyclodextrin units, obtained by radical copolymerization of vinylpyrrolidone and methacryloyl-beta-cyclodextrin (PVP-beta CD), was reported. The ability of this copolymer to act as a capillary electrophoresis (CE) chiral selector was investigated in the separation of a mixture of basic drugs. The influence of polymeric cyclodextrin concentration, temperature, and pH on the separation of the test analytes was assessed and the advantage of using the polymeric selector over native beta-cyclodextrin was demonstrated. PMID- 10532327 TI - Histamine-modified cationic beta-cyclodextrins as chiral selectors for the enantiomeric separation of hydroxy acids and carboxylic acids by capillary electrophoresis. AB - The enantiomeric separation of alpha-hydroxy acids and carboxylic acids was successfully performed by using 6-deoxy-6-N-histamino-beta-cyclodextrin (CD-hm), a monosubstituted positively charged beta-cyclodextrin (beta-CD) bearing a histamine moiety linked to the C6 of a glucose unit in the upper CD rim via the amino group. Good results were obtained at a low selector concentration (1 mM). The number of positive charges on the upper rim may be modulated as a function of pH, because of the different pKa of the amino and the imidazolyl groups, and was found to affect both the enantioselectivity and resolution factors. With the analogous 6-deoxy-[4-(2-aminoethyl)imidazolyl]-beta-cyclodextrin (CD-mh) bearing the histamine moiety linked to the C6 via the imidazolyl group, very poor results were obtained, showing that the proximity of the positive charge to the cavity plays an important role in the enantiomeric recognition. The complexation mode was studied by electrospray ionization-mass spectrometry (ESI-MS) and two dimensional nuclear magnetic resonance (2-D NMR) ROESY experiments: the recognition model is consistent with an inclusion complexation of the aromatic ring of the analyte within the CD cavity coupled to electrostatic interactions between the carboxylate and the protonated amino group of the cyclodextrin. PMID- 10532328 TI - Comparison of sulfobutylether- and sulfated-beta-cyclodextrins as additives for the chiral separation of basic spirobenzopyrans by capillary electrophoresis. AB - Three charged substituted beta-cyclodextrins (beta-CDs), sulfobutylether-beta (SBE-beta-CD), degree of substitution (DS) 4 and 7), and sulfated-beta-(S-beta CD) cyclodextrins, were compared as chiral additives in capillary electrophoresis for the enantiomeric separation of basic spirobenzopyran derivatives (pKa 9.9) which differ from each other by an N-alkyl group. The number of sulfobutylether groups attached to the cyclodextrin moiety significantly influences the enantioseparation of the basic drugs. SBE-beta-CD (DS 7) which is more strongly bound to cationic analyte than SBE-beta-CD (DS 4.6), requires smaller concentrations to achieve the same resolution. Besides, better enantioresolutions were obtained with S-beta-CD rather than with SBE-beta-CDs though higher concentrations are required, which led to high current values. However, both pairs of enantiomers cannot be resolved using S-beta-CD while SBE-beta-CDs make it possible to resolve simultaneous enantioseparation of such solutes slightly differing in hydrophobicity. This supports the hypothesis that hydrophobic interactions (outside of the CD cavity) between the butyl group attached to SBE beta-CD and the N-alkyl group of spirobenzopyran play a role in the enantioseparation. On the other hand, the sulfate group of S-beta-CD was directly attached to the CD moiety which means that the S-beta-CD-drug complexation mechanism arises through the combination of electrostatic and hydrophobic (inside the CD cavity) interactions. Finally, enantiomers of spirobenzopyran drugs were satisfactorily resolved by CE using a 20 mg/mL S-beta-CD concentration (resolution 4.0), 7 mg/mL SBE-beta-CD DS 4 (resolution 1.3), or 5 mg/mL SBE-beta CD DS 7 (resolution 3.3) added to the phosphate buffer (pH 2.6, 50 mM ionic strength). PMID- 10532329 TI - Application of sulfobutylether-beta-cyclodextrin with specific degrees of substitution for the enantioseparation of pharmaceutical mixtures by capillary electrophoresis. AB - In this research the separation of the enantiomers of the basic drug bidisomide (SC-40230) from five closely related known process impurities was investigated using several neutral and anionic sulfobutylether beta-cyclodextrins (SBE-beta CDs) as isomer selectors. Several novel sulfobutylether derivative mixtures and purified charge types having a specific degree of substitution were used to study the effect of selector charge on the efficiency and selectivity of both chiral and achiral separations. The effects of run buffer pH, selector type, and selector concentration on the chiral separation of bidisomide and the achiral separation of the related process impurities was also investigated. The related process impurity, SC-47500, displayed significant peak tailing with SBE-beta-CD mixtures which contained mono- to deca-substituted cyclodextrins. This problem was explored using isolated SBE-beta-CD charge types having degrees of substitution from one to seven. Peak tailing increased as the charge on the selector increased, suggesting that the distortion was due to electrodispersion and the large countercurrent mobility of the negatively charged complexes. Pure charge types having a lower degree of substitution provided adequate chiral and achiral selectivity, while eliminating the severe peak distortion caused by electrodispersion. The complete analysis of the bidisomide enantiomers and the related impurities was achieved with a pH 2.5 running buffer containing 5-10 mM of the isolated sulfobutylether charge types SBE[2]ds(1)sr-beta-CD or SBE[3]ds(1)sr-beta-CD. These conditions gave baseline resolution of bidisomide enantiomers and all five impurities, thus allowing both chiral and achiral purity to be determined in a single run. PMID- 10532330 TI - Comparison of highly sulfated alpha-, beta-, and gamma-cyclodextrins and 18-crown 6-tetracarboxylic acid for the enantiomeric separation of some amino acids and derivatives by capillary electrophoresis. AB - 18-Crown-6-tetracarboxylic acid (18C6H4) and highly sulfated cyclodextrins (HS alpha-, beta-, gamma-CDs) are highly selective chiral selectors for the enantioseparation of solutes bearing the primary amino function. Excellent resolutions were obtained for all solutes on HS-gamma-CD and on 18C6H4. The former, however, is by far the best chiral selector for the solutes studied in this work because the highest resolution is obtained with the shortest migration times. The reversal of the D- and L-migration order on HS-CDs compared to 18C6H4 is an interesting feature for the determination of enantiomeric excess. PMID- 10532331 TI - Enantioseparation of beta-blockers with two chiral centers by capillary electrophoresis using sulfated beta-cyclodextrins. AB - Capillary electrophoresis methods for the enantioresolution of two beta-blockers possessing two chiral centers--labetalol and nadolol--were developed using electrokinetic chromatography. These methods were based on the addition of sulfated beta-cyclodextrins (S-betaCD) as chiral selectors to the background electrolyte (BGE). Different operating parameters (pH and ionic strength of the BGE, concentration of S-beta-CD) were investigated using a normal or reversed polarity mode. A complete resolution of the four isomers of labetalol was obtained either at the cathode or at the anode according to the pH of the BGE. The resolution of nadolol was observed whatever polarity of the applied voltage but a baseline separation of the four enantiomers within a time of analysis appropriate to routine assay was only obtained at the anode. This optimal separation was performed using high concentrations of chiral additive in an acidic pH buffer of low molarity. Besides the complete enantiomeric separation of the beta-blockers studied, the interest of the proposed methods is to permit a reversal of the migration order of the different enantiomers. This could be of high interest in quality control for the study of enantiomeric purity, which is now required for the development of drugs and chemicals. PMID- 10532332 TI - When a truck becomes a motorcycle: the impact of sample load on a chiral capillary electrophoresis separation using mixtures of neutral and sulfated cyclodextrins. AB - Chiral capillary electrophoresis (CE) separations are useful for monitoring the presence of a minor isomer at low levels (e.g., <0.5%) in the presence of the major form. In order to quantitate these low levels, it is necessary to inject large amounts of sample. Separations which appear to have more than enough resolution ("big enough to drive a truck through) for dilute, equal-concentration mixtures of isomers can become inadequately resolved when the necessary amount of sample is injected. This paper addresses some important considerations in maintaining adequate resolution at high sample loads for chiral separations involving a dual-cyclodextrin (CD) system. For hydrophobic compounds, the use of both a neutral and a sulfated CD can be helpful in achieving a chiral separation. In such a system, the migration time and resolution can be controlled by varying the ratio of neutral to charged CD concentrations. It is demonstrated here that not only the ratio, but also the total CD concentration can significantly affect the separation. In this paper, the impact of the total CD concentration in a dual CD system (with the concentration ratio constant) is examined with respect to peak shape and resolution. The influences of temperature, capillary diameter, and current are also considered. The corresponding impact on the amount of sample which can be loaded and successfully separated determines the limit of quantitation of the minor isomer. Thus, this information is important in making such chiral separations applicable to determinations of low levels of minor isomer in the presence of large amounts of the major form. PMID- 10532333 TI - Binding constant dependency of amphetamines with various commercial methylated beta-cyclodextrins. AB - Cyclodextrins play an important role in enantioselective separations. They represent the major class of chiral selectors used by capillary electrophoresis. Unfortunately, the purity of commercial cyclodextrins is often not well characterized, and similar selectors sold by various suppliers may show totally different enantioselectivities. In this study, the composition of several commercial methylated-beta-cyclodextrins is evaluated by means of previously developed analytical methods. Then, different calculation methodologies, such as graphical determinations, as well as nonlinear or linear regression approaches, are evaluated in order to calculate the binding constants of inclusion complexes formed by some amphetamine derivatives with methylated-beta-cyclodextrins. The nonlinear curve-fitting methodology proves to be the most suitable for these determinations. Comparisons are made between the different selectors and several hypotheses are proposed concerning the formation of the inclusion complex. Finally, enantiomeric resolutions are evaluated for these selectors and conclusions drawn about the knowledge of selector composition. PMID- 10532334 TI - Potential of flow-counterbalanced capillary electrophoresis for analytical and micropreparative separations. AB - The potential of flow-counterbalanced capillary electrophoresis (FCCE) in chiral and achiral separations was investigated in this work. Unlimited increase of the separation selectivity can be achieved for binary mixtures using FCCE. This was shown for the enantioseparation of (+/-)-chlorpheniramine (CHL) with carboxymethyl-beta-cyclodextrin (CM-beta-CD) as chiral selector. The other example is the separation of alpha- and beta-isomers of a dipeptide aspartame (AS). The carrier ability of the (chiral) selector or pseudostationary phase, the electroosmotic flow (EOF), the pressure-driven flow or hydrodynamic flow can be used as a counterbalancing flow to the electrophoretic mobility of the analyte or vice versa. This mechanism can also be used for micropreparative purposes. FCCE also bears the potential for stepwise separation and fraction collection of multicomponent mixtures. PMID- 10532335 TI - Cyclodextrins as chiral selectors in capillary electrophoresis: a comparative study for the enantiomeric separation of some beta-agonists. AB - A comparative study for the enantiomeric separation of terbutaline, clenbuterol, salbutamol and dobutamine was performed by capillary electrophoresis using cyclodextrins and their derivatives as chiral selectors. Several parameters such as buffer composition and temperature were studied. Simple, fast and reliable enantioseparations were achieved for all drugs studied, especially when the isomerically pure sulfated beta-cyclodextrin derivatives were used as chiral selectors. PMID- 10532336 TI - Designed combination of chiral selectors for adjustment of enantioseparation selectivity in capillary electrophoresis. AB - In this study an attempt has been made to explain the reasons for changing the enantioseparation selectivity in some dual cyclodextrin (CD) systems compared to the use of single chiral selectors in capillary electrophoresis (CE). An explanation for selectivity changes is proposed based on the effect of the chiral selector on the mobility of the analyte. In order to support the proposed mechanism, several dual systems were designed on the basis of the known recognition pattern of enantiomers for individual CDs. In most cases the separation selectivity could be adjusted in a designed way. There was no experimental evidence for simultaneous binding of a given chiral analyte with both chiral selectors or of chiral recognition of an analyte complex with one CD by another CD. PMID- 10532337 TI - Resolution of ephedrine derivatives by means of neutral and sulfated heptakis(2,3 di-O-acetyl)beta-cyclodextrins using capillary electrophoresis and nuclear magnetic resonance spectroscopy. AB - Beta-cyclodextrin (beta-CD), heptakis(2,3-di-O-acetyl)beta-cyclodextrin (Diac beta-CD) and heptakis (2,3-di-O-acetyl-6-sulfato)beta-cyclodextrin (HDAS-beta-CD) were tested for their ability to discriminate the enantiomers of ephedrine, pseudoephedrine, norephedrine and methylephedrine. Using capillary electrophoresis (CE) under optimized conditions, with the exception of norephedrine in presence of beta-CD, all racemates could be resolved. Utilizing Job's plot by means of UV spectroscopy revealed 1:1 complexes formed with beta-CD and Diac-beta-CD. HDAS-beta-CD gave curved plots indicating mixed stoichiometry. Inspection of the cyclodextrin-induced chemical shifts (CICS) of both the ligands and the CDs showed that the ephedrine sits deeply in the cavity of beta-CD and HDAS-beta-CD. In the case of Diac-beta-CD, the ephedrine is located closely to the wider rim of the CD cavity. In conclusion, comparing the pattern of CICS of the various CD derivatives clearly indicates the differences in the complex geometry. PMID- 10532338 TI - Some factors affecting enantiomeric impurity determination by capillary electrophoresis using ultraviolet and laser-induced fluorescence detection. AB - The key factors influencing enantiomer trace determination were investigated; these include resolution capillary diameter, limit of detection, linear range and type of detection. Chiral reagents, (+)- and (-)-1-(9-fluorenyl)ethyl chloroformate (FLEC), were employed as probes to demonstrate the influence of the variables. In order to find the best resolution, separation variables were optimized in both capillary zone electrophoresis (CZE) and micellar electrokinetic capillary chromatography (MEKC) modes by the application of factorial design experiments. A highly efficient chiral separation of the (+/-) FLEC, derivatized with nonchiral amino acids, was achieved when using gamma cyclodextrin as the chiral selector. The benefits of using a small diameter capillary for direct determination of both (+) and (-)-FLEC impurity (0.05-0.1% area/area) were demonstrated using UV detection and applying a sample stacking condition. A frequency-doubled argon ion laser (244 nm) was used as light source for laser-induced fluorescence (LIF) detection. Excitation light was provided by means of an optical fiber directed into the Hewlett Packard 3D capillary cartridge. The signals from UV and LIF were monitored simultaneously. The application of LIF detection greatly improved sensitivity and linear range. Further, as a consequence of the increased sensitivity, sample loading could be decreased, which led to an improvement of separation efficiency. Direct determination of 0.005% impurity could be achieved within the linear range. PMID- 10532339 TI - pKa shift-associated effects in enantioseparations by cyclodextrin-mediated capillary zone electrophoresis. AB - Enantioselective migration of dansylated (Dns) amino acids in the presence of hydroxypropylated-beta-cyclodextrin under acidic conditions near the pI value of the analytes was investigated by means of capillary zone electrophoresis. Based on the migration data, the pH dependence of the complexation constants was evaluated, as well as the variation of the complex mobilities with pH. As a result of these data, the migration behavior in the pH region near the pI could be understood, which, in some instances, includes the reversal of migration order upon variation of selector concentration. The enantioselective pKa shifts upon complexation could be quantitated for the carboxylic and the amino group separately. pKa shifts were found in the order of 0.8 pI units, the differences between the enantiomers being up to 0.25 pH units. These data were in agreement with the pI shifts reported from isoelectric focusing experiments. The accurate determination of the pI values of the Dns amino acids makes it possible to calibrate the pI scale in isoelectric focusing in the presence of chiral selectors. PMID- 10532340 TI - Enantiomeric purity determination of propranolol by capillary electrophoresis using dual cyclodextrins and a polyacrylamide-coated capillary. AB - The use of a chirally selective capillary electrophoresis method is reported for the enantioselective purity determination of propranolol drug substance. The method employed a combination of both charged and neutral cyclodextrin. An internally coated capillary was used to suppress electroosmotic flow and potential peak tailing. The method was capable of monitoring below 0.1% m/m of the undesired impurity. Acceptable validation data was also obtained for recovery, linearity, and for both short and long-term injection precision. PMID- 10532341 TI - Stereoselective screening for and confirmation of urinary enantiomers of amphetamine, methamphetamine, designer drugs, methadone and selected metabolites by capillary electrophoresis. AB - Data presented in this paper demonstrate that a competitive binding, electrokinetic capillary-based immunoassay previously used for screening of urinary amphetamine and analogs cannot be employed to distinguish between the enantiomers of amphetamine and methamphetamine. However, capillary zone electrophoresis with a pH 2.5 buffer containing (2-hydroxypropyl)-beta cyclodextrin as chiral selector is shown to permit the enantioselective analysis of urinary extracts containing methamphetamine, amphetamine, 3,4 methylenedioxymethamphetamine (Ecstasy) and other designer drugs, and methadone together with its major metabolite, 2-ethylidene-1,5-dimethyl-3,3 diphenylpyrrolidine. In that approach, enantiomer identification is based upon comparison of extracted polychrome UV absorption data and electropherograms obtained by rerunning of spiked extracts with spectra and electropherograms monitored after extraction of fortified blank urine. The suitability of the described chiral electrokinetic capillary method for drug screening and confirmation is demonstrated via analysis of unhydrolyzed quality control urines containing a variety of drugs of abuse. Furthermore, in a urine of a patient under selegiline pharmacotherapy, the presence of the R-(-)-enantiomers of methamphetamine and amphetamine could be unambiguously identified. Direct intake of an R-enantiomer or ingestion of drugs that metabolize to the R-enantiomers can be distinguished from the intake of S-(+)-enantiomers (drug abuse) or prescribed drugs that metabolize to the S-enantiomers of methamphetamine and amphetamine. The described approach is simple, reproducible, inexpensive and reliable (free of interferences of other major basic drugs that are frequently found in toxicological urines) and could thus be used for screening for and confirmation of urinary enantiomers in a routine laboratory. PMID- 10532342 TI - Evaluation of quail egg white riboflavin binding protein as a chiral selector in capillary electrophoresis by applying a modified partial filling technique. AB - A preliminary evaluation of the enantioselective properties of quail egg yolk riboflavin binding protein (qRfBP) was carried out in capillary electrophoresis by using the complete filling technique. The most promising results obtained by this screening of nineteen chiral drugs were singled out with the aim of optimizing enantiomer separations by applying the partial filling technique, which allows operating at much higher protein concentrations without detection problems. The building of the separation zone in the partial filling technique has been modified in order to enable on-line monitoring, before each run, of the actual protein plug application velocity and, consequently, the building of a plug of the desired length. The electrophoretic conditions chosen gave opposite migration directions for the chiral selector and the analytes, with qRfBP migrating away from the detector. A polyvinyl alcohol-coated capillary was first totally filled with protein and the optimal plug length was obtained by further applying negative pressure together with positive voltage for the time needed. Separations of basic drugs were optimized by using protein concentrations ranging from 200 microM up to 900 microM and different plug lengths, while the running buffer pH (6.0), temperature (25 degrees C) and operating voltage (+20 kV) were kept constant. The enantioresolution of all solutes was affected by both the chiral selector concentration and protein plug length. Baseline separations were obtained for oxprenolol, prilocaine and bupivacaine. PMID- 10532343 TI - Enantiomeric separations of dansyl amino acids using the macrocyclic antibiotic A35512B as a chiral selector in capillary electrophoresis. AB - The macrocyclic antibiotic A35512B was examined as a chiral selector for capillary electrophoresis (CE) using thirteen racemic dansyl amino acids as test analytes. The chiral selectivity of A35512B was evaluated as a function of the run buffer pH, antibiotic concentration, and organic modifier composition. After optimizing these parameters, the macrocylic antibiotic A35512B provided high resolutions of all the enantiomers for the thirteen dansyl amino acids tested in this study. PMID- 10532344 TI - Equilibrium binding model of bile salt-mediated chiral micellar electrokinetic capillary chromatography. AB - Optimum concentration of bile salts in chiral separations depends on both the aggregation properties of the surfactant and the stability of the analyte-micelle complexes. An equilibrium model is proposed in which these two effects are treated separately. First the aggregation constants should be determined under the experimental conditions of the chiral MEKC analysis. With these data, the equilibrium concentrations of bile salt aggregates can be calculated at any total surfactant concentration. Using the Offord equation to approximate the mobilities of the enantiomer-bile salt complexes, a model function has been derived to fit the experimental mobilities. The method yields the binding constants of the enantiomers to each aggregate present. Those species are assumed to be important in the chiral recognition process, which have significantly different stability constants for the enantiomers. The method is demonstrated by the chiral separation of R- and S-1,1'-binaphthyl-2,2'-diyl hydrogen phosphate with sodium taurodeoxycholate. Based on the calculated binding constants, tetrameric aggregates are assumed to be the discriminating species, while no significant difference in enantiomer binding to dimers was found. PMID- 10532345 TI - Chiral separation of dansyl-DL-amino acids with micellar systems containing copper (II) ion and N-n-dodecyl-L-proline in electrokinetic capillary chromatography. AB - Enantiomers of dansylated DL-amino acids were resolved by chiral copper (II)-N-n dodecyl-L-proline (1) complexes incorporated in micelles of sodium dodecyl sulfate (SDS) in electrokinetic capillary chromatography (EKC). This resolution is caused by formation of diastereomeric ternary complexes consisting of chiral ligand 1, central copper (II) ion and enantiomeric amino acid derivatives in micellar phase. However, the resolution was not observed when SDS with an anionic polar head grop was replaced with dodecyl trimethylammonium brode (DTMAB) with a cationic polar head group. The ratio between copper (II) ion and 1 in the complex in either SDS or DTMAB was measured by UV-visible spectra, which respond to the d d transition of copper (II). Mechanism of separation should be discussed in terms of effect of surfactant structures on constitution of copper (II) ion and 1 in the micellar phase and that of arene substituent structures linked to sulfonamide units in amino acid derivatives to be separated. PMID- 10532346 TI - Enantioseparations by capillary electrophoresis using chiral glycosidic surfactants. I. Evaluation of cyclohexyl-pentyl-beta-D-maltoside surfactant. AB - Chiral cyclohexyl-pentyl-beta-D-maltoside (CYMAL-5) surfactant was evaluated in the enantioseparation of charged racemic species by capillary electrophoresis. CYMAL-5 is a glycosidic surfactant (GS) with a chiral maltose polar head group and a cyclohexyl-pentyl hydrophobic tail. At concentrations above its critical micellar concentration (CMC), CYMAL-5 produces neutral micelles in aqueous media. The neutral micelles migrate at the velocity of the electroosmotic flow (EOF). As expected, the CYMAL-5 system was only useful for the enantioseparation of charged chiral solutes. The enantioresolution of the CYMAL-5 can be manipulated over a wide range of electrolyte composition, e.g., pH, ionic strength and surfactant concentration. In the presence of EOF, and in all cases, there is an optimum surfactant concentration for maximum enantioresolution, which is located at low surfactant concentration for strongly hydrophobic solutes and at high surfactant concentration for relatively hydrophilic solutes. The presence of an optimum surfactant concentration for maximum enantioresolution is attributed to the EOF. At low pH values where the EOF is negligible, enantioresolution increased with increasing surfactant concentration in the useful concentration range in a way similar to chromatography. PMID- 10532347 TI - Chiral separations in capillary high-performance liquid chromatography and nonaqueous capillary electrochromatography using helically chiral poly(diphenyl-2 pyridylmethyl methacrylate) as chiral stationary phase. AB - Enantioseparations in nonaqueous capillary electrochromatography (CEC) are reported in this study for the first time, using wide-pore aminopropyl silica gel coated with helically chiral poly(diphenyl-2-pyridylmethyl methacrylate) (PDPM) as chiral stationary phase (CSP). The anodic electroosmotic flow (EOF) in a methanolic solution of ammonium acetate was used for the migration of neutral analytes through the packed bed in the capillaries. Four different techniques, high-performance liquid chromatography (HPLC) in common-size columns, capillary HPLC, pressure-assisted CEC and CEC were compared from the viewpoint of separation parameters. The latter three were performed with the same experimental setup, varying the relative contribution of the pressure-driven and the electrically driven flow to the overall mobility of the analyte. Capillary HPLC offers clear advantages compared to enantioseparations in common-size columns. However, for a given particle size of the packing material, CEC was not obviously advantageous compared to pressure-driven separations. PMID- 10532348 TI - Enantiomer separation by pressure-supported electrochromatography using capillaries packed with Chirasil-Dex polymer-coated silica. AB - Pressure-supported electrochromatography using capillaries packed with permethyl cyclodextrin covalently linked via an octamethylene spacer to dimethylpolysiloxane and immobilized on silica (Chirasil-Dex silica) has been employed as an efficient and rapid method for the enantiomer separation of various racemic compounds. By comparing this method with micropacked liquid chromatography (LC), employing the same column in a unified instrumental setup, micropacked capillary electrochromatography (CEC) shows higher column efficiencies and hence better resolution factors. The influence of type and concentration of buffer, amount and nature of organic modifier, and pressure support is investigated. PMID- 10532349 TI - Separations of enantiomers by preparative capillary isotachophoresis. AB - The use of capillary isotachophoresis (ITP), operating in a discontinuous fractionation mode, for preparative separations of enantiomers of chiral compounds was studied. The ITP separations were carried out in the column coupling configuration of the separation unit provided with the preseparation column of a 1.0 mm ID and the trapping column of a 0.8 mm ID. Such a configuration of the CE separation unit offers several working regimes suitable to preparative separations of enantiomers. 2,4-Dinitrophenyl-DL-norleucine (DNP Norleu) was employed as a model analyte in our experiments with beta-cyclodextrin serving in the electrolyte solutions as a chiral selector. The preparative separations lasting about 20 min were evaluated by ITP and (more often) by capillary zone electrophoresis (CZE). It was found that one preparative run provided up to 14 microg of pure DNP-Norleu enantiomers. This corresponded to a 75 times higher production rate of ITP relative to a maximum value of this parameter as estimated for preparative CZE runs in cylindrical capillaries (0.5 pmol/s). About 75% of the DNP-Norleu enantiomers loaded into the preparative equipment could be recovered in pure enantiomer fractions. Contiguous natures of the zones in the ITP stack and adsorption losses of the enantiomers in the isolation step were found to set practical limits for a further enhancement of the recovery rates in the isolation of pure enantiomers. PMID- 10532350 TI - Nonaqueous capillary electrophoretic separation of basic enantiomers using heptakis(2,3-dimethyl-6-sulfato)-beta-cyclodextrin. AB - The enantiomers of 40 basic analytes, mostly pharmaceuticals, were separated by nonaqueous capillary electrophoresis in acidic methanol background electrolytes using the sodium salt of heptakis(2,3-dimethyl-6-sulfato)-beta-cyclodextrin (HDMS beta-CD). The effective mobilities, separation selectivities, and peak resolution values were determined as a function of the HDMS-beta-CD concentration in the 0 40 mM range and were found to follow the theoretical predictions of the charged resolving agent migration model (CHARM model). Fast, efficient enantiomer separations were achieved for a large number of both very hydrophobic and hydrophilic weak bases. PMID- 10532351 TI - Cellular and molecular regulation of an erythropoietic inductive microenvironment (EIM). PMID- 10532352 TI - Translocation of HSP27 and MKBP in ischemic heart. AB - HSP27 and MKBP translocate from the cytosolic to myofibril fraction in ischemic rat heart as demonstrated by immunoblotting. Immunohistochemistry analysis showed that ischemia enhances the Z line labeling of HSP27 and MKBP. Two dimensional gel electrophoresis showed that ischemia increases the hyperphosphorylated form of HSP27. These data suggest that HSP27 and MKBP may be involved in the Z line protection against postischemic reperfusion injury. PMID- 10532353 TI - Effect of HSP47 on prolyl 4-hydroxylation of collagen model peptides. AB - Prolyl 4-hydroxylation, the most important post-translational modification in collagen biosynthesis, is catalyzed by prolyl 4-hydroxylase, an endoplasmic reticulum-resident enzyme. HSP47 is a collagen-binding stress protein which also resides in the endoplasmic reticulum (Nagata, K. and Yamada, K.M. (1986) J. Biol. Chem., 261, 7531-7536). Both prolyl 4-hydroxylase and HSP47 interact with procollagen alpha-chains during their folding and/or modification in the endoplasmic reticulum. Recent study has revealed that a simple collagen model peptide, (Pro-Pro-Gly)n, is recognized by HSP47 as well as by prolyl 4 hydroxylase in vitro (Koide et al., manuscript submitted). In the present study, we investigated the effect of HSP47 on the prolyl 4-hydroxylation of such collagen model peptides. To monitor the enzymatic hydroxylation of the peptides, we developed a non-RI assay system based on reversed-phase HPLC. When HSP47 was added to the reaction mixture, substrate and less-hydroxylated materials accumulated. This effect depended on the peptide-binding activity of HSP47, because a mutant HSP47 without collagen-binding activity did not show any inhibitory effect on prolyl 4-hydroxylation. Kinetic analysis and other biochemical analyses suggest that HSP47 retards the enzymatic reaction competing for the substrate peptide. PMID- 10532354 TI - Mutation of the yeast epsilon-COP gene ANU2 causes abnormal nuclear morphology and defects in intracellular vesicular transport. AB - Previously we reported an original method of visualizing the shape of yeast nuclei by the expression of green fluorescent protein (GFP)-tagged Xenopus nucleoplasmin in Saccharomyces cerevisiae. To identify components that determine nuclear structure, we searched for mutants exhibiting abnormal nuclear morphology from a collection of temperature-sensitive yeast strains expressing GFP-tagged nucleoplasmin. Four anu mutant strains (anu1-1, 2-1, 3-1 and 4-1; ANU=abnormal nuclear morphology) that exhibited strikingly different nuclear morphologies at the restrictive temperature as compared to the wild-type were isolated. The nuclei of these mutants were irregularly shaped and often consisted of multiple lobes. ANU1, 3 and 4 were found to encode known factors Sec24p, Sec13p and Sec18p, respectively, all of which are involved in the formation or fusion of intracellular membrane vesicles of protein transport between the endoplasmic reticulum (ER) and the Golgi apparatus. On the other hand, ANU2 was not well characterized. Disruption of ANU2 (delta anu2) was not lethal but conferred temperature-sensitivity for growth. Electron microscopic analysis of anu2-1 cells revealed not only the abnormal nuclear morphology but also excessive accumulation of ER membranes. In addition, both anu2-1 and delta anu2 cells were defective in protein transport between the ER and the Golgi, suggesting that Anu2p has an important role in vesicular transport in the early secretory pathway. Here we show that ANU2 encodes a 34 kDa polypeptide, which shares a 20% sequence identity with the mammalian epsilon-COP. Our results suggest that Anu2p is the yeast homologue of mammalian epsilon-COP and the abrupt accumulation of the ER membrane caused by a blockage of the early protein transport pathway leads to alteration of nuclear morphology of the budding yeast cells. PMID- 10532355 TI - Induction of apoptosis by Coprinus disseminatus mycelial culture broth extract in human cervical carcinoma cells. AB - Extract of Coprinus disseminatus (pers. Fr.) (C. disseminatus) culture broth (EDCB) inhibits proliferation and induces apoptosis in the human cervical carcinoma cells at 5 microg/ml. To determine whether the cell death induced by the EDCB recruits caspases or not, one of the exclusive pathways in cell death, we examined caspase-3 activity in this cell death process. The activity of caspase-3 was remarkably increased when the cell was treated with EDCB, and this activity was nullified by Z-VAD-FMK, a well known caspase-3 inhibitor. From these results, we would expect the EDCB to contain substances with the ability to induce apoptosis in the human cervical carcinoma cells. The extent of the EDCB induced apoptosis is cell line-dependent. PMID- 10532356 TI - Identification and immunological characterization of a novel 40-kDa protein linked to CD98 antigen. AB - Monoclonal antibodies (mAbs) were obtained from hybridoma clones established by cell fusion between mouse myeloma cells and spleen cells from a mouse immunized against an affinity-purified 40-kDa component of rat 125-kDa glycoprotein (GP125). Two mAbs designated as 3F2 and 6B4 detected a 40-kDa and a 125-kDa band under reducing and nonreducing conditions, respectively, in extracts prepared from rat, mouse and human tumor cells. Association of the 40-kDa protein with CD98 was revealed by sandwich-type enzyme-linked immunosorbent assay. The two mAbs were strongly reactive with various tumor cells and activated lymphocytes, but were only weakly reactive with resting lymphocytes. Confocal microscopy indicated colocalization of CD98 and the 40-kDa protein defined with 3F2 and 6B4 at the cell surface and perinuclear regions. On immunohistochemical analysis of frozen sections of rat tongue, the anti-rat CD98 mAb B3 selectively stained the basal layer and 3F2 stained the upper epithelial part in addition to the basal layer, indicating the existence of CD98-unlinked 40-kDa protein. PMID- 10532357 TI - Nuclear localization of gold labeled-hydrocortisone-bovine serum albumin conjugate injected intravenously into the hormone-target cells of rat. AB - We have suggested in a previous study using 2-nm colloidal gold labeled testosterone-bovine serum albumin (testosterone-BSA-gold) that 2-nm gold labeled steroid hormone-BSA conjugates would be a useful tool for analyzing the mechanism of steroid hormone action (39). In this study, we examined whether hydrocortisone BSA conjugate (hydrocortisone-BSA) showed a similar distribution to radiolabeled hydrocortisone in vivo, by injecting 2-nm colloidal gold labeled-hydrocortisone BSA (hydrocortisone-BSA-gold) into the rat tail vein. The hydrocortisone-BSA-gold with silver enhancement became visible as silver deposits under electron microscopy in the nuclei of hepatocytes and hepatic stellate cells but not in Kupffer cells in the liver, and in the thymocytes and thymic reticuloepithelial cells in the thymus of a rat killed 2 h postinjection. The percentage of nuclei showing deposits in the non-target cells, the epithelial cells of the seminal vesicle, was similar to the value in the seminal vesicle of a control rat injected with BSA labeled with 2-nm colloidal gold as reported previously. In the hepatocytes and thymocytes of a control rat not injected, the percentages of nuclei showing deposits were similar to those in the rat injected with testosterone-BSA-gold or BSA-gold as reported previously, but lower than those in the rat injected with hydrocortisone-BSA-gold. These results suggest that hydrocortisone-BSA-gold is useful for the morphological study of hydrocortisone target cells, and imply that BSA conjugated with hydrocortisone can enter the target cell nuclei of the rat. The present study further indicates that the fate of gold labeled-steroid hormone-BSA conjugates may be decided at the cell membrane level. PMID- 10532358 TI - Descartes' error. PMID- 10532359 TI - Congenital esotropia in perspective. PMID- 10532360 TI - Dissociated vertical deviation and head tilts. AB - INTRODUCTION: The association of anomalous head posture and dissociated vertical deviation does not seem to be appreciated, as evidenced by the paucity of literature linking these two conditions. METHOD: The series describes 14 patients who had an anomalous head posture and dissociated vertical deviation. The assumed head tilts appeared to decrease the magnitude and improve the motor control of dissociated vertical deviation. RESULTS: Twelve of 14 patients tilted their heads contralateral to the eye with the dissociated vertical deviation, or away from the eye with a larger amount of dissociated vertical deviation if the disorder was bilateral. Two patients tilted their heads to the same side as the eye with the dissociated vertical deviation. Forced head tilt-testing in the opposite direction showed an increase in the magnitude of the dissociated vertical deviation or poorer control of the deviation. Dissociated vertical deviation was not related to oblique muscle dysfunction. Peripheral fusion was demonstrated in 10 patients, as evidenced by low-grade stereopsis or Worth 4 dot fusion at near. One patient did not show any demonstrable fusion with conventional tests. Another did not show evidence of stereopsis, but Worth 4 dot testing was not performed. Two other patients were too young to cooperate with sensory testing. Anomalous head posture was controlled or minimized after the control of the dissociated vertical deviation by surgery in four patients. Two patients showed improved stereopsis after surgery for dissociated vertical deviation. CONCLUSION: Dissociated vertical deviation should be included in the differential diagnosis of an ocular cause of head tilts. Forced contralateral head tilttesting will confirm whether dissociated vertical deviation is the cause if motor control of the dissociated vertical deviation worsens or becomes manifest rather than latent. The presence of an anomalous head posture in patients with dissociated vertical deviation can be improved with strabismus surgery. PMID- 10532361 TI - Abnormal head posture associated with high hyperopia. AB - BACKGROUND: An abnormal head posture may be adopted for ocular or nonocular reasons. The most common ocular reasons are to maintain binocularity and to obtain the best possible visual acuity. Patients with undercorrected or overcorrected refractive errors have been reported to adopt a variety of head positions, thought to be an attempt to obtain the best possible visual acuity. METHODS: Five patients with symmetric high hyperopia (at least + 5.00 D) and an abnormal head posture are presented. RESULTS: All five patients demonstrated an abnormal head posture of chin down for fixation without the spectacle correction in place. This abnormal head posture was eliminated by occlusion of either eye and also by wearing of the refractive correction. No patient demonstrated significant strabismus. CONCLUSION: An abnormal head posture when not wearing spectacle correction can occur in children who have high hyperopia and insignificant strabismus. This may be a mechanism by which the best visual acuity is obtained (indicated by the disappearance of the abnormal head posture on wearing of the glasses) and also to maintain binocularity (indicated by the disappearance of the abnormal head posture under monocular testing conditions). The presence of a chin-down abnormal head posture should alert the examiner to the possible presence of high hyperopia and therefore the necessity for a cycloplegic refraction. PMID- 10532363 TI - Distance stereoacuity norms for the mentor B-VAT II-SG video acuity tester in young children and young adults. AB - PURPOSE: The purpose of this study was to provide normative distance stereoacuity data for the Mentor B-VAT II-SG video acuity tester (Mentor O & O, Norwell, Mass.). METHODS: Near and distance stereoacuity for 45 normal young children (5 to 6 years old, child group) and 67 normal young adults (16 to 20 years old, adult group) were evaluated. Distance stereoacuity was measured with the Random Dot and the Circles tests on the B-VAT unit. Near stereopsis was assessed by Titmus, Randot, and TNO tests. Additionally, a random subset of the subjects was retested 1 month later with the B-VAT unit. RESULTS: The mean and SD values of stereoacuity measured on the Circles and Random Dot tests were 49 +/- 33 and 98 +/- 49 seconds of arc in children (p < 0.0001) and 50 +/- 32 and 83 +/- 51 seconds of arc in adults (p < 0.0001), respectively. No significant differences were found between the same tests with respect to age. Of 112 subjects in both groups, 110 (98%) achieved 120 seconds of arc or finer stereoacuity threshold levels on the Circles test and 108 (96%) demonstrated 180 seconds of arc or finer scores on the Random Dot test. Although all but two of the subjects exhibited stereopsis both at near and at distance, there were low correlations among the different near and distance stereotests. Test and retest distance stereoacuity scores agreed closely. CONCLUSIONS: The B-VAT II-SG system produces reliable distance stereoacuity data. The norms we obtained may aid the clinician to detect binocular visual disturbances or may provide a basis for using distance stereoacuity as a screening method. PMID- 10532362 TI - Heterotopic muscle pulleys or oblique muscle dysfunction? AB - INTRODUCTION: The description of connective tissue sleeves that function as pulleys for the rectus extraocular muscles (EOMs) suggests that abnormalities of EOM pulley position might provide a mechanical basis for some forms of incomitant strabismus. Pulleys determine the paths and thus the pulling directions of EOMs. METHODS: High-resolution magnetic resonance images spanning the orbits were obtained in primary position, upgaze, and downgaze for each subject. Paths of the EOMs were measured with reference to the orbital center and permitted inference of pulley locations. RESULTS: Data from 18 orbits of orthotropic subjects defined means and SDs of normal EOM pulley coordinates. Eight patients, aged 17 to 60 years, had heterotopic EOM pulleys, defined as displaced at least 2 SDs from normal. We found one to eight heterotopic pulleys (considering both orbits) in each of four patients who had been diagnosed with marked superior oblique (SO) overaction and mild to marked inferior oblique (IO) underaction. Each patient had superior mislocation of at least one lateral rectus pulley by 1.8 to 4.9 mm. Three patients diagnosed with mild to moderate IO overaction and mild to moderate SO underaction in only one orbit had one to three heterotopic EOM pulleys. Each of those patients had at least one lateral rectus pulley inferiorly dislocated by 1.9 to 4.9 mm. The final patient, who was diagnosed with mild IO underaction and normal SO function bilaterally, had bilateral superior mislocation of the medial rectus pulleys by greater than 2 mm. Computer simulations using the Orbit program (Eidactics, San Francisco) incorporating individually measured pulley positions reproduced the clinical patterns of incomitant strabismus in all cases without postulating abnormalities of oblique muscle innervation or contractility. CONCLUSION: Heterotopic EOM pulleys can cause patterns of incomitant strabismus that have been attributed to oblique muscle dysfunction. Even isolated mislocations of less than 2 mm, coupled with smaller mislocations of the other pulleys, can produce the clinical appearance of bilateral oblique dysfunction. Pulley heterotopy should be considered in the differential diagnosis of incomitant strabismus and oblique dysfunction. PMID- 10532364 TI - Pseudotumor cerebri in children. AB - PURPOSE: Demographic and outcome data in the era of modern neuroimaging are needed to describe pseudotumor cerebri in children. METHODS: We reviewed the medical records of children less than 18 years old who were diagnosed with pseudotumor cerebri between 1977 and 1997. We defined pseudotumor cerebri as (1) increased intracranial pressure, (2) normal or small ventricles, and (3) normal cerebrospinal fluid composition. The condition might be idiopathic or the result of a nontumor etiology. RESULTS: Thirty-seven patients had an initial diagnosis of pseudotumor cerebri. Two patients were subsequently diagnosed with a central nervous system malignancy and were excluded from further analysis. The remaining 35 patients included 10 patients with idiopathic pseudotumor cerebri and 25 patients with disorders reported to be associated with pseudotumor cerebri. The mean age was 10.6 years with a range of 3 to 17 years. Twenty patients (57%) were female and 13 patients (37%) were obese. At presentation 4 patients had a visual acuity less than 20/40 in the best eye and 10 patients had visual field deficits. Seventeen patients (49%) had cranial nerve deficits, all of which resolved with normalization of the intracranial pressure. Follow-up data were obtained on 30 patients. Only one patient had a final visual acuity less than 20/40 in the best eye, whereas six patients had residual visual field deficits. Ten patients (33%) had optic nerve atrophy. CONCLUSIONS: There was no gender predominance, and associated etiologic factors were common in these children with pseudotumor cerebri. Permanent visual loss occurs in some children with pseudotumor cerebri. Quantitative perimetry and optic nerve examination were more sensitive than visual acuity determination in detecting damage to the visual sensory system. In rare instances the patient diagnosed with pseudotumor cerebri will be found after extended follow-up to harbor an intracranial neoplasm. PMID- 10532365 TI - The effect of buphthalmos on orbital growth in early childhood: increased orbital soft tissue volume strongly correlates with increased orbital volume. AB - PURPOSE: Our purpose was to evaluate the effect of increased orbital soft tissue volume on orbital growth. METHOD: Patients with unilateral or significantly asymmetric bilateral buphthalmos as determined by axial computed tomography scan were recruited. Volumetric determinations of the bony orbit with use of axial 1.5 mm sections on computed tomography were undertaken. Statistical analysis of the paired ocular length measurement and bony orbital volume measurements for each patient were performed. RESULTS: Eight patients (mean age 41 months) with a 15% or greater difference in axial length were enrolled. The mean axial length of the buphthalmic globes was 23% greater than that of the contralateral globes. Orbits harboring a buphthalmic globe had an orbital volume 11% greater than on the contralateral side. CONCLUSION: Increased orbital soft tissue volume as evidenced by buphthalmos was significantly associated with enlarged bony orbital volume. This indicates that soft tissue volume is a determinant of orbital volume and suggests that orbital tissue expanders might enhance bony development in patients with anophthalmos or microphthalmos and after early enucleation. PMID- 10532366 TI - Topical versus oral carbonic anhydrase inhibitor therapy for pediatric glaucoma. AB - PURPOSE: Our purpose was to compare, in a crossover design,the hypotensive effect of oral acetazolamide (Diamox) and topical dorzolamide (Trusopt) in patients with pediatric glaucoma. METHODS: All patients less than 18 years old who were switched from acetazolamide to dorzolamide without other intervention were reviewed. Intraocular pressures were obtained with either a Tono-Pen (Mentor Ophthalmics, Santa Barbara, Calif.) or applanation tonometer. Minimum follow-up times on acetazolamide and on dorzolamide were 1 month (mean 12.2 +/- 19.7 months) and 2 months (mean 8.2 +/- 5.1 months), respectively. The average dose of acetazolamide was 9.9 +/- 1.8 mg/kg/day. RESULTS: Eleven eyes (11 patients) were included. Indications for crossover from oral to topical carbonic anhydrase inhibitor (CAI) therapy were intolerance to acetazolamide (6 eyes) and surgical intervention in the fellow eye (5 eyes). The mean age at the time of crossover was 7.4 +/- 3.0 years. A comparison of intraocular pressure (IOP) before addition of a CAI was made in 8 eyes. The mean IOP off of a CAI was 27.8 +/- 4.9 mm Hg. The mean 10P was reduced to 18.5 +/- 4.3 mm Hg on acetazolamide (mean percent IOP reduction 35.7% +/- 15.6%, p < 0.01) and to 22.2 +/- 5.4 mm Hg on dorzolamide (mean percent IOP reduction 27.4% +/- 17.1%, p < 0.01). All 11 eyes showed an increase in IOP when switched from acetazolamide to dorzolamide, with a mean increase of 3.7 +/- 2.5 mm Hg (20.2% -/+ 13.7%, p < 0.01). Five eyes have remained controlled on dorzolamide and a topical beta-blocker. Five eyes required further intervention for the control of glaucoma. One eye was switched back to acetazolamide for better IOP control. CONCLUSION: Although not as effective as oral acetazolamide, topical dorzolamide causes a significant IOP reduction in this group of pediatric glaucoma patients and appears to be well tolerated. PMID- 10532367 TI - Acquired cataracts after argon laser photocoagulation for retinopathy of prematurity. AB - PURPOSE: Our purpose was to determine the incidence of cataract after argon laser photocoagulation of the retina in infants with retinopathy of prematurity (ROP). METHODS: We reviewed medical records of 189 consecutive infants undergoing argon laser photocoagulation for acute ROP between 1993 and 1996. Birth weight, estimated gestational age at birth, chronologic and postconceptional ages at the time of treatment, ROP outcome, and the postoperative occurrence of cataract were recorded. RESULTS: A total of 374 eyes in 189 infants were treated for threshold ROP. Mean birth weight was 916 gm (range 480 to 2500 gm), mean postconception age at birth was 26.4 weeks (range 23.5 to 34 weeks), and mean postconception age at surgery was 36.2 weeks (range 33 to 47 weeks). A favorable anatomic outcome occurred in 90% of eyes. However, severe macular traction, macular fold, or retinal detachment developed in 10% of eyes. A total of four eyes (1%) had cataracts. Laser-induced cataracts were diagnosed in only two eyes. A third eye had a posterior subcapsular cataract that may or may not have resulted from the laser treatment. In a fourth patient a unilateral punctate opacity judged to be visually insignificant was noted at surgery but it was not progressive. All patients who had cataracts had a persistent tunica vasculosa lentis; however, there was no statistically significant difference in the incidence of cataract formation in eyes with persistent tunica vasculosa lentis compared with eyes without it (p = 0.057). CONCLUSION: Argon laser photocoagulation remains an effective alternative to transscleral cryotherapy in the treatment of threshold ROP. The incidence of cataract formation is approximately 1% and may be more likely to occur when persistent hyaloidal vessels are present on the lens. PMID- 10532368 TI - Significance of isolated neovascular tufts ("popcorn") in retinopathy of prematurity. AB - BACKGROUND: The significance of isolated neovascular tufts ("popcorn") occurring in association with stage 2 retinopathy of prematurity (ROP) has not been studied. METHODS: We retrospectively reviewed the clinical courses and outcomes of all patients with zone II, stage 2 ROP with popcorn examined over the past 3 years at one institution. Eyes with zone I disease, plus disease, or stage 3 at the initial appearance of popcorn were excluded. The study group was compared with a control group of patients of similar birth weight and gestational age with zone II, stage 2 ROP without popcorn. RESULTS: Popcorn first appeared at a mean age of 36.4 (+/- 2.2) weeks after conception in 26 patients. Of these, 17 patients (65%) progressed to stage 3, 10 (38%) had plus disease, 6 (23%) reached threshold, and 9 (35%) required laser treatment. Of 19 control patients, 4 (21%) progressed to stage 3, 1 (5%) had plus disease, 1 (5%) reached threshold, and 1 (5%) required laser treatment. The popcorn group had a significantly higher incidence of progression to stage 3 (p < 0.005), plus disease (p < 0.025), and laser treatment (p < 0.025). All eyes of both groups had complete regression of disease. CONCLUSIONS: The presence of popcorn significantly increases the risk that an eye with zone II, stage 2 ROP will progress to stage 3, develop plus disease, and require laser treatment. Patients with popcorn and coexistent mild vascular dilation or tortuosity insufficient for plus disease are at particularly high risk for disease progression. PMID- 10532369 TI - The management of nasolacrimal duct obstruction in children between 18 months and 4 years old. AB - PURPOSE: Success with nasolacrimal duct probing has been shown to be inversely correlated with age. Consequently, several authors have suggested that the older child with a previously untreated nasolacrimal duct obstruction should undergo silicone intubation or a balloon catheterization as the primary surgical procedure because older children are more likely to have complicated obstructions that will not respond to simple probing. The purpose of this study was to investigate the hypothesis that older children with uncomplicated nasolacrimal duct obstruction can be successfully managed with simple probing. METHODS: A 14 year prospective study was conducted of consecutive patients older than age 18 months with nasolacrimal duct obstruction. All were treated (subject to certain exclusion criteria) with a simple nasolacrimal duct probing. Careful attention was paid to the type of obstruction encountered at surgery. Outcome evaluation included a standard ophthalmologic examination plus a dye disappearance test at 6 weeks after surgery. A follow-up examination or telephone interview was conducted 1 year after surgery. RESULTS: Of 378 children undergoing nasolacrimal duct probing, 23 met the inclusion criteria of being older than age 18 months (18 to 48 months). Seventy percent of the 23 children had a good outcome from the probing procedure. When analyzed by the type of obstruction, 12 of the 12 children (100%) with a simple membrane at the valve of Hasner had a good outcome. This contrasted with a success rate of 4 of 11 children (36%) who had complicated obstructions (p < 0.01). Complicated nasolacrimal duct obstructions were more prevalent in older children. CONCLUSION: A simple probing of the nasolacrimal duct has an excellent success rate in children up to 4 years old if an uncomplicated obstruction is found at the valve of Hasner. PMID- 10532370 TI - Trisomy 8p and Rieger malformation. PMID- 10532371 TI - From DNA sequence to application: possibilities and complications. AB - The development of sophisticated genetic tools during the past 15 years have facilitated a tremendous increase of fundamental and application-oriented knowledge of lactic acid bacteria (LAB) and their bacteriophages. This knowledge relates both to the assignments of open reading frames (ORF's) and the function of non-coding DNA sequences. Comparison of the complete nucleotide sequences of several LAB bacteriophages has revealed that their chromosomes have a fixed, modular structure, each module having a set of genes involved in a specific phase of the bacteriophage life cycle. LAB bacteriophage genes and DNA sequences have been used for the construction of temperature-inducible gene expression systems, gene-integration systems, and bacteriophage defence systems. The function of several LAB open reading frames and transcriptional units have been identified and characterized in detail. Many of these could find practical applications, such as induced lysis of LAB to enhance cheese ripening and re-routing of carbon fluxes for the production of a specific amino acid enantiomer. More knowledge has also become available concerning the function and structure of non-coding DNA positioned at or in the vicinity of promoters. In several cases the mRNA produced from this DNA contains a transcriptional terminator-antiterminator pair, in which the antiterminator can be stabilized either by uncharged tRNA or by interaction with a regulatory protein, thus preventing formation of the terminator so that mRNA elongation can proceed. Evidence has accumulated showing that also in LAB carbon catabolite repression in LAB is mediated by specific DNA elements in the vicinity of promoters governing the transcription of catabolic operons. Although some biological barriers have yet to be solved, the vast body of scientific information presently available allows the construction of tailor-made genetically modified LAB. Today, it appears that societal constraints rather than biological hurdles impede the use of genetically modified LAB. PMID- 10532372 TI - Low-redundancy sequencing of the entire Lactococcus lactis IL1403 genome. AB - Lactococcus lactis is an AT-rich gram positive bacterium phylogenetically close to the genus Streptococcus. Various strains of L. lactis are used in dairy industry as starters for cheese making. L. lactis is also one of the well characterized laboratory microorganisms, widely used for studies on physiology of lactic acid bacteria. We describe here a low redundancy sequence of the genome of the strain L. lactis IL1403. The strategy which we followed to determine the sequence consists of two main steps. First, a limited number of plasmids and lambda-phages that carry random segments of the genome were sequenced. Second, sequences of the inserts were used for production of novel sequencing templates by applying Multiplex Long Accurate PCR protocols. Using of these PCR products allowed to determine the sequence of the entire 2.35 Mb genome with a very low redundancy, close to 2. The error rate of the sequence is estimated to be below 1%. The correctness of the sequence assembly was confirmed by PCR amplification of the entire L. lactis IL1403 genome, using a set of 266 oligonucleotides. Anotation of the sequence was undertaken by using automatic gene prediction computer tools. This allowed to identify 1495 protein-encoding genes, to locate them on the genome map and to classify their functions on the basis of homology to known proteins. The function of about 700 genes expected to encode proteins that lack homologs in data bases cannot be reliably predicted in this way. The approach which we used eliminates high redundancy sequencing and mapping efforts, needed to obtain detailed and comprehensive genetic and physical maps of a bacterium. Availability of detailed genetic and physical maps of the L. lactis IL1403 genome provides many entries to study metabolism and physiology of bacteria from this group. The presence of 42 copies of five different IS elements in the IL1403 genome confirms the importance of these elements for genetic exchange in Lactococci. These include two previously unknown elements, present at seven and fifteen copies and designated IS1077 and IS983, respectively. Five potential or rudimentary prophages were identified in the genome by detecting clusters of phage-related genes. The metabolic and regulatory potential of L. lactis was evaluated by inspecting gene sets classified into different functional categories. L. lactis has the genetic potential to synthesise 20 standard amino acids, purine and pyrimidine nucleotides and at least four cofactors. Some of these metabolites, which are usually present in chemically defined media, can probably be omitted. About twenty compounds can be used by L. lactis as a sole carbon source. Some 83 regulators were revealed, indicating a regulatory potential close to that of Haemophilus influenzae, a bacterium with a similar genome size. Unexpectedly, L. lactis has a complete set of late competence genes, which may have concerted transcriptional regulation and unleadered polycistronic mRNAs. These findings open new possibilities for developing genetic tools, useful for studies of gene regulation in AT-rich gram positive bacteria and for engineering of new strains for the diary industry. PMID- 10532373 TI - Group II introns and expression of conjugative transfer functions in lactic acid bacteria. AB - The homologous lactococcal conjugative elements pRS01 and the sex factor of Lactococcus lactis strain 712 both contain a Group II intron within a gene believed to encode a conjugative relaxase enzyme. This enzyme is responsible for nicking of DNA at the origin of transfer (oriT) sequence of the sex factor DNA to initiate the strand transfer process. Group II introns have been studied in eukaryotes, and several of these elements in yeast mitochondrial genes have received considerable attention. These introns are relatively large in size and generally encode a protein within the intron sequence. In addition to splicing activity. Group II introns are mobile genetic elements. The intron-encoded proteins (IEPs) contain endonuclease and reverse transcriptase domains believed to play an enzymatic role in genetic mobility reactions, while a putative maturase domain is thought to promote splicing by stabilizing the folding of the intron RNA into an active ribozyme structure which carries out the splicing reaction. The lactococcal introns represent the first examples of Group II introns shown to be functional in vivo in prokaryotes. Because of the advantages of a bacterial system for genetic and molecular studies, the Ll.ltrB intron from pRS01 has attracted the attention of several laboratories interested in Group II intron biology. Recently, it has been shown that the system can be adapted to function in Escherichia coli (although at somewhat reduced efficiency). In addition, it has been recently proven that the best studied form of mobility, the homing of the intron into an intronless allele of the cognate exon gene, occurs via an RNA intermediate and does not require DNA homology or generalized host recombination functions. Current efforts are analysis of the role Ll.ltrB splicing in regulating expression of pRS01 conjugation functions. The lactococcal Group II introns represent the first demonstrated genetically mobile prokaryotic retroelements, and they also have considerable potential as genetic engineering tools for Lactic Acid Bacteria (LAB) and other organisms. PMID- 10532374 TI - Bacteriophage defence systems in lactic acid bacteria. AB - The study of the interactions between lactic acid bacteria and their bacteriophages has been a vibrant and rewarding research activity for a considerable number of years. In the more recent past, the application of molecular genetics for the analysis of phage-host relationships has contributed enormously to the unravelling of specific events which dictate insensitivity to bacteriophage infection and has revealed that while they are complex and intricate in nature, they are also extremely effective. In addition, the strategy has laid solid foundations for the construction of phage resistant strains for use in commercial applications and has provided a sound basis for continued investigations into existing, naturally-derived and novel, genetically-engineered defence systems. Of course, it has also become clear that phage particles are highly dynamic in their response to those defence systems which they do encounter and that they can readily adapt to them as a consequence of their genetic flexibility and plasticity. This paper reviews the exciting developments that have been described in the literature regarding the study of phage-host interactions in lactic acid bacteria and the innovative approaches that can be taken to exploit this basic information for curtailing phage infection. PMID- 10532375 TI - Acquired antibiotic resistance in lactic acid bacteria from food. AB - Acquired antibiotic resistance, i.e. resistance genes located on conjugative or mobilizable plasmids and transposons can be found in species living in habitats (e.g. human and animal intestines) which are regularly challenged with antibiotics. Most data are available for enterococci and enteric lactobacilli. Raw material from animals (milk and meat) which are inadvertantly contaminated with fecal matters during production will carry antibiotic resistant lactic acid bacteria into the final fermented products such as raw milk cheeses and raw sausages. The discovered conjugative genetic elements of LAB isolated from animals and food are very similar to elements studied previously in pathogenic streptococci and enterococci, e.g. theta-type replicating plasmids of the pAMbeta1, pIP501-family, and transposons of the Tn916-type. Observed resistance genes include known genes like tetM, ermAM, cat, sat and vanA. A composite 29,871 bp resistance plasmid detected in Lactococcus lactis subsp. lactis isolated from a raw milk soft cheese contains tetS previously described in Listeria monocytogenes, cat and str from Staphylococcus aureus. Three out of five IS elements on the plasmid are almost or completely identical to IS1216 present in the vanA resistance transposon Tn1546. These data support the view that in antibiotic challenged habitats lactic acid bacteria like other bacteria participate in the communication systems which transfer resistance traits over species and genus borders. The prevalence of such bacteria with acquired resistances like enterococci is high in animals (and humans) which are regularly treated with antibiotics. The transfer of antibiotic resistant bacteria from animals into fermented and other food can be avoided if the raw substrate milk or meat is pasteurized or heat treated. Antibiotic resistance traits as selectable markers in genetic modification of lactic acid bacteria for different purposes are presently being replaced, e.g. by metabolic traits to generate food-grade vectors. PMID- 10532376 TI - Multi-domain, cell-envelope proteinases of lactic acid bacteria. AB - The multi-domain, cell-envelope proteinases encoded by the genes prtB of Lactobacillus delbrueckii subsp. bulgaricus, prtH of Lactobacillus helveticus, prtP of Lactococcus lactis, scpA of Streptococcus pyogenes and csp of Streptococcus agalactiae have been compared using multiple sequence alignment, secondary structure prediction and database homology searching methods. This comparative analysis has led to the prediction of a number of different domains in these cell-envelope proteinases, and their homology, characteristics and putative function are described. These domains include, starting from the N terminus, a pre-pro-domain for secretion and activation, a serine protease domain (with a smaller inserted domain), two large middle domains A and B of unknown but possibly regulatory function, a helical spacer domain, a hydrophilic cell-wall spacer or attachment domain, and a cell-wall anchor domain. Not all domains are present in each cell-envelope proteinase, suggesting that these multi-domain proteins are the result of gene shuffling and domain swapping during evolution. PMID- 10532377 TI - The biosynthesis and functionality of the cell-wall of lactic acid bacteria. AB - The cell wall of lactic acid bacteria has the typical gram-positive structure made of a thick, multilayered peptidoglycan sacculus decorated with proteins, teichoic acids and polysaccharides, and surrounded in some species by an outer shell of proteins packed in a paracrystalline layer (S-layer). Specific biochemical or genetic data on the biosynthesis pathways of the cell wall constituents are scarce in lactic acid bacteria, but together with genomics information they indicate close similarities with those described in Escherichia coli and Bacillus subtilis, with one notable exception regarding the peptidoglycan precursor. In several species or strains of enterococci and lactobacilli, the terminal D-alanine residue of the muramyl pentapeptide is replaced by D-lactate or D-serine, which entails resistance to the glycopeptide antibiotic vancomycin. Diverse physiological functions may be assigned to the cell wall, which contribute to the technological and health-related attributes of lactic acid bacteria. For instance, phage receptor activity relates to the presence of specific substituents on teichoic acids and polysaccharides; resistance to stress (UV radiation, acidic pH) depends on genes involved in peptidoglycan and teichoic acid biosynthesis; autolysis is controlled by the degree of esterification of teichoic acids with D-alanine; mucosal immunostimulation may result from interactions between epithelial cells and peptidoglycan or teichoic acids. PMID- 10532378 TI - Bacteriocins: mechanism of membrane insertion and pore formation. AB - Lactic acid bacteria produce several types of pore forming peptides. Class I bacteriocins are lantibiotics that contain (methyl)lanthionine residues that may form intramolecular thioether rings. These peptides generally have a broad spectrum of activity and form unstable pores. Class II bacteriocins are small, heat stable peptides mostly with a narrow spectrum of activity. Most bacteriocins interact with anionic lipids that are abundantly present in the membranes of gram positive bacteria. 'Docking molecules' may enhance the conductivity and stability of lantibiotic pores, while 'receptors' in the target membrane may determine specificity of class II bacteriocins. Insertion into the membrane of many bacteriocins is proton motive force driven. Lantibiotics may form pores according to a 'wedge-like' model, while class II bacteriocins may enhance membrane permeability either by the formation of a 'barrel stave' pore or by a 'carpet' mechanism. PMID- 10532379 TI - Intestinal microflora and the interaction with immunocompetent cells. AB - The intestinal mucosal surface is colonised by the comensal microflora that attains very high numbers of bacterial cells in the distal intestine, more specifically in the colon. At the same time these extensive areas are the interface with the external environment, through which most pathogens initiate infectious processes in mammals. Intestinal mechanisms of defense need to discriminate accurately between comensal, symbiotic microflora, and exogenous pathogens. Today we do not fully understand the essence of the mechanism of discrimination but, probably, innate as well as adaptive immune responses participate in this process. We have explored, in in vitro models, the capacity of mucosal immunocompetent cells to discriminate amongst signals delivered by different types of bacteria. We have found at least two different patterns of innate response to gram-negative and gram-positive bacteria, and within this last group big differences are observed between species. We have only worked with non pathogenic bacteria in what may represent the modulation of the physiological host status. The understanding of these modulatory functions could render a unique possibility for the use of food-borne bacteria to prevent or correct intestinal problems associated with food allergy, inflammatory bowel disease, and autoimmunity. PMID- 10532380 TI - Bioactive peptides encrypted in milk proteins: proteolytic activation and thropho functional properties. AB - The bioactivities of peptides encrypted in major milk proteins are latent until released and activated by enzymatic proteolysis, e.g. during gastrointestinal digestion or food processing. The proteolytic system of lactic acid bacteria can contribute to the liberation of bioactive peptides. In vitro, the purified cell wall proteinase of Lactococcus lactis was shown to liberate oligopeptides from beta- and alpha-caseins which contain amino acid sequences present in casomorphins, casokinines, and immunopeptides. The further degradation of these peptides by endopeptidases and exopeptidases of lactic acid bacteria could lead to the liberation of bioactive peptides in fermented milk products. However, the sequences of practically all known biologically active peptides can also be cleaved by peptidases from lactic acid bacteria. Activated peptides are potential modulators of various regulatory processes in the body: Opioid peptides are opioid receptor ligands which can modulate absorption processes in the intestinal tract, angiotensin-I-converting enzyme (ACE)-inhibitory peptides are hemodynamic regulators and exert an antihypertensive effect, immunomodulating casein peptides stimulate the activities of cells of the immune system, antimicrobial peptides kill sensitive microorganisms, antithrombotic peptides inhibit aggregation of platelets and caseinophosphopeptides may function as carriers for different minerals, especially calcium. Bioactive peptides can interact with target sites at the luminal side of the intestinal tract. Furthermore, they can be absorbed and then reach peripheral organs. Food-derived bioactive peptides are claimed to be health enhancing components which can be used for functional food and pharmaceutical preparations. PMID- 10532381 TI - Peptidases and amino acid catabolism in lactic acid bacteria. AB - The conversion of peptides to free amino acids and their subsequent utilization is a central metabolic activity in prokaryotes. At least 16 peptidases from lactic acid bacteria (LAB) have been characterized biochemically and/or genetically. Among LAB, the peptidase systems of Lactobacillus helveticus and Lactococcus lactis have been examined in greatest detail. While there are homologous enzymes common to both systems, significant differences exist in the peptidase complement of these organisms. The characterization of single and multiple peptidase mutants indicate that these strains generally exhibit reduced specific growth rates in milk compared to the parental strains. LAB can also catabolize amino acids produced by peptide hydrolysis. While the catabolism of amino acids such as Arg, Thr, and His is well understood, few other amino acid catabolic pathways from lactic acid bacteria have been characterized in significant detail. Increasing research attention is being directed toward elucidating these pathways as well as characterizing their physiological and industrial significance. PMID- 10532382 TI - Sulfur metabolism in bacteria associated with cheese. AB - Metabolism of sulfur in bacteria associated with cheese has long been a topic of interest. Volatile sulfur compounds, specifically methanethiol, are correlated to desirable flavor in Cheddar cheese, but their definitive role remains elusive. Only recently have enzymes been found that produce this compound in bacteria associated with cheese making. Cystathionine beta- and gamma-lyase are found in lactic acid bacteria and are capable of producing methanethiol from methionine. Their primary function is in the metabolism of cysteine. Methionine gamma-lyase produces methanethiol from methionine at a higher efficiency than the cystathionine enzymes. This enzyme is found in brevibacteria, bacilli, and pseudomonads. Addition of brevibacteria containing this enzyme improves Cheddar cheese flavor. Despite recent progress in sulfur metabolism more information is needed before cheese flavor associated with sulfur can be predicted or controlled. PMID- 10532383 TI - Analysis of the intestinal microflora: a renaissance. AB - The ability of microbial ecologists to analyse the composition of complex bacterial communities has been greatly enhanced by the application of molecular methodologies. The use of these techniques should enable an accurate record of the identity and population dynamics of the inhabitants of the intestinal tract to be obtained, and should promote an improved comprehension of the relationship between the microflora and the human host. This, in turn, will lead to a new concept of the intestinal microflora of humans. PMID- 10532384 TI - Probiotics: from myth to reality. Demonstration of functionality in animal models of disease and in human clinical trials. AB - The enteric flora comprise approximately 95% of the total number of cells in the human body and are capable of eliciting immune responses while also protecting against microbial pathogens. However, the resident bacterial flora of the gastrointestinal tract (GIT) may also be implicated in the pathogenesis of several chronic conditions such as inflammatory bowel disease (IBD). The University College Cork-based Probiotic Research Group has successfully isolated and identified lactic acid bacteria (LAB) which exhibit beneficial probiotic traits. These characteristics include the demonstration of bile tolerance; acid resistance; adherence to host epithelial tissue; and in vitro antagonism of potentially-pathogenic micro-organisms or those which have been implicated in promoting inflammation. The primary objective of this report is to describe the strategy adopted for the selection of potentially effective probiotic bacteria. The study further describes the evaluation of two members of the resulting panel of micro-organisms (Lactobacillus salivarius subsp. salivarius UCC118 and Bifidobacterium longum infantis 35624) under in vitro conditions and throughout in vivo murine and human feeding trials. Specifically, an initial feeding study completed in Balb/c mice focused upon (i) effective delivery of the probiotic micro-organisms to the GIT and evaluation of the ability of the introduced strains to survive transit through, and possibly colonise, the murine GIT; (ii) accepting the complexity of the hostile GIT and faecal environments, development of a method of enumerating the introduced bacterial strains using conventional microbiological techniques; and (iii) assessment of the effects of administered bacterial strains on the numbers of specific recoverable indigenous bacteria in the murine GIT and faeces. Additional research, exploiting the availability of murine models of inflammatory bowel disease, demonstrated the beneficial effects of administering probiotic combinations of Lactobacillus salivarius UCC118 and Bifidobacterium longum infantis 35624 in prevention of illness-related weight loss. A further ethically-approved feeding trial, successfully conducted in 80 healthy volunteers, demonstrated that yoghurt can be used as a vehicle for delivery of Lactobacillus salivarius strain UCC118 to the human GIT with considerable efficacy in influencing gut flora and colonisation. PMID- 10532385 TI - Bringing a probiotic-containing functional food to the market: microbiological, product, regulatory and labeling issues. AB - Properly formulated probiotic-containing foods offer consumers a low risk, low cost dietary component that has the potential to promote health in a variety of ways. Several such products are available commercially, although markets in Japan and Europe are more developed than in the USA. Once healthful attributes of a probiotic product have been identified, there remain microbiological, product, regulatory and labeling issues to be addressed prior to marketing. Microbiological and product issues include safety, effective scale-up for manufacturing, definition of probiotic activity, probiotic stability in the product over the course of product manufacture, shelf-life and consumption, definition of effective dose and target population(s), and development of quality assurance approaches. Examples of probiotic-containing foods are given. Regulatory and labeling issues are complicated because they differ for each country, but are likewise critical because they provide the means for communication of the product benefits to the consumer. The regulatory climate worldwide appears to be one of caution about overstating the benefits of such products but at the same time not preventing corporate commitment to marketing. PMID- 10532386 TI - Lactic acid bacteria in the quality improvement and depreciation of wine. AB - The winemaking process includes two main steps: lactic acid bacteria are responsible for the malolactic fermentation which follows the alcoholic fermentation by yeasts. Both types of microorganisms are present on grapes and on cellar equipment. Yeasts are better adapted to growth in grape must than lactic acid bacteria, so the alcoholic fermentation starts quickly. In must, up to ten lactic acid bacteria species can be identified. They belong to the Lactobacillus, Pediococcus, Leuconostoc and Oenococcus genera. Throughout alcoholic fermentation, a natural selection occurs and finally the dominant species is O. oeni, due to interactions between yeasts and bacteria and between bacteria themselves. After bacterial growth, when the population is over 10(6) CFU/ml, malolactic transformation is the obvious change in wine composition. However, many other substrates can be metabolized. Some like remaining sugars and citric acid are always assimilated by lactic acid bacteria, thus providing them with energy and carbon. Other substrates such as some amino acids may be used following pathways restricted to strains carrying the adequate enzymes. Some strains can also produce exopolysaccharides. All these transformations greatly influence the sensory and hygienic quality of wine. Malic acid transformation is encouraged because it induces deacidification. Diacetyl produced from citric acid is also helpful to some extent. Sensory analyses show that many other reactions change the aromas and make malolactic fermentation beneficial, but they are as yet unknown. On the contrary, an excess of acetic acid, the synthesis of glucane, biogenic amines and precursors of ethylcarbamate are undesirable. Fortunately, lactic acid bacteria normally multiply in dry wines; moreover some of these activities are not widespread. Moreover, the most striking trait of wine lactic acid bacteria is their capacity to adapt to a hostile environment. The mechanisms for this are not yet completely elucidated. Molecular biology has provided some explanations for the behaviour and the metabolism of bacteria in wine. New tools are now available to detect the presence of desirable and undesirable strains. Even if much remains unknown, winemakers and oenologists can nowadays better control the process. By acting upon the diverse microflora and grape musts, they are more able to produce healthy and pleasant wines. PMID- 10532387 TI - Overview on applications for bacteriocin-producing lactic acid bacteria and their bacteriocins. PMID- 10532388 TI - Developing applications for lactococcal bacteriocins. AB - While much of the applied research carried out to date with bacteriocins has concerned nisin, lactococci produce other bacteriocins with economic potential. An example is the two component bacteriocin lacticin 3147, which is active over a wide pH range and has a broad spectrum of activity against gram-positive bacteria. Since the genetic determinants for lacticin 3147 are encoded on a large self-transmissible plasmid, the bacteriocin genes may be conveniently transferred to different lactococcal starters. The resulting food-grade strains can then be used to make a significant impact on the safety and quality of a variety of fermented foods, through the inhibition of undesirable microflora. The bacteriocin is heat stable so it can also be used as an ingredient in a powdered form such as a spray-dried fermentate. Given the observation that lacticin 3147 is effective at physiological pH, there is also considerable potential for biomedical applications. Field trials have demonstrated its efficacy in the prevention of mastitis infections in dairy cows. In contrast to lacticin 3147, the lactococcin bacteriocins A, B and M have a narrow spectrum of activity limited to lactococci. Strains which produce these inhibitors can be exploited in the acceleration of cheese ripening by assisting the premature lysis of starter cultures. PMID- 10532389 TI - Multidrug resistance in lactic acid bacteria: molecular mechanisms and clinical relevance. AB - The active extrusion of cytotoxic compounds from the cell by multidrug transporters is one of the major causes of failure of chemotherapeutic treatment of tumor cells and of infections by pathogenic microorganisms. The secondary multidrug transporter LmrP and the ATP-binding cassette (ABC) type multidrug transporter LmrA in Lactococcus lactis are representatives of the two major classes of multidrug transporters found in pro- and eukaryotic organisms. Therefore, knowledge of the molecular properties of LmrP and LmrA will have a wide significance for multidrug transporters in all living cells, and may enable the development of specific inhibitors and of new drugs which circumvent the action of multidrug transporters. Interestingly, LmrP and LmrA are transport proteins with very different protein structures, which use different mechanisms of energy coupling to transport drugs out of the cell. Surprisingly, both proteins have overlapping specificities for drugs, are inhibited by the same set of modulators, and transport drugs via a similar transport mechanism. The structure-function relationships that dictate drug recognition and transport by LmrP and LmrA will represent an intriguing new area of research. PMID- 10532390 TI - DNA-microarrays and food-biotechnology. PMID- 10532391 TI - Exopolysaccharides produced by Lactococcus lactis: from genetic engineering to improved rheological properties? AB - Over the last years, important advances have been made in the study of the production of exopolysaccharides (EPS) by several lactic acid bacteria, including Lactococcus lactis. From different EPS-producing lactococcal strains the specific eps gene clusters have been characterised. They contain eps genes, which are involved in EPS repeating unit synthesis, export, polymerisation, and chain length determination. The function of the glycosyltransferase genes has been established and the availability of these genes opened the way to EPS engineering. In addition to the eps genes, biosynthesis of EPS requires a number of housekeeping genes that are involved in the metabolic pathways leading to the EPS-building blocks, the nucleotide sugars. The identification and characterisation of several of these housekeeping genes (galE, galU, rfbABCD) allows the design of metabolic engineering strategies that should lead to increased EPS production levels by L. lactis. Finally, model development has been initiated in order to predict the physicochemical consequences of the addition of a EPS to a product. PMID- 10532392 TI - Anchoring of proteins to lactic acid bacteria. AB - The anchoring of proteins to the cell surface of lactic acid bacteria (LAB) using genetic techniques is an exciting and emerging research area that holds great promise for a wide variety of biotechnological applications. This paper reviews five different types of anchoring domains that have been explored for their efficiency in attaching hybrid proteins to the cell membrane or cell wall of LAB. The most exploited anchoring regions are those with the LPXTG box that bind the proteins in a covalent way to the cell wall. In recent years, two new modes of cell wall protein anchoring have been studied and these may provide new approaches in surface display. The important progress that is being made with cell surface display of chimaeric proteins in the areas of vaccine development and enzyme- or whole-cell immobilisation is highlighted. PMID- 10532393 TI - Applications of phage resistance in lactic acid bacteria. PMID- 10532394 TI - Immunomodulatory function of lactic acid bacteria. AB - Using mice, we found that oral administration of Bifidobacterium breve YIT4064 (B. breve) activated the humoral immune system, augmented anti-rotavirus IgA production or anti-influenza virus (IFV) IgG production and protected against rotavirus infection or influenza infection, respectively. Furthermore, when the B. breve was given to infants from an infant home, there was a significant reduction of the frequency of rotavirus shedding in stool samples during the administration of the bacteria. It was also found, again using mice, that oral administration of Lactobacillus casei strain Shirota (LcS) stimulated type 1 helper T (Th1) cells, activated the cellular immune system and inhibited incidence of tumors and IgE production. These results demonstrated that these two strains of lactic acid bacteria modulated the different immune systems each in its own way and prevented against various diseases. PMID- 10532395 TI - The role of lactic acid bacteria in colon cancer prevention: mechanistic considerations. AB - Colorectal cancer is one of the most important causes of cancer morbidity and mortality in Western countries. While a myriad of healthful effects have been attributed to the probiotic lactic acid bacteria, perhaps the most controversial remains that of anticancer activity. It should be pointed out already at this point that there is no direct experimental evidence for cancer suppression in humans as a result of consumption of lactic cultures in fermented or unfermented dairy products. However, there is a wealth of indirect evidence, based largely on laboratory studies, in the literature. The precise mechanisms by which lactic acid bacteria may inhibit colon cancer are presently unknown. However, such mechanisms might include: enhancing the host's immune response; binding and degrading potential carcinogens; quantitative and/or qualitative alterations in the intestinal microflora incriminated in producing putative carcinogen(s) and promoters (e.g. bile acid-degrading bacteria); producing antitumorigenic or antimutagenic compounds in the colon; alteration of the metabolic activities of intestinal microflora; alteration of physicochemical conditions in the colon; effects on physiology of the host. These potential mechanisms are discussed in the present paper. PMID- 10532396 TI - Lactic acid food fermentation in tropical climates. PMID- 10532397 TI - Non-dairy lactic fermentations: the cereal world. AB - Sourdough is the foremost cereal fermentation performed in a variety of technologies with almost any cereal. The lactobacilli studied most intensely include Lactobacillus sanfranciscensis, L. reuteri and L. pontis isolated from traditional and modern rye and wheat fermentations. Molecular biology tools are available for their rapid identification and monitoring throughout a process. The currently available insight on their metabolism can be used to explain their prevalence in this environment and their interactions. Key genes of the sugar degradation pathway were cloned and characterised from L. sanfranciscensis. In addition some strains were found to have special properties including the production of antagonistic compounds or the adhesion to human intestinal cells. PMID- 10532398 TI - Agonist-induced adherence of equine neutrophils to fibronectin- and serum-coated plastic is CD18 dependent. AB - Adherence to vascular endothelium and extracellular matrix proteins is a pre requisite for neutrophil accumulation at sites of inflammation. In this study, equine neutrophil adherence to fibronectin and autologous serum-coated plastic in response to PAF, hrIL-8, hrC5a and PMA has been measured. In addition, the mechanisms involved have been investigated using monoclonal antibodies (MoAbs) against the beta2 integrin CD18. PAF and hrC5a caused similar, concentration dependent, increases in adherence to fibronectin- and serum-coated plastic (maximum responses 19 +/- 4% and 19 +/- 3% for PAF and 15 +/- 4% and 16 +/- 2% for hrC5a on fibronectin- and serum-coated plastic, respectively). Adherence in response to PMA, although not reaching a maximum over the time course studied, was of a similar magnitude on the two surfaces (41 +/- 1% and 38 +/- 2% with 10( 7) M PMA on fibronectin- and serum-coated plastic, respectively). In contrast, the maximum adherence caused by hrIL-8 was significantly lower on fibronectin- than on serum-coated plastic (9 +/- 3% vs. 17 +/- 2%; 10(8) x M hrIL-8). Pre incubation with MoAbs against CD18 (H20A and 6.5E) caused concentration related inhibition of stimulus-induced adherence to both fibronectin- and serum-coated plastic. Equine neutrophil adherence in response to PAF, hrIL-8, hrC5a and PMA therefore appears to be mediated by a CD18 dependent mechanism. PMID- 10532399 TI - Kidney leucocytes of rainbow trout, Oncorhynchus mykiss, are activated by intraperitoneal injection of beta-endorphin. AB - The immunomodulatory effects of beta-endorphin were studied by measurements of the production of superoxide anion, phagocytosis and chemotaxis of kidney phagocytic cells in rainbow trout, Oncorhynchus mykiss. The production of superoxide anion in phagocytic cells increased significantly in rainbow trout injected with chum salmon beta-endorphin. The responses were dose-dependent. The phagocytosis and chemotaxis also significantly increased in kidney phagocytic cells of rainbow trout injected with alpha-endorphin. These results show that beta-endorphin in rainbow trout activates the function of phagocytic cells in vivo. PMID- 10532400 TI - Factor XIIIa positive dendritic cells are a major accessory cell population in the elicitation phase of DNCB-induced contact hypersensitivity. AB - The phenotypes and distribution of accessory cells in the ear skin of lambs during the elicitation phase of dinitrochlorobenzene (DNCB)-induced contact hypersensitivity (CHS) were examined using indirect immunoperoxidase histochemistry (ABC method), and a panel of antibodies. Thirty lambs, between 21 and 26 weeks of age, were divided into groups of 10. The shaved right ear of one group was treated with DNCB. Two weeks later this group was challenged with DNCB. One group was treated with the vehicle alone and the remaining group was left untreated. The lambs were slaughtered 48 h after challenge, and tissue specimens were collected from the ears of the three groups. Factor XIIIa+ (FXIIIa+) cells were prominent in the superficial dermis and showed predominantly a perivascular and subepidermal distribution. The other markers were less prominent, and whereas CD1+ cells and CD68+ cells showed a reaction pattern similar to the FXIIIa+ cells, CD14+ cells were found scattered predominantly in the deep dermis. There appeared to be an increase in FXIIIa+ cells, CD1+ cells, and CD68+ cells in the dermis of the DNCB-treated lambs 48 h after challenge. Only CD1+ cells were detected in epidermis of normal controls, and these cells appeared to be decreased in number in the two treated groups. Computer-assisted morphometric analysis was used to estimate the relative presence of the accessory cell subpopulations in the superficial and deep dermis and the entire dermis. A statistical analysis of the relative area of immunostaining showed a significantly increased presence of FXIIIa+ cells and CD68+ cells in the dermis of the DNCB-treated lambs 48 h after challenge. Interestingly, FXIIIa+ cells and CD68+ cells were also significantly increased in the vehicle treated group compared with untreated controls. We found no significant difference in the presence of CD1+ cells or CD14+ cells in the DNCB treated group compared with the controls. The study showed that FXIIIa+ DDC are the major accessory cell population in normal ear skin of lambs and the major responsive population during the elicitation phase of CHS. The lack of response in the CD1+ cell population suggests a less prominent role for the LC-related DC in the skin during the elicitation phase. PMID- 10532401 TI - Differential complement activation by bovine IgG2 allotypes. AB - Immunoglobulin allotypes and complement (C) are known to be related to susceptibility to infection. Because bovine IgG2 is important in resistance to pyogenic infections and because its two allotypes, IgG2a and IgG2b, differ in sequence in the CH1, hinge, CH2, and CH3 regions, we tested the ability of these allotypes to initiate the bovine C cascade. Bovine IgG2a and IgG2b were standardized according to specific anti guinea pig red blood cell (GPRBC) ELISA activity using anti IgG2 reagents shown essentially unbiased for allotype. Complement activating activity of the allotypes was quantitated in a GPRBC lysis assay. With this system, IgG2b consistently had more than twice the activity in bovine C mediated lysis as compared with IgG2a. The fact that both EDTA and EGTA/Mg almost completely inhibited C mediated lysis of GPRBCs indicated that lysis was due to the classical pathway. Since antibody usually activates C by the classical pathway, this supports the supposition that activation was by the IgG2 GPRBC complexes. Flexibility analyses showed that IgG2b had a more rigid hinge than IgG2a, perhaps partially explaining the greater efficiency of IgG2b in C activation. Other mechanisms may include differences in glycosylation and in the amino acid at position 332. The difference in ability to activate C may mean that animals of the IgG2a allotype could be more susceptible to infection with extracellular pyogenic pathogens which are killed by C or by phagocytes after opsonization with IgG2 and C. PMID- 10532402 TI - Involvement of reactive oxygen species in TNF-alpha mediated activation of the transcription factor NF-kappaB in canine dermal fibroblasts. AB - The cytokine tumor necrosis factor-alpha (TNF-alpha) plays a major role in inflammatory and immune-pathological reactions of the skin. With respect to a possible therapeutic modulation of TNF-alpha mediated activation of Nuclear Factor-kappa B (NF-kappaB) in canine cutaneous inflammation, we investigated the role of NF-kappaB and the involvement of reactive oxygen species (ROS) in the TNF alpha signalling pathway in dermal fibroblasts of the dog. TNF-alpha treatment resulted in the activation of NF-kappaB as assessed by electrophoretic mobility shift assay (EMSA). Additionally, NF-kappaB translocation was induced with butylhydroperoxide and antimycin A, but not with hydrogen peroxide. TNF-alpha stimulated NF-kappaB activation was partially inhibited by preincubation with the antioxidants alpha-lipoic acid and butylated hydroxyanisol (BHA). No superoxide generation following TNF-alpha stimulation could be detected in the supernatant of canine fibroblasts with the superoxide dismutase-inhibitable cytochrome c reduction test. In contrast, production of TNF-alpha dependent intracellular hydrogen peroxide, the dismutation product of the superoxide radical, was demonstrated spectroscopically by formation of electron dense cerium hydroperoxide precipitates. With electron energy loss spectroscopy (EELS) significant cerium deposits were detected in the mitochondria, the endoplasmatic reticulum, the cytosol and to a lesser extent on the plasma membrane of canine fibroblasts indicating multiple hydrogen peroxide production sites. Peroxides, therefore, possibly play an important part in the redox-sensitive pathway of TNF alpha dependent NF-kappaB activation in canine skin. An adjunctive therapy with appropriate antioxidants modulating NF-kappaB overactivation in cutaneous inflammation in the dog is promising. PMID- 10532403 TI - Binding of bovine IgG2a and IgG2b allotypes to protein A, protein G, and Haemophilus somnus IgBPs. AB - Immunoglobulin binding proteins (IgBPs) are thought to be virulence factors which enable pathogens to evade the host's immune response. Since bovine IgG2 is important in protection against pyogenic infections, the binding characteristics of Staphylococcus aureus protein A (PrA), streptococcal protein G (PrG), or Haemophilus somnus high molecular weight IgBPs to the two bovine IgG2 allotypes were examined. For PrA or PrG binding of IgG2, guinea pig red blood cells coated with specific IgG2a or IgG2b antibodies were used in a competitive binding inhibition assay with unlabeled and horseradish peroxidase-labeled PrA or PrG. To determine which sizes of H. somnus. IgBPs bind to the two IgG2 allotypes, immunoblots with H. somnus culture supernatant were probed with anti-DNP IgG2a and IgG2b. This detects only Fc binding because anti-DNP does not cross-react with H. somnus antigens. Both IgG2 allotypes bound equally well to PrA and PrG. However, IgG2b but not IgG2a bound to H. somnus high molecular weight IgBPs. The lack of differential binding of bovine IgG2 allotypes to PrA and PrG means that these IgBPs can be considered to be unbiased reagents in assays for detection of bovine IgG2 or for immunoaffinity purification. The differential binding of H. somnus IgBPs to the IgG2 allotypes indicates that animals having one allotype may be more resistant to H. somnus infection than animals having the other allotype. PMID- 10532404 TI - Parallel visual processes in health and disease. Symposium for the Francqui International Interuniversity Chair in Biomedical Sciences 1997. PMID- 10532405 TI - Dissecting the dark-adapted electroretinogram. AB - Although gross recordings of the ganzfeld flash-evoked electroretinogram (ERG) can potentially provide information about the activity of many, if not all, retinal cell types, it is necessary to dissect the ERG into its components to realize this potential fully. Here we describe various procedures that have been used in intact mammalian eyes to identify and characterize the contributions to the dark-adapted ERG of different cells in the retinal rod pathway. These include (1) examination of the very early part of the response to a flash (believed to reflect directly the photocurrent of rods), (2) application of high-energy probe flashes to provide information about the underlying rod photoreceptor response even when this component is obscured by the responses of other cells, (3) pharmacological suppression of responses of amacrine and ganglion cells to identify the contribution of these cells and to reveal the weaker responses of bipolar cells, (4) use of pharmacological agents that block transmission of signals from rods to more proximal neurons to separate responses of rods from those of later neurons, (5) examination of the ERG changes produced by ganglion cell degeneration or pharmacological block of nerve-spike generation to identify the contribution of spiking neurons, (6) modeling measured amplitude-energy functions and timecourse of flash responses and (7) using steady backgrounds to obtain differential reductions in sensitivity of different cell types. While some of these procedures can be applied to humans, the results described here have all been obtained in studies of the ERG of anaesthetized cats, or macaque monkeys whose retinas are very similar to those of humans. PMID- 10532406 TI - Electrophysiological findings in dominant optic atrophy (DOA) linking to the OPA1 locus on chromosome 3q 28-qter. AB - Pattern and flash visual evoked cortical potentials (PVEP, FVEP), and pattern electroretinograms, (PERG) were recorded in 13 affected individuals from 8 families with DOA. These were selected as representative from 87 affected members of 21 pedigrees with DOA who were examined, and who underwent genetic linkage analysis. Linkage to the OPA1 locus on chromosome 3q 28-qter was demonstrated in all families. VA ranged from 6/9 to HM: visual fields showed a variable centro caecal defect; SLO (when performed) showed diffuse nerve fibre loss; MRI (when performed) showed small intra-orbital optic nerves. In 9/13 patients the PVEP was absent in one or both eyes. Most recordable PVEPs were of abnormal latency, but the delays were not marked (peak times 116-135 msec); amplitudes were low or subnormal. PERG fell within the normal range in 9 eyes of 7 patients. 14 eyes showed an abnormal N95:P50 ratio in keeping with ganglion cell dysfunction. Some severely affected eyes showed P50 component involvement, but in no eye was the PERG extinguished. Significant interocular asymmetries in at least one electrophysiological measure were present in 6/13 patients. Colour contrast thresholds were significantly elevated for all three colour confusion axes, with tritan being most affected. PMID- 10532407 TI - Spatial and temporal response properties of the major retino-geniculate pathways of Old and New World monkeys. AB - Old World monkeys, apes and humans all enjoy trichromatic colour vision, and the absorption spectra of the photopigments are very similar in all species and all individuals. Colour vision in New World monkeys however, is very heterogeneous. In many species, the majority of individuals is dichromatic. Recently, anatomical and electrophysiological studies revealed that the retinal organisation in Old World monkeys and New World monkeys is very similar, although the cells belonging to the parvocellular pathway do not show any colour opponency and their spectral sensitivity is similar to that of the magnocellular cells. Apparently, the magnocellular and parvocellular pathways in the retina and the LGN have not developed as an adaptation to luminance and chromatic processing. It is more likely that the two pathways originally evolved to cover different ranges in the spatio-temporal domain. In the present paper, several spatial and temporal properties of parvo- and magnocellular cells (which are identical for dichromatic and trichromatic animals) are compared. PMID- 10532408 TI - Assessment of stereopsis in rhesus monkeys using visual evoked potentials. AB - Rhesus monkeys can have deficiencies in stereo vision, making it necessary to screen monkey subjects intended for single cell studies of stereo-based depth processing. We measured VEPs in two monkeys using a dynamic random-dot display in which a stereo-defined checkerboard reversed in depth. Monkeys fixated upon a small dot during stimulus presentation. One monkey showed clear evoked potentials in response to changes in disparity that were similar to those obtained in human subjects, using an identical stimulus paradigm. Controls with presentations of the monocular stimulus sequences (in which no depth reversal can be perceived) yielded no or much weaker VEPs. In the other animal, however, there was no difference in evoked potential between the two conditions. These electrophysiological findings closely match the performance of these same two subjects in a disparity discrimination task in which they were previously trained. We conclude that VEPs using this type of stimulus display can be used to screen monkeys for single cell or behavioral studies of stereopsis. PMID- 10532409 TI - Maturation of binocular luminance interaction in normal young and adult rhesus monkeys. AB - PURPOSE: Our aim was to answer three questions 1) Do adult rhesus monkeys have binocular luminance interactions (BLIs) similar to those found in adult humans? 2) Is BLI in very young rhesus monkeys functionally mature? 3) If not, how does it change with age? METHODS: We recorded visually evoked potentials (VEPs) in response to sinusoidally modulated uniform fields. The fields were presented dichoptically by varying the relative temporal phase between the two eyes. Monkeys varied in age from 5.6 weeks to 5.25 years. RESULTS: VEPs were Fourier analyzed and the relative second harmonic amplitudes were taken as the response measure. The second harmonic amplitudes in adult monkeys had an asymmetrical 'V shaped' function as interocular phase difference (IPD) varied from 0 degrees to 180 degrees, as had been observed previously in adult humans [1]. The youngest monkeys exhibited a symmetrical pattern which became more asymmetrical at older ages and was adult like by about 19 weeks. Asymmetry magnitude and log age correlated 0.97 (p < 0.05) in the monkeys younger than 19 weeks. CONCLUSIONS: The adult rhesus data are consistent with a model derived from humans which involves two types binocular luminance processing. One combines monocular responses nonlinearly (MNL) and a second combines monocular responses linearly followed by a binocular nonlinearity (MLBNL). These two processes have been associated with the parvocellular (P-) and magnocellular (M-) streams. Within this framework, the data from the youngest monkeys indicate that BLI in the P-stream is relatively less mature at birth than that in the M-stream and develops reaching functional maturity on these measures by around 19 weeks. PMID- 10532410 TI - The separation of parallel visual systems by disease processes. AB - AIM: to investigate psychophysics and electrophysiology in disease states with altered retinal function. METHODS: determination of colour and luminance contrast sensitivity functions using computer controlled VDUs: recording of flash and pattern ERGs to short and long flashes of coloured lights under differing conditions of light adaptation: recording of pattern ERG. RESULTS: In melanoma associated retinopathy (MAR) low spatial frequency temporal flicker loss occurs for achromatic Gaussians, but colour Gaussians are seen normally. Low spatial frequency luminance contrast sensitivity and motion losses are severe while red green gratings are seen normally. In Cuban Tropical Amblyopia, achromatic luminance contrast sensitivity may be normal, in the presence of considerable losses of colour vision, to spatial frequencies as high as 32 c/degree. There are supernormal cone ERGs. CONCLUSIONS: these 2 conditions represent highly selective loss of 'M' and 'P' pathways respectively. Almost the entire 'bell shaped curve' normally represents M activity. In CTA, there may be a selective loss of the receptive field surrounds of P ganglion cells. PMID- 10532411 TI - Parallel pathways, noise masking and glaucoma detection: behavioral and electrophysiological measures. AB - PURPOSE: We tested the hypothesis that because of their reduced neural efficiency, glaucoma patients should have increasingly impaired thresholds as external noise is added to a stimulus. METHOD: We compared the performance of 20 normals (mean age = 39 years) with that of 15 patients with early glaucoma or at very high risk for glaucoma (mean age 45 years). All patients had normal visual acuity. Contrast thresholds were measured on two sets of tasks: (1) behavioral and (2) sweep visually evoked potentials (VEPs). Two stimuli were used (a) 7.5 Hz reversing gratings of 0.69 cpd, and (b) 5.5 cpd gratings. Noise was binary and contrast varied from 0 to 80%. Psychophysical thresholds were determined using a staircase which employed a spatial four alternative forced choice procedure (4AFC) and converged on 50% correct. Sweep VEP thresholds were determined by extrapolation to zero volts as a function of log contrast. RESULTS: Differences between normal subjects and patients with early glaucoma were not significant without noise. Both the absolute size of the difference and its significance increased as noise level increased. For the behavioral thresholds these trends were clearer with the 5.5 cpd grating, while for the sweep VEPs they were more clear for the 0.69 cpd grating. CONCLUSION: The performance deficit of glaucoma patients which may be minimal under normal testing conditions is magnified when external noise is added to the stimulus. VEPs and psychophysical thresholds show interesting differences in their sensitivity to this effect. Implications for the early detection of glaucoma are discussed. PMID- 10532412 TI - Reduced duration of a visual motion aftereffect in congenital nystagmus. AB - Congenital nystagmus (CN) is a primarily horizontal, involuntary, conjugate eye movement which can be observed soon after birth or during the first half-year of life. Individuals with CN rarely complain of oscillopsia. Using a motion aftereffect (MAE), we investigated if individuals with CN have abnormalities in motion perception and if any such abnormality could be due to nystagmus or to compensatory mechanisms to avoid oscillopsia. In task A, patients (n=10) and control subjects (n=10) indicated the direction, duration and relative velocity of MAEs. The subjects binocularly viewed a high contrast, grey scale grating (0.23 cyc/deg; visual angle: 18.3 deg) moving upward or downward at 1, 3, and 6 deg/sec for 60 sec. Vertical optokinetic nystagmus (OKN) was monitored. In task B, patients (n=8) and control subjects (n=8) viewed similar spatial frequency gratings (visual angle: 40.7 degs; 0.5, 0.2, 0.08 cyc/deg) which moved at 4, 10, and 16 deg/sec for 60 sec. In task C, five control subjects, with induced vestibular nystagmus, viewed a grating (0.2 cyc/deg; visual acuity: 28.5 deg), moving upward for 40 sec. In all three tasks, after adaptation with the moving grating, subjects viewed the then static grating and reported the duration and direction of the MAE. One CN patient and eight control subjects reported MABs at all three test velocities in task A. When patients exhibited OKN, the gain was close to one, as in the control group. In task B, seven of the eight patients and all of the control subjects had MABs at the faster adaptation velocities. CN patients had less MAEs at an adaptation velocity of 4 deg/sec and when MAEs were observed, the duration of the illusory motion was reduced by approximately 48%. Control subjects, with induced vestibular nystagmus, reported MAEs at 4 deg/sec (task C). These findings indicate that nystagmus cannot be the only factor accounting for the suppression of motion perception and suggest that compensatory mechanisms used to avoid oscillopsia contribute to the differences found between the groups. PMID- 10532413 TI - Visually evoked potentials evoked by moving unidimensional noise stimuli: effects of contrast, spatial frequency, active electrode location, reference electrode location, and stimulus type. AB - We determined the relative importance of electrode derivation, stimulus type, spatial frequency and contrast in determining the relative size of the late negative and early positive responses of motion elicited VEPs. Seven subjects aged 22-48 years with normal vision were tested binocularly. Motion onset and motion reversal were employed as modes of stimulus presentation. For both, pseudo random one-dimensional noise patterns whose peak power was at 5.2, 2.6, 1.3, 0.325 and 0.1625 cycles per degree (cpd) were stimuli. Contrasts were 70% and 5%. Active electrodes were placed at Oz, 5 cm to the left of Oz, 5 cm to the right of Oz and a frontal midline position (Fpz) and referenced to linked mastoids. Transient motion reversal elicited a prominent positive response present in all subjects and at low contrasts. Motion onset VEPs have a complex waveform which may be either predominantly positive or negative. The most important variables in determining whether a prominent positivity or negativity is present in the motion onset VEP are the contrast and the spatial frequencies. Data such as these are first efforts in developing recommendations for the motion VEP. PMID- 10532414 TI - Electrophysiological correlates of human texture segregation, an overview. AB - 'Texture segregation' results from parallel processing in the visual cortex. It occurs when the stimulus contains spatial gradients within a visual dimension. We here present an introductory overview of the field, concentrating on electrophysiological correlates in the human EEG ('tsVEPs') of the neuronal processes underlying texture segregation. We describe the isolation of the tsVEP from the background EEG, give examples of the correlation between saliency and tsVEP amplitude and compare texture segregation between visual dimensions. PMID- 10532415 TI - Parallel visual and memory processes. AB - We studied parallel processes: visual processes with the prosaccade, the no saccade and the antisaccade task on the one hand and memory processes with the random tap task on the other hand. The random tap task is believed to be a pure interference task for the central executive component of working memory. The number of saccadic errors was found not to be influenced by taxing the central executive, while the latency times were significantly increased both in the prosaccade and in the antisaccade task. The effect seen in the antisaccade task was expected since it is a non-automatic activity under central executive control. Because the prosaccade task is an automatic activity, an effect of central executive load was not expected. As an explanation for our findings, we postulate that the prosaccade task is brought under willed control of the central executive. PMID- 10532416 TI - Parallel visual processes in symmetry perception: normality and pathology. AB - Mirror symmetry is one of those regularities for which the visual system seems to have developed a special sensitivity. It is detected robustly and efficiently in a single glance, suggesting that the basic processes do not perform a serial, pointwise comparison of structural elements but rather operate in parallel. Psychophysical evidence relating to the processing mechanisms will be reviewed. Although the focus will be on symmetry perception in normal vision, interesting findings on symmetry perception in observers with deficient vision (e.g., retinitis pigmentosa, visual hemineglect) will also be touched upon briefly. PMID- 10532417 TI - Pathobiology of the pineal gland with emphasis on parenchymal tumors. PMID- 10532418 TI - Expression of apoptosis and its related protein in astrocytic tumors. AB - The relationship between malignant potential and apoptosis in astrocytic tumors has not been clearly defined, and further classification of astrocytic tumors is necessary. To elucidate the relationship between the histopathological grade of astrocytic tumors and apoptosis, we studied 25 cases of astrocytic tumors, comprising 10 cases of glioblastoma (GB), 7 cases of anaplastic astrocytoma (AA), and 8 cases of astrocytoma (AC). We detected apoptosis using the TdT-mediated dUTP-biotin nick-end labeling (TUNEL) method. We studied immunohistochemical expression of bcl-2 protein and p53 protein, which are apoptosis-related factors, and cell proliferative activity using Ki-67 antibody. No significant change was noted between apoptotic index and the histological grade of the tumors. In GB, apoptotic cell-rich foci were present at the pseudopalisading necrosis. No correlation between histopathological grades and expression of either p53 or bcl 2 was observed. In GB, however, poor distribution of bcl-2 was found in the areas of pseudopalisade formation. bcl-2 is one of the regulatory factors in the cell cycle and inhibits apoptosis. Expression of apoptosis had no correlation with histopathological grade. However, in GB, the distribution of apoptotic cells showed a correlation with bcl-2-poor foci. It was thought that apoptosis was one of the regulatory factors in the formation of pseudopalisading necrosis in GB. PMID- 10532419 TI - Anomalous p27kip1 expression in a subset of malignant gliomas. AB - p27Kip (p27) expression was immunohistochemically investigated in 28 astrocytic tumors, and compared with the cell proliferation index (MIB-1 staining index). Normal rat brains and surgical specimens from human nonneoplastic brain lesions were used as controls. In the rat brains, the astrocytes were exclusively p27 positive. The reactive astrocytes in various disease processes sometimes lacked p27 expression. The distribution of p27-positive cells was uniform in low-grade astrocytomas and heterogeneous in high-grade tumors. Double staining of p27 and MIB-1 showed a reciprocal pattern in most cases. The frequency of p27 expression was inversely correlated with MIB-1 staining index and tumor grade. However, several malignant gliomas showed high p27 expression in spite of high MIB-1 staining indices. In such cases, MIB-1-positive cells were occasionally p27 positive. In this paper we discuss the etiology of the anomalous p27 expression in a subset of malignant gliomas. PMID- 10532421 TI - Intracranial malignant lymphomas: clinicopathological study of 26 autopsy cases. AB - We examined 26 autopsy-proven cases of intracranial malignant lymphoma (IML) in immunocompetent patients to determine the extent of neoplastic involvement of the central nervous system (CNS) and to evaluate the effects of radiation on the tumor and brain tissue. All tumors were identified as diffuse non-Hodgkin's lymphomas of B-cell origin. In six patients who had not received radiotherapy, the clinical course of the disease was short and extensive infiltration of the tumor was seen. The remaining 20 patients were treated with radiotherapy and had a longer survival time. Leptomeningeal involvement was common, but extensive subarachnoid proliferation of the tumor was seen in only two cases. The posterior, but not anterior, lobe of the pituitary was involved in 5 of 22 cases, and choroid plexus involvement was seen in 4 of 21. Direct invasion of the tumor into the spinal cord, which tended to occur in patients with posterior fossa masses, was observed in 5 of 21 cases. Following irradiation, coagulation necrosis was frequently found in the invading zone as well as in the tumor mass, and degeneration of the white matter was also seen. We suggest that IML can extensively infiltrate into the CNS, including the posterior lobe of the pituitary and spinal cord, and that radiation injury to the brain appears to occur relatively easily in this disease. PMID- 10532420 TI - Expression of the multidrug-resistance P-glycoprotein (Pgp, MDR-1) by endothelial cells of the neovasculature in central nervous system tumors. AB - To elucidate the expression of the MDR1 gene products P-glycoprotein (Pgp) in endothelial cells of newly formed blood microvessels in brain tumors, 30 brain tumors were examined by immunohistochemistry using an anti-Pgp monoclonal antibody, JSB-1. Positive reactions for JSB-1 were detected in endothelial cells in newly formed microvessels in all 16 cases of glioma but not in the 4 meningiomas. Although endothelial cells in newly formed microvessels of all 10 metastatic carcinomas showed positive reactions, negative reactions were seen in those of the primary carcinomas. Compared with reactions of the endothelial cells of normal cerebral capillaries, weak reactions were found in the endothelial cells forming glomeruloid proliferation in newly formed microvessels in the eight glioblastomas and at the border of the surrounding cerebral tissue of the metastatic carcinomas. Since the endothelial cells showing glomeruloid proliferation also had a high proliferative cell nuclear antigen labeling index, the present findings demonstrate a negative relationship between positive reactions for Pgp and the proliferative activities of endothelial cells in cerebral capillaries. PMID- 10532422 TI - Prognostic value of tumor-associated antigens immunoreactivity and apoptosis in medulloblastomas. An analysis of 73 cases. AB - Medulloblastomas (MB) are the most common central nervous system malignancies in children. Numerous publications describe efforts to identify the predictive value of various patterns of MB pathology and immunohistochemistry, but received data appear to be controversial. Seventy-three patients with cerebellar MB were studied retrospectively. Tumor specimens were immunohistochemically examined with antibodies to various tumor-associated antigens. Also, apoptosis detection by the in situ end-labeling method was performed. Survival analysis was made using univariate and multivariate models. Tenascin immunoreactivity and apoptotic index (AI) > or = 1.5% were found to be closely associated with poor prognosis according to an univariate analysis (P = 0.008 and 0.003, respectively). The multivariate Cox proportional hazard model exhibited independent prognostic value for the apoptotic rate only (P = 0.023). Tumors with tenascin expression and AI > or = 1.5% significantly prevailed among MB with metastatic dissemination, whereas expression of c-erbB2 oncoprotein and epidermal growth factor receptor was found to be more typical for cases with local tumor recurrence. We came to the conclusion that tenascin immunoreactivity and AI were useful for individual MB prognosis. PMID- 10532423 TI - Glioblastoma multiforme with epithelial appearance: a case report. AB - A case of glioblastoma multiforme with epithelial appearance, which was difficult to diagnose at first operation, is described. Microscopy revealed small, darkly staining anaplastic cells which were densely packed. In some areas, these cells were arranged in a tubular, gland-like pattern mimicking a poorly differentiated epithelial neoplasm. Immunostaining of glial fibrirally acidic protein (GFAP) was negative in the densely compact anaplastic areas and within the epithelial patterns, except for a small number of cells in one area. Further pathological study at the second and third operations indicated that the tumor consisted of neoplastic astrocytes and had characteristic features of glioblastoma multiforme, including necrosis, pseudopalisading, and endothelial proliferation. Many of the tumor cells were GFAP-positive. This rare case of glioblastoma multiforme was compared with cases reported in the literature. PMID- 10532424 TI - Trigeminal ganglioneuroma. AB - We present the case of an 8-year-old girl with a ganglioneuroma in the left cerebellopontine angle region. The tumor originated from the sensory root of the trigeminal nerve. Histopathologically, it was composed of neoplastic ganglion cells and Schwann cells, leading us to the diagnosis of ganglioneuroma. Intracranial ganglioneuroma is very rare. To our knowledge, this is the first report of a trigeminal ganglioneuroma. The nature and origin of this tumor are discussed and the literature reviewed. PMID- 10532425 TI - Solitary eosinophilic granuloma of the temporal lobe: case report and review of the literature. AB - A solitary eosinophilic granuloma of the central nervous system is an unusual manifestation of histiocytosis X. A unique case of a solitary eosinophilic granuloma of the right temporal lobe without osseous involvement is described. A 20-year-old man presented with a grand mal seizure. Magnetic resonance imaging demonstrated an intraaxial enhancing mass in the right temporal lobe with marked vasogenic edema. A right temporal craniotomy was performed for resection of the lesion and the diagnosis of an eosinophilic granuloma was confirmed by histopathology. Follow-up MR imaging obtained 5 years following resection demonstrated no recurrence. Solitary eosinophilic granuloma should be considered in the differential diagnosis of enhancing mass lesions affecting the central nervous system. Although the natural history of solitary eosinophilic granulomas remains poorly defined, surgical treatment still remains the mainstay of therapy for these unifocal cerebral lesions. PMID- 10532426 TI - Sulfate regulation: native kidney vs dialysis. PMID- 10532427 TI - Hepatitis E virus in end-stage renal disease. PMID- 10532428 TI - Evaluation of the PD adequest program in Japanese patients on peritoneal dialysis. AB - To perform adequate peritoneal dialysis, it is necessary to individualize the dialysis regimen. PD Adequest is a software program based on the three-pore model that can be used to predict solute clearance and ultrafiltration volume during peritoneal dialysis. We evaluated the ability of this program to predict the solute clearance and ultrafiltration volume in Japanese patients on peritoneal dialysis. The weekly creatinine clearance and weekly urea Kt/V were determined in 45 patients. The PD Adequest was used to simulate their current dialysis regimens and the predicted values of these parameters were calculated. Strong positive correlations were obtained between the actual and predicted weekly creatinine clearance (r = 0.993) and weekly urea Kt/V (r = 0.991). In conclusion, this program is potentially useful for designing peritoneal dialysis regimens in Japanese patients, even though they have a smaller body mass than Canadians and Americans for whom it was designed. PMID- 10532429 TI - In vitro and in vivo evaluation of a new polysulfone membrane for hemodialysis. Reference methodology and clinical results. (Part 1: in vitro study). AB - Different high flux membranes have been recently developed. The present study is aimed at describing the technical features and the clinical performances of a new high flux polysulfone membrane (T-sulfone, Toray, Japan). The study has been carried out on two different dialyzers (surface area = 1.3 and 1.8 m2). The filters have been tested in vitro under definite experimental conditions. The hydraulic flow resistance, the pressure drop in the blood compartment and the hydraulic permeability have been determined in a wide range of in vitro experimental conditions. The in vitro sieving coefficients for various solutes have also been determined utilizing human blood. Hydraulic permeability was found in the range of 28.4 ml/h/mm Hg/m2 and sieving coefficients were between 0.96 and 1.0 for all low molecular weight solutes. The sieving coefficient for inulin was 0.95. The pressure drop in the filter at 300 ml/min of blood flow was 95 mm Hg for the 1.3 m2 and 57 mm Hg for the 1.8 m2. The filters are then designed to operate in the presence of high blood flows without excessive resistance in the blood compartment. The blood compartment analyzed by means of a special radiological sequence obtained with a helical scanner after dye injection confirmed the homogeneous distribution of the blood flow in several cross sections of the bundle. Adequate distribution of dialysate was confirmed with a similar method applied to the dialysate compartment. The new imaging techniques utilized were greatly helpful to determine adequacy of filter design and flows distribution. PMID- 10532430 TI - In vitro and in vivo evaluation of a new polysulfone membrane for hemodialysis. Reference methodology and clinical results. (Part. 2: in vivo study). AB - Different high flux membranes have been recently developed. The present study is aimed at describing the technical features and the clinical performances of a new high flux polysulfone membrane (T-sulfone, Toray, Japan). The study has been carried out on two different dialyzers (surface area = 1.3 and 1.8 m2). The filters have been tested in vivo during hemodialysis and hemodiafiltration. The in vivo study was carried out on 12 ESRD patients on regular hemodialysis treatment. The protocol was reviewed and approved by the local ethical committee. The in vivo clearances (K) at 300 ml/min of blood flow are reported in the following Table: [Table in text]. Beta-2-m reduction ratio exceeded 50% in all sessions. Beta-2-m mass balance executed by collection of spent dialysate and elution from the used filters evidenced that removal is obtained mostly by filtration while absorption is negligible. Excellent tolerance and hemocompatibility was observed in all the studied sessions. PMID- 10532431 TI - Intra-aortic balloon: evaluation of heparin-coating under various experimental conditions. AB - In a calf model, heparin coated intra-aortic balloon (IAB) was compared with standard balloon. In group 1, 9 of each IAB type were set to the automatic mode for 15 min, 45 min and 6 hours respectively, while in group 2, 3 of each IAB type were left deflated during 20 minutes to simulate balloon dysfunction. At the end of the procedures, 3 samples of each IAB were analyzed with scanning electron microscopy (SEM) for surface deposits. Macroscopically, the 12/12 heparin coated IAB of both groups and the 9/9 standard IAB of group 1 were free of deposits, whereas the 3/3 standard IAB of group 2 exhibited clot deposits. SEM revealed deposit-free surfaces in the 36/36 heparin coated samples of both groups, while 14/27 standard samples of group 1 (p<0.01 when compared with heparin coated samples) and 8/9 standard samples of group 2 (p = 0.02, same comparison) disclosed blood cells and fibrin deposits. Morphometrically, the proportion of standard sample surfaces covered with deposits, estimated according to a score system (0% = 0; 0.1-25% = 1; 25.1-50% = 2; 50.1-75% = 3; 75. 1-100% = 4), was 0.69+/-0.82 in group 1 (p<0.01 when compared with heparin coated samples) and 1.22+/-0.83 in group 2 (p<0.01, same comparison). Thus heparin coated IAB presents no deposits either after 6 hours of intravascular ballooning or after 20 minutes of stagnation. It seems to be a promising strategy for patients with absolute or relative contraindications to systemic heparinization. PMID- 10532432 TI - Trillium coating of cardiopulmonary bypass circuits improves biocompatibility. AB - Coating of cardiopulmonary bypass circuits may be a solution to prevent adverse effects induced by contact of blood elements with foreign surfaces. Using an animal model, we investigated the Trillium TM coating of cardiopulmonary bypass circuits (a new process involving polyethylene oxide, sulphonate groups and heparin) at low systemic heparinization, focusing on haemolysis and clot formation. Cardiopulmonary bypass was initiated through jugulo-carotid access with ACT maintained around 180 sec. Treated circuits (Trillium group) were evaluated in 3 calves (mean weight of 66.0+/-8.7 kg), vs. untreated circuits in 3 control calves (mean weight of 60.7+/-7.5 kg). Blood samples were drawn at regular intervals for biochemical, hematological and blood gas analyses. After 6 consecutive hours, the animals were weaned from CPB and were awakened. The circuits were analyzed for clot deposits. After 7 days the animals were sacrificed and an autopsy was carried out. Red cell and white cell counts did not change over the 6 hours. Platelet counts dropped to 75.9+/-7.3% of the baseline value in the Trillium group after 6 hours whereas counts dropped to 57.2+/-26.0 in the control group (p<0.05). Plasma free Hb remained constant in the Trillium group but increased significantly to 280+/-65% of baseline value in the control group (p<0.05). The amount of clots were significantly higher in the control group, in the connectors, the reservoir, the heat exchanger, and the oxygenator. No renal emboli were seen in the Trillium group whereas the mean number of emboli was 3.0+/-2.4 in the control group. We conclude that Trillium coating significantly improves the biocompatibility of artificial surfaces exposed to blood. PMID- 10532433 TI - Effects of predeposit and intentional perioperative haemodilution on blood saving program in major orthopaedic surgery. AB - In this study we evaluated the effects of predeposit and intentional perioperative haemodilution on a blood saving program in major orthopaedic surgery. We demonstrated that autologous blood phlebotomy and maintenance of optimal levels of perioperative haemodilution by delaying blood transfusion, even autologous, are efficient techniques in reducing homologous, red blood cell (HRBC) transfusion. Patients who received autologous red blood cell (ARBC) or HRBC more than one day after surgery, while having Hb values <8 g/dl, are less at risk of needing the first or additional HRBCs. In conclusion, predeposit and intentional haemodilution obtained by delaying blood transfusions, even autotransfusional, is a correct way of conducting a blood saving program (BSP) in major orthopaedic surgery. These techniques are clinically effective in avoiding or reducing HRBC transfusion. PMID- 10532434 TI - Changes in HCV viremia following LDL apheresis in a HCV positive patient with familial hypercholesterolemia. AB - It has been suggested that hepatitis C virus (HCV) can be associated with beta lipoprotein in human serum. According to this, the LDL receptor could promote endocytosis of such a virus. In the present study, we evaluated the changes in HCV viremia in a HCV positive patient with familial hypercholesterolemia, undergoing both selective (DALI System, Fresenius) and non-selective (plasma exchange) LDL apheresis. HCV-RNA levels did not decrease following selective LDL apheresis, on the contrary showed a random, odd variation pattern (from -35% to +72%). Conversely, plasma exchange steadily induced a drop in HCV viremia ( 35/43%), to a lower extent than that of a totally intravascular plasmaprotein, i.e., alpha 2-macroglobulin (-53/54%). These data indicate that beta-lipoprotein may not function as a plasma carrier of HCV, at least in the present case. Moreover, a continuous, quantitatively unforeseeable circulation of HCV virions from the intravascular plasma compartment to other extravascular and intracellular sites, seems to occur during an apheresis session. PMID- 10532435 TI - Ozonation of blood during extracorporeal circulation. I. Rationale, methodology and preliminary studies. AB - We investigated whether exposure of blood ex-vivo to oxygen-ozone (O2-O3) through a gas exchanger is feasible and practical. We first evaluated the classical dialysis-type technique but we soon realized that semipermeable membranes are unsuitable because they are hydrophilic and vulnerable to O3. We therefore adopted a system with hydrophobic O3-resistant hollow fibers enclosed in a polycarbonate housing with a membrane area of about 0.5 m2. First we tested the system with normal saline, determining the production of hydrogen peroxide (H2O2) at O3 concentrations from 5 to 40 microg/ml. We then evaluated critical parameters by circulating swine blood in vitro; this revealed that heparin is not an ideal anticoagulant for this system. Finally, we performed several experiments in sheep and defined optimal anticoagulant dose (sodium citrate, ACD), priming solution, volume of blood flow per min, volume and concentration of O2-O3 mixture flowing countercurrent with respect to blood and the time necessary for perfusion in vivo. The biochemical parameters showed that an O3 concentration as low as 10 microg/ml is effective; this means that gas exchange and O3 reactivity are rapid and capable of inducing biological effects. The sheep showed no adverse effects even after 50 min of extracorporeal circulation at higher O3 concentrations (20 to 40 microg/ml) but the exchanger became less effective (low pO2 values) due to progressive clogging with cells. PMID- 10532436 TI - Amniotic membrane transplantation for symptomatic bullous keratopathy. AB - OBJECTIVE: To determine whether amniotic membrane transplantation can be used to treat symptomatic bullous keratopathy displaying poor visual potential. METHODS: Amniotic membrane transplantation was performed at 5 centers on 50 consecutive eyes (50 patients) with symptomatic bullous keratopathy and poor visual potential. The underlying causes of bullous keratopathy included aphakia (9 eyes), pseudophakia (19 eyes), failed grafts (9 eyes), and others (13 eyes). RESULTS: During the follow-up period of 33.8 weeks (3-96 weeks) after amniotic membrane transplantation, 43 (90%) of 48 eyes with intolerable pain preoperatively became pain free postoperatively. Among the 5 eyes with residual pain, 3 received repeated amniotic membrane transplantation, 1 required a conjunctival flap for pain relief, and 1 had reduced pain. Epithelial defects in 45 (90%) of 50 eyes created and covered by amniotic membrane healed rapidly within 3 weeks. Only 4 eyes (8%) showed recurrent surface breakdown. Epithelial edema or bullae recurred in a smaller area in 5 eyes (10%) and pseudopterygium developed in 1 eye. CONCLUSION: Amniotic membrane transplantation can be considered as an alternative to conjunctival flaps in alleviating pain, promoting epithelial healing, and preserving cosmetic appearance in patients with symptomatic bullous keratopathy and poor visual potential. PMID- 10532437 TI - Accuracy of scanning laser polarimetry in the diagnosis of glaucoma. AB - OBJECTIVE: To determine the diagnostic accuracy of scanning laser polarimetry. SUBJECTS AND METHODS: A total of 95 healthy subjects and 102 patients with glaucoma met all inclusion criteria. Data collected on each participant included an automated visual field examination, stereoview optic nerve head photographs, intraocular pressure measurement, and a screening and full scanning laser polarimetry study. Each participant was classified as "normal," "glaucoma," or "uncertain" by each of 3 ophthalmologists based on all available clinical information, with the exception of the scanning laser polarimetry results. Before data analysis, 4 diagnostic algorithms for the full-test mode and 2 for the screening mode were chosen to be evaluated for their sensitivity and specificity in detecting glaucoma. RESULTS: Of the 4 algorithms tested for the full-test mode, "the number" (abnormal test score, >35) had sensitivities of 57%, 71%, and 81% for early, moderate, and severe glaucoma, respectively. Specificity was 89%. For the screening test, sensitivities were much lower, particularly for those with severe glaucoma damage. CONCLUSIONS AND CLINICAL RELEVANCE: Scanning laser polarimetry can help to differentiate subjects with normal findings from patients with glaucomatous damage. Even the best algorithm tested, however, failed to detect a substantial number of subjects with severe damage. Further study is needed before scanning laser polarimetry can be recommended as a screening method for glaucoma. PMID- 10532438 TI - Latanoprost and respiratory function in asthmatic patients: randomized, double masked, placebo-controlled crossover evaluation. AB - OBJECTIVE: To evaluate the effects of single and multiple administrations of the ocular hypotensive drug latanoprost on respiratory function, asthma symptoms, and use of asthma medication in patients with bronchial asthma. METHODS: Twenty-four stable patients with asthma (forced expiratory volume in 1 second: 70% to 90% of predicted and a minimum of 10% reversibility after inhalation of albuterol sulfate) with no previous exposure to inhaled corticosteroids participated in this randomized, double-masked crossover trial. Patients received latanoprost, 0.005%, or placebo, 1 drop per day, in each eye during two 6-day treatment periods separated by a 2-week washout period. Acute latanoprost or placebo provocation, methacholine chloride airway reactivity, and 12-stimulator reversibility tests were performed. MAIN OUTCOME MEASURES: Morning and evening peak expiratory flow, spirometric performance throughout treatment periods and during different provocation tests, asthma symptoms, and use of asthma medications were evaluated. RESULTS: There were no statistically significant differences between treatments in morning and evening peak expiratory flow, scored daytime and nocturnal asthma symptoms, or daily consumption of asthma medication. During placebo provocation, there was a small increase in forced expiratory volume in 1 second that was not seen during latanoprost provocation. This small difference (-0.09 L) was statistically significant but without clinical importance. CONCLUSIONS: Resting and provoked airway function and asthma symptoms were unaffected by latanoprost treatment in patients with asthma with no previous exposure to corticosteroids. Latanoprost can be used in patients with glaucoma and concomitant bronchial asthma. PMID- 10532439 TI - Comparative study of trabecular aspiration vs trabeculectomy in glaucoma triple procedure to treat pseudoexfoliation glaucoma. AB - OBJECTIVES: To establish the relative safety and effectiveness of trabecular aspiration in combination with phacoemulsification and intraocular lens (IOL) implantation (asp+IOL) for decreasing intraocular pressure (IOP), and to compare the outcome of this method of treatment with that of phacoemulsification and IOL implantation alone (IOL-alone) or standard filtering glaucoma triple procedure (triple procedure). DESIGN: Prospective, controlled study randomized with respect to assignment to trabecular aspiration, with a case-control design between the asp+IOL and triple procedure groups. PARTICIPANTS: Seventy-four eyes of 74 patients with uncontrolled pseudoexfoliation glaucoma without a history of previous intraocular or extraocular surgery and in need of cataract surgery. Forty-eight patients were randomized to those receiving adjunctive trabecular aspiration (asp+IOL group of 26 eyes) and those given no trabecular aspiration (IOL-alone group of 22 eyes). The triple procedure group consisted of 26 cases, closely matched to the asp+IOL cases in terms of age, sex, cup-disc ratio, glaucoma medication requirements, and systemic diseases. INTERVENTIONS: Temporal clear corneal phacoemulsification and foldable IOL implantation was performed in all eyes. In the asp+IOL group, trabecular aspiration was performed with a suction force of 100 to 200 mm Hg under light tissue-instrument contact using a modified intraocular aspiration probe. A modified Cairns-type trabeculectomy without adjunctive antimetabolites was performed superiorly in the triple procedure eyes after IOL implantation. MAIN OUTCOME MEASURES: Surgical outcome was assessed in terms of IOP change, need of adjunctive glaucoma medication, visual acuity outcome, and complications. Surgical failure was defined as an outcome requiring additional surgical intervention or more than 1 medication to achieve IOP control to the target pressure. RESULTS: Two years after surgery, success rates were 36%, 64%, and 78% in the IOL-alone, asp+IOL, and triple procedure groups, respectively (P<.001). Hyphema (46%) and ocular hypotony (11%) were observed in the triple procedure group only, whereas blood reflux (61%) and descemetolysis (19%) were seen exclusively in the asp+IOL group. CONCLUSIONS: In pseudoexfoliative eyes, asp+IOL is significantly more effective than cataract surgery alone in reducing postoperative IOP and the necessity for glaucoma medication. Although trabecular aspiration in the triple procedure did not achieve pressure control in all patients, especially in the low target pressure range, the risk profile appears to be more favorable in the trabecular aspiration treated eyes than in the filtering glaucoma triple procedure group. Trabecular aspiration in the glaucoma triple procedure could serve as a possible first-line treatment for pseudoexfoliative eyes with coexisting cataract and glaucoma. PMID- 10532440 TI - The relationship between glaucoma and pseudoexfoliation: the Blue Mountains Eye Study. AB - OBJECTIVE: To quantify the relationship between pseudoexfoliation (PXF) and open angle glaucoma, ocular hypertension, and intraocular pressure (IOP) in a defined older population. METHODS: A cross-sectional study of 3654 people aged 49 to 97 years identified subjects with PXF during slitlamp examination. The IOP was measured by applanation tonometry. Glaucoma was diagnosed from characteristic visual field loss combined with optic disc cupping and rim thinning, without reference to IOP. Ocular hypertension was diagnosed if IOP was greater than 21 mm Hg in either eye, without field and disc changes. General estimating equation models were used to assess associations between eyes with PXF and glaucoma or ocular hypertension. RESULTS: Pseudoexfoliation was diagnosed in 2.3% of subjects, and both prevalence and bilaterality increased with age. Glaucomatous damage was present in 14.2% of eyes with PXF compared with 1.7% of eyes without PXF (age- and sex-adjusted odds ratio (OR), 5.0; 95% confidence interval (CI), 2.6-9.6). This was almost unchanged (OR, 4.8) after adjustment for glaucoma risk factors and was also relatively unaffected by IOP adjustment (OR, 3.7; 95% CI, 1.8-7.6). For subjects with PXF, the relationship with glaucoma persisted, but was weaker (OR, 2.3; 95% CI, 1.0-5.0) in the multivariate model. However, the population attributable risk from PXF was only 2.7%. Ocular hypertension was also more frequent in eyes with PXF (9.3%) than in eyes without PXF (3.1%) but was of borderline significance in the multivariate model (OR, 2.3; 95% CI, 0.9-5.7). CONCLUSIONS: This study confirmed the strong relationship between glaucoma and PXF. Subjects with PXF had an increased risk of glaucoma, while eyes with PXF had a higher risk, which was independent of other glaucoma risk factors, including IOP. PMID- 10532441 TI - Photodynamic therapy of subfoveal choroidal neovascularization in age-related macular degeneration with verteporfin: one-year results of 2 randomized clinical trials--TAP report. Treatment of age-related macular degeneration with photodynamic therapy (TAP) Study Group. AB - OBJECTIVE: To determine if photodynamic therapy with verteporfin (Visudyne; CIBA Vision Corp, Duluth, Ga) can safely reduce the risk of vision loss in patients with subfoveal choroidal neovascularization (CNV) caused by age-related macular degeneration (AMD). DESIGN: Two multicenter, double-masked, placebo-controlled, randomized clinical trials. SETTING: Twenty-two ophthalmology practices in Europe and North America. PARTICIPANTS: Patients with subfoveal CNV lesions caused by AMD measuring 5400 microm or less in greatest linear dimension with evidence of classic CNV and best-corrected visual acuity of approximately 20/40 to 20/200. METHODS: Six hundred nine patients were randomly assigned (2: 1) to verteporfin (6 mg per square meter of body surface area) or placebo (5% dextrose in water) administered via intravenous infusion of 30 mL over 10 minutes. Fifteen minutes after the start of the infusion, a laser light at 689 nm delivered 50J/cm2 at an intensity of 600 mW/cm2 over 83 seconds using a spot size with a diameter 1000 microm larger than the greatest linear dimension of the CNV lesion. At follow-up examinations every 3 months, retreatment with the same regimen was applied if angiography showed fuorescein leakage. The primary outcome was the proportion of eyes with fewer than 15 letters lost (approximately <3 lines of loss), adhering to an intent-to-treat analysis. RESULTS: In each group, 94% of patients completed the month 12 examination. Visual acuity, contrast sensitivity, and fluorescein angiographic outcomes were better in the verteporfin-treated eyes than in the placebo-treated eyes at every follow-up examination through the month 12 examination. At the month-12 examination, 246 (61%) of 402 eyes assigned to verteporfin compared with 96 (46%) of 207 eyes assigned to placebo had lost fewer than 15 letters of visual acuity from baseline (P<.001). In subgroup analyses, the visual acuity benefit (< 15 letters lost) of verteporfin therapy was clearly demonstrated (67% vs 39%; P<.001) when the area of classic CNV occupied 50% or more of the area of the entire lesion (termed predominantly classic CNV lesions), especially when there was no occult CNV. No statistically significant differences in visual acuity were noted when the area of classic CNV was more than 0% but less than 50% of the area of the entire lesion. Few ocular or other systemic adverse events were associated with verteporfin treatment, compared with placebo, including transient visual disturbances (18% vs 12%), injection-site adverse events (13% vs 3%), transient photosensitivity reactions (3% vs 0%), and infusion related low back pain (2% vs 0%). CONCLUSIONS: Since verteporfin therapy of subfoveal CNV from AMD can safely reduce the risk of vision loss, we recommend verteporfin therapy for treatment of patients with predominantly classic CNV from AMD. PMID- 10532442 TI - Diffuse bilateral subacute neuroretinitis: first patient with documented nematodes in both eyes. AB - OBJECTIVE: To describe the first patient with documented evidence of diffuse unilateral subacute neuroretinitis (DUSN) in both eyes. METHODS: A 10-year-old healthy Brazilian girl was first seen with signs of late-stage DUSN in both eyes. A careful search for a nematode was performed in each eye. RESULTS: A motile 550- to 660-microm nematode was found in the inferotemporal retina of the left eye. A similar-sized motile nematode was found in the superotemporal retina of the right eye. Both nematodes were treated with argon green laser applications with bilateral improvement of visual function. CONCLUSION: Although most patients with DUSN do not develop the disease in the fellow eye, this case demonstrates that DUSN can occasionally affect both eyes. PMID- 10532443 TI - A fluorescein and indocyanine green angiographic study of choriocapillaris in age related macular disease. AB - OBJECTIVE: To examine the phenomenon of a prolonged choroidal filling phase (PCFP) as seen on fluorescein and indocyanine green (ICG) angiography in patients with early age-related macular disease (AMD). METHODS: One hundred eyes of consecutive patients with early AMD were studied. Patchy and slow choroidal filling in early fluorescein and distinct areas of reduced choroidal fluorescence in ICG angiography were interpreted as PCFP. In addition, associated drusen characteristics and the AMD status of the fellow eye were recorded. RESULTS: A PCFP was observed in 26% of eyes using fluorescein and 32% of eyes using ICG angiography, with good concordance between findings using both techniques (K = 0.9). A PCFP was associated with confluent drusen (P = .01), the presence of focal retinal pigment epithelial-atrophic patches in the study eye (P=.005), and geographic atrophy in the fellow eye (P=.03). Other drusen characteristics and the distribution of visual acuity (P = .90) were not different between eyes with and without PCFP. CONCLUSIONS: A PCFP on fluorescein and ICG angiography is a common feature in early AMD. This sign has been interpreted as indicating reduced choroidal perfusion caused by change in diffusional characteristics of Bruch membrane. A PCFP is a clinical marker for diffuse deposits in Bruch membrane and a risk factor for the development of geographic atrophy. PMID- 10532444 TI - Is vascular regulation in the central retinal artery altered in persons with vasospasm? AB - OBJECTIVE: To assess the relation between ocular perfusion pressure and blood flow velocity in the central retinal artery in patients with acral vasospasm. SUBJECTS AND METHODS: Eighteen otherwise healthy subjects with acral vascular dysregulation, as demonstrated by nail-fold capillaroscopy, and 18 age- and sex matched healthy volunteers without vasospasm were recruited. After subjects had rested for 20 minutes in a supine position, intraocular pressure and blood flow velocity in the central retinal artery were determined by applanation tonometry and color Doppler imaging, respectively. The peak systolic velocity, end diastolic velocity, and resistivity index were assessed. Correlations between the mean ocular perfusion pressure (2/3 x [2/3 x diastolic blood pressure + 1/3 x systolic blood pressure] - intraocular pressure) and blood flow velocities in the central retinal artery were determined by the Pearson linear correlation factor. The Student t test was used to evaluate differences between controls and subjects with vasospasm. RESULTS: The mean +/- SD ocular perfusion pressure was 42.0 +/- 7.4 mm Hg in the group with vasospasm and 47.1 +/- 6.5 mm Hg in the control group (P= .04). In the subjects with vasospasm, the peak systolic and end-diastolic velocities and the resistivity index of the central retinal artery correlated significantly with the mean ocular perfusion pressure (R = 0.49, P = .04 P = .01; and R = -0.67, P = .002, respectively). Such correlations were not found in the control group. CONCLUSIONS: An altered blood flow regulation is suggested in the ocular circulation of patients with acral vasospasm. PMID- 10532445 TI - Effect of latanoprost on regional blood flow and capillary permeability in the monkey eye. AB - OBJECTIVE: To evaluate the effects of latanoprost on regional blood flow and capillary permeability in the monkey eye. METHODS: Anesthetized cynomolgus monkeys were unilaterally treated with a single dose containing 6 pg of latanoprost; or 10 microg of PhXA34 (13,14-dihydro-15R, S-17-phenyl-18,19,20 trinor-prostaglandin F2alpha [PGF2alpha]-isopropyl ester), which contains about 50% latanoprost. Regional blood flow in the eye was measured with radioactively labeled microspheres; capillary permeability was measured by determining the extravascular plasma-equivalent albumin space using 125I-albumin, 131I-albumin, and 51Cr-labeled erythrocytes. RESULTS: Latanoprost or PhXA34 had no or only a slight effect on the regional blood flow when measured 1, 2 1/2, 3, 4 1/2, and 6 hours after dose administration, with the exception of the anterior sclera, in which a moderate increase in blood flow was detected. No effect on capillary permeability to albumin was detected when studied 30 minutes to 2 1/2 hours and 5 to 6 hours after dose administration. CONCLUSION: Latanoprost, a selective prostaglandin F receptor agonist, exerted no or only slight vascular effects for up to 6 hours after dose administration in the monkey eye, with the exception of the anterior sclera, in which a moderate increase in blood flow was detected. CLINICAL RELEVANCE: Naturally occurring prostaglandins may cause marked microcirculatory changes in the eye that could be of clinical concern. Latanoprost, a selective prostaglandin F receptor agonist, seems to be devoid of such effects. PMID- 10532446 TI - Cataractous changes in rat lens following cigarette smoke exposure is prevented by parenteral deferoxamine therapy. AB - OBJECTIVES: To test whether iron accumulation in the lens following cigarette smoke exposure is the principal mechanism in smoke-related cataractogenesis and to assess the possible protective effect of deferoxamine mesylate treatment against lenticular degeneration with in vivo exposure to cigarette smoke. METHODS: Thirty-two male Wistar rats were randomly divided into 4 equal groups. Groups 3 and 4 rats were exposed to cigarette smoke for 1 hour each day for 90 consecutive days, and groups 1 and 2 rats were treated in a similar manner but exposed only to room air. In addition, deferoxamine was given subcutaneously to groups 2 and 4 rats. Both eyes of all the animals were then enucleated and 1 eye prepared for histopathological examination. The fellow eye was used to measure iron, calcium, zinc, and copper levels. RESULTS: Significantly higher iron and calcium and lower zinc levels were observed in the lenses of group 3 rats compared with those in the other groups. Similar comparisons performed between groups 1 and 2, 1 and 4, and 2 and 4 did not show any significant difference. Copper concentrations did not differ between groups. Distinct histopathological changes in the anterior lens epithelium, such as hyperplasia, hypertrophy, and epithelial multilayering, and the presence of swollen epithelial cells overlying the posterior lens capsule, observed in group 3 rats, were not present in the other groups. CONCLUSIONS: Cataractogenesis following cigarette smoke exposure in rats was associated with the accumulation of iron, and concurrent deferoxamine therapy prevented such cataract formation. CLINICAL RELEVANCE: Our results may apply to human cataract formation associated with cigarette smoking, so such pathogenesis may be prevented by concurrent parenteral deferoxamine treatment. Clinical studies are needed, however, to determine the value of this suggestion. PMID- 10532447 TI - Clinical features of codon 172 RDS macular dystrophy: similar phenotype in 12 families. AB - OBJECTIVE: To report the phenotype associated with the codon 172 RDS (gene for retinal degeneration slow) mutation in 11 separate families with an arginine-to tryptophan substitution with common ancestry, and 1 family with an arginine-to glutamine transition. PATIENTS: Screening for RDS gene mutations was performed in 400 subjects with autosomal dominant retinal degeneration. Twelve families were identified with a mutation in codon 172. Haplotype analysis was performed. Full ophthalmic evaluation was performed, including electrophysiologic and psychophysical investigation and imaging of autofluorescence using confocal laser scanning ophthalmoscopy. RESULTS: Haplotype analysis demonstrated that the 11 families were ancestrally related. All 12 families showed a common phenotype of macular dysfunction, with the deficit increasing with age. Abnormally high autofluorescence predated loss of visual acuity or visual field changes. Pattern electroretinographic (PERG) findings were affected early in disease. There was high intrafamilial and interfamilial consistency of phenotype. CONCLUSION: These families demonstrate a striking conformity of symptoms and signs. CLINICAL RELEVANCE: In the codon 172 RDS mutation, unlike disease resulting from other RDS mutations, prediction of approximate age of onset and progression of visual deficit is possible. This should assist diagnosis and counseling. PMID- 10532448 TI - Age-related macular degeneration and antioxidant status in the POLA study. POLA Study Group. Pathologies Oculaires Liees a l'Age. AB - OBJECTIVE: To give the levels of antioxidant nutrients in relation to age-related macular degeneration (AMD). METHODS: Pathologies Oculaires Liees a l'Age is a population-based study on cataract and AMD and their risk factors, carried out on 2584 inhabitants of Sete, France. Age-related macular degeneration was defined by findings from fundus photographs according to an international classification. Biological measurements were taken from fasting blood samples. RESULTS: After multivariate adjustment, plasma alpha-to-copherol levels showed a weak negative association with late AMD (P = .07). Lipid-standardized plasma alpha-tocopherol levels showed a significant negative association with late AMD (P= .003): the risk of late AMD was reduced by 82% in the highest quintile compared with the lowest. Similarly, lipid-standardized plasma alpha-tocopherol levels were inversely associated with early signs of AMD (odds ratio, 0.72 [95% confidence interval, 0.53-0.98]; P=.04). No associations were found with plasma retinol and ascorbic acid levels or with red blood cell glutathione values. COMMENT: These results suggest that vitamin E may provide protection against AMD. Only randomized interventional studies could prove the protective effect of vitamin E on AMD. PMID- 10532449 TI - Low uptake of eye services in rural India: a challenge for programs of blindness prevention. AB - OBJECTIVES: To investigate service uptake in a rural Indian population served by outreach eye camps and to identify barriers to uptake. PARTICIPANTS AND METHODS: A routine eye camp was conducted within 5 km of each of 48 randomly selected villages of typically Hindu, backward-caste communities. Subsequently, participatory rural appraisal-community mapping, focus groups, matrix ranking, and semistructured interviews-was undertaken to explore community views of eye problems. An eye examination was conducted on persons with eye problems who did not attend the eye camp. Predictors of attendance were identified by multilevel regression analysis. RESULTS: Of 749 adults with an eye problem, 51 (6.8%) attended the eye camp. Independent predictors of attendance were being male (odds ratio = 2.3; 95% confidence interval, 1.2-4.5) and living within 3 km of the camp (odds ratio = 4.5; 95% confidence interval, 1.7-12.5). Of the 552 persons who did not attend the eye camps and had an eye examination, 242 (43.8%) had low vision (visual acuity <6/18 to > or =3/60 in presenting better eye) and 38 (6.9%) were blind in both eyes. Cataract surgery was recommended for 197 (35.8%) of the persons who did not attend the eye camps. Of 109 persons with a previous cataract operation, 42 (38.5%) had low vision and 11 (10.1%) were blind. Fear (principally of eye damage), cost (direct and indirect), family responsibilities, ageism, fatalism, and an attitude of being able to cope (with low or no vision) were the principal barriers to attending the eye camps. CONCLUSIONS: A high proportion of people who could have benefited from eye treatment were not using available services. Poor visual outcomes were observed in surgically treated persons. PMID- 10532450 TI - Photodynamic therapy with verteporfin is effective for selected patients with neovascular age-related macular degeneration. PMID- 10532451 TI - Evaluating the retinal nerve fiber layer in glaucoma with scanning laser polarimetry. PMID- 10532452 TI - Ophthalmology in the United Kingdom. PMID- 10532453 TI - Canadian ophthalmology. PMID- 10532454 TI - Corneal epithelial toxic effects and inflammatory response to perfluorocarbon liquid. AB - We report an unusual case of corneal epithelial toxic effects associated with perfluorocarbon liquids (PFCLs). The clinical and histopathologic findings are described. An elderly man underwent vitreoretinal surgery for a complicated retinal detachment. Perfluorodecalin was used to repair the retina. It was left in situ for 8 weeks and removed via the pars plana. One month after removal of heavy liquids the patient developed a nonhealing corneal epithelial defect associated with limbitis. Perfluorodecalin was found under the superior conjunctiva. A conjunctival biopsy revealed the presence of vacuoles in the conjunctival stroma surrounded by an inflammatory response that consisted of lymphocytes, macrophages, and giant cells. On surgical removal of the PFCL from the subconjunctival space, the epithelial defect healed. The histopathologic and clinical evidence suggest that the inflammatory response and corneal epithelial ulceration were caused by the prolonged presence of PFCL in the subconjunctival space. To the best of our knowledge, PFCLs have not previously been reported to cause corneal epithelial defects or incite an inflammatory response in the human eye. PMID- 10532455 TI - Corneal stromal calcification after topical steroid-phosphate therapy. AB - Secondary corneal calcification involving the full thickness of the stroma is a rare potential complication of severe dry eye conditions, recurrent corneal ulcerations, chronic ocular inflammation, or multiple surgical procedures. We describe on a patient with unusual, hitherto unreported calcareous degeneration of the corneal stroma after topical steroid-phosphate therapy for chronic keratoconjunctivitis after Stevens-Johnson syndrome. The patient's serum levels of calcium and phosphorus were normal. Histopathologic and electron microscopic examination of the corneal button revealed mainly intracellularly located crystalline calcium deposits throughout all layers of the corneal stroma but sparing the Bowman layer. Energy-dispersive x-ray analysis confirmed the presence of calcium phosphate. The calcium deposits were closely associated with intracellular and pericellular accumulations of glycosaminoglycans. Our findings indicate that corneal stromal calcification may develop after topical steroid phosphate medication, and suggest a possible role of alterations in the glycosaminoglycan metabolism of stromal keratocytes in the calcification process. PMID- 10532456 TI - Orbital metastasis due to interval lobular carcinoma of the breast: a potential mimic of lymphoma. AB - A 53-year-old woman had an orbital mass composed of a neoplastic small round cell infiltrate and no apparent extraorbital primary tumor. Although the initial diagnosis was primary orbital lymphoma, a combination of mucin histochemistry and immunohistochemical staining for cytokeratin and estrogen receptors led to the discovery of an impalpable lobular carcinoma of the breast. We discuss how detailed histopathological assessment can lead to beneficial therapy. PMID- 10532457 TI - Medical treatment of operative corneal perforation caused by laser in situ keratomileusis. PMID- 10532458 TI - Infectious ulcerative keratitis after laser in situ keratomileusis. PMID- 10532459 TI - Use of a polyurethane patch for temporary closure of a sterile corneal perforation. PMID- 10532460 TI - Sterile mucopurulent conjunctivitis associated with the use of dorzolamide eyedrops. PMID- 10532461 TI - The first South American case of diffuse unilateral subacute neuroretinitis caused by a large nematode. PMID- 10532462 TI - Bilateral massive retinal hemorrhages in a 6-month-old infant: a diagnostic dilemma. PMID- 10532463 TI - Retinal periphlebitis in a patient with pineal germinoma. PMID- 10532464 TI - Massive orbital myiasis infestation. PMID- 10532466 TI - New support for ophthalmic drapes. AB - A new flexible support for ophthalmic drapes with the possibility of continuous oxygen supplementation was designed for use in patients undergoing eye surgery under local anesthesia. This new equipment is easy to handle and prevents contact between the patient's face and the ophthalmic drape. To prevent hypoxia of spontaneously breathing patients, the ambient air under the drapes can be supplemented with oxygen using this new equipment and no other devices. The equipment described here is advantageous for practical use in patients undergoing eye surgery under retrobulbar anesthesia. PMID- 10532465 TI - Ocular injuries caused by airsoft guns. PMID- 10532467 TI - Cystic epithelial ingrowth as a late complication of penetrating keratoplasty. PMID- 10532468 TI - Ectopic caruncle. PMID- 10532470 TI - Misleading figure legend PMID- 10532469 TI - Pneumatic displacement of subretinal hemorrhage without tissue plasminogen activator. PMID- 10532471 TI - Histological evidence of hydrogel fragmentation. PMID- 10532472 TI - Preface: Civic support and leadership as a catalyst for addressing the problem of urban asthma: the Otho S.A. Sprague Memorial Institute's Asthma Initiative. PMID- 10532473 TI - Chicago's response to the public health challenge of urban asthma. PMID- 10532474 TI - Asthma hospitalizations and mortality in Chicago: an epidemiologic overview. AB - STUDY OBJECTIVES: To characterize the patterns and correlates of asthma hospitalizations and mortality in Chicago. DESIGN: Cross-sectional analysis of discharge data for 1996 and mortality time trend data for the period from 1990 to 1997. SETTING: The city of Chicago, IL, with Cook County, IL, and US data employed for comparisons. POPULATION STUDIED: People who were hospitalized with a primary diagnosis of asthma and people whose underlying cause of death was asthma. INTERVENTIONS: None. MEASUREMENTS AND RESULTS: The 1996 asthma hospitalization rate for Chicago was 42.8 per 10,000, more than twice as high as suburban Chicago or US rates. Medicaid patients were overrepresented. Length of stay was longer for older patients and Medicaid patients. Age-adjusted asthma mortality in Chicago was 4.7 times higher in non-Hispanic blacks than in non Hispanic whites. The black/white asthma mortality ratio is 2.5:1 for the nation overall. Asthma mortality rates for Hispanics in Chicago were between those of non-Hispanic whites and blacks but have almost doubled during this decade. CONCLUSIONS: The rising asthma mortality and high asthma hospitalization rates in Chicago constitute a significant public health problem. Comorbidities more common in urban environments, such as substance abuse, may play a unique role in determining the distribution of adverse outcomes within Chicago's population. Asthma hospitalizations and deaths may vary in their risk profiles, and this should be taken into account when developing research and intervention strategies. PMID- 10532475 TI - The Chicago Asthma Surveillance Initiative: a community-based approach to understanding asthma care. AB - Nearly all of the asthma surveillance literature focuses on national-, regional-, or state-based estimates of prevalence, health-care utilization (specifically hospitalizations, emergency department, and ambulatory care visits), and mortality. Although these are important events, they reveal little about asthma's impact at the community level and provide little information that could be used to design specific interventions for improving clinical outcomes. A useful representation of asthma care across a community could guide an effective community response to the burden of asthma. The goal of the Chicago Asthma Surveillance Initiative (CASI) is to develop a community-wide surveillance program that characterizes and monitors asthma care in the Chicago area, beyond existing public health surveillance. To accomplish this, CASI surveyed Chicago area hospitals, emergency departments, primary care physicians, specialty care physicians, pharmacists, managed care organizations, the general public, and persons or families affected by asthma to learn about asthma care and its outcomes. A variety of techniques (including brochures, slide kits, and the Internet) were used to achieve rapid public dissemination of study findings. The value of this comprehensive community-based data surveillance effort will rest on how the community uses this information to stimulate new efforts to improve asthma care and reduce untoward outcomes. PMID- 10532476 TI - Asthma care practices, perceptions, and beliefs of Chicago-area primary-care physicians. Chicago Asthma Surveillance Initiative Project Team. AB - INTRODUCTION: Although primary-care physicians were a principal target audience for the National Asthma Education and Prevention Program (NAEPP), there is little published information describing the postguideline asthma care practices of these physicians or their willingness to embrace the NAEPP guidelines. This study examines asthma care practices of Chicago-area primary-care physicians and assesses these practitioners' perceptions and beliefs about several aspects of the NAEPP guidelines. METHODS: In 1997, a self-administered survey was mailed to a randomly selected 10% sample of Chicago-area general pediatricians, internists, and family practitioners. RESULTS: Surveys were returned by 244 of the 405 eligible Chicago-area primary-care physicians (60.2%) in the sample. Of these, 66 (27.6%) were pediatricians, 83 (34.7%) were general internists, and 90 (37.7%) were family practitioners. Physicians reported that 54.6 +/- 2.7% (mean +/- SE) of patients with newly diagnosed asthma have spirometry performed as part of their initial evaluation. For patients with moderate persistent asthma, prescribing of inhaled corticosteroids varied by patient age, with 60.5% of physicians routinely prescribing them for patients < 5 years, compared with 95.7% of physicians prescribing them for patients > or = 5 years. Awareness of the NAEPP guide-lines among these physicians was high, with 88.5% reporting that they have heard of the guidelines, and 73.6% reporting having read them. Of patients with moderate or severe persistent asthma, physicians estimated that 47.7 +/- 2.7% were given written treatment plans. CONCLUSION: Several aspects of the NAEPP guidelines appear to have been incorporated into clinical practice by Chicago area primary-care physicians, whereas other recommendations do not appear to have been readily adopted. This information suggests areas for interventions to improve primary care for asthma in the Chicago area. PMID- 10532477 TI - Asthma care practices, perceptions, and beliefs of Chicago-area asthma specialists. Chicago Asthma Surveillance Initiative Project Team. AB - INTRODUCTION: Few studies have closely explored how well physicians who consider themselves specialists in asthma adhere to national guideline recommendations for the diagnosis and treatment of asthma. The purpose of this study is to characterize current knowledge, attitudes, beliefs, and self-reported treatment practices of the asthma specialists working in one large metropolitan area. METHODS: In 1997, a cross-sectional survey was mailed to asthma specialists (allergists or pulmonologists) engaged in direct patient care with a practice location in the Chicago area (Cook County or one of the five surrounding counties). An approximately 50% random sample of asthma specialists was surveyed. The survey included items on (1) asthma diagnosis; (2) clinical monitoring of asthma patients; (3) pharmacologic and nonpharmacologic asthma treatment; (4) opinions and beliefs about asthma treatment options and reasons for referrals; (5) involvement in continuing medical education; (6) experiences with managed care; (7) use of asthma practice guidelines; (8) demographic information about the respondents; and (9) characteristics of the practice settings. RESULTS: A total of 113 eligible surveys were returned (response rate, 72.0%). Ninety-nine percent of the respondents indicated they would prescribe inhaled corticosteroids for patients > or = 5 years old with moderate persistent asthma, and 85.5% would prescribe them for patients < 5 years old. The respondents reported that 71.2% of their patients with moderate or severe persistent asthma were routinely given written treatment plans. The use of these plans was reported more frequently by allergists than pulmonologists (77.6% vs 58.9%, p = 0.01). Nearly half of the respondents were involved in the development of hospital-based asthma programs; fewer (14.9%) were involved in developing asthma programs for managed care organizations. A majority (63.4%) of the physicians had given a formal professional education presentation on asthma in the past year. A majority of the respondents who care for patients under managed care contracts reported that these patients have encountered barriers to access in seeking specialty care. CONCLUSION: The results suggest that asthma specialists in the Chicago area are providing asthma care that is, in many ways, consistent with national guidelines. However, there are also important differences in care that are not consistent with the guideline recommendations. Perhaps even more notable are differences in reported asthma care between the two subspecialty groups of allergists and pulmonologists. The effect of these differences on the management of persons with asthma is not known. It is hoped that information from this community-based survey will serve to catalyze discussions among Chicago-area asthma specialists as to how they might envision improving care for persons with asthma in their community. PMID- 10532478 TI - Characteristics of asthma care provided by hospitals in a large metropolitan area: results from the Chicago Asthma Surveillance Initiative. AB - INTRODUCTION: Little is known of the approaches of United States hospitals to the management of persons with asthma. The purpose of this study is to characterize the extent to which hospitals within a large community have implemented various types of asthma-specific health-care delivery processes. METHODS: A cross sectional, self-administered survey was mailed to a "key informant" in asthma care at each of the hospitals in the Chicago area. The survey instrument covered the following content areas: asthma-related inpatient services, asthma-related outpatient services, selected asthma-related quality improvement activities, and asthma-related community outreach. The survey was administered between August 1996 and January 1997. RESULTS: Data were collected from respondents at 59 of the 89 eligible hospitals, yielding a response rate of 66.3%. Of the responding hospitals, 42.4% indicated they had clinical practice guidelines for inpatient asthma management, and 37.3% reported using critical pathways. Four selected aspects of bedside care were also explored. All of the responding hospitals reported routine provision of nebulization therapy at the bedside, and nearly all routinely obtained peak flow measurements (96.6%). In the area of patient instruction, 93.2% provided bedside evaluation of proper inhaler technique, and 86.4% routinely provided instruction on the use of peak flowmeters. Only 54.0% of the hospitals reported routinely administering some type of asthma education program prior to discharge. The hospitals with clinical practice guidelines in place were also more likely to have critical pathways (p < 0.01); to have asthma specific ICU policies/guidelines/critical pathways (p < 0.01); to provide bedside instruction on the use of peak flowmeters (p < 0.01); to provide an asthma education (p < 0.01) prior to discharge; and to conduct utilization review. Very few hospitals indicated that they had community outreach programs for asthma care. CONCLUSION: The results of this survey suggest that among Chicago-area hospitals appropriate bedside care for persons with asthma is provided, but there are large variations in other types of asthma services and programs. The hospitals that have adopted asthma clinical practice guidelines are more likely to have other asthma-specific quality improvement activities than hospitals without guidelines. This relationship between use of guidelines and quality of services needs further exploration, as it may prove to be an important marker for hospitals with staff that are interested in improving asthma care. PMID- 10532479 TI - Asthma care practices in Chicago-area emergency departments. Chicago Asthma Surveillance Initiative Project Team. AB - INTRODUCTION: Emergency departments (EDs) represent an important source of asthma care, yet there are few studies detailing how ED asthma practices vary and to what extent EDs meet expectations of national asthma guidelines. The purpose of this study is to characterize ED care for persons with asthma in a single large community. METHODS: During 1996 and 1997, a cross-sectional, self-administered survey to characterize asthma care practices was conducted among medical directors of the 89 EDs serving the Chicago metropolitan area (six counties). The survey topic areas included asthma-specific demographics and selected utilization statistics; assessment practices; treatment practices; discharge and follow-up activities; and familiarity with, attitudes toward, and utilization of guidelines/ protocols. RESULTS: Sixty-four EDs completed surveys, for a response rate of 71.9%. Ninety-four percent of the respondents were ED medical directors. As part of assessment, peak flow measurements, while common, were used less frequently than pulse oximetry. The average (+/- SE) estimated length of stay for asthma care was 3.0 +/- 0.1 h, and average disposition time (ie, the decision to admit) was 2.5 +/- 0.2 h. Systemic steroids (either i.v. or p.o.) were estimated to be given to 73.2 +/- 3.9% of patients during their ED visits. Systemic steroids were prescribed for 55.9 +/- 3.5% of patients at time of discharge. Only 57.0 +/- 5.4% of patients were estimated to have received any type of written asthma educational materials. Approximately 25% of patients were reported to have been given a detailed follow-up appointment at the time of discharge. CONCLUSION: The results reveal that the medical directors reported many of the Chicago-area EDs as providing asthma care that is consistent with key aspects of national guidelines. However, in certain critical areas of care, the EDs demonstrate a high degree of variation, often with the community falling short of guideline recommendations. By identifying these variations in asthma care, it is now possible to target specific goals for community-wide asthma quality improvement among the EDs in the Chicago metropolitan area. PMID- 10532480 TI - A survey of asthma care in managed care organizations: results from the Chicago Asthma Surveillance Initiative. AB - INTRODUCTION: Managed care, both via staff model health maintenance plans and nonstaff model plans, has become a major source of health-care funding in the United States. However, very little is known about the asthma-specific products and services offered by these plans. The purpose of this study is to examine the asthma-specific products and services offered by managed care within the Chicago area. METHODS: Between December 1997 and February 1998, a self-administered survey was mailed to the medical directors of the 19 managed care organizations (MCOs) in the Chicago area. The survey covered the following content areas: general characteristics of the MCOs, asthma-related services, monitoring of asthma care, and asthma-related quality improvement efforts. The medical directors were asked to respond separately for staff model capitated plans, nonstaff model capitated plans, and noncapitated plans. RESULTS: Responses were received from 13 of the 19 eligible Chicago-area MCOs (a response rate of 68.4%). Three of the responding MCOs (23.1%) offered a staff model plan, 11 (84.6%) offered a nonstaff model capitated plan, and 6 offered some type of noncapitated plan. Asthma education programs, although available in all plan types, were offered much less frequently in the nonstaff model capitated and noncapitated plans, 36.4% and 33.3%, respectively. Asthma case management programs were also available in some, but not all of the health plans. Only 54.5% of the nonstaff model capitated health plans promoted the use of asthma practice guidelines. Among the responding MCOs, asthma quality improvement efforts related to National Committee on Quality Assurance accreditation were infrequent in 1995. Sixty-one percent of the MCOs reported that program development for improving asthma care was a very high priority relative to programs for other health conditions. CONCLUSION: The results of this study suggest that many, but not all, of the basic elements of asthma care services are offered by the MCOs in the Chicago area. Findings from this study also suggest ways in which asthma-related product and service delivery might be changed to improve outcomes for asthma in this community. PMID- 10532481 TI - Development of a survey of asthma knowledge, attitudes, and perceptions: the Chicago Community Asthma Survey. Chicago Asthma Surveillance Initiative Project Team. AB - Little is known about the general public's perception of the diagnosis of asthma and the impact of asthma on individuals, their families, and their communities. In addition, there appear to be no published survey instruments specifically designed to gain insights into how the general public perceives asthma. The purpose of this paper is to describe the development of such an instrument, the Chicago Community Asthma Survey (CCAS)-32. Development began with two qualitative steps. First, a review of the published literature guided the initial instrument construction (Step 1). Content domains were chosen based on clinical input and the Health Belief Model. Most items were derived from existing instruments. To assess content validity, cognitive interviews and expert reviews were conducted (Step 2). Items were added, modified, and deleted based on the information gathered at each of these steps. In the next step, item performance measurement (Step 3), testing of two samples provided quantitative data to further inform item reduction. Items with uniform correct responses or responses lacking in variability were excluded. The result of this three-step process was a 32-item survey of asthma knowledge, attitudes and perceptions, the CCAS-32. The introduction to the survey was subsequently modified to minimize respondent bias (Step 4). In conclusion, the CCAS-32 was constructed with input from experts in asthma and individuals from the Chicago area. The items in the CCAS-32 appear to have both face validity and acceptable performance characteristics. PMID- 10532482 TI - The effects of asthma experience and social demographic characteristics on responses to the Chicago Community Asthma Survey-32. Chicago Asthma Surveillance Initiative Project Team. AB - INTRODUCTION: The Chicago Community Asthma Survey (CCAS-32) is an instrument for characterizing the general public's knowledge, attitudes, and beliefs related to asthma. The purpose of this study was to examine the effects of asthma experience and social demographic characteristics on asthma awareness among the general public. METHODS: The CCAS-32 consists of 21 dichotomous items, designed primarily to test asthma knowledge, and 11 Likert-scale items, focusing on asthma attitudes and beliefs. From December 1997 through February 1998, a random-digit dialing method was used to administer the CCAS-32 via a telephone survey of Chicago-area (seven-county) residents > or = 18 years. Each respondent's asthma experience was classified as "person with asthma," "family/household experience," or "no/low asthma experience." Demographic variables included sex, age, education, race/ethnicity, urban vs suburban residence, and income. RESULTS: Five hundred sixty-eight Chicago-area residents completed the survey (response rate of 40.6%). Of these, 43.3% were aged 35 to 64 years, 71.3% were women, 66.7% were white, and 71.3% had completed at least some college. Sixty-two percent had no or low asthma experience, 28.5% had family or household experience, and 9.5% were persons with asthma. The mean percentage (+/- SE) of correct, or desirable, responses to asthma knowledge questions was 71.9 +/- 0.5%, with a range from 31.9 to 95.1%. The mean percentage of desirable responses differed significantly between persons with no or low asthma experience, family or household asthma experience, and persons with asthma (70.0 +/- 0.6%, 74.0 +/- 0.9%, and 77.7 +/- 1.2%, respectively, p < 0.01 for trend). Social demographic factors also appeared to result in statistically significant differences in the responses to many items. Of the demographic variables studied, age and education appeared to have the strongest effect on responses to knowledge items, with statistically significant differences in responses seen for 10 (47.6%) and 8 (38.1%) of the 21 dichotomous items. Race or ethnicity and education were each associated with differences in responses for 7 of the 11 Likert-scale items (63%). CONCLUSIONS: The results of this study suggest that the CCAS-32 can detect meaningful differences between groups with different degrees of asthma experience (ie, discriminative validity). Using the CCAS-32, it may be possible to identify subpopulations with differences in asthma awareness, thus providing guidance for the design of messages to target community and public awareness of asthma. PMID- 10532483 TI - The Chicago Asthma Consortium: a community coalition targeting reductions in asthma morbidity. AB - The problem of asthma in Chicago remains a complex one, and it is too early to know whether any programs and efforts have had a discernible effect, but the Chicago Asthma Consortium continues to expand its membership and to define its mission. The successes have come from harnessing the passion of the individual members to move the projects forward. As the focus of the consortium moves to addressing system-wide problems in asthma care and the delivery of that care, the consortium is undertaking the construction of a guide for future efforts. In this way, the consortium will fulfill its vision of creating a comprehensive, community-wide plan for the management of asthma, impacting on the unacceptable current levels of morbidity and mortality of the disease. PMID- 10532484 TI - A collaborative model to enhance the functioning of the school child with asthma. PMID- 10532485 TI - Identifying asthma patient education materials that support National Heart, Lung and Blood Institute guidelines. PMID- 10532487 TI - Targeting asthma in Chicago: community stories. PMID- 10532486 TI - The Chicago Emergency Department Asthma Collaborative. PMID- 10532488 TI - Health education program to control asthma in multiethnic, low-income urban communities: the Chicago Health Corps Asthma Program. PMID- 10532489 TI - The American Red Cross of Greater Chicago's Asthma Program. PMID- 10532490 TI - The Lawndale Christian Health Center Asthma Education Program. PMID- 10532491 TI - The Infant Welfare Society's Asthma Management Project. PMID- 10532492 TI - Bethany Hospital's asthma program. PMID- 10532493 TI - The Henry Horner Pediatric Asthma Program. PMID- 10532494 TI - Advocate Health Care's approach to adult asthma. PMID- 10532495 TI - United HealthCare of Illinois: working to improve asthma care. PMID- 10532496 TI - Restructuring asthma care in a hospital setting to improve outcomes. AB - STUDY OBJECTIVES: To restructure asthma care as the pilot program in hospital wide redesign aimed at providing better and more standardized care. We chose asthma care to begin our reorganization because it is the highest-volume diagnosis at our hospital and it involves a broad spectrum of services. DESIGN: Key elements of our restructuring included the following: (1) establishing a pulmonary unit with expanded bed capacity from 8 to 22 beds for asthma patients; (2) standardized treatment protocols; (3) availability of direct admission by primary care physicians who maintained management of their patients with the option of consultation with a specialist; and (4) use of case managers who helped patients and their families overcome obstacles to optimum care. SETTING: A hospital serving a high proportion of Medicaid patients. PATIENTS/PARTICIPANTS: Children with asthma and their families. INTERVENTIONS: Standardized care for asthma; use of case managers to facilitate adherence to treatment. RESULTS: With the restructured asthma care program, parent satisfaction with treatment was sustained; the average length of stay and use of the emergency department (ED) were reduced; observation unit use increased; and there were fewer readmissions to both the inpatient unit and the ED. CONCLUSIONS: We conclude that an inner city hospital can provide optimum care for asthma patients by standardizing treatment, aggregating asthma patients in one location, and providing education and follow-up through the use of case managers. The protocol shifts some costs from expensive services such as the pediatric ICU and the ED to less costly case management and outreach personnel. In the long run, this allocation of resources should help to lower costs as well as improve quality of care. PMID- 10532497 TI - Chicago community-based asthma intervention trial: feasibility of delivering peer education in an inner-city population. AB - The most effective means of educating children with asthma and their families has not been clearly demonstrated in previous studies. Peer education is uniquely suited to the complex problems encountered in underserved populations. The purpose of this study was to show the feasibility of delivering a peer education program for children with asthma and the effect of the program on indoor allergen levels in an inner-city population in Chicago. Overall, the program was well received. Baseline allergen levels were consistent with some previous studies in showing low levels of mite allergens and high levels of cockroach allergens, with 79.6% of samples having levels > 8 U/g. A total of 28.2% of samples had cat allergen levels > 2 microg/g, although only 9.7% of homes had cats, confirming previous reports that cat allergen is ubiquitous. Mold levels were seasonal, with the highest levels in the summer. Results from this study suggest that intervention programs should focus more on elimination of cockroaches than was previously appreciated, while minimizing the use of pesticides, and on identification of the sources of cat allergen. Structural and psychosocial issues in homes need to be addressed in future studies. This study has demonstrated the feasibility of delivering peer education in a inner-city population and highlighted the need for comprehensive intervention strategies addressing complex issues facing underserved neighborhoods. PMID- 10532498 TI - Validation of the Brief Pediatric Asthma Screen. AB - STUDY OBJECTIVES: The purpose of this study was to confirm the validity of a brief screen for pediatric asthma in schools. BACKGROUND: Asthma is the most common chronic disease of childhood, yet the frequency with which this condition is recognized among school-aged children varies widely. Several methods are used to increase the accuracy of detection of asthma, but many are cumbersome and difficult to apply on a large scale. DESIGN: We elected to validate a five question instrument, the Brief Pediatric Asthma Screen (BPAS), to screen for the presence of asthma among children attending school in Region 5 of the Chicago school district, where the schools report a 2.7% frequency of asthma. The questionnaire was distributed to the parents of grade-school children at the time of report-card pick-up. SETTING: A clinical assessment was performed on a selected group of children whose parents completed the questionnaire in a school and in a hospital outpatient clinic. PARTICIPANTS: Of 4,147 questionnaires that we distributed, 1,796 (43%) were returned. We excluded 341 children (19% of the total sample) whose parents reported that they had been diagnosed with asthma. The remaining pool indicated that the children of 183 responders (10%) had symptoms suggestive of asthma, while 1,272 parents (71%) indicated that their children did not have symptoms of asthma. MEASUREMENTS AND RESULTS: We selected 90 of the respondents who did not indicate that their children had a diagnosis of asthma. Of this group, 81 completed the validation, in which their responses suggested symptoms of asthma (n = 34) or no asthma symptoms (n = 47). The children of these respondents were given a blinded clinical evaluation consisting of history, physical examination, and spirometry. The survey demonstrated a sensitivity of 75% and a specificity of 81.2% for the presence of asthma among those who were unaware of the diagnosis. CONCLUSIONS: The BPAS is brief, can be filled out by parents, and appears accurate in detecting asthma. PMID- 10532499 TI - A pilot study describing local residents' perceptions of asthma and knowledge of asthma care in selected Chicago communities. AB - STUDY OBJECTIVES: To understand inner-city Chicago residents' perception of the prevalence and severity of asthma as well as their knowledge of asthma control and management. DESIGN: Cross-sectional survey using a random digital telephone dialing method. SETTINGS: Five inner-city Chicago communities where a high prevalence and mortality of asthma have been recognized. PARTICIPANTS: All the residents in the selected communities with a residential telephone had an equal opportunity to be surveyed. MEASUREMENTS AND RESULTS: The unit of measurement was the household. Only one adult member (age 18 or older) in any randomly selected household was interviewed. The survey included questions modified from the Chicago Asthma Surveillance Initiative study. A total of 2,322 phone calls with 527 successful contacts were made over 1,938 distinct phone lines, resulting in a response rate of 175 of 527 calls (33.2%). Seventy-nine of the participants (45.1%) reported that at least one of their family members (including themselves) has asthma. Eight persons (4.6%) reported asthma as one of the top three health concerns in their community. Of the top three health reasons mentioned for children's being absent from school, only seven persons (4%) mentioned asthma. Participants were unlikely to perceive that the problems with access to asthma care and environmental triggers for asthma in their communities were any worse compared with other communities. Participants having family members with diagnosed asthma scored no better when asked general-knowledge questions about asthma or its signs and triggers than those without a family member having asthma. CONCLUSIONS: The participants' knowledge and beliefs about the seriousness of asthma revealed in this study appeared unlikely to enhance or support compliance with the challenging requirements of the National Asthma Education and Prevention Panel guidelines. The study was conducted with a small sample, and the results should be carefully interpreted. PMID- 10532500 TI - The National Asthma Education and Prevention Program: partnering with local asthma coalitions to implement the guidelines. PMID- 10532501 TI - Gender differences in the relation between ST-T-wave abnormalities at baseline electrocardiogram and stress myocardial perfusion abnormalities in patients with suspected coronary artery disease. AB - The presence of ST-T-wave abnormalities in the resting electrocardiogram was reported as a predictor of coronary artery disease (CAD) and increased morbidity and mortality. However, the independent value of ST-T abnormalities for predicting the presence and severity of perfusion abnormalities during stress testing has not been studied in a homogenous patient group without known CAD. We evaluated the relation between resting ST-T abnormalities and myocardial perfusion abnormalities in 246 patients (age 59 +/- 13 years, 114 men and 132 women) without known CAD or previous myocardial infarction referred for evaluation of possible myocardial ischemia by dobutamine (up to 40 microg/kg/min) stress sestamibi or tetrofosmin single-photon emission computed tomographic imaging. Resting ST-T abnormalities were present in 123 patients, whereas 123 patients with normal resting electrocardiograms served as a matched control group. Abnormal myocardial perfusion (fixed or reversible perfusion defects) was detected in 72% of men with and in 35% of men without resting ST-T abnormalities (p <0.0001), whereas the prevalence of myocardial perfusion abnormalities was not different in women with and without resting ST-T abnormalities (27% vs 23%, p = NS). In the entire population, independent predictors of an abnormal perfusion by multivariate analysis of clinical characteristics and risk factors were male gender (p <0.001, chi-square 10.5) and resting ST-T abnormalities (p <0.05, chi square 3). Separate analysis of patients based on gender revealed resting ST-T abnormalities as independent predictors of abnormal perfusion in men (p <0.05, chi-square 4) but not in women. Stress perfusion defect score was higher in men with than without ST-T abnormalities (887 +/- 545 vs 207 +/- 180, p <0.001). It is concluded that resting ST-T wave abnormalities are associated with a higher prevalence and severity of resting and dobutamine-induced myocardial perfusion abnormalities in men but not in women. Resting ST-T wave abnormalities are powerful predictors of compromised myocardial perfusion independent of other clinical risk factors of CAD in men. PMID- 10532502 TI - Comparison of insulin response to intravenous glucose in healed myocardial infarction, in "cooled-off" unstable and stable angina pectoris, and in healthy subjects. AB - Fasting and postglucose hyperinsulinemia are recognized risk factors for acute coronary events. The insulin reactivity of patients with acute coronary syndromes, however, has not been carefully compared with that of patients with chronic stable angina. We used Bergman's minimal model to analyze the insulin response to intravenous glucose in 21 subjects: 8 patients with previous (>3 months) acute coronary syndrome but no effort-related angina; 6 patients with stable effort angina but no prior acute event; and 7 healthy controls. Diabetes mellitus, systemic hypertension, dyslipidemias, and obesity were excluded. All patients underwent coronary angiography. Insulin sensitivity, glucose effectiveness, and glucose tolerance were determined from insulin and glucose concentrations measured frequently up to 3 hours after a 0.33 g/kg intravenous glucose bolus. Patients with previous unstable angina or acute myocardial infarction had less extensive disease at angiography than patients with stable angina (p = 0.007). Both patient groups had higher basal and 180-minute insulinemia than controls (p <0.0007). However, patients with stable angina did not differ significantly from controls with regard to early and late insulinemic response to glucose. In contrast, patients with previous acute onset of ischemia had significantly greater 180-minute integrated insulinemia (p = 0.04) and reduced insulin sensitivity (p = 0.05) after the glucose challenge than did the stable angina group. These data suggest that patients with acute presentation of coronary artery disease, compared with patients with uncomplicated chronic stable angina, have an impaired insulin response to glucose despite less extensive coronary disease at angiography. PMID- 10532503 TI - Comparison of clinical and genetic variables of cardiac events associated with loud noise versus swimming among subjects with the long QT syndrome. AB - Acute auditory stimuli and swimming activities are frequently associated with syncope, aborted cardiac arrest, and death in the long QT syndrome (LQTS). We investigated the clinical and genetic findings associated with cardiac events precipitated by these arousal factors. The study population involved 195 patients with an index cardiac event associated with a loud noise (n = 77) or swimming activity (n = 118). Patients with events associated with loud auditory stimuli were older at their index event and were more likely to be women than patients who experienced events during swimming-related activities. Patients with an index event associated with loud noise were likely to have subsequent events related to auditory stimuli; patients with an index event associated with swimming were likely to have recurrent events related to swimming or physical activities. Family patterning of auditory and swimming and/or physical activity-related events was evident. Genotype analyses in 25 patients revealed a significant difference in the distribution of index cardiac events by genotype (p <0.001), with all 19 patients with swimming-related episodes associated with LQT1 genotype and 5 of 6 patients with auditory-related events associated with LQT2 genotype. The clinical profile and genotype findings of patients with LQTS who experience cardiac events related to acute auditory stimuli are quite different from those who experience events accompanying swimming activities. PMID- 10532504 TI - Regional endocardial mapping of spontaneous and induced atrial fibrillation in patients with heart disease and refractory atrial fibrillation. AB - We performed simultaneous catheter mapping of right and left atrial regions at onset and during sustenance of spontaneous atrial fibrillation (AF) in patients with ischemic and/or hypertensive heart disease. Seventeen patients with structural heart disease had spontaneous and electrically induced AF episodes mapped from their onset simultaneously in multiple right and left atrial regions. Atrial premature complexes (APCs) that initiated spontaneous AF had coupling intervals ranging from 260 to 400 ms (mean 332 +/- 61), most commonly arising from the lateral right atrium (31%), right atrioventricular junction (13%), atrial septum (6%), superior left atrium (25%), or inferior left atrium (25%). APC morphology on surface electrocardiograms did not correlate with origin in specific atrial regions. The earliest regions of atrial activation for the first AF cycle were the lateral right atrium (n = 5), superior left atrium (n = 4), distal or mid coronary sinus (n = 4), atrial septum (n = 2), and right atrioventricular junction at the His bundle location (n = 2). Spontaneous AF at onset usually showed discrete but irregular electrograms at virtually all right and left atrial sites mapped, with a reproducible region of AF initiation in all 8 patients with multiple events. The region of earliest atrial activation at spontaneous AF onset was in close proximity to the APC origin in 15 of 16 patients (94%), and 39 of 40 episodes (97%) mapped. Stable patterns of right and left atrial activation were observed at AF onset in 14 patients. Induced AF elicited with right atrial stimulation demonstrated different sites of earliest regional atrial activation at onset compared with spontaneous AF events in 4 of 8 patients. However, discrete intracardiac electrograms were also present in induced AF in all of the mapped atrial regions. Furthermore, the site of extrastimulus delivery in induced AF was also found to be in close proximity to the earliest region of atrial activation for the first AF beat. We conclude that spontaneous AF is initiated by APCs arising in different right or left atrial regions in patients with structural heart disease and the initial region of atrial activation in AF is in proximity to the region of APC origin. Organized and repetitive electrical activation is frequently observed in both right and left atria at AF onset. Although electrically induced AF may have different activation patterns than spontaneous AF at onset in many patients, both types of AF demonstrate organization and earliest atrial activation in proximity to the initiating APC. PMID- 10532505 TI - Relation of maximum blood pressure during exercise and regular physical activity in normotensive men with left ventricular mass and hypertrophy. MARATHOM Investigators. Medida de la Actividad fisica y su Relacion Ambiental con Todos los Lipidos en el HOMbre. AB - The relation between maximum systolic blood pressure (BP) during exercise and left ventricular (LV) mass is controversial. Physical activity also induces LV mass increase. The objective was to assess the relation between BP response to exercise and LV mass in normotensive men, taking into account physical activity practice. A cross-sectional study was performed. Three hundred eighteen healthy normotensive men, aged between 20 and 60 years, participated in this study. The Minnesota questionnaire was used to assess physical activity practice. An echocardiogram and a maximum exercise test were performed. LV mass was calculated and indexed to body surface area. LV hypertrophy was defined as a ventricular mass index > or =134 g/m2. BP was measured at the moment of maximum effort. Hypertensive response was considered when BP was > or =210 mm Hg. In the multiple linear regression model, maximum systolic BP was associated with LV mass index and correlation coefficient was 0.27 (SE 0.07). Physical activity practice and age were also associated with LV mass. An association between hypertensive response to exercise and LV hypertrophy was observed (odds ratio 3.16). Thus, BP response to exercise is associated with LV mass and men with systolic BP response > or =210 mm Hg present a 3-times higher risk of LV hypertrophy than those not reaching this limit. Physical activity practice is related to LV mass, but not to LV hypertrophy. PMID- 10532506 TI - Tolerability of extended duration intravenous milrinone in patients hospitalized for advanced heart failure and the usefulness of uptitration of oral angiotensin converting enzyme inhibitors. AB - Milrinone is a phosphodiesterase inhibitor that has been shown to improve hemodynamic parameters in patients with class III to IV heart failure when administered intravenously for < or =48 hours. This study examines the tolerability of long-term intravenous milrinone therapy and assesses its utility in allowing upward titration of oral vasodilator agents. A retrospective review of hospital records identified 63 patients who underwent hemodynamic monitoring and received intravenous milrinone for >24 hours in a critical care setting. Hemodynamics and medications were recorded before and after 24 hours of milrinone therapy. Additional medications, as well as any adverse events, were recorded throughout milrinone therapy. The mean dose of milrinone was 0.43 +/- 0.10 microg/kg/min, with a mean duration of 12 +/- 15 days (range 1 to 70). Therapy was continued for >48 hours in 89% of patients. After 24 hours of milrinone therapy, patients exhibited significant improvements in pulmonary artery pressures, pulmonary capillary wedge pressures, and cardiac index. When compared with baseline, significantly more patients received angiotensin-converting enzyme (ACE) inhibitors after 24 hours of milrinone and at the end of milrinone therapy (67% vs 86%, p <0.01). Likewise, significantly more patients also received oral hydralazine and/or nitrates at the end of milrinone therapy (38% vs 65%, p <0.01) when compared with baseline. The mean doses of most oral medications at the 3 time periods were similar. The ACE inhibitor dose was significantly higher at the end of milrinone therapy when compared with baseline, and hydralazine dose was significantly higher at the end of therapy when compared with 24 hours. Few adverse effects were noted, with only 10% of patients experiencing symptomatic ventricular tachycardia and 2 patients with significant hypotension requiring discontinuation of the drug. The adverse events were similar in the group of patients who received milrinone for > or =7 days compared with the entire cohort. Milrinone was well tolerated over the long term in a controlled inpatient setting, and allowed uptitration of oral vasodilator therapy. PMID- 10532507 TI - Association between hemodynamic impairment and Cheyne-Stokes respiration and periodic breathing in chronic stable congestive heart failure secondary to ischemic or idiopathic dilated cardiomyopathy. AB - Irregular breathing occurs frequently in patients with congestive heart failure (CHF) both during daytime and nighttime. Many factors are involved in the genesis of these breathing abnormalities, but the role of the hemodynamic impairment remains controversial. This study investigated the relation between worsening ventricular function and the frequency of respiratory disorders in patients with mild to severe CHF. One hundred fifty patients with CHF (mean age 53 +/- 8 years, left ventricular (LV) ejection fraction 26 +/- 7, in New York Heart Association [NYHA] classes II to IV, and who underwent stable therapy for > or =2 weeks) were studied. Analysis of instantaneous lung volume signal and arterial oxygen saturation during awake daytime revealed a normal respiratory pattern in 63 patients, whereas 87 had a persistent alteration of breathing, with a typical Cheyne-Stokes respiration (CSR) in 42 and periodic breathing (PB [oscillation of tidal volumes without apnea]) in 45 patients. Patients with PB and CSR showed a more pronounced hemodynamic impairment with a significantly reduced cardiac index, an increased pulmonary arterial wedge pressure, and a longer lung-to-ear circulation time (LECT) compared with patients with normal respiratory patterns. In a logistic regression model that included all of the variables significantly associated with breathing disorders, cardiac index and LECT emerged as the major determinants of CSR. In those patients with LECT > or =30 seconds (upper quartile) and cardiac index < or =1.9 L/min/m2 (lower quartiles), the incidence of CSR was significantly higher (69%) than in patients with lower LECT and higher cardiac index (14%, p <0.001). In conclusion, abnormalities of breathing activity during daytime are significantly associated with a prolonged circulation time and a more severe impairment of systolic and diastolic LV indexes. PMID- 10532508 TI - Exaggerated initial response to warfarin following heart valve replacement. AB - The response to initiation of oral anticoagulants at a usual dose of 5 mg of warfarin has been retrospectively evaluated in patients following heart valve replacement (HVR). Patients starting oral anticoagulants after HVR have a lower target International Normalized Ratio (INR) (1.5 to 2.6) until the pacing wires are removed after operation. The mean daily doses and INR responses after HVR and nonsurgical patients were retrospectively compared during the first 5 days of warfarin treatment. In a subset from both groups, the mean dose of warfarin was correlated with age, body weight, and albumin levels. Eighty-four HVR and 32 nonsurgical patients were studied. The mean daily warfarin dosage was 3.29 +/- 1.29 mg after HVR and 4.96 +/- 1.76 mg in controls (p <0.001), and the mean INRs 2.08 +/- 0.60 and 1.60 +/- 0.54, respectively (p <0.001). Of the HVR patients and controls, 48.8% and 21.8%, respectively, exceeded the upper level of the targeted range (p = 0.014), 86.9% and 40.6% had the dose reduced after the first 5 mg (p <0.001), and 54.7% and 28.1%, respectively, had warfarin withheld for at least 1 day (p = 0.015). Thirty-nine patients were included in the subset analysis. Patients with serum albumin levels <35 g/L required significantly less warfarin (3.84 mg/day) than patients with levels > or =35 g/L (5.37 mg/day; p <0.05). Thus, patients starting oral anticoagulation after HVR are significantly more sensitive to warfarin than nonsurgical patients. Patients with serum albumin levels below the normal values require less warfarin than patients with normal values during the initial phase of treatment. PMID- 10532509 TI - Angiotensin-converting enzyme gene polymorphism influences degree of left ventricular hypertrophy and its regression in patients undergoing operation for aortic stenosis. AB - Insertion (I)/deletion (D) polymorphism of the angiotensin-converting enzyme (ACE) gene has been associated with increased left ventricular hypertrophy (LVH) in patients with cardiomyopathy and congestive heart failure. Patients with aortic stenosis (AS) have varying degrees of LVH at a given valve area. The aim of this study was to examine the relation between ACE gene polymorphism and the degree of LVH in patients undergoing operation for AS. Eighty-two patients who underwent operation for AS with a stentless valve were followed prospectively with echocardiographic assessments of left ventricular mass index (LVMI). ACE gene polymorphism was determined by polymerase chain reaction. The genotype (DD, ID, and II) frequency was the same as in healthy controls. The pressure difference across the aortic valve did not differ between genotypes. Patients with the DD genotype of the ACE gene had a higher LVMI (197 +/- 47 g/m2) preoperatively than those with ID (175 +/- 41 g/m2) or II (155 +/- 43 g/m2) genotypes (p = 0.01). LVMI decreased significantly in DD (p <0.001) and ID (p <0.001) genotypes but not in the II genotype during follow-up (mean 15 months). There was a significant difference in regression of LVMI over time between genotypes (p = 0.0056), with no significant difference between genotypes at follow-up. The DD genotype of the ACE gene is associated with increased preoperative LVH in patients treated surgically for AS. The DD genotype appears to be an important factor which increases hypertrophic myocardial reactivity to pressure overload. PMID- 10532510 TI - Immediate effect of aortic valve replacement for aortic stenosis on left ventricular diastolic chamber stiffness. AB - Diastolic dysfunction is common after coronary artery bypass surgery, and we hypothesized that left ventricular (LV) hypertrophy associated with aortic stenosis may lead to worsening LV diastolic function after aortic valve replacement for aortic stenosis. Transesophageal echocardiographic LV images and simultaneous pulmonary arterial wedge pressures were used to define the LV diastolic pressure cross-sectional area relation before and immediately after aortic valve replacement for aortic stenosis in 14 patients. In all patients, LV diastolic chamber stiffness increased, as evidenced by a leftward shift in the LV diastolic pressure cross-sectional area relation. At comparable LV filling (pulmonary arterial wedge) pressures the mean LV end-diastolic cross-sectional area preoperatively was 17.9 +/- 1.7 cm2, but decreased by 32% after aortic valve replacement to 12.1 +/- 1.2 cm2 (p = 0.0001). In conclusion, after aortic valve replacement, diastolic chamber stiffness increased in all patients. PMID- 10532511 TI - Minimally invasive direct access for repair of atrial septal defect in adults. AB - This report documents our early experience with minimally invasive direct-access surgical repair of atrial septal defect (ASD) in adults. We have developed minimally invasive techniques for direct-access ASD repair in adults while maintaining the efficacy of the open operative procedure. Between June 1996 and September 1998, 59 consecutive patients underwent repair of ASD, 34 (58%) of whom underwent minimally invasive direct-access surgical closure of ASD through a right parasternal, submammary, or upper hemisternotomy incision. Twenty-three (68%) were secundum type ASD, 5 (15%) were sinus venosus types, 2 (6%) were primum types, and 4 (122%) were patent foramen ovales. Twenty-six (77%) were women (mean age 39 +/- 15 years, range 18 to 79). The mean pulmonary-to-systemic shunt ratio (Qp/Qs) was 2.3 +/- 0.6 (n = 15). There were no operative or late deaths. Follow-up was 100% complete. Four patients (12%) developed major complications. All were alive and well at the time of follow-up and there was 1 late arrhythmia (atrial fibrillation). In all but 1 patient, New York Heart Association functional class was improved or unchanged (1.47 +/- 0.51 vs 1.06 +/- 0.25, p = 0.0001). These results indicate that minimally invasive direct-access repair of ASD in adults is safe and effective, and is broadly applicable to the entire spectrum of defects. PMID- 10532512 TI - Comparison of outcomes research with clinical trials using preexisting data. AB - Outcomes research using analysis of preexisting data is a relatively new field with the potential to improve the quality and effectiveness of medical care, and may provide a useful complement to randomized studies. Motivated by the growth of this research in the cardiovascular literature, this review offers a framework to identify the core concepts of outcomes research from database analyses by comparing and contrasting it with the randomized clinical trial. PMID- 10532513 TI - Effect of prehospital thrombolysis on aborting acute myocardial infarction. AB - On administering thrombolysis in a prehospital setting, we found a threefold increase in the incidence of abortion of myocardial infarction, compared with the in-hospital program of a nearby hospital. Assessment of aborted myocardial infarction may be a better criterion for the efficacy of early thrombolysis than mortality data. PMID- 10532514 TI - Frequency of silent myocardial ischemia following coronary stenting. AB - To detect silent myocardial ischemia, 12-lead continuous electrocardiographic monitoring was performed in patients undergoing 1-vessel coronary stenting. Despite successful angiographic results, one third of the patients experienced silent myocardial ischemia during the postprocedural period. PMID- 10532515 TI - Prevalence of coronary artery disease, ischemic stroke, peripheral arterial disease, and coronary revascularization in older African-Americans, Asians, Hispanics, whites, men, and women. AB - The prevalence of coronary artery disease and of peripheral arterial disease was similar in older African-Americans, Asians, Hispanics, and whites, and the prevalence of ischemic stroke was lower in older whites than in older African Americans and Hispanics. The prevalence of coronary revascularization in older persons with coronary artery disease was lower in African-Americans than in whites and Hispanics and was lower in women than in men. PMID- 10532516 TI - Effects of fluvastatin on prothrombotic and fibrinolytic factors in type 2 diabetes mellitus. AB - The effects of fluvastatin therapy on parameters of coagulation and fibrinolysis were evaluated in patients with diabetic dyslipidemia in a randomized, placebo controlled study. Fluvastatin therapy was associated with a small reduction in factor VII coagulant activity, von Willebrand factor, and in plasminogen activator inhibitor 1 and tissue plasminogen activator antigens, but the effects of fluvastatin on hemostatic factors were much less marked than its effects on plasma lipids. PMID- 10532518 TI - Long-term hemodynamic responses to vasodilator therapy in patients with severe left ventricular dysfunction. AB - Maximal oral vasodilator therapy resulted in long-term reduction of initially elevated pulmonary vascular resistance in 10 of 13 patients with severe heart failure who tolerated inotrope-supported uptitration of afterload reduction. Eleven patients were unable to tolerate vasodilator therapy and required inotropic support for successful cardiac transplantation. PMID- 10532517 TI - Transcranial Doppler evaluation of microembolism immediately after direct-current cardioversion for atrial fibrillation in anticoagulated patients. AB - Transcranial Doppler monitoring showed no evidence of microemboli for 10 minutes after direct-current cardioversion for atrial fibrillation. PMID- 10532520 TI - Role of sinus wall compliance in aortic leaflet function. AB - Functional morphology of the normal aortic root was studied in vitro using 500 frames/s cinematography. Stiffening of the aortic wall by spraying its exterior with plastic adhesive led to compromised function of the leaflets, a phenomenon that may play a role in valve degeneration as it occurs in old-age fibrosis, atherosclerosis, or in connection with certain surgical procedures. PMID- 10532519 TI - Tailored therapy using dobutamine and nitroglycerin in advanced heart failure. AB - In 29 patients with advanced heart failure, therapy tailored to hemodynamic goals was attempted using an initial infusion of dobutamine and nitroglycerin (the latter in those with pulmonary hypertension) followed by escalating doses of oral vasodilators. In the 23 patients who were weaned from inodilator therapy, significant improvements in hemodynamic parameters and a low 90-day hospital readmission rate were documented. PMID- 10532521 TI - Usefulness of antimyosin antibody imaging for the detection of active rheumatic myocarditis. AB - Myocarditis constitutes an important component of rheumatic carditis. Antimyosin scintigraphy, which allows noninvasive assessment of myocyte damage, can be used for documentation of cardiac involvement in patients with rheumatic fever where clinical diagnosis is not unequivocal. PMID- 10532522 TI - Pneumatic external counterpulsation: a new noninvasive method to improve organ perfusion. AB - Pneumatic external counterpulsation, which is operated by applying electrocardiographic-triggered diastolic pressure via air-filled cuffs to the vascular limbs of lower limbs, is a relatively new therapeutic option for patients with angina pectoris and cerebrovascular diseases like transient ischemic attacks or sudden deafness. In this study, an augmentation in flow volume in the carotid, renal, and hepatic arteries from 20% to 25% and in the coronary arteries from 20% to 40%, as well as an increase in stroke volume by 12% was demonstrated; this shows the therapeutic results in patients with diseases caused by disturbed organ perfusion. PMID- 10532523 TI - Comparison of nitroglycerin lingual spray and sublingual tablet on time of onset and duration of brachial artery vasodilation in normal subjects. AB - This study compared the rapidity of onset, the magnitude, and the duration of action of 2 short-acting nitroglycerin preparations using high-resolution brachial ultrasound. Both sublingual tablet and lingual spray formulations caused maximal vasodilation at 3 minutes; however, the spray provided faster (at 2 minutes), greater, and more prolonged (15 minutes) vasodilation than the tablet. PMID- 10532524 TI - Weight loss of 146 kg with diet and reversal of severe congestive heart failure in a young, morbidly obese patient. AB - This case report describes a weight loss of 146 kg primarily due to diet in a very obese patient with reversal of severe congestive heart failure. PMID- 10532525 TI - T cell receptor repertoire usage in allotransplantation: an overview. AB - Lymphocytes express antigen receptors that allow the immune system to specifically recognize antigens. In transplantation, T cells play a critical role in the rejection process, and different protocols inhibiting T cell-mediated alloreactivity efficiently achieve prolongation of allograft survival. T cells can interact with alloantigens by two ways, either by the "indirect" pathway that correspond to the physiological mechanism of T cell immune recognition, or through the "direct" pathway where they recognize alloantigens directly on the surface of donor cells. If some T cells are specifically activated in allorecognition, one should be able to indirectly detect this "selection" by analyzing the T cell receptor usage that could be biased and reflect the preferential amplification of alloreactive lymphocyte subsets. Nevertheless compared with disease states such as cancer or autoimmunity the T cell receptor repertoire is still largely uncharacterized. We review the current results available on T cell repertoire usage in transplantation studies involving humans or various animal models. The T cell receptor repertoire involved in transplantation (restricted or unrestricted) and the features potentially common to alloimmune responses will be discussed. PMID- 10532526 TI - Viewing "donor potential" with realism. PMID- 10532527 TI - Obliterative bronchiolitis: the Achilles heel of lung transplantation. PMID- 10532528 TI - Prolonged cold ischemia, late graft dysfunction, and cyclosporine in renal transplantation. PMID- 10532529 TI - Improvements in diabetic microangiopathy after successful simultaneous pancreas kidney transplantation: a computer-assisted intravital microscopy study on the conjunctival microcirculation. AB - A computer-assisted intravital microscopy technology has been developed to noninvasively and objectively study diabetic microangiopathy in the conjunctival microcirculation of type-1 diabetics. Quantitative characterization of the conjunctival microcirculation was performed on 12 patients pre- and 18 months postsimultaneous pancreas-kidney transplantation (SPK). Healthy nondiabetic volunteers (n=12), solitary kidney (K) transplanted type-1 diabetics (n=5), and nontransplanted type-1 diabetics (n=12) served as controls. Pre-SPK diabetics showed abnormal-sized venules (diameter=66+/-7 microm) and reduced presence of arterioles (arteriole length/area=18+/-6 microm(-1)) compared with nondiabetic controls (53+/-4 microm; 31+/-8 microm(-1); P<0.05). The computed vascular perfusion capacity of the conjunctival microvasculature was diminished in the same patients (pre-SPK diabetics=49+/-9%; nondiabetic healthy controls=71+/-6%; P<0.05). Significant improvement in microangiopathy was observed in all post-SPK diabetics (diameter=58+/-6 microm; arteriole length/area=26+/-9 microm(-1); vascular perfusion=63+/-8%; P<0.05) 18 months post-SPK. Blood flow velocities in the conjunctival microcirculation in the same post-SPK patients showed noticeable but not significant improvements (nondiabetic controls=2.94+/-0.57 mm/sec; pre SPK=1.23+/-0.49 mm/sec; post-SPK=1.65+/-0.42 mm/sec). The solitary kidney transplant controls (post-K) showed no significant improvements in diabetic microangiopathy, confirming the unique role of the pancreas in SPK. In general, significant improvements (P<0.05) in diabetic microangiopathy were observed in all 12 diabetics 18 months post-SPK but not in the controls. PMID- 10532531 TI - Membrane stabilizing effects of calcium and taxol during the cold storage of isolated rat hepatocytes. AB - BACKGROUND: Calcium plays an important role in liver preservation and preservation induces depletion of cellular Ca. This may affect hepatocyte cytoskeleton integrity necessary for maintaining cell shape and organ viability. We tested the effects of a microtubular stabilizer (Taxol) in liver cell preservation. METHODS: Isolated rat hepatocytes were preincubated with or without a microtubule stabilizing agent, 100 microM Taxol, at 37 degrees C for 20 min, then stored in the University of Wisconsin (UW) solution +/-1.5 mM CaC12 at 4 degrees C for up to 48 hr. After storage, the cells were rewarmed in Krebs Henseleit buffer with air at 37 degrees C for 1 hr. Morphological changes in the plasma membrane (scanning electron microscopy) and cell viability (percentage of lactate dehydrogenase [LDH] release) before and after rewarming were studied. RESULTS: Hepatocytes showed time-dependent increase in bleb formation (cytoskeleton disruption) during cold storage. Rewarming the cells caused even greater bleb formation and increased LDH release (cell death). Pretreatment of cells with Taxol and cold storage in the UW solution with 1.5 mM Ca suppressed both bleb formation and LDH release in 48-hr coldstored cells. CONCLUSIONS: Cold storage of hepatocytes leads to reperfusion injury and cell death. This can be suppressed with Taxol and Ca. This suggests that hypothermia induces changes in cellular Ca and a disruption of the microtubules, leading to loss of cell viability. Improved liver preservation may require suppression of Ca-dependent disruption of the cytoskeleton system of liver cells. PMID- 10532530 TI - Analysis of Fas system in pulmonary injury of graft-versus-host disease after rat intestinal transplantation. AB - The lung is one of the primary targets of acute graft-versus-host disease (GVHD), which is the principal complication that occurs after allogeneic intestinal transplantation. The purpose of this study is to investigate the involvement of Fas/Fas ligand system in pulmonary injury after rat semi-allogeneic intestinal transplantation. The lungs were serially harvested from LEW x BN F1(LBNF1) recipients of either LEW heterotopic intestinal allografts or LBNF1 isografts, on days 1, 3, 5, 9, and 13 posttransplant. In light microscopy, pulmonary injury became apparent on day 13 in the allogeneic combination, showing a thickening of the alveolar septa. The incidence of apoptosis, examined by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate (dUTP) biotin nick end-labeling, was observed to increase steadily in the alveolar cells accompanied by a progression of GVHD. In an immunohistochemical study, Fas was constitutively expressed in the lung, although Fas ligand was expressed most extensively on day 9. The immunoreactivity of both Fas and Fas ligand were observed in alveolar cells, in addition to leukocytes. An analysis by reverse transcription polymerase chain reaction also revealed that the expression of Fas mRNA was constitutive without any significant change, although that of Fas ligand mRNA increased substantially and peaked on day 9, which was significant compared to the isogeneic combination. In conclusion, transcriptionally up-regulated Fas ligand and increased number of apoptosis suggests that the Fas system may play a role in the pathophysiology of GVHD-induced pulmonary injury. PMID- 10532532 TI - Role of tumor necrosis factor receptors TNFR-I (P55) and TNFR-II (P75) in corneal transplantation. AB - BACKGROUND: To determine the role of tumor necrosis factor-alpha (TNF-alpha) receptor (TNFR) function in corneal allograft immunology. METHODS: Animals with gene-targeted deficiency in TNFR-I (p55-/-), TNFR-II (p75-/-), or combined TNFR I/TNFR-II deficiency (p55-/-p75-/-) and their wild-type controls were used as recipients of fully-mismatched (BALB/c; n=88) or multiple minor alloantigen mismatched (BALB.b; n=62) orthotopic corneal transplants to determine the effect of selective deficiency in one or both TNF-alpha receptors on corneal allograft survival. Grafted recipients were followed biomicroscopically for signs of rejection, and survival data were analyzed by the Kaplan-Meier method. RESULTS: There was no discernible difference in survival of fully-mismatched BALB/c corneal grafts in p55-/- (n=12; P=0.76) or in double-knockout p55-/-p75-/- (n=13; P=0.41) as compared with wild-type C57BL/6.129 hosts. However, the survival of BALB/c allografts was lower in p75-/- (n=10; median survival 20 days) as compared with control C57BL/6 (n=30; median survival 30 days) hosts (P=0.02). In contrast, there was no discernible effect in survival of minor alloantigen-disparate BALB.b corneal grafts in p75-/- (n=13; P=0.95) or in combined p55-/-p75-/-(n=10; P=0.17) hosts as compared with C57BL/6 (n=9) and C57BL/6.129 (n=10) wild-type controls, respectively. However, there was a profound enhancement in the survival of BALB.b allografts in p55-/- recipients (n= 10; median survival 35 days) as compared to wild-type C57BL/6.129 (n=10; median survival 25 days) controls (P<0.01). CONCLUSIONS: Our data suggest that the two TNF-alpha receptors largely play discrete roles in mediating rejection of murine corneal allografts. TNFR-I (p55) function seems to be integral to the rejection of minor-disparate grafts, and its selective suppression leads to enhancement of allograft survival. In contrast, TNFR-II (p75) function appears to be associated with enhanced survival of major histocompatibility complex-disparate allografts. The combined deletion of TNFR functionality in p55-/-p75-/- confers no net advantage or disadvantage to major histocompatibility complex or minor alloantigen-disparate grafts. PMID- 10532533 TI - Participation of the liver in generation of a vigorous anti-donor response after inoculation of donor spleen cells. AB - BACKGROUND: There is a general agreement that a preferential accumulation of alloantigens within the liver could induce hyporesponsiveness to the inoculated antigens. Entrapment of antigens in the liver may evoke an unique immune response in the organ and play a key role in determination of the fate of the transplanted grafts. To understand the immune response in the liver after inoculation of allogeneic donor antigens, we examined the immune response to systemically inoculated alloantigen in rats whose sensitized liver was replaced with that of naive rats or in naive rats whose liver was replaced with that of sensitized rats. METHODS: Using implantation of syngeneic liver (alloantigen accumulated/naive) in rats (naive/alloantigen-sensitized), we compared the immune responses to alloantigen between rats with hepatic/extrahepatic alloantigen at 24 hr after alloantigen inoculation. This was called sensitized-liver-grafted (SLG)/sensitized-liver-removed (SLR) treatment. The immune response to donor alloantigen in this model was evaluated by survival of skin or heart grafts, complement-dependent cytotoxicity (CDC) titer and delayed-type hypersensitivity (DTH) response. RESULTS: Compared with the mean survival time (MST) in donor spleen cell inoculated (DSI) rats (skin and heart, MST: 8.2+/-1.1 and 10.7+/-2.3 days), SLG rats rejected allografts in an accelerated fashion (skin and heart, MST: 5.5+/-0.5 and 4.2+/-0.8 days), associated with higher CDC titer and DTH response. In contrast, allograft survival was moderately prolonged in SLR (skin and heart, MST: 16.5+/-2.6 and 29.5+/-3.7 days) associated with suppressed CDC titer and DTH response. The survival of third-party allograft after SLG or SLR treatment (skin, MST: 9.3+/-1.5 or 9.7+/-0.6 days) indicated that immunological hyper/hyporesponsiveness was donor-specific. CONCLUSIONS: A strong anti-donor immune response was induced by the transfer of donor antigen-baring liver to naive rats 24 hr after alloantigen inoculation, whereas removal of the liver suppressed alloimmune response. Our results indicate that vigorous anti alloimmune response occurred in the liver after systemic inoculation of donor spleen cells. PMID- 10532534 TI - The portosystemic shunt protects liver against ischemic reperfusion injury. AB - BACKGROUND: The goal of this study was to characterize the importance of splanchnic viscera in liver ischemic reperfusion injury and to enhance the tolerance of liver to warm ischemia injury with portosystemic shunt. METHODS: The hepatic blood flow of male Sprague Dawley rats was subjected to 45, 60, 120, and 150 min liver warm ischemia with or without portosystemic shunt (splenic-caval shunt). The production of tumor necrosis factor a (TNFa), nuclear factor-kappaB activation, inducible NO synthase (iNOS) expression, and apoptosis were examined. RESULTS: A total of 67% of rats with 45 min liver warm ischemia (n=6) and 100% of rats with 60 min liver warm ischemia (n=6) died within 1 day. However, all rats with 120 min (n=8) liver warm ischemia in splenic-caval shunt group survived for over 1 day, 6/8 for over 3 days, and 5/8 for over 5 days without significant histological changes of the liver. Serum tumor necrosis factor levels in liver warm ischemic rats were increased, This increase was significantly reversed after portosystemic shunt. After challenge with lipopolysaccharide (1 mg/kg, p.v.), naive rats survived for over 5 days (n=4) with the peak value of rat tumor necrosis factor (240 pg/ml) at 90 min. In contrast, all rats died within one day (n=5) with the peak value of rat tumor necrosis factor a (465 pg/ml) at 45 min after administration of lipopolysaccharide in the rats with liver warm ischemia plus splenic-caval shunt. iNOS expression and nuclear factor-kappaB activation were very strongly increased in the hepatocytes after liver warm ischemia with portosystemic shunt, compared with liver ischemia without portosytemic shunt. CONCLUSIONS: We conclude that the splanchnic viscera can contribute to liver ischemic reperfusion injury. Portosystemic shunt enhances the tolerance of liver to warm ischemia through the protective role of iNOS and nuclear factor-kappaB (NF-kappaB). PMID- 10532535 TI - Detection of minor alloantigen-specific cytotoxic T cells after rejection of murine orthotopic corneal allografts: evidence that graft antigens are recognized exclusively via the "indirect pathway". AB - BACKGROUND: The immune mechanisms by which corneal allografts are rejected in normal ocular graft beds have not been identified. Both acceptors and rejectors of these types of grafts display donor-specific delayed hypersensitivity and in vitro proliferating primed T cells, yet neither develop conventional, donor specific cytotoxic T cells. We wished to determine whether unconventional donor specific cytotoxic T cells are generated in rejector mice that recognize donor minor alloantigens presented by recipient major histocompatibility complex (MHC) molecules. METHODS: BALB/c mice received orthotopic corneal allografts from C57BL/10 donors in normal eyes. At 4 weeks (when 50% of grafts can be designated as rejected), primed cytotoxic T lymphocyte (CTL) activity in draining lymph nodes and spleen was assayed on targets selected to present donor-type minor H antigens on recipient MHC molecules. Control mice received heterotopic corneal allografts and were similarly examined. RESULTS: Lymphoid organs of recipients that rejected orthotopic or heterotopic corneal allografts contained CTL that lysed targets expressing donor-type minor H antigens presented by recipient MHC molecules. By contrast, no CTL activity was detected from lymphoid cells of recipients that accepted orthotopic corneal allografts. Rejection of orthotopic corneal allografts placed into normal mouse eyes correlates directly with the generation of donor-specific CTL that recognize minor H antigens in the context of recipients MHC molecules. CONCLUSIONS: These results indicate the indirect pathway of alloantigen presentation is the only pathway operative in the process by which orthotopic corneal allografts are rejected. The roles of emigrant Langerhans cells and corneal lymphatics in the indirect pathway are discussed. PMID- 10532536 TI - Alloimmune injury preceding airway obliteration in porcine heterotopic lung implants: a histologic and immunohistologic study. AB - BACKGROUND: Obliterative bronchiolitis (OB), the major long-term complication of lung transplantation, has thus far lacked a good large-animal model. Our goal was to develop such a model on the basis of previous rodent models with tracheal implants. METHODS: Fragments of pulmonary tissue with structures of terminal bronchi were subcutaneously transplanted to four random-bred domestic piglets. Each animal received 10 autograft and 10 allograft implants. The histologic findings were graded from 0 to 3 for implants harvested repeatedly over 2 months. RESULTS: In autografts, partial destruction of the respiratory epithelium and gradual luminal obliteration as well as mild damage to the cartilage and the bronchial wall underwent rapid reversal after initial ischemic injury. In the allografts, epithelial destruction and gradual obliteration were total within 14 days, the difference being statistically significant (P<0.05) in both. The histologic features of the obliterative plug were similar to those of human OB. In the allografts, cartilaginous destruction and pericartilaginous inflammation increased gradually to severe levels, significantly worse than in the autografts (P<0.05). Necrosis and inflammation of the bronchial wall were also more severe in the allografts (P<0.05). CONCLUSIONS: At the end of follow-up, all autografts were vital, whereas the allografts were almost totally rejected and were without native structures. All bronchi in the allografts exhibited accelerated obliteration with histologic features characteristic of human OB, thus providing a model for research into OB and its prevention. PMID- 10532537 TI - The impact of azathioprine on chronic viral hepatitis in renal transplantation: a long-term, single-center, prospective study on azathioprine withdrawal. AB - BACKGROUND: In transplanted patients, viral hepatitis progresses to chronic liver disease and patient's death after many years of transplantation. Also, it is well known that azathioprine (AZA) is harmful to the liver of these patients. However, it is unclear whether a low dose of AZA still represents a threat to the viral liver disease. METHODS: A total of 79 patients with hepatitis C, B, or both, transplanted between 1973 and 1990, were grouped according to whether they had AZA either withdrawn from the immunosuppressive regimen [group (G) I, n=45] or a dosage reduction only (group II, n=34). The decision to remove or to keep AZA was restricted to the patient's doctor. Patients records were reviewed by April 1997. RESULTS: After an equal time of follow-up, after the AZA changing (64+/-26 vs. 58+/-29 months), patients in GI showed a significant decrease in the serum liver parameters when compared to baseline [alanine aminotransferase (ALT): P=0.001; gamma-glutamyl transferase (gamma-GT): P=0.001 and total bilirubin: P=0.002], whereas in GII only ALT decreased (P=0.04) although gamma-GT and total bilirubin did not. Compared to baseline, serum creatinine (SCr) increased only in GI (P=0.001) but, at last follow-up, did not differ from GII. The intention-to perform liver biopsies was equal in GI and GII (16 vs. 14) but the hystological findings of severe chronic liver disease (either chronic active hepatitis or cirrhosis) were more frequent in GII (P=0.004). Death with a functioning graft was much more frequent in GII than in GI (P=0.001). Infection and cirrhosis were more common as a cause of death in GII than in GI. CONCLUSIONS: The use AZA is harmful to renal transplantation patients with both chronic hepatitis C and B and, therefore, should be avoided. AZA withdrawal, but not dose adjustments, diminishes the serum liver enzymes and the progression rate of the chronic viral liver disease as well as the rate of death secondary to infection and cirrhosis. PMID- 10532538 TI - Respiratory viruses in adult liver transplant recipients. AB - BACKGROUND: The contribution of respiratory viruses to respiratory disease in adult liver transplant (LT) recipients has not been studied. We performed a prospective audit to document the incidence of respiratory syncytial viruses ([RSVs], parainfluenza virus, influenza virus, and adenovirus) after LT, and to determine their contribution to respiratory disease in this setting. METHODS: Consecutive adult recipients were followed for 8 months after LT. Throat swabs were collected weekly for up to 12 weeks after LT, and virological surveillance was performed using conventional techniques (direct immunofluorescence and cell culture). A polymerase chain reaction assay for RSV was subsequently performed on selected specimens. Clinical data, including episodes of respiratory disease, were also recorded. RESULTS: During the study period, 51 patients received 53 LT. Five patients died, but no viruses were isolated from these patients at any stage. A total of 323 swabs were examined by conventional techniques and yielded 35 viral isolates (10.8%). Herpes simplex virus (type 1) accounted for 33 isolates, none of which were associated with respiratory disease. Two of 323 swabs (0.62%), in 2 patients, yielded respiratory viruses (both RSV); both patients had self-limiting, mild, upper respiratory tract symptoms. In these 2 patients, the polymerase chain reaction assay was more sensitive than conventional techniques and was able to detect extended RSV excretion. Of 51 recipients, 31 (61%) were always negative for viruses. Of 51 recipients, 10 developed respiratory failure, but no respiratory viruses were isolated from any of these patients. CONCLUSIONS: Respiratory viruses are rarely isolated from adult recipients after LT and are not associated with serious morbidity or with mortality. Routine surveillance for respiratory viruses in this patient population is not justified on the basis of this study. PMID- 10532540 TI - Clinical utility of a quantitative polymerase chain reaction for diagnosis of cytomegalovirus disease in solid organ transplant patients. AB - BACKGROUND: Accurate and rapid diagnosis of human cytomegalovirus (HCMV) disease in solid organ transplant patients remains a challenge. We evaluated the clinical utility of a quantitative polymerase chain reaction (QPCR) method to diagnose transplant patients with HCMV disease. METHODS: A total of 429 plasma samples from 121 solid organ transplant patients were prospectively collected and evaluated for HCMV using a QPCR assay. To enhance the sensitivity of the QPCR assay, plasma samples were centrifuged in a manner designed to concentrate the virions before nucleic acid extraction. Quantitation was achieved by co amplifying an internal quantitative standard (IS) that contained the same primer sequences as for HCMV. Polymerase chain reaction products were detected by hybridization to 96-well microtiter plates coated with either a HCMV- or an IS specific probe. RESULTS: A total of 103 patients had all samples negative by QPCR. None of the 103 patients developed HCMV disease during the study. In contrast, 18 patients showed at least 1 sample positive by the QPCR assay, but only 8 of these developed HCMV disease. The mean viral load value for patients without HCMV disease was 93 viral copies (vc) per ml of plasma (range: 35-325 vc/ml plasma) and for the 8 patients with HCMV disease was 67,686 vc/ml plasma (range: 167-1,325,000 vc/ml plasma) (P<0.001). Using a cut-off value of 100 vc/ml plasma and clinical diagnosis of HCMV disease, the QPCR assay showed a sensitivity of 100% and specificity of 99.1%. CONCLUSION: HCMV viral load may be useful in the diagnosis of HCMV disease in solid organ transplant patients. PMID- 10532539 TI - A European perspective on organ procurement: breaking down the barriers to organ donation. AB - A worldwide shortage of donor organs has led to the development of national and international systems for organ procurement and allocation. Such systems promote organ donation and ensure fair distribution of available donor organs through a combination of legislation, organ exchange organizations (OEOs), transplant coordinators, publicity campaigns, donor cards, and professional training programs. The development of national and international OEOs is central to this process because they maintain waiting lists and allocate organs in the most appropriate way. Most countries also employ transplant coordinators whose role involves promoting links between transplant centers and intensive care units, establishing protocols for organ donation, and helping hospital staff deal with the sensitive issues involved in organ donation. Educational initiatives, such as the European Donor Hospital Education Programme developed by Eurotransplant is now used in over 30 countries worldwide. The program aims to improve professionals' understanding of the legal and ethical issues involved in transplantation, to help them communicate effectively and sympathetically with bereaved families, and to increase organ donation rates. Other initiatives include programs such as the Donor Action Programme, which was set up by professional organizations in the US and Europe aiming to help hospitals establish tailor-made organ procurement policies to ensure that all potential donors can be identified and reported and the needs of unfortunate families can be met in a caring and sensitive manner. PMID- 10532541 TI - Epstein-Barr virus-related disorders in children undergoing renal transplantation with tacrolimus-based immunosuppression. AB - In children undergoing renal transplantation, Epstein-Barr virus- (EBV) related disorders, including posttransplant lymphoproliferative disorder, constitute a major complication associated with tacrolimus-based immunosuppression. In this study, we reviewed the EBV complications in 81 children, all of whom had EBV serological studies before renal transplantation. We also highlight the data in a subgroup of 30 children transplanted more recently who were monitored sequentially for EBV symptoms and signs and with immunological studies, and in whom the donor EBV serology was also determined. During a mean follow-up time of 3.9+/-2.3 years, 19 children developed symptomatic Epstein-Barr virus (EBV*) infection. This consisted of the clinical syndrome of infectious mononucleosis in 7 children; in addition, 10 children developed posttransplant lymphoproliferative disorder (PTLD), which was histologically confirmed in 8, and 2 others developed malignant lymphoma. Recipient seronegativity (EBV-) and donor EBV seropositivity (EBV+) predicted a high probability for seroconversion (P=0.0072) and for developing PTLD or malignancy (P<0.01). In the subgroup of 30 children studied prospectively, seroconversion occurred in 15 of 19 seronegative recipients of EBV seropositive grafts at 6.6+/-2.6 months (mean+/-SD) after transplantation. Seven children developed symptomatic EBV infection (including three with PTLD) in association with seroconversion and a rise in EBV viral load in the peripheral blood, demonstrated by an EBV-specific polymerase chain reaction (EBV-PCR). Of 15 seroconverters, 7 who developed symptomatic infection had received EBV+ grafts; 8 others with EBV+ grafts seroconverted but did not become symptomatic. These two subgroups did not differ in age, rejection rate, antiviral prophylaxis, or level of immunosuppression. In the overall group of 81 children, only the two with malignant lymphoma who were managed with chemotherapy had substantial morbidity. The 10 individuals with PTLD received a regimen combining i.v. ganciclovir and CytoGam, and stopping or reducing the tacrolimus. Four children with associated marked tonsilar growth underwent tonsillectomy. All 19 individuals with EBV disorders resolved their symptoms and signs, and all have maintained good allograft function during a follow-up time of 3.0+/-2.5 years (mean+/-SD) after the development of symptomatic EBV infection, PTLD, or malignancy. We conclude that seronegative recipients of EBV+ grafts are at high risk for developing EBV related disorders after renal transplantation under tacrolimus-based immunosuppression, although the ultimate clinical outcomes have been remarkably good. These data form the basis for formulating strategies for early identification of children at risk for EBV complications, and for instituting preventive and treatment strategies that permit these children to realize the substantial benefits offered by tacrolimus-based immunosuppression. PMID- 10532542 TI - Late graft dysfunction after prolonged cold ischemia of the donor kidney: inhibition by cyclosporine. AB - BACKGROUND: The present study was devised to elucidate the influence of prolonged cold ischemia on the development of chronic transplant dysfunction (CTD) in kidney isografts (Brown Norway-->Brown Norway; BN-->BN) and in kidney allografts (BN-->Wistar Agouti/ Rij [WAG]) under temporary cyclosporine (CsA) therapy. METHODS: To induce ischemic injury, BN donor kidneys were preserved for 24 hr in 4 degrees C University of Wisconsin solution before transplantation. Renal function (proteinuria), histomorphology according to the BANFF criteria for CTD, and infiltrating cells were assessed. Grafts were examined both early at days 2, 3, 6, and 10, and late at week 26 (allografts) or at week 52 (isografts). RESULTS: Nonischemic isografts preserved a normal function and morphology. Ischemic isografts developed a progressive proteinuria over time and demonstrated significantly more glomerulopathy with macrophage (Me) infiltration and intimal hyperplasia than nonischemic controls at week 52. During the initial 10 days, there was an increased infiltration of MHC class II+ cells, predominantly CD4+ cells and Mphi, coinciding with up-regulated intercellular adhesion molecule-1 expression. CsA treatment in ischemic isografts inhibited infiltration of MHC II+ cells in the early stage, which was accompanied by significantly less renal damage at week 52 compared with untreated controls (proteinuria: 59+/-8 vs. 134+/ 19 mg/24 hr; BANFF score: 2.8+/-0.4 vs. 4.3+/-1.0). Under CsA therapy, 24-hr cold ischemia of the allograft affected neither the onset or progress of proteinuria, nor the histomorphology (BANFF score: 7.8+/-2.4 vs. 7.3+/-1.9). In both ischemic and nonischemic allografts, intercellular adhesion molecule-1 expression and mononuclear cell infiltration (CD4, CD8, Mphi was abundantly present during the first 10 days and function deteriorated rapidly. CONCLUSIONS: Prolonged cold ischemia plays a role in the induction of CTD, but its deleterious effect can be successfully inhibited by CsA. Therefore, the alloantigeneic stimulus is the overriding component in the multifactorial pathogenesis of CTD. PMID- 10532543 TI - Prevention of renal allograft rejection in primates by blocking the B7/CD28 pathway. AB - BACKGROUND: There is accumulating evidence that blockade of the costimulatory pathways offers a valid approach for immune suppression after solid organ transplantation. In this study, the efficacy of anti-CD80 and anti-CD86 monoclonal antibodies (mAbs) in combination with cyclosporine (CsA) to prevent renal allograft rejection was tested in non-human primates. METHODS: Rhesus monkeys were transplanted with a partly major histocompatibility complex-matched kidney on day 0. Anti-CD80 and anti-CD86 mAbs were administered intravenously daily for 14 days starting at day - 1. CsA was given intramuscularly for 35 days starting just after transplantation. The kidney function was monitored by determining serum creatinine levels. RESULTS: The combination of anti-CD80 and anti-CD86 mAbs completely abrogated the mixed lymphocyte reaction. Untreated rhesus monkeys rejected the kidney allograft in 5-7 days. Treatment with anti CD80 plus anti-CD86 mAbs resulted in a significantly prolonged graft survival of 28+ 7 days (P=0.025). There were no clinical signs of side effects or rejection during treatment. Kidney graft rejection started after the antibody therapy was stopped. The anti-mouse antibody response was delayed from day 10 to 30 after the first injection. No difference in graft survival was observed between animals treated with CsA alone or in combination with anti-CD80 and anti-CD86 mAbs. However, treatment with anti-CD80 and anti-CD86 mAbs reduced development of vascular rejection. CONCLUSIONS: In combination, anti-CD80 and antiCD86 mAbs abrogate T-cell proliferation in vitro, delay the anti-mouse antibody response in vivo, and prevent graft rejection and development of graft vascular disease in a preclinical vascularized transplant model in non-human primates. PMID- 10532544 TI - FK506 markedly enhances apoptosis of antigen-stimulated peripheral T cells by down-regulation of Bcl-xL. AB - BACKGROUND: FK506 is a clinically effective immunosuppressive agent and promoter of immunologic tolerance. However, limited information is available about the mechanism of FK506-induced immunosuppression. METHODS: In the present study, we investigated the molecular mechanism of FK506-mediated enhancement of apoptosis using in vivo activated T lymphocytes. We examined the effects of FK506 on apoptosis-related proteins in superantigen-stimulated peripheral T cells. RESULTS: Injection of staphylococcal enterotoxin B (SEB) into BALB/c mice resulted in a selective apoptosis of splenic Vbeta8-positive T cells after 48 hr. Injection of FK506 within 36 hr of SEB injection resulted in a marked enhancement of DNA fragmentation of splenic Vbeta8+ T cells. FK506 did not affect the expression of Fas antigen on SEB-activated Vbeta8+ T cells. As Bcl-2-related proteins are involved in apoptotic process, we also evaluated their role by examining the expression of Bcl-2, Bcl-X(L), and Bax on SEB-FK506-treated murine splenic T cells. Although SEB injection slightly increased the expressions of Bcl 2 and Bax on V138+ T cells, FK506 did not modulate Bcl-2 or Bax expression in these cells. In contrast, the expression of Bcl-x(L) on Vgamma8+ T cells, which was markedly induced by SEB, was abrogated by FK506. CONCLUSIONS: Our findings indicate FK506-induced enhancement of apoptosis of activated T cells is mediated by down-regulation of Bcl-X(L) expression on these cells. Our results also suggest that Bcl-x(L) is a critical determinant of apoptosis of activated T cell and may represent a potential target for new therapies designed to achieve immunological tolerance. PMID- 10532545 TI - Purified donor T cells alone activate transplantation immunity to the male antigen but induce tolerance in combination with Mac-1+ donor cells. AB - BACKGROUND: In most experimental systems examined, "professional" antigen presenting cells (APCs), such as dendritic cells, have been found to activate T cells, whereas "nonprofessional" antigen-bearing cells (nonAPC) may induce tolerance. Some recent studies have suggested that nonAPCs may under certain conditions prime a T-cell immune response. We have attempted to separate the roles of transplanted T cells and monocytic/dendritic cells in activating or tolerizing antigen-specific T cells in vivo, by examining the consequences of parenteral exposure to male antigen in anti-male TCR transgenic female mice. METHODS: Qualitative and quantitative changes in the large population of male reactive transgenic T cells to various male donor cell populations in transgenic female mice were followed after injections of highly purified male lymphoid cells. Changes in male-reactive T cells with time and the long-term outcome of male skin grafts were measured. RESULTS: When a nonAPC population consisting of highly purified male T cells alone was injected intravenously into H-Y antigen specific TCR transgenic female mice, the number of host transgenic T cells was sustainably increased, and male graft rejection was accelerated. Injection of a combination of purified T cells and purified Mac-l+ cells induced massive and permanent deletion of the host male-reactive T-cell population and permanent graft tolerance. Mac-l+ cells alone gave no appreciable change in responsive T cells or graft rejection times. CONCLUSIONS: The data indicate that highly purified T cells engrafted alone induce rapid sensitization toward the male antigen. They also show that both male donor T cells and a population of male monocytic/ dendritic cells are required to induce peripheral tolerance toward this antigen and that this tolerance is related to permanent peripheral deletion of male-reactive T cells. PMID- 10532546 TI - Characterization and manipulation of T cell immunity to skin grafts expressing a transgenic minor antigen. AB - BACKGROUND: Minor histocompatibility antigens play a significant role in allograft rejection when donor and recipient are matched at MHC loci. An improved understanding of T cell immunity directed toward a model minor antigen may provide new approaches for preventing graft rejection. METHODS: C57BL/6 (B6) recipient mice were engrafted with skin from B6 beta-galactosidase transgenic (beta-gal tg) donors and the induced T cell immune responses were characterized by cytokine ELISA spot assay. beta-gal-specific immunity was manipulated prior to transplant through preinjection with beta-gal in complete Freund's adjuvant (CFA) or through preinjection with soluble beta-gal i.v. RESULTS: B6 mice rejected beta gal tg skin by day 25. Rejection was associated with a low frequency of predominantly CD8+, interferon-gamma-producing T cells capable of directly recognizing both beta-gal tg cells and an immunodominant major histocompatibility complex I-restricted peptide derived from the beta-gal protein. Rejection of multiple minor antigen disparate skin and major histocompatibility complex disparate skin occurred significantly faster, and was associated with a 10- to 30 fold higher frequency of alloreactive T cells, than rejection of beta-gal tg skin. Prepriming of recipients with beta-gal in complete Freund's adjuvant resulted in an increased frequency of beta-gal-specific T cells and accelerated rejection of beta-gal tg skin. Intravenous injection of soluble beta-gal-induced graft tolerance and a lack of detectable beta-gal-specific immunity. CONCLUSIONS: The findings reveal that transgenically expressed beta-gal behaves as a minor transplantation antigen and that manipulation of the beta-gal-specific T cell repertoire can dramatically affect rejection of beta-gal tg skin grafts. The work provides the foundation for mechanistic studies of tolerogenesis to minor antigenic determinants. PMID- 10532547 TI - Comparison of chimeric acid and non-chimeric tolerance using posttransplant total lymphoid irradiation: cytokine expression and chronic rejection. AB - BACKGROUND: Previous studies showed that an intravenous infusion of donor blood cells facilitates tolerance to ACI heart allografts in Lewis rat hosts given posttransplant total lymphoid irradiation (TLI) and anti-thymocyte globulin (ATG). The object of the current study was to compare tolerance induction using donor cells that do or do not induce chimerism. METHODS: Normal peripheral blood mononuclear cells (PBMC), granulocyte colony-stimulating factor (G-CSF)-mobilized PBMC, and bone marrow (BM) cells from ACI donors were tested for their capacity to prolong ACI heart allograft survival in Lewis hosts. Chimerism, anti-donor cell reactivity, and cytokine gene expression in grafts were determined. RESULTS: Intravenous injections of equal numbers of all three donor cells markedly prolonged graft survival (median: >164 to >175 days) as compared to uninjected controls (median: 53 days). Chimerism among T and B cells in the blood was determined by immunofluorescent staining in hosts bearing long-term (> 150 days) grafts. Although no chimerism was detected in hosts given normal or G-CSF mobilized PBMC, chimerism was detected at variable levels in all hosts given BM cells. Vigorous anti-donor reactivity in the mixed leukocyte reaction was present only in non-chimeric hosts. Long-term grafts from hosts given normal ACI PBMC developed chronic rejection, but those from hosts given ACI BM cells did not. The latter hosts showed the lowest levels of intragraft cytokine mRNA. CONCLUSIONS: Chimeric tolerance is more robust than non-chimeric tolerance in the model of posttransplant TLI, ATG, and donor cell infusion, and is associated with less chronic rejection. PMID- 10532549 TI - Gemcitabine--a novel immunosuppressive agent--prevents rejection in a rat cardiac transplantation model. AB - BACKGROUND: 2',2' -difluorodeoxycytidine (dFdC, gemcitabine) is a pyrimidine antimetabolite with antineoplastic activity against a wide range of solid tumors. The immunosuppressive activities of this compound have not been described to date. METHODS: The in vitro effects on activated T lymphocytes were studied with a lymphocyte colony-forming assay in a microagar culture system. Heart transplantations were performed in the fully allogeneic Lewis/ Brown Norway combination. dFdC was administered once daily at various dosages from the time of surgery until day 50. RESULTS: Phytohemagglutinin-induced lymphocyte proliferation was inhibited 50% by dFdC at a concentration of 3.25+/-0.9 nmol/L. Allografts of untreated animals survived for 7.5 (7-8) days and those with 25, 50, and 75 microg/kg body weight dFdC for 7.3 (7-8), 9.3 (8-10), and 16.3 (10-38) days, respectively. Treatment with 100 or 125 microg/kg body weight of dFdC, however, prolonged allograft survival until day 152.8 (129-178). Dose-dependent leukopenia was the main toxicity. CONCLUSIONS: DFdC is a new immunosuppressive agent that can successfully prevent cardiac rejection in a rat transplantation model. PMID- 10532548 TI - The probability of HLA-C matching between patient and unrelated donor at the molecular level: estimations based on the linkage disequilibrium between DNA typed HLA-B and HLA-C alleles. AB - BACKGROUND: Recent evidence suggests a more significant role of HLA-C as a target of alloreactions after bone marrow transplantation than previously suspected. Although linkage disequilibrium (LD) between HLA-B and -C serogroups is well documented, the level of LD at the allelic level is not known. In this study, we determine the LD between HLA-B and -C alleles and estimate the probability of molecular HLA-C matching between unrelated individuals who match for both HLA-B alleles. METHODS: The study included 727 haplotypes from 849 individuals who were HLA-A, -B, -C and -DRB1 typed by high-resolution PCR-SSOP technique. Zelterman's statistic was used to test for global LD between HLA loci. LD between specific HLA-B and -C allelic combinations was calculated from their observed and expected frequencies in the study haplotypes. The probability of HLA-C matching for specific HLA-B allele was estimated from contingency table generated from the HLA B and -C haplotypes. RESULTS: HLA-C was found to exist in LD with HLA-A and -B, as well as -DRB1, loci; however, it was strongest between HLA-B and -C loci. A marked variability in the level of LD between specific HLA-B and -C alleles was noticed. A strong LD was seen in some allele pairs like B*0702-C*w0702, B*3501 Cw*0401, and B*0801-Cw*0701. The overall estimated probability of HLA-C matching between unrelated individuals that match for both HLA-B alleles is 42.25%. For 237 (72.9%) of 325 combinations involving the 25 commonest HLA-B alleles, the estimated probability that the HLA-B-matched unrelated individuals will match for both HLA-C alleles is less than 50%. In addition, a 100% probability of matching for both HLA-C alleles is expected only if both individuals bear either B*0801/ B*0801 or B*4901/B*4901 or B*0801/B*4901. Probability tables for common alleles are presented. CONCLUSIONS: We conclude that, despite matching for both HLA-B alleles by high resolution DNA typing and the presence of a strong LD between HLA B and HLA-C loci, unrelated individuals are more likely to mismatch rather than match for one or both HLA-C alleles. PMID- 10532550 TI - Inhibitor against coagulation factor V after liver transplantation. AB - A new case of anti-factor V inhibitor is described in a 46-year-old man, who received a liver transplantation for hepatocellular carcinoma, without exposure to bovine thrombin or fibrin glue during the operative course. The inhibition occurred on the 14th postoperative day, while the patient was being treated with oxacillin, azathioprine, and a new immunosuppressive drug, FK506. The inhibition was of short duration (3 days), and no bleeding complication occurred despite a very low plasmatic level of factor V activity and antigen (<5%). Plasma samples drawn after cessation of FK506 disclosed a dose-dependent inhibitory activity when alcoholic solutions of FK506 were exogeneously added; this suggests a possible role of the FK506 drug in the occurrence of this anti-factor V inhibitor. PMID- 10532551 TI - Extracorporeal photochemotherapy for Epstein-Barr virus-associated lymphoma after lung transplantation. AB - Posttransplantation lymphoproliferative disorder (PTLD) is a serious complication after transplantation of solid organs. Highest incidence rates have been reported for lung transplant recipients. With the current treatment strategy for early onset PTLD, a reduction of immunosuppressive drugs, mortality of lung transplant recipients with PTLD remains high, due to both, incomplete control of PTLD and transplant rejection. We present a lung transplant recipient with a history of acute rejection and Epstein Barr virus-associated posttransplantation malignant non-Hodgkin's lymphoma. Extracorporeal photochemotherapy, in combination with a moderate reduction of immunosuppressive therapy, resulted in complete disappearance of PTLD. After a first year of follow-up, no further rejection and no recurrence of PTLD have occurred. Treatment with ECP, with its beneficial effects on both, rejection after organ transplantation and malignant lymphoma, may be a particularly valuable approach for the treatment of PTLD in patients after lung transplantation, with its increased risk for transplant rejection. PMID- 10532553 TI - Primary Vibrio vulnificus bacteremia in a liver transplant recipient after ingestion of raw oysters: caveat emptor. AB - Vibrio vulnificus is responsible for severe infections in chronically ill patients. Organ transplant recipients are also at risk for severe infections due to V vulnificus. We report here the first case of V. vulnificus primary bacteremia due to raw shellfish consumption in a liver transplant recipient. All transplant patients should be cautioned against consuming uncooked seafood and warned about the risk of severe Vibrio infections from seemingly innocuous wounds acquired in a salt water environment. PMID- 10532552 TI - Prevention of de novo hepatitis B infection in recipients of hepatic allografts from anti-HBc positive donors. AB - BACKGROUND: The shortage of donor organs occasionally mandates the use of hepatic allografts from anti-HBc+ donors in recipients who are susceptible to de novo hepatitis B virus (HBV) infection. The efficacy of hepatitis B immune globulin and lamivudine to prevent de novo HBV infection in anti-HBs negative recipients of allografts from anti-HBc+ donors has not been investigated. METHODS: After liver transplantation with an allograft from a donor positive for anti-HBc, recipients who were anti-HBs-, HbsAg- received hepatitis B immune globulin (HBIG) 10,000 IU i.v. daily for 7 days and monthly for 6 months. After 6 months, 1000 IU of HBIG was given IM. every 2 weeks for 18 months. Patients transplanted after 4/1/97 were given lamivudine 150 mg daily starting postoperative day 1. RESULTS: Between 8/14/96 and 6/10/98, 264 orthotopic liver transplants were performed and 16 anti-HBs-, HbsAg- patients received an hepatic allograft from a donor positive for anti-HBc. HBIG mono-therapy was administered to one patient. HBIG and lamivudine combination therapy was administered to 15 patients. Of the 16 patients, 8 were positive only for anti-HBc before transplant, and 8 were naive (anti-HBs-, anti-HBc-). The single patient who received HBIG monotherapy became HbsAg+ at 6 months. All patients receiving combination therapy with HBIG and lamivudine have remained HbsAg-. The average follow-up is 459 days (range 170 754). Two patients died from unrelated causes. CONCLUSIONS: Combination therapy with HBIG and lamivudine may prevent de novo HBV infection in anti-HBs-, HbsAg- recipients of hepatic allografts from anti-HBc+ donors. PMID- 10532554 TI - Hemodynamic follow-up of cardiac allografts from donors undergoing hemodialysis for chronic renal failure. PMID- 10532555 TI - Is living donor nephrectomy a "23-hr stay" procedure? PMID- 10532556 TI - Donor liver allocation: facts and fiction. PMID- 10532557 TI - Current state of the national transplantation system. PMID- 10532559 TI - Bioassayed demineralized bone matrix and calcium sulfate: use in bone-grafting procedures. AB - BACKGROUND AND AIMS: A combination product of bioassayed, demineralized bone matrix (AlloGro, AlloSource, Denver CO) and calcium sulfate pellets (OsteoSet, Wright Medical Technology, Arlington TN) was utilized in a prospective clinical study in 50 patients in need of bone-grafting procedures. It was proposed that the osteoinductive activity of the demineralized bone matrix combined with the osteoconduction and rapid dissolution of the calcium sulfate pellets would complement each other in promoting bone formation. MATERIALS AND METHODS: The patients were evaluated clinically and radiographically at regular intervals post operatively by an independent clinician. A total 10-point healing score was used to determine healing characteristics and progress. Fifty patients (24 males and 26 females) were treated for benign bone lesions (35), nonunion (11), osteomyelitis (3), and acute fracture (1). The average age was 33 years (range, 3 64 years). Lesions were located in the femur (16), tibia (15), humerus (7), and other sites (12). RESULTS: The average length of follow-up was 14 months (range, 6-32 months). Forty-nine of 50 patients healed their lesions (98%), requiring an average time to heal of 11.8 weeks (range, 3-48 weeks). There were no graft related complications. CONCLUSIONS: The results of this preliminary clinical study suggest that a combination of bioassayed demineralized bone matrix and calcium sulfate is very effective in treating benign lesions of bone, as well as nonhealing fractures, which is comparable to grafting with autograft. Future studies have been undertaken utilizing this combination in all acute operative settings and fracture management situations. PMID- 10532558 TI - Dermatofibrosarcoma protuberans at the site of arteriovenous fistula in a renal transplant recipient. PMID- 10532560 TI - Autogenous cultured bone graft--bone reconstruction using tissue engineering approach. AB - Maniatopoulos et al. reported the formation of calcified bone-like tissue when rat bone marrow cells were cultured in the presence of dexamethasone and beta glycerophosphate. We have succeeded to construct the in vitro cultured bone on the porous framework of hydroxyapatite ceramics (HA). After 2 weeks of the culture, the construct showed the existence of mineralized collagen fibers on the surface of HA determined by Scanning Electron Microscopy. The construct also demonstrated high alkaline phosphatase (ALP) activity together with noticeable level of osteocalcin. The results indicate that the construct consisted of thin layer of bone matrix covered hydroxyapatite surface and abundant bone forming active osteoblasts. After the in vivo implantation of the construct, volume of the matrix increased and obvious bone tissue was detected by ordinal microscopy even one-week after implantation and its high osteogenic activity was maintained for a long term (one year). The in vivo bone is biologically active tissue evidenced by Northern blot analysis of the implanted construct which showed ALP and osteocalcin mRNA expression comparable to those of normal cancellous bone. These results demonstrate that the in vitro fabricated cultured bone/HA construct can possess new bone forming capability in in vivo situations. We have also succeeded to fabricate the construct using aged human marrow cells and the construct showed thick lamellar bone formation after the in vivo implantation. Based on these findings, we propose alternative approach for bone reconstruction surgery using the autogenous cultured bone/HA construct. Importantly, we can fabricate the implantable autogenous bone tissue derived from patient's marrow cells and the cells can be obtained by needle aspiration without damaging the patient's normal tissue. PMID- 10532561 TI - The influence of hydroxyapatite granules on the healing of a segmental defect filled with autologous bone marrow. AB - Hydroxyapatite(HA) ceramics are frequently used as a bone graft substitutes for the filling of bony defects. The addition of autologous bone marrow to HA ceramics does improve defect healing. There is conflicting evidence in the literature whether autologous bone marrow transplantation alone is as effective as the combination of HA ceramics and bone marrow combined. It was the purpose of this study to identify the role of additional HA ceramic granules on the healing of a sheep tibia segmental defect filled with autologous bone marrow. After permission of the local animal rights committee was obtained, a 3 cm segmental defect in the midshaft of 31 adult sheep was stabilized with an unreamed tibia nail. The animals were divided into 4 groups according to the mode of defect filling: HA plus autologous bone marrow (HA + MAR) (n = 8), autologous bone marrow (MAR) (n = 9), empty defect (DEF) (n = 6), cancellous bone graft (CAN) (n = 8). After three months follow up animals were sacrificed and analysed for the key parameters of union and maximum torque at failure. One nonunion was present in each of the HA + MAR, MAR, and CAN groups. Four of the six animals in the DEF group developed a nonunion. Maximum torque at failure was reported as percentage of the intact contralateral tibia: HA + MAR 39% +/- 24%, MAR 26% +/- 17%, DEF 22% +/- 13%, CAN 41% +/- 20%. The difference between the groups was statistically significant, but appeared to be relevant. We conclude from our data, that HA ceramics do improve healing of a segmental defect in the sheep tibia filled with autologous bone marrow. The results of this combination are comparable to cancellous autograft. PMID- 10532562 TI - Hydroxyapatite fully coated conic hip prosthetic stem: a long term animal study. AB - The purpose of this paper is to evaluate in an animal model the long term results obtained with a prosthetic stem fully coated with hydroxyapatite. The cup was manufactured in polyethylene and was cemented. Six arthroplasties were performed in six sheep. After twelve months, the animals were euthanized and the femurs were harvested and processed for undecalcified sectioning. Twelve cross sections were cut perpendicularly to the longitudinal stem axis. Sections one to five corresponded to the area of the stem which, at the time of surgery, had a full initial contact between the bone and the prosthesis; sections six to ten corresponded to the area of the stem which, at the time of surgery, had a gap from 0 to 2 mm between the bone and the prosthesis; sections eleven and twelve had an initial gap larger than 2 mm. At one year after implantation, in the sections one to five, morphological analyses showed extensive direct contact between the bone and the hydroxyapatite coating. Bone prosthesis contact was lower in the sections six to ten. No contact was seen in sections eleven and twelve. Comparing bone to prosthesis contact of each subsequent section, from proximal to distal, the difference becomes significant with section five compared to section six (p < 0.00005). No detachment of the hydroxyapatite coating from the metallic substrate was observed in any section. In conclusion, this study shows that a conic shaped femoral stem, fully coated with hydroxyapatite gives very good histological and histomorphometric results at one year. Prosthesis osteointegration showed to be influenced by the initial bone to prosthesis contact. No direct bone to prosthesis contact was achieved if the initial bone to prosthesis gap was larger than 2 mm. PMID- 10532563 TI - Resorbable bone cement for augmentation of internally fixed hip fractures. PMID- 10532564 TI - Musculoskeletal tissue banking in Europe--regulations and quality assurance. AB - This paper is confined to the use of human musculoskeletal tissue in the treatment of patients. Its focus is on the safety and quality dimension of human tissue transplantation, including the ethical and legal aspects, the regulations and standards from the European perspective, quality assurance and quality management in tissue banking and as a special subject, tissue sterilisation and the validation of sterilisation methods. PMID- 10532565 TI - Distal femoral osteoarticular allografts in limb salvage surgery. PMID- 10532566 TI - Bone implants--a challenge to materials science. AB - The review paper discusses the conditions for obtaining in vivo a composite from a synthetic, inorganic material and collagen fibres. Bone itself is a composite containing collagen fibres and hydroxoapatite crystals. The crystal size is, however, far smaller than can be made using conventional methods of ceramics technology. The paper discusses therefore the possibilities to synthesize the apatite in a gel. The requirements of initial load bearing can be met by forming the gel on the surface of glass particle, sintered to a porous body. In the pores calcium ions react with the gel to form (= SiO)Ca+ complexes. When the phosphate concentration in the pore is high enough to exceed the solubility product of apatite, the calcium-gel complexes release the calcium and highly dispersed apatite crystallites are precipitated in the gel. These give the gel the osteoconductive properties observed for bioactive glasses. Glass compositions, which give a gel surface enough hydrated to form calcium complexes, are discussed. PMID- 10532567 TI - Porous bioactive glass matrix in reconstruction of articular osteochondral defects. AB - BACKGROUND AND AIMS: This study was carried out to investigate the use of porous bioactive glass implants in promotion of articular cartilage and subchondral bone repair in large osteochondral joint defects. MATERIAL AND METHODS: Two conical osteochondral defects (top diameter 3.0-3.2 mm) were drilled into the patellar grooves of the distal femurs in the rabbit. The defects, extending (approximately 6-7 mm) from the surface of the articular cartilage to the subchondral marrow space, were reconstructed with size-matched porous conical implants made of sintered bioactive glass microspheres (microsphere diameter 250-300 microm, structural implant compression strength 20-25 MPa) using press-fit technique. The implant surface was smoothened to the level of the surrounding articular cartilage. One of the two defects in each femur was left empty to heal naturally and to serve as the control. At 8 weeks, the defect healing was analyzed with use of a semiquantitative histological grading system, histomorphometry of subchondral bone repair, back-scattered electron imaging of scanning electron microscopy (BEI-SEM), and a microindentation test for characterization for the stiffness properties of the cartilage repair tissue. RESULTS: The porous structure of the bioactive glass implants, extending from the articular defect of the patellar groove into the posterior cortex of the femur, was extensively filled by new bone. Cartilage repair varied from near-complete healing by hyaline cartilage to incomplete healing predominantly by fibrocartilage or fibrous tissue. There were, however, no statistical differences in the histological scores of repair between the glass-filled and control defects, although the sum of the averages of each category was lowest for the bioactive glass filled defects. The indentation stiffness values of all the defects were also significantly lower than that of normal cartilage on the patellar groove. CONCLUSIONS: Porous textures made by sintering bioactive glass microspheres may expand the opportunities in reconstruction of deep osteochondral defects of weight-bearing joints. The implants act mechanically as a supporting scaffold and facilitate the penetration of stromal bone marrow cells and their chondrogenic and osteogenic differentiation. Ionic properties of the bioactive glasses make the substances highly potential even as delivery systems for adjunct growth factor therapy. PMID- 10532568 TI - Can prism adaptation for acquired esotropia be accomplished in a shorter time frame? AB - PURPOSE: Results of the Prism Adaptation Study (PAS) indicated that surgical success rates were highest when augmented surgery was performed for the increased angle of deviation in the prism adaptation responders who underwent surgery for the prism-determined angle of esotropia. The purpose of this study was to see if the prism adaptation response process could be performed in a shorter time span than dictated in the PAS protocol (minimum, 4-7 days). METHODS: After the prescription of appropriate spectacles, patients with acquired stable esotropia were prism adapted and then reexamined and readapted if necessary at 24 hours and 4 to 7 days. The 4- to 7-day visit was the determining visit in the PAS. The question in this study was whether the 24-hour visit would provide the same end point and allow adaptation in a shorter period of time. In addition, a motor end point was used in this study, whereas in the PAS, the end point was both motor and sensory. RESULTS: Thirty-two patients met the entry criteria. Nineteen of the 32 patients built up their entry angle during the prism adaptation process. Thirty of the 32 patients who left the office stable at the 24-hour visit remained stable at the 4- to 7-day visit. CONCLUSION: If the 4- to 7-day visit is used as the "gold" standard, 94% of the patients in this study would not have benefited from a longer (beyond 24 hours) wear time of prisms. With the use of motor stability, prism adaptation can be carried out in a reproducible and timely fashion. PMID- 10532569 TI - Combined resection and recession of a single rectus muscle for the treatment of incomitant strabismus. AB - BACKGROUND: The treatment of incomitant strabismus is challenging. Traditional approaches include the use of asymmetric bilateral surgery and the fadenoperation (posterior fixation suture). We report our experience with a different approach: combined resection and recession of a single rectus muscle. METHODS: The charts of 12 patients who underwent resection of a single rectus muscle with an equal or greater amount of recession of the same muscle were identified. In 5 patients, the procedure was performed using the adjustable suture technique, and the adjustment was performed later the same day (Group 1). In the remaining 7 patients, permanent sutures were placed at the time of surgery (Group 2). The procedure was performed for horizontal and vertical gaze incomitance, dissociated horizontal deviation, and distance-near disparity. RESULTS: Four of the 5 patients in Group 1 showed stable, long-term correction of their incomitance, both in primary gaze and in gaze in the direction of the muscle operated on. The results for patients in Group 2 showed stable, long-term correction of incomitance in 3 patients; however, these patients also had slight overcorrections in the direction of gaze opposite to the muscle operated on. An additional patient in Group 2 had a shift of her distance-near disparity, shifting from relatively exotropic to relatively esotropic disparity postoperatively. All patients in Group 2 showed at least some decrease in the amount of measured incomitance. We did not encounter complications such as muscle slippage or loss, scleral perforation, or late overcorrection in the field of gaze of the operated muscle. CONCLUSIONS: The technique of combined resection and recession of a single rectus muscle shows promise in the treatment of incomitant strabismus. It offers the advantages of posterior fixation combined with the greater technical ease of a standard hangback recession. The muscle may also be placed on an adjustable suture, allowing for postoperative adjustment in selected patients. PMID- 10532570 TI - Role of botulinum toxin A in surgically overcorrected exotropia. AB - PURPOSE: The purpose of this study was to define the role of botulinum toxin type A (BTXA) in surgically overcorrected exotropia. METHODS: A retrospective review was performed using the BTXA clinic database of more than 3500 patients to identify patients with a consecutive esotropia. RESULTS: Sixty patients met the inclusion criteria; the patients' ages ranged from 5 to 80 years. Before toxin treatment, an average of 1.8 operations had been performed per patient. The mean distance deviation was 17 PD base out and near deviation was 18 PD base out. The time from the last operation to an injection of BTXA averaged 28.3 months. We divided our patient population into 2 groups: those with fusion potential and those with no expected fusion potential. Of the 36 patients with fusion potential, 15 patients achieved and maintained good ocular alignment and resolution of their diplopia with an injection of BTXA. In the 24 patients with no expected fusion potential, 4 patients (17%) achieved and maintained good alignment with an injection of BTXA. Although they were not cured, 10 additional patients chose to have repeated BTXA injections to maintain their ocular alignment, whereas only 2 patients required occlusive methods to eradicate intractable diplopia. Five patients had additional surgery, of which 3 patients obtained a functional result. CONCLUSIONS: BTXA has a role in surgically overcorrected exotropia for patients in whom a functional result may be obtained. BTXA is of less value for patients with poor binocular function. It has proved especially useful as a treatment given only once for 42% of patients who could regain high-quality stereopsis. The safety and ease of administration of this treatment add to its merit. PMID- 10532571 TI - What is the role of botulinum toxin in the treatment of dysthyroid strabismus? AB - BACKGROUND: Botulinum toxin A has been used in the treatment of dysthyroid strabismus primarily as a temporary measure during the active phase of the disease. We report on our experience with 65 patients. METHOD: We review the records of 65 patients with dysthyroid strabismus who were treated with botulinum toxin A at Moorfields Eye Hospital between 1984 and 1996. CONCLUSIONS: Patients with a short duration of relatively mild dysthyroid strabismus have a chance of long-term benefit with botulinum toxin A. There is little use for botulinum toxin A in cases of severe dysthyroid disease. PMID- 10532572 TI - Evaluation of methods for assessing visual function of infants. AB - PURPOSE: Commonly used behavioral and electrical testing methods for estimation of visual acuity and visual function in infants yield different estimates and may not accurately predict visual acuity and visual function in later life. Moreover, neither test-retest variability nor side-by-side comparisons of the various methods have been thoroughly evaluated in the same infant population. The purpose of this study was to provide such an evaluation. METHOD: The test-retest variability of visual acuity and visual function was evaluated for the Teller Acuity Card (TAC) procedure, sweep visual evoked potential (VEP), as well as latency and amplitude measured by transient pattern VEP. Groups of approximately 20 infants contributed test-retest data. Visual function estimated by the various methods in a larger group of infants (n = 118) was compared. Correlations between methods and the validity of the various methods to detect maturational changes between 4 and 8 months of age were also assessed. Administration of these tests was according to standard and usual procedures. RESULTS: The average percent difference between test and retest estimates of acuity as well as the SD was lowest for transient VEP latency (3%, 7% SD). The other methods were markedly more variable: sweep VEP (2%, 22% SD), TAC procedure (8%, 20% SD), and transient VEP amplitude (7.5%, 39% SD). Average coefficients of variation showed a similar trend: transient VEP latency, 8%; sweep VEP, 15%; TACs, 30%; and transient amplitude, 53%. Correlations among estimates by the methods were poor, but expected changes in visual maturation from 4 to 8 months of age were detected with all methods. CONCLUSIONS: All methods evaluated provide valid and reliable test-retest data for a group, but are less valid for estimating visual acuity and visual function of an individual subject. The poor correlations between any 2 of the testing methods suggest that each test assesses a different aspect of vision. Nonetheless, expected maturational changes between 4 and 8 months of age were readily detectable by all methods evaluated. PMID- 10532573 TI - Measuring quality in pediatric ophthalmology. AB - Pediatric ophthalmologists should respond to quality expectations inherent in all disciplines of medicine. The specialists must be able to collect data to document and prove the quality of their work if it anticipates continued reimbursement for services. Quality of medical practice is assessed by outcomes research, which is different from clinical research. All specialists are systemically measuring individual and group outcomes across the spectrum of American health care systems. Pediatric ophthalmologists, both in private practice and at academic centers, must concentrate their research efforts on the assessment of cost, quality, utilization, and patient-centered health-related quality of life for the most common pathologies in pediatric ophthalmology practice. PMID- 10532574 TI - Scleral fixated (sutured) posterior chamber intraocular lens implantation in children. AB - BACKGROUND: Optical rehabilitation of unilateral aphakia in eyes with no capsular support is problematic in pediatric patients who cannot tolerate contact lenses. Possible options include a unilateral aphakic spectacle, an anterior chamber intraocular lens (IOL), or a scleral fixated posterior chamber IOL. Of these choices the posterior chamber IOL is the most physiologic. Experience in adults shows increased complications with this technique. OBJECTIVE: The objective of this study was to report the short-term results and complications of unilateral scleral fixated posterior chamber IOLs in the pediatric population. METHODS: All patients with scleral fixated lenses younger than 16 years were retrospectively reviewed. Nine patients aged 12 months to 15 years underwent unilateral scleral fixated posterior chamber lens implantation using buried polypropylene fixation sutures. Follow-up averaged 24 months. RESULTS: Postoperative visual acuity improved in all patients. Refractive goals were achieved in all but 1 patient. Complications included elevated intraocular pressure controlled with medications (1 patient), anterior uveitis (1 patient), and mild IOL decentration (1 patient). CONCLUSIONS: Although short-term visual results appear encouraging, this procedure is technically more difficult and has an increased incidence of postoperative complications when compared with secondary sulcus-fixated IOLs supported by capsular remnants. Caution should be exercised when recommending this procedure for pediatric patients because long-term risks are unknown. PMID- 10532575 TI - The IOLAB, Inc pediatric intraocular lens study. AAPOS Reasearch Committee. American Association for Pediatric Ophthalmology and Strabismus. AB - PURPOSE: This report is a summary of the data of the IOLAB, Inc pediatric intraocular lens (IOL) implantation investigation. The goal of this study was to evaluate the safety and efficacy of IOL implantation for the treatment of pediatric aphakia, pending approval by the Food and Drug Administration. METHODS: From May 1981 to July 1994, a total of 1260 pediatric eyes received 171 styles of IOLs implanted by 361 US investigators. Preoperative, operative, and postoperative status reports over the first year were required for each eye entered into the study. Annual visit reports were requested thereafter to determine the long-term effects. The study was terminated in November 1995. All IOLs were obtained from IOLAB, Inc (now Chiron Vision Corp). RESULTS: Reporting compliance was 98.3% for the preoperative and operative reports, 45.1% at 1 year, and 13.8% at 3 years. The subjects' ages ranged from younger than 1 yearto 17 years. Nine subjects (0.7%) were younger than 1 year, with the largest group of 533 subjects (42.3%) aged between 6 and 12 years atthe time of surgery. Cataract types were congenital (45.6%), traumatic (37.1%), secondary (11%), senile (0.95%), and unrecorded (5.4%). The IOL was implanted primarily in 74.8% of cases and secondarily in 21.4% of cases. There was no record in 3.8% of the cases. IOL types included anterior chamber (4.1%), iridocapsular (0.71%), posterior chamber (93.6%), and unrecorded (1.59%). There were 130 adverse reactions that required secondary surgical intervention. The most frequently performed surgical procedures included lens removal without replacement, vitrectomy, lens repositioning, and lens replacement. More than half (52%) of all eyes had a visual acuity of 20/200 or worse before surgery; amblyopia was reported in 21.1% of all participants at baseline. Postoperative visual acuity data were available on 563 eyes at 1 year after surgery. Overall, 52.8% of all eyes attained a visual acuity of 20/40 or better by the 1-year visit, and only 15.5% had visual acuity worse than 20/200. In general, the older patient, traumatic cataract, and secondary cataract categories were overrepresented in the better visual acuity outcome group. CONCLUSION: The IOLAB, Inc pediatric IOL study is the first multiple-practitioner, national study designed to evaluate the safety and efficacy of IOL implantation in children. The study results are compromised by the almost 50% loss of follow-up at the 1-year evaluation. Other variables that most likely influenced outcome results were the methods of cataract extraction, medical management, and IOL design, all of which evolved dramatically over the time course of the study. Despite these issues, pediatric IOL implantation seems to be a reasonable treatment modality for aphakia, on the basis of the available 1-year follow-up data of the remaining 45.1% of eyes in the study. PMID- 10532576 TI - Anterior hyaloid face opacification after pediatric Nd:YAG laser capsulotomy. AB - PURPOSE: The purpose of this study was to examine the clarity of the visual axis after Nd:YAG laser capsulotomy following cataract extraction and primary intraocular lens implantation in a pediatric population. METHODS: A retrospective review was performed of all cases of cataract extraction and primary intraocular lens implantation over a period of 5 years. A group of children who had been treated by primary surgical posterior capsulotomy and anterior vitrectomy (Group 1) was used as the "gold standard," with whom the children treated with Nd:YAG laser capsulotomy (Group 2) were compared. The groups were studied for the incidence of opacification of the visual axis after the primary procedure. RESULTS: Data on 78 eyes were reviewed, and 56 eyes met inclusion criteria. Of these, 33 eyes were treated with primary posterior capsulotomy and anterior vitrectomy (Group 1) and 23 eyes were treated with Nd:YAG laser capsulotomy (Group 2). One eye (3%) of Group 1 experienced postoperative visual axis reopacification. Thirteen (57%) of 23 eyes in Group 2 experienced reopacification, requiring retreatment. Four eyes (17%) treated with Nd:YAG laser required a third treatment. CONCLUSIONS: In our series, 57% of patients treated with Nd:YAG laser capsulotomy experienced reopacification across the anterior hyaloid face. With the removal of the anterior vitreous at the time of cataract extraction, the scaffolding for cell migration is removed and reopacification of the visual axis is rarely seen. For patients in whom slit-lamp capsulotomy is not possible, especially if there is no Nd:YAG laser available for use in the operating room or when loss to follow-up may be an issue, primary posterior capsulotomy and anterior vitrectomy should be strongly considered. PMID- 10532578 TI - An uncommon cause of ophthalmia neonatorum: Neisseria meningitidis. AB - Ophthalmia neonatorum is defined as conjunctivitis appearing during the first month of life. The differential diagnosis includes chemical, bacterial, viral, and other pathogens, including Neisseria gonorrhoeae, herpes simplex, and Chlamydia trachomatis. Neisseria meningitidis is not commonly specifically included in the differential. PMID- 10532577 TI - The epidemiology of pediatric glaucoma: the Toronto experience. AB - BACKGROUND: This study was conceived to provide an insight into the spectrum of glaucoma in the pediatric population. We also set out to compare the success of disease control and the prognosis for vision within the different diagnostic subgroups. This is the largest single population of children with glaucoma that has been so described and compared. METHODS: The charts of children who were first seen between birth and age 16 years and who attended the Hospital for Sick Children with any form of glaucoma between January 1974 and January 1995 were reviewed and entered into the study. RESULTS: Data are presented for 306 children. Congenital glaucoma was the most common subtype, accounting for 38%. Patients with congenital glaucoma were young, had surgery, and had more operations than any other group except those with aniridia. Goniotomy offered a cure in 47.8% of the patients. A bimodal distribution reflected their visual performance. Patients with aphakic glaucoma, the next most prevalent group (20%), presented at an older age (4.5 years). Surgical intervention was performed in 50% of these children. Nearly all patients with Sturge-Weber syndrome (80%) had surgery. The following glaucoma groups were associated with a poor visual outcome: aniridia, anterior segment developmental anomalies involving the cornea, uveitis with glaucoma other than steroid induced, retinopathy of prematurity, and persistent hyperplastic primary vitreous. Steroid-induced glaucoma and anterior segment dysgenesis, excluding Peters anomaly, had uniformly good outcomes. CONCLUSION: The ability to control glaucoma in childhood and visual prognosis is highly variable. Particular diagnostic categories do consistently well and some do poorly. PMID- 10532579 TI - Ocular injuries caused by fireworks. AB - What are the consequences of suddenly legalizing fireworks sales in a largely rural society? Would the spectrum of ocular injuries caused by fireworks differ from those found in the Western world? This is the first study on ocular injuries caused by fireworks conducted in the Republic of South Africa. We analyzed the presenting features and prospectively followed up all patients who presented to the casualties served by our ophthalmic department over the New Year celebrations of 1996-1997. The sale of fireworks to the public had been deregulated the previous year. Ocular injuries caused by fireworks had not been reported before 1995. We found that ocular injuries caused by fireworks occurred mainly in young male patients. The injuries were usually unilateral and responded to treatment. This mirrors worldwide studies that show that it is children who are frequently harmed by fireworks injury. Two of our patients were blinded by their injuries. Our findings echo those found in Western countries where fireworks have not been restricted by law. We suggest that young boys, regardless of race, nationality, literacy, or social circumstances, are at risk for ocular injuries caused by fireworks. Countries planning to unban fireworks should aim their education program at this target group. PMID- 10532580 TI - Congenital malignant rhabdoid tumor of the orbit. AB - Malignant rhabdoid tumor is a rare and highly malignant renal tumor of infancy. Extrarenal tumors involving the orbit have been reported, but never at birth. The authors describe a primary malignant rhabdoid tumor of the orbit in a neonate who had massive unilateral proptosis at birth. Clinical, radiographic, and histologic features of the tumor are discussed. PMID- 10532581 TI - Incidence and severity of retinopathy of prematurity. PMID- 10532582 TI - Hashimoto's thyroiditis: an organ-specific autoimmune disease--pathogenesis and recent developments. PMID- 10532583 TI - Direct effects of endotoxin on the endothelium: barrier function and injury. AB - LPS directly disrupts EC barrier function in vitro and in vivo. This barrier dysfunction has been reported to occur in EC derived from both the macro- and microvasculature of varying species, including humans. Unlike other EC responses, LPS-induced loss of endothelial barrier function is protein-synthesis independent. In fact, protein synthesis inhibition enhances the LPS effect. The lipid A moiety is responsible for LPS-induced activation of the non-CD14-bearing EC, and agents that bind to and neutralize this highly conserved portion of the LPS molecule can crossprotect against EC barrier dysfunction elicited by LPS derived from diverse species of Gram-negative bacteria. Although the presentation of LPS to CD14-bearing cells such as macrophages and monocytes has been well characterized, far less is known about the interactions of LPS with the non-CD14 bearing EC. An EC receptor involved in LPS binding and cellular activation has yet to be identified. The presence of the accessory molecules, LBP and sCD14, are prerequisite to LPS-induced activation of EC at clinically relevant LPS concentrations. As with monocytes and macrophages, the CD14 dependence of LPS induced endothelial barrier dysfunction can be overcome with high concentrations of LPS. In the absence of LBP and sCD14, a 200,000-fold increase in LPS concentration is required to elicit the same increments in EC monolayer permeability relative to when these accessory molecules are present. Within 30 minutes after LPS exposure, PTK activation is observed. PTK inhibition blocks LPS induced EC actin depolymerization and endothelial barrier dysfunction which are seen only after a > or = 2-hour stimulus-to-response lag time. Furthermore this LPS-induced actin depolymerization is a prerequisite to opening up the paracellular pathway and loss of monolayer integrity. Interestingly LPS-induced increments in transendothelial 14C-BSA flux and EC detachment parallel caspase mediated cleavage of ZA and FA proteins that participate in cell-cell and cell matrix adhesion. The cleavage of the ZA components, beta- and gamma-catenin, does not affect their ability to bind the transmembrane protein, cadherin, or the actin-binding protein, alpha-catenin, suggesting that the linkage of the ZA to the actin cytoskeleton remains intact. LPS-induced cleavage of the FA protein, FAK, leads to dissociation of its catalytic domain from paxillin substrate and decreased paxillin phosphotyrosine content. Caspase inhibition protects against LPS-provoked apoptosis, cleavage of adherens junction proteins, paxillin dephosphorylation, cell-shape changes, and EC detachment. In contrast it fails to block LPS-induced increments in transendothelial 14C-BSA flux. PTK inhibition, which does protect against increased transendothelial 14C-BSA flux, does not block LPS-induced proteolytic cleavage events and only partially inhibits EC detachment. These findings suggest that the EC detachment and endothelial barrier dysfunction elicited by LPS are mediated through distinct pathways (Fig. 6). Much of the work to date has focused on LPS interactions with mCD14-bearing cells, such as monocytes and macrophages, which are central to the inflammatory response elicited by endotoxin. EC, which line the vasculature, are one of the first host tissue barriers to encounter circulating LPS. Because damage to the endothelium is known to contribute to the development of multiorgan failure, including ARDS, understanding LPS-induced EC dysfunction in the setting of Gram-negative septicemia has clear pathophysiologic implications. (ABSTRACT TRUNCATED) PMID- 10532584 TI - Analysis of nucleotides and aromatic amino acids in normal and neoplastic colon mucosa by ultraviolet resonance raman spectroscopy. AB - The objective of this study was to explore the potential of using ultraviolet resonance Raman (UVRR) spectroscopy to analyze normal and neoplastic colon tissue. Ultraviolet light at 251 nm, generated from the third harmonic of a Titanium:Sapphire laser, was used to irradiate the surfaces of surgically resected human colon specimens from six patients, five clinically diagnosed with adenocarcinoma, and one with familial adenomatous polyposis. All grossly neoplastic samples found to contain mucosal dysplasia or invasive adenocarcinoma upon histologic evaluation, were analyzed in parallel with normal tissue obtained from the same specimen and located at least 1 cm away from grossly neoplastic tissue. The colon spectra were modeled as a linear combination of nucleotide, aromatic amino acid, and lipid lineshapes, using chemical standards as a reference. Nucleotide and amino acid contributions to the UVRR spectra were quantified by a least squares minimization method. The least squares minimization spectral model was verified in aqueous solutions, where relative concentrations of free nucleotides and DNA were quantified with < 10% error. Of the 11 neoplastic samples studied from the 6 specimens, 10 showed either a lower amino acid/nucleotide ratio, a lower level of adenyl (A) signal, or both when compared with their normal counterpart. Lower amino acid/nucleotide ratio was present in five of six samples containing only dysplasia, and three of the five samples containing invasive adenocarcinoma. Lower A was present in all five samples containing invasive cancer, and in three of the six samples containing only dysplasia. This lower level of A corroborates previously published biochemistry work showing a lower level of total adenylates in tumor homogenates compared with normal tissue. Our data indicate that surface UVRR may provide unique information about site-to-site changes in cellular metabolites during colon carcinogenesis. PMID- 10532586 TI - Lysophosphatidic acid as a regulator of endothelial/leukocyte interaction. AB - Lysophosphatidic acid (LPA) is produced by a variety of activated cell types and acts as an intercellular mediator of processes associated with inflammation and repair including platelets aggregation, and smooth muscle and fibroblast proliferation. However no previous studies have examined the effects of LPA on endothelial cell leukocyte interactions. We have examined the ability of LPA to activate human aortic endothelial cells (HAEC) to bind monocytes, neutrophils, and HL60 cells (a neutrophil surrogate). Treatment of HAEC for 4 hours with 10 microM LPA caused an increase in the binding of monocytes, neutrophils, and HL60. LPA but not phosphatidic acid dose-dependently increased E-selectin and vascular cell adhesion molecule-1 (VCAM-1) cell surface expression. We performed several studies to characterize the receptor mediating the LPA effect. We demonstrate that at least five potential LPA receptors are expressed by HAEC: Edg-1, -3, -4, and -5 as well as PSP24. Cyclic phosphate-containing phosphatidic acid analogue, an agonist for the type 3 low affinity LPA receptor, was not effective in activating HAEC to bind leukocytes, excluding a role for this receptor. The selective receptor antagonists N-palmitoyl-serine and N-palmitoyl-tyrosine (which inhibits PSP24) completely inhibited LPA-induced VCAM expression; however these antagonists inhibited E-selectin expression by only 30%, suggesting a role for at least one additional LPA receptor mediating E-selectin expression. We propose that Edg-1 might be the second receptor, because this receptor, when expressed in HEK293 cells, similarly to the PSP24 receptor, caused ERK activation to nanomolar concentration of LPA. Exposure of HAEC to sphingosine-1-phosphate, another Edg-1 receptor agonist, increased surface expression of E-selectin and to a much smaller extent VCAM-1. The effects of both LPA and sphingosine-1-phosphate on the induction of both VCAM-1 and E-selectin expression was abolished by pretreatment with pertussis toxin suggesting that both LPA receptors in HAEC couple to a Gi pathway. These findings reveal an important and novel role for LPA and its receptors in inflammatory processes. PMID- 10532585 TI - Nitric oxide synthase in human breast cancer is associated with tumor grade, proliferation rate, and expression of progesterone receptors. AB - Nitric oxide (NO) is generated by a family of isoenzymes named nitric oxide synthases (NOS) which includes a cytokine-inducible form, NOSII. NO is a free radical known to inhibit cell proliferation, to induce apoptosis, and to be a mediator of macrophage cytostatic and cytotoxic effects. We investigated NOS in 40 human breast carcinomas and 8 benign breast lesions. NOSII was localized in tumor cells by immunohistochemistry. NOS activity, measured with the citrulline assay, was detected in 27 of 40 tumors. Neither immunohistologic labeling nor NOS activity was detected in benign samples. NOS labeling and activity were significantly related (p < 0.02). For the first time, a significant negative relationship between NOS activity and tumor cell proliferation (p < 0.002) was found. We also showed that tumors with high NOS activity expressed progesterone receptors (p < 0.04). These results are consistent with the observation of high NOS activity in tumors with low grade (p < 0.05). These in vivo observations were related to in vitro data: cytokines (IL-1beta, IFN-gamma, and TNF-alpha) induced NOSII expression in human MCF-7 breast cancer cells, and NO inhibited their proliferation. Thus, we show herein that tumors with high NOS activity have low proliferation rate and low grade, which correlates with the in vitro observation of the inhibition of proliferation of human breast cancer cells by NO. These results may have future therapeutic implications. PMID- 10532587 TI - Identification of the region in the N-terminal domain responsible for the cytoplasmic localization of Myoc/Tigr and its association with microtubules. AB - Mutations in the MYOC/TIGR gene are associated with juvenile open-angle glaucoma and in some cases may be involved in the formation of sporadic primary open-angle glaucoma in humans. To better understand the functions of the MYOC/TIGR protein, its intracellular distribution was investigated using green fluorescent protein (GFP) as a marker. The results indicated that the recombinant mouse and human Myoc/Tigr-GFP proteins are located in the cytoplasm of the transfected cells in which they colocalize with microtubules. Deletion analysis demonstrated that the N-terminal region (positions 1-124 and 15-138 in the mouse and human proteins, respectively) encoded by exon 1 is critical for the cytoplasmic localization of Myoc/Tigr. Most of the known mutations in the human MYOC/TIGR gene implicated in juvenile and sporadic primary open-angle glaucoma formation are located outside the region responsible for the cytoplasmic localization of the protein. However, some of these mutations may alter the tertiary structure of the protein and subsequently modify its interaction with microtubules. PMID- 10532588 TI - Differential expression of individual complement regulators in the brain and choroid plexus. AB - Membrane bound regulators of complement (C) control the system at key points during activation. To determine whether C regulators were expressed in the central nervous system, temporal cortex, and choroid plexus, tissues from eight adult humans were obtained at postmortem and surgery. Tissue was taken fresh for total RNA isolation, snap freezing, or processing in paraffin wax for immunocytochemistry and in situ hybridization. Immunocytochemistry of temporal cortex using anti-CD59 stained microglia intensely; astrocytes and neurons weakly. Microglia were unequivocally stained with anti-membrane cofactor protein (MCP) whereas staining on astrocytes and neurons was weak. Decay accelerating factor (DAF) was strongly expressed by microglia but weakly by astrocytes. Neurons expressed neither DAF nor complement receptor 1 (CR1). CR1 was also absent on astrocytes and microglia. The choroid plexus epithelium revealed intense apical staining with antibodies to CD59, less strongly with anti-MCP and weakly with anti-DAF. CR1 was detected only on phagocytic Kolmer cells in the choroid plexus. Reverse transcriptase-polymerase chain reaction revealed CD59, MCP, and to a lesser degree, DAF mRNA both in the choroid plexus and temporal cortex. CR1 mRNA was detected in choroid plexus samples only. Digoxigenin-UTP labeled riboprobes to all four membrane regulators were used for in situ hybridization. DAF, MCP, and CD59 mRNA were expressed by epithelial cells of the choroid plexus and CR1 mRNA was found only in Kolmer cells. In the temporal cortex, MCP and CD59 mRNA were expressed by glia and at low level by neurons, but DAF was not detected. Previous studies have suggested that C produced in inflamed brains in conditions such as Alzheimer's and Huntington's diseases can be specifically toxic to neurons. The demonstration herein that neurons express only very low levels of CD59 and MCP and lack both CR1 and DAF might explain their susceptibility to C damage. PMID- 10532589 TI - Analysis of mammary carcinoma onset and progression in HER-2/neu oncogene transgenic mice reveals a lobular origin. AB - Morphologic examinations of mammary neoplasias arising in BALB/c (H-2d) mice carrying the activated rat HER-2/neu oncogene (BALB-NeuT), and in FVB (H-2q) mice bearing the wild-type proto-oncogene (FVB-NeuN), indicate that both conditions result in a very human-like lobular carcinoma of alveolar type, whose histotype is the result of the preferential expression of HER-2/neu products in the epithelium of lobular ducts and lobules. Detailed analysis of tumor progression indicates that transition from lobular hyperplasia to overt carcinoma is associated with a high epithelial proliferation rate, as assessed by anti proliferating cell nuclear antigen immunostaining, and coincides with the activation and maximal extension of tumor angiogenic process as assessed by microvessel count (anti-CD31), anti-beta3 integrin, and anti-laminin immunostaining. Neovascularization is accompanied by vascular endothelial cell growth factor and basic fibroblast growth factor production by hyperplastic epithelial cells. By contrast with the BALB-NeuT tumors, E-cadherin expression is almost nonexistent in those arising in FVB-NeuN mice and this may explain their high metastatic potential. Despite their different kinetics, however, the lung metastases observed in both strains are histologically similar and resemble the primary tumor. Both strains can thus be proposed as models for "in vivo" investigation of the origin and progression of the alveolar type of lobular mammary carcinoma and the testing of new therapeutic approaches. PMID- 10532590 TI - The betaA4 amyloid peptide complexes to and enhances the uptake of beta-very low density lipoproteins by the low density lipoprotein receptor-related protein and heparan sulfate proteoglycans pathway. AB - We recently suggested that soluble beta-amyloid (betaA4) is a ligand of the low density lipoprotein receptor-related protein and heparan sulfate proteoglycan pathway. In the blood and in the cerebrospinal fluid, betaA4 is bound to apolipoprotein E containing lipoproteins. We examined how binding of betaA4 to beta-very low density lipoproteins (betaVLDL) alters their cellular metabolism. Compared with betaVLDL alone, complexes of betaVLDL and betaA4 were internalized, but not degraded at increased rates in fibroblasts and in rat hippocampal cells. The uptake of complexes of betaVLDL and betaA4 was not mediated by the low density lipoprotein receptor. BetaA4 not complexed to betaVLDL competed with the endocytosis of alpha2-macroglobulin and apolipoprotein E-enriched betaVLDL. The uptake of complexes of betaVLDL and betaA4 was inhibited by heparin, suramin, lactoferrin, the 39-kd receptor-associated protein, and alpha2-macroglobulin. Complexes of betaVLDL and betaA4 were taken up at reduced rates in Chinese hamster ovary cells partially (pgsB-650) or completely lacking (pgsA-745) proteoglycans. BetaA4 in which the positively charged amino acids between positions 13 and 17 (HHQKL) were replaced by glycine (GGQGL) failed to enhance the uptake of betaVLDL. Together, the data suggest that binding of betaA4 to betaVLDL produces particles that are endocytosed by low density lipoprotein receptor-related protein and HSPG. Complexes of betaVLDL and betaA4 had an intracellular half-life 4-fold that of native betaVLDL, did not undergo lysosomal degradation, and were resecreted into the culture medium. These findings represent the first identification of an endocytotic pathway for betaA4 and may be of relevance to the pathobiochemistry of neurodegenerative disorders. PMID- 10532591 TI - Differential expression of the C5a receptor on the main cell types of rat liver as demonstrated with a novel monoclonal antibody and by C5a anaphylatoxin-induced Ca2+ release. AB - The C5-anaphylatoxin (C5a) is a protein of 74 (human) or 77 (rat) amino acid residues, respectively, which is generated by limited proteolysis upon activation of the fifth component of complement. Its generation may be induced by both the classical and alternative pathways. C5a has been shown to indirectly increase glucose output from hepatocytes (HC) in perfused rat liver by inducing prostanoid release from Kupffer cells (KC) and hepatic stellate cells (HSC). A direct action of C5a on hepatocytes would require their expression of the specific C5a receptor (C5aR). In former studies using quantitative reverse transcription polymerase chain reaction (RT-PCR) it was shown that HC lack this receptor in contrast to KC, HSC and, probably, sinusoidal endothelial cells (SEC), all of which contained mRNA for the C5aR in decreasing amounts. Using a novel monoclonal antibody (mAb R63) against the rat receptor, expression of the rat receptor on the four cell types was investigated by FACS analysis, immunohistochemistry, and immunocytochemistry. The data obtained were confirmed by functional studies in which the Ca2+ response after stimulation of the isolated cells with recombinant rat C5a (rrC5a), the ligand for the receptor was recorded. The FACS and the immunocytochemical data presented here clearly indicate that rat HC do not express the C5aR, whereas KC have the highest expression level followed by HSC. SEC expressed the receptor only weakly. In line with these findings, a strong Ca2+ response was observed after stimulation of KC and HSC, and a weak one with SEC. However, no signal was obtained upon stimulation of HC. The results of this study support the indirect stimulation of glucose output from HC via prostanoid release from nonparenchymal liver cells and contradict the formerly proposed hypothesis of a direct action of C5 anaphylatoxin on hepatocytes. PMID- 10532592 TI - Interferon-gamma up-regulates expression and activity of P-glycoprotein in human peripheral blood monocyte-derived macrophages. AB - P-glycoprotein (Pgp), the product of the multidrug resistance (MDR1) gene, is expressed in a variety of normal tissues but very little is known about its expression and function in cells of the immune system. In this study, we investigated the effect of interferon-gamma (IFN-gamma) on the expression and activity of Pgp in human peripheral blood monocyte-derived macrophages (MDM). We report that IFN-gamma up-regulated Pgp expression in a dose- and time-dependent manner. We show that IFN-gamma slightly increased the accumulation of MDR1 mRNA and induced a polarized redistribution of Pgp, as well as of some cytoskeletal proteins (ie, ezrin, actin, and alpha-actinin) on cell pseudopodia. Notably, confocal microscopy studies showed that Pgp and ezrin colocalized in these cellular structures. The IFN-gamma-induced Pgp up-modulation was a specific response of primary macrophages, as IFN-gamma treatment of primary lymphocytes and monocytic cell lines did not result in any increase of Pgp expression. Finally, IFN-gamma stimulated the Pgp transport activity in MDM, as rhodamine 123 efflux increased in treated cells as compared with control cultures. These results indicate that Pgp expression and activity can be up-regulated in human MDM in response to IFN-gamma. We suggest that IFN-gamma may be involved in the induction of multidrug resistance in macrophages. PMID- 10532593 TI - The PKD1 gene product, "polycystin-1," is a tyrosine-phosphorylated protein that colocalizes with alpha2beta1-integrin in focal clusters in adherent renal epithelia. AB - Mutations in the PKD1 gene are responsible for autosomal dominant polycystic kidney disease (ADPKD). Although PKD1 has been cloned and shown to be expressed at high levels in the fetal ureteric bud and ADPKD cystic epithelia in the human kidney, the function of its encoded protein, "polycystin-1" is unknown. In this study we used primary and immortalized human renal epithelial cell lines derived from normal fetal, adult, and ADPKD kidneys, that endogenously express PKD1, to study the biologic function of the polycystin-1 protein. ADPKD renal epithelial cells expressed high levels of polycystin-1 protein and showed increased adhesion to type I collagen by comparison with normal adult human renal epithelia that expressed little polycystin. Adherent ADPKD cells also expressed high levels of alpha2beta1-integrin and their attachment was inhibited by a functional monoclonal antibody to alpha2-integrin. Double labeling and confocal microscopy as well as coimmunoprecipitation analysis showed overlapping colocalization of polycystin-1 with alpha2beta1-integrin as well as with the focal adhesion proteins vinculin and paxillin in multiprotein clusters localized to focal areas of cell membrane contact with type I collagen matrix after short periods of attachment. Immunoprecipitation and Western immunoblot studies also showed that polycystin-1 was posttranslationally modified by tyrosine phosphorylation. These studies suggest that the PKD1-encoded protein is part of a large multiprotein complex in epithelial cells that functions in the regulation of extracellular matrix interactions with the plasma membrane and cell cytoskeleton. PMID- 10532594 TI - The status of new methods for the detection of red cell agglutination. PMID- 10532595 TI - Foundling viruses and transfusion medicine. PMID- 10532596 TI - Comparison of the performance of four microtube column agglutination systems in the detection of red cell alloantibodies. AB - BACKGROUND: The purpose of this study was to compare the performance of four currently available microtube column agglutination systems in the detection of red cell alloantibodies to that of the standard tube low-ionic-strength solution (LISS) indirect antiglobulin test (IAT) (tube LISS-IAT). STUDY DESIGN AND METHODS: In a comparative study, 172 sera, previously demonstrated to contain red cell alloantibodies, were tested in parallel by the tube LISS-IAT and three microtube column agglutination techniques (DiaMed-ID, Ortho BioVue, and Sanofi Pasteur Scangel) and one affinity-adherence test system (Gamma ReACT). Tests were performed simultaneously by a single person on freshly thawed sera that had been frozen at -20 degrees C. RESULTS: The rate of detection of clinically significant alloantibodies (n = 154) in microtube column systems was very similar. One hundred forty-one sera (91.6%) reacted in the DiaMed-ID, 139 (90.3%) in the ReACT, 139 (90.3%) in the BioVue, and 142 (92.2%) in the Scangel. Only 117 (76.0%) of these sera reacted in the tube LISS-IAT. The detection rates for 18 antibodies of minor clinical significance (anti-M, -N, -P1, -Le(a), and -Le(b)) varied among the test systems: DiaMed-ID, 5 (28%); ReACT, 7 (39%); BioVue, 14 (78%); Scangel, 10 (56%); and tube LISS-IAT, 6 (33%). Antibody reactivity as determined by titer and score was very similar in all microtube column systems and higher in these systems than in the tube LISS-IAT. CONCLUSION: The sensitivity of all four microtube column systems in the detection of clinically significant red cell alloantibodies was similar and was markedly superior to that of the tube LISS-IAT. An individual cost-benefit analysis should be performed in every institution to decide whether a microtube column system should be applied. If so, the antibody screen in the microtube column agglutination system should ideally be performed in advance of the crossmatch to provide time to screen for compatible units. PMID- 10532597 TI - Ultraviolet and visible light spectrophotometric approach to blood typing: objective analysis by agglutination index. AB - BACKGROUND: A new blood typing technology based on ultraviolet (UV) and visible light spectroscopy (UV/visible spectroscopy) has been developed. Blood groups and types are determined by quantifying reproducible changes in the UV and visible light spectra of blood in the presence of agglutinating antibodies. STUDY DESIGN AND METHODS: Samples of red cells in the presence and absence of agglutinating antibodies were examined by UV/visible spectroscopy. Blood groups and types were determined by comparing the optical density spectra obtained between 665 and 1000 nm. These comparisons generate numbers (agglutination index) ranging from 0 to 100, with smaller numbers corresponding to lack of agglutination and larger numbers corresponding to agglutination. RESULTS: The optical density of agglutinated blood is dramatically different from that of unagglutinated blood. The agglutination index derived from the relative slopes of the spectra is an objective indicator of agglutination strength. An agglutination index greater than 17 consistently and accurately established blood group- and type-specific agglutination. CONCLUSION: The method accurately predicted A, B, and O blood groups, and D type in over 275 samples. Scattering theory-based calculations of relative volumes of red cells before and after agglutination show a direct correlation with the agglutination index and provide the theoretical basis of the analysis. This quantitative technique is reproducible and has the potential for automation. PMID- 10532598 TI - A comparison of the Aplysia lectin anti-I specificity with human anti-I and several other I-detecting lectins. AB - BACKGROUND: Lectins displaying blood group specificity are important for blood group typing and antigen recognition. Their use in blood banks is especially widespread in situations where there is a shortage of specific antisera. This report describes the efficiency of Aplysia gonad lectin as a reliable reagent for the detection of I antigen, which is common on adult human cells but reduced in fetal, newborn, and rare adult red cells. STUDY DESIGN AND METHODS: The selective hemagglutinating activity of the Aplysia lectin was compared with that of human anti-I and several I-reactive lectins, including two plant lectins, one galactophilic microbial lectin, and bovine spleen galectin. RESULTS: The comparison has revealed that Aplysia gonad lectin, like human anti-I, strongly agglutinates and adsorbs to adult I-positive red cells, differentiating between them and fetal or rare I-negative adult red cells (although with less of a difference). In contrast to the plant and microbial lectins examined, its I affinity does not depend on the presence of ABH or P system antigens and it clearly detects higher I antigen expression in Oh red cells. The hemagglutinating activity of Aplysia lectin as that of all the I-detecting proteins is enhanced at 4 degrees C, but unlike the human anti-I Aplysia lectin-induced hemagglutination is stable at room temperature. CONCLUSIONS: The Aplysia lectin is a reliable anti I reagent, which strongly agglutinates I-positive adult human red cells irrespective of their ABH or P system antigens. This lectin is usable at room temperature. PMID- 10532599 TI - Antibodies provoked by the transfusion of biotin-labeled red cells. AB - BACKGROUND: Biotin-labeled (biotinylated) red cells (B-RBCs) offer a technique by which to study RBC volume and circulating kinetics without in vivo radiation. The immunogenicity of B-RBCs is undefined. STUDY DESIGN AND METHODS: To determine if biotinylation renders RBCs immunogenic, autologous B-RBCs were transfused to 20 healthy subjects, and plasma samples were obtained before transfusion and serially for up to 6 months after transfusion. These serial samples, plus plasma from 20 normal control subjects not given B-RBCs, were screened for antibodies to B-RBCs by use of an antiglobulin technique against aliquots of group O RBCs from a single donor-one aliquot biotinylated and one aliquot not biotinylated (i.e., test and control RBCs). Posttransfusion recovery and survival of B-RBCs were also determined. RESULTS: Plasma from none of 20 normal nontransfused subjects reacted with B-RBCs. Similarly, none of the 20 subjects given autologous B-RBC transfusions exhibited antibodies before transfusion. However, 3 of the 20 subjects transiently produced antibodies to B-RBCs after transfusion. Antibodies disappeared within 6 months in 2 of these 3 subjects and within 12 months in the third. Antibody reactivity was not reduced by dithiothreitol, but in 2 of the 3 subjects, B-RBC antibodies were neutralized by incubation with biotin solution. Circulating RBC kinetics were not altered in the 3 subjects with antibody. The significance of these observations is unclear, because antibodies were just beginning to emerge during the studies. CONCLUSIONS: Biotinylation does not render RBCs reactive with normal human plasma (i.e., presumably does not evoke neoantigens). Transfused B-RBCs occasionally provoke IgG antibodies in healthy subjects. Because the biologic effects of B-RBC antibodies currently are unknown, testing for them is recommended when multiple B-RBC transfusions are given to study RBC volume or circulating kinetics. PMID- 10532600 TI - Lowering the hemoglobin threshold for transfusion in coronary artery bypass procedures: effect on patient outcome. AB - BACKGROUND: There is controversy regarding the application of transfusion triggers in cardiac surgery. The goal of this study was to determine if lowering the hemoglobin threshold for red cell (RBC) transfusion to 8 g per dL after coronary artery bypass graft surgery would reduce blood use without adversely affecting patient outcome. STUDY DESIGN AND METHODS: Consecutive patients (n = 428) undergoing elective primary coronary artery bypass graft surgery were randomly assigned to two groups: study patients (n = 212) received RBC transfusions in the postoperative period if the Hb level was < 8 g per dL or if predetermined clinical conditions required RBC support, and control patients (n = 216) were treated according to individual physician's orders (hemoglobin levels < 9 g/dL as the institutional guideline). Multiple demographic, procedure-related, transfusion, laboratory, and outcome data were analyzed. Questionnaires were administered for patient self-assessment of fatigue and anemia. RESULTS: Preoperative and operative clinical characteristics, as well as the intraoperative transfusion rate, were similar for both groups. There was a significant difference between the postoperative RBC transfusion rates in study (0.9 +/- 1.5 RBC units) and control (1.4 +/- 1.8 RBC units) groups (p = 0.005). There was no difference in clinical outcome, including morbidity and mortality rates, in the two groups; group scores for self-assessment of fatigue and anemia were also similar. CONCLUSIONS: A lower Hb threshold of 8 g per dL does not adversely affect patient outcome. Moreover, RBC resources can be saved without increased risk to the patient. PMID- 10532601 TI - Soluble vascular endothelial growth factor in various blood transfusion components. AB - BACKGROUND: Blood transfusion may reduce survival after curative surgery for solid tumors. This may be related to extracellular content of cancer growth factors present in transfusion components. Vascular endothelial growth factor (VEGF) is a potent stimulator of angiogenesis in solid tumors. The potential content of VEGF in various blood components for transfusion was evaluated. STUDY DESIGN AND METHODS: Soluble VEGF (sVEGF, isotype 165) was determined by an enzyme linked immunosorbent assay (EIA) in serum and plasma samples and in lysed cells from healthy volunteers. Subsequently, total content of sVEGF was determined in nonfiltered and prestorage white cell-reduced whole blood (WB), buffy coat depleted saline-adenine-glucose-mannitol (SAGM) blood, platelet-rich plasma (PRP), and buffy coat-derived platelet (BCP) pools obtained from volunteer, healthy blood donors. As a control, total content of platelet-derived soluble plasminogen activator inhibitor type 1 (sPAI-1) was determined by an EIA in the same samples. Finally, the extracellular accumulation of sVEGF was determined in nonfiltered WB and SAGM blood during storage for 35 days and in BCP pools during storage for 7 days. RESULTS: In the healthy volunteers, median total sVEGF content was 97 (range, 20-303) pg per mL in serum and 19 (13-37) pg per mL in plasma (n = 12, p < 0.002) and 445 (280-990) pg per mL in lysed cells. Median total sPAI-1 content was 94 (64-127) ng per mL in serum, 8 (6-11) ng per mL in citrated plasma, and 95 (78-123) ng per mL in lysed cells. In SAGM blood, the median total sVEGF content was 25.3 (3.3-48.4) ng per unit in nonfiltered units and undetectable in white cell-reduced units. Median total sVEGF content was 29.2 (24.8-124.9) ng per unit in nonfiltered PRP and 28.7 (24.5-118.6) ng per unit in white cell-reduced PRP. The sVEGF accumulated significantly in WB, SAGM blood, and BCP pools, depending on the storage time. CONCLUSION: The sVEGF (isotype 165) appears to be present in various blood transfusion components, depending on storage time. PMID- 10532602 TI - Anomaly of the des-Arg9-bradykinin metabolism associated with severe hypotensive reactions during blood transfusions: a preliminary study. AB - BACKGROUND: Severe hypotensive reactions have been described after the transfusion of platelets or red cells through negatively-charged bedside white cell-reduction filters. The possibility of a role for bradykinin (BK) in the genesis of these reactions has been raised. STUDY DESIGN AND METHODS: To understand if an anomaly of BK metabolism is associated with these reactions, the metabolism of BK and des-Arg9-BK was studied in the sera of four patients who presented with a severe hypotensive transfusion reaction. Tests were performed in the absence and the presence of complete in vitro inhibition of angiotensin converting enzyme (ACE) activity by enalaprilat. RESULTS: In the presence of ACE inhibition (enalaprilat), the half-life (t1/2) of BK measured in the sera of patients who presented with a severe hypotensive transfusion reaction (361 +/- 90 sec) was not significantly different from that measured in the sera of normal controls (249 +/- 16 sec). In the presence of ACE inhibition (enalaprilat), the t1/2 of des-Arg9-BK was significantly greater in patients who presented with a severe hypotensive transfusion reaction (1549 +/- 319 sec) than in normal controls (661 +/- 38 sec) (p < 0.001). CONCLUSION: A metabolic anomaly mainly affecting the degradation of des-Arg9-BK could be responsible for its accumulation in vivo. Des-Arg9-BK could be responsible, at least in part, for severe hypotensive transfusion reactions. PMID- 10532603 TI - Immunologic changes after transfusion of autologous or allogeneic buffy coat-poor versus white cell-reduced blood to patients undergoing arthroplasty. I. Proliferative T-cell responses and the balance of helper and suppressor T cells. AB - BACKGROUND: Donor white cells (WBCs) contained in red cell (RBC) transfusions are thought to provoke down-regulation of T-cell-mediated immunity. This study investigated this topic in otherwise healthy patients receiving buffy coat depleted or WBC-filtered RBCs and undergoing standardized perioperative management. STUDY DESIGN AND METHODS: Patients undergoing elective orthopedic surgery (primary hip and knee replacement surgery) were enrolled in a prospective study. Perioperative changes in T-cell proliferation (stimulation with phytohemagglutinin and mixed lymphocyte culture) and T-cell balance (T lymphocytes, helper T cells, and suppressor T cells) were compared after random assignment to allogeneic buffy coat-depleted (Group 2, n = 8) or WBC-reduced RBC (Group 3, n = 11) transfusion regimens. Recipients of autologous buffy coat depleted RBC transfusions (n = 15) served as controls (Group 1). RESULTS: Compared to that in autologous transfusion recipients, alloantigen-induced T-cell proliferation was significantly reduced in recipients of allogeneic WBC-reduced RBCs (Day 3, p = 0.0274). After the transfusion of allogeneic buffy coat-depleted RBCs, a weak trend toward decreased T-cell proliferation was observed (p = 0.0933) and the numbers of CD4+ T cells were also significantly lower (Day 7, p = 0.0389). On Day 10, alloantigen-induced T-cell proliferation remained significantly below baseline after transfusion of WBC-reduced RBCs (p = 0.05), the numbers of CD3+ cells decreased in allogeneic RBC recipients (Group 2, p = 0.078; Group 3, p = 0.05), and those of CD8+ cells decreased significantly after the transfusion of allogeneic buffy coat-depleted RBCs (p = 0.0234) concomitant with an increased CD4:CD8 ratio (p = 0.0391). CONCLUSION: Results of the present study confirm the hypothesis of impaired T-cell-mediated immunity after allogeneic transfusion. PMID- 10532604 TI - Trends in the incidence of delayed hemolytic and delayed serologic transfusion reactions. AB - BACKGROUND: An increasing incidence of delayed hemolytic and delayed serologic transfusion reactions (DHTRs/DSTRs) has been seen at the Mayo Clinic since 1978. Recently, the average length of stay (LOS) for inpatients and the average number of red cell transfusions per inpatient (TPI) decreased, and the albumin and papain technique for RBC antibody detection was replaced by a polyethylene glycol technique. These changes may have affected the incidence of DHTRs/DSTRs. STUDY DESIGN AND METHODS: The diagnoses of DHTR and DSTR made at the Mayo Clinic from 1993 through 1998 were reviewed. These data were compared with previously published Mayo Clinic data from 1980 through 1992. The LOS for inpatients and the average TPI were also obtained from hospital data. RESULTS: The incidence of DHTR/DSTR increased from 1 in 1899 in 1980 through 1992 to 1 in 1300 in the 1993 through 1998 period (p < 0.05). Similarly, DSTR increased from 1 in 2990 in 1980 through 1992 to 1 in 1612 in the 1993 through 1998 period (p < 0.05). The incidence of DHTR showed a trend toward decrease, from 1 in 5405 in 1980 through 1992 to 1 in 6715 in 1993 through 1998. No alloantibody specificities were statistically associated with DHTRs in 1993 through 1998, unlike in the 1980 through 1992 period. Moreover, the incidence of Jka antibodies increased in 1993 through 1998, while the incidence of other alloantibodies remained stable. Average LOS and TPI declined by 24.5 percent and 8.8 percent, respectively, between the two periods. CONCLUSION: Recently, a trend toward a decrease in the incidence of DHTR and a significant increase in DSTRs has occurred at the Mayo Clinic. These changes are most likely due to a combination of factors, including a decrease in average LOS and the adoption of the polyethylene glycol antibody detection system. PMID- 10532605 TI - Efficacy of donor screening for HTLV-I and the natural history of transfusion transmitted infection. AB - BACKGROUND: It has been 10 years since the implementation in Japan of donor blood screening for human T-cell lymphotropic virus type I (HTLV-I). This report reviews the effectiveness of screening in preventing transmission of HTLV-I through blood transfusion and the current status of patients with confirmed seroconversion due to transfusions given before the implementation of screening. STUDY DESIGN AND METHODS: Patients who received blood at Kyushu University Hospital from 1990 to 1997 were followed. Serum samples were collected before transfusion and 60 days or more after transfusion. Seroconversion was determined by a second-generation particle agglutination test. Confirmation tests were an immunofluorescence assay, enzyme-linked immunosorbent assay, and immunoblotting. Confirmed seroconverted patients were followed by a search of hospital records. RESULTS: Seroconversion was found in one of 4672 transfused patients, but the donor was identified and confirmed to be negative for anti-HTLV-I and virus genome by nested polymerase chain reaction. A total of 23,323 red cell concentrates and 17,237 platelet concentrates were transfused to these 4672 patients. Therefore, the anti-HTLV-I prevalence in blood for transfusion after screening was estimated at 1 in 45,560 (0.0022%; the upper 95% CI was 0.0080%). One hundred two seroconverted patients who were transfused before donor screening for HTLV-I were followed. One patient developed HTLV-I-associated myelopathy, diagnosed 18 weeks after seroconversion, and another patient developed uveitis 1 month after seroconversion. No patients developed adult T-cell lymphoma, and the survival rate of seroconverted patients was 92.5 percent 15 years after transfusion. CONCLUSION: This study confirmed that the present donor screening program for HTLV-I by the new particle agglutination test can almost completely prevent virus transmission by transfusion. Complications of HTLV-I transmission were at lower rates than expected. PMID- 10532606 TI - Simultaneous extraction of hepatitis C virus (HCV), hepatitis B virus, and HIV-1 from plasma and detection of HCV RNA by a reverse transcriptase-polymerase chain reaction assay designed for screening pooled units of donated blood. AB - BACKGROUND: Testing for viral nucleic acids should reduce the residual risk of transmitting viral infections by transfusion of blood components. The AmpliScreen Hepatitis C (HCV) Test, Version 2.0, was designed for screening pools composed of samples from individual units of blood or plasma. STUDY DESIGN AND METHODS: An ultracentrifugation step during sample processing simultaneously extracts and concentrates HCV, HIV type 1, and hepatitis B virus particles from plasma. An HCV internal control RNA serves as an extraction and amplification control for each processed sample. Processed samples are amplified by reverse transcriptase polymerase chain reaction and detected by hybridization of the amplified products to HCV- and internal-control-specific oligonucleotide probes. Plasma samples containing known quantities of HCV were used to evaluate analytical sensitivity and precision. RESULTS: The analytical sensitivity of the test was 25 IU of HCV per mL of pooled plasma; all HCV genotypes were detected with similar efficiency. The test did not react with other blood-borne viruses. The within-run and total coefficients of variation were 1.3-13.0 percent and 1.7-22.0 percent, respectively, with low copy samples producing the more variable results. The test performed well using plasma collected in either EDTA, ACD, or PPT Vacutainer tubes. Plasma samples containing elevated levels of hemoglobin, albumin, triglycerides, or bilirubin did not interfere with the test. The test detected HCV RNA 23 to 32 days prior to seroconversion for four of the five seroconversion panels tested. CONCLUSION: The AmpliScreen HCV Test, Version 2.0 provides a reproducible and specific method for screening pooled blood units for HCV. Theoretically, this test has sufficient sensitivity to detect a single infected unit containing 2.4 x 10(3) IU of HCV per mL in a pool with 95 uninfected units and should reduce the window period by at least 20 to 30 days. PMID- 10532608 TI - Analysis of donor return behavior. Retrovirus Epidemiology Donor Study. AB - BACKGROUND: Efforts to provide a safe, adequate blood supply have been inhibited by persistent shortages attributed to a lack of motivation on the part of the general public and inefficiency in recruiting processes. This study examined whether frequency of donations and/or timing of subsequent donations by first time donors related to donor demographics. STUDY DESIGN AND METHODS: Characteristics of 879,816 first-time donors making at least one whole-blood donation were analyzed. Cox proportional-hazards regression models evaluated the first 10 return times separately, and a recurrent-event Cox model was applied to simultaneously evaluate the first five returns. RESULTS: The shorter the donation interval between the first two donations, the more likely the donor was to make subsequent donations. The proportion of repeat donors increased with education level. Rate of donation increased with age and education. The recurrent-event Cox regression model showed that Rh-negative donors, older donors, and donors who had completed college had higher donation return rates. CONCLUSION: Time to return for second donation was associated with total number of donations made and with return rate for subsequent returns. Age was the strongest predictor of high donation frequency and early-return rate. Relationships between interdonation interval and the number of future donations may prove useful in understanding return behavior and developing donor recruitment and retention strategies. PMID- 10532607 TI - A prospective, randomized, sequential, crossover trial of large-volume versus normal-volume leukapheresis procedures: effect on progenitor cells and engraftment. AB - BACKGROUND: The influence of leukapheresis size on the number of harvested peripheral blood progenitor cells is still unclear. A prospective randomized crossover trial was thus performed, to evaluate the effect of large-volume leukapheresis (LVL) versus normal-volume leukapheresis (NVL) on progenitor cells and engraftment in 26 patients with breast cancer and 15 patients with non Hodgkin's lymphoma who were eligible for peripheral blood progenitor cell transplantation. STUDY DESIGN AND METHODS: Patients were randomly assigned to undergo either LVL on Day 1 and on Day 2 or vice versa. The number of progenitor cells was evaluated in the harvest and before and after leukapheresis in the peripheral blood. RESULTS: The number of harvested CD34+ cells (4.8 x 10(6) vs. 3.4 x 10(6)/kg body weight, p < 0.001) and colony-forming units-granulocyte macrophage (3.1 x 10(5) vs. 2.4 x 10(5)/kg body weight, p = 0.0026) was significantly higher for LVL procedures than for NVL procedures. The median extraction efficacy, defined as the difference between the yield in the harvest and the decrease in the total number of CD34+ cells in peripheral blood during leukapheresis, was significantly (p < 0.0001) higher for LVL than for NVL (2.6 x 10(8) and 8 x 10(7), respectively). In patients with breast cancer, the median amount of CD34+ cells in the harvest and the median extraction efficacy were higher for LVL than for NVL (p < 0.0001). This was not found for patients with non-Hodgkin's lymphoma. CONCLUSION: LVL results in a higher yield of CD34+ cells and colony-forming units-granulocyte-macrophage than NVL, but only in patients with breast cancer and with high numbers of CD34+ cells in the peripheral blood before leukapheresis. PMID- 10532609 TI - Effects of granulocyte-colony-stimulating factor on potential normal granulocyte donors. AB - BACKGROUND: The use of granulocyte-colony-stimulating factor (G-CSF) to increase the granulocyte count and the yield from leukapheresis in normal donors is leading to renewed interest in granulocyte transfusion. Therefore, it is important to understand the side effects of G-CSF. STUDY DESIGN AND METHODS: We studied the effect of G-CSF on peripheral blood counts and recorded the side effects experienced 24 hours after an injection of G-CSF in normal subjects donating peripheral blood progenitor cells for research. RESULTS: Following administration of G-CSF to 261 donors, the neutrophil count increased to 20.6 to 24.5 x 10(9) per microL depending on the dose of G-CSF. This represented a 6.2 to 7.4-fold increase over the neutrophil count before G-CSF administration. Of all donors, 69 percent experienced one or more side effects. The most common effects were: muscle and bone pain, headache, fatigue, and nausea. There was a relationship between the dose of G-CSF and the likelihood of experiencing a side effect. Most side effects were mild, but about 75 percent of donors took analgesics because of them. CONCLUSIONS: In a granulocyte donation program involving G-CSF stimulation, about two-thirds of donors would experience one or more side effects, but these would usually be mild and well tolerated. PMID- 10532610 TI - Direct HIV testing in blood donations: variation of the yield with detection threshold and pool size. AB - BACKGROUND: This study was designed to evaluate the potential yield of introducing nucleic acid amplification testing (NAT) in blood donation, according to the detection threshold and the pool size. STUDY DESIGN AND METHODS: A mathematical model of early HIV-1 population dynamics in blood has been developed and is used to predict the window period for NAT, according to the detection threshold and the pool size. The corresponding number of undetected, infected blood donations and the residual risk are estimated by using a previously published simulation model for the United States (9.96 million blood donations from regular donors, and an observed rate of HIV antibody-positive blood donations of 3.18/100,000) and for France (2.32 million blood donations, and a rate of antibody-positive donations of 0.9/100,000). RESULTS: The average window period from infection predicted by the mathematical model for NAT ranges from 8.4 to 15.6 days, according to the detection threshold and the pool size. The maximum yield of adding NAT to the current antibody tests is estimated at 14 donations for the United States and 2 for France. The maximum yield of adding NAT to the newly developed combined HIV antibody and p24 antigen tests is 7 donations for the United States and 1 for France. CONCLUSION: NAT at blood donation could reduce the HIV-1 window period to a minimal value of 8 days without pooling the blood samples, but the yield of NAT would be close to that of combined HIV antibody and p24 antigen tests for high values of the detection threshold and the pool size. PMID- 10532611 TI - Seroindeterminate patterns and seroconversions to human T-lymphotropic virus type I positivity in blood donors from Martinique, French West Indies. AB - BACKGROUND: Screening for human T-lymphotropic virus type I (HTLV-I) antibodies in volunteer blood donors has been systematic in the French West Indies since 1989. Western blot-indeterminate results are commonly obtained. The significance of these indeterminate serologic patterns in HTLV-I-endemic areas is still unclear. STUDY DESIGN AND METHODS: During a 2-year period, 9759 blood donors were tested for HTLV-I antibodies. The epidemiologic features of HTLV-I-seropositive, seroindeterminate, and -seronegative donors were compared. A lookback investigation was performed for the HTLV-I-seropositive donors, and the HTLV-I seroindeterminate individuals were followed up. RESULTS: Thirty-nine donors (0.4%) were HTLV-I seropositive and 49 (0.5%) were seroindeterminate. The age and sex ratio characteristics of the seroindeterminate donors are divergent from those of the HTLV-I-seropositive group and are more like those of the seronegative population. However, during the study period, three cases of seroconversion after an initial seroindeterminate profile were reported. Two cases were detected through follow-up of 38 HTLV-I-seroindeterminate donors over a mean of 8 months (2-24 months). The third seroconverter belonged to the HTLV-I seropositive group and was identified through lookback investigation. This case is atypical, with p19 reactivity for several months before HTLV-I seropositivity. CONCLUSION: These findings indicate that, although HTLV-I-seroindeterminate donors mainly are HTLV-I-noninfected, the rate of seroconversion in a repeat blood donor population from an endemic region must be taken into consideration. Moreover, the case of delayed seroconversion observed in this study suggests the difficulty of counseling seroindeterminate blood donors in endemic regions. PMID- 10532612 TI - Coagulation factor levels in solvent/detergent-treated plasma. PMID- 10532613 TI - Successful management of pregnancy and hemolytic disease of the newborn due to anti-HrO in a woman of the D--phenotype. PMID- 10532614 TI - Estimated risk of transfusion-transmitted HIV infection in Sao Paulo, Brazil. PMID- 10532615 TI - Neurodevelopmental processes in the ontogenesis and epigenesis of psychopathology. AB - Dramatic gains in knowledge have been made in the fields of neuroscience, human development, and developmental psychopathology during the past quarter of a century. Despite the advances that have been achieved in each discipline separately, considerably less progress has occurred in understanding the relation between neurobiological and behavioral development in normal and atypical populations. Research has increasingly demonstrated that abnormalities that occur early in development may result in the emergence of aberrant neural circuitry that eventuates in relatively enduring forms of psychopathology. Knowledge of normal neurobiological development provides a powerful foundation for understanding the contributions that neurodevelopmental processes make to the etiology and sequelae of psychopathology across the life course. An integrated perspective wherein an appreciation of the complex neural, psychological, and social-contextual processes that cohere to bring about normal and pathological outcomes is necessary in order to advance understanding of the genesis and epigenesis of mental disorders. Such an approach will require a reduction of the schisms that so often separate neurobiological and behavioral research. PMID- 10532616 TI - CNS development: an overview. AB - The basic principles of the development of the central nervous system (CNS) are reviewed, and their implications for both normal and abnormal development of the brain are discussed. The goals of this review are (a) to provide a set of concepts to aid in understanding the variety of complex processes that occur during CNS development, (b) to illustrate how these concepts contribute to our knowledge of the normal anatomy of the adult brain, and (c) to provide a basis for understanding how modifications of normal developmental processes by traumatic injury, by environmental or experiential influences, or by genetic variations may lead to modifications in the resultant structure and function of the adult CNS. PMID- 10532617 TI - Cortical plasticity in normal and abnormal cognitive development: evidence and working hypotheses. AB - In this paper I review evidence and viewpoints on developmental plasticity in the cerebral cortex. Although there is some degree of plasticity in the cortex during early postnatal life in the human infant, this plasticity is constrained by various factors. Three working hypotheses about postnatal cortical specialization of function are advanced, and some specific predictions about the limits and extent of plasticity are assessed through both empirical evidence from infants and simulations on simple cortical network models. PMID- 10532618 TI - Neurodevelopmental abnormalities in schizophrenia: insights from neuropathology. AB - Growing epidemiological, genetic, and clinical neurobiological evidence indicates that abnormalities in brain development play determining roles in the pathobiology of schizophrenia. Neuropathological research has made significant progress in delineating cellular and molecular abnormalities in schizophrenia that have relevance to neurodevelopment. This paper reviews the neurodevelopmental processes of neurogenesis, neuronal migration, differentiation, synaptogenesis, neuron and synaptic pruning, and myelination and the reported neuropathological findings in schizophrenia that may be a consequence of disturbances in these processes. While many neuropathological findings in schizophrenia are controversial or await confirmation, reported abnormalities in neuron density, number and morphology, cytoarchitecture, dendritic arbors and spines, synapse-related proteins, and the well-established absence of gliosis or any other evidence of neurodegeneration or neural injury all provide support for the neurodevelopmental model of schizophrenia. PMID- 10532619 TI - Prenatal teratogens and the development of adult mental illness. AB - Our findings in the Helsinki Influenza Study and the Danish Forty Year Study lead us to conclude that a 2nd-trimester maternal influenza infection may increase risk for adult schizophrenia or adult major affective disorder. More recently we have also reported an increase of unipolar depression among offspring who were exposed prenatally to a severe earthquake (7.8 on the Richter scale) in Tangshan, China. Among the earthquake-exposed males (but not the females), we observed a significantly greater depression response for those individuals exposed during the 2nd trimester of gestation. These findings suggest that maternal influenza infection and severe maternal stress may operate (in different ways) as teratogens, disrupting the development of the fetal brain and increasing risk for developing schizophrenia or depression in adulthood. PMID- 10532620 TI - A prospective cohort study of neurodevelopmental processes in the genesis and epigenesis of schizophrenia. AB - A number of lines of evidence converge in implicating neurodevelopmental processes in the etiology and epigenesis of schizophrenia. In this study we used a prospective, longitudinal design to examine whether adverse obstetric experiences predict schizophrenia and whether there is a deviant functional developmental trajectory during the first 7 years of life among individuals who manifest schizophrenia as adults. The 9,236 members of the Philadelphia cohort of the National Collaborative Perinatal Project were screened for mental health service utilization in adulthood, and chart reviews were performed to establish diagnoses according to DSM-IV criteria. The risk for schizophrenia increased linearly with the number of hypoxia-associated obstetric complications but was unrelated to maternal infection during pregnancy or fetal growth retardation. Preschizophrenic cases (and their unaffected siblings who were also cohort members) manifested cognitive impairment, abnormal involuntary movements and coordination deficits, and poor social adjustment during childhood. There was no evidence of intraindividual decline in any domain, but preschizophrenic cases did show deviance on an increasing number of functional indicators with age. Together, these findings suggest that both genetic and obstetric factors participate in creating a neural diathesis to schizophrenia, the phenotypic expressions of which are age dependent, probably reflecting the maturational status of a number of interconnected brain systems. PMID- 10532621 TI - Cognitive and behavioral precursors of schizophrenia. AB - Attentional deficits are well-established characteristics of patients with schizophrenia and their at-risk offspring, suggesting a biological connection between attention and schizophrenia. The goal of this study is to clarify the developmental role of attention in the illness. Data has been collected from 87 subjects at high and low risk for schizophrenia who have participated in the New York High-Risk Project from 1977 to the present. Individuals are considered to be at high risk if either or both of their parents has schizophrenia. Analyses of attention and global behaviors, measured at intervals from about 12 to 26 years of age, indicate (a) attentional deficits can be reliably detected in high-risk children who will develop future schizophrenia-spectrum disorders (the prespectrum [PSP] group); (b) these deficits are stable, enduring over time, and appear to reflect a compromised attentional capacity; (c) attention is not affected by the onset of illness in the PSP group; (d) for all subjects, attention and global behaviors follow independent developmental pathways; and (e) behavioral difficulties, but not attention deficits, appear to be highly sensitive to environmental factors, especially rearing by a mentally ill parent. It is concluded that in PSP individuals impaired attention probably results from prenatal developmental abnormalities (possibly on the cellular level) and is likely to be a marker of a biological vulnerability to schizophrenia. In addition, attentional deficits, as opposed to early behavioral difficulties, are concluded to be a useful first step in screening for youngsters in need of early intervention. PMID- 10532622 TI - Motor dysfunction and risk for schizophrenia. AB - Motor dysfunction is associated with schizophrenia, and recent longitudinal studies indicate that it precedes the onset of clinical symptoms. Of particular interest is the heightened occurrence of involuntary movements, which are apparent as early as infancy and suggest the presence of subcortical brain abnormalities. In this article, we present the results of a study of spontaneous movements in adolescents with schizotypal personality disorder (SPD). SPD is a syndrome that has been shown to be genetically linked with schizophrenia and is often observed prior to the early adult onset of schizophrenia. Systematic coding of videotapes of diagnostic interviews revealed that the SPD group showed significantly more involuntary movements of the head, trunk, and upper limbs than did normals and adolescents with other personality disorders. There were no diagnostic group differences in the rate of voluntary movements. Salivary cortisol, measured before the interview, was positively correlated with involuntary movements. Taken together, the findings provide further support for the hypothesized etiologic relation between SPD and schizophrenia. Based on a neural diathesis-stress model, potential underlying mechanisms are discussed. PMID- 10532623 TI - Brain maturational processes and delayed onset in schizophrenia. AB - The central feature of schizophrenia is its onset in adolescence. Although this clinical observation is consistent with the view that schizophrenia may be a neurodevelopmental disorder, debate has focused on when the proposed brain maturational deviations may begin and what might be the nature of such defective development. Conflicting models of this illness (e.g., the early and late neurodevelopmental models) have been proposed. In this paper, we will first review concepts from basic developmental neurobiology pertinent to these issues; we then summarize aspects of the neurobiology of schizophrenia that have a particular bearing on the adolescent onset of this illness. We propose that the schizophrenic syndrome may result from early brain adversity and late maturational processes of brain development interacting with adverse humoral, biochemical, and psychosocial factors during adolescence and early adulthood. The onset of schizophrenia in adolescence may be related to the "plasticity switch" secondary to the peripubertal brain maturational changes, perhaps involving an alteration in glutamate receptor function. This loss of plasticity could result in social and nonsocial cognitive deficits that are central to the pathophysiology of schizophrenia; the vulnerable person may therefore utilize prepubertal processing styles that are insufficient to the adaptive and "gistful" abstraction requirements of adult cognition. Schizophrenia onset might occur in the context of psychosocial developmental challenges to a delayed social cognitive capacity among neurodevelopmentally compromised individuals. We review therapeutic implications as well as testable predictions generated by this model, and discuss research strategies that might further our understanding of the brain maturational abnormalities in schizophrenia. PMID- 10532624 TI - The effects of neonatal stress on brain development: implications for psychopathology. AB - Recent studies have focused on the behavioral and neurobiological sequella of exposure to early adverse events. We hypothesize that early adverse experiences result in an increased sensitivity to the effects of stress later in life and render an individual vulnerable to stress-related psychiatric disorders. This vulnerability may be mediated by persistent changes in corticotropin-releasing factor (CRF)-containing neurons, the hypothalamic-pituitary-adrenal axis, and the sympathetic nervous system. We therefore present an overview of the CRF system and its role as a mediator in the development of the stress response, major depression, and posttraumatic stress disorder. The literature pertaining to behavioral and neurobiological alterations associated with exposure to early adverse life events in rodents, nonhuman primates, and humans is reviewed. We focus on animal models that precipitate depressive and anxiety symptoms while producing neuroendocrine alterations that mimic those seen in adults with those disorders. The literature integrating neurobiological and behavioral consequences of early life stress is also reviewed, focusing primarily on infants born to mothers with depression and on infants who were abused or neglected. PMID- 10532625 TI - Neurodevelopmental processes and psychological functioning in autism. AB - Autism is a developmental disorder with variable severity, occurring at all levels of cognitive ability and having a number of slightly different clinical presentations. It is associated with neuropsychological deficits that occur in other conditions also, but its pattern may be specific to autism. Genetic and environmental early insults to brain development are etiological determinants of the disorder. Brain circuitries important for social, communicative, and integrational purposes have been suggested to be dysfunctional in autism. There could be at least two different pathways to autism, one connected with primary temporofrontal dysfunction (and late prenatal-early postnatal origins) and another linked to primary brain-stem dysfunction (and early prenatal origins). Further study of neurodevelopmental and neuropsychological processes in autism will help elucidate not only the pathological mechanisms involved in the specific syndromes but also the underpinnings of normal brain development. PMID- 10532627 TI - Developmental neuropsychopathology of attention deficit and impulsiveness. AB - Recent research on the disorders of attention and activity has indicated inherited variants of genes controlling aspects of neurotransmission, abnormalities of structure and function in regions of frontal lobes and basal ganglia, failures to suppress inappropriate responses, and a cascade of failures in various kinds of cognitive performance and organization of behavior. This review integrates the neurodevelopmental findings with findings from developmental psychopathology. It outlines several developmental tracks by which constitutional factors interact with the psychological environment. In one set of tracks, altered brain states lead to cognitive alteration. An understimulating environment is evoked by (and may be genetically associated with) an inattentive and cognitively impulsive style during early childhood. In another track, impulsive and inattentive behavior shows direct continuity through childhood into late adolescence. In yet another track, impulsiveness evokes (and may be genetically associated with) critical expressed emotion from parents and inefficient coping strategies, which in turn contribute to the development of antisocial conduct. This formulation emphasizes the need for several types of research: the mapping of biological findings onto different components of disorder, the combination of genetically informative designs with direct measurement of relevant aspects of the environment, and the use of longitudinal studies to examine predictive and mediating factors separately for different aspects of outcome. PMID- 10532626 TI - Frontal brain electrical activity in infants of depressed and nondepressed mothers: relation to variations in infant behavior. AB - In previous studies, infants of depressed mothers have been found to exhibit reduced left frontal brain electrical activity (EEG). The left frontal region has been hypothesized to mediate social approach behaviors and positive affective expression. These findings raise important questions about the cause and nature of atypical EEG patterns in infants of depressed mothers. The present study begins to address some of these questions by examining whether or not variations in patterns of frontal brain activity in infants of depressed and nondepressed mothers are related to variations in infant behavior as observed in naturalistic situations. If such relations exist, are they specific to certain behaviors hypothesized to be mediated by the frontal region (i.e., positive approach behaviors)? Frontal and parietal brain electrical activity was recorded from 14- to 15-month old infants of depressed versus nondepressed mothers during a baseline condition and during conditions designed to elicit interest and positive affect. Infant behavior was observed in naturalistic play conditions, with and without mother, on a separate day from EEG testing. Mothers provided information on infant temperament. Infants of depressed mothers showed less affection and touching of their mothers. For infants of depressed mothers only, reduced left frontal brain activity was found to be related to lower levels of affection toward mother, but not to infant temperament. Furthermore, increased generalized frontal activation was found to be related to higher levels of negative affect, hostility, and tantrums and aggression. Relations between infant brain activity and behavior were not found for parietal EEG activity. These results suggest that infant frontal electrical brain activity is related to variations in infant behavior, especially those involved in positive affiliative behavior and the expression and regulation of negative affect. The nature and cause of atypical patterns of brain activity and question of whether such atypical patterns of frontal brain activity predispose infants to affective disorders in later life are discussed. PMID- 10532628 TI - Toward an integrated understanding of dyslexia: genetic, neurological, and cognitive mechanisms. AB - This paper reviews what is known about developmental dyslexia at three levels of analysis: cognitive, neurological, and genetic. It also considers the difficult problem of establishing causal links between these levels of analysis, and argues that solving the gene-behavior problem is paradoxically easier than solving the brain-behavior problem. PMID- 10532629 TI - Estrogen reduces leukocyte adhesion in the cerebral circulation of female rats. AB - In this study the authors addressed the hypothesis that estrogen (i.e., 17beta estradiol) acts to repress leukocyte adhesion. The experiments involved comparing leukocyte adhesion in cerebral venules in vivo, in intact ovariectomized and 17beta-estradiol-treated (100 microg/kg/day for 1 week) ovariectomized female rats using topical applications of the adhesion-promoting drug, phorbol 12 myristate 13-acetate (PMA). Adherent Rhodamine-6G-labeled leukocytes were viewed through a closed cranial window using intravital microscopy/videometry. Leukocyte dynamics were recorded at baseline and after each dose of PMA. The PMA was suffused (1.0 mL/min) at increasing concentrations (0.01, 0.1, and 1.0 micromol/L, 15 minutes at each level). A videotape record of each experiment was made for subsequent analysis of leukocyte adhesion. The data showed that the percentage venular area occupied by adherent leukocytes at baseline was significantly greater in the ovariectomized compared to the intact and 17beta estradiol-treated groups (12.2%, 3.4%, and 4.2% respectively). That relationship was maintained during PMA treatments to the extent that the percentage venular area occupied by adherent leukocytes increased to 26.4% in the untreated ovariectomized group compared to 14.4% and 11.3% in the intact and 17beta estradiol-treated groups, respectively. In conclusion, the authors found chronic estrogen depletion enhances leukocyte adhesion in the rat cerebral circulation. Estrogen repletion in such animals is accompanied by a significant reduction in leukocyte adhesion. These findings could, at least in part, account for the ischemic brain damage seen in ovariectomized versus intact females, and the restored neuroprotection observed upon 17beta-estradiol treatment reported in earlier studies. PMID- 10532630 TI - Gender differences in cerebral blood flow and oxygenation response during focal physiologic neural activity. AB - Using functional magnetic resonance imaging techniques CBF and oxygenation changes were measured during sustained checkerboard stimulation in 38 right handed healthy volunteers (18 men and 20 women). The average blood oxygenation level dependent (BOLD) contrast technique signal intensity change was 1.67 +/- 0.6% in the group of male volunteers and 2.15 +/- 0.6% in the group of female volunteers (P < .05). Baseline regional CBF (rCBF) values in activated gray matter areas within the visual cortex were 57 +/- 10 mL x 100 g(-1) x min(-1) in women and 50 +/- 12 mL x 100 g(-1) x min(-1) in men, respectively (P = .09). Despite a broad overlap between both groups the rCBF increase was significantly higher in women compared to men (33 +/- 5 mL x 100 g(-1) x min(-1) versus 28 +/- 4 mL x 100 g(-1) x min(-1), P < .01). The increase of rCBF was not correlated with the baseline rCBF (mL x 100 g(-1) x min(-1)) (r(s) = 0.01, P = .9). Moreover, changes of rCBF were not correlated with changes in BOLD signal intensities (r(s) = 0.1, P = .7). Enhanced rCBF response in women during visual stimulation could be related to gender differences in visual physiology or may reflect gender differences in the vascular response to focal neuronal activation. Gender differences must be considered when interpreting the results of functional magnetic resonance imaging studies. PMID- 10532632 TI - Group II selective metabotropic glutamate receptor agonists and local cerebral glucose use in the rat. AB - The novel mGluR agonist LY354740 and a related analogue LY379268 are selective for mGluR2/3 receptors and are centrally active after systemic administration. In this study, rates of local cerebral glucose use were measured using the [14C]2 deoxyglucose autoradiographic technique to examine the functional consequences of their systemic administration in the conscious rat. Both LY354740 (0.3, 3.0, 30 mg/kg) and LY379268 (0.1, 1.0, 10 mg/kg) produced dose-dependent changes in glucose use. After LY354740 (3.0mg/kg), 4 of the 42 regions measured showed statistically significant changes from vehicle-treated controls: red nuclei ( 16%), mammillary body (-25%), anterior thalamus (-29%), and the superficial layer of the superior colliculus (+50%). An additional 15 regions displayed significant reductions in function-related glucose use (P < .05) in animals treated with LY354740 (30 mg/ kg). LY379268 (0.1, 1.0, 10 mg/kg) produced changes in glucose metabolism in 20% of the brain regions analyzed. Significant increases (P < .05) in glucose use were evident in the following: the superficial layer of the superior colliculus (+81%), locus coeruleus (+57%), genu of the corpus callosum (+31%), cochlear nucleus (+26%), inferior colliculus (+20%), and the molecular layer of the hippocampus (+14%). Three regions displayed significant decreases: mammillary body (-34%), anteroventral thalamic nucleus (-28%), and the lateral habenular nucleus (-24%). These results show the important functional involvement of the limbic system together with the participation of components of different sensory systems in response to the activation of mGluR2 and mGluR3 with LY354740 and LY379268. PMID- 10532631 TI - Postischemic steroid modulation: effects on hippocampal neuronal integrity and synaptic plasticity. AB - Elimination of corticosteroids after ischemia, by removal of the adrenals, has been reported to preserve neuronal integrity later. To establish the therapeutic potential of this observation, the authors address two questions: first, whether clinically more relevant steroid manipulations after ischemia exert similar protective effects, and second, whether changes in synaptic functioning occur along with structural alterations. To test this, the authors treated animals immediately after hypoxia-ischemia with (1) the steroid synthesis inhibitor metyrapone, (2) the synthetic glucocorticoid receptor agonist dexamethasone, (3) the selective glucocorticoid antagonist RU 38486, or (4) corticosterone. Metyrapone, but none of the other compounds, attenuated the occurrence of seizures immediately after ischemia. Twenty-four hours after hypoxia-ischemia, CAI hippocampal field potentials in response to stimulation of Schaffer/commissural fibers were found to be reduced. The attenuation of synaptic transmission was partly prevented by metyrapone. None of the other experimental treatments influenced the impaired synaptic function. Gross morphologic analysis revealed no differences in the loss of neuronal structure between the experimental groups at this time point. Taken together, these data suggest that metyrapone preserves neuronal functioning despite loss of neuronal structure. The authors tentatively conclude that preventing the ongoing production of steroids shortly after ischemia can delay and attenuate the appearance of ischemia-related pathology. PMID- 10532633 TI - Larger anastomoses in angiotensinogen-knockout mice attenuate early metabolic disturbances after middle cerebral artery occlusion. AB - Abnormalities in the homeostasis of the renin-angiotensin system have been implicated in the pathogenesis of vascular disorders, including stroke. The authors investigated whether angiotensinogen (AGN) knockout mice exhibit differences in brain susceptibility to focal ischemia, and whether such differences can be related to special features of the collateral circulation. Wild-type and AGN-knockout mice were submitted to permanent suture occlusion of the middle cerebral artery (MCA). The collateral vascular system was visualized by systemic latex infusion, and the ischemic lesions were identified by cresyl violet staining. The core and penumbra of the evolving infarct were differentiated by bioluminescence and autoradiographic imaging of ATP and protein biosynthesis, respectively. In wild-type mice, mean arterial blood pressure was 95.0 +/- 8.6 mm Hg, and the diameter of fully relaxed anastomotic vessels between the peripheral branches of the anterior and middle cerebral arteries 26.6 +/- 4.0 microm. In AGN knockouts, mean arterial blood pressure was significantly lower, 71.5 +/- 8.5 mm Hg (P < .01), and the anastomotic vessels were significantly larger, 29.4 +/- 4.6 microm (P < .01). One hour after MCA occlusion, AGN-knockout mice exhibited a smaller ischemic core (defined as the region of ATP depletion) but a larger penumbra (the area of disturbed protein synthesis with preserved ATP). At 24 hours after MCA occlusion, this difference disappeared, and histologically visible lesions were of similar size in both strains. The observations show that in AGN-knockout mice the more efficient collateral blood supply delays ischemic injury despite the lower blood pressure. Pharmacologic suppression of angiotensin formation may prolong the therapeutic window for treatment of infarcts. PMID- 10532634 TI - Mice deficient in interleukin-1 converting enzyme are resistant to neonatal hypoxic-ischemic brain damage. AB - Interleukin-1 (IL-1) converting enzyme (ICE) is a cysteine protease that cleaves inactive pro-IL-1beta to active IL-1beta. The pro-inflammatory cytokine IL-1beta is implicated as a mediator of hypoxic-ischemic (HI) brain injury, both in experimental models and in humans. ICE is a member of a family of ICE-like proteases (caspases) that mediate apoptotic cell death in diverse tissues. The authors hypothesized that in neonatal mice with a homozygous deletion of ICE (ICE KO) the severity of brain injury elicited by a focal cerebral HI insult would be reduced, relative to wild-type mice. Paired litters of 9- to 10-day-old ICE-KO and wild-type mice underwent right carotid ligation, followed by 70 or 120 minutes of exposure to 10% O2. In this neonatal model of transient focal cerebral ischemia followed by reperfusion, the duration of hypoxia exposure determines the duration of cerebral ischemia and the severity of tissue damage. Outcome was evaluated 5 or 21 days after lesioning; severity of injury was quantified by morphometric estimation of bilateral cortical, striatal, and dorsal hippocampal volumes. In animals that underwent the moderate HI insult (70-minute hypoxia), damage was attenuated in ICE-KO mice, when evaluated at 5 or 21 days post lesioning. In contrast, in mice that underwent the more severe HI insult (120 minute hypoxia), injury severity was the same in both groups. Reductions in intra HI CBF, measured by laser Doppler flow-metry, and intra- and post-HI temperatures did not differ between groups. These results show that ICE activity contributes to the progression of neonatal HI brain injury in this model. Whether these deleterious effects are mediated by pro-inflammatory actions of IL-1beta and/or by pro-apoptotic mechanisms is an important question for future studies. PMID- 10532635 TI - Expression of tumor necrosis factor-alpha and intercellular adhesion molecule-1 after focal cerebral ischemia in interleukin-1beta converting enzyme deficient mice. AB - Interleukin-1beta (IL-1beta) is expressed after cerebral ischemia and blocking its action reduces subsequent ischemic brain injury. However, the mechanisms by which IL-1beta affects ischemic brain are not understood. To investigate the role of IL- 1beta in regulation of tumor necrosis factor-alpha (TNF-alpha) and intercellular adhesion molecule-1 (ICAM-1) during focal cerebral ischemia, the authors studied mutant mice deficient in the IL-1 converting enzyme (ICE) gene (ICE knockout [KO] mice). Ninety-four adult male ICE KO and wild-type mice underwent 3, 6, 12, and 24 hours of permanent middle cerebral artery occlusion using the suture method. Expression of TNF-alpha and ICAM-1 protein in ischemic brain was examined using immunohistochemistry and Western blot analysis. Neither ICE KO nor wild-type mice had significant differences in CBF and body temperature measurements during the ischemic procedure. TNF-alpha expression increased in the ipsilateral hemisphere after 3 hours of occlusion, peaked at 12 hours and decreased at 24 hours of ischemia in both ICE KO and wild-type mice. ICAM-1 immunohistochemistry showed that the number of ICAM-1-positive vessels in the ischemic hemisphere was reduced in ICE KO mice (P < .05). Western blot analysis showed that ICAM-1 protein expression was significantly attenuated in the ipsilateral hemisphere in the ICE KO mice, which paralleled the immunohistochemistry results. The authors' results indicate that TNF-alpha expression is increased in both ICE KO and wild-type mice suggesting that TNF alpha expression is not related to or upregulated by IL-1beta . ICAM-1 expression is significantly reduced in the ICE KO mice suggesting that IL-1beta plays an important role in the upregulation of adhesion molecules during focal cerebral ischemia. PMID- 10532636 TI - Interleukin-1 receptor antagonist attenuates regional neuronal cell death and cognitive dysfunction after experimental brain injury. AB - The effect of systemic administration of human recombinant interleukin-1 receptor antagonist (rhIL-1ra) on behavioral outcome and histopathologic damage after lateral fluid-percussion brain injury of moderate severity was evaluated. In study 1, brain-injured Sprague Dawley rats received timed subcutaneous injections beginning 15 minutes after injury of either 100 mg/kg rhIL-1ra (high dose, total dose = 1900 mg/kg), 10 mg/kg rhIL-1ra (low dose, total dose = 190 mg/kg), or vehicle over 7 days. No effect of low-dose rhIL-1ra was observed in study 1. High dose rhIL-1ra significantly attenuated posttraumatic neuronal loss in the injured hippocampal CA3 region (P < 0.05), dentate hilus (P < 0.05), and cortex (P < 0.05) but impaired recovery of motor function at 7 days after trauma (P < 0.05). In study 2, rats were pretrained to learn a visuospatial task in a Morris water maze, subjected to fluid-percussion brain injury or sham treatment, and randomly assigned to receive multiple subcutaneous injections at timed intervals of 100 mg/kg rhIL-1ra (total dose = 900 mg/kg) or vehicle over 42 hours, followed by continuous infusion of a lower concentration of rhIL-1ra (20 mg/kg/day, total dose = 100 mg/kg), or vehicle for 5 days using subcutaneously implanted osmotic minipumps. Postinjury administration of rhIL-1ra significantly attenuated cognitive deficits compared with vehicle-treated animals at 42 hours (P < 0.05) but did not affect motor function at 48 hours, 1 week, and 2 weeks. These results suggest that inhibitors of cytokine pathways may be therapeutically useful for the treatment of brain trauma. PMID- 10532637 TI - Survival- and death-promoting events after transient cerebral ischemia: phosphorylation of Akt, release of cytochrome C and Activation of caspase-like proteases. AB - Release of cytochrome c (cyt c) into cytoplasm initiates caspase-mediated apoptosis, whereas activation of Akt kinase by phosphorylation at serine-473 prevents apoptosis in several cell systems. To investigate cell death and cell survival pathways, the authors studied release of cyt c, activation of caspase, and changes in Akt phosphorylation in rat brains subjected to 15 minutes of ischemia followed by varying periods of reperfusion. The authors found by electron microscopic study that a portion of mitochondria was swollen and structurally altered, whereas the cell membrane and nuclei were intact in hippocampal CA1 neurons after 36 hours of reperfusion. In some neurons, the pattern of immunostaining for cyt c changed from a punctuate pattern, likely representing mitochondria, to a more diffuse cytoplasmic localization at 36 and 48 hours of reperfusion as examined by laser-scanning confocal microscopic study. Western blot analysis showed that cyt c was increased in the cytosolic fraction in the hippocampus after 36 and 48 hours of reperfusion. Consistently, caspase-3 like activity was increased in these hippocampal samples. As demonstrated by Western blot using phosphospecific Akt antibody, phosphorylation of Akt at serine 473 in the hippocampal region was highly increased during the first 24 hours but not at 48 hours of reperfusion. The authors conclude that transient cerebral ischemia activates both cell death and cell survival pathways after ischemia. The activation of Akt during the first 24 hours conceivably may be one of the factors responsible for the delay in neuronal death after global ischemia. PMID- 10532638 TI - Spectral analysis of arterial blood pressure and cerebral blood flow velocity during supine rest and orthostasis. AB - This study evaluates the effect of orthostasis on the low frequency (LF, 0.04 to 0.15 Hz) fluctuations in the blood flow velocity of the middle cerebral artery (MCAFV) in relation to its arterial blood pressure (ABP) equivalent to further define and quantify this relationship in cerebrovascular regulation. Spectral analysis was performed on 22 healthy subjects during supine rest and head-up tilt. The power in the LF range can be used to quantify the LF fluctuations, and four types of LF power data could be obtained for each individual: LF power of supine MCAFV, LF power of supine ABP, LF power of tilt MCAFV, and LF power of tilt ABP. By comparing LF power of MCAFV with LF power of ABP, two power ratios could be generated to describe the flow-pressure relationship during supine rest and head-up tilt, respectively, supine power ratio (LF power of supine MCAFV/ LF power of supine ABP) and tilt power ratio (LF power of tilt MCAFV/ LF power of tilt ABP). In addition, an index for dynamic autoregulation in response to orthostasis can be calculated from these two power ratios (tilt power ratio/supine power ratio). The authors found that this index was dependent on the extent of orthostatic MCAFV changes, and the dependency could be mathematically expressed (r = 0.61, P = .0001), suggesting its involvement in cerebrovascular regulation. Moreover, these data further support the previous observation that the LF fluctuations of MCAFV might result from modulation of its ABP equivalent, and the modulation effect could be quantified as the power ratio (LF power of MCAFV/ LF power of ABP). These observations could be an important step toward further insight into cerebrovascular regulation, which warrants more research in the future. PMID- 10532639 TI - A comparison of 11C-labeled L-DOPA and L-fluorodopa as positron emission tomography tracers for the presynaptic dopaminergic system. AB - 11C-labeled 3,4-Dihydroxy-phenyl-L-alanine (L-DOPA) and L-fluorodopa were used as tracers for the functional state of the presynaptic dopamine system in anesthetized monkeys with positron emission tomography. The radiotracer disposition in brain tissue and plasma were studied and effects induced by pharmacologic challenges were evaluated. 6R-L-erythro-5,6,7,8-tetrahydrobiopterin (6R-BH4) increased the striatal influx rate constant, e.g., striatal K(i) for L [beta-11C]DOPA, but it induced no effect on the K(i)-value using L-[beta-11C]-6 fluorodopa. Studies of radiolabeled tracer and metabolites in plasma showed substantial differences between the two tracers. At baseline conditions, 60% unchanged L-[beta-11C]DOPA was detected in plasma 50 minutes after tracer injection and the 3-O-methylated fraction accounted for 25% of total radioactivity. For L-[beta-11C]-6-fluorodopa, the relation was inverse; about 25% unchanged tracer and 60% 3-O-methyl metabolite were present in plasma after 50 minutes. A site-specific 11C-labeling in the carboxylic position in the molecules revealed a significant specific retention of radioactivity in striatum with L [car-boxy-11C]-6-fluorodopa but not with L-[carboxy-11C]DOPA. The 3-O-methyl metabolite of L-DOPA is known to pass the blood-brain barrier and may interfere with the calculation of the K(i)value using a brain reference region. Thus, extensive 3-O-methylation in circulation of the fluorinated analog could obscure the detectability of potential functional change in striatal K(i) of the tracer when using a reference tissue model for calculation. PMID- 10532640 TI - Kinetic modeling of N-[11C]methylpiperidin-4-yl propionate: alternatives for analysis of an irreversible positron emission tomography trace for measurement of acetylcholinesterase activity in human brain. AB - N-[11C]Methylpiperidin-4-yl propionate ([11C]PMP) is a substrate for hydrolysis by acetylcholinesterase (AChE). This work evaluates kinetic analysis alternatives for estimation of relative AChE activity using dynamic positron emission tomography (PET) studies of [11C]PMP. The PET studies were performed on three groups of subjects: (1) 12 normal volunteer subjects, aged 20 to 45 years, who received a single intravenous injection of 16 to 32 mCi of [11C]PMP; (2) six subjects, aged 21 to 44 years, who received two 16-mCi injections of [11C]PMP (baseline and visual stimulation, respectively); and (3) five subjects, aged 24 to 40 years, who received two 16-mCi injections separated by 200 minutes (baseline and after a 1-hour constant infusion of 1.5 mg of physostigmine, respectively). Dynamic acquisition consisted of a 17-frame sequence over 80 minutes. All analysis methods were based on a first-order kinetic model consisting of two tissue compartments with the parameter k3, representing PMP hydrolysis, being the index of AChE activity. Four different schemes were used to estimate k3: (1) an unconstrained non-linear least-squares fit estimating blood brain barrier transport parameters, K1 and k2, in addition to the hydrolysis rate constant k3; (2) and (3), two methods of constraining the fit by fixing the volume of distribution of free tracer (DVfree); and (4), a direct estimation of k3 without use of an arterial input function based on the shape of the tissue time-activity curve alone. Results showed that k3 values from the unconstrained fitting and no input methods were estimated with similar accuracy, whereas the two methods using DVfree constraints yielded similar results. The authors conclude that the optimal analysis method for [11C]PMP differs as a function of AChE activity. All four methods gave precise measures of k3 in regions with low AChE activity (approximately 10% coefficient of variation in cortex), but surprisingly, with unconstrained methods yielding estimates with lower variability than constrained methods. In regions with moderate to high AChE activity, constrained methods were required to yield meaningful estimates and were superior to the unconstrained methods. PMID- 10532641 TI - Quantification of [11C]FLB 457 binding to extrastriatal dopamine receptors in the human brain. AB - Positron emission tomography (PET) has hitherto been used to examine D2 dopamine receptor binding in the striatum, a region with a high density of receptors. Research has been hampered by the lack of suitable radioligands for detection of the low-density D2 dopamine receptor populations in the limbic and cortical dopamine systems that are implicated in the pathophysiology of schizophrenia. [11C]FLB 457 is a new radioligand with the very high affinity of 20 pmol/L (K(i)) for the D2 and D3 dopamine receptor subtypes. This study in eight healthy subjects was designed to evaluate the suitability of [11C]FLB 457 for quantification of extrastriatal D2/D3 dopamine receptors. PET-data were acquired in the three-dimensional mode and the arterial input function was corrected for labeled metabolites. The standard three-compartment model and four derived approaches were applied to calculate and compare the binding potentials. Besides the striatum, conspicuous radioactivity was found in extrastriatal regions such as the thalamus, the anterior cinguli, and the temporal and frontal cortices. The time activity curves could be described by the three compartment model. The different approaches gave similar binding potential values and the rank order between regions was consistent with that found in vitro. The short time of a PET measurement using [11C]FLB 457 (63 minutes) seemed not to be sufficient for reliable determination of the high binding potential in the striatum. These results are of principal importance because they show the potential for PET quantification of minute receptor populations in the human brain. PMID- 10532642 TI - Advances in stroke management: update 1998. PMID- 10532643 TI - Thrombolysis in acute ischemic stroke: controlled trials and clinical experience. AB - Thrombolytic therapy with recombinant tissue plasminogen activator (rtPA) is approved in the United States for treatment of acute ischemic stroke. Approval was granted after a large, randomized, placebo-controlled study by the National Institute of Neurological Disorders and Stroke (NINDS) showed a significant improvement in 3-month outcomes with rtPA despite a significant risk for symptomatic hemorrhage. Two other trials, the first and second European Cooperative Acute Stroke Study (ECASS I and II), have shown comparable results, but neither was statistically positive for the predefined primary end point. An analysis of the risk/benefit profile of rtPA therapy based on the results of these three trials indicates that the treatment is effective and, when administered within 3 hours of symptom onset at a dose of 0.9 mg/kg, the benefits by far outweigh the risks for eligible patients. Even with the 6-hour time window of the two ECASS trials, a combined analysis of the three studies shows the number of disabled or dead patients to be significantly reduced. Preliminary data collected on the use of rtPA outside of clinical trials in the United States and Europe suggest that, when rtPA is used according to the trial protocol, the risks and benefits are similar to those observed in clinical trials. However, even within the United States, rtPA is underutilized. The most substantial treatment barrier is the narrow time window, which may be expanded if long-term experience shows that this is possible. Most stroke patients arrive at the hospital too late to be eligible for screening and treatment. Education of the public and physicians may help to overcome this difficulty. PMID- 10532644 TI - Risk factors and their management for stroke prevention: outlook for 1999 and beyond. AB - Stroke is a leading cause of death and morbidity, but incidence rates vary dramatically from one population to another. The reasons for this heterogeneity are being explored in several large-scale epidemiologic studies around the world. Much of the heterogeneity in stroke can be related to the prevalence of risk factors, but some populations have a higher stroke incidence than would be predicted from risk factor levels. Hypertension, including borderline hypertension, is probably the most important stroke risk factor based on degree of risk and prevalence. However, cardiac morbidity, cigarette smoking, diabetes, physical inactivity, and high levels of alcohol consumption are also strongly related to stroke risk. High levels of blood cholesterol and homocysteine may also increase stroke risk. Mortality after stroke is highest within the first 30 days but remains elevated to a degree that depends on the presenting stroke syndrome, stroke subtype, and other co-morbidities. Lacunar strokes have the best short- and long-term prognoses. Strokes due to large-vessel atherosclerosis frequently worsen; these and cardioembolic strokes have the poorest long-term prognosis. The risk for recurrence is also highest within 30 days after a first stroke, depending on the type of infarct, history of hypertension, and blood glucose levels on admission. Predictors of late recurrence include cardiac disease, hypertension, and heavy alcohol use. Only about half of stroke survivors are independent 6 months after a stroke, and quality of life is decreased. Understanding factors that predispose to stroke and determine its outcome will help in the design of acute stroke trials and in prevention programs. PMID- 10532645 TI - Antiplatelet therapy: new foundations for optimal treatment decisions. AB - Individuals who experience a stroke or a transient ischemic attack require long term treatment to prevent a subsequent stroke. According to the current guidelines, patients with a first cerebrovascular event due to cardioembolism should be treated with oral anticoagulants, barring any contraindications. Individuals with ischemic cerebral events due to atherothrombosis should typically receive antiplatelet agents. Aspirin is the best-studied antiplatelet agent and has been used in stroke prevention for many years. Trials evaluating aspirin have, over time, enrolled more patients and tested lower aspirin doses. No individual trial conducted in cerebrovascular patients has established the optimal aspirin dose for prevention of vascular events, but meta-analyses of trials at different dose ranges and the two single trials that directly compared different doses strongly suggest that the benefit of aspirin is independent of dose in this patient population. Lower doses (50-325 mg daily) are now recommended because of their more favorable side-effect profiles. Because its value is established, aspirin has been used as a control to evaluate other antiplatelet agents. On the basis of large clinical trials versus aspirin, three other antiplatelet agents (ticlopidine, clopidogrel, and the combination of aspirin plus extended-release dipyridamole) have all been shown to be effective for stroke prevention. Physician opinions regarding the efficacy of these agents in indirect comparisons and the differences in their safety profiles, availability, and cost will influence the choice of agent for the individual patient. PMID- 10532646 TI - Antiplatelet therapy: views from the experts. AB - Antiplatelet therapy is recommended for stroke prevention in persons with a history of thromboembolic stroke or transient ischemic attack (TIA) that is not of cardiac origin. Aspirin was the first antiplatelet agent to be used in this context and is still the most frequently prescribed preventive treatment for ischemic stroke. However, because the results of clinical studies with aspirin have been inconsistent, the dose of aspirin required for stroke prevention in persons with cerebrovascular disease has been a subject of debate among stroke neurologists. In the present discussion, low-dose aspirin is generally regarded by the experts as equivalent in effectiveness to high-dose aspirin, and its higher tolerability has the potential to significantly increase compliance with long-term therapy. Higher aspirin doses may have clinical utility in particular settings, but this requires further study. Despite the controversy, aspirin is now recognized as the treatment standard against which other antiplatelet agents are compared. Antiplatelet agents that may be more effective than aspirin have now been developed. Although each of these agents has been directly compared with aspirin in a large, randomized clinical trial, the lack of direct comparisons among these alternative antiplatelet therapies complicates decisions regarding long-term care of patients with cerebrovascular disease. An international panel of stroke neurologists reports that their selection of antiplatelet therapies for patients with prior history of TIA or stroke depends most heavily on drug efficacy and safety issues and is limited by availability (approval status of alternatives). PMID- 10532647 TI - A fatal relationship--influenza virus interactions with the host cell. AB - Influenza A viruses are important worldwide pathogens for humans and different animal species. The infectious agent is the prototype of the orthomyxoviridae which are characterized by a segmented negative strand RNA genome that is replicated in the nucleus of the infected cell. The genome has a combined coding capacity of about 13 kb and contains the genetic information for ten viral proteins. Despite this relatively small coding capacity--large DNA viruses like herpes or poxviruses express about 150-200 gene products--influenza A viruses are able to successfully infect and multiply in a wide range of mammalian and avian species. It is therefore not surprising that influenza A viruses extensively use and manipulate host cell functions. This includes multiple interactions of viral proteins with cellular proteins. In recent years an increasing amount of information about the identity of the cellular factors that are involved in viral transcription and replication, intracellular trafficking of viral components and assembly of the virus particle has accumulated. This article aims to review recent developments in this field with a focus on cellular factors and processes which are activated by the virus to either support viral replication or to counteract host-cell defense mechanisms. PMID- 10532648 TI - Plasma lactoferrin levels are decreased in end-stage AIDS patients. AB - The antimicrobial protein lactoferrin (Lf) is present in plasma and in mucosal secretions. Using ELISA we analysed plasma and saliva of HIV-infected patients, patients with AIDS, and healthy controls for the presence of secreted Lf. The plasma Lf levels of AIDS patients (classification C3) were significantly lower (p < 0.001) as compared to asymptomatic and symptomatic HIV infected patients, or controls. In addition, plasma Lf levels closely correlated with neutrophilic granulocyte counts in the HIV-infected patients. Thus, basal plasma Lf levels are likely the result of Lf release by neutrophilic granulocytes. The Candida titres present in the oral cavity were determined in a part of the HIV-infected patient group. As it appeared, the presence of this opportunistic pathogen always coincided with low levels of salivary Lf levels. We conclude that Lf, as part of the nonspecific immune system, might play an important role in the first line of defense against opportunistic microbial infections in AIDS patients. PMID- 10532649 TI - Immunogenicity of full length and truncated SIV envelope proteins. AB - We have compared the immunogenicity of the full-length (FL) SIV envelope (Env) protein and a truncated (T) form of the Env protein which has a short cytoplasmic tail. The Env(T) protein was previously shown to be more fusogenic than Env(FL), has a higher level of incorporation into virus-like particles (VLPs) and membrane vesicles, and expands the viral host range. We have found that mice immunized with VLPs which contained an equal amount of Env(FL) or Env(T) produced similar titres of neutralizing antibody. Comparison of immune responses between animals that received DNA vaccines encoding Env(T) vs. Env (FL) by epidermal delivery demonstrated that a higher level of specific antibody was elicited by Env(T) than Env(FL). This result correlated with a higher level of expression of pCMVEnv(T) than pCMVEnv(FL) observed in vitro. DNA immunization combined with VLP boosting elicited a similar level of neutralizing antibody with both forms of Env proteins. These data indicate that the immunogenicity of Env(FL) and Env(T) is similar, and that either form of Env protein appears to be potentially suitable for use in further development of vaccine preparations. PMID- 10532651 TI - Differential neutralizing antibody responses to varicella-zoster virus glycoproteins B and E following naked DNA immunization. AB - The only available vaccine against varicella-zoster virus (VZV) consists of the VZV-Oka attenuated but persistent virus strain. Development of a safer, subunit vaccine is therefore desirable. In this prospect, nucleic acid vaccines, expressing truncated forms of VZV glycoproteins B (recgB) and E (recgE) from which the anchor and the cytoplasmic domains were deleted, were used to immunize mice. Vaccination with recgB encoding plasmid elicited a strong and specific humoral immune response. Total IgG and neutralizing titres were comparable to those previously obtained by vaccination with purified and adjuvanted native recgB. In contrast, mice immunization with recgE encoding plasmid only induced a very weak immune response whereas we previously showed that vaccination with adjuvanted native or denatured recgE protein led to high neutralizing titres. The weakness of the immune response induced by recgE-encoding plasmid depended neither on the deletion of the anchor domain in the gE gene nor on the animal model. Analysis of antibody isotypes produced by plasmid immunizations revealed a response slightly dominated by IgG2a. Taken together, the data indicate that a VZV subunit vaccine based on adjuvanted recombinant glycoprotein E is more promising than a nucleic acid-based vaccine strategy. As regards recgB, both vaccination approaches might be appropriate. PMID- 10532650 TI - Plasmid expressing the influenza HA gene protects old mice from lethal challenge with influenza virus. AB - Virus-based influenza vaccines induce less protection in old compared to young subjects due, in part, to age-associated alterations in the immune response. This study shows that old mice produce a less diverse HI antibody response after immunization than adult mice. However, immunization of old and young mice with plasmids expressing the HA gene induced comparable clearance of influenza virus from the lungs and the same level of protection from a lethal challenge with live WSN influenza virus. Thus, genetic immunization may offer advantages for the elderly over virus-base vaccines. PMID- 10532652 TI - Increased neoplasm development due to immunosuppressive treatment with FK-506 in BALB/C mice persistently infected with the mouse herpesvirus (MHV-72). AB - BALB/c mice were infected with the lymphotropic mouse gammaherpesvirus (MHV-72). At late (7-12 months) post-infection intervals the latent virus was detected in the cells of lymphatic system (peripheral blood, lymphocytes and macrophages, thymocytes, lymph nodes, bone marrow, and peritoneal macrophages,) and in the spleen, lungs, liver, and kidney by cocultivation as well as by explantation. The MHV-72 infected mice, in which latency had been established, were treated with the immunosuppressive (IS) drug FK-506 (2 mg/kg/mouse for 30 days). This treatment increased the probability of virus reactivation by over two-fold. During the post-treatment observation period of 19 months, the incidence of lymphomas and the development of MHV-related lymphoproliferative and hemoblastic disorders raised to nearly five-fold in the drug treated mice as compared to untreated animals. PMID- 10532653 TI - Constitutive expression of human cytomegalovirus (HCMV) glycoprotein gpUL75 (gH) in astrocytoma cells: a study of the specific humoral immune response. AB - The humoral immune response to gpUL75 (gH) was determined in different groups of human cytomegalovirus (HCMV) infected subjects using a full-length glycoprotein constitutively expressed in an astrocytoma cell line. The recombinant molecule consisted of two distinct isoforms resembling the authentic protein of infected cells. Separated from the interactions of other viral gene products gH failed to form an oligomeric complex, thus exhibiting exclusively epitopes present on the monomer. Ninety five percent of serum samples from latently-infected healthy adults revealed the presence of gH-specific IgG. Moreover, examination of sequential sera from immunocompromised and immunocompetent individuals undergoing active HCMV infection demonstrated that antibodies to gH occurred in most cases simultaneously with those to the abundant surface antigen gpUL55 (gB) and at similar titres. Appearance of this response was correlated with a considerable increase of the virus-neutralizing activity and most likely associated with restriction of viral dissemination during subsequent viremic episodes. Together, these results suggest that glycoprotein H of HCMV is like gB, a highly immunogenic component of the infectious particle. PMID- 10532654 TI - Herpes simplex virus type 2 infection induced apoptosis in peritoneal macrophages independent of Fas and tumor necrosis factor-receptor signaling. AB - Freshly isolated macrophages from mature mice are poorly or nonpermissive for infections with HSV. However, despite lack of significant viral replication, HSV infection has been demonstrated to induce substantial cell death among macrophages. To determine if HSV-induced cytotoxicity of macrophages is due to apoptosis, peritoneal macrophages were obtained from C57BL/6 (B6) mice, and apoptosis was analyzed following HSV-2 infection in vitro. Macrophages underwent apoptosis upon HSV-2 infection indicated by annexin V staining, labeling of DNA strand breaks and electronmicroscopy. Apoptosis was associated with macrophage activation demonstrated by upregulation of MHC class II and Mac-1 surface expression. Though there was also an upregulation of Fas (Apo-1/CD95) and tumor necrosis factor (TNF)-receptor 1 (TNF-R1) pathways, inhibition of Fas by soluble Fas and blocking of TNF-alpha using a TNF-binding protein did not prevent HSV induced apoptosis. Moreover, apoptosis was not impaired in HSV-2 infected macrophages from Fas-deficient B6-lpr/lpr mice suggesting involvement of other apoptosis pathways, or activation of Fas or TNF-R pathways downstream of the receptor level. The present results demonstrate that HSV-2 infection leads to activation and subsequent apoptosis in peritoneal macrophages independent of Fas or TNF-R1 signaling. PMID- 10532655 TI - Childhood accidental spiral tibial (CAST) fractures. AB - OBJECTIVE: To further define and describe the spectrum of presentations for accidental spiral tibial fractures of childhood. DESIGN: A retrospective review. METHODS: Children 8 years of age or younger who had sustained a tibial fracture within the last five or ten years were collected from the patient populations of two large tertiary medical centers in Southern California, Riverside General Hospital (RGH) and Loma Linda University Medical Center (LLUMC). A total of 189 tibial fractures were documented from both locations. Of the 189 patients, the 55 children with isolated spiral tibial fractures and no criteria for exclusion were selected for further review and analysis. These patients were reviewed for age at time of injury, gender, specific extremity involved, mechanism of injury, fracture location, degree of displacement, and whether child protective services involvement occurred. RESULTS: Patients with isolated spiral tibial fractures ranged in ages from 12 months to 94 months (7 years 10 months). The mean age was 50.7 months. Eighteen (32.7%) of the patients were less than or equal to 36 months of age. No patient was under one year of age. Males (38/55 or 69%) sustained the fracture slightly more frequently than females. The right extremity was injured slightly less frequently (45.5%) than the left extremity (54.5%). Overall, the lower two thirds of the tibia contained the fracture in 95% of the injuries. Displacement of the fracture segments was most frequently none or minimal. While Child Protective Service referrals or investigations were not accomplished on the majority of the children, no injury was confirmed to have occurred as a result of non-accidental trauma. CONCLUSION: Isolated spiral tibial fractures are a common injury of children less than 8 years of age and are most frequently accidental. The original description of a distinct clinical entity matching the original definition of the toddler's fracture does not appear to exist. Instead, the previously defined toddler's fracture is simply part of a spectrum of presentations of childhood accidental spiral tibial, or CAST, fractures. Consequently, our findings further support new nomenclature suggested for this fracture (1, 2). PMID- 10532656 TI - Factors associated with significant injuries in youth ice hockey players. AB - STUDY OBJECTIVE: To assess the implementation of published injury prevention strategies in injured youth ice hockey players, to examine factors contributing to current youth ice hockey injuries, and to assess attitudes of participants toward injury-risk activities. METHODS: Case series describing a convenience sample of 103 children presenting to a children's hospital emergency department with an injury sustained playing youth ice hockey. Using a questionnaire, patients self-reported their compliance with protective equipment guidelines, the circumstances of injury, and their attitudes toward risk-taking in youth ice hockey. RESULTS: A total of 103 patients suffered 113 injuries. For each piece of required equipment, compliance approached 100%. Penalties were assigned on 4% of plays causing injury. An additional 36% of patients injured during game play felt that a penalty should have been assigned. Fifty-seven percent of injuries were caused by checking. Fifty-eight percent of injuries caused by checking met our criteria for significant injury. Significant injury was more likely when initiating or receiving a check perceived to be legal than when receiving a check perceived as illegal (P = 0.032). Twenty-four percent of patients stated spinal cord injury and 45% stated brain injury was not possible given their usual protective equipment. To win, 32% of patients stated that they would check illegally, and 6% stated that they would purposely injure. CONCLUSION: While compliance with protective equipment requirements was good, rule enforcement was perceived to be inadequate. Elimination of checking would potentially reduce the number of significant injuries more than would the enforcement of existing rules. Injured youth hockey players are underinformed about the hazards of their sport and are too willing to engage in potentially injurious activities. PMID- 10532657 TI - Management of children with aseptic meningitis in the emergency department. AB - OBJECTIVE: To review the emergency department management of children with aseptic meningitis and compare the clinical features, laboratory findings, and short-term follow-up of those who were hospitalized or discharged to determine guidelines for discharge. DESIGN: Retrospective chart review study. SETTING: Emergency department of an inner-city teaching, level III, children's hospital during an outbreak of aseptic meningitis from, March through December 1993. METHODS: The medical records of children < or = 18 years of age diagnosed in the emergency department with aseptic meningitis after lumbar puncture were retrospectively reviewed and analyzed. Thirty-four parameters were recorded for each child including demographic (2), epidemiologic (5), clinical (2), laboratory (10), and short-term follow-up data (5). RESULTS: Of the 158 eligible patients, 99 (62.7%) were hospitalized, and 59 (37.3%) were discharged. Compared to the hospitalized group, children who were discharged were significantly older (5.7 years vs. 4.7 years, P < 0.05) and experienced a more benign course, with lower rates of headache (54.7 vs. 85.7%, P < 0.05), vomiting (38.2 vs. 69.7%, P < 0.05), and irritability (1.8 vs. 8.1%, P < 0.05). They also had significantly (P < 0.05) lower mean peripheral and cerebrospinal fluid leukocyte counts (13,233 vs. 11,498/mm3 and 293.91 vs. 105.29/mm3, respectively). Interestingly, 30 (50.8%) of children in the discharged group had over 50% polymorphonuclears in their cerebrospinal fluid (CSF) cell count. The hospitalization rate during the day was significantly (P < 0.05) lower than that for the evening and night shifts (51.5 vs. 66.7%, respectively). In the discharged group, symptoms of headache, fever, and vomiting resolved after an average of 3.05 days, 2.25 days, and 1.3 days, respectively. The average hospitalization time was 3.5 days. There were no significant complications in either group. More important, in neither group were there any misdiagnoses of bacterial meningitis as aseptic meningitis. During the study period, the ambulatory management of the 59 patients cost $51,625 less than the hospitalization of an equal number of children. CONCLUSION: It is feasible, clinically safe and less costly to treat a subgroup of children with aseptic meningitis in an ambulatory setting. Although absolute criteria for ambulatory follow-up could not be defined, age >1 year, a nontoxic clinical appearance, normal white blood cell count, mild cerebrospinal fluid pleocytosis (even with a high percentage of polymorphonuclear cells), negative CSF Gram stain, and a reliable family setting could serve as guidelines for decision-making regarding emergency department discharge. Further prospective research is needed to better specify these criteria. PMID- 10532658 TI - Insertion of long lines in the pediatric emergency department. AB - OBJECTIVE: The purpose of this study was: 1) to evaluate the role of the pediatric emergency department (PED) in placing peripherally inserted midline or central catheters (long lines), and 2) to review indications and complications to use this technology to reduce the number and duration of admissions and provide an alternative method for administering intravenous therapy. METHODS: Retrospective chart review of all patients taken from a procedure log who had long lines placed in the emergency department of a children's hospital. RESULTS: Twenty-eight patients had 30 long line insertions over a 36-month period. Fourteen were female; age ranged from 1 to 36 years with a median of 9 and a mean of 11.1 +/- 8.4. The indication for insertion was for parenteral antibiotics in 27 of 28 (96%) patients and for parenteral nutrition in 1 (4%) patient. The catheters varied in length from 8 to 60 cm. Twelve of 30 (40%) catheters terminated centrally in the subclavian or superior vena cava, while 18 (60%) were in the peripheral cephalic, basilic, or axillary veins. Chest radiography confirmed positioning in 12 of 12 inserted centrally and in 15 of 18 (83%) in the peripheral circulation. Half of the patients received no premedication for the procedure; 10 (33%) received topical anesthetic cream; 2 (7%) local infiltration of anesthetic, and 2 (7%) parenteral sedation. Twenty-one of 30 (70%) patients were discharged directly from the emergency department; 3 (10%) were discharged after admission to the hospital to complete treatment at home with their long lines, and 6 (20%) used their long lines for in-hospital therapy only. Eight of 30 (27%) placements were for patients specifically referred to the PED for placement or replacement of a long line. Twelve of 30 (40%) lines were placed in children presenting for intravenous therapy for cellulitis. These patients received a long line with home i.v. therapy instead of the traditional admission. The duration of intravenous treatment documented for all patients ranged from 1 to 62 days with a median of 10.5, and a mean of 16.2 +/- 17.8, compared with the duration of the line ranging from 1 to 28 days with a median of 4, and a mean of 7.3 +/- 8.0. Ten of 30 (33%) had their line for 3 days or less. The short duration was due to problems with line function in 5 of 10, and intentional removal secondary to improved cellulitis in 5. There were no significant complications with the lines reported during placement or while in use; however, 8 of 30 (27%) of the lines placed developed problems with function, requiring repair or replacement. CONCLUSIONS: 1) Long lines can be inserted in the pediatric emergency department by physicians with different levels of training with minor complications and no adverse clinical effects; 2) the placement of long lines can eliminate the need for hospitalization in some cases, reduce the duration of hospitalization in others, and lessen the need for repeated venipunctures for routine peripheral catheter replacement in patients requiring i.v. therapy; 3) the planned duration of therapy as well as other factors not analyzed in this study should be considered when selecting patients for long line placement in the emergency department. PMID- 10532659 TI - Bilateral localized tension pneumothoraces refractory to needle decompression. AB - We present the unusual case of a 12-year-old child with bilateral localized tension pneumothoraces that were initially both difficult to diagnose and refractory to needle decompression. This case illustrates several important variations in the diagnosis and treatment of tension pneumothorax. To the best of our knowledge, it also represents the only reported case of bilateral localized tension pneumothoraces presenting in the pediatric age group. PMID- 10532660 TI - Cyproheptadine for serotonin syndrome in an accidental pediatric sertraline ingestion. PMID- 10532661 TI - Intraosseous infusion of iodinated contrast in an abused child. PMID- 10532662 TI - Serous fluid leakage after a modified Blalock-Taussig shunt: a cause of hypercyanotic episodes. AB - We report a case of a 10-week-old girl, with tetralogy of Fallot and a Blalock Taussig shunt, who presented with hypercyanotic episodes. She was found to have serous fluid leakage around her shunt, causing compression of her trachea. Hypercyanotic episodes resulting from shunt leakage have not previously been reported. An awareness of this possible complication of a Blalock-Taussig shunt will allow the emergency physician to consider it in the differential diagnosis of hypercyanotic episodes. PMID- 10532663 TI - Tubulointerstitial renal failure in childhood leptospirosis. AB - We report three children with tubulointerstitial renal failure following leptospirosis. All had acute nonoliguric renal failure with mild hypocalemia and mild metabolic acidosis. Maximum blood urea nitrogen (BUN) and creatinine were 217 and 7.1 mg/dl, respectively, on the 6th day of disease, and no patient required dialysis. They presented with acute febrile illness and dehydration, and required intravenous fluid supplements. Myalgia, vomiting, and bleeding were found in two children. Abdominal pain, arthralgia, diarrhea, and conjunctival suffusion were found in one child. Only one child, who had an underlying disease of beta-thalassemia/Hb E, had jaundice, hepatosplenomegaly, anemia, and thrombocytopenia. Penicillin treatment was given in one case. All recovered, with normal renal function. The leptospirosis complement fixation test was used to confirm diagnosis. L. batavia was considered the etiologic agent in two of the children. PMID- 10532664 TI - Hematuria in two school-age refugee brothers from Africa. PMID- 10532665 TI - Detection of group A streptococci in children under 3 years of age with pharyngitis. AB - OBJECTIVE: To determine the frequency of group A streptococcal pharyngitis in young preschool children presenting to the emergency department with upper respiratory tract infection. METHODS: A prospective, observational study performed between September 1995 and September 1997. Throat swabs were obtained on young children less than 3 years old with pharyngeal erythema as well as age- and time-matched controls without pharyngeal erythema or exudate. Signs and symptoms that were recorded included: age, temperature, pharyngeal erythema, tonsillar exudate, cervical adenopathy, scarlatini-form rash, rhinorrhea, school aged child in the home, day care attendance. Swabs were inoculated on 5% sheep blood agar and incubated for 48 hours. Beta-hemolytic colonies were sero-grouped by latex agglutination. RESULTS: Seventy-eight children with pharyngeal erythema, and 152 controls had pharyngeal specimens obtained and signs or symptoms recorded. Under 2 years of age, the detection of group A streptococci was similar to controls. Detection of group A streptococci was significantly different from controls in children over 2 years of age. Ten (29%) of 35 children over 2 years were positive for group A streptococci compared to 2 (7%) of 29 controls of the same age group (P = 0.03, odds ratio 5, 95% CI: 1.2-24). Findings on clinical examination in children with pharyngeal erythema did not distinguish those that would be culture-positive for group A streptococci. CONCLUSION: In our emergency department, group A streptococci caused 30% of pharyngitis seen in children between 2 and 3 years of age. Diagnostic testing is recommended because physical examination may not accurately distinguish etiology in this age group. PMID- 10532667 TI - An unusual case of acetaminophen overdose. PMID- 10532666 TI - Descending suppurative mediastinitis: nonsurgical approach to this unusual complication of retropharyngeal abscesses in childhood. AB - OBJECTIVE: To alert the pediatric emergency physician about suppurative mediastinitis as an unusual, life-threatening complication of retropharyngeal abscesses in children and to report an alternative therapeutic option for these cases. METHODS: We describe a case of suppurative mediastinitis secondary to a retropharyngeal abscess in a 19-month-old girl and discuss the pathophysiology, diagnosis, and treatment of this disease. RESULTS: Prompt diagnosis, based on clinical, radiographic, and CT findings, followed by immediate retropharyngeal drainage and appropriate antibiotic therapy, allowed conservative management of the mediastinal abscess, without the need for surgery. The child presented a good outcome and was discharged on hospital day 14. CONCLUSIONS: When evaluating a retropharyngeal abscess, the pediatric emergency physician should be aware of its complications. A chest radiograph should be prescribed for each patient presenting with an indolent course. Widening of the mediastinum should be considered as strong evidence of a mediastinal abscess for which the best therapeutic option is aggressive surgical drainage. In the rare cases in which marked improvement is achieved after retropharyngeal drainage, a nonsurgical approach to the mediastinal abscess could be attempted. CT scan and a simple chest radiograph have proved to be useful for diagnosis and follow-up. PMID- 10532668 TI - Posttraumatic subgaleal hematoma: a case report and review of the literature. AB - INTRODUCTION: A subgaleal hematoma or subaponeurotic hemorrhage occurs infrequently and is usually seen in pediatric patients, especially in the neonatal period. It may be associated with coagulation disorders. CASE REPORT: We report on a previously healthy 19-month-old patient who presented with an extensive subgaleal hematoma and significant anemia secondary to a fall. DISCUSSION: A literature review was conducted, and the etiology, diagnosis, and treatment of the subgaleal hematoma are discussed. CONCLUSION: Conservative treatment, except in select severe cases, is recommended for this condition. PMID- 10532670 TI - Pediatric emergency medicine: legal briefs. PMID- 10532669 TI - Current income profile for academic pediatric emergency medicine faculty. AB - STUDY OBJECTIVES: To survey academic pediatric emergency medicine (PEM) programs for information on financial compensation and patient care activities of PEM faculty and compare the results to the financial data published by the AAEM, AAAP, and MGMA. METHODS: A survey was mailed to program directors requesting information on medical school affiliation, ED census, recruitment, patient care activity and annual income for each academic rank. The survey also included questions on CME benefits, and income adjustment mechanisms/bonus plans for PEM faculty. The survey income data were stratified by program size and geographic region and then compared to income data from the AAMC, AAAP, and MGMA. RESULTS: Of 47 eligible programs, 37 (78.7%) responded,and four were excluded. Mean number of clinical hours per week for academic faculty and clinical faculty were 27.9 +/ 3.5 and 32.4 +/- 3.9, respectively, (P = 0.000). Clinical appointments in academic departments were offered by 82% of the programs. Mean annual income for all academic ranks was $121,503 +/- $15,795, and is nearly $37,000 less than the annual income for academic adult emergency medicine (AEM) faculty. Compared to medium and large programs, small programs are offering higher salaries to recent fellowship graduates (P = 0.004). When income data were stratified by program size or geographic region, no significant difference in average annual income was observed. Bonus or incentive plans were available only in 45.5% of the programs. CONCLUSION: Direct patient care responsibility of PEM academic faculty has not changed significantly in the past 13 years, despite the availability of clinical appointments within most of the surveyed programs. Our data indicate that the annual income for PEM faculty in academic institutions is significantly less than AEM faculty. No significant difference was observed between programs at the assistant, associate, or full professor level when stratified by size or geographic region. Bonus/incentive plans for exceptional patient care or scholarly activity were available in less than half of the surveyed programs. PMID- 10532671 TI - Stomachache and vomiting for 1 day. PMID- 10532672 TI - Acute pediatric digoxin ingestion. AB - Although most acute pediatric ingestions of digoxin or other related cardiac glycosides result in minimal or no symptoms, occasionally a child is symptomatic. Gastrointestinal complaints or first-degree AV block are the most common presenting symptoms. Children can generally be given a single dose of activated charcoal, observed, and discharged without any subsequent problems. However, some patients will be toxic and require monitoring, medication, and possibly digoxin specific antibody fragments. The most important role of the clinician is to recognize the clinical manifestations and institute the appropriate therapy. As in the case presented, the history of an ingestion may not always be obtained initially. Thus, the physician should maintain a high index of suspicion for acute digoxin ingestion and order the appropriate confirmatory tests (eg, a digoxin level, a potassium level, and a 12-lead ECG) when necessary. PMID- 10532673 TI - Sports-specific concerns in the young athlete: football. AB - There are over 1.5 million males playing American football at all levels in the United States. American football is the most common participant sport among high school-aged males. Owing to its high rate of injury per exposure hour, American football injuries are commonly treated in the emergency department during the autumn sports season. This article will review the history, epidemiology, and specific injury patterns seen in American football, with a focus on head and shoulder injuries. PMID- 10532674 TI - Acute care of the child with a tracheostomy. PMID- 10532675 TI - Can parents predict a child's taste in antibiotics? PMID- 10532676 TI - Obesity is an environmental issue. AB - Obesity is an environmental issue. Societies that are transitioning to westernized lifestyles are experiencing substantial increases in its prevalence. The primary environmental determinants of obesity are high calorie intake and low levels of activity. Socioeconomic status and place of residence are important contributors. These factors together comprise an obesogenic or 'toxic' environment where the development of obesity is the expected course for humans leading lifestyles incompatible with their evolutionary development. Only by addressing and modifying the toxic environment will we be able to stem the obesity epidemic. PMID- 10532677 TI - Oxidation of heparin-treated low density lipoprotein by peroxidases. AB - Low density lipoproteins (LDL) can bind to glycosaminoglycans and proteoglycans rich in heparin and chondroitin sulphate in the arterial intima and may become a target for atherogenic modification by myeloperoxidase activity. We have examined the susceptibility of resolubilized LDL, that has been precipitated from serum with heparin (HepLDL), to peroxidase-H2O2-catalysed oxidation and the effects of antioxidants and components of human serum on the oxidation. HepLDL was oxidised rapidly by horse radish peroxidase (HRP) and H2O2 (mean t1/2max for conjugated diene formation, 3 min) while there was little oxidation of native LDL or native LDL precipitated with polyethyleneglycol and resolubilised during the 30 min incubation period. The formation of thiobarbituric acid reacting substances (TBARS) essentially paralleled that of conjugated dienes during oxidation of HepLDL. HepLDL was also more rapidly oxidised than native LDL by myeloperoxidase H2O2. Oxidation of HepLDL by peroxidases did not require free tyrosine, was almost totally inhibited by butylated hydroxytoluene (BHT) and ascorbate, and was unaffected by vitamin E and urate. Increasing concentrations (0-14.9%) of beta lipoprotein deficient serum (BLPDS) significantly (P<0.0001) inhibited the formation of TBARS during HepLDL oxidation catalysed by HRP and partially inhibited the corresponding myeloperoxidase-catalysed oxidation. This inhibitory activity was removed by dialysis and gel-filtration of BLPDS and was not restored by addition of magnesium ions used in the isolation of BLPDS, or physiological levels of ascorbate, tyrosine and reduced thiols (cysteine) to gel-filtered BLPDS. The results indicate that LDL from complexes with glycosaminoglycans are highly susceptible to oxidation by peroxidases, particularly at low levels of water soluble antioxidants, and that vulnerability of these LDL to myeloperoxidase oxidation remains in the presence of serum components that should exist in the arterial intima. These findings may be relevant to the oxidative modification of LDL that becomes trapped by binding to arterial proteoglycans and to the formation of myeloperoxidase-modified LDL in the artery wall. PMID- 10532678 TI - A simple and sensitive method in using the ratios of cholesteryl ester molecular species as indexes of oxidative stress in plasma and lipoprotein fractions. AB - To develop a simpler method for lipid peroxidation which may replace the classic gold standard of measuring the loss of polyunsaturated fatty acids, we investigated the use of ratios of molecular species of neutral lipids (MSNL) separated by gas chromatography in a single step. We sub-fractionated lipoproteins and subjected each to oxidation. Among different combinations of ratios of MSNL, we found that the ratios of cholesteryl esters (CE) containing C20/C16 (R1) and C18/C16 (R2) decreased most rapidly with increasing Cu2+ concentration and with increasing oxidation time, for all lipoproteins. We suggest that these two CE ratios can be used as oxidative indexes for all lipoproteins and thus for intact plasma. Experimental evidence showed that the oxidative index of whole plasma is the weighted average index of its lipoproteins. This method was validated with thiobarbituric acid method. Normal healthy asymptomatic males had R1 value 0.496 +/- 0.130, and R2 4.674 +/- 0.848, and females had R1 0.535 +/- 0.117 and R2 4.569 +/- 0.733 with no significant gender differences. Both ratios showed low variabilities within each individual. The method was tested to be feasible in monitoring vitamin E supplementation study. This CE-ratio method is concentration independent. It is simple, rapid, and highly reproducible, and, suitable for screening plasma on large scale. PMID- 10532679 TI - Inhibitory effect of TS-962 on the formation of early atherosclerotic lesions in high fat-fed hyperlipidemic hamsters. AB - The hypocholesterolemic and anti-atherosclerotic effect of TS-962, a specific ACAT inhibitor, was investigated in a hamster model fed a high fat diet containing 10% coconut oil and 0.05% cholesterol. Lipid accumulated atherosclerotic lesions were detected by using oil red O staining in the lesion prone aortic arch. A high dose, estimated to be 15 mg/kg, of TS-962 decreased serum cholesterol to normal levels with reduction of liver cholesterol contents below normal levels, and as a consequence, entirely inhibited the lipid accumulation in the aortic arch. Furthermore, a low dose, estimated to be 1.5 mg/kg, of TS-962 remarkably inhibited aortic lipid accumulation by 73% compared with the control group, without changing either serum cholesterol level or liver cholesterol content. These findings suggest that TS-962 is effective in the primary prevention of atherosclerosis by directly suppressing the formation of foam cells in arteries. PMID- 10532680 TI - Common carotid intima-media thickness in patients with peripheral arterial disease or abdominal aortic aneurysm: the SMART study. Second Manifestations of ARTerial disease. AB - Evidence is emerging that the contribution of atherosclerosis to the development of abdominal aortic aneurysm may differ from that of other manifestations of arterial disease. B-mode ultrasound may be helpful in understanding the characteristics and factors that contribute to the development of different manifestations of arterial disease. We examined whether there is a difference in common carotid intima-media thickness (IMT), an indicator of generalized atherosclerosis, in patients with peripheral arterial disease (PAD) and abdominal aortic aneurysm (AAA). IMT of the left and right common carotid artery was measured in the first 172 patients (123 PAD and 49 AAA) enrolled in the Second Manifestations of ARTerial disease (SMART) study, a cohort study among patients with a manifestation of atherosclerotic vascular disease or risk factors for atherosclerosis. Mean IMT was 0.98 +/- 0.34 mm in patients with PAD and 0.91 +/- 0.20 mm in patients with AAA, with an age and sex adjusted mean difference of 0.18 mm (95% CI 0.08; 0.28). After additional adjustments for cardiovascular risk factors, the difference remained 0.11 mm (95% Cl 0.01; 0.21). Common carotid IMT in patients with AAA is on average smaller than in patients with PAD, independent of other determinants of IMT. These findings support the view that the development of AAA cannot completely be explained by atherosclerosis and is in part due to other pathophysiological mechanisms. PMID- 10532681 TI - Angiotensin II atherogenicity in apolipoprotein E deficient mice is associated with increased cellular cholesterol biosynthesis. AB - Angiotensin II (Ang II) was shown to be an important risk factor for accelerated atherosclerosis. Inhibition of Ang II action on the arterial wall by blocking its production with angiotensin converting enzyme (ACE) inhibitors, or by blocking binding to its receptors on cells with antagonists was shown to attenuate atherogenesis in animal model of atherosclerosis. We questioned whether Ang II atherogenicity is related to a stimulatory effect of Ang II on macrophage cholesterol biosynthesis. Angiotensin II injected intraperitoneally once a day (0.1 ml of 10(-7) M per mouse) for a period of 30 days, to the apolipoprotein E deficient mice increased the atherosclerotic lesion area by 95% (P < 0.01 vs. control), compared to placebo-injected mice, with no significant effect on blood pressure or on plasma cholesterol levels. On using mouse peritoneal macrophages (MPMs) that were harvested after intraperitoneally injection of Ang II, an increased rate of cellular cholesterol biosynthesis (measured as incorporation of [3H]acetate into cholesterol) by up to 90% (P < 0.01 vs. control) was observed. In mice treated with the ACE inhibitor, Fosinopril (25 mg/kg per day) a reduction in their MPM's cholesterol synthesis by up to 70% (P < 0.01 vs. control) was obtained. In vitro studies in human monocyte-derived macrophages (HMDM), in MPMs from control BALB/c mice, and in J-774 A.1 macrophage-like cell line demonstrated up to 44, 34 and 30% stimulation of macrophage cholesterol biosynthesis, respectively, following cell incubation with 10(-7) M Ang II for 18 h at 37 degrees C. The stimulatory effect of Ang II on macrophage cholesterol biosynthesis could be related to its interaction with the macrophage AT1 receptor, as Losartan (10(-5) M), an AT1 blocker, but not PD 123319 (10(-5) M), an AT2 blocker, prevented the stimulatory effect on macrophage cholesterol synthesis. Furthermore, in cells that lack the AT1 receptor (RAW macrophages), Ang II did not increase cellular cholesterol synthesis. Ang II increased macrophage 3-hydroxy-3-methyl glutaryl CoA (HMG CoA) reductase mRNA levels in a dose dependent manner in J-774 A.1 macrophages and in MPM. Losartan, the AT1 receptor antagonist clearly attenuated this mRNA induction. We thus conclude that Ang II stimulation of macrophage cholesterol biosynthesis is related to its interaction with the AT1 receptor, followed by stimulation of macrophage HMG CoA reductase gene expression, which leads to increased cellular cholesterol biosynthesis, and can possibly result in macrophage cholesterol accumulation and foam cell formation. PMID- 10532682 TI - Hypolipidemic effect of NK-104, a potent HMG-CoA reductase inhibitor, in guinea pigs. AB - The hypolipidemic effect of NK-104 and its mechanisms of action (effects on hepatic sterol synthesis, low density lipoprotein (LDL)-receptor expression and very low density lipoprotein (VLDL) secretion) were studied in guinea pigs using simvastatin as a reference substance. There was a dose-dependent and significant reduction of both plasma total cholesterol (17.4, 24.5 and 45.3% at 0.3, 1 and 3 mg/kg, respectively) and triglycerides (21.1 and 32.2% at 1 and 3 mg/kg, respectively) after 14-day administration of NK-104. Simvastatin at 30 mg/kg lowered plasma total cholesterol (25.0%) but not triglyceride levels. NK-104 (3 mg/kg) and simvastatin (30 mg/kg) inhibited hepatic sterol synthesis by approximately 80%, 3 h after dosing, and enhanced LDL receptor binding-capacity of liver membranes 1.5-fold after 14-day dosing. The former group accelerated LDL clearance somewhat more markedly than the latter, and increased fractional catabolic rate 1.8-fold (vs. 1.4-fold). Furthermore, only the NK-104 (3 mg/kg) suppressed VLDL secretion into the liver perfusate (triglyceride. 19.9%; apoB, 24.2%) with extensive reduction of hepatic sterol synthesis caused by prolonged action. These results indicate that NK-104 and simvastatin at 10 times the dosage of the former, similarly enhances hepatic LDL receptor; however, only NK-104 with prolonged action suppresses VLDL secretion to show higher cholesterol-lowering potency and triglyceride-reducing effect. PMID- 10532683 TI - Differences in intima-media thickness in the carotid and femoral arteries in familial hypercholesterolemic heterozygotes with and without clinical manifestations of cardiovascular disease. AB - It is unknown whether the variation in severity of cardiovascular disease (CVD), seen in patients with familial hypercholesterolemia (FH), is reflected in the intima-media thickness (IMT) of carotid and femoral arteries. We measured IMT in both these arteries in 248 consecutive patients with FH, attending our Lipid Clinic. One hundred and six patients were classified as having CVD, while the remaining FH subjects had no clinical evidence of CVD. IMT measurements of 20 prespecified carotid and femoral arterial wall segments of the FH groups with and without CVD were compared. All IMTs in both groups were severely thickened with respect to normal controls. Furthermore, the highest IMTs and the largest absolute differences were observed in the common femoral artery (1.23 +/- 0.46 mm vs 1.10 +/- 0.51 mm; P = 0.006). In subjects with CVD, the distributions of IMT within tertiles for both arterial segments were opposite to those found in FH patients without CVD, (P < 0.05, for both segments). The mean IMT of, in particular, the common femoral artery is thicker in FH individuals with CVD compared with those without. Some FH patients have abnormal IMT of the femoral artery, whereas in others the carotid artery is more affected. Therefore, in FH patients, combined assessment of the carotid and femoral arterial walls provides a more accurate estimation of total atherosclerotic burden in FH. PMID- 10532684 TI - Up-regulation of low density lipoprotein receptor by a novel isobenzofranone derivative, MD-700. AB - Stimulatory effects of a novel isobenzofranone, MD-700, on low density lipoprotein (LDL) receptor activity were investigated in vitro and in vivo. MD 700 at 0.03 microg/ml elevated the expression of LDL receptor in HepG2 cells within 4 h. Corresponding to this, uptake of fluorescent labeled-LDL (3,3' dioctadecylindocarbocyanine-LDL) by the cells increased linearly in time- and dose-dependent manner by MD-700 for up to 12 h. In the experiment using HepG2 cells transiently transfected with promoter-luciferase gene constructs, MD-700 increased luciferase activity in a dose-dependent manner from 0.03 to 0.1 microg/ml. In contrast, luciferase activity was not stimulated by MD-700 in construct with a deleted sterol regulatory element (SRE)-1, suggesting importance of SRE-1 in stimulation of the LDL receptor gene promoter by MD-700. Binding experiments on liver membranes from MD-700-treated hamsters showed about a 60% increase in 125I-labeled LDL binding. A Scatchard plot revealed that MD-700 increased the maximal binding without affecting binding affinity. In contrast to findings with an inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A reductase, pravastatin, MD-700 had no effect on the sterol synthesis in hamster liver homogenates. These results suggest that MD-700 stimulates the expression of LDL receptor, presumably in a manner independent of change in sterol metabolism, and thereby promotes LDL clearance. Hypocholesterolemic actions of MD-700 in hamsters were then examined. MD-700 lowered serum cholesterol levels in hamsters fed normal chow or a high-fat diet. Fractionation of serum lipoproteins demonstrated that MD-700 selectively decreased LDL and very low density lipoprotein cholesterol. Dose-dependent decrease in serum cholesterol was also seen in hypercholesterolemic rats. Thus, the hypocholesterolemic action of MD-700 may be attributed to up-regulation of the LDL receptor, based on stimulation of the transcription of the LDL receptor gene. Although pravastatin stimulates LDL uptake and lowers serum cholesterol in a manner similar to that seen with MD-700, the mechanism responsible for hypocholesterolemic action appears to differ. PMID- 10532686 TI - Expression and localization of the proteoglycan decorin during the progression of cholesterol induced atherosclerosis in Japanese quail: implications for interaction with collagen type I and lipoproteins. AB - The temporal and spatial distribution, and relative levels of the proteoglycan decorin and collagen type I were examined during the progression of atherosclerosis in the dorsal aortas of Japanese quail selected for cholesterol induced atherosclerosis. The quail were placed on either a control or 0.5% added cholesterol diet at approximately 16 weeks of age. Dorsal aortas were collected at 1- or 2-week intervals over a 15-week period after initiating cholesterol feeding. Biochemical analysis for decorin and collagen type I showed that both increased in the cholesterol-fed birds compared to control-fed birds beginning at 9 weeks and continued through the duration of the study. Through immunohistochemical staining for decorin and collagen type I, the spatial localization of decorin and collagen type I in control and less severe plaques in cholesterol-fed birds was most prominent in the arterial adventitia. However, in severe atherosclerotic plaques, decorin was localized in foam cell regions and collagen type I was found surrounding the foam cell regions where decorin accumulated. These results demonstrated a localization of decorin in the core of the atherosclerotic plaque foam cell region with collagen type I being located on the plaque surface. PMID- 10532685 TI - Secretion of prebeta HDL increases with the suppression of cholesteryl ester transfer protein in Hep G2 cells. AB - Prebeta HDL are small, protein rich lipoproteins that are predominantly composed of apo A-I, without apo A-II. Prebeta HDL are secreted from the liver as nascent HDL and/or are produced in the incubated plasma by cholesteryl ester transfer protein (CETP). However, the role of CETP in the secretion of HDL from the liver has yet to be determined. In the present study, we examined the effect of the suppression of hepatic CETP by antisense oligodeoxynucleotides (ODNs) against CETP targeted to the liver on the secretion of apo A-I using a Hep G2 cell culture. The ODNs against CETP were coupled to asialoglycoprotein (ASOR) carrier molecules, which serve as an important method for the regulation of liver gene expression. Hep G2 cells were cultured in DMEM supplemented with 10 FBS. After 2 days, the medium was changed to DMEM with EGF and the cells were divided into three groups. The control group received saline, while the sense group was mixed with the sense ODNs complex and the antisense group was mixed with the antisense ODNs complex, respectively, for 2 days. Both the hepatic CETP mRNA and the CETP mass in the medium in the antisense group decreased significantly more than in the sense and the control groups (CETP mass: 1.697 + /- 0.410 ng/mg cell protein vs. 2.367 + /- 0.22 and 2.360 + /- 0.139, n = 3 in each determination). In contrast, both the hepatic apo A-I mRNA and the apo A-I mass in the medium in the antisense group were significantly higher than those in the sense and the control groups (apo A-I mass; 1.877 + /- 0.215 micro/mg cell protein vs. 1.213 + /- 0.282 and 1.097 + /- 0.144, n = 3 in each determination). The increase in apo A-I was mainly due to the increase in prebeta apo A-I. These findings may partly explain why HDL and apo A-I increase in patients with CETP deficiency, while also indicating the possibility that the original level of prebeta HDL is sufficient in such patients. PMID- 10532688 TI - Lack of antioxidant activity of the antiatherogenic compound L-arginine. AB - L-Arginine, the physiologic substrate of nitric oxide synthase, has antiatherogenic properties in animal models of atherosclerosis, and improves endothelial function in hypercholesterolemic humans. Some of these effects may be mediated by increased production of nitric oxide; however, some investigators have postulated a direct antioxidant action related to its aminoguanidine moiety. We aimed therefore, to assess the antioxidant properties of L-arginine. The antioxidant properties of 200 microM L-arginine. 200 microM D-arginine and 200 microM L-glutamate were compared with the powerful antioxidant ascorbate and a control (phosphate-buffered saline). Compounds were tested using four in vitro methods: (1) the Esterbauer technique (which tests the ability of the compounds to scavenge free radicals or chelate transition metals); (2) the effect on the autoxidation of ascorbate; (3) anti-tocopherol mediated peroxidation (which tests the compound's ability to synergize with alpha-tocopherol to prevent mild chemically induced LDL oxidation); and (4) the ability of the compounds to attenuate alpha-tocopherol radical in micellular emulsions (TRAA). The above methods were repeated using the metabolites of the test compounds after incubation with human endothelial cells. Ex vivo studies were then carried out by measuring levels of lipid peroxide production (using HPLC with UV and chemiluminescence detection) in three healthy volunteers before and 2 h after a single 7-g oral dose of L-arginine. By the Esterbauer technique, L-arginine increased lag time by 45% compared to control, as did D-arginine by 50%; L glutamate had no effect and ascorbate increased lag time by 325%. Neither L arginine, D-arginine or L-glutamate had significant effects on the autoxidation of ascorbate or anti-tocopherol mediated peroxidation. By the TRAA method, L arginine had a small effect on preventing the decay of tocopherol. The results were similar for the studies of the compound's metabolites. In ex vivo studies, no changes were seen in lipid peroxide levels following acute dosage with L arginine. L-Arginine has only weak and non-specific antioxidant effects, suggesting that its major cardioprotective benefits occur through other mechanisms, such as via the nitric oxide pathway. PMID- 10532687 TI - Plasma levels of the soluble fraction of tumor necrosis factor receptors 1 and 2 are independent determinants of plasma cholesterol and LDL-cholesterol concentrations in healthy subjects. AB - In the last few years, it has been demonstrated that tumor necrosis alpha (TNF alpha) has important effects on whole-body lipid metabolism. TNF-alpha administration has been found to produce an increase in serum cholesterol levels and increased hepatic hydro-3-methyl-glutaryl coenzyme A (HMG-CoA) reductase activity in mice. The purpose of this study was to test whether plasma levels of the soluble forms of the TNF-alpha receptors 1 and 2 (sTNFR1, sTNFR2) are associated with lipid abnormalities. A total of 36 healthy subjects (19 males, mean age 36.2 +/- 1.9, and 17 females, mean age 34.9 +/- 1.4) were studied. Plasma sTNFR1 levels correlated with total (r = 0.43, P = 0.01) and LDL cholesterol (r = 0.52, P = 0.002) levels, but not with total or HDL2-HDL3 subfractions of HDL-cholesterol, total plasma triglycerides, VLDL-cholesterol or VLDL-triglycerides (all r < 0.11, P = NS). Plasma sTNFR2 levels also correlated with total (r = 0.44, P = 0.009) and LDL-cholesterol (r = 0.57, P < 0.0001) levels, and negatively with HDL2-cholesterol (r = -0.37, P = 0.029). A stepwise multiple linear regression was constructed to predict total cholesterol levels, with BMI, sex, age, sTNFR1 or sTNFR2 as independent variables. Both sTNFR1 and sTNFR2 were significantly associated with total cholesterol (P = 0.031 and 0.009), contributing to 26 and 19%, respectively, of its variance. In another model in which LDL-cholesterol was substituted for total cholesterol, sTNFR1 or sTNFR2 (P = 0.0084 and 0.0005) were significantly associated with LDL cholesterol, contributing to 39 and 32% of its variance. In summary, plasma levels of sTNFR1 and sTNFR2 circulate in proportion to total and LDL-cholesterol in healthy subjects. PMID- 10532689 TI - Spectrum of LDL receptor gene mutations in Denmark: implications for molecular diagnostic strategy in heterozygous familial hypercholesterolemia. AB - Heterozygous familial hypercholesterolemia (FH) is one of the most common potentially fatal single-gene diseases leading to premature coronary artery disease, but the majority of heterozygous FH patients have not been diagnosed. FH is due to mutations in the gene coding for the low-density lipoprotein (LDL) receptor, and molecular genetic diagnosis may facilitate identification of more FH subjects. The Danish spectrum of 29 different mutations, five of which account for almost half of heterozygous FH, is intermediate between that of countries such as South Africa, where three mutations cause 95% of heterozygous FH in the Afrikaners, and Germany or England, where there are many more mutations. In clinical practice, a strategy for the genetic diagnosis of heterozygous FH, tailored to the mutational spectrum of patients likely to be seen at the particular hospital/region of the country, will be more efficient than screening of the whole LDL receptor gene by techniques such as single-strand conformation polymorphism (SSCP) analysis in every heterozygous FH candidate. In Aarhus, Denmark, we have chosen to examine all heterozygous FH candidates for the five most common LDL receptor gene mutations (W23X, W66G, W556S, 313 + 1G --> A, 1846 1G --> A) and the apoB-3500 mutation by rapid restriction fragment analysis. Negative samples are examined for other mutations by SSCP analysis followed by DNA sequencing of the exon indicated by SSCP to contain a mutation. If no point mutation or small insertion/deletion is detected, Southern blot or Long PCR analysis is performed to look for the presence of large gene rearrangements. In conclusion, our data suggest that an efficient molecular diagnostic strategy depends on the composition of common and rare mutations in a population. PMID- 10532690 TI - Peripheral blood mononuclear cell production of interleukin-8 and IL-8-dependent neutrophil function in hypercholesterolemic patients. AB - Interleukin-8 is a cytokine produced by mononuclear cells that is involved in polymorphonuclear neutrophil leukocyte (PMN) recruitment and activation. Several studies have previously demonstrated a leukocyte activation during hypercholesterolemia and 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors have been found to play a role in the prevention of atherothrombotic disease. The purpose of this study was to determine interleukin 8 (IL-8) mRNA expression and ex vivo production from peripheral blood mononuclear cells (PBMCs) and IL-8-dependent PMN activation of hypercholesterolemic (HC) patients with respect to normocholesterolemic (NC) subjects. Using Northern blot analysis, we found a four- and threefold increase in the amount of IL-8 transcript in PBMC from HC patients, in unstimulated and LPS stimulated cultures, respectively. A specific immunoassay showed a correspondingly significant increase of IL-8 immunoactivity in the conditioned medium of PBMC from HC subjects as compared with controls (unstimulated PBMC: 15 +/- 4 vs. 4.2 +/- 3 ng/ml; P < 0.0001; LPS stimulated PBMC: 65.3 +/- 8 vs. 36.6 +/- 9 ng/ml; P < 0.0001). PMN of HC patients stimulated with IL-8 showed a reduced elastase release with respect to NC subjects before physiological granule release after f Met-Leu-Phe (fMLP) treatment. These results indicate an upregulation of the IL-8 system in dyslipidemic patients and provide evidence for ongoing in vivo IL-8 dependent PMN activation during hypercholesterolemia. PMID- 10532692 TI - Long-term effects of fish oil on lipoprotein subfractions and low density lipoprotein size in non-insulin-dependent diabetic patients with hypertriglyceridemia. AB - The effects of fish oil on lipoprotein subfractions and low density lipoprotein (LDL) size in non-insulin-dependent diabetes mellitus (NIDDM) patients with hypertriglyceridemia are unknown. To elucidate this, 16 NIDDM hypertriglyceridemic patients (plasma triglyceride 2.25- 5.65 mmol/l, plasma cholesterol < or = 7.75 mmol/l) were randomly assigned to a 6-month period with either moderate amounts of fish oil (n = 8) or placebo (n = 8) after 4 weeks of wash-out and 3 weeks of run-in. Diet and hypoglycemic treatment were unchanged throughout the experiment. LDL size were evaluated at baseline and after 6 months. Three VLDL and LDL subfractions were measured at the end of the two periods. The total lipid concentration of all very low density lipoprotein (VLDL) subfractions was lower at the end of fish oil treatment compared with placebo (large VLDL 124.3 +/- 19.7 mg/dl vs 156.7 +/- 45.5 mg/dl; intermediate VLDL 88.5 +/- 9.5 mg/dl vs 113.9 +/- 23.2 mg/dl; small VLDL 105.9 +/- 9.7 mg/dl vs 128.9 +/ 40.7 mg/dl) (mean +/- SEM), although the difference was not statistically significant. Moreover, at the end of the two treatments, the percentage distribution of VLDL subfractions was very similar (large 37.5 +/- 3.3% vs 37.6 +/- 2.6%, intermediate 27.6 +/- 0.9% vs 31.0 +/- 2.4%; small 34.9 +/- 3.7% vs 31.4 +/- 2.1%). Concerning LDL, no significant change in LDL size was observed after the two treatments (255.4 +/- 2.2 A vs 254.2 +/- 1.7 A, fish oil; 253.7 +/- 2.0 A vs 253.3 +/- 1.7 A, placebo). LDL subfraction distribution was also very similar (large 17 +/- 3% vs 17 +/- 2%; intermediate 62 +/- 3% vs 65 +/- 3%; small 21 +/- 3% vs 18 +/- 2%), at the end of the two periods, confirming the lack of effects on LDL size. In conclusion, our study indicates that in NIDDM patients with hypertriglyceridemia, fish oil does not induce any improvement in LDL distribution and LDL size despite its positive effects on plasma triglycerides. PMID- 10532691 TI - Endothelin-1 stimulates proliferation of human coronary smooth muscle cells via the ET(A) receptor and is co-mitogenic with growth factors. AB - We investigated the effects of endothelin-1 (ET-1) on growth of cultured human coronary artery smooth muscle cells (cSMC). ET-1 alone stimulated DNA synthesis in growth-arrested cSMC as measured by [3H]thymidine incorporation, with a maximum 63 +/- 23% increase above control by 10(-7) M (P < 0.05). ET-1 (10(-7) M) also stimulated increases in cyclin D1 protein levels after 24 h, and in absolute cell number after 4 days. Furthermore, ET-1 stimulated protein synthesis (maximum 73 +/- 32% increase in [3H]leucine incorporation by 10(-7) M (P < 0.05)), as well as triggering intracellular calcium transients in human cSMC, as visualised under fura-2 fluorescence microscopy. The selective ET(A) receptor antagonist BQ123 inhibited the increases in DNA synthesis, cell number, protein synthesis and intracellular calcium concentration in response to ET-1, whereas the ET(B) receptor antagonist BQ788 had no such effects. Furthermore, the ET(B) agonist sarafotoxin 6c had no effect on cSMC DNA synthesis. In addition, co-incubation of ET-1 with threshold concentrations of the growth factors, platelet-derived growth factor-BB (PDGF-BB), basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF), resulted in pronounced synergistic increases in DNA synthesis over that observed with the factors alone. In conclusion, we have shown that ET-1 stimulates proliferation of human cSMC via the ET(A) receptor and is also a co mitogen with the growth factors tested. These findings indicate a role for ET-1 in the development of coronary intimal hyperplasia in man. PMID- 10532693 TI - The effect of aggressive and moderate lowering of LDL-cholesterol and low dose anticoagulation on plasma lipids, apolipoproteins and lipoprotein families in post coronary artery bypass graft trial. AB - The reported results (The Post Coronary Artery Bypass Graft Trial Investigators. The effect of aggressive lowering of low-density lipoprotein cholesterol levels and low-dose anticoagulation on obstructive changes in saphenous-vein coronary artery bypass grafts. New Engl J Med 1997;336:153-162) of the Post Coronary Artery Bypass Graft (Post CABG) trial have shown that aggressive lowering was more effective than moderate lowering of low density lipoprotein (LDL) cholesterol in reducing the progression of atherosclerosis in saphenous-vein grafts (27 vs. 39%; P < 0.001); low dose warfarin had no effect on the progression of atherosclerosis. The present report describes the effect of long term (an average of 4.3 years) aggressive treatment with high (40-80 mg/day) and moderate treatment with low (2.5-5 mg/day) doses of lovastatin on lipids, apolipoproteins (apo) and apoA- and apoB-containing lipoprotein families. To achieve the target LDL-cholesterol levels (60-85 mg/dl for aggressive group and 134-140 mg/dl for moderate group), cholestyramine (8 g/day) was given to 25% of subjects on aggressive and 5% of subjects on moderate treatment. Although with both treatment strategies there were significant decreases (P<0.001) in the levels of total cholesterol, LDL-cholesterol, apoB, LDL-apoB and cholesterol-rich Lp-B family, percent changes in the levels of these variables were greater in the aggressive- than in the moderate-treatment groups. These treatments had only marginal effects in increasing the levels of high density lipoprotein cholesterol, apoA-I and Lp-A-I and Lp-A-I:A-II families. The long-term aggressive treatment exerted no effect on the concentrations of triglycerides, apoC-IlI, apoC-III in VLDL + LDL and triglyceride-rich Lp-Bc families. Neither treatment affected the levels of Lp(a). The potentially modifying influence of warfarin and apoE phenotypes on lovastatin-induced changes in lipoprotein variables was found to be of little significance. It is likely that the beneficial effect of lovastatin in reducing the progression of atherosclerosis in grafts is mediated through its specific lowering effect on cholesterol-rich Lp-B particles. PMID- 10532695 TI - Are all low-density lipoprotein particles partially desialylated in plasma? PMID- 10532694 TI - Effect of heparin-stimulated plasma lipolytic activity on VLDL APO B subclass metabolism in normal subjects. AB - Heparin given intravenously enhances lipolysis, although fasting lipids are not markedly altered in long-term administration. In the present study we investigated heparin-induced acute perturbation of VLDL subclass metabolism. Eight men were examined during a control study and during an 8.5 h infusion of heparin. 2H3-leucine was used as tracer and kinetic constants derived using a non steady-state model. Heparin infusion increased both plasma lipoprotein and hepatic lipase activity and raised plasma FFAs two-fold (P < 0.001). The fractional catabolic rate (FCR) of VLDL1 apo B increased on heparin (25.7 +/- 4.2 and 10.8 +/- 1.7 pools/d, heparin vs. control, P < 0.02). The FCR of VLDL2 apo B increased to 12.6 +/- 1.9 pools/d on heparin vs. 8.8 +/- 1.1 pools/d during the control (NS). Total VLDL apo B production was not significantly changed (824 +/- 45 and 692 +/- 91 mg/d, heparin vs. control, NS). We conclude that during heparin infusion, the catabolism of especially large triglyceride-rich VLDL1 apo B is greatly increased. However, although the FFA levels were high during the heparin study, the production of total VLDL apo B did not rise. These findings are consistent with the known action of heparin on lipoprotein lipase but indicate that acute increase in plasma FFA levels does not lead to a rise in VLDL apo B production. PMID- 10532696 TI - Reduced frequency of the thermolabile methylenetetrahydrofolate reductase genotype in the elderly. PMID- 10532698 TI - Is the pathogen of prion disease a microbial protein? AB - Though considerable circumstantial evidence suggests that the pathogen of prion disease is proteinaceous, it has not yet been conclusively identified. Epidemiological observations indicate that a microbial vector is responsible for the transmission of natural prion disease in sheep and goats and that the real causative agent may correspond to a structural protein of that microorganism. The microbial protein should resemble prion protein (PrP) and may replicate itself in the host by using mammalian DNA. A similar phenomenon was already described with a protein antigen of the ameba Naegleria gruberi. The various serotypes of the microbial protein may account for the existence of scrapie strains. It is proposed that many microbial proteins may be capable of replicating themselves in mammalian cells eliciting and sustaining thereby degenerative and/or autoimmune reactions subsequent to infections with microorganisms. PMID- 10532697 TI - Cytotoxic cholesterol is generated by the hydrolysis of cytoplasmic cholesteryl ester and transported to the plasma membrane. AB - The present study examines the fate and effects of free cholesterol (FC) generated by the hydrolysis of cytoplasmic cholesteryl esters (CE) in model macrophage foam cells. J774 or elicited mouse peritoneal macrophages (MPM) were enriched with CE by incubating with acetylated low density lipoprotein (acLDL) and FC/phospholipid dispersions, thus creating model foam cells. Treatment of the foam cells with the acyl coenzyme-A:cholesterol acyltransferase (ACAT) inhibitor, CP-113,818, in the absence of any extracellular cholesterol acceptors, resulted in cellular toxicity. This was accompanied by an increase in the amount of FC available for oxidation by an exogenous cholesterol oxidase. Furthermore, cellular toxicity was proportional to the size of the oxidase susceptible pool of FC over time. Morphological analysis and in situ DNA fragmentation assay demonstrated the occurrence of apoptosis in the ACAT inhibited cells. Co treatment with the hydrophobic amine U18666A, an intracellular cholesterol transport inhibitor, led to a dose dependent reduction in cytotoxicity and apoptosis, and blocked the movement of FC into the oxidase susceptible pool. In addition, treating model foam cells with CP-113,818 plus chloroquine, a compound that inhibits the function of acidic vesicles, also diminished cellular toxicity. Staining with the cholesterol binding dye filipin revealed that the macrophages treated with CP-113,818 contained a cholesterol oxidase accessible pool of FC in the plasma membrane. These results suggest that FC generated by the hydrolysis of cytoplasmic CE is transported through acidic vesicles to the plasma membrane, and accumulation of FC in this pool triggers cell death by necrosis and apoptosis. PMID- 10532699 TI - Postoperative delirium and plasma melatonin. AB - The molecular mechanisms of postoperative delirium are not understood in detail yet. This condition is similar to cases of mental symptoms in interferon therapy or hemodialysis. We propose that postoperative delirium is caused by a deficiency of serotonin, an important neurotransmitter, and review the evidence supporting our hypothesis. The serotonin deficiency results from a decrease in tryptophan, the serotonin precursor, and from an increase in melatonin, a serotonin metabolite. Our hypothesis may be applicable to the mechanisms of mental symptoms in interferon therapy and hemodialysis. In addition, we discuss to the relationship between homeostasis and biorhythms because postoperative delirium may be a dysrhythmia. PMID- 10532700 TI - Secretion of cytokines and modulation of function of immunologically competent cells. AB - Cytokines and their effects are widely studied in immunology, but the cellular mechanisms responsible for producing these substances are not yet described or proven. Antigen presenting cells (APCs), macrophages and dendritic cells are suitable subjects for such a study. One of the many papers illustrating the interplay between phagocytosis, cytokine secretion, and cell adhesion is one by Kitajima et al. The intracellular processes involve both cytokine secretion and mobilization of the membrane system of the cell. Five pieces of evidence are presented to propose that the Golgi complex is the source both of the membrane translocated and cytokine production. Morphologic evidence is cited that reverse pinicytosis does not play a significant role in the mouse macrophage system. PMID- 10532701 TI - On the role of neurobiological and genetic factors in the etiology and pathogenesis of suicidal behavior among immigrants. AB - Immigrants have higher rates of suicidal behavior than those in their countries of origin and their new countries. Immigration is a stressful life event which may lead to depression and suicidal behavior. Depressive disorders and suicidal behavior have a firm neurobiological and genetic basis. The author suggests that most immigrants who exhibit suicidal behavior in the new country had suicidal tendencies, and/or some degree of depression, and/or certain maladaptive personality traits in their countries of origin. The paper emphasizes the role of genetic and neurobiological factors in the human response to stressful events. PMID- 10532702 TI - Effects of psychological factors on the development of cardiovascular pathology: role of the immune system and infection. AB - Psychological factors affect the condition of both healthy and sick people. Acute and chronic stress, depression, anxiety, maladaptive personality traits, and other behavioral disorders cause the diseases of the heart and blood vessels. The same psychological factors suppress the immune system and promote infection. Recent studies have demonstrated that infections can play an important role in the development of coronary artery disease. The author suggests that the immune system may mediate the harmful effects of certain psychological factors on the cardiovascular system. PMID- 10532703 TI - Early cancer detection by microsatellite marker analysis. AB - A major goal of tumor biology has been the development of tumor markers that are useful for the diagnosis and management of cancer. A drawback associated with many of these markers is that they are not unique to tumor cells, but rather are normal or developmental antigens which are overexpressed in tumor tissue Therefore, determination of possible malignancy is based on a designated expression level rather than a clear-cut difference. Studies have shown that DNA from tumor cells has a pattern of chromosomal deletion clearly distinguishable from normal cell DNA, and more importantly, DNA from tumor cells can be detected in the serum of cancer patients. Particular chromosomal deletion patterns are associated with specific tumor types. It is hypothesized that individuals at risk for certain genetically well-characterized cancers, could be successfully screened for those cancers by a PCR-based blood test. In this way, neoplasia could be detected at earlier, more treatable stages of development. PMID- 10532704 TI - Information and redundancy: key concepts in understanding the genetic control of health and intelligence. AB - A model is proposed in which information from the environment is analysed by complex biological decision-making systems which are highly redundant. A correct response is intelligent behaviour which preserves health; incorrect responses lead to disease. Mutations in genes which code for the redundant systems will accumulate in the genome and impair decision-making. The number of mutant genes will depend upon a balance between the new mutation rate per generation and systems of elimination based on synergistic interaction in redundant systems. This leads to a polygenic pattern of inheritance for intelligence and the common diseases. The model also gives a simple explanation for some of the hitherto puzzling aspects of work on the genetic basis of intelligence including the recorded rise in IQ this century. There is a prediction that health, intelligence and socio-economic position will be correlated generating a health differential in the social hierarchy. Furthermore, highly competitive societies will place those least able to cope in the harshest environment and this will impair health overall. The model points to a need for population monitoring of somatic mutation in order to preserve the health and intelligence of future generations. PMID- 10532705 TI - Pre-eclampsia: a mistake of trophoblastic cells for tumour cells? AB - Pre-eclampsia is a severe form of hypertension induced by pregnancy. In pre eclampsia, there is deficient trophoblast invasion of the spiral arteries terminating in the placental bed. Perhaps some abnormality occurs in the immunosuppressive process as the maternal immune system encounters paternal antigens expressed by immunosuppressive decidual cells. This immunosuppressive abnormality might cause the deficient trophoblast invasion. Abnormal placentation might then lead to maternal endothelial cell damage by an ongoing process. There might be a recurring sequence of 4 steps: (1) The placenta releases trophoblastic cells with potentially cytotoxic characteristics. These circulating trophoblastic cells have an abnormal pattern of expression of integrins and perhaps other glycoproteins or proteins. (2) The circulating trophoblastic cells loosely bind to maternal endothelial cells, targeting them for anti-tumourigenesis. (3) The maternal immune system reacts against targeted maternal endothelial cells through anti-tumourigenic mechanisms. (4) Widespread maternal endothelial damage causes the characteristic kidney lesion called glomerula endotheliosis. PMID- 10532706 TI - Cellular pathogenesis of Alzheimer's disease. AB - The author proposes that paired helical filaments, which contain the protein tau in the fibrillar or beta-pleated sheet conformation, compete with microtubules for binding to nascent, soluble tau. Binding of nascent tau to tau in the beta pleated sheet conformation autocatalyzes the conformational change into the beta pleated sheet conformation. As long as sufficient tau is present to stabilize microtubules, neuronal function is normal. However, because paired helical filaments are resistant to proteolysis, they accumulate and eventually bind the bulk of nascent tau. This results in progressive microtubule instability and eventually neuronal death. Senile plaques are involved in Alzheimer's disease pathogenesis in that they contain fibrillar proteins which may function as heteronucleants, catalyzing the fibrillogenesis of other proteins such as tau. In this paradigm, apolipoprotein E4 serves as a heteronucleant for fibrillogenesis of tau. PMID- 10532707 TI - A novel tumor-specific gene therapy for bladder cancer. AB - Gene therapy has been successfully used to treat genetic diseases. Currently, much investigation involves the role of gene therapy in malignant tumors. One problem associated with the retroviral system used for gene therapy is its non specificity. Herein a vector delivery system is described, using human telomerase reverse transcriptase (hTRT) promotor, which can specifically affect telomerase positive tumor cells while sparing nearby telomerase-negative cells. By combining a self-containing Cre/loxP site-specific recombination system into the design, this vector will destroy telomerase-positive, p53-negative tumor cells, while sparing normal cells which are telomerase-positive with wild type p53 (such as activated lymphocytes). This vector design appears especially suited to bladder transitional cell carcinoma, because of easy access transurethrally and high rate of local recurrence, and biologically secondary to high proportion of telomerase activity and p53 dysfunction. PMID- 10532708 TI - Human chorionic gonadotropin: a secretory hormone. AB - There are several physiological and pathophysiological situations where there is an apparent fluid flux across plasma membranes at the time when human chorionic gonadotropin (hCG) levels are high. These fluxes may take the form of a fluid loss from gastrointestinal tract (e.g. emesis/hyperemesis gravidarum) or accumulations in enclosures (e.g. amniotic fluid or hydatidiform mole). What is not obvious though is whether hCG is the cause of these fluid fluxes. Although glycoprotein hormones like hCG are mainly hormonogenic, their action in the latter process involves the efflux or conductance of halide ions. Since the basis of fluid secretion is an active efflux of ions such as chloride stimulated by a humoral agent, accompanied by a passive diffusion of water across a cell wall, I hypothesize that hCG is also a secretory hormone and responsible for fluid fluxes in the above and other clinical situations. PMID- 10532709 TI - Prediction of ulcer formation on the skin. AB - The study reported in this paper investigates the feasibility of developing criteria for predicting the formation of ulcers on the human skin. Published data on the effect of pressure, friction, shear, oxygen tension, temperature, and exposure time on ulcer formation are analyzed, combined, and incorporated in developing the criteria. The results illustrate the relative contributions of the different parameters to the formation of ulcers. The developed relationships can provide guidelines for more rigorous clinical studies which would take into consideration the effect of age, sex, and physical conditions on the formation of ulcers at different sites of the body. PMID- 10532710 TI - Drugs that induce neutropenia/agranulocytosis may target specific components of the stromal cell extracellular matrix. AB - The etiology of drug-induced agranulocytosis is poorly understood. Many drugs that induce neutropenia or agranulocytosis can be metabolized to reactive intermediates that covalently bind to macromolecules. Until now, the myeloid precursor cell or an earlier committed progenitor cell has been favoured as the target for toxicity, due to evidence in some cases of cytotoxic action or antibodies against neutrophils. In the bone marrow, where neutrophils mature, certain components of the stromal microenvironment, e.g. intracellular adhesion molecule 1, vascular cell adhesion molecule 1, CD11b/CD18, heparan sulfate proteoglycans, fibronectin and hemonectin are essential for normal myeloid maturation. This article proposes that drugs implicated in agranulocytosis, or more likely their reactive metabolites, interact with specific components of the extracellular matrix and interfere with the normal regulation of granulopoiesis. PMID- 10532711 TI - Hysteria: a delusional state. AB - This paper explores the hypothesis that hysterical symptoms may best be regarded as the product of delusional thinking. Hysteria has generally been interpreted as a process quite distinct from other psychiatric disorders. An attempt is made to bring it within more conventional bounds. PMID- 10532712 TI - Research-overview article: membrane lipolysis and cholinergic priority. AB - Cholinergic drugs and antigenic and toxicological challenges induced lipolysis in twelve sheep. A lipolytic end product, the PGF2alpha metabolite, was found to be a reliable non-specific cholinergic marker. The lipolytic membrane alterations supported the concept of a general priority of the cholinergic system. A main feature is the breaking of molecular stability in dynamic hydrogen-bond interactions. Both acetylcholine and dioxygen reactivity are apparently moderated by cholinesterases. Free radicals appeared to be normal intermediates of catabolism, serving to neutralize excess protons. Antioxidants regenerate molecular oxygen, and so counteract part of excess activated oxygen. Intrinsic reactivity against its own structures characterizes the immuno-cholinergic system. Genetic priority could be assumed for cholinergic constituents and constitutions. A broad spectrum of etiologies was suggested. Lasting or repeated challenges may cause heterochiral conversions of vital proteins. The priority aspect of cholinergism also suggested methods to rank among the multitude of secondary biomolecules. PMID- 10532713 TI - Porphyria and chemicals. AB - Porphyria is a genetic family of diseases that is most frequently described as neuropsychiatric or toxogenetic. It is well known to be initiated by drugs, infections, heavy metals, hormones, chemicals and fasting. There are extensive lists of drugs that have been known to cause attacks. Others are thought to be likely to cause attacks on the basis of animal studies or in vitro studies. It has become obvious that lists of chemicals capable of causing illness in porphyrics are sorely lacking. Chemicals that have the same base as drugs that are labeled in the PDR (Physicians Desk Reference) as porphyrogenic have no such labeling in their MSDS (Material Safety Data Sheet). This article is intended to point out why porphyria needs to be considered when illness occurs after chemical exposure. The capability of testing enzymes in the porphyrin pathway allows us to evaluate these patients more thoroughly, for we are now aware that the standard measures for recognizing these diseases are often inadequate. Three examples where illness has occurred after environmental exposure to chemicals will serve as illustrations. One, a documented porphyria, is the Turkish porphyria. The other two are not yet documented as porphyria, but may be some day. One is Agent Orange which caused illness in Vietnam, and the other is exposure to unknown sources of what has been named the Gulf War syndrome. PMID- 10532714 TI - Weight loss in cancer and Alzheimer's disease is mediated by a similar pathway. AB - Weight loss, despite adequate caloric intake, occurs in Alzheimer's disease and cancer. Similar alterations of biochemistry accompany the weight loss in both diseases, and this suggests a common pathway of weight loss. If this hypothesis is correct, then drugs that prevent weight loss in cancer should also prevent weight loss in Alzheimer's disease. Megestrol acetate prevents weight loss in human and animal cancers. Omega 3 fatty acids prevent weight loss in animal models of cancer. Both compounds might prevent weight loss in Alzheimer patients. PMID- 10532715 TI - Ocular malformations and developmental genes. AB - New insights into the pathogenesis of ocular malformations came with the discovery of transcription factors that determine the fate of cells in the developing eye. Several malformations have been matched to individual developmental genes that share conserved DNA sequences such as the homeobox. These disease/gene matches include the oculorenal syndrome and PAX2; aniridia and PAX6; Rieger syndrome and RIEG1/PITX2; cyclopia and Sonic hedgehog; cone-rod dystrophy, Leber's congenital amaurosis and CRX; and recessive septooptic dysplasia and HESX1. Gene mapping and mutation analysis have allowed a more accurate and meaningful classification of genetically heterogeneous diseases such as the anterior segment dysgenesis syndromes. This paper reviews current information on the genetics of ocular malformations. PMID- 10532716 TI - Retinopathy of prematurity. PMID- 10532717 TI - The natural history of infantile esotropia during the first six months of life. Pediatric Eye Disease Investigator Group. AB - PURPOSE: The present study addresses the natural history ocular alignment in infantile esotropia that presents at 2 to 4 months of age. METHODS: Eye alignment during the first 6 months of life was evaluated in two cohorts of healthy infants who initially had esotropia at 2 to 4 months of age; 80 infants were enrolled in a prospective study at the Retina Foundation of the Southwest (RFSW), and 41 infants were reviewed retrospectively as a pilot study for the Early Surgery for Congenital Esotropia (ESCET) multicenter trial. In addition, 79 of the 80 children in the RFSW cohort were reexamined at 4.5 years of age or older for ocular alignment and stereopsis. RESULTS: Among infants who initially had constant esotropia > or = 40 PD, 0 of 45 children in the RFSW cohort and 0 of 21 children in the ESCET cohort showed resolution to orthophoria. In addition, only 2 infants showed a reduction in angle of deviation below 40 PD (one to 35 PD and one to 20 PD). Resolution to orthophoria was noted in a few infants who initially had small angle or variable angle esotropia. On follow-up at 4.5 years of age or greater, 91% of the children in the RFSW cohort had alignment within 8 PD of orthoposition and 30% had stereoacuity of 3000" to 60". Children who underwent surgical alignment at 6 months of age had a higher prevalence of coarse stereopsis than children who underwent alignment at 7 to 15 months of age. CONCLUSIONS: Taken together, these results suggest that infants who present at 2 to 4 months of age with constant esotropia of 40 PD or greater are valid candidates for surgical treatment. In addition, data from long-term follow-up support the hypothesis that early surgical alignment may promote the development of at least coarse stereopsis in these infants. PMID- 10532718 TI - Surgical management of intermittent exotropia with high AC/A ratio. AB - BACKGROUND: A subgroup of patients with intermittent exotropia have a high AC/A ratio, which places them at risk for esotropia at near after surgical treatment of their distance deviation. METHODS: A retrospective review of six patients with intermittent exotropia and a high AC/A ratio who were simultaneously treated with lateral rectus recessions to fully correct their exotropia at distance and placement of posterior fixation sutures on both medial rectus muscles in hopes of preventing an esotropia at near after surgery. RESULTS: Despite the persistence of a high postoperative AC/A ratio as assessed by the gradient method, five of six patients achieved stable postoperative alignment at distance and near without bifocals. One patient required postoperative bifocal correction for intermittent esotropia at near. CONCLUSION: In patients with intermittent exotropia and a high AC/A ratio, posterior fixation of the medial rectus muscles at the time of lateral rectus recessions enables the surgeon to fully correct the distance deviation and minimizes the risk of postoperative esotropia at near. PMID- 10532719 TI - Bilateral recession of superior rectus muscles: its influence on A and V pattern strabismus. AB - PURPOSE: To determine whether bilateral superior rectus recession modifies A and V pattern strabismus. PATIENTS AND METHODS: Three patients with V patterns and eight patients with A patterns underwent bilateral superior rectus recession with neither oblique muscle surgery nor vertical displacement of the horizontal rectus muscles. Another three patients with A patterns underwent simultaneous superior oblique tenotomy and superior oblique recession. RESULTS: Two of the patients with V patterns demonstrated an increase in the V pattern after surgery; one was unchanged. Five of the patients with A patterns converted to V patterns after surgery. In the remaining three patients the pattern was eliminated. The three patients who underwent superior oblique tenotomy along with superior rectus recession had large shifts toward V pattern (mean shift, 40 delta). CONCLUSION: Bilateral superior rectus recession tends to increase V patterns and reduce A patterns. Superior rectus recession may be synergistic with superior oblique tenotomy in collapsing an A pattern. PMID- 10532720 TI - Results of a combined adjustable recession and posterior fixation suture of the same vertical rectus muscle for incomitant vertical strabismus. AB - PURPOSE: The posterior fixation suture (fadenoperation) is an effective treatment for complicated incomitant vertical strabismus. Traditional operative methods do not permit the simultaneous use of an adjustable recession of the same muscle. METHODS: Seven patients with incomitant vertical strabismus and diplopia were treated with a combined adjustable recession and posterior fixation suture of the same vertical rectus muscle. Preoperative vertical misalignments in the primary position ranged from 4 to 10 prism diopters. Vertical incomitance from the primary position into the field of action of the recessed vertical rectus muscle ranged from 6 to 30 prism diopters (mean, 17 prism diopters). This was the sole operation in five patients and was combined with other vertical muscle surgery in two others. RESULTS: All seven patients experienced improvement in their diplopia. Five of 7 patients (71%) required postoperative adjustments to achieve orthophoria in the primary position. This combined procedure reduced large deviations in the field of action of the recessed vertical muscle in all cases. Six of 7 patients (86%) did not require prismatic correction after this operation. One patient required prism only in his reading glasses. CONCLUSIONS: A combined adjustable recession and posterior fixation suture of the same vertical rectus muscle was effective in reducing or eliminating vertical incomitant strabismus. PMID- 10532721 TI - Comparison of anterior transposition and recession of the inferior oblique muscle in unilateral superior oblique paresis. AB - PURPOSE: Both anterior transposition and graded recession have been shown to be effective procedures in weakening the inferior oblique muscle. Anterior transposition may work in part by converting the inferior oblique muscle from an elevator to a depressor of the globe. In theory, this would be useful in treating the inferior oblique overaction associated with superior oblique paresis. We compared inferior oblique recession and anterior transposition for the surgical correction of Knapp's class III unilateral superior oblique paresis. METHODS: Four patients underwent 14 mm recession, and five underwent anterior transposition of the inferior oblique muscle for the hypertropia in superior oblique paresis. Prism cover test measurements were made in all cardinal fields of gaze and were compared before and after operation between the two groups. RESULTS: The mean preoperative hyperdeviation in the primary position was 12 prism diopters in the recession group and 15 prism diopters in the anterior transposition group. The mean postoperative hyperdeviation was 1 prism diopter in the recession group and 3 prism diopters in the anterior transposition group. Postoperative results in the inferior oblique field of action demonstrated a mean 3 prism diopter hypertropia in the recession group and a 2 prism diopter hypotropia in the anterior transposition group. CONCLUSIONS: Anterior transposition and graded recession gave similar results in correcting the primary position hyperdeviation in Knapp's class III superior oblique paresis. Both procedures also markedly improved the hyperdeviation in the field of action of the inferior oblique muscle and superior oblique muscle. However, anterior transposition was more likely to result in postoperative hypodeviation in upgaze. PMID- 10532722 TI - Vertical latent nystagmus component and vertical saccadic asymmetries in subjects with dissociated vertical deviation. AB - PURPOSE: This study was conducted to quantify the vertical component of a latent nystagmus observed in subjects with dissociated vertical deviation (DVD), as well as to provide further evidence for vertical saccadic asymmetries in these individuals. METHODS: Binocular eye movements of subjects with DVD were recorded in two dimensions using a noninvasive video-based eye tracker while cover/uncover tests, alternate cover tests, and vertical saccades were performed. RESULTS: A small amplitude (1.5 degrees or less) vertical component of latent nystagmus can be observed in some subjects with DVD and is larger in the deviating eye than in the viewing eye. The frequency of the vertical nystagmus component is the same in each eye for any given fixation condition but may change depending on which eye is fixating. DVD in the presence of a vertical component of latent nystagmus can be adequately modeled by the algebraic sum of an exponentially decreasing velocity DVD and a nystagmus with an exponentially decreasing slow phase velocity. In general, the occluded eyes of DVD subjects make smaller downward saccades than the viewing eyes. CONCLUSIONS: It is possible but not obligatory that DVD subjects will have a vertical component of latent nystagmus. Algebraic summation of an exponentially decreasing velocity DVD and a vertical component of latent nystagmus provides a more parsimonious explanation of the observed saccadic eye movements than modeling the DVD itself as a combination of vergence and saccadic movements. Subjects with DVD show a range of saccadic yoking from nearly complete saccadic conjugacy to nearly complete dissociation. PMID- 10532723 TI - Medical treatment of pediatric subperiosteal orbital abscess secondary to sinusitis. AB - BACKGROUND: Subperiosteal abscess may accompany orbital cellulitis secondary to sinusitis. Common surgical principles include incision and drainage of all abscesses. Previous evidence suggests that some orbital abscesses may be treatable with intravenous antibiotics, especially in young children. Children's hospital records were reviewed to determine which abscesses may be treated medically. PATIENTS AND METHODS: Records of patients admitted for orbital cellulitis from 1993 to 1996 were reviewed. Patients with subperiosteal abscess on CT scan were included. Clinical outcomes for initial surgical versus medical management of medial abscesses were compared. Differences in age, hospital stay, and intracranial involvement were analyzed for medial versus nonmedial abscesses. RESULTS: All patients had abscesses adjacent to infected sinuses. Eighteen young children had medial abscesses. Twelve of 13 were cured by initial medical treatment; 4 of 5 underwent successful initial drainage. Outcomes were not statistically different (P > .490). Seven children with nonmedial abscesses were older (P < .001) and had more complicated courses than those with medial abscess. Three of 6 children with superior orbital abscess also had intracranial abscess. Intracranial complication was more likely with superior versus medial orbital abscess (P < .01). CONCLUSIONS: Medial subperiosteal orbital abscesses secondary to sinusitis in children 6 years of age and younger are highly amenable to treatment with intravenous antibiotics. Older children and children with nonmedial abscesses may have more complicated infections. Children with superior orbital abscesses are at higher risk for intracranial abscess. PMID- 10532724 TI - Visual prognosis of Coats' disease. AB - PURPOSE: Our purpose was to determine the visual prognosis of retinal telangiectasia (Coats' disease). METHODS: We performed a retrospective review of 35 patients with Coats' disease seen at the Hospital for Sick Children, Toronto, Canada between 1987 and 1996. Ten patients were excluded because of incomplete records. Treatment modalities consisted of no treatment, cryotherapy with and without 532 nm laser through the indirect ophthalmoscope, and enucleation. Visual outcome was determined where possible. RESULTS: Median follow-up was 4.5 years. Deterioration in visual acuity was associated with the presence of greater than 5 clock hours of involved retina and retinal detachment at diagnosis. Final visual acuity did not correlate with age of onset of disease. No eye treated with cryotherapy progressed to retinal detachment. CONCLUSIONS: Aggressive treatment of Coats' disease with cryotherapy with or without 532 nm laser, before retinal detachment, is likely to stabilize vision and decrease the risk of future total retinal detachment. PMID- 10532725 TI - A comparison of cryotherapy versus diode laser retinopexy in 100 consecutive infants treated for threshold retinopathy of prematurity. AB - PURPOSE: The purpose of this paper is to present a series of patients who were treated for threshold retinopathy of prematurity with either cryotherapy or diode laser. Complications and unfavorable outcomes during the first year after treatment will be compared for the two procedures. METHODS: The clinical courses of a consecutive series of 100 infants (192 eyes) were reviewed. All infants had their threshold status confirmed by a second examiner. Infants were treated with cryotherapy through 1993 and with diode laser thereafter. One hundred two eyes of 54 patients were treated with cryotherapy. Ninety eyes of 46 patients were treated with laser retinopexy. Two of the patients who were treated with laser (4 eyes) did not survive to the 3-month follow-up visit, and their results are not included here. The two groups of infants were comparable in their birth weight, adjusted gestational age at treatment, and severity of disease as determined by zone and sectors of stage 3 retinopathy of prematurity. RESULTS: Unfavorable outcome (total retinal detachment) was seen in 25.4% of eyes treated with cryotherapy (26 of 102), as compared with 15% of eyes treated with laser (13 of 86). Two cataracts were seen in our patients: one patient 22 weeks after cryotherapy, and the other 7 months after diode laser. CONCLUSIONS: No statistically significant difference was found in the rate of retinal detachments in the two groups (X2 = 3.05; P = .08). PMID- 10532726 TI - The efficacy of goniotomy/trabeculotomy in early-onset glaucoma associated with the Sturge-Weber syndrome. AB - PURPOSE: To assess the efficacy of goniotomy/trabeculotomy as the initial surgical procedure in early-onset glaucoma associated with Sturge-Weber syndrome. METHODS: We retrospectively analyzed 16 eyes of 14 consecutive patients with Sturge-Weber syndrome-associated glaucoma diagnosed before 4 years of age. All subjects were seen at a single institution from 1978 to 1996 and underwent goniotomy or trabeculotomy as their initial surgical procedure. RESULTS: Twelve eyes underwent initial goniotomy, and 4 eyes underwent initial trabeculotomy. One subject was lost to follow-up after surgery, resulting in 15 eyes for analysis. Of the initial goniotomy eyes, two thirds required a second surgical procedure. In the initial trabeculotomy eyes, half required a second procedure. Intraocular pressure was controlled (intraocular pressure < or = 22 mm Hg) in 66.7% of the eyes (10 of 15) after one or more goniotomy or trabeculotomy procedures for a median follow-up of 5.4 years (range, 1.4 to 15 years). For eyes with only one surgical procedure, 4 of 6 eyes had controlled intraocular pressure over a median follow-up of 3.4 years (range, 3 to 12 years). Seven of the 9 eyes that required more than one procedure had controlled intraocular pressure after all procedures over a median follow-up of 4.5 years (range, 1.4 to 15 years). CONCLUSION: Initial or repeated goniotomy or trabeculotomy may be an effective management choice for treatment of glaucoma associated with Sturge-Weber syndrome presenting in early childhood. PMID- 10532727 TI - Uric acid in the aqueous humor and tears of retinoblastoma patients. AB - PURPOSE: Malignancy can be associated with high levels of catabolic products. We performed a two-part study. Part 1 measured levels of uric acid and xanthine in the aqueous humor of eyes with malignant and nonmalignant diagnoses. Part 2 measured the levels of uric acid in tears of retinoblastoma patients. If compounds in high concentrations inside the eye could be detected outside the eye, via diffusion, in high concentrations in the tears, then a tear screening test for retinoblastoma could be developed. METHODS: High-performance liquid chromatography measured levels of uric acid and xanthine in aqueous humor samples of patients with retinoblastoma, melanoma, Coats' disease, adult cataract, and congenital cataract. Tear sampling was performed on patients with retinoblastoma and on normal eyes, and samples were assayed for uric acid. RESULTS: Part 1--Uric acid was elevated in the aqueous humor of eyes with retinoblastoma, melanoma, and Coats' disease compared with eyes with cataracts. Xanthine was elevated in retinoblastoma and Coats' disease and was lower in adult and congenital cataracts and melanoma. Part 2-No significant difference was found in the concentrations of uric acid in the tears of patients with retinoblastoma and those of normal patients. CONCLUSIONS: High levels of uric acid and xanthine present in the aqueous humor of patients with malignancy are consistent with the destructive nature of these conditions. Although uric acid was not elevated in the tears of retinoblastoma patients, continued investigation into substances that might be measurably different in the tears may yield a useful screening test in the future. PMID- 10532728 TI - A new school-based program to provide eyeglasses: childsight. AB - OBJECTIVE: To address the unmet need for glasses encountered in an urban school setting by developing and implementing a school-based, cost-effective program that provides appropriate spectacle correction to needy children. METHODS: A total of 5851 students 9 to 15 years of age in 4 middle schools in northern Manhattan were screened for vision. Those with vision worse than 20/40 were examined, given glasses if appropriate, or referred for additional evaluation. RESULTS: Of the 5851 children screened, 1614 (28%) had a failing result, with visual acuity less than 20/40 in the worse eye. Of this group, 1082 were given glasses that were assembled at the school within 1 hour of testing. Ten percent of the group that required glasses already had them, and the remaining were referred for a complete ophthalmic examination that was completed in 58 cases. Only 14 of these had vision loss unrelated to refractive error. CONCLUSIONS: The program successfully treated 88.3% of the children within the school who needed glasses. Given that only 10% of children who needed glasses had them, it indicates a huge need to provide glasses to at least a million children in this age group in the United States. PMID- 10532729 TI - Cyclic esotropia after a traumatic sixth nerve palsy in a child. AB - Cyclic esotropia is a rare phenomenon in which esotropia and orthophoria alternate over a period of 48 to 96 hours. The mechanism that underlies the phenomenon is unknown. Cyclic esotropia often occurs after a fusion-disrupting event. We report an unusual case of cycling esotropia with onset after a traumatic sixth nerve palsy. The cyclic phase persisted for 2 years, following a 48-hour alternate-day pattern. After strabismus surgery for the esotropic angle, the deviation disappeared and the patient remained orthotropic, with 1 year of follow-up to date. PMID- 10532730 TI - Unifocal Langerhans' cell histiocytosis in the clivus of a child with abducens palsy and diplopia. AB - Histiocytosis X, described by Lichtenstein in 1953, is an uncommon disorder that is characterized by an abnormal proliferation of Langerhans' cells. The Langerhans' cell normally occurs in the epidermis and T-cell-dependent areas of lymph nodes and functions as a macrophage. Histiocytosis X is predominantly a disease of childhood but can occasionally be seen in adults. Eosinophilic granuloma of the skull is the most common presentation of the disease, and the associated symptoms depend on the location of the lesion. It has been reported to occur in the temporal bone, including the petrous apex. We present the first reported case, to our knowledge, of eosinophilic granuloma, or unifocal Langerhans' cell histiocytosis, in the clivus of a child. PMID- 10532731 TI - A technique for repair of iridodialysis in children. AB - Iridodialysis can occur with blunt or penetrating trauma or inadvertently during intraocular surgery. A small dialysis may not need treatment. A larger iridodialysis that causes polycoria and diplopia, or a grossly eccentric pupil, needs to be reapproximated. A number of surgical techniques for repair, using double-armed polypropylene suture, have been reported. The suture is either left external on the surface of the sclera with the knot buried, covered with a triangular scleral flap, or retrieved with special forceps and buried in a scleral "groove." These techniques have also been used to allow posterior fixation of intraocular lens implants in the absence of capsular support; a complication of this approach is suture erosion through sclera, conjunctiva, or both. The pediatric sclera is softer and more elastic than adult sclera, and surgical repairs must last for decades. Concern about late suture erosion, and a desire for minimal scleral manipulation, led me to develop a simple technique for iridodialysis repair using a scleral tunnel incision and double-armed 10-0 polypropylene suture. PMID- 10532732 TI - Angle-closure glaucoma after laser treatment for retinopathy of prematurity. AB - Laser photocoagulation has become the standard for treatment of retinopathy of prematurity. In general, it has been found to be a safe and effective means of retinal ablation. We report a case of angle-closure glaucoma in an infant after diode laser treatment for retinopathy of prematurity, which required bilateral surgical peripheral iridectomies. PMID- 10532733 TI - Retinoblastoma with acute lymphoblastic leukemia, polyposis coli, and multiple hamartomas. AB - It has long been recognized that compared with their age- and sex-matched controls, survivors of hereditary retinoblastoma have a considerably higher risk of the development of second malignancies (10% at 20 years and 15% at 30 years of follow-up), including osteosarcoma, leiomyosarcoma, melanoma, fibrosarcoma, and other rare spindle cell sarcomas. Patients with the nongenetic variety of retinoblastoma do not particularly seem to have an increased incidence of other malignancies than the general population. However, it should be noted that a child with unilateral disease carries a 15% chance of having germline mutation. The cumulative mortality rate from second malignancies was 1.5% at 40 years after unilateral retinoblastoma diagnosis and 26% for bilateral cases in a large survey of 1458 patients. A child with unilateral retinoblastoma, cafe au lait spots, hairy nevus, and grouped pigmentation of retina in the fellow eye is described who furthermore developed acute leukemia and polyposis coli. PMID- 10532734 TI - What is popcorn? PMID- 10532735 TI - Visual functions in college baseball pitchers and batters. PMID- 10532736 TI - Long-term results of botulinum toxin in consecutive and secondary exotropia: outcome in patients initially treated with botulinum toxin. AB - INTRODUCTION: Long-term ocular alignment can be difficult to achieve in patients with consecutive and secondary (sensory) exotropia, and botulinum neurotoxin A (BTXA) is a recognized alternative to surgery in this group. PATIENTS AND METHODS: We reviewed the results of 44 patients aged 15 to 77 years (mean 31 years) who underwent their first BTXA injections from 1989 to 1990. In 30% of cases the choice of toxin treatment was made by the patient. In the remainder BTXA was recommended by the clinician to assess the risk of postoperative diplopia. Thirty-three patients (75%) were consecutively exotropic and 68% of patients had had previous strabismus surgery. The mean preinjection deviation was 41 delta of exotropia (range 12 to 85 delta exotropia) and the minimum mean angle change after 1 injection was 27 delta (range 0 to 57 delta). The average number of injections was 3 (range 1 to 17). RESULTS: Of the patient group, 59% went on to strabismus surgery, 14% continued to attend for maintenance treatment, and 9% were discharged with a small, stable deviation. The remainder were either followed up elsewhere or failed to reattend. CONCLUSIONS: Botulinum toxin appears to be a satisfactory treatment for constant exotropia in patients at risk of postoperative diplopia who have undergone multiple operations but, because more than half the group went on to surgery, surgery as a first therapy may be preferable in uncomplicated cases. PMID- 10532737 TI - Effect of graded anterior transposition of the inferior oblique muscle on versions and vertical deviation in primary position. AB - INTRODUCTION: There are various methods for weakening the inferior oblique muscle; here we describe the results of a graded anterior transposition. METHODS: Charts of 21 children (37 eyes) who underwent graded anterior transposition of the inferior oblique muscle were reviewed. Graded anterior transposition consisted of reinsertion of the inferior oblique muscle at various points along the temporal aspect of the inferior rectus muscle; the more severe the overaction, the more anterior the placement of the new insertion. In all cases the new inferior oblique insertion line was oriented parallel to the inferior rectus muscle axis. We analyzed the preoperative to postoperative change in inferior oblique overaction (versions) and vertical alignment in primary position. RESULTS: Postoperatively, 18 of 21 patients had normal versions, 2 patients had -1 underaction of 1 eye, and 1 patient had +1 overaction of both eyes. Eleven patients (15 eyes) had a preoperative vertical deviation in primary position of 4 PD or more. Three of these patients had unilateral congenital superior oblique palsy and a preoperative hypertropia of 20 PD. They underwent unilateral graded anterior transposition with a mean postoperative vertical change of 18 PD. Three patients had asymmetric primary inferior oblique overaction with true hypertropia, 1 patient had amblyopia and primary inferior oblique overaction, and 4 patients had dissociated vertical deviation associated with inferior oblique overaction. All patients had improvement after surgery, with no significant vertical deviation in primary position. CONCLUSIONS: Graded anterior transposition of the inferior oblique muscle is effective in normalizing versions and correcting vertical deviations in primary position. PMID- 10532738 TI - Dysversion of the optic disc and axial length measurements in a presumed amblyopic population. AB - PURPOSE: Our purpose was to evaluate anatomic variations of eyes presumed to be amblyopic. METHOD: Computer imaging and photography of the optic discs of 205 amblyopic subjects were performed and the axial lengths of 183 of the subjects were measured. The paired optic nerve images were evaluated for symmetry of disc contours and orientation of central blood vessels to detect optic nerve head dysversion. Dysversion of the optic nerve head, which is also referred to as segmental hypoplasia, is a congenital disorder characterized by the central retinal vessels emerging temporal to the vertical midline of the disc and being directed nasally or the nerve head tilting in a vertical direction resulting in a downward or oblique tilting of the discs with the blood vessels emerging at the superior or inferior disc rim. RESULTS: Ninety-three subjects had optic nerve dysversion. There was a greater degree of anisometropia (P< or =.004) in subjects with dysversion (anisometropia factor of 2.51+/-2.15) than in the subjects with symmetric discs (anisometropia factor of 1.76+/-1.63). Axial lengths of the amblyopic eyes were significantly smaller (P<.0001) than those of the nonamblyopic eyes. There was no statistical difference (P< or =.879) in length between amblyopic eyes with dysversion and those with symmetric discs. CONCLUSION: Optic disc dysversion was identified in 45.4% of patients who were previously assumed to be amblyopic. There are anatomic malformations in the eyes of a significant proportion of the presumed amblyopic population. This suggests that, in these individuals, congenital peripheral factors rather than impaired cortical development may be responsible for decreased unilateral acuity. PMID- 10532739 TI - Polydiaxonon prevents adhesions in the rabbit model: a pilot report. AB - PURPOSE: Restrictive strabismus is a common and difficult problem confronted by strabismologists. Several materials have been used to minimize the formation of adhesions after strabismus surgery with varying degrees of success. Polydiaxonon (PDS, Ethicon) is an absorbable, nontoxic, nonporous material. We used it as 25 and 50 microm thick sleeves to study its effectiveness in the prevention of adhesions. METHOD: Eight eyes of four adult New Zealand White rabbits were used. To simulate the adhesions, a raw scleral bed was created under the superior rectus insertion in study animals and the muscle capsule facing the sclera was opened. Four study eyes had PDS sleeves inserted around the superior rectus; the other four served as controls. After 4 months the animals were killed. The surgical sites were inspected for adhesions. Light microscopy was also performed. RESULTS: Virtually no adhesion formation was noted in the study eyes. In the control group, however, dense adhesions were seen. Light microscopy confirmed these results. No significant amount of foreign material was found. There was no toxicity resulting from PDS. CONCLUSIONS: This demonstrated nearly complete prevention of adhesions in the rabbit model. PDS sleeves appear to have potential in surgery for restrictive strabismus. PMID- 10532740 TI - Relationships among visual acuity demands, convergence, and nystagmus in patients with manifest/latent nystagmus. AB - BACKGROUND: We investigated the role convergence plays in nystagmus dampening, in particular, relationships among visual acuity demands, convergence, and nystagmus. Previously we showed that subjects with idiopathic infantile nystagmus exhibit a range of responses to acuity targets, one of which is nystagmus blockage syndrome. We report herein eye movement responses to acuity targets of patients with manifest/latent nystagmus. METHODS: Fourteen patients, 11 with latent or manifest latent nystagmus and 3 with combined manifest latent with infantile nystagmus, were asked to indicate the direction of the gap in Landolt C optotypes while their eye movements were recorded. RESULTS: The tested patients exhibited various responses to acuity demands: (1) dampening of nystagmus with convergence (i.e., nystagmus blockage syndrome) (5/14 patients), (2) changes in vergence without nystagmus dampening (2 patients), (3) decrease of nystagmus without convergence (2 patients), and (4) little change in nystagmus or vergence (5 patients). In nystagmus blockage syndrome the amount of convergence increased with acuity demands in two of five patients and the convergence duration in four of five patients; nystagmus dampening increased with acuity demands in one of five patients and the blockage duration in four of five patients. CONCLUSIONS: Many, but not all, patients with manifest/latent nystagmus, similar to those with infantile nystagmus, used convergence to dampen their nystagmus. The convergence response tended to increase with acuity demands, but the amount of dampening was idiosyncratic and not predictably related to the measured convergence across patients. PMID- 10532741 TI - Rectus muscle recession and resection without scleral sutures. AB - PURPOSE: To eliminate the risk of scleral perforation during strabismus surgery in susceptible patients, we introduce a technique to allow predictable rectus muscle recession and resection without the placement of scleral sutures. METHODS: Three patients with thin sclera underwent rectus muscle surgery by use of a double-arm suture technique that avoids placement of sutures directly into the sclera. Two of the patients had esotropia and underwent bilateral lateral rectus muscle resections and a unilateral recess/resect operation, respectively. One of the patients had exotropia and underwent bilateral lateral rectus muscle recessions. RESULTS: All three patients achieved postoperative alignment to within 15 PD of orthotropia and had no evidence of slipped or lost muscle when examined 2 months postoperatively. The appearance of the ocular surface was excellent in all three cases. CONCLUSIONS: Predictable and secure rectus muscle recession and resection can be performed without the placement of scleral sutures in patients in whom scleral suturing may be hazardous. PMID- 10532742 TI - Open globe injuries in children. AB - PURPOSE: A retrospective review of open globe injuries in children was performed to identify the common types of injury and to correlate features of the injuries and surgical management with visual prognosis. METHODS: The hospital records of 70 patients were reviewed to determine demographic data, the nature and location of the injuries, the anatomic and functional status of the injured eye before the initial repair, the details of all primary and subsequent surgical procedures, and the final visual outcome. RESULTS: Fifty of the patients studied (71%) were male and 20 (29%) were female. The average age of the patients was 5 years. Sharp objects caused the majority of injuries (67%). Most of the injuries happened at home (72%). The cornea was involved in a majority of the injuries (92%). Thirty two eyes (46%) required only primary repair; 15 eyes (21%) required primary repair with anterior vitrectomy and primary lensectomy; 17 eyes (24%) underwent secondary lensectomy or vitreoretinal surgeries, and 5 eyes (7%) were enucleated. Visual acuity of 20/40 or better was achieved by 45% of those patients who required only primary repair. Of those patients requiring a second procedure, 19% had a visual acuity of 20/40 or better. Initial clinical findings associated with an unfavorable visual outcome were retinal detachment, relative afferent pupillary defect, vitreous hemorrhage, and hyphema. CONCLUSION: The prognosis after an open globe injury in children is strongly influenced by the nature of the injury and the extent of the initial damage. Visual outcome is better in eyes that require only primary repair. PMID- 10532743 TI - Quantitative forced ductions in an animal model--characterization of passive forces. AB - PURPOSE: Our purpose was to characterize the passive tissue forces involved in ocular rotation in a controlled animal model and to evaluate the influence of manual versus mechanized ductions, repeated measurements, speed of rotation, and the influence of the nondepolarizing muscle relaxant mivacurium. METHODS: Forced ductions were performed under general anesthesia on 20 eyes of 10 pigs, with or without mivacurium, with use of a highly sensitive force gauge attached to the eye by a traction suture. The eye was moved either manually or at constant speed with a motorized platform. Eyes were rotated a total of 8 mm from their resting position under anesthesia. The force-displacement relationship was analyzed and compared between groups. RESULTS: A linear (elastic) relationship between force and displacement was noted, with a slope of 0.4 g per degree with use of the mechanized technique. Neither speed of rotation, use of mivacurium, nor repeated ductions significantly influenced the shape or slope of the relationship. Hysteresis averaged 2 to 4 g. Measurements performed with use of the motorized platform showed significantly improved reliability over those made manually. CONCLUSIONS: The passive length-tension data correlate well with data reported by others in humans. Within a wide range of eye movement, this force is elastic in nature. For relatively low angular velocities, such as might be produced in smooth pursuit, the passive forces do not change appreciably with changes in velocity. The nondepolarizing muscle relaxant mivacurium has no effect on accurate performance of passive forced ductions under general anesthesia. Studies collecting quantitative data on passive orbital forces should be performed, when feasible, with an automated duction and recording apparatus. PMID- 10532744 TI - Restricted ocular motility after orbital trauma--studies with an animal model. AB - OBJECTIVE: Our purpose was to develop a quantitative model of restricted ocular motility (fat adherence syndrome) in the pig orbit to facilitate research into pathogenesis and treatment. METHODS: Twenty eyes of 10 pigs were used in an attempt to create a fibrous adhesion between either the inferior rectus or the medial rectus muscle and the adjacent periorbita. Quantitative forced ductions were performed preoperatively and 6 weeks postoperatively with an electronic force gauge. RESULTS: Forced ductions displayed a linear relationship to displacement over the range tested, both preoperatively and postoperatively. Although increased resistance to forced ductions was produced in 10 of the 20 (50%) eyes, the average changes were not statistically significant. The absolute change in force (Postoperative--Preoperative) was linearly related to the amount of ocular rotation. Reproducibility of measurements was markedly improved by the use of a motorized forced duction apparatus compared with manual rotation. CONCLUSIONS: The results from this study, and those from previous work, suggest that the classic fat adherence syndrome is an uncommon event, even after significant soft tissue injury in the orbit. Further studies are needed to more completely define the risk factors and pathogenesis of the fat adherence syndrome and the suitability of the pig orbit as a model. PMID- 10532745 TI - Neonatal asteroid hyalosis. AB - Asteroid hyalosis, noted in 0.83% of routine eye examinations, is uncommon in younger patients and is more frequently seen in patients more than 60 years old. It has been considered to be related to an aging process, and when it occurs in younger patients, ocular disease is typically associated. We report a 4-week-old patient with Down syndrome and bilateral congenital cataracts who had unilateral asteroid hyalosis. PMID- 10532746 TI - Pediatric Horner syndrome and neuroblastoma threat. PMID- 10532747 TI - The Philip Knapp Lectureship. PMID- 10532748 TI - Slipped and lost muscles and other tales of the unexpected. Philip Knapp Lecture. AB - Loss of a rectus muscle may occur as a rare complication of strabismus surgery. In addition, extraocular muscles may become traumatically detached from the globe when they rupture or are transected as the result of an injury or during the course of retinal detachment or paranasal sinus or orbital surgery. Although the clinical features of a slipped muscle may resemble those of a lost muscle, the findings at the time of reoperation are distinct. Also the etiology, and therefore the prevention, of a slipped muscle differs from that of a lost muscle. It is likely that slipped muscles and even some lost muscles are underdiagnosed and represent a significant cause of unexpected overcorrection or undercorrection. Unless the displaced muscle is appropriately advanced, it can be extremely difficult to correct the associated strabismus, yet locating and repairing either a lost or slipped muscle can be challenging and is by no means always successful. Current concepts pertaining to the etiology, recognition, and management of slipped and lost muscles will be discussed. PMID- 10532749 TI - Large-angle exotropia corrected by intraoperative botulinum toxin A and monocular recession resection surgery. AB - PURPOSE: Surgical correction of large-angle exotropia, greater than 70 PD, traditionally requires operating on three or four horizontal muscles. However, in secondary exotropia from monocular visual loss, it is advisable to operate only on the eye with poor vision. We used intraoperative botulinum toxin as an adjunct to the monocular recession-resection procedure for large-angle sensory exotropia, therefore operating only on the visually impaired eye. METHODS: Three patients underwent monocular recession (10 mm) and resection (10 mm) along with intraoperative botulinum toxin A injection of 10 units into the recessed muscle. All had desired cosmetic repair of long-standing large-angle exotropia (range 100 to 110 PD) with amblyopia and vision worse than 20/200 in the deviated eye. RESULTS: Within 4 days after operation all patients demonstrated maximal paresis of the lateral rectus muscle. This lasted 8 to 12 weeks and resulted in stable orthotropia at 2.5 years in case 1 and stable 8 PD exotropia at 4 years in case 2. The third case demonstrated a stable 18 PD exotropia by 7 months with a satisfactory cosmetic result. CONCLUSION: This technique provides an alternative for the surgical correction of large-angle exotropia by operating only on two horizontal muscles. In sensory exotropia it also avoids subjecting a normal eye to an operative risk. PMID- 10532750 TI - Results of superior oblique posterior tenotomy. AB - PURPOSE: The purpose of the study was to evaluate the results obtained by the superior oblique posterior tenotomy close to its insertion in patients with A pattern strabismus and vertical strabismus who had bilateral or unilateral superior oblique overaction. METHODS: We retrospectively analyzed the results of superior oblique posterior tenotomy close to its insertion performed bilaterally in 13 consecutive patients with A-pattern strabismus and unilaterally in 14 consecutive patients with hypotropia performed between March 1989 and October 1996. RESULTS: The mean preoperative A-pattern deviation was 25.31 PD and the mean postoperative A-pattern deviation was 3.23 PD with a mean reduction of 22.08 PD. The mean follow-up for the above group was months (range 3 to 58 months). In the hypotropia group the mean preoperative vertical deviation was 11.07 PD and the mean postoperative vertical deviation was 4.28 PD with a mean reduction of 6.78 PD. The mean follow-up for this group was 16.21 months (range 3 to 72 months). CONCLUSIONS: Superior oblique posterior tenotomy selectively weakens its vertical action. Bilateral weakening showed marked and consistent improvement in A-pattern deviation and unilateral weakening showed definite but less consistent reduction in deviation in cases of hypotropia with superior oblique overaction. PMID- 10532751 TI - Needle sterility during strabismus surgery. AB - BACKGROUND: Infection after strabismus surgery is uncommon and its cause remains unanswered. The source of the bacteria and the manner in which it enters the eye is often unknown. Most pediatric ophthalmologists now use 5% povidone-iodine to reduce the bacterial population before surgery. The needles used during strabismus surgery may be a source of bacterial contamination. METHODS: One hundred six needles were cultured after their use in strabismus surgery. RESULTS: Sixteen of 106 needles (15.1%) and 15 of 61 cases (24.6%) were culture positive. The organisms recovered closely resembled indigenous bacterial flora. CONCLUSION: This study suggests that the needles used during strabismus surgery may be the source of bacteria that can lead to infections after strabismus surgery. PMID- 10532752 TI - Asymmetric motion visually evoked potentials in infantile strabismus are not an artifact of latent nystagmus. AB - BACKGROUND: Patients who have infantile strabismus exhibit a directional asymmetry of motion visually evoked potentials (MVEPs) recorded under conditions of monocular viewing. The majority of these patients also have latent nystagmus, raising the possibility that the MVEP asymmetry is an artifact of the nystagmus. To explore this issue, we correlated MVEPs and eye movements under conditions that eliminated or increased latent nystagmus. METHODS: MVEPs and eye movements were recorded under conditions of monocular viewing in three adults who had combined infantile-onset strabismus and latent nystagmus. The subjects viewed vertically oriented grating stimuli that oscillated horizontally at temporal frequencies of 6.6 to 11.0 Hz by use of spatial frequencies of 1 to 3 cycles/degree. Quantitative eye movement recordings of latent nystagmus and horizontal pursuit/optokinetic nystagmus were also obtained. RESULTS: Eye movement recordings showed that the latent nystagmus was absent or markedly diminished when the viewing eye was in a 45-degree adducted position, whereas nystagmus velocity increased 10 to 40 times (to 2.2 to 4.5 degrees/second) when the viewing eye was in an abducted position (p < 0.05). MVEPs were abnormal (asymmetry indices > 0.40) when the viewing eye was in an adducted or abducted position of gaze. No correlation was found between the MVEP asymmetry index and the velocity of latent nystagmus. CONCLUSIONS: MVEP asymmetries in infantile strabismus remain robust under conditions that eliminate or greatly reduce the oscillations of latent nystagmus. MVEP asymmetries and ocular motor abnormalities both characterize infantile strabismus, but the ocular motor defects do not cause the MVEP asymmetries. The nasotemporal asymmetry of MVEPs and the nasotemporal asymmetry of pursuit and latent nystagmus are likely caused by deficits in related but separate binocular visual cortical circuits. PMID- 10532754 TI - Visual outcomes after surgery for unilateral cataract in children more than two years old: posterior chamber intraocular lens implantation versus contact lens correction of aphakia. AB - PURPOSE: We sought to determine whether posterior chamber intraocular lens implantation yields better visual acuity and binocular vision than does conventional contact lens correction of aphakia in similar groups of pediatric cataract patients. METHODS: We reviewed the medical records of children aged 2 to 16 years who had unilateral cataract surgery by a single pediatric ophthalmologist between 1986 and 1996. Before 1992 all patients underwent standard lensectomy with vitrectomy. Beginning in 1992 posterior chamber intraocular lens (IOL) implantation was offered as a choice to families and was performed on most patients. RESULTS: Monocular vision outcomes were not significantly different in 20 IOL and 31 lensectomy-vitrectomy patients, with 85% of the IOL group and 77% of the lensectomy-vitrectomy group showing better than 20/100 final acuity. Binocularity, however, was much better in the IOL group, with 90% demonstrating at least 400 seconds of arc stereopsis, as opposed to 39% in the lensectomy-vitrectomy group (p = 0.003). Subgroups of patients with traumatic or nontraumatic cataract origin, age at surgery less than 7 years, and preoperative visual acuity less than 20/100 compared very similarly. CONCLUSION: Posterior chamber IOL implantation appears to provide significantly better binocular function than conventional management of unilateral cataract in childhood but does not substantially improve visual acuity results. PMID- 10532753 TI - Pediatric Horner syndrome. AB - INTRODUCTION: The purpose of this study was to define the etiologies of Horner syndrome in the pediatric population. METHODS: A retrospective review was performed of the medical records of all pediatric Horner syndrome patients (< 18 years old) examined by the pediatric ophthalmology services at two large referral centers. RESULTS: Seventy-three pediatric Horner syndrome patients were identified. Of these, 31 (42%) were congenital, 11 (15%) were acquired without surgical intervention, and 31 (42%) were acquired after a surgical procedure of the thorax, neck, or central nervous system. Of the congenital Horner syndrome patients, a history of delivery with the use of forceps, vacuum extraction, shoulder dystocia, fetal rotation, or postterm delivery was elicited in 16 patients (53%). Concomitant brachial plexus injury was identified in only 3 patients. Two patients had congenital varicella syndrome and 1 patient was diagnosed with neuroblastoma. This patient had a palpable supraclavicular mass and stridor. Diagnosis of the patients with acquired Horner syndrome included neuroblastoma (2), trauma (1), rhabdomyosarcoma (1), brainstem vascular malformation (1), disseminated sclerosis (1), and not determined (5). CONCLUSION: In children with congenital Horner syndrome, a history of forceful manipulation of the infant during birth may reduce the need for extensive systemic evaluation. Without such history, a decision to proceed with further evaluation is made with consideration of the relative incidence of neuroblastoma by age and the physical findings. All acquired pediatric Horner syndrome patients without a known etiology require thorough evaluation because of the frequent association of serious underlying disease. PMID- 10532755 TI - Ocular disease in children with vertically acquired human immunodeficiency virus infection. AB - BACKGROUND: Clinical reports suggest that ocular disease in infants and children vertically infected with human immunodeficiency virus (HIV) is different from that in adults. Pediatric patients with acquired immunodeficiency syndrome (AIDS) and HIV infection are being treated more aggressively and are living longer, but current literature on the incidence of AIDS-related ocular disease in vertically acquired HIV infection is limited. METHODS: Thirty-three children with culture positive, vertically acquired HIV infections were prospectively followed with ophthalmic examinations between September 1991 and August 1996 at the University of Massachusetts. Patients were categorized as having symptomatic or asymptomatic HIV disease according to the U.S. Centers for Disease Control and Prevention guidelines. Absolute CD4 counts and other measures of immune function were obtained. RESULTS: The average length of follow-up was 30 months, and the average number of ophthalmic examinations per patient was 4.8. Ten patients developed ophthalmic findings. Nine of 18 (50%) patients with symptomatic AIDS disease developed ophthalmic findings. One of 15 asymptomatic HIV-infected patients developed ocular findings. Two patients with absolute CD4 counts less than 10 developed cytomegalovirus retinitis. CONCLUSIONS: These results suggest that AIDS related ophthalmic disease is less common in vertically infected children than in adult AIDS patients. It also supports intensified clinical surveillance for cytomegalovirus retinitis in children with end-stage disease and very low CD4 counts. PMID- 10532757 TI - The Tundra Swan strabismus retractor. PMID- 10532756 TI - Normal reading despite limited eye movements. AB - Many investigators have demonstrated that poor readers exhibit abnormal eye movements during reading. An association between defective vergence, accommodation, and poor reading skills has also been noted by some investigators. Children who are poor readers have been subjected to therapeutic interventions on the basis of the assumption that improving their eye movements, as part of a multifaceted program of "vision therapy," will yield commensurate improvement in reading performance. This approach has been controversial, and other authors have expressed opposing views. We report the ophthalmologic and reading assessments of two children with Mobius' syndrome who were average to above-average readers despite essentially absent horizontal eye movements. PMID- 10532758 TI - Repeatability of the hand-held Nidek auto-keratometer in children. AB - Measurement of the anterior corneal radius of curvature is important in the pediatric population for proper fitting of contact lenses, calculation of intraocular lens power in patients undergoing cataract surgery, and monitoring changes in shape of the cornea in keratoconus. Because astigmatism in preschool children is mainly the result of corneal asphericity, measurement of the anterior corneal radius of curvature may also be used in screening in children at risk for amblyopia. The aim of this study was to determine the repeatability of the hand held automated Nidek keratometer (Nidek Co. Ltd, Tokyo, Japan) and its ease of use in children. PMID- 10532759 TI - Anterior segment dysgenesis and congenital glaucoma associated with partial trisomy of chromosome 1 (1q32-qter). AB - A child born with partial trisomy of chromosome 1 (1q32-qter) survived and was seen for anterior segment dysgenesis and congenital glaucoma. Pure trisomy 1q is rarely seen in live-born infants and has not previously been described in association with congenital glaucoma. The genetic basis for glaucoma is complicated and multifactorial and probably determined by a number of genes on a variety of chromosomes. The current case provides some evidence that part of chromosome 1 may be involved with the etiology of a glaucomatous process. PMID- 10532760 TI - Childhood hemianopia: the bigger picture. PMID- 10532761 TI - Neurologic toxicity associated with interferon alfa treatment of capillary hemangioma. PMID- 10532762 TI - The effect of anterior transposition of the inferior oblique muscle on ocular torsion. AB - INTRODUCTION: The effects of anterior transposition of the inferior oblique on elevation in adduction have been studied, but changes in objective ocular torsion have not been investigated. METHODS: A prospective study on the effect of anterior transposition of the inferior oblique on objective torsion with use of fundus photography was undertaken in 24 eyes of 13 patients. The amount of oculartorsion was determined by measuring the angle formed by a horizontal line drawn across the geometric center of the disc and a second line connecting the geometric center to the foveola. RESULTS: The decrease in excyclotorsion at 6 to 8 weeks after surgery was 6.9+/-5.0 degrees (34%) (p = 0.006) and 2.8+/-4.4 degrees (13%) (not significant) after 10 weeks. An overall net change of 6.2+/ 4.8 degrees (33%) was obtained after anterior transposition of the inferior oblique adjacent or anterior to the inferior rectus insertion (p = 0.006). Transpositions done 1 to 3 mm behind the inferior rectus insertion showed a negligible torsional change. Torsion where inferior oblique function normalized after anterior transposition of the inferior oblique (8.5+/-2.9 degrees) was not different from control, whereas torsion where inferior oblique function recurred after surgery (15.9+/-7.2) was significantly different from control (p = 0.002). Regression analysis showed that only preoperative inferior oblique overaction and preoperative degree of torsion predicted the change in torsion after anterior transposition of the inferior oblique. CONCLUSION: Anterior transposition of the inferior oblique muscle initially decreased objective excyclotorsion, but the effect decayed beyond 10 weeks. At least in the short term only anterior transposition of the inferior oblique muscle done adjacent or anterior to the inferior rectus insertion affected torsion. PMID- 10532763 TI - Intraoperative traction testing to detect incomplete inferior oblique myotomy/myectomy. AB - PURPOSE: Incomplete transection of the inferior oblique muscle during myotomy or myectomy is an important preventable cause of ineffective inferior oblique weakening surgery. Intraoperative traction testing has been suggested as a means of detecting an incomplete inferior oblique transection. The usefulness of intraoperative traction testing to detect incomplete myotomy was evaluated. METHODS: The subjective "tightness" of the inferior oblique muscle was evaluated after partial and complete myotomy to determine how partial myotomy affected inferior oblique traction testing and to determine whether intraoperative traction testing is an effective means of ensuring that a complete myotomy has been achieved. Serial traction testing was performed on 10 inferior oblique muscles at the time of myotomy and scored, 0+ to 4+ tightness. Testing was performed before myotomy and after 50%, 90%, and total myotomy. RESULTS: In 8 of 10 eyes traction testing remained strongly positive until the inferior oblique muscle had been completely transected. The average inferior oblique muscle tightness of these eight muscles was +1.69 before myotomy and after 50% and 90% myotomy. The tightness decreased to 0 after 100% myotomy. In two cases no inferior oblique muscle could be detected after 50% and 90% myotomy, respectively. These two muscles had preoperative traction testing of +0.5 and +1.00, respectively. CONCLUSIONS: Intraoperative traction testing is a practical and effective method of detecting incomplete inferior oblique myotomy/myectomy, but the test should be interpreted with caution in patients with loose premyotomy muscles. PMID- 10532764 TI - Variants of congenital ocular motor apraxia: associations with hydrocephalus, pontocerebellar tumor, and a deficit of vertical saccades. AB - BACKGROUND: Congenital ocular motor apraxia (COMA) is characterized by the inability to generate volitional horizontal saccadic eye movements in the absence of other focal neurologic abnormalities. SUBJECTS: We report on two children (ages 5 months and 3 years) whose COMA did not adhere to these classic criteria. The children were followed up clinically with serial ocular motor examinations and neuroimaging over a period of 3 years. RESULTS: In the first child horizontal COMA was associated initially with neonatal communicating hydrocephalus. Two and one half years after the first signs of COMA, a fourth ventricle medulloblastoma appeared. The second child, who recovered from a periventricular hemorrhage caused by perinatal asphyxia, manifested vertical COMA and compensatory vertical head thrusts. CONCLUSIONS: COMA may be associated with hydrocephalus, pontocerebellar tumor, and periventricular hemorrhage. These rare variants of COMA emphasize that the eye movement deficits may arise from several locations, cerebral as well as pontocerebellar, in the neuronal pathways generating saccades. PMID- 10532765 TI - Anomalous head posture with early-onset homonymous hemianopia. AB - PURPOSE: We have noted the frequent finding of an ipsilateral head turn in children with early-onset homonymous hemianopia. We report a series of patients with these findings and propose a theory to explain this association. METHODS: Ten patients with early-onset homonymous hemianopia and anomalous head posture were examined. Head computed tomography and magnetic resonance imaging confirmed a cerebral lesion as the cause of the hemianopia in all patients. RESULTS: All patients had onset of central nervous system disease prenatally or before age 18 months. A head turn toward the visual field defect with a gaze preference contralateral to the visual field defect was present in all patients. CONCLUSIONS: Early-onset homonymous hemianopia should be included in the differential diagnosis of anomalous head posture. PMID- 10532766 TI - Poverty predicts amblyopia treatment failure. AB - OBJECTIVE: Clinical impressions suggested a hypothesis that poverty is associated with poorer results in amblyopia therapy. To test this hypothesis, we compared patients with amblyopia who had Medicaid assistance with those who did not. METHODS: Of 1272 patients recorded to have amblyopia in the eye center computer, 280 met inclusion criteria of first visit under age 10 years and had treatment instituted and visual acuities recorded then and at follow-up visits. Seventy-one had Medicaid assistance, and 209 did not. Age at first visit, age at final visit, severity of amblyopia as measured by visual acuity at the first visit, and number of visits were all statistically indistinguishable. A large difference in final visual acuity, number of missed visits, and the parent's estimate of compliance was found. RESULTS: The likelihood of good final visual acuity of 20/30 or better was 26.8% in the Medicaid group and 58% in the non-Medicaid group. The likelihood of a poor final visual acuity of 20/70 or worse was 33.8% in the Medicaid group versus 11.5% in the non-Medicaid group. CONCLUSION: These results established socioeconomic status, measured by qualification for Medicaid assistance, to be an important predictor for success for amblyopia therapy. Work is in progress to better understand more specific factors and to meet the therapeutic challenge of amblyopia therapy for children. PMID- 10532767 TI - The neurofibrovascular bundle of the inferior oblique muscle as the ancillary origin of that muscle. AB - PURPOSE: To establish that the neurovascular bundle (NVB) of the inferior oblique muscle has ligamentous qualities that enable it to function as an ancillary origin to the muscle, particularly after anterior transposition of its insertion. METHODS: Fresh cadaveric eyes and eyes of surgical patients were studied. Eighteen orbits were dissected to demonstrate the linear course of the NVB and its adjacent fibrous bands. Intact orbits were analyzed histologically, as were autopsy and surgical specimens, to evaluate the capsule of the NVB and the adjacent fibrous bands. The elastic modulus was measured in NVB specimens and in superior oblique tendons. Six eyes in which recurrent inferior oblique muscle overaction developed after an anterior transposition procedure were surgically explored to determine the structure that was serving as its ancillary origin. RESULTS: Gross anatomic and microscopic studies showed a linear orientation of the NVB,with adjacent fibrous bands anteriorly joining the inferior oblique and inferior rectus muscle capsules. The NVB showed about 50% fibrocollagenous capsule, with the collagen fibers aligned parallel to the NVB. The elastic modulus was highest (stiffest) in the NVB and lowest in the superior oblique tendon. In patients who had undergone anterior transposition operations, the NVB served as the ancillary origin of the inferior oblique muscle. CONCLUSION: The name of the NVB should be changed to neurofibrovascular bundle because it has a prominent fibrocollagenous capsule and is encased in fibrous tissue bands anteriorly. The neurofibrovascular bundle has a linear course and is relatively stiff. It binds the midposterior portion of the inferior oblique muscle posteriorly. Its ligamentous qualities enable it to function as an ancillary origin for the inferior oblique muscle. PMID- 10532769 TI - Strabismus surgery among aged medicare beneficiaries. AB - OBJECTIVES: The purpose of this study was to investigate the incidence of strabismus surgery among aged patients in the United States. METHODS: The Medicare Part B claims experience (physician professional fee billing) for 1995 was reviewed for the number of times each strabismus surgical procedure recognized in Physicians' Current Procedural Terminology (CPT) was performed. To determine the indications for the procedures that were performed, a 5% sample of claims was reviewed for the pertinent International Classification of Diseases, Ninth Revision, Clinical Modification, diagnostic codes. RESULTS: There were 27 million aged Medicare beneficiaries eligible for Part B benefits in 1995 in a fee for-service setting. During that year physicians reported 9497 strabismus physician services. These represented 6585 separate procedures (CPT codes 67311 to 67343) and 277 botulinum toxin (Botox) injections for strabismus (CPT 67345) performed during 1995. Sixty-nine percent of the surgical procedures were for horizontal correction and 28% were for vertical correction. Adjustable sutures were used for only 1240 cases (1 9%). The add-on procedural code for reoperation surgery or surgery in the presence of restriction of the extraocular muscles was used in just 930 cases (14%). The most common diagnosis for horizontal surgery was exotropia. Paralytic strabismus and thyroid disease were identified for 17% of cases. Three percent of the diagnoses were inappropriate for the procedures performed and may have been reported in error. CONCLUSIONS: These data confirm a very low incidence of strabismus surgical procedures (2/10,000) and injections (1/100,000) among aged Medicare beneficiaries. The strabismus surgery was most often performed to repair a horizontal deviation. The adjustable suture technique was used infrequently. These data may be extrapolated into the future to aid in determining the strabismus services that will be needed early in the next century. PMID- 10532768 TI - Recombinant interferon alfa-2b in the treatment of vision-threatening capillary hemangiomas in childhood. AB - INTRODUCTION: Hemangiomas of the orbit and eyelids may cause serious ocular problems usually related to amblyopia and astigmatism. Steroids have become the accepted treatment. However, some hemangiomas are resistant to steroids or require prolonged use,with unacceptable side effects. Interferon alfa-2b, an antiangiogenic protein, was used in this prospective study to treat visually threatening hemangiomas that were unresponsive to oral or intralesional steroid treatment. METHODS: Forty patients aged 2 to 36 months with life- or organ threatening hemangiomas were prospectively enrolled to evaluate the efficacy and safety of interferon alfa treatment for hemangiomas. Sixteen of these 40 patients had hemangiomas causing serious ocular dysfunction. The patients were treated with 3 x 10(6) U/m2 interferon alfa-2b subcutaneously daily for 3 months; treatment was then tapered or retreated according to response and protocol. Therapeutic responses were documented. RESULTS: Fifteen patients with ocular hemangiomas have finished treatment. The pretreatment volume measured by computed axial tomographywas an average of 22.3 cm3. Clinical response with eye opening was observed at an average of 6 weeks. There was a significant regression of the hemangioma in all patients, with an average 82% reduction in volume. Patients were treated with glasses and occlusion therapy as appropriate. Final visual acuities with a follow-up averaging 14 months after cessation of interferon treatment were normal, except that five of 15 patients had amblyopia; one of these patients had 20/40, two had 20/60, and two had 20/70. There were no major illnesses or serious adverse side effects. CONCLUSION: Interferon alfa-2b treatment resulted in good to excellent regression of all the hemangiomas. This regression was clinically significant,with patients able to open the affected eye an average of 6 weeks into treatment. Visual results were good, with moderate amblyopia occurring only in patients treated at a later age. Interferon alfa-2b was well tolerated by these young patients, and no significant illness or side effect has occurred. PMID- 10532770 TI - Extracorporeal membrane oxygenation causing asymmetric vasculopathy in neonatal infants. AB - BACKGROUND: Extracorporeal membrane oxygenation (ECMO) is a form of cardiopulmonary bypass therapy used in term or near-term infants with severe cardiorespiratory disorders not responsive to conventional intensive care interventions. An ECMO-associated retinal vasculopathy has been described with little reference to the specific condition of the patient. We examined the eyes of 91 infants who underwent ECMO treatment. An assessment was made of the following: (1) when retinal changes occurred, (2) whether there was a particular systemic disorder or ECMO approach associated with these retinal findings, and (3) whether there may be ocular sequelae from this development. METHODS: Ninety one neonates were treated with ECMO for meconium aspiration syndrome (MAS), primary persistent pulmonary hypertension of the newborn, sepsis, congenital diaphragmatic hernia (CDH), respiratory distress syndrome (RDS), and blood aspiration. Venoarterial bypass was performed in 73 patients. The remaining 18 patients underwent venovenous bypass. Ophthalmologic examinations were performed during bypass in 6 infants and within 3 weeks of ECMO in the remainder. RESULTS: Asymmetric retinopathy (left eye > right eye) was discovered in six infants with CDH and in one infant with RDS within a 2-week period after bypass, demonstrating venous tortuosity with or without intraretinal hemorrhages. One infant treated for MAS had a left eye intraretinal hemorrhage only. All patients with the noted retinal changes underwent venoarterial cannulation. Six of 9 patients with CDH had retinal findings noted compared with 1 of 10 patients with RDS and 1 of 35 patients with MAS. CONCLUSION: Because we were able to examine infants while they were receiving ECMO or shortly after termination of bypass, asymmetric vasculopathy was found in a greater percentage of our patients compared with a similar large case series. ECMO-associated retinal vasculopathy appeared to disproportionately occur in those patients with CDH who underwent venoarterial bypass. Further study of retinal vascular changes in patients with CDH should be performed to assess long-term effects. PMID- 10532771 TI - Balloon dilatation for treatment of resistant nasolacrimal duct obstruction. AB - PURPOSE: Our purpose was to report our experience with balloon catheter dilatation for resistant nasolacrimal duct obstruction. METHODS: Patients enrolled had symptoms of nasolacrimal duct obstruction and (1) had failed previous probing or (2) were more than 2 years old. Balloon dilatation was performed with a LacriCATH lacrimal catheter (Atrion Medical Products, Birmingham, Ala.). A subset of patients had Silastic silicone rubber (Dow Corning, Midland, Mich.) intubation after balloon dilatation. Success was determined by clinical examination a minimum of 6 weeks later. RESULTS: Twenty one lacrimal systems of 12 patients were treated (age range 4 months to 7 years). Of the patient subset treated with a LacriCATH lacrimal catheter alone, 9 of 18 systems demonstrated complete resolution of symptoms. Three of the 12 patients underwent balloon dilatation intraoperatively after attempts at Silastic silicone rubber intubation were unsuccessful. In two of these patients, who were younger, Silastic silicone rubber tubes passed easily after balloon treatment; however, in an older patient, age 5 years, intubation still could not be accomplished. CONCLUSION: Common clinical strategy for treatment of resistant nasolacrimal obstruction includes repeat probing, intubation of the nasolacrimal system with Silastic silicone rubber tubes, or dacryocystorhinostomy. Balloon catheter dilatation is an alternative or adjunct to consider. Factors that may affect the success of treatment include the age of the patient, the complexity of the nasolacrimal anatomy, and use of adjunctive systemic antibiotics and steroids. PMID- 10532772 TI - Medial rectus fadenoperation for esotropia only at near fixation. AB - PURPOSE: The patient with a large esodeviation at near and a minimal if any deviation at distance who is unable to discontinue bifocal wear or demonstrates a decrease in binocular function presents a difficult therapeutic dilemma to the strabismologist. The medial rectus fadenoperation has been proposed as a method of reducing the accommodative convergence/accommodation (AC/A) ratio and may be a reasonable approach to correct a deviation that is present only at near fixation. We examined our results with this type of operation in these difficult patients. METHODS: A review of consecutive patients who underwent a bilateral medial rectus fadenoperation (no recession) for high AC/A ratio esotropia with a significant esodeviation only at near was performed. Preoperative and postoperative motility and sensory status were evaluated. RESULTS: Sixteen patients meeting the criteria were identified. The preoperative mean distance deviation was 8.8 PD of esodeviation and the mean near deviation was 33 PD of esodeviation. The mean AC/A ratio (by the gradient method) preoperatively was 7.4 PD/D. The average age at surgery was 8 years. Six (38%) had undergone prior strabismus surgery. Average follow-up was 36 months (range 12 to 98 months). Postoperatively the mean distance deviation was 3.2 PD of esodeviation and the mean near deviation was 10.3 PD of esodeviation. The mean decrease in the near deviation was 22.8 PD. The mean postoperative AC/A ratio was 2.9. Ninety-five percent of the patients normalized their AC/A ratios and 70% attained 400 arc-sec of stereopsis postoperatively (vs 44% preoperatively. The majority of patients (86%) were able to maintain satisfactory near ocular alignment without bifocals. CONCLUSIONS: The fadenoperation appears to be useful in this patient population by reducing the distance-near disparity and the AC/A ratio, by decreasing the necessity for bifocal wear, and by improving binocular function at near. PMID- 10532773 TI - Combined bilateral superior rectus muscle recession and inferior oblique muscle weakening for dissociated vertical deviation. AB - PURPOSE: Dissociated vertical deviation (DVD) is a common disorder that is often difficult to treat satisfactorily with extraocular muscle surgery. Weakening both elevators in a single eye is uncommonly performed because of possible severe upgaze deficiency or chin-up head posture postoperatively. METHODS: A retrospective review of medical records was performed that yielded 14 patients who had undergone bilateral superior rectus muscle recessions (mean 8.1 mm, range 5-10 mm) and bilateral inferior oblique muscle recession, myectomy, or anterior transposition in the treatment of DVD. Three additional patients with asymmetric inferior oblique muscle overaction or true hypertropia in primary gaze position were identified who had bilateral superior rectus muscle recessions combined with unilateral inferior oblique muscle weakening. RESULTS: Mild-to-moderate elevation deficiencies were common postoperatively but never exceeded -2 up-gaze limitation (scale 0 to -4) except in the immediate postoperative period and were not associated with persistent chin-up head posturing. Cosmetically objectionable upper eyelid retraction occurred in one patient after re-recession of a superior rectus muscle but before inferior oblique muscle surgery. Only three patients undergoing four vertical muscle surgeries had residual DVD >10 PD in primary gaze position, and none exhibited manifest dissociated strabismus warranting further treatment. CONCLUSION: Bilateral superior rectus muscle recession of up to 10 mm combined with inferior oblique muscle weakening appears to be a safe surgical approach in the management of patients with large angle or recurrent DVD. Our data further suggest that simultaneous four vertical muscle surgery may be preferred in some patients to weakening the superior rectus or inferior oblique muscles alone. PMID- 10532774 TI - Autogenous fascia augmentation of a partially extirpated muscle with a subperiosteal medial orbitotomy approach. AB - INTRODUCTION: Endoscopic sinus surgery can result in serious extraocular muscle dysfunction. The medial rectus muscle is more frequently affected than other extraocular muscles. METHODS: A transconjunctival subperiosteal medial orbitotomy was successful in retrieving a partially extirpated medial rectus muscle after endoscopic sinus surgery. RESULTS: A previous attempt to localize this muscle by conventional surgery with extensive exploration was unsuccessful. A Hummelsheim procedure was also abandoned after a rupture of the nasal aspect of the inferior rectus muscle occurred. CONCLUSION: The approach we describe allowed adequate visualization of the posterior orbital content, as well as adequate space for suture placement. PMID- 10532775 TI - Comitant esodeviation resulting from neurologic insult in children. AB - PURPOSE: Our purpose was to establish whether comitance was a common or uncommon finding in children with esodeviation associated with a neurologic insult. METHODS: A retrospective chart review was performed of children with acquired esodeviation associated with an identifiable neurologic insult. RESULTS: Examinations of 30 children seen over a 2-year period were analyzed. Twenty-two (73%) had brain tumors. Twelve (40%) had comitant esodeviation, and the other 18 (60%) had incomitant measurements. Of the patients with comitant esodeviation, 6 had normal abduction OU (two after recovery from bilateral sixth nerve palsies), and 6 had mild or minimal abduction deficits. Moderate or severe abduction deficits were associated with incomitant measurements. CONCLUSIONS: Comitant esodeviation can be common in children with identifiable neurologic insults. PMID- 10532776 TI - Visual outcome in high hypermetropia. AB - INTRODUCTION: We wished to determine whether final visual acuity is dependent on age at optical correction or presence of esotropia in children with bilateral high hypermetropia. METHODS: We reviewed the charts of all patients at Childrens Hospital Los Angeles Division of Ophthalmology with bilateral hypermetropia of greater than or equal to 5D who were able to provide objective visual acuity outcomes with Snellen letters or linear E. RESULTS: One hundred thirteen patients met entry criteria. The age at first optical correction ranged from 8 months to 141 months (average 45 months). Initial visual acuity (before optical correction) was obtainable in 82 patients. Initial visual acuity ranged from 20/20 to 20/200, with 57% of patients having acuity better than or equal to 20/40. Final visual acuity (after optical correction) ranged from 20/20 to 20/70, with 109 patients (96%) having acuity better than or equal to 20/40 and 104 patients (92%) having acuity better than or equal to 20/30. There was no relationship between final visual acuity and age that spectacles were first worn. Ninety-five patients (84%) had esotropia with or without glasses, and six of these (6%) had final visual acuity less than 20/30. Of the 18 patients with orthotropia, three (16%) had final visual acuity less than 20/30. The prevalence of ametropic amblyopia in patients with esotropia and orthotropia was not significantly different (p = 0.18). CONCLUSION: Visual acuity outcome in children with high hypermetropia is generally good regardless of age at initial optical correction or presence of strabismus. A significantly increased risk for ametropic amblyopia was not found in those patients with orthotropia. PMID- 10532777 TI - Sleep disorders in children with congenital anophthalmia and microphthalmia. AB - PURPOSE: The increased incidence of sleep disorders among blind patients has been documented in the sleep medicine literature. Blind patients lack the normal regulatory control of retinal input over their circadian rhythms, which can lead to abnormalities in their sleep-wake cycles. Our study was conducted to determine the incidence of sleep disorders in children with anophthalmia or microphthalmia and to offer therapeutic alternatives. METHODS: A 13-question survey was distributed to families of children with anophthalmia, microphthalmia, or both identified through the Anophthalmia/Microphthalmia Registry in Philadelphia, Pennsylvania. The survey included questions regarding the children's medical and ocular histories and any sleep disorders they may have experienced. Questions regarding daily schedules, family history, and social history were also included. RESULTS: Surveys were returned from 13 children with bilateral anophthalmia or microphthalmia. Ten of 13 (77%) anophthalmic/microphthalmic children were reported to have frequent early-morning waking and extensive daytime sleeping. Specific medical and social problems did not appear to be associated with the development of these sleep disorders. Strict daily schedules were often helpful in entraining the children's sleep-wake cycles. CONCLUSION: Without the contribution of retinal input to help regulate circadian rhythms, most children with bilateral anophthalmia or microphthalmia will experience sleep disorders. These children may benefit from the introduction of strict daily schedules, medical therapy (melatonin), or both. An attempt should also be made to preserve any existing light perception. PMID- 10532778 TI - Implant brachytherapy: a novel treatment for recurrent orbital rhabdomyosarcoma. AB - BACKGROUND: Orbital rhabdomyosarcoma is the most common primary malignancy of the orbit in childhood. Tumor resection and exenteration were the preferred treatment modalities for rhabdomyosarcoma. In the past 20 years, however, combined local radiation and systemic chemotherapy have shown excellent survival results. Tumor recurrence after any of the aforementioned therapies is almost always fatal. We have developed a novel treatment for recurrent disease that has resulted in long term survival for three patients. METHODS: Three patients with recurrent orbital rhabdomyosarcoma were previously treated with primary radiation and chemotherapy. At the time of recurrence, exenteration and localized brachytherapy were performed. An individually molded poly(methylmethacrylate) (Lucite; E. I. du Pont de Nemours & Co., Wilmington, Del.) device loaded with radioactive iodine seeds delivered localized high-dose radiation, 6000 cGy over 6 days, to the orbit without irradiating the brain. RESULTS: All patients are alive and free of disease with follow-up ranging from 4 years and 4 months to 8 years and 4 months. CONCLUSION: A novel technique of delivering localized radiation to the orbit of three children with recurrent orbital rhabdomyosarcoma appears curative. PMID- 10532779 TI - Positron-emission tomographic study of human amblyopia with use of defined visual stimuli. AB - PURPOSE: The purpose of this study was to use positron emission tomography (PET) to evaluate effects of amblyopia on cerebral blood flow and glucose metabolism in humans viewing defined visual stimuli and to correlate these effects with specific behavioral and electrophysiologic measures of visual function. METHODS: One subject with normal vision and five patients with amblyopia were prospectively studied. During monocular viewing of a checkerboard reversal stimulus by each subject, we performed PET imaging of relative cerebral glucose metabolism with use of [18F]fluorodeoxyglucose, PET imaging of relative cerebral blood flow with use of H2(15)O, and visual evoked potentials. Control studies were also performed with use of binocular occlusion and during presentation of stationary and horizontally drifting checkerboards. These data were correlated with letter acuities and contrast sensitivity functions for each eye. RESULTS: Although spatial resolution was superior for glucose metabolic imaging, PET readily demonstrated activation of calcarine cortex with use of both metabolic and blood flow tracers. Even in patients with mild amblyopia, functional activation of calcarine cortex was reduced in amblyopic eyes compared with sound eyes to a degree more closely correlated with visual acuity than were visual evoked potential amplitudes to the same stimulus. When responses to drifting versus stationary stimuli were compared, a putative motion processing center was identified in the right temporoparietal region. Activity in this motion center was relatively preserved during viewing of drifting stimuli by the affected eye of an anisometropic amblyopic subject, but was attenuated during viewing of the same stimulus by the affected eye of a strabismic amblyopic subject. CONCLUSIONS: PET imaging of blood flow and metabolism can quantitatively evaluate functional deficits resulting from amblyopia in striate as well as extrastriate visual areas. Calcarine cortical function correlates closely with severity of amblyopia, but function in a putative motion processing area may vary according to the type of amblyopia present. PMID- 10532780 TI - Electroretinography in incontinentia pigmenti. PMID- 10532781 TI - Intraoperative dehiscence of a rectus muscle: report of two cases. PMID- 10532782 TI - A new needle holder for use in hang-back strabismus surgery. PMID- 10532783 TI - Congenital iris ectropion associated with ocular albinism, foveal hypoplasia, and keratoconjunctivitis sicca. PMID- 10532784 TI - Acute serous detachment with argon laser photocoagulation in retinopathy of prematurity. PMID- 10532785 TI - Ocular injury caused by war games. PMID- 10532786 TI - Improving the social responsiveness of medical schools. Proceedings of the 1998 Educational Commission for Foreign Medical Graduates/World Health Organization Invitational Conference. PMID- 10532787 TI - Improving the social responsiveness of medical schools: summary of the conference. PMID- 10532788 TI - Prometheus through the looking glass: reflections on the hepatic immune system. PMID- 10532789 TI - Cruel antibody fictions! Cellular antigen enumeration by 'saturation' binding. PMID- 10532790 TI - Biologic processing of fossil fuels. Rsume of the Bioconversion Session of ICCS'97. PMID- 10532791 TI - [Otomastoiditis by Aspergillus fumigatus in a patient with AIDS]. PMID- 10532792 TI - Food Micro '99. Ecology and Physiology of Food Related Micro-Organisms. Proceedings of the 17th International Conference of the International Committee on Food Microbiology and Hygiene (ICFMH). Veldhoven, The Netherlands, September 13-17, 1999. PMID- 10532793 TI - Current awareness in NMR in biomedicine. PMID- 10532794 TI - 2nd Joint Meeting of the Leeds Castle Polyposis Group and International Collaborative Group for Hereditary Non-Polyposis Colorectal Cancer. Melbourne and Lorne, Australia, 1-6 March 1999. Abstracts. PMID- 10532795 TI - [Care of hypertension among the aged]. PMID- 10532796 TI - Development of fat emulsions. 1981. PMID- 10532797 TI - Response to Dr. Brent's commentary on "Dr. Brent and scientific debate". PMID- 10532798 TI - Proceedings of a conference on stress-inducible gene expression. Danvers, Massachusetts, USA. November 12-15, 1998. PMID- 10532799 TI - 17th International Symposium of the European Cancer Prevention Organization in Collaboration with the Japanese Society for Cancer Prevention and IPATIMUP. Gene environmental interactions in the digestive tract: their role in human cancer. Porto, Portugal, 13-15 May 1999. Abstracts. PMID- 10532800 TI - Proceedings of a joint symposium of the Phycological Society of America, International Society for Evolutionary Protistology and the Society of Protozoologists. Evolution of Mitochondria and Chloroplasts. Flagstaff, Arizona, USA. 1-8 August 1998. PMID- 10532802 TI - Proceedings of a symposium on uncoupling proteins and obesity. Quebec City, Canada, November 20-21, 1998. PMID- 10532801 TI - Evidence of actin in the cytoskeleton of microsporidia. AB - Using transmission electron microscopy, immuno-electron microscopy, and biochemical techniques such as 2-D electrophoresis and immunoblotting, actin was found in all biological stages of the microsporidia Encephalitozoon hellem and Encephalitozoon cuniculi. PMID- 10532803 TI - Role of G proteins and nitric oxide in seizures induced by different chemoconvulsants in mice. PMID- 10532804 TI - Proceedings of the International Ion Chromatography Symposium 1998. Osaka, Japan, 28 September-1 October 1998. PMID- 10532805 TI - Molecular cloning, tissue-specific expression, and chromosomal localization of a novel nerve growth factor-regulated G-protein- coupled receptor, nrg-1. AB - A novel and differentially expressed gene, named nrg-1, was identified by EST expression profiling and subsequently isolated as a 2.2-kb full-length clone from a rat PC12 cell cDNA library. Sequence analysis reveals that nrg-1 encodes a putative seven transmembrane spanning domain protein with structural features characteristic of receptors belonging to the G-protein-coupled receptor gene superfamily. The 400-amino-acid protein encoded by nrg-1 exhibits a high degree of sequence identity (40-44%) to the Edg receptor family; members include Edg-1, Edg-2, Edg-3, Edg-4, and H218. Both Northern analysis andEST expression profiling revealed that whole-tissue distribution of nrg-1 mRNA is restricted, found almost exclusively in brain. Transcripts of nrg-1 could be ubiquitously detected in different regions, with very prominent expression in lower brain regions such as the midbrain, pons,medulla, and spinal cord. In PC12 cells, nerve growth factor induces neuronal differentiation and repressed expression of nrg-1. Two other agents that differentiate PC12 cells, fibroblast growth factor and dibdutyryl cAMP, down-regulated nrg-1 mRNA levels. Epidermal growth factor, and agent that does not induce differentiation, did not repress nrg-1 mRNA levels. In a PC12 cell mutant that is deficient in protein kinase A activity (AB.11), all three differentiating agents were unable to down-regulate nrg-1 mRNA. Hence, protein kinase A appears to be an obligatory cellular component in nrg-1 mRNA regulation. Chromosomal mapping employing a rat somatic cell readiation hybrid panel demonstrated that nrg-1 is linked to marker D8Rat54 and tightly associated with H218 on chromosome 8. PMID- 10532806 TI - Distinctive patterns of PDGF-A, FGF-2, IGF-I, and TGF-beta1 gene expression during remyelination of experimentally-induced spinal cord demyelination. AB - Although remyelination is a well-recognized regenerative process following both experimental and naturally occurring CNS demyelination, remarkably little is known about the molecules involved in its orchestration. In this study we have examined the mRNA expression of seven growth factors that influence oligodendrocyte lineage cells, during the remyelination of lysolecithin-induced demyelination in the rat spinal cord. These lesions involve rapid demyelination of axons, which undergo extensive remyelination between 10 and 28 days. The distribution and levels of expression of PDGF-A, IGF-I, CNTF, FGF-2, TGF-beta1, GGF-2, and NT-3 mRNAs were examined at 2, 5, 7, 10, 14, 21, and 28 days post lesion induction, both within the lesion and within dorsal root ganglia whose axons transverse the lesion, by quantitative in situ hybridization using 35S labeled oligonucleotide probes. large increases in IGF-I and TGF-beta1 mRNAs were evident within the spinal cord by 5 days. These levels peaked at 10 days at a time when new myelin sheaths appear and had declined by 28 days. Increases in FGF 2 and PDGF-A mRNAs were less intense and less widely distributed than those of IGF-I and TGF-1, but remained elevated for a longer duration. There were no changes in expression of CNTF, NT-3, or GGF-2 mRNAs within the lesioned cords; neither were ther changes in levels of expression of any growth factor mRNAs in the dorsal root ganglia. This work therefore indicates that some but not all members of the family of growth factors that affect the oligodendrocyte lineage are expressed during remyelination of demyelinated spinal cord axons and provides the data on which future studies on the specific roles of these factors in orchestrating this important regenerative process will be based. PMID- 10532807 TI - B-50/GAP-43 potentiates cytoskeletal reorganization in raft domains. AB - B-50 (GAP-43) is a neural, membrane-associated protein that has been implicated in neurite outgrowth and guidance. Following stable transfection of Rat1 fibroblasts with B-50 cDNA we observed a dispersed distribution of B-50 immunoreactivity in flattened resting cells. In contrast, motile cells exhibited high concentrations of B-50 at the leading edge of ruffling membranes, coinciding with actin polymerization. Time-lapse studies on Rat1 fibroblasts transiently transfected with B-50/EGFP revealed that large vesicles originated from the ruffling membranes. These large vesicles (pinocytes) were found positive for Thy 1, a GPI-anchored protein, but negative for rab-5, an early endosome marker. In primary hippocampal neurons B-50 also colocalized completely with the raft marker Thy-1. Antibody-mediated cross-linking of Thy-1 in hippocampal neurons resulted in a redistribution of the intracellular protein B-50 to Thy-1-immunopositive membrane patches, whereas syntaxin was mainly excluded from the patches, showing that B-50 is associated with rafts. Academic Press. PMID- 10532808 TI - Mice lacking alpha-calcitonin gene-related peptide exhibit normal cardiovascular regulation and neuromuscular development. AB - alpha-Calcitonin gene-related peptide (alphaCGRP) is a pleiotropic peptide neuromodulator that is widely expressed throughout the Central and peripheral nervous systems. CGRP has been implicated in a variety of physiological processes including peripheral vasodilation, cardiac acceleration nicotinic acetylcholine receptor (AChR) synthesis and function, testicular descent, nociception, carbohydrate metabolism, gastrointestinal motility, neurogenic inflammation, and gastric acid secretion. To provide a better understanding of the physiological role(s) mediated by this peptide neurotransmitter, we have generated alphaCGRP null mice by targeted modification in embryonic stem cells. Mice lacking alpha CGRP expression demonstrate no obvious phenotypic differences from their wild type littermates. Detailed analysis of systemic cardiovascular function revealed no differences between control and mutant mice regarding heart rate and blood pressure under basal or exercise-induced conditions and subsequent to pharmacological manipulation. Characterization of neuromuscular junction in morphology including nicotinic receptor localization, terminal sprouting in response to denervation, developmental regulation of AChR subunit expression, and synapse elimination also revealed no differences in alphaCGRP-deficient animals. These results suggest that alphaCGRP is not required for the systemic regulation of cardiovascular hemodynamics or development of the neuromuscular junction. PMID- 10532809 TI - Renal immunology and pathology. PMID- 10532810 TI - Mental lexicon. Introduction. PMID- 10532811 TI - The official issue of the Japanese Society of Sleep Research. Proceedings of the 23rd annual meeting of the JSSR. Akita, Japan, 4-5 June 1998. PMID- 10532812 TI - Chemical biology: the promise, and confusion, of adolescence. AB - It takes time for any new scientific discipline to gain momentum, and chemical biology is no exception. But with the formation of new training programs and interdisciplinary departments, the changes are coming. PMID- 10532813 TI - Cell biology goes commercial. Cytokinetics, Inc. PMID- 10532814 TI - Response: finding the disorder in gender identity disorder. PMID- 10532815 TI - High fibre diet. PMID- 10532817 TI - True arbiters of prosperity. PMID- 10532816 TI - Biliary sphincter balloon dilation; who, when and how? AB - Biliary sphincter balloon dilation for biliary stone removal was introduced in 1983. In the early 1990s, several groups studied this technique further. The success rate of stone removal is comparable with that of endoscopic sphincterotomy in patients with fewer than three stones that are less then 1 cm in diameter. Fewer complications after balloon dilation than after endoscopic sphincterotomy have been noted in most studies. One study, however, showed a higher incidence of pancreatitis and, in particular, severe pancreatitis. Therefore, there is still some reluctance among endoscopists to promote balloon dilation as a routine first choice treatment. The technique, however, is accepted as the treatment of choice in patients with a bleeding tendency and those in whom the local anatomy is associated with an increased risk of complications with endoscopic sphincterotomy, such as patients with periampullary diverticula or Billroth II gastrectomy. PMID- 10532818 TI - Tuberculosis in the 1990's: From bedside to bench and back. PMID- 10532819 TI - Bernstein's heritage: The active search for information: from reflexes to the model of the future (1966). PMID- 10532820 TI - From reflexes to the model of the future. 1966. PMID- 10532821 TI - Pictures in cell biology. Pak1 kinase activity affects the character of cell morphology and movement. PMID- 10532822 TI - Setting the clock in Madrid. PMID- 10532823 TI - Documentation for the sake of documentation? PMID- 10532824 TI - 1st International Conference on Bartonella as Emerging Pathogens. Tubingen, Germany, March 5-7, 1999. Abstracts. PMID- 10532825 TI - Alphonse Jaminet on caisson disease: 1871--a commentary. PMID- 10532826 TI - Current literature in diabetes. PMID- 10532827 TI - Nicholas T. Zervas. PMID- 10532828 TI - Proceedings of the 47th Annual Meeting of the Congress of Neurological Surgeons. New Orleans, Louisiana, USA. 1997. PMID- 10532829 TI - The function of PS integrins in Drosophila wing morphogenesis. AB - Integrins are found on many cell types during the development of most organisms. In Drosophila their functions can be analysed genetically. An analysis of lethal mutations in a PS integrin gene showed that the integrins were required for muscle attachment and for certain cell sheet migrations during embryogenesis. In this paper we use viable mutations in integrin component genes to look at integrin function in the later stages of development of one adult structure, the wing. We show that two known viable mutations, one which has its primary effect on the fly's escape response, the other on wing morphogenesis, are mutations in the beta and PS2alpha subunits, respectively, of the PS integrins. The mutation non-jumper (mys(mj42)) in the beta subunit leads to wasting of the thoracic jump muscles. Flies in which the dosage of this allele is reduced (and no wildtype copy is present) show defects also in wing morphogenesis. The two surfaces of the wing fail to connect properly, resulting in 'blistering' of the wing and the formation of extra crossveins. The mutation in the gene for the PS2alpha integrin subunit, inflated, also leads to a failure in wing surface apposition and consequent wing blistering. When the two mutations are combined, the mutant phenotype is greatly enhanced. Thus, one of the roles of the PS integrins in late Drosophila development is to ensure the correct apposition and patterning of the wing epithelia. PMID- 10532830 TI - Special issue in memoriam John S. Fawcett. PMID- 10532831 TI - Intracortical hemangioma of the mandible. Case report. AB - We describe the first reported case of an intracortical hemangioma of the mandible in a 13-year-old Japanese girl. Panoramic radiography and CT demonstrated a small osteolytic lesion which had expanded and thinned the cortex at the inferior border of the left mandible. The lesion enhanced on post-contrast T1-weighted MRI. The diagnosis was confirmed by histopathology following block excision. Intracortical hemangioma should be considered in the differential diagnosis of radiolucent lesions orf the mandible PMID- 10532832 TI - European General Practice Research Workshop. Norway, 8-10 May 1998. Abstracts. PMID- 10532833 TI - Flat adenomas exist in asymptomatic people: important implications for colorectal cancer screening programs. PMID- 10532834 TI - Mucin-producing pancreatic tumors: comparison of MR cholangiopancreatography with endoscopic retrograde cholangiopancreatography. PMID- 10532835 TI - Twenty-six cases of mucinous ductal ectasia of the pancreas. PMID- 10532836 TI - News from the IAG. International Association of Gerontology. PMID- 10532837 TI - 27th Annual Conference on Yeasts. Smolenice, Slovak Republic, May 13-15, 1998. Abstracts. PMID- 10532838 TI - Proceedings of the 10th Symposium on Signals and Signal Processing in the Immune System. Balatonoszod, Hungary, 13-17 September 1998. PMID- 10532839 TI - [Gaceta Sanitaria: a renovation for the future]. PMID- 10532840 TI - [Gaceta Sanitaria: a messenger in the house of public health]. PMID- 10532841 TI - [Guidelines for prescription drugs; special interests are counterproductive]. PMID- 10532842 TI - [Change of course in health policy; from responsible citizen to dependent patient?]. PMID- 10532843 TI - [Speech given by Andrea Fischer, Minister of Health on the 102. German Physicians' Day in Cottbus 1999]. PMID- 10532844 TI - [The significance of leptin for the pathogenesis of diabetes mellitus type 2. Direct effects on endocrine pancreas]. PMID- 10532845 TI - [Multidisciplinary structure requires interprofessional cooperation. Report from seminar-congress 'Interdisciplinary intensive care' Garmisch-Partenkirchen, March 3-12, 1999]. PMID- 10532846 TI - [Atherosclerosis-state of the art. International Expert Meeting on Vascular Calcifications, Wittenberg, April 9-10, 1999]. PMID- 10532847 TI - XII International Conference on Event Related Potentials of the Brain, EPIC XII. Boston, Massachusetts, USA. 19-23 July 1998. Abstracts. PMID- 10532848 TI - 1998 Annual Meeting on Muscle and Cell Motility Physiology. Tokyo, Japan, 27-28 November 1998. Abstracts. PMID- 10532849 TI - Current issues in dermatologic office-based surgery. Joint American Academy of Dermatology/American Society of Dermatologic Surgery Liaison Committee. PMID- 10532850 TI - The 1998 Kodak Awards to the Institute of Medical Illustrators. PMID- 10532851 TI - Development of a scale for the comparison of metals in enzyme action. PMID- 10532852 TI - Chance and design. PMID- 10532853 TI - Harassment of lesbians as medical students and physicians. PMID- 10532854 TI - Perinatal HIV prevention. PMID- 10532855 TI - High expression of erythropoietin receptor in human chronic progressive glomerulonephritis. PMID- 10532856 TI - Research in medical education. PMID- 10532857 TI - The core content of the undergraduate curriculum in Manchester. PMID- 10532858 TI - Training medical students about HIV/AIDS? PMID- 10532859 TI - [Transcutaneous creation of a left ventricular aneurysm model in mongrel dogs and experimental study of cardiomyoplasty]. PMID- 10532860 TI - Reconstitution of active dimeric ribulose bisphosphate carboxylase from an unfoleded state depends on two chaperonin proteins and Mg-ATP. AB - In vitro reconstitution of active ribulose bisphosphate carboxylase (Rubisco) from unfolded polypeptides is facilitated by the molecular chaperones: chaperonin 60 from Escherichia coli (groEL), yeast mitochondria (hsp60) or chloroplasts (Rubisco sub-unit-binding protein), together with chaperonin-10 from E. coli (groES), and Mg-ATP. Because chaperonins are ubiquitous, a conserved Mg-ATP dependent mechanism exists that uses the chaperonins to facilitate the folding of some other proteins. PMID- 10532862 TI - [Archil Karpezovich Dondua (on his 70th birthday)]. PMID- 10532863 TI - The Therapeutic Orphan--30 Years Later. Proceedings of a joint conference of the Pediatric Pharmacology Research Unit Network, the European Society of Developmental Pharmacology, and the National Institute of Child Health and Human Development. Washington DC, USA, May 2, 1997. PMID- 10532864 TI - IMpact-RSV Study Group report. PMID- 10532865 TI - IMpact-RSV Study Group report. PMID- 10532866 TI - IMpact-RSV Study Group report. PMID- 10532867 TI - Tanner staging and pornography. PMID- 10532868 TI - Tanner staging and pornography. PMID- 10532869 TI - Tanner staging and pornography. PMID- 10532870 TI - Observer bias in acellular pertussis vaccine trials. PMID- 10532871 TI - Foster care problem. PMID- 10532872 TI - Teaching awards and departmental longevity: is award-winning teaching the "Kiss of Death" in an academic department of medicine? PMID- 10532873 TI - Index of suspicion. Case 2. Inborn errors of bile salt metabolism. PMID- 10532874 TI - Antifungal biflavones from Cupressocyparis leylandii. AB - From the leaves of Cupressocyparis leylandii (Cupressaceae) cupressuflavone, 4-O methylcupressuflavone, amentoflavone, 7-O-methylamentoflavone, 4-O methylamentoflavone and hinokiflavone were isolated. 1H- and 13C-NMR data for 4-O methylcupressuflavone are given for the first time. The biflavones from cultivar varieties of C. leylandii (Naylor's Blue, Castlewellan Gold) were chromatographicaly (HPLC) compared. The antifungal activity of cupressuflavone and 4-O-methylcupressuflavone against Alternaria alternata, Cladosporium oxysporum, Fusarium culmorum and F. avenaceum was assayed. PMID- 10532875 TI - Protective effects of the aerial parts of Salvia officinalis, Melissa Officinalis and Lavandula angustifolia and their constituents against enzyme-dependent and enzyme-independent lipid peroxidation. AB - The antioxidant effects of aqueous methanolic extracts from three medicinal Lamiaceae species were investigated in enzyme-dependent and enzyme-independent lipid peroxidation systems. All these extracts caused a considerable concentration-dependent inhibition of lipid peroxidation. Phenolic components present in the plant extracts were evaluated for antioxidant activity and were found effective in both tests. Their concentrations in each extract were determined by TLC-densitometry. PMID- 10532877 TI - ICH Harmonised Tripartite Guideline. Statistical principles for clinical trials. International Conference on Harmonisation E9 Expert Working Group. PMID- 10532876 TI - Bioactive constituents from Pteris multifida. PMID- 10532878 TI - [Proceedings of the 51st meeting of the American Academy of Neurology. Toronto, Canada, 17-24 April 1999]. PMID- 10532879 TI - Neanderthals were cannibals, bone show. PMID- 10532881 TI - Home again? PMID- 10532880 TI - Senate tops house panel in raising NIH's budget. PMID- 10532882 TI - Evolution. Handsome finches win a boost for their offspring. PMID- 10532883 TI - Rice genome. U.S. adds $12 million to global sequencing push. PMID- 10532884 TI - Neptune may crush methane into diamonds. PMID- 10532885 TI - India creates novel brain research center. PMID- 10532886 TI - Gene defect linked to Rett syndrome. PMID- 10532887 TI - Ethical loophole closing up for stem cell researchers. PMID- 10532888 TI - Permafrost comes alive for Siberian researchers. PMID- 10532889 TI - Intracytoplasmic sperm injection. PMID- 10532890 TI - Switching on the infant brain. PMID- 10532891 TI - Chemical diversity in RNA cleavage. PMID- 10532892 TI - Expanding the habitable zone. PMID- 10532893 TI - [Vascular cells and estrogens. Proceedings of the 3rd annual meeting in experimental and clinical pharmacology. 26-28 November 1998]. PMID- 10532894 TI - Biological monitoring in occupational and environmental toxicology. Proceedings of the 4th International Symposium on Biological Monitoring. Seoul, Korea, 23-25 September 1998. PMID- 10532895 TI - Repeat meeting's repeat performance. The Second International Conference on Unstable Microsatellites and Human Disease, Chapel Hill, North Carolina, USA, 17 20 April 1999. PMID- 10532896 TI - Genomics and psychiatry. Genes and the Neurobiology of Susceptibility: the Society of Biological Psychiatry 54th Annual Scientific Convention, Mayflower Renaissance Hotel, Washington, DC, USA, 13-15 May 1999. PMID- 10532897 TI - [Continuing medical education. Proceedings of the XXVIII Medical Congress of Maghrebin. Alger, 21-23 June 1999]. PMID- 10532898 TI - Proceedings of a conference on equine cyathostomes. Athens, Georgia, USA. 7-8 November 1998. PMID- 10532899 TI - Cirrhosis mortality and per capita consumption of distilled spirits, United States, 1949-1994: trend analysis. AB - OBJECTIVE: To describe, evaluate, and suggest interpretations for an observed aggregate-level relation between trends in mortality from cirrhosis and per capita consumption of distilled spirits in the United States. DESIGN: Trend analysis using data on US cirrhosis mortality and per capita alcohol consumption. RESULTS: There is a consistent long-term trend relation between mortality from cirrhosis and per capita consumption of distilled spirits in the United States from 1949 to 1994. Two instances of comparatively sharp drops in the consumption of spirits in the 1940s generated mixed results in predicting changes in cirrhosis mortality. CONCLUSIONS: An aggregate-level relation between trends in long-term cirrhosis mortality and the consumption of spirits falls considerably short of establishing a direct causal link between the two for individuals. Moreover, two sharp drops in the consumption of spirits generated only mixed results with respect to the short-term trend in cirrhosis. Nevertheless, the observed relation between the consumption of spirits and cirrhosis mortality merits further investigation. PMID- 10532901 TI - European Concerted Action (COST 917). Biogenically active amines in food. Polyamines, the gut and cancer. Bad Nauheim, Germany, 30 April-2 May 1999. Abstracts. PMID- 10532900 TI - Proceedings of the Enterochromaffin-like Symposium. New Haven, Connecticut, USA. February 1-2, 1997. PMID- 10532902 TI - Current awareness on yeast. PMID- 10532903 TI - Current awareness on yeast. PMID- 10532904 TI - Cholesterol in a historic perspective. Report of the professor C.D. de Langen symposium. The Netherlands, December 4, 1998. PMID- 10532905 TI - Current progress in the understanding of secondary brain damage from trauma and ischemia. Proceedings of the 6th International Symposium: Mechanisms of Secondary Brain Damage--Novel Developments. Mauls/Sterzing, Italy, February 1998. PMID- 10532906 TI - Proceedings of the 3rd International Congress on Vegetarian Nutrition. Loma Linda, California, USA. March 24-26, 1997. PMID- 10532907 TI - Mosaicism in human skin. Proceedings of a symposium in honor of Rudolf Happle. Marburg, Germany, May 1998. PMID- 10532908 TI - Neurofibromatosis 1. Introduction. PMID- 10532909 TI - [Delegation of responsibility from physicians to physician assistants]. PMID- 10532910 TI - New concepts in antimicrobial therapy for emergency department infections. PMID- 10532911 TI - Clinical pathways for general surgeons: abdominal revascularization. PMID- 10532912 TI - Albumin: saint or sinner? PMID- 10532913 TI - Attention deficit hyperactivity disorder. PMID- 10532914 TI - Botryoid neutrophils in unexpected heat stroke. PMID- 10532915 TI - CMV coinfection and disease progression in vertically acquired HIV infection. PMID- 10532916 TI - Doctors as expert witnesses. PMID- 10532917 TI - EEG and epilepsy. PMID- 10532918 TI - Infant feeding and atopic disease. PMID- 10532919 TI - Is prolonged rotavirus infection a common cause of protracted diarrhoea? PMID- 10532920 TI - Pretrial liaison between doctors in alleged child abuse. PMID- 10532921 TI - Rectal biopsy in the investigation of constipation. PMID- 10532922 TI - Rectal biopsy in the investigation of constipation. PMID- 10532923 TI - Recurrent apparent life threatening events and intentional suffocation. PMID- 10532924 TI - The way ahead with meningococcal disease. PMID- 10532925 TI - Transitional care of young disabled people. PMID- 10532926 TI - Allicin: a possible answer to antibiotic resistant campylobacter diarrhoeal infection? PMID- 10532927 TI - ACTH treatment for gelastic seizures. PMID- 10532928 TI - Less diarrhoea but no change in growth: 15 years' data from three Gambian villages. PMID- 10532929 TI - BCG and tuberculosis. PMID- 10532930 TI - Need to consider other causes of poor growth in Gambian children. PMID- 10532931 TI - Varicella: to vaccinate or not vaccinate? PMID- 10532932 TI - Nurse-led asthma education and childhood asthma readmission rates. PMID- 10532933 TI - Kearns Sayre syndrome initially presenting as hypomelanosis of Ito. PMID- 10532934 TI - Controlled trials from history. PMID- 10532936 TI - The role of the Glasgow meningococcal septicaemia prognostic score in the emergency management of meningococcal disease. PMID- 10532935 TI - The vitamin K debacle and infants with cholestatic liver disease. PMID- 10532937 TI - Prolonged QTc interval as an important factor in sudden infant death syndrome. PMID- 10532938 TI - Umbilical cord blood transplantation. PMID- 10532939 TI - No strings attached: preventing deaths from children's clothing. PMID- 10532940 TI - Thyroid dysfunction in Down's syndrome: relation to age and thyroid autoimmunity. PMID- 10532941 TI - Where should paediatric surgery be performed? PMID- 10532942 TI - Celebration of the 80th birthday of C. Walton Lillehei, PhD, MD. Minneapolis, Minnesota, USA. October 23-25, 1998. PMID- 10532943 TI - Thematic MiniReviews : A preview PMID- 10532944 TI - Ca(2+) release mechanisms, Ca(2+) sparks, and local control of excitation contraction coupling in normal heart muscle. PMID- 10532945 TI - The myocardial Na(+)-H(+) exchange: structure, regulation, and its role in heart disease. AB - The Na(+)-H(+) exchange (NHE) is a major mechanism by which the heart adapts to intracellular acidosis during ischemia and recovers from the acidosis after reperfusion. There are at least 6 NHE isoforms thus far identified with the NHE1 subtype representing the major one found in the mammalian myocardium. This 110 kDa glycoprotein extrudes protons concomitantly with Na(+) influx in a 1:1 stoichiometric relationship rendering the process electroneutral, and its activity is regulated by numerous factors, including phosphorylation-dependent processes. There is convincing evidence that NHE mediates tissue injury during ischemia and reperfusion, which probably reflects the fact that under conditions of tissue stress, including ischemia, Na(+)-K(+) ATPase is inhibited, thereby limiting Na(+) extrusion, resulting in an elevation in [Na(+)](i). The latter effect, in turn, will increase [Ca(2+)](i) via Na(+)-Ca(2+) exchange. In addition, NHE1 mRNA expression is elevated in response to injury, which may further contribute to the deleterious consequence of pathological insult. Extensive studies using NHE inhibitors have consistently shown protective effects against ischemic and reperfusion injury in a large variety of experimental models and has led to clinical evaluation of NHE inhibition in patients with coronary artery disease. Emerging evidence also implicates NHE1 in other cardiac disease states, and the exchanger may be particularly critical to postinfarction remodeling responses resulting in development of hypertrophy and heart failure. PMID- 10532946 TI - Transcriptional activation of the zinc finger transcription factor BTEB2 gene by Egr-1 through mitogen-activated protein kinase pathways in vascular smooth muscle cells. AB - We have recently demonstrated that a developmentally regulated zinc finger protein, basic transcription regulatory element binding protein 2 (BTEB2), is induced in neointimal smooth muscle in response to vascular injury. In this study, we investigated the molecular mechanisms regulating BTEB2 expression in vascular smooth muscle cells (SMCs) in vitro. BTEB2 mRNA expression is rapidly and persistently induced in SMCs by phorbol 12-myristate 13-acetate (PMA) and basic fibroblast growth factor. We have isolated and characterized the promoter region of the human BTEB2 gene to determine the regulatory network controlling expression of this gene in vascular SMCs. Functional studies on the BTEB2 promoter coupled to a luciferase reporter gene demonstrated activation of the promoter by PMA and basic fibroblast growth factor. Both characterization of DNA protein complexes in vitro and site-specific mutation analysis of the BTEB2 promoter have defined a 9-bp sequence, 5'-CGCCCGCGC-3', located at -25, as the Egr-1 binding site mediating an induction of the BTEB2 promoter activity by PMA. In addition, we show that this site mediates inducible expression through the mitogen-activated protein kinase pathways. These results indicate that BTEB2 is a target of the early-response gene Egr-1, and mitogen-activated protein kinase pathways directly or indirectly activate BTEB2 expression. Given a rapid induction of Egr-1 on stimulation with growth factors or injury, these findings may represent at least one of the molecular mechanisms underlying phenotypic modulation of smooth muscles after vascular injury. PMID- 10532947 TI - Identification and characterization of a new growth hormone-releasing peptide receptor in the heart. AB - Hexarelin, a synthetic hexapeptide of the growth hormone-releasing peptide (GHRP) family with strong growth hormone (GH)-releasing activity, features protecting activity against postischemic ventricular dysfunction in hearts from GH-deficient and senescent rats. To document whether hexarelin action is mediated through specific cardiac receptors, perfusion of Langendorff rat hearts with hexarelin and binding studies were carried out. In the Langendorff rat heart system, hexarelin induced a dose-dependent increase in coronary perfusion pressure. Nifedipine, chelerythrine, and bisindolylmaleimide partially inhibited the vasoconstriction induced by hexarelin, suggesting that this effect was mediated at least in part by L-type Ca(2+) channels and protein kinase C. In contrast, diclofenac and 1-(7-carboxyheptyl)imidazole were without effect, suggesting that prostaglandins and thromboxanes were not involved in the coronary vasoconstriction induced by hexarelin. To characterize the hexarelin binding sites in the rat heart, [(125)I]Tyr-Bpa-Ala-hexarelin was used as photoactivatable radioligand in saturation and competitive binding studies. We specifically labeled a hexarelin receptor with an M(r) of 84 000 in rat cardiac membranes. Saturation binding curves revealed a single class of binding sites with a K(d) of 14.5 nmol/L and a density of 91 fmol/mg of protein. Competition binding studies gave an IC(50) of 2.9 micromol/L for hexarelin; MK-0677 and EP51389, both potent GH secretagogues, did not displace the binding of the photoactivatable derivative from rat cardiac membranes. Interestingly, both compounds were devoid of any vasoconstrictive activity. These results suggest the existence of a new class of hexarelin receptor in the heart, whose role in the regulation of the coronary vascular tone is yet to be determined. PMID- 10532948 TI - Ionic mechanisms responsible for the electrocardiographic phenotype of the Brugada syndrome are temperature dependent. AB - The Brugada syndrome is a major cause of sudden death, particularly among young men of Southeast Asian and Japanese origin. The syndrome is characterized electrocardiographically by an ST-segment elevation in V1 through V3 and a rapid polymorphic ventricular tachycardia that can degenerate into ventricular fibrillation. Our group recently linked the disease to mutations in SCN5A, the gene encoding for the alpha subunit of the cardiac sodium channel. When heterologously expressed in frog oocytes, electrophysiological data recorded from the Thr1620Met missense mutant failed to adequately explain the electrocardiographic phenotype. Therefore, we sought to further characterize the electrophysiology of this mutant. We hypothesized that at more physiological temperatures, the missense mutation may change the gating of the sodium channel such that the net outward current is dramatically augmented during the early phases of the right ventricular action potential. In the present study, we test this hypothesis by expressing Thr1620Met in a mammalian cell line, using the patch-clamp technique to study the currents at 32 degrees C. Our results indicate that Thr1620Met current decay kinetics are faster when compared with the wild type at 32 degrees C. Recovery from inactivation was slower for Thr1620Met at 32 degrees C, and steady-state activation was significantly shifted. Our findings explain the features of the ECG of Brugada patients, illustrate for the first time a cardiac sodium channel mutation of which the arrhythmogenicity is revealed only at temperatures approaching the physiological range, and suggest that some patients may be more at risk during febrile states. PMID- 10532949 TI - Regulation of the transient outward K(+) current by Ca(2+)/calmodulin-dependent protein kinases II in human atrial myocytes. AB - Ca(2+)/calmodulin-dependent protein kinases II (CaMKII) have important functions in regulating cardiac excitability and contractility. In the present study, we examined whether CaMKII regulated the transient outward K(+) current (I(to)) in whole-cell patch-clamped human atrial myocytes. We found that a specific CaMKII inhibitor, KN-93 (20 micromol/L), but not its inactive analog, KN-92, accelerated the inactivation of I(to) (tau(fast): 66.9+/-4.4 versus 43.0+/-4.4 ms, n=35; P<0.0001) and inhibited its maintained component (at +60 mV, 4.9+/-0.4 versus 2.8+/-0.4 pA/pF, n = 35; P<0. 0001), leading to an increase in the extent of its inactivation. Similar effects were observed by dialyzing cells with a peptide corresponding to CaMKII residues 281 to 309 or with autocamtide-2-related inhibitory peptide and by external application of the calmodulin inhibitor calmidazolium, which also suppressed the effects of KN-93. Furthermore, the phosphatase inhibitor okadaic acid (500 nmol/L) slowed I(to) inactivation, increased I(sus), and inhibited the effects of KN-93. Changes in [Ca(2+)](i) by dialyzing cells with approximately 30 nmol/L Ca(2+) or by using the fast Ca(2+) buffer BAPTA had opposite effects on I(to). In BAPTA-loaded myocytes, I(to) was less sensitive to KN-93. In myocytes from patients in chronic atrial fibrillation, characterized by a prominent I(sus), KN-93 still increased the extent of inactivation of I(to). Western blot analysis of atrial samples showed that delta-CaMKII expression was enhanced during chronic atrial fibrillation. In conclusion, CaMKII control the extent of inactivation of I(to) in human atrial myocytes, a process that could contribute to I(to) alterations observed during chronic atrial fibrillation. PMID- 10532950 TI - A flow-activated chloride-selective membrane current in vascular endothelial cells. AB - Shear stress-induced activation of endothelial ion channels, one of the earliest responses to flow, is implicated in mechano-signal transduction that results in the regulation of vascular tone. The effects of laminar flow on endothelial membrane potential were studied in vitro using both fluorescent potentiometric dye measurements and whole-cell patch-clamp recordings. The application of flow stimulated membrane hyperpolarization, which was reversed to depolarization within 35 to 160 seconds. The depolarization was caused by a Cl(-)-selective membrane current activated by flow independently of the K(+) channel-mediated hyperpolarization. Thus, flow activated both K(+) and Cl(-) currents, with the net membrane potential being determined by the balance of the responses. Membrane potential sensitivity to flow was unchanged by flow preconditioning that elongated and aligned the cells. PMID- 10532951 TI - Cytokine-mediated apoptosis in cardiac myocytes: the role of inducible nitric oxide synthase induction and peroxynitrite generation. AB - Increased production of nitric oxide (NO) after induction of the cytokine inducible isoform of nitric oxide synthase (iNOS or NOS2) in cardiac myocytes and other parenchymal cells within the heart may in addition to contributing to myocyte contractile dysfunction also contribute to the induction of programmed cell death (apoptosis). To investigate the mechanism(s) by which increased NO production leads to apoptosis, we examined the role of NO in primary cultures of neonatal rat ventricular myocytes (NRVMs) after induction by the cytokines interleukin-1beta (IL-1beta) and interferon gamma (IFNgamma) or exposure to the exogenous NO donor S-nitroso-N-acetylcysteine (SNAC) or peroxynitrite (ONOO(-)). Both SNAC (1 mmol/L) and ONOO(-) (100 micromol/L), but not their respective controls (ie, N-acetylcysteine and pH-inactivated ONOO(-)), induced apoptosis in confluent, serum-starved NRVMs at 48 hours. Similarly, incubation of NRVMs with IL-1beta and IFNgamma for 48 hours resulted in an increase in iNOS expression, nitrite production, and programmed cell death. Both the cytokine-induced nitrite accumulation and myocyte apoptosis could be completely prevented by the nonselective NOS inhibitor L-nitroarginine (3 mmol/L) or the specific iNOS inhibitor 2-amino-5, 6-dihydro-6-methyl-4H-1,3-thiazine (AMT, 100 micromol/L). NO mediated myocyte apoptosis was not attenuated by the inhibition of soluble guanylyl cyclase with ODQ, nor could apoptosis be induced by the incubation of NRVMs with 1 mmol/L 8-bromo-cGMP, a cell-permeant cGMP analogue. However, NO mediated apoptosis was significantly attenuated by the superoxide dismutase mimetic and ONOO(-) scavenger Mn(III)tetrakis (4-benzoic acid) porphyrin (MnTBAP, 100 micromol/L). NO/ONOO(-)-mediated apoptosis was associated with increased expression of Bax with no change in Bcl-2 mRNA abundance. Furthermore, apoptotic cell death was also confirmed in adult rat ventricular myocytes (ARVMs) when grown in heteroculture with IL-1beta- and IFNgamma-treated rat cardiac microvascular endothelial cells. Therefore, cytokine-induced apoptosis in NRVMs and ARVMs is mediated by iNOS induction, ONOO(-), and associated with an increase in Bax levels. PMID- 10532952 TI - In vivo modeling of myosin binding protein C familial hypertrophic cardiomyopathy. AB - Myosin binding protein C (MyBP-C) is an integral part of the striated muscle sarcomere. As is the case for other sarcomeric genes in human populations, multiple mutations within the gene have been linked to familial hypertrophic cardiomyopathy. Although some MyBP-C lesions are the result of missense mutations, most show truncated polypeptides lacking either the myosin or myosin and titin binding sites. Previously, we generated transgenic (TG) mice with cardiac-specific expression of a MyBP-C mutant lacking the myosin and titin binding domains. Surprisingly, the mutant protein was stable and made up a majority of the MyBP-C species, with concomitant reductions in endogenous MyBP-C such that overall MyBP-C stoichiometry was conserved. In the present study, we created a second series of TG mice that express, in the heart, a mutant MyBP-C lacking only the myosin binding site. In contrast to the previous data for the MyBP-C lacking both titin and myosin binding sites, only very modest levels of protein were found, consistent with data obtained from human biopsies in which mutated MyBP-C could not be detected. Despite normal levels of wild-type MyBP-C, there were significant changes in the structure and ultrastructure of the heart. Fiber mechanics showed decreased unloading shortening velocity, maximum shortening velocity, and relative maximal power output. PMID- 10532953 TI - Diminished basal phosphorylation level of phospholamban in the postinfarction remodeled rat ventricle: role of beta-adrenergic pathway, G(i) protein, phosphodiesterase, and phosphatases. AB - Three weeks after myocardial infarction (MI) in the rat, remodeled hypertrophy of noninfarcted myocardium is at its maximum and the heart is in a compensated stage with no evidence of heart failure. Our hemodynamic measurements at this stage showed a slight but insignificant decrease of +dP/dt but a significantly higher left ventricular end-diastolic pressure. To investigate the basis of the diastolic dysfunction, we explored possible defects in the beta-adrenergic receptor-G(s/i) protein-adenylyl cyclase-cAMP-protein kinase A-phosphatase pathway, as well as molecular or functional alterations of sarcoplasmic reticulum Ca(2+)-ATPase and phospholamban (PLB). We found no significant difference in both mRNA and protein levels of sarcoplasmic reticulum Ca(2+)-ATPase and PLB in post MI left ventricle compared with control. However, the basal levels of both the protein kinase A-phosphorylated site (Ser16) of PLB (p16-PLB) and the calcium/calmodulin-dependent protein kinase-phosphorylated site (Thr17) of PLB (p17-PLB) were decreased by 76% and 51% in post-MI myocytes (P<0.05), respectively. No change was found in the beta-adrenoceptor density, G(salpha) protein level, or adenylyl cyclase activity. Inhibition of phosphodiesterase and G(i) protein by Ro-20-1724 and pertussis toxin, respectively, did not correct the decreased p16-PLB or p17-PLB levels. Stimulation of beta-adrenoceptor or adenylyl cyclase increased both p16-PLB and p17-PLB in post-MI myocytes to the same levels as in sham myocytes, suggesting that decreased p16-PLB and p17-PLB in post-MI myocytes is not due to a decrease in the generation of p16-PLB or p17-PLB. We found that type 1 phosphatase activity was increased by 32% (P<0.05) with no change in phosphatase 2A activity. Okadaic acid, a protein phosphatase inhibitor, significantly increased p16-PLB and p17-PLB levels in post-MI myocytes and partially corrected the prolonged relaxation of the [Ca(2+)](i) transient. In summary, prolonged relaxation of post-MI remodeled myocardium could be explained, in part, by altered basal levels of p16-PLB and p17-PLB caused by increased protein phosphatase 1 activity. PMID- 10532955 TI - The role of cell death in heart failure. PMID- 10532954 TI - Myocyte death in the failing human heart is gender dependent. AB - Cardiovascular disease is delayed and less common in women than in men. Myocyte death occurs in heart failure, but only apoptosis has been documented; the role of myocyte necrosis is unknown. Therefore, we tested whether necrosis is as important as apoptosis and whether myocyte death is lower in women than in men with heart failure. Molecular probes were used to measure the magnitude of myocyte necrosis and apoptosis in 7 women and 12 men undergoing transplantation for cardiac failure. Myocyte necrosis was evaluated by detection of DNA damage with blunt end fragments, whereas apoptosis was assessed by the identification of double-strand DNA cleavage with single base or longer 3' overhangs. An identical analysis of these forms of cell death was performed in control myocardium. Heart failure showed levels of myocyte necrosis 7-fold greater than apoptosis in patients of both sexes. However, cell death was 2-fold higher in men than in women. Heart failure resulted in a 13-fold and 27-fold increase in necrosis in women and men, respectively. Apoptosis increased 35-fold in women and 85-fold in men. The differences in cell death between women and men were confirmed by the electrophoretic pattern of DNA diffusion and laddering of isolated myocytes. The lower degree of cell death in women was associated with a longer duration of the myopathy, a later onset of cardiac decompensation, and a longer interval between heart failure and transplantation. In conclusion, myocyte necrosis and apoptosis affect the decompensated human heart; each contributes to the evolution of cardiac failure. However, the female heart is protected, at least in part, from necrotic and apoptotic death signals. PMID- 10532956 TI - Parsing the effects of nitric oxide, S-nitrosothiols, and peroxynitrite on inducible nitric oxide synthase-dependent cardiac myocyte apoptosis. PMID- 10532957 TI - Sodium "channelopathies" and sudden death: must you be so sensitive? PMID- 10532958 TI - Circulation research online only : october 29, 1999UltraRapid CommunicationsA mouse model of arterial gene TransferAntigen-specific immunity is a minor determinant of the early loss of adenovirus-mediated transgene expression AB - We developed a murine model of arterial gene transfer and used it to test the role of antigen-specific immunity in the loss of adenovirus-mediated transgene expression. Adenoviral vectors encoding either beta-galactosidase (beta-gal) or green fluorescent protein were infused to the lumen of normal common carotids of CD-1 and C57BL/6 mice and atherosclerotic carotids of Apoe(-/-) mice. At 3 days after gene transfer, significant reporter gene expression was detected in all strains. Transgene expression was transient, with expression undetectable at 14 days. Next, a beta-gal-expressing vector was infused into carotids of ROSA26 mice (transgenic for, and therefore tolerant of, beta-gal) and RAG-2(-/-) mice (deficient in recombinase-activating gene [RAG]-2 and therefore lacking in antigen-specific immunity). beta-Gal expression was again high at 3 days but declined substantially (>90%) by 14 days. In vivo labeling with bromodeoxyuridine revealed that carotid endothelial proliferation was increased dramatically by the gene-transfer procedure alone, likely leading to the loss of episomal adenoviral DNA. Gene transfer to normal and atherosclerotic mouse carotids can be accomplished; however, elimination of antigen-specific immune responses does not prevent the early loss of adenovirus-mediated transgene expression. Efforts to prolong adenovirus-mediated transgene expression in the artery wall must be redirected. These efforts will likely include strategies to avoid the consequences of increased cell turnover. Nevertheless, despite the brevity of expression, this mouse model of gene transfer to normal and severely atherosclerotic arteries will likely be useful for investigating the genetic basis of vascular disease and for developing gene therapies. The full text of this article is available at http://www.circresaha. org. PMID- 10532959 TI - A mouse model of arterial gene transfer: antigen-specific immunity is a minor determinant of the early loss of adenovirus-mediated transgene expression. AB - We developed a murine model of arterial gene transfer and used it to test the role of antigen-specific immunity in the loss of adenovirus-mediated transgene expression. Adenoviral vectors encoding either beta-galactosidase (beta-gal) or green fluorescent protein were infused to the lumen of normal common carotids of CD-1 and C57BL/6 mice and atherosclerotic carotids of Apoe(-/-) mice. At 3 days after gene transfer, significant reporter gene expression was detected in all strains. Transgene expression was transient, with expression undetectable at 14 days. Next, a beta-gal-expressing vector was infused into carotids of ROSA26 mice (transgenic for, and therefore tolerant of, beta-gal) and RAG-2(-/-) mice (deficient in recombinase-activating gene [RAG]-2 and therefore lacking in antigen-specific immunity). beta-Gal expression was again high at 3 days but declined substantially (>90%) by 14 days. In vivo labeling with bromodeoxyuridine revealed that carotid endothelial proliferation was increased dramatically by the gene-transfer procedure alone, likely leading to the loss of episomal adenoviral DNA. Gene transfer to normal and atherosclerotic mouse carotids can be accomplished; however, elimination of antigen-specific immune responses does not prevent the early loss of adenovirus-mediated transgene expression. Efforts to prolong adenovirus-mediated transgene expression in the artery wall must be redirected. These efforts will likely include strategies to avoid the consequences of increased cell turnover. Nevertheless, despite the brevity of expression, this mouse model of gene transfer to normal and severely atherosclerotic arteries will likely be useful for investigating the genetic basis of vascular disease and for developing gene therapies. PMID- 10532960 TI - Molecular cloning and characterization of the intermediate-conductance Ca(2+) activated K(+) channel in vascular smooth muscle: relationship between K(Ca) channel diversity and smooth muscle cell function. AB - Recent evidence suggests that functional diversity of vascular smooth muscle is produced in part by a differential expression of ion channels. The aim of the present study was to examine the role of Ca(2+)-activated K(+) channels (K(Ca) channels) in the expression of smooth muscle cell functional phenotype. We found that smooth muscle cells exhibiting a contractile function express predominantly large-conductance ( approximately 200 pS) K(Ca) (BK) channels. In contrast, proliferative smooth muscle cells express predominantly K(Ca) channels exhibiting a much smaller conductance ( approximately 32 pS). These channels are blocked by low concentrations of charybdotoxin (10 nmol/L) but, unlike BK channels, are insensitive to iberiotoxin (100 nmol/L). To determine the molecular identity of this K(+) channel, we cloned a 1.9-kb cDNA from an immature-phenotype smooth muscle cell cDNA library. The cDNA contains an open reading frame for a 425 amino acid protein exhibiting sequence homology to other K(Ca) channels, in particular with mIK1 and hIK1. Expression in oocytes gives rise to a K(+)-selective channel exhibiting intermediate-conductance (37 pS at -60 mV) and potent activation by Ca(2+) (K(d) 120 nmol/L). Thus, we have cloned and characterized the vascular smooth muscle intermediate-conductance K(Ca) channel (SMIK), which is markedly upregulated in proliferating smooth muscle cells. The differential expression of these K(Ca) channels in functionally distinct smooth muscle cell types suggests that K(Ca) channels play a role in defining the physiological properties of vascular smooth muscle. PMID- 10532961 TI - Normal and mutant rhodopsin activation measured with the early receptor current in a unicellular expression system. AB - The early receptor current (ERC) represents molecular charge movement during rhodopsin conformational dynamics. To determine whether this time-resolved assay can probe various aspects of structure-function relationships in rhodopsin, we first measured properties of expressed normal human rhodopsin with ERC recordings. These studies were conducted in single fused giant cells containing on the order of a picogram of regenerated pigment. The action spectrum of the ERC of normal human opsin regenerated with 11-cis-retinal was fit by the human rhodopsin absorbance spectrum. Successive flashes extinguished ERC signals consistent with bleaching of a rhodopsin photopigment with a normal range of photosensitivity. ERC signals followed the univariance principle since millisecond-order relaxation kinetics were independent of the wavelength of the flash stimulus. After signal extinction, dark adaptation without added 11-cis retinal resulted in spontaneous pigment regeneration from an intracellular store of chromophore remaining from earlier loading. After the ERC was extinguished, 350-nm flashes overlapping metarhodopsin-II absorption promoted immediate recovery of ERC charge motions identified by subsequent 500-nm flashes. Small inverted R(2) signals were seen in response to some 350-nm flashes. These results indicate that the ERC can be photoregenerated from the metarhodopsin-II state. Regeneration with 9-cis-retinal permits recording of ERC signals consistent with flash activation of isorhodopsin. We initiated structure-function studies by measuring ERC signals in cells expressing the D83N and E134Q mutant human rhodopsin pigments. D83N ERCs were simplified in comparison with normal rhodopsin, while E134Q ERCs had only the early phase of charge motion. This study demonstrates that properties of normal rhodopsin can be accurately measured with the ERC assay and that a structure-function investigation of rapid activation processes in analogue and mutant visual pigments is feasible in a live unicellular environment. PMID- 10532962 TI - Properties of the mutant Ser-460-Cys implicate this site in a functionally important region of the type IIa Na(+)/P(i) cotransporter protein. AB - The substituted cysteine accessibility approach, combined with chemical modification using membrane-impermeant alkylating reagents, was used to identify functionally important structural elements of the rat type IIa Na(+)/P(i) cotransporter protein. Single point mutants with different amino acids replaced by cysteines were made and the constructs expressed in Xenopus oocytes were tested for function by electrophysiology. Of the 15 mutants with substituted cysteines located at or near predicted membrane-spanning domains and associated linker regions, 6 displayed measurable transport function comparable to wild-type (WT) protein. Transport function of oocytes expressing WT protein was unchanged after exposure to the alkylating reagent 2-aminoethyl methanethiosulfonate hydrobromide (MTSEA, 100 microM), which indicated that native cysteines were inaccessible. However, for one of the mutants (S460C) that showed kinetic properties comparable with the WT, alkylation led to a complete suppression of P(i) transport. Alkylation in 100 mM Na(+) by either cationic ([2 (trimethylammonium)ethyl] methanethiosulfonate bromide (MTSET), MTSEA) or anionic [sodium(2-sulfonatoethyl)methanethiosulfonate (MTSES)] reagents suppressed the P(i) response equally well, whereas exposure to methanethiosulfonate (MTS) reagents in 0 mM Na(+) resulted in protection from the MTS effect at depolarized potentials. This indicated that accessibility to site 460 was dependent on the conformational state of the empty carrier. The slippage current remained after alkylation. Moreover, after alkylation, phosphonoformic acid and saturating P(i) suppressed the slippage current equally, which indicated that P(i) binding could occur without cotransport. Pre-steady state relaxations were partially suppressed and their kinetics were significantly faster after alkylation; nevertheless, the remaining charge movement was Na(+) dependent, consistent with an intact slippage pathway. Based on an alternating access model for type IIa Na(+)/P(i) cotransport, these results suggest that site 460 is located in a region involved in conformational changes of the empty carrier. PMID- 10532963 TI - Divalent cation interactions with light-dependent K channels. Kinetics of voltage dependent block and requirement for an open pore. AB - The light-dependent K conductance of hyperpolarizing Pecten photoreceptors exhibits a pronounced outward rectification that is eliminated by removal of extracellular divalent cations. The voltage-dependent block by Ca(2+) and Mg(2+) that underlies such nonlinearity was investigated. Both divalents reduce the photocurrent amplitude, the potency being significantly higher for Ca(2+) than Mg(2+) (K(1/2) approximately 16 and 61 mM, respectively, at V(m) = -30 mV). Neither cation is measurably permeant. Manipulating the concentration of permeant K ions affects the blockade, suggesting that the mechanism entails occlusion of the permeation pathway. The voltage dependency of Ca(2+) block is consistent with a single binding site located at an electrical distance of delta approximately 0.6 from the outside. Resolution of light-dependent single-channel currents under physiological conditions indicates that blockade must be slow, which prompted the use of perturbation/relaxation methods to analyze its kinetics. Voltage steps during illumination produce a distinct relaxation in the photocurrent (tau = 5-20 ms) that disappears on removal of Ca(2+) and Mg(2+) and thus reflects enhancement or relief of blockade, depending on the polarity of the stimulus. The equilibration kinetics are significantly faster with Ca(2+) than with Mg(2+), suggesting that the process is dominated by the "on" rate, perhaps because of a step requiring dehydration of the blocking ion to access the binding site. Complementary strategies were adopted to investigate the interaction between blockade and channel gating: the photocurrent decay accelerates with hyperpolarization, but the effect requires extracellular divalents. Moreover, conditioning voltage steps terminated immediately before light stimulation failed to affect the photocurrent. These observations suggest that equilibration of block at different voltages requires an open pore. Inducing channels to close during a conditioning hyperpolarization resulted in a slight delay in the rising phase of a subsequent light response; this effect can be interpreted as closure of the channel with a divalent ion trapped inside. PMID- 10532964 TI - Synergistic activation of G protein-gated inwardly rectifying potassium channels by the betagamma subunits of G proteins and Na(+) and Mg(2+) ions. AB - Native and recombinant G protein-gated inwardly rectifying potassium (GIRK) channels are directly activated by the betagamma subunits of GTP-binding (G) proteins. The presence of phosphatidylinositol-bis-phosphate (PIP(2)) is required for G protein activation. Formation (via hydrolysis of ATP) of endogenous PIP(2) or application of exogenous PIP(2) increases the mean open time of GIRK channels and sensitizes them to gating by internal Na(+) ions. In the present study, we show that the activity of ATP- or PIP(2)-modified channels could also be stimulated by intracellular Mg(2+) ions. In addition, Mg(2+) ions reduced the single-channel conductance of GIRK channels, independently of their gating ability. Both Na(+) and Mg(2+) ions exert their gating effects independently of each other or of the activation by the G(betagamma) subunits. At high levels of PIP(2), synergistic interactions among Na(+), Mg(2+), and G(betagamma) subunits resulted in severalfold stimulated levels of channel activity. Changes in ionic concentrations and/or G protein subunits in the local environment of these K(+) channels could provide a rapid amplification mechanism for generation of graded activity, thereby adjusting the level of excitability of the cells. PMID- 10532966 TI - The urology of Pharaonic Egypt. PMID- 10532967 TI - Extra-anatomic stents in ureteric obstruction: experience and complications. AB - OBJECTIVE: To assess the role of extra-anatomic stents (EAS) as a means of urinary diversion in patients with ureteric obstruction secondary to malignancy. PATIENTS AND METHODS: The technique for inserting EAS in patients with ureteric obstruction was described previously; to date, 13 patients (seven women and six men, mean age 45.3 years, range 22-78) have been treated. All patients had ultrasonographic evidence of hydronephrosis and/or significant biochemical evidence of renal impairment. Patients had advanced malignancy and one patient an abdominal aortic aneurysm. RESULTS: Urinary diversion was successful in all patients; two survived for more than 1 year, with stent changes at 6-monthly intervals. In three patients the stents were replaced by percutaneous nephrostomies because of problems with leakage or infection. The remaining patients died with functioning EAS in situ. CONCLUSIONS: In patients with ureteric obstruction secondary to malignancy or medical conditions excluding them from more invasive surgery, EAS provide a further therapeutic option instead of a permanent nephrostomy, which has associated inherent problems. This technique is not without potential problems and careful selection of patients remains vital in this difficult area. PMID- 10532968 TI - Laparoscopic ureterolithotomy: the Edinburgh experience. AB - OBJECTIVE: To review our experience with laparoscopic ureterolithotomy. PATIENTS AND METHODS: Since 1993, we have performed laparoscopic ureterolithotomy in 14 patients with ureteric stones. Laparoscopy was carried out in nine patients as a salvage procedure after failed ureteroscopy (six), shock wave lithotripsy (two), or both (one), and in five patients as a primary procedure for large stones (mean 27.2 mm, range 18-40). Patients in the former group had already undergone a mean of 1.88 procedures (range 1-4). Laparoscopic ureterolithotomy was carried out via a transperitoneal approach. Associated ureteric strictures were incised at the time of ureterotomy. RESULTS: All procedures were completed laparoscopically and all patients were rendered stone-free after a single procedure. The mean operative duration was 105 min. Ureteric strictures were incised in three patients, in two of whom dilatation was subsequently required; all three had a successful result. There were three minor complications. CONCLUSIONS: Laparoscopic ureterolithotomy can be a safe and effective procedure; it should be considered as a primary procedure for large mid- and upper ureteric stones. PMID- 10532965 TI - A mutation linked with Bartter's syndrome locks Kir 1.1a (ROMK1) channels in a closed state. AB - Mutations in the inward rectifying renal K(+) channel, Kir 1.1a (ROMK), have been linked with Bartter's syndrome, a familial salt-wasting nephropathy. One disease causing mutation removes the last 60 amino acids (332-391), implicating a previously unappreciated domain, the extreme COOH terminus, as a necessary functional element. Consistent with this hypothesis, truncated channels (Kir 1.1a 331X) are nonfunctional. In the present study, the roles of this domain were systematically evaluated. When coexpressed with wild-type subunits, Kir 1.1a 331X exerted a negative effect, demonstrating that the mutant channel is synthesized and capable of oligomerization. Plasmalemma localization of Kir 1.1a 331X green fluorescent protein (GFP) fusion construct was indistinguishable from the GFP wild-type channel, demonstrating that mutant channels are expressed on the oocyte plasma membrane in a nonconductive or locked-closed conformation. Incremental reconstruction of the COOH terminus identified amino acids 332-351 as the critical residues for restoring channel activity and uncovered the nature of the functional defect. Mutant channels that are truncated at the extreme boundary of the required domain (Kir 1.1a 351X) display marked inactivation behavior characterized by frequent occupancy in a long-lived closed state. A critical analysis of the Kir 1.1a 331X dominant negative effect suggests a molecular mechanism underlying the aberrant closed-state stabilization. Coexpression of different doses of mutant with wild-type subunits produced an intermediate dominant negative effect, whereas incorporation of a single mutant into a tetrameric concatemer conferred a complete dominant negative effect. This identifies the extreme COOH terminus as an important subunit interaction domain, controlling the efficiency of oligomerization. Collectively, these observations provide a mechanistic basis for the loss of function in one particular Bartter's causing mutation and identify a structural element that controls open-state occupancy and determines subunit oligomerization. Based on the overlapping functions of this domain, we speculate that intersubunit interactions within the COOH terminus may regulate the energetics of channel opening. PMID- 10532969 TI - Primary in situ extracorporeal shock wave lithotripsy in the management of ureteric calculi: results with a third-generation lithotripter. AB - OBJECTIVE: To review the results of primary in situ extracorporeal shock wave lithotripsy (ESWL) for the treatment of ureteric stones using a third-generation lithotripter, the Dornier MFL 5000 (Dornier Medizentechnic, Germany). PATIENTS AND METHODS: The study comprised a retrospective review of treatment outcome in 180 patients with 196 stones who were treated with primary in situ ESWL, assessing the success of this approach and establishing reasons for failure. RESULTS: At the 3-month follow-up, 88% of patients were stone-free; 21 patients failed ESWL and were treated by ureteroscopic stone extraction with no complications. Stone-free rates were 90% for upper ureteric, 89% for middle-third and 86% for lower-third calculi. Twenty-one patients required auxiliary procedures in the form of JJ stenting or nephrostomy. Failure of ESWL was associated with stone size (>1.3 cm) but not location or inadequate treatment. CONCLUSION: Where prompt access to ESWL is available, primary in situ ESWL remains an effective form of treatment for all ureteric calculi, although stone free rates are lower for larger stones. PMID- 10532970 TI - Chromosomal aberrations in transitional cell carcinoma that are predictive of disease outcome are independent of polyploidy. AB - OBJECTIVE: To determine whether aneusomy for chromosomes 7, 9 and 17 (reported to predict recurrence in up to 65% of patients with superficial transitional cell bladder cancer and thus providing the opportunity for early and effective treatment) reflects specific genetic events on these chromosomes or merely wider unspecific genetic damage to the cell, e.g. that increased copy numbers for 7 and 17 reflect tumour polyploidy. MATERIALS AND METHODS: The study comprised 25 primary tumours; 6 microm thick sections from formalin-fixed and paraffin embedded tumours were analysed. Chromosome copy numbers were determined by fluorescence in situ hybridization (FISH) using pericentromeric probes for chromosomes 7, 8, 9, 10, 11 and 17. A minimum of 200 nuclei per tumour area were scored by two independent observers. RESULTS: Eight of the 25 tumours examined (32%) showed no evidence of chromosomal abnormalities as detected by FISH for any chromosomes analysed. Twelve tumours (48%) showed abnormalities for one or two chromosomes, five tumours (20%) showed abnormalities for multiple chromosomes and one tumour showed abnormalities for all chromosomes analysed, suggestive of polyploidy. CONCLUSIONS: Chromosomal abnormalities predictive of recurrence occur largely in the absence of other gross chromosomal lesions. In a small proportion of cases other chromosomes are affected, but this is almost always distinct from tumour polyploidy. PMID- 10532971 TI - Assessment of a new bone anchor system for the treatment of female genuine stress incontinence. AB - OBJECTIVE: To evaluate the efficacy of the In-Tac bone-anchor system (using shape memory metal bone anchors, Influence Medical Technologies, Lancs, UK) as a vaginal procedure for the treatment of female genuine stress incontinence (GSI). PATIENTS AND METHODS: Between January 1997 and April 1998, 30 patients with GSI were recruited into the study (age range 36-74 years); patients who had undergone previous failed continence surgery were not excluded. All patients underwent a urodynamic assessment before and 3 months after surgery. All In-Tac bone-anchor procedures were performed under general anaesthesia. Patients were reviewed after surgery at 6 weeks, 3, 6 and 12 months, and yearly thereafter. RESULTS: The mean (range) operative duration was 42 (20-75) min and the blood loss 60 (10-200) mL (median 30). There were no intraoperative complications and minimal analgesia was required postoperatively. At 6 weeks, 27 patients (90%) were subjectively cured. The urodynamic assessment at 3 months revealed that 22 patients (73%) were objectively cured; at 6 months and one year the subjective cure rate was 80%. CONCLUSION: The In-Tac bone-anchor system is simple and safe, the procedure easily learned and the operation brief. It offers promise as an incision-less vaginal procedure that may have wide application for the treatment of women with GSI. A longer follow-up is needed to fully confirm its durability and effectiveness. PMID- 10532972 TI - Failure of allograft suburethral slings. AB - OBJECTIVES: To report our experience of using freeze-dried irradiated fascia lata allografts for suburethral sling procedures. PATIENTS AND METHODS: Between December 1996 and September 1998, 35 patients (mean age 60.25 years, range 37-79) underwent suburethral sling placement with fascia allograft. These patients were reviewed, with the findings at the time of any surgical re-exploration. Eleven (31%) had undergone prior surgery for genuine stress incontinence and 32 (91%) had a preoperative diagnosis of intrinsic sphincter deficiency. RESULTS: On re operation for persistent or recurrent stress incontinence, the allograft was present but grossly degenerated in two (6%) patients and completely absent in five (14%) patients. Histology of a retrieved graft fragment showed both fibroblast proliferation and degeneration within the graft. CONCLUSION: The use of freeze-dried, irradiated fascia lata for suburethral sling procedures was associated with a material failure rate of >/=20%. We caution against its use in this setting. PMID- 10532973 TI - Intraurethral sphincter prosthesis to treat hyporeflexic bladders in women: does it work? AB - OBJECTIVE: To assess the efficacy and safety of a new intraurethral sphincter prosthesis to treat hyporeflexic bladders in women. PATIENTS AND METHODS: Between July 1997 and December 1998, 18 women (mean age 45.8 years, range 26-84) with neurogenic voiding disorders and a hyporeflexic bladder were examined prospectively. All but one patient (who used the Crede manoeuvre) emptied their bladder using clean intermittent catheterization. The women were fitted with a prosthesis consisting of a valve and a pump inside a short self-retaining silicone device, which was activated by a magnetic remote control unit. The evaluation before implantation comprised a medical history, a neurological evaluation, urodynamic recordings and urine cultures. The follow-up after implantation included a monthly clinical assessment, a symptom questionnaire, urine culture, ultrasonographic examinations and replacement of the device if necessary. Urine samples were also cultured if there was any discomfort or fever. RESULTS: At 16 months of follow-up, only six of 18 patients continued to use the implant (mean follow-up 9.6 months) and were satisfied. In 10 patients major incontinence around the catheter, or irritation, led to removal of the device. Two patients were unable to learn how to transfer to the toilet to empty their bladder; therefore, their implant was removed and they were treated with a suprapubic catheter. Two patients died from unrelated causes. Twelve patients had positive bacteriuria while fitted with the device; six of them became symptomatic. Usually, only symptomatic lower urinary tract infections were treated. Technical problems occurred often; 14 catheters showed technical dysfunction and had to be replaced early and three external remote control units broke for no reason. Despite normal short-term cystoscopic findings (three patients), long-term urethral damage cannot be excluded. Indeed, the six patients who remain fitted with the device already show some widening of their urethra. CONCLUSION: This experience with the new device was disappointing; in eight of the 18 patients incontinence appeared or worsened and led to removal of the device. Technical problems were common. The other concern is that significant long-term urethral damage could be expected, as observed with indwelling catheters over time. Thus, although this device may function for a short period, it is unsuitable for long-term use. PMID- 10532974 TI - How is follow-up after transurethral prostatectomy best performed? AB - OBJECTIVE: To evaluate the role of the nurse practitioner (NP) in screening patients for potential discharge after routine transurethral prostatectomy (TURP) or bladder neck incision (BNI) where, although urologists continue to follow such patients, the trend is away from clinic attendance. PATIENTS AND METHODS: The NP telephoned 70 patients 4 weeks after surgery; information about expected postoperative problems, change in symptoms and the need to visit their general practitioner (GP) was recorded. A doctor then saw all the patients in a clinic 3 months after TURP or BNI. RESULTS: Complete records were available for 66 patients (TURP 56, BNI 10). Four weeks after their operation, 39 (59%) patients still had one or more significant symptoms but only nine (23%) had consulted their GP. After a telephone interview the NP considered that 38 of the 66 patients were fit to be discharged. At the 3-month outpatient appointment, 37 of these 38 patients were subsequently discharged. Of the remaining 29 patients, 15 (seven with carcinoma of the prostate and eight with significant symptoms) were given follow-up appointments. CONCLUSIONS: The persistence of significant symptoms in 12% of patients 3 months after TURP justifies the follow-up of all patients. A telephone interview by the NP at one month is recommended. This could result in safe discharge of more than half the patients and allow follow-up of those who need specialist input. PMID- 10532975 TI - The follow-up of patients with unfavourable early results of transurethral prostatectomy. AB - OBJECTIVE: To determine the natural course of patients with subjectively disappointing early results after transurethral prostatectomy (TURP), who experience prolonged discomfort and an initial deterioration in symptoms. PATIENTS AND METHODS: A consecutive series of 127 patients undergoing urodynamic studies and TURP were assessed 3 months after surgery using symptom scores and measurements of urinary flow rate; 107 patients reported improved symptom and quality-of-life scores, but 20 did not improve, with no change or a deterioration. These 20 patients were followed for several months using symptom scores, and measurements of flow rates and residual urine volumes. Baseline variables, including preoperative urodynamic studies, were compared between those who improved and those who did not. RESULTS: Over a mean (range) follow-up of 10.6 (6-15) months, all those initially not improving showed spontaneous improvement in all three variables with no further treatment and eventually achieved the same significant degree of improvement as those who improved soon after TURP. Preoperatively, those initially not improving had mean lower symptom scores, more bladder irritability and less obstruction than did those who improved. CONCLUSION: A significant proportion (approximately 15%) of patients with obstructive symptoms will experience considerable symptomatic discomfort for a prolonged period after an uncomplicated TURP and will not gain the full symptomatic benefit from the procedure until 6-9 months afterward. PMID- 10532976 TI - Hybrid laser treatment compared with transurethral resection of the prostate for symptomatic bladder outlet obstruction caused by a large benign prostate: a prospective, randomized trial with a 6-month follow-up. AB - OBJECTIVE: To compare the efficacy and safety of hybrid laser treatment, i.e. the combination of visual Nd-YAG laser ablation of prostate and contact Nd-YAG laser vaporization of prostate, with transurethral resection of the prostate (TURP) in the treatment of patients with symptomatic bladder outlet obstruction secondary to a benign high-volume prostate. PATIENTS AND METHODS: Forty-five symptomatic patients with hyperplastic prostates of >40 mL were randomized to undergo either hybrid laser treatment (21) or TURP (24). All patients were evaluated before and after treatment with a complex urodynamic assessment, and were accepted into the study only if they had infravesical obstruction in the pressure-flow study. In the hybrid method, Nd-YAG laser energy was first delivered by an 'adenoma dependent' approach to all areas of the obstructing lateral lobe tissue through a side-firing gold-alloy tip fibre at 40 W for 90 s of 'burn'. The prostatic urethra was then opened and the median lobe vaporized using the a contact probe at 40 W. Patients were re-evaluated 3 and 6 months after treatment. RESULTS: Both treatments proved to be safe, and improved the subjective and objective outcome measures at 3 and 6 months compared with baseline values. After 3 months, there was a greater improvement in the TURP group in peak urinary flow rate (Qmax; P<0.01), mean urinary flow rate (Qave; P<0.01) and postvoid residual urine volume (P<0.05) than in the hybrid laser group. After 6 months, there was a greater improvement in the TURP group in detrusor pressure at Qmax (P<0.01), Qave (P<0.05) and prostate size (P<0.001) than in the hybrid laser group. In the pressure-flow study at 6 months, a higher proportion of patients (seven of 19) were still obstructed in the hybrid laser group than in TURP group (two of 21; P<0.05). TURP caused more intraoperative blood loss (P<0.001) and postoperative problems associated with bleeding; 38% of hybrid laser patients were discharged with a suprapubic catheter, whereas all TURP patients could urinate at discharge (P<0.01). The duration of bladder drainage was longer after hybrid laser treatment (P<0.001). CONCLUSION: The hybrid laser method was a safe but less effective treatment than TURP for benign prostatic enlargement in patients with prostates of >40 mL. PMID- 10532977 TI - The stability of free and bound prostate-specific antigen. AB - OBJECTIVE: To determine if the assay for free prostate specific antigen (fPSA) and the calculated ratio of fPSA to total PSA (f/tPSA) is stable in conditions likely to be met in routine clinical practice. MATERIALS AND METHODS: Two blood samples were obtained from 27 patients attending a routine urology clinic. Sample 1 was centrifuged immediately, assayed for fPSA and tPSA, and the f/tPSA calculated. This sample was then stored at 4 degrees C for 24 h, 48 h and 1 week, or at -20 degrees C for 24 h, 1 week and 1 month before the assays for fPSA and tPSA were repeated. The second sample was left at room temperature for 24 h before assay and processing, as for sample 1. RESULTS: tPSA is a highly stable analyte; if whole blood samples are processed immediately, fPSA is stable for 24 h at 4 degrees C and 1 month at -20 degrees C. There was a significant reduction in the calculated f/tPSA in samples stored for >/=24 h at 4 degrees C (P<0.01); if the sample was stored at -20 degrees C the calculated f/tPSA was stable. After 24 h storage at room temperature, fPSA decreased by 6.3% and f/tPSA by 6.4%. Subsequent storage of serum at 4 degrees C for 1 week resulted in a 25% decrease from the baseline value. After 1 month at -20 degrees C the fPSA value was 13% lower than the baseline value. CONCLUSION: These results indicate that if there is to be confidence in the accuracy of the f/tPSA value, then blood samples must be handled and processed correctly. Total PSA is sufficiently stable to permit whole blood samples to remain at room temperature for 24 h before serum is separated. If fPSA is to be determined accurately then the whole blood sample must be centrifuged promptly. As the fPSA values in blood samples left at room temperature for 24 h are up to 25% lower than those on immediate assay, and the subsequent f/tPSA 29% lower, then for the optimum use of this test, these samples should also be handled appropriately. PMID- 10532978 TI - Efficacy and limitations of delayed/salvage radiation therapy after radical prostatectomy. AB - OBJECTIVE: To assess the kinetics of prostate specific antigen (PSA) and the degree of PSA suppression, to better understand the efficacy and limitations of delayed/salvage radiation therapy after radical prostatectomy. PATIENTS AND METHODS: The PSA doubling time was calculated in patients with biochemical failure after radical prostatectomy and in those who underwent delayed/salvage radiation therapy. Patients in whom PSA was undetectable by conventional assay after irradiation were followed using a hypersensitive PSA assay. RESULTS: Of 125 patients who underwent radical prostatectomy for clinically resectable prostate cancer, 47 developed biochemical failure at a mean of 11.8 months after surgery; 38 of these patients underwent radiotherapy (36 for isolated biochemical failure and two for local progression with elevated PSA levels). The mean (sd) PSA doubling time after surgery was 14.6 (16.2) months (n=44) and after radiation therapy it was 13.3 (23.9) months (n=32). Eleven of 30 evaluable patients (37%) had a sustained PSA suppression lasting at least 12 months after radiotherapy. Only the time to biochemical failure after surgery approached statistical significance for predicting a durable response to radiotherapy (P=0.08). The mean nadir value of PSA in the 11 patients with at least 12 months of sustained PSA suppression was 0.032 ng/mL at 26.9 months. CONCLUSIONS: The rapidity with which PSA levels double after surgery may provide a clinically significant indication of the nature of these recurrent tumours, which deserve the best possible attempt at cure. Slow-growing tumours with longer PSA doubling times may be better candidates for delayed/salvage radiation therapy. Larger studies involving more patients are needed to determine whether the PSA doubling time can define subgroups for which specific treatment strategies should be developed. PMID- 10532979 TI - An interstitial light assembly for photodynamic therapy in prostatic carcinoma. AB - OBJECTIVE: To develop an interstitial laser light delivery system using multiple optical fibres for photodynamic therapy (PDT) in the treatment of prostate cancer. PATIENTS AND METHODS: A laser beam was divided equally with a 1 x 4 fibre splitter to deliver PDT simultaneously through four 2-cm long, flexible cylindrical optical diffusers. Biplanar transrectal ultrasonography (TRUS) and a template were used to position the optical fibres percutaneously. In vivo measurements of light penetration depth (1/micro[eff] ) in prostate tissue were made in seven patients, using a sheathed isoprobe to measure light fluence rates at varying radial distances from the diffuser. The prostate was fixed with stabilization needles to minimize displacement during needle placement. RESULTS: The mean (sd, range) micro(eff) in the prostates of the seven patients was 0.35 (0.07, 0.22-0.44) mm-1, which produced closely parallel slopes of light attenuation. However, there was up to a 10-fold variation in absolute light levels at the same diffuser-detector separation distances amongst the seven patients, probably caused by blood pooling around the diffuser light source. A similar problem around the isoprobe detector was overcome by sheathing the probe in clear plastic tubing. By stabilizing the prostate, the optical fibre positioning was precise to within 2 mm. CONCLUSION: Although this light delivery and TRUS assembly were developed for clinical PDT in the prostate, the same instrumentation can be used reliably for in vivo light-penetration studies. Haemorrhage was unpredictable and highlighted one of the main problems which needs to be overcome. PMID- 10532981 TI - A comparison of the sperm mixed-agglutination reaction test with the peroxidase labelled protein A test for detecting antisperm antibodies in infertile men with varicocele. AB - OBJECTIVE: To compare the sperm mixed-agglutination reaction (sMAR) with the peroxidase-labelled protein A method (POPA) in infertile patients with varicocele. PATIENTS, SUBJECTS AND METHODS: The study comprised 30 men with a history of varicocele-associated infertility and 30 fertile men (control group). Antisperm antibodies against spermatozoa in the semen and against progenitor spermatozoa in testicular tissue were detected using the two methods. RESULTS: The tests were positive in 15 (50%) of patients with both the sMAR and the POPA methods, while no autoantibodies were detected in the control group. There were no significant differences between the methods. The sensitivity and specificity of both tests was approximately 93%, with no significant difference between them (P>0. 05). CONCLUSION: Both methods may be used for detecting sperm autoantibodies in infertile patients with varicocele. PMID- 10532980 TI - A double-blind, randomized, controlled multicentre study to compare the efficacy of ciprofloxacin with pivampicillin as oral therapy for epididymitis in men over 40 years of age. AB - OBJECTIVE: To compare the efficacy and safety of ciprofloxacin 500 mg orally twice daily with pivampicillin 700 mg orally twice daily for 10 days in men with acute epididymitis and over 40 years of age. PATIENTS AND METHODS: The study comprised 172 men who entered a prospective, controlled, randomized, double blind, trial of pivampicillin and ciprofloxacin. The median (range) age of the 158 patients eligible for the efficacy analysis was 58 (41-85) years; 41% had previously had a urinary tract infection and 27% had previously had epididymitis. Only one patient had a urethral catheter and 38% were sexually active. About half of the patients were admitted to hospital. RESULTS: No bacteria could be cultured from samples in 53% of the patients; Escherichia coli could be cultured from 35% and the remaining isolates were the expected urinary pathogens. None of the patients had Gonococci and only one in each group had Chlamydia. Mycoplasma hominis was detected in three patients only and M. genitalium was detected in three, while Ureaplasma was detected in 24 (15%). The treatment failed in 48 patients; in 15 of 76 (20%) receiving ciprofloxacin and in 33 of 82 (40%) receiving pivampicillin. This corresponds to a reduction in the risk of failure of 20.5% (95% confidence limits 6.6-40.2%, P=0. 006). The principal cause of failure was an unsatisfactory clinical response requiring changed antibiotic treatment in 27 patients; adverse events were responsible for failure in 14. The in vitro resistance of cultured bacteria was low in both groups, at approximately 4%. Adverse events, mainly gastro-intestinal, occurred in 17 of 83 (21%) patients starting on ciprofloxacin and in 33 of 89 (37%) receiving pivampicillin (P=0.04). CONCLUSION: For epididymitis in men over the age of 40 years ciprofloxacin 500 mg orally twice daily is more effective than pivampicillin 700 mg orally twice daily. Furthermore, ciprofloxacin has a lower incidence of adverse events. PMID- 10532982 TI - The preventive effect of systemic treatment with interferon-alpha2B for infertility from mumps orchitis. AB - OBJECTIVE: To evaluate the effect of interferon-alpha2B on mumps orchitis, often caused by postpubertal mumps and which can result in permanent testicular atrophy. PATIENTS AND METHODS: The study included 21 patients with mumps orchitis, treated between May 1990 and June 1997. Patients were randomly assigned into two groups: in group 1, 13 patients received therapy with interferon-alpha2B (3 x 10(6) IU per day) and group 2 did not, acting as controls. All were evaluated by measurements of testis size, mumps virus titre, hormone level and semen analysis. RESULTS: In group 1, the patients' symptoms resolved within 2-3 days and the volume of the testes returned to normal within 11 days; there was no testicular atrophy in any patient during the follow-up. However, asthenospermia continued to be detected in four patients (unilateral in two, bilateral in two). In group 2, the patients' symptoms resolved within 5-6 days and the volume of the testes returned to normal within 10 days; testes atrophied in three patients (unilateral in two, bilateral in one) during the follow-up. Asthenospermia continued in four patients (unilateral in two, bilateral in two). CONCLUSION: These results suggest that treatment with systemic interferon-alpha2B is effective in preventing testicular atrophy when combined with standard symptomatic treatment. PMID- 10532983 TI - Suture tracks after hypospadias repair. AB - OBJECTIVE: To determine the incidence of suture tracks after hypospadias repair in which 6-0 chromic catgut was used to close the skin. PATIENTS AND METHODS: From an initial series of 72 boys undergoing tubularized, incised-plate hypospadias repair, 23 (32%) were evaluated 1 year or more after surgery. RESULTS: Of the 23 boys, 10 (43%) were found to have suture tracks. In all cases tracks were located on the ventral shaft skin and did not involve the glans or dorsal aspect of the circumcision. These tracks had not been detected at an earlier follow-up. CONCLUSIONS: Tracks resulting from small absorbable sutures may not be apparent soon after surgery because the sinus is small. With time, the sinuses fill with keratin and thereby become obvious. The location of these tracks suggests that relative hypovascularity of ventral shaft skin may contribute to their development. The use of subcutaneous sutures may diminish the incidence of this minor complication of hypospadias surgery. PMID- 10532984 TI - Effects of a phytotherapeutic agent, PC-SPES, on prostate cancer: a preliminary investigation on human cell lines and patients. AB - OBJECTIVES: To evaluate the in vitro activity of PC-SPES, a complex phytotherapeutic agent, against prostate cancer cell lines, and to assess its activity in suppressing serum prostate specific antigen (PSA) level in patients with prostate cancer. PATIENTS AND METHODS: Four variant prostate cancer cell lines (LNCaP and an apoptosis-resistant derivative, LNCaP-bcl-2, PC3 and DU145) were exposed to three different concentrations of PC-SPES extract. Cell viability was measured at 3, 4 and 5 days of exposure using a colorimetric assay and was compared with control cultures receiving aliquots of the ethanolic extraction medium alone. Clinically, a prospective study was initiated in patients with prostate cancer who refused conventional therapy or who had failed previous cryosurgery, radiation therapy and/or hormonal therapy. The patients were treated with PC-SPES (three capsules of 320 mg/day). The serum PSA responses and side effects were evaluated. RESULTS: All cultured prostate cancer cell lines showed a significant dose-dependent reduction in cellular viability (compared with control cultures) by exposure to 4 and 6 microL of PC-SPES extract/mL of culture medium (P<0.001). In contrast to the hormone-insensitive cell lines tested (LNCaP-bcl-2, PC-3 and DU-145), only the hormone-sensitive cell line LNCaP was affected by the lowest dose of PC-SPES extract tested (2 microL/mL medium). In the prospective clinical trial of 33 patients, with a mean (range) follow-up of 6.8 (2-24) months after initiating PC-SPES therapy, serum PSA levels were lower in 87% at 2 months and in 78% at 6 months (n=18, P=0.026). The side-effects in these patients were nipple tenderness in two (6%) and leg clots requiring heparinization in two (6%). No gynaecomastia or hot flashes were observed in this group and the treatment was well tolerated. CONCLUSIONS: In this preliminary study, an extract of the phytotherapeutic agent PC-SPES was active in suppressing the growth of cultured hormone-sensitive and -insensitive prostate cancer cell lines. In the small clinical study, PC-SPES therapy decreased serum PSA levels in most patients. However, a longer follow-up and more patients will be required to evaluate the long-term efficacy of this new phytotherapy. PMID- 10532985 TI - Mast cell variations in tumour tissue and with histopathological grading in specimens of prostatic adenocarcinoma. AB - OBJECTIVE: To determine whether the number of mast cells (MCs) varies in and around prostatic carcinoma tissue, and with histopathological grading. MATERIALS AND METHODS: Specimens of 27 prostatic carcinomas were stained with the toluidine blue and staged histopathologically using the Gleason grading system. The MCs were counted in five high-power microscopy fields within and around the tissue samples, and the results analysed using the Wilcoxon signed-rank test and Spearman's correlation. RESULTS: The mean number of MCs was 1.7 per field within and 3.31 per field around the cancer tissue, the difference between these being significant (P<0. 05). There was a close correlation between the numbers of MCs within the tumour and Gleason grade (r=0.56, P=0.002) but not between grade and counts around the tumour (r=-0.18, P=0.35). CONCLUSION: These results suggest that some MCs aggregate at the periphery of the prostatic adenocarcinoma and that the number of MCs within the tumour tissue is related to tumour differentiation. PMID- 10532986 TI - Purinergic sensory neurotransmission in the urinary bladder: an in vitro study in the rat. AB - OBJECTIVES: To determine the response of mechanosensitive pelvic nerve afferents, arising from the rat urinary bladder, to the purinergic agonist alpha,beta methylene ATP and to the purinergic antagonist suramin. MATERIALS AND METHODS: Using a newly developed in vitro bladder-pelvic nerve afferent model, multiunit recordings were taken from mechanosensitive pelvic nerve afferents arising from the rat urinary bladder, in response to bladder distension. Control experiments were performed by distending the bladder with saline at 0.04 mL/min, and recording the total afferent nerve activity and the bladder pressure response to the distension. Bladder distensions were then repeated using a solution of the stable purinergic agonist alpha,beta-methylene ATP (10 micromol/L), which is known to desensitize P2X-purinoceptors after prolonged exposure, and the total afferent activity and bladder pressure response were again measured. In a separate series of experiments the afferent nerve activity and bladder pressure response to bladder distension with saline was determined in the presence of the purinergic antagonist suramin (10 micromol/L) and repeated after washout of the drug. In both series of experiments, afferent nerve responses were compared with control using the paired t-test, whilst the bladder pressure responses were compared using one-way analysis of variance. RESULTS: Bladder distension with alpha,beta-methylene-ATP produced a statistically significant reduction in afferent nerve activity, by up to 75% compared with the control, whilst having no significant effect on the bladder pressure response. Bladder distension with saline in the presence of suramin (10 micromol/L) produced a significant reduction in the resultant afferent nerve activity, by 50%, which returned to normal after washout of the drug. CONCLUSION: These findings are consistent with the notion that ATP is released endogenously during bladder distension in the rat and is involved significantly in the activation of pelvic nerve afferents arising from the rat urinary bladder. PMID- 10532987 TI - Pharmacological and urodynamic changes in rat urinary bladder function after multiple pregnancies. AB - OBJECTIVE: To investigate the changes in bladder function after multiple pregnancies and parturition in rats, and to establish links between the changes in voiding profiles and in vitro pharmacological responses. MATERIALS AND METHODS: Cystometry was used in conscious virgin and multiparous female Wistar rats (2 weeks after the last parturition) with chronically implanted lines to continuously record bladder pressure during five reproducible voiding cycles. In vitro, detrusor muscle contractile responses induced by cumulative concentrations of KCl, carbachol, noradrenaline and alpha, beta-methylene-ATP (mATP) were compared. RESULTS: In multiparous rats, there was a significant increase in the amplitude of voiding pressure (+31%), bladder capacity (+83%) and residual volume (about threefold); 60% of the multiparous rats but only 10% of the virgin rats showed bladder instability during the filling phase. Cumulative concentration response curves to KCl, expressed as the tension developed vs tissue weight, were identical in the two groups of rats. Contractile responses induced by carbachol (0.1-30 micromol/L) were significantly larger in multiparous than in virgin rats. Similarly, noradrenaline-induced contractions (0.3-10 micromol/L) were significantly higher for multiparous animals. However, the sensitivity of the detrusor muscle to mATP was not modified by multiple pregnancies. CONCLUSION: After multiple gestations, female rats develop bladder hypertrophy, bladder instabilities and a higher amplitude of voiding pressure associated with an increased residual volume. These altered patterns are similar to those found in rats after chronic infravesical outlet obstruction. We propose that pregnancies and parturition modify urinary bladder function, leading to a dysfunction similar to that induced by obstruction, and involving an increased sensitivity to adrenergic and cholinergic stimulation. PMID- 10532988 TI - Testicular prosthesis placement: a new technique. PMID- 10532989 TI - Unusual phimosis: a simple surgical solution. PMID- 10532990 TI - Recurrent granular cell tumour of the bladder in a patient with von Recklinghausen's disease. PMID- 10532991 TI - Infrahepatic renal cell carcinoma tumour thrombus: vena caval reconstruction with ovarian vein. PMID- 10532992 TI - Right pelvic ectopic kidney with pelvi-ureteric obstruction causing contralateral obstruction to kidney and ureter: a novel presentation of a pelvic ectopic kidney. PMID- 10532993 TI - Synchronous malignant melanoma of the male bulbar urethra and transitional cell carcinoma of the bladder. PMID- 10532994 TI - Gossypiboma: migration of retained surgical gauze and spontaneous transurethral protrusion. PMID- 10532995 TI - Posterior urethral polyp in a boy, diagnosed by colour Doppler ultrasonography. PMID- 10532996 TI - Gastroureteroplasty in a woman with bilateral ureteric strictures after pelvic radiotherapy. PMID- 10532997 TI - Traumatic urethral diverticula. PMID- 10532998 TI - Subumbilical ectopic testis. PMID- 10532999 TI - Long-term survival after surgery for recurrent advanced sarcomatoid renal carcinoma. PMID- 10533000 TI - Urinary diversion in childhood: indications for different techniques. PMID- 10533001 TI - Refluxing or nonrefluxing ureteric anastomosis. PMID- 10533002 TI - The management of ureteric stones. Part I: diagnosis. PMID- 10533003 TI - The management of ureteric stones. Part II: therapy. PMID- 10533004 TI - Flow requirements in ventricular fibrillation: An in vivo nuclear magnetic resonance analysis of the left ventricular high-energy phosphate pool. AB - STUDY OBJECTIVE: We sought to determine whether flow rates of approximately 60% of normal values are sufficient to preserve the left ventricular myocardial high energy phosphate pool during ventricular fibrillation (VF). METHODS: Mixed-breed swine (weight 22. 4+/-2.5 kg) were anesthetized with alpha-chloralose, placed in a state of VF, and perfused with extracorporeal circulation at a target flow of 50 mL.kg(-1).min(-1). In vivo whole-wall (average of left ventricular wall) and spatially localized phosphorous-31 nuclear magnetic resonance (NMR) spectra were acquired at baseline and during VF. RESULTS: Mean flow during VF was 58+/-20 mL.kg(-1). min(-1) (+/-SD; 95% confidence interval, 44 to 71) or about 60% of baseline cardiac output (n=13). Whole-wall adenosine triphosphate (ATP) decreased during perfused VF (P <.05), whereas creatine phosphate (CP) remained unchanged from baseline. With spatially localized NMR, the ratios of CP/ATP were similar at baseline in all layers (endocardium --> epicardium) of the left ventricular wall. However, during perfused VF, subepicardial CP/ATP ratios increased by 14% to 40% compared with baseline values, whereas subendocardial CP/ATP ratios remained unchanged (1% to 3% increase). An additional 4 animals perfused at 72+/-10 mL.kg( 1).min(-1) (+/-SD; 95% confidence interval, 56 to 92) during VF had preservation of CP and ATP levels. CONCLUSION: Flow levels equivalent to 60% of baseline cardiac output were insufficient to maintain normal high-energy phosphate levels in the in vivo fibrillating myocardium. At this level of flow, myocardial high energy phosphate loss is nonhomogeneous within the left ventricular wall. PMID- 10533005 TI - History and physical examination to estimate the risk of ectopic pregnancy: validation of a clinical prediction model. AB - STUDY OBJECTIVE: To prospectively validate a clinical prediction model for ectopic pregnancy (EP). METHODS: Prospective cohort with 14-month derivation and 12-month validation phases. All hemodynamically stable, first-trimester patients with abdominal pain or vaginal bleeding who presented to a military teaching hospital emergency department underwent follow-up until an outcome of intrauterine pregnancy (IUP) or EP was established. Patients were separated into the high-risk group, defined as having either peritoneal signs or definite cervical motion tenderness; intermediate-risk group, defined as the presence of pain or tenderness, other than midline cramping, plus absence of fetal heart tones, and absence of tissue visible at the cervical os; and low-risk group (neither high- nor intermediate-risk) using recursive partitioning. RESULTS: Summarizing both phases, 915 patients had 845 (93%) IUPs and 70 (7.6%) EPs, with 18 (1.9%) lost to follow-up. The clinical prediction model classified 75 (8.2%) into the high-risk group (sensitivity 31%, 95% confidence interval [CI] 21% to 44%; specificity 94%, 95% CI 92% to 95%); and 644 (70%) in the intermediate-risk group (sensitivity 98%, 95% CI 89% to 100%; specificity 25%, 95% CI 22% to 29%). The remaining 196 (21%) patients who met neither high-risk nor intermediate-risk criteria were classified into the low-risk group. On the basis of EP prevalence of 7.7%, the risk of EP was less than 1% (95% CI 0% to 3%) for the low-risk group, 7% (95% CI 5% to 10%) for the intermediate-risk group, and 29% (95% CI 19% to 41%) for the high-risk group. CONCLUSION: This clinical prediction model is useful for estimating the risk of EP in first-trimester patients, particularly when ancillary testing is equivocal or not readily available. PMID- 10533006 TI - Older patients' health-related quality of life around an episode of emergency illness. AB - STUDY OBJECTIVES: We sought to describe older patients' health-related quality of life during a 4-month period surrounding a visit to the emergency department and to identify factors associated with less recovery. METHODS: We prospectively studied 983 patients 65 years or older who presented to an urban academic ED in 1995 and 1996. Eighty percent of the patients were African American, and 63% were women. The primary outcome measures were the Katz Index of Activities of Daily Living and revised validated questions from the Medical Outcomes Study Health Survey at 1 month before the ED visit, the time of the ED visit, and 2-week and 3 month follow-up periods. RESULTS: In general, patients worsened markedly during the illness and then improved, although not to baseline levels. After adjustment for demographic and social factors, the most consistently powerful predictors of poor recovery were more deficiencies in activities of daily living at baseline, reports of needing more help with everyday tasks, increasing Charlson Comorbidity Index scores, and requiring a proxy for the initial survey. CONCLUSION: Emergency physicians and primary care physicians should consider inquiring about functional status and the adequacy of help at home in addition to comorbid conditions for their acutely ill older patients to target those at greatest risk for poor recovery. Future work needs to test interventions that may improve the health related quality of life of these vulnerable patients. PMID- 10533007 TI - A comparison of emergency medicine ultrasound training with guidelines of the Society for Academic Emergency Medicine. AB - STUDY OBJECTIVES: To compare the current state of emergency medicine residency ultrasound training with guidelines for that training from the Society for Academic Emergency Medicine (SAEM). METHODS: A brief questionnaire was sent to program directors from 119 emergency medicine residency programs in the United States. Responses were compared with the SAEM guidelines for clinical experience (150 total ultrasounds) and didactic experience (40 hours of didactic instruction). RESULTS: The overall response rate was 92%. Seventy-six (69%) of the programs own an ultrasound machine (ownership defined as 24-hour availability and complete discretion over use). Of these, 12 (16%) indicated that their average 1998 graduate had done at least 150 total ultrasound scans during residency, although none of the programs had average numbers that exceeded the minimum guidelines for all 4 procedure categories. Information on didactic curriculum was available from 74 ultrasound-owning programs: the duration was 0 to 20 hours in 49 (66%), 20 to 40 hours in 19 (26%), and 40 to 100 hours in 6 (8%). Only 1 program's average graduate met or exceeded the SAEM guidelines for both didactic and clinical training. CONCLUSION: Most emergency medicine residency programs own at least 1 ultrasound machine, with more than half of these obtaining their first machine within the past 3 years. Only 1 program currently meets SAEM training guidelines. PMID- 10533008 TI - How well do patients obtain short-term follow-up after discharge from the emergency department? AB - STUDY OBJECTIVE: We sought to determine the follow-up rate of discharged emergency department patients who were instructed to obtain reevaluation within 48 hours at our ED, a clinic, or a private physician's office and to determine the reasons why patients do not obtain short-term follow-up when instructed. METHODS: Emergency physicians prospectively enrolled a convenience sample of patients discharged from a university hospital ED who were believed to be at risk for clinical deterioration. Patients were instructed to obtain reevaluation within 48 hours at a public clinic, private physician's office, or our ED (without charge). A telephone interview was conducted after 48 hours had elapsed. RESULTS: Three hundred twenty-five patients were enrolled, 300 were included in data analysis, and 203 (67.7%) of these obtained follow-up as instructed. Those referred to the ED had a higher follow-up rate (105/127 [82.7%]) than those referred to clinics (59/99 [59.6%]) or private physicians (39/74 [52.7%]). Inability to obtain an appointment was cited by 34.3% of those who did not obtain follow-up care as instructed. CONCLUSION: Many patients discharged from the ED who were believed to be at risk for clinical deterioration did not obtain medical follow-up within 48 hours when so instructed. Free ED follow-up resulted in a better rate of short-term follow-up than that for clinics and private physicians and may be especially useful if a patient's ability to obtain follow-up is uncertain or if timely reevaluation is particularly imperative. PMID- 10533009 TI - Incidence of immediate and delayed hypersensitivity to Centruroides antivenom. AB - STUDY OBJECTIVE: To assess the incidence and course of immediate and delayed hypersensitivity to Centruroides antivenom. METHODS: We performed a 12-month prospective observation study, with telephone follow-up, evaluating the incidence of anaphylaxis or anaphylactoid reactions and serum sickness after Centruroides antivenom administration. The setting for the study was a poison control center and tertiary care toxicology treatment center. Participants included all patients who received Centruroides antivenom, and no interventions were performed. RESULTS: For immediate hypersensitivity reactions, 116 patients with grade III or IV envenomation received Centruroides antivenom; 77 of these patients were younger than 13 years. Three patients completed the infusion despite development of rash. A fourth patient with a history of atopy and asthma received epinephrine infusion and an inhaled beta-agonist for transient wheezing that quickly resolved; she was admitted for observation. Nine patients without hypersensitivity reactions were admitted for social reasons, for inappropriate sedation from drugs used before antivenom, or to rule out aspiration; all were discharged within 24 hours. The remaining 106 patients were discharged from the emergency department after resolution of symptoms. Thus 4 of 116 patients had immediate reactions. For patients with delayed reactions, 17 patients were lost to follow-up. Of 99 remaining patients, serum sickness developed in 61% (n=60), as defined by using liberal criteria. Serum sickness responded to oral steroids, antihistamines, or both; mean duration of symptoms with medication was 2.8 days. CONCLUSION: Anaphylactic reactions are uncommon after Centruroides antivenom infusion. Self-limited serum sickness that is easily controlled with corticosteroids and antihistamines commonly follows the use of Centruroides antivenom. PMID- 10533010 TI - Continuous intravenous midazolam infusion for Centruroides exilicauda scorpion envenomation. AB - STUDY OBJECTIVE: We sought to describe the effects of continuous intravenous midazolam infusion as therapy for severe bark scorpion (Centruroides exilicauda) envenomation. METHODS: A retrospective chart review from July 1, 1993, through January 1, 1998, identified all patients treated at a university hospital with International Classification of Diseases, Ninth Revision, codes 989.5 (toxic effect of venom) or E905.2 (scorpion sting causing poisoning). By using standardized collection forms, data were extracted from the medical record of every patient who had a grade III or IV envenomation and was treated with a continuous intravenous midazolam infusion. RESULTS: Our search identified 104 patients; 34 had grade III or IV envenomation. Of these, 33 were treated in the ICU with continuous intravenous midazolam infusion. Median patient age was 4 years (range, 1 to 68 years). Midazolam dosage was adjusted to induce a light sleep state to control agitation and involuntary motor activity. The median amount of midazolam resulting in the first recorded decrease in agitation and involuntary motor activity was 0.30 mg/kg (range, 0.03 to 1.76 mg/kg). This first evidence of clinical improvement was recorded as 1.00 hour (median), with a range of 0.00 to 3.75 hours. The initial midazolam infusion rate was 0.10 mg x kg(-1) x h(-1) (median), with a range of 0.01 to 0.31 mg x kg(-1) x h(-1). The maximal midazolam infusion rate was 0.30 mg x kg(-1) x h(-1) (median), with a range of 0.06 to 1.29 mg x kg(-1) x h(-1). The median time until the maximal midazolam infusion rate was 2.5 hours (range, 0.00 to 8.50 hours). The median duration of infusion was 9. 50 hours (range, 4.25 to 20.50 hours). The median length of stay in the ICU was 15.17 hours (range, 6.0 to 28.0 hours), and 85% of patients were discharged directly home. All patients had resolution of abnormal motor activity and agitation during their midazolam infusion. Transient hypoxemia without evidence of end-organ dysfunction was documented in 4 patients during midazolam therapy. CONCLUSION: A continuous intravenous midazolam infusion can be a safe, effective, and readily available treatment option for patients with grade III or IV C exilicauda envenomation. PMID- 10533011 TI - Pregnancy-associated injury hospitalizations in Pennsylvania, 1995. AB - STUDY OBJECTIVE: To estimate the frequency of pregnancy-associated injury hospitalization and compare rates between pregnant women and all women of reproductive age by age, race, injury mechanism, intent, and other variables. METHODS: Using International Classification of Diseases, Ninth Revision-Clinical Modification (ICD-9-CM) selection criteria applied to Pennsylvania's 1995 acute hospital discharge data, all resident women ages 15 to 44 with coexistent pregnancy and injury-related diagnoses were identified for descriptive and comparative rate calculations. RESULTS: Seven hundred sixty-one (4. 6%) of the discharges to injured women of reproductive age were associated with pregnancy. The leading injury causes among pregnant women were transportation-related (234 [33.6%]), falls (192 [26.4%]), poisonings (116 [16.0%]), and "struck by" (83 [11.4%]). Among all women 15 to 44 years, poisoning was the leading cause (32.6%) of injury, followed by transportation-related injuries (25.7%). The hospitalized injury incidence was 868 per 100,000 person-years for pregnant women versus 641 for all women ages 15 to 44 (rate ratio 1. 35, 95% confidence interval [CI] 1.25 to 1.45). Pregnant women were younger (median age 24.9 years versus 30.0 years), their mean length of stay was shorter (2.5 days versus 3.7 days), the mean injury severity score was less (3.2 versus 4.8), and the median charge per stay was lower ($4,164 versus $6,051). Rate ratios (pregnant versus all women in same age group) were significantly higher for younger women 15 to 19 years (rate ratio 2.69, 95% CI 2.49 to 3.14). Rate ratios were significantly higher for assaults (rate ratio 3.04, 95% CI 2.45 to 3.78), falls (rate ratio 2.33, 95% CI 2.01 to 2.70), motor vehicle occupant (rate ratio 2.0, 95% CI 1.73 to 2.31), and struck by (rate ratio 3.73, 95% CI 2.97 to 4.69). Rate ratios were lower for poisonings (rate ratio 0.71, 95% CI 0.59 to 0.86) and self-inflicted injuries (rate ratio 0.62, 95% CI 0.50 to 0.77). CONCLUSION: Pregnant women were more likely than all women 15 to 44 years to be hospitalized for injury and more likely to be hospitalized for assaults, falls, transportation-related, and less severe injuries, but less likely for poisonings and self-inflicted injuries. Much of the increased risk appears to be concentrated in young women. Further work is needed to establish to what extent the observed increases are the result of increased injury rates or increased hospitalization rates. PMID- 10533013 TI - Methemoglobinemia: etiology, pharmacology, and clinical management. AB - Methemoglobin (MHb) may arise from a variety of etiologies including genetic, dietary, idiopathic, and toxicologic sources. Symptoms vary from mild headache to coma/death and may not correlate with measured MHb concentrations. Toxin-induced MHb may be complicated by the drug's effect on other organ systems such as the liver or lungs. The existence of underlying heart, lung, or blood disease may exacerbate the toxicity of MHb. The diagnosis may be complicated by the effect of MHb on arterial blood gas and pulse oximeter oxygen saturation results. In addition, other dyshemoglobins may be confused with MHb. Treatment with methylene blue can be complicated by the presence of underlying enzyme deficiencies, including glucose-6-phosphate dehydrogenase deficiency. Experimental antidotes for MHb may provide alternative treatments in the future, but require further study. PMID- 10533012 TI - Expanding the National Electronic Injury Surveillance System to monitor all nonfatal injuries treated in US hospital emergency departments. AB - STUDY OBJECTIVE: Injury is a major cause of morbidity and mortality in the United States. Although the National Vital Statistics System provides data on injury related deaths, a national surveillance system is needed for timely identification of emerging nonfatal injury problems and continuous monitoring of severe nonfatal injuries. This work assesses the feasibility of expanding the National Electronic Injury Surveillance System (NEISS) to monitor all types and causes of nonfatal injuries treated in US hospital emergency departments and reports national estimates generated by a pilot study of this system. METHODS: At a stratified sample of US hospital EDs, persons receiving first-time treatment for an injury were monitored from May 1 through July 31, 1997. National estimates of the annual number and rate of ED-treated injuries overall, by patient characteristics, injury diagnosis, and external cause of injury were generated, and the sensitivity of the system for detecting ED-treated injuries was assessed. RESULTS: An estimated 29. 1 million injuries were treated in US EDs in 1997 (rate of 108.6/1, 000 population). The leading causes of injury were falls, being struck by or striking against an object or person, cutting or piercing, and motor vehicle traffic. Of 593 cases of injury detected by investigators from the Centers for Disease Control and Prevention during visits to 6 of the 21 NEISS hospitals in the study, 490 were also detected by NEISS coders for an overall sensitivity of 82.6%. CONCLUSION: Expanding the NEISS is a feasible means of timely and continuous monitoring of all types and causes of nonfatal injuries treated in US hospital EDs. PMID- 10533014 TI - Evidence-based emergency medicine: updates, feedback, and links. PMID- 10533015 TI - Application of likelihood ratios to clinical decision rules: defining the limits of clinical expertise. PMID- 10533016 TI - Evidence-based emergency department discharge planning: how do we get there? PMID- 10533017 TI - A scorpion by any other name is still a scorpion. PMID- 10533018 TI - Rectus sheath hematoma. AB - We describe 3 patients with rectus sheath hematoma presenting to the emergency department. Prompt consideration of this uncommon cause of abdominal pain may prevent more expensive and invasive diagnostic tests and, in some cases, unnecessary hospitalization and laparotomy. PMID- 10533019 TI - Rapid reversal of life-threatening diltiazem-induced tetany with calcium chloride. AB - We describe a patient who developed tetany with sudden respiratory arrest after the infusion of intravenous diltiazem. The administration of calcium chloride rapidly resolved the patient's tetany with prompt recovery of respiratory function, averting the need for more aggressive airway management and ventilatory support. The emergency physician should be aware that life-threatening tetany may accompany the administration of intravenous diltiazem and that calcium chloride may be a rapid and effective remedy. PMID- 10533021 TI - A bunker in the storm PMID- 10533020 TI - Update on emerging infections from the Centers for Disease Control and Prevention. Outbreak of Vibrio parahaemolyticus infection associated with eating raw oysters and clams harvested from Long Island Sound--Connecticut, New Jersey, and New York, 1998. AB - This article is part of a regular series on emerging infections from the Centers for Disease Control and Prevention (CDC) and the EMERGEncy ID NET, an emergency department-based and CDC-collaborative surveillance network. Important infectious disease public health information with relevance to emergency physicians is reported. The goal of this series is to advance knowledge about communicable diseases in emergency medicine, and foster cooperation between the front line of clinical medicine and public health agencies. PMID- 10533022 TI - Occult pneumonias in febrile children with leukocytosis. PMID- 10533023 TI - Wellens' syndrome. PMID- 10533024 TI - Polytopic anomalies with agenesis of the lower vertebral column. AB - We describe clinical, pathological and radiological findings in 15 cases of sporadic and familial lower spine agenesis with additional anomalies of the axial skeleton and internal organs and speculate about the cause and pathogenesis of this malformation complex. We show that all of these findings are defects of blastogenesis, originate in the primary developmental field and/or the progenitor fields, thus representing polytopic field defects. This concept appears applicable in our cases and makes such terms such as "caudal regression syndrome" or "axial mesodermal dysplasia spectrum" redundant. PMID- 10533025 TI - Caudal dysgenesis in staged human embryos: Carnegie stages 16-23. AB - The severity of expression of malformations of the median axis in the caudal region of human embryos is highly variable and ranges from caudal dysgenesis and sirenomelia to simple sacral hypoplasia. Several forms of sacral dysgenesis may be discovered later in life. This shows that caudal malformations of relatively lesser severity should occur at a greater frequency than actually reported. In the present study we looked at the morphology and histology of some human embryos with caudal dysgenesis. Several developmental alterations of the median axis were observed. These included significant reduction in the craniofacial mesenchyme characterized by hypoplasia of the pharyngeal arches, palatal shelves, and agenesis or hypoplasia of the auricular hillocks at the rostral end, absence of the caudal trunk from midsacral to all coccygeal segments, vertebral fusion or agenesis, defective development of the primary and secondary neural tubes, rectal and urinary tract dysgenesis, and deficiency, malrotation, and deficiency of the limbs at the caudal end. Hindlimb malformations included bilateral agenesis (one case), meromelia, and various forms of abnormal rotation, but no instances of sirenomelia were present. Radial dysgenesis has been reported to be associated with caudal dyplasia in the literature, however, we observed agenesis of the ulna in one and of the fibula in another embryo. There was an impressive association between limb malformations and body wall defects. The histological studies demonstrated caudal vascular deficiency and hemorrhagic lesions in the limbs of the dysplastic embryos. The data suggest that these polytopic field defects arise very early in development possibly as result of disturbances to fundamental developmental events that share common molecular and cellular mechanisms. PMID- 10533026 TI - Filippi syndrome: report of three additional cases. AB - Filippi syndrome is an autosomal recessive condition characterized by variable soft tissue syndactyly of the fingers and toes, microcephaly, pre- and postnatal growth retardation, mildly abnormal craniofacial appearance, and mental retardation. We report on three unrelated individuals with Filippi syndrome. All have microcephaly, minor facial anomalies, variable syndactyly of digits, growth impairment, and developmental delay. One patient also has polydactyly, which has not been reported previously in the Filippi syndrome. PMID- 10533027 TI - Skin elastic fibers in Williams syndrome. AB - The elastin gene is consistently deleted in Williams syndrome and as this protein represents the major component of the elastic fibers of the dermis, we sought to investigate skin elastic fibers in Williams syndrome as a key to unraveling extracellular matrix disorganization in this condition. Both morphometric parameters analyzed by using automated image analysis and immunofluorescence labeling with monoclonal antibodies against elastin and fibrillin 1 showed a disorganized pre-elastic (oxytalan and elaunin) and mature elastic fibers in the dermis of 10 Williams syndrome patients compared with five healthy children and one patient with isolated supravalvular aortic stenosis. Skin biopsies in Williams syndrome patients provide a simple mean to elucidate extracellular matrix anomalies. Hopefully, this method could give clues to the understanding of the elastic network anomalies in this condition and even to the consequences of these latter on elasticity and resilience of other tissues such as the arterial tree. PMID- 10533028 TI - Rare interstitial deletion (2)(p11.2p13) in a child with pericentric inversion (2)(p11.2q13) of paternal origin. AB - An unbalanced 46,XY,der(2)del(2)(p11.2p13) inv(2)(p11.2q13) karyotype was found in a phenotypically abnormal child with a de novo interstitial deletion of band 2p12 associated with an inv(2)(p11.2q13) inherited from the father. The inv(2) is generally considered a benign familial variant without significant reproductive consequences. However, our findings led us to consider a previously proposed mechanism of unequal meiotic crossing over at the base of a parental inversion loop, which could lead to either a deletion or duplication of a segment adjacent to the inverted region in the offspring. This phenomenon has been reported in other inversions of chromosomes 1, 7, 13, 15, and 17 and may explain the origin of the deletion in our patient. Although repetitive sequences might be present around such inversions, which could predispose to de novo deletions independently of the inversion, current evidence including this case favors a proposed causal relationship between the parental inversion and the deletion in the child. Our review and results suggest there could be a small risk for a related imbalance to couples with an inv(2)(p11.2q13). For del(2)(p11.2p13), which is rare, a more distinct phenotype has been proposed herein. Our patient shared several findings with the three previously published cases, namely the broad nasal bridge, abnormal ears, high-arched palate, psychomotor retardation, and micrognathia. However, our patient also had sensorineural hearing loss and significant hypotonia, which have not been previously reported, thereby expanding our understanding of this rare deletion. Am. J. Med. Genet. 87:139-142, 1999. Published 1999 Wiley-Liss, Inc. PMID- 10533029 TI - Apparent lability of neural tube closure in laboratory animals and humans. AB - Neural tube defects (NTDs), a set of structural abnormalities affecting the brain, spinal cord, and the skeletal and connective tissues that protect them, are common malformations among humans and laboratory animals. The embryogenesis of the neural tube is presented to convey the complexity of the phenomenon, the multiplicity of requisite cellular and subcellular processes, and the precise timing of events that must occur for successful neural tube development. Interruption, even transitory, of any of these intricate processes or disruption of an embryo's developmental schedule can lead to an NTD. The population distribution of human NTDs demonstrates that genetic predisposition functions in susceptibility to NTDs. Data from animal studies support these concepts. NTDs are common outcomes in developmental toxicity safety assessments, occurring among control and treated groups. Numerous agents have caused increased levels of NTDs in laboratory animals, and species with shorter gestational periods appear more prone to toxicant-induced NTDs than those with longer gestations. Data from post implantation whole embryo culture, although not predictive of human risk, are useful in studying neurulation mechanisms and in demonstrating the importance of maintaining embryonic schedules of development. We conclude that the concept that NTDs are produced by only a few toxicants that selectively target the developing nervous system is untenable. Rather, the combination of the time in gestation that an agent is applied, its dose, and its ability to disrupt critical processes in neurulation leads to NTDs. We further conclude that, because of both the relatively high prevalence and the multifactorial nature of NTDs, the mere occurrence of an NTD is insufficient for inferring that the defect was caused by an exogenous agent. PMID- 10533030 TI - Two distinct phenotypes caused by two different missense mutations in the same codon of the VHL gene. AB - We have identified a family segregating von Hippel-Lindau (VHL) disease with a previously unreported T547A mutation in exon 1 of the VHL gene that causes a Tyr112 to Asn missense alteration in the protein. The mutation was identified by nucleotide sequencing and confirmed by restriction enzyme digestion. The mutation cosegregated with the disease in all five tested affected individuals from the extended family. The family consists of more than 100 at-risk individuals over seven generations. To date, we have identified 13 affected individuals of whom seven have had renal cell carcinoma and one has had a pheochromocytoma. No other case of a neuroendocrine tumor of the pancreas or adrenal gland (pheochromocytoma) was found or recognized retrospectively. Other manifestations in this family include retinal angioma and hemangioblastoma of the central nervous system. We also found the T547A mutation in three asymptomatic members of the family, ages 12, 19, and 20. Another mutation, T547C, which causes Tyr112 to His, has been seen at the same position and has been associated with VHL type 2A (pheochromocytoma, but no renal cell carcinoma) in two families with a total of 22 affected individuals [Chen F, Slife L, Kishida T, Mulvihill J, Tisherman SE, Zbar B, 1996: J Med Genet 33:716-717]. Thus, different amino acid changes at the same position can cause very distinct clinical phenotypes. It will be interesting to elucidate the functional differences that underlie the different phenotypes. PMID- 10533031 TI - Clinical and genetic study of Friedreich ataxia in an Australian population. AB - Friedreich ataxia is an autosomal recessive disorder caused by mutations in the FRDA gene that encodes a 210-amino acid protein called frataxin. An expansion of a GAA trinucleotide repeat in intron 1 of the gene is present in more than 95% of mutant alleles. Of the 83 people we studied who have mutations in FRDA, 78 are homozygous for an expanded GAA repeat; the other five patients have an expansion in one allele and a point mutation in the other. Here we present a detailed clinical and genetic study of a subset of 51 patients homozygous for an expansion of the GAA repeat. We found a correlation between the size of the smaller of the two expanded alleles and age at onset, age into wheelchair, scoliosis, impaired vibration sense, and the presence of foot deformity. There was no significant correlation between the size of the smaller allele and cardiomyopathy, diabetes mellitus, loss of proprioception, or bladder symptoms. The larger allele size correlated with bladder symptoms and the presence of foot deformity. The duration of disease is correlated with wheelchair use and the presence of diabetes, scoliosis, bladder symptoms and impaired proprioception, and vibration sense but no other complications studied. PMID- 10533033 TI - Association of microphthalmia with esophageal atresia: report of two new patients and review of the literature. AB - We report on two new patients who had unilateral microphthalmia and esophageal atresia. A similar association was previously described in six patients. The accumulation of the eight affected patients provides further support for recognizing this association as a distinct syndrome. PMID- 10533032 TI - Familial patent ductus arteriosus and bicuspid aortic valve with hand anomalies: a novel heart-hand syndrome. AB - The association between cardiac and limb defects, particularly those affecting the hand, has been well documented by the delineation of several heart-hand syndromes. Based on observations with a three-generation family with seven affected individuals, we describe a novel heart-hand syndrome comprising patent ductus arteriosus, bicuspid aortic valve, 5th metacarpal hypoplasia, and brachydactyly. The inheritance pattern was consistent with autosomal dominance, although X-linked dominance could not be excluded. Penetrance appeared to be complete, but there was variability of the cardiac and hand phenotypes. Because this new syndrome closely resembled Char syndrome (patent ductus arteriosus, 5th finger middle phalangeal hypoplasia, and minor facial anomalies), multipoint linkage analysis was performed using polymorphic DNA markers spanning the recently identified Char syndrome critical region at chromosomal bands 6p12 p21.1. This analysis formally excluded this 3-cM region, documenting that the two traits are not allelic. In sum, a novel heart-hand syndrome involving left ventricular outflow and aortic arch as well as an ulnar ray derivative has been identified. Because the hand anomalies can be subtle, thorough evaluation is suggested for families inheriting these cardiac defects as a mendelian trait. PMID- 10533034 TI - Postnatal structure of the sella turcica in Down syndrome. AB - The purpose of this study was to analyze the shape of the sella turcica in a group of patients with Down syndrome and compare the findings with those made earlier in human fetuses with Down syndrome. Profile radiographs from 78 patients (age 4 months to 50 3/12 years) were analyzed. A tracing was made of each sella turcica, and the shape was compared with that of a normal sella, including the normal growth pattern from childhood to adulthood. Sella turcica structure could be classified into three morphological types, defined as: type I, almost normal appearance; type II, deviations in the anterior wall; and type III, deviations in the floor of the sella turcica. Compared with previously registered prenatal structural deviations in the sella turcica, it can be concluded that the postnatal radiographic material reflects the prenatal findings, because type I, both prenatally and postnatally, is by far the most common, whereas the remaining types are uncommon, both prenatally and postnatally. The study confirms the relevance of prenatal investigations for postnatal diagnostics as previously documented in sella turcica analyses of prenatal and postnatal myelomeningocele cases. PMID- 10533035 TI - Submicroscopic Xpter deletion in a boy with growth and mental retardation caused by a familial t(X;14). AB - In a 3-year-old boy with short stature, developmental delay, and dry skin, steroid sulphatase deficiency and a submicroscopic terminal deletion of Xp were found. Except for the short stature, no major clinical signs of X-linked recessive chondrodysplasia punctata could be observed. His mother had lowered steroid sulphatase activity compatible with carriership for X-linked ichthyosis and a submicroscopic translocation (X;14)(p22.31;p11.1). This finding combined with a normal amplification of exons 1, 5, and 10 of the STS gene from propositus' DNA suggested a breakpoint upstream of the STS gene. The submicroscopic maternal translocation had important implications for genetic counseling. This case report illustrates that contiguous gene syndrome related to the Xpter region may have an atypical clinical presentation and the usefulness of combined clinical, biochemical, molecular, and fluorescence in situ hybridization analysis. PMID- 10533036 TI - Inlet ventricular septal defect is not a partial atrioventricular septal defect. PMID- 10533038 TI - Role of the C677T polymorphism at the MTHFR gene on risk to nonsyndromic cleft lip with/without cleft palate: results from a case-control study in Brazil. PMID- 10533040 TI - Corelease of two functionally opposite neurotransmitters by retinal amacrine cells: experimental evidence and functional significance. AB - The Dale's law postulates that a neuron releases the same neurotransmitter from all its branches. In the case of multiple neurotransmitters it would require all transmitters to be released from all branches. The retinal cholinergic amacrine cells contain and release gamma-aminobutyric (GABA) and, therefore, if GABA and acetylcholine (ACh) are released at the same sites, this could mean that amacrine cells simultaneously excite and inhibit postsynaptic cells. Conversely, if the two neurotransmitters are released at different synapses, or if their release is regulated in a distinct manner, they may play different physiological roles. Recent studies carried out in cultured cholinergic amacrine-like neurons showed that Ca(2+)-dependent release of ACh and GABA have a different sensitivity to membrane depolarization, to the effect of blockers of voltage gated Ca(2+) channels (VGCC) and to the effect of presynaptic A(1) adenosine receptors. Therefore, it is proposed that in retinal amacrine cells the Ca(2+)-dependent release of ACh and GABA occurs at distinct cellular locations. The possible nature of these release sites and the physiological significance of this model are discussed in this review. PMID- 10533041 TI - Subcellular fractionation and association with the cytoskeleton of messengers encoding myelin proteins. AB - The targeting of polypeptides to restricted cytoplasmic domains by means of mRNA sorting is a widespread phenomena utilized by many cell types. In the central nervous system, in situ hybridization analysis has shown previously that the mRNAs encoding several myelin-specific proteins are specifically located within the myelinating processes of oligodendrocytes. Here, by means of biochemical and subcellular fractionation methods, we show that a myelin fraction is selectively enriched in those mRNAs. The four major myelin basic protein (MBP) mRNAs that arise by alternative splicing of exons II and VI of the MBP gene are concentrated in this subcellular fraction. Furthermore, an interaction of MBP and MOBP 81A mRNAs with the cytoskeleton was observed. This interaction might serve to mediate the anchoring of these messengers after translocation to the subcellular site of translation. PMID- 10533042 TI - Quantitation of spinal cord demyelination, remyelination, atrophy, and axonal loss in a model of progressive neurologic injury. AB - Spinal cord pathology, such as demyelination and axonal loss, is a common feature in multiple models of central nervous system (CNS) injury and disease. Development of methods to quantify spinal cord pathology objectively would aid studies designed to establish mechanisms of damage, correlate pathology with neurologic function, and assess therapeutic interventions. In this study, we describe sensitive methods to objectively quantify spinal cord demyelination, remyelination, atrophy, and axonal loss following the initiation of a progressive inflammatory demyelinating disease with Theiler's murine encephalomyelitis virus (TMEV). Spinal cord demyelination, remyelination, and atrophy were quantified from representative 1-microm-thick cross sections embedded in Araldite plastic using interactive image analysis. In addition, this study demonstrates novel, automated methodology to quantify axonal loss from areas of normal-appearing white matter, as a measure of secondary axonal injury following demyelination. These morphologic methods, which are applicable to various models of CNS injury, provide an innovative way to assess the benefits of therapeutic agents, to determine mechanisms of spinal cord damage, or to establish a correlation with sensitive measures of neurologic function. J. Neurosci Res 58:492-504. PMID- 10533043 TI - Metabotropic glutamate receptors modulate [(3)H]acetylcholine release from cultured amacrine-like neurons. AB - Retinal amacrine cells express metabotropic glutamate receptors (mGluRs), but their physiological role is unknown. We investigated the effect of mGluR on [(3)H]acetylcholine release ([(3)H]ACh) from cultured chick amacrine-like neurons. Activation of group III mGluR with the agonist L(+)-2-amino-4 phosphonobutyric acid (L-AP4) inhibited [(3)H]ACh release evoked by 25 mM KCl in a dose-dependent manner, and this effect was sensitive to pertussis toxin. In contrast, activation of group I or II mGluR with (S)-3, 5-dihydroxyphenylglycine (DHPG) and (2S,2'R,3'R)-2-(2', 3'-dicarboxycyclopropyl)glycine (DCG-IV), respectively, did not affect significantly [(3)H]ACh release. The effect of L-AP4 on [(3)H]ACh release was sensitive to nitrendipine, suggesting that it is, at least in part, due to inhibition of L-type Ca(2+) channels. Activation of group III mGluR also partly inhibited omega-conotoxin GVIA-sensitive Ca(2+) channels, coupled to [(3)H]ACh release. The L-AP4 did not affect the cAMP levels measured in amacrine-like neurons depolarized with 25 mM KCl or stimulated with forskolin, indicating that the effect of group III mGluR on [(3)H]ACh release is not due to inhibition of adenylyl cyclase activity. Inhibition of protein kinase A with KT 5720 was without effect on [(3)H]ACh release evoked by 25 mM KCl, further indicating that the effect of group III mGluR on [(3)H]ACh release cannot be attributed to the inhibition of the kinase. The effect of L-AP4 on [(3)H]ACh release was reversed by DHPG or by DCG-IV, and activation of group II mGluR also partially inhibited cAMP production stimulated by forskolin. Taken together, our results show that the effect of group III mGluR on [(3)H]ACh release may be due to a direct inhibition of L- and N-type Ca(2+) channels and is modulated by group I and group II mGluR. PMID- 10533044 TI - Enhanced sensitivity to N-methyl-D-aspartate receptor activation in transgenic and knockin mouse models of Huntington's disease. AB - We used two mouse models of Huntington's disease (HD) to examine changes in glutamate receptor sensitivity and striatal electrophysiology. One model, a transgenic, consisted of mice expressing exon 1 of the human HD gene and carrying 141-157 CAG repeat sequences (R6/2 line). The second model, a CAG repeat "knockin," consisted of mice with different lengths of CAG repeats (CAG71 and CAG94 repeats). The effects of glutamate receptor activation were examined by visualizing neurons in brain slices with infrared videomicroscopy and differential interference contrast optics to determine changes in somatic area (cell swelling). Striatal and cortical neurons in both models (R6/2 and CAG94) displayed more rapid and increased swelling to N-methyl-D-aspartate (NMDA) than those in controls. This effect was specific as there were no consistent group differences after exposure to alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) or kainate (KA). Intracellular recordings revealed that resting membrane potentials (RMPs) in the R6/2 transgenics were significantly more depolarized than those in their respective controls. RMPs in CAG94 mice also were more depolarized than those in CAG71 mice or their controls in a subset of striatal neurons. Confirming previous results, R6/2 mice expressed behavioral abnormalities and nuclear inclusions. However, CAG71 and CAG94 knockins did not, suggesting that increased sensitivity to NMDA may occur early in the disease process. These findings imply that NMDA antagonists or compounds that alter sensitivity of NMDA receptors may be useful in the treatment of HD. PMID- 10533045 TI - mGluR3 and mGluR5 are the predominant metabotropic glutamate receptor mRNAs expressed in hippocampal astrocytes acutely isolated from young rats. AB - The metabotropic glutamate receptors (mGluRs) that are expressed and not expressed on astrocytes in the brain have not been defined. While immunohistochemistry and in situ mRNA hybridization have been used on a limited basis to address this question, they do not readily enable the proportion of astrocytes expressing a particular mRNA or protein to be determined. Also, for many receptors, expression by cultured astrocytes does not reflect in situ expression. In this study, therefore, we examined expression of mRNA for all the mGluRs except mGluR6 by single-cell reverse transcriptase-polymerase chain reaction (RT-PCR) in freshly isolated hippocampal astrocytes from postnatal day P1-10 rats, as an additional approach to address the question of which mGluRs are expressed on astrocytes in situ. The astrocytic nature of the cells was supported by simultaneously measuring mRNA for the astrocytic marker glial fibrillary acidic protein (GFAP) from the same cells. In these studies, the percentage of cells showing GFAP mRNA expression was the same as the percentage of cells showing immunocytochemical staining for GFAP. We found that only mGluR3 and mGluR5 mRNAs were significantly present in GFAP mRNA(+) cells. The mGluR5 PCR products were primarily of the "a" splice variant. mGluR1, 2, 4, 7, and 8 were very rarely or never detected. mGluR6 mRNA level was too low in whole rat brain and hippocampus to warrant examination. These results show that interpretation of effects involving mGluR3 or 5 activation in the hippocampus of young rats needs to also consider effects due to astrocytes. PMID- 10533046 TI - Pharmacological and molecular evidence for dopamine D(1) receptor expression by striatal astrocytes in culture. AB - The neurotransmitter dopamine (DA) at a 10 microM concentration elicited a stimulation of intracellular cyclic AMP (cAMP) accumulation in cultured astrocytes derived from embryonic rat striatum. This accumulation was partially blocked by the beta-adrenergic receptors antagonist propranolol, mimicked by the D(1) agonist SKF 38393 and by the mixed D(1)/D(2) agonist apomorphine. A regional heterogeneity in the magnitude of dopamine-induced cAMP accumulation was observed in cultured astrocytes obtained from different brain areas. The maximum effect was observed in striatal astrocytes, a lower effect in cortical astrocytes, and no increase was detected in cerebellar astrocytes. Reverse transcription polymerase chain reaction (RT-PCR) coupled to Southern blot hybridization demonstrated that striatal astrocytes express only D(1) receptor mRNA and Western blot analysis confirmed the expression of the D(1) receptor protein in striatal astrocytes. In contrast to what found in neurons, the D(1)-dependent cAMP formation in striatal astrocytes is partially reduced by pertussis toxin (PTX) treatment. The stimulation of D(1) receptors or the activation of adenylyl cyclase by forskolin led to an increase of cytosolic and nuclear protein kinase A (PKA) catalytic activity. The presence of dopamine D(1) receptors in cultured striatal astrocytes suggests a role of dopamine in the regulation of cellular processes in striatal astrocytes. PMID- 10533047 TI - Anatomical and functional reconstruction of the nigrostriatal system in vitro: selective innervation of the striatum by dopaminergic neurons. AB - To study development of the nigrostriatal pathway in an in vitro model system, organotypic slices obtained from rat pups (P4) and containing the striatum and the cortex were grown together with apposed embryonic (E13.5) mesencephalic blocks according to the static slice culture method of Stoppini et al. (1991; J. Neurosci. Methods 37:173-182). Under these conditions, mesencephalic dopaminergic (DA) fibers rapidly grow through the slice, preferentially its striatal portion. This innervation provides a true synaptic innervation to the striatum, as shown by the presence of DA terminals on striatal neurons. DA fibers are able to exert a functional influence, as seen by their ability to modulate c-Fos expression in striatal neurons in the same way as in vivo. Thus, blockade, under basal conditions, of the effect of spontaneously released dopamine by the D2 receptor antagonist haloperidol leads to the activation of c-Fos expression in the striatum. Furthermore, stimulation of DA release by amphetamine induces striatal c-Fos expression in a D1 receptor-dependent manner. Next, the mechanisms of the selective striatal innervation were examined. Indeed, DA fibers innervated specifically the striatum, avoiding the cortical portion of the slice. This selectivity seems to be specific for DA neurons; no selectivity could be observed when noradrenergic neurons were substituted for DA neurons. Short-term cocultures in a collagen gel of mesencephalic blocks with striatal blocks failed to reveal any oriented outgrowth of DA fibers from the mesencephalon, suggesting that the selective innervation observed in the organotypic slices results from some contact-dependent, presumably adhesive interactions rather than from the presence of some diffusible substance orienting the growth of DA fibers towards the striatum. On the other hand, DA neurons seeded onto striatal slices did not attach selectively onto the striatal portion of the slice, indicating that the putative specific adhesive interactions governing the selective striatal innervation are not the same as those determining the adhesion of the DA neurons. These results show that cocultures of cortex-striatum and mesencephalic slices result in a system that displays a number of the morphological and functional traits of the normal nigrostriatal system and that can be relied on as a good in vitro model of in vivo development. PMID- 10533048 TI - KIAA0369, doublecortin-like kinase, is expressed during brain development. AB - During embryonic development, the cerebral cortex attains its characteristic adult laminated structure. The finding that X-linked lissencephaly patients harbor mutations in the doublecortin gene implicated this gene product in the process of corticogenesis. An autosomal human gene, KIAA0369, with a high level of similarity to doublecortin, has been cloned from human adult brain. This gene product contains a kinase domain in addition to a doublecortin-like domain. In order to evaluate whether this doublecortin-like kinase also plays a role during brain development, we cloned and studied the expression pattern of the mouse homolog. Three cDNA products of this gene were cloned: one, doublecortin-like kinase, the second containing only the doublecortin-like region, and the third containing only the kinase domain, a homolog of the previously cloned rat CPG16 gene. We studied doublecortin-like kinase expression in mouse using Northern blot analysis, in situ hybridization, and Western blot analysis, and conclude that doublecortin-like kinase is expressed in multiple regions of embryonic brain including the developing cerebral cortex. PMID- 10533049 TI - Oxidative stress-induced metabolic alterations in rat brain astrocytes studied by multinuclear NMR spectroscopy. AB - Oxidative stress in cultured astrocytes exerted by 30-min treatment with 50-200 microM H(2)O(2) caused time- and concentration-dependent effects on cellular metabolism. These changes were accompanied by alterations in cellular morphology. Using (31)P nuclear magnetic resonance (NMR) spectroscopy, the data demonstrate that the energy status of the cells was greatly affected directly after the stress, as indicated by the loss of high energy phosphates, i.e., phosphocreatine (PCr) and nucleoside triphosphates (NTP). Oxidative stress also involves a dysregulation of the osmotic control in astrocytes, which is accompanied by a dramatic loss of myo-inositol, taurine, and hypotaurine, as monitored by (1)H and (13)C NMR spectroscopy. While the energy state of the cells was essentially restored during a 7-hr recovery period, the changes in osmolyte concentrations lasted longer and went on throughout the recovery period. Even after 24-hr recovery, organic osmolyte concentrations were still below the control levels. (13)C NMR spectra of astrocyte cell extracts also demonstrated an enhanced glucose metabolism via the pentose phosphate pathway (PPP) and a reduced glycolysis. Additionally, the appearance of (13)C glutamate points to a distortion of glutamine synthetase (GS), leading to the accumulation of glutamate. Glycolysis as well as GS activity were back to control levels after 7 hr recovery. Thus, in contrast to the energy metabolism, osmoregulatory processes and complex glucose metabolism was impaired not only directly after oxidative stress, but occurred with a later onset during a 2-hr recovery period, and cells only slowly recovered during the next 24 hr. PMID- 10533050 TI - Distinct alterations in phospholipid metabolism in brains of apolipoprotein E deficient mice. AB - Apolipoprotein E (ApoE) is the major brain lipoprotein and plays an important role in lipid transport. ApoE-deficient mice whose apoE gene has been knocked out have distinct cognitive and neurochemical deficits, and their recovery from brain injury is impaired. In the present study we examined the possibility that the neuronal derangements of apoE-deficient mice are related to impairments in their phospholipid metabolism. This was performed by comparison of the phospholipid, fatty acid, and cholesterol compositions of distinct membranal brain fractions of apoE-deficient and control mice. Analysis of the microsomal membrane fraction P(3) revealed that, in apoE-deficient mice, these membranes contain significantly lower levels of phosphatidylcholine (PC) than those of control mice. This effect was specific to PC and thus resulted in a twofold decrease of the PC to phosphatidylethanolamine (PE) ratio in apoE-deficient mice compared to the corresponding control ratio. In contrast, the cholesterol levels of the microsomal membranes of the two mice were similar, and the fatty acid composition of their PC was unchanged. There were, however, changes in the fatty acid composition of PE and phosphatidylserine (PS), which resulted in a lower ratio of polyunsaturated to saturated fatty acids in PE and in a higher ratio in apoE deficient mice compared to the corresponding control values. These effects were specific to the microsomal fraction P(3) and were not observed with the brain subcellular membrane fraction P(2), which is composed mainly of plasma and mitochondrial membranes and whose phospholipid, fatty acid, and cholesterol levels were similar in apoE-deficient and control mice. These findings show that apoE deficiency results in specific and intracellular compartment-dependent changes in phospholipid metabolism, which may play an important role in mediating the neuronal effects of apoE. PMID- 10533051 TI - Interleukin-1 beta activates expression of cyclooxygenase-2 and inducible nitric oxide synthase in primary hippocampal neuronal culture: platelet-activating factor as a preferential mediator of cyclooxygenase-2 expression. AB - Interleukin-1 beta (IL-1beta) is an inflammatory cytokine whose expression is elevated in brain during seizures, ischemia, and injury. Expression of IL-1beta and its receptor can also be observed in normal brain. Platelet-activating factor (PAF) is also a dual mediator that promotes neuronal plasticity responses as well as inflammation. We have determined the role of PAF in the regulation of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) genes by IL 1beta in rat primary hippocampal cultures. As assessed by reverse transcriptase/polymerase chain reaction (RT/PCR), recombinant mouse IL-1beta (1 nM) led to an induction of COX-2 mRNA which peaked at 2 hours, declined to baseline levels by 4 hours, began to rise again by 6 hours, and remained elevated at 24 hours post-treatment. iNOS mRNA was also induced, but unlike COX-2, its abundance peaked at 4 hours and decreased by 6 hours to a plateau lasting through 24 hours. Pretreatment with PAF antagonist BN50730 blocked induction of COX-2 mRNA by 2-hour IL-1beta treatment, and 2-hour treatment with the PAF analog mcPAF mimicked the effects of IL-1beta on COX-2 mRNA levels. Following injury, synaptic plasticity changes may be affected by IL-1beta-PAF-COX-2 neuronal signaling. PMID- 10533052 TI - Noradrenaline effects on pyruvate decarboxylation: correlation with calcium signaling. AB - Noradrenaline effects on the rate of metabolism of pyruvate to acetyl coenzyme A, catalyzed by the pyruvate dehydrogenase complex, was measured in primary cultures of mouse astrocytes as rate of production of labeled CO(2) from 1-[(14) C]pyruvate in the absence of competing glucose in the medium. The subtype specificity of a noradrenaline-stimulated increase in rate of CO(2) formation was identical to that for noradrenaline-induced increase in free intracellular calcium ([Ca(2+)](i)), suggesting a causal relationship between these two phenomena. The noradrenaline-induced stimulation of pyruvate decarboxylation was abolished in the presence of 10 mM magnesium chloride in the medium, combined with the omission of calcium, a procedure known to prevent an increased [Ca(2+)] in the cytosol from raising intramitochondrial [Ca(2+)]. Thus, the stimulation of metabolic flux through the reaction catalyzed by the pyruvate dehydrogenase complex appears to result from an increase in intramitochondrial [Ca(2+)] ions in astrocytes. Such a mechanism for stimulation of the same enzyme has been convincingly demonstrated in other cell types, primarily heart muscle and hepatic cells, but it has not previously been demonstrated in any cell type from the central nervous system. PMID- 10533053 TI - Astrocytes prevent neuronal death induced by reactive oxygen and nitrogen species. AB - Reactive oxygen and nitrogen species (RO/NS) such as nitric oxide (NO), hydroxyl radical (OH.), and superoxide anion (O(2)(-)) are generated in a variety of neuropathological processes and damage neurons. In the present study, we investigated the neuroprotective effects of rat astrocytes against RO/NS-induced damage using neuron-glia cocultures, and the effects were compared to those of microglial cells. Sodium nitroprusside (SNP), 3-morpholinosydnonimine (SIN-1), and FeSO(4) were used to generate NO, O(2)(-) and NO, and OH., respectively. Solely cultured neurons, which were transiently exposed to these agents, degenerated, possibly through apoptotic mechanisms as revealed by in situ detection of DNA fragmentation, whereas neurons cocultured with either astrocytes or microglial cells were viable even after exposure to RO/NS. In contrast, most neurons cocultured with meningeal fibroblasts degenerated. Astrocyte-conditioned medium partially attenuated RO/NS-induced neuronal damage. When neurons were cultured on astrocyte-derived extracellular matrix (AsECM), neuronal death induced by SNP and FeSO(4) was almost completely inhibited. AsECM contained significant amounts of laminin and fibronectin, and pure fibronectin and laminin also protected neurons against RO/NS-induced damage in the same manner as AsECM. These results suggest that astrocytes can protect neurons against RO/NS-induced damage by secreting soluble and insoluble factors. PMID- 10533054 TI - Intercellular Ca(2+) waves induce temporally and spatially distinct intracellular Ca(2+) oscillations in glia. AB - Mechanically induced intercellular Ca(2+) waves propagated for approximately 300 microm in primary glial cultures. Following the wave propagation, 34% of the cells displayed Ca(2+) oscillations in a zone 60-120 microm from the stimulated cell. The initiation, frequency, and duration of these Ca(2+) oscillations were dependent on the cells' distance from the wave origin but were not dependent on the cell type nor on the magnitude of the Ca(2+) wave. When an individual cell propagated two sequential intercellular Ca(2+) waves originating from different sites, the characteristics of the Ca(2+) oscillations initiated by each wave were determined by the distance of the cell from the origin of each wave. Each Ca(2+) oscillation commonly occurred as an intracellular Ca(2+) wave that was initiated from a specific site within the cell. The position of the initiation site and the direction of the intracellular Ca(2+) wave were independent of the orientation of the initial intercellular Ca(2+) wave. Because initiation and frequency of Ca(2+) oscillations are dependent on the intracellular inositol trisphosphate concentration ([IP(3)](i)), we propose that the zone of cells displaying Ca(2+) oscillations is determined by an intercellular gradient of [IP(3)](i), established by the diffusion of IP(3) through gap junctions during the propagation of the intercellular Ca(2+) wave. Exposure to acetylcholine, a muscarinic agonist that initiates IP(3) production, shifted the zone of oscillating cells about 45 microm farther away from the origin of the mechanically induced wave. These findings indicate that a glial syncytium can resolve information provided by a local Ca(2+) wave into a distinct spatial and temporal pattern of Ca(2+) oscillations. PMID- 10533056 TI - Monoclonal antibody detects oligodendroglial cell surface protein exhibiting temporal regulation during development. AB - As tools to study stage-specific surface molecules expressed during the development of oligodendrocytes, we have generated monoclonal antibodies against peanut agglutinin (PNA)-binding glycoproteins isolated by affinity chromatography from the oligodendroglial precursor cell line Oli-neu. In this paper we report the characterization of the monoclonal antibody 7D10. The 7D10 antibody recognizes a 145-kD cell surface glycoprotein expressed by postmitotic multibranched cells of the oligodendroglial lineage. The antigen stains subpopulations of myelin-associated glycoprotein (MAG) and O4-positive cells and is subsequently down-regulated during further differentiation in vitro. The 7D10 antigen is also expressed by a subpopulation of astroglial cells but not by neurons. A truncated form of the protein is released by antigen-expressing cells into the culture supernatant. PMID- 10533055 TI - Neuronal death in cytokine-activated primary human brain cell culture: role of tumor necrosis factor-alpha. AB - We examined cytokine-mediated neuronal death in neuron-astrocyte cultures from second trimester human fetal cerebrum. In these cultures, high-output inducible nitric oxide synthase (NOS) and tumor necrosis factor-alpha (TNFalpha) are expressed in astrocytes after exposure to IL-1beta/IFNgamma. Neuronal cell death was evident at >/=48 h following cytokine stimulation. Neutralizing anti-TNFalpha antiserum inhibited ( approximately 48%) neurotoxicity in IL-1beta/IFNgamma treated cultures, demonstrating a role for endogenously produced TNFalpha. Interestingly, the degree of neuroprotection conferred by superoxide dismutase or N-methyl D-aspartate (NMDA) receptor antagonists in these cultures was smaller and variable. Similarly, the effect of the NOS inhibitor, N(G)-monomethyl L arginine (NMMA) on IL-1beta/IFNgamma-induced neuronal death was variable, showing no statistically significant effect when results from more than 30 independent cultures were averaged. Neurons die by apoptosis in cytokine-treated human fetal CNS cultures as shown by the characteristic nuclear morphology as well as positive labeling for TUNEL. Our results demonstrate a potent neurotoxicity mediated by the cytokine combination IL-1beta/IFNgamma in primary human neuron astrocyte cultures and a crucial role for endogenous TNFalpha in mediating neurotoxicity in this system. These results firmly establish the neurotoxic potential of the inflammatory cytokines IL-1beta and TNFalpha in the human CNS. PMID- 10533057 TI - Suppression of nidogen-1 translation by antisense targeting affects the adhesive properties of cultured astrocytes. AB - The multidomain glycoprotein nidogen-1 is a common component of basal membranes. Nidogen-1 is produced by the endothelial cells and the mesenchymal cells of the developing central nervous system. Recent results give evidence that nidogen-1 may also be secreted by cultured Schwann cells to basement membranes of peripheral nerves. We were interested in ascertaining whether astrocytes, which have the capacity to produce laminin and fibronectin and are an important source of extracellular matrix (ECM) molecule secretion in the brain, might also produce nidogen-1. Immunocytochemistry, in combination with polymerase chain reaction and in situ hybridization techniques, revealed that astrocytes in culture synthesize nidogen-1. To show the functional significance of the nidogen-1 secretion by astrocytes, antisense targeting techniques were applied. These experiments showed that nidogen-1 may be an essential modulator of astrocytic adhesion to the substrate. The suppression of nidogen-1 synthesis by the application of antisense oligonucleotides induced a morphological transition from a flat, polygonal to a round cell and was accompanied by the detachment of the astrocytes from the substrate. Hence, nidogen-1 might be an important component of the ECM secreted by astrocytes. The suppression of nidogen-1 synthesis may disturb the aggregation of ECM molecules to a functional basement membrane and thus reduce the astrocytic adhesion to the substrate. Nidogen-1 secretion to basement membranes by astrocytes may have important functional implications during blood-brain barrier and scar formation. PMID- 10533058 TI - Plasma membrane calcium ATPase isoforms in astrocytes. AB - The plasma membrane Ca(2+)-ATPase (PMCA) is an essential component of the machinery responsible for cellular Ca(2+) homeostasis. Together with the Na(+)/Ca(2+) exchanger, the plasma membrane Ca(2+)-ATPase (PMCA) is responsible for the extrusion of Ca(2+) from the cytosol. Although both PMCAs and Na(+)/Ca(2+) exchangers are present in high amounts in the brain, it is thought that only the latter localize to glia. This study investigates whether PMCAs are also present in astrocytes and thus are components of Ca(2+) signalling in this cell type. Membrane proteins and mRNA were isolated from primary cultures of rat cortical astrocytes and C6 glioma cells. PMCA isoforms were investigated with isoform specific antibodies and the splice variant pattern was studied in RT-PCR experiments using specific oligonucleotides. The PMCA1, 2, and 4 isoforms were detected in rat cortical astrocytes, whereas only PMCA1 and 2 were found in C6 cells. While neurons express both the CI and CII splice variants, only the splice variant CI of PMCA1, 2, and 4 was detected in astrocytes. Thus, the PMCA pump is present in mammalian glial cells. These results also show that the amounts of PMCA1 and 4 isoforms in astrocytes are comparable to those found in neurons. In contrast, astrocytes contain smaller amounts of PMCA2. Furthermore, PMCA2 and PMCA4 underwent an evident time dependent up-regulation in astrocytes cultured in vitro. PMID- 10533059 TI - Effects of schwann cell suspension grafts on axon regeneration in subacute and chronic CNS traumatic injuries. AB - In a previous study, we have shown that microtransplanted Schwann cell suspensions foster structural recovery of the acutely transected postcommissural fornix. The emphasis of the present study was to examine whether subacutely and chronically injured axons also demonstrate significant responsiveness to implanted Schwann cells. Microinjected suspensions of cultured Schwann cells i) elicited a growth response and attracted axons in a subacute and chronic traumatic lesion but ii) failed to stimulate regrowth of the postcommissural fornix projection at any nonacute postlesion stage. In conclusion, the single intervention strategy of Schwann cell microimplantation is not sufficient to ensure regeneration of the subacutally or chronically transected postcommissural fornix. The use of Schwann cells as stimulators of axon regrowth depends on the neuronal cell type and the appropriate postinjury time point. PMID- 10533060 TI - Lesion-induced changes of electrophysiological properties in astrocytes of the rat dentate gyrus. AB - Reorganization of the adult dentate gyrus following unilateral entorhinal cortex lesion (ECL) is a well-established model for studying mechanisms of trauma induced neuronal plasticity. The lesion induces deafferentiation of the outer molecular layer, which is accompanied by a strong astroglial reaction. This glial response is thought to contribute to subsequent repair processes, but the underlying mechanisms are poorly understood. In this study we addressed the question whether denervation leads to modifications in the electrophysiological properties of astrocytes, assuming that such changes might be involved in the remodeling of neural circuitry. Patch-clamp recordings were obtained from astrocytes in the dentate gyrus of adult rats that underwent ECL and compared to corresponding data from control animals. We observed a significant reduction of inward rectifier K(+) current densities, a positive shift of resting potentials, and an increase in input resistance in astrocytes of the denervated molecular layer. Current densities were reduced between 6 and 19 days postlesion (dpl), reaching a minimum at 10 dpl. Voltage-gated outward K(+) currents were not affected by the lesion. Inward rectifier K(+) currents increase with maturation in astrocytes. Thus, our results provide evidence that, following ECL, mature astrocytes dedifferentiated and readapted an immature current pattern. Presumably, these changes lead to stronger and prolonged depolarization of glial cells and neurons in response to activity-dependent K(+) release, which in turn might enhance the synthesis of neurotrophic factors and contribute to a permissive environment for neuronal reorganization. PMID- 10533061 TI - Point mutations in the dystrophin gene: evidence for frequent use of cryptic splice sites as a result of splicing defects. AB - Ten different mutations have been identified in patients with Becker (n = 1) or Duchenne (n = 9) muscular dystrophy using reverse transcription of total RNA, polymerase chain reaction amplification of the whole coding region of the gene and protein truncation test (PTT) analysis. Seven mutations had not been reported previously, and these consist in three nonsense mutations (Q2522X, E2726X, R3381X), three frameshifting deletions (3686-3687delGT, 5126delA, 5759delC), and four splicing defects of which the effects on the muscle dystrophin mRNA transcripts have been analyzed. In one case, a 3' splice-site mutation (IVS74-2A- >G) resulted in a complex pattern of exon skipping involving exons of the C terminal domain. In the three other cases, nucleotide substitutions in splice donor (IVS26+2T-->A, IVS65+1G-->A) or acceptor (IVS8-15A-->G) recognition sequences led to the use of cryptic splice sites, with consequent insertions of intronic sequences in the processed mRNA. Up to 34% (70/203) of the point mutations reported to date in the dystrophin database (http://www.dmd.nl) affect splice sites of the dystrophin gene. However, altered mRNA splicing has been confirmed experimentally in only 23% of cases (16/70). Combined with PTT, the transcript analysis protocol defined in this study permits direct determination of the impact of intronic variations on the structure of dystrophin mRNA and of the resulting consequences on the translational reading frame. We present evidence for a frequent use of cryptic splice sites as a result of splicing defects. PMID- 10533062 TI - Homonucleotide expansion and contraction mutations of PAX2 and inclusion of Chiari 1 malformation as part of renal-coloboma syndrome. AB - Renal-Coloboma syndrome, an autosomal dominant disorder characterized by colobomatous eye defects, vesicoureteral reflux, and abnormal kidneys, results from mutations in PAX2. The purpose of this study was to identify mutations in PAX2 and understand the associated patient phenotypes. We report a severely affected girl and a mildly affected mother and daughter, all of whom have PAX2 homoguanine tract (7 G) missense mutations. The mother and daughter have optic nerve colobomas and the daughter has vesicoureteral reflux. The severely affected girl developed renal failure and has bilateral colobomatous eye defects. Additionally, this girl developed hydrocephalus associated with platybasia and a Chiari 1 malformation. We examined genomic DNA from these individuals by SSCP and sequencing. The mother and daughter had a novel mutation: a contraction in a string of 7 G's to 6 G's in one allele of PAX2, leading to a premature stop codon two amino acids downstream. The severely affected girl had an expansion to 8 G's, leading to a premature stop codon 27 amino acids downstream. The 8 G expansion has been found in other patients without brain anomalies and has occurred spontaneously in a mouse model, PAX2(1Neu). We expand the known phenotype associated with mutations in PAX2 to include brain malformations. The homoguanine tract in PAX2 is a hot spot for spontaneous expansion or contraction mutations and demonstrates the importance of homonucleotide tract mutations in human malformation syndromes. PMID- 10533063 TI - Townes-Brocks syndrome: detection of a SALL1 mutation hot spot and evidence for a position effect in one patient. AB - Townes-Brocks syndrome (TBS) is an autosomal dominant developmental disorder characterized by anal and thumb malformations and by ear anomalies that can affect the three compartments and usually lead to hearing loss. The gene underlying TBS, SALL1, is a human homolog of the Drosophila spalt gene which encodes a transcription factor. A search for SALL1 mutations undertaken in 11 unrelated affected individuals (five familial and six sporadic cases) led to the detection of mutations in nine of them. One nonsense and six different novel frameshift mutations, all located in the second exon, were identified. Together with the previously reported mutations [Kohlhase et al., 1999], they establish that TBS results from haploinsufficiency. The finding of de novo mutations in the sporadic cases is consistent with the proposed complete penetrance of the disease. Moreover, the occurrence of the same 826C>T transition in a CG dimer, in three sporadic cases from the present series and three sporadic cases from the other series [Kohlhase et al., 1999] (i.e., six of the eight mutations identified in sporadic cases), reveals the existence of a mutation hotspot. Six different SALL1 polymorphisms were identified in the course of the present study, three of which are clustered in a particular region of the gene that encodes a stretch of serine residues. Finally, the chromosome 16 breakpoint of a t(5;16)(p15.3;q12.1) translocation carried by a TBS-affected individual was mapped at least 180 kb telomeric to SALL1, thus indicating that a position effect underlies the disease in this individual. PMID- 10533064 TI - Molecular studies in 20 submicroscopic neurofibromatosis type 1 gene deletions. AB - Neurofibromatosis type 1 (NF1) is a common autosomal dominant disorder characterized by a marked variability in expression. A more severe phenotype is frequently observed in the group of patients carrying a large NF1 deletion. To study the extent of the microdeletion in these NF1 patients, we generated a partial physical map of the NF1 flanking region. We describe seven PACs and three new polymorphic dinucleotide repeats located outside the NF1 gene and analyzed 20 unrelated individuals with an NF1 microdeletion in a collaborative study. We detected one individual with a substantially smaller deletion including only the NF1 gene and its three embedded genes. In the other 19 patients, the deletion extended at least 1 Mb. The parental origin of the deletion was determined in 15 individuals and was maternal in 13 and paternal in two cases. The new molecular tools described here can be used to unequivocally diagnose a possible extragenic extension of an NF1 deletion. PMID- 10533066 TI - Mutation screening of the entire coding regions of the TSC1 and the TSC2 gene with the protein truncation test (PTT) identifies frequent splicing defects. AB - Mutation analyses in tuberous sclerosis (TSC) have reported a wide variety of disease-causing aberrations in the two known predisposing genes, TSC1 and TSC2 on chromosomes 9q34 and 16p13, comprising mainly small mutations distributed over the entire genes. So far, all known TSC1 mutations as well as the majority of TSC2 mutations truncate the proteins hamartin and tuberin, respectively. We describe for the first time an RNA-based screening of the entire coding regions of both TSC genes for truncating mutations applying the protein truncation test (PTT). Simultaneous investigation of both TSC genes in a group of 48 unassigned TSC patients, which were previously tested to exclude large intragenic TSC2 rearrangements, revealed aberrant migrating polypeptides resulting from truncating mutations in nine TSC1 cases and in 16 TSC2 cases while three TSC2 cases showed enlarged proteins. TSC1 mutations include two nonsense mutations, four insertions, and three splice mutations. Nineteen mutations identified in TSC2 were composed of four different nonsense mutations in five patients, one deletion, one insertion, and seven different splicing aberrations due to at least eight different mutations found in 12 patients. Additional predicted truncating mutations according to PTT without possible identification of the causative alteration allowed assignment to TSC1 in one and TSC2 in seven cases. Twelve patients without abnormalities in the PTT are assumed to harbor missense mutations, probably in TSC2. The high proportion of TSC2 splicing aberrations strengthens the importance of intronic disease-causing mutations and the application of RNA-based screening methods to confirm their consequences. PMID- 10533065 TI - Jagged-1 mutation analysis in Italian Alagille syndrome patients. AB - Alagille syndrome (AGS) is an autosomal dominant disorder with developmental abnormalities affecting the liver, heart, eyes, vertebrae, and craniofacial region. The Jagged-1 (JAG1) gene, which encodes a ligand of Notch, has recently been found mutated in AGS. In this study, mutation analysis of the JAG1 gene performed on 20 Italian AGS patients led to the identification of 15 different JAG1 mutations, including a large deletion of the 20p12 region, six frameshift, three nonsense, three splice-site, and two missense mutations. The two novel missense mutations were clustered in the 5' region, while the remaining mutations were scattered throughout the gene. The spectrum of mutations in Italian patients was similar to that previously reported. We also studied in detail a complex splice site mutation, 3332dupl8bp, which was shown to lead to an abnormal JAG1 mRNA, resulting in a premature stop codon. With the exception of the missense mutations, the majority of the JAG1 mutations are therefore likely to produce truncated proteins. Since the phenotype of the patient with a complete deletion of the JAG1 gene is indistinguishable from that of patients with intragenic mutations, our study further supports the hypothesis that haploinsufficiency is the most common mechanism involved in AGS pathogenesis. Furthermore, our data confirmed the absence of a correlation between the genotype of the JAG1 gene and the AGS phenotype. PMID- 10533067 TI - Analysis of both TSC1 and TSC2 for germline mutations in 126 unrelated patients with tuberous sclerosis. AB - Tuberous sclerosis complex (TSC) is an autosomal dominant disorder characterized by the development of multiple hamartomas involving many organs. About two-thirds of the cases are sporadic and appear to represent new mutations. With the cloning of two causative genes, TSC1 and TSC2 it is now possible to analyze both genes in TSC patients and identify germline mutations. Here we report the mutational analysis of the entire coding region of both TSC1 and TSC2 genes in 126 unrelated TSC patients, including 40 familial and 86 sporadic cases, by single-stranded conformational polymorphism (SSCP) analysis followed by direct sequencing. Mutations were identified in a total of 74 (59%) cases, including 16 TSC1 mutations (5 sporadic and 11 familial cases) and 58 TSC2 mutations (42 sporadic and 16 familial cases). Overall, significantly more TSC2 mutations were found in our population, with a relatively equal distribution of mutations between TSC1 and TSC2 among the familial cases, but a marked underrepresentation of TSC1 mutations among the sporadic cases (P = 0.0035, Fisher's exact test). All TSC1 mutations were predicted to be protein truncating. However, in TSC2 13 missense mutations were found, five clustering in the GAP-related domain and three others occurring in exon 16. Upon comparison of clinical manifestations, including the incidence of intellectual disability, we could not find any observable differences between TSC1 and TSC2 patients. Our data help define the distribution and spectrum of mutations associated with the TSC loci and will be useful for both understanding the function of these genes as well as genetic counseling in patients with the disease. PMID- 10533068 TI - Novel mutations in XLRS1 causing retinoschisis, including first evidence of putative leader sequence change. AB - Juvenile retinoschisis is an X-linked recessive disease caused by mutations in the XLRS1 gene. We screened 31 new unrelated patients and families for XLRS1 mutations in addition to previously reported mutations for 60 of our families (Retinoschisis Consortium, Hum Mol Genet 1998;7:1185-1192). Twenty-three different mutations including 12 novel ones were identified in 28 patients. Mutations identified in this study include 19 missense mutations, two nonsense mutations, one intragenic deletion, four microdeletions, one insertion, and one intronic sequence substitution that is likely to result in a splice site defect. Two novel mutations, c.38T-->C (L13P) and c.667T-->C (C223R), respectively, present the first genetic evidence for the functional significance of the putative leader peptide sequence and for the functional significance at the carboxyl terminal of the XLRS1 protein beyond the discoidin domain. Mutations in 25 of the families were localized to exons 4-6, emphasizing the critical functional significance of the discoidin domain of the XLRS1 protein. PMID- 10533069 TI - Protein truncation test for screening hamartin gene mutations and report of new disease-causing mutations. AB - Considering the prevalence of truncating mutations in the tuberous sclerosis (TSC) hamartin gene (TSC1), we devised a protein truncation test (PTT) to analyze the full length coding sequence of TSC1. Studying 12 sporadic cases and three familial forms by a combination of PTT and single-strand conformation polymorphism analysis (SSCA), we found 5/15 mutations while PTT alone detected 4/15 truncating mutations, two of which escaped SSCA analysis. SSCA alone picked up one missense mutation and two mutations also detected by PTT. Interestingly, a TSC1 mutation was identified in all three familial forms (3/3) while the rate of mutation detection was lower in sporadic cases (2/12). In conclusion, PTT proves to be a useful technique for the rapid detection of disease-causing mutations in the TSC1 gene. PMID- 10533070 TI - DGGE method for the mutational analysis of the coding and proximal promoter regions of the Alzheimer's disease presenilin-1 gene: two novel mutations. AB - Many different mutations that cause Alzheimer's disease (AD) have been found in the presenilin-1 gene (PSEN1) and are associated with the most aggressive forms of the disease. With the aim of screening for PSEN1 genetic variations, we developed a method based on denaturing gradient gel electrophoresis (DGGE) that allows the mutational analysis of all the coding exons and the proximal promoter of PSEN1 using only four DGGE gels. The analysis by this methodology of a sample of 58 early-onset AD (EOAD) patients nonselected for family history resulted in finding four genetic variants within the PSEN1 coding region, two of which are novel mutations (M233L and A409T), whereas the other two have been reported previously (L282R and E318G). We also found a novel mutation within the PSEN1 proximal promoter (-280 C-->G) that, interestingly, provokes significant changes in the transcriptional activity of the gene in cell lines of neuronal and astrocytic, but not hepatic origin. These data strongly suggest that the region around -280 of PSEN1 promoter contains a regulatory element that controls its transcription specifically in neural cells. PMID- 10533071 TI - Comparison of heteroduplex analysis, direct sequencing, and enzyme mismatch cleavage for detecting mutations in a large gene, FBN1. AB - Analysis of large genes for mutations of clinical relevance is complicated by intragenic heterogeneity, sensitivity, and cost of the methods available, and in the case of many conditions, specificity of the genetic alterations detected. We examined the FBN1 gene for mutations in people who had Marfan syndrome using three methods: single-chain polymorphism analysis (SSCP) with heteroduplex (HA) analysis, enzyme-mediated cleavage (EMC) of heteroduplexes, and direct sequencing. We also used these methods to search for mutations in the P53 gene in patients with hepatocellular carcinoma. The results showed that EMC was most efficient for detecting mutations. However, the cost favored SSCP with heteroduplex analysis, provided conditions did not need to be optimized to detect a mutation. Until more cost-effective and sensitive methods are developed to detect unknown mutations in large genes, diagnosis of many genetic disorders will depend on the willingness of an investigator who is studying a particular disorder to perform clinical molecular testing and have the laboratory accredited. PMID- 10533072 TI - Metachromatic leucodystrophy: a newly identified mutation in arylsulphatase A, D281Y, found as a compound heterozygote with I179L in an adult onset case. AB - The majority of mutations identified in patients with Metachromatic leucodystrophy are unique to individual families. We report here a new mutation in the arylsulphatase A gene (D281Y) identified in a patient with late-onset Metachromatic leucodystrophy. This mutation was inherited in cis with the common pseudo-deficiency allele and in trans with the previously described I179S (250100.0008) mutation which complicated the enzymatic diagnosis of this condition. Sequence comparison shows D281 to be highly conserved amongst the arylsulphatases. The clinical features of this patient which are predominantly of a slowly progressive psychiatric and intellectual deterioration rather than rapid neurological impairment are typical of I179S compound heterozygotes. PMID- 10533073 TI - A novel complex mutation of the OTC (ornithine transcarbamylase) gene in a Malaysian pedigree. PMID- 10533074 TI - Two novel mutations in the MPZ gene coding region in Charcot-Marie-Tooth type 1 patients of Turkish origin: S54P, [I30del; GVYI29ins]. PMID- 10533075 TI - Identification of three novel mutations in the major human skeletal muscle chloride channel gene (CLCN1), causing myotonia congenita. AB - Myotonia congenita (MC) is a genetic disease characterized by mutations in the CLCN1 gene (OMIM*118425) encoding the skeletal muscle voltage-gated chloride channel (ClC-1). Autosomal dominant and recessive forms are observed, characterized by impaired muscle relaxation after forceful contraction (myotonia), which is more pronounced after inactivity and improves with exercise. We report three novel and one known mutations of the CLCN1 gene in four unrelated MC families. In two families the mutations were missense: 803C>T (T268M) and 1272C>G (I424M) in exons 7 and 12, respectively. The third was a splice mutation in intron 5 (696+2T>A), which induced a frame shift with a stop codon in exon 6 (fs213X). In the fourth family the previously-reported missense mutation 689G>A (G230E) was found. We also report two known polymorphisms: 261C>T (T87T) and 2154T>C (D718D) in exons 2 and 17 of two MC families; also found in 14 (33%) and 28 (67%) of 42 healthy controls, respectively. These findings expand our knowledge of mutations responsible for myotonia congenita, reducing the proportion of MC patients in whom genetic alterations have not been found. PMID- 10533076 TI - A polymorphic microsatellite XNP-GT in the XNP/ATRX gene's promotor allows familial indirect diagnosis. PMID- 10533077 TI - A germline mutation in the von Hippel-Lindau disease gene (L178Q) detected by denaturing gradient gel electrophoresis in a large Jewish-Yemenite family. PMID- 10533078 TI - Event-related potentials and functional MRI: a comparison of localization in sensory, perceptual and cognitive tasks. AB - Combining ERP and fMRI methods to elucidate the time course and anatomical basis of information processing may provide a powerful new tool for understanding human brain function. Attempts to model the time course of fMRI activations by recording surface electromagnetic fields require a better understanding of the relationship between ERPs and fMRI activations. The results presented here show that good correspondence can be obtained between the location of ERP generators and fMRI activations in sensorimotor cortex, and in face perception and target detection tasks. However, difficulties in obtaining somatosensory fMRI activations with stimuli known to evoke robust SEPs, and the lack of fMRI activations within the hippocampus in tasks that elicit large hippocampal field potentials suggest that not all stimuli or tasks that evoke focal ERPs will evoke fMRI activations. This may be related to differences in the sensitivity of the fMRI that will be overcome with better technique and with more sensitive instruments. However, caution must be exercised in developing models that assume a one-to-one correspondence between ERP generators and fMRI activations. PMID- 10533079 TI - Evoked potentials in diagnosis and monitoring of multiple sclerosis. PMID- 10533080 TI - An integrative neuroimaging approach to restorative neurology. PMID- 10533081 TI - Neural mechanisms of BAEP. PMID- 10533082 TI - Short and middle latency auditory evoked potentials in non-smokers and tobacco smokers. PMID- 10533083 TI - Divisions of the auditory cortex suggested by EPs to synthesised instrumental tones. PMID- 10533084 TI - How much has been solved regarding SEP generators? PMID- 10533085 TI - High frequency components in somatosensory evoked potentials. PMID- 10533086 TI - High frequency (600 Hz) bursts of spike-like activities generated in the human cerebral somatosensory system. PMID- 10533087 TI - Direct recording studies of spinal and subcortical somatosensory evoked potentials in humans after median and posterior tibial nerve stimulations. PMID- 10533088 TI - Changes of somatosensory evoked potentials after decision of voluntary movement. PMID- 10533089 TI - Somatosensory evoked potentials elicited by motor point stimulation. PMID- 10533090 TI - The palmomental reflex elicitation by electrical stimulation of the median nerve. PMID- 10533091 TI - Giant somatosensory evoked potentials in different clinical conditions: scalp topography and dipole source analysis. PMID- 10533093 TI - Visual evoked potentials: advances in clinical and basic sciences. PMID- 10533092 TI - Somatosensory evoked high-frequency oscillation in movement disorders. PMID- 10533094 TI - Electrophysiological studies of parallel visual processing in humans. PMID- 10533095 TI - Temporal analysis of the VEP and parallel processing. PMID- 10533096 TI - Detection of M and P pathway deficits of amblyopia by multifocal VEPs. PMID- 10533097 TI - Development of the magnocellular VEP in children: implications for reading disability. PMID- 10533098 TI - Narrow band (approximately 17-46 Hz) oscillatory response to pattern stimulation and the visual P300 of healthy subjects. PMID- 10533099 TI - Cortical blindness and visual processing. PMID- 10533100 TI - Developmental changes in early cognitive processes. AB - The three paradigms presented in this paper demonstrate the value of ERPs in examining the development of early cognitive processes. Although we have presented only three examples, early cognitive processing could be investigated in a wide range of paradigms using ERPs, in normal as well as clinical populations. Clearly, a next step in understanding the age-related changes in these cognitive processes is to employ dipole source localization to examine the involvement of different generators and their maturation. The final conclusion is that developmental studies are important as they can contribute to our understanding of models of processing and of the generators of ERPs, in terms of cognition as well as neuroanatomy. Hence, the models of cognitive processes in adults should include the development of those processes through childhood. PMID- 10533101 TI - A study of task relevance and novelty for the P3a component. PMID- 10533102 TI - Event-related potentials reflecting sequential information processing during a mental arithmetic task. AB - The sequence of brain activities was identified with respect to the behavioral output indicated by the reaction time. The early portion of ERPs suggests two automatic processes irrespective of the adding task; one is identification of physical attributes of digits and patterns and the other is recognition of the numeric meaning specific to digits. The elicitation of the subsequent positive slow potential depends on the adding task, suggesting mental processes associated with calculation and memorizing the result of the sum. These stages of information processing concatenate sequentially during the time course of performing the adding task. PMID- 10533103 TI - The duration of the integrating window in auditory sensory memory. PMID- 10533104 TI - Phoneme representations of the human brain as reflected by event-related potentials. PMID- 10533105 TI - Evoked magnetic responses during a word completion task. PMID- 10533106 TI - Where do perception and recognition of words take place in the brain? A neuromagnetic approach. PMID- 10533107 TI - Prefrontal cortex contributions to automatic and controlled attention. PMID- 10533108 TI - Source modelling of the EEG and MEG oddball response in a subject with a large P300. PMID- 10533109 TI - Auditory verbal and non-verbal mismatch negativity (MMN) in patients with severe motor and intellectual disabilities. PMID- 10533110 TI - Nogo event-related potentials in Parkinson's disease. PMID- 10533111 TI - Changes in event-related cerebral potentials (P3a) in patients with lesions of the pontomesencephalic junction. PMID- 10533112 TI - Topography of the post-imperative negative variation in schizophrenic patients and controls obtained from high-resolution ERP maps. PMID- 10533113 TI - The physiological basis of transcranial magnetic stimulation. PMID- 10533114 TI - Magnetic cerebellar stimulation. PMID- 10533115 TI - Corticospinal volleys underlying the EMG responses to transcranial stimulation of the human motor cortex. PMID- 10533116 TI - Changes in EEG after transcranial magnetic stimulation in children with cerebral palsy. PMID- 10533117 TI - Magnetic brain stimulation: a review after 10 years experience. PMID- 10533118 TI - Pain-related brain activities: magnetoencephalographic studies. PMID- 10533119 TI - Pain-related evoked potentials from parasylvian cortex in humans. PMID- 10533121 TI - Evoked potentials to painful heat stimulation. PMID- 10533120 TI - The role of parietal opercular and insular cortex in pain sensation in humans: data from PET activation studies and intracortical recordings of CO2 laser evoked potentials (LEPs). PMID- 10533122 TI - From input to output in the somatosensory system for natural air-puff stimulation of the skin. AB - We have demonstrated in this review that the air-puff is a useful tool for combined psychophysical and neurophysiological studies of skin sensation. The major disadvantage of the previous air-puff stimulation techniques can be overcome with our air-puff system, because rigorous control of physical parameters such as velocity and force has been achieved, and synchronous activation of mechanoreceptors is now possible. It is hoped that this technique of skin stimulation will be widely used, not only by basic scientists in the field of sensory physiology, but also by clinicians who are involved in testing skin sensation during induced local anesthesia [24], or in various neurological diseases affecting skin sensation. It has the major advantage of being completely non-invasive. PMID- 10533123 TI - Magnetoencephalographic investigations of cortical reorganization in humans. PMID- 10533124 TI - Development of cortical reorganization in the somatosensory cortex of adult Braille students. PMID- 10533125 TI - Wavelet entropy: a measure of order in evoked potentials. PMID- 10533126 TI - Waveform analysis in a volume conductor. PMID- 10533127 TI - Quantitative analysis of ERD during odorous sensation. PMID- 10533128 TI - ICU monitoring. PMID- 10533129 TI - Renaissance of human brain rhythms: from description to functional significance. PMID- 10533130 TI - [Cancer of the rectum: development of surgical treatment]. PMID- 10533131 TI - [Endoscopy and hepatitis C virus: what risk? Administration Council of the French Society of Digestive Endoscopy]. PMID- 10533132 TI - [Treatment of low rectal cancer by conservative rectal resection after preoperative irradiation. Long term results and prospective study]. AB - AIMS: A prospective study was undertaken to assess the feasibility and long term results of sphincter-preserving rectal excision after preoperative radiation (35 Gy). PATIENTS AND METHODS: From 1986 to 1990, 42 patients were included in the study. Thirty four (81%) could be managed by rectal excision and stapled coloanal anastomosis. They had an adenocarcinoma located at a mean distance of 55 +/- 13 mm (range: 20-80) from the anal verge. RESULTS: Eight specimens were free of tumor. The 26 others were tabulated as follows according to the Astler-Coller staging: A = 2, B1 = 15, B2 = 5, C1 = 1, C2 = 3. The mean distal free margin was 16 +/- 11 mm (range: 1-40). The follow-up period ranged from 5 to 9 years. Six patients (18%) experienced postoperative complications including minor anastomotic leakage (n = 3), bowel obstruction (n = 2), major diarrhea requiring fecal diversion (n = 1). The functional result was good in all but 3 patients (9%) who experienced a supra anastomotic stenosis and underwent a permanent colostomy. A pelvic recurrence was observed in 5 patients (15%) after a postoperative delay ranging from 11 to 50 months. At 5 years, 17 patients (50%) were alive free of cancer, 14 (41%) of them having a good functional result without colostomy. CONCLUSION: This work demonstrates that in most cases low rectal carcinoma can be safely managed by sphincter-preserving rectal excision after preoperative radiation. It strongly suggests that the long-term pelvic recurrence rate is similar to the one observed after abdomino-perineal excision. However both procedures and patients selection must be carefully performed. PMID- 10533134 TI - [Cephalic duodenopancreatectomy for endocrine tumor of the ampulla of Vater and of the minor papilla]. AB - OBJECTIVES: Endocrine tumors of the ampulla of Vater and minor papilla are rare. This study describes the mode of presentation and evaluates the correlation between pathological features and prognosis. PATIENTS: Between 1982 and 1998, 6 patients (3 M, 3 F, mean age: 47.6 years, range: 36-58) for whom a diagnosis of endocrine tumor of the ampulla of Vater or minor papilla was made between 1982 and 1998 after histological examination of an operative specimen of pancreaticoduodenectomy. RESULTS: One patient was detected incidentally, two had a Zollinger-Ellison syndrome, two had pain and one had obstructive jaundice with pain. The tumor was located in the ampulla of Vater in 5 cases and at the minor papilla in 1 case. All patients underwent a pancreaticoduodenectomy, with histological examination showing tumor diameter varying from 5 to 40 mm and positive lymph nodes. Five patients had a well differentiated endocrine tumor and one a poorly differentiated tumor. All patients had positive Grimelius staining. The secretory profile analyzed by immunohistochemistry was heterogeneous. Median duration of follow-up was 51 months (range: 6 months-16 years) with all patients currently still alive. The patient with a poorly differentiated tumor had diffuse liver metastases, the others were disease-free. CONCLUSION: This study demonstrates the frequency of metastatic spread to adjacent lymph nodes and the inconsistent secretory profiles of these tumors. Pancreaticoduodenectomy may offer long term disease-free survival in well differentiated tumors, and such histology may be useful in advising on prognosis. PMID- 10533133 TI - [Modulatory effects of polyamines and GABA on rat ileal motility in vitro]. AB - AIM: The aim of our work was to determine the effects of polyamines and GABA on rat ileum motility in vitro. METHODS: Longitudinal strips dissected from control ileum or ileum without myenteric plexus after benzalkonium chloride (BAC) treatment were placed in organ bath chambers. RESULTS: Spermine significantly inhibited spontaneous activity and nerve stimulation-induced response. Inhibition of spontaneous activity was not altered by BAC treatment or tetrodotoxin but was antagonized by BAY K 8644, a L-type calcium channel agonist. Spermine inhibitory effect on nerve-stimulation induced response disappeared after BAC treatment. GABA enhanced the response induced by nerve stimulation but did not antagonize spermine effects; its action was inhibited in presence of atropine and was mimicked by baclofen, a GABAB agonist. CONCLUSION: Polyamines and GABA can modulate rat ileum motility in vitro. GABA acted via neural GABAB receptors. We demonstrate for the first time that spermine exerts a dual action through different mechanisms on both smooth muscle cells and probably intramural neurons. PMID- 10533135 TI - [Adenomas and other dysplastic flat polyps of the colon]. PMID- 10533136 TI - [Objectives, indications and modalities of weaning the alcohol dependent patient. Conclusions and recommendations of the jury: long text]. PMID- 10533137 TI - [Objectives, indications and modalities of weaning the alcohol dependent patient. Conclusions and recommendations of the jury: short text]. PMID- 10533138 TI - [Endoscopic band ligation versus beta-blockers in the prevention of primary digestive hemorrhage by rupture of esophageal varices: are controlled studies necessary?]. PMID- 10533139 TI - [Coinfection of hepatitis C virus and human immunodeficiency virus and antiretroviral multitherapies]. PMID- 10533140 TI - [Percutaneous catheterization of the intestinal loop of hepatico- jejunostomy: a new possibility in the treatment of complex biliary diseases]. AB - OBJECTIVES: After hepatico-jejunostomy, endoscopic exploration of the biliary tract is not possible, and percutaneous transjejunal catheterization seems to be an attractive option. PATIENTS: This is a 10 year prospective evaluation of 55 percutaneous transjejunal biliary interventions in 53 patients. RESULTS: Thirty nine patients had biliary lithiasis, 10 had suspected recurrent biliary cancer, 5 biliary stenosis, and 1 angiocholitic intrahepatic abscess. Initial success was obtained in all patients and 155 procedures were performed. Interventions included strictures, dilatation, stone extraction, stent insertion and tumor biopsy. The complication rate was 15% (mainly benign biliary sepsis) with no deaths and no surgical reoperations. Thirty two of the 39 patients with biliary lithiasis had successful extraction. Eight of the 10 patients with cancer had an endoscopic biopsy and the 2 others underwent drainage. The 5 patients with benign strictures underwent dilatation and stenting. The intrahepatic abscess was treated completely by drainage. CONCLUSIONS: The feasibility of this technique, the low morbidity and the lack of mortality has been demonstrated. This technique is well accepted by patients and may be an alternative to open surgery which is known to be very difficult and risky in patients who have had one or several prior operations. PMID- 10533141 TI - [Treatment of hepatitis C virus infection in the Poitou- Charentes region]. AB - OBJECTIVES: The prognosis of hepatitis C virus infection could be improved by early treatment. However, this is only possible if most patients with hepatitis C consult a specialized institution. The aim of this study was to evaluate the modalities of care of hepatitis C virus infection in one French district. METHODS: Between November and December 1997, 89 biological laboratories from the "Poitou-Charentes" district were asked to provide results of hepatitis C virus serology tests performed during this period. A questionnaire concerning epidemiological and follow-up data was sent to the medical practitioner who prescribed the test, for all positive tests. RESULTS: Seventy eight out of 89 (88%) laboratoires agreed to participate in the study. During the study period, 6,168 subjects were tested and 196 (3.2%) were positive. This test was a diagnostic test in 69 cases (53%) and a confirmation test in 61 cases (47%). The epidemiological questionnaire was filled out in 130 cases. The main putative factors of viral contamination were: intravenous or nasal drug addiction in 69 cases (53%), blood transfusion in 39 cases (30%), and a nosocomial risk factor in 16 cases (12%). Treatment and care of virus infection was evaluated in 113 cases from the follow-up questionnaire: a liver biopsy was performed in 30 cases (27%) and interferon therapy was administered in 13 cases (12%). Liver biopsy was not performed in 83 cases (73%) due to normal transaminase levels or a contraindication to interferon therapy. The main causes of an absence of care or follow-up were: fear of complications of liver biopsy and/or side effects to interferon therapy (19%), chronic alcoholism (18%) and active drug addiction (8%). CONCLUSION: The main causes of failure to administer adequate care in hepatitis C patients were chronic alcoholism, drug addiction and fear of liver biopsy or side effects of interferon therapy. These data should be taken into account for future screening or information compaigns for the general population. PMID- 10533142 TI - [Nitric oxide and liver diseases: deleterious or hepatoprotective role]. PMID- 10533143 TI - [Drug-induced hepatitis: diagnosis and treatment]. PMID- 10533144 TI - [Drug-induced liver disorders]. PMID- 10533145 TI - [Drug-induced liver disorders. Questions for Professor Daniel Dhumeaux]. PMID- 10533146 TI - [Acute pancreatitis revealing cystic fibrosis in an adult]. AB - We report a case of pancreatitis as the first symptom of cystic fibrosis in a 22 year-old woman. The diagnosis was suspected upon the history of nasal polyposis and severe episodes of respiratory infections in infancy. The diagnosis was confirmed by sweat test. Genotyping showed a compound heterozygosity for mutations delta F508 and 5T. Acute pancreatitis is a rare manifestation of cystic fibrosis and an exceptional mode of initial presentation of the disease. It occurs in pancreatic sufficient patients, especially in young adults. This exceptional case shows that cystic fibrosis should be suspected in each case of idiopathic pancreatitis. Indications of cholecystectomy in patients with associated gallbladder lithiasis is discussed. PMID- 10533147 TI - [Pancreatic necrosis infection by Candida parapsilosis associated with fungemia]. AB - Pancreatic necrosis infection is the most common complication affecting mortality of severe acute pancreatitis (death rate 80%). Bacterial infections particularly with coliforms or anaerobes account for the majority of cases of infected necrosis. Fungal pancreatic infections with Candida species are rare and often nosocomial. We report herein the first case of pancreatic necrosis infection with Candida parapsilosis associated with fungemia confirmed by molecular typing. PMID- 10533148 TI - [Intra-hepatic biliary cystadenocarcinoma]. AB - Biliary cystadenocarcinoma is a rare tumor of the intrahepatic biliary tract, which frequently develops in a preexisting benign biliary cystadenoma. In the present case, diagnosis was difficult because of the lack of specificity of clinical, biological and radiological findings. The correct diagnosis was only achieved by histological examination of the resected lesion. Macroscopically, the right lobe of the liver showed evidence of a whitish, multilolobed, malignant mass of about 6 cm in diameter. Upon light microscopic analysis, cysts were found to be lined with papillary forms. In some areas, epithelial cells were clearly malignant contrasting with persistent non dysplasic areas, suggesting the presence of underlying cystadenomas. Eleven months after complete surgical resection, the patient is in good condition with no evidence of recurrence. PMID- 10533149 TI - [Efficiency and reproducibility of an intraperitoneal chemo-hyperthermia technique after complete resection of peritoneal carcinoma]. PMID- 10533150 TI - [Transmission by colonoscopy of hepatitis C virus: apropos of a group of 97 patients at "presumed risk"]. PMID- 10533151 TI - [Transmural esophageal dissection after iatrogenic perforation of esophagitis dissecans visualized by thoracic x-ray computed tomography]. PMID- 10533152 TI - [Small-cell carcinoma o the esophagus: presentation of a case and review of the literature]. PMID- 10533153 TI - [Vesicular metastasis of a prostatic adenocarcinoma]. PMID- 10533155 TI - [Granular cell tumor of the biliary tract and bile ducts]. PMID- 10533154 TI - [Hepatic leiomyosarcoma revealed by cutaneous metastasis]. PMID- 10533156 TI - Uterine and cerebral vascularization in postmenopausal women treated with hormone replacement therapy. AB - The effects of hormone-replacement therapy on the Doppler flow parameters of the ophthalmic artery in postmenopausal women were studied and compared with those registered at the level of the internal carotid and uterine arteries. Fifty-seven postmenopausal patients were submitted to continuous estradiol transdermal supplementation and 12-day courses of medroxyprogesterone acetate. During the estrogen phase of hormone-replacement therapy all patients underwent (at 1, 3 and 6 months after the beginning of hormone-replacement therapy) transvaginal ultrasonographic evaluation of the pelvic organs and of endometrial thickness. On the same day, they underwent color Doppler analysis of the blood flow impedance of the uterine, internal carotid and ophthalmic arteries. Estradiol plasma concentrations were assayed on the day that ultrasonographic and Doppler examinations took place. The pulsatility index of all the arteries improved, from baseline values, during the therapy and attained stable values compared to those after the first month of treatment. Furthermore, at the level of the internal carotid and ophthalmic arteries, a significant increase of the peak systolic blood flow velocity (Vmax) was observed over the 6 months of therapy. Doppler studies of the ophthalmic artery are capable of affording specific and precise pathophysiologic information to assess peripheral intracranial blood flow variations. Furthermore, such studies may be useful in monitoring hormone replacement therapy effects on cerebral perfusion. PMID- 10533157 TI - Double-blind, placebo-controlled study of the effects of tibolone on bone mineral density in postmenopausal osteoporotic women with and without previous fractures. AB - A 2-year placebo-controlled, randomized, two-center prospective study was carried out to assess the effects of tibolone (Org OD14, Livial) on trabecular and cortical bone mass and bone biochemistry parameters in elderly postmenopausal women with and without previous fractures. In total, 107 subjects, 71 with fractures and 36 without fractures, were randomized to tibolone (n = 64) or placebo (n = 43). Their mean age was 63.1 years. Bone mineral density (BMD) (g/cm2) was assessed at baseline and every 6 months for 2 years by dual-energy X ray absorptiometry (DXA). Mean baseline values were 0.79 and 0.80 for the lumbar spine in the tibolone and placebo groups, respectively, and for the femoral neck 0.64 in both groups. Serum and urinary bone biochemistry parameters were measured concurrently. An analysis of variance (ANOVA) model including center and group was applied. The completers' group was the primary subset for the analysis; the intention-to-treat (ITT) group was also analyzed. Results are expressed as the percentage change at 24 months and the annual rate of change percentage year. The tibolone group showed an overall mean increase (vs. placebo) in BMD at the lumbar spine of 7.2% (p < 0.001) and for the femoral neck 2.6% (p < 0.001). In subjects with previous fractures increases were 6.0% and 4.0% for the lumbar spine and femoral neck, while in those with no fractures, respective changes were 8.9% and 1.1%. Overall changes in the placebo group were 0.9% and -1.6% for the lumbar spine and femoral neck, respectively. A significant fall in bone biochemistry parameters showed that tibolone inhibits osteoclastic activity. In conclusion we have found that tibolone 2.5 mg induces significant increases of trabecular and cortical bone mass in elderly postmenopausal osteoporotic women with and without previous fractures. PMID- 10533158 TI - The influence of dehydroepiandrosterone on histology of selected organs in rabbits on an atherogenic diet. AB - The influence of dehydroepiandrosterone (DHEA) in fodder on the histology of selected organs in rabbits with induced hypercholesterolemia and in healthy rabbits was studied. Rabbits were randomly assigned into four groups: (1) control; (2) atherogenic diet; (3) atherogenic diet with addition of DHEA; (4) normal diet with addition of DHEA. After 12 weeks, the rabbits were bled. Tissue samples were collected, fixed in a 0.4% solution of buffered formalin, dehydrated and embedded in paraffin. Fragments of 5-7 microns were stained with hematoxylin and eosin as well as according to the van Gieson method. Histological analysis showed features of steatosis and intense degenerative changes in analyzed organs of animals from group 2, i.e. liver, kidneys, adrenal glands, lungs and bone. The degenerative changes in the group which in addition to a fat-rich diet received DHEA, were similar to group 2, but much less intense. Histological pictures of organs of the rabbits which received DHEA and normal diet did not differ significantly from the control group. In animals with experimental hyperlipidemia, DHEA acts protectively, decreasing degenerative changes in internal organs caused by an atherogenic diet. DHEA does not change the histological picture of organs in healthy animals. PMID- 10533159 TI - Calcium antagonistic effect of 17 alpha-estradiol derivatives: in vitro examinations. AB - The calcium antagonistic effect of 17 alpha-estradiol derivatives was investigated in vitro in human aortic smooth muscle cells. The substances tested were 17 alpha-estradiol, 17 alpha-ethinylestradiol, the scavestrogens J811 and J861, and 17 beta-estradiol. Examinations were carried out by measuring 45Ca influx into the cells. All compounds showed a significant inhibition of calcium influx at the concentration of 10(-6) M. The strongest effect could be registered for the scavestrogens. Since these substances are virtually devoid of estrogenic activity, they may offer advantages over 17 beta-estradiol in the therapy and prevention of cardiovascular disease. PMID- 10533160 TI - Raloxifene analog LY 117018 effects on central and peripheral beta-endorphin. AB - Raloxifene is a selective estrogen receptor modulator with a benzothiophene structure, that exerts an estrogen-like action on some target tissues and an anti estrogenic action on the uterus and breasts. A limited number of data are available on the effect of raloxifene on neuroendocrine function. Since beta endorphin (beta-EP) is considered a marker of neuroendocrine function, the aim of the present study was to evaluate the effects of a 14 day treatment with a raloxifene analog, LY 117018, on beta-EP content in the hypothalamus, hippocampus, anterior and neuro-intermediate pituitary lobe, and in the plasma of fertile and ovariectomized (ovx) rats. The effect of LY 117018 in ovx rats was compared to that of 17 beta-estradiol. beta-EP contents were measured by a specific radioimmunoassay. While ovariectomy determined a significant decrease in beta-EP levels in the anterior and neurointermediate pituitary lobe and plasma (p < 0.01), no changes of beta-EP content in the hypothalamus and hippocampus were found. The administration of 17 beta-estradiol or LY 117018 in ovx rats significantly increased beta-EP concentration in the anterior and neurointermediate pituitary lobe, in the hypothalamus and plasma (p < 0.01), though they did not significantly modify hippocampal beta-EP content. When LY 117018 was administered together with 17 beta-estradiol in ovx animals, a clear anti-estrogenic effect in all organs and in plasma was observed, resulting in significantly lower beta-EP content with respect to the group treated with 17 beta-estradiol alone (p < 0.01). The chronic administration of LY 117018 in fertile rats significantly decreased beta-EP content in the anterior pituitary, hippocampus and plasma (p < 0.01), while it increased beta-EP hypothalamic content and did not change beta-EP content in the neurointermediate lobe. In conclusion, raloxifene analog LY 117018 has an estrogen-like action on neuroendocrine opiatergic pathways when administered alone in ovx rats, while it exerts an anti-estrogen effect in fertile or in ovx rats treated with 17 beta estradiol. PMID- 10533161 TI - Hormone patterns after induction of ovulation with clomiphene citrate: an age related phenomenon. AB - Since the introduction of clomiphene citrate (CC), more than three decades ago, a discrepancy has been observed between ovulation and pregnancy rates for which as yet no explanation exists. To investigate if ovulation disorders or abnormal hormonal patterns occur more often in CC-stimulated seemingly ovulatory cycles, we performed hormonal and sonographic monitoring in first cycles of oligo- or amenorrheic patients who were stimulated with 50 mg CC, and compared the hormonal patterns to those in natural cycles of age-matched proven fertile women. Twenty four first CC cycles were monitored. Twelve cycles appeared to be ovulatory, eleven showed no follicle development and one cycle exhibited the luteinized unruptured follicle (LUF) phenomenon. Ten ovulatory cycles were compared with 27 unstimulated control cycles. In four cycles stimulated by CC, a temporary decline in estradiol levels was apparent. In these cycles, estradiol reached a higher level on cycle day (CD) 7 or 8 compared to cycles without a decline. Such an estradiol decline was seen in only one control cycle. Furthermore, the estradiol levels on CD 7 or 8 appeared to be age-related. We conclude that the estradiol decline in CC-stimulated ovulatory cycles may be a consequence of a sharp rise after CC stimulation, and such a rise may be age-related and coincide with a diminished follicle quality. If this phenomenon is associated with a suboptimal cycle, and so contributes to the suboptimal pregnancy rates after ovulation induction treatment with clomiphene citrate, is still unknown. PMID- 10533162 TI - Influence of nicotine, cotinine, anabasine and cigarette smoke extract on human granulosa cell progesterone and estradiol synthesis. AB - To reveal the well known effect of smoking on the incidence of early abortion, the possible effects of cigarette alkaloids on progesterone and estradiol synthesis were investigated. A suspected cause for early spontaneous abortion is corpus luteum insufficiency. The present experiments evaluate the effects of cigarette smoke alkaloids on progesterone and estradiol biosynthesis. Human granulosa cells were obtained from patients undergoing in vitro fertilization and embryo transfer treatment because of infertility. Incubation of the granulosa cells with cotinine, anabasine, with the combination of nicotine, cotinine and anabasine, or with an aqueous extract of cigarette smoke resulted in inhibition of progesterone synthesis. The alkaloids and smoke extract decreased the DNA content of the culture dish. These latter findings suggested a cytotoxic effect of the alkaloids. Both cotinine and anabasine slightly stimulated the synthesis of normalized estradiol. However, nicotine, combination of all three alkaloids, and cigarette smoke extract had no significant influence on estradiol production. Taken together, these data would suggest that cigarette alkaloids inhibit cellular progesterone synthesis both by inhibiting progesterone synthesis and by causing less specific toxic effects to the cell. In contrast, cigarette smoke alkaloids slightly stimulated or had no effect on estradiol production. These concomitant actions of cigarette alkaloids partly explain the higher incidence of early abortion in pregnant women who smoke. PMID- 10533163 TI - Interstitial collagenase (MMP-1) activity in human ovarian tissue. AB - The collagen content and collagenase activity were estimated in human ovarian interstitial tissue devoid of all visible follicles in menstruating, fertile as well as climacteric women. The mean total collagenase activity in ovarian specimens taken during both follicular (n = 10, 3.97 +/- 0.58 U/g wet weight, ww) and luteal phase (n = 10, 3.39 +/- 1.24 U/g ww) of the normal menstrual cycle along with total collagen concentration (184.8 +/- 41.0 vs. 194.4 +/- 30.5 micrograms/mg ww, respectively) did not differ. Total collagenase activity of climacteric gonads (n = 5, 1.55 +/- 0.71 U/g ww) was lower than in specimens collected during both follicular and luteal phase (p = 0.0002 and p = 0.017, respectively). About 23% of the total collagenase activity in follicular phase ovarian extracts and only about 1% in luteal phase ovarian preparations was found in the latent form. The percentage of latent collagenase in ovarian tissue during the follicular phase was negatively correlated with the day of the menstrual cycle (r = -0.93, p = 0.007). Extracellular matrix remodelling in the human ovary can be correlated with the functional status of the follicular unit. PMID- 10533164 TI - Prevalence of pregnancy and postpartum thyroid dysfunction in a homogeneous population of Spain. AB - To determine the prevalence of pregnancy and postpartum thyroid dysfunction in the Spanish population, 691 women divided into six cohorts were studied. Biochemical hypothyroidism was observed in 9.4%, 4.4%, 1.9%, 4.6% and 4.8% at the first trimester of pregnancy, at delivery and at the second, fourth and sixth to twelfth postpartum months, respectively. Biochemical hyperthyroidism was observed in 6.7%, 0.8%, 1.9%, 4.6% and 2.4% in the same cohorts of subjects. Thyroiditis was observed in 4%, 0%, 0.6%, 6.2% and 7.3% in the cohorts studied. No depression or psychological alterations were detected either in pregnancy or in the postpartum period. Our data suggest that if screening for thyroid dysfunction is to be recommended in the postpartum period, this must be based on biochemical parameters. PMID- 10533165 TI - Adrenal adenocarcinoma and empty sella syndrome in a 37-year-old woman. AB - The case of a 37-year-old woman with secondary amenorrhea and clear signs of hyperandrogenism is reported. The patient underwent hormonal evaluation including circadian rhythm of cortisol, gonadotropin-releasing hormone/thyroid-stimulating hormone (GnRH/TRH) test, corticotropic-releasing hormone (CRH) test and dexamethasone suppression test. She also underwent pelvic and adrenal ultrasound examination, adrenal computed axial tomography (CAT) scan and cranial nuclear magnetic resonance (NMR). A mass about 10 cm in size was detected in the left adrenal region. The sella was empty and the pituitary displaced downward. Suspected adrenal adenocarcinoma was confirmed by histological examination after surgical removal of the mass. This case is of interest for physicians because of the mixed androgen and cortisol secretion of the adenocarcinoma in a hyperprolactinemic patient with empty sella. Moreover, it suggests the need to investigate the adrenal gland in patients with hyperprolactinemia and hirsutism. PMID- 10533166 TI - Health communication and professional preparation: health educator credibility, message learning, and behavior change. AB - Health education graduate students were surveyed to assess perceptions of their professional responsibility to be role models of healthy behaviors, characteristics of a professional role model, and related socializing experiences during professional preparation. A total of 233 randomly selected health education graduate students participated in this study nationwide. Significant inverse associations were found between students' year in graduate school and sense of excellence as a role model, graduate program satisfaction, and professional commitment (all ps < 0.05). Students' sense of professional marketability and competence to role model were statistically significant in predicting their perception that role modeling healthy behaviors is a professional responsibility, F(2, 215) = 110.25, p = 0.00001. Positive associations also were found between students' desire to improve fitness behavior, nutrition, and weight and/or body fat ratio with self-ratings as role models (all ps < 0.05). Implications for the profession and preparation are provided. PMID- 10533167 TI - Role modeling: a dilemma for the health education specialist. PMID- 10533168 TI - Role modeling: an opportunity for the health education specialist. AB - It is the responsibility of the profession to determine what is right, reasonable, and ethical health education practice. Opportunities within the profession abound to deliberate about the responsibility of health education specialists to role model positive health behaviors. Davis summarized the espoused perspectives nicely: We owe it to our profession and to our students to personally travel as far on the wellness continuum as behavioral choices will permit.... Health educators do have a special responsibility to be positive health role models by fulfilling their health potential and modeling the healthiest behaviors of which they are capable. All health educators need to accept for themselves the responsibilities that we assign to others. PMID- 10533169 TI - Psychosocial factors associated with the use of breast cancer screening by women age 60 years or over. AB - This study examined psychosocial factors related to breast cancer screening among older women. Data for the study were obtained from interviews with 719 women age 60 years or over attending rural and urban primary care clinics in North Carolina. The results indicated that 50% of the women had mammograms in the past year, 65% reported clinical breast examinations in the past year, and 31% said they practiced breast self-examinations once a month. Several psychosocial factors were significant predictors of a lower likelihood of being screened. Multivariate analysis confirmed the importance of psychosocial factors as predictors of breast cancer screening. Educational intervention to increase screening for breast cancer in this population is needed, and the results provide specific suggestions regarding the content of effective educational materials and approaches for older women. PMID- 10533170 TI - The use of significant reduction rates to evaluate health education methods for pregnant smokers: a new harm reduction behavioral indicator? AB - This article evaluates the evidence to support the use of biochemical measurement of significant reduction (SR) rates among pregnant smokers as a new behavioral indicator of "harm reduction" (HR). The results of four studies--three randomized patient education clinical trials of pregnant smokers (Trials I, II, and III) and an epidemiological study (Study IV)--are presented. Among Trial I, II, and III cohorts of pregnant smokers, control group SR rates of 7% (I), 9% (II), and 20% (III) were increased among experimental groups to 17% (I), 18% (II), and 32% (III) by the same patient education methods. Analyses of infant birthweight data in Study IV found that a patient SR rate representing a 50% or more decrease between a baseline and follow-up test was associated with an increase in adjusted birthweight of 92 grams. PMID- 10533171 TI - The African American church and university partnerships: establishing lasting collaborations. AB - This article highlights the experiences of the Health Wise Church Project, a community outreach initiative between a diverse group of African American churches and a university health education program. The objective of the program was to develop early detection and illness prevention networks among older church members. Not all partnerships results in long-term collaborations, and this article makes a clear distinction between different types of "working together" arrangements. The project discussed in this article presents a four-stage model to illustrate how organizations achieve collaborative partnerships. Involving community partners in the early phase of project planning contributed to the success of the church-university collaboration. This type of shared planning helped to sustain community interest during the project's implementation. PMID- 10533172 TI - Predicting breast-feeding intention among low-income pregnant women: a comparison of two theoretical models. AB - This study examined the applicability of the transtheoretical model and a model derived from the theory of reasoned action for predicting breast-feeding intention among low-income pregnant women. Participants completed a 70-item self report questionnaire assessing their breast-feeding attitudes, intentions, and support. A positive correlation existed between Stages of Change for breast feeding and the number of Processes of Change used by respondents. A negative correlation existed between Stages of Change for breast-feeding and the number of negative breast-feeding beliefs held by respondents. Furthermore, women's normative beliefs and outcome beliefs were significantly correlated with breast feeding intention in manners consistent with the model developed from the theory of reasoned action. After accounting for significant sociodemographic and lifestyle factors, the Processes of Change and outcome beliefs remained independently correlated with breast-feeding intention. These models are capable of predicting the intention to breast-feed and might offer an innovative approach for further breast-feeding research and intervention development. PMID- 10533173 TI - Implementation of outreach telephone counseling to promote mammography participation. AB - To increase mammography participation, the authors implemented an outreach intervention translating concepts from expectancy value theory into a motivational interviewing telephone intervention that included the opportunity to schedule a screening appointment. Process data are presented from 491 women who had not scheduled a mammogram within 2 months of receiving a mailed invitation from a managed care organization's centralized breast cancer screening program. A total of 83% of targeted women accepted the counseling calls. Counselors rated 84% of completed calls as either receptive or neutral in tone. Women with prior mammography experience were more likely to be receptive and to schedule a screening appointment during the calls than were women with no prior experience. Topics discussed during the calls also differed between women with and without prior mammography experience. Implications for dissemination of counseling interventions in health care organizations are discussed. PMID- 10533176 TI - Factors associated with occupational exposure and compliance with universal precautions in an urban school district. AB - Factors associated with occupational exposure and universal precautions (UP) compliance were assessed among employees in one urban school district. Half of the employees surveyed reported responding to bleeding injuries and cleaning blood or other body fluids (e.g., vomit, urine) during the previous school year. Also, 1 in 4 custodians and 1 in 10 teachers/teacher's aides had direct contact with blood or body fluids without protection. In multivariate logistic regression analyses, direct contact was most likely among secondary school employees in unpredictable situations who did not have protective equipment or comply with UP. UP compliance was greater among those who had protective equipment available and felt self-confident. Self-confidence was associated with having received training or protective equipment. Routine communications between administrators and employees, staff training, provision of protective equipment, and exposure incident monitoring are essential to effective implementation of UP policies in schools and work settings where occupational exposure could occur. PMID- 10533174 TI - Do black and white adults use the same sources of information about AIDS prevention? AB - Although AIDS prevention campaigns need to target population segments that are at highest risk to be effective, little is known about how various sources of AIDS information vary by race, education, and age. To determine the most common communication channels for AIDS information reported by Blacks and Whites, the authors interviewed 1,769 adults in Baltimore, Maryland, to obtain data on nine common sources of information about AIDS and analyzed their reports by race, age, and education. Television and newspapers were the most common sources but varied little across groups. National and local public health agencies, as well as medical doctors and dentists, were more commonly reported by Blacks than by Whites. Religious organizations were much more commonly reported by Blacks than by Whites. Public health organizations working collaboratively with religious organizations and health care providers might be more effective in developing AIDS prevention strategies than has been considered previously. PMID- 10533177 TI - Oral contraceptives, combined. PMID- 10533175 TI - Health care providers' perspectives on patient delay for seeking care for symptoms of acute myocardial infarction. AB - To inform intervention development in a multisite randomized community trial, the Rapid Early Action for Coronary Treatment (REACT) project formative research was undertaken for the purpose of investigating the knowledge, beliefs, perceptions, and usual practice of health care professionals. A total of 24 key informant interviews of cardiologists and emergency physicians and 15 focus groups (91 participants) were conducted in five major geographic regions: Northeast, Northwest, Southeast, Southwest, and Midwest. Transcript analyses revealed that clinicians are somewhat unaware of the empirical evidence related to the problem of patient delay, are concerned about the practice constraints they face, and would benefit from concrete suggestions about how to improve patient education and encourage fast action. Findings provide guidance for selection of educational strategies and messages for health providers as well as patients and the public. PMID- 10533178 TI - Hormonal contraceptives, progestogens only. PMID- 10533179 TI - Post-menopausal oestrogen therapy. PMID- 10533180 TI - Post-menopausal oestrogen-progestogen therapy. PMID- 10533181 TI - Physics support in nuclear medicine. PMID- 10533182 TI - Guidelines for the provision of physics support to nuclear medicine. Report of a Joint Working Group of the British Institute of Radiology, British Nuclear Medicine Society. PMID- 10533183 TI - South Thames quality standard for nuclear medicine equipment. South Thames Region Nuclear Medicine Specialty Service Committee. AB - A survey of the quality assurance of the nuclear medicine equipment in use in the South Thames Region was undertaken as part of the clinical audit process within the region. The results revealed the variation in practice across the region and highlighted the need for an agreed quality standard, together with the appropriate level of physics support to provide that standard. PMID- 10533185 TI - Rationalization of the optimal views in planar lung scintigraphy. AB - A knowledge of the segmental anatomy of the lungs is the cornerstone for interpreting lung scintigraphy. Many attempts have been made to determine the best views for the appreciation of segmental defects and various theories have been formulated to explain the mechanisms of this process. In earlier work, we hypothesized that the arrangements of the segments was the principal determinant of this process. However, data subsequently derived from work on a model of diffuse lung disease indicates that the external shape of the lobes and lungs may be the most significant contributor to the optimal views of the lungs. PMID- 10533184 TI - Administered activity of 99Tcm-MAG3 for gamma camera renography in children. AB - A method of calculating the activity of 99Tcm-MAG3 to be administered to children of different ages has been developed and evaluated. The suggested administered activity schedule is only valid for estimation of split renal function. The activity required to obtain the same count rate over the kidneys for all ages was calculated as a fraction of the activity administered to an adult by using a biokinetic model and taking attenuation effects into account. The activity schedule is based on the age of the child and was tested using renograms from patients of different ages. Statistical noise was added to the smoothed renograms simulating an injected activity corresponding to 45 MBq for an adult. The precision in the determination of split renal function calculated with four different methods was determined for 500 simulated renograms. The precision was approximately the same for all ages, but varied with the method used. The activity to be administered to a very small child is 90% of the adult activity, decreases to less than 50% between 2 and 5 years of age, and then slowly increases to 100% as the child grows to adulthood. PMID- 10533186 TI - Effect of unilateral breathing exercises on regional lung ventilation. AB - We investigated the effect of a unilateral thoracic expansion exercise (TEE), a breathing manoeuvre used by physiotherapists, on regional lung ventilation. Nine trained physiotherapists aged 22-37 years completed the study. Technegas lung ventilation scans were used to determine the effect of a right unilateral TEE performed when sitting. This was compared with a maximal deep breath. Total radioactivity in each lung was determined. Each lung was sectioned into three equal zones (upper, middle and lower) and the ratio of radioactivity for each of the corresponding lung zones calculated. Ventilation was preferentially distributed to the right lung in all participants during both breathing manoeuvres. The mean (+/- S.E.M.) radioactivity ratios (right/left lung) were greater during a unilateral TEE (1.17 +/- 0.02) than during a deep breath (1.07 +/- 0.01). Seven participants achieved significantly greater ventilation to the right middle (1.15 +/- 0.03, P = 0.02) and lower zones (1.34 +/- 0.03, P = 0.02) during a unilateral TEE than to the corresponding zones on the left; this was evident soon after the initiation of the breath. The findings of this study show that relative regional ventilation to the ipsilateral lung can be increased during a unilateral TEE in trained individuals. PMID- 10533187 TI - A comparison of the renal handling of 99Tcm-DTPA and 99Tcm-MAG3 in hypertensive patients using an uptake technique. AB - The renal uptake and outflow of 99Tcm-DTPA and 99Tcm-MAG3 were compared by analysing renal studies performed in two different departments, but with analysis techniques and computer programs using algorithms that were almost identical. Comparison was performed by a retrospective review of results from patients who were referred for renal investigations because of hypertension but who had apparently normal kidneys. The analysis of tracer outflow rates in the form of whole-kidney transit times and renal cortical transit times showed no significant difference between the two tracers. The fractional uptake rate of tracer for each patient (both kidneys) indicated that MAG3 was extracted from the blood 3.3 times faster than DTPA in patients aged 20-69 years, with a lower ratio above the age of 70. When used to measure relative renal function, there was no overall difference between the two tracers. The fractional uptake rates were also converted to flow rates, producing values of 95.8 +/- 28.0 ml.min(-1).1.73 m-2 for DTPA and 320 +/- 75 ml.min(-1).1.73 m-2 for MAG3, in hypertensive patients aged 20-40 years. These values showed a good correlation with other published GFR and MAG3 clearance rates (obtained using blood sampling methods) in normal patients of similar ages. PMID- 10533188 TI - Absolute 24 h quantification of 99Tcm-DMSA uptake in patients with severely reduced kidney function: a comparison with 51Cr-EDTA clearance. AB - The aim of this study was to determine whether absolute 24 h DMSA uptake measurements (%DMSA) correlate well with 51Cr-EDTA clearance measurements in patients with severely reduced kidney function (SRKF). Between 1990 and 1997, 55 of 482 patients who underwent EDTA clearance measurements also underwent %DMSA within 1 week. Of these, 31 were women and 24 were men (mean age 60 years; range 19-77 years). EDTA clearance was determined using the slope-intercept method. Absolute depth- and background-corrected %DMSA were determined 24 h following the injection of 185 MBq per 1.73 m2 freshly prepared 99Tcm-DMSA. All patients had EDTA clearance < or = 60 ml.min-1. Eighteen patients (group A: 9 men and 9 women, mean age 55.8 years, range 28-73 years) had EDTA clearance > 20 ml.min-1 (mean +/ S.D. = 30.9 +/- 13.8 ml.min-1), whereas 37 patients (group B: 22 women and 15 men, mean age 62.0 years, range 19-77 years) had EDTA clearance < 20 ml.min-1 (mean +/- S.D. = 10.2 +/- 6.6 ml.min-1). EDTA clearance correlated well with %DMSA for the patients as a whole and for group A (r = 0.87, P = 0.73; r = 0.79, P = 0.0001 respectively). The regression equation suggests that %DMSA is not a marker of early renal dysfunction. In group B, the r-value (r = 0.48, P = 0.004) suggests that %DMSA is reliable as a marker of severe renal dysfunction to the extent that it provides rough information. In conclusion, %DMSA may not be used as a marker of early renal impairment. Additionally, in patients with severely reduced kidney function (EDTA clearance < 20 ml.min-1), it only provides a rough estimate. PMID- 10533189 TI - Can bone marrow scintigraphy predict platelet toxicity after treatment with 186Re HEDP? AB - The toxicity of 186Re-1,1-hydroxyethylidene diphosphonate (186Re-HEDP) therapy in patients with painful bone metastases is mainly limited to thrombocytopaenia. The aim of this study was to investigate the influence of bone marrow scintigraphy on the prediction of decreased platelet counts after 186Re-HEDP therapy. Twenty-nine prostatic cancer patients with multiple painful bone metastases were included in the study. From a pre-therapy nanocolloid bone marrow scintigram, the bone marrow index (BMI) was determined as an indicator of the extent of bone marrow involvement. The influence of the BMI on the prediction of percent decrease in platelet counts was investigated. The mean BMI was 59 +/- 20. Regression analysis showed that the BMI does not improve the relationship between percent reduction in platelet count and administered dose. In contrast, we previously showed that the bone scan index (BSI) does predict the percent reduction in platelet counts before treatment. We conclude that bone marrow scintigraphy does not provide any additional information on platelet toxicity after a therapeutic dose of 186Re HEDP. Bone scintigraphy is preferred in the prediction of reduced platelet counts. PMID- 10533190 TI - Assessment of glioma viability by estimating 201Tl SPET tumour uptake volume. AB - The aim of this study was to develop a quantitative method to assess viable tumour based on post-operative 201Tl single photon emission tomography (SPET). We studied 15 patients with histologically defined highly malignant gliomas in the post-operative phase before initiation of adjuvant treatment. A 201Tl index was calculated in two ways: maximal counts versus mean counts within a region of interest (ROI). The tumour uptake volume (TUV) within the lesion was calculated from the number of voxels that had 201Tl uptake above a threshold calculated from the uptake on the contralateral side. The threshold was set at three levels: A = 1.4 times the mean 201Tl uptake in a three-dimensional reference ROI + 96.7% confidence interval (the TUV was corrected by subtraction of the volume in the reference ROI that had uptake above the threshold with compensation for unequal ROI sizes); B = 1.4 times the mean reference ROI + 99% confidence interval; and C = maximum 201Tl uptake in the reference ROI. The SPET results were compared with the tumour volumes calculated from CT scans. Thirteen tumours showed high post operative 201Tl uptake. The 201Tl index was not significantly correlated with histological grade within the group of highly malignant gliomas. 201Tl SPET tumour uptake volume method B was highly significantly correlated with CT estimated tumour volume. In conclusion, the measurement of post-operative 201Tl SPET tumour uptake volume demonstrates metabolically active glioma tissue and is an alternative method for the monitoring of glioma treatment response. PMID- 10533191 TI - Fluorodeoxyglucose PET in the evaluation of amputations for soft tissue sarcoma. AB - The aims of this study were to evaluate the uptake of fluorodeoxyglucose (FDG) in the stumps of patients who have had amputations for soft tissue sarcoma and assess its utility in identifying local recurrence of disease. Sixteen patients who had either an upper or a lower limb amputation were evaluated. FDG PET scans (half body scans to the stump +/- local emission transmission views of the stump) were performed as part of their routine follow-up for evidence of metastases over a number of years (mean = 2.6 years; range 0.25-8 years). Diffuse uptake was found in 10 stumps for up to 18 months post-surgery without any evidence of disease recurrence. Focal areas of uptake were associated with known pressure areas with skin breakdown that could be seen clinically. In the absence of localized clinical changes, these areas represented a recurrence and needed a biopsy. PMID- 10533192 TI - Accuracy of standardized uptake value measured by simultaneous emission and transmission scanning in PET oncology. AB - We assessed the accuracy of the standardized uptake value (SUV) measured by simultaneous emission and transmission scanning in cancer patients using FDG positron emission tomography (PET). Conventional, independent emission (E)/transmission (T) scans and simultaneous E/T scans were conducted consecutively in 30 patients who underwent FDG PET examinations. The SUVs of 35 mass lesions and 34 selected normal tissues were derived from the independent E/T scan and simultaneous E/T scan. Experimental studies using a cylindrical phantom were also conducted to evaluate the accuracy and reproducibility of the SUV derived from a simultaneous E/T scan. The SUVs of 18F solution in the phantom were estimated to be approximately 1, with high reproducibility in the range of total counts observed in the clinical examinations. There were no significant differences in the SUVs in 35 tumours derived from simultaneous E/T scans and those derived from independent scans, and there was a strong positive correlation between the two (r = 0.99, P < 0.01). There were also no significant differences in the SUVs in 34 normal tissue regions derived from simultaneous E/T scans and those derived from independent scans. In conclusion, simultaneous E/T scanning with FDG in patients with malignant tumours is a valid method, since the SUV derived from the simultaneous scan is accurate and reproducible. PMID- 10533193 TI - Immunoscintigraphy with 99Tcm-labelled monoclonal anti-CEA BW 431/26 antibodies in patients with suspected recurrent and metastatic colorectal carcinoma: two year follow-up. AB - The aim of this study was to evaluate immunoscintigraphy with BW 431/26 anti-CEA antibody in the follow-up of 15 patients with colorectal carcinoma. A whole-body scan followed by SPET imaging of the abdomen and pelvis was performed 4-6 and 20 24 h after the intravenous infusion of 0.6-1.0 mg of intact anti-CEA monoclonal BW 431/26 antibody labelled with 814-1110 MBq of 99Tcm. The HAMA response and serum CEA levels were determined. Immunoscintigraphic findings were verified by biopsy, radiologically and/or by 2 year follow-up. On an individual patient basis, immunoscintigraphy demonstrated an overall sensitivity of 83%, specificity of 100%, accuracy of 87%, positive predictive value of 100% and negative predictive value of 60%. Better results were achieved in the pelvic region than in the liver or in the extra-hepatic abdominal region. We evaluated 40 lesions; on an individual lesion basis, immuno-scintigraphy gave a sensitivity of 80% and an accuracy of 80%. SPET images detected significantly more lesions than whole body planar images (P < 0.05). SPET at 20-24 h detected significantly more 'hot' lesions than at 4-6 h (P < 0.01). No correlation between CEA serum levels and immunoscintigraphy was observed (r = 0.376, P > 0.05). One of nine patients (11%) developed HAMA after immunoscintigraphy. We conclude that immunoscintigraphy with BW 431/26 antibody appears able to differentiate between tumour recurrence and scar tissue, and to evaluate liver metastases of colorectal carcinoma. Serum CEA levels appear not to influence the result of immunoscintigraphy and the HAMA response is minimal. A delayed SPET scan should be part of an immunoscintigraphic imaging protocol when 99Tcm-labelled BW 431/26 monoclonal antibody is used. PMID- 10533194 TI - MAP kinases in plant signal transduction. AB - Mitogen-activated protein kinase (MAPK) pathways are modules involved in the transduction of extracellular signals to intracellular targets in all eukaryotes. Distinct MAPK pathways are regulated by different extracellular stimuli and are implicated in a wide variety of biological processes. In plants there is evidence for MAPKs playing a role in the signaling of abiotic stresses, pathogens, plant hormones, and cell cycle cues. The large number and divergence of plant MAPKs indicates that this ancient mechanism of bioinformatics is extensively used in plants and their study promises to give molecular answers to old questions. PMID- 10533195 TI - MAP kinases in plant signal transduction: how many, and what for? AB - Mitogen-activated protein kinase (MAPK) pathways are protein kinase cascades that have a function in the transduction of extracellular signals to intracellular targets in all eukaryotes. Distinct MAPK pathways are regulated by different signals and have a role in a wide variety of physiological processes. In plants there is evidence for a role of MAPKs in the signaling of pathogens, abiotic stresses, plant hormones, and cell cycle cues. A large number of distinct MAPKs in plants have been identified that are all most similar to the animal ERK MAPKs. By sequence alignment all available full length plant MAPKs can be grouped into five subfamilies. Functional data exist for members of four subfamilies and show that different subfamilies encode MAPKs for specific functions. Analysis of partial MAPK sequences from full length, truncated cDNAs and expressed sequence tags (ESTs) revealed the presence of two new subfamilies in the plant MAPK superfamily. Signature sequences valid for the superfamily of plant MAPKs and each subfamily were derived and should help in future classification of novel MAPKs. The future challenge is to unambiguously assign functions to each MAPK and decipher the other partners of their signaling pathways. PMID- 10533196 TI - MAP kinase cascades in Arabidopsis: their roles in stress and hormone responses. AB - Mitogen-activated protein kinase (MAPK) cascades have essential roles in diverse intracellular signaling processes in plants, animals and yeasts. In plants, MAPK and MAPK-like kinase activities are transiently activated in response to environmental stresses and plant hormones. In addition, transcription of genes encoding protein kinases involved in MAPK cascades in upregulated by environmental stresses. A possible MAPK cascade of Arabidopsis was identified based on both the yeast two-hybrid analysis and functional complementation analysis of yeast mutants. This MAPK cascade may have important roles in stress signal transduction pathways in Arabidopsis. PMID- 10533197 TI - MAP kinases in pollen. AB - The male gametophyte of flowering plants has a highly regulated developmental programme to ensure efficient fertilization of the ovule and the faithful transmission of genetic material to the offspring. Cell cycle control mechanisms dictate the formation of the vegetative and generative (sperm) cells, while an increase in transcriptional/translational activity and the accumulation of stored proteins and mRNA is followed by a quiescent state at maturation. A switch to a new developmental programme occurs after the pollen tube lands on the stigma with the formation of the pollen tube, growth through the style, and subsequent fertilization. Apart from the internal control mechanisms involved in this developmental programme, pollen grains must cope with physical changes during development within the anther (desiccation) and subsequently during germination on the stigma (rehydration). The metabolic and structural changes that occur throughout these processes should require signaling mechanisms to co-ordinate the appropriate response, and recent data demonstrate the presence in pollen of an array of molecules belonging to diverse signalling pathways, including mitogen activated protein (MAP) kinases. The role of MAP kinases in pollen is discussed in the context of the various developmental and physical changes that occur throughout pollen maturation and germination. PMID- 10533199 TI - Pathogen-induced MAP kinases in tobacco. AB - The activation of two tobacco MAP kinases, SIPK and WIPK, by a variety of pathogen-associated stimuli and other stresses have been analyzed (Table 1). SIPK was activated by SA, a CWD carbohydrate elicitor and two elicitins from Phytophthora spp, bacterial harpin, TMV, and Avr9 from Cladosporium fulvum. In addition to these pathogen-associated stimuli, wounding also activated SIPK, suggesting that this enzyme is involved in multiple signal transduction pathways. In all cases tested, SIPK activation was exclusively post-translational via tyrosine and threonine/serine phosphorylation. WIPK was activated by only a subset of these stimuli, including infection by TMV or harpin-producing Pseudomonas syringae (preliminary unpubl. result) and treatment with the CWD elicitor, elicitins or Avr9. In contrast to SIPK, WIPK was activated at multiple levels. Low level activation (e.g. by the CWD elicitor) appeared to be primarily post-translational whereas dramatic increases in kinase activity (e.g. by TMV or elicitins) required not only post-translational phosphorylation, but also preceding rises in mRNA levels and de novo synthesis of WIPK protein. Interestingly, under conditions where the same stimulus activated both of these kinases, their kinetics of activation appeared to be distinct. SIPK was the first to be activated. Activation of the low basal level of WIPK protein present before treatment exhibited similar kinetics to that of SIPK; however, the appearance of high levels of WIPK enzyme activity was delayed, perhaps reflecting the need for WIPK transcription and de novo protein synthesis. PMID- 10533198 TI - Mitogen-activated protein kinases and wound stress. AB - One of the most severe environmental stresses that plants encounter during their life cycle is wounding. Plants respond to wound stress by activating a set of genes that encode proteins involved in healing injured tissues. In recent years, mitogen-activated protein kinases have been implicated to be key signal molecules in the initial signal transduction pathways that mediate this stress to expression of genes. PMID- 10533200 TI - Receptor-mediated MAP kinase activation in plant defense. AB - Plants mount a complex array of defense reactions in response to attack by pathogens. Initiation of these events depends on perception and signal transduction of elicitors, which are plant-derived or pathogen-derived signals, that give rise to transcriptional activation of defense-related genes as well as to changes in activities of enzymes involved in cell wall reinforcement and oxygen radical formation. An oligopeptide, identified within a 42 kDa glycoprotein elicitor from Phythophthora sojae, activates in parsley cells typical plant defense reactions, enabling researchers to study plant-pathogen interaction at the single cell level. The oligopeptide elicitor was found to be necessary and sufficient to stimulate a complex defense response in parsley cells, comprising H+/Ca2+ influxes, K+/Cl- effluxes, activation of a mitogen activated protein (MAP) kinase, an oxidative burst, defense-related gene activation, and phytoalexin formation. PMID- 10533201 TI - Regulation of cell division and the cytoskeleton by mitogen-activated protein kinases in higher plants. AB - The microtubule-associated protein 2 kinase (MAP2-kinase), now better known as mitogen-activated protein kinase (MAPK), was initially discovered in association with the cytoskeleton, and was later also implicated in cell division. The importance of mitogenic stimulation in plant development roused interest in finding the plant homologues of MAPKs. However, data on plant MAPKs in cell division are rather sparse and fragmentary. Therefore we place the available information on cell cycle control of MAPKs in plants into a broader context. We discuss four aspects of cell division control: cell proliferation and the G1/S phase transition, G2-phase and mitosis, cytokinesis, and cytoskeletal reorganisation. Future work will reveal to what extent plants use signalling pathways that are similar or different to those of animal or yeast cells in regulating cell divisions. PMID- 10533202 TI - The MAP kinase cascade that includes MAPKKK-related protein kinase NPK1 controls a mitotic proces in plant cells. AB - The tobacco NPK1 cDNA was the first-isolated plant cDNA for a homolog of mitogen activated protein kinase kinase kinases (MAPKKKs). The kinase domain of the NPK1 protein can replace the functions of MAPKKKs in yeasts, while the amino acid sequence of the kinase-unrelated region does not have any homology to those of MAPKKKs from other organisms. Transcription of the NPK1 gene takes place in meristematic tissues or immature organs in a tobacco plant. During a tobacco cell cycle, transcriptional and translational products of NPK1 are present from S to M phase and decrease after the M phase. Expression of the NACK1 gene, which is predicted to encode a novel kinesin-like microtubule-based motor protein capable of activating NPK1, is specific to M phase, suggesting that activation of NPK1 occurs in M phase. Characterization of cDNAs for a MAPKK and a MAPK which can act downstream of NPK1 makes a proposition that the MAP kinase pathway involving NPK1 regulates a mitotic process associated with microtubules. PMID- 10533203 TI - Mitogen-activated protein kinase and abscisic acid signal transduction. AB - The phytohormone abscisic acid (ABA) is a classical plant hormone, responsible for regulation of abscission, diverse aspects of plant and seed development, stress responses and germination. It was found that ABA signal transduction in plants can involve the activity of type 2C-phosphatases (PP2C), calcium, potassium, pH and a transient activation of MAP kinase. The ABA signal transduction cascades have been shown to be tissue-specific, the transient activation of MAP kinase has until now only been found in barley aleurone cells. However, type 2C phosphatases are involved in the induction of most ABA responses, as shown by the PP2C-deficient abi-mutants. These phosphatases show high homology with phosphatases that regulate MAP kinase activity in yeast. In addition, the role of farnesyl transferase as a negative regulator of ABA responses also indicates towards involvement of MAP kinase in ABA signal transduction. Farnesyl transferase is known to regulate Ras proteins, Ras proteins in turn are known to regulate MAP kinase activation. Interestingly, Ras like proteins were detected in barley aleurone cells. Further establishment of the involvement of MAP kinase in ABA signal transduction and its role therein, still awaits more study. PMID- 10533204 TI - Signal transduction of ethylene perception. AB - Ethylene signal transduction pathway regulates various aspects of plant physiology and development. Studies of mutants defective in the ethylene response, has led to the elaboration of key genes involved in the perception of ethylene. Among them are putative ethylene receptors, Raf-like kinases, nuclear targeted proteins and transcription factors. The gene products share common motifs found in other signaling-cascade pathways in organisms ranging from bacteria to mammals. Recent biochemical studies provide insight into the function and regulation of the components of the ethylene cascade and make ethylene perception a paradigm for signal transduction in multicellular organisms. PMID- 10533205 TI - Stroke registers--why bother? PMID- 10533206 TI - Cardiac services: bigger is better but managed clinical networks are best. PMID- 10533207 TI - The Dundee Stroke Register: experience of the first ten years. AB - The objective to establish a register of all patients admitted to Dundee hospitals with acute stroke has been achieved. To do this a computerised database system has been established and a prospective survey has been conducted of clinical data of patients at time of admission, with follow-up at one and three years. All patients with a WHO diagnosis of acute stroke (excluding patients with subarachnoid haemorrhage), from January 1988 have been recorded. By the end of 1998, 3222 patients had been registered. The Dundee Stroke register database is the largest in Scotland and one of the largest in the world. The establishment of the register and database has required substantial investment of resources and the collaboration of the NHS, universities and the private sector. The operation of the register has heightened interest in stroke. It also provides present and future opportunities for clinical research and audit studies and the monitoring of outcomes. PMID- 10533208 TI - A mobile screening unit, practicalities and problems. AB - The need for a mobile screening unit to travel throughout Scotland to measure and modify cardiovascular risk factors in the apparently healthy working population was fulfilled by the conversion of a double-decker bus. The aim of the project was to assess the incidence of risk factors in healthy individuals and to modify their risk factors by individual counselling. Many unforeseen practical problems were encountered and overcome during the planning stage and during the project. The team of specially trained staff spent three and a half years screening more than 20,000 people throughout Scotland. This project provides a model for other screening projects involving a multicentre/scattered cohort. PMID- 10533209 TI - Emergency psychiatric detentions: inter regional comparison of sections 24 and 25 of the Mental Health (Scotland) Act 1984. AB - There are few published studies examining the emergency compulsory admission of patients to hospital under sections 24 and 25 of the Mental Health (Scotland) Act 1984. This retrospective case-note study reviewed 200 case-notes in Fife and Edinburgh to investigate the type of patients detained by general practitioners (GPs) and psychiatrists, in neighbouring health regions in which emergency psychiatric service provision differs. There were significant differences between the patients so detained, with regard to demographic variables and outcome of detention, but not for diagnoses. These were consistent findings for the two health regions investigated. There was evidence of mis-application of the 'emergency recommendations'. Situational factors are important in the decision to detain a person. Differences in psychiatric provision in the community may have an influence on the outcome of these detentions. Further studies would help to further define these variables. PMID- 10533210 TI - Fingertip trauma in children from doors. AB - Fingertip and nailbed trauma caused by doors is common in children, occurring when fingers are either shut in the door itself or are trapped in the hinge as the door is closed. An audit was carried out over five months of all fingertip and nailbed injuries due to trauma from a door. One hundred and eighty eight children, 2% of all attendances in this period, had sustained such trauma, 39% of these occurring in children under four years of age. One hundred and forty seven children (75%) had sustained relatively minor soft-tissue injury to the finger, However the remaining forty seven (25%) of the injuries sustained were more serious e.g. Avulsion of the nail from the nailbed or amputation of part of the fingertip and 29 (15%) of all the cases required a general anaesthetic for exploration, cleaning and repair. The Plastic Surgery department followed up these 29 children and 71 Accident & Emergency follow-up appointments were generated by the remaining injuries. The incidence of significant injury was higher than expected and caused considerable distress to both the children and their parents, It is suggested that home safety protocols should feature advice on how to avoid these injuries. PMID- 10533211 TI - Right heart failure as the dominant clinical picture in a case of primary amyloidosis affecting the pulmonary vasculature. AB - A 91-year-old female patient died of right heart failure and pulmonary hypertension. The autopsy revealed multi-organ vascular amyloidosis and pulmonary alveolar septal amyloidosis with no evidence of parenchymal myocardial amyloid deposition. This is a rare example of cor pulmonale secondary to pulmonary amyloidosis. PMID- 10533212 TI - Listeria endocarditis causing aortic root abscess and a fistula to the left atrium. AB - We report the case of a 74-year-old man who presented with endocarditis on a porcine aortic valve replacement. Five of six blood cultures grew listeria monocytogenes. Transoesophageal echocardiography demonstrated the presence of a cavity posterior to the aortic annulus, apparently communicating with the left atrium. The patient underwent successful aortic valve re-replacement. Listeria endocarditis is rare with only 58 reported cases in the literature and is associated with high mortality. PMID- 10533213 TI - [Nikolai Nikolaevich Samarin (1888-1954)]. PMID- 10533214 TI - [Results of carotid endarterectomy in patients with obliterating atherosclerosis based on data of neuropsychological status and electrophysiologic indices]. AB - The clinical picture and functional indices were investigated in 22 patients with hemodynamically significant stenosis of the major arteries of the carotid basin before and after carotid endarterectomy at the period up to 3 years. It was shown that medical efficiency of the carotid endarterectomy was more pronounced in patients with the intact compensatory blood flow. PMID- 10533215 TI - [Diagnosis and correction of pathological changes of the tracheobronchial tree during lung surgery]. AB - In 32% of patients operated on the lungs various changes in the tracheobronchial tree develop as early as during the surgical intervention. They are of determining significance in pathogenesis of postoperative disorders in the external breathing function. Bronchoscopy is the most effective method of diagnosis and of timely correction of these disorders during operation on the lungs. PMID- 10533216 TI - [Choice of the method of plasty in hiatal hernia]. AB - Results of operative treatment of patients with esophageal hernias of the diaphragm can be substantially improved in cases of differential approach to choice of the plasty method. Choice of the method of plasty depends on the hernia type, topographic-anatomical structure of the esophageal opening of the diaphragm, concomitant diseases and age of the patient. Preference should be given to methods of non-complete fundoplication performed from the transabdominal access. PMID- 10533217 TI - [Characteristics of Helicobacter infections in gastroduodenal ulcers and their complications]. AB - The presence of Helicobacter pylori (HP) was studied in 143 patients with ulcer disease of the stomach and duodenum. The urease test, immunoenzyme analysis, "AEROTEST" and culture of HP in laboratory conditions were used for diagnosis of HP. On the grounds of the culture method three degrees of colonization of the gastroduodenal mucosa were established. It was found that 96.3% of the patients were infected with HP, 86.3% of them having massive colonization. All the patients with complicated ulcers of the duodenum were HP-positive. A severe degree of the HP-infection with the predominant involvement of the antral part was characteristic of patients with bleeding ulcers; in patients with perforative ulcers and stenosis of the pyloric part the HP status is notable for considerably greater variability. Among the patients with gastric ulcers there was less amount of cases with HP (88.2%) as compared with those having duodenal ulcers. All patients with complicated disease were infected with HP, but there was nothing specific in colonization depending on. PMID- 10533218 TI - [Combined vagotomy in the surgical treatment of patients with duodenal ulcer]. AB - Short-term and long-term results of surgical treatment of 487 patients with duodenal ulcer were studied. In 337 patients (69.2%) truncal vagotomy was performed; in 94 patients (19.3%)--combined vagotomy; in 56 patients (11.5%)- selective vagotomy. With no lethal outcomes, the frequency of general surgical complications was 5.6% which did not depend on the type of vagotomy. According to the Visick criteria long-term excellent and good results were noted after truncal vagotomy in 90.4% of the patients, after combined vagotomy--in 88% and after selective proximal vagotomy--in 70% of the patients. With the same frequency of recurrent ulcers, diarrhea and dumping-syndrome combined vagotomy has some advantages in recovery of the motor evacuating function as compared with truncal vagotomy; yielding in frequency of diarrhea it has better long-term results by the Visick criteria as compared with selective proximal vagotomy and less frequency of recurrent ulcers. PMID- 10533219 TI - [Effects of balloon occlusion of the inferior vena cava on indices of regional and systemic hemodynamics]. AB - The influence of interruption of blood flow along the vena cava inferior on the indices of portohepatic and systemic hemodynamics was studied in 10 patients with a combined portal and caval hypertension and in 5 patients without such combinations. It was found that occlusion of the vena cava inferior was followed by some changes to the hemodynamic induces, their degree being dependent on the occlusion level and the diagnosed portal and caval hypertension. Patients with hepatic cirrhosis and symptoms of caval hypertension overcome the interruption of circulation along the vena cava inferior better than those without signs of portal and caval hypertension. PMID- 10533220 TI - [Problem of acute appendicitis]. AB - A combination of various kinds of allergic reactions of both the specific immediate type and of the local reaction of hypersensitivity of the slow type plays its role in the development of acute appendicitis. As a rule, the process is developing by the mixed variant with the involvement of the autoimmune component. Clinico-laparoscopic verification of the character of alterations in the vermiform process allows differentiation of primary (true) appendicitis and secondary (false) appendicitis. The present-day conception of pathogenesis of acute appendicitis determines the necessity of a differentiated approach when choosing the strategy of treatment. PMID- 10533221 TI - [Evaluation of functional potentials of microcirculation in patients with obliterating diseases of lower limb vessels]. AB - An analysis of the data of laser doppler fluorometry in 136 patients has shown that different obliterating diseases have different typical fluorometric features of changes in microcirculation. Results of the postischemic test are of greater informative value in the noninvasive assessment of the functional state of microcirculation. The significance of the functional tests was particularly increasing in critical ischemia of the lower extremities. The intraoperative determination of the muscle blood flow allowed performing the control of the lower extremity revascularization and determining the optimum level of amputation. PMID- 10533222 TI - [Surgical treatment of patients with critical ischemia of the lower limbs of atherosclerotic etiology]. AB - The authors analyze the results of conservative and operative treatment of 546 patients. The results of operative treatment (reconstructive operations on the arteries, osteo-trepanation) were shown to be considerably better in cases of critical ischemia than those of conservative treatment (infusion therapy, hemodilution, plasmapheresis). PMID- 10533223 TI - [Use of arthroscopic arthroplasty in the treatment of patients with degenerative diseases of the knee joint]. AB - An analysis of results of treatment of 38 patients with 40 degenerative injuries of the knee joint by arthroscopic arthroplasty shows clinical effectiveness of this method only in cases with light and medium degrees of gravity of the disease. The absence of clinical effects of the complex conservative treatment of patients with osteochondrosis and persistent painful syndrome, mechanical symptoms of disturbances in the joint, recurring synovitis can be considered as indications for arthroscopic operations. The probability of achievement of positive results of arthroscopic treatment is higher in patients with the I-III degrees of injuries of the cartilage and IV degree--with a full thickness defect of the articulation surface as big as 1 cm in diameter in one part of the knee joint and on one of the joining condyles. PMID- 10533224 TI - [Surgical methods in the treatment of patients with sequelae of hip joint injuries]. AB - The article analyzes results of treatment of 64 patients with sequelae of traumas of the hip joint. They showed that arthrodesis of the hip joint was indicated in treatment of young patients with severe posttraumatic coxarthrosis, destruction of the cotyloid cavity or the femur head including those after fighting injuries of the joint, purulent complications after traumas or preceding operations. Intertrochanteric osteotomy of the femur is reasonable in treatment of young patients with the initial stages of coxarthrosis and preserved amplitude of movement of the joint. Endoprosthesis of the hip joint is the operation of choice in patients older than 40-45 years with posttraumatic coxarthrosis and aseptic necrosis of the femur head, as well as in patients with the recurrent pain syndrome after intertrochanteric osteotomy. PMID- 10533225 TI - [Treatment of patients with postgastrectomy syndromes]. AB - An experience with treatment of 545 patients with organic, functional and combined postgastroresectional syndromes (PGRS) is analyzed. The patients are divided into groups depending on the character of the syndromes and their combinations. The most frequent causes of the development of PGRS are formulated. Special methods of preoperative examination and preparing are described which allow individual approach to choice of the method of reconstructive-restorative operations. Some technical details of the reconstructive interventions are described. In 95% of the patients operated upon long-term results of treatment were good and satisfactory. PMID- 10533226 TI - [Diagnosis and treatment of recurrence of gastroduodenal hemorrhage in the early postoperative period]. AB - Results of treatment of 68 patients with early postoperative gastrointestinal bleedings operated upon for acute ulcerous bleedings were analyzed. Among the most frequent sources of the bleeding were sutured ulcer and acute postoperative ulceration of the mucosa. Therapeutic treatment was effective but in 42.6% of the patients which is related with the development of disseminated intravascular blood coagulation. Operations were performed on 36 patients, 9 of them died. Total lethality was 27.9%. Surgical measures for the postoperative gastrointestinal bleedings are discussed. PMID- 10533228 TI - [Determination of the volume of infusion therapy]. AB - A method of determining the diurnal volume of infusion therapy is described which takes into account the age and weight of the patients, normal indices of the diurnal requirements of water, values of morbid losses of water in enteroparesis, fever, dyspnea and vomiting. The calculation normogram proposed is based on the Aberdeen normogram and is recommended for wide clinical practice. PMID- 10533227 TI - [Problems of development, production and clinical use of highly effective blood substitutes]. AB - An analysis of biological properties of blood substitutes has shown the expedience of wider clinical use instead of polyglucin of such substitutes as polyglucol, polifer, rondex, neorondex, polyoxidin, volecam; instead of rheopolyglucin--the rheopolyglucin with glucose, rheogluman and rheoglusol prepared for clinical tests; instead of haemodesin--haemodesin-H, neohaemodesin, gluconeodesin; instead of protein hydrolysates--polyamine and purified from peptides infusamine, hydromine, amicin; instead of 0.9% solution of sodium chloride--disol, quintasol, lactasol, mafusol. They have polyfunctional properties and considerably increase the effectiveness of the infusion- transfusion therapy. PMID- 10533229 TI - [Method of puncture electrocoagulation in the treatment of patients with varicose veins]. AB - The author proposes to treat varicose veins by electrocoagulation using the needles of different diameter connected with the device EC 500 M. The method was used in treatment of 91 patients mostly with the 2nd and 3rd stages of chronic venous insufficiency. Ligation of the opening of the great subcutaneous vein and coagulation of it was simultaneously fulfilled in 56 patients. The treatment was performed in the out-patient clinic. In most of the patients (97.9%) good clinical and cosmetic results were noted. The method is sufficiently radical and allows the complications characteristic of sclerotherapy to be avoided. PMID- 10533230 TI - [Sanative postoperative laparoscopy using rapid bacteriological methods in comprehensive treatment of patients with diffuse peritonitis]. AB - In treatment of patients with diffuse peritonitis the authors have been using postoperative sanative laparoscopy which proved to provide a means not only for rapid evaluating the dynamics of peritonitis course, but also for carrying out a series of therapeutic manipulations aimed at proper management of infections and inflammatory complications. Sanative laparoscopy was accompanied by taking samples of peritoneal exudate to study sensitivity of microflora to antibiotics and antiseptic substances, ensuring rational antibacterial treatment of peritonitis. This therapeutic method was used in 38 patients who underwent 60 examinations. Sanative laparoscopy was of a planned character in 30 patients, and in 8 cases it was carried out for emergency indications. All the studies were performed within 12-23 hours. A single examination was carried on in 11 subjects, the rest of the patients demanding from 2 to 3 examinations. No complications related to sanative laparoscopy were observed. Positive outcome was noted in the majority of the patients, re-laparoscopy being indicated but for 3 patients. Two patients died due to causes not related to the pathology involved. PMID- 10533231 TI - [Are all possibilities of using standard instruments explored in difficult cannulations of the major duodenal papilla?]. AB - An analysis of 132 patients with the most difficult cannulation of the major duodenal papilla was made. The procedure efficiency increased by 15.2% after using a Dormia basket. Different kinds of endoscopic papillotomy used with the application of standard instrument have also increased the efficiency of cannulation from 70.5 to 95.4%. Complications and lethality made 11.4 and 1.5% respectively. These results are considered to be not high under conditions of the most difficult cannulation of the major duodenal papilla. The opportunities of application of standard instruments for difficult cannulations have not been exhausted yet. Continuous search for new technical developments can considerably improve the results. PMID- 10533232 TI - [Acute anal fissures in puerperants]. AB - The author shares his experiences with treatment of 236 puerperas with anal fissures. Three types of anal fissures are established. The appearance of anal fissures can be caused by precipitated labor, large fetus, episio- and perineotomy. The main attention in cases with postpartum anal fissures was given to local treatment by different means which included arrest of the pain syndrome and formation of the granulation barrier till the appearance of regular stool and prevention of constipation. Long-term results were good. PMID- 10533233 TI - [Pericardial echinococcosis]. PMID- 10533234 TI - [Hemobilia in cholelithiasis]. PMID- 10533235 TI - [Gangrenous pansigmoiditis]. PMID- 10533236 TI - [Traumatic rupture of the vermiform process]. PMID- 10533237 TI - [Traditions and innovations in teaching surgery at medical universities]. PMID- 10533238 TI - [The old goal, the new tasks. Teaching surgery to students of medical universities]. PMID- 10533239 TI - [International Conference, "Wounds and wound infection" (Moscow, 11-13 November 1998)]. PMID- 10533241 TI - [The question of the 1st in the world cadaveric kidney allotransplantation to man]. PMID- 10533240 TI - [Prognostically significant factors of complications after liver resection]. AB - The results of investigation on prognosing complications after resection of the liver in 165 patients are presented. A uni- and multifactorial regression analysis included 31 preoperative and 17 intraoperative indices. The results obtained have shown that cirrhosis, the level of bilirubin before operation and the volume of resection of the liver are independent factors which are of importance for prognosis of complications after resection of the liver. PMID- 10533242 TI - [Formation of surgical deontology in Russia]. PMID- 10533243 TI - [Infusion-transfusion therapy in the system of intensive care in traumas and hemorrhage]. PMID- 10533244 TI - [Ultrasonic diagnosis of intestinal obstruction]. PMID- 10533245 TI - [Ivan Petrovich Pavlov and contemporary science. For the 150th anniversary of his birth]. PMID- 10533246 TI - [Memory disorders and deep structures of the brain]. AB - Clinical and neuropsychologic examination of 141 patients with arteriovenous malformations (AVM) was examined according to A.R.Lurye method. AVM of caudate nucleus were found in 27 patients, of thalamus in 34 ones, of hippocampal formation in 39 individuals, of gyrus cinguli in 41 cases. 102 patients were operated. Both total impairment of the memory and its peculiarities in damages of different structures were found in patients with AVM. A common peculiarity was the development of the amnestic symptom complex similar to Korsakov's syndrome. Such disorders occur only in combined damages of the deep structures (before the operation in patients with ventricular hemorrhages), excluding patients with AVM of caudate nucleus. The memory impairments were modal-nonspecific; in all the patients an audio-speech delayed memory was altered and reproduction in visual memory. Peculiarities of memory impairments in damage of separate structures were functional asymmetry of the defects of memory in AVM of caudate nucleus and thalamus (if present) as well as permanent inclusions and contaminations in AVM of gyrus cinguli. During process of evolution separate sides of memory function might be doubled in different structures, and each of them might have its own contribution to memory function. PMID- 10533247 TI - [Chronic anxiety and phobic disorders with persistent agoraphobia: clinical and follow-up study]. AB - There were studied 2 groups of the patients with a diagnosis of agoraphobia (according to ICD = 10). The first group included 34 patients which didn't use a specialized psychiatric service; the second one included 25 patients which needed an active therapy under conditions of psychiatric hospital. Dynamics of a disease was investigated by the method of retrospective (3 years) and following prospective (3 years) evaluation. The first group was characterized by relatively favourable outcome of chronic anxious-phobic disorders (APD) with the phenomena of a stable agoraphobia (5.8% of patients with a decrease of social adaptation): a limited agoraphobic avoidance (2 cases in the average), a rare and only psychogenic exacerbation (23 cases). Comorbid disorders were presented as minor depression (53%), somatophormic disorders (single isolated cardialgias and the conversive disorders--28%), personal disorders of hyperthimic (53%) and hysteric (35.5%) type. The second group was characterised by relatively worse outcome of chronic APD with the phenomena of a stable agoraphobia (32.0% of the patients with a decrease of social adaptation), that was associated with more generalized avoidance behaviour (more than 2 cases), with a gradual increase of both the severity of panic attacks and agoraphobia in limits of either periodic long-term aggravations (46%) or a continuous progredient course (29%). As compared with the 1-st group the second group was also characterised by significantly higher average number in a month of the panic attacks (4.9 + 1.1 vs 2.4 + 0.4; p < 0.01) and hospitalization (2.5 + 0.6 vs 0.2 + 0.2 + 0.1; p < 0.05) during all period of prospective observation. More severe comorbid disorders were revealed: slow progredient schizophrenia (20% vs 0% in the first group; p < 0.01), a major depressive disorder (28% vs 3%; p < 0.01), dysthymic disorder (32% vs 3%; p < 0.05); personal disorders were presented mostly by the deviations of schizoid type (59%). PMID- 10533248 TI - [Interdisciplinary physio-reflexotherapy in the treatment of headache]. AB - To treat the patients with a headache (HA) and with the signs of muscular articular disorders in a cranio-cervical region a complex physioreflexotherapeutic approach was applied. It was directed to the relaxation of pericranial muscles, to the restoration of a physiologic mobility in a zone of cranio-cervical transition and to the action to the irritative zones of these regions. 60 patients with a headache of tension (HAT) and 48 ones with migraine (M) were treated. A therapy included laser and manual physioreflexotherapy and an education to the muscular autorelaxation technique. An estimation after 6 months of therapy gave good and excellent results in 61% of the patients with HAT and in 37% of the patients with M. A number of the days with HA, a duration and an intensity of the attacks decreased. A conclusion was made that the disorders of craniocervical structures may make worse a course of HA. In such cases there was indicated a described therapeutic approach. PMID- 10533250 TI - [Diagnostic value of rosette-forming acetylcholine specific lymphocytes in the blood of myasthenia gravis patients]. AB - In a blood of 37 patients with different forms of myasthenia (M) there was found an increase of a number of rosette-forming lymphocytes specific to acetylcholine in a specific rosette-forming reaction with acetylcholine erythrocytes' diagnostic kit. A value of this index correlated with a duration of the disease, with a degree of its generalization, with a degree of its severity and with the type of myasthenic process' course. Meanwhile, it wasn't depended on the age of the patients. In a blood of both healthy individuals and the patients with some other neurologic diseases a number of rosette-forming lymphocytes specific to acetylcholine was considerably lower. A number of the lymphocytes specific to acetylcholine was increased even in the debut of M and remained altered during all the period of the disease. That may serve as an additional diagnostic marker of M in any age, at different character and severity of the disease, in any phase of this pathology. PMID- 10533251 TI - [A structural organization of the sensory projections on a reticular formation of the brain stem]. AB - The animal experiments were performed by the cutting the sensory afferent tracts which had the projections on the reticular formation (RF) of brain stem, namely- visual, acoustic and taste tracts. Brain was investigated by methods of Marki, Nauta, Fink-Haimer. A quantitative evaluation of the voluminous fractions of neurons and gla was carried out by using "Classimat" device. Capillary density was also counted. A morphologic substrate for the conduction of the specific influences was found in form of both afferent fibers entering in brain stem and the cell groups associated with them. Between the specific brain stem and reticular structures of medulla oblongata, brain stem and midbrain (n.cuneiformis, n. tegmenti pedunculo-ponticus, n.centralis oralis and caudalis pontis, n. papillioformis, n. parabrachialis medialis and n. parvocellularis) intermediate zones were determined which were localized on the junction between sensory and reticular nuclei and had specific afferentation. Intermediate zones were also found in RF between the nuclei in both oral-caudal and mediolateral directions. An idea was formulated about the differencies in a structural organization of RF in medulla oblongata, brain stem and midrain, that were conditioned by a multicentral localization of RF functions. PMID- 10533249 TI - [Clinical peculiarities of post-withdrawal disorders in heroin abuse and approaches to their treatment]. AB - On the basis of the observation of 62 patients with heroin addiction there was described the post-withdrawal syndrome. It was characterised by a lability of psychopathologic symptomatology and by undulate course (with exacerbation and desactualization of the drive to narcotic). Clinically there were determined 3 types of post-withdrawal syndrome's course. The 1-st type was characterised by irresistible drive to narcotic, followed by psychopathic-like behaviour. For the 2-nd type a prevalence of the depressive symptomatology with different intensity of the disorders of the affect was more typical. The 3-d type was presented by unclear polymorphic manifestations and rather deep disorders of a sleep. It was established that clinical picture of the syndrome was determined mostly by a degree of a pathological drive to heroin. Differentiated approaches to a treatment of different types of post-withdrawal disorders were described. A duration of a syndrome was 3-4 weeks. PMID- 10533252 TI - [Adaptive-compensatory changes of autonomic balance in children with infantile cerebral paralysis]. AB - In children of 6-7 years old with infantile cerebral paralysis the heart muscle mobilization is observed that displays in heart rate acceleration, decrease of its variability and some diminuation of adaptive reserves of cardiovascular system. As a result of such adaptation strain the adequate maintenance of the principal hemodynamic parameter (average arterial pressure) is ensured. PMID- 10533253 TI - [A size of neurological and psychiatric problems in the last decade of the 20th century and their trends in the light of statistical epidemiological data provided by WHO]. PMID- 10533254 TI - [Lipid peroxidation and functional state of erythrocytes in children with attention-deficit disorders]. PMID- 10533255 TI - [Comparative diagnostic value of methods of visual evoked potentials registration during the stimulation by reversible chess pattern and by flash in patients with demyelinating pathology]. PMID- 10533257 TI - [The influence of aluminum ions on the phosphorylation of tubulin and brain microtubular proteins]. PMID- 10533256 TI - [Angiotensin-converting enzyme gene as a possible risk factor or protective factor in Alzheimer's disease]. PMID- 10533259 TI - [Motor potential]. PMID- 10533258 TI - [A case of subependymal heterotopia diagnosed by means of magnetic-resonance tomography]. PMID- 10533261 TI - [Scientific-practical conference "Problem of infancy"]. PMID- 10533262 TI - [The conference of interregional association of the funds for the aid to patients with nervous-muscular diseases "Hope"]. PMID- 10533260 TI - [Brain as an organ of immunity]. PMID- 10533264 TI - Binding of nystatin and amphotericin B with sterol-free L dilauroylphosphatidylcholine bilayers resulting in the formation of dichroic lipid superstructures. AB - Interactions of multilamellar vesicles (MLV) of dilauroylphosphatidylcholine (DLPC) with the polyene antibiotics, amphotericin B (AmB) and nystatin (Ny), were followed by circular dichroism (CD). These interactions proceed with both antibiotics through a slow association with high [DLPC]/[antibiotic] stoichiometric molar ratios (> or = 130), at room temperature for which DLPC membranes are in a fluid state. Microscopic investigations of the spatial distributions of the antibiotic and the MLV in the mixtures revealed that MLV form clusters inside which the antibiotic is strongly concentrated and lipid superstructures appear. Concomitantly with the appearance of these superstructures a DLPC dichroic signal emerges. This observation indicates that the chiral properties of antibiotic oligomers can induce a chirality of the DLPC molecules which are bound to them. These results support the hypothesis of a recent molecular modeling of AmB oligomers which postulates that their chiral properties result from a chiral assemblage of antibiotic molecules (Millie et al., J. Phys. Chem. B, in press). PMID- 10533263 TI - The reaction of thiolates with 2,3-dibromo-1-propanol revisited: application to the synthesis of bis(fattyalkylthio)propanols. AB - This work compares two reaction schemes for preparing 2,3-bis(fattyalkylthio)-1 propanols for further synthetic adaptation as hydrophobic analogs of lung surfactant phosphatidylcholines. An attempt to prepare 2,3-bis(fattyalkylthio)-1 propanols based on the previously published methods of Bell and co-workers (B.R. Ganong, C.R. Loomis, Y.A. Hannun, R.M. Bell, 1986. Proc. Natl. Acad. Sci. USA 83, 1184-1188; B.R. Ganong, R.M. Bell, 1987. Methods Enzymol. 141, 313-320; J.P. Walsh, L. Fahrner, R.M. Bell, 1990. J. Biol. Chem. 265, 4374-4381) was found to give the rearranged 1,3-bis(fattyalkylthio)-2-propanols as major products. As a reliable alternative, the reaction of ethyl 2,3-dibromopropionate with 2 equivalents of long chain sodium n-alkanethioates gave the corresponding ethyl 2,3-bis(n-alkylthio)propionates, which were then reduced with LiAlH4 to yield the desired 2,3-bis(fattyalkylthio)-1-propanols. Both 13C and 1H NMR spectroscopy were used to differentiate the two possible 1,3- and 2,3-dithio substituted alcohol products and to rigorously assign their structures. PMID- 10533265 TI - Resonance energy transfer study using a rhenium metal-ligand lipid conjugate as the donor in a model membrane. AB - We measured steady state and time-resolved resonance energy transfer between donors and acceptors in model membranes. The donor was a long lifetime rhenium lipid complex, which displayed a mean lifetime of 1 microsecond and lifetime components as long as 3 microseconds in the labeled DOPC membranes. The transfer efficiencies were found to be substantially larger than those predicted without consideration of lateral diffusion. The larger transfer efficiencies are consistent with a mutual lateral diffusion coefficient in the membrane near 2 x 10(-8) cm2/s. These results demonstrate that lateral diffusion in membranes can be detected with microsecond lipid probes. PMID- 10533268 TI - The karyotype of the South American rodent Kunsia tomentosus (Lichtenstein, 1830). AB - Chromosomal analysis of Kunsia tomentosus showed a karyotype with 2n = 44, constituted by 21 pairs of acrocentric autosomes. The X chromosome was a median acrocentric, between pairs 3 and 4 in size, and the Y chromosome was a small acrocentric (between pairs 19 and 20). Five pairs with nucleolus organizer regions were located at the short arms. C-banding showed blocks of constitutive heterochromatin occurring in the centromeres of all autosomes and of the X chromosome. The Y chromosome was entirely heterochromatic. In order to identify possible homologies, karyotypes of Kunsia and Scapteromys, the phyletically related taxa, were compared. No autosome shared by either genus was found by G band comparisons. The C-band patterns and those produced by Alu I, Mbo I, Rsa I and Hae III restriction endonucleases were also different. The results of FISH indicated a different composition of the telomeric regions of the chromosomes of both taxa, since in Scapteromys the probes hybridized in both telomeres, and in Kunsia this hybridization only occurred in one of the telomeres. These differences also occurred in the localization and number of nucleolus organizer regions. PMID- 10533267 TI - Impact of monocrotophos on protein and carbohydrate metabolism in different tissues of albino rats. AB - The impact of monocrotophos on protein and carbohydrate metabolism in different tissues of albino rats was investigated. The monocrotophos (0.25 mg/ml) was orally intubated into an experimental group of rats. In another group, the same amount of water was orally intubated (control group) for 29 days. The protein content was increased in liver, serum and spleen of albino rats after treatment with monocrotophos. The protein content decreased in muscle and kidney, and overall the free sugar level decreased in all tissues. The glycogen content increased in muscle, serum and kidney after treatment with monocrotophos, and the glycogen content and reducing sugar level decreased in liver and spleen. The significance of these results is discussed. PMID- 10533269 TI - Apoptosis as a mediator of hyperplastic recovery in human prostate lesions: cytochemical and immunocytochemical evaluation. AB - The present study was carried out to investigate the occurrence of apoptosis in human prostatic lesions with emphasis on nodular hyperplasia and adenocarcinomas, using cytochemistry and immunocytochemistry. The results showed that apoptosis is a common event on nodular hyperplasia but not in adenocarcinomas. This led to the hypothesis that apoptosis may represent an important factor on the localized recovery response of the hyperplastic acini. PMID- 10533270 TI - Spontaneous germ cell death by apoptosis in epididymis of the adult bat Artibeus lituratus. AB - Many factors can lead cells to apoptosis during the various stages of cell life. This study was undertaken to characterize germ cell death in the epididymis of the adult Artibeus lituratus by histochemical and immunohistochemical techniques using light microscopy and transmission electron microscopy. The results showed that cells with a nuclear phenotype and ultrastructural characteristics of chromatin compaction were common in apoptosis. The Apoptag test confirmed that the suspected cells were apoptotic. It is suggested that immature germ cells, when released from the germinative epithelium, may be directed towards the epididymis instead of being disposed of in the testicle. Furthermore, intact immature cells can leave the testicle in the initial phases of apoptosis and complete this phenomenon in the epididymis. PMID- 10533271 TI - Modification of proteins and polynucleotides by peroxynitrite. AB - Varied intensities of nitrotyrosine immunoreactivity were detected by Western blots after the reaction of proteins or enzymes with peroxynitrite (PN), a strong oxidant derived from nitric oxide. Intense immunoreactivity of cAMP-dependent protein kinase, calmodulin and most histones may depend on greater access to tyrosine residues in the reaction, whereas the absence of immunoreactivity of caspase-3, ubiquitin and S-100 proteins may reflect lack of accessibility. In addition, the changes in UV/visible absorbency were observed after PN-treatment of polynucleotides, polypeptides or proteins. Brief PN-treatment of invertase increased its enzymatic activity. Furthermore, PN-treatment of rabbit IgG decreased its recognition by anti-IgG. The results suggest that PN may chemically modify polypeptides, proteins and polynucleotides and may subsequently alter their biological activity. PMID- 10533273 TI - Jordanian population data on the PCR-based loci: LDLR, GYPA, HBGG, D7S8 and GC. AB - Genotype and allele frequency distributions for PM polymerase chain reaction (PCR)-based genetic markers were determined in a Jordanian sample population. Results were obtained using the AmpliType PM PCR Amplification and typing kit. All loci were in agreement with the Hardy-Weinberg equilibrium expectations. The predominant alleles for LDLR, GYPA, HBGG, D7S8 and GC loci were B, A, B, A and C respectively. No statistically significant variation was detected in allele frequencies of these loci in Jordanians compared to that in Israeli Arab, U.S Caucasian and Japanese populations. Data presented here can be used to estimate the frequency of a specific DNA profile in the Jordanian population for forensic analyses and paternity testing. PMID- 10533272 TI - Adolescent girls investigated for sexual abuse: history, physical findings and legal outcome. AB - OBJECTIVE: The aim of the study was to summarize the history of assault and record the results of medicolegal examination in adolescent girls under investigation for alleged sexual abuse, and to monitor the outcome of the legal process. The investigation period was 1990-94. METHOD: A consecutive series of 94 0-para girls, aged 9-22, median age 15.0 years, were examined in the head-to-toe manner including anogenital examination. Girls were referred from investigating police and social authorities. Only non-acute examinations were performed. Findings considered consistent with abusive vaginal penetration were hymenal distortion including deep clefts, hymenal and vestibular scarring, and introital diameter permitting vaginal inspection with a 17 mm speculum in the absence of consensual intercourse. Perianal scarring was recorded. STD sampling was made on indication. Findings were documented on body sketches. Medicolegal conclusions were grouped into three categories according to history and physical findings. Information on the outcome of legal procedures was collected from referring authorities. RESULTS: For 82% (77/94) of the girls, referring agencies provided examining physicians with a detailed and consistent history of abuse, presented results comprise these 77 girls. Intrafamiliar abuse was alleged by 81% (62/77), onset prior to menarche by 53% (41/77), and repeated abuse by 74% (57/77) of the girls. Abusive genital penetration was reported by 77% (59/77) and anal penetration by 19% (14/77). Sequelae after admitted self-inflicted injury were found in 15% (12/77). Deep hymenal clefts and/or vestibular scars were found in 59% (35/59) of the girls reporting penetrative abuse, compared with 6% (1/16) when non-penetrative abuse was alleged, P < 0.001. Girls with experience of voluntary intercourse could all be examined with a 25 mm speculum. Of the 17 girls without experience of consensual intercourse but alleging abusive penetration, 47% (17/36) could easily be examined with a 17 mm speculum, compared to none of 13 reporting non-penetrative abuse, P < 0.001. Non-specific anal abnormalities occurred in 10 (13%) girls; more often when anal abuse was reported, P < 0.001. No specific STDs were found. The medicolegal conclusion supported a history of abusive genital penetration in 41 (69%) cases; findings were non-specific in 11 cases and a normal anogenital status was found in 25 cases. The alleged abuse of 34 of the 77 (44%) girls was tried in court. One suspect was acquitted, 32 men were convicted of the abuse of 33 girls. Eleven perpetrators admitted abuse, and their histories were in concordance with the abuse alleged by the victims, as well as with the physical findings. CONCLUSION: A medicolegal diagnosis of alleged non-acute cases of sexual abuse relies on a detailed history. Adolescent girls alleging abuse may exhibit signs of admittedly self-inflicted extragenital injury. Our findings confirm that non-penetrative sexual acts leave no lasting genital signs, but that repeated abusive genital penetration significantly more often than non-penetrative abuse leaves deep posterior hymenal clefts and/or vestibular scarring, and a hymenal opening allowing examination with 17-25 mm specula also in girls without experience of voluntary intercourse. In cases with a confessing perpetrator, no discordance was found between the history of the victim, medicolegal conclusion and the history of the perpetrator. PMID- 10533266 TI - Optimisation of plant sterols incorporation in human keratinocyte plasma membrane and modulation of membrane fluidity. AB - The in vitro effects of plant sterols were investigated with regard to their uptake and membrane lipid fluidity in human keratinocytes. Among the different media tested to transport sterols (liposomes, micelles and organic solvents), the best results in terms of incorporation and viability were obtained by the use of the organic solvents dimethylsulfoxide and ethanol. After 48 h incubation exogenous sterol can account for about 30% of the total cell sterol content. The total sterol amount in plasma membranes increased 2-fold after incubation with cholesterol, whereas it was not altered when phytosterols were incorporated. The incorporation of cholesterol, sitosterol and stigmasterol led to an increase in the percent of unsaturated fatty acid C18:1 in the plasma membrane. The effect of this uptake on membrane fluidity was studied by means of fluorescence polarisation using DPH and TMA-DPH as fluorescent probes. Whereas cholesterol and sitosterol had no significant effect on the DPH fluorescence anisotropy (rs), the presence of stigmasterol induced a 12% decrease of rs reflecting an increase in membrane fluidity. We can conclude from this study that in the presence of sitosterol, the mean fluidity of the membrane is regulated whereas stigmasterol triggers a looseness of molecular packing of phospholipids acyl chains, in accordance with previous results obtained on purely lipid model membranes. PMID- 10533276 TI - Incidence of xenobiotics among drivers killed in single-vehicle crashes. AB - The authors have performed a study of single-vehicle crashes (SVCs) in order to verify a correlation between the loss of vehicle control and the presence of drugs in the body. Overall, 129 cases were recorded and occurred in the catchment area of the Institute of Legal Medicine in Milan between 1986 to 1996. Among the 129 cases under study, respectively 121 men and eight women, 101 were car-drivers and 28 motor-cyclists. The median age was equal to 29 years, while the average age to 32.0 years (range 15-65 years). Fifty eight cases (45.0%) were "positive" for the presence of ethanol > or = 0.8 g/l or other drugs. The sample of "positive cases" was studied according to sex, age, day, hour and type of vehicle. Considering the cases with presence of ethanol, although under the legal limit (20 cases), the total amount of cases (78) becomes even more consistent. The amount of ethanol was found to be respectively 0.34 g/l in daily drivers and 0.87 g/l in nightly drivers (p < 0.01). Our considerations confirm the importance of toxicological analyses in the forensic investigation of traffic deaths being the sample under study recorded following criteria which minimised other possible factors effecting road accidents. PMID- 10533277 TI - Skin and soft tissue artifacts due to postmortem damage caused by rodents. AB - Five cases of postmortem bite-injuries inflicted by rodents are presented (five males between 41 and 89 years; three cases caused by mice, one case by rats, one case of possible mixed rodent activity by rats and mice). The study presents a spectrum of phenomenological aspects of postmortem artifacts due to rodent activity to fresh skin and soft tissue: the majority of the injuries have a circular appearance. The wound margins are finely serrated with irregular edges and circumscribed 1-2 mm intervals within, partly showing protruding indentations up to 5 mm. Distinct parallel cutaneous lacerations deriving from the biting action of the upper and lower pairs of the rodents incisors are diagnostic for tooth marks of rodent origin but cannot always be found. No claw-induced damage can be found in the skin beyond the wound margins. Areas involved in the present study were: exposed and unprotected parts of the body, such as eyelids, nose and mouth (representing moist parts of the face); and the back of the hands. Postmortem rodent activity may occasionally be expected on clothed and therefore protected parts of the body. The phenomenon of postmortem rodent activity to human bodies can be found indoors especially under circumstances of low socioeconomic settings; outdoors this finding is particularly observed among fatalities among homeless people. PMID- 10533275 TI - Population data on the D1S1656 and D12S391 STR loci in Andalusia (south Spain) and the maghreb (north Africa). AB - Allele and genotype frequencies for two tetrameric short tandem repeat (STR) loci were determined in two population samples, from Andalusia (S Spain; n = 127) and the Maghreb (N Africa; n = 40). After denaturing polyacrylamide gel electrophoresis, 14 alleles were identified for D12S391 and 13 alleles for D1S1656. No deviations from the Hardy-Weinberg equilibrium were detected. Some statistical parameters of forensic interest (H, PD, EC) were also calculated, and the data obtained for both populations were compared. Sequencing data of several intermediate D12S391 alleles designated 17.3, 18.3, and 19.3 are also presented. PMID- 10533274 TI - Population genetic studies on the tetrameric short tandem repeat loci D3S1358, VWA, FGA, D8S1179, D21S11, D18S51, D5S818, D13S317 and D7S820 in Egypt. AB - The short tandem repeat loci (STRs) D3S1358, VWA, FGA, D8S1179, D21S11, D18S51, D5S818, D13S317, D7S820 and a locus allowing for sex-discrimination (amelogenin) can be co-amplified by the polymerase chain reaction using a commercially available kit (AmpFlSTR Profiler plus, Perkin-Elmer Biosystems, San Jose, CA) and subsequently typed using capillary electrophoresis (ABI Prism 310 Genetic analyzer, Perkin Elmer Applied Biosystems, San Jose CA). To establish databases for these loci for an Arab population sample from Egypt, 140 unrelated persons were typed. Analysis of these data revealed that all loci except for VWA were in Hardy-Weinberg equilibrium, that the combined mean paternity exclusion chance (MEC) was 0.999875 and that the combined discriminating power (DP) was 2.635 x 10(-11). The allelic distributions found in the Egyptian sample were significantly different at four loci from those found for an Austrian Caucasian population, at all nine loci from an African-American sample and at six of six loci from a Chinese sample. No evidence of linkage equilibrium between any of the co-amplified loci was found. Our results support that the combination of multiplex PCR and capillary electrophoresis can both save time and yield excellent results for paternity testing and stain analysis. PMID- 10533281 TI - Emerin. AB - Emerin encoded by the STA gene is the first nuclear protein linked with a muscular dystrophy. Emerin is a 34 kDa, predominantly hydrophilic protein with a single hydrophobic region supposed to serve as a transmembrane domain. It was classified as a type II integral membrane protein localized at the inner nuclear membrane/nuclear lamina with an ubiquitous tissue distribution. It is speculated that emerin is required for the stability and normal function of rigorously moving nuclei in skeletal muscle and heart. During mitosis, emerin is cell-cycle dependent phosphorylated and shows stage-dependent changes in distribution and localization suggesting that it plays a role in re-assembly of nuclear membranes. Mutations of the emerin gene have been associated with X-linked Emery-Dreifuss muscular dystrophy clinically defined by early joint contractures, progressive muscle weakness, and cardiomyopathy. Hopefully, identification of the protein defect may promote new therapeutic strategies concerning muscle fiber development and stability. PMID- 10533278 TI - Testing for alpha-chloralose by headspace-GC/MS. A case report. AB - A case is presented involving an acute fatality resulting from self-administered alpha-chloralose, a rodenticide. A 18-year-old man was found dead at home, with several stains of vomit on the carpet. An empty box of three bags of 5 g 100% alpha-chloralose (Corbeaux nuisibles, Rhone Poulenc) was found near the body. The compound was identified and quantified by headspace gas chromatography coupled to mass spectrometry. Alpha-chloralose was converted by concentrated sulphuric acid into chloral, a volatile compound, that was, after chromatography on a HP5-MS capillary column, identified by the following ions, m/z 82, 111 and 148. Peripheral blood concentration was 175.7 mg/l. Alpha-chloralose was quantified in several tissues, indicating kidney sequestration. No other drugs, including ethanol, were detected. PMID- 10533280 TI - Amyloid beta. AB - Amyloid beta (A beta) is a 39-43 residue amyloidogenic peptide that is deposited into the extracellular amyloid plaques which characterize an Alzheimer's disease (AD) brain. A beta is derived from the amyloid precursor protein (APP) and undergoes a toxic conformational change (gain of toxic function). The length of the A beta peptide dramatically influences its properties with the longer 42 and 43 residue species being more amyloidogenic. The genetics of familial AD (FAD) supports a central role for A beta in AD since mutations in the FAD causing genes APP and the presenilins (PS1 and PS2) increase the formation of A beta 42,43. Considerable activity is directed towards A beta as a therapeutic target. These strategies aim to inhibit A beta synthesis, A beta fibril formation, its toxic actions on cells or promote its clearance from the brain. PMID- 10533282 TI - Identification of the essential EPE1 gene involved in retention of secreted proteins on the cell surface of Saccharomyces cerevisiae cells. AB - Saccharomyces cerevisiae yeast cells secrete extracellularly low amounts of a few proteins. The reasons for retardation of secreted proteins on the cell surface remain obscure. We describe here a mutant able to export enhanced amount of proteins. Classical genetic methods, nucleic acids manipulations and cloning procedures were used to isolate and characterize the mutant and to clone and sequence the corresponding wild type gene. The isolated Saccharomyces cerevisiae mutant MW11, is temperature sensitive and exports on average twenty-fold more proteins at 37 degrees C than parental wild type strain (80 micrograms of proteins/1 x 10(8) mutant cells, SEM +/- 5, n22; versus 3 micrograms of proteins/1 x 10(8) parental cells, SEM +/- 1, n22). Protein overexport in the mutant requires a functional SEC1 pathway and is independent of cell lysis. Cloning and sequencing of the corresponding wild type gene identified an open reading frame of 786 bp coding for a hydrophilic protein with predicted molecular mass of 30 kDa and cytosolic localization. The newly identified gene, designated EPE1, is an essential gene. Its DNA and amino acids sequence showed no homology with other yeast genes and proteins. It is concluded that the function of unknown yet genes, such as EPE1 is needed for retention of secreted proteins on the surface of Saccharomyces cerevisiae cells. PMID- 10533279 TI - Adam (MDMA) and Eve (MDEA) misuse: an immunohistochemical study on three fatal cases. AB - Three fatal cases of MDMA/MDEA misuse have been examined. These referred to white males between 19 and 20 years of age, in which post-mortem toxicology showed the presence of MDMA (in one case), MDEA (in one case) and both (in one case). The clinical data were analysed and the histopathological findings were studied following immunohistochemical investigations. A complete immunohistochemical study has made it possible to demonstrate rhabdomyolysis and myoglobinuria with alterations of the organs typical of a DIC. Clinical, histopathological and toxicological data suggest that severe or fatal complications following ecstasy ingestion could be related to idiosyncratic response. PMID- 10533283 TI - Low density lipoprotein uptake: holoparticle and cholesteryl ester selective uptake. AB - Low density lipoproteins (LDL) contain apolipoprotein B-100 and are cholesteryl ester-rich, triglyceride-poor macromolecules, arising from the lipolysis of very low density lipoproteins. This review will describe the receptors responsible for uptake of whole LDL particles (holoparticle uptake), and the selective uptake of LDL cholesteryl ester. The LDL-receptor mediates the internalization of whole LDL through an endosomal-lysosomal pathway, leading to complete degradation of LDL. Increasing LDL-receptor expression by pharmacological intervention efficiently reduces blood LDL concentrations. The lipolysis stimulated receptor and LDL receptor related protein may also lead to complete degradation of LDL in presence of free fatty acids and apolipoprotein E- or lipase-LDL complexes, respectively. Selective uptake of LDL cholesteryl ester has been demonstrated in the liver, especially in rodents and humans. This activity brings five times more LDL cholesteryl ester than the LDL-receptor to human hepatoma cells, suggesting that it is a physiologically significant pathway. The lipoprotein binding site of HepG2 cells mediates this process and recognizes all lipoprotein classes. Scavenger receptor class B type I and CD36, which mediate the selective uptake of high density lipoprotein cholesteryl ester, are potentially involved in LDL cholesteryl ester selective uptake, since they both bind LDL with high affinity. It is not known whether they are identical to the uncloned lipoprotein binding site and if the selective uptake of LDL cholesteryl ester produces a less atherogenic particle. If this is verified, pharmacological up-regulation of LDL cholesteryl ester selective uptake may become another therapeutic approach for reducing blood LDL-cholesterol levels and the risk of atherosclerosis. PMID- 10533288 TI - The unmet challenges of hepatitis C. PMID- 10533286 TI - Density and substrata are important in lung type II cell transdifferentiation in vitro. AB - Morphological techniques and metabolic cell marker assays were used to study the transdifferentiation of pulmonary type II epithelial cells to type I-like cells in vitro. In the lung this process is important during remodelling and alveolar repair. Type II cell phenotype was best maintained over eight days when densely packed cells were plated out on a commercially available extracellular matrix. Such cells retained type II cell characteristics (lamellar bodies, high activities of gamma glutamyl transpeptidase and alkaline phosphatase) but expressed low levels of rT1(40) a surface protein marker of type I cells. In contrast, low density cultures, irrespective of substratum, exhibited rapid cell spreading, loss of lamellar bodies, loss of type II cell enzyme markers and expressed high levels or rT1(40). Conditions have been described whereby the same isolate of type II cells can be used to produce differential epithelial phenotypes and use can be made of this for further characterisation or to investigate the effect of toxins on different lung cell types in vitro. PMID- 10533292 TI - Bibliotherapy: an indirect approach to treatment of childhood aggression. AB - The process of group therapy with five aggressive young boys, utilizing bibliotherapy as its primary mode of intervention, was investigated and is illustrated in this paper. The rationale for using affective bibliotherapy in a group context is given, the content of the program is described, and the process is fully displayed. The effectiveness of the treatment was studied in a single subject design, comparing treatment children with their matched counterparts. Results pointed to reduced aggression of all the five treatment students, compared with no change in the control children, by self- and teacher report. In addition, results based on an analysis of transcripts showed increased constructive behavior in group for all participants. Although these results should not be generalized, they suggest an interesting line of research for future investigation. PMID- 10533284 TI - AML1, the target of chromosomal rearrangements in human leukemia, regulates the expression of human complement receptor type 1 (CR1) gene. AB - The human CR1 gene is expressed specifically in hematopoietic cells. It is suggested that some cell-type specific factors which involve in gene-specific activation or repression exist in cells according to the result that the gene expression varies differently depend on differentiation stage. Here, we demonstrate that the integrity of a polyomavirus enhancer core sequence, 5' TGTGGT-3', is critical to the human CR1 promoter activity. AML1 is a site specific DNA-binding protein that recognizes the enhancer core motif TGTGGT. We show that the AML1 binds specifically to this site and activates the human CR1 promoter. Furthermore, we demonstrate that the Ets binding site (GGAA) located 2 bp upstream of the AML1 site is also involved in the regulation of the human CR1 promoter activity. Point mutations of either the AML1 or the Ets binding site that abolish the binding of the respective factors result in significant decreases of the human CR1 promoter activity. These results suggest that AML1 and Ets proteins direct the expression of the human CR1 promoter. PMID- 10533293 TI - What is worrying children in the Gaza Strip? AB - This study used a four-part method in order to identify and understand the things that worry children in the Gaza Strip. One hundred and ninety four Gazan children, between the ages of 8 and 14, were firstly asked to generate lists of the things about which they worried. These data were then used to construct a survey questionnaire which allowed a rank ordering of children's worries and the exploration of developmental and gender differences. The final part of the study employed focus groups in which the children elaborated on their worries, spoke about the strategies they use to manage these concerns, and proffered advice for younger children who might have to face similar concerns in the future. PMID- 10533289 TI - Children at risk: outcome and cost measures needed. AB - Sporadic reports in the media focus on the difficulty of America's social welfare leadership to protect children at risk and to allocate scarce resources. These criticisms suggest the need for valid conclusions in both socio-psychological and economic terms for evaluating the efficacy of three key strategies used for children at risk: reunification, foster and kinship care, and adoption. This article calls for creating a comprehensive data base that supplies the most critical variables leading to reasonable successes and the average cost per case when comparing children reunified with a biological parent to those who are placed into out-of-home settings and to those who are adopted. This analysis to include public and private expenditures for services provided by human services- welfare, special education, judicial, correctional, mental health, medical, and other related organizations. PMID- 10533290 TI - Adolescent defense style and life stressors. AB - This study examines the relationship between stressful life events and defense mechanisms. Eighty seven female adolescent patients completed the Adolescent Family Inventory of Life Events and Changes (A-FILE) assessing stressors in six domains of family life, and the Defense Style Questionnaire (DSQ) assessing 19 defense mechanisms grouped into Immature, Prosocial, and Mature clusters. Increasing stressors are significantly positively correlated with a more immature defense style. Results support the hypothesis that there is an iterative relationship between immature defenses and life stressors. These findings are compatible with a regression model of defense functioning and complement our previous results linking defenses to temperament. PMID- 10533294 TI - Intrarenal infusion of supernatant from cytokine-activated human mesangial cells may cause glomerular damage. AB - BACKGROUND: The pathogenesis of glomerular damage in glomerulonephritis (GN) is not fully understood. Several studies have suggested that reactive oxygen molecules play a role in renal disease. It is known that, during GN, mesangial cells are activated. In a previous study, we demonstrated that in vitro interleukin (IL)-1 plus IL-6 stimulated cultured human mesangial cell (HMC) activation to release free oxygen radicals. METHODS: In this study, we measured hydrogen peroxide (H2O2) and superoxide anion (O2-) levels after stimulation by IL-1 plus IL-6 in cultured HMCs. We then infused H2O2 directly into the left renal arteries of Sprague-Dawley rats. We also infused the culture supernatants of HMCs after stimulation by IL-1 plus IL-6 into the left renal arteries of rats. Two hours after stopping the infusion, the kidneys were removed and fixed using Carson's modified Millonig's buffer for electron microscopy. RESULTS: Both 100 microM H2O2 and supernatants of HMCs stimulated by IL-1 10 U/ml plus IL-6 1,000 U/ml caused similar glomerular damage, including blebbing and sloughing of endothelial cells, and denuded basement membrane in glomeruli. When 100 microM H2O2 or supernatants of cytokine-activated HMCs were infused into renal arteries, they caused hematuria and proteinuria. CONCLUSIONS: These results suggest that activated HMCs may secrete free radicals and cause glomerular damage. PMID- 10533285 TI - Subcellular immunolocalization of protein kinase CK2 in normal and carcinoma cells. AB - CK2 is a messenger-independent protein serine/threonine kinase that has been implicated in cell growth and proliferation. Our recent analysis of squamous cell carcinomas of the head and neck (SCCHN) revealed a significant elevation in CK2 activity in these tumor cells relative to normal mucosa of the upper aerodigestive tract and suggested a correlation with aggressive tumor behavior and poor clinical outcome. In order to further define the distribution of CK2 in these tissues, we have examined the immunohistochemical staining pattern of surgical specimens of both SCCHN tumors and normal upper aerodigestive tract mucosa using a monoclonal antibody directed against the catalytic subunit CK2 alpha of the kinase, and have compared these data with the subcellular distribution of CK2 activity in these same tissues. These measurements showed that CK2 is predominantly localized to the nuclei of the tumor cells, which agreed closely with the immunohistochemical staining pattern of CK2-alpha in tumor cells. The chiefly nuclear distribution of CK2-alpha immunostaining found consistently in SCCHN tumor cells and tumor-infiltrating lymphocytes contrasted with a relatively more predominant cytosolic staining pattern exhibited by various cellular constituents of normal oropharyngeal mucosa. The immunostaining pattern of CK2-alpha revealed that staining was observed in the cells stained for the proliferation-marker Ki-67; however, strong distinct immunostaining for CK2 alpha was also observed in large numbers of other cells in these same tumors, suggesting that CK2 elevation in these tumors is not a reflection of proliferative activity alone, but may also relate to the pathobiological behavior of the tumor. PMID- 10533287 TI - Fumarate metabolism and the microaerophily of Campylobacter species. AB - (1) The role of fumarate metabolism in the microaerophily of the Campylobacter genus and the effects of therapeutic agents against it were investigated. (2) NMR spectroscopy was employed to determine the properties of Campylobacter fumarase (Fum) and fumarate reductase (Frd). Radiotracer analysis was used to determine the production of carbon dioxide by Campylobacter cells. Standard microbiological techniques were used to measure the effects of environmental conditions and inhibitors on bacterial growth. (3) All Campylobacter species tested showed both Fum and Frd activities. Frd activity was observed with or without the addition of an exogenous electron donor in the particulate fractions obtained from lysates. Fumarate was oxidized to carbon dioxide via the acetyl-CoA cleavage pathway. The genes encoding proteins involved in fumarate metabolism were identified in the Campylobacter jejuni genome. Cells grew better in atmospheres with 5 and 10% oxygen levels. Fum activity was the same in cultures grown under different oxygen tensions and did not vary with the age of cultures. Frd activity was higher in cultures which grew at faster rates and decreased with the age of cultures. Four Frd inhibitors showed bactericidal effects against Campylobacter spp. with different potencies. The relative strengths of inhibition of the compounds followed the same order as the bactericidal effects. (4) The results suggested that Frd and Fum are constitutive and play a fundamental role in these microaerophiles which show characteristics of anaerobic metabolism, and that the Frd inhibitors tested would not be of therapeutic use. PMID- 10533291 TI - Bullies, victims and bystanders: a method of in-school intervention and possible parental contributions. AB - This study developed a method of in-school intervention that dramatically reduced the suspension rate and violence in elementary schools. It suggests that children who were not read to by their parents often become bullies and/or victims of bullies. Other parental practices, including inconsistent discipline in the home, also may be contributing factors. PMID- 10533296 TI - Development of a new prognostic system and validation of APACHE II for surgical ICU mortality: a multicenter study in Taiwan. AB - BACKGROUND: To develop and to validate a new prognostic prediction system for patients admitted to the surgical intensive care unit (ICU), and to compare its performance with the Acute Physiology and Chronic Health Evaluation (APACHE) II system. METHODS: The database was derived from three surgical ICUs in three hospitals. For each patient, demographic data, diagnosis, APACHE II score and hospital survival data were collected. The accuracy in outcome prediction of the APACHE II was assessed by means of receiver operating characteristic (ROC) analysis. The new prognostic system was developed by using a multiple logistic regression in the developmental data set and validated with the validation data set. RESULTS: A total of 1,248 patients were included from three ICUs. The area under the ROC curve was 0.74 for the APACHE II score. The new prognostic system includes 18 variables. Goodness-of-fit tests indicated that the model performed well in the developmental and validation samples (p = 0.235 in the developmental data set and p = 0.297 in the validation set). The area under the ROC curve was 0.84 in the developmental sample and 0.77 in the validation sample for the new prognostic score. The area under the ROC curve was 0.71 in the validation sample for the APACHE II score. CONCLUSIONS: Although APACHE II correlates with mortality for surgical ICU patients in Taiwan, its accuracy is not as good as in the original study. Mortality prediction performance improved with the use of the new, local scoring system. PMID- 10533295 TI - Long-term effect of large biliary endoprostheses in high-risk surgical patients with irretrievable common bile duct stones. AB - BACKGROUND: Endoscopic sphincterotomy and basket extraction are currently used to remove bile duct stones, with a duct clearance rate of 85% to 90%. A biliary endoprosthesis (stent) is an alternative method to treat difficult cases, especially high-risk surgical patients. The aim of this study was to investigate the long-term effect of biliary endoprostheses in patients with irretrievable common bile duct stones. METHODS: From December, 1990, to November, 1998, 546 patients were referred to the Veterans General Hospital-Kaohsiung for endoscopic removal of common bile duct stones. Of them, 12 received long-term biliary endoprosthesis because endoscopic removal or surgery was not suitable. Large caliber stents (> 10 French) were inserted into 12 patients (5 women and 7 men, mean age, 78.4 years) and they were followed up with regular clinical visits. Quantitative cholescintigraphy was performed in four patients to evaluate biliary emptying after liver function returned to normal. RESULTS: No early complications from stent insertion occurred and a satisfactory resolution of jaundice, pain and fever were noted in all patients. One patient had repeated cholangitis due to stent occlusion and five died from unrelated causes. The median effective period of stent placement was 11 months (range, 1-38 months). Quantitative cholescintigraphy revealed delayed biliary drainage in four patients despite an absence of symptoms after their liver function returned to normal. CONCLUSIONS: Large-caliber stents are a safe and effective treatment for long-term palliation in high-risk patients with retained common bile duct stones. PMID- 10533299 TI - Expression and prognostic significance of proliferating cell nuclear antigen and Ki-67 in malignant ovarian germ cell tumors. AB - BACKGROUND: Both proliferating cell nuclear antigen (PCNA) and Ki-67 are proliferative markers known to correlate with the cell proliferative state. The aim of this study was to evaluate the usefulness of PCNA and Ki-67 immunoreactivity in the assessment of clinicopathologic features and prognosis in patients with malignant ovarian germ cell tumors. METHODS: Thirty-one patients with surgically resected malignant ovarian germ cell tumors were investigated. The clinicopathologic features and survival data of these patients were recorded. Immunohistochemical staining with monoclonal antibodies (PC 10 for PCNA, and MIB 1 for Ki-67) were performed on paraffin embedded tissue from each patient. The correlation of the immunoreactivity of these two markers with the clinicopathologic features and prognosis were studied. RESULTS: All of the tumors were positive for PCNA and Ki-67, but the intensity of expression varied widely. The immunoreactivity in each tumor was also heterogeneous. The scoring of PCNA and Ki-67 was determined by a semiquantitative method. Both advanced tumor stage (stages III and IV) and high PCNA score (scores 3 and 4) indicated a poorer prognosis for survival than did early stage (stages I and II) and a low PCNA score (scores 1 and 2) (p = 0.017 and p = 0.008, respectively). In addition, the proportion of tumor relapse and tumor-induced death was more accurately predicted by PCNA and Ki-67 scoring than by tumor staging (chi 2 = 0.3159, chi 2 = 0.7186 and chi 2 = 1.9689, respectively). CONCLUSIONS: PCNA and Ki-67 proliferative markers appear promising to differentiate patients into low- and high-risk groups. In the presence of a high PCNA or Ki-67 score, aggressive postoperative chemotherapy is warranted, even if the disease is in a very early stage. PMID- 10533298 TI - Surgical treatment of bronchiectasis: 10 years' experience. AB - BACKGROUND: Since antibiotic therapy and vaccination have been widely used in medical practice, the incidence of bronchiectasis has decreased steadily. The principal role of surgery associated with this disease is for the treatment of complications. We present an analysis of surgical results during a 10-year period. METHODS: The medical records of bronchiectasis patients who were surgically treated were retrospectively reviewed from July, 1987, to March, 1998. The surgical indications, complications and recurrences of bronchiectasis were evaluated. RESULTS: A total of 41 bronchiectasis patients underwent surgical treatment from 1987 to 1998 at our hospital. There were 18 males (mean age, 37.8 +/- 15.3 years; range, 16-73 years), and 23 females (mean age, 33 +/- 7.1 years; range, 21-46 years). The indications for surgery were hemoptysis in 30, failed medical treatment in eight, suspected neoplasm in two and retention of a foreign body in one patient. Anatomic resections of the diseased sites were carried out more frequently on the left lower lobes of the lungs. In total, there were 20 left lower lobes, five right lower lobes, 10 left lingular lobes, five right middle lobes, four left upper lobes and one right upper lobe that required surgery. Surgical complications included hemorrhage in one patient, bronchopleural fistula in one and galactorrhea in one patient. The follow-up intervals were from two to 131 months (mean, 72.5 +/- 37.6 months; median, 74 months). There were two cases of recurrent symptoms and six cases of recurrent hemoptysis; all were easily controlled by medication. There were no mortalities. CONCLUSIONS: Surgical treatment of bronchiectasis yields immediate resolution of symptoms, better quality of life and no mortalities. Cessation of smoking, avoiding air pollution and careful medical follow-up are mandatory. PMID- 10533297 TI - Gene therapy with tumor vaccine increases the survival of hepatoma-bearing mice. AB - BACKGROUND: Tumor vaccines combined with cytokine gene therapy and Bacillus Calmette-Guerin (BCG) were tested for prevention and therapeutic effects in the H6 mouse hepatoma model. METHODS: Plasmid DNA of expression vectors carrying cDNA of mouse interleukin (IL)-2 and mouse granulocyte-macrophage colony-stimulating factor (mGM-CSF) were used for transfection to obtain H6 mouse hepatoma cells that secreted IL-2 (H6/IL-2) or GM-CSF (H6/GM-CSF). For tumor prevention, groups of mice were immunized twice with irradiated tumor cells with untransduced H6, H6/IL-2, H6/GM-CSF, or an equal mixture of H6/IL-2 and H6/GM-CSF. Three weeks later, these mice were inoculated subcutaneously with live H6 hepatoma cells, and tumor growth was measured. For therapeutic studies, mice first inoculated with live H6 cells were treated three days later with various irradiated tumor cell vaccines alone or in combination with BCG. Subsequent tumor growth was measured. RESULTS: In tumor prevention studies, significant protection from tumor growth has been observed in animals vaccinated with irradiated cytokine-secreting H6 cells compared with those immunized with irradiated parental H6 cells. In tumor therapy studies, subsequent administration of irradiated H6/GM-CSF cells in combination with BCG impeded the tumorigenicity of preinoculated live H6 hepatoma cells. CONCLUSIONS: These results suggest that cytokine-secreting tumor vaccines have a prophylactic effect and BCG, in combination with irradiated H6/GM-CSF cells, shows a synergistic effect on delaying the growth of H6 mouse hepatomas. PMID- 10533301 TI - Effect of oncostatin-M on proliferation and activity in osteoblastic MC3T3-E1 cells. AB - BACKGROUND: Osteogenic cells (osteoprogenitor cells and osteoblasts) respond to specific cytokines, growth factors and hormones. Understanding which cytokines affect osteogenic cells and the consequences of those effects are central to understanding normal and pathologic bone remodeling. Oncostatin-M (OSM) is a glycoprotein interleukin-6 cytokine known to inhibit bone resorption in fetal long bone cultures. However, it is still unclear whether OSM affects bone formation. The aim of this study was to investigate the effects of OSM on bone formation (bone cell proliferation, differentiation and function). METHODS: For the in vitro bone formation bioassay, MC3T3-E1 cells were plated into 35 mm Petri dishes and cultured in Dulbecco's modified eagle medium (DMEM). Various concentrations (1, 10, 100 or 1,000 units/ml) of OSM were added daily to the experimental dishes, while only DMEM was added in the control dishes during the proliferative and differentiated stages of MC3T3-E1 cell growth. The effect of OSM on bone formation was evaluated using 3H-thymidine incorporation, alkaline phosphatase expression, type I collagen synthesis assay and phase-contrast light microscopy. RESULTS: OSM significantly decreased osteoprogenitor cell proliferation and alkaline phosphatase activity in a dose-related fashion. There was no effect on type I collagen synthesis noted at any OSM dose. Thus, OSM exerted significant inhibition on bone formation. CONCLUSIONS: OSM is a potent inhibitor of bone formation by decreasing both osteoprogenitor cell proliferation and alkaline phosphatase activity. PMID- 10533300 TI - Effects of latanoprost 50 micrograms/ml on Chinese patients with primary open angle glaucoma and ocular hypertension. AB - BACKGROUND: This study was designed to determine the efficacy and safety of latanoprost 50 micrograms/ml in Chinese patients with primary open-angle glaucoma (POAG) and ocular hypertension (OH). METHODS: A 14-day randomized, double-masked, parallel-group study comparing topical latanoprost with placebo was followed by a 10-week, one-armed, open-labeled latanoprost treatment study. Intraocular pressure (IOP), visual function, ocular manifestations and miscellaneous adverse effects were evaluated at baseline, and days 1, 7, 14, 15, week 6 and week 12 visits. RESULTS: Twenty-six eligible patients were enrolled in the study. The mean IOP was significantly reduced from baseline only in the latanoprost-treated eyes during the first study period (p = 0.003 on day 1; p = 0.004 on day 7; p < 0.001 on day 14). Meanwhile, the mean IOP was significantly lower in the latanoprost-treated eyes than that in the placebo-treated eyes (p = 0.03 on day 1; p = 0.001 on days 7 and 14). Eyes in both groups showed significantly reduced IOP at each visit of the secondary 10-week latanoprost treatment period, with a mean decrease of 6.12 mmHg. The IOP-lowering effect showed no diminution throughout the study course. Conjunctival hyperemia occurred in 16 of all patients who underwent latanoprost treatment and was the most frequent adverse effect observed. No patient withdrew from the study because of intolerable side effects. CONCLUSIONS: Topical latanoprost is effective in reducing IOP for patients with POAG and OH. The pressure-lowering effect lasts for at least 24 hours after 1 drop instillation and no drift of effect is noted during this 12 week study. Conjunctival hyperemia was the most common side-effect, which was mild in degree and recovered after discontinuation of the medication. PMID- 10533302 TI - Analysis of prognostic factors in Chinese women with breast cancer in southern Taiwan. AB - BACKGROUND: We conducted a retrospective review of all early-stage breast cancer patients treated at the Veterans General Hospital-Kaohsiung to determine overall and disease-free survival rates, and to evaluate prognostic factors for these outcomes. METHODS: During the period of October, 1990, to December, 1997, 332 patients with early-stage breast cancer were treated at our institution. Cox's multivariate regression analysis was used to select prognostic factors significant for overall survival and disease-free survival. RESULTS: The survival rate for breast cancer patients was 88.35% at five years. Prognostic factors predicting breast cancer mortality included poorly differentiated histologic grade, four or more lymph nodes positive for metastasis and negative progesterone receptor status. For disease recurrence, prognostic factors included positive nodes, aneuploidy and poorly differentiated histologic grading. CONCLUSIONS: We conclude that a combination of lymph node status, DNA ploidy, histologic grading and progesterone-receptor status help to evaluate the possible outcomes for patients with breast cancer and to plan for optimal therapy. PMID- 10533303 TI - Ethambutol-induced psychosis: a case report. AB - Clinically, ethambutol (EMB)-induced psychosis is rare. In our review of the literature, most cases of antituberculosis agent-associated psychoses were caused by isoniazid (INH). We report the case of a 51-year-old man with suspected tuberculosis (TB) pleurisy. An anti-TB trial with INH, rifampicin and EMB was given initially. Dizziness, disorientation, and auditory and visual hallucinations developed after seven days of therapy. Laboratory examinations, including routine biochemistry tests, serum titer of antinuclear antibodies, cerebrospinal fluid analysis and computerized tomography of the head showed no abnormal findings. Following discontinuation of anti-TB agents, the psychiatric symptoms subsided. When the patient was challenged with EMB, the same psychiatric symptoms recurred, but resolved again after discontinuation of EMB. It is important to be aware that EMB can induce psychosis when anti-TB medications are prescribed. PMID- 10533304 TI - Accidentally delayed diagnosis of ruptured ovarian carcinoma in a young woman: a care report. AB - Ovarian carcinoma commonly occurs in postmenopausal women and often presents with an insidious course. Acute abdomen is rarely an initial symptom. When these patients present with abdominal discomfort, the disease has already spread throughout the peritoneal cavity. We present a case of mucinous cystadenocarcinoma in a young woman who presented with acute abdomen and intra abdominal bleeding. This 24-year-old woman was previously diagnosed with a ruptured left ovarian cystic tumor at a primary clinic. She underwent emergency exploratory laparotomy, followed by unilateral salpingo-oophorectomy at the clinic. No thorough examination of the peritoneal cavity was done during surgery. The diagnosis of mucinous cystadenocarcinoma was accidentally over-looked until one month later when she returned for routine follow-up. Upon referral to our clinic, the patient underwent a repeat laparotomy. The surgicopathologic diagnosis was intraperitoneal carcinomatosis stage IIIC that could not be excised completely, even though rigorous staging surgery including washing cytology, total abdominal hysterectomy, salpingo-oophorectomy, retroperitoneal lymphadenectomy, appendectomy, infracolic omentectomy and excision of any suspicious and removable lesions were performed. This case alerts us to consider the possibility of ovarian malignancy when a young woman presents with an acute abdomen secondary to ruptured ovarian cystic tumor and intraperitoneal hemorrhage. Careful preoperative preparation and thorough intrasurgical examination of the peritoneal cavity along with a prompt pathologic diagnosis of suspicious lesions will prevent missed diagnoses. PMID- 10533305 TI - Percutaneous transluminal coronary angioplasty for renovascular hypertension in a child: a case report. AB - Since its introduction, percutaneous transluminal coronary angioplasty has become an alternative therapeutic modality to surgical and medical treatment for renovascular hypertension. We report the case of a nine-year-old boy who had hypertension caused by renal arterial stenosis. The patient's high blood pressure was 164/100 mmHg, which was discovered incidentally during a physical check-up. A selective renal angiography showed a severe short-segment stenosis with post stenotic dilatation of the left renal artery. A 4-mm balloon catheter was advanced through the stenotic area and was inflated five times to dilate the stenosis. After the procedure, the selective renal angiography showed a significant increase in the diameter of the left renal artery. Blood pressure decreased to normal immediately after the procedure. During the one-year follow up period, the patient remained normotensive without the use of antihypertensive drugs. PMID- 10533308 TI - Signal transduction and the coordination of B lymphocyte development and function I. Transduction of BCR signals from the cell membrane to the nucleus. Introduction PMID- 10533306 TI - Giant cell reparative granuloma: a case report. AB - Giant cell reparative granuloma (GCRG) is an infrequent benign lesion with undetermined etiopathogenesis affecting the maxillary and mandibular bone and, rarely, the skull. It is also extremely rare in the sphenoid bone. GCRG is usually diagnosed by histologic examination of bone lesions. We report a case of GCRG originating from the sphenoid bone. Computerized tomography revealed an expansile lesion with thinning or destruction of the cortical bone. The lesion itself was slightly hyperdense with good but inhomogeneous contrast enhancement. Reported magnetic resonance image findings showed hyperintensity on both T1 weighted and T2-weighted images and variable contrast enhancement. Plain skull radiographs usually reveal a lytic lesion within the bone. PMID- 10533307 TI - Glycogen storage disease type IV: a case report. AB - Glycogen storage disease type IV (GSD-IV) is a rare autosomal recessive disease caused by a deficiency of glycogen branching enzyme (GBE) activity. This results in the accumulation of abnormal glycogen in the liver and other organs. We report the case of a 14-month-old female patient with typical hepatic pathologic findings of GSD-IV. The patient suffered from decreased muscle tone and progressive hepatosplenomegaly since birth. A wedge biopsy of the liver showed enlarged hepatocytes with colorless to faintly eosinophilic ground glass intracytoplasmic inclusions. Portal fibrosis and lobular, fibrous septa were present. Ultrastructure of the inclusions revealed non-membrane-bound fibrillar material 5 nm in maximal diameter. Enzyme study revealed a total deficiency of GBE activity. PMID- 10533310 TI - The B-cell antigen receptor: formation of signaling complexes and the function of adaptor proteins. PMID- 10533312 TI - Involvement of the lymphocyte cytoskeleton in antigen-receptor signaling. PMID- 10533311 TI - Intermediary signaling effectors coupling the B-cell receptor to the nucleus. PMID- 10533313 TI - Pax-5/BSAP: regulator of specific gene expression and differentiation in B lymphocytes. PMID- 10533309 TI - Signal transduction via the B-cell antigen receptor: the role of protein tyrosine kinases and protein tyrosine phosphatases. PMID- 10533314 TI - Receptor modulators of B-cell receptor signalling--CD19/CD22. PMID- 10533317 TI - Signaling pathways that control V(D)J recombination. PMID- 10533318 TI - B-cell-receptor-dependent positive and negative selection in immature B cells. AB - This review touches on only a small part of the complex biology of B cells, but serves to illustrate the point that the antigen receptor is the most important of many cell-surface receptors affecting cell-fate decisions. Receptor expression is necessary, but not sufficient, for cell survival. It is also essential that a B cell's antigen-receptor specificity be appropriate for its environment. The need to balance reactivity with self tolerance has resulted in an intricate feedback control (affected by both the recombinase and cell survival) that regulates independent selection events at the level of the receptor and the cell. PMID- 10533316 TI - B-cell antigen receptor signaling in immature-stage B cells: integrating intrinsic and extrinsic signals. PMID- 10533321 TI - Molecular processes that regulate class switching. PMID- 10533315 TI - Positive and negative signaling in B lymphocytes. PMID- 10533324 TI - Uterine leiomyoma in pregnancy: its influence on obstetric performance. AB - OBJECTIVE: To assess the effects of uterine leiomyoma on obstetrical performance. METHODS: We reviewed the medical records of 102 women with singleton pregnancies who were found ultrasonographically to have uterine leiomyomas during the first half of their pregnancy and who gave birth at our hospital at > or = 22 weeks of gestation between January 1990 and December 1997. RESULTS: The 102 women gave birth to 101 healthy infants, weighing 2,974 +/- 579 g at 38.8 +/- 2.6 weeks of pregnancy. One woman experienced an unexplained antepartum fetal death at 24 weeks of gestation. Bleeding at the first trimester occurred in 16% of the women. Pain localized in the lower abdomen and requiring relief occurred in 28% of the women during the first or second trimester. Tocolytic treatment was required in 25% of the pregnancies, and preterm delivery occurred in 12% thereof. A cesarean section was performed in 39% of the pregnancies. Bleeding > or = 500 ml occurred at delivery in 48% of the cases. The largest fibroid, > 6 cm in diameter, which was seen in 51 women, was associated with higher frequencies of tocolytic treatment (41%), preterm delivery (24%), bleeding > or = 500 ml at delivery (59%), and cesarean delivery (51%). In 76 women (75%) who attempted vaginal delivery, the obstetrical outcome was comparable to that of 115 control women who were matched regarding age, parity, and gestational week. CONCLUSIONS: Although pain in the lower abdomen, the requirement of tocolytic treatment, preterm delivery, and cesarean delivery were common, the neonatal outcome was fairly good in women with uterine leiomyomas. The present data might be encouraging to pregnant women with uterine leiomyomas. PMID- 10533319 TI - CD40-CD154 interactions in B-cell signaling. PMID- 10533323 TI - Placenta previa increta penetrating the entire thickness of the uterine myometrium: ultrasonographic and magnetic resonance imaging findings. AB - This is the first report of placenta previa increta in which the placenta villi penetrated the entire thickness of the uterine myometrium, but did not invade the pubocervical fascia. Ultrasonographic and magnetic resonance imaging findings are described. PMID- 10533322 TI - Low-dose GnRH agonist therapy for the management of endometriosis. AB - OBJECTIVE: In order to examine whether treatment with a GnRH agonist alone can maintain estrogen levels within the "estrogen window" that inhibits endometriosis without influencing bone-mineral density, we studied the effects of GnRH agonist therapy and changes in bone-mineral density. METHODS: Buserelin acetate nasal spray was administered 3 times a day for 8 weeks (daily dose, 900 micrograms) to 21 women with endometriosis. The drug was then given twice a day for 16 weeks (daily dose, 600 micrograms). The total duration of treatment was 24 weeks. The bone-mineral density of the lumbar vertebrae was measured by dual-energy X-ray absorptiometry before treatment (baseline), at the end of treatment, and 24 weeks after the end of treatment. RESULTS: The bone-mineral density of the lumbar vertebrae at the end of treatment was 2.44% +/- 0.46% (mean +/- standard error) lower than the baseline value. The value at 24 weeks after the end of treatment was 1.10% +/- 0.64% lower than the baseline value. More than 80% of the patients had serum-estradiol levels of 45 pg/ml or less. During treatment, more than 90% of the patients had serum-estradiol levels of 60 pg/ml or less. Genital bleeding was inhibited in 90% of the patients. After 8 weeks of treatment, the clinical symptoms improved in 75% of the patients; such improvement persisted for the duration of the treatment. CONCLUSION: Decreasing the dose of GnRH agonist during treatment can minimize the loss of bone-mineral density without lessening the beneficial effects on endometriosis. This technique might be useful in the management of endometriosis. PMID- 10533329 TI - Prenatal diagnosis of Meckel syndrome: a case report. AB - OBJECTIVE: To demonstrate the major sonographic findings associated with Meckel syndrome and to emphasize the importance of prenatal sonography in helping to establish the correct diagnosis. SUBJECTS: Two fetuses with prenatal diagnosis of Meckel syndrome were sonographically evaluated. RESULTS: Both fetuses were demonstrated to have evidence of renal cystic dysplasia, occipital cephalocele and postaxial polydactyly. One case was diagnosed at 16 weeks of gestation whereas the other was detected at 36 weeks. Of interest, the first case had only unilateral renal cystic dysplasia and contralateral renal agenesis and mild degree of oligohydramnios. The other related anomalies which were not detected prenatally included cerebellar hypoplasia in case 1 and micrognathia in case 2. CONCLUSION: The main sonographic findings included renal cystic dysplasia, occipital cephalocele and postaxial polydactyly. PMID- 10533325 TI - Recent management of malignant ovarian germ cell tumors: a study of 34 cases. AB - OBJECTIVE: To review the outcome of the treatment in patients with malignant ovarian Germ cell tumors with respect to survival and surgical management at a single institution during 1990-1996. METHODS: Thirty-four patients with malignant ovarian Germ cell tumors were studied retrospectively for their surgical management. Fourteen patients had pure dysgerminoma, 11 endodermal sinus tumor, 6 immature teratoma, and 3 mixed Germ cell tumors. Nine patients had stage IA, 8 stage IC, 2 stage IIC, 8 stage III, 3 stage IV, and 4 referred patients with recurrent diseases. RESULTS: Nineteen patients underwent primary conservative surgery, 11 had primary nonconservative surgery. Twenty-two patients were treated with chemotherapy (BEP or EP or PVB regimen). Five patients with pure dysgerminoma received adjuvant radiotherapy. Persistent remission was achieved in 26 patients. Two patients (7.4%) had recurrence after remission. Seven patients had died of the diseases. Patients with complete clinical remission did not undergo second-look surgery. The overall survival was 78.8%, 100% for immature teratoma, 84.6% for pure dysgerminoma, 72.8% for endodermal sinus tumor, and 33.3% for mixed Germ cell tumors, with median follow-up time 31 (3-93) months. CONCLUSION: Patients with limited diseases regardless of histologic types can be safely managed by unilateral salpingo-oophorectomy followed by, if indicated, 3-4 courses of cisplatin-based chemotherapy. For advanced diseases, conservative surgery is advisable in patients with endodermal sinus tumor. PMID- 10533328 TI - Efficacy of magnesium sulphate and phenytoin in the management of eclampsia. AB - Fifty pregnant women admitted with diagnosis of eclampsia were randomly allocated to magnesium sulphate (Group A) or phenytoin sodium (Group B) treatment group. Incidence of recurrence of seizures maternal as well as perinatal morbidity and mortality were compared in both the groups. Mean maternal age, parity and gestational age was similar in both the groups. Mean birth weight was significantly lower in Group B compared to Group A. Seizure frequency prior to hospitalization was 5.4 +/- 4.7 in Group A and 4.8-3.6 in Group B. Mean time interval between occurrence of first seizure and hospitalization was 9.6 +/- 3.5 hours in Group A and 11.8 +/- 9.3 hours in Group B, the difference was not statistically significant. Women treated with phenytoin had a higher incidence of recurrent seizures (10/25-40%) than those treated with magnesium sulphate (2/25 8%). Majority of the women treated with phenytoin (6/10-60%) had single convulsion after initiation of anticonvulsant therapy and 1 woman of each group had recurrent convulsions (75). There was no significant difference in perinatal outcome in both the groups. Maternal morbidity was comparable in both the groups and there was no maternal death in either of the groups. PMID- 10533327 TI - The use of rectal misoprostol as active pharmacological management of the third stage of labor. AB - OBJECTIVE: To compare the effectiveness of rectal Misoprostol versus combined intramuscular oxytocin and ergometrine (O-E) in the management of the third stage of labor. METHODS: Low-risk women in 3rd stage of labor were allocated to receive either rectal Misoprostol [200micrograms (n = 25), 400 micrograms (n = 45)] or 5 units oxytocin and 0.2 mg ergometrine intramuscularly (n = 75). Clinical and hematological parameters were compared using t and chi-square tests. RESULTS: Both groups were well matched and had similar duration of the 3rd-stage of labor. Misoprostol users had lower 3rd-stage estimated blood loss and needed less further ecbolics compared to O-E group. Postpartum Hb and Hct levels were significantly lower in O-E group than Misoprostol group. Postpartum hypertension occurred more in O-E group. Subjects in Misoprostol group had more shivering. Subjects receiving 200 micrograms and 400 microgram Misoprostol had similar outcome variables. CONCLUSION: Rectal Misoprostol may be used safely in the management of the third stage of labor. PMID- 10533326 TI - A case of primary transitional cell carcinoma of the fallopian tube. AB - The primary carcinoma of the fallopian tube is the rarest of all gynecologic malignancies and histologically most of them are adenocarcinomas. Primary transitional cell carcinomas are extremely rare in the fallopian tube. A 63-year old postmenopausal woman presenting with lower abdominal pain was found to have a left adnexal mass. Exploratory laparotomy revealed a mass arising from the fallopian tube with the histologic features of transitional cell carcinoma. Light and electron microscopic studies supported the notion of transitional cell carcinoma. The tumor was extended to the muscle layer and confined to the left fallopian tube without metastasis. The patient received 3 courses of systemic cisplatin-based chemotherapy and has been well with no evidence of recurrence until August, 1998. PMID- 10533330 TI - Hyperbaric oxygenation for rectovaginal fistula: a report of two cases. AB - A rectovaginal fistula after delivery is a rare complication, and its management can become difficult if infection occurs. In two such cases, we administered hyperbaric oxygenation (HBO) treatment against complicated infections, and we obtained a good outcome in each case. PMID- 10533320 TI - B-lymphocyte signaling receptors and the control of class-II antigen processing. PMID- 10533331 TI - Immunohistochemical studies concerning cathepsin D in endometrial carcinomas. AB - OBJECTIVES: To study the prognostic value of immunohistochemical detection of cathepsin D and laminin in endometrial carcinomas. METHODS: Immunohistochemical staining of cathepsin D was performed on paraffin sections of 111 endometrial carcinomas, and laminin deposition was studied in 65 endometrial carcinomas. RESULTS: Of 111 tissue specimens, 55 showed a positive reaction for cathepsin D. The incidence of cathepsin D-positive staining increased with the extension of the primary tumor (p < 0.01). Patients with vessel invasion and pelvic lymph-node metastasis had a higher incidence of cathepsin D-positive staining than patients without these findings (p < 0.05). A favorable prognosis was obtained in a negative case of cathepsin D in comparison with a positive case (p < 0.01). Of 9 patients in whom laminin was detected in the cytoplasm of cancer cells, 5 showed poor prognoses and died from a primary disease. CONCLUSION: Cathepsin D and laminin status might represent possible prognostic factors. PMID- 10533333 TI - Effects of low oxygen condition on the generation of reactive oxygen species and the development in mouse embryos cultured in vitro. AB - OBJECTIVE(S): To elucidate the relationship between intracellular H2O2 production and embryo development in different oxygen culture conditions. METHODS: Pronuclear stage embryos were obtained from C57BL/CBA F1 and ICR mice. Measurement of H2O2 level was performed with 2',7'-dichlorodihydroflourescein diacetate and the number of blastomeres was counted after staining with 4',6' diamidino-2-phenylindole. RESULTS: Regardless of strains, H2O2 level reached a peak at the 2-cell stage in 20% O2. But in embryos cultured in 5% O2 it was significantly lower at the 2-cell and 4-cell stages compared to those from 20% O2. The embryos cultured in 20% O2 showed developmental delay or block, but in 5% O2 these phenomena were overcome and the development was significantly increased with an infrequent fragmentation. CONCLUSIONS: Our data suggest that the 5% O2 decreases the relative concentration of H2O2 and results in improved embryo development in terms of quantity and quality without regard to type of strains. PMID- 10533336 TI - Pharmacogenomics: tailoring drug therapy. PMID- 10533337 TI - Utah school offers new insights into chemical dependency. PMID- 10533335 TI - Who is managing our services? PMID- 10533341 TI - Meaningful managed care reform: still elusive, but not out of sight. PMID- 10533340 TI - Carpe datum ... carpe diem. PMID- 10533342 TI - Pharmacist intervention to control lice resistance. PMID- 10533343 TI - Pharmacist counseling on nutrition and physical activity--Part 2 of 2: Helping patients make changes. PMID- 10533346 TI - Washington State CARE Project: downstream cost changes associated with the provision of cognitive services by pharmacists. AB - OBJECTIVE: To determine the changes in drug costs associated with drug therapy changes resulting from pharmacists' cognitive services (CS) provided to Medicaid recipients during a 1-year period following the documented CS. DESIGN: A study control group analysis of documented pharmacists' CS interventions linked to Medicaid prescription claims. Each CS resulting in a drug therapy change was linked to an index prescription claim and all refills for the same drug within 365 days. The drug cost change associated with the CS was calculated as the difference between the estimated cost of the prescription as originally written less the actual cost to Medicaid for the stream of refills dispensed. SETTING: Pharmacies serving ambulatory Medicaid patients in the state of Washington, excluding staff-model health maintenance organization pharmacies and pharmacies predominantly serving long-term care residents. PARTICIPANTS: Approximately 200 community pharmacies participating in the Washington State Cognitive Activities and Reimbursement Effectiveness (CARE) Project. Pharmacies were randomly assigned to a group that was paid a fee for each CS provided or a group that was not paid. INTERVENTION: Payment for CS. MAIN OUTCOME MEASURES: Downstream drug costs associated with CS resulting in a drug therapy change. RESULTS: CS resulting in drug therapy changes accounted for 5,417 out of 20,240 (27%) documented CS in the CARE Project. Of the 2,002 CS records analyzed in this study, 76% indicated a change in the prescribed drug or drug regimen, 9% indicated that a drug was added, 5% indicated that a current drug was discontinued, and 10% indicated that an originally prescribed drug was never dispensed. Only 9% involved generic substitution; all other changes would have necessitated prior prescriber approval. Overall, CS resulting in a drug therapy change generated a mean drug cost savings of $13.05 per CS intervention. There were no significant differences in average savings per intervention between the paid and nonpaid groups. CONCLUSION: For all result categories except "add drug therapy," the extrapolated cost savings in the paid group exceeded the savings estimated from the nonpaid group, sometimes by a considerable amount. At the payment rate used in this study, paying for CS that result in a drug therapy change (except add drug therapy) is estimated to save an additional $10 per 1,000 prescriptions dispensed. Those CS that result in addition of drug therapy are estimated to add an incremental cost of about $13 per 1,000 prescriptions. A sensitivity analysis revealed that a higher intervention rate would lead to a higher potential savings. This finding suggests that efforts to encourage CS interventions may lead to greater savings. PMID- 10533332 TI - Effect of methamphetamine on male mice fertility. AB - The effect of methamphetamine (MAMP) on the ability of males to mate with females and to impregnate them was examined in 8-week-old ICR mice. Male mice that had been administered an intraperitoneal injection of MAMP (15 mg/kg, 7.5 mg/kg, or 3.75 mg/kg) or saline were housed with females 24 or 48 hours later. The vaginal plugs were checked, and the number of births was counted. The effect of MAMP on sperm motility and the serum testosterone (TS) concentration was also examined. Methamphetamine was observed to have harmful effects only at the highest dose level. In the mice housed 24 hours after the injection of 15 mg/kg MAMP, the increase in the cumulative number of vaginal plugs lagged, and the total number of vaginal plugs decreased significantly. A significant decrease was also observed in the total number of births. Methamphetamine, at a dose of 15 mg/kg, decreased sperm motility. The TS concentration decreased initially, then increased. PMID- 10533334 TI - The contribution of menopause to changes in body-fat distribution. AB - OBJECTIVE: To investigate whether menopause contributes to changes in body-fat distribution, irrespective of aging or obesity. METHODS: The subjects were 545 premenopausal (aged 16-55 years; mean +/- standard deviation, 37.7 +/- 9.1 years) and 219 postmenopausal (aged 45-65 years, 58.0 +/- 5.0 years) women. Baseline characteristics included age, body mass index (BMI), and menopausal status (premenopause or postmenopause). The ratio of trunk fat to leg fat (trunk-leg ratio) was estimated by dual-energy X-ray absorptiometry. The trunk-leg ratio and baseline characteristics were compared between the 2 groups. In all subjects (n = 764), possible correlations between the trunk-leg ratio and the baseline characteristics were determined using univariate and multivariate analysis. In postmenopausal women, the relationship of the trunk-leg ratio to YSM or age after adjusting for BMI was investigated. RESULTS: The trunk-leg ratio and BMI were significantly higher in postmenopausal women than in premenopausal women. In all subjects, age and BMI were positively correlated with the trunk-leg ratio (r = 0.445 and 0.587, respectively, p < 0.0001). Menopause was also positively correlated with the trunk-leg ratio on univariate regression analysis (standardized regression coefficient = 0.369, p < 0.0001). On multiple regression analysis, age, BMI, and menopause were independently correlated with the trunk leg ratio (p < 0.05). In postmenopausal women, age and YSM were positively correlated with the trunk-leg ratio, independent of the BMI (p < 0.01). CONCLUSIONS: Menopause contributes to a change in body-fat distribution, irrespective of aging or obesity. PMID- 10533339 TI - Lessons learned from the Washington State CARE Project. PMID- 10533344 TI - Influence of a financial incentive on cognitive services: CARE project design/implementation. AB - OBJECTIVE: To describe the design and methods of the Washington State Cognitive Activities and Reimbursement Effectiveness (CARE) Project, a demonstration project in which community pharmacies were paid for cognitive services (CS) provided to Medicaid patients, its evaluation objectives, and the extent to which implementation objectives were achieved. DESIGN: Prospective randomized trial. Community pharmacies were allocated to a documentation-and-payment group, documentation-only group, and "silent" control group. CS were reported using a problem-intervention-result classification system embedded within a pseudo National Drug Code format. Management strategies included use of area coordinators. SETTING: Pharmacies serving ambulatory Medicaid patients in the state of Washington, excluding staff-model health maintenance organization pharmacies and pharmacies predominantly serving long-term-care residents. PARTICIPANTS: 200 community pharmacies (110 treatment; 90 control), with another 100 randomly selected pharmacies as a silent control group. INTERVENTIONS: A modest monthly stipend. The treatment group billed Medicaid for each documented CS associated with a drug therapy-related problem. All participants received training in documentation methods. A unique coding scheme allowed documentation of CS within the constraints of the Medicaid program. Data edit checks and feedback were used to ensure data quality and completeness. Area coordinators were used to facilitate training, compliance with study procedures, and participation. MAIN OUTCOME MEASURES: Participation rates, documentation rates, coding scheme revision, data quality and completeness rates, and effectiveness of area coordinators. RESULTS: Pharmacists documented more than 20,240 CS records. Approximately 89% of records passed edit checks, and 94% did so after modification. Nearly 83% could be linked to a paid drug or CS claim. The coding system was sufficient, with minor modifications, to account for all interventions documented. Area coordinators did not function as expected. CONCLUSION: A system for documentation and payment of pharmacists' CS to Medicaid recipients was implemented successfully and relatively easily in community pharmacies. PMID- 10533347 TI - Patients' perceived benefit from and satisfaction with asthma-related pharmacy services. AB - OBJECTIVE: To determine whether patients targeted to receive intervention from an asthma management program reported receiving more services and had greater perceived benefit and satisfaction with those services compared with asthma patients not targeted by the program. DESIGN: Mailed survey. SETTING: Community pharmacy. PATIENTS: 471 community-based patients receiving asthma medications from 44 intervention pharmacies and 1,164 patients from 46 usual care (control) pharmacies. MAIN OUTCOME MEASURES: Five-point agreement scale measuring asthma services received, perceived value of the services, and satisfaction. RESULTS: Usable surveys were received from 39.0% of intervention patients and 42.4% of controls. There were no statistically significant differences between groups in the frequency of provision of listed services. Approximately 60% of respondents from both groups received written materials on asthma medications and 54% received inhaler counseling; both were rated high for perceived benefit. Fewer than 20% reported being counseled about asthma triggers. Fewer than 5% reported pharmacists talking to physicians on their behalf. General satisfaction with pharmacy services was high (78.2% agree or strongly agree), but not statistically different between groups. More than 65% believed that pharmacists spend enough time counseling patients. Several comments indicated that patients did not expect or ask for information because they were unaware that services were available and/or they had already been counseled by their physician. Responses to the statement "my asthma is better controlled because of help given to me by the pharmacist" were equivocal and not different between groups. CONCLUSION: Overall, there were few differences between groups. General satisfaction with pharmacy services is high, but patients' perceived benefit and satisfaction with cognitive services is lower. Increased public awareness of pharmacists' capabilities and a more proactive approach to providing cognitive services is needed. PMID- 10533338 TI - The Utah School experience. PMID- 10533351 TI - Management of insomnia. AB - OBJECTIVE: To review current issues in the pharmacologic and nonpharmacologic management of insomnia. DATA SOURCES: Controlled trials and case studies identified via MEDLINE for 1990 through April 1999 under the search terms insomnia, hypnotics, flurazepam, quazepam, estazolam, temazepam, triazolam, zolpidem, zaleplon, L-846, CL-284,846, melatonin, and valerian. DATA SYNTHESIS: Insomnia is a common, undertreated disorder. Nonpharmacologic management strategies (e.g., stimulus control, relaxation therapy, sleep hygiene) are therapeutic options that, compared with medication use, provide more sustained effects. The benzodiazepines and zolpidem are the most commonly prescribed hypnotic agents, but their use is associated with tolerance and central nervous system adverse effects. A new nonbenzodiazepine hypnotic agent, zaleplon, was very recently approved in the United States. Because of its short half-life, zaleplon will be useful in patients experiencing difficulty in falling asleep and in those who wake up at night and have trouble falling back to sleep. Antidepressants, antihistamines, and alternative medications are other treatment options. To avoid complications of therapy, hypnotic agents should be used at their lowest possible doses and for limited treatment durations. CONCLUSION: Pharmacotherapy is currently the most common treatment modality for insomnia, but long-term use of hypnotic agents can become complicated by drug tolerance, dependence, or rebound insomnia. Nonpharmacologic options--including combinations of behavioral interventions, sleep-restriction therapy, and patient education- provide longer-lasting benefits. PMID- 10533350 TI - Evaluation of consumer drug information databases. AB - OBJECTIVES: To evaluate prescription drug information contained in six consumer drug information databases available on CD-ROM, and to make health care professionals aware of the information provided, so that they may appropriately recommend these databases for use by their patients. DESIGN: Observational study of six consumer drug information databases: The Corner Drug Store, Home Medical Advisor, Mayo Clinic Family Pharmacist, Medical Drug Reference, Mosby's Medical Encyclopedia, and PharmAssist. SETTING: Not applicable. PATIENTS OR OTHER PARTICIPANTS: Not applicable. INTERVENTIONS: Information on 20 frequently prescribed drugs was evaluated in each database. The databases were ranked using a point-scale system based on primary and secondary assessment criteria. MAIN OUTCOME MEASURES: For the primary assessment, 20 categories of information based on those included in the 1998 edition of the USP DI Volume II, Advice for the Patient: Drug Information in Lay Language were evaluated for each of the 20 drugs, and each database could earn up to 400 points (for example, 1 point was awarded if the database mentioned a drug's mechanism of action). For the secondary assessment, the inclusion of 8 additional features that could enhance the utility of the databases was evaluated (for example, 1 point was awarded if the database contained a picture of the drug), and each database could earn up to 8 points. RESULTS: The results of the primary and secondary assessments, listed in order of highest to lowest number of points earned, are as follows: Primary assessment--Mayo Clinic Family Pharmacist (379), Medical Drug Reference (251), PharmAssist (176), Home Medical Advisor (113.5), The Corner Drug Store (98), and Mosby's Medical Encyclopedia (18.5); secondary assessment--The Mayo Clinic Family Pharmacist (8), The Corner Drug Store (5), Mosby's Medical Encyclopedia (5), Home Medical Advisor (4), Medical Drug Reference (4), and PharmAssist (3). CONCLUSION: The Mayo Clinic Family Pharmacist was the most accurate and complete source of prescription drug information based on the USP DI Volume II and would be an appropriate database for health care professionals to recommend to patients. PMID- 10533349 TI - Frequency and severity of sexual harassment in pharmacy practice in Ohio. AB - OBJECTIVE: To determine the frequency and severity of sexual harassment in the pharmacy workplace for both male and female pharmacists, and to identify: (1) instigators, (2) places of occurrence, and (3) pharmacists' responses. DESIGN: Mailed survey using elements of the Sexual Experience Questionnaire (SEQ). One repeat mailing to nonrespondents. SETTING: Community pharmacies, hospital pharmacies, other pharmacies in the state of Ohio. PATIENTS AND OTHER PARTICIPANTS: 789 randomly selected pharmacists registered in Ohio. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Amount of gender harassment, unwanted sexual attention, and sexual coercion; differences in occurrences of sexual harassment between men and women; identification of instigators as colleagues, patients, or supervisors; identification of place of occurrence as community pharmacy, hospital pharmacy, or elsewhere; pharmacists' responses and reactions. RESULTS: After two mailings, 265 usable surveys were returned for a response rate of 34%. Women differed significantly from men in total occurrences of sexual harassment, with men reporting 183 instances of sexual harassment and women reporting 281 such experiences. Instigators were colleagues (43%), patients (30%), and superiors (27%). Men reported 143 experiences of unwanted sexual attention, whereas women reported 272 such occurrences. Colleagues were responsible for 47% of instances of unwanted sexual attention, patients were responsible for 37%, and superiors 16%. No significant differences were found between men and women in total number of occurrences of sexual coercion. CONCLUSION: Sexual harassment in the workplace has been experienced by both male and female pharmacists. Women experienced more hostile work environment harassment than did men. However, quid pro quo harassment did not differ significantly between the sexes. PMID- 10533345 TI - Characteristics of pharmacies and pharmacists associated with the provision of cognitive services in the community setting. AB - OBJECTIVE: To determine the influence of payment, pharmacy setting characteristics, pharmacist demographics, practice setting, and attitudinal characteristics on whether cognitive services (CS) were performed by pharmacists, and the volume of CS performed. DESIGN: Prospective randomized trial. Community pharmacies were randomized to a documentation-and-payment group (study group) and a documentation-only group (control). Participating pharmacies and pharmacists were surveyed by mail, and responses were linked to a documented CS database. SETTING: Community pharmacies serving ambulatory Medicaid patients in the state of Washington, excluding health maintenance organization pharmacies and pharmacies predominantly serving long-term care residents. PARTICIPANTS: 200 community pharmacies and their pharmacists (110 study, 90 control) participating in the Washington State Cognitive Activities and Reimbursement Effectiveness (CARE) Project. INTERVENTION: Payment for CS. All participants documented CS. Study group pharmacies billed Medicaid for services performed in identifying and resolving drug therapy-related problems. MAIN OUTCOME MEASURE: Documentation of CS. RESULTS: Documentation of CS was more likely if the pharmacist was an owner or manager, if documentation was not perceived as burdensome, and if the pharmacy had a low ratio of prescription to total sales. Higher documentation rates were associated with study group status, lower pharmacy prescription volume as a percentage of total sales, and a higher percentage of prescriptions billed to Medicaid. Among pharmacists, two setting variables--medical center location and rural location--were associated with higher documentation rates. CONCLUSION: Performance of CS was strongly affected by payment and other situational factors, including practice setting and volume of prescriptions dispensed. Pharmacies and pharmacists were also more likely to perform CS if the target patient population represented a relatively large percentage of that pharmacy's patient clientele. PMID- 10533353 TI - Pharmaceutical care interventions--a closer look. PMID- 10533348 TI - Pharmacists' attitudes regarding quality of worklife. AB - OBJECTIVE: To examine the quality of worklife of pharmacists across practice environments. DESIGN: Written survey mailed to a stratified random sample of pharmacists. PARTICIPANTS: 2,014 pharmacist-members of the American Pharmaceutical Association (APhA) residing in the United States. MAIN OUTCOME MEASURES: Work-related attitudes including job satisfaction, career satisfaction, organizational commitment, turnover intention, likelihood of voting for a union, and patient care issues. RESULTS: Usable surveys were returned by 1,199 practicing APhA members (60% response rate). Overall, work-related attitudes were generally positive. However, clear differences were identified in quality-of worklife perceptions associated with practice setting, area of primary responsibility, and several demographic variables. Some quality-of-worklife concerns were found in all practice settings. While a majority of respondents indicated that they would not vote for a union at their place of employment if given the opportunity, 43% of staff employee pharmacists (who would likely be targeted in any union campaign) indicated that they would definitely or probably vote for a union. CONCLUSION: From the pharmacist's perspective, important quality-of-worklife issues include job and career satisfaction, turnover intention, and patient care concerns. The data provide a point of departure for future dialogue, action, and research aimed at understanding and enhancing the quality of pharmacists' worklife. PMID- 10533354 TI - Oprelvekin (Neumega), first platelet growth factor for thrombocytopenia. AB - Oprelvekin decreases the need for platelet transfusion in nontransplant patients receiving myelosuppressive chemotherapy. Until pharmacoeconomics studies determine the most cost-effective strategy for use of oprelvekin, it is likely to be used primarily in patients receiving dose-intensive chemotherapy to maintain the high-dose regimen that may provide a survival advantage for the patient with cancer. Additional product information is available at www.ahp.com/products/neumega.htm. PMID- 10533355 TI - Low immunization rates in ethnic minorities--a problem pharmacists can help solve. PMID- 10533360 TI - Serum and the Soluvac: the Australian approach to whole blood substitutes and blood transfusion during the Second World War. PMID- 10533356 TI - Computerized POE: changing roles for the clinical pharmacist. PMID- 10533352 TI - Pharmacy service alliances: a tool to reduce uncertainty and create new revenue streams. AB - OBJECTIVES: (1) Present conceptual support for a type of pharmacy network, a pharmacy service alliance (PSA), (2) describe the development of a PSA in eastern Iowa, and (3) discuss how other types of PSAs can be developed. DESIGN AND PARTICIPANTS: 12 independent pharmacies in eastern Iowa. SETTING: Community pharmacy practice. MAIN OUTCOME MEASURES: Formation of a PSA. RESULTS: Pharmacy members of the Certified Pharmaceutical Care Network, a PSA, have jointly developed new pharmacy services. Collaborative efforts have involved disease state management programs, group marketing activities, and a quality improvement process. CONCLUSION: PSAs offer an organizational model that pharmacies could use to successfully develop new pharmacy services. PMID- 10533359 TI - Filtering the city's image: progressivism, local control, and the St. Louis water supply, 1890-1906. PMID- 10533358 TI - Syphilization: human experimentation in the search for a syphilis vaccine in the nineteenth century. PMID- 10533362 TI - [Recurrent profuse hemorrhage in gastric angiodysplasia]. AB - Dieulafoy's disease--i.e. arteriovenous malformation--is a rare cause of gastric bleedings, which makes up 1.5-3% of all sources of hemorrhage. All the armory of diagnostic modalities may require to reveal arteriovenous malformations. Angiography of the stomach appeared to be the most informative method for diagnosis of this disease in 2 patients. Radical treatment of patients with Dieulafoy's disease implies resection of the proximal part of the stomach or gastrectomy. Endoscopic hemostasis (with the use of coagulation and injections of sclerogenous preparations around the diseased area) results in temporary effect. PMID- 10533357 TI - "A kindly, useful mentor": applying the history of medicine to public policy. PMID- 10533361 TI - Autobiographical letter from Horatio Robinson Storer, M.D., to his son, Malcolm Storer, M.D., discussing the "History of gynaecological teaching". PMID- 10533363 TI - [Triangular anastomoses in surgery of the stomach]. AB - The mode of triangulation in the anastomoses of the stomach and the duodenum (thrice-repeated usage of linear suturing apparatuses in shaping anastomoses) is a perspective way for improvement of the results of surgical treatment of these organs. The results of 216 cases have shown that the use of the triangular anastomoses in gastric surgery allows a decrease in the rate of complications because of anastomosis failure in early postop period more than 3.5 times. The healing of such anastomoses proceeds by primary intention and in short terms and is accompanied by minimal inflammatory reaction. The course of restoration of the motor and evacuation function of the stomach after creation of such anastomoses is characterized by early, partial and timely evacuation. In remote postoperative period mechanical everted anastomoses, made up by linear suturing apparatuses, provide absence of the tendency for cicatricial strictures and stipulation of motor and evacuation functional characteristics of the stomach. The authors believe that beneficial impact of this method on gastric function allows to recommend it for wider use in surgery of the stomach. PMID- 10533366 TI - [Treatment and prevention of postoperative eventration]. AB - The proposed mode for the treatment of eventrations is based on the leading of Kischner needles through the rectus abdominis muscles vaginas and is successfully used in the treatment for 7 patients. This mode if indicated may be applied for prophylaxis of eventrations. It has some advantages compared to other methods. PMID- 10533367 TI - [Long-term results of balloon dilatation in the treatment of esophageal achalasia]. AB - 25 patients (aged from 32 to 58 years) with achalasia of the esophagus during 1985-1997 years underwent balloon dilatation of the esophagus. 18 patients had stage IV, 5--stage III and 2--stage II of the disease. Mean diameter of the stricture's area in the esophagus made up. 7.2 +/- 2.0 mm. Balloon dilatation was performed in 4 patients by 2-4 balloons d = 10 mm in one stage, and in the test patients by balloon "Rigiflex" d = 40 mm. 2-3 procedures were carried out with the interval 7-10 days. In all cases balloon dilatation was successful. Mean diameter of the esophageal lumen after dilatation has increased to 16.0 +/- 2.5 mm. In 2 patients with IV stage of the disease relapse was detected within 6-8 months. 5 year follow-up results were satisfactory in 4 patients, from 5 to 10 years--in 14 patients, and over 10 years--in 5 patients. Prolonged clinical follow-up (for 7.5 years) demonstrated complete absence of dysphagia and normal regime of nutrition. Balloon dilatation is safe, available and effective method of nonoperative treatment for achalasia of the esophagus. PMID- 10533364 TI - [Method of protecting of sutures in the stomach and intestines]. AB - The authors have developed and described the method for intraoperative prophylaxis of complications caused by incompetence of the sutures on the stomach and bowels. It consists in closure of the sutures along the line of contiguity by two-layers of compact-porous collagenous explants, which possess antibacterial properties, the period of resorption in abdominal cavity being 15-20 days. The characteristics of the explants are presented and their advantages compared to commonly used methods covering by pellicle or glue film peritonization by sero muscular sutures. Application of collagenous explants from "Sanguicol" preparation in surgical treatment of 155 patients with external intestinal fistulas, compared to peritonization by sero-muscular sutures (66 patients), has demonstrated a high effectiveness of proposed explants in prevention of postoperative intraabdominal complications. PMID- 10533365 TI - [Lipid peroxidation in acute surgical diseases of abdominal cavity organs]. AB - The study of lipid peroxidation (LPO) characteristics in patients with acute mesenterial circulatory disturbances (64), acute pancreatitis (44) and acute bowel obstruction (46) allowed the conclusion on informative value of determination of malonic dialdehyde and diene conjugates in the blood for staging these diseases and the degree of severity of endotoxicosis. On the basis of the state of LPO it became possible to evaluate the extent of bowel failure. According to LPO in postoperative period the process of development of vascular rethrombosis in the bowels could be assessed. The varieties of lipid peroxidation products in peripheral venous blood can be used for differential diagnosis of such diseases, similar in their clinical symptoms, as acute pancreatitis and acute bowel obstruction. PMID- 10533369 TI - [Tests of durability characteristics of osteosynthesis of femoral neck fractures by different fixators under static and dynamic loading]. AB - Test bench experiments on the models of femoral neck bone fractures with the use of various kinds of latches has been carried out for evaluation of durability characteristics of the osteosynthesis, using universal test machine "Zwick-1464". 40 preparations of proximal-diaphysis parts of the femoral bone from 28 females and 12 males (mean age 71 years) were used one day after death. The models of basal, transcervical and subcapital fracture were formed with the angle between its planes and horizontal plane (the angle of Pauel's) 30, 40 and 70 degrees. The models of fractures underwent static and dynamic loads, the results being fixed automatically in graphics. The obtained data have confirmed the dependence of the stability of the osteosynthesis by tested fixators on the size of Pauels' angle: the more was this value the less was the firmness of the junction. Dependence of the firmness of fixation of bone fragments on location of the fracture in femoral neck was noted. The differences of durability characteristics of created constructions in basal, and transcervical fractures of types I and II by Pauels are insignificant. The authors think that the least invasive mode for fixation among tested constructions is osteosynthesis with V-shaped bunch of needles, which produced minimal trauma for bone tissue and created resilient tension in the system of bone fixator (latch), which is especially important in osteoporosis. PMID- 10533375 TI - [Tracheobronchoscopy in injuries of the thorax and neck]. AB - The results of examination and treatment of patients with penetrating wounds of the thorax and the neck are analysed. Diagnostic potential abilities of roentgenography and tracheobronchoscopy are evaluated comparatively. High diagnostic and curative effectiveness of tracheobronchoscopy is detected, which is indicated in diagnosis of the wounds of the larynx and the trachea as well as in cases of technical difficulties during endotracheal intubation. Besides, tracheobronchoscopy is indicated in postoperative period for sanation of the tracheobronchial tree under visual control for prevention of broncho-pulmonary complications. PMID- 10533377 TI - [Autotransplantation of splenic tissue after splenectomy]. PMID- 10533371 TI - [Surgical treatment of chronic post-traumatic osteomyelitis of the femur in pseudarthrosis, ununited fractures and defects]. AB - The authors have analyzed the results of treatment for chronic posttraumatic osteomyelitis of the femoral bone in 69 cases of ununited fractures, false joints ad femoral bone defects. The treatment included surgical sanation of the osteomyelitic nidus together with aspiration lavage drainage and extranidal osteosynthesis by external fixation as well as local and general antibacterial therapy. In 13 cases free bone autoplasty of the osteomyelitic cavities or defects was used together with spongious structures of iliac bone (crest and wing), in 2 of these patients it was applied for replacement of the large defects of the diaphysis of the femoral bone (8 and 12 cm). In 57 patients surgical treatment was carried out in one stage, a major requisite for this was the opportunity for joining bone fragments inside the open wound after sequestrectomy as well as stable external fixation by the apparatus. This treatment gave successful results in 97% of patients. PMID- 10533370 TI - [Main principles of etiotropic therapy in chronic osteomyelitis]. AB - The results of prospective trial in 352 patients treated surgically for chronic osteomyelitis were presented: 151 patients with hematogenous and 201--with posttraumatic osteomyelitis. Various antibacterial treatment modalities were used for sanation of fistulas and purulent foci--chemical antiseptics, ultrasound, laser, flowing--lavage drainage, immobilized antibacterial drugs on various filling materials. Bacteriological examinations, including verification of pathogens and quantitative measurement of microbes in the tissues, has definitely proved effectiveness of such combined methods of treatment. The author suggests that sanation of the purulent focus should be applied in all stages of surgical treatment for such patients: during preparation for surgery, intraoperatively and in postoperative period. PMID- 10533368 TI - [Bilateral reconstruction of the renal arteries by laparotomy approach]. AB - Bilateral disease of renal arteries due to renovascular hypertension is encountered, according to the authors' data, in 23% of cases. The standard method of surgical treatment, generally accepted in Russia, consists in staged reconstruction of renal arteries through the thoracophrenolumbotomy approach. For the first time, one stage reconstruction of both renal arteries by type of patch "Gore-Tex" isthmoplasty through laparotomy approach was performed in RRCS RAMS. Technical aspects of the procedures performed in 2 patients are described in details. The authors suggest that one stage reconstruction of both renal arteries through the laparotomy approach is the method of choice in bilateral stenosis of renal arteries and allows to perform not only isthmoplasty but also prosthetic reconstruction of renal arteries up to their bifurcation without intersection of the right renal vein and its branches. PMID- 10533376 TI - [Stimulation of medical personnel work in operating rooms]. AB - Preliminary results of experimental trial on stimulation of medical staff's labour in the operation unit, held in medical institutions of Chelyabinsk, has been analysed. As a basis for calculation of the payments for the work, the labour expenditures of each member of surgical team (surgeon, assistant, anesthesiologist, surgical nurse, anesthesist, litter bearer nurse) and the cost of 1 hour work were assessed, the degree of the difficulty of each operation, as well as the total cost of the procedure was determined. Registration of the work of all members of the surgical team and its payment, according to the quantity and the quality of the performed operations, demonstrates the stage of transition to the system of payment for the labour, i.e.--for the concrete work, done by somebody of the staff (for the treatment of some patient, etc.). The evaluation of the work according to the number of treated patients (operations being done) and to the quality of the treatment provides the conditions for personal financial incentive of surgical staff to intensification of their work and contributes to the achievements of favourable final results and in the same time- it provides decrease of the terms for patients' stay in the hospital, saving money and material resources. PMID- 10533372 TI - [Treatment of ununited fractures and pseudarthrosis of long bones of the lower limbs complicated by osteomyelitis]. AB - 123 patients with ununited diaphyseal fractures and false joints of the shin bones (93) and the hip (30), complicated by purulent infection, were treated in the clinic for pyogenic complications and consequences of locomotor traumas of the CITO from 1988 to 1994 years. Most patients were men (110 patients). The posttraumatic period varied from several months to 12 years. Many patients before their admittance to the clinic were not once unsuccessfully operated 69 (56.1%) patients had osseous defects of the thigh and the shin, all patients contractures of joints of the lower extremities as well as substantial trophic disturbances and atrophy of the muscles. 12 patients underwent closed transosseous osteosynthesis by the apparatus of an external fixation. One stage surgery was carried out in 111 patients, which included radical fistulosequestrenecretomy (and when indicated--extended segmental sequestrectomy) and transosseous osteosynthesis by the apparatus of external fixation with the use of the needles, rods, or their combination. The feature of the osteosynthesis depended on the extent of bone defect, which has been developed after sequestrectomy, in the defect up to 4 cm monolocal osteosynthesis was used, in the defect from 5 to 8 cm -bilocal and if it were over 8 cm--multilocal osteosynthesis was preferable. One stage treatment resulted in favourable outcomes in 95.9% of patients: the osteomyelitis was eliminated, the integrity of the bone was restored and in a great majority of cases--the length of the extremity was preserved. The treatment duration was 1.5-2 times less than in commonly used methods. PMID- 10533374 TI - [Results of the treatment of pseudarthrosis of the tibia by subsurface and extrafocal osteosynthesis]. AB - The results of treatment of 417 patients with pseudoarthroses of the tibia (aged from 17 to 71 years) with the use of various methods: by internal fixation with auto- and allografts, metal plates as well as in the use of extranidal (extralocal) osteosynthesis were analysed. The number of consolidation failures was minimal after autoplasty by Khakhutov and bone autoplasty. The rate of healing after osteosynthesis was substantially lower in internal fixation, than in extranidal (extralocal) osteosynthesis. Inflammatory complications occur less frequently when osteoplasty by Khakhutov was used. The highest rate of complications was observed in tibialization and in bilocal osteosynthesis. PMID- 10533378 TI - [Reversal of drug resistance in bacteria and tumor cells. (Drug research based on computer-predicted activity)]. AB - This paper present a short overview on the new perspectives in drug-design against drug resistance phenomena focusing on the infectious antibiotic resistance of bacteria. The role of plasmids in resistance of bacteria, the mode of transmissions were analyzed in details. The elimination of plasmids--the genetic basis of bacterial resistance located extrachromosomally--was studied systematically with several heterocyclic compounds. Correlations were analyzed between the anti-plasmids++ effects and chemical structure of tricyclic compounds in quantitative structure activity relationship (QSAR) studies. A hypothesis was presented to exploit the analogous mechanisms of multidrug resistance in bacterial and cancer cells by inhibiting the action of drug exporter proteins in the presence of compounds synthetized on the basis of computer aided drug design. PMID- 10533381 TI - [Meckel diverticulum as a possible source of complications]. AB - The authors performed a retrospective analysis of 108 operations performed because of Meckel's diverticulum from Jan. 1, 1980 until Dec. 31, 1997. 87 incidental operations were performed (80.55%) and 21 operations were performed (19.54%) because of complications of Meckel's diverticulum. Of the 87 incidental cases, 44 were males and 43 were females. The diverticulum was resected in 70 cases, was inverted in 16 patients and was left untouched in 1 patient. There were postoperative complications in 10 cases (11.5%), however, only one complication was related to the diverticulum operation. There was one death due to pneumonia in the incidental group (1.14%). Of the 21 cases having complications there were 5 females and 16 males. The following complications were observed: 4 cases of ileus, 7 cases of perforation (a foreign body was found in two cases) and 10 cases of inflammation. The Meckel's diverticulum was resected in 19 cases and the small intestine was resected in 2 cases. Postoperative complications occurred in 3 cases (14.3%), however these were not related to the diverticulum operations. There were 2 deaths due to pneumonia and pulmonary embolism (9.5%). Histological examination revealed 7 cases of heterotopic gastric mucosa in both the incidental group and the group with complications. There was one case of heterotopic pancreatic tissue in the group with complications. The incidence of heterotopy in the incidental group was 8%, whereas in the group with complications the incidence was 33.3%. Histological examination also revealed reflux gastritis-like diverticulitis in 5 of the patients with heterotopic gastric mucosa. Meckel's diverticulum was diagnosed in 2 patients preoperatively by upper gastrointestinal series. PMID- 10533373 TI - [Surgical treatment of chronic osteomyelitis of the sternum and the ribs]. AB - Transsternal approach is commonly used in majority of operations in heart surgery. In 0.5-5.9% of patients after median sternotomy osteomyelitis of the sternum and the ribs develops. The authors set forth their experience in surgical treatment of 182 such patients. In 97% of them total resection of the sternum was combined with simultaneous resection of costal cartilages which were involved in pyogenous inflammatory process. Costal cartilages were resected during subtotal resection of the sternum in 95% of cases and ribs--in 25%. Limited resection of the sternum was used only in patients with suppuration in the area of sutures in the sternum. Radical one-stage resection of the pyogenous site at the anterior thoracic wall was carried out in 62 patients. In 120 patients in poor condition and who previously underwent an opening of the abscess, staged surgery (2 or more operations) was performed. The authors suggest that in patients with cardio pulmonary and hepato-renal insufficiency it is advisable to repair the defect, developed after resection of the thoracic wall, by split and perforated cutaneous flap. Muscular flap (32 patients) and greater omentum flap (30 patients) with vascular pedicles or local tissues (28 patients) were used for plastics of the thoracic wall defects. In 98 patients for the closure of the defect autodermoplasty with free split perforated cutaneous flap was used, otherwise the wound of the thoracic wall in them recovered by secondary intention type. 168 (92%) patients have recovered, 4 patients developed recurrence of osteomyelitis, 10 patients died. The authors suggest that the treatment of such serious patients should be carried out in specialized departments which have experience in thoracic, purulent and plastic surgery. PMID- 10533380 TI - [New diagnostic possibilities in dermatologic diseases]. AB - The author reviews the latest developments regarding the most important immunological methods for the diagnosis of dermatological diseases. The salt split-skin, fluorescence overlay antigen mapping and immunoblot techniques are useful methods for the differentiation of autoimmune bullous skin diseases. The use of recombinant fusion products of antigenic epitopes in patients with pemphigus and bullous pemphigoid affords a possibility for the differentiation or diagnosis of the disease by means of a simple ELISA technique. In situ hybridization and the polymerase chain reaction (PCR) technique are very useful in many dermatological diseases to demonstrate infective agents or genetic abnormalities. PMID- 10533379 TI - [Percutaneous ultrasound-guided ethanol sclerotherapy of autonomous thyroid nodules]. AB - Percutaneous ethanol injection therapy for autonomously functioning thyroid nodules has been performed in 53 patients. 36 patients suffered from hyperthyroidism, and 17 patients had subclinical hyperthyroidism. Ethanol was administered under ultrasonographic guidance in 2-6 sessions depending on the size of the nodule Local neck pain was the most often adverse effect. Transient dysphonia occurred in 3 patients. A subacute granulomatous thyroiditis-like reaction within 1 week after the last session occurred in 4 patients. During a 10 day steroid administration this reaction was stopped. After ethanol sclerotherapy reduction of thyroid nodular volume can be achieved. The reduction of the nodules was between 36 and 75% (mean 55 +/- 15%) of the pre-treatment volume at 6 week after therapy. In 27 of 36 hyperthyroid patients the FT4- and T3-levels became normal. Repeated sclerotherapy was successfull in 6 of the remaining 9 hyperthyroid patients. No relapse of hyperthyroidism was observed. The scintiscan showed a complete cure in 10 of 23 patients one year after PEI-therapy, while in 11 patients partial normalization of the scintiscan was observed. In 2 of 23 patients the scintiscan remained unchanged. Indication of ethanol sclerotherapy is not clear. The method appears an effective alternative procedure in patients with large nodules at high surgical risk. Under special circumstances (pregnancy or iodine-induced hyperthyroidism) ethanol sclerotherapy may be a practical alternative for toxic autonomously functioning thyroid nodules. PMID- 10533382 TI - [Percutaneous embolization of hemorrhaging renal arteriovenous fistula]. AB - A 67 year old male patient with macroscopic hematuria was treated by percutaneous intraarterial embolization with Gianturco-coil (Cook). The patient had a percutaneous stone removal from the left kidney two weeks before the bleeding. The emergency angiography showed an arteriovenous shunt in the left kidney. A superselective embolization was performed. There was no bleeding on the control angiography. The embolization was performed without complication. PMID- 10533383 TI - [A hypothetical model of the biomechanical interaction of the teeth and the abutment tissues of the jaws during masticatory loading of different values]. AB - Biomechanical model "tooth-jaw" in considered. This biomechanical model takes into account local nonhomogeneous mechanical constructions tooth-jaw, its special geometry, physical properties of firm tissues of tooth, periodontium and jaw. Using mathematical methods, determination of supporting jaw tissues depending by computer tomograms was grounded. The system SPLEN-K has calculated tension deformed condition of dental tissues in area of a group of chew teeth. PMID- 10533385 TI - [The effect of immobilized salivary alkaline phosphatase on remineralization processes]. AB - Relationship between alkaline phosphatase immobilized on apatite and remineralization processes has been studied. The enzyme notably accelerated the remineralization. The course of enzymatic remineralization depended on the source of phosphorus (inorganic phosphorus or beta-glycerophosphate). Calcium incorporation in remineralizing surface was the most rapid in the presence of beta-glycerophosphate. This may be explained by a higher rate of formation of apatite under the effect of immobilized alkaline phosphatase, than of brushite or vitlokite. PMID- 10533384 TI - [The sources of active forms of oxygen in the normal oral tissues and in pathology]. AB - In the experiment on rats the changes of the superoxydanionradical production from microsomal and mitochondrial oxidation and tissue phagocytes of the saliva gland and parodontium in contact sialadenitis, spontaneous parodontitis, after ionizing radiation have been investigated. It is supposed that an increase or decrease of superoxydanionradical production from different sources demands a differentiated corrective approach. PMID- 10533386 TI - [The experimental and clinical validation of the use of a polikatan preparation in periodontal diseases]. AB - Polycatane, a bischofite-based drug (bischofite is a mineral containing up to 95% dry residue of MgCl2 x 6H2O), exerted antiinflammatory and bacteriostatic effects and stimulated phagocytosis and healing of burn infected ulcer in the buccal mucosa of experimental animals. The drug was effective in the treatment of patients with chronic catarrhal gingivitis and generalized periodontitis of light and medium severity. By its antiinflammatory and wound-healing effects polycatane is superior to polyminerol, the reference drug. PMID- 10533387 TI - [The characteristics of the immune status of patients with lichen ruber planus]. AB - All three main components of the immune system (phagocytosis, humoral and cell mediated immunity) are changed in patients with lichen ruber planus (LRP). Changes in the functional activity of phagocytes manifest by a decreased phagocytic activity of the peripheral blood monocytes towards Staphylococcus aureus and increased production of active oxygen forms during phagocytosis, which are registered by luminol-dependent chemiluminescence. Changes in the humoral immunity consist in increased serum levels of IgG and IgA, and shifts in the cell mediated immunity manifest by increased count of cytotoxic CD8+ T lymphocytes and decreased functional activity of T cells recorded by the proliferative response to phytohemagglutinin. The greatest shifts in the immune status are observed in patients with erosive ulcerative LRP, which is characterized by the greatest severity and intensity of inflammation. PMID- 10533390 TI - [The effect of ozone therapy on the dynamic spectrum of the peripheral blood proteins in patients with phlegmons of the face and neck]. AB - The influence of ozone therapy (parenteral and local application of ozonized solutions) and conventionally used methods of treatment on spectrum dynamics of peripheral blood proteins of 59 patients with phlegmon of the face and the neck was studied. Application of ozone in combined treatment of the given pathology was found to contribute to correction of protein synthetic liver function. Manifested by the rise in albumin level and by decrease in globulin synthesis activity. The effect is explained by hepatocyte metabolism change because of activation of enzymatic intracellular processes. PMID- 10533388 TI - [A trial of using Solcoseryl dental adhesive paste in the surgical treatment of patients with chronic periodontitis]. PMID- 10533389 TI - [The monitoring of immunological indices in odontogenic abscesses and phlegmons and their differential diagnostic meaning]. AB - The immune homeostasis parameters have been monitored in 31 patients with odontogenic abscesses and phlegmons. Immunological values possessing differential diagnostic significance have been detected. Activation of phagocytosis, spontaneous and stimulated activity of phagocytes in parallel with a moderate decrease in the cellular functional reserve, increased immunoregulatory index, and moderate shifts in the concentrations of IgG and IgM are characteristic of odontogenic abscesses, while odontogenic phlegmons are characterized by deep depression of the phagocyte function and failure of cellular and humoral immunity. The disorders in the immune functions of patients with odontogenic phlegmons normalized 2 weeks later than in those with odontogenic abscesses. Both diseases run a wave-like course and are characterized by phasic changes in immunological parameters, which is significant for determining pathogenesis and choosing appropriate immunocorrective therapy. PMID- 10533391 TI - [Low-frequency pulsed magnetotherapy combined with electrostimulation of biologically active points in the combined treatment of traumatic mandibular osteomyelitis]. AB - The results of treatment are analyzed in 51 patients (35 with exacerbation of chronic traumatic mandibular osteomyelitis and 16 with chronic traumatic mandibular osteomyelitis). Low-intensity pulsed magnetic therapy of the focus in combination with electric stimulation of segmentary bioactive points, synchronized by the patient's pulse, are proposed to be added to the therapeutic complex. Such a modality improved the regional hemodynamics, promoted liquidation of the postoperative edema on days 1-2 after intervention, and sooner than after traditional therapy repaired the energy of the patient's organism. PMID- 10533393 TI - [The choice of dental implants in different types of atrophy of the jaw bones]. AB - Atrophic sites of the jaw bones with complete and partial adentia are classified. This classification helps the practitioner properly choose the dental implant for orthodontic treatment. PMID- 10533392 TI - [The etiology of salivary calculi]. AB - Ninety-four patients with chronic sialodochitis, 45 of these with sialolithiasis, have been followed up for a long time. Sialolithiasis develops in patients with congenital defects in the ductal system of the salivary gland presenting as ectasias and can be regarded as calculous sialoductitis. The etiological factors of sialolithiasis (calculous sialoductitis) are strictures (stenosis) of the ducts (distal portions) and specific topography of the duct (a "broken" duct with abrupt curves in which the concrements may form). PMID- 10533395 TI - [The characteristics of the clinical status of the periodontal tissues in children with gingival recession]. AB - Periodontal tissues were examined in 29 healthy children and in 91 children with gingival recession aged 6-14 years. The routine indices were higher in patients with gingival recession associated with chronic catarrhal gingivitis and maxillodental abnormalities than in those with only one oral disease. PMID- 10533394 TI - [The correction of a high ridge and bridge of the nose]. AB - The proposed method for correcting congenital deformations of the nose is less traumatic and its cosmetic results are more stable. The procedure is as follows. After endonasal dissection and wide stratification of the skin of the ridge and bridge of the nose, the osseocartilagenous protrusion of the ridge and the bridge are removed not by depression, but by resection and removal with subsequent bilateral paramedial and transverse-lateral osteotomy of the nasal bones and their mobilization into a proper position. The method may be used for correcting a high ridge of the nose in any of its deformations: an osseocartilagenous protrusion, high wide ridge, uneven ridge after an injury, etc. PMID- 10533396 TI - [Maxillodental anomalies and consanguineous marriages]. AB - Families of 549 probands and families of 123 probands with cleft lip and/or palate were examined in order to evaluate the relationship between marriages between close relatives and the incidence and structure of maxillodental diseases. Clinical and genealogical analysis of families of probands with maxillodental abnormalities and cleft lip and/or palate showed a significantly higher incidence of marriages between close relatives and an inbreeding coefficient in these families. Analysis of the population and familial incidence of maxillodental abnormalities and the inbreeding coefficient will help the physicians consulting such families more accurately evaluate the risk and improve the efficacy of prevention of such conditions. PMID- 10533397 TI - [The time periods for the formation of the permanent occlusion in children born and permanently living in a region close to a former nuclear test range]. AB - The terms of eruption of permanent teeth were analyzed in 2000 subjects aged 3-19 years born and permanently living in a region adjacent to a former nuclear testing field in Kazakhstan. All groups of permanent teeth erupted sooner, and the rate of eruption was higher in girls than in boys. Except the 4th and 5th teeth, all groups of permanent teeth erupted sooner on the mandible than in the maxilla. The formation of occlusion was delayed in children born and living in the studied region in comparison with the children in central Russia and in the Extreme North. Eruption of some groups of teeth was delayed in older age groups. These data may be used at all pedodontics departments in organization of sanitation of children and in evaluation of the physical development and formation of orthodontic status of children. PMID- 10533398 TI - [A system for feeding an antiseptic solution to the Ultradent apparatus]. PMID- 10533399 TI - [The main trends in the development of the computerization of public health and the system of mandatory medical insurance for 1999-2000 and the tasks of the dental service of Russia]. PMID- 10533400 TI - [Planning for the raising of dentists' qualifications by taking into account the need of practicing physicians for consultative help]. PMID- 10533401 TI - [The Nordic article of the month]. PMID- 10533403 TI - [Genetic epidemiology and twin research]. PMID- 10533405 TI - [Radiologic intervention in the biliary tract performed at a central hospital]. AB - During the ten-year period 1987-96, 131 diagnostic percutaneous transhepatic cholangiographies (PTC) were performed in 103 patients due to obstructive jaundice. 54 cases of percutaneous transhepatic bile drainage (endoprosthesis) (PTBD) and 53 of percutaneous transhepatic external drainage (PTED) were also performed. In 89 patients (86%) the obstructive jaundice was caused by malignant disease. Puncture was done under fluoroscopic or ultrasonographic control. An 8 Fr. plastic endoprosthesis of 15 cm length was used for internal bile drainage, and a 7.6 Fr. 60 cm long catheter was used for external drainage. For combined external/internal drainage an 8.4 Fr. catheter of 60 cm length was used. 24 patients (23%) developed complications. Eight of these complications were serious, and three patients (2.5%) died as a result of the procedure. Three patients developed duodenal perforations. 11 out of 51 patients (20%) treated with endoprosthesis died within 30 days. Mean functioning time for endoprostheses was 128 days. Progress in radiologic intervention technique has changed the treatment of obstructive jaundice. In our view, endoscopic bile drainage should be the treatment of choice. Percutaneous transhepatic bile drainage is an alternative in cases where endoscopic therapy fails. PMID- 10533406 TI - [Percutaneous drainage of the gallbladder in acute cholecystitis]. AB - Percutaneous cholecystostomy has replaced surgical treatment for acute cholecystitis in surgical high-risk and critically ill patients. We wished to assess the procedures performed at Lillehammer County Hospital. We report 32 drainages performed in the last three years. Clinical records and radiological reports were reviewed retrospectively. Technical problems and complications during and after the procedure were recorded. In spite of relatively few procedures per radiologist, all were initially successful. No serious complications related to the procedure were encountered. Dislocation of the catheter occurred in 5 out of 32 drainages. This corresponds well to earlier reports. We conclude that percutaneous cholecystostomy in acute cholecystitis is a safe procedure in this group of high-risk patients at our hospital. PMID- 10533404 TI - [Magnetic resonance tomography of biliary and pancreatic ducts]. AB - Magnetic resonance imaging of the biliary and pancreatic ducts, MRCP, is a technique developed over the last few years. Using strongly T2-weighted sequences, images of the biliary and pancreatic ducts similar to ERCP can be obtained within one single inhalation. No contrast media or medication is required. In 23 patients 25 MRCP examinations were retrospectively compared with ERCP or PTC. One patient had normal findings; three had gallbladder stones. Eight out of nine common bile-duct stones were shown. MRCP after papillotomy in one patient showed a common bile-duct stone; ERCP seven days later was normal. MRCP correctly showed obstruction and dilatation of the bileducts in ten patients with tumor and in one patient with chronic pancreatitis. Two of these were erroneously interpreted as caused by stone. 21 of 25 MRCPs were consistent with the final diagnosis. We consider MRCP a promising method which may replace diagnostic ERCP in majority of patients. Stones in the gallbladder and bile-ducts can be diagnosed. The method also shows obstructions and other lesions affecting pancreatobiliary ducts. PMID- 10533407 TI - [Diagnostic imaging in intestinal pneumatosis]. AB - Abnormal air in intestinal walls, mesentery and portal veins can be caused by serious illness like gangraena or necrosis. It can also accompany other conditions that may be treated conservatively. We describe three patients with pneumatosis intestinalis. Special emphasis is given to imaging techniques, particularly computer tomography. We recommend continuous axial slices and the use of an oral contrast agent. Correct settings of window and level is important for diagnosis of this rare disease. PMID- 10533402 TI - [Are too few liver transplantations performed in Norway?]. PMID- 10533408 TI - [Primary lymphoma of the central nervous system]. AB - Primary lymphoma of the central nervous system is an important diagnosis to consider in any patient with an expansive lesion of unknown origin. The incidence of this cancer is reported to be increasing. We collected retrospective data from the files on all patients with brain tumours treated surgically at the University Hospital of Tromso in the 1986-98 period (n = 513). Of 283 patients operated for brain tumours from 1986 to 1994, only one (0.4%) had a primary lymphoma. Of 230 patients treated from 1995 to 1998 seven (3%) suffered from the same condition. Two of the eight patients had T-cell lymphomas. Primary lymphoma of the central nervous system is a highly invasive tumour and should preferably be treated with chemotherapy and irradiation. The use of surgery is controversial. In most cases, surgery should be restricted to biopsy only. PMID- 10533409 TI - [Diagnostic imaging of parathyroid tumors]. PMID- 10533410 TI - [Anxiety and depression--a common mixture]. AB - Mixed symptoms of anxiety and depression are highly prevalent among patients with psychic complaints. The number and duration of symptoms and the degree of impairment determines whether the condition fulfils the criteria for an anxiety and depressive disorder. Increasing attention is now being given to the clinical significance of subsyndromal anxiety and depression because of the functional impairment inflicted. Epidemiological studies and clinical experience show that comorbidity of anxiety and depressive disorders is highly prevalent. This may partly be attributed to common genetic causes; partly to environmental causes. Concurrent anxiety and depression pose a challenge in term of differential diagnoses. The treatment of such co-morbid disorders also calls for some special precautions. PMID- 10533411 TI - [Medullary compression in metastatic cancer]. AB - Cancer patients with spinal cord compression may develop irreversible neurological deficit. The clinical picture implies back pain and subsequent neurological deficit. There is always a danger of rapid deterioration of the patient's condition. If spinal cord compression is suspected, the case is an emergency. MRI should be preferred in the diagnostic work-up, and corticosteroids be administered promptly. Radiation therapy or surgical treatment should be started as soon as possible. Patient outcome is related to the degree of neurological deficit at the start of treatment. PMID- 10533412 TI - [Borderline primary hyperparathyroidism]. AB - The clinical presentation of primary hyperparathyroidism has changed considerably after the application of biochemical autoanalysers. The condition was previously found with characteristic symptoms in bone, kidneys and the gastrointestinal tract; less than 1% of patients were assumed to be asymptomatic. Today, most patients have mild and uncharacteristic symptoms. Primary hyperparathyroidism has been detected with increasing frequency in the western world, but there are large variations within demographically otherwise comparable areas. There are also regional differences regarding the incidence of surgical treatment of the disease. Demographic studies have shown increased morbidity and mortality primarily from cardiovascular and possibly malign diseases related to hypercalcemia. Follow-up studies based on surgical series have shown increased mortality related to preoperative serum calcium level. In view of the altered clinical picture of primary hyperparathyroidism, treatment of moderate and possibly "benign" primary hyperparathyroidism has been actualised. At an international consensus conference in 1990, this patient group was defined by an arbitrary serum calcium limit of 3.0 mmol/l, but this limit has been found to be unacceptably high. Regardless of where the upper limit is set, there are so far well accepted indications for surgery, but only one in two patients fulfil these criteria. A working group of endocrinologists and endocrine surgeons from nine university clinics in Scandinavia have recently initiated a prospective, randomised study evaluating surgical treatment and a systematic follow-up of patients with borderline primary hyperparathyroidism. PMID- 10533413 TI - [A man with nausea and pain under his right heel]. PMID- 10533414 TI - [Pancreas transplantation--a review]. AB - According to the International Pancreas Transplant Registry, more than 1,000 pancreas transplantations are now performed annually. When performed simultaneously with a kidney transplant, pancreas transplantation is a generally accepted treatment for type 1 diabetic patients with end stage renal disease, and the pancreas graft survival in this setting is equivalent to that of the kidney graft. The reported results of solitary pancreatic transplantation in non-uraemic diabetic patients have been less favourable due to a high rejection rate in combination with difficulties in rejection diagnosis and treatment. However, the introduction of new immunosuppressive drugs over the last few years has drastically improved the survival of solitary pancreatic grafts and allows for a rising enthusiasm. The majority of the 122 transplantations performed in Oslo so far were simultaneous kidney and pancreas transplantations. Duct occluded segmental grafts were used until 1988 when the pancreatico-duodenal technique with bladder drainage was introduced. In March 1998 we changed to enteric drainage of the exocrine pancreas due to a high percentage of lower urinary tract problems and bicarbonate loss associated with the bladder drainage technique. The positive impact of a functioning pancreas transplant on the recipient's quality of life is well recognised, whereas long-term beneficial effects on the secondary complications of diabetes are not well documented. A short overview of various aspects of pancreas transplantation is given, including experience with the 122 transplantations performed at the National Hospital, Oslo, Norway. PMID- 10533415 TI - [Liver transplantation--development and experiences]. AB - A liver transplant program was established in Norway in 1984, and until March 1999 200 liver transplantations have been carried out. Data for these 200 consecutive patients are briefly outlined with emphasis on survival. Relevant data are also given from the Nordic Liver Transplant Registry (NLTR), the European Liver Transplant Registry (ELTR) and from United Network for Organ Sharing (UNOS). Future trends and potential advances in liver transplantation are briefly discussed. One-year and three-year survival rates for Norwegian patients have increased markedly over the years and were 85% and 75% respectively for the 1995-98 period. The number of liver transplantations per million population per year was 3.4 in Norway, 7.8 in Sweden, 5.7 in Finland and 5.4 in Denmark (1990 98). The low number of liver transplantations in Norway warrants attention. It is possible that some patients with end stage liver disease have not been offered this treatment modality. Monitoring of results and active participation in international liver transplant registries like NLTR and ELTR is an important quality control instrument. PMID- 10533416 TI - [What can twin research reveal about the causes of diseases?]. AB - The purpose of this paper is to describe the place of twin studies in etiological research on somatic diseases, using asthma as an example. Twin studies provide answers to the relative importance of genes and environment in the development of disease, but have been criticised for systematic biases especially linked to unusual conditions in twin pregnancies. For asthma, classical twin studies show that genes explain the largest part of the interindividual variability, and that common family environment does not explain similarity in asthma in siblings. The assumptions underlying classical twin studies are discussed. A few other twin designs are mentioned, and we conclude that twin studies still play an important role in understanding causes of somatic diseases. PMID- 10533417 TI - [Twin studies in psychiatry]. AB - Twin research in medicine and psychology has blossomed during the last decades. A brief presentation of twin research methods is given, followed by a review of psychiatric studies, particularly Norwegian ones. The Nordic investigations are of great importance because many of these studies have been able to avoid shortcomings in sampling by linking national twin registers to disease registers. This procedure is of consequence for the estimation of heritability. Twin research shows that the genetic contribution to Alzheimer's disease, manic depressive (bipolar) disorder, schizophrenia and childhood autism is relatively strong, whereas genetic factors play a minor role in common depressions, anxiety and somatoform disorders, and alcohol abuse. Since the concordance rates are far from 100 percent in most of these illnesses, environmental factors must also be important. PMID- 10533418 TI - [Mortality in Norwegian hospitals--a critical comment]. PMID- 10533419 TI - [The identity of clones]. PMID- 10533420 TI - [Intravenous drug addicts]. PMID- 10533421 TI - [Internship at a regional hospital]. PMID- 10533422 TI - [Diagnostic imaging in acute renal infarction]. PMID- 10533423 TI - [Is quality assurance of registration and publishing of therapeutic results sufficient?]. PMID- 10533424 TI - Job insecurity in a broader social and health context. PMID- 10533425 TI - New patterns of employment in Europe. PMID- 10533426 TI - New dimensions of labour market citizenship. PMID- 10533427 TI - Health consequences of job insecurity. PMID- 10533428 TI - The downsizing epidemic in the United States: towards a cultural analysis of economic dislocation. PMID- 10533429 TI - Models of job insecurity and coping strategies of organizations. PMID- 10533430 TI - Major technological change, socioeconomic change and society's response. PMID- 10533431 TI - Labour participation and work quality policy: outline of an alternative economic vision. PMID- 10533432 TI - Health and work: concluding remarks. PMID- 10533433 TI - Reaction kinetics, molecular action, and mechanisms of cellulolytic proteins. AB - Cellulolytic proteins form a complex of enzymes that work together to depolymerize cellulose to the soluble products cellobiose and glucose. Fundamental studies on their molecular mechanisms have been facilitated by advances in molecular biology. These studies have shown homology between cellulases from different microorganisms, and common mechanisms between enzymes whose modes of action have sometimes been viewed as being different, as suggested by the distribution of soluble products. A more complete picture of the cellulolytic action of these proteins has emerged and combines the physical and chemical characteristics of solid cellulose substrates with the specialized structure and function of the cellulases that break it down. This chapter combines the fundamentals of cellulose structure with enzyme function in a manner that relates the cellulose binding and biochemical kinetics at the catalytic site of the proteins to the macroscopic behavior of cellulase enzyme systems. PMID- 10533434 TI - Genetic engineering for improved xylose fermentation by yeasts. AB - Xylose utilization is essential for the efficient conversion of lignocellulosic materials to fuels and chemicals. A few yeasts are known to ferment xylose directly to ethanol. However, the rates and yields need to be improved for commercialization. Xylose utilization is repressed by glucose which is usually present in lignocellulosic hydrolysates, so glucose regulation should be altered in order to maximize xylose conversion. Xylose utilization also requires low amounts of oxygen for optimal production. Respiration can reduce ethanol yields, so the role of oxygen must be better understood and respiration must be reduced in order to improve ethanol production. This paper reviews the central pathways for glucose and xylose metabolism, the principal respiratory pathways, the factors determining partitioning of pyruvate between respiration and fermentation, the known genetic mechanisms for glucose and oxygen regulation, and progress to date in improving xylose fermentations by yeasts. PMID- 10533435 TI - Successful design and development of genetically engineered Saccharomyces yeasts for effective cofermentation of glucose and xylose from cellulosic biomass to fuel ethanol. AB - Ethanol is an effective, environmentally friendly, nonfossil, transportation biofuel that produces far less pollution than gasoline. Furthermore, ethanol can be produced from plentiful, domestically available, renewable, cellulosic biomass. However, cellulosic biomass contains two major sugars, glucose and xylose, and a major obstacle in this process is that Saccharomyces yeasts, traditionally used and still the only microorganisms currently used for large scale industrial production of ethanol from glucose, are unable to ferment xylose to ethanol. This makes the use of these safest, most effective Saccharomyces yeasts for conversion of biomass to ethanol economically unfeasible. Since 1980, scientists worldwide have actively been trying to develop genetically engineered Saccharomyces yeasts to ferment xylose. In 1993, we achieved a historic breakthrough to succeed in the development of the first genetically engineered Saccharomyces yeasts that can effectively ferment both glucose and xylose to ethanol. This was accomplished by carefully redesigning the yeast metabolic pathway for fermenting xylose to ethanol, including cloning three xylose metabolizing genes, modifying the genetic systems controlling gene expression, changing the dynamics of the carbon flow, etc. As a result, our recombinant yeasts not only can effectively ferment both glucose and xylose to ethanol when these sugars are present separately in the medium, but also can effectively coferment both glucose and xylose present in the same medium simultaneously to ethanol. This has made it possible because we have genetically engineered the Saccharomyces yeasts as such that they are able to overcome some of the natural barrier present in all microorganisms, such as the synthesis of the xylose metabolizing enzymes not to be affected by the presence of glucose and by the absence of xylose in the medium. This first generation of genetically engineered glucose-xylose-cofermenting Saccharomyces yeasts relies on the presence of a high copy-number 2 mu-based plasmid that contains the three cloned genetically modified xylose-metabolizing genes to provide the xylose-metabolizing capability. In 1995, we achieved another breakthrough by creating the super-stable genetically engineered glucose-xylose-cofermenting Saccharomyces yeasts which contain multiple copies of the same three xylose-metabolizing genes stably integrated on the yeast chromosome. This is another critical development which has made it possible for the genetically engineered yeasts to be effective for cofermenting glucose and xylose by continuous fermentation. It is widely believed that the successful development of the stable glucose-xylose-cofermenting Saccharomyces yeasts has made the biomass-to-ethanol technology a step much closer to commercialization. In this paper, we present an overview of our rationales and strategies as well as our methods and approaches that led to the ingenious design and successful development of our genetically engineered Saccharomyces yeasts for effective cofermentation of glucose and xylose to biofuel ethanol. PMID- 10533436 TI - Ethanol production from renewable resources. AB - Vast amounts of renewable biomass are available for conversion to liquid fuel, ethanol. In order to convert biomass to ethanol, the efficient utilization of both cellulose-derived and hemicellulose-derived carbohydrates is essential. Six carbon sugars are readily utilized for this purpose. Pentoses, on the other hand, are more difficult to convert. Several metabolic factors limit the efficient utilization of pentoses (xylose and arabinose). Recent developments in the improvement of microbial cultures provide the versatility of conversion of both hexoses and pentoses to ethanol more efficiently. In addition, novel bioprocess technologies offer a promising prospective for the efficient conversion of biomass and recovery of ethanol. PMID- 10533437 TI - Production of multifunctional organic acids from renewable resources. AB - Recently, the microbial production of multifunctional organic acid has received interest due to their increased use in the food industry and their potential as raw materials for the manufacture of biodegradable polymers. Certain species of microorganisms produce significant quantities of organic acids in high yields under specific cultivation conditions from biomass-derived carbohydrates. The accumulation of some acids, such as fumaric, malic and succinic acid, are believed to involve CO2-fixation which gives high yields of products. The application of special fermentation techniques and the methods for downstream processing of products are described. Techniques such as simultaneous fermentation and product recovery and downstream processing are likely to occupy an important role in the reduction of production costs. Finally, some aspects of process design and current industrial production processes are discussed. PMID- 10533438 TI - The use of virtual reality exposure in the treatment of anxiety disorders. AB - One possible alternative to standard in vivo exposure may be virtual reality exposure. Virtual reality integrates real-time computer graphics, body tracking devices, visual displays, and other sensory input devices to immerse a participant in a computer-generated virtual environment. Virtual reality exposure (VRE) is potentially an efficient and cost-effective treatment of anxiety disorders. VRE therapy has been successful in reducing the fear of heights in the first known controlled study of virtual reality in the treatment of a psychological disorder. Outcome was assessed on measures of anxiety, avoidance, attitudes, and distress. Significant group differences were found on all measures such that the VRE group was significantly improved at posttreatment but the control group was unchanged. The efficacy of virtual reality exposure therapy was also supported for the fear of flying in a case study. The potential for virtual reality exposure treatment for these and other disorders is explored. PMID- 10533439 TI - A cognitive model of generalized anxiety disorder. AB - A cognitive model of generalized anxiety disorder (GAD) is described. The model asserts that generalized anxiety is an abnormal worry state. In this model, GAD results from the usage of worrying as a coping strategy and subsequent negative evaluation of worrying. The use of worry as a strategy is supported by positive metabeliefs concerning worry, whereas the negative appraisal of worrying (worry about worry) is linked to negative metabeliefs developed out of previous experience. These beliefs center on the themes of uncontrollability of worries and the dangerous consequences of worrying. Negative appraisal of worrying is associated with behavioral and cognitive responses that serve to maintain unwanted thoughts, and preserve dysfunctional beliefs. A review of the literature indicates that the model is consistent with existing data. Predictions and treatment implications of the model are discussed. PMID- 10533440 TI - Cognitive bias in eating disorders: implications for theory and treatment. AB - Research testing the predictions of cognitive-behavioral theory related to the psychopathology of eating disorders has lagged behind treatment outcome research. Central to cognitive theories of eating disorders is the hypothesis that beliefs and expectancies pertaining to body size and to eating are biased in favor of selectively processing information related to fatness/thinness, dieting, and control of food intake or body weight. In recent years, controlled investigations of the predictions of cognitive theories of eating disorders have yielded empirical support for these theories. This paper reviews research which has tested the predictions of cognitive-behavioral theory and discusses the implications of these findings for the treatment of eating disorders. Understanding of information processing biases may assist the clinician in understanding a range of psychopathological features of anorexia and bulimia nervosa, including denial, resistance to treatment, and misinterpretation of therapeutic interventions. PMID- 10533441 TI - A systematic analysis of the influence of prior social context on aggression and self-injury within analogue analysis assessments. AB - Experimental analogue analyses are sometimes used to identify the operant function of aberrant behavior for individuals with developmental disabilities. These methods are proposed to be superior to other assessment techniques because they systematically control for the presence and absence of hypothesized maintaining contingencies within the analogue analysis. Analogue analyses are usually performed over a brief period of time with little reference to the broader social context within which they are conducted. Social interactions that a person experiences prior to an analogue analysis assessment might influence performance within the assessment. The authors examined the influence of prior social conditions on levels of aberrant behavior under analogue assessments with two individuals with severe developmental disabilities. Results indicated that prior social conditions significantly influenced analogue analysis results in idiosyncratic ways for both. These findings are discussed in terms of the need to examine broad contextual influences when conducting functional analyses of aberrant behavior. PMID- 10533442 TI - A palmtop computer program for the treatment of generalized anxiety disorder. AB - This is the first report of a palmtop computer program developed to increase the efficiency and cost-effectiveness of cognitive behavioral therapy for generalized anxiety disorder (GAD). The computer program offers advantages to researchers, therapists, and clients. These advantages include continuous, unobtrusive collection of process data on treatment adherence as well as on the impact of cognitive behavioral therapy techniques in the client's natural setting. In addition, the computer extends treatment beyond the therapy hour and motivates clients to comply with homework assignments by prompting practice of cognitive behavioral strategies. The successful application of the palmtop computer program reported in this integrated series suggests a new line of research directed toward increasing the cost-effectiveness of what is currently the gold-standard treatment for GAD. PMID- 10533443 TI - Two-year follow-up of behavioral treatment and maintenance for body dysmorphic disorder. AB - Recent research has suggested that body dysmorphic disorder (BDD) is part of the obsessive-compulsive compulsive spectrum of disorders. As such, it has been hypothesized that these disorders respond in a similar manner to obsessive compulsive disorder when behavioral interventions are used. A continuation of follow-up was conducted with a group of patients with BDD following treatment. Ten patients completed an intensive behavioral therapy program and either participated in a 6-month maintenance program or served as controls. At 12-, 18-, and 24-month follow-up assessments, patients participating in the maintenance program were more effective at managing limited symptom return and had significantly lower anxiety and depression. Both groups remained improved for acute symptomatology and behavioral avoidance. The results suggest that maintenance programs following behavioral treatment are effective in preventing symptom relapse and assist in patient self-management of lapses typically associated with BDD. PMID- 10533444 TI - Teaching study skills and test-taking strategies to elementary school students. The Testbusters Program. AB - A pilot program to reduce test anxiety and related social-evaluative concerns is presented. Testbusters is a program designed specifically for elementary and middle school children in grades 4 through 7 that teaches effective study habits, study skills, and test-taking strategies and includes a behavioral contract to ensure consistent study behavior. The assessment strategy includes self-report instruments, a behavioral assessment, and children's grade point averages. After a 6-month waiting period, eight children with moderate to severe test anxiety participated in the 11-week program. The results indicated that Testbusters decreased general levels of test anxiety and self-ratings of distress when taking a test. Overall grade point average improved significantly and grades in the majority of the subjects showed positive improvement. There was no change in overall self-esteem or judgments of cognitive competence. The results are discussed in terms of the use of a skills strategy to decrease anxiety and improve academic achievement. PMID- 10533445 TI - Reliability of the Matson Evaluation of Social Skills in Individuals with Severe Retardation (MESSIER). AB - Recently, the concepts of social competence and social skills have become important aspects of the evolving definitions of mental retardation. However, no psychometrically sound instruments exist for assessing social skills in lower functioning developmentally disabled people. This study examined the psychometric properties of the Matson Evaluation of Social Skills for Individuals with Severe Retardation (MESSIER)--a new scale designed to measure social skills in adults with severe developmental disabilities. The authors conducted a preliminary evaluation of the test-retest and interrater reliability of the MESSIER. It was determined that the MESSIER has high stability across raters and good stability over time. In addition, good internal consistency was established with coefficient alpha. Potential uses for the scale and directions for future research are discussed. PMID- 10533446 TI - Familial risks of cancer. PMID- 10533447 TI - Cancer prevention: epidemiology and perspectives. AB - Following increases up until the late 1980s, some decline in cancer mortality has been observed in North America and in Western Europe. Approximately half the decline can be attributed to the levelling off in lung and other tobacco-related cancer epidemics and the rest to several factors, including reduced exposure to occupational carcinogens, prevention and early diagnosis, and improved treatment. Between 25 and 30% of all cancer deaths in Europe are due to tobacco smoking. In this review the effect of tobacco smoking on cancer incidence and mortality is examined, together with other important aetiological factors including alcohol, diet and environmental and occupational carcinogens. The effect of new treatments and the potential for prevention of cancer are also discussed. PMID- 10533448 TI - Adjuvant chemotherapy after curative resection for gastric cancer in non-Asian patients: revisiting a meta-analysis of randomised trials. AB - The aim of this study was to assess whether adjuvant chemotherapy after curative resection of gastric cancer increases survival rates. DATA SOURCES: MEDLINE (1966 1999), CancerLit (1983-1999), bibliographies, personal reprint files, and review articles were searched for relevant articles. Studies had to be randomised controlled trials of adjuvant chemotherapy versus observation following curative resection of stomach cancer that took place in non-Asian countries. Two reviewers independently evaluated the trials for eligibility, quality assessment and data abstraction, 13 trials met the eligibility criteria. The odds ratio for death in the treated group was 0.80 (95% confidence interval (CI) 0.66-0.97), corresponding to a relative risk of 0.94 (95% CI 0.89-1.00). Subgroup analyses showed a trend towards a larger magnitude of the effect when analysis was restricted to trials in which at least 2/3 of patients had node-positive disease. Our results suggest that adjuvant chemotherapy may produce a small survival benefit of borderline statistical significance in patients with curatively resected gastric carcinoma. Continued trials to find and confirm an effective adjuvant strategy are warranted. PMID- 10533449 TI - Lymphatic relapse in women with early breast cancer: a difficult management problem. AB - The aim of this study was to review the ability to control symptoms of regional lymphatic relapse in women with early breast cancer. A retrospective study was made of 759 consecutive women presenting with stage 1 or 2 breast cancer treated by breast conserving surgery and radiotherapy between June 1984 and December 1994, 291 (38.3%) of whom were managed by a policy of observation on the lymphatic pathways. Patterns of lymphatic relapse, relapse management and morbidity caused by recurrent malignancy were reviewed from the case notes. The overall rate of relapse in the ipsilateral axilla and/or supraclavicular fossa was 76/759 (10%) at any time prior to death or last follow-up. 34 of 65 patients who relapsed in the axilla did so despite prior axillary surgery and/or radiotherapy. 41 of 76 patients with regional recurrence presented with symptoms, including lymphoedema, arm pain or sensory motor changes. These symptoms were poorly controlled by palliative surgery, radiotherapy or systemic therapy in 23 cases, including 12 who progressed to arm paralysis. Symptomatic control of patients with regional lymphatic relapse can be very difficult, even in women under regular surveillance in a multidisciplinary breast cancer clinic. PMID- 10533450 TI - Effects of treatment with megestrol acetate on the insulin-like growth factor system: time and dose dependency. AB - This study was designed to evaluate time and dose dependency of alterations in insulin-like growth factor (IGF)-I, free IGF-I and the functional status of IGF binding protein (IGFBP)-3 in breast cancer patients during treatment with megestrol acetate (MA). In 16 patients receiving MA 160 mg daily, total IGF-I levels increased gradually (significant after 3 days on treatment) by a maximum of 2.66-fold after 5-6 months on treatment. However, free (readily dissociable) IGF-I levels increased to a smaller extent (1.23-2.15-fold). This discrepancy may be due to an increase in intact IGFBP-3 determined by Western ligand blotting (WLB). Similar findings were observed in 12 patients treated with MA in escalating doses from 40-800 mg daily. A dose-dependent increase in IGF-I was observed up to a dose level of 120 mg daily. We conclude that treatment with MA caused a profound increase in plasma levels of total IGF-I accompanied by a moderate increase in free IGF-I. This may explain the anabolic effects of MA in patients suffering from cachexia, but refute the hypothesis that alterations in the IGF-system may contribute to the antitumour effects of MA in breast cancer patients. PMID- 10533451 TI - 18F-FDG whole body positron emission tomography (PET) in patients with unknown primary tumours (UPT). AB - The management of patients with unknown primary tumours (UPT) often includes a large number of radiographical studies and invasive procedures, but the occult primary tumour is detected in less than 25%. In this prospective study we explored whether non-invasive whole body PET scans using FDG (18-F fluorodeoxyglucose) are of clinical value in detection of UPT. Whole-body FDG-PET scans were performed in 20 patients following standard staging procedures according to histology. PET results were verified either histologically or by the clinical course of the disease. 11 patients had neck metastases (5 squamous cell, 5 adenocarcinomas and 1 poorly differentiated carcinoma). The remaining patients had metastases located in bone (3), bone marrow (1), brain (1), pericardium (1), skin (1), pleura (1) and chest wall (1). All metastatic lesions were visible with PET. In 13 patients PET suggested the site for the primary tumour and this was verified in 9 (45%), either histologically or by the clinical course of disease. 8 of these had primary lung cancer and 1 had carcinoma at the basis of the tongue. In most patients PET had no treatment related implications. 3 patients with non-small cell lung cancer (NSCLC) received chemotherapy prompted by the PET result. The rest received either radical radiotherapy to the head and neck region (7), palliative radiotherapy to the metastatic lesion (8), chemotherapy based on signet ring cell carcinoma in bone marrow (1) or no therapy (1). These results indicates that PET is useful in UPT preceding expensive and invasive diagnostic procedures and can result in a faster diagnosis in approximately one third of the patients who then avoid unnecessary extensive procedures. Furthermore, a larger proportion of patients will receive treatment aimed at the correct diagnosis. A prospective cost-effectiveness analysis of PET in this setting is warranted. PMID- 10533452 TI - Expression of p53 and mdm2 mRNA and protein in colorectal carcinomas. AB - The aim of our study was to investigate the expression of p53 and mdm2 mRNA and protein in colorectal adenocarcinoma. For the detection of mRNA, 60 fresh frozen human tumour samples and 12 samples of corresponding normal tissue were examined. After total RNA extraction, reverse transcription (RT) was performed followed by cDNA amplification with specific primers using RT-polymerase chain reaction (PCR). Immunohistochemical detection of protein was examined in 81 formalin-fixed and paraffin-embedded human tumour specimens as well as 15 samples of adjacent normal colorectal tissue. p53 mRNA was detected in 80% (48/60) of the tumours and in 67% (8/12) of normal tissue samples; 87% (52/60) of tumours had mdm2 mRNA in contrast to only 17% (2/12) of normal tissue specimens. Nuclear p53 protein expression was observed in 52% (42/81) of the tumour specimens and in none of the 15 normal specimens, whereas mdm2 protein was found in the nucleus (31%, 25/81) and also in the cytoplasm (86%, 70/81) of tumour samples. In normal tissue, mdm2 protein expression was only observed in the cytoplasm (13%, 2/15) and not in the nucleus. There was a significant correlation between coexpression of p53 and mdm2 protein and the occurrence of lymph node metastases (P = 0.03) as well as between p53 protein expression and the occurrence of distant metastases (P = 0.007). Additionally, significant associations were found between p53 mRNA and p53 protein, p53 mRNA and mdm2 mRNA or protein, and also between mdm2 mRNA and mdm2 protein expression, supporting the existence of a regulatory mechanism involving p53 and mdm2. PMID- 10533453 TI - Cell-retained isoforms of vascular endothelial growth factor (VEGF) are correlated with poor prognosis in osteosarcoma. AB - Vascular endothelial growth factor (VEGF) is a major angiogenic factor. Osteosarcoma is characterised by hypervascularity and metastatic potential. We examined VEGF mRNA expression, VEGF isoform pattern and VEGF receptor (flt-1 and KDR) by RT-PCR analysis in 30 osteosarcomas. All 30 osteosarcomas expressed VEGF mRNA. 17 osteosarcomas (57%) expressed flt-1 mRNA, whilst 20 (67%) expressed KDR mRNA. 6/30 (20%) osteosarcomas were positive for VEGF121 only, 8 (27%) for VEGF121 + VEGF165, and 16 (53%) for VEGF121 + VEGF165 + VEGF189. Patients with osteosarcomas with VEGF165 (n = 24) had significantly poorer prognosis in comparison with those without VEGF165 (P = 0.022, Wilcoxon's test). The osteosarcomas with VEGF165 had significantly increased vascularity assessed on sections immunostained for CD34 (P < 0.001, Mann-Whitney U test). Although VEGF165 is a soluble isoform, it is also retained on the cellular surface. These results suggest that cell-retained VEGF isoforms (VEGF165, VEGF189) might be essential for neovascularisation in osteosarcoma, whilst the soluble VEGF121 isoform is not sufficient to stimulate neovascularisation in this type of neoplasm. PMID- 10533454 TI - A randomised dose-comparison trial of granisetron in preventing emesis in children with leukaemia receiving emetogenic chemotherapy. AB - This randomised study was performed to assess the anti-emetic efficacy and tolerability of two-dose regimens of granisetron in children with leukaemia. 49 children with leukaemia were treated with three consecutive courses of high-dose methotrexate or cytarabine regimen. During the first course, patients were evaluated regarding the emetogenicity of each regimen. They were randomised in a crossover manner to receive 20 or 40 micrograms/kg of granisetron before the second and third course of chemotherapy. Neither emesis nor severe appetite loss were observed in over 80% of patients within the first 24 h in all treatment groups. There was no significant difference in the anti-emetic efficacy between the two-dose regimens of granisetron. However, complete protection was achieved less frequently on days 2 and 3. Older children and girls appeared to be less well protected. No adverse events attributable to granisetron were observed. Granisetron dose regimens of 20 and 40 micrograms/kg are, comparably, well tolerated and effective in controlling chemotherapy-induced emesis in the first 24 h, though this protection fails thereafter, particularly in older patients and girls. PMID- 10533456 TI - Familial cancers in a nationwide family cancer database: age distribution and prevalence. AB - We calculated sex- and age-specific familial relative risks (FRRs) of cancer in offspring of cancer probands at 19 male and 20 female cancer sites, based on the nationwide Family Cancer Database from Sweden. The proportion of familial cancers among all cancers was also determined. The database contained 550,000 primary cancers. The familial risk at known sites: colon, breast, ovary, testis, skin (melanoma), nervous system, thyroid and other endocrine glands were confirmed. The FRR of thyroid cancer exceeded any other cancer and was over twice as high for male as for female offspring, and appeared to constitute an early- and late onset component. Novel register-based findings were familial risks in cervical and uterine cancer, and in male offspring of male probands kidney and skin (mainly squamous cell) cancer. Familial risks were noted also for lung cancer, lymphoma and leukaemia but they may have largely environmental causes. The proportion of familial cancers depended on the site, ranging from 11% in prostate to 8.7% in female breast and to well below 1% at many sites. PMID- 10533455 TI - Cerebral blood flow and glucose metabolism in long-term survivors of childhood acute lymphoblastic leukaemia. AB - Central nervous system treatment for childhood acute lymphoblastic leukaemia (ALL) has been reported to cause changes in cerebral blood flow and glucose metabolism. Little is known about the association of these functional changes with neuropsychological defects and structural changes. The aim of the present study was to assess the relationship between changes in regional cerebral blood flow and glucose utilisation in long-term survivors of ALL, and the association of these functional abnormalities with neurocognitive and structural defects. 8 survivors of childhood ALL were studied with single photon emission tomography (SPECT) using Tc99m-ethyl cysteinate dimer (ECD) as tracer and with positron emission tomography (PET) using 18F-fluorodeoxyglucose (FDG) as tracer. 8 healthy controls also underwent FDG-PET. All subjects also underwent magnetic resonance imaging and neuropsychological assessment 5 years after cessation of the therapy. Focal cerebral blood flow abnormalities were found in ECD-SPECT in 5 of the 8 survivors. Glucose utilisation appeared normal in the corresponding regions. However, glucose utilisation was decreased in thalamus and cerebellum in the survivors of ALL as compared with healthy controls. 3 patients had severe and 5 patients mild neurocognitive difficulties. The changes in cerebral blood flow and FDG uptake did not correspond neuroanatomically with the neurocognitive defects. Focal defects in cerebral blood flow in long-term survivors of ALL are not associated with changes in local cerebral glucose utilisation. Neurocognitive difficulties are not consistently associated with either changes in cerebral blood flow or with decreased glucose utilisation. Therefore, based on the present set of studies FDG-PET and ECD-SPECT cannot yet be recommended for the evaluation of long-term neurocognitive defects associated with treatment of ALL. PMID- 10533457 TI - The visibility of cancer on earlier mammograms in a population-based screening programme. AB - The aim of this study was to examine how frequently the later-round screen detected and interval breast cancers were visible in earlier screening mammograms by retrospective review and to compare their radiological and clinicopathological features with those diagnosed by primary screening. In a population-based mammography screening programme 63,731 women aged 50-59 years were invited and 56,158 examinations were carried out in the period 1987-1992 in the Tampere area in Finland. A total of 276 breast cancers were detected, of which 131 were diagnosed on later screening rounds or were interval cancers. A retrospective review of previous screening mammograms was carried out in 130 cases by the radiologist who diagnosed the breast cancer and thus knew the exact location of the tumour, no blinded review was carried out. 43 (33%) cancers were visible, 84 (65%) were not visible and 3 (2%) not included on the mammogram in a retrospective review. Later round screen-detected cancers were statistically significantly more often visible in earlier screening mammograms (43%) than interval cancers (19%) (P = 0.002). Tumours missed by screening mammography but which were visible on retrospective review were often histologically well differentiated and were more often diagnosed in the subsequent screening round than by clinical diagnosis as interval cancers. If all retrospectively visible interval cancers had been diagnosed by screening 19% (10/54) of the interval cancers could have been avoided. If all retrospectively visible cancers had been diagnosed at the time of false-negative screening or assessment 65% (84/130) of all patients would have benefitted from an earlier diagnosis compared with the actual figure of 31% (41/130). PMID- 10533458 TI - Antitumour immunity of Bacillus Calmette-Guerin and interferon alpha in murine bladder cancer. AB - Intravesical Bacillus Calmette-Guerin (BCG) immunotherapy is currently the optimal choice for aggressive superficial bladder cancer, with a 70% response rate. This study investigated whether the antitumour response elicited by BCG could be improved by the addition of recombinant interferon alpha (IFN alpha) in the subcutaneous murine MB49 bladder tumour model. The combination of BCG and IFN alpha had superior and earlier antitumour activity than BCG alone for MB49 cells in culture. A total of 14/15 BCG plus interferon-treated mice and 8/16 BCG treated mice became tumour free after treatment. BCG or the combination treatment significantly raised the T-helper 1 (Th1) cytokine IFN gamma levels compared with levels in all other groups. Whilst BCG therapy alone increased CD4+ and CD8+ populations in spleens, the combination of BCG and IFN alpha also increased alpha beta+ T cells significantly. Our results suggest that the combination of BCG and IFN alpha may represent a more efficacious therapeutic than BCG alone for superficial bladder cancer. PMID- 10533460 TI - Superior gene transfer into solid tumour cells than into human mobilised peripheral blood progenitor cells using helpervirus-free adeno-associated viral vector stocks. AB - Autologous peripheral blood progenitor cell (PBPC) grafts can be contaminated with tumour cells that potentially give rise to relapse following myeloablative therapy and PBPC transplantation. Adeno-associated virus (AAV)-based vectors produced by a new adenovirus-free technique are a gene delivery system which may be applicable for tumour cell purging. To test for the host range of these vectors, solid tumours of clinical relevance and normal CD34+ PBPC were selected as target cells for an AAV-vector, encoding the green-fluorescent protein (GFP) as the indicator gene. At a multiplicity of infection (MOI) of 100: 79.94% +/- 14.36% (mean +/- SEM) of the connective tissue sarcoma cell line (HS-1) and 64.84% +/- 6.91% of the cervical carcinoma cell line cells (HeLa-RC) expressed GFP while the other cell lines tested (1 ovarian tumour, 1 germ cell tumour, 1 osteosarcoma, 2 small cell lung cancer) ranged between 2.82% and 11.94%. Optimising the transduction protocol by use of higher MOIs of up to 500 and by pretreatment with the tyrosine kinase inhibitor, genistein, resulted in up to 95.97% and 94.10% green-fluorescent HS-1 and HeLa-RC cells, respectively. In contrast, only 1.39% +/- 0.51% of the normal haematopoietic CD34+ progenitor cells expressed GFP at a MOI of 100. The differential infectivity between HS-1 and CD34+ cells was maintained after tumour cell spiking in leucapheresis products. Our observations suggest that AAV-based vectors may prove useful for purging of autologous PBPC grafts from solid tumour cells. PMID- 10533459 TI - ATM heterozygote cells exhibit intermediate levels of apoptosis in response to streptonigrin and etoposide. AB - Ataxia-telangiectasia (A-T) is a rare recessive disease characterised by cerebellar ataxia, immunodeficiency, sensitivity to ionising radiation and increased cancer risk. Heterozygotes have an increased risk of cancer and may comprise 1% of the population. In vitro, A-T heterozygote cell lines show radiosensitivity intermediate between normal and A-T homozygotes. Furthermore, in A-T homozygotes, hypersensitivity to chemical agents which cause DNA damage, similar to that produced by ionising radiation, has been observed. To investigate the chemosensitivity of A-T heterozygote cell lines, we used TUNEL to analyse the level of apoptosis after drug treatment with etoposide and streptonigrin. Our samples included four normal, eight A-T heterozygote and 10 A-T homozygote lymphoblastoid cell lines. All cell lines were exposed to drugs for 24 h, then cultivated in fresh media for 0 and 72 h. The levels of apoptosis increased significantly in all cell lines, with the greatest increase in homozygote cells and an intermediate increase in heterozygote cells (P values of < 0.01 for etoposide treatment and < 0.02 for streptonigrin treatment were obtained using the Kruskal-Wallis H-test). Our results indicate that A-T heterozygotes express intermediate sensitivity to etoposide and streptonigrin similar to that observed in response to ionising radiation. PMID- 10533461 TI - Superior therapeutic efficacy of N-L-leucyl-doxorubicin versus doxorubicin in human melanoma xenografts correlates with higher tumour concentrations of free drug. AB - N-L-leucyl-doxorubicin (Leu-DOX), a prodrug of doxorubicin (DOX), has previously shown antitumour activity against human ovarian, breast and lung carcinomas in nude mice. In the present study, the efficacy of Leu-DOX was compared with free DOX in inhibiting the growth of four DOX-sensitive and -resistant malignant melanoma xenografts. In an attempt to elucidate mechanisms underlying any differential effect, a sensitive high-performance liquid chromatography (HPLC) method was established for measuring plasma and tumour concentrations of the two drugs and their main metabolites. Leu-DOX was more effective than free DOX in inhibiting xenograft growth. At equitoxic intravenous doses of Leu-DOX (28 mg/kg) and DOX (8 mg/kg) administered to tumour-bearing nude mice, comparable levels of DOX were found in plasma, whereas differences were seen in tumour tissue concentrations. Thus, in animals carrying highly sensitive (LOX) and resistant (THX) melanomas, higher tumour concentrations of free DOX were observed in the Leu-DOX treated group from 24 up to 240 h after drug injection. Notably, the difference in drug-induced tumour growth inhibition correlated with differences in tumour exposure to free DOX, assessed as area under the curve (AUC) calculated over the first 48 h. In conclusion, the results confirm the prodrug nature of Leu DOX and provide a possible explanation for its increased antitumour efficacy. PMID- 10533462 TI - Cisplatin (CDDP)-induced radiation resistance is not associated with CDDP resistance in 86HG39 and A172 malignant glioma cells. AB - Malignant gliomas are often treated with cisplatin (cis diamminedichloroplatinum(II), CDDP) and radiation but results remain unsatisfactory. The development of resistance might explain the poor prognosis. The aim of this study was to investigate whether two human malignant glioma cell lines, 86HG39 and A172, develop resistance to CDDP and/or radiation after CDDP pretreatment. The cells are incubated three times with 10(-6) M CDDP for 24 h, allowed to recover from the treatment and then tested for sensitivity to CDDP and radiation (9 Gy, 60Co) using a colorimetric assay (MTT). A172 pretreated and wild type cells showed no difference in sensitivity to CDDP, whilst 86HG39 cells became more sensitive following pretreatment. This indicates that no resistant phenotype towards CDDP developed in any of the cell lines. In contrast, the CDDP pretreated cells, after radiation, had significantly higher growth rates compared with the wild-type cells in both cell lines (A172: 4.4 +/- 0.5 versus 2.5 +/- 0.3, respectively, 192 h after radiation; 86HG39: 6.2 +/- 0.7 versus 2.3 +/- 0.3, respectively, 216 h after radiation; P < 0.05) indicating resistance against radiation. The level of glutathione (GSH), which is known to mediate resistance against radiation, was 157.2 +/- 61.3 ng/mg protein in A172 cells and 93.2 +/- 16.9 ng/mg protein in 86HG39 cells, and there was no significant difference between levels in wild-type and pretreated cells (A172: 130.1 +/- 34; 86HG39: 81.6 +/- 10.4). These data show that CDDP pretreatment can induce resistance against radiation in vitro independently of CDDP cross-resistance mediated by a mechanism different from GSH. PMID- 10533464 TI - Comments on: Survival of colorectal cancer patients in Europe during the period 1978-1989, Gatta, et al., Eur J Cancer 1998, 34, 2176-2183. PMID- 10533463 TI - A mechanism behind the antitumour effect of 6-diazo-5-oxo-L-norleucine (DON): disruption of mitochondria. AB - 6-diazo-5-oxo-L-norleucine (DON) exerts a growth inhibitory effect selectively on the neuroendocrine tumour cell line BON and is proposed as an antitumour drug. The mechanism behind this has not yet been clarified. In the present study, transmission electron microscopy was used for the assessment of changes in cellular organelles. Furthermore, the methylthiazolyldiphenyl tetrazolium (MTT) assay for mitochondrial enzymatic activity, a fluorescent marker (rhodamine 123) for mitochondrial integrity and [2-(11)C]-acetyl-carnitine which is a substrate of the tricarboxylic acid cycle of mitochondria were employed. The studies were performed in parallel in BON and in a neuroblastoma cell line LAN, with the cells grown as monolayers or as multicellular aggregates. Severe morphological changes of intracellular organelles were observed in BON aggregates treated with low doses of DON. Especially striking was the disruption of mitochondrial internal membrane structures. Other features included the swelling of endoplasmic reticulum, autophagocytosis of secretory granules and nuclear condensation (apoptosis). In LAN cells, no ultrastructural changes were seen after DON treatment. The MTT assay indicated inhibition of mitochondrial enzymatic activity in BON cells but not in LAN cells after 5 h treatment with DON. The mitochondrial damage was also demonstrated as a reduced metabolism of [2-(11)C]-acetyl carnitine. The observations revealed mitochondrial damage by DON treatment and suggest that the mitochondria might be a primary target for the antitumour effect in neuroendocrine cells. PMID- 10533465 TI - Comments on: Treatment of metastatic malignant melanoma with dacarbazine plus fotemustine, Seeber, et al., Eur J Cancer 1998, 34, 2129-2131. PMID- 10533466 TI - Gene therapy for cancer. PMID- 10533467 TI - Does breast cancer exist in a state of chaos? PMID- 10533468 TI - Non steroidal anti-inflammatory drugs and colorectal cancer: is there a way forward? AB - Non steroidal anti-inflammatory drugs (NSAIDs) have diverse clinical applications through modulation of oxidative processes and cell signalling. Observations that these agents may inhibit human colorectal carcinogenesis have produced great excitement. However, comparative data relating to their chemopreventative effectiveness or to relevant mechanisms of action remains unclear. This review considers the clinical and epidemiological evidence for colorectal tumour prevention by NSAIDs against current concepts of drug mechanisms. We also propose areas of further research for potential therapeutic advancement. PMID- 10533469 TI - The immunohistochemical expression of desmoplakin and its role in vivo in the progression and metastasis of breast cancer. AB - Desmoplakin (DP) is a protein located at the inner plaque of desmosomes where it associates with the desmosomal cadherins to form a cell adhesion complex. Reduced expression of DP has been correlated with the progression of several cancers, but its role in in vivo breast cancer is yet to be established. The aim of this present paper was to determine the immunohistochemical (IHC) expression of DP in breast cancer specimens (n = 75) in comparison with ductal carcinoma in situ (DCIS) (n = 26), tumour associated normal (n = 29) and normal breast tissue (n = 7). DP expression was correlated with that of desmosomal cadherin, Desmoglein 2 (Dsg2) and other clinical and IHC prognostic markers. DP staining occurred at the sub-plasma membrane level. There was no significant correlation between the level of DP (as assessed by the H-score) and that of Dsg. Significantly stronger staining was demonstrated in normal breast tissue and well differentiated tumours compared with more moderately or poorly differentiated tumours (P = 0.04). A significant inverse correlation was seen between DP staining and tumour size (P = 0.01). In a limited series of 8 cases, primary tumours demonstrated significantly stronger staining than the matched metastatic lymph nodes (P = 0.046). Of all the IHC markers examined, only Ki-67 showed a significant inverse relationship with DP staining (P = 0.01). In summary, the data suggest that loss of DP may be of potential importance in progression of breast cancer in vivo from normal, DCIS, well differentiated through to poorly differentiated, large tumours. In addition, this loss may be associated with metastasis. PMID- 10533470 TI - The prognosis of small primary breast cancers. AB - The Nottingham Prognostic Index (NPI) is an integrated prognostic index used to predict patient survival for women with invasive breast cancer. The index is based on invasive tumour size, histological lymph node stage and tumour grade. The value of such an index has been questioned in small invasive breast cancers and it has been suggested that size is the only necessary prognostic determinant. The aims of this study were to determine the extent of regional lymph node involvement and survival in women with small invasive breast cancers and to assess the value of the NPI. Between 1976 and 1994, 2684 women aged < or = 70 years were treated for primary operable invasive breast cancers of < or = 5 cm in maximum diameter, of which 318 measured < or = 1 cm. Follow-up data were evaluated to determine histological factors important in predicting survival outcomes in women with cancers < or = 1 cm in diameter and comparing their survival according to the NPI with all women treated for primary operable breast cancers < or = 5 cm in maximum diameter. Histological lymph node involvement was demonstrated in 56/318 (18%) of cancers of < or = 1 cm in diameter. Significant survival differences were demonstrated for small breast cancers according to lymph node stage, vascular invasion and histological tumour grade. Only lymph node stage and histological tumour grade were independent prognostic indicators using a multivariate Cox model. The survival curves for small tumours stratified by the NPI were similar to those of cancers up to 5 cm in diameter. The results indicate that lymph node staging and histological grading are still important prognostic determinants for breast cancers < or = 1 cm in diameter. An axillary node staging procedure should be performed for all invasive breast cancers < or = 1 cm in diameter. The NPI remains relevant for small breast cancers. PMID- 10533471 TI - Quality of life in postmenopausal patients with breast cancer after failure of tamoxifen: formestane versus megestrol acetate as second-line hormonal treatment. Swiss Group for Clinical Cancer Research (SAKK). AB - The Swiss Group for Clinical Cancer Research (SAKK) compared efficacy and toxicity of formestane (250 mg intramuscularly (i.m.) every 2 weeks) versus megestrol acetate (MGA; 160 mg orally daily) as second-line treatment in postmenopausal patients with advanced breast cancer and disease progression while on tamoxifen treatment in a randomised trial (Thurlimann B, Castiglione M, Hsu Schmitz SF, et al. Eur J Cancer 1997, 33, 1017-1024). Quality of life (QL) was evaluated as a secondary endpoint (n = 177). Overall, 83% (669/805) of expected QL forms were received, 88% (155/177) at baseline, 88% (402/457) on study treatment, and 65% (112/171) at treatment failure. Patients with no impairment in performance status reported better physical well-being (P = 0.0001), mood (P = 0.0007) and coping (P = 0.03), and less tiredness (P = 0.0001) and appetite/sense of taste disturbance (P = 0.0001) at baseline. After adjustment for baseline, there was no statistically significant difference in QL by treatment. Baseline QL was strongly predictive for QL under treatment but not for time to treatment failure. In conclusion, the question of whether oestrogen deprivation (e.g. formestane) or addition of progesterone (MGA) has a more beneficial impact on QL needs further investigation. The subjective experience of second-line endocrine treatment varies considerably as a consequence of the large variation in the individual course of the disease and has to be judged on an individual basis. PMID- 10533472 TI - Causes for increased myelosuppression with increasing age in patients with oesophageal cancer treated by chemoradiotherapy. AB - The aim of this study was to identify why increasing myelosuppression accompanies increasing age in patients treated for oesophageal cancer by chemoradiation. Weekly neutrophil and platelet counts were obtained throughout treatment in 86 patients undergoing chemoradiation without surgery for oesophageal cancer. One or two cycles of cisplatin 80 mg/m2/day followed by 5-fluorouracil 800 mg/m2/day for 4-5 days were administered during the first and fourth or fifth week of radiotherapy using 2 Gy daily fractions. 44 of the patients underwent 5 fluorouracil pharmacokinetic studies. Multiple regression procedures were used to determine the strength of factors that contribute to initial and nadir neutrophil and platelet counts. The kinetics of myeloid response were evaluated from the rates of disappearance and re-appearance of neutrophils and platelets during treatment. Age, fluorouracil dose (or AUC), baseline body weight and neutrophil (or platelet) count were found to be powerfully and independently predictive of both first neutrophil and platelet nadir count. Baseline neutrophil and platelet counts were also found to correlate negatively with advancing age independently of other factors. The rate of descent of both indices, however, was independent of age, baseline count and fluorouracil dose suggesting that variations in the size of the myeloproliferative compartment prior to treatment were responsible for interpatient variations. In addition, the rate of recovery of both indices was not influenced by age amongst patients in whom data was assessable suggesting that proliferation of surviving marrow elements is not compromised by age. These data are compatible with the hypothesis that a progressive depletion of the myeloid stem cell compartment accompanies advancing age, and that this is responsible for increasing myelotoxicity. PMID- 10533473 TI - Surgery plus chemotherapy or chemotherapy alone for primary intermediate- and high-grade gastric non-Hodgkin's lymphoma: the Royal Marsden Hospital experience. AB - Primary gastric lymphomas (PGL) have traditionally been treated with surgery followed by chemotherapy or radiotherapy. Surgery was thought to improve staging, optimise local disease control and reduce risk of perforation or bleeding, but recent studies question its role. In this study, patients with intermediate- or high-grade PGL who received chemotherapy from 1985 to 1996 at the Royal Marsden Hospital were identified using a prospectively accrued database. A total of 37 patients (6 with low-grade mucosa-associated lymphoid tissue lymphoma (MALT-L), 9 with high-grade MALT-L, 20 with diffuse large B-cell (DLBC) lymphoma and 2 other histologies), 17 of whom had localised disease, were treated with either surgery plus chemotherapy or chemotherapy alone. 5-year overall survival for localised and advanced PGL was 94 and 50%, respectively, with no differences between the two treatments over a 53 months median follow-up. No perforations or serious bleeding occurred. Surgery is associated with important morbidity and we detected no benefit of surgery prior to chemotherapy in this limited series of patients. PMID- 10533474 TI - Multidrug resistance-associated protein (MRP) expression is correlated with expression of aberrant p53 protein in colorectal cancer. AB - Multidrug resistance-associated protein (MRP) is one of the major factors responsible for non-P-glycoprotein (Pgp)-mediated multidrug resistance of human tumour cells. In this study, we examined MRP and aberrant p53 expression in 54 colorectal cancers (CRC), 35 carcinoma in adenomas (CIA) and 40 adenomatous polyps by immunohistochemical procedures. 38 of 54 (70%) CRCs, 16 of 35 (46%) CIAs and 3 of 40 (8%) adenomatous polyps were MRP positive (chi 2 test, P < 0.0001). 36/54 (67%) CRCs, 10/35 (29%) CIAs and 0/40 adenomatous polyps were p53 positive. 30 of the 36 p53-positive CRCs were also MRP positive and 8/10 CIAs were both p53 and MRP positive. MRP overexpression correlated with aberrant p53 accumulation in CRCs and CIAs (chi 2 test, P < = or 0.01). Coexpression of MRP and p53 in the same cells was confirmed in the CRCs and CIAs by double staining procedures. These results suggested that MRP overexpression is related to aberrant p53 expression in CRC. PMID- 10533475 TI - Development of a disease specific quality of life (QoL) questionnaire module to supplement the EORTC core cancer QoL questionnaire, the QLQ-C30 in patients with pancreatic cancer. EORTC Study Group on Quality of Life. AB - There is overwhelming consensus that quality of life assessment is urgently required in pancreatic cancer, yet little research has been conducted. We report on the development of a disease specific questionnaire module to supplement the EORTC core cancer module, the QLQ-C30 in patients with pancreatic cancer, using EORTC quality of life study group guidelines for module development. Relevant QoL issues were generated from literature searches and interviews with health professionals and patients with pancreatic cancer. Issues were constructed into items and provisionally translated. The provisional module was pretested in patients in 8 European centres. The resulting module the QLQ-PAN26 includes 26 items related to disease symptoms, treatment side-effects and emotional issues specific to pancreatic cancer. This should ensure that the module will be sensitive to assess the small but important disease and treatment related QoL changes in pancreatic cancer. The use of the QLQ-C30 and QLQ-PAN26 will provide a comprehensive system of QoL assessment in international trials of pancreatic cancer. PMID- 10533476 TI - Microsatellite instability is rare in rectal carcinomas and signifies hereditary cancer. AB - We analysed microsatellite instability (MSI) in a consecutive series of 165 rectal carcinomas. Data on a personal and/or family history of cancer were collected from all patients and revealed metachronous cancer in 9 patients, 2 of whom had developed colorectal cancer, and a suspected familial aggregation of colorectal cancer in three families. Only three of the 165 (2%) rectal cancers showed MSI. The patients whose tumours displayed MSI had clinical histories suggesting hereditary cancer--a family history of colorectal cancer and/or synchronous colorectal cancers. Denaturing gradient gel (DGGE) analysis was used to screen the MSI+ patients for mutations in the hMLH1 and hMSH2 genes and revealed two new germline mutations; a 1 bp deletion in exon 10 of hMSH2 creating a premature stop-codon and a splice donor site mutation in intron 16 of hMLH1. Considering colorectal carcinomas as a group, MSI has been reported to occur in approximately 10-20% of the tumours and thus can not, per se be used for clinical detection of hereditary tumours. This study shows, however, that MSI is rare in rectal carcinomas and when present strongly suggests a hereditary predisposition for colorectal cancer development. PMID- 10533477 TI - Vaccination with HPV16 peptides of patients with advanced cervical carcinoma: clinical evaluation of a phase I-II trial. AB - A phase I-II clinical trial was performed involving vaccination with HPV16 E7 peptides of patients suffering from HPV16 positive cervical carcinoma which was refractory to conventional treatment. Patients receiving the vaccine were HLA A*0201 positive with HPV16 positive cervical carcinoma. The clinical trial was designed as a dose-escalation study, in which successive groups of patients received 100 micrograms, 300 micrograms or 1000 micrograms of each peptide, respectively. The vaccine consisted of two HPV16 E7 peptides and one helper peptide emulsified in Montanide ISA 51 adjuvant. 19 patients were included in the study, no adverse side-effects were observed. 2 patients showed stable disease for 1 year after vaccination; 15 patients showed progressive disease of whom 1 died during the vaccination treatment due to progressive disease; and 2 patients showed tumour-regression after chemotherapy following vaccination. A relative low count of lymphocytes before and after vaccination was present in 11/19 patients indicating that these patients were immunocompromised. This study shows that HPV16 E7 peptide vaccination is feasible, even in a group of patients with terminal disease. This paves the way for vaccinating patients with less advanced disease, whose immune system is less compromised by progressive disease. PMID- 10533478 TI - Liver tumours. AB - Primary hepatic tumours in children represent an heterogeneous group of neoplasms. Malignant tumours are more common (60% of primary liver tumours), but account for only 1.2-5% of all paediatric neoplasms. There are two main types of malignant tumour, those of epithelial origin, hepatoblastoma (HB) and hepatocellular carcinoma (HCC), and the rarer mesenchymal tumours, e.g. rhabdomyosarcoma and undifferentiated sarcoma, (Weinberg AG, Finegold, MJ. Primary hepatic tumours of childhood. Hum Pathol 1983, 14, 512-532). Vascular tumours e.g. haemangioendotheliomas are the most common of the benign tumours followed by mesenchymal hamartoma and the rare hepatic adenoma and focal nodular hyperplasia. This article will concentrate on the malignant epithelial tumours. PMID- 10533479 TI - Effects of a third intensification block of chemotherapy on bone and collagen turnover, insulin-like growth factor I, its binding proteins and short-term growth in children with acute lymphoblastic leukaemia. AB - Children with acute lymphoblastic leukaemia (ALL) have reduced bone turnover caused by the disease itself and early intensive chemotherapy, but the effects of later chemotherapy using different drug combinations are uncertain. We report here a longitudinal study on 9 children with ALL randomised to receive an additional third intensification block of chemotherapy, compared with 9 children receiving continuing chemotherapy over the same period. During third intensification, bone alkaline phosphatase, procollagen type I C-terminal propeptide, the carboxyterminal propeptide of type I collagen, procollagen type III N-terminal propeptide and lower leg length all decreased in response to dexamethasone, then returned to (but not beyond) baseline levels after dexamethasone was stopped and other drugs started. These changes were unrelated to circulating insulin-like growth factor (IGF)-I, IGF binding protein (IGFBP)-3 or IGFBP-2. In all children, bone alkaline phosphatase remained below the population mean throughout. We conclude that dexamethasone decreased bone and soft tissue turnover, probably through direct effects on target tissues. The postdexamethasone phase of third intensification and continuing chemotherapy had no major deleterious effect on collagen turnover, but there was evidence of continuing suboptimal bone mineralisation. PMID- 10533480 TI - Incidence of and survival from upper aerodigestive tract cancers in the U.K.: the influence of deprivation. AB - The aim of the study was to investigate the effect of social deprivation on the incidence of and survival from upper aerodigestive tract (UAT) cancers in the U.K. Incidence was calculated on 25,903 cases of malignant upper aerodigestive tract cancers collected from four cancer registries in the U.K. for the period 1984-1993. A Cox proportional hazard model was used to determine the influence of deprivation, measured in Carstairs quintiles for crude and cause-specific survival on 17,393 of these cases. Patients with UAT cancers who were younger, males or of South Asian origin were more likely to live in a deprived area than in an affluent area. The incidence of UAT cancers in a district was correlated with deprivation score for the district for both men (r = 0.78) and for women (r = 0.60). People who lived in deprived areas had a relative risk of 1.25 (95% confidence interval (CI): 1.15-1.35) of dying from their cancer and of 1.24 (95% CI: 1.13-1.35) of dying from all causes compared with people who lived in affluent areas. People living in deprived areas were more likely to get UAT cancer and were more likely to die from their cancer than people living in affluent areas. PMID- 10533481 TI - Reducing colorectal cancer mortality by repeated faecal occult blood test: a nested case-control study. AB - Randomised trials have shown the efficacy of faecal occult blood testing (FOBT) in reducing colorectal cancer mortality, but observational studies are needed to monitor such efficacy in population programmes. We conducted a nested case control study on a cohort of 21,879 subjects who participated in a colorectal screening programme from 1978 to 1995, undergoing at least one FOBT test. 95 fatal cases of colorectal cancer were eligible for the study. For each fatal case, 5 non-fatal matched controls were randomly selected from the cohort. FOBT screening history was less common among cases than controls. The odds ratio of colorectal cancer mortality among 'attenders' (defined as those who underwent a second FOBT within 2 years of study entry) with respect to 'non-attenders' was 0.64 (95% confidence interval 0.36-1.15). We also computed odds ratios defining exposure as one or more tests in the detectable preclinical period, hypothesising various lengths for the latter, which, however, yielded an efficacy estimate biased towards the null. A strong inverse relationship was observed between mortality and the number of tests, but this phenomenon is interpretable as 'healthy screenee bias'. The results suggest that the potential efficacy in preventing colorectal cancer mortality through annual FOBT screening may be of the order of one third. PMID- 10533482 TI - Cell cycle arrest and induction of apoptosis by beta galactoside binding protein (beta GBP) in human mammary cancer cells. A potential new approach to cancer control. AB - Conflict between mitogenic pressure, as is the case in tumour cells and an imposed inability to proceed through the cell cycle may result in cell death. In the present study we examined the effect of beta galactoside binding protein (beta GBP), a negative growth factor which controls cell cycle transition from S phase into G2, on three human mammary cell lines which differ for oncogenic potential, oestrogen receptor expression and expression of the EGF receptor family. We found that in all cases beta GBP induced a cell cycle block prior to the cells' entry into G2 and that this was followed by progressive apoptotic death. This evidence on epithelial cancer cells parallels previous data on tumour cells of mesenchymal origin and suggests that beta GBP has potential therapeutic implications in the treatment of cancers. PMID- 10533483 TI - Characterisation of a synergistic interaction between a thymidylate synthase inhibitor, ZD1694, and a novel lipophilic topoisomerase I inhibitor karenitecin, BNP1100: mechanisms and clinical implications. AB - We developed a combination protocol for inhibitors of thymidylate synthase (TS) and DNA topoisomerase I (Topo I) that can exert highly lethal effects in vitro against HCT-8 human colorectal cancer cells. The specific schedule was constructed so that a TS inhibitor could induce not only primary DNA damage but also cellular conditions optimal for the efficient action of a Topo I inhibitor. The initial drug treatment consisted of a brief exposure to a quinazoline-based antifolate, ZD1694. After an interval of approximately one cell-doubling time, cells were exposed for 8-24 h to BNP1100, a Karenitecin-class 7-thiomethyl camptothecin, in the presence of 1-10 microM thymidine; the latter acted as a crucial factor to promote the collision of moving replication forks with the drug stabilised DNA-Topo I cleavable complexes even under continuous TS inhibition. Clonogenic analyses confirmed that these mechanistically distinct drugs at clinically achievable concentrations worked in a highly synergistic manner, with a maximum effect abolishing the viability of virtually all cancer cells (> 99.9%). The pretreatment with ZD1694 increased the amount of DNA-bound Topo I by up to 4-fold and the DNA-damaging capability of BNP1100 by up to 15-fold. The possibility of at least four DNA-damaging pathways is proposed which might have resulted from the individual actions of TS and Topo I inhibitors as well as their concerted actions. Taken together, the present findings provided a logically permissible explanation as to why TS and Topo I inhibitors in concerted interactions induced a highly lethal effect which was more than a simple additive effect. Since these drugs are effective specifically on actively proliferating cancer cells, but not on non-cycling G0/G1 cells, this mechanism-based protocol may warrant consideration for clinical verification. PMID- 10533484 TI - HPMA copolymer platinates as novel antitumour agents: in vitro properties, pharmacokinetics and antitumour activity in vivo. AB - The aim of this study was to compare in vitro and in vivo HPMA copolymer platinates with cisplatin in terms of platinum release, toxicity and antitumour activity. N-(2-hydroxypropyl)methacrylamide (HPMA) conjugates containing peptidyl side-chains (Gly-Gly or Gly-Phe-Leu-Gly) terminating in either carboxylate or amino species were prepared. The carboxylate polymeric intermediate was reacted with cisplatin, and the polymeric diamine with potassium tetrachloroplatinate to produce HPMA copolymer platinates of Mw 25,000-31,000 Daltons with a platinum loading of 3-7 wt%. The diglycyl spacer was selected because it is non biodegradable, whereas the tetrapeptide spacer is known to be cleaved by the lysosomal thiol-dependent proteases. In vitro the HPMA copolymer platinates displayed a range of platinum release rates at pH 7.4 and 5.5; from < 5%/24 h in the case of the diamino species which require enzymatic activation, to > 80%/24 h in the case of the carboxylate. Cisplatin and the fast releasing carboxylate species displayed IC50 values of 10 micrograms/ml Pt-equivalent against B16F10 cells in vitro, whereas the slow releasing conjugates were not cytotoxic over the dose range studied. Antitumour activity of HPMA copolymer platinates was first evaluated against L1210 and B16F10 tumours inoculated intraperitoneally (i.p.). When conjugates were administered i.p., the antitumour activity observed against L1210 tumours was within the range seen for free cisplatin (ratio of mean survival of treated animals to mean survival of controls, T/C, 1.20-1.70). Neither cisplatin nor HPMA copolymer platinates were active against intraperitoneal (i.p.) B16F10 tumours when administered i.p. However, when conjugates were administered intravenously (i.v.) to treat subcutaneous (s.c.) B16F10 tumours grown to palpable size, free cisplatin was still not active but the HPMA copolymer platinates bearing carboxylate and diamine platinates showed significant antitumour activity (T/C > 1.35). Throughout these studies, the polymer platinates were 5-15-fold less toxic than cisplatin in vivo. After i.v. administration, the blood clearance of HPMA copolymer platinate was considerably slower (t1/2 alpha approximately 10 h) than seen for free cisplatin (t1/2 alpha < 5 min). HPMA copolymer platinates (15 mg/kg Pt-equivalent) gave rise to an approximately 60-fold increase in Pt AUC in B16F10 tumour tissue than was achieved after administration of cisplatin at its maximum tolerated dose (MTD) (1 mg/kg). PMID- 10533485 TI - All-trans retinoic acid enhances gap junctional intercellular communication among renal epithelial cells in vitro treated with renal carcinogens. AB - Epidemiological and clinical studies imply that retinoids have a chemopreventative action against cancer and can suppress the growth of cancer cells. The regulation of connexin (Cx) expression by retinoids varies among tissues and organs. In this study, we investigated whether all-trans retinoic acid (ATRA) upregulates gap junctional intercellular communication (GJIC) in renal epithelial cells exposed to renal carcinogens. Madin Darby canine kidney (MDCK) cells were incubated with ATRA for 3 days, then briefly exposed to 12-O tetradecanoyl-phorbol-13-acetate (TPA) or renal carcinogens potassium bromate (KBrO3) and dimethylnitrosamine (DMN). ATRA increased the expression of connexin 43 mRNA and protein without affecting Cx 43 phosphorylation and prevented inadequate Cx 43 localisation caused by TPA/KBrO3 or DMN. Consequently, ATRA prevented the disruption of GJIC in MDCK cells. These data suggest that ATRA enhanced GJIC by upregulating Cx 43 expression and that ATRA might be useful for prevention of renal cell carcinoma. PMID- 10533486 TI - Preclinical activity of 17 beta-[N-[N'-(2-chloroethyl)-N'-nitrosocarbamoyl]-L alanyl]-5 alpha-dihydrotestosterone (E91) against tumour colony forming units and haematopoietic progenitor cells. AB - E91 (17 beta-[N-[N'-(2-chloroethyl)-N'-nitrosocarbamoyl]-L-alanyl]-5 alpha dihydrotestosterone) (CNC-ala-DHT) is a newly synthesised alkylating compound consisting of N-[N'-(2-chloroethyl)-N'-nitrosocarbamoyl]-L-alanine (CNC-ala) as the alkylating moiety and of 5 alpha-dihydrotestosterone (DHT) as a steroid carrier molecule. We studied the antitumour activity of E91 (final concentrations: 0.1, 1, 10 and 30 mumol/l) against freshly explanted human tumours, using an in vitro soft agar cloning system. A total of 54 tumour samples was evaluated using 1 h-exposure and 51 tumour specimens were studied using a continuous exposure for 21-28 days. In addition, the compound's activity was compared with other clinically used anticancer agents. After short-term exposure, 49 of 53 evaluable specimens (92%) had adequate colony formation, as compared with 49 of 50 (98%) after long-term exposure. After short-term exposure, E91 exhibited only marginal antitumour activity. However, in long-term exposure experiments, E91 had marked and concentration-dependent antitumour activity (P < 0.001). At concentrations of > 10 mumol/l, E91 was as active as the other clinically used antineoplastic agents and at 30 mumol/l, E91 was significantly more active than 5-fluorouracil (P = 0.041). E91 showed activity against a wide spectrum of tumour types. The highest activity was observed against colorectal carcinomas (3/4 tumour specimens inhibited at 30 mumol/l). Sensitivity was also high remarkable in breast cancer specimens with 3/6 specimens inhibited at 30 mumol/l. In vitro myelotoxicity was less than that of doxorubicin. At 30 mumol/l, E91 induced a reduction of colony forming units-granulocyte macrophage (CFU-GM) to only 53% of control and of CFU-GEMM to 20% of control. We conclude that because of broad activity and reduced myelotoxicity further clinical development of E91 appears warranted. PMID- 10533487 TI - XR3054, structurally related to limonene, is a novel inhibitor of farnesyl protein transferase. AB - In this report, a novel inhibitor of farnesyl protein transferase (FPTase) is described. The compound, XR3054, is structurally similar to farnesol, a component of the reaction in which FPTase catalyses transfer of farnesol pyrophosphate to the CAAX recognition motif on proteins. The compound was selected initially because of its ability to inhibit in vitro farnesylation of CAAX recognition peptides with an IC50 of 50 microM. The farnesylation of p21 ras was reduced in a dose-dependent manner in the presence of XR3054. Similarly XR3054 was able to reduce the anchorage-independent growth of V12 H-ras transformed NIH 3T3 cells in a focus formation assay in soft agar, with an IC50 value of 30 microM, whilst not affecting the anchorage-independent growth of v-raf transformed cells. XR3054 reduced the phosphorylation of p42 mitogen activated protein (MAP) kinase in parental NIH 3T3 cells and V12 H-ras transformed NIH 3T3 cells, but constitutively active v-raf transformed cells showed no reduction in phosphorylation of ERK2 in the presence of XR3054. XR3054 inhibited the proliferation of the prostatic cancer cell lines LnCAP and PC3 and the colon carcinoma SW480 and HT1080 (IC50 values of 12.4, 12.2, 21.4 and 8.8 microM, respectively) but was relatively inactive when tested against a panel of breast carcinoma cell lines. The activity did not relate to the presence of mutant or wild-type ras in the cell lines tested. In conclusion XR3054 inhibits ras farnesylation, MAP kinase activation and anchorage-independent growth in NIH 3T3 transformed with v12 H-ras. Since the antiproliferative effect of the compound is not related to the ras phenotype, XR3054 may also have effects on other cell signalling mechanisms. PMID- 10533488 TI - Dipyridamole-mediated reversal of multidrug resistance in MRP over-expressing human lung carcinoma cells in vitro. AB - Expression of the multidrug resistance-associated protein (MRP) is widespread in human malignancies, high levels are associated with poor prognosis and may be responsible for intrinsic and radiotherapy-induced chemoresistance. In this study, the nucleoside transport inhibitor, dipyridamole (DP), was investigated as a chemosensitiser of MRP. In growth inhibition assays MRP-over-expressing COR L23/R cells were 20 times more resistant to VP16 and doxorubicin compared with the parental COR L23/R human lung carcinoma cells. DP caused an approximately 8 fold sensitisation of the resistant cells and a 2-fold sensitisation of the parental cells. DP enhanced the accumulation of VP16 1.5 to 2-fold in the parental cells, but had only a modest effect on VP16 accumulation in the resistant cells. VP16 efflux was rapid in both cell lines. DP caused a modest and transient inhibition of the initial efflux in the resistant cells but not the parental cells. Incubation with DP caused a progressive decrease in GSH levels which was more rapid and profound in COR L23/R cells than in COR L23/P cells. Thus, chemosensitisation to VP16 by DP in MRP-overexpressing COR L23/R cells appears to be caused by depletion of cellular GSH rather than a direct effect of DP on MRP-mediated drug accumulation and efflux. PMID- 10533490 TI - No association between the presence of a constitutional RB1 gene mutation and age in 68 patients with isolated unilateral retinoblastoma. PMID- 10533489 TI - Inhibition of telomerase activity and cell proliferation by a reverse transcriptase inhibitor in gynaecological cancer cell lines. AB - Telomerase is a ribonucleoprotein which has a RNA template to bind and extend telomere ends, so prolonging the life of tumour cells. The aim of this study was to determine whether transcriptase function of telomerase could be inhibited by the reverse transcriptase inhibitors (RTI); azydothymidine (AZT), dideoxyinosine (ddI) and AZT-5' triphosphate (AZT-TP). We examined their effects on the proliferation of cancer cells and the antitumour effects of cisplatin in vitro. The three agents did not cause major changes in telomerase activity or telomere length in MCAS cells. However, in HEC-1 cells changes in telomerase activity and telomere length were observed that were dependent on the RTI concentration and duration of exposure. ddI and AZT-TP reduced telomerase activity and shortened the length of the telomere. In the presence of RTI, the antitumour effects of cisplatin were enhanced. This was particularly evident in HEC-1 cells where there was a marked reduction in cell proliferation, appearance of morphological changes and senescent-like cells in the presence of ddI or AZT-TP. In MCAS cells, TP53 expression was increased by ddI and AZT-TP, while p21 expression was unchanged. In HEC-1 cells the expression of both TP53 and P21 was increased by ddI. Continuous administration of RTI enhanced the cell growth inhibition of cisplatin. RTI also inhibited the proliferation of some cells. PMID- 10533491 TI - You must be joking! PMID- 10533492 TI - Is healthcare reengineering resulting in union organizing of registered nurses? PMID- 10533493 TI - Conceptualizing nursing work-force redevelopment. PMID- 10533494 TI - Healthcare in Burma. PMID- 10533495 TI - Knowledge workers and knowledge-intense organizations, Part 3. Implications for preparing healthcare professionals. AB - We have outlined a framework for understanding knowledge workers, knowledge intense organizations, and the promise of sophisticated, interdisciplinary knowledge work teams because we believe that healthcare is the quintessential knowledge-based service industry. These changes will revolutionize healthcare. We have choices to make, as individuals, and as leaders of the nursing profession. We can choose to help drive and shape the changes needed to realize the potential of this framework, or we can decide to wait and see what happens. We must find the courage and the vision to move nursing and healthcare into this knowledge intense, interdisciplinary future. We end this series as we started it, with a quote from Peter Drucker, who said: "The best way to cope with the radically changing future is to help shape it." PMID- 10533496 TI - Vertical systems integration. AB - Healthcare organizations are vertically integrating into systems for operational, economic, and quality incentives. These system transformations require that the nurse executive assume new roles, responsibilities, and skills. The authors review the trends in system integration and discuss implications for nurse administrators based on literature and structured interviews with nurse executives who have experience in integrated systems or who are engaged in integrated system planing and development. PMID- 10533498 TI - Quantum leadership. New roles for a new age. AB - The quantum age is upon us, changing everything before our very eyes. No part of human life is unaffected by the age change and its new realities. The impact on management and leadership is just beginning to be defined. If leaders and their organizations are to thrive in the new era, a whole new mind-set and skill-set must emerge in the manager. The author identifies characteristics of management role changes necessary to thrive on the journey into the quantum age. PMID- 10533497 TI - High-resource hospital users in an integrated delivery system. AB - OBJECTIVES: With decreasing healthcare reimbursement, nurse administrators need to aggressively manage care for high-resource users of hospital services to ensure the viability of their healthcare organization. The objective of this study was to (1) investigate frequent Medicare inpatient admission and emergency department users, (2) investigate Medicare day outliers, and (3) examine Medicare reimbursement/charge ratios. BACKGROUND: Although much research has focused on patients who have been readmitted frequently to the hospital, little research has examined patients who are frequent users of both emergency departments and inpatient services. METHODS: In this study, all 4,920 elderly Medicare inpatient admissions and emergency department visits for 1 year in a 222-bed general hospital were included. Patient profiles of two categories of high resource users were created. RESULTS: Results showed the frequent high user group (n = 75), who had six or more combined emergency department and inpatient admissions per year, had cardiac, diabetic, and chronic respiratory conditions, and came to the hospital from their homes. The day outlier profile (n = 148) consisted of older patients who have neoplasms, and respiratory and circulatory diseases. The mean Medicare reimbursement/charge ratio varied for high volume diagnosis-related groups (DRGS.) IMPLICATIONS: From the study, implications include refining case management, monitoring high-resource patients by computer tracking, analyzing high-user trends by several different methods, incorporating many facets of an integrated healthcare delivery into their care, expanding patient, outpatient, and community support programs, and continually monitoring revenue for organizational viability. PMID- 10533499 TI - Developing staffing standards. Statistical considerations for patient care administrators. AB - To demonstrate accountability and responsibility for patient care operations, patient care leaders are re-evaluating staffing standards. Typically, activity studies are conducted and statistical methods correlate patient acuity levels to hours per patient day (HPPD). The authors discuss statistical considerations that allow patient care leaders to evaluate the appropriateness of HPPD generated by activity study data. PMID- 10533500 TI - Influence of nursing management education on management direction and role. AB - The authors discuss how 13 nurse managers in Sweden experienced management direction and the management role before and 1 year after a professional development course in advanced nursing at the masters-degree level. The nurse managers related theoretical knowledge to an increased awareness of further possibilities inherent in managerial positions. They stressed the importance of networking for future support and growth. PMID- 10533501 TI - Changing your patients' attitudes toward healthcare insurance. Payment and coverage decisions should be their responsibility. PMID- 10533502 TI - Pharmaceutical representatives' role questioned. PMID- 10533503 TI - How much steroid for poison ivy? PMID- 10533504 TI - What technology has brought to the telephone. You can do a lot more than call home these days. PMID- 10533505 TI - Follow-up to initial episode of atrial fibrillation. PMID- 10533506 TI - 'Hot tub folliculitis'. Test the waters--and the patient--for Pseudomonas. AB - A healthy 10-year-old boy presented with a widespread, nonpruritic, pustular rash of 3 days' duration. He reported having no fever, chills, or other systemic symptoms. Physical examination revealed perifollicular pustules concentrated in the swimsuit area, with fewer widely scattered lesions on the trunk and extremities (figure 1). His mother reported that he had bathed in a hot tub on the evening before onset of the rash. A similar rash had developed in two other boys who had bathed with the patient. No laboratory tests or cultures were performed. The patient was empirically treated with erythromycin tablets, 250 mg twice daily for 10 days. His mother was instructed to contact the owners of the hot tub so that it could be properly cleaned and maintained. Within 1 week the rash had completely resolved. PMID- 10533507 TI - Atopic dermatitis. Perspectives on a manageable disease. AB - Although there is no cure for atopic dermatitis, it can be successfully controlled in most patients. Topical corticosteroids are usually effective, but some patients with recalcitrant disease require phototherapy or use of more potent immunosuppressive agents. Improved understanding of the immunologic basis of atopic dermatitis may provide opportunities for the development of new therapies. PMID- 10533508 TI - The many 'faces' of Graves' disease. Part 1. Eyes, pulse, skin, and neck provide important clues to diagnosis. AB - The variety in clinical appearance of patients with Graves' disease suggests that predominant symptoms affecting one body system may be either compelling or misleading to clinicians during the initial assessment. However, each patient in these case presentations had additional bedside evidence of many of the recognized multisystem manifestations of hyperthyroidism, as detected on more extensive evaluation. Laboratory tests confirmed the diagnosis in each instance. While these cases are not meant to represent all reported manifestations of Graves' disease, grouping of symptoms into memorable "faces" is helpful for definitive clinical recognition and precise diagnosis of this complex and challenging syndrome. PMID- 10533509 TI - Regional anesthesia. Nerve blocks of the extremities and face. AB - Regional nerve blocks are useful for anesthetizing the hand and fingers, the foot and toes, and the face and mouth. These simple procedures may be performed prior to wound repair or surgery of the affected area. The authors discuss the indications, techniques, dosages, and potential complications of regional anesthesia. PMID- 10533510 TI - New treatments for rheumatoid arthritis. Available and upcoming slow-acting antirheumatic drugs. AB - No single therapeutic agent has been found to be universally effective for rheumatoid arthritis, so regimens using combinations of drugs have become the rule. Recently, several new agents with unique mechanisms of action have been introduced and found to produce various degrees of clinical benefit. Among these agents are folate and purine antagonists, alkylating agents, and antipyrimidines. Chimeric (mouse/ human) monoclonal antibody to tumor necrosis factor-alpha and human recombinant interleukin-1 receptor antagonist await approval for general use but have undergone considerable study. PMID- 10533511 TI - When to try COX-2-specific inhibitors. Safer than standard NSAIDs in some situations. AB - COX-2-specific inhibitors, by sparing COX-1 enzyme and its physiologic functions, are a safer option than regular NSAIDs in patients who are at risk for gastrointestinal bleeding (e.g., patients with a history of peptic ulcer disease, gastritis, alcoholism, concomitant corticosteroid or anticoagulant use). They have been approved for use in arthritis, and their efficacy is comparable to that of other NSAIDs. Further clinical data are needed to establish the long-term safety profile of these newly introduced drugs. PMID- 10533512 TI - A practical approach to gout. Current management of an 'old' disease. AB - Gouty arthritis is the culmination of a number of physiologic mechanisms that ultimately result in deposition of uric acid within joints and soft tissues. Decreased uric acid clearance through the kidney is the most common cause of gout. Tophaceous gout occurs in less than 10% of patients. Acute episodes are treated with NSAIDs or colchicine. Low-dose therapy with these agents can also prevent recurrent attacks. Most patients with gout need long-term treatment with either uricosuric agents or xanthine oxidase inhibitors. PMID- 10533513 TI - Getting control of osteoarthritis pain. An update on treatment options. AB - Osteoarthritis consists of a heterogeneous group of disorders that result in articular cartilage degeneration and is diagnosed on the basis of clinical findings. Pathogenesis involves an imbalance between the synthetic and degradative processes that occur in joints. Current interest in the role of cytokines and metalloproteinases may lead to improved treatment of osteoarthritis. For now, management consists of combinations of pharmacologic and nonpharmacologic therapies. A general pharmacologic approach is to begin with acetaminophen and add a low-dose NSAID, nonacetylated salicylate, selective COX-2 inhibitor, or topical capsaicin cream if needed. If pain persists, full-dose NSAID therapy, with the addition of a protective agent in patients at risk for gastrointestinal bleeding, or full-dose COX-2 inhibitor therapy may be tried. Joint injections, irrigation, or arthroscopy may be beneficial in some cases. In patients who continue to have pain and limited function despite these measures, surgical intervention should be considered. PMID- 10533514 TI - Thoracoscopic lung biopsy. Five commonly asked questions about video-assisted thoracic surgery. AB - VATS has proved to be an extremely useful diagnostic tool. Perhaps its most frequent application has been in lung biopsy to diagnose indeterminate solitary pulmonary nodules and interstitial infiltrates. In many institutions, VATS procedures have largely replaced previous methods of attempting to establish the nature of a solitary pulmonary nodule. In ambulatory patients with indeterminate infiltrates, VATS techniques have prompted earlier referral to establish a tissue diagnosis, with apparently decreased morbidity. VATS has clearly found a place in the modern practice of thoracic surgery and is likely to play an ever-increasing role in the management of diseases of the chest. PMID- 10533515 TI - Link between clove cigarettes and urticaria? PMID- 10533516 TI - Assessing the aorta with transesophageal echocardiography. Update on imaging capabilities with today's technology. AB - Improved ultrasound imaging with transesophageal echocardiography is now readily available, so prompt, portable, convenient, and safe assessment of aortic disease is possible. The procedure offers a diagnostic approach that may be more useful and less expensive than other imaging studies. Many experts recommend transesophageal echocardiography as the procedure of choice in patients with possible aortic dissection, especially those who are in clinically unstable condition. Most patients can successfully undergo transesophageal echocardiography. The procedure is also an excellent method for evaluating the success of surgical repair of aortic dissection, and it can be used in long-term patient follow-up. Transesophageal echocardiography can assess atherosclerotic lesion size and composition, the dynamic effect of flow, and the aortic intima and lumen in evaluation of aortogenic embolization. It may help identify patients at high risk of stroke during cardiac surgery. In addition, the procedure is sensitive and specific in diagnosing penetrating aortic ulcers, which occur when an atheromatous lesion pierces the internal elastica and extends into the aortic wall media. The process may cause formation of intramural hematoma, identified on transesophageal echocardiography as homogeneous mottled thickening of the aortic wall and inward displacement of intimal calcification. PMID- 10533517 TI - Motion sickness. How to help your patients avoid travel travail. AB - Motion sickness is an exceedingly common disorder about which primary care physicians are likely to be consulted for advice and treatment. Appropriate management should be based on patient characteristics and the type and length of the exposure and should include general preventive recommendations and directed pharmacologic agents. Education for patients about the causes of motion sickness and how to prevent it can alleviate anxiety and enhance their enjoyment of travel and recreation. PMID- 10533518 TI - Bleeding problems in patients with liver disease. Ways to manage the many hepatic effects on coagulation. AB - Liver disease is associated with a wide variety of abnormalities of the coagulation system and often with severe bleeding. Knowledge of the effects of liver disease and its complications on the various clotting factors may be useful in differential diagnosis. Therapy is difficult but should be approached in a stepwise fashion, with attention to the potential hazards of each intervention. The coagulation system and its interrelationship with liver function can be complex. To ensure the best care of patients with liver disease and bleeding, primary care physicians should not hesitate to enlist the assistance of specialists in clotting disorders and a reliable coagulation laboratory. PMID- 10533519 TI - How to spot illicit drug abuse in your patients. AB - Illicit drug abuse continues to become more widespread, especially in teenagers. Therefore, it is important for physicians to recognize the signs and symptoms of abuse in their patients. Drug abuse should be considered in differential diagnosis of many physical and nearly all psychiatric complaints. An understanding of the pharmacologic mechanisms and adverse effects of illicit drugs can enhance overall care of patients who abuse drugs. The primary classes of drugs of abuse--cannabinoids, opiates, stimulants, hallucinogens, and inhalants--produce clinically diverse presentations. By recognizing these unique signs and symptoms, physicians can differentiate between drug-induced conditions any psychiatric illness. PMID- 10533520 TI - The 'sweet killer'. Can you recognize the symptoms of ethylene glycol poisoning? AB - Ingested ethylene glycol is readily absorbed and metabolized into toxic metabolites that can cause CNS depression, cardiopulmonary failure, and renal failure. Thorough history taking, physical examination, and laboratory testing are essential for diagnosis. Careful differential diagnosis is important because symptoms of ethylene glycol poisoning are similar to those of other intoxicants. Early, aggressive treatment with appropriate therapies, such as ethanol therapy, hemodialysis, vitamin cofactors, and antidotal agents, is necessary to prevent permanent disability or death. PMID- 10533522 TI - Hyperthyroidism. PMID- 10533521 TI - A painful precursor to skin necrosis. Ecthyma gangrenosum. PMID- 10533524 TI - Signaling events regulating the neurodevelopmental triad. Glutamate and secreted forms of beta-amyloid precursor protein as examples. AB - During development of the nervous system a common set of signal transduction pathways appear to regulate growth cone behaviors, synaptogenesis and natural cell death, three fundamental processes that comprise the "neurodevelopmental triad". Among the intercellular signals that coordinate the developmental triad in the mammalian brain are glutamate (the major excitatory neurotransmitter) and beta-amyloid precursor protein (beta APP). Localization of ionotropic glutamate receptors to dendritic compartments allows for selective regulation of dendrite growth cones and spine formation by glutamate released from axonal growth cones and presynaptic terminals. Expression of particular subtypes of glutamate receptors peaks during a developmental time window within which synaptogenesis and natural neuronal death occur. Calcium is the preeminent second messenger mediating both acute (rapid remodelling of the microtubule and actin cytoskeletal systems) and delayed (transcriptional regulation of growth-related proteins; e.g., neurotrophins) actions of glutamate. The expression of beta APP in brain is developmentally regulated and it is expressed ubiquitously in differentiated neurons. beta APP is axonally transported and secreted forms of beta APP (sAPPs) are released from neurons in an activity-driven manner. Secreted APPs modulate neuronal excitability, counteract effects of glutamate on growth cone behaviors, and increase synaptic complexity. Acute actions of sAPPs appear to be transduced by cyclic GMP which promotes activation of K+ channels and reduces [Ca2+]i. Delayed actions of sAPPs may involve regulation of gene expression by the transcription factor NF kappa B. Finally, the striking effects of glutamate, neurotrophic factors, and sAPPs on synaptogenesis and neuronal survival in cell culture systems and in vivo suggest that each of these signals plays major roles in the process of natural cell death. The same signalling mechanisms that mediate adapative regulation of neuroarchitecture during brain development appear to play prominent roles in maladaptive neurodegenerative processes in an array of disorders ranging from stroke to epilepsy to Alzheimer's disease. PMID- 10533523 TI - Why do neurotransmitters act like growth factors? AB - It is now well established that neurotransmitters act as growth-regulatory signals for neuronal and non-neuronal cells of both primitive and higher organisms, where they control cell proliferation, motility, survival, growth, differentiation, and gene expression. Many of these actions are reminiscent of the actions of other growth-regulatory signals such as growth factors, neurotrophins, and proto-oncogenes. How, then, do neurotransmitters exert these effects? Although some information is available concerning second messengers activated by these neurotransmitters in developing cells, little is known about subsequent steps involving signal transduction cascades leading to their final outcomes. This review attempts to provide testable hypotheses regarding possible cellular and molecular mechanisms downstream of second messengers activated by neurotransmitters, based on recent insights into signal transduction cascades activated by classical growth-regulatory signals. In many cases, there are clear points of convergence between these pathways, raising the interesting possibility that neurotransmitters and other growth-regulatory signals may cooperate to regulate developmental functions of cells and tissues. PMID- 10533525 TI - Modulation of intracellular calcium in early neural cells by non-NMDA ionotropic glutamate receptors. AB - Non-NMDA ionotropic receptor channels were longtime thought to be impermeable to calcium. There is now increasing evidence that this is not a general feature. Neural cells bearing non-NMDA receptors permeable to divalent cations can be found not only in the adult CNS, but also at surprisingly early stages of development (well before the onset of synaptogenesis). Since modulation of cytosolic calcium is known to trigger numerous transcription, translation, and post-translation mechanisms, molecules acting at non-NMDA ionotropic receptors may profoundly affect the fate of individual cells, brain regions, and finally the whole nervous system. However, present knowledge of transduction mechanisms and possible roles (ranging from transcription of immediate early genes to regulation of cell survival) is still very fragmentary. These receptors can be activated and modulated by endogenous molecules, but also by exogenous naturally occurring or pharmaceutical substances. If significant amounts of these substances can pass the human placental barrier, their consumption or accidental intake during pregnancy may constitute a risk for the developing embryonic and foetal CNS. PMID- 10533526 TI - Serotonin regulation of neurite outgrowth in identified neurons from mature and embryonic Helisoma trivolvis. AB - Neurite outgrowth and growth cone motility are among the many aspects of neuronal development that can be affected by specific neurotransmitters. This was first demonstrated in experiments on identified molluscan neurons that were isolated from mature ganglia and cultured under conditions that promote the regeneration of new neurites. The application of serotonin to a regenerating Helisoma neuron B19 produced an abrupt, reversible cessation of neurite outgrowth and growth cone motility. While this type of response would subsequently be demonstrated for other neurons and neurotransmitters in many different invertebrate and vertebrate species, experiments on Helisoma neurons have continued to play a pivotal role in advancing this field. In this paper, the mechanisms and sites of serotonin action and how these responses are manifested in vivo during embryonic development are discussed. Experiments primarily on neuron B19 have shown that serotonin acts on a novel serotonin receptor that is coupled to the elevation of cyclic AMP. This intracellular messenger directly activates a class of cyclic-nucleotide-gated sodium channels, leading to sodium influx, membrane depolarization, and activation of voltage-gated calcium channels. The resulting elevation of intracellular calcium acts through a calcium/calmodulin-dependent pathway to inhibit neurite outgrowth and growth cone motility. Although the final steps have yet to be completely resolved, they undoubtedly involve calcium-dependent regulation of cytoskeletal components. Regarding the sites of serotonin action, serotonin responses have been localized to growth cones and even filopodia in specific neurons. However, some studies suggest that neurite development may actually be regulated by serotonin in a paracrine, non-localized manner in a surprisingly large percentage of Helisoma neurons. Finally, experiments on Helisoma embryos have investigated how serotonin actually regulates the in vivo development of specific neurons. Pharmacological treatments that reduce the serotonin concentration in embryos affected the neurite morphology and synaptic efficacy of neuron B19 and the amount of neurite branching in embryonic neuron C1. All of these responses were consistent with the primary action of serotonin being the inhibition of neurite outgrowth, as predicted by the original cell culture studies. PMID- 10533527 TI - Neurotransmitters and neurotrophins collaborate to influence brain development. AB - It is well recognized that the neurotrophin family of factors as well as neurotransmitters play critical roles in the ontogeny of the brain. Moreover, a growing literature suggests that these environmental signals do not operate individually, but interact in critical ways to enhance maturation. This review focuses on three brain systems where this collaboration is particularly evident: the cerebellum, the basal forebrain-hippocampus and locus coeruleus-hippocampus. The material presented indicates that cross-talk between neurotransmitters and neurotrophins may be a mechanism common to the development of multiple neuronal groups throughout the central nervous system. Moreover, this cross-talk appears to involve the interaction of both neuronal and glial cell populations. PMID- 10533528 TI - Cholinergic regulation of cortical development and plasticity. New twists to an old story. AB - Cholinergic afferents innervate cerebral cortex during the most dynamic period of neuronal differentiation and synapse formation, suggesting they play a possible regulatory role in these events. A number of in vivo studies have shown over the last decade that alterations in cholinergic innervation during early postnatal development can change various features of cortical ontogeny. In particular, neonatal lesions to basal forebrain cholinergic afferents result in delayed cortical neuronal development and permanently altered cortical cytoarchitecture and cognitive behaviors. Likewise, cholinergic manipulations affect morphological plasticity in cat visual cortex as well as in the somatosensory cortex of rodents. Furthermore, augmentation of cholinergic function by means of perinatal choline treatment enhances cognitive performance in a sex specific manner. Additional indications for a sexual dimorphism in cortical cholinergic innervation and resulting function are gathered from a variety of paradigms. Recent information about effects of NGF, BDNF and NTB-4/5 on cortical morphogenesis and plasticity reveals complex interactions between the cholinergic basal forebrain afferents and this neurotrophin family. Detailed studies on the expression of cholinergic receptor proteins in cortical development and their associated signal transduction pathways strongly point towards a morphogenetic function of muscarinic receptors, in particular. Transient receptor localization in thalamocortical terminal fields and on a variety of other non-cholinergic fiber bundles suggest a cholinergic role in target finding and/or synapse formation for cortical afferents and efferents. We propose a hypothesis regarding the mechanisms for cholinergic regulation of neuronal differentiation and synapse formation on the level of the individual growth cone and discuss possibilities for cholinergic interactions with differential gene expression. We conclude that understanding the precise role of the cholinergic system in cortical morphogenesis and its relationship to neurotrophin function will be of clinical relevance for a number of developmental brain disorders, including Down Syndrome and Rett Syndrome. PMID- 10533529 TI - Somatostatin as a neurotrophic factor. Which receptor/second messenger transduction system is involved? AB - A variety of studies support a trophic role for somatostatin in the developing nervous system, evidenced as stimulation of neurite outgrowth and axonal or neuronal migration in both in vivo and culture models. Cloning experiments have now demonstrated the existence of five subtypes of somatostatin receptor, differentially distributed in the nervous system, differentially linked to specific signal transduction systems and in certain cases differentially expressed during development. The combination of the differential and developmental regulation of expression of both the somatostatin peptides and their receptors thus provides great potential in terms of trophic effects. To substantiate trophic effects of somatostatin, data are presented from two different model systems, cultures of cerebellar granule cells as well as transgenic mice in which somatostatin is expressed under the control of the glial fibrillary acidic protein promoter. Finally, potential receptor subtypes and second messenger systems involved in these trophic effects are addressed. PMID- 10533531 TI - Neurotransmitters and neurodevelopment. Role of dopamine in neurite outgrowth, target selection and specific synapse formation. AB - Neurotransmitters and their receptors appear early during nervous system development and are thought to play important roles in neurite outgrowth, growth cone motility, target cell selection and synaptogenesis. In vivo studies in both vertebrates and invertebrates have shown that the perturbations of embryonic transmitter expression result in abnormal morphological and synaptic development. In vitro studies have further revealed that transmitters are capable of affecting neurite outgrowth and growth cone behaviour. The precise cellular mechanisms by which neurotransmitters affect these developmental steps are, however, poorly defined. In vitro, a presynaptic neuron from the mollusc Lymnaea stagnalis releases dopamine, which induces both growth cone attraction and growth cone collapse of target and non-target cell growth cones, respectively. We propose that the ability of dopamine to differentially affect growth cone motility of two cell types results from a divergence of the dopamine receptor-activated second messenger pathways at the G-protein level. Such transmitter-receptor interactions between growth cones of specific neurons may not only induce changes in the growth cone motility, but may subsequently play an important role in target cell selection and specificity of synaptogenesis. PMID- 10533530 TI - Diversity of the endogenous opioid system in development. Novel signal transduction translates multiple extracellular signals into neural cell growth and differentiation. AB - This review explores the role of individual opioid receptor types and signal transduction pathways on cell growth and differentiation. The findings reviewed herein provide suggestive evidence that while no single opioid receptor or peptide type exclusively regulates growth, depending on the cell type, the activation of all three (mu, delta, or kappa) opioid receptor types can affect maturation in a cell type-dependent manner. Specific developmental responses are determined primarily by how a particular opioid receptor type is coupled to intracellular signaling effectors. Moreover, the coupling of opioid receptors appears to be developmentally regulated, and these protein-protein interactions change during ontogeny. The diversity of opioid receptor types and intracellular effectors may be a mechanism by which individual cells discriminate among different opioid signals, and may permit diverse opioid signals to be translated into a unique developmental logic in distinct neuronal and glial subpopulations. PMID- 10533532 TI - Functional coupling of neurotransmitters with second messengers during cleavage divisions: facts and hypotheses. AB - Current data on the role of classical neurotransmitters as multiple and multifunctional regulators of early embryogenesis are reviewed. It is shown that these developmental regulators are coupled with second messengers. Peculiarities of this prenervous coupling emphasized and are used as the basis for discussing the problem of the evolutionary origin of cell regulatory systems. PMID- 10533533 TI - Poverty, disease progression and employment among people living with HIV/AIDS in Australia. AB - A national survey of 925 people living with HIV/AIDS (PLWHA) in Australia is used to examine the relationship between disease progression, employment status, poverty and economic hardship. While disease progression has some impact on economic hardship, employment status is found to be the strongest determinant of both poverty and economic hardship. The most commonly cited reasons for leaving work were psychosocial (71%), with declining health cited by half of respondents. It is therefore argued that psychosocial issues are at least as important as changes in health in causing unemployment and therefore poverty and economic hardship among PLWHA in Australia. PMID- 10533534 TI - Psychological and social correlates of high-risk sexual behaviour among men and women living with HIV/AIDS. AB - Men and women living with HIV/AIDS who experience difficulty maintaining safer sex practices place their sex partners as well as themselves at considerable risk for sexually transmitted infections. Psychological correlates of continued sexual risk behaviours provide important information for intervention development. Continued sexual risk behaviour was investigated in a sample of 203 HIV-positive men and 129 HIV-positive women recruited from infectious disease clinics and AIDS service agencies. The study showed that 42% of men and 42% of women reported at least one occasion of unprotected anal or vaginal intercourse in the preceding six months. Unprotected intercourse frequently occurred outside of long-term relationships and with partners who were not known to be HIV-infected. Similar to populations at primary risk, HIV-infected men and women reported alcohol and drug use, including use before sexual episodes. However, the association between substance use and unprotected sex was modest for men and absent for women. Contrary to previous research, emotional distress and maladaptive coping were not related to continued sexual risk. Interventions are urgently needed to support men and women living with HIV/AIDS in maintaining long-term safer sex practices. PMID- 10533535 TI - Lack of legal income is strongly associated with an increased risk of AIDS and death in HIV-infected injecting drug users. AB - The aim of the study was to analyze the impact of soci-economic status in addition to other risk factors in the progression of HIV disease in a cohort of injecting drug users (IDUs) with a mean follow-up of two years. Between 1989 and 1992, 124 HIV-infected IDUs were recruited in a primary care outpatient clinic providing free consultations and free access to therapy. The main outcome measures were death and AIDs-defining events. The proportion of current daily injectors at entry in the study and at the end of follow-up was 67.7% and 57.2%, respectively. The proportion of individuals on maintenance opioid therapy at entry in the study and at the end of follow-up was 0 and 12.1%, respectively. CD4 cell counts below 200 x 10(6)/L at baseline, positive p24 antigenemia at baseline, the lack of legal income and occasional drug use at entry were risk factors for clinical progression and death. When adjusted in a multivariate analysis, the absence of legal income remained associated with death and occurrence of an AIDS-defining event with a relative risk of 5.2 (1.5-18.1) (p = 0.004). Lack of legal income is a strong risk factor for progression of HIV disease in IDUs, that is independent of CD4 cell count and p24 antigenemia. PMID- 10533536 TI - Geographic proximity, policy and utilization of syringe exchange programmes. AB - The objective of the research was to assess the effects of geographic proximity on the utilization of syringe exchange among injection drug users (IDUs) in New York City. Between 1994 and 1996, 805 IDUs were interviewed with a structured questionnaire. Geographic proximity was defined as living within a ten-minute walk. Eighty-one per cent of IDUs who lived close typically used a syringe exchange compared to 59% of those who lived further away. In multiple logistic regression analysis, those who lived close remained (adjusted odds ratio of 2.89; 95% CI 2.06 to 4.06, p = 0.001) more likely to use syringe exchange. Those who lived close were less likely to have engaged in receptive syringe sharing at last injection (adjusted odds ratio = 0.45, 95% CI 0.24 to 0.86, p = 0.015). In conclusion, locating exchange services in areas convenient to large numbers of IDUs may be critical for prevention of HIV infection. PMID- 10533539 TI - Development of the Impact of Weight Loss Scale (IWLS): a psychometric study in a sample of men with HIV/AIDS. AB - The purpose of this study was to assess the psychometric properties of the newly developed Impact of Weight Loss Scale (IWLS), a subjective measure of perceived weight loss and related behaviour, body image perception, and affect, as well as the scale's relationship to mental health and nutritional measures in an HIV sample. Seventy-five HIV-positive men were administered the IWLS along with measures of depression, quality of life and nutritional status. Half (51%) of the sample had a CD4 count below 200 cells/cu.mm, and 80% had significant loss of body weight (> or = 10% body weight loss) or body cell mass (< or = 90% of normative body cell mass). The IWLS demonstrated good internal consistency reliability (Cronbach's alpha = 0.88) and had a unidimensional factor structure. Higher IWLS scores, which indicate a more detrimental impact of weight loss, were correlated with greater depression (r = 0.29), reduced quality of life (r = 0.37), and objective nutritional deficits (ratio of body cell mass/height, r = 0.28; amount of weight loss, r = 0.30). The IWLS is a brief self-report measure with good psychometric characteristics and has potential utility in both research and clinical applications. PMID- 10533538 TI - The parent disclosure interview. AB - As the number of HIV-infected women and children in the USA has increased, clinicians and researchers have debated the benefits and risks of disclosure of parental HIV status to children. Disclosure is usually ascertained through interviews of unknown reliability. Given the need to advance knowledge regarding the benefits and risks of disclosure of parental HIV status to children, a reliable and comprehensive disclosure interview is needed. The Parent Disclosure Interview (PDI) was developed for this purpose. In order to study its reliability, 29 HIV-infected mothers were administered the PDI twice, on average one week apart, by two different female interviewers. Kappa statistics indicate that the PDI is highly reliable in most content areas. Researchers may use the interview for comparing the prevalence of disclosure among different groups of HIV-infected parents. Practitioners who assist parents in making decisions about disclosure of HIV status to children may use the interview to obtain a baseline assessment of the clients' disclosure history and attitudes towards disclosure. PMID- 10533537 TI - AIDS-related information exposure in the mass media and discussion within social networks among married women in Bombay, India. AB - Married women are at high risk of acquiring HIV infection in India and health education remains the most feasible preventive tool in their context. In a survey conducted among 350 married women in Bombay, it was found that a majority had acquired information about AIDS from the mass media, especially television. Although 87% of women who knew of AIDS had been exposed to AIDS-related information in the mass media in the past four weeks, only 57% had discussed it within their social networks. Those with more exposure to AIDS information in the mass media were significantly more likely to discuss AIDS within social networks. The women were most likely to discuss AIDS with their husbands as a general social issue, followed by friends and family members and least likely to talk to husbands about AIDS as a personal issue relating to their sexual relationship. Increased frequency and duration of AIDS messages on television will have a positive influence on AIDS knowledge in this group. PMID- 10533540 TI - Knowledge, attitudes and practices concerning HIV/AIDS among sex workers in Phnom Penh, Cambodia. AB - The human immunodeficiency virus (HIV)/AIDS epidemic is currently spreading faster in Cambodia than anywhere else in Asia. Heterosexual transmission of HIV through prostitution is believed to be catalyzing the epidemic, and sex workers (SWs) are at a very high risk for becoming infected with HIV and subsequently developing AIDS. In order to gain a better understanding of the knowledge, attitudes and practices of this highly vulnerable population, face-to-face interviews were conducted with SWs (N = 502) in the capital city, Phnom Penh. The SWs surveyed were predominantly young, uneducated, poor women from rural areas, many of whom remain isolated in brothels. Brothel-based SWs are probably at greatest risk for acquiring HIV. They reported twice as many sexual contacts per day and used condoms less frequently than community-based SWs. The majority of SWs surveyed knew that condoms offered protection against HIV/AIDS, although one quarter of SWs did not always use condoms. Despite their high level of baseline HIV/AIDS knowledge, nearly all SWs requested that additional health education materials be made available to them and their customers. PMID- 10533541 TI - Contrasting strategies used by young people to ensure condom use: some findings from a qualitative research project. AB - Interviews conducted among 56 young men and women (aged 16-19) reveal two contrasting strategies used to ensure condom use at first intercourse with a new partner. These are defined as verbal communication based strategies (involving some explicit discussion about contraception before intercourse) and non-verbal communication based strategies (where one partner takes responsibility for using condoms without discussing this with their partner). Whilst the former is argued as being the more effective strategy, this paper suggests an important role for the latter, particularly when young people find themselves in situations where initiating discussions about condom use is perceived as being particularly difficult. PMID- 10533543 TI - The History of Lunacy and the scholarship of Edward H. Hare. PMID- 10533542 TI - Caring for people with HIV in Zambia: are traditional healers and formal health workers willing to work together? AB - This study of traditional healers and formal health workers determined their knowledge and practices in the field of HIV/AIDS and examined their training needs and attitudes to collaboration, in preparation for planning joint training workshops. Several misconceptions concerning symptoms and transmission of HIV disease were found in both groups, particularly among traditional healers. Twenty healers (51%) and four formal health workers (15%) claimed a cure existed for AIDS. The majority of traditional healers interviewed expressed difficulties discussing a diagnosis of HIV directly with patients, mainly due to fear of the patient becoming depressed and suicidal. Most interviewees wanted more training- the majority of traditional healers in recognizing symptoms of HIV/AIDS and their treatment, and the majority of formal health workers in HIV counselling. Most were interested in supplying condoms. Almost all healers and half of the formal health workers were keen to collaborate in training and patient care. The study indicates that there is willingness amongst Zambian traditional healers and formal health workers to collaborate in training and patient care in the field of HIV/AIDS. As well as covering symptoms, transmission and prevention of HIV/AIDS, training should aim to increase ability to openly discuss HIV with patients, which many traditional healers and some formal health workers find difficult. Involving traditional healers in supplying condoms may improve acceptability and availability, particularly in rural areas. PMID- 10533544 TI - EURODEP Consortium and late-life depression. PMID- 10533545 TI - Hypertension and cognitive decline. PMID- 10533546 TI - 'Subthreshold' mental disorders. A review and synthesis of studies on minor depression and other 'brand names'. AB - BACKGROUND: Subthreshold conditions (i.e. not meeting full diagnostic criteria for mental disorders in DSM-IV or ICD-10) are prevalent and associated with significant costs and disability. Observed more in primary care and community populations than in speciality settings, varying conceptualisations have been applied to define these conditions. AIMS: To examine definitional issues for subthreshold forms of depression (e.g. minor depression) and to suggest future directions for research and nosology in psychiatry and primary care. METHOD: A Medline search was conducted. The relevant articles were reviewed with regard to specific categories of information. RESULTS: Studies applied a myriad of names and definitions for subthreshold depression with varying duration, symptom thresholds and exclusions. Prevalence rates also vary depending upon the definitions, settings and populations researched. CONCLUSIONS: Future research needs to apply methodological and intellectual rigour and systematically consider a broader clinical and nosological context. In addition, collaboration between psychiatry and primary care on research and clinical issues is needed. PMID- 10533547 TI - Effectiveness of antidepressants. Meta-analysis of dose-effect relationships in randomised clinical trials. AB - BACKGROUND: Antidepressant drugs are usually prescribed at low doses, possibly to avoid adverse reactions. No comprehensive review has addressed the issue of dose, clinical response and tolerability in a quantitative way. AIMS: To determine whether high doses of antidepressants are more effective than low doses, and how safety is affected by dose. METHOD: Trials comparing two or more doses of the same antidepressant were located, and all antidepressants administered were converted to the equivalent dose of imipramine. Generalised estimating equations were used to analyse percentage improvement and adverse event rate according to dose level. RESULTS: Thirty-three studies were identified. The dose level 100-200 mg imipramine equivalents showed an average improvement of 53% by 'intention-to treat'. Higher doses were not accompanied by increased efficacy, while lower doses showed reduction in efficacy. Adverse events significantly increased with dose. CONCLUSIONS: With a low dose of antidepressants, clinicians trade off a slightly reduced chance of improvement for a higher chance of avoiding adverse reactions. PMID- 10533548 TI - Depression of older age. Origins of the study. AB - BACKGROUND: The EURODEP collaboration was formed to take advantage of existing studies of random community samples of older people in Europe, using GMS-AGECAT for case identification and diagnosis. Later, other centres joined, and the EURO D scale was developed to harmonise the different methods used with the GMS. Previous studies had revealed different levels of depression in Europe but had been confounded by the use of unreconcilable methods. These studies attempt to overcome this problem. AIMS: To introduce the first set of publications from the EURODEP collaboration. METHOD, RESULTS AND CONCLUSIONS: Presented in five accompanying papers (pp. 307-345, this issue). PMID- 10533549 TI - Review of community prevalence of depression in later life. AB - BACKGROUND: Despite considerable interest, there is no consensus regarding the prevalence of depression in later life. AIMS: To assess the prevalence of late life depression in the community. METHOD: A systematic review of community-based studies of the prevalence of depression in later life (55+). Literature was analysed by level of caseness at which depression was defined and measured. RESULTS: Thirty-four studies eligible for inclusion were found. The reported prevalence rates vary enormously (0.4-35%). Arranged according to level of caseness, major depression is relatively rare among the elderly (weighted average prevalence 1.8%), minor depression is more common (weighted average prevalence 9.8%), while all depressive syndromes deemed clinically relevant yield an average prevalence of 13.5%. There is consistent evidence for higher prevalence rates for women and among older people living under adverse socio-economic circumstances. CONCLUSIONS: Depression is common in later life. Methodological differences between studies preclude firm conclusions about cross-cultural and geographical variation. Improving the comparability of epidemiological research constitutes an important step forward. PMID- 10533550 TI - Depression in Europe. Geographical distribution among older people. AB - BACKGROUND: This is the first report of results from the EURODEP Programme. AIMS: To assess the prevalence of depression judged suitable for intervention in randomised samples of those aged > or = 65 in nine European centres. METHOD: The GMS-AGECAT package. RESULTS: Differences in prevalence are apparent, 8.8% (Iceland) to 236% (Munich). When sub-cases and cases are added together, five high- and four low-scoring centres emerge. Women predominated over men. Proportions of sub-cases to cases revealed striking differences but did not explain prevalence. There was no constant association between prevalence and age. A meta-analysis (n = 13,808) gave an overall prevalence of 12.3%, 14.1% for women and 8.6% for men. CONCLUSIONS: Considerable variation occurs in the levels of depression across Europe, the cause for which is not immediately obvious. Case and sub-case levels taken together show greater variability, suggesting that it is not a matter of case/sub-case selection criteria, which were standardised by computer. Substantial levels of depression are shown but 62-82% of persons had no depressive level. Opportunities for treatment exist. PMID- 10533551 TI - Cross-cultural comparison of depressive symptoms in Europe does not support stereotypes of ageing. AB - BACKGROUND: Stereotypes of older people suggest that they are depressed. AIMS: To examine depression symptoms among people aged > or = 65 in the general population and to ask the following questions. Are there high proportions of depressive symptoms among otherwise well people? Do these levels reflect the prevalence of depression? Do key symptoms vary with age and do they confirm stereotypes? METHOD: Nine centres contributed data from community-based random samples, using standardised methods (GMS-AGECAT package). RESULTS: Proportions of depressive symptoms varied between centres. Some often associated with ageing were rare. Many were more common in women. Low-prevalence centres tended to have fewer symptoms among 'well' people, but there were inconsistencies. Low levels of symptoms among the well population of a centre did not necessarily predict lower levels in the depressed. CONCLUSIONS: Variations in the prevalence of depressive symptoms occurred between centres, not always related to levels of illness. There was no consistent relationship between proportions of symptoms in well persons and cases for all centres. Few symptoms were present in > 60% of the older population--stereotypes of old age were not upheld. PMID- 10533552 TI - Development of the EURO-D scale--a European, Union initiative to compare symptoms of depression in 14 European centres. AB - BACKGROUND: In an 11-country European collaboration, 14 population-based surveys included 21,724 subjects aged > or = 65 years. Most participating centres used the Geriatric Mental State (GMS), but other measures were also used. AIMS: To derive from these instruments a common depression symptoms scale, the EURO-D, to allow comparison of risk factor profiles between centres. METHOD: Common items were identified from the instruments. Algorithms for fitting items to GMS were derived by observation of item correspondence or expert opinion. The resulting 12 item scale was checked for internal consistency, criterion validity and uniformity of factor-analytic profile. RESULTS: The EURO-D is internally consistent, capturing the essence of its parent instrument. A two-factor solution seemed appropriate: depression, tearfulness and wishing to die loaded on the first factor (affective suffering), and loss of interest, poor concentration and lack of enjoyment on the second (motivation). CONCLUSIONS: The EURO-D scale should permit valid comparison of risk-factor associations between centres, even if between-centre variation remains difficult to attribute. PMID- 10533553 TI - Depression symptoms in late life assessed using the EURO-D scale. Effect of age, gender and marital status in 14 European centres. AB - BACKGROUND: Data from surveys involving 21,724 subjects aged > or = 65 years were analysed using a harmonised depression symptom scale, the EURO-D. AIMS: To describe and compare the effects of age, gender and mental status on depressive symptoms across Europe. METHOD: We tested for the effects of centre, age, gender and marital status on EURO-D score. Between-centre variance was partitioned according to centre characteristics: region, religion and survey instrument used. RESULTS: EURO-D scores tended to increase with age, women scored higher than men, and widowed and separated subjects scored higher than others. The EURO-D scale could be reduced into two factors: affective suffering, responsible for the gender difference, and motivation, accounting for the positive association with age. CONCLUSIONS: Large between-centre differences in depression symptoms were not explained by demography or by the depression measure used in the survey. Consistent, small effects of age, gender and marital status were observed across Europe. Depression may be overdiagnosed in older persons because of an increase in lack of motivation that may be affectively neutral, and is possibly related to cognitive decline. PMID- 10533554 TI - Cost-effectiveness of assertive community treatment for homeless persons with severe mental illness. AB - BACKGROUND: Homelessness is a major public health problem among persons with severe mental illness (SMI). Cost-effective programmes that address this problem are needed. AIMS: To evaluate the cost-effectiveness of an assertive community treatment (ACT) programme for these persons in Baltimore, Maryland. METHODS: A total of 152 homeless persons with SMI were randomly allocated to either ACT or usual services. Direct treatment costs and effectiveness, represented by days of stable housing, were assessed. RESULTS: Compared with usual care, ACT costs were significantly lower for mental health in-patient days and mental health emergency room care, and significantly higher for mental health out-patient visits and treatment for substance misuse. ACT patients spent 31% more days in stable housing than those receiving usual care. ACT and usual services incurred $242 and $415 respectively in direct treatment costs per day of stable housing, an efficiency ratio of 0.58 in favour of ACT. Patterns of care and costs varied according to race. CONCLUSION: ACT provides a cost-effective approach to reducing homelessness among persons with severe and persistent mental illnesses. PMID- 10533555 TI - Predictors of chronic post-traumatic stress disorder. A prospective study. AB - BACKGROUND: Most individuals who, shortly after trauma, express symptoms of post traumatic stress disorder (PTSD) recover within one year of their traumatic experiences. In contrast, those who remain ill for one year rarely recover completely. The early identification of the latter is, therefore, very important. AIMS: To prospectively evaluate predictors of PTSD at four months and one year. METHOD: We followed 236 trauma survivors recruited from admissions to a general hospital's emergency room for four months, at which point 41 (17.4%) met diagnostic criteria for PTSD. Twenty-three of these individuals, and 39 individuals without PTSD at four months, were assessed again at one year. RESULTS: Depressive symptoms were the best predictors of PTSD at both time points. Intrusive symptoms and peri-traumatic dissociation were better at predicting four-month PTSD than one-year PTSD. CONCLUSIONS: The occurrence of depression during the months that follow a traumatic event is an important mediator of chronicity in PTSD. PMID- 10533556 TI - Neurobehavioural symptoms one year after a head injury. AB - BACKGROUND: Neurobehavioural symptoms are common immediately after a minor head injury but have not been studied one year after the injury. AIMS: To estimate the rate and pattern of neurobehavioural symptoms one year after a head injury of varying severity. METHOD: Adults who had been hospitalised after a head injury (n = 196, 164 of whom had a face-to-face interview) and showed indirect evidence of brain assault were assessed for the presence of neurobehavioural symptoms with the help of a behaviour rating scale. RESULTS: About 40% had three or more symptoms. Individual symptoms varied among 3% (social disinhibition), 15% (lack of initiative) and 35% (irritability) of the cohort. Premorbid factors such as lower social class and lower educational achievement, head-injury-related factors such a low Glasgow coma score, and outcome-related factors such as the presence of a disability according to the Edinburgh Rehabilitation Status Scale and psychiatric caseness according to the Clinical Interview Schedule--Revised, significantly influenced the rate and the pattern of behavioural symptoms. The pattern of symptoms varied between age groups and according to the severity of the head injury. CONCLUSIONS: A significant proportion of patients with varying degrees of severity of head injury showed behavioural symptoms after one year of head injury. PMID- 10533557 TI - Risperidone treatment of amphetamine psychosis. PMID- 10533558 TI - Amiodarone-induced depression. PMID- 10533559 TI - Psychological treatments for hypochondriasis. PMID- 10533560 TI - Antipsychotic polypharmacy and early death. PMID- 10533561 TI - Shotguns and blunderbusses: suicide in farmers. PMID- 10533562 TI - Satisfaction of carers at home. PMID- 10533564 TI - In memoriam PMID- 10533563 TI - Hair analysis for substance use. PMID- 10533565 TI - A spotlight on electrophysiological remodeling and the molecular biology of ion channels. PMID- 10533566 TI - Electrophysiological remodeling in hypertrophy and heart failure. PMID- 10533567 TI - Electrical remodeling in ischemia and infarction. AB - This is a review of the electrophysiologic changes occurring at different times following myocardial infarction, both in the infarcted region (substrate) and in areas remote from the infarct. Regulators of channel function which might contribute to re-modeling, including autocrine/paracrine factors involved in ion channel gene regulation, are discussed. PMID- 10533568 TI - Atrial electrophysiological remodeling caused by rapid atrial activation: underlying mechanisms and clinical relevance to atrial fibrillation. AB - One of the most exciting developments in our understanding of atrial fibrillation (AF) over the last several years has been the recognition that AF itself modifies atrial electrical properties in a way that promotes the occurrence and maintenance of the arrhythmia, a process termed 'atrial remodeling'. The principle stimulus for AF-induced atrial remodeling is the rapid atrial rate that results: rapid regular atrial pacing produces changes similar to those caused by AF in animal models. The mechanisms of atrial tachycardia-induced remodeling have been extensively explored, and involve changes in atrial electrophysiology associated with altered ion channel function. The most important ionic change is a reduction in L-type Ca2+ current, which reduces action potential duration (APD) and APD adaptation to rate. AF-induced changes in ion channel function appear to be due both to rapid voltage- and time-dependent alterations in channel availability caused by tachycardia and to slower downregulation of messenger RNA concentrations encoding alpha-subunits of specific ion channels. Atrial remodeling likely contributes importantly to a wide variety of clinical phenomena of previously unrecognized mechanism, including atrial dysfunction after cardioversion of AF, the increasing resistance to therapy of longer-standing AF, the association of AF with other forms of supraventricular tachyarrhythmia and the tendency of paroxysmal AF to become chronic. The present paper reviews the state of knowledge regarding the mechanisms and clinical consequences for AF of atrial remodeling caused by rapid atrial activation. PMID- 10533569 TI - Mechanisms of remodeling of gap junction distributions and the development of anatomic substrates of arrhythmias. PMID- 10533570 TI - From genes to channels: normal mechanisms. AB - Electrophysiologic remodeling is a process whereby heart disease alters the electrophysiologic properties of cardiac tissue. These alterations, in turn, can cause or exacerbate disease-related arrhythmias. Ion channels are the fundamental molecular units underlying cardiac electrophysiology, and it therefore follows that electrophysiologic remodeling represents alterations in the function or expression of genes encoding ion channels or other proteins crucial for cardiac electrophysiologic activity. This review will describe the mechanisms whereby normal function of these proteins arises from the processes of gene transcription, mRNA processing, and protein transport, post-translational modification, assembly with other proteins, and degradation. Identification of entirely novel targets for drug intervention should result from further understanding of the fundamental mechanisms underlying remodeling. PMID- 10533571 TI - Structure and function of the cardiac sodium channels. AB - The coincident cloning of the voltage-gated Na channel from the electric eel electroplax and development of patch-clamp methodology has allowed an explosive phase of investigation into the structural basis of cardiac Na channel function. Recognizing the importance of structural motifs that underlie gating (charged S4 segments, III-IV linker) and permeation (P-loops) have complemented new molecular information surrounding inherited cardiac arrhythmias, such as the chromosome 3 linked form of the long QT syndrome. Although the proarrhythmic potential recognized in the CAST trial [1] slowed the development of class I antiarrhythmic agents, our emerging understanding of the molecular pharmacology of Na channels may motivate strategies for Na-channel drug discovery that involve targeting particular structural domains. PMID- 10533572 TI - Ca channels in cardiac myocytes: structure and function in Ca influx and intracellular Ca release. AB - There are Ca channels in the plasma membrane and also the sarcoplasmic reticulum (SR) membrane in cardiac myocytes. The relationship between channel structure, associated proteins and function of these Ca channels is discussed. The sarcolemmal Ca channels are crucial both to the basic cellular electrophysiological properties and control of cardiac contractility (via excitation-contraction coupling). The intracellular Ca release channels (or ryanodine receptors) respond to triggering events mediated by sarcolemmal ion currents and are largely responsible for releasing Ca which activates the myofilaments to produce contraction. Several possible mechanisms of excitation contraction coupling are discussed. The Ca released from the SR can also feedback on several sarcolemmal ion currents and alter action potential configuration as well as contribute to arrhythmogenesis. PMID- 10533573 TI - Insights into the structure, distribution and function of the cardiac chloride channels. AB - This review describes the properties and distribution of the three major types of chloride currents that have been studied in cardiac tissue. These include a cAMP- and protein kinase A-dependent current, a calcium-activated current and a swelling-induced current. The study of cardiac anion currents is a less mature field than the study of cardiac cation currents. Consequently, less is known regarding the structure, molecular identity and physiological role of anion currents in comparison to cardiac cation currents. Where known, the available molecular and structural information is also discussed. Although there is no proven physiological role for cardiac chloride currents, the possible clinical electrophysiological roles of cardiac chloride currents are discussed. PMID- 10533574 TI - Structure and function of cardiac potassium channels. AB - Recent advances in molecular biology have had a major impact on our understanding of the biophysical and molecular properties of ion channels. This review is focused on cardiac potassium channels which, in general, serve to control and limit cardiac excitability. Approximately 60 K+ channel subunits have been cloned to date. The (evolutionary) oldest potassium channel subunits consist of two transmembrane (Tm) segments with an intervening pore-loop (P). Channels formed by four 2Tm-1P subunits generally function as inwardly rectifying K(+)-selective channels (KirX.Y): they conduct substantial current near the resting potential but carry little or no current at depolarized potentials. The inward rectifier IK1 and the ligand-gated KATP and KACh channels are composed of such subunits. The second major class of K+ channel subunits consists of six transmembrane segments (S1-S6). The S5-P-S6 section resembles the 2Tm-1P subunit, and the additional membrane-spanning segments (especially the charged S4 segment) endow these 6Tm-1P channels with voltage-dependent gating. For both major families, four subunits assemble into a homo- or heterotetrameric channel, subject to specific subunit-subunit interactions. The 6Tm-1P channels are closed at the resting potential, but activate at different rates upon depolarization to carry sustained or transient outward currents (the latter due to inactivation by different mechanisms). Cardiac cells typically display at least one transient outward current and several delayed rectifiers to control the duration of the action potential. The molecular basis for each of these currents is formed by subunits that belong to different Kvx.y subfamilies and alternative splicing can contribute further to the diversity in native cells. These subunits display distinct pharmacological properties and drug-binding sites have been identified. Additional subunits have evolved by concatenation of two 2Tm-1P subunits (4Tm 2P); dimers of such subunits yield voltage-independent leak channels. A special class of 6Tm-1P subunits encodes the 'funny' pacemaker current which activates upon hyperpolarization and carries both Na+ and K+ ions. The regional heterogeneity of K+ currents and action potential duration is explained by the heterogeneity of subunit expression, and significant changes in expression occur in cardiac disease, most frequently a reduction. This electrical remodelling may also be important for novel antiarrhythmic therapeutic strategies. The recent crystallization of a 2Tm-1P channel enhances the outlook for more refined molecular approaches. PMID- 10533575 TI - Normal regional distribution of membrane current density in rat left ventricle is altered in catecholamine-induced hypertrophy. AB - OBJECTIVE: To test the hypothesis that changes in the normal regional distribution of potassium and calcium currents contribute to the different regional changes in action potential duration in isoprenaline-induced hypertrophy in rats. METHODS: Hypertrophy was elicited in rats by seven daily injections of isoprenaline. Left ventricular myocytes were isolated from basal sub-endocardial, basal mid-myocardial and apical sub-epicardial tissue. Membrane currents were measured using the whole-cell patch-clamp technique at 35 +/- 1 degrees C. RESULTS: Cell membrane capacitance was similar in all three groups and was increased by 17% in hypertrophy (P < 0.001, t-test). Changes in the calcium independent transient outward current (Ito1) density in hypertrophy were different in the three regions (P < 0.05, ANOVA). Ito1 was reduced in sub epicardial (control, 23.4 +/- 2.0 pA pF-1; hypertrophy, 15.8 +/- 1.5 pA pF-1, P < 0.01 ANOVA) and in mid-myocardial myocytes (control, 24.0 +/- 2.8 pA pF-1; hypertrophy, 13.8 +/- 1.3 pA pF-1, P < 0.01 ANOVA) and was not significantly altered in sub-endocardial myocytes (control, 8.5 +/- 0.7 pA pF-1; hypertrophy, 7.4 +/- 1.8 pA pF-1). Steady-state background current density was reduced in hypertrophy (P < 0.05, ANOVA). The regional difference in steady-state background current in control hearts (P < 0.05, ANOVA) was altered in hypertrophy. Calcium current (ICa) density was similar in the three regions studied in both control and hypertrophied hearts. ICa was reduced in hypertrophy (P < 0.05, ANOVA). CONCLUSION: The normal regional differences in Ito1 are reduced, in steady-state background current are altered and in ICa are unchanged in catecholamine-induced hypertrophy in the rat left ventricle. These data may in part explain the reduction in the normal regional differences in APD observed in hypertrophy. PMID- 10533576 TI - Ionic basis of ventricular arrhythmias in remodeled rat heart during long-term myocardial infarction. AB - OBJECTIVE: Deleterious electrical abnormalities evolve during myocardial infarction. The goal of this study was to analyse current changes during the late decompensated phase of heart disease induced by coronary ligation and to compare them in various heart regions. METHODS: Young rats were submitted to left coronary ligature. After 4-6 months, cells were enzymatically dissociated and isolated from the upper part basal region of the left ventricle, as well as from the septum, apex and the right ventricle before being studied under whole-cell patch-clamp. RESULTS: Basal L-type Ca2+ current, ICaL elicited at +10 mV did not exhibit regional dependence neither in control nor after post-myocardial infarction (PMI). ICaL showed both a significantly reduced peak amplitude (17.1 +/- 2.8 pA/pF versus 9.9 +/- 1.4 pA/pF in seven control and seven PMI hearts, n = 32 and 40, respectively) and a slower inactivation, such that the amount of inward charges during a 200 ms-depolarizing pulse was nearly unchanged. beta Adrenergic stimulation was less effective in increasing ICaL in PMI cells but it slowed inactivation further. Significant differences in the K+ currents were observed. A regional distribution was seen for Ito only, with the largest amplitude in the right ventricle (in pA/pF: 23.1 +/- 2.4, 18.2 +/- 3.9, 14.8 +/- 2.4, 8.3 +/- 1.7 in the right ventricle, apex, septum and left ventricle, respectively n = 8, 7, 8 and 9). This was also true in failing heart cells despite Ito being halved in each of the four regions (in pA/pF: 12.2 +/- 2.5, 11.2 +/- 1.9, 5.1 +/- 1.0 and 4.8 +/- 1.0, respectively n = 12, 12, 11 and 13). IK1 was also significantly reduced by 20% in the PMI cells. Two-way analyses of variance demonstrated the absence of interaction between the topographical origin of the cells and the physiological state of the rats. The alpha 1-adrenergic agonist, methoxamine significantly reduced Ito and IK1 to the same extent in both sham and PMI cells, by about 35% and 20% respectively. CONCLUSIONS: Long-term left coronary occlusion induces significant alterations in both Ca2+ and K+ currents that occur with similar amplitude in both ventricles. They include a marked reduction in Ito amplitude as well as a slowing of ICaL inactivation. Both factors could contribute to the disturbances in cellular electrical behaviour and the occurrence of arrhythmias in the post-myocardial infarcted heart. PMID- 10533577 TI - Influence of postnatal-development on I(f) occurrence and properties in neonatal rat ventricular myocytes. AB - OBJECTIVE: I(f) is a hyperpolarization-activated current, which plays a key role in determining the spontaneous rate of cardiac pacemaker cells. We have previously shown that I(f) is also expressed in left ventricular myocytes isolated from spontaneously hypertensive rats; in these cells, its occurrence and density is linearly related with the severity of myocardial hypertrophy. Since hypertrophy induces a re-expression of genes encoding fetal proteins, we investigated changes in I(f) properties during post-natal development. METHODS: Fresh ventricular myocytes were enzymatically isolated from the heart of 1-2- to 28-day-old Wistar rats. The whole-cell configuration of the patch-clamp technique was employed to record the action potential and I(f). RESULTS: Membrane capacitance, an index of cell size, progressively increased from 13 +/- 1 pF at 1 2 days to 66 +/- 4 pF at 28 days of age (p < 0.01). At 1-2 days, a cesium sensitive hyperpolarization-activated inward current (I(f)) was recorded in the majority of tested cells (n = 51). The midpoint of the activation curve (V1/2) was -78 +/- 2 mV (n = 32), and specific current conductance of fully activated I(f) (gf.max) was 60 +/- 11 pS/pF. Reversal potential (Vrev) measured by tail current analysis was -24 +/- 3 mV (n = 8). Reduction of extracellular Na+ from 140 to 35 mM or extracellular K+ from 25 to 5.4 mM caused a shift of -12 +/- 1 mV (n = 3) or -11 +/- 2 mV (n = 5) of Vrev, respectively. Occurrence of I(f) decreased with aging, being present in 64%, 48% and 32% of cells at 10, 15 and 28 days, respectively. When present, I(f) density was significantly smaller than at 1-2 days (p < 0.05), reaching a value of 8 +/- 2 pS/pF at 28 days. However, V1/2 did not change in the older rats, being -80 +/- 2, -83 +/- 4 and -85 +/- 3 mV at 10, 15 and 28 days, respectively. Vrev at 10 and 15 days was -27 and -28 mV, respectively, thus suggesting that channel selectivity did not change. CONCLUSIONS: The pacemaker current, I(f), is expressed in ventricular myocytes from neonatal rats and progressively disappears; when present, it shows electrophysiological properties similar to I(f) re-expressed in hypertrophied adult rat ventricular myocytes. Thus, it is likely that the occurrence of I(f) in ventricular myocytes of hypertrophied and failing hearts is due to the re expression of a fetal gene. PMID- 10533578 TI - Decreased inward rectifier current in adult rabbit ventricular myocytes maintained in primary culture: a single-channel study. AB - OBJECTIVE: Regulation of ion channel function in heart has been shown to be affected by changes in the cellular environment. Recently it was shown that rabbit ventricular myocytes kept in primary culture, show a strong reduction in inward rectifier current (IK1). The aim of the present study was to elucidate the mechanism underlying this decrease in IK1, using single-channel measurements. In addition, we studied the effects of primary culture on the ATP-regulated K+ (K.ATP) channel, also a member of the inwardly rectifying K+ channel family. METHODS: Adult rabbit ventricular myocytes were cultured for up to 3 days in Ham's F-10 medium complemented with 1% rabbit serum and 5% glutamine. IK1 and K.ATP channel activity was studied in the inside-out patch configuration of the patch-clamp technique with equimolar K+ concentrations (140 mM K+) on the intra- and extracellular side. Single channel characteristics were determined at various times during culture and compared to those present in freshly isolated myocytes. RESULTS: IK1 channels in freshly isolated myocytes (day 0) had a single-channel conductance of 56.1 +/- 2.5 pS (mean +/- SEM) and an open probability of 0.64 +/- 0.05 (mean +/- SEM). Neither the single-channel conductance nor the open probability (Po) underwent significant changes during culture. The mean number of channels per patch, however, was drastically reduced from 1.2 +/- 0.3 (mean +/- SEM) at day 0 to 0.17 +/- 0.06 at day three. K.ATP channel density and open probability, on the other hand, were both increased with an optimum at day two. Po increased from 0.27 +/- 0.06 at day 0 to 0.63 +/- 0.06 at day three. The mean number of channels per patch was 2.29 +/- 0.57 and 3.25 +/- 0.48 at days 0 and 3 respectively. The unitary current amplitude at -50 mV remained unchanged, suggesting no change in the K.ATP single-channel conductance. CONCLUSIONS: The decrease in IK1 in rabbit ventricular myocytes as has been observed during primary culture is the result of a reduction in the number of active channels and not of altered kinetic or conductive channel properties. The increase in K.ATP channel activity under the same conditions suggests that gene expression of both channel types is differently regulated. PMID- 10533579 TI - Dihydropyridine and beta adrenergic receptor binding in dogs with tachycardia induced atrial fibrillation. AB - BACKGROUND: We have shown that rapid atrial activation, as occurs during atrial fibrillation (AF), reduces L-type Ca2+ current (ICa) and that this is the principal mechanism of the action potential duration and refractoriness changes that characterize tachycardia-induced atrial remodeling. The present study was designed to determine whether atrial tachycardia alters biochemical indices of the number of L-type Ca2+ channels and/or of the number and binding affinity of beta-adrenergic receptors. METHODS: In canine atrial sarcolemmal preparations, the number and binding affinity of dihydropyridine receptors were determined with the use of 3H-nitrendipine and that of beta-adrenergic receptors with 125I iodocyanopindolol. Results were obtained with preparations from dogs paced at 400/min for 1 (P1, n = 20), 7 (P7, n = 9), and 42 (P42, n = 9) days, and compared with observations in sham-operated controls (P0, n = 14). RESULTS: Pacing reduced the Bmax of dihydropyridine receptors, from 157 +/- 18 fmol/mg (P0) to 116 +/- 9 fmol/mg (P1, P < 0.05), 100 +/- 14 fmol/mg (P7, P < 0.05) and 94 +/- 9 fmol/mg (P42, P < 0.01). The affinity of dihydropyridine receptors was unchanged, with the Kd averaging 711 +/- 102 pM. 656 +/- 74 pM, 633 +/- 155 pM and 585 +/- 92 pM in P0, P1, P7 and P42 dogs. Neither Bmax nor Kd of beta-adrenergic receptors was altered by rapid pacing. Values of Bmax of dihydropyridine receptors correlated with atrial ICa current density (r2 = 0.95) and ERP (r2 = 0.99). CONCLUSIONS: Rapid atrial activation results in downregulation in the number of dihydropyridine receptors without altering the number or affinity of beta adrenergic receptors. The reductions in ICa that play an important role in the atrial electrical remodeling by which 'AF begets AF' appear to be due at least in part to a decrease in the number of L-type Ca2+ channels in cardiac cell membranes. PMID- 10533580 TI - Gene expression of proteins influencing the calcium homeostasis in patients with persistent and paroxysmal atrial fibrillation. AB - OBJECTIVE: Persistent atrial fibrillation (AF) results in an impairment of atrial function. In order to elucidate the mechanism behind this phenomenon, we investigated the gene expression of proteins influencing calcium handling. METHODS: Right atrial appendages were obtained from eight patients with paroxysmal AF, ten with persistent AF (> 8 months) and 18 matched controls in sinus rhythm. All controls underwent coronary artery bypass grafting, whereas most AF patients underwent Cox's MAZE surgery (n = 12). All patients had a normal left ventricular function. Total RNA was isolated and reversely transcribed into cDNA. In a semi-quantitative polymerase chain reaction the cDNA of interest and of glyceraldehyde-3-phosphate dehydrogenase were coamplified and separated by ethidium bromide-stained gel electrophoresis. Slot blot analysis was performed to study protein expression. RESULTS: L-type calcium channel alpha 1 and sarcoplasmic reticulum Ca(2+)-ATPase mRNA (-57%, p = 0.01 and -28%, p = 0.04, respectively) and protein contents (-43%, p = 0.02 and -28%, p = 0.04, respectively) were reduced in patients with persistent AF compared to the controls. mRNA contents of phospholamban, ryanodine receptor type 2 and sodium/calcium exchanger were comparable. No changes were observed in patients with paroxysmal AF. CONCLUSIONS: Alterations in gene expression of proteins involved in the calcium homeostasis occur only in patients with long-term persistent AF. In the absence of underlying heart disease, the changes are rather secondary than primary to AF. PMID- 10533581 TI - Steady-state and nonsteady-state action potentials in fibrillating canine atrium: abnormal rate adaptation and its possible mechanisms. AB - OBJECTIVE: Our goal was to study rate adaptation of atrial action potentials in non-steady and steady states to further our understanding of mechanisms determining inducibility and stability of atrial fibrillation. METHODS: We used standard microelectrode techniques to examine the characteristics of steady-state action potentials paced at regular cycle lengths (CL) and of nonsteady-state action potentials observed after an abrupt change of CL in atria from canine hearts that had been rapidly paced. RESULTS: We compared action potential characteristics among normal atria, atria in which chronic atrial fibrillation (cAF, lasting more than 3 days) had been induced and atria in which only nonsustained atrial fibrillation (nAF, lasting less than 12 h) had been induced. In steady-state, the rate adaptation of maximum diastolic potential (MDP) and action potential duration (APD) and markedly reduced in both cAF and nAF. Action potential characteristics did not differ between cAF and nAF atria, suggesting that factors other than electrophysiological properties determine the chronicity of AF. The time course of change in APD after an abrupt change of CL was altered in nAF/cAF atria; i.e., when CL was prolonged, APD also prolonged at the first beat, and then shortened during several subsequent beats (initial phase). Thereafter, APD slowly prolonged to a new steady-state (slow phase). In nAF/cAF atria, the initial phase was enhanced (greater shortening of APD) and the slow phase was reduced (less prolongation of APD). This latter phase was modified by ryanodine. CONCLUSIONS: Thus the reduced rate adaptation of steady-state APD is explained mainly by the loss of a slow phase of APD adaptation in nAF/cAF which is reversed in the presence of ryanodine. Therefore, in both nAF and cAF atria, rate adaptation of MDP as well as APD are reduced, nonsteady state as well as steady state, AP characteristics are markedly altered and these changes are partially explicable by Ca, -dependent processes. PMID- 10533582 TI - Reversal of atrial electrical remodeling following cardioversion of long-standing atrial fibrillation in man. AB - BACKGROUND: In animal studies, atrial fibrillation has been shown to shorten the atrial refractory period and impair its rate adaptation. However, little is known about the effects of chronic atrial fibrillation on atrial electrophysiology and its recovery course in humans. METHODS AND RESULTS: Nineteen patients, mean age 64 +/- 14 years, with chronic atrial fibrillation of more than six months duration were included in this study. All of them were successfully converted to sinus rhythm with an external defibrillator. Atrial effective refractory periods at right atrial appendage and distal coronary sinus were determined with five pacing cycle lengths (300, 400, 500, 600 and 700 ms) at 30 min after cardioversion and once a day for four days. The atrial conduction properties, including P wave duration of surface ECG, and right and left atrial conduction times, were also measured at the same time interval. Twenty age-matched patients without a history of atrial tachyarrhythmia were evaluated as controls. In comparison with controls, chronic atrial fibrillation significantly shortened the atrial effective refractory period, impaired its rate adaptation response, especially at distal coronary sinus, and depressed the conduction properties of atria. The atrial conduction properties did not change during the four-day follow up period; however, the atrial effective refractory period was gradually prolonged and its rate adaptation response improved after restoration of sinus rhythm. CONCLUSIONS: In humans, chronic atrial fibrillation significantly shortened the atrial effective refractory period, and impaired its rate adaptation response. Restoration and maintenance of sinus rhythm could reverse these electrophysiological changes. PMID- 10533583 TI - Ionic targets for drug therapy and atrial fibrillation-induced electrical remodeling: insights from a mathematical model. AB - Recent advances in molecular electrophysiology have made possible the development of more selective ion channel blockers for therapeutic use. However, more information is needed about the effects of blocking specific channels on repolarization in normal human atrium and in atrial cells of patients with atrial fibrillation (AF). AF-induced electrical remodeling is associated with reductions in transient outward current (Ito), ultrarapid delayed rectifier current (IKur), and L-type calcium current (ICa,L). Direct evaluation of the results of ion channel depression is limited by the nonspecificity of the available pharmacological probes. OBJECTIVES: Using a mathematical model of the human atrial action potential (AP), we aimed to: (1) evaluate the role of ionic abnormalities in producing AP changes characteristic of AF in humans and (2) explore the effects of specific channel blockade on the normal and AF-modified AP (AFAP). METHODS: We used our previously developed mathematical model of the normal human atrial AP (NAP) based on directly measured currents. We constructed a model of the AFAP by incorporating experimentally-measured reductions in Ito (50%), IKur (50%), and ICa,L (70%) current densities observed in AF. RESULTS: The AFAP exhibits the reductions in AP duration (APD) and rate-adaption typical of AF. The reduction in ICa,L alone can account for most of the morphological features of the AFAP. Inhibition of Ito by 90% leads to a reduction in APD measured at -60 mV in both the NAP and AFAP. Inhibition of the rapid component of the delayed rectifier (IKr) by 90% slows terminal repolarization of the NAP and AFAP and increases APD by 38% and 34%, respectively. Inhibition of IKur by 90% slows early repolarization and increases plateau height, activating additional IK and causing no net change in APD at 1 Hz in the NAP. In the presence of AF induced ionic modifications, IKur inhibition increases APD by 12%. Combining IKur and IKr inhibition under both normal and AF conditions synergistically increases APD. In the NAP, altering the model parameters to reproduce other typical measured AP morphologies can significantly alter the response to K(+)-channel inhibition. CONCLUSIONS: (1) The described abnormalities in Ito, IKur and ICa,L in AF patients can account for the effects of AF on human AP properties; (2) AP prolongation by IKur block is limited by increases in plateau height that activate more IK; (3) Blockers of IKur may be more effective in prolonging APD in patients with AF; 4) Inhibition of both IKur and IKr produces supra-additive effects on APD. These observations illustrate the importance of secondary current alterations in the response of the AP to single channel blockade, and have potentially important implications for the development of improved antiarrhythmic drug therapy for AF. PMID- 10533584 TI - beta-Adrenergic action on wild-type and KPQ mutant human cardiac Na+ channels: shift in gating but no change in Ca2+:Na+ selectivity. AB - OBJECTIVE: Prior studies of the modulation of the Na+ current by sympathetic stimulation have yielded controversial results. Separation of the Na+ and Ca2+ currents poses a problem in myocyte preparations. The gating of cloned Na+ channels is different in oocytes compared with mammalian expression systems. We have examined the sympathetic modulation of the alpha-subunit of the wild-type human cardiac Na+ channel (hH1) and the long QT-associated mutant, delta KPQ, expressed in human embryonic kidney cells. METHODS: Stable cell lines of hH1 and delta KPQ were established in human embryonic kidney cells. Whole-cell and single channel currents were measured with the patch-clamp technique. Sympathetic stimulation was effected by exposure to isoproterenol or 8-bromo-cAMP. Na+ channel activation and inactivation were determined using standard voltage clamp protocols. Ca2+:Na+ permeability ratio was determined under bi-ionic conditions. RESULTS: We observed a qualitatively different effect of sympathetic stimulation on the cardiac Na+ current from that reported in frog oocytes: activation and inactivation kinetics were shifted to more negative potentials. This shift was similar for both hH1 and delta KPQ. [delta V0.5 for inactivation: 8.3 +/- 1.7 mV, p < 0.001 (hH1); 6.8 +/- 0.9 mV, p < 0.001 (delta KPQ)]. Increased rate of closed state inactivation contributed to the shifting of the inactivation-voltage relationship. Open-state inactivation was not affected as mean open times were unchanged. Reversal potential measurement in hH1 suggested a low Ca2+:Na+ permeability ratio of 0.017, uninfluenced by sympathetic stimulation. In delta KPQ, the size of the persistent relative to the peak current was increased with 8 bromo-cAMP from 3.0 +/- 0.7% to 4.3 +/- 0.6% (p = 0.056). CONCLUSIONS: Sympathetic stimulation exerts multiple effects on the gating of hH1. Similar effects are also seen in delta KPQ which may increase arrhythmia susceptibility in long QT syndrome by modifying the Na+ channel contribution to the action potential. PMID- 10533585 TI - Intrinsic lidocaine affinity for Na channels expressed in Xenopus oocytes depends on alpha (hH1 vs. rSkM1) and beta 1 subunits. AB - OBJECTIVE: The affinity of lidocaine for the alpha-subunit of the Na channel has been reported to be greater for heart than for non-heart alpha-subunits, and also to be no different. Lidocaine block has a complex voltage dependence caused by a higher affinity for the inactivated state over the resting state. Inactivation kinetics, however, depend upon the alpha-subunit isoform and the presence of the auxiliary beta 1-subunit and will affect measures of block. METHODS: We studied the voltage dependence of lidocaine block of Na currents by a two microelectrode voltage clamp in oocytes injected with RNA for the Na channel alpha-subunits of human heart (hH1a) or a rat skeletal muscle (rSkM1) alone, or coexpressed with the beta 1-subunit. RESULTS: The midpoints of availability for a 25-s conditioning potential in control solutions were -65 mV for rSkM1, -50 for rSkM1 + beta 1, -78 mV for hH1a and -76 for hH1a + beta 1. The Kd of tonic lidocaine block was measured at -90, -100, -110, -120 and -130 mV in the same oocytes. The apparent Kd for both isoforms +/- beta 1 became greater with more negative holding potentials, but tended to reach different plateaus at -130 mV (Kd = 2128 microM for rSkM1, 1760 microM for rSkM1 + beta 1, 433 for hH1a, and 887 microM for hH1a + beta 1). Inactivated state affinities, assessed by fitting the shift in the Boltzmann midpoint of the availability relationship to the modulated receptor model, were 4 microM for rSkM1, 1 microM for rSkM1 + beta 1, 7 microM for hH1a and 9 microM for hH1a + beta 1. CONCLUSION: The heart Na channel alpha subunits expressed in oocytes have an intrinsically higher rest state affinity for lidocaine compared to rSkM1 after the voltage- and state dependence of block are considered. Coexpression with beta 1 modestly increased the rest affinity of lidocaine for rSkM1, but had the opposite effect for hH1a. PMID- 10533586 TI - Benzocaine enhances and inhibits the K+ current through a human cardiac cloned channel (Kv1.5). AB - OBJECTIVE: The aim of this study was to analyze the effects of a neutral local anaesthetic, benzocaine, on a cardiac K+ channel cloned from human ventricle. METHODS: Experiments were performed on hKv1.5 channels stably expressed on mouse cells using the whole-cell configuration of the patch clamp technique. RESULTS: At 10 nM, benzocaine increased the current amplitude ("agonist effect") by shifting the activation curve 8.4 +/- 2.7 mV in the negative direction, and slowed the time course of tail current decline. In contrast, benzocaine (100-700 microM) inhibited hKv1.5 currents (KD = 901 +/- 81 microM), modified the voltage dependence of channel activation, which became biphasic, and accelerated the channel deactivation. Extracellular K+ concentration ([K+]o) also affected the channel gating. At 140 mM [K+]o, the time course of tail currents deactivation was significantly accelerated, whereas at 0 mM [K+]o, it was slowed. At both [K+]o the activation curve became biphasic. Benzocaine accelerated the tail current decay at 0 mM but not at 140 mM [K+]o. The reduction in the permeation of K+ through the pore did not modify the blocking effects of micromolar concentrations of benzocaine, but suppressed the agonist effect observed at nanomolar concentrations. CONCLUSIONS: All these results suggest that benzocaine blocks and modifies the voltage- and time-dependent properties of hKv1.5 channels, binding to an extracellular and to an intracellular site at the channel level. Moreover, both sites are related to each other and can also interact with K+. PMID- 10533587 TI - Cysteine scanning analysis of the IFM cluster in the inactivation gate of a human heart sodium channel. AB - The conserved isoleucine-phenylalanine-methionine (IFM) hydrophobic cluster located in the III-IV linker of voltage-gated sodium channels has been identified as a major component of the fast inactivation gate in these channels. OBJECTIVES: The aim of our study was to probe the contribution of each amino acids of the IFM cluster to the inactivation. METHODS: A combination of site-directed mutagenesis, cysteine covalent modification and electrophysiological recording techniques were used to elucidate the role of isoleucine1485 and methionine1487 on hH1 sodium channels expressed in tsA201 cells. RESULTS: Mutant I1485C behaves like mutant F1486C studied earlier: producing an incomplete inactivation (residual current), a slowing and change in the voltage-dependence of the time constants of current decay, a shift of the steady-state inactivation to more depolarized voltages, and a faster recovery from inactivation than the wild-type hH1. The electrophysiological parameters of mutant M1487C are similar to those of wild type hH1 except for the presence of a residual current. Exposure of the cytoplasmic surface of the mutants to MTS reagents MTSES, MTSET and MTSBn further disrupted inactivation. In order to explain differences in the amplitude of the sustained currents recorded in the presence of MTSES or MTSET, we studied the effects of exposure of mutants 11485C, F1486C and M1487C to acidic and basic pH in the absence and presence of MTSES and MTSET. The effects of MTSES [negatively charged (-)] and MTSET (+) on the amplitude of the residual current of mutant F1486C were modulated by changes in intracellular pH. CONCLUSION: Isoleucine1487 and methionine1485, which surround phenylalanine1487 contribute to stabilizing the inactivation particle for fast inactivation. PMID- 10533588 TI - Pause induced early afterdepolarizations in the long QT syndrome: a simulation study. AB - OBJECTIVE: The long QT syndrome (LQTS) is characterized by prolonged repolarization and propensity to syncope and sudden death due to polymorphic ventricular tachycardias such as torsade de pointes (TdP). The exact mechanism of TdP is unclear, but pause-induced early afterdepolarizations (EADs) have been implicated in its initiation. In this study we investigate the mechanism of pause induced EADs following pacing at clinically relevant rates and characterize the sensitivity of different cell types (epicardial, midmyocardial, and endocardial) to EAD development. METHODS: Simulations were conducted using the Luo-Rudy (LRd) model of the mamalian ventricular action potential (AP). Three cell types- epicardial, midmyocardial (M), and enocardial--are represented by altering the channel density of the slow delayed rectifier current, IKs. LQTS is modelled by enhanced late sodium current (LQT3), or reduced density of functional channels that conduct IKr (LQT2) and IKs (LQT1). The cell is paced 40 times at a constant Basic Cycle Length (BCL) of 500 ms. Following a 1500 ms pause, an additional single stimulus is applied. RESULTS: Our results demonstrate that pause-induced EADs develop preferentially in M cells under conditions of prolonged repolarization. These EADs develop at plateau potentials ('plateau EADs'). Mechanistic investigation shows that prolongation of the plateau phase of the post-pause AP due to a smaller delayed rectifier potassium current, IKs' and enhancement of the sodium-calcium exchange current, INaCa, allows for the reactivation of the L-type calcium current, ICa(L), which depolarizes the membrane to generate the EAD. CONCLUSIONS: APD is a very important determinant of arrhythmogenesis and its prolongation, either due to acquired or congenital LQTS, can result in the appearance of EADs. The formation of pause-induced EADs preferentially in M cells suggests a possible role for these cells in the generation of arrhythmias that are associated with abnormalities of repolarization (e.g., TdP). The ionic mechanism of pause-induced EADs involves reactivation of the L-type calcium current during the prolonged plateau of the post-pause AP. PMID- 10533589 TI - Evidence for a role of Trp proteins in the oxidative stress-induced membrane conductances of porcine aortic endothelial cells. AB - OBJECTIVE: Expression of homologues of the Drosophila transient receptor potential (Trp) protein has recently been demonstrated for vascular endothelium. Some Trp isoforms such as Trp3, are known to constitute cation conductances with biophysical properties similar to those of the endothelial oxidant-activated cation conductance. Therefore we tested whether Trp proteins provide the molecular basis of the oxidant-induced membrane currents in porcine aortic endothelial cells (ECAP). METHODS: Expression of the Trp3 isoform in ECAP was tested by RT-PCR and subsequent southern blot analysis. In order to knock-out the function of endogenous Trp channels, ECAP were transiently transfected to express NTRP3, a dominant negative fragment of Trp3. Oxidative-stress was introduced by exposure of cells to tert-butylhydroperoxide (tBHP; 400 microM), and membrane current as well as membrane potential were recorded using the conventional whole cell patch-clamp technique. RESULTS: RT-PCR experiments demonstrated the expression of a Trp3 isoform in ECAP. The oxidant tert.-butylhydroperoxide (tBHP) completely depolarized endothelial cells by activation of a cation conductance which allowed significant Na+ inward currents at negative potentials (mean inward current 462 pA at -80 mV). The tBHP-induced currents resembled Trp-related currents in terms of cation selectivity, La3+ sensitivity and lack of voltage dependence. Expression of the N-terminal fragment of hTrp3 (NTRP3), but not of a C-terminal fragment of hTrp3 (CTRP3), abolished the oxidant-induced cation current and reduced membrane depolarization. CONCLUSION: Our results strongly suggest Trp proteins as the molecular basis of endothelial oxidant-activated cation channels. It is concluded that Trp proteins play an important role in the redox sensitivity of the vascular endothelium. PMID- 10533590 TI - Crimson scalenes in Indigo Apple. PMID- 10533591 TI - New insights into the pathophysiological role for cytokines in heart failure. PMID- 10533592 TI - Guidelines to the use of laboratory animals: what about neuromuscular blocking agents? PMID- 10533594 TI - Preconditioning and limitation of stunning: one step closer to the protected protein(s)? PMID- 10533593 TI - Respiratory control in normal and hypertrophic hearts. PMID- 10533595 TI - Cytokine-induced free radicals and their roles in myocardial dysfunctions. PMID- 10533596 TI - Homocysteine and endothelial function. PMID- 10533597 TI - Cardiac alternans: mechanisms and pathophysiological significance. PMID- 10533598 TI - Pacing-induced heart failure: a model to study the mechanism of disease progression and novel therapy in heart failure. PMID- 10533599 TI - Spotlight on microcirculation: an update. PMID- 10533600 TI - Beneficial effects of propionyl L-carnitine on sarcolemmal changes in congestive heart failure due to myocardial infarction. AB - OBJECTIVE: Earlier studies have revealed sarcolemmal (SL) defects in congestive heart failure due to myocardial infarction; however, the mechanisms of SL changes in the failing heart are poorly understood. Since congestive heart failure is associated with various metabolic abnormalities including a deficiency of carnitine, we examined the effects of propionyl L-carnitine, a carnitine derivative, in animals with congestive heart failure. METHODS: For this purpose, heart failure in rats was induced by occluding the coronary artery and 3 weeks later the animals were treated with 100 mg/kg (i.p. daily) propionyl L-carnitine for 4 weeks. The sham control group received saline injections. The animals were assessed for their left ventricular function. SL membranes were examined for Na(+)-K+ ATPase, Na(+)-Ca2+ exchange and adenylate cyclase activities. RESULTS: A marked improvement in the attenuated left ventricular function of the experimental animals was seen upon treatment with propionyl L-carnitine. The SL adenylyl cyclase activities in control, untreated failing hearts and treated failing hearts were 590 +/- 36, 190 +/- 22 and 320 +/- 21 pmol cAMP/mg/10 min, whereas the SL Na(+)-K+ ATPase activities were 35.7 +/- 2.8, 22.5 +/- 2.4 and 30.1 +/- 2.8 mumol Pi/mg/h, respectively. Furthermore, the SL Na(+)-dependent Ca(2+)-uptake activity, which decreased in the failing hearts (4.6 +/- 0.4 vs. 9.3 +/- 0.7 nmol Ca2+/mg/2 s for control), was improved (6.8 +/- 0.5 nmol Ca2+/mg/2 s) significantly following treatment with propionyl L-carnitine. CONCLUSION: These results indicate that metabolic therapy with propionyl L carnitine may attenuate defects in the SL membrane and thus may improve heart function in congestive heart failure due to myocardial infarction. PMID- 10533601 TI - Myocardial oxygenation at high workstates in hearts with left ventricular hypertrophy. AB - BACKGROUND: High cardiac workloads produced by catecholamine infusion result in loss of myocardial phosphocreatine (PCr) and accumulation of inorganic phosphate (Pi) which are more prominent in heart with left ventricular hypertrophy (LVH) than in normal hearts. Since ischemia can cause changes in phosphorylated compounds similar to those during catecholamine stimulation, this study tested the hypothesis that the exaggerated depletion of PCr and accumulation of Pi during high workloads in LVH is the result of impaired myocyte oxygenation. METHODS AND RESULTS: 31P- and 1H-NMR spectroscopy were used to determine myocardial high energy phosphate levels and myoglobin desaturation, respectively, in eight normal dogs and nine dogs with LVH produced by ascending aortic banding. The mean LV weight/body weight ratio was approximately twice normal in the LVH group. Infusion of dobutamine (15 and 30 micrograms/kg/min), and dobutamine + dopamine (each 20 micrograms/kg/min) caused progressive similar increases in the heart rate x systolic LV pressure product to a maximum of 57.4 +/- 3.3 x 10(3) in normal and 63.9 +/- 2.7 x 10(3) in LVH animals, while myocardial oxygen consumption increased from 0.09 +/- 0.01 to 0.24 +/- 0.04 in normals and from 0.10 +/- 0.02 to 0.25 +/- 0.03 ml/min/g in LVH. PCr/ATP ratios during basal conditions were lower in LVH hearts (1.73 +/- 0.10, 1.61 +/- 0.09 and 1.51 +/- 0.09 in subepicardium, midwall and subendocardium, respectively) as compared with normals (2.24 +/- 0.09, 2.01 +/- 0.08 and 1.89 +/- 0.07; each p < 0.01 normal vs. LVH). Catecholamine infusions caused dose-related decreases in PCr/ATP and appearance of Pi which was more marked in LVH than in normal hearts. 1H-NMR spectroscopy did not detect deoxymyoglobin in either normal or LVH hearts even during the highest workloads. In contrast, occlusion of the anterior descending coronary artery resulted in a large deoxymyoglobin signal. CONCLUSIONS: Increases of cardiac work produced by catecholamine stimulation resulted in greater decreases of PCr and greater increases of Pi in hypertrophied than in normal hearts. These abnormalities were not the result of inadequate intracellular oxygen availability and consequently cannot be ascribed to demand ischemia. PMID- 10533602 TI - Effects of endothelial and inducible nitric oxide synthases inhibition on circulatory function in rats after myocardial infarction. AB - OBJECTIVES: To examine the relative roles of eNOS and iNOS (endothelial and inducible nitric oxide synthases) on basal and beta-adrenergic receptor (beta-AR) stimulated arterial hemodynamic responses after myocardial infarction (MI). METHODS: Left ventricular (LV) pressures and steady-state and pulsatile arterial hemodynamics were measured at baseline, and after acute NOS inhibition with either NG-nitro-L-arginine methyl ester (L-NAME, 100 mg/kg) or iNOS inhibition with aminoguanidine (AG, 75 mg/kg) in sham-operated and MI Sprague-Dawley rats. RESULTS: In sham rats, L-NAME decreased (P < 0.05) peak positive LV dP/dt and aortic blood velocity by 19% and 53%, respectively, and increased (P < 0.05) mean arterial pressure (MAP); systemic vascular resistance, and LV end-diastolic pressure (EDP) by 20, 189 and 89%, respectively. The frequency-dependent components of hemodynamics including aortic input impedance modulus, characteristic impedance, and phase shift were increased (P < 0.05) with L-NAME, while pulsatile power was decreased (P < 0.05). AG increased (P < 0.05) aortic input impedance modulus and characteristic impedance but had no effect on any other hemodynamic variable. In MI rats, L-NAME decreased (P < 0.05) LV dP/dt and aortic blood velocity by 22 and 55%, respectively, and increased (P < 0.05) SVR by 108%. There was no effect of L-NAME on MAP or LV EDP in MI rats. After MI, AG increased (P < 0.05) heart rate and LV dP/dt but had no effect on other LV or pulsatile hemodynamic variables. Compared to sham rats, heart rate, LV dP/dt, and blood velocity-isoproterenol dose responses were shifted downward (P < 0.05), while SVR-isoproterenol dose response was shifted upward (P < 0.05) in MI rats. In sham rats, L-NAME potentiated (P < 0.05, at > 10(-2) micrograms/kg) the isoproterenol-induced increase in LV dP/dt and aortic blood velocity, and potentiated (P < 0.05) the isoproterenol-induced decline in SVR. As expected, AG had no effects on isoproterenol-stimulated hemodynamics in sham rats. After MI, there was no effect of L-NAME or AG on isoproterenol-stimulated hemodynamics. CONCLUSIONS: (1) Circulatory and cardiac responses to inhibition of NO by L-NAME suggest that eNOS, but not iNOS, is the principal regulator of integrated arterial hemodynamic function in rats. (2) Both basal and beta-AR-stimulated NO regulation of hemodynamic are attenuated after MI. (3) The attenuation of arterial hemodynamic effects after isoproterenol is mediated, in part, by alterations in the beta-AR-activation of eNOS system after MI. PMID- 10533603 TI - Preservation of myofilament calcium responsiveness underlies protection against myocardial stunning by ischemic preconditioning. AB - OBJECTIVE: Whereas diminution of infarct size by ischemic preconditioning (IP) is well-accepted, protection against stunning is controversial. Since stunning is characterized by decreased myofilament Ca2+ responsiveness, we investigated whether IP would preserve myofilament responsiveness in a model of stunning. METHODS: Rat hearts were retrogradely perfused with Krebs-Henseleit (K-H) solution for 20 min and then subjected to 20 min of no-flow global ischemia, followed by 20 min of reperfusion in the absence (stunning) or in the presence (IP) of a previous 5-min period of ischemia followed by 15 min of reperfusion. A group of hearts perfused under non-ischemic conditions served as control. Thin ventricular trabeculae were dissected from each of the experimental groups and loaded with fura-2 to measure intracellular calcium concentration ([Ca2+]i) and developed force. RESULTS: After 20 min of reperfusion, left ventricular developed pressure decreased in stunned hearts to 61 +/- 5% of control (P < 0.01), whereas recovery was complete in the IP hearts (97 +/- 4%). Steady-state [Ca2+]i-force relationships revealed a decreased maximal Ca(2+)-activated force in stunned hearts relative to control, but no change in the IP group. The Ca2+ required for 50% activation increased in stunning but not in IP. CONCLUSIONS: These results show that the decrease in myofilament responsiveness that characterizes stunning is prevented by ischemic preconditioning. PMID- 10533604 TI - Regulation of glycogen utilization in ischemic hearts after 24 hours of fasting. AB - INTRODUCTION: Fasting protects the ischemic heart from injury and infarction. Previous studies have shown that hearts from fasted animals have greater glycogen utilization and a lower cytosolic redox state (NADH/NAD+) during global ischemia. While the mechanisms of increased glycogen utilization in fasted animals have not been elucidated, animals that hibernate or are tolerant of anoxia are known to increase the tissue content of the active form of glycogen phosphorylase, phosphorylase a. Therefore, this study was designed to (a) determine whether hearts from fasted animals have increased activity of glycogen phosphorylase during ischemia and (b) define those mechanisms responsible for this increase. METHODS: Hearts isolated from either fed or fasted (24 h) rats were perfused and freeze-clamped at baseline, and after 1 and 10 min of ischemia, for measurement of phosphorylase activity, phosphorylase kinase activity, and glucose-6-phosphate concentrations. RESULTS: Fasting increased the phosphorylase a/b ratio under both baseline and ischemic conditions. This increase was not accompanied by an increase in the activity of phosphorylase kinase, either with maximal [Ca2+] or under physiologic [Ca2+]. Glucose 6-phosphate concentrations were lower in hearts from fasted animals under baseline, but not ischemic, conditions. CONCLUSIONS: Fasting enhances glycogen utilization during ischemia by increasing the active form of glycogen phosphorylase. This increase is not due to a change in phosphorylation by phosphorylase kinase nor end-product inhibition by G-6P. While the precise mechanism of increased glycogen phosphorylase activity in fasted animals is not clear, one likely explanation may be the lower cytosolic redox state demonstrated in the myocardium of fasted animals. PMID- 10533606 TI - Angiotensin IV has mixed effects on left ventricle systolic function and speeds relaxation. AB - OBJECTIVE: A novel angiotensin receptor has been described and named AT4. Ligands for this receptor include the angiotensin II (Ang II) metabolite Ang II (3-8), known as angiotensin IV (Ang IV). There is 10-fold more AT4 receptor than AT1 receptor in rabbit myocardium. The AT4 receptor has a high affinity for Ang IV (Ki in rabbit myocardium < 2 x 10(-9)) and similar ligands, but very low affinity for Ang II (Ki in rabbit myocardium > 10(-6)). Although several functions have been attributed to the novel Ang IV peptide/AT4 receptor system, the effect of this system on left ventricular (LV) function has not been studied. We hypothesized (1) that Ang IV would affect LV function and (2) that any effects would be opposite to those of Ang II. METHODS: Using the buffer-perfused (30 degrees C) isolated rabbit heart, we studied the effect of the AT4 agonist Nle1 Ang IV on LV systolic function, quantified using both Frank-Starling and end systolic pressure-volume relationships, and relaxation. We also studied the effect of the AT1/AT2 agonist, Sar1-Ang II on LV function. Finally, because the profile of effect of Nle1-Ang IV was similar to the reported effect of nitric oxide (NO), we also studied the effect of Nle1-Ang IV in the presence of the NO synthase inhibitor NG-monomethyl-L-arginine. RESULTS: Nle1-Ang IV reduced LV pressure-generating capability at any volume but increased the sensitivity of pressure development to volume change. Nle1-Ang IV reduced LV ejection capability. Sar1-Ang II had the opposite effect-increasing both pressure generation and ejection capability. Finally, both Sar1-Ang II and Nle1-Ang IV speeded LV relaxation. Inhibition of NO synthase did not alter the effect of Nle1 Ang IV on LV systolic function or relaxation. CONCLUSIONS: AT4 receptor agonism has mixed effects on LV systolic function, depressing pressure-generation and ejection capabilities, but enhancing the sensitivity of pressure development to volume change. It also speeds relaxation. The effect of Ang IV on systolic function is generally opposite to the effect of Ang II, whereas the Ang IV influence on relaxation is similar to the effect of Ang II. PMID- 10533605 TI - Role of superoxide anion in the pathogenesis of cytokine-induced myocardial dysfunction in dogs in vivo. AB - OBJECTIVE: Although studies in vitro have implicated oxygen-derived free radicals as possible mediators of inflammatory cytokine-induced cell injury, the role of the radicals in the cytokine-induced myocardial dysfunction in vivo remains unclear. The present study was designed to address this point in our novel canine model of cytokine-induced myocardial dysfunction in vivo. METHODS: Studies were performed in mongrel dogs, in which microspheres (MS, 15 microns in diameter) with and without interleukin-1 beta (IL-1 beta) were injected into the left main coronary artery (control and IL-1 beta group). Left ventricular ejection fraction (LVEF) was evaluated by echocardiography for 1 week. RESULTS: Immediately after the intracoronary injection of MS (10(6)/kg), LVEF equally decreased to approximately 30% in both the control and IL-1 beta group. While LVEF rapidly recovered within 2 days in the control group, it remained depressed in the IL-1 beta group until day 7 (p < 0.0001 vs. control group). Pretreatment with OPC-6535 (an inhibitor of superoxide production) before (2 mg/kg i.v.) and 1 and 2 days after IL-1 beta MS application (1 mg/kg i.v.) prevented the IL-1 beta-induced myocardial dysfunction. Superoxide production in the myocardium was significantly higher in the IL-1 beta group than in the control group at day 2 (p < 0.01), and OPC-6535 significantly suppressed the IL-1 beta-induced superoxide production (p < 0.01). An HPLC assay showed that nitrotyrosine, a marker of the formation of peroxynitrite by superoxide anion and nitric oxide, was present in the myocardium treated with IL-1 beta but not in that with control MS. OPC-6535 abolished the IL 1 beta-induced formation of myocardial nitrotyrosine. CONCLUSION: These results indicate that superoxide anion and the resultant formation of peroxynitrite may substantially be involved in the pathogenesis of the cytokine-induced myocardial dysfunction in dogs in vivo. PMID- 10533608 TI - Ischemic preconditioning limits infarct size following regional ischemia reperfusion in in situ mouse hearts. AB - OBJECTIVE: Ischemic preconditioning has been demonstrated in a wide variety of animals, from dogs to rats. Experimentally-induced murine myocardial ischemia reperfusion has been described, but ischemic preconditioning has not been reported in mouse hearts. To test the hypothesis that mouse hearts exhibit preconditioning-induced protection, experiments were conducted in anesthetized open chest mice subjected to regional myocardial ischemia-reperfusion. METHODS: Following barbiturate anesthesia the FVB and C57BL/6J mice underwent a tracheostomy and were mechanically ventilated. The heart was exposed via a left thoracotomy performed with the aid of a dissecting microscope. A 7-0 silk suture on a curved taper needle was passed under the proximal left anterior descending coronary artery to form a snare. Mice were then randomly assigned to receive either no preconditioning or preconditioning. All mice were subjected to 60 min regional myocardial ischemia followed by 2.5 h of reperfusion. Ischemic preconditioning (IP) was induced with two (FVB mice) or three (C57BL/6J mice) cycles of 5 min coronary occlusion and 5 min reperfusion. Control animals did not receive preconditioning ischemia. Area-at-risk was assessed with fluorescent particles. Infarct size was assessed with triphenyl tetrazolium chloride, and is expressed below as a percentage of the area-at-risk. RESULTS: In FVB mice preconditioning reduced infarct size 49%, from 36.7 +/- 4.5% to 18.9 +/- 2.8% (P < 0.05). In C57BL/6J mice preconditioning reduced infarct size by 66%, from 56.4 +/- 8.3% to 18.9 +/- 4.2% (P < 0.05). CONCLUSION: From these data we conclude that the infarct limiting effect of ischemic preconditioning is demonstrable in murine hearts. PMID- 10533607 TI - The role of Bcl6 in mature cardiac myocytes. AB - OBJECTIVE: The Bcl6 gene encodes a sequence-specific transcriptional repressor and is ubiquitously expressed in adult murine tissues including heart muscle. The objective of this study was to examine the role of Bcl6 in cardiac myocytes. METHOD: We developed Bcl6-deficient (Bcl6-/-) mice and histologically examined hearts from these mice. RESULTS: Massive myocarditis with eosinophilic infiltration occurred in Bcl6-/- mice after 4-6 weeks of age. Since expression of the Bcl6 gene was induced in normal cardiac myocytes after 2 weeks of age and thereafter detected through adulthood, loss of Bcl6 in mature cardiac myocytes may be related to the induction of eosinophilic myocarditis. To examine the effects of eosinophils from Bcl6-/- mice on normal hearts, bone marrow cells from Bcl6-/- mice were adoptively transferred into sublethally irradiated RAG1 deficient mice. Although massive eosinophilic infiltration was detected in conjunctivas and spleens from the chimeric mice, myocarditis was never observed. Electron microscopic analysis of cardiac myocytes from Bcl6-/- mice revealed a spectrum of degenerative changes prior to eosinophilic infiltration. CONCLUSION: Bcl6 maynot be essential for the maturation of cardiac myocytes but may play a role in protecting mature cardiac myocytes from eosinophilic inflammation. PMID- 10533609 TI - Myocardial injury leads to a release of heat shock protein (hsp) 60 and a suppression of the anti-hsp65 immune response. AB - OBJECTIVE: While atherosclerosis is associated with high titers of autoantibodies to bacterial hsp65 crossreacting with human hsp60 (anti-hsp60 autoantibodies), myocardial infarction entails decreased humoral immune response to hsp65. We previously hypothesized that myocardial ischemia and subsequent infarction not only induce myocardial hsp60 expression, but also trigger release of myocardial hsp60 into the circulation, influencing the systemic hsp immune response via immune complex formation. METHODS: In the present study, organ culture of rat hearts under circulatory arrest provided a model of myocardiocyte injury due to ischemia. RESULTS: Reperfusion of ischemic hearts confirmed the occurrence of myocardial injury by a rise of heart enzymes. Myocardial hsp60 expression was induced up to threefold in response to ischemia, and most of hsp60 expression was localized to the muscle fibers. Analysis of coronary eluate revealed release of hsp60 from myocardium. In addition, hsp60-containing, but not hsp60-free, coronary eluate was recognized by anti-hsp65 serum antibodies and induced proliferation of hsp65-specific T cells. When hsp60-containing coronary eluate was reinjected into an hsp65-primed rat, both humoral and cellular hsp65-immune responses were strongly downregulated. CONCLUSION: Our findings demonstrate the release of highly immunogenic and crossreactive hsp60 into the circulation in response to myocardial ischemia and myocardiocyte injury. PMID- 10533610 TI - Overexpression of long or short FGFR-1 results in FGF-2-mediated proliferation in neonatal cardiac myocyte cultures. AB - OBJECTIVE: The type 1 fibroblast growth factor receptor (FGFR-1) is the only high affinity receptor for fibroblast growth factor-2 (FGF-2) in the rat myocardium, and is essential for normal growth and development of the heart. Levels of FGFR-1 are developmentally regulated, being high in embryonic cardiac myocytes. Also, FGFR-1 exists as both 'long' and 'short' isoforms, and there is a switch from predominant expression of the 'long' isoform in the embryo to the 'short' isoform in the adult heart. Both the decrease in receptor levels and the isoform switch in postnatal cardiac myocytes correlate with a loss of proliferative potential. We investigated whether an increase in either 'long' or 'short' FGFR-1 isoforms could stimulate proliferation in postnatal rat cardiac myocyte cultures. METHODS AND RESULTS: Previously we cloned cDNAs corresponding to 'long' (L) and 'short' (S) FGFR-1 isoforms from embryonic mouse hearts. Hybrid FGFR-1(L) and (S) genes, directed by a myosin light chain-2 promoter and SV40 enhancer sequences, were generated and used to transiently transfect neonatal rat cardiac myocytes. Overexpression of FGFR-1 mRNA and protein was detected by RNA blotting and immunocytochemistry. Ligand-crosslinking confirmed the presence of specific receptors capable of binding FGF-2 on the cell membrane. Overexpression of either FGFR-1(L) or (S) was associated with stimulation of proliferation as assessed by significant increases in bromodeoxyuridine uptake (DNA synthesis) and cell number. To determine whether this response was FGF-2 specific, the level of FGF-2 was assessed in the culture medium of cardiac myocytes overexpressing FGFR-1 isoforms. A three-fold increase was detected in the media of cardiac myocytes overexpressing either FGFR-1(L) or (S) compared to control levels. Neutralization of this FGF-2 with antibodies inhibited the proliferative response. CONCLUSION: Overexpression of either FGFR-1(L) or (S) resulted in an increase in FGF-2 mediated proliferation of postnatal rat cardiac myocytes. PMID- 10533611 TI - Isometric tension development and its calcium sensitivity in skinned myocyte sized preparations from different regions of the human heart. AB - OBJECTIVE: In this study we investigated whether differences exist or develop in patients with aortic or mitral valve disease in myofibrillar contractile function and contractile protein composition between subendo- and subepicardial human ventricular tissue. Isometric tension, its calcium sensitivity and contractile protein composition were studied in left ventricular subendo- and subepicardial and in atrial biopsies obtained during open heart surgery from 24 patients with mitral or aortic valve disease. METHODS: Isometric tension was measured in mechanically isolated skinned myocyte-sized preparations at different free calcium concentrations at 15 degrees C. Protein composition was analysed by one dimensional gel electrophoresis. A comparison was made between the results of subendo- and subepicardial ventricular tissue within each New York Heart Association class and within the different hemodynamically overloaded groups. RESULTS: Maximal isometric tension was significantly lower in atrial than in ventricular preparations. The concentration of calcium required for half-maximal activation was significantly higher in atrial than in ventricular preparations. Within the ventricle no differences were found in contractile protein composition, isometric tension and its calcium sensitivity between subendo- and subepicardial tissue when all patients were treated as one group or when patients were subdivided according to severity of heart disease or hemodynamic overload. CONCLUSIONS: In this group of patients with ventricular volume or pressure overload no regional differences exist or develop during cardiac disease in left ventricular myofibrillar protein composition and force production. Maximal isometric tension and its calcium sensitivity are smaller in atrial than in ventricular preparations. PMID- 10533613 TI - Detection of microsatellite instability in sporadic cardiac myxomas. AB - OBJECTIVE: Microsatellite instability (MIN) is an early event in DNA repair deficient associated diseases and reflects an elevated mutation rate in the genome of neoplastic cells. Sporadic cardiac myxomas are the most common primary heart tumours and their aetiopathology remains obscure. This study investigates the incidence of MIN in sporadic cardiac myxomas as a possible genetic mechanism of tumour pathogenesis. METHODS: Eleven surgically excised sporadic cardiac myxomas were assessed for MI using twenty-two highly polymorphic microsatellite markers, located on a wide range of chromosomal arms. DNA was extracted from myxoma tissue specimens as well as the respective normal tissue and subjected to polymerase chain reaction. RESULTS: The microsatellite analysis revealed that seven myxoma specimens (64%) exhibited MIN in at least one marker. One tumour specimen exhibited evidence of MIN in four microsatellite markers, while the most frequently affected marker was D17S855 (27%), located on chromosome 17q. DISCUSSION: We have detected a considerable incidence of MIN in sporadic cardiac myxomas indicating that decreased fidelity in DNA replication and repair is common in these tumours. To the best of our knowledge this is the first report describing MIN in sporadic cardiac myxomas, as a possible pathogenetic mechanism of these rare neoplasms. PMID- 10533612 TI - Distribution and function of recombinant endothelial nitric oxide synthase in transplanted hearts. AB - Introducing recombinant genes into donor hearts may offer a therapeutic intervention that could potentially attenuate the complications of heart transplantation, including rejection, infection and accelerated atherosclerosis. In the cardiovascular system, reduced bioactivity of endothelial nitric oxide is a feature of atherosclerosis and vascular injury. Nitric oxide is an arterial vasodilator that also inhibits proliferation of vascular smooth muscle cells and platelet aggregation. Experiments were designed to determine the distribution of adenoviral-mediated transfer of recombinant endothelial nitric oxide synthase gene (eNOS) and the effect of recombinant gene expression on the function of transplanted hearts. Adenoviral vectors for (a) bovine eNOS (AdeNOS) or (b) beta galactosidase (AdLacZ; control) were infused into two groups (n = 12, per group) of explanted rat hearts. The transduced hearts were then implanted heterotopically into the abdomen of syngeneic recipient rats. After four days, the hearts were excised and examined for distribution and function of the recombinant genes. Polymerase chain reaction (PCR) verified the presence of the recombinant eNOS gene in eNOS-transduced but not in beta-galactosidase-transduced hearts; reverse transcriptase-PCR identified mRNA for eNOS in AdeNOS-transduced hearts. NOS activity (conversion of tritiated L-arginine to citrulline) was greater in homogenates of AdeNOS-compared to AdLacZ-transduced hearts. Positive immunoreactivity for eNOS was present in cardiomyocytes predominantly in eNOS transduced hearts. Myocardial contractility and coronary blood flow, as determined using a Langendorff preparation, were not different between hearts transduced with AdeNOS or AdLacZ. These results suggest that, up to four days post transplantation, adenoviral-mediated transfer of eNOS into transplanted hearts is possible. However, expression of the recombinant protein did not result in measurable changes in myocardial contractility or coronary perfusion. PMID- 10533614 TI - Angiotensin inhibition and atrial natriuretic peptide release after acute volume expansion in rats with aortocaval shunt. AB - OBJECTIVE: In heart failure atrial natriuretic peptide (ANP) release in response to volume expansion is impaired while the renin-angiotensin system is activated. This study was designed to test the hypothesis that ANP release in heart failure is dependent on an activated angiotensin system. METHODS: We studied the ANP and renin-angiotensin systems in a rat model of shunt-induced high-output heart failure, in which we rapidly increased circulating fluid volume with a 5 ml, hyperoncotic infusion, and evaluated the effects of acute inhibition of the angiotensin converting enzyme as well as of the blockade of the angiotensin II type 1 receptors on the ANP release and on renal excretory function. RESULTS: ANP and angiotensin II plasma concentrations prior to volume expansion were elevated (p < 0.05) in rats with aortocaval shunt compared to controls. The diuretic response to acute volume expansion (18.5 +/- 1.5 vs. 48.2 +/- 2.4 microliters/min, p < 0.001) was markedly blunted. ANP release was attenuated in rats with aortocaval shunt, as was the increase of its second messenger cGMP in plasma and urine. The blunted increase in ANP plasma levels was not due to depleted cardiac stores as cardiac ANP content, as well as ANP synthesis, were increased (p < 0.05). Acute inhibition of the angiotensin converting enzyme as well as blockade of the angiotensin II type 1 receptors restored ANP release in response to volume expansion (p < 0.01). Moreover, acute inhibition of the renin angiotensin system completely normalized the diuretic response. CONCLUSIONS: Our data suggest that the ANP system is impaired in rats with aortocaval shunt. The activation of the angiotensin system contributes to the impairment of the ANP system. Acute inhibition of the angiotensin II system significantly improved the ability of the ANP system to respond to acute volume expansion. Our findings indicate a hitherto fore unappreciated interaction between both systems and suggest additional mechanisms for the beneficial effects of angiotensin converting enzyme inhibition or angiotensin II type 1 receptor antagonists in heart failure. PMID- 10533615 TI - Familial hyperhomocysteinaemia and endothelium-dependent vasodilatation and arterial distensibility of large arteries. AB - OBJECTIVES: Mild hyperhomocysteinaemia, fasting as well as after a methionine load, occurs in families and is associated with premature atherosclerosis. We hypothesised that endothelial dysfunction plays a role in the relation between hyperhomocysteinaemia and clinical vascular disease. METHODS: In this study flow mediated, endothelium-dependent vasodilatation of the brachial artery and, as a marker of biophysical changes of the vessel wall such as increased smooth muscle cell tone or collagen formation, arterial distensibility of the common carotid artery were investigated in 123 healthy first-degree relatives of patients with mild hyperhomocysteinaemia and coronary, cerebral or peripheral artery disease. RESULTS: In multiple linear regression analyses, the increase in the homocysteine concentration after a standard methionine load was a significant determinant of an impaired flow-mediated vasodilatation of the brachial artery (measured on a separate day). The only other predictors were the baseline vessel diameter and age. Fasting homocysteine level was not associated with flow-mediated vasodilatation in the brachial artery. There was no relationship between homocysteine levels and nitroglycerine-induced, endothelium-independent vasodilatation of the brachial artery. Arterial distensibility of the carotid artery was also not related to homocysteine levels. CONCLUSIONS: In healthy first degree relatives of patients with mild hyperhomocysteinaemia, the increase in homocysteine level after a methionine load is an independent predictor of endothelial dysfunction. The results also suggest that fasting and post methionine homocysteine levels may reflect distinct disturbances in methionine metabolism, which may be linked to vascular dysfunction through distinct mechanisms. PMID- 10533616 TI - Nifedipine improves endothelial function in hypercholesterolemia, independently of an effect on blood pressure or plasma lipids. AB - OBJECTIVE: Dihydropyridine calcium antagonists have been shown to retard atherogenesis in animal models and to prevent the development of early angiographic lesions in human coronary arteries. Endothelial dysfunction is an early event in the pathogenesis of cardiovascular disease. We investigated whether nifedipine could improve endothelial function in hypercholesterolemia, independently of changes in blood pressure or plasma lipids. METHODS: First, we compared in vivo forearm vascular responses to the endothelium-dependent and independent vasodilators serotonin (5-HT) and sodium nitroprusside (SNP) in 11 patients with familial hypercholesterolemia before and after 6-weeks treatment with nifedipine GITS (60 mg, OD) and in 12 matched controls. In a subgroup of six control subjects forearm vascular function was also assessed before and after 6 weeks nifedipine GITS treatment. In vitro, we subsequently explored possible mechanisms underlying the effect of nifedipine on endothelial function. We investigated the effects of nifedipine on both NO production by recombinant endothelial NO synthase (eNOS) and endothelial cells, using 3H-arginine conversion, as well as on superoxide generation by endothelial cell lysates, using lucigenin enhanced chemiluminescence. RESULTS: In hypercholesterolemia 5-HT induced vasodilation was impaired (47 +/- 9% increase in forearm bloodflow vs. 99 +/- 8% in controls). Treatment with nifedipine completely restored 5-HT-induced vasodilation (113 +/- 13%), whereas it did not influence basal forearm vasomotion or SNP-induced vasodilation. Nifedipine did not alter forearm vascular responses in control subjects and did not alter blood pressure or plasma lipids. In vitro, we found no direct effect of nifedipine on NO production by recombinant eNOS or endothelial cells. However, we did observe a reduction in endothelial superoxide generation. CONCLUSIONS: Our data show that nifedipine improves endothelial function in hypercholesterolemia. It is suggested from our in vitro experiments that this effect is due to reduced NO degradation. PMID- 10533617 TI - Angiotensin II receptor antagonists prevent neointimal proliferation in a porcine coronary artery organ culture model. AB - OBJECTIVES: Angiotensin II (AngII) generation in response to vascular injury has long been assumed to influence neointimal proliferation contributing to restenosis. This concept has been supported by evidence that ACE inhibition and AT1 receptor blockade effectively limits restenosis in the rat. On the other hand, ACE inhibition has proven ineffective in clinical trails. The present study examines the response of the porcine coronary artery after angioplasty in vitro and compares the actions of an ACE inhibitor to AngII receptor antagonists. METHODS AND RESULTS: Captopril, an ACE inhibitor, and the AngII receptor antagonists, losartan and PD123319, were evaluated for their ability to attenuate neointimal proliferation in a porcine organ culture model of coronary restenosis. The neointima was significantly increased by 300% after angioplasty compared to non-angioplasty controls. The AT1 receptor antagonist, losartan, produced a significant reduction in neointimal index at 10(-5) mol/l, while its in vivo metabolite, EXP3174, reduced neointimal proliferation at 10(-6) mol/l. PD123319, a selective antagonist of the AT2 receptor, also restricted neointimal proliferation at 10(-5) mol/l. Treatment with captopril (10(-6) mol/l) increased the neointimal proliferation by approximately 200% after angioplasty. CONCLUSIONS: Direct blockade of AngII receptors effectively inhibits cell proliferation and restenosis post-angioplasty in vitro. ACE inhibition, exclusive of flow, does not attenuate proliferative restenosis. These data suggest that AngII contributes to neointimal proliferation and validates the concept that receptor antagonists could contribute to the therapeutic management of restenosis. PMID- 10533618 TI - Arterial function in nitric oxide-deficient hypertension: influence of long-term angiotensin II receptor antagonism. AB - OBJECTIVE: Since the effects of angiotensin II receptor antagonism on arterial function in nitric oxide (NO)-deficient hypertension are unknown, we investigated the influence of losartan therapy (20 mg kg-1 day-1) on the control of arterial tone in NG-nitro-L-arginine methyl ester (L-NAME; 20 mg kg-1 day-1)-induced hypertension. METHODS: Forty Wistar rats were divided into four groups: control, losartan, L-NAME, and losartan + L-NAME. The responses of isolated mesenteric arterial rings were examined in standard organ chambers after 8 treatment weeks. RESULTS: Losartan therapy prevented the development of L-NAME-induced hypertension and the associated impairments of endothelium-independent relaxations to nitroprusside, isoprenaline, and cromakalim, vasodilators acting via the formation of NO, activation of beta-adrenoceptors and opening of K+ channels, respectively. In addition, endothelium-dependent relaxations of noradrenaline-precontracted rings to acetylcholine during NO synthase inhibition in vitro were decreased in L-NAME rats, and clearly improved by losartan therapy. The inhibition of cyclooxygenase by diclofenac improved the responses to acetylcholine more effectively in L-NAME than losartan + L-NAME rats, but the relaxations remained decreased in L-NAME rats when compared with losartan + L NAME rats. When hyperpolarization of smooth muscle was prevented by precontractions induced by high concentration of KCl, the responses to acetylcholine during combined NO synthase and cyclooxygenase inhibition were similar and almost abolished in all groups. Furthermore, superoxide dismutase, a scavenger of superoxide anions, enhanced the acetylcholine-induced relaxations more effectively in L-NAME than losartan + L-NAME rats, although plasma antioxidant capacity was similar in all study groups. CONCLUSION: Chronic L-NAME induced hypertension was associated with attenuated arterial relaxation via endothelium-dependent and -independent mechanisms, both of which were improved by the losartan treatment. The mechanisms whereby losartan enhanced arterial relaxation in this model of experimental hypertension may have included enhanced hyperpolarization and increased sensitivity to NO in smooth muscle, and decreased vascular production of superoxide and vasoconstrictor prostanoids. PMID- 10533619 TI - Chronic selective hypertriglyceridemia impairs endothelium-dependent vasodilatation in rats. AB - OBJECTIVE/METHODS: In order to investigate whether selective hypertriglyceridemia impairs endothelium-dependent vasodilatation in the rat hindlimb, rats were selectively bred to establish two strains, one with a pronounced hypertriglyceridemia (HT) and the other with normal plasma levels of triglycerides (LT). RESULTS: Carotid arteries and aortae removed from 3, 6, 9 and 12 month old LT- and HT-rats exhibited a normal morphology. However, marked morphological differences were observed between vessels from 18-20 month old HT- and LT-rats. The endothelium-dependent vasodilator acetylcholine (2 to 50 micrograms/kg), administered into the iliac artery, elicited a concentration dependent increase in hindlimb blood flow which was not different in 3, 6 and 9 month old LT- or HT-rats but was impaired in 12 and 18-20 month old HT-rats. In contrast the endothelium-independent vasodilator sodium nitroprusside enhanced blood flow in both strains to a similar extent. Neither administration of the nitric oxide (NO) synthase (NOS) substrate, L-arginine, nor the NOS inhibitor NGnitro-L-arginine, affected the responsiveness to endothelium-dependent vasodilators in 12 month old HT-rats. These attenuated responses could not be attributed to a decrease in endothelial NOS expression as Western blot analysis revealed identical levels of this enzyme in the aortae and carotid arteries from LT- and HT-rats. Determination of superoxide anion (O2-) formation however, demonstrated a markedly elevated production of O2- in aortae from HT-rats. CONCLUSION: We conclude that chronic selective hypertriglyceridemia, an independent risk factor in the development and progression of atherosclerosis, leads to an endothelial dysfunction which is associated with an increased vascular O2- production and a subsequent decrease in bioavailable NO. PMID- 10533620 TI - Phenotypic changes in rat and guinea pig coronary microvascular endothelium after culture: loss of nitric oxide synthase activity. AB - OBJECTIVES: Coronary microvascular endothelial cells (CMVEs) can modulate the contractile performance of the adjacent myocardium by the release of agents such as nitric oxide (NO). Most previous studies using CMVEs have been done in situ, in the intact organ. We set out to study possible differences in NO synthase (NOS) regulation between freshly isolated and cultured rat and guinea pig CMVEs. METHODS: CMVEs were isolated from Wistar rats and Dunkin Hartley guinea pigs and then grown in culture for varying times. Fura-2 fluorescence was used to measure agonist-induced changes in CMVE intracellular calcium levels. Agonist-induced changes in CMVE cGMP levels were measured by commercial radioimmunoassay kit. Western blot analysis was used to measure endothelial, constitutive NOS (ecNOS) and soluble guanylate cyclase (sGC) protein levels. Reverse transcription, polymerase chain reactions and Southern blotting were used to measure ecNOS mRNA transcripts. RESULTS: In both fresh (1 h post-isolation) and cultured (14 days with one passage) CMVEs of the rat and guinea pig, bradykinin (BK) and the calcium ionophore A23187 (both 1 microM) elicited significant (P < 0.01) increases in the fura-2 340/380 fluorescence ratio. In cultured CMVEs, basal cGMP levels were unaffected by exposure to BK or A23187. Exposure to sodium nitroprusside (SNP) or atrial natriuretic peptide (ANP) (both 1 microM) induced significant (P < 0.01) increases in cGMP in guinea pig cells, whereas in rat cells only ANP produced a significant (P < 0.01) response. By contrast, freshly isolated CMVEs of both species had higher basal cGMP levels than cultured cells, and on exposure to BK and A23187, responded with significant (P < 0.01) increases in cGMP. Moreover, exposure of both fresh rat and guinea pig CMVEs to SNP or ANP also resulted in significant (P < 0.01) increases in cGMP. Western blot analysis demonstrated that ecNOS and sGC protein were lost from the rat CMVEs following culture. Furthermore, there was also a significant loss of ecNOS mRNA from the rat cells following culture. CONCLUSIONS: These data demonstrate that freshly isolated rat and guinea pig CMVEs possess ecNOS activity, and that this activity is downregulated following culture. At least for the rat, this effect would seem to lie at both the transcriptional and translational level. Furthermore, rat CMVEs have reduced activity of sGC following culture. PMID- 10533621 TI - Tumor necrosis factor-alpha enhances hypoxia-reoxygenation-mediated apoptosis in cultured human coronary artery endothelial cells: critical role of protein kinase C. AB - BACKGROUND: Ischemia and tumor necrosis factor-alpha (TNF alpha) released during ischemia both cause apoptosis and necrosis of myocardial tissues. Since endothelium may be critically important in determination of cardiac function, we examined the interaction between TNF alpha and hypoxia-reoxygenation with regard to induction of apoptosis and underlying signaling pathway in cultured human coronary artery endothelial cells (HCAECs). METHODS AND RESULTS: HCAECs were cultured and exposed to hypoxia alone, hypoxia-reoxygenation, TNF alpha alone, TNF alpha plus hypoxia-reoxygenation, or TNF alpha only during the period of reoxygenation. Apoptosis was evaluated by transmission electron microscopy, DNA nick-end labeling and DNA laddering. Hypoxia alone caused modest time-dependent apoptosis of cultured HCAECs, and reoxygenation increased the number of apoptotic cells (P < 0.01 vs. hypoxia alone). TNF alpha induced concentration-dependent apoptosis, and enhanced reoxygenation-mediated apoptosis in cultured HCAECs (P < 0.01 vs. hypoxia-reoxygenation alone). As expected, monoclonal antibody to TNF alpha significantly blocked the pro-apoptotic effect of TNF alpha-induced apoptosis (P < 0.01). TNF alpha-induced apoptosis was found to be associated with marked activation of protein kinase C (PKC), and pretreatment of cells with a specific PKC inhibitor markedly reduced TNF alpha-mediated PKC activity and apoptosis. CONCLUSION: These observations indicate that hypoxia alone causes modest apoptosis, reoxygenation increases apoptosis beyond that caused by hypoxia in cultured HCAECs. TNF alpha alone causes apoptosis, and further enhances apoptosis caused by hypoxia-reoxygenation. The activation of PKC plays a critical role in TNF alpha-induced apoptosis of cultured HCAECs. PMID- 10533622 TI - Endothelin converting enzyme is located on alpha-actin filaments in smooth muscle cells. AB - OBJECTIVE: Endothelin converting enzyme is the key enzyme in the generation of endothelin-1 from big-endothelin-1. The mature endothelin-1 is a potent vasoconstrictor which also promotes mitogenesis and proliferation of smooth muscle cells. The objectives were to demonstrate in smooth muscle cells the presence of a phosphoramidon-sensitive endothelin converting enzyme activity, reveal the subcellular localization of the enzyme protein and determine the effects of the metalloproteinase inhibitor, phosphoramidon, the lysosomotrophic drug, chloroquine, and colchicine on the cycling pathway of the enzyme. METHODS: Subcellular localization of endothelin converting enzyme on human smooth muscle cells and the rat cell line, A7r5, was by immunofluorescence and confocal microscopy or by biotinylation of cell cultures and immunoblotting, after treatment of cell cultures with cytochalasin D, colchicine, chloroquine and phosphoramidon. Converting enzyme activity was determined by high performance liquid chromatographic assay. RESULTS: We detected phosphoramidon-sensitive endothelin converting enzyme activity in smooth muscle cells. In addition to its plasma membrane location, for the first time we revealed a striking co localization of endothelin converting enzyme and alpha-actin filaments in smooth muscle cells. Colchicine treatment results in a perinuclear accumulation of endothelin converting enzyme. An increased level of endothelin converting enzyme protein was shown to be present in smooth muscle cells which had been grown in the presence of phosphoramidon or chloroquine. CONCLUSION: The 120 kDa endothelin converting enzyme co-localizes with alpha-actin in smooth muscle cells and resembles that found in endothelial cells in that it is present on both the plasma membrane and intracellularly. PMID- 10533623 TI - Procoagulant and proinflammatory activity in acute coronary syndromes. PMID- 10533624 TI - Aspirin and plasma interleukin-6 acute coronary syndromes. PMID- 10533626 TI - All doctors are disabled, but some are more disabled than others. PMID- 10533625 TI - Medical research in the Rhondda valleys. AB - This brief review has highlighted some of the research done by the MRC in the South Wales valleys. The two MRC Units published, in total, over 2000 reports ranging from letters to journals to conference proceedings, with around 200 reports appearing in the BMJ, Lancet and Nature alone. The expertise gained in South Wales meant that the Pneumoconiosis Research Unit team was involved internationally in co-ordinating research on coal workers' lung disease, and later on the health effects of asbestos and other respirable dusts. It is remarkable that the early large-scale studies were conducted and analysed without the benefits of modern computers and statistical packages. The varied Epidemiology Unit research programme enabled Cochrane to develop his ideas on defining health and evaluating health services, and also Elwood, who directed the Unit from 1974 to 1995, to pioneer studies of aspirin prophylaxis in cardiovascular disease. A steady stream of occupational health studies were carried out by Epidemiology Unit staff and many other large surveys were conducted in other parts of South Wales and across the country. Later MRC Epidemiology Unit work has focused on the town of Caerphilly where a prospective study of some 2500 men, established in 1979, has so far resulted in over 100 papers, notably on haemostatic factors related to heart disease. The study has run in tandem with a similar survey in the Speedwell area of Bristol which was established by former Epidemiology Unit staff working for the health authority in that area. Other medical research groups have also worked in the South Wales valleys. In 1961 Julian Tudor Hart left the Epidemiology Unit after a year's research to enter general practice, and establish the famous research practice at Glyncorrwg, over the mountain from the Rhondda Fawr. This year the extensive collection of MRC survey records is being transferred to the Department of Social Medicine at Bristol, and it is quite likely that further research will be undertaken relating Rhondda survey data collected over 40 years ago to subsequent mortality. The South Wales valleys will continue to contribute to medical research into the next millennium. Archie Cochrane's papers have been catalogued and are available for study at the Cochrane Archive established at Llandough Hospital. PMID- 10533627 TI - Cogan's syndrome: an oculo-audiovestibular disease. AB - Typical Cogan's syndrome is a rare disease of young adults consisting of flares of interstitial keratitis and sudden onset of Meniere-like attacks (nausea, vomiting, tinnitus, vertigo and hearing loss). Life-threatening aortic insufficiency develops in 10% of reported cases. Atypical Cogan's syndrome (audiovestibular dysfunction with other types of inflammatory eye disease) is associated with vasculitis in 20% of cases and has a less favourable prognosis than typical Cogan's syndrome. PMID- 10533628 TI - Anthracycline-induced cardiomyopathy. AB - Anthracycline cardiomyopathy is less frequently encountered nowadays, due to the well-recognised dose limitations and cardiac monitoring protocols used by chemotherapy centres. However, it is a condition that will persist due to the sensitivity of some patients to these drugs and the necessity for large doses to be used for certain individuals. We have demonstrated the benefit of angiotensin converting enzyme inhibitor therapy and would consider introducing these compounds at the earliest opportunity. The use of probucol and vitamins as antioxidants capable of preventing the onset of cardiomyopathy in humans appears to require further investigation but may significantly reduce the incidence of this condition in the future. PMID- 10533629 TI - Can the use of low-dose dopamine for treatment of acute renal failure be justified? AB - The use of dopamine for the prevention and treatment of acute renal failure is widespread. Its use is based on physiology suggesting selective renal vasodilation when it is infused at low dose. This article reviews the available data on the clinical use of dopamine. When used to prevent acute renal failure in high-risk treatments there is no evidence of benefit of dopamine but, given the low incidence of significant renal failure, the studies are underpowered. In treatment of acute renal failure, the quality of the data is poor. Only in one small randomised trial of moderate acute renal failure in patients with malaria was a clinically significant benefit of dopamine shown. The rest of the data, in the form of case series, showed either no benefit of dopamine or small benefits of little clinical significance. Again, these studies are of insufficient power for conclusions to be drawn as to the overall benefits and risks. We conclude that benefits of dopamine use cannot be ruled out by currently available data but its use cannot be advised until trials examining clinically important endpoints in large numbers of patients have been performed. PMID- 10533630 TI - Evaluation of renal function in elderly heart failure patients on ACE inhibitors. AB - A total of 187 heart failure patients aged 65-92 years, with pretreatment serum creatinine levels below 200 mumol/l, were monitored for more than 12 months on angiotensin-converting enzyme (ACE) inhibitor therapy. Optimal ACE inhibitor dosage was found in 27% of patients, while a significant deterioration in renal function, characterised by > 20% increase in serum creatinine to > 200 mumol/l, occurred in 25 patients. This was most closely attributable to ACE inhibitor treatment per se (implying co-existence of bilateral renal artery stenosis) in only four cases, including one in whom renal deterioration was reproducible on inadvertent rechallenge. In the other 21, renal deterioration was attributable to diuretic-related blood volume depletion (two cases), nonsteroidal anti inflammatory drugs (two cases), obstructive uropathy (two cases), preterminal renal shutdown (two cases), and the interaction between diuretic and ACE inhibitor dosage (including long-acting vs short-acting drugs) (13 cases). This study could serve as the basis for future comparisons of ACE-inhibitor-related renal deterioration when the entry requirement is optimal ACE inhibitor dosage. PMID- 10533631 TI - Efficacy and safety of intravenous amiodarone in recent-onset atrial fibrillation: experience in patients admitted to a general internal medicine department. AB - We examined the efficacy and safety of intravenous amiodarone in 20 unselected patients with recent-onset atrial fibrillation who were admitted to a general internal medicine department during a 6-month period. The treatment protocol included a loading dose of 1200 mg intravenous amiodarone in 24 hours, after which amiodarone treatment was continued orally. Eleven of the 20 patients (55%) converted to sinus rhythm within 48 hours of intravenous amiodarone treatment and were discharged in sinus rhythm, while 9/20 (45%) patients failed to convert during hospitalisation. Six patients (30%) failed to convert to sinus rhythm even after one further month of oral treatment. There was one death and a high frequency (25%) of thrombophlebitis during hospitalisation. The in-hospital non convertors had a significantly lower ejection fraction and initial low ventricular response rate than the convertors. In conclusion, the acute conversion rate by intravenous amiodarone was at best modest. It is suggested that intravenous amiodarone is probably more effective in patients with rapid recent-onset atrial fibrillation and good left ventricular function. PMID- 10533632 TI - Funduscopy: a forgotten art? AB - Funduscopy is an integral part of the physical examination, especially in older patients in whom visual problems and systemic diseases affecting the fundi (e.g., diabetes mellitus) are more common. We have assessed the views of hospital doctors to funduscopy via a questionnaire survey, reviewed the case notes to see whether or not funduscopy is carried out on older patients, and assessed the views of older patients on vision via a questionnaire survey. Review of the case notes showed only three of 100 patients had had funduscopy. Most patients reported a visual problem on specific enquiry. Whilst most hospital doctors believed funduscopy was important, many felt they had insufficient training in this procedure and felt their skills could be improved. We conclude that older patients are missing out on routine funduscopy. Hospital doctors should be aware that not all patients complain of visual problems and specific enquiry should be made. The issue of training and encouragement to perform funduscopy needs to be addressed before funduscopy becomes a forgotten art. PMID- 10533633 TI - Diagnostic approach to patients with suspected pulmonary embolism: a report from the real world. AB - This study was carried out to examine the diagnostic approach to patients with suspected pulmonary embolism (PE) in a university hospital. A retrospective case record review of 251 patients with suspected pulmonary embolism was carried out according to a standard protocol, which looked at the utilisation of imaging techniques and compared clinical diagnoses with a standardised diagnosis established according to current recommendations. Isotopic lung scan was the most commonly used technique (73%), followed by leg vein sonography (36%) and contrast venography (31%). Lung arteriography was done in only 7% of patients. Among the 205 patients with a clinical diagnosis of PE, 115 (56%) would be diagnosed as having PE according to the standard criteria, 84 (41%) would be unclassified, and six (3%) would not be regarded as having PE. Among patients who were diagnosed as having PE and received anticoagulant therapy, 32% did not have the diagnosis confirmed by an imaging technique. Most of these had a non-diagnostic lung scan which, despite evidence to the contrary, seemed to be interpreted as confirmation of PE. We conclude that clinicians do not seem to follow current recommendations when approaching patients with suspected PE. In particular, there is an over reliance on lung scans, and the significance of non-diagnostic scans was often misinterpreted. Arteriography was underused. These results emphasise the need to take measures to implement practice guidelines and to explore the usefulness of newer non-invasive techniques. PMID- 10533634 TI - Spuriously elevated plasma calcitonin in a patient with a thyroid nodule not associated with medullary thyroid carcinoma. AB - An increase in plasma calcitonin concentration is widely regarded as a specific and sensitive indication of underlying medullary thyroid carcinoma (MTC). We present a case in which the association of increased plasma calcitonin concentration and a thyroid nodule was not due to MTC. Subsequent measurement of plasma calcitonin by a variety of methods highlighted the variability that exists in calcitonin measurement and the potential for clinically misleading results. The rationale for investigation and treatment of MTC, including a recommendation to screen all patients with thyroid nodules using plasma calcitonin measurement, is based on the use of specific two-site calcitonin assays which are not universally used in the UK or USA. PMID- 10533635 TI - Diabetic ketoacidosis precipitated by thyrotoxicosis. AB - We report two patients with type 1 diabetes mellitus, previously well controlled with good compliance, presenting with unexplained diabetic ketoacidosis. Following initial correction of the metabolic disorder, persisting tachycardia lead to the diagnosis of thyrotoxicosis. In both cases, treatment with propranolol and carbimazole helped in the stabilization of their metabolic states. Although thyrotoxicosis is known to destabilize diabetes control, we can find no reports of it precipitating diabetic ketoacidosis. PMID- 10533636 TI - Lobular carcinoma-in-situ within a fibroadenoma of the breast. AB - We present a case of an in-situ lobular carcinoma within an otherwise benign fibroadenoma in a 45-year-old woman. PMID- 10533637 TI - Hypoxaemia--think of the liver! Every internist should be aware of the hepatopulmonary syndrome. AB - Hepatopulmonary syndrome is characterised by arterial hypoxaemia, liver disease, and intrapulmonary vascular dilatation. A case is reported in which severe hypoxaemia, detected by chance, led to the diagnosis of liver disease and hepatopulmonary syndrome. PMID- 10533638 TI - Massive pleural effusions in cryptococcal meningitis. AB - Cryptococcal infection uncommonly presents with pulmonary manifestations and even more rarely so as massive bilateral effusions. Pleural involvement is usually associated with underlying pulmonary parenchymal lesions and is unusual while on antifungal therapy. We report a patient with cryptococcal meningitis who, while on intravenous 5-flucytosine and amphotericin B, developed life-threatening bilateral massive pleural effusions with evidence of spontaneous resolution, consistent with prior hypothesis of antigenic stimulation as the cause of pleural involvement. PMID- 10533639 TI - Unusual findings in a patient taking warfarin. PMID- 10533640 TI - New weakness in a critically ill patient. PMID- 10533641 TI - A woman with amenorrhoea. PMID- 10533642 TI - Acute epigastric pain and recurrent vomiting in an elderly man. PMID- 10533643 TI - An elderly man with dysphasia and pyrexia. PMID- 10533644 TI - Sudden hearing loss following acute hepatitis. PMID- 10533645 TI - Haemolytic anaemia associated with indinavir. PMID- 10533646 TI - Multiple left ventricular thrombi in dilated cardiomyopathy. PMID- 10533647 TI - Nasal ventilation. PMID- 10533648 TI - Time delay to thrombolytic therapy. PMID- 10533649 TI - Can combined defects be diagnosed on spirometry alone? PMID- 10533650 TI - Interprofessional learning. PMID- 10533651 TI - Metabolic acidosis. PMID- 10533652 TI - Prevalence of Helicobacter pylori infection in an Albanian population. PMID- 10533653 TI - [Anti HCV in hemodialyzed patients: reduction of the prevalence and association with epidemiological variables]. AB - In order to establish the present prevalence of HCV in hemodialyzed patients (HD) from the city of La Plata, and to know the association between the prevalence and different variables, we have studied 217 hemodialyzed patients belonged to the Hemodialysis Unit of a Public Hospital and other 7 private Units. Serological reactivity to Anti HCV and Anti HBc IgG were investigated in all of them, as well as age, sex, number of transfusions, the time under dialysis treatment, the use of erythropoietin and hepatitis episodes were taken into account. We have found a prevalence of Anti HCV of 18.4% which was significantly superior to the blood donors' prevalence (P < 0.01) and significantly inferior to the one found in 1993 in HD patients of the same city (p = 0.002). We have found an association between the presence of Anti HCV and transfusions exposure, as well as more hemodialysis time. Our results may agree with the ones that suggest the aminotransferases are not good markers for hepatocellular injuries in Anti HCV (R) HD patients. PMID- 10533654 TI - [Pancreatic and colonic simultaneous or successive resections of tumors in both organs, duodenum infiltrating colon carcinoma and pancreas tail carcinoma invading left colon. Report 10 cases]. AB - Two patients with cancer of the right colon fistulized in duodenum underwent simultaneous right hemicolectomies and cephalic duodenopancreatectomies. There were no important post-operative complications and they are both alive eleven and ten years after surgery. In another case a piece of duodenum adherent to colon was resected and the duodenum was transversally sutured, segments IV and V of the liver were also resected because of a infiltration. The patient is well more than three years after surgery. Another similar patient who underwent resection of a part of the duodenum during a right hemicolectomy due to adenocarcinoma, developed the head of the pancreas ten months later which was resected by means of a cephalic duodenopancreatectomy. He survived the second operation for three years and a half and died with carcinomatosis. A fourth female patient who had been referred due to jaundice caused by cephalopancreatic cancer was reoperated two months later with resection of the tumor Seven months after duodenopancreatectomy she underwent another operation for resection of a bleeding cancer of the rectum by means of anterior resection. She survived for four and a half years. Cancers of body and tail of the pancreas sometimes invade the left colon and make necessary resection in the same operation. This happened in four other cases. In two of them it was necessary to resect stomach and the first jejunal loop. Respective survivals were of two years and nine months in two cases, others are 4111 alive more than two years after surgery. But the fourth case, who had an acinar carcinoma, died a month later with en hepatic metastasis which grew rapidly. The opposite happened to the tenth patient because colon cancer invaded pancreas and stomach, that were partially resected. The patient is alive two and a half years later. PMID- 10533655 TI - [Liver cyst sclerotherapy: report of a case and review of the literature]. AB - We present a case of a 49-year-old woman with a single hepatic symptomatic cyst. We performed US and CT examinations for correct diagnostic purposes and we approached it with a 18 G Chiba neddle. There after a 6 F "pig tail" catheter was inserted using Seldinger's method and after performing a cystography 95% ethilic alcohol was instilled within a 48 hs period. The third day the patient was discharged with no complication. The control examination revealed no cyst what suever four months later. PMID- 10533656 TI - [Liver pseudotumor: a diagnostic problem]. AB - The Authors presents 3 cases of Hepatic Pseudotumours, analyzed of the diagnostics procedures, concluding that a correct interpretation of the Sonography and CT avoid the utilization of invasive methods. PMID- 10533657 TI - [Barrett's esophagus: diagnosis and coexistent diseases in childhood]. AB - From 1990 to 1998, 12 patients with columnar metaplasia of the lower esophagus were diagnosed. Only 4 of them displayed a Barrett's epithelium (BE) with a specialized columnar epithelium and globet cells. As it is already published, a male predominance was observed. In three of these patients some predisposing factor to develop BE was detected (i.e. severe central nervous system damage, chronic pulmonary disease, esophageal atresia and chemotherapy). All patients had severe gastroesophageal reflux (GER) with abnormal pH probe. Diagnosis was suggested by characteristic endoscopy images in 2 patients and was confirmed by biopsy in all cases. All patients had been primary treated with proton-bomb inhibitors. Two patients were treated by a Nissen fundoplication, 4 and 6 month after diagnosis, respectively. One patient with severe neurological damage will undergo the same surgery soon. In another patient with caustic esophageal injury, the affected esophagus will be resected, restoring continuity with stomach or colon. In view of the potential oncogenous transformation of BE, the importance of not overlooking this anomaly in all patients with severe GER is highlighted. All cases with predisposing factors to develop BE should be closely followed by periodic examination and multiple biopsies. PMID- 10533658 TI - [Amebic liver abscess in newborn. Report of a case]. AB - The amebic liver abscess in newborns is an uncommon disease. There are few cases reported in the literature. This is a case of a 20 day-old female newborn who presented an abdominal mass, yellowish diarrheic depositions and jaundice. An abdominal CT scan showed a cystic mass located in the right hepatic lobe. The laboratory exams confirmed the amebic etiology. The clinical treatment failed and the surgery was decided. The pathologic results confirmed the diagnosis of an amebic liver abscess. Eight days after the resection the patient died because of a necrotizing enterocolitis. PMID- 10533659 TI - [Gastric solitary plasmacytoma associated with +Helicobacter pylori infection]. AB - Solitary gastric plasmacitomas are infrequent tumors. They account for 5% of the extramedullary plasmacitomas. We report an unusual case in a 14 years old boy. The patient has had gastric symptoms for 2 years prior to an endoscopic examination. A fungating, ulcerated lesion was observed in the antrum. The biopsies showed a monoclonal, Lambda positive, diffuse, plasmocitic proliferation infiltrating the mucosa. Also a moderate number of Helicobacter pylori were identified in the gastric pits and numerous lymphoid follicules were observed in the deep portion of the mucosa. In view of the presence of HP infection the patient was treated with Orneprazole and Clarithromycin. Endoscopic examination and biopsies performed 3 and 5 months later showed a complete remission of the gastric lesion. At the time of this report the patient is in good physical condition, has recovered his weight and has grown 5 cm. Differential diagnosis with plasmo limpho in chronic gastritis and with lympheicitic lymphoma with plasmocitoid features had to be done. The macroscopic appearance of the gastric lesion, the absence of other inflammatory cells and monoclonality of the plasmocitic infiltration ruled out chronic gastritis. The negative staining for CD 20 as well as the abscence of lymphoid cells in the mucosal infiltrate give support: to the diagnosis of plasmocitoma. The close association between gastric MALT lymphoma and HP infection has been reported as well as its regression after antibiotic treatment for its erradication. In our review of the literature we failed to find any references to the association of HP with gastric plasmocitoma nor to its regression after antibiotic therapy. PMID- 10533660 TI - [The physician-patient relationship in post-modernism]. AB - The doctor-patient relationship represents a particular link that goes beyond formality and it is projected in time, space and emotionalism. It takes place in the midst of a cultural, social, physiological, scientific and technical context which at the same time is conditioned by local, regional, national and foreign influences as a consequence of globalization. Different ethnical groups, social stratification, economic structures, philosophical concepts, religious beliefs and moral values play a pivotal role. The technological advances have shown an exponential increase during the last forty years. This phenomenon was accompanied by deep changes that varied from philosophical to strictly technical aspects which were significatively intensified during the post Second World War period. The consequences have been unfortunate for physicians and particularly for patients. Those consequences came out by philosophical reviews, the economical liberalism and neoliberalism development, a new concept for science, the priorization of technique development together with changes in both ethical value scales and economic & politic power hegemonies. The great reports of philosophy have tried to justify these social changes. The end of modern meta discourses such as Iluminism, Idealism, and Marxism, incited the liberal and neoliberal discourse. Knowledge has been affected not only in the research field but in the teaching field as well. Today, the discourse in vogue is the performance. The "organic conjugation" of technique and profit precedes its union with science. The State disengagement in managing the great social problems in many countries, the globalization and the concentration of the capital has redefined the power. Neither the patient nor the medicine escapes from this reality. The final results are generations of physicians quite right informed but unable to solve or even face the minimum problems of a patient, and what is worst, they run the risk of feeling frustrated and resentful. Everything is thought and done through technique, and there is a fact that appears to justify this action "The malpractice judgement industry". The economical monopoly of private or sindical Healthcare Providers and self-managed Hospitals projects are rapidly and notoriously modifying the medical labour market. There seems to exist only one element that the doctor and the patient have in common in this new culture: loneliness. There is also a human communication failure that separates them: the patient is in more need of human comprehension than ever before and the doctor has a cultural inability to offer it. To sum up, the relationship between the doctor and the patient is still a theme of deep preoccupation, even more nowadays when everything seems to indicate that this relationship has suffered a significant impairment. PMID- 10533661 TI - [Markers of hepatitis C virus infection in dialysis units. A decreasing problem but still present in Argentina]. PMID- 10533662 TI - [Barrett's esophagus in children]. PMID- 10533663 TI - [Peptic ulcer at the beginning of the 3rd millennium. Perspectives of its treatment]. PMID- 10533664 TI - [Functional capacity of the heart in cardiac insufficiency. A French method for evaluation]. PMID- 10533666 TI - [Preservation of the native aortic valve in the treatment of aortic aneurysmal dilatation]. AB - Mechanical valve conduit replacement of the aortic root is a durable and appropriate procedure for aortic root dilatation with or without aortic aortic insufficiency. But this procedure may sacrifice an anatomically salvageable aortic valve and requires a life-long anticoagulation with its attendant thromboembolic versus haemorrhagic risks, which is not ideal for young active patients. Recently, two techniques of aortic root replacement with aortic valve sparing have been described, based on experimental data. The first one is Yacoub's procedure (1983), where the correction of the aortic root is performed by correcting the sinotubular junction and replacement of the aortic sinuses with an appropriately tailored Dacron graft (remodelling). The second technique was described by David (1992). In this one, the aortic root reconstruction is performed by reimplanting the aortic valve in a tubular Dacron graft (reimplantation). Since 1993, we have been interested in these procedures and the aim of this study was to examine the perioperative and intermediate term results of these techniques. From 1993 to 1998, 14 patients had either reimplantation of the aortic valve (3 patients) or remodelling of the aortic root (11 patients). Patients' ages ranged from 17 to 57 years (40.2 +/- 7.9 years). Four patients had Marfan's syndrome (29%). There were 11 cases of aortic insufficiency, three 1+ (21.4%), 3+ (21.4%), seven 2+ (50%) and one 3+ (7.2%). All the patients had morphologically normal aortic leaflets. The mean diameter of the sinuses was 57 +/- 4 mm. There was no acute or chronic dissection. The left ventricular function was measured as the percentage of the fractional shortening of the left ventricular diameter. The mean of the fractional shortening was 38.3 +/- 4%. There was no mortality and all patients underwent early and late follow-up echocardiography. The 14 patients have only mild or no insufficiency, which has not progressed in any patient. No other valve-related complication has occurred. Aortic valve-sparing replacement of the aortic root is an excellent procedure for patients with aortic root dilatation and anatomically salvageable valves. The long-term results are still unknown but it seems an attractive alternative to composite replacement of the aortic valve and descending aorta. Morbidity and mortality rates are very low, even lower than a Bentall procedure, while the savings in cost and human lives due to the absence of a mechanical valve prosthesis are significant. PMID- 10533665 TI - [Evaluation of a specific French scale of activity in chronic heart failure. A national multicenter study. Group for Cardiac Insufficiency and Cardiomyopathy of the French Society of Cardiology]. AB - Many systems have been proposed to evaluate the functional incapacity caused by chronic cardiac failure. The classification of the New York Heart Association (NYHA) is the best known. It is subjective, poorly reproducible and has a poor predictive value on effort. The authors propose a Specific French Scale of Activity with the object of a more accurate functional evaluation of cardiac failure, easier to use by the doctor and more specific to French patients and their life styles. A French multicentre study was set up in hospital departments by the French Society of Cardiology working group on Cardiomyopathy and Cardiac Failure to assess this new classification with respect to the NYHA classification and peak VO2 (Weber's classification). Eight centres participated in the study. A total of 124 patients with chronic cardiac failure and a mean age of 61 years (102 men) were included. Cardiac failure was due to ischaemic heart disease in 72 cases, hypertension in 10 cases, dilated cardiomyopathy in 40 cases and aortic regurgitation in 2 cases. Eighty-two patients underwent a double evaluation using the French Scale: 40 patients by 2 physicians and 42 patients by a physician and a nurse. Good reproducibility was found between the assessment by the 2 physicians in 35 cases (87%) and between the physician and nurse in 30 cases (71%). When compared with peak VO2, the classification was concordant in 47% of cases using the NYHA and in 61% of cases using the French Scale, with variation of one class in 40% of cases with the NYHA and 35% of cases with the French Scale. These results show good reproducibility and correspondence of classification with the exercise test which was better using the French Scale than the NYHA classification. PMID- 10533667 TI - [MRI quantification of regional variations of left ventricular parietal stress in normal subjects]. AB - The object of this study was to analyse regional variations in end systolic left ventricular wall stress in normal subjects using three-dimensional magnetic resonance imaging (MRI) with excellent spatial resolution. Eight to 12 contiguous short axis sections of the left ventricle were acquired from the apex to the base in apnoea with a rapid echo-gradient sequence in 15 healthy volunteers. The end systolic wall stress was calculated by three methods: Grossman's formula (CR) using the wall thickness and radius of curvature, Janz's formula (CS) using the surfaces, and a three-dimensional approach (C3D) providing a precise calculation of the radius of curvature. The values of wall stress obtained by CS and CR were lower (p < 0.001) at the apex (3.2 and 3.3 10(3) newton/m2 respectively) than at the base (6.9 and 7.1 10(3) newton/m2). There was no difference between the base and apex with the C3D method (8.0 and 9.0 10(3) newton/m2 respectively, NS). The same results were observed at the inferior, lateral, anterior and septal segments with an increase at the base using the CS and CR formulae, the C3D remaining homogeneous in the left ventricle except for the interventricular septum. The lateral wall stress was significantly lower with respect to the interventricular septum in all sections from the apex to the base, irrespective of the method of calculation used. The differences in regional wall stress from the base to the apex reported in healthy subjects seem to be related to an underestimation of left ventricular wall thickness and an underestimation of the radius of curvature rather than to a physiological phenomenon. PMID- 10533668 TI - [Thoracic radiodermatitis in interventional cardiology. Apropos of 6 cases]. AB - The aim of this study is to describe thoracic radiodermatitis, a rare but not to be forgotten complication of interventional cardiology. The appearances are variable, from often oval-shaped erythema to cutaneous necrosis, with risk of chronic ulceration and malignant degeneration. The authors report 6 cases observed in 1997 after coronary angioplasty. Complex and long procedures are the main causes of this complication. Prevention requires a contribution from all the medical cardiological team, for the diagnosis, determining the indication of the type of revascularisation and for limiting the dose of X-radiation administered. PMID- 10533669 TI - [Cytomegalovirus and arterial disease. Current aspects]. AB - The cytomegalovirus (CMV) is the only microbial agent implicated in three particular types of arterial disease: coronary disease of the transplanted heart, post-angioplasty restenosis and atherosclerosis. The object of this article is to analyse the recent data on the role of CMV in these pathologies with an exhaustive review of the literature. The available data is mainly epidemiological but the interpretation is difficult because of the multiplicity and imperfections of the diagnostic techniques of the infection. However, the results are quite concordant in favour of a real association. Different physiopathological mechanisms are proposed. In coronary disease of the transplanted heart, the lesions could be initiated by an inflammatory process. In post-angioplasty restenosis, the virus seems able to trigger cellular proliferation by inhibiting the mechanisms of apoptosis. Finally, in atherosclerosis, CMV infection seems to promote atherothrombotic processes and accelerate the progression of atherosclerotic plaques by activating inflammatory cells. Direct methods of detection of viral DNA show the presence of the virus within these lesions. There is, therefore, epidemiological, anatomo-pathological and physiopathological evidence in favour of a relationship between CMV infection and these three forms of arterial disease. PMID- 10533671 TI - [Mycotic aneurysm of the splenic artery. A rare complication of surgically treated infectious endocarditis and its causative cardiac lesion]. AB - The authors report the case of a 59 year old woman with mitral valve streptococcal endocarditis complicating rheumatic valvular disease with several metastatic septic complications. In addition to ocular and cerebral localisations, the patient developed a very rare mycotic aneurysm of the splenic artery. Mitral valve replacement was necessary because of severe mitral regurgitation with major dilatation of the left heart chambers. This surgery was performed under high dose heparin therapy. Large aneurysms of the splenic artery carry a high risk of rupture. This splenic artery aneurysm was treated in the same operative session as the valvular disease by a sternolaparotomy: the aneurysm was operated first of all, and then valvular replacement was performed. Three years later, the patient is well and cured of the endocarditis. To the authors' knowledge, this is the third report of mycotic aneurysm of the splenic artery and the first case combined with surgery of the infectious valvular disease and the gastro-intestinal artery aneurysm. PMID- 10533670 TI - [Appetite suppressants and heart valve disorders]. AB - Serotoninergic appetite-suppressant drugs, fenfluramine and dexfenfluramine, were withdrawn from the market in September 1997 on account of two major cardiopulmonary complications: primary pulmonary hypertension and valvular regurgitation. The valvular heart diseases involve mainly left-sided valves, and contrary to physiological valvular regurgitations, they appear mostly on the aortic valve. Prolonged exposure (> 3 months) appears to confer a higher risk of cardiac valve involvement. Pathological features are similar to carcinoid or ergot alkaloid-induced valve diseases, and suggest a common pathophysiological mechanism which would also explain pulmonary hypertension by the toxic effect of high levels of circulating serotonin. After the first reports documenting a dramatically high prevalence of valvular side effects (up to 33% according to the Food and Drug Administration), recent studies reported a lower prevalence and severity. The long-term outcome and the real incidence are unknown and require further research and epidemiological data. A clinical survey of the patients exposed to serotoninergic appetite-suppressants is necessary, to be repeated 6 to 8 months later in the absence of an initial cardiac murmur. Doppler echocardiographic examination should be performed after prolonged exposure (> 3 months) or a high dosage of these drugs, in circumstances such as the presence of cardiovascular symptoms, a cardiac murmur, or an uncertain cardiac examination because of weight of patients. PMID- 10533672 TI - [Extensive mycotic coronary aneurysm detected by echocardiography. Apropos of a case]. AB - The authors report the case of a large mycotic right coronary aneurysm detected at echocardiography in a 45 year old patient with AIDS. Although emergency surgery was planned, the patient died of rupture of the aneurysm with cardiogenic shock and sudden pericardial tamponade. This case underlines the diagnostic value of echocardiography, by the transthoracic approach for para-cardiac masses and with the transoesophageal probe for accurate localisation and demonstration of the coronary origin. In this case, the CT scan was less useful than transthoracic echocardiography. Coronary angiography confirmed the strongly suggestive echocardiographic diagnosis and helped decide management strategy. Atheromatous coronary aneurysms may be treated by stenting but mycotic aneurysms require surgical management. PMID- 10533673 TI - [Paroxysmal ischemic mitral valve insufficiency]. AB - The prognosis of patients with coronary artery disease may be threatened by ischaemic mitral regurgitation. Apart from rupture of a papillary muscle which requires rapid valve replacement. Chronic ischaemic papillary muscle dysfunction can often be a severe complication of ischaemic heart disease. The authors report the case of a patient with dyspnoea but no angina of effort. Cardiovascular investigations with right heart catheterisation demonstrated the occurrence of severe mitral regurgitation only during angioplasty of the left marginal artery. PMID- 10533674 TI - Difference in bronchoprotective effects of bronchodilators on postallergic propranolol-induced bronchoconstriction. AB - Administration of propranolol provokes bronchoconstriction only in asthmatic patients. It is unknown whether bronchodilator therapy can prevent the propranolol-induced bronchoconstriction (PIB). We previously reported an animal model of PIB in which bronchoconstriction is caused by propranolol when inhaled 20 minutes after an antigen provocation in passively sensitized guinea pigs. Our goal was to evaluate the bronchoprotective effects of bronchodilators on the PIB in our animal model. Propranolol was inhaled 20 minutes after an antigen challenge in passively sensitized, anesthetized, and artificially ventilated guinea pigs. Atropine (5 mg/kg) and equipotent doses of salbutamol (0.6 microgram/kg) and aminophylline (25 mg/kg), which were determined by the dose response curves for inhibition of histamine-induced bronchoconstriction, were intravenously administered 5 minutes before the propranolol inhalation. Treatment of the animals with 25 mg/kg of aminophylline, but not with 0.6 microgram/kg of salbutamol or 5 mg/kg of atropine, significantly prevented the PIB. These results show that our animal model is an experimental model of PIB which is resistant to beta 2-agonists or anticholinergics and suggest that aminophylline may be useful to prevent and treat PIB resistant to beta 2-agonists. PMID- 10533676 TI - Assessment of long-term bronchiolar clearance of particles from measurements of lung retention and theoretical estimates of regional deposition. AB - Twelve healthy nonsmokers inhaled monodisperse Teflon particles labelled with 51Cr (half-life 27.8 days) with an aerodynamic diameter (dae) of 6.1 microns, 5 at a normal flow, 0.5 L/s, and 7 at an extremely slow flow, 0.05 L/s. Lung retention after 24 hours was measured for about 6 months and could be well described by a 2-component exponential function. After the normal inhalation, 14% of the particles retained after 24 hours cleared with a half-time of 3.7 days and 86% with a half-time of 217 days. After the slow inhalation, 35% of the particles retained after 24 hours cleared with a half-time of 3.6 days and 65% with a half time of 170 days. Deposition was calculated using 3 different models including the recent Human Respiratory Tract Model (HRTM), adopted by the International Commission on Radiological Protection (ICRP), and a model based on Monte Carlo particle transport, together with an asymmetric lung model. Generally, the 3 models agreed fairly well and predicted a considerably higher deposition in the bronchiolar region (generations 9-15) at the slow flow than at the normal flow. Together, the experimental data and the predictions of the deposition models indicate that about 40% of the particles deposited in the conducting airways during the slow inhalation were retained after 24 hours. They also strongly indicate that the particles which cleared with a half-time of about 4 days were mainly deposited in the bronchiolar region, and that about 25% of the particles deposited in the bronchiolar region cleared in this phase. The experimental data agreed quite well with the HRTM predictions made using its default parameter values for slow clearance in the bronchial tree. PMID- 10533675 TI - Antagonistic effects of pyrrolidine dithiocarbamate and N-acetyl-L-cysteine on surfactant protein A and B mRNAs. AB - Pulmonary surfactant, a mixture of phospholipids and specific associated proteins, reduces surface tension at the air-liquid interface of the lung and protects the large epithelial surface of the lung from infectious organisms. Surfactant proteins, SP-A and SP-B, are required for normal surfactant function. In the current work, increased levels of oxidized glutathione (GSSG) are demonstrated at doses of pyrrolidine dithiocarbamate (PDTC) which decrease SP-A and SP-B mRNAs, suggesting that cellular oxidation reduces surfactant protein expression. Similarly, reduction of SP-A and SP-B mRNA levels following accumulation of GSSG induced by glutathione reductase inhibitor 1,3-bis-(2 chloroethyl)-1-nitrosourea (BCNU), supports the hypothesis that surfactant protein synthesis is reduced in response to oxidation of pulmonary epithelial glutathione. Concurrent induction of apolipoprotein J (apoJ) mRNA by PDTC demonstrates the selectivity of pulmonary gene regulation by the dithiocarbamate. In contrast, the glutathione precursor N-acetyl-l-cysteine (NAC) prevented PDTC dependent increase in GSSG/GSH ratio, inhibition of SP-A and -B mRNAs, and induction of apoJ. Insufficiency of SP-A and -B, which occurs in inflammatory lung diseases, may result from the exposure of the pulmonary epithelium to oxidant stress and may be reversed by the antioxidant NAC. PMID- 10533677 TI - Indomethacin does not influence alveolar liquid clearance in anesthetized sheep or rats. AB - Active transport of sodium has been shown to be the predominant mechanism involved in alveolar liquid clearance. One regulatory mechanism involved in the modulation of this transport system is cAMP. Although it was initially thought that cAMP could directly modulate transepithelial Na+ transport, recent data suggest that this cAMP modulation could be secondary to the production of arachnidonic acid metabolites. The purpose of this study was thus to evaluate if prostaglandin products could have an indirect or direct role to play in lung liquid clearance. Addition of 10(-5) M salmeterol, known to increase intracellular cAMP, to the instilled fluid in rats stimulated lung liquid clearance. However, addition of indomethacin did not influence the stimulating effect of salmeterol. Furthermore, addition of prostaglandin E2 to the instilled fluid did not stimulate alveolar fluid clearance. In order to determine if this response could be species related, we evaluated if indomethacin could modulate alveolar liquid clearance in sheep. Presence of cAMP and aminophylline stimulated lung liquid clearance in sheep, but indomethacin did not inhibit this response. The present study demonstrates that cyclooxygenase products are not involved in the modulation of basal or stimulated alveolar or lung liquid clearance in sheep or rats. PMID- 10533678 TI - The effect of enoxaparin on bleomycin-induced lung injury in mice. AB - We have evaluated the effect of enoxaparin, a potent antithrombotic drug, on bleomycin (Bleo)-induced pulmonary inflammation in mice. Pulmonary injury was induced by a single intratracheal (i.t.) instillation of Bleo. Four groups of female C57BL/6 mice, each received one of four treatments: (1) i.t. Bleo and daily intraperitoneal (i.p.) injections of enoxaparin (EN) starting one day before i.t. instillation of Bleo (Bleo-EN); (2) i.t. Bleo and i.p. injections of saline (Bleo-Sal); (3) i.t. saline and i.p. enoxaparin (Sal-EN); (4) i.t. saline and i.p. saline (Sal-Sal). Animals were sacrificed 14 days after i.t. treatment. Lung injury was evaluated by analysis of bronchoalveolar lavage fluid and histologically by an overall semiquantitative index of lung injury and a quantitative image analysis assessing alveolar wall area fraction and fibrosis fraction. Treatment of mice with enoxaparin did not ameliorate Bleo-induced lung injury. Our study does not establish a critical role of procoagulant activity in the evolution of Bleo-induced lung injury and does not support the use of antithrombotic therapy for the prevention of pulmonary fibrosis. PMID- 10533680 TI - Needlestick-prevention devices. PMID- 10533681 TI - Cell phones and walkie-talkies. Is it time to relax your restrictive policies? AB - In this article, ECRI revisits the issue of whether--and to what degree- healthcare facilities should restrict the use of cell phones and walkie-talkies. Although interference between these communications devices and medical devices has been demonstrated, the risk to patients appears to be minimal, as well as manageable. Based on the evidence available to date, we believe that restrictive policies can be relaxed to allow the use of cell phones and walkie-talkies in certain situations. (This is not to say that all restrictions should be lifted, however.) As always, we welcome your questions and comments, especially as they relate to experiences at your facility. PMID- 10533679 TI - Tyloxapol confers durable protection against hyperoxic lung injury in the rat. AB - We tested the hypothesis that the nonlipid components of Exosurf, tyloxapol (TY), and cetyl alcohol (CA), protect against hyperoxic lung injury by induction of the animals' endogenous antioxidant defenses. Adult rats were intratracheally instilled with escalating doses of TY and CA (n = 20) or TY alone (n = 32) and immediately exposed to 100% oxygen. Intratracheal instillation of TY alone or in combination with CA protected against lethal hyperoxic injury in the rat in a dose-dependent fashion. To assess the effects of timing, rats were instilled with TY plus CA 24 hours before (n = 6) and 24 hours after (n = 6) exposure to 100% oxygen, with time to death determined. Rats were also instilled with TY alone at 0 hour (n = 6), 48 hours (n = 3), 96 hours (n = 3), and 186 hours (n = 4) prior to exposure to 100% oxygen. Lungs were assayed for superoxide dismutase, glutathione peroxidase, and catalase activities. Finally, the pharmacokinetics of TY in the rat lung were determined by instilling radiolabelled TY intratracheally. TY has a prolonged half-life in the rat lung, and protection against lethal hyperoxic injury was achieved by a single intratracheal dose delivered up to 186 hours prior to injury. Antioxidant enzymes were not induced in protected animals. We conclude that TY provides durable protection against hyperoxic lung injury without induction of antioxidant enzymes. It is present in the lung for sufficient duration to invoke a direct mechanism of protection, possibly as a radical scavenger. These findings raise the prospect of a therapeutic application for TY as prophylaxis in patients at risk for oxygen toxicity and adult respiratory distress syndrome. PMID- 10533683 TI - Lorad T-350 mobile mammography system does not indicate failed clinical exposure. PMID- 10533682 TI - Y2K compliance report. Problems affecting clinical laboratory equipment. PMID- 10533684 TI - Long reset time of Hamilton Medical Galileo intensive care ventilator puts patients at risk. PMID- 10533685 TI - Failure of GE Marquette Solar 8000 physiologic monitor display. PMID- 10533686 TI - Cracking cases on Linvatec Hall Versipower Plus battery packs. PMID- 10533687 TI - The consequences and opportunities of shortened lengths of stay for cardiovascular patients. AB - Length of hospitalization for many cardiovascular conditions has decreased dramatically recently. Many attribute this increase to the cost constraints imposed by managed care and to improved technologies. Although technological developments have contributed to cost inflation, they have also reduced the need for long hospitalizations and improved patient outcomes. This new context for delivering cardiovascular services is discussed in terms of the challenges and opportunities for nursing leadership in several aspects of care. PMID- 10533688 TI - Preoperative psychosocial predictors of hospital length of stay after heart transplantation. AB - The effect of psychosocial factors on hospital length of stay (LOS) after heart transplantation has not been reported. This study examines relationships between preoperative psychosocial variables and LOS and identifies preoperative psychosocial predictors of LOS after transplant. A nonrandom sample of 307 patients at two medical centers completed a self-administered booklet of psychosocial measures. A chart review was also conducted. Psychosocial problems included anxiety, stress, and inadequate coping; questionable understanding of heart failure and treatment; substance abuse; and noncompliance. Self-care disability, a history of noncompliance, and more emotional disability predicted 8% of LOS. This supports the inclusion of psychosocial issues and functional disability in post-heart transplant clinical pathways. PMID- 10533689 TI - Learning retention in patients receiving midazolam during permanent pacemaker implantation. AB - This pilot study (N = 20) tested the effects of intravenous midazolam administration on learning retention after pacemaker implantation. Patients were randomized to receive teaching at 1 or 3 hours after the last dose of midazolam. Using a standardized teaching format, one of two study nurses performed the teaching that included incision care, activity restrictions, environmental factors potentially affecting pacemaker function, and follow-up requirements. Learning was evaluated by one of the investigators blinded to teaching time. Subjects in the 1-hour group retained significantly less information than those taught at 3 hours after drug administration. Patients taught later answered similar numbers of questions correctly, whereas there was much more variability in correct responses for the group taught earlier. This article reviews the effects of midazolam on memory and learning as well as provides suggestions for alterations in patient education protocols for patients receiving midazolam for pacemaker implantation. The effect of shortened length of stay on care practices is also discussed. PMID- 10533690 TI - Caregiver satisfaction with preparation for discharge in a decreased-length-of stay cardiac surgery program. AB - Preparing caregivers of cardiac surgery patients for early discharge is an essential component of patient care. This study examined caregiver satisfaction with preparation for discharge in a decreased length of stay cardiac surgery program. Data were obtained from caregivers (N = 53) of cardiac surgery patients discharged on postoperative day 4 or 5. Data were analyzed with regard to caregiver satisfaction with preparedness for discharge, preference for a longer hospitalization, benefit of an earlier discharge, and patient care expectations. Results indicted that the majority of caregivers preferred earlier discharge but did not feel prepared for the responsibility of patient care. Implications for education and support for the caregiver before early discharge are addressed. PMID- 10533691 TI - The effects of a discharge planning and home follow-up intervention on elders hospitalized with common medical and surgical cardiac conditions. AB - This study was a secondary analysis of data collected on 202 patients hospitalized with common medical or surgical cardiac conditions who completed a 24-week postdischarge follow-up program as part of a large-scale randomized clinical trial. Subjects were age 65 years or older, admitted from their homes with one of the following diagnosis-related groups: heart failure, angina, myocardial infarction, coronary artery bypass graft surgery, or cardiac valve replacement. The intervention consisted of comprehensive discharge planning and home follow-up by an advanced practice nurse (APN) for 4 weeks after discharge. Control subjects received usual care. Findings indicated that medical patients in the intervention group had fewer multiple readmissions during the 24 weeks of follow-up and a reduced total number of days of rehospitalization. There were fewer hospital readmissions in the surgical group when measured from discharge to 6 weeks. There were no differences in functional status between intervention and control groups for either population. The findings of this study suggest that high-risk elders with significant cardiac problems may benefit from a care program that emphasizes collaborative, coordinated discharge planning and home follow-up that includes telephone and home visits by APNs. PMID- 10533692 TI - Implementing a congestive heart failure disease management program to decrease length of stay and cost. AB - Congestive heart failure (CHF) is the most common reason for a hospital admission in the Medicare age group and is nearly double the rate of pneumonia, the next highest volume diagnosis. The economic burden of this debilitating, chronic disease demands a mechanism to improve quality of care while preventing unnecessary hospitalizations. Beginning in 1995, Evanston Northwestern Healthcare (ENH) created a disease management program involving a multidisciplinary team designed to decrease length of stay (national average = 6.2 days; ENH = 4 days), reduce costs, prevent readmissions (national 30-day readmission rate = 23%; ENH CHF Program = 2.3%), and improve compliance with the treatment regimen. Compliance monitoring through an automated telemanagement program reinforces education, identifies early warning signs and reduces the likelihood of hospitalization. After 18 months, telemanagement participants' compliance rate averages 89.5%. CHF hospitalization rates are 0.6/patient/year compared with the national benchmark of 1.7/patient/year. A disease management program consists of inpatient consultation, education, outpatient CHF clinic, cardiac home care, and compliance monitoring. Throughout this continuum, education must be communicated consistently by all team members. A CHF Assessment Guide assists the multidisciplinary team to thoroughly complete all education and address unique solutions to patients' needs. PMID- 10533693 TI - Envisioning a network of care for at-risk patients after myocardial infarction. AB - Myocardial infarction (MI) continues to be a significant health care issue because of its prevalence. As treatment options improve the survival rate, an increasing number of individuals have to learn how to adjust to this major life event and prevent recurrence. Recovery can be difficult. Many patients experience emotional distress, fear of dying, and family turmoil, fail to return to work when physiologically capable of doing so, are unable to return to their previous levels of sexual activity, and are not capable of making the necessary diet and exercise changes. Acute management strategies continue to be aimed at limiting the infarct size, whereas holistic approaches to the patient and family adjustment must target seeking prompt treatment when symptoms present, psychologic adjustment, stress reduction, and patient and family education for self-care and risk reduction. As hospital length of stay for acute MI decreases, health care professionals must provide an interdisciplinary, collaborative approach to ensure that the at-risk MI patient is provided all of the information and support needed to lead a satisfying, productive, healthy life. An excellent way for nurses to not only address this challenge, but to lead the effort, would be to develop a network of care for the at-risk MI patient. PMID- 10533694 TI - Outcomes measurement. AB - Reductions in hospital lengths of stay (LOS) for patients with cardiovascular conditions can be a cost-effective in-hospital outcome, but the effect of shortened hospital LOS on patient and family outcomes after discharge needs to be evaluated. Suggestions for the use of appropriate data to evaluate LOS and outcomes that need study are presented. PMID- 10533695 TI - Pharmacokinetic and pharmacoimmunodynamic interactions between prednisolone and sirolimus in adrenalectomized rats. AB - Prednisolone (Pred) and sirolimus (SIR) are immunosuppressive compounds acting through different mechanisms with moderate synergism found in vitro. Both drugs are metabolized partly by CYP3A enzymes. After i.v. administration of placebo, Pred (5 mg/kg), SIR (1 mg/kg), or Pred with SIR (5 and 1 mg/kg doses) to adrenalectomized male rats, Pred plasma and SIR whole blood concentrations were followed for 48 hr along with circulating T-helper and T-cytotoxic cell counts. Ex vivo whole blood lymphocyte proliferation marked host responsiveness. An extended indirect PK/PD model was used to describe responses to these drugs, alone or combined. An interactive two-stage population analysis showed no modification in drug PK. Mean Pred plasma clearance was 0.655 L/hr (interrat++ variability: 11%) and significantly increased with weight. Mean SIR whole blood volume of distribution and clearance were 5.6 L (62%) and 0.28 L/hr (32%), and animal scaling showed weight-power proportionality. In vitro metabolism studies showed no significant inhibition of Pred or prednisone CYP3A metabolism by SIR (50 microM), but this pathway accounted for less than 5% of Pred metabolism. Pred decreased numbers of T-helper lymphocytes with a mean IC50 of 37.8 nM (21%) alone or 12.3 nM (130%) with SIR. Results for T-cytotoxic lymphocytes were similar. SIR increased lymphocyte numbers with a mean IC50 of 52.2 nM (24%) for T-helper and 28.8 nM (51%) for T-cytotoxic cells. Taking into account drug effects on lymphocyte trafficking, Pred directly inhibited ex vivo lymphocyte proliferation with a mean IC50 of 1.08 nM (38%). SIR, after a transduction step, inhibited proliferation with a mean IC50 of 1.00 nM (26%). Responses measured after drug coadministration were reasonably quantitated by addition of single drug effects. Since, at pharmacologic concentrations in rats, Pred and SIR did not interact in their PK but synergistically or additively interact in their dynamics, their joint therapeutic use is promising. The adrenalectomized rat may be a suitable animal model to characterize drug effects on lymphocyte trafficking and reactivity. PMID- 10533696 TI - Mathematical modeling of circadian cortisol concentrations using indirect response models: comparison of several methods. AB - Six mathematical functions to describe the chronobiology of cortisol concentrations were assessed. Mean data from a dose-proportionality study of inhaled fluticasone propionate were fitted with an indirect response model using various biorhythmic functions (single cosine, dual ramps, dual zero-order, dual cosines, and Fourier series with 2 and n-harmonics) for production rate. Data with known parameters and random variation were also generated and fitted using the ADAPT II program. Fitted parameters, model estimation criteria, and runs tests were compared. Models with preassigned functions: the dual ramps, the dual zero-order and the dual cosines provide maximum and minimum times for cortisol release rate, were suitable for describing asymmetric circadian patterns and yielding IC50 values. Fourier analysis differs from the other methods in that it uses the placebo data to recover equations for cortisol secretion rate rather than by postulation. Nonlinear regression for Fourier analysis, instead of the L2 norm method, was useful to characterize the baseline cortisol data but was restricted to a maximum of two harmonics. Apart from the single cosine function, which predicts symmetrical cortisol concentrations, all methods were satisfactory in describing the baseline and suppressed cortisol concentrations. On the other hand, Fourier series with L2-norm produced the best unbiased estimate for baseline patterns. The Fourier method is flexible, accurate, and can be extended to other drug-induced changes in normal periodic rhythms. PMID- 10533697 TI - In vitro characterization of the erythrocyte distribution of methazolamide: a model of erythrocyte transport and binding kinetics. AB - The rate and extent of binding of methazolamide to human erythrocytes was studied in vitro. All experiments were carried out at physiological temperature (37 C) and pH (7.4). Methazolamide (MTZ) buffer concentrations were analyzed by HPLC. Distributional equilibrium between buffer and washed red blood cells was achieved after 1 hr. Results of equilibrium studies were consistent with two classes of binding sites for MTZ within the erythrocyte: a low affinity, high capacity site (CA-I) and a high affinity, low capacity site (CA-II). A two-binding site model was fitted to experimental data generating estimates for binding parameters Ka1 (0.0017 +/- 0.00022 microM-1) nM1 (636 +/- 5.23 microM), Ka2(0.46 +/- 0.0083 microM-1), and nM2(80.9 +/- 0.389 microM). Based upon these findings, kinetic studies were performed in order to characterize the rate of drug distribution. The rate of erythrocyte uptake of MTZ was mathematically modeled using a series of differential equations describing drug diffusion across the red blood cell membrane and subsequent complexation with intracellular binding sites. The model assumed that penetration of MTZ into the red blood cells was passive but drug binding to the carbonic anhydrase isozymes was not instantaneous. Using a novel curve fitting technique, parameter estimates of RBC membrane permeability (0.0102 +/- 0.000618 cm/min), and binding rate constants k-1(0.254 +/- 0.0213 min-1), k1 (0.0022 +/- 0.00020 ml/microgram-min), k-2(1.59 +/- 0.0358 min-1), and k2(3.1 +/- 0.035 ml/microgram-min) were obtained. The model characterized the observed biphasic decline of MTZ buffer concentrations over time and may help explain the prolonged residence of MTZ in vivo. PMID- 10533699 TI - Optimal sampling times for Bayesian estimation of the pharmacokinetic parameters of nortriptyline during therapeutic drug monitoring. AB - Sampling times for Bayesian estimation of the pharmacokinetic parameters of an antidepressant drug, nortriptyline, during its therapeutic drug monitoring were optimized. Our attention was focused on designs including a limited number of measurements: one, two, and three sample designs in which sampling times had to be chosen between 0 and 24 hr after the last intake of a test-dose study. The optimization was conducted in four groups of patients defined by their gender and the administration or not of concomitant drugs inhibiting the metabolism of nortriptyline. The Bayesian design criterion was defined as the expected information provided by an experiment. A stochastic approximation algorithm, the Kiefer-Wolfowitz algorithm, was used for the criterion maximization under experimental constraints. Results showed that optimal Bayesian sampling times differ between patients in monotherapy and polytherapy. For one-sample designs the measurements have to be performed either at the lower (0 hr) or at the upper (24 hr) bound of the admissible interval. Replications were often found for 2- and 3-point designs. Other sampling designs can lead to criterion close to the optimum and can therefore be performed without great loss of information. In contrast, we found that several designs lead to low values of the information criterion, which justifies the approach. PMID- 10533698 TI - Comparison of different methods to evaluate population dose-response and relative potency: importance of interoccasion variability. AB - Different mixed-effects models were compared to evaluate the population dose response and relative potency of two albuterol inhalers. Bronchodilator response was measured after ascending doses of each inhaler in 37 asthmatic patients. A linear mixed-effects model was developed based on the approach proposed by Finney for the evaluation of bioassay data. A nonlinear mixed-effects (Emax) model with interindividual and interoccasion variability (IOV) in the different pharmacodynamic parameters was also fit to the data. Both methods produced a similar estimate of relative potency. However, the estimate of relative potency was 22% lower with the nonlinear mixed-effects model if IOV was not taken into account. Monte Carlo simulations based on a similar study design demonstrated that more biased and variable estimates of ED50 and relative potency were obtained when the nonlinear mixed-effects model ignored the presence of IOV in the data. Furthermore, the linear mixed-effects model that did not account for IOV produced confidence intervals for relative potency that were too narrow and thus could lead to erroneous conclusions. These problems were avoided when the estimation model could account for IOV. Results of the simulations were consistent with those of the experimental data. Although the linear or the nonlinear mixed-effects model may be used to evaluate population dose-response and relative potency, there are important differences in the assumptions made by each method. PMID- 10533700 TI - A comparison of methods for estimating individual pharmacokinetic parameters. AB - Characteristics of the methods for estimating individual pharmacokinetic parameters are compared both theoretically and numerically. The methods examined represent the range of most of modern methods and include the ordinary least squares, iteratively reweighted least squares, extended least squares, generalized least squares, maximum quasi-likelihood and its extended scheme, and minimum relative entropy methods. When the function representing the mean itself is used as a variance function, which may be then related to a Poisson distribution, the iteratively reweighted least squares estimator and maximum quasi-likelihood estimator are both identical to that of the minimum relative entropy method. These methods work by minimizing a kind of relative entropy between observed data and corresponding theoretical values. Furthermore, these methods guarantee agreement between the sum of the observed values and the estimate of the sum. This relation does not hold in general for the other estimators. The sum can, in a sense, be viewed as an approximation of the area under the curve. In addition, it is shown by numerical study that these methods are robust against the misspecification of the variance model and work as effectively as such sophisticated methods as the extended least squares, generalized least squares, and maximum extended quasi-likelihood methods. These sophisticated methods require complicated numerical optimization techniques and should be used only in cases where the estimation of the variance function is demanded. In the other cases, the method of minimum relative entropy or its equivalent is sufficient or even preferable for estimating individual pharmacokinetic parameters. PMID- 10533701 TI - Effect of sperm immobilisation and demembranation on the oocyte activation rate in the mouse. AB - To analyse the effect of the state of the sperm plasma membrane on oocyte activation rate following intracytoplasmic sperm injection (ICSI), three types of human and mouse spermatozoa (intact, immobilised and Triton X-100 treated) were individually injected into mouse oocytes. At 30, 60 and 120 min after injection, maternal chromosomes and sperm nuclei within oocytes were examined. Following human sperm injection, the fastest and the most efficient oocyte activation and sperm head decondensation occurred when the spermatozoa were treated with Triton X-100. Intact spermatozoa were the least effective in activating oocytes. Thus, the rate of mouse oocyte activation following human sperm injection is greatly influenced by the state of the sperm plasma membrane during injection. When mouse spermatozoa were injected into mouse oocytes, the rates of oocyte activation and sperm head decondensation within activated oocytes were the same irrespective of the type of sperm treatment prior to injection. We witnessed that live human spermatozoa injected into moue oocytes often kept moving very actively within the ooplasm for more than 60 min, whereas motile mouse spermatozoa usually became immotile within 20 min after injection into the ooplasm. In 0.002% Triton X-100 solution, mouse spermatozoa are immobilised faster than human spermatozoa. These facts seem to suggest that human sperm plasma membranes are physically and biochemically more stable than those of mouse spermatozoa. Perhaps the physical and chemical properties of the sperm plasma membrane vary from species to species. For those species whose spermatozoa have 'stable' plasma membranes, prior removal or 'damage' of sperm plasma membranes would increase the success rate of ICSI. PMID- 10533702 TI - Influences of zinc on fertilisation and development of bovine oocytes in vitro. AB - The effects of zinc (as ZnCl2) on in vitro production of bovine embryos (IVMFC) and components of the procedure, that is in vitro oocyte maturation (IVM), fertilisation (IVF) and embryo development in culture (IVC), and the effect of added zinc on sperm motility were studied. Immature cumulus oocyte complexes (COCs) were aspirated from ovarian follicles (2-5 mm diameter) at slaughter, and matured, fertilised and cultured in chemically defined conditions. The presence of zinc (10, 100 or 1000 micrograms added per millilitre) throughout IVMFC inhibited fertilisation. After addition of 10 micrograms zinc per millilitre separately to media for IVM and IVF, fertilisation was inhibited only when zinc was present for IVM. When present for IVF, 80% of oocytes selected for IVM reached 2- to 4-cell stages by 46 h after insemination whereas 67% of control oocytes (inseminated without added zinc) cleaved. Higher zinc concentrations (100 and 1000 micrograms added per millilitre) for IVF inhibited fertilisation. Sperm motility was reduced with addition of 10 micrograms per millilitre of zinc for sperm preparation (i.e. capacitation interval). Addition of 1.0 microgram zinc per millilitre to media used through IVMFC, or to the IVC medium alone, resulted in inhibition of development after 2- to 4-cell stages. When added to IVM or to both IVM and IVF media 1.0 microgram/ml of zinc compromised development to the morula stage and beyond. Maturing bovine oocytes may be more sensitive to 1.0 microgram ml of zinc in vitro than in vivo because a concentration of 3.0 micrograms/ml has been reported for bovine follicular fluid. Fertilisation was not adversely affected by 10 micrograms/ml of zinc; however, higher concentrations were inhibitory. PMID- 10533703 TI - Glutathione content and embryo development after in vitro fertilisation of pig oocytes matured in the presence of a thiol compound and various concentrations of cysteine. AB - The present study examined the effect of different concentrations of cysteine in the presence of a thiol compound, beta-mercaptoethanol (BME), during in vitro maturation (IVM) of pig oocytes on cumulus expansion, nuclear maturation, intracellular glutathione (GSH) level and subsequent embryonic development after in vitro fertilisation (IVF). In experiment 1, oocytes were matured in NCSU 23 medium containing 10% porcine follicular fluid, 25 microM BME, 0.5 microgram/ml LH, 0.5 microgram/ml FSH and 0, 0.1, 0.2 or 0.4 mg/ml cysteine for 20-22 h and then without hormonal supplements for an additional 20-22 h. After culture, cumulus cells were removed and a proportion of oocytes fixed to examine the rate of nuclear maturation. The remaining oocytes were co-incubated with spermatozoa for 5-6 h and putative zygotes were transferred to NCSU 23 medium containing 0.4% bovine serum albumin for 144 h. A proportion of putative zygotes were fixed 12 h after insemination to examine fertilisation parameters. In experiment 2, oocytes were matured as in experiment 1 and the GSH content was measured by a DTNB-GSSG reductase recycling assay. No mean differences among treatments were observed in nuclear maturation (78-89%). The mean differences in penetration rate (69-77%), polyspermy rate (31-40%), male pronuclear formation rate (93-96%) or mean number of sperm per oocyte (1.5-1.8) were not affected by the presence or absence of cysteine during oocyte maturation. Also no difference was observed in cleavage rates 48 h after insemination. However, compared with no addition (19%), the presence of 0.1-0.4 mg/ml cysteine during IVM increased (p < 0.001) the proportion of blastocysts (32-39%) at 144 h. In comparison with controls (5.6 pmol/oocyte), the GSH content of oocytes matured in the presence of cysteine was significantly (p < 0.001) higher (13-15 pmol/oocyte) with no mean differences among different cysteine concentrations. The results indicate that in the presence of a thiol compound, supplementation of IVM medium with cysteine can increase the GSH level and improve the developmental competence of pig oocytes following fertilisation. Further, no effect on either GSH level or embryo development was observed by increasing the levels of cysteine supplementation from 0.1 to 0.4 mg/ml. PMID- 10533704 TI - Hyaluronic acid and the cumulus extracellular matrix induce increases in intracellular calcium in macaque sperm via the plasma membrane protein PH-20. AB - The hyaluronic acid (HA)-rich extracellular matrix (ECM) of the cumulus oophorus is known to facilitate fertilization. It has been suggested that HA may enhance fertilisation in a number of species, and in macaque sperm, HA has been shown to increase the number of acrosome reactions that follow sperm binding to the zona pellucida. In this study, we investigated the effects of HA on intracellular Ca2+ in capacitated cynomolgus macaque sperm. Fluorometry studies using the intracellular Ca2+ indicator Fluo-3 showed that addition of 100 micrograms/ml of HA induced a rapid increase in intracellular Ca2+. This Ca2+ increase (approximately 2-3 times above basal levels) was inhibited by preincubation of sperm with Fab fragments of anti-recombinant PH-20 IgG. The frequency of acrosome reactions in sperm exposed to HA was not above control levels. A synthetic gel was prepared with similar viscosity to the cumulus and with HA trapped in its matrix. Video imaging of individual sperm was used to demonstrate that capacitated sperm swimming into the HA gel had increased intracellular Ca2+ levels. Preincubation of sperm with Fab fragments of anti-PH-20 IgG inhibited the increased intracellular Ca2+ levels induced by the HA gel. Sperm in control gel (no HA) did not show increased intracellular Ca2+, while sperm in gel containing anti-PH-20 IgG showed increased Ca2+ (positive control). Sperm loaded with Fluo-3 were allowed to interact with cynomolgus macaque cumulus masses, and sperm within the cumulus ECM clearly showed increased intracellular Ca2+ that was inhibited when sperm were preincubated in anti-PH-20 Fab. Fluorescein isothiocyanate (FITC) HA was found to bind to sperm over the acrosomal region, corresponding to PH-20 localisation, and this binding could be inhibited by preincubation of sperm with anti-PH-20 fragments. The results of this study show that HA increases intracellular Ca2+ in macaque sperm through interaction with plasma membrane PH 20. We propose that HA binding to plasma membrane PH-20 induces an aggregation of receptors that in turn results in intracellular signalling. As a result, sperm have higher basal CA2+ levels and are more responsive to induction of the acrosome reaction after binding to the zona pellucida. PMID- 10533705 TI - Effect of antibodies against seminal vesicle secretion on fertility in the rat. AB - Fertile female Wistar rats were immunised against rat and mouse seminal vesicle secretion (SVS) to test the production of allo-antibodies and the effect of the antibodies elicited on fertility. Twenty-six per cent of the rat and mouse SVS immunised females were infertile after the treatment. The sera were titrated by ELISA and used in Western blots to detect the proteins recognised. Although neither the antibody titres nor the proteins recognised by the sera showed a close relation with the degree of fertility, in all females the highest antibody titre in the fluids from the genital tract was found in the oviductal fluid and during the night of oestrus. This fact suggested that the site of action of the antibody could be the oviduct. Similar results were obtained using mouse SVS as immunogen--a fact that can be related to the antigenic similarity between the SVS of the two species. The antibodies react with the spermatozoa but not with eggs or embryos. Analyses performed on embryos collected from sterile females showed that there was a delay in fertilisation and normal embryogenesis. Our results suggest that SVS proteins are antigenic and that these antigens are bound to the spermatozoa and could take part in early pre-fertilisation events such as capacitation or sperm transport. PMID- 10533706 TI - Oocyte activation and parthenogenetic development of bovine oocytes following intracytoplasmic sperm injection. AB - Development of bovine oocytes after intracytoplasmic sperm injection (ICSI) was investigated. Oocytes were matured for 24-26 h in vitro and injected with isolated sperm heads. When treated with 7% ethanol (v/v) for 5 min, 71.7% of ICSI oocytes were activated as shown by the resumption of meiosis and the formation of female pronuclei. However, 41.5% of injected sperm heads remained condensed at 18 20 h after injection into the ooplasm. The incidence of decondensing sperm and that of male pronuclei at this stage were 15.1% and 26.4%, respectively. A total of 55.5% of oocytes reached the 2-cell stage following sperm head injection and 54.7% after sham-ICSI; these percentages were not significantly different from those following in vitro fertilisation (IVF) (73.1%). The percentage of 2-cell embryos reaching the 8-cell stage following ICSI was 37.5%, and 27.6% after sham ICSI, which were significantly lower (p < 0.01) than the equivalent percentage following IVF (62.4%). The percentages of parthenogenetic embryos reaching the 2 cell, 4-cell and 8-cell stages following ICSI were 56.4%, 48.9% and 30.0%, respectively. These results indicate that the low rate of normal embryonic development of bovine oocytes following ICSI is largely due to the parthenogenetic activation of the oocytes. PMID- 10533707 TI - Isolation and characterisation of zona pellucida A (ZPA) cDNAs from two species of marsupial: regulated oocyte-specific expression of ZPA transcripts. AB - The zona pellucida (ZP) is an extracellular glycoprotein coat that is deposited around the oocyte during folliculogenesis and performs several functions that relate to fertilisation and preimplantion development. In eutherian mammals it consists of three major glycoproteins--ZPA, ZPB, and ZPC--but little is known about its molecular constitution in marsupials. We have isolated the cDNA encoding the ZPA homologue in two distantly related marsupial series: the possum, Trichosurus vulpecula (a phalangerid) and the dunnart Sminthopsis crassicaudata (a dasyurid). The two cDNA sequences were 86% identical and showed extensive regions of homology to eutherian ZPA proteins, particularly in the central region of the molecule. Many other features of the ZPA protein, except the positioning of the N-linked glycosylation sites, were also conserved between marsupials and eutherians. ZPA expression was shown to occur maximally in the cytoplasm of the oocyte primary follicles with a little, but significant, expression in oocytes of both primordial follicles and in the cytoplasm of the oocyte in follicles with an antral cavity. No expression was seen in surrounding follicle or granulosa cells. PMID- 10533708 TI - DNA stability and thiol-disulphide status of rat sperm nuclei during epididymal maturation and penetration of oocytes. AB - DNA stability and thiol-disulphide status of rat sperm nuclei was observed in vivo during maturation in the epididymis and penetration of oocytes. When spermatids and spermatozoa were stained with acridine orange after fixation with acetic alcohol, the red/green fluorescence ratio observed under a confocal microscope was not different between spermatids (3.81 +/- 0.16) and testicular spermatozoa (4.03 +/- 0.34), and then decreased sharply (p < 0.01) as the spermatozoa descended the epidymis to the caput epididymis (1.13 +/- 0.03). However, the ratio was not different among corpus (0.69 +/- 0.01), cauda epididymis (0.68 +/- 0.11) and ejaculated spermatozoa (0.63 +/- 0.01). On the other hand, when spermatozoa were labelled with monobromobimane (mBBr), the fluorescence intensities gradually decreased (p < 0.01) during passage of spermatozoa from testis (4.74 +/- 0.16) through epididymis (caput, 2.72 +/- 0.08; corpus, 1.07 +/- 0.03; cauda, -0.05 +/- 0.05; ejaculated, 0.08 +/- 0.03). The acridine orange red/green fluorescence ratio increased (p < 0.01) during zona penetration (binding sperm, 0.52 +/- 0.09; perivitelline sperm, 0.64 +/- 0.16) and sperm decondensation (decondensed sperm, 0.69 +/- 0.12). When spermatozoa in the perivitelline space were labelled with mBBr, the fluorescence was detected. These results demonstrate that DNA stability against acid appears to be ahead of the oxidation of protamine during sperm maturation in the epididymis and is an initial event of the unpackaging process in rat genome occurring during or just after zona penetration but before ooplasm penetration. PMID- 10533709 TI - Nicotinamide alters the calcium release pattern and the degradation of MPF activity after fertilisation in ascidian oocytes. AB - Fertilisation in ascidian oocytes triggers a plasma membrane current, the release of intracellular calcium and the degradation of Maturation Promoting Factor (MPF) activity leading to the completion of meiosis and the initiation of embryo development. We have previously shown that the fertilisation current in ascidians is produced through the metabolism of nicotinamide nucleotide (NN) metabolites to ADP ribose. In this study we have used nicotinamide to test whether NN metabolism plays additional roles in fertilisation in ascidians. Nicotinamide treatment blocked calcium-induced calcium release (CICR) and arrested the cell cycle prior to the completion of meiosis I. Nicotinamide further prevented the abolition of MPF activity after fertilisation. Interestingly, nicotinamide treatment caused ascidian oocytes to form interphase-like pronuclei after fertilisation, despite the high MPF activity. The data demonstrate that NN metabolism is involved in calcium signalling through CICR and further suggest that a NN metabolite acts as a messenger connecting MPF activity to the formation of the meiotic apparatus. PMID- 10533710 TI - Ultrastructural changes during nuclear maturation in Bufo arenarum oocytes. AB - During progesterone-induced nuclear maturation the oocytes of Bufo arenarum undergo a series of nuclear and cytoplasmic changes. The breakdown of heterocellular communications between the follicular cell projections and the oocyte microvilli, and the consequent enlargement of the perivitelline space, were observed at the animal pole. The more evident cytoplasmic feature during nuclear maturation comprised the gathering of glycogen granules in clusters, some phagocytosed by empty vesicles. With respect to the location of these vesicles, some were observed in close proximity to the oolemma and others were freely suspended in the perivitelline space, extruded from the oocyte. Other visible events were the disruption of the annulate lamellae, the formation of an elaborate cortical endoplasmic reticulum and the rearrangement of the cortical granules in a monolayer immediately beneath the oolemma together with aggregates of endoplasmic reticulum cisternae. Our results show that during nuclear maturation the nuclear oocyte changes include a flattening of the spherical oocyte nucleus, its migration towards the surface of the animal pole, the disappearance of the nucleoli and the dissolution of the nuclear envelope. PMID- 10533711 TI - Fungal morphology in submerged cultures and its relation to glucose oxidase excretion by recombinant Aspergillus niger. AB - The effect of culture conditions such as medium composition and shear stress on the fungal pellet morphology in shake-flask cultures and its relation to glucose oxidase (GOD) excretion by recombinant Aspergillus niger NRRL 3 (GOD 3-18) was investigated. It was shown that culture conditions resulting in the formation of smaller fungal pellets with an increased mycelial density result in higher yields of exocellular GOD. The pellets obtained in shake-flask cultures showed distinct layers of mycelial density with only the thin outer layer consisting of a dense mycelial network. The performance of the recombinant strain and the process of pellet formation was also analyzed during batch cultivation in a stirred-tank bioreactor. It was shown that the process of pellet formation occurred in two steps: (1) aggregation of free spores to spore clusters with subsequent germination and formation of small aggregates surrounded by a loose hyphal network, and (2) aggregation of the primary aggregates to the final full-size pellets. The fungal pellets formed during bioreactor cultivation were smaller, did not show large differences in mycelial density, and were more efficient with respect to the production of exocellular GOD. The decreasing pellet size also correlated with an increased mycelial density, indicating an improvement of the transport of nutrients to the inner parts of the pellet. PMID- 10533712 TI - Characterization of invertase entrapped into calcium alginate beads. AB - A solution of 10 g/L of sodium alginate (Satialgine types used [Sanofi trademark]: SG800 and S1100 with manuronic/guluronic ratio of 0.5 and 1.2, respectively) containing invertase (0.08 g of protein/L) was dropped into 0.1 M CaCl2 solution buffered at pH 4.0, 7.0, or 8.0. The beads were left to harden in CaCl2 solution for 24 h. The high immobilization yield of 60% occurred with SG800 at pH 8.0. The activity of soluble and insoluble invertase was measured against pH (2.8-8.0), sucrose concentration (4.5-45 mM), and temperature (30-60 degrees C). Both forms presented an optimum pH of 4.6. However, the soluble invertase was stable at the overall pH interval studied, whereas insoluble invertase lost 30% of its original activity at pH > 5.0. At temperatures above 40 degrees C, the insoluble form was more stable than the soluble one. The kinetic constants and activation energies (Ea) for free invertase were KM = 41.2 mM, Vmax = 0.10 mg of TRS/(min.mL), and Ea 28 kJ/mol for entrapped invertase they were (KM)ap = 7.2 mM, (Vmax)ap = 0.060 mg of TRS/(min.mL), and (Ea)ap = 24 kJ/mol. PMID- 10533713 TI - Recent developments in microbial inulinases. Its production, properties, and industrial applications. AB - Microbial inulinases are an important class of industrial enzymes that have gained much attention recently. Inulinases can be produced by a host of microorganisms, including fungi, yeast, and bacteria. Among them, however, Aspergillus sp. (filamentous fungus) and Kluyveromyces sp. (diploid yeast) are apparently the preferred choices for commercial applications. Among various substrates (carbon source) employed for their production, inulin-containing plant materials offer advantages in comparison to pure substrates. Although submerged fermentation has been universally used as the technique of fermentation, attempts are being made to develop solid-state fermentation technology also. Inulinases catalyze the hydrolysis of inulin to D-fructose (fructose syrup), which has gained an important place in human diets today. In addition, inulinases are finding other newer applications. This article reviews more recent developments, especially those made in the past decade, on microbial inulinases--its production using various microorganisms and substrates. It also describes the characteristics of various forms of inulinases produced as well as their applications. PMID- 10533714 TI - Leaching kinetics of water soluble organic carbon (WSOC) from upland soil. AB - The kinetics of the leaching process of Water Soluble Organic Carbon (WSOC) from an upland soil were studied in a column leaching experiment. The pattern of the leaching curve indicated multiphasic release kinetics. A conceptual model was proposed to divide capillary and gravitational pores. The translocation of organic carbon in soil during leaching occurs in four sequential steps. 1) degradation of insoluble organic carbon to form WSOC; 2) desorption into capillary pore water; 3) diffusion towards gravitation pore water; and 4) leaching by convection flow. The overall kinetics of the leaching process can be depicted using an equation with three additive terms for degradation, desorption/diffusion, and convection, respectively. PMID- 10533715 TI - Occurrence and behavior of wastewater indicators in the Santa Ana River and the underlying aquifers. AB - The occurrence and behavior of wastewater indicator compounds in the Santa Ana River (SAR) water and the underlying aquifer recharged by the SAR has been studied. The SAR contains a high proportion of tertiary treated wastewater effluents, up to 100% during summer and fall. The following water quality parameters were quantified: four specific wastewater indicator compounds, ethylene diaminetetraacetic acid (EDTA), nitrilotriacetic acid (NTA), a naphthalene dicarboxylate (NDC) isomer, alkylphenol polyethoxy carboxylates (APECs), and selected haloacetic acids (HAAs), nitrate, dissolved oxygen (DO), DOC, total carbohydrate, and phenolic substances. Statistical analysis indicated that normal distribution was adequate to describe the probability distribution of the constituents in most cases. In the river, the concentrations of wastewater indicator compounds decreased as the fraction of storm runoff increased. EDTA and NDC were detected in a monitoring well near the river and in two production wells 1.8 and 2.7 km down gradient with little apparent attenuation. By contrast, NTA, APECs, bromochloro- and dibromoacetic acids appeared to be attenuated significantly during infiltration of river water and groundwater transport. PMID- 10533716 TI - Synergistic effects of combination of photolysis and ozonation on destruction of chlorophenols in water. AB - Synergistic effects including TOC elimination, ozone consumption and microtoxicity reduction for combination of photolysis and ozonation compared to those of direct photolysis and ozonation alone on destruction of chlorophenols including 2-chlorophenol, 4-chlorophenol and 2,4-dichlorophenol were studied. It was found that the synergistic effects of combination of photolysis and ozonation increased obviously with increasing initial pH of solution to basic pH levels. Results showed that the synergistic effects of photolytic ozonation under the conditions imposed was notable with mineralization rate enlarging more than 100%, oxidation index (OI) decreasing 50%, and microtoxicity being reduced by 30%, indicating that the potentialities of photolytic ozonation compared to direct photolysis and ozonation alone was remarkable for treatment of industrial wastewater containing chlorophenols. PMID- 10533717 TI - Toxic effect of surfactants and probable products of their biodegradation on methanogenesis in an anaerobic microbial community. AB - Surfactants used in household and various industries, are rather toxic; therefore, the accumulation of these compounds in the environment through wastewaters has challenged the problem of their biodegradation. In this research, an attempt was made to assess the toxic effect of various surfactants and the likely products of their biodegradation on the acetoclastic methanogens of an anaerobic microbial community. Among the substances investigated, cationic surfactants were found to be most toxic to methanogens: 154 mg/l alkamon DS and 345 mg/l catamin AB induced a 50% inhibition of methanogenesis. Toxicity studies of some aromatic and cyclic compounds, as the probable products of biodegradation of alkylbenzene sulfonate surfactants, showed that methanogenesis in the microbial community under study are rather tolerant to high concentrations of these compounds. PMID- 10533718 TI - A laboratory study of the mineralization and binding of 14C-labeled herbicide rimsulfuron in a rendzina soil. AB - The fate of pyrimidine-2-14C-rimsulfuron in a rendzina soil was investigated using a laboratory microcosm approach. Measurement of CO2 evolution suggested that rimsulfuron applied at 5 times the recommended dose did not affect soil respiration. Under abiotic conditions, no mineralization of 14C-rimsulfuron into 14C-CO2 occurred and under biotic ones it was very low reaching 0.75% of the applied 14C-rimsulfuron after 246 days of incubation. The analysis of data showed that a three-half order model provided the best fit for the mineralization curve. Extractable 14C-residues decreased over time to 70-80% of the applied 14C rimsulfuron at the end of the incubation. After 246 days of incubation, non extractable residues (NER) accounted for up to 24.7% of the applied 14C rimsulfuron and were distributed according to organic carbon in soil size fractions, suggesting a progressive incorporation process of NER to soil humus. PMID- 10533719 TI - 17 beta-estradiol: behavior during waste water analyses. AB - The distribution between the aqueous and solid phase of 17 beta-estradiol in the ng-range in waste water was investigated by spiking experiments with a radio labeled hormone. The distribution was measured by liquid scintillation counting. The major part of 17 beta-estradiol remained in the aqueous phase and did not adsorb to the solids. The fraction in the aqueous phase could be enriched by a conventional solid phase extraction with a recovery rate of up to 100%. Losses were negligible and the compound did not adsorb to the surfaces of glass bottles. PMID- 10533720 TI - Biodegradation kinetics of surfactants in seawater. AB - In this paper, a general kinetic model for degradation processes of surfactants is proposed. The model equation is v = K2S2 + K1S + K0, where v is the substrate consumption rate in the biodegradation process, S is the surfactant concentration in the medium and K2, K1, and K0 are kinetic constants. From this general expression, different simplified equations can be obtained (where K0 = 0; K2 and K0 = 0; K2 = 0; K2 and K1 = 0), which are representative of the process for different operating conditions. This model was tested by measuring the degradation of two different surfactants (Sodium dodecyl benzene sulfonate, LAS; and Sodium dodecyl sulfate, DSNa) under two different temperatures (5 and 20 degrees C). Values predicted by the model are close to experimental data obtained. PMID- 10533721 TI - Adenovirus-mediated wild-type p53 expression induces apoptosis and suppresses tumorigenesis of experimental intracranial human malignant glioma. AB - Adenoviral-mediated gene transfer for the treatment of experimental intrinsic malignant brain neoplasms holds promise. The role, however, of intracellular, adenoviral-mediated p53 expression to inhibit growth of experimental human intracranial malignant gliomas remains largely unexplored. Using the AdCMV.p53 vector we measured the in vitro expression of p53 and the resultant effect upon U251 human malignant glioma cellular proliferation. We further measured the survival of nude mice after intracranial injection of the infected vs. control U251 cells. The growth of the infected U251 cells was inhibited when compared to both the uninfected cells and cells infected with the control vector (AdCMV.Null). Agarose gel electrophoresis confirmed the AdCMV.p53-dependent cellular apoptosis. Nude mice having intracranial injections of the U251 cells infected with the control (AdCMV.Null) vector showed diminished survival. In contrast, mice having intracranial injections of the cells infected with the AdCMV.p53 vector showed 100% survivorship measured 100 days after treatment. Gene therapy via the AdCMV.p53 viral vector holds promise for the clinical treatment of human malignant gliomas. PMID- 10533722 TI - Thalidomide and a thalidomide analogue inhibit endothelial cell proliferation in vitro. AB - Angiogenesis is a crucial process in inflammatory reactions as well as in tumor implantation and growth. Tumors with high rates of invasion and recurrence such as gliomas, are specially dependent on neovascularization. This suggests that inhibition of angiogenesis might reduce the growth of these tumors. Thalidomide has been previously shown to inhibit angiogenesis induced by basic fibroblast growth factor in vivo, using the rabbit corneal micropocket assay. Therefore, the effect of thalidomide and a thalidomide analogue (cc-1069) on the proliferation in vitro of endothelial and glioma cells was tested. We observed a decrease in endothelial cell proliferation in cultures treated with thalidomide or the thalidomide analogue cc-1069. The analogue inhibited endothelial cell proliferation more efficiently than thalidomide. The inhibition occurred in association with a marked decrease in the activity of the nuclear factor SP1 and a moderate inhibition of NF-kappaB activation in nuclear extracts of endothelial cells. The drugs did not impair cell viability. There was no effect of thalidomide or the thalidomide analogue on the proliferation of the glioma cell line (U251) in vitro. PMID- 10533723 TI - Soluble factors, including TNF alpha, secreted by human T cells are both cytotoxic and cytostatic for medulloblastoma cells. AB - We studied the effect of the treatment of a medulloblastoma cell line by human T cells derived soluble factors. Medulloblastoma is one of the more common aggressive solid neoplasms in children for which there is no adequate therapy. Cell lines established from such tumours may be helpful to test the effect of various molecules on cell proliferation. Previous studies have suggested that T cell-derived factors may be toxic for the medulloblastoma cell line Dev. Cytokines were thought to mediate this effect. In this paper, we described changes in morphology, survival and cell cycle induced in Dev cells cocultured with human T cell lines chronically infected with a retrovirus (HTLV-I) and known to secrete high level of cytokines TNF alpha, IL1alpha and IL6. Such cocultures resulted in the death of a part of Dev cells and in decreased proliferation of surviving cells, associated with morphological changes and increase in vimentin expression. Treatment with conditioned medium from infected Dev cells, containing virus induced cytokines, triggered the same effect. Reduction of these effects by TNF alpha deprivation of conditioned medium suggested that this cytokine may be implicated. Direct treatment of Dev cells with recombinant cytokines indicated that TNF alpha, but not IL1 or IL6, is associated with Dev cell alterations. TNF alpha was shown to induce the death of Dev cells by an apoptotic pathway. Furthermore, TNF alpha had a bimodal effect on the cell cycle of surviving Dev cells. These differential effects of such cytokines on medulloblastoma cells could be therefore of interest for immunotherapy of these tumours. PMID- 10533725 TI - Correlation of clinical features and telomerase activity in human gliomas. AB - Telomerase is a ribonucleoprotein containing an RNA template that synthesizes telomeric DNA. The expression of telomerase activity is concomitant with the attainment of immortality in tumor tissues and cells. In this report, we analyzed telomerase activity in 39 human gliomas with different histological, and in 10 meningiomas, 3 neurinomas, and 2 normal brain tissues by using a polymerase chain reaction (PCR)-based telomeric repeat amplification protocol (TRAP) assay. Telomerase activity was detectable in almost all of the gliomas (36 of 39), but not in any of the meningiomas, neurinomas, or normal brain tissues. In addition, we also analyzed the level of telomerase activity in the 36 gliomas with positive telomerase activity. The relative telomerase activity of the glioma showed a clear association with the pathological grade of glioma; i.e., most of the tumors with high telomerase activity were pathologically of high grade. And also the relative level of telomerase activity could be correlated with the survival time of the patients. These results suggest that the level of telomerase activity in brain tumors is a diagnostic marker indicating the prognosis of the patient as well as the malignant potential of the tumor. PMID- 10533724 TI - Transforming growth factor-alpha antisense vectors can inhibit glioma cell growth. AB - The effects of transforming growth factor-alpha (TGF-alpha) on cell growth were studied in human glioma U251 cells transfected with antisense TGF-alpha vectors (pcDNAI.neo). Several antisense clones showed a marked decrease in growth rate in serum-free medium but not in medium containing 10% FBS, compared with those of parental cells and clones from sense or vector transfectants. Antisense clones also produced fewer and smaller colonies in anchorage-independent growth assays. Moreover, there was a reduction in TGF-alpha expression in these antisense clones at both the protein and mRNA levels, as determined by enzyme linked immuno sorbent assay and reverse transcriptase polymerase chain reaction analysis. A U251 clone transfected by TGF-alpha antisense in a different vector (pMT/Ep) also showed a marked suppression in cell growth and TGF-alpha mRNA level. Finally, transfected clones with either vector system, showed decreased tumorigenicity in nude mice. In summary, a strong correlation between the inhibition of glioma cell growth and TGF-alpha expression was obtained from two different plasmid vectors, indicating that the expression of TGF-alpha could be specifically and effectively down-regulated by TGF-alpha antisense vector, which in turn led to growth inhibition. These studies suggests that TGF-alpha plays an essential role in controlling human glioma cell proliferation and may serve as a potential target for treatment of malignant glioma. PMID- 10533726 TI - Selective transvascular delivery of oligodeoxynucleotides to experimental brain tumors. AB - Optimal therapeutic strategy for malignant brain tumors is controversial. Recent studies of viral or nonviral gene therapy in rats emphasize the need for a selective delivery system. We examined whether phosphorothioate oligodeoxynucleotides (lacZ 2157, 5'-GTGGCGTCTGGCGGAAAACC-3') could be selectively delivered transvascularly into experimental brain tumors following intracarotid infusion of bradykinin, a specific blood-tumor barrier opener. The specificity of 32P-labeled complementary antisense lacZ 2157 and the stability of lacZ 2157 in vivo were confirmed using slot-blotting hybridization method and polyacrylamide gel electrophoresis. Concentrations of lacZ 2157 after intracarotid injection (2 mg/kg, 10 microg/kg/min) with or without bradykinin were determined in the brain, tumor tissue, liver, kidney, and plasma. The transfer ratio of lacZ 2157 from the plasma to the tissues was calculated and expressed as tissue content relative to plasma content of lacZ 2157 per mg tissue (Do, microl/mg). Delivery of lacZ 2157 to tumor tissue increased 3.24 times with bradykinin over delivery in controls (0.0243 +/- 0.0176 vs. 0.00750 +/- 0.00389; p < 0.05). Delivery of lacZ 2157 to ipsilateral and contralateral cerebral cortex to the tumor, and delivery to the contralateral basal ganglia, did not increase significantly with bradykinin. These results indicate that such transvascular delivery with bradykinin can deliver a relatively large amount of oligodeoxynucleotide selectively to brain tumors without affecting normal brain. PMID- 10533727 TI - A new xenograft model of primary central nervous system lymphoma. AB - The management of primary lymphoma of the central nervous system (PCNSL) remains controversial and patients' outcome dismal. In order to investigate new selective therapeutic strategies in a controlled system, a reproducible model of PCNSL in nude rats was developed and characterized. Human B lymphoma cells (BL2) were implanted in the brain frontal area in New Zealand nude rats through a silastic device sealed to the skull. Fifteen and 30 days post-implantation, animals were sacrificed. An autopsy was performed. Representative brain sections were cut and examined for the presence of lymphoma. Immunohistochemistry was performed for proliferation (MIB1-Ki67), a B-cell marker (L26-CD20), a T-cell marker (UCHL1 CD45RO). The analysis of the brains showed tumor growth in 88% of the rats. No mortality was observed. At autopsy no extracerebral, spinal or cerebellar metastasis were found. Microscopically the brain tumors appeared non encapsulated, highly vascularized, with a characteristic perivascular and diffuse lymphomatous spread in the parenchyma. Immunohistochemistry showed a marked positivity of the tumor cells for L26. Tumor cells were negative for UCHL1. Mean proliferation rate was 30%. The device was well tolerated and caused no local infection. Controlled studies on PCNSL in animal models are lacking. This PCNSL model in nude rats reproduces the histology and location of human CNS lymphoma. Tumor dimensions are within the resolution limits of CT and MRI and therefore suitable for stereotactic therapy. This model provides a tool to test new chemo and radiotherapeutical strategies in a controlled fashion. PMID- 10533728 TI - Treatment of newly diagnosed glioblastoma multiforme with carmustine, cisplatin and etoposide followed by radiotherapy. A phase II study. AB - A meta-analysis and several studies of patients with grade III and IV gliomas have indicated that the addition of nitrosurea based chemotherapy to surgery and radiation may improve survival. We performed a phase II study of pre-irradiative chemotherapy with BCNU, cisplatin and etoposide. This implies a short total treatment duration and a reliable response evaluation. The treatment schedule was three cycles of BCNU 200 mg/m2 i.v. on day 1, cisplatin 20 mg/m2 i.v. on day 1-5 and etoposide (VP-16) 100 mg/m2 i.v. on day 1-5, given every five weeks and followed by localized radiation, 60 Gy in 30 fractions. Twenty-nine patients with newly diagnosed glioblastoma multiforme (GBM), mean age 50 (27-66) and performance status (PS) 0-2 were included. Using the Macdonald criteria 33% had partial remission (PR), 41% stable disease (SD) and 26% progressive disease (PD) after chemotherapy. After additional radiation 44% had PR, 37% SD and 19% PD. Non hematological toxicity and leukopenia was mild, but thrombocytopenia (TP) frequent. Grade III and IV TP occurred in 25% and 57% respectively, and grade III bleeding in 45%. No severe or fatal complications was seen. Median time to progression (TTP) was 7.6 months (6.0-9.1) and median survival was 11.4 months (10.1-12.7). We conclude that this regimen is effective and feasible in patients with GBM. The short course pre-irradiatory chemotherapy may be less cumbersome than adjuvant chemotherapy and the regimen may be even more active in grade III gliomas. PMID- 10533729 TI - Volumetric reduction of a choroid plexus carcinoma using preoperative chemotherapy. AB - We report for the first time a measured volumetric reduction of a choroid plexus carcinoma utilizing preoperative chemotherapy. Histologically proven choroid plexus carcinoma was diagnosed in a fifteen month old female. She was treated with three courses of chemotherapy including etoposide (VP16), cyclophosphamide, vincristine, and cisplatin. Computer-assisted three dimensional reconstruction of the tumor volume was performed prior to and after three courses of chemotherapy. An overall reduction of 29.5% of tumor volume was accomplished preoperatively. Staged surgical procedures resulted in a complete resection of her lesion and she has remained disease-free for 31 months. A volumetric measurement as a response to preoperative chemotherapy may prove valuable in determining future optimal treatment regimens for choroid plexus carcinoma of childhood. PMID- 10533730 TI - Management of cerebral metastases from malignant melanoma: results of a combined, simultaneous treatment with fotemustine and irradiation. AB - We report results of a conservative treatment for brain metastases from malignant melanoma with a combination of irradiation and chemotherapy (fotemustine and/or DTIC). To date, 12 patients have been treated. There was a complete remission of the brain metastases in four patients. In two patients a partial remission was observed. The mean survival of the responder was 8.2 months (95% confidence interval 3.8-12.6 months). The most common side effects were thrombocytopenia, leukopenia, and alopecia. Altogether, the treatment was well tolerated. As the outcome of patients with brain metastases from malignant melanoma is generally poor, this combined chemo- and radiation therapy may provide improved care for such patients. PMID- 10533733 TI - The histological basis of detection of adenoma and cancer in the colon by autofluorescence endoscopic imaging. AB - BACKGROUND AND STUDY AIMS: The reason for the difference in fluorescence between normal and diseased tissues (carcinoma and adenoma) in the colon observed on autofluorescence endoscopy is unclear, flavins, NADPH and collagen being regarded as possible major sources of fluorescence. The purpose of this study was to identify the reason for this difference in fluorescence. PATIENTS AND METHODS: Samples of human colonic tissues (adenoma: n = 6, cancer: n = 11, normal: n = 11) were obtained from resected specimens. The flavin content of human colonic tissue was measured by high performance liquid chromatography. Fluorescence microscopy under blue light excitation (400-440 nm) was performed using frozen sections of normal, adenomatous and cancerous tissues, and examining them for the presence and characteristics of fluorescence. RESULTS: The flavin content of normal and diseased tissue was not significantly different. Fluorescence microscopy of normal colonic tissue revealed strong fluorescence in the submucosal layer, which corresponded to collagen. Tissue fluorescence did not decrease in reducing agent or acid solution. No difference in fluorescence was detected in normal mucosa, adenoma or cancerous tissue on fluorescence microscopy. These findings indicate that flavins and NADPH do not affect tissue fluorescence, and that submucosal collagen is the main source of tissue fluorescence in the colon. CONCLUSION: The reason for the decreased fluorescence in diseased tissues appears to be a decrease in collagen fluorescence due to the screening effect of mucosal thickening or replacement of submucosa by cancer cells. PMID- 10533731 TI - Combined treatment modality for anaplastic oligodendroglioma: a phase II study. AB - PURPOSE: To investigate feasibility, toxicity and antitumor activity of combined surgery, postoperative radiation therapy (RT) and adjuvant chemotherapy (CHT) in adult patients with pure anaplastic oligodendroglioma (PAO) or mixed anaplastic oligoastrocytoma (MAO). METHODS: Between January 1988, and June 1993, 23 patients entered into a phase II study. After surgery, postoperative RT was administered with 60 Gy in 30 daily fractions in 30 treatment days in 6 weeks. Two weeks after RT, adjuvant 'modified' PCV (mPCV) (Procarbazine, 60 mg/m2, days 1-14; CCNU, 100 mg/m2, day 1; and vincristine, 1.4 mg/m2 (max. 2 mg), days 1 and 8) was administered every six weeks up to six cycles or until progression occurred. RESULTS: Median survival time is not attained yet, while 1-5 year survival rates are 100%, 100%, 78%, 61%, and 52%, respectively. Median time to tumor progression is not attained yet, while 1-5 year progression-free survival rates are 100%, 100%, 70%, 52%, and 52%, respectively. On univariate analysis of potential prognostic factors, sex, tumor location (frontal versus other), and histology (pure versus mixed anaplastic oligodendroglioma) were not found to influence survival. Age of < 50 years carried improved prognosis as well as Karnofsky performance status (KPS) 90-100 when compared to KPS of 70-80. Patients having tumors < or = 4 cm did better than those with tumors > 4 cm as well as those with total tumor resection when compared to those with subtotal tumor resection or biopsy only. Acute high-grade (> or = 3) CHT-related toxicity was mainly hematological with only 3 (13%) patients experiencing acute grade 4 toxicity. CONCLUSIONS: Combined treatment modality consisting of surgery, postoperative high-dose RT and mPCV chemotherapy for patients with anaplastic oligodendroglioma was effective with acceptable toxicity. Further studies are needed with more patients and longer follow-up to verify these results in this rare disease. PMID- 10533732 TI - Treatment of malignant gliomas in the elderly. AB - The benefit of standard treatment of malignant glioma in older patients is debated. In order to assess the effect of a combination of surgery, radiotherapy and chemotherapy on survival of elderly patients with high grade gliomas, 30 consecutive patients older than 70 years with malignant supratentorial gliomas were studied between 9/93 and 9/96. Median age was 73 years (70-79). The mean Karnofsky performance status (KPS) was 66 (30-100). Patients underwent maximum possible surgery, followed by a course of radiotherapy (45 Gy/25 fractions/5 weeks) with 3 or 4 orthogonal beams and a 2 cm margin around the tumor bed. The administration of chemotherapy was left at the discretion of the responsible physician and 12 patients received reduced dose nitrosourea-based chemotherapy. The overall median survival was 36 weeks. The median time to progression was 26 weeks. Three months after surgery, 26 patients were alive, 5 were in complete response, 2 in partial response and 10 were stabilized. Preradiotherapy KPS was the only significant prognostic factor with a median survival of 40 weeks in patients with KPS > or = 70 and 25 weeks when KPS was < 70 (logrank test, p = 0.05). In responding and stable patients (57% of the group) the median KPS was 68 and 66 at 1 and 3 months after the completion of radiotherapy. There was no case of radiotherapy-induced dementia with this regimen. Four out of 12 patients who received chemotherapy, experienced WHO grade 3/4 hematotoxicity. This study suggest that some patients older than 70 years with KPS > or = 70 may benefit from the treatment of malignant gliomas with surgery followed by reduced dose of limited field radiotherapy. Further studies are needed to define the most appropriate dose of radiotherapy and to evaluate further the risk/benefit ratio of a reduced dose chemotherapy in this population. PMID- 10533735 TI - Quality assurance and colonoscopy. AB - BACKGROUND AND STUDY AIMS: Little is known concerning the usefulness and feasibility of quality assurance programs in gastrointestinal departments. The aim of this study was to identify the indicators of quality in colonoscopy, to check their use in clinical practice, and to identify their threshold values. MATERIALS AND METHODS: A prospective study was performed in four endoscopic units. In the first phase, a questionnaire was used to identify the indicators that were considered important and easy to record; in the second phase, the selected items were prospectively recorded. RESULTS: Data from 603 colonoscopies were evaluated. The selected indicators were: rate of cecal intubation, rate of examinations with normal findings, rates of complications, appropriateness of indications, use of a washing machine for disinfection, duration of the disinfection procedure, rate of procedures repeated due to poor colon cleansing, rate of operative procedures, length of waiting time, rate of procedures performed for follow-up of known disease, experience of the operator, and rate of procedures performed with the patient under conscious sedation. A striking difference emerged between the technical standards at three centers, which were fairly good, and the standard at the fourth center, which was less satisfactory. The length of the waiting time was high in all centers, as well as the rate of examinations conducted with an inappropriate indication. The rate of procedures performed under conscious sedation varied widely between the centers. CONCLUSIONS: The study of the indicators of quality of colonoscopy is feasible and easy to perform in clinical practice, and can be useful for quality assurance programs. PMID- 10533734 TI - Esophageal achalasia: intrasphincteric injection of botulinum toxin A versus balloon dilation. AB - BACKGROUND AND STUDY AIMS: In patients with esophageal achalasia, pneumatic dilation is the treatment of choice, but it bears the risk of perforation in about 4% of cases. A new nonsurgical method, intrasphincteric injection of botulinum toxin A, has shown promising initial results, and we thus undertook this study to compare the long-term outcome of these two methods. PATIENTS AND METHODS: In a prospective randomized study, 24 patients with definitive esophageal achalasia were divided into two equal groups and underwent either balloon dilation or injection of botulinum toxin (20 U injected into each of the four quadrants in the lower esophageal sphincter). The outcome was assessed on the basis of a symptom score documented before treatment and at regular intervals for two and a half years thereafter. Complications associated with the two techniques were also documented. RESULTS: No relevant complication occurred in either of the treatment groups. Initially, dilation was successful in 10 of 12 patients (83%), and botulinum toxin injection in 9 of 12 patients (75%). The symptom scores showed no significant differences between the two groups before and one month after treatment. Over the two and a half year follow-up, however, all nine successfully treated patients in the botulinum toxin group experienced recurrence of symptoms, but only four of the ten patients (40%) in the dilation group. CONCLUSIONS: The two treatment methods initially had equal success rates, but in the long term the effect of the botulinum toxin injection was statistically significantly shorter than that of balloon dilation. As fewer risks are associated with the injection treatment, studies should be undertaken either to identify patient subgroups in whom botulinum toxin can be effective long-term or to test substances with longer-lasting effects. PMID- 10533737 TI - Distant lymph node metastases in esophageal cancer: impact of endoscopic ultrasound-guided biopsy. AB - BACKGROUND AND STUDY AIMS: The aim of this retrospective study was to evaluate the impact of endoscopic ultrasound (EUS)-guided biopsy in patients with esophageal carcinoma where distant lymph nodes which were possibly metastatic were visualized using EUS. PATIENTS AND METHODS: Out of 198 patients (150 men, mean age 66 years) examined over a 4-year period by EUS for local staging of esophageal cancer (121 squamous cell carcinomas and 77 adenocarcinomas), there was EUS visualization of distant lymph nodes in 40 (20%). EUS-guided biopsy was carried out in the latter patients, of cervical nodes with mediastinal tumors (n = 19), of celiac nodes with cervical tumors (n = 2) or superior mediastinal tumors (n = 9), and upper mediastinal lymph nodes in the case of distal adenocarcinomas (n = 10). RESULTS: On EUS-guided biopsy, results were positive in 31 patients, eight were correctly negative (as confirmed by surgery), and in one patient there was a technical failure, with positive findings on subsequent surgery. The sensitivity and specificity of the diagnosis of malignant lymph nodes were therefore 97% and 100% respectively. The positive results of EUS guided biopsy modified the tumor staging in 31 of these cases (77.5%), proving distant lymph node metastasis which is classified as stage M1. With regard to actual clinical management, surgery was withheld from 24 patients (60% of 40 cases) who were then treated with concomitant radiotherapy and chemotherapy. CONCLUSION: EUS-guided biopsy of distant lymph nodes was indicated in 20% of patients with esophageal cancers, and the biopsy results led to upgrading of the tumor stage in about 80% of cases and influenced the treatment decision in about 60%. PMID- 10533736 TI - Endoscope disinfection using acidic electrolytic water. AB - BACKGROUND AND STUDY AIMS: The aim of the present study was to evaluate a new endoscope disinfector (WM-1) that uses acidic electrolytic water (AEW). MATERIALS AND METHODS: AEW was produced by electrolysis of a 0.05% NaCl-water mixture, with a redox potential greater than 1000 mV and a pH lower than 2.7. In the first study, an endoscope artificially contaminated with 15 species of bacteria and four strains of viruses was treated using the WM-1. In the second study, endoscopic contamination after clinical use was examined by culture for Helicobacter pylori and other bacteria, and by polymerase chain reaction for the H. pylori urease gene and hepatitis C virus. The extent of contamination was then examined after exposing the WM-1 to AEW. The safety of AEW was examined using both in vivo and in vitro studies. RESULTS: All of the bacteria and viruses were destroyed or inactivated after the instrument had been exposed to AEW. Clinical contamination was detected from the instrument in 19 of 30 endoscopic procedures, whereas no bacteria or viruses were detected after five minutes' exposure to AEW. AEW was found to be nonirritant, nontoxic to cells, and nonmutagenic. CONCLUSION: The WM-1 successfully and safely disinfected the endoscopes. With running costs of yen 24 per day ($0.21 per day), the WM-1 provides an effective and inexpensive alternative to conventional disinfection equipment. PMID- 10533738 TI - Surface-rendering imaging of gastrointestinal lesions by three-dimensional endoscopic ultrasonography. AB - BACKGROUND AND STUDY AIMS: In three-dimensional endoscopic ultrasonography (3D EUS), a surface-rendering method can provide both a surface image and a cross sectional ultrasonographic image. We evaluated the usefulness of this imaging method for digestive tract lesions. PATIENTS AND METHODS: A total of 30 patients underwent 3D-EUS with surface-rendering using a 3D probe system which arranged individual radial scanning images into 128 points on a computer monitor to outline the surface of a lesion. A complete surface image of the lesion was displayed on the computer monitor using lines obtained from 40 radial scanning images. RESULTS: Surface-rendering images of lesions were similar to endoscopic images. The surface-rendering method permitted precise correlation of two dimensional images depicting a slice of a lesion with the corresponding surface of the lesion. Unlike conventional endoscopy, this approach permitted observations of lesions at any desired angle. Complete images of lesions were achieved in 14 patients and were half-completed in another five, but could not be obtained in 11 patients: in six the distance between the lesion and the 3D probe was too short to avoid artifacts, while in five the lesion was larger than the longitudinal scanning length of 4 cm or greater than 90 in extent in radial scanning images. Artifacts caused by heartbeat led to irregular images in four patients, including three with esophageal cancer and one with gastric cancer. CONCLUSIONS: Despite some problems, surface-rendering imaging should prove useful for diagnosis, and the method will improve as software is perfected. PMID- 10533739 TI - Small-bowel perforations related to endoscopic retrograde cholangiopancreatography (ERCP) in patients with Billroth II gastrectomy. AB - BACKGROUND AND STUDY AIMS: Endoscopic retrograde cholangiopancreatography (ERCP) is one of the mainstays in the diagnosis and treatment of hepatobiliary and pancreatic diseases, and is also increasingly used for patients with previous Billroth II gastrectomy. The aim of this study was to review our experience of ERCP in patients with Billroth II gastrectomy, and the complications associated with this procedure. PATIENTS AND METHODS: The records of 110 patients with Billroth II gastrectomy, treated between January 1993 and December 1997, were received retrospectively. Details noted included indications for ERCP, therapeutic interventions, causes of failure, and complications. RESULTS: A total of 110 patients underwent ERCP; the total number of ERCP attempts was 185. The major indications for ERCP were cholangitis (31%), common bile duct stones (22%), and jaundice (15%). The endoscope was successfully passed up to the papilla in 134 examinations (71%), and selective cannulation was successful in 122 of these (66%). There were 63 (34%) failed ERCP attempts. Causes of failure were: difficulty in entering the afferent loop (n = 19, 10%), failure to enter the duodenum (n = 23, 12%), endoscope-related bowel perforation (n = 9, 5%), and failed cannulation (n = 10, 6%). The other two failures were caused by desaturation in the patient, and inability to distend the duodenum. The major complication of the procedure was perforation, which occurred in 11 examinations (6%). Of these perforations, nine occurred in the small bowel while the endoscope was being manipulated through the afferent loop; these patients required laparotomy. Two patients had retroduodenal perforations, one occurring after sphincterotomy and one after cannulation. Both patients were successfully managed conservatively. Three patients suffered bleeding after sphincterotomy (3/185 procedures, 1.6%), and one patient developed acute pancreatitis. These were managed conservatively. The overall complication rate was 8%. There were two deaths among the patients with small-bowel perforations, and consequently an overall mortality rate of 1% (2/185 procedures). CONCLUSIONS: Most complications of ERCP in patients with previous Billroth II gastrectomy were caused by bowel perforation while the endoscope was being manipulated through the afferent limb. Such perforations are intraperitoneal and require surgical intervention. PMID- 10533740 TI - Improved technique for performing endoscopic ultrasound guided fine needle aspiration of lymph nodes. AB - BACKGROUND AND STUDY AIMS: Trans-esophageal real-time endoscopic ultrasound (EUS) guided fine needle aspiration (FNA) has emerged as an important technique for sampling perigastrointestinal lymph nodes. The purpose of this study was to compare the yield of EUS-guided FNA of mediastinal lymph nodes using different techniques. MATERIALS AND METHODS: A 2 cm mediastinal lymph node was dissected at autopsy. FNA was performed on this lymph node with a 21 gauge needle which is used clinically for EUS-guided FNA (GIP-Mediglobe). FNA of the lymph node was performed for 60 sec, while continuous or intermittent suction was applied with a 10 ml, 20 ml and 30 ml syringe. The pathologist was blinded to the technique used for FNA of the lymph node. The slides were examined and the results recorded independently by two pathologists who were blinded to each other's findings. A similar procedure was repeated in a 2 cm lymph node removed during another autopsy. RESULTS: Pathologic examination revealed metastatic transitional cell bladder carcinoma in the first lymph node, and metastatic non-small cell lung carcinoma in the second lymph node. The cellularity and quality of FNA performed with the 10 ml syringe was better than with the 20 ml or 30 ml syringe. With the 10 ml syringe, continuous suction for one minute provided a better sample than intermittent suction. FNA with a 20 ml or 30 ml syringe was more cumbersome, as it required more physical force. CONCLUSIONS: Our study reveals that continuous rather than intermittent suction with smaller syringes (5-10 ml) provides optimal cellularity in EUS-guided FNA of mediastinal lymph nodes and that use of larger (20-30 ml) syringes does not improve the rate of obtaining a diagnostic specimen. PMID- 10533741 TI - An alternative approach to the inaccessible intradiverticular papilla. AB - BACKGROUND: In patients with a narrow-necked juxtapapillary diverticulum the endoscopic cannulation of the papilla of Vater and the subsequent biliary therapy is sometimes impossible. PATIENTS AND METHODS: Three patients referred for endoscopic retrograde cholangiography and stone extraction were included. Earlier attempts to cannulate failed because visual control was impeded by narrow-necked juxtapapillary diverticula with the papilla located in the fundus. Endoscopic balloon dilation of the narrow diverticular neck, using a 15-mm stone retrieval balloon, was carried out. RESULTS: In all three cases the papillary orifice was readily brought into view after balloon dilation of the diverticular opening. Subsequent endoscopic treatment to the bile duct was successful without any complications. CONCLUSION: Balloon dilation of a narrow-necked juxtapapillary diverticulum is a safe and easy procedure, which facilitates both cannulation of the papilla and subsequent biliary endoscopic treatment. PMID- 10533742 TI - The role and future of endoscopic imaging systems. AB - Visual perception is the main sensory input from the environment in most situations of daily life. It is the only sensory input from the operating field in endoscopic surgery, and thus the qualities of the optical imaging system have a considerable impact on the course of the surgical intervention. Significant improvements have been made recently in various fields of science and engineering, influencing endoscopic imaging systems in experimental and clinical use. Among these are technologies that improve the endoscope itself in terms of providing new visual features, such as fogging prevention and plastic images, using new illumination techniques. Other developments concern the improvement of image resolution and color fidelity through new charge-coupled device (CCD) sensors or alternative techniques for image creation. Finally, the combination of endoscopic technologies with robotics provides for intuitive and more efficient direction of the line of sight. PMID- 10533743 TI - Gastric hyperplastic polyp occurring at the resection site of gastric adenoma. AB - Endoscopic mucosal resection was carried out in a 70-year-old man with a gastric adenoma. Endoscopy 1 year later revealed a subpedunculated polyp about 1 cm in diameter at the resection site. Pathological examination of the resected specimen showed hyperplasia of the regenerative epithelium. The mechanism of occurrence of hyperplastic polyp at the resection site is discussed. PMID- 10533744 TI - Corrosive-induced gastric outlet obstruction. PMID- 10533745 TI - Overexpression of p53 protein and colonic adenoma recurrence. PMID- 10533746 TI - Circumscribed papillomatosis of the common bile duct treated by pancreatoduodenectomy. PMID- 10533747 TI - An unusual case of endometriosis causing rectal bleeding. PMID- 10533749 TI - Early rebleeding after successful hemoclipping of a postpolypectomy rectal ulcer. PMID- 10533748 TI - Limited duodenal pneumatosis during needle-knife sphincterotomy. PMID- 10533750 TI - Rectal ultrasound in the diagnosis of localized colitis cystica profunda (mucosal prolapse-related disease). PMID- 10533751 TI - Asymptomatic right colon ischemia associated with colonic wall calcification: report of a case. PMID- 10533752 TI - Adenocarcinoma arising from a hyperplastic polyp with adenomatous foci. PMID- 10533753 TI - Ventriculoperitoneal shunt mimicking a colonic polyp. PMID- 10533754 TI - Intra-abdominal complications after cardiac surgery. AB - Gastrointestinal complications such as peptic ulcer disease, pancreatitis, acute cholecystitis, bowel ischaemia, and diverticulitis are rare after cardiac surgery (< 1%), but are associated with high morbidity and mortality (about 30%). Hypoperfusion during cardiopulmonary bypass seems a possible aetiological factor. As many patients may be mechanically ventilated and sedated, the usual symptoms and signs of an abdominal complication may be masked. It is necessary to keep this possibility in mind in patients with abdominal pain or tenderness, and the usual diagnostic measures should be undertaken if time permits. Initial treatment is usually conservative, but when it fails, prompt intervention is obligatory. Unfortunately surgeons are often reluctant to submit patients to major abdominal operations immediately after cardiac surgery. However, effective and timely intervention may be life-saving in patients who are poorly able to compensate for the major haemodynamic disturbances of the untreated serious bleeding or sepsis. Although the cardiac condition must be taken into consideration, most patients' cardiac function will have improved since their open-heart surgery and they should be able to withstand general anaesthesia and most operations. PMID- 10533755 TI - "Avoidable" deaths in two areas of Sweden - analysis of deaths in hospital after injury. AB - OBJECTIVE: To describe causes of death and other characteristics of "avoidable" deaths in patients admitted to hospital after trauma, and estimate and analyse changes in the avoidable death rate during the years studied. DESIGN: Retrospective analysis of medico legal autopsy material. SETTING: One northern and one western area in Sweden 1988-1996. SUBJECTS: 335 cases who died in hospital after trauma. MAIN OUTCOME MEASURES: Avoidable death, defined as an Injury Severity Score (ISS) of 35 or less and Abbreviated Injury Scale (AIS) head of 4 or less and cause of death. RESULTS: We found 70 avoidable deaths (21%). Among these, 15 (21%) died of head injuries, 17 (24%) of thoracic, abdominal, or pelvic injuries, and 38 (54%) of medical complications. The number of deaths after trauma decreased considerably from 1988-90 to 1994-96, but the proportion who died in hospital remained almost constant. The proportion of avoidable deaths decreased from 22% to 17%, mainly because the proportion of deaths from medical complications was halved. CONCLUSION: The standard of Swedish in-hospital trauma care has improved, particularly with a reduction in post-traumatic complications. However, there is still room for improvement in the treatment of complications among elderly people. PMID- 10533756 TI - Treatment with hyperbaric oxygen affects endothelial cell fibrinolysis. AB - OBJECTIVE: To investigate the effect of hyperbaric oxygen treatment (HBO) on the thrombolytic properties of endothelial cells. SETTING: University hospital, Sweden. INTERVENTIONS: Human endothelial cells were derived from saphenous veins, and exposed to oxygen in a compression chamber at 2.5 atmospheres absolute (ATA, =250kPa). Cells exposed to 2.5 ATA with a gas mixture similar to air (HB Air), and unpressurised air-exposed cells served as controls. MAIN OUTCOME MEASURES: Tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor type 1 (PAI-1). RESULTS: Immediately after treatment there was a significant increase in t-PA protein in the medium in cultures treated with HBO compared with HB Air (p = 0.015, n = 6), and untreated controls (p = 0.015, n = 6). The PAI-1 concentration in media was also higher in the HBO-treated group compared with HB Air (p = 0.004, n = 6) and untreated controls (p = 0.004, n = 6). Six hours after treatment there was still a significant increase in PAI-1 in the HBO-treated group compared with untreated controls (p = 0.007, n = 6), but not with the pressure control. t-PA concentrations were similar. Specific mRNA for t-PA and PAI-1 was detectable immediately after treatment and six hours later in all experimental groups as assessed by reverse transcriptase polymerase chain reaction (RT-PCR). HBO increased the gene expression for both t-PA and PAI-1. CONCLUSIONS: HBO affects endothelial cell function and its fibrinolytic response. These findings may have clinical relevance in hyperbaric medicine and trauma care. PMID- 10533758 TI - Complications after bilateral adrenalectomy for phaeochromocytoma in multiple endocrine neoplasia type 2--a plea to conserve adrenal function. AB - OBJECTIVE: To evaluate the complications of the adrenocortical supplementation that is needed after bilateral adrenalectomy for phaeochromocytoma in patients with multiple endocrine neoplasia (MEN) type 2 syndrome. DESIGN: Retrospective study. SETTING: University hospital, The Netherlands. MATERIALS: 28 patients with MEN 2 who underwent total adrenalectomy for phaeochromocytoma between 1972 and 1996. MAIN OUTCOME MEASURES: Perioperative morbidity and mortality, histopathological findings, complications of adrenocortical supplementation therapy. RESULTS: 22 patients had bilateral phaeochromocytomas on histopathological examination (79%) and 6 patients had initially unilateral phaeochromocytomas There was no operative mortality or substantial morbidity except for one splenic injury that necessitated splenectomy. During a mean follow up period of 14 years (range 1-26) nine patients (32%) had a total of 19 Addisonian crises that necessitated admission to hospital. One patient died of an unrecognised Addisonian crisis. CONCLUSION: Complications of adrenocortical supplementation therapy are considerable, but they can be reduced when unilateral adrenalectomy is done for a unilateral phaeochromocytoma in patients with MEN 2 syndrome, provided that they are carefully followed up. PMID- 10533757 TI - Extent of thyroidectomy in nodular thyroid disease. AB - OBJECTIVE: To find out which procedure was the safest for each indication for operation in diseases of the thyroid gland. DESIGN: Retrospective study. SETTING: Two teaching hospitals, The Netherlands. SUBJECTS: 599 consecutive patients who had 601 thyroid operations between 1 October 1985 and 1 June 1993. MAIN OUTCOME MEASURES: Incidence of complications, particularly postoperative hypocalcaemia and injuries to the recurrent laryngeal nerve. RESULTS: Accidental injuries to the recurrent laryngeal nerve occurred in 0.7% of the nerves at risk (7/948) and the incidence of permanent hypocalcaemia was 5.2% (31/599). In subtotal procedures (bilateral subtotal thyroidectomy with the remnant left dorsally or total hemithyroidectomy combined with subtotal hemithyroidectomy on the other side with a remnant left at the upper pole) the rate was 11/390 (2.8%) compared with 18/525 (3.4%) after total resections. The corresponding numbers of accidental injuries to the recurrent laryngeal nerve were 2 and 4. CONCLUSIONS: Total thyroidectomies are more likely to be done for malignant disease, so the slightly higher complication rates probably reflect the nature of the disease, which requires more radical resection. Both subtotal procedures can be done with comparable low morbidity. PMID- 10533760 TI - Effects of prolonged stay in hospital on gastric flora. AB - OBJECTIVE: To find out if a long preoperative stay in hospital results in the introduction of micro-organisms into the gastric flora that are resistant to the usual perioperative chemoprophylaxis. DESIGN: Observational study. SETTING: Teaching hospital, Greece. SUBJECTS: 145 consecutive patients admitted for abdominal or pelvic operations. INTERVENTIONS: Aspiration of gastric fluid for measurement of pH and culture. MAIN OUTCOME MEASURES: Growth and type of pathogens in gastric aspirate. RESULTS: 125 patients spent a mean (range) of 13.6 days (7-69) in hospital preoperatively and 20 patients spent 3.1 (1-7) days. In the long-stay group 103 had a gastric pH of <4 and in 22 it was 4 or more; in 40 patients pathogens were grown from the gastric aspirate but resistant Pseudomonas cepacia was grown from only 2. In the short-stay group 16 patients had a pH of <4 and in 4 it was 4 or more; in 4 patients pathogens were grown from the aspirate. CONCLUSION: Prolonged stay in hospital preoperatively does not seem to affect the gastric flora, so routine chemoprophylaxis should be sufficient in abdominal operations. PMID- 10533759 TI - Detection of cancer in augmented breasts by positron emission tomography. AB - OBJECTIVE: To assess the diagnostic efficiency of positron emission tomography with 18-fluorine fluorodeoxyglucose in detecting breast cancer in augmented breasts. DESIGN: Retrospective study. SETTING: University hospital, Korea. SUBJECT: 9 cases or 8 patients with breasts augmented with paraffin or silicone. INTERVENTION: FDG-PET, mammography, and ultrasonography RESULTS: The mammogram detected the breast cancer in only 1 of 3 patients, and ultrasonography gave a false positive result in 1 patient with an augmented breast. In contrast, PET predicted all the cancers and 5/6 benign lesions. 2/3 breast cancers had axillary FDG uptake interpreted as showing metastatic involvement, and in 1 case with cancer with no axillary lymph node involvement there was no FDG uptake in the axilla, which correlated with the pathological finding. CONCLUSIONS: Although the high cost of PET makes its use as a screening test for all patients with augmented breasts unrealistic, it would be the best diagnostic choice if other methods failed. PMID- 10533762 TI - Quantitative measurements of venous reflux by duplex scanning of the incompetent long saphenous vein before and after high ligation at the saphenofemoral junction. AB - OBJECTIVE: To assess the effect of high ligation of the saphenofemoral junction (SFJ) on the amount of reflux in the long saphenous vein using quantitative duplex scanning. DESIGN: Prospective study. SETTING: Teaching hospital, The Netherlands. SUBJECTS: 23 patients presented with 29 limbs showing signs of isolated insufficiency of the long saphenous vein. INTERVENTION: High ligation of the SFJ. MAIN OUTCOME MEASURES: Duration of reflux (s), peak velocity (m/s), mean velocity (m/s) and mean flow (ml/s) at the SFJ, and in the mid-thigh, the distal thigh and the proximal lower leg before and four weeks after high ligation. RESULTS: All variables except duration of reflux improved significantly at all levels in the 29 limbs studied. One limb showed no improvement in flow at the level of the SFJ, 9 in the mid-thigh, 13 in the distal thigh, and 15 in the proximal lower leg (just below the knee). CONCLUSION: High ligation was effective in reducing reflux at the SFJ, but in about half the limbs the distal reflux in the long saphenous vein remained. PMID- 10533761 TI - Obesity surgery: discouraging long term results with Mason's vertical banded gastroplasty. AB - OBJECTIVE: To evaluate the long term results of Mason's vertical banded gastroplasty (VBG) using accepted criteria, and to find out which factors predicted success. DESIGN: Retrospective survey of a cohort of 40 severely obese patients (mean initial body mass index (BMI in kg/m2): 43, range 34-62). SETTING: General teaching hospital, The Netherlands. MAIN OUTCOME MEASURES: Success according to three definitions: weight loss of more than 25%; percentage of excess weight 50% or less; and BMI < 30. RESULTS: Mean follow-up was 7.4 years (range 0.5-10) or 85%. The distribution over MacLean and Reinhold criteria shows a shift towards unfavourable categories. The consecutive percentages of success at five years were 35%, 62%, and 35%. Logistic regression analysis of success at 5 years shows that the following factors significantly predicted success: Definition I: age odds ratio (95% confidence interval): 0.88 (0.78 to 0.99). Definition II: age: 0.84 (0.69-1.01), outlet > or = 5 cm: 176 (2.4 to 12774), percentage of ideal weight > or = 100%: 0.03 (0.002 to 0.48). Definition III: age: 0.86 (0.75 to 0.99), pouch size > or = 15 ml: 10.64 (1.48 to 76.6). CONCLUSION: The long term results of VBG are disappointing when assessed by the standard criteria. PMID- 10533763 TI - Implications of histological grade of tumour for the prognosis of radically resected periampullary adenocarcinoma. AB - OBJECTIVE: To study the influence of histological grade of tumour on the prognosis of radically resected periampullary cancers. DESIGN: Retrospective study. SETTING: Teaching hospital, Austria. SUBJECTS: 156 patients (papilla of Vater, n = 34, head of the pancreas, n = 105, and distal common bile duct, n = 17) who underwent partial pancreaticoduodenectomy for periampullary adenocarcinoma between 1 January, 1967 and 31 December, 1996. OUTCOME MEASURES: The relation between grade of tumour and site, T and N classification, extramural growth, invasion of vessels and resection margins, tumour volume, and survival time. RESULTS: Well differentiated lesions were significantly more common in the papilla of Vater (n = 15, 44%, p = 0.01) than in the pancreatic head or the common bile duct (n = 20, 19%, and n = 5, 29%, respectively). Only in ampullary lesions did the grade of tumour significantly affect the incidence of other histopathological risk factors (T p = 0.003; nodal status p = 0.01; extramural growth p = 0.0001; tumour volume p = 0.02) and survival time (p = 0.02); no significant correlations were found in cancers of the head of the pancreas or common bile duct. CONCLUSIONS: There was a significant difference in the distribution of grade of tumour between the different sites of origin of resected periampullary cancers. Grade of tumour correlated with T and N classification, tumour volume, extramural growth, and survival only in ampullary lesions. PMID- 10533764 TI - Lipid peroxidation and antioxidant state after laparoscopic and open cholecystectomy. AB - OBJECTIVE: To measure the amount of lipid peroxidation and erythrocyte antioxidation in patients undergoing laparoscopic and open cholecystectomy and healthy controls. DESIGN: Non-randomised study. SETTING: University hospital, Istanbul. SUBJECTS: 31 patients, of whom 14 underwent open and 17 laparoscopic cholecystectomy, and 15 healthy controls. INTERVENTIONS: Heparinised blood samples were taken from the patients immediately after operation and from the healthy controls. MAIN OUTCOME MEASURES: Lipid peroxidation index as expressed by thiobarbituric-acid-reactive substances (TBARS) and components of the erythrocyte antioxidant defence system, namely reduced glutathione, reduced glutathione peroxidase (glutathione-Px) and CuZn superoxide dismutase (CuZn SOD) in patients undergoing open or laparoscopic cholecystectomy and healthy controls. RESULTS: All 4 variables were significantly higher in the cholecystectomy groups than in controls (p < 0.001), and laparoscopic cholecystectomy caused significantly less oxidative stress than the open operation (p < 0.001). CONCLUSION: Both types of cholecystectomy cause oxidative stress and lead to an adaptive antioxidant response in the body. However; both oxidative stress and the antioxidant response are more pronounced after traditional open cholecystectomy. PMID- 10533765 TI - Piperacillin/tazobactam compared with cefuroxime/ metronidazole in the treatment of intra-abdominal infections. AB - OBJECTIVE: To assess the effect of piperacillin/tazobactam compared with cefuroxime/metronidazole in the treatment of patients with intra-abdominal infections. DESIGN: Randomised open study. SETTING: 16 Swedish and 6 Norwegian hospitals. SUBJECTS: 269 patients with intra-abdominal infections were randomised and treated with at least one dose of each study drug. 205 patients, 105 treated with piperacillin/tazobactam and 100 with cefuroxime, were clinically evaluable for follow up (had been given the full course of treatment). INTERVENTION: Patients were given piperacillin 4g/tazobactam 0.5 g every 8 hours or cefuroxime 1.5 g every 8 hours plus metronidazole 1.5 g every 24 hours. Each patient was to be treated for a minimum of 3 days and not more than 10 days. MAIN OUTCOME MEASURES: Clinical evaluation of infection at the end of and 4-6 weeks after treatment. Evaluation of safety and tolerance to the drugs and bacteriological susceptibility to the treatment drugs. RESULTS: In the intention to treat analysis treatment was equally successful for piperacillin/ tazobactam (103/140, 74%) and the cefuroxime/metronidazole groups (90/129, 70%) (p = 0.6). Corresponding figures for the clinically evaluable group were 102/105 (97%) and 94/100 (94%) for piperacillin/tazobactam and cefuroxime/metronidazole groups, respectively, at the end of treatment. At late follow up, 92/105 (88%) and 83/100 (83%) in the two groups, respectively, remained free of infection. The side effects of the treatment were mild and evenly distributed between the two groups. Most pathogens were susceptible to the drugs in both treatment groups. CONCLUSION: Both piperacillin/tazobactam and cefuroxime/metronidazole are well suited to the treatment of patients with intra-abdominal infections, and we found no significant difference between the two. The drugs were safe and well tolerated in the regimens used. PMID- 10533766 TI - Subcutaneous tissue oxygen and carbon dioxide tensions during hyperbaric oxygenation: an experimental study in rats. AB - OBJECTIVE: To investigate the response of subcutaneous tissue oxygen (O2) and carbon dioxide (CO2) tensions to hyperbaric oxygenation. DESIGN: Experimental study. SETTING: University hospital, Finland. SUBJECTS: 10 Wistar rats. INTERVENTION: Subcutaneous tissue PO2 and PCO2 were directly measured with an implanted Silastic tube tonometer and capillary sampling technique while breathing air and exposed to hyperbaric oxygen (HBO) at 2.5 or 2.8 ATA pressure. Hyperbaric exposures were carried out in a large multiplace chamber pressurised with air. MAIN OUTCOME MEASURES: Subcutaneous tissue PO2 and PCO2. RESULTS: The mean subcutaneous PO2 rose from the baseline of 8 kPa (60 mmHg) to 16 kPa (112 mmHg) when rats breathed room air during pressurisation to 2.8 atm. When the rats breathed oxygen at 2.5 ATA the maximal mean tissue PO2 was four times higher than the mean starting value. During the HBO treatment at 2.8 ATA the tissue PO2 rose to a value about five times above baseline. The tissue PCO2 values almost doubled during the exposure to HBO at 2.5 ATA, probably because elimination of carbon dioxide was impaired. CONCLUSION: Measurements of tissue PO2 and PCO2 with an implanted Silastic tonometer and a capillary sampling technique can successfully be adapted to hyperbaric conditions. The method yielded reproducible results and is applicable to clinical use in hyperbaric medicine. PMID- 10533768 TI - Immune function of the upper splenic remnant supplied by short gastric vessels. AB - OBJECTIVE: To study the immune function of the upper third of the spleen supplied by short gastric vessels after two thirds partial splenectomy. DESIGN: Experimental study. SETTING: Teaching hospital, Turkey. MATERIAL: Sixty Wistar albino rats, 20 in each group. INTERVENTIONS: Control = sham laparotomy; partial splenectomy = the upper third of the spleen supplied by short gastric vessels was preserved after two thirds partial splenectomy and dividing the main vascular supply; and total splenectomy. At the end of the sixth week postoperatively, antigenic stimulation was induced with an injection of pneumococcal suspension in 10 animals from each group. 0.5 ml of diluted Indian ink was injected into the aorta. MAIN OUTCOME MEASURES: Histological architecture of splenic tissue, and changes in the white pulp after antigenic stimulus. Bacteriological analysis with aerobic blood culture. Phagocytic activity as counted by Indian-ink-laden macrophages. The ability to produce antibodies as measured by serum IgM concentrations. RESULTS: Histological architecture of splenic tissue was normal. Germinal centres (p = 0.02), lymphoid follicles (p = 0.09), and their ratio (p = 0.0006) in the white pulp of the splenic remnant was significantly increased after antigenic stimulus compared with normal spleen. Significantly more animals without spleens developed bacteraemia (p = 0.02). Phagocytic activity of the upper splenic remnant was 89% that of normal spleen. Serum IgM concentrations without antigenic stimulus were 144, 138.2 (p = 0.6), and 86.2 (p < 0.001) mg/L; and with antigenic stimulus 263, 201.7 (p < 0.0001), and 98.1 (p < 0.0001) mg/L in groups 1, 2, and 3, respectively. The increase in serum IgM concentrations as a response to antigen was significant in the control (p < 0.0001) and in the partial splenectomy group (p < 0.0001), but not in the splenectomy group (p = 0.1). CONCLUSIONS: After reduction of its volume, the upper splenic remnant remained adequately supplied by the short gastric vessels. The upper part of the spleen preserved its normal histological architecture, had considerable phagocytic activity, possessed the ability to produce antibodies, and created a satisfactory immune response to antigenic stimulus. In rats, a considerable volume of functional and well perfused splenic tissue is preserved even after dividing the main vessels. PMID- 10533767 TI - Effects of octreotide on acute pancreatitis of varying severity in rats. AB - OBJECTIVE: To find out the effects of the octreotide on the course of acute pancreatitis in rats. DESIGN: Prospective laboratory study. SETTING: Medical school, Turkey ANIMALS: 184 Sprague-Dawley rats, 120 of which were randomly allocated into 8 groups of 15 each for the survival study, and the remainder of which were randomly allocated into 8 groups of 8 rats each for assessment of biochemical variables and histological score. INTERVENTIONS: The same 8 groups were used for the two parts of the study: saline alone (control), octreotide alone (control), oedematous pancreatitis induced by cerulein with and without octreotide, moderate pancreatitis induced by low-dose glycodeoxycholic acid and cerulein with and without octreotide, and severe pancreatitis induced by high dose glycodeoxycholic acid and cerulein with and without octreotide. MAIN OUTCOME MEASURES: Mortality, results of biochemical tests, and histological score. RESULTS: No rats in the control groups died. Of those with oedematous pancreatitis 1 died that had not been given octreotide (7%) and 2 that had (13%). In the moderate pancreatitis groups 4 that had not been given octreotide died (27%) compared with one that had (7%). In the severe pancreatitis group 7 that had not had octreotide died (46%) compared with 6 that had (40%). Octreotide caused a reduction in serum amylase and lactate dehydrogenase activity in all groups, but reduced aspartate aminotransferase only in those rats with moderate pancreatitis. It prevented hypocalcaemia in rats with severe pancreatitis, but had no effect on serum electrolyte concentrations, alkaline phosphatase activity, or blood gas analyses. Rats with moderate pancreatitis that had been given octreotide had less tissue oedema, acinar necrosis, and inflammatory cell infiltration. In those with severe pancreatitis there was less tissue oedema but more acinar necrosis. CONCLUSION: If octreotide is given early in the course of the disease it may result in improved outcome, but it seems to be ineffective in severe pancreatitis in which acinar necrosis is already established. PMID- 10533769 TI - Pneumomediastinum after nasal fracture. PMID- 10533770 TI - Anal implantation of exfoliated tumour cells from a rectal adenocarcinoma after colorectal stapled anastomosis. PMID- 10533771 TI - Retrocaecal hernia: an unusual case with a giant sac and volvulus of the herniated small bowel. PMID- 10533772 TI - Subtotal cystectomy as a complication of hernia repair in infants. PMID- 10533773 TI - Improved ventilatory function after combined operation for pulmonary emphysema and lung cancer. AB - BACKGROUND: Smoking is the leading cause of both lung cancer and emphysema. Therefore, some patients with stage I and II disease will present with contra indications to resection including a predicted postoperative FEV1 of less than 0.81 or a VO2max of less than 10 ml/kg/min. Recently, lung volume reduction surgery (LVRS) has re-emerged in the management of emphysema with excellent results. METHODS AND PATIENTS: 2 patients are reported with lung cancer in the left lower lobe and emphysematous destruction in both upper lobes. They, respectively, had a predicted postoperative FEV1 of 0.9211 and 0.6851. No metastases were present. Pre-operatively, a COPD index of 0.9 and 0.7 was calculated. A left lower lobectomy together with volume reduction of the left upper lobe was performed through a standard posterolateral thoracotomy. RESULTS: Pathological examination showed, respectively, stage IIb and stage Ib disease. The postoperative course was uneventful and 3 months later a FEV1 of 1.441 for patient 1 and 1.041 for patient 2 were recorded. CONCLUSION: These findings suggest that pulmonary function criteria for pulmonary resection have to be revised when patients can undergo simultaneous lung cancer resection and LVRS. The pre-operatively calculated COPD index can be used to predict which patients may not have a decrease in ventilatory function. PMID- 10533774 TI - Surgical treatment of the substernal goitre. AB - This report describes the surgical experience gained from 32 patients with substernal goitre, operated on during January 1995 to December 1997. The material corresponds to 15.7% of the total thyroidectomies performed. The diagnosis was clinical. In spite of 65.6% of the patients being asymptomatic, breathing problems, dysphagia and hyperthyroidism were observed. The patients underwent tests of thyroid function, simple chest radiograph and computerized tomography of mediastinum. All patients underwent surgical treatment through a transverse cervical incision. Two patients (6.25%) needed median sternotomy. Vocal cord palsy (3.12%), transient hypocalcemia (6.25%) and one death due to cardiac causes (3.12%) were the complications that occurred. PMID- 10533775 TI - Survival and prognostic factors in patients with resected epidermoid oesophageal carcinoma. AB - To determine long-term survival of patients with thoracic oesophageal tumor who underwent resection, and to identify possible associated prognostic factors, 58 patients underwent oesophagectomy alone (group A) and 16 combined with neo adjuvant chemoradiotherapy (group B). Univariate and multivariate analysis of prognostic factors were performed for age, depth of oesophageal wall tumour penetration, node involvement, type of resection, TNM stage and degree of tumour differentiation. Long-term survival rates at 1-5 years were 81% versus 89%; 56% versus 67%; 30% versus 67%; 12% versus 44%; and 0% versus 33% for group A and B, respectively (P = 0.0543, NS). Univariate analysis revealed only depth of invasion (P = 0.0076) and TNM stage (P = 0.0452) as isolated prognostic factors for long-term survival and multivariate analysis did not demonstrate any independent factor. Despite the small number of cases, neoadjuvant chemoradiotherapy seems to improve prognosis as well as to allow resection in a greater number of cases due to tumor downstaging. PMID- 10533776 TI - Hepatic functional reserve in patients with biliary malignancies: an assessment by technetium 99m galactosyl human serum albumin hepatic scintigraphy. AB - PROBLEM: Technetium 99m galactosyl human serum albumin (99mTc-GSA) is a hepatic scintigraphy agent that binds to the asialoglycoprotein receptors. We evaluated the clinical use of the scintigraphy for the pre-operative assessment of biliary malignancies. PATIENTS AND METHODS: Scintigraphy was performed before operation in 56 patients with biliary malignancies. A hepatic uptake ratio of 99mTc-GSA (LHL15; the count ratio of the liver to the sum of the heart and liver 15 min after injection of 99mTc-GSA) was calculated. RESULTS: LHL15 was significantly associated with bilirubin half-life in patients treated before operation with percutaneous transhepatic biliary drainage (P = 0.007). After operation, 4 of 18 patients with LHL15 < 0.925 died within 30 days. The postoperative mortality was significantly greater in patients with LHL15 < 0.925 than in patients with LHL15 > or = 0.925 (P = 0.033). CONCLUSION: 99mTc-GSA scintigraphy is useful in evaluating the hepatic functional reserve in patients with biliary malignancies. PMID- 10533777 TI - Impact of bactibilia on the development of postoperative abdominal septic complications in patients with malignant biliary obstruction. AB - This study was intended to clarify the role of bile bacteria in the development of postoperative septic complications in patients with malignant biliary obstruction. A total of 116 patients with malignant biliary obstruction underwent surgical intervention after biliary decompression. The relation between contaminated ductal bile and postoperative abdominal septic complications was analyzed retrospectively. Such complications developed in 49 patients (42%). Bile contaminated operations (n = 93) resulted in a higher incidence of septic complications than non-bile-contaminated operations (n = 23; P = 0.009). Patients with pre-operative positive bile culture had a higher incidence of septic complications than those with negative bile culture (P = 0.06). There was a positive correlation between the presence of pre-operative cholangitis and the occurrence of septic complications (P = 0.007). Bacteria found in pre-operative ductal bile were also detected in infected sites of 80% of patients with septic complications. Intra-operative contamination from infected ductal bile plays a critical role in the development of postoperative abdominal septic complications. PMID- 10533778 TI - The different manifestation and outcome between pancreatitis and pancreatic malignancy with left-sided portal hypertension. AB - Left-sided portal hypertension can be induced by isolated splenic venous obstruction due to various etiologies, such as chronic pancreatitis and pancreatic malignancy. The patients may present with bleeding isolated gastric varices and hypersplenism in addition to their pancreatic lesions. In the past 3 years, we have encountered 24 patients with left-sided portal hypertension. They were diagnosed with an abdominal echogram, CT or splenoportography. Twelve patients had histories of acute pancreatitis for a few months to years. Eleven of them were found to have isolated gastric varices. Six of them underwent operation due to hypersplenism or pseudocyst. The postoperative courses were smooth and the gastric varices subsided after splenectomy. The other 12 patients with left-sided portal hypertension were diagnosed as having pancreatic malignancy. Only two of them were found to have isolated gastric varices. Seven of them received operations and only two patients with their tumors located at the pancreatic body and tail could be resected. The other 5 patients were diagnosed with abdominal CT and high serum CA 19-9. We concluded that the patients with left-sided portal hypertension can be suspected by isolated gastric varices without liver cirrhosis. The diagnosis can be confirmed by abdominal CT or splenoportography. The incidence of isolated gastric varices are significantly lower in the patients with pancreatic malignancy than those with chronic pancreatitis. The gastric varices subsided after splenectomy. The prognosis of pancreatic malignancy is poor and most of them are inoperable. PMID- 10533779 TI - Pancreatic carcinoma: < or = 2 cm versus > 2 cm in size. AB - BACKGROUND: Despite recent progress in diagnosis and therapy, the clinical course of patients with pancreatic carcinoma remains dismal. There have been several approaches to improve the clinical course of patients with pancreatic carcinoma, namely: (i) detection of small pancreatic carcinoma; (ii) radical resection with retroperitoneal clearance and portal vein resection; (iii) multidisciplinary therapy including chemoradiation; and so forth. METHODS: In this series, eight Japanese patients with small pancreatic carcinoma measuring less than 2 cm in diameter (including two with non-invasive carcinoma and one with minimally invasive carcinoma) and 53 with larger pancreatic carcinoma were reviewed to find the diagnostic and therapeutic clues to improve the clinical course of patients with pancreatic carcinoma. RESULTS: Lymphatic (ly) and perineural (pn) permeation was significantly more frequent and extensive in the 53 patients with large pancreatic carcinoma than in the eight with small pancreatic carcinoma (ly 0/1/2/3:3/24/18/8 versus 5/2/1/0, P = 0.0284; pn 011/2/3:4/29/14/6 versus 3/2/3/0, P = 0.0491). The surgical margin was affected by malignant cells in 18 (34%) of the 53 patients with large carcinoma but none (0%) of the eight with small carcinoma (P = 0.0004). The comprehensive stage was significantly earlier in the eight with small carcinoma than in the 53 with large carcinoma (comprehensive stage I/II/II/IV:4/0/3/1 versus 0/3/26/24, P < 0.0001). Comprehensive curability of the eight small carcinoma cases was significantly higher than that of the 53 large carcinoma cases (comprehensive curability A/B/C:5/3/0 versus 9/5/39; P = 0.0003). 1-year and 3-year cumulative survival rates of the eight patients with small carcinoma were 100% and 82%, whereas those of the 53 with large carcinoma were 51% and 17%, respectively (P = 0.0207). However, the eight small carcinoma cases already showed frequent invasion to the vascular (v), lymphatic (ly) and perineural (pn) structure [v(+): 3/8, ly(+): 5/8, pn(+):5 /8] and lymph node metastasis (n) [n(+): 3/8]. Out of the eight small pancreatic carcinomas, one minimally invasive carcinoma and two non invasive carcinomas showed no vascular, lymphatic or perineural invasion or lymph node metastasis. All the three patients have been doing well 19, 32 and 44 months after the operation. The diagnostic clues in the three patients were dilatation of the main pancreatic duct in one and of the branch duct in the other two. CONCLUSIONS: These findings suggest that surgical resection frequently cures patients with small pancreatic carcinoma but more effective adjuvant therapy should be developed to control lymphatic permeation, venous invasion or perineural infiltration in surgical resection of large pancreatic carcinoma. The supreme goal is to detect non-invasive or minimally invasive pancreatic carcinoma with a dilatation of the main or branch pancreatic duct as a diagnostic aid. PMID- 10533780 TI - Prognostic value of peritoneal lavage cytology and chemotherapy during surgery for advanced gastric cancer. AB - BACKGROUND: The prognostic value of intra-operative peritoneal lavage cytology and chemotherapy was evaluated retrospectively. METHOD: Lavage cytology was performed in 257 patients. Prognosis was investigated in 85 of pT3 and pT4 patients with radical gastrectomy. Intra-operative chemotherapy was selected according to the cytology and exploration of the peritoneal cavity. For patients forecasted to have peritoneal recurrence, intraperitoneal injection of cisplatin was performed. RESULTS: No free cancer cells (cy(-)) were found in pTis, pT1 and pT2. In pT3 and pT4, cy(-) were 82.8% of the cases without macroscopic metastasis (P(-)), and the presence of free cancer cells (cy(+)) were 89.3% of the cases with macroscopic metastasis (P(+)). Intraperitoneal injection was performed in about 60% of P(-)/cy(+) and P(+) cases. Five-year survival rate of P(-)/cy(-) was 41.7% and that of P(-)/cy(+) was 33.3%. All of P(+) died within 3 years. CONCLUSION: Patients of P(-)/cy(+) probably had microscopic residual disease and might benefit from intraperitoneal chemotherapy. PMID- 10533781 TI - Computer-aided diagnosis of the character of bowel obstruction. AB - A computer programme for the differential diagnosis of bowel obstruction was created and put into clinical practice to accelerate and simplify the diagnostic process. The training material was 503 cases of bowel obstruction admitted to the Surgical Clinic of Kaunas Medical University during 1990-1996. Based on 36 statistically significant anamnestical, clinical, laboratory investigations and plain abdominal X-ray findings, the computer programme was built up using a Bayesian formula. Retrospectively, the prognostic diagnosis was compared with the final clinical diagnosis based on instrumental or operative findings. Then, in a control group of 136 patients, the prospective prognostic diagnosis was obtained. The accuracy of the prognostic diagnosis in the control group of patients with complete small bowel obstruction amounted to 88.7% and for the patients with complete large bowel ileus 95.8%. Prognostic accuracy for partial small bowel obstruction was the most precise (96.1%) and for partial large bowel ileus (87.5%). The overall diagnostic accuracy of the computer algorithm was 92.6%. All cases were classified. This computer algorithmic programme for the differential diagnosis of the character of mechanical bowel obstruction has markedly shortened and facilitated the process of diagnosis of ileus. PMID- 10533782 TI - Effects of small bowel exclusion on intestinal myoelectrical activity pattern: comparison between innervated and denervated (transplanted) Thiry-Vella loops in rats. AB - Small bowel transplantation (SBT) leads to several changes in normal intestinal physiology with special reference to lymphatic disruption and graft denervation. Intestinal myoelectrical activity (MA) has been studied in different conditions, but little is known about MA in excluded bowel segments without the influence of nutrients. We performed this study to evaluate the effects of bowel exclusion on MA pattern. Fifteen Wistar rats were divided into two groups: five were used as donors and five as recipients for SBT; the remaining five underwent isolation of a jejunal segment as Thiry-Vella loop (TVL). On the 20th postoperative day, four bipolar electrodes were implanted in the small bowel of each rat: proximally and distally on the transplanted and the native intestine (SBT group); proximally and distally on the TVL and across the jejunal anastomosis (TVL group). On the 30th postoperative day, MA was recorded for 30 min after a 12 h fast. MA pattern was not altered by the exclusion of innervated jejunal segments (TVLs) with maintenance of high amplitude and migrating myoelectric complex (MMC) occurrence independent of MA in the continuity bowel. The characteristic regular spiking activity was not observed in transplanted grafts and MA analysis showed slow waves containing superimposed irregular spiking activity. PMID- 10533783 TI - Surgical treatment for colorectal endometriosis. AB - The authors studied the surgical treatment of patients with intestinal endometriosis. A total of 10 patients, with a median age range of 43 years, underwent an operation. Cramp abdominal pain (100%), diarrhea (30%), constipation and enterorrhagia (20%) dominated the clinical picture. At the time of surgery, four patients presented intestinal obstructive symptoms. Five (50%) patients reported gynecological complaints. Four patients were infertile and five had prior surgical gynaecological events. Seven cases presented sigmoid involvement, and three had involvement of the cecal appendix. Pre-operative diagnosis was carried out in two patients only. Surgical indications were due to suspicion of cancer (4 patients), appendicitis (3 patients), diverticular disease (1 patient) and unmanageable pain (2 patients). The following procedures were performed: left colectomy (2 cases), rectosigmoidectomy (3 cases), sigmoidectomy (3 cases), colostomy (2 cases) and three appendicectomy cases associated with concomitant gynecological interventions. No postoperative complications or deaths were observed. The authors emphasize that intestinal stenotic lesions should be treated by means of extirpation while the parietal nodule should be treated by exeresis. Intestinal endometriosis should be suspected in cases of lower abdomen recurrent pain in premenopausal infertile women or with previous surgical, gynecological events associated with intestinal symptoms or distal colon stenosis. PMID- 10533784 TI - Techniques for determining the ideal stoma site in laparoscopic colostomy. AB - The best placement of the stoma is one of the important issues in colostomy. We developed a new laparoscopic technique for colostomy. In the technique, because the port for the laparoscope was placed in the right side of the abdomen, it was easy to separate the descending colon from lateral peritoneum and to lift up the colon loop to the stoma site. In our experiences using this technique, patients had no serious complications. This technique enables the performance of safe, secure, and effective temporary colostomies. PMID- 10533785 TI - Evaluation and management of malfunctionings following implantation of the artificial urinary sphincter. AB - The authors report their experience of mechanical malfunctionings after placement of the AMS 800 artificial urinary sphincter. The aim of the study was to suggest outlines for the evaluation and management of these complications. From 1991 to 1998, 42 patients (39 men, 3 women with a mean age of 67.5 years) underwent artificial sphincter implantation of whom 5 patients required explantation of the prosthesis because of infection. The following mechanical malfunctionings occurred: 2 cases of air bubbles; 3 cases of pump overturning with tube kinking; 16 cases of persistent or recurrent incontinence 6-61 months postoperatively; and 1 case of cuff opened during sexual intercourse. Surgical revision of the prosthetic components was performed in the patients who presented mechanical malfunctionings. Several complications can arise after artificial sphincter placement (infection, urethral erosion, mechanical malfunction). In the last case, the authors stress the importance of an accurate diagnosis for the choice of exact treatment. PMID- 10533786 TI - Non-absorbable prosthetic meshes: which is the best option in the repair of abdominal wall defects? AB - The mechanical properties and macroscopic behaviour of non-absorbable materials have been widely studied. Nevertheless, biological tissue response to contact with these prostheses is not well-known. Our purpose was to compare the microscopic behaviour of two non-absorbable materials. Polypropylene and mersilene meshes were implanted on 36 female Wistar rats each (PPL and ME groups) . Six animals per group were sequentially sacrificed at 1, 2, 3, 5, 10 and 15 weeks. Global cell density and number of polymorphonuclear leukocytes, giant cells, fibroblasts and histiocytes were compared for every studied phase. The polypropylene group showed higher cell density and polymorphonuclear response in the initial phases, while scores for giant cells were higher in the mersilene group. Fibrohistiocytic reaction was increased in the polypropylene group. Polypropylene tends to provoke higher acute inflammatory reaction and connective tissue formation than mersilene. The latter induces higher foreign body reaction. PMID- 10533787 TI - Spontaneous hematoma of the rectus abdominis sheath: a review of 177 cases with report of 7 personal cases. AB - Hematoma of the rectus abdominis muscle sheath is a little known and rarely diagnosed condition, in spite of its definite clinical setting and treatment. It is very important to the surgeon, as it may be mistaken frequently for acute inflammatory abdominal conditions and should be considered in the differential diagnosis of intra-abdominal tumors. The literature on 177 cases of non-traumatic hematomas of the rectus abdominis muscle sheath is reviewed, including seven personal cases reported by the authors. Its epidemiology, pathogenesis, clinical features, diagnostic examinations and treatment are discussed. PMID- 10533788 TI - Donor selection for xenotransplantation: detection of Galalpha1-3Gal on different porcine organs. AB - The expression of the major porcine xenoantigens (Galalpha1-3Gal) in different tissues varies between species. The selection of suitable donors and the interpretation of studies which attempt to prevent hyperacute rejection are dependent on donor expression of Galalpha1-3Gal. Screening of large number of animals to find potential Galalpha1-3Gal negative donors requires a robust, tissue-based and practical method of assessing Galalpha1-3Gal expression. In this study, we have assessed the expression of Galalpha1-3Gal in a variety of pig organs using anti Galalpha1-3Gal antibody. Biopsies of heart, kidney, ear and tail were obtained from 20 outbred pigs. Biopsies were fixed in formalin and stained with a human anti Galalpha1-3Gal antibody obtained from pooled human AB serum passed down a Galalpha1-3Gal immunoadsorbent column. Tissue from all 4 organs from all 20 pigs expressed Galalpha13Gal. This study shows that detection of Galalpha1-3Gal on an ear or tail biopsy is a simple but very reliable method for assessing Galalpha1-3Gal expression on the heart and kidney and facilitates donor selection for xenotransplantation. PMID- 10533789 TI - Intra-operative hyaluronic acid clearance predicts postoperative graft function in living related liver transplantation. AB - Although intra-operative hyaluronic acid (HA) clearance has been reported to be a predictive parameter for early graft function after the implantation of a cadaveric hepatic graft, it has yet to be evaluated as a parameter in assessing graft function in living related liver transplantation. The aim of this study was to evaluate whether intra-operative HA clearance can be a predictive parameter of early graft function in living related liver transplant patients. Eight consecutive patients, who underwent a living related liver transplantation, were entered into the study. The HA clearance 180 min after reperfusion of the graft was evaluated. Significantly higher serum HA levels were found in the patients with fulminant hepatitis than in the patients with non-fulminant hepatitis before operation (P < 0.01), just before reperfusion (P < 0.01), and 180 min after reperfusion (P < 0.05). The HA clearance correlated with the peak total bilirubin within 5 postoperative days (P < 0.05) and the lactic acid one day after operation (P < 0.01). The intra-operative HA clearance serves as a sensitive parameter for assessing the postoperative graft function after the implantation of the new liver. Based on our findings, measuring the HA clearance was thus found to be clinically useful in the assessment of graft function in living related liver transplantation. PMID- 10533790 TI - Hepatic injuries: management and outcome. AB - A total of 52 patients with different grades of liver injuries were treated in a 1 year period: 32 patients had penetrating injuries and in 20 patients injuries resulted from blunt trauma. Blunt trauma victims were frequently associated with chest and head injuries and with skeletal fractures. A 4-fold incidence of associated intra-abdominal injuries was encountered in penetrating trauma victims. Blunt and gunshot victims commonly had severe grades of liver injuries (55% and 83.3%, respectively). Stab wounds caused simpler grades of injuries (86.7%). Suture hepatorraphy was the commonest procedure performed (57.7%).The other procedures applied to treat the injured liver were: selective ligation of the injured blood vessels (15.4%); packing (7.7%); and liver resection (3.8%). Two patients died on the operating table before any remedy was applied. The overall morbidity was 40.4%. The liver related complications constituted 17.3%. The total mortality was 13.5% and the liver related mortality was 9.6%. PMID- 10533791 TI - Factors effecting mortality in urban vertical free falls: evaluation of 180 cases. AB - AIM: The aim of this retrospective study was to evaluate the factors on mortality in urban free vertical falls. PATIENTS AND METHODS: A total of 180 urban vertical free fall victims who survived transport to the emergency room between the period of 1980-1998 were evaluated. Minor bruises, abrasions, haematomas, and soft tissue injuries were not encountered. Serious injuries such as bone fractures, liver lacerations, epi-subdural haematomas, haemothorax, haemomediastinum, retroperitoneal haematomas were evaluated. RESULTS: Of the total, 23% (n = 41) of patients were female and 73% (n = 139) were male. The mean age was 22.3 years (4 75 years). Extremity fractures were found in 6.7% (n = 12), cranial trauma in 14.4% (n = 26), thoracic trauma in 2.2% (n = 4) retroperitoneal trauma in 2.8% (n = 5), vertebral column trauma in 1.7% (n = 3) of cases. The overall number of the pathologies was 59. In-hospital mortality was 8.9% (n = 16). The injury severity scores (ISSs) of non-survivors and survivors were 33 +/- 4, and 5 +/- 0.6, respectively (P = 0.0001). The heights fallen were 8.6 +/- 2.3 m for non survivors and 5.2 +/- 0.2 m for survivors (P = 0.022). The mean ages of non survivors and survivors were 41.6 +/- 5.9 years and 20.4 +/- 1.2 years, respectively (P = 0.003). Serious cranial trauma was found in 68.7% (n = 11) and 9.1% (n = 5) of non-survivors and survivors, respectively (P = 0.0001). Extremity trauma was encountered in 31.2% (n = 5) and 4.2% (n = 7) of non-survivors and survivors, respectively (P = 0.0015). The ISSs were 6.8 +/- 1.0 and 8.9 +/- 1.1 for cases under the age of 6 years and others, respectively (P = 0.15). Using logistic regression analysis, ISS, height and age were found to be significant factors in mortality. CONCLUSION: Vertical deceleration injury represents a distinct form of trauma. With the results of this study, it can be concluded that ISS, height and age are significant factors in determining the severity of trauma. PMID- 10533792 TI - Mitochondrial respiratory chain disorders and the liver. AB - Mitochondrial respiratory chain disorders are an established cause of liver failure in early childhood but they are probably under-diagnosed, partly due to under-recognition and partly due to the difficulty of investigation. It is particularly important to look for mitochondrial disorders if the liver disease presents with hypoglycaemia and lactic acidaemia or if it is accompanied by neurological, muscle or renal tubular abnormalities. Respiratory chain defects have been demonstrated in a number of patients who die of liver failure following severe epilepsy; this includes at least some cases of Alpers syndrome or 'progressive neuronal degeneration of childhood'. In mitochondrial liver disease, histology usually shows steatosis, often accompanied by fibrosis, cholestasis and loss of hepatocytes. Unless the clinical picture suggests a particular syndrome, such as Pearson syndrome, biochemical assays and histochemistry should be the initial investigations. Ideally, investigations should be carried out on liver as well as more standard tissues, such as muscle, since defects can be tissue specific. Nuclear defects and mtDNA point mutations are probably responsible for many cases of mitochondrial liver disease but, as yet, the only identified molecular abnormalities are mtDNA rearrangements and mtDNA depletion. Treatment of mitochondrial liver disease is unsatisfactory. If the disease is confined to the liver, transplantation may be appropriate but in several patients transplantation has been followed by the appearance of disease in other organs, particularly the brain. PMID- 10533793 TI - Hepatic expression of viral antigens, hepatocytic proliferative activity and histologic changes in liver biopsies of children with chronic hepatitis B after interferon-alpha therapy. AB - AIMS/BACKGROUND: The purpose of the present study was to evaluate the effects of interferon-alpha (IFNalpha) treatment on necro-inflammatory changes, viral antigen expression and hepatocytic proliferation rate assessed by Ki-67 protein in liver biopsies of children with chronic hepatitis B virus (HBV) infection. METHODS: The study included 18 children at prepubertal age. Histologic changes were assessed using a modified scoring system for grading of histological activity index and staging of fibrosis. The hepatocytic expression of Ki-67 and HBV antigens including HBV surface antigen (HBsAg) and HBV core antigen (HBcAg) were evaluated using a semi-quantitative immunohistochemical scoring system. RESULTS: We found a significant decrease in the scores of intralobular confluent and spotty necrosis, periportal piecemeal necrosis, and in Ki-67 immunopositivity after treatment. Serologic response with clearance of HBV e antigen in 9 patients (50%) was associated with this improvement. The extent of fibrosis and scoring of portal inflammation, however, did not show any difference. HBcAg expression showed a significant decrease after treatment, whereas HBsAg expression either increased or remained unchanged. CONCLUSION: We conclude that IFNalpha treatment might provide an improvement in hepatic histology with a reduction in hepatocytic injury. It might also provide a serologic response associated with a decrease in hepatocytic HBV replication. PMID- 10533794 TI - Determination of serum hepatitis C virus (HCV) core protein using a novel approach for quantitative evaluation of HCV viraemia in anti-HCV-positive patients. AB - AIMS/BACKGROUND: Hepatitis C virus (HCV) infection is frequently diagnosed by detection of antibody to the HCV (anti-HCV). Recently, a method for detection of HCV core protein, "Imucheck F-HCV Ag Core Kokusai", has been developed. In this study, we evaluate the utility of this method. METHODS: HCV core protein levels in sera were determined using this following method; anti-HCV titres were measured by particle agglutination (PA) test and then quantitative HCV-RNA values were investigated using a competitive reverse transcription-polymerase chain reaction (RT-PCR) test. RESULTS: The HCV core protein was detected only in anti HCV-positive sera. Of 490 anti-HCV-positive sera, 130 (26.5%) were positive by this method. Of 144 anti-HCV-positive/HCV-RNA-positive sera, 130 (90.3%) were positive by it. A significant correlation between the HCV core protein levels and quantitative HCV-RNA values was recognized (n= 110, r=0.86, p<0.01). A significant correlation between the HCV core protein levels and alanine aminotransferase titres was also observed (n=67, r=0.72, p<0.05). All 71 patients with chronic active hepatitis, cirrhosis and hepatocellular carcinoma were positive with this method, whereas 18 of 32 patients with chronic inactive hepatitis were positive. Twenty-three patients with chronic active hepatitis were treated with interferon-alpha. During therapy, some patients showed a negative conversion of HCV core protein. One (7.7%) of 13 patients with HCV genotype 1 and 5 (62.8%) of 8 patients with genotype 2 remained negative for 6 months after the therapy. CONCLUSION: This method may be useful for quantitative evaluation of HCV viraemia in anti-HCV-positive patients. PMID- 10533795 TI - Long-term treatment of chronic hepatitis C with ursodeoxycholic acid: influence of HCV genotypes and severity of liver disease. AB - AIMS/BACKGROUND: Current therapy for chronic hepatitis C virus (HCV) infection is based on the administration of interferon alpha (IFN) alone or in combination with other anti-viral agents. However, such therapy is effective in only a minority of selected patients. Long-term ursodeoxycholic acid (UDCA) treatment has been reported to improve liver function and structure especially in cholestatic disorders. We investigated the effect of long-term UDCA treatment on liver function in respect to the severity of chronic liver disease and HCV genotypes. METHODS: Forty-five patients with non-cholestatic laparoscopy-biopsy proven HCV-associated chronic hepatitis (n=16) or cirrhosis (n=29) who had not responded to, or were unsuitable for IFN, were randomly assigned to receive UDCA (600 mg/day; n=23) or no therapy (n=22) for 12 months. At entry, all patients were evaluated by means of conventional and quantitative liver function tests (LFTs), including galactose elimination capacity and antipyrine clearance, HCV antibodies, HCV-RNA and HCV genotypes. LFTs were measured at 6 and at 12 months, whereas HCV-RNA was determined again after treatment. RESULTS: Baseline characteristics were comparable in the two study groups. Long-term UDCA therapy was well tolerated. Based on the analysis of variance, there was a significant decrease in serum transaminase, LDH and GGT levels in UDCA treated patients. By contrast, the activities of these enzymes increased in untreated patients, with AST levels reaching statistical significance only. Statistical analysis also showed that the improvement in biochemical markers was more pronounced in UDCA treated patients with liver cirrhosis than in those with chronic hepatitis but was similar in patients with HCV genotype 1b and non-1b. However, HCV-RNA was positive in all patients after treatment. Quantitative LFTs remained, on average, stable over the 12 months of the trial in all groups. CONCLUSIONS: Long-term UDCA treatment is well tolerated in patients with HCV-associated chronic liver disease. The effect appears to be greater in cirrhotics than in patients with chronic hepatitis but is independent of HCV genotypes. Thus, long-term UDCA treatment, despite the absence of an anti-viral effect, seems beneficial in reducing disease activity in patients with chronic hepatitis or cirrhosis who are unsuitable for IFN therapy. PMID- 10533797 TI - Fcgamma receptor expression on hepatic macrophages and histological activity of chronic hepatitis. AB - AIMS/BACKGROUND: Activated liver macrophages in chronic hepatitis express a high affinity receptor for IgG named FcgammaRI. This study was performed to find the difference in FcgammaRI expression between chronic hepatitis B (CHB) and C (CHC) with reference to histological activity. METHODS: Consecutive patients with CHB (20 cases) and CHC (25 cases) were enrolled in the study. Inflammatory activity was evaluated using the modified histological activity index (HAI). FcgammaRI positive macrophages were quantitatively measured by computer assisted morphometry. RESULTS: Total HAI score was significantly higher in CHB than in CHC. Confluent necrosis was observed in significantly higher frequency in CHB at Stages 3 5 than in CHC. The percentage area of FcgammaRI-positive macrophages was significantly higher in CHB than in CHC. In CHB, the percentage area of FcgammaRI positive macrophages correlated with total HAI (<0.01) as well as the degree of confluent necrosis (<0.01), interface hepatitis (<0.05) and portal inflammation (<0.05). FcgammaRI-positive macrophages accumulated mainly at the site of confluent necrosis. In CHC, no correlation was observed between activated macrophages and any histological categories. CONCLUSION: These results suggest that FcgammaRI-positive macrophages are associated with confluent necrosis in CHB, which is more common in CHB patients than in CHC. PMID- 10533796 TI - Cerebral proton and phosphorus-31 magnetic resonance spectroscopy in patients with subclinical hepatic encephalopathy. AB - BACKGROUND/AIMS: In vivo magnetic resonance spectroscopy can be used to study cerebral metabolism non-invasively. We aimed to correlate 1H and 31P magnetic resonance spectral abnormalities in the brains of patients with subclinical hepatic encephalopathy. METHODS: Eighteen patients were studied at 1.5T, with combined 1H and 31P magnetic resonance spectra obtained from multiple voxels in the cerebral cortex and basal ganglia. Peak area ratios of choline, glutamine/glutamate, relative to creatine in the 1H spectra and percentage phosphomonoesters, phosphodiesters and betaNTP signals relative to total 31P signals in the 31P spectra were measured. RESULTS: Six patients did not complete the full examination - 31P results are available from 12 patients only. Relative to creatine, there were reductions in choline and elevations in glutamine/glutamate, varying across the brain with choline significantly reduced in occipital cortex (p<0.05) and glutamine/glutamate most significantly elevated in temporo-parietal cortex (p<0.0001). Percentage phosphomonoester (p<0.05), phosphodiester (p<0.05) and betaNTP (p<0.005) signals were significantly decreased in basal ganglia spectra. No correlation was found between the magnitude of 1H and 31P MRS changes, except between percentage phosphodiester decrease and glutamine/glutamate to creatine increase in occipital cortex. CONCLUSION: The results of this study point to a multifactorial aetiology for this condition. PMID- 10533798 TI - Immunolocalisation of PIVKA-II in paraffin-embedded specimens of hepatocellular carcinoma. AB - AIMS/BACKGROUND: Although serum PIVKA-II has been widely used as a tumor marker for hepatocellular carcinoma (HCC), little information is available concerning tissue PIVKA-II, and detection of tissue PIVKA-II in paraffin-embedded tissue specimens of HCC is also considered difficult. METHODS: Paraffin-embedded HCC tissues obtained at autopsy from 22 patients were subjected to immunohistochemical staining using the antibody MU-3 (Eisai Co., Ltd.) after microwave antigen retrieval. RESULTS: PIVKA-II was mainly detected in the cytoplasm of HCC cells. The intensity of tissue PIVKA-II staining was not correlated with serum PIVKA-II levels or with the histotogical differentiation of HCC. However PIVKA-II staining tended to be more intense in tissue from patients with portal tumor thrombus, distant metastases, or a longer duration of HCC. CONCLUSION: This method of immunohistochemical staining is easy and simple to use and may be helpful for detecting tissue PIVKA-II in paraffin-embedded HCC specimens. PMID- 10533799 TI - T1762/A1764 variants of the basal core promoter of hepatitis B virus; serological and clinical correlations in Chinese patients. AB - BACKGROUND: A double variant in the basal core promoter, converting nucleotide 1762 from A to T (T1762) and nucleotide 1764 from G to A (A[764), has been described in patients with chronic hepatitis B infection. Its prevalence and significance in Chinese chronic HBV carriers are unknown. METHODS: We studied 177 Chinese patients with chronic HBV infection (chronic hepatitis/asymptomatic: 89/88; hepatitis B e antigen positive/negative: 84/93). The double variant was detected by mismatched polymerase chain reaction and restriction fragment length polymorphism analysis. The reliability of this method was verified by sequencing in 41 serum samples with 100% matching. RESULTS: The double variant T1762/ A1764 was found in 52 of 89 patients with chronic hepatitis, but in only 6 of 59 asymptomatic carriers (p<0.001). The prevalence was significantly lower in hepatitis B e antigen positive patients (23/84) than in hepatitis B e antigen negative patients (35/64) (p<0.005). Precore variant, A1896 was detected in 40 individuals; 31 of them suffered from chronic hepatitis and 9 were asymptomatic (p<0.001). A combination of both variants T1762/A1764 and A1896 was seen in 3 of 59 asymptomatic and 22 of 89 patients with chronic hepatitis (p<0.005). CONCLUSIONS: Mismatched polymerase chain reaction with restriction fragment length polymorphism provides a reliable, easy and fast method for detection of the presence of the T1762/A1764 variant. In Chinese chronic hepatitis B carriers, T1762/A1764 variant was associated with both active liver disease and hepatitis B e antigen negativity. PMID- 10533800 TI - An in vitro model for the study of phagocytosis of damaged hepatocytes by rat Kupffer cells. AB - AIMS/BACKGROUND: One function of Kupffer cells is the phagocytosis of nonviable hepatocytes. Our aims were to develop a model for phagocytosis of damaged hepatocytes by rat Kupffer cells in vitro, and to characterise prostaglandin E2 (PGE2), prostacyclin (PGI), and tumour necrosis factor-alpha (TNF) production in this model. METHODS: Kupffer cells were incubated alone or with damaged hepatocytes for up to 18 h, then washed and cultured for up to 66 h. To compare mediator responses produced during inert particle phagocytosis, Kupffer cells were also incubated with latex beads. RESULTS: Phagocytic uptake of hepatocyte debris was confirmed in at least 50% of Kupffer cells. A dissociation between TNF and PGI responses was found for both latex beads and damaged hepatocytes, such that a TNF secretory response was not triggered by either stimulus whereas PGI production was increased for both. Although phagocytosis of beads increased PGE2 production, phagocytosis of hepatocytes did not. CONCLUSIONS: Phagocytosis of damaged hepatocytes by Kupffer cells results in the production of PGI but not PGE2 or TNF. PMID- 10533801 TI - Altered thyroid status modulates portal pressure in normal rats. AB - BACKGROUND & AIMS: Disturbances in thyroid function in humans and experimental animal models have been associated with alterations in liver function and portal circulation. We have previously shown that hypothyroidism can significantly reduce portal pressure in portal vein ligated rats as well as inhibit the development of cirrhosis and fulminant hepatic failure following toxic liver injury. The aim of this study was to determine the effects of increased and decreased thyroid function on portal pressure in rats with normal liver histology and portal circulation. METHODS: Three groups of 12 Wistar rats each were studied over a 30 day period: euthyroid (Group 1), hyperthyroid (Group 2) and hypothyroid (Group 3). Hyperthyroidism was induced by subcutaneous injection of triiodothyronine (400 microg/100g body weight) every ten days during the study period. Hypothyroidism was induced by methimazole (0.04% in drinking water) from 2 weeks prior to and throughout the 30 day study. Serum triiodothyronine (T3) and thyroid stimulating hormone (TSH) levels were determined to confirm the induction of hyper- and hypothyroidism. Portal pressure was assessed by direct catheterization of the portal vein prior to sacrifice. Indirect confirmation of changes in portal circulation was obtained by determining splenic weight at the time of sacrificing the animals. Animals were sacrificed at 10 day intervals throughout the 30 day study. RESULTS: Triiodothyronine treated rats were hyperthyroid compared to controls, with an elevation in serum T3 levels (3.8+/ 0.9 mmol/L vs 1.3+/-0.4 mmol/L, p<0.05). In rats treated with methimazole, hypothyroidism was confirmed by a 7-fold increase in serum TSH compared to controls (1.8+/-0.4 vs 0.24+/-0.04 mmol/L, p<0.01). Portal pressure was significantly higher in the triiodothyronine treated rats compared to controls (12.8+/-1.7 and 9.6+/-0.75 cm H2O, p<0.001). Splenic weights in hyperthyroid rats were significantly higher than in controls (579+/-44 vs 478+/-46 mg, p<0.01). Portal pressure was significantly lower in the methimazole treated group compared to the control group (8.13+/-0.68 vs 9.6+/-0.75 cm H2O, p<0.01) as were splenic weights (400+/-33 vs 478+/-46 mg, p<0.01). CONCLUSION: These studies demonstrate that disturbed thyroid function exerts significant hemodynamic effects on the portal circulation in normal rats and complements results from previous similar studies in cirrhotic animals. PMID- 10533802 TI - Hepatitis C infection in an Italian population not selected for risk factors. AB - AIMS/BACKGROUND: This study estimated the prevalence of HCV infection and relationship with viremia in a general population. The inhabitants of Albavilla town were personally invited to participate. METHODS: Out of 3997 inhabitants falling within the age range 18-85 years, 2403 (participation rate 60.1%) were examined for transaminases, HCVAb, HCVRNA, genotype and immunoblot assay. The following information was collected: sex, age, blood transfusions, surgery, use of glass syringes, drug addiction, alcohol consumption, tattoos and body mass index. RESULTS: 115 (4.8%) were HCVAb+, the prevalence being 1.2% under 40 years. Transfusion in the past was the only risk factor for HCV infection. Among the HCVAb+ subjects, 71 (61.7%) were HCVRNA+. 40.8% of the HCVAb+/HCVRNA+ group had normal ALT, compared with 68% of those with HCVAb+/HCVRNA-. The HCV genotypes in the 71 HCVRNA+ subjects were: 2a/2c in 58 (81.7%), 40% of them with normal ALT;1b in 11 (15.5%), none with normal ALT; genotype 3 in two (2.8%). CONCLUSION: The prevalence of HCVAb in this general population was 4.8%. About 3% were HCVRNA positive and of these genotype 2a/ 2c was present in 81.6%. PMID- 10533803 TI - Steatosis and bile duct damage in chronic hepatitis C: distribution and relationships in a group of Northern Italian patients. AB - BACKGROUND/AIMS: Hepatitis C virus (HCV) related disease follows a long, benign course and most affected patients have mild disease. Liver biopsy is mandatory to grade and stage the disease. Characteristic, though non-specific, HCV histological lesions such as bile duct damage and steatosis have been singled out but their association with non-histological parameters has not been completely defined. Our aim was to study the relationships among these histological lesions and clinical, biochemical, functional and virological characteristics in a group of Northern Italian patients with chronic hepatitis. METHODS: We studied 172 patients with HCV-related chronic hepatitis. Patients were divided into groups on the basis of histology including bile duct damage and steatosis. Clinical, biochemical, functional and virological profiles were related to histological findings. RESULTS: Histological grading and staging of disease increased as the age of patients increased. Steatosis was present in 70% of our patients and was related to a higher degree of fibrosis and to decreased functional activity. The prevalence of bile duct damage was 20%. This lesion was present in older patients with higher staging and impaired liver function. Biochemically it was associated with an increase in aspartate aminotransferase, gammaglutamyltranspeptidase, alkaline phosphatase, and total bilirubin. CONCLUSIONS: In the population we studied, HCV chronic hepatitis was predominantly a mild disease. Moreover both steatosis and bile duct damage were also mild. Steatosis was associated with fibrosis and this might influence liver metabolic function. Bile duct lesions were found in older patients with advanced disease showing biochemical evidence ofcholestasis. The molecular role HCV might play in the pathogenesis of these histological features should be addressed in further studies. PMID- 10533804 TI - Interferon treatment of two patients with quadruple infection with hepatitis C virus (HCV), human immunodeficiency virus (HIV), hepatitis G virus (HGV), and TT virus (TTV). AB - We administered interferon (IFN) to two patients who had quadruple infection with hepatitis C virus (HCV), human immunodeficiency virus (HIV), hepatitis G virus (HGV), and TT virus (TTV), a recently isolated novel DNA virus. Nine mega-units of natural alpha-IFN were administered daily during the first two weeks and thrice weekly during the following 22 weeks (total dose, 720 mega-units). In both cases, serum alanine aminotransferase (ALT) levels decreased during IFN administration but increased thereafter. The concentrations of HCV, HIV, HGV, and TTV declined with the administration of IFN. However, the concentrations of these 4 viruses increased after the cessation of IFN with the except of TTV in patient 2 which disappeared during treatment and did not subsequently reappear. IFN reduced the concentrations of 4 viruses, in an apparently independent manner. PMID- 10533805 TI - Imbalance between cell proliferation and cellular DNA fragmentation in hepatocellular carcinoma. AB - AIM/BACKGROUND: Hepatocellular carcinoma (HCC) is known for its rapid growth. This study was undertaken to determine the expression of proliferative markers, apoptosis (DNA fragmentation) and oncogene products known to regulate apoptosis (p53, bcl-2) in HCC. METHODS: 150 Chinese patients with HCC were studied (M:F 128:22, age 14-88 years). Immunohistochemistry was employed to detect cell proliferative markers (PCNA, Ki67), and oncogene products known to regulate apoptosis (p53, bcl-2). DNA fragmentation was determined by terminal dUTP nick end labeling (TUNEL). RESULTS: 98% and 95% of HCC had PCNA (median 2+) and Ki67 (median 2+) detected respectively. TUNEL labeling was detected in only a small number of tumor cells (no labeling in 11%, median 1/1000 cell labeled, range: 0 70/1000 cells). There was no correlation between TUNEL labeling and the clinical parameters (sex, age, cirrhosis, and survival) and the expression of cell proliferative markers. p53 was detected in 53% of the patients (median 1+, range: 0-4+) and bcl-2 was detected in a small proportion of tumor cells in only 13% of the HCCs (range: 0-1 +). The expression of p53 and Bcl-2 did not correlate with TUNEL labeling or the natural survival. CONCLUSIONS: Cell proliferation in HCC is unmatched by apoptosis, accounting for the rapid growth of this tumor. This lack of apoptosis in HCC is unrelated to the expression of p53 or bcl-2 over expression. PMID- 10533806 TI - A new class of diacidic nonpeptide angiotensin II receptor antagonists: candesartan cilexetil. AB - Blockade of the action of angiotensin ii (AII) has long been a target for development of novel antihypertensive agents. We recently discovered a novel class of potent non-peptide AII receptor antagonists, benzimidazole-7-carboxylic acids including candesartan. Candesartan is a highly potent and insurmountable antagonist selective in the angiotensin II type-I receptor (AT1). Structure activity relationship (SAR) studies revealed that the adjacent arrangement of a lipophilic substituent, a tetrazolylbiphenylmethyl moiety and a carboxyl group was the important structural requirement for potent AII antagonistic activity. Especially, the presence of a carboxyl group at the 7-position was found to be essential for insurmountable antagonism. To improve bioavailability of candesartan, chemical modification was examined to yield candesartan cilexetil, a prodrug of candesartan. Candesartan cilexetil is a potent and long-acting blocker that, when given once-daily to patients, provides effective 24 hr blood pressure control. PMID- 10533807 TI - Three-dimensional quantitative structure-activity relationships (3D-QSAR) of antidiabetic thiazolidinediones. AB - Thiazolidine-2,4-diones (TZDs) are novel insulin resistance releasing compounds that serve as orally active antidiabetic agents. Many TZDs have been synthesized and evaluated by in vivo screening for over 20 years. Recently, TZDs have been found to be selective agonists of peroxisome proliferator-activated receptor-y (PPAR-gamm), which is believed to work in the regulation of insulin resistance. This paper reports our efforts for the pharmacophore modeling study through 3D (three-dimensional) structure-activity relationship of TZDs to gain an insight into their molecular mechanism as well as the relation between antihyperglycemic and PPAR-gamma agonistic activities. The modeling study was carried out by conformational analysis along with the Apex-3D QSAR method to identify molecular features common to a series of 7 selected TZDs. Although the number of compounds included in the study was rather low, the variations in the activities were nicely elucidated with 3D-site specific physiochemical parameters significantly. PMID- 10533808 TI - Structure-activity relationship of hydroxamate-based inhibitors on membrane-bound Fas ligand and TNF-alpha processing. AB - Both Fas ligand (FasL) and tumor necrosis factor-alpha (TNF-alpha) are type II integral membrane proteins. Recently, we have reported that FasL is processed to a soluble form by an unknown metalloproteinase at the cell surface and some hydroxamate matrix metalloproteinase (MMP) inhibitors inhibit the processing similar to the case observed with TNF-1alpha. We studied the inhibitory effects of various hydroxamate MMP inhibitors on FasL and TNF-alpha processing in order to characterize the processing enzymes using human FasL cDNA transfectants and LPS-stimulated THP-1 cells. It turned out that (1) the P1' group of hydroxamates was very important for the selective inhibitory activity toward TNF-alpha and FasL processing, (2) P1' 3-phenylpropyl group was favorable for the inhibition of FasL processing, and (3) P1' isobutyl and isopropyl groups were favorable for that of TNF-alpha processing. These differences in sensitivity to inhibitors imply that (1) membrane-bound FasL and TNF-alpha might be processed by distinct metalloproteinases, (2) the S1' site of FasL processing enzyme differs from that of MMP-1 and MMP-9, but appears to be similar to that of MMP-3, and (3) the S1' site of TNF-alpha processing enzyme is smaller than that of FasL processing enzyme. These results would be helpful in designing a more selective inhibitor. PMID- 10533809 TI - The role of water molecules in the deacylation of acylated structures of class A beta-lactamase. AB - Molecular dynamics simulation of the penicillin- and penem-acylated enzymes reveals that the conformational flexibility of the acyl moieties in the binding cleft and the conformational change of the acyl moieties are crucial for deacylation. The water molecule adjacent to the Glu 166 residue is not the nucleophile for deacylation, but construction of a model of the oxyanion tetrahedral intermediate suggested a plausible role of the water molecule as a proton donor for the oxyanion to facilitate the deacylation. PMID- 10533810 TI - Analysis of affinities of penicillins for a class C beta-lactamase by molecular dynamics simulations. AB - We present a calculation for the binding free energy difference between two complexes of the class C beta-lactamase from Enterobacter cloacae with foramidocillin (FOPC) and with piperacillin (PIPC). The calculation was carried out by means of the thermodynamic integration (TI) method implemented with molecular dynamics (MD). By use of the available crystal structure of the class C beta-lactamase from E. cloacae, the structures of the beta-lactamase-FOPC (FOPC complex) and beta-lactamase-PIPC (PIPC complex) complexes were built by molecular modeling and equilibrated with MD simulations. FOPC were gradually converted into PIPC in both the solution and the enzyme system by means of MD/TI methods during the MD simulation. The structure of the PIPC complex as derived by the MD/TI simulation was similar to that of the PIPC complex obtained from molecular modeling. The calculated difference in the free energy of binding (deltadeltaGbind) was -2.2 kcal/mol. This compares well with the experimental value of -1.5 kcal/mol. The results indicate that the binding affinity of FOPC is lower than that of PIPC because of the greater difficulty of desolvation for FOPC upon binding to the enzyme. This calculation suggests that the desolvation of the ligand, as well as its interaction with the beta-lactamase, is important in understanding the relative affinity of the ligands with beta-lactamase. PMID- 10533811 TI - 3D-pharmacophore analyses of aldose reductase inhibitory spiroquinazolinones. AB - In order to get an insight for designing novel inhibitors of aldose reductase, we analyzed relationships between structures of spiroquinazolinones and their inhibitory activities against rabbit aldose reductase by comparative molecular field analysis and molecular modeling of the enzyme-inhibitor complex. It was revealed that the following interactions were operative for the enhancement of inhibitory activity; 1) the hydrophobic interaction between substituents at the 6'- and 7'-position of quinazolinone and the hydrophobic residues such as Trp20, Val47, Tyr48, Tyr121 and Phe122; 2) the electrostatic interaction formed between electronegative substituents at the 6'-position and the side chain of Gln49; 3) the complementary fit of sterically small 6'-substituents to the active site. PMID- 10533812 TI - Syntheses and SH-enzyme inhibitory activities of new epoxysuccinic acid piperazine derivatives against mu-calpain and cathepsin B. AB - New chiral epoxysuccinic acid derivatives 5 approximately 23 bearing various amino acids and N-substituted piperazines were synthesized to evaluate their inhibitory activities against mu-calpain and cathepsin B. After screening these compounds, 1-[(2S,3S)-epoxysuccinyl-L-leucyl]-4-(2-chlorophenyl)piperazine 9 proved to exhibit fairly strong inhibitory activity against both cysteine proteases. L-Valyl derivative 19 exhibited selective inhibitory activity against cathepsin B in comparison with that against mu-calpain. PMID- 10533813 TI - Synthesis and pharmacological activity of pyrazolopyrrolopyrimidine derivatives having vasorelaxing activity. AB - A new series of 5-substituted and 5-nonsubstituted pyrazolopyrrolopyrimidine derivatives were synthesized, and their vasorelaxing and hypotensive activities were evaluated. The syntheses were efficiently accomplished through the use of three key intermediates (7, 16, and 24), as shown in Schemes I-III. The desired pharmacological activities were confirmed on the basis of vasorelaxing activity in rat aorta (in vitro) and hypotensive activity in rats (in vivo). Specifically, compound 25 exhibited the strongest activity and appears to be a promising clinical lead for the development of a new antihypertensive agent. PMID- 10533814 TI - The future of transfusion medicine. PMID- 10533815 TI - New variant Creutzfeldt-Jakob disease and white cell reduction: risk assessment and decision making in the absence of data. PMID- 10533816 TI - The cost-effectiveness of reducing donor exposures with single-donor versus pooled random-donor platelets. AB - BACKGROUND: Single-donor platelets (SDPs) are frequently preferred over pooled random-donor platelets (RDPs) to reduce donor exposures and the risk for virus transmission or HLA alloimmunization. Transfusion-associated virus-transmission risks have significantly decreased, which suggests that white cell reduction by filtration eliminates any difference in the risk of alloimmunization in transfused leukemic patients. Health care reform pressures of make it appropriate to examine the cost-effectiveness of SDPs versus RDPs in reducing donor exposures. STUDY DESIGN AND METHODS: A decision analysis model was developed and sensitivity analyses were used to assess the incremental cost (dollars/quality adjusted life-year) associated with the use of SDPs versus RDPs for adult patients undergoing hematopoietic progenitor cell transplantation or primary coronary artery bypass grafting (CABG). RESULTS: Among transplant patients, the incremental cost of choosing SDPs as opposed to RDPs ranged from $168,700 to $519,822 per quality-adjusted life-year. For patients undergoing primary CABG, the incremental cost was $192,415 (females) and $216,280 (males). Variations in the cost differential between SDPs and RDPs, the number of random-donor platelets in the RDP, and the risk of bacterial sepsis markedly influenced cost effectiveness. The model was minimally affected by variations in the risks of transmission of HIV and hepatitis B and C, and human T-lymphotropic viruses. CONCLUSION: In comparison with other accepted medical interventions, the use of SDPs as opposed to RDPs may not be a cost-effective method of reducing donor exposures in the adult patient populations studied. SDPs were more cost-effective in patients undergoing primary CABG than in leukemia patients undergoing hematopoietic progenitor cell transplantation. Regardless of diagnosis, decreasing the acquisition cost differential would have the greatest impact on improving the cost-effectiveness of SDPs, as opposed to RDPs, to decrease donor exposures. PMID- 10533817 TI - Erythropoietin and preoperative autologous blood donation in the prevention of hepatitis C infection: necessity or luxury? AB - BACKGROUND: Prevention of exposure to allogeneic blood transfusion during surgery is an important financial issue when recombinant human erythropoietin (rHuEPO) is used in addition to preoperative blood donation. STUDY DESIGN AND METHODS: The aim of this study was to carry out a cost-effectiveness analysis of the use of rHuEPO in preoperative blood donation in orthopedic surgery. The study, based on a decision tree analysis of the use of rHuEPO, was conducted from the perspective of the French health care system. The efficacy criterion was the number of hepatitis C infections prevented. The decision tree analysis was constructed as follows: the residual risk of hepatitis C infection was 8.26 per million units transfused, and the chance node was defined according to the number of units transfused. RESULTS: With the use of rHuEPO in preoperative blood donation, 0.30562 cases of hepatitis C infection per 100,000 patients were prevented. The incremental cost of one prevented hepatitis C infection amounted to $888,000,000 (US). CONCLUSION: Despite the limitations of our model, the cost-effectiveness ratio was so large that variations only slightly modified the size of the result. From the societal perspective, it was not cost-effective to add rHuEPO to preoperative blood donation. PMID- 10533818 TI - Cost-effectiveness of a limited-donor blood program for neonatal red cell transfusions. AB - BACKGROUND: Very-low-birthweight infants have typically been given fresh red cells (RBCs), a practice in which aliquots of RBCs for several infants were issued each day from a single unit. Recently, to limit donor exposures, large volumes of RBCs are reserved for the long-term transfusion support of individual infants. STUDY DESIGN AND METHODS: Medical records were examined retrospectively to assess the costs of a limited-donor program for providing RBC transfusions to very-low-birthweight infants. Costs of multiple- and limited-donor programs were compared by using two samples of 30 consecutive infants treated at The University of Iowa Hospitals and Clinics in 1993 and 1997. Effectiveness was evaluated with respect to the number of donor exposures per infant. RESULTS: The cost, in 1997 dollars, of preparing each small-volume transfusion in the multiple-donor program was $27.86 per transfusion, while that in the limited-donor program was $34.83. This difference was largely attributable to use of white cell reduction in association with the limited-donor program in 1997. Eliminating the costs associated with white cell reduction rendered the costs of the limited- and multiple-donor transfusions comparable. The limited-donor program had donor exposures of 2.0 per infant, while the multiple-donor program had 3.6 exposures per infant (p<0.002). CONCLUSION: The limited-donor blood program reduces donor exposure without adversely affecting costs. PMID- 10533819 TI - Transfusion-related acute lung injury due to granulocyte-agglutinating antibody in a patient with paroxysmal nocturnal hemoglobinuria. AB - BACKGROUND: Transfusion-related acute lung injury (TRALI) is usually reported after the transfusion of blood components from donors with white cell (WBC) antibodies, but only very rarely if the patient has these antibodies. The pathogenesis of TRALI is not fully understood. Not all transfusion recipients develop TRALI, even though WBC antibodies are present in the donor or the recipient. CASE REPORT: A patient with paroxysmal nocturnal hemoglobinuria (PNH) who developed TRALI after the transfusion of non-WBC-reduced red cells is described. Granulocyte-agglutinating anti-5b was detected in his serum, and the crossmatch with the donor granulocytes was positive. The patient also developed a severe exacerbation of hemolysis with renal failure; serologic results excluded an immune hemolytic posttransfusion reaction. The patient recovered from both events after about 1 week. CONCLUSION: Granulocyte-agglutinating antibodies present in the recipient play an important role in TRALI, and also other factors may contribute to its pathogenesis. The reaction between the PNH patient's antibody (anti-5b) and transfused WBCs was found not only to be responsible for the respiratory distress but also to have triggered, through the innocent bystander mechanism of complement activation, an intensive hemolysis, which was very likely a contributing factor in the development of TRALI. PMID- 10533820 TI - Hemolytic transfusion reaction due to anti-Tc(a). AB - BACKGROUND: Anti-Tc(a) detects a high-incidence antigen in the Cromer blood group system. Cromer system antibodies have not usually been associated with hemolytic transfusion reactions or hemolytic disease of the newborn. CASE REPORT: Anti Tc(a) (initially identified in the patient's serum in 1982) was not detected when she was admitted to the hospital with upper gastrointestinal. bleeding. Three units of red cells were administered. The patient was discharged, but was readmitted to the hospital after her hemoglobin fell to 7.1 g per dL. Antibody detection tests remained negative and three additional units were transfused. Over the next 7 days, her hemoglobin steadily fell to 5.5 g per dL. The level of lactate dehydrogenase rose to 1257, the plasma hemoglobin rose to >16 mg per dL, and the haptoglobin decreased to <6 mg per dL. Five days after transfusion, her direct antiglobulin test was weakly reactive with complement-specific antiglobulin reagents. Eluates were nonreactive. Anti-Tc(a) was detected in her serum; no other antibodies were detected. Differential typing failed to detect any circulating Tc(a+) red cells. The antibody was strongly reactive in a monocyte monolayer assay. CONCLUSION: Although Cromer system antibodies have generally not been proven to be clinically significant in transfusion therapy, the destruction of red cells from six units of transfused Tc(a+) red cells in this patient indicates that anti-Tc(a) may have destructive potential in some patients. PMID- 10533821 TI - Platelet concentrates prepared and stored under currently optimal conditions: minor impact on platelet adhesive and cohesive functions after storage. AB - BACKGROUND: The effect on platelets of two standard methods of platelet concentrate (PC) preparation was studied by flow cytometry. The findings were correlated with those obtained in an experimental in vitro perfusion model. STUDY DESIGN AND METHODS: PCs were prepared from whole blood by the platelet-rich plasma (PRP) or buffy coat (BC) method and placed on a flatbed platelet agitator at 22 degrees C for up to 5 days. Platelet glycoproteins (GP)lbalpha, GPIIb/IIIa, and GPIV, p-selectin and lysosomal integral membrane protein, and the binding of von Willebrand factor, fibrinogen, fibronectin, and coagulation factor Va were measured with the corresponding specific conjugated antibodies. Perfusions were carried out in an annular chamber with citrated blood depleted of platelets and white cells by filtration, to which samples from PCs were added. RESULTS: PRP-PC production provoked intense platelet activation. In contrast, in BC-derived PCs, platelet activation was milder, and only a significant increase in bound fibrinogen was seen. After 1 day of storage, differences between the methods that had been observed immediately after separation had almost disappeared. During the remaining storage period, increases in activation-dependent antigens and in procoagulant activity were measured. Of the studied platelet GPs, only GPIIIb/ IIIa decreased by 25 percent in PRP-PCs. Differences in covered surface were not significant in perfusion studies performed on Day 0 and after 5 days of storage in PRP-PCs (26.8 +/- 6.9 vs. 20.5 +/- 5.8) or BC-PCs (23.8 +/- 11 vs. 24.8 +/- 10.2). CONCLUSION: Platelet activation occurred during the separation and storage of PCs prepared by both methods, and it was higher in PRP-PCs only in samples obtained immediately after preparation. Despite these changes, platelet adhesive and cohesive functions were similar in both types of PCs and remained basically unchanged after storage. PMID- 10533822 TI - Quality and clinical response to transfusion of prestorage white cell-reduced apheresis platelets prepared by use of an in-line white cell-reduction system. AB - BACKGROUND: This study evaluated the quality and clinical effectiveness of white cell (WBC)-reduced apheresis platelets collected by the use of a new technology, fluidized particle-bed separation. STUDY DESIGN AND METHODS: In phase 1, six suitable donors underwent two separate plateletpheresis procedures on one occasion, each separated by less than 10 minutes. In random order, a control unit was collected with the COBE Spectra and a test unit with the Spectra Leukocyte Reduction System (LRS). The quality of apheresis platelet components was assessed by an in vitro test panel, and residual WBCs were counted by Nageotte chamber and flow cytometric methods. For the in vivo studies, the test and control units were randomly labeled with either 51Cr or 111In at the end of storage and transfused simultaneously to the donor. Samples were taken for calculation of platelet survival and recovery. In phase II, 109 thrombocytopenic patients were given platelets collected by use of the Spectra LRS. RESULTS: Test platelets had significantly fewer residual WBCs (median 7.6 x 10(4)) than control platelets (median 3.9 x 10(5)), with equivalent in vitro function values. Test and control platelets had similar recovery and survival. Transfused platelets collected by use of the LRS achieved a mean 1-hour corrected-count increment of 19.3. CONCLUSION: The LRS collects platelet components with significantly lower WBC contamination without adverse effects on the function or in vivo survival of the platelets. PMID- 10533823 TI - Reduction of blood loss by infusion of human platelets in a rabbit kidney injury model. AB - BACKGROUND: As a first step toward testing the efficacy of stored platelets or platelet substitutes in vivo, a kidney injury model was developed to assess the hemostatic properties of human platelets in normal and thrombocytopenic rabbits. STUDY DESIGN AND METHODS: New Zealand white rabbits were made thrombocytopenic by two consecutive injections of busulfan. Two weeks later, human platelets were transfused to animals whose reticuloendothelial systems were inhibited by the administration of ethyl palmitate. The left kidney was exposed and a slice excised from the anterior pole. The blood was contained in a parafilm boat and absorbed by preweighed gauze to assess blood loss. The percentage of human platelets transfused to the rabbit was determined by flow cytometry on blood collected from the cut site using anti-CD42a (marker for human platelets). The degree of activation of the human platelets was determined using anti-CD62a (marker specific for human p-selectin). RESULTS: Blood loss was similar in normal animals treated with saline alone (35.4 +/- 5.8 g; n = 4); ethyl palmitate and saline (42.5 +/- 5.7 g; n = 6, p = 0.4); or ethyl palmitate and fresh human platelets (45.7 +/- 7.9 g; n = 6, p = 0.3). Bleeding in thrombocytopenic rabbits infused with saline was increased (75.6 +/- 3.9 g; n = 7) as compared with nonthrombocytopenic animals. A significant reduction in blood loss was noted in thrombocytopenic rabbits given fresh human platelets (51.6 +/- 4.5 g; n = 6, p = 0.0023). Transfusion of human platelets to rabbits did not cause activation of the platelets. Furthermore, transfusion of thrombin-activated platelets (60-98% activated) to thrombocytopenic rabbits reduced blood loss (54 +/- 7.3 g; n = 7) to the same extent as fresh platelets. CONCLUSIONS: This is the first report describing a kidney injury model developed to assess the efficacy of fresh and activated human platelets in reducing blood loss in thrombocytopenic rabbits. This model could monitor the efficacy of human platelets prepared by various preservation protocols in suppressing bleeding in rabbits. PMID- 10533824 TI - Coexpression of tissue factor and tissue factor pathway inhibitor by human monocytes purified by leukapheresis and elutriation. Response of nonadherent cells to lipopolysaccharide. AB - BACKGROUND: Counterflow centrifugal elutriation is the method of choice for obtaining a large quantity of highly purified monocytes. In spite of the fact that this technique has been used for many years to isolate monocytes for cellular immunotherapy, it is not known whether the process of elutriation can stimulate tissue factor (TF) expression and therefore trigger coagulation in patients receiving these cell preparations. The aim of the present study is thus to identify TF and TF pathway inhibitor (TFPI) in elutriated monocytes and to evaluate their ability to trigger thrombin generation. STUDY DESIGN AND METHODS: Human monocytes are separated by leukapheresis and elutriation in sterile, endotoxin-free conditions. TF and TFPI mRNA is detected by reverse transcription polymerase chain reaction. TF and TFPI are measured by enzyme-linked immunosorbent assay in cell lysates. TF antigen expression on cell surface is evidenced by direct-flow cytometry. Two functional tests (a chronometric test and an amidolytic assay) assess the capacity of monocytes to trigger thrombin generation. The response to lipopolysaccharide (LPS) is evaluated with each technique. Monocytic cell line THP-1 is used as a positive control. RESULTS: Elutriated monocytes coexpress TF mRNA and TFPI mRNA. The expression of TF mRNA is dramatically increased by LPS activation. This is correlated with a 100-fold increase in the amount of TF antigen in monocyte lysates. Flow immunocytometry confirms the expression of TF antigen on cell membrane in response to LPS stimulation, whereas TFPI mRNA is slightly increased after LPS stimulation. The amount of TFPI antigen in cell lysates is small when compared to that in plasma. Elutriated monocytes have a strong potential to trigger thrombin generation in response to LPS. CONCLUSION: In spite of the coexpression of TF mRNA and TFPI mRNA, elutriated monocytes are capable of supporting prothrombinase activity. This should be taken into account in the evaluation of the safety of adoptive cellular immunotherapy. PMID- 10533825 TI - Granulocyte storage and antigen stability. AB - BACKGROUND: Current methods for the detection of granulocyte antibodies require panels of freshly isolated cells. This makes these assays time-consuming, costly, and technically difficult. STUDY DESIGN AND METHODS: The immunofluorescence method of detecting the binding of antibodies to granulocytes was modified for use with a flow cytometer, and methods were tested to store granulocytes for use in that assay. Granulocytes were stored at 4 degrees C for 7 days under three conditions: 1 -percent formaldehyde-fixed cells were stored in Hanks' balanced salt solution (HBSS); untreated cells were stored in tissue culture medium (RPMI 1640); and cells were fixed and stored with a commercial white cell-storage solution (Cyto-Chex Reagent). Antigen stability was evaluated by using monoclonal antibodies (MoAbs) and alloantibodies. Serologic studies were done by an indirect immunofluorescence assay and assessed by flow cytometric analysis. RESULTS: On Day 2, only 2 to 7 percent of granulocytes stored in RPMI-1640 remained. On Day 7, 67 to 76 percent of granulocytes fixed in formaldehyde and stored in HBSS remained, and 47 to 87 percent of granulocytes stored in a white cell-storage solution remained. All antigens were detectable by the MoAbs and alloantisera on Day 7. However, nonspecific staining by the fluorescein isothiocyanate (FITC) conjugated secondary antibody hindered interpretation of test results on Day 4. Non-specific staining occurred over time and was associated with increased cell permeability during storage. Two sources of nonspecific staining were identified. The first source was the FITC-conjugated secondary antibody; it was eliminated by switching to a phycoerythrin conjugate. The second source was factors in human serum; it was resolved by examining only viable, impermeable cells identified by using 7-aminoactinomycin-D. CONCLUSION: Granulocytes and their antigens can be preserved for at least 7 days, but evaluation of antibody reactions was possible for only 4 days as a result of non-specific staining due to enhanced membrane permeability of dying cells. PMID- 10533826 TI - The viability of autologous human red cells stored in additive solution 5 and exposed to 25 degrees C for 24 hours. AB - BACKGROUND: No data exist on the viability of red cells (RBCs) stored in modern additive solution systems and allowed to warm above 10 degrees C. STUDY DESIGN AND METHODS: In a randomized crossover study, 3 units of blood were collected at least 8 weeks apart from 11 volunteer donors and stored in additive solution 5 (AS-5). Of 3 units from each volunteer, 1 was stored for 6 weeks at 4 degrees C, 1 for 5 weeks at 4 degrees C except for 24 hours at 25 degrees C on Day 14, and 1 for 5 weeks at 4 degrees C except for 24 hours at 25 degrees C on Day 28. Units were sampled periodically during storage; at the end of storage, viability was measured by the 99mTc/51 Cr double-label method. RESULTS: RBC viability was not significantly different in the storage protocols. Less than 1 percent of stored cells hemolyzed. RBC ATP concentrations at the end of storage correlated with viability and were approximately equal in the warmed units after 30 days' storage and the conventionally stored units after 42 days. CONCLUSIONS: The data suggest that RBCs stored in AS-5 and allowed to warm to 25 degrees C for 24 hours lose about 12 days of their shelf life. PMID- 10533827 TI - Comparison of Adsol and CPDA-1 blood preservatives during simulated massive resuscitation after hemorrhage in swine. AB - BACKGROUND: In recent years, there has been a change from the use of blood stored in CPDA-1 to the use of red cells (RBCs) stored in electrolyte mixtures, such as Adsol (AS-1 RBCs). However, because Adsol contains mannitol, as well as increased amounts of glucose relative to CPD and CPDA-1, concerns have been expressed as to possible harmful effects (recipient hyperglycemia, inappropriate osmotic diuresis) that it might induce under conditions of massive RBC transfusion. STUDY DESIGN AND METHODS: A hemorrhagic shock animal model was used to evaluate the effects of large-volume infusion of CPDA-1 or Adsol on glucose homeostasis and on urinary output under conditions that were devoid of extensive surgical manipulation. Hemorrhage was induced in 10 female Pitman-Moore mini-pigs to maintain mean arterial blood pressure at 55 mmHg for 90 minutes. After the return of autologous RBCs plus 1 L of 0.9-percent sodium chloride, the animals were given solution equivalent to the solute load in either 20 units of CPDA-1 whole blood (63 mL x 20 = 1260 mL) or 20 units of AS-1 RBCs (100 mL x 20 = 2000 mL) over a period of 90 minutes. Animals were monitored to determine physiologic and blood chemical responses to infusion of the solutions and to determine if there was hyperglycemia or inappropriate diuresis in the Adsol-treated group. RESULTS: Animals that received CPDA-1 developed significant hypocalcemia, arterial hypotension, and elevated blood glucose concentrations; two of five animals died of circulatory collapse. In contrast, glucose metabolism in the Adsol recipients was well-regulated, serum ionized calcium concentration was not significantly altered, and all animals survived. No evidence of inappropriate diuresis was observed. CONCLUSION: Administration of large amounts of Adsol was not associated with hyperglycemia or inappropriate osmotic duiresis in hemorrhaged and resuscitated minipigs. These data suggest that fewer physiologic changes may be associated with the massive transfusion of AS-1 RBCs than with that of CPDA-1 whole blood. PMID- 10533829 TI - Molecular analysis of secretor type alpha(1,2)-fucosyltransferase gene mutations in the Chinese and Thai populations. AB - BACKGROUND: The human Lewis histo-blood group system belongs to a family of structurally related oligosaccharides. The mutations of fucosyltransferase genes alpha(1,2)-fucosyltransferase (FUT2 or Se) and alpha(1,3/1,4)-fucosyltransferase (FUT3 or Le), are responsible for the polymorphism of Lewis blood group phenotypes. However, a population study of the FUT2 mutation in Chinese and Thais has not yet been done, and there is some controversy about the phenotypes of Le(a+b+) and Le(a+b-). STUDY DESIGN AND METHODS: One hundred twentyfour Chinese and 70 Thais were phenotyped for Lea and Le(b). DNA samples were studied by polymerase chain reaction and then by a restriction enzyme digestion method to distinguish wild-type and six known mutations. Direct sequencing was done for controls and some uncertain cases. RESULTS: A new mutation, C302T mutation, was found in 2 of 136 chromosomes in the Thai population; none were discovered in Chinese. The frequencies of the normal and six mutant alleles among Chinese and Thais, respectively, were as follows: 134 (54.0%) of 248 and 58 (41.4%) of 140 were wild-type (Se); 0 of 248 and 2 of 140 (both 1.4%) had the G428A mutation; 120 (48.4%) of 248 and 75 (53.6%) of 140 had the A385T mutation; 2 (0.81%) of 248 and 0 of 140 had the C571T mutation; and 1 (0.4%) of 248 and 3 (2.2%) of 140 had the G849A mutation. Only 1 Chinese (0.4%) of 248 had the C628T mutation, and none had fusion gene mutation. CONCLUSION: The FUT2 genes encoding for the phenotypes Le(a+b+) and Le(a+b-) are the same. The function and character of the mutant enzyme may play an important role in the phenotype. The methods used in this study are clinically applicable in population studies of the FUT2 gene polymorphism to explore relationships among different ethnic groups and correlations between phenotype and genotype. PMID- 10533828 TI - Evaluation of a panel of human monoclonal antibodies to D and exploration of the synergistic effects of blending IgG1 and IgG3 antibodies on their in vitro biologic function. AB - BACKGROUND: The D immunoprophylaxis program has successfully reduced the incidence of Rh hemolytic disease of the newborn (HDN), but it has also reduced the availability of plasma-derived polyclonal anti-D, which constitutes the current therapeutic product. Human monoclonal anti-D from hybridoma cell lines may be an acceptable alternative, and clinical efficacy of each anti-D is being evaluated in several centers. STUDY DESIGN AND METHODS: This study represents the largest assessment (outside of the International Workshops) of human D monoclonal antibodies for potential therapeutic use. The in vitro biologic activity and immunologic and serologic reactivity of a coded panel of 20 D antibodies (THERAD) was investigated. The bioassays used were lymphocyte (K-cell) antibody-dependent cell-mediated cytotoxicity (ADCC), monocyte ADCC, and monocyte chemiluminescence, which together reflect the processes involved in antibody-coated red cell destruction in vivo. From this panel, six antibodies (THERADs 14, 19, 22, 23, 27, and 28, comprising 3 IgG1 and 3 IgG3 D monoclonal antibodies) were further selected to investigate the effects of blending in the three bioassays. RESULTS: Several THERAD blends displayed greater activity than their component parts, in the range of 6 to 124 percent. There was no evidence to suggest functional blocking effects with this restricted panel of antibodies. CONCLUSION: The THERAD blends containing both IgG1 and IgG3 anti-D appeared to be the most functionally active, as did blends containing antibodies to two distinct D epitopes. This in vitro evidence has important implications for the future formulation of an effective monoclonal preparation for the prevention of Rh HDN. PMID- 10533831 TI - Integrating the new generation of blood components into transfusion practice. PMID- 10533830 TI - Use of blood therapeutically drawn from hemochromatosis patients. Council on Scientific Affairs, American Medical Association. PMID- 10533832 TI - What is a transfusion medicine specialist? PMID- 10533833 TI - Immune tolerance in hemophiliacs: can we afford it? Is it worth it? Baxter Previously Untreated Patient Study Group. PMID- 10533834 TI - Absence of intrauterine transmission of TT virus. PMID- 10533835 TI - Inhibited and disinhibited temperament and autonomic stress reactivity. AB - We examined the relationship of temperament dimensions serving as markers for Gray's behavioral activation system (BAS) and behavioral inhibition system (BIS) with autonomic stress reactivity in 35 middle-aged men. Temperament was measured using the Strelau Temperament Inventory--Revised. Skin conductance responses and inter-beat interval were measured during administration of the Rorschach test. The results showed that temperamental activation was positively related to the task-level of and task-induced change in respiratory sinus arrhythmia (RSA) amplitude, but unrelated to heart rate (HR) reactivity. Temperamental inhibition was negatively associated with the task-level of electrodermal activity and task induced change in RSA amplitude, and positively associated with HR reactivity. The findings are in part contrary to the hypotheses presented in the literature. They also suggest that the temperamental inhibition-HR reactivity relationship is mediated by the parasympathetic nervous system. PMID- 10533836 TI - Anxiety and arousal: tests of a new six-system model. AB - The present experiment was a test of a new six-system model of anxiety, which includes physiological, behavioral, cognitive, affective, trait, and state components of anxiety and also differentiates between direct and mediated responses. The State-Trait Anxiety Inventory was used to screen 795 undergraduates at the University of Maryland. Of the 52 subjects chosen, half were high trait anxious and half low. The two groups were further divided into high and low situational stress conditions. Subjects in the high-stress condition were exposed to two types of stressful cognitive and two types of stressful affective tasks. Subjects in the low-stress condition were exposed to the same four tasks with the stressful aspect removed. Prior state anxiety and cognitive or affective sensitivity were also considered. It was found that the most influential factor in resultant arousal was situational stress. Trait anxiety, state anxiety, and cognitive vs. affective sensitivity also significantly influenced both direct and mediated physiological and subjective anxiety responses. In addition, rather than leading to increased arousal, as hypothesized, the presence of trait and state anxiety reduced arousal under certain conditions. PMID- 10533838 TI - Diurnal variations of tonic electrodermal activity. AB - Diurnal variability of skin conductance level (SCL) was examined in two complementary experiments, simultaneously with variability of skin temperature (ST) and that of simple reaction time (RT) which was recorded as a behavioural index of arousal. In Experiment I, 6 subjects spent 6 days in the laboratory in homogeneous conditions. Three recording sessions, each lasting 2 h, began, respectively, at 9:00 a.m. (morning), 1:00 p.m. (afternoon) and 5:00 p.m. (evening). Results indicated that SCL increased linearly throughout the day. Experiment II was undertaken to test whether this effect could still be observed in more heterogeneous conditions. Subjects (n = 12) attended to their own activities between the two 30-min sessions beginning, respectively, at 9:30 a.m. and 6:00 p.m. during a single experimental day. Again, SCL was higher in the evening than in the morning. In both experiments the SCL pattern seemed to be asynchronous with ST and RT variations. Taken as a whole, these data bring additional evidence of temporal electrodermal variation, a phenomenon which should be further taken into account in EDA research. PMID- 10533837 TI - Gender dependent EEG-changes during a mental rotation task. AB - This study is based on 30 students, 15 females and 15 males. EEG was recorded with 19 electrodes according to the international 10/20 system against averaged signals picked up from both ear lobes. Averaged power spectra and cross-power spectra between all electrode-positions were computed. Data were reduced to five frequency bands (theta, alpha1, alpha2, beta1 and beta2) and finally mean amplitude and coherence were computed. EEG-recordings were made during a mental rotation task (Shepard-figures). The obtained spectral parameters were compared with the corresponding values of a baseline activity with eyes open at rest. The results of statistical evaluations (paired Wilcoxon tests, Wilcoxon tests of independent samples) were presented in error probability maps. In males, local coherence decreased between the posterior left electrodes in the theta-band. In females, a decrease of coherence between the posterior right electrodes were observed in this frequency-band. In contrast to females, males showed an increase of coherence between frontal, central and parietal electrode positions in both hemispheres in the alpha1-band. The increase of local coherence in the beta1-band over the right temporo-parietal sites in the male group was more pronounced than in the female group. Especially in females, interhemispheric coherence increased between the posterior electrodes in the theta-, beta1- and beta2-ranges. This study suggests the involvement of many brain areas during mental rotation. Females show a rather symmetrical allocation of coherence increase in theta, beta1 and beta2. PMID- 10533839 TI - Slow brain potentials in a visual monitoring task. AB - The purpose of this study was to analyse slow brain potentials (SPs) during visual monitoring, needed to perform the clock-monitoring task (CMT). This task was successfully applied in behavioural studies, in which attention capabilities were investigated in dependence on individual factors. Clearly pronounced preparatory SPs were observed during the CMT reflecting a high level of preparation and attention, necessary for achieving high performance accuracy. The well-known influence of task repetition on SPs and the relation between SP and behaviour were confirmed. The practicability of the CMT, the clear and easily detectable SP components and the sensitivity of the present approach makes it recommendable for further applications, e.g. for the evaluation of the effects of occupational/environmental exposures on processes of information processing. PMID- 10533841 TI - Anxiety moderates cardiovascular responses to painful stimuli during sphygmomanometry. AB - Sphygmomanometry is the most common technique of blood pressure (BP) determination. We were interested in the role of anxiety as a predictor of BP changes induced by painful stimuli during sphygmomanometry. We studied 141 normotensive healthy subjects who were asked to complete a State-Trait-Anxiety Inventory (STAI) prior to the experiment. BP was determined continuously and non invasively using a Finapres device (Penaz-technique) and by arm sphygmomanometry. Five sphygmomanometric measurements took place, during the fourth the arm cuff was inflated to 300 mmHg (unpredictable to the subject), the others were done with a pressure of 175 mmHg. State and trait anxiety (STAI X1 and X2) correlated positively with diastolic BP changes during inflation of the arm cuff (state anxiety: r = 0.26, P < 0.05; trait anxiety: r = 0.20, P < 0.05). Our results suggest that anxiety may have an impact on cardiovascular responses following painful stimuli. PMID- 10533840 TI - Automatic attentional shifts induced by a noradrenergic drug in Alzheimer's disease: evidence from evoked potentials. AB - Prior research showed that attentional deficits are observed in Alzheimer's disease (AD). These deficits can further impair other cognitive processes. The present experiment was designed to study the shifts in attention induced by a noradrenergic drug (S 12024-2) through their electrophysiological correlates in 12 outpatients with mild AD, using an auditory oddball paradigm. The P3a component, known to be related to automatic attentional processing, was increased by the drug, whereas no changes occurred either in PN or in P3b, which are considered to reflect conscious processing. These results point to an involvement of the noradrenergic system in the modulation of automatic attentional processing, and provide evidence for weakening of the orienting reflex in AD, due to a possible noradrenergic deficit in patients with mild AD. PMID- 10533842 TI - Spatiotemporal organization of brain dynamics and intelligence: an EEG study in adolescents. AB - This study investigated relationships between global dynamics of brain electric activity and intelligence. EEG was recorded monopolarly from 10 symmetric leads (10-20 system) in 37 (17 males) healthy subjects (mean age 13.7 years) at rest and during performance of two visually presented cognitive tasks, verbal (semantic grouping) and spatial (mental rotation). On another occasion, the subjects were administered the Intelligence Structure Test (IST). Both total IST score and some individual subtests of specific abilities showed significant positive correlations with EEG coherence in the theta band and significant negative relationships with EEG dimension, a measure of complexity and unpredictability of neural oscillatory dynamics underlying the EEG time series. Furthermore, EEG coherence and dimensional complexity were inversely related. Taken together, these EEG metrics accounted for over 30% of the variability of the total IST score in this sample. No significant effects of the task type (spatial vs. verbal) or specific abilities were observed. Long-distance theta coherence between frontal and parieto-occipital areas showed the most consistent relationship with cognitive abilities. The results suggest that order to chaos ratio in task-related brain dynamics may be one of the biological factors underlying individual differences in cognitive abilities in adolescents. PMID- 10533843 TI - Retinoylation of proteins in rat hepatocytes following uptake of chylomicron remnant retinyl ester. AB - Several proteins may covalently bind retinoic acid, a process called retinoylation. Recently, we have demonstrated that proteins were retinoylated in vivo in liver, kidney and lung. In order to gain further knowledge about the mechanism of this process, we studied retinoylation in rat hepatocytes administered vitamin A as [3H]retinyl esters in chylomicron remnants. This resembles the normal physiological uptake of vitamin A. After 24 h incubation, about 0.0017 mol [3H]retinoid was covalently bound per mol protein. Citral, an inhibitor of the oxidation of retinol to retinoic acid, reduced retinoylation about 40%, indicating that oxidation of retinol to retinoic acid is necessary for a large fraction of the observed covalent modification of proteins. When cells were incubated with physiological concentrations of [3H]retinol or [3H]retinoic acid dissolved in ethanol, much less retinoid was covalently bound per mol protein compared with cells incubated with chylomicron remnant. Saturation of the retinoylation was apparent with retinoic acid around the physiological concentration. Retinoylated proteins were also analysed by SDS-PAGE. In general, the same protein bands were labelled with both [3H]retinol and [3H]retinoic acid, although the intensity of the bands varied. Major bands had an apparent molecular weight of about 16, 35, 50 and 120 kDa. In a parallel experiment in which liver stellate cells were incubated with [3H]retinol, major retinoylated protein bands were about 35, 60 and 65 kDa. Thus, different proteins appear to be retinoylated in hepatocytes and liver stellate cells, suggesting that protein retinoylation is a cell specific phenomenon. These results demonstrate that retinoids presented to hepatocytes as chylomicron remnant retinyl esters are covalently linked to proteins. We therefore suggest that retinoylation of proteins represents a minor but significant pathway whereby cells metabolize vitamin A. PMID- 10533844 TI - Plasma levels of cyclic GMP and endothelin in postmenopausal women with unstable coronary artery disease. AB - Many women with typical anginal chest pain have normal coronary angiograms, which may be due to altered endothelial function. We evaluated the endothelial markers cyclic GMP (cGMP) and immunoreactive endothelin (ir-ET) regarding presence of coronary atherosclerosis in women with clinical signs of unstable coronary artery disease (CAD). Plasma levels of cGMP and ir-ET were determined in 118 patients and 84 controls. Ischaemia was evaluated at an exercise test. Of the patients 20% had normal vessels, 14% insignificant CAD and 66%, significant stenosis at coronary angiography. Mean (95% CI) concentration of cGMP (nmol/l) was higher in patients than in controls (5.05 (4.53; 5.58) vs. 3.79 (3.34; 4.23)). Separating patients according to daily intake of nitroglycerin, only patients with this medication had significantly higher cGMP level (5.73 (4.88; 6.58)), whereas the difference between those without (4.35 (3.76; 4.94)) and controls disappeared. Patients with ischaemia at exercise test had higher cGMP level than those without (6.01 (5.13; 6.88) vs. 4.30 (3.66; 4.94)), even after adjusting for nitroglycerin treatment. ir-ET (pmol/l) was lower in patients with normal vessels than patients with coronary atherosclerosis (0.83 (0.78; 0.88) vs. 0.98 (0.92; 1.04)) and than the control group (0.91 (0.87; 0.94)). The difference between the control group and patients with atherosclerosis was also significant. Patients with unstable CAD and long-term nitroglycerin treatment have increased cGMP level. Patients with exercise-induced ischaemia have higher cGMP level than those without, irrespective of nitroglycerin treatment, which may reflect a general compensatory mechanism. Patients with normal vessels have low level of ir-ET, indicating different mechanisms for ischaemia/angina in these patients compared with patients with atherosclerosis. PMID- 10533845 TI - Determination of coagulation factor VIII activity by a chromogenic substrate method on STA, an automated coagulation analyzer. AB - This study was aimed at evaluating the performance of a chromogenic factor VIII assay on STA, an automated coagulation analyzer. Additionally, a correlation study was conducted with an aPTT-based one-stage factor VIII clotting assay. Throughout the study the performance of the chromogenic assay was tested in two ranges of factor VIII activity: a high range with activity between 20% and 150% and a low range with activity below 20%. Inter-assay coefficient of variation ranged from 1.9% to 8.9% and intra-assay coefficient of variation from 0.5% to 11.4%, depending on factor VIII concentration. Day-to-day reproducibility was tested over a 5-day period; between-day imprecision ranged from 7.1% to 9.4%. The chromogenic factor VIII assay showed a good correlation with the clotting assay in both ranges. The correlation coefficients were 0.924 and 0.792 for the high and low range, respectively. A statistically significant difference in mean values was observed in the high range (p < 0.0001). Comparison between the chromogenic assay on STA versus on microplates showed a high correlation (0.991), which was highly significant (p < 0.0001). In conclusion, the chromogenic assay for factor VIII on STA shows good analytical performance. It correlates well with the one-stage factor VIII clotting assay, although significant differences between individual samples occur. Probably these are partly related to differences in measurement principle and standardization. Altogether, this precise and rapid assay is suitable for determination of factor VIII by an automated procedure. PMID- 10533846 TI - Serum hyaluronan as a marker of liver fibrosis in asymptomatic chronic viral hepatitis B. AB - Increased concentrations of serum hyaluronan, a polysaccharide widely distributed in the extracellular space, have been demonstrated in liver disease of various aetiologies and proposed as a useful marker of liver fibrosis. The aim of the present study was to evaluate the association of serum hyaluronan with the extent of hepatic fibrosis in asymptomatic cases of chronic hepatitis B viral infection. The study was conducted in a consecutive sample of 111 asymptomatic chronic carriers of hepatitis B surface antigen. Liver function tests, alcohol consumption and cigarette smoking were determined and, for 84 subjects, liver biopsy was performed and degrees of inflammation and fibrosis were scored. Hyaluronan was measured using a radiometric assay. Mean serum hyaluronan increased with increasing fibrosis score (from 22.2 +/- 4.8 to 50.6 +/- 12.7 microg/l, p = 0.058) or pathological severity (from 18.8 +/- 5.9 to 50.6 +/- 12.5 microg/l, p = 0.048), even after adjusting for the effect of age. No such correlation was found with portal inflammation. The study showed that, in asymptomatic chronic carriers of hepatitis B, serum hyaluronan concentration correlates with hepatic fibrosis, a known marker of disease prognosis. This finding supports the hypothesis that hyaluronan might be of use in assessing and monitoring time trends in liver disease, substituting for repeated biopsies. PMID- 10533847 TI - Principal component analysis of proton nuclear magnetic resonance spectra of lipoprotein fractions from patients with coronary heart disease and healthy subjects. AB - Blood plasma was drawn from 12 healthy subjects and 12 patients with coronary heart disease (CHD). The lipoproteins were fractionated by serial ultracentrifugation. The methyl and methylene regions of proton nuclear magnetic resonance (1 H NMR) spectra of the lipoproteins very low density lipoprotein (VLDL), low density lipoprotein (LDL) and high density lipoprotein (HDL) were analysed by principal component analysis (PCA). Grouping patterns in the score plots and the profiles of the principal components revealed several characteristics of the spectra. LDL subparticle size among the CHD group was consistently skewed against smaller, denser subparticles. This feature was independent of the concentration of LDL cholesterol. Analysis of the LDL spectra by soft independent modelling of class analogy (SIMCA) showed that none of the samples from the CHD group could be assigned as healthy subjects (p < 0.05). We also found that the samples from the healthy subjects were associated with a higher concentration of HDL cholesterol and larger VLDL subparticles. The approach presented, in which PCA is used in combination with NMR spectroscopy, might be implemented in clinical studies to give information about lipoprotein subparticle distribution and lipid content. PMID- 10533848 TI - Electrochemical detection of nitric oxide production in human polymorphonuclear neutrophil leukocytes. AB - The detection of nitric oxide (NO) release by human polymorphonuclear neutrophil leukocytes (PMNs) presents several difficulties, mainly due to concomitant production of O2- and H2O2, which could interfere with the measurements. A Nafion and nickel porphyrin-coated microelectrode was used to measure NO production in PMNs in vitro. It allowed detection of 6.3 +/- 1.9 nM NO in a PMN-containing system and was unaffected by added chemicals. Addition of the chemotactic oligopeptide f-met-leu-phe (fMLP; 100 nM) induced a NO release which reached a value of 71 +/- 30 pmol NO/10(6) PMN x ml(-1) 5 min after stimulation in the presence of SOD (150 U/ml). If SOD was omitted, the corresponding value was 36 +/ 20 pmol NO/10(6) PMN x ml(-1). Presence or absence of catalase did not alter the amount of NO measured. Addition of the NO-synthase inhibitor N(G)-monomethyl-L arginine (LNMMA; 1 mM) reduced the current by 82 +/- 20%. These results agree with the rate of NO production in human PMNs when measured spectrophotometrically using the NO-dependent oxidation of oxyhaemoglobin to methaemoglobin. The NO production in human PMN was dependent on fMLP concentrations, but independent of cell-concentrations of 0.5-3.5 x 10(6)/ml. This paper shows that a electrochemical method, e.g. Nafion and porphyrin-coated microelectrode, is suitable for studies of NO release from stimulated human PMNs. PMID- 10533849 TI - Comparison of Abbott IMx total homocysteine assay with a high pressure liquid chromatography method for the measurement of total homocysteine in plasma and serum from a Norwegian population. AB - In this study 189 samples were analysed for the measurement of homocysteine by the Abbott IMx homocysteine assay and an HPLC method. A strong correlation was obtained between the homocysteine measurements performed by the Abbott IMx homocysteine assay and the HPLC method (coefficient of correlation, r2 = 0.947, p < 0.0001). The plot of the difference for the homocysteine measurements between the two methods against the average of the two measurements resulted a mean difference of 0.80 +/- 6.66 (mean +/- 2SD), and 0.008 +/- 0.126 (mean +/- 2SD) for log converted values of homocysteine. The concentrations of homocysteine measured in all the samples by the two methods were not significantly different. However, the Abbott IMx homocysteine assay measured the concentrations of total homocysteine in hyperhomocysteinemia as significantly higher than the HPLC method (25.00 micromol/l vs. 23.12 micromol/l, p < 0.0001). More studies may be required to explore factors that may influence measurements of homocysteine by the Abbott IMx homocysteine assay and the HPLC method. PMID- 10533850 TI - Growth hormone response to the insulin tolerance and clonidine tests in type 1 diabetes. AB - The insulin tolerance test (ITT) is regarded the gold standard for assessing growth hormone (GH) release in adult patients with suspected GH deficiency. Some of these patients also have diabetes mellitus. There are contradictory reports regarding the GH response to ITT in type 1 diabetic patients with varying degrees of metabolic control. This is also true for the clonidine test. We studied ten patients with uncomplicated type 1 diabetes mellitus during periods of poor and improved metabolic control and compared them with ten healthy control subjects. The GH secretion was assessed by an ITT with an insulin infusion of 2.5 mU/kg/min and an intravenous clonidine test. The GH response to ITT was similar during poor and improved metabolic control (mean +/- SEM) (AUC 2327 +/- 616 vs. 2649 +/- 508 microg/l x min) and did not differ from the response in control subjects (AUC 2587 +/- 343 microg/1 x min). The clonidine test induced a significantly greater GH response during poor than during improved metabolic control in the diabetic patients (AUC 2598 +/- 492 vs. 1508 +/- 368 microg/l x min, p < 0.05); this response was greater than in the control subjects (670 +/- 226 x microg/l x min, p < 0.005 vs. improved metabolic control). Thus, the GH response to ITT is similar in diabetic patients with varying degrees of metabolic control and healthy subjects, while the GH response to the clonidine test is higher in the type 1 diabetic patients than in healthy controls. PMID- 10533851 TI - Analysis of tissues that reflect central nervous system disease by mass spectrometry. PMID- 10533852 TI - A nasty start for NICE. PMID- 10533853 TI - Genetically modified foods: "absurd" concern or welcome dialogue? PMID- 10533854 TI - Adequacy of methods for testing the safety of genetically modified foods. PMID- 10533855 TI - Does biocompatibility of dialysis membranes affect recovery of renal function and survival? PMID- 10533856 TI - Drug-resistant Pneumocystis carinii. PMID- 10533857 TI - The nurse practitioner in emergency medicine--valuable but undefined. PMID- 10533858 TI - Modification of attitudes to influence survival from breast cancer. PMID- 10533859 TI - Care of minor injuries by emergency nurse practitioners or junior doctors: a randomised controlled trial. AB - BACKGROUND: We aimed to assess the care and outcome of patients with minor injuries who were managed by a nurse practitioner or a junior doctor in our accident and emergency department. METHODS: 1453 eligible patients, over age 16 years, who presented at our department with minor injuries were randomly assigned care by a nurse practitioner (n=704) or by a junior doctor (n=749). Each patient was first assessed by the nurse practitioner or junior doctor who did a clinical assessment; the assessments were transcribed afterwards to maintain masked conditions. Patients were then assessed by an experienced accident and emergency physician (research registrar) who completed a research assessment, but took no part in the clinical management of the patient. A standard form was used to compare the clinical assessment of the nurse practitioner or junior doctor with the assessment of the research registrar. The primary outcome measure was the adequacy of care (history taking, examination of patient, interpretation of radiographs, treatment decision, advice, and follow-up). FINDINGS: Compared with the rigorous standard of the experienced accident and emergency research registrar, nurse practitioners and junior doctors made clinically important errors in 65 (9.2%) of 704 patients and in 80 (10.7%) of 749 patients, respectively. This difference was not significant. The nurse practitioners were better than junior doctors at recording medical history and fewer patients seen by a nurse practitioner had to seek unplanned follow-up advice about their injury. There were no significant differences between nurse practitioners and junior doctors in the accuracy of examination, adequacy of treatment, planned follow-up, or requests for radiography. Interpretation of radiographs was similar in the two groups. INTERPRETATION: Properly trained accident and emergency nurse practitioners, who work within agreed guidelines can provide care for patients with minor injuries that is equal or in some ways better than that provided by junior doctors. PMID- 10533860 TI - Effectiveness of therapeutic plasma exchange in the 1996 Lanarkshire Escherichia coli O157:H7 outbreak. AB - BACKGROUND: The largest number of adult cases of haemolytic uraemic syndrome (HUS)/thrombotic thrombocytopenic purpura (TTP) during an Escherichia coli O157 outbreak occurred in 1996 in central Scotland. Adults who develop HUS/TTP induced by E. coli O157 tend to be elderly and have a historical mortality rate of almost 90% when treated conservatively. Therefore the decision was made to treat adults who developed HUS/TTP during this outbreak with therapeutic plasma exchange (TPE). We report our outcome with this controversial treatment. METHODS: A case definition for HUS/TTP was developed at the beginning of the outbreak. All cases meeting this definition were considered for TPE. Information on demographics, clinical features, treatment and outcome of patients was obtained by retrospective case note review. FINDINGS: 22 adults developed HUS/TTP. They had a mean age of 71 years. 16 cases received TPE. Six cases had contraindications to TPE or died before the procedure could be done. Ten of the 22 (45%) adults with HUS/TTP died. Five of the 16 (31%) TPE-treated cases died, four of eight aged over 70 years compared with one of eight aged less than 70 years. Premorbid illness, neurological features, treatment with ciprofloxacin or prostacyclin, and the laboratory severity of HUS/TTP were not associated with death; the number of cases, however, was too small to allow statistical conclusion. INTERPRETATION: The mortality rate is high in adults who develop HUS/TTP induced by E. coli O157. TPE appears to be a promising treatment that was well tolerated in our elderly patients. A national register of adult cases of HUS/TTP induced by E. coli O157 should be established. PMID- 10533861 TI - Influence of psychological response on survival in breast cancer: a population based cohort study. AB - BACKGROUND: The psychological response to breast cancer, such as a fighting spirit or an attitude of helplessness and hopelessness toward the disease, has been suggested as a prognostic factor with an influence on survival. We have investigated the effect of psychological response on disease outcome in a large cohort of women with early-stage breast cancer. METHODS: 578 women with early stage breast cancer were enrolled in a prospective survival study. Psychological response was measured by the mental adjustment to cancer (MAC) scale, the Courtauld emotional control (CEC) scale, and the hospital anxiety and depression (HAD) scale 4-12 weeks and 12 months after diagnosis. The women were followed up for at least 5 years. Cox's proportional-hazards regression was used to obtain the hazard ratios for the measures of psychological response, with adjustment for known clinical factors associated with survival. FINDINGS: At 5 years, 395 women were alive and without relapse, 50 were alive with relapse, and 133 had died. There was a significantly increased risk of death from all causes by 5 years in women with a high score on the HAD scale category of depression (hazard ratio 3.59 [95% CI 1.39-9.24]). There was a significantly increased risk of relapse or death at 5 years in women with high scores on the helplessness and hopelessness category of the MAC scale compared with those with a low score in this category (1.55 [1.07-2.25]). There were no significant results found for the category of "fighting spirit". INTERPRETATION: For 5-year event-free survival a high helplessness/hopelessness score has a moderate but detrimental effect. A high score for depression is linked to a significantly reduced chance of survival; however, this result is based on a small number of patients and should be interpreted with caution. PMID- 10533862 TI - Haemodialysis-membrane biocompatibility and mortality of patients with dialysis dependent acute renal failure: a prospective randomised multicentre trial. International Multicentre Study Group. AB - BACKGROUND: There is controversy as to whether haemodialysis-membrane biocompatibility (ie, the potential to activate complement and neutrophils) influences mortality of patients with acute renal failure. We did a prospective randomised multicentre trial in patients with dialysis-dependent acute renal failure treated with two different types of low-flux membrane. METHODS: 180 patients with acute renal failure were randomly assigned bioincompatible Cuprophan (n=90) or polymethyl-methacrylate (n=90) membranes. The main outcome was survival 14 days after the end of therapy (treatment success). Odds ratios for survival were calculated and the two groups were compared by Fisher's exact test. Analyses were based on patients treated according to protocol (76 Cuprophan, 84 polymethyl methacrylate). FINDINGS: At the start of dialysis, the groups did not differ significantly in age, sex, severity of illness (as calculated by APACHE II scores), prevalence of oliguria, or biochemical measures of acute renal failure. 44 patients (58% [95% CI 46-69]) assigned Cuprophan membranes and 50 patients (60% [48-70]) assigned polymethyl-methacrylate membranes survived. The odds ratio for treatment failure on Cuprophan compared with polymethyl-methacrylate membranes was 1.07 (0.54-2.11; p=0.87). No difference between Cuprophan and polymethyl-methacrylate membranes was detected when the analysis was adjusted for age and APACHE II score. 18 patients in the Cuprophan group and 20 in the polymethyl-methacrylate group had clinical complications of therapy (mainly hypotension). INTERPRETATION: There were no differences in outcome for patients with dialysis-dependent acute renal failure between those treated with Cuprophan membranes and those treated with polymethyl methacrylate membranes. PMID- 10533863 TI - Mutations in the ABC1 gene in familial HDL deficiency with defective cholesterol efflux. AB - BACKGROUND: A low concentration of HDL cholesterol is the most common lipoprotein abnormality in patients with premature atherosclerosis. We have shown that Tangier disease, a rare and severe form of HDL deficiency characterised by a biochemical defect in cellular cholesterol efflux, is caused by mutations in the ATP-binding-cassette (ABC1) gene. This gene codes for the cholesterol-efflux regulatory protein (CERP). We investigated the presence of mutations in this gene in patients with familial HDL deficiency. METHODS: Three French-Canadian families and one Dutch family with familial HDL deficiency were studied. Fibroblasts from the proband of each family were defective in cellular cholesterol efflux. Genomic DNA of each proband was used for mutation detection with primers flanking each exon of the ABC1 gene, and for sequencing of the entire coding region of the gene. PCR and restriction-fragment length polymorphism assays specific to each mutation were used to investigate segregation of the mutation in each family, and to test for absence of the mutation in DNA from normal controls. FINDINGS: A different mutation was detected in ABC1 in each family studied. Each mutation either created a stop codon predicted to result in truncation of CERP, or altered a conserved aminoacid residue. Each mutation segregated with low concentrations of HDL-cholesterol in the family, and was not observed in more than 500 control chromosomes tested. INTERPRETATION: These data show that mutations in ABC1 are the major cause of familial HDL deficiency associated with defective cholesterol efflux, and that CERP has an essential role in the formation of HDL. Our findings highlight the potential of modulation of ABC1 as a new route for increasing HDL concentrations. PMID- 10533864 TI - Effects of mutations in Pneumocystis carinii dihydropteroate synthase gene on outcome of AIDS-associated P. carinii pneumonia. AB - BACKGROUND: Sulpha drugs are widely used for the treatment and long-term prophylaxis of Pneumocystis carinii pneumonia (PCP) in HIV-1-infected individuals. Sulpha resistance in many microorganisms is caused by point mutations in dihydropteroate synthase (DHPS), an enzyme that is essential for folate biosynthesis. We assessed whether mutations in the DHPS gene of P. carinii were associated with exposure to sulpha drugs and influenced outcome from PCP. METHODS: We studied bronchoalveolar samples collected in 1989-99 from a prospective cohort of HIV-1-infected patients who had PCP. In 144 patients with 152 episodes of PCP, we analysed portions of DHPS using PCR and direct sequencing. The relation between survival, P. carinii DHPS mutations, and other predictors of treatment failure was assessed by Kaplan-Meier and multivariate Cox regression analysis. FINDINGS: P. carinii DHPS mutations were found in 31 (20.4%) of 152 PCP episodes. 3-month survival was significantly lower in patients infected with mutant P. carinii DHPS strains than in those with wild-type strains (p=0.002). After adjustment for other prognostic variables, presence of DHPS mutations remained the most important predictor of mortality (hazard ratio 3.1 [95% CI 1.2-8.1]). DHPS mutations were significantly more common in patients who had previous exposure to sulpha drugs (18 of 29 [62%]) than in those who had no exposure (13 of 123 [10.5%]; p<0.0001). A significant increase with time in the rate of DHPS mutations (p=0.01 for trend) was closely correlated with the rate of previous or current use of sulpha drugs as chemoprophylaxis. INTERPRETATION: Mutations in DHPS are associated with impaired prognosis in PCP, and may develop as a result of exposure to sulpha drugs. PMID- 10533865 TI - Weight loss and purpura. PMID- 10533866 TI - Effect of diets containing genetically modified potatoes expressing Galanthus nivalis lectin on rat small intestine. AB - Diets containing genetically modified (GM) potatoes expressing the lectin Galanthus nivalis agglutinin (GNA) had variable effects on different parts of the rat gastrointestinal tract. Some effects, such as the proliferation of the gastric mucosa, were mainly due to the expression of the GNA transgene. However, other parts of the construct or the genetic transformation (or both) could also have contributed to the overall biological effects of the GNA-GM potatoes, particularly on the small intestine and caecum. PMID- 10533867 TI - Differential binding of the insecticidal lectin GNA to human blood cells. AB - Evidence of snowdrop lectin binding to human white cells supports the need for greater understanding of the possible health consequences of incorporating plant lectins into the food chain. PMID- 10533868 TI - Resurgence of mumps in Singapore caused by the Rubini mumps virus vaccine strain. AB - The measles, mumps, and rubella vaccine containing the highly attenuated Rubini mumps virus strain conferred no protection against acute parotitis in vaccinated children in Singapore. Its introduction into the national childhood immunisation programme has resulted in a reduction in the seroprevalence of mumps to prevaccination levels. PMID- 10533869 TI - Intracluster correlation of STD prevalence in a community intervention trial in Kenya. AB - This study is a cluster-randomised, community intervention trial to measure the impact of female condom introduction on STD prevalence among Kenyan agricultural workers. The intracluster correlation coefficient of baseline STD prevalences at the 12 sites was 0.0011. PMID- 10533870 TI - Differences in lung bioavailability between different propellants for fluticasone propionate. AB - The lung bioavailability (as adrenal suppression) of fluticasone propionate was about twofold greater with chlorofluorocarbons than hydrofluoroalkane as propellant. Direct switching between formulations on a microg equivalent may therefore be inadvisable. PMID- 10533871 TI - Are UK doctors particularly stressed? AB - Doctors are thought to have higher levels of stress than the general population. However, in a survey of a nationally representative sample of UK doctors, we found that stress levels were equivalent to those in the general population; previous findings may have been biased by inappropriate use of the general health questionnaire. PMID- 10533872 TI - Extent of antibiotic use in Taiwan shown by antimicrobial activity in urine. AB - Taiwan has high prevalence of antibiotic resistant, community-acquired respiratory pathogens. We investigated whether there was a high frequency of antibiotic use in the community. Antimicrobial activity in urine was detected in 55.2% of 1182 patients on arrival at an emergency department, 25.1% of 203 internal medicine out-patients, 7.6% of 471 high school students, and 7.4% of 202 people at a centre for senior citizens. PMID- 10533873 TI - International research agenda set for end-of-life care. PMID- 10533874 TI - Surgery patients at risk for herb-anaesthesia interactions. PMID- 10533875 TI - Gene-directed cancer therapy starts to find its way. PMID- 10533876 TI - Angola: a forgotten emergency. PMID- 10533877 TI - European Commission proposes new strategy against HIV/AIDS. PMID- 10533878 TI - Depressive illness. AB - WHO estimate that by the beginning of the next century major unipolar depression will be one of the most important causes of ill health overall. Whereas the cause of depression is still obscure, it is becoming clear that a number of diverse factors are likely to be implicated, both genetic and environmental. Effective treatment of depression similarly involves a variety of methods, from electro convulsive therapy to inter-personal psychotherapy. The pathophysiology of depression is gradually becoming accessible through research strategies, such as functional neuroimaging paired with mood altering interventions. PMID- 10533879 TI - Ether and the chemical-contact dissolution of gallstones. PMID- 10533881 TI - Genetically modified potatoes. PMID- 10533880 TI - Shifts in mortality curves: saving or extending lives? . PMID- 10533882 TI - Earthquake in Turkey. PMID- 10533883 TI - Earthquake in Turkey. PMID- 10533884 TI - Earthquake in Turkey. PMID- 10533885 TI - Intensive case management for severe psychotic illness. PMID- 10533886 TI - Intensive case management for severe psychotic illness. PMID- 10533887 TI - Intensive case management for severe psychotic illness. PMID- 10533889 TI - Mycophenolate mofetil in IBD patients. PMID- 10533888 TI - HTLV-I-associated infective dermatitis. PMID- 10533890 TI - Safety and efficacy of vigabatrin and carbamazepine. CPMP. The Committee for Proprietary Medicinal Products. PMID- 10533891 TI - Immunotherapy with Mycobacterium vaccae. PMID- 10533892 TI - Averting a malaria disaster. PMID- 10533893 TI - Rotavirus vaccine put on hold. PMID- 10533895 TI - Effect of simple interventions on necropsy rate when active informed consent is required. PMID- 10533894 TI - Smoking in pregnancy. PMID- 10533896 TI - Humanised maternity care. PMID- 10533897 TI - Depression, physical illness, and the faces of Rembrandt. PMID- 10533898 TI - The Nobel chronicles. 1976: Baruch S Blumberg (b 1925); Daniel Carleton Gajdusek (1923). PMID- 10533899 TI - Changes in resistive index following extracorporeal shock wave lithotripsy. AB - BACKGROUND: Extracorporeal shock wave lithotripsy (ESWL) has replaced most surgical and endourologic forms of therapy for upper urinary tract stone disease. Despite its proved safety and efficacy, its adverse effects on renal function are still to be identified. A newer diagnostic technique, color Doppler ultrasonography, has brought a new insight into renal function. It enables precise evaluation of the renal vascular supply. Changes in intrarenal vascular resistance after ESWL were studied with Doppler ultrasound techniques. METHODS: In 70 consecutive patients the resistive index (RI) was measured at an interlober artery before and 30 min after ESWL in the treated and contralateral kidneys. In 17 patients, a follow-up Doppler study was performed 1 week after ESWL. RESULTS: In the treated kidneys, the RI significantly increased from 0.656+/-0.053 (mean +/- SD) at baseline to 0.682+/-0.053 (P<0.0001). There was no significant correlation of increase in RI with patient age (r = 0.010) or with pre-ESWL blood pressure (r = 0.002). Elderly patients (> or =60 years old, n = 31) had higher RI levels on baseline than younger patients (<60 years old, n = 39). In 18 of the 31 (58.1%) elderly cases the RI were elevated to greater than 0.7, indicating pathologic changes. In younger patients, only 9 (23.1%) experienced increase in RI up to 0.7 or greater. The contralateral untreated kidneys showed significant change in RI before (0.664+/-0.045) and after (0.679+/-0.049) lithotripsy in elderly patients (P<0.005). A follow-up Doppler study showed that the mean RI returned to pretreatment levels after 1 week. CONCLUSIONS: Because of higher RI levels on baseline, elderly patients have a higher risk of post-ESWL renal tissue damage than younger patients. Clinical implication of RI change in the contralateral kidneys in this study remains to be answered. The measurement of changes in RI with Doppler ultrasound techniques after ESWL may provide useful information for clinical diagnosis of renal tissue damage. PMID- 10533900 TI - Prospective evaluation of prostate cancer detection by prostate-specific antigen related parameters. AB - BACKGROUND: The diagnostic value of prostate-specific antigen (PSA) for differentiating prostate cancer from benign prostatic conditions is limited by its lack of specificity. Several PSA-related parameters have been suggested as enhancing the discriminatory power of total PSA values, but their clinical utility should be considered preliminary until established in a prospectively evaluated cohort. METHODS: In a prospective cohort study, results of ultrasound guided biopsy and/or transurethral resection of the prostate gland were assessed in 706 consecutive Japanese men. The clinical usefulness of total PSA, free PSA, percentage of free PSA, PSA density (PSAD), PSA density for transition zone (PSADT) and gland volume for predicting prostate cancer was investigated using receiver operating characteristic (ROC) curve analysis in 16 different patient subgroups. RESULTS: Overall, 150 of the 706 patients (21.2%) had prostate carcinoma. The ROC curve analysis showed that PSAD and PSADT were more powerful predictors of prostate cancer than total PSA in most of the 16 patient subgroups tested. The improvement in performance was modest, however. No substantial difference was noted between PSAD and PSADT. Total gland volume did not significantly affect the performance of these parameters. The use of a PSAD threshold value of 0.11-10.15 ng/mL per cm3 (or a PSADT value of 0.23-0.27 ng/mL per cm3) would have avoided 24-48% (or, for PSADT, 34-40%) of unnecessary biopsies at the cost of missing 5-10% of detectable cancers in a patient subgroup with intermediate total PSA levels. The performance of free PSA and percentage of free PSA was worse than that of any other test in this study. This may be due to inappropriate handling of sera prior to measurement. CONCLUSIONS: The discriminatory potential of total PSA for predicting prostate cancer was modestly improved by the use of PSAD and PSADT. No substantial advantage of PSADT over PSAD could be demonstrated. Stringent and standardized storage conditions should always be maintained when applying free PSA-related parameters. PMID- 10533901 TI - Significance of hematoma size for evaluating the grade of blunt renal trauma. AB - BACKGROUND: The hematoma size relative to the body size was measured on computed tomography films using a personal computer system in order to define whether that parameter is useful for decision-making in the management of blunt renal trauma. METHODS: From 1982 to 1997, 33 patients with intermediate or severe grade blunt renal trauma were retrospectively divided into three groups: group 1, managed conservatively without transcatheter embolization; group 2, managed by bedrest after selective transcatheter embolization; and group 3, managed operatively. In these three groups, the hematoma area (H) and the ratio of hematoma area to body area on CT (H/B) were measured and the chronological changes of the H/B in groups 1 and 2 were studied. RESULTS: The H and H/B of group 3 were significantly larger than those of group 1. The H/B was more clearly distinguished for each group compared with the H alone. Well-preserved kidney integrity, despite the presence of a large hematoma in group 2, allowed the conservative treatment following transcatheter embolization of the bleeding site. The H/B of all group 1 patients gradually decreased and on the 40th or 50th day after injury they reached a level equivalent to the ratio of contra-lateral normal kidney area to body area. CONCLUSION: The ratio of hematoma area to body area on CT was very useful in evaluating the grade of blunt renal trauma. In conservative treatment for blunt renal trauma changes of the hematoma size is a useful indicator for management. PMID- 10533902 TI - Proliferation and differentiation of rat dorsal prostatic epithelial cells in collagen gel matrix culture, focusing upon effects of adipocytes. AB - BACKGROUND: Prostatic epithelial cells organize functional acinus structures under epithelial extracellular matrix and epithelial-stromal cell interactions. Recently, the adipose tissue, which surrounds and even exists within the prostate, has been suggested to affect the differentiation and proliferation of some cell types. Therefore, tissue fragments, which consist mainly of epithelial and fibromuscular stromal cells, were cultured in three-dimensional collagen gel matrix culture with adipocytes. METHODS: Tissue fragments of rat dorsal prostate, including both epithelial and fibromuscular stromal components, were cultured in collagen gel with or without adipocytes. Epithelial cell differentiation was evaluated with the reconstruction of acinus-like structures and with immunohistochemistry of rat dorsal prostate-specific proteins, dorsal protein-1 and probasin. The proliferation was examined by uridine uptake. RESULTS: Under coculture of the fragments and adipocytes, epithelial cells reconstructed more differentiated acinus-like structures surrounded by fibromuscular stromal cells than tissue fragment culture without adipocytes. Dorsal protein-1 and probasin expressions of epithelial cells in this coculture system were the same as in rat prostate in vivo. In the coculture, epithelial cells had a higher proliferation activity. CONCLUSION: These results indicate that adipocytes promote proliferation and differentiation of prostatic epithelial cells. Our new culture model with adipocytes suggests the importance of cell-cell interactions, including those of epithelial cells and adipocytes. PMID- 10533903 TI - Effect of Bcl-2 overexpression in human prostate cancer cells in vitro and in vivo. AB - BACKGROUND: Cancer cells often develop mechanisms to resist apoptosis and the extent of such anti-apoptotic ability has been shown to parallel tumor progression in various malignancies. Among various molecules implicated in regulating apoptosis pathway, bcl-2 and its family members are best characterized. METHODS: To investigate the effect of bcl-2-mediated anti apoptotic ability on tumor growth and progression in prostate cancer, a cell line overexpressing bcl-2 (LNCaP/bcl-2) was established by genetically engineering a prostate cancer cell line LNCaP. Tumor growth of LNCaP/bcl-2 was compared with the parental cell line in vitro and in vivo. RESULTS: LNCaP/bcl-2 cells show resistance to apoptosis caused by nutrient deprivation and did not arrest when cultured in serum-free or androgen-free medium, while parental LNCaP cells or LNCaP cells transfected with the vector only (LNCaP/control) underwent extensive apoptosis on nutrient deprivation and sustained growth suppression in serum-free or androgen-free medium. When injected subcutaneously into nude mice, tumors deriving from LNCaP/bcl-2 cells grew faster compared with LNCaP/control for about 3 weeks (P = 0.02), but this effect was not evident after 5 weeks. Upon castration, the control tumors regressed but LNCaP/bcl-2-derived tumors showed resistance, as was previously reported. CONCLUSIONS: These data confirm the notion that anti-apoptotic function of bcl-2 is oncogenic and confers resistance to androgen deprivation and also indicate that it may also play a critical role in earlier stages of tumorigenesis. PMID- 10533904 TI - The study of PSA gene expression on urogenital cell lines. AB - PURPOSE: Reverse transcriptase-polymerase chain reaction (RT-PCR) offers a potentially more sensitive assay for detecting cells expressing prostate-specific antigen (PSA) mRNA in peripheral circulation. But the sensitivity and specificity are variable depending on the position of the PSA amplification. To increase sensitivity and specificity, the whole PSA cDNA (1466 bp) was separated into eight different parts. METHODS: We examined RT-PCR on 12 urogenital cell lines, including three prostate cancer (LNCaP, PC3, DU145), five human renal cell carcinoma (SMKT-R3, TOS-1, TOS-2, R4, ACHN), two urinary bladder cancer (YTS-1, KK-47) and two testicular cancer (NEC8, NEC14) cell lines. The sizes of the eight fragmented PSA used in the experiment were PSA-1 (1-257bp), PSA-2 (1-322bp), PSA 3 ( 172-507bp), PSA-4 (172-851 bp), PSA-5 (595-1347 bp), PSA-6 (682 967 bp), PSA 7 (682-1347 bp) and PSA-8 (863-1466 bp). RESULTS: All cell lines had positive signals from PSA-6, PSA-7 and PSA-8. The positive signals from PSA-1, PSA-2 and PSA-3 were detected in some other cell lines in addition to the three prostate cancer cell lines. Only LNCaP which produces the PSA protein had a positive signal from PSA-5. PC3 and DU145 (which do not produce PSA) and LNCaP had a positive signal from PSA-4. Therefore, the inner primer PSA-4' (578-782 bp) used to increase sensitivity and specificity. Nested RT-PCR on the 12 cell lines, using the PSA-4 and 4' primers, detected more clear bands in the three prostate cancer cells. CONCLUSION: Nested RT-PCR using PSA-4 (outer primer) and PSA-4' (inner primer) may be useful for detecting prostate cancer cells in the peripheral blood. PMID- 10533905 TI - Common sheath reimplantation with ureteral plication: a useful technique for the management of ectopic ureterocele. AB - BACKGROUND: When salvaging the upper pole kidney in duplex ectopic ureterocele, primary bladder level surgery with common sheath ureteral reimplantation has the definite advantage of allowing the reconstruction of the entire collecting system through a single lower abdominal incision. However, there are several complications associated with a common sheath reimplantation in a child with a very dilated upper pole ureter, such as vesicoureteral reflux or ureterovesical stenosis. METHODS/RESULTS: To avoid these complications, ureteral plication over the common ureteral sheath in two children with duplex ectopic ureterocele was used. Postoperatively, neither child showed reflux or recurrent urinary tract infection and both showed a marked improvement of the upper pole collecting system. CONCLUSION: This technique allows for a simple and definitive reconstruction in cases of duplex ectopic ureterocele, particularly with dilated upper pole ureter. PMID- 10533906 TI - Leiomyosarcoma of the spermatic cord. AB - BACKGROUND: A rare case of leiomyosarcoma from the spermatic cord is described. A 72-year-old man complained of a hard, golf-ball sized mass in the right inguinal canal. METHODS: Transinguinal radical orchiectomy was performed and histologic examination revealed leiomyosarcoma originating from the spermatic cord. Distant metastases were not observed by further examinations. Radiation as adjuvant therapy was carried out in order to prevent the local recurrence. RESULTS/DISCUSSION: The patient has been alive for 16 months with no evidence of disease. In leiomyosarcoma of the spermatic cord, locoregional recurrence is common. In addition to transinguinal radical orchiectomy, local radiation therapy should be carried out as an adjuvant. PMID- 10533907 TI - Midodrine in intradialytic hypotension. PMID- 10533908 TI - Destructive spondyloarthropathy: an overview. PMID- 10533909 TI - Leukocyte adhesion molecules and leukocyte-platelet interactions during hemodialysis: effects of different synthetic membranes. AB - Membranes made from synthetic polymers, in general, are considered as being biocompatible membranes and tend to be treated as a homogeneous group. However, all of these membranes have multiple and different characteristics that may contribute to interactions with blood components. As a consequence, the biocompatibility profile of synthetic membranes may vary. In the present cross over study, we examined by flow cytometry the effects (expressed as % change from predialysis values) of three different synthetic polymers (polysulfone, PSF; polyacrylonitrile-co-sodium methallyl sulfonate, AN69; ethylenevinylalcohol, EVAL) on the expression of leukocyte adhesion molecules (CD11b/CD18, CD15s) and the interactions between leukocytes and platelets under conditions of routine clinical use. For neutrophils, a statistically significant difference was found in CD15s expression for EVAL as compared to AN69 (p<0.05) and in CD11b/CD18 expression for PSF as compared to both EVAL (p<0.01) and AN69 (p<0.05). No difference between membranes was found on the expression of such adhesive molecules on monocytes. Significant differences in platelet-neutrophil (but not in platelet-monocyte) coaggregate formation were observed between PSF and both EVAL (p<0.001) and AN69 (p<0.01). Reactive oxygen species production by neutrophil population during hemodialysis was significantly different between each pair of synthetic polymers (PSF vs EVAL, p<0.001; PSF vs AN69, p<0.001; AN69 vs EVAL, p<0.05). Our data demonstrate that in terms of leukocyte adhesion receptors and platelet-leukocyte interactions, the biocompatibility profile of the synthetic membranes polysulphone, AN69 and EVAL shows many similarities but also several significant differences. Our results support the concept that biocompatibility evaluation of each membrane should be based exclusively on data generated by that membrane in order to avoid errors based on assumptions about group characteristics. PMID- 10533910 TI - A case of antiphospholipid antibody syndrome diagnosed after thrombosis of an arteriovenous shunt. AB - A 32-year-old male dialysis patient with lupus nephritis was admitted because of shunt obstruction. The arteriovenous fistula was reconstructed, but obstruction recurred twice within several hours after surgery. A high blood level of anticardiolipin beta2-glycoprotein I antibody suggested that shunt obstruction was caused by a thrombotic tendency related to the antiphospholipid antibody syndrome. Accordingly, for the third shunt procedure, antiplatelet therapy (which had been commenced for systemic lupus erythematosus) was combined with dalteparin sodium from before surgery and warfarin was added postoperatively. This regimen prevented shunt obstruction. In conclusion, hemodialysis patients who suffer repeated shunt obstruction should be examined for antiphospholipid antibody syndrome. PMID- 10533911 TI - Surface-immobilized biomolecules on albumin modified porcine pericardium for preventing thrombosis and calcification. AB - The search for a noncalcifying tissue material to be used for valve replacement application continues to be a field of extensive investigation. A series of porcine pericardial membranes was prepared by modifying the glutaraldehyde- treated tissues with albumin and subsequently immobilizing bioactive molecules like PGE1, PGI2 or heparin via the carbodiimide functionalities. The in vitro calcification and collagenase degradation of these modified tissues were studied as a function of exposure time. Furthermore, the biocompatibility aspects of such novel interfaces were established by platelet adhesion and fibrinogen adsorption. The results reported in this article propose that the treatment with antiplatelet agents such as albumin, heparin and prostaglandins (PGE1 or PGI2) change the surface conditioning of pericardial tissues, suggesting a possible role of deposited serum components in affecting mineralization process on bioprosthesis. Therefore, it is worthy to hypothesize that besides inhibiting the accumulation of calcium in the devitalized cells, the early formation of a conditioning layer on the bioprosthesis surface may affect salt precipitations, determining the propensity of the implant to calcify. More detailed studies are needed to understand the involvement of plasma proteins and cellular components of the recipient blood in tissue-associated calcification. PMID- 10533913 TI - A simple method for hepatocyte attachment in hollow fibre bioreactors. AB - A new method for hepatocyte attachment in hollow fibre (HF) bioreactors was proposed and verified. A flow of medium with suspended hepatocytes, evoked by transmembrane pressure (TMP), and directed across the membrane into the fibre lumen, has accelerated and improved hepatocyte contact with the HF. It was found that seeding of hepatocytes onto the membrane was optimal at TMP of 50-80 mmHg. Ammonia utilisation and ureagenesis rates in hepatocytes seeded in the bioreactor suggests that the proposed method warrants proper conditions for cell functionality and allows for extended culture of hepatocytes in HF bioreactors. It is speculated that time cutback between introduction of hepatocytes into the bioreactor and the start of the cell attachment process, accomplished by the presented method, leads to substantially improved recovery of freshly isolated hepatocytes, and consequently to better overall performance of HF bioreactor. PMID- 10533912 TI - Platelet function and hemolysis in centrifugal pumps: in vitro investigations. AB - The effects of centrifugal pumps on blood components other than erythrocytes, namely platelets and their interaction with the coagulation system, are not very well known. In a comparative study with three centrifugal pumps (BioMedicus BP 80, St. Jude Isoflow, and Sarns Delphin) and the Stockert roller pump hemolysis, platelet counts, thromboplastin and partial thromboplastin times, as well as resonance thrombography (RTG) parameters for the assessment of platelet and coagulation function were evaluated in vitro. Normalized indices of hemolysis (NIH) with ACD anticoagulation after 360 minutes were 0.008+/-0.004 (Isoflow), 0.018+/-0.017 (BP-80), 0.085+/-0.051 (Delphin), and 0.049+/-0.010 g/1001 (roller pump). Plasmatic coagulation was activated in all circuits. Platelet function was severely inhibited by the BP-80, indicated by increase in RTG platelet time to 358%+/-150% of initial values compared to 42%+/-29% (Isoflow), 40%+/-20% (Delphin), and 12%+/-10% (roller pump). Fibrin polymerization was affected similarly. The large surface area of the BP-80 leads to an extensive activation of platelets and plasminogen. PMID- 10533914 TI - Microspheres based detoxification system: in vitro study and mathematical estimation of filter performance. AB - Because of the closed plasma (secondary) circuit in the Microspheres based Detoxification System (MDS), a convective blood purification system, the same amount of filtrated plasma is backfiltrated into the blood circuit. Therefore, there is no direct way to determine the ultrafiltrate production rate, which is an important factor of efficiency. The only possible way to estimate the filtration properties of the filter is to consider pressure values. In this study the pressure distribution in the filter was investigated in vitro. To explain the results and to calculate inaccessible parameters, a mathematical model was established which also considered the asymmetric behaviour of the filter membrane. The result was a linear pressure gradient, agreement with the measurements was reasonably good (calculated primary pressure loss differs <13% from measured value when using mean measured filter resistance as model parameter). Linear pressure distribution offers the possibility of easily calculating the filtration length, a parameter which can be used to estimate the filter condition. The comparison between calculated filtration and backfiltration rates offers an instrument of control for these values. PMID- 10533915 TI - The application of two different blood cell separators to harvest CD34+ cells in patients suffering from non Hodgkin's lymphoma. AB - From January 1996 until now, thirty-eight PBSC procedures were carried out on 20 patients suffering from NHL, mobilized by polichemotherapy regimens plus recombinant human Granulocyte-Growth Factor (rhG-CSF). Patients were enrolled in PBSC procedures using Dideco Excel (group A) and Cobe Spectra v.4.7 (group B) blood cell separators. Twelve patients were enrolled in group A (6 males and 6 females, median age 33) and 9 patients in group B (5 males and 4 females, median age 55). The mean White Blood Cell (WBC) and Mononuclear Cells Fraction (MNC) peripheral blood counts were not statistically different in either group and neither were blood CD34+ cell peripheral counts. CD34+ cell peripheral value was predictive of the CD34+ yield while mean values of harvested CD34+ cells were not significantly different. CD34+ cell efficiencies were statistically the same. The CD34+ cell purity of the apheresis harvest was statistically different between the two groups (group A = 3.0+/-2.2%; group B = 1+/-0.9%) p = 0.001. High CD34+ cell yields were observed in both groups which confirms that both blood cell separators are able to harvest hematopoietic progenitor cells from peripheral blood. PMID- 10533916 TI - Calculation of change in brain temperatures due to exposure to a mobile phone. AB - In this study we evaluated for a realistic head model the 3D temperature rise induced by a mobile phone. This was done numerically with the consecutive use of an FDTD model to predict the absorbed electromagnetic power distribution, and a thermal model describing bioheat transfer both by conduction and by blood flow. We calculated a maximum rise in brain temperature of 0.11 degrees C for an antenna with an average emitted power of 0.25 W, the maximum value in common mobile phones, and indefinite exposure. Maximum temperature rise is at the skin. The power distributions were characterized by a maximum averaged SAR over an arbitrarily shaped 10 g volume of approximately 1.6 W kg(-1). Although these power distributions are not in compliance with all proposed safety standards, temperature rises are far too small to have lasting effects. We verified our simulations by measuring the skin temperature rise experimentally. Our simulation method can be instrumental in further development of safety standards. PMID- 10533917 TI - Optical scattering in camera-based electronic portal imaging. AB - An investigation of scintillation light scattering within camera-based electronic portal imaging devices is presented. A simple analytical scatter model is adapted for the precise geometries of two different camera-based imaging systems and the results of modelling and experimental measurements are compared. The results of the study strongly suggest that the main source of optical scattering is multiple reflection between the scintillation screen and 45 degree mirror in both systems. The scattered light has a highly non-uniform distribution, which is strongly dependent upon field size. For large radiation fields the scatter contribution can be over 20% of the primary signal scintillation light intensity in the centre of the field. A purely optical method of removing the scattered light signal using a louvre grid on the surface of the scintillation screen is then presented and experimentally demonstrated to be effective. PMID- 10533919 TI - Resistivity and phase in localized BIA. AB - We describe a system for highly reproducible non-invasive rf impedance measurements as a function of position along body segments such as the thigh. Results are reported for mainly healthy male and female subjects ranging in age from 19 to 65 and in body-mass index from 15 to 40. A principal conclusion is that the phase of the impedance falls monotonically with increasing distance from the knee, with average values substantially above what is found using standard, whole-body bioelectrical impedance analysis (BIA). To compensate for thigh shape, the data are further analysed using an anatomical model based on reasonable approximations for the distributions of muscle, fat and bone, yielding values of the effective resistivity for current flow parallel to the muscle fibres. The phase and resistivity results are discussed with reference to the whole-body BIA study of maintenance haemodialysis patients by Chertow et al, and in regard to possible physiological correlations observed in this work. PMID- 10533918 TI - Fast spectroscopic imaging for non-invasive thermometry using the Pr[MOE-DO3A] complex. AB - The praseodymium complex of 10-(2-methoxyethyl)-1,4,7,10-tetraaza-cyclododecane 1,4,7-tr iacetate) was evaluated as a temperature-sensitive contrast agent using the temperature dependence (approximately 0.12 ppm degrees C(-1)) of the chemical shift of its methoxy side group signal. Pr[MOE-DO3A] was employed in combination with spectroscopic imaging (SI) methods for the determination of spatially resolved 2D and 3D temperature distributions in phantoms. Conventional SI and fast echo planar SI sequences (EPSI) were implemented on a 4.7 T MR imaging system fulfilling the demands for non-invasive thermometry (NIT) with respect to thermal and temporal resolution, being <1 degree C and <20 s total measuring time, respectively. The sequences are based on a fast spin echo SI method taking into account the very short relaxation times of the Pr complex methoxy group (T1 = 28 ms, T2 = 13 ms) and its chemical shift difference (-24 ppm) from water. Calibration curves were measured in a uniformly heated water phantom and 2D SI methods were applied to dynamic heating experiments. The average differences between the temperatures measured via fibreoptic thermometer and those derived from the spectroscopic methods were < or =0.2 degrees C. Furthermore, 3D EPSI experiments with a 16 x 16 x 16 matrix size yielded temperature measurements within 17 s from voxels of size 3 x 3 x 3 mm3. PMID- 10533920 TI - Experimental 3D dosimetry around a high-dose-rate clinical 192Ir source using a polyacrylamide gel (PAG) dosimeter. AB - It is well known that the experimental dosimetry of brachytherapy sources presents a challenge. Depending on the particular-dosimeter used, measurements can suffer from poor spatial resolution (ion chambers), lack of 3D information (film) or errors due to the presence of the dosimeter itself distorting the radiation flux. To avoid these problems, we have investigated the dosimetry of a clinical 192Ir source using a polyacrylamide gel (PAG) dosimeter. Experimental measurements of dose versus radial distance from the centre of the source (cross line plots) were compared with calculations produced with a Nucletron NPS planning system. Good agreement was found between the planning system and gel measurements in planes selected for analysis. Gel dosimeter measurements in a coronal plane through the phantom showed a mean difference between measured absorbed dose and calculated dose of 0.17 Gy with SD = 0.13 Gy. Spatially, the errors at the reference point remain within one image pixel (1.0 mm). The use of polymer gel dosimetry shows promise for brachytherapy applications, offering complete, three-dimensional dose information, good spatial resolution and small measurement errors. Measurements close to the source, however, are difficult, due to some of the limiting properties of the polyacrylamide gel. PMID- 10533921 TI - Transmission measurements in air using the ESTRO mini-phantom. AB - The aim of this work is to study the possibility of using the ESTRO mini-phantom for transmission measurements of primary kerma in water at a point free in air. We discuss in-air measurements in general, with special attention given to in-air equivalent measurements using a water equivalent mini-phantom. The study includes four different photon energies (4, 6, 10 and 18 MV), where scoring of dose and primary kerma inside a mini-phantom in narrow beam geometry is performed with the Monte Carlo code EGS4. The results reveal that relative measurements (i.e. with and without a water absorber present) at 10 cm depth in a mini-phantom do not represent the variation of primary kerma in water at a point free in air (deviations as large as 7% at 4 MV are observed). Minimum deviations are obtained at depths somewhat larger than the depth of dose maximum. The observed deviations are due to a considerable beam hardening in the water absorber, which changes the amount of attenuation and scatter inside the mini-phantom. PMID- 10533922 TI - Near-infrared optical properties of ex vivo human uterus determined by the Monte Carlo inversion technique. AB - The optical properties, absorption (mua) and reduced scattering coefficient (mu's), of ex vivo human myometrium and leiomyoma (fibroid) have been determined by the Monte Carlo inversion technique over the wavelength range 600-1000 nm. This region is currently of interest for new, minimal-access, surgical laser procedures such as photodynamic therapy (PDT) for abnormalities of the uterus, and interstitial laser photocoagulation (ILP) for the thermal ablation of fibroids. In the region 630-675 nm (corresponding to PDT), the optical coefficients of myometrium are mua = 0.041+/-0.012 mm(-1) and mu's = 1.37+/-0.19 mm(-1). For the wavelength range 800-1000 nm (associated with infrared lasers for ILP), the optical coefficients of fibroid were found to be mua = 0.020+/-0.003 mm(-1) and mu's = 0.56+/-0.03 mm(-1). Overall, the optical properties of fibroid were found to be lower than myometrium, and this was attributed to the differences in both anatomy and vascularity. The results show that PDT for ablation of the uterine endometrium is most unlikely to affect any tissues beyond the myometrium, and that the region around 800 nm is the most effective for ablation of fibroids using ILP as the penetration depth of light is greatest at this wavelength. PMID- 10533923 TI - A new statistical method for transfer coefficient calculations in the framework of the general multiple-compartment model of transport for radionuclides in biological systems. AB - A new and simple statistical procedure (STATFLUX) for the calculation of transfer coefficients of radionuclide transport to animals and plants is proposed. The method is based on the general multiple-compartment model, which uses a system of linear equations involving geometrical volume considerations. By using experimentally available curves of radionuclide concentrations versus time, for each animal compartment (organs), flow parameters were estimated by employing a least-squares procedure, whose consistency is tested. Some numerical results are presented in order to compare the STATFLUX transfer coefficients with those from other works and experimental data. PMID- 10533924 TI - Attenuation correction in SPECT using consistency conditions for the exponential ray transform. AB - Using data consistency conditions for the exponential ray transform, a method is derived to correct SPECT data for attenuation effects. No transmission measurements are required, and no operator-defined contours are needed. Furthermore, any 3D parallel-ray geometry can be considered for SPECT data acquisition, even unconventional geometries which do not lead to a set of 2D parallel-beam sinograms. The method is presented for both the 2D parallel-beam geometry and a particular 3D case, called the rotating slant hole geometry. Full details of the algorithms are given. Implementation has been carried out and results are presented in 2D and in 3D using simulated data. PMID- 10533925 TI - Optimization of the arthroscopic indentation instrument for the measurement of thin cartilage stiffness. AB - Structural alterations associated with early, mostly reversible, degeneration of articular cartilage induce tissue softening, generally preceding fibrillation and, thus, visible changes of the cartilage surface. We have already developed an indentation instrument for measuring arthroscopic stiffness of cartilage with typical thickness >2 mm. The aim of this study was to extend the applicability of the instrument for the measurement of thin (<2 mm) cartilage stiffness. Variations in cartilage thickness, which will not be known during arthroscopy, can nonetheless affect the indentation measurement, and therefore optimization of the indenter dimensions is necessary. First, we used theoretical and finite element models to compare plane-ended and spherical-ended indenters and, then, altered the dimensions to determine the optimal indenter for thin cartilage measurements. Finally, we experimentally validated the optimized indenter using bovine humeral head cartilage. Reference unconfined compression measurements were carried out with a material testing device. The spherical-ended indenter was more insensitive to the alterations in cartilage thickness (20% versus 39% in the thickness range 1.5-5 mm) than the plane-ended indenter. For thin cartilage, the optimal dimensions for the spherical-ended indenter were 0.5 mm for diameter and 0.1 mm for height. The experimental stiffness measurements with this indenter correlated well with the reference measurements (r = 0.811, n = 31, p < 0.0001) in the cartilage thickness range 0.7-1.8 mm. We conclude that the optimized indenter is reliable and well suited for the measurement of thin cartilage stiffness. PMID- 10533926 TI - Optimization of importance factors in inverse planning. AB - Inverse treatment planning starts with a treatment objective and obtains the solution by optimizing an objective function. The clinical objectives are usually multifaceted and potentially incompatible with one another. A set of importance factors is often incorporated in the objective function to parametrize trade-off strategies and to prioritize the dose conformality in different anatomical structures. Whereas the general formalism remains the same, different sets of importance factors characterize plans of obviously different flavour and thus critically determine the final plan. Up to now, the determination of these parameters has been a 'guessing' game based on empirical knowledge because the final dose distribution depends on the parameters in a complex and implicit way. The influence of these parameters is not known until the plan optimization is completed. In order to compromise properly the conflicting requirements of the target and sensitive structures, the parameters are usually adjusted through a trial-and-error process. In this paper, a method to estimate these parameters computationally is proposed and an iterative computer algorithm is described to determine these parameters numerically. The treatment plan selection is done in two steps. First, a set of importance factors are chosen and the corresponding beam parameters (e.g. beam profiles) are optimized under the guidance of a quadratic objective function using an iterative algorithm reported earlier. The 'optimal' plan is then evaluated by an additional scoring function. The importance factors in the objective function are accordingly adjusted to improve the ranking of the plan. For every change in the importance factors, the beam parameters need to be re-optimized. This process continues in an iterative fashion until the scoring function is saturated. The algorithm was applied to two clinical cases and the results demonstrated that it has the potential to improve significantly the existing method of inverse planning. It was noticed that near the final solution the plan became insensitive to small variations of the importance factors. PMID- 10533927 TI - Optimal photon energies for IUdR K-edge radiosensitization with filtered x-ray and radioisotope sources. AB - The purpose of this work is to determine the most physically effective radiation energy for K-edge absorption of x- or gamma-rays by iododeoxyuridine (IUdR) on Chinese hamster ovary (CHO) cells. Brachytherapy sources (Sm-145, I-125, Yb-169 and Am-241) and x-ray beams (30 kVp, 100 kVp and 100 kVp with gold, gadolinium, lead or tungsten filtration) were investigated for their preferential absorption qualities by IUdR sensitized DNA. The 30 kVp, 100 kVp and 100 kVp with tungsten filtration were then used to irradiate CHO cells, with or without IUdR incorporation (i.e. 10(-5) M of IUdR for 3 days). Radiation absorption calculations were performed to determine the increase in energy absorption in DNA with and without IUdR incorporated. In order to measure the in vitro biological effects of K-edge absorption, cell survival experiments were performed. The radiation physics calculations yielded an iodine dose enhancement ratio (DER) of 1.4+/-0.15. 1.8+/-0.15 and 2.7+/-0.15 for the 30 kVp, 100 kVp and tungsten filtered 100 kVp respectively, for 18% IUdR replacement of thymidine in DNA. The corresponding cell sensitization enhancement ratios (SER), determined from the cell survival assay, were determined to be 1.24+/-0.2, 1.8+/-0.2 and 2.3+/-0.3 for the 30 kVp, 100 kVp and tungsten filtered 100 kVp respectively, for cells with 18+/-2% IUdR incorporation. These SER values are in reasonable agreement with the DER values of 1.4, 1.8 and 2.7. From these radiation calculations and radiobiology experiments we confirm that using x-radiation energies above the K edge of iodine (33.2 keV) can have a significant effect on cell survival. This effect is due mainly to the increase in the local dose to the DNA for IUdR sensitized cells compared with the normal DNA which lacks the iodine contrast agent. Our results support the clinical application of IUdR and low-energy brachytherapy, perhaps using new technologies such as the x-ray needle or new isotopes such as Yb-169. PMID- 10533928 TI - Cardiac biomagnetic source estimation with a heart-torso model and a trained neural network. AB - The intensity of the cardiac sources for normal adult subjects was estimated from given magnetic field profiles with a trained neural network based on the relationship of the electrical activity of the heart to the cardiac magnetic fields. The input for training the neural network consisted of the magnetic field profiles above the torso during the heartbeat. The outputs were the dipole intensities which produced those magnetic field profiles. A back propagating algorithm with bias and momentum was utilized for training. The measured and simulated torso magnetic field profiles and magnetocardiograms were used for training the neural network. Estimation of the dipole intensities was performed for unknown magnetic field profiles with the trained neural network. The estimated cardiac dipole intensities were reasonably close to the true dipole intensities. These results show the feasibility of the estimation of cardiac dipole intensities with a trained neural network under a very restricted forward model of the cardiac magnetic fields. Generalization of the results to cover a large population base could be difficult because the activation isochrones are different from subject to subject. PMID- 10533929 TI - Bioelectromagnetic localization of a pacing catheter in the heart. AB - The accuracy of localizing source currents within the human heart by non-invasive magneto- and electrocardiographic methods was investigated in 10 patients. A non magnetic stimulation catheter inside the heart served as a reference current source. Biplane fluoroscopic imaging with lead ball markers was used to record the catheter position. Simultaneous multichannel magnetocardiographic (MCG) and body surface potential mapping (BSPM) recordings were performed during catheter pacing. Equivalent current dipole localizations were computed from MCG and BSPM data, employing standard and patient-specific boundary element torso models. Using individual models with the lungs included, the average MCG localization error was 7+/-3 mm, whereas the average BSPM localization error was 25+/-4 mm. In the simplified case of a single homogeneous standard torso model, an average error of 9+/-3 mm was obtained from MCG recordings. The MCG localization accuracies obtained in this study imply that the capability of multichannel MCG to locate dipolar sources is sufficient for clinical purposes, even without constructing individual torso models from x-ray or from magnetic resonance images. PMID- 10533930 TI - Effect of dominant features on neural network performance in the classification of mammographic lesions. AB - Two different classifiers, an artificial neural network (ANN) and a hybrid system (one step rule-based method followed by an artificial neural network) have been investigated to merge computer-extracted features in the task of differentiating between malignant and benign masses. A database consisting of 65 cases (38 malignant and 26 benign) was used in the study. A total of four computer extracted features--spiculation, margin sharpness and two density-related measures--was used to characterize these masses. Results from our previous study showed that the hybrid system performed better than the ANN classifier. In our current study, to understand the difference between the two classifiers, we investigated their learning and decision-making processes by studying the relationships between the input features and the outputs. A correlation study showed that the outputs from the ANN-alone method correlated strongly with one of the input features (spiculation), yielding a correlation coefficient of 0.91, whereas the correlation coefficients (absolute value) for the other features ranged from 0.19 to 0.40. This strong correlation between the ANN output and spiculation measure indicates that the learning and decision-making processes of the ANN-alone method were dominated by the spiculation measure. Three-dimensional plots of the computer output as functions of the input features demonstrate that the ANN-alone method did not learn as effectively as the hybrid system in differentiating non-spiculated malignant masses from benign masses, thus resulting in an inferior performance at the high sensitivity levels. We found that with a limited database it is detrimental for an ANN to learn the significance of other features in the presence of a dominant feature. The hybrid system, which initially applied a rule concerning the value of the spiculation measure prior to employing an ANN, prevents over-learning from the dominant feature and performed better than the ANN-alone method in merging the computer extracted features into a correct diagnosis regarding the malignancy of the masses. PMID- 10533931 TI - Megavoltage CT on a tomotherapy system. AB - A megavoltage computed tomography (MVCT) system was developed on the University of Wisconsin tomotherapy benchtop. This system can operate either axially or helically, and collect transmission data without any bounds on delivered dose. Scan times as low as 12 s per slice are possible, and scans were run with linac output rates of 100 MU min(-1), although the system can be tuned to deliver arbitrarily low dose rates. Images were reconstructed with clinically reasonable doses ranging from 8 to 12 cGy. These images delineate contrasts below 2% and resolutions of 3.0 mm. Thus, the MVCT image quality of this system should be sufficient for verifying the patient's position and anatomy prior to radiotherapy. Additionally, synthetic data were used to test the potential for improved MVCT contrast using maximum-likelihood (ML) reconstruction. Specifically, the maximum-likelihood expectation-maximization (ML-EM) algorithm and a transmission ML algorithm were compared with filtered backprojection (FBP). It was found that for expected clinical MVCT doses enough imaging photons are used such that little benefit is conferred by the improved noise model of ML algorithms. For significantly lower doses, some quantitative improvement is achieved through ML reconstruction. Nonetheless, the image quality at those lower doses is not satisfactory for radiotherapy verification. PMID- 10533932 TI - A spline-regularized minimal residual algorithm for iterative attenuation correction in SPECT. AB - In SPECT, regularization is necessary to avoid divergence of the iterative algorithms used for non uniform attenuation compensation. In this paper, we propose a spline-based regularization method for the minimal residual algorithm. First, the acquisition noise is filtered using a statistical model involving spline smoothing so that the filtered projections belong to a Sobolev space with specific continuity and derivability properties. Then, during the iterative reconstruction procedure, the continuity of the inverse Radon transform between Sobolev spaces is used to design a spline-regularized filtered backprojection method, by which the known regularity properties of the projections determine those of the corresponding reconstructed slices. This ensures that the activity distributions estimated at each iteration present regularity properties, which avoids computational noise amplification, thus stabilizing the iterative process. Analytical and Monte Carlo simulations are used to show that the proposed spline regularized minimal residual algorithm converges to a satisfactory stable solution in terms of restored activity and homogeneity, using at most 25 iterations, whereas the non regularized version of the algorithm diverges. Choosing the number of iterations is therefore no longer a critical issue for this reconstruction procedure. PMID- 10533933 TI - Implementation and quantitative evaluation of analytical methods for attenuation correction in SPECT: a phantom study. AB - Three differing exact methods of inverting the two-dimensional (2D) exponential Radon transform were implemented and evaluated quantitatively with a phantom study. The phantom had the shape of a pie-chart divided into six cavities, each 480 ml in volume and 10 cm in height, that were symmetrically positioned in a cylinder that was 20 cm in diameter and 10 cm in height. This phantom tests for linearity between true activity concentration and measured activity concentration, and it is denoted as a linearity phantom in the present study. Each cavity contained a different concentration of a homogeneous solution of 99mTc (74, 148, 222, 296, 370 and 444 kBq ml(-1)). Data acquisition was performed with two energy windows: a 20% photopeak energy window set symmetrically over the 140 keV of 99mTc and a secondary 5% energy window set over the 122 keV peak. We optimized a triple-energy window scatter correction method for a gamma camera collimator system to obtain accurate scatter-corrected projections. A circular ROI 3 cm in diameter was identified over each cavity region, and count density (counts per pixel) was calculated. This value was converted to activity concentration (kBq ml(-1)) using a cross-calibration coefficient between SPECT counts and the gamma well counter. The relation between true activity (x) and measured activity concentration (y) was fitted to a line using the least-squares method. Regression lines were y = 0.63 + 1.0255x (R2 = 0.9987), y = -2.62 + 1.0278x (R2 = 0.9995), and y = 0.092 + 1.0241x (R2 = 0.9989) for the Bellini, Inouye and Metz-Pan methods respectively. In another phantom study using two different types of phantoms, contrast of a cold region in the two was 96% and 101% for all three methods. Combined optimized scatter correction and analytical attenuation correction methods achieve good accuracy in quantification of activity distribution with a uniform attenuating medium. PMID- 10533934 TI - Geometrical models of left ventricular contraction from MRI of the normal and spontaneously hypertensive rat heart. AB - This study develops a quantitative analysis and model for the differences in left ventricular dynamics in normal and spontaneously hypertensive rats, as determined using non-invasive magnetic resonance imaging (MRI). We emerge with a characterization of the geometrical changes in the left ventricle resulting from hypertension. In addition, the techniques we have adopted are potentially applicable to the study of other disease models for important human cardiac pathologies. A gradient-echo multislice imaging sequence (echo time 4.3 ms) achieved complete image coverage of the heart at high time resolution (13 ms) through the cardiac cycle. Cardiac anatomy in two age-matched groups of young adult (8 and 12 weeks old) normal Wistar-Kyoto (WKY, n = 8) and spontaneously hypertensive rats (SHR, n = 8) was imaged in synchrony with the electrocardiographic R wave in defined planes both parallel and perpendicular to the principal cardiac axis. The transverse left ventricular image sections were circularly symmetrical; this permitted application of different analytical models for the three-dimensional geometry of the epi- and endocardial borders. An ellipsoidal figure of revolution offered an effective description of the three dimensional left ventricular geometry throughout the cardiac cycle in both normal WKY and SHR animals. The model successfully characterized both the dynamic changes in the shape of the left ventricle through the cardiac cycle and the pathological alterations resulting from spontaneous hypertension. The elliptical model also formed the basis of a simple stress distribution analysis. Such parametric descriptions thus provided a useful alternative to more complex finite element analyses of cardiac function. The eccentricity of the ventricle was characterized by an ellipticity factor a, where a = 1 for a sphere and a < 1 for a prolate ellipsoid. At end systole, the endocardial surface of the left ventricle gave a = 0.43+/-0.02 and 0.49+/-0.02 for the WKY and SHR animals respectively (probability, P < 0.05). At end diastole, the endocardial surface of the left ventricle gave a = 0.58+/-0.02 and 0.63+/-0.02 for the WKY and SHR animals respectively (P < 0.05). Such a difference in ventricular shape was a potential adaptation to increased blood pressure. Hypertension thus altered the left ventricular ellipticity to give a more spherical geometry compared with the normal rats. PMID- 10533935 TI - A new polymer gel for magnetic resonance imaging (MRI) radiation dosimetry. AB - New composition polymer gels, the N-vinylpyrrolidone argon (VIPAR) gels, were developed and investigated as MRI dosimeters. VIPAR gels were irradiated in the dose range of 0-12 Gy by a 6 MV x-ray linear accelerator and MR-scanned in a 1.5 T magnetic resonance imager. A linear relationship was found between absorbed dose and spin spin relaxation rate R2. The dose sensitivity was found to be approximately 0.1 s(-1) Gy(-1) for a gel composition of 4% w/w in N vinylpyrrolidone, 4% w/w in N,N'-methylene-bisacrylamide, 5% w/w in gelatine type A and 87% w/w in water. This dose sensitivity was stable with time and did not deteriorate even when a boost radiation dose of 2.5 Gy was applied 15 days after the first irradiation. Good reproducibility of these results was observed when a new batch of gels was produced and used for corresponding measurements and analysis. PMID- 10533936 TI - Dose quality assurance for stereotactic radiotherapy treatments. AB - A simple Perspex phantom is presented for routine dosimetry checks of stereotactic radiotherapy procedures, along with a procedure for checking the planning process. The phantom is hemispherical and water filled. It can be fitted into the Brown-Robert-Wells fiducial system for CT scanning and attached to the stereotactic frame for treatment. Both arcing and fixed field plans can be carried out on the phantom. We present our results, performed on a Varian 600C linac, which were found to be within 2% of the expected dose. PMID- 10533937 TI - An equation describing spread of membrane potential changes in a short segment of blood vessel. AB - The spread of membrane potential changes throughout certain cells and tissues plays an important role in their physiology. The attenuation of such changes in any tissue is usually characterized by the cable length constant lambda, which can be determined experimentally if the equations describing membrane potential spread in the tissue are known. Here we derive an equation describing spread of membrane potential changes in a short cable, which is an appropriate model for short segments of blood vessels. This equation is more general than those already published in that the positions of both the current source that gives rise to a potential change, and the point at which the change is measured, can be anywhere along the cable. PMID- 10533938 TI - Development of collimator insert for linac based stereotactic irradiation. AB - The aim of this study is to develop collimator inserts of various sizes which are either not commercially available or are expensive to import. The dosimetry parameters such as tissue maximum ratio (TMR), off-axis ratio (OAR) and output factor of the developed collimator insert are compared with that of the commercial collimator insert (Radionics). In order to check the suitability of the collimator insert developed locally for clinical use and to standardize the method of development, a collimator insert of 15 mm identical to the one supplied by Radionics is developed with low-melting alloy (Cerrobend). Moreover for the clinical use of the developed collimator insert, certain acceptance tests are performed which include a collimator concentricity test, beam size check and radiation leakage test. The dose verification is carried out with a thermoluminescent dosimeter (7LiF rods) and an FBX chemical dosimeter in a human head-shaped Perspex phantom filled with water. The variation between the calculated and measured dose is found to be within +2.4% for 7LiF rods and -2.0% for the FBX chemical dosimeter thus ensuring the suitability of the developed collimator insert for clinical use. This has encouraged us to standardize the method adapted to develop the collimator insert and to develop collimator inserts of different field sizes. PMID- 10533939 TI - A checklist for reporting of thermoluminescence dosimetry (TLD) measurements. PMID- 10533940 TI - Reliability of published data on radionuclide half lives--relevance to the use of reference sources for checking instrument performance. PMID- 10533941 TI - GPs' management of women seeking help for familial breast cancer. AB - OBJECTIVE: We aimed to ascertain how often patients seek help for familial breast cancer in primary care, and to identify GPs management of these patients, in order to see whether guidelines are followed. METHODS: This was a descriptive study. GPs (n = 202) attending a postgraduate education programme were asked to fill in a questionnaire which included questions about the number of patients seeking help for familial breast cancer within the last 3 months and about their management strategies. RESULTS: About 80% of the GPs reported that they referred women with concerns about familial breast cancer for further diagnostics (mammography or ultrasound). For half these referrals a plan of regular appointments was set up, and one-eighth of the referrals included breast examination by a physician. Breast self-examination was advised in 50% of the cases. Estimates given to women regarding their breast cancer risk varied considerably. There was a strong relationship between risk estimates and management strategies. CONCLUSIONS: Current guidelines regarding surveillance of women with breast cancer in the family were only partly followed. These guidelines do not give sufficient information to define whether there is an increased risk for breast cancer. These guidelines need to be refined. PMID- 10533942 TI - GPs' views on their role in cancer genetics services and current practice. AB - BACKGROUND: Increasing demand for cancer genetics services has necessitated an urgent review of how these services are organized and, in particular, identification of an effective role for primary care. OBJECTIVES: We aimed to assess the views of GPs on their role in cancer genetics services and their confidence in performing that role; to assess their understanding of cancer genetics, current practice and referral behaviour; and to identify needs for information and training to enable GPs to play an effective role in these services. METHOD: A cross-sectional questionnaire survey of GPs was conducted through general practices in SE Scotland; 397 (response rate 59.3%) GPs returned a completed questionnaire. Outcome measures were: responders' perceptions of their role in cancer genetics services; confidence within that role; understanding of cancer genetics; current practice regarding patients presenting with concerns about their family history of cancer; and perceived information and training needs. RESULTS: GPs identified their role to be: taking a family history; making appropriate referrals to specialist services; providing emotional support; teaching breast self-examination; and discussing need for screening. Lack of confidence within this role was reflected in low levels of understanding of cancer genetics and in inappropriate referral practices. Concerns were expressed about the increasingly specialist role demanded of primary care. A desire for referral guidelines and community genetics clinics was identified. CONCLUSIONS: GPs readily identify a role for themselves in cancer genetics services, but admit to a lack of confidence in this area, calling for clear referral guidelines and specialist community support. Current inappropriate referral to specialist services results from a lack of confidence in estimating cancer risk, highlighting the need for the development of clear referral criteria. Given the rapidly increasing demand for cancer genetics services and the vital role of primary care, it is important to identify a model of these services that facilitates effective involvement of GPs without further increasing their workload. PMID- 10533943 TI - Chronic pain in primary care. AB - Chronic pain is a very common cause of suffering, disability and economic adversity in the community. It is a complex problem that needs to be understood in a multi-dimensional way for effective management. Most research to date has been based in specialist clinics rather than in primary care, with consequently limited findings. Chronic pain differs from acute pain in that management follows a rehabilitative rather than a treatment model, though these are not mutually exclusive. Full assessment of the patient, preferably multi-disciplinary, will improve his or her outlook. Management should be holistic, rigorous in the application of conventional therapies (including analgesics and physical therapy) and ready to admit an improved understanding of psychological and social techniques. There may be a role for complementary therapies. As a large proportion of chronic pain presents only in the community, there may be a role for greater primary care input to management. PMID- 10533944 TI - Helicobacter treatment with quadruple therapy in primary health care for patients with a history of ulcer disease. AB - BACKGROUND: Few patients with a history of peptic ulcer are treated by their GP for H. pylori infection, even though theoretical evidence supports such an approach. OBJECTIVES: We aimed to determine the validity of this recommendation and to test the feasibility of quadruple therapy in primary health care. METHODS: In this prospective, non-randomized intervention study, 51 unselected patients with a history of proven ulcer disease received a 7-day quadruple therapy (lansoprazole, colloidal bismuth subcitrate, tetracycline and metronidazole) from their GP. Main outcome measures were: (i) endoscopically confirmed cure of the infection; (ii) results of serology at entry and at 6 months follow-up; (iii) quality of life at entry, at 6 weeks and at 6 months follow-up; (iv) gastric symptoms at entry, at 6 weeks and at 6 months follow-up; and (v) medication at entry and at 6 months follow-up. RESULTS: Quadruple therapy was well tolerated and there were no drop-outs with this regimen. Intention to treat cure rate was 48/51 (94%, 95% CI 87-100%), per protocol cure rate was 48/49 (98%, 95% CI 94 100%). 45/50 (90%) had positive serology at entry. IgG antibody titres decreased > 40% in 95.2% of patients. Quality of life improved significantly after treatment, gastric symptoms decreased and medication use decreased. CONCLUSIONS: GPs should be encouraged to identify patients with a history of ulcer disease and chronic use of acid suppressants and offer them treatment for H. pylori infection. This approach will cure the infection in almost all patients, it will improve the quality of life and decrease costs. Quadruple therapy does not lose efficacy when employed in primary care. Pre-treatment serological testing is potentially useful for narrowing down the treatment group to those with actual infection, and serology is promising as an easy and cheap follow-up instrument in primary health care. PMID- 10533945 TI - Use of multiple methods to determine factors affecting quality of care of patients with diabetes. AB - BACKGROUND: The process of care of patients with diabetes is complex; however, GPs are playing a greater role in its management. Despite the research evidence, the quality of care of patients with diabetes is variable. In order to improve care, information is required on the obstacles faced by practices in improving care. Qualitative and quantitative methods can be used for formation of hypotheses and the development of survey procedures. However, to date few examples exist in general practice research on the use of multiple methods using both quantitative and qualitative techniques for hypothesis generation. OBJECTIVES: We aimed to determine information on all factors that may be associated with delivery of care to patients with diabetes. METHODS: Factors for consideration on delivery of diabetes care were generated by multiple qualitative methods including brainstorming with health professionals and patients, a focus group and interviews with key informants which included GPs and practice nurses. Audit data showing variations in care of patients with diabetes were used to stimulate the brainstorming session. A systematic literature search focusing on quality of care of patients with diabetes in primary care was also conducted. RESULTS: Fifty-four potential factors were identified by multiple methods. Twenty (37.0%) were practice-related factors, 14 (25.9%) were patient-related factors and 20 (37.0%) were organizational factors. A combination of brainstorming and the literature review identified 51 (94.4%) factors. Patients did not identify factors in addition to those identified by other methods. CONCLUSION: The complexity of delivery of care to patients with diabetes is reflected in the large number of potential factors identified in this study. This study shows the feasibility of using multiple methods for hypothesis generation. Each evaluation method provided unique data which could not otherwise be easily obtained. This study highlights a way of combining various traditional methods in an attempt to overcome the deficiencies and bias that may occur when using a single method. Similar methods can also be used to generate hypotheses for other exploratory research. An important responsibility of health authorities and primary care groups will be to assess the health needs of their local populations. Multiple methods could also be used to identify and commission services to meet these needs. PMID- 10533946 TI - Influencing antibiotic prescribing in general practice: a trial of prescriber feedback and management guidelines. AB - BACKGROUND: The extent of use of antibiotics to treat upper respiratory infections in general practice is an area for concern due to the increasing problem of bacterial resistance. Effective educational strategies to promote rational prescribing are needed. OBJECTIVES: We aimed to examine the effectiveness of prescriber feedback and management guidelines in reducing antibiotics prescribing by GP trainees for undifferentiated upper respiratory tract infection, and in improving the choice of antibiotic for tonsillitis/streptococcal pharyngitis. The research tested a stepwise approach to targeting educational input to high prescribers. METHOD: General Practice trainees in New South Wales (n = 157) were randomly allocated to a treatment group (n = 78) which received an education intervention on antibiotic use, or to a control group (n = 79) which received an intervention on an unrelated topic. Trainees completed three practice activity surveys, each of 110 consecutive patient encounters, with 6-month intervals between surveys. Prescriber feedback and management guidelines on use of antibiotics for URTI and choice of antibiotic for tonsillitis/streptococcal pharyngitis were delivered in a written form between surveys 1 and 2. An educational outreach visit to high prescribers occurred between surveys 2 and 3. Outcome measures were the rate of antibiotic prescribing for all indications, for URTI and prescribing of select antibiotics for tonsillitis/streptococcal pharyngitis. RESULTS: Antibiotic prescribing by the intervention group declined over three occasions from 25.0 to 23.3 to 19.7 per 100 URTI problems, while the control group increased from 22.0 to 25.0 to 31.7 per 100 URTI problems (P = 0.002). Prescribing in agreement with accepted guidelines for tonsillitis/streptococcal pharyngitis increased over time in the intervention group from 55.6 to 69.8 to 73.0 per 100 problems, but decreased in the control group from 59.6 to 57.5 to 58.5 (P = 0.05). CONCLUSION: Prescriber feedback and management guidelines were shown to influence antibiotic prescribing for URTI and choice of antibiotic for tonsillitis/streptococcal pharyngitis. This study provides a model for targeting educational input to those prescribers who most need to change their behaviour. PMID- 10533948 TI - Urinary incontinence: an unexpected large problem among young females. Results from a population-based study. AB - BACKGROUND: The International Continence Society has defined urinary incontinence as a condition in which involuntary loss of urine is objectively demonstrable and is a social or hygiene problem. Urinary incontinence is presumably a common health problem among women even in younger ages. OBJECTIVES: The primary aim was to investigate the prevalence of urinary incontinence (UI) in a female population with a special focus on younger women (18-30 years old). The secondary aim was to investigate the association between UI and number of deliveries, use of contraceptives or oestrogen substitutions, and urinary tract infections (UTIs). METHODS: A population-based study with a self-administered questionnaire was set in the community of Surahammar, Sweden. Subjects were all women (3493) aged 18-70 years living in Surahammar during 1995. The main outcome measures were the prevalence of UI and variables such as number of deliveries, use of contraceptives or oestrogen substitutions, and UTIs. RESULTS: Twenty-six per cent of the women reported problems of UI. The prevalence of UI in younger women was 12%. The number of reported complaints of UTIs was significantly higher in the women with UI compared with women without urinary incontinence (wUI). In the younger women UTI, nulliparous or having given birth to one or two children were most frequent in those with UI. The use of contraceptives was more common in younger women without UI (P < 0.05). However, the use of oestrogen was more common in older women in the age group 51-70 years with UI (P < 0.01). CONCLUSION: Our findings have shown that 26% of the women who took part in the survey reported problems of UI. Among women below 30 years of age, 12% reported complaints of UI. We found a high prevalence of UI in younger women with a UTI, not taking oestrogen, nulliparous or having given birth to one or two children. There are needs for further investigations with a special focus on younger women. PMID- 10533947 TI - The advisability of implementing cholesterol screening in school-age children and adolescents with a family history of cardiovascular disease and hyperlipidaemia. AB - BACKGROUND: The family basis of coronary heart disease is well recognized and it is important for family physicians to assess whether children have elevated cholesterol levels. OBJECTIVES: We aimed to evaluate the advisability of implementing cholesterol screening in children with a family history of cardiovascular disease and hyperlipidaemia. METHODS: We conducted a cross sectional study in Taiwan from February to June 1996. There were 47,800 students in the population. A total of 4520 students were recruited by two-stage sampling. All the participants were required to fill out a structured questionnaire. RESULTS: The response rate was 92.5%. Our results show that 16-18% of the children had a positive family history of cardiovascular disease or hyperlipidaemia. Children with a family history of hyperlipidaemia were significantly more likely to have elevated total cholesterol and low-density lipoprotein cholesterol than those without such a history (both odds ratios: 1.4, P < 0.05). Positive predictive values of hyperlipidaemia were less than 13% based on family history. More than 75% of children with abnormal lipid levels would be missed. CONCLUSIONS: The data suggest that parents' self-reported family history is an ineffective means of identifying children with elevated serum lipid levels in Taiwan. Further research and modification of current National Cholesterol Education Program Panel guidelines for selective cholesterol screening in children may be warranted. PMID- 10533949 TI - Randomized controlled trial of the effect of medical audit on AIDS prevention in general practice. AB - OBJECTIVE: We aimed to evaluate the effect of a medical audit on AIDS prevention in general practice. METHODS: We conducted a prospective randomized controlled study performed as 'lagged intervention'. At the time of comparison, the intervention group had completed 6 months of audit including a primary activity registration, feedback of own data and a meeting with colleagues and experts, and had received brief summaries of the meetings and reminders about the project (a full 'audit circle'). The participants were from general practices in Copenhagen and the Counties of Funen and Vejle, Denmark. One hundred and thirty-three GPs completed the project. The main outcome measures were the number of consultations involving AIDS prevention and the number of talks about AIDS initiated by the GP, and some elements of the content were registered on a chart. RESULTS: No statistically significant difference was observed in the frequency of consultations involving AIDS prevention between the intervention group (1.2% of consultations) and the control group (1.4%). Furthermore, no significant differences were observed regarding the content of these consultations or regarding the fraction of such consultations initiated by the GPs. CONCLUSIONS: Medical audit had no observed effect on AIDS prevention in general practice. PMID- 10533950 TI - The impact of patients' influence on recovery in a group of patients with dyspepsia. AB - BACKGROUND: The approach to health and disease can either be salutogenic (origins of health) or pathogenic (disease causing), which thus makes recovery a concept featuring several different angles. Antonovsky, with his concept of salutogenesis, tried to reach a more complete understanding of its favourable effects on health. OBJECTIVE: We aimed to investigate, understand and learn from the experiences of a small group of patients about factors leading to recovery. METHODS: A qualitative approach was used to explore patient experiences. One semi structured interview was conducted by one of the authors (BN) with each of the 18 patients suffering from dyspepsia who had been investigated by means of gastroscopy at a university hospital clinic 12-15 years previously. The interviews were recorded either in written notes composed directly after the interviews or tape-recorded and subsequently transcribed. A modified form of grounded theory according to Strauss-Corbin was used to analyse the data. RESULTS: A pattern featuring five types of patients' influence on their lives was discerned, ranging from "a sense of no possibility of having an influence on existence/life" to "having influence". Strategies used by patients to maintain health could be categorized into four types: "extremists", "oscillators", "leapers" and "full-scalers". CONCLUSIONS: Listening to patients who had experiences with dyspepsia brought patient influence on their own lives and on the care process into focus. We consider that there might be a link between patients having an influence on their lives and their being healthy today. In clinical practice, patient recovery and health promotion could gain from a perspective where patient influence is treated with esteem and emphasized in the consultation. In the future, research design could benefit from taking patient influence on the care process into consideration. However, no causal linkage between patient influence and patient outcome was established in this study. In order to do that, studies with quantitative design should be undertaken in the future. PMID- 10533951 TI - Promoting medical self-care: evaluation of a family intervention implemented in the primary health care by pharmacies. AB - BACKGROUND: Medical self-care is the range of behaviours undertaken by people to promote or restore health when dealing with a medical problem. OBJECTIVES: The aim of the study was to evaluate medical self-care effects of a family intervention implemented in primary health care by pharmacies, in terms of non professional and professional involvement. METHODS: The intervention was implemented in one of two primary health care areas during a 4-month period and involved consecutive families acting as an intervention (IG, n = 94) or a control (CG, n = 93) group. Eight telephone interviews were conducted with each family. The families were asked about complaints of illness, how long they prevailed and how they were treated. RESULTS: The results showed (P < 0.05-0.0001) that the IG had more medical problems (931 versus 621) compared with the CG, were less hospitalized (4 versus 10), stayed at home more to take care of sick children (84 versus 40), read more medical brochures (121 versus 31), tried more non-medical treatments (228 versus 116), and had fewer visits to the department of paediatrics but more visits to primary health care (69 and 98 versus 90 and 68). CONCLUSIONS: Due to the non-randomization procedure, some caution with regard to generalization of the results must be taken, but they are in concordance with established knowledge of the usefulness of medical self-care. The results indicate that a brief intervention for families can change the use of health authorities. It therefore seems meaningful to implement the intervention in a more comprehensive way in the primary health care setting, while at the same time trying to implement it as a large-scale randomized experimental study, comprising aspects such as the individual's need for care, the use of the right organization level and the assessment of economic costs and savings. PMID- 10533952 TI - Comparison of symptoms in Japanese and American depressed primary care patients. AB - BACKGROUND: Depression is a highly prevalent, worldwide problem with multiple social and health consequences. It often presents in primary care with physical symptoms. Little research has been done on cross-cultural expression of depression in primary care. This paper examines the hypothesis that depressed Japanese patients present with more and with more distinct somatic complaints than depressed American patients. METHODS: Data were collected by chart audit for patients with a diagnosis of depression at two sites: Minamikawachi Tochigi, Japan and Cleveland, Ohio, USA. Patient demographics and type and number of presenting symptoms in the two populations were compared. Logistic regression was used to determine whether there were differences between countries in physical symptoms and to adjust for relevant demographic characteristics. RESULTS: Japanese family physicians charted more somatic complaints from patients diagnosed as depressed than did American family physicians. Specific physical symptoms differed by country: Japanese patients had more abdominal distress, headaches, and neck pain. These symptoms have strong cultural significance for Japanese patients. CONCLUSIONS: This study clearly indicates the prominence and importance of physical symptoms in the presentation of depression in Japanese primary care patients. Their physicians must be alerted to the possibility of depression, especially when patient complaints include abdominal, neck or head pain. PMID- 10533953 TI - Selections from current literature: effects of hawthorn on the cardiovascular system. PMID- 10533954 TI - Selections from current literature: focus group technique in chronic illness. PMID- 10533955 TI - Mildly dyskaryotic smear results. PMID- 10533956 TI - Histologic changes in three explanted native cardiac valves following use of fenfluramines. AB - Use of fenfluramines, either alone or co-administered with phentermine ("fen phen") as anorexic agents in obesity, has been associated with the development of clinically significant cardiac valve disease. We present the macroscopic and histologic findings in cardiac valves explanted from three patients who presented with valvular disease after fenfluramine or fenfluramine-phentermine use and underwent single valve replacement surgery. Paraffin sections were prepared with hematoxylin and eosin, trichrome, elastic-van Gieson, and Giemsa stains, as well as immunostains using antibody to CD3 and CD20. All three patients (two females, ages 37 and 43, and a 49-year-old male) developed progressively symptomatic mitral (2 patients) or aortic (1 patient) valvular insufficiency following dexfenfluramine (2 patients) or fenfluramine-phentermine (1 patient) use. Macroscopic changes included irregular leaflet thickening, accompanied by chordal fusion in the mitral valves, but without vegetations, commissural fusion, or evidence of annular dilation. Histologically, fibromyxoid plaques and nodules just below the valve surface, superficial to a generally intact elastic fiber layer, were associated with CD3-positive lymphocytes. Valves from all three patients had central myxoid degenerative changes, which were focal/mild in one mitral valve, diffuse/moderate in one mitral valve, and diffuse/marked in one aortic valve. Focal areas of superficial fibromyxoid change or intimal thickening may also be seen in cardiac valves from patients with drug-unrelated processes leading to symptomatic or asymptomatic valvulopathy. Therefore, when valve tissue is available for histopathologic examination, valvular disease can be attributed to use of fenfluramines only if the following criteria are satisfied: (i) the macroscopic and microscopic features are consistent with fenfluramine-related valvulopathy, (ii) clinical, echocardiographic, and intraoperative findings support the diagnosis, and (iii) the history of drug exposure predates the development or exacerbation of valvular dysfunction. PMID- 10533957 TI - Lipolysis is an important determinant of isoproterenol-induced myocardial necrosis. AB - The cardiotoxic effect of isoproterenol (ISO) is associated with, and possibly due to, calcium overload. Prior work suggests that calcium entry into cardiac myocytes after ISO administration occurs in two phases: an early rapid phase, followed by a slow phase beginning about 1 hour after ISO injection, leading to a peak myocardial calcium level after about 4 hours. We have tested the relationship of these phases to myocardial necrosis (MN) by determining the time after ISO administration at which the commitment to MN occurs. This was done by administration of propranolol at various times before and after ISO. In addition, since ISO induces lipolysis, and lipids can be toxic, experiments were conducted to determine if adrenergically-activated lipolysis could play a significant role in ISO-MN. We found that propranolol protected the myocardium equally well when administered anytime within 2 hours of ISO injection, but had no effect when given 4 hours after ISO. This showed that metabolic events taking place more than two hours after ISO injection are required for ISO-MN. As expected from prior work, there was a small and consistent amount of propranolol-resistant ISO-MN. Lipolysis, assessed by measuring serum glycerol levels, increased to tenfold above base line at one hour after ISO administration and returned to near basal levels at 4 hours. Potentiation of lipolysis by intravenous injections of phospholipase A2 (PLA2) or lipoprotein lipase (LPL) to rats treated with ISO substantially augmented MN. Propranolol completely blocked the increase in necrosis produced by PLA2 when given with ISO. Lipases induced only minimal necrosis in the absence of ISO. Administration of adenosine (an anti-lipolytic agent), oxfenicine (an inhibitor of mitochondrial palmitoyl carnitine transferase), or vitamin C (an anti-oxidant) resulted in a 55-60% reduction in MN. These results suggest that critical necrosis-determining events occur between 2 and 4 hours after ISO administration and imply a relationship between ISO induced lipolysis, calcium influx, and ISO-MN. We hypothesize that importance of lipolysis as a determinant of ISO-MN is related to the generation of free fatty acids, their oxidized/metabolic products, or direct damage to plasma membrane. PMID- 10533958 TI - A postmortem review of congenital cardiac malformations in a series of 180 adults, over the age of 16 years, born between 1865 and 1980. AB - The evolution of diagnosis and treatment of congenital heart malformations can be traced through patients surviving into adulthood. We reviewed the heart specimens from 180 patients aged 16 to 86 years and considered the morphological features, the nature of any interventional procedures, and the events leading to death. Based on the mode of clinical presentation, 33 cases were considered covert, and the remaining 147 cases were known or suspected to have a cardiac abnormality during life. Of the symptomatic cases, 60 had no surgical intervention, whereas 167 surgical procedures had been performed in the remaining 87 cases. Acquired heart disease was noted in 7 of the covert cases and in 16 of the symptomatic cases. Overall, there were only 3 instances of errors in clinical identification of significant morphological abnormalities, and 2 cases related to surgical procedures. This review emphasizes the value of autopsy examination for clinicopathologic correlations and the case for retention of cardiac specimens for teaching purposes. PMID- 10533959 TI - Clinical and pathologic study of two siblings with arrhythmogenic right ventricular cardiomyopathy. AB - ARVC is a cardiomyopathy in which the right ventricular myocardium is replaced by fibroadipose tissue. Males are affected slightly more often than females and, in those cases which are familial, the pattern of inheritance is usually autosomal dominant with incomplete penetrance. We examined the hearts of two sisters, ages 17 and 14, with no family history of heart disease. The older sibling, who was previously considered healthy, died suddenly, while the younger sibling developed congestive heart failure and received a cardiac transplant. An autopsy of the older sibling and examination of the younger sibling's excised heart revealed severe examples of ARVC with minor differences. A thick cap of fibroadipose tissue covered most, if not all, of each right ventricle and was transmural in some areas. Microscopically, lelt ventricles contained extensive myocyte disarray and multifocal fibrosis. The coronary arteries displayed intimal hyperplasia with disruption of the internal elastic lamina, similar to fibromuscular dysplasia. These two cases comprise a unique familial grouping in a polymorphic disease. Despite the male predominance and autosomal dominant inheritance in ARVC, the only members affected in this family were female, and an autosomal dominant pattern of inheritance, even with incomplete penetrance, would be unusual. In addition, we identified changes in the coronary arteries similar to fibromuscular dysplasia and corroborated recently reported changes in the left ventricle of patients with ARVC, providing evidence that this disease, in its most severe form, involves the entire heart. PMID- 10533960 TI - Ultrastructural alterations during the critical phase of reperfusion: a stereological study in buffer-perfused isolated rat hearts. AB - The present study focuses on myocardial ultrastructural alterations during the early phase of reperfusion. Isolated buffer-perfused rat hearts were exposed to standard perfusion (control group,n = 10); 60 min of global ischemia (n = 10); 60 min of global ischemia followed by 2 min of reperfusion (n = 10); or 60 min of global ischemia followed by 10 min of reperfusion (n = 10). The hearts were perfusion-fixed for electron microscopy, and ultrastructural evaluation was performed using stereological technique in order to obtain an estimate of the volume fraction and absolute volume of different tissue components. EFFECT OF ISCHEMIA: Neither the ventricular nor the myocytic volume differed significantly from the respective control values. Both the myocytic mitochondrial volume (135+/ 8 vs control 89+/-6 microl) and the volume of myocytic clear space (35+/-6 vs control 10+/-2 microl) were significantly increased. The capillary volume (22+/-4 vs control 58+/-6 microl) and the volume of the capillary lumen (15+/-3 vs control 48+/-5 microl) were significantly decreased. The volume of the capillary wall, however, was not altered after exposure to ischemia (7+/-3 vs control 10+/ 1 microl). ADDITIVE EFFECT OF ISCHEMIA AND REPERFUSION: Both the ventricular volume (755+/-28 vs control 600+/-32 microl) and the myocytic volume (396+/-24 vs control 287+/-16 microl) were significantly increased after 10 min of reperfusion. EFFECT OF REPERFUSION: The ischemic-induced myocytic mitochondrial swelling and increase of clear space were not reinforced during reperfusion. Furthermore, the volume of the capillary lumen and the capillary wall did not alter significantly in the groups exposed to reperfusion compared to the ischemic hearts. In conclusion, stereological evaluation did not reveal significant aggravation of ischemic-induced myocardial injury during the early phase of reperfusion. PMID- 10533962 TI - Eosinophilic infiltration immediately following transplantation: recurrent hypersensitivity reaction? AB - Hypersensitivity myocarditis is a well-known complication of pharmaceutical therapy, often requiring heart transplantation. We report the unusual case of pre transplant hypersensitivity myocarditis with eosinophilic myocardial infiltration in the donor heart, demonstrated by needle biopsy at the time of transplant ('time-zero' biopsy). At first the myocarditic process was temptatively attributed to a pre-transplant pathology in the donor heart, but the close similarity between the pre-transplant and the post-transplant infiltrate and the clinical data of an eosinophilic peak of the recipient during the transplant procedure brought to the diagnosis of early recurrent hypersensitivity myocarditis. PMID- 10533961 TI - Enhanced embryonic nonmuscle myosin heavy chain isoform and matrix metalloproteinase expression in aortic abdominal aneurysm with rapid progression. AB - Abdominal aortic aneurysms (AAAs) are characterized by structural deterioration of aortic wall leading to progressive dilatation. The histopathological changes in AAAs are particularly evident within the elastic media, which is normally comprised mainly of vascular smooth muscle cells (SMCs). There are vascular myosin heavy chain (MHC) isoforms; SM2 is specifically expressed in differentiated SMCs and SMemb is a nonmuscle-type MHC abundantly expressed in SMCs of the fetal aorta with an immature phenotype. Although AAA altered expression of matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs), pathophysiological role of SMC phenotypic modulation in the AAA progression remains uncertain. To determine whether phenotypic modulation in vascular SMCs contributes to arterial medial degeneration, we examined MHC expression in SMCs of AAA. Aortic specimens were obtained from patients with slowly progressed AAA (n = 12) and rapidly progressed AAA (n = 5), and compared with normal aortic tissue (n = 3). Immunohistochemical staining was performed for detection of SMemb, SM2, MMP (types 2 and 9) and TIMP (types 1 and 2). Faint SMemb and abundant SM2 were observed in normal aorta, while the balance shifted to SMemb predominance in AAAs. Compared with slowly progressed AAA tissue, rapidly expanded AAA tissue demonstrated marked increases in SMemb expression with suppressed SM2. Predominant SMemb expression indicates presence of phenotypic modulated SMCs and enhanced MMP; while abundant TIMP was seen in mature SMCs expressing SM2. SMemb expression is markedly increased in AAA with MMP enhancement, and a significant imbalance between SMemb and SM2 results in rapid progression of AAA. PMID- 10533963 TI - The challenge of managed care in drug abuse treatment. PMID- 10533964 TI - A process evaluation of the San Francisco Target Cities Project in the first year. PMID- 10533965 TI - Creating more with less: implementation issues in the Newark Target Cities Project. PMID- 10533966 TI - Miami's Target Cities Project: past and present lessons. PMID- 10533967 TI - Establishing a Target Cities model in Cleveland. PMID- 10533968 TI - Target Cities as an effective solution to the special problems of treatment and recovery in the city of Laissez Le Bon Temps Roulle. PMID- 10533969 TI - Improving substance abuse treatment for indigent clients in Detroit. AB - It has been clearly established that substance abuse treatment works (De Leon 1988). Thus, activities which increase the proportion of indigent clients in Detroit who actually get into treatment and activities which help indigent clients stay in treatment are likely to significantly improve treatment outcomes. The Target Cities projects in general and the Detroit Target Cities project in particular represent some of the few efforts currently underway to determine intervention activities which significantly improve treatment outcomes for indigent substance abusing clients. Subsequent evaluation has shown that the proportion of clients referred by the CDRS who were actually admitted into a treatment program increased significantly after full implementation of the Detroit Target Cities screening and pretreatment case management activities. Furthermore, the average time between referral by the CDRS and admission into a treatment program decreased significantly (Tucker 1997). Evaluation of the Detroit Target Cities jail-based substance abuse treatment program also showed a significant increase in the proportion of clients who remained drug free after full implementation of the program (Tucker 1998). PMID- 10533970 TI - The Portland Target Cities Project: emerging patterns of service in a managed care environment. PMID- 10533971 TI - A chronology of the Dallas Target Cities model, with emphasis on the impact of a state funding crisis. PMID- 10533972 TI - Albuquerque Target Cities: preliminary findings. AB - Overall, there appeared to be a decline in frequency of use in the substance patterns of the 46 cases investigated for this analysis. The rising frequency with which prescription drugs are reported may be explained by the fact that these clients are in treatment, a situation which frequently necessitates the prescription of various medications. Additionally, the rise in methadone use can also be explained by the fact that it is prescribed to individuals who are undergoing treatment for a heroin addiction. Finally, it is probable that polysubstance use is underreported in the intake data, thus explaining the apparent increase reported at follow-up. PMID- 10533974 TI - Reorganizing publicly-funded drug abuse treatment: the experience of ten Target Cities Projects. PMID- 10533973 TI - The Boston Target Cities program: overview and evaluation results. PMID- 10533975 TI - Biopsychosocial characteristics and treatment outcomes of pregnant cocaine dependent women in residential and outpatient substance abuse treatment. AB - The purpose of this descriptive study was to compare the characteristics and treatment outcomes of pregnant cocaine-dependent women and their infants enrolled in residential (N=32) and outpatient (N=32) treatment settings. Biopsychosocial characteristics and issues that influenced the women's multiple treatment outcomes are highlighted. Comparisons of retention and infant birth outcomes found no significant differences between treatment programs, whereas abstinence and patterns of attrition showed meaningful differences favoring residential treatment. Further research is needed to evaluate whether the reported treatment outcomes are markers of improved life functioning that hold promise for the women in treatment, their families and the community. PMID- 10533977 TI - Evaluating corrections-based treatment for the drug-abusing criminal offender. AB - The recent increase in drug abusers in the criminal justice system has led to the expansion of corrections-based drug treatment facilities. Although three key evaluations have provided consistent support for the effectiveness of drug treatment within the criminal justice system, direct comparisons of outcomes across these evaluations are limited by variations in their measurement systems and the structure of official records on which they are based. This article addresses some of the issues relating to the assessment of treatment outcomes for the drug-abusing offender and provides several recommendations for future research. PMID- 10533976 TI - HIV needle risk behaviors and drug use: a comparison of crack-smoking and nonsmoking injection drug users in Ohio. AB - This study compares the drug use and needle risk behaviors among 733 crack smoking injection drug users (IDUs) and 518 nonsmoking IDUs. Participants were recruited in Dayton and Columbus, Ohio, for the Cooperative Agreement for AIDS Community-Based Outreach/Intervention Research Program from 1992 to 1996. Crack smoking IDUs were more likely to be male, African-American, and 30 to 40 years of age, but less likely to be married or living with a sex partner compared to nonsmokers. Daily crack users were less likely to be daily injectors but more likely to use alcohol daily when compared to non-crack users and less-than-daily crack smokers. IDUs who smoked crack less than daily were more likely to have injected with needles and syringes used by others. There is an urgent need for additional research on the relationship between drug injection and crack smoking as well as improved HIV risk-reduction interventions that include drug abuse treatment components focusing on issues surrounding crack-cocaine addiction. PMID- 10533978 TI - Detecting cocaine and opiates in urine: comparing three commercial assays. AB - Urine screening is a potentially useful tool for detecting drugs of abuse in treatment, criminal justice, and other human service settings. This article examines the relative accuracy and other features of three drug screening assays sold by commercial laboratories: (1) Abbott Diagnostics ADx machine and reagents; (2) ONTRAK, manufactured by Roche Diagnostics; and (3) EZ-SCREEN, manufactured by Environmental Diagnostics. Urine samples (n=345) were collected from indigent men and women in a work and life skills program, and tested for cocaine and opiates with each of the kits. The ADx fluorescent immunoassay was presumed to be the most sensitive and specific screening method, and comparisons with the two visually-determined test kits supported this assumption. Of the two visual test kits, ONTRAK was the more specific assay, and was lower in cost and simplest to perform. Decision makers could employ similar evaluative methods in selecting drug testing materials. PMID- 10533979 TI - Ionizing radiation stimulates existing signal transduction pathways involving the activation of epidermal growth factor receptor and ERBB-3, and changes of intracellular calcium in A431 human squamous carcinoma cells. AB - Previous studies demonstrated that ionizing radiation activates the epidermal growth factor receptor (EGFR), as measured by Tyr autophosphorylation, and induces transient increases in cytosolic free [Ca2+], [Ca2+]f. The mechanistic linkage between these events has been investigated in A431 squamous carcinoma cells with the EGFR Tyr kinase inhibitor, AG1478. EGFR autophosphorylation induced by radiation at doses of 0.5-5 Gy or EGF concentrations of 1-10 ng/ml is inhibited by >75% at 100 nM AG1478. Activation of EGFR enhances IP3 production as a result of phospholipase C (PLC) activation. At the doses used, radiation stimulates Tyr phosphorylation of both, PLCgamma and erbB-3, and also mediates the association between erbB-3 and PLCgamma not previously described. The increased erbB-3 Tyr phosphorylation is to a significant extent due to transactivation by EGFR as >70% of radiation- and EGF-induced erbB-3 Tyr phosphorylation is inhibited by AG 1478. The radiation-induced changes in [Ca2+]f are dependent upon EGFR, erbB-3 and PLCgamma activation since radiation stimulated IP3 formation and Ca2+ oscillations are inhibited by AG1478, the PLCgamma inhibitor U73122 or neutralizing antibody against an extracellular epitope of erbB-3. These results demonstrate that radiation induces qualitatively and quantitatively similar responses to EGF in stimulation of the plasma membrane associated receptor Tyr kinases and immediate downstream effectors, such as PLCgamma and Ca2+. PMID- 10533980 TI - Endothelin-1 effect on tyrosine phosphorylation and on tyrosine phosphatase (PTP 1C) translocation in rabbit platelets. AB - This study examined the temporal relationships of endothelin-1-stimulated rabbit platelets tyrosine phosphorylated proteins. The effect of endothelin-1 on tyrosine phosphorylation was dose- and time-dependent and caused a rapid tyrosine phosphorylation of three groups of proteins in the molecular mass range 70-100 kDa, 100-150 kDa and 150-200 kDa. Significant protein tyrosine phosphatase activity and amount were found to be associated with the cytoskeleton of endothelin-1-stimulated rabbit platelets. Under our experimental conditions, translocation from the cytosolic fraction to the cytoskeleton reached its highest levels within 10-20 sec of endothelin-1 stimulation. Endothelin-1-induced translocation of protein tyrosine phosphatase, associated with the increase in its activity was demonstrated by immunoblotting and immunoelectron microscopy. PMID- 10533981 TI - Use of bioluminescent aequorin for the pharmacological characterization of 5HT receptors. AB - A convenient functional assay for 5HT2a and 5HT2c receptors is reported utilizing the bioluminescent aequorin to detect intracellular calcium changes. Using this assay, the pharmacological properties of many 5HT ligands can be determined in a 96-well format. The data indicate that the aequorin detection method is superior to the inositol phosphate assay with regard to speed and scope. This system is also appropriate for kinetic studies of receptor desensitization. We showed that the human 5HT2c receptor desensitizes in a biphasic manner, with a fast desensitization of approximately 90% of the total response occurring within 15 minutes while the remaining 10% response remains for at least 3 hours. PMID- 10533982 TI - Chemotactic effect of urokinase plasminogen activator: a major role for mechanisms independent of its proteolytic or growth factor domains. AB - Urokinase type plasminogen activator (uPA) converts plasminogen to plasmin and is highly chemotactic for many cell types. We examined, using recombinant wild type and mutated forms of uPA, the extent to which its proteolytic properties, its growth-like domain (GFD) and/or interactions with the specific receptor (uPAR) contribute to the chemotactic activity towards vascular smooth muscle cells (SMC). Recombinant wild type uPA (r-uPA) stimulated cell migration nearly 5.8 fold, inactive r-uPA, with a mutation in the catalitic domain (r-uPA(H/Q)), 3 fold, uPA without growth factor like domain (r-uPA(GFD )), 2.6-fold, and a form containing both mutations (r-uPA(H/Q, GFD ), 3.3-fold. All recombinant forms of uPA, wild type and those with mutations were equally and highly effective (IC50 approximately 20 nM) in displacing 125I-r-uPA bound to SMC. These results indicate that additional mechanisms, not dependent on uPA's proteolytic activity or the binding ability of its GFD to uPAR, are the major contributors to its chemotactic action on SMC. PMID- 10533983 TI - The TrkB receptor tyrosine kinase regulates cellular proliferation via signal transduction pathways involving SHC, PLCgamma, and CBL. AB - The TrkB protein tyrosine kinase is a high affinity receptor for brain derived neurotrophic factor (BDNF) and neurotrophin-4 (NT-4). TrkB autophosphorylation occurs on five cytoplasmic tyrosines: Y484, Y670, Y674, Y675, and Y785. Using site directed mutagenesis, we have assessed the importance of TrkB tyrosines 484 and 785 in affecting TrkB-mediated signaling events leading to NIH 3T3 cell mitogenesis and survival. Mutation of TrkB tyrosine 484, while having no affect on BDNF-inducible PLCgamma and Cbl tyrosine phosphorylation, is essential for the phosphorylation of Shc, the complete activation of extracellular regulated kinase 1/2 (ERK1/2) and the induction of c-fos protein synthesis. In contrast, mutation of Y785 does not significantly affect BDNF-inducible Shc phosphorylation, ERK1/2 activation, or c-fos protein synthesis, but completely inhibits the tyrosine phosphorylation of PLCgamma and Cbl. These data indicate that both ERK-dependent and ERK-independent signaling pathways lead to BDNF-inducible mitogenesis and survival. PMID- 10533985 TI - Agonist induced conformation alteration of neurotensin receptor and the mechanism behind Na+ inhibition of 125I-NT binding. AB - In the absence of Na+, 125I-Neurotensin (125I-NT) binding to the Neurotensin receptor (NTR) produces a stable noncovalent 125I-NT-NTR complex whose dissociation rate is extremely low even after the addition of 1 microM NT, 100 microM SR48692 (antagonist), 100 microM GPPNHP or 100 mM NaCl. Lowering the medium pH to 4.5 enhances the process (approximately 70% in 10 minutes). Labeling by photoactivatable 125I-Tyr3-Azo4-NT identifies a approximately 50 KD Mr band along with several other minor components. Interestingly, the labeling intensity is drastically reduced when binding is performed in the presence of Na+ or GPPNHP. However, a minor reduction is noticed when Na+ or GPPNHP is added to the medium after binding. The binding kinetics indicates that Na+ lowers the rate of 125I-NT association by acting as a noncompetitive inhibitor. On the contrary, Na+ favors the interaction of antagonist, SR48692 by lowering the value of Ki. GTPgamma35S binding to membranes in the presence of 30 mM NaCl suggests that Na+ inhibition of 125I-NT binding is due to the uncoupling of NTR associated G protein(s). In order to explain the entire phenomenon, a two-step, binding model has been proposed. In Step-1, interaction between NT and NTR produces a transient complex, which attains a stable state in the absence of NaCl via step-2, thereby altering the native NTR conformation. The presence of Na+ prevents step-2 by dissociating the transition complex. PMID- 10533984 TI - Tyrosine kinase inhibition affects type 1 angiotensin II receptor internalization. AB - Growth factor receptors activate tyrosine kinases and undergo endocytosis. Recent data suggest that tyrosine kinase inhibition can affect growth factor receptor internalization. The type 1 angiotensin II receptor (AT1R) which is a G-protein coupled receptor, also activates tyrosine kinases and undergoes endocytosis. Thus, we examined whether tyrosine kinase inhibition affected AT1R internalization. To verify protein tyrosine phosphorylation, both LLCPKCl4 cells expressing rabbit AT1R (LLCPKAT1R) and cultured rat mesangial cells (MSC) were treated with angiotensin II (Ang II) [1-100 nM] then solubilized and immunoprecipitated with antiphosphotyrosine antisera. Immunoblots of these samples demonstrated that Ang II stimulated protein tyrosine phosphorylation in both cell types. Losartan [1 microM], an AT1R antagonist, inhibited Ang II stimulated protein tyrosine phosphorylation. LLCPKAT1R cells displayed specific 125I-Ang II binding at apical (AP) and basolateral (BL) membranes, and both AP and BL AT1R activated tyrosine phosphorylation. LLCPKAT1R cells, incubated with genistein (Gen) [200 microM] or tyrphostin B-48 (TB-48) [50 microM], were assayed for acid-resistant specific 125I-Ang II binding, a measure of Ang II internalization. Both Gen (n = 7) and TB-48 (n = 3) inhibited AP 125I-Ang II internalization (80+/-7% inhibition; p<0.025 vs. control). Neither compound affected BL internalization. TB-1, a non-tyrosine kinase-inhibiting tyrphostin, did not affect AP 125I-Ang II endocytosis (n = 3), suggesting that the TB-48 effect was specific for tyrosine kinase inhibition. Incubating MSC with Gen (n = 5) or herbimycin A [150 ng/ml] (n = 4) also inhibited MSC 125I-Ang II internalization (82+/-11% inhibition; p<0.005 vs. control). Thus, tyrosine kinase inhibition prevented Ang II internalization in MSC and selectively decreased AP Ang II internalization in LLCPKAT1R cells suggesting that AP AT1R in LLCPKAT1R cells and MSC AT1R have similar endocytic phenotypes, and tyrosine kinase activity may play a role in AT1R internalization. PMID- 10533986 TI - Children with recurrent abdominal pain: issues in the selection and description of research participants. AB - Research criteria proposed by Apley for identification of children with recurrent abdominal pain (RAP) have been widely adopted, but many researchers have deviated from the original definition. Ambiguities in Apley's criteria and problems caused by departures from his definition are discussed in this article. Many inconsistencies in research results may have been created by the decision of some researchers to exclude children with presumed organic causes for their pain from RAP study samples. Further precision and comparability of research results may be gained by using the two-stage approach to classification that is proposed in this article. The first stage requires no medical evaluation but is based strictly on the correspondence of RAP symptoms to temporal and severity criteria. At the second stage, subgroups of RAP are specified by using results from medical assessments or detailed patterns of symptoms. It is also suggested that researchers specify the temporal features of RAP and provide measures of impairment. Like Apley's criteria, the revised approach to classification allows the comparison of community- and school-based samples with children who have undergone medical evaluation for RAP. PMID- 10533987 TI - Infant colic and childhood recurrent abdominal pain syndromes: is there a relationship? PMID- 10533988 TI - Childhood recurrent abdominal pain and subsequent adult irritable bowel syndrome. PMID- 10533989 TI - Pathways between recurrent abdominal pain and adult functional gastrointestinal disorders. PMID- 10533990 TI - Coping and responses to stress among children with recurrent abdominal pain. PMID- 10533991 TI - Prenatal cocaine and neuromotor outcome at four months: effect of duration of exposure. AB - The effect of prenatal cocaine exposure on the motor development of full-term infants was examined in a prospective study, controlling for maternal characteristics and exposure to other substances. Intrauterine cocaine exposure was determined at birth by maternal self-report and was verified by hair analysis. At 4 months, 120 cocaine-exposed (COC) and 186 non-cocaine-exposed (NON COC) infants were assessed by blinded examiners using a standard evaluation of neuromotor function, the Movement Assessment of Infants (MAI). Relative to NON COC infants, COC infants had significantly higher full-scale MAI total risk scores after adjusting for covariates (p = .05). Infants exposed through the third trimester of pregnancy (n = 48) had higher MAI scores for both total risk (p = .02) and Volitional Movement (p = .01), and when compared with infants exposed only within the first two trimesters (n = 72), they had significantly more deficits in Volitional Movement (p = .03). Although MAI scores for the majority of exposed infants were within the normal range, infants exposed through the third trimester were at significantly increased risk for motor dysfunction (relative risk = 1.6; 95% confidence interval = 1.1, 2.8). Intrauterine cocaine exposure had an adverse effect on infant motor development after the neonatal period; this association was related to the timing and duration of gestational exposure. Further study is needed to evaluate the long-term clinical implications of neuromotor abnormalities in prenatally exposed infants. PMID- 10533992 TI - Knowledge of physician prescriptions and adherence to treatment among children with cystic fibrosis and their mothers. AB - This investigation examined factors related to adherence to treatment regimens for children with cystic fibrosis (CF) and their mothers. Subjects were 45 children with CF who ranged in age from 6 to 10 years and their mothers. Findings revealed that children's and parents' reports of level of adherence were related to their knowledge of the specific details associated with medically prescribed treatments. In this sample, 12% to 32% of mothers did not have an accurate understanding of physician recommendations for their children's treatments. When controlling for individual differences in the prescribed treatment regimens, parents' and children's knowledge of what had been prescribed accounted for a significant portion of the variance in the children's reported treatment-related behaviors. Results are discussed in terms of implications for future intervention research aimed at enhancing adherence to treatment as well as for future directions for clinical efforts in this area. PMID- 10533993 TI - Motor organization in very low birth weight infants during caregiving: effects of a developmental intervention. AB - The purpose of this study was to determine whether an individualized approach to handling very low birth weight (VLBW) infants designed to support development would result in less motor disorganization than the task-oriented approach in traditional use. Using a quasi-experimental crossover design, motor responses were investigated in 38 infants (< or = 1700 g, 53% male, 89% white) observed at 28, 32, and 36 weeks post-conceptional age. Subjects served as their own controls. Motor responses were coded from direct observation and videotapes. Results demonstrated that during developmental handling, (1) the overall amount of movement was less, the number of organized movements was greater, and the number of disorganized movements was less than during traditional handling; and (2) the amount of movement increased over time, but in the traditional condition, it peaked at 32 weeks. Results support positive effects of developmental handling and suggest the potential for reducing the prevalence of minor motor impairments in VLBW infants. PMID- 10533994 TI - Effects of baby walkers on motor and mental development in human infants. AB - Because baby walkers enable precocious locomotion in very young, otherwise prelocomotor infants, walker experience might be conceptualized in terms of early enrichment. However, walker devices prevent visual access to the moving limbs by design. Therefore, prelocomotor walker experience may be conceptualized in terms of early deprivation, reminiscent of that created in a classic series of animal experiments on the critical role of visual feedback in developing motor systems. This study analyzed motor and mental development in 109 human infants, with and without walker experience, between the ages of 6 and 15 months. Walker experienced infants sat, crawled, and walked later than no-walker controls, and they scored lower on Bayley scales of mental and motor development. Significant effects of walker type, frequency, and timing of walker exposure were observed. Considering the injury data along with the developmental data, the authors conclude that the risks of walker use outweigh the benefits. PMID- 10533995 TI - Perceived role restriction and depressive symptoms in mothers of children with chronic health conditions. AB - This study examined role restriction in 365 inner-city mothers of 5- to 8-year old children with chronic health conditions and tested whether it could account for a previously reported relationship between children's functional limitations and maternal psychological distress. Functional limitations in the children were related to maternal role restriction with sociodemographic factors controlled. Children's functional limitations also independently predicted maternal Depression subscale scores in a regression model. Adding role restriction to this model significantly increased explained variance in Depression scores, indicating that it also is directly related to maternal distress symptoms. However, adding role restriction only slightly reduced the impact of functional limitations in the model, suggesting that it plays a small role, if any, in explaining the relationship between the other two variables. Because perceived role restriction independently predicts maternal depressive symptoms and represents a potentially modifiable risk factor, it warrants attention as a useful target for intervention. PMID- 10533996 TI - Identifying fetal alcohol syndrome among youth in the criminal justice system. AB - A disproportionately large number of youth and adults with fetal alcohol syndrome (FAS) and fetal alcohol effects (FAE) seem to be coming into conflict with the legal system. Learning and behavioral difficulties associated with FAS/FAE may make them more susceptible to criminal behavior. This study determined the prevalence of FAS/FAE among youth who were remanded for a forensic psychiatric/psychological assessment. All youth remanded to a forensic psychiatric inpatient assessment unit over a 1-year period were evaluated for FAS/FAE. Of the 287 youth, 67 (23.3%) had an alcohol-related diagnosis: 3 (1.0%) had a diagnosis of FAS and 64 (22.3%) had a diagnosis of FAE. Thus, this group is disproportionately represented in the juvenile justice system, indicating the need for increased education and awareness among those in the criminal justice system involved with these youth. PMID- 10533997 TI - Longitudinal follow-up of the intellectual and academic functioning of children receiving central nervous system-prophylactic chemotherapy for leukemia: a four year final report. AB - This longitudinal investigation extends our prospective study of the intellectual and academic functioning of children treated for cancer to 4 years after diagnosis. In the longer term, the children who received central nervous system (CNS) chemotherapy experienced greater neurocognitive deficits, particularly in the area of academic achievement, than did the children who did not receive CNS chemotherapy. Specifically, the CNS chemotherapy-treated children scored lower on academic tests of reading at 3 and 4 years after diagnosis. The results suggest that CNS chemotherapy prophylaxis may adversely effect the development of higher order mental abilities and cognitive skills during the late-effects period and may also impair academic achievement. PMID- 10533998 TI - Changes in the practice of child and adolescent psychiatry: are our patients better served? PMID- 10533999 TI - A school-aged child with delayed reading skills. AB - During a health supervision visit, the father of a 7.5-year-old African American second-grader asked about his son's progress in reading. He was concerned when, at a recent teacher-parent conference to review Darren's progress, the teacher remarked that Darren was not keeping up with reading skills compared with others in his class. She said that he had difficulty sounding out some words correctly. In addition, he could not recall words he had read the day before. The teacher commented that Darren was a gregarious, friendly child with better-than-average verbal communication skills. His achievement at math was age-appropriate; spelling, however, was difficult for Darren, with many deleted letters and reversals of written letters. A focused history did not reveal any risk factors for a learning problem in the prenatal or perinatal periods. Early motor, language, and social milestones were achieved on time. Darren had not experienced any head injury, loss of consciousness, or chronic medical illness. He had several friends, and his father denied any behavioral problems at home or at school. His teacher completed a DSM-IV-specific behavioral survey for attention deficit/hyperactivity disorder (ADHD). It did not show any evidence of ADHD. Darren's father completed 1 year of college and is currently the manager of a neighborhood convenience store. His mother had a high school education; she recalled that she found it difficult to complete assignments that required reading or writing. She is employed as a waitress. Darren does not have any siblings. The pediatrician performed a complete physical examination, the results of which were normal, including visual acuity, audiometry, and a neurological examination. It was noted that Darren seemed to pause several times in response to questions or commands. On two occasions, during finger-nose testing and a request to assess tandem gait, directions required repetition. Overall, he was pleasant and seemed to enjoy the visit. His pediatrician concluded that he had a learning problem but she was uncertain about the next step. She asked herself, "Is there anything else I can do in the office to evaluate Darren's problem with learning? Should I quickly refer him for educational testing or encourage a reading tutor? What questions can I ask his teacher that would be helpful? Am I missing a medical disorder?" PMID- 10534000 TI - Purification and partial characterization of a cadmium-binding protein from the liver of rainbow trout (Onchorynchus mykiss). AB - This study describes the isolation and partial characterization of a low molecular weight (approximately 14 kDa), cadmium-binding protein from rainbow trout (Onchorynchus mykiss) liver. Rainbow trout were injected intraperitoneally with 3.5 mg/kg cadmium chloride (total body dose) twice weekly for 3 wk. Livers were removed and a cadmium-binding protein was isolated. Monoclonal antibodies produced against this protein were used in the affinity purification process. Amino acid analysis showed the protein contained 3.8 mol% cysteine, 3.5 mol% phenylalanine, 2.2 mol% tyrosine and 1.9 mol% histidine. The low cysteine content suggests that it was distinct from metallothionein. The monoclonal antibodies were also used to identify the protein in liver homogenates from both cadmium exposed and control fish and in the testes of cadmium-exposed mice lacking the gene for both metallothionein-1 and metallothionein-II. The compound identified in this study represents a non-metallothionein cadmium-binding protein that appears to be highly conserved. PMID- 10534001 TI - Detection and comparison of nitric oxide in clinically normal horses and those with naturally acquired small intestinal strangulation obstruction. AB - The purpose of this study was to determine whether nitric oxide (NO) is present in clinically normal horses under basal conditions and if it increases secondary to naturally acquired small intestinal strangulation obstruction. Thirty-one horses were used; 20 horses with naturally acquired small intestinal strangulation obstruction and 11 clinically normal horses with no signs of gastrointestinal tract disease. Jugular venous blood, abdominal fluid, and urine were collected for NO quantification. Plasma, abdominal fluid, and urine were stored at -70 degrees C until analyzed for NO using a chemiluminescent method. Biopsy specimens collected from the affected jejunal segment, during anesthesia or after immediately after euthanasia, or from the midjejunum of control horses, were divided into subsections for fixation in zinc formalin and cryopreservation in OCT gel. Nicotinamide adenine dinucleotide phosphate (reduced) (NADPH) diaphorase histochemical stains were performed on cryopreserved tissues and inducible nitric oxide synthase (iNOS) and nitrotyrosine immunohistochemical stains were performed on formalin-fixed, paraffin-embedded tissues. There were significantly greater plasma and abdominal fluid NO concentrations in affected horses as compared with controls, but there were no significant differences between horses for urine NO concentrations. There was a significant decrease in NADPH diaphorase stain in mucosal epithelium, vasculature, and leukocytes, and in submucosal plexi in affected horses compared with control horses. There was a significant increase in iNOS staining in mucosal and submucosal leukocytes and in mucosal leukocyte nitrotyrosine staining of the affected compared with control horses. Endothelial NOS and neuronal NOS are present under basal conditions in the jejunum of horses and probably mediate physiologic or cytoprotective effects. Plasma and abdominal fluid, but not urine, NO concentrations increase subsequent to small intestinal strangulation obstruction; this may be associated with increased mucosal and submucosal iNOS staining in leukocytes, which was likely due to increased expression subsequent to stimuli associated with ischemia. The increased nitrotyrosine staining in mucosal leukocytes of affected horses likely reflects the presence of peroxynitrite subsequent to increased NO and superoxide production and may reflect a cytotoxic role of NO in small intestinal strangulation obstruction in horses. PMID- 10534002 TI - Effects of muscle glycogen depletion on some metabolic and physiological responses to submaximal treadmill exercise. AB - The aim of this study was to investigate the effects of reduced muscle glycogen concentration on some physiological and metabolic responses during moderate intensity treadmill exercise in horses. Six Thoroughbred geldings were randomly allocated to 2 treatments (protocols A and B) or control in a 3 x 3 replicated Latin square design. In protocol A, horses performed low intensity exercise while horses in protocol B performed short bursts of high intensity exercise. Protocol A was designed to induce glycogen depletion mainly of slow twitch muscle fibers while protocol B aimed to deplete mainly fast twitch muscle fibers. Horses in the control group did not undergo exercise prior to the exercise test. Five hours after glycogen depletion, horses performed treadmill exercise at 60% VO2max at a treadmill slope of 10% until fatigue (20-30 min). The induced glycogen depletion prior to exercise had no significant effect on plasma glucose, insulin, or lactate concentrations during the exercise test, and there was no effect on glycogen utilization rate, although respiratory exchange ratios were lower in the glycogen-depleted groups. The VO2, heart rate and central blood temperature did not vary significantly between the protocols A and B and control throughout the exercise test. It was concluded that 20-30% depletion of glycogen concentration in the middle gluteal muscle resulted in a shift towards fat metabolism, but does not significantly affect heart rate, oxygen uptake, or concentrations of plasma glucose and lactate during moderate intensity exercise. PMID- 10534003 TI - Evaluation of lansoprazole (an H+/K+-ATPase inhibitor) and azithromycin (an antibiotic) for control of gastric ulceration in swine during periods of feed deprivation. AB - Helicobacter-like organisms as well as fermentative bacteria have been implicated in gastric ulcer production in swine. Irregular feeding schedules are also considered a major risk factor. A research trial was conducted to determine whether medication with an acid secretion inhibitor (lansoprazole), either alone or in combination with an antibiotic (azithromycin), would protect pigs from gastric ulceration if the animals were subjected to a 48 h period of fasting. In a 2 x 3 factorial design, 48 pigs were fasted, while an equal number were fed ad libitum. Within these 2 study groups, pigs were randomly assigned to 1 of 3 treatments: control, 30 mg lansoprazole s.i.d. for 7 d, or lansoprazole (30 mg s.i.d. for 7 d) and azithromycin (500 mg s.i.d. for 3 d). Overall, fasted pigs were 1.9 times more likely to develop erosive or ulcerative lesions of the pars esophagea (chi2 = 9.89, P < 0.002). Treatment with an acid secretion inhibitor alone or in combination with an antibiotic did not protect pigs from developing gastric lesions. Helicobacter-like organisms were not detected in any of the stomachs. Possibly, the lansoprazole dose of 30 mg given once per day was insufficient to prevent pH levels from becoming low enough to cause damage to epithelial tissue. Alternatively other substances such as bile acids may have caused the ulcerative lesions, even though stomach acid production was suppressed. PMID- 10534004 TI - Prognostic factors affecting survival of 507 horses with joint disease: (1983 to 1990). AB - Between July 1, 1983 and December 31, 1990, risk factors were determined for all horses with joint disease presented to a referral center, of being discharged, of ever becoming sound, or of being alive at 3 mo follow-up. Logistic multiple regression models were done separately for foals (< or = 4 mo), yearlings (> 4-24 mo) and racing or nonracing adult horses (> 24 mo). The breakdown in this study was 53 foals, 87 yearlings, 141 nonracing adults, and 226 racing adults. Thirty one foals (58%), 68 yearlings (78%), 119 non-racing adults (84%), and 213 racing adults (94%) were discharged. Foals with a less severe lameness, duration of illness of > 1 d, and infectious arthritis had increased odds of discharge. At follow-up, 12 of 18 (67%) were alive, 10 (56%) of which were sound. Yearlings with osteochondrosis had higher odds of discharge; at follow-up, 38 of 49 (78%) were alive, 32 (65%) of which were sound. For non-racing adults, horses with less severe lameness, without a miscellaneous diagnosis, or intended for pleasure use had increased odds of discharge. At follow-up, 55 of 78 (70%) were alive and 33 of 58 (57%) with soundness data became sound. Risk factors for higher odds of being alive at follow-up were carpal lameness, arthroscopic surgery, a prognosis other than poor, became sound, above-median hospitalization costs, and duration of follow-up. The 161 racing adults (76% of discharges), with follow-up, were more likely to have had osteoarthritis, higher hospital costs, hospitalization > 1 d, and arthroscopy. Sixty-four (60%) of these became sound; the odds increased if the horse was not severely lame at admission or was hospitalized for > 1 d. Risk factors and prognosis differed by age-use group among horses seen at our hospital. PMID- 10534005 TI - Effect of administration of oat beta-glucan on immune parameters of healthy and immunosuppressed beef steers. AB - In order to assess the effect of oat beta-glucan (ObetaG) administration on immune parameters of beef steers, 3 experiments were carried out. In experiment 1, the in vitro effect of ObetaG on the proliferation of blood lymphocytes, with or without the presence of dexamethasone (DXM), was evaluated. In experiment 2, groups of 12 healthy steers were administered ObetaG or saline solution and immunized with ovalbumin (OVA). Immune parameters studied included IgG antibody levels to OVA, proliferation responses of blood lymphocytes to OVA, and blood leukocyte differential cell counts. For experiment 3, groups of 10 steers were treated with ObetaG and DXM, DXM only, or saline solution, and immunized with OVA and keyhole limpet hemocyanin (KLH). Serum antibody responses to OVA and KLH, serum IgG concentration levels, blastogenic responses of blood lymphocytes to OVA and KLH, differential blood leukocyte numbers, and iron and zinc concentration in serum were tested to evaluate the effect of ObetaG to overcome immunosuppression. The in vitro treatment of naive blood lymphocytes with ObetaG did not increase their ability to proliferate; however, when ObetaG was added to cultures of DXM treated lymphocytes, a significant (P < 0.05 to P < 0.001) reversion of the immunosuppressive effect of DXM occurred. Administration of ObetaG to clinically healthy steers did not induce significant changes on any of the immune parameters studied. The administration of ObetaG to DXM-treated steers provoked, on Day 25, a significant increase in IgG anti-OVA (P < 0.01) and anti-KLH (P < 0.05) responses vs the DXM only group. On Day 25, the specific proliferation responses of lymphocytes, to both OVA and KLH, were significantly increased (P < 0.05) in ObetaG+DXM group compared to DXM group. On Day 4, a significant increase in the number of leukocytes (P < 0.01) and neutrophils (P < 0.001), and a significant decrease in the number of monocytes (P < 0.05) were observed in the group treated with DXM only compared to ObetaG+DXM group. No significant differences were observed in iron and zinc concentration between ObetaG+DXM and DXM groups. These results indicated that ObetaG did not influence immune responses of naive cells in vitro or of healthy steers in vivo; however, when cells or animals were treated with DXM, ObetaG significantly restored some of the specific and non specific immune parameters studied. PMID- 10534006 TI - Experimental exposure of young pigs using a pathogenic strain of Streptococcus suis serotype 2 and evaluation of this method for disease prevention. AB - Control of Streptococcus suis infections and associated disease have proven to be a difficult challenge under most farm conditions. The objective of this study was to experimentally expose young pigs with a pathogenic strain of S. suis serotype 2 as a means of controlling the disease in a commercial swine farm. Prior to the start of the study, the pathogenic S. suis strain responsible for mortality in the farm was identified and used to experimentally inoculate baby piglets. Over a 3-week period, groups of pigs were selected (100 pigs/wk) and divided into 2 groups: control (50 pigs/week) and experimentally exposed (50 pigs/week). Pigs in the experimentally exposed group were inoculated at 5 d old by tonsillar swabbing with the pathogenic S. suis farm isolate. The effect of exposure with this pathogenic strain was evaluated during the nursery and finishing stages and was based on: morbidity (pigs with central nervous signs (CNS) and/or lameness), mortality and number of treatments required by pigs that had either CNS or lameness. The relative risk (RR) of acquiring disease due to S. suis infection was also calculated. Results showed that morbidity in the experimentally exposed groups was lower than in the control group and these results were statistically different (P = 0.006). Experimentally exposed pigs also showed a statistically significant reduction in lameness problems (P = 0.012), but not in CNS (P = 0.20) or mortality (P = 0.59). Pigs in the control group had an increased RR of 4.76, 8.77 and 2.7 for morbidity, to have lameness or to have CNS signs, respectively. In conclusion, experimental exposure of young pigs with the farm's pathogenic S. suis strain at a young age, had a positive effect in reducing clinical signs characteristics of S. suis infection. This method constitutes a novel approach to the control of S. suis infections in swine farms. PMID- 10534007 TI - Nested reverse transcriptase-polymerase chain reaction (RT-PCR) for typing ruminant pestiviruses: bovine viral diarrhea viruses and border disease virus. AB - A nested reverse transcription (RT) polymerase chain reaction (PCR) assay was evaluated for differentiating reference bovine viral diarrhea virus (BVDV) strains, BVDV from diagnostic accessions, modified-live virus (MLV) BVDV strains in bovine viral vaccines, and a reference border disease virus (BDV). The detection level of this assay was compared to viral infection in cell culture. The PCR assay was used to distinguish 3 ruminant pestiviruses, types 1 and 2 BVDV, and type 3 BDV. The consensus (first) PCR assay detected all 3 ruminant pestiviruses, a result of the shared sequence homology. The consensus PCR product was subjected to a second (nested) PCR which used type-specific primers. The nested PCR was able to differentiate the 3 ruminant pestiviruses. Viral stocks of BVDV were diluted 10-fold and processed for the 2-step PCR assay. The sensitivity of this 2-step PCR assay was compared to viral infectivity in cell culture based on identical volumes of the system tested (cell culture assay and processing for RNA). The RT-PCR type-specific assay differentiated BVDV laboratory reference strains (12), diagnostic laboratory isolates (15), 2 MLV BVDV vaccine strains, and a BDV strain. The 30 ruminant pestiviruses typed included: (1) 27 reference strains and diagnostic laboratory isolates; 18 cytopathic (CP) type 1 strains, 3 CP type 2 strains, 3 noncytopathic (NCP) type 1 strains, and 3 NCP type 2 strains; (2) 2 MLV strains, type 1; and (3) 1 CP BDV type 3. The PCR assay had a detection limit of 10 TCID50/0.025 mL of virus when 3 separate BVDV were tested. This 2 step RT-PCR assay would be useful for the typing of ruminant pestiviruses, particularly BVDV isolates from the diagnostic laboratory. PMID- 10534008 TI - Nuclear proliferation in syncytia during avian reovirus replication. AB - Cultured chick embryonic fibroblasts formed syncytia after infection with avian reovirus (ARV) strain 58-132. Mitotic figures were occasionally observed within the syncytia. In addition, many nuclei in the syncytia incorporated 5-bromo-2' deoxyuridine (BrdU), a DNA replication marker, indicating that they were in the S phase of the cell cycle. These observations suggested that the nuclei within ARV induced syncytia originated from nuclear endomitosis without cell division, as well as from cell fusion. PMID- 10534009 TI - The adjuvant effect of a single dose of interleukin-12 on murine immune responses to live or killed Brucella abortus strain RB51. AB - This study was designed to determine if a single 0.5 microg administration of recombinant murine interleukin-12 (IL-12) would influence immune responses of mice vaccinated with live or killed Brucella abortus strain RB51 (SRB51). Mice were vaccinated intraperitoneally with 5 x 10(8) cfu of live or gamma-irradiated SRB51 bacteria alone, or in combination with 0.5 microg of IL-12. Control mice received saline or 0.5 microg of IL-12. Serologic responses and spleen weights after vaccination were greater in mice vaccinated with live SRB51 when compared to mice receiving killed SRB51 or control treatments. Administration of a single dose of IL-12 as a vaccine adjuvant did not influence immune responses, clearance of live SRB51, or resistance against B. abortus strain 2308 (S2308) challenge. The results of this study suggest that a single administration of 0.5 microg of IL-12 at the time of vaccination does not have significant adjuvant effects on vaccine-induced immune responses against live or killed Brucella. PMID- 10534010 TI - Effect of longeing and glucosamine supplementation on serum markers of bone and joint metabolism in yearling quarter horses. AB - The effect of longeing and glucosamine supplementation on known biological markers of joint disease was studied in yearling quarter horses. Twenty-one yearling quarter horses were randomly assigned to one of 4 treatments: 1) longeing (longeing 20 min daily) supplement control (LN); 2) longeing/glucosamine (LG); 3) walking (mechanical walker for 120 min daily (WN)); and 4) walking/glucosamine (WG). Oral glucosamine was administered at 5.5 g b.i.d. weeks 1-4, 3.5 g b.i.d. during weeks 5-6, and 2.0 g b.i.d. during weeks 7-8. Serum was obtained weekly for 8 wk and analyzed for keratan sulfate and osteocalcin concentrations. Walked horses receiving glucosamine showed slight elevation in serum keratan sulfate compared to controls (P = 0.04). Glucosamine or longeing exercise had no significant effect (6 > or = 0.08) on serum osteocalcin concentrations. Under these conditions, longeing and/or glucosamine supplementation did not significantly alter serum concentrations of keratan sulfate or osteocalcin. PMID- 10534011 TI - Theoretical aspects of neuroplasticity. AB - The authors propose an integrative theory of the organization of neuroplastic processes. Neuroplasticity is assumed to be one of the essential characteristics of the nervous tissue which may be manifested comparatively rapidly and result in reversible changes (functional plasticity). It may also modulate the expression of genotype into phenotype (adaptation) and thus bring about long-lasting effects. Neuroplastic mechanisms are triggered by various natural or artificial stimuli, which may arise in the internal or external environment, and they may differ quantitatively or qualitatively. The effects of plasticity can lead to either positive or negative changes during development (evolutionary plasticity), after short-term exposition (reactive plasticity), after long-term or continuous stimuli (adaptational plasticity), and during functional or structural recovery of damaged neuronal circuits (reparation plasticity). Manifestations of plasticity have probably the same basis, irrespective of the cause which triggered them or the brain region where they were accomplished. Neuroplastic mechanisms are based on the modulation of signal transmission across synapses. They can be related to interneuronal relations. The resulting changes may occur in the communication between neurons (synaptic level), in the activity of local neuronal circuits (at the level of local circuits) or in the relations between individual functional brain systems (multimodular level). PMID- 10534012 TI - Association analysis of 24-h blood pressure records with I/D ACE gene polymorphism and ABO blood group system. AB - The purpose of this study was to analyze the association between 24-h blood pressure parameters, insertion/deletion polymorphism of the angiotensin I converting enzyme gene and the ABO blood group system in a sample of the general Czech population. Fourteen parameters describing the 24-h blood pressure readings were obtained by analyzing blood pressure records in 243 volunteers, 119 men and 124 women. These parameters were adjusted for sex and body mass index (BMI) by means of multiple regression test. All subjects were genotyped for the insertion/deletion polymorphism of angiotensin I converting enzyme (I/D ACE) gene and routinely examined for the blood group phenotype in the ABO system. An association was found between the interaction of I/D ACE gene polymorphism and ABO blood group system on the one hand and mean values of systolic (p=0.016) or mean arterial (p=0.027) blood pressures and the phase shift of 24-h BP rhythm (p=0.036) on the other hand. Among three I/D ACE variants the DD genotype was associated with the highest values of mean blood pressure in blood group A and AB carriers. The same genotype was associated with the lowest blood pressures in blood group B and O carriers. In subjects with the DD genotype, the earlier daily position of the maximum of 24-h BP rhythm was found in blood group B, AB and O carriers. On the contrary, blood group A was associated with the latest position of maximum of the 24-h BP rhythm in the DD genotypes. PMID- 10534013 TI - Effect of somatotropin on adipose tissue net glucose-stimulated lipogenesis in young goats. AB - Net glucose-stimulated lipogenesis (NGSL: the rate of lipogenesis in the presence of glucose minus the rate of lipogenesis in the absence of glucose) in omental adipose tissue explants from young castrated male goats was evaluated in control animals (n = 3; placebo-treated) and in animals treated with the sustained release of recombinant bovine somatotropin (n = 4; bST; 100 mg at 7-day intervals in a 147 days lasting experiment). The rate of fatty acid synthesis was determined in acute incubations in both freshly prepared and chronically cultured explants. Adipose explants remained metabolically active and retained their ability to respond to hormones when maintained in a tissue culture medium. NGSL in explants cultured for 24 h in the presence of insulin alone or bST alone, was non-significantly increased (more in the controls) and decreased (more in bST treated animals), respectively. However, cortisol alone decreased (P<0.05) NGSL in explants from both control and bST-treated animals. In tissues from bST treated animals, cortisol acted synergistically with insulin to produce a higher rate of NGSL than that observed in cultures with insulin alone. bST inhibited insulin plus cortisol-stimulated lipogenesis significantly (P<0.05) in explants from bST-treated animals but non-significantly in control animals. The rates of NGSL were decreased (P<0.05) by catecholamines in explants from both control and bST-treated animals. Norepinephrine (NE) and isoprenaline (ISO) were equally effective in the controls, whereas isoprenaline was more effective than norepinephrine in bST-treated animals. PMID- 10534014 TI - Lipolysis induced by alloxan in rat adipocytes is not inhibited by insulin. AB - Isolated rat adipocytes were incubated with adrenaline, adrenaline plus insulin, alloxan or alloxan plus insulin. Glycerol release was taken as a measure of lipolysis. It was observed that alloxan in the concentration of 3, 10 and 20 mmol/l intensifies lipolysis in adipocytes in the absence of adrenaline. Insulin (10(-6) mol/l) treatment of cells did not inhibit lipolysis caused by this compound, but significantly restricted lipolysis induced by adrenaline (10(-6) mol/l). It was also shown that alloxan in the concentration of 3 and 10 mmol/l intensified lipolysis stimulated by adrenaline (10(-6) mol/l). Addition of 20 mmol/l of alloxan strongly inhibited glycerol release in the presence of adrenaline. The results presented here clearly indicate that the action of alloxan concerns cells of the white adipose tissue. PMID- 10534015 TI - Inhibitory effect of gossypol on basal and luteinization factor-stimulated progesterone synthesis in porcine granulosa cells. AB - Gossypol, a polyphenolic aldehyde, inhibits steroidogenesis and the reproductive system in both sexes. The present study was undertaken to investigate whether gossypol may affect progesterone biosynthesis in cultured porcine granulosa cells isolated from small (1-2 mm) follicles (SGC). SGC were cultured with gossypol, NO donor S-nitroso-N-acetylpenicillamine (S-NAP) or the specific NO-synthase inhibitor N-nitro-L-arginine methyl ester (L-NAME), in the presence or absence of follicular fluid isolated from large (5-8 mm) follicles (LFF) or conditioned media (CM) of granulosa cells isolated from large follicles (LGC). Gossypol enhanced the nitrite content in culture media of SGC and inhibited basal progesterone secretion by SGC. S-NAP (10(-3) M) inhibited progesterone secretion and enhanced the formation of cGMP by SGC. L-NAME had no effect on progesterone accumulation by SGC. The stimulatory effect of LFF or CM media on progesterone production by SGC in culture was also inhibited by S-NAP (10(-3)) and gossypol (10(-4) M). Moreover, gossypol inhibited forskolin-stimulated progesterone secretion, as well as substrate-enhanced conversion of 22-OH-cholesterol and pregnenolone to progesterone. These results suggest that the inhibitory effect of gossypol on progesterone secretion in culture of SGC may be mediated via NO generation. PMID- 10534016 TI - Serum opioid activity after physical exercise in rats. AB - In order to study the effect of exercise on the total serum opioid activity, female rats were trained for 3 weeks on a motor-driven treadmill and the experiment was ended by a final strenuous run until exhaustion. The serum samples were taken immediately after the final run and were analyzed by radioreceptor assay. Despite considerable interindividual variations, serum opioid activity, expressed in met-enkephalin equivalents (ME eq +/- S.D.), was significantly higher in the exercising group (74.5+/-50.5 pmol ME eq/ml) than in the control group (35.7+/-20.2 pmol ME eq/ml). Because of the much lower molar levels of beta endorphin and met-enkephalin, this result suggests that many other opioid peptides might be involved in that increase. PMID- 10534018 TI - Ganglioside content and composition in rat cerebellum after prolonged diazepam treatment. AB - The main purpose of this study was to determine the content and composition of cerebellar gangliosides after prolonged diazepam treatment and their possible recovery after diazepam withdrawal. Male Wistar rats were administered diazepam in a dose of 10 mg/kg/day in drinking water for 3, 5 or 6 months. A additional group of rats had a one-month recovery period after five months of diazepam treatment. Control animals were age-matched and pair-fed. At the end of the experiment, the animals were sacrificed and the total cerebellar contents of ganglioside-NeuAc as well as its content in particular ganglioside fractions were estimated. After three months of diazepam consumption, no changes of ganglioside NeuAc in investigated fractions (G(Q1b), G(T1b), G(D1b), G(D1a), G(M1), G(M2), and G(M3)) were observed. Five months of diazepam treatment caused a significant decrease in the total amount of gangliosides, which was evident in most of the investigated fractions, with the exception of the monosialoganglioside G(M2). Six months of treatment induced a generalized decrease in all the investigated ganglioside fractions. The diazepam-induced ganglioside reduction found after five months of treatment was also present after a one-month recovery period. The only fraction, which recovered and reached its control value, was monosialoganglioside G(M3). PMID- 10534017 TI - Hypericin-induced phototoxicity of human leukemic cell line HL-60 is potentiated by omeprazole, an inhibitor of H+K+-ATPase and 5'-(N,N-dimethyl)-amiloride, an inhibitor of Na+/H+ exchanger. AB - Hypericin, an antiretroviral and antineoplastic agent, seems to have multiple modes of light-induced biological activity connected with the production of single oxygen and/or excited-state proton transfer and a consequent pH drop of pH formation in the hypericin environment. In the present study omeprazole, an inhibitor of H+K+-ATPase, and amiloride, an inhibitor of the Na+/H+ exchanger, have been used for testing the hypothetical pH decreasing effect of hypericin in its antineoplastic action. The results of our experiments have shown that in the HL-60 cell line the effect of hypericin (10(-6) mol.l(-1)) was significantly potentiated by omeprazole and 5'-(N,N-dimethyl)-amiloride. The effect of omeprazole seemed to be less specific than that of 5'-(N,N-dimethyl)-amiloride. Our results support the hypothesis that the excited-state proton transfer and the consequent acidification of the hypericin environment could play a role in the biological activity of hypericin. Moreover, both omeprazole and 5'-(N,N-dimethyl) amiloride are effective potentiating agents of hypericin cytotoxic effect in the HL60 cell line. PMID- 10534019 TI - Inhibition of metaphit-induced audiogenic seizures by APV in rats. AB - The influence of APV ((+/-)-2-amino-5-phosphonovaleric acid) on EEG activity and behavior was studied on a model of epilepsy induced by intraperitoneal administration of metaphit (1-(1-(3-isothiocyanatophenyl)-cyclohexyl) piperidine). Male Wistar rats received an injection of metaphit (10 mg/kg) and were subjected to intense audio stimulation (100+/-3 dB, 60 s) at hourly intervals during the experiment. The seizures were classified according to a four point scale ranging from 0 (no seizure) to 3 (tonic convulsions). In our report we studied the time course which revealed the maximum incidence and severity of seizures 7-12 h after the injection (10 out of 12 rats, with severity of 2.25+/ 0.32). APV (0.05, 0.1, 0.2 and 0.3 micromol) was injected intracerebroventricularly at the time of fully developed convulsions. APV inhibited seizures in a dose-dependent manner. The minimum dose, which completely blocked seizures in all animals, was 0.3 micromol, while ED50 were 0.11, 0.10 and 0.07 micromol against running, clonus and tonus, respectively. In contrast to behavioral inhibition of convulsions, metaphit-provoked epileptiform activity was not abolished by APV, and represented a prerequisite for the reappearance of behavioral seizures. It is suggested that APV is rather an anticonvulsant than an antiepileptic agent in this model of epilepsy. PMID- 10534020 TI - Assessment of EEG frequency dynamics using complex demodulation. AB - The complex demodulation (CD) approach was applied to human EEG recorded during a cognitive task performance, including voluntary goal-directed movements. The standard CD algorithm was extended by a simple procedure using frequency histograms and power spectra to select the characteristic frequencies of EEG segments around the task performance. In the majority of records, amplitude modulation was found, which decreased or disappeared in the period prior to and at the very beginning of the task performance. It was found that the decrease of modulation in fast beta and gamma components begins approximately one second before that of the alpha components. Frequency modulation appeared in some records at the end of the task in beta and gamma components. The results showed that a cognitive task performance is accompanied by non-linear processes in the frequency components of EEG. These dynamic changes could extend the findings of event-related desynchronization obtained by linear methods. PMID- 10534021 TI - The effect of triiodothyronine on changes of membrane fluidity in regenerating rat liver. AB - The increase of the membrane fluidity during the early phase of liver regeneration after partial hepatectomy was described in literature in plasma membrane and in microsomes. We found similar changes also in isolated mitochondria and in crude total membrane fraction of the liver homogenate. The administration of triiodothyronine to rats before partial hepatectomy diminished the increase of the membrane fluidity in the regenerating liver by 50%. Triiodothyronine effect is explained by hormonal modification of lipid metabolism in the regenerating liver. PMID- 10534022 TI - Reciprocal adaptive response of human peripheral lymphocytes induced by bleomycine or gamma rays. AB - The adaptive response and reciprocal adaptive response induced in vitro by exposure to low doses of gamma rays (0.05 Gy) or bleomycin (0.05 microg/ml) in human peripheral blood lymphocytes were assessed by the frequency of chromosome aberrations. Gamma rays (1.5 Gy) or bleomycin (1.5 microg/ml) were used as the challenge doses. In the experiments, blood samples from 5 healthy donors were investigated. It has been found that low doses of bleomycin and gamma rays induced a reciprocal adaptive response to high doses of gamma rays or bleomycin. Moreover, the results confirmed that the adaptive response did not correlate with the radiosensitivity of the peripheral blood lymphocytes. PMID- 10534023 TI - Introduction to the proceedings of the Turtle Creek Consensus Conference on Prehospital Care. PMID- 10534024 TI - Confirmation of airway placement. AB - Proper airway management in the prehospital setting is essential. Recent data from Orange County, Florida, suggest that the problem of misplaced endotracheal tubes may be greater than previous studies have indicated. Strong medical direction, strict protocols, and active continuous quality improvement programs are needed to ensure that paramedics learn the correct techniques of endotracheal intubation, and that they verify tube placement with an end-tidal carbon dioxide monitor, and maintain ongoing monitoring of tube placement during transport. PMID- 10534025 TI - Pharmacologic treatment of cardiac arrest. AB - Antiarrhythmic drugs currently recommended in the American Heart Association's Advanced Cardiac Life Support (ACLS) guidelines for the treatment of cardiac arrest have not been proved in controlled clinical trials to improve survival in patients with ventricular fibrillation (VF) or pulseless ventricular tachycardia (VT). Intravenous amiodarone is a promising agent for the treatment of VF and VT. Based on available evidence, amiodarone should be considered for use in patients with shock-refractory ventricular arrhythmias. PMID- 10534026 TI - Biphasic truncated exponential waveform defibrillation. AB - This paper presents data from studies that have compared the efficacies of biphasic truncated exponential (BTE) and monophasic damped sine (MDS) waveform defibrillation in patients with out-of-hospital cardiac arrest and in in-hospital defibrillation. When a shock is delivered, rhythms evolve rapidly in a variety of directions and take different courses, even over a short time. When defibrillation is defined as termination of ventricular fibrillation at 5 seconds postshock, whether to an organized rhythm or asystole, low-energy BTE shocks appear to be more effective than high-energy MDS shocks in out-of-hospital arrest. For future research, the terms associated with defibrillation should be standardized and used uniformly by all investi-gators. In particular, there should be an agreed-upon definition of shock efficacy. PMID- 10534027 TI - Management of difficult airways in the field. AB - Establishing an airway is a critical first step in emergency management of comatose patients and those who have suffered head trauma, cardiac arrest, or respiratory failure. The use of succinylcholine, a paralytic, to assist with intubation is a safe and effective way to help establish an airway under difficult circumstances, in the prehospital setting. It requires excellent intubation skills, a thorough knowledge of the indications and contraindications of its use, and similar knowledge of any other medications employed. Succinylcholine-assisted intubation should never be implemented without close physician monitoring. Therefore, under the auspices of strong medical control, it is an effective way to establish adequate oxygenation and to control ventilation in some of the most critical patients encountered in the field. Additionally, because physical examination alone is not dependable for ensuring proper endotracheal tube placement, an objective confirmatory device such as an end tidal carbon dioxide detector should be used. PMID- 10534028 TI - Public-access defibrillation. AB - This paper provides a general overview of the current status of public-access defibrillation (PAD). The objectives are to describe the rationale for PAD, to define different levels of PAD as enumerated by the American Heart Association AED Task Force, to review the development of PAD over the past decade, and to discuss the cost-benefit of PAD, mainly from models and a few small implementations throughout the country. A prospective, randomized, controlled clinical trial (the Public Access Defibrillation Phase I Trial) has been designed to compare nontraditional targeted bystander CPR using AEDs with that not using AEDs. It is expected that the trial will resolve many of the uncertainties about the benefits of PAD. PMID- 10534029 TI - Public-access defibrillation: where do we place the AEDs? AB - BACKGROUND: Many prehospital cardiac arrests occur in public places. Even the best EMS systems have a finite response time. Therefore, it has been recommended that automated external defibrillators (AEDs) be placed in public areas for immediate access by trained members of the general public. OBJECTIVE: To determine the locations of multiple cardiac arrests in order to plan for placement of public-access AEDs. METHODS: Retrospective review of all primary cardiac arrests in calendar year 1997. Cardiac arrests in which resuscitation was not attempted (DOA), traumatic cases, pediatric cases, and those due to "other" causes were excluded. Location of the cardiac arrest was obtained from the ambulance run ticket. The EMS system is an urban, Midwestern, all-ALS, public utility model system with fire department first responders that transports approximately 58,000 patients annually. RESULTS: There was scene response to 922 cardiac arrests. 377 DOAs and 219 nonprimary cardiac arrests were excluded. There were 326 primary cardiac arrests. Sixteen locations had more than one cardiac arrest: 11 locations had two cardiac arrests, four locations had three cardiac arrests, and one location had four cardiac arrests. The airport, an airline overhaul facility, a casino, and two hotels each had two cardiac arrests; the other locations of multiple cardiac arrests were in nursing homes. The professional sports stadiums had no cardiac arrests. CONCLUSIONS: Since very few locations had more than one cardiac arrest, it may be difficult to identify high yield public places in which to place an AED. Nursing homes may want to consider AED availability. PMID- 10534030 TI - Prehospital consideration of sildenafil-nitrate interactions. AB - OBJECTIVE: To determine whether paramedics and on-line physicians screen patients for use of sildenafil citrate (Viagra) prior to prehospital administration of nitrates. METHODS: A prospective, observational study was performed over a one month period in three EMS systems. Consecutive radio communications between on line physicians and paramedics concerning male patients with cardiac complaints were monitored. Investigators observed the frequency with which on-line physicians screened for sildenafil use prior to ordering nitrates. After observation of the radio communications was completed, a written survey was distributed to all paramedics in the three EMS systems. RESULTS: Seventy-six physician-paramedic interactions were monitored. Nitrates were ordered by on-line physicians in 56 cases. No paramedic reported sildenafil use/nonuse, and no on line physician inquired about the patient's potential use of the drug. Only half of the surveyed paramedics reported that they routinely screen for sildenafil use, and approximately a fourth reported that its use would not alter their management of chest pain patients. CONCLUSION: In this study, on-line physicians in three EMS settings did not screen for sildenafil use prior to ordering nitrates. While some paramedics do screen for sildenafil use, practice patterns among paramedics in these three systems were inconsistent. PMID- 10534031 TI - Flight crew airway management in four settings: a six-year review. AB - OBJECTIVE: To analyze flight crew airway management in four different settings (in flight, at trauma scenes, in ambulance, and in referring hospitals) and in two different aircraft used by the same helicopter EMS (HEMS) service. The null hypothesis was that there was no association between practice setting, or aircraft, and airway practices or success rate. METHODS: This retrospective study analyzed all patients in whom advanced airway management was attempted by the HEMS service during the study period October 1991 through October 1997. Data used were from flight records of Boston MedFlight Critical Care Transport Service, which uses a nurse/paramedic crew and had a paralytic-assisted intubation protocol in place. Data were analyzed with chi-square and Fisher's exact testing, risk ratio analysis, and logistic regression. RESULTS: Advanced airway management was attempted in 722 patients, with an airway placed in 705 (97.8%). Intubation success was unrelated to site of airway management (p = 0.14), but patients were more likely to have intubation attempted prior to flight (as opposed to in flight) if the crew were in the AS365N2 Dauphin as compared with the BK-117 (p<0.0001). In addition, patients were 0.77 times as likely (95% confidence interval, 0.68-0.88) to receive paralytic-facilitated intubation if airway management occurred in the hospital setting as compared with other sites. CONCLUSIONS: While HEMS crew airway management success rates are equally high in all practice settings, airway management decision making and practice appear to be significantly influenced by practice setting and aircraft type. These data support contentions that nonphysician HEMS crews can effectively manage airways in a variety of circumstances. PMID- 10534032 TI - Characteristics of state legislation governing medical care at mass gatherings. AB - Organized mass gathering medical care (MGMC) has existed in the United States for 30 years, but there is little evidence to support any standard of care or uniformity in its delivery. OBJECTIVE: To determine whether MGMC regulations exist within state EMS legislation and to describe the characteristics of any such regulations. METHODS: The authors conducted a cross-sectional survey of U.S. state EMS directors in fall 1998 to determine the prevalence of formal legislation governing MGMC. The lead author received copies of legislation from every state EMS office that indicated such legislation existed. RESULTS: Responses were obtained from all 50 state EMS offices and that of the District of Columbia (n = 51). Only six (12%) states provide regulatory guidance for MGMC. These regulations reside within departments of health in all six states and within the divisions of EMS in three of these six. Only one state requires physician oversight of a medical action plan and minimum staffing by EMS personnel, respectively. No state addresses early defibrillation capability or EMS scope of practice. There is no agreement on the definition of either a mass gathering or minimum resource deployment. Public health and hygiene practices at mass gatherings also lack uniformity. CONCLUSION: Few states regulate MGMC. Existing regulations are poorly developed and lack both standardized terminology and content. PMID- 10534033 TI - Managed care enrollee utilization of 911 medical services. AB - OBJECTIVE: To determine the mechanism by which managed care organization (MCO) enrollees enter the emergency medical services (EMS) system. METHODS: All enrollees belonging to the region's largest MCO and transported to emergency departments by a paramedic-level municipal EMS system were identified from billing records. Dispatch logs were examined to determine the time and origin of the call to the 911 communication center. Patient care records were used to obtain age, the level of care delivered (advanced or basic life support), and whether the patient received any medications while out of hospital. Hospital admission was also determined. RESULTS: Over a six-month period, 195 enrollees were transported. Three modes of 911 EMS system entry were identified: group I enrollees who called 911 directly; group II-enrollees who called the MCO triage center, who then called 911 on behalf of the patient; and group III--enrollees who were sent to the MCO health center for evaluation, and subsequently the MCO called 911 to transfer the patient to the hospital. Of the 195 patients transported to the emergency department, the dispositions of 108 (55%) patients were obtained. Group I (n = 109) patients were more likely to be transported in the evening (3 PM to 11 PM), less likely to require advanced life support therapies, and less likely to be admitted to the hospital when compared with groups II (n = 32) and III (n = 54) patients. Group III patients were the most likely to receive advanced life support care and require admission to the hospital. CONCLUSION: The majority of MCO enrollees called 911 directly, and were most likely to do so during evening hours. Enrollees who called 911 directly (group I) had a trend toward lower acuity, based on the lowest ALS utilization of any group. Those enrollees who most frequently required advanced life support were those who received initial treatment at the MCO center prior to EMS transport. Though EMS system-specific, this type of descriptive analysis is helpful in assisting both EMS systems and MCOs to better assess utilization of 911 EMS resources by MCO enrollees. This study also challenges the prudent layperson paradigm. PMID- 10534034 TI - Measuring quality and effectiveness of prehospital EMS. AB - INTRODUCTION: In today's health care environment, the demand for objective comparative information about the performance of health care organizations and providers has created a need for data-driven evaluation processes. In response, national organizations and federal agencies have established quality indicators, created tools to measure performance according to those indicators, and issued report cards for individual providers, as well as health care organizations. PURPOSE: Emergency medical services (EMS) systems are no different from other health care systems in the need for objective comparative system information to assist government officials at all levels in establishing relevant policy, selecting appropriate system design, and monitoring system quality and effectiveness. Governmental decision makers, payers, and consumers are demanding objective evidence that they are receiving value and quality for the cost of EMS. EMS systems administrators also require objective feedback about performance that can be used internally to support improvement efforts and externally to demonstrate accountability to the public and other stakeholders. To date, there are few validated indicators of effectiveness and quality in EMS systems. Moreover, most potential indicators have not been studied for use in systemwide evaluation. As a result, there are no universally accepted methods of measurement. The following paper examines traditional efforts to assure quality in EMS systems, while assessing the need to go beyond the traditional to establish measurable indicators of system quality. Valid and measurable indicators will provide a basis for establishing benchmarks of performance. In the future, these benchmarks will facilitate comparisons of a system with itself, as well as with other systems. PMID- 10534035 TI - The reliability of prehospital clinical evaluation for potential spinal injury is not affected by the mechanism of injury. AB - INTRODUCTION: Traditional EMS teaching identifies mechanism of injury as an important predictor of spinal injury. Clinical criteria to select patients for immobilization are being studied in Michigan and have been implemented in Maine. Maine requires automatic immobilization of patients with "a positive mechanism" clearly capable of producing spinal injury. OBJECTIVE: To determine whether mechanism of injury affects the ability of clinical criteria to identify patients with spinal injury. METHODS: In this multicenter prospective cohort study, EMS personnel completed a check-off data sheet for prehospital spine-immobilized patients. Data included mechanism of injury and yes/no determinations of the clinical criteria: altered mental status, neurologic deficit, evidence of intoxication, spinal pain or tenderness, and suspected extremity fracture. Hospital outcome data included confirmation of spinal injury and treatment required. Mechanisms of injury were tabulated and rates of spinal injury for each mechanism were calculated. The patients were divided into three different high risk and low-risk groups. RESULTS: Data were collected for 6,500 patients. There were 209 (3.2%) patients with spinal injuries identified. There were 1,058 patients with 100 (9.4%) injuries in the first high-risk mechanism group, and 5,423 patients with 109 (2%) injuries in the first low-risk group. Criteria identified 97 of 100 (97%) injuries in the high-risk group and 102 of 109 (94%) in the low-risk group. Two additional data divisions yielded identical results. CONCLUSION: Mechanism of injury does not affect the ability of clinical criteria to predict spinal injury in this population. PMID- 10534036 TI - Evaluation of a new EMS asthma protocol in New York City: a preliminary report. AB - BACKGROUND: In July 1996, the New York City (NYC) regional EMS implemented a new protocol whereby EMS personnel in the prehospital setting could administer 125 mg of intravenous methyl prednisolone to asthma patients as one of their medical options following telephone consultation with a medical control physician. OBJECTIVE: To determine whether this protocol had any effect on hospital admission rates or the emergency department (ED) length of stay. METHODS: This retrospective chart review focused on the 219 (of 603 total) patients who arrived to the ED by ambulance over a two-year period whose ED diagnosis was asthma. There were 81 patient encounters in year 1, and 138 in year 2. Eleven of the year 2 group received prehospital steroids. The study took place at an urban 911 receiving, Level 2 ED. RESULTS: Of the group who received prehospital steroids, none resulted in hospital admission. Due to the small sample size in the steroid receiving group, the differences in these admission rates are not yet significant. No differences were detected in the ED length of stay between the two patient groups (157 vs. 160 minutes in year 2, p = 0.9). CONCLUSION: The differences in admission rates suggested by this study suggest a simple yet potentially powerful tool for improving patient outcome in the treatment of asthma. PMID- 10534037 TI - A rural emergency medical technician with selected advanced skills. AB - OBJECTIVE: To educate rural emergency medical technician basics (EMTs) in selected advanced skills, and then evaluate the safety and effectiveness of practice. METHODS: After a minimum 72 hours of training, EMTs employed three skills (Combitube, glucometry, automated external defibrillation) and seven medications (albuterol, nitroglycerin, naloxone, epinephrine, glucagon, activated charcoal, and aspirin). Written patient care records and audiotapes were reviewed. Congruence between prehospital assessment and emergency department (ED) diagnosis was assessed, along with correct use of airway skills (18 of 36 months). The completeness of documentation, appropriateness of treatment, and patient response (by explicit criteria) were determined. Errors and complications were recorded. RESULTS: During three years of the program, 266 patients were treated, primarily for chest pain and respiratory distress. No significant errors or complications occurred. Treatment was judged 94% appropriate, with improvement in 60% of patients. Documentation had major omissions in 3% of cases. Field and ED diagnostic congruence was present in 97/129 (75%) when evaluated during the first 18 months. EMT skill levels were maintained. The mean time to traditional advanced life support (ALS) care was 41 minutes. CONCLUSIONS: Basic-level EMTs in rural areas can be trained in selected advanced skills, and provide ALS-level care quickly and appropriately. Close medical oversight involving review of care and follow-up education is an important part of the program. PMID- 10534038 TI - Respiratory effects of spinal immobilization. AB - OBJECTIVE: To evaluate the effect of whole-body spinal immobilization on respiration. METHODS: This was a randomized, crossover laboratory study with 39 human volunteer subjects (20 males; 19 females) ranging in age from 7 to 85 years. Respiratory function was measured three times: at baseline (seated or lying), immobilized with a Philadelphia collar on a hard wooden backboard, and on a Scandinavian vacuum mattress with a vacuum collar. The comfort levels of each of the two methods were assessed on a forced Likert scale. RESULTS: Both immobilization methods restricted respiration, 15% on the average. The effects were similar under the two immobilization conditions, although the FEV1 was lower on the vacuum mattress. Respiratory restriction was more pronounced at the extremes of age. The vacuum mattress was significantly more comfortable. CONCLUSION: This study confirmed the previously reported respiratory restriction caused by spinal immobilization. Vacuum mattresses are more comfortable than wooden backboards. PMID- 10534039 TI - The impact of television public service announcements on the rate of bystander CPR. AB - OBJECTIVE: To determine whether televised public service announcements (PSAs) demonstrating the fundamentals of CPR were effective in increasing the rate of layperson bystander-initiated CPR. METHODS: Two 30-second PSAs were shown 597 times from September 8, 1996, through April 12, 1997. In each, CPR was given to one member of an older couple by the other in the home. The authors measured rates of bystander CPR in communities that were exposed to the PSA and in communities that were not exposed in two time periods, a before-airing period, January 1, 1993, through September 7, 1996, and a during-airing period, September 8, 1996, through April 12, 1997. A case was defined as a patient with a nontraumatic cardiac arrest that occurred before arrival of EMS personnel, and for whom CPR was initiated by EMS personnel or lay bystanders. RESULTS: There were 1,786 cardiac arrests in the "before" period and 289 in the "during" period. The rate of bystander CPR increased from 43% to 55% (p<0.05) in the intervention community and remained the same in the comparison community (33%). CONCLUSION: Airing of the PSA was accompanied by an increase in the rate of bystander CPR, though the increase may be attributable to a secular trend. PMID- 10534040 TI - Gamma hydroxybutyrate (GHB): report of a mass intoxication and review of the literature. PMID- 10534041 TI - Inadvertent endotracheal needle migration during resuscitation. PMID- 10534042 TI - Initial survival following massive crush injury, leg avulsion, and hemicorporectomy. PMID- 10534043 TI - Finding some good in a tragedy. PMID- 10534045 TI - Organ procurement and EMS: expanding the scope of your practice and your life. PMID- 10534044 TI - The National Organ and Tissue Donation Initiative: working together to save lives. PMID- 10534046 TI - Organ procurement background and misconceptions. PMID- 10534047 TI - Paramedic use of succinylcholine. PMID- 10534048 TI - Exact rebinning methods for three-dimensional PET. AB - The high computational cost of data processing in volume PET imaging is still hindering the routine application of this successful technique, especially in the case of dynamic studies. This paper describes two new algorithms based on an exact rebinning equation, which can be applied to accelerate the processing of three-dimensional (3-D) PET data. The first algorithm, FOREPROJ, is a fast forward projection algorithm that allows calculation of the 3-D attenuation correction factors (ACF's) directly from a two-dimensional (2-D) transmission scan, without first reconstructing the attenuation map and then performing a 3-D forward projection. The use of FOREPROJ speeds up the estimation of the 3-D ACF's by more than a factor five. The second algorithm, FOREX, is a rebinning algorithm that is also more than five times faster, compared to the standard reprojection algorithm (3DRP) and does not suffer from the image distortions generated by the even faster approximate Fourier rebinning (FORE) method at large axial apertures. However, FOREX is probably not required by most existing scanners, as the axial apertures are not large enough to show improvements over FORE with clinical data. Both algorithms have been implemented and applied to data simulated for a scanner with a large axial aperture (30 degrees), and also to data acquired with the ECAT HR and the ECAT HR+ scanners. Results demonstrate the excellent accuracy achieved by these algorithms and the important speedup when the sinogram sizes are powers of two. PMID- 10534049 TI - Penalized-likelihood estimators and noise analysis for randoms-precorrected PET transmission scans. AB - This paper analyzes and compares image reconstruction methods based on practical approximations to the exact log likelihood of randoms-precorrected positron emission tomography (PET) measurements. The methods apply to both emission and transmission tomography, however, in this paper we focus on transmission tomography. The results of experimental PET transmission scans and variance approximations demonstrate that the shifted Poisson (SP) method avoids the systematic bias of the conventional data-weighted least squares (WLS) method and leads to significantly lower variance than conventional statistical methods based on the log likelihood of the ordinary Poisson (OP) model. We develop covariance approximations to analyze the propagation of noise from attenuation maps into emission images via the attenuation correction factors (ACF's). Empirical pixel and region variances from real transmission data agree closely with the analytical predictions. Both the approximations and the empirical results show that the performance differences between the OP model and SP model are even larger, when considering noise propagation from the transmission images into the final emission images, than the differences in the attenuation maps themselves. PMID- 10534050 TI - Multiobjective genetic optimization of diagnostic classifiers with implications for generating receiver operating characteristic curves. AB - It is well understood that binary classifiers have two implicit objective functions (sensitivity and specificity) describing their performance. Traditional methods of classifier training attempt to combine these two objective functions (or two analogous class performance measures) into one so that conventional scalar optimization techniques can be utilized. This involves incorporating a priori information into the aggregation method so that the resulting performance of the classifier is satisfactory for the task at hand. We have investigated the use of a niched Pareto multiobjective genetic algorithm (GA) for classifier optimization. With niched Pareto GA's, an objective vector is optimized instead of a scalar function, eliminating the need to aggregate classification objective functions. The niched Pareto GA returns a set of optimal solutions that are equivalent in the absence of any information regarding the preferences of the objectives. The a priori knowledge that was used for aggregating the objective functions in conventional classifier training can instead be applied post optimization to select from one of the series of solutions returned from the multiobjective genetic optimization. We have applied this technique to train a linear classifier and an artificial neural network (ANN), using simulated datasets. The performances of the solutions returned from the multiobjective genetic optimization represent a series of optimal (sensitivity, specificity) pairs, which can be thought of as operating points on a receiver operating characteristic (ROC) curve. All possible ROC curves for a given dataset and classifier are less than or equal to the ROC curve generated by the niched Pareto genetic optimization. PMID- 10534051 TI - Geometrically correct 3-D reconstruction of intravascular ultrasound images by fusion with biplane angiography--methods and validation. AB - In the rapidly evolving field of intravascular ultrasound (IVUS), the assessment of vessel morphology still lacks a geometrically correct three-dimensional (3-D) reconstruction. The IVUS frames are usually stacked up to form a straight vessel, neglecting curvature and the axial twisting of the catheter during the pullback. Our method combines the information about vessel cross-sections obtained from IVUS with the information about the vessel geometry derived from biplane angiography. First, the catheter path is reconstructed from its biplane projections, resulting in a spatial model. The locations of the IVUS frames are determined and their orientations relative to each other are calculated using a discrete approximation of the Frenet-Serret formulas known from differential geometry. The absolute orientation of the frame set is established, utilizing the imaging catheter itself as an artificial landmark. The IVUS images are segmented, using our previously developed algorithm. The fusion approach has been extensively validated in computer simulations, phantoms, and cadaveric pig hearts. PMID- 10534053 TI - Nonrigid registration using free-form deformations: application to breast MR images. AB - In this paper we present a new approach for the nonrigid registration of contrast enhanced breast MRI. A hierarchical transformation model of the motion of the breast has been developed. The global motion of the breast is modeled by an affine transformation while the local breast motion is described by a free-form deformation (FFD) based on B-splines. Normalized mutual information is used as a voxel-based similarity measure which is insensitive to intensity changes as a result of the contrast enhancement. Registration is achieved by minimizing a cost function, which represents a combination of the cost associated with the smoothness of the transformation and the cost associated with the image similarity. The algorithm has been applied to the fully automated registration of three-dimensional (3-D) breast MRI in volunteers and patients. In particular, we have compared the results of the proposed nonrigid registration algorithm to those obtained using rigid and affine registration techniques. The results clearly indicate that the nonrigid registration algorithm is much better able to recover the motion and deformation of the breast than rigid or affine registration algorithms. PMID- 10534052 TI - On the Laplace-Beltrami operator and brain surface flattening. AB - In this paper, using certain conformal mappings from uniformization theory, we give an explicit method for flattening the brain surface in a way which preserves angles. From a triangulated surface representation of the cortex, we indicate how the procedure may be implemented using finite elements. Further, we show how the geometry of the brain surface may be studied using this approach. PMID- 10534054 TI - A projector/backprojector with slice-to-slice blurring for efficient three dimensional scatter modeling. AB - Scatter correction is an important factor in single photon emission computed tomography (SPECT). Many scatter correction techniques, such as multiple-window subtraction and intrinsic modeling with iterative algorithms, have been under study for many years. Previously, we developed an efficient slice-to-slice blurring technique to model attenuation and system geometric response in a projector/backprojector pair, which was used in an ML-EM algorithm to reconstruct SPECT data. This paper proposes a projector/backprojector that models the three dimensional (3-D) first-order scatter in SPECT, also using an efficient slice-to slice blurring technique. The scatter response is estimated from a known nonuniform attenuation distribution map. It is assumed that the probability of detection of a first-order scattered photon from a photon that is emitted in a given source voxel and scattered in a given scatter voxel is proportional to the attenuation coefficient value at that voxel. Monte Carlo simulations of point sources and an MCAT torso phantom were used to verify the accuracy of the proposed projector/backprojector model. An experimental Jaszczak torso/cardiac phantom SPECT study was also performed. For a 64 x 64 x 64 image volume, it took 8.7 s to perform each iteration per slice on a Sun ULTRA Enterprise 3000 (167 MHz, 1 Gbyte RAM) computer, when modeling 3-D scatter, attenuation, and system geometric response functions. The main advantage of the proposed method is its easy implementation and the possibility of performing reconstruction in clinically acceptable time. PMID- 10534055 TI - Acoustic noise analysis in echo planar imaging: multicenter trial and comparison with other pulse sequences. AB - The purpose of this study was to evaluate acoustic noise in echo planar imaging (EPI) at various magnetic resonance imaging (MRI) centers and to compare EPI acoustic noise with that in other fast pulse sequences. We measured A-weighted root-mean-square sound pressure levels and peak impulse sound pressure levels for EPI, under the same conditions, in eleven clinical super-conducting MRI systems. We also compared sound pressure levels for the EPI and six different pulse sequences and analyzed the acoustic noise spectra. Sound pressure levels during the use of the EPI differed greatly among institutions. Moreover, sound pressure levels of the EPI were not significantly different from those of other fast pulse sequences and were within permissible noise exposure levels. In comparison to other fast sequences, the EPI had significantly greater acoustic noise in the high-octave band frequency. PMID- 10534056 TI - Gene therapy. AB - For the past two decades, concerted efforts have been made to treat human disease by replacing nonfunctioning genes in cells or by correcting mutations that produce disease. Successful application of these methods could lead to effective therapies for a variety of genetic and acquired diseases, many of which are not treatable today. Genes can be transferred into cells using either viral vectors, which have evolved the ability to bring their DNA into the host cell, or nonviral vectors, which, until recently, have been much less efficient in their ability to introduce exogenous DNA into cells. Whereas cell transduction is possible in vivo, it is much simpler to accomplish in vitro; for this reason the hematopoietic stem cell has been a favorite target in attempts to implement gene therapy. Stem cells can be partially purified from the blood or marrow, transduced with the gene of interest, and reintroduced into the body. Any genetic disease that responds to transplantation would be expected also to respond to gene therapy. The progress made over the past two decades has resulted in better vectors and ingenious new techniques for correcting mutations in endogenous genes. Although effective gene therapy in humans has continued to be an elusive goal, recent advances lead us to hope that this goal will be realized within the next few years. PMID- 10534057 TI - Dose and timing of interleukin (IL)-12 and timing and type of total-body irradiation: effects on graft-vs.-host disease inhibition and toxicity of exogenous IL-12 in murine bone marrow transplant recipients. AB - Paradoxically, a single injection of recombinant murine interleukin (IL)-12 on the day of bone marrow transplantation (BMT) inhibits graft-vs.-host disease (GVHD) while preserving graft-vs.-leukemia (GVL) effects in lethally irradiated mice receiving fully MHC-mismatched bone marrow and spleen cells. These protective effects are mediated by interferon (IFN)-gamma, whose early secretion is induced by IL-12 treatment. We investigated the relationship of IL-12 dose and timing of administration, as well as timing and type of total-body irradiation (TBI), with the ability of IL-12 to inhibit GVHD or mediate toxicity. The results show that a relatively low dose of IL-12 (as little as 50 U in a single injection) can mediate significant GVHD protection. The timing of IL-12 administration, however, is a critical factor. IL-12 administered 1 hour before BMT was most protective, but protection was still observed when it was administered 1-12 hours after BMT. Delaying IL-12 administration to 36 hours post BMT completely obviated its protective effect. Administration of a second IL-12 injection 6 days after BMT negated the protective effect of an initial injection at the time of BMT. While IL-12 protection was evident when TBI was administered by 137Cs-irradiator in one or two fractions on day -1 or day 0, the use of an X irradiator to deliver TBI on day -1 was associated with marked IL-12 toxicity. Whereas the protective effect of IL-12 against GVHD depended on donor-derived IFN gamma, toxicity depended on the ability of host cells to produce IFN-gamma. Careful studies are warranted to test the effects of IL-12 in the context of BMT with various conditioning regimens in large animal preclinical models before this novel approach to GVHD protection can be applied clinically. PMID- 10534058 TI - Cyclosporine, methotrexate, and prednisone compared with cyclosporine and prednisone for prevention of acute graft-vs.-host disease: effect on chronic graft-vs.-host disease and long-term survival. AB - Graft-vs.-host disease (GVHD) is a major predictor of outcome following allogeneic bone marrow transplantation (BMT). For patients alive at day 100 after BMT, the presence or absence of chronic GVHD is one of the most important determinants of survival and quality of life. We wished to determine the effects on chronic GVHD of two regimens used for the prophylaxis of acute GVHD: cyclosporine, methotrexate, and prednisone (CSA/MTX/PSE) and cyclosporine and prednisone (CSA/PSE). One hundred forty-nine evaluable patients were entered into the acute GVHD study. As of 31 March 1997, 63 months after the last patient underwent BMT, the median survival time was 4.5 years (range 0.09-9.9). The incidence of chronic GVHD was independent of the prophylactic regimen (55 vs. 54%), and extensive chronic GVHD occurred in 25 and 24% of patients receiving CSA/MTX/PSE and CSA/PSE, respectively. Of note, the median Karnofsky performance status of both groups was 100% (range 70-100%), reflecting the low incidence of extensive chronic GVHD. Survival rates free of chronic GVHD were 52 vs. 42% (p = 0.29) for patients receiving CSA/MTX/PSE vs. CSA/PSE. The incidence of relapse was also similar in both groups of patients. These data suggest that the combinations of CSA/MTX/PSE and CSA/PSE result in comparable chronic GVHD-free survival without an increase in leukemic relapse. PMID- 10534059 TI - High-dose chemo/radiotherapy and autologous bone marrow or stem cell transplantation for poor-risk advanced-stage Hodgkin's disease during first partial or complete remission. AB - Complete remission rates of 70-90% can be achieved following combination chemotherapy for patients with advanced-stage Hodgkin's disease (HD). Patients who present with unfavorable poor prognostic factors, however, have a 5-year disease-free survival of only 40-50%. In an attempt to improve the prognosis of 20 patients with poor-risk advanced-stage HD, we evaluated the role of early high dose therapy (HDT) and autologous bone marrow/stem cell transplantation (ASCT) during the first complete or partial remission (CR/PR). Patients were eligible for ASCT if they either achieved a PR (defined as > 50% regression) (six patients), or achieved a CR (14 patients) but had presented with three or more of the following unfavorable features: stage IV disease with bone marrow involvement or > or = 2 extranodal sites of involvement; bulky mass > 10 cm or bulky mediastinal mass > 1/3 of mediastina/thoracic ratio; B symptoms; and elevated serum lactate dehydrogenase (LDH) level. The study included 11 men (55%) and 9 women (45%). The median age was 37 years (range 20-57). Seventeen patients (85%) had stage IV disease; 14 (70%) had B symptoms; 13 (65%) had bulky mass > 10 cm; 14 (70%) had > or = 2 extra nodal sites involvement; and eight patients (40%) had elevated LDH levels. All patients were treated with standard four or 7-8 drug combination chemotherapy regimens until they achieved maximal response prior to ASCT with a median of six cycles (range 4-11). Six patients also received involved field radiotherapy to residual bulky mass > 5 cm or bony lesions before ASCT. The median time from diagnosis to ASCT was 8.6 months (range 5.5-18.9). Preparative regimens consisted of fractionated total body irradiation (FTBI) 1200 cGy in combination with etoposide 60 mg/kg and cyclophosphamide 100 mg/kg in all patients except one who had borderline pulmonary function and received lomustine 15 mg/kg instead of FTBI. All patients engrafted and there was no transplant related mortality. One patient developed congestive cardiomyopathy at 4 years post-ASCT. All patients remain alive and in remission at a median follow-up of 42.8 months (range, 13.2-149.2). These preliminary results suggest that HDT and ASCT can be performed safely during first CR/PR in selected patients with advanced-stage HD who have an unfavorable prognosis. Further randomized studies comparing HDT and ASCT during first CR with conventional chemotherapy and ASCT at relapse in poor-risk advanced-stage HD should be conducted. The prognostic factors and risk groups described recently by an international prognostic study can be used to identify high-risk patients who may be candidates for more intensive therapy. PMID- 10534060 TI - Impact of admission body weight and chemotherapy dose adjustment on the outcome of autologous bone marrow transplantation. AB - We performed a retrospective analysis of 473 consecutive adult patients undergoing autologous bone marrow transplantation for hematologic malignancies between 1988 and 1995. The analysis examined whether significant deviation from ideal body mass index is associated with a decrease in event-free survival (EFS), an increase in nonrelapse mortality (NRM) including late toxicities and second malignancies, or relapse. Chemotherapy dosing in underweight and overweight patients is administered based on the relationship of admission body weight (ABW) to ideal body weight (IBW). Doses were adjusted for obesity; however, the adjustment did not obviate increased risk for NRM. Patients were categorized into five groups according to the relationship of ABW to age-adjusted body mass index (aBMI) as a percent of actual BMI, as follows: group I, 70-79%; group II, 80-99%; group III, 100-119%; group IV, 120-139%; and group V, 140-199% aBMI. When body weight was expressed as percent BMI adjusted for age, there was a significantly increased risk for NRM in groups I and IV (p = 0.03 and 0.02, respectively). A trend toward greater NRM in group V (p = 0.10) was also noted. Multivariate analysis confirmed that the risk of NRM for extremely underweight and overweight patients is almost three times that of patients close to ideal body weight. Age adjusted BMI was an independent predictive factor for NRM but not associated with increased relapse. We determined that dose adjustment could be safely used without significant increase of relapse. In patients with significant deviation of BMI from aBMI, dose adjustment and possible weight normalization should be considered. PMID- 10534061 TI - Veno-occlusive disease of the liver after busulfan, melphalan, and thiotepa conditioning therapy: incidence, risk factors, and outcome. AB - The purpose of this study was to determine the incidence of veno-occlusive disease (VOD) after a high-dose regimen of busulfan, melphalan, and thiotepa and the risk factors for a more severe outcome. We followed 253 consecutive patients with malignant disorders who received autologous transplants after stem cell harvest followed by 12 mg/kg busulfan, 100 mg/m2 melphalan, and 500 mg/m2 thiotepa. Diagnosis of VOD was based on weight gain, hepatomegaly, and jaundice. Risk factors for moderate or severe VOD were identified using logistic regression models. VOD occurred in 70 of 253 patients (28%), of whom 31 (12%) had moderate and 11 (4%) severe VOD. The median day of onset of hyperbilirubinemia was day 9, significantly later than the onset of jaundice after our cyclophosphamide-based regimens (p < 0.001). Resolution of weight gain and jaundice, followed by their reappearance several weeks later, occurred in 23 of 70 patients with VOD and was an adverse prognostic sign. Risk factors for moderate or severe VOD were a diagnosis of lymphoma or myeloma (odds ratio [OR] 2.65 compared with breast cancer), tumor involvement in the liver (OR 3.95), fever in the month before transplant (OR 3.32), and prior radiation therapy (OR 2.70). We conclude that VOD after busulfan, melphalan, and thiotepa was less frequent and less severe and developed later than VOD after our historical cyclophosphamide-based regimens. Significant risk factors included a diagnosis other than breast cancer, hepatic metastases, persistent fever, and prior radiation therapy. This study suggests that alkylating agents of comparable overall toxicity differ in their liver toxicity. PMID- 10534063 TI - Thiotepa-associated cardiomyopathy during blood or marrow transplantation: association with the female sex and cardiac risk factors. AB - Thiotepa (TT) has not been reported to cause cardiomyopathy, whereas cyclophosphamide (Cy)-related cardiomyopathy is well characterized. To search for cases of acute onset cardiomyopathy associated with TT, we retrospectively reviewed 171 patients who received TT-containing conditioning regimens for blood or marrow transplantation (BMT). Nine of 171 patients (5.3%) developed clinical congestive heart failure in the post-BMT period. The median time to onset of heart failure was 15 days after BMT (range 5-30). The median pre-BMT left ventricular ejection fraction (LVEF) was 50% (range 42-65%) as determined by two dimensional echocardiogram, or gated blood pool scan. At the time of cardiomyopathy onset, LVEF was 30%. Six patients died of causes unrelated to heart failure. All affected patients who developed congestive heart failure following administration of TT had some evidence of cardiac dysfunction prior to transplantation. Significant risk factors for the development of cardiomyopathy included low pre-BMT-LVEF and female sex--particularly in females receiving allogeneic transplantation. The incidence of congestive heart failure with TT containing regimens was similar to the incidence using other regimens with and without Cy. The mean time to clinical evidence of TT-associated cardiomyopathy was longer than the mean time reported with Cy. We recommend caution in using high-dose TT-containing regimens for patients with histories of cardiac dysfunction. PMID- 10534062 TI - Myeloablation by intravenous busulfan and hematopoietic reconstitution with autologous marrow in a canine model. AB - We have previously described pharmacokinetic studies with a dimethylsulfoxide based intravenous busulfan preparation in a canine model and in preliminary clinical trials. Using the same intravenous busulfan preparation, we carried out a dose escalation study to determine a marrow-ablative dose and to test the ability of autologous marrow to reconstitute hematopoiesis in dogs so treated. Busulfan was given intravenously at doses of 3.75 to 40 mg/kg. Marrow ablation was achieved at 20 mg/kg given either as a single dose or in four daily increments of 5 mg/kg each. There was a relative sparing of lymphocytes. A busulfan dose of 40 mg/kg resulted in severe central nervous system toxicity. Otherwise, nonhematopoietic toxicity was minimal and restricted to mild hepatic abnormalities. Four dogs were given busulfan at 20 mg/kg followed 30 hours later by infusion of autologous marrow, and all showed prompt and complete hematopoietic reconstitution. The area under the curve (AUC) determined by busulfan concentration in plasma over time was dose dependent, ranging from 12 to 100 microg x h/mL for busulfan doses of 3.75-20 mg/kg. There was a suggestion that the plasma half-life increased at the highest busulfan doses used. Intravenous administration of busulfan circumvented differences in bioavailability; nevertheless, considerable variations in the pharmacokinetic parameters were observed between individual animals. Thus, intravenous busulfan can be given safely and is effective in ablating hematopoiesis. However, factors other than absorption influence the AUC, and individualization of dosing may be required even with intravenous administration of the drug. PMID- 10534064 TI - Comparison of the distribution of progenitor cells in G-CSF-mobilized peripheral blood and steady-state bone marrow after counterflow centrifugal elutriation. AB - Blood-derived progenitor cells obtained following mobilization with granulocyte colony-stimulating factor (MoPBSC) are increasingly being used as an alternative to bone marrow (BM) in allogeneic stem cell transplantation. The higher numbers of mature T lymphocytes in MoPBSC grafts may increase the risk of (chronic) graft vs.-host disease. Counterflow centrifugal elutriation (CCE) is an effective method for T-cell depletion of BM grafts. The elutriation characteristics of steady-state BM and MoPBSC were compared using a CCE procedure in which fractions were obtained after small incremental increases in flow rate with constant centrifugal force. Counterflow centrifugal elutriation experiments with MoPBSC from six healthy volunteers showed that 54% of all cells collected were recovered in the < or = 15 mL/minute fractions, whereas experiments with mononuclear BM cells from five healthy volunteers resulted in recovery of 52% of collected cells from the > or = 19 mL/minute fractions. The peak concentrations of CD34+ cells were found in the same fraction (18 mL/minute), but more CD34+ cells from MoPBSC were recovered from the small (< or = 16 mL/minute) fractions (54% for MoPBSC, 26% for BM; p = 0.08). The small CD34+ cells from BM were more frequently lacking CD38 and human leucocyte antigen-DR expression than the small CD34+ cells from MoPBSC. Mature T-cells (CD3+) in BM and MoPBSC samples had similar CCE features, as did early (long-term culture initiating cells, high-proliferative potential colony-forming cells) and more mature (colony-forming units granulocyte/macrophage, BFU-e) hematopoietic progenitor cells. The results of this study suggest that T-cell depletion by CCE of MoPBSC as compared to BM products, may lead to a greater loss of CD34+ cells, but not of immature hematopoietic progenitor cells. PMID- 10534065 TI - Effects of occlusal loading on ankylosis, bone, and cementum formation during bone morphogenetic protein-2-stimulated periodontal regeneration in vivo. AB - BACKGROUND: The purpose of this study was to investigate the influence of occlusal loading on recombinant human bone morphogenetic protein-2 (BMP-2) induced bone and cementum formation in a previously established rat model of periodontal regeneration during the early and late stages of wound healing. METHODS: 64 Wistar rats were divided into 8 groups and had surgically created fenestrated defects on the right side of the mandible involving the removal of bone and exposure of the first and second molar roots. Four groups had their right maxillary molars extracted 2 weeks prior to surgery. Ten microl of 100 ug/ml BMP-2 in a collagen membrane was placed in extracted (hypofunctional) and non-extracted (functional) groups (BMPe and BMPf, respectively) while control groups had collagen membrane only (CONe and CONf). Groups were sacrificed at 10 (BMPe, BMPf, CONe, CONf) or 35 days (BMP35e, BMP35f, CON35e, CON35f) postoperatively and tissues processed for histological examination. Transverse 5 microm sections were stained for identification of new bone, ankylosis and cementum formation. RESULTS: At 10 days, CONe developed greater bone growth compared with CONf (P<0.05), while both BMP groups developed greater bone compared with controls. However, BMPe developed more ankylosis compared with both CONe and CONf while BMPf was significantly greater than CONf only (P<0.05). BMPf only developed significantly greater new cementum compared with controls. At 35 days, BMP35f developed greater bone growth compared with all other groups including BMP35e (P<0.05) and unlike results at 10 days, no differences were apparent between CON35f and CON35e. Unwanted bone growth beyond the defect margin anteriorly was significantly greater in BMP35f. CONCLUSIONS: Results suggest hypofunction stimulates early bone formation. Furthermore, hypofunction and BMP-2 increase the development of transient ankylosis. However, after wound healing is complete, function augments the early effects of BMP-2-induced new bone growth indicating remodeling to physiological levels does not occur. Finally, occlusal loading is both an important stimulus for remodeling and establishment of the periodontal ligament space during early wound healing as well as enhancing BMP-2 induced cementogenesis. PMID- 10534066 TI - Implications of minocycline, platelet-derived growth factor, and transforming growth factor-beta on inflammatory repair potential in the periodontium. AB - BACKGROUND: Semisynthetic tetracyclines used in the adjunctive treatment of inflammatory periodontal disease enhance collagen expression in induced periodontal lesions of rats. Polypeptide growth factors regulate key cellular events in tissue repair. The physiologically active androgen 5 alpha dihydrotestosterone (DHT) stimulates bone and connective tissue turnover. It was relevant to study the effects of transforming growth factor beta (TGF beta)/platelet-derived growth factor (PDGF) and minocycline alone and in combination on the formation of biologically effective androgens which can influence repair. METHODS: Confluent monolayer cultures of human gingival fibroblasts of the fifth through the ninth passage were incubated in Eagle's minimum essential medium, with 14C-testosterone/14C-4-androstenedione in the presence or absence of optimal concentrations of TGF-beta/PDGF/minocycline (M), alone and in combination. At the end of a 24-hour incubation period, the medium was analyzed for steroid metabolites and quantified using a radioisotope scanner. RESULTS: The androgen substrates 14C-testosterone (14C-T) and 14C-4 androstenedione (14C-4-A) were metabolized to DHT and 4 androstenedione/testosterone respectively. There were significant increases in the formation of DHT from 14C-T in response to M, TGF-beta, and PDGF, alone and in combination (13 to 48%), compared with controls (n = 4; P<0.01). The yields of 4-androstenedione were also greater in response to these agents (31%; 3-fold). When 14C-4-A was used as substrate, there were 21 to 80% increases in the formation of DHT in response to these agents alone and in combination (n = 4; P<0.01). CONCLUSIONS: The biologically effective androgen metabolites formed in response to minocycline, TGF-beta, and PDGF can contribute to reparatory events in the inflamed periodontium. Judicious, adjunctive usage of the chemically modified tetracyclines in the treatment of periodontal diseases can obviate the risk of microbial resistance, with potential applications of their anti inflammatory and proanabolic effects in regenerative technology. PMID- 10534067 TI - Human periodontal ligament fibroblast behavior on chemically conditioned dentine: an in vitro study. AB - BACKGROUND: Chemical root conditioning is widely used in an attempt to improve the outcome of regenerative periodontal surgery, but its effect on connective tissue cell proliferation and biosynthetic activity has been poorly studied. The goal of the present study was to test in vitro the consequences of conditioning human dentine by citric acid or minocycline on the behavior of attached human periodontal ligament (HPDL) cells in terms of proliferation, protein synthesis and morphological appearance. METHODS: HPDL cells were seeded on powdered human dentine, either untreated or conditioned for 3 minutes with 3% citric acid or 2.5% minocycline HCI. Scanning (SEM) and transmission (TEM) electron microscopic observations were performed, and 3H-thymidine and 3H-proline incorporation tests were used to evaluate the proliferative and the biosynthetic activities. RESULTS: Cell spreading was already evident and the penetration of cytoplasmic processes into dentinal tubules were frequently observed on all dentine types after 2 hours of attachment. After 24 hours of incubation, citric acid conditioning promoted an intense spreading of the cells, while minocycline HCI conditioning induced the formation of a dense feltwork of cellular processes. HPDL fibrolasts adherent to both types of surface-conditioned dentine exhibited a significantly higher rate of proliferation (P<0.01) as well as a significantly higher level of total protein and of collagen synthesis (P<0.01) than on untreated dentine. CONCLUSIONS: These data suggest that during periodontal surgery a conditioning of the root surface by citric acid or by minocycline HCI could promote the attachment, the proliferation, and the biosynthetic activity of HDPL, prerequisites to periodontal regeneration. PMID- 10534068 TI - Relationship between a self-reported health questionnaire and laboratory tests at initial office visits. AB - BACKGROUND: Dental patients routinely complete a medical questionnaire and have an oral interview during their initial visit, but may have undiagnosed systemic problems which can affect their dental treatment. METHODS: Thirty-nine consecutive patients referred for a periodontal evaluation completed a medical questionnaire and an oral interview. They were referred to a hospital laboratory for an urinalysis, complete blood count, and a standard blood chemistry panel. RESULTS: The self-reported medical history responses were compared with the laboratory data and several abnormalities were noted. Abnormal levels were found with cholesterol, (26/39 patients, 67%); triglycerides, (13/39, 33%); glucose, (6/39, 15%); eosinophils, (18/39, 46%); and monocytes, (10/39, 26%). Smokers (17/39, 44%) had a higher number of abnormal levels or percentages of cholesterol, triglycerides, basophils, lymphocytes, eosinophils, and monocytes. Gender differences were seen in elevated triglyceride levels, abnormal aspartate aminotransferase (AST) and alanine aminotransferase (ALT), and self-reported cardiovascular disease. CONCLUSIONS: This study demonstrated that many patients are unaware of their current medical status and a significant number had undiagnosed abnormalities. PMID- 10534069 TI - Antisense oligonucleotide of tissue inhibitor of metalloproteinase-1 induces the plasminogen activator activity in periodontal ligament cells. AB - BACKGROUND: Matrix metalloproteinases (MMPs), produced by both infiltrating and resident cells of the periodontium, play a role in physiologic and pathologic events. It is recognized that an imbalance between activated MMPs and their endogenous inhibitors leads to pathologic breakdown of the extracellular matrix during periodontitis. Although it is known that pro-MMPs are activated by the plasminogen activator (PA)/plasmin system, and that the activated MMPs are inactivated by tissue inhibitor of metalloproteinases (TIMPs), participation of TIMPs in the PA/plasmin system has not been defined. METHODS: We investigated the effects of the antisense oligonucleotide, consisting of a 21-base sequence from the human TIMP-1 gene including the first ATG initiation codon, on PA/plasmin activities in the cultured medium of periodontal ligament (PDL) fibroblastic cells. Antisense or sense oligonucleotides were directly added into cell-cultured medium, and enzyme activities from the PDL cells were measured. RESULTS: Antisense TIMP-1 oligonucleotide specifically stimulated the PA activity dose dependently. Other oligonucleotides, sense TIMP-1 or antisense TIMP-2, did not affect PA activity in PDL cells. The PA activity increased by antisense TIMP-1 oligonucleotide was due to an increase of urokinase-type PA (uPA) protein, but not that of tissue-type PA by means of immunoblotting. Furthermore, the stimulation of PA activity in the conditioned medium by adding antisense oligonucleotide for TIMP-1 was not due to the decreasing levels of PA inhibitor 1, an inhibitor of PA. CONCLUSIONS: TIMP-1 controls the synthesis of uPA in the PDL cells. Control of the TIMP-uPA system is important in inflammatory periodontal ligament healing. PMID- 10534070 TI - Smoking influences decision making in periodontal therapy: a retrospective clinical study. AB - BACKGROUND: Mechanical periodontal therapy consists of a non-surgical course, followed by surgical treatment to eliminate or reduce remaining pathological pockets. Only if diligent mechanical therapy fails are additional measures considered. It has been documented that smoking interferes with the host defense mechanisms. This study addresses the question is meticulous non-surgical periodontal therapy equally successful in smokers and non-smokers? If not, is a thorough and cumbersome non-surgical approach in smokers worth undertaking? METHODS: Thirty-five smokers and 35 non-smokers were selected retrospectively from a pool of 306 patients treated in a private practice over a 17-month period. All had at least 14 teeth present with 8 presenting with gingival pockets > or =6 mm. Non-surgical treatment was performed in 6 to 10 appointments and results were evaluated 6 to 12 weeks after therapy. Bleeding on probing sites with probing depths > or =5 mm were then considered for surgical treatment. RESULTS: Before treatment smokers had statistically significantly higher mean percent of pockets 4 to 5 mm and > or =6 mm (40.36+/-10.65 and 26.51+/-11.95, respectively, compared to 30.38+/-7.57 and 20.42+/-10.03 for non-smokers) and showed significantly lower proportional reduction of these parameters with treatment (50.80+/-33.76 and 81.36+/-19.82 for pocket 4 to 5 mm and 6 mm, compared to 68.43+/-21.23 and 91.7+/ 8.92 for nonsmokers). A multivariate analysis gave smoking, plaque control, and initial percent of sites > or =6 mm to be significant predictors of the percent of teeth in need of further therapy. In non-smokers, treatment was apparently successful in all tooth types with the exception of upper first and second molars (28.5% failure) and lower second molar (20% failure). In smokers, rates of further treatment needs were particularly high in the premolar-molar area in both jaws, ranging from 31.4% to 48.5% for an individual tooth type; 42.8% of smokers and 11.5% of non-smokers needed further treatment in 16% of their teeth (pretest probability). A decision analysis showed that for smokers with at least 1 of 5 sites > or =6 mm, one should initiate surgical treatment, rather than first treat non-surgically. If the point of indifference that the decision is correctly set at 95%, the pretest probability should be >12%. There is a higher risk that non surgical therapy will fail, for instance if we lower the point of indifference to 60%, the pretest probability should be >31%. CONCLUSIONS: It is concluded that smoking impairs healing after nonsurgical periodontal therapy. The decision analysis of this study questions the need for a thorough course of non-surgical treatment in smokers with advanced periodontal disease. PMID- 10534071 TI - An 18-year longitudinal study of untreated mucogingival defects. AB - BACKGROUND: A study was conducted to observe the changes in areas with untreated mucogingival defects over an 18-year period. The results in this group after 4 and 10 years were previously published. METHODS: Upon entering dental school, a group of 39 freshman dental students were assessed for plaque index, gingival index, probing depth, and width of keratinized tissue. At that time, 112 sites of inadequate keratinized gingiva were found. Seventeen of the original 39 participants with a total of 61 sites were reassessed for the same parameters after 18 years. RESULTS: The results revealed that 19 sites showed a slight increase in keratinized tissue, 35 were unchanged (for a total of 54 stable sites), and 7 sites showed a slight decrease in keratinized tissue. The mean width of keratinized tissue at the beginning of the study was 1.74+/-0.545 mm and 2.02+/-0.885 mm after 18 years. This represented a small, but statistically significant increase in the width. The plaque index (PI) and gingival index (GI) of this group at baseline (PI = 0.77+/-0.439 and GI = 0.93+/-0.447) and at 18 years (PI = 0.36+/-0.344 and GI = 0.65+/-0.303) indicated a high level of oral hygiene and gingival health. CONCLUSIONS: It was concluded that in the absence of gingival inflammation, areas with small amounts of keratinized tissue may remain stable over long periods of time. PMID- 10534072 TI - Spontaneous repositioning of pathologically migrated teeth. AB - BACKGROUND: There is limited clinical evidence that pathologically migrated teeth may reposition themselves after conventional periodontal treatment. The current research was carried out to determine the frequency of spontaneous repositioning of pathologically migrated teeth after routine periodontal therapy, and to study the relation between the severity of migration and the degree of repositioning following treatment. METHODS: Sixteen patients with moderate to severe periodontal disease and presenting 33 diastema sites secondary to pathologic migration participated in this study. After conventional periodontal treatment had been performed, reactive repositioning was assessed by measuring the space between pathologically migrated teeth and adjacent teeth on study models obtained at baseline, re-evaluation at 6 weeks after scaling and root planing, and 4 months after surgery. RESULTS: After scaling and root planing only, 48.5% of all sites exhibited some degree of repositioning with 36.4% of all sites closing completely. After surgery (6 months after baseline observations), 69.7% of all sites exhibited some degree of repositioning with 51.5% of all sites closing completely. When only small to moderate diastemata were considered (<1 mm), 77.8% of sites closed completely. CONCLUSIONS: The findings of this study support the hypothesis that spontaneous repositioning after conventional periodontal treatment is likely, particularly when only light to moderate degrees of pathologic migration are considered. We hypothesize that this spontaneous movement is due to wound contraction during healing. PMID- 10534073 TI - Humoral immunity to stress proteins and periodontal disease. AB - BACKGROUND: There is evidence that microbial heat shock (stress) proteins (Hsp) are immunodominant antigens of many microorganisms. Immunity to these proteins has been shown in non-oral infections to contribute to protection. This study was undertaken to assess the relationship(s) between immunity to human and microbial heat shock proteins, periodontal disease status, and colonization by periodontal disease-associated microorganisms. METHODS: Subgingival plaque and blood samples obtained from 198 patients during an earlier clinical study were examined for the presence of specific periodontal disease-associated microorganisms and antibodies to selected human and microbial heat shock proteins (Hsp70, Hsp90, DnaK, and GroEL). Particle concentration immunofluorescence assay (PCFIA) was used to detect anti-Hsp antibodies and slot immunoblot assay (SIB) was used to detect subgingival plaque species. Regression models were used to examine the contribution of age, gender, gingival index, probing depth, attachment loss, calculus index, plaque index, and microbial colonization to the anti-Hsp antibody concentrations. RESULTS: Our studies demonstrated that, when evaluated by ANOVA, patients with higher anti-Hsp (Hsp90, DnaK, and GroEL) antibody concentrations tended to have significantly (P< or =0.05) healthier periodontal tissues. This was most obvious when the relationship between mean probing depths and antibody concentrations were studied. For Hsp90 antibodies, 2 variables (probing depth and P. gingivalis concentration) were found to have significant contributions (R = 0.293, P<0.0002). The equation derived from the regression model was y = 12558 2070*PD +1842*PG. This confirmed the inverse relationship with probing depth and the positive relationship with colonization by P. gingivalis. Attempts to model the other stress protein antibodies were not successful. CONCLUSIONS: We believe that the present observations reflect the presence of protective anti-Hsp antibodies, rather than simply the presence of the microorganism in the gingival sulcus. The clinical significance of these observations lies in the potential of identifying patients who are at risk for developing periodontal disease based on their inability to mount an immune response to specific Hsp or Hsp epitopes, as well as the development of vaccines based on Hsp epitopes. PMID- 10534075 TI - Arbitrarily primed-polymerase chain reaction for identification and epidemiologic subtyping of oral isolates of Fusobacterium nucleatum. AB - BACKGROUND: Fusobacterium nucleatum is the most frequently isolated bacterium in periodontal disease and plays an important role in serious infections in other parts of the body. Arbitrarily primed-polymerase chain reaction (AP-PCR) was used to construct primers for specific identification and subtyping of F. nucleatum. Subtypes may differ in virulence and, hence, are important as periodontal pathogens. Subtypes also may differ in antibiotic susceptibility; therefore, knowing the subtypes may influence choice of treatment. METHODS: We analyzed 70 DNA samples of F. nucleatum isolated from patients with periodontal disease (PD) (N = 32) or AIDS-related PD (N = 8) and from healthy carriers (N = 30). From 90 AP-PCR primers screened, five amplification products were selected, cloned in pCR II vector, and sequenced. These sequences were used to design new pairs of specific primers. Sequences were compared to GenBank entries with BLAST and showed no significant matches. RESULTS: Three primer pairs produced bands of approximately 1 Kb (primer 5059S) or 0.5 Kb (primers FN5047 or M1211) with all F. nucleatum DNAs tested. PCR amplification using primer pair M8171 produced a 1 Kb band with isolates from 7 (22%) PD and 5 (63%) PD-AIDS patients and 9 (30%) healthy controls. Using the same primer pair, 2 other bands of approximately 0.5 Kb and 0.4 Kb were observed with DNA from isolates from 2 (6%) PD and all PD-AIDS patients, but were not observed with DNA samples from healthy controls (P<0.0001). All the primer pairs produced no or different amplicon profiles with DNA samples from bacterial species other than F. nucleatum. CONCLUSIONS: Our results suggest that PCR primer pairs 5059S, FN5047 or M1211 can be used to specifically identify F. nucleatum isolates and distinguish them from other bacteria. The primer pair M8171 could also be used to differentiate F. nucleatum isolated from periodontal patients or healthy individuals. These specific primers can be used in PCR analysis for specific identification of F. nucleatum and to distinguish it from other bacteria associated with human periodontitis. These approaches appear promising in facilitating laboratory identification, molecular subtyping, and taxonomy of putative periodontopathogens. PMID- 10534074 TI - Single nucleotide polymorphisms of the N-formyl peptide receptor in localized juvenile periodontitis. AB - BACKGROUND: Neutrophils from patients with localized juvenile periodontitis (LJP) exhibit decreased binding and responsiveness to various chemotactic agents, including N-formyl-1-methionyl-1-leucyl-1-phenylalanine (FMLP). This altered reaction of neutrophils is thought to account in part for the increased susceptibility of LJP patients to infections by periodontal organisms. Receptors for FMLP are involved in the activation and the subsequent response to certain chemotactic stimuli. METHODS: In order to determine if this decreased response is due to a genetic variation in the receptor, we directly compared DNA encoding the FMLP receptor from controls matched for gender and race and LJP patients by single-strand conformation polymorphism analysis (SSCP). RESULTS: Using this technique, we observed a characteristic SSCP pattern in 29 out of 30 patient samples in the FMLP receptor DNA. This pattern differed from those obtained from the 20 control subjects as well as 31 patients with adult periodontitis. DNA sequencing of 30 patients indicated single nucleotide polymorphisms (SNPs) in the FMLP receptor DNA from the LJP patients when compared to 20 controls (P = 0.0005). Two single nucleotide base alterations were consistently seen: either a thymine to cytosine substitution at base 329 in 17 LJP patients or a cytosine to guanine substitution at base 378 in 5 LJP patients. A combination of both alterations were seen in 7 LJP patients. Both alterations resulted in amino acid changes in the second intracellular loop of the receptor, specifically phenylalanine to serine at residue 110 and cysteine to tryptophan at residue 126. This region of the FMLP receptor has recently been shown to play a role in ligand binding and G-protein activation. CONCLUSIONS: This study suggests that a molecular alteration in the second intracellular loop of the FMLP receptor molecules in LP patients may play a role in the decreased chemotactic activity reported for some LJP patients. PMID- 10534076 TI - Progression and treatment of chronic adult periodontitis. AB - BACKGROUND: The periodontal status of 41 medically healthy adults with untreated chronic periodontitis was monitored before and after scaling and root planing (SRP). METHODS: During a 6-month pretreatment phase, clinical measurements, digital subtraction radiography (DSR) analysis of alveolar bone, and measurement of gingival crevicular fluid (GCF) prostaglandin E2 (PGE2) levels were undertaken. SRP was provided during a 1-month treatment phase. Clinical, radiographic, and biochemical analyses were repeated in a 6-month post-treatment healing period. RESULTS: Pretreatment: no clinically significant changes in mean plaque indices (PI), probing depths (PD), bleeding on probing (BOP), or relative clinical attachment levels (CAL) were detected (P>0.05). DSR revealed small but statistically significant bone height (0.04 mm) and mass (0.97 mg) loss (P<0.001). GCF PGE2 levels gradually increased from 38.8 ng/ml at month 1 to 79.4 ng/ml at month 6. Post-treatment: statistically and clinically significant reductions were observed in mean PI, BOP, and PD (P<0.05). A statistically significant reduction in CAL was noted (P<0.05). The trend towards progressive bone loss was halted and reversed, and a statistically significant decrease in GCF PGE2 concentrations was detected (P<0.001). Smokers, non-smokers, and ex smokers did not differ significantly in PI, BOP, CAL, radiographic, or biochemical parameters at any time. Mean PD was significantly greater in current smokers than in non- and ex-smokers (P<0.005). PD reduced comparably in all 3 smoking subgroups following treatment (P<0.01). CONCLUSIONS: Conventional clinical measurements failed to identify disease progression over a 6-month period. Significant improvements were observed in clinical parameters after SRP, and a trend towards progressive bone loss was halted and reversed. Regular and frequent maintenance visits are important following treatment to maintain improvements in clinical parameters. Smokers had deeper probing depths than non- and ex-smokers, but pockets were reduced significantly and comparably in all 3 smoking subgroups following efficacious treatment. PMID- 10534077 TI - Changes in gingival crevicular fluid levels of immunoglobulin A following therapy: association with attachment loss. AB - BACKGROUND: In previous studies, we demonstrated that increased levels of immunoglobulin A (IgA) in gingival crevicular fluid (GCF) may be "protective", while increased levels of the polymorphonuclear lysosomal enzyme, beta glucuronidase, in GCF were associated with increased risk of disease activity. In this study, we examined the effect of scaling and root planing (SRP) on the levels of beta-glucuronidase, IgG, and IgA in GCF over a 24-week period and compared these to clinical attachment loss (CAL). METHODS: Twenty-nine patients with periodontal disease were examined for attachment level, probing depth, plaque, and bleeding on probing at 6 sites per tooth. GCF was collected from the mesial aspect of all teeth excluding third molars and analyzed for beta glucuronidase, IgG, and IgA. After baseline data were collected, each patient received SRP, and GCF was collected again at 2, 4, 6, 8, 12, and 24 weeks post SRP while clinical data were obtained at 4, 8, 12, and 24 weeks. In addition, we analyzed whether the magnitude of the IgA response to SRP would affect the rate of periodontal disease progression by examining GCF IgA levels at 2 time intervals: 2 to 4 weeks post-SRP and 6 to 12 weeks post-SRP. RESULTS: Seventeen patients (58.6%) exhibited at least 1 site losing > or =2.5 mm of CAL during the 24-week study. Beta-glucuronidase in GCF was significantly decreased at 2 weeks following SRP and then demonstrated a gradual increase throughout the study period. Levels of IgA in GCF significantly increased following SRP, reaching a peak at 6 weeks and then gradually decreasing throughout the study. Furthermore, we found an inverse relationship between GCF IgA levels at 6 to 12 weeks post-SRP and the occurrence of CAL. CONCLUSIONS: These results support the hypothesis that maintenance of high levels of IgA in GCF may be "protective" against periodontal attachment loss. Furthermore, levels of beta-glucuronidase appear to be a more sensitive indicator of gingival inflammation than clinical measures. PMID- 10534078 TI - Qualitative analysis of peripheral peri-implant bone and influence of alendronate sodium on early bone regeneration. AB - BACKGROUND: Alendronate sodium increases alveolar bone density with systemic use. It inhibits osteoclast activity and is thought to result in a net increase in osteoblastic activity. However, little is known about local in vivo use. The purpose of this study was to evaluate the effect of local delivery of alendronate on bone regeneration within peri-implant defects. Peri-implant bone was examined histomorphometrically to evaluate the amount of supporting bone peripheral to the bone-implant interface. METHODS: Six adult hound dogs were evenly divided into 2 groups, with one group receiving alendronate-coated dental implants and the other group serving as controls. Dental implants were placed immediately after extraction of right and left second, third, and fourth mandibular premolars. Forty-eight dental implants were placed (2 types in each dog: 24 hydroxyapatite [HA]-coated and 24 titanium machine-polished [TMP]), for a total of 4 variables. A bioabsorbable collagen membrane was secured over the implants and defects, and the flaps closed primarily. The dogs were sacrificed on day 28. Specimens were sectioned, mounted, and stained with Stevenel's blue and van Gieson's picric fuchsin. The amount of bone adjacent and 1 mm peripheral to the implant surface was recorded with a computerized microscopic digitizer. RESULTS: Locally applied alendronate resulted in significantly increased amounts of bone (P<0.0002, ANOVA) in the peripheral area with both HA and TMP implants. However, the most influential factor in the amount of peripheral bone was the type of implant surface (P<0.0001). CONCLUSIONS: Local application of alendronate is useful in increasing the amount of peripheral peri-implant bone. Also, the amount of supporting bone was not related to the bone-to-implant contact but to the surface characteristics of the implant. The findings of the present study indicate that the evaluation of dental implant-supporting bone should include peripheral bone as well as bone-to-implant interface. PMID- 10534079 TI - Culture of gingival fibroblasts on bioabsorbable regenerative materials in vitro. AB - BACKGROUND: The use of membranes in guided tissue regeneration (GTR) can limit the apical migration of gingival cells and favor the establishment of new attachment by periodontal ligament fibroblasts. However, gingival recession during healing following GTR has been described as a frequent complication. The purpose of this study was to determine if gingival fibroblasts are affected by the composition of the bioabsorbable membranes used in mucogingival surgery. METHODS: Two type of bioabsorbable regenerative materials were used as cell carriers. Wistar rat gingival fibroblasts (RGF) were obtained from attached gingiva, cut into small fragments, and placed in culture dishes. When confluent, cells were detached using trypsin and identified as "first transferred cells" (P1). At the third passage (P3), cell count, trypan blue exclusion test, acid phosphatase activity, DNA synthesis, phase contrast microscopy, and scanning electron microscopy were performed. The cells were then placed in wells containing the membranes and incubated for 72 hours. RESULTS: When examined under microscopy, the control wells (without membranes) showed one cell type with the elongated appearance characteristic of fibroblasts. The wells with membranes showed an altered cell morphology with a high proportion of cell fragments regardless of the type of membrane used. CONCLUSIONS: These results suggest that cell carrier membranes could affect RGF morphology and thus alter gingival tissue healing following GTR. PMID- 10534080 TI - The effect of smoking and periodontal treatment on salivary composition in patients with established periodontitis. AB - BACKGROUND: The purpose of this study was to evaluate the effect of smoking on the periodontal status and the salivary composition in subjects with established periodontitis before and after periodontal therapy. METHODS: Our study group included 26 healthy subjects, 12 smokers and 14 non-smokers with established periodontitis. Clinical measurements and non-stimulated whole saliva were obtained and analyzed at baseline and after scaling and root planing. Smokers presented at baseline with significantly greater probing depth (4.16+/-0.26) compared to non-smokers (3.52+/-0.32) which was statistically significant (P = 0.0268); likewise, baseline clinical attachment level was greater in smokers (4.49+/-0.31 compared to non-smokers 3.87+/-0.13; P = 0.0620). Mean plaque index was also greater in smokers compared to non-smokers (0.86 and 0.65, respectively; P = 0.0834). Baseline pretreatment sodium values were significantly greater in non-smokers (14.36 mEq/l compared to 9.31 mEq/l in smokers; P = 0.0662); likewise non-smokers exhibited 50% greater salivary calcium levels (6.04 mg/100 ml compared to 4.32 mg/100 ml in smokers; P = 0.0133). RESULTS: Post-treatment probing depth and clinical attachment level were not different between smokers and non-smokers; this in spite of significant difference in plaque index in smokers (0.35 compared to 0.13 in non-smokers; P = 0.0135). Post-treatment, smokers had reduced calcium concentration (3.58 mg/100 ml compared to 5.11 mg/100 ml in non-smokers; P = 0.0438). Treatment affected albumin level in smokers only, consequently non-smokers had significantly greater salivary albumin concentration (1.1 mg/100 ml compared to 0.38 mg/100 ml in smokers; P = 0.0274). CONCLUSIONS: Subjects with established periodontitis exhibited elevated concentrations of salivary electrolytes and proteins. Within this study group, smokers exhibited greater disease level but reduced sodium, calcium, and magnesium concentrations. Smokers responded favorably to treatment. The clinical improvement eliminated the differences in salivary composition. PMID- 10534081 TI - Gingival hemorrhage, myelodysplastic syndromes, and acute myeloid leukemia. A case report. AB - Myelodysplasia syndrome (MDS) presenting as spontaneous gingival hemorrhage is described. Gingival hemorrhage is recognized as a symptom of MDS, a rare group of potentially fatal hematological disorders, but it has not previously been documented as a presenting sign. The diagnostic pitfalls are discussed with the case, and the need for careful interpretation of laboratory findings in conjunction with clinical signs is emphasized. Finally, the MDSs are defined, classified and discussed with respect to their relevance to the clinical periodontist, from a diagnostic, therapeutic, and management standpoint. PMID- 10534082 TI - Periodontal and computer tomography scanning evaluation of endosseous implants in conjunction with sinus lift procedure. A 6-case series. AB - The aim of the present study was to evaluate endosseous implants in conjunction with the sinus lift procedure using periodontal parameters and computed tomography (CT) scanning. In 6 patients, 8 sinus lift procedures were performed to the atrophic posterior maxilla. Fourteen endosseous implants were placed. The condition of the peri-implant mucosa was assessed postoperatively at an average of 31 months using periodontal parameters such as plaque index, probing depth, clinical attachment level, and bleeding on probing. All the implants were maintained in a clinically healthy condition throughout the observation period. CT images revealed the difference in the morphology of the sinus floor between the locations of the block-shaped bone and the small pieces of bone which were both grafted. The results indicated that endosseous implants in conjunction with the sinus lift procedure could be maintained by monitoring the periodontal parameters. PMID- 10534083 TI - Family practice patients' attitudes toward firearm safety as a preventive medicine issue: a HARNET Study. Harrisburg Area Research Network. AB - BACKGROUND: Firearm-related deaths are expected to outnumber motor-vehicle related deaths within the next 5 years. The goal of this project was to document gun ownership and safety habits among patients of family physicians and to determine patients' attitudes toward physician counseling about firearm safety. METHODS: We conducted a cross-sectional survey of patients of 11 family practices affiliated with a research network. Patients (or parents of children) were asked to complete a 23-item questionnaire that asked about gun ownership, storage, sources of information about gun safety, and attitudes about gun safety, and the role of their physician. RESULTS: Of the 1359 questionnaires distributed, 1214 (89%) were returned. Gun ownership varied by location (urban, suburban, and rural) and ranged from 16% to 59%. Only 8% of respondents believed that their physician has a responsibility to discuss firearm safety. Most (91%) patients did not view their physician as a source of firearm safety information, and only 14% thought their physician was knowledgeable regarding gun safety. Most gun owners (71%) did not think they would follow their physicians' advice about gun storage. Most patients (5%) responded that gun safety should not be discussed during the office visit. CONCLUSION: With regard to firearm safety, family physicians lack credibility in the eyes of their patients. These patients do not appear to be receptive to information about firearm safety, and efforts to decrease firearm related injury might be more effective if focused elsewhere. PMID- 10534084 TI - A comparison of flat and shallow conical tips for cervical cryotherapy. AB - BACKGROUND: Although two types of cervical cryotherapy tips are widely used, there have been no randomized prospective comparison studies reported in the medical literature. Shallow conical tip proponents theorize that a greater depth of freeze near the os yields better treatment of cervical intraepithelial neoplasia (CIN) without elevating the squamocolumnar junction into the cervical canal. Flat tip proponents theorize an equally effective CIN treatment with lower incidence of posttreatment squamocolumnar junction location in the cervical canal. METHODS: A comparative descriptive study was performed to evaluate 117 cryotherapy candidates with biopsy-proved CIN who were classified by location of their squamocolumnar junctions (ectocervix, at the os, or in the canal). They were then randomized to receive double-freeze cervical cryotherapy with either a flat or shallow conical tip. Four or more months later, repeated colposcopy and Papanicolaou smears were performed to assess resolution of CIN and posttreatment location of the squamocolumnar junction. RESULTS: Eighty-four patients (71%) completed the study. Analysis indicated no important difference between the two tips in eliminating CIN. The squamocolumnar junction was colposcopically visualized at all posttreatment examinations. When the pretreatment squamocolumnar junction location was on the ectocervix, data analysis indicated that squamocolumnar junction movement was greater with the shallow conical tip (P = .037), particularly into the canal, where it is clinically more difficult to visualize (P = .019). There were no significant differences in movement of the squamocolumnar junction when it was originally at the os or in the canal. CONCLUSIONS: This study found no significant difference in effectiveness of the two types of tips in eliminating CIN and supports the practice of using one type- either flat or shallow conical tips--to treat all candidates for cervical cryotherapy. Using the flat tip when the pretreatment squamocolumnar junction is on the ectocervix will allow easier posttreatment visualization of the squamocolumnar junction. Further studies with a greater number of subjects are indicated. PMID- 10534085 TI - Multiple marker screening for Down syndrome--whom should we screen? AB - BACKGROUND: The multiple marker test highlights the complexities of an expanding number of genetic tests that family physicians can offer to their patients. Many concerns surround the use of the multiple marker test, including a poor understanding of the test by physicians and patients, limited sensitivity, increased anxiety among women, and the risks of amniocentesis in patients who have false-positive results. METHODS: An online search of the medical literature was used to select English-language articles addressing the burden of suffering from Down syndrome, and efficacy, cost-effectiveness, and psychological effects of screening. RESULTS AND CONCLUSIONS: Down syndrome occurs in 0.99 per 1000 births, and the risk increases with age. Between 10% and 20% of infants die in the first year, predominantly from heart defects. Special education, the move away from institutionalization, and more employment opportunities have improved the outlook for these persons. The multiple marker test has an overall detection rate for Down syndrome of 56% with a 7% false-positive rate. A laboratory cutoff ratio of 1:190 is the most efficient for optimizing the detection rate and minimizing the false-positive rate. Limited screening to women older than 30 years appears most cost-effective, reducing losses of normal fetuses secondary to amniocentesis and the number of pregnant patients requiring pretest counseling. The family physician needs to inform the patient of the nature, purpose, and risks of screening including the psychological effects. Pretest counseling should be nondirective. PMID- 10534087 TI - Adrenal insufficiency: an uncommon cause of fatigue. AB - BACKGROUND: Adrenal insufficiency is a rare condition that can cause common and nonspecific symptoms. One such symptom, reported by all patients with adrenal insufficiency, is fatigue. On the other hand, up to 20% of patients seeking care from primary care physicians will have fatigue as a complaint. Only a small percentage of patients are found to have underlying medical disease. METHODS: A MEDLINE literature search was performed from 1966 to the present using the key words "fatigue," "adrenal insufficiency," and "polyglandular autoimmune endocrinopathy." Major endocrinology textbooks were also referenced. In addition, references were obtained from bibliographies of available articles. RESULTS AND CONCLUSIONS: This article describes a patient with adrenal insufficiency and fatigue as the primary complaint. A brief discussion of fatigue and clues to organic causes follows, along with a more detailed discussion of adrenal insufficiency. Important medical history or signs and symptoms of organic disease suggest the need for screening tests and more detailed evaluation to uncover the uncommon medical causes of fatigue. PMID- 10534086 TI - Diagnosis and management of Alzheimer disease. AB - BACKGROUND: Alzheimer disease afflicts millions of older Americans, with an estimated cost to society approaching $100 million annually. Family physicians will care for an increasing number of patients with Alzheimer disease as well as their caregivers and families. METHODS: A comprehensive and systematic review of the literature published between 1985 and 1998 about diagnosing and treating Alzheimer disease was conducted, using "dementia," "Alzheimer's disease," and "treatment" as search strategy key words. Data and information that reported significant conclusions were critically reviewed. Potentially important new data about new agents that might be of benefit when caring for patients with Alzheimer disease are discussed. RESULTS AND CONCLUSIONS: The primary goals when treating Alzheimer disease patients are enhancing autonomy and functional abilities and maintaining quality of life for patients and caregivers. In addition to diagnostic and pharmacologic treatment, primary care physicians will be called upon to provide nonpharmacologic support to assist with behavioral, social, and living environment problems faced by these patients and their families. The most common pharmacologic treatment is cholinesterase inhibition. Two cholinesterase inhibitors, tacrine and donepezil, are effective in treating cognitive and global function. Newer cholinesterase inhibitors should soon be available that might offer safety advantages as well as efficacy in treating behavioral and psychiatric symptoms related to Alzheimer disease. Other agents, including vitamin E, nonsteroidal anti-inflammatory drugs, estrogen, and Ginkgo biloba, are under investigation. Nonpharmacologic measures are important components in the management of Alzheimer disease. Support groups can help to diminish behavioral problems, maintain the patient's independence, and provide relief for caregivers and families. PMID- 10534088 TI - Atypical infective endocarditis. AB - BACKGROUND: Although infective endocarditis has changed in the recent past as a result of microbiologic and risk factors, it continues to be clinically challenging. The disease is characterized by the formation of septic masses of platelets on the surfaces of heart valves. Several mechanisms can cause or contribute to the development of endocarditis. Although risk factors for infective endocarditis are well known, patients with atypical signs and symptoms continue to challenge us. METHODS: We describe a case report of a patient admitted to our inpatient service with back pain and presumed pyelonephritis. A MEDLINE literature search was conducted, using the key words "endocarditis," "back pain," and "bacterial," for the years 1986 to the present. RESULTS AND CONCLUSIONS: A 42-year-old woman with a history of intravenous drug abuse was admitted to the family practice service with back pain and pyelonephritis. She developed hypoxia and a new heart murmur and had continued fevers. Blood cultures drawn in the emergency department grew methicillin-resistant Staphylococcus aureus. A bone scan and magnetic resonance imaging led to the diagnosis of epidural abscess. What appeared to be a simple case of pyelonephritis with back pain became a case of infective endocarditis complicated by an epidural abscess. PMID- 10534090 TI - Thoracic outlet compression syndrome: a rare cause of upper extremity symptoms in children. PMID- 10534089 TI - Evaluating evidence from a decision analysis. PMID- 10534091 TI - Neurosarcoidosis: case report and brief literature review. PMID- 10534092 TI - Enhanced obstetrics training for family practice residents: a unique collaborative program. PMID- 10534093 TI - Gun safety and the family physician. PMID- 10534094 TI - Good things still come in old packages: cryosurgery vs LEEP. Loop electrosurgical excision procedure. PMID- 10534096 TI - In-flight radiation. PMID- 10534095 TI - Economics of treatment guidelines for hypertension. PMID- 10534097 TI - Inpatient care of children. PMID- 10534098 TI - Professional identity and names. PMID- 10534099 TI - Traffic in the brain: how life experience can govern physiological function and personal behavior and disturbances as well. PMID- 10534100 TI - Odogenic change of blood coagulation in humans. AB - The blood coagulation time as an integral sign of human homeostasis is revealed to be influenced by conscious or unconscious olfactory stimuli through autonomic mechanisms. The hemostasis of subjects with predominance of sympathetic activity is more sensitive to the olfactory stimuli than that of subjects with vagal prevalence. The effects of the olfactory stimuli upon hemostasis in humans support the viewpoint that simultaneous information and physico-chemical processes act together in parallel to play and important role in life activities of human organism (Simonov, 1981; Sudakov, 1995). Emotions, therefore, are essential in the transfer of information from the environment (Anokhin, 1966; Miltner et al., 1994). It is not excluded that even very weak stimuli which are subthreshold for conscious perception might be informational for the organism as those engaging emotional response. PMID- 10534101 TI - Challenging dogmas and the advance of knowledge. PMID- 10534102 TI - Heterotopic and homotopic classical conditioning of the baroreflex. PMID- 10534103 TI - Stimulation-induced behavioral inhibition: a new model for understanding physical violence. AB - Physical violence is widely considered to result from action carried out with the intention of causing injury; that is, from aggression. However, the "hypothesis" of aggression is inapplicable in all but a few instances as well as inappropriate for many destructive, rage-associated responses directed at inanimate objects. This paper outlines a new perspective on physical violence, reinterpreting many behaviors hitherto labeled aggressive as stimulation-seeking behaviors (SSBs) above an arbitrary level of intensity. It is further proposed that: 1) physical violence is a by-product of SSB, driven in part by brain catecholaminergic (CA) systems, and the direct result of exchanges of energy that exceed the body's tolerance threshold; 2) allegedly discrete categories of motor-motivational behavior represent overlapping bands of intensity on a continuous spectrum of SSB; and 3) the sensory input derived from SSB is fed back into the central nervous system where it activates brain serotonergic and/or cholinergic systems, which in turn inhibit CA systems, resulting in a general state of behavioral quiescence. In addition to accounting for a number of previously unexplained observations, the model suggests that physical violence could be prevented by providing groups at high risk with extensive opportunities for therapeutic sensory stimulation to substitute for that derived from excessive SSB. For people at especially high risk, portable devices could be developed that would allow the user to self-administer desired levels of sensory stimulation at moments of intense anger, thereby preventing potentially dangerous outbursts of SSB prior to the onset of the behavior. PMID- 10534104 TI - The university and corporate biotechnology: profit margins and epistemology. PMID- 10534105 TI - Functional coupling and differential regulation of the phospholipase A2 cyclooxygenase pathways in inflammation. AB - Prostaglandins generated by the phospholipase A2 (PLA2)/cyclooxygenase (COX) pathway are well known to mediate diverse intracellular and extracellular effects that regulate mammalian development, vascular function, renal physiology, parturition, and immune responses to infection or wounding. In immune-mediated diseases and in certain cancers, this pathway is aberrantly up-regulated and excessive prostaglandin production contributes to the pathology. It is now known that there are two isoforms of COX and multiple secreted and intracellular PLA2 enzymes. The use of isoform-specific inhibitors, coupled with antisense and in vivo gene deletion experiments, has identified independent pathways of arachidonic acid metabolism, which are differentially regulated at the levels of gene expression, protein phosphorylation, and cellular localization. There is cross-talk between the pathways at the level of PLA2 and substrate supply to the two isoforms of COX is apparently compartmentalized. Knockout studies have shown that the two COX isoforms play independent roles in immediate and delayed agonist induced prostaglandin synthesis. Cytosolic PLA2-alpha is essential for both responses. Inducible secreted forms of PLA2 are, as yet, not essential for either response with the exception of the in vitro murine mast cell immediate response and instances of murine macrophage prostaglandin synthesis. These enzymes amplify the delayed response and are likely to modulate the severity of immune-mediated diseases. PMID- 10534106 TI - The actions of bacterial DNA on murine macrophages. AB - Murine macrophages are able to distinguish bacterial from mammalian DNA. The response is mimicked by single-stranded oligonucleotides containing unmethylated CG dinucleotides ("CpG" motifs) in specific sequence contexts. The dose-response curve for activation is influenced by variation in the sequence flanking the core CpG motif. CpG or bacterial DNA activates several signaling pathways in common with bacterial lipopolysaccharide (LPS), leading to induction of cytokine genes such as tumor necrosis factor alpha. Pretreatment with LPS causes desensitization to subsequent activation by CpG DNA. Both stimuli also cause cell cycle arrest in macrophages proliferating in response to the macrophage growth factor colony stimulating factor-1 (CSF-1), but prevent apoptosis caused by growth factor removal. In part, cell cycle arrest by CpG DNA and LPS may be linked to rapid down-modulation of the CSF-1 receptor from the cell surface, a response that occurs in an all-or-nothing manner. The response of macrophages to CpG DNA has aspects in common with the DNA damage response in other cell types, which may provide clues to the underlying mechanism. PMID- 10534107 TI - S100A8: emerging functions and regulation. AB - The functional importance of members of the S100 Ca2+-binding protein family is becoming apparent. Murine (m)S100A8 (initially named CP-10) is a potent chemoattractant (10(-13) to 10(-11) M) for myeloid cells and the chemotactic activity of other S100s has since been reported, suggesting a new class of chemoattractants. Murine S100A8 has been associated with a number of acute and chronic inflammatory conditions including bacterial infection, atherogenesis, and cystic fibrosis. It is expressed constitutively with S100A9 in neutrophils and is regulated by inflammatory stimulants in macrophages and microvascular endothelial cells. The lack of co-expression of S100A9 with S100A8 in activated macrophages suggests distinct functions for the proteins expressed by different cell types. Glucocorticoids up-regulate induction of mS100A8 by inflammatory mediators, and its exquisite sensitivity to oxidation suggests that it may protect against oxidative tissue damage. Inactivation of the mS100A8 gene is embryonic lethal, providing the first evidence for non-redundant function of a member of the S100 gene family. S100A8 may have an immunoregulatory role by contributing to the regulation of fetal-maternal interactions. It may play a protective role and its absence may allow infiltration by maternal cells, a process eventually manifesting as resorption. This review focuses on the variety of emerging functions attributed to murine S100A8, a protein implicated in embryogenesis, growth, differentiation, and immune and inflammatory processes. PMID- 10534108 TI - The macrophage in atherosclerosis: modulation of cell function by sterols. AB - Lipid-laden macrophage foam cells are an early and persistent component of atherosclerotic lesions. As such they are likely to play a key role in disease progression, both as scavengers of lipid and as inflammatory mediators. The sterol content of macrophage foam cells is largely native cholesterol together with a small but significant proportion of oxidized cholesterol (oxysterols). Few in vitro investigations of the influence of sterol accumulation on macrophage function have used cells that contain physiologically or even pathologically representative amounts of cholesterol or, more particularly, oxysterols. However, recent studies, using macrophages with a sterol content much closer to that of authentic foam cells, show that the presence of oxysterols causes an impairment in macrophage cholesterol export, suggesting a key role for oxysterols in the maintenance of the foam cell phenotype. The implications of physiologically relevant levels of oxysterols on a wider range of macrophage function remain to be investigated. PMID- 10534109 TI - Impact of the -308 TNF promoter polymorphism on the transcriptional regulation of the TNF gene: relevance to disease. AB - A biallelic G (TNF1 allele) to A (TNF2 allele) polymorphism 308 nucleotides upstream from the transcription initiation site in the tumor necrosis factor (TNF) promoter is associated with elevated TNF levels and disease susceptibilities observed in human subjects. The TNF2 allele is strongly associated with the high-TNF-producing autoimmune MHC haplotype HLA-A1, B8, DR3, with elevated serum TNF levels and a more severe outcome in infectious diseases, such as cerebral malaria. A number of groups have set out to determine whether the -308 polymorphism could affect transcription factor binding and hence influence TNF transcription and expression levels. Although some studies have failed to show any functional difference between the two allelic forms, others have shown that the -308 polymorphism effected transcription factor binding to the region encompassing -308, with the region in the TNF2 allele showing altered binding characteristics. The -308 polymorphism also has been found by some groups to be functionally significant in reporter gene assays in Raji B cells, Jurkat T cells, and U937 pre-monocytic cells. Up to fivefold differences can be measured between TNF1 and TNF2 allelic constructs when the TNF 3'UTR is present, indicating a role in the expression of the polymorphism. Although controversial, the majority of the data support a direct role for the TNF2 -308 allele in the elevated TNF levels observed in TNF2 homozygotes and HLA-A1, B8, DR3 individuals. Elevated TNF levels due to the -308 polymorphism may alter the immune response such that it confers susceptibility to certain autoimmune and infectious diseases. PMID- 10534110 TI - The amyloid precursor protein of Alzheimer disease in human brain and blood. AB - Studies of the metabolism and function of the amyloid precursor protein (APP) and its proteolytic fragment A beta in cultured cells, transgenic mice, and post mortem brain tissue have advanced our understanding of Alzheimer disease (AD). However, the molecular pathogenesis of the disease is still not clear, and we are a long way from finding a cure for the disease. Studies carried out on human platelets and leukocytes have also helped shed light on APP and A beta metabolism and function. Platelet and leukocyte APP isoforms are processed using mechanisms similar to those in neuronal cells to generate A beta and soluble forms of APP. The activation of platelets and leukocytes leads to the secretion of APP and A beta, resulting in higher levels of these proteins in serum. APP and A beta in the circulation may be involved in the regulation of platelet function and in the modulation of immune responses. Because human platelets and lymphocytes produce all forms of APP and secrete amyloidogenic A beta peptides, these tissues may be useful in monitoring responses to therapeutic interventions directed at APP metabolism. Although not of neuronal origin, further studies on the more accessible ex vivo tissues, including platelets and leukocytes and other blood components, may reveal potential peripheral markers for AD and will further our understanding of the molecular pathogenesis of AD. PMID- 10534112 TI - Macrophages--proliferation, activation, and cell cycle proteins. AB - Our understanding of mammalian cell proliferation has increased enormously over the past decade. A major advance has been identification and characterization of cyclins and their catalytic partners, cyclin-dependent kinases (cdks). The following brief review highlights the role of macrophages as a cell model for many of the major advances in this field. Macrophages were central to the identification of D-type cyclins and cdk4. In addition, it appears the first work showing that cell cycle proteins are the targets for anti-proliferative agents was performed in macrophages. In these latter studies, and a number of subsequent studies in other cell types, it was shown that many antimitogenic agents repressed cyclin and/or cdk expression. However, recent work in this laboratory suggests macrophage D-type cyclins may also be involved in processes other than proliferation. We have unexpectedly found that macrophages treated with lipopolysaccharide (LPS) express high levels of cyclin D2, even though LPS simultaneously represses cyclin D1 levels and potently blocks proliferation. These data, and those showing the yeast extract Zymosan A also raises cyclin D2 levels, suggest cyclin D2 plays a role in macrophage activation. PMID- 10534111 TI - Differential responses of human monocytes and macrophages to IL-4 and IL-13. AB - The primary interleukin-4 (IL-4) receptor complex on monocytes (type I IL-4 receptor) includes the 140-kDa alpha chain (IL-4R alpha) and the IL-2 receptor gamma chain, gamma(c), which heterodimerize for intracellular signaling, resulting in suppression of lipopolysaccharide (LPS)-inducible inflammatory mediator production. The activity of IL-13 on human monocytes is very similar to that of IL-4 because the predominant signaling chain (IL-4R alpha) is common to both receptors. In fact, IL-4R alpha with IL-13R alpha1 is designated both as an IL-13 receptor and the type II IL-4 receptor. When the anti-inflammatory activities of IL-4 and IL-13 were investigated on synovial fluid macrophages and compared with the responses by monocytes isolated from the patients at the same time as joint drainage, the response profiles differed with some responses similar in the two cell populations, others reduced on the inflammatory cells. Similar differences were recorded in the response profiles to IL-4 and IL-13 by monocytes and monocytes cultured for 7 days in macrophage colony-stimulating factor (M-CSF) or granulocyte-macrophage CSF (GM-CSF) (monocyte-derived macrophages, MDMac). MDMac have reduced gamma(c) mRNA levels and reduced expression of the functional 64-kDa gamma(c). There was a similar loss of IL-13R alpha1 mRNA on monocyte differentiation. In turn, there was a significant reduction in the ability of IL-4 and IL-13 to activate STAT6. These findings suggest that different functional responses to IL-4 and IL-13 by human monocytes and macrophages may result from reduced expression of gamma(c) and IL-13R alpha1. PMID- 10534113 TI - Inflammatory processes in a murine model of intra-abdominal abscess formation. AB - Abscess formation has been viewed as a host defense strategy to contain the spread of infection. However, abscesses are also serious and life-threatening manifestations of persisting microbial infection. The initiation of abscess formation, both clinically and experimentally, involves the release of bacteria and an abscess-potentiating agent (e.g., fecal fiber or an analog) into a sterile site, with host defense mechanisms being unable to eliminate the infecting organisms. Abscess formation is aided by a combination of factors that share a common feature: impairment of phagocytic killing and hence clearance of microorganisms. These include bacterial virulence factors (e.g., capsule formation, succinic acid production); complement activation by the abscess potentiating agent; fibrin deposition; and microbial sequestration within abscess neutrophils. Recruitment of cells into the peritoneal cavity follows mast cell activation in the pathogenesis of infection: histamine and tumor necrosis factor alpha can be detected in the peritoneal cavity within minutes of challenge with an abscess-inducing mixture. However, the role of mast cells in host defense is made less clear by the finding of diminished abscess formation (but no mortality or increased morbidity) in mast-cell-depleted mice. This may indicate that mast cell products have a role in not only the initiation of an inflammatory response but also the promotion of fibrin deposition and abscess formation. PMID- 10534114 TI - Suppressors of cytokine signaling (SOCS): negative regulators of signal transduction. AB - SOCS-1 was originally identified as an inhibitor of interleukin-6 signal transduction and is a member of a family of proteins (SOCS-1 to SOCS-7 and CIS) that contain an SH2 domain and a conserved carboxyl-terminal SOCS box motif. Mutation studies have established that critical contributions from both the amino terminal and SH2 domains are essential for SOCS-1 and SOCS-3 to inhibit cytokine signaling. Inhibition of cytokine-dependent activation of STAT3 occurred in cells expressing either SOCS-1 or SOCS-3, but unlike SOCS-1, SOCS-3 did not directly interact with or inhibit the activity of JAK kinases. Although the conserved SOCS box motif appeared to be dispensable for SOCS-1 and SOCS-3 action when overexpressed, this domain interacts with elongin proteins and may be important in regulating protein turnover. In gene knockout studies, SOCS-1(-/-) mice were born but failed to thrive and died within 3 weeks of age with fatty degeneration of the liver and hemopoietic infiltration of several organs. The thymus in SOCS 1(-/-) mice was small, the animals were lymphopenic, and deficiencies in B lymphocytes were evident within hemopoietic organs. We propose that the absence of SOCS-1 in these mice prevents lymphocytes and liver cells from appropriately controlling signals from cytokines with cytotoxic side effects. PMID- 10534115 TI - Expression of serum amyloid A3 mRNA by inflammatory macrophages exposed to membrane glycoconjugates from Trypanosoma cruzi. AB - Differential display reverse transcriptase-polymerase chain reaction (PCR) was used to identify genes expressed by murine macrophages exposed to glycosylphosphatidylinositol-anchored mucin-like glycoproteins isolated from Trypanosoma cruzi trypomastigotes. Among the different PCR product bands identified in the differential display gel, one showed high homology with the serum amyloid A3 protein (SAA3). Northern blot assays showed augmentation of SAA3 mRNA expression by inflammatory macrophages exposed to live trypomastigotes or parasite glycolipids, as compared to unstimulated macrophages. Our results also showed the expression of SAA3 mRNA, in liver and heart from animals in the acute phase of Chagas disease. It is important that expression of SAA3 mRNA was closely associated with tissue parasitism and presence of inflammatory cells. Together, our findings indicate the possible involvement of SAA3 protein on immunopathology of Chagas disease and establish a new infectious disease model to study the pathophysiological role of this acute-phase protein. PMID- 10534116 TI - Attenuated liver fibrosis and depressed serum albumin levels in carbon tetrachloride-treated IL-6-deficient mice. AB - Chronic intermittent injection of carbon tetrachloride (CCl4) for more than 10 weeks induced liver fibrosis in mice, as evidenced by positive Azan staining and increased intrahepatic collagen content. Preceding the onset of liver fibrosis, interleukin-6 (IL-6) gene expression was enhanced in liver and immunoreactive IL 6 was detected in infiltrating inflammatory cells. To delineate the role of IL-6 in this process, we treated IL-6-deficient mice with CCl4 in a similar manner for 12 weeks, after which fibrotic changes were less evident and serum albumin levels were lower in IL-6-deficient than wild-type mice. Moreover, CCl4-induced expression of transforming growth factor beta1 and hepatocyte growth factor genes in liver was significantly reduced in IL-6-deficient mice. Thus, IL-6 may be vitally involved in fibrotic changes and maintenance of serum albumin levels, partly by modulating intrahepatic expression of these cytokines. PMID- 10534117 TI - Differentiating agents regulate cathepsin B gene expression in HL-60 cells. AB - We utilized HL-60 cells as a model system to examine the regulation of ctsb gene expression by differentiating agents. Inducers of monocytic differentiation [phorbol ester (PMA), calcitriol (D3), and sodium butyrate (NaB)] and inducers of granulocytic differentiation [all-trans retinoic acid (RA) and 9-cis retinoic acid (9-cis RA)] increase ctsb mRNA levels in a dose-dependent manner as determined by Northern blot hybridization. D3 and retinoids exert additive effects, suggesting that these agents act in part through distinct pathways. Actinomycin D decay experiments indicate that D3, NaB, RA, and 9-cis RA do not alter mRNA stability. In contrast, PMA markedly increases the half-life of ctsb mRNA. In transient transfection assays, PMA and NaB both stimulate transcription of the luciferase reporter gene placed under the control of ctsb promoter fragments. Thus, inducers of HL-60 cell differentiation can regulate the expression of the ctsb gene at both transcriptional and posttranscriptional levels. PMID- 10534118 TI - Polarized expression of immunoglobulin, spectrin, and protein kinase C beta II occurs in B cells from normal BALB/c, autoimmune lpr, and anti-ssDNA transgenic, tolerant mice. AB - The rapid redistribution of B cell surface immunoglobulin to a cap upon cross linking treatment is a well-described phenomenon, the physiological significance of which is unknown. We describe the observation that splenic B cells from unimmunized normal, autoimmune, and tolerant mice express naturally occurring capped immunoglobulin in the absence of exogenous stimulation. The percentage of capped B cells increases to 20% of B cells by age 16 weeks in the progressive autoimmune lpr mouse. Transgenic, tolerant mice expressing lpr-derived genes for ssDNA-binding antibody also demonstrate a large percentage (35-75%) of immunoglobulin-capped splenic B cells. In these capped B cells, protein kinase C beta II, the cytoskeletal proteins spectrin and ankyrin, and the lipophilic probe diI are enriched beneath the site of the immunoglobulin cap. These data suggest that polarization of surface receptors, signaling molecules, anionic phospholipid domains, and cytoskeletal proteins may be an important part of the B cell immune response in vivo. PMID- 10534119 TI - Murine fetal natural killer cells are functionally and structurally distinct from adult natural killer cells. AB - Natural killer (NK) cell phenotype and activity was studied by analyzing uncultured and short-time-cultured murine NK cells from fetal day 17 spleen and thymus. In contrast to NK cells from adult mice, freshly sorted fetal NK cells did not contain NK receptor transcripts for Ly-49A, B, C/I, D, F, G2, or H. The only NK receptor transcripts that could be detected were Ly-49E and CD94. It is important that Ly-49E was present at a 10- to 30-fold higher level compared with uncultured NK cells from adult mice. After short-time interleukin-2 culture, the level of Ly-49E mRNA was comparable between fetal and adult NK cells. Functionally, fetal NK cells only killed MHC class I-negative tumor cells when activating NK receptors were cross-linked with antibody. We show that fetal NK cells are mature but are different from NK cells in adult mice regarding their NK cell receptor repertoire and function. PMID- 10534121 TI - Coexpression of binding sites for A(B) histo-blood group trisaccharides with galectin-3 and Lag antigen in human Langerhans cells. AB - Galectin-3 is an immunomodulatory protein with binding capacity for various glycoconjugates including IgE. It has been shown to be produced by epidermal keratinocytes and is present on the surfaces of skin Langerhans cells (LC). Therefore, it may have a role in the pathogenesis of various skin diseases, such as atopic dermatitis. To study the expression of galectin-3 in LC, we used, in addition to specific antibodies, a panel of synthetic, carrier-immobilized, specific oligosaccharides of the A- and B-histo-blood group, which are recognized by this lectin. In the mean time, Birbeck granules were visualized with an anti Lag antibody. The double labeling experiments showed a remarkable colocalization of signals for Lag antigen (Birbeck granules) and galectin-3, as well as the binding sites for A- and B-histo-blood group trisaccharides. The specificity of the oligosaccharide binding was demonstrated by the lack of binding by Le(c), Le(d) (H blood group antigen), and sLe(x), which are not recognized by galectin 3. These results suggest that galectin-3 is present in Birbeck granules, where it retains reactivity for its glycoligands. PMID- 10534120 TI - Characterization of the biosynthesis, processing, and sorting of human HBP/CAP37/azurocidin. AB - Azurocidin is a multifunctional endotoxin-binding serine protease homolog synthesized during the promyelocytic stage of neutrophil development. To characterize the biosynthesis and processing of azurocidin, cDNA encoding human preproazurocidin was stably transfected to the rat basophilic leukemia cell line RBL-1 and the murine myeloblast-like cell line 32D cl3; cell lines previously utilized to study the related proteins cathepsin G and proteinase 3. After 30 min of pulse radiolabeling, two forms of newly synthesized proazurocidin (34.5 and 37 kDa), differing in carbohydrate content but with protein cores of identical sizes, were recognized. With time, the 34.5-kDa form disappeared, while the 37 kDa form was further processed proteolytically, as judged by digestion with N glycosidase F. Conversion of high-mannose oligosaccharides into complex forms was shown by acquisition of complete resistance to endoglycosidase H. Radiosequence analysis demonstrated that the amino-terminal seven amino acid propeptide of proazurocidin was removed in a stepwise manner during processing; initial removal of five amino acids was followed by cleavage of a dipeptide. Presence of the protease inhibitors Gly-Phe-diazomethyl ketone, bestatin, or leupeptin inhibited only the cleavage of the dipeptide, thus indicating the involvement of at least two amino-terminal processing enzymes. Translocation of azurocidin to granules was shown by subcellular fractionation. Similar results, with efficient biosynthesis, processing, and targeting to granules in both cell lines, were obtained with a mutant form of human preproazurocidin lacking the amino-terminal heptapropeptide. In conclusion, this investigation is an important addition to our previous studies on related azurophil granule proteins, and provides novel information concerning the biosynthesis and distinctive amino-terminal processing of human azurocidin. PMID- 10534122 TI - Morphine promotes apoptosis in Jurkat cells. AB - Patients with intravenous heroin addiction are prone to recurrent infections and at times these infections are fatal. We evaluated the effect of morphine on the apoptosis of Jurkat cells and freshly isolated human T lymphocytes. Morphine promoted apoptosis of both the Jurkat cells and the freshly isolated T lymphocytes in a dose-dependent manner. DAGO, a specific mu receptor agonist, also promoted Jurkat cell apoptosis. DNA isolated from morphine-treated Jurkat cells and T lymphocytes also showed integer multiples of 200 base pairs. Superoxide dismutase (SOD) enhanced lymphocyte apoptosis; whereas catalase attenuated the morphine-induced apoptosis of Jurkat cells as well as of T lymphocytes. Morphine-treated Jurkat cells also showed a decreased expression of bcl-2 and an enhanced expression of bax. In addition, morphine-treated Jurkat cells showed activation of caspase-3. These results indicate that morphine induced T lymphocyte apoptosis may be mediated through the generation of reactive oxygen species. The change in ratio of bax and bcl-2 seems to tilt the balance toward apoptosis, leading to the activation of caspase-3. This study provides further support for the hypothesis that morphine may be directly compromising immune function by enhancing apoptosis of T lymphocytes in patients with heroin addiction. PMID- 10534123 TI - The generation of human dendritic and NK cells from hemopoietic progenitors induced by interleukin-15. AB - Interleukin-15 (IL-15) is a pleiotropic cytokine that induces the generation and differentiation of lymphoid cells and shares many biological activities with IL 2. We have shown here the development of dendritic cells (DC) from human CD34+ hemopoietic precursor cells cultured for 2-4 weeks with IL-15 alone. DC generated with IL-15 have typical morphological, immunocytochemical, phenotypic, and functional characteristics of mature DC. Dual flow cytometry analysis performed weekly demonstrated increasing co-expression of CD1a or CD83 with HLA-DR, CD80, CD86, IL-2R alpha, beta, and gamma. Two populations of cells were distinguished among CD34+ progeny. Small and medium-size cells were mainly natural killer (NK) cells (72.6-85.2% CD56+) and low numbers of DC (9.1-21.3% CD1a+). Large cells were mostly DC (75.4-95.4% CD1a+). Isolated CD34+ cells did not express IL-2R subunits but after 2-3 days in culture with IL-15, they were found to express IL 2Rgamma. Induced expression of IL-2Rgamma on CD34+ cells may explain the primary mechanism of IL-15-regulated differentiation of hemopoietic precursor cells. Thus, our data suggest that IL-15 stimulates CD34+ cells to differentiate into NK and DC and may represent a new growth and survival factor for lymphoid DC. PMID- 10534124 TI - Maturation decreases responsiveness of human bone marrow B lineage cells to stromal-derived factor 1 (SDF-1). AB - We compared the chemotactic responsiveness of different subsets of human B lineage cells to stromal derived factor-1 (SDF-1). High percentages (30-40% of input) of purified bone marrow progenitors including non-B lineage progenitors, pro-B cells, and pre-B cells migrated to SDF-1alpha, demonstrating that SDF-1 is an efficacious chemoattractant of these cells. Pro-B cells responded optimally to a lower concentration of SDF-1 than other subsets, demonstrating that SDF-1 is a more potent chemoattractant of this subset. A lower percentage (10-15% of input) of mature B lymphocytes migrated to SDF-1alpha than pro-B cells, demonstrating that responsiveness of B lineage cells to SDF-1 decreases during differentiation. Inhibition by anti-CXCR4 mAb demonstrated that migration of B lineage cells to SDF-1 was completely dependent on CXC chemokine receptor-4 (CXCR4). Mature B cells expressed higher levels of CXCR4 receptors than uncommitted progenitors and pro-B cells, despite differences in responsiveness to SDF-1. CXCR4 receptors expressed by unresponsive and SDF-1-responsive B cells bound SDF-1alpha with similar affinities (K(D) = 1.7-3.3 x 10(-9) M). Therefore, elements downstream from CXCR4 appear to regulate responsiveness of B cells to SDF-1. We speculate that SDF-1 and CXCR4 direct migration of progenitor cells in microenvironments that promote B lymphopoiesis. PMID- 10534125 TI - MIP-3alpha induces human eosinophil migration and activation of the mitogen activated protein kinases (p42/p44 MAPK). AB - The CC chemokine macrophage inflammatory protein-3alpha (MIP-3alpha) is the product of recent electronic cloning efforts, however, little characterization of its spectrum of biological effects has been undertaken. Human eosinophils exhibited pertussis-toxin-sensitive migration in response to human recombinant (hr)MIP-3alpha. Messenger RNA for the MIP-3alpha receptor, CCR-6, and low levels of surface expression were demonstrated by reverse transcriptase-polymerase chain reaction and FACS analysis. Analyses of cell signaling revealed dose-dependent increases in intracellular calcium mobilization, calcium transients that were, however, greatly reduced when compared with MCP-3-induced responses. Further investigations of MIP-3alpha-induced signal transduction revealed time- and dose dependent, partially pertussis toxin-dependent, increases in phosphorylation of the p42/p44 mitogen-activated protein kinases (MAPK) that occurred at 10- to 100 fold lower concentrations, and that were linked to a phosphoinositide 3-kinase pathway. These results suggest that MIP-3alpha can regulate multiple, parallel signal transduction pathways in eosinophils, and suggest that MAPK activation by MIP-3alpha in eosinophils is a significant signaling pathway for migration induction. PMID- 10534126 TI - C/EBP regulates the promoter of the eosinophil-derived neurotoxin/RNS2 gene in human eosinophilic cells. AB - The eosinophil-derived neurotoxin (EDN), a member of the mammalian ribonuclease family, is found in the large specific granules of human eosinophilic leukocytes. We have investigated the role of the C/EBP transcription factor family in the regulation of EDN promoter activity. Here we show that the C/EBP family is involved in intrinsic regulation of EDN promoter activity. We have identified a C/EBP binding site located at -124 in the proximal promoter of the EDN gene. Mutation of this C/EBP site results in a decrease of promoter activity in HL-60 eos cells as well as in eosinophils differentiated in vitro from CD34+ cells. Different C/EBP proteins are able to bind to the C/EBP site as shown by gel shift assay. Our results indicate the importance of the C/EBP family in the regulation of the EDN gene in eosinophils. PMID- 10534127 TI - LPS-induced signals in activation of caspase-3-like protease, a key enzyme regulating apoptotic cell damage into a macrophage-like cell line, J774.1, in the presence of cycloheximide. AB - The earliest observed apoptotic change in a macrophage-like cell line, J774.1, treated with lipopolysaccharide (LPS) in the presence of cycloheximide (CHX) was a selective increase in caspase-3-like activity. The addition of polymyxin B, TPCK, herbimycin A, or genistein, all of which inhibited LPS-induced tumor necrosis factor alpha (TNF-alpha) production by macrophages, suppressed the activation of the caspase-3-like protease in these macrophages treated simultaneously with CHX. However, SB202190 and SB203580, inhibitors of MAP kinase, and PD98059, an inhibitor of MAP-kinase kinase (MEK), showed no effect on the activation of the caspase-3-like protease or on the cell damage of the macrophages treated with LPS and CHX, whereas they inhibited LPS-induced TNF alpha production. These results suggest that some of the early signals in LPS treated macrophages are common to the subsequent pathways for TNF-alpha production and caspase-3-like protease activation, but the later signals, like MAP-kinase kinase or MAP-kinase, are not involved in the pathways for caspase-3 like protease activation. PMID- 10534128 TI - Smoking and ischaemic heart disease. Overview. PMID- 10534129 TI - Calculation of risk for the development of acute myocardial infarction in the normal population based on long-term follow-up studies: smokers compared with non smokers. AB - The objective was to study the association between smoking habits and mortality from coronary heart disease among men and women aged 35-49 years. Almost 45000 individuals (50% women) from three counties in Norway attended a screening programme during 1974-1978. The participation rate was almost 90% of the population. These individuals have been followed for an average of 18 years through the Norwegian death register. There were 1021 and 193 deaths from coronary heart disease among men and women, respectively. The mortality rate among current smokers was three times higher than that among those who had never smoked cigarettes. The relative risk between low-dose smokers and non-smokers was higher than that between high- and low-dose smokers. Among men, the relative risk between smokers and non-smokers was lower in the age group 45-49 years than in the two younger 5-year age groups. Among women, the relative risks did not vary significantly across the age groups. The relative risk between smokers and non smokers remained the same in the first and second half of the observation period in men. A stronger association in the second half of the observation period was suggested among women (P = 0.08). The duration of smoking was an independent predictor of coronary heart disease among men, but not among women. Duration of smoking and length of follow-up seemed to have a different relationship with mortality from coronary heart disease in men and women. The dose-response relationship was the same in men and women, on the relative scale. Age affected the magnitude of relative risk between smokers and non-smokers among men, but not among women. PMID- 10534130 TI - Is the effect of smoking on the risk for coronary heart disease even stronger than was previously thought? AB - OBJECTIVE: To assess the extent to which smoking cessation among cohort members during the follow-up affects the estimated association between smoking and coronary heart disease (CHD) risk in an observational study with a single determination of the smoking status at baseline. DESIGN: Cross-sectional trend and prospective cohort analyses. SETTINGS: North Karelia and Kuopio provinces in eastern Finland. SUBJECTS: Men (n = 13542) aged 30-59 years who participated in risk factor surveys between 1972 and 1992. Of these, 3937 men belonged to the 1972 cohort with the prospective follow-up. MAIN OUTCOME MEASUREMENTS: Smoking prevalence, CHD mortality, non-CHD mortality, and total mortality. RESULTS: Smoking prevalence in the population decreased from 53% to 37% between 1972 and 1992. In the follow-up of the 1972 study cohort, the association between baseline smoking status and CHD weakened markedly when the follow-up time from the baseline measurement was extended. The risk ratio of cumulative CHD mortality associated with smoking was 6.96 in the first 2-year follow-up and it decreased gradually to 2.06 at the 20-year follow-up. A similar decrease in the risk ratio was observed when the analysis was performed during periods of the follow-up. The association between smoking and non-CHD mortality weakened only slightly when the follow-up time became longer. CONCLUSION: The results from prospective observational studies conducted in populations with a decreasing smoking prevalence may be biased by the misclassification of study subjects during the follow-up as a result of smoking cessation. This results in an underestimation of the risk of CHD caused by smoking. Thus, smoking may be even more harmful for health than the estimates from prospective studies suggest. PMID- 10534131 TI - Pathology of the coronary arteries in smokers and non-smokers. AB - Cigarette smoking is known to be a strong risk factor for several cardiovascular diseases such as ischaemic heart disease, stroke, intermittent claudication and aortic aneurysm. Atherosclerosis, often with superimposed thrombosis, has been shown to be the underlying disease process in all of these diseases. This fact has led to the assumption that smoking accelerates the atherosclerotic process and thereby promotes premature cardiovascular disease. However, increased occurrence of smoking-mediated thrombosis might also play a causative role. This review therefore considers clinical and experimental evidence regarding the atherogenicity and thrombogenicity of cigarette smoking in the development and occurrence of cardiovascular disease. On the basis of the currently available literature, it is concluded that evidence for the atherogenetic effect of smoking is scarce, and that the effect on the athrosclerotic process in the coronary arteries is only moderate if present at all. On the other hand, both clinical and experimental data strongly support the notion that thrombogenetic factors are responsible for the increased occurrence of ischaemic heart disease, and particularly acute coronary syndromes, among chronic smokers. PMID- 10534132 TI - The effect of cigarette smoking and nicotine chewing gum on platelet function and fibrinolytic activity. PMID- 10534133 TI - Age-distribution, risk factors and mortality in smokers and non-smokers with acute myocardial infarction: a review. TRACE study group. Danish Trandolapril Cardiac Evaluation. AB - Smoking is a risk factor for acute myocardial infarction; paradoxically, many studies have shown a lower post-infarct mortality among smokers. There are some important differences between smokers and non-smokers, which might explain the observed difference in mortality: smokers have less multivessel disease and atherosclerosis but are more thrombogenic; thrombolytic therapy seems to be more effective among smokers; smoking might result in an increased out-of-hospital mortality rate, by being more arrhythmogenic; and smokers are on average a decade younger than non-smokers at the time of infarction, and have less concomitant disease. Adjusting for these differences in regression analyses shows that smoking is not an independent risk factor for mortality after acute myocardial infarction. The difference in age and risk factors are responsible for the lower mortality among smokers. PMID- 10534134 TI - Low birth weight and risk of hypertension in African school children. AB - BACKGROUND: In accordance with Baker's programming hypothesis, many studies have demonstrated a relationship between low birth weight (LBW) and high risk of hypertension in adulthood. The present study examines a possible association between LBW and the risk of a child having hypertension later in life. METHODS: The study was a cross-sectional, semi-urban survey. Information on the perinatal characteristics of 2648 randomly sampled school children was collected retrospectively in Kinshasa town, Democratic Republic of Congo. RESULTS: High risk of hypertension in these African school children was related to LBW (<2.500 g); the odds ratio was 2 (95% confidence interval 0.9-8.2, P<0.01) and 2.3 (95% confidence interval 0.6-11.5, P<0.01) for systolic and diastolic hypertension respectively. Birth weight was inversely related with both blood pressure and heart rate; the strongest association was shown in females and adolescents. CONCLUSIONS: Antenatal stress leading to LBW may be associated with programming induced by foetal undernutrition, which in turn leads to the emergence of cardiovascular disease and increased risk of hypertension. PMID- 10534135 TI - Collaborative overview ('meta-analysis') of prospective observational studies of the associations of usual blood pressure and usual cholesterol levels with common causes of death: protocol for the second cycle of the Prospective Studies Collaboration. AB - BACKGROUND: Many prospective studies have been reported on the associations of blood pressure and blood cholesterol with cardiovascular disease, but few have been large enough to provide reliable estimates of the nature of these associations in different circumstances or to characterize the associations with non-vascular causes of death. Moreover, almost all such reports have related risk to baseline measurements of the risk factor of interest, which can lead to substantial underestimation of the importance of the risk factor due to 'regression dilution' bias. OBJECTIVE: By appropriate combination of data on individual participants from all such studies in a systematic 'meta-analysis', with correction for regression dilution, to characterize more precisely than has previously been possible the age-specific and sex-specific relevance of blood pressure and blood cholesterol levels to particular causes of death. METHODS: Information has been made available on about 175000 deaths (45% due to vascular disease and 20% due to cancer) among more than 1.4 million individuals followed for an average of 14 years in over 60 prospective studies of blood pressure and blood cholesterol (with data on high-density lipoprotein cholesterol available for a substantial subset of individuals). This represents approximately 95% of the data from all relevant studies. PMID- 10534136 TI - Upper-body adiposity and risk of myocardial infarction. AB - BACKGROUND: The relation between obesity and coronary heart disease (CHD) has long been studied, but no convincing conclusion has been drawn. OBJECTIVE: To estimate the relative risk associated with upper-body adiposity which is at present believed to be a better predictor of CHD. DESIGN: This was a community based case-control study. METHODS: We studied 338 consecutively admitted patients who had had their first acute myocardial infarction (AMI) and 662 community controls who had not suffered AMI selected as a random sample of adults living in the catchment area of the hospital. We defined three classes of body mass index (BMI) and waist: hip circumference ratio on the basis of tertiles of distribution for controls. Odds ratios (OR) were estimated using unconditional logistic regression. Separate models were built for men and women. RESULTS: In univariate analysis we found a higher risk of AMI for men and women with hypertension, dyslipidaemia, diabetes and lower levels of education, for older women, for male smokers and for men with family histories of CHD. Both for men and for women a higher BMI was associated with a slightly higher risk, whereas the adjusted risk of AMI increased with increasing waist: hip circumference ratio [for men OR (second tertile)= 2.5, 95% confidence interval (CI) 1.3-4.9 and OR (third tertile)= 11.1, 95% CI 6.0-20.6; for women OR (second tertile) = 3.0, 95% CI 0.6 14.8 and OR (third tertile) = 14.1,95% CI 3.2-62.7]. This relation held for each BMI class and was stronger for classes of lower BMI. CONCLUSIONS: Distribution of body fat rather than BMI is a strong marker of risk for AMI and there is a clear interaction between these two variables. PMID- 10534137 TI - Low free-thyroxine levels are a risk factor for subclinical atherosclerosis in euthyroid hyperlipidemic patients. AB - BACKGROUND: Patients displaying overt and subclinical hypothyroidism have more cardiovascular risk factors. Consequently, they are more likely to develop atherosclerosis and cardiovascular diseases. OBJECTIVE: To analyze whether low free-thyroxine levels (FTL) would also be associated with atherosclerosis in euthyroid patients. METHODS: We selected a group of 1434 healthy euthyroid male patients without known histories of thyroid disease and with levels of thyroid stimulating hormone values within the normal range (mean 1.70+/-0.76 mUl/l, range 0.13-4.01 mUl/l). Mean age of these patients who had been referred for assessment of hyperlipidemia was 44.6 years and mean FTL was 14.25+/-3.06 pmol/l. We divided the population according to the degree of atherosclerosis in the carotid arteries. RESULTS: Mean age, body mass index, systolic blood pressure, cigarettes/day, blood level of glucose, cholesterol levels, and fibrinogen levels were significantly higher for the patients with atherosclerotic lesions whereas mean FTL was lower for patients with carotid atherosclerosis (P = 0.0002). The relationship between FTL and carotid atherosclerosis was independent from the following cardiovascular risk factors: age, hypertension, amount of excess weight, cholesterol level, fibrinogen level, smoking status, and presence versus absence of diabetes mellitus. CONCLUSIONS: Low FTL is a risk factor for atherosclerosis in male euthyroid hyperlipidemic patients. PMID- 10534138 TI - The role of cigarette smoking in atherosclerotic disease: an epidemiologic analysis. AB - The Bowling Green Study of the Primary and Secondary Prevention of Atherosclerotic Disease has accumulated an age-sex register of 668 consecutive patients who developed some form of atherosclerotic disease between 4 November 1974 and 1 January 1997. Observational data relating to levels of lipids and glucose, blood pressure, body-mass index, and cigarette-smoking status are included in the age-sex register. Analysis of this database clearly shows that cigarette smoking is the main cause of atherosclerotic disease, including events in each of the vascular trees, as well as multiple-system disease and death, at ages much earlier than the ages at which similar atherosclerotic events occur in ex-smokers. Ex-smokers, in turn, suffer corresponding events at earlier ages than do never-smokers. Cigarette smoking produces atherosclerotic events at roughly the same age irrespective of status of other risk factors and is therefore the single most important risk factor for atherosclerosis. PMID- 10534139 TI - A review of metabolic and cardiovascular effects of oral antidiabetic agents: beyond glucose-level lowering. AB - It has become evident that cardiovascular disease is the major cause of morbidity and mortality in type 2 diabetes mellitus. This raises the possibility that glucose lowering agents with other nonglucose-lowering effects, might have added benefits. In this review, we focus on the metabolic and cardiovascular effects of oral antidiabetic agents that go beyond glucose-level lowering. Such effects include lipid modifying actions, antithrombotic and profibrinolytic activities, and direct action at the level of the vessel wall to improve endothelial function or prevent smooth muscle hyperplasia. These additional activities, particularly those seen with the newer oral antidiabetic agents, hold the promise of reducing cardiovascular complications beyond that achievable by glucose lowering alone. PMID- 10534140 TI - Bibliography. Current world literature. PMID- 10534141 TI - Nitric oxide modulates HIV-1 replication. AB - Although nitric oxide (NO) production is increased in HIV-1-infected patients, and NO is known to inhibit the replication of several viruses, very little is known about the effects of NO on HIV-1 replication. In the present studies, we find that S-nitrosothiols (RSNOs), a class of NO donor compounds present in the human circulatory system, inhibit HIV-1 replication in acutely infected human peripheral blood mononuclear cells (PBMCs) and have an additive inhibitory effect on HIV-1 replication in combination with 3'-azido-3'-deoxythymidylate (AZT). RSNOs inhibit HIV-1 replication in acutely infected PBMCs at a step in the viral replicative cycle after reverse transcription, but before or during viral protein expression through a cGMP-independent mechanism. In the latently infected U1 cell line, NO donor compounds and intracellular NO production stimulate HIV-1 reactivation. These studies suggest that NO both inhibits HIV-1 replication in acutely infected cells and stimulates HIV-1 reactivation in chronically infected cells. Thus, NO may have a physiologic role in HIV-1 replication, and NO donor compounds, which have been used for decades in the treatment of coronary artery disease with limited toxicity, might be useful in the treatment of HIV-1 disease by inhibiting acute infection, reactivating latent virus, or both. PMID- 10534142 TI - Frontal lobe neuronal injury correlates to altered function in FIV-infected cats. AB - Six cats infected intravenously at 8 weeks of age with feline immunodeficiency virus Maryland isolate (FIV-MD), were evaluated at 8 and 14 months of age (6 months and 12 months postinfection, respectively) with high spatial resolution proton magnetic resonance spectroscopy (MRS) of the frontal cortex. Two separate control cat groups were evaluated at 8 months and 16 months of age. Single voxel two-dimensional high-resolution proton magnetic resonance imaging was performed using the PRESS sequence by selecting a 0.125 ml volume of interest in the medial frontal cortex. A significant reduction in both N-acetylaspartate (NAA) and NAA: choline ratio was found in the FIV 14-month-old group compared with FIV 8-month old cats, and to the respective age-matched control 16-month-old cats. A negative correlation between NAA and CD4 lymphocyte count was seen in the FIV-14 group only. This group of FIV cats also exhibited a higher proportion of quantitative electroencephalographic relative slow wave activity (RSWA) that correlated to lower NAA content in the frontal cortical voxel. Although peripheral blood proviral load increased over time of infection, no correlation was found between proviral blood or lymph node load and NAA values, CD4 lymphocyte counts, or frontal cortical RSWA. Thus, this study demonstrated that neurologic functional disruption of the frontal cortex correlated strongly with neuronal injury and/or loss in FIV-MD-infected cats independent of peripheral proviral load. The ability to define in vivo neurodegeneration further in this animal model helps in understanding the neuropathogenesis of lentivirus infection, and possibly, a means to follow progression and reversibility during the initial stages of brain infection as therapeutic agents are identified. PMID- 10534143 TI - HIV-1-specific cytolytic T-lymphocyte activity correlates with lower viral load, higher CD4 count, and CD8+CD38-DR- phenotype: comparison of statistical methods for measurement. AB - OBJECTIVES: The objective of this study was to use novel statistical methods to determine the correlation between HIV-1-specific cytolytic T-lymphocyte (CTL) activity and HIV-1 plasma viral load, in a blinded study of HIV-infected patients at various stages of clinical disease. METHODS: Peripheral blood mononuclear cells (PBMC) were collected and stored at enrollment and 2 weeks later, from 15 HIV-infected individuals who were receiving stable antiretroviral therapy for the previous 6 weeks and during the study period. HIV-1-specific CTL activity was measured using an antigen-specific PBMC in vitro stimulation method. Measurements of plasma viral load, as well as CD4+ and CD8+ T lymphocytes expressing T-cell activation markers (DR and CD38) were also performed at each time point. CTL activity was quantified using three separate statistical methods: area under the net HIV-specific lysis curve (AUC), lytic units (LU20), and linear regression (LR) of net HIV-specific lysis. RESULTS: HIV-1 nef-, pol- and gag-specific CTL activity (AUC method) was significantly higher in subjects with a plasma viral load < or = 30,000 RNA copies/ml, than in those with viral load >30,000 RNA copies/ml. When plasma viral load was analyzed as a continuous variable, there was a strong correlation between higher CTL activity and lower viral load for nef (r2 = .77; p < .001), pol (r2 = .63; p < .001) and gag (r2 = 0.75; p < .001) targets by the AUC, but not for the LU20 analysis. Using the LR analysis, which is less dependent on in vitro PBMC growth than the AUC analysis, an independent association was demonstrated between nef- and gag-specific CTL activity and lower viral load. Measurement of CTL activity was also significantly correlated with a higher percentage of circulating CD8+DR-CD38- T lymphocytes. CONCLUSIONS: In this blinded study using an in vitro stimulation of frozen PBMC, higher HIV-1 nef-, pol-, and gag-specific CTL activity correlated with lower plasma viral load, particularly in patients with a CD4 count <500 cells/mm3. Two new statistical methods for estimating CTL activity, AUC and LR analyses, were superior to the standard lytic unit (LU20) method for demonstrating this correlation. These data also demonstrated that higher circulating CD8+ T lymphocytes with a DR-CD38 phenotype, correlate with a lower plasma viral and load and higher HIV-specific CTL activity. This suggests that lymphocytes with this double-negative phenotype may include circulating HIV-specific CD8+ CTL. PMID- 10534144 TI - Acute activation of CD8+ T lymphocytes in interleukin-2-treated HIV-infected patients. ANRS-048 IL-2 Study Group. Agence Nationale de Recherches sur le SIDA. AB - CD8+ T lymphocytes play a key role in the control of HIV infection, through both cytotoxic and noncytotoxic mechanisms. To study in vivo effects of interleukin-2 (IL-2) treatment on this cell compartment, the level of activation of CD8+ T lymphocytes was evaluated before and just after 5-day administration of IL-2 in 16 HIV-infected patients. The serum level of soluble CD25 and of soluble CD8 significantly increased following IL-2 administration. The number of mRNA molecules coding for perforin and granzyme B, two enzymes that are contained in granules of cytotoxic cells, also significantly increased in peripheral blood mononuclear cells and in purified CD8+ cells (p < .001). Variations of plasma HIV viremia and perforin gene expression following IL-2 administration were inversely correlated (p = .023), suggesting that IL-2-induced activation of CD8+ T lymphocytes contributes to limit HIV replication in vivo. In contrast to perforin and granzyme B gene expression, IL-2 administration did not increase the expression of macrophage inhibitory protein-1alpha (MIP-1alpha), MIP-1beta, and regulated-on-activation normal T-expressed and secreted (RANTES) genes. These findings indicate that CD8+ T lymphocytes in HIV-infected patients are acutely activated by IL-2 treatment, which may improve long-term control of HIV infection. PMID- 10534145 TI - Safety and antiretroviral effects of combined didanosine and stavudine therapy in HIV-infected individuals with CD4 counts of 200 to 500 cells/mm3. AB - The safety and antiretroviral effects of didanosine and stavudine in combination were evaluated in 86 people infected with HIV with CD4 counts between 200 and 500 cells/mm3 who had received <7 days of prior nucleoside analogue antiretroviral treatment. Patients were randomized to receive blinded treatments with one of five weight-adjusted, twice-daily regimens of didanosine and stavudine (100 + 10 mg, 100 + 20 mg, 100 + 40 mg, 200 + 20 mg, and 200 + 40 mg) for up to 1 year. Dosages were adjusted appropriately for patients weighing <60 kg and reduced in response to adverse effects. No clear dose-related differences among treatment groups were detected with regard to suppression of plasma HIV RNA level or reduction in infectious titers in peripheral blood mononuclear cells (PBMCs), improvement in CD4 count, or adverse effects. However, trends toward greater decreases in viral load and increases in CD4 count were detected when treatment groups containing the full recommended dosage of one or both agents (high-dose subgroup; arms 3, 4, and 5) were compared with the groups receiving lower dosages. At 28 weeks the mean log 10 HIV RNA decrease was 1.12 (n = 52) and at 52 weeks it was 0.97 (n = 32). Combination therapy was well tolerated, with no apparent dose-related adverse effects. Peripheral neuropathy occurred in 2 of 86 (2.3%) of patients. Didanosine and stavudine together appear to be a good nucleoside analogue foundation for aggressive triple- or quadruple-drug therapy. Full therapeutic doses of each of these two agents should be used to achieve optimal suppression of HIV replication. PMID- 10534146 TI - Insulin-like growth factor system in patients with HIV infection: effect of exogenous growth hormone administration. AB - The purpose of this study was to characterize changes in the levels of insulin like growth factor-I (IGF-I) and IGF binding proteins (BP) 1, 2, and 3 in HIV infected adults throughout the course of their disease, and to assess the responsiveness of the IGF system components to growth hormone (GH) administration (6 mg/day) for 2 weeks. Healthy control study subjects (n = 10) were compared with patients who were either HIV-positive (n = 9), had AIDS without weight loss (n = 13), or had AIDS with >10% weight loss (n = 6), all of whom had been free of acute illness for at least 3 months. Under basal conditions, fasting serum concentrations of epinephrine, norepinephrine, cortisol, glucagon, insulin, IGF I, and IGFBP-3 were not significantly different among the four groups. The serum concentrations of IGFBP-1 and IGFBP-2 were significantly higher in AIDS patients with wasting than in the other three groups (p < .05). In addition, there was a statistically significant positive correlation between the levels of IGFBP- 1 (p = .004) and IGFBP-2 (p = .03) and the stage of disease. Following GH administration, the serum concentrations of insulin and IGF-I were increased in all groups (p < .05). In addition, the increases in insulin levels correlated with stage of disease (p = .004). The responses of the IGFBPs were more variable. GH administration significantly increased the levels of IGFBP-3 in all groups except the patients with AIDS wasting, whereas the levels of IGFBP-1 were significantly decreased in controls and AIDS patients. These results demonstrate that there is a continuum of both elevations in the IGFBPs and altered metabolic responsiveness in patients infected with HIV that increases with the severity of the disease. These data also demonstrate that AIDS patients, who are free from secondary infection, respond to administration of GH by significantly increasing hepatic IGF-I production. PMID- 10534147 TI - Randomized, placebo-controlled trial of Chinese herb therapy for HIV-1-infected individuals. AB - CONTEXT: Alternative medicine or complementary remedies that have not been scientifically tested are nonetheless widely used to treat chronic illnesses, particularly if curative options are limited. OBJECTIVES: To assess the effectiveness of Chinese medicinal herbs in reducing symptoms and improving the quality of life of HIV-infected persons. DESIGN: Prospective, placebo-controlled double-blind study. SETTING: University-based HIV outpatient clinic. PATIENTS: 68 HIV-infected adults with CD4 cell counts <0.5 x 10(9)/L. INTERVENTION: Participants were randomized to receive four daily doses of seven pills containing a standardized preparation of 35 Chinese herbs or placebo for 6 months. MAIN OUTCOME MEASURES: Symptoms, HIV disease progression, HIV-1 RNA plasma viral loads, CD4 and CD8 cell counts, and scores on standard questionnaires for quality of life, depression, anxiety, and coping. RESULTS: Intervention and placebo groups were equivalent at baseline regarding, respectively, previous antiretroviral therapy (74% versus 79%), median CD4 cell counts (0.20 x 10(9)/L versus 0.25 x 10(9)/L), and median HIV-1 plasma viral loads (35,612 copies/ml versus 52,027 copies/ml). At enrollment, none of the study subjects was seriously ill or depressed, and average coping and quality of life scores were in the normal range. In all, 53 (78%) participants completed the study. Patients taking Chinese herbs reported significantly more gastrointestinal disturbances (79% versus 38%; p = .003) than those receiving placebo. No therapy related toxicities were observed. At completion of the study, no significant differences between the intervention and placebo groups were found regarding plasma viral loads, CD4 cell counts, symptoms, and psychometric parameters. HIV-1 RNA level was unchanged at study end. Among participants who were not on concomitant antiretroviral therapy, median CD4 cell counts declined by 0.05 x 10(9)/L in both the intervention and placebo groups. CONCLUSIONS: This standardized formulation of Chinese herbs for HIV-infected individuals did not improve quality of life, clinical manifestations, plasma virus loads, or CD4 cell counts. The data suggest that this formulation of Chinese herbs is not effective when administered in a Western medicine setting. PMID- 10534148 TI - Temporal changes in the rate of progression to death among Italians with known date of HIV seroconversion: estimates of the population effect of treatment. Italian HIV Seroconversion Study (ISS). AB - OBJECTIVE: To evaluate changes in survival among HIV-positive individuals with known date of seroconversion (SC). DESIGN: Prospective cohort study. METHODS: Follow-up lasted from SC to death or to the end of 1997. A multivariate Cox model was applied to estimate relative hazards (RH) of death. The year of SC (as a categoric fixed variable) and calendar year (as a time-dependent variable) were considered to evaluate, respectively, cohort and prevalent changes in the rate of death. A separate Cox model was used to assess the association between survival and new combination therapies, using an "intention to treat" approach. RESULTS: The study included 1535 individuals (53.9% injecting drug users, 25.3% homosexuals, 19.5% heterosexuals); 75.8% seroconverted between 1980 and 1991, and 24.2% seroconverted between 1992 and 1997. When adjusting for year of SC, the RH of death (and that of AIDS) was significantly lower in 1997, compared with before 1991 (RH = 0.54; 95% confidence interval, 0.30-0.98). Adjusted RHs of death were significantly lower for combination antiretroviral therapy, compared with no therapy. When combining the two Cox models, the 1997 reduction in risk of death was largely due to antiretroviral therapies; similar results were obtained when the endpoint was AIDS. CONCLUSIONS: A reduction in the risk of death, probably due to combination antiretroviral therapy, was observed in 1997 after having adjusted for age at SC and year of SC. PMID- 10534149 TI - Unreported AIDS-defining opportunistic illnesses in persons reported with HIV related severe immunosuppression. AB - To better estimate the distribution of AIDS cases after the 1993 change in the case definition, we assessed the proportion of persons whose AIDS diagnosis was based on laboratory criteria for severe immunosuppression (CD4 count <200 cells/microl or <14%) and who also had an unreported opportunistic illness (OI) at the time of the CD4 report. Five U.S. reporting sites (Arizona; Los Angeles County, California; New Jersey; Oregon; and Washington State) reviewed AIDS cases reported between January 1 and June 30, 1993. From these sites, 3289 immunologic cases were reported; of these cases, 322 (9.8%; range, 1.6%-16.1%) were in persons who had an unreported OI. More of those who had an unreported OI were male, members of racial groups other than white, injection drug users, and had a CD4 count of <50 cells/microl at AIDS diagnosis. Because of recent advances in OI prophylaxis and treatment of HIV infection, studies monitoring HIV-related morbidity should assess the occurrence of OIs in a sample of persons reported with HIV and severe immunosuppression. Such assessment will ensure representative ascertainment of initial AIDS-defining OIs and thus improve the usefulness of the data for public health planning and the allocation of resources for patient care. PMID- 10534150 TI - Phenotypic expressions of CCR5-delta32/delta32 homozygosity. AB - OBJECTIVE: As blockade of CC-chemokine receptor 5 (CCR5) has been proposed as therapy for HIV-1, we examined whether the CCR5-delta32/delta32 homozygous genotype has phenotypic expressions other than those related to HIV-1. DESIGN: Study subjects were white homosexual men or men with hemophilia who were not infected with HIV-1. In this study, 15 CCR5-delta32/delta32 homozygotes were compared with 201 CCR5 wild-type (+/+) subjects for a wide range of clinical conditions and laboratory assay results ascertained during prospective cohort studies and routine clinical care. CCR5-delta32 genotype was determined by polymerase chain reaction, followed by single-stranded conformational polymorphism analysis. RESULTS: Hypertension and conditions attributable to hemophilia were the only diagnoses frequently found in clinical records of CCR5 delta32/delta32 study subjects. Based on blood pressure measurement and treatment history, CCR5-delta32/delta32 homozygotes had a 2.8-fold higher prevalence of hypertension than age-matched CCR5-+/+ study subjects (95% confidence interval [CI], 1.2-6.4; p = .01); none of the homozygotes had severe hypertension. Hematologic measures were generally similar across the genotypes, but total lymphocyte counts were approximately 20% higher in CCR5-delta32/delta32 study subjects than in CCR5-+/+ study subjects (p < .05). Among patients with hemophilia who were infected with hepatitis C virus (HCV), mean alanine aminotransferase levels were 117% higher among CCR5-delta32/delta32 homozygotes (p < .05), but serum HCV levels did not differ by CCR5-delta32 genotype. CCR5 delta32/delta32 homozygous study subjects had a lower prevalence of antibodies to measles virus than those with other genotypes, but this association was not confirmed in a group of blood donors. The prevalence of antibodies to nine other common viruses, HBV, and HCV was not related to CCR5 genotype. CONCLUSIONS: CCR5 delta32/delta32 homozygotes are generally similar to wild-type persons. Confirmatory investigations are required to determine whether hypertension, increased lymphocyte counts, and higher hepatic enzyme levels in the presence of HCV infection represent true phenotypic expressions of this genotype. CCR5 delta32/delta32 homozygosity does not provide broad protection against viral infections. PMID- 10534151 TI - Similarities and differences by race/ethnicity in changes of HIV seroprevalence and related behaviors among drug injectors in New York City, 1991-1996. AB - OBJECTIVE: To measure differences and similarities in the prevalence of HIV infection and of related risk and protective behaviors among New York City black, white, and Hispanic drug injectors during a period of decreasing HIV prevalence. METHODS: Drug injectors were interviewed at a drug detoxification clinic and a research storefront in New York City from 1990 to 1996. All subjects had injected drugs within the last six months. Phlebotomy for HIV testing was conducted after pretest counseling. Analysis compares the first half (period) of this recruitment interval with the second half. RESULTS: HIV seroprevalence declined among each racial/ethnic group. In each period, white drug injectors were significantly less likely to be infected than either blacks or Hispanics. Similar declines were found in separate analyses by gender, length of time since first injection, and by recruitment site. After adjustment for changes in sample composition over time, blacks and Hispanics remained significantly more likely to be infected than whites. Interactions indicate that the decline may be greatest among Hispanics and slowest among blacks. A wide variety of risk behaviors declined in each racial/ethnic group; and syringe exchange use increased in each group. Few respondents reported injecting with members of a different racial group at their last injection event. CONCLUSIONS: HIV prevalence and risk behaviors seem to be falling among each racial/ethnic group of drug injectors. Black and Hispanic injectors continue to be more likely to be infected. Declining prevalence among whites poses some risk of politically based decisions to reduce prevention efforts. Overall, these results show that risk reduction can be successful among all racial/ethnic groups of drug injectors and suggest that continued risk reduction programs may be able to attain further declines in infection rates in each group. PMID- 10534152 TI - Prospective study of HTLV-I infection in an initially asymptomatic cohort. AB - A prospective clinical study of 20 initially asymptomatic HTLV-I-seropositive carriers was commenced in 1991 to determine the natural history of the infection in relation to HTLV-I proviral load, immune responses, and lymphocyte phenotype. Proviral load varied widely between carriers but was relatively constant within an individual over time. The lymphocyte phenotype and prevalence of activated lymphocytes were not predictive of disease and the magnitude of the cytotoxic T lymphocyte response to HTLV-I was independent of proviral load. Incident conditions, some related to HTLV-I infection, including a case of HTLV-I associated myelopathy (HAM), were documented in 9 carriers. Development of myelopathy and uveitis was associated with high peripheral blood HTLV-I proviral load that predated symptoms. Persistently high proviral load appears to predate the development of HTLV-I-associated inflammation in neuro-ophthalmic tissue. PMID- 10534153 TI - Evidence of stavudine-related phenotypic resistance among zidovudine-pretreated HIV-1-infected subjects receiving a therapeutic regimen of stavudine plus lamivudine. PMID- 10534154 TI - Once-daily administration of didanosine in combination with stavudine in antiretroviral-naive patients. The STADI Group. PMID- 10534155 TI - Oral thrush and HIV protease inhibitors. PMID- 10534156 TI - Apolipoprotein E and the neuropathology of Alzheimer's disease. PMID- 10534157 TI - Expression of insulin-like growth factor-1 receptor in human colorectal cancer. AB - The activation of the insulinlike growth factor 1/IGF-1 receptor system (IGF1/IGF1-R) has recently emerged as critical event in transformation and tumorigenicity of several murine and human tumors. Expression of IGF1 and of IGF1 R has been demonstrated in normal and neoplastic intestinal cell lines of rats and humans. However, the modulation of IGF1-R expression during the progression from normal colonic mucosa to adenoma, to carcinoma, and to metastasis, has not been evaluated. In this retrospective study, we investigated the expression of IGF1-R in 12 colonic adenomas (AD), 36 primary colorectal adenocarcinomas (CA), and in 27 corresponding metastases (MT). Normal colonic mucosa (N) was adjacent to the CA in 34 cases. Formalin-fixed, paraffin-embedded tissues of each case were immunostained using the avidin-biotin-peroxidase method. We used an anti IGF1-R rabbit polyclonal antibody (Santa Cruz Biotechnology, CA; dilution 1:100). Positive staining was quantitated by CAS-200. Moderate to strong cytoplasmic immunostaining was observed in 34 of 36 CA (96%), and in 25 of 27 MT (93%). In all of the positive MTs, the intensity of the staining was always strong. In 10 of 12 ADs (83%), only a faint cytoplasmic stain was identified. Normal mucosa when present was negative. Strong IGF1-R positivity correlated with higher grade and higher-stage tumors (P < .01). These data suggest a role of IGF1-R expression during the progression of colorectal adenoma to carcinoma. An increased number of IGF1-R receptors may favor the metastasis of colorectal cancer. PMID- 10534158 TI - Myoepithelial differentiation in high-grade invasive ductal carcinomas with large central acellular zones. AB - We hypothesized that invasive ductal carcinomas (IDCs) with large central acellular zones comprising necrosis, tissue infarction, collagen, and hyaline material on their cut surfaces are formed in association with myoepithelial differentiation of the carcinoma cells. To verify this, the expression of S100 protein, alpha-smooth muscle actin (alpha-SMA), and glial fibrillary acidic protein (GFAP) and keratin 14, which has been shown to represent the myoepithelial immunophenotype, was examined immunohistochemically in 18 IDCs with such central zones covering more than 30% of each tumor area, 18 IDCs without such areas as negative controls, and 10 metaplastic carcinomas as positive controls for myoepithelial differentiation. Expression of S100, detected with a polyclonal antibody, S100-alpha, S100-beta, alpha-SMA, GFAP, and keratin 14, was observed in 61%, 83%, 39%, 33%, 28%, and 39% of the IDCs with large central acellular zones, 17%, 44%, 6%, 6%, 0%, and 6% of the IDCs without such zones, and 80%, 70%, 50%, 100%, 80%, and 50% of the metaplastic carcinomas, respectively. We concluded that IDCs with large central acellular zones frequently contain carcinomas showing myoepithelial differentiation. Such histological and immunohistochemical features in IDCs would be expected to be clinicopathologically significant. PMID- 10534159 TI - Clinical relevance of the histological classification of sinonasal intestinal type adenocarcinomas. AB - To verify the prognostic value of the histological typing of intestinal-type adenocarcinoma (ITAC) of the sinonasal tract, 41 such lesions were separately classified by 2 pathologists both according to the World Health Organization (WHO) criteria for the histological typing of large intestine adenocarcinomas (well, moderately, poorly differentiated adenocarcinomas, and mucinous adenocarcinomas) and to the criteria of Kleinsasser and Schroeder (papillary tubular cylinder cell type, alveolar-goblet cell type, signet-ring cell type, and transitional type). Using the WHO classification, 7 adenocarcinomas (17.1%) were well differentiated, 15 (36.6%) were moderately differentiated, and 6 (14.6%) were poorly differentiated; 13 cases (31.7%) were classified as mucinous adenocarcinoma. Following the Kleinsasser and Schroeder classification, 28 adenocarcinomas were of the papillary-tubular cylinder cell type (PTCC), 6 tumors (14.6%) were of the alveolar-goblet cell type (AGC), 1 (2.4%) was of the signet ring cell (SRC) type, and 6 (14.6%) were of the transitional (TR) type. The interrater agreement in subtyping for both histological classifications was 92.6% (kappa, 0.89; P < .001). Kaplan-Meier analysis of cases stratified according to WHO classification showed that patients with mucinous and poorly differentiated adenocarcinomas had a significantly shorter disease-free interval and survival rate than patients with well and moderately differentiated adenocarcinomas (P = .02 and P < .001, respectively; log-rank test). Separate evaluation of survival for patients with AGC and TR adenocarcinomas did not show any statistically significant difference (P = .5). Clinical stage was advanced in most cases (92.6% of patients had T3 or T4 carcinomas) and had no prognostic relevance in the current series, as did treatment and occupational exposure. We conclude that the histological typing of ITACs is highly reproducible and appears to be related to the clinical outcome of the tumors. Adenocarcinomas with a prominent (>50%) mucinous component tend to have a similar clinical behavior, irrespective of their cyto-architectural features and the presence of an associated tubulopapillary component. Therefore, the separation into alveolar-goblet, signet ring, and transitional forms has no prognostic impact. PMID- 10534160 TI - Viral genotypes and p53 expression in Epstein-Barr virus-associated primary malignant lymphomas of the intestines. AB - A small number (4% to 6%) of primary malignant lymphomas arising in the intestines express the EBV genome. However, in these tumors, the viral genotype and the role of tumor suppressor gene p53 have not been investigated. We sought to determine what genotype of EBV is frequently involved and whether the expression of p53 is related to these tumors. We used EBER-1 in situ hybridization and polymerase chain reactions (PCRs) for EBNA-1, EBNA-2A, and EBNA 2B to detect latent infection with EBV and to determine the genotype, respectively. In addition, we performed p53 PCR-SSCP (exons 5 through 9) and immunohistochemical analysis for p53. We found that EBV type B was present in 4 of 6 cases (67%); the genotype of the remaining cases could not be determined. The p53 PCR-SSCP indicated normal migration patterns in all malignant lymphomas, despite the fact that the tumor cells were strongly immunostained for p53 protein in 5 of the 6 cases. Thus, our study demonstrates that EBV-associated non Hodgkin's lymphomas of the intestines in Korea are strongly related to the type B EBV and not to mutations of p53 gene. We suggest that EBV-associated intestinal lymphomas may arise through an interaction between the latent proteins of EBV and the wild-type p53 protein. PMID- 10534161 TI - Splenic marginal zone lymphoma with increased number of blasts: an aggressive variant? AB - Splenic marginal zone lymphoma (SMZL) is a recently described and distinctive type of splenic lymphoma and is characterized by an indolent clinical course. By analyzing a large series of SMZL cases, we recognized the existence of a subset of 6 cases characterized by an aggressive clinical course that led to death caused by the tumor in 5 of 6 cases, whereas the remaining patient showed signs of tumor progression. The morphological, immunohistological, and molecular study of these cases has allowed us to detect precise distinctive features of this SMZL variant. The cases included here were characterized by massive splenomegaly and a morphological picture showing a micronodular pattern of splenic involvement with follicle replacement, biphasic cytology, and marginal zone differentiation. Unlike classical SMZL cases, a conspicuous component of larger lymphocytes was distributed in the marginal zone ring, occasionally overrunning it, with isolated presence of the same cells within the central small cell component and also in the red pulp. The bone marrow and peripheral lymph nodes showed similar histological findings to those described for SMZL in these locations. The genetic and molecular study of these cases showed no alterations specific to other lymphoma types, such as t14;18 and t11;14. Instead of this, it showed 7q loss in 3 of 5 cases, p53 inactivation in 2 of 6 cases, cyclinD1 overexpression in 2 of 6 cases, and the presence of translocations involving the 1q32 region in 2 of 4 cases. The recognition of this aggressive variant, besides offering a prognostic indication, could lead to a more suitable form of clinical management of these patients. Further molecular studies would clarify the role of the different genetic alterations found. PMID- 10534162 TI - Atypical immature metaplastic-like proliferations of the cervix: diagnostic reproducibility and viral (HPV) correlates. AB - Although some immature squamous lesions (papillary immature metaplasias) of the cervix have been described and associated with human papillomaviruses (HPV), nonpapillary atypical immature squamous proliferations (AISPs) are a poorly defined entity and range from atypical reactive metaplasias to squamous intraepithelial lesions resembling immature metaplasia. This study examined the diagnostic reproducibility of AISPs and their relationship to HPV nucleic acids. Forty-four diagnostically problematic AISPs were studied. Based on nuclear density (crowding), chromasia, variation (anisokaryosis) in nuclear size, and surface cytoplasmic maturation, cases were independently scored by 2 observers as (1) probably reactive (Rx), (2) not otherwise specified (NOS), and (3) squamous intraepithelial lesion (SIL). Extracted archival DNA was scored for HPV by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis. Interobserver reproducibility (kappa statistic) and HPV correlates (chi square) were computed. Approximately one third of cases were classified in each category by the observers. Interobserver reproducibility was excellent (0.80), poor (0.23), and fair to good (0.41) for a diagnosis of Rx, NOS, and SIL, respectively. Differences in HPV DNA positivity between Rx and SIL were significant for both observers (5.8% to 6.7% v 38.4% to 50.0%, respectively); however, differences between NOS and SIL (30.7% to 42.8% v 38.4% to 50.0%) were not, even when cases were limited to those in which both observers agreed (28.6% v 37.5%). By light microscopy, AISPs exceeding the threshold for presumed reactive changes (NOS or SIL) are a morphologically heterogeneous group that defy precise classification. Furthermore, their histopathologic appearance, even when there is diagnostic agreement, does not consistently correlate with their HPV status. The laboratory management of AISPs should take into account the uncertainty of this diagnosis. PMID- 10534163 TI - Differential expression of cytokeratins in follicular variant of papillary carcinoma: an immunohistochemical study and its diagnostic utility. AB - The follicular variant of papillary carcinoma (FVPTC) is characterized by follicular growth pattern and tumor cells with appropriate nuclear features of papillary carcinoma. However, occasionally these lesions may show focal or multifocal instead of diffuse distribution of nuclear features of papillary carcinoma. Such lesions can be underdiagnosed as benign follicular nodule. Previous studies have shown that cytokeratins, especially 19, are helpful in differentiating papillary carcinoma from other benign and malignant follicular patterned lesions. In this study, we applied monoclonal antibodies to CK5/6/18, CK18, CK10/13, CK20, CK17, and CK19 to paraffin sections of formaldehyde-fixed tissue from 26 cases of FVPTC with multifocal distribution of papillary cancer nuclei, 10 cases of usual variant of papillary carcinoma, 1 case of Warthin's tumor-like papillary carcinoma, and 2 cases of the columnar cell carcinoma. CK19 stained strongly and diffusely all cases of papillary carcinoma. FVPTC cases showed strong staining of the areas with papillary cancer nuclei in all cases and moderate to strong staining in areas of tumor without obvious nuclear features of papillary cancer. Normal thyroid parenchyma adjacent to the tumor nodule showed focal staining in most cases; however, tissue away from the tumor nodule failed to show any staining. All cases of usual type of papillary carcinoma, 2 of columnar cell carcinoma, and 1 Warthin's tumor-like papillary carcinoma showed strong and diffuse staining with CK19 and failed to show any staining of adjacent normal thyroid parenchyma. Similar but less intense staining patterns were seen with CK17 and CK20. The control group, consisting of cases of follicular adenoma, follicular carcinoma, and hyperplastic nodule, showed no staining with CK19. We suggest that if one is using immunohistochemistry to aid in the diagnosis of cases of FVPTC with multifocal distribution of nuclear features of papillary cancer, an antibody panel comprising CKs 17, 19, and 20 may prove helpful. In addition, we hypothesize that the staining of adjacent nontumorous thyroid parenchyma with CK19, seen only in cases of FVPTC, suggests that some factors secreted/produced by this particular tumor may lead to modification in keratin expression of surrounding follicular epithelium. PMID- 10534164 TI - Apolipoprotein E genotype and its pathological correlation in Chinese Alzheimer's disease with late onset. AB - In this study, we attempted to find a relationship between apoliprotein E (ApoE) genotypes and Alzheimer's disease (AD) pathology in different areas of the brain in Chinese. We also studied the borderline group of possible AD (Poss). There were 34 definite or probable AD (Ad), 18 Poss, and 123 brains from age-matched normal subjects (N). ApoE genotype was determined by nested polymerase chain reaction on genomic DNA extracted from archival paraffin-embedded materials. Hippocampus (including entorhinal cortex), amygdala, superior temporal lobe, middle frontal gyrus, and inferior parietal lobule of the brains of Ad and Poss were examined with beta amyloid (A beta) immunostaining, and the same regions plus medial occipital lobe were examined with tau immunostaining. The percentage of plaque area stained for A beta in each brain region was obtained by an image analyzer, and the average number of neurofibrillary tangles stained for tau was counted with an eyepiece graticule. ApoE epsilon4 frequency was increased in both Ad (22.1%, chi2, df = 1, P = .00005), and Poss (33.3%, P = .000005) compared with N (5.3%). A beta load was significantly increased in the neocortex in Ad examined with at least 1 copy of epsilon4 compared with subjects without epsilon4 (Mann Whitney, P = .014). The same trend, though not statistically significant, occurred in Poss (P = .15). Tau expression was associated with ApoE epsilon4 in neither Ad nor Poss. Poss is genetically and histologically similar to Ad, although the overall A beta load is significantly increased in the latter. These findings support the recent Consensus Report's findings that all Alzheimer-type pathology may be significant. PMID- 10534165 TI - Grading dysplasia in colorectal adenomas by means of the quantitative binding pattern determination of Arachis hypogaea, Dolichos biflorus, Amaranthus caudatus, Maackia amurensis, and Sambucus nigra agglutinins. AB - The current study deals with the setting up of a new tool that enables the benign versus the malignant nature of colorectal adenomas to be determined accurately. The 2 objectives are to determine (1) whether adenomas should, or should not, be included in a 2- or a 3-tier grading system, and (2) whether severe dysplasias and carcinomas in situ share common or different biological characteristics. The levels of expression of different types of glycoconjugates were characterized in a series of 166 colorectal specimens, including 14 normal, 90 dysplastic, and 62 cancerous cases. The glycoconjugate expressions were demonstrated for 5 lectins, namely, Arachis hypogaea (PNA), Dolichos biflorus (DBA), Amaranthus caudatus (ACA), Maackia amurensis (MAA) and Sambucus nigra (SNA). The glycoconjugates demonstrated by these 5 lectins belong to the family of the Thomsen-Friedenreich antigens. The binding patterns of the 5 lectins were quantitatively determined by means of computer-assisted microscopy. The quantitative data were submitted to discriminant analyses. Our results show that the specific glycochemical staining patterns could be identified unambiguously and without misclassification between benign (normal and low dysplasia) and malignant (ie, either as moderate/severe dysplasia, carcinoma in situ, or cancer) cases. The data also strongly suggested that (1) dysplasias seem to be distinguishable in 2 instead of 3 groups, that is, low versus moderate/severe (high); and (2) moderate/severe dysplasias are biologically distinct from carcinomas in situ. The methodology developed can be applied directly in routine diagnosis to identify moderate/severe dysplasia specimens already exhibiting features common to carcinomas, and which therefore should be treated consistently in view of the fact that our data strongly suggest that most moderate/severe dysplasias are still benign, whereas carcinomas in situ are real carcinomatous lesions. PMID- 10534166 TI - Intraluminal fibromyxoid lesions in bronchiolitis obliterans organizing pneumonia are highly capillarized. AB - Bronchiolitis obliterans organizing pneumonia (BOOP) and usual interstitial pneumonia (UIP; ie, cryptogenic fibrosing alveolitis of mural type, CFA) are clinically and histologically distinguishable interstitial lung diseases. Both contain intraluminal lesions of newly formed fibromyxoid connective tissue. In BOOP, the fibromyxoid lesions are susceptible to complete reversal, but in UIP they are supposed to participate in the remodeling of the interstitium. Our hypothesis was that capillarization of the intraluminal fibromyxoid lesions is more frequent in BOOP compared with UIP. In this study, we stained diagnostic thoracoscopic or open lung biopsy specimens of patients with BOOP (n = 9) and UIP (n = 10) with antibodies against human laminin, von Willebrand factor, and CD34 to reveal the microvasculature of intraluminal fibromyxoid lesions. Our results show that in BOOP there is abundant capillarization in the newly formed intraluminal fibromyxoid lesions often reminiscent of granulation tissue. The mean number of capillaries per area unit (mm2) was 107 +/- SD 74 in samples stained for laminin, 103 +/- SD 46 for von Willebrand factor, and 63 +/- SD 36 for CD34. In marked contrast, in UIP, the corresponding accounts were significantly lower, being 14 +/- SD 15 for laminin (P < .003), 11 +/- SD 14 for von Willebrand factor (P < .001) and 6 +/- SD 6 for CD34 (P < .001). The intraobserver (P < .001) and interobserver correlations (P < .002) were highly significant, showing that our results are reproducible. We conclude that the content and nature of the newly formed intraluminal connective tissue, for example, in the form of vascular growth factors, are different in BOOP and in UIP, and this partly leads to the different clinical course of these diseases. PMID- 10534167 TI - Identical clonal origin of synchronous and metachronous low-grade, noninvasive papillary transitional cell carcinomas of the urinary tract. AB - One of the most important biological features of papillary transitional cell carcinoma (pTCC) of the urinary tract is its multicentricity and its tendency for recurrences. Two possible mechanisms, field effect and intramucosal seeding/spreading, have been proposed. The former theory hypothesizes that carcinogenic agents cause synchronous or metachronous malignant transformation of multiple urothelial cells (independent clonal origin), and the latter speculates that synchronous and metachronous tumors are derived from implantation or direct spreading of tumor cells (identical clonal origin). We tested these hypotheses by analyzing the methylation patterns of the androgen receptor gene (HUMARA) located at the X-chromosome. Thirty-five metachronous and synchronous, low-grade (grade 1 or 2), noninvasive pTCCs of the urinary tract from 10 heterozygous female patients were successfully analyzed using formalin-fixed, paraffin-embedded tissue. These included 16 recurrent bladder tumors from 4 patients, 10 metachronous bladder and ureter/renal pelvis tumors from 4 patients, and 9 multifocal tumors from 2 patients. All tumors are monoclonal as indicated by unbalanced methylation of HUMARA. Furthermore, same methylated allele was detected in multiple recurrent or multifocal tumors from any given patient, indicating their identical clonal origin. We conclude that low-grade, noninvasive pTCCs are monoclonal in nature. Synchronous or metachronous pTCCs have an identical clonal origin, strongly supporting the intramucosal seeding/spreading hypothesis. PMID- 10534168 TI - The testis-specific expression pattern of the growth hormone/placental lactogen (GH/PL) gene cluster changes with malignancy. AB - Growth hormone (GH) and placental lactogen (PL) gene transcription patterns in testicular germ cell tumors (GCT) and normal testicular tissue were comparatively investigated to identify GH/PL gene products associated with the development of GCT. This was done by nondiscriminative reverse transcriptase-polymerase chain reaction (RT-PCR), amplifying all major transcripts of any of the 5 GH/PL genes- GH-N(ormal), GH-V(ariant), PL-A, PL-B, PL-L(ike)--and subsequent analytical restriction enzyme analyses of 5'-end radioactively labeled cDNA. Surprisingly, all nonseminomatous GCT (NSGCT; n = 9) expressed GH-N, PL-A/B, and PL-L transcripts (9 of 9). Seminoma (n = 7) showed a distinctly unique pattern of GH-N and PL-A/B. GH-V products, which are hallmarks of the normal healthy testis, were not detected in any testicular cancer specimen (0 of 16). The fact that both seminomatous and NSGCT showed alterations in the same gene cluster indicates a pathogenetic relationship. Two choriocarcinoma cell lines of conceptus origin, BeWo and JAR, clearly differing from the male counterparts, exhibited a placental derived pattern of PL-A/B and GH-V. Obviously, profound differences exist between conceptus and male germ cell GH/PL gene cluster transcription. In summary, the unique testicular pattern of GH/PL gene expression changes significantly and in directed ways with malignancy. Loss of GH-V gene expression in testicular GCT compared with normal testis and loss (seminoma) or mutation (NSGCT) of PLL gene products might have significance in terms of the relationship between these tumors and for testicular GCT development. PMID- 10534169 TI - Expression of neural cell adhesion molecules and neurofilament protein isoforms in Ewing's sarcoma of bone and soft tissue sarcomas other than rhabdomyosarcoma. AB - In a previous study, it was shown that rhabdomyosarcomas widely express "neural" markers, such as neural cell adhesion molecules (N-CAM) and neurofilament protein isoforms. In the current study, a series of Ewing's sarcomas of bone and soft tissue sarcomas other than rhabdomyosarcoma was probed for the same antigens. It was found that N-CAM was widely expressed in the various sarcoma types, except Ewing's sarcomas, though less than in rhabdomyosarcomas. Similar to rhabdomyosarcomas, neurofilament isoforms were found throughout all sarcoma types but were largely restricted to those expressed early in neurogenesis, that is, poorly phosphorylated medium-weight isoforms. Neurofilament expression was most extensive in Ewing's sarcomas. It was concluded that the expression of "neural" markers (N-CAM, neurofilaments) is widespread in sarcomas and does not signify a neural tumor. The absence of N-CAM expression in Ewing's sarcoma may be helpful in its distinction from other sarcomas. PMID- 10534170 TI - Gastrointestinal stromal tumors: recent advances in understanding of their biology. AB - Gastrointestinal stromal tumor (GIST) is the preferred term for mesenchymal tumors specific for the gastrointestinal tract (60% in stomach, 30% small intestine, 10% elsewhere). GISTs include most tumors previously designated as leiomyoma, cellular leiomyoma, leiomyoblastoma, and leiomyosarcoma. However, in the esophagus, leiomyoma is the most common mesenchymal tumor. GISTs are composed of spindle (70%) or epithelioid (30%) cells, and 10%-30% are malignant showing intra-abdominal spread or liver metastases. They are immunohistochemically positive for c-kit (CD117), CD34, and sometimes for actin but are almost always negative for desmin and S100-protein. The malignant GISTs especially show activating mutations in the c-kit gene. GISTs and gastrointestinal autonomic nerve tumors (GANT) overlap. The cell of origin is not fully understood, but resemblance to the interstitial cells of Cajal, expression of some smooth muscle markers, and occurrence outside of the GI-tract suggest origin from multipotential cells that can differentiate into Cajal and smooth muscle cells. PMID- 10534171 TI - Immunohistochemical detection of p53 protein accumulation in head and neck cancer: correlation with p53 gene alterations. AB - The genetic and functional status of the p53 gene may be an important factor in guiding therapeutic strategies for patients with cancer. The purpose of this study was to determine whether p53 immunohistochemistry (IHC) accurately reflects the mutational status of the p53 gene and to determine whether p53 IHC independently predicts tumor responsiveness to radiation therapy for patients with HNSCC. p53 IHC was performed using the monoclonal antibody DO7 on tumors from 85 patients with HNSCC treated with primary or adjuvant radiation. The p53 status in all of these tumors was previously assessed by direct sequence analysis of exons 5 through 9: 49 tumors were p53 wild-type, and 36 harbored p53 gene mutations. All patients were well characterized with respect to locoregional recurrence, distant spread, and survival. Positive p53 staining was observed in 53 of the 85 cases (62%). Only 27 (51%) of these 53 IHC-positive cases harbored gene mutations in exons 5 through 9; 23 (72%) of the 32 IHC-negative cases did not harbor mutations. The overall correlation rate between IHC and sequencing was 59% (P < .04, chi2). Discordant results were observed for 35 (41%) cases, including 26 IHC-positive cases and 9 IHC-negative cases. In 7 of 9 cases, false negative staining was due to a nonsense or splice-site mutation. p53 IHC was not predictive of overall survival (P = .37) or disease-free survival (P = .95). In a sizable number of cases, p53 IHC does not reflect the mutational status of the p53 gene. Specific types of alterations (eg, truncating mutations) and other factors may contribute to this poor correlation. Moreover, p53 IHC does not appear to be an independent predictor of tumor responsiveness to radiation in patients with HNSCC. PMID- 10534172 TI - Confocal laser scanning microscopic observation of angioarchitectures in human thyroid neoplasms. AB - Confocal laser scanning microscopy (CLSM) was employed to study the blood vascular system of human thyroid tumors. CLSM observation combined with immunohistochemistry for type IV collagen clearly visualized 3-dimensional images of the microvascular structures. CLSM observation showed that normal thyroid follicles were tightly covered by branching microvessels, whereas microvessels in follicular adenomas were more prominent and more irregular in shape. Microfollicular adenomas showed that neoplastic small follicles were attached to the capillaries and had a "grape-like" appearance. Strikingly well-developed vascular networks were seen in neoplastic follicles of papillary carcinomas. Interestingly, papillae of papillary carcinoma occasionally contained contained aggregated vascular complexes (glomeruloid structure) composed of tortuous, densely packed, and irregularly arranged small vessels. Such aggregated vascular complexes were seen in 5 of 7 papillary carcinoma tissues but not in other histological thyroid tumors. Our findings indicate that the fundamental vascular pattern correlates well with the growth pattern, suggesting an interdependence between parenchyma and stroma characteristic for thyroid tumors. CLSM observation combined with immunohistochemistry may contribute to a better understanding of morphological characteristics of angioarchitecture in human surgical materials. PMID- 10534173 TI - Angiogenesis in carcinosarcomas of the uterus: differences in the microvessel density and expression of vascular endothelial growth factor between the epithelial and mesenchymal elements. AB - Carcinosarcoma of the uterus is a highly aggressive neoplasm. However, the angiogenesis of this neoplasm is still unknown. This is the first study to examine the differences in angiogenesis between the epithelial and mesenchymal elements of this biphasic neoplasm. Surgical specimens from 21 primary uterine carcinosarcomas were histopathologically evaluated, and then immunohistochemically analyzed for tumor angiogenesis, using an anti-vascular endothelial growth factor (VEGF) antibody. The microvessel density (MVD) was also measured in each element of these neoplasms, using anti-CD34 monoclonal antibody. The MVD in the epithelial element was found to be higher than that of the mesenchymal element in 20 of 21 (95.2%) primary tumors. The epithelial elements showed a higher MVD (mean, 81.6 +/- 41.1) than the mesenchymal elements (mean, 36.7 +/- 23.8) in these primary tumors (P < .0001). Moreover, the epithelial elements showed a higher VEGF expression (mean, 0.78 +/- 0.23) than the mesenchymal elements (mean, 0.37 +/- 0.20) (P < .0001). The tumors with lymph vascular invasion showed a higher VEGF expression (n = 17; mean, 0.85 +/- 0.17) than the tumors without lymph-vascular invasion (n = 4, mean, 0.47 +/- 0.12) (P < .01). Microscopically, neither lymph-vascular space invasion nor metastatic tumors consisted of sarcoma alone in this series. In addition, a decrease in the VEGF expression was found in the transitional areas between carcinomatous and sarcomatous elements in all 10 homologous and 4 heterologous tumors evaluated. These results suggest that the tumor angiogenesis in the epithelial element may be more active than that of the mesenchymal element and also substantiated the high metastatic potential of the epithelial element in uterine carcinosarcoma. Based on these findings, carcinoma cells thus may play a key role in the angiogenesis of this biphasic neoplasm. PMID- 10534174 TI - Analysis of chromosome 9p21 deletion and p16 gene mutation in salivary gland carcinomas. AB - The chromosomal locus 9p21 contains the p16(INK4a/CDKN2/MTS1) tumor suppressor gene that has been implicated in a variety of tumor types, including carcinomas of the head and neck, esophagus, and pancreas. To determine whether the loss of this gene is involved in salivary gland cancers, 35 carcinomas and paired nonneoplastic specimens were analyzed for loss of heterozygosity (LOH) of polymorphic genetic markers located in the region of interest. Five types of salivary gland tumors were studied: mucoepidermoid carcinoma, salivary duct carcinoma, adenoid cystic carcinoma, acinic cell carcinoma, and polymorphous low grade adenocarcinoma. Seven of 9 salivary duct carcinomas showed LOH of 1 or more polymorphic markers. In 1 case of salivary duct carcinoma with LOH, we confirmed a deletion of bp 240-254 within exon 2. In addition, another salivary duct carcinoma showed a homozygous deletion of p16 in differential polymerase chain reaction analysis. Loss of heterozygosity was found in 1 of 10 adenoid cystic carcinomas and 1 of 8 mucoepidermoid carcinomas and was absent in the remaining subtypes. No mutations in exon 1 or exon 2 or homozygous deletion of p16 were found in these 2 particular neoplasms with LOH. These results suggest that inactivation of p16 is important in the development or progression of at least some salivary duct carcinomas, but we found no evidence that its alteration plays a role in the other subtypes examined. PMID- 10534175 TI - Clinical significance of genetic imbalances revealed by comparative genomic hybridization in chondrosarcomas. AB - DNA copy number changes were studied by comparative genomic hybridization (CGH) in 50 chondrosarcoma samples from 45 patients. Mean number of genetic aberrations in primary tumors was 4.8 +/- 1.8. The most frequently gained regions were 20q12 qter (37%), 20q (32%), 8q24.1-qter (27%), 20p (24%), and 14q24-qter (24%). Losses were 5.5 times less frequent than gains and observed mainly at Xcen-q21, 6cen q22, and 18cen-q11.2 (11% each). Recurrent and metastatic tumors showed a mean of 4.0 +/- 2.2 aberrations per sample. The most frequently gained regions were chromosome 7 (4 cases), 5q14-q32 (4 cases), 6p (3 cases), and 12q (3 cases). Losses of DNA sequences were 3.4 times less frequent than gains. Histological tumor grade was significantly associated with metastasis-free survival (P = .002) and overall survival (P = .003), being the strongest prognostic factor tested. A statistically significant correlation was found between gain at 8q24.1-qter and shorter overall survival (P = .01) but not with local recurrence or metastasis free survival. Gain at 14q24-qter was associated with a trend to shorter overall survival (P = .05) but neither with an increased risk for local recurrence nor with metastasis-free survival. In a multivariate analysis, only the tumor grade associated with overall survival (P = .02). In a multivariate analysis together with the tumor grade, gain at 8q24.1-qter did not retain its significance (P = .44), indicating that this imbalance is not an independent prognostic factor. PMID- 10534176 TI - Large cell, epithelioid, telangiectatic osteoblastoma: a unique pseudosarcomatous variant of osteoblastoma. AB - A previously undescribed large-cell, epithelioid, and aneurysmal variant of osteoblastoma with minimal osteoid-production--simulating telangiectatic osteosarcoma, epithelioid angiosarcoma, and metastatic carcinoma is reported. The tumor occurred in the mandible of a 14-year-old girl. The light microscopic, immunohistochemical, ultrastructural, cell proliferation, and DNA-ploidy studies, as well as the 7-year disease-free follow-up period all indicate a benign osteoblastic tumor. Cytogenetically, the tumor had a pseudodiploid karyotype, distinguished by a complex t(1;5;17;22) and a terminal 1q deletion. Recognition of this unique, pseudomalignant variant of osteoblastoma is important to avoid an erroneous diagnosis of malignancy. PMID- 10534177 TI - Nasal glomus tumors: report of two cases with emphasis on immunohistochemical features and differential diagnosis. AB - We describe 2 cases of nasal glomus tumor that presented as nasal polyps. Grossly, each of the polypectomy specimens consisted of small fragments of polypoid soft tissue with glistening mucosa. Histopathological examination of each of the specimens showed sheets and nests of monomorphic round cells intimately associated with capillary-sized blood vessels. The tumor cells were strongly cytoplasmic positive for vimentin, smooth-muscle specific actin, muscle specific actin, and CD34. Collagen IV showed pericellular positivity. Nasal glomus tumors are extremely rare and represent less than 0.5% of nasal nonepithelial tumors. Nasal polyps are common surgical pathological specimens, with the majority of nasal polyps being inflammatory polyps or a respiratory epithelial proliferation. Histologically, many nasal polyps show vascular proliferation with an inflammatory cell infiltrate, which may be confused with the rare glomus tumor. In addition, other nasal vascular tumors, in particular nasal hemangiopericytoma and neural tumors, may histologically mimic nasal glomus tumors. PMID- 10534178 TI - Recurrent Epstein-Barr virus-associated post-transplant lymphoproliferative disorder: report of a patient with histologically similar but clonally distinct metachronous abdominal and brain lesions. AB - A liver transplant patient developed a single central nervous system (CNS) intraparenchymal lesion 5 months after the diagnosis of an intraabdominal diffuse large B-cell post-transplant lymphoproliferative disorder (PTLD). Biopsy of the new CNS lesion showed a diffuse large B-cell PTLD morphologically and immunohistochemically indistinguishable from the abdominal lesion. In addition, both lesions were positive for Epstein-Barr virus (EBV) DNA by polymerase chain reaction (PCR) and for EBV-encoded RNA by in situ hybridization. Although these results were consistent with a metastatic origin for the CNS lesion, the finding of an intraparenchymal lesion without leptomeningeal or dural spread was suggestive of a new primary CNS lymphoma. Proof that the brain lesion was a second primary and not a metastasis was obtained by immunoglobulin gene rearrangement studies and assessment of EBV clonality. Multiple primary lymphoid neoplasms arise at higher frequency in the setting of immunosuppression, and molecular investigations of tumor clonality can provide clinically relevant staging and prognostic information. PMID- 10534179 TI - Pulmonary angiomyolipoma and multifocal micronodular pneumocyte hyperplasia associated with tuberous sclerosis. AB - A 36-year-old woman with a long-standing diagnosis of tuberous sclerosis was found dead. A yellow-tan 0.4 cm-diameter pulmonary tumor was identified at autopsy which had typical microscopic features of an angiomyolipoma (AML). Immunohistochemical stains showed reactivity for actin, but not HMB-45, Melan-A, and tyrosinase (despite reactivity of the patient's renal AML for HMB-45 and Melan-A), perhaps owing to the small size of the lesion and the sometimes focal nature of the reactivity for these markers. Additional lung nodules proved to be multifocal micronodular pneumocyte hyperplasia. This report highlights the first occurrence of a pulmonary angiomyolipoma in the setting of tuberous sclerosis. PMID- 10534180 TI - Lack of Kaposi's sarcoma-associated virus (KSHV) and detection of human herpes virus 6 and 7 by PCR in African Burkitt's lymphoma from HIV-negative patients. PMID- 10534181 TI - MUC 5 expression in breast carcinomas. PMID- 10534182 TI - Experience with Salmonella typhi Vi capsular polysaccharide vaccine. AB - Typhoid fever remains an important health threat in many parts of the world, with an estimated 16 million cases and 600,000 deaths occurring each year. The emergence of Salmonella typhi strains multiply resistant to antibiotics has complicated the treatment of this disease. Field experience of 8 years shows that a vaccine composed of purified Vi capsular polysaccharide of Salmonella typhi, given as a single intramuscular or deep subcutaneous injection, has consistent immunogenicity and efficacy. Side effects, based on reports since 1989, are infrequent and mild. Furthermore, the Vi vaccine may be administered simultaneously with other common "travel" vaccines, at two different sites of injection, without affecting immunogenicity and tolerability. This review presents an update of the development and clinical experience with the Salmonella typhi Vi polysaccharide vaccine (Typhim Vi; Pasteur Merieux Connaught, France). PMID- 10534183 TI - Evaluation of treatment responses in late Lyme borreliosis on the basis of antibody decrease during the follow-up period. AB - The purpose of this study was to identify a serological marker of successful treatment as distinct from treatment failure in late Lyme borreliosis. Consecutive serum samples from 68 treated patients with late Lyme borreliosis were analyzed during a 1-2 year follow-up period after the start of treatment. End-point enzyme immunoassay titres of IgG1, IgG2, IgG3, and combined IgG1+3 subclasses against a sonicate antigen of Borrelia burgdorferi were determined and compared to the IgG antibody response against Borrelia burgdorferi flagella. Individual half-lives of the antibody levels were calculated for each patient. The half-life values were compared to the patients' clinical outcome in order to find a serological marker of remaining disease activity or relapse. The levels of combined IgG1+3 subclass antibodies against the sonicate antigen and the individual levels of IgG1, IgG2, and IgG3 antibodies did not change significantly after treatment. In contrast, antibodies to flagella decreased markedly after successful treatment, with a half-life of 112+/-92 days (arithmetic mean value +/ SD). This was significantly shorter than the half-life after unsuccessful treatment (271+/-151 days), (P<0.0001). The decrease was observed mainly in IgG1 and IgG4 responses to flagella, less so for IgG2 or IgG3. The results suggest that a rapid decrease in flagella antibodies can serve as a marker for a successful treatment of Lyme borreliosis. PMID- 10534184 TI - Usefulness of pp65 antigenemia for the early diagnosis of cytomegalovirus disease in patients with AIDS. AB - An observational cohort study was performed to assess the effectiveness of a cytomegalovirus antigenemia (CMV-Ag) assay designed to predict clinical CMV disease in patients with AIDS. Eighty-six HIV-infected patients with CD4+ cell counts of < 100/mm3, positive CMV IgG, and no previous CMV disease were enrolled. Thirty-eight (44%) patients had at least one positive CMV antigenemia test, ten of whom eventually developed CMV focal disease. CMV disease was diagnosed in 13 (15%) patients. The CMV antigenemia assay was positive in ten of these 13 patients. Using a cut-off value of five positive cells in every 150,000 leukocytes sampled, the CMV antigenemia assay had a positive predictive value of 89% and a negative predictive value of 94%. The median time from the first positive CMV antigenemia test to the onset of CMV disease was 102 days. CMV disease probability at 6 months in patients with a CMV antigenemia value > or = 5 was 77.8% versus 6% in patients with CMV antigenemia value < 5 (log-rank test = 48.345; P < 0.001). Several independent factors were associated with the development of CMV disease: CMV antigenemia > or = 5 cells (hazard ratio: 20.44), CD4+ count < or = 25/mm3 (HR: 3.12), and sexual transmission of HIV infection (hazard ratio, 3.15). CMV antigenemia seems to be a good predictor of CMV disease in patients with AIDS. PMID- 10534185 TI - Immune response to natural and recombinant antigens of Helicobacter pylori in patients with dyspeptic complaints. AB - Sera of 223 dyspeptic patients with endoscopic findings of nonulcer dyspepsia (72%), gastric ulcer (15%) and duodenal ulcer (13%) were tested for antibodies against Helicobacter pylori with an enzyme immunoassay and an immunoblot technique using lysates of Helicobacter pylori cells as antigen source. One hundred and fifty-one (68%) sera were found to be positive for Helicobacter pylori IgG with both methods; 5% of the positive results in the enzyme immunoassay were false-positive due to cross-reactions mainly of proteins with a molecular mass of 43-66 kDa. Since cross-reactivity not only reduces the diagnostic value of the immunoassay but also complicates evaluation of the immunoblot results, an attempt was made to overcome these problems by using specific purified recombinant proteins instead of the crude cell preparations as antigens. Of the commonly recognised immunogens of Helicobacter pylori, antibodies against a cell surface protein of 26 kDa, the small urease subunit (29 kDa) and the cytotoxin-associated protein (130 kDa) were identified as highly sensitive serological markers for inclusion in a recombinant antigen mixture for Helicobacter pylori screening. Only the cytotoxin-associated protein was confirmed to be an indicator immunogen for ulcerogenic strains. To assess the reliability of recombinant fragments of this protein in serological screening, the reactivity of antibody to purified fragments of the cytotoxin-associated protein was compared with that to the natural protein. A C-terminal recombinant fragment of 58 kDa showed results identical to those obtained with the natural protein and was thus considered to be an appropriate component of an antigen mixture for serological detection of Helicobacter pylori. PMID- 10534186 TI - Evaluation of the mecA femB duplex polymerase chain reaction for detection of methicillin-resistant Staphylococcus aureus. AB - This study systematically evaluated a recently described duplex polymerase chain reaction test for methicillin-resistant Staphylococcus aureus with 25 different German epidemic strains of methicillin-resistant Staphylococcus aureus and 66 staphylococci other than methicillin-resistant Staphylococcus aureus, including 17 different coagulase-negative staphylococcal species and subspecies, that were either oxacillin susceptible or oxacillin resistant. The results were compared with those of conventional cultural identification and susceptibility testing. Of the 91 isolates tested, all 25 confirmed strains of methicillin-resistant Staphylococcus aureus were identified correctly. None of the remaining strains of methicillin-susceptible Staphylococcus aureus was misidentified as methicillin resistant Staphylococcus aureus. It was concluded that the duplex polymerase chain reaction appears to offer a time-saving and accurate method of detection of methicillin-resistant Staphylococcus aureus. PMID- 10534187 TI - Performance of a Western blot assay to compare mother and newborn anti-Toxoplasma antibodies for the early neonatal diagnosis of congenital toxoplasmosis. AB - The aim of this study was to retrospectively evaluate the performance of a Western blot assay to compare mother and newborn anti-Toxoplasma gondii antibodies for the early neonatal diagnosis of congenital toxoplasmosis. Since specific anti-Toxoplasma IgM or IgA is detected inconstantly at birth in the neonate, the diagnosis of congenital toxoplasmosis is often delayed until 6-9 months, after IgG titers have been observed persistently. In this study, 81 paired samples from 60 mother/child pairs were tested for IgG and IgM patterns. All mothers had (or were strongly suspected to have) acquired toxoplasmosis during pregnancy. Specific IgM and IgA were simultaneously detected by immunocapture tests, and IgG was titrated. A serological and clinical follow-up of infants was conducted during the first year of life until the diagnosis of congenital toxoplasmosis could be either confirmed or ruled out. Seventeen of the 60 newborns were congenitally infected. Specific IgM or IgA was detected by immunocapture at birth in 76.5% and 70.6% of cord sera from infected neonates, respectively, with an equal specificity of 77.5%. Comparative Western blot allowed the detection of neosynthesized IgG and IgM in the cord blood of 50% and 78.6% of infected infants, respectively, with a specificity of 100%. The combination of IgA and IgM immunocapture tests, the analysis of IgG and IgM Western blot patterns, and the combination of both techniques allowed the detection of 94%, 94%, and 100% of cases within the first 3 months of life, respectively. In conclusion, Western blotting seems to be a useful complementary tool for the early postnatal diagnosis of congenital toxoplasmosis. PMID- 10534188 TI - Infective endocarditis due to Fusobacterium nucleatum in an intravenous drug abuser. AB - Infective endocarditis due to anaerobic non-spore-forming gram-negative bacilli in intravenous drug abusers is exceedingly rare, with only two cases being previously reported in the literature. A case of endocarditis due to Fusobacterium nucleatum in an intravenous drug abuser is reported, accompanied by a review of the literature. PMID- 10534189 TI - A case of HIV-associated cerebral histoplasmosis successfully treated with fluconazole. AB - Clinically apparent involvement of the central nervous system is a rare event in cases of disseminated histoplasmosis, even in HIV-infected persons. Despite therapy with amphotericin B, mortality remains very high. Reported here is the case of an HIV-infected patient with a 3-month history of fever, cough, weight loss and miliary lung infiltrates. Four weeks after initiation of tuberculostatic therapy, high-grade fever, neurological symptoms, personality changes and respiratory deterioration occurred. Magnetic resonance imaging of the brain showed multiple mass lesions, and a chest radiograph revealed worsening of pulmonary infiltrates. Methenamine silver staining of a lung biopsy specimen demonstrated Histoplasma capsulatum. Subsequently, this pathogen was cultured from lavage fluid. Following high-dose intravenous fluconazole therapy (800 mg once daily), the patient's condition improved markedly within 10 days, followed by an almost complete resolution of pulmonary and cerebral mass lesions. This is believed to be the first documented case of rapid improvement of disseminated histoplasmosis with central nervous system involvement in an HIV-infected patient upon induction of therapy with fluconazole. PMID- 10534190 TI - A case of disseminated histoplasmosis likely due to infection from a liver allograft. AB - Disseminated histoplasmosis in immunosuppressed patients rarely occurs in nonendemic areas. Reported here is a case of disseminated histoplasmosis in a patient who had undergone orthotopic liver transplantation and had never traveled outside of France. The infection was most likely transmitted via the liver allograft, since the organ donor had lived in an area highly endemic for the disease (French Guiana) 20 years earlier. Two years post-transplantation, none of the other patients who received allografts (heart, cornea, kidneys) from the same donor had developed signs of infection. PMID- 10534191 TI - Lack of induction by rhinoviruses of systemic type I interferon production or enhanced MxA protein expression during the common cold. AB - To study whether MxA protein expression is systemically upregulated during rhinovirus infection, blood specimens were collected from 40 patients with common cold and MxA expression in mononuclear cells analyzed by flow cytometry. None of the patients with a confirmed rhinovirus infection (n = 15) or with an infection of unknown etiology (n = 20) had elevated expression of the MxA protein (median fluorescence intensity, 549 and 582, respectively) when compared to healthy controls (n = 11, median 590). Patients with influenza infections had significantly elevated values (n = 5, median 750), and interferon could be detected only in serum samples from influenza patients. In conclusion, expression of MxA in blood lymphocytes and an apparently systemic type I interferon response is not induced during rhinovirus infection or during most other cases of common cold in young adult patients. PMID- 10534192 TI - Sensitivity and specificity of a rapid immunochromatographic test for diagnosis of visceral leishmaniasis. AB - A rapid immunochromatographic dipstick test has become available for the qualitative detection of total anti-Leishmania immunoglobulins using the recombinant K39 antigen. To evaluate the test, 96 serum specimens from patients with a variety of tropical infections were tested. Fourteen of the specimens derived from patients with parasitologically confirmed kala-azar, and all were strongly positive for antibodies to Leishmania donovani complex using the immunofluorescence test. Although all 82 samples negative by the immunofluorescence test were confirmed as negative by the dipstick assay, only 10 (71.4%) of the 14 positive samples were reactive. These results indicate that the test in its current form lacks sufficient sensitivity to be recommended as a screening tool, but it might be useful for indicating further diagnostic procedures in a clinical setting. PMID- 10534193 TI - First report of the presence of human T-cell lymphotropic virus I infection in Italian drug addicts. PMID- 10534194 TI - Management of Babesia divergens babesiosis without a complete course of quinine treatment. PMID- 10534195 TI - Polymicrobial endocarditis caused by methicillin-resistant Staphylococcus aureus and glycopeptide-resistant enterococci. PMID- 10534196 TI - Diabetes mellitus associated with protease inhibitor use. PMID- 10534198 TI - Seroepidemiological study of herpes simplex virus in the female population in the autonomous region of Madrid, Spain. PMID- 10534197 TI - Lack of evidence of heterozygosity for cystic fibrosis as a risk factor for invasive aspergillosis in haematological patients. PMID- 10534199 TI - Plasmodium falciparum malaria acquired by accidental inoculation. PMID- 10534200 TI - Predictors of condom use among patients with sexually transmitted diseases in Uganda. AB - BACKGROUND AND OBJECTIVE: Patients with sexually transmitted diseases (STDs) are at an increased risk of HIV infection and they must be targeted for increased condom use. GOAL: To identify predictors of condom use among patients with STDs. STUDY DESIGN: In a cross-sectional survey, an interview-administered questionnaire was administered to 138 patients at the STD clinic, Mulago, and the outpatients department, Mbarara Hospital, in Uganda. Data were collected on socio demographic situations, STD symptoms, type of sexual partners, and use of condoms. Multivariate logistic regression models were used to identify independent predictors of condom use. RESULTS: Of the 138 patients, 87 (66%) knew how to use condoms, 81 (59%) ever used a condom, 34 (25%) used a condom at least once in the previous 3 months, 20 (15%) used a condom during the last sexual intercourse, and 80 (58%) accepted a free supply of condoms. Reasons for not using condoms among the 57 who had never were: having a regular partner or spouse (28, 49%), partner does not approve (17, 30%), reduced sexual pleasure (5, 9%), and no answer (7, 12%). The independent predictors of condom use were: being a man, not having a regular partner, having had sex with a casual partner, being able to read English, having at least secondary education, and using electricity for lighting. CONCLUSION: Providing health promotion messages in local languages may improve condom use in this population. There is a need for complementary HIV prevention strategies for women and for regular sexual partnerships. PMID- 10534201 TI - Patterns of Chlamydia trachomatis testing and follow-up at a University Hospital Medical Center. AB - OBJECTIVE: Although testing for Chlamydia trachomatis is encouraged and increasingly practiced at sexually transmitted disease (STD) and family planning clinics, patterns of testing and follow-up in other settings are not well described. To begin to address these issues, we performed a chart review of patients with a positive laboratory test for C. trachomatis at a major university medical center. METHODS: Chart review of medical records for all patients with positive laboratory tests for C. trachomatis during calendar year 1996. RESULTS: Of 326 patients with positive tests, 95% were female and 5% were male. Median age was 22 for females and 25 for males. Most positive C. trachomatis test results were from the emergency room (ER)/walk-in clinic (55%) or patients receiving obstetric/gynecologic (OB/GYN) care (31%). While most C. trachomatis tests performed were on patients who had symptoms, patterns of treatment varied between sites. Fifty-seven percent of ER/walk-in patients received empiric antibiotics at the initial visit versus 36% of patients under OB/GYN care. Among patients with positive screening tests seen in the ER/walk-in clinic, 32% of patients had no treatment documented versus 14% of OB/GYN patients. Four percent of women with positive tests who did not receive therapy at the time of their initial evaluation developed pelvic inflammatory disease in the interval between testing and return to the medical center. CONCLUSIONS: Of the patients with positive chlamydial screening tests, the proportion not treated was similar to that found in studies performed in STD clinics. PMID- 10534202 TI - Disproportionate high incidence of invasive cervical cancer as an AIDS-indicative disease among young women in Catalonia, Spain. AB - BACKGROUND: Spain has the highest rate of AIDS in Europe since 1990, mainly at the expenses of the intravenous drug user (IDU) transmission group, and women account for 20% of all the cases. Because cervical lesions may be altered by HIV, invasive cervical cancer (ICC) is included in the AIDS case definition worldwide. GOAL OF STUDY: To assess the impact of the HIV epidemic on the incidence of ICC and to describe the characteristics of ICC in the spectrum of AIDS-defining diseases in Catalonia, Spain. STUDY DESIGN: Descriptive study of women notified through the AIDS surveillance system during 1994 to 1996. Age-specific ICC incidence rates from a population-based Cancer Registry were used to calculate the population attributable risk percent (PAR%). RESULTS: Fifty-six cases (6.8%) with ICC were reported to the AIDS Registry, 64.3% of those women were IDU with a mean age of 32. Among women aged 20 to 49, the PAR% was 12.2% (95% CI: 7.9-16.5%) and the incidence rate ratio was 18.5 (95% CI: 11.2-29.2). CONCLUSIONS: HIV is having an impact on ICC epidemiology among young women in Catalonia. The policy of cervical screening should be modified and offered every 6 to 12 months to women at risk for or with HIV infection. PMID- 10534203 TI - The impact on accuracy and cost of ligase chain reaction testing by pooling urine specimens for the diagnosis of Chlamydia trachomatis infections. AB - BACKGROUND AND OBJECTIVES: Nucleic acid amplification testing is the most accurate approach to diagnosing Chlamydia trachomatis infections. Our objective was to compare the accuracy and cost savings of pooling urines as opposed to individual testing. STUDY DESIGN: Strategies of pooling urine specimens into groups of four (4x pool) or eight (8x pool) followed by testing the positive pools individually were compared to individual specimen testing to determine if significant cost savingS could be realized without compromising the sensitivity and specificity of the LCx C. trachomatis Assay (Abbott Laboratories, Abbott Park, Chicago, IL) performed in a busy private medical laboratory. RESULTS: A total of 1,220 patient urine samples, 1,187 male (97%) and 33 female (3%), were tested using the normal LCx specimen to cutoff ratio (S/CO) of 1.0 and a decreased S/CO value of 0.2. Individual testing identified 98.2% (109/111) of positive urines. The 4x pooling maneuver identified 92.8% (103/111) of positive patients with the regular cutoff and 96.4% (107/111) when the cutoff was decreased. These values were 95.9% (47/49) and 97.9% (48/49), respectively, when eight urines were pooled. Both pooling and individual testing strategies identified all the negative samples accurately. Cost savings of pooling were calculated to be 44.5% for pools of four and 37.5% for pools of eight, applying the lowered cutoff. CONCLUSIONS: Pooling urine specimens for testing with the C. trachomatis LCx system is a simple, accurate, and cost-saving approach that can significantly reduce the cost of amplified nucleic acid testing with minimal sacrifice of testing accuracy. PMID- 10534204 TI - Providing low-cost sexually transmitted diseases services in two semi-urban health centers in Central African Republic (CAR): characteristics of patients and patterns of health care-seeking behavior. AB - BACKGROUND: While treatment of symptomatic sexually transmitted diseases (STDs) has been shown to reduce the incidence of HIV infection, there are few published reports describing the delivery of high quality STD care in Africa. GOAL: To test the feasibility of providing comprehensive, affordable STD services through the existing primary care infrastructure. DESIGN: STD treatment services using a syndromic' approach were established in two semi-urban hospital outpatient departments (OPD) in Central African Republic (CAR). A dedicated paramedical provider took a clinical history, performed an examination, explained the diagnosis and the importance of referring partners, dispensed drugs, and offered partner referral vouchers. A fee-for-service system was used to resupply drugs initially purchased with project funds. RESULTS: Of 9,552 visits by index patients and partners over a 28-month period starting in October 1993, 60% were made by women; of these women, 90% were symptomatic, 77% had "vaginal discharge," 70% "lower abdominal pain," and 7% "genital ulcer." Among men, 64 % were symptomatic, 38 % had "urethral discharge," and 14% "genital ulcer." Half of all symptomatic patients presented within 1 week of the onset of symptoms; 44% of men compared to 18% of women had sought care elsewhere before the clinic visit. The average cost per STD treated with recommended drugs was $3.90. Etiologic data from subpopulations in both sites suggest that a high proportion of patients was infected with an STD. CONCLUSIONS: Comprehensive yet affordable care for STDs in persons (and their partners) who recognize symptoms is feasible and should be widely implemented in primary care systems to prevent the spread and complications of STDs and HIV in Africa. PMID- 10534205 TI - Sexually transmitted diseases services in societies with limited resources. PMID- 10534207 TI - Genital ulcer caused by human bite to the penis. AB - BACKGROUND AND OBJECTIVES: Human bite injuries, while less frequent than cat or dog bites, usually stem from aggressive behavior, sports, or sexual activity. It has been thought that human bites have a higher rate of infection than animal bites, but this view is likely skewed because of the frequency of closed fist injuries presenting to emergency rooms. Human bites to the genitalia also occur, but are not often reported because of embarrassment. GOAL OF THE STUDY: We report a genital ulceration after a human bite to the penis and review appropriate diagnostic and therapeutic maneuvers. STUDY DESIGN: This article reports the development of a severe genital ulcer associated with a human bite to the penis secondarily infected, as verified by culture, with an oral flora organism Eikenella corrodens. RESULTS: The genital ulceration healed after appropriate antibiotic therapy. CONCLUSIONS: Treatment of human bites focuses on obtaining an accurate history and performing a salient physical examination, as well as early irrigation and debridement. Transmission of communicable disease should be considered as a possible consequence. Prophylactic antibiotic treatment and primary closure of wounds continue to be areas of controversy. PMID- 10534206 TI - Control of Chlamydia trachomatis infections in female army recruits: cost effective screening and treatment in training cohorts to prevent pelvic inflammatory disease. AB - CONTEXT: Chlamydia trachomatis genitourinary infections in females can lead to serious and costly sequelae. Programs such as basic (initial entry) military training with controlled points of entry offer an opportunity to screen large cohorts of women at risk for infection. OBJECTIVE: To assess the cost effectiveness of three interventions for C. trachomatis infections in women beginning Army training: 1) screening using urine ligase chain reaction (LCR) by age, 2) unrestricted testing using urine LCR, and 3) universal antibiotic treatment with azithromycin. DESIGN: Cost-effectiveness analysis from a military perspective. SETTING AND PATIENTS: A hypothetical cohort of 10,000 women who intended to complete at least 2 years of military service was studied. Analysis was based on data from 13,204 female trainees screened for chlamydial infection at Fort Jackson, SC. OUTCOMES: Program and training costs, cost of illness averted, and pelvic inflammatory disease (PID) prevented were determined for a 1 year follow-up period. Using sensitivity analysis, outcomes over 2 years were studied. RESULTS: At a 9.2% prevalence, no screening resulted in $220,900 in training and sequelae costs and 276 cases of PID. Screening by age produced the lowest cost $217,600, over a 1-year period and prevented 222 cases of PID for a cost-savings of $15 per case of PID prevented. Universal testing prevented an additional 11 cases of PID at a cost of $226,400, or costing $800 per additional case of PID prevented over age-targeted screening. Universal treatment prevented an additional 32 cases of PID and cost $221,100, saving $167 per additional cases of PID prevented over universal screening. Over a 2-year period, universal treatment provided the highest cost-savings and prevented the most disease. CONCLUSION: Screening by age provided a cost-savings to the Army over a 1-year period. Other organizations accessing large cohorts of young women could also benefit, even in the short term, from implementation of an age-based chlamydial screening program. Universal testing or universal treatment may be warranted in which long-term societal goals, such as maximum reduction of PID, are relevant. PMID- 10534208 TI - Double-blind comparison of trovafloxacin and doxycycline in the treatment of uncomplicated Chlamydial urethritis and cervicitis. Trovafloxacin Chlamydial Urethritis/Cervicitis Study Group. AB - BACKGROUND: Chlamydia trachomatis is among the most common sexually transmitted bacteria worldwide. With excellent activity against C. trachomatis and Neisseria gonorrhoeae and prolonged elimination half-life allowing once-daily dosage, the fluoroquinolone trovafloxacin has potential advantages in the treatment of uncomplicated chlamydial infection. GOAL OF THIS STUDY: This study compared the efficacy of trovafloxacin with that of doxycycline for the treatment of uncomplicated chlamydial infection. STUDY DESIGN: In a double-blind, multicenter trial, trovafloxacin 200 mg was administered once daily for 5 days and doxycycline 100 mg was administered twice daily for 7 days to patients with uncomplicated chlamydial urethritis or cervicitis. Follow-up visits were conducted 10, 21, and 35 days after enrollment. RESULTS: Of the 970 patients (403 men, 567 women) observed, 511 were microbiologically evaluable and 360 were clinically evaluable. C. trachomatis eradication rates in the trovafloxacin and doxycycline groups were equivalent in women (95% and 97%, respectively), but not in men (89% and 99%). Similarly, rates of clinical success (cure plus improvement) demonstrated equivalence of trovafloxacin and doxycycline in women (96% and 94%), but not in men (94% and 100%). The most frequent treatment-related adverse events were dizziness, nausea, and headache in patients given trovafloxacin, and nausea, vomiting, and headache in patients given doxycycline. Treatment-related discontinuations were comparable between the drug groups. CONCLUSION: Trovafloxacin given once daily for 5 days was clinically and bacteriologically equivalent to doxycycline given twice daily for 7 days in women with uncomplicated chlamydial cervicitis. This equivalence was not demonstrated in men with uncomplicated chlamydial urethritis. PMID- 10534209 TI - Privatization of STD services in Tacoma, Washington: a quality review. AB - BACKGROUND AND OBJECTIVES: This study describes key features of the privatized model of sexually transmitted disease (STD) service provision in Tacoma, Washington. GOALS: To assess the operational characteristics and the quality of care provided in the privatized STD clinics. STUDY DESIGN: Key informant interviews and surveys were conducted. Medical records were reviewed and compared to a standardized STD clinic medical record. Client treatment was compared to the Centers for Disease Control and Prevention (CDC) 1993 STD Treatment Guidelines. RESULTS: In 1997, the contract clinics provided 3,275 publicly sponsored STD visits. The goals of multiple access sites, STD screening, appropriate therapy for laboratory-diagnosed, bacterial STD were achieved. However, medical records were frequently incomplete and Gram stains were not done. Significant deviations from CDC STD Treatment Guidelines were documented in the medical management of pelvic inflammatory disease (PID). CONCLUSIONS: Privatized STD services would be improved by use of a standardized medical record. Availability of a Gram stain and management of PID as recommended by the CDC would augment quality of care. PMID- 10534210 TI - Partner notification for chlamydial infections among private sector clinicians in Seattle-King County: a clinician and patient survey. AB - BACKGROUND & OBJECTIVES: To describe partner notification practices for chlamydial infections among private sector clinicians. STUDY DESIGN: Telephone interviews of clinicians and patients identified through public health case reports in Seattle-King County, August-October 1998. RESULTS: Clinicians reported advising 135 of 150 (90%) patients to notify their sex partners, but knew that all partners of only 26 (17%) patients received treatment. While 71 (57%) clinicians acknowledged ever providing medicine-to a patient to give to a partner, only 6 (4%) so treated a patient about whom they were interviewed. Most (87%) clinicians believed the health department should routinely contact all patients about partner notification. Almost all patients (72/76-95%) reported that their provider had advised them to notify their partners and 59 (78%) stated they did so. Most patients (11/17-65%) who did not notify all of their partners would have been willing to allow their clinician or the heath department to do it for them. CONCLUSION: Private sector clinicians and their patients are generally unaware of chlamydial partner notification outcomes but are receptive to expanded partner notification services. PMID- 10534211 TI - Websites and STD services. PMID- 10534213 TI - Health-related quality of life in primary care patients with gastroesophageal reflux disease. AB - OBJECTIVE: To describe the clinical characteristics and health-related quality of life of family medicine patients with clinically diagnosed gastroesophageal reflux disease (GERD). METHODS: The study involved the baseline assessment of 268 patients enrolled in a randomized clinical trial comparing treatments for GERD. The study was conducted in a five-center, university-based family practice in southeastern West Virginia. Patients with a clinical diagnosis of GERD and who had not received treatment in the past 30 days were eligible; pregnant and lactating women and patients with severe renal or hepatic insufficiency were excluded. RESULTS: Two hundred sixty-eight patients were included in the analysis. Mean +/- SD age was 44.9 +/- 14.1 years; 61.2% were women and 91.4% were white. Mean +/- SD body mass index was 30.3 +/- 6 kg/m2, and >15.3% of patients had no insurance. One hundred seventy-four (64.9%) patients were enrolled from nonurban primary care clinics. One hundred sixty-four patients (61.2%) were prescribed at least one medication prior to study enrollment (mean +/- SD 2.88 +/- 1.71; range 1-9). When adjusted for age, gender, comorbidity status, and rural status, severity of GERD was associated with decreased health related quality of life. GERD patients without comorbidity demonstrated decrements in health-related quality of life when compared with the US general population. When compared with another GERD population, the study patients reported fairly consistent GERD symptomatology and health-related quality of life. CONCLUSIONS: GERD symptom severity was associated with impaired health related quality of life in a predominantly rural primary care population. PMID- 10534212 TI - Evaluation of a computer-assisted antibiotic-dose monitor. AB - OBJECTIVE: To examine the effect of a computer-assisted antibiotic-dose monitor used to reduce the number of days that patients receive excessive dosages of antibiotics and the number of adverse drug events (ADEs) secondary to antibiotics. DESIGN: Descriptive epidemiologic study of a two-year preintervention period and one-year intervention period. SETTING: The LDS Hospital, a tertiary care center in Salt Lake City, UT. PATIENTS: All patients aged > or = 18 years, admitted to LDS Hospital from April 1, 1993, to March 31, 1996, who received at least one of five targeted antibiotics (vancomycin, gentamicin, imipenem, cefazolin, cefuroxime), who had a serum creatinine or a urine creatinine clearance test result before antibiotic therapy, and who were never admitted or transferred to the shock/trauma/respiratory intensive care unit. METHODS: Each morning during the 12-month intervention period, the antibiotic-dose monitor checked the renal function of all patients who were receiving any of the five antibiotics. Pharmacists received a computer listing of patients who may have been receiving excessive dosages. The antibiotic-dose monitor suggested an alternate dosage and a pharmacist contacted the patient's physician if the suggested change in the dosage was appropriate. RESULTS: During the intervention period, 4483 patients received at least one of the five study antibiotics and 1974 (44%) were identified as receiving an excessive dosage, compared with 4494 (50%) of 8901 patients during the preintervention period (p < 0.001). The patients receiving excessive dosages received an excessive dosage for an average of 2.9 days during the intervention period, compared with 4.7 days (p < 0.001) during the preintervention period. In addition, these same patients during the intervention period received fewer doses of antibiotics (10.9 vs. 13.4; p < 0.001), fewer grams of antibiotics (10.4 vs. 12.0; p < 0.02), at less cost ($98 vs. $128; p < 0.004) than the patients during the preintervention period. Moreover, there were 14 ADEs (0.3%) secondary to the five study antibiotics during the intervention period, compared with 82 (0.9%; p < 0.001) for the two-year preintervention period. The study also found that significantly more patients identified as receiving excessive dosages had experienced decreases in renal function, compared with patients who were not identified as receiving excessive dosages (25% vs. 12% during preintervention period and 23% vs. 16% during intervention period; p < 0.001). CONCLUSIONS: Many patients experience decreases in renal function after antibiotics are ordered. The use of the computer-assisted antibiotic-dose monitor appears to be a promising method to help reduce the excessive use and cost of antibiotic therapy and reduce the number of ADEs secondary to antibiotics. PMID- 10534214 TI - Factors influencing the incidence of lamotrigine-related skin rash. AB - OBJECTIVE: To determine the incidences of serious and nonserious lamotrigine related rash, determine the risk factors for lamotrigine-related rash, and evaluate the impact on the incidence of rash of the manufacturer's recommendation to reduce the starting dose of lamotrigine. METHODS: This was a retrospective case record survey at five tertiary referral epilepsy centers in the UK. The risk factors for lamotrigine-related rash were identified by logistic regression. The independent factors tested were gender, age, epilepsy type, concurrent medication, and starting dose of lamotrigine. The incidences of rash before and after the recommendation of reduction in starting dose were compared by chi2 analysis. RESULTS: A total of 1050 patients were included. The incidences of serious and nonserious rash were 1.1% (95% CI 0.5% to 1.8%) and 7% (95% CI 5.5% to 8.6%), respectively. Females were at higher risk of developing rash than were males, with a relative risk of 1.8 (95% CI 1.2 to 2.8). The starting dose of lamotrigine was reduced in response to the manufacturer's recommendation, and there was a significant reduction (p = 0.045) in the incidence of serious rash, from 1.5% (12/805) to 0% (0/245). However, there was no reduction in the overall incidence of lamotrigine-related rash, with 63/805 (8%) before and 23/245 (9%) after the recommendation. CONCLUSIONS: Failure to detect a reduction in the incidence of lamotrigine-related rash since the new (reduced) recommended starting dose of lamotrigine may arise from failure to reduce the starting dose below a critical threshold level, incomplete compliance with current recommendations, or insufficient sample size. The results of this and other studies show that the starting dose of lamotrigine is a significant factor affecting the incidence of rash; furthermore, this study also shows that significant reduction in the incidence of serious rash can be achieved by reducing the starting dose. Therefore, clinicians should not deviate from the recommendations. PMID- 10534215 TI - Life-threatening reaction to vancomycin given for noninfectious fever. AB - OBJECTIVE: To report a case of vancomycin-induced anaphylaxis (or anaphylactoid reaction) in a patient with a fever of unrecognized noninfectious origin. CASE SUMMARY: An 83-year-old white man, who was a patient of the Veterans Affairs Medical Center, developed a serious anaphylactic (or anaphylactoid) reaction while receiving intravenous vancomycin as empiric therapy for a nosocomial fever of unknown origin. The fever was subsequently proved to have been due to acute polyarticular gout rather than an infection. DISCUSSION: This patient developed respiratory distress and an increased serum troponin concentration, suggestive of a myocardial enzymatic leak as a result of vancomycin therapy. Vancomycin was given before the noninfectious cause of his fever was recognized. CONCLUSIONS: Even with cautious slow infusion, intravenous vancomycin can precipitate life threatening infusion-related reactions in some patients. Because of this, and to reduce selective pressure for vancomycin resistance, sources of fever that do not require treatment with vancomycin should be diligently investigated prior to the institution of empiric vancomycin therapy in febrile patients, particularly when the past medical history is suggestive of an alternative diagnosis. PMID- 10534216 TI - Haloperidol-induced torsade de pointes. AB - OBJECTIVE: To report a case of torsade de pointes related to the administration of high-dose intravenous haloperidol for the treatment of severe agitation. CASE SUMMARY: Reports in the literature of intravenous haloperidol-induced torsade de pointes are rare. We describe the case of a 41-year-old white woman with no predisposing factors who developed torsade de pointes 55 minutes after a dose of intravenous haloperidol 80 mg (total dosage 915 mg over 7 d). The results of the electrocardiogram were consistent with torsade de pointes and showed a prolonged QTc interval of 610 milliseconds. Intravenous magnesium sulfate 2 g/100 mL NaCl 0.9% was administered, which controlled the arrhythmia. The patient received one additional 80-mg haloperidol dose six hours after the arrhythmia-triggering dose, without reoccurrence of torsade de pointes. Haloperidol was then discontinued, and the patient had no further arrhythmias. CONCLUSIONS: Our case report and others from the literature suggest that intravenous haloperidol administration may prolong QT intervals in some patients, precipitating the potentially life threatening arrhythmia torsade de pointes. Clinicians should be aware of haloperidol's potential to induce torsade de pointes, since it is used regularly for agitation and delirium in the critical care arena. PMID- 10534217 TI - Chronic constrictive pericarditis induced by long-term bromocriptine therapy: report of two cases. AB - OBJECTIVE: To report two cases of chronic constrictive pericarditis that appear to be related to the intake of bromocriptine for Parkinson's disease. CASE SUMMARY: Two white men (aged 63 and 69 y) were treated with bromocriptine for four (40 mg/d) and two years (30 mg/d), respectively, with a cumulative dose intake of 58.4 and 21.9 g, respectively. The patients experienced dyspnea with bilateral lower-limb edema and pleural effusion, suggesting right cardiac dysfunction. Echocardiography, computed tomography, and cardiac catheterization results were compatible with a diagnosis of constrictive pericarditis, so pericardectomy was performed on both patients. The anatomic pathology examination showed a fibrous pericardium; cultures were sterile. In the first case, pleural effusion recurred seven months after the pericarditis; bromocriptine was suspected and treatment was discontinued. In the second case, just prior to the pericardectomy, an episode of mental confusion occurred and prompted the cessation of bromocriptine therapy. DISCUSSION: To the best of our knowledge, only one case of constrictive pericarditis induced by bromocriptine therapy has previously been described in the literature. CONCLUSIONS: Our cases call attention to a possible association between bromocriptine use in patients who have Parkinson's disease and constrictive pericarditis. PMID- 10534218 TI - Treatment of Wegener's granulomatosis with immune globulin: CNS involvement in an adolescent female. AB - OBJECTIVE: To describe the use of intravenous immune globulin (IVIG) to treat Wegener's granulomatosis (WG) in an adolescent female with an abnormal magnetic resonance imaging (MRI) scan and electroencephalogram (EEG), as well as central nervous system involvement manifesting as generalized seizures. CASE SUMMARY: A 15-year-old white girl diagnosed with WG and receiving prednisone and cyclophosphamide was admitted with new-onset tonic-clonic seizures. The patient received phenobarbital and phenytoin to control seizures and was receiving cyclophosphamide and corticosteroids for WG. She developed cyclophosphamide induced cystitis and was started on a four-day therapeutic course of IVIG following the discontinuation of cyclophosphamide. After 16 days of hospitalization, repeat EEG and MRI were within normal limits, and laboratory and clinical improvement was evident in at least nine of the affected organ systems including pulmonary, hematologic, renal, ocular, cutaneous, musculoskeletal, central nervous system, gastrointestinal, and genitourinary. The patient was discharged with clinical involvement of WG documented in two organ systems, hematologic and renal. DISCUSSION: WG is a form of vasculitis believed to develop due to an autoimmune disorder. The diagnosis is based on radiographic and histopathologic findings, as well as the presence of elevated antineutrophil cytoplasmic antibodies and a suggestive clinical presentation. The presentation is widely variable and is most commonly associated with upper-airway involvement such as sinusitis, cough, pulmonary infiltrates, and cavitary nodules. Renal involvement signifies generalized disease. Conventional treatment for WG includes cyclophosphamide and prednisone. Little information is available describing therapeutic alternatives. Cytotoxicity related to immunosuppressant regimens limits continuous treatment and may necessitate the use of alternative agents. CONCLUSIONS: This case describes the use of IVIG in an adolescent patient presenting with WG as a generalized, active disease with neurologic complications. IVIG may be useful in generalized, active WG complicated by intolerance to cyclophosphamide and seizures, but further study is necessary to define its role. PMID- 10534219 TI - Oxacillin-induced tissue necrosis. AB - OBJECTIVE: To report a case of oxacillin-induced tissue necrosis in which recommended concentration guidelines for dilution and administration were used. Oxacillin concentration data, potential risk factors, and treatment options for extravasation injuries are also briefly reviewed. CASE SUMMARY: Oxacillin was infused peripherally by infusion pump in a 79-year-old white woman as prophylactic antibiotic coverage for permanent pacemaker placement. Oxacillin extravasation occurred after the second postoperative dose. A dime-sized area of necrosis was noted at the heparin-lock insertion site. DISCUSSION: Only one case of oxacillin-induced necrosis has been reported. The degree of damage and concentration of drug used were not specifically described. Concentration may play a role in the appearance or absence of tissue damage after an antibiotic extravasation and should be taken into consideration when evaluating a drug's tissue toxicity potential. CONCLUSIONS: The potential exists for oxacillin 50 mg/mL to cause tissue damage in humans if an extravasation occurs. This reaction may be avoided with use of a less-concentrated preparation, avoidance of infusion pump administration, and identification of high-risk patients. PMID- 10534220 TI - Prevention of suboptimal beta-blocker treatment in patients with myocardial infarction. AB - OBJECTIVE: To review the published data and clinical guidelines on the use of beta-blockers in myocardial infarctions (MIs) and contrast that with actual clinical practice. DATA SOURCES: A MEDLINE search (January 1970-June 1999) was performed to identify all relevant articles. References from these articles were also evaluated for review if deemed important. DATA SYNTHESIS: Intravenous and oral beta-blockers have been proven to improve outcomes in patients with MIs in numerous clinical trials. In current clinical practice, only 15% of MI patients receive intravenous beta-blockers and long-term beta-blocker therapy is used in <40% of patients without contraindications. However, they could be safely administered to 40% and 70% of these patients, respectively. Furthermore, most of these patients are receiving doses far below those found beneficial in clinical trials. Many of the real and perceived contraindications to beta-blockers are reviewed to allow the practitioner to identify patients who are incorrectly excluded from beta-blocker therapy. Also discussed are special clinical situations in which the benefits observed during clinical trials may not apply. CONCLUSIONS: Beta-blockers are valuable drugs in the treatment of peri- and post MI. In clinical practice, most patients are not treated or are inadequately treated with beta-blockers. Pharmacists should ensure that such patients actually have an absolute contraindication or unusual situation where therapy is not firmly indicated. Patients without absolute contraindications warrant titration to specific target doses or a target heart rate of 55-60 beats/min. PMID- 10534221 TI - Tolterodine use for symptoms of overactive bladder. AB - OBJECTIVE: To describe the pharmacology, pharmacokinetics, clinical efficacy, and safety of tolterodine for the treatment of overactive bladder. DATA SOURCES: Published articles and abstracts were identified from a MEDLINE search (January 1980-October 1998) using the terms tolterodine, PNU-200583E, urge incontinence, overactive bladder, detrusor instability, detrusor overactivity, and antimuscarinic. Pertinent articles written in English were considered for review. Additional articles were identified from the bibliographies of retrieved articles. Data from the Food and Drug Administration-approved product labeling and the manufacturer were also used in the absence of published data. STUDY SELECTION AND DATA EXTRACTION: Clinical studies of tolterodine involving human subjects were evaluated. DATA SYNTHESIS: Tolterodine is a competitive muscarinic receptor antagonist with relative functional selectivity for bladder muscarinic receptors. It is metabolized in the liver by CYP2D6 to an active metabolite (DD 01), which is partially responsible for its pharmacologic activity. Those who are genetically devoid of CYP2D6 will have higher concentrations of the parent compound and virtually undetectable concentrations of DD 01; however, the clinical efficacy does not appear to be altered. In dosages of 2 mg twice daily, tolterodine has shown consistent reductions in the number of micturitions per 24 hours and less consistently decreased incontinence episodes in patients with detrusor overactivity. The functional selectivity of tolterodine for bladder muscarinic receptors results in fewer systemic adverse effects, such as dry mouth, than occur with comparable nonselective antimuscarinic agents. CONCLUSIONS: Clinical studies have shown that the effectiveness of tolterodine for symptoms of overactive bladder is similar to that of oxybutynin. The adverse effect profiles of tolterodine and oxybutynin are similar; however, comparative clinical trials have shown significantly fewer patients taking tolterodine require dosage reductions or discontinue therapy due to antimuscarinic adverse effects such as dry mouth. Although more costly than oxybutynin, tolterodine represents a modest improvement over oxybutynin with respect to adverse effect profile, which may allow more patients with incontinence to tolerate therapeutic doses. Further research is necessary to determine whether tolterodine has clinical advantages over similar agents in patients with other muscarinic adverse effects, such as constipation or cognitive impairment. PMID- 10534222 TI - Micronized fenofibrate: a new fibric acid hypolipidemic agent. AB - OBJECTIVE: To review the efficacy and safety of fenofibrate in the management of hyperlipidemias. DATA SOURCES: A MEDLINE search (1974-October 1998), Current Contents search, additional references from article bibliographies, and the package insert from the manufacturer were used to identify data for evaluation. Studies evaluating fenofibrate (peer-reviewed publications, package insert data) were considered for inclusion. Abstracts and data on file with the manufacturer were not considered for inclusion. STUDY SELECTION: English-language literature was reviewed to evaluate the pharmacology, pharmacokinetics, clinical use, and tolerability of fenofibrate. Data from animals and in vitro systems were included only when necessary to explain the drug's pharmacology. DATA SYNTHESIS: Micronized fenofibrate is a fibric acid derivative approved by the Food and Drug Administration (FDA) in February 1998 for the treatment of types IV and V hyperlipidemia. Data from the peer-reviewed literature also support the use of fenofibrate in types IIa, IIb, and III hyperlipidemias. Micronized fenofibrate 67 201 mg/d is useful as monotherapy or as an adjunct to other hypolipidemics and dietary therapy. In placebo-controlled clinical trials, regular formulation fenofibrate 300-400 mg/d lowered serum triglyceride (TG) concentrations by 24 55%, total cholesterol by 9-25%, low-density lipoprotein cholesterol (LDL-C) concentrations by 6-35%, and raised high-density lipoprotein cholesterol (HDL-C) concentrations by 8-38%. Few comparative data exist regarding fenofibrate versus clofibrate and gemfibrozil. In noncomparative and comparative clinical trials, fenofibrate appeared to be well tolerated. The most common causally related adverse events were digestive, musculoskeletal, and dermatologic in nature. Concurrent use of fenofibrate and a hydroxymethylglutaryl-coenzyme A inhibitor may increase the risk of myopathy and/or rhabdomyolysis, although recent data suggest that concurrent use of fenofibrate with low-dose simvastatin or pravastatin is safe. Fenofibrate may enhance the effect of oral anticoagulants. CONCLUSIONS: Fenofibrate reduces serum TG, total cholesterol, and LDL-C, and raises HDL-C to clinically relevant degrees. Its spectrum of activity appears to exceed that recommended for types IV and V hyperlipidemia to encompass types IIa, IIb, and III hyperlipidemias as well. To this extent, it may be considered a broader-spectrum fibrate than is indicated by its FDA approval. Adverse effects of fenofibrate appear to be similar to those of other fibrates and require routine monitoring (clinical, liver function). Long-term safety data are readily available from drug registries in many countries where the product has been available for nearly two decades. Cost-effectiveness studies comparing fenofibrate with other hypolipidemics demonstrate benefits of fenofibrate over simvastatin in types IIa and IIb hyperlipidemia. The need for dosage titration of the micronized preparation from 67 mg/d upward to a final dose of 200 mg/d is also not supported by peer-reviewed literature (except in the case of renal impairment). Although preliminary data on plaque regression are encouraging, published clinical studies evaluating the impact of fenofibrate on cardiovascular morbidity and mortality are awaited. Micronized fenofibrate is worthy of formulary inclusion. PMID- 10534223 TI - Role of lamivudine in the treatment of chronic hepatitis B virus infection. AB - OBJECTIVE: To examine the evidence regarding the therapeutic effectiveness of lamivudine in the treatment of chronic hepatitis B virus (HBV) infection in immunocompetent patients. DATA SOURCES: Using chronic hepatitis B and lamivudine as MeSH headings, MEDLINE was searched from 1966 to September 1998 for all published randomized controlled trials evaluating lamivudine in chronic HBV infection. Relevant articles from selected bibliographies were also retrieved. STUDY SELECTION: Only randomized, single- and double-blind trials in human HBV carriers published in the English language were included. DATA SYNTHESIS: Evidence from the controlled trials suggests that lamivudine has a therapeutic effect in suppressing HBV replication in immunocompetent patients. Lamivudine 100 mg/d appears to suppress HBV replication in as many as 97% of patients within two weeks after the initiation of therapy and is capable of suppressing histologic damages. However, viral suppression is effective only during the therapy; on discontinuation of lamivudine therapy, most patients return to the pretreatment condition. Viral resistance to lamivudine has been observed. Most patients with chronic HBV infection appear to tolerate 100 mg/d of lamivudine therapy. CONCLUSIONS: Evidence has shown that oral lamivudine 100 mg/d will produce rapid and significant suppression of viral replication in immunocompetent patients with chronic HBV infections. Treatment periods up to one year have been effective and well tolerated. The suppression of viral replication may not be sustained after cessation of lamivudine therapy, and very few patients have complete elimination of HBV during therapy. Therefore, long treatment periods may be necessary. Efficacy and tolerability of treatment beyond one year need to be investigated. Resistance to lamivudine has been reported in patients receiving therapy. A combination anti-HBV regimen using lamivudine and other agents with different mechanisms of action should be investigated to maximize the elimination of the viral infection while minimizing or preventing damage to the liver cells and tissues and the development of viral resistance. PMID- 10534224 TI - The use of valproic acid in HIV-positive patients. AB - OBJECTIVE: To review the use of valproic acid in HIV-positive patients. DATA SOURCES: Clinical literature was accessed through a MEDLINE search (January 1966 November 1998). Key search terms included HIV, AIDS, seizures, valproic acid, and glutathione. DATA SYNTHESIS: Patients with HIV often develop neurologic manifestations; therefore, valproic acid may be considered in the management of this population. It has been demonstrated that valproate may increase viral burden by potentiating replication. An evaluation of studies addressing the use of valproic acid in HIV-positive patients was conducted. CONCLUSIONS: The potential for valproate-induced increases in viral replication exists. Although further studies are warranted, clinicians should exercise caution when using valproate in HIV-positive patients. PMID- 10534225 TI - Role of anagrelide in the treatment of thrombocytosis. AB - OBJECTIVE: To review the role of anagrelide in the management of essential thrombocythemia. DATA SOURCES: A MEDLINE search (January 1966-August 1998) was performed using the key terms anagrelide, thrombocytosis, and essential thrombocythemia. In addition, the package insert, product monograph, and patient information pamphlets were reviewed. DATA SYNTHESIS: Recently, there has been a trend toward the use of anagrelide in the management of thrombocythemia. Anagrelide lacks leukemogenic and mutagenic potential and possesses a more favorable adverse effect profile compared with other therapeutic agents. At recommended doses, anagrelide induces thrombocytopenia in thrombocythemic patients with a concomitant reduction in the incidence of disease-related symptoms. CONCLUSIONS: Although the treatment of choice for thrombocythemia is still an area of debate, anagrelide has distinct advantages over alternative therapies. Anagrelide represents an important therapeutic option in the treatment of patients with thrombocythemia. PMID- 10534226 TI - Trovafloxacin-associated exfoliative dermatitis in a homecare patient. PMID- 10534227 TI - Inadvertent epidural gentamicin administration. PMID- 10534228 TI - Pyridoxine ineffective in isoniazid-induced psychosis. PMID- 10534229 TI - Effects of itraconazole on tacrolimus blood concentrations in a renal transplant recipient. PMID- 10534230 TI - Cost-effectiveness of macrolides in lower respiratory tract infections. PMID- 10534231 TI - Severe thrombocytopenia associated with amlodipine treatment. PMID- 10534232 TI - The role of nuclear cardiology in clinical decision making. AB - This review suggests that the field of nuclear cardiology is alive, well, and thriving, providing relevant information that aids in everyday clinical decision making for nuclear medicine and referring physicians alike. Despite the competition from other modalities, the clinically appropriate applications of nuclear cardiology techniques are likely to increase. The foundation of this optimism is based on the vast amount of data documenting cost-effective clinical applications for diagnosis, risk stratification, and assessing therapy in both chronic and acute coronary artery disease (CAD), the powerful objective quantitative analysis of perfusion and function provided by the technique, and the increasing general availability of the approach. PMID- 10534233 TI - Pharmacological stress testing. AB - Pharmacological stress in conjunction with radionuclide myocardial perfusion imaging has become a widely used noninvasive method of assessing patients with known or suspected coronary artery disease. In the United States, over one third of perfusion imaging studies are performed with pharmacological stress. Pharmacological stress agents fall into two categories: coronary vasodilating agents such as dipyridamole and adenosine, and cardiac positive inotropic agents such as dobutamine and arbutamine. For both, in the presence of coronary artery disease (CAD), perfusion image abnormalities result from heterogeneity of coronary blood flow reserve. Vasodilating agents work directly on the coronary vessels to increase blood flow, whereas inotropic agents work indirectly by increasing myocardial work load, which then leads to an increase in coronary blood flow. Both classes of agents have high accuracies for diagnosing coronary artery disease, and they have excellent safety records with acceptably low occurrences of side effects. For dipyridamole planar thallium imaging, pooled analysis yields a sensitivity of 85% and a specificity of 87% for diagnosis of coronary disease, but there is a large variation in reported values depending on various factors, such as the extent of postcatheterization referral bias, the type of imaging (planar versus single photon emission computed tomography [SPECT]), the types of patients being studied (single versus multivessel disease, men versus women), and the imaging agent used (thallium versus one of the technetium-based agents). Diagnostic accuracies for adenosine are similar to those of dipyridamole, with reported overall sensitivities ranging from 83% to 97%, and specificities ranging from 38% to 94%. For dobutamine, pooled analyses yield a sensitivity of 82% and a specificity of 75%. There is some concern that dobutamine may interfere with uptake of technetium-99m sestamibi, lowering the sensitivity for detection of disease, and thus the vasdodilating agents are generally preferred. Pharmacological stress testing has high clinical use for risk stratifying patients with known or suspected CAD, in patients after myocardial infarction, and in patients needing noncardiac surgery. Vasodilating agents are particularly advantageous in assessing post-myocardial infarction patients, allowing testing as soon as 2 days after the event. Like patients undergoing exercise stress testing, patients with normal perfusion images by pharmacological stress have a <1% annual incidence of cardiac events. The likelihood of an event increases with the extent and severity of perfusion abnormalities. However, it is important to consider clinical variables when using perfusion imaging for risk stratification, particularly in the presurgery patients. As with exercise testing, adjunct markers such as ST segment depression during testing, lung uptake of radiotracer (if thallium is used), and ventricular cavity dilatation add additional prognostic information to that available from the perfusion images alone. The aim of current research is to find better agents that are easier to use and that have fewer side effects. MRE-0470 is an experimental vasodilating agent that is more receptor selective than adenosine and promises a lower incidence of hypotension. Arbutamine more closely simulates exercise than dobutamine, and it can be administered by a closed-loop computerized delivery device. Work is also underway to look at novel uses of pharmacological stress agents, such as acquiring gated SPECT images during dobutamine infusion to enhance detection of myocardial viability. With increasing use of noninvasive testing in elderly patients and in patients with comorbidities that preclude adequate exercise, pharmacological stress testing has become an indispensable tool for radionuclide myocardial perfusion imaging studies. A good understanding of pharmacological stress testing is essential for performing high quality nuclear cardiology PMID- 10534234 TI - Stress myocardial perfusion imaging versus echocardiography for the diagnosis and risk stratification of patients with known or suspected coronary artery disease. AB - Stress perfusion imaging and stress echocardiography (ECHO) are both very useful for assessment of diagnosis and risk stratification of patients with coronary artery disease (CAD). Both techniques have been well validated during exercise and inotropic stress, but coronary vasodilation stress is better used in combination with perfusion imaging. The overall sensitivity for detection of CAD is slightly higher by single photon emission computed tomography (SPECT) than by two-dimensional (2D) ECHO during all stress modalities, whereas the specificity is slightly higher by ECHO, although the differences in general are not statistically significant. SPECT, however, appears to be superior to ECHO in the diagnosis of isolated circumflex stenosis, as well as for the correct identification of multivessel CAD. A substantially greater amount of information is available regarding risk stratification with SPECT than with 2D ECHO. Although the data suggest that both techniques are very useful for risk stratification of patients with stable CAD, after myocardial infarction, and for preoperative risk stratification, the risk for cardiac events is lower in the presence of a normal stress SPECT study than of a normal stress ECHO. PMID- 10534235 TI - The potential for myocardial imaging with hypoxia markers. AB - Direct "hot spot" imaging of myocardial tissue hypoxia is potentially of great clinical importance because available noninvasive approaches for the detection of myocardial ischemia have generally been based on the detection of flow heterogeneity or identification of regional alterations of myocardial metabolism. These existing approaches provide only an indirect assessment of regional myocardial ischemia, and may be affected by either sympathetic activation or substrate availability. The assessment of tissue oxygenation with hypoxic compounds may be the best indicator of the balance of flow and oxygen consumption. These compounds may provide a means of identifying dysfunctional chronically ischemic but viable "hibernating" myocardium and find a critical place in the assessment of angiogenesis. Nitroimidazole compounds hold promise for positive imaging of hypoxia in the heart. However, refinement of these compounds is needed to improve target specificity. The potential of technetium 99m (Tc99m) complexes derived from removal of the nitroimidazole moiety from a nitroimidazole-containing ligand is interesting and warrants further investigation. Experimental studies support the possibility of identifying myocardial hypoxia with the positron-emitting compound F18-fluoromisonidazole noninvasively. The potential of a Tc99m labeled nitroimidazole for positive imaging of myocardial ischemia is tremendous because single-photon imaging is more widely available. The true clinical potential of these nitroimidazole compounds can only be defined with future experimental and clinical studies. Ideally, these studies should include comparisons of tracer uptake with independent measures of regional ischemia or measures of oxygen tension, potentially using magnetic resonance imaging. PMID- 10534236 TI - Current status of radionuclide tracer imaging of thrombi and atheroma. AB - The imaging of thrombi and atherosclerotic plaques has great potential for decision making in the management of patients with all types of disease within the circulatory system. This importance is owing to the developments showing that areas of moderate stenosis with underlying atheroma are physiologically reactive and capable of causing reversible clinical symptoms that can progress to irreversible end-organ damage if not effectively treated. Identifying and quantifying areas of smaller vulnerable plaque and areas of acute thrombosis will assist in identification of patients at risk and help determine when and how to treat these patients. Initial efforts in this area used nonspecific constituents of thrombi and atheroma that were radiolabeled using long-lived isotopes, which had high background activity that required imaging over 48 to 72 hours. Newer approaches have focused on the use of small antibody fragments or small peptides, so-called molecular recognition units, that specifically target antigens present only in areas of thrombosis or active atherogenesis. These compounds are labeled Technetium-99 m (99mTc) and provide excellent images. Efforts to image thrombi have been directed at the IIB/IIIA receptor, which is present in low concentration on the cell membrane of circulating quiescent platelets, but on stimulation and active thrombosis, more than 80,000 potential binding sites per platelet appear. One such peptide has been clinically approved for imaging of deep vein thrombophlebitis. Parallel efforts are being made for imaging areas of active atherogenesis by targeting smooth muscle cells and other constituents unique for vulnerable plaques. Efforts in developing these modalities are important to expand the applications to new areas in nuclear cardiology. PMID- 10534237 TI - Matrix metalloproteinases in multiple sclerosis: targets of therapy or markers of injury? PMID- 10534238 TI - Candidate genes and Parkinson's disease: where to next? PMID- 10534239 TI - New developments in the neurobiology of the tuberous sclerosis complex. AB - OBJECTIVE: To outline recent developments in the neurobiology of the tuberous sclerosis complex (TSC). BACKGROUND: TSC may be associated with neuropsychiatric disorders including epilepsy, mental retardation, and autism. The uncontrolled growth of subependymal giant cell astrocytomas may lead to hydrocephalus and death. The recent identification of mutations in two genes (TSC1 and TSC2) that cause TSC has led to rapid progress in understanding the molecular and cellular pathogenesis of this disorder. How distinct mutations lead to the varied clinical phenotype of TSC is under intense investigation. RESULTS: We report the recent diagnostic criteria for TSC and provide an overview of the molecular genetics, molecular pathophysiology, and neuropathology of TSC. Important diagnostic criteria for TSC include facial angiofibromas, ungual fibromas, retinal hamartomas, and cortical tubers. Both familial and sporadic TSC cases occur. Approximately 50% of TSC families show genetic linkage to TSC1 and 50% to TSC2. Among sporadic TSC cases, mutations in TSC2 are more frequent and often accompanied by more severe neurologic deficits. Multiple mutational subtypes have been identified in the TSC1 and TSC2 genes. The TSC1 (chromosome 9) and TSC2 (chromosome 16) genes encode distinct proteins, hamartin and tuberin, respectively, which are widely expressed in the brain and may interact as part of a cascade pathway that modulates cellular differentiation, tumor suppression, and intracellular signaling. Tuberin has a GTPase activating protein-related domain that may contribute to a role in cell cycle passage and intracellular vesicular trafficking. CONCLUSION: Identification of tuberous sclerosis complex (TSC) gene mutations has fostered understanding of how brain lesions in TSC are formed. Further characterization of the roles of hamartin and tuberin will provide potential therapeutic avenues to treat seizures, mental retardation, and tumor growth in TSC. PMID- 10534240 TI - Altered brain activation in cognitively intact individuals at high risk for Alzheimer's disease. AB - OBJECTIVE: To determine whether brain function is altered in cognitively normal individuals at high risk for AD several years before the typical age at onset for this illness. BACKGROUND: Neuropathologic alterations in AD precede cognitive impairment by several years. It is unknown whether functional alterations in neural circuitry accompany these neuropathologic changes, and if so, whether they may be detectable before onset of symptoms. METHODS: We used functional MRI to compare cortical activation between two groups of cognitively normal women differing only in their risk for developing AD. Visual naming and letter fluency tasks were used to activate brain areas subserving object and face recognition, previously described sites of hypometabolism and neuropathologic alteration in AD. The risk groups differed in family history of AD and apolipoprotein E allele status, but were matched in age, education, and measures of cognitive performance. Average age of the study participants was 52 years. RESULTS: The regional patterns of brain activation were similar between groups. However, the high risk group showed areas of significantly reduced activation in the mid- and posterior inferotemporal regions bilaterally during both tasks despite identical naming and letter fluency performance. CONCLUSIONS: Cognitively normal individuals at high risk for AD demonstrate decreased brain activation in key areas engaged during naming and fluency tasks. Decreased activation in the high risk group may be a consequence of the presence of subclinical neuropathology in the inferotemporal region or in the inputs to that region. If so, these findings provide evidence of a window of opportunity for disease-modifying treatment before the onset of symptomatic AD. PMID- 10534241 TI - Serum MMP-9 and TIMP-1 levels are related to MRI activity in relapsing multiple sclerosis. AB - OBJECTIVE: To 1) compare monthly serum levels of matrix metalloproteinase-9 (MMP 9) and tissue inhibitor of MMP-type 1 (TIMP-1) in patients with relapsing remitting MS (RRMS) versus healthy controls and 2) determine the relationship among monthly serum levels of MMP-9 and TIMP-1 and MRI activity. BACKGROUND: Activated T-cells and macrophages secrete MMPs that may facilitate their migration across vascular subendothelial basement membranes into the CNS. The serum concentration of MMP-9 is reported to be higher in patients with RRMS than healthy controls. METHODS: Monthly evaluations including gadolinium-enhanced (Gd+) brain MRI and measures of serum MMP-9 and TIMP-1 were performed for up to 15 months in 24 patients with RRMS and for up to 4 months in 10 controls. RESULTS: Serum MMP-9 but not TIMP-1 levels are elevated in RRMS patients compared to healthy controls (p = 0.025, p = 0.61). In a univariate analysis, high MMP-9 and low TIMP-1 levels precede appearance of new Gd+ lesions (respectively; odds ratio = 3.3, p = 0.008; odds ratio = 2.2, p = 0.086). In a multivariate analysis, in comparison to months when MMP-9 is low and TIMP-1 high, MRI scans obtained the month following high MMP-9 and low TIMP-1 serum concentrations are more likely to report new Gd+ lesions (p = 0.0006, odds ratio = 21.5). CONCLUSION: An increase in the activity of matrix metalloproteinase-9 (MMP-9) relative to tissue inhibitor of MMP-type 1 (TIMP-1) may be related to formation of new MS lesions, suggesting that serum levels of MMP-9 and TIMP-1 may be surrogate markers of disease activity in relapsing-remitting MS. PMID- 10534242 TI - Changes of serum sICAM-1 and MMP-9 induced by rIFNbeta-1b treatment in relapsing remitting MS. AB - OBJECTIVE: To correlate changes in serum levels of intercellular adhesion molecule-1 (sICAM-1) and matrix metalloproteinases (MMP) with clinical and MRI evidence of disease activity in MS patients receiving treatment with interferon beta (rIFNbeta)-1b. BACKGROUND: rIFNbeta reduces the frequency of gadolinium enhancing (Gd+) MRI in relapsing-remitting MS (RRMS). Its mechanism of action on improving the integrity of the blood-brain barrier remains unclear. METHODS: sICAM-1 and MMP-9 and MMP-2 serum levels were longitudinally (24 months) investigated (ELISA; zymography) in correlation with the modifications of the integrated area under the curve of Expanded Disability Status Scale scores normalized to entry baseline (deltaEDSS AUC) and of GD+ MRI scans, and with neutralizing antibodies (NAB) to rIFNbeta-1b production (MxA) in 36 RRMS patients. RESULTS: During the first 12 months of treatment, levels of sICAM-1 increased and MMP-9 decreased significantly. After 12 months, levels returned toward baseline. Levels of sICAM-1 and MMP-9 were significantly negatively correlated. MMP-2 levels did not change significantly during the same period. During the second semester of the study, deltaEDSS AUC was significantly reduced. The percentage of patients with Gd+ MRI decreased significantly in the first (33%), second (29%), third (20%), and fourth (28%) semesters of treatment compared to baseline (62%). The NAB+ patients (14%) tended to have lower sICAM-1 levels at the ninth month; a higher MMP-9 activity at the sixth, 12th, and 18th months; and a greater deltaEDSS AUC in the third semester of treatment in comparison with the NAB- patients. CONCLUSIONS: These results show that rIFNbeta 1b therapy increases sICAM-1 serum levels and reduces serum MMP-9 activity. PMID- 10534243 TI - Shedding of TNF receptors in multiple sclerosis patients. AB - OBJECTIVE: To evaluate the rate of shedding of tumor necrosis factor (TNF) receptors (TNFRs) in MS patients. BACKGROUND: It was previously suggested that TNF might play a significant role in the immunopathologic mechanism of MS. TNF mediates its biologic effects by interacting with two distinct receptors: TNFR p55 and TNFR-p75. Both of these receptors exist in soluble and membrane-bound forms. Soluble receptors have been shown to influence TNF activity in vitro and in vivo and maintain balance between active, free TNF and inactive form of this cytokine bound to its soluble receptors. METHODS: In the current study, the authors measured shedding of TNFRs from cell surface of peripheral blood mononuclear cells, peripheral blood lymphocytes, and monocytes in three groups of MS patients: relapsing-remitting in relapse, relapsing-remitting in remission, and chronic progressive. RESULTS: The authors observed a significant distortion in generation of both soluble TNF receptors. Whereas the TNFR-p55 was shed at lower rate compared with healthy volunteers, the shedding of TNFR-p75 was significantly higher in MS patients. CONCLUSION: Disturbance in TNFR shedding might contribute to the distortion of a fine balance between circulating TNF and its natural inhibitors in MS. PMID- 10534244 TI - A study of five candidate genes in Parkinson's disease and related neurodegenerative disorders. European Study Group on Atypical Parkinsonism. AB - OBJECTIVE: To determine whether reported genetic association of polymorphisms in the CYP2D6, CYP1A1, N-acetyltransferase 2 (NAT2), DAT1, and glutathione s transferase M1 (GSTM1) genes with PD were evident in a population of 176 unrelated patients with sporadic PD and to extend these findings to other disease groups (familial PD [n = 30], ALS [n = 50], multiple system atrophy [n = 38], progressive supranuclear palsy [n = 35], and AD [n = 23]). METHODS: A combination of allele-specific PCR and analysis of restriction fragment length polymorphisms were performed. RESULTS: We genotyped 1,131 individuals. After matching each patient with a control subject by age, sex, ethnicity, and geographic origin, there was no association of any allele/genotype with any of the six disease groups. There was an increased frequency of NAT2 slow acetylators in the ALS group compared with controls (70% versus 50%; OR 2.33 [95% CI, 1.03 to 5.30]), but this was not significant after adjusting for multiple testing. CONCLUSIONS: This is one of the most extensive candidate gene studies performed in PD and the first time that some of these loci have been studied in multiple system atrophy and progressive supranuclear palsy. In contrast with previous studies, we found no role for these polymorphisms in the etiology of PD, ALS, multiple system atrophy, progressive supranuclear palsy, or AD. PMID- 10534245 TI - Mutational analysis of the tau gene in progressive supranuclear palsy. AB - OBJECTIVE: To identify genetic mutations in the coding regions and the splice donor sites of the tau gene on chromosome 17q in individuals with progressive supranuclear palsy (PSP). BACKGROUND: Several studies provide evidence for linkage disequilibrium between a PSP disease gene and allelic variants of the tau gene. However, a causative mutation has not been identified. METHODS: We designed a study to search for genetic mutations in 15 coding regions of the tau gene including the splice-donor sites in 22 patients with PSP by comparing the mobility shifts on single-strand conformation analysis with those of age-matched controls. Fragments with altered migration were sequenced directly and compared for differences in nucleotide composition. Restriction enzyme digests were used to confirm single base-pair substitutions. RESULTS: Significant differences in mobility shifts were found in exons 1, 4A, and 8 between affected individuals and age-matched controls. All individuals with PSP had a common extended haplotype characterized by a homozygous polymorphism in the 5' splice site untranslated region of exon 1, two missense mutations in exon 4A (Asp285Asn, Ala289Val), and a nonsense mutation in the 5' splice site of exon 8. CONCLUSIONS: This study demonstrates that 22 unrelated progressive supranuclear palsy (PSP) patients have four identical sequence variants within the tau gene that are not present in 24 age-matched controls. Although the functional significance of these results on tau protein expression is unknown, the presence of this "susceptibility" haplotype in individuals may place them at risk for developing PSP. PMID- 10534246 TI - Dopamine D2 receptor gene polymorphism and the risk of levodopa-induced dyskinesias in PD. AB - OBJECTIVE: To investigate whether polymorphisms in the genes for dopamine receptors D1 and D2 are associated with the risk of developing peak-dose dyskinesias in PD. BACKGROUND: Peak-dose dyskinesias are the most common side effects of levodopa therapy for PD. The identified predictors may only partially account for the risk of developing peak-dose dyskinesias because a substantial proportion of patients never develop peak-dose dyskinesias. Genetic factors could play a role in determining the occurrence of peak-dose dyskinesias. METHODS: A case-control study of 136 subjects with sporadic PD and 224 population control subjects. We studied three polymorphisms involving the dopamine receptor D1 gene and one intronic short tandem repeat polymorphism of the dopamine receptor D2 gene. RESULTS: The polymorphisms of the dopamine receptor D1 gene were not associated with the risk of developing PD or peak-dose dyskinesias. The 15 allele of the polymorphism of the dopamine receptor D2 gene was more frequent in parkinsonian subjects than in control subjects. More important, the frequency of both the 13 allele and the 14 allele of the dopamine receptor D2 gene polymorphism was higher in nondyskinetic than in the dyskinetic PD subjects. The risk reduction of developing peak-dose dyskinesias for PD subjects carrying at least 1 of the 13 or 14 alleles was 72% with respect to the PD subjects who did not carry these alleles. CONCLUSIONS: Certain alleles of the short tandem repeat polymorphism of the dopamine receptor D2 gene reduce the risk of developing peak dose dyskinesias and could contribute to varying susceptibility to develop peak dose dyskinesias during levodopa therapy. PMID- 10534247 TI - Safety and efficacy of NeuroBloc (botulinum toxin type B) in type A-resistant cervical dystonia. AB - OBJECTIVE: To determine the safety and efficacy of botulinum toxin type B (BoNT/B) in patients with type A-resistant cervical dystonia (CD). BACKGROUND: Local intramuscular injections of BoNT are an effective therapy for CD. After repeated use, some patients become resistant to therapy. BoNT/B, effective in type A toxin-responsive patients, is proposed as an alternative therapy for type A-resistant patients. METHODS: The authors performed a 16-week, double-blind, placebo-controlled trial of BoNT/B in type A-resistant patients with CD. After resistance to therapy was confirmed with the frontalis-type A test, placebo or 10,000 U BoNT/B was administered in a single session into two to four clinically involved muscles. The Toronto Western Spasmodic Torticollis Rating Scale (TWSTRS) was the primary efficacy measurement. TWSTRS-Total, three visual analog scales (Patient Global Assessment of Change, Principal Investigator Global Assessment of Change, Patient Analog Pain Assessment), and adverse events were assessed at baseline and weeks 2, 4, 8, 12, and 16. RESULTS: A total of 77 patients participated (38 placebo, 39 active). Improvements in severity, disability, and pain were documented in the BoNT/B-treated group. TWSTRS-Total scores were improved in the BoNT/B-treated group at weeks 4 (p = 0.0001), 8 (p = 0.0002), and 12 (p = 0.0129). All three visual analog scales demonstrated improvements at week 4 (p < 0.0001, 0.0001, and 0.001). A Kaplan-Meier analysis supported a duration of effect of 12 to 16 weeks in the active group. Dry mouth and dysphagia were self-limited adverse effects, reported more commonly in the BoNT/B group. CONCLUSIONS: Botulinum toxin type B (BoNT/B) (NeuroBloc) is safe and efficacious for the management of patients with type A-resistant cervical dystonia with an estimated duration of treatment effect of 12 to 16 weeks. PMID- 10534248 TI - Safety and efficacy of NeuroBloc (botulinum toxin type B) in type A-responsive cervical dystonia. AB - OBJECTIVE: To determine the safety and efficacy of botulinum toxin type B (BoNT/B) in patients with cervical dystonia (CD). BACKGROUND: BoNT/B is a form of chemodenervation therapy for the treatment of patients with CD. METHODS: The authors performed a 16-week, randomized, multicenter, double-blind, placebo controlled trial of BoNT/B in patients with CD who continue to respond to botulinum toxin type A. Placebo, or 5,000 U or 10,000 U of BoNT/B was administered in two to four muscles involved clinically in CD. The Toronto Western Spasmodic Torticollis Rating Scale (TWSTRS)-Total score at week 4 was the primary efficacy measure. Clinical assessments and adverse events were recorded for treatment day 1 and at weeks 2, 4, 8, 12, and 16. RESULTS: A total of 109 patients were enrolled randomly across all three treatment groups. The mean improvement in the TWSTRS-Total scores in each group at week 4 was 4.3 (placebo), 9.3 (5,000 U), and 11.7 (10,000 U). For the prospectively defined primary contrast (10,000 U versus placebo), highly significant differences were noted for the primary (TWSTRS-Total, baseline to week 4, p = 0.0004) and supportive secondary (Patient Global Assessment, baseline to week 4, p = 0.0001) outcome measures. Improvement in pain, disability, and severity of CD occurred for patients who were treated with BoNT/B when compared with placebo-treated patients. Overall, improvements associated with BoNT/B treatment were greatest for patients who received the 10,000-U dose. The duration of treatment effect for BoNT/B was 12 to 16 weeks for both doses. CONCLUSION: Botulinum toxin type B (NeuroBloc) is safe and efficacious at 5,000 U and 10,000 U for the management of patients with cervical dystonia. PMID- 10534249 TI - Bilateral thalamic stimulation for the treatment of essential tremor. AB - OBJECTIVE: To determine the safety and efficacy of bilateral thalamic stimulation in the treatment of essential tremor (ET). METHODS: Nine ET patients with disabling tremor refractory to pharmacotherapy underwent bilateral staged implants. Tremor was assessed by the Fahn-Tolosa-Marin Tremor Rating Scale at baseline 1 (before first implant), baseline 2 (before second implant), and at 6 month and 1-year follow-up. Blinded evaluations were performed at 3 months. Associated changes in speech were evaluated in six patients. There were seven men and two women with a mean age of 73.8 years. RESULTS: There was a significant improvement in the mean total tremor score from a baseline of 66.1+/-11.6 to 28.4+/-12.8 12 months after the second surgery. Similarly, the mean motor tremor subscore was 20.1+/-5.0 before the first surgery and improved significantly to 14.1+/-3.6 before the second surgery. Motor tremor scores 6 months after the second surgery (6.0+/-3.7) and 12 months after the second surgery (7.5+/-3.9) also improved significantly relative to the preoperative scores. The mean activities of daily living (ADL) subscore at baseline was 18.2+/-2.9 and improved significantly before the second surgery to 9.0+/-3.2. These ADL scores further improved 6 months (6.2+/-5.2) and 12 months (7.9+/-5.7) following the second surgery, but these gains were not significant. Blinded evaluations also revealed a similar degree of improvement. Complications were noted in five patients: asymptomatic intracranial hematoma (1), postoperative seizures (1), a hematoma over the implanted pulse generator (IPG) (1), lead repositioning (1), and IPG malfunction (1). Adverse effects related to stimulation were mild and resolved with adjustment of the stimulation parameters. Three of the six patients demonstrated worsening of dysarthria with both stimulators on. CONCLUSIONS: Bilateral thalamic stimulation is effective in reducing tremor and functional disability in ET; however, dysarthria is a possible complication. PMID- 10534250 TI - Time course of frontal somatosensory evoked potentials. Relation to L-dopa plasma levels and motor performance in PD. AB - OBJECTIVE: To verify whether the change in L-dopa plasma levels after a single dose of carbidopa/L-dopa 50/200 (controlled-release) transiently modifies frontal components of somatosensory evoked potentials (SEPs) in patients with PD in parallel with improvement of motor performance. BACKGROUND: Apomorphine, a potent dopamine-agonist drug, transiently increases frontal SEP components, which may be depressed in PD; however, relationships between clinical status, frontal SEPs, and therapy are still unclear. METHODS: Nineteen PD patients (mean age 65.9 years, range 52 to 77, responders to L-dopa therapy, were studied in the same day at times T0 (baseline predose level), T1 (presumed L-dopa peak time), and T2 (end of dose-induced motor response). The following were monitored: L-dopa plasma concentration, tapping test, reaction times, peak latency (with central conduction times), and amplitude of cervical, subcortical, as well as cortical parietal and frontal SEP components elicited by median nerve stimulation of the more clinically affected arm. RESULTS: The average amplitude of frontal components of PD patients was significantly reduced at T0 with respect to control subjects. A significant and transient amplitude increase of frontal SEPs was found at T1, in parallel with the L-dopa peak concentration and improvement in motor performance (tapping and reaction times), without significant changes in amplitude of parietal SEP waves. No latency shifts were observed in brain and spinal waves. CONCLUSIONS: L-Dopa may influence the responsiveness of the parkinsonian brain as assessed by frontal somatosensory evoked potentials. The time course of these modifications coincides with that of the clinical response in the motor performance. PMID- 10534251 TI - Critical decline in fine motor hand movements in human aging. AB - BACKGROUND: Slowing of motor movements in human aging is a well-known occurrence, but its biologic basis is poorly understood. Reliable quantitation may refine observations of this phenomenon to better aid research on this entity. METHODS: A panel equipped with timing sensors under computer control was used to measure upper extremity movement times in two groups of healthy individuals: adults younger than 60 years of age (n = 56; range, 18-58 years) and adults older than 60 years of age (n = 38; range, 61-94 years). RESULTS: Fine motor performance was better in the dominant hand (p = 0.0007) regardless of age. Adult and aged groups differed on two basic timing measures, which reflect coarse motor and fine motor performance (p < 0.0001). There were no gender differences on either measure. There was a strong effect of task difficulty with age on coarse motor (p < 0.01) and fine motor (p < 0.0001) measures. The fine motor measure of hand performance in healthy individuals correlated in a nonlinear fashion with age for more difficult tasks (r2 = 0.63) but showed a simple linear relation for less demanding tasks (r2 = 0.5). CONCLUSION: This technique sensitively detects age related motor performance decline in humans. There may be a critical period in late midlife when fine motor performance decline either begins or abruptly worsens. PMID- 10534252 TI - Motor cortex localization using functional MRI and transcranial magnetic stimulation. AB - OBJECTIVE: Congenital brain lesions producing focal seizures may be accompanied by reorganization of the areas responsible for motor and sensory functions within the brain due to a phenomenon that has been termed "neuronal plasticity." This can be studied using functional MRI (fMRI) and transcranial magnetic stimulation (TMS). Using either method, the motor cortex can be localized noninvasively, but to date there have been few studies correlating the level of agreement between the two techniques. METHODS: We used fMRI and TMS to localize the motor cortex in a young woman with intractable focal seizures, congenital left arm weakness, and a dysplastic right hemisphere on MRI. RESULTS: There was excellent agreement in the localization of motor representation for each hand. Both were predominantly located in the left hemisphere. fMRI also showed an area of posterior activation in the right hemisphere, but there was no evidence of descending corticospinal projections from this site using TMS, direct cortical stimulation, and Wada testing. CONCLUSIONS: Functional MRI (fMRI) and transcranial magnetic stimulation (TMS) were successfully used to localize cortical motor function before epilepsy surgery. Each technique demonstrated migration of motor function for the left hand to the left motor cortex. After resection of the dysplastic right precentral gyrus there was no permanent increase in weakness or disability. The two techniques are complementary; fMRI indicates all cortical areas activated by the motor task, whereas TMS identifies only those areas giving rise to corticospinal projections. PMID- 10534253 TI - Increasing incidence of medically recognized migraine headache in a United States population. AB - OBJECTIVE: To investigate trends in the incidence of medically recognized migraine in Olmsted County, Minnesota over approximately a decade. METHODS: The authors used the records-linkage system of the Rochester Epidemiology Project to identify individuals whose records included any diagnostic rubric related to headache for the 3-year period 1979 through 1981 and the 2-year period 1989 through 1990. A nurse abstracter and a neurologist (J.W.S.) reviewed the complete history of each potential case and assigned a diagnosis using the International Headache Society classification (IHS, modified). Only patients who consulted a doctor for their headache and had their initial visit for migraine within the study years were considered as incident cases. RESULTS: The incidence of medically recognized migraine increased in female subjects between the 1979 through-1981 period and the 1989-through-1990 period for all ages, but particularly among those who were aged 10 to 49 years. The peak incidence rate at age 20 to 29 years increased from 634.5 new cases per 100,000 person-years in 1979 through 1981 to 986.4 in the 1989-through-1990 period (absolute increase 351.9; relative increase 56%). The rise in incidence in female subjects was most sizable for migrainous disorder (IHS code 1.7); smaller increases were noted for migraine without aura and with typical aura. Only a slight absolute increase in migraine incidence rates was observed in male subjects, restricted to those 10 to 19 years of age (absolute increase 174.7; relative increase 89%). CONCLUSIONS: Although the incidence rates reported here are restricted to patients who consulted a doctor for their headache, the authors suggest that the incidence of migraine has increased over time in female subjects, especially those of reproductive age. The increase was most pronounced for migrainous disorder. Incidence rates were more stable in male subjects over time. PMID- 10534254 TI - Increased brain serotonin synthesis in migraine. AB - OBJECTIVE: To measure brain serotonin synthesis with PET using the tracer alpha [11C]methyl-L-tryptophan in migraine patients. BACKGROUND: Although the cause of migraine remains poorly understood, there is considerable evidence to support a role of the neurotransmitter serotonin in the pathophysiology of migraine. METHODS: We studied 11 women (aged 33+/-7.7 years) with a diagnosis of migraine according to International Headache Society criteria and 8 healthy women (aged 29+/-9.2 years). Five patients were studied before and after chronic treatment with propranolol or nadolol. RESULTS: Serotonin synthesis capacity (K-complex) values in migraine patients were higher than those measured in controls throughout the brain (p = 0.016); mean K-complex for whole brain was 0.0077 + 0.0020 mL/g/min in patients with migraine and 0.0054+/-0.0003 mL/g/min in controls. The regional pattern did not differ between the two groups. However, the K-complex for whole brain in the subgroup of migraine patients with aura (n = 3) did not differ from that of the control group (p = 0.32). In the five patients studied twice (before and after treatment), we found a trend of increased whole brain K-complex after drug treatment (p = 0.06). CONCLUSIONS: Our findings indicating increased brain serotonin synthesis capacity in migraine patients are consistent with previous reports of systemic alteration of serotonin metabolism in patients without aura. Our results also suggest that the mechanism of action of beta-adrenergic antagonists for migraine prophylaxis may involve regulation of serotonin synthesis. PMID- 10534255 TI - Impaired olfaction as a marker for cognitive decline: interaction with apolipoprotein E epsilon4 status. AB - OBJECTIVE: To determine whether olfactory status predicts cognitive decline (CD) over a 2-year follow-up period. METHODS: The authors enrolled individuals in a community-based longitudinal study of memory and aging in the Japanese-American community in King County, WA, between 1992 and 1994. At baseline they screened 1,985 persons using the Cognitive Abilities Screening Instrument (CASI) and the 12-item Cross-Cultural Smell Identification Test (CC-SIT). Of these 1,985 people, 1,836 were found not to be demented. Two years later the authors rescreened 1,604 participants with the CASI. They defined CD as a 2-year loss of > or =5.15 points/100 on the CASI. They genotyped 69% of the 1,604 people completing both examinations for apolipoprotein E (apoE). RESULTS: After adjusting for age, CASI score at baseline, education, smoking, sex, and follow-up time, the authors determined an odds ratio (OR) for CD of 0.90 (95% CI, 0.84 to 0.97) for an increase in each correct point on the CC-SIT (range, 0 to 12). Compared with normosmics, the OR for persons with impaired olfaction (microsmics) was 1.25 (95% CI, 0.83 to 1.89) and for anosmics the OR was 1.92 (95% CI, 1.06 to 3.47). Persons who were anosmic at baseline and who had at least one APOE-epsilon4 allele had 4.9 times the risk of CD (95% CI, 1.6 to 14.9) compared with normosmics without the epsilon4 allele. The estimated relative risk among women was 9.7 (95% CI, 1.3 to 70.4), and for men the risk was 3.2 (95% CI, 0.8 to 12.6). Receiver operating characteristic (ROC) curves showed that although the area under the curve (AUC) for baseline CASI was only 0.51, the AUC for CC-SIT alone was 0.62. Adding CC-SIT to the ROC model with CASI improved the AUC curve from 0.51 to 0.62. CONCLUSIONS: Unexplained olfactory dysfunction in the presence of one or more APOE-epsilon4 alleles is associated with a high risk of cognitive decline. Cross-Cultural Smell Identification Test classifies people with cognitive decline correctly to a greater degree than a global cognitive test. PMID- 10534256 TI - Sensitivity, specificity, and stability of CSF-tau in AD in a community-based patient sample. AB - OBJECTIVE: To evaluate the sensitivity and specificity of CSF-tau in clinical practice as a diagnostic marker for AD compared with normal aging and depression, to study the stability of CSF-tau in longitudinal samples, and to determine whether CSF-tau levels are influenced by different covariates such as gender, age, duration or severity of disease, or possession of the APOE-epsilon4 allele. METHODS: Consecutive AD patients from a community-based sample were studied, including 407 patients with AD (274 with probable AD and 133 with possible AD), 28 patients with depression, and 65 healthy elderly control subjects. A follow-up lumbar puncture was performed in 192 AD patients after approximately 1 year. CSF tau was determined using a sandwich ELISA, which was run as a routine clinical neurochemical analysis. RESULTS: CSF-tau was increased in probable (690+/-341 pg/mL; p < 0.0001) and possible (661+/-447 pg/mL; p < 0.0001) AD, but not in depression (231+/-110 pg/mL) compared with control subjects (227+/-101 pg/mL). Receiver operating characteristics analysis showed that a cutoff level of 302 pg/mL resulted in a sensitivity of 93% (95% CI, 90-96%) and a specificity of 86% (95% CI, 75-94%), with an area under the curve of 0.95 to discriminate AD from control subjects. Within the AD group, CSF-tau did not differ significantly between baseline and follow-up investigations, and was relatively stable between baseline and 1-year follow-up levels, with a coefficient of variation of 21.0%. High CSF-tau levels were also found in most AD patients with very short duration of dementia, and with Mini-Mental State Examination scores >23 (n = 205). In total, 193 of 205 patients (sensitivity, 94%) had a CSF-tau level higher than 302 pg/mL. CONCLUSIONS: CSF-tau has a high sensitivity and specificity to differentiate AD from normal aging and depression, as demonstrated in a large community-based series of consecutive AD patients during which analyses were run continually in a clinical neurochemical laboratory. The increase in CSF-tau is found very early in the disease process in AD, is stable over time, and has a low interindividual variation on repeated sampling. Although high CSF-tau is found in some neurologic conditions (e.g., stroke), these findings suggest that CSF-tau may be of use to help in differentiating AD from normal aging and depression, especially early in the course of the disease, when the symptoms are vague and the diagnosis is especially difficult. PMID- 10534257 TI - Elevated CSF prostaglandin E2 levels in patients with probable AD. AB - OBJECTIVE: To determine CSF eicosanoid concentrations and brain cyclo-oxygenase activity in AD patients and age-matched control subjects. BACKGROUND: Nonsteroidal anti-inflammatory drugs may benefit AD patients by inhibiting cyclo oxygenases and thereby reducing prostaglandin (PG) production or oxidant stress in the CNS. METHODS: CSF eicosanoid and F2-isoprostane (IsoP) levels were determined in seven probable AD patients and seven age-matched control subjects. Cyclo-oxygenase activity was determined in microsomes prepared from the hippocampus of 10 definite AD patients and 8 age-matched control subjects. All measurements were made using gas chromatography/mass spectrometry. RESULTS: CSF concentrations of prostaglandin (PG) E2 were increased fivefold (p < 0.01) and 6 keto-PGF1alpha was decreased fourfold (p < 0.01) in probable AD patients. There was no change in total CSF eicosanoid concentration in probable AD patients. CSF F2-IsoP, a quantitative marker of lipid peroxidation in vivo, was increased in probable AD patients (p < 0.05). Cyclo-oxygenase activity in the hippocampus from definite AD patients was not different from age-matched control subjects. CONCLUSIONS: These data suggest that cyclo-oxygenase activity may not contribute significantly to CNS oxidative damage in AD. Increased CSF PGE2 concentration in probable AD patients suggest that cyclo-oxygenase inhibitors may benefit AD patients by limiting PG production. PMID- 10534258 TI - Transthyretin amyloidosis and superficial siderosis of the CNS. AB - OBJECTIVE: To describe a previously unreported clinical and radiologic presentation of hereditary transthyretin (TTR)-related amyloidosis. BACKGROUND: Unexplained cerebellar ataxia, pyramidal syndrome, and hearing loss are observed in some patients with TTR-related amyloidoses. METHODS: We performed clinical, radiologic, and pathologic examinations of three family members with TTR-related (Ala36Pro) amyloidosis. RESULTS: The patient was a 69-year-old woman with vitreal amyloid deposits, progressive sensorineural deafness, cerebellar ataxia, pyramidal syndrome, and recurrent transient neurologic symptoms. Cranial MRI showed symmetric thin rims of low signal intensity in T2- and T2*-weighted images in the cortex of the sylvian fissures, of the cerebellar hemispheres and vermis, and in the quadrigeminal plate consistent with superficial siderosis of the CNS. Her older daughter had vitreal amyloid deposits, acute Brown-Sequard syndrome at C4, acute sensorineural deafness, and recurrent transient neurologic symptoms. Cranial MRI at age 48 revealed a rim of low signal intensity in T2- and T2* weighted images in the superior vermis folia and the right sylvian cortex. In addition, two small hemosiderin deposits were seen in the left parietal cortex. Lumbar puncture yielded colorless CSF with increased ferritin content and was followed by fourth ventricle hemorrhage. Cranial MRI 11 months later showed progression of brain hemosiderin deposits. The younger daughter had vitreal deposits, sensorimotor polyneuropathy, and acute sensorineural hearing but no evidence of siderosis on cranial MRI. She died at age 43 years of posterior fossa subarachnoid hemorrhage, and the neuropathologic examination showed amyloid deposition in the leptomeningeal spaces and vessels. CONCLUSION: Transthyretin related amyloidosis may cause superficial siderosis of the CNS through subarachnoid bleeding related to meningovascular amyloid deposition. PMID- 10534259 TI - Psychiatric adverse events during vigabatrin therapy. AB - OBJECTIVE: To determine the incidence of psychiatric adverse events associated with vigabatrin therapy, we reviewed data from US and non-US double-blind, placebo-controlled trials of vigabatrin as add-on therapy for treatment refractory partial epilepsy. METHODS: "Verbatim" terms from investigators' reports had been translated into standard "preferred" terms using an adverse event dictionary. Terms for psychiatric events were then combined into categories for analysis of rates during vigabatrin versus placebo treatment. RESULTS: Compared with placebo, vigabatrin subjects had a higher incidence of events coded as depression (12.1% versus 3.5%, p < 0.001) and psychosis (2.5% versus 0.3%, p = 0.028); there were no significant differences between treatment groups for aggressive reaction, manic symptoms, agitation, emotional lability, anxiety, or suicide attempt. Although depression and psychosis were typically observed during the first 3 months, most studies were 12 to 18 weeks long, so that definitive conclusions could not be reached about timing of events. Psychosis was generally transient and reported to be responsive to reduction or discontinuation of vigabatrin or to neuroleptic treatment. Depression was typically mild. Serious depression, defined as discontinued from the study, hospitalized, or suicide attempt, or coded as psychotic depression, occurred in only 9 of the 49 vigabatrin-treated patients with depression. CONCLUSIONS: Vigabatrin use in treatment-refractory partial epilepsy is associated with increased occurrence of depression and of psychosis, although the frequency of psychosis is apparently lower than previously reported. Clinical experience suggests that these adverse events respond to reduction of vigabatrin dose or to counteractive psychotropic treatment. PMID- 10534260 TI - Subjective versus objective memory change after temporal lobe epilepsy surgery. AB - OBJECTIVE: To examine subjective versus objective memory change after anterior temporal lobectomy (ATL). METHODS: A prospective, controlled study. Controls included 39 unoperated patients with intractable temporal lobe epilepsy (TLE) who were administered a series of cognitive and health-related quality of life measures at baseline and at 12-month follow-up intervals. The surgery sample included 65 patients with intractable, focal TLE who had undergone either a right or left ATL. These patients were tested preoperatively and at 6-month follow-up intervals. Subjective and objective memory change was quantified using a newly developed methodology to control for practice effect and regression to the mean. RESULTS: Measures of subjective and objective memory change were not significantly related in the surgery sample. Prevalence of significant subjective memory decline 1 year after surgery ranged from 3 to 7%, whereas prevalence of significant objective memory decline ranged from 26 to 55%. Postoperative levels of emotional distress significantly predicted self-reported memory decline 1 year after ATL. Postoperative medication side effect and seizure outcome were also related significantly to subjective memory change in patients who had undergone left ATL. CONCLUSIONS: Subjective and objective memory change after temporal lobectomy are not related. Complaints of significant memory decline after ATL are infrequent and may serve as a marker for depression or other mood disorder rather than organically based memory decline. PMID- 10534261 TI - The effect of vigabatrin (gamma-vinyl GABA) on cerebral blood flow and metabolism. AB - OBJECTIVE: To investigate the effect of vigabatrin (VGB; gamma-vinyl gamma aminobutyric acid [GABA]), a selective irreversible GABA-transaminase inhibitor, on cerebral metabolic rate for glucose (CMRGlc) and cerebral blood flow (CBF) measured with 18F-fluorodeoxyglucose (FDG) PET and 15O water PET. BACKGROUND: Antiepileptic drugs (AEDs) reduce CMRGlc to varying degrees. Phenobarbital causes a mean decrease of 30 to 40%. Phenytoin, carbamazepine (CBZ), and valproate (VPA) cause milder reductions in CMRGlc. The combination of VPA with CBZ results in a greater decrease than either drug alone. The effect of novel AEDs on both CBF and CMRGlc has not been studied extensively. METHODS: Fourteen patients with refractory complex partial seizures on CBZ monotherapy for 4 weeks were included in the study. All patients had baseline 18F-FDG and 15O water PET studies followed by double-blind randomization to placebo (PLC) or VGB while on continuous CBZ treatment. PET scans were repeated after an interval of 2 months on target dose of VGB (50 mg/kg) or PLC. Quantitative PET data analysis was performed using a region of interest template. Significance was tested with the Wilcoxon rank sum test. RESULTS: No statistically significant difference in age, duration of epilepsy, or CBZ levels was observed in the two patient groups. VGB reduced global CMRGlc by 8.1+/-6.5% and global CBF by 13.1+/-10.4%. The change in CMRGlc was different in patients taking VGB compared with those on PLC (p < 0.04). VGB patients showed regional decreases in both CMRGlc and CBF, particularly in temporal lobes. CSF total GABA increased in the VGB patient group (1.48+/-1.06 versus 4.03+/-4.19 nm/mL). The increase differed from the PLC group (p < 0.03). We found a strong relation between decreased total CSF GABA and increased CMRGlc in the VGB patient group (R2 = 0.82, p < 0.01). CONCLUSIONS: Vigabatrin (VGB) causes mild reductions in both cerebral blood flow (CBF) and cerebral metabolic rate for glucose (CMRGlc) in contrast to other drugs such as barbiturates, which are direct agonists at the gamma-aminobutyric acid benzodiazepine receptor complex. Conventional AEDs depress CBF and CMRGlc to a greater degree than does VGB. The relatively mild reduction could be due to pre- as well as postsynaptic effects or a use-dependent mechanism. PMID- 10534262 TI - Antiphosphatidyl serine antibodies are independently associated with ischemic stroke. AB - OBJECTIVE: To determine whether elevated titers of antiphosphatidyl serine antibodies (aPS) are associated with an increased risk of ischemic stroke in a general stroke population. BACKGROUND: aPS are members of the family of antiphospholipid antibodies that has been associated with increased stroke risk. Although aPS have been demonstrated to occur in 18% of a group of young patients with cerebrovascular symptoms, their prevalence in the general stroke population is unknown, and no controlled study to assess the strength of their association with ischemic stroke has been undertaken previously. METHODS: A case-control study comparing 267 acute ischemic stroke patients and 653 community controls. Sera were obtained immediately after acute stroke in patients. Titers of IgG aPS >16 IgG phospholipid units or IgM aPS >22 IgM phospholipid units were considered positive. Odds ratios (ORs) were obtained by logistic regression, adjusting for age, gender, race/ethnicity, history of hypertension, diabetes mellitus, cardiovascular disease, and cigarette smoking. RESULTS: The adjusted OR was 5.6 (95% confidence interval [CI] 1.8, 18.0) for IgG aPS and 2.9 (95% CI 1.6, 5.3) for IgM aPS. The adjusted OR for either an elevated IgG or IgM aPS was 3.2 (95% CI 1.8, 5.5). CONCLUSIONS: This study demonstrates that elevated IgG and IgM antiphosphatidyl serine antibodies titers are associated with increased risk of ischemic stroke. The prevalence of these antibodies is lower, but the associated stroke risk is comparable with that of anticardiolipin antibodies. PMID- 10534263 TI - The ischemic penumbra: operationally defined by diffusion and perfusion MRI. AB - BACKGROUND: Identifying tissue at risk for infarction is important in deciding which patients would benefit most from potentially harmful therapies and provides a way to evaluate newer therapies with regard to the amount of ischemic tissue salvaged. OBJECTIVE: To operationally define and characterize cerebral tissue at risk for stroke progression. METHODS: We retrospectively selected 25 patients with an acute onset of a hemispheric stroke from our database who had undergone a combination of two diffusion-weighted MRI studies and a perfusion-weighted MRI study. We applied a logistic regression model using maps of the relative mean transit time and relative cerebral blood flow (rCBF) as well as three different maps of the relative cerebral blood volume (rCBV) to predict an operationally defined penumbra (region of mismatch between the diffusion lesion on day 1 and its extension 24 to 72 hours later). RESULTS: Maps of the rCBF and initial rCBV were significant predictors for identifying penumbral tissue. Our operationally defined penumbral region was characterized by a reduction in the initial rCBV (47% of contralateral control region [CCR]), an increase (163% of CCR) in the total rCBV, and a reduction (37% of CCR) in the rCBF, whereas the operationally defined ischemic core showed a more severe reduction in the rCBF (12% of CCR) and in the initial rCBV (19% of CCR). CONCLUSION: These MR indexes may allow the identification and quantification of viable but ischemically threatened cerebral tissue amenable to therapeutic interventions in the hyperacute care of stroke patients. PMID- 10534264 TI - Isolated intracranial hypertension as the only sign of cerebral venous thrombosis. AB - BACKGROUND: Cerebral venous thrombosis (CVT) is often overlooked when intracranial hypertension (ICH) is isolated, hence mimicking idiopathic intracranial hypertension (IIH). OBJECTIVE: To describe the characteristics of patients with CVT and ICH. METHODS: We examined 160 consecutive patients with CVT between 1975 and 1998. They were separated into two groups according to their clinical presentation--isolated ICH and other neurologic symptoms and signs. RESULTS: Fifty-nine patients with CVT (37%) presented with isolated ICH. Neuroimaging showed involvement of more than one sinus in 35 patients (59%). Brain CT was normal in 27 of 50 patients (54%). Lumbar puncture was performed in 44 patients and showed elevated opening pressure in 25 of 32 (78%) and abnormal CSF content in 11 (25%). Etiologies and risk factors included local causes in 7 of 59 (12%), surgery in 1, inflammatory diseases in 18 (30.5%), infection in 2, cancer in 1, postpartum state in 1, coagulopathies in 11 (19%), oral contraception in 7 (12%), and remained unknown in 11 (19%). Anticoagulants were used in 41 of 59 patients (69.5%), steroids or acetazolamide in 26 (44%), therapeutic lumbar puncture in 44 (75%), and surgical shunt in 1. Three patients had optic atrophy with severe visual loss, 1 died from carcinomatous meningitis, and 55 (93%) had complete recovery. CONCLUSIONS: Central venous thrombosis (CVT) can present with all the classical criteria for idiopathic intracranial hypertension (IIH), including normal brain CT with normal CSF content. Because the recognition of CVT has crucial prognostic and therapeutic implications, MRI, with magnetic resonance venography when necessary, should be performed in patients with isolated intracranial hypertension. The outcome of CVT is unpredictable, and management of patients with CVT should not differ whether they present with isolated raised intracranial pressure or with other neurologic symptoms and signs. Therefore, isolated raised intracranial pressure from CVT differs in management from IIH and should be classified neither as "IIH" nor "pseudotumor cerebri." PMID- 10534265 TI - Neurologic complications of pediatric lung transplantation. AB - OBJECTIVE: To report neurologic complications in a large population of pediatric lung transplantation patients. METHODS: A retrospective review of the first 135 patients to undergo lung transplantation at St. Louis Children's Hospital from July 1990 to December 1997. RESULTS: Sixty-one (45%) patients had neurologic complications. The most common presenting symptoms were seizures (27%), followed by encephalopathy, headache, depression, and focal neurologic deficits. Cyclosporine toxicity (7%) and hypoxia-ischemia (7%) constituted the most commonly identified etiologies, followed by stroke, metabolic, and infectious causes. Risk factor analysis found that patients with interstitial lung disease had a higher frequency of hypoxic-ischemic events and patients with seizures had significantly elevated trough cyclosporine levels. Patients with stroke and hypoxia had a poor neurologic prognosis, whereas patients with cyclosporine toxicity uniformly had a good outcome. CONCLUSIONS: Neurologic complications occur frequently after lung transplantation in pediatric patients, with seizures being the most common presenting symptom. Except in patients with stroke and hypoxia, prognosis is generally favorable. Seizures not accompanied by an irreversible structural etiology are unlikely to require long-term treatment with antiepileptic medications. Cyclosporine neurotoxicity typically resolves without requiring discontinuation of immunosuppressive therapy. PMID- 10534266 TI - Lack of sodium channel mutation in an Italian family with paramyotonia congenita. AB - OBJECTIVE: To conduct the genotype-phenotype correlation in a family in which several individuals share clinical and electrophysiologic features of paramyotonia congenita (PC). BACKGROUND: PC, hyperkalemic periodic paralysis (HyperPP), and potassium-aggravated myotonias form the group of hereditary sodium channelopathies. Each of these disorders is associated with different point mutations in SCN4A, the gene encoding the alpha-subunit of the adult human skeletal muscle sodium channel. However, in HyperPP families, evidence of a causative gene different from SCN4A has been found. METHODS: We conducted direct clinical examination, electrophysiologic (EMG/electroneurographic) and cardiologic studies, as well as laboratory screening in several affected and nonaffected members of the family. We performed the genotype-phenotype correlation by microsatellite linkage and cDNA-mutation analyses of the SCN4A gene. RESULTS: Affected members in this family showed clinical and electrophysiologic features typical of PC. The disease phenotype segregated with the chromosomal region that includes the SCN4A gene. Analysis of the entire cDNA sequence of the SCN4A gene in the index case disclosed a G3826A transition, which results in the Val1276Ile substitution. However, PCR-single-stranded confirmation polymorphism and direct sequencing analysis of the segment coding for Val-1276 on genomic DNA confirmed the G3826A transition in the index case but was negative in 11 affected members of the family; however, neither mutations nor aberrant splicings causative of the PC phenotype in this family were found on SCN4A. CONCLUSION: The existence of a second gene different from SCN4A that can give rise to a clinical PC phenotype can be speculated upon. PMID- 10534267 TI - Insulin acts in hypokalemic periodic paralysis by reducing inward rectifier K+ current. AB - OBJECTIVE: To define how insulin acts in hypokalemic periodic paralysis (HypoPP). BACKGROUND: HypoPP results from point mutations of the skeletal muscle L-type Ca2+ channel. Attacks of flaccid paralysis are associated with hypokalemia and triggered by insulin. A persistent inward current causes depolarization-induced paralysis. The relationships of the Ca2+ channel mutations to the persistent inward current and how insulin triggers paralytic attacks are not yet known. METHODS: Intercostal muscle fibers from HypoPP and normal subjects were studied in vitro at 37 degrees C using two electrodes to determine action potential thresholds and a three-electrode voltage clamp to study membrane currents. RESULTS: HypoPP fibers were depolarized in bathing solution with 4 mM K+. Reducing K+ from 4.0 mM to 2.5 or 1.0 mM depolarized HypoPP fibers but hyperpolarized normal fibers. Adding 12 mU/mL of insulin to bathing fluids increased the depolarization of HypoPP fibers and increased the hyperpolarization of normal fibers. Depolarized HypoPP had increased action potential thresholds. The fraction of excitable muscle fibers decreased with increasing fiber depolarization. Blocking Na+ channels or L-type Ca2+ channels did not prevent depolarization induced by hypokalemia or by insulin. Insulin reduced the conductance of the inward rectifier K+ channel for outward-flowing currents. CONCLUSIONS: Insulin potentiates depolarization of hypokalemic periodic paralysis (HypoPP) fibers by reducing inward rectifier K+ conductance. The Ca2+ mutations in HypoPP indirectly derange membrane excitability by altering the function of other membrane channels. PMID- 10534268 TI - A common mutation (epsilon1267delG) in congenital myasthenic patients of Gypsy ethnic origin. AB - OBJECTIVE: Mutation analysis of the acetylcholine receptor (AChR) epsilon subunit gene in patients with sporadic or autosomal recessive congenital myasthenic syndromes (CMS). BACKGROUND: The nicotinic AChR of skeletal muscle is a neurotransmitter-gated ion channel that mediates synaptic transmission at the vertebrate neuromuscular junction. Mutations in its gene may cause congenital myasthenic syndromes. A recently described mutation in exon 12 of the AChR epsilon subunit (epsilon1267delG) disrupts the cytoplasmic loop and the fourth transmembrane region (M4) of the AChR epsilon subunit. METHODS: Forty-three CMS patients from 35 nonrelated families were clinically classified as sporadic cases of CMS (group III according to European Neuromuscular Centre consensus) and were analyzed for epsilon1267delG by PCR amplification and sequence analysis. RESULTS: The authors report the complete genomic sequence and organization of the gene coding for the epsilon subunit of the human AChR (accession number AF105999). Homozygous epsilon1267delG was identified in 13 CMS patients from 11 independent families. All epsilon1267delG families were of Gypsy or southeastern European origin. Genotype analysis indicated that they derive from a common ancestor (founder) causing CMS in the southeastern European Gypsy population. Phenotype analysis revealed a uniform pattern of clinical features including bilateral ptosis and mild to moderate fatigable weakness of ocular, facial, bulbar, and limb muscles. CONCLUSIONS: The mutation epsilon1267delG might be frequent in European congenital myasthenic syndrome patients of Gypsy ethnic origin. In general, patients (epsilon1267delG) were characterized by the onset of symptoms in early infancy, the presence of ophthalmoparesis, positive response to anticholinesterase treatment, and the benign natural course of the disease. PMID- 10534270 TI - Diagnostic value of myotactic reflexes in axonal and demyelinating polyneuropathy. AB - In 11 patients with chronic inflammatory demyelinating polyneuropathy (CIDP) and 11 patients with chronic idiopathic axonal polyneuropathy (CIAP), absent myotatic reflexes were significantly associated more often with CIDP than with CIAP, an absent biceps-reflex having the highest sensitivity and specificity for the diagnosis of CIDP. In CIDP, the latencies of electromyographically recorded myotatic reflexes often indicated demyelination, notwithstanding normal clinically assessed myotatic reflexes. Myotatic reflexes may therefore be useful for the distinction between axonal and demyelinating polyneuropathy. PMID- 10534269 TI - Alternative splicing patterns of CYP2D genes in human brain and neurodegenerative disorders. AB - The expression patterns of alternatively spliced forms of the CYP2D (6, 7, 7A, 7B) gene were analyzed in the brains of individuals with Lewy body disease (LBD) and correlated with CYP2D6 polymorphisms. Five different alternatively spliced transcripts were identified. The most common was the deletion of exon 6 (87.3% of cases), followed by a 91-base pair fragment deletion at the 3' end of the gene (63.9% of cases). There was no correlation between the polymorphisms in the CYP2D6B gene or presence of LBD and these five alternatively spliced transcripts. Susceptibility to LBD may occur through mechanisms other than altered mRNA splicing of the CYP2D6 gene. PMID- 10534271 TI - The effect of hyperventilation on downbeat nystagmus in cerebellar disorders. AB - Hyperventilation can affect nystagmus in patients with vestibular disorders. However, the effects on nystagmus in patients with cerebellar disease have not been systematically studied. Using the magnetic field search coil technique, we studied the effects of hyperventilation on nystagmus in a series of cerebellar patients. In four of eight patients, hyperventilation produced an increase in the slow-phase velocity of downbeat nystagmus. We speculate that this effect may be mediated through metabolic effects on cerebellar calcium channels. PMID- 10534272 TI - Nonvasculitic autoimmune inflammatory meningoencephalitis (NAIM): a reversible form of encephalopathy. AB - Five patients, age 54 to 80 years, presented between 3 weeks and 18 months after symptomatic onset of progressive cognitive decline, psychosis, and unsteady gait that proved to be due to a steroid-responsive nonvasculitic autoimmune inflammatory meningoencephalitic syndrome. CSF examination showed elevated immunoglobulin (Ig)G index and IgG synthesis rate in all three patients in whom it was checked, and brain biopsy revealed perivascular lymphocytic infiltrates without vessel wall invasion. PMID- 10534273 TI - Heavy nonencephalitic cerebral cysticercosis in tapeworm carriers. The Cysticercosis Working Group in Peru. AB - We describe 11 patients with massive brain infection with viable cysticerci, undetectable inflammatory reaction on CNS imaging, and an unexpectedly high (82%) prevalence of tapeworm infection. With the exception of two individuals with heavy parasite loads, patients had a relatively benign clinical course and tolerated the use of cysticidal drugs. This group of patients represents a particular presentation of neurocysticercosis, different from the previously described syndromes of cysticercotic encephalitis and disseminated systemic cysticercosis. PMID- 10534274 TI - Clinical outcome in pneumococcal meningitis correlates with CSF lipoteichoic acid concentrations. AB - Lipoteichoic and teichoic acids are components of the cell wall of Streptococcus pneumoniae. A recently developed enzyme immunoassay was used in patients with pneumococcal meningitis to investigate lipoteichoic and teichoic acid concentrations in CSF at the first lumbar puncture in relation to the clinical outcome determined by the Glasgow Outcome Scale. Lipoteichoic and teichoic acid concentrations in CSF were significantly associated with neurologic sequelae and mortality in S. pneumoniae meningitis. PMID- 10534275 TI - Use of Bacille Calmette-Guerin (BCG) in multiple sclerosis. AB - We studied the effect of Bacille Calmette-Guerin (BCG) vaccine as an immunomodulator in MS. According to the guidelines for clinical trials in MS, a single crossover, MRI-monitored trial was performed in 14 patients with relapsing remitting MS. After treatment, MRI activity was significantly reduced. No major adverse effects were reported. Adjuvant therapy with BCG vaccine was safe and merits study in MS. PMID- 10534276 TI - Postictal heart rate oscillations in partial epilepsy. AB - We report postictal heart rate oscillations in a heterogeneous group of patients with partial epilepsy. This pattern is marked by the appearance of transient but prominent low-frequency heart rate oscillations (0.01 to 0.1 Hz) immediately after 5 of 11 seizures recorded in 5 patients. This finding may be a marker of neuroautonomic instability and, therefore, may have implications for understanding perturbations of heart rate control associated with partial seizures. PMID- 10534277 TI - The effect of APOE on dementia is not through atherosclerosis: the Rotterdam Study. AB - The APOE genotype is involved in atherosclerosis, and atherosclerosis increases the risk of dementia, in particular among carriers of the APOE-epsilon4 allele. We studied, in a population-based setting (244 dementia cases; 1,002 nondemented controls), whether APOE is associated with dementia through atherosclerosis. As neither adjusting nor stratification for atherosclerosis altered the association of APOE with dementia, our study suggests that atherosclerosis is not an intermediate factor. PMID- 10534278 TI - Crossmodal and sensorimotor integration in tactile awareness. AB - We investigated tactile awareness in three patients with tactile extinction of stimuli located on contralesional somatotopic space. Contralesional tactile awareness was enhanced when they gazed to the left and when they moved their limbs. We suggest that personal somatotopic and peripersonal space are integrated by polymodal and sensorimotor links, which allow us to be aware of and act effectively on stimuli in space. PMID- 10534279 TI - Perceptual processing of stereopsis in humans: high-field (3.0-tesla) functional MRI study. AB - Cortical activation associated with stereopsis was studied in eight right-handed neurosurgeons professionally trained in stereoscopic vision. The activation map associated with viewing three-dimensional images, as contrasted to viewing the corresponding two-dimensional images of identical contents (images of MR angiography), showed consistent activation in the cortex adjacent to the intraparietal sulcus. The study further demonstrated a dominant role of the right hemisphere in perceptual processing of stereopsis in humans. PMID- 10534280 TI - Autosomal-recessive juvenile parkinsonism in a Jewish Yemenite kindred: mutation of Parkin gene. AB - We report a Jewish family of Yemenite origin in which three brothers born from a consanguineous marriage had juvenile parkinsonism. The DNA samples from three affected brothers and one healthy brother were analyzed for the linkage to markers covering the autosomal-recessive juvenile parkinsonism (AR-JP) locus. A perfect homozygous cosegregation to the markers was found, giving a maximal lod score of 3.11 at D6S1579, D6S305, and D6S411, all of which are 0 cm apart from each other (nonparametric linkage score, 8.041; p = 0.000977). Exon 3 of the Parkin gene was homozygously deleted in all patients. The AR-JP gene also exists in the Jewish population. PMID- 10534281 TI - Long-lasting improvement following (-)-OSU6162 in a patient with Huntington's disease. PMID- 10534282 TI - Subacute thyroiditis in a patient with MS treated with interferon beta-1a. PMID- 10534283 TI - Cerebellar ataxia associated with subclinical celiac disease responding to gluten free diet. PMID- 10534284 TI - A medullary syndrome characterized by wild arm ataxia. PMID- 10534285 TI - Possible usefulness of lamotrigine in the treatment of SUNCT syndrome. PMID- 10534286 TI - High prevalence of neurovascular instability in neurodegenerative dementias. PMID- 10534287 TI - Dopamine agonists reorient visual exploration away from the neglected hemispace. PMID- 10534288 TI - Dopamine agonists reorient visual exploration away from the neglected hemispace. PMID- 10534289 TI - Incidence and clinical consequence of the purple glove syndrome in patients receiving intravenous phenytoin. PMID- 10534290 TI - Toward optimal health: the experts respond to fibroids. Interview by Jodi Godfrey Meisler. PMID- 10534291 TI - Innovating in new market spaces. PMID- 10534292 TI - The future of weight management. PMID- 10534293 TI - Pharmacotherapy of dyslipidemia in postmenopausal women: weighing the evidence. AB - In the United States, coronary heart disease (CHD) is the leading cause of death in women. The incidence of CHD rises dramatically in women following menopause, which can be partially attributed to a more atherogenic lipoprotein profile. For years, observational and epidemiological data have suggested that estrogen and progesterone therapy reduced CHD end points. However, the first prospective trial that evaluated hormone replacement therapy (HRT) for secondary CHD prevention demonstrated no positive cardiovascular benefit of HRT compared with placebo. In interventional studies, the 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA)reductase inhibitors significantly reduced CHD outcomes in postmenopausal women, and these agents have emerged as the drugs of choice for primary and secondary CHD prevention. The selective estrogen receptor modulators (SERMs) may have a role in CHD prevention, but long-term clinical trials evaluating end points are needed. An evidence-based approach is necessary when deciding the appropriate pharmacotherapy of dyslipidemia in postmenopausal women. PMID- 10534294 TI - Menstrual migraine. AB - Migraine in women is influenced by hormonal changes throughout the life cycle: menarche, menstruation, oral contraceptive use, pregnancy, menopause, and hormonal replacement therapy (HRT). Based on clinical experience, the frequency of menstrual migraine has been reported to be as high as 60%-70%. Most women have increased headache and migraine attacks (usually without aura) at the time of menses. Attacks occurring only with menstruation, even if infrequent, are called true menstrual migraine. Attacks occurring both at menstruation and at other times of the month could be called "menstrually triggered migraine." Menstrual migraine occurs at the time of the greatest fluctuation in estrogen levels. Estrogen withdrawal is probably the trigger for migraine attacks in susceptible women. Drugs that are proven effective or commonly used for the acute treatment of menstrual migraine include nonsteroidal anti-inflammatory drugs (NSAIDs), dihydroergotamine, the triptans, and the combination of aspirin, acetaminophen, and caffeine. The goal of standard continuous preventive therapy is to reduce the frequency, duration, and intensity of attacks. Preventive therapy may eliminate all headaches except those associated with menses. Women already using prophylactic medication who continue to have menstrual migraine can increase the dose of their medication prior to their menses. Women who do not use preventive medicine or have migraine exclusively with their menses can be treated perimenstrually with short-term prophylaxis. If severe menstrual migraine cannot be controlled by acute and preventive treatment, hormonal therapy may be indicated. PMID- 10534295 TI - Cosmetic and postmastectomy breast implants: Finnish women's experiences. AB - The purpose of this study was to compare women's satisfaction with and short-term problems of silicone breast implants after cosmetic breast augmentation and after mastectomy. Women (n = 224) were recruited through advertising in mass media, and 91% responded to a questionnaire asking for their experiences, both positive and negative, with silicone breast implants. Approximately equal numbers of women received their implants for cosmetic reasons (augmentation group) and postmastectomy (113 and 111, respectively). Mean time from first implantation was 9 years (SD 7.3) in the cosmetic group and 8 years (SD 4.9) in the postmastectomy group. Women in the postmastectomy group received their implants at an older age than women in the cosmetic group (percent of women 45 and older, 59% and 3%, respectively). Women's overall preoperative knowledge of and postoperative satisfaction with their implants were similar in the two groups; 58% of women said that they had insufficient knowledge of breast implants preoperatively, 26% of women said they would not choose the implants again, and 44% of women expressed no dissatisfaction with their breasts. However, women in the cosmetic group were better informed about possible physical problems. One third of the women in the postmastectomy group had one or more reoperations, most frequently because of implant slippage (30%), encapsulation (26%), or implant size and shape (23%). Because insertion of breast implants is a lifelong decision, in-depth counseling about complication rates and possible risks should be given to women before implantation, and nonsurgical alternatives should be discussed, particularly for cosmetic implantation. PMID- 10534296 TI - Patient preference for the management of mildly abnormal Papanicolaou smears. AB - Our objective was to investigate patient knowledge, desire for participation in medical decision making, and preference for the management of mildly abnormal Papanicolaou (Pap) smears (low-grade squamous intraepithelial lesions [LGSIL]) in the context of the continuing controversy between active (immediate colposcopy and biopsy) and surveillance (repeat Pap smears) management strategies. One hundred thirty-six women referred for a diagnostic colposcopy with a first-time mildly abnormal Pap smear result completed questionnaires before contact with either the nurse or physician. They were given the State-Trait Anxiety Inventory, the CESD Depression scale, and a knowledge about dysplasia quiz. They were then presented with the two management options and asked to state a preference, if any. They then completed the Problem Solving-Decision Making Scale, a measure of desire for involvement in medical decision making. The majority of women in this sample opted for the active management strategy. Management preference was related to anxiety, with the most anxious women more likely to choose the active management strategy. Management preference was not related to knowledge or to desire for an active role in decision making, although the more knowledgeable women also reported a desire for an active role in the decision-making process. Given the current controversy over the management of mildly abnormal Pap smears (LGSIL), as well as the fact that there is no conclusive evidence to support one strategy over another, the informed management preference of women affected by these decisions should be factored into the equation. PMID- 10534297 TI - The risk of premature menopause induced by chemotherapy for early breast cancer. AB - The objectives of this retrospective case series were to determine the prevalence and timing of menstrual abnormalities in early-stage breast cancer patients undergoing adjuvant methotrexate or anthracycline-based chemotherapy and to more fully assess the possible mechanism of the amenorrhea reported after chemotherapy. One hundred forty-two premenopausal patients undergoing adjuvant chemotherapy were analyzed for patient age, breast cancer stage, type of chemotherapy, and menstrual abnormalities before, during, and after chemotherapy completion. A 24-month minimum follow-up after chemotherapy completion was available for all patients. One hundred nine of 142 patients were evaluable. Sixty-nine patients (46 node negative, 23 node positive) received methotrexate based chemotherapy, 33 patients (3 node negative, 30 node positive) received anthracycline-based chemotherapy, and 7 patients received both treatments (all node positive). Amenorrhea occurred in about a third of patients during chemotherapy (methotrexate groups 31%, anthracycline group 33%), and a higher proportion were amenorrheic 1 year after chemotherapy was completed (methotrexate group 45%, anthracycline group 46%). Abnormalities were more likely to occur in older premenopausal patients (Chi square = 6.18, p < 0.05), although 28% of patients under age 35 developed persistent abnormal menses. In some amenorrheic patients, follicle-stimulating hormone (FSH) levels were decreased within 6 months of chemotherapy (24.4 IU). The levels tended to be higher after chemotherapy (59.1 IU), suggesting ovarian failure. Menstrual abnormalities and menopause will frequently occur in premenopausal early-stage breast cancer patients, with 30% of all patients amenorrheic 1 year after chemotherapy. PMID- 10534298 TI - Domestic violence and sexual abuse in women physicians: associated medical, psychiatric, and professional difficulties. AB - Physicians have been called on to identify victims of domestic violence (DV) and sexual abuse (SA). Few data exist, however, on the prevalence of DV and SA in physicians themselves or on the personal or professional sequelae of such experiences. We determined the reported lifetime prevalence of DV and SA among women physicians and the personal characteristics, health-related factors, and work-related factors associated with these forms of abuse. We used data from the Women Physicians' Health Study, a large (n = 4501 respondents), nationally distributed questionnaire study that included questions on DV and SA histories, personal characteristics, and psychiatric, medical, and work-related histories. We compared the characteristics of women physicians with and without histories of DV or SA. The logistic models indicate that women physicians reporting DV histories (3.7% of the population) were significantly (p < 0.05) less likely to be single and significantly more likely to report depression histories, suicide attempts, substance abuse, current or past cigarette smoking, severe daily stress at home, chronic fatigue syndrome, and DV experienced by their mothers. Women physicians reporting SA histories (4.7% of the population) were significantly more likely to be younger than 60 years, identify themselves as homosexual or bisexual, to have specialized in psychiatry, obstetrics and gynecology, or emergency medicine, and to report histories of depression, suicide attempts, eating disorders, and fair or poor perceived health status. Although the reported lifetime prevalence of DV and SA among women physicians is below other reported figures, such experiences are associated with medical and psychiatric difficulties that could negatively affect them personally and professionally. PMID- 10534299 TI - Where nontradition is the norm: are sex and age determinants of practicing primary care specialties? AB - We studied the gender, age at graduation, and specialty of 2329 graduates of The Medical College of Pennsylvania (MCP) to determine if women and older graduates of a historically female institution tend to practice primary care specialties. Four of the primary care specialties studied, obstetrics and gynecology, family practice, general internal medicine, and pediatrics, are actively engaged in promoting women's health. MCP graduates were selected for study because of the institution's commitment to women's health and its association with admitting qualified, nontraditional students whose gender and age may have inhibited acceptance elsewhere. Seventy-two percent (1672) of the 1970-1992 graduates responded to an alumnae/i questionnaire. Chi-square tests revealed that female graduates were more likely to practice family practice, pediatrics, and obstetrics-gynecology but not more likely to practice general internal medicine. There was no relationship between age and practicing any of the four specialties. As more females graduate from U.S. medical schools, it is likely that they will retain their tendency to practice primary care specialties. These specialties offer women the opportunity to practice various aspects of comprehensive, lifelong women's healthcare. We should not expect older graduates schooled in environments favorable to women's health and careers to practice primary care medicine. PMID- 10534301 TI - Strategies for building a multidisciplinary academic program in women's health. AB - During the decade of the 1990s, women's health has received unprecedented attention from government, industry, marketers of healthcare, and academic medical centers. An assessment of research, education, and healthcare delivery has exposed gaps in our knowledge about gender-related issues. Recognition of gender as a rich frontier for innovation and discovery has resulted in widespread and varied responses and a commensurate increase in activity in the field. However, this diversity of effort has created the new challenge of effectively communicating strategies of response to the multiple disciplines invested in women's health. This article describes a strategy used at Duke University Medical Center to build awareness of women's health through a highly visible and successful Women's Health Seminar Series. The series serves as a focal point for broader efforts to build a comprehensive, multidisciplinary, academic program in women's health with initiatives in clinical care, research, faculty development, provider education, and community outreach. PMID- 10534300 TI - Lack of effect of short-term micronized progesterone on bone turnover in postmenopausal women. AB - A number of studies suggest that progestogens have beneficial effects on bone in postmenopausal women, particularly in combination with estrogen, although these studies have used derivatives that may have estrogenic and androgenic properties in addition to effects mediated by progesterone receptors. Progesterone itself affects only progesterone and glucocorticoid receptors. However, until the development of micronized progesterone (MP), absorption of progesterone preparations was too low to be clinically useful. MP has similar protective effects on the uterus and fewer effects on the lipid profile than other preparations, but its effects on bone are unknown. We tested the hypothesis that MP would alter bone turnover, as measured by serum and urine biochemical markers, in postmenopausal women. Fourteen women aged 65 or over who were not on estrogen replacement received a 6-week course of daily MP (200 mg). Markers of bone turnover were measured in serum and urine collected at baseline, at 6 weeks on MP, and 6 weeks after termination of MP. We also measured total and high-density lipoprotein (HDL) cholesterol and progesterone levels during the study. Markers of bone resorption were urinary free deoxypyridinoline cross-linked N telopeptides and C-telopeptides of type I collagen. Markers of bone formation were serum osteocalcin, bone alkaline phosphatase, and type I C-terminal and N terminal procollagen peptides. Using repeated measures analysis of variance, markers of bone formation and resorption did not change with MP treatment in spite of an increase in progesterone levels in all women. We conclude that 6-week treatment with MP alone does not have an effect on bone turnover in postmenopausal women in spite of high physiological levels. These data suggest that effects on bone demonstrated using other progestogen preparations might be due to androgenic or estrogenic effects or that progesterone may not affect bone in estrogen-deficient women. PMID- 10534302 TI - Lessons learned about research on premenstrual syndrome. AB - We discuss specific problems in implementing research to evaluate exercise treatment for premenstrual syndrome (PMS). Modifications of lifestyle, such as implementing exercise regimens, frequently are recommended as treatment for PMS, but evidence supporting this treatment is largely anecdotal. Originally, we designed a study to examine the effects of physical exercise on the symptoms of PMS. Despite initial enthusiasm, the majority of participants dropped out before beginning the active intervention segment of the study. This unexpected attrition resulted in a review of methodology, including recruitment and study design, in an attempt to understand factors related to research on exercise-based treatments of PMS so future researchers would be cognizant of the obstacles inherent in such research. Such understanding will allow research to advance more efficiently by enabling investigators to avoid the pitfalls we identified. PMID- 10534304 TI - Women's Health LiteratureWatch. PMID- 10534303 TI - WebWatch--Women's Health & Gender-Based Medicine. Eating disorders. PMID- 10534305 TI - Correlation between in vivo and in vitro hepatic uptake of metabolic inhibitors of cytochrome P-450 in rats. AB - To predict the degree of accumulation of hepatic metabolic inhibitors in the liver from the in vitro data, we investigated the relationship between cell/medium concentration ratios (C/M ratios) in isolated rat hepatocytes and liver/blood unbound concentration (K(Bf)) after i.v. administration of various metabolic inhibitors such as itraconazole, ketoconazole, verapamil, diltiazem, enoxacin, ciprofloxacin, clarithromycin, cimetidine, and nizatidine. The C/M ratios of itraconazole were approximately 6,000 and 200 at the concentrations of 0.1 and 10 microg/ml, respectively, and the uptake of ketoconazole and verapamil into the hepatocytes also showed a concentration dependence, although the degree was smaller than that of itraconazole. The uptake of diltiazem, enoxacin, ciprofloxacin, and clarithromycin into the hepatocytes showed linear profiles on concentration dependence. There was an excellent correlation between C/M ratios and K(Bf) values of all nine drugs with a slope of 1. This finding suggested the possibility of predicting drug concentrations in the liver (C(H)) from C/M ratios, the blood concentrations of drugs (C(B)) and unbound fraction in blood (f(B)), which was expressed by C(H) = (C/M). C(B). f(B). It may be possible to predict the drug concentrations in human liver from K(Bf) values estimated with isolated human hepatocytes and concentrations in the blood in a similar manner as in rats. PMID- 10534306 TI - Preclinical pharmacokinetics and interspecies scaling of a novel vitronectin receptor antagonist. AB - Allometric scaling may be used in drug development to predict the pharmacokinetics of xenobiotics in humans from animal data. Although allometry may be successful for compounds that are excreted unchanged or that are oxidatively metabolized (with corrections for metabolic capacity), it has been more challenging for compounds excreted primarily as conjugates in bile. (S)-10, 11-Dihydro-3-[3-(pyridin-2-ylamino)-1-propyloxy]-5H-dibenzo[ a, d]cycloheptene-10 acetic acid (SB-265123) is a novel alphavbeta3 ("vitronectin receptor") antagonist. In this study, the in vivo pharmacokinetics and in vitro plasma protein binding of SB-265123 were examined in four species: mice, rats, dogs, and monkeys. In monkeys and dogs, SB-265123 exhibited moderate clearance, whereas low clearance (<20% hepatic blood flow) was observed in the rat, and high clearance (>70% hepatic blood flow) was seen in the mouse. The concentration-time profiles indicated the possibility of enterohepatic recirculation; subsequent studies in bile duct-cannulated rats demonstrated extensive biliary excretion of an acyl glucuronide of SB-265123. In allometric scaling to predict the disposition of SB 265123 in humans, various standard correction factors were applied, including protein binding, maximum lifespan potential, and brain weight; each failed to produce adequate interspecies scaling of clearance (r(2) < 0.72). Consequently, a novel correction factor incorporating bile flow and microsomal UDP glucuronosyltransferase activity in each species was applied, demonstrating substantial improvement in the correlation of the allometric plot (r(2) = 0.96). This study demonstrates a novel allometric correction that may be applicable to compounds that undergo conjugation and biliary excretion. PMID- 10534307 TI - Effect of a ligand selective for peripheral benzodiazepine receptors on the expression of rat hepatic P-450 cytochromes: assessment of the effect in vivo and in a hepatocyte culture system. AB - The peripheral benzodiazepine receptor plays a role in the translocation of cholesterol into mitochondria where steroidogenesis occurs. Sterols have been suggested to be involved in the regulation of the cytochrome P-450 (CYP)2B subfamily as the endogenous suppressor of this CYP. To investigate the role of cholesterol metabolites on the expression of CYPs, the effect of PK11195, a specific ligand of the peripheral benzodiazepine receptor and a stimulator of cholesterol transportation, on CYP expression was examined in rats in vivo and in cultured hepatocytes. As judged by the change in testosterone metabolic activity catalyzed by liver microsomes, i.p. injection of PK11195 into rats increased the CYP2B subfamily significantly. A trend in the induction of the CYP2A1, 2C11, and 3A isozymes was also observed. When PK11195 was given to rats together with phenobarbital, an additive effect of these compounds on testosterone metabolic activity was observed. In cultured hepatocytes, PK11195 exhibited the same effect on CYP expression as seen in vivo, but the magnitude of the effect was much greater than that observed in vivo. The inductive effect of PK11195 toward the CYP2B and 3A subfamilies was 2.3- and 6.5-fold greater, respectively, than that with phenobarbital. The inductive effect of PK11195 was confirmed by immunoblotting with antibodies against CYP2A, 2B, 2C, and 3A proteins. These results indicate that PK11195 has an inductive effect on several subfamilies of CYPs by directly acting on liver cells and has no ability to suppress the expression of these CYPs. This observation suggests that, if certain sterols play a role in the suppressive control of the CYP2B subfamily, they are produced in organelles other than the mitochondria. PMID- 10534308 TI - Direct detection of antipyrine metabolites in rat urine by (13)C labeling and NMR spectroscopy. AB - Antipyrine is a useful probe to evaluate variation of in vivo activities of oxidative hepatic drug-metabolizing enzymes. Here we describe a new approach using (13)C labeling and NMR spectroscopy for the direct and simultaneous detection of all phase I and phase II metabolites of antipyrine in rat urine. [C methyl-(13)C]Antipyrine was synthesized and administered orally to rats (100 mg/kg), and the 0- to 24-h postdose urine was analyzed by 100-MHz (13)C NMR spectroscopy under the conditions of distortionless enhancement by polarization transfer without any pretreatments such as deconjugation, chromatographic separation, and solvent extraction. Consequently, all the major metabolites in urine were successfully detected with favorable signal-to-noise ratios in the limited acquisition time (30 min). The assignments of the resonances were performed by enzymic modification and spiking authentic samples. The reproducibility of the NMR detection was sufficient for the quantitative evaluation of the metabolic profile. Effects of 3-methylcholanthrene on antipyrine metabolism were examined by this approach to evaluate variation of in vivo phase I and phase II metabolism of antipyrine in rats. The present approach is useful and practical to evaluate variation of in vivo activities of conjugation enzymes as well as oxidation enzymes responsible for the formation of antipyrine metabolites in rats. This direct approach would enhance the value of the antipyrine test because of the simplicity and convenience. PMID- 10534309 TI - Identification of human cytochrome P-450 isoforms involved in metabolism of R(+)- and S(-)-gallopamil: utility of in vitro disappearance rate. AB - Isoforms of cytochrome P-450 (CYP) involved in the metabolism of gallopamil enantiomers were identified by measuring the disappearance rate of parent drug from an incubation mixture with human liver microsomes and recombinant human CYPs. Mean (+/- S.D.) intrinsic clearances (CL(int)) of R(+)- and S(-)-gallopamil in human liver microsomes were 0.320 +/- 0.165 and 0.205 +/- 0.107 ml/min/mg protein, respectively. These values were highly correlated with the 6beta hydroxylation activity of testosterone, a marker substrate of CYP3A4 (r = 0.977 and 0.900 for R(+)- and S(-)-gallopamil, respectively, p <.001). Ketoconazole and troleandomycin, selective inhibitors of CYP3A4, and polyclonal antibodies raised against CYP3A4/5 markedly reduced the CL(int) of gallopamil enantiomers in human liver microsomes. Among the 10 recombinant human CYP isoforms, CYP3A4 exhibited the highest CL(int) of gallopamil enantiomers, and CYP2C8 and CYP2D6 also exhibited appreciable activity. When the contribution of CYP3A4 to the total metabolic clearance of gallopamil enantiomers in human liver microsomes was estimated by relative activity factor, the mean (+/- S.D.) contributions were 92 +/- 18 and 68 +/- 19% for R(+)- and S(-)-gallopamil, respectively. These values were comparable to the rates of immunoinhibition by antibodies raised against CYP3A4/5 observed in human liver microsomes. The present study suggests that CYP3A4 is a major isoform involved in the overall metabolic clearance of gallopamil enantiomers in the human liver, and that the present approach based on disappearance rate may be applicable to identify major isoforms of CYP involved in the metabolism of a drug in human liver microsomes. PMID- 10534310 TI - Oxidation of troglitazone to a quinone-type metabolite catalyzed by cytochrome P 450 2C8 and P-450 3A4 in human liver microsomes. AB - Troglitazone, a new oral antidiabetic drug, is reported to be mostly metabolized to its conjugates and not to be oxidized by cytochrome P-450 (P-450) enzymes. Of fourteen cDNA-expressed human P-450 enzymes examined, CYP1A1, CYP2C8, CYP2C19, and CYP3A4 were active in catalyzing formation of a quinone-type metabolite at a concentration of 10 microM troglitazone, whereas CYP3A4 had the highest catalytic activity at 100 microM substrate. In human liver microsomes, rates of the quinone type metabolite formation (at 100 microM) were correlated well with rates of testosterone 6beta-hydroxylation (r = 0.98), but those at 10 microM troglitazone were not correlated with any of several marker activities of P-450 enzymes. Quercetin efficiently inhibited quinone-type metabolite formation (at 10 microM troglitazone) in human samples that contained relatively high levels of CYP2C, whereas ketoconazole affected these activities in liver microsomes in which CYP3A4 levels were relatively high. Anti-CYP2C antibodies strongly inhibited quinone-type metabolite formation (at 10 microM troglitazone) in CYP2C-rich human liver microsomes (by approximately 85%); the intensity of this effect depended on the human samples and their P-450 status. The results suggest that in human liver both CYP2C8 and CYP3A4 have major roles in quinone-type metabolite formation and that the hepatic contents of these two P-450 forms determine which P-450 enzymes play major roles in individual humans. CYP3A4 may be expected to play a role in formation of quinone-type metabolite from troglitazone even at a low concentration in humans. PMID- 10534311 TI - Identification of metabolites of octamethylcyclotetrasiloxane (D(4)) in rat urine. AB - Octamethylcyclotetrasiloxane (D(4)) is an industrial chemical of significant commercial importance. In this study, its major urinary metabolites were identified. The urine samples described here were collected from male and female Fischer rats (F-344) administered [(14)C]D(4) i.v. The metabolite profile was obtained using an HPLC system equipped with a radioisotope detector. HPLC analysis was performed on a C18 column, using an acetonitrile/water mobile phase. The HPLC radiochromatogram revealed two major and at least five minor metabolites. The two major metabolites, constituting 75 to 85% of the total radioactivity, were identified as dimethylsilanediol [Me(2)Si(OH)(2)] and methylsilanetriol [MeSi(OH)(3)]. Formation of MeSi(OH)(3) clearly established demethylation at the silicon-methyl bonds of D(4). No parent D(4) was present in urine. The minor metabolites identified were tetramethyldisiloxane-1,3-diol [Me(2)Si(OH)-O-Si(OH)Me(2)], hexamethyltrisiloxane-1,5-diol [Me(2)Si(OH)-OSiMe(2) OSi(OH)Me(2)], trimethyldisiloxane-1,3,3-triol [MeSi(OH)(2)-O-Si(OH)Me(2)], dimethyldisiloxane-1,1,3,3-tetrol [MeSi(OH)(2)-O-Si(OH)(2)Me], and dimethyldisiloxane-1,1,1,3,3-pentol [Si(OH)(3)-O-Si(OH)(2)Me]. The structural assignments were based on gas chromatography-mass spectrometry analysis of the tetrahydrofuran metabolite extracts, which were derivatized using bis(trimethylsiloxy)triflouroacetamide, a trimethylsilylating agent. The structures were confirmed by synthesizing (14)C-labeled standards and comparing their HPLC radiochromatograms with the corresponding components in the rat urine. GC-MS spectral comparisons of the trimethylsilylated derivatized standards and urinary components also were made to further confirm their identities. Finally, several of the urinary metabolites were fractionated using HPLC, and GC-MS comparisons were again made for positive structural identification. The pathways for metabolite formation are not yet understood, but a mechanistic hypothesis has been proposed to account for the various metabolites observed thus far. PMID- 10534312 TI - Prediction of human liver microsomal oxidations of 7-ethoxycoumarin and chlorzoxazone with kinetic parameters of recombinant cytochrome P-450 enzymes. AB - Different roles of individual forms of human cytochrome P-450 (CYP) in the oxidation of 7-ethoxycoumarin and chlorzoxazone were investigated in liver microsomes of different human samples, and the microsomal activities thus obtained were predicted with kinetic parameters obtained from cDNA-derived recombinant CYP enzymes in microsomes of Trichoplusia ni cells. Of 14 forms of recombinant CYP examined, CYP1A1 had the highest activities (V(max)/K(m) ratio) in catalyzing 7-ethoxycoumarin O-deethylation followed by CYP1A2, 2E1, 2A6, and 2B6, although CYP1A1 has been shown to be an extrahepatic enzyme. With these kinetic parameters (excluding CYP1A1) we found that CYP1A2 and 2E1 were the major enzymes catalyzing 7-ethoxycoumarin; the contributions of these two forms were dependent on the contents of these CYPs in liver microsomes of different humans. Similarly, chlorzoxazone 6-hydroxylation activities of liver microsomes were predicted with kinetic parameters of recombinant human CYP enzymes and it was found that CYP3A4 as well as CYP1A2 and 2E1 were involved in chlorzoxazone hydroxylation, depending on the contents of these CYP forms in the livers. Recombinant CYP2A6 and 2B6 and CYP2D6 had considerable roles (V(max)/K(m) ratio) for 7-ethoxycoumarin O-deethylation and chlorzoxazone 6-hydroxylation, respectively; however, these CYP forms had relatively minor roles in the reactions, probably due to low expression in human livers. These results support the view that the roles of individual CYP enzymes in the oxidation of xenobiotic chemicals in human liver microsomes could be predicted by kinetic parameters of individual CYP enzymes and by the levels of each of the CYP enzymes in liver microsomes of human samples. PMID- 10534313 TI - Conjugation of the enantiomers of ketotifen to four isomeric quaternary ammonium glucuronides in humans in vivo and in liver microsomes. AB - The antiallergic drug ketotifen is chiral due to a nonplanar seven-membered ring containing a keto group. Earlier studies have revealed glucuronidation at the tertiary amino group as a major metabolic pathway in humans. Chemical synthesis of glucuronides from racemic ketotifen now led to four isomers separable by HPLC of which two each could be ascribed to (R)-(+)- and (S)-(-)-ketotifen by synthesis from the enantiomers. According to (1)H NMR analysis of the (S) ketotifen N-glucuronides, the conformation of the piperidylidene ring differs between the two isomers. Enzymatic hydrolysis with Escherichia coli beta glucuronidase proceeded at a lower rate with the slower eluting (S)-ketotifen glucuronide than with the three other isomers. On incubation of the ketotifen enantiomers (0.5-200 microM) with human liver microsomes in the presence of UDP glucuronic acid and Triton X-100, the N-glucuronides of (R)-ketotifen were produced with an apparent K(M) 15 microM and V(max) 470 pmol/min/mg protein. The two (S)-ketotifen glucuronides were formed by two-enzyme kinetics with K(M1) 1.3 microM and K(M2) 92 microM and V(max) values of 60 and 440 pmol/min/mg protein. After ingestion of 1 mg of racemic ketotifen, 10 healthy subjects excreted in urine 17 +/- 5% of the dose in the form of N-glucuronides. The (R)-ketotifen glucuronide isomers contributed one-sixth only, whereas the remainder consisted primarily of the (S)-ketotifen glucuronide isomer, which eluted last. Differential hydrolysis or membrane transport may be responsible for the discrepancy between N-glucuronide isomer ratios in vitro and in vivo. PMID- 10534314 TI - Propofol hydroxylation by dog liver microsomes: assay development and dog breed differences. AB - Pharmacokinetic studies indicate that clearance of propofol, an anesthetic agent, is slower in greyhounds compared with other dog breeds. Biotransformation of propofol to 2,6-diisopropyl-1,4-quinol (4-hydroxypropofol) by cytochrome P-450 in the liver is proposed as a critical initial step in the elimination of this drug in dogs. Breed differences in the activity of this enzyme could therefore explain pharmacokinetic differences. An in vitro propofol hydroxylase assay was developed and then used to compare enzyme activities in liver microsomes from male greyhound, beagle, and mixed-breed dogs (five each). HPLC of incubate identified only one NADPH-dependent metabolite, which had a chromatographic retention time and UV absorbance, fluorescence, and mass spectra that were identical with authentic 4-hydroxypropofol standard. HPLC with fluorescence detection provided a highly sensitive quantitation method for 4-hydroxypropofol with a quantitation limit of 8 ng/ml using optimized excitation/emission wavelengths (288 nm/330 nm, respectively). Estimates of apparent K(m) and V(max) for propofol hydroxylation by microsomes from a male beagle dog were 7.3 microM and 3.8 nmol/mg/min, respectively. At a substrate concentration of 20 microM, propofol hydroxylase activity was significantly lower (p =.032) in greyhound microsomes (1.7 +/- 0.4 nmol/mg/min) compared with beagle microsomes (5.1 +/- 1.3 nmol/mg/min) but was not statistically different (p =.42) compared with mixed-breed microsomes (3.1 +/ 1.2 nmol/mg/min). These results indicate that there are breed differences in propofol hydroxylase activity and that deficient hydroxylation of propofol by one or more hepatic cytochrome P-450 isoforms may contribute to slow pharmacokinetic clearance of propofol by greyhounds. PMID- 10534315 TI - Disposition of [G-(3)H]paclitaxel and cremophor EL in a patient with severely impaired renal function. AB - In the present work, we studied the pharmacokinetics and metabolic disposition of [G-(3)H]paclitaxel in a female patient with recurrent ovarian cancer and severe renal impairment (creatinine clearance: approximately 20 ml/min) due to chronic hypertension and prior cisplatin treatment. During six 3-weekly courses of paclitaxel at a dose level of 157.5 mg/m(2) (viz. a 10% dose reduction), the renal function remained stable. Pharmacokinetic evaluation revealed a reproducible and surprisingly high paclitaxel area under the plasma concentration time curve of 26.0 +/- 1.11 microM.h (mean +/- S.D.; n = 6; c.v. = 4.29%), and a terminal disposition half-life of approximately 29 h. Both parameters are substantially increased ( approximately 1.5-fold) when compared with kinetic data obtained from patients with normal renal function. The cumulative urinary excretion of the parent drug was consistently low and averaged 1.58 +/- 0.417% (+/- S.D.) of the dose. Total fecal excretion (measured in one course) was 52.9% of the delivered radioactivity, and mainly comprised known mono- and dihydroxylated metabolites, with unchanged paclitaxel accounting for only 6.18%. The plasma area under the plasma concentration-time curve of the paclitaxel vehicle Cremophor EL, which can profoundly alter the kinetics of paclitaxel, was 114.9 +/- 5.39 microl.h/ml, and not different from historic data in patients with normal or mild renal dysfunction. Urinary excretion of Cremophor EL was less than 0.1% of the total amount administered. These data indicate that the substantial increase in systemic exposure of the patient to paclitaxel relates to decreased renal metabolism and/or urinary elimination of polar radioactive species, most likely lacking an intact taxane ring fragment. PMID- 10534316 TI - Catalysis of drug oxidation during embryogenesis in human hepatic tissues using imipramine as a model substrate. AB - We investigated the catalysis of drug monooxygenation by human embryonic hepatic tissues at a very early stage of gestation (days 52-59). Imipramine was used as a model substrate and the metabolites generated were identified and quantified by electrospray mass spectroscopy and HPLC. The primary metabolite generated was desipramine. It was reported previously from this and other laboratories that cytochrome P-450 monooxygenase (CYP) 1A1, 1B1, 2E1, and 3A7 are each expressed in human embryonic hepatic tissues, and selective inhibitors were therefore used to elucidate their respective roles. Furafylline did not inhibit the reaction, supporting that CYP1A2 was not expressed in human embryonic hepatic tissues. Diethyldithiocarbamate also failed to inhibit the same reaction, suggesting that CYP2E1 did not play a significant role in catalyzing the reaction. Triacetyloleandomycin inhibited the reaction by approximately 90%, suggesting that CYP3A7 was primarily responsible for catalyzing the reaction. However, alpha naphthoflavone inhibited the same reaction by approximately 70%, suggesting that CYP1A1 and/or CYP1B1 may also catalyze the reaction substantially. To explore this issue more, a cDNA-expressed human CYP3A7 (CYP3A7 SUPERSOMES) was incubated with alpha-naphthoflavone (1 microM). Generation of desipramine was inhibited by approximately 40 to 50%. The addition of the CYP3A subfamily selective inhibitor triacetyloleandomycin (1 microM) produced no statistically significant inhibition in reactions catalyzed by CYP1A1 or 1B1 SUPERSOMES. Taken together, the results indicated that CYP3A7 was the major if not sole isoform responsible for catalysis of the N-demethylation of imipramine in human hepatic tissues during embryogenesis. PMID- 10534317 TI - Stereoselective metabolism of ifosfamide by human P-450s 3A4 and 2B6. Favorable metabolic properties of R-enantiomer. AB - The anticancer prodrug ifosfamide (IFA) contains a chiral phosphorous atom and is administered clinically as a racemic mixture of R and S enantiomers. Animal model studies and clinical data indicate enantioselective differences in cytochrome P 450 (CYP) metabolism, pharmacokinetics, and therapeutic efficacy between the two enantiomers; however, the metabolism of individual IFA enantiomers has not been fully characterized. The role of CYP enzymes in the stereoselective metabolism of R-IFA and S-IFA was investigated by monitoring the formation of both 4-hydroxy (activated) and N-dechloroethyl (DCl) (inactive, neurotoxic) metabolites. In the 4-hydroxylation reaction, cDNA-expressed CYPs 3A4 and 3A5 preferentially metabolized R-IFA, whereas CYP2B6 was more active toward S-IFA. Enantioselective IFA 4-hydroxylation (R > S) was observed with six of eight human liver samples. In the N-dechloroethylation reaction, CYPs 3A4 and 2B6 both catalyzed a significantly higher intrinsic metabolic clearance (V(max)/K(m)) of S-IFA compared with R-IFA. Striking P-450 form specificity in the formation of individual DCl metabolites was evident. CYPs 3A4 and 3A5 preferentially produced (R)N2-DCl-IFA and (R)N3-DCl-IFA (derived from R-IFA and S-IFA, respectively), whereas CYP2B6 correspondingly formed (S)N3-DCl-IFA and (S)N2-DCl-IFA. In human liver microsomes, the CYP3A-specific inhibitor troleandomycin suppressed (R)N2- and (R)N3-DCl-IFA formation by >/=80%, whereas (S)N2- and (S)N3-DCl-IFA formation were selectively inhibited (>/=85%) by a CYP2B6-specific monoclonal antibody. The overall extent of IFA N-dechloroethylation varied with the CYP3A4 and CYP2B6 content of each liver, but was significantly lower for R-IFA (32 +/- 13%) than for S-IFA (62 +/- 17%, n = 8; p <.001) in all livers examined. R-IFA thus has more favorable liver metabolic properties than S-IFA with respect to less extensive N-dechloroethylation and more rapid 4-hydroxylation, indicating that R IFA may have a distinct clinical advantage over racemic IFA. PMID- 10534318 TI - Identification and characterization of efavirenz metabolites by liquid chromatography/mass spectrometry and high field NMR: species differences in the metabolism of efavirenz. AB - Efavirenz (Sustiva, Fig. 1) is a potent and specific inhibitor of HIV-1 reverse transcriptase approved for the treatment of HIV infection. To examine the potential differences in the metabolism among species, liquid chromatography/mass spectrometry profiles of efavirenz metabolites in urine of rats, guinea pigs, hamsters, cynomolgus monkeys, and humans were obtained and compared. The metabolites of efavirenz were isolated, and structures were determined unequivocally by mass spectral and NMR analyses. Efavirenz was metabolized extensively by all the species as evidenced by the excretion of none or trace quantities of parent compound in urine. Significant species differences in the metabolism of efavirenz were observed. The major metabolite excreted in the urine of all species was the O-glucuronide conjugate (M1) of the 8-hydroxylated metabolite. Efavirenz was also metabolized by direct conjugation with glucuronic acid, forming the N-glucuronide (M2) in all five species. The sulfate conjugate of 8-OH efavirenz (M3) was found in the urine of rats and cynomolgus monkeys but not in humans. In addition to the aromatic ring-hydroxylated products, metabolites with a hydroxylated cyclopropane ring (at C14) were also isolated. GSH-related products of efavirenz were identified in rats and guinea pigs. The cysteinylglycine adduct (M10), formed from the GSH adduct (M9), was found in significant quantities in only rat and guinea pig urine and was not detected in other species. In vitro metabolism studies were conducted to show that the GSH adduct was produced from the cyclopropanol intermediate (M11) in the presence of only rat liver and kidney subcellular fractions and was not formed by similar preparations from humans or cynomolgus monkeys. These studies indicated the existence of a specific glutathione-S-transferase in rats capable of metabolizing the cyclopropanol metabolite (M11) to the GSH adduct, M9. The biotransformation pathways of efavirenz in different species were proposed based on some of the in vitro results. PMID- 10534319 TI - Cytochrome P-450-mediated metabolism of the individual enantiomers of the antidepressant agent reboxetine in human liver microsomes. AB - In vitro studies were conducted to identify the hepatic cytochrome P-450 (CYP) enzymes responsible for the oxidative metabolism of the individual enantiomers of reboxetine. In human liver microsomes, each reboxetine enantiomer was metabolized to one primary metabolite, O-desethylreboxetine, and three minor metabolites, two arising via oxidation of the ethoxy aromatic ring and a third yet unidentified metabolite. Over a concentration range of 2 to 200 microM, the rate O desethylreboxetine formation for either enantiomer conformed to monophasic Michaelis-Menten kinetics. Evidence for a principal role of CYP3A in the formation of O-desethylreboxetine for (S, S)-reboxetine and (R,R)-reboxetine was based on the results from the following studies: 1) inhibition of CYP3A activity by ketoconazole markedly decreased the formation of O-desethylreboxetine, whereas inhibitors selective for other CYP enzymes did not inhibit reboxetine metabolism, 2) formation of O-desethylreboxetine correlated (r(2) = 0.99; p <.001) with CYP3A selective testosterone 6-beta-hydroxylase activity across a population of human livers (n = 14). Consistent with inhibition and correlation data, O desethylreboxetine formation was only detectable in incubations using microsomes prepared from a Baculovirus-insect cell line expressing CYP3A4. Furthermore, the apparent K(M) for the O-desethylation of reboxetine in cDNA CYP3A4 microsomes was similar to the affinity constants determined in human liver microsomes. In addition, (S,S)-reboxetine and (R,R)-reboxetine were found to be competitive inhibitors of CYP2D6 and CYP3A4 (K(i) = 2.5 and 11 microM, respectively). Based on the results of the study, it is concluded that the metabolism of both reboxetine enantiomers in humans is principally mediated via CYP3A. PMID- 10534321 TI - Prediction of human clearance of twenty-nine drugs from hepatic microsomal intrinsic clearance data: An examination of in vitro half-life approach and nonspecific binding to microsomes. AB - Twenty-nine drugs of disparate structures and physicochemical properties were used in an examination of the capability of human liver microsomal lability data ("in vitro T(1/2)" approach) to be useful in the prediction of human clearance. Additionally, the potential importance of nonspecific binding to microsomes in the in vitro incubation milieu for the accurate prediction of human clearance was investigated. The compounds examined demonstrated a wide range of microsomal metabolic labilities with scaled intrinsic clearance values ranging from less than 0.5 ml/min/kg to 189 ml/min/kg. Microsomal binding was determined at microsomal protein concentrations used in the lability incubations. For the 29 compounds studied, unbound fractions in microsomes ranged from 0.11 to 1.0. Generally, basic compounds demonstrated the greatest extent of binding and neutral and acidic compounds the least extent of binding. In the projection of human clearance values, basic and neutral compounds were well predicted when all binding considerations (blood and microsome) were disregarded, however, including both binding considerations also yielded reasonable predictions. Including only blood binding yielded very poor projections of human clearance for these two types of compounds. However, for acidic compounds, disregarding all binding considerations yielded poor predictions of human clearance. It was generally most difficult to accurately predict clearance for this class of compounds; however the accuracy was best when all binding considerations were included. Overall, inclusion of both blood and microsome binding values gave the best agreement between in vivo clearance values and clearance values projected from in vitro intrinsic clearance data. PMID- 10534320 TI - Metabolites of [(14)C]-5-(2-ethyl-2H-tetrazol-5-yl)-1-methyl-1,2,3, 6 tetrahydropyridine in mice, rats, dogs, and humans. AB - The M1 muscarine agonist, 5-(2-ethyl-2H-tetrazol-5-yl)-1-methyl-1,2, 3,6 tetrahydropyridine (Lu 25-109), is extensively metabolized in mice, rats, dogs, and humans. The metabolite profile after an oral dose of [(14)C]Lu 25-109 was determined in plasma and in urine. Lu 25-109 was metabolized by N-demethylation (Lu 25-077), N-oxidation (Lu 32-181), and N-deethylation (Lu 31-126). In addition, combined N-demethylation and N-deethylation (Lu 31-190), and formation of a pyridine derivative took place (Lu 31-102). Lu 25-109 was also oxidized to pyridinium (Lu 29-297), 3-hydroxy-pyridinium (Lu 35-080), N-deethyl-2-pyridone (Lu 35-026), and a glucuronide of a 4, 6-dihydroxy-pyridinium ("m/z 398") compounds. A glucuronide of a dihydroxylated dihydro-pyridine compound ("m/z 400") was isolated from human urine, but not fully identified. In vitro studies were undertaken to elucidate the order of formation of the metabolites. In human plasma, the concentrations of Lu 25-109 and the pharmacologically active N demethyl metabolite (Lu 25-077) were small compared with the N-oxide (Lu 32-181) and the N-deethyl-2-pyridone (Lu 35-026) at the first sample time (0.75 h). The N deethyl metabolite (Lu 31-126) was the major component in human plasma between 3 and 10 h postdose. The major human metabolites in urine (Lu 32-181, Lu 35-026, and Lu 31-126) and the minor metabolites (Lu 25-077, Lu 35-080, Lu 31-190, and Lu 29-297) were all present in urine from rats, dogs, and mice, whereas m/z 398 was present in only mice and humans, and Lu 31-102 in only rats. The minor human metabolite m/z 400 was not detected in mice, rats, or dogs. PMID- 10534322 TI - In vitro and in vivo studies on the metabolism of tirofiban. AB - Tirofiban hydrochloride [L-tyrosine-N-(butylsulfonyl)-O-[4-(4-piperidinebutyl)] monohydrochloride, is a potent and specific fibrinogen receptor antagonist. Radiolabeled tirofiban was synthesized with either (3)H-label incorporated into the phenyl ring of the tyrosinyl residue or (14)C-label in the butane sulfonyl moiety. Neither human liver microsomes nor liver slices metabolized [(14)C]tirofiban. However, male rat liver microsomes converted a limited amount of the substrate to a more polar metabolite (I) and a relatively less polar metabolite (II). The formation of I was sex dependent and resulted from an O dealkylation reaction catalyzed by CYP3A2. Metabolite II was identified as a 2 piperidone analog of tirofiban. There was no evidence for Phase II biotransformation of tirofiban by microsomes fortified with uridine-5'-diphospho alpha-D-glucuronic acid. After a 1 mg/kg i.v. dose of [(14)C]tirofiban, recoveries of radioactivity in rat urine and bile were 23 and 73%, respectively. Metabolite I and unchanged tirofiban represented 70 and 30% of the urinary radioactivity, respectively. Tirofiban represented >90% of the biliary radioactivity. At least three minor biliary metabolites represented the remainder of the radioactivity. One of them was identified as I. Another was identified as II. When dogs received 1 mg/kg i.v. of [(3)H]tirofiban, most of the radioactivity was recovered in the feces as unchanged tirofiban. The plasma half-life of tirofiban was short in both rats and dogs, and tirofiban was not concentrated in tissues other than those of the vasculature and excretory organs. PMID- 10534323 TI - Substance P receptor antagonist I: conversion of phosphoramidate prodrug after i.v. administration to rats and dogs. AB - A water-soluble phosphoramidate prodrug (L-758,298, compound I) of the potent and selective human Substance P receptor antagonist L-754, 030 (compound II) is under development as an i.v. drug for treatment of emesis, migraine, and chronic pain. Compound I undergoes hydrolysis readily to II under acidic conditions. In the studies reported herein, we investigated the stability of I in blood and hepatic subcellular fractions from rats, dogs, and humans as well as the conversion of I to II in rats and dogs after i.v. dosing. Compound I was converted to II rapidly in rat blood but was stable in dog and human blood. However, the conversion was rapid in liver microsomes prepared from dogs and humans. As expected from the results of in vitro studies, the in vivo conversion of I to II was rapid after i.v. dosing of I to rats and dogs. The relative extent of exposure of II after i.v. dosing of I was estimated by comparing the dose-adjusted area under the plasma concentration versus time curve values of II after i.v. dosing of I with those after i.v. dosing of II. In rats, the extent of exposure was estimated to be approximately 90 and approximately 100% at 1 and 8 mg/kg, respectively; in dogs, that was approximately 59% at 0.5 mg/kg. A nonproportional increase in the area under the concentration versus time curve value of II with dose was observed after i.v. administration of I in dogs from 0.5 to 32 mg/kg, suggesting that the elimination of II might have been saturated at higher doses. PMID- 10534325 TI - Echocardiographic pitfalls in the diagnosis of hypertrophic cardiomyopathy. PMID- 10534324 TI - Pathophysiology of myocardial hibernation. Implications for the use of dobutamine echocardiography to identify myocardial viability. PMID- 10534326 TI - Clinical and economic outcomes assessment with myocardial contrast echocardiography. PMID- 10534329 TI - Dissection of the multifunctional "Receptor-Recycling" endocytic compartment of hepatocytes. PMID- 10534327 TI - Added value of contrast echocardiography in assessing myocardial viability. PMID- 10534330 TI - Long-term results of patients undergoing liver transplantation for primary sclerosing cholangitis. AB - Liver transplantation is the only effective therapeutic option for patients with end-stage liver disease due to primary sclerosing cholangitis (PSC). In this study, we analyzed a single center's experience with 150 consecutive PSC patients who received 174 liver allografts. Mean follow-up was 55 months. Actuarial patient survival at 1, 2, 5, and 10 years was 93.7%, 92.2%, 86.4%, and 69.8%, respectively, whereas graft survival was 83.4%, 83.4%, 79.0%, and 60. 5%, respectively. The main indication for retransplantation was hepatic artery thrombosis, and the major cause of death was severe infection. Patients with PSC had a higher incidence of acute cellular and chronic ductopenic rejection compared to a non-PSC control group. Chronic ductopenic rejection adversely affected patient and graft survival. Biliary strictures, both anastomotic and nonanastomotic, were frequent and occurred in 16.2% and 27.2% of patients, respectively. The incidence of recurrent PSC was 20%. A negative impact on patient survival was not seen in patients with either postoperative biliary strictures or recurrence of PSC. Six patients (4%) had cholangiocarcinoma and 1 patient died related to recurrence of malignant disease. Seventy-eight percent of PSC patients had associated inflammatory bowel disease, most commonly chronic ulcerative colitis, which did not adversely impact patient outcome posttransplantation. Nine patients required proctocolectomy after liver transplantation; 5 because of intractable symptoms related to inflammatory bowel disease and 4 due to the development of colorectal carcinoma/high-grade dysplasia. Our data show that liver transplantation provides excellent long-term patient and graft survival for patients with end-stage PSC. PMID- 10534331 TI - Inhibition of interleukin 6-mediated mitogen-activated protein kinase activation attenuates growth of a cholangiocarcinoma cell line. AB - Biliary tract malignancies represent challenges because of the lack of effective therapy and poor prognosis, in part because of the paucity of information regarding the mechanisms regulating their growth. We have recently identified a critical role for the p44/p42 mitogen-activated protein kinase (MAPK) pathway in interleukin 6 (IL-6)-stimulated growth of human cholangiocytes. Although IL-6 is a potential mitogen for cholangiocarcinoma, the role of this cytokine and its intracellular signaling pathways in cholangiocarcinoma growth is unknown. Thus, our aims were to determine the role of IL-6-mediated signaling mechanisms, and in particular the MAPK pathways, in the growth regulation of human cholangiocarcinoma. KMCH-1 cells (malignant cholangiocyte cells) secreted IL-6 constitutively, and increased IL-6 secretion in response to inflammatory cytokines such as tumor necrosis factor alpha (TNF-alpha) and IL-1beta. Stimulation with IL-6 resulted in proliferation of malignant cholangiocytes. These cells also possessed the IL-6 receptor complex subunits as directly assessed by immunoblot analysis. Furthermore, proliferation was completely inhibited by preincubation with anti-IL-6 neutralizing antibodies, indicating that the proliferative response to IL-6 involved receptor-mediated signaling. Both p38 and p44/p42 MAPKs were constitutively present and active in malignant cholangiocytes, and increased activity of both was observed within 15 minutes of stimulation with IL-6. Selective inhibition of either the p44/p42 MAPK pathway, by PD098059, or of the p38 MAPK pathway, by SB203580, blocked proliferation in response to IL-6. Thus, IL-6 can contribute to the autocrine and/or paracrine growth stimulation of malignant cholangiocytes via activation of either p38 or p44/p42 MAPK signaling pathways. PMID- 10534332 TI - Randomized clinical trials in HEPATOLOGY: predictors of quality. AB - Evidence shows that the quality of randomized clinical trials (RCTs) affects estimates of intervention efficacy, which is significantly exaggerated in low quality trials. The present study examines the quality of all 235 RCTs published in HEPATOLOGY from the initiation in 1981 through August 1998. Quality was assessed by means of a validated 5-point scale and separate quality components associated with empirical evidence of bias. Only 26% of all RCTs reported sample size calculations, 52% adequate generation of the allocation sequence, 34% adequate allocation concealment and 34% double-blinding. The median quality score of all trials was 3 points (range, 1-5 points). Multiple logistic regression analysis explored the association between quality and therapeutic areas, number of centers, external funding, year of publication, and country of origin. High quality trials were most likely to investigate portal hypertension (odds ratio [OR]: 2.4; 95% CI: 1.1-5.5; P =.03), be multicentered (OR: 3.4; 95% CI: 1.3-8.9; P =.01), sponsored by public organizations (OR: 4.2; 95% CI: 2.1-8.6; P =.0001), or the drug and device industry (OR: 4.7; 95% CI: 2.2-10.2; P =.0001) compared with other therapeutic areas, single-center trials, and trials with no external funding. Quality did not improve with time and was not associated with country of origin. The main conclusions are that the quality of RCTs in HEPATOLOGY needs improvement and that the probability of high quality increased with the number of centers involved and external funding. PMID- 10534333 TI - Transjugular intrahepatic portosystemic shunt in the treatment of refractory bleeding from ruptured gastric varices. AB - The optimal management of ruptured gastric varices in patients with cirrhosis has not been codified yet. The present study reports the use of transjugular intrahepatic portosystemic shunt (TIPS) in patients with refractory gastric variceal bleeding. Thirty-two consecutive patients were included. All had been unresponsive to vasoactive agents infusion, sclerotherapy, and/or tamponade and were considered poor surgical candidates. They were followed-up until death, transplantation, or at least 1 year (median: 509 days; range 4 to 2,230). Hemostasis was achieved in 18 out of 20 patients actively bleeding at the time of the procedure. In the whole sample of 32 patients, rebleeding rates were 14%, 26%, and 31%, respectively at 1 month, 6 months, and 1 year. De novo encephalopathy was observed in 5 (16%) patients. Seven patients experienced complications and consequently 4 of these patients died. TIPS primary patency rates were 84%, 74%, and 51%, respectively, at 1 month, 6 months, and 1 year. For the same periods of time, survival rates were 75%, 62%, and 59%. These results suggest that TIPS can be used in cirrhotic patients with refractory gastric variceal bleeding and are effective in achieving hemostasis as well as in preventing rebleeding. PMID- 10534334 TI - "Silent" presentation of veno-occlusive disease after liver transplantation as part of the process of cellular rejection with endothelial predilection. AB - Hemorrhagic centrilobular necrosis and fibrous stenosis of hepatic venules, suggesting veno-occlusive disease (VOD) have rarely been observed after orthotopic liver transplantation (OLT). The aim of this study was to determine the prevalence of this syndrome after OLT in relation to the course with particular reference to acute rejection and to azathioprine administration. VOD was identified in 19 of 1,023 patients transplanted over a 9-year period. VOD occurred at a median of 30 days posttransplantation, without clear cut clinical evidence for hepatic vein outlet obstruction. Seventeen of the 19 patients had an episode of acute rejection before or at the time of VOD. These episodes were compared with that of patients without VOD. In patients with VOD, portal inflammation and endothelialitis were enhanced (P =.014 and P =.048) and endothelialitis was also higher than bile duct damage (P =.03). The incidence of a centrilobular endothelialitis for both groups was not different although an increased trend was observed in the study group (64% vs. 46%; P =.18). The incidence of persistent rejection was similar between both groups (47% vs. 41%). The incidence of chronic rejection was higher in the study group (29% vs. 10%; P =. 04). All patients with VOD received azathioprine as part of immunosuppressive regimen. Despite azathioprine withdrawal, zone 3 changes persisted in 57% of patients. In conclusion, the incidence of VOD was 1.9% after OLT. The association of prominent endothelial involvement and VOD with acute rejection in most cases suggests an immunological phenomenon. PMID- 10534335 TI - Natural history of liver disease in cystic fibrosis. AB - The median age of the population with cystic fibrosis (CF) has increased worldwide, which has led to the suggestion that the prevalence of liver disease would increase. The aim of this study was to evaluate the natural history of CF associated liver disease over a 15-year period in a well-controlled population of patients with CF. During the years 1976 through 1993, 124 patients were followed up by yearly liver function tests (LFTs). Fifteen patients were followed up with liver biopsies throughout the whole study period. More than 50% of the patients had pathological LFTs in infancy, later being normalized. Approximately 25% of children 4 years of age or older had biochemical markers of liver disease during the study period. In about 10% of the patients, cirrhosis or advanced fibrosis was confirmed at biopsy and 4% of patients had cirrhosis with clinical liver disease. Severe liver disease developed mainly during prepuberty and puberty. Of the 15 patients prospectively followed up with liver biopsies, only 3 had progressive fibrosis. No specific risk factor was identified, but deficiency of essential fatty acids was found more often in patients with marked steatosis (P <.05). No patient developed clinical liver disease in adulthood and the histological changes in the liver biopsies were usually not progressive. Liver disease was no more frequent at the end of the study period although the median age of the patient population had increased. Modern treatment might positively influence liver disease because it seemed less common, less progressive, and less serious than previously reported. PMID- 10534336 TI - Differential regulation of extracellular matrix synthesis during liver regeneration after partial hepatectomy in rats. AB - Little is known about the modulation of the extracellular matrix (ECM) during liver regeneration. We studied the temporospatial expression of procollagens and of matrix metalloproteinases (MMPs) and their physiological antagonists, the tissue inhibitors of metalloproteinases (TIMPs) after two-thirds partial hepatectomy (PH) by Northern blot analysis and in situ hybridization. The entry of hepatocytes into the S-phase at 24 hours after PH was accompanied by a peak (sixfold induction) of hepatic TIMP-1 RNA levels that steadily declined thereafter to reach normal levels 144 hours after PH. Moderate MMP-2 and TIMP-2 RNA levels remained constant up to 144 hours after PH, and MMP-1 and -13 RNA were always undetectable. In situ hybridization showed a dramatic upregulation of TIMP 1 RNA transcripts in mesenchymal cells of portal, perisinusoidal and, to a lesser extent, pericentral areas. In contrast, scattered hepatocytes represented only a minor fraction (below 10%) of TIMP-1 RNA positive cells. When hepatocytes stopped DNA synthesis at 72 hours after PH, an upregulation of procollagen alpha1(I) and alpha2(III) transcripts was observed paralleled by threefold increased PIIINP levels in the sera. Our data reveal a tightly regulated program of de novo matrix synthesis after PH. Whereas interstitial procollagens appear to participate in the induction and maintenance of the quiescent hepatocyte phenotype, the early and localized expression of TIMP-1 indicates a role unrelated to its function as a general MMP-antagonist, e.g., as a growth promoting agent for hepatocytes. PMID- 10534337 TI - Decreased p27(Kip1) expression and cyclin D1 overexpression, alone and in combination, influence recurrence and survival of patients with resectable extrahepatic bile duct carcinoma. AB - This study was undertaken to identify potential abnormalities of p27(Kip1) and cyclin D1 expression in extrahepatic bile duct carcinomas and to assess the prognostic significance of p27(Kip1) and cyclin D1 levels for patients with this disease. Decreased p27(Kip1) expression (<50% nuclei staining) and cyclin D1 overexpression (>5% nuclei staining) was observed immunohistochemically in 19 (56%) and 23 (68%) of the 34 tumors examined, respectively. Both decreased p27(Kip1) and cyclin D1 overexpression were associated with relapse (P =.0005 for p27(Kip1) and P =.0004 for cyclin D1). Kaplan-Meier curves showed that both decreased p27(Kip1) and cyclin D1 overexpression correlate significantly with shortened survival rates (for p27(Kip1), P =.0419 and P =.002 for overall and disease-free survival; for cyclin D1, P =.0392 and P =.0021 for overall and disease-free survival). Cox regression model analyses identified decreased p27(Kip1) and cyclin D1 overexpression as independent markers predicting death from relapse (P =.0371, risk ratio: 3.891 for p27(Kip1); P =.0429, risk ratio: 8.31 for cyclin D1). Decreased p27(Kip1) was associated with cyclin D1 overexpression (P =.0202), and coincident abnormalities of the 2 proteins occurred in 16 of the 34 (47%) tumors, indicating that extrahepatic bile duct carcinoma progression may require synchronous dysfunction of p27(Kip1) and cyclin D1 in about half of patients. Patients with tumors showing coincident abnormalities of p27(Kip1) and cyclin D1 showed even more frequent recurrence than patients with an alteration in only 1 of the 2 proteins. In conclusion, decreased p27(Kip1) expression and cyclin D1 overexpression, alone and in combination, predict poor prognosis in patients with resectable extrahepatic bile duct carcinoma. PMID- 10534338 TI - The vascular profile of regenerative and dysplastic nodules of the cirrhotic liver: implications for diagnosis and classification. AB - We investigated the angiogenic phenotype of regenerative and dysplastic hepatocellular nodules to assess whether these lesions have distinct vascular profiles compared with the adjacent nonneoplastic or malignant liver. Forty-three liver nodules surgically removed from 18 patients were classified into regenerative and dysplastic categories. Serial sections of each nodule, adjacent cirrhotic liver (16 patients), and associated hepatocellular carcinoma (HCC) (6 patients), have been immunostained against CD31 and alpha-smooth muscle actin (alphaSMA) to detect capillary and muscular vessels. The study included 20 large regenerative nodules (LRNs), 13 low-grade dysplastic nodules (LGDNs), and 10 high grade dysplastic nodules (HGDNs). The number of both capillary units and unpaired arteries was significantly increased in HGDNs and malignant lesions over LGDNs, regenerative, and cirrhotic nodules (P <.01), which showed an overlapping vascular profile. In addition, the number of capillary units, but not that of unpaired arteries, was significantly increased in HCC compared with HGDNs (P <.01). These results show that certain angiogenic features segregate HGDNs from other nonmalignant nodules such as LRNs and LGDNs. The former group of lesions is similar to HCC whereas the latter group is undistinguishable from the adjacent cirrhosis as far as their vascular profile is concerned. The adopted investigative approach does not support the morphological distinction between LRNs and LGDNs although it suggests that HGDNs are likely advanced precursors of HCC. An abnormal number of capillary units and/or unpaired arteries in a nonmalignant hepatocellular nodule can be diagnostically helpful to identify a precancerous lesion. PMID- 10534339 TI - KDR/Flk-1 is a major regulator of vascular endothelial growth factor-induced tumor development and angiogenesis in murine hepatocellular carcinoma cells. AB - Vascular endothelial growth factor (VEGF), which is one of the most potent angiogenic factors, has been shown to play a pivotal role in tumor angiogenesis, including hepatocellular carcinoma (HCC). The effects of VEGF are mediated mainly through two distinct receptors, flt-1 and KDR/Flk-1. It has been suggested that KDR/Flk-1 plays an important role in tumor development. However, the role of KDR/Flk-1 in HCC has not been examined. We previously reported that VEGF tightly regulated murine HCC development, based on the results of a study using a retroviral tetracycline-regulated (Retro-Tet) gene expression system. This system allows VEGF gene expression to be manipulated in vivo by providing tetracycline in the drinking water. In the present study, we combined the KDR/Flk-1-specific neutralizing monoclonal antibody (KDR/Flk-1mAb) and the Retro-Tet system to elucidate the role of KDR/Flk-1 in VEGF-induced tumor development and angiogenesis in a murine HCC experimental model. In a xenograft study, tumor augmentation induced by VEGF overexpression was almost abolished by means of KDR/Flk-1mAb treatment, with accompanying inhibition of angiogenesis, KDR/Flk-1 autophosphorylation, but not interference of flt-1 activation. This inhibitory effect was achieved even on established tumors and regardless of whether the tumor size was small or large. On the contrary, KDR/Flk-1mAb treatment significantly increased the apoptosis in the tumor. With orthotopic transplantation, KDR/Flk-1mAb also inhibited HCC development in the liver. These results suggest that KDR/Flk-1 is a major regulator of VEGF-mediated HCC development and angiogenesis not only at the initial stage, but also after the tumor has fully developed. PMID- 10534340 TI - Liver-specific and proliferation-induced deoxyribonuclease I hypersensitive sites in the mouse insulin-like growth factor binding protein-1 gene. AB - The insulin-like growth factor binding protein-1 (IGFBP-1) gene is highly expressed in fetal, perinatal, and regenerating liver. Up-regulation is transcriptionally mediated in regenerating liver and occurs in the first few minutes to hours after partial hepatectomy. In transgenic mice a 970-bp region from -776 to +151 of the IGFBP-1 promoter was sufficient for tissue-specific and induced expression of the gene in fetal and hepatectomized livers. However weak and/or poorly regulated expression in some transgenic lines suggested the existence of other regulatory regions. Here, genomic clones containing large regions 5' of the mouse IGFBP-1 gene sequence were isolated, subcloned, and sequenced. Deoxyribonuclease I (DNaseI) hypersensitivity analyses identified clusters of tissue-specific nuclease-sensitive sites in the promoter region, -100 to -300, -2,300, -3,100, and -5,000 along with other weak sites. After partial hepatectomy, enhanced sensitivity and/or novel sites were detected in the -100/ 300, -5,000, and -3,100 regions, the promoter region remaining the most hypersensitive. A subset of these sites was present in fetal and perinatal livers. Novel tissue-specific sites that interacted with C/EBP and hepatic nuclear factor 3 (HNF3) transcription factors were identified in the -3,100 region. A hepatectomy-induced DNA binding complex containing the transcription factor USF1 was identified within the -100 to -300 region of the promoter. These results suggested that a complex array of tissue-specific and hepatic proliferation-induced transcription factors combine to regulate both the proximal promoter and more distal regulatory elements of the IGFBP-1 gene. PMID- 10534341 TI - Increased sensitivity to endotoxemia in the bile duct-ligated cirrhotic Rat. AB - Sepsis is a common complication of cirrhosis with a high mortality. In this study, we have investigated some of the pathways that may be involved in tissue injury and death. Bile duct-ligated (BDL) cirrhotic and control rats were challenged with lipopolysaccharide (LPS). Sensitivity to LPS was markedly enhanced in the BDL group, and was associated with increased liver injury and mortality. There was a 5-fold constitutive activation of nuclear factor kappa B (NFkappaB) in the liver of BDL rat controls (P <.001), and this was activated further, but to a similar extent, in the liver of both sham and BDL rats after injection of LPS. Plasma tumor necrosis factor alpha (TNF-alpha) increased more markedly in the BDL cirrhotic rats (2,463 +/- 697 pg/mL in BDL rats versus 401 +/ 160 pg/mL in the controls at 3 hours; P <.01). Plasma nitrite/nitrate concentrations were increased in the BDL controls at baseline, and increased further after LPS (P <.05), but did not differ from sham controls at 6 hours. Plasma F(2)-isoprostanes increased 6-fold in the cirrhotic rats and 2-fold in the controls (P <.01) indicative of lipid peroxidation. Esterified F(2)-isoprostanes in the liver increased 2- to 3-fold at 1 hour in control and BDL rats, but returned to baseline levels by 3 hours. Esterified F(2)-isoprostanes in the kidney increased by 2-fold in the BDL rats after LPS administration, but remained unchanged in sham controls. We conclude that there is a marked increase in sensitivity to LPS in BDL cirrhotic rats. This is associated with an enhanced TNF alpha response and increased lipid peroxidation. These may be directly and causally related to mortality. PMID- 10534342 TI - Suppression of lipopolysaccharide-induced nitric oxide synthase expression by platelet-activating factor receptor antagonists in the rat liver and cultured rat Kupffer cells. AB - Excessive nitric oxide (NO) generated by hepatic cells in response to lipopolysaccharide (LPS) and inflammatory substances (e.g., platelet-activating factor [PAF]) is a key contributor to the pathophysiological outcomes observed in the liver during sepsis. In rats subjected to liver-focused endotoxemia, inducible nitric oxide synthase (iNOS) levels in the intact liver were elevated by 6 hours; cell-specific expression of iNOS messenger RNA (mRNA) was Kupffer cells (KCs), endothelial cells, and hepatocytes. Elevated serum alanine transaminase (ALT) levels at 6 hours confirmed hepatic damage. Pretreatment of endotoxemic rats with PAF receptor antagonists BN 50739 or WEB 2170 reduced serum ALT and iNOS mRNA levels in the intact liver. Pretreatment of cultured KCs with BN 50739 or WEB 2170 inhibited both LPS and PAF-induced iNOS mRNA formation. In addition, LPS-induced iNOS protein levels in KCs pretreated with BN 50739 or WEB 2170 were decreased. Exposure of KCs to either LPS or PAF caused the translocation of the p65 subunit of nuclear factor kappa B (NF-kappaB) into the nucleus and this process was attenuated by BN 50739 and WEB 2170. There was concomitant inhibition of LPS-dependent degradation of the inhibitory protein IkappaBalpha and increase in intracellular Ca(2+) in KC treated with BN 50739 or WEB 2170. Also, in KCs, LPS was able to induce iNOS mRNA expression independent of CD14. This response was inhibited by pretreatment of KCs with either BN 50739 or WEB 2170. Our findings indicate that PAF receptor antagonists convey protection against hepatocellular injury accompanied by a decrease in nitric oxide (NO) formation in the livers of endotoxemic rats. PMID- 10534343 TI - Activation of caspase-8 in transforming growth factor-beta-induced apoptosis of human hepatoma cells. AB - Transforming growth factor-beta1 (TGF-beta1) has been shown to induce apoptosis in normal or transformed hepatocytes. To elucidate the biochemical pathways leading to apoptosis induced by TGF-beta1 in human hepatoma cells (HuH-7), we examined the expression of Bcl-2-related proteins and X-chromosome-linked inhibitor of apoptosis (XIAP), and activation of the caspase cascade following TGF-beta1 treatment. Bcl-xL expression began to decline at 12 hours after TGF beta1 treatment and progressively decreased to very low levels in a time dependent manner. Bax expression showed a little change throughout the experiment. On the other hand, activation of caspase-8 was clearly observed at 36 hours after TGF-beta1 treatment, followed by activation of caspase-9, and caspase 3 was activated at 48 hours after treatment at which time apoptosis of HuH-7 cells was observed. TGF-beta1 significantly decreased XIAP expression in HuH-7 cells. Addition of an inhibitor of caspase-8 or caspase-3 (IETD-FMK or DEVD-CHO) markedly inhibited TGF-beta1-induced apoptosis of HuH-7 cells. Fas/Fas ligand (FasL) interactions in HuH-7 cells were not involved in the apoptotic process. Furthermore, epidermal growth factor (EGF) also completely inhibited TGF-beta1 induced apoptosis of HuH-7 cells by inhibiting activation of the caspase cascade. Our results suggested that activation of caspase-3 initiated through caspase-8 activation is involved in the apoptotic process induced by TGF-beta1 in HuH-7 cells. Our results also showed that down-regulation of the expression of Bcl-xL and XIAP by TGF-beta1 may facilitate activation of caspase-3 in these cells. PMID- 10534344 TI - Ischemic preconditioning protects the mouse liver by inhibition of apoptosis through a caspase-dependent pathway. AB - A short period of ischemia and reperfusion, called ischemic preconditioning, protects various tissues against subsequent sustained ischemic insults. We previously showed that apoptosis of hepatocytes and sinusoidal endothelial cells is a critical mechanism of injury in the ischemic liver. Because caspases, calpains, and Bcl-2 have a pivotal role in the regulation of apoptosis, we hypothesized that ischemic preconditioning protects by inhibition of apoptosis through down-regulation of caspase and calpain activities and up-regulation of Bcl-2. A preconditioning period of 10 minutes of ischemia followed by 15 minutes of reperfusion maximally protected livers subjected to prolonged ischemia. After reperfusion, serum aspartate transaminase (AST) levels were reduced up to 3-fold in preconditioned animals. All animals subjected to 75 minutes of ischemia died, whereas all those who received ischemic preconditioning survived. Apoptosis of hepatocytes and sinusoidal endothelial cells, assessed by in situ TUNEL assay and DNA fragmentation by gel electrophoresis, was dramatically reduced with preconditioning. Caspase activity, measured by poly (adenosine diphosphate ribose) polymerase (PARP) proteolysis and a specific caspase-3 fluorometric assay, was inhibited by ischemic preconditioning. The antiapoptotic mechanism did not involve calpain-like activity or Bcl-2 expression because levels were similar in control and preconditioned livers. In conclusion, ischemic preconditioning confers dramatic protection against prolonged ischemia via inhibition of apoptosis through down-regulation of caspase 3 activity, independent of calpain like activity or Bcl-2 expression. PMID- 10534345 TI - Phospholipid metabolism of hypothermically stored rat hepatocytes. AB - Isolated rat hepatocytes were suspended and stored in either Liebovitz-15 medium (37 degrees C or 4 degrees C) or University of Wisconsin (UW) solution (4 degrees C) containing [(3)H] arachidonic acid (AA). At varying times, membrane phospholipids were separated by thin layer chromatography. AA labeled phospholipids similarly at both 4 degrees C and 37 degrees C. Analysis of the ratios of [(3)H] AA and [(14)C] glycerol incorporated into phosphatidic acid or other phospholipids in dual-labeled cells indicated that the deacylation/reacylation cycle was the major route of AA incorporation at hypothermia. This was supported by showing that blocking phospholipase A(2) (PLA(2)) activity by trifluoperazine suppressed AA incorporation into phospholipids. PLA(2) activity, measured by determining the release of AA, was slow during 48-hour cold storage, but increased significantly when ATP was depleted by inhibition of mitochondria and glycolysis. In the whole rat liver, there was no significant loss of phospholipids during 48-hour storage (total phospholipids [micromol phosphorus/L/mg] : 0.197 +/-. 001 at 0 hours) unless energy blockers were used (0.155 +/-.005 at 48 hours) or glycogen depleted by fasting the rat (0.167 +/-.001 at 48 hours). This study shows that a net PLA(2) stimulated hydrolysis of phospholipids is seen only when ATP is depleted and its generation from anaerobic glycolysis inhibited. Thus, PLA(2) hydrolysis of phospholipids is not a significant cause of liver cell injury during cold storage when livers are obtained in optimal condition. However, conditions affecting the generation of ATP during cold storage could alter PLA(2) leading to membrane damage. PMID- 10534346 TI - Concanavalin A hepatotoxicity in mice: tumor necrosis factor-mediated organ failure independent of caspase-3-like protease activation. AB - Several models of tumor necrosis factor (TNF)/TNF-receptor 1 (TNF-R1)-dependent liver injury in mice were investigated with respect to caspase-3-like protease activation representing a pivotal mechanism of apoptotic cell death. Injection of TNF or T-cell-activating agents (i.e., agonistic anti-CD3 antibody or staphylococcal enterotoxin B [SEB]) into galactosamine (GalN)-sensitized mice caused TNF/TNF-R1-dependent liver injury. Intravenous concanavalin A (Con A) alone induced TNF-mediated hepatotoxicity dependent on both TNF-R1 and TNF-R2. Hepatic caspase-3-like proteases were activated in GalN/TNF, GalN/anti-CD3, or GalN/SEB-treated mice, but not in Con A-treated mice. Consistently, the broad spectrum caspase inhibitor, benzoyloxycarbonyl-val-ala-asp-fluoromethylketone (zVADfmk), prevented TNF-mediated hepatotoxicity in all GalN-dependent models, but failed to protect against Con A. Under transcriptional arrest, however, Con A induced TNF-R1-dependent, but not TNF-R2-dependent, activation of caspase-3-like proteases, and zVADfmk prevented animals from Con A-mediated liver injury under this condition. Histological analysis revealed distinct differences between Con A and GalN/Con A-induced liver injury regarding apoptotic morphology of hepatocytes. We conclude that impaired transcription induces a switch of Con A hepatotoxicity toward a caspase-3-like protease-dependent pathway. The observation that the functional state of the transcriptional machinery decides whether TNF-driven hepatocyte apoptosis involves activation of caspase-3-like proteases or alternative signaling pathways in vivo might be of relevance for the immunopathology of the liver. PMID- 10534347 TI - Endotoxin suppresses mouse hepatic low-density lipoprotein-receptor expression via a pathway independent of the toll-like receptor 4. AB - Endotoxin provokes an inflammatory state in the infected host. C3H/HeJ mice are tolerant to endotoxin because of an Lps gene mutation. Recent studies have identified that this gene encodes the Toll-like receptor 4. Endotoxin also induces hyperlipidemia and suppresses hepatic low-density lipoprotein (LDL) receptor expression. In the current study, we investigated whether a defective Lps gene would impair the hepatic LDL-receptor response to endotoxin in C3H/HeJ mice. Eighteen hours after an intraperitoneal injection of endotoxin, the hepatic LDL-receptor expression and the plasma lipoprotein pattern were analyzed. Endotoxin increased plasma triglyceride and apoE in very low-density lipoproteins (VLDL) and intermediate-density lipoproteins, and decreased apoAI in high-density lipoproteins (HDL) in the endotoxin-sensitive mice (C3H/HeN), but not in the endotoxin-resistant mice (C3H/HeJ). These data indicate that a defective Lps gene impairs the endotoxin signaling to alter these lipoproteins. However, the hepatic LDL-receptor response to endotoxin in the endotoxin-resistant mice was similar to that in the endotoxin-sensitive mice. Thus, at a dose of 5 microg/mouse, endotoxin reduced hepatic LDL-receptor expression by 35% in C3H/HeN mice and by 52% in C3H/HeJ mice. At a dose of 50 microg/mouse, endotoxin reduced hepatic LDL receptor expression by 61% in C3H/HeN mice and by 63% in C3H/HeJ mice. It is concluded that endotoxin suppresses hepatic LDL-receptor expression in vivo via a pathway independent of the Toll-like receptor 4. PMID- 10534348 TI - Hepatobiliary transport of bile acid amino acid, bile acid peptide, and bile acid oligonucleotide conjugates in rats. AB - Uptake of drugs by bile acid carriers could account for the selectivity of drug actions in the gut and liver. We have previously shown that conjugation of xenobiotics with bile acids facilitates their transfer to hepatocytes and ileal enterocytes. In this study L-alanine and 2 biooligomers, the tetrapeptide L (ala)(4) and a 15 mer oligodeoxynucleotide (ODN) were coupled covalently via linker molecules to the 3-position of bile acids. The L-alanine-coupled bile acid conjugates were rapidly taken up by the liver and efficiently eliminated into bile. These compounds mimicked hepatic transport of bile acids. Also in case of the tetrapeptide (ala)(4), bile acid conjugation significantly improved hepatic and intestinal cell uptake and rendered the peptide conjugate resistant to peptidases. Because uptake by isolated hepatocytes was not dependent on sodium ions and was blocked by ochratoxin A, we assume basolateral transport by an oatp type bile acid carrier. In the case of the 15 mer ODN, normal and bile acid conjugated oligodeoxynucleotide appeared intact in bile but without marked improvement of hepatocellular uptake and biliary elimination. We conclude that bile acids can deliver small peptides to gut and parenchymal liver cells via bile acid transport pathways, whereas in the case of oligonucleotides an attached bile acid was not sufficient to shuttle them successfully into hepatocytes. PMID- 10534349 TI - Hepatic jagged1 expression studies. AB - Mutations in Jagged1, a Notch ligand, have been shown to result in Alagille syndrome (AGS), however, the causal link between haploinsufficiency of Jagged1 and intrahepatic ductal paucity is unknown. This survey was performed to determine the expression pattern of Jagged1 in the fetal and postnatal liver. Reverse transcription polymerase chain reaction (RT-PCR) showed Jagged1 expression in all samples studied including rat liver embryonic days 16 to 21, 1 day-old, 1-week-old, and 2-month-old adult rats. RT-PCR detected Jagged1 in total liver RNA extracted from cadaver organ donor samples from reduced human grafts and explanted native livers from a variety of pediatric disorders including AGS, biliary atresia, congenital hepatic fibrosis, sclerosing cholangitis, cystic fibrosis, fulminant hepatic failure, tyrosinemia, and chronic rejection. Immunohistochemistry showed Jagged1 expression in human fetal samples localized to the ductal plate from 14-week gestation onward. Expression in the postnatal liver was seen in biliary epithelium and zone 3 hepatocytes. In conclusion, these studies show that Jagged1 is expressed in the fetal and postnatal liver in health and disease. We show localization of expression by immunohistochemistry to ductal plate epithelium in human fetal samples and to the biliary epithelium and zone 3 hepatocytes in human postnatal samples. Our results show the localization of Jagged1 in fetal liver and demonstration of Jagged1 expression in postnatal rat and human liver specimens. Further studies of Jagged1 and the Notch signaling pathway are expected to elucidate mechanisms of the regulation of biliary epithelial growth and development. PMID- 10534350 TI - Characterization of a hyaluronan receptor on rat sinusoidal liver endothelial cells and its functional relationship to scavenger receptors. AB - Hyaluronan is a widely distributed extracellular component of connective tissue with several mechanical and cell biological functions. The serum level of hyaluronan is elevated in rheumatic and liver diseases and in certain malignancies. The major route of hyaluronan clearance from the blood is via the liver, taken up predominantly by sinusoidal liver endothelial cells. We have purified a novel hyaluronan binding protein from liver that also has an affinity for the N-terminal propeptide of type I procollagen, a physiological scavenger receptor ligand. A polyclonal antibody raised against the protein was found to inhibit the binding and degradation of hyaluronan as well as two scavenger receptor ligands by cultured sinusoidal liver endothelial cells. Immunostaining of nonpermeabilized liver cells and liver sections showed that the antibody specifically stains the surface of sinusoidal liver endothelial cells. After pretreatment with monensin to block the recirculation of endocytic receptors, the immunostaining was specifically associated with early endosomes of these cells. Thus, this rat sinusoidal liver endothelial cell hyaluronan receptor shares functional properties with the scavenger receptor family, a group of proteins shown to play a key role in the uptake of atherogenic lipids and other waste products from the tissues. PMID- 10534351 TI - Characterization of the reactivity pattern of murine monoclonal antibodies against wild-type hepatitis B surface antigen to G145R and other naturally occurring "a" loop escape mutations. AB - The hepatitis B surface antigen (HBsAg) "a" domain harbors major B-cell epitopes. Viruses with mutations in this region emerge after vaccination or during hepatitis B immune globulin (HBIg) prophylaxis. A strain with G145R replacement has been almost invariably isolated as a major escape mutant. We investigated mutant antigen-antibody interactions with direct binding assays. G145R and 16 other naturally occurring recombinant HBsAg mutants were expressed in mammalian Cos-1 cells. The reactivity of a panel of 28 murine anti-hepatitis B surface antigen (anti-HBs) monoclonal antibodies to mutant antigens was measured with enzyme immunoassay and expressed as percentage compared with the wild-type (wt) HBsAg signal for each antibody. All point-mutated proteins displayed diffuse intracellular immunofluorescent labeling corresponding to a secretory pathway. Monoclonal antibodies (mAbs) were classified according to different binding patterns. The effect of mutations on antibody binding differs depending on the amino acid involved and on the location within the "a" loop. As expected, most antibodies had absent or negligible binding (<40%), notably with residue 145 replacements. However, we identified antibodies that reacted with conformational epitopes but nevertheless had adequate reactivity (>40%) with all mutant antigens, including G145R. The effect of G145R was more pronounced than that of G145A. A subgroup of antibodies had substantially increased recognition (>120%) of antigens with mutations in the first loop. We demonstrated that antibodies can be selected or combined that react with all mutants investigated, including G145R. These data offer perspectives for improving anti-HBs-based protection against hepatitis B. PMID- 10534352 TI - Outcome of de novo hepatitis C virus infection in heart transplant recipients. AB - The outcome of de novo hepatitis C virus (HCV) infection in heart transplant recipients of HCV-antibody positive organs is not known. The aim of the study was to determine the short-term outcome of de novo HCV infection in recipients of HCV positive donor organs. HCV-antibody negative recipients of HCV-antibody positive hearts were identified from January 1, 1991 to February 28, 1998. Control patients matched for year of transplantation were also identified. Twenty-eight patients (22 males, mean age of 56 +/- 11 SD) received hearts from HCV-antibody positive donors. The control group was similar to the patients in all clinical and demographic aspects. Twenty-three patients had detectable viremia by reverse transcription polymerase chain reaction (RT-PCR). Of these 23 patients with de novo HCV infection, 7 (30%) developed HCV-related liver disease. Three patients (13%) had chronic hepatitis and 4 patients (17%) developed severe acute cholestatic hepatitis (ACH). Mycophenolate mofetil (MMF) use (P =.04) and high viral load at onset of acute liver disease (P =.02) were associated with ACH. Overall survival was similar between patients with de novo HCV infection and controls (P =.20). Development of ACH (P =.02) and MMF use (P =.0009) were associated with decreased survival in patients with de novo HCV infection. The present study showed that survival of patients with de novo HCV infection was similar to a matched control group. HCV-related severe ACH is associated with a poor short-term outcome in patients with de novo HCV infection. MMF use may be associated with a higher incidence of HCV-related severe ACH and a poor short term outcome. PMID- 10534353 TI - The impact of diagnosis of hepatitis C virus on quality of life. AB - The aim of this study was to examine the effects of diagnosis of hepatitis C virus (HCV) infection on quality of life in a cohort admitted to Fairfield Infectious Diseases Hospital with acute hepatitis from 1971 to 1975. Sera stored from the original admission were tested for antibody to HCV. Systematic approaches were used to locate anti-HCV-positive individuals and outcomes assessed by the Short Form 36 (SF-36) scale and a study-specific questionnaire as well as clinical review. Study subjects' SF-36 scores were compared with Australian population norms. Anti-HCV and HCV-RNA positive individuals (n = 15) aware of their serostatus rated significantly worse on 7 of 8 SF-36 scales compared with population norms. However, HCV-seropositive and RNA-positive individuals unaware of their HCV serostatus (n = 19) scored significantly worse in only 3 scales. Those aware of their serostatus did not differ sociodemographically, clinically, virologically, or serologically from those who were unaware, nor was there a link between quality of life (QOL) scores and objective measures of ill health. All subjects had injected drugs in the past. In conclusion, HCV-RNA and anti-HCV-positive individuals in our study have significantly poorer subjective health status 26 years after original infection compared with population norms. QOL measures were significantly worse for HCV seropositive individuals aware of their serostatus compared with those unaware. We feel that the reduced QOL in the diagnosed group may be partially an effect of labeling and that the impact of the diagnostic process per se on QOL in individuals with HCV requires further evaluation. PMID- 10534354 TI - Long-term incidence of hepatitis B virus resistance to lamivudine in human immunodeficiency virus-infected patients. AB - Hepatitis B virus (HBV) resistance to lamivudine has not been extensively documented in human immunodeficiency virus (HIV)-infected patients. We studied the long-term incidence of HBV resistance to lamivudine in HIV-positive patients. Sixty-six HIV-HBV-coinfected patients were studied while receiving lamivudine (150 mg twice daily) as a part of antiretroviral therapy. All these patients had a detectable serum HBV DNA at the beginning of lamivudine therapy. Serum HBV DNA was quantified by molecular hybridization. Sequence analysis of the HBV polymerase was performed in patients who became resistant to lamivudine. After 2 months of lamivudine, HBV DNA became undetectable in 57 patients (86.4%, 95% CI: 75.7%-93.6%). After 2 years of lamivudine, 47% +/- 18.6% of the patients, had sustained HBV-DNA suppression. All the 22 tested patients with HBV resistance developed mutation at position 550 in the YMDD motif of the DNA polymerase. None of the following variables were associated with an increased risk of lamivudine resistance: age, associated protease inhibitor therapy, Center for Disease Control (CDC) stage C, known HIV-infection duration, serum HBV-DNA level at baseline, CD4 cell count and serum alanine transaminase levels at baseline and at HBV-replication suppression (2 months of lamivudine). Lamivudine (300 mg/d) is effective for the inhibition of HBV replication in HIV-infected patients. However, emergence of lamivudine-resistant HBV may occur in 20% of patients per year. PMID- 10534355 TI - Clinical features of hepatitis C-infected patients with persistently normal alanine transaminase levels in the Southwestern United States. AB - Approximately one third of patients with chronic hepatitis C virus (HCV) infection have normal alanine transaminase (ALT) levels. We studied the clinical, biochemical, virological, and histological features in patients with persistently normal ALT. A case-control study was conducted on 275 patients with chronic HCV infection, including 75 patients with persistently normal ALT and 200 patients with abnormal ALT. Persistently normal ALT was defined as 4 consecutive ALT values in each patient within a period of 12 months. The average age of the patients was 44 years (range 18 to 69 years). More non-Hispanic whites had persistently normal ALT. The mean serum ferritin level was significantly lower in patients with persistently normal ALT as compared with abnormal ALT (128 +/- 92 ng/mL and 224 +/- 128 ng/mL), respectively (P =.017). The mean HCV-RNA level was significantly lower in patients with persistently normal ALT as compared with abnormal ALT (12 x 10(5) +/- 2.8 x 10(6) copies/mL and 33 x 10(5) +/- 8.0 x 10(6)), respectively (P =.02). Histologically, patients with persistently normal ALT had less severe portal inflammation (P <.05), lobular inflammation (P =.003), piecemeal necrosis (P =.002), fibrosis (P <.05), lower prevalence of cirrhosis (P =.007), as well as a slower fibrosis progression rate (P <.001). Chronic hepatitis C patients with persistently normal ALT have low-activity grade and stage on liver biopsy. In these patients the hepatitis C RNA level was lower compared with abnormal ALT patients, which may explain the slower fibrosis progression rate. PMID- 10534356 TI - Changes of hepatitis B surface antigen variants in carrier children before and after universal vaccination in Taiwan. AB - Mutants of a determinant of hepatitis B surface antigen (HBsAg) identified in vaccinated children pose a potential threat to long-term success of vaccination programs. We examined the mutants of a determinant (residues 110-160) of HBsAg in hepatitis B virus (HBV) DNA-positive children identified during previous serosurveys in Taipei undertaken just before (1984), 5 years after (1989), and 10 years after (1994) universal vaccination began. In HBV DNA-positive children from 3 surveys, the prevalence of a determinant mutants increased from 8 of 103 (7.8%) in 1984 to 10 of 51(19.6%) in 1989 and 9 of 32 (28.1%) in 1994 and was higher in those fully-vaccinated than unvaccinated (12/33 vs. 15/153, P =. 0003). Most amino acid changes of the variants clustered in residues 125-129 and 140-149. In all 27 children with detectable mutants, the mean age of those vaccinated was younger than those unvaccinated (4. 8 +/- 3.8 vs. 7.9 +/- 2.3 yrs, P <.05); and mutations occurred in a region with greatest local hydrophilicity (residues 140 149) more frequently in those vaccinated than in those unvaccinated (10/12 vs. 6/15, P =.0253). More mutated residues and more mutations at neutralizing epitopes, such as N146, C147, T148, and C149, were found in the 1994 survey. Vaccinated children may contract variant infections through vertical or horizontal transmission. Universal vaccination has accelerated an accumulation of HBsAg a determinant mutants with amino acid changes critical for immune escape in vaccinated children who became carriers, suggesting that new vaccination strategies should be considered. PMID- 10534357 TI - Cost effectiveness of interferon alpha2b combined with ribavirin for the treatment of chronic hepatitis C. AB - Treatment of chronic hepatitis C with Interferon (IFN) alpha2b monotherapy results in 10% to 15% sustained virological response (SVR). Combining IFN with ribavirin increases this response. In this analysis, using the Markov model, 6 treatment strategies for chronic hepatitis C (previously untreated) were compared on the basis of incremental cost per additional quality-adjusted life years ($/QALY). Our results showed that the no treatment strategy was associated with a cost of $38,747 and 13.10 QALYs. The strategy using IFN alone for 48 weeks was associated with a cost of $35,642 and 14.05 QALYs. The strategy using IFN monotherapy followed by combination therapy for nonresponders and relapsers was associated with a cost of $34, 561 and 15.53 QALYs. A similar strategy, but limiting combination to relapsers only, was associated with a cost of $34,758 and 14.40 QALYs. The strategy using IFN with ribavirin as the initial therapy for all patients was associated with a cost of $34,792 and 15.31 QALYs. Finally, the strategy using viral genotyping first and then adjusting the duration of combination therapy based on genotype was associated with a cost of $37,263 and 15.89 QALYs. The strategy using genotyping to guide duration of combination therapy was the most cost-effective approach with an incremental cost effectiveness ratio of $7,500 per QALY. Sensitivity analyses confirmed the robustness of these results. We conclude that combination of IFN and ribavirin with duration of therapy based on the viral genotype, is a cost-effective approach in treating patients with chronic hepatitis C. PMID- 10534358 TI - Serum levels of soluble interferon Alfa/Beta receptor as an inhibitory factor of interferon in the patients with chronic hepatitis C. AB - Human serum contains a soluble form of interferon alfa/beta (sIFN alpha/beta) receptors, the functional and clinical significance of which has not been investigated in patients with chronic hepatitis C. In the present study, serum levels of sIFN alpha/beta receptor were assessed in 81 patients with chronic hepatitis C and correlated with the effectiveness of IFN therapy in these patients. Serum levels of sIFN alpha/beta receptor were significantly higher in patients with chronic hepatitis C than in healthy control patients (P <.0001). In these patients, serum levels of sIFN alpha/beta receptor were correlated with those of alanine transaminase (ALT) (P <.05), (2'-5')serum oligo(A) synthetase (2 5AS) (P <.0001), and pathological stages of liver fibrosis (P <.01). In 55 patients with chronic hepatitis C who underwent IFN therapy, there was an inverse correlation between the pretherapeutic serum levels of sIFN alpha/beta receptor and the rate of increase in serum levels of 2-5AS after the start of IFN (P <.01). Pretherapeutic serum levels of sIFN alpha/beta receptor were significantly lower in patients who showed sustained response to IFN therapy compared with those who did not respond to the therapy (P <.05). Multivariate analysis showed that low levels of serum sIFN alpha/beta receptor (C&dbond;O moiety for both 3-buten-2-one and 2-butanone. At simulated positive potentials, 3 buten-2-one adsorption occurred on Pt(111) and Pt(100) clusters through the pi orbitals of the >C&dbond;C< moiety, whereas at negative potentials 3-buten-2-one was adsorbed via u-bridging or di-varsigma configuration(s). Copyright 1999 Academic Press. PMID- 10534386 TI - On the Use of Washburn's Equation in the Analysis of Weight-Time Measurements Obtained from Imbibition Experiments. AB - A careful analysis of the experimental results from imbibition experiments, expressed as the increased weight of a porous layer once it was in contact with a liquid, vs time, has been done on the basis of Washburn's equation, which is usually the main tool for that analysis. It has been found that the experimental results reflect other physical phenomena more than imbibition, among them the effect of the initial contact between plate and liquid and the evaporation of liquid from the wetted porous layer. The unavoidable disturbance introduced by the first of these phenomena leads to the need for rescaling of the experimental data. Once this is done, Washburn's equation has to be applied accordingly to the new reference system chosen. Also, it has been shown that if evaporation is present, experimental data deviate from Washburn's predictions. This effect disappears if the porous layer is covered. Copyright 1999 Academic Press. PMID- 10534387 TI - The Dynamic Adsorption of Charged Amphiphiles: The Evolution of the Surface Concentration, Surface Potential, and Surface Tension. AB - In this work the evolution of the surface concentration, surface potential, and surface tension for adsorption of a charged amphiphile at an interface is studied numerically. While the results are of interest for any amphiphile, the simulations are performed for typical surfactant material parameters. The surface potential is related at each time step to the instantaneous surface charge density determined by the surfactant surface concentration using the Gouy-Chapman model. The sublayer concentration at each time step is a Boltzmann distribution in instantaneous equilibrium with the surface potential. At equilibrium, the surfactant is assumed to obey the Davies adsorption isotherm. The model is integrated first for diffusion-controlled adsorption, in which the surfactant diffuses to the sublayer and adsorbs onto the interface in local equilibrium according to the adsorption isotherm. In this limit, since the equilibrium adsorption is strongly reduced by the repulsive electrostatic potential, the time required to deliver the surfactant by diffusion is also reduced. The greater the electrical repulsion, the faster the diffusion-controlled adsorption at a given surfactant concentration. Because less surfactant adsorbs, the surface tension reduces less at equilibrium. Counterions of greater valence than the surfactant are more effective at screening the surface potential. Equilibrium adsorption, surface tension reduction, and diffusion time scales increase. As the surfactant valence increases, so does the repulsion; the opposite trends in surface tension and diffusion time scales are predicted. The model is also integrated including both diffusion and adsorption-desorption kinetic barriers. In experiment, adsorption-desorption kinetic barriers have been shown to control the mass transfer of non-ionic surfactants at elevated bulk concentration. The ability of the interface to deplete the bulk reduces with concentration. Therefore, diffusion time scales are reduced. In these regimes, adsorption-desorption kinetics can be rate determining. In simulation, the occurrence of the shift of the controlling mechanism from pure diffusion control at dilute concentration to mixed kinetic-diffusion control at elevated concentration is strongly influenced by ionic strength and surfactant valence. As the electrostatic adsorption increases, kinetic barriers are apparent at lower concentrations. Finally, a simple time scale argument that has previously proven useful in predicting a priori the time required for diffusion-controlled adsorption to an interface for nonionic surfactant adsorption is extended to include electrostatic effects. Copyright 1999 Academic Press. PMID- 10534388 TI - Experimental Determination of Drop Shape in Slow Steady Shear Flow. AB - Theoretical predictions for the 3D shape of a Newtonian drop immersed in a Newtonian fluid under slow flow have been available for a long time (C.E. Chaffey and H. Brenner, J. Colloid Interface Sci. 24, 258 (1967); J.M. Rallison, J. Fluid Mech. 98, 625 (1980)). The predictions contain four scalar coefficients, given in terms of the viscosity ratio of the component fluids. In this work, experimental values of such coefficients were measured for the case of two Newtonian fluids in simple shear flow. The experiments were carried out in a parallel plate apparatus, equipped with video-enhanced microscopy, and the measurements were performed by image analysis techniques. The results were also compared to theoretical calculations and provide the first complete assessment of shape predictions for the case of Newtonian fluids. Copyright 1999 Academic Press. PMID- 10534389 TI - Transport Properties of Cation Exchange Membranes in the Presence of Ether Compounds in Electrodialysis. AB - Ether compounds such as ethylene glycols with different molecular weights and crown ethers have good affinity to the cation exchange membranes, sulfonated styrene-divinylbenzene membrane (sodium ion form) and perfluorocarbon sodium sulfonate membrane. The impregnated amount of ethylene glycols in the membranes was higher than the water content of the membranes. After the ether compounds had been impregnated in the cation exchange membranes, electrodialysis of mixed salt solutions (1:1 mixture of alkaline earth metal ions or potassium ions and sodium ions) was carried out in the presence of the compounds to observe the change in permselectivity between two cations. Though current efficiency did not change in the presence of the compounds, transport numbers of alkaline earth metal ions relative to sodium ions decreased. Namely, sodium ions permeated through the membrane more selectively than alkaline earth metal ions. This is due mainly to a decrease in the mobility of alkaline earth metal ions in the membrane phase and partially to a decrease in the ion exchange equilibrium constant of alkaline earth metal ions to sodium ions with the membrane. This originates from the difference in ion-dipole interaction between cations and ether groups. The transport number of potassium ions relative to sodium ions also decreased in the presence of the compounds. In particular, the permeation of potassium ions relative to sodium ions remarkably decreased in the presence of 18-crown-6 in the membrane and in the solution due to the formation of a strong complex between potassium ions and 18-crown-6. Copyright 1999 Academic Press. PMID- 10534390 TI - Equilibrium Shapes of Solid Particles on Elastically Mismatched Substrates. AB - Small particles or islands bonded to a substrate can be profoundly influenced by both interfacial and elastic driving forces that tend to have opposing influences on the apparent wetting behavior. The superposition of these two driving forces can therefore lead to a rich set of particle properties, most notably their equilibrium shapes. Here we present a variational analysis leading directly to an Euler-Lagrange equation that can be solved to yield the equilibrium shapes of partially wetting particles as a function of their size, interface energy densities, and elastic interaction with a rigid substrate. The solutions are used to gain insight into the variables that most significantly influence the equilibrium morphology, and to derive the approximate driving force for surface area reduction by coarsening among a dispersion of unequally sized particles. The relatively simple analytical model can also form a foundation upon which more realistic numerical simulations may be built and compared. Copyright 1999 Academic Press. PMID- 10534391 TI - Initial Heats of H(2)S Adsorption on Activated Carbons: Effect of Surface Features. AB - The sorption of hydrogen sulfide was studied on activated carbons of various origins by means of inverse gas chromatography at infinite dilution. The conditions of the experiment were dry and anaerobic. Prior to the experiments the surface of some carbon samples was oxidized using either nitric acid or ammonium persulfate. Then the structural parameters of carbons were evaluated from the sorption of nitrogen. From the IGC experiments at various temperatures, heats of adsorption were calculated. The results showed that the heat of H(2)S adsorption under dry anaerobic conditions does not depend on surface chemistry. The dependence of the heat of adsorption on the characteristic energy of nitrogen adsorption calculated from the Dubinin-Raduskevich equation was found. This correlation can be used to predict the heat of H(2)S adsorption based on the results obtained from nitrogen adsorption. Copyright 1999 Academic Press. PMID- 10534392 TI - A Kinetic Model of Protein Adsorption/Surface-Induced Transition Kinetics Evaluated by the Scaled Particle Theory. AB - The adsorption of proteins and other large molecules at the liquid-solid interface often involves a surface-induced transition in either internal conformation or molecular orientation. Recently, Van Tassel et al. modeled this adsorption/transition process as the sequential surface placement of spreading disks. In this work, we employ the scaled particle theory (SPT) to derive approximate analytical expressions for the probability functions appearing in the kinetic equations for this model system. Specifically, the probability functions governing the adsorption and spreading events are calculated in terms of the reversible work required to create cavities in a binary system of spread and unspread disks. Compared to those derived earlier via a density expansion theory (DET), the SPT approximated probability functions are simpler and more accurate (compared to simulation), and are applicable over a wider set of parameter values. Copyright 1999 Academic Press. PMID- 10534393 TI - Chemical Charge Regulation and Charge Renormalization in Concentrated Colloidal Suspensions. AB - This article discusses the charging behavior of particles in a colloidal suspension allowing for a simple model reaction at the surface of the colloidal particles that regulates the total number Z(f) of free counterions. This number is to be distinguished from an effective number of charges Z* calculated in the Wigner-Seitz cell model to determine the interaction between the colloidal particles. We discuss the problem in terms of the full free energy of the system. The number of free counterions Z(f) as a function of the structural charge of the colloid is derived from the minimum of this free energy. Our results are compared with experimental data. Copyright 1999 Academic Press. PMID- 10534394 TI - Surface Complexation of Hg(2+) on Goethite: Mechanism from EXAFS Spectroscopy and Density Functional Calculations. AB - EXAFS spectra of Hg(2+) sorbed on goethite at pH 4.60 has revealed the existence of an inner-sphere complex with an immediate coordination shell of 2 O atoms at 2.04 A and a further shell of 1 Fe atom at 3.28 A. Using density functional calculations, we determined the optimized geometries of small clusters representing surface complexes. These results shows that adsorption of Hg(2+) occurs via two oxygen atoms bound to edge-sharing Fe sites on the (110) surface. This is the same surface site we find for Cd (1, 2) and Sr (3). The short Hg-O bond length shows that the oxygens bridging Hg and Fe atoms are deprotonated. Copyright 1999 Academic Press. PMID- 10534395 TI - Reactions of a Trifunctional Silane Coupling Agent in the Presence of Colloidal Silica Sols in Polar Media. AB - Reactions of 3-glycidoxypropyltrimethoxysilane (GPS) in a water-rich environment with silica colloids were studied. (29)Si NMR spectroscopy was used to monitor quantitatively the variation in GPS hydrolysis, condensation, and adsorption with pH and water concentration. The results show the competition between adsorption on the silica surface and condensation polymerization of GPS in the bulk liquid and are used to identify conditions favorable to surface modification. The presence of colloidal silica was shown to increase the apparent GPS dimerization rate, suggesting either surface catalysis or an increased local reactant concentration in the electrical double layer around the silica colloids. Copyright 1999 Academic Press. PMID- 10534396 TI - The Effect of Electromagnetic Retardation on the Rupture Process of a Very Thin Liquid Film. AB - Effects of electromagnetic retardation on the rupture process of a very thin liquid film coated on a flat plate are studied. The analysis results indicate that the electromagnetic retardation effect (D) for the case A > 0 (attraction) is a stabilization factor, which prolongs the rupture time. The wavenumber of the most unstable mode is decreasing as D increases. It is also found that the linear solution of rupture time T(LM) is larger than the nonlinear rupture time T(NM), but the gradient of T(LM) to D is comparable to that of T(NM). Copyright 1999 Academic Press. PMID- 10534397 TI - Insertional mutation of the collagen genes Col4a3 and Col4a4 in a mouse model of Alport syndrome. AB - Mice homozygous for the transgenic insertion in line OVE250 exhibit severe progressive glomerulonephritis. Ultrastructural changes in the glomerular basement membrane (GBM) at 2 weeks of age resemble those in Alport syndrome. The transgenic insertion site was mapped by FISH to mouse chromosome 1 close to Pax3. Genetic and molecular analyses identified a deletion of genomic DNA at the transgene insertion site. Exons 1 through 12 of the collagen IV gene Col4a4, exons 1 and 2 of the adjacent Col4a3 gene, and the intergenic promoter region are deleted. Transcripts of Col4a3 and Col4a4 are undetectable in mutant kidney, and both proteins are missing from the GBM. Persistent cellular proliferation in mutant kidneys suggests that interaction with the extracellular matrix may be important for cell maturation. Evolutionarily conserved sequence elements in the promoter regions of human and mouse Col4a3 and Col4a4 include a 19-bp element that was tandemly duplicated in the human lineage and a CTC box element common to several genes encoding extracellular matrix proteins. This new animal model of Alport syndrome, Col4Delta3-4, lacks both alpha3 and alpha4 chains of collagen IV and exhibits an earlier disease onset than mice lacking alpha3 only. PMID- 10534398 TI - Human and mouse XAP-5 and XAP-5-like (X5L) genes: identification of an ancient functional retroposon differentially expressed in testis. AB - Although most retroposons that arose by reverse transcription of cellular mRNAs and by reintegration into the genome are nonfunctional, several examples exist in which the retroposon acquired a novel function and became fixed in the genome as a functional gene. We identified another such case: the ubiquitously expressed X linked XAP-5 gene with unknown function gave rise to its retroposed counterpart, XAP-5-like (X5L), which has an intronless open reading frame and is autosomal in human. Phylogenetic analysis of the human and mouse XAP-5 and X5L genes shows that the retroposition most likely took place before the radiation of eutherian mammals. The XAP-5 and X5L genes are expressed in a wide range of tissues but are differentially expressed in testis. The ancient origin and broad expression of the X5L retroposon indicate that the XAP-5 and X5L genes may have assumed different functions in somatic cells. In addition to this, because of its autosomal location and its high level and particular pattern of expression in spermatogenic cells, the X5L expression in testis may compensate for the X-linked XAP-5 gene, which may be silenced during spermatogenesis. PMID- 10534400 TI - Reciprocal chromosome painting reveals detailed regions of conserved synteny between the karyotypes of the domestic dog (Canis familiaris) and human. AB - The domestic dog is increasingly being recognized as a useful model for human disease. The aim of this study was to conduct the first detailed whole-genome comparison of human and dog using bidirectional heterologous chromosome painting (reciprocal Zoo-FISH) analysis. We used whole-chromosome paint probes produced from degenerate oligonucleotide-primed PCR amplification of high-resolution bivariate flow-sorted human and dog chromosomes. No fewer than 68 evolutionarily conserved segments were identified between the dog and the human karyotypes. The use of elongated metaphase chromosomes for both species allowed the boundaries of each evolutionarily conserved segment to be determined to subband resolution. The distribution of conserved segments is discussed, as are the applications of these data in refining the current status of the dog genome map. PMID- 10534399 TI - Molecular characterization of a MSRV-like sequence identified by RDA from monozygotic twin pairs discordant for schizophrenia. AB - Retroviral-related amplicons were used in modified RDA to identify four sequences from affected members of three pairs of monozygotic twins discordant for schizophrenia. One sequence (schizophrenia associated retrovirus, SZRV-1, GenBank Accession No. AF135487) is characterized here. It is similar to two known sequences of retroviral origin: multiple sclerosis-associated retrovirus, MSRV (GenBank Accession No. AF009668), and ERV-9 (GenBank Accession No. S77575). It is present in multiple copies in the human genome and has been localized to six different chromosomal sites. A zooblot shows that this multicopy sequence is predominant in the primate lineage and present in rhesus monkeys and humans. SZRV 1 is expressed as a 9-kb RNA band in the placenta. This could offer support to the hypothesis that retroviral sequences transposing during fetal growth may alter neurodevelopmental genes and cause diseases, although its direct involvement in the causation of schizophrenia remains to be established. PMID- 10534401 TI - Cloning, characterization, and localization of mouse and human SPO11. AB - Spo11 is a meiosis-specific protein in yeast that has been found covalently bound to DNA double-strand breaks (DSBs) during the early stages of meiosis. These DSBs initiate homologous recombination, which is required for proper segregation of chromosomes and the generation of genetic diversity during meiosis. Here we report the cloning, characterization, tissue expression, and chromosomal localization of both mouse and human homologues of Spo11. The putative mouse and human proteins are 82% identical and share approximately 25% identity with other family members. Northern blot analysis revealed testis-specific expression for both genes, but RT-PCR results showed ubiquitous expression of at least a portion of Spo11 in mouse. Human SPO11 was also detected in several somatic tissues. Mouse Spo11 was localized to chromosome 2H4, and human SPO11 was localized to chromosome 20q13.2-q13.3, a region amplified in some breast and ovarian tumors. PMID- 10534402 TI - A mouse homolog of the Saccharomyces cerevisiae meiotic recombination DNA transesterase Spo11p. AB - The Saccharomyces cerevisiae Spo11 protein is thought to catalyze formation of the DNA double-strand breaks that initiate meiotic recombination. We have cloned cDNA and genomic DNA for a mouse gene encoding a protein with significant sequence similarity to conserved domains found in proteins of the Spo11p family. This putative mouse Spo11 gene maps to the distal region of chromosome 2 (homologous to human chromosome 20q13.2-q13.3) and comprises at least 12 exons, spanning approximately 15-18 kb. Strong expression of the Spo11 message is seen in juvenile and adult testis by RNA in situ hybridization, RT-PCR, and Northern blot, with much weaker expression in thymus and brain. In situ hybridization detects expression in oocytes of embryonic ovary, but not of adult ovary. RT-PCR and in situ hybridization analyses of a time course of juvenile testis development indicate that Spo11 expression begins in early meiotic Prophase I, prior to the pachytene stage, with increasing accumulation of mRNA through the pachytene stage. Taken together, these results strongly suggest that this gene encodes the functional homolog of yeast Spo11p, which in turn suggests that the mechanism of meiotic recombination initiation is conserved between yeast and mammals. PMID- 10534403 TI - Structural analysis and mapping of DNase I hypersensitivity of HS5 of the beta globin locus control region. AB - The beta-globin locus control region (LCR) is a cis regulatory element that is located in the 5' part of the locus and confers high-level erythroid lineage specific and position-independent expression of the globin genes. The LCR is composed of five DNase I hypersensitive sites (HSs), four of which are formed in erythroid cells. The function of the 5'-most site, HS5, remains unknown. To gain insights into its function, mouse HS5 was cloned and sequenced. Comparison of the HS5 sequences of mouse, human, and galago revealed two extensively conserved regions, designated HS5A and HS5B. DNase I hypersensitivity mapping revealed that two hypersensitive sites are located within the HS5A region (designated HS5A(major) and HS5A(minor)), and two are located within the HS5B region (HS5B(major), HS5B(minor)). The positions of each of these HSs colocalize with either GATA-1 or Ap1/NF-E2 motifs, suggesting that these protein binding sites are implicated in the formation of HS5. Gel retardation assays indicated that the Ap1/NF-E2 motifs identified in murine HS5A and HS5B interact with NF-E2 or similar proteins. Studies of primary murine cells showed that HS5 is formed in all hemopoietic tissues tested (fetal liver, adult thymus, and spleen), indicating that this HS is not erythroid lineage specific. HS5 was detected in murine brain but not in murine kidney or adult liver, suggesting that this site is not ubiquitous. The presence of GATA-1 and NF-E2 motifs (which are common features of the DNase I hypersensitive sites of the LCR) suggests that the HS5 is organized in a manner similar to that of the other HSs. Taken together, our results suggest that HS5 is an inherent component of the beta-globin locus control region. PMID- 10534404 TI - The promoter of the Poly(A) binding protein 2 (Pabp2) retroposon is derived from the 5'-untranslated region of the Pabp1 progenitor gene. AB - The mouse Pabp2 retroposon encodes an isoform of poly(A) binding protein that is expressed in meiotic and early haploid spermatogenic cells. In the present study, we have determined the transcription start site of the Pabp2 gene to clarify the source of its promoter, a prerequisite for expression of retroposons and preservation of their function by natural selection. The 5' end of the mouse Pabp2 retroposon exhibits extensive similarity to the entire 5' UTR of the human PABP1 mRNA, but there is no similarity upstream of the transcription start site of the human PABP1 mRNA, indicating that the Pabp2 gene lacks 5' flanking sequences of the parental PABP1 gene. Oligonucleotide-directed RNase H cleavage and 5' rapid amplification of cDNA ends both indicate that the transcription start site of the mouse Pabp2 gene is located approximately 330 bases downstream of the capsite of the PABP1 mRNA, indicating that the Pabp2 promoter is derived from the PABP1 5' UTR. PMID- 10534405 TI - Genomic structure and functional expression of a human alpha(2)/delta calcium channel subunit gene (CACNA2). AB - CACNA2 encodes the alpha(2)/delta subunit of the human voltage-gated calcium channels and is located in the candidate region of malignant hyperthermia susceptibility type 3 (MHS3). We determined the structural organization of CACNA2 by isolation of overlapping genomic DNA clones from a human phage library. The gene consists of at least 40 exons, 2 of which are alternatively spliced, spanning more than 150 kb of genomic DNA. Exons range from 21 to 159 bp, and introns range from 98 bp to at least more than 20 kb. We constructed a full length cDNA and cloned it into a mammalian expression vector. Cotransfection of the CACNA2 cDNA with alpha(1A) and beta(4) cDNA into HEK293 cells led to the expression of Q-type calcium currents. The alpha(2)/delta subunit enhanced the current density 18-fold compared to cells transfected with only alpha(1A) and beta(4) cDNA. The sequence analysis provides the basis for comprehensive mutation screening of CACNA2 for putative MHS3 individuals and patients with other channelopathies. PMID- 10534406 TI - Human eukaryotic initiation factor EIF2C1 gene: cDNA sequence, genomic organization, localization to chromosomal bands 1p34-p35, and expression. AB - We report the cloning and characterization of the human eukaryotic protein translation initiation factor EIF2C1 gene. The human EIF2C1 gene consists of 19 exons and 18 introns that span a region of almost 50 kb. It is located on the short arm of chromosome 1 in the region 1p34-p35. This genomic region is frequently lost in human cancers such as Wilms tumors, neuroblastoma, and carcinomas of the breast, liver, and colon. The human EIF2C1 gene is ubiquitously expressed at low to medium levels. Differential polyadenylation and splicing result in a complex transcriptional pattern. The cDNA sequence is 7478 bp long and contains an extremely large 3' untranslated region of 4799 bp with multiple, short repeated segments composed of mono-, tri-, or quattronucleotides interspersed throughout. The human EIF2C1 gene belongs to a multigene family in human. It is highly conserved during evolution, sharing about 90% identity with rabbit eIF2C and 70% identity with plant AGO1 at the amino acid level. These facts suggest that human EIF2C1 might play an important physiological role. PMID- 10534407 TI - Characterization of copine VII, a new member of the copine family, and its exclusion as a candidate in sporadic breast cancers with loss of heterozygosity at 16q24.3. AB - In a search for candidate tumor suppressor genes within a 650-kb common region of loss of heterozygosity (LOH) at 16q24.3 in breast cancer tissues, a 2.6-kb cDNA, named copine VII (CPNE7), was characterized. The gene is 2654 bp and codes for a 633-residue protein with high homology to the other members of the copine family, such as copine I, copine III, and N-copine. The predicted amino acid sequence contains two copies of a C2 domain in the N-terminus. Since these domains have been found in several membrane-binding proteins involved in different intracellular processes, copine VII was viewed as a potential tumor suppressor gene. Mutation analysis was carried out by single-strand conformation polymorphism analysis of 18 breast tumor tissue samples with ascertained LOH on chromosome 16q24.3. Since only two polymorphisms were identified, no evidence was found to indicate that copine VII is the tumor suppressor gene at 16q24.3 involved in breast cancer. PMID- 10534409 TI - Internal defenses of the snail Biomphalaria glabrata. AB - We describe three distinct types of cells among Biomphalaria glabrata hemocytes: large cells with a tubulo-vesicular compartment, a component of the endocytic system, and with numerous mitochondria and large aggregates of glycogen particles; medium-size cells poor in organelles and glycogen; and small cells with organelles and few secretory granules. Other small hemocytes can be interpreted as juvenile cells. B. glabrata hemocytes contain few enzymes and do not show specific secretory granules, except for a subpopulation of large cells richer in acid phosphatase vesicles. Hemocytes have different aspects corresponding to different physiological states and their transitions: in quiescent hemocytes, the cell cortex is narrow and organelles are scattered in the cytoplasm, both in circulating cells characterized by thin-folded filopods and large macropinocytic vacuoles and in sedentary cells in which extended filopods connect to the extracellular matrix. In stress-activated hemocytes, the cortical region is thickened by polymerization of actin, and organelles are gathered around the nucleus. Fixed phagocytes are components of the connective tissue; the presence of numerous lysosomes and residual bodies and of acid phosphatase and peroxidase activities suggests a high phagocytic activity. PMID- 10534408 TI - The role of destruxins in the pathogenicity of metarhizium anisopliae for three species of insect AB - The entomopathogenic fungus Metarhizium anisopliae var anisopliae produces a family of cyclic peptide toxins, destruxins (DTX), both in culture and in vivo in mycosed insects. The contribution of these insecticidal toxins to the disease process has been investigated in the tobacco hornworm, Manduca sexta (Lepidoptera), the desert locust, Schistocerca gregaria (Orthoptera), and the vine weevil, Otiorhynchus sulcatus (Coleoptera). A significant negative correlation was found between the titer of DTX production in vitro of isolates of M. anisopliae var anisopliae pathogenic for Otiorhynchus and the median lethal time, suggesting a role for the toxin in isolate virulence. The same was true for Manduca-active isolates. A key exception was isolate 703. This is highly virulent for M. sexta, yet does not produce DTX in vitro, grows largely as hyphal fragments in the hemolymph of infected insects, and does not cause host paralysis. These results are discussed in the light of the hypothesis that there are at least two possible virulence strategies among isolates of M. anisopliae var anisopliae pathogenic for Manduca viz the "toxin strategy" and the "growth strategy." For locusts, a strong positive correlation was found only between in vitro toxin production and percentage mortality of individuals in which sporulation did not occur on the cadaver. To account for this, it is suggested that if DTX kills locusts before the fungus has established itself, then the pathogen may not compete effectively with the saprophytic flora and, as a result, fails to sporulate. It is concluded that, in the pathogenesis of M. anisopliae var anisopliae for all three insects, there is a relationship between the titer of DTX production of isolates in vitro and the killing power. Copyright 1999 Academic Press. PMID- 10534411 TI - Internal defenses of the snail Biomphalaria glabrata. AB - Injection of foreign particles (zymosan, latex beads, living bacteria) and substances (ferritine, horseradish peroxidase) in Biomphalaria glabrata induces the rapid appearance of tubular double-helical filaments in the hemolymph. This rapid induction may be due to the polymerization of precursor proteins of the hemolymph. Tubular helical filaments are also observed in normal snails, especially in the hemal spaces of the proximal kidney; they seem to belong to the extracellular matrix and form bundles of a few parallel units associated with fine fibrils. Hemocytes adhere to these bundles by means of the filopodia tip or by small areas of the cell surface; consequently, they may become temporarily sedentary in the hemal spaces. The deposition of helical filaments seems increased in ageing snails and in snails parasitized by Echinostoma caproni. PMID- 10534412 TI - Bioassay to assess the toxicity of bacillus thuringiensis to daikon leaf beetle (Coleoptera: chrysomelidae) AB - We developed a new method to rear daikon leaf beetles, Phaedon brassicae Baly (Coleoptera: Chrysomelidae), on a diet of Cruciferae leaves in a growth chamber. Eggs were stored at 4 degrees C for 30 days without significant loss of viability. A bioassay using artificial diet was developed to standardize the assessment of toxicity of a powdered crystal-spore preparation of Bacillus thuringiensis strain YBT-0618 to P. brassicae larvae. When the diet containing this powder preparation was stored at 4 degrees C for a period of 7 days, the coefficient of variation of the median lethal concentration (LC(50)) values in three replicates was 0.060. The reproducibility, or precision, of the bioassay results was optimal when, at each dilution, 30 neonates were exposed and incubated for 72 h at 25 degrees C, 65-75% RH, and a photoperiod of 12:12 (L:D). Copyright 1999 Academic Press. PMID- 10534413 TI - Water and temperature relations of growth of the entomogenous fungi beauveria bassiana, metarhizium anisopliae, and paecilomyces farinosus AB - The effects of temperature (5-50 degrees C), water availability (0. 998-0.88 water activity, a(w)), and a(w) x temperature interactions (15-45 degrees C) on growth of three entomogenous fungi, Beauveria bassiana, Metarhizium anisopliae, and Paecilomyces farinosus, were evaluated on a Sabouraud dextrose-based medium modified with the ionic solute KCl, the non-ionic solute glycerol, and an inert solute, polyethylene glycol (PEG) 600. The temperature ranges for growth of B. bassiana, M. anisopliae, and P. farinosus were 5-30, 5-40, and 5-30 degrees C, and optimum growth temperatures were 25, 30, and 20 degrees C, respectively. All three species grew over a similar a(w) range (0.90-0.998) at optimum temperatures for growth. However, there were significant interspecies variations in growth rates on media modified with each of the three a(w)-modifying solutes. Growth a(w) optima ranged between 0.99 and 0.97 on KCl-, glycerol-, and PEG 600-modified media for M. anisopliae and P. farinosus. B. bassiana grew optimally at 0.998 a(w), regardless of a(w). Comprehensive two-dimensional profiles of a(w) x temperature relations for growth of these three species were constructed for the first time. The results are discussed in relation to the environmental limits that determine efficacy of entomogenous fungi as biocontrol agents in nature. Copyright 1999 Academic Press. PMID- 10534410 TI - Internal defenses of the snail Biomphalaria glabrata. AB - The three populations of hemocytes and the fixed phagocytes of Biomphalaria glabrata show different affinities for the following injected foreign materials: ferritin, horseradish peroxidase, zymosan particles, latex beads, and live bacteria. Ferritin and horseradish peroxidase, which are internalized by micropinocytosis, induce a moderate activation of hemocytes. Particles trigger a strong activation that consists of mobilization of the cytoskeleton, with formation of pseudopodia and filopodia and modifications of the plasma membrane that enable hemocytes to adhere together, as in encapsulations. Phagocytosis of zymosan and live bacteria induces alterations in mitochondria that may reflect changes in the respiratory activity. Phagocytosis of particles follows different processes: fixed phagocytes ingest latex beads of two different sizes according to the "zipper model", which implies sequential binding between phagocyte receptors and opsonized ligands; zymosan particles are ingested by medium-size hemocytes by a variant of the zipper model in which a limited close contact between cell and particle resembles focal contacts in cell spreading; bacteria or large latex beads are ingested by large hemocytes without apparent close contact following a "trigger model" which resembles macropinocytosis. PMID- 10534414 TI - A noda-like virus isolated from the sweetpotato pest spodoptera eridania (Cramer) (Lep.; noctuidae) AB - A small isometric virus has been isolated from larvae of the sweetpotato pest Spodoptera eridania (Cramer) collected near Pariacoto, Ancash province, Peru. It is designated the Pariacoto virus (PaV). In addition to its high pathogenicity on its natural host Spodoptera eridania, PaV was found to replicate in Spodoptera ochrea (Hampson) larvae but not in Spodoptera frugiperda (Smith) larvae. The size of the viral particle was estimated to be about 30 nm in diameter. Polyacrylamide gel electrophoresis showed a protein of approximately 40.5 kDa. After agarose gel electrophoresis, the viral genome appeared to be bipartite RNA. Gel immunodiffusion tests showed no serological relationship between PaV and Nodamura virus, the type species for insect nodaviruses. Electron microscopy confirmed that viral replication occurs in the cytoplasm. These properties are similar to those of other members of family Nodaviridae, to which the virus is currently assigned. Copyright 1999 Academic Press. PMID- 10534416 TI - Genetic transformation and mutagenesis of the entomopathogenic fungus paecilomyces fumosoroseus AB - We have developed a DNA-mediated transformation system for the entomopathogenic fungus Paecilomyces fumosoroseus based on resistance to the herbicide glufosinate ammonium. The resistance is provided by the bar gene from Streptomyces hygroscopicus and is under the control of the Aspergillus nidulans trpC promoter and terminator sequences. Frequencies of up to 110 transformants/&mgr;g of linearized plasmid DNA were obtained aided by the addition of the nuclease inhibitor, aurintricarboxylic acid (ATA). The transformants were stable for drug resistance for five consecutive vegetative transfers under selective and nonselective conditions. Southern analyses revealed that the transforming DNA integrated into the P. fumosoroseus genome in single and multiple copies. Single copy integration events were generated with higher efficiency by restriction enzyme-mediated integration (REMI), although rates of transformation were generally not as high as those treatments containing ATA. Two mutant strains altered in sporulation capacity and virulence toward the Russian wheat aphid were recovered using this approach. This transformation-based manipulation of P. fumosoroseus will facilitate insertional mutagenesis and the functional analysis of various genes. PMID- 10534415 TI - Bacteria, granulocytomas, and trematode metacercariae in the digestive gland of Mytilus edulis: seasonal and interpopulation variation. AB - During the period October 1983-September 1984, mussels were sampled at monthly intervals from three populations in Denmark. The mussels were prepared for histopathological studies and 20 individuals were randomly chosen from each month and each population for a study of the prevalence and variation of the histological changes. The aim of the study was to test the hypothesis that the prevalence of parasites, bacteria, and granulocytoma differs among blue mussel populations during a year, due to the reproductive cycle and the environment. The only bacterial infection found was Chlamydia sp. and this was rare. Neither sex, month, nor population was an important determinant of the prevalence of bacteria. Metacercariae of the trematode Renicola roscovita had a prevalence of almost 54% in a population from Lilla Baelt but only 4% in a population from Isefjorden. A statistically significant interaction between month and population indicated that the prevalence varied over months but the pattern of variation was different among populations. No other parasites were found. The prevalence of granulocytomas within populations was positively correlated with the degree of anthropogenic contamination of the localities, which might suggest a functional connection. Neither month nor sex had a significant effect. PMID- 10534417 TI - Characterization of a nucleopolyhedrovirus from the black cutworm, agrotis ipsilon (Lepidoptera: noctuidae) AB - The black cutworm, Agrotis ipsilon (Hufnagel) (Lepidoptera: Noctuidae), is a serious localized pest of vegetable and field crops. We have characterized a newly discovered baculovirus, the Agrotis ipsilon multicapsid nucleopolyhedrovirus (AgipMNPV), that was isolated from A. ipsilon in Illinois. Restriction enzyme fragment profiles of AgipMNPV DNA were distinct from those of previously described nucleopolyhedroviruses. Electron microscopy of AgipMNPV infected tissues indicated that nucleocapsids of this virus are multiply enveloped. A. ipsilon was highly susceptible to infection by AgipMNPV and significantly more susceptible to infection by AgipMNPV than by Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV). Host range studies showed that Heliothis virescens and Helicoverpa zea were moderately susceptible to infection; Pseudaletia unipuncta and Spodoptera frugiperda were only partially susceptible, and Anticarsia gemmatalis, Spodoptera exigua, Trichoplusia ni, and Ostrinia nubilalis were not susceptible to infection by AgipMNPV. Because of its high virulence, AgipMNPV has potential as an alternative to chemical insecticides for control of A. ipsilon. Copyright 1999 Academic Press. PMID- 10534418 TI - PPL1c, a virulent mutant bacteriophage useful for identification of paenibacillus larvae subspecies larvae PMID- 10534419 TI - cGMP is decreased after acute ischemia in chronically ischemic canine limbs. AB - BACKGROUND: A chronic partially ischemic state may alter the skeletal muscle response to acute ischemia and free radical formation. METHODS: In order to investigate this hypothesis, a chronic ischemic state was established by ligating the right femoral artery of four mongrel dogs. ABIs were decreased from 1.05 +/- 0.25 preligation to 0.54 +/- 0.14 at 6 weeks (P = 0.04). At the end of 8 weeks, the hindlimb was subjected to 3 h of acute ischemia by clamping the iliac artery. The clamp was then released for 2 h of reperfusion. Plasma samples from the right iliac vein were taken during the ischemia-reperfusion period for analysis of cGMP. Tibialis anterior biopsies for Western analysis of eNOS and iNOS were taken upon completion of reperfusion. Comparisons to control dogs subjected to the acute ischemia and reperfusion without prior femoral artery ligation were made. RESULTS: cGMP levels were increased in the controls at 3 h of ischemia (3539 +/- 350) and 2 h of reperfusion (2880 +/- 269). The chronic ischemia group did not develop a corresponding increase in cGMP at 3 h of ischemia (2762 +/- 251) or after 2 h of reperfusion (2102 +/- 130). Western analysis of eNOS and iNOS revealed similar levels in both groups. Analysis of eNOS revealed 0.6429 +/- 0.086 and 0.5916 +/- 0.072 (densitometric units +/- SEM) for study and control dogs, respectively. Analysis of iNOS revealed 0.3401 +/- 0.067 and 0.2475 +/- 0.066 for study and control dogs, respectively. CONCLUSION: Previous ligation of the femoral artery resulting in chronic partial ischemia in this model demonstrated no increase in cGMP following acute ischemia that was not accompanied by a change in eNOS or iNOS levels. Nitric oxide activity is reflected by cGMP levels, which may increase in response to free radicals in the acute setting of complete ischemia. PMID- 10534421 TI - Jejunal pouch interposition after pylorus-preserving gastrectomy. AB - BACKGROUND: To improve the quality of life of patients undergoing gastrectomy, a nerve-conserving jejunal pouch was interposed after pylorus-preserving gastrectomy (PPG) with vagal nerve preservation. We report the details of the operative technique and the outcome. METHODS: PPG with lymph node dissection was performed, preserving the hepatic, pyloric, and celiac branches of vagal nerve. The jejunum was cut approximately 20 cm distal to the ligament of Treitz. Marginal vessels were not divided in order to preserve the nerves in the jejunum that were used to construct the pouch. A linear stapler was used to perform a side-to-side jejunojejunostomy. A 12-cm-long pouch was formed by firing the stapler twice. The pouch was interposed between the residual parts of the stomach. Postoperatively, the patients were interviewed periodically. A dual phase, dual-isotope radionucleid pouch-emptying study was performed 6 months after surgery. RESULTS: A total of 13 patients underwent the operation. No complication developed. During the first 6 months after surgery, the body weight of the patients was maintained at 91% of the preoperative level. The radioisotope retention rate for the combined pouch and residual stomach was 46% for liquid food and 76% for solid food 120 min after ingestion. The pattern of its emptying was similar to that in healthy individuals. CONCLUSIONS: The pouch-emptying test demonstrated a satisfactory retention capacity and acceptable emptying for the gastric substitute. A reasonably good quality of life has been obtained for patients undergoing PPG with interposition of a nerve-preserving jejunal pouch. PMID- 10534420 TI - Exogenous calcium preconditions myocardium from patients taking oral sulfonylurea agents. AB - We have previously reported that atrial trabeculae from patients taking oral sulfonylurea hypoglycemic agents cannot be preconditioned by transient ischemia, which may, in part, explain the increased cardiovascular mortality historically associated with the use of these agents (J. C. Cleveland et al., 1997, Circulation 96, 29-32). Recently, we reported that clinically accessible and acceptable exogenous Ca(2+) pretreatment protects human atrial trabeculae from subsequent ischemia (B. S. Cain et al., 1998, Ann. Thoracic Surg. 65, 1065-1070). It remains unknown whether this preconditioning strategy could confer protection to trabeculae from patients taking oral sulfonylurea drugs. We therefore hypothesized that exogenous Ca(2+) confers ischemic protection to trabeculae from patients taking oral sulfonylureas. Human atrial trabeculae were suspended in organ baths and field stimulated at 1 Hz, and force development was recorded. Following 90 min equilibration, trabeculae from patients taking oral sulfonylurea agents (n = 6 patients) were subjected to ischemia/reperfusion (I/R; 45/120 min) with or without Ca(2+) (1 mM increase x 5 min) 10 min prior to I/R. I/R decreased postischemic human myocardial contractility in trabeculae from patients on oral hypoglycemics to 15.3 +/- 2.0% baseline developed force (%BDF). Ca(2+) pretreatment increased postischemic human myocardial developed force to 35.3 +/- 2.9 %BDF in these patients (P < 0.05 vs I/R, ANOVA and Bonferroni/Dunn). We conclude that atrial muscle from patients taking oral hypoglycemic agents can be preconditioned with exogenous Ca(2+). This therapy may offer a clinically relevant means to precondition the myocardium of diabetics taking oral hypoglycemic agents prior to clinical interventions such as coronary angioplasty or cardiac bypass. PMID- 10534422 TI - The 21-aminosteroid U74389G enhances hepatic blood flow and preserves sinusoidal endothelial cell function and structure in endotoxin-shocked dogs. AB - BACKGROUND: 21-Aminosteroids are potent anti-inflammatory and antioxidant drugs that provide remarkable endothelial protection in different models of tissue ischemia-reperfusion and inflammation. The effects of 21-aminosteroids in sepsis, a highly inflammatory condition leading to panendothelial activation and injury, are largely uninvestigated. We therefore explored the effects of the 21 aminosteroid U74386G on hepatic blood flow, endothelial cell function, and sinusoidal structure in a canine model of fluid-resuscitated, hyperdynamic endotoxic shock. MATERIALS AND METHODS: Following invasive hemodynamic monitoring and placement of ultrasonic flow probes around the common hepatic artery and the portal vein, 12 anesthetized dogs received 2 mg/kg iv of Escherichia coli endotoxin, followed by generous saline infusion, before randomization into two groups. One group (N = 6) received U74389G as an iv bolus of 80 microg/kg, followed by a continuous infusion of 10 microg/kg. min. The other group (N = 6) received an equivalent volume of vehicle. Hyaluronic acid was measured in plasma for in vivo evaluation of endothelial cell function. Liver biopsies were taken after 4 h of endotoxic shock and prepared for light and electron microscopic examination. RESULTS: Compared with the vehicle-treated controls, U74389G maintained a higher blood flow in the hepatic artery and in the portal vein, without markedly influencing the systemic hemodynamic response. The endotoxin induced increase in plasma hyaluronic acid levels was significantly attenuated following U74389G treatment (70 +/- 14 vs 188 +/- 24 ng/mL after 3 h of endotoxic shock; P < 0.05). Morphological studies showed that the U74389G-treated group had less sinusoidal endothelial cell damage together with a dramatic reduction of neutrophil infiltration into the liver tissue. CONCLUSION: U74389G can preserve the functional and structural integrity of endothelial cells in the hepatic sinusoid during hyperdynamic endotoxic shock. This endothelial-protective effect was associated with a better maintained hepatic blood flow and a significant attenuation of inflammatory liver injury. PMID- 10534423 TI - Differential expression of ileal Na(+)/H(+) exchanger isoforms after enterectomy. AB - BACKGROUND: Na(+)/H(+) exchangers (NHE) are transporters involved in the absorption of NaCl along the gastrointestinal tract. The aim of this study was to determine the expression pattern of the intestinal brush border NHE isoforms 2 and 3 following massive small bowel resection (SBR). Additionally, the effect of epidermal growth factor (EGF) and salivarectomy (removal of the primary source of EGF) on the expression pattern was studied. MATERIALS AND METHODS: ICR mice underwent a proximal SBR or sham surgery and then received either orogastric saline or EGF (50 microg/kg/day). In separate experiments mice underwent salivarectomy followed by SBR or sham. Postoperatively the remaining ileum was isolated and levels of NHE-2 and NHE-3 mRNA and protein were resolved. RESULTS: Following SBR, the expression of both mRNA and protein for NHE-3 increased by approximately 2.5-fold. Treatment with EGF enhanced NHE-3 mRNA in sham animals with further elevation following SBR. The expression of NHE-2 mRNA demonstrated minimal change while protein marginally increased (40%) following SBR. EGF did not affect the expression of NHE-2 mRNA. Salivarectomy did not influence NHE-2 protein expression and inhibited the increased NHE-3 protein expression following SBR. CONCLUSIONS: Following SBR, the expression pattern for brush border NHE isoforms is distinctive. Increased expression of NHE-3 secondary to SBR and/or EGF treatment with loss of this increase following salivarectomy implies a common mechanism to enhance enterocyte proliferation and luminal absorption of NaCl and water. These results suggest that NHE-3 is an important ileal exchanger following SBR. PMID- 10534425 TI - Hypervolemic hemodilution: an alternative to acute normovolemic hemodilution? A mathematical analysis. AB - BACKGROUND: Hypervolemic hemodilution has been proposed as an alternative to normovolemic hemodilution to reduce homologous blood transfusions. So far, convincing data supporting this concept are unknown. MATERIALS AND METHODS: We therefore present a mathematical model calculating the efficacy of hypervolemic, normovolemic, and "no" hemodilution. Hypervolemic hemodilution constituted volume expansion (20% of estimated blood volume) maintained throughout surgery. Normovolemic hemodilution contained isovolemic exchange of blood (40% of estimated blood volume) vs colloid as well as retransfusing blood plus colloid to maintain minimal acceptable hematocrit, e.g., transfusion trigger. To determine the efficacy of each technique maximal allowable blood loss and final postoperative hematocrit were calculated. Maximal allowable blood loss referred to the amount of blood lost during surgery after which homologous blood transfusion became necessary. RESULTS: Recalculating published clinical data strongly validated the formulas used for our model. Hypervolemic hemodilution always revealed lowest maximal allowable blood losses. Normovolemic hemodilution constantly ensured highest maximal allowable blood losses. For blood losses <40% of blood volume, hypervolemic and normovolemic hemodilution provided almost identical final postoperative hematocrits. But in contrast to normovolemic hemodilution, hypervolemic hemodilution did not carry the risk of severe transient, retransfusion-induced hypervolemia. "No" hemodilution always gave lowest final postoperative hematocrits. CONCLUSIONS: Thus, hypervolemic hemodilution cannot replace normovolemic hemodilution to reduce homologous transfusions, but for blood losses <40% of blood volume hypervolemic hemodilution appears to be superior. PMID- 10534424 TI - Echocardiographic and survival studies in mice undergoing endotoxic shock: effects of genetic ablation of inducible nitric oxide synthase and pharmacologic antagonism of platelet-activating factor. AB - Evidence implicating inducible nitric oxide synthase (iNOS) in the alterations of cardiac function characteristic of septic shock has come mostly from studies on anesthetized animals, isolated hearts, cultured myocytes, or hosts treated with pharmacologic inhibitors that lack complete specificity for iNOS. Platelet activating factor (PAF) can participate in the induction of iNOS and has also been implicated in cardiac dysfunction in sepsis. The present studies assessed cardiac function in a model of sepsis in awake mice in which the gene for iNOS was either normal or selectively disrupted. Mice of each genotype were treated with parenteral fluids or with a highly specific antagonist of PAF. Endotoxic shock was induced by challenge with bacterial lipopolysaccharide (LPS) after priming with heat-killed Propionobacterium acnes. Wild-type mice increased stroke volume and cardiac output in response to LPS. These changes were absent in iNOS deficient mice. When treated with parenteral fluids, LPS-challenged wild-type and iNOS-deficient mice both had a marked reduction in cardiac output. Antagonism of PAF had no effect on echocardiographic indices in wild-type mice, but selectively overcame the bradycardia and reduced cardiac output elicited by fluid administration in LPS-shocked, iNOS-deficient mice. Thus, there are major cardiovascular effects of PAF that are shared by rather than mediated by iNOS. Neither complete iNOS deficiency nor antagonism of PAF improved survival, whether tested as single or combined intervention. On the contrary, complete deficiency of iNOS was detrimental to survival. Finally, we tested the hypothesis that iNOS deficiency might improve survival if the deficiency were specific but partial. For this, we used mice with one normal and one disrupted gene for iNOS. No survival advantage was evident for these iNOS heterozygotes. Thus, partial or complete inhibition of iNOS, with or without antagonism of PAF, afforded no evident benefit beyond the previously demonstrated reduction in hypotension. Finally, these studies demonstrate that echocardiography preceded by acclimatization is feasible in unanesthetized mice, a finding which should expand the value of genetically manipulated animals for analysis of cardiac function. PMID- 10534427 TI - The association for academic surgery PMID- 10534426 TI - Glutamine is essential for nitric oxide synthesis by murine macrophages. AB - BACKGROUND: Recent studies suggest an interaction between l-arginine (Arg) and l glutamine (Gln) in the control of nitric oxide (NO) synthesis. Endotoxemia enhances Gln demand and NO production. This study was initiated to investigate the effects of altered Gln availability on the capacity of macrophages to produce NO and the interaction of Gln with l-citrulline (Cit) and Arg in the regulation of endotoxin-stimulated NO synthesis. METHODS: Cultures of RAW 264.7 macrophages in MEM containing Gln (0 to 100 mM) or Arg (0 or 0.6 mM) and supplemented or not with Cit (0.31 to 10 mM) were exposed to Escherichia coli lipopolysaccharide (LPS) at 0.001 and 1 microg/ml. After 24-h incubation, supernatants were evaluated for nitrite concentrations by Greiss reaction as a measure of NO synthesis. RESULTS: LPS stimulated nitrite synthesis in a dose-dependent fashion. Macrophages cultured in Gln-free medium containing Arg (0.6 mM) did not produce NO when stimulated with LPS. In contrast, in the presence of Arg and 0.001 microg/ml LPS, adding as little as 0.31 mM Gln resulted in a 23-fold increase in NO production (from 0.13 +/- 0. 02 to 2.92 +/- 0.06 nmol/ml) (P < 0.0001). Furthermore, a dose-dependent increase in LPS-stimulated nitrite release was observed with increasing amounts of Gln to as much as 1 mM. LPS-stimulated macrophages cultured in Arg-free medium containing Gln (0.31-10 mM) did not produce significant amounts of nitrite. However, in the absence of Arg, increasing extracellular Gln levels to 100 mM in the culture medium resulted in nitrite synthesis (2.39 +/- 0.11 nmol/ml). Detectable levels of nitrite (2.84 +/- 0.21 nmol/ml) were also documented when stimulated macrophages were incubated in culture medium lacking Arg but containing Cit (0.31 mM) and Gln (2 mM). Increasing Cit levels (0.63 to 10 mM) significantly augmented nitrite release (P < 0.05). Once again, no detectable levels of nitrite were observed when macrophages were cultured in Gln-free medium, even when Arg and Cit were present. CONCLUSION: These results suggest that Gln is an essential amino acid for NO synthesis by macrophages and raise the strong possibility that Gln acts with nitric oxide synthase to catalyze the conversion of Arg to NO. The consumption of Gln during sepsis may represent NO production. PMID- 10534428 TI - Association for academic surgery 33rd annual meeting PMID- 10534429 TI - AAS 1999 membership list PMID- 10534430 TI - The association for academic surgery newsletter, volume 8, number 2, fall 1999 PMID- 10534431 TI - Coevolution of a plant host-pathogen gene-for-gene system in a metapopulation model without cost of resistance or cost of virulence AB - A metapopulation model of a one-locus gene-for-gene system in a plant host and a biotrophic pathogen is described. The model allows subpopulations to go extinct, and, due to characteristic differences in life-history strategies, the plant host is assumed to be recolonized from a seed bank, whereas the pathogen is recolonized by migration. It is shown that variation in the gene-for-gene system can be maintained at a noticeable level without assuming cost of resistance or cost of virulence, if the probability of extinction depends on the host mean fitness in the subpopulation. The level of variation in the pathogen population increases with increasing extinction rate, genetic drift and fitness of the infected host, but decreases with increasing migration rate. Generally, these effects are magnified for life cycles in which selection occur before genetic drift and after migration. The metapopulation model generates positive associations between the virulence allele and the resistance allele without assuming cost of resistance or cost of virulence. Copyright 1999 Academic Press. PMID- 10534432 TI - Three mechanisms of host resistance to microparasites-avoidance, recovery and tolerance-show different evolutionary dynamics. AB - Parasite resistant hosts may avoid becoming infected, recover more quickly after infection or survive longer once infected. A model is constructed to examine the evolution of costly host resistance to directly transmitted microparasites and these three distinct mechanisms of avoidance, recovery and tolerance are compared. In each case polymorphism is more likely between very dissimilar strains and resistance (by which we mean the resistant strain alone) is always more likely to occur in hosts with high intrinsic productivity. However, the region where polymorphism occurs is relatively much smaller when resistance is through reduced pathogenicity. In particular, polymorphism with highly resistant strains requires correspondingly high costs. This is in contrast to avoidance or recovery resistance, where polymorphism can also occur when high resistance is associated with small costs due to the inability of highly resistant strains with low susceptibility or high recovery to support the parasite alone and hence resist invasion by the susceptible strain. Relatedly, resistance through avoidance and recovery is favoured in response to less pathogenic parasites. PMID- 10534433 TI - Investigating autocatalytic gene expression systems through mechanistic modeling. AB - A structured model of gene expression, which incorporates the stochastic behavior of cellular processes, was developed to examine the "all-or-none" phenomenon observed in autocatalytic systems (e.g. the lac operon). Autocatalytic expression systems typically have the genes encoding the inducer transport proteins controlled by internal inducer levels, so that transport of the inducer increases production of the transport protein. The model was able to predict the unique behaviors of autocatalytic expression systems that have been experimentally observed and provided valuable insight into the role of population heterogeneity in these systems. The simulations substantiate the importance of stochastic processes on induction of gene expression in autocatalytic systems. The simulation results show that the all-or-none phenomenon is governed largely by random cellular events, and that population-averaged variations in gene expression are due to changes in the frequency of full gene induction in individual cells rather than to uniform variations in gene expression across the entire population. In addition, the model shows how concentrations of inducer too low to induce expression in uninduced cells can maintain induction in pre-induced cultures. A comparison of induction behaviors from an autocatalytic system and a system having constitutive synthesis of the transport protein showed that transport protein levels must be decoupled from inducer control to achieve homogeneous expression of a gene of interest in all cells of a culture. PMID- 10534434 TI - Adaptation of passive rat left ventricle in diastolic dysfunction. AB - This article deals with providing a theoretical explanation for quantitative changes in the geometry, the opening angle and the deformation parameters of the rat ventricular wall during adaptation of the passive left ventricle in diastolic dysfunction. A large deformation theory is applied to analyse transmural stress and strain distribution in the left ventricular wall considering it to be made of homogeneous, incompressible, transversely isotropic, non-linear elastic material. The basic assumptions made for computing stress distributions are that the average circumferential stress and strain for the adaptive ventricle is equal to the average circumferential stress and strain in the normotensive ventricle, respectively. All the relevant parameters, such as opening angle, twist per unit length, axial extension, internal and external radii and others, in the stress free, unloaded and loaded states of normotensive, hypertensive and adaptive left ventricle are determined. The circumferential stress and strain distribution through the ventricular wall are also computed. Our analysis predicts that during adaptation, wall thickness and wall mass of the ventricle increase. These results are consistent with experimental findings and are the indications of initiation of congestive heart failure. PMID- 10534435 TI - Two levels of information in DNA: relationship of Romanes' "intrinsic" variability of the reproductive system, and Bateson's "residue" to the species dependent component of the base composition, (C+G)%. AB - In 1886 Charles Darwin's research associate George Romanes published a paper entitled "Physiological Selection: An Additional Suggestion on the Origin of Species". This was criticized by his Victorian contemporaries and largely ignored by those who followed. However, the recent recognition of two levels of information in DNA suggests that Romanes had solved the major problems with Darwin's theory. It was apparent from the outset that the form of reproductive isolation likely to apply most generally to initial species divergence (hybrid sterility), would depend on differences, not in "primary" information ("genic"), but in "secondary" information ("chromosomal"). This viewpoint, further elaborated by Bateson & Saunders (1902), White (1978), and King (1993), is criticized by the genic school (Coyne & Orr, 1998) because it requires visible differences between chromosomes, and appears not to explain Haldane's rule. However, chromosomal differentiation with respect to the species-dependent component of base composition [(C+G)%; Forsdyke, 1996] appears to resolve these problems. Because it explained so much, it was easy to believe that the genic viewpoint explained everything. Romanes and Bateson thought otherwise. We are only just beginning to recognize what they were trying to tell us. PMID- 10534436 TI - The role of prestress and architecture of the cytoskeleton and deformability of cytoskeletal filaments in mechanics of adherent cells: a quantitative analysis. AB - Mechanical properties of adherent cells were investigated using methods of engineering mechanics. The cytoskeleton (CSK) was modeled as a filamentous network and key mechanisms and corresponding molecular structures which determine cell elastic behavior were identified. Three models of the CSK were considered: open-cell foam networks, prestressed cable nets, and a tensegrity model as a special case of the latter. For each model, the modulus of elasticity (i.e. an index of resistance to small deformation) was given as a function of mechanical and geometrical properties of CSK filaments whose values were determined from the data in the literature. Quantitative predictions for the elastic modulus were compared with data obtained previously from mechanical tests on adherent cells. The open-cell foam model yielded the elastic modulus (10(3)-10(4)Pa) which was consistent with measurements which apply a large compressive stress to the cell. This suggests that bending of CSK filaments is the key mechanism for resisting large compression. The prestressed cable net and tensegrity model yielded much lower elastic moduli (10(1)-10(2)Pa) which were consistent with values determined from equilibrium measurements at low applied stress. This suggests that CSK prestress and architecture are the primary determinants of the cell elastic response. The tensegrity model revealed the possibility that buckling of microtubules of the CSK also contributed to cell elasticity. PMID- 10534437 TI - Maintaining life and health by natural selection of protein molecules. AB - A concept for a life and health-preserving principle is presented, with reference to evolutionary, medical, and biochemical observations. Life comprises two basic phenomena: it unfolds over longer periods at the population level, and is sustained for the duration of individual life spans. The evolution of life within populations by means of natural selection of individuals is central to Darwin's theory of evolution. An important component of maintaining individual life is proposed here to be the natural selection of molecular components-the proteins, a process of preferred removal of denatured and old, synonymous with the selection of younger, functional molecules. The proteins of the cell are committed to fulfilling all the tasks programmed by the genome while continuously maintaining all appropriate cellular functions, including protecting the DNA. Physiological and environmental influences accelerate the breakdown of aged protein molecules, driving this renewal process so that the cell can maintain its protein stock at high-performance levels. The principle of selection makes the incredible dynamics of continual protein turnover, and hence not only the preservation of life, but the maintenance of health in individual beings, comprehensible. Arguments are presented to counter the hypothesis that protein breakdown is a stochastic, random process governed by first-order kinetics. PMID- 10534438 TI - Measuring molecular information. PMID- 10534439 TI - The direct release of nitric oxide by gypenosides derived from the herb Gynostemma pentaphyllum. AB - Herbal medicines are increasingly being utilized to treat a wide variety of disease processes. Gypenosides are triterpenoid saponins contained in an extract from Gynostemma pentaphyllum and are reported to be effective in the treatment of cardiovascular diseases; however, the mechanism underlying the therapeutic effect is not known. We tested the hypothesis that gypenosides extracted from G. pentaphyllum elicit vasorelaxation through the direct release of endothelium derived nitric oxide. Nitric oxide production in bovine aortic endothelial cells grown under standard tissue culture conditions was quantitated using a chemiluminescence method. Arterial vasomotion was assayed using isolated porcine coronary artery rings under standard isometric recording conditions. The extract of G. pentaphyllum at 0.1-100 microg/ml elicited concentration-dependent vasorelaxation of porcine coronary rings that was antagonized by the nitric oxide synthase inhibitor N(G)-nitro-l-arginine methyl ester. Indomethacin had no significant effect on G. pentaphyllum-induced relaxation. The G. pentaphyllum extract elicited a concentration-dependent increase in nitric oxide production from cultured bovine aortic endothelial cells. At the concentrations utilized, there was no morphological evidence for cellular toxicity. These results demonstrate that extracts of G. pentaphyllum directly stimulate nitric oxide release, but not prostanoid production. Nitric oxide production in response to gypenosides may be one mechanism whereby this herbal medicine elicits its therapeutic effects. PMID- 10534440 TI - Reproductive function in female mice lacking the gene for endothelial nitric oxide synthase. AB - Nitric oxide (NO) acts as a neuronal messenger in both the central and peripheral nervous systems and has been implicated in reproductive physiology and behavior. Pharmacological inhibition of nitric oxide synthase (NOS) with the nonspecific NOS inhibitor, l-N(G)-nitro-Arg-methyl ester (l-NAME), induced deficits in both the number of ovarian rupture sites and the number of oocytes recovered in the oviducts of mice. Female neuronal NOS knockout (nNOS-/-) mice have normal numbers of rupture sites, but reduced numbers of oocytes recovered following systemic injections of gonadotropins, suggesting that NO produced by nNOS accounts, in part, for deficits in ovulatory efficiency observed after l-NAME administration. Additionally, endothelial NOS knockout (eNOS-/-) mice have reduced numbers of ovulated oocytes after superovulation. Because endothelial NOS has been identified in ovarian follicles, and because of the noted reduced breeding efficiency of eNOS-/- mice, the present study sought to determine the role of NO from eNOS in mediating the number of rupture sites present after ovulation. Estrous cycle length and variability were consistently reduced in eNOS-/- females. The number of rupture sites was normal in eNOS-/- mice under natural conditions and after administration of exogenous GnRH. After exogenous gonadotropin administration, eNOS-/- females displayed a significant reduction in the number of ovarian rupture sites. Female eNOS-/- mice also produced fewer pups/litter compared to WT mice. These data suggest that NO from endothelial sources might play a role in mediating rodent ovulation and may be involved in regulation of the timing of the estrous cycle. PMID- 10534441 TI - Direct effect of temperature on the catalytic activity of nitric oxide synthases types I, II, and III. AB - The effect of temperature (between 5.0 and 45.0 degrees C) on the catalytic activity of nitric oxide synthases types I, II, and III (NOS-I, NOS-II, and NOS III, respectively) has been investigated, at pH 7.5. The value of V(max) for NOS I activity increases from 1.8 x 10(1) pmol min(-1) mg(-1), at 5.0 degrees C, to 1.8 x 10(2) pmol min(-1) mg(-1), at 45.0 degrees C; on the other hand, the value of K(m) (=4.0 x 10(-6) M) is temperature independent. Again, the value of V(max) for NOS-II activity increases from 8.0 pmol min(-1) mg(-1), at 7.0 degrees C, to 5.4 x 10(1) pmol min(-1) mg(-1), at 40.0 degrees C, the value of K(m) (=1.8 x 10( 5) M) being unaffected by temperature. Temperature exerts the same effect on NOS I and NOS-II activity, as shown by the same values of DeltaH(V(max)) (=4.2 x 10(1) kJ mol(-1)), DeltaH(K(m)) (=0 kJ mol(-1)), and DeltaH((V(max))(/K(m))()) (=4.2 x 10(1) kJ mol(-1)). On the contrary, the value of K(m) for NOS-III activity decreases from 3.8 x 10(-5) M, at 10.0 degrees C, to 1.6 x 10(-5) M, at 40.0 degrees C, the value of V(max) (=6.8 x 10(1) pmol min(-1) mg(-1)) being temperature independent. Present results indicate that temperature influences directly NOS-I and NOS-II activity independently of the substrate concentration, the values of K(m) being temperature independent. However, when l-arginine level is higher than 2 x 10(-4) M, as observed under in vivo conditions, NOS-III activity is essentially unaffected by temperature, the substrate concentration exceeding the value of K(m). As a whole, although further studies in vivo are needed, these observations seem to have potential physiopathologic implications. PMID- 10534442 TI - Different distributions of nitric oxide synthase-containing neurons in the mouse and rat hypothalamus. AB - The distribution of nitric oxide synthase-containing neurons was studied in the rat and mouse hypothalamus by immunohistochemistry and NADPH-diaphorase histochemistry. Immunostaining and NADPH-diaphorase staining of hypothalamic neurons were comparable in all hypothalamic nuclei of either species except in the arcuate nucleus that stained positive for nitric oxide synthase immunoreactivity but negative for NADPH-diaphorase reactivity. The presence of nitric oxide synthase-immunopositive neurons in the arcuate nucleus was confirmed by nitric oxide synthase immunofluorescence viewed under the confocal microscope at 1 microm thickness. Cross-species comparison showed that, in general, the number and intensity of nitric oxide synthase-containing neurons were much higher in the rat than in the mouse hypothalamus. Differences in the distribution of nitric oxide synthase-containing neurons between these two rodents were found in most hypothalamic nuclei. In particular, two dense clusters of nitric oxide synthase-containing neurons were found in the paraventricular and supraoptic nuclei of the rat hypothalamus in contrast to their scarcity in the same nuclei of the mouse hypothalamus. PMID- 10534443 TI - Production and storage of nitric oxide in adaptation to hypoxia. AB - Adaptation to hypobaric hypoxia is known to exert multiple protective effects related with nitric oxide (NO). However the effect of adaptation to hypoxia on NO metabolism has remained unclear in many respects. In the present work we studied the interrelation between NO production and storage in the process of adaptation to hypoxia. The NO production was determined by the total nitrite/nitrate concentration in rats plasma. The volume of NO store was evaluated in vitro by the magnitude of isolated aorta relaxation to diethyldithiocarbamate. It was shown that both the nitrite/nitrate level and the NO store increased as adaptation to hypoxia developed. Furthermore, the NO store volume significantly correlated with plasma nitrite/nitrate. Therefore, adaptation to hypoxia stimulates NO production and storage and these effects can potentially underlie NO-dependent beneficial effects of adaptation. PMID- 10534444 TI - Diminished nitric oxide production following administration of transforming growth factor-beta1 using a noninflammatory wound repair model. AB - The effects of transforming growth factor-beta1 (TGF-beta1) on systemic nitric oxide (NO) production and wound repair were evaluated using a noninflammatory mouse perforated mesentery connective tissue repair model. Urinary nitrates were monitored as a measure of NO production. TGF-beta1 [bovine serum albumin (BSA) carrier and TGF-beta1 BSA carrier free] were administered on days 0 and 1 and were evaluated in mice over a 10-day period. A significant decrease in the average postwounding urinary nitrate levels compared to prewounding levels was observed within the TGF-beta1 treatment group animals (P 5 keV) by as much as 1.3 rad. The phase lags of the second type of QPO are more complex. The soft photons (<5 keV) of the 3 and 12 Hz QPOs lag the hard photons (>5 keV) by as much as 1.0 rad. However, the soft photons of the 6 Hz QPO precede the hard ones by as much as 0.6 rad. This means that different harmonics of this type of QPO have different signs for their phase lags. This unusual behavior is hard to explain when the lags are due to light-travel time differences between the photons at different energies, e.g., in a Comptonizing region surrounding the area in which the QPOs are formed. PMID- 10534456 TI - A New Supernova Remnant Coincident with the Slow X-Ray Pulsar AX J1845-0258. AB - We report on Very Large Array observations in the direction of the recently discovered slow X-ray pulsar AX J1845-0258. In the resulting images, we find a 5&arcmin; shell of radio emission; the shell is linearly polarized with a nonthermal spectral index. We classify this source as a previously unidentified, young (<8000 yr) supernova remnant (SNR), G29.6+0.1, which we propose is physically associated with AX J1845-0258. The young age of G29.6+0.1 is then consistent with the interpretation that anomalous X-ray pulsars (AXPs) are isolated, highly magnetized neutron stars ("magnetars"). Three of the six known AXPs can now be associated with SNRs; we conclude that AXPs are young ( less, similar10,000 yr) objects and that they are produced in at least 5% of core collapse supernovae. PMID- 10534457 TI - The ISO/SWS Spectrum of IRC +10216: The Vibrational Bands of C2H2 and HCN. AB - We present the Infrared Space Observatory/Short-Wavelength Spectrometer full grating resolution spectrum of IRC +10216, which is dominated by strong absorption/emission bands of C2H2 and HCN. All C2H2 bands and the strong near infrared stretching bands of HCN are observed in absorption, whereas the fundamental, hot, and combination bands of HCN involving the nu2 bending mode around 14 um are observed in emission. Particularly strong is the HCN nu2=20- >nu2=11 vibrational transition at 14.3 um. The most plausible mechanism for such emission is the radiative pumping of molecules from the ground to the nu2=20 state (7.1 um) followed by radiative decay: nu2=20-->nu2=11. We present detailed models for HCN that verify the efficiency of the mentioned effect. The HCN abundance inferred from these models is &parl0;1.5-3&parr0;x10-5. PMID- 10534458 TI - Turbulent Convection: Comparing the Moment Equations to Numerical Simulations. AB - The nonlocal hydrodynamic moment equations for compressible convection are compared to numerical simulations. Convective and radiative flux typically deviate less than 20% from the three-dimensional simulations, while mean thermodynamic quantities are accurate to at least 2% for the cases we have investigated. The moment equations are solved in minutes rather than days as required on standard workstations. We conclude that this convection model has the potential to considerably improve the modeling of convection zones in stellar envelopes and cores, in particular those of A and F stars. PMID- 10534459 TI - Origin of the Universal Correlation between the Flare Temperature and the Emission Measure for Solar and Stellar Flares. AB - We present a theory to explain the observed universal correlation between flare temperature T and emission measure EM=n2V for solar and stellar flares (including solar microflares observed by Yohkoh as well as protostellar flares observed by ASCA), where n is the electron density and V is the volume. The theory is based on a magnetic reconnection model with heat conduction and chromospheric evaporation, assuming that the gas pressure of a flare loop is comparable to the magnetic pressure. This theory predicts the relation EM~B-5T17/2, which explains well the observed correlation between EM and T in the range of 6x106 K < T<108 K and 104450% by QCA. IVUS predictors were minimum and mean in-stent area, stent length, and in-stent diameter. Multiple models were constructed with multivariate logistic regression analysis. The model containing minimum in-stent area and stent length best fit the Hosmer-Lemeshow goodness-of-fit test. This model was used to construct a reference chart to calculate the expected 6-month restenosis rate. CONCLUSIONS: The expected 6-month in-stent restenosis rate after stent implantation for short lesions in relatively large vessels can be predicted by use of in-stent minimal area (which is inversely related to restenosis) and stent length (which is directly related to restenosis), both of which can be read from a simple reference chart. PMID- 10534465 TI - Transient entrainment of bundle-branch reentry by atrial and ventricular stimulation: elucidation of the tachycardia mechanism through analysis of the surface ECG. AB - BACKGROUND: Different responses to entrainment have been reported in relation to the pacing site of a variety of tachycardias. However, transient entrainment of bundle-branch reentrant tachycardia (BBRT) has not been investigated systematically. METHODS AND RESULTS: We attempted entrainment of 13 BBRTs in 9 patients by pacing first the right ventricle and then the right atrium. The initial pacing cycle length (CL) was 10 ms faster than the tachycardia CL. Subsequent pacing sequences were performed with 5- to 10-ms CL decrements until tachycardia termination or loss of postatropine 1:1 AV conduction. Both full ventricular-paced and AV-conducted QRS complex references were obtained during sinus rhythm pacing from the same sites and with similar CL as during entrainment. Transient entrainment was achieved by ventricular and atrial stimulation in 11 and 8 tachycardias, respectively. Constant fusion was always present during entrainment by ventricular stimulation. There was no change in the QRS complex (orthodromically concealed fusion) during entrainment by atrial stimulation in 6 of 6 tachycardias with left bundle-branch block morphology and in 1 of 2 tachycardias with right bundle-branch block morphology. CONCLUSIONS: BBRT, especially if it has a left bundle-branch block morphology, can be differentiated from other wide-QRS-complex tachycardia mechanisms through analysis of the ECGs recorded during tachycardia entrainment by atrial and ventricular stimulation. This diagnostic approach may be especially useful when it is difficult to record a stable or sufficiently sized His bundle electrogram or when spontaneous changes in the ventricular CL precede similar changes in the His bundle CL. PMID- 10534467 TI - Usefulness of a tilt training program for the prevention of refractory neurocardiogenic syncope in adolescents: A controlled study. AB - BACKGROUND: Recurrent syncope represents a debilitating disorder and quality of life deteriorates as a function of recurrence of symptoms. Although the administration of beta-blockers, vasoconstrictors, fludrocortisone, and serotonin reuptake inhibitors may be helpful in preventing episodes, many patients are intolerant of or respond poorly to these agents. Orthostatic training has been reported to be effective in preventing refractory syncope. Thus, to determine whether a tilt training program could prevent symptoms in adolescents, the following controlled study was undertaken. METHODS AND RESULTS: Forty-seven consecutive adolescents (18 male and 29 female, mean age 16.0+/-2.2 years) with recurrent syncope and positive head-up tilt test refractory to previous traditional therapies were distributed between 2 groups, depending on their consent (24 patients) or refusal (controls, 23 patients) to enter the program. Orthostatic training was started, in the presence of a family member, with a series of 5 in-hospital sessions. The 24 patients and their relatives were then instructed to perform the tilt training at home by standing against a wall twice a day for a planned duration of up to 40 minutes, depending on the in-hospital orthostatic tolerance. Head-up tilt response was reevaluated after 1 month, and the clinical effect was noted over a mean follow-up of 18. 2+/-5.3 months (range 15 to 23); 26.1% of patients in the control group and 95.8% of patients in the training group became tilt-negative (P<0.0001). Spontaneous syncope was observed in 56.5% versus 0% in the control and training group, respectively (P<0.0001). CONCLUSIONS: Orthostatic training was found to significantly improve symptoms of adolescents with neurocardiogenic syncope unresponsive to or intolerant of traditional medications. Twice-a-day training sessions of 40 minutes were well accepted by patients. PMID- 10534468 TI - Circulating insulin and insulin growth factor-1 are independent determinants of left ventricular mass and geometry in essential hypertension. AB - BACKGROUND: It is unclear whether insulin and insulin-like growth factor-1 (IGF 1) are independent determinants of left ventricular (LV) mass in essential hypertension. METHODS AND RESULTS: We studied 101 never-treated nondiabetic subjects with essential hypertension. All had 24-hour noninvasive ambulatory blood pressure (ABP) monitoring and a 75-g oral glucose tolerance test. We determined fasting glucose, insulin, and IGF-1 and postload glucose and insulin 2 hours after glucose. Insulin resistance was estimated by the homeostasis model assessment (HOMA(IR)) formula. LV mass showed an association with body mass index (BMI) (r=0.47; P<0.01), postload insulin (r=0.54; P<0.01), HOMA(IR) (r=0.39; P<0.01), and IGF-1 (r=0. 43; P<0.01) and a weaker association with average 24 hour systolic and diastolic ABPs (r=0.29 and r=0.26; P<0.05) and basal insulin (r=0.31; P<0.05). Relative wall thickness was positively related to IGF-1 (r=0.39; P<0.01) but not to fasting or 2-hour postload insulin, HOMA(IR), and glucose. In a multiple regression analysis, the final LV mass model (R(2)=0.64) included IGF-1, postload insulin, average 24-hour systolic ABP, sex, and BMI. IGF 1 and postload insulin accounted for >40% of variability of LV mass. The final model (R(2)=0.36) for relative wall thickness included IGF-1 (16% total explained variability), average 24-hour systolic ABP, sex, BMI, and age but not insulin and HOMA(IR). CONCLUSIONS: These data indicate that insulin and IGF-1 are powerful independent determinants of LV mass and geometry in untreated subjects with essential hypertension and normal glucose tolerance. PMID- 10534466 TI - Use of electroanatomic mapping to delineate transseptal atrial conduction in humans. AB - BACKGROUND: Interaction between wave fronts in the right and left atrium may be important for maintenance of atrial fibrillation, but little is known about electrophysiological properties and preferential routes of transseptal conduction. METHODS AND RESULTS: Eighteen patients (age 44+/-12 years) without structural heart disease underwent right atrial electroanatomic mapping during pacing from the distal coronary sinus (CS) or the posterior left atrium. During distal CS pacing, 9 patients demonstrated a single transseptal breakthrough near the CS os, 1 patient in the high right atrium near the presumed insertion of Bachmann's bundle and 1 patient near the fossa ovalis. The mean activation time from stimulus to CS os was 48+/-15 ms compared with 86+/-15 ms to Bachmann's bundle insertion (P<0.01) and 59+/-23 ms to the fossa ovalis (P=NS and P<0.01, respectively). During left atrial pacing, the earliest right atrial activation was near Bachmann's bundle in 5 and near the fossa ovalis in 4 patients. The activation time from stimulus to CS os was 70+/-15 ms compared with 47+/-16 ms to Bachmann's bundle (P<0.01) and 59+/-25 ms to the fossa ovalis (P=NS). Whereas the total septal activation time was not significantly different during CS pacing compared with left atrial pacing (41+/-16 versus 33+/-17 ms), the total right atrial activation time was longer during CS pacing (117+/-49 versus 79+/-15 ms; P<0.05). CONCLUSIONS: Three distinct sites of early right atrial activation may be demonstrated during left atrial pacing. These sites are in accord with anatomic muscle bundles and may have relevance for maintenance of atrial flutter or fibrillation. PMID- 10534469 TI - Central pulmonary artery lesions in chronic obstructive pulmonary disease: A transesophageal echocardiography study. AB - BACKGROUND: In patients with acute pulmonary embolism, transesophageal echocardiography (TEE) often reveals presumably thrombotic lesions within the central pulmonary arteries (CPAs). These CPA lesions, when found in patients with primary pulmonary hypertension, have been attributed to in situ thrombosis or atherosclerosis. We hypothesized that similar CPA lesions may also develop in patients with chronic obstructive pulmonary disease (COPD) in the absence of pulmonary embolism. METHODS AND RESULTS: We examined by TEE 25 patients with COPD and 27 control patients with left heart disease. None of the patients had previous pulmonary embolism or ileofemoral and popliteal vein thrombosis. By use of TEE, CPA lesions were found in 12 COPD patients (48%) and 2 control patients (7.4%) (P<0.01). When CPA lesions were subdivided into types 1 (protruding and mobile) and 2 (wall-adherent), type 1 lesions proved to be uncommon, being found within the pulmonary trunk in 12% and 3.7% of COPD and control patients, respectively (P=NS). Conversely, type 2 lesions, which were always localized in the right pulmonary artery, were frequent in COPD patients (36%) and rare in control patients (3.7%) (P<0.01). When available, helical CT and MR angiography confirmed TEE findings, supporting an atherosclerotic origin of type 2 lesions, which were different from typical thrombotic lesions. FEV(1)/FVC ratio, RV/TLC ratio, PaO(2), hematocrit value, and pulmonary artery systolic pressure were not significantly different in COPD patients with and without CPA lesions. At TEE, however, COPD patients with CPA lesions showed a larger size of the main and right pulmonary arteries. CONCLUSIONS: TEE often reveals CPA lesions in stable patients with COPD even in the absence of significant pulmonary hypertension and not in close relation with the severity of pulmonary dysfunction. PMID- 10534470 TI - Regression of atherosclerosis induced by liver-directed gene transfer of apolipoprotein A-I in mice. AB - BACKGROUND: The ability of apolipoprotein (apo)A-I to induce regression of preexisting atherosclerotic lesions has not been determined, and a mouse model of atherosclerosis regression has not yet been reported. METHODS AND RESULTS: LDL receptor-deficient mice were fed a western-type diet for 5 weeks to induce atherosclerotic lesions. A second-generation recombinant adenovirus encoding human apoA-I or a control adenovirus were injected intravenously in order to express apoA-I in the liver. Three days after injection, total apoA-I levels in mice injected with the apoA-I-expressing adenovirus were 216+/-16.0 mg/dL, compared with 68.0+/-3.0 mg/dL in control virus-injected mice (P<0.001). HDL cholesterol levels in mice injected with the AdhapoA-I vector 7 days after injection were 189+/-21.0 mg/dL, compared with 123+/-8.0 mg/dL in control virus injected mice (P<0.02). Total and non-HDL cholesterol levels did not differ between the 2 groups. Atherosclerotic lesion area was quantified by en face analysis of the aorta and cross-sectional analysis of the aortic root. Compared with baseline mice, atherosclerosis progressed in mice injected with the control adenovirus. In contrast, in mice expressing apoA-I compared with baseline mice, total en face aortic lesion area was reduced by 70% and aortic root lesion was reduced by 46%. Expression of apoA-I was associated with a significant reduction in the fraction of lesions occupied by macrophages and macrophage-derived foam cells. CONCLUSIONS: Liver-directed gene transfer of human apoA-I resulted in significant regression of preexisting atherosclerotic lesions in LDL receptor deficient mice as assessed by 2 independent methods. PMID- 10534471 TI - Contribution of endothelin-1 to myocardial injury in a murine model of myocarditis: acute effects of bosentan, an endothelin receptor antagonist. AB - BACKGROUND: Endothelin (ET) is one of the most important contributing factors in the pathophysiology of cardiovascular diseases. However, little is known about its role in myocarditis. METHODS AND RESULTS: Four-week-old DBA/2 mice were inoculated with the encephalomyocarditis virus. Expression levels of ET converting enzyme-1 (ECE-1) and prepro-ET-1 mRNA were significantly increased at 7 and 14 days after virus inoculation. Plasma and myocardial ET-1 levels were significantly higher in infected than noninfected mice between 5 and 14 days after virus inoculation. Immunohistochemical analyses revealed that not only endothelial cells and myocytes but also infiltrating mononuclear cells produced ET-1 protein at 7 days. Oral bosentan, a mixed ET-1 receptor antagonist, was administered after virus inoculation in doses of 0 (control group), 10, or 100 mg. kg(-1). d(-1), and the animals were killed on day 14. Mean heart weight/body weight ratios were 8.3+/-1.8 versus 11.2+/-2.4 versus 10. 8+/-2.4 in the bosentan 100 mg. kg(-1). d(-1) versus 10 mg. kg(-1). d(-1) versus control groups, respectively (P<0.05). Corresponding histological scores for myocardial necrosis were 2.0+/-0.2 versus 2. 9+/-0.3 versus 3.0+/-0.4 (P<0.05), and cellular infiltration scores were 2.3+/-0.3 versus 2.9+/-0.4 versus 3.3+/-0.4 (P<0.05). Animals killed on day 5 had significantly smaller necrotic areas after treatment with bosentan 100 mg. kg(-1). d(-1) than the group treated with a lower dose or the control group, despite the absence of differences in virus titers. CONCLUSIONS: This study suggests that ET-1 plays an important pathophysiological role in viral myocarditis. Treatment with bosentan had a cardioprotective effect without modifying viral replication. PMID- 10534472 TI - Local L-arginine delivery after balloon angioplasty reduces monocyte binding and induces apoptosis. AB - BACKGROUND: Local administration of L-arginine after balloon angioplasty has been shown to enhance NO generation and inhibit lesion formation. In this study, we assessed the mechanisms by which local delivery of L-arginine inhibits lesion formation. METHODS AND RESULTS: New Zealand White rabbits (n=56) were fed a 1% cholesterol diet. After 1 week, both iliac arteries were balloon-denuded, and a local drug delivery catheter was introduced into both iliac arteries to deliver either L-arginine (800 mg/5 mL with and without 100 microCi L-[2,3-(3)H] arginine) or saline. Monocyte-endothelial interaction was assessed by functional binding assay; NO activity was measured by chemiluminescence. Intramural administration of radioactively labeled L-arginine led to significantly higher counts in comparison to the contralateral segment for up to 1 week after delivery (676+/-223 versus 453+/-93 cpm/mg; P<0.02); this was associated with significantly higher NO levels in the L-arginine-treated segments (394.4+/-141.6 versus 86.3+/-34.3 nmol/mg; P<0.01). Even after 2 to 3 weeks, monocyte binding was significantly decreased by treatment with L-arginine as compared with saline infusion (P<0.01). After 4 weeks, there was a 9-fold greater number of apoptotic cells in the vessel wall of L-arginine as compared with the saline-treated segments (P<0.05). CONCLUSIONS: Intramural delivery of L-arginine immediately after angioplasty causes a sustained increase in tissue L-arginine levels associated with enhancement of local NO synthesis. The local increase in NO synthesis is associated with an attenuation of monocyte binding and increased apoptosis of resident macrophages. This treatment strategy could be valuable for the prevention and management of restenosis. PMID- 10534474 TI - Images in cardiovascular medicine. Sinus of valsalva aneurysm with rupture into the interventricular septum and left ventricular cavity. PMID- 10534475 TI - Transatlantic Conference on Clinical Trial Guidelines in Peripheral Arterial Disease: clinical trial methodology. Basel PAD Clinical Trial Methodology Group. AB - Guidelines for the clinical development of drugs in peripheral arterial disease (PAD) have been issued by the Food and Drug Administration for the United States and by the regulatory agency of the European Union for Europe. With increasing globalization, transatlantic cooperation in drug research and development is essential for the future and would be substantially facilitated by the existence of transatlantic guidelines. A conference was held in Basel, Switzerland, in November 1997 to discuss the scientific background of the existing guidelines on the basis of published evidence and the extensive knowledge of clinical investigators and experienced regulators. The meeting was attended by 52 invited experts from the United States and Europe, as well as by representatives from the 2 regulatory authorities. The main conclusions from the meeting are presented and may serve as a reference for the future development of transatlantic guidelines for the evaluation of pharmacotherapy in PAD. PMID- 10534476 TI - Ulceration and stenosis of internal carotid artery imaged by convergent color Doppler. PMID- 10534473 TI - Digoxin delays recovery from tachycardia-induced electrical remodeling of the atria. AB - BACKGROUND: Atrial fibrillation (AF) induces electrical remodeling, which is thought to be responsible for the low success rate of antiarrhythmic treatment in AF of longer duration. Electrical remodeling seems to be related to tachycardia induced intracellular calcium overload. Due to its vagomimetic action, digoxin is widely used to control the ventricular rate during AF, but it also increases intracellular calcium. On the basis of these characteristics, we hypothesized that digoxin would aggravate tachycardia-induced electrical remodeling. METHODS AND RESULTS: We analyzed the atrial effective refractory period (AERP) at cycle lengths of 430, 300, and 200 ms during 24 hours of rapid atrio/ventricular (300/150 bpm) pacing in 7 chronically instrumented conscious goats treated with digoxin or saline. Digoxin decreased the spontaneous heart rate but had no other effects on baseline electrophysiological characteristics. In addition to a moderate increase in the rate of electrical remodeling during rapid pacing, digoxin significantly delayed the recovery from electrical remodeling after cessation of pacing (at 430, 300, and 200 ms: P=0. 001, P=0.0015, and P=0.007, respectively). This was paralleled by an increased inducibility and duration of AF during digoxin. Multivariate analysis revealed that both a short AERP and treatment with digoxin were independent predictors of inducibility (P=0.001 and P=0.03, respectively) and duration (P=0.001 for both) of AF. CONCLUSIONS: Dioxin aggravates tachycardia-induced atrial electrical remodeling and delays recovery from electrical remodeling in the goat, which increases the inducibility and duration of AF. PMID- 10534477 TI - Percutaneous transluminal coronary angioplasty reverses vasoconstriction of stenotic coronary arteries in hypertensive patients. PMID- 10534478 TI - Renin-angiotensin system and blood pressure. PMID- 10534479 TI - Increased membrane and soluble P-selectin in atrial fibrillation. PMID- 10534480 TI - Familial dilated cardiomyopathy. PMID- 10534481 TI - Novel approaches to unravel the genesis of glomerulosclerosis by new methodologies in molecular biology and molecular genetics. PMID- 10534482 TI - The proinflammatory role of hyaluronan-CD44 interactions in renal injury. PMID- 10534483 TI - What is new in primary hyperoxaluria? PMID- 10534484 TI - V2-vasopressin receptor antagonists-mechanism of effect and clinical implications in hyponatraemia. PMID- 10534485 TI - Blood pressure control in type 2 diabetes--what does the United KingdomProspective Diabetes Study (UKPDS) tell us? PMID- 10534486 TI - Assessing ambulatory blood pressure in renal diseases: facts and concerns. PMID- 10534487 TI - Dosing angiotensin II blockers--beyond blood pressure. PMID- 10534488 TI - Dialysing the patient with acute renal failure in the ICU: the emperor's clothes? PMID- 10534489 TI - Selective percutaneous ethanol injection therapy (PEIT) of the parathyroid in chronic dialysis patients--the Japanese strategy. Japanese Working Group on PEIT of Parathyroid, Tokyo, Japan. PMID- 10534490 TI - Which diet for diabetic patients with chronic renal failure? PMID- 10534491 TI - Chloramine, a sneaky contaminant of dialysate. PMID- 10534492 TI - The disappointing results of PTCA in the uraemic patient--are stents the answer? PMID- 10534493 TI - Angiotensin II and oxidized LDL: an unholy alliance creating oxidative stress. PMID- 10534494 TI - Critical care dialysis--a Gordian knot (but is untying the right approach?). PMID- 10534495 TI - Is there material hazard to treatment with intravenous iron? PMID- 10534496 TI - Can antihypertensive medications control BP in haemodialysis patients: yes or no? PMID- 10534497 TI - Festschrift for Professor Claude Jacobs. Nephrotoxicity of contrast media. PMID- 10534498 TI - Festschrift for Professor Claude Jacobs. Recent developments in conductivity monitoring of haemodialysis session. AB - On-line monitoring of dialysate conductivity is now a standard equipment (called 'Diascan') of the dialysis monitor Integra (Hospal, Italy). From the record of the dialysate conductivity at the dialyser inlet and outlet, the Diascan calculates the values of patient's plasma conductivity and of ionic dialysance which is a weighed average of the dialysances of all ions of quantitative importance in plasma and dialysate. Because there is an equivalence between the transfer characteristics of urea and electrolytes, the ionic dialysance reflects the urea clearance corrected for recirculation. Because the conductivity of a solution is related to the concentrations of the ions and thus to the effective osmolality, the plasma conductivity is a reflection of the plasma sodium concentration. The determination of ionic dialysance and plasma conductivity by the Diascan module is fully automatic and totally inexpensive, does not require any blood or dialysate sampling and therefore can be repeated every 15 or 30 min during each dialysis session. Some clinical applications of conductivity modelling are presented: (i) the repeated measurement of ionic dialysance allows the quantification of the dialysis dose actually delivered to the patient from the beginning of the session; (ii) the measurement of ionic dialysance with blood lines in normal and reversed positions permits the easy estimation of the blood flow rate in the vascular access of the haemodialysed patient; (iii) the on-line monitoring of ionic dialysance allows the development of new methods of haemodialysis with simultaneous infusion of ions; (iv) the on-line monitoring of ionic dialysance and patient's plasma conductivity facilitates the automatic optimization of the dialysate conductivity for each individual patient. PMID- 10534499 TI - Preliminary report of the action in Turkey of the International Society of Nephrology Renal Disaster Relief Task Force. PMID- 10534500 TI - Opinions regarding outcome differences in European and US haemodialysis patients. AB - STUDY GOAL AND DESIGN: The aim of this evaluation was to understand why outcomes seem to be different in different parts of the world. In an attempt to look at this question from a point of view other than that necessarily adopted by epidemiological studies, we decided to explore the personal opinion of a selected group of American (US) and European (EU) experts by means of a simple questionnaire. A 13-item questionnaire was sent to 14 internationally recognized opinion leaders in the field of haemodialysis: all seven Europeans and five of the seven Americans responded. The answers to each question were stratified in order to highlight the key differences between the experts in the different continents. RESULTS: Ten of the 12 respondents (six EU and four US) said that dialysis outcomes are better in Europe; nine (six EU and three US) confirmed their opinion after taking patient characteristics into account. When asked to suggest reasons for this difference, the highest score was given to the quality of procedures and medical training with no differences between EU and US physicians. This was followed by three other factors that received the same overall score (financial issues, doctor bedside time and quality of pre-dialysis care), but it is interesting to note that the Europeans attributed considerably greater importance to bedside time than their US counterparts. CONCLUSION: It seems that the reported difference in dialysis outcomes between Europe and the US is a widely accepted fact. Although directed towards few respondents, our questionnaire does suggest some differences in the approach towards dialysis and end-stage renal disease patients. PMID- 10534501 TI - Living non-related versus related renal transplantation--its relationship to the social status, age and gender of recipients and donors. AB - BACKGROUND: Persistent differences between social classes and genders exist in the quality of medical care due to disparities in need and access. METHODS: 149 haemodialysis (HD) patients including 114 renal transplant candidates, and their proposed live donors were interviewed and followed for 4 years. Differences in need and access were analysed among the living non-related compared to related renal transplant according to social status, age and gender of recipients and donors. Also the motive for organ-donation as well as the recipient's survival was compared between living non-related and related renal transplantation. RESULTS: The proportion of females among renal transplant candidates was significantly lower than among HD-patients. Females were significantly less likely to be recipients, but more likely to be donors of renal allografts, particularly if they were unemployed. Initially all of the living non-related donors claimed to have altruistic motives for organ-donation but gift rewarding, drug abuse, unemployment, and economical deadlock, urgent need of money were significantly frequent than among living related donors. The donation process lasted significantly longer in females and in living non-related donors and there was a trend for higher mortality in recipient of living non-related grafts. Almost all of the living non-related donors disappeared after organ-donation without subsequent follow-up. CONCLUSIONS: Females are transplanted less frequently, but donate kidneys more frequently than males in living non-related transplantation programmes. There is an excess of vulnerable people among living non-related donors. PMID- 10534502 TI - Erythropoietin resistance due to dialysate chloramine: the two-way traffic of solutes in haemodialysis. PMID- 10534503 TI - Secretion of chemokines and cytokines by human tubular epithelial cells in response to proteins. AB - BACKGROUND: Chronic interstitial scarring contributes to the progression of renal failure in glomerular disease but its cause is unknown. The development of proteinuria could stimulate tubular cells to release cytokines, chemoattractants and matrix proteins into the interstitium, thus contributing to interstitial disease. METHODS: Polarized human tubular epithelial cells were grown on permeable supports and exposed to serum proteins on their apical surface. The release of tumour necrosis factor alpha(TNFalpha), platelet derived growth factor (PDGF) and monocyte chemoattractant protein-1 (MCP-1) by the cells was measured using immunoassays. RESULTS: Under control conditions there was polarized release of PDGF-AB with predominant basolateral secretion (basolateral to apical ratio 4.7+/-1.6). MCP-1 release was less polarized (ratio 1. 7+/-0.5). TNFalpha was not detected. Exposure of the cells to normal human serum proteins on their apical side increased basolateral release of PDGF-AB (1.7+/-0.4 fold) and MCP-1 (2.4+/ 0.2 fold). Fractionation of the serum showed that this effect on human tubular epithelial cells was reproduced by a fraction of molecular weight 40-100 kDa. The predominant proteins in this fraction were albumin and transferrin but these purified proteins alone did not alter secretion of PDGF-AB or MCP-1. CONCLUSION: This data demonstrates that human tubular cells exposed to proteins, which would be filtered in glomerular disease, produce inflammatory mediators with the potential to stimulate inflammation and scarring in the interstitium of the kidney. PMID- 10534504 TI - Effects of L-arginine on cyclosporin-induced alterations of vascular NO/cGMP generation. AB - BACKGROUND: Cyclosporin (CsA)-induced vascular dysfunction has been attributed to a diminished role of the nitric oxide (NO)/cGMP-mediated vasodilator mechanism. The present study was aimed at investigating whether L-arginine, the substrate of NO synthesis, ameliorates CsA-induced vascular dysfunction. METHODS: Male Sprague Dawley rats were used throughout the study. The thoracic aorta was isolated from normal rats and acutely treated with CsA (10(-4) mol/l, 60 min) in vitro, or the aorta was taken from rats treated with CsA (25 mg/kg/day, i.m., 1 week). The vascular relaxation response to acetylcholine, and tissue levels of NO metabolites and cGMP were determined. The vascular expression of NO synthase (NOS) isoforms was also determined by western blot analysis. RESULTS: Acute treatment with CsA in vitro markedly attenuated the vasorelaxation response to acetylcholine, which was completely restored by L-arginine. The vascular accumulation of NO metabolites in response to acetylcholine was decreased significantly by CsA, which was prevented by cotreatment with L-arginine. CsA decreased the cGMP accumulation in response to both acetylcholine and sodium nitroprusside. L-Arginine restored, although not completely, acetylcholine stimulated cGMP generation, whereas it did not affect sodium nitroprusside stimulated cGMP generation. Following chronic CsA treatment in the whole animal, the vasorelaxation response to acetylcholine was decreased significantly along with tissue levels of NO metabolites; this was preserved by L-arginine supplementation. Vascular expression of iNOS protein was decreased by CsA treatment along with decreased tissue accumulation of NO metabolites. L-Arginine supplementation did not modify the altered expression of NOS proteins. CONCLUSION: These results suggest that CsA causes an L-arginine-sensitive vascular dysfunction which is associated with impaired generation of NO and cGMP. PMID- 10534505 TI - Hereditary renal amyloidosis associated with variant lysozyme in a large English family. AB - BACKGROUND: Two kindreds with hereditary systemic amyloidosis caused by the first two mutations to be described in the human lysozyme gene were discovered recently and study of the variant lysozyme has been powerfully informative about mechanisms of amyloid fibrillogenesis. However, the clinical manifestations in these families, additional members of which have lately been identified, have not previously been reported in detail. METHODS: The proband presented with proteinuria aged 50 and a family history of amyloidosis, and underwent renal biopsy, whole-body serum amyloid P component (SAP) scintigraphy, and sequencing of the lysozyme gene. Her family history and the phenotype of hereditary lysozyme amyloidosis were thoroughly documented and compared with the presentation and natural history of all other known patients with this condition. RESULTS: The proband belonged to an extended English family other members of which were known to have hereditary lysozyme amyloidosis. Those with amyloid in previous generations presented with renal involvement, frequently developed complications due to gastrointestinal amyloid, and died before age 60. All amyloid deposits were composed of lysozyme and complete concordance was established between amyloid and heterozygosity for a point mutation in the lysozyme gene, encoding the previously reported Asp67His substitution in the mature protein. CONCLUSION: The phenotype, reported for the first time in this extended kindred, contrasts with that of an apparently unrelated family carrying the same mutation who presented with spontaneous hepatic haemorrhage and rupture, and with the manifestations in a family with the lysozyme Ile56Thr variant who presented with dermal petechiae before proceeding to fatal visceral amyloidosis. A remarkably wide spectrum of disease can be caused by the same amyloid fibril protein, although renal involvement predominates in all cases except those dying of hepatic rupture. PMID- 10534506 TI - Antibodies to Tamm-Horsfall protein in endemic nephropathy. AB - BACKGROUND: The aim of the study was to investigate the possible role of antibodies to Tamm-Horsfall protein (anti-THP) in the early diagnosis of endemic nephropathy (EN). METHODS: Anti-THP (IgA, IgM, IgG classes) antibodies were determined by direct ELISA in a random sample of 159 subjects from the endemic village of Kaniza who were divided into four groups according to the WHO criteria, i.e., 'diseased', 'suspect', 'at risk', and 'others'. These groups were compared to subjects from the non-endemic village of Klakar and healthy subjects from Zagreb. RESULTS: No differences for all the classes of antibody were observed among the groups of subjects from the endemic village of Kaniza (P>0.05) or between these subjects and those from the non-endemic village of Klakar (P>0.05). The values of IgM anti-THP antibodies exceeded those of the IgA and IgG classes in all groups of subjects (P<0.05). The values for all three classes of antibodies were higher in the rural than the urban population (P<0. 05). CONCLUSION: Determination of anti-THP antibodies was not found to be useful in the early diagnosis of endemic nephropathy. The results suggest that most of the anti-THP antibodies are 'natural' and/or cross reactive. The highest values observed in the rural population could probably be explained by exposure to some ubiquitous antigen or more likely they are consequences of fever. PMID- 10534507 TI - Recurrent focal glomerulosclerosis: natural course and treatment with plasma exchange. AB - BACKGROUND: Focal glomerulosclerosis (FGS) can recur after renal transplantation and prognosis is poor in untreated patients. A circulating plasma factor has been implicated in the pathogenesis of a recurrent FGS and treatment with plasma exchange has proven effective in decreasing proteinuria in some patients. METHODS: We retrospectively studied the course of disease in patients with recurrent FGS, transplanted in our centre. Seven patients transplanted between 1991 and 1997, received treatment with plasma exchange, whereas 10 patients, transplanted between 1973 and 1991, were left untreated and served as historical controls. RESULTS: The time of onset of proteinuria (>3.5 g/day) was comparable in the untreated and treated patients (9 and 10 days respectively), as was the average proteinuria at that time (5.5 and 5.8 g/day respectively). In the untreated patients, proteinuria persisted and eventually all grafts were lost, on average 43 months after the diagnosis of a recurrence. In five cases (50%) the recurrence was the single cause of graft loss. The clinical course was different in the seven patients who were treated with plasma exchange. In five of these patients, the recurrence occurred within 3 weeks after transplantation. Plasma exchange was started 1-14 days after onset of proteinuria in these patients. Two lost their grafts after 0.7 and 1.0 months because of untreatable rejection. In the remaining three patients the plasma exchange resulted in abrupt disappearance of the proteinuria, and the response has been lasting for 2-3.2 years. In these patients the only histological abnormality was foot effacement on electron microscopy. In two patients the recurrence became manifest at 9 weeks and 5.8 years after transplantation respectively. These two patients relapsed after the initial course of plasma exchange, but responded to repeated session, and are currently being treated once a month. They have been followed for 1. 7 and 1.4 years after the onset of proteinuria and their urinary protein levels are 0.23 and 1.2 g/10 mmol creatinine. CONCLUSIONS: The prognosis of untreated recurrent FGS is poor. Treatment with plasma exchange can lead to complete remission of proteinuria and relapsing patients may respond to repeated sessions. Best results are obtained when plasma exchange is started early, when there are no visible lesions on light-microscopy. PMID- 10534508 TI - Functional correlates of alpha(2A)-adrenoceptor gene polymorphism in the HANE study. AB - BACKGROUND: The aim of this study was to test an association between alleles of the alpha(2A)-adrenoceptor gene with hereditary hypertension and with alterations of lipid and carbohydrate metabolism. METHODS: Genomic DNA was isolated from 147 hypertensive patients and digested with DraI. Genotypes at the alpha(2A) adrenoceptor were identified by restriction fragment length polymorphism. Genotype at each locus was related to blood pressure, family history of hypertension and various clinical chemistry parameters. RESULTS: The alpha(2A) adrenoceptor polymorphism was not significantly associated with blood pressure or a family history of hypertension. Patients with the d allele of the alpha(2A) adrenoceptor had significantly lower HbA(1) (5.6 vs 6.2%, P=0.0344) and HbA(1c) (3.4 vs 3.9%, P=0.0237) and total cholesterol (212 vs 229 mg/dl, P=0.0333) than those without. Similar trends, which failed to reach statistical significance, were seen for glucose, triglycerides and LDL cholesterol. CONCLUSIONS: We propose that alleles at the alpha(2A)-adrenoceptor locus might contribute to interindividual differences in the regulation of human lipid and glucose metabolism. PMID- 10534509 TI - Disturbed LDL and scavenger receptor functions in monocytes from chronic haemodialysed patients. AB - BACKGROUND: The most frequent complication in patients with end-stage renal failure on chronic haemodialysis (HD) treatment is atherosclerosis, i.e. the different forms of heart and vascular diseases. The complete disorder of serum lipid and lipoprotein patterns is well demonstrated, whereas our knowledge about the low-density lipoprotein (LDL) and scavenger receptor expression and function are poorly understood. METHODS: In our current work, LDL and scavenger receptor expression and functions were simultaneously studied in monocytes obtained from 15 healthy male control subjects and from 11 chronic HD male patients applied with (125)I-labelled LDL, isolated from healthy volunteers. To study the scavenger LDL receptors, labelled acetylated LDL (acLDL) was used. RESULTS: LDL binding to the monocytes of the HD-group was found to be decreased in comparison to that of the controls. As a result, the 50 microg LDL protein-induced inhibition of endogenous cholesterol synthesis was also diminished. In contrast, acLDL binding was greatly increased, though it could trigger only a low apoE synthesis. Consequently the number of cholesterol inclusions in monocytes was increased. CONCLUSIONS: The disturbed expression and function of LDL and scavenger receptors both may play significant roles in pathogenesis of cardiovascular complications in chronic HD patients. Based on our present results, it can be assumed that dysfunction of scavenger receptors is at the centre of cardiovascular complications of HD patients with renal failure. PMID- 10534510 TI - Significance of serum pepsinogens and their relationship to Helicobacter pylori infection and histological gastritis in dialysis patients. AB - BACKGROUND: Previous investigations reported that patients undergoing dialysis therapy had significantly higher serum pepsinogen (PG) levels than patients with normal renal function. However, in dialysis patients, the relationship between serum PG levels and Helicobacter pylori infection remains unknown. METHODS: Sixty three maintenance dialysis patients (54 haemodialysis and nine continuous ambulatory peritoneal dialysis) who required endoscopic examination were enrolled in the study. Sixty four age- and sex-matched patients with normal renal function served as controls. We performed endoscopic examination and obtained both the gastric antral and corpus mucosa for histopathological evaluation and H. pylori identification. Twenty three patients on dialysis underwent H. pylori eradication therapy. RESULTS: In dialysis patients, H. pylori-positives had significantly higher serum PG II levels than H. pylori-negatives (26.6+/-21.5 vs 14.1+/-7.1 ng/ml, P<0.05), but no significant difference was found in serum PG I between H. pylori-positives and H. pylori-negatives (228.8+/-158.5 vs 179. 4+/-113.5 ng/ml). There was no significant difference in serum PG II between dialysis patients and controls (19.9+/-16.5 vs 18.6+/-14.9 ng/ml), while serum PG I levels were significantly higher in dialysis patients than in controls (201.7+/-136.8 vs 77.6+/-85.8 ng/ml, P<0.05). Serum PG II levels, but not those of PG I, significantly correlated with the inflammation and activity scores of antrum in dialysis patients, and these scores were highly influenced by H. pylori infection. Dialysis patients in whom H. pylori was eradicated successfully showed significant reductions of serum PG II levels but not of PG I. CONCLUSIONS: In dialysis patients, high serum levels of PG II, but not PG I, are significantly related to H. pylori infection and mucosal inflammation. A significant decrease in serum PG II levels could be used as a predictor of the eradication of H. pylori infection in dialysis patients. PMID- 10534511 TI - Ambulatory blood pressure monitoring in patients receiving long, slow home haemodialysis. AB - BACKGROUND: Good blood pressure (BP) control has been reported previously in haemodialysis (HD) patients receiving 8-h dialysis sessions. Home HD allows patients to dialyze for long periods, but there are few data on the BP control achieved by these patients. We studied BP control, using ambulatory blood pressure monitoring (ABPM), in our home-HD patients who were receiving long-hours dialysis. METHODS: Twenty-four patients aged 52.7+/-11 years underwent ABPM. They had been on home HD for 52.9+/-39 months and dialysed for 7.2+/-1.1 h thrice weekly. Two patients were taking antihypertensive drugs. Historical data on BP and weight gains were obtained from the patients' own records. Left ventricular (LV) mass was assessed by echocardiography and total body water (TBW) by bioelectrical impedance. RESULTS: The mean 24-h BP was 129+/-17 mmHg (systolic) and 83+/-14 mmHg (diastolic). The daytime BP was 131+/-17 mmHg (systolic) and 84+/-14 mmHg (diastolic), while the night-time BP was 126+/-22 mmHg (systolic) and 81+/-17 mmHg (diastolic). Six patients (25%) had a normal circadian BP rhythm, but the rest showed a subnormal fall or an increase in BP at night. Mean 24-h BP did not correlate significantly with time on dialysis, dialysis session length, Kt/V, haemoglobin, interdialytic weight gain, or TBW. Twenty-one patients (87%) had LV hypertrophy and 16 of these had diastolic dysfunction. LV mass index was inversely correlated with nocturnal BP fall (r=-0.54, P=0.03). Non-dippers had been treated longer than dippers (29 vs 59.2 months, P=0.03) but they were similar in respect to age, dialysis session length or Hb concentration. CONCLUSIONS: Long, slow haemodialysis at home provides satisfactory daytime BP control in the majority of patients without the need for antihypertensive drugs but abnormal circadian BP rhythm and LV hypertrophy remain common. PMID- 10534512 TI - Enhanced oxidative stress in haemodialysis patients receiving intravenous iron therapy. AB - BACKGROUND: Iron balance is critical for adequate erythropoiesis and there remains much debate concerning the optimal timing and dosage of iron therapy for haemodialysis patients receiving recombinant human erythropoietin therapy. METHODS: In this study, we examined the influence of baseline ferritin level and intravenous infusion of 100 mg ferric saccharate on the oxidative status of the patients on maintenance haemodialysis. The levels of antioxidant enzymes and lipid peroxides were determined in erythrocytes and plasma of 50 uraemic patients on haemodialysis. These patients were divided into groups 1, 2, and 3, based on their baseline serum ferritin levels of <300, 301-600, and >601 microg/l, respectively. RESULTS: We found that the mean superoxide dismutase (SOD) activities in the erythrocytes were similar in the three groups of patients and did not differ from those of the age-matched controls. On the other hand, all the haemodialysis patients showed significantly higher plasma SOD activity as compared to controls. After intravenous iron infusion, group 3 patients showed the largest decrease in plasma SOD activity. The plasma glutathione peroxidase (GSHPx) activities of the patients in all three groups and the erythrocyte GSHPx activities of the patients in the groups 2 and 3 were lower than those of the healthy controls. In all three groups of patients, no difference in GSHPx activity was found before and after intravenous iron infusion. On the other hand, we found that the average baseline levels of plasma lipid peroxides of all three groups of patients were significantly higher than that of the controls. The patients in group 3 with the highest serum ferritin levels showed the highest levels of plasma lipid peroxides. More importantly, we found that after iron infusion, the patients in all three groups, particularly those in group 3, showed significantly elevated levels of plasma lipid peroxides. CONCLUSION: We demonstrated that increased oxidative stress in the blood circulation of the uraemic patients on haemodialysis is exacerbated by the elevated baseline serum ferritin levels and intravenous iron infusion. The resultant oxidative damage may contribute to the increased incidence of atherosclerosis in the patients with end stage renal disease on long-term haemodialysis. PMID- 10534513 TI - Impact of Helicobacter pylori infection on serum gastrin in haemodialysis patients. AB - BACKGROUND: Helicobacter pylori infection is associated with increased gastrin release in patients with normal renal function. Hypergastrinaemia is a common finding in haemodialysis patients and, in many cases, may be linked to H. pylori infection. The aim of this study was to examine the effect of H. pylori infection, and its eradication, on elevated gastrin levels in haemodialysis patients. METHODS: Eighty-nine dyspeptic patients were included in the study. While 44 patients had normal renal function, the remaining 45 were end-stage renal failure patients. Patients were assigned to one of four groups according to their H. pylori and renal function status. Infected patients were re-evaluated after 2 months following eradication treatment. Serum gastrin levels were measured in these groups both before and after eradication treatment. RESULTS: Haemodialysis patients with H. pylori infection had higher serum gastrin levels than did H. pylori negative haemodialysis patients (321+/-131 pg/ml vs 154+/-25 pg/ml) (P<0.05). Mean serum gastrin concentration was 152+/-21 pg/ml in the non uraemic H. pylori-positive group. This value was 58+/-17 pg/ml in the non-uraemic H. pylori-negative group (P<0.05). There were significant decreases in serum gastrin levels from pre- to post-eradication of H. pylori in the infected haemodialysis and non-uraemic patient groups (312+/-131 pg/ml to 179+/-85 pg/ml and 152+/-21 pg/ml to 72+/-2.4 pg/ml respectively, P<0.05). Four patients in group Ib and 5 patients in group IIb who had persistent infection did not have a decrease in serum gastrin level. All patients with successful eradication had a decrease in serum gastrin concentration. CONCLUSION: Our findings suggest that H. pylori infection contributes to hypergastrinaemia in haemodialysis patients. More research is needed regarding the clinical consequences of hypergastrinaemia in these individuals. PMID- 10534514 TI - Prevalence of present and past hepatitis G virus infection in a French haemodialysis centre. AB - BACKGROUND: Previous studies, detecting GB virus-C (GBV-C) or hepatitis G virus (HGV) RNA by using reverse transcriptase polymerase chain reaction (RT-PCR), have shown that haemodialysis (HD) patients had a high risk of being infected and viraemic with this virus. A past GBV-C/HGV contact can now be detected by testing for antibodies directed against the GBV-C/HGV envelope protein E2 (anti-E2). METHODS: In order to evaluate GBV-C/HGV contact, 120 patients undergoing chronic HD were tested for GBV-C/HGV RNA by RT-PCR and anti-E2 antibodies by ELISA. GBV C/HGV viraemic patients were followed prospectively for 18 months, and retrospectively when sera were stored. The total follow-up was between 18 and 78 months. RESULTS: GBV-C/HGV RNA was detected in 17 patients (14%), and 18 patients (15%) had a significant level of anti-E2 antibodies. No positive anti-E2 specimens were also positive for GBV-C/HGV RNA and vice versa. A total of 35 patients (29%) were contaminated with GBV-C/HGV. Sixteen of the 17 viraemic patients had a persistent viraemia (follow-up 18-78 months) and one cleared the virus during the study period. A past or present GBV-C/HGV contact was statistically correlated with the duration of HD and hepatitis C virus (HCV) infection, but was independent of age, hepatitis B virus (HBV) infection, and alanine aminotransferase (ALT) level. CONCLUSIONS: Twenty-nine per cent of patients who underwent HD in our centre have been infected by GBV-C/HGV, 49% were still viraemic and 51% have developed anti-E2 antibodies, indicating a past contact with GBV-C/HGV. Our results demonstrate that the prevalence of GBV-C/HGV contact in HD was underestimated when only RT-PCR was used. Therefore GBV-C/HGV contact is probably much more frequent in HD than previous studies would suggest and is at this time not correlated with hepatotoxicity. Anti-HCV antibodies blood screening since 1990 and recent changes in managing HD patients have probably reduced GBV-C/HGV contact in the same way. PMID- 10534515 TI - Comparison of low-molecular-weight heparin (enoxaparin sodium) and standard unfractionated heparin for haemodialysis anticoagulation. AB - BACKGROUND: Low-molecular-weight heparin (LMWH) has been suggested as providing safe, efficient, convenient and possibly more cost-effective anticoagulation for haemodialysis (HD) than unfractionated heparin, with fewer side-effects and possible benefits on uraemic dyslipidaemia. METHODS: In this prospective, randomized, cross-over study we compared the safety, clinical efficacy and cost effectiveness of Clexane (enoxaparin sodium; Rhone-Poulenc Rorer) with unfractionated heparin in 36 chronic HD patients. They were randomly assigned to either Clexane (1 mg/kg body weight, equivalent to 100 IU) or standard heparin, and followed prospectively for 12 weeks (36 dialyses) before crossing over to the alternate therapy for a further 12 weeks. Heparin anticoagulation was monitored using activated coagulation times. RESULTS: Dialysis with Clexane resulted in less frequent minor fibrin/clot formation in the dialyser and lines than with heparin (P<0.001), but was accompanied by increased frequency of minor haemorrhage between dialyses (P<0.001). Clexane dose reduction (to a mean of 0.69 mg/kg) eliminated excess minor haemorrhage without increasing clotting frequencies. Mean vascular compression times were similar in both groups. Over 24 weeks, no changes in standard serum lipid profiles were observed. CONCLUSIONS: This study suggests that a single-dose protocol of Clexane is an effective and very convenient alternative to sodium heparin, but currently direct costs are about 16% more. We recommend an initial dose of 0.70 mg/kg. PMID- 10534516 TI - Efficacy and tolerance of interferon-alpha(2b) in the treatment of chronic hepatitis C virus infection in haemodialysis patients. Pre- and post-renal transplantation assessment. AB - BACKGROUND: Hepatitis C virus (HCV) infection represents an important problem for the dialysis population due to its high prevalence and the long-term development of chronic liver disease, particularly following renal transplantation. METHODS: In order to assess the efficacy and tolerance of interferon (IFN) in the treatment of chronic hepatitis C in haemodialysis (HD) patients and their clinical course following renal transplantation, a multicentre, randomized, open label study was conducted to compare IFN therapy vs a control group. RESULTS: Nineteen HCV RNA-positive patients received 3 x 10(6) U of IFN s.c., three times a week (post-HD), and 17 HCV RNA-positive patients were assigned to the control group. Tolerance to IFN therapy was good in nine patients, while treatment was discontinued in the other 10 due to the occurrence of side effects. HCV RNA was negative at the end of treatment in 14 out of 19 patients (74%) receiving IFN and in one patient (5%) in the control group. Six out of the 14 patients who initially responded to IFN therapy had a virological relapse (43%). Eight patients (42%) remained HCV RNA-negative, three of them until the day that renal transplantation (RT) was performed (7, 12 and 27 months, respectively), as did five patients on HD during the follow-up (27+/-5 months). Eight out of the nine patients (89%) who completed therapy were HCV RNA-negative at the end of treatment, and seven of them (78%) remained HCV RNA-negative during the follow-up on dialysis (21+/-8 months). Mean transaminase (ALT) values were significantly decreased following IFN therapy, while no changes were observed during the follow up period in the control group. Fifteen patients (10 in the treatment group and five in the control group) underwent RT. Three patients in the treatment group were HCV RNA-negative at RT, and one of them had a virological relapse 20 months after RT, while the other two remained HCV RNA-negative at 3 months and 24 months after RT, respectively. In contrast to the control group, transaminase (ALT) remained within normal limits in all patients in the treatment group. Finally, during the post-RT follow-up, the transaminase mean values were significantly lower in treated patients vs patients in the control group (P<0.05). CONCLUSIONS: It is concluded that the biochemical and virological response to IFN therapy is good in HD patients. In addition, IFN therapy appears to exert a beneficial effect on the course of liver disease following RT, regardless of the virological response. Despite the fact that IFN therapy was discontinued in 10 out of the 19 patients due to the occurrence of side effects, these disappeared following discontinuation of therapy. Therefore, IFN therapy is advisable for HCV-infected dialysis patients who are candidates for RT. PMID- 10534517 TI - Effect of mycophenolate mofetil on erythropoiesis in stable renal transplant patients is correlated with mycophenolic acid trough levels. AB - BACKGROUND: Both mycophenolate mofetil (MMF) and azathioprine (AZA) are immunosuppressive drugs that inhibit purine synthesis. In theory, MMF selectively inhibits lymphocyte proliferation, while AZA has well-known effects on red blood cells and thrombocytes as well. In renal transplant recipients we replaced CsA therapy by MMF in an attempt to reduce the immunosuppressive load 1 year after kidney transplantation. During this study we observed the effect of MMF on haematological parameters such as haemoglobin (Hb), leukocytes, and thrombocytes. METHODS: One year after kidney transplantation 26 stable patients were converted from cyclosporin A (CsA) to MMF (2 g/day). Thereafter, these patients were tapered twice in their MMF dose from 2 g to 1.5 g (4 months after conversion) and from 1.5 to 1 g (8 months after conversion) per day. The Hb levels, leukocyte and thrombocyte counts, and mycophenolic acid (MPA) trough levels were routinely measured. RESULTS: After conversion from CsA to MMF not only creatinine levels and the number of leukocytes, but also the haemoglobin (Hb) level significantly decreased in 21/26 patients (P=0.0004). In eight patients the Hb level dropped more than 1 mmol/l (=1.61 g/dl). Only in two of eight patients was an explanation for blood loss found. The effect on Hb level did not ameliorate after the first MMF dose reduction to 1.5 g/day. After tapering the MMF dose to 1 g/day, the Hb approached the pre-conversion level. Not only the MMF dose but also the mycophenolic acid (MPA) trough level correlated with the Hb level. CONCLUSIONS: After conversion from CsA to MMF 1 year after kidney transplantation, a decrease in Hb level and leukocyte count was observed. The MPA trough level correlated also with the Hb level. The effect on the Hb level was reversible after dose reduction. This finding suggests that MMF exerts a negative effect on erythropoietic cells. PMID- 10534518 TI - Sheehan syndrome presenting as early post-partum hyponatraemia. PMID- 10534519 TI - Progressive tumoral calcinosis as the presenting feature of sarcoidosis in a patient on haemodialysis treatment. PMID- 10534520 TI - Renal zygomycosis: an under-diagnosed cause of acute renal failure. AB - BACKGROUND: Invasive zygomycosis (mucormycosis) occurs predominantly in immunocompromised patients in whom it carries a grave prognosis. While renal involvement is not so uncommon in disseminated infection, isolated renal zygomycosis is rare. METHODS AND RESULTS: Forty-five patients with systemic zygomycosis were seen over a 12-year period from January 1986 to December 1997. Among these, 18 had renal involvement, nine with disseminated disease and nine with isolated renal zygomycosis. No underlying predisposing disease was identified in the majority of patients (72%). Renal involvement was confirmed at autopsy in 13 and by ante-mortem renal biopsy in five patients. The infection involved one kidney in five patients and was bilateral in the remaining. The manifestations included fever, flank pain, haematuria and pyuria with evidence of enlarged non-functioning kidneys on computerised tomography (CT). Of those with bilateral disease, 12 (92.3%) had anuric acute renal failure. Anti-fungal therapy was given to six patients (amphotericin B in mean total dose of 1.1 g) and of these only two with unilateral disease who also underwent nephrectomy survived while all the other 16 died. CONCLUSION: This study shows that renal zygomycosis has emerged as a cause of acute renal failure in the last decade since no patient with renal involvement was identified at our centre prior to 1986 even though autopsies have been done regularly in patients dying of unknown causes. Bilateral renal zygomycosis should be suspected in any patient who presents with haematuria, flank pain and otherwise unexplained anuric renal failure. Characteristic CT findings and an early renal biopsy can confirm the diagnosis and help in effective management of this serious disease. PMID- 10534521 TI - Endovascular treatment of arteriovenous fistulas complicating percutaneous renal biopsy in three paediatric cases. AB - DESIGN: We evaluated the incidence and history of arteriovenous fistula (AVF) after kidney biopsy and assessed the use of superselective embolization for treatment. OBSERVATIONS: During the last 10 years, 896 kidney biopsies (age range of the patients: 1 month-18.6 years) have been performed in our institution under real-time ultrasonographic guidance with a 14 gauge cutting biopsy needle, and 32 of the patients had renal allografts (3.4%). We observed three cases of AVF (two in allograft kidneys, one in a native kidney) among all biopsies (0.34%), and the incidence of developing AVF after renal allograft biopsy was 6.3%. All three patients with AVF were symptomatic, and intravascular therapy was indicated. INTERVENTIONS: An angiographic study combined with endovascular treatment of the intrarenal AVF and pseudoaneurysm was performed in all three patients. Embolization was performed with bucrylate and lipiodol in two patients and with micro-coils in one. After successful embolization, all three patients became asymptomatic (in two renal bleeding stopped, in one patient with severe uncontrollable hypertension blood pressure returned to normal limits). No complications were observed secondary to the embolization procedure. CONCLUSION: The technique of superselective embolization using a coaxial catheter is an effective and safe method in the treatment of post-biopsy AVFs and pseudoaneurysm. PMID- 10534522 TI - Mobilization of lead from bone in end-stage renal failure patients with secondary hyperparathyroidism. AB - BACKGROUND: It is now recognized that long-term exposure to even low levels of lead may increase bone lead content. Lead can then be released in toxicologically significant amounts during critical states of increased bone turnover. METHODS: Two patients with end-stage renal failure, one on haemodialysis and the other on continuous ambulatory peritoneal dialysis (CAPD), had been exposed to lead and developed secondary hyperparathyroidism. An edetate calcium disodium (EDTA) test was performed in combination with haemofiltration or CAPD before and after parathyroidectomy. RESULTS: Before parathyroidectomy, both patients had low delta aminolaevulinic acid dehydrase (ALA-D) and high concentrations of chelated lead. After parathyroidectomy, there was a dramatic decrease in chelated lead and the ALA-D returned to normal. CONCLUSION: Secondary hyperparathyroidism increases mobilization of bone lead in dialysis patients with an elevated lead burden. This may cause toxic effects. PMID- 10534523 TI - A young woman with high blood pressure on haemodialysis: it is never too late to evaluate hypertension. PMID- 10534524 TI - Images in nephrology. Recurrence of psoriasis in an arteriovenous fistula. PMID- 10534525 TI - Horseshoe kidney. PMID- 10534526 TI - Liver transplantation for dialysis dependent hepatorenal failure. PMID- 10534527 TI - Common errors in diagnosis and management of urinary tract infection. I: pathophysiology and diagnostic techniques. PMID- 10534528 TI - Common errors in diagnosis and management of urinary tract infection. II: clinical management. PMID- 10534529 TI - No 'Stinkefinger' and renal failure--do you see a link? Diagnosis: acrorenal syndrome. PMID- 10534530 TI - Gordon Murray and the artificial kidney in Canada. PMID- 10534531 TI - Bone markers in the diagnosis of low turnover osteodystrophy in haemodialysis patients. PMID- 10534532 TI - Tubulointerstitial nephritis with anti-neutrophil cytoplasmic antibody following indomethacin treatment. PMID- 10534533 TI - Unilateral form of polycystic kidney disease. PMID- 10534534 TI - Dihydropyridine calcium channel blockers: a rare and reversible cause of hepatotoxicity with cholestasis in a CAPD patient. PMID- 10534535 TI - Vascular steal syndrome. PMID- 10534536 TI - Acute renal failure from intoxication by Cortinarius orellanus: recovery using anti-oxidant therapy and steroids. PMID- 10534537 TI - IgA myeloma: a potential outcome of IgA nephropathy. PMID- 10534538 TI - Pseudoproteinuria following gelatin infusion. PMID- 10534540 TI - Management of crystal arthritis. PMID- 10534539 TI - Targeted therapies in rheumatoid arthritis: the need for action. PMID- 10534541 TI - A review of gastrointestinal manifestations of systemic lupus erythematosus. AB - In this review, we analyse critically the effects of systemic lupus erythematosus (SLE) on the gastrointestinal (GI) tract from mouth to anus, attempting to distinguish the features that are most likely to be due to therapy. GI manifestations of SLE include mouth ulcers, dysphagia, anorexia, nausea, vomiting, haemorrhage and abdominal pain. GI vasculitis is usually accompanied by evidence of active disease in other organs. Early recognition of the significance of these symptoms offers the best opportunity to improve the symptoms and to aid long-term survival. PMID- 10534542 TI - Complement activation in patients with systemic lupus erythematosus without nephritis. AB - OBJECTIVE: To study the association between disease activity and complement activation prospectively in patients with systemic lupus erythematosus (SLE). PATIENTS AND METHODS: Twenty-one SLE patients were examined monthly for 1 yr. Disease activity, autoantibodies, conventional complement tests and the following complement activation products were investigated: C1rs-C1inh complexes, C4bc, Bb, C3a, C3bc, C5a and the terminal SC5b-9 complement complex (TCC). RESULTS: Modest variation in disease activity was noted. None of the patients had nephritis. Flare was observed at 27 visits. Four patients had anti-C1q antibodies in conjunction with modestly low C1q concentrations. The complement parameters were rather constant during the observation period. Slightly to moderately decreased C4 (0.05-0.10 g/l) was found in 10 patients and severely decreased C4 (<0.05 g/l) in seven patients. Decreased C4 was not associated with increased complement activation. Complement activation products were either normal or slightly elevated. TCC was the only activation product correlating significantly with score for disease activity at flare. None of the variables tested predicted flares. CONCLUSION: Complement tests are of limited importance in routine examination of SLE without nephritis, although TCC is suggested to be one of the most sensitive markers for disease activity. PMID- 10534543 TI - Reliability and sensitivity to change of a simplification of the Sharp/van der Heijde radiological assessment in rheumatoid arthritis. AB - OBJECTIVE: To determine the reliability and sensitivity to change of a simplified radiological scoring method [simple erosion narrowing score (SENS)] for rheumatoid arthritis (RA). SENS was compared to the Sharp/van der Heijde score (SHS) as a gold standard. METHODS: Sets of seven radiographs of hands and feet were taken of 20 RA patients with a wide spectrum of radiological damage. For 14 patients, these seven radiographs were taken during a follow-up period of 5 yr, and for six patients during a follow-up of 10 yr. Each set of radiographs was scored twice by the same observer (DvdH). Erosions and joint space narrowing were scored with SHS (range 0-448) in 32 and 30 joints in the hands, respectively, and both in 12 joints in the feet. SENS gives a score of 1 if there is any erosion in a joint and also 1 if there is any narrowing in the joint (range 0-86). In each case, SENS was derived from SHS. To analyse data, generalizability theory and repeated measurements ANOVA were used. RESULTS: The overall reliability coefficient was 0.81 for SHS and 0.80 for SENS. Intra-observer reliability [intraclass correlation coefficient (ICC)] was 0.99 and 0.98 for SHS and SENS, respectively. The ICC for the sensitivity to change was 0.84 for SHS and 0.88 for SENS. The smallest detectable difference (SDD) could be determined for both methods. The presence of progression based on this SDD was very comparable between the two methods. CONCLUSION: The measurement properties of SENS are good and comparable to SHS. This makes SENS suitable for use in clinical practice and in large (epidemiological) studies, especially in the first years of disease. PMID- 10534544 TI - Changes in plasma free fatty acid concentrations in rheumatoid arthritis patients during fasting and their effects upon T-lymphocyte proliferation. AB - OBJECTIVE: To measure whether changes in the concentrations of circulating free fatty acids (FFAs) after a 7 day fast in rheumatoid arthritis (RA) patients would inhibit in vitro T-lymphocyte proliferation. METHODS: The concentration and composition of plasma FFAs were measured in nine RA patients at the conclusion of a 7 day fast. A FFA mixture was made up based on these findings (20% linoleic, 43% oleic, 10% stearic, 27% palmitic acid). Mitogen-induced lymphocyte proliferative responses were measured after co-culture of peripheral blood mononuclear cells (PBMC) from healthy individuals in the presence of increasing concentrations of this FFA mixture (from 0 to 2000 microM) and in the presence of FFA mixtures where the relative proportions of fatty acids varied. RESULTS: Both the concentration of the FFA mixture and the ratio between the unsaturated and saturated fatty acids significantly influenced in vitro lymphocyte proliferation (P<0.0001). Unexpectedly, the highest concentrations of the FFA mixture increased lymphocyte proliferation. At equimolar concentrations (600 microM), manipulating the amounts of oleic and linoleic fatty acids relative to stearic and palmitic fatty acids had a potent inhibitory effect upon lymphocyte proliferation. CONCLUSION: Fasting-associated increases in total plasma FFA concentrations do not inhibit, but rather enhance, in vitro lymphocyte proliferation. An inhibitory effect could only be achieved by manipulating the balance between the unsaturated and saturated fatty acids. PMID- 10534545 TI - Systemic lupus erythematosus: clinical features in patients with a disease duration of over 10 years, first evaluation. AB - OBJECTIVE: Most information available about the disease course of patients with systemic lupus erythematosus (SLE) is restricted to the first 5 yr after disease onset. Data about the disease course 10 yr after disease onset are rare. The aim of this multicentre study was to describe the outcome of SLE patients with a disease duration of >10 yr. METHODS: Outcome parameters were the SLE Disease Activity Index (SLEDAI), the European Consensus Lupus Activity Measure (ECLAM), the Systemic Lupus International Collaborative Clinics/American College of Rheumatology Damage Index (SLICC/ACR), a global damage index (DI) and required treatment. In 10 different European rheumatology centres, all SLE patients who were evaluated in the last 3 months of 1994, and who had been diagnosed with SLE at least 10 yr ago, were included in the study. RESULTS: It should be stressed that our results are confined to a patient cohort, defined by a disease duration of at least 10 yr, and who are still under clinical care at the different centres in Europe. These SLE patients still showed some disease activity, related to symptoms of the skin and musculoskeletal systems, next to the presence of renal involvement. A total of 72% of the patients needed treatment with prednisolone (/=4 Beighton criteria) was 40.5% in girls and 12.9% in boys. Despite slight trends for hypermobile subjects to be less active in sports and to report more joint pain, no correlation could be found between hypermobility and musculoskeletal symptoms. CONCLUSIONS: An unusually marked sex difference in hypermobility exists among Icelandic 12-yr-olds, but hypermobility does not seem to affect joint symptoms or leisure activities at this age. PMID- 10534558 TI - Lung function in Adamantiades-Behcet disease. PMID- 10534560 TI - Rheumatoid arthritis in beta-thalassaemia trait. PMID- 10534559 TI - Surfactant treatment for osteoarthritis. PMID- 10534561 TI - Th1-type cytokine mRNA in rheumatoid arthritis mononuclear cells induced by streptococcal pyrogenic exotoxin A. PMID- 10534562 TI - Intravenous immunoglobulins for adult Still's disease and pregnancy. PMID- 10534563 TI - A search for Pneumocystis carinii DNA by polymerase chain reaction on bronchoalveolar lavage fluids from patients with Wegener's granulomatosis. PMID- 10534564 TI - Re: Tench et al. An insight into rheumatology resources available on the World Wide Web. PMID- 10534565 TI - An insight into rheumatology resources available on the World Wide Web (http://www.ilar.org) PMID- 10534566 TI - Melioidosis presenting as septic arthritis in Bengali men in east London. PMID- 10534567 TI - Heat shock protein 70 and ATP as partners in cell homeostasis (Review). AB - Heat shock proteins (HSP) are molecular chaperones, involved in many cellular functions such as protein folding, transport, maturation and degradation. Since they control the quality of newly synthesized proteins, HSP take part in cellular homeostasis. The Hsp70 family in particular exerts these functions in an adenosine triphosphate (ATP)-dependent manner. ATP is the main energy source used by cells to assume fundamental functions (respiration, proliferation, differentiation, apoptosis). Therefore, ATP levels have to be adapted to the requirements of the cells and ATP generation must constantly compensate ATP consumption. Nevertheless, under particular stress conditions, ATP levels decrease, threatening cell homeostasis and integrity. Cells have developed adaptive and protective mechanisms, among which Hsp70 synthesis and overexpression. In this review, we focus on the mechanisms which regulate Hsp70 expression under ATP depletion, using ischaemia as a paradigmatic model for ATP depletion in vivo, and analyze the molecular targets for Hsp70-mediated protection against ATP depletion. We also consider how these Hsp70-mediated protective effects could be applied in the therapeutical approaches of human diseases associated with cellular ATP depletion. PMID- 10534568 TI - Donor bone marrow and transplantation tolerance: historical perspectives, molecular mechanisms and future directions (review). AB - The induction of transplantation tolerance, defined as the indefinite existence of an intact, functioning allograft, has been a goal of transplantation immunologists for decades because of the morbid effects of immunosuppressive drugs required to maintain graft function. One promising method of tolerance induction involves the peritransplant administration of cytoablative drugs followed by administration of a low dose of donor bone marrow cells. A subpopulation of donor bone marrow cells bearing the CD8 accessory molecule cause the functional deletion of graft reactive T cells via a CD95 dependent mechanism that also involves the cytokine TGFbeta. This interaction is bi-directional in which the bone marrow cell deletes the graft reactive T cell that recognizes it. Therefore, only T cells that recognize the donor antigens are deleted, leaving the immune response to other antigens intact. This protocol has been successfully used in rodents and an outbred large animal model. Clinical trials of donor bone marrow administration are now underway and initially indicate that administration of donor bone marrow is clinically safe. PMID- 10534569 TI - Germline mutations in the MEN1 gene: creation of a new splice acceptor site and insertion of 7 intron nucleotides into the mRNA. AB - The MEN1 tumor predisposition syndrome is caused by mutations in the MEN1 gene on human chromosome 11q13. We screened MEN1 gene exons 1-10 and flanking intron sequences from four different MEN1 families for mutations. In three families, heterozygous germline mutations within the exons were found, two of these representing novel mutations. In another family, all clinically affected members were heterozygous for a point mutation Gright curved arrow A within intron 4. Sequence analysis of cDNA from lymphocytes of the affected patients revealed that the intron mutation created a new acceptor splice site, leading to the inclusion of 7 bp of intronic sequence into the mRNA. The resulting frameshift generates a premature stop in codon 271. Intron borders should thus be screened for mutations in MEN1 diagnostics and cDNA sequence analysis is helpful in identifying pathophysiological consequences of intron mutations. PMID- 10534571 TI - Cortical motor potential alterations in chronic fatigue syndrome. AB - Premovement, sensory, and cognitive brain potentials were recorded from patients with Chronic Fatigue Syndrome (CFS) in four tasks: i) target detection, ii) short term memory, iii) self-paced movement, and iv) expectancy and reaction time (CNV). Accuracy and reaction times (RTs) were recorded for tasks i, ii, and iv. Results from CFS patients were compared to a group of healthy normals. Reaction times were slower for CFS patients in target detection and significantly slower in the short-term memory task compared to normals. In target detection, the amplitude of a premovement readiness potential beginning several hundred milliseconds prior to stimulus onset was reduced in CFS, whereas the poststimulus sensory (N100) and cognitive brain potentials (P300) did not differ in amplitude or latency. In the memory task, a negative potential related to memory load was smaller in CFS than normals. The potentials to self-paced movements and to expectancy and RT (CNV) were not different between groups. The findings in CFS of slowed RTs and reduced premovement-related potentials suggest that central motor mechanisms accompanying motor response preparation were impaired in CFS for some tasks. In contrast, measures of neural processes related to both sensory encoding (N100) and to stimulus classification (P300) were normal in CFS. PMID- 10534570 TI - Proteolytic degradation of the retinoblastoma family protein p107: A putative cooperative role of calpain and proteasome. AB - p107 protein, a member of the retinoblastoma family protein, suppresses growth promotion in cancer cells. We have already reported evidence that calpain, a calcium dependent protease is involved in the cleavage of p107 protein. We show here that p107 protein can also be a substrate for ubiquitination. A negative growth regulator, the HMG-CoA reductase inhibitor lovastatin was found to induce loss of p107 protein which was reversible by a specific protease inhibitor lactacystin as well as calpain inhibitor. Following treatment with lovastatin higher molecular weight ubiquitinated forms of p107 were detected by anti-p107 immunoprecipitation and anti-ubiquitin Western blotting. These forms further increased when lactacystin was added to culture medium. These results indicate that ubiquitin-proteasome pathway plays a potential role in the degradation as well as calpain. The data presented here suggest a model in which calpain and ubiquitin-proteasome system possibly play a cooperative role in targeting the protein under certain conditions. PMID- 10534572 TI - Molecular effects of taxol and caffeine on pancreatic cancer cells. AB - Pancreatic cancer is the fifth leading cause of cancer related deaths in the United States. Despite many recent advances in the treatment modalities, the mortality rate still remains very high. Paclitaxel (Taxol) and Caffeine have been used for the treatment of this disease, however the molecular mechanisms of these agents are not fully understood, which may be partly responsible for the failure of these agents in the treatment of pancreatic cancer. Human pancreatic adenocarcinoma cell lines, HPAC and PANC-1 containing wild-type and mutant p53 respectively, were used to investigate the effects of Taxol and Caffeine on cell growth, and their effects on the modulation of cell cycle and apoptosis related genes. Protein extracts from these cells treated with 100 nM of Taxol or 4 mM of Caffeine were subjected to Western blot analysis for this study. Drug treated cells were also analyzed to calculate the number of cells undergoing apoptosis. Dose and time dependent growth inhibition was observed in both PANC-1 and HPAC cells when treated with either Taxol or Caffeine. Western blot analysis showed an up-regulation of p21WAF1 in both cell lines treated with either Taxol or Caffeine. Furthermore, down-regulation of cyclin B and cdk1 was observed in Taxol and Caffeine treated HPAC cells. However, the results were drastically different in PANC-1 cells where cyclin B was down regulated only by Caffeine treatment and the level of cdk1 protein was undetectable in this cell line. Moreover, up regulation of p53 and down-regulation of Bcl-2 was observed only in HPAC cells treated with Taxol. Apoptotic cell death analysis showed increasing number of cells undergoing apoptosis between 24 and 48 h of Caffeine treatment, however only Taxol showed greater than 50% cells under-going apoptosis only in HPAC cells. The up-regulation of p21WAF1 and down-regulation of cyclin B and cdk1 suggest their possible roles in G2/M cell cycle arrest caused by both Taxol and Caffeine as reported earlier. From these results we conclude that the differential molecular changes observed in this study may determine the cellular effects of these agents on pancreatic adenocarcinoma cells and that the effects of chemotherapeutic agents may be determined by the endogenous status of p53 mutation and, in turn, may determine the therapeutic effects of these agents in the treatment of pancreatic cancer. PMID- 10534573 TI - Sensitivity of shuttle vector plasmid pZ189 DNA to carbon ion beams. AB - The sensitivity of shuttle vector plasmid pZ189 DNA to carbon ion beams was studied under dry conditions. This plasmid contains the supF gene that can be used in mutagenesis analysis. The survival curve of the plasmid had both a shoulder and exponential parts, and showed a D10 value of about 17 kGy for both Escherichia coli wild-type and DNA repair-deficient mutant (UV-sensitive) strains. Therefore, it was assumed that the excision repair system was not effective for repairing DNA lesions induced by irradiation with carbon ion beams. PMID- 10534574 TI - Telomerase activity in the synovial tissues of chronic inflammatory and non inflammatory rheumatic diseases. AB - Telomerase is a ribonucleoprotein complex which can compensate for telomeric loss originating from each cell division, and its activation plays a critical role in cellular immortality. We previously found that telomerase is activated not only in immortal cancer cells but also in activated lymphocytes. To assess the diagnostic significance of telomerase activity in RA synovial tissues, we quantitatively examined telomerase activity in synovial tissue samples obtained from 47 patients with RA, 31 with osteoarthritis (OA), and 23 with other joint diseases. Telomerase activity in synovial tissues was detected in 28 of 47 (59.6%) patients with RA, including monoarticular-type RA, but in none of those with other joint diseases except one case each of synovial chondromatosis and OA. Thus, the specificity of telomerase activity in synovial tissues for RA among joint diseases was 96.3% (52/54). In RA samples, the telomerase activity was detected in 14 of 27 (51. 9%) patients with total joint replacement, 7 of 12 (58.3%) open synovectomy cases, and 7 of 8 (87.5%) arthroscopic synovectomy cases. Detection of telomerase activity in synovial tissues is considered to be useful for diagnosis of RA, including monoarticular-type RA, or active inflammation with lymphocyte infiltration, and arthroscopy can be applied for this purpose. PMID- 10534575 TI - Single cell analysis of T cells infiltrating labial salivary glands from patients with Sjogren's syndrome. AB - Sjogren's syndrome (SS) is an autoimmune disease characterized by lymphocytic infiltration into the lacrimal and salivary glands leading to symptomatic dry eyes and mouth. To analyze the function of T cells infiltrating the labial salivary glands, we analyzed T cell receptor (TCR) beta and alpha chains, the expression of various cytokine mRNAs, and apoptosis associated genes in predominant TCR BV2+ T cells in the labial salivary glands of patients with SS at the single cell level. TCR BV2+ T cells in the labial salivary glands were sorted as single cells by flow-cytometry, and then examined by a single cell polymerase chain reaction (PCR). We isolated 18 TCR BV2+ T cell clones from three patients with SS. In six clones, there were highly conserved amino acid motifs (RDxG, GNT, QGxxQETQ) in the CDR3 region of the TCR beta chain. Three of the six clones showed conserved amino acids (EDxTG, or ExxTG) in the CDR3 region of the TCR alpha chain, suggesting restricted T cell epitopes. All TCR BV2+ clones expressed IL-2 mRNA, and six clones were able to produce IL-4, indicating that the cells were Th0 type T cells. All TCR BV2+ clones in the labial salivary glands were CD4+ T cells, and ten clones overexpressed Fas antigen at the mRNA level. In contrast, only one clone expressed Fas-ligand (Fas-L) mRNA, and neither perforin nor granzyme A/B was expressed. In conclusion, these findings support the notion that TCR BV2+ T cells that infiltrate labial salivary glands recognize restricted epitopes and function as CD4+ Th0 type T cells in the induction phase of autoimmunity. PMID- 10534576 TI - Familial Alzheimer's disease: genetic influences on the disease process (Review). AB - Alzheimer's disease (AD) has both genetic and environmental etiologies. Genetic causes include presenilin (PS) mutations on chromosomes 1 and 14, and amyloid precursor protein (APP) mutations on chromosome 21. At least two susceptibility genes also exist. In this review phenotypic differences in AD groups are described and possible differences in the mechanism(s) by which AD mutations lead to dementia are reviewed. Clinical, pathological and biochemical phenotypes distinguish AD cases with different etiologies. For example, age-at-onset and age at-death between PS-1, PS-2, APP and sporadic AD groups differ. Also, some forms of AD are associated with more Abeta deposition others, and some AD groups have morphologically distinct Abeta deposits or other unique histopathologic features. APP-related AD mutations always occur within the Abeta portion of the APP gene, adjacent to sites where alpha-, beta- and gamma-secretase breakdown pathways operate in the expressed protein. These mutations alter APP metabolism leading to increased Abeta production. It is unknown if other AD groups are subject to identical changes in APP metabolism. Activation of apoptosis pathways, more general defects in protein transport or metabolism, differential regulation of tau kinases or other factors may also be important. Overall, data support the notion that differences occur in the disease process in etiologically distinct AD groups. PMID- 10534577 TI - Protease activity induced by nicotine in human cells. AB - Nicotine has a wide range of biological effects, and proteases have been extensively studied for their biological roles in living creatures. The aim of this study is to determine whether nicotine can induce proteolytic protease activity in cultures of various human cell lines. Plasminogen activator-like fibrinolytic protease activity, using 125I-fibrin as substrate in the presence of plasminogen, was estimated in cells with and without nicotine treatment. Among 16 cell lines tested, APr-1 cells were found to have the highest induced protease activity. Partial purification of the proteases was carried out by high performance liquid chromatography gel filtration on TSKG2000SW. Protease inhibitor tests indicated that the proteases induced by nicotine are serine proteases. PMID- 10534578 TI - Changes of E-cadherin and beta-catenin levels during induced differentiation of colorectal carcinoma cells. AB - Following the finding of a great increase in cell-cell adhesion in several colorectal carcinoma cell lines after induced differentiation, the expression of E-cadherin-catenin complexes was analyzed. The sensitivity of cell lines to the differentiation induced by sodium butyrate differed. Nevertheless, all cells growing for 5 days in the medium containing 2 mM sodium butyrate changed their morphology and adherent properties. The expression of E-cadherin and catenins participating in its function were analyzed. A significant increase in E-cadherin level after butyrate treatment was found in HT29 and LS174T cell lines only. However, a high decrease in beta-catenin level was detected in all cell lines treated with butyrate. Further analysis showed regulation of beta-catenin at the level of mRNA. PMID- 10534579 TI - Expression of the gadd153 gene in normal and tumor breast tissues by a sensitive RT-PCR method. AB - The gadd153 gene belongs to the CCAAT/enhancer binding protein (C/EBP) family of transcription factors. The role of these proteins in the control of proliferation and differentiation have mostly been studied in vitro. The involvement of gadd153 gene expression in human disease, and most particularly in breast cancer, is largely unknown. Since gadd153 gene is normally expressed at very low levels in most cells, we developed a sensitive reverse transcriptase-polymerase chain reaction (RT-PCR) technique that permits the detection of low amounts of RNA. In these conditions, 24 breast tumors and 14 corresponding normal samples were analysed, and levels of expression between tumor and normal tissues were compared. Statistical analysis indicated a significant induction of gadd153 gene expression in tumor samples in comparison to normal ones (p<0. 01). Thus, overexpression of gadd153 may inhibit the cellular differentiation process and facilitate breast tumorigenesis. Further studies are needed with larger number of cases to determine the specific prognostic role of gadd153 in breast cancer. PMID- 10534580 TI - DNA vaccines: basic mechanism and immune responses (Review). AB - DNA vaccines raise immune responses by expressing proteins in vaccinated hosts. Responses are raised by nanogram levels of protein expression. Popular methods of DNA delivery include intramuscular (i. m.) injections of DNA in saline and gene gun delivery of DNA-coated gold beads to the epidermis. Professional antigen presenting cells derived from the bone marrow present DNA-expressed antigens to T cells. Following gene gun immunizations directly transfected dendritic cells present antigens, whereas following i.m. immunizations both directly transfected dendritic cells and macrophages can present antigen. For both methods of DNA delivery, non-lymphoid cells can serve as factories of antigen for professional antigen presenting cells. Gene gun immunizations depend on antigen expression at the skin target whereas i.m. immunizations are largely independent of DNA expression in the muscle target. For both methods, antigen expression capable of initiating an immune response persists for about one month. intramuscular deliveries of DNA tend to raise type 1 T-cell help for intracellular and plasma membrane antigens but type 2 T-cell help for secreted antigens. Gene gun immunizations tend to raise type 2 T-cell help for both cell-associated and secreted antigens. In mice, DNA-raised immune responses can be equivalent to those raised by viral infections for both the height and longevity of antibody and cytotoxic T-cell responses. PMID- 10534581 TI - Inhibition of cGMP accumulation in mesangial cells by bradykinin and tyrosine kinase inhibitors. AB - We examined the effect of bradykinin (BK) on the accumulation of cGMP of the mesangial cell (MC), a smooth muscle-like cell of the renal glomerulus. BK caused a time- and concentration dependent reduction of the cGMP concentration. In addition, BK inhibited total protein tyrosine kinase (PTK) activity. Two tyrosine kinase inhibitors (TKI) genistein and tyrphostin also reduced the cGMP concentration. The inhibition of BK and TKI were not additive. The inhibition of PTK by BK, mediated through activation of the B2-receptor, was unaffected by inhibitors of Gi/o proteins, phospholipase C, protein kinase C, cyclooxygenase and Ca2+ release from intracellular stores. Only IBMX a broad spectrum inhibitor of phosphodiesterases (PDE) and 8-methoxymethyl IBMX a specific type-1 PDE inhibitor prevented the inhibitory effects of BK and TKI indicating the involvement of type-1 PDE. In addition, BK had no effect on soluble guanylate cyclase (sGC) and nitric oxide synthase activity. In freshly isolated glomeruli, which represent the physiological environment of MC, BK also reduced the cGMP concentration. Like in MC, the inhibitory effect was suppressed by IBMX. These data demonstrate that BK suppresses a PTK-dependent pathway of cGMP production in rat MC at a level downstream of NO synthase and sGC. It is suggested that BK and TKI inhibitors decrease cGMP levels by preventing tyrosine phosphorylation of type-1 PDE activity, thereby leading to enzyme activation. PMID- 10534582 TI - Lesions of the dorsolateral funiculus block supraspinal opioid delta receptor mediated antinociception in the rat. AB - Previous experiments have demonstrated that [D-Ala(2), Glu(4)]deltorphin (DELT) produces delta-receptor mediated antinociceptive effects when microinjected into the rat lateral ventricle and ventral medial medullary reticular formation (MRF), but not in the periaqueductal grey region (PAG). The present experiments were undertaken to further characterize the role of delta opioid agonists microinjected into the MRF and to explore the possibility of a descending pain modulatory system which might be linked to supraspinal delta opioid receptors. Rats received formalin into the dorsum of the right hindpaw and flinching responses were recorded. DELT given intracerebroventricularly (i.c.v.), intrathecally (i.th.) or into the MRF before formalin produced a dose-dependent and delta opioid receptor-mediated attenuation of both the first and second phases of the formalin-induced foot flinch response. DELT given i.c.v., i.th., or into the MRF also blocked formalin-induced increase in Fos-like immunoreactivity (FLI) in the dorsal horn of lumbar spinal cord ipsilateral to the formalin injection. Unilateral lesioning of the ipsilateral dorsolateral funiculus (DLF) did not alter nociceptive responses to formalin alone, but blocked the antinociceptive effect of DELT administered into the MRF; DELT was fully active in sham-DLF lesioned rats. Additionally, rats with DLF lesions did not show decreases in formalin-induced FLI in the ipsilateral lumbar spinal cord after injection of DELT into the MRF. These data suggest that delta opioid receptors in the MRF may be involved in activation of a descending inhibitory pain pathway projecting through the DLF to modulate tonic nociceptive input at the spinal level. PMID- 10534583 TI - Cold-evoked pain varies with skin type and cooling rate: a psychophysical study in humans. AB - The psychophysical responses to noxious cold stimulation of the skin in normal human subjects are not well understood. Continuous pain ratings with the visual analogue scale is an important method to assess these responses. In this study, we addressed several important issues about the parameters with which stimuli are delivered: the type of skin stimulated, the rate with which the stimulus temperature decreases, and the dimension of the pain rated by subjects. Cold stimuli were delivered to the thenar eminence (glabrous skin) and the dorso lateral hand (hairy skin) via a 4 cm(2) Peltier-type stimulator. Cold and pain thresholds were determined by the method of limits (MOL). A computerized visual analogue scale (VAS) was used to obtain continuous ratings of pain intensity and affect. The McGill Pain Questionnaire (MPQ) was used to assess the quality of cold-evoked pain. Supra-threshold stimuli (34 degrees C base) were delivered at 0.5, 1 or 2 degrees C/s to 2 degrees C, held for 20s and returned to baseline at 9 degrees C/s. These studies revealed: (1) Cold thresholds, measured with MOL, were lower (i.e. occurred at higher absolute temperatures) for the hairy skin of the dorso-lateral hand compared to the glabrous skin of the thenar eminence. (2) A similar pattern was evident for cold induced pain thresholds with MOL at 1.5 degrees C/s and with intensity and affect VAS scales at 0.5 and 1 degrees C/s. (3) Exponents for supra-threshold ratings fit to power functions were larger for the glabrous skin site than the hairy skin site regardless of cooling rate or dimension of pain measured. (4) All pain indices were higher for slower cooling rates. (5) No significant differences were found in the pain indices for pain ratings of intensity and affect. (6) A substantial proportion of subjects chose words representing paradoxical heat with the MPQ. (7) Painful paradoxical heat sensations occurred most often during cooling, while innocuous warm sensations mainly occurred during the rewarming phase. PMID- 10534584 TI - Effects of noradrenergic and serotonergic antidepressants on chronic low back pain intensity. AB - To understand the relative efficacy of noradrenergic and serotonergic antidepressants as analgesics in chronic back pain without depression, we conducted a randomized, double-blind, placebo-control head-to-head comparison of maprotiline (a norepinephrine reuptake blocker) and paroxetine (a serotonin reuptake blocker) in 103 patients with chronic low back pain. Of these 74 completed the trial; of the 29 who did not complete, 19 were withdrawn because of adverse effects. The intervention consisted of an 8-week course of maprotiline (up to 150 mg daily) or paroxetine (up to 30 mg daily) or an active placebo, diphenhydramine hydrochloride (up to 37.5 mg daily). Patients were excluded for current major depression. Reduction in pain intensity (Descriptor Differential Scale scores) was significantly greater for study completers randomized to maprotiline compared to placebo (P=0.023), and to paroxetine (P=0.013), with a reduction of pain by 45% compared to 27% on placebo and 26% on paroxetine. These results suggest that at standard dosages noradrenergic agents may provide more effective analgesia in back pain than do selective serotonergic reuptake inhibitors. PMID- 10534585 TI - An analysis of factors that contribute to the magnitude of placebo analgesia in an experimental paradigm. AB - Placebo analgesia was produced by conditioning trials wherein heat induced experimental pain was surreptitiously reduced in order to test psychological factors of expectancy and desire for pain reduction as possible mediators of placebo analgesia. The magnitudes of placebo effects were assessed after these conditioning trials and during trials wherein stimulus intensities were reestablished to original baseline levels. In addition, analyses were made of the influence of these psychological factors on concurrently assessed pain and remembered pain intensities. Statistically reliable placebo effects on sensory and affective measures of pain were graded according to the extent of surreptitious lowering of stimulus strength during the manipulation trials, consistent with conditioning. However, all of these effects were strongly associated with expectancy but not desire for relief. These results show that although conditioning may be sufficient for placebo analgesia, it is likely to be mediated by expectancy. The results further demonstrated that placebo effects based on remembered pain were 3 to 4 times greater than those based on concurrently assessed placebo effects, primarily because baseline pain was remembered as being much more intense than it actually was. However, similar to concurrent placebo effects, remembered placebo effects were strongly associated with expected pain levels that occurred just after conditioning. Taken together, these results suggest that magnitudes of placebo effect are dependent on multiple factors, including conditioning, expectancy, and whether analgesia is assessed concurrently or retrospectively. PMID- 10534586 TI - Comparative reliability and validity of chronic pain intensity measures. AB - Reliable and valid measures of pain are essential for conducting research on chronic pain. The purpose of this longitudinal study was to compare the reliability and validity of several measures of pain intensity. One hundred twenty-three patients with chronic pain were administered telephone interview versions of 0-10 scales of current, worst, least and average pain, immediately prior to beginning a multidisciplinary treatment program. The measures were administered again to these subjects 2 weeks (n=108), 1 month (n=106) and 2 months (n=105) after the end of treatment. The validity (defined as ability to detect changes in pain intensity over the course of treatment up to the 2-month follow-up assessment) and reliability (defined as stability over time in the 2 months after treatment) of these four measures and of composite combinations of these measures were examined. Contrary to prediction, the composite measures did not show a statistically significant superiority to the individual ratings in terms of their ability to detect change in pain intensity from pre-treatment to various points after treatment. The composite scores did, however, show greater stability than did the individual ratings after treatment. The practical conclusions of this study are; (1), individual 0-10 pain intensity ratings have sufficient psychometric strengths to be used in chronic pain research, especially research that involves group comparison designs with relatively large sample sizes, but, (2), composites of 0-10 ratings may be more useful when maximal reliability is necessary, (e.g. in studies with relatively small sample sizes, or in clinical settings where monitoring of changes in pain intensity in individuals is needed). PMID- 10534587 TI - Evidence for an inflammation-induced change in the local glutamatergic regulation of postganglionic sympathetic efferents. AB - Sympathetic efferents are involved in the pain of inflammation. Thus the control of these fibers is a matter of considerable importance. In this regard, postganglionic sympathetic fibers in normal rats express ionotropic glutamate receptors. The present study tests the hypothesis that inflammation leads to a significant increase in numbers of sympathetic efferents that express these receptors. In normal rats, the percentage of fibers in the L4 and L5 sympathetic gray rami immunostained with antibodies against subunits of NMDA (NMDAR1), AMPA (GluR1), or kainate (GluR5,6,7) receptors are 29, 5 and 5%, respectively. Forty eight hours following injection of complete Freund's adjuvant into one hindpaw, the percentages of fibers in the ipsilateral gray rami immunostained for NMDA, AMPA or kainate are 57, 52 and 48%, respectively. Thus, following inflammation there is a two-fold increase in axons expressing NMDA receptors and a ten-fold increase in axons expressing AMPA or kainate receptors. These data suggest that postganglionic activity may be enhanced by glutamate receptor activation during inflammation. Increased activity in postganglionic fibers could lead to an increased release of NE and other substances in postganglionic efferents such as prostaglandins which in turn could enhance nociceptor activity. This change in glutamate receptor organization offers a possible site of pharmacological intervention for the maladaptive symptoms that often arise following peripheral inflammation. PMID- 10534588 TI - Neuropathic pain from an experimental neuritis of the rat sciatic nerve. AB - Painful peripheral neuropathies involve both axonal damage and an inflammation of the nerve. The role of the latter by itself was investigated by producing an experimental neuritis in the rat. The sciatic nerves were exposed at mid-thigh level and wrapped loosely in hemostatic oxidized cellulose (Oxycel) that on one side was saturated with an inflammatory stimulus, carrageenan (CARRA) or complete Freund's adjuvant (CFA), and on the other side saturated with saline. In other rats, a myositis was created by implanting Oxycel saturated with CFA into a pocket made in the biceps femoris at a position adjacent to where the nerve was treated. Pain-evoked responses from the plantar hind paws were tested before treatment and daily thereafter. Statistically significant heat- and mechano hyperalgesia, and mechano- and cold-allodynia were present on the side of the inflamed nerve (CARRA or CFA) for 1-5 days after which responses returned to normal. There were no abnormal pain responses on the side of the saline-treated nerve, and none in the rats with the experimental myositis. The abnormal pain responses were inhibited by N-methyl-D-aspartate receptor blockade with MK-801, but were relatively resistant to the dose of morphine tested (10 mg/kg). Light microscopic examination of CARRA-treated nerves, harvested at the time of peak symptom severity, revealed that the treated region was mildly edematous and that there was an obvious endoneurial infiltration of immune cells (granulocytes and lymphocytes). There was either a complete absence of degeneration, or the degeneration of no more than a few tens of axons. Immunocytochemical staining for CD4 and CD8 T-lymphocyte markers revealed that both cell types were present in the epineurial and endoneurial compartments. The endoneurial T-cells appeared to derive from the endoneurial vasculature, rather than from migration across the nerve sheath. We conclude that a focal inflammation of the sciatic nerve produces neuropathic pain sensations in a distant region (the ipsilateral hind paw) and that this is not due to axonal damage. The neuropathic pain is specific to inflammation of the nerve because it was absent in animals with the experimental myositis and in those receiving sham-treatment. These results suggest that an acute episode of neuritis-evoked neuropathic pain may contribute to the genesis of chronically painful peripheral neuropathies, and that a chronic (or chronically recurrent) focal neuritis might produce neuropathic pain in the absence of significant (or clinically detectable) structural damage to the nerve. The model that we describe is likely to be useful in the study of the neuroimmune factors that contribute to painful peripheral neuropathies. PMID- 10534589 TI - Why is depression comorbid with chronic myofascial face pain? A family study test of alternative hypotheses. AB - A number of explanations have been proposed to account for findings that rates of depression are elevated in persons with chronic, non-malignant pain disorders (CNPDs); for example, that CNPDs are variants of depression (e.g. 'masked depression'), that the stress of living with CNPDs contribute to the onset of depression ('diathesis-stress'), or that the correlation of CNPDs and depression is a methodological artifact of studying treatment-seeking samples. These alternative hypotheses are tested for one specific CNPD, chronic myofascial face pain, using a family study methodology. The procedure was to conduct direct psychiatric interviews with 106 patients with a history of carefully diagnosed myofascial face pain, 118 acquaintance controls without personal histories of myofascial face pain, and a random sample of adult first degree relatives of these case and control probands. The probands were further subdivided into four roughly equal samples consisting of cases with and without personal histories of major depressive disorder (MDD), and controls with and without personal histories of MDD. Dates of initial onsets of myofascial face pain and MDD in patient probands were obtained from interviews and records. The main results were that, compared to control probands without personal histories of MDD, MDD and depressive spectrum disorders (DSD) were elevated in the first degree relatives of control probands with personal histories of early onset MDD, but not in the first degree relatives of myofascial face pain probands with or without personal histories of early or late onset MDD. This outcome is consistent with the hypothesis that living with chronic myofascial face pain contributes to elevated rates of depression. It is inconsistent with the alternative hypotheses that this CNPD is a variant of depression or that the elevated MDD rates are simply an artifact of selection into treatment. The implications of these results and additional results consistent with them are discussed. PMID- 10534590 TI - Percutaneous electrical nerve stimulation: an alternative to TENS in the management of sciatica. AB - Sciatica is a common pain problem and current pharmacologic therapies have proven inadequate for many patients. The objective of this sham-controlled investigation was to compare a novel non-pharmacologic technique, percutaneous electrical nerve stimulation (PENS), to transcutaneous electrical nerve stimulation (TENS) in the management of the radicular pain associated with sciatica. Sixty-four consenting patients with sciatica due to lumbar disc herniation were treated with PENS, TENS and sham-PENS according to a randomized, single-blinded, cross-over study. All patients had been maintained on a stable oral non-opioid analgesic regimen for at least 6 weeks prior to entering the study. Each treatment modality was administered for a period of 30 min three times per week for 3 weeks, with 1 week 'off' between each modality. Both PENS and TENS treatments were administered using a stimulation frequency of 4 Hz. The pre-treatment assessment included the health status survey short form (SF-36), as well as visual analog scales (VAS) for radicular pain, physical activity and quality of sleep. The pain VAS was also repeated after each treatment session. At the end of each 3-week treatment block, the SF-36 was repeated. After receiving all three treatment modalities, a global assessment questionnaire was completed. Both PENS (42%) and TENS (23%) were significantly more effective than the sham (8%) treatments in decreasing VAS pain scores. The daily oral analgesic requirements were also significantly reduced compared to the pre-treatment values with PENS (P<0.01) and TENS (P<0.05). However, PENS was significantly more effective than TENS (and sham-PENS) in improving physical activity and quality of sleep. The SF-36 evaluation confirmed the superiority of PENS (versus TENS and sham-PENS) with respect to post treatment functionality. In the overall assessment, 73% of the patients reported that PENS was the most desirable modality (versus 21% for TENS and 6% for sham PENS). Finally, 71% of the patients stated that they would be willing to pay extra to receive PENS therapy compared to 22% and 3% for TENS and sham-PENS, respectively. In this sham-controlled study, we concluded that PENS was more effective than TENS when administered at a stimulation frequency of 4 Hz in providing short-term pain relief and improved functionality in patients with sciatica. PMID- 10534591 TI - Responsiveness of general health status in chronic low back pain: a comparison of the COOP charts and the SF-36. AB - The objective of this study was to compare the responsiveness and assess the concurrent validity of two functional health status instruments, the Dartmouth COOP charts and the SF-36 in chronic low-back pain (CLBP) patients. The data came from 129 of 174 patients who participated in a randomized clinical trial of the therapeutic management of CLBP. Reliable and valid disease-specific outcomes, patient-rated low-back pain and disability, were used as external criteria (EC) to identify improved and non-improved patients. Unpaired t-statistics and receiver operating characteristic (ROC) curve calculations were used to quantify responsiveness. The two instruments had sufficient and very similar responsiveness using both EC. Comparisons between improved and non-improved patients for the COOP charts and SF-36, respectively, using pain as EC, yielded differences which translated into large effect sizes (0.8 and 0.7) (P=0.0008 and 0.003). Using disability as EC, differences of moderate effect size were found (0.5 and 0.6) (P=0.02 and 0.002). The ROC curve calculations using pain as EC resulted in areas under the curve of 0.76 (95% CI: 0.64, 0.88) for the COOP charts, and 0.74 (95% CI: 0.60, 0.88) for the SF-36. The corresponding areas using disability as EC were 0.67 (95% CI: 0.55, 0.79) and 0.72 (95% CI: 0.60, 0.84). The best cut-off point in both instruments for differentiating between improved and non-improved patients was approximately six percentage points. The constructs of functional health status, as reflected in the global scores of the two instruments, are highly correlated (r=0.82). Six of the instruments' nine dimensions are moderately to highly correlated (r=0.52 to 0.86), and the overall canonical correlation was high (R=0.9). In conclusion, both instruments seem equally suitable for use as outcome measures in clinical trials on CLBP. The COOP charts are faster to fill out and score. PMID- 10534592 TI - Complex regional pain syndrome: are the IASP diagnostic criteria valid and sufficiently comprehensive? AB - This is a multisite study examining the internal validity and comprehensiveness of the International Association for the Study of Pain (IASP) diagnostic criteria for Complex Regional Pain Syndrome (CRPS). A standardized sign/symptom checklist was used in patient evaluations to obtain data on CRPS-related signs and symptoms in a series of 123 patients meeting IASP criteria for CRPS. Principal components factor analysis (PCA) was used to detect statistical groupings of signs/symptoms (factors). CRPS signs and symptoms grouped together statistically in a manner somewhat different than in current IASP/CRPS criteria. As in current criteria, a separate pain/sensation criterion was supported. However, unlike in current criteria, PCA indicated that vasomotor symptoms form a factor distinct from a sudomotor/edema factor. Changes in range of motion, motor dysfunction, and trophic changes, which are not included in the IASP criteria, formed a distinct fourth factor. Scores on the pain/sensation factor correlated positively with pain duration (P<0. 001), but there was a negative correlation between the sudomotor/edema factor scores and pain duration (P<0.05). The motor/trophic factor predicted positive responses to sympathetic block (P<0.05). These results suggest that the internal validity of the IASP/CRPS criteria could be improved by separating vasomotor signs/symptoms (e.g. temperature and skin color asymmetry) from those reflecting sudomotor dysfunction (e.g. sweating changes) and edema. Results also indicate motor and trophic changes may be an important and distinct component of CRPS which is not currently incorporated in the IASP criteria. An experimental revision of CRPS diagnostic criteria for research purposes is proposed. Implications for diagnostic sensitivity and specificity are discussed. PMID- 10534593 TI - Prostaglandin E(2) has antinociceptive effect through EP(1) receptor in the ventromedial hypothalamus in rats. AB - The effects of microinjection of prostaglandin E(2) (PGE(2)) (50 fg-50 ng/0.2 microl) into the ventromedial hypothalamus (VMH) on nociception were studied using a hot-plate test in rats. Microinjection of PGE(2) (5-500 pg and 50 ng/0.2 microl) into the VMH significantly prolonged the paw-withdrawal latency on a hot plate 5 and 10 min after injection, respectively. Maximal prolongation was obtained 5 min after the injection of PGE(2) at 5 pg. Subsequently, to determine whether the PGE(2) receptor subtype EP(1) is involved in the PGE(2)-induced antinociceptive effect in the VMH, we observed the changes in nociception after intraVMH microinjection of SC19220, an EP(1) receptor antagonist, and 17-phenyl omega-trinor PGE(2), an EP(1) receptor agonist. Simultaneous injection of SC19220 (150 ng) with PGE(2) (500 pg) into the VMH blocked the PGE(2)-induced prolongation of the paw-withdrawal latency. Moreover, an intraVMH microinjection of 17-phenyl-omega-trinor PGE(2) (500 pg) prolonged it. These results indicate that PGE(2) in the VMH has antinociceptive effect through its actions on EP(1) receptors in rats. PMID- 10534594 TI - Generalised muscular hyperalgesia in chronic whiplash syndrome. AB - The whiplash syndrome has immense socio-economic impact. Despite extensive studies over the past years, the mechanisms involved in maintaining the pain in chronic whiplash patients are poorly understood. The aim of the present experimental study was to examine the muscular sensibility in areas within and outside the region involved in the whiplash trauma. Eleven chronic whiplash patients and 11 sex and age matched control subjects were included in the study. Before the experiment, the whiplash patients had pain in the neck and shoulder region with radiating pain to the arm. Five patients reported pain that was more widespread. The somatosensory sensibility in the areas over the infraspinatus, brachioradial, and anterior tibial muscles was assessed by pressure stimulation, pin-prick stimulation, and cotton swap stimulation. Infusion of hypertonic saline (5.85%, 0.5 ml) into the infraspinatus and anterior tibial muscles was performed to assess the muscular sensibility and referred pain pattern. The saline-induced muscle pain intensity was assessed on a continuous visual analogue scale (VAS). The distribution of pain was drawn on an anatomical map. The pressure pain thresholds were significantly lower in patients (P<0. 01) compared with controls: infraspinatus (mean 152.2 vs. 172.7 kPa), brachioradial (mean 70.0 vs. 363.8 kPa), and anterior tibial muscle (mean 172.7 vs. 497.8 kPa). The skin sensibility to pin-prick stimulation and cotton swap stimulation was not different between patients and controls. Infusion of hypertonic saline caused significantly higher VAS scores with longer duration in patients compared to control subjects (P<0.01). The area under the VAS-time curve was significantly (P<0.01) increased in patients compared to control subjects after injection into the infraspinatus muscle (mean 4138.1 vs. 780.0 cm s) and anterior tibial muscle (mean 4370.8 vs. 978.7 cm s). The saline infusion caused local pain defined as pain located around the injection site and referred pain areas not included in the local pain area. The area of local and referred pain were significantly larger in patients compared to control subjects (P<0.01). In the control group, the referred pain areas to infusion of hypertonic saline into the anterior tibial muscle were found at the dorsal aspect of the ankle. In contrast, the areas of referred pain were quite widespread in the patient group with both distal and proximal referred pain areas. In the present study, muscular hyperalgesia and large referred pain areas were found in patients with chronic whiplash syndrome compared to control subjects both within and outside the traumatised area. The findings suggest a generalised central hyperexcitability in patients suffering from chronic whiplash syndrome. This indicates that the pain might be considered as a neurogenic type of pain, and new pharmacological treatments should be investigated accordingly. PMID- 10534595 TI - Randomised clinical trial comparing the effects of acupuncture and a newly designed placebo needle in rotator cuff tendinitis. AB - Acupuncture has gained increasing attention in the treatment of chronic pain. The lack of a satisfying placebo method has made it impossible to show whether needling is an important part of the method or whether the improvement felt by the patient is due to the therapeutic setting and psychological phenomena. Also, the effectiveness of acupuncture has not been demonstrated sufficiently. We treated 52 sportsmen with rotator cuff tendinitis in a randomised single-blind clinical trial using a new placebo-needle as control. Patients were treated for 4 weeks. The primary endpoint of the trial was the change in the modified Constant Murley-score from the baseline. Assessment of the treatment outcome was made by experienced orthopaedists not informed of the treatment allocation. Acupuncture with penetration of the skin was shown to be more effective than a similar therapeutic setting with placebo needling in the treatment of pain. The acupuncture-group improved 19.2 Constant-Murley-score points (SD 16.1, range from -13 to 50), the control-group improved 8.37 points (SD 14.56, range from -20 to 41), (P=0.014; C.I. 2.3;19.4). This study showed that needling is an important part of the acupuncture effect in the treatment of chronic shoulder pain in athletes. No conclusions can be derived from this study concerning the importance of choosing points and the rules of Traditional Chinese Medicine. Using the new placebo method as control for other ailments could improve the evidence of specific acupuncture effects beyond pain treatment. PMID- 10534596 TI - Estrogen-induced alterations of spinal cord enkephalin gene expression. AB - Enkephalin-synthesizing neurons in the superficial laminae of the spinal and trigeminal dorsal horn are critical components of the endogenous pain-modulatory system. We have previously demonstrated that these neurons display intracellular estrogen receptors, suggesting that estrogen can potentially influence their enkephalin expression. By using Northern blot, we now show that a bolus injection of estrogen results in a rapid increase in spinal cord enkephalin mRNA levels in ovariectomized female rats. Thus, 4 h after estrogen administration the enkephalin mRNA-expression in the lumbar spinal cord was on average 68% higher (P<0.05) than in control animals injected with vehicle only. A small increase in the amount of enkephalin mRNA was also seen after 8 h (P<0.05), whereas no difference between estrogen-injected and control animals was found after 24 h or at time periods shorter than 4 h. Taken together with the previous anatomical data, the present findings imply that estrogen has an acute effect on spinal opioid levels in areas involved in the transmission of nociceptive information. PMID- 10534597 TI - Effects of intraplantar morphine on paw edema and pain-related behaviour in a rat model of repeated acute inflammation. AB - Recent studies suggest that peripheral morphine may represent a valuable treatment in inflammatory painful diseases. This study examined effects of intraplantar morphine against noxious pressure and paw edema in rats with repeated acute inflammation induced by two carrageenin injections 7 days apart. This model mimics at least partly some aspects of recurrent inflammatory pain encountered in the clinical situation. In the first part of the experiment, the effect of intraplantar morphine into the inflamed hindpaw was determined 3 h after carrageenin injection. Intraplantar morphine (50-200 microg) produced significant elevations of vocalization thresholds to paw pressure in inflamed but not in non-inflamed paws after both carrageenin injection; these effects were reversible by intraplantar naloxone methiodide (40 microg). The effects of intraplantar morphine (150 microg) were similar in magnitude to that of intravenous morphine (1 mg/kg) after first carrageenin injection. In contrast, at doses of 150-200 microg, they were significantly lower after second ipsilateral carrageenin injection 7 days later, than first injection. Intraplantar morphine (100-200 microg) had no effect on paw edema associated with both carrageenin injections. In the second part of the experiment, intraplantar morphine was injected 10 min before the first injection of carrageenin. Intraplantar morphine (50 microg) was ineffective, whereas morphine (100-200 microg) prevented reduction of vocalization thresholds to paw pressure of inflamed hindpaw for 3 h. The intraplantar injection of morphine (100 and 150 microg) produced a transient increase in the volume of inflamed hindpaw, not reversible by intraplantar naloxone methiodide (40 microg). Pretreatment with intraplantar morphine had no effect on reduction of vocalization thresholds to paw pressure and edema related to a second ipsilateral injection of carrageenin 7 days later. These findings suggest that peripheral morphine may be useful for the clinical management of acute inflammatory pain rather than in recurrent inflammatory painful situations. PMID- 10534598 TI - Electrical stimulation of motor cortex for pain control: a combined PET-scan and electrophysiological study. AB - Although electrical stimulation of the precentral gyrus (MCS) is emerging as a promising technique for pain control, its mechanisms of action remain obscure, and its application largely empirical. Using positron emission tomography (PET) we studied regional changes in cerebral flood flow (rCBF) in 10 patients undergoing motor cortex stimulation for pain control, seven of whom also underwent somatosensory evoked potentials and nociceptive spinal reflex recordings. The most significant MCS-related increase in rCBF concerned the ventral-lateral thalamus, probably reflecting cortico-thalamic connections from motor areas. CBF increases were also observed in medial thalamus, anterior cingulate/orbitofrontal cortex, anterior insula and upper brainstem; conversely, no significant CBF changes appeared in motor areas beneath the stimulating electrode. Somatosensory evoked potentials from SI remained stable during MCS, and no rCBF changes were observed in somatosensory cortex during the procedure. Our results suggest that descending axons, rather than apical dendrites, are primarily activated by MCS, and highlight the thalamus as the key structure mediating functional MCS effects. A model of MCS action is proposed, whereby activation of thalamic nuclei directly connected with motor and premotor cortices would entail a cascade of synaptic events in pain-related structures receiving afferents from these nuclei, including the medial thalamus, anterior cingulate and upper brainstem. MCS could influence the affective-emotional component of chronic pain by way of cingulate/orbitofrontal activation, and lead to descending inhibition of pain impulses by activation of the brainstem, also suggested by attenuation of spinal flexion reflexes. In contrast, the hypothesis of somatosensory cortex activation by MCS could not be confirmed by our results. PMID- 10534599 TI - Acute pain threshold in subjects with chronic pain following spinal cord injury. AB - Studies of pain perception in patients with chronic pain have yielded contradictory results. While several studies found that acute pain threshold is raised in chronic pain subjects, others showed that these subjects exhibit a decreased pain threshold compared to pain free subjects. The aim of this study was to further examine this topic by studying pain perception in subjects with chronic pain following partial or complete spinal cord injury (SCI). We found a significant elevation of heat-pain threshold (measured above the level of lesion) in complete SCI subjects with chronic pain (CSCIP) as opposed to complete SCI subjects without pain, incomplete SCI subjects with (ISCIP) and without chronic pain and normal controls. This elevation of pain threshold was completely reversed following a complete relief of the chronic pain by DREZ lesion. Moreover, the CSCIP exhibited significantly higher scores in the McGill pain questionnaire compared to ISCIP, indicative of a more intense chronic pain perceived by these subjects. In addition, the chronic pain below the level of spinal lesion, reported by CSCIP originated from a significantly larger body area than that of ISCIP. These results indicate that a critical level of chronic pain must be perceived in order to induce an elevation in acute pain threshold. PMID- 10534600 TI - N of 1 randomised controlled trials of oral ketamine in patients with chronic pain. AB - Anecdotal reports suggest that the general anaesthetic drug ketamine, taken orally in low doses, can give rise to some extra analgesia in patients with refractory neuropathic pain. This study was designed to determine the proportion of patients with chronic neuropathic pain responding to oral ketamine, and then to separate the true treatment effect from non-specific effects by means of an n of 1 randomised controlled trial. Twenty-one patients gave informed consent and completed daily pain diaries and continued on their usual treatments (drug and non-drug) for the duration of the study. After a 'baseline' week, oral ketamine was taken once a day for 1 week. The dose of 20 mg was increased each day until an analgesic effect was noticed or adverse effects occurred, or until a maximum of 100 mg was reached. Those patients responding to oral ketamine were then entered into the n of 1 randomised trial which consisted of three treatment/placebo week pairs. Twelve patients did not progress to the n of 1 trial because of no benefit and/or intolerable adverse effects (dizziness, drowsiness etc.). Nine patients completed the n of 1 trial; there was no difference between the ketamine and placebo weeks in six patients; one patient demonstrated effective analgesia with ketamine, but it was of short duration and marred by unpleasant adverse effects; two patients showed some evidence of a beneficial response to ketamine, and continued with the oral ketamine after the trial. We conclude that oral ketamine only gave rise to an extra analgesic response in three out of 21 patients with chronic neuropathic pain (14%). Adverse effects limited the use of the drug in almost half of the patients. The n of 1 trial was useful in demonstrating no true therapeutic effect for the ketamine in two thirds of the patients progressing to that part of the trial. PMID- 10534601 TI - Heat-induced release of CGRP from isolated rat skin and effects of bradykinin and the protein kinase C activator PMA. AB - In the skin, noxious heating induces an axon reflex response which is commonly accepted to be due to the release of vasodilatory neuropeptides from polymodal nociceptors. In the present study, the quantitative assessment of calcitonin gene related peptide (CGRP) release from rat skin serves as an integrative measure of primary afferent activation by noxious heat and the presumed sensitising action of bradykinin and an activator of protein kinase C (PKC). The isolated rat hairy skin of either hind paw was mounted on acrylic rods and exposed for 5 min periods to synthetic interstitial fluid of either 32 degrees C for control or of higher temperatures up to 59 degrees C during stimulation. In addition, experiments were performed in calcium free solution (containing 10 mM EGTA) or the skin was preloaded with the membrane permeant calcium chelator BAPTA-AM (1 mM). To look for modulatory effects on the heat responses, bradykinin or polymyristate-acetate (PMA) were added during heat stimulation in further experiments. Heating the skin induced a temperature-dependent release of CGRP from a threshold of 43 degrees C which was absent in calcium free solution. Only at the highest temperatures (55 and 59 degrees C) was a partially calcium-independent release observed. Inhibition of the release was also obtained with the intracellular calcium buffer BAPTA-AM. Bradykinin 10 but not 1 microM as well as PMA 1 and 10 microM significantly facilitated the heat-induced CGRP release at 47 degrees C whereby BK caused a marginal and PMA a significant CGRP release by itself. Our results indicate that moderate noxious heat induces calcium-dependent CGRP release and this can be facilitated by bradykinin and by the activation of PKC. This suggests the same sensitising mechanism that affects nociceptor heat responses. PMID- 10534602 TI - Postoperative recovery evaluated with a new, tactile scale (TaS) in children undergoing ophthalmic surgery. AB - Following pediatric eye surgery, visual scales for assessment of recovery are inadequate due to impairment of vision. A tactile scale (TaS) was therefore developed and tested in a pilot-trial. Fifty children, 23 girls and 27 boys undergoing different types of ophthalmic surgery used TaS to rate postoperative pain and nausea. TaS was easy to explain to children and easy to use. Ratings ranging from light discomfort to severe pain were made by 49 children, whereas one child reported no pain at any time following surgery. Analgesics were administered to 38% of the patients and the administration of analgesics was significantly (P=0.01, simple regression) related to the ratings of pain by TaS. The mean ratings of pain by TaS were significantly (P<0.05, General Linear Model followed by Dunnett's t-test) lower up to 3 h after the administration of analgesics compared to ratings before analgesics were given, indicating that ratings by TaS were related to the children's actual level of pain. Nausea and/or vomiting was common and was reported or recorded in 28 children (56%). After further validation, TaS may be a useful tool for assessment of postoperative pain and efficacy of given treatments in children and adults, not only after eye procedures, but also following other types of surgery. PMID- 10534603 TI - Detection of static and dynamic components of mechanical allodynia in rat models of neuropathic pain: are they signalled by distinct primary sensory neurones? AB - In the present study, chronic constrictive injury (CCI model) of the sciatic nerve or tight ligation of L5 and L6 spinal nerves (Chung model) produced both dynamic and static components of mechanical allodynia in rats. The two responses were detected, respectively, by lightly stroking the hind paw with cotton wool or application of pressure using von Frey hairs. Animals with spinal nerve ligation developed both types of responses at a faster rate compared to animals with the CCI. Morphine (1-3 mg/kg, s.c.) dose-dependently blocked static but not dynamic allodynia. In contrast, pregabalin (previously S-isobutylgaba and CI-1008) dose dependently (3-30 mg/kg, p.o.) blocked both types of allodynia. In CCI animals, two administrations of capsaicin (100 microg/50 microl) into the plantar surface of the ipsilateral paw at 1-h intervals blocked the maintenance of thermal hyperalgesia without affecting either static or dynamic allodynia. The similar administration of a further two doses of capsaicin into the same animals blocked the maintenance of static allodynia without affecting the dynamic response. These data indicate that thermal hyperalgesia, static and dynamic allodynia are respectively signalled by C-, Adelta- and Abeta/capsaicin insensitive Adelta- primary sensory neurones. It is suggested that pregabalin possesses a superior antiallodynic profile than morphine and may represent a novel class of therapeutic agents for the treatment of neuropathic pain. PMID- 10534605 TI - The NMDA receptor antagonist MK-801 attenuates c-Fos expression in the lumbosacral spinal cord following repetitive noxious and non-noxious colorectal distention. AB - The effects of pretreatment with an NMDA receptor antagonist, MK-801, on c-Fos (Fos) expression in the lumbosacral spinal cord following repetitive, noxious (80 mmHg) or non-noxious (20 mmHg) colorectal distention (CRD) was examined immunocytochemically in awake and urethane anesthetized rats. In awake rats, noxious CRD induced Fos expression in the lumbosacral spinal cord. Pretreatment with MK-801 (0.1-1.0 mg/kg, i.p.) produced no change or an increase in noxious CRD induced-Fos expression and caused aversive side effects. In order to examine greater doses of MK-801, further experiments were performed in rats anesthetized with urethane. Both noxious and non-noxious CRD induced Fos in the lumbosacral spinal cord. Pretreatment with MK-801 (0.5, 1.0, 5.0 mg/kg, i.p.) dose dependently attenuated noxious CRD-induced Fos by 20-40%. Five mg/kg MK-801 attenuated non-noxious CRD-induced Fos by 20%. Lesser doses did not significantly attenuate Fos expression. The laminar distribution of Fos following MK-801 pretreatment revealed a tendency towards the deeper laminae showing the greatest attenuation at the highest dose of MK-801. Protein plasma extravasation in the colon measured with Evan's blue dye showed no difference between rats without balloons, rats with balloons that were not distended and non-noxious CRD. There was significantly more extravasation following noxious CRD. Pretreatment with systemic MK-801 had no effect on plasma extravasation produced by noxious CRD. These data suggest that the induction of Fos in the lumbosacral spinal cord by noxious and non-noxious CRD is partially NMDA receptor mediated. However, NMDA receptor activation contributes significantly more to noxious than non-noxious CRD-induced Fos. Inflammation of the colon following noxious CRD likely contributes to sensitization of colonic afferents which may contribute to the increased NMDA receptor-mediated Fos following the noxious stimulus. PMID- 10534604 TI - Hyposecretion of adrenal androgens and the relation of serum adrenal steroids, serotonin and insulin-like growth factor-1 to clinical features in women with fibromyalgia. AB - Neuroendocrine deficiencies have been implicated in fibromyalgia (FM). In the present study, adrenal androgen metabolites and their relationship with health status in FM were investigated. For comparison, serum levels of other implicated neuroendocrine mediators were correlated with health status. Fifty-seven consecutive women with FM completed the Fibromyalgia Impact Questionnaire (FIQ). Fasting blood samples were taken for measurement of dehydroepiandrosterone sulphate (DHEAS), free testosterone (T), cortisol, serotonin and insulin-like growth factor-1. Normal value for DHEAS and T were obtained from 114 controls. DHEAS levels were decreased significantly in pre- and postmenopausal patients (P<0.0001 and P<0.0005, respectively). T levels were decreased significantly in premenopausal and insignificantly in postmenopausal patients (P<0.0001 and P=0.06, respectively). The following correlations between neurohormonal levels and FIQ scores were found: DHEAS (after adjustment for age) vs. pain (P<0.001) and T (after adjustment for age) versus physical functioning (P=0.002). None of the other neurohormonal levels correlated significantly with any of the FIQ scores. IGF-1 levels were lower in the obese patients as compared to those who were non-obese (P=0.03). The BMI correlated positively with pain (P<0. 001) and inversely with DHEAS levels (P=0.006). After further adjustment for BMI, the correlation between age adjusted DHEAS and pain was no longer significant. Hyposecretion of adrenal androgens was documented in FM. This was more pronounced in obese patients. Low serum androgen levels correlated with poor health status in FM. Longitudinal studies are needed to elucidate whether these are cause and/or effect relationships. PMID- 10534606 TI - Sex differences in pain perception and cardiovascular responses in persons with parental history for hypertension. AB - We investigated gender differences in cardiovascular and pain responses to the cold pressor (CP) test in persons with positive (PH+) or negative parental history (PH-) for hypertension. Previous work has suggested an attenuated sensitivity to painful stimulation in hypertensive men and more recently in men with parental disposition for hypertension. It is not known whether this hypoalgesic effect is present in PH+ women. In this study, we evaluated differences in pain perception between men and women with PH+ or PH- using an assessment method to measure current as well as delayed pain. Participants rated their pain every 15 s during a 90-s hand CP (0-4 degrees C) and a 90-s post-CP rest period. Systolic and diastolic blood pressures (SBP, DBP) and heart rate (HR) were measured before, during, and after the CP. PH+ and PH- groups did not differ in age, height, weight, education, resting SBP, DBP, or HR. PH+ men showed greater DBP responses to the CP than PH- men, while female groups did not differ in cardiovascular responses to the CP. Although pain ratings during the CP did not differ between groups, post-CP reported pain receded faster in the PH+ men than in the PH- men. PH+ women, on the other hand, tended to report greater pain than PH- women. These findings question the generalizability of the hypoalgesic effects in hypertension-prone women. PMID- 10534607 TI - The kappa opioid nalbuphine produces gender- and dose-dependent analgesia and antianalgesia in patients with postoperative pain. AB - Nalbuphine, pentazocine, and butorphanol, mixed agonist/antagonist opioids that induce analgesia by acting predominantly at kappa opioid receptors, have recently been shown in single-dose studies to have greater analgesic efficacy in women than in men. In the current experiments, the first placebo controlled dose response study of opioid analgesic efficacy that examines for gender differences, nalbuphine (5, 10, or 20 mg) and placebo were evaluated in 62 men and 69 women for the treatment of moderate to severe postoperative pain following extraction of impacted wisdom teeth. In a randomized, open injection, double blind experimental design, pain intensity was recorded on a 10 cm visual analog scale (VAS) immediately prior to drug administration (baseline) and at 20 min intervals thereafter. Although responses to placebo were similar in men and women, for all doses of nalbuphine women exhibited significantly greater analgesic response than men, compatible with our previous results. Unexpectedly, men receiving the 5 mg dose of nalbuphine experienced significantly greater pain than those receiving placebo; only the 20 mg dose of nalbuphine in men produced significant analgesia compared to placebo. While a similar antianalgesic effect was not observed in women, only the 10 mg dose of nalbuphine produced significant analgesia compared to placebo. These results suggest that the optimal analgesic dose of nalbuphine for women is lower than the highest dose that can be safely administered. In contrast, the antianalgesic effect of nalbuphine suggests avoidance of its routine use for postoperative analgesia in men until further studies clarify this issue. Because gender differences in other mixed kappa agonists/antagonists (i.e. pentazocine and butorphanol) have previously been shown, these results may generally apply to this class of opioid analgesics. PMID- 10534608 TI - Parenterally administered kainic acid induces a persistent hyperalgesia in the mouse and rat. AB - Nociceptive primary afferent C-fibers express a subset of glutamate receptors that are sensitive to kainic acid. Thus, we tested the possibility that activation of these receptors alters nociception. Intraperitoneal (i.p.) injection of kainic acid induced a persistent thermal hyperalgesia, when tested using the hot plate (mice) and tail flick (mice and rats) assays, and mechanical hyperalgesia when tested using von Frey monofilaments (rats), but had no effect on acetic acid-induced chemical nociception (mice). When administered i. p., 6 cyano-7-nitroquinoxaline-2,3-dione (CNQX), an (R, S)-alpha-amino-3-hydroxy-5 methylisoxazole-4-proprionic acid HBr/kainate (AMPA/KA) antagonist, completely blocked hyperalgesia. When injected intrathecally (i.t.), kainic acid itself failed to induce hyperalgesia and AMPA/KA antagonists given i.t. also failed to attenuate the hyperalgesic effect of kainic acid administered i.p. , indicating that the spinal cord is not the primary site of action. Kainic acid injected subcutaneously in the back of mice decreased response latencies in the hot plate and tail flick assays, indicating that hyperalgesia is achieved by a variety of parenteral routes of injection. Histological evaluation of rat spinal cord and dorsal root ganglia revealed no neurodegenerative changes 24 h after kainic acid. Together these data suggest that a persistent hyperalgesia results from the transient activation of AMPA/KA receptors that are located outside the spinal cord, perhaps on the distal projections of primary afferent fibers. PMID- 10534609 TI - Gestational and ovarian sex steroid antinociception: relevance of uterine afferent and spinal alpha(2)-noradrenergic activity. AB - Pregnancy is associated with an antinociception that is multifactorial and results from spinal (kappa/delta) opioid antinociceptive pathways as well as peripheral processes (ovarian sex steroids, uterine afferent neurotransmission). The present results provide the first indication that the full manifestation of pregnancy-induced analgesia also requires a supraspinal component. The analgesia of gestation or its hormonal simulation (via estrogen and progesterone administration; HSP) is substantially attenuated (>/=60%) following blockade of spinal alpha(2) (but not alpha(1)) adrenergic receptors. HSP antinociception is also attenuated by transection of the hypogastric nerve, the magnitude of which is indistinguishable from that produced by spinal alpha(2) receptor blockade. Additionally, hypogastric neurectomy abolishes the component of the antinociception associated with HSP that is mediated by spinal alpha(2) receptors. This suggests that the augmented spinal noradrenergic activity during HSP is not due to activation at the terminal of noradrenergic spinal projection neurons but requires supraspinal activity. It is suggested that enhanced spinal noradrenergic activity amplifies ongoing spinal kappa/delta antinociception as has been observed following the concomitant intrathecal application of alpha(2) and opioid agonists. The current observations underscore the importance of visceral afferent activity as well as its modulation by a female-specific hormonal milieu to the efficacy of endogenous spinal opioid antinociception. PMID- 10534610 TI - Prediction of physician visits and prescription medicine use for back pain. AB - The primary purpose of this study was to examine the extent to which specific patient attitudes and beliefs about medical care and self-care for back pain predict future healthcare use. An automated database allowed examination of the predictive relationships in two primary care patient samples. In general, beliefs that physicians should find a definitive cause and permanent cure for back pain predicted neither physician visits nor prescription medication fills. Patient attitudes endorsing the benefits of medical treatment for back pain (as opposed to a permanent cure) predicted the use of these specific healthcare services. In a third sample of primary care back pain patients, we assessed whether a four session self-care intervention modified those attitudes and beliefs shown to predict future healthcare use. The group intervention was associated with changes in attitudes about use of physician services but not medication use. A secondary purpose was to examine initial psychometric properties of a proposed back pain Self-Care Orientation Scale made up of the original 11 items. Factor analyses of the item set yielded three factors, but inconclusive results; the internal consistency of the identified sub-scales was only moderate. However, findings that a subset of items predicted physician visits and prescriptions medication fills, and was sensitive to change following a self-care intervention, suggest avenues for improving measurement of self-care orientation. These findings help clarify specific patient attitudes and beliefs that are related to healthcare utilization and suggest that a subset of these beliefs can be modified through a brief educational intervention. PMID- 10534611 TI - Supraclavicular glomus tumor, 20 year history of undiagnosed shoulder pain: a case report. AB - A long-standing case of severe dysesthesia due to a supraclavicular glomus tumor is presented. Chronic pain caused by a subcutaneous glomus (non-chemodectoma) tumor is rare and usually misdiagnosed. The supraclavicular location, presentation, and coincidence of trauma history are unique in this case. A 62 year-old male complained of 20 years of intractable right shoulder and supraclavicular region pain, which started 6 months after a fall. The pain was unrelieved by repeated and extensive physical therapy, chiropractic manipulation, local steroid injections, and two shoulder operations. The cause of the condition remained undiagnosed and obscure. Local surgical exploration revealed a subcutaneous grayish mass with pathologically proven glomus tumor. Immediate alleviation of the pain and tenderness followed complete resection of the mass. The patient remained free of pain at a 2-year follow-up. Subcutaneous glomus (non chemodectoma) tumors can occur in unusual sites, and should be considered in chronic regional pain syndromes. Immediate cure is generally achieved by local resection. Pertinent literature is reviewed. PMID- 10534612 TI - Positive effect of regional analgesia (RA) in terminal stage paediatric chondrosarcoma: a case report and the review of the literature. AB - A 10-year-old girl was treated for progressive left pelvic chondrosarcoma and severe local pain radiating to the ipsilateral lower extremity. Despite high doses of opioids, pain was poorly controlled and treatment resulted in urine retention and constipation. Positive effect on pain (143 out of 181 days) was obtained by regional analgesia. Continuous lumbar epidural opioid infusion led to pain relief and disappearance of symptoms. Port-catheter dysfunction necessitated a change of epidural catheter and the patient was treated that with morphine, bupivacaine and clonidine plus clonazepam which resulted in relief of constipation and restoration of urinary function. The patient subsequently developed an abscess required or subarachoid infusion (morphine associated with clonazepam, clomipramine and corticosteroids). Later bilateral controlateral cordotomy was performed due to absence of analgesia and the patient subsequently died of tumour progression. PMID- 10534613 TI - Ectodysplasin, a protein required for epithelial morphogenesis, is a novel TNF homologue and promotes cell-matrix adhesion. AB - In the mouse Tabby (Ta) mutant and human X-linked anhidrotic ectodermal dysplasia (EDA) syndrome development of several ectodermal organs such as hair, teeth, and sweat glands is impaired. The gene behind Tabby and EDA has been cloned, and several alternative transcripts have been isolated. The protein product named ectodysplasin had no obvious function or prominent homology to other known gene products apart from a short collagen-like sequence. We have isolated two novel Ta transcripts which are variants of the longest isoform of Tabby, named Ta-A. In situ hybridizations revealed Ta-A to be the major transcript in the developing embryo. It was detected in the endoderm of early embryos and subsequently in specific locations in the neuroepithelium and ectoderm. Unexpectedly, sequence analysis of the most C-terminal domain of Ta revealed that ectodysplasin is a novel member of the tumor necrosis factor (TNF) ligand superfamily. Mouse ectodysplasin was biochemically and functionally characterized, and shown to be a glycosylated, oligomeric type II membrane protein (N-terminus inside), all characteristics typical to TNF-like proteins. Members of the TNF family are critically involved in host defence and immune response often mediating either apoptosis or cell survival. Expression of Ta in several epithelial cell lines did not result in prominent changes in cell morphology and did not promote apoptosis. Instead, it was shown to promote cell adhesion to extracellular matrix, a function consistent with its postulated role in epithelial-mesenchymal interactions regulating the development of ectodermal appendages. Ectodysplasin is the first TNF-like signaling molecule described known to be required for epithelial morphogenesis. PMID- 10534614 TI - Disrupting the establishment of polarizing activity by teratogen exposure. AB - Between days 9.5 and 10, the forelimb buds of developing murine embryos progress from stage 1 which are just beginning to express shh and whose posterior mesoderm has only weak polarizing activity to stage 2 limbs with a distinguishable shh expression domain and full polarizing activity. We find that exposure on day 9.5 to teratogens that induce the loss of posterior skeletal elements disrupts the polarizing activity of the stage 2 postaxial mesoderm and polarizing activity is not subsequently restored. The ontogeny of expression of the mesodermal markers shh, ptc, bmp2, and hoxd-12 and 13, as well as the ectodermal markers wnt7a, fgf4, fgf8, cx43, and p21 occurred normally in day 9.5 teratogen-exposed limb buds. At stage 3, the treated limb apical ectodermal ridge usually possessed no detectable abnormalities, but with continued outgrowth postaxial deficiencies became evident. Recombining control, stage matched limb bud ectoderm with treated mesoderm prior to ZPA grafting restored the duplicating activity of treated ZPA tissue. We conclude that in addition to shh an early ectoderm-dependent signal is required for the establishment of the mouse ZPA and that this factor is dependent on the posterior ectoderm. PMID- 10534616 TI - Syndactyly of Ft/+ mice correlates with an imbalance in bmp4 and fgf8 expression. AB - The most obvious phenotype of Ft/+ mice is a syndactyly of fore limbs characterised by a fusion of the tips of digits 1 to 4. The tempospatial expression of genes involved in limb development revealed that patterning of Ft/+ limb buds is not affected by the mutation. However, an upregulation of Bmp4 in the anterior-distal region of the limb bud at d12.0 of embryonic development is accompanied by a loss of Fgf8 expression in the distal part of the AER. Downstream target genes of Bmp action such as Msx1 and 2 are upregulated. This induction of the signalling cascade indicates ectopic expression of functional Bmp4. Nevertheless, analysis of physical parameters of bones from adult mice revealed a reduction of the bone mass of the autopod. The data suggest a negative effect of Bmp4 on Fgf8 expression and a positive influence on the induction of bone elements. PMID- 10534615 TI - Induction of apoptosis in the germline and follicle layer of Drosophila egg chambers. AB - The reaper and head involution defective genes can induce apoptotic death in several Drosophila cell types, including portions of the embryo and eye. By a combination of FLP recombinase and the yeast Gal4/UAS transcription activation system, we expressed both cell death genes in discrete clones in the adult ovarian follicle cell layer. The expression of either reaper or head involution defective induced follicle cell apoptosis during all oogenic stages. Unexpectedly, the disruption of the follicle layer led to the induced degeneration of the nurse cells in an apoptotic manner, demonstrating a germline somatic interaction required for germ cell viability. The germline apoptosis initiates at a specific time in oogenesis, coinciding with the beginning of vitellogenesis. This observation is intriguing given previous suggestions of a process to eliminate defective egg chambers at these same oogenic stages. The induce germline degeneration initiates with the transient formation of a network of filamentous actin around the nurse cell nucleus, in close association with a product of the adducin-related hu-li tai shao gene. This was immediately followed by nuclear condensation and DNA fragmentation, both characteristics diagnostic of apoptosis. Occurring concomitantly with the nuclear phenotypes were the disorganization of ring canals, and the degradation of Armadillo protein (a beta catenin homolog) and filamentous actin. Germ cells degenerating as a normal consequence of oogenesis displayed a similar set of phenotypes, suggesting that a common apoptotic mechanism may underlie these different germline death phenomena. PMID- 10534617 TI - In vivo analysis of Frat1 deficiency suggests compensatory activity of Frat3. AB - The Frat1 gene was first identified as a proto-oncogene involved in progression of mouse T cell lymphomas. More recently, FRAT/GBP (GSK-3beta Binding Protein) family members have been recognized as critical components of the Wnt signal transduction pathway. In an attempt to gain more insight into the function of Frat1, we have generated Frat1-deficient mice in which most of the coding domain was replaced by a promoterless beta-galactosidase reporter gene. While the pattern of LacZ expression in Frat1(lacZ)/+ mice indicated Frat1 to be expressed in various neural and epithelial tissues, homozygous Frat1(lacZ) mice were apparently normal, healthy and fertile. Tissues of homozygous Frat1(lacZ) mice showed expression of a second mouse Frat gene, designated Frat3. The Frat1 and Frat3 proteins are structurally and functionally very similar, since both Frat1 and Frat3 are capable of inducing a secondary axis in Xenopus embryos. The overlapping expression patterns of Frat1 and Frat3 during murine embryogenesis suggest that the apparent dispensability of Frat1 for proper development may be due to the presence of a second mouse gene encoding a functional Frat protein. PMID- 10534618 TI - The LIM homeodomain protein dLim1 defines a subclass of neurons within the embryonic ventral nerve cord of Drosophila. AB - Members of the LIM homeodomain family of transcription factors have been implicated in specifying cell identity in a range of species. In Drosophila three LIM homeobox genes, apterous, lim3 and isl, have been shown to control axon pathfinding of subsets of neurons within the embryo. Here we describe the isolation and characterization of another LIM homeobox gene in Drosophila termed dlim1, a homolog of the vertebrate Lim-1 gene. The sequence and expression of dLim1 is highly related to its vertebrate homologs. Within the Drosophila embryo, dLim1 is expressed in the head primordia, the brain lobes, and in distinct sets of motorneurons and interneurons within the ventral nerve cord. Comparatively in vertebrates, Lim-1 (Lhx1) along with Lim-3 (Lhx3), Gsh-4 (Lhx4), Isl-1 and Isl-2 are expressed in developing motorneurons along the spinal column, where their overlapping expression suggests a role for these genes in the establishment of specific motorneuron subtypes. dLim1 is absent from all cells expressing Isl, Lim3, and Apterous, indicating that these proteins function independently within the Drosophila embryo. To investigate the function of dlim1, we generated loss-of function mutations within the locus. Our findings show that dlim1 is an essential gene that when mutated results in lethality near the larval-pupal boundary. In contrast to vertebrate Lim-1, dlim1 has no apparent role in anterior patterning of the Drosophila embryo. Our analysis shows that dlim1 has been evolutionarily conserved, however the Drosophila lim1 gene exhibits unique properties that distinguishes it from its vertebrate homologs. PMID- 10534619 TI - Neural cell fate in rca1 and cycA mutants: the roles of intrinsic and extrinsic factors in asymmetric division in the Drosophila central nervous system. AB - In the central nervous system (CNS) of Drosophila embryos lacking regulator of cyclin A (rca1) or cyclin A, we observe that several ganglion mother cells (GMCs) fail to divide. Whereas GMCs normally produce two sibling neurons that acquire different fates ('A/B'), non-dividing GMCs differentiate exclusively in the manner of one of their progeny ('B'). In zygotic numb mutants, sibling neuron fate alterations ('A/B' to 'A/A') occur infrequently or do not occur in some sibling pairs; we have determined that depletion of both maternal and zygotic numb causes sibling neurons to acquire equalized fates ('A/A') with near-complete expressivity. In rca1, numb mutant embryos, we observe binary cell fate changes ('B' to 'A') in several GMCs as well. Finally, we have demonstrated that expression of Delta in the mesoderm is sufficient to attain both sibling fates. Our results indicate that the intrinsic determinant Numb is absolutely required to attain differential sibling neuron fates. While the extrinsic factors Notch and Delta are also required to attain both fates, our results indicate that Delta signal can be received from outside the sibling pair. PMID- 10534620 TI - Gut specific expression using mammalian promoters in transgenic Xenopus laevis. AB - The recent development of transgenic methods for the frog Xenopus laevis provides the opportunity to study later developmental events, such as organogenesis, at the molecular level. Our studies have focused on the development of the tadpole gut, where tissue specific promoters have yet to be identified. We have used mammalian promoters, for the genes elastase, pancreatic duodenal homeobox-1, transthyretin, and intestinal fatty acid binding protein to drive green fluorescent protein expression in live tadpoles. All of these were shown to drive appropriate tissue specific expression, suggesting that the molecular mechanisms organising the gut are similar in amphibians and mammals. Furthermore, expression from the elastase promoter is initiated in the pancreatic buds before morphological definition becomes possible, making it a powerful tool for the study of pancreatic determination. PMID- 10534621 TI - GFP-tagged balancer chromosomes for Drosophila melanogaster. AB - We constructed green fluorescent protein (GFP)-expressing balancer chromosomes for each of the three major chromosomes of Drosophila melanogaster. Expression of GFP in these chromosomes is driven indirectly by a Kruppel (Kr) promoter, via the yeast GAL4-UAS regulatory system. GFP fluorescence can be seen in embryos as early as the germ band extension stage, and can also be seen in larvae, pupae, and adults. We show the patterns of GFP expression of these balancers and demonstrate the use of the balancers to identify homozygous progeny. PMID- 10534622 TI - Expression of the Ets transcription factors erm and pea3 in early zebrafish development. AB - Here we report the cloning of zebrafish erm and pea3 cDNAs, which are members of the PEA3 subgroup of Ets transcription factors. The expression patterns of these two genes were examined during zebrafish embryogenesis. Maternal mRNAs of both genes are detectable and transcripts are found ubiquitously until the late blastula, in the marginal zone of gastrula stages and in the presumptive fore- and hindbrain and in the trunk region of early somite stages. Later, erm expression is observed in distinct regions of the forebrain, the mid-/hindbrain boundary, the differentiating rhombomeres, branchial arches, otic vesicles, the pectoral fins, the somites and the tailbud in a dynamic fashion. In contrast, pea3 is not expressed in the rhombomeres but in the Rohon-Beard neurons, the sensory lateral line placodes and the heart. PMID- 10534624 TI - The spatial and temporal pattern of C-Lmx1 expression in the neuroectoderm during chick neurulation. AB - C-Lmx1 has been shown to be a key regulatory gene for specification of dorsoventral pattern during vertebrate limb development. Here, we describe its earlier pattern of expression during and shortly after neurulation. Transcripts are first expressed in the mesoderm of the head process and rostral tip of the primitive streak at the late gastrula/early neurula stage (stage 4). As neurulation occurs with shaping of the neural plate, C-Lmx1 is expressed in a butterfly-like pattern in the lateral neuroectoderm. During bending of the neural plate, C-Lmx1 expression becomes localized to three areas of the bending neuroectoderm: the median hingepoint (future floor plate of the neural tube) and the paired dorsolateral regions of the neuroepithelium, including the dorsolateral hingepoints and the adjacent neuroectodermal and epidermal ectodermal components of the neural folds. After closure of the neural groove and formation of the primary brain vesicles, C-Lmx1 is expressed in the dorsal neural tube along the entire length of the neuraxis, as well as in the floor plate at the brain but not spinal cord levels. At the midbrain and rostral hindbrain levels, C-Lmx1 is heavily expressed. Here, in addition to expression in the dorsal neural tube and floor plate, it is expressed in the lateral walls of the neural tube, with the exception of the levels of rhombomeres 2 and 4. C-Lmx1 is also expressed in several other discrete domains during and shortly after neurulation, including the prechordal plate and rostral head mesenchyme, foregut endoderm, otic placode and vesicle, dorsal somitic mesoderm, midline endoderm at the level of the caudal spinal cord, mesonephroi and limb bud mesoderm. PMID- 10534623 TI - Characterization of npas3, a novel basic helix-loop-helix PAS gene expressed in the developing mouse nervous system. AB - Here we describe the cloning and expression pattern of a new bHLH-PAS domain gene, Npas3. Npas3 shares 50.2% amino acid sequence identity with Npas1 and a lesser similarity with other members of the bHLH-PAS domain family of transcription factors. Northern blot analysis detected Npas3 mRNA between 11.5 and 17.5 d.p.c. in embryonic development and exclusively in the adult brain. Whole-mount and section in situ hybridization assays revealed expression of Npas3 between 9.5 and 11.5 d.p.c. in the developing neural tube. In addition, Npas3 mRNA was expressed throughout the neuroepithelium of the developing central nervous system between 10. 5 and 12.5 d.p.c. Interestingly, at 14.5 d.p.c., the expression of Npas3 mRNA became restricted to the neopallial layer of the cortex. At 12.5 d.p.c., Npas3 mRNA was evident in nonneural tissues such as the developing dermis and mesenchyme surrounding the otic and nasal placodes. Expression was also detected in the developing cardiac valves, limb and developing kidney. PMID- 10534625 TI - Xpitx-1: a homeobox gene expressed during pituitary and cement gland formation of Xenopus embryos. AB - Pitx-1 is a member of the family of bicoid-related vertebrate homeobox genes; it was originally identified as a tissue-specific transcriptional regulator of the proopiomelacortin gene. Here we report on the embryonic expression of Xpitx-1, which is expressed in the anterior neural ridge and in the cement gland Anlage during late gastrulation/early neurulation. In tadpole stage embryos Xpitx-1 transcripts are primarily detected in the cement gland, stomodeal-hypophyseal Anlage, oral epithelia and lens placode. Therefore, Xpitx-1 may be part of the genetic network that controls the early development of these structures. PMID- 10534626 TI - Expression of prestalk and prespore proteins in minute, two-dimensional Dictyostelium slugs. AB - We show that exceedingly small two-dimensional slugs of Dictyostelium differentiate normally and have an anterior prestalk zone and a posterior prespore zone. Using GFP as a marker attached to the appropriate promoter, prestalk expression is concentrated in the anterior, while prespore expression is produced in the posterior, closely resembling what is found in normal, large slugs. PMID- 10534627 TI - An overview of sexually transmitted diseases. Part II. AB - Sexually transmitted diseases are a persistent problem in the United States and throughout the world. Many of these infections involve the skin and may be encountered in the field of dermatology. This 3-part review highlights the cutaneous features, diagnosis, and treatment of 11 of the most common sexually transmitted diseases, other than AIDS. The second part of this series focuses on anogenital warts, chronic viral hepatitis, molluscum contagiosum, scabies, and pediculosis pubis. Additional features, such as epidemiology and transmission of the organism, are discussed when applicable. (J Am Acad Dermatol 1999;41:661-77.) LEARNING OBJECTIVE: At the conclusion of this learning activity, participants should be familiar with the clinical features, diagnosis, and treatment of sexually transmitted diseases (excluding AIDS) which have cutaneous presentations or involvement. PMID- 10534628 TI - Muir-Torre syndrome: case report of a patient with concurrent jejunal and ureteral cancer and a review of the literature. AB - BACKGROUND: Muir-Torre syndrome is a rare autosomal dominant genodermatosis, first described in 1967, characterized by the presence of sebaceous tumors and an internal malignancy in the absence of other predisposing factors. OBJECTIVE: Our purpose was to review and update published literature on Muir-Torre syndrome. METHODS: We describe a 66-year-old white man with a history of sebaceous tumors and newly diagnosed transitional cell cancer of the right ureter and adenocarcinoma of the jejunum. The literature on Muir-Torre syndrome is reviewed by means of MEDLINE search and available published reports and updated. RESULTS: Only 205 cases of Muir-Torre syndrome with 399 internal malignancies have been reported. The common presentation is the presence of sebaceous tumors along with a low-grade visceral malignancy. Sebaceous tumors appeared before the internal malignancy in 45 cases (22%), concurrently in 12 (6%), and after the internal malignancy in 114 (56%). In 33 (16%) of 205 patients, a temporal relationship was not reported. The total number of sebaceous gland carcinomas reported is 44; 17 of 44 were neoplasms of the meibomian gland. Keratoacanthomas have been noted in 48 (23%) of 205 patients. Gastrointestinal cancers are the most common internal malignancies (61%), followed by genitourinary (22%). CONCLUSION: The presence of sebaceous tumors warrants a search for an internal malignancy. In patients with Muir-Torre syndrome, regular follow-up and search for new malignancy is mandatory. Evaluation and monitoring of the family members of patients are also necessary. Patients and their families should be counseled for genetic testing. Genetic analysis of the primary tumor and skin lesions should be arranged as an added research tool if possible to better understand the disease. PMID- 10534629 TI - A noninvasive method for quantifying and distinguishing inflammatory skin reactions. AB - BACKGROUND: The ribonuclease protection assay (RPA) represents a technology that allows detection of small amounts of intact RNA. Recent progress in understanding cytokine networks in the skin suggests that measurements of cytokine mRNA levels could provide a method to distinguish various reactions such as irritant contact dermatitis and allergic contact dermatitis that can occur in the skin. OBJECTIVE: We attempted to differentiate and quantitate irritant and immunologic skin reactions by measuring mRNA levels. METHODS: We have used the technique of tape stripping human skin to remove superficial cell layers and have extracted RNA from these skin samples. This RNA was used for RPA analysis. RESULTS: By means of RPA analysis, we have demonstrated distinct cytokine profiles that appear to discriminate, for example, irritant from immunologic skin reactions. CONCLUSION: We have shown that multiple cytokine mRNA levels can be defined in these RNA samples obtained from the skin. This approach assesses not only the cytokine gene profiles, but at the same time may quantify the severity of common irritant versus allergic skin reactions. PMID- 10534630 TI - Reliability and accuracy of dermatologists' clinic-based and digital image consultations. AB - BACKGROUND: Telemedicine technology holds great promise for dermatologic health care delivery. However, the clinical outcomes of digital image consultations (teledermatology) must be compared with traditional clinic-based consultations. OBJECTIVE: Our purpose was to assess and compare the reliability and accuracy of dermatologists' diagnoses and management recommendations for clinic-based and digital image consultations. METHODS: One hundred sixty-eight lesions found among 129 patients were independently examined by 2 clinic-based dermatologists and 3 different digital image dermatologist consultants. The reliability and accuracy of the examiners' diagnoses and the reliability of their management recommendations were compared. RESULTS: Proportion agreement among clinic-based examiners for their single most likely diagnosis was 0. 54 (95% confidence interval [CI], 0.46-0.61) and was 0.92 (95% CI, 0. 88-0.96) when ratings included differential diagnoses. Digital image consultants provided diagnoses that were comparably reliable to the clinic-based examiners. Agreement on management recommendations was variable. Digital image and clinic-based consultants displayed similar diagnostic accuracy. CONCLUSION: Digital image consultations result in reliable and accurate diagnostic outcomes when compared with traditional clinic-based consultations. PMID- 10534631 TI - Lymph node metastases of cutaneous melanoma: diagnosis by B-scan and color Doppler sonography. AB - BACKGROUND: Sonography is a sensitive, noninvasive method that can be used to detect regional lymph node metastases. Color Doppler sonography (CDS) can supply further information on lymph node perfusion. OBJECTIVE: We evaluated the usefulness of CDS for differentiating between benign lymphadenopathies and lymph node metastases of cutaneous melanoma. METHODS: In a prospective study, reactive inflammatory lymph nodes (rLN) and lymph node metastases of cutaneous melanoma (mLN) were examined by sonography and CDS. Lymph node echogenicity and shape (length/depth ratio) were determined by sonography. The vascularization pattern of the lymph nodes was established with CDS. We recorded the Doppler frequency spectra at the hili of the lymph nodes and then calculated the resistance and pulsatility indices (RI, PI). RESULTS: The echogenicity of the lymph node centers had a sensitivity of 96% and a specificity of 100%. The shape differed highly significantly between the two groups (P <.001). The criterion length/depth ratio < 2 had a sensitivity of 85% and specificity of 86%. Hilus vessels could be detected in 14 of 22 rLN (64%). These vessels, however, were not present in any of the metastases. The RI and the PI in detectable lymph node vessels differed between the two groups (RI: P <.05; PI: not significant), but because of the overlap between the two groups, these indices were of no diagnostic value. In the presence of 2 or more of the following 3 criteria: length/depth ratio < 2, hypoechoic center, and the absence of hilus vessels, diagnosis of metastasis of malignant melanoma had a sensitivity of 100% and a specificity of 96%. CONCLUSION: CDS improves the diagnostic accuracy of conventional sonography. The measurement of Doppler curves in lymph node vessels and the calculation of pulsatility and resistance indices, on the other hand, is time-consuming and seems to be of no diagnostic value. PMID- 10534632 TI - Digital fluorescence photography can assess the suppressive effect of benzoyl peroxide on Propionibacterium acnes. AB - BACKGROUND: Porphyrins produced by Propionibacterium acnes exhibit an orange-red fluorescence under UVA light. The amount of fluorescence can be estimated by digital fluorescence photography. OBJECTIVE: We thought that digital fluorescence photography would be a quicker and simpler method than bacteriologic culture to demonstrate depopulation of P acnes in sebaceous follicles. We used benzoyl peroxide to bring about rapid suppression of P acnes. METHODS: Benzoyl peroxide 10% was applied twice daily for 7 days to the faces of 9 subjects. Five subjects were untreated controls. Digital fluorescence photographs of cheek and nose, and scrub samples for quantitative recovery of P acnes from the cheek were taken at baseline, day 3, day 7 (end of treatment), and day 16 (regression phase). RESULTS: The effect of benzoyl peroxide against P acnes was clearly demonstrated both by culture and by fluorescence photography after only 3 days. Image analysis of porphyrin fluorescence correlated well with the decrease in P acnes density from scrub cultures. No further decrease was observed at day 7 (end of therapy). Ten days later there was a return to baseline values, although in some subjects these remained lower. CONCLUSION: Digital fluorescence photography is a reliable, fast, and easy screening technique to demonstrate the suppressive effect of topical antibacterial agents on P acnes. PMID- 10534633 TI - Changes in hair weight and hair count in men with androgenetic alopecia, after application of 5% and 2% topical minoxidil, placebo, or no treatment. AB - Quantitative estimation of hair growth using hair weight and number was recorded for 120 weeks in 4 groups of 9 men with androgenetic alopecia. Three double-blind groups applied either 2% or 5% minoxidil solution, or vehicle. The fourth group, unblinded, received no treatment. Measurements of hair weight and number were continued for 96 weeks, when treatment (if any) was stopped, though measurements were continued for another 24 weeks. Although not compared statistically, the placebo and untreated groups behaved in a similar fashion. In contrast, the 5% and 2% minoxidil treatment groups showed a statistically significant increase in mean percentage change in interval weight from baseline compared with placebo; results for number counts were usually less significant. Over 96 weeks, topical minoxidil induced and maintained an increase in interval weight over baseline of about 30%. After treatment was stopped, hair weight and number counts for the minoxidil groups returned to about the same levels as placebo in 24 weeks. PMID- 10534634 TI - Cutaneous CD8 T cell infiltrates in advanced HIV infection. AB - BACKGROUND: Aggressive non-Hodgkin's lymphomas are common among patients infected with HIV. Although such lymphomas are mostly of the B-cell type, various cases of cutaneous T-cell lymphoma (CTCL) have also been reported. Recent reports suggest that some HIV-related lymphoproliferative conditions may not be clonal processes, but polyclonal lymphoid proliferations. OBJECTIVE: We reviewed our experience with HIV patients seen at the dermatology clinics for possible CTCL. METHODS: A retrospective study was performed to evaluate clinical, laboratory, and histologic findings of HIV-infected patients with atypical T-cell cutaneous infiltrates. RESULTS: We observed 9 patients with advanced HIV infection and a cutaneous eruption characterized by a dense infiltrate of lymphocytes resembling mycosis fungoides histopathologically, but composed of CD8(+) cells. Although clonality was not identified in any of the 6 cases tested, 3 patients had similar CD8(+) infiltrates involving lymph nodes or bone marrow. Of the 9 patients, 8 died of AIDS wasting syndrome or infections in less than 1 year. CONCLUSION: Cutaneous and systemic infiltrates with polyclonal CD8 T lymphocytes can be seen in patients with advanced HIV infection and profound CD4 lymphopenia. The clinical presentation may resemble CTCL and is associated with a poor outcome. PMID- 10534635 TI - A randomized comparison of narrow-band TL-01 phototherapy and PUVA photochemotherapy for psoriasis. AB - BACKGROUND: Although PUVA treatment of psoriasis is more effective than conventional or broad-band UVB phototherapy, two small studies have suggested that narrow-band or TL-01 phototherapy may have a therapeutic effect equal to PUVA. If confirmed, this would be of considerable importance as TL-01 therapy is likely to be considerably safer in the long term than PUVA. OBJECTIVE: The purpose of this study was to compare PUVA with narrow-band (TL-01) phototherapy in psoriasis. METHODS: We studied 100 patients with plaque-type psoriasis who were randomly allocated to twice-weekly treatment with PUVA or narrow-band UVB. RESULTS: Clearance of psoriasis was achieved in a significantly greater proportion of patients treated with PUVA (84%) than with TL-01 (63%) (P =.018), and with significantly fewer treatments (median number of treatments for clearance with PUVA, 16.7; with TL-01, 25.3; P =.001). Only 12% of those treated with TL-01 were clear of psoriasis 6 months after finishing treatment compared with 35% for PUVA (P =.002). CONCLUSION: When given twice weekly, PUVA is more effective for psoriasis than narrow-band UVB phototherapy. PMID- 10534636 TI - Bullous erysipelas: A retrospective study of 26 patients. AB - BACKGROUND: Erysipelas is a superficial form of cellulitis caused by a variety of microbes, and it responds to antibiotic treatment. During the past few years we treated several patients with a bullous form of erysipelas involving the lower legs. We believe their disease had a more protracted course than patients with nonbullous erysipelas. OBJECTIVE: We studied bullous erysipelas by conducting a retrospective analysis of 26 patients with bullous erysipelas of the legs treated by the authors during a 5-year period. METHODS: We conducted a retrospective review of the records of all patients with a diagnosis of bullous erysipelas who were treated at the Department of Dermatology, Hadassah Medical Center, Jerusalem, between the years 1992 and 1996. Data regarding patients with nonbullous erysipelas were obtained from the medical center's computerized data pool. RESULTS: A total of 26 cases of bullous erysipelas were found, comprising 22 women and 4 men whose ages ranged from 28 to 87 (mean, 58.8) years. The average hospital stay was 20.57 days (range, 12 to 46 days). The average hospital stay for patients with nonbullous erysipelas and cellulitis treated in the same department by the authors during the study period was 10.6 days (range, 2 to 54 days). CONCLUSION: Bulla formation is a complication of erysipelas, seen in our series in 5.2% of the patients (26 of 498 admissions for erysipelas and cellulitis). The course of the disease is protracted, requiring longer medical attention. PMID- 10534637 TI - Screening for malignant melanoma: A cost-effectiveness analysis. AB - BACKGROUND: Skin cancer is the most common cancer in the United States. Increasing evidence suggests that screening for malignant melanoma is effective, but its cost-effectiveness has not been determined. OBJECTIVE: We attempted to determine the effectiveness and costs of a visual screen to diagnose malignant melanoma in high-risk persons. METHODS: We developed a decision analysis comparing no skin cancer screen with a single screen by a dermatologist. Clinical outcomes included malignant melanoma, nonmelanoma skin cancer, or no skin cancer. Life expectancy and costs of care were projected on the basis of clinical findings. RESULTS: Skin cancer screening increased average discounted life expectancy from 15.0963 years to 15.0975 years. Based on the prevalence of malignant melanoma, however, this translates into an increased discounted life expectancy of 0.9231 years for each person with diagnosed melanoma. Using a cost of $30 per screen, total skin cancer-related costs for a cohort of 1 million people increased from $826 million with no screen to $861 million with screening, with an increase of 1200 years of life. This results in an incremental cost effectiveness ratio of $29,170 per year of life saved (YLS) with screening. Sensitivity analysis showed that the cost-effectiveness ratio for screening remained below $50,000/YLS if the prevalence of melanoma in the screened population was at least 0. 0009, the probability that a melanoma detected in screening was localized was at least 94.8%, or the cost of each screen was below $57. CONCLUSION: Skin cancer screening in high-risk patients is likely to be associated with a small increase in discounted life expectancy and is reasonably cost-effective compared with other cancer screening strategies. PMID- 10534639 TI - The uses of digital photography in dermatology. AB - Digital photography is a powerful tool that is transforming the specialty of dermatology by integrating patient and practice management. The fundamentals of digital imaging are discussed, and an approach to the selection of a digital camera and its associated hardware and software is provided. The applications of this technology to patient and practice management are addressed, and the ethical implications of digital tampering are also discussed. PMID- 10534638 TI - Melkersson-Rosenthal syndrome in childhood: successful management with combination steroid and minocycline therapy. AB - The Melkersson-Rosenthal syndrome consists of a triad of recurrent lip and/or face swelling, fissured tongue, and intermittent facial palsy. Onset of the symptoms may occur during childhood, and treatment of the condition is difficult. We describe two children with Melkersson-Rosenthal syndrome in whom combination treatment with prednisone and minocycline proved effective and well tolerated. PMID- 10534640 TI - Propylthiouracil hypersensitivity: report of two patients with vasculitis and review of the literature. AB - Two patients with a hypersensitivity vasculitis in association with propylthiouracil (PTU) administration are described. Although both patients presented with a cutaneous eruption, our first patient suffered severe systemic manifestations and the second patient's involvement was primarily limited to the skin. Patients with a vascular hypersensitivity reaction to PTU typically present with constitutional symptoms, acral purpuric skin lesions, and variable involvement of multiple organ systems. The reaction is treated by urgent withdrawal of PTU and implementation of supportive measures and immunosuppressive agents, as necessary. Prompt recognition of this condition and initiation of appropriate therapy lead to complete recovery in most cases. PMID- 10534641 TI - Diagnostic pearl: unmagnified diascopy for large pigmented lesions reveals features similar to those of epiluminescence microscopy. PMID- 10534642 TI - A new tissue adhesive for laceration repair in children AB - Bruns TB, Robinson BS Smith RJ, Kile DL, Davis TP, Sullivan KM, et al. J Pediatr 1998;132:1067-70. PMID- 10534643 TI - The basic principles of computed tomography and magnetic resonance imaging. PMID- 10534644 TI - Sturge-Weber syndrome. PMID- 10534645 TI - Treatment of rosacea-like demodicidosis with oral ivermectin and topical permethrin cream. AB - A 32-year-old man presented with a chronic rosacea-like dermatitis of the facial skin and the eyelids. The skin disorder had been present for 4 years and was unresponsive to multiple previous treatment attempts. Skin scrapings and a histologic examination of a biopsy specimen from the affected area revealed the presence of numerous Demodex mites. The patient was treated with oral ivermectin and subsequent topical permethrin resulting in complete and rapid clearing of the folliculitis. We believe that this case supports the view that Demodex mites may be pathogenic when they are present in large numbers. Oral treatment with 200 microg/kg ivermectin with subsequent weekly topical permethrin showed impressive treatment efficacy in a case refractory to conventional treatment. PMID- 10534646 TI - Accelerated growth of dermatofibrosarcoma protuberans during pregnancy. AB - Dermatofibrosarcoma protuberans (DFSPs) are uncommon skin tumors that have a high incidence of local recurrence even with wide surgical margin, but DFSPs rarely metastasize. Previous reports have suggested that DFSPs may enlarge more rapidly during pregnancy. We report 3 additional cases of DFSPs that showed accelerated growth during pregnancy. Immunohistochemical stains for CD34, S-100 protein, factor XIIIa, and estrogen and progesterone receptors were performed on biopsy specimens. The tumors in all 3 patients, and 4 additional DFSPs from 2 male and 2 female subjects, showed expression of progesterone receptor. As with many other stromal neoplasms, DFSPs appear to express low levels of hormone receptors, which may be one factor that accounts for their accelerated growth during pregnancy. PMID- 10534647 TI - Patients can accurately identify when they have a dermatologic condition. AB - The use of "gatekeepers" may be rational when patients do not know what specialist to see for their condition. Using representative data on outpatient practice in the United States, we tested the hypothesis that when patients believe they have a skin condition, they have a dermatologic disease. At least 95% of visits for skin symptoms are associated with dermatologic diagnoses. Patients who believe they have skin conditions do have dermatologic diseases and are able to choose the appropriate specialist. PMID- 10534648 TI - Hepatitis C virus and lichen planus: A case-control study of 340 patients. AB - Lichen planus (LP) has been associated with liver disease, particularly hepatitis C virus (HCV) infection, in several studies to date. Most of these reports, especially the larger series, were conducted in Europe. In addition, the type of LP associated with HCV was reported to be oral LP in most studies. We conducted a case-control study in a US metropolitan population known to have a low seroprevalence of HCV infection to explore the impact of geography and endemicity of HCV on the association between cutaneous LP and HCV. The study comprised 340 patients with cutaneous LP (as case ) against 577 patients with psoriasis (as control-1 ). Hepatitis antibody titers and liver function abnormalities were our main outcome measures. Prevalence of HCV antibody among 149,756 regional volunteer blood donors (as control-2 ) was also used for comparison. Twelve (55%) of 22 patients with LP tested for HCV were antibody positive. This was significantly higher than 25% of 40 psoriasis patients tested for HCV antibody (P =.04) or than 0.17% of blood donors tested for HCV antibody (P < 10(-5)). Thus a small but significant percentage of US patients with cutaneous LP had HCV antibody. Because LP may be the first presentation of HCV infection, it is important for clinicians to actively look for such infection in patients with LP. PMID- 10534649 TI - Lafora disease: diagnosis by skin biopsy. AB - Lafora disease is a fatal neurometabolic disorder characterized by progressive myoclonic epilepsy. Diagnosis relies upon the discovery of specific inclusion bodies in any of several organs. Dermatologists and dermatopathologists should be familiar with this condition because axillary skin biopsy is useful to diagnose this disorder. We present a case of Lafora disease diagnosed by axillary skin biopsy and review the condition's clinical, histologic, and ultrastructural features. PMID- 10534650 TI - Papular-purpuric "gloves and socks" syndrome: polymerase chain reaction demonstration of parvovirus B19 DNA in cutaneous lesions and sera. AB - We report a typical case of papular-purpuric "gloves and socks" syndrome (PPGSS) in which primary infection by parvovirus B19 was demonstrated by seroconversion to this virus; parvovirus B19 DNA was also identified by polymerase chain reaction (PCR) methods in the sera of the patient and in the cutaneous biopsy specimen, both taken 4 days after the onset of clinical manifestations. To our knowledge, this is the fourth published case in which parvovirus B19 DNA has been recovered from the skin by PCR. Serologic studies and PCR investigations in cutaneous biopsy for other viruses including herpes simplex virus types 1 and 2, varicella zoster virus, Epstein-Barr virus, cytomegalovirus, and human herpesvirus 6, 7, and 8 were negative. Clinically, our case presented some additional features, which have not been previously described in cases of PPGSS, namely dysuria with vulvar edema and erythema, and unilateral petechial rash on the breast. The histopathologic findings of our case were nonspecific and consisted of an interface dermatitis with slight vacuolar degeneration at the dermoepidermal junction and a superficial perivascular inflammatory infiltrate mostly composed of lymphocytes, with numerous extravasated erythrocytes. We review the cases of PPGSS published in the literature with respect to the different viruses that have been proposed as etiologic agents and conclude that acute infection by parvovirus B19 is the only one that has been adequately proved. PMID- 10534651 TI - The Muir-Torre syndrome in kindreds with hereditary nonpolyposis colorectal cancer (Lynch syndrome): A classic obligation in preventive medicine. PMID- 10534653 TI - A 30-year-old woman with melanoma metastases and a positive pregnancy test. PMID- 10534655 TI - Self-assessment examination of the american academy of dermatology (Identification no. 899-211) AB - Learning objectives: At the conclusion of this self-assessment learning activity, physician participants should be able to assess their own diagnostic and patient management skills with respect to those of their colleagues in the field, use the results of the self-assessment to help determine personal learning needs that can be addressed through subsequent CME involvement, and enhance their ability to comply with the requirements for certification in the specialty of dermatology.Instructions for Category I CME credit appear in the front advertising section. See last page of Contents for page number. INSTRUCTIONS: In answering each question, refer to the specific directions provided. Because it is so often necessary to provide information in questions occurring later in a series that give away answers to earlier questions, please answer the questions in each series in sequence. PMID- 10534654 TI - Narrow-band UVB-associated lesional blisters in pityriasis rubra pilaris. PMID- 10534652 TI - p53, p54, PUVA, and counting. PMID- 10534656 TI - Answers to self-assessment examination of the journal of the american academy of Dermatology:Identification no. 899-211 PMID- 10534657 TI - Necrobiotic cutaneous T-cell lymphoma. AB - We report 3 patients with granulomatous cutaneous T-cell lymphoma (CTCL) who showed necrobiosis histologically with palisading granulomas. Although granulomatous change may be present in up to 4% of cases of CTCL, necrobiosis is rare. Misdiagnosis may occur if epidermotropism is minimal or if atypical cells are masked by the granulomatous infiltrate. T-cell receptor gene analysis confirmed the presence of clonal T-cell populations in lesional skin from all 3 cases. PMID- 10534658 TI - Acute generalized exanthematous pustulosis induced by chromium picolinate. AB - A case of acute generalized exanthematous pustulosis (AGEP) due to chromium picolinate is described. This supplemental form of chromium has received a great deal of interest recently because of its possible beneficial effects on both muscle strength and body composition. There have been no previous reports to our knowledge of adverse cutaneous reactions to this agent. Various aspects of AGEP are reviewed. PMID- 10534659 TI - Adult onset verrucous epidermal nevus. AB - Epidermal nevi are hamartomas of the skin and have multiple clinical variants, including a verrucous type. Usually verrucous epidermal nevi present at an early age, are found on the lower extremities, have a female predominance, and are refractory to treatment. Only very rarely do verrucous epidermal nevi have an adult onset. We describe a 60-year-old woman with a 5-year history of multiple verrucous plaques on her face, scalp, and neck. Histologic examination revealed the characteristic findings of verrucous epidermal nevus. This is the oldest patient reported with an adult onset verrucous epidermal nevus. PMID- 10534660 TI - Glipizide-induced pigmented purpuric dermatosis. AB - Pigmented purpuric dermatosis can occasionally be caused by various medications. No reported cases of oral hypoglycemic agents causing pigmented purpuric dermatosis exist. We report a case of glipizide-induced pigmented dermatosis. PMID- 10534661 TI - Sudden onset of disseminated porokeratosis of Mibelli in a renal transplant patient. AB - Porokeratosis is a disorder of epidermal keratinization of uncertain cause. Five clinical variants of porokeratosis have been described. These include porokeratosis of Mibelli, punctate porokeratosis, linear porokeratosis, porokeratosis palmaris plantaris et disseminata, and disseminated superficial porokeratosis. Disseminated superficial porokeratosis and single plaque porokeratosis of Mibelli have each been documented to occur in association with immunosuppression. To our knowledge, only 5 cases of disseminated porokeratosis of Mibelli in transplant recipients have been reported. We present a patient who developed explosive onset of disseminated porokeratosis of Mibelli shortly after renal transplantation. It is important to differentiate this unusual variety of porokeratosis from other cutaneous manifestations in transplant patients so that appropriate therapy can be instituted. PMID- 10534662 TI - Unusual eruption as a presenting symptom of cat scratch disease. AB - Cat scratch disease (CSD) is a common infectious cause of subacute regional lymphadenopathy. Bartonella henselae is the principal etiologic agent. About 10% of CSD patients experience atypical manifestations, including rashes. The most common cutaneous manifestation of CSD is a papule at the inoculation site. We report a case of CSD presenting with an eruption on the upper trunk, reminiscent of Sweet's syndrome, accompanied by lymphadenopathy, arthralgia, and fever. Response to systemic corticosteroids was remarkable. Histopathologic findings refuted the diagnosis of Sweet's syndrome. Identification of anti-B henselae antibodies and B henselae DNA in the affected lymph node confirmed the diagnosis of CSD. This is a first report of extensive papuloedematous eruption as a cutaneous manifestation of CSD. Accurate diagnosis is possible due to the availability of serological tests and DNA amplification techniques. PMID- 10534663 TI - Acute infantile hemorrhagic oedema. AB - Acute infantile hemorrhagic oedema (AIHO) was first described in 1913 but, despite frequent reports in the European literature, it is not well recognized in the English language literature. It is considered by many to be a variant of Henoch Schonlein Purpura (HSP) because of similarities in cause and histopathology. However, because of the benign nature of this condition and frequent absence of IgA associated with HSP, it may be sensible to consider this as a distinct variety of cutaneous small vessel vasculitis (CSVV). We report this case to highlight the condition and emphasize its benign nature. PMID- 10534664 TI - Ecthyma gangrenosum in an AIDS patient with normal neutrophil count. AB - Ecthyma gangrenosum is the cutaneous manifestation of Pseudomonas aeruginosa septicemia, typically affecting immunosuppressed patients, particularly those with neutropenia. Association with HIV disease has been rarely reported. We describe an unusual presentation of solitary ecthyma gangrenosum on the face of a non-neutropenic patient with AIDS. PMID- 10534665 TI - Livedo reticularis revealing a latent infective endocarditis due to Coxiella burnetti. AB - We report the first case of livedo reticularis revealing a latent infective endocarditis due to Coxiella burnetti. The patient, a 54-year-old woman, also had chronic thrombocytopenia and mixed cryoglobulinemia. Chronic Q fever was confirmed by serodiagnosis and livedo regressed totally with doxycycline and hydroxychloroquine. PMID- 10534666 TI - Three cases of pustulotic arthro-osteitis associated with episcleritis. AB - Three cases of pustulotic arthro-osteitis (PAO) associated with episcleritis were described. In each patient, the episcleritis developed more than 10 years after the onset of PAO. These episcleritis were treated with topical corticosteroids. PAO is classified as a member of the seronegative spondylarthritis group of diseases. Though complications of seronegative spondylarthritis include uveitis and episcleritis. PAO associated with episcleritis was not reported. Episcleritis should be considered as a complication of PAO. PMID- 10534667 TI - Unusual cutaneous lesions in two patients with visceral leishmaniasis and HIV infection. AB - Two HIV infected patients with visceral leishmaniasis and unusual cutaneous lesions are described. The first patient developed linear brown macules containing Leishmania parasites on the fingers and palms of the hands. This patient never received highly active antiretroviral treatment and the visceral leishmaniasis could not be cured even with liposomal amphotericin. In the second patient, Leishmania parasites were present in a skin biopsy of a fibrous histiocytoma. After completing visceral leishmaniasis treatment, a discrete elevation of one of his tattoos was seen. A biopsy specimen of this tattoo revealed Leishmania amastigotes. In this patient the visceral leishmaniasis was finally cured with meglumine antimoniate, followed by pentacarinat isothianate as maintenance therapy in conjunction with highly active antiretroviral treatment. PMID- 10534668 TI - Pustular eruption induced by olanzapine, a novel antipsychotic agent. AB - Pustular eruptions caused by medications are unusual. Antipsychotic agents have not been previously reported to cause pustular reactions. We report a case of olanzapine-induced pustular drug eruption. PMID- 10534669 TI - Localized primary cutaneous Mycobacterium kansasii infection in an immunocompromised patient. AB - Mycobacterium Kansasii is a rare primary cutaneous pathogen, most commonly affecting persons exposed to contaminated water, particularly after local trauma. Most patients who present with localized primary cutaneous M kansasii infection are immunocompetent, whereas the majority of patients with disseminated or pulmonary infection are immunocompromised. We describe a primary cutaneous M kansasii infection in an iatrogenically immunosuppressed patient. PMID- 10534670 TI - Fibrous hamartoma of infancy. AB - Fibrous harmartoma of infancy is a benign soft tissue tumor that occurs in the first few years of life. Although the lesion is not distinctive clinically, it has a characteristic microscopic appearance. Only 12 cases of fibrous hamartoma of infancy have been reported in the dermatology literature. PMID- 10534671 TI - Cutaneous inoculation tuberculosis in a child. AB - Cutaneous tuberculosis remains a rare entity in the United States. We describe a case of cutaneous tuberculosis in a child. PMID- 10534672 TI - Dermatofibrosarcoma protuberans of the oral cavity. AB - Dermatofibrosarcoma protuberans (DFSP) is a low grade, malignant spindle cell tumor with an infiltrative growth pattern and a high rate of local recurrence. This tumor's cell of origin is controversial. DFSP usually presents in adult life and is most frequently located on the trunk and proximal extremities. Although 10% to 15% of cases involve the head and neck, this tumor has not been previously described in the oral cavity. PMID- 10534673 TI - Isolated extragenital bowenoid papulosis of the neck. AB - We report a case of extragenital bowenoid papulosis (BP) in a healthy immunocompetent 42-year-old man. The lesions occurred on the anterolateral aspects of the neck and were not associated with genital, oral, or periungual lesions. Lesional skin tested positive with the Digene hybrid capture system cocktail assay that identifies infection with a mixture of high to intermediate oncogenic human papillomavirus (HPV) types, including types 16, 18, 31, 33, 35, 45, 51, 52, and 56. This cocktail assay identifies infection with HPV types typically associated with high-grade squamous intraepithelial lesions and invasive carcinoma. This case represents the sixth case of isolated cutaneous BP occurring a significant distance from the genital region. PMID- 10534674 TI - Acute periorbital mucinosis in discoid lupus erythematosus. AB - Periorbital edema associated with lupus erythematosus is not frequently reported. To our knowledge, periorbital edema from increased dermal mucin has not been reported with any form of lupus. We present a patient with discoid lupus exhibiting periorbital edema from massive mucinosis. PMID- 10534675 TI - Paraneoplastic amyopathic dermatomyositis associated with breast cancer recurrence. AB - Amyopathic dermatomyositis, also known as dermatomyositis sine myositis, describes patients with the typical cutaneous features of dermatomyositis but without evidence of inflammatory myopathy. There have been 3 reported cases of amyopathic dermatomyositis associated with breast cancer in patients with no prior malignancy. We describe the first case of paraneoplastic amyopathic dermatomyositis associated with breast cancer recurrence. PMID- 10534676 TI - Exuberant granulation tissue associated with chronic graft-versus-host disease after transplantation of peripheral blood progenitor cells. AB - Angiomatous lesions appearing in chronic graft versus host disease (cGVHD) has been reported previously. We report a case of a patient in whom cutaneous exuberant granulation tissue developed in the course of a cGVHD after allo peripheral blood progenitor cells transplantation. PMID- 10534677 TI - Durable loss of a malignant T-cell clone in a stage IV cutaneous T-cell lymphoma patient treated with high-dose interferon and photopheresis. AB - Stage IV cutaneous T-cell lymphoma (CTCL) has a notoriously poor prognosis and many treatment options exist. We describe the successful treatment of a case of stage IV CTCL with combination photopheresis and high-dose interferon alfa (IFNalpha). The patient was treated with combination therapy of monthly photopheresis and daily doses of IFNalpha up to 36 MU. Response to therapy was followed by clinical observation and Southern blot analysis for the detection of a malignant clone. The findings of this case were compared with others using a computer-based literature review. A complete clinical response lasting 64 months was achieved. Clinical relapse was preceded by an increase in the CD4/CD8 ratio and by reappearance of the T-cell receptor gene rearrangement. Combined photopheresis and high-dose IFNalpha led to a durable and sustained complete response in stage IV CTCL. CD4/CD8 ratios and T-cell gene rearrangements may be helpful in patient management. PMID- 10534678 TI - Primary cutaneous B-cell lymphoma with a dermatomal distribution. AB - Primary cutaneous B-cell lymphomas (CBCLs) are indolent lymphoproliferative skin disorders with the only exception of the variant arising on the legs, which is characterized by a more aggressive clinical course. We report a case of CBCL unusual in that it presented with a dermatome-like distribution and showed early peripheral lymph node involvement. The other noteworthy finding is that rearrangement of the bcl-2 gene was detected by polymerase chain reaction in skin tissue samples. The bcl-2 rearrangement, which is usually absent in the classic CBCLs, is likely to play a role in the aggressive clinical behavior of the tumor, although further studies are necessary to confirm it. The hypothesis of a virus causing the dermatomal presentation is intriguing but there is no proof for such speculation at this point. PMID- 10534679 TI - Tamoxifen-induced remission of psoriasis. AB - Psoriasis vulgaris may be worsened or precipitated by numerous factors, including hormonal influences. Several lines of evidence suggest that female sex hormones such as estrogen and progesterone affect this condition. We describe a patient whose psoriasis responded favorably to administration of the antiestrogen compound tamoxifen. PMID- 10534680 TI - Adult-onset angiofibroma and multiple endocrine neoplasia type I. AB - Multiple endocrine neoplasia type 1 (MEN 1) predisposes affected persons to neoplasms of the parathyroid glands, the endocrine pancreas, the anterior pituitary, and the duodenum. We report the first case of adult-onset multiple angiofibromas of the central face associated with MEN 1. Seven siblings also developed adult-onset angiofibromas, none with evidence of tuberous sclerosis. Basic fibroblast growth factor (BFGF) is elevated in many patients with MEN 1 and may play a pathogenetic role. PMID- 10534681 TI - Successful treatment of Kimura's disease with cyclosporine. AB - We report the case of a 29-year-old Japanese woman presenting with recurring Kimura's disease. We began treatment with cyclosporine within 7 days, the nodular lesion had almost cleared. The cyclosporine dose was then gradually reduced and discontinued after 6 months. The patient was reassessed 18 months after the cessation of treatment and there was no evidence of recurrence of the disease. We speculate that the effects of cyclosporine on T helper-2 cells improves Kimura's disease. PMID- 10534682 TI - 1998 AATS Scientific Achievement Award recipient-Norman E. Shumway, MD, PHD. American Association for Thoracic Surgery. PMID- 10534683 TI - Blood versus crystalloid cardioplegia for myocardial protection of donor hearts during transplantation: A prospective, randomized clinical trial. AB - OBJECTIVE: To assess the safety and efficacy of myocardial protection of the donor heart during transplantation with the use of blood cardioplegia, a prospective randomized clinical trial was undertaken between January 1997 and March 1998. METHODS: Forty-seven consecutive patients were assigned either to crystalloid (27 patients; group 1) or blood cardioplegia (20 patients; group 2). Comparison of recipient age (54 +/- 11 years vs 55 +/- 7 years; P =. 9), sex (89% vs 90% male patients; P =.9), diagnosis (63% vs 65% dilated cardiomyopathy; P =.8), elevated pulmonary vascular resistance (30% vs 30%; P =.9), prior cardiac operations (22% vs 30%; P =.5), need for urgent heart transplantation (7% vs 20%; P =. 2), donor age (32 +/- 11 years vs 31 +/- 13 years; P =.7), cause of death (33% vs 40% vascular; P =.5), and global myocardial ischemia (176 +/- 51 minutes vs 180 +/- 58 minutes; P =.5) showed no difference. Hemodynamically unstable donors (15% vs 45%; P =.02) were more prevalent in group 2. RESULTS: Operative mortality rates (4% vs 5%; P =.8), high-dose inotropic support (41% vs 30%; P = 0.6), and postoperative mechanical assistance (11% vs 10%; P = 0.9) were comparable in the 2 groups. Prevalence of acute right heart failure (27% vs 0; P =.02) and of temporary complete atrioventricular block (52% vs 20%; P =.02) were greater in group 1. Spontaneous sinus rhythm recovery was more prevalent in group 2 (11% vs 40%; P =.02). Higher peak creatine kinase (1429 +/- 725 u/L vs 868 +/- 466 u/L; P =.01) and creatine kinase MB (144 +/- 90 u/L vs 102 +/- 59 u/L; P =. 06) levels suggested more severe ischemic injury in group I. CONCLUSION: Use of blood cardioplegia was associated with a lower prevalence of right heart failure, cardiac rhythm dysfunction, and laboratory evidence of ischemia. PMID- 10534684 TI - Lung transplantation in very young infants. AB - INTRODUCTION: Established successes with adult lung transplantation have laid the foundation for extension of this therapeutic modality to infants and children dying of end-stage pulmonary disease. The purpose of this report is to convey our experience with 19 infants undergoing lung transplantation before the age of 6 months. METHODS: Six patients with predominantly pulmonary vascular disease and 13 patients with primarily pulmonary parenchymal disease have undergone bilateral sequential lung transplantation at our institution since 1990. Mean age at transplant was 104 +/- 44 days, and mean weight was 4.9 +/- 1.6 kg. RESULTS: Although early mortality (32%, 6/19) was higher than that previously reported for older pediatric age groups, long-term survival was similar (44% at a maximum follow-up of 6 years). Although anastomotic complications and infections occurred at a rate approximating that seen in older pediatric age groups, episodes of acute rejection appear to occur with decreased frequency. Similarly, at a mean follow-up of 3 years, only 2 (15%) of 13 long-term survivors have evidence of bronchiolitis obliterans. The functional residual capacity, as measured on infant pulmonary function tests, has gradually increased as the children have grown, suggesting that lung growth is occurring. CONCLUSIONS: Bilateral lung transplantation is a viable alternative in infants dying of end-stage pulmonary disease. Efforts directed toward avoiding the complications that lead to early posttransplant mortality combined with the seemingly lower incidence of early and late rejection may provide long-term results better than those in other age groups. PMID- 10534685 TI - Characterization of a pig-to-goat orthotopic lung xenotransplantation model to study beyond hyperacute rejection. AB - BACKGROUND: A pig-to-goat orthotopic lung xenograft model was developed to test whether depletion of goat xenoreactive antibodies against pig red blood cells would prolong pig lung xenograft survival. METHODS: Adult goats with anti-pig xenoreactive antibodies underwent left pneumonectomy followed by orthotopic transplantation of pig left lung (group 1) or immunodepletion of their xenoreactive antibodies by extracorporeal right pig lung perfusion before transplantation without (group 2) or with (group 3) complete clampage of the right pulmonary artery. In group 4, goat left lungs were orthotopically transplanted into pigs and served as negative controls (pig serum does not have anti-goat xenoreactive antibodies). Each study group included 5 animals. Immunosuppression in surviving recipients included cyclosporine and azathioprine. RESULTS: Group 1 recipients died 7 +/- 3 hours after xenograft reimplantation of severe pulmonary hypertension and dysfunction and vasogenic shock, with little evidence of histologic xenograft injury. Group 2 xenografts had a stable circulatory and respiratory function on reperfusion and survived 9 +/- 4 days. Group 3 animals also tolerated complete occlusion of the right pulmonary artery, and xenografts assured the total respiratory support for 4 +/- 1 days. After immunodepletion, goat serum showed no detectable titers of xenoreactive antibodies, which began to reappear by postoperative day 2, where xenografts showed histologic stigmata of acute (humoral and cellular-mediated) rejection that evolved to a complete xenograft necrose at death. Group 4 xenografts showed scattered features of acute rejection 5 +/- 1 days after the operation. CONCLUSIONS: Pig left lung xenografts can provide prolonged and complete respiratory support after depletion of goat xenoreactive antibodies, but they ultimately necrose once recipient xenoreactive antibodies return to pretransplantation values. PMID- 10534686 TI - Impact of small prosthetic valve size on operative mortality in elderly patients after aortic valve replacement for aortic stenosis: does gender matter? AB - OBJECTIVE: Ideal management of the elderly patient with a small aortic root remains controversial. This retrospective analysis was performed to determine whether small prosthetic valve size is related to outcome in patients 70 years of age or older undergoing aortic valve replacement for aortic stenosis. METHODS: Between December 1991 and July 1998, 366 patients 70 years of age or older (median age 77 years, range 73-81 years, 49% male) underwent isolated aortic valve replacement or aortic valve replacement with coronary bypass grafting with standard Carpentier-Edwards bovine pericardial valves (Baxter Healthcare Corp, Edwards Division, Santa Ana, Calif) (n = 277; 76%) or St Jude Medical mechanical valves (St Jude Medical, Inc, St Paul, Minn) (n = 89; 24%). Propensity scoring and multivariable regression models were used to evaluate the risks associated with implantation of 19-mm valves. RESULTS: Operative mortality was 16.7% (17/102) in patients who received 19-mm valves and 3% (8/264) among those receiving >/=21-mm valves (P /=21-mm valves was 6.4 (95% CI 2.7, 15.4; P 30); preoperative serum albumin was used to quantify nutritional status and underlying disease. Data were gathered between 1993 and 1997 from 5168 consecutive patients undergoing coronary artery bypass or valve operations, or both. RESULTS: No significant correlations were observed between body mass index and preoperative albumin levels. Low body mass index (<20) and low albumin level (<2.5 g/dL) were each independently associated with increased mortality after cardiopulmonary bypass (P T)/1766(C-->G) and 1766(C-->T)/1768(T-->A) in case 1 and 2, respectively. In the BCP from patient 1, a putative new binding site for the transcription factor hepatocyte nuclear factor 3 (HNF3) was generated. The functional analyses of the mutant showed a 2.8-fold increase of core promoter activity, whereas the BCP variant of patient 2 was also identified to result in enhanced promoter activity. The alignment of full-length genomes from child 1 to the reference sequence showed 61 nucleotide substitutions. Furthermore, the time of reactivations from child 1 was always accompanied by selection of a precore mutation at nucleotide position 1899. In liver tissue of patient 1 before development of hepatocellular carcinoma only free viral sequences were found, whereas a single site integration of HBV was detected in hepatocytes after activation of carcinogenesis. Specific mutations in the HBV BCP of the two patients that are rarely present in chronic carriers were identified to increase the core promoter activity possibly by altering transcription factor binding, suggesting that these variants may be involved in the pathogenesis of frequent HBV reactivations. PMID- 10534722 TI - A novel hepatitis B virus variant S 129 (Gln-->Leu): lack of correlation between antigenicity and immunogenicity. AB - A point mutation has been detected in the "a" determinant of hepatitis B surface antigen (HBsAg) in an infant immunised with hepatitis B vaccine after exposure to hepatitis B virus (HBV). This A-to-T point mutation at nucleotide 540 resulted in a glutamine-to-leucine substitution at amino acid residue 129 (129L). The S gene fragment (nucleotide 58-1072) of this isolate was cloned and used to substitute the wild-type S gene in a plasmid (p3.8II), containing 1.2 copy of full-length HBV genome with expression and replication efficiency. This plasmid p3.8II-129L was used to transfect HepG2 cells. HBsAg expressed by p3.8II-129L showed higher binding efficiency compared with the original plasmid containing the wild-type clone. A panel of 24 anti-HBs monoclonal antibodies (MAbs) was used to characterise the binding efficiency of HBsAg expressed by p3.8II-129L. Eighteen showed higher binding to the antigen, whereas HBsAg expressed by p3.8II-145R gave a consistently lower absorbance or were negative. Surprisingly, when the immunogenicity of plasmid constructs was used for DNA immunisation in Balb/c mice, the anti-HBs response induced by p3.8II-129L was significantly lower than that of the wild-type p3.8II. The lack of correlation between the antigenicity profile (binding of expressed HBsAg to anti-HBs in vitro), and the immunogenicity (induction of anti-HBs by plasmid DNA in vivo) of HBsAg with leucine substitution at position 129 indicates that biological characteristics other than the binding efficiency of HBsAg to anti-HBs could occur in HBsAg variants. These different aspects of the biological characteristics of S mutants merit further investigation. PMID- 10534723 TI - Clinical implications of coinfection with a novel DNA virus (TTV) in hepatitis C virus carriers on maintenance hemodialysis. AB - A novel hepatitis-associated DNA virus, designated as transfusion-transmitted virus (TTV), was identified recently. We investigated the frequency of TTV viremia in hepatitis C virus (HCV) carriers on maintenance hemodialysis to determine whether TTV coinfection has any clinical relevance. The subjects were 50 hemodialysis patients who had been followed over 4 years after diagnosis of HCV infection. Stored serum samples derived from each patient every 12th month after enrollment were subjected to polymerase chain reaction to amplify TTV DNA and HCV RNA. At enrollment, TTV viremia was detected in 24 (48%) HCV-positive patients irrespective of the number of previous blood transfusions and the duration of hemodialysis. The presence of TTV viremia had no relation to serum alanine aminotransferase (ALT) levels, HCV viremic levels or HCV genotypes. After enrollment, HCV infection persisted in all patients over the 4-year follow-up period, whereas spontaneous resolution of TTV infection was observed in 7 (29%) of the 24 TTV viremic cases (annual rate 7.3%, 95% confidence interval [CI] 0.8 25.5%). Evidence for TTV infection was found in 4 (15%) of the 26 TTV nonviremic patients (annual incidence 3.9%, 95% CI 0.1-19. 6%). The relationship between the ALT profile and TTV infection during follow up was not evident. Active TTV coinfection occurs frequently in HCV carriers undergoing hemodialysis but exerts no biochemical or virological influence on the underlying hepatitis C. Lack of disease association and the frequent spontaneous resolution of infection suggest that the clinical significance of TTV infection remains unclear. PMID- 10534724 TI - Artificial NS4 mosaic antigen of hepatitis C virus. AB - An artificial antigen composed of 17 small antigenic regions derived from the NS4 protein of hepatitis C virus (HCV) genotypes 1 through 5 was designed and constructed. Eleven antigenic regions were derived from the 5-1-1 region, and 6 others were derived from the C-terminus of the NS4-protein of different genotypes. The gene encoding for this artificial antigen was assembled from synthetic oligonucleotides by a new approach designated as restriction enzyme assisted ligation (REAL). The full-length synthetic gene was expressed in Escherichia coli as a fusion protein with glutathione S-transferase. By the use of site-specific antibodies raised against synthetic peptides, it was shown that all regions for which sequence-specific antibodies were obtained were accessible to antibody binding. The diagnostic relevance of the NS4 artificial antigen was demonstrated by testing this antigen with 4 HCV seroconversion panels and a panel of previously tested and stored serum specimens. The artificial antigen was found to specifically detect anti-NS4 antibodies in a number of specimens that were previously found to be anti-NS4 negative. Furthermore, this antigen detected anti NS4 activity earlier in 2 of 4 seroconversion panels than did the antigen used in a commercially available supplemental assay. Equally important is the observation that the artificial NS4 antigen demonstrated equivalent anti-NS4 immunoreactivity with serum specimens obtained from patients infected with different HCV genotypes, whereas the NS4 recombinant protein derived from genotype 1, used in the commercial supplemental test, was less immunoreactive with serum specimens containing HCV genotypes 2, 3, and 4. Collectively, these data support the significant diagnostic potential of the NS4 mosaic antigen. The strategy employed in this study may be applied to the design and construction of other artificial antigens with improved diagnostically pertinent properties. J. Med. Virol. 59:437 450 1999. PMID- 10534725 TI - Anti-HCV RIBA/LiaTek reactivity and HCV genotype in EIA-negative patients with viremia. AB - Individuals infected with hepatitis C virus (HCV) usually produce anti-HCV antibodies detectable by enzyme immunoassay (EIA); however, in certain viremic cases this antibody does not appear. To investigate whether anti-HCV in these cases is detectable by Western blot (WB), 38 HCV RNA positive/anti-HCV EIA negative sera were tested by RIBA 3.0 or LiaTek III. The HCV genotypes (INNO LiPA) were analyzed to determine whether the variance in these genotypes can be the reason for the late, weak antibody production or its absence. As the control group, 282 EIA-positive/HCV RNA-positive patients were examined. A single band reactivity of various intensities by RIBA or LiaTek was observed in 16/38 EIA negative sera. Positive results with NS3 were detected in 4 sera and weak positive (+/-) with core, NS3, and NS5 in 5, 6, and 1 sera, respectively. In 3 cases with anti-NS3, the seroreversion was observed in follow-up. The distribution of genotypes in anti-HCV-negative versus anti-HCV-positive groups was: 1b alone, 50.0% vs. 78.0%; 3a alone, 13.2% vs. 15.6%; and mixed (1b+3a), 36.8% vs. 5.0%, respectively. The follow-up studies showed that viremia was lost spontaneously in 12/35 patients. In some patients infected with two genotypes, the spontaneous loss of the 3a genotype was observed. The study showed that WB tests are useful for serological confirmation of HCV infection in some EIA negative/HCV RNA-positive patients but, because seroreversion may occur, sequential sera samples should be tested. No unusual HCV genotype was detected in anti-HCV-negative/HCV RNA-positive cases, but the frequency of mixed infection with the 1b+3a genotypes in this group was found to be higher than that in anti HCV-positive hepatitis patients. PMID- 10534726 TI - Subcellular localisation of NS3 in HCV-infected hepatocytes. AB - Hepatitis C virus (HCV) NS3 is a multifunctional protein with both protease and helicase activities and has been shown to interact with host cell proteins. It is shown that NS3 is present in the hepatocytes from patients with chronic HCV infection by using anti-NS3 antisera. NS3 is detectable in approximately 4% of the hepatocytes from these patients. In most infected cells, NS3 is present in the cytoplasm; however, in a minority of HCV-infected cells, both the cytoplasm and the nucleus or the nucleus on its own are positive for NS3. The presence of NS3 in the nuclei of hepatocytes in chronically infected patients indicates that the protein may play a role other than in virus replication, such as in persistence of HCV infection. PMID- 10534727 TI - Hepatitis B vaccination in patients with chronic hepatitis C. AB - The aim of the study was to evaluate the safety, immunogenicity, and possible therapeutic effect of hepatitis B vaccine in patients with chronic hepatitis C. The subjects studied included three groups: group I, 26 patients with chronic hepatitis C who were susceptible to hepatitis B virus infection; group II, 35 healthy subjects who were susceptible to both hepatitis B and hepatitis C virus infection; and group III, 30 patients with chronic hepatitis C receiving no hepatitis B vaccination as controls. Three 20 microg/dose of recombinant hepatitis B vaccines were given to subjects of groups I and II in months 0, 1, and 6. Blood samples from the subjects were collected before and 1 month after each dose of vaccination for serological testing. The subjects of groups I and II had similar antibody to hepatitis B surface antigen (anti-HBs) response rates after the first (30.8% vs. 17.1%), second (61.5% vs. 60.0%), and third (88.5% vs. 91.4%) doses of vaccination. Also, their geometric mean titers of anti-HBs did not differ much when vaccination completed in 7 months (360 vs. 581 mIU/ml). During vaccination period, patients with chronic hepatitis C demonstrated no significant change of serum cytokines and HCV RNA levels, but significantly lowered ALT levels after three doses of vaccination. Hepatitis B vaccination is safe and immunogenic in patients with chronic hepatitis C. It did not significantly affect their levels of HCV RNA, but tended to lower ALT levels. PMID- 10534728 TI - Cervical shedding of cytomegalovirus in human immunodeficiency virus type 1 infected women. AB - Cervical shedding of cytomegalovirus (CMV) is important in transmission of CMV to exposed sexual partners and neonates. We evaluated prevalence and correlates of CMV DNA shedding in cervical secretions from a large cohort of HIV-1-seropositive women. Using polymerase chain reaction (PCR) assays, CMV DNA was detected in 183 (59%) cervical swab samples from 311 women. Cervical shedding of CMV DNA was significantly associated with shedding of HIV-1 DNA (odds ratio 1.8; 95% confidence interval 1.1-2.8). CMV shedding was also more frequent in women with Neisseria gonorrhoeae and Trichomonas vaginalis infections, but these associations were not statistically significant. Cervical shedding of CMV in HIV 1-infected women is very frequent and may reflect higher risk of transmission to sexual partners and neonates than previously appreciated. PMID- 10534729 TI - Prevalence of long-term BK and JC excretion in HIV-infected adults and lack of correlation with serological markers. AB - The natural history of polyomavirus infection, and sensitivity of diagnostic assays remain unclear. A stratified group of 94 human immunodeficiency virus (HIV)-infected patients was studied for both virological and serological markers of active infection with both JC virus and BK virus. JC DNA was detected in the urine of 18 of 81 (22%) patients and BK DNA in 30 (37%) patients. Whilst patients with a low CD(4) cell count (P =.009), CD(4)/CD(8) ratio (P =.031) and beta2M concentration (P =.042) were significantly more likely to be excreting BK, JC excretion did not correlate with any of the immunological markers measured. Furthermore, when all the immunological factors were taken into account, there was no association between either BK or JC excretion and age of the patient (P =.149 for BK, P = 0.891 for JC). BK IgM antibody was detected in only 3 of 30 (10%) BK excretors. JC IgM was detected in 5 of 18 (27. 7%) JC excretors but also in 11 of 63 (17.5%) patients without demonstrable JC excretion. Therefore IgM was a very poor indicator of viruria. One year follow-up on a subset of patients showed that both DNA detection in urine and IgM antibody remain stable over many months despite falling CD(4) cell counts, and would indicate that events leading to enhanced viral production probably occur early after HIV infection. Replication of JC virus in the brain leading to the onset of progressive multifocal leukoencephalopathy (PML) could not be predicted using any of the markers studied. PMID- 10534730 TI - Emergence and genetic evolution of HIV-1 variants with mutations conferring resistance to multiple reverse transcriptase and protease inhibitors. AB - The emergence of genotypic resistance in protease and reverse transcriptase (RT) gene regions was longitudinally evaluated in plasma samples from a group of 12 HIV-1-infected patients treated with different combination of antiretroviral therapies and selected on the basis of their clinical failure. Complex mutational patterns in the reverse transcriptase gene were observed. In particular, combinations of AZT (41L, 67N, 70R, 210W, and 219Q/E) and 3TC (184M) were seen in 10 patients. Two patients presented codon 151 multinucleoside analogue resistance (MNR). Additionally, seven patients harbored RT nonnucleoside analogue-related resistance substitutions (98G, 103N, and 181C). Multiple protease-selected mutations were found in each patient with an average of six substitutions per patient, with 10I/F/V, 63P, 71V, 82A/T, 84V, and 90M being the most prevalent substitutions. Overall, these results showed that for most patients virological failure was coupled with detectable genotypic resistance. Furthermore, most patients exhibited genotypic resistance to almost all available anti-HIV-1 drugs. The high viral loads found in most patients at the end of the study suggest that the replication of these multidrug resistant viruses are not severely compromised. Phylogenetic analysis of these pol sequences revealed that a specific HIV-1 genotype prone to develop multidrug resistance was not found. PMID- 10534731 TI - Presence of human herpesviruses 6, 7, and 8 DNA sequences in normal brain tissue. AB - The three novel human herpesviruses (HHV) 6, 7, and 8 are predominantly, but not exclusively, lymphotropic. In an attempt to elucidate their neurotropism in vivo, viral DNA sequences present in fresh autopsy cortical brain tissues obtained from 84 consecutive Chinese subjects (mean age, 66.9 years; range, 21-98 years) were detected by a nested polymerase chain reaction. These patients were apparently immunocompetent and free of clinical signs of viral diseases. HHV-6 DNA was detected in 36 of 84 (42.9%) patients, and the DNA-positive and -negative groups did not show a significant difference in age or sex distribution. Of the 36 HHV-6 DNA-positive cases, 9 (25%) were variant A and 27 (75%) were variant B. In view of the lower prevalence of variant A than variant B in the adult population, the two variants may share a comparable neuroinvasive potential. HHV-7 and HHV-8 DNA were detected respectively in three and two patients. The low positive rates of HHV-7 and HHV-8 may represent a relatively lower neuroinvasive potential of the viruses. Alternatively, the localization of HHV-7 and HHV-8 may be more restricted and the sampled cortical tissues may not represent the most abundant site of persistence in the nervous system. The results provide molecular evidence of the presence of the three newly identified herpesviruses in brain tissue. The pathogenic role for HHV-7 and HHV-8, as with HHV-6, in neurological diseases should not be overlooked. PMID- 10534732 TI - Human herpesviruses 6 and 7 as potential pathogens after liver transplant: prospective comparison with the effect of cytomegalovirus. AB - Because cytomegalovirus (CMV) is an important opportunistic infection after liver transplant, we conducted a prospective study to see if the same applied to human herpesviruses (HHV)-6 and -7. We used polymerase chain reaction (PCR) methods optimised to detect active, not latent, infection and studied patients not receiving antiviral prophylaxis for CMV. Post-transplant, 536 blood samples were tested by PCR (median 7; range 4-50). Active infection with CMV was detected in 28/60 (47%), HHV-6 in 19/60 (32%), and HHV-7 in 29/60 (48%) of patients. The PCR positive samples were tested by quantitative-competitive PCR to measure the virus load of each betaherpesvirus. The median peak virus load for CMV was significantly greater than that for HHV-6 or HHV-7. Detailed clinicopathological analyses for the whole population showed that CMV and HHV-6 were each significantly associated with biopsy-proven graft rejection. Individual case histories suggested that HHV-6 and HHV-7 may be the cause of some episodes of hepatitis and pyrexia. It is concluded that HHV-6 is a previously unrecognized contributor to the morbidity of liver transplantation, that HHV-7 may also be important and that both viruses should be included in the differential diagnosis of graft dysfunction. PMID- 10534733 TI - Whole cell lysate enzyme immunoassays vs. recombinant glycoprotein G2-based immunoassays for HSV-2 seroprevalence studies. AB - Seroepidemiology studies of herpes simplex virus type 2 (HSV-2) infections have been difficult to carry out because antibodies to HSV type 1 (HSV-1) show an extensive cross-reactivity with HSV-2 antigens. Many kits available currently are not entirely type specific for serodiagnosis of HSV-2 infections and therefore do not allow reliable discrimination of past exposure to these closely related alphaherpes viruses. Attempts to develop type-specific antigens have focused on the envelope glycoproteins, particularly glycoprotein G (gG). A cross-sectional study was carried out to examine the seroprevalence of antibodies to HSV-2 among healthy university students, using different methods: a whole cell lysate enzyme linked immunosorbent assay (ELISA), two different ELISAs, and a newly developed immunoblot assay, the last three based on recombinant gG2. HSV-2 prevalence was 24 times higher with the whole cell lysate ELISA (31%; 95% confidence interval [CI]: 27-35%) than the ELISAs and the immunoblot assay based on recombinant gG2 (1.3%; 95% CI: 0.1-2.5%), thus showing the inaccuracy of commercial tests based on whole-antigen preparations for epidemiological studies. Laboratories should be cautious and ensure that commercial tests for HSV typing are based on type specific glycoproteins. PMID- 10534734 TI - Mutations conferring resistance to zidovudine diminish the antiviral effect of stavudine plus didanosine. AB - This study evaluated the influence of zidovudine (ZDV) resistance mutations on the antiviral effect of the combination of stavudine (D4T) plus didanosine (ddI) in patients treated previously with ZDV plus zalcitabine (ddC). Twenty patients who had been treated with ZDV plus ddC for a median duration of 11 months (range, 7-42 months) were switched to D4T (40 mg twice a day [BID]) + ddI (200 mg BID) in an open pilot study lasting 6 months. The CDC classes were A (n = 10) and B (n = 10). The median baseline CD4 count was 285/mm(3) and the median baseline plasma virus RNA (Amplicor HIV Monitor RT-PCR assay) was 4.6 log copies/ml. Population based sequence analysis detected mutations associated with resistance to reverse transcriptase (RT) inhibitors in the RT domain of virus RNA from baseline plasma samples in 13/20 (65%) patients. Twelve patients had mutations associated with zidovudine resistance (3 T215Y - M41L - L210W; 3 T215Y - M41L; 2 T215Y - L210W; 3 T215Y; 1 K70R) and 1 patient had a multi-dideoxynucleoside resistance mutation (QI5IM). Patients with a resistance mutation had a significantly lower RNA suppression after 3 and 6 months (median RNA reduction -0.5 log and -0.1 log) than the remaining patients (-1.6 log and -2 log). Fifty percent of patients with wild-type viruses had undetectable plasma RNA after 24 weeks of D4T plus ddI therapy, whereas all those with mutated viruses had HIV RNA concentration > 3 log copies/ml at week 24 (P <.05). Our finding may have implications when deciding on a second line therapy with three or four drugs that includes two new nucleoside analogues. Cross-resistance between nucleoside analogues deserves maximal attention to ensure optimal antiretroviral therapy and design algorithms for antiretroviral management based on genotypic antiretroviral resistance testing. PMID- 10534735 TI - Latency of Epstein-Barr virus is stabilized by antisense-mediated control of the viral immediate-early gene BZLF-1. AB - The ability of the Epstein-Barr virus (EBV) to avoid lytic replication and to establish a latent infection in B-lymphocytes is fundamental for its lifelong persistence and the pathogenesis of various EBV-associated diseases. The viral immediate-early gene BZLF-1 plays a key role for the induction of lytic replication and its activity is strictly regulated on different levels of gene expression. Recently, it was demonstrated that BZLF-1 is also controlled by a posttranscriptional mechanism. Transient synthesis of a mutated competitor RNA saturated this mechanism and caused both expression of the BZLF-1 protein and the induction of lytic viral replication. Using short overlapping fragments of the competitor, it is shown that this control acts on the unspliced primary transcript. RT-PCR demonstrated unspliced BZLF-1 RNA in latently infected B lymphocytes in the absence of BZLF-1 protein. Due to the complementarity of the gene BZLF-1 and the latency-associated gene EBNA-1 on the opposite strand of the genome, we propose an antisense-mediated mechanism. RNase protection assays demonstrated transcripts in antisense orientation to the BZLF-1 transcript during latency, which comprise a comparable constellation to other herpesviruses. A combined RNAse protection/RT-PCR assay detected the double-stranded hybrid RNA, consisting of the unspliced BZLF-1 transcript and a noncoding intron of the EBNA 1 gene. Binding of BZLF-1 transcripts is suggested to be an important backup control mechanism in addition to transcriptional regulation, stabilizing latency and preventing inappropriate lytic viral replication in vivo. PMID- 10534736 TI - Epidemiological features of rotavirus infection in Caracas, Venezuela: implications for rotavirus immunization programs. AB - The epidemiological features of rotavirus infection may be quite relevant for evaluation of the performance of a rotavirus vaccine in different settings, as well as for monitoring its impact during vaccination under routine conditions. This article describes some important issues regarding rotavirus epidemiology in Venezuela, where major field trials of rotavirus vaccine have been carried out. Rotaviruses was significantly more frequently observed in inpatient (43%) than in outpatient (21%) consultations for diarrhea in infants and young children. There was a high prevalence of rotavirus illness, regardless of socioeconomic conditions, but the risk of dehydration was greater among the lower socioeconomic groups. Rotavirus disease occurs year-round, with a slight seasonal pattern. Eighty-five percent of rotavirus-positive diarrheal episodes, as well as 86% of cases of dehydration due to rotavirus, occurred during the first year of life. However, rotavirus illnesses occur less commonly during the first months of life (0-2 months), which may be a result of protection by transplacental antibodies. The pattern of acquisition of rotavirus antibody was consistent with this age distribution of disease and with optimal age for vaccination. Thus, regional epidemiological characteristics of rotavirus infection may affect optimal performance of rotavirus vaccine. PMID- 10534737 TI - Genetic investigation of novel hantaviruses causing fatal HPS in Brazil. AB - Although hantavirus pulmonary syndrome (HPS) was discovered in North America in 1993, more recent investigations have shown that the disease is a much larger problem in South America, where a greater number of cases and HPS-associated viruses have now been detected. Here we describe the genetic investigation of three fatal HPS cases from Brazil, including a 1995 case in Castelo dos Sonhos (CAS) in the state of Mato Grosso and two 1996 cases in the counties of Araraquara (ARA) and Franca (FRA), in the state of Sao Paulo. Reverse transcription-polymerase chain reaction (RT-PCR) products representing fragments of the hantavirus N, G1, and G2 coding regions were amplified from patient acute phase serum samples, and the nucleotide (nt) sequences (394, 259, and 139 nt, respectively) revealed high deduced amino acid sequence identity between ARA and FRA viruses (99.2%, 96.5%, and 100%, respectively). However, amino acid differences of up to 14.0% were observed when ARA and FRA virus sequences were compared with those of the geographically more distant CAS virus. Analysis of a 643-nt N coding region and a 1734-nt predominantly G2-encoding region of ARA and CAS virus genomes confirmed that these Brazilian viruses were distinct and monophyletic with previously characterized Argentinean hantaviruses, and suggested that Laguna Negra (LN) virus from Paraguay was ancestral to both the Brazilian and Argentinean viruses. The phylogenetic tree based on the N coding fragment also placed LN in a separate clade with Rio Mamore virus from Bolivia. At the amino acid level, ARA and CAS viruses appeared more closely related to the Argentinean viruses than they were to each other. Similarly, analysis of the diagnostic 139-nt G2 fragment showed that the Juquitiba virus detected in a 1993 fatal HPS case close to Sao Paulo city, Brazil was closer to Argentinean viruses than to ARA or CAS viruses. These data indicate that at least three different hantavirus genetic lineages are associated with Brazilian HPS cases. PMID- 10534738 TI - Semiautomated typing of human papillomaviruses by restriction fragment length polymorphism analysis of fluorescence-labeled PCR fragments. AB - A simplified version of a PCR-based reductional restriction fragment length polymorphism (rRFLP) approach for typing of human papillomaviruses (HPVs) is described previously [Wang et al., 1997]. It is achieved by the use of a biotin labeled primer in PCR which, on restriction digestion and staining, is associated with only a single restriction fragment. In this report, we describe a further development of the rRFLP approach with the use of a fluorescence-labeled primer in PCR and fragment detection by laser scanning in an automatic sequencer. HPV typing is achieved by computer-assisted matching of the fluorescence-labeled rRFLP patterns with a database of rRFLP patterns of all known anogenital HPV types. On analysis of the typing of 133 HPV-positive cases using this procedure, 20 different HPV types were detected in exfoliated cervical cells in PAP smear samples derived from Taiwanese women. The results indicate the existence of a heterogeneous population of HPV types in Taiwan. Although most cases were associated with the more common HPV types, a significant fraction (about 20%) of the HPV types detected was related to the less common genotypes, which are often not included in commercial kits available for HPV typing. The results indicate the importance of covering as many HPV types as possible in clinical HPV genotyping protocols. PMID- 10534739 TI - Estimating the time of HTLV-I infection following mother-to-child transmission in a breast-feeding population in Jamaica. AB - Mother-to-child transmission of human T-cell lymphotropic virus type I (HTLV-I) is primarily due to prolonged breast-feeding (>6 months) in the postnatal period. Most infant infections are not identifiable until 12 to 18 months of age by available whole virus Western blot serologic tests because of their inability to distinguish passively transferred maternal antibody from infant antibody. We investigated two methods to assess more accurately the time of infant infection. In prospectively collected serial biospecimens, HTLV-I-specific immunoglobulin (Ig) isotypes of IgM and IgA were determined by Western blot and HTLV-I proviral DNA was detected by polymerase chain reaction (PCR). IgA and IgG reactivity was assessed in periodic serum samples from 16 HTLV-I-seropositive children while IgM reactivity was assessed in 9 of the 16 children. Approximately three to five samples were tested for each child. IgG reactivity was observed in 100% of children at 24 months of age and 73% of children at 6-12 months of age; however, this could represent maternal and not infant antibody. Both IgA and IgM reactivity were insensitive indicators of infection, with only 50% of children showing reactivity at 24 months of age. PCR testing was performed in biospecimens obtained from 11 of these children. An estimated median time of infection of 11.9 months was determined by PCR, which was similar to the median time to infection determined by whole virus Western blot (12.4 months; P = 0.72). PCR tests support a median time to infection that is similar to that estimated by whole virus Western blot. PMID- 10534740 TI - MxA protein in capillary blood of children with viral infections. AB - Capillary blood of febrile children was lysed by using a lysis buffer containing ascorbic acid. MxA quantitation was performed by an immunochemiluminescent assay. The MxA values were significantly higher in capillary blood of infants with viral infections due to adenovirus (n = 5), rotavirus (n = 15), or respiratory syncytial virus (n = 28), than in capillary whole blood from infants with bacterial infections (n = 6) and healthy control patients (n = 20). A strong correlation was found between the MxA values in capillary whole blood and peripheral whole blood (r' = 0.86, P < 0.0001, n = 48). The MxA values found at these two sites were compared with the levels of IFN-alpha obtained by a dissociation enhanced lanthanide fluoroimmunoassay. A correlation between these two values was found. The results show that the combination of collection of blood by finger prick and specific immunochemiluminescent assay for MxA protein measurement may be of value for the diagnosis of viral infections in children. PMID- 10534742 TI - Impact of the introduction of A/Sydney/5/97 H3N2 influenza virus into South Africa. AB - In 1998 South Africa experienced a major influenza epidemic that was characterized by extensive illness and an unusually early season. The impact of the epidemic was charted by measuring proxy indexes of influenza activity such as school absenteeism and excess mortality in persons older than 65 years. Viruses isolated from patients of all age groups were analyzed both antigenically and at the molecular level to determine the characteristics of the influenza strain responsible for the outbreaks. The study revealed that influenza activity was detected as early as the middle of April and peaked toward the end of May and early June. School absenteeism correlated with a sharp rise in virus isolation during this period. Consumption of influenza-related pharmaceuticals, as well as mortality figures, also corresponded to the increased absenteeism and virus isolation. Characterization of the viruses isolated during 1997 and 1998 showed clearly that the epidemic was caused by the introduction of the A/Sydney/5/97 like H3N2 influenza strain into South Africa in 1998. With no prior exposure to this virus strain, which is antigenically distinct from the viruses that had been present in this country in 1997, the population was highly susceptible, resulting in an early, rapid spread of influenza. This epidemic has highlighted the importance of having an influenza vaccine specifically formulated for the Southern Hemisphere. If the 1998 vaccine had not contained the A/Sydney/5/97 strain, the widespread outbreaks in South Africa would have been far worse in terms of morbidity, mortality, and economic loss. This, in turn, emphasizes the need for increased influenza surveillance and international cooperation. PMID- 10534743 TI - A suitable model for wound healing: how many times are we to stumble over the same block? PMID- 10534744 TI - A suitable model for wound healing: how many times are we to stumble over the same block? PMID- 10534741 TI - Lassa and Mopeia virus replication in human monocytes/macrophages and in endothelial cells: different effects on IL-8 and TNF-alpha gene expression. AB - Cells of the mononuclear and endothelial lineages are targets for viruses which cause hemorrhagic fevers (HF) such as the filoviruses Marburg and Ebola, and the arenaviruses Lassa and Junin. A recent model of Marburg HF pathogenesis proposes that virus directly causes endothelial cell damage and macrophage release of TNF alpha which increases the permeability of endothelial monolayers [Feldmann et al. , 1996]. We show that Lassa virus replicates in human monocytes/macrophages and endothelial cells without damaging them. Human endothelial cells (HUVEC) are highly susceptible to infection by both Lassa and Mopeia (a non-pathogenic Lassa related arenavirus). Whereas monocytes must differentiate into macrophages before supporting even low level production of these viruses, the virus yields in the culture medium of infected HUVEC cells reach more than 7 log10 PFU/ml without cellular damage. In contrast to filovirus, Lassa virus replication in monocytes/macrophages fails to stimulate TNF-alpha gene expression and even down regulates LPS-stimulated TNF-alpha mRNA synthesis. The expression of IL-8, a prototypic proinflammatory CXC chemokine, was also suppressed in Lassa virus infected monocytes/macrophages and HUVEC on both the protein and mRNA levels. This contrasts with Mopeia virus infection of HUVEC in which neither IL-8 mRNA nor protein are reduced. The cumulative down-regulation of TNF-alpha and IL-8 expression could explain the absence of inflammatory and effective immune responses in severe cases of Lassa HF. PMID- 10534745 TI - Effects of temperature on tissue thermal injury and wound strength after photothermal wound closure. AB - BACKGROUND AND OBJECTIVES: Our goal was to determine the effect of temperature on the induction of tissue damage after laser-welded wound closure with and without albumin solder. STUDY DESIGN/MATERIALS AND METHODS: Multiple full-thickness skin incisions were made in a porcine model. Incisions were repaired by using a 1.32 microm laser at temperatures of 65 degrees C, 75 degrees C, 85 degrees C, or 95 degrees C with and without a 50% human albumin solder. The rate of apoptosis (programmed cell death) was quantified by counting the proportion of cells that stained positively for nuclear DNA fragmentation (nick end labeling). The distance that necrosis extended from the wound edge was also measured. The strength of the weld was measured with a tensiometer. RESULTS: For laser-welded repairs with solder, the amount of apoptosis at 65 degrees C and 75 degrees C was comparable to that of controls but became significantly elevated at 85 degrees C and 95 degrees C. The extent of necrosis was similar to that of controls at low temperature but also increased at 95 degrees C. Incisions repaired without solder showed increased necrosis compared with those repaired with solder at temperatures of 65 degrees C, 75 degrees C, and 95 degrees C at 0-0.5 mm from the incision. Wounds repaired at 85 degrees C and 95 degrees C showed more apoptosis in the absence of solder. The increased cell death at higher temperatures correlated with significantly decreased wound strengths at 3 days after repair in the solder group. A lower rate of cell death was observed in the solder group, which correlated with superior wound strength when compared with repairs without solder at days 0 (65-95 degrees C) and 3 (95 degrees C). CONCLUSION: Both apoptotic and necrotic cell death were used as quantitative measures of tissue injury and were accurate predictors of short-term wound strength. The addition of albumin solder decreased overall tissue injury. These results suggest that temperatures of 65-75 degrees C with solder provide the optimal conditions for maximizing acute wound strength and minimizing tissue injury. PMID- 10534746 TI - Radiometric surface temperature measurements during dye-assisted laser skin closure: in vitro and in vivo results. AB - BACKGROUND AND OBJECTIVE: A thermal camera was used to measure surface temperatures during laser skin welding to provide feedback for optimization of the laser parameters. STUDY DESIGN/MATERIALS AND METHODS: Two-centimeter-long, full-thickness incisions were made in guinea pig skin in vitro and in vivo. India ink was applied to the incision edges, which were then mechanically apposed. Continuous-wave, 1.06-microm Nd:YAG laser radiation was scanned over the incisions, producing an effective pulse duration of approximately 100 msec. Cooling durations between scans of 1.6, 4.0, and 8.0 sec were studied in vitro. A 5-mm-diameter laser spot was used with the power kept constant at 10 W. Thermal images were obtained at 30 frames per second with a thermal camera detecting 3-5 microm radiation. Surface temperatures were recorded at 0, 1, and 6 mm from the center line of the incision. RESULTS/CONCLUSIONS: Cooling durations of 1.6 and 4.0 seconds in vitro resulted in temperatures at the weld site that remained above approximately 65 degrees C for prolonged periods of time. Cooling durations of 8.0 seconds were sufficient both in vitro and in vivo to prevent a significant rise in baseline temperatures at the weld site over time. PMID- 10534747 TI - Simplified treatment planning for interstitial laser thermotherapy by disregarding light transport: a numerical study. AB - BACKGROUND AND OBJECTIVE: The objective was to investigate the effect of light transport on the temperature distribution and the coagulated volume under conditions relevant to interstitial laser thermotherapy (ILT) of tumors in the human liver. STUDY DESIGN/MATERIALS AND METHODS: Temperature distributions and coagulated volumes produced with a diffusing laser fiber or a conductive heat source, at equal output power, were numerically calculated for tissue with different optical penetration depths. Four irradiation times (5, 10, 20, and 30 min) were studied. A three-dimensional finite-element model was used to calculate the temperature distribution during heating with four conductive heat sources (no light emission). Results were compared with measured temperature distributions during laser irradiation in a gel phantom with known optical properties. RESULTS: Numerical calculations showed that the influence of light transport on the coagulated volume was negligible in tissue with optical penetration depths below 3-4 mm at all studied irradiation times. The phantom experiment indicated good agreement with the calculated temperature distribution, both with a single diffusing laser fiber and with four fibers. CONCLUSION: Light transport influences coagulated volumes only slightly under conditions presented in this work, which is relevant to ILT of tumors in the human liver. PMID- 10534748 TI - Evaluation of novel nonlaser light source for endometrial ablation using 5 aminolevulinic acid. AB - BACKGROUND AND OBJECTIVE: This research evaluated the effectiveness of a new nonlaser prototype short-arc lamp to achieve photodynamic ablation of endometrium in a rat. STUDY DESIGN/MATERIALS AND METHODS: Thirty female Sprague-Dawley rats were divided into two groups. 5-Aminolevulinic acid (ALA), the precursor to the photosensitizer protoporphyrin IX, was injected into the left uterine horn and vehicle alone (Hyskon) was injected into the right horn of 23 rats (group 1). An additional seven rats received vehicle only into both uterine horns (group 2). Three hours later, a cylindrical diffusing optical fiber was inserted into the lumen of the uterine horns, and light treatment was delivered from either a laser or a nonlaser light source. Rats in group 1 received either 1 hour (n = 15) or 10 minutes (n = 8) of light treatment into both uterine horns. In rats in group 2, the left horn was exposed to 1 hour of light treatment. Uterine tissues were examined histologically 4 days after light treatment. RESULTS: One hour of light exposure to the uterine horns injected with ALA produced extensive necrosis of the rat uterine wall. No difference in the magnitude of destruction was seen between the groups treated with the laser and nonlaser light sources. Ten minutes of light exposure resulted in endometrial ablation that was comparable in both the laser- and the prototype-treated groups, but the destruction of the deepest layers of the uterine wall was more consistent in the group treated with the nonlaser prototype. One hour of light treatment from either light source did not result in any histological changes in the uterine horns not exposed to ALA. CONCLUSION: The extent of endometrial ablation in the rat uterine horn achieved with the nonlaser prototype was comparable to that achieved with the laser. Thus, the nonlaser prototype may provide a less expensive approach to photodynamic endometrial ablation. PMID- 10534749 TI - Autofluorescence imaging and spectroscopy of normal and malignant mucosa in patients with head and neck cancer. AB - BACKGROUND AND OBJECTIVE: An early detection of oral cancer might improve the patient's prognosis. We present preliminary results of autofluorescence photodetection of cancerous oral mucosa. MATERIALS AND METHODS: 49 patients were investigated altogether. In 30 patients, malignant and healthy oral mucosa were excited with violet light (lambda = 375 to 440 nm). Images were recorded by a sensitive CCD camera. Spectrophotometric analysis in the green spectral range was performed on tumorous and innocuous mucosa in 36 patients. RESULTS: In 13 patients (43.3%), tumors were subjectively better distinguishable from their surroundings through a reduction of green autofluorescence than by ordinary inspection. Tumor detection abilities varied for different locations and tumor morphologies. Spectral analysis showed contrasts in autofluorescence intensities between tumor and normal tissues in 34 patients (94.4%). Autofluorescence spectra of normal mucosa varied both inter- and intraindividually. CONCLUSIONS: Using violet excitation light, camera-based autofluorescence photodetection in the green spectral range presented a highly promising tool for the diagnosis of oral malignomas in almost half of the cases examined. The possible ways on how the obtained results could serve to find a more advanced method for a precise tumor detection in the oral cavity are being discussed. PMID- 10534750 TI - Modeling the modification depth of carbon dioxide laser-treated dental enamel. AB - BACKGROUND AND OBJECTIVES: Many studies of laser-induced thermal decomposition of dental enamel have demonstrated a reduction in the rate of acid dissolution, size of artificial caries-like lesions, and acid reactivity. Additionally, studies have correlated the loss of carbonate from dental enamel with a reduction in acid dissolution. Dental mineral consists of hydroxyapatite with many substitutions, the major one being carbonate ( approximately 3-5% by weight), which markedly affects acid reactivity. The principle objective of the present work was to determine the precise depth of modification, i.e. , thermally induced decomposition of dental enamel (carbonate loss), at the predicted optimum laser irradiation parameters. STUDY DESIGN/ MATERIALS AND METHODS: Bovine enamel blocks were irradiated at lambda = 9.6 microm with 2-microsec and 100-microsec pulses and at lambda = 10.6 microm with 2-microsec pulses. Carbonate loss was calculated from infrared spectra as a function of depth and compared to numerical simulations of the maximum temperature rise. RESULTS: Carbonate loss was initiated at temperatures greater than 400 degrees C, but was complete only after repeated irradiation of the surface above the melting threshold. Carbonate loss of dental enamel irradiated at 9.6 microm with a 100-microsec pulse and at 10.6 microm with a 2-microsec pulse was greater than that of enamel irradiated at 9.6 microm with a 2-microsec pulse. The depth of carbonate loss in dental enamel irradiated with a 2-microsec pulse was greater for lambda = 10.6 microm than for lambda = 9.6 microm. CONCLUSION: The depth of modification is consistent with the presented model that incorporates the absorption depth and thermal relaxation time/pulse duration. However, repeated irradiation is required for complete removal of carbonate, depending on absorption depth and pulse duration. PMID- 10534751 TI - Optical properties of human trabecular meshwork in the visible and near-infrared region. AB - BACKGROUND AND OBJECTIVES: Despite disparate treatment parameters, similar success in laser trabeculoplasty (LT) is attained using the argon (514.5 nm) and diode (810 nm) laser. However, the mechanism of this success remains unresolved. To further understand LT, this study characterizes the optical properties of trabecular meshwork (TM). STUDY DESIGN/MATERIALS AND METHODS: Reflectance was measured from 10 TM samples over wavelengths of 400-820 nm, using an integrating sphere/spectrophotometer. Corrections were made for reflections at boundaries of refractive index mismatch. Kubelka-Munk coefficients were calculated and converted to linear transport coefficients. RESULTS: Scattering greatly dominated absorption. The scattering and absorption coefficients were, respectively, 141.20 +/- 15.80 cm(-1) and 4.89 +/- 1.95 cm(-1) at 514.5 nm, and 94.44 +/- 15.03 cm(-1) and 0.0874 +/- 0.111 cm(-1) at 810 nm (estimated anisotropy of 0.90). The corresponding penetration depths (1/e) were 69 microm (514.5 nm) and 106 microm (810 nm). CONCLUSION: The absorption coefficient of 514 nm energy is two orders of magnitude greater than 810 nm energy, while scattering coefficients are much closer. The fluence used at 514.5 nm is higher at the surface than that at 810 nm, but falls below it deep within the TM due to the differential absorption. Therefore, similar initial therapeutic effects are obtained with 810 nm using less total absorbed energy. Thermal damage resultant from excess energy deposited at 514.5 nm may be related to the lack of success in repeat argon LT, pointing out the need for studies of repeat diode LT. PMID- 10534752 TI - Determination of berberine in Rhizoma coptidis and its preparations by non aqueous capillary electrophoresis. AB - A non-aqueous capillary electrophoretic method was established for the determination of berberine in Rhizoma coptidis and its preparations. The effects of organic solvent and the concentrations of sodium acetate were studied, which showed that berberine in extracts of traditional Chinese medicine can be separated successfully in a buffer solution of 75 mmol/L of sodium acetate in methanol containing 1 mol/L of acetic acid. The simple and rapid method was linear in the range 25-200 microgram/mL of berberine and had a good reproducibility, with the relative standard deviation below 2%. Non-aqueous capillary electrophoresis is a satisfactory system for the analysis of alkaloids in traditional Chinese medicine. PMID- 10534753 TI - Relationship between the clinical effects of berberine on severe congestive heart failure and its concentration in plasma studied by HPLC. AB - It has been reported that berberine is valuable for long-term treatment of ventricular premature beats (VPBs) and leads to a decrease in mortality for patients with congestive heart failure (CHF). In order to improve its therapeutic value and reduce its side effects, it is necessary to study the relationship between its activity and plasma concentration in patients with CHF. Patients with CHF were treated with conventional therapy for 2 weeks. Immediately after the data from a dynamic electrocardiogram (DCG) and left ventricular ejection fraction (LVEF) were obtained, 1.2 g/day of oral berberine was given. After 2 weeks of berberine therapy, the DCG data and LVEF were reassessed and the plasma berberine concentration was measured by HPLC. Plasma samples were pretreated by extraction with chloroform. Berberine in all samples was determined using a mu Bondapak C(18) column, a mobile phase of acetonitrile:0.02 mol/L phosphoric acid (45:55, v/v), and a UV detector at 346 nm. The mean recovery was 96.5%. The linear range was 40-1600 ng/mL. The detection limit for berberine in plasma was 0. 4 ng. The decrease in frequency and complexity of VPBs and the increase in LVEF in patients with plasma berberine concentrations higher than 0.11 mg/L (n = 31, group B) were more significant than at concentrations lower than 0.11 mg/L (p < 0.01 vs p < 0.05). PMID- 10534754 TI - Interaction of carbamazepine and chlorpromazine in rabbits. AB - The interaction of carbamazepine and chlorpromazine in rabbits has been studied. The drugs were administrated as single oral doses (200 mg of each drug). The sequence of administration of the drugs was varied. It has been established that by simultaneous administration these drugs decrease absorption of each other in plasma. This may be explained by competition of the drugs to transfer from the gastrointestinal tract into plasma, as well as by the formation of complexes, more or less stable and more or less bound to gastrointestinal tissues. Carbamazepine intensifies the biotransformation of chlorpromazine, which may be caused by the ability of carbamazepine to induce microsomal liver enzymes. Chlorpromazine suppresses the biotransformation of carbamazepine, however. This may be caused by intensive capture of chlorpromazine by liver tissues and by its intensive biotransformation, which in turn is conditioned by its surface-active nature and by the increase of its metabolism with carbamazepine. Therefore the biotransformation of chlorpromazine is increased and metabolism of carbamazepine is reduced. The sequence of administration of the drugs affects their pharmacokinetics significantly. PMID- 10534755 TI - Direct HPLC analysis of ketoprofen in horse plasma applying an ADS-restricted access-phase. AB - Making up part of the unique family of restricted access materials (RAM) the Lichrospher ADS (alkyl-diol silica) sorbents have been developed as special packing materials for precolumns used for LC-integrated sample processing of biofluids. The advantage of such phases consists of direct injection of untreated biological fluids without sample clean-up and elimination of the protein matrix together with an on-column enrichment. The plasma samples, with internal standard phenacetin added (not essential), were brought onto the precolumn (C-18 ADS, 25 micron, 25 x 4 mm i.d.) using a phosphate buffer, 0.1 M, pH 7.0. After washing with the buffer, the ADS column was backflushed with the mobile phase phosphate buffer 0. 05 M pH 7.0: acetonitrile (80:20), thus transporting the analytes onto a reversed-phase column Ecocart 125-3 HPLC cartridge with a LiChrocart 4-4 guard column, both packed with LiChrospher 5 micron 100 RP-18; after separation detection was performed in UV at 260 nm. Essential features of the method include the novel precolumn packing, the absence of sample pretreatment, a quantitave recovery, good precision and accuracy, as well as a considerable reduction of analysis time compared to conventional manual methods applied in bioavailability studies. PMID- 10534756 TI - Quantitative analysis of cimetidine in human plasma using LC/APCI/SRM/MS. AB - A quantitative method was developed and validated for rapid and sensitive analysis of cimetidine in human plasma. The method involved the use of liquid chromatography (LC) coupled with atmospheric pressure chemical ionization (APCI) and selected reaction monitoring (SRM) mass spectrometry (MS). A cimetidine analog, SKF92374, was used as the internal standard. Separation of cimetidine and the internal standard was accomplished using a reverse-phase HPLC column (C18). The eluted components were ionized by the APCI source and subsequently detected by a highly selective triple quadrupole mass spectrometer in the SRM mode. Linear standard curves were obtained from 5 ng/mL (lower limit of quantitation) to 10,000 ng/mL. The results demonstrated excellent precision (%RSD 1. 1-8.9%) and accuracy (94.7-108.0%) over this range. In addition, the amount of plasma sample needed for analysis was small (50 muL), and the plasma pretreatment (analyte recovery >94%) was simple and time saving. This assay was used to evaluate cimetidine levels in premature infants following intravenous infusion of cimetidine. PMID- 10534757 TI - Enhanced theophylline metabolism in patients with bronchial asthma at age 4 and under. AB - The plasma levels of theophylline (TP) and its metabolites were measured in patients with bronchial asthma who were treated with a slow-release preparation of TP. The ratios of the plasma levels of these metabolites to TP levels in the group aged 1-4 years were larger than those in the group aged 5 years and older, suggesting enhanced activity of drug-metabolizing enzymes during infancy. PMID- 10534758 TI - Purification and analysis of the neutral glycan moiety of glycosyl phosphatidylinositol from porcine thyroid cells. AB - Metabolic labelling of inositolphosphate glycan with radioactive precursors is not sufficient to characterize and assess the involvement of the glycosyl phosphatidylinositol/inositolphosphate glycan (GPI/IPG) system in porcine thyroid cell signal transduction machinery. A protocol is described for the isolation and purification of free GPI using differential polarity of lipids and sequential thin layer chromatography. The purification until homogeneity of GPI constitutes a required step for gas chromatographic analysis. Next, successive chemical treatments allowed us to remove the neutral glycan moiety of thyroidal GPI, and its composition was obtained by gas chromatography. The proposed structure is consistent with data available for GPI anchor, but differs from compositional analysis data reported for insulin-sensitive GPI. Our results support the existence in porcine thyroid cells of the GPI/IPG system, which can take part in TSH-dependent signal transduction processes. PMID- 10534759 TI - Protein A tangential flow affinity membrane cartridge for extracorporeal immunoadsorption therapy. AB - Tangential flow affinity membrane cartridge (TFAMC) is a new model of immunoadsorption therapy for hemoperfusion. Recombinant Protein A was immobilized on the membrane cartridge through Schiff base formation for extracorporeal IgG and immune complex removal from blood. Flow characteristics, immunoadsorption capacity and biocompatibility of protein A TFAMC were studied. The results showed that the pressure drop increased with the increasing flow rate of water, plasma and blood, demonstrating reliable strength of membrane at high flow rate. The adsorption capacities of protein A TFAMC for IgG from human plasma and blood were measured. The cartridge with 139 mg protein A immobilized on the matrix (6 mg protein A/g dry matrix) adsorbed 553 mg IgG (23.8 mg IgG/g dry matrix) from human plasma and 499.4 mg IgG (21.5 mg IgG/g dry matrix) from human blood, respectively. The circulation time had a major influence on IgG adsorption capacity, but the flow rate had little influence. Experiments in vitro and in vivo confirmed that protein A TFAMC mainly adsorbed IgG and little of other plasma proteins, and that blood cell damage was negligible. The extracorporeal circulation system is safe and reliable. PMID- 10534761 TI - Study on the bioavailability of baicalin-phospholipid complex by using HPLC. AB - A complex of baicalin with soy phospholipid was prepared to improve the bioabsorption of baicalin. A simple and sensitive HPLC method was developed for baicalin determination in rat plasma. It was shown that its plasma concentration reached a peak of 0.42 microgram/mL 5.3 h after oral administration, 600 mg/kg. However, after intake of its phospholipid complex, a peak of 0.90-microgram/mL occurred at a later time, 6.1 h. The elimination of the complex tended to be slower than that of the free drug. There was a significant difference in the mean area under the concentration-time curves (AUC) between the free drug and the complex (p < 0.01). PMID- 10534760 TI - Development of predictive retention-activity relationship models of antipsychotic drugs by micellar liquid chromatography. AB - The predictive and interpretative capability of quantitative chromatographic retention-biological activity models is supported by the fact that in adequate experimental conditions the solute partitioning into the chromatographic system can emulate the solute partitioning into lipid bilayers of biological membranes, which is the basis of drug and metabolite uptake, passive transport across membranes and bioaocumulation. The use of micellar solutions of Brij35 as mobile phases in reversed liquid chromatography has proven to be valid in predicting some biological activities of different kinds of drugs. In this paper, the correlations between the logarithm of capacity factors and pharmacokinetic, preclinical pharmacology and therapeutic efficacy parameters of phenothiazines are studied. Parabolic quantitative retention-activity relationship models with predictive and interpretative ability have been obtained. PMID- 10534762 TI - Genes, chromosomes, and rhabdomyosarcoma. AB - Rhabdomyosarcomas are a heterogeneous group of malignant tumors and are the most common soft-tissue sarcoma of childhood. Rhabdomyosarcomas resemble developing skeletal muscle, notably in their expression of the MRF family of transcription factors and the PAX3 and PAX7 genes. These PAX genes are also involved through specific translocations, t(2;13)(q35;q14) and variant t(1;13)(p36;q14) in the alveolar subtype, which result in PAX3-FKHR and PAX7-FKHR fusion genes, respectively. The fusion genes are thought critically to affect downstream targets of PAX3 and PAX7 or possibly have novel targets. Similar downstream changes may also be involved in embryonal and fusion gene negative cases. Genomic amplification of such genes as MYCN, MDM2, CDK4, and PAX7-FKHR is a feature mainly of the alveolar subtype, while specific chromosomal gains, including chromosomes 2, 8, 12, and 13, are associated with the embryonal subtype. Loss of alleles and imprinting at 11p15.5 and disruption of genes such as IGF2, ATR, PTC, P16, and TP53 have also been implicated in rhabdomyosarcoma development. Whereas there is now a realistic possibility of cure in the majority of cases, there remains a subset that is resistant to multimodality therapy, including high-dose chemotherapy. Characterization of the defining molecular features of tumors that are likely to behave aggressively represents a particular challenge. Current research is leading toward a better understanding of rhabdomyosarcoma tumorigenesis, which may ultimately result in novel therapeutic strategies that increase the overall cure. Genes Chromosomes Cancer 26:275-285, 1999. PMID- 10534764 TI - Nonrandom karyotypic features in squamous cell carcinomas of the skin. AB - We report the finding of clonal chromosome abnormalities in 13 short-term cultured squamous cell carcinomas (SCCs) of the skin. Intratumor heterogeneity, in the form of cytogenetically related (subclones) or unrelated clones, was detected in six tumors. Whereas clones with complex karyotypic changes were found in 6 tumors, clones with simple anomalies were observed in 10 tumors, and sometimes these clones coexisted with highly abnormal clones. Rearrangement of chromosome 8, in the form of isochromosome i(8q) or whole arm translocation, was the most common aberration, found predominantly in complex clones. Another recurrent feature, i.e., the centromeric rearrangement of chromosome 1, as isochromosome i(1q) or i(1p), or whole arm translocations, was always part of a complex karyotype. Homogeneously staining regions were found in two cases, one with a highly complex karyotype and the other with a simple karyotype. In order to obtain an overall karyotypic picture in SCC of the skin, the cytogenetic findings in 10 SCCs reported earlier were reviewed. The chromosomes most commonly affected were, in decreasing order, chromosomes 1, 11, 8, 9, 5, 3, and 7. Chromosomal sites most frequently rearranged were almost all pericentromeric: they were 8q10-q11, 1p10-q12, 5p10-q11, 11p15, and 9p10-q10. Recurrent anomalies were i(1q), i(8q), i(5p), i(1p), i(9p), and i(9q). Among them, only i(8q) and i(9q) might be assumed to be early genetic events, considering the fact that they could occasionally be identified in simple clones. The most frequent losses included part of or the entire chromosomes 2, 4, 9, 11, 14, 18, and 21, arm 8p, and chromosomes X, Y, and 13. Overrepresentation most frequently involved 1q, chromosome 7, and 8q. The characteristic karyotypic pattern observed in skin SCC was in line with the experience in several other carcinomas. Genes Chromosomes Cancer 26:295-303, 1999. PMID- 10534763 TI - Identification of germline missense mutations and rare allelic variants in the ATM gene in early-onset breast cancer. AB - Epidemiological studies have shown an increased risk of breast cancer in obligate ataxia telangiectasia (A-T) heterozygotes. We analyzed 100 samples from young breast cancer patients for mutations in ataxia-telangiectasia mutated (ATM), the gene responsible for the autosomal recessive condition, A-T, to determine whether A-T heterozygosity predisposes such individuals to develop breast cancer. These patients were selected from families with a moderate or absent family history of breast cancer and included a subset of 16 radiosensitive patients. Forty-four germline sequence variants were detected by fluorescent chemical cleavage of mismatch of RT-PCR products. These included seven rare variants found in nine patients (three described for the first time), but no truncating mutations. Although three variants were detected in the radiosensitive subset, this was not statistically significant compared to the nonradiosensitive group. One variant, G2765S, is likely to be a missense mutation, but the other six variants probably represent rare polymorphisms. However, five of the seven rare germline variants detected showed loss of heterozygosity of the wild-type ATM allele for one or more markers close to the ATM locus in matched tumor DNA. This high rate of somatic inactivation of ATM may indicate either that these rare variants play a role in breast cancer development or alternatively that a neighboring tumor suppressor gene is important for tumorigenesis. We found germline truncating ATM mutations to be rare in these young breast cancer patients and therefore they are unlikely to play a role in the etiology of their disease. Genes Chromosomes Cancer 26:286-294, 1999. PMID- 10534765 TI - Dominant genetic alterations in immortalization: role for 20q gain. AB - Gain of 20q has been observed in many cancer types, including bladder cancers. However, the biological significance of low-copy-number 20q gain in human cancer pathogenesis has not yet been defined. We reported that immortalization of human uroepithelial cells (HUC) transformed with human papillomavirus 16 (HPV 16) E7 is associated with single-copy 20q gain (P = 2 x 10(-7)). We also observed 20q13.2 amplification in some cell lines, but only after 20 passages. Thus, we hypothesized that low-copy gain of 20q gene(s) contributes in a dominant way to bypassing HUC senescence. To test this hypothesis, we fused precrisis E7 transformed HUCs (pcE7s) with three independent immortal E7-HUCs that acquired a single-copy 20q gain at immortalization. In one of these lines, a single-copy gain of 20q and a 10p12.1-pter loss were the only cytogenetic alterations. Immortal cell hybrids were obtained with all three crosses. Southern analysis for unique HPV16 insertion sites, as well as fluorescence in situ hybridization (FISH) with whole chromosome 20 painting probes (WCP20) for marker chromosomes in the immortal clones, confirmed the hybrid and independent nature of representative immortal clones. In contrast, when we used the same protocol, no immortal somatic cell hybrids were obtained when HPV16 E6 immortal HUC (E6-HUC) that showed 3p and 9p losses, but no 20q gain, were fused with precrisis E6 transformed HUC (pcE6s). This latter observation is consistent with many results demonstrating that recessive changes are required for cell immortalization. Therefore, the new results reported herein for the first time demonstrate that dominant changes can contribute to bypassing senescence, and that such genes may be located on 20q. Genes Chromosomes Cancer 26:304-311, 1999. PMID- 10534766 TI - Nonrandom chromosomal aberrations and cytogenetic heterogeneity in gallbladder carcinomas. AB - Chromosome banding analysis of 11 short-term cultured gallbladder carcinomas revealed acquired clonal aberrations in seven tumors (five primary and two metastases). Three of these had one clone, whereas the remaining four were cytogenetically heterogeneous, displaying two to seven aberrant clones. Of a total of 21 abnormal clones, 18 had highly complex karyotypes and three exhibited simple numerical deviations. Double minutes and homogeneously staining regions were observed in one and two carcinomas, respectively. To characterize the karyotypic profile of gallbladder cancer more precisely, we have combined the present findings with our three previously reported cases, thereby providing the largest cytogenetic database on this tumor type to date. A total of 287 chromosomal breakpoints were identified, 251 of which were found in the present study. Chromosome 7 was rearranged most frequently, followed by chromosomes 1, 3, 11, 6, 5, and 8. The bands preferentially involved were 1p32, 1p36, 1q32, 3p21, 6p21, 7p13, 7q11, 7q32, 19p13, 19q13, and 22q13. Nine recurrent abnormalities could, for the first time, be identified in gallbladder carcinoma: del(3)(p13), i(5)(p10), del(6)(q13), del(9)(p13), del(16)(q22), del(17)(p11), i(17)(q10), del(19)(p13), and i(21)(q10). The most common partial or whole-arm gains involved 3q, 5p, 7p, 7q, 8q, 11q, 13q, and 17q, and the most frequent partial or whole-arm losses affected 3p, 4q, 5q, 9p, 10p, 10q, 11p, 14p, 14q, 15p, 17p, 19p, 21p, 21q, and Xp. These chromosomal aberrations and imbalances provide some starting points for molecular analyses of genomic regions that may harbor genes of pathogenetic importance in gallbladder carcinogenesis. Genes Chromosomes Cancer 26:312-321, 1999. PMID- 10534767 TI - Fine-structure deletion mapping of 10q22-24 identifies regions of loss of heterozygosity and suggests that sporadic follicular thyroid adenomas and follicular thyroid carcinomas develop along distinct neoplastic pathways. AB - Previous studies have demonstrated frequent loss of heterozygosity (LOH) of markers on chromosome arm 10q in both follicular thyroid carcinomas (FTCs) and follicular thyroid adenomas (FAs). A novel tumor suppressor gene, PTEN, has been mapped to 10q23.3 and is the susceptibility gene for Cowden syndrome, an autosomal dominant disorder characterized by multiple hamartomas and a risk of benign and malignant tumors of the breast and thyroid. Studies examining the relationship of somatic PTEN status and follicular thyroid neoplasms have only demonstrated a variable subset of tumors that have somatic monoallelic deletions of PTEN, suggesting that other tumor suppressor genes may be present in this region. We therefore sought to conduct a detailed examination of LOH of 20 polymorphic markers in a 19-cM region spanning 10q22-24, including PTEN, in 44 FAs and 17 FTCs. Using this fine-structure somatic mapping approach, we defined at least two novel regions of LOH in follicular adenomas and follicular carcinomas, suggesting the presence of at least two distinct tumor suppressor genes that may play a role in thyroid neoplasia. Furthermore, the difference in patterns of LOH in adenomas versus carcinomas lends additional support to the hypothesis that adenomas and carcinomas can develop along two separate, nonserial pathways. Genes Chromosomes Cancer 26:322-328, 1999. PMID- 10534768 TI - Isolation and characterization of a novel TP53-inducible gene, TP53TG3. AB - We applied the differential mRNA display method to isolate genes regulated by wild-type TP53 in cells of a colon-cancer line (SW480) in which we had established an inducible TP53 expression system under the control of the lactose operon. Here we report isolation and characterization of a novel TP53-inducible gene, termed TP53TG3 (TP53 target gene 3). Its DNA sequence was identical to sequences present in two BAC clones that had been mapped to chromosome band 16p13. The gene expressed several transcripts by alternative splicing; the two major transcripts, TP53TG3a and TP53TG3b, encoded 124- and 132-amino-acid peptides that were expressed predominantly in testis. Immunohistochemical analysis using cancer cells (HeLa or H1299) that had been transfected with plasmid DNA designed to express the MYC-fused TP53TG3 proteins indicated that these products were present mainly in the cytoplasm 20 hr after transfection. However, 40 hr after transfection, the recombinant proteins had accumulated in the nuclei of some cells. Because no known nuclear localization domain was present in the amino acid sequence, we suspect that this protein plays an important role in the TP53-mediated signaling pathway, when it forms complexes with other protein(s) and is transferred by them into the nucleus. Genes Chromosomes Cancer 26:329-335, 1999. PMID- 10534769 TI - Combined spectral karyotyping and DAPI banding analysis of chromosome abnormalities in myelodysplastic syndrome. AB - Spectral karyotyping (SKY) is a new molecular cytogenetic technique that allows simultaneous visualization of each chromosome in a different color. We have used SKY for comprehensive analysis of 20 myelodysplastic syndromes (MDSs) (13 primary MDSs, 3 therapy-related MDSs, and 4 acute leukemias developed from MDS, including 1 cell line established from a secondary leukemia), previously analyzed by G banding. To locate the chromosomal breakpoints, DAPI-counterstained band images from all metaphases were transformed to G-band-like patterns. By using SKY, it was possible to identify the origin and organization of all clonal marker chromosomes (mar), as well as the origin of all abnormalities defined as additional material of unknown origin (add) or homogeneously staining regions (hsr) by G-banding. In total, SKY identified the chromosomal basis of 38 mar, add, and hsr, corrected 8 abnormalities misidentified by G-banding, and revealed 6 cryptic translocations in 5 cases. Total or partial chromosomal loss (mainly of -5/5q- and -7/7q-) is the most frequent cytogenetic abnormality in MDS. In 3 of 11 cases with -5/5q- and in 4 of 8 with -7/7q-, lost material was detected by SKY in unbalanced translocations. A total of 60 chromosomal losses were identified by G-banding in 16 cases with multiple chromosome abnormalities involving at least 3 chromosomes. For 26 of these losses (43%), SKY analysis suggested that the losses were not complete, but had been translocated to a variety of partner chromosomes. Moreover, SKY analysis revealed that a ring chromosome in a case of acute leukemia developed from MDS contained three to six segments that originated from chromosome 21 material. Fluorescence in situ hybridization showed the amplification of the AML1 gene on regions derived from chromosome 21, providing the first evidence of amplification involving this gene in MDS. Genes Chromosomes Cancer 26:336-345, 1999. PMID- 10534770 TI - Genetic alterations during the progression of squamous cell carcinomas of the uterine cervix. AB - Most cervical carcinomas appear to arise from cervical intraepithelial neoplasia (CIN) lesions. In addition to infection with high-risk human papilloma viruses, which is indicative of an increased risk of progression, alterations of oncogenes and tumor suppressor genes play a role. Genetic studies of CIN lesions, primary cervical carcinoma, and metastases may shed light on the relative importance of various genetic alterations involved in the progression of CIN to invasive carcinoma. We examined tumor material from 10 patients with squamous cell carcinoma of the uterine cervix and synchronous CIN lesions and lymph node metastases. The CIN component, invasive carcinoma, and lymph node metastases were analyzed separately for loss of heterozygosity (LOH) on the following loci: VHL (3p21), HLA region (6p22-23), PGL (11q 22-24), E6 associated protein (15q11-13), TP53 (17p13), DCC (18q21.1), and chromosomes 1, 2, 4, 9, 20, and X. Using immunohistochemistry, the expression of the EGF receptor, ERBB2, and TP53 was determined. In CIN lesions, frequent LOH was found at chromosome arms 3p, 6p, and 11q. Primary invasive carcinoma showed additional LOH at chromosome arms 6q, 17p, and 18q. In lymph node metastases, an additional locus on the X chromosome displayed LOH. All carcinomas and synchronous lesions but one showed high expression levels of the EGF receptor. TP53 staining, when present, was found in all synchronous lesions. Focal staining of ERBB2 was found in one CIN lesion, two invasive carcinomas, and four metastases. The molecular alterations accumulated in a fashion that paralleled the progression of the tumors. These results indicate that cervical tumorigenesis occurs in a stepwise fashion, including infection and integration of oncogenic HPV and several specific genetic alterations. Genes Chromosomes Cancer 26:346-354, 1999. PMID- 10534771 TI - Rapid and sensitive minimal residual disease detection in acute leukemia by quantitative real-time RT-PCR exemplified by t(12;21) TEL-AML1 fusion transcript. AB - Because previous PCR-based methodologies for detection of minimal residual disease (MRD) in leukemia patients have been too cumbersome to allow for widespread clinical usefulness, we have employed a real-time quantitative PCR (RQ PCR) system to develop an MRD assay for t(12;21). We initially determined the expression of the different alternatively spliced TEL-AML1 mRNAs found in t(12;21) breakpoint variants I and II. We then optimized PCR primers for the RQ PCR system and, using the t(12;21)+ REH cell line in spiking experiments, found a linear detection of TEL-AML1 over at least five logs. Moreover, 1 malignant cell in a background of 1,000,000 normal cells could be detected. The expression of the GAPDH, ABL, and beta(2)-microglobulin (beta2M) housekeeping genes were then compared in normal donors and in leukemic patients, and the very stably expressed beta2M was selected as an internal reference gene, allowing us to compensate for variation in RNA quality and day-to-day variation. In 12 samples from t(12;21) positive patients at diagnosis, the levels of the TEL-AML1 fusion transcripts were found to vary up to 14-fold after normalization to beta2M. Interestingly, in samples obtained from seven patients at diagnosis, during induction chemotherapy, or relapse, the level of TEL-AML1 in peripheral blood (PB) and bone marrow (BM) was found to differ only by threefold, suggesting that MRD may be evaluated in PB samples in most patients. We conclude that this assay could set new standards for t(12;21) MRD detection with its accuracy, its high throughput, and its short turnover time for samples. Genes Chromosomes Cancer 26:355-365, 1999. PMID- 10534772 TI - 3' BCR recombines with IGL locus in BCR-ABL-positive philadelphia-negative chronic myeloid leukemia. AB - We have isolated the 3' BCR breakpoint junction of a complex BCR-ABL1 rearrangement found in leukemic cells with a cytogenetically normal karyotype, and the corresponding germline fragment that spanned the 3' BCR recombination site. Fluorescence in situ hybridization localized the 3' BCR recombination site to 22q11, about 350-600 kb proximal to BCR. Restriction map and DNA sequence comparisons indicated that 3' M-Bcr had recombined at a site within the variable region (Itv Region IV) of the immunoglobulin lambda (IGL) locus. Somatic rearrangement of DNA sequences (variable, joining, and constant regions) within the IGL locus, as in other Ig and TCR loci, represents the basis for human antibody diversity. Misrecombination of these somatically rearranging sites has been associated with chromosomal rearrangements in lymphoid leukemia and lymphoma, but there are no previous descriptions of IGL involvement in genomic aberrations associated with myeloid leukemia. Genes Chromosomes Cancer 26:366 371, 1999. PMID- 10534773 TI - Missense and nonsense mutations in codon 659 of MLH1 cause aberrant splicing of messenger RNA in HNPCC kindreds. AB - Germline mutations that give rise to premature termination codons in mRNAs have frequently been associated with aberrant processing of the nascent transcripts. This can take the form either of nonsense-mediated mRNA decay or of aberrant splicing of the pre-mRNA. In a family affected by hereditary nonpolyposis colorectal cancer, a two-nucleotide deletion in codon 659, which introduces a frameshift and a new stop codon in exon 17 of the DNA mismatch repair gene MLH1, has been reported to lead to skipping of the exon. We now report that this phenomenon occurs also when there are missense or nonsense mutations in this codon. Our results thus suggest that in aberrant splicing the nature of the mutation may be less important than its position within the exon. These findings are of importance to mutation interpretation, as they imply that aberrant splicing could be associated even with silent mutations that do not lead to amino acid substitutions. Genes Chromosomes Cancer 26:372-375, 1999. PMID- 10534774 TI - Germline mutations in NF1 patients with malignancies. AB - We have analyzed 98.5% of the coding region of the NF1 gene at the cDNA level in seven NF1 patients who developed malignant peripheral nerve sheath tumors. Seven germline mutations were detected in six individuals: a 6-bp in-frame deletion in exon 28, a splice acceptor mutation in intron 31 resulting in a premature stop of translation, a missense mutation in exon 38, and three total NF1 gene deletions. In one of the patients with a total NF1 gene deletion, a missense mutation in exon 16 on the other NF1 allele was detected. These data indicate that NF1 patients developing malignant neoplasms can have any type of NF1 germline mutation such as a total gene deletion, a frameshift mutation, an in-frame deletion, or a missense mutation. We conclude that in our series no specific type of NF1 germline mutation was found in NF1 individuals with malignancies, but that large NF1 gene deletions were more frequently found in this group than reported for the general population of NF1 individuals. Genes Chromosomes Cancer 26:376 380, 1999. PMID- 10534775 TI - Splice variant lacking the transactivation domain of the BRCA2 gene and mutations in the splice acceptor site of intron 2. PMID- 10534776 TI - Primary parauterine leiomyoma with a t(6;14) PMID- 10534777 TI - Sail your way to a healthy heart. The Mediterranean diet. PMID- 10534778 TI - Countering untreatable angina. PMID- 10534779 TI - Cereal fiber reduces coronary risk in women too. PMID- 10534780 TI - Fish-oil supplements slow atherosclerosis. PMID- 10534781 TI - Sudden death runs in families. PMID- 10534782 TI - Ask the doctor. I frequently get a dull chest pain that goes on for hours at a time. My doctor tells me that it is not coming from my heart and that I shouldn't worry, but how can he be sure that such severe pain isn't dangerous to me? PMID- 10534783 TI - Illness without disease - part II. PMID- 10534784 TI - Neurobiological effects of early trauma. PMID- 10534785 TI - Lost in the shuffle. PMID- 10534786 TI - A new treatment for pedophilia. PMID- 10534787 TI - What is Asperger's disorder? PMID- 10534788 TI - Gender and stress. PMID- 10534789 TI - Women and sleep. PMID- 10534790 TI - What minerals do we need? PMID- 10534791 TI - Overactive bladder. PMID- 10534792 TI - Too many mastectomies for early breast cancers. PMID- 10534793 TI - By the way, doctor, I'm thinking of trying a laser to remove the fine hair above my upper lip. How do lasers measure up to electrolysis, waxing, and other methods of removing hair? PMID- 10534794 TI - Immelmann's indignation. PMID- 10534795 TI - Cough threshold in people with spinal cord injuries. AB - BACKGROUND AND PURPOSE: The purpose of this study was to compare the cough threshold between people with and without spinal cord injury (SCI). The effect of smoking on cough threshold was also investigated. SUBJECTS: The participants were 26 people with SCI (15 smokers, 11 nonsmokers) and 18 people without SCI (9 smokers, 9 nonsmokers). METHODS: Aerosols of citric acid were delivered with incremental doubling concentration from 62.5 mmol to 2 mol. Cough threshold was defined as the first concentration of citric acid that induced at least 2 coughs, which is associated with large chest excursion and concurrently acoustic response. RESULTS: The mean cough thresholds of smokers and nonsmokers with SCI (209 and 417 mmol, respectively) were lower than those of smokers and nonsmokers without SCI (467 and 1,072 mmol, respectively). The mean citric acid cough thresholds decreased in smokers with and without SCI when compared with nonsmokers with and without SCI. CONCLUSION AND DISCUSSION: The cough sensitivity increased in subjects with SCI, and smoking could also increase the cough sensitivity. Training about the frequency and technique of cough in patients with SCI should be carefully monitored. PMID- 10534796 TI - Endurance training of the trunk extensor muscles in people with subacute low back pain. AB - BACKGROUND AND PURPOSE: Clinicians treating patients with low back pain often use exercise to reduce pain and improve function. The aim of this study was to evaluate the effectiveness of trunk extensor endurance training in reducing pain and decreasing disability in subjects with subacute low back pain (ie, onset of back pain within 7 days to 7 weeks). SUBJECTS AND METHODS: Patients were randomly assigned to either an experimental group or a control group. A visual analog scale and the pain rating index (PRI) of the McGill Pain Questionnaire (MPQ) were used to obtain baseline measurements of pain. The Roland Morris Disability Questionnaire (RMDQ) was used to measure disability, and the Sorensen Test was used to measure trunk extensor endurance. Subjects in the experimental group attended exercise sessions 3 times a week for 6 weeks. Subjects in the control group did not do exercises. Both groups were given back care advice and hot packs for 15 minutes, 3 to 5 times per week. Reassessments were carried out at 3 and 6 weeks. RESULTS: There were differences between the 2 groups at 3 weeks in regard to pain intensity during the evaluation session and pain experienced over the preceding 24 hours, the total MPQ PRI, the sensory component of the MPQ PRI, and the RMDQ. At 6 weeks, no differences were found for pain measurements, disability scores, and holding time on the Sorensen Test. CONCLUSION AND DISCUSSION: Trunk extensor endurance training reduced pain and improved function at 3 weeks but resulted in no improvement at 6 weeks when compared with the control group. Endurance exercise is considered to expedite the recovery process for patients with an acute episode of low back pain. PMID- 10534798 TI - Thirtieth Mary McMillan Lecture: PT 2000: nurturing the profession. PMID- 10534797 TI - Four clinical tests of sacroiliac joint dysfunction: the association of test results with innominate torsion among patients with and without low back pain. AB - BACKGROUND AND PURPOSE: The purpose of this study was to assess the association between innominate torsion (asymmetric anteroposterior positioning of the pelvic innominates) and Gillet, standing forward flexion, sitting forward flexion, and supine-to-sit tests. SUBJECTS: A sample of 21- to 50-year-old patients with low back pain (n=150) and a comparison group of patients with upper-extremity impairments (n=138) were recruited from outpatient physical therapy facilities. METHODS: The association of single and combined test results with innominate torsion (calculated from pelvic landmark data) and with presence or absence of low back pain were estimated via odds ratios, sensitivities, specificities, and predictive values. RESULTS: Individual test sensitivities were low (8%-44%), as were negative predictive values (28%-38%), for identifying the presence of innominate torsion. Combining tests and controlling for sex, age group, leg length difference, or iliac crest level did not improve performance characteristics. The associations of test results with low back pain were weak, with the exception of the Gillet test (odds ratio=4.57). CONCLUSION AND DISCUSSION: The data do not support the value of these tests in identifying innominate torsion, although the use of these tests for identifying other phenomena (eg, sacroiliac joint hypomobility) cannot be ruled out. Further exploration of the association of Gillet test results with low back pain is warranted. PMID- 10534799 TI - 1999 APTA Presidential Address: leaders who inspire change. American Physical Therapy Association. PMID- 10534800 TI - More on the clinical doctorate. PMID- 10534801 TI - More on the clinical doctorate. PMID- 10534802 TI - Methods of literature review. PMID- 10534804 TI - Insertion/deletion polymorphism of the angiotensin-converting enzyme gene and hypertension. PMID- 10534803 TI - Tumor necrosis factor-alpha and interleukin-10 genotypes in congestive heart failure. PMID- 10534805 TI - The diagnostic dilemma of the unilateral cystic kidney-ADPKD with aplasia of one kidney. PMID- 10534806 TI - Acute renal failure following cardiac surgery. PMID- 10534807 TI - Haemodialysis in India. PMID- 10534808 TI - An insight into rheumatology resources available on the world wide web. Reply To letters by McNally et al. and armstrong et al PMID- 10534809 TI - Growth hormone replacement and the risk of malignancy in children with neurofibromatosis AB - Howell SJ, Wilton P, Lindberg A, Shalet SM. J Pediatr 1998;133:201-5. PMID- 10534810 TI - Actuarial and kaplan-meier survival analysis: there is a difference PMID- 10534812 TI - Drug therapy before coronary artery operations PMID- 10534811 TI - Emerging new concepts of myocardial laser revascularization PMID- 10534813 TI - The not-so-hidden persuaders. PMID- 10534814 TI - Ask the doctor. I signed up to be part of an epidemiological study and had a test called electron beam CT done on my heart. I do not have any symptoms of heart disease, but a note was sent to my doctor saying that I had a lot of calcium in my coronary arteries, and my doctor isn't sure what to do. What does this test result mean? PMID- 10534815 TI - Trauma and post-traumatic stress in severe mental illness. PMID- 10534816 TI - Does smoking lead to crime? PMID- 10534817 TI - Children's faces, east and west. PMID- 10534819 TI - By the way, doctor. Whenever my doctor prescribes a medication for me, I ask my pharmacist for the package insert describing the drug. However, I often find the information more alarming than helpful. How can supposedly safe drugs have so many side effects? PMID- 10534818 TI - Scant evidence for progesterone cream. PMID- 10534820 TI - Occupational injury. PMID- 10534821 TI - Patient cost-benefit analysis of teledermatology measured in a randomized control trial. AB - A randomized controlled trial of the costs and benefits of teledermatology consultations compared with traditional hospital consultations was carried out. Over a nine-month period, 197 patients were referred from general practice for a dermatological opinion, 98 for a teledermatology consultation and 99 for a hospital consultation. Eighty patients required an additional subsequent hospital appointment. Patients were asked to complete an economic questionnaire after each consultation, and 164 questionnaires were returned: 62% of those randomized to the teledermatology consultation responded compared with 58% of those randomized to the hospital consultation. Patients seen by teledermatology at their own health centre had shorter distances to travel and spent less time overall attending the appointment compared with those seen at the hospital. However, the teledermatology consultations were more time-consuming for the general practitioner and dermatologist. These findings indicate that teledermatology has more benefits for the patient than for the health-care delivery team. PMID- 10534822 TI - Applications of low-cost eye tracking in telemedicine. AB - A low-cost, high-resolution system for measuring eye position information and eye movement has been developed. The Vision Control System (VCS) equipment can be worn as a lightweight headset. The ability to operate equipment via eye movement empowers individuals by keeping hands free to perform other tasks. Potential applications of the VCS equipment include its use in surgical training as an on screen pointing device to highlight areas of interest, use by paramedics to relay information at the scene of an accident, use to examine a radiologist's visual scanning technique in reading films, and use in ophthalmology to monitor eye behaviour. PMID- 10534823 TI - Telemedical record documentation: a preliminary survey. AB - Telemedical records are the weak link in telemedicine. With a number of medicolegal and reimbursement issues still unresolved, telemedical record documentation will be a critical piece of the puzzle when the patient's original legal medical record must be produced. A questionnaire was distributed to active telemedicine programmes in the USA to determine how interactive teleconsultations were being documented. Forty programmes completed the questionnaire. The responses indicated a lack of consistency and a need for documentation guidelines for telemedical records. PMID- 10534824 TI - The legal and risk management conundrum of telemedicine. AB - A store-and-forward teledermatology service was established between two general practices and the department of plastic surgery at Derriford Hospital in Plymouth. An academic lawyer and an expert in risk management made an assessment of the service. They looked at the following issues: medicolegal problems; security; confidentiality; and risk. None of them was considered insoluble and there is no reason why such issues should prevent the National Health Service from developing telemedicine services. All organizations considering telemedicine need to ensure that the proposed telemedicine service satisfies the issues raised in this study. PMID- 10534825 TI - Business risks in telemedicine. AB - There is a wide range of potential risks associated with telemedicine. Medical professionals and managers who are contemplating the use of telemedicine are strongly advised to undertake a full risk analysis before, not after, implementation. PMID- 10534826 TI - The enhancement of audience participation in telemedicine education by the use of electronic voting. AB - The rotation of trainee registrars in obstetrics and gynaecology to peripheral centres in South Australia was introduced for the first time in January 1998. A pilot study of the use of telemedicine to maintain the trainees' involvement in the established weekly training sessions conducted at the Women's and Children's Hospital (WCH) was also commenced at the same time. Sessions were problem based and required the trainees to be actively involved throughout. An electronic voting system (EVS) was used by the WCH audience, with the results of all responses being seen at the peripheral centres. Initial evaluation of the EVS as an educational tool was sufficiently positive to justify further studies of its use in this environment. The availability of voting units at the peripheral sites, used in conjunction with the main centre, will provide an exciting opportunity for further research. PMID- 10534827 TI - Ten years' clinical experience with telemedicine in prenatal care in Hungary. AB - A multicentre randomized clinical trial of prenatal home care of pregnant women was carried out in Hungary. Pregnant women registered contraction activity of the uterus daily using a portable contraction monitor. The data were transmitted directly to the physician's PC for analysis. Of 748 women who entered the study, only 263 fulfilled all the requirements of randomization, monitoring and treatment. The preterm birth rate in the study group was half that of the control group. Telemedical prenatal monitoring improves perinatal results by providing more intensive and better observation of pregnant women. PMID- 10534828 TI - Evaluation of the first international teleconference in ophthalmology. AB - The First International Teleconference in Ophthalmology was held during March 1998 between five sites in the UK, USA, Greece and Malaysia. ISDN transmission at 128 kbit/s was used to reduce costs while maintaining the clarity of the presented material. Specialized lecture theatres were not available at all sites and conventional halls had to be adapted for videoconferencing. For this reason initial point-to-point testing was carried with Bristol to simplify problem solving. Thereafter, a multipoint bridge was used to connect all sites together. During the conference a number of individual presentations were given, all followed by extensive discussion periods. Special instructions were given beforehand on the production of slide material, with particular reference to font sizes and colour combinations. Full use was made of various presentation media, including slides, videos and live demonstrations. The conference was attended by over 500 delegates, all of whom were specialists in ophthalmology. The technology employed was ideal for teaching purposes. However, if used in a clinical field, it should be kept in mind that the choice of transmission rate makes certain features not easily apparent in images but they become clearer when pointed out by the presenter. PMID- 10534829 TI - An evaluation of an Australian videoconferencing project for child and adolescent telepsychiatry. AB - A user satisfaction survey of videoconferencing services for child and adolescent mental health was carried out over a two-year period in Victoria, Australia. The aims of the survey were to evaluate key utilization areas, effect on professional practice, and advantages and disadvantages of the videoconferencing service. Eighty questionnaires were sent out and 58 (73%) were returned. Consultations were reported as the most frequent use of videoconferencing equipment (62%), followed by clinical use (59%), supervision (36%), teaching (19%) and administration (14%). Fifty-seven per cent of respondents reported that videoconferencing had affected professional practice. Advantages of the services included cost savings (52%) while disadvantages included technological problems (40%). The findings show the benefits of videoconferencing for improving the delivery of mental health care in rural Australia. PMID- 10534830 TI - School-based telemedicine: using technology to bring health care to inner-city children. AB - In the USA, children in low-income, medically understaffed areas have difficulty accessing health care. A pilot telemedicine service was established in Kansas, linking four elementary schools with a local hospital at 128 kbit/s. PC-based videoconferencing systems, a stethoscope, an otoscope and fax machine were installed for each school's nurse. During the first three months of operation, 187 consultations were conducted by a hospital physician over the video-link. Ear, nose and throat problems were the most common reason for referral (29%), followed by dermatological problems (20%) and mandatory school physicals (20%). Prescriptions were issued in 67% of cases. Eighteen per cent of children required referrals for specialist care. The pilot service successfully enabled medically disadvantaged children to receive health care and prompted four additional schools to join the project. PMID- 10534831 TI - Telemedicine and the organization of the health sector. AB - The widespread introduction of telemedicine would have organizational consequences for the health sector. However, the problem of health-sector organization is complex. A general framework for analysis of organizational consequences has been developed and consists of eight points. These are: (1) the starting point; (2) mapping of applications of telemedicine where organizational consequences are possible; (3) characterizing the nature of larger organizational consequences; (4) the consequences for the health sector in the chosen area; (5) possible consequences for the number and placement of geographical subunits of the health sector; (6) possible consequences for health sector organizational types; (7) the consequences for the tools of health policy; and (8) design of the change process towards a new organization. PMID- 10534832 TI - A three-year review of telemedicine at the community level--clinical and fiscal results. AB - The Telemedical Emergency Neurosurgical Network (TENN) was developed to establish community-level access to neurosurgical critical care services for an underserved urban population. Since October 1994, the system has provided round-the-clock connectivity between a single peripheral hospital and a 10-station receiving network which includes physicians' homes and offices. Data from the first 35 months of operation confirm clinical efficacy and cost-effectiveness. The primary benefit was in providing rapid access to care for an 'at-risk' underserved population in emergencies. This undoubtedly resulted in the actual saving of life and prevention of permanent disability. The total costs over the 35 months were calculated as $64,375 and the total savings (mainly in avoided air transport) were $626,149, a net saving of $561,774. PMID- 10534833 TI - Telemedicine: capabilities of telecommunications in clinical practice in Russia. AB - Successful implementation of telemedicine services depends on an adequate telecommunications infrastructure. At present the main infrastructure in Russia and the countries of the former Soviet Union consists of analogue telephone lines. These are sufficient for e-mail and access to the World Wide Web but do not provide sufficient bandwidth for the transmission of realtime teleconsultations. ISDN lines and fibre optic cables are becoming more widespread in the major cities but have yet to be installed in the regions. PMID- 10534834 TI - The North Norwegian Health Net. AB - The North Norwegian Health Net is a comprehensive scheme to connect all health care institutions in the area to a national computer network. Services available include telemedicine, e-mail and Web access. A general practitioner has responsibility for ensuring that the clinical information is correct. Medical departments are responsible for the content of their own Web pages. All institutions require authorization before connection to ensure data protection and security. Changes in communication between primary- and secondary-care sectors are being monitored. To date the implementation of the network programme has gone smoothly. PMID- 10534835 TI - Low-cost ISDN videoconferencing equipment for orthopaedic second opinions. AB - We validated the application of low-cost ISDN videoconferencing equipment for telemedicine. A telemedicine benchmark was designed and four different office videoconferencing systems were evaluated, all using basic-rate ISDN connections (128 kbit/s). All the low-cost systems showed generally good or acceptable performance for clinical use. We also investigated the feasibility of videoconferencing for an orthopaedic second-opinion service. Eight point-to-point conferences were conducted to discuss real clinical cases by use of interactive sharing of medical images. The average duration of each session was 35 min. Encouraging results were obtained. PMID- 10534836 TI - Design and implementation of a telemedicine system in Palestine. AB - The shortage of expert physicians and health-care providers in Palestine has created the need for a telemedicine system. The difficulty and the cost of transfer between Palestinian cities in the current political situation makes it important to use such systems. This paper describes the project's progress. The software and hardware required were developed in our laboratories. The software included modules for the preprocessing, enhancement, manipulation, compression and decompression of medical images. The hardware included the communication protocols, physical layer interface circuit and selection of a suitable medium for communication. In tests a radiograph was transmitted from one computer to another. The second and third phases will be to establish a communication link between Palestinian hospitals at different locations and to examine the realtime transfer of medical images. PMID- 10534838 TI - The uneven diffusion of telemedicine services in Australia. AB - The diffusion of telemedicine services in Australia over the last four years has been uneven. Some of the major barriers to the adoption of telemedicine relate to the nature of the industry, including its immaturity, the limited telecommunications infrastructure, the lack of appropriate dialogue between vendors and buyers about solutions required, and the lack of partnerships in the industry. Remuneration is only one barrier. There are, of course, other substantial organizational, financial and attitudinal barriers to the adoption of telemedicine. Further development of telemedicine in Australia will require detailed assessment of these matters. PMID- 10534837 TI - The Internet and an orthopaedic discussion group. AB - Since its inception, the Internet has created many new opportunities for the dissemination of knowledge. We have developed an Internet-based orthopaedic discussion group. There are 93 members from Ireland and elsewhere in the world. Precautions are taken to ensure the confidentiality of all cases submitted to the group. All patient identifiers are removed from images, radiographs and reports before submission. In the first 18 months more than 100 cases were discussed by the group. The use of an Internet-based discussion group in orthopaedics has expanded the educational and communication opportunities for the Irish orthopaedic profession both nationally and internationally. PMID- 10534839 TI - Nuclear medicine teleradiology in regional and rural centres of Western Australia. AB - We have established a nuclear medicine teleradiology service for rural and regional areas of south-west Western Australia. This study aimed to evaluate whether existing treatment models had altered in ways which were directly related to improved medical outcomes and reduced cost. Preliminary observation into musculoskeletal injury of the ankle joint included diagnosis of 10 radio-occult talar dome lesions which in some cases had not been identified until six months after the trauma. Several cases suggest that nuclear medicine provided a more timely diagnosis and actively altered treatment. Further, the potential role of radiologists as primary-care extenders in guiding appropriate investigations and defining the nature of the illness before specialist consultation may lessen the economic burden on public health care while creating economies of scale by expanding the clinical usefulness of the nuclear physician over a large rural area. PMID- 10534841 TI - Burns follow-up: an innovative application of telemedicine. AB - We assessed the efficiency and effectiveness of burn consultations via telemedicine. The Regions Hospital Burn Center completed 87 follow-up visits with 40 patients via telemedicine from March 1997 to August 1998. These consultations involved burn physicians, occupational therapists and a clinical psychologist. Patients were seen at 15 telemedicine sites in six states (Minnesota, Iowa, Montana, North and South Dakota, and Wisconsin). Telemedicine burn consultations were cost-effective for the patient, but were more time consuming for the physician and therapist. As remote sites become more familiar with preparing patients for teleconsultations, telemedicine will become more efficient for the physician while remaining cost-effective for the patient. PMID- 10534840 TI - Telemedicine on a small island. AB - Information on the health status of the population of a small Greek island was collected. The information consisted of personal data, clinical history, physical examination, blood pressure evaluation, electrocardiography, and ultrasound scans of neck, breast and abdomen. Ninety-six per cent of the entire island population (280 inhabitants) participated in the study. Two per cent were at risk of serious complications of pathological disease and were immediately referred to a regional hospital for adequate care while 25% had minor pathological problems. Our experience suggests that health-care workers on small islands should be trained in the use of technology as a means of communication with mainland hospitals. PMID- 10534842 TI - Dynamic robotic telepathology: a preliminary evaluation on frozen sections, histology and cytology. AB - We evaluated the diagnostic efficacy of a dynamic robotic telepathology system for the delivery of pathology services to distant hospitals. The system provided static/dynamic features and the remote control of a robotic microscope using four ISDN lines. For evaluation purposes, 184 consecutive cases were diagnosed at distance using the system. The cases were 60 frozen sections, 64 cases of gastrointestinal pathology and 60 cases of urinary cytology. The telemedicine diagnoses obtained in this way were compared with traditional microscopic diagnosis. Diagnostic agreement ranged from 90% in urinary cytology to 100% in frozen sections. The results suggest that a dynamic robotic telepathology system can be a useful tool for supporting the pathology practice of isolated hospitals. PMID- 10534843 TI - Digital teledermatology for skin tumours: a preliminary assessment using a receiver operating characteristics (ROC) analysis. AB - A low-cost store-and-forward teledermatology system using digital images for the remote diagnosis and management of skin tumours was evaluated. Two hospitals participated in the trial. Patients were seen face to face at one hospital, and had their images and clinical history viewed remotely by a different dermatologist at a second hospital. A preliminary receiver operating characteristics (ROC) analysis revealed clinical agreement between the teledermatologist and face-to-face dermatologist in 93% of cases in terms of their assessment of the benign/malignant nature of the lesions. Sensitivity of the judgements was 88% and specificity was 80%. These preliminary findings indicate the potential for remote management of skin tumours using a low-cost system in the National Health Service. PMID- 10534844 TI - Clinical outcomes in telepsychiatry. AB - A retrospective review of patient records comparing clinical outcomes of patients seen by interactive television (IATV) and those seen in person was carried out to determine whether IATV reduced the quality of psychiatric care. A Global Assessment of Functioning (GAF) score was determined for each patient in both the study and control groups at the initial visit, and at subsequent visits. Forty nine patients with either major depression or schizoaffective disorder were studied. No significant difference was found in the percentage change in GAF scores between the two groups, which suggests that clinical outcomes were not affected by the use of IATV. Patients seen by IATV had a greater attendance rate and follow-up visits took less than half the time compared with in-person visits, indicating that IATV was an acceptable and efficient method of providing psychiatric services. PMID- 10534845 TI - Telemedicine and distance learning--experience in Georgia and future prospects. AB - Telemedicine and distance learning have a high priority in Georgia owing to the uneven distribution of health-care resources. Outside the country's capital, Tbilisi, resources and manpower are scarce, so telemedicine may be the best option for health-care delivery in spite of the costs. However, an adequate infrastructure must be in place to make this possible. The National Information Learning Centre (NILC) provides access to the Internet and MEDLINE and is currently assessing the best software for low-bandwidth distance learning and videoconferencing. PMID- 10534846 TI - Telemedicine in Sweden--a diffusion study. AB - A survey of telemedicine use in Sweden was carried out. The response rate by hospital management was high at almost 90%. Sixty per cent of hospitals were involved in some kind of telemedicine activity in Sweden, with a further 15% planning telemedicine implementation. In rural areas of north Sweden good links exist between primary and secondary care. In the more populated southern region good links exist between the county hospitals and specialist university hospitals. Fifty-four per cent of telemedicine applications were used for specialist consultations, 13% for consultation between paramedics and hospitals and 10% for rounds. Teleradiology was the most frequent service. PMID- 10534847 TI - The role of telecare in the management of exacerbations of chronic obstructive pulmonary disease in the home. AB - We examined home care as an alternative to hospital admission in exacerbations of chronic obstructive pulmonary disease (COPD). We performed a pilot study to investigate the feasibility of using telecommunications technology to assist in the support of acutely ill patients with exacerbations of COPD at home. Realtime, interactive video, via an analogue video-phone, was used to allow patients in their own homes to obtain nursing support from a nurse located at a distant base station. Six individuals, four male and two female, had video-phones installed in their homes by members of the nursing intervention team. The age range was 52-72 years, mean 61.5. These patients used the system on 18 occasions. Experience in home telecare, via interactive video, has been limited to provision of ongoing support for relatively stable individuals with chronic illness. This pilot project represents the first attempt at providing home telecare in the UK to those experiencing an acute exacerbation of their chronic illness, who would otherwise have merited acute hospital admission. PMID- 10534848 TI - Sigmoidoscopy in a nurse-practitioner community clinic using telemedicine. AB - We evaluated the feasibility of remote endoscopy in a community setting. Realtime teleconsultation and telesigmoidoscopy were carried out by a nurse practitioner at a community clinic while a colorectal specialist was present at the main hospital. Rigid video-sigmoidoscopes were used and the images were transmitted via ISDN lines at 384 kbit/s. Over three months, 32 patients (mean age 35 years; 19 men and 13 women) with bleeding per rectum took part in the study. Evaluation was carried out using satisfaction questionnaires for the patients, the nurse practitioner and the clinicians. The mean grade for clarity of intraluminal views was 3.5 (1 poor, 4 excellent). Only two cases had views graded less than 3, due to the presence of excessive faecal residue. All the patients were satisfied with the teleconsultation and video-endoscopy and would return for a similar visit. User satisfaction was also high on the part of the nurse practitioner and the clinician. PMID- 10534849 TI - Realtime remote consultation in the outpatient clinic--experience at a teaching hospital. AB - We used an outpatients' telemedicine clinic to evaluate remote consultation in a London hospital practice. Video-images were relayed by ISDN at 384 kbit/s between the main hospital site and the sister hospital. Teleconsultation was evaluated in three different specialties: pain management, orthopaedics and general surgery. Medical personnel involved in the consultation consisted of a resident or senior house officer at the remote site and a senior registrar or consultant at the main hospital. A total of 146 consultations was held over a six-month period in the three clinical specialties involved. Questionnaires were completed by 58% of patients and over 90% of the medical personnel involved. The mean scores (on a five-point Likert scale) for physician satisfaction at the main and distant site were 3.6 and 3.8, respectively. On a scale of 0-10, 92% of patients rated the consultation 8-10 and 90% indicated that they would return for a similar consultation. Seventy-three per cent felt that a similar consultation from their referring doctor would have been appropriate. Confidence in the diagnosis was high among the specialists at the main hospital site. PMID- 10534850 TI - A programme management model for the Nova Scotia telemedicine network. AB - Sustainable telemedicine networks are not the norm but rather the exception. Unless a formal programme management model is in place, telemedicine cannot be integrated successfully into the mainstream of modern health care. Critical factors in achieving sustainable telemedicine have been identified. The programme management model developed for the Nova Scotia Telemedicine Network incorporated the following tools: a telemedicine self-assessment indicator, service modelling, technical design and integration, application development and validation, scheduling, ongoing network management and evaluation. PMID- 10534851 TI - Low-bandwidth tele-ultrasound. AB - We examined the acceptability and diagnostic accuracy of dynamic ultrasound images transmitted at 128 kbit/s and 384 kbit/s. The gold standard was the direct recording of 200 ultrasound examinations on video-tape. The taped images were later transmitted at both 128 kbit/s and 384 kbit/s and recorded, resulting in three tapes for each case. Four observers viewed each tape individually. Ninety per cent of images transmitted at 384 kbit/s were rated as diagnostically acceptable compared with 32% of images transmitted at 128 kbit/s. Diagnostic agreement between tapes transmitted at 384 kbit/s and the gold standard was 85%, compared with 78% for 128 kbit/s transmissions. Observers were not satisfied with low-bandwidth transmission of ultrasound images despite adequate diagnostic accuracy. Dynamic ultrasound images transmitted at 384 kbit/s were viewed as both diagnostically acceptable and accurate. PMID- 10534852 TI - A business model for telemedicine. AB - The US Military Healthcare System commissioned a clinical and business case study for telemedicine in Europe. The aims of the study were to identify the major cost areas by specialty for each European location, to assess the feasibility of a telemedicine alternative and to carry out a cost-benefit analysis. We also examined 2000 consecutive air referrals to determine whether telemedicine could have avoided evacuation. The study showed that 31,000 conventional consultations could be replaced by telemedicine in Europe each year. The potential savings amounted to $3.7 million in travel costs and 25,000 working days. The business and clinical case for telemedicine was strong, thus providing one of the prerequisites for successful telemedicine. PMID- 10534853 TI - Teleconsultation for cooperative acquisition, analysis and reporting of ultrasound studies. AB - We have developed a teleconsultation system to support cooperation between doctors at different institutions in the acquisition, analysis and reporting of ultrasound studies. All the relevant activities were supported by the application. The system enabled remote supervision of the ultrasound study as it was being performed, cooperative static image acquisition and analysis, as well as report writing. Cooperative analysis of previously acquired studies was also possible. The system is undergoing evaluation in five hospitals in Portugal. PMID- 10534854 TI - The legal implications of veterinary telemedicine and telecare. AB - This paper presents an overview of some legal implications of the application of telemedicine to veterinary medicine. As in other areas of medicine, veterinary practitioners need to be aware that remote practice or consultation requires adherence to the same professional standards as in normal clinical veterinary practice. Licensing, confidentiality and malpractice are discussed. PMID- 10534855 TI - The Cornwall dermatology electronic referral and image-transfer project. AB - An evaluation of a store-and-forward electronic referral system was carried out in Cornwall to gauge its effectiveness in prioritizing dermatological patients needing secondary care. A dermatologist classified patients to be seen according to need based on information on the electronic referral form along with an image of the skin condition. Initial findings indicated that patients were more accurately triaged in at least 50% of cases and that 25% of patients did not require an outpatient dermatological appointment. Dermatologists rated image quality at 7.5 on a 10-point scale. This low-cost system has potential for more appropriate clinical management of dermatological patients in both primary and secondary care. PMID- 10534856 TI - Telemedical experiences at an Antarctic station. AB - Wintering-over in Antarctica represents a physician's most remote and inaccessible scenario, apart from a space station. Because of the harsh and unpredictable winter weather, Antarctic stations are typically inaccessible for over six months of the year. Telephone and fax communication, and recently other forms of telemedicine, have provided vital links to specialists. The author was the sole physician for more than 250 people wintering-over during the 1995 austral winter at McMurdo Station. There were several instances of serious or life-threatening illness where the author relied on teleconsultation. These cases included new-onset coronary artery disease, posterior hip dislocation, complicated Colles' fracture and acute appendicitis. There were also numerous consultations for non-emergency clinical presentations normally managed by specialists. Telemedicine was a crucial link to specialists from the remote and inaccessible environment of Antarctica. PMID- 10534857 TI - Project RATEMA--one year's experience. AB - The RATEMA project is an international cooperation between the University of Ulm in Germany and the Urals Research Centre for Radiation Medicine in Chelyabinsk, Russia. For one year we conducted weekly conferences between the two sites, based on a satellite link with a 384 kbit/s connection. During the videoconferences the physicians on both sides--experts in radiation medicine--discussed the health status of Russian patients who had been chronically exposed to ionizing radiation in the South Urals region. The German partners presented patients with comparable haematological and oncological diseases. The project has shown the advantages and difficulties of working in an international and interdisciplinary environment. The experience gained has been very valuable for planning new projects with similar tasks. The results and the contents of the RATEMA database are the basis for education and research for physicians involved in the management of radiation victims. PMID- 10534858 TI - Using cable television networks for interactive home telemedicine services. AB - Most recent cable television network infrastructures can be used to deliver broadband interactive telemedicine services to the home. These facilities allow the provision of social and health services like medical televisiting for elderly, disabled and chronically ill patients; health tele-education; and teleconsultation on demand. Large numbers of patients could benefit from these services. There is also the increasing European tendency to offer customized home care services. These applications are being developed and validated by a pilot project in Madrid as part of the ATTRACT project of the European Commission. The long-term aim is to develop broadband applications on a large scale to support low-cost interactive home telemedicine services for both patients and institutions. PMID- 10534859 TI - Tele-otolaryngology consultations between two rural primary-care centres in southern Lapland and the University Hospital of Umea. AB - In 1996 a telemedicine link was established between two primary-care centres of Vasterbotten county and the University Hospital. Specialties involved at the University Hospital were otoloaryngology, orthopaedics and dermatology. Videoconferencing used ISDN at 384 kbit/s. The primary-care centres were equipped with video-endoscopes. During the first 21 months, there were 32 otolaryngology consultation. The average time for each consultation was between 15 and 30 min. Patients, general practitioners and specialists were interviewed using questionnaires with answers on a six-point scale, in which a score of six was best. Patient satisfaction produced a mean score of 5.7. The specialist doctors rated the video-consultation satisfactory for diagnosis. Roughly 40% of the referrals could be avoided by telemedicine. The general practitioners rated the educational effect of the consultation very highly. PMID- 10534860 TI - Evaluation of a training and diagnostic ultrasound service for general practitioners using narrowband ISDN. AB - We carried out a four-month pilot study of telemedicine for the delivery of training and diagnostic ultrasound services for general practitioners (GPs). Two health centres and a district general hospital were linked using ISDN (128 kbit/s) and PC-based videoconferencing units. Sixteen videoconferencing sessions were conducted, with 64 patients scanned over the ISDN link for a range of clinical conditions. Nine cases of pathology were demonstrated and confirmed by a consultant radiologist. A total of 229 images were transmitted using the store and-forward facility, of which 194 (85%) images were of diagnostic quality and 35 (15%) were regarded as non-diagnostic. Allowing for sampling error, we would expect 79-89% of stored images to provide diagnostic information at a 95% confidence level. A total of 115 store-and-forward images (SAFI) were randomly selected and compared with hard-copy images (HCI) for technical quality. There was a significant difference in the quality of images. The results show that SAFI are far superior to HCI, traditionally used by clinicians for making primary diagnosis and that the superior quality of the SAFI improved diagnostic accuracy. This pilot study showed a high degree of confidence in videoconferencing for training GPs to carry out ultrasound clinics, enhancing their ability to make accurate diagnoses, and providing them with immediate access the second and higher opinion. PMID- 10534862 TI - A review of telemedicine. AB - Moving the information instead of the patient is often sufficient and telemedicine is a useful and potentially powerful tool which fulfils this idea. It has changed the classical form of health-care delivery by providing those people living in rural and remote areas with comparable services to those in urban areas. Equally, remote hospitals can obtain specialist consultations and diagnosis without the need for the patient to travel. We have reviewed the present status of telemedicine and its future possibilities. PMID- 10534861 TI - Integrating interactive television-based psychiatric consultation into an urban community mental health service. AB - We conducted a pilot study of urban telepsychiatry employing interactive television (IATV) over ISDN links for psychiatric consultation to support a primary-care mental health team. During the six months of the pilot phase, 30 consultations were arranged by the general practitioners for 14 different patients. Of these, 24 were completed by IATV and one by telephone when the IATV link failed. The system was used to manage patients with complex problems, many of whom were difficult to engage in standard services. The results showed high levels of user acceptance. Various problems for the further implementation of such systems were also identified. PMID- 10534863 TI - The sustainability of telemedicine projects. AB - Although there are instances where the provision of health care is successfully driven by the profit motive, in most countries it is considered a public service. The provision of telemedicine services assuredly meets an important social need to extend health care to remote and rural areas in developing countries. While there are potential advantages and benefits from telemedicine, the evidence of its cost-effectiveness and sustainability is meagre. Telemedicine undoubtedly yields cost savings in certain circumstances, but few service providers have found a way to recover their costs (and make a profit) from those to whom they provide their service. With their low expenditures per person, developing countries face a daunting challenge in making such public services sustainable. Pilot projects should be a first step in demonstrating the cost-effectiveness and benefits of telemedicine, but such projects should also be sustainable. Sponsors of such pilot projects must have a clear plan from the start about how the project can continue after the sponsorship comes to an end. This paper examines ways in which telemedicine services can be made sustainable. PMID- 10534864 TI - Training for telemedicine. AB - All technology requires training. Simply installing videoconferencing equipment in a hospital or health centre, and leaving the manufacturer's manual nearby, is not sufficient to encourage or maintain its use for telemedicine. Presenting clinicians and other users with thick policy and procedure documents to be read and understood is also not helpful. Following initial awareness raising about telemedicine and its potential, staff must be properly trained: not just in how to turn on the equipment but in how to use it effectively for consultation, education and administrative purposes. Training needs to be continual, especially where staff turnover is high. It should be practical, pitched at different levels, incorporated into mainstream activities such as orientation courses and staff development days wherever possible, and supplemented with clear, straightforward protocols and user-friendly instruction manuals. PMID- 10534865 TI - The basic diagnostic approaches used in robotic still-image telepathology. AB - We investigated the basic diagnostic processes used in telepathology with our robotic still-image system, OLMICOS, by analysing the steps and patterns used in 20 consecutive tissue section diagnoses. Three basic approaches were recognized. One was magnifying a suspect finding in a low-powered microscopic image. This approach was used mostly for confirming or characterizing a tumour. The second approach was scanning over a low-powered image by magnifying square images to form a mesh. This was found to be useful to confirm the presence or absence of signs and was mostly used as the initial step in judging the surgical margin of malignant cases or diagnosing lesions. The third approach was a combination of these two and was used for delineating the surgical margin, confirming the absence of metastases or diagnosing difficult lesions. Recognition of these three basic diagnostic approaches is important in making a rapid and correct remote diagnosis. PMID- 10534866 TI - Twelve months' experience with telemedicine for the British armed forces. AB - In 1998, a low-cost telemedicine system was established for the British armed forces in Bosnia. It involved a digital camera, computer, e-mail and a telephone link between the remote site and the UK. Images were transferred to a PC and forwarded to specialists in the UK, who replied within 24 hours. In the first 11 months of use, there were 60 referrals from one remote site. Clinical management following specialist advice was significantly altered in a third of these cases, with additional reassurance offered in 13% of cases. When other sites were connected the usage of the service doubled. This inexpensive system provided effective care to our patients, irrespective of location. PMID- 10534867 TI - MultiMed--remote interactive medical simulation. AB - MultiMed is a telemedicine and tele-education project to extend access to a sophisticated medical simulation facility in Bristol. It will allow remote users to undertake simulated medical scenarios and to access a reference database, for a comprehensive remote learning experience. The first phase will focus on the area of anaesthesia and all remote users will be based in the UK. PMID- 10534868 TI - The commissioning of renal services. PMID- 10534869 TI - A patient requests an old-style tonic. PMID- 10534870 TI - Key developments in urology. PMID- 10534871 TI - STIs are still a growing problem. PMID- 10534872 TI - The patient with haematuria. PMID- 10534873 TI - Cardiovascular disease in the elderly. PMID- 10534874 TI - Tinnitus. PMID- 10534875 TI - Who's responsible for men's health? PMID- 10534876 TI - A depressed refugee who 'aches all over'. PMID- 10534877 TI - Key issues in men's health. PMID- 10534878 TI - Testicular lumps in general practice. PMID- 10534879 TI - Managing erectile dysfunction. PMID- 10534880 TI - Prostatic disease: a rational approach. PMID- 10534881 TI - The new guidelines on cardiac arrest. PMID- 10534882 TI - Perennial rhinitis. PMID- 10534883 TI - Passing the MRCGP. Preparation. PMID- 10534884 TI - The difficult asthmatic. PMID- 10534885 TI - A pill request from a troubled mother of two. PMID- 10534886 TI - Key developments in respiratory medicine. PMID- 10534887 TI - Current asthma management. PMID- 10534888 TI - When is asthma related to work? PMID- 10534889 TI - Thromboembolic disease: the new risk factors explained. PMID- 10534890 TI - Pleural disease--diagnosis and management. PMID- 10534891 TI - When should you suspect sick sinus syndrome? PMID- 10534892 TI - Unexplained chronic neurological disorders. PMID- 10534893 TI - Research and fertility regulation. PMID- 10534894 TI - Safety and efficacy of fertility-regulating methods: a decade of research. AB - An international venture was launched in 1985 to fill a recognized gap in post marketing surveillance of fertility-regulating methods. For this purpose a new task force was set up by the Special Programme of Research, Development, and Research Training in Human Reproduction, which is cosponsored by the United Nations Development Programme, the United Nations Population Fund, the World Bank, and WHO. Research priorities were chosen and epidemiological studies inaugurated, involving a total of 47 countries--mostly from the developing world. Important progress has been made, especially in helping to define the beneficial and possible adverse effects of oral contraceptives on the risk of neoplasia; in showing that the injectable contraceptive depot-medroxyprogesterone acetate protects against endometrial cancer and does not increase the overall risk of breast cancer, in clarifying which groups of women are susceptible to the rare cardiovascular complications of oral contraceptives (myocardial infarction, stroke, and venous thromboembolism); and in establishing the long-term effectiveness and safety of intrauterine devices. The research has already made a significant impact on family planning policies and practice. Critical appraisal of this venture, which has been modestly funded, confirms the value of mission oriented research. It also illustrates the potential of collaboration that bridges the global divide between developing and developed countries. PMID- 10534895 TI - Quality of vaccination services and social demand for vaccinations in Africa and Asia. AB - For immunization to be effective in the long run as a major global disease control intervention it is important to provide good quality vaccination services. Studies carried out in three countries in Asia (Bangladesh, India, and the Philippines) and two countries in Africa (Ethiopia and Malawi), and reported on in this article, document the fact that parents are willing to invest considerable effort in having their children vaccinated; however, there are a number of serious shortcomings in the quality of the routine vaccination services and strains are apparent at the interface between the vaccination providers and the users. These shortcomings are detracting from the sustainability of routine vaccination programmes and are promoting the growth of pools of nonimmunized and partially immunized children. To safeguard the continued operation and to enhance the coverage of routine vaccination programmes it is crucial that these difficulties be addressed. PMID- 10534896 TI - Serological diagnosis of human immuno-deficiency virus in Burkina Faso: reliable, practical strategies using less expensive commercial test kits. AB - Reported are the results of a cross-sectional survey in Burkina Faso to identify reliable, practical strategies for the serological diagnosis of HIV-1 and/or HIV 2 infections, using less-expensive commercial test kits in various combinations, as an alternative to the conventional Western blot (WB) test, which costs US$ 60. Serum samples, collected from blood donors, patients with acquired immunodeficiency syndrome (AIDS) and pregnant women, were tested between December 1995 and January 1997. Twelve commercial test kits were available: five Mixt enzyme-linked immunosorbent assays (ELISA), three Mixt rapid tests, and four additional tests including monospecific HIV-1 and HIV-2 ELISA. The reference strategy utilized a combination of one ELISA or one rapid test with WB, and was conducted following WHO criteria. A total of 768 serum samples were tested; 35 were indeterminate and excluded from the analysis. Seroprevalence of HIV in the remaining 733 sera was found to be 37.5% (95% confidence interval: 34.0-41.1). All the ELISA tests showed 100% sensitivity, but their specificities ranged from 81.4% to 100%. GLA (Genelavia Mixt) had the highest positive delta value, while ICE HIV-1.0.2 (ICE) produced the most distinct negative results. Among the rapid tests, COM (CombAIDS-RS) achieved 100% sensitivity and SPO (HIV Spot) 100% specificity. Various combinations of commercial tests, according to recommended WHO strategies I, II, III, gave excellent results when ICE was included in the sequence. The best combination of tests for strategy II, which achieved 100% sensitivity and specificity, was to use ICE and COM, the cost of which was US$ 2.10, compared with US$ 55.60 for the corresponding conventional strategy. For strategy III, the best combination, which achieved 100% sensitivity and specificity, was to use ICE, ZYG (Enzygnost Anti HIV-1/HIV-2 Plus) and COM, the cost of which was US$ 2.90 (19.2 times lower than the corresponding strategy requiring WB). No rapid test combination showed 100% sensitivity and specificity. Our results indicate that the serodiagnosis of HIV in Burkina Faso is possible by using reliable, less-expensive strategies which do not require Western blot testing. Moreover, there is a choice of strategies for laboratories working with or without an ELISA chain. PMID- 10534897 TI - Current clinical efficacy of chloroquine for the treatment of Plasmodium falciparum infections in urban Dar es Salaam, United Republic of Tanzania. AB - Reported is the use of a 14-day WHO protocol, which takes into account the clinical, parasitological and haematological responses to antimalarial drugs, to determine the efficacy of chloroquine in the treatment of uncomplicated malaria in young children (n = 200) in urban Dar es Salaam. Chloroquine failure was found in 43% of the children. Of these, 12.5% were considered to be early treatment failures and were given a single dose of sulfadoxine-pyrimethamine. Fever subsided in all children treated with sulfadoxine-pyrimethamine and there were no parasitological failures. In addition, children treated with sulfadoxine pyrimethamine because of early treatment failure with chloroquine had better haematological recovery than the chloroquine-sensitive group. It is concluded that chloroquine can no longer be considered an effective therapy for P. falciparum malaria in young children in Dar es Salaam. PMID- 10534898 TI - Use of polymerase chain reaction in human African trypanosomiasis stage determination and follow-up. AB - Stage determination of human African trypanosomiasis is based on the detection of parasites and measurements of biological changes in the cerebrospinal fluid (CSF) (concentration of white blood cells > 5 cells per mm3 and increased total protein levels). The patient is treated accordingly. Demonstration of the absence or presence of trypanosomes by the double centrifugation technique is still the only test available to clinicians for assessing treatment success. In this study, however, we evaluate the polymerase chain reaction (PCR) as a tool for assessing the disease stage of trypanosomiasis and for determining whether treatment has been successful. All 15 study patients considered to be in the advanced stage of the disease were PCR positive; however, trypanosomes were demonstrated by double centrifugation in only 11 patients. Of the five remaining patients, who were considered to be in the early stage, PCR and double centrifugation were negative. Following treatment, 13 of the 15 second-stage patients were found to be negative for the disease in at least two samples by PCR and double centrifugation. Two others were still positive by PCR immediately and one month after the treatment. Trypanosome DNA detection using PCR suggested that the two positive patients were not cured but that their possible relapse could not be identified by a search for parasites using the double centrifugation technique. Further evaluation of the PCR method is required, in particular to determine whether PCR assays could be used in studies on patients who fail to respond to melarsoprol, as observed in several foci. PMID- 10534900 TI - Toxic effects of mycotoxins in humans. AB - Mycotoxicoses are diseases caused by mycotoxins, i.e. secondary metabolites of moulds. Although they occur more frequently in areas with a hot and humid climate, favourable for the growth of moulds, they can also be found in temperate zones. Exposure to mycotoxins is mostly by ingestion, but also occurs by the dermal and inhalation routes. Mycotoxicoses often remain unrecognized by medical professionals, except when large numbers of people are involved. The present article reviews outbreaks of mycotoxicoses where the mycotoxic etiology of the disease is supported by mycotoxin analysis or identification of mycotoxin producing fungi. Epidemiological, clinical and histological findings (when available) in outbreaks of mycotoxicoses resulting from exposure to aflatoxins, ergot, trichothecenes, ochratoxins, 3-nitropropionic acid, zearalenone and fumonisins are discussed. PMID- 10534899 TI - Adaptation of cytochrome-b5 reductase activity and methaemoglobinaemia in areas with a high nitrate concentration in drinking-water. AB - An epidemiological investigation was undertaken in India to assess the prevalence of methaemoglobinaemia in areas with high nitrate concentration in drinking-water and the possible association with an adaptation of cytochrome-b5 reductase. Five areas were selected, with average nitrate ion concentrations in drinking-water of 26, 45, 95, 222 and 459 mg/l. These areas were visited and house schedules were prepared in accordance with a statistically designed protocol. A sample of 10% of the total population was selected in each of the areas, matched for age and weight, giving a total of 178 persons in five age groups. For each subject, a detailed history was documented, a medical examination was conducted and blood samples were taken to determine methaemoglobin level and cytochrome-b5 reductase activity. Collected data were subjected to statistical analysis to test for a possible relationship between nitrate concentration, cytochrome-b5 reductase activity and methaemoglobinaemia. High nitrate concentrations caused methaemoglobinaemia in infants and adults. The reserve of cytochrome-b5 reductase activity (i.e. the enzyme activity not currently being used, but which is available when needed; for example, under conditions of increased nitrate ingestion) and its adaptation with increasing water nitrate concentration to reduce methaemoglobin were more pronounced in children and adolescents. PMID- 10534901 TI - Some thoughts on ICPD+5. PMID- 10534902 TI - Integrating reproductive health: myth and ideology. AB - Since 1994, integrating human immunodeficiency virus/sexually transmitted disease (HIV/STD) services with primary health care, as part of reproductive health, has been advocated to address two major public health problems: to control the spread of HIV; and to improve women's reproductive health. However, integration is unlikely to succeed because primary health care and the political context within which this approach is taking place are unsuited to the task. In this paper, a historical comparison is made between the health systems of Ghana, Kenya and Zambia and that of South Africa, to examine progress on integration of HIV/STD services since 1994. Our findings indicate that primary health care in Ghana, Kenya and Zambia has been used mainly by women and children and that integration has meant adding new activities to these services. For the vertical programmes which support these services, integration implies enhanced collaboration rather than merged responsibility. This compromise between comprehensive rhetoric and selective reality has resulted in little change to existing structures and processes; problems with integration have been exacerbated by the activities of external donors. By comparison, in South Africa integration has been achieved through political commitment to primary health care rather than expanding vertical programmes (top-down management systems). The rhetoric of integration has been widely used in reproductive health despite lack of evidence for its feasibility, as a result of the convergence of four agendas: improving family planning quality; the need to improve women's health; the rapid spread of HIV; and conceptual shifts in primary health care. International reproductive health actors, however, have taken little account of political, financial and managerial constraints to implementation in low-income countries. PMID- 10534903 TI - ICPD and its aftermath: throwing out the baby? PMID- 10534904 TI - Cost-effective HIV treatment for developing countries. PMID- 10534906 TI - Clinico-polysomnographic diagnostics of narcolepsy-cataplexy. AB - INTRODUCTION: The introduction of polysomnography changed considerably our knowledge about narcolepsy. The aim of our study was to present the clinical picture and diagnostic criteria of narcolepsy based on our own research data. MATERIAL AND METHODS: The study comprises 22 consecutive patients with a mean age of 39.8 years, SD = 16.4, age range 7-78 years. The following methods were used: neurological, physical and mental status examination, specially designed questionnaires, nocturnal polysomnography, MSLT. RESULTS: The following clinical manifestations were observed: excessive daytime sleepiness (EDS) in 95.45% of the cases, cataplexy attacks (CA) in 77.27%, hypnagogic hallucinations (HH)--in 77.27%, sleep paralysis (SP)--in 54.54% and subjectively disturbed nocturnal sleep--in 72.72%. The mean ages of occurrence of the respective symptoms were as follows: EDS--32.14 +/- 10.15 years, CA--28.70 +/- 11.70 years, HH--36.70 +/- 8.82 years, SP--35.53 +/- 15.50 years and for the disease as a whole--39.82 +/- 15.50 years. At MSLT the mean sleep latency was 165.91 +/- 16.37 seconds and the mean REM latency--207.96 +/- 14.09 seconds. During the nighttime sleep the mean sleep latency was less than 5 minutes and the mean REM latency--less than 7 minutes. The sleep structure showed: NREM sleep, stage 1,2--66.30 +/- 4.1%, stage 3,4--5.52 +/- 4.1%, REM sleep--25.65 +/- 2.3%. The clinical picture presented 4 symptoms in 31.81%, 3 symptoms--in 50.00%, 2 symptoms--in 9.09% and 1 symptom in 9.09% of the cases. CONCLUSIONS: 1. The onset of narcolepsy is at about 30 years of age. 2. The clinical picture of narcolepsy is characterized by 2 major symptoms--excessive daytime sleepiness, attacks of cataplexy and 3 minor symptoms -sleep paralysis, hypnagogic hallucinations and disturbed nighttime sleep which may be found in various combinations. 3. Nocturnal polysomnography demonstrates characteristic changes in sleep structure. 4. MSLT ascertains shorter sleep latency and sleep onset rapid eye movements periods (SOREMs) of less than 5 minutes. 5. The presence of one of the major symptoms and SOREMs is sufficient for confirming the diagnosis of narcolepsy. PMID- 10534905 TI - Impact of home gardening and nutrition education in a district of rural India. PMID- 10534907 TI - Third malignancy after treatment of Hodgkin's disease. AB - We present a case of 36-year-old man who was treated for IIB supradiaphragmatic lymphocyte-predominant Hodgkin's disease in 1972 (at the age of 11). The patient remained relapse-free after combined radiotherapy (irradiation of the right supraclavicular field) and chemotherapy (six MOPP cycles) and a 3-year supporting chemotherapy (Velbe). In 1993 he underwent spinal cord surgery for a right-sided "hour glass" schwannoma at C4-5 level. In 1996 a large formation histologically verified as "chondrosarcoma" occurred in the right supraclavicular and axillary regions. The pathogenesis of the second and third malignancies may be attributed to the histologic pattern of HD with long-term survival and increased cumulative risk, non-alternating MOPP courses and continued supporting therapy and radiotherapy given to the involved fields. PMID- 10534908 TI - Surgical complications in hematological malignancies. AB - Surgical complications in oncohematological practice appear a primary problem for their high incidence, diagnostic and therapeutic difficulties and unfavourable prognosis. The authors present data from a retrospective study of 93 patients from the Clinic of Hematology and Surgical Clinic of the Higher Medical Institute in Plovdiv for the period from 1990 to 1997. Surgical complications occurred in 9.9% of the patients treated for hematological malignancies. Most frequent and hazardous were the cases of acute abdomen, followed by soft tissue purulent infections. The authors suggest that high risk of surgical complications exists in patients undergoing vigorous chemotherapy. Patients are particularly vulnerable in the neutropenic period after chemotherapy when the abdominal and anorectal complications may be fatal. PMID- 10534909 TI - Myotonic dystrophy: a report of two cases from one family. AB - The authors report two cases of a brother and sister who suffered myotonic dystrophy and had a family history of the disease. Muscular dystrophy--more severe in the boy and mild in the girl--was the clinical manifestation. EMG findings corresponded to a primary muscle disorder, coinciding with myotonic activity in case 1. The genealogical study of four generations of this family revealed a descent pattern of inheritance. Anticipation--a characteristic phenomenon of the disease expressed in the occurrence of the disorder at earlier ages and in more severe forms in the successive generations--was also observed. PMID- 10534910 TI - Anatomo-topographic landmarks on the lateral wall of nasal cavity used in endonasal surgery. AB - INTRODUCTION: A dissection of the lateral wall of nasal cavity that imitates a true endonasal operation was the design of the present study. The object of the study was by measuring the distance between a pivot landmark and important anatomical structures revealed successively during an endonasal operation to create additional highlights that will minimise the hazard of injuring vital structures. METHODS: Twenty anatomical preparations of sagittally sectioned heads from cadavers (elderly males of Bulgarian origin) were dissected to study the topographic relationships between the great paranasal cavities (maxillary, frontal, and sphenoidal sinus) and the ethmoidal labyrinth. Using punctum subnasale as a pivot landmark, the distances to crucial microendoscopic landmarks (i.e., nasolacrimal duct, uncinate process, ethmoidal bulla, base of the skull, anterior wall of the sphenoid sinus, orifice of the maxillary sinus) were measured at different angles to the horizontal plane. RESULTS: The mean distances from punctum subnasale to the natural ostium of the maxillary sinus and anterior wall of the sphenoidal sinus, measured at an angle of 30 degrees to the floor of the nasal cavity, were 48.25 +/- 0.75 and 67.105 +/- 0.794 mm, respectively. At an angle of 45 degrees, the mean distance to the nasolacrimal duct was 38.056 +/- 0.591 mm; to the uncinate process 45.25 +/- 0.57 mm; to the ethmoidal bulla 50.25 +/- 0.57 mm; and to the base of the skull 66.053 +/- 0.818 mm. Mean distance of 43.158 +/- 0.568 mm to the lacrimal sac and 60.25 +/- 0.68 mm to the base of the skull were measured at an angle of 60 degrees. CONCLUSION: The analysis of the results imply that the selected distances will contribute to a more precise application of this contemporary surgical method. PMID- 10534911 TI - Microendoscopic endonasal approach in treating diseases of nasal cavity, paranasal sinuses, and adjacent structures--a preliminary communication. AB - Since the beginning of 1997 the authors has introduced routinely in their practice a method of combined nasal endoscopy and microrhinoscopy in the diagnosis and minimally invasive surgery of diseases of nasal cavity, paranasal sinuses, and adjacent structures. The results of the method applied in 25 patients (19 males and 6 females) aging from 2 months to 77 years with doubtful initial diagnosis are analysed. In 17 of the patients the initial diagnosis was confirmed, in 5 the diagnosis was revised, and in 3 no pathology of the nasal cavity and paranasal sinuses was found. In 13 patients minimally invasive surgery was successfully performed using this modern method. The results obtained are compared to the related literature data. PMID- 10534912 TI - A model of a computer file for the intensive neonatal practice "Neonatal Intensive Computer File". AB - INTRODUCTION: The use of computer methods for processing the databases gathered in a neonatal intensive care unit has more than 20 years experience. During the past years various systems for data management in accordance with the nature of the work have been designed. AIM: The aim was to create a method for quick and easy processing of variable clinical material in an uniform format. REQUIREMENTS: The method allows the arrangement of the entire primary information into a standardized database suitable for computer processing, following up an unlimited number of parameters, calculating derivative indices, representing and evaluating data by means of graphic trends, surveying current clinical information not only for routine practice, but also for research purposes. RESULTS: We developed a computer file type method. This is a method enabling the systematic and well arranged structuring of the newborn's history, the recordings of the monitoring and life-supporting equipment, the data from clinical, laboratory examinations and all the other implemented diagnostic-therapeutic procedures, family, social and possibly legal background. CONCLUSION: The proposed method Neonatal Intensive Computer File (NICF) was applied successfully in the management the data of the above mentioned project (Grant L-444, MES-Sofia): organizes the entire information into a form suitable for the everyday clinical practice and for research studies. Neonatal Intensive Computer File (NICF) gives ample possibilities for quick and reliable evaluation of the condition of the newborn infant and the effect of the applied treatment. The introduced panel (complex) of indices with possibilities for its expansion allows early detection of advancing and threatening disturbances, characterizes the type and pinpoints the exact time and location of the disturbances occurring in the system ventilator-patient physician. The graphic trends visualize the variation of the indices and contributes to the quick evaluation and orientation in them. They are considerably more informative than the single time registered values of any indicator or index. Optimizes the management of the applied ventilation/oxygenation technique and overall treatment, facilitates the timely decision-making process (the change and/or the choice of new therapeutic alternatives) Facilitates the statistical analysis of the total accumulated data or a selected part of it. Allows the use of the same clinical material for the needs of multiple research projects guaranteeing precision and reliability. The accumulated databases (information bank) are important for future prospective and retrospective studies not only in the clinical but also in the social field. The information yielded by Neonatal Intensive Computer File (NICF) can be used for prognostic projections and conclusions in terms of the current condition and outcome as well as ad vitam. PMID- 10534913 TI - Comparative study of the incidence and prevalence of dental caries in 12-year old children from Plovdiv. AB - INTRODUCTION: The necessity of refreshing the data determines the purpose of the present study--to study the incidence and prevalence of dental caries in 1997 and to compare the results with the results of previous studies. METHODS: The study of dental caries is clinico-statistical and comprises 200 12-year-old children from Plovdiv. The sample selection is random. The diagnosis of dental caries is visual-tactile. The incidence (DMFT) and prevalence of dental caries is determined by person (Ep). The results are compared with the results of studies of children of the same age group carried out in 1981, 1989, 1992. RESULTS: The incidence of caries in 12-year old children from Plovdiv is 3.03 and the prevalence rate by persons is 80.0%. Comparing the results with previous studies shows a trend of steadily decreasing values with 1992 exhibiting the lowest levels of these indicators. A trend of increasing DMFT and Ep between 1992 and 1997 is observed. CONCLUSIONS: The trends of incidence and Ep in 12-year old children in Plovdiv show the favorable effect of the preventive programs till 1992 and the necessity of undertaking urgent measures for overcoming the unfavorable trends set in the last years. PMID- 10534914 TI - Some chronobiological characteristics of cerebral strokes. AB - INTRODUCTION: The chronobiological studies of cerebro-vascular diseases, including acute cerebro-vascular incidents are scanty. METHODS: The authors study retrospectively the case histories of 352 patients with CS and lethal outcome during a 5-year period /1991-1995/. The data was analyzed statistically by determining percentages and standard errors and using graphical analysis. RESULTS: The peaks of mortality rate of cerebral stroke coincide with that of the incidence rate. The onset of ICS is most frequently in the hour intervals 8-12 and 14-17 o'clock, while of HCS--8-12 o'clock. The lethal outcome from ICS is predominantly in the hour interval 5-7 o'clock, while of HCS--4-9 o'clock. CONCLUSIONS: The hour of onset and the hour of death from ischemic and hemorrhagic cerebral stroke have discrepant peak hours. PMID- 10534915 TI - Effect of complex sanatorium treatment on some dyspnea indices in patients with chronic obstructive pulmonary disease. AB - The complex treatment of patients with chronic obstructive pulmonary disease (COPD) administered in a sanatorium environment is an effective therapeutic option for this condition as it involves conduction of a combination of climatic, therapeutic and rehabilitation procedures for a longer period of time. We studied the effect of this therapeutic modality on the dyspnea indices of COPD patients and the implication these indices have for the outcome of the treatment. The study was performed in the sanatorium of the State Hospital for Lung Diseases in Raduntzi, Bulgaria. It included 75 patients (65 men, 11 women) with different forms of stable COPD (mean age, 56.8 +/- 1.0 years, mean +/- SEM, FEV1% predicted -37.3 +/- 1.6%, mean duration of sanatorium stay 14 +/- 0.4 days). During the stay the patients received anti-obstructive and anti-inflammatory therapy based on the clinical discretion of the attending physician. All patients attended a rehabilitation programme according to their conditions; oxygen therapy was used in three patients. Of the 75 patients, 50 (66.7%) showed improvement at discharge, 21 (28%) had no improvement, and 4 (5.3%) were discharged in deteriorated condition. The complex treatment resulted in a minimal but statistically significant improvement of the basic spirographic and dyspnea parameters. This improvement depended not so much on the type of therapy administered as on the initial dyspnea and blood gas parameters. The body mass index (BMI) can also be used as a prognostic indicator especially if it is lower than 20 kg/m2. The lower this index is the lower the basic functional parameters are--in spite of using all available treatments, our patients with low body mass index rarely showed any noticeable improvement. PMID- 10534916 TI - Ultrasound follow-up study of arthroscoped patients with gonitis. AB - The purpose of the present study was to evaluate by ultrasound methods and carry out a one-year follow-up study of synovial proliferation in the knee joint of patients with rheumatoid arthritis prior to and after arthroscopy and arthroscopic (AS) synovectomy. MATERIAL AND METHODS: 24 patients with a proven rheumatoid arthritis and affected knee joints were recruited for the study. Arthroscopic synovectomy of one of the affected joints was performed in all of them. The synovitis was evaluated clinically and ultrasonographically in 24 knee joints prior to the AS synovectomy and 7 days, and 3, 6 and 12 months after it. RESULTS: Either synovial thickening or villonodular proliferation in the knee joints were the findings in the examined patients using arthrosonography. All patient showed improvement of the disease activity index after the arthroscopic synovectomy and weakening of the baseline ultrasound evidence for synovial thickening and villonodular proliferation (p < 0.001). During the 12-month follow up study two patients were found at ultrasonography to have (without any clinical evidence for that) recurrence of the synovial thickening three months after arthroscopy. Ten patients had synovial thickening or villonodular proliferation recurrence during the 12 months of follow up with clinical evidence of gonitis; one case showed ultrasonographic evidence of hydrops. There was a complete consistency between the ultrasound and arthroscopic protocols with respect to the presence of synovial proliferation and intra-articular effusion. CONCLUSION: Arthosonography is an easy, safe, low-cost, non-invasive modality for diagnosis and follow-up of patients with rheumatoid arthritis prior to and after arthroscopic synovectomy and for assessment of the clinical prognosis in such patients. PMID- 10534917 TI - An attempt to elucidate the concept of "folk dietary tradition". AB - Nutrition is a complex interdisciplinary science. It is an intersection point of a number of other sciences such as biology, economics, technology, ethnography, medicine, history, linguistics, politics. Folk dietary tradition is a normative nutritional system that is used by an already formed community. As no definition is available we analysed the term and, on the basis of our extensive research, came to the following conclusion: folk dietary tradition is constituted together with the establishment of the community for the purpose of consolidating it as well as for differentiating it from other communities; it reflects the historical development and the destiny of the specific community; it changes, becomes enriched or poorer, or loses its originality; it can be formed using previous, local dietary traditions that precede the formation of a community, or can be borrowed by other peoples which are in the process of introducing a new religion. During its golden age Bulgaria introduced its folk dietary tradition to the peoples converted by Bulgaria to Christianity. In the present study I give a definition of the basic characteristics of the Bulgarian folk dietary tradition. This definition can be used to analyse all folk dietary traditions and make comparison between them. PMID- 10534918 TI - Basic principles of Bulgarian folk dietary traditions. AB - Bulgarian folk dietary tradition is one of the most ancient traditions in the world. Its origin can be traced back to prehistoric times. The contribution of ancient Thracians to it, for instance, with respect to some major foods, preparation of foods, practices and customs, is so deeply ingrained in it that to a large extent it determines its overall character. And this is actually an all European legacy. The Proto-Bulgarian legacy in the field of nutrition is also considerable. Of particular importance here is the correlation of meals with time and with the so called good and bad periods of time. In the present study we have attempted to formulate some of the important principles of the Bulgarian folk dietary tradition. They are only a small part of the vast realm of principles concerning the diet of Bulgarians. All Bulgarian customs, rules and bans in the field of nutrition are based on them. Until quite recently they have had an obligatory character. They have coded in them the thousand-year-long experience of the Bulgarian people. All important rational aspects repeatedly verified in real life under different conditions and situations have been included in them. Thus a complete system of rules and norms of behaviour have been obtained giving exhaustive answers to all questions related to nutrition. It is designed to help people and future generations to avoid risk situations and prevent catastrophes resulting from malnutrition. PMID- 10534920 TI - [Ways of delivery after cesarean section]. AB - OBJECTIVE: To analyse deliveries after a previous cesarean section. STUDY DESIGN: The way of birth of 262 patients between 1993-1995 in the Ist Department of Obstetrics and Gynaecology of Warsaw University Hospital was analysed. Material was divided into two groups: an elective cesarean section operation was performed on 117 women, while 145 women underwent a trial of labour. RESULTS: The most common indications for an elective cesarean section were fetal distress and maternal diseases. A trial of vaginal delivery was successful in 55.2%. The repeated operation was performed on 44.8% of women. The most frequent indication for a cesarean section during a trial of labour was failure to progress. CONCLUSIONS: Women after a previous cesarean section, who underwent a trial of labour delivered vaginally in 55.2%-30.5% of all the patients in the analysed material. The indications for the repeated operation were the same as for the first one in 27% of the cases. PMID- 10534919 TI - [Analysis of perinatal death at the Institute of the Health Center of the Polish mother in 1995, 1996 and 1997. The reason for making changes in the accountability for perinatal death]. AB - Perinatal death's causes of fetuses and newborns from single and twin pregnancies delivered at the PMMHI from 1995-1997 were discussed. Data from the Pathology Department were analysed and compared to information regarding prenatal US + ECHO diagnoses coming from the Department for Diagnoses of Congenital Malformations at the PMMHI. The most frequent cause of death of fetuses and newborns from single pregnancies were congenital malformations (42%). In twins there prevailed such typical for multiple pregnancies' death causes as TTTS (27%), intrauterine demise of one of the twins (17%). Premature labor occupies the second most frequent cause of death both in single and multiple pregnancies. Most of perinatal deaths may be predicted prenatally by means of ultrasound and fetal echocardiography. PMID- 10534921 TI - [Changes in arylsulphatase activity (EC 3.1.6.1) in the blood serum and urine in women during pregnancy and in the course of delivery]. AB - By the determination of arylsulphatase A activity (EC 3.1.6.1) in the blood serum and urine obtained from 66 women using the modified method by Lee-Vaupel and Conzelmann it was noticed the increase in the enzyme activity during the pregnancy comparing to the non-pregnant group. The highest enzyme activity was observed in the III trimester of pregnancy. In the following stages of delivery (I, II, III) it was assumed the increase in enzyme activity in urine. The highest enzyme activity in urine was observed in the stage III, and in the serum--in the stage II. It was compared the enzyme activity in primiparae and multiparae proving, that in the serum nd urine this activity is higher in the stages I and II in multiparae, and in the stage III in primiparae. PMID- 10534922 TI - [The concentration of alpha-2-antiplasmin in sera of women in post partum]. AB - Alpha-2-antiplasmin is a main inhibitor of plasmin and play a crucial role in regulation of intravascular fibrinolysis. It is important to know, in pregnant women as well in the immediate post partum period the activation of hemostasis takes place and is observed thrombotic incidences. In our studies we have included 33 pregnant women whose delivered eutrophic newborns alpha-2-AP 7.79 mg/dl and 36 pregnant women whose delivered small-for-date newborns, alpha-2-AP 7.49 mg/dl. We found no significant statistically differences between tested groups and in comparison to control group 6.0 mg/dl. The presented results seems to indicate, the determination of concentration of alpha-2-AP in sera of women in immediate post partum period does not reflect the changes in the mechanisms of fibrinolytic system in the course of the partum. PMID- 10534923 TI - [The influence of hormonal replacement therapy on the certain collagen metabolites]. AB - RATIONALE OF THE STUDY: Collagen is the most abundant protein within the human body. It constitutes about 80% of the organic part of all tissues. Type I collagen constitutes 95% of the organic part of the bone. Therefore the measuring of the carboxyterminal telopeptide of type I collagen (ICTP), cross-linked via pyridinoline and liberated during the degradation of type I collagen, enables monitoring of the bone collagen degradation. Biosynthesis of collagen includes several posttranslational modifications, among them the propeptides cleavage from the N and C terminal parts of the molecule. Measuring of these products is a useful indicator of the collagen biosynthesis rate. THE AIM OF THE STUDY: To evaluate the influence of hormonal replacement therapy on degradation of type I collagen as measured by ICTP and biosynthesis of type III collagen as measured by PIIINP (N terminal propeptide of type III collagen). MATERIAL AND METHODS: The above mentioned markers of collagen metabolism were measured in sera of 25 women (mean age 49.12 +/- 3.01 years) who underwent abdominal hysterectomy with bilateral salpinpo-ophorectomy because of benign diseases. The control group included 25 women matched for age and body mass who did not receive any treatment. Immediately after surgery HRT constituted of Estraderm TTS50 and Provera 10 mg sequentially was established in every patients. Blood for biochemical investigations was collected before surgery and after 6 and 12 months of the therapy. Serum ICTP and PIIINP concentrations were measured by RIA method using commercially available kits from Orion Diagnostica-Finland. RESULTS: It was found that HRT significantly decreased serum ICTP concentrations and this was accompanied by the increase in the biosynthesis of type III collagen as measured by PIIINP. Moreover, a strong negative correlation was found between bone mineral density and ICTP in both investigated groups. CONCLUSIONS: 1) HRT significantly decreased type I collagen breakdown as measured by ICTP concentrations. 2) Type III collagen biosynthesis as measured by PIIINP concentration steadily increased during HRT. 3) The diagnostic use of ICTP measurement altogether with the bone mineral density could be a useful predictor for the risk of the bone fracture during menopause. PMID- 10534924 TI - [Y-specific sequences in Turner syndrome]. AB - Turner Syndrome (TS) is the only one monosomy that occurrs+ in humans. The cytogenetics of TS is very well known from years. It has been estimated that almost 98-99% of TS foetuses end in abortion. It was suggested that the monosomy arises relatively late during embryonal development and survived TS individuals could be mosaics. It has been proved that mosaic karyotype mos 45,X/46X, + mar(Y) occurrs++ in 2% to 11% of TS patients. The patients having additional cell line containing der(Y) are at increased risk of gonadoblastoma development. In these cases gonadectomy should be considered. Therefore detection of mosaic and establishing the origin of marker chromosome (specially containing Y-specific sequences) is of special importance. The aim of present study was to detect the small mosaics, containing mar(Y) in TS patients, by using PCR and FISH techniques. Eight Y sequences for the PCR analyses as well as bicolor in situ hybridisation with painting probes for Y and X chromosomes have been applied. The positive amplification for Y-specific sequences has been detected in 7% of TS patients. Our results support the thesis that searching for the Y sequences should be introduced to routine genetic TS diagnosis. PMID- 10534925 TI - [Vulvar lichen sclerosus in girls treated locally with clobetasol propionate]. AB - Vulvar lichen sclerosus is rare disease in children. 5 girls (age 5-16) with LS were treated locally with clobetasol propionate. All patients showed clinical improvement or full regression of symptoms without side effects. PMID- 10534926 TI - [Heterotopic osteogenesis in postoperative care in ovarian cancer treatment]. AB - The case of calcification in postoperative scare in patient with ovarian cancer was described. The patient was operated because of ovarian cancer and had subsequently chemiotherapy. Afterwards the second look operation was performed which showed calcification (size 9.5 x 2 x 0.4 cm) in postoperative scare. The calcification wasn't a metastasis from the ovary. The etiology of calcification in unclear. The heterotopic osteogenesis in postoperative scars is quite rare. They usually appear as a result of calcification of connective tissue. Sometimes the relation between osteogenesis and neoplasmatic disease is supposed. Till now the reason of these kind of calcification is still unknown. PMID- 10534927 TI - [A case of extended hysterectomy in stage IB-1 cervical cancer after aorto bifemoral reconstruction]. AB - We describe a cervical cancer case stage IB-1 according to FIGO classification surgically treated with extended hysterectomy and lymphadenectomy. The patient was four years after aorto-femoral Dallon bypass on account of Leriche syndrome and from this time she was treated with oral anticoagulant therapy. During the routine diagnostic investigations the invasive cervical cancer IB-1 was diagnosed. According to the oncological protocol she has had extended hysterectomy Piver III type and partial pelvic lymphadenectomy. Extended hysterectomy did not cause technical troubles. Pelvic lymphadenectomy was only partially possible to realizing. Compact connective tissue around the artificial vessels did not permit safe preparation of pelvic lymphatic system. PMID- 10534928 TI - [Metabolism of mevalonate in human placenta]. AB - Prenylation of protein rather than cholesterol synthesis, seems to be main "metabolic target" of mevalonate metabolism in the human placenta. It seems very probable that cytological transformation of cytotrophoblast cells to syncytiotrophoblast which occur during placental development, "switched off" activity of mevalonate biosynthesis in term placenta. PMID- 10534929 TI - [The influence of hormone replacement therapy containing transdermal 17-beta estradiol and oral medroxyprogesterone acetate on coagulation and fibrinolysis]. AB - OBJECTIVE: The aim of the study was to evaluate some coagulative and fibrinolytic changes in women taking hormone replacement therapy (HRT). MATERIALS AND METHODS: In 29 women aged 45-64 years with osteoporosis and climacteric symptoms several fibrinolytic and coagulative parameters were measured. These measurements were performed three times in each women--before and after three and six months of HRT containing transdermal 17 beta-estradiol (50 mg per day) and oral medroxyprogesterone acetate (2.5 mg per day). RESULTS: PAI-1 (antigen) and factor VII (activity) were decreased significantly during the trial. No other significant modifications in the examined parameters (fibrinogen, antithrombin III, protein C, protein S, thrombin-antithormbin III complexes, t-PA and D dimers) were detected. CONCLUSIONS: The hormone replacement therapy can reduce the risk of coronary heart disease thanks to decreased levels of PAI-1 and factor VII. No other changes in coagulation and fibrinolysis were observed during the treatment. PMID- 10534930 TI - [Diagnostic micro-laparoscopy in gynecology]. AB - Diagnostic microlaparoscopy was performed on 30 patients aged 17-41. Indications++ for these procedures were: sterility (17), tumor of the ovary (5), endometriosis (3), uterine myoma (2), pelvic pain syndrome++ (1), carcinoma of the ovary--second look procedures (1), operative hysteroscopy (1). During these procedures we examined small pelvis and 17 patients were additionally undergone chromotubation. These procedures lasted 8-24 minutes. No complications were noticed during and after operations. Microlaparoscopy were verified by laparoscopy with positive correlation of data. Advantages and limitations of microlaparoscopy were taken into consideration. We presented possibilities of the new diagnostic technique in gynaecology. PMID- 10534932 TI - [The role of cytogenetic studies in prognosis of treatment results in patients with ovarian cancer]. AB - The aim of this study was to determine the correlation between chromosome aberrations complexity in carcinomatous cells and survival in 38 patients with ovarian cancer. The material of cytogenic studies were 38 specimens taken from primary neoplastic ovarian tumors, which were than crumbled enzymatically. The suspension of neoplastic cells was used to set up primary cell culture in vitro. Only numerical aberrations or one of structural aberrations were assumed to be simple aberrations. More than one of numerical aberrations or structural aberrations were thought to form complex aberrations. The curve of survival in patients were determined using nonparametrical method of Kaplan and Meier and the differences between the curves were compared using logrank test. CONCLUSIONS: 1. The existence of complex chromosome aberrations in ovarian cancer cells is unfavourable prognostic factor. 2. The cytogenic analysis of neoplastic ovarian tumors together with numerous clinical and histological factors is an important prognostic factor. PMID- 10534931 TI - [The correlation between clinical and histological features and complexity of chromosomal changes in primary ovarian cancer]. AB - The aim of the study was to determine the correlation between the complexity of chromosome aberrations in ovarian cancer and its clinical stage and histological grade. 38 primary ovarian cancer tumors were analysed cytogenetically. The chromosomes were obtained from primary cell cultures in vitro and were stained using GTW technique. Advanced clinical stage correlated with complex chromosomal rearrangements. Similarly, histologically poorly differentiated tumors more often displayed complex chromosomal rearrangements (p = 0.004). Simple chromosome aberrations or normal karyotype were more frequently detected in well differentiated tumors. The analysis of the results revealed that the increase in histological grade and clinical stage correlates with the increase in the number and complexity of chromosomal rearrangements. PMID- 10534933 TI - [Successful pregnancy term of a patient treated for congenital urinary bladder exstrophy in childhood]. AB - The case of successfully terminated pregnancy and labour at 32 years old patient of her multiple surgical procedure for congenital bladder extrophy at her childhood was described. Procedures during pregnancy and labour course were presented. PMID- 10534935 TI - Voluntary and involuntary sterilization: medical, ethical, legal and religious aspects. AB - Surgical voluntary sterilization has become one of the most widely used methods of contraception, with vasectomy and tubal sterilization being the most commonly employed techniques, associated with a low failure, morbidity, mortality, and long-term sequelae rate. As sterilization is related with the elimination of the possibility for procreation, a number of ethical, legal and religious issues have arisen, leading often to personal misjudgements, legal disputes, and failures in applying family planning. Involuntary sterilization is currently not practiced, except in cases of severely mentally retarded people, who are unable to appreciate the consequences of their acts or care for their children and who may have a high likelihood of propagating hereditary disease. PMID- 10534934 TI - [Successful pregnancy after conservative surgery and chemotherapy for dysgerminoma of the ovary: case report]. AB - OBJECTIVES: Considering the important improvement of surgical techniques and chemotherapy in the last few years, it is possible today, in selected cases of patients previously treated for ovarian cancer, to support their desire for motherhood, thus improving the quality of life for them. The major problem for the Gynecologic Oncologist in treating young women for ovarian tumor is the lack of statistically significant experience world-wide, because of the very few cases in which the reproductive function is preserved, and pregnancy is subsequently possible. STUDY DESIGN: The aim of this study was presentation the successful pregnancy after conservative surgery and chemotherapy for dysgerminoma of the ovary stage Ia. RESULTS: The patient age 23 was admitted to Department of Gynecology & Obstetric in Hospital of Slupsk in 1995 year with diagnosis right ovarian tumor. Right-side adnexectomy was performed initially. Histopathological examination of surgical specimen revelated dysgerminoma ovary stage Ia. Due to the clearly elevated tumor marker levels, indicating an involvement of other germ cell elements, and necrobiosis of tumor, we opted for postoperative chemotherapy instead of radiotherapy. Six cycles of BEP protocol chemotherapy was given. The follow-up included regular monitoring of the tumor markers. In the year 1998 the patient conceived and had a delivery of a normal infant at term. To date, there has been no indication towards tumor recurrence. CONCLUSION: After adequate staging and accurate information is given to the patient, conservative treatment may be safe in some women with early ovarian cancer. In early stage pure ovarian dysgerminoma conservative surgery combined with radiotherapy or chemotherapy showed high complete remission rates and excellent survival rates. For younger patients and those with gestation desires as well as patients with advanced diseases, adjuvant chemotherapy following surgery might be a better choice. PMID- 10534936 TI - [The report of UNICEF/WHO Joint Consultation on iron deficiency anemia,Geneva, February 3-5, 1999]. PMID- 10534937 TI - Air travel and chronic respiratory diseases. PMID- 10534938 TI - Pattern of opportunistic pulmonary infections in HIV sero-positive subjects: observations from Pondicherry, India. AB - Prospective analysis for a period of six-and-a-half years was done in 190 patients with HIV infection, which showed post-primary tuberculosis with sputum positive for acid-fast bacilli in 65% of cases. Extrapulmonary forms of tuberculosis especially lymph nodes infection was more frequent. Cervical group of lymph node involvement was the commonest presentation. Procedures such as FNAC/biopsy of lymph nodes and pleura provided the immediate diagnostic yields. These procedures must be considered early in the course of illness of HIV infected patients with suspected extrapulmonary and disseminated tuberculosis. Tuberculosis constitutes a common pathology with an appreciable mortality in disseminated subjects. Majority of patients with tuberculosis responded to 2EHRZ/7HR therapy indicating infection by Mycobacterium tuberculosis rather than by atypical mycobacteria, without any serious adverse reactions. Retrospective analysis of two groups (February 1991-May 1994) and (June 1994-October 1997) shows a significant increase in disseminated tuberculosis and Pneumocystis carini pneumonia indicating late stage of HIV disease. PMID- 10534939 TI - Pulmonary sarcoidosis in a south Indian hospital: clinical and lung function profile. AB - Twenty-two histopathologically proven cases of sarcoidosis were analyzed to determine the clinical presentation, lung function and the response to treatment. Laboratory data, chest x-ray and pulmonary function tests (PFT) were analyzed. Sarcoidosis was found to be more common in females in this study. Cough, breathlessness and weight loss were the predominant symptoms. Serum angiotensin converting enzyme (SACE) was elevated in 50% patients. Comparison of chest radiographs and PFT at the time of diagnosis revealed that stage I disease was associated with normal pulmonary function, 50% patients with stage II disease had mildly impaired PFT and 75% patient with stage III disease had severely impaired PFT. The indication for oral steroid treatment was respiratory symptoms in 58.8% of cases. Of the 13 patients who were available for follow up 10 (76.9%) had subjective improvement in symptoms. Majority of patients showed regression on chest radiograph but one patient progressed to stage IV disease. Pulmonary function data of the patients who were followed up showed improvement but this was not significant statistically. Oral corticosteroids improved symptoms but changes in pulmonary function seemed to be independent of steroid therapy. Further study of a larger number of patients over a longer period would be necessary. PMID- 10534940 TI - Clinical profile of pulmonary nocardiosis. AB - Pulmonary nocardiosis mimics pulmonary tuberculosis in both clinical symptoms, being chronic in nature and radiological characteristics, and it is often wrongly treated with anti-tuberculosis drugs. The present study was undertaken to determine the prevalence of pulmonary nocardial infection in patients having chronic chest symptoms and to study their clinical response to specific chemotherapeutic agents. All the patients, who had a negative sputum for AFB on direct smear examination consecutively, were investigated for nocardiosis by examining the sputum with KOH preparation and modified Ziehl-Neelsen method. This was later confirmed by fungal culture of the sputum and inoculation on McClung's broth for paraffin baiting technique. Fibreoptic bronchoscopy was performed on all the suspected cases and the bronchial aspirate was examined similarly. The confirmed cases of nocardiosis were treated with cotrimaxazole and doxycycline for a total duration of six months. The prevalence of pulmonary nocardiosis in the present study was 1.9 percent. All the patients were immunocompetent. All the patients showed a good clinical response to chemotherapy at the end of six months of treatment. No relapse has been observed on follow up. PMID- 10534941 TI - Tracheal duplication as a cause of congenital stridor. AB - A six-week-old child with tracheal duplication presenting with congenital stridor is being reported. To the best of our knowledge, a tracheal web or tracheal duplication (as the present condition may be named) has not been described as a causative factor of infantile stridor. PMID- 10534942 TI - An unusual mediastinal mass due to metastasis. AB - During childhood the common anterior middle mediastinal masses are either lymph node enlargement or teratomas/dermoids. A case of ossific mediastinal metastasis and pleural metastases from osteosarcoma, presenting three years after the primary disease (late metastasis) is reported for its rarity. PMID- 10534943 TI - A case of motor neurone disease with sleep apnoea syndrome. AB - A case of a 63-year-old patient with motor neurone disease (amyotrophic lateral sclerosis) with central sleep apnoea syndrome is being reported. His sleep architecture was fragmented with a high apnea-hypopnea index of 65 per hour and maximum oxygen-desaturation of 78 percent. Total correction of sleep pattern with nasal non-invasive ventilation (BiPAP-ST) was demonstrated. PMID- 10534944 TI - Environmental pollution control authority of the National Capital Region. June 5, 1999, New Delhi. PMID- 10534945 TI - Characterization of pyrethroid resistance and susceptibility to coumaphos in Mexican Boophilus microplus (Acari: Ixodidae). AB - Two patterns of pyrethroid resistance were characterized from Boophilus microplus (Canestrini) collected in Mexico. One was characteristic of a kdr mutation and the other involved esterase and cytochrome P450 enzyme systems. Very high resistance to permethrin, cypermethrin, and flumethrin, not synergized by TPP and PBO and high resistance to DDT, characterized the kdr-like pattern found in the Corrales and San Felipe strains. Esterase and cytochrome P450-dependent resistance was found in the Coatzacoalcos strain. It was characterized by resistance to permethrin, cypermethrin, and flumethrin, synergized by TPP and PBO, but no resistance to DDT. The Coatzacoalcos strain also showed 3.6-fold resistance to the organophosphate coumaphos. This factor appeared to be independent of pyrethroid resistance. Pyrethroid resistance patterns found in Mexico were similar to those found earlier in Australia. The significance of pyrethroid and coumaphos resistance to the U.S. cattle fever tick quarantine is discussed. PMID- 10534946 TI - Sequence analysis of the knockdown resistance-homologous region of the para-type sodium channel gene from pyrethroid-resistant Boophilus microplus (Acari: Ixodidae). AB - Using reverse transcription polymerase chain reaction and degenerate oligonucleotide primers, a partial para-homologous sodium channel cDNA was obtained from the southern cattle tick, Boophilus microplus (Canestrini). The cDNA sequence encoded the region in which knockdown resistance (kdr)-type mutations have been identified in numerous insect species. Comparison of deduced amino acids from the cDNA sequence showed high similarity with sodium channels from other species, particularly in highly conserved repeat domains of the sodium channel. Analysis of the kdr-homologous region of the genomic DNA sequences from several susceptible and pyrethroid-resistant tick strains did not detect mutations. The result suggests novel mutations in the sodium channel gene or metabolic detoxification may be involved in the resistance to pyrethroids in this tick. PMID- 10534947 TI - Identification and purification of a 16-kDa allergen from Psoroptes ovis (Acarina: Psoroptidae). AB - Psoroptes ovis, (Hering), the sheep scab mite, is the causative agent of an allergic dermatitis of sheep and cattle. Recent studies of the host immune response to this ectoparasite have provided information that suggests control may be achieved by immune intervention. A significant effector in protection of the host from clinical lesions is host behavioral grooming. Host grooming is believed to be intensified by a pruritic immediate hypersensitivity response to mite allergens. Knowledge of potential P. ovis allergens is limited. This article reports on the identification and SDS-PAGE continuous elution purification of a 16-kDa polypeptide that elicits immediate type hypersensitivity in calves and has sequence homology with known group II mite allergens, Lep d 2 of Lepidoglyphus destructor (Schrank), and Der f II of Dermatophagoides farinae (Hughes). This P. ovis allergen appears to be a good vaccine candidate for further study and cloning. PMID- 10534949 TI - Use of orally administered chitin inhibitor (lufenuron) to control flea vectors of plague on ground squirrels in California. AB - The efficacy of orally administered lufenuron, a chitin inhibitor, to control fleas on California ground squirrels, Spermophilus beecheyi (Richardson), was evaluated during a 2-yr study in Santa Barbara County, CA. Results demonstrated that use of a host-targeted feed cube containing lufenuron was effective in significantly reducing the burden of Oropsylla montana (Baker) and Hoplopsyllus anomalus (Baker) fleas on ground squirrels. A flea index that indicated a mean number of fleas per squirrel of 10.0 decreased to 1.3 after 2 treatments in season 1, and to 0.7 and 0.2 after the 3rd and 4th treatments, respectively, in season 2. A cost comparison of this new method compared with a traditional reactive, emergency, insecticide-based plague control program demonstrated a cost reduction of approximately 90%. The results of this study indicated that a lufenuron feed cube was an effective, cost-saving, and proactive technique for controlling fleas on California ground squirrels, and thus reducing the risk of disease transmission in plague endemic regions. PMID- 10534948 TI - Association of Ixodes pacificus (Acari: ixodidae) with the spatial and temporal distribution of equine granulocytic ehrlichiosis in California. AB - This study was conducted to determine if the biology of certain ticks associated with horses regulates the spatial and temporal distribution of equine granulocytic ehrlichiosis (EGE) in California north of Monterey County. We compared the spatial and temporal distribution of EGE cases with the seasons of activity and life histories of ticks that infest horses. Spatially, cases collected from equine veterinarians clustered around each other in a manner different from the way in which control cities of practice were distributed, with foci limited to the Sierra Nevada and coastal foothills. Cases also clustered seasonally: most were diagnosed between November and April. The spatial and temporal pattern of EGE cases closely parallels the well-characterized life history and distribution of Ixodes pacificus Cooley & Kohls, but not other ticks commonly associated with horses. Building on previous studies, there is compelling evidence that this tick has the vectorial capacity to transmit Ehrlichia equi to horses. Based on the life history and distribution of I. pacificus in relation to EGE cases, we reason that this tick is the only biologically plausible vector of E. equi in California, and provide evidence for a tightly linked association between I. pacificus and the epidemiology of EGE. PMID- 10534950 TI - Spatio-temporal dynamics of resident and immigrating populations of Carcinops pumilio (Coleoptera: Histeridae) in high-rise poultry facilities. AB - The histerid beetle Carcinops pumilio (Erichson) is an important natural predator of the house fly, Musca domestica L., in accumulated poultry house manure. We examined the spatio-temporal dynamics of establishing adult C. pumilio in high rise poultry facilities using conventional and geostatistical approaches. The growth curves of resident and immigrating populations followed logistic and exponential equations, respectively, and their rates of establishment were statistically the same. Frequency distributions for both populations were strongly positively skewed, and approximately 53% of sampling intervals were significantly modeled by the negative binomial. Taylor's power law indicated both populations to be aggregated, and gave excellent least squares regression fits to both populations. Correlograms, a geostatistical tool, suggested little local spatial structure (e.g., 2nd order effects) for either population. The resident population was 'randomly' aggregated: beetles were clustered around randomly distributed aggregations of house fly immatures. The immigrating population exhibited significant spatial trends (e.g., 1st order effects) consistently seen at all sampling intervals. C. pumilio spatial structure was represented primarily by this spatial trend; thus, immigration of C. pumilio may have been either a singular event in time, or initiating at 1 or multiple times from a singular location. PMID- 10534951 TI - Ticks and antibodies to Borrelia burgdorferi from mammals at Cape Hatteras, NC and Assateague Island, MD and VA. AB - Results of a survey for ixodid ticks and/or serum antibodies to Borrelia burgdorferi from 14 species of small to large mammals from eastern coastal areas of the United States are presented. Most samples were obtained from July 1987 through June 1989 (excluding December-March) at 3 locales: Assateague Is. National Seashore, Worcester Co., MD., and Accomack Co., VA. (approximately 38 degrees 05' N 75 degrees 10' W), and Cape Hatteras National Seashore, Dare Co., NC (approximately 35 degrees 30' N 76 degrees 35' W). Hosts sampled included opossums (Didelphis virginiana), least shrews (Cryptotis parva), gray foxes (Urocyon cinereoargenteus), red foxes (Vulpes vulpes), raccoons (Procyon lotor), feral cats (Felis sylvestris), feral horses (Equus caballus), sika deer (Cervus nippon), rice rats (Oryzomys palustris), white-footed mice (Peromyscus leucopus), meadow voles (Microtus pennsylvanicus), house mice (Mus musculus), norway rats (Rattus norvegicus) and jumping mice (Zapus hudsonius). An indirect fluorescent antibody test was used for testing sera from opossums, raccoons, and feral cats; enzyme-linked immunosorbent assays were used for sera from foxes, horses, deer, and house and white-footed mice. Antibodies to B. burgdorferi were found in all species tested from each locale. Seasonal data reinforce the contention that P. leucopus is a suitable sentinel species for B. burgdorferi. Ticks on hosts included Ixodes scapularis Say, I. texanus Banks, Dermacentor variabilis (Say), D. albipictus (Packard), and Amblyomma americanum (L.). Males comprised approximately 0-22 and 60-81% of Ixodes sp. and Amblyomma-Dermacentor adults collected from hosts, respectively. All stages of A. americanum, adult D. variabilis, and larval I. scapularis were collected from vegetation. The highest seropositivity rate (67%) was recorded for 45 P. leucopus at Assateague during July, approximately 1 mo. after peak nymphal I. scapularis intensity. Borrelia burgdorferi was isolated from 6 nymphal and 12 female I. scapularis collected from P. leucopus and C. nippon, respectively, on Assateague. PMID- 10534952 TI - Population ecology of Phlebotomus argentipes (Diptera: Psychodidae) in West Bengal, India. AB - The population abundance of Phlebotomus argentipes Annandale & Brunetti was studied between January 1986 and December 1987 at 2 sites in West Bengal, India, in relation to 8 ecological parameters (air temperature, rainfall, windspeed, relative humidity, soil moisture, soil temperature, soil pH, and soil organic carbon). Sand flies were present throughout the year with minimum abundance in winter months and maximum during monsoon and postmonsoon months. Correlation analysis examined pairwise relationships among the 8 ecological parameters and P. argentipes abundance. Multiple linear regression of sand fly abundance on the 8 parameters showed that average soil temperature and soil moisture, both recorded 1 mo earlier, were associated positively with sand fly abundance. These findings have important implications for Indian kala-azar disease control and prevention. Effective vector management programs are needed most when weather conditions favor increased sand fly abundance, given that greater sand fly abundance increases the likelihood of host-vector contact and the transmission of Leishmania. PMID- 10534953 TI - Evidence for a new malaria vector species, species E, within the Anopheles culicifacies complex (Diptera: Culicidae). AB - Female Anopheles culicifacies Giles from Ramanathapuram district, Tamil Nadu state, India, were examined for oocysts and sporozoites and their larval progeny for mitotic karyotype. Collections were made from Mandapam and Uchipuli on the mainland, and Thangachimadam and Pamban on Rameshwaram Island. Of the 451 An. culicifacies females that were collected and dissected, 24 were found positive for Plasmodia (21 for sporozoites and 3 for oocysts). Both acrocentric and submetacentric Y-chromosome karyotypes were observed among the progeny of females from all villages. All 11 iso-female lines whose parental females were positive for sporozoites or oocysts had progeny with submetacentric Y-chromosomes. Total absence of sporozoite-positives among mothers of acrocentric males was evidence of assortative mating between these 2 sympatric populations (i.e., 2 species). We propose that the nonvector population with acrocentric Y-chromosome sons retain the original designation of species B and that the vector population with the submetacentric Y-chromosome sons be designated as species E, a new species. PMID- 10534954 TI - Mark-release-recapture experiment with adult Anopheles minimus (Diptera: Culicidae) on Ishigaki Island, Ryukyu Archipelago, Japan. AB - Movement of adult An. minimus Theobald between larval habitats and feeding sites was investigated by mark-release-recapture in March 1993 on Ishigaki Island, Ryukyu Archipelago, Japan. Unfed (998 females) and fed (1,485 females) cohorts from a laboratory colony were marked with different colors of fluorescent dye, and released near Nishihama stream where larvae were abundant. Overall, 42 fed (2.82%) and 13 unfed (1.30%) females were recaptured. Among these, 12 females were recaptured at a cowshed, the nearest blood meal source. Most females in the fed cohort recaptured at the release point had developing or matured ovaries, whereas 55% at the cowshed were parous. The movement of released females from larval habitat to the cowshed for blood feeding indicated a flight distance > 1 km. PMID- 10534955 TI - Evaluation of various substances to increase adult Stomoxys calcitrans (Diptera: Muscidae) collections on alsynite cylinder traps in north Florida. AB - During 1993-1995, field studies evaluated various volatile substances to increase the catch of adult stable flies, Stomoxys calcitrans L., on adhesive-coated translucent fiberglass (Alsynite) cylinder traps. Dry ice, 1-octen-3-ol (referred to as octenol), acetone, 4:1:8 mixture of 1 octen-3-ol: 3-n-propylphenol: 4 methylphenol, and an eye gnat (Hippelates) attractant were tested. Using dry ice as a baseline, the latter 4 treatments also were considered as possible alternatives to carbon dioxide. Dry ice significantly increased fly collections on cylinders as much as 25-fold compared with cylinders with no odor. Although trap collections increased by approximately 4% with addition of octenol (release rate approximately 18.0 mg/h), it was not significantly different when compared with dry ice alone. Fly collections on cylinders baited with octenol only were significantly lower than dry ice and not significantly different from cylinders with no odor. Collections from Alsynite cylinders baited with either acetone released at approximately 62.0 mg/h or eye gnat bait plus sand caught significantly more stable flies than no odor. However, neither substance increased fly collections as much as dry ice. The 4:1:8 phenolic mixture (released at either 0.7 mg/h or 20.0 mg/h) significantly increased fly collection on cylinders nearly 6-fold compared with no odor and warrants further investigation as an alternative to carbon dioxide for sampling stable flies. PMID- 10534956 TI - Detection of cryptic species in the Tabanus nigrovittatus (Diptera: Tabanidae) complex in Massachusetts by morphometric and cuticular hydrocarbon analysis. AB - Greenhead flies of the Tabanus nigrovittatus complex from Massachusetts salt marshes were identified as T. nigrovittatus Macquart and T. conterminus Walker using the morphometric model developed by Sofield et al. Four body measurements from a total of 5,983 female flies collected over 2 consecutive years yielded canonical scores producing a unimodal rather than the expected bimodal distribution. The lack of bimodality indicated that both species were not present at the study site. This was substantiated by cuticular hydrocarbon (CHC) analysis of a subsample of these specimens. Fifteen, female flies of the Tabanus nigrovittatus complex from the same site were identified to species using the Sofield et al. morphometric model and validated using CHC analysis. Two individuals of the T. nigrovittatus complex were identified incorrectly as T. conterminus by the morphometric model. The tendency of this model to incorrectly classify some individuals of T. nigrovittatus as T. conterminus brings into question the identity of the Walker syntypes of T. conterminus. Based on CHC analysis, our study shows that both species coexist within our study area. PMID- 10534957 TI - Avian serology in a St. Louis encephalitis epicenter before, during, and after a widespread epidemic in south Florida, USA. AB - Blood and serum from 3,915 wild and domestic birds (2,590 resident, 139 migrant, and 1,186 captive), representing 56 species collected in central Florida from 1989 through 1997, were analyzed for evidence of St. Louis encephalitis (SLE) virus transmission. All sera were tested for SLE hemagglutination inhibition (HI) antibody. Selected sera and bloods were tested for SLE neutralizing (NT) antibody and virus. The reproductive success of resident birds was highest from 1990-1992 and lowest from 1994-1997. Transmission of SLE to resident birds, especially mourning doves (Zenaida macroura), peaked during the summer of 1990, a year during which a widespread SLE epidemic was recorded in central Florida. The SLE antibody-positive resident birds 1st appeared during September of the epidemic year. Some SLE, HI antibody-positive resident birds were captured throughout 1991, but only 5% were yearlings, compared with 36% in 1990. By 1993, wild resident birds expressing HI and NT antibodies to SLE had nearly disappeared. None of the migrant birds tested were SLE-positive. Sentinel chickens maintained in Indian River County during the epidemic year seroconverted to SLE starting in early July with peak seroconversion rates in August, September, and October. High (> or = 50%) SLE seroconversion rates in sentinel chickens preceded those in wild birds by 10 wk and preceded peak human SLE transmission by at least 8 wk. Major SLE epidemics in south Florida depend on abundant wild bird populations, especially during the amplification phase of the transmission cycle. We propose that hard winter freezes along the temperature-subtropical climatic zone interface in central Florida, at approximately 27 degrees 30' North Latitude, opens foraging and nesting habitats for ground-feeding birds, resulting in high reproductive success and an abundance of seronegative individuals that rapidly amplify the SLE later in the year. PMID- 10534958 TI - Repellency of essential oils to mosquitoes (Diptera: Culicidae). AB - The repellency of different concentrations (5, 10, 25, 50, 75, and 100%) and combinations of 5 essential oils (Bourbon geranium, cedarwood, clove, peppermint, and thyme) to Aedes aegypti (L.) and Anopheles albimanus Wiedemann when applied to human skin was determined in laboratory tests. Cedarwood oil failed to repel mosquitoes and only high concentrations of peppermint oil repelled Ae. aegypti. None of the oils tested prevented mosquito bite when used at the 5 or 10% concentration. Thyme and clove oils were the most effective mosquito repellents and provided 1 1/2 to 3 1/2 h of protection, depending on oil concentration. Clove oil (50%) combined with geranium oil (50%) or with thyme oil (50%) prevented biting by An. albimanus for 1 1/4 to 2 1/2 h. The potential for using essential oils as topical mosquito repellents may be limited by user acceptability; clove, thyme, and peppermint oils can be irritating to the skin, whereas both human subjects in this study judged the odor of clove and thyme oils unacceptable at concentrations > or = 25%. PMID- 10534959 TI - Megaselia scalaris (Diptera: Phoridae) causing myiasis in Crotalus durissus terrificus (Serpentes: Viperidae) in Brazil. AB - We describe a case of myiasis in Crotalus durissus terrificus (Laurenti) caused by Megaselia scalaris (Loew). The snake was found in Anhembi, Sao Paulo, Brazil, with a lesion measuring 25 mm in diameter where the larvae of M. scalaris had penetrated the ribs. The opportunistic behavior of the larvae of M. scalaris is discussed. PMID- 10534960 TI - Analysis of ITS2 DNA sequences from Brazilian Anopheles darlingi (Diptera: Culicidae). AB - Specimens of Anopheles darlingi Root, the major vector of malaria in Brazil, were collected from several states in Brazil: Sao Paulo (Dourado), Bahia (Itabela), Rondonia (Porto Velho), Roraima (Boa Vista), and Acre (Placido de Castro). Sequence divergence in the 2nd internal transcribed spacer (ITS2) was examined. The ITS2 sequences of mosquitoes captured in the Amazon region (Porto Velho, Boa Vista and Placido de Castro) and in the northeast of Brazil (Itabela) were almost identical; however, a 4-5% sequence divergence was observed in the ITS2 of mosquitoes captured in the southeast (Dourado). Further analysis is needed to determine if these differences indicate that Dourado population may be a separate species. PMID- 10534961 TI - Preliminary survey for entomopathogenic fungi associated with Ixodes scapularis (Acari: Ixodidae) in southern New York and New England, USA. AB - Free-living larval, nymphal, and adult Ixodes scapularis Say were collected from scattered locales in southern New England and New York to determine infection rates with entomopathogenic fungi. Infection rates of larvae, nymphs, males, and females were 0% (571), 0% (272), 0% (57), and 4.3% (47), respectively. Two entomopathogenic fungi were isolated from field-collected I. scapularis females from Fire Island, NY. Isolates were identified as Verticillium lecanii (Zimmermann) Viegas and Verticillium sp. (a member of the Verticillium lecanii species complex). PMID- 10534962 TI - Key to third-instar chrysomyinae (Diptera: Calliphoridae) from carrion in the continental United States. AB - The introduction of 4 Chrysomya Robineau-Desvoidy spp. to the Americas has made obsolete previously published keys to Nearctic calliphorid larvae, particularly those covering the subfamily Chrysomyinae. To assist forensic entomologists, ecologists, and public health workers, we provide a key to 3rd instars of 8 chrysomyine species reported from or likely to occur in carrion within the continental United States. The rare (in the United States) species Cochliomyia aldrichi Del Ponte, C. minima Shannon, and Chloroprocta idioidea (Robineau Desvoidy) are not included because specimens and suitable descriptions were unavailable. PMID- 10534963 TI - Fatigue in women receiving adjuvant chemotherapy for breast cancer: characteristics, course, and correlates. AB - This study investigated the characteristics, course, and correlates of fatigue in women receiving adjuvant chemotherapy for breast cancer. Fifty-four patients were assessed before the start of chemotherapy and during the first three treatment cycles. An age-matched sample of women with no cancer history was assessed at similar time intervals for comparison purposes. Results indicated that breast cancer patients experienced worse fatigue than women with no cancer history. These differences were evident before and after patients started chemotherapy. In addition, fatigue worsened among patients after treatment started. More severe fatigue before treatment was associated with poorer performance status and the presence of fatigue-related symptoms (e.g., sleep problems and muscle weakness). Increases in fatigue after chemotherapy started were associated with continued fatigue-related symptoms and the development of chemotherapy side effects (e.g., nausea and mouth sores). These findings demonstrate the clinical significance of fatigue in breast cancer patients before and during adjuvant chemotherapy treatment. Results also suggest that aggressive management of common side effects, such as nausea and pain, may be useful in relieving chemotherapy-related fatigue. PMID- 10534964 TI - Terminal cancer patients and timing of referral to palliative care: a multicenter prospective cohort study. Italian Cooperative Research Group on Palliative Medicine. AB - This study describes the characteristics of a representative sample of terminally ill cancer patients at admission to Italian palliative care programs, the rate and reasons for discontinuation of care, and survival after enrollment. All Italian palliative care units (PCUs) specifically committed to palliative care were asked to consecutively register all new patients (n = 3901) between January and June, 1995. Fifty-eight of the 62 PCUs contacted by the Steering Committee completed the study. A random sample of 589 evaluable patients was prospectively selected from the 2667 eligible patients. Patients were mostly referred by a general practitioner (31.2%) or a specialist (42.1%). Most patients (84.7%) were followed until death. Seventy-seven discontinued care because of hospital admission (6.6%), change of residence (3.9%), refusal (1.7%), or improvement (0.8%). Median survival was 37.9 days; 14.3% of the patients died within 7 days, and 15.3% lived longer than 180 days. A statistically significant association between survival and gender, cancer type, setting of the first visit, and type of unit was observed. In Italy, as in other countries with different health systems, referral of cancer patients to palliative care tends to occur late in the course of the disease. This study suggests that the process of enrollment and the duration of patients' survival in palliative care, when studied in large unselected populations, can provide important information relevant to the care of terminally ill patients. PMID- 10534965 TI - Symptom prevalence, characteristics, and distress in AIDS outpatients. AB - Symptom distress is an important but poorly characterized aspect of quality of life in AIDS patients. To assess and characterize the symptoms and symptom distress associated with AIDS, 504 ambulatory patients with AIDS were evaluated between December, 1992 and December, 1995. The assessment included measures of symptom distress, physical and psychosocial functioning, and demographic and disease-related factors. Patients described symptoms during the previous week using the Memorial Symptom Assessment Scale Short Form (MSAS-SF), a validated measure of physical and psychological symptom distress. The mean age was 38.6 years (range 18-69); 56% were male. African-Americans comprised 40% of the sample, Caucasians 35%, and Hispanics 23%. Ninety-three percent had CD4+ T-cell counts below 500, and 66% had counts below 200; 69% were classified in CDC category C (history of AIDS-defining conditions). Fifty-two percent reported intravenous drug use. Karnofsky performance status was > or = 70 in 80% of the patients. No patients were taking protease inhibitors. The mean (+/- SD) number of symptoms was 16.7 +/- 7.3. The most prevalent symptoms were worrying (86%), fatigue (85%), sadness (82%), and pain (76%). Patients with Karnofsky performance scores < 70 had more symptoms and higher symptom distress scores than patients with scores > or = 70 (21.2 +/- 6.5 vs. 15.6 +/- 7.1 symptoms/patient; 2.3 +/- 0.8 vs. 1.6 +/- 0.8 on the Global Distress Index [GDI] of the MSAS-SF; P < 0.0001 for both). Patients who reported intravenous drug use as an HIV transmission factor reported more symptoms and higher overall and physical symptom distress than those who reported homosexual or heterosexual contact as their transmission factor (17.8 +/- 7.5 vs. 15.4 +/- 6.9 symptoms/patient, P = 0.0002; 1.9 +/- 0.9 vs. 1.6 +/- 0.8 on the MSAS-GDI, P = 0.002). Both the number of symptoms and symptom distress were highly associated with psychological distress and poorer quality of life; for example, r = -0.69 (P < 0.0001) between GDI scores and scores on a validated measure of quality of life. Neither gender nor CD4+ T-cell count was associated with symptom number or distress. Responses from this self referred sample of AIDS outpatients indicate that AIDS patients experience many distressing physical and psychological symptoms and a high level of distress. Both the number of symptoms and the distress associated with them are associated with a variety of disease-related factors and disturbances in other aspects of quality of life. Symptom assessment provides information that may be valuable in evaluating AIDS treatment regimens and defining strategies to improve quality of life. PMID- 10534966 TI - Physicians' recognition of the symptoms experienced by HIV patients: how reliable? AB - To assess how well physicians recognize common symptoms in HIV patients and identify factors associated with symptom recognition, a multicenter cross sectional survey was performed in a random sample of 118 hospitalized and 172 ambulatory HIV patients, and their attending physicians. Patients' reports of 16 different symptoms were compared to physicians' reports of whether each symptom was present and/or specific treatments prescribed. Overall, fatigue, anxiety, skin problems, fever, and weight loss were more often recognized by physicians than other symptoms. Agreement between patients and physicians was poor to moderate, with Kappa statistics ranging from 0.17 (dry mouth) to 0.58 (fever). Recognition was independently more likely for ambulatory patients (adjusted odds ratio 1.69, P < 0.001) and for patients seen as sicker (adjusted odds ratio 1.88, P < 0.001). Appropriate symptom management requires improved symptom recognition. More systematic clinical examinations, including attentive patient interview, are needed. PMID- 10534967 TI - Can a controlled-release oral dose form of oxycodone be used as readily as an immediate-release form for the purpose of titrating to stable pain control? AB - Two separate trials compared controlled-release (CR) oral oxycodone (administered every 12 hours) with immediate-release (IR) oxycodone (4 times a day) to determine whether patients with chronic pain could be titrated to stable pain control as readily with the CR as with the IR formulation. In one study, 48 patients with cancer pain were randomized to open-label titration with either CR or IR oxycodone (maximum dose, 400 mg/day) for a period of up to 21 days. In a study of similar design, 57 patients with low back pain were titrated with either CR or IR oxycodone (maximum dose, 80 mg/day) for a period of up to 10 days. The majority of patients in both studies were converted to oxycodone from other opioid analgesics. Results of both studies showed no difference between CR and IR oxycodone with respect to both the percentage of patients achieving stable pain control, the time to achieve stable pain control, and the degree of pain control achieved. Among cancer patients, 85% achieved stable analgesia, 92% with the CR formulation and 79% with the IR formulation. Among noncancer patients, 91% achieved stable pain control, 87% with the CR formulation and 96% with the IR formulation. The most commonly reported adverse effects in both studies were similar for the two formulations and were those anticipated with opioids: nausea, vomiting, constipation, somnolence, dizziness, and pruritus. Nausea and vomiting were the most frequently cited reasons for treatment discontinuations. These studies suggest that dose titration can be accomplished as readily with oral CR oxycodone as with IR oxycodone in patients with chronic, moderate to severe pain. PMID- 10534969 TI - Nurses' knowledge about pharmacological and nonpharmacological pain management in children. AB - The purpose of this study was to investigate the knowledge base and practices of Finnish nurses in the area of children in pain. The convenience sample consisted of 265 nurses working on children's wards in university hospitals. Data were collected using an instrument designed for the study. The results showed that there remain gaps in the knowledge base of nurses with regard to both pharmacological and nonpharmacological pain management in children. The education and the area of expertise were significant influences on knowledge scores. Nurses used a fairly wide range of nonpharmacological pain alleviation methods but most of these were such that the nurse was in an active role and the child was passive. There is a clear need for further education. Nurses should take a more active role in seeking new information and also should be encouraged to use nonpharmacological methods that let the children be active participants in their own care. PMID- 10534968 TI - Opioid-induced emesis among hospitalized nonsurgical patients: effect on pain and quality of life. AB - There is very little information in the medical literature regarding opioid induced emesis and its relationship to patient outcomes. Two-hundred and six nonsurgical patients in a 400-bed teaching hospital with minimal known risks of disease-associated emesis were interviewed to examine emesis and associated outcomes following the administration of opioids for acute pain management. The mean age, weight, and height of the study group were 54.4 (+/- 19.6) years, 175.8 (+/- 45.7) pounds, and 67.1 (+/- 4.4) inches, respectively. Seventy-three (35.4%) patients experienced nausea; 28 (13.6%) patients vomited; and 15 (7.3%) patients retched following the opioid therapy. These symptoms were mild and discomforting for relatively short periods of time. The patients' ability to concentrate and eat was affected by the incidence of nausea/vomiting. The intensity, duration, and severity of nausea were positively associated with the magnitude of the functional limitations. The symptoms also influenced patients' ratings of various hospital satisfaction measures. In conclusion, emesis due to opioids represents a notable burden on nonsurgical patients. Successful therapies that prevent opioid induced emesis are likely to positively influence patient outcomes by reducing adverse effects, improving functional outcomes, and enhancing quality of life. PMID- 10534970 TI - Oral glycopyrrolate alleviates drooling in a patient with tongue cancer. AB - Although sialorrhea and drooling are uncommon symptoms in cancer patients, they can cause considerable discomfort, inconvenience and social embarrassment. In this article we describe a patient with tongue cancer who was successfully treated with oral glycopyrrolate 0.4 mg 3 times daily. Glycopyrrolate is a quaternary ammonium compound. In contrast to the recommended treatment with scopolamine, glycopyrrolate is virtually without side effects to the central nervous system because it penetrates the blood-brain barrier poorly. Glycopyrrolate has a slow and erratic absorption from the gastrointestinal system, but even low plasma levels are associated with a distinct and long lasting antisialogic effect. PMID- 10534971 TI - Reduction in constipation and laxative requirements following opioid rotation to methadone: a report of four cases. AB - Constipation is a common symptom in cancer patients, especially in those who are receiving opioid analgesics for pain. Although several articles have recently examined constipation with respect to causation and treatment, little research has been done to elucidate the effects of different opioids on the bowel. Recent research has found that laxative doses may be lower in patients using methadone as an analgesic, but changes in constipation were not measured. We report here on four cases in which patients had improvement in constipation and decreased laxative requirements following opioid rotation to methadone. PMID- 10534972 TI - The highly integrated dialogue between neurons and astrocytes in brain function. AB - For decades neurons have been regarded as the only cells involved in the generation and control of brain signalling, while the surrounding glia was supposed to provide structural and metabolic support to neuronal function. However, based on a number of recent findings, a new view is emerging: astrocytes, the glial cells ensheathing synaptic specializations, are active and integrated participants of neurotransmission. Not only do astrocytes take up and remove synaptically released glutamate (the major excitatory neurotransmitter), thus ending its stimulatory action and preventing neuronal damage, but also and outstandingly, they are able to undergo rapid bidirectional communication with neurons, based on reciprocal glutamatergic signalling. Thus, release of glutamate from synaptic terminals, in addition to postsynaptic neurons, turns on the astrocytes nearby which respond by liberating the same neurotransmitter via a novel Ca(2+)-dependent mechanism and thereby signal back to neurons. The present review discusses the above findings and their important implications as well as additional evidence supporting the new concept of an integrated neuron-astrocyte communication in brain function. PMID- 10534973 TI - Force systems developed by six different cantilever configurations. AB - OBJECTIVES: To describe the force systems developed by cantilevers with different configurations during incisor intrusion. DESIGN: A two-dimensional Finite Element model. SETTING AND SAMPLE STUDIED: Laboratories of the Department of Orthodontics, Royal Dental College, Aarhus University, Denmark. The materials studied comprised cantilevers made of 0.017" x 0.025" TMA wire. EXPERIMENTAL VARIABLE: Vertical activation of the six different modelled cantilever configurations. OUTCOME MEASURE: Force level and direction during activation and deactivation analysed at the point of force application. RESULTS: The force direction was dependent on the configuration of the cantilever. Only the cantilevers with a curvature produced combined retraction and intrusion forces. All other configurations resulted in combined protrusion and intrusion, which reversed into retraction and intrusion, according to the variations in deactivation. CONCLUSION: The choice of correct cantilever configuration is important when selecting the mechanics of intrusion for a particular patient. PMID- 10534974 TI - Shape of the face and tongue in obstructive sleep apnea patients--statistical analysis of coordinate data. AB - OBJECTIVES: To determine the shape difference of the face and tongue of obstructive sleep apnea (OSA) patients, in comparison to those of non-apneic patients. DESIGN: Retrospective analysis of observational data on a cohort of patients. SETTING: A university teaching hospital and sleep referral center. SAMPLE POPULATION AND METHOD: Eighty patients referred for overnight polysomnography and lateral cephalometry and who met the selection criteria were included. Upright and supine cephalograms were obtained and subgrouped based on the severity of clinical symptoms. Shape differences between the groups were assessed by a multiple analysis of variance and a Hotelling's T2. MEASUREMENTS AND RESULTS: A set of anatomical landmarks were selected for outlines of the face and the tongue on cephalograms. X and Y coordinates of each landmark were utilized as variables. As symptoms become severe, the hyoid bone and the submental area positioned inferiorly and the fourth vertebra relocated posteriorly with respect to the lower mandibular border. When subjects changed their body position from the upright to the supine, the posterior part of the tongue appeared to sink down. The hyoid bone position to epiglottis-retrognathion line in the supine position distinguishes OSA patients from non-apneic subjects. CONCLUSION: Despite many limitations, we demonstrate that the supine cepahlometrics during wakefulness can be a useful adjunctive diagnostic tool for OSA, when cephalograms are analyzed in a coordinate data form. PMID- 10534975 TI - Dentofacial morphology and upper respiratory function in 8-10-year-old children. AB - OBJECTIVES: The aim of this preliminary study was to assess the nature of associations between selected dentofacial morphological variables and respiratory mode as measured by percent nasality (%N) as part of an ongoing longitudinal study. DESIGN: Cross-sectional cohort study. SETTING AND SAMPLE POPULATION: The Pediatric Clinical Study Center, Children's Hospital, Columbus, OH. Ninety-eight normal children were tested. EXPERIMENTAL VARIABLE: Normal variation in %N. OUTCOME MEASURE: Selected dentofacial morphological variables including total and lower anterior face heights, face width, and palatal arch width and %N were estimated. RESULTS: Small associations between morphologic features and respiratory mode were found, but none were statistically significant. CONCLUSION: No evidence exists for the classic association between 'mouth breathing' and the stereotype of the 'adenoid facies'. PMID- 10534976 TI - Stability after sagittal split ramus osteotomy without post-operative maxillomandibular fixation in the treatment of prognathic patients with asymmetric mandibles. AB - OBJECTIVE: To examine post-operative stability in prognathic patients with mandibular asymmetry who underwent sagittal split ramus osteotomy (SSRO) of the mandible and titanium screw fixation without post-operative maxillomandibular fixation (MMF). DESIGN: Skeletal and dental changes after surgery with and without post-operative MMF were compared between prognathic patients with symmetric and asymmetric mandibles. SETTING AND SAMPLE POPULATION: Second Department of Oral and Maxillofacial Surgery at Tokyo Medical and Dental University. Twenty prognathic patients were examined. EXPERIMENTAL VARIABLE: An appliance for repositioning the proximal segment was applied in all 20 patients. Ten patients with symmetric mandibles received post-operative MMF with stainless steel wires and intermaxillary rubber traction after the removal of MMF (Group I), while the other 10 patients with asymmetric mandibles underwent post operative intermaxillary rubber traction only (Group II). OUTCOME MEASURE: Posteroanterior cephalograms were obtained pre-operatively; 2-3 days post operatively; and 3, 6, and 12 months after surgery. Changes in the positions of the gonion points (Go) and upper and lower incisors (U-1 and L-1) were examined. RESULTS: In both groups, the Go tended to shift laterally as a consequence of the operation. Although the tendencies of the post-operative changes in the Go points of Groups II and I were different, statistical analyses revealed no significant differences between the two groups. At the last stage of the follow-up period, the absolute value of the change in L-1 tended to be larger in Group II than in Group I, but without any statistical significance. CONCLUSION: This study suggests that post-operative change in prognathic patients with asymmetric mandibles treated without post-operative MMF is comparable to that in patients with symmetric mandibles treated with post-operative MMF. Accordingly, post operative MMF may be avoided, even in prognathic patients with asymmetric mandibles. PMID- 10534977 TI - The use of linear and angular measurements of maxillo-mandibular anteroposterior discrepancies. AB - OBJECTIVES: To extend the assessment of the clinical significance of the measurement of the anteroposterior discrepancy between maxilla and mandible on the bisector of the palatal plane to mandibular plane angle to a large group of randomly selected patients of both sexes, and to verify the correlation of this measurement to well established angular and linear assessments of anteroposterior discrepancy. DESIGN: Retrospective analysis of pre-treatment lateral cephalometric radiographs. SETTING AND SAMPLE POPULATION: The Laboratory of Functional Anatomy of the Stomatognathic Apparatus at Milan University, Italy. Three hundred and six orthodontic patients (165 males, 141 females) aged between 6 and 50 years. EXPERIMENTAL VARIABLE: ANB angle; corrected ANB* angle which compensates for the position of the maxilla and rotation of the mandible relative to the cranial base; Wits appraisal; MM-Wits: linear distance between the projections of A and B points on the bisector of the palatal plane to mandibular plane angle. RESULTS: The MM-Wits distance was significantly correlated to two angles (ANB and ANB*), as well as to the Wits appraisal. All the correlations performed did not show sex- or age-characteristic patterns. The correlation to the corrected ANB* was the best among the three, with a correlation coefficient of 0.915, MM-Wits (mm) = 1.497 x ANB* (degrees) -6.784. From the correlation, reference values for the new measurement have also been estimated, and found to be between -0.65 and -6.85 mm for skeletal Class I individuals. CONCLUSION: It is recommended that the diagnosis of orthodontic anomalies should be performed taking into consideration more than a single anteroposterior appraisal. PMID- 10534978 TI - Discovery: Osf2/Cbfa1, a master gene of bone formation. AB - This report reviews the current research that has impacted on our understanding of osteogenesis. Recent studies indicate that the transcription factor Osf2 (osteoblast specific transcription factor 2)/Cbfa1 (core binding factor activity 1) serves as a Master Gene regulating osteoblast-specific gene expression. The gene is expressed in cells of the osteoblast lineage only, and this expression is regulated by calciotropic agents. Moreover, when expressed in non-skeletal cells, the cells assume many of the characteristics of an osteoblast. In knockout experiments designed to assess the importance of the gene in osteogenesis, no evidence of bone formation could be observed in animals that are homozygous for the deletion. Studies of the heterozygote indicate that osteoblast function is compromised: there is a severe reduction in the number of bone cells, the tissue is deficient in bone proteins, and the activity of the enzyme alkaline phosphatase is low. It was noted that the heterozygote displays abnormalities that are remarkably similar to those exhibited by cleidocranial dysplastics. Indeed, Osf2 mapped close to a chromosomal locus on chromosome 6p21, long suspected of being involved with the disease. A search conducted for Osf2 mutations in kindreds with cleidocranial dysplasia revealed deletions, insertions, and missense mutations; these mutations are found to segregate with patients who are defined clinically as cleidocranial dysplastic. Aside from providing a new insight into a disease state that has so far avoided molecular analysis, results of the studies emphasize that the loss of a Master Gene drastically alters the development and maintenance of the appendicular skeleton and the craniofacial complex. PMID- 10534979 TI - The emerging soft tissue paradigm in orthodontic diagnosis and treatment planning. AB - Until now, orthodontic diagnosis and treatment planning has been based on hard tissue relationships and on the Angle paradigm that considers ideal dental occlusion 'nature's intended ideal form'. In this view, the clinician and nature are partners in seeking the ideal. In the modern biological model, variation is accepted as the natural form; ideal occlusion is the exception rather than the rule, and the orthodontist and nature are often adversaries. The orthodontist's task is to achieve the occlusal and facial outcomes that would most benefit that individual patient, whose esthetic concerns are often paramount. Because the soft tissues largely determine the limitations of orthodontic treatment, from the perspectives of function and stability, as well as esthetics, the orthodontist must plan treatment within the patient's limits of soft tissue adaptation and soft tissue contours. This emerging soft tissue paradigm in diagnosis and treatment planning places greater emphasis on clinical examination of soft tissue function and esthetics than has previously been the case, and new information in these areas is required. PMID- 10534980 TI - Understanding orthodontic treatment satisfaction from the patients' perspective: a qualitative approach. AB - OBJECTIVES: To apply qualitative research methods to develop hypotheses about orthodontic treatment process and outcome, as well as treatment satisfaction. DESIGN: Four focused interviews with 4-8 adolescents in retention. SETTING AND SAMPLE POPULATION: UNC orthodontic clinic; 22 patients in retention. RESULTS: Patients expressed dissatisfaction with some aspects of the treatment process, but were generally satisfied with the treatment outcome. Patients were aware of differences between clinic versus private treatment. From these discussions, we infer that there may be important differences between patients' and orthodontists' perceptions of the treatment process. CONCLUSIONS: Qualitative methods are useful tools for exploring orthodontic treatment from the patients' perspective and can be used to suggest important future areas of research. PMID- 10534981 TI - The etiology of palatal displacement of maxillary canines. AB - OBJECTIVES: To test the hypothesis that palatal displacement of the maxillary canine is completely under genetic influence. DESIGN: A randomized controlled design studied cases affected by a severe expression of lateral incisor anomaly on one side and by milder expression of the same anomaly on the other. Comparison of frequency of occurrence of unilateral palatally displaced canine measured in each. Each side acted as control for the other within the same individual. SETTING AND SAMPLE POPULATION: The Departments of Orthodontics of the Universities of Jerusalem and Tel Aviv and in private practice. From approximately 12,000 consecutively treated patients, all those exhibiting an anterior maxilla with a missing lateral incisor on one side, a peg-shaped or reduced lateral incisor on the other, and a palatally displaced canine (n = 19). OUTCOME MEASURE: Missing lateral incisors, peg-shaped, and reduced lateral incisors (all genetically determined characters) have been shown to be associated with palatal displacement of the canine. The canine displacement is presumed by some authorities to be similarly genetically determined. If this is so, then the impacted canine should occur with equal frequency on either side in the patient with a missing lateral incisor on one side and a peg-shaped or reduced lateral incisor on the other. RESULTS: The canine aberration occurred far more frequently on the side of the diminutive lateral incisor. CONCLUSION: There is an environmental factor involved in the palatal displacement of maxillary canines. PMID- 10534982 TI - A histomorphometric and scanning electron microscopy study of human condylar cartilage and bone tissue changes in relation to age. AB - OBJECTIVES: To determine the possibility for adaptive growth in human condyles, quantifying the thickness of fibrocartilage and the constitution of cells with potential activity, the trabecular bone volume, and the structural parameter: marrow space star volume in a larger sample of human autopsy condyles. EXPERIMENTAL SETTING AND DESIGN: A histomorphometric and scanning electron microscopic analysis of cartilage characteristics and bone remodelling activity. The Departments of Orthodontics and Cell Biology at Aarhus University, Denmark. An autopsy sample of condyles from 20 individuals, 18-31 years of age. Correlation analyses between ages and all parameters were conducted. RESULTS: There was a statistically significant correlation between age and the fibrocartilage thickness (r = -0.48, p < 0.05) and age and the trabecular bone volume (r = -0.52, p < 0.05). There was a statistically significant correlation between age and the hypertrophic chondrocytes (r = -0.50, p < 0.05) and age and the hypertrophic chondrocytes in bone (r = -0.48, p < 0.05). There was no significant correlation between age and the two other zones in the fibrocartilage. No statistical analyses were carried out concerning gender differences, as the sample consisted of very few females. Individual morphological description was performed, and is described in the article. Only slight age-related differences were found concerning the quantitative measurements for the fibrocartilage and the underlying bone tissue. CONCLUSIONS: Quantitative and qualitative investigations of the turnover activity in the fibrocartilage and the bone tissue, describing activity of hypertrophic chondrocytes and the trabecular bone tissue, indicated condylar growth potential in the age group until 30 years of age. The growth activity seemed to decline with age. PMID- 10534983 TI - Consistency of orthodontic treatment planning decisions. AB - OBJECTIVE: (i) To analyse the consistency of a group of orthodontists' treatment decisions and (ii) to identify factors that may influence this consistency. DESIGN: Cross-sectional observational study. SUBJECTS: Ten orthodontists. METHODS: The orthodontists examined 60 case vignettes and recorded their treatment decisions on two separate occasions. MAIN MEASURE: Inter- and intra examiner agreement was evaluated with the Kappa statistic. RESULTS: The consistency within each orthodontist was moderate. However, between-orthodontist consistency was poor. CONCLUSION: It appears that there is marked variation between orthodontists in treatment planning decisions and this may reflect the lack of evidence for the effectiveness of competing treatments. PMID- 10534984 TI - Mandibular setback osteotomy: facial soft tissue behavior and possibility to improve the accuracy of the soft tissue profile prediction with the use of a computerized cephalometric program: Quick Ceph Image Pro: v. 2.5. AB - OBJECTIVES: In part I, to derive ratios of soft to hard tissue profile changes after mandibular setback surgery and to report postoperative changes in soft tissue thickness at the lip and chin areas and, furthermore, to test the hypothesis that soft tissue thickness can act as one of the predictors of soft tissue response after surgery. In part II, to compare the predicted profile lines, using either the customized or the pre-programmed ratios, with the actual postsurgical outcomes. DESIGN: A retrospective study with the sample divided into two groups for different purposes. SETTING AND SAMPLE POPULATION: Department of Orthodontics and Department of Oral and Maxillofacial Surgery at Goteborg University, Sweden. Forty-one Caucasian subjects in need of mandibular setback surgery only. EXPERIMENTAL VARIABLE: Hard and soft tissue movements and changes in soft tissue thickness after surgery were calculated using a customized analysis. Comparisons of the predicted profile outcomes with the actual postsurgical outcomes were carried out with another customized analysis. OUTCOME MEASURE: Distance measurements of certain landmarks in relation to constructed reference lines, both in horizontal and vertical planes, were calculated. RESULTS: The upper lip thickness decreased and the lower lip thickness increased after surgery. The hypothesis that the soft tissue thickness at the lip and chin areas could act as predictors of the ratios of soft to hard tissue changes after surgery, was not supported. Ratios for the lower lip of about 83% of the horizontal and 14% of the vertical movement of the PM point on the bony chin were found in Part I. In Part II, these ratios were introduced and the predicted profile moved significantly closer to the actual postsurgical outcome than if using the pre-programmed ratios (p < 0.05). CONCLUSION: Being able to customize the ratios of soft to hard tissue changes after a particular type of orthognathic surgery will enhance the accuracy of a patient's predicted profile outcome. These ratios should be pre-programmed in future versions of the software. PMID- 10534985 TI - Piggyback archwires. AB - Commonly, clinicians' principal tool in the alignment phase of orthodontic mechanotherapy is the nickel-titanium wire. During the course of orthodontic treatment, however, there are times when some segments of the dental arch require flexible wires, while the rest would benefit from rigid wires. In this report, we describe a technique where both of these needs are satisfied simultaneously. Specifically, a segment of nickel-titanium wire is piggybacked onto a stainless steel wire in regions where flexibility is desired. This method eliminates the problems associated with the activation, de-activation forces created along a continuous archwire and might be more economical. Clinical pictures illustrate the point. PMID- 10534986 TI - Patient and parent preferences for orthodontic practices. AB - This study was designed to identify who chooses an orthodontic office and what factors might induce the attraction. Patients and parents from the lists provided by suburban orthodontic offices were contacted. A mail-out survey instrument was used to gather the data. Results revealed that the reputation of the practitioner was most important along with the level of caring attitude the office projected. It was also important that the office is located near home, interestingly, the mother is the most significant decision-maker in the family in choosing an orthodontic office. Moreover, not the cost of treatment but the payment plan was the critical element in the decision process. The higher income families with three or less children were attracted to office characteristics such as excellence of the orthodontist, attention, and convenience. A marketing strategy based on these elements might provide the best return on the investment. PMID- 10534987 TI - The effect of surgically induced anterior disc displacement of the temporomandibular joint on the midface and cranial base. AB - OBJECTIVE: The purpose of this study is to investigate the effect of unilateral TMJ disc displacement on the midface and cranial base. STUDY DESIGN: Thirteen 10 week-old rabbits, eight controls and five experimental were included in this study. The five experimental rabbits had surgically created unilateral disc displacement. Animals were sacrificed at 22 weeks and frontal, occlusal, and lateral oblique radiographs were made of the skulls. RESULTS: The occlusal radiograph demonstrated that the glenoid fossa on the experimental side was located more anterior. The oblique radiograph demonstrated the root of the zygomatic arch the experimental side was inferior. The anterior aspect of the fossa was more inferior on the frontal radiograph. CONCLUSIONS: A previous study suggested a shortening in the ramal height. This study suggests an alteration of the cranial articular fossa. Thus, it is suggested that disc displacement is capable of producing asymmetry in the developing mandible and cranial base. PMID- 10534988 TI - Condylar bony change and craniofacial morphology in orthodontic patients with temporomandibular disorders (TMD) symptoms: a pilot study using helical computed tomography and magnetic resonance imaging. AB - OBJECTIVE: To investigate how condylar bony changes relate to craniofacial morphology using helical CT and MRI. DESIGN: Craniofacial morphology of orthodontic patients with condylar bony changes was compared with Japanese standard. SETTING AND SAMPLE POPULATION: The Department of Orthodontics, Niigata University School of Dentistry, Twenty-nine subjects were selected from orthodontic patients (six males and 23 females, a mean age of 18.8 +/- 6.3 years) who were diagnosed by helical CT as having condylar bony changes. EXPERIMENTAL VARIABLE: Subjects were divided into two groups: a unilateral condylar bony change group (unilateral group) (four males and nine females) and a bilateral condylar bony change group (bilateral group) (two males and 14 females). OUTCOME MEASURE: Condylar bony changes were evaluated on reconstructed coronal and sagittal CT scans. Disk positions were evaluated by MRI scans. Five linear and four angular measurements in lateral and posteroanterior cephalograms were compared with those of an age- and sex-matched 'standard population' from the Japanese standard. RESULTS: In the bilateral group, osteophyte formation and erosion were the common bony changes and were present in adult as well as juvenile subjects. In the unilateral group, flattening was the most common features and erosion was only present in subjects below 19 years. Disk displacement without reduction was seen in 90.6% of the bilateral group, and in 76.9% of the unilateral group. Retrognathic mandibles were shown in the bilateral group. All subjects exhibited a lateral shift of the menton toward the condylar bony changed side in the unilateral group. CONCLUSION: Condylar bony changes might be progressive and unstable in adults of the bilateral group as well as in juveniles of the both groups. It appears that condylar bony changes may be related to a lateral shift of the mandible and a retrognathic mandible in orthodontic patients with TMD symptoms. PMID- 10534989 TI - An in situ caries model to study demineralisation during fixed orthodontics. AB - OBJECTIVES: To investigate de/remineralization of enamel during the early stages of orthodontic treatment using the in situ caries model. DESIGN: A prospective, longitudinal study, using the in situ caries model. SETTING AND SAMPLE POPULATION: The Department of Orthodontics at the University of Liverpool School of Dentistry. Fifteen orthodontic patients undergoing fixed appliance treatment with extraction of premolar teeth. EXPERIMENTAL VARIABLE: Two enamel samples with pre-formed caries-like lesions were placed bilaterally, in specially constructed holders, on an orthodontic fixed appliance. One sample was bonded with a small bracket base. OUTCOME MEASURE: The parameters of the pre-formed carious lesion, expressed as mineral loss (delta Z), lesion depth (ld), lesion width (lw) and ratio (delta Z/ld) were compared between the bracketed, the non-bracketed and a control sample that had not been placed in the mouth. The difference between brackets place on the dominant (toothbrush hand) side and non-dominant side were also investigated. The correlation between mineral loss and length of time the sample was in the mouth was also analysed. RESULTS: There was considerable individual variation; however, a one-factor repeated analysis of variance showed a significant difference in ratio values between the three groups (p = 0.006). A pairwise comparison showed a significant reduction in ratio value for the non bracketed sample compared with the control, but not the bracketed sample. There was no significant difference in mineral loss between the dominant and non dominant sides. There was no linear correlation between the length of time the sample was in the mouth. CONCLUSION: An enamel sample with a pre-formed carious lesion, when placed in the mouth of an orthodontic patient, showed reduced remineralization in the presence of a simulated orthodontic bracket. Consistently effective preventive regimes to prevent demineralization in patients with fixed orthodontic appliances need to be developed. The technique described will be a valuable tool in this process. PMID- 10534990 TI - Quantitative description of overjet and overbite and their relationship with the craniofacial morphology. AB - OBJECTIVES: 1) To define the sagittal and vertical characteristics of anterior teeth in adults with normal occlusions; 2) to explore a relationship between the overbite and overjet; and 3) to relate overbite and overjet to the skeletal pattern. DESIGN: Prospective data collection. SETTING AND SAMPLE POPULATION: Ninety-two adult dental students from the Aristotle University of Thessaloniki (49 females and 43 males) with naturally occurring Class I occlusions. EXPERIMENTAL VARIABLE AND OUTCOME MEASURES: Cephalometric data were collected for overbite, overjet, and skeletal relationships. These were then correlated for potential association between front teeth and vertical and horizontal skeletal relationships. RESULTS: The overjet measures were equally distributed among men and women, but overbite was higher in women. Facial proportions were also bigger in men, but the Mediterranean face was bigger than Northern American Caucasian. The mandibular plane angle could be associated with either increased or decreased overjet and overbite. CONCLUSION: The overbite and overjet features of an occlusion cannot be predictably associated with any particular craniofacial pattern. PMID- 10534991 TI - The mechanical advantage of the masseter muscle in subjects with different vertical and sagittal facial morphology. AB - OBJECTIVES: To study the relationships between the mechanical advantage of masseter muscle (the ratio between the muscular and the bite force moment arms) and two parameters describing vertical and sagittal facial morphology. DESIGN: Retrospective analysis of pre-treatment lateral cephalometric radiographs. SETTING AND SAMPLE POPULATION: The Laboratory of Functional Anatomy of the Stomatognathic Apparatus at Milan University, Italy. Four hundred and sixty-eight orthodontic patients (209 males, 259 females) aged between 6 and 50 years. MEASUREMENTS: Mechanical advantage of masseter muscle, posterior-to-anterior facial height ratio, corrected ANB* angle were measured in all cephalograms. RESULTS: Masseter muscle mechanical advantage was, on average, 79.5% (SD 6.1), ranging between 53 and 94%. No sex-related characteristics were found. A moderate effect of age was found (r = 0.19), with a certain increment in the mean mechanical advantage in adults. No relationship with the corrected ANB* angle was found (r = 0.099), while a significant effect of the posterior-to-anterior facial height ratio was detected (r = 0.542). CONCLUSION: The lack of correlation between the mechanical advantage of masseter muscle and corrected ANB* angle might be explained by: (1) when the occlusal point moves depending upon Angle skeletal classification, the ramus width might also modify, and both arms change; (2) the present assessment analysed only the lateral two-dimensional projection of a three-dimensional structure: the retroposition of occlusal point involves also a modification of line slope, and a subsequent reduction of its projection and arm. PMID- 10534992 TI - New horizons in understanding early tooth development. AB - Information on the mechanism that governs tooth shape, size, and location in the developing jaw is sparse. One hypothesis holds that exogenous signals from the ectomesenchyme and epithelium of the embryonic jaw induce a specific pattern of cellular development that leads to tooth formation. A second hypothesis is that a specific tooth type develops from select clones of cells. The objective of this review is to consider these ideas in light of recent studies on genes that regulate embryonic patterning. Special attention is directed at the notion that there is restricted expression of specialized genes. These genes are expressed in overlapping domains along the jaw axis. Genes expressed within a region provide a unique homeobox code that serves to specify a tooth type and ultimately determines tooth form and position in the developing jaw. Expression of this genetic 'bar code' is carefully controlled by the dental epithelium, neural crest derived cells, and growth factors. The latter agents also probably serve as epithelial signals that activate events that lead to odontogenic differentiation and morphogenesis. PMID- 10534993 TI - Update on the Fujita lingual bracket. PMID- 10534994 TI - Osseous ridge augmentation with orthodontic extraction. PMID- 10534995 TI - Lever-hook edgewise arch for midfacial protraction. PMID- 10534996 TI - Polyolefin foam protective sports mouthguard. PMID- 10534997 TI - Temporary bite raiser. PMID- 10534998 TI - Improving incisor torque control with nickel titanium Torque Bars. PMID- 10535000 TI - The .018" nickel titanium stop for prevention of archwire crawl. PMID- 10535001 TI - By the numbers. AB - Any statement about the ability of the specialty to meet the future demand for orthodontic care based merely on a ratio of the projected number of orthodontists to the projected population is an oversimplification. Still, some inferences can be drawn from the data presented in this article. To be as prudent as possible, we will limit the scope of the discussion to the next 10 years. 1. It seems likely that the annual number of orthodontic graduates will remain about the same. 2. Considering the slow increase in the average age of orthodontists, the death rate will increase slightly, but not significantly. 3. The average annual number of retirees will be somewhere between 125 (the current rate) and 359 (the maximum projected by the JCO Retirement Survey). 4. The above assumptions would leave no less than 7,500 practicing orthodontists in the United States in 10 years. 5. The number of children age 9-17 will increase by 5-10%. In a worst-case scenario, could 7,500 orthodontists meet the orthodontic demand 10 years from now? The 1997 JCO Orthodontic Practice Study reported the annual median number of case starts to be 180. Multiplied by 9,000 orthodontists, that would equal 1,620,000 case starts. If we project a 10% increase in demand, there could be 1,782,000 case starts 10 years from now--an average of 238 starts per year for 7,500 orthodontists. Respondents to the Practice Studies have consistently indicated that they could handle 50 additional case starts with no increase in staff or facilities, which would mean current orthodontists could accommodate an average of 230 case starts. Moreover, orthodontic productivity is likely to increase due to delegation and improved technology. In conclusion, even if the sharp increase in the number of annual retirements anticipated in the JCO Retirement Survey turns out to be correct, it seems unlikely that it will affect the ability of the specialty to accomplish its mission in the foreseeable future. PMID- 10534999 TI - CAD/CAM fabrication of occlusal splints for orthognathic surgery. PMID- 10535002 TI - Efficacy of open-bite treatment with the Thera-spoon. PMID- 10535004 TI - Nonsurgical treatment of severe mandibular prognathism. PMID- 10535003 TI - SEM evaluation of a new technique for interdental stripping. PMID- 10535005 TI - A new and improved indirect bonding technique. PMID- 10535006 TI - An orthodontic attachment for patients with fixed prosthetic restorations. PMID- 10535007 TI - The 30-second difference. PMID- 10535008 TI - Glass ionomer cement dressing for surgically exposed impacted teeth. PMID- 10535009 TI - The SUPERspring II: a new appliance for non-compliant Class II patients. PMID- 10535011 TI - Alignment of impacted canines with cantilevers and box loops. PMID- 10535010 TI - Complex surgical-orthodontic case with a transposed cuspid and mandibular asymmetry. PMID- 10535012 TI - Criteria used by general dentists to choose an orthodontist. PMID- 10535014 TI - Precision finishing in lingual orthodontics. PMID- 10535015 TI - The SPEED bracket auxiliary slot. PMID- 10535013 TI - Effects of head posture on headgear force application. PMID- 10535016 TI - The First Class Appliance for rapid molar distalization. PMID- 10535017 TI - Retention strategies: a pilgrim's progress. PMID- 10535018 TI - Bleaching teeth during supervised retention. PMID- 10535019 TI - Interdisciplinary treatment of a Class II deep-bite patient with missing and impacted teeth. PMID- 10535020 TI - A new supraconstruction for palatal orthodontic implants. PMID- 10535021 TI - Ethics in orthodontic practice, Part 5. PMID- 10535022 TI - The dental VTO: an analysis of orthodontic tooth movement. PMID- 10535023 TI - Australian uprighting spring for partially impacted second molars. PMID- 10535024 TI - The DenOptix digital radiographic system. PMID- 10535025 TI - Early treatment in mixed dentition. PMID- 10535026 TI - Retention: a long and constant contract. PMID- 10535027 TI - Case report: transmigration of an impacted mandibular cuspid. PMID- 10535028 TI - The latest in hi-tech photography. PMID- 10535029 TI - Considerations and treatments to erupt impacted teeth. PMID- 10535030 TI - The new 4 D Arch Developer System. PMID- 10535031 TI - Twin block functional therapy: a modified technique. PMID- 10535032 TI - Twelve rules of orthodontic treatment during mixed dentition. PMID- 10535033 TI - The need for an orthopedic occlusal analyzer in orthodontics. PMID- 10535034 TI - Cephalometric variation in patients with and without intraoral neuromuscular repositioning appliance. PMID- 10535035 TI - Maximizing the benefits of resin-modified glass ionomer orthodontic adhesives. PMID- 10535036 TI - [Data without gaps: a genuine contribution to quality assurance in heart surgery]. PMID- 10535037 TI - [30-day mortality after heart surgery: a pilot project of the Working Party of Directors of Hospital Cardiology Departments]. AB - BACKGROUND AND OBJECTIVE: 30-day mortality after operation is generally accepted as a central standard of quality, especially in regard to cardiac operations. The Working Party of Directors of Hospital Cardiology Departments (Arbeitsgemeinschaft Leitender Kardiologischer Krankenhausarzte, ALKK) in Germany set up a pilot project to analyse whether by direct communication with patients by a database centre the expenditure incurred in collecting complete data can be decisively reduced and full documentation of outcome can in this way be obtained even for a large multi-centre patient cohort. PATIENTS AND METHODS: Between 1.6.1997 and 31.3.1998, data were consecutively collected by questionnaire on all patients registered for a cardiac operation at 85 of the 135 ALKK centres. The questionnaire included data about each patient and the indication for operation as well as the estimate of operative risk, assigned to one of five risk categories by the referring cardiologist either alone or in conjunction with the cardiac surgeon. RESULTS: Until 30.9.1998, the data were obtained on 11,349 patients who had given informed consent (response rate 99.99%), including survival figures. 824 (7.3%) patients had not undergone the planned cardiac surgery, 134 having died before the data of operation. The 30-day postoperative mortality, obtained in 99.99% of the 10,525 patients, was 3.92%. The operative mortality was lowest, at 3.73%, for aortocoronary bypass only (n = 7932), highest for aortocoronary bypass plus valvular operation (n = 785), at 8.04%. There was good agreement between the cardiologists' preoperative risk assessment and the observed mortality. CONCLUSIONS: The 30-day mortality after cardiac operation can be obtained almost completely and with reasonable expenditure even for a large patient cohort. The results confirm that hospital mortality data definitely understate the overall operative risk. The methodology used in this pilot project, namely the inclusion of information from the patients by questionnaire, can also be applied to clinical results in other areas. PMID- 10535038 TI - [Transitory left ventricular outflow tract obstruction after mitral valve reconstruction]. AB - HISTORY AND ADMISSION FINDINGS: A few days after uneventful surgical reconstruction of the mitral valve a 43-year-old man was found to have a systolic murmur due to prolapse of the posterior leaflet, suggesting renewed mitral regurgitation. INVESTIGATIONS: Echocardiography revealed haemodynamically significant left ventricular outflow tract obstruction (LVOT) with a left ventricle to aorta systolic gradient of 83 mm Hg. In addition there was moderately severe mitral regurgitation as well as a pericardial effusion but no signs of tamponade. TREATMENT AND COURSE: The obstruction was at first treated with verapamil, later with sotalol. The pericardial effusion was interpreted as part of a postcardiotomy syndrome. The effusion regressed under steroid administration, and the LVOT and mitral regurgitation also decreased. A provocation test five months postoperatively no longer brought about an outflow gradient. The good results were still present 12 months postoperatively. CONCLUSION: The described, rarely seen form of LVOT was probably caused by a combination of a very large anterior mitral leaflet, postoperative pericardial effusion and pharmacological effects. If the obstruction first occurs postoperatively, appropriate medication may improve the cardiac status and reoperation may be avoided. Echocardiography is an important method of diagnosis and serial monitoring. PMID- 10535039 TI - [Methodical improvements of exercise echocardiography]. PMID- 10535040 TI - [Hyperhomocysteinemia, a risk factor for atherosclerosis: causes and effects]. PMID- 10535041 TI - [Medical care in the Federal Republic of Germany with special consideration of cardiology as a priority subject. Current state and prospects]. PMID- 10535042 TI - [Transesophageal echocardiography as a standard examination in sepsis?]. PMID- 10535043 TI - [Evidence based medicine using heart failure as a paradigm]. PMID- 10535044 TI - [Treatment of chronic heart failure with digitalis: are there still indications?]. PMID- 10535045 TI - [ACE inhibitors or AT1 receptor antagonists?]. PMID- 10535046 TI - [Value of diuretics and aldosterone antagonists in the treatment of heart failure]. PMID- 10535047 TI - [Drug therapy of chronic heart failure: when are beta receptor blockers indicated?]. PMID- 10535048 TI - [Emergency treatment of acute heart failure]. PMID- 10535049 TI - [Diagnosis of acute and chronic heart failure]. PMID- 10535051 TI - Two neuraminidase inhibitors for treatment of influenza. PMID- 10535050 TI - [The preoperative care in noncardiac surgery of patients with heart failure]. PMID- 10535052 TI - Zaleplon for insomnia. PMID- 10535054 TI - [Influenza pandemics: past and future]. AB - The circulation of numerous influenza virus subtypes in water birds constitutes a continuous threat with respect to emergence of a future influenza pandemic. The interspecies barrier is high but has already been breached 3 times this century, notably in 1918, 1957 and 1968. Still, it is impossible to predict when another pandemic will occur. Continuous and intensive surveillance of influenza viruses in man, pigs, and birds, especially in China, may provide the opportunity to prepare a vaccine timely in sufficient amounts, at least for the industrialised countries. PMID- 10535053 TI - [Unobserved death of an infant: cot death?]. AB - Three children, two girls aged 8 and 12 months and one boy aged 7 weeks, were found dead unexpectedly. Autopsy revealed pneumonia in two children, following which the diagnosis of 'natural, explained death' was made; one child showed no abnormalities and the diagnosis read 'natural, unexplained death' (cot death). Autopsy may currently only be performed with parental permission or, in case of doubt about unnatural cause of death, by order of the public prosecutor. The authors propose routine performance of a protocolled autopsy by GP, pediatrician, pathologist and medical examiner in order to avoid subsequent and possibly incorrect doubt about the cause of death. PMID- 10535055 TI - [Role of chemo(radio)theray prior to surgery for stage IIIa non-small cell lung carcinoma (localized non-resectable tumor)]. AB - Neoadjuvant chemotherapy with or without combined radiotherapy followed by surgery may yield a survival profit for selected patients with non-small cell lung carcinoma in stage IIIA. It has not yet been established which is the most efficacious neoadjuvant chemotherapy nor what is the best postneoadjuvant chemotherapy, especially resection or radiotherapy. Comparative clinical studies to find the answer to these two questions are in progress. New forms of treatment are required in order to achieve 5-year survival in more stage IIIA patients than is currently the case. PMID- 10535056 TI - [Roaming through methodology. XVI. What to do about missing data]. AB - Medical scientific research involving multiple measurements in patients is usually complicated by missing values. In case of missing values the choice is to limit the analysis to the complete cases or to analyse all available data. Both methods may suffer from substantial bias and may only be applied in a valid way if the rather strong assumption of 'missing completely at random' holds for the missing values, i.e. the missing value is not related to the other measured data nor to unmeasured data. Two other statistical methods may be applied to deal with missing values: the likelihood approach and the multiple imputation method. These methods make efficient use of all available data and take into account information implied by the available data. These methods are valid under the less stringent assumption of 'missing at random', i.e. the missing value is related to the other measured data, but not to unmeasured data. The best approach is to ensure that no data are missing. PMID- 10535057 TI - [Prevalence of asymptomatic cardiac ischemia in men with diabetes mellitus type I]. AB - OBJECTIVE: To establish the prevalence of 'silent ischaemia' of the myocardium in male patients with type 1 diabetes mellitus using a non-invasive cardiac examination, and to determine what clinical variables are related to silent ischaemia. DESIGN: Prospective, cross-sectional. METHOD: Males aged 20-69 years who visited the outpatient department of Internal Medicine of the De Weezenlanden Hospital in Zwolle between 1 February 1992 and 31 January 1995, and who showed no symptoms of ischaemic cardiopathy (angina pectoris, myocardial infarction or arrhythmias) or of chronic obstructive pulmonary disease, were examined for cardiac ischaemia by means of a 24-hour Holter registration and a perfusion scintigram after administration of dipyridamol. In order to demonstrate a possible connection between cardiovascular risk factors and silent ischaemia, the patients with an abnormal and those with a normal scintigram were compared by means of multivariate analysis. RESULTS: Data were collected on 92 successive patients, with a median age of 40 years (range 22-69). There were 19 patients (21%) with an abnormal myocardial scintigram. On average they were older and had a longer history of diabetes mellitus. An abnormal Holter registration was observed in 14 patients (15%), abnormality of either the Holter registration or the myocardial scintigram in 28 patients (30%) and abnormality of both the myocardial scintigram and the Holter registration in 5 patients (5%). The duration of the diabetes mellitus, and a diastolic blood pressure > or = 90 mm Hg were statistically significant and independent predictors of an abnormal myocardial scintigram (relative risks 1.08 and 3.4 per year, respectively). CONCLUSIONS: The prevalence of cardiac ischaemia in males with type 1 diabetes mellitus without cardiac symptoms is approximately 20%. Abnormal test results were associated with a longer duration of the diabetes mellitus and a diastolic blood pressure > or = 90 mm Hg. PMID- 10535058 TI - [Metabolic acidosis caused by hyperalimentation; a dangerous complication of parenteral nutrition]. AB - A man aged 46 developed oesophageal obstruction due to carcinoma. He was admitted with dehydration following strongly reduced fluid intake leading to prerenal kidney failure. He had had a renal transplantation, after which a moderate renal insufficiency persisted. During 12 days he received parenteral nutrition upon which he developed severe hyperchloraemic metabolic acidosis. The treatment of this condition consists of discontinuation of the parenteral nutrition and administration of bicarbonate. If the parenteral nutrition needs to be continued, the amount of chloride must be diminished and acetate be added. It is important to monitor the acid-base balance and plasma electrolytes in patients with parenteral nutrition, especially in the presence of renal failure. The patient recovered from the metabolic acidosis but later died of metastatic oesophageal carcinoma. PMID- 10535059 TI - [From Miescher to molecular DNA technology; a chapter from the medical history of the past century]. AB - Although molecular biology is a young discipline, it originated in the second half of the 19th century with the simultaneous discoveries of the laws of heredity by Mendel and of nucleic acid by Miescher. It was not until about 1950 that the structure of DNA was determined and it was proved that DNA governs the hereditary properties. Subsequently, the developments followed in rapid succession with the unravelling of the hereditary code, the elucidation of the mechanism of the translation of DNA into proteins, the discovery of the structure of genes and the finding of the methods for genetic manipulation. These have proved essential for the evolution of modern biotechnology. PMID- 10535060 TI - [Influenza season 1998/99; composition of vaccine for 1999/2000]. AB - The first indication of influenza activity in the Netherlands in the 1998/'99 season was the isolation of an influenza B virus in week 47 of 1998. In subsequent weeks influenza activity slowly increased, reaching a peak in week 6 of 1999. After a gradual decline for three weeks a second peak was reached in week 8 of 1999. The first wave of influenza activity was primarily caused by influenza B viruses, whereas during the second wave predominantly influenza A viruses of the A/H3N2 subtype were isolated. The antigenic properties of the influenza A viruses resembled those of the viruses isolated in the previous season and the vaccine strain A/Sydney/5/97. The influenza B viruses did not completely match with B/Harbin/7/94 which is most commonly used for vaccine production. The vaccine, however, did provide good protection against the epidemic strains of influenza. This season influenza A/H1N1 viruses did not play a significant part and only a small number of viruses of this subtype were isolated at the end of the season. For the influenza season 1999/2000 it is recommended by the World Health Organization that the vaccines contain the following (or similar) virus strains: A/Sydney/5/97 (H3N2), A/Beijing/262/95 (H1N1) and B/Beijing/184/93. PMID- 10535061 TI - ['Osteoporosis' guideline from the Dutch Society for General Practitioners]. PMID- 10535062 TI - [Missed injuries upon admission of severely injured trauma patients to the emergency department]. PMID- 10535063 TI - [Concepts for measuring quality of life in patients with chronic sinusitis]. AB - BACKGROUND: In 1997, Benninger developed the Rhinosinusitis Disability Index (RSBI) for patients who suffered from chronic sinusitis. Its content related validity and construct related validity were established, as were its sensitivity and reliability. The aim of our study is to introduce a concept to measure quality of life in these patients according to circumstances prevalent in Germany. METHODS: The questionnaire (Rhinusitis-Beeintrachtigungs-Index, RSBI) contains 30 questions that describe the 5 aspects of quality of life. Moreover we used a standardized data sheet to acquire further information about history and diagnostic results. Using this method it is possible to specificity more precisely the patients current complaints and condition. Evaluation is possible according to complex of questions as well as simple items. The influence of treatment on quality of life can be measured as the sum of the specific scores and is expressed comprehensively in the total score. The concept of the study includes a prospective inquiry as well as before and after surgical intervention. CONCLUSION: A disadvantage of the American RSDI is that the clinical symptoms are not described in a detailed catalogue which allows the correlation of the clinical factors. Therefore we added a detailed questionnaire concerning the clinical symptoms and the individual treatment of the patient (RSBI). That permits comprehensive analysis of quality of life as it relates to different aspects of disease and different strategies in treatment. PMID- 10535064 TI - [Cytokines and chemokines in paranasal sinus diseases]. AB - BACKGROUND: New insights into inflammatory processes became possible by investigating the pattern of cytokines and chemokines as well as adhesion molecules in different acute and chronic sinus diseases since the last decade. This review aims to update and discuss findings of in vitro and in vivo studies concerning the role of cytokines and chemokines in inflammatory sinus diseases during the last years. RESULTS: Discrepancies in research findings may be due to the small available databases today, the use of different techniques for investigation, and the lack of a valuable classification of sinus diseases. Despite this discrepancies, there is evidence that in acute bacterial and viral sinusitis, proinflammatory cytokines play a dominant role in initiating and sustaining the inflammation, which is especially characterized by neutrophil tissue infiltration. In chronic sinusitis IL-3 dominates the cytokine profile, giving support to a variety of inflammatory cells. IL-3 may also contribute to fibrosis and constant thickening of the mucosa leading to an obstruction of the ostiomeatal complex. In most bilateral nasal polyps, tissue eosinophilia is a striking finding which is believed to play a central role in pathogenesis, as it does in asthma. Eosinophilia may be explained by increased migration and prolonged eosinophil survival. Activation and survival of eosinophils in nasal polyps are thought to be regulated by autocrine stimulation by IL-5. Therefore, IL-5 represents the main target for future therapy in nasal polyposis. CONCLUSIONS: Defining cytokine and chemokine patterns in inflammatory sinus diseases leads to a better understanding of immunologic processes in nasal mucosa. Results of cytokine and chemokine research are of paramount interest in developing new therapeutic approaches. PMID- 10535065 TI - [What are the side-effects of nocturnal continuous positive pressure ventilation (nCPAP) in patients with sleep apnea for the head-neck region?]. AB - BACKGROUND: Nasal continuous positive airway pressure ventilation therapy is the present gold standard in the treatment of obstructive sleep apnea. Depending on the definitions used, about 60% of the patients tolerate nCPAP therapy. The reason for this limited tolerance is a varying number of side effects. The aim of the present retrospective study was to analyze the incidence and intensity of otorhinolaryngological side effects of nCPAP therapy. METHOD: Questionnaires inquiring about the frequency of using nCPAP and objective and subjective complaints were sent to 92 patients who were treated with nCPAP in our department within the last years. Six of 92 patients also used a heated humidifier. A telephone interview was added to complete the questionnaires as correctly as possible. RESULTS: Eighty questionnaires were sent back completely answered. The mean frequency of using nCPAP (+/- standard deviation) was 6.8 +/- 1.6 hours per night and 6.5 +/- 1.4 nights per week. The mean duration of nCPAP therapy was 28.0 +/- 21.0 months; the mean pressure used was 6.8 +/- 1.2 cm H2O. The following side effects were specified most frequently: disturbance of sleep during the night (71.3%), dry mouth (47.5%), dry nose (46.3%), pressure marks by the mask (41.3%), crusts within the nasal cavity (38.8%), and hearing loss (26.3%). Dryness within the nose and mouth was considered the most irritating side effects. CONCLUSIONS: NCPAP therapy has a number of different side effects in the head and neck. These side effects are seen frequently. Prospective analysis must show whether there are correlations between the intensity and frequency of side effects and the duration of therapy, and whether technical improvements (quality of masks, noise reduction, humidifiers) are able to reduce the frequency of side effects. PMID- 10535066 TI - [Role of diagnostic imaging in chronic recurrent parotitis in childhood]. AB - BACKGROUND: Chronic recurrent parotitis represents a disease of still uncertain etiology that can be divided into a childhood and an adult form. Sialography is still the imaging procedure of choice. METHOD: We report about a nine-year-old girl who had had six episodes of parotitis in the past two years. MR sialography is presented as an alternative to conventional sialography. MR was performed with a 1.5 T wholebody tomograph using a high resolution 3D-CISS sequence. RESULTS AND CONCLUSION: MR sialography does not require any contrast medium. It primarily images liquid structures, and the flow of saliva can be adequately demonstrated after stimulation with ascorbic acid. These special features of MR sialography allow its use also during acute episodes of sialadenitis, thus providing a significant improvement over conventional sialography. Furthermore, no overlapping occurs as in conventional summation images, due to the multiplanar reconstruction features. PMID- 10535067 TI - [Immunohistochemical detection of extracellular matrix proteins in the irradiated rat submandibular gland]. AB - BACKGROUND: Extracellular matrix proteins (EMP) form the main components of basement membranes (BM) and associated structures. Basement membranes influence tissue structure, exchange of substances, and cell growth and differentiation. While BM changes have been reported in malignant tumors and in inflammatory diseases, systematic studies of BM changes following irradiation are rare. Because functional and histomorphological damage to the salivary glands is a well known sequela of radiotherapy in humans, our goal was to describe possible BM changes using an experimental model and immunohistochemistry (IH). METHODS: In 59 rat mandibular glands we investigated the distribution of EMP by IH. The animals differed in age from 3 months to 2 years and pre-treatment status (irradiation vs no irradiation). The analyses were performed at selected time points after completion of external irradiation (< 4 months [M]/> or = 4-6 M/> 6 M). RESULTS: The polyclonal antibodies (anti-laminin [AL], anti-fibronectin [AF], anti collagen III [AC-III], anti-collagen IV [AC-IV]) identified components of BM (laminin, collagen IV) and BM-associated structures (fibronectin, collagen III) in glandular tissue, in vascular walls, in nerve tissue, and the interstitial connective tissue. Various EMP were detected in different patterns. AL stained nerve tissue moderately and both different gland tissue structures and vascular walls (without adventitia) slightly, while excretory ducts and blood vessel adventitia were distinctly positive for AF. Fibronectin was also present in the connective tissue stroma. Significant differences were seen between irradiated and nonirradiated glands, often with generally stronger and more extensive staining in the irradiated group. For example most tissue structures showed distinctly increasing immunostaining for AL in positive correlation with increasing latency of irradiation up to 6 months. The age was of minor importance. CONCLUSIONS: Previous irradiation has to be considered when interpreting the EMP profile of salivary gland tissue, especially in studies both on chronic degenerative diseases and tumor differentiation developing in an area that has undergone radiation therapy. PMID- 10535068 TI - [Indications for vocal cord augmentation with collagen]. AB - BACKGROUND: Vocal fold augmentation through an indirect laryngoscopic injection has been largely supplanted by the external approach of vocal fold medialization. Vocal fold augmentation with collagen is still of clinical importance for the temporary treatment of glottic insufficiencies. The different elimination time of certain collagen compounds in the vocal fold tissue enables individual therapeutic applications. PATIENTS AND METHODS: The collagen compounds Zyderm I and II and Zyplast were injected into the vocal fold in 37 patients with neurogenic glottic insufficiency and 2 patients with vocal fold atrophy during indirect laryngoscopy. The choice of compound was made depending on the case, prognosis, severity of the disorder, the persistence of the glottic insufficiency, and the patient's age and condition. RESULTS: Thirty-three patients showed an significant improvement of the glottic closure. However, 6 patients showed hardly any change. CONCLUSION: Collagen augmentation can be used for the temporary medialization of the vocal fold. It is a suitable method for the treatment of vocal fold paralysis a) within the neural regeneration time, and b) in patients with reduced condition and/or a short life expectancy due to severe diseases. PMID- 10535069 TI - [Comparative voice quality evaluation after laser surgical versus fronto-lateral partial laryngectomy in T1b and T2 vocal cord carcinoma]. AB - BACKGROUND: We conducted a study to compare the voice quality after transoral endolaryngeal laser surgery versus anterolateral partial laryngectomy in terms of ability to communicate effectively. MATERIAL AND METHODS: Two groups each of 8 patients were reexamined at least 6 months after either a laser surgical or a partial anterolateral laryngectomy for a T1b or T2 vocal chord carcinoma was performed by the same surgeon. The following examinations were conducted: video laryngoscopy, video-stroboscopy, phonetogram when reading a standard text or when speaking and shouting, voice load test, respiratory flow measurement, auditive voice rating following the RBH model, and auditive rating of the voice by the patients according to a questionnaire. RESULTS: A voice disorder with a medium grade dysphony and an essential limitation of the speaking and shouting voice function and voice quality was detected in both groups of patients. No significant differences were observed in any of the parameters evaluation in both groups of patients. CONCLUSIONS: Both surgical techniques appear to be equivalent in terms of postoperative voice function. However, the tracheotomy was avoided in the patients undergoing laser surgery. PMID- 10535070 TI - [Functional esophageal scintigraphy (99mTc-DTPA)1.2. Presentation of a method for diagnosis of effective deglutition in patients with oral and hypopharyngeal tumors]. AB - BACKGROUND: Today more than 60% of 65-year-olds complain of dysphagia or odynophagia. The aim of this study was to use esophageal scintigraphy as a means of estimating quantitative, reproducible, and comparable parameters of the swallowing ability in patients with tumors of the oropharynx and hypopharynx before and after surgery. Another group of patients suffering from larynx tumors was also examined prior to and following laryngectomy. METHODS: Fifteen male patients with T3/4 tumors of the oropharynx or hypopharynx, which were dissected followed by plastic surgery with a radialis flap, were examined before and after surgery. Esophageal scintigraphy was performed in 10 patients in this age group with T3/4 larynx tumors. The mean transit time (MTT in seconds) and clearance (in%) were the parameters evaluated. Condensed images were determined by averaging. Analog Polaroid images were also obtained. RESULTS: The 15 patients who underwent radialis flap surgery as a reconstructive measure showed a 60% improvement in the ability to swallow. Four of the 10 patients showed an obvious postoperative improvement in the ability to swallow as determined by clearance. With respect to the mean transit time, 2 cases out of 10 showed an improvement in the swallowing process. CONCLUSIONS: In patients who underwent reconstructive radialis flap surgery, it was possible to reestablish the continuity of the orohypopharynx, which is a prerequisite for the restoration of the swallowing facilitation. As a result one can expect an improvement in swallowing kinetics postoperatively. The patients who underwent laryngectomy required adaptation time to learn how to swallow under altered anatomical conditions. The primary reason for this is the altered pressure in the hypopharynx. Esophageal scintigraphy provides easily comparable parameters that serve as an ideal method for the objective determination of the ability to swallow in patients with these tumors. This method is neither poorly tolerated nor invasive and compared to other methods is a less costly alternative for examining the swallowing process. PMID- 10535071 TI - [Halitosis--foetor ex ore]. AB - BACKGROUND: An overview is presented on the etiology, diagnosis, and therapy of halitosis. METHODS: Results are given of our multidisciplinary halitosis outpatient department started in 1994. The team consists of ENT specialists and paradontologists, occasionally assisted by a psychiatrist. The oral odor is confirmed with a halitometer (Interscan Corporation, Model RH-17E USA). 491 Patients, nearly the same number of males as females, mostly between 20 and 50 years of age were seen. RESULTS: Oral causes (87%) were due to tongue coating (51%), gingivitis (17%), paradontitis (15%), or combinations of factors (17%). The other 13% involved causes related to ENT problems (4%), both ENT and oral (3%), digestive tract (1%), and presumed psychiatric pathology (5%). CONCLUSIONS: Many patients underwent diagnostic and therapeutic aimed interventions to no avail prior to their arrival in our halitosis clinic. Usually advising the patient to maintain better oral hygiene is sufficient. PMID- 10535073 TI - [Surgical therapy of malignant and benign diseases of the hypopharynx. I]. PMID- 10535072 TI - [Interesting case no. 28. Orbital metastasis of established breast cancer]. PMID- 10535074 TI - [Instrumentation in craniovertebral junctional surgery]. PMID- 10535075 TI - [Influence of scalp shaving on prevention of postoperative intracranial infection]. AB - Shaving of the whole scalp is ordinarily performed prior to neurosurgical operation. Although it is performed in order to prevent postoperative intracranial infection, there has been no apparent basis for this practice published in previous reports. We examined whether shaving the whole scalp reduced the rate of postoperative infection or not. We divided 274 cases, who received their first intracranial operation in the last 2 years, into two groups; a whole shaving group and a partial shaving group. We compared the rate of postoperative intracranial infection between the two groups according to age, diagnosis, operation, operation time and placement of drainage. Overall, 12 cases out of 274 (4.38%) had postoperative intracranial infection. The long operation time and the long term placement of drainage mechanism increased the postoperative intracranial infection rate. There was no postoperative intracranial infection in 74 patients who received burr-hole/twist-drill operation. As for craniotomy/craniectomy operations, 7 cases out of 83 (8.4%) in the partial scalp-shaving group whole scalp shaving group and 5 cases out of 117 (4.2%) had postoperative intracranial infections. Thus, there was no significant difference in the rate of postoperative intracranial infection between the two groups, if anything, the whole scalp shaving group tended to show a higher rate. According to these results, partial scalp shaving did not increase the rate of postoperative intracranial infection. Considering that patients who have lost their hair find it embarrassing to return to society, it is well to know that the whole scalp shaving is not absolutely necessary for all first craniotomy. PMID- 10535076 TI - [A case with traumatic internal carotid artery dissection in which the extent of the pseudolumen was defined by MR angiography]. AB - We report a 29-year-old male with traumatic internal carotid artery (ICA) dissection who presented with cerebral ischemia developed after removal of a left acute subdural hematoma and external decompression. CT scans 4 days after the operation showed infarctions of the distribution of the bilateral cerebral hemispheres. Cerebral angiography on the 11th hospital day demonstrated narrowing of the extracranial internal carotid artery at C1-C2 vertebral levels. Slight arterial dilatation and retention of the contrast medium were found just above the narrowing segment, which was suspected to be a pseudolumen. Three D time-of flight MRA showed an intramural hematoma corresponding to the narrowing on the angiography. Original axial MRA images showed that narrowing of the lumen was surrounded by a crescent hematoma and that two flow velocity areas were in the area distal to the narrowing. High flow seemed to be ordinary artery flow rate. The low flow area, including turbulent flow, led to the retention of contrast medium mimicking a pseudolumen. Precise MRA imaging will bring us an accurate diagnosis of extracranial carotid dissections. PMID- 10535077 TI - [Congenital arteriovenous malformation of the scalp with a drainage to the transverse sinus: a case report]. AB - Arteriovenous malformation (AVM) of the scalp is uncommon, and a subtype which has connection with the intracranial dural sinus is extremely rare. Only 3 cases have been reported. We present a case of congenital AVM of the scalp which was connected with the intracranial venous sinus. A 27-year-old woman had been noted as having a pulsatile soft mass in the midline of the occipital region since her birth. She visited our hospital because of pain and enlargement of the mass. The patient had had no history of trauma. Physical examination revealed a pulsatile scalp mass in the midline of the occipital region, measuring 3.5 x 3.5 x 1 cm. A loud bruit was ausculated. Tenderness was noted. The skin over the mass was slightly reddish. No focal neurological deficits were noted. Plain skull films demonstrated a round defect in the midline of the occipital bone. Magnetic resonance imaging (MRI) demonstrated a subcutaneous mass with low signal intensity and an infratentorial mass with flow void. 3D CT angiograms demonstrated a subcutaneous vascular mass with a single large vein draining into the right transverse sinus. External carotid angiograms revealed a vascular lesion within the scalp with supply from the branches of the bilateral occipital arteries and the meningeal arteries. The nidus penetrated the skull and connected to a dilated varix, which had a draining vein shunting into the right transverse sinus. After embolization of the right meningeal feeding arteries, surgery was performed. The vascular lesion penetrated the skull and the dura. The infratentorial mass was in the epiarachnoid space and was fed by a small pial artery. The mass was excised completely after interruption of the pial artery and the draining vein. Postoperative course was uneventful. Histologically, the subcutaneous mass and infratentorial vascular mass were shown to be AVM and varix, respectively. PMID- 10535078 TI - [A surgically treated case of giant pseudoaneurysm arising as a complication following carotid endarterectomy]. AB - We reported a case of giant pseudoaneurysm (PA) originating at the left extracranial internal carotid artery after carotid endarterectomy (CEA). A 60 year-old man underwent CEA for severe stenosis of the left internal carotid artery. A day after the operation, a left cervical painful and pulsatile mass beneath the operative scar was noticed. Soon after that, the patient showed dyspnea, and intratracheal intubation was carried out. 6 days after CEA, a painful swelling reoccurred. A left carotid angiogram visualized a 5 x 5 cm sized PA in the region of the left carotid bifurcation and demonstrated patency of the two major carotid branches. After a balloon catheter had been inserted and inflated at the carotid bifurcation, the patient underwent a second operation. The bleeding from arteriotomy was well controlled by the inflated balloon catheter at the carotid bifurcation. The continuous SURGILENE suture appeared to be disrupted and a saccular PA was found to have arisen from a defect on the lateral surface of the carotid bifurcation. The suture was completely removed and arteriotomy was closed with a continuous SURGILENE suture again and some single sutures were added. A PA arising as a complication following CEA is rare, but the possibility of PA after CEA should be considered in any patient presenting with a painful or/and pulsatile mass in the neck at any time interval following the operation. Angiography at the early post-op stage is necessary to rule out the various complications after CEA. To control the bleeding, the balloon catheter was thought to be very useful. PMID- 10535079 TI - Spinal epidural varices. AB - A 16-year-old male experienced a sudden attack of back pain while walking through the corridor of school which required emergent hospitalization. Except for the back pain, no neurological symptoms were noted. Magnetic resonance (MR) imaging indicated an angiopathy-like flow void in the epidural region at Th 3-5 which seemed to explain the patient's back pain. Thoracic laminectomy at Th 3-5 and resection of the affected site were performed. Pathologically, the resected lesion only had a dilated normal vein and no findings indicating vascular deformity. The patient's outcome was good and no relapse of pain has occurred for about 2 years since the operation. Although some authors have reported vascular deformity with spinal epidural hemorrhage or varices with lumbar hernia of the intervertebral disc, there is no report concerning spinal epidural varices with pain only. The present case seemed to be a rare event and is reported here. PMID- 10535080 TI - [Cortical deafness due to bilateral temporal subcortical hemorrhages associated with moyamoya disease: report of a case]. AB - A 51-year-old male with moyamoya disease had experienced a left putaminal hemorrhage 14 years previously, in 1983. Left encephalo-duro-arterio-synangiosis (EDAS) was performed in that year. The left putaminal hemorrhage reoccurred in 1990 and in 1994. Left superficial temporal artery-middle cerebral artery anastomosis (STA-MCA anastomosis) was performed. He was symptom free after the surgery. However, he suddenly suffered from deafness in 1997, when computerized tomography (CT) revealed the right temporal subcortical hemorrhage. Deafness continued for three days. It gradually improved and completely disappeared in two months. The damage to the bilateral auditory radiation was suggested as the cause of deafness in this patient. Cortical deafness is a rare symptom that is most often associated with cerebral infarction. This may be the first report of moyamoya disease which developed cortical deafness. PMID- 10535081 TI - [Infratentorial hemorrhage following supratentorial surgery]. AB - Hemorrhage in regions remote from the site of initial intracranial operations is rare, but does occur. We report three cases of cerebellar hemorrhage that developed after supratentorial surgery, all of which had similar clinical findings and CT images. The first case was a 37-year-old man with a craniopharyngioma in the suprasellar lesion. Partial removal of the tumor was performed through frontal craniotomy and the translaminaterminals approach. A large quantity of cerebospinal fluid (CSF) was suctioned from the third ventricle during the operation, resulting in marked brain shrinkage. The second and third cases were 34- and 51-year-old women with unruptured right middle cerebral aneurysms. Clipping of the aneurysms through the pterional approach was performed in both cases. In the second case, CSF was suctioned in large quantity from the carotid and prechiasmal cistern at the operation, resulting in marked brain shrinkage. In the third case, however, only a small volume of CSF was suctioned from the carotid and prechiasmal cistern during the operation, and no marked brain shrinkage was observed. CT scan showed that the hematomas were located mainly in the subdural or the subarachnoid spaces over the cerebellar hemisphere and partially extending into the cerebellar cortex. The mechanism of cerebellar hemorrhage in these series of patients was thought to be multifactorial. The possible etiology for cerebellar hemorrhage in the three cases presented was examined, including the role of CSF suction during surgery and disturbance of venous circulation in the posterior fossa. Suction of the CSF may cause intracranial hypotension. Further reduction of intracranial pressure leads to an increased transluminal venous pressure. There was no episode of hypertension or disturbed blood coagulation during or after the operation. The preoperative angiogram also revealed no abnormality at the region of the posterior fossa. Neuroimaging of infratentorial hemorrhage after supratentorial craniotomy is obviously different from that of hypertensive cerebellar hemorrhage. From the shape or extension of the hemorrhage, the main vessels of hemorrhage are the superior vermian vein and their tributaries damaged by stretching and tearing of these vessels. These vessels are not demonstrable in the angiogram, therefore there is no evidence for this hypothesis and the etiology is still unclear. There is no doubt, however, that there was a disturbance of venous circulation in this complication. We would like to emphasize the possibility of this complication. Patients who show signs of difficulty in awaking from anesthesia or the development of new neurological deficits not attributed to direct operative procedure after supratentorial craniotomy must be evaluated early, with adequate imaging including the posterior fossa. PMID- 10535082 TI - [A case of neurofibromatosis type 1 associated with arteriovenous fistula caused by re-bleeding of a vertebral dissecting aneurysm]. AB - We described a dissecting aneurysm of the vertebral artery (VA), which was associated with neurofibromatosis type 1 (NF1). A 41-year-old man was referred to our hospital because of abrupt, severe headache. A CT scan revealed diffuse subarachnoid hemorrhage (SAH) predominantly in the prepontine cistern. The angiograms showed a string sign in the left VA, just distal to the posterior inferior cerebellar artery (PICA). The vertebral dissection was considered responsible for SAH, and endovascular occlusion of the left VA was attempted. During the intervention, the patient complained of severe neck pain at the time of selective vertebral angiography, which revealed an arteriovenous fistula. The VA was occluded proximal to the PICA with GDC, which covered the fistula. Open surgery confirmed the two unruptured aneurysms. Intracranial dissection is rarely reported in association with NF1. However, ateriovenous fistula is not an uncommon combination with dissecting aneurysm and the extracranial segment of the VA is a characteristic target. Anatomical feasibility is conceivably the pathogenesis. PMID- 10535083 TI - [Persistent primitive olfactory artery: report of 5 cases]. AB - Five cases of a persistent primitive olfactory artery are reported. The anomalous artery coursed anteromedially along the ipsilateral olfactory nerve and made a hairpin turn, supplying the circulation of the anterior cerebral artery. It was proposed that a primitive embryonic olfactory artery had remained as this artery. A cerebral aneurysm was located at the hairpin curve of the artery. Hemodynamic stress is considered to have an important role in aneurysm formation. Any aneurysm occurring at an artery should be considered for surgical treatment. PMID- 10535084 TI - [Endovascular surgery for untreated ruptured aneurysm with symptomatic vasospasm]. AB - It is difficult to treat ruptured aneurysms with symptomatic vasospasm. Although direct surgery for such cases is associated with poor outcomes, conservative therapy has the risk of both rerupture and infarction. In two cases of ruptured aneurysms with symptomatic vasospasm, we performed aneurysmal coil embolization with Guglielmi electrodetatchable coils (GDC). At the same time we performed percutaneous transluminal angioplasty (PTA) with papaverine infusion. In both cases, rerupture did not occur and PTA was effective angiographically. A good outcome was achieved in case 1. However, broad cerebral infarction occurred in case 2, in which the patient had shown severe symptomatic vasospasm on admission. In advanced cases, such as in case 2, the outcome is poor. The aneurysm may not be able to be approached before PTA because of severe vasospasm. In such cases, PTA must be performed carefully to avoid aneurysmal rerupture. Intraarterial papaverine infusion is safer than PTA for severe spasm in distal vessels. However the efficacy of papaverine is known to be transient in many cases. It is often difficult to determine the exact relationship between branches and the aneurysm in the presence of vasospasm. In such cases, we recommend that the rupture point be packed and that the aneurysmal neck remain unpacked. After vasospasm is cured and good general condition has been recovered, direct surgery can be performed. In summary, endovascular surgery is an effective option for treatment of ruptured aneurysm with symptomatic vasospasm. PMID- 10535085 TI - [A case of chordoid meningioma]. AB - Chordoid meningioma is a very rare variant, especially in adults. We report an adult case of chordoid meningioma. A 52-year-old man was admitted to our hospital due to right hemiparesis. MRI revealed a left temporal convexity mass that showed two structures; the cerebral layer was shown as an isointensity area on T1WI and T2WI, and the dural layer seemed relatively hypointense on T1WI and hyperintense on T2WI. The tumor was well enhanced with gadolinium, especially on the dural side. A left external carotid arteriogram showed tumor stains with feeding vessels from the left middle meningeal artery. The tumor was totally removed via fronto-temporal craniotomy. Histological examination of the surgical specimen revealed two different structural components, meningothelial meningioma on the cerebral side, and chordoid meningioma on the dural side, consisting of clustering spindled cells and partly vacuolated ones in the mucoid stroma. In immunohistochemical examination, tumor cells showed positive staining for vimentin and epithelial membrane antigen (EMA), and negative for cytokeratin. From the above findings, this case was diagnosed as chordoid meningioma in an adult, a very rare variant of meningioma. PMID- 10535086 TI - [An etymological consideration on the nomenclatures of academic meetings and academic societies]. PMID- 10535087 TI - [Is vocational integration in the competitive labor market a realistic goal for the chronically mentally ill?]. AB - A wide range of sheltered jobs has been created in the second or special labour market with the aim of enabling the chronically mentally ill to participate in working life. However, having a job in the special labour market often precludes the chance of obtain-ing employment in the competitive labour market. To date, vocational integration programs enable only a small number of persons with psychiatric disabilities to attain reintegration into competitive employment. The predictors of successful vocational integration are subsequently discussed. A substantial increase in both sheltered and competitive jobs on the common labour market could be achieved for mentally ill and disabled people by adapting the "supported employment" model, as widely practised in the USA, to European labour market standards and appropriately funding its implementation. The utilisation of this model could serve to reduce the necessity of further expansion in the special labour market. PMID- 10535088 TI - [Effective use of time in community psychiatry--utopia and the reality of network management. Semmler W: Psychiat. Prax. 26 (1999) 67-70]. PMID- 10535089 TI - [Predictors of success in supported employment programmes--results of a prospective study]. AB - BACKGROUND: Vocational integration of mentally ill patients is confronted not only with illness-related restrictions but also with the current situation on the labour market and prejudices on the part of potential employers. The Consultancy Centres for Vocational Integration work with a supported employment approach which was adapted to the German situation. It includes prevocational training, assistance in finding appropriate jobs, cooperation with employing companies and long-term job site support. METHODS AND PATIENTS: Within the scope of a major study on vocational rehabilitation in the German region of Westphalia-Lippe, we carried out a prospective study of the further vocational course of 61 patients who had already been placed in competitive employment within the framework of the Consultancy Centres. These patients (30 men, 31 women) had a mean age of 31 years (+/- 6.9); 54% of them were suffering from schizophrenic disorders; mean duration of illness was 8.2 years (+/- 6.8). RESULTS: After two years (and for a subsample of 33 patients also after 3 years) two-thirds of patients were still in competitive employment. Predictors of success proved to be 1) a higher ability to cope with vocational stress on introduction of the measure, 2) an earlier start to rehabilitation, and 3) financial assistance for the company. A close correlation was recorded between course of illness (rehospitalizations) and success of rehabilitation. CONCLUSIONS: Even in the current situation of high unemployment rates vocational integration of mentally ill patients is possible and stable in medium term. Especially highly-motivated patients with favourable preconditions (early start of rehabilitation measures, higher ability to cope with work stress) are successful if intensive long-term support--including efforts involving employers--is provided. PMID- 10535090 TI - [The connection between labor integration and prognosis of schizophrenic patients: an 8-year follow-up study]. AB - This study reflects the labour-situation of 67 DSM-III schizophrenic outpatients in an 8-year follow-up. About 50% of the patients had less than 20 hours of work a week on the average--most of them in work therapy or sheltered work-places. Many of them changed their location of work. Patients with an initially good prognosis (MPS) had a good outcome, regardless if they had a normal job and worked about 30 hours a week or worked about 20 hours in a sheltered place. For patients with a poor prognosis even the integration in a sheltered working place was difficult. PMID- 10535091 TI - [Twelve conditions of successful vocational rehabilitation]. AB - This paper presents the hypothesis that the mentally ill (and impaired) can be successfully and permanently restored to the general job market, if a given region provides specific functions and institutions and develops styles and structures of interaction that take into account the specific needs of persons with psychosis. Twelve hypotheses are presented as to which factors enhance the chances of successful and long-term job-integration. The concept presented here appears to draw its advantage over others from its strictly regional as well as its interactionally oriented approach. PMID- 10535092 TI - [Comparison of need for care of sheltered home residents and patients in an ambulant psychiatric care unit]. AB - OBJECTIVE: Are sheltered home residents and patients of an ambulant care unit distinguishable by measuring their need for care? METHOD: N = 45 patients (30 patients of an ambulant care unit, among them 15 former residents of a sheltered home, and a parallelized sample of currently sheltered home residents) were examined with the "Integrierter Behandlungs- und Rehabilitationsplan" of the "Aktion Psychisch Kranke". Additionally, we assessed sociodemographic and illness related data and the reasons actually speaking against ambulant care. RESULTS: In average need for care was estimated significantly higher for the residents of the sheltered home than for the patients of the ambulant care unit (F = 6.383; p = 0.004). However, 6 sheltered home residents could be identified, who had less need for care, but were not motivated to get discharged. CONCLUSION: In the mean sheltered home residents and patients of an ambulant care unit are distinguishable by their need for care. However, other important factors for an ambulant care are the discharge motivation and the patients' ability to live alone respectively the existence of his or her social net. PMID- 10535093 TI - [Standard treatment of schizophrenia and legality of personnel equipment in psychiatric hospitals (German Psych-PV): possibilities and limitations]. AB - BACKGROUND: Comprehensive insight regarding better treatment and social reintegration of patients with schizophrenia has been gained over the past decade. Implementation of this knowledge into everyday's practice should be a major aim prior to the research on new variants of rehabilitative measures. To which extent this takes place has been poorly discussed until now. METHOD: Our comprehensive treatment program includes psychoeducation for patients and relatives, cognitive training, social skills training, additional psychoeducation for patients with dual diagnosis and a variety of a traditional group therapies like work therapy. An intensive cooperation is practised with complementary psycho-social services. In n = 89 consecutive admissions lasting at least 2 weeks we examined which patients were reached by these treatment offers. RESULTS: 84% of the patients participated in group therapies, where they passed 6.9 hours weekly on average. Most patients were reached by occupation therapy (62%), followed by physical therapy (54%), unspecific conversation group (54%), psychoeducation (35%), cognitive training (30%), work therapy (26%) and social skills training (7%). The PANSS Positive Scale at admission was negatively correlated with participation in group therapies. DISCUSSION: The implementation of a variety of psychotherapeutic offers for psychotic patients adequate to the state of the art can be achieved under conditions of standard hospital care in Germany (Psych-PV-law). Required organizational measures therefore are programs offered for patients of several wards in union and participation of employees in education. However, even by such efforts a considerable part of patients is not reached. Strategies to improve treatment results should consider these problems. PMID- 10535094 TI - [The police in psychiatric daily routine]. AB - PURPOSE: The following is a description of those patients brought by the police to the emergency room of a general hospital of Berlin for a psychiatric exploration. METHOD: The medical records of those patients brought in to the emergency room during one year were analyzed retrospectively. RESULTS: Within one year, 317 (10.8%) out of 2903 patients were brought by the police to have an emergency psychiatric exploration. The ratio of men to women was 2:1, and the median age was 41. Most illnesses diagnosed were schizophrenia, disorders caused by psychotropic substances, alcoholism, alcohol intoxication, and adjustment disorders/stress disorders. One third of the 317 patients were under influence of alcohol. 21.8% of the 317 patients were suicidal. Over two thirds of the patients were admitted to the psychiatric ward. Only 31 patients were not admitted to the ward for lack of indication. The most frequent reason for intervention of police was suicidal behaviour (35%), "strange behaviour" (28.7%) or aggressive behaviour (23%). CONCLUSION: The results show that those patients brought in by the police are in acute crisis situations often involving suicidal tendencies and suffer from more severe psychiatric illnesses. The psychiatric emergency exploration initiated by the police was generally justifiable. PMID- 10535095 TI - [Unimaginable violence in psychiatry--some reflections after a visit to Poland]. AB - What was the situation of the staff, when the Germans began, in the autumn of 1939, to murder systematically the patients in Polish clinics? A documentation of a fact-finding committee and a novel by Lem are studied for answers to this question. The patients themselves tried to resist. Whereas the staff are described as having been in the hopeless position of being unable to take any action. However, in very special situations the possibility of resistance was found. PMID- 10535097 TI - [Dysmorphophobia: between neurosis and psychosis]. PMID- 10535096 TI - [Acute paranoid hallucinatory psychosis following mefloquine prophylaxis (Lariam)]. AB - Mefloquine is a drug of choice for malaria prophylaxis in Africa because of the spread of chloroquine resistant plasmodium falciparum. On the other hand there are some reports about severe neuropsychiatric side effects associated with the intake of mefloquine medication. In our paper we present a case-report of a patient suffering for the first time from an acute paranoid psychosis induced by mefloquine prophylaxis. PMID- 10535098 TI - [Maniform psychosis in neurolues]. PMID- 10535099 TI - Understanding mechanisms in a cooperative protein: the CO ligation intermediates of hemoglobin. AB - Hemoglobin is a regulatory component of the oxygen transport to the tissues, and for decades has been a prototype to develop new strategies for the study of the structure/function relationships in proteins. One of the most difficult, and so far, unattained objectives of hemoglobin research has been the study of the hemoglobin molecules in a state of partial ligation with oxygen, or intermediates, as a means of testing theories of cooperativity. A cryogenic technique has been developed for the isolation, identification and quantification of the reaction intermediates of hemoglobin and CO, which in many aspects is a close approximation to the physiological ligand. The technical features that are crucial for the evaluation of the significance of the experimental data obtained using this technique and various approaches to the analysis of the data are reported. The discussion points out the importance of accessing direct information on the nature and concentrations of the intermediates in solution to clarify mechanisms of cooperativity as opposed to the less informative studies of the bulk properties of the solution. PMID- 10535100 TI - Interaction of a protein, BSA, and a fluorescent probe, Mag-Indo-1, influence of EDTA and calcium on the equilibrium. AB - Recent findings indicate that ion-chelator probes with tetracarboxylate structure bind proteins. It was suggested that these fluorescent probes are valuable tools to gain information on protein structure through the energy transfer from tryptophans to the bound probe. Here, the binding of the fluorescent probe Mag Indo-1 to bovine serum albumin (BSA) was investigated. Mag-Indo-1 was reported previously to serve as a probe for magnesium cations (Kd = 2.8 x 10(-4) M for zero ionic strength) which can also interact with calcium cations (Kd = 7.5 x 10( 7) M). Probe complexation with protein results in a shift of the emission fluorescence spectrum of the probe from 480 to 457 nm. We used emission fluorescence techniques to monitor this interaction. Computational resolution of the complex fluorescence spectra and a new software to test the theoretical model were developed in our laboratory. This enabled us to calculate the number of interacting sites and the dissociation constants. The fluorescent probe Mag-Indo 1 binds at a singular site with high affinity (Kd = 1.8 x 10(-7) M) to bovine serum albumin (BSA). Since proteins are known to bind several compounds unspecifically, we have studied the influence of EDTA as a competitor of the probe. Our findings suggest that the BSA binding site is identical for both Mag Indo-1 and EDTA. We found that EDTA binds the protein with Kd = 0.4 x 10(-3) M. We studied the influence of calcium and found that Mag-Indo-1 does not bind the calcium free Apo-protein anymore. PMID- 10535101 TI - Comparison of the thermal denaturation behaviour of DNA-solutions and metaphase chromosome preparations in suspension. AB - Hyperchromicity measurements are well established to analyse the thermal denaturation behaviour of pure DNA sequences in solution. Here, we show that under appropriate experimental conditions this technique can also be applied to study thermally controlled conformation changes of higher order DNA-protein complexes as for instance metaphase chromosome preparations in suspension. A computer controlled sensitive, upright double beam photometer with a heatable cuvette was constructed. Measurements of the temperature dependent extinction of both, solutions and particle suspensions are possible, since sedimentation effects of particles can be neglected due to the vertical optical axis in the probe cuvette. Thermal denaturation of metaphase chromosome preparations of human and Chinese hamster cells was investigated and compared to melting profiles of DNA solutions for two excitation wavelengths, 256 and 313 nm. The influence of neutral and low pH was considered. The results indicate that metaphase chromosome preparations show a thermal denaturation behaviour different from pure DNA. Whereas DNA solutions showed one pH dependent melting peak at 256 nm only, the peak pattern of metaphase chromosome preparations showed a large variability both at 256 and 313 nm. At neutral pH, in two temperature regions (40-55 degrees C and 75-82 degrees C) peaks were found indicating chromosome typical conformation changes independently from the mammalian cell species (Chinese hamster, human). In contrast to pure DNA, no typical reduction in the temperatures of peak maxima with decreasing pH was found for metaphase chromosome preparations of both cell types. These results may be relevant for further systematic studies of efficient thermal probe/target denaturation procedures in non enzymatic DNA-chromosome in situ hybridisation. PMID- 10535102 TI - Changes of SOD-like substances in mouse organs after low-dose X-ray irradiation. AB - We demonstrated that low-dose irradiation with 50 cGy of X-ray induces in vivo production of superoxide dismutase (SOD)-like substances and accelerates antioxidant activity. To elucidate the defense mechanism against X-ray radiation, we examined which components among these SOD-like substances, such as SOD, vitamin C and celuroplasmin, are produced by low-dose irradiation. Our study revealed that SOD-like specific activity hardly involved SOD-like substances other than SOD. Moreover, it is suggested that low-dose irradiation induced synthesis and production of SOD itself, leading to elevation of SOD-specific activity. PMID- 10535103 TI - Arginine acts as a protective and reversal agent against glycolytic inhibitors in spermatozoa. AB - It is known that the amino acid arginine stimulates sperm motility and glycolytic activity. We have earlier studied its efficacy as a stimulator of glycolysis in goat spermatozoa under anaerobic conditions. Here, we have assessed the influence of arginine in reversing the impairment caused by glycolytic inhibitors, iodoacetamide and iodoacetic acid. Glycolysis has been monitored by measuring the consumption of 13C labeled glucose and the amount of 13C labeled lactate produced under different experimental conditions, using 13C NMR. It is observed that both L- and D-arginine are able to prevent and reverse the inhibitory action of glycolytic inhibitors. The reversal effect of arginine gives rise to about eight times higher metabolic activity as compared to the inhibited cells while structurally related amino acids such as nitro-arginine, homo-arginine, lysine and ornithine are ineffective. The energetics of spermatozoa as measured by 31P NMR show a reduction in ATP level in cells incubated with iodoacetamide. Treatment of these cells with both L- and D-arginine restores the ATP level. The results may have significance in the treatment of male infertility. PMID- 10535104 TI - Ridges and rivers: a test of competing hypotheses of Amazonian diversification using a dart-poison frog (Epipedobates femoralis). AB - Mitochondrial DNA cytochrome b sequence data from a dart-poison frog, Epipedobates femoralis, were used to test two hypotheses of Amazonian diversification: the riverine barrier and the ridge hypotheses. Samples were derived from sites located on both banks of the Rio Jurua and on both sides of the Iquitos Arch in western Amazonia. The phylogeographic structure was inconsistent with predictions of the riverine barrier hypothesis. Haplotypes from opposite river banks did not form monophyletic clades in any of our phylogenetic analyses, nor was the topology within major clades consistent with the riverine hypothesis. Further, the greatest differentiation between paired sites on opposite banks was not at the river mouth where the strongest barrier to gene flow was predicted to occur. The results instead were consistent with the hypothesis that ancient ridges (arches), no longer evident on the landscape, have shaped the phylogeographic relationships of Amazonian taxa. Two robustly supported clades map onto opposite sides of the Iquitos Arch. The mean haplotypic divergence between the two clades, in excess of 12%, suggests that this cladogenic event dates to between five and 15 million years ago. These estimates span a period of major orogenesis in western South America and presumably the formation of these ancient ridges. PMID- 10535105 TI - X chromosome DNA variation in Drosophila virilis. AB - Comparisons of polymorphism patterns between distantly related species are essential in order to determine their generality. However, most work on the genus Drosophila has been done only with species of the subgenus Sophophora. In the present work, we have sequenced one intron and surrounding coding sequences of 6 X-linked genes (chorion protein s36, elav, fused, runt, suppressor of sable and zeste) from 21 strains of wild-type Drosophila virilis (subgenus Drosophila). From these data, we have estimated the average level of DNA polymorphism, inferred the effective population size and population structure of this species, and compared the results with those obtained for other Drosophila species. There is no reduction in variation at two loci close to the centromeric heterochromatin, in contrast to Drosophila melanogaster. PMID- 10535106 TI - Facial attractiveness, symmetry and cues of good genes. AB - Cues of phenotypic condition should be among those used by women in their choice of mates. One marker of better phenotypic condition is thought to be symmetrical bilateral body and facial features. However, it is not clear whether women use symmetry as the primary cue in assessing the phenotypic quality of potential mates or whether symmetry is correlated with other facial markers affecting physical attractiveness. Using photographs of men's faces, for which facial symmetry had been measured, we found a relationship between women's attractiveness ratings of these faces and symmetry, but the subjects could not rate facial symmetry accurately. Moreover, the relationship between facial attractiveness and symmetry was still observed, even when symmetry cues were removed by presenting only the left or right half of faces. These results suggest that attractive features other than symmetry can be used to assess phenotypic condition. We identified one such cue, facial masculinity (cheek-bone prominence and a relatively longer lower face), which was related to both symmetry and full- and half-face attractiveness. PMID- 10535108 TI - Emissions reduction of high and low polluting new technology vehicles equipped with a CeO2 catalytic system AB - The efficiency of a catalytic system that provides the vehicle combustion chamber with an adequate amount of cerium oxide (CeO2), without affecting the operational parameters of the engine, has been studied for pollutant emissions reduction. New technology gasoline engine passenger cars, equipped with this catalytic system, have been driven on their normal route by the same drivers, in order to obtain the same traffic conditions and the same driving mode, as far as possible. These vehicles were tested for CO and HC emissions before and after the installation of the catalytic system. The new technology passenger cars examined have been divided into two categories using as criterion the level of their initial emissions (before the installation of the CeO2 catalytic system). The two categories are: (a) high polluting cars with CO and HC emissions above the implemented standards; and (b) low polluting cars with emissions below the standards. Both car categories equipped with the CeO2 system, present important reduction of the pollutant emissions. The results are interpreted statistically and they are correlated with other studies. PMID- 10535107 TI - The kinetics of proteinase K digestion of linear prion polymers. AB - Transmissible spongiform encephalopathies such as scrapie are caused by a protein only infectious agent, known as a prion. It is not clear how a protein can be capable of replicating itself, and the mechanism remains controversial. One influential model hypothesizes that prions are nucleated, macroscopically linear polymers. We investigated the theoretical kinetics of this model and derived predictions which could be used to test the model. In the model, the polymerization and depolymerization rates are independent polymer size. This leads to an exponential size distribution at equilibrium. In agreement with a prediction stemming from this size distribution, the average size of PrP-res polymers was proportional to the square root of the concentration of PrP-res in a published study of in vitro conversion. Prion digestion by proteinase K (PK) is predicted to be biphasic. The second phase of digestion should be virtually independent of the PK concentration and should depend on the initial size distribution of prion polymers. For initially equilibrated polymers with an exponential size distribution, phase two digestion is exponential at a predicted rate. This rate varies in a defined way with the concentration used for equilibration and with other parameters which affect the average polymer size. PMID- 10535109 TI - Driving cycles for measuring passenger car emissions on roads with traffic calming measures AB - Although local authorities in the UK need to be aware of any air quality impacts resulting from their traffic calming operations, there is little information relating to the effects of different traffic calming measures. The effects on air quality on this scale are complex, and so TRL is providing guidance by developing performance indices for different measures based on their effects on vehicle emissions. The emissions indices for passenger cars are based on tests conducted on a chassis dynamometer, and this paper describes the development of the methodology for constructing the driving cycles to be used. The technique involves the measurement of the speed profiles of a large number of vehicles using a roadside LIDAR system, and the determination of typical gear selections using three-instrumented cars. PMID- 10535110 TI - Trends in hydrocarbon fleet emissions at four UK highway sites. AB - The on-road hydrocarbon emissions of vehicles at four sites in the UK have been monitored using remote sensing equipment. The single lane highway sites in London (Haringey and Southwark), Middlesborough and Leicester were monitored between May 1994 and October 1995. Analysis of the results shows that there is both a large majority of low emitting vehicles which contribute little to fleet hydrocarbon emissions and a small minority of high emitting vehicles which contribute significant proportions to fleet hydrocarbon emissions at all sites. This also results in a skewing of the data set so that a pattern of high mean values and lower median values is consistently observed. Analysis of model year data suggests a low association between vehicle age and mean hydrocarbon emissions for vehicles produced prior to 1983 but the relationship improves after 1983 with regression analyses giving r2 values as high as 0.96. Relatively new high polluting vehicles are the greatest contributors to fleet emissions with, on average, 52% of hydrocarbon fleet emissions being produced by these vehicles from model years 1985-1991. Therefore, fleet emissions could be significantly reduced if new highly polluting vehicles were subject to regular emissions testing followed by appropriate remedial action or were removed from the highway by the withdrawal of their vehicle registration. Older vehicles play a minor role in fleet emissions with, on average, only 13% of hydrocarbon fleet emissions being produced by vehicles registered prior to 1983. PMID- 10535111 TI - Actual car fleet emissions estimated from urban air quality measurements and street pollution models. AB - A method to determine emissions from the actual car fleet under realistic driving conditions has been developed. The method is based on air quality measurements, traffic counts and inverse application of street air quality models. Many pollutants are of importance for assessing the adverse impact of the air pollution, e.g. NO2, CO, lead, VOCs and particulate matter. Aromatic VOCs are of special great concern due to their adverse health effects. Measurements of benzene, toluene and xylenes were carried out in central Copenhagen since 1994. Significant correlation was observed between VOCs and CO concentrations, indicating that the petrol engine vehicles are the major sources of VOC air pollution in central Copenhagen. Hourly mean concentrations of benezene were observed to reach values of up to 20 ppb, what is critically high according to the WHOs recommendations. Based on inverse model calculation of dispersion of pollutants in street canyons, an average emission factor of benzene for the fleet of petrol fuelled vehicles was estimated to be 0.38 g/km in 1994 and 0.11 in 1997. This decrease was caused by the reduction of benzene content in Danish petrol since summer 1995 and increasing percentage of cars equipped with three way catalysts. The emission factors for benzene for diesel-fuelled vehicles were low. PMID- 10535112 TI - Updated urban emission inventory with a high resolution in time and space for the city of Graz. AB - Emission inventories are a tool to monitor the development of the emission situation in urban areas. Therefore, periodic updates of the emission inventory are required. An update of an emission inventory is always combined with new or improved emission factors and models. In order to define the emission trend it is necessary to distinguish between methodological and real changes. This means using the most recent data (models and emission factors) and applying it to the new and old base year. Taking the emission inventory for the city of Graz as an example the effects of methodological changes on emission estimates are shown. Due to the fact that this emission inventory is also used for urban air quality modelling it has a high temporal and spatial resolution. Emphasis was therefore put on an accurate description of the time behaviour of the emission sources. The use of a geographical information system ensures an exact location of the emission source. The new emission inventory serves as a basis for an urban air quality model. The work done so far is part of the 'DATE-Graz project' where emission inventory dispersion models and measurement campaigns act together to provide a better understanding of urban air problems in the city of Graz. PMID- 10535113 TI - Estimation of spatially resolved road transport emissions for air quality management applications in the north west region of England AB - Spatially resolved estimates of combustion and non-combustion related emissions of CO, NOx, VOCs and PM from road transport sources have been made for the North West region of England in 1994. These have been generated using detailed emissions models for combustion related emissions of CO, NOx, VOCs and PM which take into account the different emissions profiles associated with particular vehicle groups, different road types and journeys under cold start conditions. Emissions estimates have been generated for a 1 x 1 km grid covering the region's urban and industrial zones and a 5 x 5 km grid for the whole study area. Emissions models have been generated and applied within a Geographical Information System (GIS) environment. Areas of uncertainty in the estimation procedure have been examined and the results compared with alternative data sources. Although the work centres upon the North West region of England, as far as possible the methods and data sources used are intended to be generic, particularly in respect to other administrative areas of the UK. In this way, this work can be considered to be of wider interest than at the local level alone. PMID- 10535114 TI - Reliability of the current models of instantaneous pollutant emissions AB - Pollutant emissions and fuel consumption of passenger cars are usually assessed as a function of average as a function of average speed. Nevertheless a number of attempts were made to take into account the driving pattern using emission vs. instantaneous speed and acceleration models. Five quite similar models were developed in Europe. To which extent these instantaneous models are innovative as compared to conventional models? In a first step the various development stages of an emission-based model and the various associated errors are presented. Among the possible sources of error, we selected modelling-induced errors, enabling to compare rigorously the various model types in terms of performance. The analysis was performed using the Modem model, developed from the measurement results obtained over a sample of 150 vehicles, representative of French, English and German fleets, over 14 representative driving cycles under urban conditions. Average emissions as measured over these 14 cycles are compared to emissions calculated over same cycles using this model. Modelling errors range from -51% to +57% as a function of the considered cycle and the vehicle type. This inaccuracy is quite similar for the other four European models. This demonstrates the low reliability of the models used, which cannot be used to assess the impact of slight changes in the driving pattern, sometimes leading to completely false conclusions. These models are barely more precise than average speed based models. In a second step, a number of alternatives liable to improve the reliability of instantaneous models are contemplated: for example using another method for calculating acceleration, or increasing the number of speed and acceleration classes significantly. But this does not improve significantly the model reliability. Limited measurements performed on a catalyst vehicle demonstrated that very high engine loads, even if not frequent, play a significant role in emissions: sometimes they yield emission values a thousand times higher. It is therefore of prime interest to study them in a comprehensive manner. In addition, this demonstrates that each model should be developed from measurements carried out over a set of representative driving cycles under real world driving conditions. PMID- 10535115 TI - Evaluation of pollutant loadings in the runoff waters from a major rural highway AB - The quality of pavement runoff water from a 275-m motorway section has been studied for 1 year, during which approximately 50 rain events have been sampled. Two different types of pollution have been revealed. One type can be defined as chronic and includes suspended solids, chemical oxygen demand, total hydrocarbons, zinc and lead. The second type can be considered to be seasonal and incorporates chlorides, sulfates, suspended solids and heavy metals due to the use of deicing salt in winter. Pollutant loading as regards lead appears lower than in previous studies because of the increasing number of vehicles using unleaded gasoline. The study conducted on the sources of pollution and on heavy metal fluxes (Pb, Cu, CD, Zn) released by the traffic has been used to assess a mass balance with respect to pollutant loadings removed by runoff waters. It seems that a large proportion of the lead concentration may disperse in the atmosphere, whereas cadmium sources may be ill-identified or underestimated. PMID- 10535116 TI - Highway runoff and potential for removal of heavy metals in an infiltration pond in Portugal AB - Highway runoff from IP 4, a mountain road in the north-east of Portugal, has been monitored using a system integrating a raingauge, a flowmeter and an automatic water sampler. Average daily traffic (ADT) is 6000 and the study catchment has 5970 m2 of total area and 2500 m2 of road pavement. A single stormwater outlet discharges into an infiltration pond with overflow to a creek. Sampling was carried out before the runoff water entered the pond. Among the parameters analysed in the runoff water, the heavy metals cadmium (Cd), chromium (Cr), copper (Cu), lead (Pb) and zinc (Zn) were emphasised because of their toxicity. Concentrations of Cd and Cr were usually lower than the detection limit (1 microgram/l). Copper levels found were between 1 and 54 micrograms/l; lead from 1 to 200 micrograms/l and zinc from 50 to 1460 micrograms/l. A first flush effect was observed, meaning that the first 50% of the runoff volume for each event typically transported between 61 and 69% of the total suspended solids, Zn, Cu and Pb loads. Runoff water is totally infiltrated into the pond and heavy metals are being sorbed to the soil. Soils used in infiltration ponds should have specific characteristics in order to perform effectively and ensure groundwater protection. Not only well-known soil texture and composition characteristics are relevant: the soil sorption capacity--the extension and reversibility of the processes--is of main importance in this kind of highway runoff treatment. Experiments concerning the sorption of Zn, Cu and Pb to soils, followed by desorption at pH values of 2, 4 and 6 were conducted in the laboratory. These experiments were performed with the soil existing at the highway IP 4 infiltration pond and with two other common types of Portuguese soils. The three types of soil showed different behaviours, which must be related to their characteristics; the soil pH seemed to play a significant role in controlling the Zn, Cu and Pb sorption processes. As expected, as lowering of the pH value increased the desorption rate. The infiltration pond soil is the one with the lowest sorption capacity, however, it showed a relatively high sorption strength which means that it is considered reliable, concerning highway runoff treatment and groundwater protection. PMID- 10535117 TI - Comparison of the heavy metal content of motorway stormwater following discharge into wet biofiltration and dry detention ponds along the London Orbital (M25) motorway AB - The Surrey section of the London Orbital M25 motorway uses mainly detention pond facilities for the treatment of stormwater runoff. A majority of these implement the use of dry detention basins. However, in a few locations biofiltration facilities operate through the use of reed bed systems. An assessment of the removal efficiencies for both wet biofiltration and dry pond treatment facilities was undertaken. Motorway-derived contaminants, including V, Cr, Mn, Co, Ni, Cu, Zn, Mo, Cd, Sb and Pb, were measured in unfiltered stormwater collected during the initial stages of a storm event using inductively coupled plasma mass spectrometry (ICP-MS). Results suggest that a higher level of motorway-derived heavy metal contamination exists in stormwater runoff from a road section with a higher average daily traffic density. In addition, a comparison of both sites shows a higher percentage removal efficiency of heavy metals in stormwater from the biofiltration facility. PMID- 10535118 TI - Adsorptive infiltration of metals in urban drainage--media characteristics AB - Urban pavement drainage often contains significant quantities of anthropogenic metal elements, including Cd, Cu, Pb and Zn that exceed surface water discharge standards. In many urban areas low rainfall pH, results in predominately dissolved metal element mass. Such partitioning has critical implications for the selection of in-situ treatment. One such category of treatment is engineered infiltration systems. To be effective, such systems must adsorb dissolved metal elements to their fixed media while also acting as filters for particulate-bound fractions. One such strategy is called a partial exfiltration trench (PET). The PET contains oxide-coated sand (OCS); an amphoteric media of high surface area (5 15 m2/g) as compared to uncoated silica sand (0.01-0.05 m2/g). OCS was generated through heating a mixture of silica sand and ferric nitrate solution to dryness. This paper presents results of both media characterization and bench scale PET simulations. Media tested were OCS and plain silica sand. Media testing was carried out until capacity was exhausted, using both synthetic and actual stormwater loadings. Testing was conducted for pH levels of 6.5 and 8.0. Results indicated that OCS had greater capacity than silica sand for all dissolved fractions. As the pH was raised from 6.5 to 8.0, OCS capacity was improved. A PET configuration with porous pavement resulted in the highest in-situ treatment capacity for metal element bearing storm water. PMID- 10535119 TI - The design of vegetative constructed wetlands for the treatment of highway runoff. AB - The Environment Agency for England and Wales are responsible for assessing the effects of highway runoff and for monitoring the treatment systems/procedures which have been introduced for the reduction of deleterious effects. The Agency is looking into the improvement of surface water management in terms of best management practices and plans to work in partnership with the Highways Agency to achieve this aim. Among the treatment options being considered are constructed wetlands. Draft Guidelines have been developed to provide information on their design. This paper describes procedures for carrying out an Environmental Sensitivity Analysis to determine whether treatment by a constructed wetland is appropriate. Information on water quality and quantity is required as well as the sensitivity of the receiving environment. The legislative position, particularly in relation to the discharge quality of the water and the conservation status of the receiving environment, needs also to be considered. The factors that will determine the most appropriate wetland design criteria include traffic loadings, road drainage area, land availability, cost and the size/extent and type of the receiving water body. The following structures are recommended for incorporation in the overall design; oil separator and silt trap, spillage containment, settlement pond, vegetative wetland and final settlement tank. The operation and maintenance procedures and the monitoring requirements for a functioning wetland are described. PMID- 10535120 TI - Measuring and modelling the airborne particulate matter mass concentration field in the street environment: model overview and evaluation. AB - This paper discusses the outline structure and preliminary evaluation of an emission-dispersion model for predicting the temporal and spatial distribution of vehicle-derived airborne particulate matter mass concentration in street canyons. The model is called Street Level Air Quality (SLAQ). SLAQ is semi-empirical, in that it uses not only results from field and wind tunnel experiments but also theory and models derived from multiple runs of numerical routines in order to simulate the basic physical processes within the street canyon. A combination of a plume model, for the direct contribution of vehicle exhaust, and a box model for the recirculating part of the pollutants in the street, is used to predict concentration for receptors within the canyon. Emission rates of vehicle-derived particulate matter are calculated within SLAQ, which serve as input to the dispersion module. Exhaust emission rates are scaled element by element along the street for each of the lanes according to the direction of traffic flow to account for modal operation of vehicles near signalised intersections. This refinement allows SLAQ to account for non-uniformity in along-canyon emission rates and to model a street that has several intersections along its length. Thermal turbulence due to environmental surface sensible heat and vehicle generated heat is accounted for in the model. Other features of SLAQ include correction for the urban heat island effect, dry deposition, wet deposition, particle settling and estimation of wind direction standard deviation, when this latter data is not available. SLAQ has been evaluated in a street in Loughborough, Leicestershire, United Kingdom and correlation coefficient of 0.8 between the modelled and measured concentrations has been obtained. PMID- 10535121 TI - A street deposit sampling method for metal and hydrocarbon contamination assessment. AB - Urban surface contamination, by atmospheric deposits as well as human activities, is a major concern for urban pollution management. Besides coarse street deposits which are clearly perceived and easily removed, suspended solid (SS) surface loads and contamination by heavy metals and hydrocarbons are rarely assessed although they could be of major importance with regards to combined or separate server overflow (CSO and SSO) impacts. Both dry and wet vacuum sampling procedures have been first compared, in the laboratory, using dry and sieved clay or street deposits. Then the wet vacuum sampling procedure has been refined, coupling the injection of water and the hand-brushing of the surface prior to its vacuum cleaning, and evaluated on a car parking area close to the University. Finally this procedure has been assessed in Bearn Street within the 'Le Marais' district in Paris centre, and 34 samples have been analysed for metal and eight for aromatic hydrocarbon contamination. Heavy metal concentrations (0.1-1.7 g kg 1 dry wt. Cu, 0.9-6.1 g kg-1 dry wt. Pb and 1.5-4.6 g kg-1 dry wt. Zn) within street deposit samples collected in Paris centre, indicate a high contamination, especially for copper and zinc, as compared to reported data. Total polyaromatic hydrocarbons (PAHs) are in the 3-11 mg kg-1 dry wt. range, thus approximately 10 times less contaminated than dry atmospheric deposits. This paper presents data obtained and discusses the difficulties encountered when sampling street deposits in busy areas of a city like Paris. The water jet street cleaning procedure used by Paris city workers was tested for its efficiency, by comparison of surface loads before and after the cleaning procedure. Although solids cleaning efficiency is highly variable (20-65%) and somewhat higher for particles larger than 100 microns, particulate metal cleaning efficiency is even more variable (0 75%) and particulate PAHs appear not to be significantly removed. PMID- 10535122 TI - Measured and modelled concentrations and vertical profiles of airborne particulate matter within the boundary layer of a street canyon. AB - Concentrations and vertical profiles of various fractions of airborne particulate matter (suspended particulate matter (SPM), PM10 and PM2.5) have been measured over the first three metres from ground in a street canyon. Measurements were carried out using automated near real-time apparatus called the Kinetic Sequential Sampling (KSS) system. KSS system is essentially an electronically controlled lift carrying a real-time particle monitor for sampling air sequentially, at different heights within the breathing zone, which includes all heights within the surface layer of a street canyon at which people may breathe. Data is automatically logged at the different receptor levels, for the determination of the average vertical concentration profile of airborne particulate matter. For measuring the airborne particle concentration, a Grimm Dust Monitor 1.104/5 was used. The recorded data also allows for time series analysis of airborne particulate matter concentration at different heights. Time series data and hourly-average vertical concentration profiles in the boundary layer of the confines of a street are thought to be mainly determined by traffic emissions and traffic associated processes. Hence the measured data were compared with results of a street canyon emission-dispersion model in time and space. This Street Level Air Quality (SLAQ) model employs the plume-box technique and includes modules for simulating vehicle-generated effects such as thermally- and mechanically-generated turbulence and resuspension of road dust. Environmental processes, such as turbulence resulting from surface sensible heat and the formation of sulphate aerosol from sulphur dioxide exhaust emissions, are taken into account. The paper presents an outline description of the measuring technique and model used, and a comparison of the measured and modelled data. PMID- 10535123 TI - Benzene in blood as a biomarker of low level occupational exposure. AB - The occupational airborne exposure to benzene of 150 workers employed in petrol stations and a refinery plant was assessed using personal sampling pumps. All workers provided blood samples after the end of work and on the following morning before resuming work. Benzene concentrations in the blood of 243 non occupationally-exposed subjects were also measured. The median occupational benzene exposure for all 150 workers studied was 80 micrograms/m3. Overall median blood benzene of all workers was 251 ng/l at the end of the shift, and 174 ng/l the following morning. The benzene concentrations measured in blood collected the following morning proved to be significantly lower than those measured at the end of the shift. Median blood benzene for the 243 'normal' subjects was 128 ng/l, which was significantly lower than that measured in the workers before a new work shift. The median blood benzene concentration was significantly higher in smokers than in non-smokers, both in the general population (210 ng/l vs. 110 ng/l) and in the exposed workers at the end of the shift (476 ng/l vs. 132 ng/l) and the following morning (360 ng/l vs. 99 ng/l). End-of-shift blood benzene correlated significantly with environmental exposure; this correlation was better in the 83 non-smokers than in the 67 smokers. In non-smokers with the median benzene occupational exposure of 50 micrograms/m3, no difference was found in blood benzene concentration in exposed and non-exposed subjects. PMID- 10535124 TI - Personal exposures to airborne metals in London taxi drivers and office workers in 1995 and 1996. AB - In 1995, a petroleum marketer introduced a diesel fuel additive in the UK containing Mn as MMT (methylcyclopentadienyl manganese tricarbonyl). A small study of personal exposures to airborne Mn in London was conducted before and after introduction of the additive to identify any major impact of the additive on exposures. In 1995, personal exposures to Mn were measured in two groups, taxi drivers and office workers (10 subjects per group) for two consecutive 7-day periods. A similar study was carried out in 1996 to determine if exposures had changed. Samples were also analyzed for Ca, Al, Mg and Pb. In 1996, exposures to aerosol mass as total suspended particulates (TSP) and PM2.5 were measured in addition to the metals. Manganese exposures in this cohort did not increase as a result of introduction of the additive. However, a significant source of Mn exposure was discovered during the conduct of these tests. The mean exposure to Mn was higher among the office workers in both years than that of the taxi drivers. This was due to the fact that approximately half of the office workers commuted via the underground railway system where airborne dust and metal concentrations are significantly elevated over those in the general environment. Similar results have been noted in other cities having underground rail systems. Exposure to Mn, Pb, Ca, and Mg were not significantly different between the 2 years. Taxi drivers had higher exposures than office workers to Mg and Pb in both years. Commuting via the underground also had a significant impact on exposures to TSP, PM2.5, Al, and Ca, but had little effect on exposures to Mg. The aerosol in the underground was particularly enriched in Mn, approximately 10-fold, when compared to the aerosol in the general environment. There are several possible sources for this Mn, including mechanical wear of the steel wheels on the steel rais, vaporization of metal from sparking of the third rail, or brake wear. PMID- 10535125 TI - Platinum uptake by the freshwater isopod Asellus aquaticus in urban rivers. AB - Platinum has been increasing in the environment as a result of emissions from catalytic converters. The platinum emitted is principally located in the vicinity of roads but might be transported to urban rivers through highway and urban run off water. Platinum concentrations in the freshwater isopod Asellus aquaticus were measured for two urban rivers and a stormwater detention pond. Concentrations ranged from 0.04 to 12.4 micrograms g-1 for direct analysis and from 0.16 to 4.5 micrograms g-1 after depuration. Analyses of water, pore water and sediments indicate that platinum in urban rivers is mostly found in the sediments and these provide the major contribution of platinum to Asellus aquaticus. Exposure experiments showed the importance of platinum speciation for uptake. PMID- 10535126 TI - Social awareness of air quality information. AB - In the UK, air quality information is made available through a range of media. However, limited attention has been paid to ensuring that the information is provided to the public in a format that is understandable and relevant to their needs. This research has begun the task of determining the nature and extent of public air quality requirements by performing a social survey (using a postal questionnaire) to provide a basic snapshot of the public's views and by determining the views of information providers and interested professionals. The paper identifies the main shortcomings in current public air quality information provision. The social survey and workshop results demonstrate that current information provision and dissemination does not match public requirements; the depth and breadth of local information needs to be enhanced. Local authorities need to improve their co-ordination and collaboration, the role of the mass-media needs to be considered carefully, air quality needs to be better described and its implications for individuals spelled out and certain public groups need special consideration. In addition, local authorities need more guidance on communicating air quality, possibly through a best practice guide. Further research is required to identify the best descriptors for air quality, to improve the effectiveness of public advice during episodes of poor air quality and to use public air quality information to effect behavioral changes. PMID- 10535128 TI - Seasonal analysis of air pollution levels in Madrid. AB - Madrid city has a high density of population and suffers from chronic congestion problems. It means that some pollutants could produce atmospheric emergency situations when weather stability periods last longer. Due to the low level of industry in the region, mobile sources have an important contribution to total emissions. Madrid has a 20-year-old air pollution control network which is composed of 24 permanent stations which control all pollutants and atmospheric variables. This paper analyses inmission values from 1990 to 1997. The analysis covers main pollutant values and their variations within the week and between seasons. The study has a twofold approach: mean-daily values and semi-hourly values. The results allow to draw some conclusions about inmission values in different areas of the city and how traffic-flow contributes to them. PMID- 10535127 TI - Traffic related air pollution in city districts near motorways. AB - Traffic related air pollution near major motorways was measured and results are presented. PMID- 10535129 TI - Dispersion of PAH and heavy metals along motorways in The Netherlands--an overview AB - In the past years the Road and Hydraulic Engineering Division (DWW) of the Dutch Directorate-General of Public Works and Water Management has supervised research after the pollution of the vicinity of motorways in the Netherlands with micropollutants such as heavy metals and PAH. The results of these and additional studies have been summarized and further analyzed, so that we would be able to design optimal measures and devices to reduce pollution. PMID- 10535130 TI - Urban air quality management: the traditional vs. an exposure-based approach. AB - This paper presents thoughts about a different form of air quality management that focuses on long-term personal exposure of individuals and goes beyond the traditional system of air quality standards. The exposure-based approach is a customer-oriented methodology with the aim of individually lowering the personal intake of chronically acting air pollutants and hence improve human health conditions more directly. PMID- 10535131 TI - Estimation of trends in urban traffic NOx emissions by an empirical model AB - A model has been developed to estimate trends in urban traffic NOx emissions by measured NO2 concentrations. PMID- 10535132 TI - ISIS-Traffic, a monitoring model for car induced air pollution in build-up areas AB - ISIS-Traffic is an easy-to-use model for simulations of the air pollution caused by cars in urban districts. With the model even large networks of streets can be evaluated in relation to their pollution load. PMID- 10535133 TI - Local air quality management: a practical approach to air quality assessment and emissions audit. AB - This paper reviews the results of the practical application of national UK tools of air quality management on a local scale, to show their usefulness and application. Such tools include an urban emissions inventory, dispersion modelling and the review and assessment guidance procedures of the National Air Quality Strategy. PMID- 10535134 TI - Air quality index and its use in Italy's management plans. AB - The paper defines a specific Air Quality Index (AQI) introduced to evaluate the general trend of air quality within the elaboration of air quality management plans and discusses its application in the Trento (Italy) plan. PMID- 10535135 TI - Health effects associated with Madrid air pollution levels. AB - The purpose of this study was to determine the relationship between the pollution levels recorded in Madrid and the number of hospital admissions made on the grounds of respiratory disorders. PMID- 10535137 TI - Air quality legislation and controls--imperfectly conceived solutions to imperfectly understood problems? AB - The development of legislative controls for the improvement of air quality is reviewed. In particular, the controls relating to motor vehicle emissions are discussed, and the effects of moving from direct measures relating to vehicle construction and manufacture, to indirect controls on vehicle operation are considered. PMID- 10535136 TI - Monitoring children's personal exposure to airborne particulate matter in London, UK--method development and study design. AB - The paper outlines the methodology selected for identifying the personal exposure of children to airborne particulate matter in a UK urban environment. PMID- 10535139 TI - Steps towards an environmentally sustainable transport system AB - Motorists may assist in going towards sustainable transport if given a wider basis for choice between vehicles, to include a number of important environmental aspects. PMID- 10535138 TI - Commuter exposure to respirable particles inside buses and by bicycle. AB - The exposure of bus commuters and a cyclist to respirable particles in the city of Manchester has been evaluated, using personal sampling pumps installed in the cabs of the vehicles and carried by the cyclist. These have provided an estimate of the average exposure of commuters using bus services and cycling in a congested European city. PMID- 10535140 TI - The influence of wooded shelterbelts on the deposition of copper, lead and zinc at Shakerley Mere, Cheshire, England AB - Research was undertaken to determine concentrations of Cu, Pb and Zn in soils and vegetation in shelterbelts and open field sites adjacent to the M6 motorway, a major arterial route, near Middlewich, Cheshire, UK. PMID- 10535141 TI - Environmental health concerns: optimised use of available knowledge. PMID- 10535142 TI - The fate of metals in Arctic surface waters. Method for defining critical levels. AB - Based upon studies in the industrially developed Arctic region, Russian Kola, here we discuss the fate of metals in high latitude surface water. Mainly, attention is paid to the priority pollutants from copper-nickel smelters. The influence of accompanying processes, such as acidification and eutrophication, on metal behavior is considered. The dramatic situation for fauna of Arctic latitudes is illustrated: (i) during the snow-melt, due to the pulse of ionic metal forms; and (ii) during the long polar winter in lower water layers, due to the involvement of a wide spectrum of metals in the redox-cycle under eutrophication and oxygen deficiency. Here we identify fish pathologies, which are related to the influence of metals. Generalizing the data on metal behavior, an original approach to define the integrated impact dose of metals--a toxicity index--has been developed. It presents a visualization of the integrated toxicity index for surface waters of the Russian Kola (based on the data for a 460-lake survey). As shown, there is a risk of fish diseases, due to both airborne contamination by metals and an indirect leaching by acid runoff over almost 30% of the area of the Russian Kola. For the Arctic region, polar winter stress syndrome will be repeatedly significant. During the polar night, as well as the spring, the vulnerability of the Arctic biota to toxic impact is higher. The accompaniment of water metal-pollution by two or more stressors would occur simultaneously, thereby multiplying the risk that it could develop. PMID- 10535143 TI - PCP in the freshwater and marine environment of the European Union. AB - This study collates monitoring data principally from 1991 to 1996, relating levels of pentachlorophenol (PCP) in the freshwater and marine environments of the European Union (EU) Member States of Belgium, France, Germany, the Netherlands and the UK. In general, PCP levels in the open freshwater and marine environments displayed a downward trend in these countries, and where increased PCP concentrations were observed, these were tentatively linked to specific point discharges. The monitoring data were compared against the relevant predicted no effect concentration (PNEC) for PCP in a particular environmental medium. On this basis, the study suggests that PCP does not pose a risk to the coastal, estuarine, marine waters or marine sediments of the above countries, and that any risks posed by PCP to the freshwater environment can largely be attributed to sediments contaminated with PCP from historic usage, or due to the continued use of small quantities of sodium pentachlorophenyl laurate in northern France. Further analysis of the data was not possible owing to the lack of systematic reporting by Member States, the lack of information on usage quantities and patterns, and inconsistent environmental surveillance and reporting of environmental data. This is particularly relevant for south-west France, Portugal and north-east Spain, the region which accounts for almost 90% of the EU's consumption of NaPCP. These issues need to be addressed at a pan-European level to better inform the selection of risk management measures and to improve the effectiveness of such measures. PMID- 10535144 TI - Assessment of pollution aerosols sources above the Straits of Dover using lead isotope geochemistry. AB - We assess the capability of lead isotopes to study the transport of pollution aerosols above the Straits of Dover by collecting atmospheric aerosols above the Eastern Channel and the Southern Bight of the North Sea. During the same period, we characterized the lead isotopic signature of the main industrial sources on the French coast near the Straits of Dover. Urban and automobile-derived aerosols were also collected. Due to the phasing out of lead in gasoline, the urban isotopic composition (206Pb/207Pb = 1.158 +/- 0.003) has become more radiogenic, although it is highly variable. On a regional scale, major industrial emissions have a well-defined isotopic composition (1.13 < 206Pb/207Pb < 1.22), more radiogenic than the petrol-lead signature (1.06 < 206Pb/207Pb < 1.12). These results together with those measured near the main coastal highway show that the automobile source has become a minor component of particulate lead in air. On a local scale, Dunkerque, the most urbanized and industrialized area along the Straits of Dover, may transiently control elevated lead concentrations. Except for the occurrence of local and regional range transport episodes, lead concentrations in the Straits of Dover can be related to remote or semi-remote pollution source emissions. Combining air mass retrospective trajectories and related lead abundances and isotopic compositions, it can be shown that lead aerosols originating from eastern Europe have an isotopic signature (1.145 < 206Pb/207Pb < 1.169) different from the isotopic composition of west-European lead aerosols (1.111 < 206Pb/207Pb < 1.142). The influence of remote North American sources is suggested, with caution, due to uncertainties in meteorological calculations. PMID- 10535145 TI - Air-pollutant dispersal patterns and vegetation damage in the vicinity of three aluminium smelters in Norway. AB - Dispersal patterns for fluoride and damage to vegetation was studied near three aluminium smelters in Norway. Leaf samples from three broad-leaved species (Betula pubescens Ehrh., Salix caprea L. and Sorbus aucuparia L.) were collected and leaf injury and the plants overall vitality were evaluated systematically in areas with different distance and direction from the emission sources. Both dispersal patterns and the distribution of damages were mainly determined by the predominant wind direction in the growing season and partly by topography. Damage was restricted to the areas closest to the smelters, within 2 km from the emission sources. Even with the average F- emission as low as 7.1 kg h-1, serious damage to vegetation was observed in built-up areas within 1 km south of the smelter in Mosjoen. Regression analysis showed a correlation between leaf injury and fluoride content in leaves within a locality, but great variation between localities. Leaf injury appeared at concentrations as low as 30 mg kg-1 in some species. An average leaf injury of 1 (scale 0-9) appeared at approximately 100 mg kg-1 in the vicinity of the northernmost smelter, Mosjoen, compared to approximately 150 and 300 mg kg-1 at Husnes and Ovre Ardal, respectively. Fluoride uptake and leaf injury differed among species. Approximately 100 species, mostly woody plants were evaluated for leaf injury and overall vitality. Conifers such as Pinus sylvestris L. and Picea omorika (Pan_) Purk. were sensitive. Except for Populus tremula L., most broad-leaved indigenous species were less sensitive. Species composition in natural areas as well as vegetation use in gardens were affected by pollution. In old gardens at the polluted site Ovre Ardal both the number of plant species and the area used for edible plants was lower compared to old gardens in an unpolluted nearby community. In newer gardens the use of both ornamentals and edible plants was more similar. PMID- 10535146 TI - History of accumulation of mercury and nickel in Thane Creek, Mumbai, using 210Pb dating technique. AB - The study area Thane Creek, lies on the southern part of the Deccan belt of India between latitude 18 degrees 53'-19 degrees 04' N and longitude 72 degrees 48'-72 degrees 53' E and includes the Ulhas river estuaries. Lead-210 (half-life of 22.3 years) is used for an estimation of recent sedimentation rate in Thane Creek using radiochemical separation and alpha counting of its grand-daughter 210Po. The concentration of Hg and Ni in vertical depth profiles of sediment is estimated to assess the anthropogenic input of these nuclides due to large-scale industrialization which has taken place at this site during last two decades. Depth-wise concentration profiles of Hg and Ni indicate positive evidence of continued fresh inputs into the Airoli section of Thane Creek. PMID- 10535147 TI - Arsenic in the Meager Creek hot springs environment, British Columbia, Canada. AB - Levels of arsenic in water from Meager Creek hot springs, British Columbia, Canada, were found to be naturally elevated. Biota including microbial mats, green algae, sedge, cedar, fleabane, monkey flower, moss, mushrooms and lichens, that were expected to be impacted by the water, were analyzed for total levels of arsenic and for arsenic species. The major arsenic species extracted from all samples were arsenate and arsenite, which are toxic forms of arsenic. Additionally, small amounts of arsenosugars X and XI were detected in microbial mats and green algae, implying that cyanobacteria/bacteria, and possibly green algae are capable of synthesizing arsenosugars from arsenate. Low to trace amounts of arsenosugars X and XI were detected in lichens and the fungus Tarzetta cupularis. A large fraction (on average, greater than 50%) of arsenic was not extracted by using methanol/water (1:1) and the chemical and toxicological significance of this arsenic remains unknown. PMID- 10535148 TI - Atmospheric heavy metal deposition plumes adjacent to a primary lead-zinc smelter. AB - A method for the determination of atmospheric heavy metal deposition rates has been developed using 0.5-m2 deposition trays at 0.1 m from the ground. Trays were spaced at 150-m intervals along a 1500-m line 500 m east of a Pb-Zn smelter. Ten sampling events of 1-3-h duration were conducted under westerly wind conditions so as to determine the sources of heavy metals deposited near the smelter. Deposited materials were sampled from the trays using wipes. There was good agreement between deposition trays placed side-by-side and exposed in pairs. Under certain conditions, however, the method is not appropriate owing to the potential for local contamination. Geometric mean deposition rates for Pb, Zn, Fe, Cu, As and Cd averaged over a nominal plume width of 600 m amounted to 18.8, 22.2, 12.2, 0.614, 0.403 and 0.052 mg m-2 day-1, respectively. Gaussian deposition profiles were seen for Pb, Zn, Fe, Cu, As and Cd downwind from the blast furnace, sinter plant, and refinery area. Zinc deposition could also be attributed to a northern Zn production area. This northern site was not generally associated with elevated Pb deposition. On the basis of this work, the deposition of heavy metals in residential areas adjoining the smelter is likely to occur downwind from the smelter site, with deposition rates increasing with wind speed. The strategic measurement of heavy metal dry-deposition rates over short periods of time using large collection surfaces provides source-specific information not obtainable by conventional long-term 'passive' deposition sampling. Lower detection limits than those achieved here are likely to be achieved in non smelter settings. Previous suggestions implicating a sink of city surface dusts as the probable source of Pb recontamination of residential settings in the absence of ongoing smelter emissions are not supported by this work. PMID- 10535149 TI - Indoor air quality at the Correr Museum, Venice, Italy. AB - Two multidisciplinary field surveys, one in winter and the other in summer have monitored the indoor microclimate, air pollution, deposition and origin of the suspended particulate matter and microorganisms of the Correr Museum, Venice. In addition, this study was focused to identify the problems caused by the heating and air conditioning system (HAC) and the effects due to the presence of carpets. Heating and air conditioning systems (HACs), when chiefly designed for human welfare, are not suitable for conservation and can cause dangerous temperature and humidity fluctuations. Improvements at the Correr Museum have been achieved with the assistance of environmental monitoring. The carpet has a negative influence as it retains particles and bacteria which are resuspended each time people walk on it. The indoor/outdoor pollutants ratio is greater in the summertime, when doors and windows are more frequently open to allow for better ventilation, illustrating that this ratio is mainly governed by the free exchange of the air masses. The chemical composition, size and origin of the suspended particulate matter have been identified, as well as the bacteria potentially dangerous to the paintings. Some general suggestions for improving indoor air quality are reported in the conclusions. PMID- 10535150 TI - Fish model for assessing the in vivo estrogenic potency of the mycotoxin zearalenone and its metabolites. AB - The in vivo estrogenic potency of zearalenone (ZEA), a mycotoxin produced by different strains of Fusarium fungi, and its metabolites (alpha- and beta zearalenol), have been studied in fish. Estrogenicity was evaluated using an in vitro competitive receptor binding assay and in vivo induction of vitellogenesis and zonagenesis, two estrogen receptor (ER)-mediated responses that are integral aspects of fish oogenesis. The ER binding affinities of alpha-zearalenol and ZEA in rainbow trout (Oncorhynchus mykiss) were approximately 1/150 and 1/300 to that of estradiol, respectively. Juvenile salmon (Salmo salar) were exposed to a single intraperitoneal injection of ZEA, alpha-zearalenol and beta-zearalenol (each at 1 and 10 mg/kg) and compared to fish injected with estradiol-17 beta (E2; 5 mg/kg) and controls. Using indirect enzyme-linked immunosorbent assay (ELISA) with homologous antibodies, a dose-dependent induction of vitellogenin (Vtg) and eggshell zona radiata proteins (Zr-proteins) were observed 7 days after exposure to ZEA and alpha-zearalenol. beta-Zearalenol did not elevate plasma Vtg levels, but a non-significant elevation of plasma Zr-proteins levels was observed at the highest dose (10 mg/kg). Generally, alpha-zearalenol and ZEA possess estrogenic potencies that are approximately 50% compared to that of E2, and their order of estrogenic potency (in both in vitro receptor competitive binding and in vivo induction of Vtg and Zr-proteins levels) is: alpha-zearalenol > ZEA > beta zearalenol. Our results show that blood plasma analysis of Vtg and Zr-proteins levels provides a suitable in vivo fish model for assessing the estrogenic potencies of ZEA and its metabolites. PMID- 10535151 TI - Influence of microbes on the mobilization, toxicity and biomethylation of arsenic in soil. AB - To understand the effects of microbial activity on the mobilization and speciation of arsenic in soil, the cycling of arsenic was studied in microcosm experiments under laboratory conditions. Particular attention was paid to the biomethylation of arsenic and to the toxicity of inorganic and organic arsenic species for microbes. Microbes enhanced mobilization of arsenic from soil by 19 24% compared to formaldehyde inhibited controls. Formation of dissolved methylated arsenic species by microbes was low (< 0.1%) during the 5-day incubation. Even though methylation may function as a detoxification method, it was of minor importance in the soil tested. PMID- 10535152 TI - Heavy metal release from phosphorite tailings into seawater: a simulated laboratory study. AB - A number of laboratory experiments were carried out to assess the desorption of trace metals from sedimentary phosphorite samples into artificial seawater. Shaking and percolation experiments have been performed on phosphorite samples from the Cd-rich phosphorite deposits of Hahotoe-Kpogame (Togo, West Africa) using artificial seawater in the ratio 1:10. The results show that elevated concentrations of trace elements Cd(17-256 micrograms/l), Ni (12-193 micrograms/l), Zn (21-200 micrograms/l) and major elements Al (6-1915 micrograms/l) and Fe (30-1264 micrograms/l) are released into seawater by desorption processes. Maxima of trace metals with high mobility are reached within 4 h indicating the fast desorption kinetics in seawater. Thus, the direct disposal of potentially toxic metal-rich mine tailings may lead to regional coastal water pollution. PMID- 10535153 TI - Trends in alkanes and PAHs in airborne particulate matter from Oporto and Vienna: identification and comparison. AB - A total of 56 weekly samples from Oporto and 40 from Vienna were collected and analysed, for 23 n-alkanes and 16 polycyclic aromatic hydrocarbons (PAH), by GC MS after extraction with a toluene/methanol mixture. Total and elemental carbon were in the same range of values for both sampling sites. Although parts of the spectrum of species in both sampling sites were constant over the sampling period there is no evidence for suggesting a universal tracer for alkanes. For both Oporto and Vienna, Principal Component Analysis (PCA) identified two PCs and in both cases the first PC contained only PAHs while the second contained only alkanes. This separation between alkanes and PAHs and the observation of an alternating pattern with higher concentrations of odd carbon numbered from C27 to C30 (natural emissions from biological origin) is believed to result from a separation between anthropogenic and biological contributions associated with the first and second PC, respectively. PMID- 10535154 TI - [Human in vitro fertilization: technics and ethics]. AB - IVF is certainly one of the new medical technologies where ethical debates are the most frequent and heavy. Three illustrations will be presented: The improvement of ovulation induction techniques have increased the number of supernumerary embryos. How to deal with these embryos, on which principles and what do the patients think about it? Introduction in the early 90' of assisted fertilization techniques (ICSI) have largely extended IVF indications and improved the success++ rate. Nevertheless, it will be demonstrated that it's implementation was a hard process for which the creation of embryos for research was a crucial step, opening another passionate ethical debate. Oocyte donation which came true thanks to IVF, will give the opportunity to illustrate an unusual consequence of ethics on technical aspect; due to an ethical choice (anonymous donation) we have been able to improve by a factor 3 the clinical efficiency of our oocyte donation program. An example of enhancement of the technique due to ethics, a very important fact in the situation of shortage of oocytes for donation. In conclusion, it appears that the ethical debate does not only focus on the legitimacy of this "medicine of desire" but also demonstrate that the development as well as the daily practice of IVF makes a clear vision of ethical choices mandatory and indissociable of technical aspects themselves. PMID- 10535155 TI - [The evolution of man]. PMID- 10535156 TI - [In memoriam Jean Lecomte]. PMID- 10535157 TI - [The C. Heymans Memorial International Symposium 1938-1969-1998. The Knokke Casino Auditorium--17 October 1998]. PMID- 10535158 TI - [Observations on the Federation of National Academies of Medicine of the European Union. Extraordinary session of the Portuguese Academy of Medicine (Lisbon, 25 July 1998)]. PMID- 10535159 TI - [Myocardial viability in man: state of the art and directions to pursue]. AB - The advent of modern coronary revascularization procedures has profoundly modified the prognosis of patients with acute myocardial infarction or that of patients with chronic left ventricular ischemic dysfunction. Presumably, the beneficial effects of revascularization result from improving blood supply to dysfunctional but viable regions with subsequent improvement in regional and global left ventricular function. Various approaches have been proposed to predict the reversibility of left ventricular dysfunction after coronary revascularization. For the most part, these techniques allow detection of viable myocardium with a high sensitivity (80%). However, specificity has been more varied (+/- 55% for, +/- 75% for PET and +/- 85% dobutamine echocardiography). All these techniques also bear important prognostic implications that are independent and complementary to those usually available in these patients. PMID- 10535160 TI - [Intracerebral grafts in Parkinson disease. Current experiences and perspectives]. AB - Six patients with advanced Parkinson's disease have been grafted with human fetal neuronal cells. This clinical work is based on the technique originally described by the groups of Lund, Sweden and Creteil, France, with which we closely collaborate. A first group of 3 patients was grafted between 1995 and 1996, with neuronal tissue obtained from same-day abortions. We have observed bilateral improvement of motor functions after unilateral transplantation into the putamen. One patient had a second graft, into the heterolateral putamen, one year after the first one. Another group of 3 patients was operated between 1997 and 1998, and received grafts of hibernated tissue, in order to facilitate the technical organization of the transplantation procedure. Altogether, these clinical results are encouraging, but the technical and ethical limitations related with the use of human fetal tissue prompted us to develop alternative solutions, such as the use of xenografts and the transplantation of genetically modified cells. PMID- 10535162 TI - [Development of the scientific basis of Czech pharmacy 1939-1945]. AB - The German occupation of the Czechoslovak Republic in 1938-1945 and the worldwide war conflict which limited the import of drugs and medicines into the Czech Lands were a stimulus for the development of Czech pharmaceutical industry and research, which incorporated experts-lecturers and students from the closed Czech universities. Important roles in obtaining and disseminating scientific knowledge in pharmaceutical sciences were played by the Institute for Drug Research in Prague, Central Union of Pharmacists, and a number of pharmacies. In drug research, several new preparations were devised; the greatest achievement was the method of manufacture of "Czech" penicillin, which was used already during the war. The inland basis of raw materials was enlarged by cultivation and collection of medicinal plants. Scientific knowledge gained during the war in pharmaceutical chemistry, pharmacognosy and galenical pharmacy contributed after the liberation in 1945 to the establishment of pharmaceutical sciences, which had not had a sufficient basis before. PMID- 10535161 TI - [Postantibiotic effects of antimicrobial agents]. AB - The present authors review data from the literature concerning the postantibiotic effect (PAE) and the postantibiotic effect of subinhibitory concentrations (PA SME) of some antibiotics on clinically significant G- and G+ bacterial species. Attention is paid to the influence on their physiological characteristics with regard to virulence factors. PMID- 10535163 TI - [From Purkinje's pharmacologic observations to molecular drug interactions]. AB - The 650th anniversary of the foundation of Charles University (7 April 1348) in Prague has initiated a number of historical surveys of the subjects which has been taught at the University for a longer period of time. The disciplines connected with pharmacotherapy were being developed in an empirical conception at the University from the second half of the 14th century but the beginnings of experimental drug research date as late as the mid-19th century. The present survey of the history of "the sciences of medicaments" therefore attempts to outline in short entries the developmental stages of pharmaceutical and pharmacological investigations in the territory of Bohemia and Moravia in about recent 150 years. The arrangement of data is chronological; in the part covering the second half of the 20th century the research of a predominantly exploratory character (universities and academic institutions and their representatives) and research aimed primarily to innovate medicaments (research institutions of pharmaceutical industry and clinical pharmacology and some of their representatives) are treated separately. PMID- 10535164 TI - [Prediction of pregnancy-induced hypertension in women with risk factors]. AB - The objective of the presented work was to find possible predictive factors of pregnancy-induced hypertension (PIH) in women with the risk of gestational diabetes (GDM) during pregnancy. A group of women with the risk of development of gestational diabetes was selected because it is known that in women with GDM a hypertension is encountered 2-4 times more frequently than in women without disorders of glucose tolerance during pregnancy. The patients were divided into four groups with regard to the risk of development of hypertension. The examinations were made once during the 20th week of gestation. PIH is manifested in the great majority of patients only after the 20th week of pregnancy. All patients were normotensive at the time of examination. During the subsequent course of pregnancy in all patients the diagnosis of gestational diabetes and pregnancy-induced hypertension was investigated. The investigated parameters were fasting insulin levels during the 20th week of gestation, coagulation factors- antithrombin III (AT III), D-dimer, number of thrombocytes. The authors investigated also anamnestic data on the presence of type II diabetes and hypertension in close relatives of the patients. RESULTS: No differences were found in the insulin, antithrombin III and D-dimer levels nor in the number of thrombocytes. A significant relationship was revealed between the incidence of hypertension during pregnancy and the family history of the patient. In women where there was diabetes type 2 and hypertension in the family, there was a markedly more frequent incidence of pregnancy-induced hypertension after the 20th week of pregnancy (p > 0.0002) than in the other investigated groups. PMID- 10535165 TI - [Importance of anticardiolipin antibody screening in pregnancy]. AB - We performed 1698 examinations of anticardiolipin antibodies (ACLA) among pregnant women with gestational age from 16 to 37 weeks of pregnancy during eleven months of 1996. The ACLA levels above normal range were found in patients with pregnancy induced hypertension, preeclampsia, gestational diabetes, diabetes mellitus type I, venous thrombosis, thrombocytopenia and rheumatological diseases. The following results show the possible relations between high ACLA levels and pregnancy induced hypertension and gestational diabetes. The screening test for ACLA is recommended as a supportive method for prenatal follow-up of pregnant patients. PMID- 10535166 TI - [Effect of combined administration of TRH and dexamethasone in pregnant women and the course of their pregnancy]. AB - Extremely immature neonates are threatened during the first days after delivery by many conditions which are due to incomplete development.--A key role is played during the first days of extrauterine life by the incidence and degree of the respiratory distress syndrome (RDS). Its incidence in neonates born before the completed 32nd week of gestation is very common. Causal treatment of RDS is not known. To overcome it the neonatologist must use in the majority of infants invasive techniques of controlled ventilation which are associated with the risk of further complications such as barotrauma, retinopathy and later the development of bronchopulmonary dysplasia. Attempts to influence intrauterine maturation of the lungs were started in the fifties. As a routine procedure nowadays corticoids are administered antenatally. Their limited effect divert the attention of perinatologists to other substances which could enhance maturation of pulmonary tissue. In human medicine ambroxol was introduced, in animals opiates are tested as well as beta-mimetics, aminophylline. The greatest hopes were aroused by trials with the use of T-hormones. T-hormones have a maturating regulating function in the foetal organism. They have an affinity for pneumocytes and in animal experiments they have a positive effect on surfactant formation. Moreover they act synergically when combined with corticoids. OBJECTIVE OF STUDY: a) to evaluate the safety of the method from the aspect of undesirable side effects of hormone administration to the mother b) evaluation of hormone levels: TSH, total T4, total T3, TRH and prolactin in maternal serum. PMID- 10535167 TI - [Hormone therapy and urogynecology]. AB - The female genital and urinary systems exists in close anatomical and functional proximity, disorders of one resulting in dysfunction of the other. The investigation and management of lower urinary tract disorders must take this important relationship into consideration, as neither can be viewed in isolation. The value of estrogen replacement therapy as a treatment of urinary incontinence is controversial and until today there is a little substantial evidence to conclude that estrogen therapy alone is of value in the treatment of this symptom. This conflicting evidence concerning the therapeutic benefit of estrogen therapy in stress urinary incontinence seems to be outweighed with other advantages of estrogen replacement therapy. Clear evidence exists to suggest that recurrent urinary tract infections can be prevented or even treated by the use of estrogen therapy. Systemic estrogen replacement appears to relieve the symptoms of urgency, urge incontinence, frequency, nycturia and dysuria, and low-dose topical estrogen is effective in the management of atrophic vaginitis. Even with postulating the HRT to be of enormous therapeutic value to postmenopausal women in urogynecology it may stay only a mean of support of other causal methods of treatment of dysfunction of lower urinary tract. PMID- 10535168 TI - [Endometrial ablation]. PMID- 10535169 TI - [Importance of choosing a pathology laboratory for objective evaluation of findings in women after conization]. PMID- 10535170 TI - [Initial experience with Mirena in the Czech Republic]. PMID- 10535171 TI - [The Lahodny method of reconstructive correction of genital prolapse and urinary incontinence]. PMID- 10535173 TI - [The 110th anniversary of the founding of the Medical School Maternity Hospital in Brno]. PMID- 10535172 TI - [Historical legacy of Schauta's operation and its modification today]. PMID- 10535174 TI - [Birth rate in the Czech Republic within the European context: demographic analysis for 1990-1996]. PMID- 10535175 TI - [Pharmacologic and toxicologic results in genetic polymorphisms (review)]. AB - Paper provides a survey of the basic knowledge on the genetic polymorphism of enzymes involved in the metabolism of medicinal drugs and other xenobiotics. The major implications of this phenomenon have been described, namely the interindividual variability in the therapeutic effect of drugs differently metabolised by the polymorphically determined enzymes, or the interindividual variability in the susceptibility to neoplastic processes related to the polymorphically determined enzymes metabolising xenobiotics. Recent advances in the understanding of the molecular genetics of enzymes metabolising drugs and other xenobiotics (particularly cytochrome P-450, glutathione S-transferase, N acetyltransferase, UDP-glucoronosyltransferases and others), help to understand the molecular basis of several genetic polymorphisms. Present article reviews the current status of studies on polymorphism of xenobiotics-metabolising enzymes and brings a discussion to their pharmacotherapeutical relevance and to their significance for identification of susceptible individuals/subgroups in the carcinogen exposed population. PMID- 10535176 TI - A 64-year-old woman with fever and acute polyarthritis. PMID- 10535177 TI - Minimizing the risk of NSAID-induced GI bleeding. AB - Although nonsteroidal anti-inflammatory drugs (NSAIDs) have definite indications and can offer relief for many conditions, many patients have severe gastrointestinal problems after taking them. Physicians should prescribe these drugs more selectively and advise patients to limit their use of over-the-counter NSAIDs. For patients at high risk who need an NSAID, a prophylactic drug or a cyclooxygenase 2-selective NSAID can decrease the risk. PMID- 10535178 TI - In diagnosing hepatitis C, which patient needs which test? PMID- 10535179 TI - Human papillomavirus typing and the reduction of cervical cancer risk. AB - Minor cervical cytologic abnormalities are common, but knowing which low-grade lesions will progress to cervical cancer--and therefore deserve biopsy and excision--is difficult. Since some human papillomavirus (HPV) types are strongly associated with cervical cancer, HPV typing may be a means of determining which patients with minor abnormalities require biopsy and treatment and which need only follow-up smears. This paper reviews the association between cervical cancer and HPV infection, the pathogenesis of HPV infection, the utility of HPV typing in training patients with a diagnosis of atypical squamous cells of undetermined significance, and the prospects for the development of an HPV vaccine. PMID- 10535180 TI - Searching the Internet for medical information: practical tips. AB - The Internet can be useful for physicians, but beginning users are apt to encounter two problems: too much irrelevant information, and difficulty finding the desired information. This paper offers some practical tips that can prevent frustration and wasted time when searching for medical information on the Internet. PMID- 10535181 TI - Keeping our patients' secrets. AB - Protecting the privacy of the patient's medical record is a central issue in current discussions about a patient bill of rights, and controversy over a proposed "unique health identifier" has raised the decibel level of these discussions. At the heart of the debate is how best to resolve the inherent conflict between the individual's right to privacy and the need for access to patients' health information for reasons of public health, research, and health care management. PMID- 10535182 TI - The pathogenesis and spectrum of acute coronary syndromes: from plaque formation to thrombosis. AB - Acute coronary syndromes occur when an unstable atherosclerotic plaque erodes or ruptures, exposing the highly thrombogenic material inside the plaque to the circulating blood and triggering rapid formation of a thrombus that occludes the artery. This understanding is leading to a wide variety of new therapies. It is hoped that further research into the pathogenesis of acute coronary syndromes will lead to more accurate markers of risk and more specific preventions and treatments. PMID- 10535183 TI - [Utility of clinical evaluation in the diagnosis of community-acquired pneumonia]. PMID- 10535184 TI - [Strains of ganciclovir-resistant cytomegalovirus]. AB - BACKGROUND: In countries where the resistance of cytomegalovirus to ganciclovir has been studied, strains resistant to therapeutic doses of this drug have been isolated. When a change in treatment has been impossible the patient has shown bad clinical evolution. The aim of this study was to investigate the presence of these strains in our medium, observe whether the resistances appear in patients previously treated with ganciclovir and determine its implication in the evolution of cytomegalovirus infection. PATIENTS AND METHODS: One hundred twenty three stains of cytomegalovirus, isolated during the period 1990-1998, corresponding to the following 94 patients were studied: 17 breast feeding children of healthy parents who were negative controls (sensitive strains), 43 organ transplant recipients, 29 AIDS patients, 2 with other immunodeficiencies and 3 children with intrauterine infection. Seventeen patients were studied due to the insidious course of the infection despite treatment. The remaining were random. The technique used was that of growth inhibition of the strains seeded on different gradients of ganciclovir: 0, 1, 5, 10 and 20 microM. The inoculate consisted in a cellular suspension evaluated according to the degree of viral growth. The strains presenting an inhibitory doses 50% (ID 50%) greater than 10 microM were considered as resistant. RESULTS: Eighty-two strains presented an ID 50% lower than 5 microM, 24 from 5 to 10 microM and in the 17 remaining strains, corresponding to 12 patients, the ID 50% was greater than 10 microM. The evolution of these latter 12 patients with strains considered to be resistant to ganciclovir was of death in 8. All were immunodepressed and with a history of having previously received ganciclovir. Another currently has a chronic evolution and the three remaining patients, who presented better immunity, became cured. All patients has a chronic evolution and the three remaining patients, who presented better immunity, became cured. All patients had undergone previous treatment with ganciclovir except two: one patient with Wegener disease treated with acyclovir 15 days before, and the other was an infant of an HIV positive mother who had received the drug. CONCLUSIONS: The presence of cytomegalovirus strains resistant to ganciclovir was confirmed in our patients. The previous use of ganciclovir and, in one case of acyclovir, appears to be implicated in the appearance of resistance. The evolution of the immunodepressed patients with infection by resistant strains was mortal except when their immunity was improved. PMID- 10535185 TI - [Vancomycin-resistant enterococci: in vitro activity of quinupristin / dalfoprostin (RP 59500)]. AB - BACKGROUND: The recent emergence of glycopeptide-resistant enterococci limits the treatment of enterococcal infections. The aim of this study was to evaluate the in vitro activity of a new streptogramin, quinupristin/dalfopristin, against 30 clinical isolates of vancomycin-resistant enterococci and compared with those of other 15 antimicrobials. MATERIAL AND METHODS: Enterococci were identified by using Rapid ID 32 Strep system. Genotyping of the isolates was performed by PCR. The MICs of quinupristin/dalfopristin were determined by the agar dilution technique recommended by the NCCLS. Susceptibilities to the rest of antibiotics tested (teicoplanin, ampicillin, penicillin, imipenem, doxycicline, chloramphenicol, gentamicin, streptomycin, rifampin, levofloxacin, fleroxacin, trovafloxacin, sparfloxacin, pefloxacin and clinafloxacin) were determined by using the E test. beta-lactamase production was examined with nitrocefin disks. RESULTS: Quinupristin/dalfopristin has demonstrated excellent activity against Enterococcus faecium (MIC90' 2 micrograms/ml). Enterococcus faecalis was considerably less susceptible than E. faecium, at concentration of 4 micrograms/ml inhibited only 31% of tested strains. For doxycicline 77% of strains were susceptible. Only five isolates were susceptible to clinafloxacin; the other quinolones tested displayed poor activity. Resistance to chloramphenicol was detected in 47% of isolates. None of the isolates produced beta-lactamase. CONCLUSIONS: This study indicates that vancomycin-resistant enterococci are often concomitantly resistant to multiple antibiotics. Quinupristin/dalfopristin was the most active agent tested against E. faecium strains. On the basis of these results quinupristin/dalfopristin could be a therapeutic option for the treatment of vancomycin-resistant E. faecium infections. PMID- 10535186 TI - [Corynebacterium amycolatum: sepsis in hematologic patients]. AB - BACKGROUND: To evaluate clinical and microbiological characteristics of sepsis due to Corynebacterium amycolatum, because few cases have been reported in human infections by this bacteria. METHODS: We report two cases of sepsis due to Corynebacterium amycolatum, the patients were diagnosed as having leukaemia. The identification procedures are discussed. RESULTS: We describe 2 patients with sepsis in neutropenic phase. Antibiotic treatment was successful in both cases. Clinical isolates were classified as Corynebacterium xerosis by the Api Coryne system Version 1.0. The chemotaxonomic characteristics of the isolates were determined by thin-layer chromatographic analysis. Isolates did not contain mycolic acids, they were identified as Corynebacterium amycolatum. CONCLUSIONS: Sepsis due to Corynebacterium amycolatum is a rare disorder. A test for mycolic acid could be sufficient to make a distinction between Corynebacterium amycolatum and other diphteroid bacteria. Such a test involves thin-layer chromatography. PMID- 10535187 TI - [The Helicobacter pylori adhesion gene: relation with the origin of the isolates and associated disease]. AB - BACKGROUND: The adhesion gene (hpaA) of Helicobacter pylori isolates from different Spanish regions was studied by PCR-RFLP in order to group the strains according to the origin of the strains and to relate it with ulcer or gastritis production. METHODS: One hundred and forty three Helicobacter pylori clinical isolates were obtained from the culture of gastric biopsies (29 from Ibiza, 39 from Madrid, 36 from Almeria and 39 from Aviles). After DNA extraction, a PCR was performed to amplify a 375 pb fragment of the hpaA gene which was digested with Hinfi and SauIIIa. RESULTS: The gene was detected in all of the strains studied. After digestion with Hinfi and SauIIIa, three and four patterns called 1, 2 and 3 and A, B, C and D were obtained, respectively. Strains from Madrid and Almeria showed the highest hpaA genetic variability. In the relationship with the pathology, patterns 1 and A were the most prevalent in strains obtained from patients with ulcer and with gastritis. CONCLUSIONS: The results of hpaA gene study show the genetic variability of Helicobacter pylori and slight differences according to the origin were found. PMID- 10535189 TI - [Molecular typing and sensitivity of Candida albicans isolates proceeding from critically ill patients]. AB - BACKGROUND: Nosocomial infection due Candida albicans in immunocompromised patients are recognized as a significant cause of morbidity and mortality. The endogenous forms of infections may require effective strategies for prevention which are different from those for exogenous infections due to transmission of any organism from patient to patient. Typing by PCR of isolates of C. albicans maybe useful for that. METHODS: Twenty-four isolates in blood cultures of 24 critically ill patients were studied. Typing by interrepeat PCR and in vitro antifungal susceptibility tests was performed by a microdilution. RESULTS: Twenty one different genotypes were obtained. The isolates with same genotype shown different patterns of susceptibility. With one strain a band pattern very different from that obtained with the remaining isolates. CONCLUSIONS: No were relation between strain of same unit. The isolates with same genotype was different critically unit or year of isolation. PMID- 10535190 TI - [How to administer indinavir. Consensus group]. PMID- 10535191 TI - [Arthropathy in an Equatorial Guinea patient undergoing kidney transplantation]. PMID- 10535188 TI - [Pyrimethamine and sulfadoxine as a preventive treatment for Pneumocystis carinii and Toxoplasma encephalitis]. AB - BACKGROUND: Few studies have been reported on the potential usefulness of Fansidar combination at a fixed doses of pyrimethamine and sulfadoxine as prophylaxis for pneumonia by Pneumocystis carinii and toxoplasmic encephalitis. METHODS: The clinical histories of the patients seen in our department from January, 1992 to December, 1994 who were prescribed pyrimethamine and sulfadoxine for the above prophylactic treatment were reviewed. RESULTS: One hundred fifty clinical histories fulfilled the requisites for evaluation. Thirty-seven patients (24%) had a previous diagnosis of AIDS, with the median CD4 count being of 134/mm3. The median follow up of treatment with Fansidar was of 31 months. Twenty eight cases of pneumonia by Pneumocystis carinii (6.9 cases for 100 patients years) and 10 cases of toxoplasmic encephalitis (2.5 cases for 100 patients years) were diagnosed. CONCLUSIONS: These values are similar to those presented with respect to other pharmacologic interventions commonly used for the prevention of pneumonia by Pneumocystis carinii and toxoplasmic encephalitis and suggest that pyrimethamine and sulfadoxine is a useful alternative and requires consideration in individualized cases for the prophylaxis of these diseases. PMID- 10535192 TI - [Subcutaneous infection in the foot of an immunosuppressed patient]. PMID- 10535193 TI - [Antibodies against human parvovirus B19 in the Madrid community]. PMID- 10535194 TI - [High incidence of Creutzfeldt-Jakob disease in Soria]. PMID- 10535195 TI - [The persistent and notable sensitivity of Brucella to tetracyclines]. PMID- 10535196 TI - [Hemophagocytic syndrome associated with lymphoma in an HIV-infected patient: presentation of a case and review of the literature]. PMID- 10535197 TI - [Lemierre syndrome caused by Porphyromonas endodontalis]. PMID- 10535198 TI - [Parenteral antibiotics at home]. PMID- 10535199 TI - [Comparison between indirect immunofluorescence and shell vial cultured in the detection of adenovrius in respiratory specimens]. PMID- 10535200 TI - [Hepatic fascioliasis resistant to bithionol treatment but responsive to triclabendazole]. PMID- 10535201 TI - [Re-emergence of acute lymphocytic meningitis caused by mumps virus in Spain]. PMID- 10535202 TI - Proposal of a new hypothesis for the psychosomatic treatment of obesity and its application. AB - Dieting or a change in eating habits is the most widely used approach aimed at reducing body weight. However, it is also well known that many obese people cannot reduce body weight substantially, no matter how hard they try, and that they soon regain whatever they do lose. The conventional approach to the treatment of obesity is to control it by prohibition or suppression of overeating, and by orders to change eating habits. This paper presented and examined a new psychosomatic approach for obesity (NPAO). Taking the story of "The North Wind and the Sun" from Aesop's Fables as a metaphor, this hypothesis is based on the reduction of overstressors through a "Sun"-type approach as opposed to a "North Wind"-type approach. This "Sun"-type approach, which incorporates 2 principles and 3 basic rules, is useful in decreasing stressors such as prohibition, suppression and orders, and increasing pleasantness, which competes with unpleasant stress. The treatment based on this hypothesis was applied to 77 subjects: 62 men (age 46.2 +/- 8.0 years) and 15 women (age 50.6 +/ 4.5 years). All subjects were given medical checks just before and 6 months after the psychosomatic approach for obesity. For a proportion of cases, maximal oxygen uptake (VO2max) was measured before and after. In the practiced group (48 cases) except for three persons who had stopped the program within 3 months after the start, body weight and body mass index fell significantly by 5.2 kg (p < 0.001) and 2.0 kg/m2 (p < 0.001) respectively, after 6 months. There were significant reductions in total cholesterol and triglyceride (p < 0.01, p < 0.01 respectively). VO2max, however, increased significantly (p < 0.05). The subjects' impressions of this therapy, collected after 6 months were as follows: "It was comfortable" 67.7%, "It was hard going" 8.8%, "My body has become lighter" 79.4%, "I have become more energetic" 70.5%, and "I have become happier" 64.7%. During the period of the therapy, there was no report of any appearance of new physical or mental abnormalities such as fatigue or uncomfortableness. On the other hand, there were no significant changes in any parameters except for an increase of blood sugar in the non-practiced group (26 cases). These results strongly indicate that the NPAO is easy in practice, has a high success rate, shows no rebounding, reduces body weight safely, and results in an increase of vigor. PMID- 10535203 TI - [Incidence of inflammatory bowel disease in Aragon: outcome of a prospective population-based study]. AB - OBJECTIVE: To identify every new case of ulcerative colitis (UC) (including ulcerative proctitis), Crohn's disease (CD) and indeterminate colitis (IC) in Aragon, in Spain (population: 1,189,000, area: 47,719 km2) and to compare the incidence in this region with that in the rest of Spain and Europe. PATIENTS AND METHODS: We designed a prospective, population-based study based on inception cohorts. During a 3-year predetermined period (1st February 1992-31st January 1995) we identified every new case of inflammatory bowel disease in Aragon by checking the records in all the hospitals, outpatient clinics and private practices in this region. RESULTS: The overall adjusted incidence rate per 100,000 inhabitants/year was 7.2 for UC (a figure lower than the average for Europe) and 3.9 for Crohn's disease (a rate similar to that of Southern Europe). These rates are much higher than those previously described in Spanish studies, probably due to the design and methods used as well as to a real increase in the incidence of CD in Spain. The age and sex pattern was similar to those other studies. CONCLUSIONS: The incidence rates for inflammatory bowel disease in Aragon are higher than those previously described, the incidence of UC being inferior to that of Europe. Nevertheless, the incidence of CD is similar the average for southern Europe, which suggests that there has been a recent increase in the incidence of this disease in Aragon. PMID- 10535204 TI - [Liver retransplantation in adults: clinical course and results of 13 years' experience]. AB - INTRODUCTION: Liver retransplantation is the only alternative to irreversible graft failure. However, it remains a controversial treatment. The aim of this study was to analyze the clinical course and the results of liver retransplantation in our center. PATIENTS AND METHODS: The actuarial survival in a series of 54 retransplantations in 49 patients between February 1984 and December 1997 was analyzed. The retransplantations were grouped according to period: group A (n = 16) 1984-1992, group B (n = 22) 1993-1995 and group C (n = 16) 1996-1997. RESULTS: The actuarial survival per group according to year was: 31.25%, 54.55% and 62.50% for groups A, B, and C, respectively, which shows a clear improvement with time, although differences were not statistically significant. CONCLUSIONS: The results of liver transplantation in our series show a lower actuarial survival rate than those of primary transplantation but these results have improved in recent years. PMID- 10535205 TI - [Accuracy of imaging techniques and tumor markers in the diagnosis of pancreatic cancer]. AB - OBJECTIVE: The aim of this study was to evaluate the diagnostic accuracy of: a) three imaging techniques: ultrasonography (US), computed tomography (CT) and endoscopic retrograde cholangiopancreatography (ERCP), and b) the serum tumor markers CEA, CA 19-9 and CA 125, in the diagnosis of pancreatic cancer. PATIENTS AND METHODS: A total of 137 patients were prospectively evaluated. Pancreatic cancer was diagnosed in 25 patients; chronic pancreatitis in 24; acute pancreatitis in 22; extrapancreatic malignancies in 24, and benign digestive disease in 42. The reference interval for each marker was determined in 36 healthy volunteers. Diagnostic accuracy was determined using receiver operating characteristic (ROC) curves. Each diagnostic test was used in order: a) to diagnose pancreatic cancer when the disease was clinically suspected, and b) to differentiate between pancreatic cancer and chronic pancreatitis. RESULTS: CT (0.976-0.888) and US (0.857) diagnostic accuracy resulted greater than the best serum tumor marker CA 19-9 (0.755-0.786) (p = NS), in both diagnostic conditions. To obtain a diagnostic specificity of 90% in pancreatic cancer, the CA 19-9 cutoff level should increase up to 3 and 7.5 folds being in this case better than CEA and CA 125 (p < 0.05). CONCLUSIONS: CT scan offers the greatest accuracy in the diagnosis of PC. Among the tumor markers, CA 19-9 offers the best accuracy in the diagnosis of pancreatic cancer. PMID- 10535206 TI - [Multidrug-resistant Salmonella a significant clinical problem?]. AB - The incidence of infections caused by Salmonella strains resistant to antibiotics, including ampicillin, chloramphenicol, streptomycin, sulfonamides, tetracycline and even amoxicyllin-clavulanate, is increasing. We present two cases that illustrate the potential severity of infections caused by multidrug resistant Salmonella and also the difficulty of reaching a differential diagnosis with inflammatory bowel disease. PMID- 10535207 TI - [Non-aneurysmatic aortic dysphagia]. AB - Esophageal compression by a vascular structure is a rare cause of dysphagia, the aberrant right subclavian artery being the most common congenital abnormality. Aortica dysphagia is usually observed in the elderly, especially in hypertensive women with cardiopathy and degenerative osteopathy. We report a 73-year-old woman with dysphagia, caused by a non-aneurysmatic aortic elongation, who presented progressive dysphagia, which ended in aphagia associated with heart failure. The diagnostic approach to these patients is discussed. The patient received cinitapride and, following treatment for heart failure, remains asymptomatic after a 3-year follow-up period, although manometric alterations persist. PMID- 10535208 TI - [Hepatic hemorrhagic infarction in eclampsia and HELLP Syndrome associated with the antiphospholipid syndrome]. AB - A 33 year-old woman developed eclampsia with HELLP syndrome. Laboratory results revealed lupus anticoagulant and anticardiolipin antibodies. Imaging tests showed liver and spleen infarctions. The patients was given enoxaparin and supportive care and there was a good evolution. We discuss some aspects about liver infarction and its association with toxemia of pregnancy and the antiphospholipid syndrome. PMID- 10535209 TI - [Eosinophilic enteritis causing intestinal obstruction]. AB - Eosinophilic enteritis is an uncommon disorder of unknown etiology in which the digestive symptoms are associated with eosinophilic infiltration of the different layers of the intestinal wall. Clinical symptoms depend on the layers involved and are usually characterized by peripheral eosinophilia. Radiological findings depend on the layers involved. Definitive diagnosis is based on clinical and histopathological findings. Treatment of choice is currently with corticoids and prognosis is benign with relapses. The pathogenesis remains unclear. We report a patient who presented with intestinal obstruction and describe the follow-up. PMID- 10535211 TI - [Cost effectiveness of the eradication of Helicobacter pylori in dyspepsia]. PMID- 10535210 TI - [Paracentesis in the treatment of ascites in cirrhotic patients]. PMID- 10535212 TI - [Usefulness of echography in the diagnosis of biliary ileus]. PMID- 10535213 TI - [Hemoperitoneum as a complication of diagnostic colonoscopy]. PMID- 10535215 TI - [The Freiburg Public Health Congress]. PMID- 10535214 TI - [Gastric schwannoma: the importance of immunohistochemistry for the differential diagnosis with other mesenchymal tumors]. PMID- 10535217 TI - [A different approach--a new instrument for expert assessment of disability stages?]. AB - A New Tool for Expertising Nursing-Care Need? Since the introduction of the long term care insurance in April 1995 criticism of the mode of expertise used to assign a particular nursing status to a patient has not ceased. Nursing status is required to receive monetary benefit from the long-term care insurance. According to the code of social law (SGB XI) nursing status is based on the total amount of time in minutes a care-needing person requires help to perform certain activities of daily living (ADL). This time-based classification is the bone of contention. As the currently used classification instrument has not been validated and is not reliable, it has been the aim of the present study to introduce a more objective mode of expertise. In our study we used the Barthel-Index as an instrument that allows to assign nursing status to a person in agreement with the person's respective ability to fulfil ADL. In a prospective study 252 patients were examined by the raters of the MDK Bavaria (Medical Service of Health Insurance) according to the SGB XI and the Barthel-Index. On the basis of the Barthel-Index and a certain discriminatory function more than 86% of the patients were assigned the same nursing status as with the normally used mode of expertise. Most of the differing results were borderline cases between two levels of nursing status. The new mode of expertising results in a more balanced distribution of different levels of nursing status. Therefore, the Barthel-Index in combination with the discriminatory function is as effective as the normally used mode of expertising. It is a valid and reliable instrument which could well replace the mode of expertising used so far. PMID- 10535218 TI - [Development of information, counseling and treatment status for patients with hypertension from 1984-91 in West Germany]. AB - Data from three nationally representative cross-sectional health surveys from 1984-86 (T0, n = 4790), 1987-88 (T1, n = 5335), and 1990-91 (T2, n = 5311) were analysed to determine changes in the utilisation of ambulatory services, degree of monitoring, risk factor awareness, drug treatment, and control, and medical counselling of hypertensive patients (actual hypertension prevalence) in West Germany. The percentage of hypertensives who had consulted a physician within the last year increased from 87% (T0) to 92% (T2) in men (p < 0.01), and from 94% to 96% in women (p > 0.05). The proportion of male (female) hypertensive patients who had their blood pressure measured by a physician within the last year was 87% (92%) in T2 (p-value for seven-year changes > 0.05). At T2, 50% (58%) of all male (female) hypertensives, whose blood pressure had been measured, were aware of their hypertension, with no significant changes during the study period. At T2, male (female) hypertensives in contact with a physician reported on medical recommendation of more physical activity in 16% (14%), reduced salt intake in 21% (25%), and reduced smoking/smoking cessation (only actual smokers) in 35% (31%) of cases. Counselling improved significantly only in hypertensive women. Although females had their hypertension more frequently treated with antihypertensive drugs (T0: 53%, T1: 53%, T2: 55%, p > 0.05), the increase of pharmaceutical treatment was stronger in men (T0: 34%, T1: 42%, T2: 45%, p < 0.01). Of those hypertensives treated with antihypertensive drugs, 55% (men) and 50% (women) had their blood pressure controlled below the 160/95 mmHg threshold in T2 with no significant changes during the study period. Suggestions are made for improving risk-factor awareness, and a higher quality and better integration of pharmaceutical treatment and non-pharmaceutical counselling of hypertensive patients. PMID- 10535216 TI - [Managed care--a possibility for German Health Insurance?]. AB - Health services systems in nearly all developed countries face similar problems. This fact raises the question whether concepts used in different countries can be changed without affecting the historically grown foundations of national health services as have been accepted by majority. A group of experts of the German Medical Services of the Statutory Health Insurance were asked to analyse whether the managed care approach could play a substantial role in reforming the German "Bismarck model", given that the advisory responsibility of the social medical service for the sick funds is respected in such a consideration and that basic essentials of the system are maintained. The group concludes, in brief, that managed care and the "German model" are contradictory in respect of preconditions, aims and assumed results. Furthermore, the experts share the view that the "German model" incorporates sufficient to cope with its problems without changing the nature of health services, based on principles of solidarity in Germany. The system needs structural reforms rather than changes in monetary mechanisms. PMID- 10535219 TI - [Epidemiologic and serologic studies of pneumococcal infections with reference to the new STIKO recommendations]. AB - Pneumococcal diseases play an important role especially for babies and toddlers (otitis media) and for older persons (pneumonia). 28% of the 481 reported cases of bacterial meningitides (without meningococcus) in Mecklenburg-Vorpommern were caused by pneumoniae streptococcus. A pneumococcus antibody study by the land register confirms the high contamination in older people. Therefore STIKO recommends since March 1998 to effect pneumococcal vaccination with every person from the age of 60 and dove as well as for children, adolescents and adults with higher risk due to a primary disease. PMID- 10535221 TI - [Mendel-Mantoux tuberculin test with GT 1 in the public health office]. AB - Within the German Public Health Service, the recommendation of the German Central Committee fur Combatting Tuberculosis to use the intracutaneous tuberculin test according to Mendel-Mantoux as a standard test has led to demands for additional investigations on sensitivity and specificity of different tuberculin doses. In routine everyday work of the Public Health Office of the German provincial capital Stuttgart the Mendel-mantoux tuberculin test with tuberculin GT 1 Behring was examined in respect of sensitivity and incidence of excessive reactions and compared to a former study with GT 5. 2,592 persons for whom the tuberculin test was prescribed according to the German infectious diseases law were tested for tuberculosis by the Stuttgart Public Health Office between May and October 1998, using the Mendel-Mantoux test with GT 1 Behring. Of these, 202 persons did not return for checking the result. The 2390 checked persons were 28.4 years old on the average, the majority being female (59.0% against 41.0% males) and of German nationality (59.6% against 40.1% other nationals). A positive reaction was seen in 16.2% (induration > or = 6 mm), in 7.2% Germans against 29.6% other nationals. In 51% of the positive results the indurations had a diameter of more than 10 mm, in 12.6% more than 15 mm, 9 probands, i.e. 0.4% of all tested persons, had an excessive reaction (formation of blisters with local necrosis). Especially among the other nationals, the share of strongly positive results (> 15 mm) did not decrease to the same extent as did the share of positive results as a whole. 94 persons belonging to the surrounding of a bacillary index case got a second test with GT 10. Of these, 30 (31.9%) had a positive reaction. The use of a low tuberculin concentration (GT 1) for the intracutaneous test according to Mendel Mantoux cannot be recommended. The sensitivity is significantly lower than with GT 5 which leads to seeming tuberculin conversions (because of the low sensitivity of GT 1) without avoiding excessive reactions. For tuberculin surveys in areas adjacent to bacillary index cases, the intracutaneous test according to Mendel-Mantoux should be performed with the usual dose (GT 10) as a standard test. For other purposes, the multipuncture-stamp test may be sufficient. PMID- 10535220 TI - [Surgical gloves--how well do the protect against infections?]. AB - Health care workers (HCW) in surgery are at high risk for bloodborne infections (BBI) e.g. by hepatitis-B(HB)-, hepatitis-C(HC)- and HI-virus. On the other hand, infectious medical staff can cause nosocomial BBI in patients, too. Intact gloves provide an efficient barrier against BBI but glove perforations are common during several surgical procedures. A review of studies on glove perforations during different surgical operations was carried out with special regard to user, number and location of perforations, duration and kind of operation. We compared the results with the frequency of glove perforation during operations in 1938 single used gloves. They were collected after different surgical procedures in a department for general surgery and tested for perforation by 1-liter-water filling method according to DIN 455/1. The product used during the period of this investigation was Sempermed sterile latex surgical glove. Most perforations were found on the index finger and thumb of the non-dominant hand. Duration of operation, role of the user (primary surgeon) and kind of operation are predictors for the incidence of glove perforations. Double gloving, endoscopic and no-touch techniques decrease the possibility of blood contact during operation. Indicator systems are useful for detection of the loss of glove integrity. Due to the high perforation rate found in this study glove change as a routine during surgical procedures should be discussed (e.g. every 30 minutes). PMID- 10535222 TI - [Heroin-assisted treatment of opiate addicts--former and current research emphasis]. AB - Heroin-assisted treatment has been examined broadly in Switzerland since 1994 within the context of scientific studies. The goal was to clarify the suitability of this treatment for opiate addicts whom previous therapy had failed to reach. Results of the initial research phase show that the target group could be reached for treatment extending 18 months with a satisfactory retention rate of 69%. The patients could improve their health and social situation during treatment and reduce illegal consumption of narcotics. Studies during the initial years primarily examined the viability of heroin-assisted treatment and its effects on the patients' psychosocial and somatic development. A second study phase ongoing since 1998 pursues the specific importance of medical and psychosocial treatment for patients' health and social development in heroin-assisted treatment. The focal point is the effort to optimise treatment of patients with comorbidity of psychiatric disorders and severe somatic diseases, particularly AIDS. Investigations carried out in Switzerland have been discussed broadly at an international level. Studies on heroin-assisted treatment are also being conducted at present in various countries. In future, co-operation should be further intensified with researchers on an international scale. PMID- 10535223 TI - [Multimedia learning program for medical consultation]. AB - To improve communicative competence during physicians' consultations, we developed a computer-assisted multimedia learning programme. This programme can be used in medical education as well as in continuing education for physicians. Dependent on individual skills and needs, the system, based on hypertext links, can be tailored to expand and improve communicative competence in a series of steps. The programme is based on a manual of medical interviewing and breaking bad news. The user gains information about theory and techniques of verbal intervention. He is trained to be able to reflectively observe and interactively intervene in video-taped and transcribed physician-patient dialogues. In a contrast-typological approach to model learning, in which negative and positive cases can be compared in a problem-based concept, alternative types of interventions can be recognised and evaluated. Finally, the learning goals in verbal intervention techniques should be subject of self-evaluation which can be objectified by further evaluations. PMID- 10535224 TI - [Some comments on the 2nd European drinking water guideline]. AB - It took 18 years before issuing a revised version of the first EU Drinking Water Guideline. As is well known, it did not receive unanimous acclaim neither by the water supply nor by the public health authorities. The second guideline has now been released and can be welcomed as a quite logically constructed and consistent version in comparison to its predecessor. Borderline values are stated only for those microbiological and chemical ingredients of water that are relevant to health. These borderline values must be considered as minimum deadlines and may not be raised by a member country although their severity may be increased. Basing on recent toxicological findings some of the borderline values have been raised or lowered compared to the previous version. On the other hand, however, the aesthetic aspects of drinking water have been neglected. In this respect we may look forward with interest as to how German legislation will implement the new guideline. This will have to be done at the latest by 2000 A.D. end. PMID- 10535225 TI - [Comparison of European standards of swimming pool safety]. AB - European countries follow different procedures to monitor the quality of the water of swimming pools open to the public: some use generally accepted technical standards, some officially recommended guidelines, some rules or regulations by local authorities and some have national laws or regulations. These agree insofar as in every country the water of swimming pools must be disinfected. The microbiological limits are to a certain extent identical with drinking water threshold values, e.g., no faecal coliforms must be present in 100 ml. Other microorganisms as for instance legionellae, staphylococci or Pseudomonas aeruginosa ar not uniformly considered. In all countries the water must be disinfected by chlorination. Big differences exist, however, in respect of the maximum admissible concentrations of free or total chlorine, haloforms and organic carbons. With regard to the common goals, harmonization of the regulations appears possible. PMID- 10535226 TI - [Bone and Joint Decade 2000-2010--prevention and treatment of diseases of skeletal muscles]. PMID- 10535227 TI - [Effects of globalization of structure and function of public health service in Germany]. AB - This essay discusses the effects of globalization on the German health care system. Globalization is regarded as a process to a "One-World-Society" without limits or borders to communication. Especially the economic effects of globalization are discussed, and the reduction of working places and wage levels in developed industrial societies, caused by globalization. Considering this development, it will not be possible to uphold present health care standards. PMID- 10535228 TI - [Developing a small-area cancer atlas: process, validity and possible applications]. PMID- 10535229 TI - Assessing a public health approach to delay onset and progression of adolescent substance use: latent transition and log-linear analyses of longitudinal family preventive intervention outcomes. AB - This study examined the effects of the Iowa Strengthening Families Program (ISFP) and the Preparing for the Drug-Free Years program (PDFY) on young adolescent transitions from nonuse of substances to initiation and progression of substance use. Analyses incorporated 3 waves of data collected over a 2.5-year period from 329 rural young adolescents. Outcomes were analyzed by using log-linear models that incorporated substance use status frequencies derived from latent transition analyses. Effects on delayed substance use initiation were shown for both the ISFP and the PDFY at a 2-year follow-up. Also at this follow-up, the PDFY showed effects on delayed progression of use among those previously reporting initiation. PMID- 10535232 TI - Expressed emotion and behavior therapy outcome: a prospective study with obsessive-compulsive and agoraphobic outpatients. AB - The relationship of expressed emotion (EE) to behavior therapy outcome for obsessive-compulsive disorder (n = 60) and panic disorder with agoraphobia (n = 41) was investigated. Relatives' emotional overinvolvement and hostility predicted higher rates of treatment dropout. Higher hostility, as assessed by the Camberwell Family Interview (CFI), was related to poorer outcome for target ratings and for the Social Adjustment Scale; higher perceived criticism was also predictive of worse response on target ratings. In contrast, nonhostile criticism on the CFI was associated with better outcome on the behavioral avoidance test. In general, the relationship of EE to outcome was not moderated by type of relative, diagnosis, amount of contact with the relative, or use of psychotropic medication. PMID- 10535230 TI - Initial impact of the Fast Track prevention trial for conduct problems: I. The high-risk sample. Conduct Problems Prevention Research Group. AB - Fast Track is a multisite, multicomponent preventive intervention for young children at high risk for long-term antisocial behavior. Based on a comprehensive developmental model, intervention included a universal-level classroom program plus social skills training, academic tutoring, parent training, and home visiting to improve competencies and reduce problems in a high-risk group of children selected in kindergarten. At the end of Grade 1, there were moderate positive effects on children's social, emotional, and academic skills; peer interactions and social status; and conduct problems and special-education use. Parents reported less physical discipline and greater parenting satisfaction/ease of parenting and engaged in more appropriate/consistent discipline, warmth/positive involvement, and involvement with the school. Evidence of differential intervention effects across child gender, race, site, and cohort was minimal. PMID- 10535231 TI - Initial impact of the Fast Track prevention trial for conduct problems: II. Classroom effects. Conduct Problems Prevention Research Group. AB - This study examined the effectiveness of the universal component of the Fast Track prevention model: the PATHS (Promoting Alternative THinking Strategies) curriculum and teacher consultation. This randomized clinical trial involved 198 intervention and 180 comparison classrooms from neighborhoods with greater than average crime in 4 U.S. locations. In the intervention schools, Grade 1 teachers delivered a 57-lesson social competence intervention focused on self-control, emotional awareness, peer relations, and problem solving. Findings indicated significant effects on peer ratings of aggression and hyperactive-disruptive behavior and observer ratings of classroom atmosphere. Quality of implementation predicted variation in assessments of classroom functioning. The results are discussed in terms of both the efficacy of universal, school-based prevention models and the need to examine comprehensive, multiyear programs. PMID- 10535233 TI - Demand-withdraw interaction in couples with a violent husband. AB - This study examined the relationship between demand-withdraw interaction and battering in couples with a violent husband. The authors compared the interaction patterns of 47 couples with a violent husband with the interaction patterns of 28 distressed but nonviolent couples and 16 happily married nonviolent couples. All couples engaged in videotaped discussions of problem areas in their marriage. Both batterers and battered women showed less positive communication and more negative communication than did their nonviolent counterparts. Additionally, batterers showed significantly higher levels of both demanding and withdrawing than did other men. Battered women demanded more change than did women in nonviolent marriages but were significantly less inclined to withdraw than were their husbands. The discussion of these findings focuses on the interactional dynamics between batterers and battered women and how these interactions might be understood. PMID- 10535235 TI - Engaging the unmotivated in treatment for alcohol problems: a comparison of three strategies for intervention through family members. AB - In a randomized clinical trial, 130 concerned significant others (CSOs) were offered 1 of 3 different counseling approaches: (a) an Al-Anon facilitation therapy designed to encourage involvement in the 12-step program, (b) a Johnson Institute intervention to prepare for a confrontational family meeting, or (c) a community reinforcement and family training (CRAFT) approach teaching behavior change skills to use at home. All were manual-guided, with 12 hr of contact. Follow-up interviews continued for 12 months, with 94% completed. The CRAFT approach was more effective in engaging initially unmotivated problem drinkers in treatment (64%) as compared with the more commonly practiced Al-Anon (13%) and Johnson interventions (30%). Two previously reported aspects of the Johnson intervention were replicated: that most CSOs decide not to go through with the family confrontation (70% in this study) and that among those who do, most (75%) succeed in getting the drinker into treatment. All 3 approaches were associated with similar improvement in CSO functioning and relationship quality. Overall treatment engagement rates were higher for CSOs who were parents than for spouses. On average, treatment engagement occurred after 4 to 6 sessions. PMID- 10535234 TI - Contingency management, self-control, and education support in the treatment of childhood phobic disorders: a randomized clinical trial. AB - This study evaluated the relative efficacy of an exposure-based contingency management (CM) treatment condition and an exposure-based cognitive self-control (SC) treatment condition relative to an education support (ES) control condition for treating children with phobic disorders. Eighty-one children and their parents completed a 10-week treatment program in which children and parents were seen in separate treatment sessions with the therapist, followed by a brief conjoint meeting. Children in both the CM and SC conditions showed substantial improvement on all of the outcome measures. These gains were maintained at 3-, 6 , and 12-month follow-ups. Interestingly, children in the ES condition also showed comparable improvements at posttreatment and at 3-, 6-, and 12-month follow-ups. Implications of the findings are discussed with respect to knowledge development and clinical practice. PMID- 10535236 TI - Predicting patients' responses to psychotherapy: are some more predictable than others? AB - Patient profiling predicts improvement across psychotherapy sessions on the basis of intake clinical characteristics. Patients completed questionnaires before their 1st session and at other points during treatment. After using hierarchical linear modeling to calculate expected courses of improvement, 1/2 of the sample was divided into 1 of 2 groups: those whose treatment responses across sessions matched or exceeded their expected courses (75% of the sample) and those whose responses failed to match expectations. A discriminant function analysis indicated that the groups could be differentiated on the basis of intake characteristics; that is, patients with higher discriminant scores were more likely than those with low discriminant scores to have responded to psychotherapy as predicted. Successful cross-validation of the predictor weights was performed with the other 1/2 of the sample. PMID- 10535237 TI - Risk recognition and trauma-related symptoms among sexually revictimized women. AB - This study used experimental methodology to investigate the differential impact of various levels of sexual victimization on women's perceptions of risk and evaluative judgments of sexual assault within a dating interaction. Single- and multiple-incident victims were compared with nonvictims. Results supported the hypothesis that revictimized women would exhibit longer latencies than either single-incident victims or nonvictims in signaling that an audiotaped date rape should be halted. Revictimized women with greater posttraumatic stress disorder (PTSD) symptoms, arousal symptoms in particular, exhibited latencies similar to those of nonvictims, whereas revictimized women with lower levels of PTSD symptoms had significantly longer latencies. Dissociative symptoms were not related to latency. These findings suggest that PTSD-related arousal symptoms may serve a buffering effect, increasing sensitivity to threat cues that portend a sexually coercive interaction. PMID- 10535238 TI - Parenting through change: an effective prevention program for single mothers. AB - This randomized experimental prevention study (a) evaluated the effectiveness of a parent-training program in a sample of 238 divorcing mothers with sons in Grades 1-3 and (b) provided an experimental test of coercion theory. The intervention produced reductions in observed coercive parenting, prevented decay in positive parenting, and generally improved effective parenting practices in comparisons of mothers in experimental and control groups. Moreover, coercion theory was supported. Improved parenting practices correlated significantly with improvements in teacher-reported school adjustment, child-reported maladjustment, and mother-reported maladjustment. The intervention indirectly benefitted child outcomes through improved parenting practices for a model based on child report and, to a lesser extent, on teacher report. The intervention did not produce direct effects on child outcomes. PMID- 10535239 TI - Effective treatment relationships for persons with serious psychiatric disorders: the importance of attachment states of mind. AB - Participants were 54 clients with serious psychiatric disorders and 21 clinical case managers. Clients' serious psychiatric disorders included Axis I diagnoses, such as schizophrenia and bipolar disorder. This study examined how attachment states of mind of both clients and case managers influenced the effectiveness of therapeutic relationships and client functioning. Client and case manager attachment states of mind interacted in predicting the working alliance and client functioning. Specifically, clients who were more deactivating with respect to attachment had better alliances and functioned better with less deactivating case managers, whereas clients who were less deactivating worked better with more deactivating case managers. These findings highlight the importance of clinicians and clients being matched in ways that balance their interpersonal and emotional strategies. PMID- 10535240 TI - The efficacy of cognitive-behavioral and interpersonal treatments for depression in Puerto Rican adolescents. AB - This study evaluated the efficacy of cognitive-behavioral therapy (CBT) and interpersonal psychotherapy (IPT) with depressed adolescents in Puerto Rico. Seventy-one adolescents meeting Diagnostic and Statistical Manual of Mental Disorders (3rd ed., rev.; American Psychiatric Association, 1987) criteria for a diagnosis of depression were randomly assigned to 1 of 3 conditions: CBT, IPT, or wait list (WL). Pretreatment, posttreatment, and 3-month follow-up measures of depression symptoms, self-esteem, social adjustment, family emotional involvement and criticism, and behavioral problems were completed. Results suggest that IPT and CBT significantly reduced depressive symptoms when compared with the WL condition. IPT was superior to the WL condition in increasing self-esteem and social adaptation. Clinical significance tests suggested that 82% of adolescents in IPT and 59% of those in CBT were functional after treatment. The results suggest that both IPT and CBT are efficacious treatments for depressed Puerto Rican adolescents. IPT's impact in other levels of outcome is discussed in terms of its consonance with Puerto Rican cultural values. PMID- 10535241 TI - Multilevel daily process designs for consulting and clinical psychology: a preface for the perplexed. AB - The next 3 articles in this issue use multilevel statistical procedures to analyze data collected in daily process studies of (a) stress and coping, (b) binge eating, and (c) chronic pain experience. Important differences in the methods and procedures of these studies illustrate the many options available to investigators and data analysts. This article serves as a preface to help readers who are new to these studies' methodology appreciate their novel contributions to the literature in consulting and clinical psychology. Four frequently asked questions are addressed concerning the design of daily process studies, the distinctive meaning of a within-person finding, the possibility that self monitoring studies are measurement reactive, and complexities in the use of multilevel statistical procedures for analyzing person-day data sets. PMID- 10535242 TI - Coping with daily events and short-term mood changes: an unexpected failure to observe effects of coping. AB - This study examined the relationship between coping efforts and stress-related mood changes. Men and women with high levels of work or marital stress reported stress and coping efforts approximately once an hour for 2 days using an electronic diary. Stress episodes were identified as a stress-free time followed by a stressor at the next time point. Analyses examined how appraisals and coping influenced pre- to poststress mood change and how problem appraisals were related to coping efforts. Greater mood changes were associated with appraisals of high stress and high disruptiveness. Appraisals of high control and high desirability were associated with more planning, direct action, and fewer acceptance coping efforts. Coping failed to predict any pre- to poststressor mood changes. Possible explanations for the overall failure of coping to predict momentary mood changes are discussed. PMID- 10535243 TI - Hypersensitivity to social interactions in bulimic syndromes: relationship to binge eating. AB - This study used a 6- to 22-day experience-sampling procedure to test for hypersensitivity to social interactions in bulimic individuals. Ratings on daily social interactions, self-concepts, moods, and eating behaviors from 55 actively bulimic, 18 formerly bulimic, and 31 noneating-disordered women were obtained. Hierarchical linear modeling analyses showed negative social interactions to be associated with significant increases in self-criticism (SC) and deteriorations in mood in all participants. However, bulimic participants (active or in remission) showed larger increases in SC following negative social interactions than did normal eaters (and thus seemed to be hypersensitive to interpersonal experiences). Additional analyses indicated that binge episodes tended to be preceded by poorer than average social experiences, self-concepts, and moods and followed by deteriorations in self-concept, mood, and social perception. The authors discuss possible perpetuating influences of interpersonal sensitivity in bulimic syndromes. PMID- 10535244 TI - Pain, negative mood, and perceived support in chronic pain patients: a daily diary study of people with reflex sympathetic dystrophy syndrome. AB - Chronic pain patients show substantial psychological distress, including depressed mood, anxiety, and anger. Nevertheless, the causal role of negative mood in the course of chronic pain conditions remains unclear. This study prospectively investigated the relationship between daily pain, negative mood, and social support in 109 people with reflex sympathetic dystrophy syndrome. Participants completed 28 daily diaries that included questions about pain, mood, and perceived support. Time-lagged within-subject analyses indicated that pain led to increases in depressed, anxious, and angry mood. Depressed mood, but not anxiety or anger, contributed to increases in pain. Perceived support had both main and buffering (interaction) effects on negative mood and a main effect on pain. PMID- 10535245 TI - Trauma exposure among children with oppositional defiant disorder and attention deficit-hyperactivity disorder. AB - Consecutive admissions to an outpatient child psychiatry clinic diagnosed with oppositional defiant disorder (ODD), attention deficit-hyperactivity disorder (ADHD), or adjustment disorder were assessed for trauma exposure by a structured clinical interview and parent report. Controlling for age, gender, severity of internalizing behavior problems, social competence, family psychopathology, and parent-child relationship quality (assessed by parent report), an ODD diagnosis, with or without comorbid ADHD, was associated with increased likelihood of prior victimization (but not nonvictimization) trauma. ADHD alone was not associated with an increased likelihood of a history of trauma exposure. Traumatic victimization contributed uniquely to the prediction of ODD but not ADHD diagnoses. Children in psychiatric treatment who are diagnosed with ODD, but not those diagnosed solely with ADHD, may particularly require evaluation and care for posttraumatic sequelae. PMID- 10535246 TI - Should patients' religiosity influence clinicians' referral to 12-step self-help groups? Evidence from a study of 3,018 male substance abuse patients. AB - Twelve-step self-help organizations maintain that anyone, regardless of his or her religious beliefs, can benefit from participation in their groups. Yet many addiction professionals have reservations about referring nonreligious patients to 12-step groups. The present study examined the influence of patients' religiosity on whether they were referred to and benefited from 12-step groups. Participants were 3,018 male substance abuse inpatients. Individuals who engaged in fewer religious behaviors in the past year were referred to 12-step groups less frequently by clinicians. However, referrals to 12-step groups were effective at increasing meeting attendance, irrespective of patients' religious background, and all experienced significantly better substance abuse outcomes when they participated in 12-step groups. The viewpoint that less religious patients are unlikely to attend or benefit from 12-step groups may therefore be overstated. PMID- 10535247 TI - Differentiation of malignant lymphoma and non-lymphoma by an occlusive dressing method employing bromodeoxyuridine. AB - Although lymphoma cells can proliferate in skin tissue, lymphocytes which have infiltrated the skin due to inflammatory changes are generally unable to do so. It may be possible to differentiate malignant lymphomas from benign lymphocytic infiltrations in skin tissue by detection of cell cycle-related antigens. We developed a novel in vivo method for studying the cell kinetics of human skin using bromodeoxyuridine (BrdU) with an occlusive dressing. Following the application of BrdU, BrdU-labeled lymphoid cells were counted in the dermis. The patients studied included 22 with various types of lymphomas (lymphoma group) and 22 with different forms of inflammatory skin disease (non-lymphoma group). Skin specimens were obtained after applying occlusive adhesive plasters containing 2% BrdU dissolved in 0.9% sodium chloride solution and were then immunostained with anti-BrdU monoclonal antibody. The average percentages of labeled cells were 15.9 +/- 7.6% for the lymphoma group and 4.7 +/- 2.6% for the non-lymphoma group. This difference was statistically significant (p < 0.0001). Using this method, malignant lymphomas and benign lymphocytic infiltrations of the skin could be differentiated by counting BrdU-labeled lymphoid cells, and this method could help in determining a prognosis. PMID- 10535248 TI - Malassezia spp carriage in patients with seborrheic dermatitis. AB - The role of Malassezia spp in seborrheic dermatitis (SD) is controversial. To compare the cutaneous density and the cultural characteristics of Malassezia in persons with or without SD, quantitative cultures were obtained by stripping the forehead with a tape placed on Leeming and Notman medium. Plates were incubated at 37 degrees C in a plastic bag, and colonies were counted after 14 days. High yeast density was defined as > 100 colony forming units (CFU)/tape. Volunteers were divided into four groups depending on their HIV serology [HIV (+) versus HIV (-) or unknown] and their clinical status (with or without SD). 126/129 cultures were positive (97.7%). Malassezia spp density was low on clinically normal skin in all HIV (-) persons (40/40) but was high in 8/34 (24%) HIV (+) persons without SD (p < 0.001). In SD patients, high densities were found in 10/22 (45%) HIV (+) and in 17/33 (52%) HIV (-) persons. The strains could be divided into three basic groups on the basis of their cultural characteristics. Colony morphology type A predominated on normal skin (72%), and morphology type C predominated on persons with SD (78%). High yeast density can be present without skin symptoms. The pathogenicity of Malassezia seems more likely to be determined by the subtype present on the skin rather than by its density. PMID- 10535249 TI - Twice weekly 5 mg dexamethasone oral pulse in the treatment of extensive alopecia areata. AB - Thirty patients (20 males, 10 females) with widespread alopecia areata (25 extensive alopecia areata, 5 alopecia areata) for a mean period of 4.2 years were included in the study. All patients above 12 years were administered 5 mg dexamethasone oral pulse on two consecutive days every week. Three children (< 12 years) received 2.5 mg to 3.5 mg dexamethasone oral biweekly pulse. Patients who had received treatment for a minimum period of 12 weeks were evaluated for terminal hair growth. Complete to excellent (75-95%) hair growth was observed in 16 (63.3%) patients. Growth was good (50-74%) in 2 cases and poor (< 50%) in 3 (10%) cases. Six (20%) patients has no growth of terminal hair. Complete to excellent growth of hair was obtained after a mean period of 5.35 months (range 3 10 months). Relapse occurred in one case each after three and six months but hair regrew with re-treatment. Side effects of corticosteriods were frequent, seen in 8 (26.6%) patients, but were mild. In only one case, treatment had to be discontinued. We propose that twice weekly 5 mg dexamethasone oral pulse for six months may be considered as one of the modalities in the treatment of extensive long standing alopecia areata. PMID- 10535250 TI - Neck location of basal cell carcinomas: a new-significant variant? AB - Basal cell carcinoma of the neck has been suggested to have a distinct clinical, histological and prognostic significance. The aim of our study is to verify this assumption. Among 1,185 basal cell carcinomas, we have observed 42 tumors localized on the neck. The mean age did not differ from that of patients with basal cell carcinomas on other sites, and there was no difference in incidence between sexes. The frequence of recurrence was 4.76%, similar to the recurrence rate of basal cell carcinomas on other sites. The lesions were typed histologically according to accepted criteria. The patterns observed were: nodular (62.8%), superficial (21.1%), mixed (4.7%), micronodular (5.9%), and infiltrative (5.7%). These frequencies did not differ significantly from those observed in the basal cell carcinomas on other sites. Thus, in comparison with the overall basal cell carcinoma population, the tumors of the neck do not represent a particular clinical and histologic subtype and do not have a particular recurrence potential. We think that basal cell carcinomas localized on the neck do not represent a particular subtype of these tumors; their treatment can be the same as that of basal cell carcinomas on other regions. PMID- 10535251 TI - Macular lesions in leprosy: a clinical, bacteriological and histopathological study. AB - Among 150 untreated patients of leprosy, 19 had only macular lesions; three were of the indeterminate type, and eight each were of the tuberculoid and the borderline types, according to the Indian Association of Leprologists (IAL, 1981) classification. The clinical, bacteriological, and histopathological parameters of these 19 patients were studied both before and after six months of WHO Multi Drug Therapy (MDT/1982). A single macule was present in seven (36.84%) patients. In twelve (63.16%), two or more were seen. In eighteen (94.74%), one or more peripheral nerves were enlarged. The size of the macules varied from 5 to 15 cm, and there were no changes seen even after treatment. In most (94.74%) of the patients, the macules were hypopigmented. The surfaces were rough and dry in seven (36.84%) but smooth in the other twelve (63.16%). The margins were well defined in the seven (36.84%) patients with single macules but ill defined in the other twelve (63.16%). After six months of antileprosy treatment, the single macules showed some resolution. Slit skin smear examination was negative in all cases before and after treatment. Clinico-histopathological correlations were seen in only six (31.58%) patients; the clinical diagnoses were indeterminate and tuberculoid leprosy in three (15.79%) patients each. In the indeterminate group, the clinico-histopathological correlation was 100%; it was 37.50% in the tuberculoid group. There were no correlations between the clinical and histopathological parameters in thirteen (68.42%) cases. After six months of treatment, the histopathology became nonspecific in all patients. The lepromin test was positive in six (31.58%) patients; four were of the tuberculoid group and one each from the indeterminate and borderline leprosy groups. Hence, although macular lesions can be seen throughout the leprosy spectrum, it is difficult to correlate their clinical, bacteriological and histopathological parameters. PMID- 10535252 TI - Mast cell "densities" in vascular proliferations: a preliminary study of pyogenic granuloma, portwine stain, cavernous hemangioma, cherry angioma, Kaposi's sarcoma, and malignant hemangioendothelioma. AB - The "densities" of mast cells (MCs) in six kinds of vascular proliferation, pyogenic granuloma, portwine stain, cavernous hemangioma, cherry angioma, Kaposi's sarcoma, and malignant hemangioendothelioma (MHE), measured per mm2 were studied using respective specimens prepared with tryptase stain and a personal computer. The average densities of MCs in pyogenic granuloma and MHE were 103.5 +/- 25.2/mm2 (n = 10) and 106.3 +/- 40.2/mm2 (n = 10) [mean +/- standard deviation (SD)]; that in normal skin was 6.85 +/- 4.9/mm2 (n = 20) (mean +/- SD). is a significant difference [t-test (p < 0.0001) and Wilcoxon-test (p < 0.01)]. The results in portwine stain (n = 4), cavernous hemangioma (n = 9), cherry angioma (n = 4), and Kaposi's sarcoma (n = 4) were 68.6 +/- 28.9/mm2, 105.7 +/- 56.9/mm2, 85.3 +/- 45.6/mm2, 82.2 +/- 28.4/mm2 (mean +/- SD), respectively, all of which were greater than that in normal skin by a simple comparison. The results of immunofluorescence microscopy were positive with basic fibroblast growth factor staining in the tissues of pyogenic granuloma, Kaposi's sarcoma and MHE. These facts may morphologically indicate a role of MCs in the angiogenesis of these vascular tumors. PMID- 10535253 TI - Metastatic cutaneous plasmacytoma: a case report associated with IgA lambda multiple myeloma and a review of the literature of metastatic cutaneous plasmacytomas associated with multiple myeloma and primary cutaneous plasmacytomas. AB - We present the case of a 67-year-old Japanese woman with immunoglobulin A lambda (IgA lambda) multiple myeloma (MM). She had firm nodular cutaneous lesions on the trunk and scalp without adjacent bone involvement. The patient was diagnosed as having IgA lambda MM of stage IIIA with 52% plasmacytosis in the bone marrow six months before the appearance of the cutaneous lesions. The abnormal plasma cells showed moderate to marked dysplasia in both the bone marrow and skin lesions. The abnormal plasma cells in the bone marrow exhibited abnormal karyotypes: 41, XX, der (1) t (1p; 1q), -4, -10, -14, -16, -17, 17p+, that differed from the "unfavorable" karyotype reported previously. We reviewed the cases of metastatic cutaneous plasmacytoma in MM and cases of primary cutaneous plasmacytoma that have been reported in English or Japanese and identified the Ig class. Among the 83 cases of metastatic cutaneous plasmacytomas in MM, IgG, IgA, IgD, and Bence Jones protein were found in 52%, 23%, 16%, and 6%, respectively. A disproportionately high frequency of IgD lambda MM was found to have spread to the skin, compared with the frequency of IgD MM itself, which was present in only around 2% of the MM cases. Among the 18 primary cutaneous plasmacytomas, IgG, IgA, and Bence-Jones protein were found in 56%, 11%, and 17%, respectively, but no IgD was found. PMID- 10535254 TI - Infantile myofibromatosis: a case study and review of literature. AB - Infantile myofibromatosis is an unusual condition generally presenting in the newborn period. The case being reported is that of a female newborn who had multiple lesions that involved skin, subcutaneous tissue, skeletal muscles, bone, and lungs. The disease was diagnosed because of the easily palpable skin tumors and subcutaneous nodules that were obvious immediately after birth. The diagnosis was established by histopathological examination of one nodule that showed a spindle-celled mesenchymatogenic lesion demonstrating the morphological and immuno-phenotype characteristics of myofibroblastic differentiation. The histologic picture, combined with the clinical manifestations and the imaging findings, are consistent with infantile myofibromatosis. The physical condition of the newborn was excellent and remains so six months later. The tumors of the skin and the subcutaneous nodules have gradually regressed without therapy. At the age of six months, four (4) nodules are palpable; the infant is under continuous observation. PMID- 10535255 TI - Adult onset of inflammatory linear verrucous epidermal nevus. AB - Adult onset of inflammatory linear verrucous epidermal nevus (ILVEN) is reported in a 44-year-old Japanese man. A mild pruritic eruption appeared one year earlier and extended from the left dorsal foot to the gluteal region. Histologically, acanthosis and papillomatous thickening of epidermis as well as spongiotic edema and exocytosis with lymphocytes and neutrophils were observed. Topical tacalcitol was not effective, but the pruritus as well as the eruption slightly improved with topical corticosteroid and vaseline containing salicylic acid. This adult onset of ILVEN is considered to be a rare case. PMID- 10535256 TI - A case of lipodystrophia centrifugalis abdominalis infantilis with morphea. AB - A 5-year-old female developed three depressed lesions sequentially on her left hip, left upper arm and left Achilles tendon; we diagnosed them as lipodystrophia centrifugalis abdominalis infantilis (LCAI). She also developed two sclerotic lesions, at almost the same time, on her left upper arm and left forearm; we diagnosed them as morphea. Clinical and histopathological examination revealed that the LCAI and morphea seen in our case were completely different. Although two other cases of co-existing LCAI and morphea have already been reported, they were not described in detail and their morphea lesions were adjacent to their LCAI lesions. We herein report a case of LCAI with morphea in which the lesions were not adjacent. PMID- 10535257 TI - Panniculitis showing membranocystic changes in the dermatomyositis. AB - Clinically obvious panniculitis in association with dermatomyositis is rare. We reported a patient with panniculitis showing membranocystic changes in the dermatomyositis. PMID- 10535259 TI - Idiopathic acquired anhidrosis: reversible generalized anhidrosis after sunstroke. AB - Idiopathic acquired generalized anhidrosis is a very rare condition in which the pathogenesis is still unknown. Although varied findings have been reported, no consistent abnormal findings in sweat glands have been established by histological study or electronmicroscopic study. An 18-year-old Korean boy suffered from generalized anhidrosis after sunstroke. Physical and neurologic examination could not reveal any abnormal findings. There were no specific changes in the sweat glands, but amorphous vacuole-like structures were seen in the cytoplasm of secretary cells. We reviewed the previous reports about this rare condition and compared them with our patient. PMID- 10535258 TI - A case of systemic sclerosis associated with gastric cancer. AB - We report the case of a 47-year-old woman who began to experience stiffness and Raynaud's phenomenon of her fingers and toes as well as easy fatigability approximately one year before initial evaluation. Histopathologic examination revealed dense fibrosis and perivascular lymphoid cell infiltration in the dermis. The patient's diagnosis was systemic sclerosis (SSc). Esophageal sclerosis was not revealed, but an early type IIb carcinoma of the gastric antrum was noted by endoscopic examination. Other recent reports have discussed the association of SSc with various malignant carcinomas. We also reviewed the literature for associations between SSc and gastric cancer in Japanese patients. PMID- 10535260 TI - Sebaceous hyperplasia in youngsters. PMID- 10535261 TI - Letter to the editor regarding the article: "Erythema dyschromicum perstans in early childhood" by Sang-Ju Lee and Kee-Yang Chung. PMID- 10535262 TI - [Effect of complement on the production of tumor necrosis factor alpha by human leukocytes with culture supernatants of clinical isolates of Staphylococcus aureus]. AB - Induction of tumor necrosis factor alpha (TNF-alpha) of human leukocytes by overnight culture supernatants of Staphylococcus aureus (S. aureus) (CS) (4 strains) and then the effect of the complement on production of TNF alpha by the treated cells were examined. One-tenth diluted solution of CS was added to Heparinized blood and was incubated for 6 hrs at 37 C in a 5% CO-air humidified incubator. TNF alpha level in the plasma was measured by ELISA. The level in the plasma was different in different CS. The induction of TNF alpha was not found in EDTA-added blood but was found in the EGTA-added them treated with CS, though different levels were showed. Western blotting assay of these plasma samples without EDTA-added blood found the presence of both C5a and C3a. Neither C5a and C3a was found in Heparinized blood treated with a relative concentration to TSST 1, enterotoxin C and alpha (alpha) homolysin contained in CS. Mixture of these toxins induced only 1/6.5 of the TNF alpha amount obtained with the CS. Human leukocytes isolated by Mono.-Poly. resolving solution were cultured with CS in RPMI 1640 medium with or without 10% fresh or heated serum. Production of TNF alpha by the isolated leukocytes was not found under the condition of serum free. But it was found in the presence of fresh serum and also in the presence of heated serum. The addition of CS to RPMI 1640 containing heated serum did not occur naturally in the production of C5a. These results suggest that there is a difference in TNF alpha inductivility to human leukocytes in each strain and that other bacterial components without TSST-1, SEC and alpha-hemolysin are more important to induce TNF alpha. Serum may be necessary to produce TNF alpha by CS treated cells as a source of factor(s) to interact with bacterial components rather than a source of complement. PMID- 10535263 TI - [Clinical analysis of pneumonia in the elderly in a community hospital- comparison of community-acquired pneumonia and nosocomial pneumonia]. AB - We experienced 530 elderly cases with pneumonia among 930 patients with pneumonia in Kawasaki Medical School Kawasaki Hospital between April 1986 and September 1998. Clinical analysis of all these patients and a comparison of one group consisting of 418 patients with community-acquired pneumonia and another group composed of 112 patients with nosocomial pneumonia were performed. In all of the elderly patients with pneumonia, respiratory symptoms and inflammatory findings were less frequent, but were frequent for those in poor general and nutritional condition. The causative microorganism was isolated in 42% of these patients. Streptococcus pneumoniae, MSSA and Klebsiella pneumoniae were frequently isolated from the sputum of the patients with community-acquired pneumonia, while Methicillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa, Methicillin-sensitive Staphylococcus aureus (MSSA) were frequently isolated from that of nosocomial pneumonia patients. Mycoplasma pneumoniae, Chlamydia pneumoniae and some viruses were less frequent for patients in both groups. Although many intravenous antibiotics, such as cephem or carbapenem were administered to patients in both groups, the prognosis was relatively good for those with community acquired pneumonia but was extremely poor for those with nosocomial pneumonia despite mechanical ventilation or steroid pulse therapy for many patients. PMID- 10535264 TI - [A new exanthematous disease in newborn infants caused by exotoxins producing Staphylococcus aureus; exotoxins production of the isolates and serum levels of antitoxin antibody in the patients and umbilical cord blood]. AB - Recently, in Japan newly neonatal exanthematous disease was elucidated to be caused by staphyloccocal supcrantigcnic exotoxins, mainly TSST-1. We studied exotoxins producibility of 43 strains of S. aureus isolated from neonates with exanthematous disease and examined antibody titers to staphylococcal enterotoxin A, B, C (SEA, SEB, SEC) and toxic shock syndrome toxin 1 (TSST-1) of the patients and control (umbilical cord blood from term infants). The results were as follows 1.34 of 43 strains (79%) isolated from the patients were SEC and TSST-1 producing MRSA, 5 strains (12%) were SEB, SEC, and TSST-1 producing MRSA, 1 strain (2%) was SEB and TSST-1 producing MRSA, 2 strains (12%) were SEB producing MSSA and did not produce TSST-1. The 1 strain (2%) was MSSA which produced SEC and TSST-1. 2. 16 neonates with exanthematous disease, who showed typical clinical signs and laboratry findings of thrombocytopenia, with SEC and TSST-1 producing MRSA isolates had significantly low anti-TSST-1 antibody titers at onset (p < 0.05), compared with the control (umbilical cord blood from term infants): TSST-1 appeared to the causative agent for the disease. In two neonates with exanthematous disease, with SEB- and non- TSST-1-producing MSSA isolates, anti SEB antibody titers were low at onset, so SEB appeared to the causative agent for the disease. 3. In Japan, low anti-TSST-1 antibody titers were found in the umbilical blood samples from about 70% of term infants; and low anti-SEB or anti SEC antibody titers were found in samples from only about 10% of them, that is, a number of term infants had anti-SEB and anti-SEC antibodies. The majority of S. aureus isolated from neonates with exanthematous disease were enterotoxin- and TSST-1-producing MRSAs. The results of our study by measuring antitoxin antibody titers suggested that SEB and SEC might not be pathogenically responsible, but TSST-1 was considered to be responsible for the majority of exanthematous disease. Prevalence of TSST-1-producing MRSA in the neonatal and premature baby ward is the main cause for the high incidence of this disease in Japan, whereas the low antibody titer to TSST-1 in the mother, in comparison with the anti enterotoxin antibody titers, may also be a predisposing factor. PMID- 10535265 TI - [Clinical analysis of patients with bacterial meningitis in childhood and reevaluation of rapid antigen detection methods]. AB - Twenty-eight cases of bacterial meningitis during the recent ten years were analyzed retrospectively, and the following results were obtained. 1. Pathogens were as follows; H. influenzae 13 (46.4%), S. pneumoniae 8 (28.6%), S. agalactiae 4 (14.3%), E. coli 2 (7.1%), and L. monocytogenes 1 case (3.6%). 2. Twelve out of the thirteen H. influenzae cases were caused by serotype b (Hib), and 2 strains were beta-lactamase producer. Fifty percent of the S. pneumoniae cases were caused by penicillin-resistant strains. And all these resistant strains belonged to serotype 19 or 23. 3. Underlying diseases related to the onset of meningitis were found in 46% of the cases, and these consisted of CNS shunt operated 5, asplenia or polysplenia 2, Mondini's anomaly 1, sacral dermal sinus 1, and neonate 4 cases. 4. Prognosis of these cases were three deaths, four with neurologic sequelae, and twenty-one complete recoveries. 5. On admission, 85% (17/20) of the cases were diagnosed correctly by the rapid antigen detection. Sensitivity and specificity of the rapid antigen detection by using latex particle agglutination is 90% and 100% in the Hib cases, and 83% and 100% in the S. pneumoniae cases respectively. Moreover, the bacteriologically unknown 2 cases caused by parenteral partial treatment were also diagnosed by the detection of antigen in concentrated urine. PMID- 10535266 TI - [Usefulness of MT-2 assay to decide when to start anti-HIV therapy]. AB - It is well known that the emergence of syncytium inducing (SI) variant correlates accelerated CD4 decline and disease progression during the course of HIV infection. We have conducted a clinical study to investigate whether or not detection of SI by MT-2 cells (MT-2 assay) can be a clinical marker to decide when to start anti-HIV therapy since 1995. We examined 483 blood samples obtained from 172 HIV-infected patients by the MT-2 assay. SI was detected from 20 patients. There were five untreated patients whose CD4 counts were 300/microliter or more. Anti-HIV combination therapies were started soon after detection of SI in all of them. We have followed their clinical courses for more than 4 years, so far, in three of them. Their CD4 counts and clinical courses were both stable. In contrast, in another patient who could not receive any anti-HIV therapy even though SI positive, decline of CD4 was very rapid (200/microliter/year) and the disease progressed to AIDS within one year as like other SI positive cases before the era of several approved anti-HIV drugs. According to these distinct prognosis, the MT-2 assay might be a useful clinical marker for deciding anti-HIV therapy at least in our limited cases. PMID- 10535267 TI - [Bacteriological survey of diarrheal epidemics in the 1998 Bangladesh floods]. AB - In 1998, the worst flood disaster in Bangladesh ravaged more than half of its land and diarrheal epidemics broke out. We examined fecal specimens of diarrheal patients at rural hospitals in Chandpor district located 140 km southeast of Dhaka to analyze the enteric bacterial pathogens in post-flood period October. Of the 76 patients stools examined, Vibrio cholerae O1 biotype El Tor, serotype Ogawa, and Vibrio cholerae O139 Bengal were detected in 25 (33%) and in 14 (18%) respectively. Other enteropathogenic bacteria confirmed were Vibrio cholerae O5, Vibrio fluvialis and Enteropathogenic Escherichia coli O44. Neither Shigella nor Salmonella species was detected in this study. A drug susceptibility test was performed using TC, DOXY, CPFX, NA, and AMPC disks to cholera Vibrios. The O1 Vibrios showed the same susceptible pattern as O139 excluding NA susceptibility. TC resistant strain among the Vibrios was not detected though TC is a common therapeutic drug for diarrhea in this area. Our result clearly suggested that the epidemic potentiality of O139 still existed in rural Bangladesh. PMID- 10535268 TI - [The molecular epidemiological analysis of Shigella sonnei isolates from outbreak cases in Yokohama]. AB - We conducted a molecular epidemiological analysis to evaluate the epidemiologic patterns of Shigella sonnei isolates from outbreak cases in Yokohama to clarify the epidemiologic linkages by contact tracing and sources of infection. In the first case (case A), all of the 6 isolates were the colicin 0 type and resistant to both streptomycin (SM) and trimethoprim/sulfamethoxazole (ST). The 5 isolates have plasmid of 230 kb. By RAPD analysis with 2 kinds of primers specific for Shigella, every 6 isolates showed the same pattern. But the DNA fingerprint analysis by PFGE that was performed according to 2 standardized restriction endonucleases revealed a discriminative pattern. However, the resemblance of all isolates, which was calculated by the UPGMA methods, was 0.90 or higher. In the second case (case B), all of the 14 isolates were the colicin 6 type and sensitive to 16 drugs. The serotype of 13 isolates was phase I. The 11 isolates have plasmids of 230 kb and 3 kb. The resemblance of all isolates, which was calculated by the UPGMA methods, was 0.89 or higher. The analysis with a combination of the plasmid, RAPD analysis and PFGE profiles may be effective in investigating detailed epidemiological features of isolates. PMID- 10535269 TI - [Two cases of suspected Bartonella henselae infection from a dog]. AB - A 55-year-old male was admitted to our hospital because of fever and left submaxillary, right axillary, and left inguinal lymphadenopathy. A presumptive diagnosis of rickettsiosis was made and treatment with oral doxycycline was started. Lymphadenopathy was partialy resolved after antibiotics treatment. Ablation of the left inguinal node was done and histopathological examination showed non-Hodgkin's lymphoma. Lymphadenopathy was resolved by chemotherapy. The second patient, a 40-year-old male, developed a tender submandibular node. Excisional biopsy of the node was performed to eliminate lymphoma. Histopathological examination revealed granulomatous lymphadenitis with follicular hyperplasia. The patients had no history of cat contact, but owned a dog. Diagnosis of both cases was confirmed by the detection of IgG antibodies to Bartonella henselae with an enzyme immunoassay. Our findings suggest that dogs are implicated in B. henselae infection and can serve as a reservoir of the organism as well as cats. In the abscence of other bacterial and especially after exposure to dogs, B. henselae should be included as possible cause of lymphadenopathy. PMID- 10535270 TI - [A case of endophthalmitis and abscesses in the liver and the lung caused by Klebsiella pneumoniae]. AB - [Case report] A 65-year-old male war admitted to a local hospital because of fever. He was treated with piperacillin and clindamycin without noticeable effect. He began to complain of loss of vision on the third hospital day and culture of the blood specimen yielded Klebsiella pneumoniae. He was diagnosed as endophthalmitis and referred to our hospital for further examination. The hematological laboratory test showed leukocytosis (12,700/microliter) and increased CRP (20.4 mg/dl). A computed tomographic (CT) scan of the thorax revealed multiple lung abscesses. An abdominal ultrasonographic scan and a CT scan of the abdomen revealed multiple liver abscesses. We drained the abscess in the liver and Klebsiella pneumoniae was detected from the sample of aspirated fluid and his sputum. Meropenem was administered intravenously. Fever started to improve on the tenth hospital day and the size of both liver and lung abscesses were reduced. He has lost vision of his right eye. He was discharged after sixty days. He did not have any immunosuppressive underlying disease including HIV infection and diabetes mellitus which cause these lesions. PMID- 10535271 TI - [A case of Chlamydia trachomatis and Mycoplasma pneumoniae coinfection in a child]. AB - Mycoplasma pneumoniae infection is often experienced as community-acquired pneumonia, whereas Chlamydia trachomatis pneumonia is sometimes experienced as a vertical infection from mother to child through delivery, but is rare after six months of age. We reported the first case to date of a seven-year-old boy with coinfection of C. trachomatis and M. pneumoniae. The patient, who was referred to our hospital because of suspected pneumonia, had had severe cough and pyrexia for four days before his admission. The coinfection was confirmed because the antibodies to both bacteria were significantly increased. In addition the PCR for C. trachomatis detected the presence of C. trachomatis in the throat even after treatment. The PCR from a cervix swab of his mother and from a throat swab of his father were negative, but their antibodies to C. trachomatis were positive. We think that this may have been community-acquired pneumonia or due to persistent infection after a vertical transmission. PMID- 10535272 TI - Mammalian neural induction and brain pattern formation. AB - In summary, as I mentioned, I have long be focusing on the cerebellum. I first talked about the IP3 receptor in relation to the Purkinje cells. Ca2+ release through IP3 receptor regulates the fertilization, cell division, and dorso ventral axis formation. It is also involved in learning and memory system. Gene called Zic is expressed in granule cells that make synaptic contact with Purkinje cells. Zic1 is one of the genes important in cerebellar pattern formation. Zic2 is essential in making our brain. Zic3 is important in the right and left axis formation. Purkinje cell and granule cell positioning are really nicely determined. But we now have two genes that determine the positioning, that may help to form a very nice and beautiful structure of the cerebellum. The principles and evidences obtained through the study of the cerebellum are found to be applied to other part of the brain, presumably because the cerebellum is phylogenetically old and plays an important information processing for motor movement. PMID- 10535273 TI - Hepatitis C and iron. AB - Hepatitis C is the commonest form of chronic viral hepatitis in most western countries. A significant proportion of patients develop cirrhosis, hepatic failure and hepatocellular carcinoma. The results of controlled trials have shown that interferon alpha is an effective treatment for hepatitis C. Treatment results in normalization of elevated transaminase levels in up to 50% of patients, although only 15-25% of patients have a sustained response. Recent studies have shown that iron influences the response of chronic hepatitis C to treatment and the natural history of hepatitis C. The mechanisms responsible for the effects of iron are not clear but emerging data suggest that the cellular location of iron within the liver lobule and the subsequent effects on immune function are likely to be critical determinants for these effects. It is likely that therapies for chronic hepatitis C which either remove iron or interfere with the action of iron at the cellular level may not only prove useful clinically but may also elucidate further the mechanisms of cellular injury in this disease. PMID- 10535274 TI - Family ward: a new therapeutic approach. AB - This article describes a new integrated child psychiatric family ward treatment model at the Tampere University Hospital. Theoretically, the treatment is based on an integration of systems and psychoanalytical theories as well as behavioral approach. A centerpiece of the model is a 3-week treatment period for the whole family at the family day ward. The work of the multidisciplinary team on the ward focuses on family relationships, on representational level, and on the interactional behavior of the family. Interaction and relationships are also used as tools, including a reflective working model and sharing concrete interaction with the family. So far, the family ward has offered 165 family treatment periods for 113 different families. Altogether in 63% of the total treatment periods one or both parents have had mental illness and in 15% of the total treatment periods there have been serious custody disputes with accusations of sexual abuse of the child. Helping these multi-problem families is a special challenge for our treatment model and at the moment we are developing new methods for assessment and support of parenthood. PMID- 10535275 TI - Ca(2+)-induced light adaptation in retinal ON-bipolar cells. AB - Retinal ON-bipolar cells possess a metabotropic glutamate receptor linked to the control of a cGMP cascade, which functions to generate high synaptic amplification of rod signals under dark-adapted conditions. We report that a major component of light adaptation of the rod visual system results from a reduction in gain in synaptic transmission from rods to ON-bipolar cells, initiated by Ca(2+)-loading when the cGMP-activated channels of the postsynaptic cell open with light. When intracellular Ca(2+)-buffering was reduced adaptation was induced in ON-bipolar cells by background light too weak to significantly desensitize rods, a property absent in OFF-bipolar cells. Desensitization of ON bipolar cell flash responses was induced by raising intracellular free Ca2+. PMID- 10535276 TI - Association between alcohol intake and subjective health: the Sotetsu Study. AB - Subjective health and alcohol intake are important predictors of mortality. There have been few epidemiological studies, however, of the relationship between alcohol consumption and subjective health among the Japanese. The purpose of this study is to investigate the association between alcohol consumption and subjective health. The study subjects were 2,020 Japanese male employees, who were free from serious disease conditions. The data on subjective health and alcohol consumption were obtained by means of self-reported questionnaire. The subjects who responded "poor health" in the answer to the question about the subjective health status were considered to be in ill-health. Ethanol intake per day was calculated by multiplying the frequency of drinking by the ethanol intake per drinking occasion and summing up for each alcoholic beverage. Age, smoking status, physical activity, and sleeping hours were treated as confounding factors. As a result, subjects who consumed 25-36 or 49 g ethanol or more per day had a significantly lower risk of self-rated ill-health compared with those who had never drunk, and a significantly inverse trend was found independent of age, smoking status, physical activity, and sleeping hours. In conclusion, moderate drinkers have a lower risk of self-rated ill-health among Japanese male employees investigated. PMID- 10535277 TI - Family therapy of schizophrenia. AB - Family therapy of schizophrenia has long been conceived and practised under etiological premises. Familial disturbances as pathological regression/fixation (psychoanalytical) and individuation-impairing family dynamics (systemic) were addressed directly in the hope of "curing" the disorder. The efforts to prove the viability of the concepts and/or the efficacy of the therapeutic approach were largely unsuccessful. Newer strategies of family therapy of schizophrenia are both more precise in their theoretical assumptions and more performing in the pursuit of their therapeutic goals. We analyse the basis of modern family therapy in the "Expressed-Emotions (EE)"--research and propose a newer, more adequate understanding of the EE phenomenon. From our own studies and from a general review of relevant studies we derive an understanding of the rationale of family work and family therapy of schizophrenia. We discuss the results of a meta analysis on the active ingredients and the conditions of efficacy of family interventions. PMID- 10535278 TI - [Internalization and replication of Mycobacterium tuberculosis and M. avium complex within type II alveolar epithelial cell line]. AB - Profiles of internalization and replication of Mycobacterium tuberculosis (MTB) or M. avium complex (MAC) within A-549 human type II alveolar epithelial cell line (A-549 cells) were studied and the results were compared with the mode of internalizing and proliferative behaviors of the organisms within murine peritoneal macrophages (M phi s) and various M phi-like cell lines (murine J774A. 1, human THP-1, and human MONO-MAC-6). First, MTB and MAC internalized not only in peritoneal M phi s and M phi-like cell lines but also in A-549 cells. Secondly, MTB and MAC replicated within A-549 cells and these organisms displayed much more vigorous intracellular multiplication in A-549 cells than in murine peritoneal M phi s and J774A. 1 M phi-like cell line. Human M phi-like cell lines (THP-1, MONO-MAC-6) allowed the growth of MTB and MAC equally or occasionally more rapidly, as compared to the case of A-549 cells. PMID- 10535279 TI - [Effects of the Chinese traditional medicines "mao-bushi-saishin-to" and "yokuinin" on the antimycobacterial activity of murine macrophages against Mycobacterium avium complex infection]. AB - We previously examined the effects of two Chinese traditional medicines "Mao Bushi-Saishin-To" (MBST) and "Yokuinin", on the therapeutic efficacies of a benzoxazinorifamycin, KRM-1648, against Mycobacterium avium complex (MAC) infection induced in mice. MBST but not Yokuinin potentiated the therapeutic activity of KRM-1648 against MAC infection. In the present study, we examined the effects of these traditional medicines on some M phi cell functions. First, MBST significantly potentiated M phi anti-MAC antimicrobial activity, while Yokuinin did so to a much lesser extent. Secondly, MBST and Yokuinin each strongly inhibited production of nitric oxide (NO) in MAC-infected M phi s. Thirdly, treatment of M phi s with MBST or Yokuinin caused reductions in the accumulation of IL-10 in culture fluids by MAC-infected M phi s during the first 2-days cultivation. On the other hand, in the separate experiment, treatment of M phi s with these drugs caused no significant change in the accumulation of TGF-beta by MAC-infected M phi s at day 7. These findings suggest that these Chinese traditional medicines, particularly MBST, potentiate M phi anti-MAC antimicrobial activity, however, NO do not appear to be crucial effectors in the anti-MAC activity of MBST- or Yokuinin-treated M phi s. Moreover, MBST- and Yokuinin mediated down-regulation of the production of IL-10 in MAC-infected M phi s may be related to their potentiating effects on M phi anti-MAC activity. PMID- 10535280 TI - [Current epidemiological trend of tuberculosis among foreigners in Japan]. AB - In Japan, the proportion of foreign patients among the total tuberculosis patients is still very small, but their problems in tuberculosis case finding and treatment require intensive control activities as in other low prevalence countries with higher proportion of foreign-born cases. The latest national survey for the foreign tuberculosis patients conducted in 1996 shows the epidemiological status between 1990 and 1993. The number of foreign tuberculosis patients in 1993 was 484, consisting of 1.0% of the total new patients in Japan. The new case rate among foreigners was estimated to be 53 per 100,000 against 38 for whole Japan in 1993. Compared with the figure in the 1993 survey, the number of foreign patients declined from 585 in 1992 to 484 in 1993. However, the number of bacillary positive tuberculosis patients in 1992 was 230 and almost the same as in 1992. The decline or stagnation of total number of tuberculosis patients can be due either to the decrease in the foreign population inflow into Japan (real decline), or partially to the reduction of overdiagnosis in X ray examination and the possible loss of some cases in the 1996 survey method. A manual sorting method from the registration cards was used at each public health center, since there is no item of country of origin in the routine tuberculosis surveillance system, and some cards might have already been displaced by the time of the survey for patients who were excluded from the registry, either cured, died or defaulted. The average treatment completion rate (1991-93) among foreign patients was 51%, which was much lower than the national figure of 81% for the same years. Moreover the rate showed deteriorating trend by year. For more accurate information, the foreigner's data must be taken in the national tuberculosis surveillance system and control activities for foreigners need to be strengthened. PMID- 10535281 TI - [Chemoprophylaxis against Mycobacterium avium complex infection induced in mice]. AB - M. avium complex (MAC) is one of the important causative agents of opportunistic infections among AIDS patients. Recent evidence showed that the entry of infection is through the gastrointestinal tract. In the present study, we compared the prophylactic effect of some antimicrobials against MAC infection induced in mice. Different groups of beige mice were fed with pellets containing 0.0067% (10 mg/kg) of KRM-1648, rifabutin (RFB) and clarithromycin (CAM). Seven days after the administration of drugs, the mice were infected with M. intracellular N-241 (5 x 10(8) CFU) orally, five times, every other day and killed one and 126 days after the last infection. The effect of drug was evaluated using the frequency and severity of gross lung lesions in the mice and by the total CFU recovered from the lungs and spleen. MAC infection was not likely to be established since there was no macroscopic evidence of lesion in organs and the recovery of cultures from lungs and spleen tested was negative, in 3 of 10 mice in the control group, 2 of 9 in the CAM group, 4 of 9 in the RFB group and 4 of 10 in the KRM group. These mice were excluded from the analysis of the study results. Thus, we examined 7 mice in the control group, 7 in the CAM group, 5 in the RFB group, and 6 in the KRM group. Tubercle-like lesions were observed in the lungs of all 7 mice in the control group (severity: 3+ in 5 mice; 4+ in 2 mice), in 5 of 7 mice (71%) in the CAM group (severity: 2+ in 1 mouse; 3+ in 4 mice), and in 4 of 5 mice (80%) in the RFB group (severity: 1+ in 1 mouse; 2+ in 1 mouse; 4+ in 2 mice), while only slight lesions (severity: 1+) were observed in 4 of 6 mice (67%) in the KRM group. There was no macroscopic evidence of lesion in spleen, liver and kidneys. The log CFU was determined at the next day of the completion of the last infection. The log CFU of the lungs was 2.49 and 2.28 in the control group and the CAM group, respectively. The bacteria were not recovered either from the lungs in the RFB and KRM groups, nor from the spleen in all the groups. The order of efficacy of the drugs on the basis of the CFUs recovered from the lungs and spleen in each group determined 126 days after the completion of the last infection was as follows; KRM > CAM > RFB in the lungs and KRM > CAM [symbol: see text] RFB in the spleen, although there was no significant difference among the three drugs (P < 0.05). However, the significantly preferable effect was obtained in the three drug groups as compared with the control group. PMID- 10535282 TI - [The 74th Annual Meeting Special Lecture. II. Present situation and problems of tuberculosis in Japan--its prevention, diagnosis and treatment]. AB - 1. Outline of tuberculosis epidemiology after the World War II Tuberculosis was surprisingly highly prevalent during the World War 2nd in Japan, and it was reported that the incidence of TB was as high as 698.4 in 1951. Strong TB control program has been carried out throughout the country after the new Anti-TB Law had been enacted in 1951. TB incidence decreased with the annual reduction rate of 11% up to 1976, it was one of the highest speed of TB decrease in the world. The decrease of TB is stagnating since 1977, and the incidence is still 33.9 in 1997. 2. Causes of stagnation of TB in Japan It is estimated that the annual risk of TB infection was so high as 4% in 1945 that many of the children had been infected with TB bacilli in those days. As a result, the majority of the people aged 60 years old or more at present are already infected with TB bacilli, and more than half of the new cases is being occurred among them at present in Japan. Moreover, the life span of those already infected people is extending so remarkably that the percentage of the aged is increasing and TB incidence is stagnating in Japan. The causes of the stagnation are not the spread of HIV infection or the increase of TB among the immigrants or refugees, because both of them are not so big problems in Japan. However, more severe problems of TB epidemiology in Japan are that 1. the incidence of smear positive cases didn't decrease more than these ten years, 2. the most marked stagnation of the incidence of TB is being observed among the young aged 20 to 39, 3. the incidence of TB in big cities doesn't decrease recently and 4. group infection and nosocomial TB infection is increasing recently. The author has analyzed the factors of the stagnation of TB in Japan, and come to the conclusions that it is caused by 1. the increase of the population by 3.0%, 2. the aging of the population by 41.0%, 3. the increase of compromised host (diabetes mellitus and so on) by 19.8%, and 4. other factors by 36.3%. It was considered that the main other factor is the marked stagnation of TB infection risk mainly by socioeconomic changes such as progressed urbanization and the spread of the modern housing with aluminum sash to increase the chances of TB infection in office buildings and/or private house. 3. Present problems of TB prevention, diagnosis and treatment in Japan As BCG vaccination is so widely and so repeatedly being carried out for children that the diagnosis of TB infection is almost impossible now in Japan. Accordingly the frequency and/or risk factors of TB infection in the community are not so clear. Preventive chemotherapy is given for high risk groups in Japan, but its diagnosis may become often unreliable. New techniques such as PCR, MTD, AccuProbe, RFLP and so on are used frequently in Japan, but the consideration on the false negative results of the examinations are lacking on occasion. The results of the cohort analysis of TB treatment among all the registered cases are rather satisfactory, but it was concluded that the shortening of the duration of the hospitalization and chemotherapy is advised if compared with those of Japan and those of the other developed countries. PMID- 10535283 TI - [The 74th Annual Meeting President Lecture. Pathogenesis and therapy of acute lung injury]. AB - Almost all of respiratory diseases except benign lung tumors and lung dysplasia entail acute lung injury (ALI). The many clinical conditions associated with acute lung injury include aspiration pneumonia, bacterial pneumonia and sepsis. Acute lung injury is the end results of common pathways initiated by a variety of local or systemic insults leading to diffuse damage to the pulmonary parenchyma. Despite the accumulation of abundant information regarding the physiological and cellular basis of lung injury and increasing sophisticated intensive care, an improvement in prognosis has lagged behind. It has become clear that there is not one mediator responsible for ALI, but rather a complex interplay exists between diverse proinflammatory (e.g., lipopolysaccharide, complement products, cytocains, chemocains, reactive oxygen species and arachidonic acid products) and anti-inflammatory (IL-10, IL-1-RA, PGI2) mediators. Early in the course of ALI, large numbers of neutrophils are sequestered in and emigrate from the pulmonary capillaries. The fundamental cause of ALI is pulmonary vascular hyperpermeability caused by the activated neutrophils which release oxygen radicals and proteases. In these processes several adhesion molecules play very important roles. Neutrophil elastase inhibitors become very useful therapeutic agents against acute exacerbation of idiopathic interstitial pneumonia (IIP), because this pathological conditions is a type of ALI. Similarly, N-acetyl cystein could also become a useful therapeutic agent against IIP, because it is a precursor of glutathione, which is the major antioxidant in the fluid lining of the bronchial epithelium. PMID- 10535284 TI - [Teeth displacements under occlusal force in view of dental arches]. PMID- 10535285 TI - [A concept on occlusion--the occlusal contacts]. PMID- 10535286 TI - [The relationship between inorganic ion composition of saliva and dental caries prevalence in children]. AB - In order to know the relationship between inorganic ion composition of saliva and dental caries prevalence, saliva from children was studied. The saliva samples were collected from 30 subjects aged 3 to 18 years. Each inorganic ion concentration and pH of saliva was measured and their relationship to dental caries was studied. The results were as follows: 1. Significant relationship was noted between ammonium ion concentration and caries prevalence. It was suggested that the ammonium ion was derived from decomposition of urea or amino acids. 2. Although a significant relationship was also noted between potassium ion concentration and caries prevalence, its direct process is unknown. 3. The sodium ion concentration in saliva increased with age, which implies its relationship to the development of the hormonal system. PMID- 10535287 TI - [A report on medical assistance for patients with cleft lip and palate and technical transfer in Vietnam]. AB - We had participated in the charitable operation for the patients with cleft lip and palate and technical suidance in Vietnam in 1998 for 2 weeks. The project was a part of the activities of the Japanese Cleft Palate Foundation, which were performed as part of the Official Development Assistance (ODA) of Japan, and was aided by state subsidy of the Ministry of Posts and Telecommunications, the Ministry of Foreign Affairs and the Ministry of Education and so on. Our team consisted of members of Tokyo Dental College, Iwate Medical University, Dalhousie University, and our university. The hospital we visited was Odonto-Maxillo-Facial Center in Ho Chi Min City, which had 7 clinical departments, 36 dental units, 60 beds in ward, and 75 doctors in total. This hospital dealt with all kinds of disease in connection with oral and maxillo-facial area including dental caries, periodontal disease, maxillo-facial injuries, and maxillo-facial anomalies. Forty five patients with cleft lip and palate were operated on during our stay in Vietnam. The age of these patients was higher than that in medical advanced countries because of economic problems and insufficient medical institutions. In some cases, surgery was performed in collaboration with local doctors, which means technical suidance leads to more effective treatment. PMID- 10535288 TI - [Anatomical names of fossae and foveae in skeleton]. AB - Latin anatomical names of Fossae and Foveae in the skeleton were analyzed and compared with Japanese anatomical names for better understanding of the structures of the human body and for possible revision in the future. The conclusions were as follows: 1. In general, round excavations were called Foveae (singular : Fovea), and nonround excavations were called Fossae (singular : Fossa). Some shallow excavations for articulation and some shallow excavations with the names which indicate their contents were called Foveae even though they were not round. 2. Each name of Fossae contained the word which indicates form, location or content of Fossa, the bone (or osseous structure) which articulates with Fossa, or the muscle which is attached to Fossa. 3. Each name of Foveae contained the word which indicates location, content or articulation of Fovea, the bone (or osseous structure) which articulates with Fovea, or the muscle (or muscular trochlea) which is attached to Fovea. 4. The Japanese name which corresponds to Fossa canina should be changed from Kenshi (canine tooth) = ka (fossa) to Kenshikin (canine muscle) = ka or Koukakukyokin (levator anguli oris muscle) = ka. 5. The Japanese name which corresponds to Fossa pterygopalatina should be changed from Yoku (wing) = kougai (palate) = ka (fossa) to Yokutotsu (pterygoid process) = kougaikotsu (palatine bone) = ka. PMID- 10535289 TI - [Statistical analysis of fabrication of indirect single restorations]. AB - A statistical survey based on laboratory records was performed on the number of indirect restorations fabricated at the dental hospital of Tokyo Medical and Dental University from April 1 to September 30, 1997. A comparison was also carried out with a previous survey, which had been carried out in 1986, in order to detect any change and possible alterations in the near future. Based on the results of this statistical survey, the conclusions were as follows: 1. A total of 9,126 indirect restorations were fabricated during the six month period in 1997; among them, 8,007 (87.7%) restorations were covered by health insurance and 1,119 (12.3%) restorations were not. 2. The most common restoration was the cast post and core (28.6%), followed by full crowns (18.5%) and removable partial dentures (15.6%). On the other hand, the least number were post crowns (0.03%) and resin jacket crowns (0.2%). 3. When making a comparison with the data in 1986, an increase in the number of removable partial dentures and a decrease in the number of inlays were the most distinctive features. 4. For anterior teeth, resin-veneered crowns were most common, especially for lower teeth. The percentage of restorations, which were not covered by health insurance, decreased from 45.0% (in 1986) to 12.3% (in 1997). PMID- 10535290 TI - [Statistical analysis of fabrication of fixed and removable partial dentures]. AB - A statistical survey based on laboratory records was performed on the number of fixed and removable partial dentures fabricated at the dental hospital of Tokyo Medical and Dental University from April 1 to September 30, 1997. A comparison was also performed with a previous survey, which had been carried out in 1986, in order to detect any change and possible alterations in the near future. From the findings of this statistical survey, the conclusions were as follows: 1. A total of 2,478 fixed and removable partial dentures were fabricated during the six month period in 1997. 2. The 3-unit fixed partial denture (bridge) was most common (63.1%) and the number of bridges decreased as the number of units increased. 3. For the single missing tooth, a fixed bridge was more popular (81.0%) than a removable partial denture. 4. For two missing teeth, there was no difference between the number of fixed bridges and removable partial dentures. 5. The percentage of fixed and removable partial dentures, which were not covered by health insurance, decreased to a large extent in comparison with the survey in 1986. PMID- 10535291 TI - [Study on oral health status and health behavior of workers at government office]. AB - The purpose of this study was to investigate the oral health status and health behavior of the adult population in the workplace. In 1998, oral health examination of 388 workers (male: 287, female: 101) at a government office was performed. A questionnaire was administered to obtain data regarding oral symptoms and health behavior. The results were as follows: Overall, 48% needed treatment for dental caries, 44% needed calculus removal, and 23% needed treatment for periodontal disease. Although there were no oral health complaints, 20% had early caries, 40% had dental calculus, and 19% had periodontitis. Compared to males, more females brushed their teeth, had home dentists (44%) and received more regular dental health check-ups at least once a year (48%). For males, those with home dentists had higher FT and DMFT in the twenties and thirties. There was no relationship between oral health status and regular check ups in both males and females. The results revealed that receiving regular dental check-ups from home dentists was not popular in Japan. Further, the role of home dentists is not preventive oriented. It was concluded that it is necessary to provide regular oral health examination and health promotion programs for adult population at the workplace in Japan. PMID- 10535292 TI - Virtual neuroendoscopy, a comparative magnetic resonance and anatomical study. AB - We evaluated the usefulness and reliability of intraventricular virtual neuroendoscopy based on a comparative anatomical study. Virtual intraventricular endoscopic images were calculated from 3D magnetic resonance images in five anatomic specimens. Contiguous 1.2 mm slices of the specimen heads were acquired at a 1.5 T MR scanner using a 3D-gradient echo sequence. The images were then transferred to an independent 3D-workstation (Sun Spark 20). After scanning the specimen heads, real endoscopy within the cerebral ventricles of these brains was performed with a standard rod lens system. Comparison between real and virtual endoscopic views of the intraventricular topography was based on the same anatomical reference and landmarks. Acquisition of MR data and virtual image post processing have been possible in all specimens. The virtual endoscopic images of the ventricles were comparable to the intraventricular views obtained by a standard rod lens system. Virtual intraventricular neuroendoscopy can be employed for planning and simulating neuroendoscopic procedures. It enables the neurosurgeon to simulate the endoscopic procedure within the cerebral ventricles on the basis of the patient's individual anatomy prior to surgery. PMID- 10535293 TI - MRI-guided brain biopsies using a 0.2 Tesla open magnet. AB - The technique of interventional on-line MR-guided procedures in a 0.2 Tesla open magnet is presented. 72 procedures were performed including 60 brain biopsies and 12 interventions as catheter placement for cyst or abscess drainage. The advantages and limitations are discussed and recent developments such as MR-coil designs and MR-sequences are presented. PMID- 10535294 TI - Intraoperative image-directed dye marking of tumor margins. AB - The incorporation of interactive image guidance during intracranial tumor surgery offers the possibilities of reduced operative trauma, shorter operation time, greater precision, and an increased understanding of complex anatomy and pathology. A basic weakness with these systems though is that they cannot account for movement of target points due to brain shift by draining of CSF or removal of pathology during the operative procedure. We have developed a stereotactic (frameless) guided injector probe for marking the tumor boundary with dye injection in conjunction with a neuronavigation system. The device consisted of a rigid blunt hollow probe (2 mm dia.) with 4 small side holes at the tip. The catheter is mounted in a holder equipped with 3 LEDs supplying guidance information for the neuronavigation system. A small manual aliquoting pump delivers a measured amount of dye in each track. Isotonic methylene blue was injected in 6 to 8 tracks around the periphery of the tumor as determined by the contract ring in MR scans. The dye was injected using image-directed guidance before resection of the tumor was started (often with the dura intact). Tumor tissue could then be resected until the dye became visible at the tumor boundary. Identification of the dye in the tissue was enhanced with the use of the operating microscope. The 3-dimensional position of the dye track could be determined at the end of tumor resection and compared with its initial position giving a good estimate of local brain shift. The method has proved especially helpful for the resection of large gliomas allowing for a more radical operative result. PMID- 10535295 TI - Endoscopic third ventriculostomy for hydrocephalus. AB - The authors report on 125 patients who underwent endoscopic third ventriculostomy for obstructive hydrocephalus in three Italian Neurosurgical Centers. The series includes 77 cases of primary aqueductal stenosis, 33 with triventricular hydrocephalus due to external tumor compression, and 15 with tetraventricular hydrocephalus. The operations were carried out mainly under general anesthesia, using a flexible endoscope. Decrease of size of the third ventricle and the presence of a signal void at the level of the fenestration are the main postoperative MRI findings. Signs of intracranial hypertension, increased head circumference and Parinaud syndrome respond more frequently to the endoscopic treatment. The overall rate of good results (shunt-independent patients) in this series is 86.4%; primary aqueductal stenosis (93.5%) and triventricular hydrocephalus due to external compression (84.8%) are associated to the higher rate of good postoperative results than tetraventricular hydrocephalus (53.3%). Because of the very low invasivity of this technique, the absence of postoperative mortality and the scarce and usually transient postoperative complications, the authors advise to enlarge the indications for endoscopic third ventriculostomy to all patients with obstructive hydrocephalus when the third ventricle is large enough and there are no alterations of the CSF resorption. PMID- 10535296 TI - The trans-supraorbital approach. AB - The trans-supraorbital approach represents the advantage of combining the keyhole principle and skull base surgery. In cadaveric work the author shows the anatomic fields visualized with this procedure, advantages and potential surgical applications. The experimental surgical procedure was performed on 8 adult skulls and on 3 cadavers with intact cerebral structures. First, a 2 cm supraorbital approach for endoscopic exploration was used. The basic anatomic landmarks and corresponding dimensions of the microsurgical field were recorded and then compared with those from the trans-supraorbital approach, after removing the orbital arch and other parts of the orbital ceiling. This technique offers a 0.5 cm average increase in surgical field from the craniotomy. The surgical distance that results is shorter (about 2 cm), as well as the length of the surgical instruments required. A better microscope illumination in the deep fields, and the possibility of access to the intraorbital region, the superior orbital fissure, the optic foramen, and the cavernous sinus through the clinoid space are also obtained, with minimal cerebral retraction and similar advantages as through the pterional trans-sylvian approach. The author concludes that by extending the craniotomy and decreasing the surgical distance, the microsurgical field is more convenient for microscope-assisted surgery without totally relying on the endoscope, and with minimal brain retraction. It also provides multiple options to approach vascular and tumor lesions found in these microsurgical corridors, combining principles of minimal invasion and skull-base surgery. PMID- 10535297 TI - Inferolateral microsurgical approach to the orbit: an anatomical study. AB - Using detailed cadaveric dissection this study has demonstrated the increase in exposure by using the inferolateral microsurgical approach for neurosurgical and ophthalmological access. The approach to surgical exploration of this region is divided into three steps. The neural, muscular and vascular structures of each step are discussed. We think that, with an intimate understanding of the anatomy of the orbit, many large intraorbital lesions located in the muscle cone and the inferior nasal compartment of the orbit can be safely removed through inferolateral orbitotomy. PMID- 10535298 TI - A novel endoscopic approach to anterior odontoid screw fixation: technical note. AB - Techniques for operative management for type II odontoid fractures have continuously been refined with anterior odontoid screw arthrodesis having a clear advantage in maintaining normal motion. We have refined the technique of odontoid screw fixation further with the introduction of an endoscopic approach developed by the senior author. The necks of two partially embalmed cadavers were slightly extended under fluoroscopic guidance to simulate a reduced, anteriorly displaced type II fracture. Using a guide wire, graduated plastic sheath and endoscopic guidance, a solid 45 mm bone screw was passed through the odontoid with the aid of biplanar fluoroscopy. There were no apparent complications and no damage to surrounding vital structures. Anterior screw fixation of the odontoid is an established technique that provides adequate fixation, but the procedure can be technically demanding secondary to awkward tissue retraction. We present a percutaneous technique that obviates the need for tissue retraction while achieving an excellent result with only a modicum of effort. PMID- 10535299 TI - Endoscopy of the spinal cord: cadaveric study and clinical experience. AB - Recent improvements in instruments permit endoscopic examination of previously inaccessible sites. We report on the clinical use of a small-diameter endoscope to examine the spinal subarachnoid space, cord surface and syrinx cavities. Prior to clinical application, three types of endoscopes with external diameters of 0.5, 1.4 or 2.2 mm were inserted percutaneously in the lumbar region of five cadavers for preclinical evaluation of the procedure and the three endoscopes. The observations permitted us to perform spinal endoscopy preoperatively or intraoperatively using the 0.5-mm instrument in seven patients with spinal cord lesions between 1995 and 1997. The patients included two with spinal cord herniation through a dural defect, two with syringomyelia, one with spinal arachnoid cyst, one with spinal epidural cyst and one undergoing lumboperitoneal shunt for hydrocephalus. In patients in whom an endoscope was used preoperatively, the endoscope provided morphological information useful in preoperative diagnosis and planning surgical strategy. When the endoscope was used intraoperatively, areas outside the field of vision of a microscope could be examined, and physiological evaluation could include visualizing improved cord perfusion from the spinal subarachnoid space after surgery. Endoscopes could be safely inserted and approached to the lesions under direct vision while avoiding blood vessels and nerve roots on the spinal cord surface. No changes in symptoms or complications occurred in association with endoscopy. Using a small-diameter endoscope, the contents of the spinal subarachnoid space could be examined. Further improvements to increase possible endoscopic manipulation and enhance safety may extend the possibilities for endoscopic examination and permit endoscopic treatment. PMID- 10535300 TI - Percutaneous nucleotomy in the treatment of cervical disc herniation: report of three cases and review. AB - We describe here three patients with cervical disc herniation who were treated at our department with manual percutaneous nucleotomy. These patients had suffered from cervical pain radiating into an upper extremity, with the duration of the conservative treatment varying between three and six months before the operation. After a follow-up of three years, radiating pain into the upper extremity was totally recovered in one patient, markedly diminished in one and remained unchanged in one. All patients were continuously working. The outcome of patients who underwent percutaneous nucleotomy for cervical disc herniation has been evaluated in only three previous papers, all focusing on automated cervical nucleotomy. We review the literature on cervical percutaneous nucleotomy and discuss the usefulness of this methodology. PMID- 10535301 TI - Multilevel vertebral body replacement with a titanium mesh spacer for aneurysmal bone cyst: technical note. AB - A 64-year-old male presented with abrupt tetraparesis caused by a minor impact. Diagnostic images obtained on admission showed an aneurysmal bone cyst visible in the cervical spine at the fourth to upper sixth level, although the patient had been wearing a halo brace to diminish the symptoms. The vertebral body from the fourth to the sixth level was dissected, and this space was packed with a titanium cage filled with ceramic bone fragments mixed with fibrin glue. The combination of a titanium cage and an anterior locking plate can be made easily for anterior spinal fusion with enough rigidity to maintain the necessary space during fusion without any major support equipment. Both edges of the titanium mesh cage cut into the vertebral body to hold the cage in place. The other part, the titanium plate, makes it secure until ceramic bone fragments in the cage promote bony ingrowth for fusion. PMID- 10535302 TI - Inverse relationship between connexin43 and desmin expression in cultured porcine aortic smooth muscle cells. AB - Our previous work has shown that in vascular tissues the elastic medial regions express high levels of the gap junctional protein, connexin43, but low levels of desmin, while the muscular medial regions express low levels of connexin43 but high levels of desmin. It is uncertain, however, whether this regional difference at the tissue level extends down to the level of the individual cell, or reflects an averaged relationship of groups of cells of different connexin43 and desmin expression. The present study has addressed this question using cultured porcine aortic smooth muscle cells. Immunoconfocal microscopic analysis of single-labeled cells showed that while smooth muscle alpha-actin, calponin and vimentin were positively labeled in the majority of medial smooth muscle cells both in intact porcine aorta and corresponding cultured cells, desmin and connexin43 labeling was highly heterogeneous. In the cultured cells, 0.3-0.5% of cells were found to be desmin-positive, and quantitative analysis after double labeling for desmin and connexin43 revealed that the desmin-positive cells were smaller, and contained significantly lower numbers and smaller sizes of connexin43 gap junctional spots than did desmin-negative cells. Our findings demonstrate that an inverse expression pattern of connexin43 and desmin holds true at the level of the individual cell. This suggests a close relationship between intrinsic phenotypic control and the regulation of connexin43 expression in the arterial smooth muscle cell. PMID- 10535303 TI - Endoglin overexpression modulates cellular morphology, migration, and adhesion of mouse fibroblasts. AB - Endoglin is the gene mutated in hereditary hemorrhagic telangiectasia type 1 (HHT1), a dominantly inherited vascular disorder. Endoglin glycoprotein is a component of the transforming growth factor type beta (TGF-beta) receptor system which is highly expressed by endothelial cells, and at lower levels on fibroblasts and smooth muscle cells, suggesting the involvement of these lineages in the HHT1 vascular dysplasia. Overexpression of endoglin in mouse NCTC929 fibroblasts led to decreased migration in chemotactic and wound healing assays, as well as changes in the cellular morphology. When plated on uncoated surfaces, endoglin transfectants formed intercellular clusters, endoglin being not specifically localized to the cell-cell junctions, but homogenously distributed on the cellular surface. Although the expression of alpha5beta1 integrin and of an activation epitope of beta1 integrin were unchanged, a polyclonal antibody to alpha5beta1 integrin was able to inhibit cluster formation, suggesting the involvement of integrin ligand/s. In fact, coating with fibronectin, laminin, or an RGD-containing 80 kDa fragment of fibronectin were able to prevent the cellular clustering. Furthermore, synthesis of plasminogen activator inhibitor 1 (PAI-1), and to a weak extent that of fibronectin, were inhibited in endoglin transfectants. Thus, the presence of endoglin in mouse NCTC929 fibroblasts is associated with reduced production of certain extracellular matrix (ECM) components, which might explain their altered morphology, migration and intercellular cluster formation. PMID- 10535304 TI - Membrane orientation of carboxyl-terminal half P-glycoprotein: topogenesis of transmembrane segments. AB - Multiple topological orientations of the carboxyl-terminal half of P-glycoprotein have been observed. One orientation is consistent with the hydropathy-predicted model and contains six transmembrane (TM)-spanning regions. In another orientation, the cytoplasmic-predicted loop between TM8 and TM9 is extracellular and glycosylated. In support of this "alternative" topology, TM8 was previously established to function as a signal-anchor sequence to insert with its amino terminal end in the cytoplasm and the carboxyl-terminal end in the extracytoplasmic space. However, it is unclear how downstream TM segments fold in the membrane when TM8 functions as a signal-anchor sequence. Here, we created several chimeric Pgp molecules to examine the membrane insertion of TM segments 9 and 10 using a cell-free system. We found that TM9 functions as a stop-transfer sequence when following the signal-anchor sequence, TM8. However, the stop transfer activity of TM9 depends on the presence of TM10. In the absence of TM10, TM9 partially translocated across the membrane into the endoplasmic reticulum lumen. In contrast, TM9 efficiently stopped the translocation event of the nascent chain in the presence of TM10. Our results suggest that the membrane insertion of TM8 and TM9 establishes the extracellular loop between TM8 and TM9. Formation of this loop apparently involves the interactions between Pgp TM segments, which facilitate proper folding of the Pgp carboxyl-terminal half. PMID- 10535305 TI - High-affinity cholecystokinin type A receptor/cytosolic phospholipase A2 pathways mediate Ca2+ oscillations via a positive feedback regulation by calmodulin kinase in pancreatic acini. AB - In rat pancreatic acini, we previously demonstrated that depending on the agonist used, activation of cholecystokinin type A (CCKA) receptor (CCK-AR) results in the differential involvement of the cytosolic phospholipase A2 (cPLA2), phospholipase Cbeta1 (PLCbeta1) and Src/protein tyrosine kinase (PTK) pathways. The high-affinity CCK-AR appears to be coupled to the Gbeta/cPLA2/arachidonic acid (AA) cascade in mediating Ca2+ oscillations. The low-affinity CCK-AR is coupled to both the Galphaq/11/PLCbeta1/inositol 1,4,5-trisphosphate (IP3) to evoke intracellular Ca2+ release and the Src/PTK pathway to mediate extracellular Ca2+ influx. The objectives of this study were to provide evidence that cPLA2 is present in pancreatic acini and to evaluate the possibility that its activation results in Ca2+ oscillations and amylase secretion. Furthermore, we investigated the mechanism of Ca2+ oscillations mediated by the high-affinity CCK-AR. In rat pancreatic acini, immunoprecipitation studies using an anti-cPLA2 monoclonal antibody, demonstrated a cPLA2 band at the location of 110 kDa. A selective inhibitor of cPLA2, AACOCF3 (100 microM), inhibited production of AA metabolites, Ca2+ oscillations and amylase secretion elicited by the high-affinity CCK-AR agonist, CCK-OPE (10-1000 nM). In addition, through the repetitive release of intracellular Ca2+, CCK-OPE enhanced phosphotransferase activities of Ca2+/calmodulin-dependent protein kinase type IV (CaMK IV), which were inhibited by AACOCF3. The CaMK inhibitor, K252-a (1-3 microM), also abolished basal and CCK OPE-stimulated CaMK IV activities. The CaM inhibitor, W-7 (100 microM), and K252 a inhibited Ca2+ oscillations and amylase secretion evoked by CCK-OPE without affecting the AA formation. Therefore, it appears that Ca2+ oscillations elicited by the high-affinity CCK-AR/Gbeta/cPLA2/AA pathway activate CaMK IV. Activated CaMK, in turn, regulates Ca2+ oscillations through a positive feedback mechanism to mediate pancreatic exocytosis. PMID- 10535306 TI - Nuclear targeting of cAMP response element binding protein 2 (CREB2). AB - The transcription factor cAMP response element binding protein 2 (CREB2) belongs to a family of proteins containing a basic region as DNA-binding domain and a leucine zipper as a dimerization domain in its C-terminus. Using indirect immunofluorescence labeling of cells we show that CREB2 is a nuclear protein. To identify the signal(s) required for nuclear targeting of CREB2, various domains of the protein were expressed in COS cells as fusion proteins with glutathione S transferase and their cellular location assayed by indirect immunofluorescence using antibodies directed against the glutathione S-transferase moiety of the fusion proteins. The results show that the nuclear targeting signal is located in the C-terminal part of the molecule. Deletion mutagenesis revealed that the basic region of CREB2, encompassing amino acids 280 to 300, is sufficient for sorting CREB2 to the nucleus. Single point mutations of basic amino acids within the basic region of CREB2 identified the sequence KKLKK (amino acids 280 to 284) as important for nuclear targeting. Thus, the basic region of CREB2 is necessary not only for tethering CREB2 to DNA but also for sorting CREB2 to the nucleus. However, sequences outside the basic region are additionally required for efficient nuclear sorting of CREB2. PMID- 10535307 TI - The synaptophysin-synaptobrevin complex is developmentally upregulated in cultivated neurons but is absent in neuroendocrine cells. AB - Regulated secretion requires the formation of a fusion complex consisting of synaptobrevin, syntaxin and SNAP 25. One of these key proteins, synaptobrevin, also complexes with the vesicle protein synaptophysin. The fusion complex and the synaptophysin-synaptobrevin complex are mutually exclusive. Using a combination of immunoprecipitation and crosslinking experiments we report here that the synaptophysin-synaptobrevin interaction in mouse whole brain and defined brain areas is upregulated during neuronal development as previously reported for rat brain. Furthermore the synaptophysin-synaptobrevin complex is also upregulated within 10-12 days of cultivation in mouse hippocampal neurons in primary culture. Besides being constituents of small synaptic vesicles in neurons synaptophysin and synaptobrevin also occur on small synaptic vesicle analogues of neuroendocrine cells. However, the synaptophysin-synaptobrevin complex was not found in neuroendocrine cell lines and more importantly it was also absent in the adrenal gland, the adenohypophysis and the neurohypophysis although the individual proteins could be clearly detected. In the rat pheochromocytoma cell line PC 12 complex formation between synaptophysin and synaptobrevin could be initiated by adult rat brain cytosol. In conclusion, the synaptophysin synaptobrevin complex is upregulated in neurons in primary culture but is absent in the neuroendocrine cell lines and tissues tested. The complex may provide a reserve pool of synaptobrevin during periods of high synaptic activity. Such a reserve pool probably is less important for more slowly secreting neuroendocrine cells and neurons. The synaptophysin on small synaptic vesicle analogues in these cells appears to resemble the synaptophysin of embryonic synaptic vesicles since complex formation can be induced by adult brain cytosol. PMID- 10535308 TI - Expression of the beta-chain of the complement regulator C4b-binding protein in human ovary. AB - Human C4b-binding protein (C4BP) is an important regulator of the complement system that also binds and inactivates the anticoagulant vitamin K-dependent protein S. These two activities are performed by two distinct polypeptides of 70 kDa and 45 kDa known as alpha and beta chains, respectively. C4BP is present in plasma in various isoforms with different alpha/beta composition. We report here that C4BPbeta, but not C4BPalpha, is expressed in adult human ovary. Expression of C4BPbeta was detected in all ovarian biopsies analyzed (n = 15), independently of age and phase of the menstrual cycle. In situ hybridization and immunostaining analyses on cryostat sections demonstrated expression of C4BPbeta in both regressing corpus luteum and corpus albicans, but not in the follicles, the corpus luteum, the ovary stroma or the vascular cells. In addition, we noted that the expression pattern of the C4BPbeta mRNA resembles that described for the connective tissue that invades the degenerating corpus luteum and causes a progressive fibrosis that gradually converts it into a scar, the corpus albicans. RT-PCR and immunostaining analyses of primary cultures derived from human ovaries demonstrated the presence of fibroblast-like cells that express C4BPbeta. As a whole, these data suggest a role for the C4BPbeta in human ovary during the healing and scar resorption processes that leads to the formation of the corpus albicans and its replacement by ovarian stroma. PMID- 10535309 TI - The Plasmodium falciparum translationally controlled tumor protein: subcellular localization and calcium binding. AB - Artemisinin derivatives are endoperoxide antimalarials widely used to treat falciparum malaria in areas where drug resistance is common. In Plasmodium falciparum-infected erythrocytes, radiolabeled artemisinin derivatives have been shown to react with malarial proteins, one of which is the Translationally Controlled Tumor Protein (TCTP). The P. falciparum TCTP was found by immunofluorescence to be located in both the cytoplasm and food vacuoles. Immunoelectron microscopy shows that it is present in the parasite cytoplasm as well as in its food vacuolar and limiting membranes. Like other TCTPs, the P. falciparum protein binds to calcium. Further studies on the physiological role of TCTP may aid in understanding the mechanism of action of endoperoxide antimalarials. PMID- 10535310 TI - Microtubule polyglutamylation in Drosophila melanogaster brain and testis. AB - The presence of glutamylated tubulin, a widespread posttranslational modification of alpha- and beta-tubulin, has been investigated in Drosophila melanogaster using the specific monoclonal antibody GT335. We show here that this modification is strongly detected in brain and testis whereas other tissues analyzed did not appear to contain any glutamylated isoforms. Neuronal microtubules are glutamylated on alpha-tubulin only whereas sperm flagella showed a strong modification of both alpha- and beta-tubulin. These results argue for an essential role for glutamylation in differentiation processes that require microtubule stabilization. PMID- 10535311 TI - Clifford Grobstein (1916-1998) and the developing kidney. PMID- 10535312 TI - Inductive tissue interactions. An interview with Chancellor Lauri Saxen. Interview by Eero Lehtonen. PMID- 10535313 TI - What is needed for kidney differentiation and how do we find it? PMID- 10535314 TI - Towards a molecular anatomy of the Xenopus pronephric kidney. AB - Kidney development is distinguished by the sequential formation of three structures of putatively equivalent function from the intermediate mesoderm, the pronephros, mesonephros, and metanephros. While these organs differ morphologically, their basic structural organization exhibits important similarities. The earliest form of the kidney, the pronephros, is the primary blood filtration and osmoregulatory organ of fish and amphibian larvae. Simple organization and rapid formation render the Xenopus pronephric kidney an ideal model for research on the molecular and cellular mechanisms dictating early kidney organogenesis. A prerequisite for this is the identification of genes critical for pronephric kidney development. This review describes the emerging framework of genes that act to establish the basic components of the pronephric kidney: the corpuscle, tubules, and the duct. Systematic analysis of marker gene expression, in temporal and spatial resolution, has begun to reveal the molecular anatomy underlying pronephric kidney development. Furthermore, the emerging evidence indicates extensive conservation of gene expression between pronephric and metanephric kidneys, underscoring the importance of the Xenopus pronephric kidney as a simple model for nephrogenesis. Given that Xenopus embryos allow for easy testing of gene function, the pathways that direct cell fate decisions in the intermediate mesoderm to make the diverse spectrum of cell types of the pronephric kidney may become unraveled in the future. PMID- 10535315 TI - A bioinformatics approach to investigating developmental pathways in the kidney and other tissues. AB - Over the past few years, large amounts of data linking gene-expression (GE) patterns and other genetic data with the development of the mouse kidney have been published, and the next task will be to integrate these data with the molecular networks responsible for the emergence of the kidney phenotype. This paper discusses how a start to this task can be made by using the kidney database and its associated search tools, and shows how the data generated by such an approach can be used as a guide to future experimentation. Many of the events taking place as the kidney develops do, of course, also take place in other tissues and organisms and it will soon be possible to incorporate relevant information from these systems into analyses of kidney data as well as the new information from microarray technology. The key to success here will be the ability to access over the internet data from the textual and graphical databases for the mouse and other organisms now being established. In order to do this, informatic tools will be needed that will allow a user working with one database to query another. This paper also considers both the types of tools that will be necessary and the databases on which they will operate. PMID- 10535316 TI - Role of BMP family members during kidney development. AB - Members of the Bone morphogenetic protein (BMP) family have been shown to be important signaling molecules throughout mouse development. Accordingly, many BMPs are also expressed during organogenesis of the metanephric kidney. However, only BMP7 has been shown to be absolutely required for proper formation of the kidney, thus the majority of information known involves this family member. BMP7 is expressed in both the ureteric epithelium and the mesenchyme throughout embryonic development and has been shown to function as a survival factor for the nephrogenic mesenchyme. However, there has been some controversy over the role of BMP7 as an inducing molecule for the metanephric mesenchyme. Recent studies have shown that BMP7 functions as an anti-differentiation factor for this mesenchyme cell population. The function of BMPs in the ureter and in the more differentiated epithelial structures of the nephron is less well understood. PMID- 10535317 TI - GDNF and its receptors in the regulation of the ureteric branching. AB - Recent transgenic and organ culture experiments have inevitably shown that glial cell line-derived neurotrophic factor (GDNF) is a mesenchyme-derived signal for ureteric budding and branching. The signalling receptor complex for GDNF includes a dimer of Ret receptor tyrosine kinase and two molecules of GDNF family receptor alpha1. Alpha-receptors are not only needed for the ligand binding and Ret activation, but they might mediate signals without Ret. While GDNF is clearly required for ureteric branching, tissue recombination studies have shown that it is not sufficient for the completion of ureteric morphogenesis, and other signalling molecules are needed. Different experimental models have resulted in somewhat contradictory results on their molecular identity, but transforming growth factor-beta1, -beta2, fibroblast growth factor-7 and hepatocyte growth factor form, obviously among others, a redundant set of growth factors in ureteric differentiation. Three other members of the GDNF family, neurturin, artemin and persephin, are also expressed in the developing kidney, and at least neurturin and persephin promote ureteric branching in vitro, but their true in vivo roles are still unclear. PMID- 10535318 TI - Wnts as kidney tubule inducing factors. AB - Since the discovery that inductive tissue interactions regulate nephrogenesis, one of the aims has been to identify the molecules that mediate this induction. The small size of embryonic tissue has limited the possibilities to identify the inducers biochemically, even though such efforts were directed to study, e.g. neural induction (for a comprehensive review, Saxen and Toivonen, Primary embryonic induction, Academic Press, London, 1962). The rapid progress in molecular biology made it possible to identify genes from minute amounts of tissue and provided techniques to generate recombinant proteins to assay their action in classic experimental systems. This led to the identification of some signals that are involved in primary and secondary inductive interactions during embryogenesis. Here, we will review evidence suggesting that secreted signaling molecules from the Wnt gene family mediate kidney tubule induction. PMID- 10535319 TI - Vesicular transport and kidney development. AB - Vesicular transport processes play crucial roles in the biogenesis of cellular membranes and in the polarized transport functions of epithelial cells. During the 1990's we have witnessed major progress in elucidation of the machineries responsible for the intracellular membrane trafficking. The components of these machineries are abundant in tissues with a high content of epithelial cells, such as the kidney. However, the developmental role of the membrane trafficking apparatus in higher eukaryotes has been addressed hardly at all. We summarize here data on the presence and the functional role of vesicle transport proteins in the kidney, and describe work addressing the developmentally regulated expression and localization of three molecules suggested to be involved in polarized trafficking in kidney epithelia, Rab17, syntaxin 3, and Munc-18-2. The results show that specialized transport machinery is induced during differentiation of renal epithelia. However, the expression levels of the components under study are highest in the mature structures, indicating that the proteins are predominantly required for the function of mature epithelia and possibly for the maintenance of the polarized phenotype of specific epithelial cells. The proteins are, however, detected at low levels already in earlier, differentiating structures, and could thus also be involved in the differentiation of kidney epithelia. PMID- 10535320 TI - Mesonephric kidney--a stem cell factory? AB - Mesonephros is a vestige, transient renal organ that functions only during embryonic development. The anatomy, position and even cellular fate of the mesonephric kidney varies drastically among mammalian species. The origin of mesonephros from intermediate mesoderm and the dependence of its differentiation on the nephric or Wolffian duct have been well established. Commonly accepted is also the mesonephric origin of epididymal ducts of the male reproductive tract. Recently, upon the more profound understanding of the molecular mechanisms involved in the development of the permanent mammalian kidney, some light has been shed over the molecular events taking place during the mesonephric development as well. Because of the functional and structural similarities between the mesonephric and metanephric kidneys, it is not surprising that many molecules regulating metanephric development are also activated during mesonephric development. However, the multifunctional nature of mesonephros has been unexpected. First, it serves as an embryonic secretory organ, in some mammalian species more so than in others. It is thereafter removed by programmed cell death. Second, it is a source of multiple stem cells including somatic cells in the male gonad, vascular endothelial cells, and hematopoietic stem cells. Thus, mesonephros is a challenging model for studies on epithelial differentiation and organogenesis, regulation of apoptosis, sex determination and stem cell differentiation. In this review, we focus in the molecular and stem cell aspects in the differentiation of the mammalian mesonephros. PMID- 10535321 TI - Dopamine in the developing kidney. AB - The adult kidney has a high rate of dopamine (DA) production, metabolism, and signalling. The non-neuronal DA system in the adult kidney is of utmost importance for the regulation of salt metabolism. DA may also act as a transcription factor and may be of importance for tissue differentiation. In the central nervous system, D1 receptors require the dopamine- and cAMP-regulated phosphoprotein with a molecular weight of 32,000 Dalton (DARPP-32) to mediate their actions. The renal D1 mediates DARPP-32 activation via a cascade involving cAMP and PKA, and protein kinase C (PKC) activation via phospholipase C. Active DARPP-32 has a specific inhibitory effect on protein phosphatase 1 (PP1), leaving, e.g. Na+,K+-ATPase in a phosphorylated, inactive, state. Thus, dopamine acts as a natriuretic hormone in the mature kidney. Here, we discuss the age dependent distribution and some functional aspects of several parts of the renal dopamine system (dopamine, AADC, COMT, D1 receptor, and DARPP-32) during renal morphogenesis. PMID- 10535322 TI - Discovery of the congenital nephrotic syndrome gene discloses the structure of the mysterious molecular sieve of the kidney. AB - The molecular nature of the glomerular slit diaphragm, the site of renal ultrafiltration, has until recently remained a mystery. However, the identification of the gene affected in congenital nephrotic syndrome has revealed the presence of a novel protein, possibly specific for the slit diaphragm. This protein, which has been termed nephrin, is a transmembrane protein that probably forms the main building block of an isoporous zipper-like slit diaphragm filter structure. Defects in nephrin lead to abnormal or absent slit diaphragm leading to massive proteinuria and renal failure. The discovery of nephrin sheds new light on the glomerular filtration barrier, provides new insight into the pathomechanisms of proteinuria, and even opens up possibilities for the development of novel therapies for this common and severe kidney complication. PMID- 10535323 TI - Influence of fetal environment on kidney development. AB - Several lines of evidence, mostly derived from animal studies, indicate that changes in fetal environment may affect renal development. Besides maternal hyperglycemia or drug exposure, that were recently found to alter nephrogenesis, changes in vitamin A supply to the fetus may prove to be responsible for most of the variations in nephron number found in the population. A low vitamin A status in the fetus may be a major cause of inborn nephron deficit, either as a feature of intrauterine growth retardation or independently of growth retardation. The possibility that vitamin A status may also influence renal vascular development is raised. We suggest that low vitamin A supply to the fetus plays a role in the intrauterine programming of chronic renal disease and hypertension. PMID- 10535324 TI - Cellular pathophysiology of cystic kidney disease: insight into future therapies. AB - Polycystic kidney disease (PKD) is a developmental kidney disorder which can be inherited as either an autosomal dominant trait, with an incidence of 1:50 to 1:1000, or as an autosomal recessive trait with an incidence of 1:6,000 to 1:40,000. Three different genes have now been cloned that are associated with mutations that cause PKD. Two of these are linked to the most common forms of the dominant disease while the third is associated with the orpk mouse model of recessive polycystic kidney disease. Advances in understanding the molecular genetics of PKD have been paralleled by new insights into the cellular pathophysiology of cyst formation and progressive enlargement. Current data suggest that a number of PKD proteins may interact in a complex, which when disrupted by mutations in PKD genes may lead to altered epithelial proliferative activity, secretion, and cell matrix biology. The identification of a unique cystic epithelial phenotype presents new opportunities for targeted therapies. These include targeted gene therapy, gene complementation, and specific immunological or pharmacological interruption of growth factor pathways. PMID- 10535326 TI - Renal cancer genetics: von Hippel Lindau and other syndromes. AB - There have been significant advances in our understanding of the genetic basis of renal carcinogenesis. In particular, research in the last five years has demonstrated a central role for the inactivation of the von Hippel-Lindau gene by mutation or hypermethylation in the formation of the conventional type of renal cell carcinoma. The von Hippel-Lindau syndrome is characterised by germ-line inactivating mutation whereas sporadic renal carcinoma is associated with somatic mutations. Tumour formation is accompanied by loss of the remaining wild-type allele. The biology of the von Hippel-Lindau gene and its normal function continued to be unravelled but a role has been demonstrated for it in the regulation of gene transcription, the regulation of oxygen-dependent genes and their expression and the control of tumour angiogenesis acting via the vascular endothelial growth factor. Another form of familial renal cancer, the hereditary papillary renal cell carcinoma, has been shown to be consequent upon activating mutations of the c-met proto-oncogene. The genetic data continue to enhance our understanding of the biology of this common set of neoplasms. PMID- 10535325 TI - Pax2 in development and renal disease. AB - Pax genes are associated with a variety of developmental mutations in mouse and man that are gene dosage sensitive, or haploinsufficient. The Pax2 gene encodes a DNA binding, transcription factor whose expression is essential for the development of the renal epithelium. Both gain and loss of function mutants in the mouse demonstrate a requirement for Pax2 in the conversion of metanephric mesenchymal precursor cells to the fully differentiated tubular epithelium of the nephron. However, Pax2 expression is down-regulated as cells leave the mitotic cycle. Humans carrying a single Pax2 mutant allele exhibit renal hypoplasia, vesicoureteric reflux, and optic nerve colobomas. Conversely, persistent expression of Pax2 has been demonstrated in a variety of cystic and dysplastic renal diseases and correlates with continued proliferation of renal epithelial cells. Thus, Pax2 misexpresssion may be a key determinant in the initiation and progression of renal diseases marked by increased or deregulated cell proliferation. PMID- 10535327 TI - Mechanisms of epithelial development and neoplasia in the metanephric kidney. AB - Recent studies on the mechanisms of normal epithelial development in the kidney, and on the aetiology of renal neoplasms, are converging to reveal remarkably close relationships between the phenotypes and behaviours of normally-developing and neoplastic cells. Normal renal epithelia arise from two sources; those of the collecting duct system develop by arborisation of an initially-unbranched ureteric bud, in a manner similar to the development of other glandular organs, while epithelial nephrons develop via an unusual mesenchyme-to-epithelial transition. Both types of development require controlled proliferation, cell-cell and cell-matrix interactions, protease activity etc., but of the two tissues, the development of the nephrons is arguably the more complex. It includes many defined stages, signals and checkpoints that ensure that events happen at the right time, and that processes such as proliferation, apoptosis and differentiation are properly balanced. Detailed investigation of renal neoplasms has revealed some to be caused by mutations in molecules with known roles in normal nephrogenesis (e.g. Wilms' tumour and the WT-1 gene, renal cell carcinoma and the c-met receptor tyrosine kinase gene), some to be caused by mutations in genes expressed during normal development (e.g. renal cell carcinoma and the TSC 2 gene, renal cell carcinoma of the clear cell variety and the VHL gene). Furthermore, these and other tumours of unknown aetiology re-express genes such as Pax-2 that are expressed during the normal mesenchyme-to-epithelium transition but are shut off during terminal differentiation. Their re-appearance in tumours suggests that the cells have 'regressed' in an ontogenic sense, and their biology may therefore be understood most clearly by reference to the properties of normal developing cells rather than cells of a mature kidney. PMID- 10535328 TI - Hypothesized neural dynamics of working memory: several chunks might be marked simultaneously by harmonic frequencies within an octave band of brain waves. AB - The capacity of working memory (WM) for up to about seven simple items holds true both for humans and other species, and may depend upon a common characteristic of mammalian brains. This paper develops the conjecture that each WM item is represented by a different brain wave frequency. The binding-by-synchrony hypothesis, now being widely investigated, holds that the attributes of a single cognitive element cohere because electroencephalogram (EEG) synchrony temporarily unifies their substrates, which are distributed among different brain regions. However, thought requires keeping active more than one cognitive element, or WM "chunk," at a time. If there is indeed a brain wave frequency code for cognitive item-representations that are copresent within the same volume of neural tissue, the simple mathematical relationships of harmonies could provide a basis for maintaining distinctness and for orderly changes. Thus, a basic aspect of music may provide a model for an essential characteristic of WM. Music is a communicative phenomenon of "intermediate complexity," more highly organized than the firing patterns of individual neurons but simpler than language. If there is a distinct level of neural processing within which the microscopic physiological activity of neurons self-organizes into the macroscopic psychology of the organism, it might require such moderate complexity. Some of the obvious properties of music--orderly mixing and transitions among limited numbers of signal lines-are suggestive of properties that a dynamic neural process might need in order to organize and reorganize WM markers, but there are a number of additional, nonobvious advantageous properties of summating sinusoids in music like relationships. In particular, harmonies register a stable periodic signal in the briefest possible time. Thus, the regularity of summating sinusoids whose frequencies bear harmony ratios suggests a particular kind of tradeoff between parallel and serial processing. When there are few copresent waves, at EEG frequencies, this sort of parallel coding retains behaviorally meaningful brief periods. A necessary companion hypothesis is that the brain wave frequencies underlying WM are confined to a single octave; that is, the upper and lower bounds of the band are in the ratio of 2:1. This hypothesized restriction, suggested by an empirical property of EEG bands that has been widely reported but rarely commented upon, has the important property of precluding spurious difference rhythms. A restriction to an octave, of "harmonious" frequency-markers for WM items, also seems consistent with a great deal of behavioral data suggesting that WM comprises a rapidly fading trace process in which only up to three or four item-representations are strongly activated simultaneously. There is also an additional, sequential renewal-or-revision process, within which up to another three or four items are being actively refreshed by rehearsal or replaced. Such serial processing may involve a less stringent octave band crowding problem. PMID- 10535329 TI - The early history of the synapse: from Plato to Sherrington. AB - One hundred years ago, in 1897, Sherrington adopted the name synapse. However, the concept of the synapse emerged from considerations of how muscles are contracted and so locomotion affected over a period of 2400 years, from the time of Plato and Aristotle in the 4th century BC to the early part of the 20th century. This early history is considered in the present review. In terms of duration of influence, the early history was dominated by Aristotle's concept of vital pneuma. This was derived from the ether which filled all space, taken in by the lungs, transformed to vital pneuma in the heart, and then conducted in the blood stream to be transmitted to muscles. The vital pneuma then initiated the final phase of the muscle's psyche, that is, its contraction leading to locomotion. Aristotle's ideas had to be modified with the discovery by Galen and his students in the 2nd and 3rd centuries AD that nerves arising from the brain and spinal cord are necessary for the initiation of muscle contraction. They modified the Aristotlean account so that the vital pneuma delivered by blood vessels to the brain was converted there to psychic pneuma, from whence it was conducted along nerves to be transmitted to muscle, so allowing the muscle to contract. There matters rested for about 1300 years until Descartes. Descartes rejected the idea of organs and muscles possessing a psyche with a final cause that was released by the conduction and transmission of psychic pneuma in nerves, emphasising that mechanical explanations must be sought when determining the function of an organ or muscle. He argued in his corpuscular theory that fine particles derived from the blood in the brain, which he gave the unfortunate name of animal spirits, were conducted and transmitted along nerves to enter muscle during transmission, so leading to the increase in width of the muscle fibres, their shortening and contraction. This description was elaborated on in great detail by Descartes, and by his contemporary Borelli, in the 17th century. In the 18th century, Swammerdam carried out a series of brilliant experiments that showed that the Descartes/Borelli theory could not be correct, muscles did not change their volume during contraction, and so could not be contracted by being swollen due to an influx of the corpuscles that made up the animal spirits. These results were published at about the time of the birth of Galvani (1737), whose work was to show that animal spirits were not corpuscular but electrical. The triumph of 19th century physiology, primarily due to Matteucci, du Bois-Reymond and Helmholtz, was to take Galvani's discoveries and show that nerves possessed a potential across their walls that could give rise to a propagating transient potential change which was transmitted to muscles with a finite velocity. Although Sherrington refined the concept and adopted the word "synapse" at the end of that century, it was not until the early part of the 20th century that a conceptual scheme for the synapse involving transmitters and receptors was developed. This clearly delineated a new period following the early history of synaptic transmission. PMID- 10535330 TI - Electrophysiological effects of opioid receptor activation on Syrian hamster suprachiasmatic nucleus neurones in vitro. AB - Entrainment of the dominant circadian pacemaker localised to the hypothalamic suprachiasmatic nuclei (SCN) is mediated partially via the indirect retino geniculo-hypothalamic projection to the SCN, which is presumed to utilise enkephalin and other neurotransmitters, to modulate circadian rhythmicity. In the present study, we have investigated electrophysiologically the currently unknown functional effects of enkephalin, and another opioid receptor agonist morphine, on hamster SCN neuronal activity in vitro. Basal or N-methyl-D-aspartate-evoked firing rates of SCN neurones were generally unresponsive (86%) to the opioid receptor agonists leucine-enkephalin, methionine-enkephalin, or morphine. Washout of the enkephalins or morphine resulted in a rebound excitatory response ("withdrawal activation") in 39% of neurones tested. Withdrawal activation was also elicited by administration of the opioid receptor antagonist naloxone, following pre-exposure to morphine, in 59% of neurones tested. These withdrawal responses were blocked or attenuated by the alpha2-adrenoceptor agonist clonidine, results which suggest a functional interaction exists between opioid receptors and alpha2-adrenoceptors in the SCN. Our observations show that opioid receptor agonists are largely devoid of actions on normal hamster SCN circadian pacemaker activity, while the occurrence of withdrawal responses may have implications on circadian function during withdrawal from opiate abuse. PMID- 10535331 TI - Beneficial effect of renin-angiotensin system for maintaining blood pressure control following subarachnoid haemorrhage. AB - Subarachnoid haemorrhage is a serious condition often accompanied by delayed cerebral ischaemia. Earlier reports have provided evidence suggesting a role for angiotensin II in the development of cerebral vasospasm following subarachnoid bleeding. We sought to examine the influence of angiotensin II blockade with losartan on blood pressure and survival in animals following experimental subarachnoid haemorrhage, induced in conscious rats by injecting homologous blood via a catheter placed along the surface of the brain. We combined measurements of plasma renin activity with blood pressure recording in order to examine renin angiotensin system activation following experimental subarachnoid haemorrhage. Following subarachnoid injury an approximately three-fold increase in plasma renin activity occurred (3.4 +/- 1.0 vs. 10.1 +/- 1.8 ng angiotensin I produced/ml/h, p < 0.01). In animals treated with losartan (20 mg/kg) prior to the induction of subarachnoid haemorrhage blood pressure fell dramatically following the cerebral injury (124 +/- 5 vs. 94 +/- 7 mmHg, p < 0.001), whereas blood pressure remained unchanged in control animals. Survival was markedly reduced in those animals treated with losartan. Given the pronounced decrease in blood pressure and impaired survival following subarachnoid haemorrhage in animals treated with losartan, it would appear that the acute activation of the renin-angiotensin system following this insult is in fact a desirable, compensatory response. PMID- 10535332 TI - Methionine residue 35 is important in amyloid beta-peptide-associated free radical oxidative stress. AB - Amyloid beta-peptide (Abeta), the central constituent of senile plaques in Alzheimer's disease (AD) brain, has been shown to be a source of free radical oxidative stress that may lead to neurodegeneration. In the current study Abeta(1 40), found in AD brain, and the amyloid fragment Abeta(25-35) were used in conjunction with electron paramagnetic resonance spin trapping techniques to demonstrate that these peptides mediate free radical production. The methionine residue in these peptides is believed to play an important role in their neurotoxicity. Substitution of methionine by structurally similar norleucine in both Abeta(1-40) and Abeta(25-35), and the substitution of methionine by valine, or the removal of the methionine in Abeta(25-35), abrogates free radical production and protein oxidation of and toxicity to hippocampal neurons. These results are discussed with relevance to the hypothesis that neurodegeneration in Alzheimer's disease may be due in part to Abeta-associated free radical oxidative stress that involves methionine, and to the use of spin trapping methods to infer mechanistic information about Abeta. PMID- 10535334 TI - The routine obstetric ultrasound. PMID- 10535333 TI - Induction of CD40 on human endothelial cells by Alzheimer's beta-amyloid peptides. AB - Growing evidence suggests that beta-amyloid (Abeta) peptides play a central role in mediating vascular endothelium dysfunction, but the extent to which immune mechanisms are involved in this process remains unclear. To explore such mechanisms, we incubated cultured human aortic endothelial cells (HAEC) with freshly solublized Abeta and examined expression of a central immunoregulatory molecule, CD40, in these cells using reverse transcriptase-polymerase chain reaction, Western immunoblotting, and Flow cytometry. Our results show that treatment of endothelial cells with Abeta1-40, Abeta1-42 or gamma interferon (IFN gamma) results in a dose-dependent induction of endothelial CD40 expression. Furthermore, ligation of endothelial CD40 and simultaneous treatment of human endothelial cells with IFN-gamma or Abeta peptides leads to a significant release of interleukin-1beta (IL-1beta), a marker for endothelial cell activation. Since IL-1beta is an important inflammatory response mediator, these findings suggest that the functional role of Abeta-induced endothelial CD40 may be promotion of the inflammatory cascade in vascular endothelial cells. PMID- 10535336 TI - Appendicitis in pregnancy: diagnosis, management and complications. AB - BACKGROUND: Acute appendicitis is the most common surgical emergency in pregnancy. The purpose of this study is to investigate the clinical presentation, management and outcome in patients who underwent appendectomy during pregnancy. MATERIAL AND METHODS: The case records of 56 women who underwent appendectomy during pregnancy between January 1985 and December 1997 were reviewed and analyzed. RESULTS: The incidence of appendicitis in pregnancy was one in 766 births. The preoperative diagnosis was correct in 75% of the cases. Uterine contractions and a history of diffuse or periumbilical pain migrating to the right lower abdominal quadrant were significantly more frequent among women with appendicitis compared to those patients where the appendectomy revealed a normal appendices. Abdominal pain, nausea, vomiting, leukocyte count, CRP and body temperature were not helpful in establishing the correct diagnosis. There was no maternal morbidity related to the appendectomy. Pregnancy complications were found to be considerable: 4/12 (33%) who underwent appendectomy for appendicitis during the first trimester aborted spontaneously. Second trimester appendectomy for appendicitis was followed by premature delivery in 4/28 (14%). However, no pregnancy complications were observed following third trimester appendectomy for appendicitis. We found no increase in pregnancy complications in cases with perforated appendicitis. CONCLUSION: Appendicitis in pregnancy should be suspected when a pregnant woman complains of new abdominal pain. No laboratory finding was found to be diagnostic for acute appendicitis during pregnancy. Considerable fetal loss was found after appendectomy during pregnancy in the first and second trimester. No increase in pregnancy complications in cases with perforated appendicitis was observed. The combination of symptoms and clinical judgement is still vital in deciding which patient needs surgical treatment. Based on the results in the present study we recommend prophylactic antibiotic treatment in all laparotomies during pregnancy when appendicitis is suspected. PMID- 10535335 TI - Iron status and iron balance during pregnancy. A critical reappraisal of iron supplementation. AB - BACKGROUND: Iron supplementation in pregnancy is a controversial issue. The aim of this review was to summarize the results of relevant papers on this subject. METHODS: Placebo-controlled studies on iron treatment in pregnancy were identified from the Cochrane database. RESULTS: Among fertile women, 20% have iron reserves of >500 mg, which is the required minimum during pregnancy; 40% have iron stores of 100-500 mg, and 40% have virtually no iron stores. The demand for absorbed iron increases from 0.8 mg/day in early pregnancy to 7.5 mg/day in late pregnancy. Dietary iron intake in fertile women is median 9 mg/day, i.e. the majority of women have an intake below the estimated allowance of 12 18 mg/day. Iron absorption increases in pregnancy, but not enough to prevent iron deficiency anemia in 20%, of women not taking supplementary iron. Iron-treated pregnant women have greater iron reserves, higher hemoglobin levels, and a lower prevalence of iron deficiency anemia than placebo-treated women both in pregnancy as well as postpartum. Furthermore, children born to iron-treated mothers have higher serum ferritin levels than those born to placebo-treated mothers. An iron supplement of 65 mg/day from 20 weeks of gestation is adequate to prevent iron deficiency anemia. CONCLUSIONS: In order to avoid iron deficiency in pregnancy, prophylactic iron supplement should be considered. Iron supplements may be administered on a general or selective basis. The selective approach implies screening with serum ferritin in early pregnancy, in order to identify women who can manage without prophylactic iron. PMID- 10535337 TI - Effect of atracurium or pancuronium on the anemic fetus during and directly after intravascular intrauterine transfusion. A double blind randomized study. AB - BACKGROUND: To determine the effect of atracurium or pancuronium on onset and duration of fetal paralysis, movements and heart rate parameters directly after transfusion, using computer analyzed fetal heart rate recording (c-FHR). METHODS: Double blind randomized study of 23 RhD alloimmunized pregnant women requiring an intravascular intrauterine fetal blood transfusion (IUT) between 24 and 36 weeks. Atracurium was injected in 11 fetuses at 17 IUT's and pancuronium in 12 fetuses at 19 IUT's. For statistical analysis the Mann-Whitney test was used. RESULTS: No statistical differences were found in fetal heart rate and movements between both groups before transfusion. The fetal movements returned more rapidly in the atracurium group when compared to the pancuronium-group (median 24 vs. 57 min, range 6-55 vs. 4-220; (p<0.02). Fetal movements did not hamper the procedure in any case. The atracurium group showed significantly more fetal movements (p<0.01), more accelerations (0<0.05) but no significant reduction of fetal heart rate variability directly after transfusion which was in direct contrast to the pancuronium group. CONCLUSIONS: Neuromuscular blockade with atracurium produces sufficient paralysis for intrauterine transfusion with minimal disturbance of the parameters used to monitor fetal wellbeing after the procedure. Although the routine use of fetal paralysis during IUT may be questionable, we believe that when it is necessary the use of atracurium is the better choice. PMID- 10535338 TI - The effect of betamethasone on fetal biophysical activities and Doppler velocimetry of umbilical and middle cerebral arteries. AB - OBJECTIVE: Preliminary reports suggest that antenatal steroid administration may confound the assessment of fetal well-being by suppressing biophysical activities, consequently drug-induced effects could prompt unwarranted delivery of premature fetuses. The purpose of this study was to examine the effect of antenatal betamethasone administration on fetal biophysical activities and Doppler flow indices of the umbilical and middle cerebral circulation. METHODS: Forty women at risk of premature delivery between 27-32 weeks gestation (mean 30.2 weeks) received two consecutive doses of intramuscular betamethasone, 24 hours apart. Ultrasonographic observations of fetal behavior for 30 minute periods and Doppler examination of the umbilical and cerebral arteries were performed prior to (0 hours), 48 hours after, and 96 hours after administration of the first dose. To account for fetal circadian rhythms and maternal prandial status, all examinations were carefully timed and performed between 1-4 pm. Analysis of Variance, chi-square test and Fisher's Exact test were used for statistical analysis, as appropriate. RESULTS: Nine patients were excluded from analysis due to delivery prior to completion of all examinations. Number of breathing episodes as well as total breathing time at 48 hours decreased by 83.0% (p<0.01) and 90.4% (p<0.01), respectively, at 48 hours in comparison to baseline. Fetal limb and trunk movements decreased by 53.2% (p<0.01) and 48.6% (p<0.01), respectively. Amniotic fluid volume and fetal tone were normal in all patients. At 48 hours, 14 of 31 fetuses and 4 of 31 fetuses had a biophysical profile score of 6/8 and 4/8, respectively, in comparison to 0 of 31 and 0 of 31 at 0 hours (p<0.05 and p<0.001, respectively). All parameters returned to baseline values at 96 h. Pulsatility indices of umbilical and middle cerebral arteries remained unchanged at 48 hours and 96 hours (p=NS). CONCLUSIONS: Betamethasone induces a profound, albeit transient, suppression of fetal breathing, limb and trunk movements, resulting in decreased biophysical profile scores. Awareness of this drug-induced effect might prevent unnecessary iatrogenic delivery of preterm fetuses. PMID- 10535339 TI - Clinical application of maternal serum cytokine determination in premature rupture of membranes--interleukin-6, an early predictor of neonatal infection? AB - BACKGROUND: In cases of premature rupture of membranes (PROM), an early detection of fetal infection is necessary in order to weigh infectious complications against prematurity. As routine parameters (leukocytes, C-reactive protein (CRP), fever, and fetal tachycardia) lack satisfactory sensitivity and specificity, this study evaluates whether the determination of interleukin-6 (IL-6), interleukin-8 (IL-8) or soluble interleukin-2 receptor (IL-2R) in maternal serum could supplement or replace routine inflammation parameters. METHODS: In this prospective study results of clinical and laboratory parameters were investigated with respect to neonatal infection in 71 patients with PROM. IL-6, IL-8 and IL-2R were determined by enzyme immunoassays. RESULTS: Best specificity and sensitivity could be demonstrated for CRP and IL-6. Both elevation of CRP and IL-6 correlated significantly (p<0.01 and p<0.001, respectively) with the onset of neonatal infection. At a cutoff of 11 pg/ml, IL-6 reaches a sensitivity of 81% and a specificity of 76%; CRP a specificity of 76% (cutoff 1.2 mg/dl) and a sensitivity of 56%. In 4/16 (25%) cases developing neonatal infection, IL-6 increased earlier than CRP. IL-8 and IL-2R results showed a less significant correlation with fetal outcome. CONCLUSIONS: Determination of IL-6 in maternal serum can significantly contribute to an earlier detection of fetal infection in patients with PROM. PMID- 10535340 TI - Modified cervical cerclage in pregnant women with advanced bulging membranes: knee-chest positioning. AB - BACKGROUND: Emergent cervical cerclage is often unsuccessful in patients with severely protruding fetal membranes for the following reasons: (1) difficulty in pushing the bulging membranes into the cervix; (2) risk of premature rupture of membranes during the operative procedures; and (3) displacement of the cerclage tape by the shortened cervix. METHODS: We performed Shirodkar's cervical cerclage with a slight modification. Using a metreurynter, a primary suture was made with the patient placed in the knee-chest position; a Shirodkar's cerclage was then performed in the dorsal lithotomy position. RESULTS: Since 1992, we have performed this procedure on five patients whose fetal membranes bulged into the vagina, at a gestational period of 20-24 weeks. When a patient was placed in the knee-chest position, the bulging membranes intruded spontaneously into the uterine cavity, but the procedure did not cause the membranes to rupture in any patient. The procedure prolonged the pregnancy to longer than 36 gestational weeks in two of the five patients. Three infants survived intact. CONCLUSIONS: Our promising experience warrants further studies on the advantages of the knee chest position during the application of cervical cerclage in pregnant women who present with fetal membranes that bulge through a widely dilated cervix. PMID- 10535341 TI - Nifedipine versus ritodrine for suppression of preterm labor; a meta-analysis. AB - BACKGROUND: Since large randomized clinical trials comparing the effectiveness of nifedipine and ritodrine in the suppression of preterm labor are lacking, we performed a meta-analysis on the subject. METHODS: We searched the databases Medline and EMBASE using the keywords 'nifedipine', 'ritodrine' and 'randomized' or 'randomised'. The studies were scored for blinding, method of randomization and type of analysis ('intention-to-treat' versus 'par protocol'). Subsequently, two by two tables were constructed using 'delay of labor by 48 hours or more', 'delay of labor beyond 36 weeks gestation', perinatal mortality, respiratory distress syndrome and admission to a neonatal intensive care unit as end points. Homogeneity between the studies was tested with a Breslow-Day test. Pooled odds ratios were calculated in case homogeneity could not be rejected. RESULTS: We could detect ten studies that were published between 1986 and 1998, incorporating data of 681 patients. Nifedipine reduced the risk of delivery within 48 hours compared to ritodrine, but this difference was not statistically significant (odds ratio 0.85, 95% confidence interval 0.54 to 1.1). Nifedipine also reduced the risk of delivery before 36 weeks compared to ritodrine, and this difference was statistically significant (odds ratio 0.59, 95% confidence interval 0.39 to 0.90). We are not aware of studies reporting on long-term outcome. CONCLUSION: Since studies reporting on long-term outcome are lacking, the choice between nifedipine and ritodrine can only be based on obstetrical and short-term neonatal outcomes. From that perspective, nifedipine should be the drug of first choice for the suppression of preterm labor. PMID- 10535342 TI - Post-cesarean section morbidity in HIV-positive women. AB - BACKGROUND: The present work is an audit of post-cesarean section morbidity in HIV-positive women in the tertiary teaching hospital La Fe, Valencia, Spain. STUDY DESIGN: Retrospective case-control study. SUBJECTS: Forty-five HIV-positive pregnant women and 90 appropriately matched controls, delivered by cesarean section in the same hospital and managed using a uniform protocol. MAIN OUTCOME MEASURES: The duration of stay in hospital after cesarean section, the need for postoperative antibiotics and the incidence of major and minor puerperal complications. Baseline characteristics of HIV-positive women were also analyzed in relation to the morbidity after surgery. STATISTICAL ANALYSIS: Chi-square analysis for categorical data and parametric and non-parametric tests for numerical data, where appropriate. RESULTS: Most HIV-positive women (86.7%) had a complicated recovery after surgery. A longer duration of stay in hospital (p<0.0005) and a greater incidence of major (p<0.003) and minor (p<0.00001) postoperative complications were observed in the HIV-positive group compared to the control group. HIV-positive women with > or =500 CD4 lymphocytes/mm3 had less post-cesarean section morbidity CONCLUSIONS: A greater post-cesarean section morbidity was found in HIV-positive women compared to the control women. Immunological status of HIV-positive women may be important in predicting puerperal morbidity after surgery. PMID- 10535343 TI - Induction of labor with mifepristone--a randomized, double-blind study versus placebo. AB - OBJECTIVE: To evaluate the efficacy of mifepristone in inducing labor in women with an unripe cervix, its effect on the cervix and on the status of the newborn. METHODS: In a prospective double-blind study, 36 post-term pregnant women with a Bishop score of 5 or less received either 400 mg mifepristone (n=24) or placebo (n=12). If, 48 hours after the treatment was started, labor had not begun or the Bishop score was 5 or less, the women were given 0.5 mg prostaglandin E2 intracervically, a treatment which was repeated 12 hours later, if necessary. RESULTS: During the first 48 hours following treatment, 19 (79.2%) of the women treated with mifepristone and two of the women (16.7%) treated with placebo went into labor. In addition, one and three women, respectively, had a ripe cervix at the end of the 48h period. The overall success rate was thus 83.3% for mifepristone and 41.7% for placebo (p=0.008; OR 14.8; 95% CI 2.1-107.6). The median time from the start of treatment to delivery was also shorter (mifepristone 36h23' and placebo 53h17'). Treatment with intracervical PGE2 was needed more often after the placebo. The duration of labor, however, tended to be shorter after placebo than after mifepristone in the women who delivered vaginally. The frequencies of instrumental delivery were similar in both treatment groups. The median Apgar score was slightly lower at 1 minute (p<0.05) following mifepristone treatment, but did not differ at 5 and 10 minutes. There was no difference between the two treatment groups in the umbilical pH at delivery. CONCLUSION: The results of the present study show that mifepristone is a simple and effective treatment for inducing labor in post-term women with an unripe cervix. PMID- 10535344 TI - The influence of prepregnancy body mass index on labor complications. AB - BACKGROUND: To investigate the influence of Body Mass Index on the incidence of labor complications in a population of women with a normal pregnancy. MATERIAL AND METHODS: From a local database, information on maternal weight and height was extracted concerning 4258 women who had an uncomplicated pregnancy. After calculation and stratification with respect to Body Mass Index, this was retrospectively related to labor interventions and complications. RESULTS: High Body Mass Index was related to more oxytocin infusion and early amniotomy, but not to vacuum extraction or cesarean section. Primary inertia and, to a minor degree, cephalopelvic disproportion and secondary inertia were seen more often in women with high Body Mass Index. CONCLUSIONS: Overweight (25.0<=BMI<30.0) and obesity (BMI>=30.0) are only weak predictors of labor complications, given a normal pregnancy. However, the heavy use of labor augmentation indicates that obese women should not be recommended to give birth in an ABC-clinic or at home. PMID- 10535345 TI - Use of obstetric forceps in Finland today--experience at Vaasa Central Hospital 1984-1998. AB - BACKGROUND: Forceps delivery has become rare in Finland since the introduction of the vacuum extractor. Our aim was to survey the number of forceps deliveries in Finland and analyze our own material of 130 forceps deliveries during a 15-year period between 1984 and 1998. During this period there were 17,887 deliveries at Vaasa Central Hospital. METHODS: A retrospective study of 130 forceps deliveries and 11 trial forceps cases, which subsequently resulted in a cesarean section. RESULTS: There was no maternal or neonatal mortality. In 39 cases a cesarean section could be avoided by use of forceps after a failed vacuum extraction. Only in one case was maternal morbidity regarded as serious. There was no serious neonatal morbidity. Anal sphincter ruptures occurred in three cases (2.3%). All the women in the trial forceps group were nulliparous, in 73% of these the fetus was in a persistent occipito-posterior position. Failed vacuum extraction and trial forceps did not significantly influence neonatal outcome. CONCLUSIONS: Forceps delivery appears to be a safe alternative in our setting. PMID- 10535346 TI - Interruption of early pregnancy with mifepristone in combination with gemeprost. AB - BACKGROUND: Mifepristone in combination with prostaglandin has been used since 1988 for induction of early abortion. The aim of the present investigation was to assess the tolerance and efficacy of 600 mg. mifepristone orally followed by gemeprost 1 mg. vaginally either 24 hours (group one) or 48 hours (group two) later. METHODS: Sixty-four healthy women applying for abortion within the first 8 weeks of pregnancy were randomly allocated to one of the two treatment groups. Intrauterine pregnancy and gestational age were verified by ultrasonography. Symptoms after administration of mifepristone and gemeprost were recorded, and the patients observed at the hospital for at least three hours after prostaglandin-insertion. Blood samples for blood group, hemoglobin, beta-chorion gonadotrophin, aspartate-aminotransferase and creatinine were drawn. RESULTS: Outcome was established by gynecological examination, the level of beta-hCG and ultrasonography, at visits one, two and if necessary three to four weeks later. Surgical curettage was performed in case of incomplete abortion, of which there were four in the 24-hour interval group and five in the 48-hour interval group with a success rate (complete abortion) of 55 out of 64 patients (86%). CONCLUSIONS: There was no difference in efficacy or side effects whether the prostaglandin was administered 24 or 48 hours after mifepristone intake, which suggests that the treatment period can be reduced from the conventional 48 hours. PMID- 10535347 TI - The significance of an 'insufficient' Pipelle sample in the investigation of post menopausal bleeding. AB - BACKGROUND: Out-patient endometrial sampling is a commonly performed procedure in the investigation of post-menopausal bleeding. We hypothesized that an 'insufficient' Pipelle sample reliably reflected the absence of serious endometrial pathology. METHODS AND RESULTS: A review of 141 consecutive cases reported as 'insufficient' for diagnostic purposes revealed 29 (20%) cases to have uterine pathology after secondary investigation. These included two cases of endometrial carcinoma and two other cases of uterine malignancy. CONCLUSION: These results do not support the initial hypothesis and suggest that in women presenting with post-menopausal bleeding, an 'insufficient' sample is an indication for further investigation. PMID- 10535348 TI - Childhood and adolescent ovarian malignant tumors in Israel. A nationwide study. AB - OBJECTIVES: To determine the incidence of ovarian malignant tumors in childhood and adolescence, to ascertain the frequency distribution of the various tumor types and to assess time trends in Israel on a nationwide basis. METHODS: The study group included all Israeli Jewish patients < or = 19 years old with histologically confirmed ovarian malignancies, diagnosed in Israel from 1970 to 1994. Data were obtained from the Israel Cancer Registry. The effects of age at diagnosis and period of diagnosis were analyzed using the Poisson regression. RESULTS: Among the 82 patients identified, the most frequent tumors (72.0%) were of germ cell origin and among those about one third were dysgerminomas. Epithelial tumors were diagnosed in 26.6% of the patients and most of these were borderline malignancies. The incidence rate (IR) for the total group of ovarian malignancies in the 0-19 age group was 0.52 and for ages 5-19 it was 0.71 per 100,000. After adjustment for age, a significant linear trend for a decrease of germ cell tumors over time was found, stemming from a decrease of dysgerminomas. A significant trend for increase in the IRs with age was also found. In addition, a steep rise in the age specific IRs of epithelial borderline tumors was noted in the last 5 year period. CONCLUSIONS: The IRs of ovarian malignancies in childhood and adolescence in Israel, as in other countries, is very low as compared to adults and the most common tumors are of germ cell origin while malignant epithelial tumors are very rare. A time period effect in the germ cell tumors that resulted from an inexplicable significant decrease in the age specific IRs of dysgerminomas, was observed. A significant increase in borderline tumors was also noted and may be attributed to greater awareness of pathologists to this entity. PMID- 10535349 TI - Pathological indications for conservative therapy in treating cervical cancer. AB - BACKGROUND: Neodymium-ytlium, argon, gadolinium (Nd-YAG) laser conization was performed in 366 patients with carcinoma in situ (CIS), and 198 with microinvasive carcinoma (MIC), and 29 with early invasive cancer (IC). The diagnostic accuracy was 72.5% in the CIS group, and 74.6% in the MIC and early IC group. RESULTS: All of the patients who obtained complete excision are now alive and doing well following the conization, with no evidence of recurrence of the disease. Incomplete excision was found in 21.7% of the MIC, 50.0% of the IC with 3 mm or less invasion, 22.2% of the IC with 3.1-4 mm, 60.0% of the IC with 4.1-5 mm invasion, 100% of the IC with 5 mm or more invasion. The residual rate of the MIC and IC with 4 mm or less invasion was 8.5%, lower than 62.5% of the IC with 4.1-5 mm invasion. The cure rate by conization in the patients with IC of 4.0 mm or less was much higher than that in the patients with IC of over 4.1 mm. CONCLUSIONS: These results suggested that the indication of laser conization for early IC was the case within 4 mm invasion, completely excised. PMID- 10535350 TI - The efficacy of intratubal silicone in the Ovabloc hysteroscopic method of sterilization. PMID- 10535351 TI - Metastasis to the breast from an ovarian carcinoma. PMID- 10535352 TI - The relevance of intrapartum fetal pulse oximetry in the presence of fetal cardiac arrhythmia. PMID- 10535354 TI - The role of the E-cadherin complex in gastrointestinal cell differentiation. PMID- 10535353 TI - One stillborn and one severely hydropic twin due to parvovirus B19 infection; successful outcome of the surviving twin. PMID- 10535355 TI - Effects of glycosaminoglycans on proliferation of epithelial and fibroblast human malignant mesothelioma cells: a structure-function relationship. AB - Proteoglycans interact with other effective macromolecules regulating a variety of cellular events via their glycosaminoglycan (GAG) chains. The effects of all known glycosaminoglycans (GAGs) produced by normal cells and tissues on the proliferation of two human malignant mesothelioma cell lines, one with fibroblast like morphology and the other with epithelial differentiation - both able to produce hyaluronan (HA), galactosaminoglycans (GalAGs) and heparan sulphate (HS) containing proteoglycans - have been studied. Cell proliferation was assessed by measuring [3H]thymidine incorporation and cell number. GalAGs, i.e. chondroitin sulphates (CSs) and dermatan sulphate (DS), strongly stimulate the proliferation of fibroblast-like cells in a dose-dependent manner (170-250% at 100 microg/ml), independently of their sulphation pattern. In epithelial cells, however, only DS stimulates cell proliferation. The effects of CSs on proliferation of epithelial cells are not depended on their sulphation pattern. Thus, CSs either with -[GlcA GalNAc-(-6-O-SO(3)-)]- or -[GlcA-GalNAc-(-4-O-SO(3)-]- as the commonest unit, had no significant effect. L-Iduronic acid (IdoA)-rich heparin and fast-moving HS (fm HS), a HS fraction with a heparin-like structure, had significant antiproliferative effects on mesothelioma cells of both types (30-70% at 1.0 microg/ml and 85-90% at 100 microg/ml, respectively). GlcA-rich HS, however, had no significant effects. HA inhibits only the proliferation of fibroblast-like cells by 25% at 50 and 100 microg/ml. Keratan sulphate suppresses cell proliferation (10-30%) in both cell lines. In the view of these findings, a structure-function relationship of GAGs on cell proliferation of the two human malignant mesothelioma cell lines is discussed. Other factors, such as chain conformation and geometry, as well as interactions of growth factors with GAGs, possibly involved in the regulation of cell proliferation, are also discussed. PMID- 10535356 TI - Lipid-induced changes in vascular smooth muscle cell membrane fluidity are associated with DNA synthesis. AB - In the present study, we examined whether changes in the membrane fluidity of vascular smooth muscle cells (VSMCs) alter their DNA synthesis. For this purpose, the membrane fluidity of the cells was modulated after treatment of VSMCs with 1,2-dioleoyl phosphatidylcholine (PC). Treatment of VSMCs with 1,2-dioleoyl PC rich medium containing 10% heat-inactivated human serum and 3 mg/ ml 1,2-dioleoyl PC for 24 h resulted in an increase in VSMC membrane fluidity at all temperatures from 15 degrees to 40 degrees C as well as a 51% inhibition of DNA synthesis, compared with untreated cells. Remarkably, enrichment of VSMCs with 1,2-dioleoyl PC/cholesterol-rich medium containing 10% human serum, 3 mg/ml 1,2-dioleoyl PC and 2 mg/ml cholesterol restored both membrane fluidity and DNA synthesis to the levels of untreated cells. The present findings show an inverse association between increased membrane fluidity and cellular DNA synthesis. PMID- 10535357 TI - Association of pS2 (TFF1) release with breast tumour proliferative rate: in vitro and in vivo studies. AB - Although cytosolic expression of the protein pS2 (TFF1) is considered to be a marker of oestrogen receptor (OR) function, there exists some clinical data to suggest an inverse relationship of cytosolic pS2 to tumour proliferation. Although secreted from breast cancer cells, the relationship of pS2 secretion to tumour natural history has been little studied. The mechanisms and kinetics of pS2 release and its relation to tumour cell proliferation were studied in a human breast cancer cell line MCF-7 and verified in a preliminary clinical study. Stimulation by stripped serum or oestradiol resulted in parallel increases of proliferation and pS2 release in both time course and dose-response experiments. Direct pharmacological alterations of proliferation were followed by identical changes in pS2 release. The relationship between serum pS2 levels and tumour proliferative activity when analysed as a function of steroid status showed a slope of 0.56 in OR+ vs. 0.19 in OR- tumours. It is concluded that pS2 release from breast cancer cells is associated with their proliferation and measurement of serum pS2 levels might be a good predictor of tumour proliferative state and could permit noninvasive monitoring of this tumour parameter. PMID- 10535359 TI - Purine nucleotides modulate proliferation of brown fat preadipocytes. AB - The hypothesis that purine nucleotides and nucleosides affect brown fat preadipocyte proliferation was tested using isolated rat interscapular brown fat preadipocytes in culture. Daily addition of 100 microM adenosine triphosphate (ATP) (n = 4) to cultures enhanced the relative DNA content by 1.5-fold compared to control cultures (P < 0.05) measured using CyQUANT-GR fluorescence. Higher concentrations of ATP inhibited growth and 500 (n = 2) or 1000 microM ATP (n = 3) almost completely inhibited growth. ATP (100 microM) did not affect while 250 1000 microM ATP decreased protein content relative to control cultures. Adenosine (100 microM; n = 3) did not affect DNA or protein content, but 500 microM and 1000 microM adenosine suppressed brown adipocyte proliferation and inhibited protein synthesis. Cultured brown adipocytes quickly removed or degraded ATP in the culture media as determined by luciferin-luciferase bioluminescence, suggesting that the inhibitory effects of high ATP concentrations may result from its breakdown to adenosine. The results support the conclusion that ATP promotes and adenosine inhibits brown adipocyte proliferation. PMID- 10535358 TI - The c-myc gene regulates the polyamine pathway in DMSO-induced apoptosis. AB - It is accepted that apoptosis is a gene-controlled process of cellular self destruction. It occurs during physiological regulation and in pathological situations in the life of a cell. In the immune system, several different intracellular and extracellular factors have been associated with the induction of apoptosis, and the final responses depend on the cell system and the acquired signals. In lymphoid cells, dexamethasone-induced apoptosis is associated with c myc downregulation in cells that remain in G0-G1 until the point of death. Ornithine decarboxylase (ODC), a key enzyme involved in polyamine biosynthesis, is regulated by c-myc, which is a transcriptional activator implicated not only in the control of cell proliferation and differentiation but also in programmed cell death. As dimethylsulphoxide (DMSO) induces apoptosis in the RPMI-8402 human pre-T cell line, the present study analysed the involvement of the c-myc proto oncogene and polyamine pathway as mediators of apoptosis. Cell growth, programmed cell death, c-myc expression, ODC activity and intracellular polyamine content were detected after DMSO and difluoromethylornithine (DFMO) treatment. DMSO treated cells exhibit a decrease in ODC activity and polyamine levels associated with cell growth arrest and programmed cell death induction. The expression of c myc proto-oncogene, as its mRNA or protein, is specifically down-regulated. DFMO, a well defined polyamine biosynthesis inhibitor, completely blocks ODC activity, resulting in growth inhibition but not apoptosis. Moreover, in these samples no evidence of changes of c-myc expression were found. The results obtained suggest that, in RPMI-8402 cells, DMSO provokes a c-myc-dependent decrease of ODC activity followed by a depletion of intracellular polyamine levels, associated with programmed cell death and cell growth arrest. PMID- 10535360 TI - The effect of tobacco smoking and of betel chewing with tobacco on the buccal mucosa: a cytomorphometric analysis. AB - The effect of tobacco use on the buccal mucosa has been assessed by cytomorphometry. Cell and nuclear diameters (CD, ND) of exfoliated oral squames were measured in tobacco smokers (S), betel chewers with tobacco (C) and those with a combined habit (S+C). Non-users (NU) served as controls. The mean CD values in S, C, S+C and NU were: 50.8 (+/-0.47), 49.39 (+/-0.48), 49.12 (+/-0.47) and 51.87 (+/-0.76) microm, and the mean ND values were: 8.83 (+/-0.07), 8.61 (+/ 0.08), 8.72 (+/-0.10) and 8.33 (-/+ 0.09) microm, respectively. The least significant difference procedure (LSD at P=0.05) showed a significant reduction for CD in C and S+C and an increase for ND in all three habit groups, compared to the controls. This study shows that the use of tobacco influences the cytomorphology of the normal buccal mucosa. Betel chewing with tobacco influences the ND and CD, while smoking influences only the ND. PMID- 10535361 TI - Telomerase activity and in situ telomerase RNA expression in oral carcinogenesis. AB - To investigate the role of telomerase in oral carcinogenesis, we assayed telomerase activity in various oral tissues by a modified telomeric repeat amplification protocol (TRAP) analysis. Also, using digoxigenin-labeled probes, we measured the in situ expression of human telomerase RNA component (hTR) in paired oral squamous cell carcinomas (OSCC) and adjacent non-cancerous matched tissue (NCMT). We detected telomerase activity in three OSCC cell lines, but not in primary oral keratinocytes. In patient samples, most OSCC (36/42, 86%) and oral premalignant lesions (8/12, 67%) possessed telomerase activity. In addition, 6 of 27 (22%) NCMT contained weak telomerase activity. In situ hybridization showed that hTR was expressed in almost all OSCC (23/27, 85%) as well as in the majority of NCMT (20/25, 80%). In most cases, accumulation of hTR was observed both in the nucleus and cytoplasm of epithelial cells. A correlation between hTR expression and more advanced tumor grade was observed. The appearance of telomerase activation and hTR expression during oral carcinogenesis was different. This study indicates that the activation of telomerase is an early and frequent event in OSCC. PMID- 10535362 TI - Loss of differentiation of 4NQO-induced rat malignant oral keratinocytes correlates with metastatic dissemination and is associated with a reduced cellular response to TGF-beta1 and an altered receptor profile. AB - This study examined the metastatic capacity of clonal populations of 4NQO-induced rat malignant oral keratinocytes following orthotopic transplantation to athymic mice. Polygonal and spindle cells formed well-differentiated squamous cell carcinomas (keratin positive and vimentin negative) and undifferentiated spindle cell tumours (keratin negative and vimentin positive), respectively, in almost 100% of animals at the site of inoculation (floor of mouth). Transplantation of 5x 10(6) cells of either cell type at high cell density resulted in approximately 50% of animals forming pulmonary metastases. By contrast, inoculation of 2x 10(6) differentiated polygonal cells resulted in the formation of significantly fewer pulmonary metastases than the undifferentiated spindle cells. A single well differentiated clone of polygonal cells and 3 of 4 of the undifferentiated spindle cell lines produced comparable levels of TGF-beta1. One undifferentiated spindle cell line expressed significantly more TGF-beta1 and, following transplantation orthotopically, fewer animals formed pulmonary metastases despite the formation of primary tumours in almost all grafted animals, suggesting that TGF-beta1 can act as a tumour suppressor in this cell type. All cell lines produced comparable amounts of TGF-beta2. The clones of polygonal cells were markedly inhibited and the spindle cells were only partially inhibited by exogenous TGF-beta1. Both cell types expressed high-affinity TGF-beta cell surface receptors; the ratio of type I to type II TGF-beta receptors was 1.0:<3.0 in the spindle cells and 1.0:17.9 in the polygonal clone. The results suggest that differentiated rat malignant oral keratinocytes are less aggressive and have a decreased potential to metastasise than their undifferentiated spindle cell counterparts. This may be attributable, in part, to a change in TGF-beta receptor profile leading to the partial loss of response to exogenous TGF-beta1. PMID- 10535363 TI - Glycaemic disorders in denture stomatitis. AB - The prevalence of glycaemic disorders was investigated in native Upper-Austrians with Candida-associated denture stomatitis. All patients with previously unknown diabetes mellitus were subjected to an oral glucose tolerance test (OGTT) and as a result diabetes was diagnosed in 13% of the patients over 50 years of age. Thirty-five percent of all inspected patients over 50 years of age with denture stomatitis had type 2 diabetes mellitus and 36% had impaired glucose tolerance (IGT). The correlation between Candida-associated denture stomatitis and diabetes mellitus indicates a means for the early diagnosis of diabetes. Hyperglycaemia could not be a predisposition to denture stomatitis, since all patients with denture stomatitis in the age-bracket 26-50 years were without diabetes and only very few of the older patients with diabetes were obese. The correlation between Candica-associated denture stomatitis and type 2 diabetes mellitus could be traced back to a reduced resistance to Candida that preceded the diabetes. PMID- 10535364 TI - Collagen fibres in the wall of odontogenic keratocysts: a study with picrosirius red and polarizing microscopy. AB - The collagen in the walls of 15 keratocysts was studied histochemically by staining sections with picrosirius red and examining them with polarizing microscopy. This was compared to 15 cases of dentigerous cyst and 15 cases of radicular cyst. Polarization colours of the collagen fibres were recorded according to their width. No differences were found between the polarization colours of thin fibres (<0.8 microm) in all three lesions; the polarization colours of thick fibres (1.6-2.4 microm) in keratocysts were significantly more greenish-yellow when compared with those of dentigerous cysts and radicular cysts. The staining of the collagen fibres in the keratocysts is similar to that reported in odontogenic neoplasms, which suggests that the stroma of keratocysts could be regarded not just as a structural support of the cyst wall, but as playing a part in the neoplastic behaviour of the cyst. PMID- 10535365 TI - Calcifications of the disc of the temporomandibular joint. AB - Calcified lesions of the temporomandibular joint discs obtained from 135 human cadavers were studied. Calcifications were observed in 92 of 250 discs by soft x ray radiography. Studies by light and electron microscopy and x-ray diffraction revealed that there were two different types of calcification in the discs: apatite crystal deposition with or without ossification, and calcium pyrophosphate dehydrate (CPPD) crystal deposition. Calcifications were recognized more frequently posteriorly than anteriorly, and were related to disc perforation. The results of this investigation suggest that disc degeneration, which may occur as a result of aging or mechanical stress, causes calcifications. PMID- 10535366 TI - Primary intraosseous carcinoma of the mandible with probable origin from reduced enamel epithelium. AB - This report describes a case of primary intraosseous carcinoma (PIOC) arising de novo in the mandible. The patient was a 74-year-old woman and an early PIOC was found incidentally during histopathological examination of the pericoronal tissue obtained at extraction of a deeply impacted third molar. The curetted soft tissues consisted of a microinvasive, keratinizing squamous cell carcinoma with scattered foci of carcinoma cells showing calcification; stromal osseous metaplasia was also observed. After additional treatment, the patient has remained free of disease for 2 years. Since the tumor was completely enclosed in the bone, the only identifiable source of the present PIOC is the reduced enamel epithelium. Despite its rarity, this case emphasizes the importance of careful histologic examination of all apparently innocuous dental follicles that are removed surgically. PMID- 10535368 TI - A perspective on postoperative atrial fibrillation. AB - Some patients develop atrial fibrillation after any type of surgery, some patients develop atrial fibrillation only after cardiac surgery, and still other patients never develop postoperative atrial fibrillation. Despite the inherent difficulty in identifying the causes of postoperative atrial fibrillation, several important observations have been made during the recent past that may play a role in treating or even preventing this common and serious postoperative problem. This communication provides a framework within which the problem of postoperative atrial fibrillation can be evaluated and treated. PMID- 10535367 TI - Maxillofacial osteonecrosis in a patient with multiple "idiopathic" facial pains. AB - Previous investigations have identified focal areas of alveolar bone tenderness, increased mucosal temperature, abnormal anesthetic response, radiographic abnormality, increased radioisotope uptake on bone scans, and abnormal marrow within the quadrant of pain in patients with chronic, idiopathic facial pain. The present case reports a 53-year-old man with multiple debilitating, "idiopathic" chronic facial pains, including trigeminal neuralgia and atypical facial neuralgia. At necropsy he was found to have numerous separate and distinct areas of ischemic osteonecrosis on the side affected by the pains, one immediately beneath the major trigger point for the lancinating pain of the trigeminal neuralgia. This disease, called NICO (neuralgia-inducing cavitational osteonecrosis) when the jaws are involved, is a variation of the osteonecrosis that occurs in other bones, especially the femur. The underlying problem is vascular insufficiency, with intramedullary hypertension and multiple intraosseous infarctions occurring over time. The present case report illustrates the extreme difficulties involved in the diagnosis and treatment of this disease. PMID- 10535369 TI - Postoperative atrial arrhythmias: risk factors and associated adverse outcomes. AB - Atrial arrhythmias are the most common complication of cardiac surgical procedures today. Because of the additional hospital costs associated with these arrhythmias, owing to increased use of antiarrhythmic medications, diagnostic studies, and prolonged hospitalization, this subject continues to draw the interest of cardiac surgeons, cardiologists, insurance companies, and hospital administrators, among others. Despite many clinical studies, there is still no consensus regarding the best prevention strategy for these arrhythmias. We recently reviewed our experience with these arrhythmias, with the intent of identifying risk factors for the development of these arrhythmias and identifying any associated, potentially adverse, outcomes. We found that the incidence of postoperative atrial arrhythmias has increased to more than 35% in recent years. Risk factors for the development of postoperative atrial arrhythmias include increasing patient age, preoperative use of digoxin, history of rheumatic heart disease, chronic obstructive pulmonary disease, and increasing aortic cross-clamp (ischemic) time. Among patients with postoperative atrial arrhythmias, there was an increased rate of perioperative stroke, increased frequency of ventricular arrhythmias, increased need for the placement of a permanent pacemaker, and prolongation of the intensive care unit and total hospital length of stay. PMID- 10535370 TI - Potential preoperative markers for the risk of developing atrial fibrillation after cardiac surgery. AB - Postoperative atrial arrhythmias after cardiac surgical procedures are common, with a reported overall incidence of approximately 50%. The pathophysiological mechanisms responsible for atrial fibrillation after a cardiac procedure remain unclear, although several clinical studies published during the past decade have identified certain preoperative risk factors associated with postoperative atrial fibrillation. In this study, we attempted to identify the histopathological changes in atrial cardiomyocytes that might predict the development of atrial fibrillation during the postoperative period. Atrial tissue from 60 patients was sampled before and after a cardiopulmonary bypass. Fifteen patients (25%) developed postoperative atrial fibrillation. The only clinical independent risk factor for the development of postoperative atrial fibrillation was chronic obstructive pulmonary disease (COPD) (P = .037). Histologically, there were 3 findings in the atrial myocardium that were more common in patients who developed postoperative atrial fibrillation: (1) vacuolation size (P = .017), (2) vacuolation frequency (P = .0136), and (3) lipofuscin content (P = .013). The identification of these histological markers for the development of postoperative atrial fibrillation may contribute not only to our understanding of the underlying pathophysiology that leads to postoperative atrial fibrillation but also to a method of preventing this troublesome complication of cardiac surgery. PMID- 10535371 TI - Treatment of postoperative atrial fibrillation. AB - Postoperative atrial fibrillation can occur in approximately 30% of patients. Although often a benign complication, it can result in significant morbidity and prolong hospitalization with attendant increased expenditure of health care resources. A rigid approach for prophylaxis and treatment is illogical, but with separate focus on rate control and cardioversion, a sinus mechanism can be safely and reliably achieved with minimal patient discomfort. PMID- 10535373 TI - Esophageal diverticula: introduction. PMID- 10535372 TI - Treatment of postoperative atrial fibrillation: a nonsurgical perspective. AB - Atrial fibrillation is a common complication of cardiovascular surgery. The 2 most important risk factors for its development are advancing age and a preoperative history of atrial fibrillation. Long-term sequelae, such as a stroke, are uncommon; however, atrial fibrillation frequently results in an increased length and cost of hospitalization. Strategies to prevent postoperative atrial fibrillation include perioperative beta-blockers, amiodarone, and atrial pacing. These strategies are most effective in high-risk patients. When atrial fibrillation does occur, treatment includes control of the ventricular rate, systemic anticoagulation, and conversion back to sinus rhythm. PMID- 10535374 TI - Esophageal diverticula: patient assessment. AB - Esophageal diverticula are best classified by their anatomic location: pharyngoesophageal (Zenker's diverticula), midthoracic, and epiphrenic. Most diverticula result from esophageal motility disorders. Although some patients are asymptomatic and diverticula are incidental findings, most patients are symptomatic. Dysphagia, regurgitation, and pain are common complaints, however, symptoms are often nonspecific and may be the result of an associated esophageal motility disorder. Contrast radiography is the prime diagnostic tool; evaluation of the diverticulum, associated esophageal abnormalities, and complications are assessed by a barium esophogram. Esophagoscopy adds little to the evaluation of the diverticulum but may be indicated in the assessment of other esophageal abnormalities. Motility studies, which may be difficult or hazardous to perform, are of little use in the diagnosis and treatment of Zenker's diverticula. Manometric evaluation of midthoracic or epiphrenic diverticula usually show an associated motility disorder and may influence treatment decisions. PMID- 10535375 TI - The treatment of Zenker's diverticula: a review. AB - A historical review reveals that the treatment of Zenker's diverticula has paralleled its presumed pathophysiology. With the development of technical facilities to better evaluate the pharyngoesophageal region, incomplete relaxation of the upper esophageal sphincter (UES) seems to represent the key element in the development of high pharyngeal pressures with a subsequent outpouching responsible for the diverticulum formation. Many studies have justified myotomy as an essential component in the treatment of pharyngoesophageal diverticula because it represents an efficient therapy with little morbidity. A diverticulopexy should be added for pouches between 1 and 4 cm and a diverticulectomy should be performed for sacs greater than 5 cm to expect the best relief of symptoms. Other treatment modalities have recently been used such as the endoscopic division of the common wall between the cervical esophagus and the diverticulum with either electrocautery (Dohlman's procedure), a laser, or a stapling device. This method is gaining popularity because it achieves a good clinical outcome, especially in high-risk patients. However, more studies are needed to confirm its long-term effectiveness. PMID- 10535376 TI - Midthoracic esophageal diverticula. AB - Periesophageal inflammation, most commonly secondary to tuberculosis, was a frequent cause of midthoracic diverticula. Today, the majority of these diverticula are the result of esophageal motility disorders. Although many patients are asymptomatic, it is the underlying motility disturbances that produce most symptoms. A barium esophagogram is the best study to show midthoracic diverticula. Esophageal manometry may be difficult to perform because of the obstruction of passage of the motility catheter by the diverticulum, but it is useful in defining the cause of the diverticulum and directing therapy. Esophagoscopy is helpful in the assessment of complications or associated esophageal abnormalities. It adds little to the evaluation of the diverticulum. In patients requiring surgery, a diverticulectomy with a myotomy performed on the esophageal wall opposite the diverticulum is the preferred treatment. Lesser procedures have been reported to be successful in select patients. PMID- 10535377 TI - Treatment of epiphrenic diverticula. AB - Epiphrenic diverticulum is a rare disorder of the lower esophagus, thought to be related to an esophageal motility disorder. Treatment should involve removal of diverticulum and myotomy. Although the surgery is technically a difficult one, the long-term outcome should be excellent. PMID- 10535378 TI - Minimally invasive treatment of esophageal diverticula. AB - Minimally invasive approaches are ideally suited to treat diverticula of the mid- and lower esophagus. The most commonly reported procedure is a laparoscopic diverticulectomy and myotomy, particularly when the diverticulum is located within 10 cm of the lower esophageal sphincter. Treatment is the same as for the open approach: Symptomatic patients are offered surgical treatment, the diverticulum is excised without compromise of the esophageal lumen, the proximal extent of the myotomy is dictated by preoperative manometry, and postoperative evaluation is performed to exclude recurrence and gastroesophageal reflux. The results of laparoscopic treatment of esophageal diverticula are similar to the results reported in the open procedure. The laparoscopic technique used to treat esophageal diverticula is described. PMID- 10535379 TI - Beneficial effects of omega-3 fatty acid treatment on the recovery of cardiac function after cold storage of hyperlipidemic rats. AB - Cardiac effects of omega-3 polyunsaturated fatty acids (PUFAs) were studied in female Wister rats fed a standard diet (control [C] diet) or a high-cholesterol (HC) diet. Subgroups of rats from these groups were treated with eicosapentaenoic acid-E (EPA) or docosahexaenoic acid-95E (DHA) for 5 weeks. Although plasma total cholesterol (TC) and triglyceride (TG) levels were higher in each group fed the HC diet versus each group fed the C diet, EPA administration with the HC diet (HC + EPA) significantly (P < .05) reduced these levels. An isolated working-heart preparation was used to determine cardiac function. Cardiac output (CO) was lower in rats fed the HC diet and HC + DHA versus any of the groups fed the C diet (P < .05). In addition, left ventricular (LV) maximum differentiation of pressure-time curve (dp/dt) was lower in the rats fed the HC diet versus any of the C diet groups (P < .05). After evaluation of cardiac function in each rat, the heart was stored in a histidine-tryptophan-ketoglutarate solution for 8 hours at 4 degrees C. The heart was then reperfused, and recovery of cardiac function was evaluated. No significant differences were observed for post-preservative cardiac function within the C diet groups. However, within the HC diet groups, HC + EPA significantly (P < .05) improved the recovery of cardiac function. In addition, HC + DHA also significantly (P < .05) improved the recovery of coronary flow (CF) and LV dp/dt. No significant differences were observed for plasma TC and TG concentrations in the C diet groups. EPA administration significantly decreased cardiac levels of palmitic, oleic, and linoleic acids in the HC diet groups. No significant differences were observed for cardiac levels of free fatty acids (FFAs) within the C diet groups. Cardiac EPA and DHA levels were significantly (P < .05) elevated in EPA- or DHA-treated rats compared with the other diet-fed rats. Cardiac EPA levels were also elevated in DHA-treated rats compared with untreated rats (P < .05). These results suggest that EPA attenuates coronary and myocardial preservation injuries through an increase in serum lipids and an accumulation of myocardial FFAs resulting from a HC diet. PMID- 10535380 TI - Dietary fat saturation, but not the feeding state, modulates rates of cholesterol esterification in normolipidemic men. AB - To determine whether the rates of cholesterol esterification in normal individuals are affected by diets differing in fats, nine men were randomly assigned to three groups receiving a diet rich in monounsaturated (MONO), polyunsaturated (POLY), or saturated (SAT) fat for 2 weeks using a crossover design. Subjects drank a dose of deuterium oxide, and the fractional esterification rate (FER) was calculated during fed and unfed periods. Total esterified cholesterol was calculated as the product of the FER and pool size, the latter obtained from a decay curve following injection of [4-14C] cholesterol. The POLY diet produced the lowest serum cholesterol concentration and the SAT diet the highest (P < .001). For cholesterol ester (CE) deuterium enrichment, an interaction was noted between diet and time (P < .01). The FER was greater (P < .003) in subjects fed the POLY diet versus either of the other diets, although the amount of esterified cholesterol produced, expressed as either milligrams per day (P < .103) or milligrams per kilogram of body weight per day (P < .100), did not differ among groups. No effect of the feeding state was found for either the FER (P < .187) or total esterified cholesterol expressed as milligrams per day (P < .146) or milligrams per kilogram of body weight per day (P < .128). The results suggest that the diet fat type, but not the feeding state, may be responsible for serum esterified cholesterol concentrations. PMID- 10535381 TI - Low serum insulin in traditional Pacific Islanders--the Kitava Study. AB - Increased serum insulin is related to abdominal obesity and high blood pressure in affluent societies where insulin, weight, and blood pressure typically increase with age. The increased insulin level has been thought to reflect insulin resistance, a well-known associated factor in the metabolic syndrome. In most nonwesternized populations, body weight and blood pressure do not increase with age and abdominal obesity is absent. However, it is not known whether serum insulin likewise does not increase with age in nonwesternized societies. Fasting levels of serum insulin were measured cross-sectionally in 164 subsistence horticulturalists aged 20 to 86 years in the tropical island of Kitava, Trobriand Islands, Papua New Guinea, and in 472 randomly selected Swedish controls aged 25 to 74 years from the Northern Sweden WHO Monitoring Trends and Determinants in Cardiovascular Diseases (MONICA) Study. In Kitava, the intake of Western food is negligible and stroke and ischemic heart disease are absent or rare. The body mass index (BMI) and diastolic blood pressure are low in Kitavans. The main outcome measures in this study were the means, distributions, and age relations of serum insulin in males and females of the two populations. Serum fasting insulin levels were lower in Kitava than in Sweden for all ages (P < .001). For example, the mean insulin concentration in 50- to 74-year-old Kitavans was only 50% of that in Swedish subjects. Furthermore, serum insulin decreased with age in Kitava, while it increased in Sweden in subjects over 50 years of age. Moreover, the age, BMI, and, in females, waist circumference predicted Kitavan insulin levels at age 50 to 74 years remarkably well when applied to multiple linear regression equations defined to predict the levels in Sweden. The low serum insulin that decreases with age in Kitavans adds to the evidence that a Western lifestyle is a primary cause of insulin resistance. Low serum insulin may partly explain the low prevalence of cardiovascular disease in Kitavans and probably relates to their marked leanness. PMID- 10535383 TI - Stimulation of oxygen consumption following addition of lipid substrates in liver and skeletal muscle from rats fed a high-fat diet. AB - We studied hepatic and skeletal muscle metabolic activity in rats fed a high-fat diet. Rats were fed a low-fat or high-fat diet for 15 days. At the end of the experimental period, full energy-balance determinations together with serum free triiodothyronine (FT3), leptin, and free fatty acid (FFA) measurements were performed. In addition, we assessed fatty acid-stimulated oxygen consumption in perfused liver and in skeletal muscle homogenate. Rats fed a high-fat diet showed a significant increase in energy intake but no variation in body energy gain, due to a significant increase in energy expenditure. Serum FT3 and FFA levels significantly increased in rats fed a high-fat diet versus rats fed a low-fat diet, while no variation was found in serum leptin levels. Perfused livers and skeletal muscle homogenates from rats fed a high-fat diet exhibited a significant increase in fatty acid-stimulated oxygen consumption. Our results suggest that the enhanced fatty acid oxidation rates in liver and skeletal muscle contribute to the maintenance of fat balance in response to increased fat intake, preventing excess fat deposition. PMID- 10535382 TI - Heparan sulfate chains with antimitogenic properties arise from mesangial cell surface proteoglycans. AB - Heparan sulfate (HS) chains accumulate in both the medium and the cell layer of mesangial cell cultures. When given in fresh medium to quiescent cultures at naturally occurring concentrations, they suppress entry into the cell cycle and progression to DNA synthesis. We have attempted to identify the proteoglycan (PG) source of the antimitogenic HS chains from mesangial cell layers (HS(c)) and medium (HS(c)). When cells were labeled for 16 hours with [35S]sulfate, 25% of the label was found in intracellular HS chains and 5% in extracellular HSPGs. Cell-surface HSPGs accounted for the remaining 70% of the label associated with cell-layer HS and were released by either trypsin or 2% Triton X-100. About 20% of this cell-surface fraction was released by treatment with phosphatidylinositol specific phospholipase C (PI-PLC), and probably represents glypican-like PG; glypican mRNA was present in the cells. The remainder of this fraction could be incorporated into liposomes, indicating the presence of hydrophobic transmembrane regions suggestive of syndecans. Upon purification and deglycosylation, an antiserum to rat liver HSPGs that reacts primarily with syndecan-2 showed a strong signal corresponding to this protein and three weaker bands that may represent additional syndecans. mRNAs for syndecan-1, -2, and -4 were present in the cultures. Syndecan-1 and -2 mRNAs were increased 30 minutes after stimulation of quiescent rat mesangial cells (RMCs) with serum. Heparin, HS(c), and HS(m) all prevented this increase. Syndecan-4 mRNA was not affected by serum, heparin, or HS. In pulse-chase experiments, the amount of 35S appearing in the cellular protein-free HS fraction was accounted for almost entirely by cell-surface PGs, as matrix-associated label was a minor contribution at the end of the pulse labeling. The appearance of [35S]HS in cell extracts was unaffected by phospholipase C treatment, indicating that turnover of the newly labeled syndecan fraction is the source of the antimitogenic HS chains. PMID- 10535384 TI - Octreotide (somatostatin analog) treatment reduces endothelial cell dysfunction in patients with diabetes mellitus. AB - Octreotide is a long-acting somatostatin analog that has been shown to have various effects in diabetes. This study was performed to evaluate whether octreotide affects the vascular complications of diabetes mellitus. Albuminuria and serum thrombomodulin were used as markers of vascular and renal dysfunction. We studied the effect of octreotide in 27 patients with insulin-dependent diabetes mellitus (IDDM). They received 200 microg octreotide per day over a period of 6 months. As a marker of endothelial cell damage, we measured the serum thrombomodulin level. We also measured urinary albumin excretion, hemoglobin A1c (HbA1c), insulin-like growth factor-1 (IGF-1), and other parameters. IGF-1 decreased from 123 ng/mL before treatment to 114 ng/mL after 6 months of octreotide treatment (P = .009), while no significant change was observed in the unblinded control group (from 103 ng/mL to 102 ng/mL after 6 months of treatment). Urinary albumin excretion in patients with macroalbuminuria declined from 1,124 mg/L before octreotide treatment to 556 mg/L after 6 months of treatment (P < .05), whereas no change was observed in the control group. There was also a reduction of the plasma thrombomodulin level from 61.8 ng/mL to 46.1 ng/mL (P < .07) after 6 months of treatment. Furthermore, HbA1c decreased from 8.75% +/- 1.27% to 8.12% +/- 1.23% (P < .07) after octreotide treatment. PMID- 10535385 TI - Disruption of filamentous actin diminishes hormonally evoked Ca2+ responses in rat liver. AB - Previous studies have suggested a role for the actin cytoskeleton in hormonally evoked Ca2+ signaling in the liver. Here, we present evidence supporting a connection between filamentous actin (F-actin) organization and the ability of vasopressin and glucagon to increase cytosolic free-Ca2+ ([Ca2+]i) levels. F actin was disrupted in hepatic cells by perfusion of rat liver with cytochalasin D. Epifluorescence microscopy of subsequently isolated cells showed reduced cortical fluorescent phalloidin staining in cytochalasin D-treated liver cells. Cytochalasin D pretreatment of liver cells reduced the vasopressin-stimulated elevation of [Ca2+]i by 60% and of glucagon by 50%. Experiments performed on cytochalasin D-treated cells using Mn2+ as an indicator of Ca2+ influx quenched fura-2 fluorescence signals following vasopressin administration. This indicates that a structurally intact cortical F-actin web is not a prerequisite for the influx of calcium. Therefore, the attenuation of the increase in cytosolic calcium observed in cytochalasin D-treated liver cells was likely caused either by the depletion of the calcium store by treatment with cytochalasin D or by the need for an intact cytoskeletal structure for its release. Because the resting level of calcium did not change in cells exposed to cytochalasin D, the latter is likely. The reduced [Ca2+]i response may be the mechanism by which cytochalasin D pretreatment inhibits vasopressin-induced metabolic effects. Cytochalasin D pretreatment also decreased the ability of glucagon to stimulate gluconeogenesis and reduced the stimulation of O2 uptake usually observed following glucagon administration. In conclusion, these results suggest that the hormonal elevation of [Ca2+]i and resultant activation of specific metabolic pathways require normal F-actin organization. PMID- 10535386 TI - Imidapril, an angiotensin-converting enzyme inhibitor, improves insulin sensitivity by enhancing signal transduction via insulin receptor substrate proteins and improving vascular resistance in the Zucker fatty rat. AB - Angiotensin-converting enzyme (ACE) inhibitors are antihypertensive agents, that inhibit the conversion of angiotensin I to angiotensin II, resulting in smooth muscle relaxation and a reduction of vascular resistance. Recently, it has been suggested that ACE inhibitors improve insulin resistance in diabetic patients. To investigate the effect of an ACE inhibitor on insulin sensitivity, insulin signaling, and circulation, imidapril was administered orally or intraduodenally to Zucker fatty rats. Oral administration of imidapril improved insulin sensitivity based on the results of an oral glucose tolerance test (OGTT) and a decrease in urinary glucose secretion. Phosphatidylinositol 3-kinase (PI 3 kinase) activity associated with hepatic insulin receptor substrate-1 (IRS-1) in the insulin-stimulated condition was significantly enhanced 110% without a significant alteration in tyrosine phosphorylation of IRS-1 in the imidapril treated group. In muscle, IRS-1 tyrosine phosphorylation and PI 3-kinase activity associated with IRS-1 in the insulin-stimulated condition were enhanced 70% and 20%, respectively, in the imidapril-treated group. In contrast, an alteration of the IRS-2 pathway was observed only in liver; a significant insulin-induced increase in the IRS-2-associated PI 3-kinase over the basal level was observed in the imidapril-treated group but not in the control. In addition, treatment with imidapril was shown to significantly reduce blood pressure and increase blood flow in the liver and muscle. These results suggest that the ACE inhibitor imidapril may improve insulin sensitivity not only by acting directly on the insulin signaling pathway but also by increasing blood flow in tissues via normalization of vascular resistance, a major cause of hypertension. PMID- 10535387 TI - Plasma levels of nitric oxide and related vasoactive factors following long-term treatment with angiotensin-converting enzyme inhibitor in patients with essential hypertension. AB - Several mechanisms other than the inhibition of systemic and local formation of angiotensin II (Ang II) have been proposed to play a role in mediating the hypotensive effects of angiotensin-converting enzyme (ACE) inhibitors. In the present study, we measured plasma levels of nitric oxide (NO) and the related vasoactive factors bradykinin, 6-keto prostaglandin F1alpha (6-keto PGF1alpha) a stable metabolite of prostacyclin, and cyclic guanosine-3',5'-monophosphate (cGMP) before and after a 4-week treatment with the ACE inhibitor lisinopril in 17 patients with essential hypertension. Plasma NO levels were measured by the Griess method after conversion of nitrate to nitrite. Long-term lisinopril treatment significantly reduced blood pressure and increased plasma NO and 6-keto PGF1alpha. The treatment also tended to increase plasma levels of bradykinin and cGMP, but not to a significant extent. The posttreatment NO level was inversely correlated with posttreatment systolic, diastolic, and mean blood pressure (n = 17, r= -.68, P< .01, n = 17, r= -.54, P < .05, and n = 17, r= -.66, P< .01, respectively). The posttreatment bradykinin level was also modestly correlated with posttreatment systolic and mean blood pressure (n = 17, r = -.51, P < .05 and n = 17, r = -.55, P < .05, respectively). In contrast, posttreatment 6-keto PGF1alpha and cGMP levels were not correlated with posttreatment systolic, diastolic, or mean blood pressure. These findings raise the possibility that increased formation of NO and bradykinin, as well as inhibition of the renin angiotensin system, contribute to the hypotensive effect of the ACE inhibitor observed in our hypertensive patients. PMID- 10535388 TI - Hepatic nonoxidative disposal of an oral glucose meal in patients with liver cirrhosis. AB - Seven patients with liver cirrhosis and five healthy subjects were studied over 4 hours after ingestion of a glucose meal to determine whether alterations of hepatic nonoxidative glucose disposal participate in the pathogenesis of impaired glucose tolerance. Hepatic uridyl-diphosphoglucose (UDPG) turnover was calculated from the isotopic enrichment of urinary acetaminophen glucuronide during continuous infusion of 13C-galactose and used as an index of hepatic glycogen synthesis. Patients with cirrhosis had postprandial hyperglycemia and decreased glucose clearance, but hepatic UDPG turnover was not altered (1.84 +/- 0.29 mg/kg fat-free mass min v 1.76 +/- 0.15 in controls, nonsignificant). It is concluded that hepatic postprandial glycogen synthesis is unaltered in patients with advanced cirrhosis, demonstrating important hepatic functional reserve. PMID- 10535389 TI - Decreased skeletal muscle capillary density is related to higher serum levels of low-density lipoprotein cholesterol and apolipoprotein B in men. AB - The relationships between skeletal muscle morphology, particularly muscle fiber capillary density, and serum lipid profiles were evaluated in 25 non-obese men aged 18 to 36 years (body mass index [BMI], 22.7 +/- 2.5 kg/m2; body fat, 13.6% +/- 4.0%, maximal oxygen uptake [VO2max], 46.2 < or = 6.3 mL/kg/min). Skeletal muscle samples were taken from the vastus lateralis using the needle-biopsy method. The fiber types (I, IIa, and IIx) and their percent distribution, the indices of capillary density, and the diffusion index expressed as the cross sectional area occupied by one capillary were determined. Blood samples were drawn from the antecubital vein after a 12-hour fast. Based on Pearson's correlation analysis, the number of capillaries around type IIx fiber correlated inversely with the serum level of low-density lipoprotein cholesterol ([LDL-C] r = -.50, P < .05). The number of capillaries per fiber (cap/fiber ratio), number of capillaries per area (cap/mm2), and capillaries around each fiber type correlated inversely with the serum level of apolipoprotein B ([apo B] r = -.40 to -.54, P < .05 to .01). Further, the diffusion index for each fiber type correlated positively with LDL-C and apo B (r = .42 to .50, P < .05 to .01). Among 14 subjects in whom high-density lipoprotein cholesterol (HDL-C) subfractions were analyzed, a positive correlation was found between cap/mm2 and HDL2-C (r = .64, P < .05). Partial correlation analysis showed that these correlations either remain or improve after adjusting for age, VO2max, and body fatness. These results indicate that skeletal muscle capillary density and diffusion capacity are related to lipid and apolipoprotein concentrations for both type I and type II fibers. PMID- 10535390 TI - Adjusted and unadjusted energy usage rates both determine body fat and plasma leptin in male Fischer 344 rats. AB - Previous studies of the relationship between plasma leptin and energy usage have yielded contradictory findings. The present study was therefore conducted to clearly distinguish and measure the energy usage rate and the energy usage rate adjusted for a surrogate of metabolically active tissue mass. We investigated the simultaneous relationships between these two measures of energy usage, leptin, and body fat in 21-month-old adult male Fischer 344 rats on three different long term dietary regimens: (1) continuous ad libitum feeding (Ad-lib); (2) ad libitum feeding until early adulthood, and then continuous 60% caloric restriction (CR); and (3) ad libitum feeding until early adulthood, then 60% caloric restriction until 16 months, and then ad libitum feeding for 5 months (CR/Ad-lib). Two versions of the daily usage rate were measured: daily dietary caloric intake (DCI), and daily energy expenditure (EE) based on indirect calorimetry. Two versions of the metabolically active tissue mass were also measured: fat-free mass (FFM), and the sum of the weight of the heart, brain, liver, and kidneys. Energy usage rates were adjusted for these measures of metabolically active tissue mass to yield measures of the energy metabolic rate. Correlation, regression, and path analyses showed that both the energy usage rate and adjusted energy usage rate played important independent roles in determining body fat and plasma leptin, but only after multivariate techniques were used to account for the simultaneous interactions between variables. Increases in the energy usage rate were associated with increases in body fat and the adjusted energy usage rate. Increases in the adjusted energy usage rate were associated with decreases in body fat and plasma leptin. These findings suggest that differences in subjects adjusted energy usage rate could explain some of the apparently contradictory findings concerning the relationship between energy usage and plasma leptin in previously published studies. In conclusion, this appears to be the first study to clearly separate and quantify the effects of the energy usage rate and adjusted energy usage rate on body fat and plasma leptin. The findings suggest that under conditions of long-term stable body weight, both of these measures of energy usage play independent simultaneous roles in determining body fat and plasma leptin. PMID- 10535391 TI - Total body fat and abdominal visceral fat response to exercise training in the HERITAGE Family Study: evidence for major locus but no multifactorial effects. AB - The familial etiology of the response in total fat mass (FM) and abdominal visceral fat (AVF) to 20 weeks of exercise training was investigated in families participating in the HERITAGE Family Study. AVF (measured by computed tomographic scanning) and FM (measured by underwater weighing techniques) were assessed at baseline (in a sedentary state) and after 20 weeks of exercise training. The response AVF (AVFdelta) and response FM (FMdelta) were computed as the simple delta values (posttraining - baseline) and adjusted for the effects of sex, generation, and a polynomial in age using multiple regression analysis. To index the AVF response independently of the response in FM and the initial level of visceral fat, the AVFdelta was also adjusted for age and baseline AVF (AVFB) and FMdelta. Familial correlation analysis was used to investigate the multifactorial familial effects (polygenic and/or familial environmental), and segregation analysis was used to search for major gene effects. For the age-adjusted AVFdelta, a putative recessive locus accounting for 18% of the variance (q2 = 1%) was detected. Adjusting AVFdelta for AVFB and FMdelta slightly increased the percentage of variance accounted for (to 26%, q2 = 3%) but did not radically alter the pattern of the parameter estimates. For FMdelta, a putative dominant locus accounting for 31% of the variance (q2 = 49%) was noted. In conclusion, the results were consistent across methods in suggesting that there is little evidence of a multifactorial heritability for either AVFdelta or FMdelta. Rather, the familial etiology of the response to exercise training appears to be primarily due to putative major genes (a recessive locus for AVFdelta and a dominant locus for FMdelta). In addition, a pleiotropic/oligogenic system underlying these variables was inferred. That is, the putative loci for FMdelta and/or AVFB also may impact the AVFdelta, with an additional independent major locus effect on AVFdelta after the former influences have been removed. PMID- 10535392 TI - Long-term infusion of norepinephrine plus serotonin into the ventromedial hypothalamus impairs pancreatic islet function. AB - To examine the possibility of a cause-effect relationship between enhanced monoamine content in the ventromedial hypothalamus ([VMH] a characteristic of hyperinsulinemic and insulin-resistant animals) and islet dysfunction, we infused norepinephrine ([NE] 25 nmol/h) and/or serotonin ([5-HT] 2.5 nmol/h) into the VMH of normal hamsters for 5 weeks and then examined insulin release from the isolated pancreatic islets. VMH infusion of NE + 5-HT, but not of either neurotransmitter alone, produced a marked leftward shift in the dose-response curve of glucose-induced insulin release (twofold to sixfold increase at 5 to 7.5 mmol/L glucose v vehicle-treated animals). In addition, the islet responsiveness to 1 micromol/L NE and 10 micromol/L acetylcholine was abolished in these NE + 5 HT VMH-infused hamsters. These findings indicate that an increase of NE and 5-HT content in the VMH can induce dysregulation of islet insulin release in response to glucose and neurotransmitters. Inasmuch as VMH NE and 5-HT levels are elevated in hyperinsulinemic and insulin-resistant animals, the present findings suggest that an endogenous increase in these hypothalamic monoamines may contribute to islet dysfunction, which is one of the characteristics of type 2 diabetes. PMID- 10535393 TI - Elevated serum leptin concentrations in type 2 diabetic patients with microalbuminuria and macroalbuminuria. AB - Leptin levels are elevated in end-stage renal disease, suggesting an impairment of renal leptin degradation. The present study aimed to determine whether leptin levels are also elevated in patients with earlier stages of renal disease, ie, microalbuminuric and macroalbuminuric nephropathy. A total of 60 subjects were assigned to two study groups. Group A contained 10 type 2 diabetics with macroalbuminuria, 10 type 2 diabetics with normoalbuminuria, and 10 healthy control subjects. Group B contained 10 type 2 diabetics with microalbuminuria, 10 type 2 diabetics with normoalbuminuria, and 10 healthy controls. The subgroups of both study groups were matched for sex and body fatness. In group A, macroalbuminuric diabetic patients had higher serum leptin levels than the normoalbuminuric diabetics (11.90 +/- 2.98 v 4.13 +/- 0.92 ng/mL, P < .002) and control subjects (4.78 +/- 1.37 ng/mL, P < .006). In group B, microalbuminuric diabetics had higher serum leptin levels than the normoalbuminuric diabetics (21.16 +/- 5.80 v8.74 +/- 1.89 ng/mL, P < .04) and control subjects (10.06 + 3.00 ng/mL, P < .06). In both groups A and B, creatinine clearance was inversely correlated with the serum leptin level after adjusting for body fat. In conclusion, serum leptin levels are elevated in type 2 diabetic patients with microalbuminuria and macroalbuminuria, suggesting that renal leptin degradation is already impaired in the early stages of renal disease. PMID- 10535394 TI - Urine and plasma galactitol in patients with galactose-1-phosphate uridyltransferase deficiency galactosemia. AB - Urinary excretion of galactitol was determined in 95 normals (N/N), 67 galactosemic (G/G), and 39 compound heterozygotes for the Duarte and galactosemia genotype (D/G). Galactitol excretion is age-dependent in both normal individuals and patients with classic galactosemia on lactose-restricted diets. In galactosemic patients who are homozygous for the Q188R mutation, urinary galactitol levels were fivefold to 10-fold higher than those of normal subjects of comparable age. All but a few patients with classic galactosemia with the Q188R mutation and another mutant G allele had urinary excretion comparable to the Q188R homozygous patients. African-American galactosemic patients with the S135L mutation of the galactose-1-phosphate uridyltransferase (GALT) gene also excreted abnormal quantities of galactitol. Most subjects with a Duarte allele and a G allele excrete normal amounts of the sugar alcohol. There is a correlation between galactitol excretion and red blood cell (RBC) galactose-1 phosphate (gal-1-P). Plasma galactitol was also elevated in galactosemic patients (3.4 to 23.2 micromol/L; undetectable in normal individuals). In contrast to the decrease in urinary galactitol with age, plasma levels remain in a narrow concentration range with no significant difference with age. Urine and plasma galactitol distinguish galactosemic patients from normals. In addition, urinary galactitol excretion may be an important parameter for the assessment of steady state galactose metabolism in galactosemia. PMID- 10535396 TI - Fatty acid inhibition of glucose-stimulated insulin secretion is enhanced in pancreatic islets from insulin-resistant rats. AB - A study was initiated to test two hypotheses. The first was the postulate that glucose-stimulated insulin secretion would be enhanced in pancreatic islets isolated from normal non-obese rats made insulin-resistant by dietary means. The second, related hypothesis was that glucose-stimulated insulin secretion by pancreatic islets isolated from insulin-resistant rats would be more vulnerable to inhibition following culture in the presence of fatty acids. For this purpose, insulin resistance was induced in normal Sprague-Dawley rats by feeding fat enriched and fructose-enriched diets. The results indicate that islets isolated from either fat-fed or fructose-fed rats secreted significantly more insulin at a glucose concentration of 2.5 to 10.0 mmol/L. In addition, the mean maximal glucose (27 mmol/L)-stimulated insulin secretion rate was significantly lower (15.3 +/- 2.5 ng/islet/h) in islets from fructose-fed rats versus chow-fed rats (25.2 +/- 3.1 ng/islet/h) following culture for 48 hours in the presence of palmitate (0.125 micromol/L). These results support the view that glucose stimulated insulin secretion is enhanced in islets from insulin-resistant rats, and that these islets are more vulnerable to the inhibitory effects of free fatty acid (FFA) on insulin secretion. PMID- 10535395 TI - Sesamin, a sesame lignan, is a potent inducer of hepatic fatty acid oxidation in the rat. AB - The effects of sesamin, one of the most abundant lignans in sesame seed, on hepatic fatty acid oxidation were examined in rats that were fed experimental diets containing various amounts (0%, 0.1%, 0.2%, and 0.5%) of sesamin (a 1:1 mixture of sesamin and episesamin) for 15 days. Dietary sesamin dose-dependently increased both mitochondrial and peroxisomal palmitoyl-coenzyme A (CoA) oxidation rates. Mitochondrial activity almost doubled in rats on the 0.5% sesamin diet. Peroxisomal activity increased more than 10-fold in rats fed a 0.5% sesamin diet in relation to rats on the sesamin-free diet. Dietary sesamin greatly increased the hepatic activity of fatty acid oxidation enzymes, including carnitine palmitoyltransferase, acyl-CoA dehydrogenase, acyl-CoA oxidase, 3-hydroxyacyl-CoA dehydrogenase, enoyl-CoA hydratase, and 3-ketoacyl-CoA thiolase. Dietary sesamin also increased the activity of 2,4-dienoyl-CoA reductase and delta3,delta2-enoyl CoA isomerase, enzymes involved in the auxiliary pathway for beta-oxidation of unsaturated fatty acids dose-dependently. Examination of hepatic mRNA levels using specific cDNA probes showed a sesamin-induced increase in the gene expression of mitochondrial and peroxisomal fatty acid oxidation enzymes. Among these various enzymes, peroxisomal acyl-CoA oxidase and bifunctional enzyme gene expression were affected most by dietary sesamin (15- and 50-fold increase by the 0.5% dietary level). Sesamin-induced alterations in the activity and gene expression of carnitine palmitoyltransferase I and acyl-CoA oxidase were in parallel with changes in the mitochondrial and peroxisomal palmitoyl-CoA oxidation rate, respectively. In contrast, dietary sesamin decreased the hepatic activity and mRNA abundance of fatty acid synthase and pyruvate kinase, the lipogenic enzymes. However, this lignan increased the activity and gene expression of malic enzyme, another lipogenic enzyme. An alteration in hepatic fatty acid metabolism may therefore account for the serum lipid-lowering effect of sesamin in the rat. PMID- 10535397 TI - Metabolic effect of decreasing nonesterified fatty acid levels with acipimox in hyperthyroid patients. AB - Glucose intolerance is often found in patients with hyperthyroidism, but the pathogenetic mechanisms are not fully understood. Since lipolysis is increased in hyperthyroidism, elevated plasma nonesterified fatty acids (NEFAs) may contribute to abnormal glucose metabolism in hyperthyroidism. The aim of this study was to investigate whether decreasing the plasma NEFA level with acipimox can affect glucose metabolism in hyperthyroidism. We performed an intravenous glucose tolerance test (IVGTT) with acipimox 250 mg or placebo in six untreated hyperthyroid men and six age- and body mass index (BMI)-matched controls. Fasting plasma NEFA levels were significantly higher in the hyperthyroid patients versus the controls (997.0 +/- 303.4 v290.5 +/- 169.1 micromol/L, P < .001). Plasma NEFAs decreased rapidly with acipimox treatment in both controls and hyperthyroid patients. In the controls, the glucose disappearance constant (K(G)) was not different for acipimox treatment versus placebo (2.18 +/- 0.62 v 2.42 +/- 1.00% x min(-1)). In hyperthyroid patients, acipimox treatment increased the K(G) significantly compared with placebo treatment (2.44 +/- 0.84 v 1.58 +/- 0.37% x min(-1), P < .05). Changes in K(G) values with acipimox treatment were inversely correlated with changes in plasma NEFA levels (r = -.65, P < .05). Acipimox treatment increased the acute insulin response (AIR) in hyperthyroid patients (943 +/- 381 v 698 +/- 279 microU/mL x min, P < .05), whereas it did not change the AIR in controls. Changes in the AIR with acipimox treatment correlated significantly with changes in the K(G) (r = .70, P < .05). There was a weak correlation between changes in the AIR with acipimox treatment and changes in plasma NEFA levels (r = -.55, P = .06). In summary, decreasing the plasma NEFA level with acipimox in hyperthyroid patients increases both the K(G) and AIR during an IVGTT. These findings suggest that the abnormal glucose metabolism in hyperthyroidism could be attributed, at least in part, to the increase of plasma NEFA. PMID- 10535398 TI - Visceral adipose tissue and low-density lipoprotein particle size in middle-aged versus young men. AB - Age is associated with increased deposition of visceral adipose tissue. We examined whether this age-related change in regional adipose tissue distribution had an impact on low-density lipoprotein (LDL) particle size. For this purpose, the plasma lipoprotein-lipid profile, including LDL peak particle diameter as determined by gradient gel electrophoresis, was assessed in 38 young men (aged 26.4 +/- 4.2 years, mean +/- SD) and compared with 40 middle-aged men (55.9 +/- 6.2 years). Middle-aged men had higher values for total body fat and visceral adipose tissue area as measured by computed tomography than young men (P < .001). Although significant differences were noted between the two age groups for plasma cholesterol, triglyceride (TG), apolipoprotein B (apo B), LDL cholesterol, and LDL apo B, as well as the cholesterol to high-density lipoprotein (HDL) cholesterol ratio (P < .001), no difference was found for LDL peak particle size between young and middle-aged men. While visceral adipose tissue was a significant correlate of plasma lipoprotein levels, the fasting TG concentration was the best predictor of LDL particle size, and the regression of TG levels on LDL peak particle diameter was not different between the two age groups. These results suggest that middle-aged men are characterized by an increased concentration of LDL particles (reflected by increased LDL apo B levels) but not by a reduced LDL peak particle size compared with young men. It is therefore proposed that in the absence of an important age-related change in TG levels, age per se is associated with an increased concentration of atherogenic LDL particles rather than a reduction of LDL particle diameter. PMID- 10535399 TI - Effect of short-term medroxyprogesterone acetate on left ventricular mass: role of insulin-like growth factor-1. AB - Previous studies using 17beta-estradiol and medroxyprogesterone acetate (MPA) have shown that hormone replacement therapy (HRT) increases left ventricular mass (LVM). To determine if insulin-like growth factor-1 (IGF-1) is associated with the increase in LVM, we measured IGF-1 and IGF-binding protein-3 (IGFBP-3) levels in 19 postmenopausal women before and after 8 weeks of oral treatment with MPA 5 mg/d. LVM was measured by two-dimensional echocardiography. Changes in IGF-1, IGFBP-3, and LVM from baseline were analyzed by paired ttest. Regression analysis was used to determine if changes in the IGF-1 axis with MPA treatment affect the increase in LVM. LVM increased 4.4% during the study (P = .006 vbaseline). IGF-1 increased 17% with MPA (P = .008), whereas IGFBP-3 did not change. The IGF 1/IGFBP-3 ratio increased 16.8% (P = .0003). Regression analysis of LVM with IGF 1, IGFBP-3, and the IGF-1/IGFBP-3 ratio suggested that IGF-1 during MPA therapy explains 2.4% and the IGF-1/IGFBP-3 ratio explains 3.2% of the variation in LVM. There was no effect of IGFBP-3 on LVM. Most of the variation in LVM with MPA (90.5%) was explained by baseline LVM. The IGF-1/IGFBP-3 ratio on MPA treatment was inversely related to the change in LVM: women with a lower LVM at baseline had the greatest increase in LVM with MPA. These findings suggest that MPA increases IGF-1 and LVM. Because the increase in IGF-1 with MPA treatment explains a fraction of the increase in LVM, other mechanisms must also be operative. PMID- 10535400 TI - Circulating tumor necrosis factor alpha concentrations are higher in abdominal versus peripheral obesity. AB - Fat tissue is a significant source of endogenous tumor necrosis factor alpha (TNFalpha), the pluripotent cytokine that plays an important role as a mediator of the peripheral insulin resistance found in obesity. The majority of evidence for this role of TNFalpha is from studies in animal models of obesity. To explore further the role of TNFalpha in the pathogenesis of obesity-related insulin resistance in humans, we compared plasma levels of TNFalpha and the other main endocrine cytokine, interleukin-6 ([IL-6] both measured by enzyme-linked immunosorbent assay), in 26 obese women (body mass index [BMI] > 30 kg/m2) and 13 female controls (BMI < 26 kg/m2) without a history of recent or active infection. Glucose and insulin levels were measured at 0, 1, and 2 hours after a 75-g oral glucose load. There was no significant difference in plasma TNFalpha or IL-6 levels between obese and non-obese subjects overall (2.10 +/- 0.19 v 1.65 +/- 0.18 pg/mL and 2.06 +/- 0.29 v 1.50 +/- 0.17 pg/mL, respectively). However, TNFalpha levels were significantly elevated in obese subjects with a 2-hour glucose level more than 140 mg/dL (n = 8) compared with the other obese subjects (n = 18) and the non-obese controls (2.88 +/- 0.46 v 1.75 +/- 0.10 and 1.65 +/- 0.18 pg/mL, respectively, P < .01). Furthermore, the TNFalpha level correlated significantly with the waist to hip ratio ([WHR] r = .53, P < .01) and fasting and post-oral glucose tolerance test (OGTT) insulin levels (r = .47, P < .02), but not with the BMI, and was higher in obese women with a WHR more than 0.90 (n = 14) in comparison to those with a WHR less than 0.90 (n = 12, 2.47 +/- 0.29 v 1.66 +/- 0.18 pg/mL, respectively, P < .03). The corresponding plasma leptin level was significantly higher in obese women versus the control group (41.6 +/- 2.5 v22.3 +/- 2.9 ng/mL, P < .001) and was related to the BMI (r = .60, P < .01) but not to TNFalpha or the WHR. There were no significant differences in the corresponding IL-6 concentration between groups, and IL-6 did not correlate with TNFalpha, leptin, BMI, WHR, or insulin levels. In conclusion, circulating TNFalpha levels are higher in abdominal obesity compared with peripheral obesity, and may contribute to the insulin resistance that more commonly complicates the former pattern of fat distribution. PMID- 10535401 TI - Further observations on the relationship between adenosine deaminase and body mass. PMID- 10535402 TI - Biological relevance of pituitary adenylate cyclase-activating polypeptide (PACAP) in the gastrointestinal tract. AB - Since its initial discovery in 1989, pituitary adenylate cyclase activating peptide (PACAP) has been noted to distribute widely in the brain, the respiratory and the gastrointestinal system. It occurs in two bioactive molecules, PACAP-27 and the C-terminally extended PACAP-38, which evoke activity by binding to three distinct types of high-affinity, G-protein coupled membrane receptors. It is present throughout the entirety of the gut but is rare in certain areas such as the intestinal mucosa and islets of Langerhans. PACAP-induced biological effects are protean and include alterations of motility in the bowel and the gallbladder, stimulation of gastric acid and intestinal secretion, hormone/enzyme release from the exocrine and endocrine pancreas, and the induction as well as inhibition of proliferation in neuroendocrine cells and tumors. Its hepatic activity has to date not been elucidated in detail. One of the interesting features of PACAP is the species and organ dependent variation of its biological effects. Of particular note is its superior potency when compared with other neuropeptides identified in the gut, and the involvement of a number of different second messenger systems upon PACAP receptor activation. PMID- 10535403 TI - Characterization of rodent liver and kidney AVP receptors: pharmacologic evidence for species differences. AB - Radioligand binding studies with [3H]vasopressin (AVP) were used to determine the affinities of AVP receptor agonists and antagonists for mouse liver and kidney plasma membrane preparations. Both membrane preparations exhibited one class of high-affinity binding site. AVP ligand binding inhibition studies confirmed that mouse liver binding sites belong to the V1A subtype while kidney binding sites belong to the V2 receptor subtype. The affinity of each ligand for mouse V1A receptors was very similar to that for rat V1A receptors, showing differences in Ki values of less than 3-fold. In contrast, several peptide (d(CH2)5Tyr(Me)AVP) and nonpeptide (OPC-21268 and SR 49059) ligands had different affinities for mouse and rat kidney V2 receptors, with differences in Ki values ranging from 14- to 17-fold. These results indicate that mouse and rat kidney V2 receptors show significant pharmacologic differences. PMID- 10535404 TI - A synthetic peptide corresponding to amino acid residues 90 to 95 of human follicle-stimulating hormone beta-subunit delays the onset of puberty in female Swiss Webster mice. AB - We have recently reported that a synthetic peptide corresponding to amino acid residues 81-95, a receptor-binding region of the human FSH-beta-subunit, and a subdomain within this region, hFSH-beta-(90-95) (DSTDCT), prolonged vaginal estrus when administered intraperitoneally (ip) to normally cycling Swiss Webster mice. These results were similar to those we reported for a synthetic peptide corresponding to hFSH-beta-(34-37) [TRDL, a subdomain within receptor-binding region hFSH-beta-(33-53)] in the same model system. TRDL also accelerated the onset of puberty in immature mice. We now report the effects of hFSH-beta-(90-95) in prepubertal female mice. In two separate experiments, a single ip injection of 200 microg/g body weight (BW) hFSH-beta-(90-95) in phosphate buffered saline (PBS, vehicle) administered to mice on day 28 delayed first vaginal estrus by 3 days in 50% (4/8) and 62.5% (6/8) when compared to mice given vehicle alone on day 28. Vaginal opening was also delayed in mice receiving hFSH-beta-(90-95) when compared to mice injected with vehicle alone. Serum estradiol levels of vehicle injected control mice in first vaginal estrus were three-fold higher than in mice treated with hFSH-beta-(90-95), whose vaginal smears showed no evidence of first estrus. No significant differences in ovarian or uterine weights, or serum progesterone levels, were observed between vehicle-injected control mice achieving first vaginal estrus and mice receiving hFSH-beta-(90-95) in whom first estrus was delayed. The contrasting effects on the onset of puberty of hFSH-beta (90-95) (delay) and hFSH-beta-(34-37) (acceleration) may reflect synthetic peptide binding to different domains of the FSH receptor, resulting in variable post-binding effects. These results are consistent with our earlier study suggesting that FSH-beta-subunit-related synthetic peptides can induce significant in vivo effects on the onset of puberty in female mice. PMID- 10535405 TI - Gastrin-induced gene expression in oxyntic mucosa and ECL cells of rat stomach. AB - The histamine-producing ECL cells are numerous in the acid-producing (oxyntic) mucosa. They respond to gastrin by secretion of histamine that acts on parietal cells to produce acid. In addition, gastrin has a trophic effect on the oxyntic mucosa which is exerted on stem cells and ECL cells. To elucidate the molecular actions of gastrin on the stomach we attempted to identify genes that are regulated by gastrin in oxyntic mucosa and in isolated ECL cells. Differential display polymerase chain reaction was used to identify mRNAs that are differentially expressed in rats that are hypergastrinemic after treatment with the proton pump inhibitor omeprazole for 48 h compared with rats that are hypogastrinemic after 24 h fasting. Differences in mRNA levels were confirmed by Northern blot analysis (comparing mRNA from fasted rats, omeprazole-treated rats and rats treated with omeprazole + the CCK2 (cholecystokinin) receptor antagonist YF476). The cDNAs were identified by sequencing followed by data base search. Hypergastrinemia induced by omeprazole treatment resulted in overexpression of mRNA for histidine decarboxylase, fetuin, pepsinogen and cytochrome P450 in the oxyntic mucosa. This was prevented by CCK2 receptor blockade. In isolated ECL cells gastrin upregulated mRNAs for histidine decarboxylase and synaptotagmin V as well as one mRNA transcript without known homology. PMID- 10535406 TI - Correlation of type I insulin-like growth factor receptor (IGF-I-R) and insulin receptor-related receptor (IRR) messenger RNA levels in tumor cell lines from pediatric tumors of neuronal origin. AB - The insulin receptor-related receptor (IRR) is a member of the insulin receptor family. So far no ligand has yet been discovered for this receptor type (orphan receptor). IRR, insulin receptor (IR), and insulin-like growth factor-I receptor (IGF-I-R) are all tyrosine kinases. The cellular function of the IRR is not known. The expression of IRR mRNA is restricted to a few, e.g. neuronal tissues, and has also been found in neuroblastomas. Since tyrosine kinase receptors, including the IGF-I-R, may be involved in tumor genesis, we examined the expression of IRR mRNA and IGF-I-mRNA in 18 tumor cell lines using RT-PCR and the solution hybridization/RNAse protection assay. In particular, the mRNA levels of IRR and IGF-I-R were compared by semi-quantitative RT-PCR in seven neuroblastomas and 11 soft tissue sarcomas (STS), five of which were of neuronal origin. In all of the seven neuroblastoma cell lines and in five of the 11 STS cell lines, the IRR mRNA was detected. In addition, the IRR mRNA was expressed in rhabdomyosarcoma, in leiomyosarcoma, in one of the Ewing sarcoma and in the neurofibrosarcoma cell line. The last two tumor cell types are of neuronal origin. The levels of expression of IGF-I-R and IRR mRNA of the neuroblastoma cell lines were closely related (r = 0.82, P < 0.002). Furthermore, IRR mRNA was found only in cell lines that also expressed IGF-I-R mRNA. In conclusion, cell lines from pediatric tumors of neuronal origin express IRR mRNA simultaneously with a another tyrosine kinase receptor (IGF-I-R) mRNA. The tight coupling of their mRNA expression suggests a functional association of both receptors in the tumor cells. PMID- 10535407 TI - C-type natriuretic peptide (CNP) effects on intracellular calcium [Ca2+]i in mouse gonadotrope-derived alphaT3-1 cell line. AB - C-type natriuretic peptide (CNP), the third member of the atrial natriuretic peptide family, acts via guanylyl cyclase containing GC-B receptors to stimulate cyclic guanosine 3',5' monophosphate (cGMP) accumulation in the gonadotrope derived alphaT3-1 cell line and rat pituitary cells. This effect is inhibited by concomitant activation of the phospholipase C (PLC)-coupled gonadotrophin hormone releasing hormone (GnRH) receptors in these cells. Since GnRH stimulates gonadotrophin secretion from gonadotropes by increasing the cytosolic Ca2+ concentration ([Ca2+]i) and natriuretic peptides have been found to influence PLC/Ca2+ signalling in other systems, we have investigated whether CNP can alter basal or GnRH-stimulated changes in [Ca2+]i in alphaT3-1 cells. In Ca 2+ containing medium, 10(-7) M CNP modestly, but significantly increased [Ca2+]i over several min, but subsequently inhibited the elevation of [Ca2+]i in response to 10(-7) M GnRH in both Ca2+-containing and Ca2+-free medium. This inhibitory effect was mimicked by 10(-6) M 8-Br-cGMP, but not by ANP, indicating mediation by cyclic GMP and the CNP-specific GC-B receptor. However, basal and GnRH stimulated inositol (1,4,5) trisphosphate (Ins(1,4,5)P3) generation were not measurably affected by CNP, and CNP failed to affect thapsigargin-induced capacitative Ca2+ entry. Thus, it appears that the cross-talk between CNP and GnRH in these cells is reciprocal in that GnRH modulates CNP effects on cGMP generation, whereas, CNP modulates GnRH effects on Ca2+ mobilisation. PMID- 10535408 TI - Angiotensin-(1-7) binds at the type 1 angiotensin II receptors in rat renal cortex. AB - Significant angiotensin (Ang) (1-7) production occurs in kidney and effects on renal function have been observed. The present study was undertaken to investigate binding characteristics of the heptapeptide to Ang II receptors present in rat renal cortex. [125I]-Ang II binding to rat glomeruli membranes was analyzed in the presence of increasing concentrations of Ang II, Ang-(1-7), DUP 753 and PD 123319. Linearity of the Scatchard plot of the [125I]-Ang II specific binding to rat glomeruli membranes indicated a single population of receptors, with a Kd value of 0.7 +/- 0.1 nM and a Bmax of 198 +/- 0.04 fmol/mg protein. DUP 753, an specific AT1 receptor antagonist, totally displaced the specific binding of [125I]-radiolabelled hormone with a Ki of 15.8 +/- 0.9 nM, while no changes were observed in the presence of the selective AT2 receptor antagonist, PD 123319. The specific [125I]-Ang II binding to rat glomerular membranes was displaced by Ang-(1-7) with high affinity (Ki = 8.0 +/- 3.2 nM). We conclude that radioligand binding assays in the presence of selective Ang II antagonists DUP 753 and PD 123319 suggest the unique presence of AT1, receptors in rat glomeruli and a possible role in the control of the biological renal effects of Ang-(1-7). PMID- 10535409 TI - Vasoactive intestinal peptide inhibits cytokine production in T lymphocytes through cAMP-dependent and cAMP-independent mechanisms. AB - Previous reports indicate that VIP and the structurally related peptide PACAP, inhibit IL-2 and IL-10 production in antigen-stimulated T lymphocytes. Intracellular cAMP elevation appears to be the primary transduction pathway involved. However, in the lower concentration range, an additional, cAMP independent transduction pathway appears to mediate the VIP inhibition of cytokine production. Here, we address this question by using VIP agonists and antagonists which act through cAMP-dependent and -independent pathways. The antagonists based on the neurotensin-VIP hybrid molecule did not affect the inhibitory effect of VIP/PACAP on IL-2 and IL-10 production, confirming that astrocytes and T lymphocytes express different receptors. A lipophilic antagonist with increased membrane permeability, partially reversed the inhibitory effect of VIP/PACAP, forskolin, prostaglandin E2, and 8-bromo-cAMP without significantly affecting cAMP levels, suggesting that it acts downstream of cAMP. Two VIP agonists inhibit IL-2 and IL-10 production. One of the agonists increases cAMP, whereas the second one does not induce cAMP/cGMP. Our results indicate that VIP inhibits cytokine production in stimulated CD4+ T cells through two separate mechanisms, which involve both cAMP-dependent and cAMP-independent transduction pathways. PMID- 10535410 TI - CRH-deficient mice have a normal anorectic response to chronic stress. AB - Many studies have implicated corticotropin-releasing hormone (CRH) as a mediator of stress-induced decreases in food intake. However, urocortin, sauvagine, and urotensin, other members of the family of CRH-like molecules, have also been shown to be potent inhibitors of food intake. This raises the possibility that a CRH-related molecule might also be responsible for stress-induced anorexia. We therefore examined the effects of three chronic stressors, repetitive daily restraint, turpentine abscess, and surgical stress, upon food intake in wildtype and CRH-deficient mice created by targeted inactivation of the CRH gene. We have found that both genotypes have similar basal food intake which initially decreases to the same degree following initiation of each stress paradigm. Food intake also recovers following the same time course and to the same degree in both genotypes. Therefore, CRH is not necessary for decreases in food-intake induced by the chronic stressors examined in this study. PMID- 10535412 TI - Demonstration of new sites of expression of the CCK-B/gastrin receptor in pancreatic acinar AR42J cells using immunoelectron microscopy. AB - The CCK-B/gastrin receptor has been characterised in both normal and tumour tissues. Endocytosis of the CCK-B/gastrin receptor has recently been demonstrated and this has similarly been described for other peptide receptors. In addition, ligand and ligand-receptor translocation to the nucleus has been demonstrated for other peptides. The aim of this study was to identify the sites of expression of the CCK-B/gastrin receptor in the known CCK-B/gastrin receptor bearing pancreatic acinar AR42J cells. The specificity of the CCK-B/gastrin receptor antibody (alpha CCKBR-Ser antibody) was demonstrated by inhibition ELISA studies, radioligand inhibition studies and immunofluorescence binding studies on AR42J cells. Western blotting and immunogold electron microscopy techniques were used to identify the receptor in AR42J cell preparations. The affinity purified alpha-CCKBR-Ser antibody was shown to be specific for the CCK-B/gastrin receptor. The receptor was expressed on the cell membrane, in the cytoplasm and within the nucleus. Isoforms of the receptor protein identified in extra-nuclear and nuclear extracts ranged in molecular weight from 58 to 66 kDa. We conclude that the CCK-B/gastrin receptor is not only expressed on the cell membrane, but also in the cytoplasm and nucleus of AR42J pancreatic acinar cells. PMID- 10535411 TI - Transglutaminase-mediated amine incorporation into substance P protects the peptide against proteolysis in vitro. AB - The in vitro metabolism of transglutaminase-synthesized substance P analogs has been characterized comparing their stability to that of the parent peptide. The major metabolites have been purified and their structures elucidated by mass spectrometry. Our results demonstrated that gln5 spermidine and spermine analogs of substance P possess an enhanced resistance to the action of proteases. Moreover spermine, a large size hydrophilic compound, specifically prevented the hydrolysis at Phe7-Phe8 bond. PMID- 10535414 TI - LHRH antagonist inhibits gastric cell proliferation in suckling rats. AB - The cell proliferation of the gastric epithelium is stimulated by food deprivation in suckling rats, and LHRH inhibits this process as it does in other hyperproliferating tissues. However, as LHRH antagonist is also being used as a potent antiproliferative agent for tumors, this study aims to investigate whether it plays any role on the cell proliferation of the gastric epithelium. Seventeen day-old rats were fasted for 20 h and treated with LHRH antagonist, LHRH, or both during this period. The following morning, animals were injected with BrDU to label proliferating cells, and were sacrificed 1 h later. Paraffin sections of the gastric mucosa were processed for immunohistochemistry and BrDU labeled and non-labeled epithelial cells were counted to determine the labeling index (LI). None of the hormone treatments changed body weight or the morphology of the stomach. Apoptotic cells were observed in the gastric gland in treated rats. The LI were inhibited by the antagonist, LHRH or both, showing that the LHRH antagonist plays an inhibitory role on the hyperproliferating gastric epithelium. These results suggest that both LHRH agonist and antagonist can influence the proliferative activity in suckling rats, though the mode of action remains to be determined. PMID- 10535413 TI - Renal effects of angiotensin II receptor subtype 1 and 2-selective ligands injected into the paraventricular nucleus of conscious rats. AB - We determined the effects of losartan and CGP42112A (selective ligands of the AT1 and AT2 angiotensin receptors, respectively) and salarasin (a relatively nonselective angiotensin receptor antagonist) on urinary volume and urinary sodium and potassium excretion induced by administration of angiotensin II (ANG II) into the paraventricular nucleus (PVN) of conscious rats. Both the AT1 and AT2 ligands and salarasin administered in the presence of ANG II elicited a concentration-dependent inhibition of urine excretion, but losartan inhibited only 75% of this response. The IC50 for salarasin, CGP42112A, and losartan was 0.01, 0.05, and 6 nM, respectively. Previous treatment with saralasin, CGP42112A and losartan competitively antagonized the natriuretic responses to PVN administration of ANG II, and the IC50 values were 0.09, 0.48, and 10 nM, respectively. The maximum response to losartan was 65% of that obtained with saralasin. Pretreatment with saralasin, losartan, and CGP42112A injected into the PVN caused shifts to the right of the concentration-response curves, but the losartan concentrations were disproportionately greater compared with salarasin or CGP42112A. The IC50 values were 0.06, 0.5, and 7.0 for salarasin, CGP42112A, and losartan, respectively. These results suggest that both AT1 and AT2 receptor subtypes in the PVN are involved in ANG II-related urine, sodium, and potassium excretion, and that the inhibitory responses to AT2 blockade are predominant. PMID- 10535415 TI - Inhibition of gastric emptying by bombesin-like peptides is dependent upon cholecystokinin-A receptor activation. AB - The amphibian peptide bombesin (BN) and the related mammalian peptides gastrin releasing peptide (GRP) and neuromedin B (NMB) inhibit gastric emptying in rats. Exogenous administration of BN stimulates the release of cholecystokinin (CCK), a gastrointestinal peptide that also potently inhibits gastric emptying. To determine whether the inhibition of gastric emptying by BN-like peptides is mediated by a CCK-dependent mechanism, we examined the ability of the CCK-A receptor antagonist, devazepide, to block the inhibition of saline gastric emptying produced by BN, GRP18-27 and NMB. Using the same dosages as in the gastric emptying experiment, we also evaluated the effect of devazepide on feeding suppression produced by systemically administered BN. Our results showed that devazepide completely blocked the suppression of gastric emptying produced by BN, GRP18-27 and NMB but had no effect on BN-induced suppression of food intake. These results suggest that BN-like peptides inhibit gastric emptying through an indirect mechanism that is dependent upon CCK-A receptor activation. In contrast, the suppression of food intake by BN, in this experimental paradigm, is independent of CCK-A receptors. PMID- 10535416 TI - Contraceptive effectiveness: what should the counseling message be? PMID- 10535417 TI - What's in a (drug) name? Plenty! PMID- 10535418 TI - R U E 4 Y2K? PMID- 10535419 TI - 1999 Albert Lasker Medical Research Awards presented. PMID- 10535420 TI - New technique treats male infertility. PMID- 10535421 TI - From the Centers for Disease Control and Prevention. Bidi use among urban youth- Massachusetts, March-April 1999. PMID- 10535422 TI - From the Centers for Disease Control and Prevention. High prevalence of chlamydial and gonococcal infection in women entering jails and juvenile detention centers--Chicago, Birmingham, and San Francisco, 1998. PMID- 10535423 TI - Cost-effectiveness of methods to enhance sensitivity of Papanicolaou testing. PMID- 10535424 TI - Cost-effectiveness of methods to enhance sensitivity of Papanicolaou testing. PMID- 10535425 TI - Recognizing abusive head trauma in children. PMID- 10535426 TI - Recognizing abusive head trauma in children. PMID- 10535427 TI - Recognizing abusive head trauma in children. PMID- 10535428 TI - Electron beam computed tomography to detect coronary artery disease. PMID- 10535429 TI - Total parenteral nutrition for critically ill patients. PMID- 10535430 TI - Total parenteral nutrition for critically ill patients. PMID- 10535431 TI - Total parenteral nutrition for critically ill patients. PMID- 10535432 TI - A population-based study of school scoliosis screening. AB - CONTEXT: Although school-based screening programs for adolescent idiopathic scoliosis are mandated in 26 states in the United States, few program outcomes data exist regarding the effectiveness of such programs. OBJECTIVE: To determine the effectiveness of a community-based school scoliosis screening program. DESIGN, SETTING, AND PARTICIPANTS: Retrospective cohort study of children who attended kindergarten or first grade at public or private schools in Rochester, Minn, during 1979-1982. Children were followed up until age 19 years or until they left the school district. MAIN OUTCOME MEASURES: Number of children diagnosed and treated for scoliosis, based on results from scoliosis screenings performed annually in grades 5 through 9, linked to community medical records data; performance characteristics of the screening program. RESULTS: Of the 2242 children screened, 92 (4.1 %) were referred for further evaluation. Of these, 68 (74%) had documented medical or chiropractic evaluation of scoliosis. School screening identified 5 of the 9 children treated for scoliosis but resulted in referrals for another 87 children who were not treated. The cumulative incidence of diagnosed scoliosis in this population was 1.8% (95% confidence interval [CI], 1.2%-2.3%) for curves of more than 10 degrees, 1.0% (95% CI, 0.6%-1.5%) for curves of at least 20 degrees, and 0.4% (95% CI, 0.1 %-0.6%) for curves of 40 degrees or more; 0.4% (0.5% of girls and 0.3% of boys) were treated for scoliosis. The positive predictive value of the school screening program for the identification of treated scoliosis was 0.05 (95% CI, 0.048-0.052), with 448 children needed to screen to identify 1 child who subsequently received treatment. The percent positive agreement across consecutive years of screening varied from 7% to 30%. CONCLUSION: In this population, school scoliosis screening identified some children who went on to receive treatment but referred many more who did not. These data should be considered in making decisions regarding school scoliosis screening. PMID- 10535433 TI - Walking compared with vigorous physical activity and risk of type 2 diabetes in women: a prospective study. AB - CONTEXT: Although many studies suggest that physical activity may reduce risk of type 2 diabetes, the role of moderate-intensity activity such as walking is not well understood. OBJECTIVES: To examine the relationship of total physical activity and incidence of type 2 diabetes in women and to compare the benefits of walking vs vigorous activity as predictors of subsequent risk of type 2 diabetes. DESIGN AND SETTING: The Nurses' Health Study, a prospective cohort study that included detailed data for physical activity from women surveyed in 11 US states in 1986, with updates in 1988 and 1992. PARTICIPANTS: A total of 70,102 female nurses aged 40 to 65 years who did not have diabetes, cardiovascular disease, or cancer at baseline (1986). MAIN OUTCOME MEASURE: Risk of type 2 diabetes by quintile of metabolic equivalent task (MET) score, based on time spent per week on each of 8 common physical activities, including walking. RESULTS: During 8 years of follow-up (534, 928 person-years), we documented 1419 incident cases of type 2 diabetes. After adjusting for age, smoking, alcohol use, history of hypertension, history of high cholesterol level, and other covariates, the relative risks (RRs) of developing type 2 diabetes across quintiles of physical activity (least to most) were 1.0, 0.77, 0.75, 0.62, and 0.54 (P for trend <.001); after adjusting for body mass index (BMI), RRs were 1.0, 0.84, 0.87, 0.77, and 0.74 (P for trend = .002). Among women who did not perform vigorous activity, multivariate RRs of type 2 diabetes across quintiles of MET score for walking were 1.0, 0.91,0.73, 0.69, and 0.58 (P for trend <.001). After adjusting for BMI, the trend remained statistically significant (RRs were 1.0, 0.95, 0.80, 0.81, 0.74; P for trend = .01). Faster usual walking pace was independently associated with decreased risk. Equivalent energy expenditures from walking and vigorous activity resulted in comparable magnitudes of risk reduction. CONCLUSIONS: Our data suggest that greater physical activity level is associated with substantial reduction in risk of type 2 diabetes, including physical activity of moderate intensity and duration. PMID- 10535434 TI - Bronchiolitis-associated hospitalizations among US children, 1980-1996. AB - CONTEXT: Respiratory syncytial virus (RSV) causes more lower respiratory tract infections, often manifested as bronchiolitis, among young children than any other pathogen. Few national estimates exist of the hospitalizations attributable to RSV, and recent advances in prophylaxis warrant an update of these estimates. OBJECTIVES: To describe rates of bronchiolitis-associated hospitalizations and to estimate current hospitalizations associated with RSV infection. DESIGN AND SETTING: Descriptive analysis of US National Hospital Discharge Survey data from 1980 through 1996. PARTICIPANTS: Children younger than 5 years who were hospitalized in short-stay, non-federal hospitals for bronchiolitis. MAIN OUTCOME MEASURE: Bronchiolitis-associated hospitalization rates by age and year. RESULTS: During the 17-year study period, an estimated 1.65 million hospitalizations for bronchiolitis occurred among children younger than 5 years, accounting for 7.0 million inpatient days. Fifty-seven percent of these hospitalizations occurred among children younger than 6 months and 81 % among those younger than 1 year. Among children younger than 1 year, annual bronchiolitis hospitalization rates increased 2.4-fold, from 12.9 per 1000 in 1980 to 31.2 per 1000 in 1996. During 1988-1996, infant hospitalization rates for bronchiolitis increased significantly (P for trend <.001), while hospitalization rates for lower respiratory tract diseases excluding bronchiolitis did not vary significantly (P for trend = .20). The proportion of hospitalizations for lower respiratory tract illnesses among children younger than 1 year associated with bronchiolitis increased from 22.2% in 1980 to 47.4% in 1996; among total hospitalizations, this proportion increased from 5.4% to 16.4%. Averaging bronchiolitis hospitalizations during 1994-1996 and assuming that RSV was the etiologic agent in 50% to 80% of November through April hospitalizations, an estimated 51, 240 to 81, 985 annual bronchiolitis hospitalizations among children younger than 1 year were related to RSV infection. CONCLUSIONS: During 1980-1996, rates of hospitalization of infants with bronchiolitis increased substantially, as did the proportion of total and lower respiratory tract hospitalizations associated with bronchiolitis. Annual bronchiolitis hospitalizations associated with RSV infection among infants may be greater than previous estimates for RSV bronchiolitis and pneumonia hospitalizations combined. PMID- 10535435 TI - Prognostic value of 24-hour blood pressure in pregnancy. AB - CONTEXT: Elevated blood pressure (BP) measured at the physician's office may reflect true hypertension or white coat hypertension (WCH). The prognostic value of WCH among pregnant women is unknown. OBJECTIVE: To assess the prognostic value of WCH in pregnancy. DESIGN: Prospective cohort study conducted between September 1994 and October 1997. SETTING: Community hospital. PATIENTS: Women without preexisting hypertension and not treated with antihypertensive drugs aid with high (n = 148) or normal (n = 106) office BP (high office BP was defined as > or =140 mm Hg systolic and/or > or =90 mm Hg diastolic) matched for gestational age during their third trimester of pregnancy. All women underwent 24-hour noninvasive BP monitoring, and women without hypertension on 24-hour monitoring (125/74 mm Hg or less for average 24-hour BP) with office hypertension were classified as having WCH. Women were followed up through the end of pregnancy. MAIN OUTCOME MEASURES: Duration of pregnancy, gestational hypertension, preeclampsia or eclampsia, cesarean delivery, placental and neonatal weight, and length of maternal and neonatal hospital stays for those with and without elevated office BP. RESULTS: After application of exclusion criteria, data for 7 women were removed from the analysis. For the remaining subjects, in the group with elevated BP, prevalence of WCH was 29.2% (42/144). Duration of pregnancy was similar in the normotensive and WCH groups (39.6 vs 39.8 weeks; P = .50), but shorter (38.3 weeks; P<.001) in the true hypertension group. Incidence of preeclampsia was similar in the normotensive and WCH groups (5.8% vs 7.1 %; P = .86) but higher in the true hypertension group (61.7%; P<.001). Frequency of cesarean delivery was lower in the normotensive (12.4%) than in the WCH (45.2%; P = .008) and true hypertension (41.1 %; P = .009) groups. Neonatal weight was lower (P<.001) in the true hypertension (mean, 2911 g) than in the normotensive (3336 g) and WCH groups (3435 g), which did not differ (P = .68). The duration of neonatal hospital stay did not differ between the normotensive and the WCH group (5.3 vs 6.9 days; P = .13) but was longer in the true hypertension group (12.3 days; P<.001). CONCLUSIONS: In women with elevated BP during their third trimester of pregnancy, 24-hour BP was superior to office BP (distinguishing true hypertension from WCH) for prediction of the outcome of pregnancy. Outcomes in the normotensive and WCH group were comparable, but the increased incidence of cesarean delivery in the WCH group may reflect decision-making processes influenced by office BP. PMID- 10535436 TI - Evaluation of conflict of interest in economic analyses of new drugs used in oncology. AB - CONTEXT: Recent studies have found that when investigators have financial relationships with pharmaceutical or product manufacturers, they are less likely to criticize the safety or efficacy of these agents. The effects of health economics research on pharmaceutical company revenue make drug investigations potentially vulnerable to this bias. OBJECTIVE: To determine whether there is an association between pharmaceutical industry sponsorship and economic assessment of oncology drugs. DESIGN: MEDLINE and HealthSTAR databases (1988-1998) were searched for original English-language research articles of cost or cost effectiveness analyses of 6 oncology drugs in 3 new drug categories (hematopoietic colony-stimulating factors, serotonin antagonist antiemetics, and taxanes), yielding 44 eligible articles. Two investigators independently abstracted each article based on specific criteria. MAIN OUTCOME MEASURE: Relationships between funding source and (1) qualitative cost assessment (favorable, neutral, or unfavorable) and (2) qualitative conclusions that overstated quantitative results. RESULTS: Pharmaceutical company-sponsored studies were less likely than nonprofit-sponsored studies to report unfavorable qualitative conclusions (1/20 [5%] vs 9/24 [38%]; P = .04), whereas overstatements of quantitative results were not significantly different in pharmaceutical company-sponsored (6/20 [30%]) vs nonprofit-sponsored (3/24 [13%]) studies (P = .26). CONCLUSIONS: Although we did not identify bias in individual studies, these findings indicate that pharmaceutical company sponsorship of economic analyses is associated with reduced likelihood of reporting unfavorable results. PMID- 10535437 TI - Why don't physicians follow clinical practice guidelines? A framework for improvement. AB - CONTEXT: Despite wide promulgation, clinical practice guidelines have had limited effect on changing physician behavior. Little is known about the process and factors involved in changing physician practices in response to guidelines. OBJECTIVE: To review barriers to physician adherence to clinical practice guidelines. DATA SOURCES: We searched the MEDLINE, Educational Resources Information Center (ERIC), and HealthSTAR databases (January 1966 to January 1998); bibliographies; textbooks on health behavior or public health; and references supplied by experts to find English-language article titles that describe barriers to guideline adherence. STUDY SELECTION: Of 5658 articles initially identified, we selected 76 published studies describing at least 1 barrier to adherence to clinical practice guidelines, practice parameters, clinical policies, or national consensus statements. One investigator screened titles to identify candidate articles, then 2 investigators independently reviewed the texts to exclude articles that did not match the criteria. Differences were resolved by consensus with a third investigator. DATA EXTRACTION: Two investigators organized barriers to adherence into a framework according to their effect on physician knowledge, attitudes, or behavior. This organization was validated by 3 additional investigators. DATA SYNTHESIS: The 76 articles included 120 different surveys investigating 293 potential barriers to physician guideline adherence, including awareness (n = 46), familiarity(n = 31), agreement (n = 33), self-efficacy (n = 19), outcome expectancy (n = 8), ability to overcome the inertia of previous practice (n = 14), and absence of external barriers to perform recommendations (n = 34). The majority of surveys (70 [58%] of 120) examined only 1 type of barrier. CONCLUSIONS: Studies on improving physician guideline adherence may not be generalizable, since barriers in one setting may not be present in another. Our review offers a differential diagnosis for why physicians do not follow practice guidelines, as well as a rational approach toward improving guideline adherence and a framework for future research. PMID- 10535438 TI - Legal issues concerning electronic health information: privacy, quality, and liability. AB - Personally identifiable health information about individuals and general medical information is increasingly available in electronic form in health databases and through online networks. The proliferation of electronic data within the modern health information infrastructure presents significant benefits for medical providers and patients, including enhanced patient autonomy, improved clinical treatment, advances in health research and public health surveillance, and modern security techniques. However, it also presents new legal challenges in 3 interconnected areas: privacy of identifiable health information, reliability and quality of health data, and tortbased liability. Protecting health information privacy (by giving individuals control over health data without severely restricting warranted communal uses) directly improves the quality and reliability of health data (by encouraging individual uses of health services and communal uses of data), which diminishes tort-based liabilities (by reducing instances of medical malpractice or privacy invasions through improvements in the delivery of health care services resulting in part from better quality and reliability of clinical and research data). Following an analysis of the interconnectivity of these 3 areas and discussing existing and proposed health information privacy laws, recommendations for legal reform concerning health information privacy are presented. These include (1) recognizing identifiable health information as highly sensitive, (2) providing privacy safeguards based on fair information practices, (3) empowering patients with information and rights to consent to disclosure (4) limiting disclosures of health data absent consent, (5) incorporating industry-wide security protections, (6) establishing a national data protection authority, and (7) providing a national minimal level of privacy protections. PMID- 10535439 TI - Political considerations for changing medical screening programs. PMID- 10535440 TI - Conflict of interest and cost-effectiveness analysis. PMID- 10535441 TI - Resolving disagreements in the patient-physician relationship: tools for improving communication in managed care. AB - Managed care uses financial incentives and restrictions on tests and procedures to attempt to influence physician decision making and limit costs. Increasingly, the public is questioning whether physicians are truly making decisions based on the patient's best interest or are unduly influenced by economic incentives. These circumstances lead to the potential for disagreements and conflict in the patient-physician relationship. We convened a group of individuals in October 1998, including patient representatives, leaders from health care organizations, practicing physicians, communication experts, and medical ethicists, to articulate the types of disagreements emerging in the patient-physician relationship as a result of managed care. We addressed 3 specific scenarios physicians may encounter, including allocation, illustrated by a patient who is referred to a different ophthalmologist based on a new arrangement in the physician's group; access, illustrated by a patient who wishes to see his own physician for a same-day visit rather than a nurse specialist; and financial incentives, illustrated by a patient who expects to have a test performed and a physician who does not believe the test is necessary but is afraid the patient will think the physician is not ordering the test because of financial incentives. Using these scenarios, we suggest communication strategies that physicians can use to decrease the potential for disagreements. In addition, we propose strategies that health plans or physician groups can use to alleviate or resolve these disagreements. PMID- 10535442 TI - JAMA Patient Page: respiratory problems. PMID- 10535443 TI - Targeting autoimmune diabetes with gene therapy. AB - The autoimmune nature of insulin-dependent, or type 1, diabetes targets the beta cells of the pancreas for destruction and results in a lifelong commitment to insulin replacement therapy. Although the number of formulations and dosing of insulin have become more sophisticated and more efficient in recent years, insulin therapy alone is unable to prevent nephropathy, retinopathy, or vascular and heart disease, which still occur in a large number of patients. Different approaches have been attempted to eliminate the requirement of exogenous insulin administration. Historically, these have included pancreatic and islet transplants, which were later combined with treatments intended to halt the destructive process directed against the islets. Despite significant advances made in all of these areas, each approach faces a hostile immunological response that frequently ends with the loss of the islets. Gene therapy-based approaches add a new dimension to the efforts aimed at specifically blocking the immunological attack against the islets in genetically at-risk individuals (autoimmunity) or the immunological response against transplanted allogeneic islets (rejection). This new technology may have an important role in the therapy and cure of type 1 diabetes. PMID- 10535444 TI - Insulin signaling in insulin receptor substrate (IRS)-1-deficient brown adipocytes: requirement of IRS-1 for lipid synthesis. AB - Immortalized fetal brown adipocyte cell lines have been generated from homozygous (-/-) and heterozygous (+/-) insulin receptor substrate (IRS)-1-deficient mice, as well as from wild-type mice (+/+). Under growing conditions, these cell lines maintained the expression of the adipogenic marker fatty acid synthase and uncoupling protein-1, a tissue-specific thermogenic marker. The IRS-1 (-/-) brown adipocytes lacked IRS-1 protein expression and had a significant increase in IRS 2 protein expression. Insulin-induced tyrosine phosphorylation of IRS-1 was reduced by 50% in heterozygous IRS-1-deficient cells and was totally absent in homozygous cells, while tyrosine phosphorylation of IRS-2 showed a gradual increase. Insulin receptor alpha-subunit protein content and beta-subunit tyrosine kinase activity remained unchanged upon insulin stimulation, regardless of the lack of IRS-1. Brown adipocytes from homozygous IRS-1-deficient mice showed no IRS-1-associated p85alpha subunit of phosphatidylinositol 3-kinase (PI 3-kinase) or IRS-1-associated PI 3-kinase activity in response to insulin, but exhibited enhanced IRS-2-associated p85alpha subunit and IRS-2-associated PI 3 kinase activity. Overall insulin-induced PI 3-kinase activity associated to antiphosphotyrosine immune complexes was decreased by 30% in the homozygous IRS-1 deficient brown adipocytes. Downstream PI 3-kinase, activated Akt (protein kinase B) was decreased by 92% in an insulin-stimulated homozygous IRS-1-deficient brown adipocyte cell line, whereas the expression of Akt was similar in the three cell lines. However, activated p70 S6 kinase (p70s6k) remained unchanged. Although brown adipocyte cell lines showed similar cytosolic lipid content in the presence of 10% fetal calf serum, cytosolic lipid content was reduced in both serum deprived heterozygous and homozygous IRS-1-deficient cells. Insulin treatment for 24 h doubled the cytosolic lipid content in wild-type and heterozygous IRS-1 deficient brown adipocyte cell lines but failed to increase the cytosolic lipid content in homozygous IRS-1-deficient cells. Our results strongly suggest that IRS-1/PI 3-kinase/Akt activation is an essential requirement for insulin stimulation of lipid synthesis in brown adipocytes. PMID- 10535445 TI - Hypothalamic orexin expression: modulation by blood glucose and feeding. AB - Orexins (hypocretins), novel peptides expressed in specific neurons of the lateral hypothalamic area (LHA), stimulate feeding when injected intracerebroventricularly. We investigated their role in feeding in the rat by measuring hypothalamic prepro-orexin mRNA levels under contrasting conditions of increased hunger. Prepro-orexin mRNA levels increased significantly after 48 h of fasting (by 90-170%; P < 0.05) and after acute (6 h) hypoglycemia when food was withheld (by 90%; P < 0.02). By contrast, levels were unchanged during chronic food restriction, streptozotocin-induced diabetes, hypoglycemia when food was available, voluntary overconsumption of palatable food, or glucoprivation induced by systemic 2-deoxy-D-glucose. Orexin expression was not obviously related to changes in body weight, insulin, or leptin, but was stimulated under conditions of low plasma glucose in the absence of food. Orexins may participate in the short-term regulation of energy homeostasis by initiating feeding in response to falls in glucose and terminating it after food ingestion. The LHA is known to contain neurons that are stimulated by falls in circulating glucose but inhibited by feeding-related signals from the viscera; orexin neurons may correspond to this neuronal population. PMID- 10535447 TI - Lack of association between duration of breast-feeding or introduction of cow's milk and development of islet autoimmunity. AB - The hypothesis that early exposure to cow's milk or lack of breast-feeding predisposes to type 1 diabetes remains controversial. We aimed to determine prospectively the relationship of, first, duration of exclusive breast-feeding and total duration of breast-feeding, and second, introduction of cow's milk protein as infant formula, cow's milk, or dairy products, to the development of islet antibodies in early life. Some 317 children with a first-degree relative with type 1 diabetes were followed prospectively from birth for 29 months (4-73). Mothers kept a home diary and answered infant feeding questionnaires at 6-month intervals. No systematic feeding advice was given. Insulin autoantibodies (normal range <5.5%), anti-GAD antibodies (<5.0 U), and anti-IA2 antibodies (<3.0 U) were measured at 6-month intervals. Cox proportional hazards model of survival analysis detected no significant difference between children who did not develop islet antibodies (225 of 317 [71%]), children with one islet antibody raised once (52 of 317 [16.4%]), children with one antibody raised repeatedly (18 of 317 [5.7%]), or children with two or more antibodies raised (22 of 317 [6.9%]), in terms of duration of exclusive breast-feeding, total duration of breast-feeding, or introduction of cow's milk-based infant formulas, cow's milk, or dairy products (relative risk: 0.91-1.09). Four of the children with two or more islet antibodies developed type 1 diabetes. We conclude that there is no prospective association between duration of breast-feeding or introduction of cow's milk and the development of islet autoimmunity in high-risk children. PMID- 10535446 TI - Diabetes-prone and diabetes-resistant BB rats share a common major diabetes susceptibility locus, iddm4: additional evidence for a "universal autoimmunity locus" on rat chromosome 4. AB - Diabetes-prone (DP) BB rats develop autoimmune type 1 diabetes spontaneously. At least five loci are linked to disease expression: the major histocompatibility complex (iddm2), two susceptibility loci (iddm4, iddm5), and, possibly, a resistance locus (iddm3). Spontaneous disease also requires homozygosity for lyp/iddm1, which causes lymphopenia. It has not been determined whether lyp/iddm1 is required for predisposition to diabetes autoimmunity in addition to being required for its spontaneous expression. We analyzed backcross rats segregating for diabetes but not lymphopenia using Wistar-Furth (WF) and diabetes-resistant (DR) BB animals. The latter are nonlymphopenic (lyp+/+) and develop diabetes only in response to immunological perturbants. Treatment of (DR-BB x WF)F1 x WF animals (all lyp+/+) using a standard induction protocol caused type 1 diabetes in 58% of progeny. Expression of type 1 diabetes was strongly linked to iddm4. The results suggest that lyp/iddm1 does not determine the predisposition to autoimmunity in BB rats and that iddm4 is a major diabetogenic locus in both DP- and DR-BB animals. The iddm4 gene maps to a region containing several major autoimmunity loci, including aia2, aia3, and cia3. We propose that BB rat diabetes requires 1) class II RT1u (iddm2) for susceptibility, 2) additional loci for disease initiation and progression in response to perturbants, and 3) lyp for spontaneous disease. PMID- 10535448 TI - Oral administration of cholera toxin B-insulin conjugates protects NOD mice from autoimmune diabetes by inducing CD4+ regulatory T-cells. AB - Restoration of peripheral tolerance to target autoantigens during autoimmune diseases has met with several limitations because of the limited efficacy of this approach in an already immune host. To optimize the induction of tolerance, we have shown that feeding insulin conjugated to cholera toxin B-subunit (CTB), a potent mucosal adjuvant, reduced by 5,000 the amounts of antigen necessary for delaying diabetes onset in NOD mice. To analyze these protective mechanisms, we have performed cotransfer experiments using splenocytes from young females fed once with 10 microg of CTB-insulin, mixed with diabetogenic T-cells, and intravenously injected into irradiated syngeneic male recipients. We demonstrated that the delayed onset of diabetes relied on CD4+ T-cells. We studied the cytokine production from plate-bound anti-CD3-stimulated cells. Higher interleukin (IL)-4 amounts were observed in both splenocytes and pancreatic lymph node (PLN) cell cultures from CTB-insulin-fed mice as soon as 4 h after the feeding. An increase in the levels of transforming growth factor-beta was seen after 24 h only in the mesenteric lymph nodes (MLN). In both of these organs, a reduction of gamma-interferon (IFN-gamma) production occurred after CTB-insulin treatment, at 24 h in the PLN and at 7 days in the MLN. Reverse transcription polymerase chain reaction analysis indicated an increase in the level of IL-4 and a reduction in IFN-gamma transcripts in the PLN of mice treated orally with CTB insulin and of the recipients of regulatory T-cells. Using different strains of congenic NOD mice at the Thy1 locus, we showed that protection was associated with the accumulation of T-cells from CTB-insulin-fed mice in the lymph nodes from draining sites containing functional islets, i.e., the PLN in normal mice and the renal lymph nodes after a syngeneic islet graft under the kidney capsule of streptozotocin-treated mice. Taken together, our results clearly indicate that oral administration of CTB-insulin conjugates in NOD mice produced a shift from a T-helper type 1 to a type 2 profile with the induction of antigen-specific regulatory CD4+ T-cells in the vicinity of the mucosal barrier and close to the inflamed islets. PMID- 10535449 TI - Peptides derived from murine insulin are diabetogenic in both rats and mice, but the disease-inducing epitopes are different: evidence against a common environmental cross-reactivity in the pathogenicity of type 1 diabetes. AB - Two rodent models of autoimmune type 1 diabetes have been used to investigate the role of insulin as an autoantigen in this disease. In lymphopoenia-induced diabetes in the PVG.RT1u rat, neonatal tolerization with insulin B-chain peptides, but not A-chain peptides, conferred significant protection from disease. After rechallenge of adult rats, neonatally B-chain-tolerized animals showed diminished B-chain-specific T-cell proliferation, interleukin (IL)-2 production, and interferon-gamma (IFN-gamma) production, as compared with control animals. The epitope recognized by the PVG.RT1u rat was mapped to residues 1-18 of the B-chain; T-cell lines specific for this epitope were generated, and these conferred diabetes upon adoptive transfer to irradiated syngeneic recipients. In adult nonobese diabetic (NOD) mice, subcutaneous immunization with B-chain peptide 9-23 emulsified in incomplete Freund's adjuvant (IFA) was also potent at preventing onset of diabetes. In contrast to PVG.RT1u rats, NOD mice recognized an epitope within residues 10-29 of the insulin B-chain. The data implicate insulin as a target autoantigen in type 1 diabetes but do not support a role for molecular mimicry to insulin in the pathogenesis of this disease. PMID- 10535450 TI - Heterophile anti-mouse immunoglobulin antibodies may interfere with cytokine measurements in patients with HLA alleles protective for type 1A diabetes. AB - Wilson and coworkers (Wilson SB, Kent SC, Patton KT, Orban T, Jackson RA, Exley M, Porcelli S, Schatz DA, Atkinson MA, Balk SP, Strominger JL, Hafler DA: Extreme Th1 bias of invariant V alpha24J alpha Q T-cells in type 1 diabetes. Nature 391:177-181, 1998) have recently reported raised serum levels of interleukin-4 (IL-4) in anti-islet autoantibody-positive first-degree relatives of patients with type 1A diabetes who did not progress to diabetes. Protection from diabetes has been noted for several human lymphocyte antigen (HLA) alleles, such as HLA DR2-DQA1*0102-DQB1*0602. We, therefore, wanted to determine whether this cytokine phenotype was associated with HLA genes protective for type 1A diabetes. We used a two-site fluoroimmunoassay with the same monoclonal antibodies as those reported by Wilson et al. Using this assay, we have found evidence for human heterophile antibodies mimicking serum IL-4: all serum IL-4 reactivity was lost if mouse serum or mouse immunoglobulin were added to the assay; serum IL-4 activity was bound and then eluted by protein A/G chromatography; and levels of anti-mouse antibodies correlated with apparent serum IL-4. This pseudo-IL-4 activity was found in a subset of control subjects, patients with type 1A diabetes, and their relatives and was primarily associated with specific HLA alleles protective for type 1A diabetes (e.g., DQB1*0602). After adjustment for HLA, positive levels of heterophile antibodies were not associated with protection from diabetes. The confounding effect of protective HLA alleles associated with heterophile antibodies could explain the previously reported association between raised serum IL-4 and protection from type 1A diabetes. The mechanism by which specific DQ alleles protect from diabetes and are associated with increased heterophile antibodies is currently unknown. PMID- 10535452 TI - Disruption of circadian insulin secretion is associated with reduced glucose uptake in first-degree relatives of patients with type 2 diabetes. AB - The objective of this study was to evaluate whether first-degree relatives (FDRs) of patients with type 2 diabetes had abnormal circadian insulin secretion and, if so, whether this abnormality affected their glucose metabolism. Six African American FDRs with normal glucose tolerance and 12 matched normal control subjects (who had no family history of diabetes) were exposed to 48 h of hyperglycemic clamping (approximately 12 mmol/l). Insulin secretion rates (ISRs) were determined by deconvolution of plasma C-peptide levels using individual C peptide kinetic parameters. Detrending and smoothing of data (z-scores) and computation of autocorrelation functions were used to identify ISR cycles. During the initial hours after start of glucose infusions, ISRs were approximately 60% higher in FDRs than in control subjects (585 vs. 366 nmol/16 h, P < 0.05), while rates of glucose uptake were the same (5.6 mmol x kg(-1) x h(-1)), indicating that the FDRs were insulin resistant. Control subjects had well-defined circadian (24 h) cycles of ISR and plasma insulin that rose in the early morning, peaked in the afternoon, and declined during the night. In contrast, FDRs had several shorter ISR cycles of smaller amplitude that lacked true periodicity. This suggested that the lack of a normal circadian ISR increase had made it impossible for the FDRs to maintain their compensatory insulin hypersecretion beyond 18 h of hyperglycemia. As a result, ISR decreased to the level found in control subjects, and glucose uptake fell below the level of control subjects (61 vs. 117 micromol x kg(-1) x min(-1), P < 0.05). In summary, we found that FDRs with normal glucose tolerance had defects in insulin action and secretion. The newly recognized insulin secretory defect consisted of disruption of the normal circadian ISR cycle, which resulted in reduced insulin secretion (and glucose uptake) during the ascending part of the 24 h ISR cycle. PMID- 10535451 TI - Multiple sites of purinergic control of insulin secretion in mouse pancreatic beta-cells. AB - In mouse pancreatic beta-cells, extracellular ATP (0.1 mmol/l) effectively reduced glucose-induced insulin secretion. This inhibitory action resulted from a direct interference with the secretory machinery, and ATP suppressed depolarization-induced exocytosis by 60% as revealed by high-resolution capacitance measurements. Suppression of Ca2+-dependent exocytosis was mediated via binding to P2Y1 purinoceptors but was not associated with inhibition of the voltage-dependent Ca2+ currents or adenylate cyclase activity. Inhibition of exocytosis by ATP resulted from G-protein-dependent activation of the serine/threonine protein phosphatase calcineurin and was abolished by cyclosporin A and deltamethrin. In contrast to the direct inhibitory action on exocytosis, ATP reduced the whole-cell ATP-sensitive K+ (K(ATP)) current by 30% (via activation of cytosolic phospholipase A2), leading to membrane depolarization and stimulation of electrical activity. The stimulatory effect of ATP also involved mobilization of Ca2+ from thapsigargin-sensitive intracellular stores. We propose that the inhibitory action of ATP, by interacting with the secretory machinery at a level downstream to an elevation in [Ca2+]i, is important for autocrine regulation of insulin secretion in mouse beta-cells. PMID- 10535453 TI - Resistance of ALR/Lt islets to free radical-mediated diabetogenic stress is inherited as a dominant trait. AB - ALS/Lt and ALR/Lt are inbred mouse strains selected for susceptibility and resistance to alloxan (AL)-induced diabetes. Within 24-h after AL administration in vivo, ALS/Lt islets were distinguished from ALR/Lt islets by more extensive necrotic changes. Within 7 days post-AL, ALS/Lt mice exhibited hyperglycemia and hypoinsulinemia, whereas ALR/Lt mice maintained normal plasma insulin and glucose levels. We have recently shown that resistance in ALR/Lt correlated with constitutively elevated systemic (and pancreatic) free radical defense status. In the present report, we examined whether ability to detoxify free radical stress extended to the level of ALR/Lt pancreatic islets. Cultured ALS/Lt islets exposed for 5 min to increasing (0-3 mmol/l) AL concentrations in vitro exhibited an 80% decline in numbers of intact islets after a subsequent 6-day culture period, as well as a 75% reduction in islet insulin content and a 94% decrease in glucose stimulated insulin secretory capacity. In contrast, ALR/Lt islets remained viable and retained glucose-stimulated insulin secretory capacity as well as normal insulin content. This ALR/Lt islet resistance extended to hydrogen peroxide, a free radical generator whose entry into beta-cells is not dependent on glucose transporters. The elevated antioxidant defenses previously found in ALR/Lt pancreas were extended to isolated islets, which exhibited significantly higher glutathione and Cu-Zn superoxide dismutase 1 levels compared with ALS/Lt islets. A dominant genetic trait from ALR/Lt controlling this unusual AL resistance was indicated by the finding that reciprocal F1 mice of both sexes were resistant to AL administration in vivo. A backcross to ALS/Lt showed 1:1 segregation for susceptibility/resistance, indicative of a single gene controlling the phenotype. In conclusion, the ALR/Lt mouse may provide important insight into genetic mechanisms capable of rendering islets strongly resistant to free radical mediated damage. PMID- 10535454 TI - Metabolic characteristics of individuals with impaired fasting glucose and/or impaired glucose tolerance. AB - With the release of the new 1997 American Diabetes Association diagnostic criteria, a new category was introduced, termed "impaired fasting glucose" (IFG). The metabolic abnormalities of individuals with IFG, compared with those with impaired glucose tolerance (IGT) (World Health Organization criteria), remain to be elucidated. We assessed insulin action (hyperinsulinemic clamp), insulin secretion (25-g intravenous glucose tolerance test), and endogenous glucose output (EGO) (3-(3)H-glucose) in 434 nondiabetic Pima Indians with either normal (NFG; <6.1 mmol/l) or impaired (IFG; 6.1-7.0 mmol/l) fasting glucose and with either normal (NGT; 2-h glucose <7.8 mmol/l) or impaired (IGT; 2-h glucose 7.8 11.1 mmol/l) glucose tolerance: NFG/NGT (n = 307), IFG/NGT (n = 11), NFG/IGT (n = 98), and IFG/IGT (n = 18). Compared with the NFG/NGT group, individuals with IFG/NGT had lower maximal insulin-stimulated glucose disposal (M; -20%, P < 0.01), a lower acute insulin response (AIR) to intravenous glucose (-33%, P < 0.05), and higher EGO (8%, P = 0.055). Individuals with NFG/IGT also had lower M (-21%, P < 0.001) and lower AIR (-8%, P < 0.05), but normal EGO (-1%, NS). Individuals with IFG/IGT showed the most severe abnormalities in M (-27%), AIR ( 51%), and EGO (+13%) (all P < 0.001 compared with NFG/NGT). These group differences could be explained by the observation that AIR and EGO, but not M, were more strongly related to the fasting than to the 2-h glucose concentration. Thus, Pima Indians with isolated IFG and isolated IGT show similar impairments in insulin action, but those with isolated IFG have a more pronounced defect in early insulin secretion and, in addition, increased EGO. More severe metabolic abnormalities are present in Pima Indians with combined IFG and IGT. PMID- 10535455 TI - Inhibitory effects of leptin-related synthetic peptide 116-130 on food intake and body weight gain in female C57BL/6J ob/ob mice may not be mediated by peptide activation of the long isoform of the leptin receptor. AB - We recently reported that intraperitoneal administration of leptin-related synthetic peptide 116-130 [LEP-(116-130)] resulted in reduced food intake and significant weight loss in homozygous female C57BL/6J ob/ob mice. In this study, we used two in vitro bioassays to show that the interaction of LEP-(116-130) with the long form of the leptin receptor (OB-Rb), the receptor isoform that is predominantly expressed in the hypothalamus, is not required for the observed in vivo effects of the peptide on energy balance. LEP-(116-130) was unable to compete the binding of alkaline phosphatase-leptin fusion protein to OB-R. Moreover, LEP-(116-130) was unable to activate signal transduction by OB-Rb in vitro. In homozygous female C57BLKS/J-m db/db mice that do not express OB-Rb, intraperitoneal administration of LEP-(116-130) reduced body weight gain and blood glucose levels but not food intake, which further supports a mechanism of action that does not require peptide interaction with OB-Rb. PMID- 10535456 TI - Phenotypic characteristics associated with insulin resistance in metabolically obese but normal-weight young women. AB - Metabolically obese, normal-weight (MONW) individuals are a hypothesized subgroup of the general population. These normal-weight individuals potentially display a cluster of obesity-related features, although this has not been systematically tested in young women. We hypothesized that MONW young women would display higher levels of total and visceral fat and lower levels of physical activity than normal women. In a cohort of 71 healthy nonobese women (21-35 years old), we identified MONW women based on cut points for insulin sensitivity (normal = glucose disposal >8 mg x min(-1) x kg(-1) of fat-free mass [FFM], n = 58; impaired = glucose disposal <8 ml x min(-1) x kg(-1) of FFM, n = 13). Thereafter, we measured body composition (dual energy X-ray absorptiometry) and body fat distribution (computed tomography), cardiorespiratory fitness (VO2max on a treadmill), physical activity energy expenditure (doubly labeled water and indirect calorimetry), glucose tolerance (oral glucose tolerance test), serum lipid profile, and dietary intake. We found a higher body fat percentage (32 +/- 6 vs. 27 +/- 6%, P = 0.01) and higher subcutaneous (213 +/- 61 vs. 160 +/- 78 cm2, P = 0.03) and visceral (44 +/- 16 vs. 35 +/- 14 cm2, P < 0.05) abdominal adiposity in the MONW group versus the normal group. The MONW group showed a lower physical activity energy expenditure (2.66 +/- 0.92 vs. 4.39 +/- 1.50 MJ/day, P = 0.01), but no difference in cardiorespiratory fitness was noted between groups. In conclusion, despite a normal body weight, a subset of young, apparently healthy women displayed a cluster of risky phenotypic characteristics that, if left untreated, may eventually predispose them to type 2 diabetes and cardiovascular disease. PMID- 10535457 TI - Diabetes induces an impairment in the proteolytic activity against oxidized proteins and a heterogeneous effect in nonenzymatic protein modifications in the cytosol of rat liver and kidney. AB - It is assumed that increased oxidative stress contributes to the development of complications in diabetes. In this study, several markers of protein structural modifications directly induced by free radicals were investigated in the liver and kidney cytosolic fractions of rats with streptozotocin-induced diabetes. Sulfydryl residue and side-chain amino group analyses, as well as immunoblotting and chromatographic measurements of protein-bound carbonyl, suggest that protein oxidative modification is not increased by diabetes, with the exception of sulfydryl groups in renal cytosol. The levels of the glycation-derived carbonyl N epsilon-fructosyl-lysine are significantly increased by diabetes. Furthermore, unchanged proteolytic activity against in vivo-oxidized proteins, significant decreases both in activity against H2O2-modified proteins and in proteasome activity, measured by the degradation of a specific fluorogenic substrate, suggest that the unchanged oxidative protein modification in the diabetic state cannot be attributed to an increased cytosolic proteolytic activity in these tissues. These results provide evidence against a generalized increase in protein oxidative damage and demonstrate a diabetes-induced alteration in cytosolic proteolytic pathways, suggesting that proteasome activity may be impaired in these organs. PMID- 10535458 TI - Modifications induced by LDL from type 1 diabetic patients on endothelial cells obtained from human umbilical vein. AB - The aim of the present work was to analyze the effect of LDL obtained from type 1 diabetic patients in good metabolic control on human umbilical vein endothelial cells (HUVECs) after a short incubation period to detect possible atherogenic modifications of endothelial properties. Cultured HUVECs were incubated for 3 h with culture medium alone (control HUVEC), with native LDL from 12 healthy men (control LDL), or with native LDL from 12 type 1 diabetic men (type 1 LDL) (100 pg/ml). After the incubation, the following parameters were evaluated: nitric oxide synthase (NOS) activity, cytoplasmic Ca2+ levels, Na+-K+-ATPase activity, plasma membrane fluidity determined by means of 1,6-diphenyl-1,3,5-hexatriene (DPH) and 1-(4-trimethylaminophenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH), and plasma membrane conjugated diene (CD) content. The same experiments were repeated after bradykinin stimulation or in the presence of the antioxidant butylated hydroxytoluene (BHT), and nitric oxide (NO) production in intact HUVECs was also evaluated. HUVECs incubated with control LDL in comparison with control HUVECs showed a decreased fluidity of the membrane surface evaluated by TMA-DPH and a higher CD content. These alterations were prevented by the presence of BHT. HUVECs incubated with type 1 LDL in comparison with both control HUVECs and cells incubated with control LDL showed 1) increased NOS and Na+-K+-ATPase activity, cytoplasmic Ca2+ levels, and CD content, and 2) decreased fluidity of the membrane surface evaluated by TMA-DPH. These modifications were blunted--but not abolished--by the presence of BHT. After bradykinin stimulation either in the absence or in the presence of BHT, both cytoplasmic Ca2+ levels and NO production were increased in control HUVECs and in HUVECs incubated with control LDL, while a reduced response was observed in HUVECs incubated with type 1 LDL. The alterations observed in the endothelial function after the cell-LDL interaction might play a central role in the atherogenic process in diabetes. PMID- 10535459 TI - Increased renal expression of vascular endothelial growth factor (VEGF) and its receptor VEGFR-2 in experimental diabetes. AB - It has been suggested that the cytokine vascular endothelial growth factor (VEGF) has an important role in the pathogenesis of diabetic retinopathy, but its role in nephropathy has not been clearly demonstrated. Assessment of VEGF, 125I-VEGF binding, and vascular endothelial growth factor receptor-2 (VEGFR-2) in the kidney was performed after 3 and 32 weeks of streptozotocin-induced diabetes. Gene expression of both VEGF and VEGFR-2 was assessed by Northern blot analysis and the localization of the ligand and receptor was examined by in situ hybridization. VEGF and VEGFR-2 protein were also evaluated by immunohistochemistry. Binding of the radioligand 125I-VEGF was evaluated by in vitro and in vivo autoradiography. Diabetes was associated with increased renal VEGF gene expression. VEGF mRNA and protein were localized to the visceral epithelial cells of the glomerulus and to distal tubules and collecting ducts in both diabetic and nondiabetic rats. Renal VEGFR-2 mRNA was increased after 3 weeks of diabetes but not in long-term diabetes. In situ hybridization and immunohistochemical studies revealed that glomerular endothelial cells were the major site of VEGFR-2 expression. In addition, VEGFR-2 gene expression was detected in cortical and renomedullary interstitial cells and on endothelial cells of peritubular capillaries. There was an increase in 125I-VEGF binding sites after 3 but not 32 weeks of diabetes. The major VEGF binding sites were in the glomeruli. 125I-VEGF binding was also observed in medullary rays and in the renal papillae. These studies indicate an early and persistent increase in renal VEGF gene expression in association with experimental diabetes. In addition, an early and transient increase in renal VEGF receptors was also observed in diabetic rats. These findings are consistent with a role for VEGF in mediating some of the changes observed in the diabetic kidney. PMID- 10535460 TI - Adverse effects of hyperglycemia on kidney development in rats: in vivo and in vitro studies. AB - Congenital malformations occur more frequently in the offspring of diabetic mothers. These in vivo and in vitro studies investigate the potential adverse effects of hyperglycemia on kidney development in the rat. Female rats were made hyperglycemic throughout gestation with a single injection of streptozotocin (STZ) on day 0 of gestation, or for a short period encompassing the early stage of renal organogenesis by infusing glucose from gestational days 12-16. Kidney development in the pups was assessed by determining the total number of nephrons formed in the kidney. The number of nephrons was significantly reduced (10-35%) in the pups from STZ-treated dams, as a function of hyperglycemia. There were also fewer nephrons in pups from dams given glucose infusion whose hyperglycemia was transiently higher on day 13 of gestation. The in vitro experiments were done on metanephroi removed from 14-day-old fetuses and grown for 6 days in medium containing 0, 6.9, 13.8, or 27.5 mmol/l glucose. The development of explants grown in 0, 13.8, and 27.5 mmol/l glucose was impaired compared with that of explants grown in the 6.9 mmol/l control medium, showing that the glucose concentration must be closely controlled to ensure optimum in vitro metanephros development. Thus, exposure to hyperglycemia in utero can cause a nephron deficit, which in turn may have renal consequences later in life. PMID- 10535461 TI - Evidence of a novel type 2 diabetes locus 50 cM centromeric to NIDDM2 on chromosome 12q. AB - To replicate the recent finding of a type 2 diabetes locus (NIDDM2) on 12q, families segregating early-onset autosomal-dominant type 2 diabetes were screened for linkage. Included were 26 Caucasian and 6 non-Caucasian pedigrees with an average age at diabetes diagnosis of 37 +/- 18 years. Affected (n = 233) and nonaffected (n = 152) family members were genotyped for 17 markers covering 90 cM on chromosome 12q. While no evidence for linkage was detected at the NIDDM2 locus, a linkage peak was observed 50 cM centromeric to NIDDM2 at markers D12S375 and D12S1052. In a nonparametric analysis, the Z(all) score was 2.9 (P = 0.015) at D12S375, and increased to 3.8 (P = 0.007) among Caucasian families. Further increase in significance was observed in pedigrees with poor insulin response, with a maximum Z(all) of 6.2 (P = 0.002) at D12S375. Suggestive evidence of linkage was also detected by the parametric analysis, with the heterogeneity logarithm of odds score peaking at 2.5 (alpha = 0.15) between D12S375 and D12S1052. In summary, our data indicate that the NIDDM2 locus does not play a major role in early-onset autosomal-dominant type 2 diabetes. Rather, they strongly suggest that a previously undetected type 2 diabetes locus exists 50 cM from NIDDM2 on 12q. PMID- 10535462 TI - Hepatocellular carcinoma after sustained response to interferon in non-cirrhotic hepatitis C: flaws in the cure, or a clue to the flaws? PMID- 10535463 TI - The haemochromatosis gene: a global perspective and implications for the Asia Pacific region. AB - Mutations in the haemochromatosis (HFE) gene cause most of the cases of hereditary haemochromatosis among people of Northern European ancestry while remaining a rare cause of iron overload among indigenous persons of the Asia Pacific region. Advances in understanding of the role of the HFE protein product and other recently cloned iron transporters signify an exciting period, as previously unknown components of the iron metabolism pathway are revealed one by one. Epidemiological studies have shown that this gene is more widespread than its phenotypic expression would suggest and that the heterozygous state may be implicated in the expression of other diseases of the liver such as porphyria cutanea tarda, hepatitis C virus infection and non-alcoholic steatohepatitis. The diagnosis, management and ethical implications for clinical practice in the aftermath of this discovery are discussed. PMID- 10535464 TI - Helicobacter pylori and extra-digestive diseases. AB - Helicobacter pylori is a recently rediscovered gram-negative bacteria that causes peptic ulcer disease, gastric lymphoma and gastric carcinoma. Helicobacter pylori achieves its pathogenetic role by triggering an intense leucocyte infiltration of the gastric submucosa which is mediated by proinflammatory cytokines. This pathogenetic mechanism is common to many other diseases and, therefore, Helicobacter pylori seroprevalence has also been investigated in other diseases. It is now known that H. pylori seropositivity is associated with an increasing number of cardiovascular, respiratory, extra-gastroduodenal digestive, neurological, skin, autoimmune, growth and miscellaneous disorders. Although the precise role for H. pylori is unknown in these diseases, it is of tremendous interest to most clinicians and scientists as H. pylori is amenable to eradication therapy using simple and reliable drug regimens. The conditions associated with H. pylori seropositivity are highlighted in this concise article. PMID- 10535465 TI - Helicobacter pylori in Africa: observations on an 'enigma within an enigma'. AB - BACKGROUND: We conducted a retrospective literature review of all the data published on Helicobacter pylori in Africa in order to test whether the prevalence of diseases associated with this organism differs from that in developed nations. METHODS: Both sero-epidemiological (n = 8) as well as prospective endoscopic studies in subjects with either dyspepsia or epigastric pain (n = 23) and one retrospective study were available for analysis. RESULTS: Sero-epidemiology confirmed both the early age of acquisition in children (50% by 10 years) as well as the high prevalence of the organism (61%) in adult asymptomatic individuals. Endoscopic studies in dyspeptic individuals revealed the presence of the organism in 72%. Duodenal ulceration was noted in 26% of 3473 cases and in these, H. pylori was present in 90%. An association of gastric metaplasia with duodenal ulceration was identified in the one study in which it was investigated. Gastric ulceration occurred approximately four-fold less frequently (7% of 2286 cases) than duodenal ulceration and the organism was evident in 75% of the gastric ulceration cases. Findings of intestinal metaplasia (14%) and gastric cancer (3.4%) were not infrequent, but the paucity of accurate epidemiological data made it difficult to establish a correlation between the two. CONCLUSION: It would appear that prospective endoscopic-based studies in African subjects may question the standard dogma of a low prevalence of H. pylori associated diseases in Africa. Further research is clearly required. PMID- 10535466 TI - Effect of lafutidine, a novel histamine H2-receptor antagonist, on monochloramine induced gastric lesions in rats: role of capsaicin-sensitive sensory neurons. AB - BACKGROUND: Lafutidine ((+/-)-2-(furfurylsulfinyl)-N-(4-(4-(piperidinomethyl)-2 pyr idyl)oxy-(Z)-2-butenyl)acetamide) is a novel histamine H2-receptor antagonist and has been shown to exhibit a potent gastroprotective activity in addition to its antisecretory action. In the present study, we examined the effects of lafutidine on the mucosal ulcerogenic and potential difference (PD) responses induced by monochloramine (NH2Cl) in rat stomachs. METHODS: Oral administration of NH2Cl at 120 mmol/L produced haemorrhagic lesions in the stomach in unanaesthetized rats. RESULTS: Lafutidine (3-30mg/kg), given p.o., showed a dose dependent and significant inhibition against damage caused by NH2Cl: the effect was significant at 10 mg/kg or greater but disappeared almost totally in the sensory deafferented animals following capsaicin pretreatment. Likewise, capsaicin (10 mg/kg, p.o.), but not cimetidine (100 mg/kg, p.o.) exhibited a potent protection against NH2Cl-induced gastric lesions. Topical application of NH2Cl (10 mmol/L) reduced transmucosal PD in ex-vivo stomachs of anaesthetized rats, but this PD response was also prevented by pre-exposure to lafutidine, in a dose-dependent and sensory neuron-sensitive manner. Mucosal exposure to NH4OH (60 mmol/L) also caused PD reduction in ex-vivo stomachs made ischaemic by bleeding from the carotid artery (1 mL/100 g bodyweight), resulting in severe gastric lesions. These ulcerogenic and PD responses caused by NH4OH plus ischaemia were attenuated by prior application of lafutidine as well as taurine, a scavenger of NH2Cl. The former effect was, again, dependent on the sensory neurons. Intraluminal capsaicin but not cimetidine was also effective in preventing a PD response to NH2Cl. CONCLUSIONS: These results suggest that lafutidine, but not cimetidine, protects the stomach against NH2Cl, whether occurring endogenously or administered exogenously and that this action may be mediated by capsaicin sensitive sensory neurons. PMID- 10535467 TI - Relationship of hydrolase activities to epithelial cell turnover in distal colonic mucosa of normal rats. AB - BACKGROUND: The relationships between changes induced by diet in colonic epithelial kinetics and in the activities of brush border hydrolases are poorly defined. The aims of this study are to define these relationships, as changes in kinetics would be expected to influence differentiation, and to determine whether the type of ingested dietary indigestible carbohydrates influences hydrolase activities. METHODS: Groups of eight rats were fed a low fibre diet +/- supplements of different types of indigestible carbohydrates for 4 weeks. Alkaline phosphatase (ALP) and dipeptidyl peptidase IV (DPPIV) activities and epithelial kinetics were measured in distal colonic mucosa. RESULTS: Median ALP activities correlated positively and DPPIV activity negatively with the median proportion of cells entering metaphase (r = 0.58 and -0.58, respectively; P < 0.05) and number of metaphase arrests per crypt column across the diets (r = 0.59 and 0.58, respectively; P < 0.05). Stepwise regression analysis showed that both hydrolases independently predicted these kinetic indices (R2 > 63% for each). Mucosal ALP activities were markedly elevated during consumption of raw potato starch, guar gum and methylcellulose, while only potato starch caused a significant elevation of DPPIV activities. CONCLUSIONS: The type of indigestible carbohydrate in the diet influences colonic mucosal hydrolase activities. The opposite relationship between kinetics and each of the two hydrolases indicates that these hydrolases do not reflect the same event; dipeptidyl peptidase IV might relate to differentiation status while ALP could also be influenced by epithelial irritation due to changes in luminal conditions. PMID- 10535468 TI - Effect of butyrate on paracellular permeability in rat distal colonic mucosa ex vivo. AB - BACKGROUND AND AIMS: The effects of butyrate on colonic epithelial barrier function are poorly understood. The aim of this study was to examine the short term effects of butyrate on paracellular permeability of rat distal colonic epithelium. METHODS: Mucosa mounted in Ussing chambers was treated with butyrate (1-10 mmol/L) for 4 h. Transepithelial conductance, [51Cr]-EDTA flux, mucosal brush border hydrolase activity and epithelial kinetics, using proliferating cell nuclear antigen (PCNA) staining, were measured. RESULTS: On exposure to butyrate (10 mmol/L, but not 1 or 5 mmol/L), transepithelial conductance was 65 +/- 2% higher (mean +/- SEM; n = 8, P < 0.05, paired t-test) and the rate coefficient for [51Cr]-EDTA flux was 65 +/- 25% higher (P = 0.03) than those of control tissue. Histologically, the epithelium exhibited no signs of injury, but butyrate treated tissue exhibited interstitial oedema consistent with water uptake in association with butyrate absorption. Butyrate caused a reduction in crypt column height to 30.6 +/- 1.6 cells from 33.4 +/- 1.8 cells in controls (n = 10, P < 0.03), but the number of cells per crypt column staining with PCNA was unchanged. Butyrate significantly reduced the mucosal activities of alkaline phosphatase by 40 +/- 16%, maltase by 54 +/- 12% and dipeptidyl peptidase IV by 41 +/- 14%. CONCLUSIONS: Acute exposure to butyrate increased paracellular permeability in rat distal colon. The mechanism involved may relate to the loss of differentiated surface epithelial cells, or as a physiological response to Na+-coupled butyrate uptake. PMID- 10535470 TI - Case report: Colonic duplication: a rare cause of obstruction. AB - Complete duplication of the entire large bowel with partial ileal involvement is very rare and diagnosis can often be difficult as illustrated by this case report. We also review the other clinical associations of this rare condition and briefly discuss the embryology of duplications of the gastrointestinal tract. PMID- 10535469 TI - Butyrate from bacterial fermentation of germinated barley foodstuff preserves intestinal barrier function in experimental colitis in the rat model. AB - BACKGROUND AND AIMS: The consumption of germinated barley foodstuff (GBF) prevents inflammation and diarrhoea in a colitis model. In this study we investigated the mechanism of the preventative effect of GBF on experimental colitis in rats, in view of production of bacterial butyrate and preservation of intestinal barrier function. METHODS: Sprague-Dawley rats administered with diets supplemented with 3.5% dextran sodium sulphate were used as an experimental colitis model. Butyrate was given to rats orally or intracaecally. Intestinal barrier function was estimated by light microscopic observation of the mucosa, intestinal permeability and bacterial translocation. RESULTS: Mucosal damage was reduced by intracaecal administration of butyrate, but not by oral administration. Bacterial butyrate production and reduction of mucosal damage depended on the dose of GBF in diets. The action of endogenous bacterial butyrate, including the reduction of intestinal permeability and bacterial translocation, was inhibited by administration of an inhibitor of beta-oxidation of short-chain fatty acids. CONCLUSIONS: The feeding of GBF promotes bacterial butyrate production and improves intestinal barrier function in rats, resulting in mitigation of experimental colitis. PMID- 10535472 TI - Transfusion-transmitted virus infection in China: prevalence in blood donors and in patients with liver diseases. AB - BACKGROUND: Prevalence of transfusion-transmitted virus (TTV) infection among blood donors and in patients with liver diseases in China was studied. METHODS: DNA was extracted from serum and amplified by seminested polymerase chain reaction with reported primer sets from a conserved region of the TTV genome. RESULTS: TT Virus DNA was detected in 55 of 196 blood donors (28%); 31% (40 of 127) in the north and 22% (15 of 69) in the south. TT Virus DNA was also detected in 14 of 31 patients (45%) with non-A-non-G fulminant hepatitis and in eight of 25 patients (32%) with non-A-non-G chronic hepatitis. The rate of TTV viraemia in these patients with liver disease was comparable to that in blood donors. TT Virus DNA sequencing of 12 isolates showed that the prevalence of genotype 2 was significantly higher than that reported in Japan (66.7 vs 2.6%, P < 0.001). Furthermore, genotyping assays based on restriction fragment length polymorphism were carried out on all 88 TTV DNA-positive samples. It was found that 42 isolates (47.7%) belonged to genotype 1 and 40 (45.5%) to genotype 2. It was of particular interest that the prevalence of genotype 1 in patients with non-A-non G fulminant hepatitis was significantly higher than that in blood donors (10/14 vs 22/55, P < 0.05). CONCLUSIONS: The data indicate that TTV infection is common in China and that the pathogenic potential of TTV toward the liver (if any) may differ between genotypes. PMID- 10535471 TI - Alpha-interferon therapy in chronic hepatitis due to active dual infection with hepatitis B and C viruses. AB - BACKGROUND: Nearly 14% of non-alcoholic chronic liver disease in India is related to hepatitis B virus (HBV) and hepatitis C virus (HCV) dual infection. There are no clear data available from the literature on the therapeutic management of these patients who often suffer an unfavourable course. METHODS: Fourteen consecutive cases of biopsy-proven chronic liver disease fulfilling the following criteria were included: Child's A or B liver disease, hepatitis B surface antigen positive, HBV-DNA positive, antibody to HCV positive and HCV-RNA positive. Seven patients had chronic liver disease (group I), while the remaining seven patients (group II) had additional disorders (non-Hodgkin's lymphoma (two), acute leukaemia (two), thalassaemia (two), chronic renal failure (one). Interferon alpha-2b (IFN) was given in a dose of 6 MIU thrice weekly for 6 months. Complete response was defined as loss of HBV-DNA and HCV-RNA at 6 months and sustained response (SR) as the sustained loss of HBV-DNA and HCV-RNA for more than 6 months during the follow-up period. RESULTS: At the end of 6 months, alanine aminotransferase (ALT) levels remained unchanged (120 +/- 40 vs 136 +/- 64 IU/L), but six of the seven (86%) patients in group I lost HBV-DNA. All three hepatitis Be antigen (HBeAg)-positive patients lost HBeAg with an early flare of ALT (at 45 +/- 12 therapy days). Two of these patients (29%) lost HCV-RNA. Thus, SR was seen in 29%, while HBV-DNA loss was found in 100% during the follow-up period. In group II patients, there was a significant decrease in ALT (308 +/- 14 vs 65 +/- 25 IU/L, P < 0.001), but only three (43%) patients lost HBV-DNA and two (29%) lost HCV-RNA. One patient with acute leukaemia and another with renal failure had a complete response to IFN, but none of the lymphoma patients showed any antiviral response. CONCLUSIONS: In chronic hepatitis due to dual infection with HBV and HCV, interferon therapy is: (i) safe; (ii) effective (more so in clearing HBV); (iii) often associated with early ALT flare; and (iv) may be less effective if non-Hodgkin's lymphoma is present. PMID- 10535473 TI - Duplex Doppler sonography in patients with Budd-Chiari syndrome. AB - BACKGROUND: Angiography has been the mainstay for diagnosis of Budd-Chiari syndrome even though other modalities are increasingly being used. We have evaluated our findings of duplex Doppler sonography (DDS) in patients with Budd Chiari syndrome. METHODS: Duplex Doppler sonography was performed in 37 consecutive angiographically proven patients with Budd-Chiari syndrome. RESULTS: Real time ultrasonography showed abnormalities of right, middle and left hepatic veins (HV) in 21, 15 and 18 patients, respectively. Duplex Doppler sonography showed abnormal flow patterns in 37, 22 and 31 patients in the right, middle and left HV, respectively, thereby increasing the diagnostic yield by 40%. An abnormal waveform in one or more HV was present in all 37 patients. Uniphasic flow was the commonest abnormality and was seen in 22, nine and 14 patients, respectively, in the right, middle and left HV, while there was no flow in five, four and seven patients in the right, middle and left HV, respectively. Intrahepatic collaterals were seen in 35 of 37 patients (94.6%). Hepatopetal flow was found in the portal vein of 21 of 23 patients (91.3%), while flow was hepatofugal in one and portal vein thrombosis was found in another. CONCLUSION: Duplex Doppler sonography is a useful procedure which helps in the diagnosis of patients with Budd-Chiari syndrome. PMID- 10535474 TI - Haemodynamic changes in non-alcoholic (viral) liver cirrhosis studied by computed tomography (CT) arterial portography and CT arteriography. AB - AIMS: To evaluate haemodynamic and vascular changes in non-alcoholic (viral) cirrhosis on conventional computed tomography (CT), CT arteriography (CTA) and CT arterial portography (CTAP), and to determine the cause of the observed reticular stain on angiography. METHODS: Using surgically resected liver specimens from 31 patients with viral hepatitis associated hepatocellular carcinoma, images of conventional CT, CTA, CTAP and the sinusoidal phase of hepatic arteriography were retrospectively analysed and compared with pathology of the non-cancerous portion of the liver. RESULTS: Computed tomography arteriography showed inhomogeneous enhancement (diffuse, low-density nodules) in a total of 16 samples (52%); in eight of 10 (80%) cirrhotic livers, three of six (50%) precirrhotic livers, five of 12 (42%) livers with chronic active hepatitis and none of three with no active liver disease. The frequency of inhomogeneous enhancement became significantly higher with increasing severity of parenchymal damage (P < 0.05). In contrast, conventional CT and CTAP showed homogeneous enhancement in all 31 (100%) patients. There was no correlation between inhomogeneous enhancement on CTA and reticular staining on sinusoidal-phase hepatic angiograms. Inhomogeneous enhancement was frequently seen in patients with hepatitis B surface antigen and/or anti-hepatitis C virus antibody compared with those without them (P < 0.05). CONCLUSION: The CTA was much more sensitive in detecting haemodynamic changes in the cirrhotic liver than CTAP, conventional CT and sinusoidal-phased hepatic angiography. Further study is required to clarify the mechanism of inhomogeneous enhancement on CTA and homogeneous enhancement on CTAP seen in cirrhosis. PMID- 10535475 TI - Parenchymal echo patterns of cirrhotic liver analysed with a neural network for risk of hepatocellular carcinoma. AB - BACKGROUND: To objectively evaluate the parenchymal echo patterns of the liver in cirrhosis, an image analysing system in which a neural network is used has been found capable of numerically calculating coarse score (CS). Using this system, we analysed whether or not CS can serve as a predictive factor for the development of hepatocellular carcinoma (HCC). METHODS: The risk factors for HCC were evaluated in 95 patients with liver cirrhosis with an average follow-up period of 2041 +/- 823 days. We used a three-layer feed-forward neural network and a back propagation algorithm to calculate CS. RESULTS: There were strong correlations between CS, alanine aminotransferase (ALT) and alpha-fetoprotein (AFP) and the average cumulative incidence rate of HCC evaluated by the Cox's proportional hazards model. The adjusted rate ratios were estimated to be 3.00, 2.80 and 2.01, respectively. The cumulative risks of HCC were significantly higher with an initial CS > or = 1.5 than with an initial CS < 1.5, with ALT > or = 80 IU/L than with initial ALT < 80 IU/L and with AFP > or = 20 ng/mL than with initial AFP < 20 ng/mL, all analysed by the log-rank test. CONCLUSIONS: Coarse score is a useful predictor for development of HCC. PMID- 10535477 TI - Case report: Occurrence of hepatocellular carcinoma 4.5 years after successful treatment with virus clearance for chronic hepatitis C. AB - A 67-year-old man with hepatitis C virus infection and histological features of chronic active hepatitis was treated with human recombinant interferon alpha-2b. He responded successfully to interferon with normalization of serum alanine aminotransferase and continuous eradication of hepatitis C virus. However, 4.5 years after the end of interferon therapy, hepatocellular carcinoma, 20 mm in diameter, was detected by routine ultrasonographic screening. The patient underwent surgical treatment for resection of the hepatocellular carcinoma. The non-cancerous liver tissue was histologically normal. This suggests that patients who have shown a complete response to interferon therapy for chronic hepatitis C should be followed up closely. PMID- 10535476 TI - Effective hepatic artery chemoembolization for advanced hepatocellular carcinoma with extensive tumour thrombus through the hepatic vein. AB - BACKGROUND AND AIMS: Advanced hepatocellular carcinoma (HCC) with extensive tumour growth through the hepatic vein still has an extremely poor prognosis, even after cancer chemotherapy and/or transarterial embolization. Although aggressive surgical treatments using extracorporeal circulation and liver transplantation have been performed by some authors, the reported results were still unsatisfactory. In this study, we report the favourable result of hepatic artery chemoembolization and subsequent surgical resection in three patients with advanced HCC with extensive tumour thrombus through the hepatic vein. METHODS AND RESULTS: Three irresectable patients with HCC with extensive tumour thrombus through the hepatic vein underwent hepatic artery chemoembolization with aclarubicin, mitomycin C, lipiodol and/or Gelfoam. After the reduction of tumour extent with hepatic artery chemoembolization, two of the three patients underwent surgical resection. These two patients are still alive at 59 and 21 postoperative months, respectively. In the other case, the extent of the tumour and functional reserve of the liver prevented us from performing surgical resection, but the patient is doing well 62 months after the initial treatment. CONCLUSIONS: Hepatic artery chemoembolization with aclarubicin, mitomycin C, lipiodol and/or Gelfoam might be an effective treatment for irresectable advanced HCC with extensive tumour thrombus into the inferior vena cava or the right atrium through the hepatic vein. Radical surgical resection might be applicable for selected patients without high surgical risk after reducing tumour extent by hepatic artery chemoembolization. PMID- 10535478 TI - Case report: Percutaneous drainage of the pancreatic head hydatid cyst with obstructive jaundice. AB - We report a rare case of a patient with a primary hydatid cyst in the head of the pancreas who presented with obstructive jaundice caused by extrinsic compression of the intrapancreatic portion of the bile duct. The patient was treated successfully by ultrasound-guided percutaneous drainage of the cyst using hypertonic (20%) saline as the scolicidal agent and albendazole chemoprophylaxis before and after the drainage. The cyst was not visible on ultrasonography at 6 months follow up. Clinical, sonographic and serological follow up to 35 months showed no evidence of cyst recurrence or dissemination. In endemic areas of hydatid disease, hydatid cyst should be a differential diagnosis in cystic lesions of the pancreas in patients presenting with obstructive jaundice. PMID- 10535479 TI - Hepatobiliary and pancreatic: a case of severe epigastric pain. PMID- 10535480 TI - Gastrointestinal: multiple lymphomatous polyposis. PMID- 10535481 TI - Helicobacter pylori infection correlates with severity of reflux esophagitis: with manometry findings. AB - The role of Helicobacter pylori infection in the development and exacerbation of reflux esophagitis was investigated. The prevalence of Helicobacter pylori infection, the severity of atrophic gastritis, and esophageal motility (determined by esophageal manometry by an infusion catheter method) were assessed in patients with mild (n = 46) and severe (n = 27) reflux esophagitis and subjects without reflux (n = 28). Compared with the prevalence of Helicobacter pylori infection in the non-reflux group, the prevalence in the mild and severe reflux groups (60.7%, 47.8%, and 14.8%, respectively) was significantly (P < 0.05) lower. Atrophic gastritis was milder in both reflux groups than in the non reflux group. The degree of gastritis was also milder in the severe reflux group than in the mild reflux group. The esophageal sphincter pressure was significantly (P < 0.05) lower in the reflux groups than in the non-reflux group, and the amplitude of primary peristalsis was significantly (P < 0.05) lower in the severe reflux group than in the non-reflux group. There were no significant differences between reflux patients with and without Helicobacter pylori infection in the parameters of esophageal manometry. These data imply that a low prevalence of Helicobacter pylori infection may result in a milder grade of atrophic gastritis, and consequently, exacerbate reflux esophagitis. PMID- 10535482 TI - The vast majority of gastric T cells are polarized to produce T helper 1 type cytokines upon antigenic stimulation despite the absence of Helicobacter pylori infection. AB - Helicobacter pylori infection is associated with chronic infiltration by various cell types, including T cells, whose cytokine production may regulate the inflammatory reaction as well as local immune response to the bacterium. We prospectively analyzed the constituents of the cellular infiltrates and the cytokines produced by T cells in antral biopsies obtained from 73 subjects with and without H. pylori infection, before and after eradication therapy, and compared them with a histological grade of gastritis. We found that T cells predominated in cell number, followed by granulocytes/monocytes and plasma cells in both H. pylori-infected and H. pylori-uninfected subjects. Despite the absence of H. pylori infection, more than 70% of gastric CD4-positive T cells obtained from uninfected tissue produced interferon-gamma (IFN-gamma) in the cytosol. Upon receptor cross-linking of a CD3 and a CD28 molecule, T cells in both infected and uninfected tissue continuously secreted a far greater amount of IFN-gamma than those in peripheral blood mononuclear cell controls for a period of cell culture, whereas the increase in interleukin-4 (IL-4) was very small, and no increase in IL-2 secretion was seen. In H. pylori-infected patients, IFN-gamma secretion was correlated with the grade of mononuclear cell infiltration and decreased to an uninfected control level after eradication therapy. We did not see the effect of eradication on IL-4 secretion. Anti-H. pylori antibody of the IgG2 subclass was remarkably increased in H. pylori-infected subjects. These results together suggest that gastric T cells are already differentiated to produce a large amount of IFN-gamma by a mechanism unrelated to H. pylori infection. H. pylori infection appeared to activate T cells to secrete even more IFN-gamma, which may contribute to maintaining a perpetual inflammation in H. pylori-infected stomach. PMID- 10535483 TI - Antibacterial action of bile acids against Helicobacter pylori and changes in its ultrastructural morphology: effect of unconjugated dihydroxy bile acid. AB - Although it has been shown that bile acids possess antibacterial activity against Helicobacter pylori, few reports on their activity have been published. We determined the minimum inhibitory concentration at 72 h of various unconjugated and conjugated bile acids against laboratory standard strains and clinical isolates of H. pylori, and studied morphologic changes of H. pylori under the scanning electron microscope during treatment with deoxycholic acid and ursodeoxycholic acid at the minimum inhibitory concentrations for 24 h. We found that only the unconjugated form of dihydroxy bile acid has antibacterial activity. The minimum inhibitory concentration of deoxycholic acid is 200-400 microg/ml, that of chenodeoxycholic acid is similar to that of deoxycholic acid, and that of ursodeoxycholic acid is 400-800 microg/ml. The morphology of H. pylori changed from its primary rodlike shape to a spherical shape with blebs on the cell surface, and was further degraded to an irregularly condensed mass, following an increase in bile acid. This morphologic change in H. pylori was different from the change to a spherical shape caused by amoxicillin. On the basis of these results, it seems that unconjugated dihydroxy bile acid might be a candidate drug for eradication of H. pylori, but further investigations of the clinical usefulness of bile acid for this purpose should be done. PMID- 10535484 TI - A new endoscopic metallic stenting method for duodenal stenosis: a preliminary report. AB - Palliative duodenal stenting was attempted in three patients with severe duodenal stenosis due to tumor invasion. Two methods were applied for duodenal stenting: the conventional method, which inserts the Ultraflex (stent for esophageal stenosis) along the guidewire under fluoroscopy, and a new method that uses a snare and an endoscope to guide the esophageal stent. The conventional method is often unsuccessful, because the delivery tube is too short, but the latter method appears to be a safe and effective duodenal stenting technique. PMID- 10535485 TI - Expression of multidrug resistance protein in metastatic colorectal carcinomas. AB - To clarify the relationship between multidrug resistance protein (MRP) and clinicopathologic features, the influence of adjuvant chemotherapy, and prognosis of patients who underwent resection of metastatic liver carcinomas originating from colorectal carcinomas, we examined the expression of MRP in tumor tissues by immunostaining. Specimens of 38 primary colorectal tumors and 44 metastatic liver tumors of colorectal origin were examined (metastatic group). We also examined 28 nonmetastatic colorectal carcinomas. The percentages of nonmetastatic tumors and of primary and metastatic tumors of the metastasis group that expressed MRP were similar. MRP expression in primary and metastatic tumors did not correlate with any clinicopathologic features. The use of adjuvant chemotherapy after operation for primary colorectal carcinomas was associated with increased MRP expression among metastatic liver tumors. Expression of MRP in the tumor did not influence the prognosis or survival rate after resection of primary or metastatic tumors. Our data suggest that MRP expression in metachronous liver metastases from colorectal carcinomas may be induced by administration of anticancer drugs but is not associated with clinicopathologic features of the tumor, liver metastasis, or prognosis. PMID- 10535486 TI - Prevalence of TT virus in patients with fulminant hepatic failure in Japan. AB - A novel virus (TT virus) was isolated from patients with posttransfusion hepatitis of unknown etiology. We studied the prevalence of TT virus in 26 patients with fulminant hepatic failure without risk factors, including blood transfusion, and also examined 106 healthy blood donors as controls. We assayed serum TT virus DNA by seminested polymerase chain reactions and also examined the genotypes of this virus. Serum was obtained at admission from patients with fulminant hepatic failure. Serum samples at admission from seven (27%) of the 26 patients were positive for TT virus DNA. There were no differences in clinical findings, duration from onset to coma, or results of laboratory tests in patients with and without TT virus DNA. However, all 7 patients with TT virus died, whereas 9 of the 19 patients without TT virus died. The outcome for patients with fulminant hepatic failure and TT virus was significantly worse than for patients without the virus (P = 0.0227). TT virus was also detected in 29 (27%) of the 106 healthy blood donors. The genotype of the TT virus was mainly 1a in both groups. There were no differences in the rate of positivity and the genotypes of TT virus between patients with fulminant hepatic failure and healthy blood donors. TT virus infection may not cause severe hepatitis, such as fulminant hepatic failure, but it may indicate a poor outcome in such patients. PMID- 10535487 TI - Clinical and virological features of chronic hepatitis C carriers with normal alanine aminotransferase levels despite positive serum HCV-RNA after interferon therapy. AB - Interferon (IFN) is the only drug that induces viral clearance, but in patients with chronic hepatitis C, HCV-RNA clearance is achieved in only 20%-40% of patients treated with IFN for 6 months. The remaining patients have positive serum HCV-RNA, but about 5%-15% of the patients with positive serum HCV-RNA after IFN therapy showed normal alanine animotransferase (ALT) levels (incomplete response; ICR). In these patients, IFN therapy is thought to be related to the suppression of necroinflammatory reaction in the liver. The aim of this study was to evaluate the demographic, clinical, histological, and virological characteristics of patients with an ICR. In this study, ICR was defined as normalization of serum ALT, but positive HCV-RNA by reverse-transcription (RT) nested PCR at two points, 3 and 6 months after cessation of IFN therapy. By multiple logistic regression analysis, the risk ratio for ICR appearance in patients treated with IFN for more than 12 months was 3.06 compared with patients treated with IFN for less than 12 months. In patients with ICR after IFN therapy, serum ALT was often reelevated during the follow-up period. The incidence of ALT re-elevation was about 10% per year in the patients treated with IFN for less than 12 months, while the incidence of ALT reelevation in the patients treated with IFN for 12 months and more was significantly lower (P = 0.0176) by the log rank test. PMID- 10535488 TI - Detection of pre-S/S gene mutants in chronic hepatitis B carriers with concurrent hepatitis B surface antibody and hepatitis B surface antigen. AB - Hepatitis B virus (HBV) with various mutations has been reported. The frequency of the natural occurrence of such variants and whether the heterogeneity of these genomic regions correlates with a specific serologic pattern of concurrent hepatitis B surface antigen (HBsAg) and hepatitis B surface antibody (anti-HBs) were investigated. We analyzed the perS/S regions of HBV in six asymptomatic HBV carriers who were seropositive for both HBsAg and anti-HBs (group A), four hepatocellular carcinoma (HCC) patients with concurrent HBsAg and anti-HBs (group B), and five asymptomatic HBV carriers without anti-HBs as controls (group C). PreS/S regions of HBV-DNA were amplified by polymerase chain reaction (PCR), cloned, and sequenced. The results showed that, in some of the samples, a few deletions and numerous point mutations were presented in preS/S regions. One of the HBV carriers with anti-HBs (group A) and an HCC patient with anti-HBs (group B) had point mutations in the "a" determinant, resulting in conversion from Ile 126 of wild-type to Asn-126. The patients with anti-HBs (groups A and B) had a significantly greater divergence rate of amino acid for the preS/S gene compared with controls. Our results suggested that the HBV mutants observed in the preS/S gene may have led to changes in the immunogenicity of the viral particles, and thus influence the viral behavior and clinical course. Therefore, some HBV patients with concurrent HBsAg and anti-HBs may be HBV S mutants. PMID- 10535489 TI - Detection of IgA, IgM, and IgG subclasses of anti-M2 antibody by immunoblotting in autoimmune cholangitis: is autoimmune cholangitis an early stage of primary biliary cirrhosis? AB - Autoimmune cholangitis (AIC) has been proposed as a distinct disease entity from primary biliary cirrhosis (PBC), without antimitochondrial antibody (AMA) and anti-M2 antibody but with a high titer of antinuclear antibody (ANA) in the serum. However, negativity for AMA and anti-M2 antibody was determined by different methods in different studies. We hypothesized that anti-M2 antibody negativity in AIC resulted from methodological differences, including selection of the immunoglobulin subclass of the autoantibody. Twenty-three patients compatible with AIC whose serum tested negative for AMA and positive for ANA (> or = 1:80) were compared with 71 AMA-positive PBC patients. Laboratory findings, histology, and the pattern of anti-M2 antibody assessed by immunoblotting were compared. Alkaline phosphatase, total bilirubin, total cholesterol, and IgM values were lower in patients with AIC (P < 0.05, 0.01, respectively). Anti smooth muscle antibody was detected more frequently in patients with AIC (P < 0.01). However, anti-M2 antibody was detected using immunoblotting not only in PBC but also in AIC cases. IgA class alone, IgM class alone, or both IgA and IgM classes of anti-M2 antibody were detected in 13%, 17%, and 22% of AIC patients, respectively, whereas they were not detected in PBC patients (P < 0.05, P < 0.01, P < 0.01). IgG class anti-M2 was detected in all patients with PBC, whereas it was detected in 48% of patients with AIC (P < 0.01). Histological evaluation showed that the early stages of disease were found more frequently in AIC (78%) than in PBC patients (39%) (P < 0.01). Anti-M2 antibody was detected by immunoblotting in all AIC patients. Hence, AIC is not a distinct disease from PBC. For diagnosing AIC and/or PBC, anti-M2 antibody should be examined by the immunoblotting assay to detect not only IgG but also IgA and IgM subclasses. PMID- 10535490 TI - Portal venous blood flow while breath-holding after inspiration or expiration and during normal respiration in controls and cirrhotics. AB - In this study, we used magnetic resonance (MR) imaging to measure portal blood flow in 12 healthy controls and 17 cirrhotics while they were breath-holding after inspiration and after expiration. We then compared the results with measurements made during normal respiration in the healthy controls and cirrhotics. Blood flow in the main portal vein under basal fasting conditions was quantitated using the cine phase-contrast MR velocity mapping method. Three measurements were made on one occasion, as follows: (1) throughout the cardiac cycle during normal respiration, (2) with the subject breath-holding after maximal inspiration, and (3) with the subject breath-holding after maximal expiration. During normal respiration, portal blood flow was 1.3 +/- 0.21/min in controls vs 1.0 +/- 0.11/min in cirrhotics (P < 0.0001); while subjects were breath-holding after inspiration, portal blood flow was 1.0 +/- 0.21/min in controls vs 0.9 +/- 0.11/min in cirrhotics; and while subjects were breath holding after expiration, portal blood flow was 1.5 +/- 0.21/min in controls vs 1.1 +/- 0.21/min in cirrhotics (P < 0.0001). The differences were primarily due to changes in flow velocity. When the magnitude of these hemodynamic changes in the three respiratory conditions was compared in controls and cirrhotics, analysis of variance (ANOVA) showed a significant difference (P < 0.0001). In controls, portal blood flow decreased during maximal inspiration relative to flow during normal respiration (-24.6 +/- 8.3%). Changes in portal blood flow in controls were greater than in cirrhotics (-13.5 +/- 4.5%) (P < 0.0001); however, the difference in blood flow increase associated with maximal expiration between the two groups (+11.8 +/- 9.4% vs +5.9 +/- 11.5%) was not significant. We found that the respiration-induced hemodynamic variation in portal blood flow was less in cirrhotics than in the healthy controls. Portal blood flow measurements made during normal respiration using MR imaging closely reflect nearly physiologic conditions. PMID- 10535491 TI - Hepatocyte growth factor effects on motility of stone-diseased and stone-free human gallbladders. AB - Hepatocyte growth factor (HGF) has unique morphogenic activity for several cell types. Besides its major effect upon liver regeneration, its motogenic activity to enhance motility has not been verified for smooth muscles. Therefore we evaluated the impact of HGF in an in-vitro model of human gallbladder motility. Twelve stone-diseased and eight stone-free muscle strips were preincubated with HGF (100 ng/ml, 200 ng/ml). For the analysis of motility, cholecystokinin (CCK) was added (0.1 nM, 0.5 nM, 2 nM, 10 nM, and 100 nM). Twelve stone-diseased and eight stone-free strips without HGF incubation served as the control group. The tone of healthy (tone/100 nM CCK: control group, 12.4 +/- 3.6 mN; HGF group, 19.5 +/- 4.5 mN) and stone-diseased (tone/100 nM CCK: control group, 10.8 +/- 3.8 mN; HGF group, 17.3 +/- 4.8 mN) muscle strips, preincubated with HGF, was increased, with a higher sensitivity to CCK. Our results suggest that there is a clear motogenic response of stone-diseased human gallbladders to HGF. PMID- 10535492 TI - Mucosal carcinoma within a colonic diverticulum. AB - We report a case of a mucosal carcinoma and adenoma within a diverticulum in the cecum. Radiographic, endoscopic, and pathologic evaluation of the tumor is presented. Surgical resection was undertaken because of the size and shape of the lesion, risk of perforation, and the possibility of malignancy. A recent review of the literature with respect to clinical signs, diagnosis, growth of the carcinoma, and treatment of tumors around or within diverticula is also presented. A carcinoma or adenoma arising within the diverticulum is very rare. Endoscopic resection of the tumor could entail the risk of perforation, because of the lack of muscular coats in the diverticula. Surgical treatment may be the procedure of choice for lesions near or within the diverticula. PMID- 10535493 TI - Sigmoid adhesion caused by a congenital mesocolic band. AB - Congenital mesocolic band is an uncommon aberration in the development of the mesentery. Large-bowel obstruction secondary to adhesion and/or congenital band is very rare in children. A 6-month-old male infant who had no history of previous surgery was admitted with unremitting crying. A barium enema showed extraintestinal compression of the sigmoid colon. Laparotomy revealed an adhesive mesocolic band compressing the proximal part of the sigmoid colon. The band was lysed. The patient has remained asymptomatic since the procedure. PMID- 10535494 TI - Two cases of inflammatory bowel disease with multiple myeloma. AB - A significant increase has been reported in reticuloendothelial neoplasms in patients with inflammatory bowel diseases. We present two rare cases of multiple myeloma in patients with inflammatory bowel diseases. One was in a 58-year-old woman with ulcerative colitis, and the other was in a 59-year old woman with Crohn's disease. In both patients, multiple myeloma occurred during long-term observation of inflammatory bowel disease and during the inactive stage of intestinal inflammation. The multiple myeloma appeared to have resulted from monoclonal gammopathy of undertermined significance in both patients, and was diagnosed by characteristic serum and bone marrow findings. Our findings suggested that multiple myeloma should be particularly considered in women of middle or advanced age with ulcerative colitis or Crohn's colitis and serum monoclonal gammopathy. PMID- 10535495 TI - Aged Budd-Chiari syndrome attributed to chronic deep venous thrombosis with alcoholic liver cirrhosis. AB - Budd-Chiari syndrome is a rare disease, but there are many known causes. Recent studies showed that it can be an acquired lesion resulting from thrombosis in some elderly patients. We report a 74-year-old man with Budd-Chiari syndrome attributed to chronic deep venous thrombosis and alcoholic liver cirrhosis. When he was aged 45 years, stasis ulcers of the lower extremities appeared. Cerebral infarction and left hemiparesis occurred at age 71. Ultrasonography, venacavography, and three-dimensional-magnetic resonance imaging on admission demonstrated total obstruction of the inferior vena cava with several massive thrombi and developed collateral vessels. Although the etiology of the thrombosis remained obscure, we made some speculative assumptions that chronic disseminated intravascular coagulation (which is frequently observed in cirrhosis) or hereditary coagulopathy could be involved, from his familial history of thrombotic phenomena and a severe deficiency of clotting inhibitors. Despite the high mortality of untreated Budd-Chiari syndrome reported in previous studies, this patient had been alive for about 30 years from the suspected onset. PMID- 10535496 TI - Granulocyte-colony-stimulating-factor-producing hepatocellular carcinoma. AB - Patients with hepatocellular carcinoma (HCC) rarely show marked granulocytosis. We report an interesting case of rapidly growing poorly differentiated HCC associated with marked granulocytosis. The blood leukocyte count decreased following several treatments for HCC, including transcatheter arterial embolization, and increased again along with tumor regrowth. The serum levels of granulocyte colony-stimulating factor (G-CSF) were markedly elevated, and immunohistochemical study showed G-CSF staining in the cytoplasm of certain HCC cells. These findings confirm that HCC in this case was a G-CSF-producing tumor. PMID- 10535497 TI - Spontaneous rupture of a nonparasitic liver cyst complicated by intracystic hemorrhage. AB - a case of spontaneous rupture of simple liver cyst complicated by intracystic hemorrhage is described. This rare condition was detected in a 61-year-old man who underwent left trisegmentectomy of liver under a suspected diagnosis of cystadenocarcinoma because of elevated serum levels of carbohydrate antigen (CA) 19-9 and DUPAN 2, and the presence of an intracystic structure. The resected specimen showed a benign liver cyst with intracystic hematoma and high levels of CA19-9 and DUPAN 2 in the cystic fluid. It is suggested that cyst rupture may increase serum levels of tumor markers whose levels are high in the cystic fluid, and that repeated observations of an intracystic structure may be the most reliable method to distinguish intracystic hemorrhage from cystic neoplasm. PMID- 10535498 TI - How does Helicobacter pylori infection affect the pathophysiology of reflux esophagitis? PMID- 10535499 TI - Overshift towards T helper (Th)1 cells induces gastric mucosal injury: immunopathology of Helicobacter pylori-infected gastric mucosa. PMID- 10535500 TI - The interaction of bile acids and Helicobacter pylori. PMID- 10535501 TI - Incomplete response of interferon treatment has an important role for the prevention of hepatocellular carcinoma. PMID- 10535502 TI - The entity "autoimmune cholangitis": hanging by a thread? PMID- 10535503 TI - Genetic diversity of major piroplasm surface protein genes and their allelic variants of Theileria parasites in Thai cattle. AB - Twenty-eight field isolated Theileria parasite DNAs obtained from dairy and beef cattle in distinct geographical areas of Thailand were characterized by using polymerase chain reaction (PCR) amplification with six sets of oligonucleotide primers. Three sets of them were modified from two genes of immunodominant major piroplasm surface protein (MPSP) coding for 32 kDa (p32) of T. sergenti and 33/34 kDa (p33/34) of T. buffeli, and MPSP of Theileria spp.(Thai-isolate). The other three sets of primers were basically generated from three alleles of MPSP which were specific for Japanese T. sergenti-Ikeda stock (I-type), Japanese T. sergenti Chitose stock (C-type) and Australian T. buffeli-Warwick stock (B1-type), respectively. The results indicated that 14 out of 28 isolates were amplified by the Thai-specific primer whereas 6 isolates were amplified by the p32 specific primer and the other 5 isolates were amplified by the p32 and Thai-specific primers. In addition, by using the allele-specific PCR, 14 out of 28 isolates contained C-type MPSP whereas 3 isolates contained B1 type parasites. Interestingly, 20 out of 28 isolates could be amplified by the Thai-specific primer. The majority of Theileria parasites distributed in Thailand contained Thai type parasites, whereas C-type parasites showed the mixed population with B1 and Thai type parasites. No I type parasite was detected. PMID- 10535504 TI - Immunogenicity of alpha-toxin, capsular polysaccharide (CPS) and recombinant fibronectin-binding protein (r-FnBP) of Staphylococcus aureus in rabbit. AB - This study was conducted to evaluate the antibody levels of alpha-toxin, capsular polysaccharides (CPS) and fibronectin-binding protein (FnBP) in rabbits immunized with an experimental vaccine against Staphylococcus aureus and to develop the bovine mastitis subunit vaccine in the future. Enzyme immunoassay was used for detection of IgG antibodies against staphylococcal CPS, alpha-toxin and FnBP. The levels of specific antibodies against CPS, alpha-toxin and FnBP in immunized rabbits were significantly increased after first immunization compared with control animals (p<0.05). Of three antigen used in vaccine, immunogenicity of CPS was relatively lower, compared with those of alpha toxin and fibronectin binding protein. Numbers of S. aureus in blood of immunized groups were lower than those of control group after bacterial challenge. But the bacterial numbers among immunized groups were not significantly different. S. aureus counts in excised organs were significantly lower in all immunized rabbits than in PBS-control group (p<0.05). The present study showed that alpha-toxin, capsular polysaccharide and fibronectin binding protein included in a subunit vaccine were protective. PMID- 10535505 TI - Longevity, lifetime pig production and productivity, and age at first conception in a cohort of gilts observed over six years on commercial farms. AB - An observational cohort study was conducted using a producer group of 33 farms selected based on their completeness of reproduction data, including dates of birth, entry to a herd, and removal. Average lifetime pig production and parity at removal in a cohort of 2,265 females born in 1990 were 67.2 pigs born alive and 5.6 parities, respectively. Approximately 90% of farrowings occurred from the second through the fourth year from birth. Farrowing rates between parities of 2 and 4 were higher than other parities, and pigs born alive from parities 3 to 5 were the greatest among parities. The 10th and 90th percentiles of age at first conception were 227 and 322 days. Increasing the age at first mating was associated with low farrowing rate (P<0.01) in parity 0. Older age at first conception was associated with lower parity at removal, shorter reproductive herd life, and fewer lifetime pigs born alive (P< 0.01). Of the 2,265 breeding females, 253 (11.2%) were re-mated at parity 0 and farrowed. These sows with a record re-mating at parity 0 had lower parity at removal, less lifetime pig production and lower lifetime productivity than those with no re-mating at parity 0 (P<0.01). It is recommended that unbred gilts 230 days of age or older should be mated soon. PMID- 10535506 TI - CT examination of the guttural pouch (auditory tube diverticulum) in Przewalski's Horse (Equus przewalskii). AB - The domestic horse (Equus caballus) have the large symmetrical guttural pouches (the auditory tube diverticulum) formed by saccate bulge of the auditory tube. In this study, CT examination was carried out in the head of Przewalski's horse (Equus przewalskii), the only true wild horse living at present. As results of the examination, Przewalski's horse possessed the large symmetrical guttural pouches divided into medial and lateral compartments by the stylohyoid bone. Moreover, the right and left guttural pouches meet each other at the median part to form a thin septum. As CT sections get close to the part of the occipital condyle, the lateral compartment disappeared, and the medial compartment gradually became small toward the base of the skull. These results indicate that the nuchal-basal part of the medial compartment is not well-developed as compared with the domestic horse. PMID- 10535507 TI - Parasitological survey on wild carnivora in north-western Tohoku, Japan. AB - In the winter of 1997-1998, we collected parasitological data from 60 wild carnivora in the north-western part of Tohoku region, Japan. These included 7 foxes (Vulpes vulpes japonica), 20 raccoon dogs (Nyctereutes procyonoides viverrinus), 29 martens (Martes melampus melampus), 3 weasels (two Mustela sibirica itatsi and one M. nivalis namiyei), and one Japanese badger (Meles meles anakuma). Roundworms (Toxocara canis in foxes and Toxocara tanuki in raccoon dogs), hookworms (Ancylostoma kusimaense and Arthrostoma miyazakiense) and Molineus sp. in the small intestine were the most prevalent in foxes and raccoon dogs. In martens, Aonchotheca putorii in the stomach, Concinnum ten in the pancreatic duct, Molineus sp. and Euryhelmis costaricensis in the small intestine were the most prevalent. Collected parasites include some new helminth species for this region or Japan; the strobilar stage of Taenia polyacantha from foxes, Pygidliopsis summa from a raccoon dog, Eucoleus aerophilus, A. putorii, and Soholiphyme baturini from martens. PMID- 10535508 TI - Geographical variation of the skull morphology of the common tree shrew (Tupaia glis). AB - Geographical variation was examined morphologically in the common tree shrew (Tupaia glis) in some Indochinese and Malayan regions. Osteometrical examination and principal component analysis elucidated the morphological differences among various populations. The populations from southern and western Thailand were distinguished morphologically from the other populations. Variation in males from south Thailand and Kuala Lumpur suggests that the Isthmus of Kra may have an influence on the variation of skull morphology. However, the Isthmus of Kra was not completely considered as a factor of geographical separation in this species, because we could not confirm the separation in skull size and shape between the localities at least in females. While, the Kanchanaburi population in western Thailand was significantly smaller than the other population in skull size, and constituted the morphologically separable group in our study. PMID- 10535509 TI - Effects of volatile anesthetics on the activity of laryngeal 'drive' receptors in anesthetized dogs. AB - Effects of halothane, isoflurane and sevoflurane on laryngeal drive receptor activity were studied in the afferent activity of the superior laryngeal nerve in anesthetized spontaneously breathing dogs. Of 40 single units recorded, most of them (65%) responded to the volatile anesthetics applied to the isolated larynx at a concentration of 5%. The exposure to the anesthetics resulted in either an inspiratory increase (15%), both inspiratory and expiratory decrease (54%), or both inspiratory increase and expiratory decrease (31%) responses. The average discharge frequency of the receptors tended to be decreased on inhalation of the anesthetics, where significant decreases were observed in both respiratory phases for halothane and at expiration for isoflurane, but in neither respiratory phase for sevoflurane. These results support an advantage of sevoflurane over halothane and isoflurane for induction of anesthesia to minimize the influence of the activity of laryngeal drive receptors on the breathing pattern and airway stability. PMID- 10535510 TI - Biphasic change in correlation between ovarian lipid peroxides and progestational activity during pseudopregnancy induced in immature rats. AB - We measured ovarian lipid peroxide (LP) levels and plasma progestins, progesterone (P4) and 20alpha-dihydroprogesterone, throughout pseudopregnancy in gonadotropin-primed immature rats. Plasma P4 fluctuated, with two peaks on days 5 (PSP5) and 8 of pseudopregnancy, and then declined to the basal level by PSP12. Ovarian LP increased from PSP1 to PSP4, decreased temporarily until PSP8, and then rose gradually until PSP14. From PSP1 through PSP7, ovarian LP was positively correlated with total progestins according to the Spearman ranked correlation coefficient (r=+0.829, p<0.05). In contrast, a negative correlation between ovarian LP and plasma P4 was apparent (r=-0.816, p<0.05) from PSP8 to PSP14. These results show the biphasic correlation of LP with luteal progestational activity depending on the luteal stage. PMID- 10535511 TI - Analysis of the N-terminal polypeptide of the capsid precursor protein and the ORF3 product of feline calicivirus. AB - The N-terminal unique polypeptide region of the capsid precursor protein of feline calicivirus (FCV) and the protein encoded by ORF3 of FCV were expressed as fusion proteins with glutathione S-transferase to analyze the expressed products in FCV-infected cells. Immunoblot analysis using a serum from a cat experimentally infected with FCV indicated relatively high immunogenicity of the N-terminal polypeptide in FCV-infected cats, as compared with the ORF3 protein. Specific antisera were prepared by immunization to mice with the fused proteins and used in immunoblot analysis. A 14 kD product corresponding to the N-terminal polypeptide and a 10 kD polypeptide of the ORF3 product were identified in the FCV-infected cells but not detected in the purified particles. No neutralization activity against FCV was detected in these antisera. The proteins identified as polypeptides of 14 kD and 10 kD in this study may have functions as non structural proteins. PMID- 10535512 TI - Unique expression patterns of myosin heavy chain genes in the ductus arteriosus and uterus of rabbits. AB - In smooth muscle tissue, two smooth muscle myosin heavy chain (MHC) isoforms (SM1, SM2) and two non-muscle MHC isoforms (NMA, NMB) have been identified. The purpose of our study was to clarify whether smooth muscle MHC mRNA expression reflects the physiological and functional state of the muscle. We studied the expression pattern of MHC mRNAs, using the S1-nuclease mapping procedure, in functionally and morphologically changeable organs; the ductus arteriosus (DA) during development (25 and 29 days of gestation, and from 3-day-old neonates) and uteri from virgin, day-10 pregnant (P10) and day-29 pregnant (P29) rabbits. The results demonstrated that SM2 expression was greater in the fetal DA than in the fetal aortic and pulmonary arteries, but that it decreased significantly following closure of DA. In the gravid uterus, SM1 expression was significantly (P<0.05) strong compared to other MHC mRNAs from virgin to P10 rabbits. During pregnancy, NMB expression showed a tendency to increase until P10, and after P10, SM2 expression increased dramatically and NMB expression decreased to give almost a mirror image of the SM2 expression. Smooth muscle type (SM1, SM2) was significantly (P<0.05) strong compared to non-muscle type expression (NMA, NMB) at P29. These data suggest that smooth muscle MHC mRNA, especially SM2 expression reflects the physiological and functional state of the smooth muscle. PMID- 10535513 TI - A case of a dog with thickened calvaria with neurologic symptoms: magnetic resonance imaging (MRI) findings. AB - A 6-year-old female mongrel dog weighing 9.0 kg was presented ananastatic, with clouding of consciousness, bilateral loss of hearing and depressed reactivity of the eyes to light. Magnetic resonance imaging (MRI) examination showed that the calvaria was markedly thickened with compression to the cerebrum and cerebellum. The case of a dog with thickened calvaria with compression of the cerebrum and cerebellum which could not be diagnosed by conventional measures was amenable to diagnosis by MRI. With increased application of MRI examination, such canine cases might increase in number. PMID- 10535515 TI - Electron microscopical observations of psittacine beak and feather disease in an Umbrella cockatoo (Cacatua alba). AB - Psittacine beak and feather disease (PBFD) was diagnosed in an umbrella cockatoo (Cacatua alba) with severe feather dystrophy and loss. Electron microscopically, the intranuclear and intracytoplasmic inclusion bodies observed by light microscopy were composed of viral particles forming paracrystalline arrays, whorls, semicircles or concentric circles. Recovered viral particles from the skin and feather follicle tracts were icosahedral and 15 to 20 nm in diameter. PMID- 10535514 TI - Measurement for breath concentration of hydrogen and methane in horses. AB - This study concerns the establishment of a simple testing method for breath concentration of hydrogen and methane in horses. Twenty-eight healthy thoroughbreds and 24 Arabians were used. Breath samples were collected using one minute closed circulatory respiration through an aluminum bag filled with 10 liters of pure oxygen, which was mounted on the subjects by means of a face mask. Breath samples obtained, were analyzed by gas chromatography. A significant correlation in both hydrogen and methane levels was observed for samples collected at separate times. These findings confirmed the usefulness of our approach for testing breath concentrations of hydrogen and methane in horses. PMID- 10535516 TI - Balloon dilation of right ventricular outflow tract in a dog with tetralogy of Fallot. AB - Balloon dilation was performed on a dog with tetralogy of Fallot. Immediately following balloon dilation, the peak systolic pressure gradient across the pulmonic valve declined from 97 to 63 mmHg. Doppler echocardiography following balloon dilation revealed increased pulmonary blood flow. Clinical symptoms obviously improved and the dog's improved condition was maintained for 4 months. There were no serious complications in performing the procedure. It was concluded that balloon dilation was a safe and effective treatment for a dog case with tetralogy of Fallot. Long-term follow-up studies will be required to identify the exact indications of balloon dilation for tetralogy of Fallot. PMID- 10535518 TI - Serotyping and pathogenicity of Erysipelothrix strains isolated from tonsils of slaughter pigs in Thailand. AB - Erysipelothrix strains were isolated from the tonsils of 46 (15.0%) of 307 apparently healthy slaughter pigs in Thailand during the period of August to September, 1997. A total of 27 of the 46 Erysipelothrix isolates could be classified into 5 serovars but the remaining 19 were untypable in this study. Of the 25 isolates serologically identified as Erysipelothrix rhusiopathiae, 20, 4, and 1 isolates belonged to serovars 2, 12, and 17, respectively. Only 2 isolates from the tonsils belonged to Erysipelothrix tonsillarum and represented either serovar 7 or 10. Although the periods and the districts of the survey were limited, the information obtained in the present investigation demonstrates the presence of a variety of serovars in pigs in Thailand. Of 29 selected isolates belonging to serovars 2, 7, 10, 12, 17, and untypable, only 5 (17.2%) were virulent for both mice and pigs. Five of these virulent isolates belonging to serovars 2 and 12 killed less than 30% of mice immunized with a swine erysipelas bacterin commercially available in Thailand, suggesting that the vaccine elicited a sufficient immunity to these field isolates. PMID- 10535517 TI - Feline coronavirus participation in diarrhea of cats. AB - Fecal samples were examined for viruses participated in gastrointestinal disorders of cats, especially focusing on feline coronavirus (FCoV) by a reverse transcriptase-polymerase chain reaction assay. It was found that a primary viral pathogen was feline panleukopenia parvovirus (FPLV; 28.5% of the positive rate) and the secondary was FCoV (10.7%). Commonly reported clinical signs of cats of which feces were FCoV-positive were vomiting, diarrhea and dehydration with an exception of one serious case with concurrent FPLV infection. PMID- 10535519 TI - Characterization of methicillin-resistant Staphylococcus aureus isolated from dogs in Korea. AB - Twelve strains of the methicillin-resistant Staphylococcus aureus (MRSA) recovered from hospitalized dogs were analyzed for in vitro antimicrobial susceptibility and virulence, and were genetically characterized by pulsed-field gel electrophoresis (PFGE). Antibiotic susceptibility test showed that nearly all isolates were resistant to beta-lactam antibiotics tested and all the strains were fully susceptible to glycopeptides. There were no inhibitory activities among the aminoglycosides. The 50% lethal dose (LD50) was determined by intraperitoneal injection of cell suspensions and estimated by the Spearman Karber method. The mouse lethality of MRSA and methicillin-susceptible S. aureus (MSSA) was not significantly different in both normal and cyclophosphamide treated mice (p>0.05), indicating that they were equally virulent. There was a great difference in the incidence of toxin production between the MRSA and MSSA group; 83.3% (10 of 12) of the MRSA and 14.3% (1 of 7) of the MSSA were toxin producers. The predominant types produced by MRSA was B. All the MRSA strains were capsular type 5 producers, while of 7 MSSA strains, four were type 5, one for type 8, and two were nontypeable. Based on the PFGE analysis, the 12 MRSA isolates generated 9 to 11 fragments in the size range of <48.5 to 630.5 kb, and yielded 6 different patterns. The results indicated that production of toxin and capsule type do not play a role in the pathogenicity to mouse and PFGE is a valuable tool for the characterization of MRSA. This report is the first such cases in the veterinary literature in Korea and may indicate the frequent emergence of MRSA in veterinary clinic hereafter. PMID- 10535520 TI - Selenium-dependent glutathione peroxidase modules encoded by RNA viruses. AB - Glutathione peroxidase (GPx) is the prototypical eukaryotic selenoprotein, with the rare amino acid selenocysteine (Sec) at the enzyme active site, encoded by the UGA codon in RNA. A DNA virus, Molluscum contagiosum, has now been shown to encode a functional selenium-dependent GPx enzyme. Using modifications of conventional sequence database searching techniques to locate potential viral GPx modules, combined with structurally guided comparative sequence analysis, we provide compelling evidence that Se-dependent GPx modules are encoded in a number of RNA viruses, including potentially serious human pathogens like HIV-1 and hepatitis C virus, coxsackievirus B3, HIV-2, and measles virus. Analysis of the sequences of multiple viral isolates reveals conservation of the putative GPx related features, at least within viral subtypes or genotypes, supporting the hypothesis that these are functional GPx modules. PMID- 10535521 TI - Study of the incorporation of selenium into peroxidase isozyme of wheat seedling. AB - The effects of selenium on the activity of peroxidase (POD) of wheat seedling and its isozyme pattern were studied using a greenhouse hydroponic experiment. The results show that the activity of POD is increased in response to higher Se concentration (approx 5.0 mg/L) in culture medium. The electrophoretic pattern of the POD isozyme was altered by growth of the wheat in a selenium medium. Se could incorporate into some POD isozymes during either seed germination or the seedling growth period. There is a dose-responsive incorporation of selenium in isozyme of POD and selenium content in the isozyme increase along with the increase of selenium concentration in the medium. PMID- 10535522 TI - Low environmental selenium availability as an additional determinant for goiter in East Java, Indonesia? AB - Iodine deficiency, which is most visibly indicated by goiter, is highly prevalent in Indonesia. Since 1994, Indonesia has a decree that all salt used for human, livestock, and industry must be iodized. However, despite the increased distribution of iodized salt, pockets with significantly higher prevalence of goiter still remain. This situation may be consequence of selenium (Se) deficiency. This study aimed to assess the Se level in the environment of goiter prevalent areas. Five hundred eleven school children participated in this study. Goiter was measured using both ultrasound and palpation. Ninety-nine eggs were collected from free-living chicken in 11 villages, and the Se contents of egg yolk and egg white were determined by neutron activation analysis. In the villages studied, Se concentration in egg yolk ranged from 0.15 to 1.52 microg/g and in egg white from 0.18 to 2.97 microg/g. The prevalence of goiter measured by palpation ranged from 18.4% to 70% and by ultrasound from 0% to 100%. Because of the inconsistency of goiter rate measured by palpation and ultrasonography, the question remains whether low availability of Se in the environment might be an additional contributing factor for goiter. PMID- 10535523 TI - Age-related changes of element contents in human mitral and tricuspid valves. AB - To examine age-related changes of human cardiac valves, mitral and tricuspid valves were analyzed by inductively coupled plasma-atomic emission spectrometry. The subjects for mitral valves consisted of 12 men and 8 women, ranging in age from 52 to 96 yr. The subjects for tricuspid valves consisted of 11 men and 6 women, ranging in age from 52 to 93 yr. Furthermore, 16 of the samples of the cardiac valves were derived from the same subjects. The contents of calcium, phosphorus, and magnesium in the mitral valves increased progressively with advancing age and reached maximum in the 80s in regard to calcium and phosphorus and maximum in the 90s in regard to magnesium. The maximum average amounts corresponded to about three times the average contents in the 60s. In contrast, the content of sulfur in the mitral valves remained constant between the 50s and 90s. Regarding tricuspid valve, the contents of calcium, phosphorus, and magnesium scarcely increased with advancing age, except for one subject who died of chronic renal failure. Histological observations of the mitral valves revealed that deposits of calcium were present in both the elastic fibers and its degenerative tissues of the mitral valve. The present study indicates that the accumulation of calcium, phosphorus, and magnesium occurs progressively in the mitral valve with aging, but does not occur in the tricuspid valve. PMID- 10535524 TI - Regulation of mitochondrial cytochrome b mRNA by copper in cultured human hepatoma cells and rat liver. AB - Copper overload and deficiency are known to cause morphological and functional mitochondrial abnormalities. The reverse transcriptase-polymerase chain reaction (RT-PCR)-based method of differential display of mRNA was used to identify genes with altered expression in cultured human hepatoma cells (Hep G2) exposed to increasing concentrations of copper (0-100 microM, 24 h). Copper regulation of a cloned PCR product, identified as the gene for the mitochondrially encoded cytochrome b, was confirmed by Northern analysis and in situ hybridization. Copper toxicity increased cytochrome b mRNA abundance up to 3.6-fold, and copper chelation reduced it by 50%. Hepatic cytochrome b mRNA was also increased in rats fed a high-copper diet. Thapsigargin treatment resulted in a significant increase in cytochrome b mRNA, suggesting that an increase in intracellular calcium may be involved in the mechanism of copper action. Furthermore, although cyclohexamide (CHX) alone did not increase cytochrome b mRNA, the addition of CHX and copper resulted in a sixfold increase. These data suggest a role for cytochrome b in the response to increases or decreases in hepatic copper. PMID- 10535526 TI - Early effects of surgery on zinc and metallothionein levels in female rats. AB - Time-response effects of experimental surgery on zinc (Zn) and metallothionein (MT) homeostasis were investigated in female rats up to 24 h. Hepatic Zn content increased at 20 and 24 h postsurgery, whereas serum Zn levels decreased. Hepatic MT increased significantly by 9 h postsurgery and peaked at up to twofold of control at 12 h after surgery. Following the peak at 12 h, hepatic MT content decreased with time but did not reach control levels at the end of this study. When MT isoforms were evaluated, MT-II levels were elevated to the highest extent by 12 h after surgery, whereas MT-I levels started to decrease after 3 h postsurgery but then increased by 20 h. The early increases in MT content are probably mediated by nonmetallic mediators released during the postsurgical inflammatory process, favoring the plasma/tissue mobilization of Zn. This process might be part of the overall mechanisms occurring in the inflammation. PMID- 10535525 TI - Urinary zinc excretion and zinc status of patients with beta-thalassemia major. AB - In this study, zinc status and urinary zinc excretion with and without desferrioxamine (DFO) infusion and the relationship between urinary zinc excretion and renal tubular dysfunction in thalassemia major (TM) patients were investigated. Forty TM patients were given four DFO infusions on alternate days over a 1-wk period prior to the transfusion. On each day that DFO was given, a 24 h urine collection initiated. DFO was omitted for 1-wk before the following transfusion and during the period four 24-h urine collections were performed. Twenty healthy children provided 24-h urine collection as controls. Blood samples were taken on each of two consecutive transfusion days of the patients and from the controls. Urinary zinc excretion was measured and plasma and red blood cell (RBC) zinc analysis were performed by inductively coupled plasma-atomic emission spectrophotometry. Urinary N-acetyl-beta-D-glucosaminidase (NAG) activity and creatinine were determined in morning urine specimens. The mean plasma zinc concentration was significantly lower in the patients not given DFO compared to the values of the patients given DFO and the control group. The mean RBC zinc concentration (micromol/g Hb) in the patients (with and without DFO) and the control group were similar. Urinary zinc excretion was significantly higher in the patients receiving DFO compared to the control group, whereas urinary zinc excretion in the patients not given DFO was not different from the controls. Urinary NAG indices (U/g Cr) were significantly higher in the patients compared to controls. Urinary zinc excretion was correlated with the urinary NAG indices. PMID- 10535527 TI - Blood chromium determination in assessing reference values in an unexposed Mediterranean population. AB - Plasma chromium levels were determined in 243 healthy subjects. The study group consisted of 134 men and 109 women, ages 19-71 yr, all residing in Barcelona in northeastern Spain. The study was designed to assess the reference levels for plasma chromium and to investigate its relationships to age and sex. The assays were performed by means of a graphite-furnace atomic absorption spectrometer. The mean plasma chromium concentration was 3.01 +/- 1.45 nmol/L, ranging from 0.6 to 6 nmol/L. The upper reference values in the 0.95 percentile for this population was 5 nmol/L. No significant differences were observed with respect to the subjects' sex. PMID- 10535528 TI - Long-term results of the use of silicone sheets after diskectomy in the temporomandibular joint: clinical, radiographic and histopathologic findings. AB - The aim of the present study was to evaluate the long-term results of a group of patients who had the disk of the temporomandibular joint (TMJ) removed and permanently replaced by a silicone sheet. The study group comprised 48 patients, treated in the period from 1983 to 1993. In eight patients, the implants had to be removed after an average interval of 5.6 years and they were submitted for histopathological examination. Twenty-five of the 40 patients with silastic implants in place, and five of the 8 patients who had their implants removed, were available for long-term follow-up (mean interval of 7.0 years, SD 2.8 years). Clinical function was rated according to the Helkimo Dysfunction Index and compared to the preoperative findings. Results showed decreased tenderness of muscles and joints to palpation and increased mouth opening, but no statistically significant improvement in joint function. In 4 patients, a decrease in condylar width was found, while another 4 patients presented with thickening of the condyle by appositional bone formation. Histopathology of the failed implants showed scattered fragments of silastic material and dacron fibers with accumulation of histiocytes in immediate contact with the silicone particles and phagocytozed intracellular material. T-lymphocytes were also present in the vicinity of the silicone particles. PMID- 10535529 TI - Full-thickness skin graft interposition after temporomandibular joint ankylosis surgery. A study of 31 cases. AB - Recurrence is a major problem after release of temporomandibular joint ankylosis. Early physiotherapy and choice of interpositional material are important in preventing recurrence. Currently, the most used technique is gap arthroplasty associated with coronoidectomy, temporalis muscle flap interposition and reconstruction of the condylar unit with a costochondral graft. Full-thickness skin graft interposition, using the technique described by Popescu & Vasiliu, can also be used. This retrospective review of 31 patients confirms the reliability of full-thickness skin graft interposition. Results were successful in 90% of the 20 patients with follow-up longer than one year. PMID- 10535530 TI - Eagle's syndrome: lesser cornu amputation: an alternative surgical solution? AB - A case is reported of a 42-year-old female patient, who presented with clinical symptoms of Eagle's syndrome, radiographic evidence of marginally elongated styloid processes as well as markedly elongated lesser cornua of the hyoid. The symptoms were successfully treated by amputating the lesser cornua of the hyoid. The patient has now been asymptomatic for more than 6 years. PMID- 10535531 TI - Cysts of the temporomandibular joint. Report of two cases. AB - Two patients with a cyst of the TMJ are presented. One had a synovial cyst, the other a ganglion cyst. The differential diagnosis of these lesions is discussed. PMID- 10535532 TI - Soft tissue management using palatal mucosa around endosteal implants in vascularized composite grafts in the mandible. AB - Excessive thickness and mobility of the skin flap and lack of a vestibular sulcus in composite vascularized bone grafts for mandibular reconstruction limit the usefulness of endosteal implants to support a prosthetic device. Palatal mucosal grafts and a simple acrylic stent are an excellent means to overcome these problems. The technique is presented in this paper. PMID- 10535533 TI - A modified submental approach for oral endotracheal intubation. AB - Hernandez, in 1986, published the first paper on "The submental route for endotracheal intubation". The technique was developed to avoid tracheostomy, particularly in maxillofacial trauma cases, where short-term intermaxillary fixation was required. Gordon & Tolstunov published 2 cases utilizing the technique. Our initial experience with the technique as presented was less than satisfactory. The technique was modified to utilize a strict midline approach. Fifteen cases utilizing this modified technique are reported, including 14 cases of craniomaxillofacial trauma and one complex orthognathic surgery case. There have been no operative or postoperative complications. Postoperative submental scarring has been acceptable. PMID- 10535534 TI - Distraction of scarred soft tissue before secondary bone grafting. A case report. AB - Mandibular distraction was performed to restore oral function in a 52-year-old man with tongue cancer, in whom a mandibular fracture developed after marginal resection of the mandible. The fracture caused the mandibular dental arch to be shorter than the maxillary arch. An external fixation device was attached to the collapsed mandible. The mandibular soft tissue was expanded by 32 mm. After gradual distraction, a vascularized iliac bone graft was transferred to the lengthened space. Subsequently, vestibuloplasty was performed and implants were inserted. A normal appearance, acceptable occlusion and satisfactory oral function were achieved. PMID- 10535535 TI - Monitoring of graft perfusion and osteoblast activity in revascularised fibula segments using [18F]-positron emission tomography. AB - The aim of the present study was to evaluate healing of revascularised fibula grafts used for mandibular reconstruction using [18F]fluoride ion and positron emission tomography (PET). Sixteen PET studies in 11 fibula grafts were analysed to determine both blood flow and fluoride influx as a measure of vascularisation and osteogenic activity. Two graft failures and three non-unions were encountered and were compared to the successfully healed grafts. In uneventful graft healing, early PET studies (on average 19 days after grafting) showed a significantly increased blood flow to the grafted bone and to the union between the grafts and the mandibles when compared to the reference region of the cervical spine. In contrast, fluoride influx was significantly lower in the grafts when compared to the plating area and the cervical spine. Six months after grafting, blood flow to the grafted bone and the mandibular bone had returned to a level comparable with the reference region. Fluoride influx remained significantly lower in the grafts than in the plating areas or cervical spines. Graft failures were associated with negligible fluoride influx near zero in early PET studies. These results suggest that revascularised fibula grafts provide a low osteogenic potential, presumably due to the pre-existing lack of cancellous bone. The relatively high frequency of non-unions makes meticulous adaptation of the graft and the mandible mandatory, particularly in patients with compromised viability of the recipient bone. PMID- 10535536 TI - Versatility of vascularized fibula and soft tissue graft in the reconstruction of the mandibulofacial region. AB - The anatomical region "laterodorsal calf" with the fibula, the flexor muscles and the overlying skin served by the peroneal vessels, is a suitable donor region particularly for combined microsurgical transfer of bone and soft tissue. In a five and a half year period, 59 reconstructions, 39 (66%) with and 20 (34%) without soft tissue transfer, were performed. The versatility and the functional and esthetic results after osseous, osteocutaneous, osteomuscular and osteomyocutaneous tissue transfer are presented. PMID- 10535537 TI - Adenoid cystic carcinoma arising in the pterygopalatine fossa presenting with visual deficit. A case report. AB - An adenoid cystic carcinoma, arising in the pterygopalatine fossa and presenting with a visual deficit as the initial symptom in a 44-year-old woman, is reported. Magnetic resonance imaging was valuable in diagnosing the primary site and extension of the tumor. PMID- 10535538 TI - Closure of an oropharyngocutaneous fistula in an irradiated patient. A case report. AB - A case of oropharyngocutaneous fistula is presented in a preoperatively irradiated patient. A double-layer closure, using a modified Owens flap, was used to obtain a satisfactory result. PMID- 10535539 TI - Etiology of fibrous dysplasia and McCune-Albright syndrome. AB - In this paper, the etiology of monostotic fibrous dysplasia and McCune-Albright syndrome is explained. Both monostotic fibrous dysplasia and McCune-Albright syndrome are sporadically occurring disorders in which a mutation in the GNAS1 gene occurs postzygotically in a somatic cell. All cells descended from the mutated cell can manifest features of McCune-Albright syndrome or fibrous dysplasia. Cells descended from non-mutated cells develop into normal tissues. Thus, the clinical pattern is variable in distribution and appearance. More generalized vs more localized expression depend on (a) how small or how large the cell mass is during embryogenesis when the mutation occurs and (b) where in the cell mass the mutation occurs. Topics discussed include G proteins and their receptors, cycling of stimulatory G protein between active and inactive forms, and specific mutations in GNAS1. We also discuss four possibilities: (a) Are there masked mutations? (b) Are there effects of imprinting? (c) Are there non classical mutations? and (d) Is fibrous dysplasia a neoplasm? PMID- 10535540 TI - Treatment of central giant cell granuloma of the jaw with calcitonin. AB - Giant cell granuloma of the jaw is a benign lesion that may cause local destruction of bone and displacement of teeth. The common therapy is curettage or resection, which may be associated with loss of teeth and, in younger patients, loss of dental germs. An alternative treatment has recently been introduced, in which patients receive a daily dose of calcitonin. Four patients who have been treated with calcitonin in various concentrations for at least 1 year are reported. In all patients, complete remission of the giant cell granuloma was observed, without signs of recurrence. The working mechanism of calcitonin is discussed, as are length of treatment and optimal dose. PMID- 10535541 TI - The platysma myocutaneous visor flap for intraoral reconstruction. A case report. AB - The use of a modified myocutaneous platysma flap is presented for a patient with a large ameloblastoma of the mandible. The possible advantages and limitations of the technique are discussed. PMID- 10535542 TI - Osteochondroma of the mandibular condyle. A case report. AB - A 52-year-old woman presented with pain on the right temporal region, restricted mandibular movement and a gradually developing malocclusion. Osteochondroma of the mandibular condyle was suspected based on diagnostic imaging. Local resection with preservation of the condylar head resulted in complete resolution of symptoms. PMID- 10535543 TI - Granular cell tumor of the parotid gland. A case report. AB - Granular cell tumor (GCT), or granular cell myoblastoma, is a relatively uncommon lesion of the soft tissues. The tumor is thought to derive from a Schwann cell or from a perineural undifferentiated mesenchymal cell. GCT can occur in any organ, but the parotid gland is very unusual. A case of GCT of the parotid gland in a 30 year-old woman is described. PMID- 10535544 TI - Facial pressure sore complicated by mandibular osteomyelitis. AB - A case is reported of an 85-year-old woman with mandibular osteomyelitis secondary to a submental pressure sore. The main aetiological factors in the development of the pressure sore were dementia and severe senile kyphosis. Pressure sores are rare in the head and neck region and, though osteomyelitis is a common complication at other anatomical sites, it has not been previously reported in the mandible. PMID- 10535545 TI - Microvascular anastomosis in oral reconstructive surgery: a comparative study of recipient vessels between Japanese and German patients. AB - Specimens of recipient vessels were obtained from the oral and maxillofacial regions when carrying out microsurgery and the condition of the vascular wall was analyzed histopathologically. This study was performed in both Japan and Germany and the results were compared. A higher percentage and a more advanced abnormality of the recipient vessel seemed to exist in the German patients as compared to the Japanese patients. This difference could be due to the preoperative multimodal chemo- and radiotherapy more often applied in Germany and the higher incidences of systemic diseases such as hypertension and diabetes in the German population. The success rate of the surgery is, however, the same in both countries. It could be concluded that the viability of the flaps was not affected by preoperative damage to the recipient vessel. PMID- 10535546 TI - Short-term effect of deep-frozen stored auricular cartilage allograft repair on the osteoarthritic sheep temporomandibular joint. AB - The initial effect of deep-frozen stored auricular cartilage replacement was tested on the progression of experimentally induced osteoarthritis in sheep. Bilateral osteoarthritis was induced in the sheep temporomandibular joint. Three months later, discectomy and deep-frozen auricular cartilage allograft repair were performed unilaterally, with the other side being left as a control. Six months post induction of osteoarthritis, 3 months post repair, the sheep were killed. The stored auricular cartilage allografts were not firmly attached to the surrounding tissue and tended to be markedly thinned and perforated. The effect of storage detrimentally affected the properties of the graft. PMID- 10535547 TI - A simple and versatile device to aid patient positioning during surgery. Technical note. AB - A simple technique to aid patient positioning during surgery is described. It has proved to be effective, versatile and cheap. PMID- 10535548 TI - Congenital submental salivary fistula. PMID- 10535549 TI - The importance of doing complete literature searches. PMID- 10535550 TI - Achilles tendon injuries: conservative versus surgical treatment. PMID- 10535551 TI - An addendum to the Hippocratic oath? PMID- 10535552 TI - Two-stage exchange is better than direct exchange in the infected THA. PMID- 10535553 TI - Careful patient selection is the key for direct exchange in the infected THA. PMID- 10535554 TI - Video-assisted thoracic diskectomy and anterior release: a biomechanical analysis of an endoscopic technique. AB - This study evaluates the residual biomechanical stability of the spine following multilevel anterior diskectomies and anterior longitudinal ligament release using video-assisted thoracoscopic surgery (VATS). Eighteen domestic pigs were randomly divided into three groups of six pigs. Group 1 underwent thoracic anterior release from T4-T9 using a left-sided VATS approach, group 2 underwent thoracic anterior release from T4-T9 via a traditional left thoracotomy (open), and group 3 did not undergo surgery and served as a control. After surgery, the animals were euthanized, and the thoracic spinal columns were harvested for biomechanical testing. Nondestructive testing was performed on all specimens in pure compression, flexion, extension, right lateral bending, and torsion. Specimens from group 1 had significantly lower stiffness values (P<.05) than the control group for all five test modes. These data demonstrate that adequate anterior release of the thoracic spine can be obtained with the VATS technique. Further prospective clinical studies on VATS are required before the widespread application of this technique. PMID- 10535555 TI - Spiral computed tomography with two- and three-dimensional reconstruction in the management of tibial plateau fractures. AB - Spiral computed tomography (CT) with three-dimensional and multiplanar reconstructions was used in the evaluation of tibial fractures in nine patients. Computed tomography added important information to that obtained by plain radiographs. Five (55%) fractures were reclassified. The degree of articular depression was often underappreciated on plain radiographs. Furthermore, the fracture complexity and the spatial relation of fragments could be readily demonstrated with 3-D reconstruction. This technique is useful in planning operative reconstruction. PMID- 10535556 TI - Anatomic considerations for anterior instrumentation of the lumbar spine. AB - One hundred seventy lumbar vertebrae from L1-L4 were used to quantitatively evaluate the lumbar vertebral body and examine the relationship of the maximum posterior angles of screw placement to the spinal canal. Anatomic evaluation included dimensions of the vertebral body. Three entrance points on the lateral aspect of the vertebral body for screw insertion and an additional point 3 mm from the posterolateral corner of the spinal canal were defined and marked. The maximum posterior screw angles were determined as the angles between the line connecting the entrance point with the additional point and the coronal plane. Results showed that vertebral body dimensions increased from L1-L4. The average vertebral body depth, width, and height were approximately 26 mm, 36 mm, and 22 mm at L1, and 30 mm, 44 mm, and 23 mm at L4, respectively. The spinal canal may be penetrated if the screws are directed posteriorly 2 degrees-5 degrees at L1 - L2 and 9 degrees - 14 degrees at L3-L4 starting at the junction between the pedicle and vertebral body, 22 degrees - 32 degrees at L1-L4 from the level of 10 mm anterior to the junction, and 43 degrees -50 degrees from the level of 20 mm anterior to the junction. Therefore, mid-body screws should be directed perpendicular to the lateral plane of the vertebral body. For a more anteriorly placed screw, slightly posterior angulation is recommended. PMID- 10535557 TI - Reduced acetabular depth in hip instability in the newborn. AB - This study assessed the frequency of ultrasonographic hip instability and compared the results with clinical and morphological ultrasonographic evaluations of the hip in the newborn infant. Hip stability in 2016 consecutive births (1074 boys and 942 girls) was assessed clinically and ultrasonographically. Ultrasonography included dynamic stability testing and morphologic evaluations. Compared with ultrasonography, clinical stability testing had a specificity of 0.998 and a sensitivity of 0.667. The incidence of instability was 0.007 for hips and 0.010 for babies. Acetabular depth in 3994 normal hips was 8.3 +/- 0.9 mm, femoral head diameter was 15.4 +/- 1.3 mm, and acetabular bony coverage of the femoral head was 54 +/- 4.3%. Hip instability was significantly associated with a reduced acetabular depth. As the femoral head diameter was within normal in the unstable hips, coverage was significantly reduced. PMID- 10535558 TI - Biomechanical and surgical solutions for patellar implant in total knee arthroplasty. PMID- 10535559 TI - Fractures of the glenoid cavity: assessment and management. PMID- 10535560 TI - Musculoskeletal trauma: high- and low-energy injuries. PMID- 10535561 TI - Periosteal osteosarcoma. PMID- 10535563 TI - Entrapment of the extensor pollicis longus after intra-articular Smith's fracture. PMID- 10535562 TI - Transarticular spread of Ewing's sarcoma across the sacroiliac joint: CT and MRI correlation. PMID- 10535564 TI - Nonunion of a medial clavicular fracture following radical neck dissection: MRI diagnosis. PMID- 10535565 TI - Spinal epidural hematoma following thrombolytic therapy for acute myocardial infarction. PMID- 10535566 TI - Radiologic case study. Traumatic myositis ossificans. PMID- 10535567 TI - Cystic sebaceous tumors as marker lesions for the Muir-Torre syndrome: a histopathologic and molecular genetic study. AB - Cystic sebaceous tumors (CST) are well-circumscribed, large, deeply located dermal sebaceous proliferations with a cystic growth pattern. We identified 12 CST in 8 of 19 patients with Muir-Torre syndrome (MTS). We interpret CST as a tumor spectrum with clearly benign cystic sebaceous adenomas at one end and proliferative atypical cystic sebaceous tumors at the other. When examining these proliferative atypical tumors on morphologic criteria alone, the possibility of an evolving cystic sebaceous carcinoma cannot be excluded. We have not observed recurrences or metastases, indicating that these lesions are not highly malignant carcinomas. In 10 of 12 cases of CST, we examined microsatellite instability (MSI). All 10 examined examples of CST from patients with MTS showed MSI characteristic for hereditary nonpolyposis colorectal cancer (HNPCC), which is caused by autosomal dominant inherited DNA mismatch repair (MMR) defects. Mutational analysis of the MMR genes hMSH2 and hMLH1 had revealed different germline mutations in the hMSH2 gene in three of six examined patients with MTS with CST. We then found four more CST in patients without a history of internal malignancy. All four CST exhibited MSI. By mutational analysis in one of these patients we identified a truncating germline mutation in the MMR gene hMLH1. We conclude that CST is a marker for the mismatch repair-deficient subtype of MTS with a high risk for later internal malignancies. By recognizing CST, the histopathologist can suggest the great likelihood of MTS to the clinician. PMID- 10535569 TI - p53, mdm-2, and p21 waf-1 in the porokeratoses. AB - The etiology of the porokeratoses is unknown. Overexpression of the p53 tumor suppressor protein and disregulated cell cycle control have been pathogenically implicated. The p53 tumor suppressor gene product is regulated by mdm2 and both gene products influence cell cycle progression through the cyclin-dependent kinase inhibitor p21. Thirty-three cases of the various types of porokeratosis were immunohistochemically studied for p53, mdm2, and p21 proteins. Each of the cases showed increased p53 and decreased mdm2 and p21 expression within keratinocytes underlying cornoid lamella. This study confirms the previous findings of increased p53 staining and expands the potential roles of mdm2 and p21 in the pathogenesis of the porokeratoses. PMID- 10535568 TI - Differential expression of CD34 and Ki-M1p in pleomorphic fibroma and dermatofibroma with monster cells. AB - Pleomorphic fibroma (PF) and dermatofibroma with monster cells (DFMC) are characterized by the presence of numerous cells with large atypical nuclei. Despite cytologic similarities, the two entities are likely to be unrelated, but their histogenesis is poorly understood. In this study, we examined six cases of PF and eleven cases of DFMC by immunohistochemistry using antibodies against vimentin, alpha-smooth muscle actin, S-100 protein, CD34, factor XIIIa, and the pan-monocytic marker Ki-M1p. Strong vimentin expression was seen in all tumors, whereas none of them expressed S-100 protein. PF consistently exhibited CD34 staining but appeared to be depleted of Ki-M1p positive cells compared with the surrounding normal skin. Conversely, all cases of DFMC contained numerous Ki-M1p positive cells including atypical multinucleate cells, but virtually no CD34 reactivity was observed. A weak staining for alpha-smooth muscle actin was occasionally seen in a subset of the cells of both entities. Our results indicate that PF and DFMC are histogenetically distinct entities that may arise from two different types of dermal dendritic cells defined by their reactivity for CD34 and Ki-M1p, respectively. Immunohistochemistry using these two antibodies permits an easy and reliable discrimination between PF and DFMC. PMID- 10535570 TI - Immunohistochemical staining for androgen receptors: a sensitive marker of sebaceous differentiation. AB - Androgen receptors (AR) are present in normal skin being localized to the basal and differentiating cells of the sebaceous gland, and as such, sebaceous glands are androgen sensitive tissue. Androgen receptor expression was examined in 43 sebaceous neoplasms including 8 sebaceous carcinomas, 22 sebaceous adenomas, 12 specimens showing sebaceous hyperplasia, and 1 sebaceous epithelioma, as well as in 14 squamous cell carcinomas, 2 clear cell acanthomas, and 35 basal cell carcinomas. Epithelial membrane antigen (EMA) expression was also examined in all of the sebaceous neoplasms. All specimens were fixed in formalin and embedded in paraffin. Diffuse positive nuclear androgen receptor antibody immunohistochemical staining was observed in all samples of sebaceous neoplasms, whereas approximately 60% of basal cell carcinomas showed only focal positivity for nuclear androgen receptor immunoreactivity. Clear cell acanthomas and squamous cell carcinomas were uniformly negative. Whereas all sebaceous neoplasms exhibited immunoreactivity for androgen receptors, the staining pattern was more marked in the nuclei of seboblasts and differentiating sebocytes in the adenomatous, hyperplastic, and epitheliomatous lesions than in the nuclei of the less differentiated sebaceous carcinoma cells. All the sebaceous neoplasms except for sebaceous carcinomas exhibited immunoreactivity for EMA. In the sebaceous carcinomas, EMA staining was absent in the most poorly differentiated specimen, but with increasing differentiation, the carcinomas became immunoreactive to EMA. We have shown that the nuclei of sebaceous neoplasms, including sebaceous gland carcinomas, show immunoreactivity for androgen receptors (AR), that immunohistochemical staining for the presence of AR may be a reliable marker of sebaceous differentiation, and that the AR may be a better marker of sebaceous differentiation than EMA, particularly in poorly differentiated sebaceous carcinomas. PMID- 10535571 TI - Secondary syphilis in persons infected with and not infected with HIV-1: a comparative immunohistologic study. AB - To better understand the cutaneous immune response to Treponema pallidum, we performed an immunohistologic study of skin biopsies from a total of 11 patients with secondary syphilis; biopsies from five persons infected with HIV-1 were included in the analysis to assess at the tissue level the impact of concomitant HIV-1 infection on disease expression. In all of the biopsies, staining for HLA DR, a marker for cellular activation, was observed among infiltrating leukocytes, dermal vascular endothelial cells, and keratinocytes. Infiltrating mononuclear cells stained positively for CD4 or CD8, with CD4+ cells always being in the majority. Surprisingly, most of the CD4+ cells had histiocytic, rather than lymphocytic, morphologic characteristics. Immunostaining for CD14 confirmed that these cells were monocytic in origin, whereas immunostaining for CD3 revealed that the lymphocytes were predominantly CD8+ cytotoxic T cells. B cells were not detected despite the presence of variable numbers of plasma cells in all specimens. By immunofluorescence, all of the specimens demonstrated perivascular deposition of immunoglobulins, complement, or fibrinogen; linear staining at the dermal-epidermal junction also was observed in most of the specimens. No differences in immunocytochemical or immunofluorescence staining patterns were observed between the specimens from patients who were HIV positive and patients who were HIV negative. In addition to providing a more precise definition of the infiltrating cells in syphilitic lesions, our results, taken as a whole, indicate that cellular immune processes are largely responsible for the development of cutaneous manifestations during syphilitic infection and that coinfection with HIV-1 has little discernible effect on the cutaneous response to T. pallidum. PMID- 10535572 TI - Increased p53 staining in normal skin of posttransplant, immunocompromised patients and implications for carcinogenesis. AB - The p53 tumor suppressor gene is a transcriptional activator involved in control of cell cycle. Nonmelanoma skin cancers and premalignant lesions in transplant patients have been associated with an increased rate of p53 mutation. It is possible that normal skin in transplant patients also has a more labile p53 tumor suppressor gene, predisposing them to the development of nonmelanocytic cutaneous malignancies. To test this hypothesis, we looked at p53 expression in normal skin from posttransplant, immunocompromised patients and compared this to p53 expression in normal skin from immunocompetent patients. Twenty-three skin biopsies of normal, non-sun-exposed skin from 23 immunosuppressed transplant patients and 6 skin biopsies of normal, non-sun-exposed skin from 3 immunocompetent patients were stained for p53 immunoreactivity. The skin biopsies from the immunocompromised patients showed increased staining for p53 when compared to the skin biopsies from the immunocompetent patients (mean = 7.52/mm for the immunocompromised patients and mean = 1.05/mm for the normal control group). Background levels of p53 mutation may be increased in normal skin of posttransplant immunocompromised patients. This background increase in p53 expression could reflect mutation of the gene, which may play a role in the subsequent development of cutaneous malignancies in this subgroup of patients. PMID- 10535573 TI - Elastic tissue in fibroepithelial polyps. AB - Fibroepithelial polyps are common cutaneous lesions with an unknown etiology. We attempted to demonstrate that fibroepithelial polyps develop secondary to a focal loss of elastic tissue. Forty-five fibroepithelial polyps were examined. All were stained with Verhoeff-van Gieson stain for elastic tissue and examined. All but one specimen had a normal amount of elastin and none revealed abnormally shaped elastic fibers. Abnormal or decreased elastic tissue is not the cause of fibroepithelial polyps. PMID- 10535574 TI - Inflammatory myofibroblastic tumor of the skin. AB - We report a case of inflammatory myofibroblastic tumor (IMF) of the skin in a female with a history of Wegeners granulomatosis. The patient had a painless, erythematous, and indurated lesion of the left elbow. The resected specimen revealed a 4 cm x 3 cm nodule involving the entire dermis and superficial portions of subcutis with a stellate profile at scanning magnification. There were spindle cells in fascicles and whorls and a mixed inflammatory cell infiltrate of plasma cells, lymphocytes, neutrophils, and eosinophils. The spindle cells were immunoreactive for vimentin, muscle specific actin, and smooth muscle actin. The polyclonal and polymorphous nature of the inflammatory cells was confirmed by immunohistochemical studies. This is the first case of IMF of the skin documented by immunostaining. PMID- 10535575 TI - Congenital multiple plaque-like glomangiomyoma. AB - Congenital glomus tumor is a rare clinical variant of glomus tumor, and glomangiomyoma is the least frequent histologic type of glomus tumor. We report a case of congenital multiple plaque-like glomangiomyoma in a 38-year-old man with multiple nodules and plaques on his left arm and forearm. Histopathologic study showed an angiomatous, nonencapsulated tumor with numerous highly folded dilated vascular lumina scattered throughout the dermis. The lumina were lined by a single layer of flat endothelial cells, and one to several rows of glomus cells were observed adjacent to the endothelial cells. Around large vessels, there was a gradual transition from glomus cells to elongated mature smooth muscle cells with thin and long "blunt-ended" nuclei. Immunohistochemically, there were strong positive reactions for cytoplasmic alpha-smooth muscle actin in glomus cells and smooth muscle cells, vimentin in glomus cells and endothelial cells, and desmin in the smooth muscle cells only. To our knowledge, this is the first case report of congenital multiple plaque-like glomus tumor with the microscopic appearance of a glomangiomyoma. PMID- 10535576 TI - Pilomatricoma associated with several hair follicles. AB - We report two cases of pilomatricoma in which the neoplasm was connected with several hair follicles. A 17-year-old boy developed an erythematous nodule, 1 cm in diameter, in the right temporal region, histologically showing basophilic cells connected with at least two hair follicles. In the other case, a 20-year old man had a centrally ulcerated tumor, 2 cm in diameter, in the left lower back. Basophilic cells proliferated in connection with at least six hair follicle like structures, accompanied by the development of shadow cells underneath. The histopathologic findings in these two cases suggest the existence of a subset of pilomatricomas that involve several pre-existing hair follicles. PMID- 10535577 TI - Spiradenoma arising in a nevus sebaceus of Jadassohn: case report and literature review. AB - Nevus sebaceus (NS) of Jadassohn is usually a verrucous plaque on the scalp or face that arises secondary to disordered development of epithelial, pilar, sebaceous, and apocrine structures. The emergence of neoplasia is a late stage in the natural history of NS. Although most neoplastic proliferations are benign, several malignant tumors have arisen in this lesion. We describe the first case of a benign spiradenoma arising in an NS on the scalp in a 72-year-old Caucasian woman. Reexcision was recommended to prevent the development of a second neoplastic process and to avoid the rare occurrence of a malignant transformation of the existing neoplasia. The patient declined reexcision and remains under observation. The spectrum of tumors arising in NS are described and are categorized according to behavior. Syringocystadenoma papilliferum is the most commonly observed benign growth, whereas basal cell carcinoma is the most frequently seen malignant process. The signs of tumor development (benign or malignant) within an NS are reviewed, and treatment recommendations are provided. The clinical course of rare and unique aggressive neoplasms originating in NS is summarized. PMID- 10535578 TI - Sebaceous carcinoma of the vulva. AB - Extraocular sebaceous carcinoma is an uncommon neoplasm usually localized on the head and neck. Sebaceous glands are abundant on the vulva, but vulvar sebaceous carcinoma is an uncommon neoplasm. To our knowledge, there are only five previously reported cases of sebaceous carcinoma on this location. We report an additional case of vulvar sebaceous carcinoma associated with Bowen's disease in the overlying epidermis. The patient also had bowenoid papulosis involving the skin of labia majora. We analyzed by immunohistochemistry, Southern blot hybridization, and polymerase chain reaction (PCR) techniques for the presence of DNA of human papilloma viruses (HPVs) in the specimen of sebaceous carcinoma and in lesions of bowenoid papulosis. Immunohistochemistry, Southern blot hybridization, and PCR studies in specimens of bowenoid papulosis lesions and sebaceous carcinoma did not detect DNA of HPVs. A significant increase in intranuclear p53 staining was demonstrated in several areas of neoplastic aggregations of sebaceous carcinoma. PMID- 10535579 TI - Basal cell carcinoma with tumor epithelial and stromal giant cells: a variant of pleomorphic basal cell carcinoma. AB - A case of basal cell carcinoma with giant cells of the central epithelial and surrounding stromal components is presented. The lesion was an 8-mm dome-shaped papule on the ear of a 66-year-old man. The giant cells of the epithelial component shared the immunophenotype of the more typical cells of the basal cell carcinoma (keratin, smooth muscle actin, and bcl-2 positive), whereas the stromal giant cells were positive only for bcl-2. This case represents a peculiar variant of pleomorphic basal cell carcinoma, the significance of which is unknown. PMID- 10535580 TI - Retiform/racemiform neoplasm with features of clear cell hidradenoma. AB - We report a case of a 78-year-old woman with a tumor of the left cheek. The tumor was a well-circumscribed cystic/solid nodule with a racemiform and reticulated pattern of growth of its epithelial cells, and mucinous and fibrocytic stroma. The epithelial cords and strands were continuous with the apocrine lining of large cystic structures. The main bulk of the epithelial component was formed by the proliferation of clear cells. This tumor is an example of an unusual benign neoplasm with racemiform and retiform patterns having a histogenetical link with the folliculo-sebaceous-apocrine unit. PMID- 10535581 TI - Epithelioid blue nevus occurring in children with no evidence of Carney complex. AB - We report two pediatric patients who had biopsies of solitary lesions diagnosed as epithelioid blue nevi. Histologically these lesions had wedge-shaped, heavily pigmented infiltrates extending to the subcutaneous fat. The infiltrate was composed of spindled and polyhedral cells that were nevomelanocytic cells with nuclear pleomorphism. Also noted were pigmented globular cells interpreted as melanophages. These lesions have the same characteristics as those blue nevi occurring in patients with Carney complex. More recently, adult patients have been identified with similar nevi, but without evidence of Carney complex. To our knowledge, pediatric patients with epithelioid blue nevi, but no evidence of Carney complex have not been described previously. PMID- 10535582 TI - Cytomegalovirus-induced syringosquamous metaplasia. AB - An unusual case of syringosquamous metaplasia of the eccrine ducts caused by cytomegalovirus (CMV) is presented. The patient was HIV positive and had extensive excoriation of the perineum and vulva. Biopsy revealed the presence of herpes simplex virus (HSV) inclusions in the necrotic exudate, a CMV vasculitis and extensive involvement of the eccrine ducts. In addition to containing typical CMV inclusions, the eccrine ducts showed proliferation and squamous metaplasia. Inclusions of HSV were not seen within the eccrine ducts by light microscopy or immunohistochemistry. The extensive proliferation with accompanying squamous metaplasia superficially can resemble an infiltrating squamous carcinoma, but this was not evident to a great extent in this case. To the best of our knowledge, our case represents the first of syringosquamous metaplasia of eccrine ducts caused by CMV infection. PMID- 10535583 TI - Pityriasis rubra pilaris with acantholysis and lichenoid histology. AB - Acantholytic foci have been reported several times in pityriasis rubra pilaris (PRP). Lichenoid tissue reactions were also mentioned in the literature regarding PRP. We report a 58-year-old patient who, after having colon cancer, had PRP with biopsies showing acantholytic lesions and a heavy lichenoid lymphocytic infiltration. Investigation by serial sectioning of the acantholytic lesion suggested an involvement of the intraepidermal eccrine duct and further investigation with carcinoembryonic antigen (CEA) staining demonstrated a CEA positive eccrine duct in the acantholytic foci. We suggest that acantholysis in PRP is induced by proteolytic enzymes, urea, and other substances in eccrine sweat in keratin-plugged acrosyringia. This patient had a combination of three relatively rare features of PRP-acantholysis, lichenoid reaction, and a cancer background. PMID- 10535584 TI - Penile intraepithelial neoplasia overlying Kaposi's sarcoma lesions: role of viral synergy? AB - Several viral agents have been detected in the lesional tissue of Kaposi's sarcoma (KS). Their precise oncogenic role remains to be determined. A 32-year old heterosexual man with acquired immunodeficiency syndrome (AIDS) who had penile lesions of KS with overlying epithelial changes characteristic of intraepithelial neoplasia associated with concurrent infection by human papillomavirus (HPV) and human herpesvirus 8 (HHV-8) is reported. The absence of viral DNA from uninvolved skin suggests that this coinfection is more than coincidental and may involve synergy between these viruses, as has already been suggested for HPV and herpes simplex 2 virus. PMID- 10535585 TI - How useful are T-cell receptor gene rearrangement studies as an adjunct to the histopathologic diagnosis of mycosis fungoides? AB - The diagnosis and differential diagnosis of mycosis fungoides (MF) is often difficult, clinically and histologically. Recent attempts to enhance diagnostic sensitivity have involved T-cell receptor (TCR) gene rearrangement studies, using Southern blot or polymerase chain reaction (PCR) technique. PCR is more widely used because of its increased sensitivity, lower labor- and cost-intensive analytic steps, lack of radioactive substances, and applicability to routinely processed biopsies. Several studies that use this technique detect clonal bands in 50% to 83% of the MF specimens from patch, patch/plaque, and erythrodermic stages, which often pose a diagnostic challenge. This compares with no clonality in the control groups of most of these studies, or with up to 13% in a few studies, although long-term follow-up reveal that some cases of so-called "clonal dermatitis" eventually progress into overt cutaneous T-cell lymphoma. Furthermore, retrospective studies on archival histologic material from patients with MF demonstrate similar rates of clonality in histologically "inconclusive," "borderline," and diagnostic MF biopsies. Therefore, in the proper clinicopathologic setting, and with consideration of the known limitations of this technique, TCR gene rearrangement studies on lesional skin using PCR may be helpful as an adjunct to the histopathologic diagnosis of MF. PMID- 10535586 TI - Arthroscopic Bankart repair of anterior detachments of the glenoid labrum. A prospective study. AB - BACKGROUND: The purpose of this study was to evaluate the results of an arthroscopic transglenoid suture-stabilization procedure in athletically active patients who had recurrent, unilateral, unidirectional anterior dislocations of the shoulder and an isolated anterior detachment of the glenoid labrum. METHODS: Forty-one patients who had unilateral, unidirectional anterior dislocations of the shoulder and an isolated anterior detachment of the glenoid labrum were managed with arthroscopic repair. All patients were athletic, and seventeen of the male patients were football players. No patient had inferior or posterior laxity or a posterior detachment. The sutures were anchored to the posterior aspect of the scapula, and the knots were tied anteriorly to secure the detached region of the labrum and the inferior glenohumeral ligament to the anterior aspect of the scapula. The mean duration of follow-up was fifty-two months (range, twenty-five months to seven years). The patients were evaluated annually with a physical examination, radiographs, isokinetic strength-testing, the modified shoulder-rating scale of Rowe and Zarins, and the scoring system of the American Shoulder and Elbow Surgeons. RESULTS: Forty (98 percent) of the forty one athletes returned to their preoperative sport postoperatively. Thirty-nine patients (95 percent) had no additional dislocations or subluxations, and two (5 percent), both of whom were football players, had a single episode of subluxation. Thirty-seven patients (90 percent) had a score of at least 80 points on the scale of Rowe and Zarins, and thirty-four (83 percent) had a score of at least 90 points. Thirty-nine patients (95 percent) had a score of at least 80 points on the scale of the American Shoulder and Elbow Surgeons, and twenty-five (61 percent) had a score of at least 90 points. Lower scores were associated with loose bodies seen on arthroscopy (p = 0.001), osseous lesions seen on postoperative radiographs (p = 0.036), and subluxation (p = 0.000). Twenty-two shoulders (54 percent) had a full range of motion in all planes, and eighteen (44 percent) had no strength deficit in any position on isokinetic testing. With the numbers available for study, no significant association was found between the presence of a Hill-Sachs or an osseous Bankart lesion on preoperative radiographs and the overall score on the scale of Rowe and Zarins or the scale of the American Shoulder and Elbow Surgeons; however, there was a significant association between the range of motion and an osseous Bankart lesion on preoperative radiographs (p = 0.002) and between decreased strength on isokinetic testing and a Hill-Sachs lesion on preoperative radiographs and an osseous lesion on postoperative radiographs (p = 0.022). There also was a significant association between a decreased range of motion (p < 0.002) and decreased strength (p = 0.014) and the arthroscopic finding of loose bodies. Muscle strength also was affected by arm dominance and the number of preoperative dislocations. CONCLUSIONS: Arthroscopic transglenoid repair of isolated anterior labral detachments restored stability of the shoulder and led to a favorable outcome in thirty-nine (95 percent) of the forty-one athletes. Only the two football players who had postoperative subluxation had a score of less than 80 points according to the scale of the American Shoulder and Elbow Surgeons. PMID- 10535587 TI - Natural history of congenital kyphosis and kyphoscoliosis. A study of one hundred and twelve patients. AB - BACKGROUND: Congenital kyphosis and kyphoscoliosis are much less common than congenital scoliosis. However, they are potentially more serious because compression of the spinal cord and paraplegia sometimes develop. The goals of the present study were to document the natural history of congenital kyphosis and kyphoscoliosis and to determine the stage at which the natural progression should be interrupted by treatment. METHODS: We reviewed the medical records and radiographs of the spine of 112 consecutive patients. Sixty-eight patients had a type-I kyphosis due to an anterior failure of vertebral-body formation, twenty four had a type-II kyphosis due to an anterior failure of vertebral-body segmentation, and twelve had a type-III kyphosis due to a combination of anomalies; the deformities of the remaining eight patients could not be classified. Eighty-five skeletally immature, untreated patients were first evaluated at a mean age of six years and nine months (range, two months to sixteen years and three months), and twenty-seven patients were skeletally mature at the time of the first visit. Sixty-three of the eighty-five skeletally immature patients were observed without any treatment for a mean period of six years and six months (range, one to sixteen years) before skeletal maturity, and the remaining twenty-two patients had a posterior arthrodesis of the spine soon after the initial visit. At skeletal maturity, forty-one patients had not been treated and sixty-eight had had an arthrodesis of the spine. The remaining three patients had not yet reached skeletal maturity at the time of the most recent follow-up. RESULTS: The apex of the kyphosis was seen at all levels but was most frequent between the tenth thoracic and the first lumbar level (seventy-four patients; 66 percent). There was no relationship between the severity of the kyphosis and its location in the spine. Progression of the curve was most rapid during the adolescent growth spurt and stopped only at skeletal maturity. Progression was most rapid and the magnitude of the curve was the greatest in type-III kyphosis (twelve patients) followed by type-I kyphosis due to a posterolateral quadrant vertebra (thirty-nine patients), a posterior hemivertebra (eight patients), a butterfly vertebra (fifteen patients), and a wedged vertebra (six patients). A kyphosis due to two adjacent type-I vertebral anomalies progressed more rapidly and produced a more severe deformity than did a similar single anomaly. The prognosis for type-II kyphosis was variable and was much more severe when an anterolateral unsegmented bar had produced a kyphoscoliosis (nine patients) than it was when a midline anterior bar had produced a pure kyphosis (fifteen patients), which usually progressed slowly. Spontaneous neurological deterioration due to compression of the spinal cord occurred in ten patients (seven of whom had a type-I kyphosis and three of whom had an unclassifiable anomaly) at a mean age of thirteen years and eight months, and one other patient (with an unclassifiable anomaly) had spastic paraparesis at the age of twenty eight years. CONCLUSIONS: Congenital kyphosis and kyphoscoliosis are uncommon deformities with the potential to progress rapidly, resulting in severe deformity and possible neurological deficits. A thorough knowledge of the natural history is essential in the planning of appropriate and timely treatment to prevent progression of the deformity and neurological complications. PMID- 10535588 TI - Long-term follow-up study of bilateral above-the-knee amputees from the Vietnam War. AB - BACKGROUND: Because caring for patients who have combat-related amputations is a discontinuous practice, military surgeons must relearn treatment techniques during each conflict. METHODS: The purpose of the present long-term study (average duration of follow-up, 27.5 years) was to document the status of patients who had sustained a bilateral above-the-knee amputation in Vietnam and had been managed by the only separate amputee service in the United States Army. A review of the records of 484 battle amputees identified thirty individuals (6 percent) who had a bilateral above-the-knee amputation. Twenty-six (87 percent) of the thirty patients had been injured by a land mine or a booby trap. Fifty three (88 percent) of the sixty limbs were amputated because of trauma, and the other seven (12 percent) were amputated secondarily because of infection. Data regarding education, employment, marriage and family life, prosthetic use, and psychological care were collected by mail or telephone for twenty-three (85 percent) of the twenty-seven surviving patients. Respondents also completed the Short Form-36 (SF-36) Health Survey. RESULTS: At the time of the study, five (22 percent) of the twenty-three respondents used prostheses for walking; the devices were used for an average of 7.7 hours per day. Sixteen respondents (70 percent) were or had been employed outside of the home since the time of discharge. The physical functioning score on the SF-36 questionnaire was significantly lower for the study group than it was for a group of age and gender-matched controls (p < 0.001; Student two-tailed t test). With the numbers available, no significant differences could be detected between the groups with regard to physical role functioning (p = 0.377), bodily pain (p = 0.603), general health (p = 0.407), vitality (p = 0.949), social functioning (p = 0.460), emotional role functioning (p = 0.029), or mental health (p = 0.102). CONCLUSIONS: The patients in the present study have led relatively normal, productive lives within the context of their physical limitations. PMID- 10535589 TI - Triple arthrodesis: twenty-five and forty-four-year average follow-up of the same patients. AB - BACKGROUND: Triple arthrodesis is used to treat major deformities of the hindfoot and is often performed in young patients. The purpose of this study was to assess the long-term outcomes of triple arthrodesis in young patients. METHODS: Sixty seven feet of fifty-seven patients were evaluated at an average of twenty-five and forty-four years after triple arthrodesis. The most common indication for the operation was neuromuscular imbalance of the hindfoot, which was secondary to poliomyelitis in thirty-seven feet (55 percent), Charcot-Marie-Tooth disease in six (9 percent), spinal cord abnormalities in four (6 percent), cerebral palsy in three (4 percent), and Guillain-Barre syndrome in one (1 percent). RESULTS: Fifty two feet (78 percent) had some residual deformity after the arthrodesis. However, these deformities appeared to be nonprogressive between 1973 and 1994. Pseudarthrosis occurred in thirteen feet. Thirty feet or ankles (45 percent) were painful at the first follow-up evaluation, and thirty-seven feet or ankles (55 percent) were painful at the second follow-up evaluation. Of the thirty feet or ankles that were painful at the first follow-up evaluation, twenty-three were painful at the second follow-up evaluation. Of the thirty-seven feet or ankles that were not painful at the first follow-up evaluation, fourteen were painful at the second follow-up evaluation. Eighteen patients (32 percent) needed walking support at the time of the first follow-up, and thirty-nine patients (68 percent) needed it at the time of the second follow-up. Two of the patients who needed support at the first follow-up evaluation did not need it at the second follow-up evaluation. At the first follow-up evaluation, twenty-one ankles (31 percent) had no radiographic evidence of degenerative changes. However, by the second follow up evaluation, all of the ankles had some degenerative changes. Similar progressive arthritic findings were noted at the naviculocuneiform and tarsometatarsal joints. According to the system of Angus and Cowell, the overall result at the time of the first follow-up was rated as good in fifty feet (75 percent) and as fair in seventeen feet (25 percent). At the time of the second follow-up, nineteen feet (28 percent) were rated as good, forty-six (69 percent) were rated as fair, and two (3 percent) were rated as poor. CONCLUSIONS: Despite progressive symptoms and radiographic degeneration in the joints of the ankle and midfoot, fifty-four patients (95 percent) were satisfied with the result of the operation. The triple arthrodesis was a satisfactory solution for imbalance of the hindfoot in this group of patients. PMID- 10535590 TI - The influence of medial and lateral placement of orthotic wedges on loading of the plantar aponeurosis. AB - BACKGROUND: Repetitive trauma and overuse of the plantar aponeurosis are believed to be causal factors of plantar fasciitis. Therefore, it is important to know how an orthosis influences loading of the plantar aponeurosis. The aim of this study was to quantify strain in the plantar aponeurosis in cadaveric feet with the use of various combinations of orthotic wedges. METHODS: An in vitro test that simulated static stance was used to determine the loading characteristics of the plantar aponeurosis. A differential variable reluctance transducer was operatively implanted into the plantar aponeurosis of nine fresh-frozen cadaveric lower limbs. Each specimen was mounted in an electromechanical testing machine that applied an axial load of as much as 900 newtons to the tibia. Eight different combinations of test conditions, in which wedges (each with a 6-degree incline) were or were not positioned under the medial and lateral aspects of the forefoot and hindfoot, were evaluated, with the plantigrade foot used as a neutral control. RESULTS: Each of the test conditions that involved a wedge under the forefoot resulted in strain that was significantly different from that in the neutral control. A wedge under the lateral aspect of the forefoot decreased strain in the plantar aponeurosis, and a wedge under the medial aspect increased strain (p < 0.05). The test conditions that involved a wedge under the hindfoot but not under the forefoot resulted in strains that were not significantly different from those in the neutral control (p > 0.05). CONCLUSIONS: A wedge under the lateral aspect of the forefoot transmits loads through the lateral support structures of the foot, locking the calcaneocuboid joint and decreasing strain in the plantar aponeurosis. A wedge under the medial aspect of the forefoot transmits loads through the medial support structures of the foot, which produces a truss-like action that increases strain in the plantar aponeurosis. PMID- 10535591 TI - Vascularized bone graft from the iliac crest for the treatment of nonunion of the proximal part of the scaphoid with an avascular fragment. AB - BACKGROUND: It was hypothesized that nonunion of the proximal third of the scaphoid associated with avascular necrosis could be treated successfully with a free vascularized bone graft obtained from the iliac crest. METHODS: Fifteen patients who had a nonunion of the proximal part of the scaphoid that had been present for an average of two years and three months (range, nine months to seven years) were managed with use of a free vascularized bone graft obtained from the iliac crest. Avascularity of the scaphoid, as assessed on preoperative radiographs, was characterized by loss of trabecular structure, collapse of subchondral bone, and formation of bone cysts. The results of the procedure were assessed in terms of osseous union, pain, active motion of the wrist, and osteoarthritis. Postoperatively, vascularity of the scaphoid was evaluated with use of magnetic resonance imaging and color Doppler ultrasonography. The average duration of follow-up was six years and one month (range, two years and one month to eight years and one month). RESULTS: Preoperatively, one patient had had pain with any movement of the wrist and fourteen had had pain after strenuous manual labor or sports activity. The average pain score, derived with use of a 10-point visual analog scale, was 2.4 points (range, 1.0 to 6.7 points). Postoperatively, union was achieved in twelve patients; six were pain-free, and six had occasional pain during strenuous manual labor or sports activity, or both. The average pain score for these twelve patients was 1.1 points (range, 0.0 to 4.2 points) on the visual analog scale. Preoperatively, osteoarthritis was limited to the region between the radial styloid process and the distal part of the scaphoid in fourteen patients and to the radioscaphoid region in one patient. Postoperatively, the degree of osteoarthritis remained unchanged in seven of the twelve patients who had union and progressed to the radioscaphoid region in five. Vascularity, as seen on the imaging studies, was restored in all twelve patients who had union. The nonunion persisted in three patients, all of whom had progressive osteoarthritis leading to carpal collapse. CONCLUSIONS: The index procedure was successful in twelve of the fifteen patients who had a symptomatic nonunion of the proximal part of the scaphoid associated with avascular necrosis and osteoarthritis that was limited to the radioscaphoid joint. PMID- 10535592 TI - The posterior fat pad sign in association with occult fracture of the elbow in children. AB - BACKGROUND: An elevated posterior fat pad visible on a lateral radiograph of a child's elbow following trauma is generally considered to be suggestive of an intracapsular fracture about the elbow. However, in previous studies, the prevalence of fracture in elbows with an elevated posterior fat pad and no other radiographic evidence of fracture has ranged from only 6 percent (two of thirty one) to 29 percent (nine of thirty-one). We are not aware of any prospective studies, limited to children, on the value of an elevated posterior fat pad as an indicator of an occult fracture about the elbow. While it is common practice to manage children who have radiographic evidence of an elevated posterior fat pad as if they have a fracture, scientific evidence for this approach is lacking. METHODS: Forty-five consecutive children who had an average age of four and a half years, a history of trauma to the elbow, and an elevated posterior fat pad without other radiographic evidence of a fracture were enrolled in the study. At an average of three weeks after the injury, anteroposterior, lateral, and two oblique radiographs were made and evaluated for evidence of fracture-healing. If there was evidence of new-bone formation on any of these four radiographs, it was considered to indicate a fracture of the elbow. RESULTS: Thirty-four (76 percent) of the forty-five patients had evidence of a fracture. Eighteen (53 percent) of the thirty-four had a supracondylar fracture of the humerus; nine (26 percent), a fracture of the proximal part of the ulna; four (12 percent), a fracture of the lateral condyle; and three (9 percent), a fracture of the radial neck. CONCLUSIONS: This prospective study demonstrated that the posterior fat pad sign was predictive of an occult fracture of the elbow following trauma in thirty-four (76 percent) of forty-five children who had no other evidence of fracture on anteroposterior, lateral, and oblique radiographs after the injury. This finding is in contrast to those of previous studies, in which the highest prevalence of fracture in elbows with an elevated posterior fat pad and no other radiographic evidence of fracture was 29 percent (nine of thirty-one elbows). Our results support the practice of managing children who have a history of trauma to the elbow, an elevated posterior fat pad, and no other radiographic evidence of fracture as if they have a nondisplaced fracture about the elbow. PMID- 10535594 TI - Early osteolysis following total hip arthroplasty with use of a Hylamer liner in combination with a modular ceramic femoral head. A case report. PMID- 10535593 TI - Infection after total knee arthroplasty. A retrospective study of the treatment of eighty-one infections. AB - BACKGROUND: The clinical presentation of an infection at the site of a total knee arthroplasty can be used as a guide to treatment, including the decision as to whether the prosthesis should be retained or removed. We reviewed the results of treatment of infection after total knee arthroplasty to evaluate the effectiveness of four treatment protocols based on the clinical setting of the infection. METHODS: We retrospectively evaluated the results of treatment of eighty-one infections in seventy-six consecutive patients who either had an infection after a total knee arthroplasty or had multiple positive intraoperative cultures of specimens of periprosthetic tissue obtained during a revision total knee arthroplasty performed because of presumed aseptic loosening. The patients were managed according to one of four protocols. Five infections in five patients who had positive intraoperative cultures were treated with antibiotic therapy alone. Twenty-three early postoperative infections in twenty-one patients were treated with debridement, antibiotic therapy, and retention of the prosthesis. Twenty-nine late chronic infections in twenty-eight patients were treated with a delayed-exchange arthroplasty after a course of antibiotics. Seven acute hematogenous infections in six patients were treated with debridement, antibiotic therapy, and retention of the prosthesis. Seventeen infections in seventeen patients were not treated according to one of the four protocols. Sixteen late chronic infections were treated either with an arthrodesis (five infections) or with debridement, antibiotic therapy, and retention of the prosthesis (eleven infections). One acute hematogenous infection was treated with resection arthroplasty because of life-threatening sepsis. RESULTS: The mean duration of follow-up was 4.0 years (range, 0.3 to 14.0 years). Eleven patients who had an arthrodesis, a resection arthroplasty, or an above-the-knee amputation after less than two years of follow-up were included in the study as individuals who had a failure of treatment. In the group of patients who were managed according to protocol, the initial course of treatment was successful for all five infections that were diagnosed on the basis of positive intraoperative cultures, five of the ten deep early infections, all thirteen superficial early infections, twenty-four of the twenty-nine late chronic infections, and five of the seven acute hematogenous infections. Only one of eleven prostheses in patients who had a late chronic infection that was not treated according to protocol was successfully retained after debridement. CONCLUSIONS: Our treatment protocols, which were based on the clinical setting of the infection, were successful for most patients. A major factor associated with treatment failure was a compromised immune status. Bone loss and necrosis of the soft tissues around the joint also complicated the treatment of these infections. PMID- 10535595 TI - Partial avulsion of the cauda equina associated with a lumbosacral fracture dislocation. A case report. PMID- 10535596 TI - Sample size and statistical power in clinical orthopaedic research. PMID- 10535597 TI - The fallacy of short-term outcomes analysis in pediatric orthopaedics. PMID- 10535598 TI - The natural history of debonding of the femoral component from the cement and its effect on long-term survival of Charnley total hip replacements. PMID- 10535599 TI - The adequacy of medical school education in musculoskeletal medicine. PMID- 10535600 TI - Use of an intramedullary hip-screw compared with a compression hip-screw with a plate for intertrochanteric femoral fractures. A prospective, randomized study of one hundred patients. PMID- 10535602 TI - Chemokines in the inflammatory bowel diseases. AB - Ulcerative colitis and Crohn's disease are characterized by chronic intestinal inflammation. Intestinal bacteria initiate the activation of intestinal inflammatory processes, which are mediated by proinflammatory cytokines and chemokines. In inflammatory bowel disease, intestinal inflammation is not downregulated, in part due to defective or absent inhibitory processes. Studies to date have demonstrated that IL-8, MCP-1, and ENA-78 are highly expressed in the intestinal mucosa in areas of active Crohn's disease and ulcerative colitis. Neutrophils and macrophages in the inflamed intestine synthesize and secrete large amounts of chemokines in patients with inflammatory bowel disease. Increased chemokine expression has also been observed in epithelial cells, endothelial cells, and smooth muscle cells. Future trials of specific agents capable of inhibiting chemokine synthesis and secretion or blocking chemokine chemokine receptor interaction will be important to study in patients with ulcerative colitis and Crohn's disease. PMID- 10535601 TI - Chemokines and chemokine receptors: their role in allergic airway disease. AB - One of the hallmarks of allergic pulmonary disorders is the accumulation of an abnormally large number of leukocytes including eosinophils, neutrophils, lymphocytes, basophils, and macrophages in the lung. There is now substantial evidence that eosinophils, under the control of T lymphocytes, are major effector cells in the pathogenesis of asthma. Therefore, understanding the mechanisms by which eosinophils accumulate and are activated in tissues is a fundamental question very relevant to allergic diseases. Another characteristic of allergic inflammation is the activation of leukocytes resulting in the release of biologically active mediators, such as histamine from mast cells and basophils. It is now apparent that chemokines are potent leukocyte chemoattractants, cellular activating factors, histamine releasing factors, and regulators of homeostatic immunity, making them particularly important in the pathogenesis of airway inflammation in asthma. In this regard, chemokines are attractive new therapeutic targets for the treatment of allergic disease. This article focuses on recently emerging data on the importance of chemokines and their receptors in allergic airway inflammation. PMID- 10535603 TI - Role of chemokines in the regulation of Th1/Th2 and autoimmune encephalomyelitis. AB - Chemokines are low molecular weight chemotactic peptides that bind seven transmembrane-spanning, G protein-coupled receptors and deliver signals leading to T cell costimulation, hematopoeisis, cytokine expression, T cell differentiation, and integrin activation. Experimental autoimmune encephalomyelitis (EAE) is a CD4+ Th1-mediated demyelinating disease of the central nervous system (CNS) that serves as a model for multiple sclerosis (MS). A hallmark in the pathogenesis of this CNS demyelinating disease is the emigration of T cells and monocytes from the blood to the CNS. There are several considerations that suggest a role for chemokines in the influx of inflammatory cells and the resulting disease process including a tight temporal expression pattern with relationship to disease activity and prevention of disease development by in vivo neutralization. We review the evidence that temporal and spatial expressions of chemokines are crucial factors, complementing adhesion molecule upregulation, that regulate EAE and potentially MS disease activity as well as the functions of chemokines in Th1 and Th2 biology. PMID- 10535604 TI - Chemokines as molecular targets for therapeutic intervention. AB - Despite the youth of the chemokine field, many antagonists of chemokine function have already been identified and tested at the preclinical level. These include neutralizing antibodies, peptidyl and non-peptidyl antagonists and non-specific immunosuppressive agents. These early studies suggest that chemokine agonists have the potential to regulate many diseases, ranging from HIV-1 infection and tumor growth to acute and chronic inflammation. Clinical application will depend on pharmaceutical development. Great strides have been made in defining structural domains of the chemokines involved in receptor binding and activation. The identification of receptors is rapidly progressing, but with 50 potential ligands and 15 characterized receptors, it is obvious that additional molecular studies are needed. The intriguing observation that several pathogens either use chemokine receptors as entry portals or produce chemokine decoys to subvert the immune system suggests that there is much to be learned about the immune system from studies of "virokines." Future studies should lead to the discovery and design of more effective inhibitors and antagonists with therapeutic benefit. PMID- 10535605 TI - HIV infection and pathogenesis: what about chemokines? AB - Several chemotactic cytokines, or chemokines. inhibit HIV replication by blocking or down regulating chemokine receptors that serve as entry cofactors for the virus. Although the role of chemokine receptors in HIV pathogenesis has been the subject of intense interest, chemokines are comparatively less seriously considered as potential correlates of protection from HIV infection and disease progression. However, a critical analysis of newly available data reveals substantial evidence to support a beneficial role for chemokines in HIV infection and disease. In this review we summarize the results of such studies and their promising implications for HIV infection. PMID- 10535606 TI - C3/C4 concentration ratio reverses between colostrum and mature milk in human lactation. AB - The levels of complement fractions C3 and C4 were assayed in human milk in a classic nephelometric assay adapted to this secretion. Concentrations of these molecules were measured in 667 milk samples obtained sequentially from 76 volunteer lactating mothers during the first 12 weeks of lactation. Immunonephelometry was performed using skimmed milk samples diluted 10 times and yielded reproducible (coefficients of variation in within- and between-run precision lower than 9% for C3 and than 14% for C4) and accurate (linear recovery in dilution-overloading assay) data. High concentrations (mean +/- SE) were found for C3 (199.32+/-16.35 mg/L) and C4 (113.42+/-11.16 mg/L) in colostrum samples (n = 159; days 1-5). A significant (P<0.001) and rapid decrease was observed in transitional milk samples (n = 198; days 6-14), containing 57.71+/-5.18 and 72.39+/-4.98 mg/L of C3 and C4, respectively. Stable lower levels were noted in mature milk samples (n = 310; days 15-84) at 30.36+/-1.57 mg/L for C3 (P<0.001) and 53.38+/-3.61 mg/L for C4 (P<0.05). The decrease rate was different for C3 and C4, yielding a reversal of the C3/C4 ratio between colostrum and more mature milk. PMID- 10535608 TI - TNF-alpha and chronic fatigue syndrome. AB - Based upon the clinical presentation of chronic fatigue syndrome (CFS), we hypothesized that proinflammatory cytokines may play a role in the pathogenesis of the disease. We therefore undertook a retrospective cross-sectional study to examine the role of TNF-alpha in patients with CFS. Our results suggest a significant increase serum TNF-alpha in patients with CFS (P<0.0001) compared to non-CFS controls. This study supports the further examination of the role of proinflammatory mediators in CFS. Furthermore, the clinical testing of TNF-alpha blockers and other antiinflammatory agents for the treatment of this disease is warranted. PMID- 10535607 TI - Long-term follow-up of the changes in circulating cytokines, soluble cytokine receptors, and white blood cell subset counts in patients with rheumatoid arthritis (RA) after monoclonal anti-TNF alpha antibody therapy. AB - To investigate the mechanism of the long-lasting efficacy of chimeric monoclonal anti-TNFalpha antibody (cA2) therapy for rheumatoid arthritis (RA), eight patients with refractory RA were treated with a single infusion of cA2 and the changes in circulating cytokines (IL-1, IL-6, TNFalpha, and IL-10), soluble cytokine receptors (TNF-RI, RII, and sIL-6R) and peripheral white blood cell (WBC) subset counts were followed up long-term (12 weeks) after cA2 therapy in them. Significant clinical responses (>20% improvement according to Paulus' criteria) were observed just after cA2 infusion and lasted more than 4 weeks in all patients, as reported elsewhere. Moreover, five of the eight patients showed prolonged clinical responses (>12 weeks). The elevated serum IL-6 and sTNF-RI (or RII) levels before treatment rapidly decreased after treatment. The serum IL-10 levels also significantly elevated before treatment. The elevations of serum IL 10 levels were augmented after treatment and stayed higher than the baseline in four patients with prolonged clinical responses. No significant TNFalpha, IL 1alpha and -beta, or sIL-6R were detected in the sera of the patients before treatment and during the whole study period. On the other hand, peripheral lymphocytes as well as total WBC and neutrophils increased for 4 weeks after treatment. However, thereafter, only the lymphocyte count decreased gradually and stayed below the baseline long-term (12 weeks). FACS analysis revealed the predominance of T lymphocytes in the decrease in lymphocyte counts. These results suggest that the augmentation of IL-10 production and the decrease in T cells might partly contribute to the long-lasting efficacy of cA2 treatment in RA. PMID- 10535611 TI - Tumor necrosis factor-alpha is decreased in the umbilical cord plasma of patients with severe preeclampsia. AB - We investigated the role of the fetal immune system in pregnancies complicated by preeclampsia by assessing umbilical cord plasma levels of the cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta). Nineteen nulliparous patients with severe preeclampsia composed the study group (group A). A comparison group was comprised of 19 healthy nulliparous patients with uneventful pregnancies (group B). Mixed umbilical cord blood was collected immediately after delivery. Plasma was prepared and all samples were assayed for TNF-alpha and IL-1beta by specific enzyme-linked immunoassays (ELISAs). Data are presented as the median with range of values. The length of labor was similar in both groups. TNF-alpha was detected less frequently in the umbilical cord plasma of preeclamptic patients than in the umbilical cord plasma of control patients (57.9 vs. 89.5%, p < 0.05), and the concentrations of TNF-alpha were significantly lower in the umbilical cord plasma of the preeclamptic patients [20 pg/ml (0-80 pg/mL) vs. 50 pg/mL (0-310 pg/mL), p < 0.05]. Umbilical cord plasma IL-1beta detection rates and concentrations from the preeclamptic and control patients were similar, [15.8 vs. 5.3%, 0 pg/mL (0-40 pg/mL) vs 0 pg/mL (0-10 pg/mL)]. The lower concentrations of TNF-alpha in umbilical cord plasma of patients with severe preeclampsia suggest that release of TNF-alpha by the fetus and mother are independent and may reflect adaptation of the fetus to reduced placental perfusion in preeclampsia. PMID- 10535610 TI - Impaired IgG antibody production to pneumococcal polysaccharides in patients with ataxia-telangiectasia. AB - Various factors seem to be etiologic in the susceptibility to sinopulmonary infections in ataxia-telangiectasia (A-T) patients, i.e., low serum and salivary IgA, low serum IgG2, and even aspiration of saliva. S. pneumoniae is a common pathogen responsible from pulmonary infections and impaired antibody response to polysaccharide antigens is seen in patients with IgG2 and IgA deficiency as well as patients with CVID and WAS. We studied IgG-type antibody production to six pneumococcal serotypes in 29 A-T patients by ELISA before and 3-4 weeks after pneumococcal vaccine. The response was considered positive when the antibody titer was >10 U/ml but weak when the titer was 10-20 U/ml. Twenty-two of 29 (76%) patients did not respond to any of the serotypes, 5 (17%) showed a positive response to one serotype, 1 (3.4%) to two serotypes, and 1 (3.4%) to four serotypes. With conversion to gravimetric units (ng IgG/ml) and >1800 ng/ml (300 ng Ab N/ml) considered a positive response, 5 of 29 (17.2%) patients showed a positive response (300 ng ab N/ml) to two or fewer serotypes. All patients tested produced IgG antibody to tetanus toxoid. Sixteen of 27 (59.3%) patients had low IgG2 and four (14.8%) had low IgG3 levels, while 18 (62.1%) of 29 patients had low serum IgA. No correlation was found either between serum Ig isotype levels and antipolysaccharide antibody response or between susceptibility to infection and antibody production. The mechanism responsible for disturbed antipolysaccharide (TI-2 antigen) antibody production in patients with A-T needs to be investigated. It may provide additional information on the function of the ATM gene product and be helpful in clarifying the role of B cells and contribution of T cells in TI-2 responses. PMID- 10535609 TI - Kinetics of lymphokine production in HIV+ patients treated with highly active antiretroviral therapy and interleukin 2. AB - This study presents the kinetics of CD4/CD25 cell numbers, serum sCD25 levels, and intracellular production and release of interleukin-2 (IL-2) and interleukin 16 (IL-16) in 11 HIV+ patients treated with six cycles of highly active antiretroviral therapy (HAART) plus six MUI of subcutaneous IL-2 compared to 10 HIV+ patients treated with HAART alone. IL-2 therapy induced moderate effects on CD4 T cell recovery and increased CD4/CD25+ cells and sCD25 levels after 2 weeks, while intracellular and secreted IL-2 was reduced and IL-16 was increased at the same time point. After 24 weeks, while HAART-treated patients had increased IL-2 production, in IL-2 treated patients, cytokine production was unaltered compared to pretreatment values. Decreased in vitro IL-2 production may depend on a feedback inhibition by IL-2 infusion. Because of its known antiviral effects, the increased IL-16 production seen after 2 weeks in IL-2-treated individuals may produce beneficial effects on HIV disease. The kinetics of cytokine production may serve to define better the use IL-2 in clinical trials. PMID- 10535612 TI - Prophylactic ranitidine in experimental fetal distress: acute phase effects on the fetal stomach. AB - Fetal distress (FD) adversely affects fetal gastric physiology and histology, increasing gastric acid secretion and disturbing gastric protective mechanism. Considering these findings, an experimental study was planned to test whether ranitidine prevents FD-related gastric physiological and histological changes during late gestational period. In this study, a rabbit model of FD was created by way of intermittent maternal aortic occlusion. In group 1 (SC), saline treated animals underwent control operation. In group 2 (SD), FD was created in saline treated animals. In group 3 (RC), ranitidine treated animals underwent control operation. In group 4 (RD), FD was created in ranitidine treated animals. Blood lactic acid levels of the fetuses were 2.3 +/- 1.0 mg/L in SC group and 4.7 +/- 1.8 mg/L in group SD (p < 0.01); 2.5 +/- 0.9 mg/L in group RC and 6.7 +/- 2.5 mg/L in group RD (p < 0.01). Fetal gastric acid secretion was 5.94 +/- 2.13 microEq/h in group SC and 8.26 +/- 2.24 microEq/h in group SD (p < 0.05); 6.63 +/ 2.3 microEq/h in group RC and 6.04 +/- 2.43 microEq/h in group RD (p < 0.05). Fetal gastric PGE2 level was 16.4 +/- 2.65 microg/g-wet weight in group SC and 7.62 +/- 1.86 microg/g-wet weight in group SD (p < 0.01); 15.6 +/- 2.61 microg/g wet weight in group RC and 8.44 +/- 1.44 microg/g-wet weight in group RD (p < 0.01). In addition, histopathological examination showed normal gastric structure in groups SC and RC, but there were mild erosive and hemorrhagic changes in groups SD and RD. Because prophylactic ranitidine significantly decreased gastric acid secretion, but did not prevent harmful histopathologic effects in FD, it is suggested that gastric damage cannot be avoided by decreasing gastric acid secretion alone. However PGE2 analogs with or without H2 receptor blockers may have a potential role to prevent FD-related gastric damage. PMID- 10535613 TI - Amniotic fluid volume and onset of labor in physiological pregnancy. AB - We measure, by means of ultrasound, the amniotic fluid volume (expressed as maximal vertical pocket or MVP) in 646 normal pregnancies at the 39th gestational week. Our aim is to evaluate the possible correlation between MVP and onset of the labor. In a 2-week follow-up, the onset of the labor is considered the "event" variable in a time-dependent statistical analysis. Univariate analysis (Kaplan-Meier algorithm) describes a different trend in predicting the onset of labor when a stratification of MVP < 50 and > or = 50 mm was performed (chi2 = 7.91 p < 0.0049 with 1 df, Breslow-Gehan test). The first category was comprised of 496 fetuses with a median (min-max) MVP of 39 mm (25-49), the second category of 150 fetuses with a MVP of 57 mm (50-100). The results suggest that lower levels of MVP are associated to a higher percentage of the onset of labor. Furthermore, in our measurement, performed at the 39th gestational week, the correlation with the events is higher within the 40th gestational week. In fact, at 7 days from the amniotic fluid measurement, the onset of labor and the subsequent delivery is observed in 80.65 and 73.00% of the cases when they are stratified according to MVP < 50 and > or = 50 mm. At the end of the follow-up, instead, the percentage of "events" is similar, 88.10 and 86.67%, respectively. Adjustment for covariates (Cox analysis), as well as maternal age, neonatal weight, and obstetrics history, show an odds ratio (95% C.I.) of 2.08 (1.61-2.69) for MVP, using the above cutoff level. In physiological pregnancy, lower levels of amniotic fluid at term correlated to a higher probability of the onset of labor. PMID- 10535614 TI - Factor VII deficiency detected in pregnancy: a case report. AB - Factor VII deficiency is a rare hereditary coagulation disorder with an incidence estimated at 1 in 500,000 individuals. In this report, we describe the 13th case in pregnancy. The diagnosis of severe factor VII deficiency (factor VII level <5%) was established at 10 weeks' gestation after initial laboratory testing showed a markedly prolonged prothrombin time and a normal activated partial thromboplastin time. There was a history of two preterm deliveries, but there was no evidence of previous bleeding manifestations. Antenatal progress of the index pregnancy was unremarkable. Prophylactic treatment with fresh frozen plasma was started at the onset of labor and the patient had a vaginal delivery of a live girl at 36 weeks' gestation. There was no postpartum hemorrhage and mother and newborn were discharged in good condition. The patient's postpartum level of factor VII remained undetectable. Two aspects are outlined: the absence of any significant increase in factor VII clotting activity during this pregnancy and the need to give replacement therapy at labor in patients with severe factor VII deficiency to decrease the risk of postpartum hemorrhage. PMID- 10535615 TI - Vanishing forceps delivery. AB - This study evaluates the effect of decreasing cesarean rates and increasing regional anesthesia use on the frequency of forceps deliveries. Data of women who delivered at our community hospital from 1990 through 1997 are reviewed. In 1994, the members of our department adopted several strategies to decrease cesarean deliveries. The cesarean rate decreased whereas regional analgesia use increased. We studied the frequency and type of vaginal operative deliveries during this 8 year period. These data were evaluated by chi2 analysis. Data of women who delivered in the first 4 years (group 1) were compared with data of those who delivered in the second 4 years (group 2). A p < 0.05 was considered significant. The demographic and clinical characteristics of these women remained unchanged during the study period. The total cesarean rate decreased from 23.2% in group 1 to 17.9% in group 2 (p < 0.0001). The proportion of women who received regional anesthesia increased from 18.8 in group 1 to 25.7 in group 2 (p < 0.0001). Vaginal operative deliveries increased from 3.6 to 5.5 (p < 0.0001), whereas the proportion of forceps deliveries decreased from 2.2 to 1.5 (p = 0.001). Perinatal morbidity and mortality did not change. The decrease in cesarean rate and increase in regional anesthesia use were associated with an increase in operative deliveries; however, forceps deliveries continue to decrease in our community hospital. PMID- 10535616 TI - Psychobiological markers of stress in pregnancy: 6-sulfatoxymelatonin--a longitudinal study. AB - Maternal stress, physical and psychological, has been associated with adverse pregnancy outcome. The pineal gland is a physiological transducer that reflects adrenergic input. In a recent pilot study, we found urinary 6-sulfatoxymelatonin, the melatonin metabolite, to be elevated after a women spent a day at work compared to levels after a day off work, a leisure day. To evaluate the value of melatonin as a marker of stress, we evaluate melatonin metabolite levels in 121 women, along with perceived anxiety levels and urinary cortisol. Urinary cortisol and maternal anxiety levels each were significantly higher after a work day compared to a leisure day p = .03 and p = .001, respectively. 6 Sulfatoxymelatonin was not significantly different between work and leisure. Changes in cortisol levels were correlated with changes in melatonin metabolite levels (r = .62, p = .001). There was no correlation between changes in anxiety between work and leisure and changes in 6-sulfastoxymelatonin. We found no correlation with 28 week 6-sulfatoxymelatonin or 28-week cortisol and birth weight or gestational age at delivery. Results of this study suggest that melatonin secretion may not be a valuable marker for stress in pregnancy. PMID- 10535617 TI - Congenital anomalies associated with esophageal atresia: Saudi experience. AB - Eighty-nine Saudi newborns with esophageal atresia/tracheoesophageal fistula (EA/TEF) were managed at King Faisal Specialist Hospital and Research Center (KFSH & RC), Riyadh, Saudi Arabia between the years 1980-1995; there were 54 boys and 35 girls. Forty-four (49%) newborns had associated congenital anomalies. Genitourinary anomalies were present in 19 (21%), cardiovascular in 17 (19%), gastrointestinal in 9 (10%), central nervous system in 8 (9%), musculoskeletal in 7 (8%), chromosomal anomalies in 4 (5%), and head and neck in 5 (6%) cases. In general, the survival rate was higher in patients without associated anomalies than with associated anomalies (93 vs. 77%, p = 0.028). However, all patients with head and neck anomalies survived, whereas all patients with chromosomal anomalies died. With these exceptions, the survival rate was similar regardless of the type or the number of associated anomalies. The average birth weight was similar between survivors and non-survivors (2572 vs. 2376 g) and between patients with or without associated anomalies (2566 vs. 2519 g). We conclude that the survival rate of newborns with EA/TEF is high, especially in the absence of associated anomalies. Investigations for possible associated genitourinary and cardiovascular anomalies should be considered for all patients with EA/TEF. PMID- 10535618 TI - EEG abnormalities in survivors of neonatal ECMO: its role as a predictor of neurodevelopmental outcome. AB - The incidence and site of electroencephalogram (EEG) abnormalities and the efficacy of post-ECMO EEG as a predictor of neurodevelopmental outcome was evaluated in survivors of neonatal extracorporeal membrane oxygenation (ECMO). All survivors of neonatal ECMO with an EEG performed prior to their discharge were included if they had at least 12 months of follow-up. The neurodevelopmental outcome was reported as normal, suspect, and abnormal on the basis of neurological examination and the scores on Bayley Scales of Infant Development or McCarthy Scale of Children's Abilities. EEG abnormalities were noted in 31 (70%) of 44 infants. The distribution of EEG abnormalities was not significantly different for right and left hemispheres. The incidence of abnormal neurodevelopmental outcome was similar in infants with a normal or an abnormal EEG (3 of 13 vs. 7 of 31; p = 0.8). EEG abnormalities had no correlation with neurodevelopmental outcome. We conclude that the high incidence of EEG abnormalities and their lack of correlation with neurodevelopmental outcome would suggest that these abnormalities do not represent permanent brain injury and a single EEG performed after decannulation from ECMO is not helpful in identifying infants at risk of subsequent abnormal neurodevelopmental outcome. PMID- 10535619 TI - Congenital orbital teratoma. AB - A case of congenital orbital teratoma is described in which there was no organized eye only microscopic evidence of ocular tissues within the disorganized teratoma. A baby boy presented at birth with a 10-x-8-cm mass extruding from the left orbit. Magnetic resonance imaging (MRI) showed a mixed cystic-solid orbital mass containing areas of calcification and deforming the bony orbit around its margins. There was no organized eye and no intracranial extension. The eye was removed with reconstruction of the eyelids. Histopathology showed representation from all three germ cell layers consistent with a teratoma. There was no organized eye, but some disorganized ocular structures within the teratoma. Follow-up has been uneventful. Neonatologists and pediatricians should be aware of the possible diagnoses in a newborn presenting with an orbital mass, so that early definitive surgery can be performed with preservation of the globe where possible. PMID- 10535620 TI - Glycosylated hemoglobin as a predictor of fetal pulmonic maturity in insulin dependent diabetes at term. AB - In insulin dependent diabetic (IDDM) gestations, fetal pulmonary maturity is delayed in the presence of suboptimal glycemic control. Serum glycosylated hemoglobin (HbA1c) provides a means of assessing glycemic control. We evaluated maternal HbA1c in IDDM pregnancies at term undergoing amniocentesis for lung maturity to establish if euglycemia is associated with improved fetal lung maturity. Between July 1995 and June 1996, IDDM patients undergoing amniocentesis at term for lung maturity studies had a maternal serum sample analyzed for HbA1c. Fetal lung maturity was established by the presence of phosphatidylglycerol (PG) in amniotic fluid. HbA1c was considered elevated if >6.2%. Mean HbA1c level was 6.8% (range 4.4 to 9.9%). PG was present in 54% of patients with elevated HbA1c (7/13) versus 80% of those with normal HbA1c (8/10) (p = 0.4). Although birth weight was higher in the elevated than in the normal HbA1c group (3770 +/- 514 vs. 3215 +/- 610 g), no association was present between birth weight and HbA1c level (r = 0.22, p = 0.4). The rate of a mature pulmonic profile at term is not significantly different between IDDM women with good or poor glycemic control. HbA1c values should not be used to predict the presence or absence of amniotic fluid PG. PMID- 10535621 TI - Medical physics graduate programs should focus on education and research and leave clinical training to residencies. PMID- 10535622 TI - Permanent prostate seed implant brachytherapy: report of the American Association of Physicists in Medicine Task Group No. 64. AB - There is now considerable evidence to suggest that technical innovations, 3D image-based planning, template guidance, computerized dosimetry analysis and improved quality assurance practice have converged in synergy in modern prostate brachytherapy, which promise to lead to increased tumor control and decreased toxicity. A substantial part of the medical physicist's contribution to this multi-disciplinary modality has a direct impact on the factors that may singly or jointly determine the treatment outcome. It is therefore of paramount importance for the medical physics community to establish a uniform standard of practice for prostate brachytherapy physics, so that the therapeutic potential of the modality can be maximally and consistently realized in the wider healthcare community. A recent survey in the U.S. for prostate brachytherapy revealed alarming variance in the pattern of practice in physics and dosimetry, particularly in regard to dose calculation, seed assay and time/method of postimplant imaging. Because of the large number of start-up programs at this time, it is essential that the roles and responsibilities of the medical physicist be clearly defined, consistent with the pivotal nature of the clinical physics component in assuring the ultimate success of prostate brachytherapy. It was against this background that the Radiation Therapy Committee of the American Association of Physicists in Medicine formed Task Group No. 64, which was charged (1) to review the current techniques in prostate seed implant brachytherapy, (2) to summarize the present knowledge in treatment planning, dose specification and reporting, (3) to recommend practical guidelines for the clinical medical physicist, and (4) to identify issues for future investigation. PMID- 10535623 TI - The roles of multileaf collimators and micro-multileaf collimators in conformal and conventional nasopharyngeal carcinoma radiotherapy treatments. AB - The purpose of this work is to study the efficacy and limitations of using standard multileaf collimators (MLCs) and micro-multileaf collimators (mMLCs) in the treatment of nasopharyngeal carcinoma (NPC) by conventional and conformal radiotherapy techniques. The penumbra characteristics of MLC, mMLC, and customized block collimated beams are measured with respect to leaf edge angle, beam energy, treatment depth, and field size and compared with those generated by a commercial three-dimensional planning computer system. Upon verification of the planning system, it is used to evaluate the treatment plans generated with these beam shapers for conventional and conformal NPC treatments. The effective penumbra of a MLC beam is strongly influenced by its edge angle, leaf width, and treatment depth. The suitability of standard MLCs in conventional NPC treatments is determined mainly by the edge angle to be used. For conformal NPC treatments involving six or more fields, dose volume histograms comparable to those of customized beam blocks are obtained with a standard MLC. The mMLC does not have the same restrictions as those on standard MLC but is limited to phase II treatment by its small usable field size. Both standard MLCs and mMLCs can be used to replace customized divergent beam blocks in both conventional and conformal NPC treatments. However, a MLC, due to its larger effective penumbra, may be unsuitable for use in cases when the tumor volumes extend very close to the critical normal structures. A mMLC, on the other hand, is limited by its small maximum field size and can only be used for collimating the facial portals in the second phase treatment. PMID- 10535624 TI - A comparison of beam characteristics for gated and nongated clinical x-ray beams. AB - Respiratory gating has only recently been applied to conventional external beam radiotherapy. In order for respiratory gating to be used clinically, an evaluation of the dosimetric effects of small units of delivered dose must be performed. The purpose of this study is to systematically evaluate the effect of various gating sequences on x-ray central axis output, ionization ratios (nominal accelerating potential), beam flatness, and beam symmetry. Measurements were taken for 6 and 18 MV photons on a linear accelerator that generates the gate by using a gridded electron gun to stop the electron flow to the wave-guide. The beam output, energy, flatness, and symmetry did not vary by more than 0.8 percent in most of the gating sequences. The maximum output deviations (0.8 percent), flatness deviations (1.9 percent), and symmetry deviations (0.8 percent) occurred when a low number of monitor units (<5 MU) were delivered in the gating window. Although these deviations are not clinically significant, each linear accelerator should be evaluated carefully before clinical implementation. PMID- 10535625 TI - Changes in incident photon fluence of 6 and 18 MV x rays caused by blocks and block trays. AB - When a block and tray are placed in a x-ray beam the dose to a point in a phantom is changed by the following factors: (1) attenuation of photon and electron fluence from the head of the accelerator by the tray and the block, (2) decrease in the scatter in the phantom by a reduction in the phantom volume that receives radiation, and (3) generation of scatter off the tray and block. This third factor is generally ignored in dosimetry calculation but has been measured in this work. Measurements of incident photon fluence for 6 and 18 MV x rays were made with a columnar miniphantom of 10 cm depth. The tray factor for a 9 mm thick Lexan tray is found to be variable and to increase by 1.8% due to scatter off the tray when the field size is increased from a 3cm x 3 cm to 40cm x 40 cm field. Also, it was found that scatter off a block could increase the incident photon fluence by as much as 2%. The magnitude of this block scatter depends on the length of the inner edge of the opening in the block and on amount of block that is being irradiated, the overlap of the block by the radiation field. The total block-tray factor can be as much as 3% larger than the single-value tray factor measured with a 10cm x 10cm field that is traditionally used. An analytical equation is developed that accurately models the block-tray factor. PMID- 10535626 TI - Effect of electron contamination on scatter correction factors for photon beam dosimetry. AB - Physical quantities for use in megavoltage photon beam dose calculations which are defined at the depth of maximum absorbed dose are sensitive to electron contamination and are difficult to measure and to calculate. Recently, formalisms have therefore been presented to assess the dose using collimator and phantom scatter correction factors, Sc and Sp, defined at a reference depth of 10 cm. The data can be obtained from measurements at that depth in a miniphantom and in a full scatter phantom. Equations are presented that show the relation between these quantities and corresponding quantities obtained from measurements at the depth of the dose maximum. It is shown that conversion of Sc and Sp determined at a 10 cm depth to quantities defined at the dose maximum such as (normalized) peak scatter factor, (normalized) tissue-air ratio, and vice versa is not possible without quantitative knowledge of the electron contamination. The difference in Sc at dmax resulting from this electron contamination compared with Sc values obtained at a depth of 10 cm in a miniphantom has been determined as a multiplication factor, Scel, for a number of photon beams of different accelerator types. It is shown that Scel may vary up to 5%. Because in the new formalisms output factors are defined at a reference depth of 10 cm, they do not require Scel data. The use of Sc and Sp values, defined at a 10 cm depth, combined with relative depth-dose data or tissue-phantom ratios is therefore recommended. For a transition period the use of the equations provided in this article and Scel data might be required, for instance, if treatment planning systems apply Sc data normalized at d(max). PMID- 10535627 TI - Measurement of the collection efficiency of a large volume spherical ionization chamber in megavoltage therapy beams. AB - The collection efficiency of a 5.7 cm diameter spherical ionization chamber has been measured in 4 MV and 10 MV x-ray beams at various distances from the source. This chamber was found to have a substantial inefficiency due to its large volume and the high dose rate and pulsed nature of the therapy beams. It was also found that the efficiency depended on the dose rate of the machine because the inter pulse separation time of the linac is significantly less than the ion transit time for this chamber. Thus, ionization from more than one beam pulse is collected by the chamber at the same time. The efficiency was determined using three techniques (i) the two-voltage technique, (ii) the voltage extrapolation technique and (iii) a method originally devised for determining the collection efficiency of large volume ionization chambers in diagnostic radiology. The results show that methods (ii) and (iii) agree well, but that the two-voltage technique predicts a much lower efficiency. At about 4 m from the source, the collection efficiency for this chamber varied between 98% and 97% for dose rates between 50 and 250 cGy/min for 4 MV and between 97% and 90% for dose rates between 100 and 600 cGy/min for 10 MV. At isocenter, the comparable figures were 78% and 56% respectively for 4 MV and 65% and 38% respectively for 10 MV. PMID- 10535628 TI - Monte Carlo simulations of the imaging performance of metal plate/phosphor screens used in radiotherapy. AB - The imaging performance of metal plate/phosphor screens which are used for the creation of portal images in radiotherapy is investigated by using Monte Carlo simulations. To this end the modulation transfer function, the noise power spectrum and the detective quantum efficiency [DQE(f)] are calculated for different metals and phosphors and different thicknesses of metal and phosphor for a range of spatial resolutions. The interaction of x-rays with the metal plate/phosphor screen is modeled with the EGS4 electron gamma shower code. Optical transport in the phosphor is modeled by simulating scattering and reabsorption events of individual optical photons. It is shown that metals with a high atomic number perform better than lighter metals in maximizing the DQE(f). It is furthermore shown that the DQE(f) for the metal plate/phosphor screen alone is nearly x-ray quantum absorption limited up to spatial frequencies of 0.4 cycles/mm. In addition, it is argued that the secondary quantum sink of optical photons imposed by the optical chain (mirror, lenses and video camera) leads to a significant degradation of the signal-to-noise ratio at spatial frequencies which are most important for successful registration of portal images. Therefore, the conclusion is that a replacement of the optical chain by a flat array of photodiodes placed directly under the phosphor will lead to a substantial improvement in image quality of portal images. PMID- 10535629 TI - A system for three-dimensional dosimetric verification of treatment plans in intensity-modulated radiotherapy with heavy ions. AB - The introduction of dynamic intensity modulation into radiotherapy using conventional photon beams or scanning particle beams requires additional and efficient methods of dose verification. Dose measurements in dynamically generated dose distributions with a single ionization chamber require a complete application of the treatment field for each single measurement. Therefore measurements are performed by simultaneous use of multiple ionization chambers. The measurement is performed by a computer controlled system and is comprised of the following steps: (a) automated positioning of the ionization chambers, (b) measurement at these points, (c) a comparison with the calculated dose from the treatment planning system, and (d) documentation of the measurement. The ionization chambers are read out by a multichannel electrometer and are densely packed into a mounting of polymethylmetacrylate, which is attached to the arm of a three-dimensional motor-driven water phantom. The measured and planned dose values are displayed numerically as well as graphically. The mean deviation between measured and planned doses as well as their standard deviation are calculated and displayed. Through printouts complete documentation of the measurement is obtained and a quick decision can be made whether the dose distribution is acceptable for the patient. The system is now routinely used for dose verification at the heavy ion therapy project at the Gesellschaft fur Schwerionenforschung in Darmstadt. Up to now 242 measurements have been performed for heavy ion treatment of 30 patients. The system allows efficient verification and documentation of carbon ion fields and is in principle also applicable to intensity-modulated photon beams. PMID- 10535630 TI - Clinical implementation of a Monte Carlo treatment planning system. AB - The purpose of this study was to implement the Monte Carlo method for clinical radiotherapy dose calculations. We used the EGS4/BEAM code to obtain the phase space data for 6-20 MeV electron beams and 4, 6, and 15 MV photon beams for Varian Clinac 1800, 2100C, and 2300CD accelerators. A multiple-source model was used to reconstruct the phase-space data for both electron and photon beams, which retained the accuracy of the Monte Carlo beam data. The multiple-source model reduced the phase-space data storage requirement by a factor of 1000 and the accelerator simulation time by a factor of 10 or more. Agreement within 2% was achieved between the Monte Carlo calculations and measurements of the dose distributions in homogeneous and heterogeneous phantoms for various field sizes, source-surface distances, and beam modulations. The Monte Carlo calculated electron output factors were within 2% of the measured values for various treatment fields while the heterogeneity correction factors for various lung and bone phantoms were within 1% for photon beams and within 2% for electron beams. The EGS4/DOSXYZ Monte Carlo code was used for phantom and patient dose calculations. The results were compared to the dose distributions produced by a conventional treatment planning system and an intensity-modulated radiotherapy inverse-planning system. Significant differences (>5% in dose and >5 mm shift in isodose lines) were found between Monte Carlo calculations and the analytical calculations implemented in the commercial systems. Treatment sites showing the largest dose differences were for head and neck, lung, and breast cases. PMID- 10535631 TI - The use of radiographic film for linear accelerator stereotactic radiosurgical dosimetry. AB - The measurement of stereotactic radiosurgical dose distributions requires an integrating, high-resolution dosimeter capable of providing a spatial map of absorbed dose. Although radiographic film is an accessible dosimeter fulfilling these criteria, for larger radiotherapy photon fields the sensitivity of film emulsion exhibits significant dependencies on both depth in phantom and field size. We have examined the variation of film sensitivity over the ranges of depths and field sizes of interest in radiosurgery with a 6 MV photon beam. While for large (20cm x 20cm) fields the potential error in dose due to the variation of the film response with depth reaches 15%, the corresponding maximum error for a 2.5 cm diameter radiosurgical beam is 1.5%. This uncertainty was observed to be comparable in magnitude to that produced by variation in processing conditions (1.1%) and by varying the orientation of the film plane relative to the beam central axis (1.5%). The dependence of emulsion sensitivity on field size has been observed to be negligible for fields ranging in diameter from 1.0 cm to 4.0 cm. The source of the dependence of film sensitivity has been illustrated by using an EGS4 Monte Carlo simulation for large fields to illustrate significant increases in the photon spectrum below 400 keV with depth in phantom. In contrast, relative increase of this low-energy component is negligible for radiosurgical photon fields. PMID- 10535632 TI - Morphology-guided radiosurgery treatment planning and optimization for multiple isocenters. AB - This work merges two distinct fields, 3D morphology and ionizing radiation dosimetry, to solve the problem of 3D-treatment planning and optimization in stereotactic radiosurgery. In Leksell Gamma Knife radiosurgery, dose delivery is based on the unit "shot," a dose distribution approximately spherical in shape. Multiple shots, or isocenters, are used in Gamma Knife treatment to deliver a conformal dose to an irregular radiosurgical target. The medial axis transformation, or skeleton, of the target, which uniquely characterizes the target volume and shape, is used to determine the optimal shot positions (isocenters), sizes (collimator helmet size and dosimetric weight), and the total number of shots that will deliver a conformal dose distribution to the target. The skeletonization approach reduces a complicated 3D-optimization problem to 1D searching with potential savings in computation time and mathematical complexity. In addition, optimization based on target shape replicates and automates manual treatment planning. This approach makes the process easily understandable. The relationship between skeleton discs and the dose distributions they predict is discussed. Results of optimal plans and corresponding dose distributions are presented. This approach is generally applicable to other types of multi isocentric stereotactic radiosurgery techniques. PMID- 10535633 TI - Treatment head design for multileaf collimated high-energy electrons. AB - This paper describes how a conventional treatment head can be modified for use of multileaf collimated electron beams. Automatic and dynamic beam delivery are possible for both electrons and photons by using the computer controlled multileaf collimator (MLC) for both photon and electron beams. Thereby, the electron beams can be mixed more freely into the treatment to take advantage of the specific depth modulation characteristics of electrons. The investigation was based on Monte Carlo calculations using the software package BEAM. The physical parameters used in this optimization were the beam penumbra and the virtual/effective point source position. These parameters are essential for shaping beams, beam matching and for dosimetry calculations. The optimization was carried out by modifying a number of parameters: replacing the air atmosphere in the treatment head with helium, adding a helium bag below the MLC, changing the position of the scattering foils, modifying the monitor chamber, and adjusting the position of the MLC. The beam characteristics for some of these designs were found to fulfil our criteria for clinically useful beams down to at least 9 MeV. PMID- 10535634 TI - Quantitative angiographic blood-flow measurement using pulsed intra-arterial injection. AB - A technique for quantitative blood-flow measurement using a novel pulsed injection of radiographic contrast agent is reported. A pressurized source of contrast agent is interrupted by a rotary valve at rates ranging from 1 to 30 Hz, producing well-defined boli at the end of a catheter. The position of these boli can be recorded by a digital radiographic system and analyzed by one of several previously reported techniques, to produce quantitative measurements of blood velocity and flow rate throughout the cardiac cycle. The contrast-agent flow wave form produced by the pulsed injector has been measured with an electromagnetic flow meter, for driving pressures ranging from 600 to 1500 kPa. Excellent modulation of the contrast agent is observed for injection frequencies up to 20 Hz, through catheters up to 100 cm in length. Preliminary in vitro angiographic flow measurements have been performed using an x-ray image intensifier, coupled to a linear photodiode array as the digital detector. Both constant flow and pulsatile human blood-flow wave forms were simulated within a 6.4-mm-diam straight tube and monitored with an electromagnetic flow meter. These experiments indicate that the pulsed injector can be used to provide estimates of arterial blood flow over the entire cardiac cycle (including reverse flow), to within about +/-11%, following injection of less than 10 ml of iodinated contrast agent. PMID- 10535635 TI - Feature selection with limited datasets. AB - Computer-aided diagnosis has the potential of increasing diagnostic accuracy by providing a second reading to radiologists. In many computerized schemes, numerous features can be extracted to describe suspect image regions. A subset of these features is then employed in a data classifier to determine whether the suspect region is abnormal or normal. Different subsets of features will, in general, result in different classification performances. A feature selection method is often used to determine an "optimal" subset of features to use with a particular classifier. A classifier performance measure (such as the area under the receiver operating characteristic curve) must be incorporated into this feature selection process. With limited datasets, however, there is a distribution in the classifier performance measure for a given classifier and subset of features. In this paper, we investigate the variation in the selected subset of "optimal" features as compared with the true optimal subset of features caused by this distribution of classifier performance. We consider examples in which the probability that the optimal subset of features is selected can be analytically computed. We show the dependence of this probability on the dataset sample size, the total number of features from which to select, the number of features selected, and the performance of the true optimal subset. Once a subset of features has been selected, the parameters of the data classifier must be determined. We show that, with limited datasets and/or a large number of features from which to choose, bias is introduced if the classifier parameters are determined using the same data that were employed to select the "optimal" subset of features. PMID- 10535636 TI - Comparison between measured and predicted attenuation curves of x-ray beams. AB - Measurements of the attenuation curves of low energy x-ray beams are used in this work as a functional description of such beams. Measurements of the transmission curves through aluminum made with a low volume ionization chamber have been used to predict transmission curves through copper. The results are compared with the data of attenuation in copper measured with the above-mentioned ionization chamber, and also with calculations of attenuation in copper made from reference x-ray spectra and tabulated values of the mass attenuation coefficients of copper. PMID- 10535637 TI - Refraction-enhanced x-ray imaging of mouse lung using synchrotron radiation source. AB - The low divergent x-ray beam from the third-generation synchrotron radiation source such as SPring-8 enables us to observe refraction of x rays that is in the range of microradians. Under an experimental condition for which ray optics is a good approximation, we found that the refraction produces a high-contrast projection image of a mouse when it was recorded at 6.5 m behind the specimen. Especially, the lung is visualized far better than with the conventional imaging which utilizes absorption of x rays. This is a promising new technique for the diagnosis of diseases in the human lung with a low radiation dose. PMID- 10535638 TI - A synchrotron radiation microtomography system for the analysis of trabecular bone samples. AB - X-ray computed microtomography is particularly well suited for studying trabecular bone architecture, which requires three-dimensional (3-D) images with high spatial resolution. For this purpose, we describe a three-dimensional computed microtomography (microCT) system using synchrotron radiation, developed at ESRF. Since synchrotron radiation provides a monochromatic and high photon flux x-ray beam, it allows high resolution and a high signal-to-noise ratio imaging. The principle of the system is based on truly three-dimensional parallel tomographic acquisition. It uses a two-dimensional (2-D) CCD-based detector to record 2-D radiographs of the transmitted beam through the sample under different angles of view. The 3-D tomographic reconstruction, performed by an exact 3-D filtered backprojection algorithm, yields 3-D images with cubic voxels. The spatial resolution of the detector was experimentally measured. For the application to bone investigation, the voxel size was set to 6.65 microm, and the experimental spatial resolution was found to be 11 microm. The reconstructed linear attenuation coefficient was calibrated from hydroxyapatite phantoms. Image processing tools are being developed to extract structural parameters quantifying trabecular bone architecture from the 3-D microCT images. First results on human trabecular bone samples are presented. PMID- 10535639 TI - Penicilliosis marneffei--West meets East. PMID- 10535640 TI - Expression in Escherichia coli of flaB, the gene coding for a periplasmic flagellin of Leptospira interrogans serovar pomona. AB - A periplasmic flagellin gene, flaB, of Leptospira interrogans serovar pomona (strain Pomona) was expressed in Escherichia coli for the production and antigenic characterisation of the protein. The flaB structural gene, which was previously cloned into pUC118, was derived by PCR from the recombinant plasmid and used to generate an expression construct with the trc promoter-driven pProEx HT system. Under the conditions employed, the flaB was expressed as inclusion bodies formed within E. coli, yielding c. 120 mg of the recombinant protein/L of culture. A polyhistidine tag introduced at the amino-acid terminus of the FlaB protein allowed for the purification of the protein by nickel-chelate affinity chromatography. The expressed protein reacted with both mouse and bovine antisera to L. interrogans on Western blots, indicating that it could be of use in the diagnosis of leptospirosis. The recombinant leptospiral flagellin may also be of value in studying its role in the pathogenesis of leptospirosis. PMID- 10535641 TI - Virulence properties of type VII Streptococcus agalactiae (group B streptococci) and immunochemical analysis of capsular type polysaccharide. AB - Strains of a new polysaccharide type of group B streptococci (GBS), type VII, have been isolated from human carriers and invasive infections. Some of these strains bear the protein antigen c or R, as do other GBS serotypes. The capsular type polysaccharide is sialylated and this residue is involved in the immunodeterminant structure. All type VII strains examined were virulent in CD-1 mice; the LD50 after intraperitoneal (i.p.) challenge was 4.57 (SD 0.12) x10(7) cfu for the reference strain and 5.49 (SD 1.5) x10(7) cfu for clinical isolates. A particular feature of this serotype was the ability to induce septic arthritis not only when injected intravenously (i.v.), but also when injected i.p. Rabbit antiserum against the capsular type VII polysaccharide exhibited opsonic activity in a phagocytosis assay and protective activity against infection. PMID- 10535642 TI - Fine structural characterisation of a Rickettsia-like organism in human platelets from patients with symptoms of ehrlichiosis. AB - Since 1982, Ehrlichia platys infection has been diagnosed in canines from Venezuela by the use of buffy coat smears. In 1992, ehrlichia-like bodies were observed in platelets from a severely ill girl by light microscopy. The patient was seropositive to E. chaffeensis by the indirect fluorescent antibody test (IFAT). Tetracycline was administered and the patient recovered. More than 400 cases with such intra-platelet organisms have been studied at this laboratory over the past 6 years, and all the patients had a good response to the treatment. To determine whether the organisms in human blood platelets were truly platelet ehrlichiae, IFAT and transmission electron microscopy (TEM) studies were undertaken in four patients. Light microscopic examination of blood samples revealed the dense organism inside platelets, and a great reactivity of the blood cells. Sera from the four patients were seronegative against E. chaffeensis and E. platys antigens. Three of four samples contained the intra-platelet organisms when examined by TEM. Electron microscopy showed platelets with vacuoles containing pleomorphic organisms. These organisms had a thickened membrane, an electron-translucent inner area and an electron-dense granular component in the periphery. An abundant electron-dense material was observed surrounding them. The ultrastructure of such micro-organisms has not been reported previously, Based on the similarity of many of their characteristics with rickettsiae, we suggest that the microorganisms found in the present study might belong to the family Rickettsiaceae. PMID- 10535643 TI - Expression of Bacteroides fragilis virulence markers in vitro. AB - Bacteroides fragilis isolates from intestinal and non-intestinal infections, normal flora and the environment were examined for properties linked with interactions among cells in vitro. Different adhesion molecules were detected in agglutination assays with human erythrocytes and tests for auto-agglutination and adherence to human colon carcinoma cells (HT29). There was no correlation between these properties, indicating that independent molecules are involved. Treatment with trypsin, heat or EDTA inhibited agglutination and adherence, suggesting that these molecules are proteins. The lack of correlation with the origin of the strains did not permit any of these activities to be recognised as virulence markers. The expression of fragilysin, a protease associated with damage to intestinal cells and bacterial translocation, was examined. Only those strains from patients with diarrhoea expressed this protease activity in assays with HT29 cells and this was confirmed by specific PCR for the bft gene. The activity of fragilysin as an enterotoxin was confirmed in the rabbit intestinal ligated loop assay. The association of this property only with strains from intestinal infections indicates that it is too early to suggest this protease as a determinant factor of B. fragilis invasiveness. PMID- 10535644 TI - Factors affecting the course and severity of transnasally induced Staphylococcus aureus pneumonia in mice. AB - In order to examine several factors that may affect the course and severity of transnasally induced Staphylococcus aureus pneumonia in mice, bacteria were prepared in a free suspension or bound to fetal mouse cells. Immunosuppression was induced in five strains of mice (ICR, C57BL/6, BALB/c, C3H/He and CBA/J) by injection of cyclophosphamide (200 mg/kg body weight), 2 days before infection. Impairment of mucociliary clearance was induced by intranasal instillation of formalin. Mice were then infected with various doses and strains of the organism. Although no significant differences were observed between either form of inoculum, pretreatment with formalin plus cyclophosphamide was associated with a significant increase in lung bacterial counts. In particular, cyclophosphamide treatment was associated with a high mortality in mice infected with several strains of S. aureus irrespective of their toxin production profiles. Histopathological examination demonstrated that in mice treated with formalin plus cyclophosphamide, clusters of bacteria were observed in lung parenchyma, associated with a mild accumulation of inflammatory cells at day 2 and extensive cell infiltration at day 7. CBA/J mice represented the most susceptible strain among those examined, with 10(4)- and 10(2)-fold higher bacterial counts in the lungs at days 3 and 5, respectively. These results indicate that neutropenia and impaired mucociliary clearance are major factors that influence the severity of S. aureus pneumonia in mice. Analysis of the role of genetic background in enhancement of vulnerability to infection is warranted in future studies. PMID- 10535645 TI - Use of PCR to identify enteroaggregative Escherichia coli as an important cause of acute diarrhoea among children living in Calcutta, India. AB - The importance of enteroaggregative Escherichia coli (EAggEC) as a possible aetiological agent of acute diarrhoea among children in Calcutta, India, was investigated. Simultaneously the use of a previously described PCR diagnostic system was assessed for its ability to identify EAggEC infection. E. coli strains isolated during a 1-year case-control study from faecal samples of 388 children aged <5 years, with or without diarrhoea, were examined for EAggEC by HeLa cell adherence assay in parallel with a PCR assay with primers generated from an EAggEC DNA probe. A blind comparison was made between the two methods to determine their diagnostic potential. E. coli isolates that adhered to HeLa cells in an aggregative pattern were the sole isolates significantly more often in 254 cases (9%) than in 134 control (2%) children. Age stratification showed that EAggEC were isolated more frequently from children aged <36 months. The EAggEC isolates belonged to several O serogroups and showed multiple drug resistance. Both methods were positive for 26 samples, nine samples were positive by PCR alone and seven samples were positive by culture alone, thus indicating a 78% sensitivity and 97% specificity for the PCR assay. EAggEC is an important aetiological agent of acute diarrhoea among infants in and around Calcutta, and the PCR diagnostic system may be useful to identify such infection in epidemiological studies. PMID- 10535646 TI - Direct detection of Prevotella intermedia and P. nigrescens in suppurative oral infection by amplification of 16S rRNA gene. AB - A specific 16S rDNA PCR and subsequent hybridisation reaction was designed to discriminate between strains of Prevotella intermedia (n = 15) and P. nigrescens (n = 15). This technique was then used to detect the presence of these two bacterial species in acute suppurative oral infection. A total of 36 pus samples aspirated from 26 peri-apical abscesses, three root canals, three periodontal abscesses, two cases of refractory periodontitis, one cyst and one haematoma was examined. A portion of the pus sample was processed by PCR and the remainder of the specimen was subjected to routine culture. The PCR-based technique gave an identical pattern of detection of P. intermedia or P. nigrescens to that obtained by culture for 30 of the 36 specimens. Either P. intermedia or P. nigrescens was present in 14 samples and neither species was detected in 16 samples. In the remaining six samples the PCR method indicated the presence of one (n = 3) or both (n = 3) of the Prevotella species but neither or only one species was isolated by culture. It is concluded that the presence of P. intermedia and P. nigrescens in pus can be detected rapidly and specifically by direct PCR amplification of 16S rDNA. P. nigrescens was detected more frequently than P. intermedia in suppurative peri-apical infection both by culture and PCR. PMID- 10535647 TI - Expression of beta-lactamases in Yersinia enterocolitica strains of biovars 2, 4 and 5. AB - Characteristic patterns of susceptibility to beta-lactam antibiotics are associated with different biovars of Yersinia enterocolitica. To elucidate the basis for these differences, the beta-lactamases of strains of Y. enterocolitica biovars 4 (n = 63), 2 (n = 12) and 5 (n = 10) were characterised. PCR fragments were generated from the beta-lactamase A (blaA) and B (blaB) genes; in addition, beta-lactamase induction tests were performed with imipenem as the inducer and beta-lactamase inhibition assays were undertaken with aztreonam and clavulanic acid. All the strains yielded PCR amplification fragments with primers to blaA and blaB. Biovar 4 strains had uniform patterns of beta-lactamase induction and inhibition: uninduced biovar 4 strains predominantly expressed BlaA, but low level expression of BlaB was also detected; after induction, biovar 4 strains predominantly produced BlaB. Beta-lactamase expression varied between and within biovars 2 and 5: uninduced strains predominantly expressed either BlaA or BlaB, or exclusively BlaB; after induction BlaB was predominantly or exclusively expressed. Both the basal and induced levels of beta-lactamase varied within biovars 2 and 5. Some biovar 5 strains were not inducible; these predominantly produced BlaA. The results of this study show that biovar 2, 4 and 5 strains contain both blaA and blaB, but that the expression of the enzymes is regulated differently between the biovars, and varies within biovars 2 and 5. There was some correlation between antibiogram and the clusters defined from the beta lactamase induction and inhibition tests, but it was not possible to predict beta lactamase expression profiles from MIC data. PMID- 10535648 TI - Comparison of three PCR techniques for detecting cytomegalovirus (CMV) DNA in serum, detection of early antigen fluorescent foci and culture for the diagnosis of CMV infection. AB - Three PCR assays were developed for detection of cytomegalovirus (CMV) DNA in serum and were evaluated with samples from organ transplant recipients. The Qiamp Blood Kit was efficient for extraction of DNA from sera. Single-round PCR of a 293-bp region of CMV DNA was sensitive and highly specific for CMV targets and was more sensitive than detection of early antigen fluorescent foci (DEAFF) testing or isolation of CMV from buffy coat by cell culture. The results of a significant proportion of buffy coat samples were not interpretable because of toxicity in conventional culture or DEAFF tests. A non-competitive quantitative PCR test and semi-quantitative PCR test for the detection of CMV DNA in serum yielded comparable results for samples taken serially from three bone marrow transplant recipients. Single-round PCR was superior to conventional techniques for the diagnosis of CMV infection, was simple to perform and was completed rapidly. The semi-quantitative technique has added advantages where quantification is important. PMID- 10535650 TI - Oral fluid antibody detection in the diagnosis of Helicobacter pylori infection. AB - The aim of this study was to evaluate an enzyme-linked immunosorbent assay (ELISA) for the detection of anti-Helicobacter pylori specific IgG antibodies in specimens of oral fluid. Antral biopsy specimens, serum and oral fluid samples were collected from 81 patients attending for upper gastrointestinal endoscopy. The presence or absence of current H. pylori infection was determined by culture, histology and urease detection. Anti-H. Pylori specific IgG was detected in serum by an established in-house ELISA and in oral fluid by an ELISA developed for this study. In all, 34 (42%) of 81 patients were positive for H. pylori by one or more of the 'gold standard' tests (culture, histology and urease detection). The oral fluid ELISA had a sensitivity of 94% and specificity of 85% with regard to current H. pylori infection. The serum ELISA had a sensitivity and specificity of 91%. There was an overall agreement of 88% between serum and oral fluid antibody detection. The detection of anti-H. pylori specific IgG in oral fluid by ELISA is comparable in sensitivity and specificity with serum-based methods. Oral fluid based ELISA could provide a reliable, non-invasive method for the diagnosis of H. pylori infection, and may be of particular benefit for population surveys. PMID- 10535649 TI - Analysis of Salmonella enterica serotype Typhimurium by phage typing, antimicrobial susceptibility and pulsed-field gel electrophoresis. AB - Three typing methods commonly used for bacteria--phage typing, antimicrobial susceptibility and pulsed-field gel electrophoresis (PFGE)- were used to characterise 64 Salmonella enterica serotype Typhimurium isolates from individual adult patients from Nairobi, Kenya. The isolates encompassed 11 definitive phage types (DTs), which fell into eight PFGE clusters; 31.3% of isolates were either untypable or reacted nonspecifically with the phages used for typing and 26.6% were of DT 56. Plasmids of c. 100 kb were responsible for self-transferable multiresistance among the isolates. Analysis by PFGE and phage type demonstrated that multiresistant Typhimurium strains causing diarrhoea and invasive disease were multiclonal. PMID- 10535651 TI - Automated rubella antibody screening:a cautionary tale. PMID- 10535652 TI - Depressed patients' perceptions of facial emotions in depressed and remitted states are associated with relapse: a longitudinal study. AB - Within the framework of interpersonal and cognitive theories of depression, we investigated whether the perception of facial emotions was associated with subsequent relapse into depression. The 23 inpatients with major depression who remitted (65 admitted patients) were studied at admission (T0), at discharge (T1), and 6 months thereafter to assess relapse. They judged schematic faces with respect to the expression of positive and negative emotions. Six patients (26.1%) relapsed. High levels of perception of negative emotions in faces, either assessed at T0 or at T1, were associated with relapse. Moreover, subjects saw more negative emotions in depressed than in remitted state. Significant results were confined to ambiguous faces, i.e., faces expressing equal amounts of positive and negative emotions. Our data support the hypothesis that a bias toward the perception of others' facial emotions as negative is an enduring vulnerability factor to depression relapse and depressed mood amplifies this negative bias in perception. PMID- 10535653 TI - Effects of lesion variables and emotion type on the perception of facial emotion. AB - The purpose of this study was to consider the effects of valence, motoric direction (i.e., approach/withdrawal), and arousal on the perception of facial emotion in patients with unilateral cortical lesions. We also examined the influence of lesion side, site, and size on emotional perception. Subjects were 30 right-hemisphere-damaged (RHD) and 30 left-hemisphere-damaged (LHD) male patients with focal lesions restricted primarily to the frontal, temporal, or parietal lobe. Patient groups were comparable on demographic and clinical neurological variables. Subjects were tested for their ability to match photographs of four facial emotional expressions: happiness, sadness, fear, and anger. Overall, RHD patients were significantly more impaired than LHD patients in perceiving facial emotion. Lesion side, but not site, was associated with motoric direction and valence dimensions. RHD patients had specific deficits relative to LHD patients in processing negative and withdrawal emotions; there were no group differences for positive/approach emotions. Lesion size was not significantly correlated with accuracy of emotional perception. PMID- 10535654 TI - Subtyping depression: testing algorithms and identification of a tiered model. AB - We seek to distinguish psychotic, melancholic, and nonmelancholic depression by clinical features and to test varying algorithm models to determine optimal criteria sets. We report a study of 269 depressed inpatients and outpatients. A latent class analysis (LCA) of 16 clinical features allowed for specificity or overrepresentation of features to be examined across the three classes. Varying algorithm models for distinguishing melancholic and nonmelancholic depression, involving endogeneity symptoms and observer-rated psychomotor disturbance (PMD) were compared. Psychotic depression was readily distinguished by the specific presence of psychotic features, and PMD was most severe in this class. Melancholic depression was most clearly distinguished from the residual nonmelancholic class by the presence of PMD. Although some endogeneity symptoms were overrepresented in the melancholic class, their specificity was unimpressive. An algorithm involving PMD components alone was highly efficient in discriminating LCA classes and, more importantly, superior to DSM-IV decision rules when examined against a range of clinical validators of melancholia. Subtyping appears assisted by a hierarchical model, based on a small set of features. The move from nonmelancholic to melancholic depression appears defined by a tier of observably rated PMD, whereas the move from melancholic to psychotic depression is determined by a tier of psychotic features and contributed to by significantly higher levels of PMD. PMID- 10535655 TI - Response to sexual assault: a relational perspective. AB - It has been suggested that although the severity of the stressor is the primary determinant of acute posttraumatic stress disorder (PTSD) symptoms, pre-existing personality patterns may be the primary contributors to the development of chronic PTSD symptomatology. The authors postulate that of the multiple personality factors that influence behavior and response to traumatic events, relational capacity or the ability to sustain interpersonal relationships provides an overarching construct for understanding the contribution of social contextual factors to post-trauma response. The results of this exploratory study support the authors' hypothesis that relational capacity is a significant factor in explaining persistent PTSD symptoms in a sample of adult women who have been raped. Significant correlations were found between measures of relational capacity, the Bell Object Relations Inventory and the Inventory of Interpersonal Problems and measures of distress, the Posttraumatic Symptom Scale, and the Beck Depression Inventory. PMID- 10535657 TI - Motivational interviewing and treatment adherence among psychiatric and dually diagnosed patients. AB - The effect of motivational interviewing on outpatient treatment adherence among psychiatric and dually diagnosed inpatients was investigated. Subjects were 121 psychiatric inpatients, 93 (77%) of whom had concomitant substance abuse/dependence disorders, who were randomly assigned to: a) standard treatment (ST), including pharmacotherapy, individual and group psychotherapy, activities therapy, milieu treatment, and discharge planning; or b) ST plus motivational interviewing (ST+MI), which involved 15 minutes of feedback on the results of a motivational assessment early in the hospitalization, and a 1-hour motivational interview just before discharge. Interviewers utilized motivational techniques described in Miller and Rollnick (1991), such as reflective listening, discussion of treatment obstacles, and elicitation of motivational statements. Results indicated that the proportion of patients who attended their first outpatient appointment was significantly higher for the ST+MI group (47%) than for the ST group (21%; chi2 = 8.87, df = 1, p<.01) overall, and for dually diagnosed patients (42% for ST+MI vs. 16% for ST only; chi2 = 7.68, df = 1, p<.01). Therefore, brief motivational interventions show promise in improving outpatient treatment adherence among psychiatric and dually diagnosed patients. PMID- 10535656 TI - Sexual assault history and headache: five general population studies. AB - Headache is a common and disabling symptom. Although it is known that sexual assault is associated with a wide range of physical symptoms, little research has addressed its association with headache. The present study evaluated this association in five independent samples of randomly selected household residents (pooled N = 7502). The weighted mean odds ratio (OR) linking sexual assault and headache, controlling age and education, was 1.70 (95% confidence interval [CI] = 1.40, 2.07). Odds ratios were homogeneous across studies, and were similar regardless of participants' gender or ethnicity. Persons sexually assaulted in childhood consistently had greater odds of headache than those first assaulted in adulthood (OR = 1.89, 95% CI = 1.19, 2.99). These results indicate that sexual assault, particularly in childhood, is associated with headache. PMID- 10535658 TI - Family accommodation of obsessive-compulsive symptoms: instrument development and assessment of family behavior. AB - Relatives frequently accommodate patients' obsessive-compulsive symptoms and clinicians hypothesize that such accommodations adversely affect patient outcome. This study's purpose was to develop a valid and reliable measure, the Family Accommodation Scale for Obsessive-Compulsive Disorder (FAS), and to investigate the family accommodation construct. We administered the FAS and additional family and patient measures to 36 adult obsessive-compulsive patients and their primary caregivers. The FAS demonstrated excellent interrater reliability and good internal consistency and performed well on assessment of its convergent and discriminant validity. Family accommodation was significantly associated with patient symptom severity and functioning, and with relatives' own obsessive compulsive symptoms. Although most relatives accommodated patient symptoms, many did not believe that such accommodations improved the patient's clinical status. The FAS will provide researchers and clinicians with a useful tool for assessing family accommodation and for identifying families who may benefit from interventions aimed at developing more adaptive coping strategies. PMID- 10535659 TI - Dose-dependent effects of nitric oxide synthase inhibition on systemic and renal hemodynamics in conscious lambs. AB - The present experiments were carried out to determine the role of nitric oxide in influencing systemic and renal hemodynamics in conscious young sheep. Parameters of cardiovascular function were measured before and for 4 h after intravenous injection of either L-NAME (NG-nitro-L-arginine methyl ester) or D-NAME (N(G) nitro-D-arginine methyl ester) at doses of 10, 20, or 40 mg/kg in 13 conscious, chronically instrumented young sheep aged 43 +/-5 days. Blood pressure increased and heart rate decreased in a dose-dependent manner following administration of L NAME. Renal vascular resistance was increased for 10 min following a dose of 10 mg/kg of L-NAME and for 120 min following a dose of 40 mg/kg of L-NAME. The renal vasodilatory response to close arterial injection of 1 microg/kg of acetylcholine was attenuated by L-NAME in a dose-dependent manner. These experiments provide the first information that under normal physiological conditions in conscious young animals, nitric oxide influences systemic and renal hemodynamics. PMID- 10535660 TI - Renal and vascular effects of chronic nitric oxide synthase inhibition: involvement of endothelin 1 and angiotensin II. AB - Endothelin 1 (ET-1) is a potent vasoconstrictor implicated in the control of blood pressure and renal function. Its effects can be modulated by nitric oxide (NO), which inhibits ET-1 production and action. Recently, we reported that ET-1 production can also be modulated by angiotensin II (AngII) in vivo. To investigate the interactions between NO, ET-1, and AngII in hypertension and renal dysfunction, we assessed immunoreactive ET-1 (ir-ET-1) concentration in plasma and urine as well as in vascular and renal tissues of rats with chronic inhibition of NO synthesis, in the presence and the absence of the AngII type 1 receptor antagonist losartan. Normal (protocols A and B) and uninephrectomized rats (protocol C) received the L-arginine analog N(G)-nitro-L-arginine methyl ester (L-NAME), an inhibitor of NO synthesis, 0.05% (protocol A) or 0.1% (protocols B and C), with or without losartan (20 mg x kg(-1) x day(-1)). After 6 weeks, systolic blood pressure was significantly increased in L-NAME rats compared with the controls (p < 0.01), while serum creatinine and urea, creatinine clearance, and proteinuria were similar to control values. However, ir ET-1 concentration in plasma and in the thoracic aorta was augmented in animals receiving 0.1% L-NAME (1 < 0.01), while it was unchanged in the mesenteric arterial bed, preglomerular arteries, and glomeruli. In contrast, ir-ET-1 concentration was decreased in the renal papilla (p < 0.05) as well as in the urine of L-NAME rats (p < 0.01). Treatment with losartan significantly attenuated the rise in systolic blood pressure induced by L-NAME (p < 0.01). Losartan also normalized the increased ir-ET-1 concentration in plasma and in the thoracic aorta, but had no effect on tissues with normal or reduced ir-ET-1 levels. These results indicate that chronic inhibition of NO synthase with L-NAME induces hypertension without renal dysfunction. Increased ET-1 production in some blood vessels and elevated circulating ET-1 concentration may contribute to the maintenance of high blood pressure. The reduction of systolic blood pressure by losartan supports a role for AngII in the pathogenesis of this form of hypertension, which may be due, at least in part, to the modulation of ET-1 production. PMID- 10535661 TI - New insights into body composition assessment in obese women. AB - During treatment of patients with non-insulin-dependent diabetes mellitus, there may be marked body weight loss. Therefore, body composition should be monitored to check for a decrease in fat mass alone, without an excessive decrease of both fat-free mass and total body water. Accordingly, it is useful to monitor the hydration of these patients. One method that allows us to check the status of body hydration is the multifrequency bioelectric impedance analysis (MFBIA). It makes use of formulas that estimate total body water on the basis of the concept that the human body may be approximated to a cylinder of length equal to body height. In normal subjects body water estimates are sufficiently accurate, but in obese subjects the true hydration status may be overestimated. In this report, we describe the accuracy of mathematical models previously described in the literature, and correct for the overestimation of total body water in obese subjects by means of a new equation based on a new model. The coefficients for each model have been recalculated by the weighing of our sample in order to test the accuracy of estimates obtained with the equations. This new model includes both body volume and two impedances at appropriate frequencies useful for identifying two terms strictly related to extra- and intra-cellular water. The new formulas do not include body weight, but they include the body volume, a parameter more closely related to the biophysical reference model. Fifty-five overweight females, body mass index ranging from 26.8 to 50.2 kg/m2, were enrolled in the study. The proposed equations, taking advantage of two impedance values at appropriate frequencies, better predict total body water in obese women. This was particularly evident when the results obtained with the multifrequency bioelectric impedance analysis and deuterium isotopic oxide dilution method were compared. Although this last method is considered the "gold standard," it is not suitable for use in routine clinical practice. In conclusion, evaluation of total body composition by means of bioelectric impedance analysis might be included in programs for the prevention of non insulin-dependent diabetes and for monitoring weight loss during overt pathology. PMID- 10535662 TI - Haloperidol-mediated phosphoinositide hydrolysis via direct activation of alpha1 adrenoceptors in frontal cerebral rat cortex. AB - In addition to its effect on D2 dopamine receptor blockades, haloperidol is able to interact with multiple neurotransmitters (NTs). Its action on phosphoinositide (PI) turnover was studied on cerebral cortex preparations. It induces an increase in inositol phosphate (IP) accumulation, which was only blunted by the alpha1 adrenoceptor blocker prazosin. Haloperidol maximal effect (Emax) was less than the effect of the full agonist norepinephrine (NE), and dose-response curves for both NE in the presence of submaximal doses of haloperidol and haloperidol in the presence of Emax doses of NE showed that haloperidol behaves as a partial agonist of cerebral alpha1-adrenoceptors. Its effect on PI hydrolysis is mediated through phospholipase C activation, as 2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate (NCDC) and 1-[6-([(17beta)-3-methoxyestra- 1,3,5(10)-trien-17-yl]amino)hexyl]-1H pyrrole-2,5-dione) (U-73122) were able to abrogate both haloperidol and NE actions. Further, the typical neuroleptic exerts a direct activation of alpha1 adrenoceptors as its actions were not modified by cocaine and persisted in spite of chemical rat cerebral denervation with 6-hydroxydopamine (6-OHDA). The possibility that this agonistic action on alpha1-adrenoceptors would be involved in haloperidol side effects is also discussed. PMID- 10535663 TI - Zidovudine (AZT) induced alterations in mitochondrial biogenesis in rat striated muscles. AB - Zidovudine (AZT) and didanosine (ddI), two drugs used in the treatment of AIDS, are also known to cause mitochondrial abnormalities. We investigated the physiological relevance of the mitochondrial defects by measuring in situ skeletal muscle performance and cytochrome c oxidase (CYTOX) enzyme activity in heart muscle, red highoxidative (RG) and white low-oxidative (WG) portions of the gastrocnemius muscle of control (n = 17), AZT-(n = 14), or ddI-treated (n = 11) rats for 28 days. We also evaluated the hypothesis that AZT treatment could alter the expression of the mitochondrial transcription factor A (mtTFA), a key molecule involved in mitochondrial DNA (mtDNA) replication and transcription. AZT had a pronounced effect on blood pressure and skeletal muscle performance, which were significantly decreased during contractile activity at 2 and 5 Hz, compared with control. A significant decrease in CYTOX activity in heart and RG, but not WG muscles, was also evident. In the heart, this was accompanied by an apparent compensatory increase in mtTFA mRNA level that could not be attributed to enhanced transcriptional activation mediated by nuclear respiratory factor 1 (NRF 1). In contrast with AZT, no effect of ddI was found on the extent of fatigue or muscle enzyme activity. These results indicate that AZT induces mitochondrial defects primarily in muscles with the highest oxidative capacities (heart and RG). The long-term effects of AZT on mitochondrial biogenesis have the potential to reduce muscle performance, but the effects on performance in this short-term study were likely due to an inability of the AZT-treated animals to maintain blood pressure during contractile activity. PMID- 10535664 TI - Concentration-dependent effects of angiotensin II on sinus rate in canine isolated right atrial preparations. AB - To investigate the concentration-response relationship of angiotensin II with respect to its chronotropic effects, the sinus rate was recorded from canine isolated right atrial preparations perfused through the sinus node artery. Nicotine (5 x 10(-5) M) injection induced an early, atropine-sensitive bradycardic response and a more delayed propranolol-sensitive tachycardic response, suggesting that the preparations contained both cholinergic and adrenergic neurons. The former response, but not the latter, was markedly reduced in preparations in which the right atrial ganglionated plexus was removed. Positive chronotropic responses were induced by angiotensin II over a wide range of concentrations (10(-12) - 5 x 10(-6) M), with a maximum increment of 29.9 +/- 9.6 beats/min. Responses to low concentrations (angiotensin II, 10(-11) M) were monophasic and were abolished by propranolol. In contrast, the responses to higher concentrations (angiotensin II, 10(-6) M) were not abolished by propranolol and were biphasic (early response, 29.9 +/- 12.1 beats/min; later response, 18.6 +/- 9.0 beats/min), the early response being blocked by losartan (AT1 antagonist) but not the later one, both being blocked by saralasin (nonselective angiotensin II antagonist). In conclusion, the data suggest that angiotensin II exerts its stimulant effects on the heart through receptors located either on cardiomyocytes or neurons, depending on the agonist concentration. PMID- 10535665 TI - Exercise promotes a subcellular redistribution of calcium-stimulated protease activity in striated muscle. AB - The aims of this study were (i) to investigate whether the contractile activity associated with running increases calcium-stimulated, calpastatin-inhibited protease activity (calpain-like) in a time-dependent manner and (ii) to determine whether the changes, if any, are proportionately distributed between soluble (cytosolic) and particulate (bound) fractions of striated muscle in vivo. Calcium dependent, calpastatin-inhibited caseinolysis (i.e., calpain-like activity) was measured in control and exercised rats (25 m/min, 0% grade) at 2, 5, 15, 30, and 60 min. Total calpain-like activity in skeletal muscle increased by 26% (13.2 +/- 1.3 vs. 17.9 +/- 2.2 U/g wet wt.) (p < 0.05) after running (60 min), accompanied by an increased activity in the particulate fraction. In cardiac muscle, exercise (60 min) increased total calpain-like activity by 33% (p < 0.05), which was attributable to increases in both the cytosolic and particulate fractions. Both tissues responded with an early (2-5 min) activation of total calpain-like activity (p < 0.05), supported by early increases for particulate fractions from skeletal muscle; whereas for cardiac muscle, a noticeable early drop (p < 0.05) occurred in the particulate fraction. Minimal changes were observed for total, cytosolic, and particulate fractions of noncontracting tissue (i.e., liver). The results of this study support the hypothesis that the total calpain-like activity increases associated with level running occur early on with exercise and that the increases are accompanied by changes in the redistribution of soluble to particulate fractions. The changes would set the stage for enhanced rates of protein degradation known to occur in striated muscle with exercise. PMID- 10535666 TI - Persistent functional and structural retinal anomalies in newborn rats exposed to hyperoxia. AB - Previous studies have shown that newborn rats exposed postnatally to hyperoxia will develop a permanent impairment of the retinal function as determined with the electroretinogram (ERG). The purpose of our study was to examine whether postnatal hyperoxia equally alters the light- and dark-adapted ERGs and oscillatory potentials (OPs) as well as leads to permanent structural modification of the retina. During the first 14 days of life, cohorts of Sprague Dawley rats were exposed to a hyperoxic environment, and ERGs were recorded at mean ages of approximately 25 and 55 days. Our results indicate that both light- and dark-adapted ERGs and OPs are already significantly altered within a few days following exposure to hyperoxia. None of the ERG and (or) OP parameters, with the exception of the a-wave, returned to normal values by 55 days of age. In fact some dark-adapted OPs were completely abolished following postnatal O2 exposure. Histological analysis revealed that the retina of rats exposed to hyperoxia failed to develop an outer plexiform layer and had a reduced count of horizontal cells, consistent with the permanent postreceptoral anomalies seen in the ERG responses. Our results suggest that postnatal hyperoxia causes a generalized retinal disorder leading to permanent structural modifications of the retinal cytoarchitecture and lasting anomalies of the rod and cone functions. PMID- 10535667 TI - Coronary endothelium-derived vasodilation during cooling and rewarming of the in situ heart. AB - The integrity of coronary vascular endothelial vasodilator function during core cooling and rewarming was investigated in a pentobarbital-anesthetized open-chest dog model. Vasodilator response was assessed as the change from baseline blood flow by injecting the endothelial-dependent vasodilator acetylcholine (ACh) (1.0 microg) or the endothelial-independent vasodilator nitroglycerin (NTG) (50 microg) into the left anterior descending (LAD) coronary artery. Change in blood flow was measured using a transit time ultrasonic volume flowmeter technique. During cooling and rewarming LAD blood flow was significantly decreased. After rewarming, aortic pressure was artificially elevated to reach control. This procedure restored heart work (LV-RPP, left ventricular rate pressure product) and coronary perfusion pressure, but LAD blood flow remained lowered. Ability to dilate the vascular bed supplied by LAD, after injections of ACh or NTG, was present both during cooling and rewarming. At 25 degrees C coronary blood flow (LAD) increased from 3 +/- 1 to 9 +/- 1 mL x min(-1) in response to both ACh and NTG. Posthypothermic blood flow increased from 7 +/- 1 to 19 +/- 2 and 20 +/- 3 mL x min(-1) in response to ACh and NTG, respectively. Measured as the percent change from baseline LAD blood flow, the response was not significantly different from the one obtained in prehypothermic hearts. In conclusion, coronary vasodilator function, both endothelium dependent and endothelium independent, is present but not maintained at the same level during cooling to 25 degrees C and rewarming. In spite of the deterioration of cardiac function, no selective defect in the endothelium-dependent response was detected, either during hypothermia or after rewarming. PMID- 10535668 TI - Heat shock does not attenuate low-frequency fatigue. AB - Whole-body hyperthermia or heat shock confers protection to myocardial contractility against reperfusion-induced injury. The purpose of this study was to determine whether heat shock could provide similar protection to skeletal muscle contractility against low-frequency fatigue. Male Sprague-Dawley rats (6 rats/group) were heat shocked at 41.5 degrees C for 15 min either 24 h or 4 days prior to fatiguing stimulation to compare the contractile responses of the plantaris muscle with those of a nonheated group. Both 24 h and 4 days after heat shock, the 72-kDa heat shock protein (HSP72) was elevated above control levels. There were no differences between the heat-shocked and non-heat-shocked animals in measures of contractility prior to fatiguing contractions or in resistance to fatigue. Heat-shock preconditioning did not lead to improved postfatigue force recovery above control responses and, in fact, delayed the recovery of force. This study does not support the use of heat-shock therapy to improve skeletal muscle contractile performance under fatiguing conditions. PMID- 10535669 TI - Modulation of vasomotion in resistance arteries of JCR:LA-cp rats: a model of insulin resistance. AB - Obesity and insulin resistance are strongly associated with an increased risk of vascular disease. Vasomotion is the cyclic variation in the diameter of arteries and is a general feature of the vasculature that may have important physiological consequences. We tested the hypothesis that obesity - insulin resistance is associated with abnormal vasomotion by comparing obese, insulin-resistant JCR:LA cp rats, known to develop vasculopathy, atherosclerosis, and ischemic lesions of the heart, with lean insulin-sensitive animals from the same strain. Vasomotion was assessed using isolated mesenteric arteries on a myograph system after preconstriction to 50% of maximal constriction with norepinephrine. The amplitude of vasomotion was enhanced by the presence of meclofenamate, a prostaglandin H synthase inhibitor, and was diminished by N(G)-nitro-L-arginine methyl ester (L NAME), a nitric oxide synthase inhibitor. Removal of the endothelium essentially abolished vasomotion, and meclofenamate had no effect on de-endothelialized arteries. Frequency was not altered by either L-NAME or meclofenamate. Although pharmacological inhibition of nitric oxide and eicosanoid production clearly altered vasomotion, there was no difference in the amplitude or frequency of vasomotion in arteries from obese rats compared with lean rats. These results indicate that the endothelium plays a central role in modulating vasomotion, involving both enhancing and inhibiting effects, and that vasomotion is similar between obese, insulin-resistant and lean, insulin-sensitive rats. PMID- 10535670 TI - Estrogen replacement increases coronary artery distensibility in ovariectomized rats. AB - The effect of estrogen on the passive characteristics of arteries is not known. We hypothesized that estrogen would increase arterial distensibility as part of its protective effect on the vasculature. Female Sprague-Dawley rats were ovariectomized at 11 weeks of age. One group received a placebo (n = 6), while two other groups (n = 5 each) of rats received a 17beta-estradiol pellet (0.15 mg or 0.5 mg with 60-day release). After 4 weeks of estrogen replacement, coronary and mesenteric arteries (<200 microm diameter) were dissected and mounted on a dual-chamber arteriograph. Lumen diameter and wall thickness were measured in pressurized arteries. The relative changes in diameter (distensibility) as well as wall thickness per unit change in pressure were significantly increased (p < 0.05) in the coronary arteries of the 0.5 mg estradiol replaced rats compared with the ovariectomized control animals and the 0.15 mg estradiol replaced rats. Surprisingly, in the mesenteric arteries from the same animals, there was no difference in distensibility or pressure - wall thickness among the groups. This study provides experimental data of a novel hypothesis that estrogen may afford part of its protection through vascular remodeling of the coronary circulation. PMID- 10535671 TI - Characterization of positive inotropic effect of colforsin daropate [correction of dapropate] hydrochloride, a water-soluble forskolin derivative, in isolated adult rat cardiomyocytes. AB - The present study was undertaken to characterize the positive inotropic action of colforsin daropate [corrected] hydrochloride (NKH477), a novel water-soluble forskolin derivative, on isolated cardiomyocytes of adult rats. Simultaneous measurements of cellular contraction and intracellular calcium concentration ([Ca2+]i) were carried out. The effects of isoprenaline and ouabain on these parameters were also determined for comparison. The contraction and maximum [Ca2+]i of NKH477-, isoprenaline-, or ouabain-treated cells were increased concentration dependently. Peak shortening of NKH477-treated cells was positively correlated with the shortening velocity and inversely with the time to peak shortening. Maximum, but not minimum, [Ca2+]i in NKH477-treated cells was correlated with the rate of increase in [Ca2+]i and inversely with the time to maximum [Ca2+]i. Similar results were obtained with isoprenaline. In contrast, ouabain increased both maximum and minimum [Ca2+]i. Treatment with either NKH477 or isoprenaline increased cellular cAMP content, but treatment with ouabain did not. These results suggest that the positive inotropic action of NKH477 is associated with an increase in [Ca2+]i and acceleration of its kinetics. PMID- 10535672 TI - Cellular localization and expression of insulin-like growth factors (IGFs) and IGF binding proteins within the epiphyseal growth plate of the ovine fetus: possible functional implications. AB - The insulin-like growth factors (IGFs) are important in the regulation of normal fetal musculoskeletal growth and development, and their actions have been shown to be modulated by IGF binding proteins (IGFBPs). Because the anatomical distribution of IGFBPs is likely to dictate IGF bioavailability, we determined the cellular distribution and expression of IGF-I, IGF-II, and IGFBP-1 to IGFBP-6 in epiphyseal growth plates of the fetal sheep, using immunocytochemistry and in situ hybridization. Little mRNA for IGF-I was detectable within the growth plates, but mRNA for IGF-II was abundant in germinal and proliferative chondrocytes, although absent from some differentiating chondrocytes and hypertrophic cells. Immunohistochemistry for IGF-I and IGF-II showed a presence of both peptides in all chondrocyte zones, including hypertrophic cells. Immunoreactive IGFBP-2 to -5 were localized within the germinal and proliferative zones of chondrocytes, but little immunoreactivity was present within the columns of differentiating cells. IGFBP immunoreactivity again appeared in hypertrophic chondrocytes. IGFBP mRNA in chondrocytes of the epiphyseal growth plate was below the detectable limit of in situ hybridization. However, low levels of mRNAs for IGFBP-2 to -6 were detected by the reverse transcriptase polymerase chain reaction. A co-localization of IGFBPs with IGF peptides in intact cartilage suggests that they may regulate IGF bioavailability and action locally. To test this hypothesis, monolayer cultures of chondrocytes were established from the proliferative zone of the growth plate, and were found to release immunoreactive IGF-II and to express mRNAs encoding IGFBP-2 to -6. Exogenous IGFBP-3, -4, and -5 had an inhibitory action on IGF-II-dependent DNA synthesis. IGFBP-2 had a biphasic effect, potentiating IGF-II action at low concentrations but inhibiting DNA synthesis at equimolar or greater concentrations relative to IGF-II. Long R3 IGF-I, which has a reduced binding affinity for many IGFBPs, was more potent than native IGF-I in promoting DNA synthesis by chondrocytes. Our findings suggest that locally produced IGF-II and IGF-I derived from the circulation can influence fetal epiphyseal chondrogenesis, and that this may be modulated locally by multiple IGFBP expression. PMID- 10535673 TI - Activation of bulbospinal serotonergic neurons during cold exposure. AB - In a four-part study, we expand on our previous report that bulbospinal serotonin (5HT) neuronal activation occurs with 24 h of cold exposure. To characterize temporal aspects, rats were exposed to 3 degrees C or were maintained at 22 degrees C for 2, 8, 48, or 96 h (experiment 1) or for 15, 30, or 60 min (experiment 2). To ensure that cold-induced changes in 5HT activity were not due to disturbances in diurnal pattern, rats in experiment 3 were exposed to cold (8 h) during the dark cycle. To explore the hypothesis that cold-induced 5HT activation is part of a broad metabolic response that includes activation of the sympathetic nervous system, metabolically impaired (hypothyroid) rats were exposed to 8 degrees C in experiment 4. Significant increments in 5 hydroxyindoleacetic acid (SHIAA) concentration were evident by 60 min of cold exposure and existed at all later time points measured. These findings were most robust in spinal cord and rostral brainstem. Activation in spinal cord was also found when rats were exposed to 8 h of cold during the dark cycle, the active period for rats. In experiment 4, hypothyroid rats exhibited significantly greater norepinephrine excretion compared with control rats exposed to the same cold stimulus; this finding was accompanied by significantly greater increments in 5HIAA concentration in rostral brainstem and spinal cord of hypothyroid rats. In addition, significant elevations in tryptophan concentration were noted throughout the brainstem and spinal cord of cold-exposed, hypothyroid rats relative to room temperature, hypothyroid rats. This finding suggested that elevations in 5HIAA concentration in these rats were due to increases in precursor availability. The implications of these findings relative to autonomic and metabolic control are discussed. PMID- 10535674 TI - Effects of adenosine and acadesine on interstitial nucleosides and myocardial stunning in the pig. AB - 5-Amino-4-imidazolecarboxamide riboside (AICAr) or acadesine has been proposed to exert cardioprotection by enhancing adenosine production in ischemic myocardium. However, there are conflicting reports on acadesine's effects in ischemic myocardium and few studies in which myocardial adenosine levels have been measured. The purpose of this study was to determine whether acadesine increases interstitial fluid adenosine levels and attenuates myocardial stunning or potentiates the effects of adenosine in the intact pig. In pentobarbital anesthetized pigs, myocardial stunning was induced by 10 min left anterior descending coronary artery occlusion and 90 min reperfusion. Regional ventricular function was assessed by measuring systolic wall thickening, and interstitial nucleosides were estimated by cardiac microdialysis. Control hearts were compared with hearts treated with acadesine, adenosine, and adenosine plus acadesine. Adenosine pretreatment (100 microg x kg(-1) x min(-1), intracoronary) immediately prior to ischemia increased interstitial adenosine levels 9-fold and improved postischemic functional recovery from a control value of 17.6 +/- 4.1% to 43.6 +/ 3.4% of preischemic systolic wall thickening. In contrast, acadesine (20 mg/kg i.v. bolus 10 min prior to ischemia + 0.5 mg x kg (-1) x min(-1), i.v. infusion through 60 min reperfusion) had no effect on interstitial fluid adenosine levels or the recovery of regional function (21.5 +/- 5.9% recovery), nor were the functional effects of adenosine potentiated by acadesine. These findings indicate that acadesine does not enhance myocardial adenosine levels, attenuate myocardial stunning, or potentiate the cardioprotective effects of adenosine in the pig. PMID- 10535675 TI - The anion transport inhibitor DIDS activates a Ba2+-sensitive K+ flux associated with hepatic exocrine secretion. AB - 4,4'-Diisothiocyanatostilbene-2,2'-disulfonate (DIDS), an anion transport inhibitor and choleretic organic anion, was used to study the relationship between putative DIDS-sensitive K channels and exocrine secretion in the isolated and bile duct cannulated perfused rat liver. Bile flow, DIDS excretion, and effluent perfusate K+ content were measured. DIDS (125 microM) caused a doubling in bile generation concomitant with its appearance in bile, confirming earlier reports. Furthermore, DIDS induced a transient increase in perfusate K+ concentration that peaked prior to the biliary parameters and, after 10 min, reversed to net uptake that fully compensated for the initial release. The K channel blocker Ba2+ (1 mM) strongly inhibited the release phase along with the accompanying choleresis and DIDS excretion. Ouabain (13.5 microM) alone was choleretic and hyperkalemic and, when applied in combination with DIDS, depressed DIDS excretion, choleresis, and DIDS-sensitive K+ uptake. To obtain further evidence for the presence of DIDS-sensitive K channels K+ flux was measured under the influence of different gradients of the cation. Perfusate K+ at 26 and 80 mM changed the DIDS-activated K+ flux from a transient outward to a sustained inward flux, and both DIDS excretion and bile flow decreased. Mean net K+ flux over 20 min DIDS perfusion changed from -1.3 +/-1.1 micromol/g with 5.9 mM K+ to -1304 +/ 55 micromol/g with 80 mM K+ in the perfusate. K+ efflux was fully and reversibly blocked by Ba2+ and influx was ouabain-insensitive, suggesting that the DIDS activated K+ flux was channel mediated. The results show that a significant fraction of DIDS-induced bile generation is associated with K+ release that may be mediated by Ba(2+)-sensitive K channels, possibly of the inward rectifying type. PMID- 10535676 TI - Two novel types of calcium release from skeletal sarcoplasmic reticulum by phosphatidylinositol 4,5-biphosphate. AB - In both the heavy and light fractions of fragmented sarcoplasmic reticulum (SR) vesicles from the fast skeletal muscle, about 27 min after beginning the active Ca2+ uptake, the extravesicular Ca2+ concentration suddenly increased to reach a steady level (delayed Ca2+ release). Phosphatidylinositol 4,5-bisphosphate (PIP2) not only shortened the time to delayed Ca2+ release but also induced prompt Ca2+ release from the heavy fraction of SR. Delayed Ca2+ release and prompt Ca2+ release stimulated by 100 microM PIP2 were not modified by ruthenium red. PIP2 (>0.1 microM) markedly accelerated the rate of 45Ca2+ efflux from SR vesicles in a concentration-dependent manner. The PIP(2)-induced 45Ca2+ efflux was potentiated by ruthenium red but profoundly inhibited by La3+. The concentration response curve for Ca2+ or Mg2+ in PIP2-induced 45Ca2+ release was clearly different from that in the Ca(2+)-induced Ca2+ release. PIP2 caused a concentration-dependent increase in Ca2+ release from SR of chemically skinned fibers from skeletal muscle. Furthermore, [3H]ryanodine or [3H]methyl-7 bromoeudistomin D (MBED) binding to SR was increased by PIP2 in a concentration dependent manner. These observations present the first evidence that PIP2 most likely activates two types of SR Ca2+ release channels whose properties are entirely different from those of Ca(2+)-induced Ca2+ release channels (the ryanodine receptor 1). PMID- 10535678 TI - Incommensurate frequencies of major vascular regulatory mechanisms. AB - The dynamic relationship among three major vascular control mechanisms that operate on large fractions of cardiac output: arterial baroreflex and renal and mesenteric autoregulation, was investigated in conscious rats. Wistar and spontaneously hypertensive rats were studied in their home cages 10 days after implantation of pulsed Doppler flow probes. There was an oscillation of blood pressure centered at 0.45 Hz that is associated with operation of arterial baroreflexes. Hindquarters blood flow displayed a featureless, "1/f' power spectrum, in which no autoregulatory or baroreflex signatures could be discerned, although active control of resistance over a wide range of frequencies was evident. The renal pressure - flow transfer function was dominated by an autoregulatory mechanism with a resonance peak at 0.25 +/- 0.01 Hz. In the mesenteric circulation an autoregulatory mechanism was seen with a resonance peak at 0.15 +/- 0.01 Hz and another active mechanism was seen above 0.2 Hz that appeared from its negative admittance phase to be a baroreflex. The center frequencies of mesenteric and renal autoregulation and of the arterial baroreflex were related in a ratio of 1 : 1.7 +/- 0.1 : 3.0 +/- 0.2 (approximately 4:7:12). Such relatively high order ratios can be expected to minimize the possibility of phase locking and (or) entrainment among the various control mechanisms. PMID- 10535677 TI - Lateral hypothalamic serotonergic responsiveness to food intake in rat obesity as measured by microdialysis. AB - The hypothalamic serotonergic system is involved in the regulation of food ingestion and energy metabolism. Since disturbances of both energy intake and expenditure can contribute to obesity, the objective of the present study was to evaluate the serotonergic response stimulated by food ingestion in two different models of obesity: the hyperphagic Zucker and the hypophagic and hypometabolic, monosodium glutamate (MSG) obese Wistar rat. For this we used microdialysis to examine the release of 5-hydroxytryptamine (serotonin, 5HT) and 5 hydroxyindoleacetic acid (5HIAA) in the lateral hypothalamus. Daily intake of MSG obese rats was 40% lower while that of Zucker obese rats was 60% higher than that of the respective lean controls. In overnight-fasted animals, 20-min microdialysate samples were collected before (basal release) and during a 2-h period of access to a balanced palatable food mash. The animals began to eat during the first 20 min of food access, and food consumption was similar among the four groups in all six individual 20-min periods recorded. Ingestion of food increased 5HT release in all groups. In MSG-obese and lean Wistar rats, 5HT levels were similarly elevated during the whole experimental period. In the Zucker strain, 5HT increments of basal release tended to be higher in obese than in lean rats at 20 and 40 min, and a significantly higher increment was observed at 60 min after food access (40 and 135% for lean and obese, respectively). The area under the curve relating serotonin levels to the 120 min of food availability was significantly higher in Zucker obese (246.7 +/- 23.3) than MSG obese (152.7 +/- 13.4), lean Wistar (151.9 +/- 11.1), and lean Zucker (173.5 +/- 24.0) rats. The present observation, of a food-induced serotonin release in the lateral hypothalamus of lean Wistar and Zucker rats, evidences that 5HT in the lateral hypothalamus is important in the normal response to feeding. In obese animals, the serotonin response was similar to (in the hypophagic-hypometabolic MSG model) or even higher than (in the hyperphagic Zucker model) that seen in the respective lean controls. This result indicates that the energy homeostasis disturbances of both these obesity models may not be ascribed to an impairment of the acute lateral hypothalamic serotonin response to a dietary stimulus. PMID- 10535679 TI - Differential response of rat skeletal muscle glycogen metabolism to testosterone and estradiol. AB - Although reports on sex steroids have implicated them as promoting protein synthesis and also providing extra strength to the skeletal muscle, it remains unclear whether sex steroids affect glycogen metabolism to provide energy for skeletal muscle functions, since glycogen metabolism is one of the pathways that provides energy for the skeletal muscle contraction and relaxation cycle. The purpose of the current study was to show that testosterone and estradiol act differentially on skeletal muscles from different regions, differentially with reference to glycogen metabolism. To study this hypothesis, healthy mature male Wistar rats (90-120 days of age, weighing about 180-200 g) were castrated (a bilateral orchidectomy was performed to test the significance of skeletal muscle glycogen metabolism in the absence of testosterone). One group of castrated rats was supplemented with testosterone (100 microg/100 g body weight, i.m., for 30 days from day 31 postcastration onwards). To test whether estradiol has any effect on male skeletal muscle glycogen metabolism 17beta-estradiol (5 microg/100 g body weight, i.m., for 30 days from day 31 postcastration onwards) was administered to orchidectomized rats. To test whether these sex steroids have any differential effect on skeletal muscles from different regions, skeletal muscles from the temporal region (temporalis), muscle of mastication (masseter), forearm muscle (triceps and biceps), thigh muscle (vastus lateralis and gracilis), and calf muscle (gastrocnemius and soleus) were considered. Castration enhanced blood glucose levels and decreased glycogen stores in skeletal muscle from head, jaw, forearm, thigh, and leg regions. This was accompanied by diminished activity of glycogen synthetase and enhanced activity of muscle phosphorylase. Following testosterone supplementation to castrated rats, a normal pattern of all these parameters was maintained. Estradiol administration to castrated rats did not bring about any significant alteration in any of the parameters. The data obtained suggest a stimulatory effect of testosterone on skeletal muscle glycogenesis and an inhibitory effect on glycogenolysis. Estradiol did not play any significant role in the skeletal muscle glycogen metabolism of male rats. PMID- 10535680 TI - Na+,K+ pump and Na+-coupled ion carriers in isolated mammalian kidney epithelial cells: regulation by protein kinase C. AB - This review updates our current knowledge on the regulation of Na+/H+ exchanger, Na+,K+,Cl- cotransporter, Na+,Pi cotransporter, and Na+,K+ pump in isolated epithelial cells from mammalian kidney by protein kinase C (PKC). In cells derived from different tubule segments, an activator of PKC, 4beta-phorbol 12 myristate 13-acetate (PMA), inhibits apical Na+/H+ exchanger (NHE3), Na+,Pi cotransport, and basolateral Na+,K+ cotransport (NKCCl) and augments Na+,K+ pump. In PMA-treated proximal tubules, activation of Na+,K+ pump probably plays a major role in increased reabsorption of salt and osmotically obliged water. In Madin Darby canine kidney (MDCK) cells, which are highly abundant with intercalated cells from the collecting duct, PMA completely blocks Na+,K+,Cl- cotransport and decreases the activity of Na+,Pi cotransport by 30-40%. In these cells, agonists of P2 purinoceptors inhibit Na+,K+,Cl- and Na+,Pi cotransport by 50-70% via a PKC independent pathway. In contrast with MDCK cells, in epithelial cells derived from proximal and distal tubules of the rabbit kidney, Na+,K+,Cl- cotransport is inhibited by PMA but is insensitive to P2 receptor activation. In proximal tubules, PKC-induced inhibition of NHE3 and Na+,Pi cotransporter can be triggered by parathyroid hormone. Both PKC and cAMP signaling contribute to dopaminergic inhibition of NHE3 and Na+,K+ pump. The receptors triggering PKC-mediated activation of Na+,K+ pump remain unknown. Recent data suggest that the PKC signaling system is involved in abnormalities of dopaminergic regulation of renal ion transport in hypertension and in the development of diabetic complications. The physiological and pathophysiological implications of PKC-independent regulation of renal ion transporters by P2 purinoceptors has not yet been examined. PMID- 10535681 TI - Nitric oxide inhibits human and canine pulmonary vascular tone via a postjunctional, nonelectromechanical, cGMP-dependent pathway. AB - We examined nitric oxide mediated regulation of pulmonary arterial and venous smooth muscle (PASM and PVSM, respectively): whether this inhibition is mediated via prejunctional receptors on adrenergic nerve endings; whether NO is neuronally derived; the relationship between degree of inhibition and vessel size; and identification of the signalling mechanisms involved. Canine pulmonary vascular tissues were generally quiescent, while human PASM exhibited spontaneous phasic activity. The nitric oxide (NO) synthesis inhibitor Nomega-nitro-L-arginine (L NNA; 10(-4) M) increased tone and enhanced phasic activity. Electrical field stimulation (EFS) evoked contractions were markedly enhanced by L-NNA in an endothelium-dependent fashion, and antagonized by the NO donor S-nitroso-N acetylpenicillamine (SNAP; 10(-7) to 10(-5) M). 8-Bromo-cGMP mimicked the effects of SNAP on basal tone and EFS contractions, while an inhibitor of soluble guanylate cyclase mimicked those of L-NNA. While mechanical responses to exogenously added norepinephrine (10(-9)-10(-4) M) were also enhanced by L-NNA and suppressed by SNAP, EFS-evoked excitatory junction potentials were unaffected by SNAP. We conclude that, in human and canine PASM and PVSM, there is a tonic generation of NO originating within the endothelium that does not mediate a prejunctional effect, but which acts postjunctionally to activate a cGMP dependent pathway within the smooth muscle. PMID- 10535683 TI - Sodium-pump potentials and currents in guinea-pig ventricular muscles and myocytes. AB - When guinea-pig papillary muscles were depolarized to ca. -30 mV by superfusion with K+-free Tyrode's solution supplemented with Ba2+, Ni2+, and D600, addition of Cs+ transiently hyperpolarized the membrane in a reproducible manner. The size of the hyperpolarization (pump potential) depended on the duration of the preceding K+-free exposure; peak amplitudes (Epmax) elicited by 10 mM Cs+ after 5 , 10-, and 15-min K+-free exposures were 12.9, 17.7, and 23.2 mV, respectively. Pump potentials were unaffected by external Cl- but suppressed by cardiac glycosides, hyperosmotic conditions, and low-Na+ solution. Using Epmax as an indicator of Na+ pump activation, the half-maximal concentration for activation by Cs+ was 12-16.3 mM. At 6 mM, Cs+ was three times less potent than Rb+ or K+ and five times more potent than Li+. From these findings, and correlative voltage clamp data from myocytes, we calculate that (i) a pump current of 7.8 nA/cm2 generates an Epmax of 1 mV and (ii) resting pump current in normally polarized muscle (approximately 0.16 microA/cm2) is five times smaller than previously estimated. PMID- 10535682 TI - Biochemical and pharmacological characteristics of a newly synthesized H+-K+ ATPase inhibitor, YJA20379-1, 2-amino-4,5-dihydro-8-phenylimidazole[2,1 b]thiazolo[5,4-g]benzothiazol e. AB - The biochemical and pharmacological characteristics of a newly synthesized H+-K+ ATPase inhibitor, 2-amino-4,5-dihydro-8-phenylimidazole[2,1-b]thiazolo[5,4 g]benzothiazole (YJA20379-1), were investigated. In the pig gastric microsomes, YJA20379-1 inhibited the gastric H+-K+ ATPase regardless of pH condition, IC50 values being 21 and 24 microM at pH 6.4 and 7.4, respectively. The inhibitory activity of YJA20379-1 was antagonized by dithiothreitol treatment but could not be reversed by dilution and washing of the enzyme preparation. In Sprague-Dawley rats, YJA20379-1, administered i.d., p.o, i.v., or s.c., significantly inhibited basal gastric acid secretion, with ED50 values of 4.7, 20.2, 6.3, and 13.4 mg/kg, respectively. The antisecretory action of YJA20379-1 was short lasting (less than 7 h at an oral dosing of 30 mg/kg). Oral administration of YJA20379-1 also prevented the formation of ethanol, indomethacin, and water immersion stress induced gastric lesions and mepirizole-induced duodenal ulcers in rats. Furthermore, YJA20379-1 accelerated the healing of acetic acid induced chronic gastric ulcers in rats. In conclusion, these results suggest that YJA20379-1 has a potent inhibitory activity on the gastric H+-K+ ATPase but much shorter duration of antisecretory action than omeprazole, thereby exerting its anti-ulcer effects partly with cytoprotective activity. PMID- 10535684 TI - Na+ channel and Na+-K+ ATPase involvement in norepinephrine- and veratridine stimulated metabolism in perfused rat hind limb. AB - In the constant flow perfused rat hind limb, norepinephrine (NE) evoked increases in oxygen uptake (VO2) and lactate efflux (LE) were inhibited by the cardiac glycoside ouabain (1 mM), without interrupting the NE-mediated vasoconstriction. The membrane labilizer veratridine, previously shown to increase VO2 and LE, without increasing perfusion pressure, was also shown to be inhibited by the cardiac glycoside ouabain, as well as by the ouabain analogues digitoxin and digoxin. The stimulatory actions of veratridine on VO2 were inhibitable by low doses of the specific sodium channel blocker tetrodotoxin (TTX), while NE effects were unaffected, suggesting that NE may be acting via a TTX-insensitive sodium channel. It is concluded that agents such as NE (a vasoconstrictor) or veratridine (a membrane labilizer), which stimulate VO2 in the perfused rat hind limb, do so by increasing Na+ influx. The observed increases in oxygen consumption and LE are due to Na+-K+ ATPase activity to pump Na+ out of the cell at the expense of ATP turnover. Energy dissipation due to Na+ cycling may be a form of facultative thermogenesis attributable to NE that can be stimulated by membrane labilizers such as veratridine in the constant flow perfused rat hind limb. PMID- 10535685 TI - Gender difference in baroreflex-mediated bradycardia in young rats: role of cardiac sympathetic and parasympathetic components. AB - In a previous clinical study we have demonstrated a significantly lower baroreflex-mediated bradycardic response in young women compared with men. The present study determined whether sexual dimorphism in baroreflex sensitivity in young rats also covers the reflex tachycardic response. The study was then extended to test the hypothesis that an attenuated cardiac cholinergic component of the baroreflex heart rate response in females may account for the gender difference. Baroreflex sensitivity (BRS) was expressed as the regression coefficient of the reciprocal relationship between evoked changes in blood pressure and heart rate. BRS measured in conscious rats with phenylephrine (BRS(PE)) and nitroprusside (BRS(NP)) represented the reflex bradycardic and tachycardic responses, respectively. Female rats exhibited significantly lower BRS(PE) compared with male rats (-1.53+/-0.1 vs. -2.36+/-0.13 beats x min(-1) x mmHg(-1); p < 0.05) but similar BRS(NP) (-2.60+/-0.20 vs. -2.29+/-0.17 beats x min(-1) x mmHg(-1)). Blockade of cardiac muscarinic receptors with atropine methyl bromide elicited greater attenuation of BRS(PE) in male than in female rats (72+/-4.6 vs. 53+/-6.7% inhibition; p < 0.01) and abolished the gender difference. In male rats cardiac muscarinic blockade attenuated BRS(PE) significantly more than did cardiac beta-adrenergic receptor blockade with propranolol (72+/-4.6 vs. 43+/-2.7; p < 0.01), which suggests greater dependence of BRS(PE) on the parasympathetic component. In females, muscarinic and beta adrenergic blockade elicited similar attenuation of BRS(PE). The findings suggest that (i) BRS is differentially influenced by gender; female rats exhibit substantially lower BRS(PE) but similar BRS(NP) compared with age-matched male rats and (ii) the sexual dimorphism in BRS(PE) results, at least partly, from a smaller increase in vagal outflow to the heart in response to baroreceptor activation. PMID- 10535687 TI - Do local immune-neuroendocrine disturbances initiate diabetes? AB - It has been suggested that there exists a local immune-neuroendocrine self regulating system in the pancreas. The system consists of beta-cells, nerve ganglia, intercellular fluid, connective tissue, and endothelial and immunocompetent cells. The local immune-neuroendocrine system governs the background level of insulin production by intrinsic mechanisms both in normal conditions and in a recovery period after different kinds of stress. The activity of this system by a complex of metabolic, environmental, nerve, and nonspecific immune factors has been determined. The local immune-neuroendocrine system is partially autonomous as a result of local integrative nerve circuits, morphological and functional substrates. Increased or decreased synthesis and release of some cytokines or biologically active substances (neurotransmitters, neuropeptides, gamma-aminobutyric acid, metabolites, nitric oxide, ions, etc.) by various cell types in the local immune-neuroendocrine system above usual levels may result in disturbances of sensitivity and functions of beta-cells. If the capability of the local immune-neuroendocrine system is insufficient for their compensation, the islet cell autoantigens may occur, the specific immune mechanisms are involved, and the pathological process becomes irreversible. Some ways for prevention of disturbances in the local immune-neuroendocrine system during the early and late phases of diabetes are presented. PMID- 10535686 TI - Functional involvement of angiotensin AT2 receptor in adrenal catecholamine secretion in vivo. AB - The aim of the present study was to analyse modulations of adrenal catecholamine secretion from the adrenal gland of anesthetized dogs in response to locally administered angiotensin II (AngII) in the presence of either PD 123319 or CGP 42112, both of which are highly specific and selective ligands to angiotensin AT2 receptor. Plasma concentrations of epinephrine and norepinephrine in adrenal venous and aortic blood were quantified by a high performance liquid chromatography coupled with electrochemical detection (HPLC-EC) method. Adrenal venous blood flow was measured by gravimetry. Local administration of AngII (0.05 microg, 0.1 microM) to the left adrenal gland increased adrenal gland catecholamine output more than 30 times that found in nonstimulated states. Administration of either PD 123319 (0.085 microg (0.23 microM) to 8.5 microg (23 microM)) or CGP 42112 (0.005 microg (0.01 microM) to 5 microg (10 microM)) did not affect the basal catecholamine output significantly. The increase in adrenal catecholamine output in response to AngII was inhibited by approximately 80% following the largest dose of PD 123319. CGP 42112 significantly attenuated the catecholamine response to AngII by approximately 70%. PD 123319 and CGP 42112 were devoid of any agonist actions with respect to catecholamine output by the adrenal gland in vivo. Furthermore, both PD 123319 and CGP 42112 inhibited the increase in adrenal catecholamine secretion induced by local administration of AngII. The present study suggests that AT2 receptors play a role in mediating catecholamine secretion by the adrenal medulla in response to AngII receptor agonist administration in vivo. PMID- 10535688 TI - Relative resistance of functional beta2-adrenoceptor-mediated smooth muscle responses to in vitro desensitization. AB - The effects of in vitro incubation of rat isolated left atria, pulmonary artery rings, and aortic rings with isoprenaline (10(-6) M for 6 h) were examined to compare the degree of desensitization of beta1- and beta2-adrenoceptor-mediated functional responses. The experimental protocols were carefully controlled to exclude influence from persistence of agonist in the tissues after the prolonged exposures, time-dependent changes in tissue sensitivity, and the methods of plotting the data. Concentration-response curves for isoprenaline were constructed before incubation with isoprenaline and, after washout during 1 h, a second curve was obtained. Two protocols were employed: firstly, the preincubation curve was constructed to ensure that a maximum response was obtained (>10(-6) M) and, secondly, the preincubation curve was constructed to a maximum isoprenaline concentration of 10(-6) M. Preincubation curves were corrected for time-dependent changes in sensitivity from sham-incubation control experiments. There was significant desensitization of the beta1-adrenoceptor mediated positive inotropic responses of the left atria, using both protocols, seen as rightward shifts (dose ratios: 4.48 +/- 1.12 and 8.39 +/- 2.3) of the concentration-response curves and depression of the maximum responses (77.0 +/- 3.2 and 60.8 +/- 5.5%). In contrast, the beta2-adrenoceptor-mediated relaxations of the noradrenaline-constricted pulmonary artery and aorta did not display a significant loss of sensitivity. When the relaxation responses were plotted as a percentage of the noradrenaline-induced tone, there was no significant rightward shift of the concentration-response curves in the pulmonary artery (dose ratios: 2.82 +/- 1.33 and 2.24 +/- 0.62) or aorta (dose ratios: 1.43 +/- 0.62 and 1.31 +/ 0.27) and thus no desensitization. PMID- 10535689 TI - Effect of alpha-phenyl-N-tert-butylnitrone on diabetes and lipid peroxidation in BB rats. AB - Oxygen free radicals have been shown to interfere with pancreatic islet beta cell function and integrity, and have been implicated in autoimmune type 1 diabetes. We hypothesized that the spontaneous autoimmune type 1 diabetes of the BB rat would be prevented by in vivo administration of a free-radical spin trap, alpha phenyl-N-tert-butylnitrone (PBN). Twenty-eight diabetes-prone (BBdp) and 13 non diabetes-prone (BBn) rats received PBN (10 mg/kg) subcutaneously twice daily, and 27 BBdp and 12 BBn rats received saline as controls. Rats were treated from age 47 +/- 6 days until diabetes onset or age 118 +/- 7 days. PBN caused no growth, biochemical, or hematological side effects. Sixteen control BBdp rats became diabetic (BBd, mean age 77 +/- 6 days) and six demonstrated impaired glucose tolerance (IGT rats). The incidence of diabetes and IGT was not different in PBN treated BBdp rats. Saline-treated rats showed no differences in pancreatic malondialdehyde (MDA) contents of BBd, IGT rats, and the BBdp that did not develop diabetes, versus BBn rats (2.38 +/- 0.35 nmoL/g). Among rats receiving PBN, BBn had lower pancreatic MDA than BBd and IGT rats (1.38 +/- 0.15 vs. 1.88 +/- 0.15 and 2.02 +/- 0.24 nmoL/g, p < 0.05), but not than BBdp rats (1.78 +/- 0.12 nmoL/g, ns). BBn rats receiving PBN also had lower pancreatic MDA than the saline controls (p < 0.05). Thus, PBN is remarkably nontoxic and is able to decrease MDA in the absence of the autoimmune process, but does not prevent diabetes. A combination of PBN with other complementary antioxidant agents may hold better promise for disease prevention. PMID- 10535690 TI - Response component analysis of simple and complex cells of area 18 during depression of area 17. AB - Simple and complex cells of visual areas of cats may be reliably classified according to the modulatory index (MI) of their responses. This investigation is aimed at analysing the MI in area 18 when a small region (about 200-400 microm in diameter) of area 17 was inactivated with a microinjection of GABA, in anesthetized cats. Cells were stimulated with sine-wave gratings whose orientation, spatial, and temporal frequencies were optimal for the studied unit. The AC and DC response components, and the MI were computed along with fast Fourier transforms of evoked discharges recorded as peristimulus time histograms. Results showed that these response components were relatively unaffected in simple cells, whereas complex cells exhibited large changes when area 17 was silenced. In particular, a large proportion of complex cells showed a MI greater than 1, thereby adopting a response pattern resembling simple cells. It is suggested that this subpopulation of complex cells receives a direct input from geniculate X cells. PMID- 10535691 TI - Effects of ethanol and diabetes on galactose oxidative metabolism and elimination in rats. AB - Blood galactose clearance after an intravenous galactose load has been widely used for years as an index of liver function. We developed a noninvasive [13C]galactose breath test, which explores galactose oxidative metabolism; this test is well correlated with liver fibrosis in patients with chronic viral hepatitis. The goal of this study was to evaluate the influence of nonhepatic factors such as diabetes and ethanol on whole-body galactose clearance (measured as the serum galactose elimination capacity test) and oxidative metabolism (measured as the [13C]galactose-induced breath 13CO2 production) in rats. Acute ethanol administration induced a significant decrease of galactose clearance and 13CO2 production. There was a significant correlation between the amount of ethanol given and the inhibition of galactose metabolism (R2 = 0.72, p < 0.0001). In streptozotocin-induced diabetic rats, the [13C]galactose-induced breath 13CO2 production was significantly reduced (p < 0.0001) and normalized by insulin treatment. However, diabetes did not decrease whole-body galactose clearance, indicating an isotopic dilution of [13C]glucose produced from [13C]galactose metabolism into the enlarged glucose pool. These results must be taken into account when using the [13C]galactose breath test as a quantitative liver function test. PMID- 10535693 TI - Mitochondrial membrane potential regulation is independent of c-fos expression. AB - Tumour cells contain mitochondria with elevated membrane potentials compared with normal cells, and thus this feature provides a selective target for destroying tumour cells. To improve mitochondrial-based therapies, a better understanding of the factors involved in regulating mitochondria are required. Since v-fos overexpression has been shown to elevate mitochondrial membrane potentials in rat fibroblasts, we investigated whether the human homologue, c-fos, was also capable of regulating the mitochondrial membrane potential in cells. Rat fibroblasts transfected with the c-fos gene did not accumulate more rhodamine 123 (Rh123) nor did they retain this Rh123 for extended periods of time compared with their parental line. Moreover, there was no difference in survival following dequalinium chloride (Deca) treatment between transfectants and controls. Similarly, reduction of c-fos expression in rat fibroblasts did not significantly alter their mitochondrial membrane potential. In addition, human ovarian carcinoma cells, which overexpress the c-fos gene, did not accumulate more Rh123 nor were they hypersensitive to Deca compared with their parental line. In another human ovarian carcinoma cell line, selection of variants with lower mitochondrial membrane potential did not alter c-fos mRNA or protein levels. These data suggest that alterations in c-fos expression do not regulate the magnitude of the mitochondrial membrane potential. PMID- 10535692 TI - Nonpeptide endothelin receptor antagonists attenuate the pressor effect of diaspirin-crosslinked hemoglobin in rat. AB - Endothelin 1 (ET-1) is a potent vasoactive and mitogenic peptide that is thought to participate in the hemodynamic effects elicited by drugs that block the biosynthesis and release of endothelium-derived nitric oxide (NO), such as NO synthase inhibitors. Using the nonpeptide endothelin receptor antagonists bosentan and LU-135252, we tested the hypothesis that endothelins contribute to the pressor activity of diaspirin-crosslinked hemoglobin (DCLHb), a hemoglobin based oxygen carrier, whose pressor activity in mammals is attributed primarily to a scavenging action towards NO. The NO synthase inhibitor nitro-L-arginine methyl ester (L-NAME), ET-1, and noradrenaline (NA) were used as reference drugs. Bosentan markedly reduced the pressor effects elicited by DCLHb, L-NAME, and ET 1, but not those evoked by NA. LU-135252 attenuated the pressor effect elicited by DCLHb and ET-1, but not that produced by L-NAME or NA. The decreases in heart rate associated with the pressor effect of DCLHb and L-NAME were reduced by LU 135252, whereas only those elicited by DCLHb were attenuated by bosentan. In contrast with bosentan, LU-135252 caused a decrease in the baseline blood pressure and heart rate. These results suggest that endothelins may participate in the pressor activity of DCLHb. They suggest also that nonpeptide endothelin receptor antagonists such as bosentan or LU-135252 may be useful to counteract endothelin-mediated undesirable hemodynamic effects of drugs that inhibit the activity of the NO system. PMID- 10535694 TI - Glutaminergic and adrenergic receptors expressed on adult guinea pig Schwann cells in vitro. AB - We have investigated the responsiveness of adult guinea pig Schwann cells to a range of neuroligands, using ratiometric calcium imaging. The majority of cells responded to ATP (90 +/- 4%), adrenaline (57 +/- 5%), and noradrenaline (61 +/- 5%), as well as glutamate (60 +/- 5%). The number of cells responding to glutamate increased significantly (90 +/- 4%; p < 0.01) when the cells were grown in excitatory amino acid (EAA) free medium, indicating EAA-induced downregulation. Only a small number of cells (9 +/- 2%) responded to acetylcholine. Agonist and antagonist experiments show that these adult Schwann cells predominantly express ionotropic glutaminergic receptors (N-methyl-D aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isooxazolepropionic acid (AMPA), and kainate) as well as alpha1-, alpha2-, and beta-adrenoreceptors. We conclude that Schwann cells derived from adult guinea pigs express a variety of neuroligand receptors when established in culture and are particularly rich in glutamate receptors. This probably reflects a de-differentiated state important to development and regeneration. PMID- 10535695 TI - Are prostaglandins involved in atrial natriuretic peptide mechanisms of cardiovascular control? AB - Atrial natriuretic peptide (ANP) can excite cardiac nerve endings and invoke a decrease in arterial blood pressure and a reduction in renal sympathetic nerve activity. Our laboratory has previously demonstrated that this renal depressor reflex was invoked by systemic injection of ANP and not by the direct application of ANP to the epicardium, a major locus for vagal afferents. We now examine whether inhibition of prostaglandin synthesis impairs reflex responses that are normally associated with ANP injections. Renal sympathetic nerve activity, arterial blood pressure, and heart rate were recorded in anesthetized rats. Indomethacin was used to inhibit prostaglandin synthesis through the cyclooxygenase pathway. The ANP-mediated decrease in arterial blood pressure and renal sympathetic nerve activity, observed when prostaglandin synthesis was inhibited, did not differ significantly from the decreases observed in these parameters when prostaglandin synthesis was not inhibited. Heart rate remained unchanged. Our results suggest that the sympatho-inhibitory effects of ANP do not require prostaglandins as intermediary compounds. PMID- 10535696 TI - Isolation of angiotensin converting enzyme (ACE) binding protein from human serum with an ACE affinity column. AB - Immobilized angiotensin-converting enzyme (ACE) was utilized as an affinity ligand to isolate a naturally occurring ACE binding protein from normal human serum. The enzyme was isolated from solubilized bovine lung membrane preparations by lisinopril affinity chromatography. It had an estimated molecular weight of 180 000 and was recognized by the anti-ACE antibody for the rabbit testicular ACE in immunoblots. ACE was immobilized onto epoxy Sepharose as well as Affi-Gel 15. Immobilized ACE on Affi-Gel 15 had higher catalytic activity (0.176 U/mL) compared with the enzyme immobilized on epoxy Sepharose (0.00005 U/mL). Immobilized ACE served as the affinity ligand for the identification of the ACE binding protein in human serum with an estimated molecular weight of 14 000 as observed by SDS polyacrylamide gel electrophoresis. The identification and further characterization of ACE binding proteins in serum and tissues may facilitate the greater understanding of the endogenous regulation of this key enzyme, which is involved in blood pressure homeostasis. PMID- 10535697 TI - Pharmacological uses and perspectives of heavy water and deuterated compounds. AB - Since the discovery of D20 (heavy water) and its use as a moderator in nuclear reactors, its biological effects have been extensively, although seldom deeply, studied. This article reviews these effects on whole animals, animal cells, and microorganisms. Both "solvent isotope effects," those due to the special properties of D20 as a solvent, and "deuterium isotope effects" (DIE), which result when D replaces H in many biological molecules, are considered. The low toxicity of D20 toward mammals is reflected in its widespread use for measuring water spaces in humans and other animals. Higher concentrations (usually >20% of body weight) can be toxic to animals and animal cells. Effects on the nervous system and the liver and on formation of different blood cells have been noted. At the cellular level, D20 may affect mitosis and membrane function. Protozoa are able to withstand up to 70% D20. Algae and bacteria can adapt to grow in 100% D2O and can serve as sources of a large number of deuterated molecules. D2O increases heat stability of macromolecules but may decrease cellular heat stability, possibly as a result of inhibition of chaperonin formation. High D2O concentrations can reduce salt- and ethanol-induced hypertension in rats and protect mice from gamma irradation. Such concentrations are also used in boron neutron capture therapy to increase neutron penetration to boron compounds bound to malignant cells. D2O is more toxic to malignant than normal animal cells, but at concentrations too high for regular therapeutic use. D2O and deuterated drugs are widely used in studies of metabolism of drugs and toxic substances in humans and other animals. The deuterated forms of drugs often have different actions than the protonated forms. Some deuterated drugs show different transport processes. Most are more resistant to metabolic changes, especially those changes mediated by cytochrome P450 systems. Deuteration may also change the pathway of drug metabolism (metabolic switching). Changed metabolism may lead to increased duration of action and lower toxicity. It may also lead to lower activity, if the drug is normally changed to the active form in vivo. Deuteration can also lower the genotoxicity of the anticancer drug tamoxifen and other compounds. Deuteration increases effectiveness of long-chain fatty acids and fluoro-D phenylalanine by preventing their breakdown by target microorganisms. A few deuterated antibiotics have been prepared, and their antimicrobial activity was found to be little changed. Their action on resistant bacteria has not been studied, but there is no reason to believe that they would be more effective against such bacteria. Insect resistance to insecticides is very often due to insecticide destruction through the cytochrome P450 system. Deuterated insecticides might well be more effective against resistant insects, but this potentially valuable possibility has not yet been studied. PMID- 10535698 TI - Nonadrenergic noncholinergic vasodilation of guinea pig pulmonary arteries is mediated by nitric oxide. AB - Nonadrenergic noncholinergic (NANC) mediated vasodilation may contribute to the maintenance of low pulmonary vascular tone. The NANC neurotransmitters, nitric oxide (NO) and the sensory neuropeptides, substance P and calcitonin gene related peptide (CGRP), were investigated as possible mediators of NANC vasodilation in guinea pig pulmonary arteries. Fresh guinea pig pulmonary artery rings, with and without an intact endothelium, were mounted in organ baths containing Krebs solution and precontracted with the prostaglandin F2alpha analogue U44069. In both endothelium-intact and denuded vessels, electrical field stimulation (1-12 Hz) in the presence of guanethidine and atropine resulted in a frequency dependent vasodilation. The peptide fragment hCGRP8-37, a competitive antagonist of the CGRP receptors, the peptide fragment NK1 antagonist SP4-11, and the nonpeptide NK1 antagonist RP67580 had no effect on NANC vasodilation. In both endothelium-intact and denuded vessels, N(G)-nitro-L-arginine methyl ester (L NAME), an inhibitor of NO synthesis, inhibited NANC vasodilation, an effect that was reversible with L-arginine. We conclude that NANC vasodilation in guinea pig pulmonary arteries is mediated predominantly through NO activity. PMID- 10535699 TI - Galactose transport inhibition by cytochalasin E in rat intestine in vitro. AB - Cytochalasins are cytoskeleton disrupters, and cytochalasin E has been reported to increase intestinal paracellular permeability. In this study, the cytochalasin E effect on galactose transport has been investigated. Ussing-type chamber experiments show an inhibitory effect of 20 microM cytochalasin E on unidirectional mucosal to serosal flux of galactose. On the contrary, the opposite unidirectional flux is not modified by the inhibitor. Results using intestinal everted sacs and rings confirm that galactose uptake by the tissue is diminished by cytochalasin E. The effect appears already after 5 min incubation, depends on cytochalasin E concentration, and does not occur in the absence of Na+. The inhibition is accompanied by an increase in the apparent K(m) of the active sugar transport (11.5 vs.15.8 mM) without significant change in the VmaX (10.6 vs. 9.1 micromol x g(-1) wet weight x 5 min(-1)). Cytochalasin E does not modify either galactose uptake by brush border membrane vesicles or Na(+)-K(+) ATPase activity in the enterocytes, indicating that the inhibitory effect on the Na(+)-dependent sugar transport cannot be explained as a direct effect on SGLT1 activity or as an indirect effect through the Na(+)-K(+) ATPase. Thus, our results suggest that cytochalasin E decreases SGLTI activity indirectly through cytoskeleton disruption. PMID- 10535700 TI - Vascular, renal, and endocrine responses to low-dose atrial natriuretic peptide in the fluid-balanced New Zealand genetically hypertensive rats with and without endogenous arginine vasopressin. AB - In hypertension, the relationship between atrial natriuretic peptide (ANP) and vasopressin (AVP) is not yet clear, although their renal actions are effectively autoregulation. To examine the possible interaction further, the responses to ANP infusion (75 ng x min (-1), i.v.) have been investigated in both hypertensive and normotensive AVP-replete (HT and NT) and AVP-deficient (HTDI and NTDI) rats. This study aimed to assess the renal function and the plasma hormone concentrations of AVP, angiotensin II (AII), ANP, aldosterone, and corticosterone in the conscious, chronically catheterized, fluid-balanced rats, and to examine the cardiovascular, renal, and endocrine responses to a constant infusion of a low-dose ANP. Data gained from the present study showed, for the first time, the hormone profile, plasma electrolyte composition, and detailed renal function of the servo controlled, fluid-balanced rats. The similarities of plasma electrolyte composition between servo-controlled and untreated rats indicated that the servo controlled fluid replacement technique maintained the differences between the strains and maintained body fluid balance during the experimental periods. Following ANP administration, there were no changes in glomerular filtration rate (GFR) in all groups, but an enduring diuresis and natriuresis were observed in HT and NT, which were milder in HTDI rats. However, the hypotensive effect of ANP was of a similar magnitude in all rat strains. HTDI rats exhibited an inhibition of the renin-angiotensin system (RAS), which may have participated in the reduced mean arterial blood pressure (MAP) and natriuresis observed in these rats. The renal actions of ANP appear to rely upon renal tubular events, as indicated by increased fractional electrolyte excretions in the AVP-replete rats. This study highlights the importance of AVP to the profile of the renal actions of ANP in normal rats. PMID- 10535701 TI - Interaction between prostanoids and nitric oxide in the control of tubular function in rats with chronic bile duct ligation. AB - Recent work indicates that both nitric oxide and cyclooxygenase products play an important role in the renal alterations of liver cirrhosis, although the interactions between them have not been completely established. The purpose of this study was to assess the effect of simultaneous blockade of nitric oxide synthase and cyclooxygenase in rats with chronic bile duct ligation and in control, sham-operated rats. Compared with control rats, chronic bile duct ligation rats, 23-25 days after surgery, showed a decreased mean arterial pressure, natriuresis, and kaliuresis, without differences in glomerular filtration rate, and an increased urinary nitrite excretion. Nitric oxide synthesis inhibition by administration of N(G)-nitro-L-arginine methyl ester induced, in control rats, an increase in mean arterial pressure, without significant changes in natriuresis or glomerular filtration rate. In chronic bile duct ligation rats, N(G)-nitro-L-arginine methyl ester induced an increase in mean arterial pressure, natriuresis, and kaliuresis, together with a reduction in urinary nitrite excretion and an increase in prostaglandin E2 excretion. Cyclooxygenase inhibition with indomethacin induced in both experimental groups a marked inhibition in urinary prostaglandin E2 excretion without significant changes in Na+ or K+ excretion, and a significant increase in urinary nitrite excretion in control rats. N(G)-Nitro-L-arginine methyl ester in addition to indomethacin prevented the indomethacin-induced increase in nitrite excretion and dramatically reduced sodium excretion in both experimental groups. Thus, the present study suggests that both nitric oxide and cyclooxygenase products interact in the control of urinary sodium excretion and that each system is activated in the absence of the other one. PMID- 10535702 TI - Insulin-sparing effect of hydroxychloroquine in diabetic rats is concentration dependent. AB - To study the effect of hydroxychloroquine (HCQ) on glucose and insulin homeostasis, healthy rats were dosed with 160 mg x kg (-1) x day(-1) of HCQ orally, and streptozocin-induced diabetic rats received 80, 120, and 160 mg x kg( 1) x day(-1) of HCQ, while controls received normal saline. Ten days after treatment with HCQ, healthy animals were challenged intravenously with insulin or glucose, while diabetic rats were given only an i.v. injection of insulin. In healthy rats, the areas within and under the glucose concentration - time curve following insulin and glucose challenge were estimated. In diabetic animals, the areas under the curve for both the percent change in serum glucose from baseline (AUG) and the percent change in serum insulin from baseline (AUI) were used as pharmacodynamic end points. In healthy rats, HCQ did not influence fasting serum glucose concentrations or glycemic profiles following i.v. administration of glucose or insulin. In diabetic rats, AUG and AUI were increased dependent on blood HCQ concentrations. The normal homeostatic mechanisms responsible for insulin-glucose regulation may compensate for possible HCQ-induced reduction of insulin metabolism in healthy rats. The HCQ dose- or concentration-effect relationships for glucose and insulin were linear over the range of HCQ concentrations tested. It is concluded that HCQ significantly elevated insulin blood concentration resulting in reduced glucose levels in a concentration dependent fashion in diabetic rats. HCQ may have therapeutic potential in the treatment of type I and type II diabetes. PMID- 10535703 TI - Hemodynamic and biochemical effects of 100% oxygen breathing in humans. AB - High concentrations of inspired oxygen have been reported to have significant hemodynamic effects that may be related to increased free radical production. If oxygen therapy increases free radical production, it may also modify hemodynamic responses to a nitric oxide donor. Twenty-nine healthy male volunteers were studied using randomized, double-blind, placebo-controlled, crossover designs to determine whether oxygen therapy is associated with hemodynamic and forearm vascular effects. We measured hemodynamic parameters and forearm vascular responses before and 1 h after exposure to 100% oxygen versus medical air. Plasma 8-iso-PGF2alpha and plasma vitamin C were measured to assess the biochemical effects of oxygen administration. Hemodynamic measurements were also made following the acute administration of sublingual nitroglycerin. Oxygen therapy caused no significant change in blood pressure, plasma 8-iso-PGF2alpha, or vitamin C. Oxygen did cause a significant reduction in heart rate and forearm blood flow, and an increase in peripheral vascular resistance. Oxygen caused no change in the hemodynamic response to nitroglycerin. Therefore, in healthy young adults, therapy with 100% oxygen does not affect blood pressure, despite causing an increase in vascular resistance, is not associated with evidence of increased free radical injury, and does not affect the hemodynamic responses to nitroglycerin. PMID- 10535704 TI - Arterial versus venous changes in vascular capacitance during nitroprusside infusion: a vascular modelling study. AB - The distributions of nitroprusside (NP) induced changes in vascular capacitance, arterial versus venous, are unknown. We measured canine ileal arterial and venous pressures and total (isolated loop) vascular volumes (scintigraphy), before and during NP infusion. NP sufficient to decrease perfusion pressure by 30% increased total vascular volume to 111 +/- 3% (+/- SEM) of control (p < 0.01). Increasing flow to restore perfusion pressure increased volume 4% more (p < 0.01). Assuming a two-compartment model and on the basis of the literature data, changes in venous capacitance were estimated and compared with arterial capacitance. During constant-flow perfusion, NP increased venous volume by 10.0% (vs. 18.1%, arterial). When flow was increased to restore pressure, venous volume increased by another 3.7% (vs. 2.6%, arterial). Assuming an original arterial to venous volume ratio of 133/1033, the final, constant-pressure increase in venous volume was almost 4 times the arterial increase. In conclusion, the increase in vascular volume during NP infusion was due primarily to similar-magnitude, active increases in venous and arterial capacitances (i.e., rightward shifts in pressure volume relations). However, as venous volume is so much larger than arterial, the NP-induced increase in venous volume was greater. PMID- 10535705 TI - Tyrosine kinase inhibitors block the glucocorticoid stimulation of prostaglandin endoperoxide H synthase expression in amnion cells. AB - Human amnion cells in primary culture respond to glucocorticoids in a characteristic fashion by the increased expression of the inducible prostaglandin endoperoxide H synthase isoenzyme, PGHS-2. Since PGHS-2 induction by agonists generally involves tyrosine kinases, we examined the possibility that the glucocorticoid stimulation of PGHS-2 in the amnion cells is tyrosine kinase dependent. PGHS-2 expression was stimulated in confluent, serum-starved amnion cells with dexamethasone, and the effect of the tyrosine kinase inhibitors herbimycin A and tyrphostins AG126, AG1288, and A1 on enzyme activity induction was determined. All four inhibitors blocked the increase of PGHS activity in a concentration-dependent manner with IC50 values of 0.077 +/- 0.05, 15.38 +/- 5.14, 20.91 +/- 3.1, and 29.77 +/- 8.21 microM, respectively (mean +/- SE, n = 4). Dexamethasone increased (approximately twofold) the tyrosine phosphorylation of 120-, 110-, and 77-kDa proteins in cell extracts, and herbimycin A selectively blocked the phosphorylation of the 110-kDa phosphoprotein. The stimulation of the steady-state level of PGHS-2 mRNA by dexamethasone was also inhibited by herbimycin A. These results suggest that glucocorticoids induce PGHS-2 expression in amnion cells with the involvement of tyrosine kinase(s). The role of tyrosine kinase dependent mechanisms in the control of amnion cell responsiveness to corticosteroids remains to be established. PMID- 10535706 TI - Increased glucose synthesis in renal tubule fragments from hyperthyroid rats. AB - Rates of glucose synthesis from several substrates were examined in renal tubule fragments from hyperthyroid rats. A hyperthyroid state was induced by daily intraperitoneal injections of thyroxine (T4) (100 microg/100 g body weight) for 14 days. At the end of the experimental period, plasma triiodothyronine and T4 levels were six and eight times higher, respectively, than initial values. Hyperthyroid rats gained less weight and had lower blood glucose despite an increased food intake. In both control and hyperthyroid rats, rates of glucose production by renal tubule fragments were higher with glutamine and glycerol than with lactate, alanine, or glutamate. T4 treatment induced a significant increase in the de novo glucose synthesis from all substrates, except glutamine. The highest percent increase was obtained with alanine (64%), compared with 31-40% for glutamate, lactate, and glycerol. The T4 treatment induced increase in glucose synthesis by renal tubule fragments suggests that renal gluconeogenesis contributes to enhance glucose production in hyperthyroidism. PMID- 10535707 TI - Physiological roles of matrix metalloproteinases: implications for tumor growth and metastasis. AB - Physiological processes involving remodelling of the extracellular matrix, such as wound healing, embryogenesis, angiogenesis, and the female reproductive cycle, require the activity of matrix metalloproteinases (MMPs). This group of proteases degrades basal membranes and connective tissues and plays an essential role in the homeostasis of the extracellular matrix. An imbalance in the expression or activity of MMPs can have important consequences in diseases such as multiple sclerosis, Alzheimer's disease, or the development of cancers. Because of the pathophysiological importance of MMPs, their activity is highly controlled in order to confine them to specific areas. An activation cascade, initiated by the proteolysis of plasminogen, cleaves proMMPs, and every step is controlled by specific activators or inhibitors. MMPs destabilize the organization of the extracellular matrix and influence the development of cancer by contributing to cell migration, tumor cell proliferation, and angiogenesis. Accordingly, these proteases possess an important role in cell-matrix interactions by affecting fundamental processes such as cell differentiation and proliferation. Therefore, the characterization of MMPs involved in specific types and stages of tumors will significantly improve the diagnosis and treatment of these cancers in humans. PMID- 10535709 TI - Sulfated cholecystokinin octapeptide in the rat: pontomedullary distribution and modulation of the respiratory pattern. AB - We performed anatomical and physiological studies to determine the site and actions of sulfated cholecystokinin octapeptide (CCK8-S) on breathing. Peptide locations were determined by combined immunodetection of CCK8-S- containing synaptic varicosities and retrograde labeling of medullary neurons projecting to the ventral respiratory group. Retrogradely labeled neurons and CCK8-S immunolabeled varicosities overlapped within the nuclei of the solitary tract, ventral respiratory group, and the Kolliker-Fuse nucleus. Additional CCK8-S immunoreactive terminals were located in the rostroventrolateral medullary reticular nucleus, lateral paragigantocellular reticular nucleus, and the caudal pontine reticular nucleus. The respiratory effects of CCK8-S, which binds to CCK(A) and CCK(B) receptors, were examined by intravenous injection in adult rats and by bath application in the in vitro neonatal rat brainstem - spinal cord preparation. CCK8-S produced an increase in the mean amplitude of diaphragmatic electromyogram (EMG) of 28 +/- 35% (SD) and a decrease in mean respiratory interval of 13 +/- 4% in vivo. In vitro, CCK8-S significantly increased inspiratory duration and decreased respiratory interval, primarily by shortening expiratory duration. CCK8-unsulfated, a specific agonist for CCK(B) receptors, did not produce these effects. CCK8-S effects in the in vitro preparation were partially blocked by the CCK receptor antagonist lorglumide (final bath concentration 600 nM). These results suggest that CCK8-S modulates the respiratory rhythm via CCK(A) receptors within one or more medullary or pontine respiratory groups in both neonatal and adult rats. PMID- 10535708 TI - Influence of some phospholipase A2 and cytochrome P450 inhibitors on rat arterial smooth muscle K+ currents. AB - The hyperpolarizing factor that is liberated by vascular endothelial cells in response to various agonists, and known to induce relaxation by opening of smooth muscle K+ channels, has been suggested to be a product of cytochrome P450 dependent arachidonic acid metabolism. In this study, the direct influence of two phospholipase A2 inhibitors and of five structurally and mechanistically different cytochrome P450 inhibitors on K+ currents in freshly isolated vascular smooth muscle cells from the rat aorta was investigated. On stepping the cell membrane potential from -70 mV to a series of depolarized test potentials, a noisy outward current developed at test potentials > +10 mV, which showed no appreciable inactivation during the voltage pulse. It was largely abolished by 3 mM external tetraethylammonium chloride (TEA), suggesting that it predominantly consisted of current through large-conductance Ca(2+)-activated K+ channels. The phospholipase A2 inhibitor quinacrine considerably inhibited this TEA-sensitive current, while 4-bromophenacylbromide exerted no effect. The cytochrome P450 inhibitors proadifen and miconazole reversibly decreased the amplitude of I(K), while clotrimazole and 1-aminobenzotriazole had no effect. Conversely, 17 octadecynoic acid increased whole-cell I(K). These results show that some phospholipase A2 and cytochrome P450 inhibitors may interfere with K+ channel activation in the rat arterial smooth muscle cell. The relevance of these findings to studies on the involvement of cytochrome P450 dependent metabolism in the generation of the endothelium-derived hyperpolarizing factor in intact arteries is discussed. PMID- 10535710 TI - Changes in spine and radius bone density during long-term hormone replacement. AB - Lumbar spine and mid-radius bone mineral density was measured repeatedly in 48 postmenopausal women who completed 7 years of taking either a 500 mg x day(-1) calcium supplement (n = 22) or calcium supplementation with hormone replacement therapy. The hormone replacement was either a low dose (n = 15) or a moderate dose (n = regime. The purpose of the measurements was to establish the long-term pattern of change in bone mineral mass produced by continued hormone replacement. The calcium-only group lost bone mineral mass at the radius, while the spine, bone was preserved. Low dose hormone replacement preserved radius bone. Moderate dose replacement increased bone mineral mass at the spine and preserved radius bone. PMID- 10535711 TI - Electrophysiologic properties and ventricular fibrillation in normal and myopathic hearts. AB - This study tests the hypothesis that moderate myocardial dysfunction is associated with altered myocardial anisotropic properties and structurally altered ventricular fibrillation (VF). Mongrel dogs were randomized to either a control group or a group that was rapidly paced at 250 beats/min until the left ventricular ejection fraction was < or = 40%. Changes in anisotropic properties and the electrical characteristics of VF associated with the development of moderate myocardial dysfunction were assessed by microminiature epicardial mapping studies. In vivo conduction, refractory periods, and repolarization times were prolonged in both longitudinal and transverse directions in myopathic animals versus controls. VF was different in myopathic versus control animals. There were significantly more conducted deflections during VF in normal hearts compared with myopathic hearts. Propagated deflection-to-deflection intervals during VF were significantly longer in myopathic hearts compared with controls (125.5 +/- 49.06 versus 103.4 +/- 32.9 ms, p = 0.009). There were no abnormalities in cell size, cell shape, or the number of intercellular gap junctions and there was no detectable change in the expression of the gap junction proteins Cx43 and Cx45. Moderate myocardial dysfunction is associated with significant electrophysiological abnormalities in the absence of changes in myocardial cell morphology or intercellular connections, suggesting a functional abnormality in cell-to-cell communication. PMID- 10535712 TI - Intracellular calcium stores are involved in growth hormone-releasing hormone signal transduction in rat somatotrophs. AB - In rat pituitary somatotrophs, the stimulation of growth hormone secretion by growth hormone-releasing hormone (GHRH) is a Ca(2+)-dependent event involving Ca2+ influx. The presence of calcium-induced calcium release (CICR) Ca2+ stores has been suggested in these cells. The aim of our study was to demonstrate the presence of CICR stores in rat somatotrophs and to determine their function in GHRH Ca2+ signalling. To this end we measured cytosolic free Ca2+ concentration ([Ca2+]i), using indo-1 in purified rat somatotrophs in primary culture, while altering intracellular Ca2+ stores. Ionomycin (10 ttM) or 4-bromo-A23187 (10 ItM), used to mobilise organelle-bound Ca2+, raised [Ca2+]i in the absence of extracellular Ca2+. Caffeine (5 to 50 mM), used to mobilise Ca2+ from CICR stores, transiently raised [Ca2+]i in 65% of cells tested. The response to 40 mM caffeine was abolished when Ca2+ stores were depleted, with 1 microM thapsigargin or with 10 microM ryanodine. All cells that responded to 40 mM caffeine responded to 10 nM GHRH. The [Ca2+]i response to 10 nM GHRH was reversible and repeatable. However, the second response was 38% smaller than the first. Ryanodine treatment abolished the reduction in the second [Ca2+]i response, while thapsigargin increased the reduction by 67%. We conclude that rat somatotrophs possess CICR Ca2+ stores and that they account for 30-35% of the GHRH-induced increase in [Ca2+]i, and that their partial depletion is involved in somatotroph desensitization. PMID- 10535714 TI - Increased flow rate and papaverine increase K+ exchange in perfused rat hind-limb skeletal muscle. AB - This study tested the hypothesis that increases in perfusate flow rate result in increased rates of unidirectional and net K+ transport in rat hind-limb skeletal muscle at rest. Ten neurally and vascularly isolated hind limbs, with arterial and venous catheters placed proximal to the popliteal region, were perfused for 10-min periods at flow rates (presented in a random order) of 0.27, 0.42, 0.63, 0.84, or 1.05 mL x min(-1) x g(-1). Potassium extraction and unidirectional K+ influx were determined using 42K, and arterial perfusion pressure was measured continuously. Increases in flow rate resulted in decreases in K+ extraction and increases in unidirectional K+ influx, unidirectional K+ efflux, and net K+ efflux. The increases in K+ flux were associated with increases in oxygen uptake, glucose uptake, and lactate release. In separate experiments (n = 5), the vasodilator papaverine (10(-4) M) did not further vasodilate the vasculature of resting hind limbs, suggesting that the hind limbs in this preparation were fully vasodilated. Papaverine, at constant flow, resulted in a nearly 1.5-fold increase in K+ extraction, a doubling of unidirectional K+ influx, and increases in unidirectional K+ efflux and net K+ efflux. It is concluded that physiological increases in flow rate result in increases in K+ transport in isolated, perfused rat hind-limb skeletal muscle. Furthermore, papaverine appeared to induce an increase in skeletal muscle membrane permeability to K+. PMID- 10535713 TI - Inhibition of matrix metalloproteinase MMP-2 activates chloride current in human airway epithelial cells. AB - Matrix metalloproteinases (MMPs) are involved in the remodeling and degradation of the extracellular matrix. Recently, it has been found that MMPs also contribute to processes not directly related to tissue remodeling, such as platelet aggregation or degranulation of airway gland cells. Since mucus secretion is closely related to ion channel function, we investigated whether MMPs could also be involved in the regulation of ion channels. We used human airway submucosal cell line Calu-3 to study the effects of MMPs on whole-cell current and transepithelial short-circuit current (I(sc)). Phenanthroline, a specific inhibitor of MMPs, increased whole-cell current with the half-maximally effective dose of 5.2 microM, and reversibly activated I(sc) in transepithelial measurements. Current stimulated by phenanthroline displayed linear current voltage relationships and had inhibitor pharmacology and ion selectivity consistent with cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel activity. Zymography and Western blot showed significant expression of MMP-2 in Calu-3 cells. Moreover, anti-MMP-2 antibodies (1 microg/mL) increased whole-cell current and I(sc), whereas human recombinant MMP-2 (10 ng/mL) reduced it. We also studied the expression of MMPs and the effects of phenanthroline on whole-cell current in A549 cells, which are derived from airway surface epithelium and do not express CFTR Cl- channels. While these cells also showed significant expression of MMP-2, inhibition of this enzyme with phenanthroline exerted no significant effect on whole-cell current. It is concluded that MMP-2 is involved in the regulation of CFTR Cl- channels in human airways. PMID- 10535715 TI - Resting metabolic rate and thermic effects of a sucrose-sweetened soft drink during the menstrual cycle in young Chinese women. AB - The resting metabolic rate (RMR) and thermic effects (TEF) of a sucrose-sweetened soft drink in a group (n = 19) of ovulating young Chinese women were determined by indirect calorimetry in the midfollicular and midluteal phases of the menstrual cycle. Urinary luteinizing hormone surge was used to confirm ovulation. The RMR was measured twice in each phase and found to be similar (F(1,18) = 0.863) across the follicular (5018 kJ/24 h) and the luteal (5098 kJ/24 h) phases. Within each phase and on separate days, subjects were given water (280 mL) or sucrose-sweetened soft drink (539 kJ). Soft drink, but not water, consumption increased energy expenditure over a period of 45 min. Compared with the follicular phase, a small but significant increase in TEF (kJ/45 min) was observed in the luteal phase (t = 2.434, p < 0.05). Energy expenditure after drinking the soft drink, however, was similar in the two phases. RMR was positively correlated with TEF (r = 0.613, p < 0.01) and net TEF (r = 0.648, p < 0.005) in the luteal but not the follicular phase. In ovulating women, the thermic effect of sucrose is influenced by the phase of the menstrual cycle. PMID- 10535716 TI - Comparison of low-molecular-weight heparin, administered primarily at home, with unfractionated heparin, administered in hospital, and subcutaneous heparin, administered at home for deep-vein thrombosis. AB - In this study, 294 patients with acute proximal DVT (deep venous thrombosis) were randomly assigned to receive intravenous standard heparin in the hospital (98 patients) or low-molecular-weight heparin (LMWH) (nadroparin 0.1 mL [equivalent to 100 AXa IU] per kg of body weight subcutaneously twice daily) administered primarily at home (outpatients) or alternatively in hospital (97 patients) or subcutaneous calcium heparin (SCHep) (99 patients, 0.5 mL bid) administered directly at home. The study design allowed outpatients taking LMWH heparin to go home immediately and hospitalized patients taking LMWH to be discharged early. Patients treated with standard heparin or LMWH received the oral anticoagulant starting on the second day, and heparin was discontinued when the therapeutic range (INR 2-3) had been reached. Anticoagulant treatment was maintained for 3 months. Patients treated with SCHep were injected twice daily for 3 months without oral anticoagulants. Patients were evaluated for inclusion and follow-up with color duplex scanning. Venography was not used. In case of suspected pulmonary embolism (PE) a ventilatory-perfusional lung scan was performed. Endpoints of the study were recurrent or extension of DVT, bleeding, the number of days spent in hospital, and costs of treatments. Of the 325 patients included, 294 completed the study. Dropouts totaled 31 (10.5%); six of the 325 included patients (1.8%) died from the related, neoplastic illness. Recurrence or extension of DVT was observed in 6.1% of patients in the LMWH group, in 6.2% in the standard heparin group, and in 7.1% in the SCHep group. Most recurrences (11/17) were in the first month in all groups. Bleedings were all minor, mostly during hospital stay. Hospital stay in patients treated with LMWH was 1.2+/-1.4 days in comparison with 5.4+/-1.2 in those treated with standard heparin. There was no hospital stay in the SCHep group. Average treatment costs in 3 months in the standard heparin group (US $2,760) were considered to be 100%; in comparison costs in the LMWH group was 28% of the standard heparin and 8% in the SCHep group. This study indicated that LMWH and SCHep can be used safely and effectively to treat patients with proximal DVT at home at a lower cost. PMID- 10535717 TI - Surgical treatment of interventional coronary angiographic accidents. AB - Newer methodologies have increased the incidence of coronary interventions. At the authors' institution, 5,614 coronary interventional procedures (28% of all catheterizations) were performed over a 3-year period, from 1995 to 1997. Eighty one patients (1.4%) suffered angiographic accidents, including coronary artery dissection, free rupture, tamponade, foreign body embolism, and wire entrapment, and were retrospectively reviewed. All patients were taken for emergency surgery in less than 4 hours. The mean age was 61.2 years, 44 (54%) were men, and 37 (46%) were in cardiogenic shock at the time of surgery. Fifty-seven patients (70%) had intraaortic balloon counterpulsation. The number of previous cardiac interventions ranged from one to four with a mean of 1.9. One to five bypass grafts (mean, 2.2) were performed, and three patients required temporary ventricular assist devices. There were six deaths for a 30-day mortality rate of 7.4%. Thirty-two patients (39.5%) suffered significant morbidity, including cerebrovascular accidents, and renal and respiratory failure. Perioperative myocardial infarctions were diagnosed in 39 (48%) patients. Average length of stay was 12.1 days. One-year survival was satisfactory at 90% (73/81), with 56 survivors (77%) regaining normal everyday activity. Early surgical intervention, rapid revascularization, and temporary mechanical support are keys to low mortality in this high-risk group. Identification of high-risk interventions and significant comorbid conditions, with concomitant surgical consultation, need to be pursued to reduce the high morbidity rate. PMID- 10535718 TI - Daytime-selective antihypertensive activity of celiprolol. AB - Day-activity rhythms of heart rate and blood pressure are thought to be mediated mainly through the sympathetic nervous system and may have greater amplitudes in patients with hypertension owing to increased daytime and largely normal nighttime values. Drug-induced nighttime hypotension in patients with chronic hypertension has been associated with the precipitation of cardiac failure and a fall in cerebral flow. The authors examined the effects of a single dose and of a 4-week treatment with different classes of antihypertensive drugs on ambulatory blood pressure (ABP) in 10 patients with mild hypertension. Data were assessed by polynomial analysis (Harvard Graphics 3). A single oral dose of enalapril 10 mg, amlodipine 5 mg, carvedilol 25 mg, and celiprolol 200 mg produced a mean reduction of 24-hour ABP compared to placebo of, respectively, 24/11, 11/5, 13/6, and 12/5 mm Hg (p values between <0.02 and <0.001). With enalapril, amlodipine, and carvedilol, between-subject variability contributed significantly to the overall variability in the measurements (p values between 0.05 and 0.01 versus zero), whereas with celiprolol this was not so. Although the beta blockers reduced daytime blood pressures similarly to the ACE inhibitor or the calcium channel blocker, they did not reduce nighttime blood pressures. These results were confirmed by an 8-week crossover trial comparing enalapril 10 mg daily with celiprolol 200 mg daily in the same group of patients. The authors conclude (1) that beta blockers produce a more stable reduction of blood pressure in patients with mild hypertension less affected by pressor effects through the sympathetic nervous system; (2) that beta blockers, unlike ACE inhibitors and calcium channel blockers, do not give rise to nighttime hypotension in this category of patients; and (3) that the selective beta blocker celiprolol may even perform better in these respects than the nonselective beta blocker carvedilol. PMID- 10535719 TI - Doppler echocardiographic study of right ventricular systolic performance in inferior myocardial infarction. AB - Twenty-four male patients with myocardial infarction (MI) without clinical and electrocardiographic signs of right ventricular (RV) involvement were selected to enter the study. All the patients were divided into two groups: Group I consisted of 12 patients with anterior MI and infarct-related left anterior descending artery and Group II included 12 patients with inferior MI and infarct-related right coronary artery. Patients of Group II had higher right atrial pressure and right atrial pressure/pulmonary capillary wedge pressure ratio (p<0.01, p<0.01) and lower values of pulmonary flow velocity, mean acceleration, and pulmonary flow velocity2/acceleration time ratio than patients of Group I (p<0.01, p<0.01, p<0.01, respectively). Pulmonary flow indices correlated inversely and significantly with hemodynamic dysfunction in patients with inferior myocardial infarction and right coronary proximal lesions (p<0.01). PMID- 10535720 TI - Pharmacogenetic analysis of the effect of angiotensin-converting enzyme inhibitor on restenosis after percutaneous transluminal coronary angioplasty. AB - Angiotensin-converting enzyme (ACE) inhibitors are reported to prevent neointimal formation after balloon injury in animal models, but in most prospective studies in humans, ACE inhibitors failed to prevent restenosis after percutaneous transluminal coronary angioplasty (PTCA). The ACE genotype assigned by an insertion/deletion (I/D) polymorphism is known to affect the potency of ACE inhibitors in several renal diseases. The authors attempted to clarify whether the effect of ACE inhibitors on restenosis might be modified by the ACE genotype. A total of 126 patients was randomly and prospectively assigned to the control group and the imidapril group. In the imidapril group, patients received 5 mg imidapril daily, starting 1 day before PTCA and continuing for 3 to 6 months. Forty-six control (65 vessels) and 32 imidapril patients (43 vessels) completed the study. The minimal lumen diameter before and after the procedure did not differ significantly among the groups with the three genotypes (II, ID, and DD) in both the control and imidapril groups. Late luminal loss during the follow-up period was not related to the ACE genotype in the control group but was significantly related in the imidapril group (II, 0.63+/- 0.19 mm; ID + DD, 1.12+/-0.14 mm, p<0.05). Furthermore, in the II genotype, imidapril significantly reduced late loss and restenosis rate as defined by most of the frequently used definitions. In conclusion the ACE I/D polymorphism may influence the effect of ACE inhibitors in preventing restenosis after PTCA. PMID- 10535721 TI - Hyperhomocyst(e)inemia is associated with carotid atherosclerosis. AB - The atherogenicity of homocyst(e)ine--H(e) --emerged from many studies showing an association between moderately elevated levels and vascular occlusive disease. The aim of this study was to evaluate whether high homocyst(e)ine levels were associated with carotid atherosclerosis. Carotid atherosclerosis was defined as an intimal media thickness of internal and carotid bifurcation of at least 2 mm on the near and far walls as determined by B-mode ultrasonography. The study population included 91 patients: group 1 (61% males, mean age 64+/-10 years, 57% with history of hypertension) with ultrasound evidence of carotid atherosclerosis and 100 with normal carotid walls--group 2 (36% males, mean age 52+/-15 years, 27% with history of hypertension). Homocyst(e)ine levels (mol/L) were determined by high-performance liquid chromatography with a fluorescent detector. Body mass index, dyslipidemia, smoking, diabetes, serum creatinine, plasma folic acid and vitamin B12 were not significantly different in the two groups. Homocyst(e)ine levels (micromol/L) were significantly higher in patients with carotid ather osclerosis than in those with normal arteries (11.7+/-6.5 micromol/L, 95% CI 10.4 13.1 vs 8.07+/-4.4 micromol/L, 95% CI 7.2-8.9, p<0.0001). By multiple regression analysis H(e) levels were positively correlated with male gender (p<0.02), age (p<0.001), and negatively with folic acid (p<0.0001). By logistic regression the independent predictors of carotid atherosclerosis were male gender (OR 2.65), hypertension (OR 2.55), age (x10 years, OR 2.15) and H(e) levels (x1 micromol/L, OR 1.11). This study confirmed homocyst(e)ine is associated with carotid atherosclerosis. Consequently the authors recommend H(e) levels be screened in all patients at risk for atherosclerosis. PMID- 10535722 TI - Air plethysmography in chronic venous insufficiency: clinical diagnosis and quantitative assessment. AB - To define the role of air plethysmography (APG) in the clinical diagnosis and quantitative assessment of chronic venous insufficiency (CVI), APG studies were performed on 582 limbs in 291 patients with signs and symptoms of CVI. One hundred and thirty-one limbs were classified into group I (no evidence of CVI), 291 into group II (mild CVI), and 160 into group III (advanced CVI). On APG, the mean venous filling index (VFI) was 1.45 mL/s, 3.90 mL/s, and 5.25 mL/s in groups I, II, and III respectively (p<0.05). The mean ejection fraction (EF) and mean residual volume fraction (RVF) also showed significant differences between the limbs with CVI and the contralateral normal limbs, but the values were similar for the different severities of CVI limbs. The amount of overlap in VFI, EF, and RVF values among the clinical groups was considerable. Discrimination analysis derived a VFI value of 2.67 mL/s as a cutoff point between normal limbs and limbs with CVI, with a positive predictive value of 96%. In conclusion, APG is a simple and noninvasive test for quantitative assessment of the different components of CVI, valvular reflux, and calf muscle pump function. However, only VFI correlated significantly with the severity of CVI. VFI, with its high positive predictive value, may be useful in diagnosis of CVI, and it may serve as an objective quantitative measurement for monitoring the effect of treatment. PMID- 10535724 TI - Magnetic resonance imaging in internal carotid artery agenesis with computed tomography and angiographic correlation--case reports. AB - The authors present two cases of agenesis of the internal carotid artery (ICA) discovered incidentally on magnetic resonance imaging and confirmed on computed tomography, magnetic resonance angiography, and conventional angiography. They also propose a clinical algorithm for the workup of patients with suspected absence of the ICA. PMID- 10535723 TI - Tissue characterization of myocardial cells by use of high-frequency acoustic microscopy: differential myocyte sound speed and its transmural variation in normal, pressure-overload hypertrophic, and amyloid myocardium. AB - The purpose of the present study was to evaluate the acoustic properties of myocytes in normal, pressure-overload hypertrophic, and amyloid myocardium. Myocardial tissue specimens at autopsy were obtained from 10 subjects without cardiovascular disease, six patients with left ventricular (LV) hypertrophy, and six patients with cardiac amyloidosis. Sound speed of myocytes was measured at subendocardial and subepicardial regions in myocardium by use of a high-frequency (450 MHz) acoustic microscope. In normal myocardium, the sound speed of myocytes was significantly higher in subendocardial region (1,728+/-19 m/sec) than in subepicardial region (1,645+/-22 m/sec) (p<0.0001). A significantly higher sound speed of myocytes was observed in the subendocardial region in LV hypertrophic myocardium (1,779+/-19 m/sec) than that in normal myocardium (p<0.001). In amyloid myocardium, a significantly lower sound speed of myocytes was observed in subendocardial (1,560+/-8 m/sec) and subepicardial (1,594+/-48 m/sec) regions than that in respective regions of the normal myocardium (p<0.0001 and p<0.05, respectively). Transmural variation in sound speed of myocytes measured by high frequency acoustic microscopy existed in normal left ventricle. The differential myocyte sound speed and its transmural variation was observed in LV hypertrophic and amyloid myocardium as compared with normal myocardium. High-frequency acoustic microscopy can be a promising technique for myocardial tissue characterization at the myocyte level. PMID- 10535725 TI - An unusual case of multiple right atrial thrombi in a patient with a dual-chamber pacemaker--a case report. AB - The authors present an unusual case of multiple large atrial thrombi attached to permanent pacemaker leads identified by transesophageal echocardiography. Pathogenesis, clinical implications, and therapeutic options of pacemaker thrombi are discussed. PMID- 10535726 TI - Regression of large atrial thrombi and coronary neovascularizations with conventional anticoagulation in mitral stenosis--a case report. AB - The authors report a case of angiographically documented multiple coronary neovascularizations originating from the left circumflex artery (LCX) and coursing toward multiple thrombi located in the left atrium in a patient with severe mitral stenosis. The thrombi as well as the neovascularizations underwent near-complete resolution with 4 weeks' anticoagulation therapy with warfarin maintaining an international normalization ratio of 3.5. Percutaneous mitral balloon valvuloplasty was performed successfully without complications. PMID- 10535727 TI - Giant unruptured aneurysm of the thoracic aorta--a case report. AB - An asymptomatic 88-year-old woman underwent a screening medical examination. The chest x-ray film showed a large mediastinal mass with calcification. Both chest computed tomography and nuclear magnetic resonance imaging revealed an unruptured aortic aneurysm, predominantly affecting the ascending aorta and the proximal part of the aortic arch. Its maximum diameter was 10.5 cm. An ascending aortic aneurysm more than 10 cm in diameter is very rare. She died of acute pulmonary embolism unrelated to the aneurysm, and autopsy indicated that the etiology of the aneurysm was atherosclerotic degeneration. Retrospectively, the natural progression of the aneurysm was able to be followed on a series of chest x-ray films obtained over 18 years. PMID- 10535728 TI - Regarding "Bifurcating aneurysm of the left main coronary artery involving left anterior descending and left circumflex arteries". PMID- 10535729 TI - Electron avenue: pathways of disulfide bond formation and isomerization. PMID- 10535730 TI - Catch the mu1B train to the basolateral surface. PMID- 10535731 TI - The rde-1 gene, RNA interference, and transposon silencing in C. elegans. AB - Double-stranded (ds) RNA can induce sequence-specific inhibition of gene function in several organisms. However, both the mechanism and the physiological role of the interference process remain mysterious. In order to study the interference process, we have selected C. elegans mutants resistant to dsRNA-mediated interference (RNAi). Two loci, rde-1 and rde-4, are defined by mutants strongly resistant to RNAi but with no obvious defects in growth or development. We show that rde-1 is a member of the piwi/sting/argonaute/zwille/eIF2C gene family conserved from plants to vertebrates. Interestingly, several, but not all, RNAi deficient strains exhibit mobilization of the endogenous transposons. We discuss implications for the mechanism of RNAi and the possibility that one natural function of RNAi is transposon silencing. PMID- 10535732 TI - Mut-7 of C. elegans, required for transposon silencing and RNA interference, is a homolog of Werner syndrome helicase and RNaseD. AB - While all known natural isolates of C. elegans contain multiple copies of the Tc1 transposon, which are active in the soma, Tc1 transposition is fully silenced in the germline of many strains. We mutagenized one such silenced strain and isolated mutants in which Tc1 had been activated in the germline ("mutators"). Interestingly, many other transposons of unrelated sequence had also become active. Most of these mutants are resistant to RNA interference (RNAi). We found one of the mutated genes, mut-7, to encode a protein with homology to RNaseD. This provides support for the notion that RNAi works by dsRNA-directed, enzymatic RNA degradation. We propose a model in which MUT-7, guided by transposon-derived dsRNA, represses transposition by degrading transposon-specific messengers, thus preventing transposase production and transposition. PMID- 10535733 TI - Heterozygous germline mutations in the p53 homolog p63 are the cause of EEC syndrome. AB - EEC syndrome is an autosomal dominant disorder characterized by ectrodactyly, ectodermal dysplasia, and facial clefts. We have mapped the genetic defect in several EEC syndrome families to a region of chromosome 3q27 previously implicated in the EEC-like disorder, limb mammary syndrome (LMS). Analysis of the p63 gene, a homolog of p53 located in the critical LMS/EEC interval, revealed heterozygous mutations in nine unrelated EEC families. Eight mutations result in amino acid substitutions that are predicted to abolish the DNA binding capacity of p63. The ninth is a frameshift mutation that affects the p63alpha, but not p63beta and p63gamma isotypes. Transactivation studies with these mutant p63 isotypes provide a molecular explanation for the dominant character of p63 mutations in EEC syndrome. PMID- 10535734 TI - Building a replisome from interacting pieces: sliding clamp complexed to a peptide from DNA polymerase and a polymerase editing complex. AB - We have solved the crystal structures of the bacteriophage RB69 sliding clamp, its complex with a peptide essential for DNA polymerase interactions, and the DNA polymerase complexed with primer-template DNA. The editing complex structure shows a partially melted duplex DNA exiting from the exonuclease domain at an unexpected angle and significant changes in the protein structure. The clamp complex shows the C-terminal 11 residues of polymerase bound in a hydrophobic pocket, and it allows docking of the editing and clamp structures together. The peptide binds to the sliding clamp at a position identical to that of a replication inhibitor peptide bound to PCNA, suggesting that the replication inhibitor protein p21CIP1 functions by competing with eukaryotic polymerases for the same binding pocket on the clamp. PMID- 10535735 TI - Crystal structure of the helicase domain from the replicative helicase-primase of bacteriophage T7. AB - Helicases that unwind DNA at the replication fork are ring-shaped oligomeric enzymes that move along one strand of a DNA duplex and catalyze the displacement of the complementary strand in a reaction that is coupled to nucleotide hydrolysis. The helicase domain of the replicative helicase-primase protein from bacteriophage T7 crystallized as a helical filament that resembles the Escherichia coli RecA protein, an ATP-dependent DNA strand exchange factor. When viewed in projection along the helical axis of the crystals, six protomers of the T7 helicase domain resemble the hexameric rings seen in electron microscopic images of the intact T7 helicase-primase. Nucleotides bind at the interface between pairs of adjacent subunits where an arginine is near the gamma-phosphate of the nucleotide in trans. The bound nucleotide stabilizes the folded conformation of a DNA-binding motif located near the center of the ring. These and other observations suggest how conformational changes are coupled to DNA unwinding activity. PMID- 10535736 TI - Essential role of phosphoinositide metabolism in synaptic vesicle recycling. AB - Growing evidence suggests that phosphoinositides play an important role in membrane traffic. A polyphosphoinositide phosphatase, synaptojanin 1, was identified as a major presynaptic protein associated with endocytic coated intermediates. We report here that synaptojanin 1-deficient mice exhibit neurological defects and die shortly after birth. In neurons of mutant animals, PI(4,5)P2 levels are increased, and clathrin-coated vesicles accumulate in the cytomatrix-rich area that surrounds the synaptic vesicle cluster in nerve endings. In cell-free assays, reduced phosphoinositide phosphatase activity correlated with increased association of clathrin coats with liposomes. Intracellular recording in hippocampal slices revealed enhanced synaptic depression during prolonged high-frequency stimulation followed by delayed recovery. These results provide genetic evidence for a crucial role of phosphoinositide metabolism in synaptic vesicle recycling. PMID- 10535737 TI - A novel clathrin adaptor complex mediates basolateral targeting in polarized epithelial cells. AB - Although polarized epithelial cells are well known to maintain distinct apical and basolateral plasma membrane domains, the mechanisms responsible for targeting membrane proteins to the apical or basolateral surfaces have remained elusive. We have identified a novel form of the AP-1 clathrin adaptor complex that contains as one of its subunits mu1B, an epithelial cell-specific homolog of the ubiquitously expressed mu1A. LLC-PK1 kidney epithelial cells do not express mu1B and missort many basolateral proteins to the apical surface. Stable expression of mu1B selectively restored basolateral targeting, improved the overall organization of LLC-PK1 monolayers, and had no effect on apical targeting. We conclude that basolateral sorting is mediated by an epithelial cell-specific version of the AP-1 complex containing mu1B. PMID- 10535738 TI - Distinct mechanisms promote polarity establishment in carpels of Arabidopsis. AB - Lateral organs of plants display asymmetry with abaxial identity being specified by members of the Arabidopsis YABBY gene family. Mutations in CRABS CLAW, the founding family member, display ectopic formation of adaxial carpel tissues only when the functions of other genes, such as GYMNOS or KANADI, are also compromised. Mutations in these genes alone do not result in loss of polar differentiation, and therefore, they act redundantly with CRABS CLAW to establish polarity. As GYMNOS encodes a uniformly expressed homolog of the chromatin remodeling protein, Mi2, we argue that the unique genetic interactions do not reflect a molecular redundancy. Rather, CRABS CLAW regulates transcription spatially, whereas GYMNOS regulates downstream targets temporally to ensure proper differentiation of the carpels. PMID- 10535739 TI - Oxygen regulation of airway branching in Drosophila is mediated by branchless FGF. AB - The Drosophila tracheal (respiratory) system is a tubular epithelial network that delivers oxygen to internal tissues. Sprouting of the major tracheal branches is stereotyped and controlled by hard-wired developmental cues. Here we show that ramification of the fine terminal branches is variable and regulated by oxygen, and that this process is controlled by a local signal or signals produced by oxygen-starved cells. We provide evidence that the critical signal is Branchless (Bnl) FGF, the same growth factor that patterns the major branches during embryogenesis. During larval life, oxygen deprivation stimulates expression of Bnl, and the secreted growth factor functions as a chemoattractant that guides new terminal branches to the expressing cells. Thus, a single growth factor is reiteratively used to pattern each level of airway branching, and the change in branch patterning results from a switch from developmental to physiological control of its expression. PMID- 10535740 TI - A transient, neuron-wide form of CREB-mediated long-term facilitation can be stabilized at specific synapses by local protein synthesis. AB - In a culture system where a bifurcated Aplysia sensory neuron makes synapses with two motor neurons, repeated application of serotonin (5-HT) to one synapse produces a CREB-mediated, synapse-specific, long-term facilitation, which can be captured at the opposite synapse by a single pulse of 5-HT. Repeated pulses of 5 HT applied to the cell body of the sensory neuron produce a CREB-dependent, cell wide facilitation, which, unlike synapse-specific facilitation, is not associated with growth and does not persist beyond 48 hr. Persistent facilitation and synapse-specific growth can be induced by a single pulse of 5-HT applied to a peripheral synapse. Thus, the short-term process initiated by a single pulse of 5 HT serves not only to produce transient facilitation, but also to mark and stabilize any synapse of the neuron for long-term facilitation by means of a covalent mark and rapamycin-sensitive local protein synthesis. PMID- 10535741 TI - Insulin resistance and antidiabetic drugs. AB - Insulin resistance describes an impaired biological response to insulin, which underpins the development of type 2 (non-insulin-dependent) diabetes mellitus (T2DM). Initially, insulin resistance causes a compensatory hyperinsulinaemia, which gives way to pancreatic beta-cell failure. Insulin resistance and hyperinsulinaemia conspire together in the development of a diverse collection of risk factors for coronary heart disease, namely obesity, T2DM, dyslipidaemia, hypertension, atherosclerosis, and a pro-coagulant state. This collection of factors is commonly found in T2DM patients, and is recognised as the Insulin Resistance Syndrome or Syndrome X. By targeting insulin resistance as a treatment strategy for T2DM, it should be possible to broaden the potential benefits, so that improved glycaemic control is complemented with improvements to other components of Syndrome X. At present, metformin and thiazolidinediones are the only therapies for T2DM that directly address aspects of insulin resistance. Increasing awareness of the clinical implications of insulin resistance, and increasing knowledge of the cellular basis of insulin resistance, provide the rationale and a means for developing an anti-insulin resistance approach to the treatment of T2DM. PMID- 10535742 TI - HIV-1-trans-activating (Tat) protein: both a target and a tool in therapeutic approaches. AB - Tat proteins (trans-activating proteins) are present in all known lentiviruses and are early RNA binding proteins that regulate transcription. Tat from the human immunodeficiency virus type-1 is a protein comprising 86 amino acids and encoded by 2 exons. The first 72 amino acids are encoded by exon 1 and exhibit full trans-activating activity. The second exon encodes a 14-amino-acid C terminal sequence that is not required for trans-activation but does contain an RGD motif, which is important in binding to alphavbeta3 and alpha5beta1 integrins. Tat has an unusual property for a transcription factor; it can be released and enter cells freely, yet still retain its activity, enabling it to up regulate a number of genes. Tat also has an angiogenic effect; it is a potent growth factor for Kaposi sarcoma-derived spindle cells, and, separately, it has been shown to bind to a specific receptor, Flk-1/KDR, on vascular smooth muscle cells, as well as to integrin-like receptors present on rat skeletal muscle cells and the lymphocyte cell line H9. It appears that the basic domain of tat is important, not only for translocation but also for nuclear localisation and trans activation of cellular genes. As such, targeting of tat protein or, more simply, the basic domain provides great scope for therapeutic intervention in HIV-1 infection. There is also opportunity for tat to be used as a molecular tool; the protein can be manipulated to deliver non-permeable compounds into cells, an approach that already has been employed using ovalbumin, beta-galactosidase, horseradish peroxidase, and caspase-3. PMID- 10535743 TI - Isolation and characterization of a thymidylate synthase-deficient human colon tumor cell line. AB - Following mutagenesis of the human colorectal tumor cell line HCT C with ethyl methanesulfonate, clonal sublines were isolated that survived on medium toxic to cells expressing thymidylate synthase (TS). The subline exhibiting the lowest TS activity, designated as C18, was characterized. Extracts from C18 cells were mixed with extracts from parental C cells to determine whether the TS-deficient phenotype is trans-acting. No effect was observed on the activity of TS in parental extracts. The levels of functional TS in C18 cells were analyzed by the binding of the mechanism-based inhibitor 5-fluoro-2'-deoxyuridylate (FdUMP) under conditions that allowed for the detection of 10 fmol of TS. Only a low level of FdUMP-TS complexes was detected in C18 extracts. The level of TS expression in C18 cells was similar to that in parental C cells, as indicated by immunoblot and RNA analyses. DNA sequence analysis of TS cDNA from C18 cells revealed the existence of a point mutation (C-->T) at nucleotide 647 that predicts the replacement of Ser216 by a leucine residue. That the C18 cell line was homozygous for this mutation was indicated by restriction fragment-length polymorphism analysis and by primer extension analysis. To provide additional evidence that substitution of Ser216 by a leucine residue created a defective protein, a TS deficient bacterial strain was transformed with an expression vector containing the mutated human TS cDNA. The transformed strain exhibited thymidine auxotrophy, indicating that the mutant TS (Leu216) is nonfunctional. PMID- 10535744 TI - Development and validation of an intact cell assay for protein tyrosine phosphatases using recombinant baculoviruses. AB - We have developed an intact cell assay to be used in the direct quantitation of protein tyrosine phosphatase (PTP) activity. Utilizing the baculovirus expression system, the assay readily allows for a direct activity readout for PTPs such as PTP1B or CD45. Infected Sf9 cells expressing either full-length PTP1B, full length CD45, CD45 catalytic domain, or hCOX-1 (mock-infected) are harvested 29 hr post-infection, at which time cells are viable and the expressed proteins are processed, as well as localized to their predicted subcellular compartments. Assays are carried out in a 96-well format, with cells expressing the PTP of interest. Cells are preincubated with or without inhibitor and challenged with substrate, and the phosphatase activity is determined spectrophotometrically by monitoring the conversion of p-nitrophenyl phosphate to p-nitrophenol at OD405. Documented PTP inhibitors have been used to validate this assay system. This study demonstrates that a direct readout of PTP activity in intact cells can be achieved, thus providing a useful cell-based screen for determining selective inhibitors of PTPs. PMID- 10535745 TI - Protective effect of ethanol against acetaminophen-induced hepatotoxicity in mice: role of NADH:quinone reductase. AB - The role of NAD(P)H:quinone reductase (QR; EC 1.6.99.2) in the alcohol-derived protective effect against hepatotoxicity caused by acetaminophen (APAP) was studied. In mice pretreated with dicoumarol (30 mg/kg), an inhibitor of QR, hepatic necrosis caused by APAP (400 mg/kg) was potentiated. Hepatocellular injuries induced by APAP, as assessed by liver histology, serum aminotransferase activities, hepatic glutathione (reduced and oxidized) contents, and liver microsomal aminopyrine N-demethylase activities, all were potentiated by pretreatment of mice with dicoumarol. Even in mice given APAP and ethanol (4 g/kg), in which APAP-inducible hepatic necrosis was abolished, the dicoumarol pretreatment again produced moderate hepatotoxicity and reversed the protective effect of ethanol. In mice pretreated with dicoumarol and ethanol, levels of APAP in blood and bile fluid between 90 and 240 min were higher than those in mice given ethanol. However, the biliary contents of sulfate and glucuronide conjugates of APAP were much lower than those in the ethanol group, particularly at early time points. In contrast, the biliary level of APAP-cysteine conjugate, which in the ethanol group was at its basal level, was increased maximally in the dicoumarol-pretreated mice. In the mice given dicoumarol and ethanol, the biliary APAP-cysteine conjugate level was increased moderately. These results suggest that ethanol inhibited not only the microsomal (CYP2E1 mediated) formation of a toxic quinone metabolite from APAP, but also accelerated the conversion of the toxic quinone metabolite produced back to APAP by stimulating cytoplasmic QR activity. In the presence of dicoumarol, however, QR activity was inhibited, and conversion of the toxic quinone metabolite back to APAP became inhibited and diminished the alcohol-dependent protective effect against APAP-induced hepatic injury. PMID- 10535746 TI - Inhibition of select mitochondrial enzymes in PC12 cells exposed to S-(1,1,2,2 tetrafluoroethyl)-L-cysteine. AB - Many halogenated foreign compounds are detoxified by conversion to the corresponding cysteine S-conjugate, which is N-acetylated and excreted. However, several halogenated cysteine S-conjugates [e.g. S-(1,1,2,2-tetrafluoroethy)-L cysteine (TFEC)] are converted to mitochondrial toxicants by cysteine S-conjugate beta-lyases. In the present work, we showed that TFEC appreciably inactivated highly purified alpha-ketoglutarate dehydrogenase complex (KGDHC) in the presence of a cysteine S-conjugate beta-lyase. Incubation of PC12 cells (which contain endogenous cysteine S-conjugate beta-lyase activity) with TFEC led to a concentration- and time-dependent loss of endogenous KGDHC activity. A 24-hr exposure to 1 mM TFEC decreased KGDHC activity in the cells by 90%. Although treatment with TFEC did not inhibit intrinsic pyruvate dehydrogenase complex (PDHC) activity, it inhibited dichloroacetate/Mg2+-mediated activation/dephosphorylation of PDHC in the PC12 cells by 90%. To determine the selectivity of enzymes targeted by TFEC, several cytosolic and mitochondrial enzymes involved in energy metabolism [malate dehydrogenase, glyceraldehyde 3 phosphate dehydrogenase, glutamate dehydrogenase, lactate dehydrogenase, cytosolic and mitochondrial aspartate aminotransferases (AspAT)] were also assayed in the PC12 cells exposed to 1 mM TFEC for 24 hr. Of these enzymes, only mitochondrial AspAT, a key enzyme of the malate-aspartate shuttle, was inhibited. The present results demonstrate a selective vulnerability of mitochondrial enzymes to toxic cysteine S-conjugates. The data indicate that TFEC may be a useful cellular/mitochondrial toxicant for elucidating the consequences of the diminished mitochondrial function that accompanies numerous neurodegenerative diseases. PMID- 10535747 TI - The integrin specificity of human recombinant osteopontin. AB - The ability of full-length human recombinant osteopontin (OPN) to support the adhesion of various alphav integrin-expressing cell lines was determined in order to characterize its integrin selectivity. The identity of this protein was assessed by cDNA sequence and mass spectroscopic analysis, and confirmed as full length OPN. Neither the human embryonic kidney 293 cell line, which expresses the alphavbeta1 integrin, nor the human colonic adenocarcinoma HT-29 cell line, which expresses the alphavbeta5 integrin, were able to adhere to OPN; both of these cell lines are deficient in the beta3 subunit. In contrast, an alphavbeta3 integrin-expressing cell line, SK-MEL-24, was able to adhere to OPN in an arginine-glycine-aspartic acid dependent manner. In addition, this OPN-mediated cellular adhesion was completely blocked with an anti-alphavbeta3 integrin antibody (LM609), confirming that only the alphavbeta3 integrin mediated this cellular adhesion. These data demonstrate that, at least among the alphav integrins, only the alphavbeta3 is able to support cellular adhesion to osteopontin. This finding may have implications for the design of therapeutics targeting OPN-integrin interactions. PMID- 10535748 TI - Increased toxicity of cocaine on human hepatocytes induced by ethanol: role of GSH. AB - Increased toxicity of cocaine to human hepatocytes is observed when cells are simultaneously incubated with ethanol. Ethanol might exacerbate cocaine hepatocyte toxicity by three different pathways: a) by increasing the oxidative metabolism of cocaine and hence the oxidative damage; b) by the formation of a more toxic metabolite, namely cocaethylene; or c) by decreasing the defence mechanisms of the cell (i.e. GSH). In the present study, experiments were conducted to investigate the feasibility of these hypotheses. In hepatocytes preincubated for 48 hr with ethanol, neither significant changes in cocaine metabolism nor cytotoxicity were found despite differences in hepatocyte p nitrophenol hydroxylase (largely CYP2E1 activity). Cocaethylene, the transesterification product of cocaine and ethanol, was found to be more toxic than cocaine for human hepatocytes (3x). However, the small amount formed when human hepatocytes were incubated with cocaine and ethanol would hardly explain the increased toxicity observed. On the other hand, the simultaneous presence of cocaine and ethanol caused a sustained decline in the intracellular GSH content that was larger than that observed in cocaine- or ethanol-treated cultures. Parallel to this phenomenon, a significant increase in lipid peroxidation was observed, as compared to cells treated with equimolar amounts of cocaine, ethanol, or cocaethylene. Finally, depletion of hepatocyte GSH with diethylmaleate down to levels similar to those found in ethanol-treated cells made hepatocytes more susceptible to cocaine. Taken together, the results of this research suggest that by decreasing GSH levels, ethanol makes human hepatocytes more sensitive to cocaine-induced oxidative damage. PMID- 10535749 TI - Differential phosphorylation of sites in the linker region of P-glycoprotein by protein kinase C isozymes alpha, betaI, betaII, gamma, delta, epsilon, eta, and zeta. AB - To determine whether individual protein kinase C (PKC) isozymes differentially phosphorylate sites in the linker region of human P-glycoprotein (P-gp), we used a synthetic peptide substrate, PG-2, exactly corresponding to amino acid residues spanning the region 656-689 of the multidrug resistance gene (MDRI). All tested PKC isozymes phosphorylated PG-2. The maximum phosphate incorporation by calcium dependent PKC isozymes alpha, betaI, betaII, and gamma was 3, 2, 2, and 3 mol phosphate/mol PG-2, respectively. The maximum phosphate incorporation by calcium independent isozymes delta, epsilon, eta, and zeta was 1.5, 0.5, 1.5, and 1.5 mol phosphate/mol PG-2, respectively. Two-dimensional tryptic phosphopeptide mapping indicated differential phosphorylation of the PKC consensus sites Ser-661, Ser 667, and Ser-671 by individual isozymes, which may be functionally significant. These data suggest that differential phosphorylation by PKC isoenzymes of PKC sites within the P-gp linker region may play a role in modulating P-gp activity. PMID- 10535750 TI - Potentiation by febrifugine of host defense in mice against Plasmodium berghei NK65. AB - The effect of febrifugine, the main alkaloidal constituent of an antimalarial crude drug, Dichroa febrifuga Lour., on protective immunity in mice infected with erythrocytic stage Plasmodium berghei NK65 was investigated. Febrifugine was administered orally, at a dose of 1 mg/kg/day, to mice before and/or after they were infected intraperitoneally with 2 x 10(6) parasitized red blood cells. Then, mortality and the levels of parasitemia and plasma NO3- [a degradation product of nitric oxide (NO)] were monitored. Febrifugine significantly reduced the mortality and the level of parasitemia. The plasma NO3- concentration began to rise within 2 days after treatment with febrifugine and declined to normal in 2 days when the mice were treated orally with febrifugine once a day for 3 consecutive days before parasite infection. This antimalarial activity of febrifugine was reduced by both N(G)-monomethyl-L-arginine and aminoguanidine. These results indicate that the increased production of NO by febrifugine plays an important role in host defense against malaria infection in mice. PMID- 10535751 TI - Cross-resistance of dideoxycytidine-resistant cell lines to azidothymidine. AB - 2',3'-Dideoxycytidine (ddC) and azidothymidine (AZT) inhibit HIV-1 replication and currently are used in AIDS therapy. Long-term use of the drugs is associated with the selection of drug-resistant HIV strains, thus limiting their effectiveness. Another mechanism, associated with their altered metabolism in host cells, also can cause "cellular" drug resistance. Human lymphocytic H9 cell lines (H9-ddC0.5w and H9-ddC5.0w) selected for ddC resistance by exposure to 0.5 and 5.0 microM ddC were found to be cross-resistant to AZT. Compared with controls, the thymidine kinase (TK) activities in H9-ddC0.5w and H9-ddC5.0w cells were 56.7 and 51.4% (with thymidine as a substrate) and 50.3 and 42% (with AZT as a substrate). Consequently the cellular incorporation of AZT and thymidine (24-hr incubation) also was reduced to 51.3 and 70.0% in H9-ddC0.5w cells and to 12.1 and 17.3% in H9-ddC5.0w cells. A 3-hr incubation with 25 microM AZT and ddC decreased their cellular incorporation to 50.5 and 76.15% in H9-ddC0.5w cells and to 12.95 and 47.8% in H9-ddC5.0w cells compared with H9 cells. Thus, the change in AZT accumulation did not correlate exactly with the decrease in TK activity and far exceeded the effect on ddC accumulation. Evidence is presented that ddC, in addition to deoxycytidine kinase, affected TK1 activity. The involvement of multidrug resistance proteins in the mechanism of the resistance was ruled out by the failure of trifluoperazine and verapamil to alter cellular accumulations of AZT, ddC, daunorubicin, and rhodamine-123. Development of cellular ddC and AZT cross-resistance may affect the therapeutic efficacy of these antiviral agents. PMID- 10535752 TI - Relative contributions of mouse liver subcellular fractions to the bioactivation of mitomycin C at various pH levels. AB - Mitomycin C (MMC) is a clinically active anticancer drug that requires reductive activation to exert its toxicity. The enzymes currently recognized as capable of activating MMC cannot account for all of the toxicity of the drug. These studies were conducted to identify and compare the subcellular compartments where MMC reduction can take place under different physiological conditions. Subcellular fractionation of mouse liver was achieved using differential centrifugation and isopycnic equilibrium gradient centrifugation. Nuclear, mitochondrial, microsomal, lysosomal, peroxisomal, and cytosolic fractions were assayed for their ability to reductively activate MMC at pH 6.0 and 7.4. MMC reductive activation was determined by its ability to generate reactive oxygen species. The results of these studies showed that MMC reductive activation by the various fractions was pH dependent. At pH 7.4, the microsomal fraction accounted for approximately 78% of the total MMC reductive activation. The peroxisomal fraction accounted for 12% and the nuclear and lysosomal fractions each accounted for 5% of the total reductive activation. At pH 6.0, the microsomes accounted for 51% and the peroxisomes for 34% of the total reductive activation. The mitochondrial fraction, which did not reductively activate MMC at pH 7.4, accounted for 9% of the total activation at pH 6.0. These results suggested that peroxisomes may be important in MMC activation at either pH and that at pH 6.0 the mitochondrial fraction may also be important for MMC reductive activation. PMID- 10535754 TI - Increased DNA-binding activity of cis-1,1 cyclobutanedicarboxylatodiammineplatinum(II) (carboplatin) in the presence of nucleophiles and human breast cancer MCF-7 cell cytoplasmic extracts: activation theory revisited. AB - The molecular mechanism of carboplatin [cis-1,1 cyclobutanedicarboxylatodiammineplatinum(II)] activation is still unresolved. We studied the binding of carboplatin to calf thymus DNA in the presence of thiourea, glutathione, and human breast cancer MCF-7 cell cytoplasmic extracts by measurement of DNA-dependent ethidium bromide fluorescence and atomic absorption spectroscopy. After a 96-hr period of reaction, the decrease in the DNA-dependent fluorescence yield of ethidium bromide due to the formation of platinum (Pt)-DNA adducts increased significantly in the presence of thiourea (6-fold) and glutathione (3- to 4-fold) as compared to the controls in the absence of the nucleophiles. There was also a marked elevation in the levels of platinum incorporated into DNA, measured by atomic absorption spectroscopy (2- to 3-fold and 5- to 7-fold for thiourea and glutathione, respectively). More remarkably, the Pt-DNA adducts formed in the presence of cytoplasmic extracts of MCF-7 human breast cancer cells also showed similar results in a dose-related fashion. Carboplatin, therefore, displayed a characteristic increase in DNA binding/damaging in the presence of the very same S-containing nucleophiles that showed the expected quenching effects in the case of cisplatin [cis diamminedichloroplatinum (II)]. We propose a nucleophile-facilitated release of the active species of carboplatin prior to binding with DNA. PMID- 10535753 TI - Discrimination among reduced folates and methotrexate as transport substrates by a phenylalanine substitution for serine within the predicted eighth transmembrane domain of the reduced folate carrier. AB - A phenylalanine substitution for serine in the reduced folate carrier at residue 309 (RFC1-S309F) was identified in a methotrexate (MTX)-resistant cell line selected with 5-formyltetrahydrofolate (5-CHO-THF) as the sole folate source. The transport characteristics of the mutated carrier were studied by transfection into the MTXrA line, which lacks endogenous RFC1 function. The level of expression of carrier in the cell lines studied was determined by specific surface binding of 5-methyltetrahydrofolate (5-CH3-THF). Influx of 5-CH3-THF and 5-CHO-THF mediated by RFC1-S309F was 20- and 7-fold greater than that of MTX, respectively. Consistent with the influx difference between 5-CHO-THF and MTX, the growth requirement (EC50) for 5-CHO-THF in MTXrA-S309F cells was decreased by a factor of 9, while the MTX IC50 was reduced by a factor of only approximately 2 as compared with the recipient MTXrA cells. The decrease in 5-CH3-THF influx mediated by the mutated carrier was attributed to a decrease in the mobility of the 5-CH3-THF-carrier complex, since the influx Kt was essentially unchanged. However, the reduction in 5-CHO-THF and MTX influx was attributed to decreases in both carrier affinity and Vmax, although the decline in the MTX influx Vmax appeared to be much greater than for 5-CHO-THF. The inhibitory effect of chloride on 5-CHO-THF influx observed for L1210 cells was eliminated in the MTXrA-S309F line. This study represents another example of a single mutation in RFC1 that markedly impairs MTX influx but partially preserves transport of reduced folates when cells are selected with 5-CHO-THF as the available folate substrate. The data indicate that residues in the predicted eighth transmembrane domain of RFC1 can play an important role in the selectivity of folate binding and the mobility of the carrier-substrate complex. PMID- 10535755 TI - Nuclear vitamin K2 binding protein in human osteoblasts: homologue to glyceraldehyde-3-phosphate dehydrogenase. AB - The importance of vitamin K in bone metabolism has been suggested previously. The binding protein of vitamin K2 (menatetrenone, 2-methyl-3-all-trans-tetraphenyl 1,4-naphthoquinone, menaquinone-4), found in nuclear extract of human osteoblasts, binds to vitamin K1 and K2, but not K3. Since the binding protein does not bind to other steroids or vitamins, such as hydrocortisone, vitamin A, 1,25(OH)2vitamin D3, trolox (a derivative of vitamin E), and warfarin, a specific binding protein to vitamin K1 and vitamin K2 in osteoblasts was suggested. The size of the specific binding protein was revealed to be 6S by sucrose density gradient and about 40,000 daltons by SDS-PAGE. Twenty amino acid residues from the N-terminal were the same as human glyceraldehyde-3-phosphate dehydrogenase (GAPDH), but the 21st residue, alanine, was replaced with serine. The binding protein was precipitated with anti-human GAPDH antibody, and authentic human GAPDH could bind vitamin K2. We propose that the nuclear binding protein for vitamin K2 exists in nuclei similarly to other vitamin receptors and that the molecular structure is very close to human GAPDH. PMID- 10535756 TI - Inhibition of 12-O-tetradecanoylphorbol-13-acetate induction of c-fos mRNA by the protein kinase A inhibitor N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinoline sulfonamide. AB - The tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) can induce expression of many immediate-early genes, such as c-fos and c-jun. In this study, TPA increased c-fos mRNA, cellular cyclic AMP, and protein kinase A (PKA) activity in the first 30 min with similar inductive time courses. Treatment of NIH 3T3 cells with N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinoline sulfonamide (H-89), a PKA specific inhibitor, suppressed TPA induction of PKA activity and c fos mRNA in a concentration-dependent manner, but did not inhibit serum-induced transcription. H-89 did not inhibit TPA and serum induction of c-jun mRNA. H-89 interfered with TPA-stimulated serum-responsive element-binding activity in a concentration-dependent manner, but did not inhibit TPA-induced mitogen-activated protein kinase 1/2 activity or Elk-1 phosphorylation. TPA stimulation of a c-fos promoter reporter construct was inhibited by overexpression of the dominant negative regulatory protein of PKA. In deletion studies, the H-89 inhibitory element was found to be localized between -563 and -379 in the c-fos promoter region. These results suggest that H-89 will be very useful for investigating the molecular mechanism of TPA induction of c-fos. PMID- 10535757 TI - Antioxidant response element-mediated 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induction of human NAD(P)H:quinone oxidoreductase 1 gene expression. AB - Antioxidant response element (ARE) is required for high basal expression of the human NAD(P)H:quinone oxidoreductase 1 (NQO1) gene in tumor cells and its induction in response to beta-naphthoflavone and phenolic antioxidants. In this study, we have demonstrated that ARE also is required for induction of human NQO1 gene expression in response to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). The various results suggest an alternate pathway for TCDD induction of human NQO1 gene expression. This pathway is independent of xenobiotic response element (XRE) and aromatic hydrocarbon (Ah) receptor. It is presumed that TCDD-induced expression of CYP1A1 leads to increased oxidative stress, resulting in transcriptional activation and/or modification of ARE-binding factors and increased expression of the human NQO1 gene. PMID- 10535758 TI - Antiplatelet effect of amlodipine: a possible mechanism through a nitric oxide mediated process. AB - The effect of amlodipine, a novel calcium channel blocker of the dihydropyridine type, on rabbit platelet aggregation, and the possible antiaggregatory mechanisms of amlodipine, especially on the nitric oxide (NO) guanosine 3',5'-cyclic monophosphate (cyclic GMP)-mediated pathway, were investigated. Other effects of amlodipine on thromboxane B2 (TXB2) formation in platelets also were examined. Amlodipine concentration-dependently inhibited rabbit platelet aggregation induced by collagen (10 microg/mL) or thrombin (0.1 U/mL) with an IC50 range of 32-69 microM. Along with this inhibition, our results also demonstrated that in the presence of L-arginine (100 IM), amlodipine (50 microM) increased nitric oxide synthetase (NOS) activity (from the resting activity of 2.05+/-0.36 to 7.11+/-0.95 pmol/mg protein/min) and NO release (by 80%), accompanied by an elevation of the cyclic GMP level (from the resting platelet level of 1.27+/-0.12 to 6.21+/-0.55 pmol/10(9) platelets) induced by collagen (10 microg/mL). However, the antiaggregatory effect of amlodipine (50 microM) could be attenuated significantly by oxyhemoglobin (5 microM), a NO scavenger, or N(G)-nitro-L arginine methyl ester (100 microM), a specific NOS inhibitor. In addition, the TXB2 production in platelets induced by collagen or thrombin was concentration dependently inhibited by amlodipine. Therefore, we propose that the antiaggregatory mechanisms of amlodipine might be mediated, in part, by a NO cyclic GMP process accompanied by the inhibition of TXB2 formation in platelets. PMID- 10535759 TI - Inhibitory effect of Ginkgo biloba extract on the expression of inducible nitric oxide synthase in endothelial cells. AB - Excessive production of nitric oxide (NO) may have cytotoxic effects through the formation of peroxynitrite with superoxide. The extract of Ginkgo biloba leaves (EGb) has been demonstrated to be a potent scavenger of free radicals. Although EGb has been shown recently to inhibit NO production in macrophages, its effect on NO production in endothelial cells is largely unknown. The objective of this study was to elucidate the mechanism by which EGb affects NO production in a human endothelial cell line (ECV304). After cells were incubated with EGb (10-100 microg/mL) for 2 or 4 hr, the amounts of NO metabolites released by the cells were quantitated, and cellular NOS activities were determined following the conversion of [3H]arginine to [3H]citrulline. NOS protein expression was determined by western immunoblotting analysis. mRNA levels were examined by reverse transcription-polymerase chain reaction (RT-PCR) analysis. EGb (50 microg/mL) caused a 30% reduction of NO metabolites released by endothelial cells. Following EGb treatment, cellular inducible NO synthase (iNOS) activity was reduced by 28% with a concomitant reduction in the levels of iNOS protein mass and mRNA. There was no change in the activity or protein mass of constitutive NO synthase in these cells. EGb inhibited NO production by attenuating the level of iNOS mRNA in ECV304 cells. Selective inhibition of iNOS by EGb may be therapeutically relevant in modulating NO production in endothelial cells. PMID- 10535760 TI - Contribution of phosphodiesterase isoenzymes and cyclic nucleotide efflux to the regulation of cyclic GMP levels in aortic smooth muscle cells. AB - Involvement of phosphodiesterase isoenzymes (PDEs) in guanosine-3',5'-cyclic monophosphate (cGMP) hydrolysis was analyzed in aortic smooth muscle cells. Four families of PDEs were separated from pig aorta: PDE1 (calcium-calmodulin activated), PDE3 (cGMP-inhibited), PDE4 (adenosine 3',5'-cyclic monophosphate [cAMP]-specific), and PDE5 (cGMP-specific). Within this PDE complement, PDE1 and PDE5 mostly contributed to the hydrolysis of cGMP both in the presence and absence of calcium-calmodulin. The role of these isoenzymes in cGMP degradation was analyzed in primary cultures of porcine aortic smooth muscle cells after stimulation with sodium nitroprusside (SNP) or atrial natriuretic factor (ANF). Pretreatment with 10 microM zaprinast, a concentration that selectively inhibits PDE5, did not potentiate the SNP- or ANF-induced rise of cGMP, questioning the widespread opinion that only PDE5 accounts for cGMP hydrolysis in this tissue. Further evidence came from experiments assessing the effect of zaprinast or 3 isobutyl-1-methylxanthine at concentrations inhibiting both type 1 and type 5 isoenzymes, in which this potentiation was clearly seen. Contribution of cGMP egression to the control of intracellular cGMP levels after SNP or ANF stimulation was also investigated. Shortly after guanylate cyclase activation, extracellular cGMP levels surpassed intracellular levels. However, comparison of the amounts of cGMP extruded to the extracellular medium with those degraded by PDEs leads to the conclusion that efflux is of relatively minor importance in regulating intracellular cGMP levels. In cells made tolerant to SNP, selective PDE5 inhibition synergistically increased intra- and extracellular cGMP amounts after SNP stimulation. These results indicate a previously undescribed greater relevance of PDE5 after tolerance development in aortic smooth muscle cells. PMID- 10535761 TI - A method of achieving physiological plasma levels of melatonin in the chicken by oral administration. AB - In avian species it has been difficult to elucidate the precise role of melatonin in the control of reproductive cycles. We have investigated ways of administering melatonin to immature chickens and laying hens to achieve physiological levels and patterns in blood simulating either short or long photoperiods. Melatonin was administered orally using different doses and various ways of applying melatonin to the feed. For subcutaneous injections, melatonin was suspended in propylene glycol or grape seed oil. Melatonin always appeared in the first blood samples taken within an hour of administration. When melatonin was absorbed into feed pellets or whole wheat, a high initial plasma concentration was reached, followed by a rapid decrease over the ensuing 2-3 hr, but was still detectable as long as 24 hr after administration. For example, doses of 300 microg/kg produced 15 nM, which is more than ten times higher than the nocturnal peak concentration. When melatonin was absorbed into cracked wheat grains that were subsequently washed with ethanol, the initial transitory peak was eliminated, levels in plasma were sustained for at least 12 hr in the normal nocturnal range (750 pM), and no melatonin (< 60 pM) was present 18 hr later. When injected (2 microg/bird), concentrations peaked (610 pM) within 30 min and decreased rapidly over the next 2-3 hr. It was concluded that melatonin-treated, ethanol-rinsed cracked wheat grains can be used to experimentally mimic long-night plasma melatonin patterns. Injections may be useful for mimicking the melatonin patterns of very short nights in chickens experiencing constant light. PMID- 10535762 TI - Estimation of frequently sampled nocturnal melatonin production in humans by deconvolution analysis: evidence for episodic or ultradian secretion. AB - In order to investigate nocturnal melatonin production and clearance rates, we applied deconvolution analysis to plasma melatonin concentration time series obtained every 20 min for 12 hr from 20:00 h to 08:00 h in two groups of healthy subjects at rest (group 1, 12 male subjects, 22-26 yr; group 2, ten female subjects, 31-42 yr). The estimated melatonin production rate from group 1 (0.55 +/- 0.21 microg/kg/night) was significantly higher than that of group 2 (0.26 +/- 0.19 microg/kg/night), as well as the mass of melatonin released per burst (275 +/- 110 vs. 145 +/- 130 pg/mL for groups 1 and 2, respectively), the amplitude of secretory bursts (7.8 +/- 3.2 vs. 4.7 +/- 3.5 pg/mL/min), and the pulsatile melatonin production rate (2.76 +/- 1.14 vs. 1.27 +/- 0.97 ng/mL/night). These differences could reflect alterations related to age and or gender. No differences were observed between the two groups in the secretory burst half duration and frequency and the interburst interval. Melatonin production rates, as estimated by deconvolution analysis, are in agreement with other independent estimates, especially the isotopic method, and disclose an ultradian rhythmicity. PMID- 10535763 TI - Hematopoietic effect of melatonin involvement of type 1 kappa-opioid receptor on bone marrow macrophages and interleukin-1. AB - Melatonin has immuno-enhancing properties and exerts colony-stimulating activity (CSA) via T-helper cell-derived opioids. Opioid agonists may mimic the CSA of melatonin with an order of potency that suggests the presence of a type 1 kappa opioid receptor (type 1 kappaOR [kappa]-OR]). The kappaOR antagonist nor binaltorphimine neutralized the in vitro effect of melatonin and inhibited regeneration of hematopoiesis in mice treated with carboplatin. The CSA of dynorphin A was abolished by incubation of adherent cells with antisense (AS) oligodeoxynucleotide to kappaOR or by addition of anti-interleukin (IL)-1 monoclonal antibody (mAb), which also neutralized the effect of melatonin. Bone marrow cells that express kappaORs were identified to be macrophages. In conclusion, we describe the presence of kappaORs in bone marrow macrophages and suggest a hematopoietic function for melatonin via endogenous kappa-opioid agonists and, possibly, IL-1. PMID- 10535764 TI - Antioxidative effects of melatonin in Drosophila melanogaster: antagonization of damage induced by the inhibition of catalase. AB - In Drosophila melanogaster strain Canton S, the catalase inhibitor 3-amino-1,2,4 triazole was used to enhance oxidative stress from endogenous sources. This treatment was chosen as an alternative to direct administration of oxidants, which would cause damage and interact with melatonin already in the extracellular space before they reach the intracellular compartments. Male flies were kept in constant darkness and fed 1% sucrose as the only diet, with or without additions of melatonin (2 mM), inhibitor (100 mM), or combinations of both. After 20 or 24 hr, most of the animals exposed to 3-amino-1,2,4-triazole only had died, whereas a large number of flies had survived the inhibitor treatment in the presence of melatonin. Protein carbonyl, an indicator of oxidative protein modification, and lipid peroxidation, as determined by the formation of malondialdehyde and 4 hydroxyalkenal, were measured in flies treated for 20 hr. Melatonin alone did not substantially change these parameters, but prevented the increase in protein carbonyl caused by the catalase inhibitor. The effect of 3-amino-1,2,4-triazole on lipid peroxidation was relatively minor, but a clear-cut inhibition was found after simultaneous administration of melatonin. The preferential suppression of oxidative damage of proteins, as compared to lipids, indicates a particular protective role for melatonin in the aqueous phase of cellular compartments. PMID- 10535765 TI - Melatonin and its precursors scavenge nitric oxide. AB - Nitric oxide (NO) scavenging activity of melatonin, N-acetyl-5-hydroxytryptamine, serotonin, 5-hydroxytryptophan and L-tryptophan was examined by the Griess reaction using flow injection analysis. 1-Hydroxy-2-oxo-3-(N-methyl-3 aminopropyl)-3-methyl-1-triazene (NOC-7) was used as NO generator. The Griess reagent stoichiometrically reacts with NO2-, which was converted by a cadmium copper reduction column from the stable end products of NO oxidation. Except for tryptophan, all the compounds examined scavenged NO in a dose-dependent manner. Melatonin, which has a methoxy group in the 5-position and an acetyl side chain, exhibited the most potent scavenging activity among the compounds tested. Serotonin, N-acetyl-5-hydroxytryptamine, and 5-hydroxytryptophan, respectively, showed moderate scavenging activity compared to melatonin. Tryptophan, which has neither a methoxy nor a hydroxyl group in the 5-position, exhibited the least NO scavenging activity. PMID- 10535766 TI - Melatonin content in plasma and large intestine of patients with colorectal carcinoma before and after surgery. AB - The distinct melatonin rhythm with higher concentrations during the darktime was found in plasma of both control patients and patients with colorectal carcinoma. Moderate surgery did not induce any changes in plasma melatonin levels, but a pronounced increase in both the day- and nighttime melatonin concentrations was found after surgical treatment for colon cancer. The melatonin content in the tumor tissue did not differ from that in the proximal and the distal parts of the resected gut, which were without signs of malignant changes. Neither concentrations of serotonin nor 5-hydroxyindole acetic acid differed among analyzed parts of the gut. Daytime melatonin concentrations in gut tissue (314.7 +/- 87.8 pg/g of wet tissue) were more than ten times higher than the daytime levels in circulation. It was hypothesized that increased levels of this hormone in the gastrointestinal tract may play an important protective role against the development of colorectal cancer via stimulation of the immune system, protection against free radicals, and interaction with fatty acid uptake and metabolism. PMID- 10535767 TI - CREB phosphorylation and melatonin biosynthesis in the rat pineal gland: involvement of cyclic AMP dependent protein kinase type II. AB - Phosphorylation of cyclic AMP response element binding protein (CREB) at amino acid serine 133 appears as an important link between the norepinephrine (NE) induced activation of second messenger systems and the stimulation of melatonin biosynthesis. Here we investigated in the rat pineal gland: 1) the type of protein kinase that mediates CREB phosphorylation: and 2) its impact on melatonin biosynthesis. Immunochemical or immunocytochemical demonstration of serine133 phosphorylated cyclic AMP regulated element binding protein (pCREB) and radioimmunological detection of melatonin revealed that only cyclic AMP-dependent protein kinase (PKA) inhibitors suppressed NE-induced CREB phosphorylation and stimulation of melatonin biosynthesis, whereas inhibitors of cyclic GMP-dependent protein kinase (PKG), mitogen-activated protein kinase kinase, protein kinase C, or calcium-calmodulin-dependent protein kinase (CaMK) were ineffective. Investigations with cyclic AMP-agonist pairs that selectively activate either PKA type I or II link NE-induced CREB phosphorylation and stimulation of melatonin biosynthesis to the activation of PKA type II. Our data suggest that PKA type II plays an important role in the transcriptional control of melatonin biosynthesis in the rat pineal organ. PMID- 10535768 TI - Melatonin-induced inhibition of proliferation and G1/S cell cycle transition delay of human choriocarcinoma JAr cells: possible involvement of MT2 (MEL1B) receptor. AB - Melatonin, the pineal neurohormone, is an evolutionarily conserved photoperiodic signaling molecule with diverse functions that include the entrainment of human circadian rhythms. Although evidence supporting a direct inhibitory action of melatonin on human cancer cell proliferation exists in the literature, the molecular and cellular signaling mechanisms involved are largely undefined. In our study, significant inhibition of human choriocarcinoma JAr cell proliferation at physiological and pharmacological concentrations of melatonin was observed. 2 Iodomelatonin, a high affinity melatonin receptor agonist, was more potent than melatonin in inhibiting JAr cell proliferation. In addition, the presence of putative melatonin receptors in choriocarcinoma was suggested by the demonstration of specific 2-[125I]iodomelatonin binding to the tumor. Interestingly, the selective MT2 melatonin receptor ligand, 4-phenyl-2 propionamidotetraline (4-P-PDOT), was found to exert not only concentration dependent anti-proliferative actions on JAr cells, but also additive effects with melatonin in inhibiting JAr cell proliferation. Furthermore, MT2 melatonin receptor gene expression by JAr cells was demonstrated by reverse transcription polymerase chain reaction (RT-PCR) and in situ hybridization (ISH). Taken together, our data suggest that the reported anti-proliferative action of melatonin on human choriocarcinoma JAr cells may be mediated, in part, by MT2 melatonin receptor. Moreover, analysis of melatonin effect on cell cycle kinetics indicated that G1/S transition delay may underlie the observed inhibition of choriocarcinoma cell proliferation by melatonin. PMID- 10535769 TI - Genetic typing of bovine pestiviruses from England and Wales. AB - Using RNA purified directly from stored clinical specimens, a collection of 62 pestiviruses were typed by RT-PCR and sequencing within the 5'-untranslated region of the genome. All the specimens had been obtained in 1966/1967 from diary cattle in England and Wales. Eight further pestiviruses, grown in cell culture, were characterised in the same way. Seven of these viruses were representatives of a panel of British isolates, obtained from cattle ten years before. The eighth was the virus used in a British bovine viral diarrhoea (BVD) vaccine. Most of the viruses were genetically unique and were of BVDV type Ia. One recent isolate was BVDV type Ib, two others were intermediate between Ia and Ib. No BVDV type II or border disease virus (BDV) isolates were found. There was no overall association between geographical and phylogenetic clustering, suggesting long-distance virus dispersal, presumably via trading of infected cattle. The sequences of the recently obtained cattle viruses were very similar or, in one case, identical to the older isolates in the region studied. Their close similarity to some previously characterised pestiviruses from British sheep suggests that a common pool of BVDV Ia is shared by these two livestock species, although another pestivirus--BVDV--is confined to sheep. The British cattle viruses were mostly distinct from continental European isolates, but more similar to type Ia isolates from North American cattle. PMID- 10535771 TI - Prevalence and infection risks of zoonotic enteropathogenic bacteria in Swiss cow calf farms. AB - A longitudinal study was performed in 67 larger Swiss cow-calf farms from September 1996 through November 1997. The objectives of the study were to estimate prevalence and risk factors for colonization with potentially zoonotic enteropathogenic bacteria in younger calves and in calves at weaning age. The study included data from 395 calves with three to four fecal samples each. Fecal samples were analyzed for Campylobacter spp., verotoxin producing E. coli (VTEC), Yersinia spp. and Salmonella sp. Possible environmental and individual factors associated with colonization of these agents were examined. The calves were housed indoor during the first 3 months of life (winter 1996/1997). The prevalences within this time period were: C. coli 3.4%, C. fetus 15.5%, C. hyointestinalis 9.6%, C. jejuni 38.5%, VTEC 44.3% and Yersinia spp. 2%. At the end of the grazing season the prevalences at weaning (8-10 months of age) were: C. coli 1.7%, C. fetus 4.0%, C. hyointestinalis 25.9%, C. jejuni 13.3%, VTEC 38.2% and Yersinia spp. 0%. No salmonellae were present at any time of the study. The prevalences of C. jejuni and VTEC increased significantly within the first 3 months of life, whereas C. hyointestinalis decreased. None of the environmental factors such as housing or feeding had any consistent influences on colonization by the bacteria studied. VTEC, Campylobacter spp. and Yersinia spp. should probably be considered as normal inhabitants of the bovine intestinal tract. However, as they represent a source of gastrointestinal infections in humans, management factors limiting intestinal colonization of these bacteria should be considered in cow-calf operations. PMID- 10535770 TI - A highly specific and sensitive competitive enzyme-linked immunosorbent assay (ELISA) based on baculovirus expressed pseudorabies virus glycoprotein gE and gI complex. AB - A direct competition enzyme-linked immunosorbent assay (ELISA) based on baculovirus expressed complex of pseudorabies virus (PRV) glycoproteins E (gE) and I (gI) has been developed. For that purpose gE and gI genes of PRV were co expressed in insect cells. Complex formation was confirmed by radioimmunoprecipitation assay. The specificity and sensitivity of the test were evaluated and compared with an ELISA using only gE as an antigen and a commercially available test. For validation, 245 negative sera and 165 positive sera have been tested. The gE/gI ELISA had a higher sensitivity and specificity when compared with the ELISA using only gE as the antigen. Both sensitivity and specificity were comparable with the commercially available test. Moreover, the test based on the baculovirus gE/gI complex allows the detection of anti-gE antibodies in pig serum as early as two weeks after infection. The gE/gI ELISA test is easy to perform; its additional advantage is that the gE/gI antigen can be produced in baculovirus system in large quantities without handling live pseudorabies virus. PMID- 10535772 TI - Colonisation of the chicken caecum by afimbriate and aflagellate derivatives of Salmonella enterica serotype Enteritidis. AB - A semi-quantitative cloacal-swab method was used as an indirect measure of caecal colonisation of one-day old and five-day old chicks after oral dosing with wild type Salmonella enterica serovar Enteritidis PT4 and genetically defined isogenic derivatives lacking the ability to elaborate flagella or fimbriae. Birds of both ages were readily and persistently colonised by all strains although there was a decline in shedding by the older birds after about 21 days. There were no significant differences in shedding of wild-type or mutants in single-dose experiments. In competition experiments, in which five-day old birds were dosed orally with wild-type and mutants together, shedding of non-motile derivatives was significantly lower than wild-type. At 35 days post infection, birds were sacrificed and direct counts of mutants and wild-type from each caecum were determined. Whilst there appeared to be poor correlation between direct counts and the indirect swab method, the overall trends shown by these methods of assessment indicated that flagella and not fimbriae were important in caecal colonisation in these models. PMID- 10535773 TI - Optimized ribotyping protocol applied to Hungarian Bordetella bronchiseptica isolates: identification of two novel ribotypes. AB - We reported previously that ribotype patterns generated with PvuII and a probe derived from the Escherichia coli rrnB gene could be used to differentiate isolates of Bordetella bronchiseptica. In the present study we report modifications made to the original ribotyping procedure that permit detection in the formerly characterized isolates of an additional 8 fragments with homology to rrnB. Ribotypes were redefined to include these fragments. Although this modification did not permit the detection of novel ribotypes from the previously characterized isolates, it did result in a more accurate reclassification of five of these isolates to other existing ribotypes. It was hypothesized that the additional fragments could form the basis for novel ribotypes in future analyses, and this was supported by the subsequent evaluation of 101 previously uncharacterized pig, rabbit, and dog B. bronchiseptica isolates from Hungary. A total of six different patterns were detected from this group, including two previously not identified that were designated ribotypes 17 and 18. The profile of ribotype 17 includes a novel fragment not associated with any other ribotype. A subset of the fragments constituting ribotype 18, essential for its differentiation from other ribotypes, is only detectable under the modified conditions reported here. Hungarian swine isolates are highly clonal, since 98.2% were identified as ribotype 3. Similarly, 83.7% of rabbit isolates from Hungary are also ribotype 3. Cluster analysis revealed that despite the existence of numerous ribotypes, B. bronchiseptica isolates display limited heterogeneity. The ability to detect additional ribotypes under the modified conditions described in this study strengthens the usefulness of ribotyping as an epidemiologic tool. PMID- 10535774 TI - Induction, characterisation and pathogenicity in rainbow trout Oncorhynchus mykiss (Walbaum) of Lactococcus garvieae L-forms. AB - Difficulties with induction and cultivation of L-forms, particularly those derived from Gram positive parent cells, have constrained to some degree the ability to evaluate the pathogenicity of these morphotypes. Induction of L-forms of Lactococcus garvieae was undertaken using either charcoal or inactivated horse serum media supplemented with ampicillin, benzylpenicillin or erythromycin, the drug of choice for treatment of infections in rainbow trout, Oncorhynchus mykiss, (Walbaum), and NaCl as an osmotic stabiliser. Lysozyme treated cells could be cultured in a cell wall deficient state using media consisting of charcoal, NaCl and either ampicillin or benzylpenicillin. The influence of some amino acids for induction of L-forms was assessed by disc diffusion and combined interaction. Analysis of variance of colony counts indicated that the amino acids glycine, DL methionine, L-threonine and L-serine (P<0.03), and the presence of charcoal were beneficial and that inactivated horse serum was detrimental to L-form development. Electron microscopy revealed that the cell wall of L-forms was missing and this cell had a greatly expanded volume compared to parent cells. Electrophoresis of whole cell proteins showed some variation of electropherotype between parent and L-form cells. L-forms expressed greater quantities of proteins with molecular mass of 36 and 66 kDa and parent cells contained greater quantities of proteins of molecular mass 29, 43 and 60 kDa. Additional proteins of molecular mass 32, 44 and 53 kDa were present in L-form extracts, and in parent cells of 34, 38, 40, 42, 85 and 123 kDa which may represent cell wall associated proteins or alterations in expression due to different growth rates. Intraperitoneal challenge of rainbow trout with L-forms failed to produce overt infection even in immune-suppressed fish, but L-forms were shown by indirect fluorescent antibody test to remain inkidney tissue. Fish were susceptible to infection when challenged with parent cells of L. garvieae. PMID- 10535775 TI - Air pollution kills babies... PMID- 10535777 TI - A proposal to calculate publication equivalents. PMID- 10535776 TI - The observational epidemiology of changing weight: an appeal for reasons. PMID- 10535778 TI - The effect of air pollution on infant mortality appears specific for respiratory causes in the postneonatal period. AB - To examine the association between individual lifetime measures of mean exposure to air pollution and postneonatal respiratory deaths, we have conducted a matched population-based case-control study covering all births registered in the Czech Republic from 1989 to 1991 that were linked to death records. For each case of infant death, we have randomly selected 20 controls from infants of the same sex born on the same day and alive when the case died. Exposure was assigned as the arithmetic mean of all 24-hour air pollution measurements in the district of residence of each case and control for the period between the birth and death of the index case. We used conditional logistic regression to estimate the effects of suspended particles, sulfur dioxide, and nitrogen oxides on risk of death in the neonatal and postneonatal period, controlling for maternal socioeconomic status and birth weight, birth length, and gestational age. There were 2,494 infant deaths with exposure data on at least one pollutant, 133 of them from respiratory causes. The effects of all pollutants were strongest in the postneonatal period and were specific for respiratory causes. For these, rate ratios for a 50 microg/m3 increase in particles, sulfur dioxide, and nitrogen oxides were 1.95 [95% confidence interval (CI) = 1.09-3.50], 1.74 (95% CI = 1.01 2.98), and 1.66 (95% CI = 0.98-2.81), respectively, after controlling for all covariates. Only particles showed a consistent association when all pollutants were entered in one model. We found no evidence of a relation between any pollutant and mortality from other causes. These results indicate that the effects of air pollution on infant mortality are specific for respiratory causes in the postneonatal period, are independent of socioeconomic factors, and are not mediated by birth weight or gestational age. PMID- 10535779 TI - Independent effects of stable and changing body weight on total mortality. AB - Most studies find that associations between mortality and either body mass index or weight change are U-shaped. Previous studies, however, have not considered independent effects of weight level and weight change while also controlling for confounding by diseases leading to weight change. We used follow-up data on average 10-year total mortality from a Danish population of 15,113 men and women, who had their weight measured at about 5-year intervals. We obtained information on preexisting disease at surveys and by linkage to hospital discharge registers. We defined subclinical disease as incident disease or death during the first 4 years of follow-up. We decomposed the mortality risk associated with weight change into a static effect, corresponding to the difference in mortality at stable weight at the initial and attained weight, and a dynamic effect, estimated as the difference between mortality after weight change and mortality at stable weight at the attained weight. Both weight level and weight change had independent effects on total mortality, with both these associations being U shaped. Adjustment for smoking and the exclusion of subjects with preexisting and subclinical disease did not alter the associations. PMID- 10535780 TI - A prospective study of vitamin supplement intake and cataract extraction among U.S. women. AB - We prospectively examined the association between vitamin supplement intake and the incidence of cataract extraction during 12 years of follow-up in a cohort of 47,152 female nurses. Women were 45 years or older and free of diagnosed cancer in 1980; others were added as they reached 45 years of age, for a total of 73,956 women. During 720,082 years of follow-up, 1,377 senile cataracts were diagnosed and extracted. Those who used multivitamins or separate supplements of vitamin C, E, or A did not have decreased risks of cataract as compared with nonusers even for use of 10 or more years. After adjusting for cataract risk factors, including cigarette smoking, body mass index, and diabetes mellitus, users of vitamin C supplements for 10 or more years had a relative risk (RR) of 0.95 [95% confidence interval (CI) = 0.76-1.20]. Associations were stronger among long-term vitamin C supplement users who were never-smokers (RR = 0.71; 95% CI = 0.47-1.08) and less than 60 years of age (RR = 0.72; 95% CI = 0.49-1.04). These findings suggest that there is little overall benefit of long-term use of vitamin supplements for risk of cataracts requiring extraction. PMID- 10535781 TI - Work-related assault injuries among nurses. AB - Work-related violence is a major public health problem; however, there is a serious deficiency in the knowledge of risk factors for this problem. The purpose of this case-control study was to identify risk factors for work-related assault injuries among nurses. We used unconditional logistic regression to model the dependence of work-related assault injuries on each exposure of interest and the respective confounders. We found a decreased rate for the presence of security personnel (RR = 0.40; 95% CI = 0.19-0.82). We found increased rates for the following factors: the perception that administrators considered assault to be part of the job (RR = 8.14; 95% CI = 3.76-17.60); having received assault prevention training in the current workplace (RR = 4.64; 95% CI = 2.33-9.23); a high (>5) vs. low (<2) patient/personnel ratio (RR = 2.54; 95% CI = 1.13-5.70); working predominantly with patients with mental illness (RR = 3.5; 95% CI = 1.41 8.85); and working with patients who had more than 1- to 4-week and more than 4 week lengths of stay in the institution vs. <1 day (RR = 8.85; 95% CI = 1.58 49.52 and 4.25; 95% CI = 1.17-15.39, respectively). PMID- 10535782 TI - The impact of parental smoking on asthma and wheezing. SIDRIA Collaborative Group. Studi Italiani sui Disturbi Respiratori nell'Infanzia e l'Ambiente. AB - To evaluate the impact of parental smoking on childhood asthma and wheezing, we studied two random samples of subjects ages 6-7 and 13-14 years in ten areas of northern and central Italy. Standardized questionnaires were completed by parents of 18,737 children and 21,068 adolescents (response rates, 92.8% and 96.3%, respectively) about their smoking habits and the respiratory health of their children. Adolescents were asked about their respiratory health and personal smoking. We compared two groups of cases with healthy subjects: (1) "current asthma" (children, 5.2%; adolescents, 6.2%) and (2) "current wheezing" not labeled as asthma (children = 4.5%, adolescents = 8.5%). Exposure to smoke of at least one parent increased the relative risk of current asthma among children [odds ratio (OR) = 1.34; 95% confidence interval (CI) = 1.11-1.62] and of current wheezing among adolescents (OR = 1.24; 95% CI = 1.07-1.44). Maternal smoking had a stronger effect than paternal smoking. Maternal smoking during pregnancy was associated with current asthma (OR = 1.62; 95% CI = 1.34-1.96) and current wheezing in children (OR = 1.31; 95% CI = 1.06-1.62); the effects were lower among adolescents. Among subjects with a negative history of parental asthma, maternal smoking was associated with current wheezing in both age groups, whereas among those with a positive history of parental asthma it was associated with current asthma in children, but not in adolescents. We estimated that 15% (95% CI = 12-19) of the current asthma cases among children and 11% (95% CI = 8.3-14) of the current wheezing cases among adolescents are attributable to parental smoking in Italy. PMID- 10535783 TI - Family size, infections, and asthma prevalence in New Zealand children. AB - We conducted a prevalence case-control study to investigate the relation between family composition, infection, and development of asthma at age 7-9 years. Potential cases (399) and controls (398) were selected from the Wellington, NZ, arm of the International Study of Asthma and Allergies in Childhood, a population based prevalence study. Further screening questions restricted cases to children with a diagnosis of asthma and current medication use (N = 233) and restricted controls to children without a history of wheezing and no diagnosis of asthma (N = 241). After controlling for confounders (including infections, atopy, and socioeconomic status), family size was strongly related to asthma. Having no siblings [prevalence odds ratio (POR) = 2.51; 95% confidence interval (CI) = 1.05 6.01] or one sibling (POR = 1.86; 95% CI = 1.14-3.03) was associated with an increased risk of asthma compared with having more than one sibling. Parent reported rubeola infection (and possibly other similar viral exanthems) was independently associated with a decreased risk of asthma (POR = 0.48; 95% CI = 0.27-0.83), but reported pertussis infection (POR = 1.57; 95% CI = 0.58-4.24) and day care attendance in the first year of life (POR = 1.81; 95% CI = 0.93-3.51) were not strongly associated with increased risks of asthma. PMID- 10535784 TI - Two centuries of mortality in ten large families with Huntington disease: a rising impact of gene carriership. AB - To estimate the impact of the Huntington gene on mortality, we studied ten families with Huntington disease, whose records started before 1800. We investigated mortality from 1800 to 1997 in 257 carriers of the Huntington gene and 474 potential carriers. Follow-up extended from age 20 years to the date of death or end-of-study date. The observed deaths were compared with those expected on the basis of the general population, adjusted for sex, age, and calendar time. To study the influence of the family and parental transmission, we calculated hazard ratios adjusted for sex, probability of carrying the gene, and year of birth. In 25,013 person-years, 420 deaths occurred, whereas 278 deaths were expected [standardized mortality ratio = 1.5; 95% confidence interval (CI) = 1.4 1.7]. Excess mortality was confined to ages 40-70 years (standardized mortality ratio = 2.2; 95% CI = 1.9-2.4). To study the evolution of mortality over time in this age group, we calculated absolute mortality rates per calendar period. From 1800 onward, mortality rates in the general population continuously declined, but among the families with Huntington disease this decline was absent. There were only small differences in risk between families, and the relative risk for paternal over maternal transmission was 1.2 (95% CI = 0.9-1.5). Our main finding is that persons who carry the Huntington gene and reach middle age have not benefited from advances in medical care and overall increase in life expectancy. PMID- 10535785 TI - Periconceptional nutrient intake and risk for neural tube defect-affected pregnancies. AB - We investigated whether intakes of nutrients, including folate, by women in the periconceptional period were associated with risks of neural tube defect (NTD) affected pregnancies. Data were part of a case-control study of fetuses and infants with NTDs among 1989-1991 California births. We conducted interviews with mothers of 409 NTD cases and 420 nonmal-formed controls. Nutrient intake for the 3 months before conception was derived from food frequency questionnaires and from questions to mothers about vitamin/mineral supplement use. We computed NTD risk for each nutrient controlling for the influence of all other studied nutrients and for maternal education, race/ethnicity, height, and prepregnancy weight. Most single nutrients reduced NTD risks when intakes were considered in quartiles and unadjusted for other nutrients. Some of the same nutrients, however, did not provide similar interpretations when we adjusted for other nutrients. Adjusted analyses revealed decreased NTD risks with increased intakes of methionine, lutein, magnesium, zinc, and thiamin for women who did not use vitamin supplements periconceptionally. We observed decreased NTD risks associated with increased intakes of linoleic acid, cysteine, calcium, and zinc for women who used supplements. We also observed increased NTD risks with increased intakes of oleic acid. For users as well as nonusers of vitamin supplements, we observed reduced risks with increased intakes of grains and dairy products. Chance was a likely alternative explanation for many of the observed risk patterns. PMID- 10535786 TI - A low pregnancy body mass index is a risk factor for an offspring with gastroschisis. AB - A mother's prepregnancy obesity has been suggested as a risk factor for having offspring with an abdominal wall defect. We evaluated this hypothesis among 104 cases of gastroschisis--a severe birth defect of the abdominal wall most prevalent in infants of young women--and 220 controls with no defect. Using Quetelet's index (QI = weight in kg/height in m2) as a measure of body mass, we found a higher risk of gastroschisis (odds ratio (OR) = 3.2; 95% confidence interval (CI) = 1.4-7.3) for underweight mothers (QI<18.1 kg/m2) and a lower risk (OR = 0.2; 0.05-0.9) for overweight mothers (QI>28.3 kg/m2) as compared with mothers of normal weight. As QI was correlated to height, with the correlation varying according to mother's ethnicity and age, we adjusted for these factors in the analysis; the adjusted values approximated the unadjusted values. Evaluation of QI as a continuous variable showed that, for every unit increase in QI, the risk for gastroschisis decreased by about 11%. Sociodemographic, pregnancy, and nutrient factors did not confound the association. These results suggest that low prepregnancy body mass rather than obesity is a risk factor for gastroschisis. PMID- 10535787 TI - Preeclampsia and breast cancer risk. AB - Breast cancer is associated with endogenous hormone levels, but the exact relation and underlying mechanisms remain unclear. Data from several recent epidemiologic studies suggest that a woman who experiences preeclampsia in her own pregnancy, or who was herself born to a preeclamptic pregnancy, is at reduced risk for breast cancer later in life. This paper reviews the evidence for a connection between preeclampsia and breast cancer risk, and discusses the hormonal mechanisms that might explain this association. Preeclampsia is characterized by reduced levels of estrogens and insulin-like growth factor-1, and by elevated levels of progesterone, androgens, human chorionic gonadotropin, IGF-1 binding protein, corticotropin-releasing factor, cortisol, and insulin. These factors may act both individually and synergistically to decrease breast cancer risk. The occurrence of preeclampsia during a woman's pregnancy may reflect an underlying hormonal profile that both predisposes her to preeclampsia and reduces her risk for breast cancer. In addition, the major hormonal alterations associated with preeclampsia during gestation may have lasting effects on subsequent breast cancer risk. Finally, the hormonal and nutritional environment of the womb, for which preeclampsia is a marker, may play an important role in programming lifelong risk for breast cancer in the female offspring. PMID- 10535789 TI - Modeling effects of age at and time since delivery on subsequent risk of cancer. AB - We describe a simple model for examining the temporal effects of childbirth on cancer risk, considering data on uniparous and nulliparous women, from either a cohort or case-control study design. For uniparous women, the expression for risk includes terms for age at delivery and time since delivery. With a suitable definition of the effect of uniparity, no terms relating to delivery are needed for nulliparous women. If the pure age effect is assumed to be the same in all women, the effects of age at delivery and time since delivery are both estimable, despite the linear dependence involving attained age in uniparous women. Omitting terms for time since delivery and considering the heterogeneity of age-specific effects of uniparity provides a valid test for the effect of time since delivery, although risk estimates are biased. Tests based on linear interaction terms for age at delivery and attained age, as applied in recent case-control studies, are not appropriate for investigating the effect of time since delivery. We show how our basic model may be applied to the analysis of case-control data from a Norwegian study of breast cancer. We then compare these results with those from other models. PMID- 10535788 TI - The additional risk of endometrial cancer associated with unopposed estrogen use in women with other risk factors. AB - We analyzed data from a population-based case-control study of endometrial cancer. Our goal was to identify a subgroup of women in whom the additional cancer risk associated with unopposed estrogen use was sufficiently small so as to not be a deterrent to taking a hormone preparation of this type. Researchers interviewed women with endometrial cancer (N = 553) and controls (N = 752) regarding hormone use. The additional risk of endometrial cancer associated with unopposed estrogen use did not vary substantially in the presence or absence of hypertension, parity, oral contraceptive use, or smoking. The results suggest that, although heavier women may experience a greater risk of endometrial cancer associated with unopposed estrogen use (8.2 per 1,000 per year) than lighter women (4.2 per 1,000 per year), long-term users in the latter group nonetheless face a substantial absolute increase in risk. We conclude that subdividing women on the basis of the presence or absence of other known risk factors for endometrial cancer fails to delineate a subgroup that is exempt from the increased risk of this cancer associated with use of unopposed estrogens. 83.6% of estrogen users reported taking conjugated estrogens. PMID- 10535790 TI - Parental age and risk of sporadic and familial cancer in offspring: implications for germ cell mutagenesis. AB - We used the nationwide Swedish Family-Cancer Database to analyze the effect of parental age on cancer in offspring at ages 15-53 years. We studied 13 cancer sites, including 37,877 people. Data on familial and sporadic cancers were analyzed separately. We adjusted for age of spouse, year of diagnosis, and birth order. Rate ratios (RRs) were calculated by Poisson regression. Maternal age was associated with sporadic melanoma and leukemia, causing a 30% excess if mothers were more than 40 years vs. less than 20 years of age. A marginal effect of about 10% of both maternal and paternal age was observed for sporadic breast cancer. Paternal age increased the RR of sporadic nervous system cancer by about 15%. Accumulation of chromosomal aberrations and mutations during the maturation of germ cells may be a mechanism for these findings. In familial cancers of colon, melanoma, and thyroid, higher age showed an apparent protective effect, which was also noted for sporadic cervical cancer and melanoma. The results argue against major age-induced mutagenic/carcinogenic effects on germ cells as well as against age-induced adverse cancer-related hormonal effects during pregnancy. Because two or more mutations are required for adult cancers, however, these cancers may be an insensitive indicator of germ cell mutagenesis. PMID- 10535792 TI - Reproductive risk factors for mucinous and non-mucinous epithelial ovarian cancer. AB - We evaluated reproductive risk factors for mucinous and non-mucinous tumors in a population-based case-control study of epithelial ovarian cancer among women ages 20-79 years. We observed a reduction in risk of tumors of both types in association with one or more full-term pregnancies and with use of oral contraceptives for 5 or more years. While findings of some previous studies support the hypothesis that certain aspects of a woman's reproductive life have a different impact on the risk of these subtypes of ovarian epithelial cancer, our data suggest that this issue is not yet resolved. PMID- 10535791 TI - Exposure to phenoxy herbicides and the risk of spontaneous abortion. AB - The Ontario Farm Family Health Study was designed to assess retrospectively the potential adverse effects of exposure to pesticides on pregnancy. Information on the health and life style of approximately 2,000 farm couples, as well as a history of use of pesticides on the farm, was collected by questionnaire. This analysis focuses on pre- and postconception exposure to phenoxy herbicides and the risk of spontaneous abortion using the complete (to date) pregnancy history for each woman. Preconception exposure (from 3 months before conception to the month of conception) was weakly associated with the risk of spontaneous abortion at <20 weeks' gestation [adjusted odds ratio (OR) = 1.1; 95% confidence interval (CI) = 0.6-1.9]. When the analyses were restricted to spontaneous abortions of <12 weeks, the risk was more than doubled (adjusted OR = 2.5; 95% CI = 1.0-6.4), but the results were sensitive to the cutpoint used. If the husband did not normally wear protective equipment during application, the crude OR for early spontaneous abortions was 5.0 (95% CI = 0.7-36.2). Exposure to phenoxy herbicides during the first trimester was generally not associated with increased risk of spontaneous abortion. The results suggest a possible role of preconception (possibly paternal) exposures to phenoxy herbicides in the risk of early spontaneous abortions. PMID- 10535793 TI - Risk of gallstone disease is associated with serum level of alpha-1-acid glycoprotein. AB - We explored the association between serum level of alpha-1-acid glycoprotein (AGP, an acute phase protein) and the risk of gallstones. AGP was determined in stored serum samples from a case-control study (cases: 113 patients who underwent cholecystectomy for gallstone disease; controls: 184 surgery patients screened by ultrasound, showing no gallstones). Serum AGP correlated negatively with HDL cholesterol and positively with triglycerides. Presence of gallstones was positively associated with serum AGP, more strongly so for cholesterol gallstones than for pigment gallstones, but not differently between solitary and multiple cholesterol stones. Asymptomatic gallstones (13 surgery patients with gallstones found by screening) were also associated with serum AGP levels. A causal role of elevation of serum AGP (or underlying immune activation) in gallstone formation is discussed. PMID- 10535794 TI - The joint impact of family history of myocardial infarction and other risk factors on 12-year coronary heart disease mortality. AB - We investigated the impact of family history of myocardial infarction on 12-year coronary heart disease mortality. Men and women with a family history had an increased risk for coronary heart disease death, irrespective of other risk factors (RR = 1.58; 95% CI = 1.17-2.13 and RR = 2.12; 95% CI = 1.11-4.05, respectively). Women with a family history seemed to be more susceptible to the detrimental effects of smoking; not to the effects of other risk factors. We found no effect modification by family history in men. PMID- 10535795 TI - Active and passive smoking and the occurrence of natural menopause. AB - We examined smoking in relation to natural menopause in 543 women who prospectively recorded menstrual data from their 20s. Mean age at natural menopause was 0.8 years younger (95% CL = -1.5, -0.0) in 98 women who smoked at menopause compared with 362 never-smokers (RR 1.3, 95% CI = 1.0-1.7). We did not observe a decrease in age at natural menopause in former smokers, a dose-response among current smokers, or a lower age at menopause with passive smoke exposure at home. These results suggest that the effect of smoking on ovarian senescence is limited to active smoking during the menopausal transition. PMID- 10535796 TI - Reproducibility and validity of maternal recall of pregnancy-related events. AB - We assessed the reproducibility and validity of a questionnaire that asks mothers to recall pregnancy-related events from thirty or more years ago. Among 146 women who completed the questionnaire twice, responses were highly reproducible for pre pregnancy height and weight (r = 0.95), pregnancy complications (r = 0.74), substance use (r = 0.80), preterm delivery (r = 0.82), birthweight (r = 0.94), and breastfeeding (r = 0.89). Among 154 women whose questionnaire responses were compared to data collected during their pregnancies, recall was highly accurate for height (r = 0.90), pre-pregnancy weight (r = 0.86), birthweight (r = 0.91), and smoking (sensitivity = 0.86, specificity = 0.94). These findings suggest that long-term maternal recall is both reproducible and accurate for many factors related to pregnancy and delivery. PMID- 10535797 TI - What you should have learned about epidemiologic data analysis. PMID- 10535798 TI - Sensitivity analysis of selective migration in ecologic comparisons of health. PMID- 10535799 TI - New options to test the melatonin hypothesis. PMID- 10535800 TI - Angiosarcoma of the scalp and use of a cordless (portable) telephone. PMID- 10535801 TI - Exposure to nitrosamines and mortality from salivary gland cancer among rubber workers. PMID- 10535802 TI - Electric blankets a warm cancerous feeling. PMID- 10535803 TI - Occupational factors for sarcoma subtypes. PMID- 10535804 TI - Further evidence that routinely reported sexually transmitted diseases presage the evolution of the AIDS epidemic. PMID- 10535805 TI - Omeprazole and risk of recurrent bleeding. PMID- 10535806 TI - Hepatitis B vaccine and liver problems in U.S. children less than 6 years old. PMID- 10535807 TI - A woman's own birth weight and gestational age and risk of preeclampsia. PMID- 10535808 TI - Why race is differentially classified on U.S. birth and infant death certificates. PMID- 10535809 TI - The modulation of corneal keratocyte and epithelial cell responses to poly(2 hydroxyethyl methacrylate) hydrogel surfaces: phosphorylation decreases collagenase production in vitro. AB - We examined the regulation of collagenase production by rabbit keratocyte, epithelial and mixed keratocyte/epithelial cell cultures which were exposed to poly(2-hydroxyethyl methacrylate) (PHEMA) hydrogel surfaces with different chemistries and morphologies (sponge and homogeneous gels). Tissue culture modified polystyrene (TCP), used as a control surface, induced the maximum collagenase response with all cell culture types. Copolymer homogeneous gels containing 2-ethoxyethyl methacrylate (EEMA) or methyl methacrylate (MMA) induced a high response in keratocyte cultures, whilst PHEMA hydrogels induced a moderate response and the phosphorylated PHEMA (phos-PHEMA) hydrogel induced no response. Epithelial cells cultured on PHEMA, copolymer and phos-PHEMA hydrogels produced less collagenase activity than the keratocyte cells. The profile of collagenases produced by epithelial cells in response to phos-PHEMA was different to that for the other hydrogels. Co-cultured cells produced higher levels of collagenase (relative to the TCP) in response to hydrogels than did either the keratocytes or epithelial cells alone, but the response of phos-PHEMA was still the lowest. The overall enzyme response to the sponge hydrogels was lower than that to the homogeneous hydrogels, although this effect was less prominent in the keratocyte cultures. The markedly reduced and alternative collagenase responses to phosphorylated surfaces was not a consequence of cell death, and may be a phenomenon related to changes in cell surface charge and morphology. PMID- 10535810 TI - Is cellulose sponge degradable or stable as implantation material? An in vivo subcutaneous study in the rat. AB - The long-term behaviour of cellulose sponge implants, 10 x 10 x 5 mm in size, and tissue reactions in and around them were examined in the subcutaneous tissue of the rat from 1 to 60 weeks after implantation. The cellulose sponge used was filled up with connective tissue 4 to 8 weeks after implantation. Histologically, moderate foreign body tissue reaction inside the implant, the appearance of cracks and fissures, spotty colouration, and softening of the pore walls were observed up to 16 weeks after implantation. Later, the foreign body reaction inside the sponge became milder, the spotty colouration disappeared and micropores enlarged in the viscose cellulose matrix. Histomorphometrically, the cross-sectional area of the implants and the size of the pore wall fragments decreased, and the number of pore wall fragments increased significantly. The cellulose sponge used can be regarded as a slowly degradable implantation material. However, the time needed for the total disappearance of the cellulose sponge from subcutaneous tissue is longer than the 60 weeks. PMID- 10535811 TI - Biocompatibility of a novel tissue connector for fixation of tracheostoma valves and shunt valves. AB - Rehabilitation after laryngectomy often includes the use of a shunt valve and a tracheostoma valve to restore voice. To improve the fixation method of these valves, a new tissue connector has been developed, basically consisting of a ring that will be integrated into surrounding tracheal soft tissue. The valves can be placed in the ring. To test the principle of the tissue connector, a prototype consisting of a subcutaneous polypropylene mesh and a percutaneous titanium stylus was implanted into the backskin of 10 rats by a two-stage surgical procedure. We reasoned that if a firm connection can be realized with the skin, a firm connection with the trachea will also be possible. The subcutaneous part was implanted first, followed by the percutaneous part after 6 weeks. The complete tissue connector with surrounding tissue was removed 8 weeks later and examined histologically. The principle of the new tissue connector proved to be effective: hardly any epithelial downgrowth appeared, and adhesion of soft tissue was demonstrated. No infection or severe inflammation reaction was detected. The tissue connector seems appropriate for its intended use. PMID- 10535812 TI - Guided tissue fabrication from periosteum using preformed biodegradable polymer scaffolds. AB - A successful tissue engineering method for bone replacement would imitate natural bone graft by providing the essential elements for new bone formation using synthetic scaffolds, osteogenic cell populations, and bone induction factors. This is a study of the suitability of various formulations of poly(DL-lactic-co glycolic acid) (PLGA) foams to provide a tissue conducting scaffold in an ovine model for bone flap fabrication. Three formulations were used of different copolymer ratio and molecular weight. Porous wafers of PLGA were stacked into rectangular chambers (volume 4 cm3) enclosed on five sides. Some chambers also contained autologous morcellized bone graft (MBG). The chambers were inserted with the open face adjacent to the cambium layer of the periosteum in rib beds of seven sheep and harvested after 8 weeks in vivo. Gross and histologic examination of the resulting tissue specimens demonstrated molded units of vascularized tissue generally conforming to the shape of the chambers and firmly attached to the periosteum. Polymer degradation appeared to occur by varying degrees based on polymer formulation. New bone formation was observed only in areas containing MBG. There was no evidence of significant inflammatory reaction or local tissue damage at 8 weeks. We conclude that a PLGA foam scaffold is (1) an efficient conductor of new tissue growth but not osteoinductive, (2) contributes to the shape of molded tissue, and (3) biocompatible when used in this model. Further studies are warranted to develop practical methods to deliver bone induction factors to the system to promote osseous tissue generation throughout the synthetic scaffold. PMID- 10535813 TI - Effects of alginate composition on the metabolic, secretory, and growth characteristics of entrapped beta TC3 mouse insulinoma cells. AB - The effects of alginate composition on cell growth as well as the metabolic and secretory profile of transformed beta-cells entrapped in alginate/poly-L lysine/alginate (APA) solid beads were investigated following entrapment of beta TC3 mouse insulinoma cells in alginate composed of either high mannuronic acid or high guluronic acid residues. Entrapped cultures were maintained in spinner flasks for 40-60 days. The pattern of cell growth and the overall rates of glucose consumption and insulin secretion were investigated. Cultures of beta TC3 cells entrapped in alginate composed predominantly of mannuronic acid units (77%) displayed a linear increase in the rates of glucose consumption and insulin secretion concomitant with an increase in cell population in the periphery of the beads. Conversely, cultures of beta TC3 cells entrapped in alginate composed predominantly of high guluronic acid units (69%) displayed a decrease in the rates of glucose consumption and insulin secretion during the first three weeks of culture, followed by a rapid recovery that surpassed the initial rates by day 40. This biphasic pattern was concomitant to a decrease in viable cells during the first three weeks as ascertained by histology, followed by an increase in cell proliferation. Cell growth in high guluronic acid alginate took place at random locations throughout the solid bead and not in the periphery, as was the case in high mannuronic acid alginate preparations. Possible reasons for these differences and the significance of these findings in the context of a bioartificial pancreas composed of APA entrapped transformed cells are discussed. PMID- 10535814 TI - Incorporation of particulated bone implants in the facial skeleton. AB - The purpose of this study was to compare the regenerative response of autogenous cortical and cancellous bone chips and a natural particulate resorbable bone mineral (RBM) (Bio-Oss, Geistlich-Pharma, Wolhusen, Switzerland) in standardized bony defects relating paranasal sinuses to one another and to bone blocks. On 13 skeletally mature female goats four standardized critical-sized full thickness bone defects were made in the frontal bone overlying the frontal sinus. These defects were filled at random with cortical bone chips, cancellous bone chips, spongiosa granules of a RBM or left empty. Fluorochrome bone markers were injected subcutaneously 1 and 5 weeks after transplantation, and one week before the animals were killed. The animals were killed at 3, 6, 12 and 24 weeks after surgery. Autogenous cancellous bone chips is the material of choice for bridging a bony defect in the maxillofacial area where there is no need for mechanical strength. They heal in the same way as cancellous bone blocks do. Cortical bone chips are not reliable enough to be used as a solitary bone-grafting material under these conditions. A cortical block as a solitary implant gives better results. RBM granules as solitary implant in a critical-sized defect do not stimulate osteoconduction but give rise to an extensive osteoclastic activity stimulated by the mutual loose relation. A solid block of RBM is in a similar case more reliable. PMID- 10535815 TI - On morphology of consolidated UHMWPE resin in hip cups. AB - The microstructure of replacement hip cups made from ultra-high molecular weight polyethylene (UHMWPE) has been revealed by permanganic etching to bring out the fine details. Specimens are examined by optical microscopy according to the Nomarski technique, and by scanning and transmission electron microscopy. In all cups the original reactor particles are visible. Optical microscopy is most productive in revealing variations not only in the texture of different individual particles, but also in the degree of consolidation of a particle with its neighbours. Both these factors differ according to the material and process. Electron microscopy reveals the existence of pockets of lower molecular weight material which has been expelled from the particles by the original sintering process. PMID- 10535816 TI - Radiopaque acrylic cements prepared with a new acrylic derivative of iodo quinoline. AB - A novel iodine-containing methacrylate, 2,5-diiodo-8-quinolyl methacrylate, has been synthesized and used in the preparation of acrylic radiopaque cements. The effect of incorporation of this monomer to the self-curing resins, on the curing parameters, swelling behaviour and mechanical properties was studied. The incorporation of the radiopaque compound 2,5-diiodo-8-hydroxyquinoline to the solid phase was also carried out for comparative experiments. A decrease in the peak temperature and an increase in the setting time was observed with the addition of the radiopaque monomer, however, the curing parameters did not appreciably change with the addition of the radiopaque compound to the solid phase. Swelling of the modified cements was in the same range as that of the radiolucent cement; however, the diffusion coefficients calculated according to the Fick's law were higher for the iodine-containing materials. The addition of 5 wt% of the iodine-containing methacrylate provided a significant increase in the tensile properties with respect to either control radiolucent formulations or BaSO4-containing formulations. Biocompatibility of the modified cements was studied by implantation of rods of the cements into rats and histological analysis of the surrounding tissue. PMID- 10535817 TI - Viscoelastic properties of some soft lining materials. II--Ageing characteristics. AB - A non-resonant forced vibration, dynamic mechanical analyser was employed to measure the viscoelastic characteristics of soft lining materials at 1 Hz, after storage in distilled water at mouth temperature for periods up to and including one year. The six commercial products included methacrylate, silicone, and phosphazine based polymers and the one experimental material was a methacrylate. Water sorption of the soft liners, recorded by change in sample mass, ranged from -4.39 to +48.57% and their solubilities from 0.13 to 13.58%, after one year. The heat-cured silicone was the most stable polymer in water in contrast to its autopolymerised counterpart. The excessive water uptake of this latter material resulted in a massive reduction in modulus. At the other extreme one methacrylate with a high plasticiser content hardened substantially after ageing (modulus changed from 5.87 to 72.3 MPa). Changes in loss tangent data were relatively small for all the polymers tested, even for materials with high water uptake. Reduced leaching and/or plasticiser content have led to a more stable generation of soft lining materials. PMID- 10535818 TI - Complement activation by model drug carriers for intravenous application: determination by two-dimensional electrophoresis. AB - The interactions of intravenously injected drug carriers with blood proteins are considered as an important factor for the fate of the particles after their administration. Protein adsorption on latex particles applied as model for intravenous drug carriers was analysed using two-dimensional electrophoresis (2 DE). The particles were incubated in citrated plasma, serum and heat-inactivated serum, respectively. Incubation in the various media resulted in clear differences in the protein adsorption patterns. Two characteristic protein spots were determined to be enriched on the 2-DE gels only after incubation of the particles in serum. Employing N-terminal microsequencing these protein spots were identified to be fragments of the complement protein C3. Enrichment of these particular spots was most likely a result of complement activation by the particles. Mechanism of C3 binding to the particle surface and subsequent inactivation by cleavage are discussed in order to explain the results. It could be demonstrated that 2-DE analysis provides the possibility to distinguish between adsorption and covalent attachment of C3 to particulate surfaces. The findings indicate that complement activation was caused by covalent binding of the C3 component C3b to the particles' surface. The influence of the incubation medium on the in vitro protein adsorption of particulate drug carriers has to be considered when a correlation between the protein adsorption pattern and the in vivo behaviour of the particles is approached. PMID- 10535819 TI - Serum albumin-alginate coated beads: mechanical properties and stability. AB - According to a previously described method, alginate beads were prepared from a Na-alginate solution containing propylene glycol alginate (PGA) and human serum albumin (HSA). The solution was added dropwise to a CaCl2 solution. The beads were treated with NaOH, which started the formation of amide bonds between HSA and PGA at the periphery, giving a membrane. Batches of beads with increasingly thick membranes were prepared using growing concentrations of NaOH, and studied with a texture analyser. When raising NaOH concentration, the rupture strength progressively increased, and the resistance strength to a deformation of 50% of total height also increased before slightly decreasing for the highest NaOH concentration. Variations of bead elasticity were also observed. When the beads were prepared with saline reducing gelation time from 10 to 5 min, and reaction time from 15 to 5 min, mechanical properties varied more progressively with the NaOH concentration, while the results became more reproducible. A series of assays conducted with 0.01 M NaOH confirmed the importance of using a short gelation time, and saline rather than water. Stability assays were also performed. The results were compared to those of alginate-polylysine coated beads and showed the interest of the transacylation method. PMID- 10535820 TI - Glass-reinforced hydroxyapatite composites: fracture toughness and hardness dependence on microstructural characteristics. AB - Fracture toughness and hardness properties of CaO-P2O5 glass-reinforced hydroxyapatite composites have been assessed using indentation techniques and results calculated according to Laugier and Evans' equations. Both properties showed to be dependent upon several microstructural characteristics, namely residual porosity and the percentage of secondary beta and alpha tricalcium phosphate phases in the structure of the composites. Composites presented a Palmqvist-type indentation crack system, which is the specific crack system addressed by Laugier's approach. Fracture toughness determinations according to Evan's equation, which is a universal one and adapted to both median and Palmqvist crack systems, did not correlate well with Laugier determinations. PMID- 10535821 TI - Developmental anomalies of the outflow tracts and aortic arch: towards an understanding of the role of deletions within the 22nd chromosome. PMID- 10535822 TI - Can transcatheter closure of atrial septal defect be regarded as a 'standard' procedure? PMID- 10535823 TI - Spontaneous occlusion of systemic-to-pulmonary arterial shunts. PMID- 10535824 TI - Interruption of the aortic arch associated with deletion of chromosome 22q11 is associated with a subarterial and doubly committed ventricular septal defect in Japanese patients. AB - OBJECTIVES: The purpose of this study was to clarify the clinical characteristics of interruption of the aortic arch associated with chromosome 22q11deletion. BACKGROUND: About half of patients with interruption of the aortic arch between the left common carotid and the left subclavian artery have deletion of chromosome 22q11. METHODS: In total, 20 patients with interruption of the aortic arch were studied with fluorescence in situ hybridization using peripheral lymphocytes and a DiGeorge syndrome chromosomal probe (Oncor N25). Cardiovascular anomalies in these patients were diagnosed by cross-sectional echocardiography and angiocardiography, and were confirmed at intracardiac repair. RESULTS: Of 13 patients with interruption between the left common carotid artery and the left subclavian artery, seven had the deletion. All 7 also showed thymic hypoplasia and hypocalcemia, together with a nasal voice and peculiar facies. Six of the seven patients had complete deficiency of the muscular outlet septum, with the defect extending to the perimembranous area. Such complete absence of the muscular outlet septum was not present in any of the patients without the deletion. CONCLUSIONS: Interruption of the aortic arch between the left common carotid and the left subclavian artery, absence of the thymus, and complete absence of the muscular outlet septum, were characteristic in Japanese patients with interruption of the aortic arch associated with deletion of chromosome 22q11. PMID- 10535825 TI - Transcatheter closure as standard treatment for most interatrial defects: experience in 200 patients treated with the Amplatzer Septal Occluder. AB - To judge whether an Amplatzer Septal Occluder can be used as standard therapy instead of surgery for closure of atrial septal defects we report our experiences in 200 patients. Of these patients, 127 had an atrial septal defect with haemodynamically significant left-to-right shunt, 68 patients a persistent oval foramen after presumed paradoxical embolism, and 5 had a fenestration after Fontan-repair. Mean age was 29.8 years (0.8 to 77.7 years). Body weight ranged from 6.9 to 120.0 kg (mean 51.5 kg). After diagnostic cardiac catheterization, and balloon-sizing of the defect, we implanted Amplatzer Septal Occluders with stents of 4 to 28 mm diameter. Follow-up studies were obtained after 48 hours, and one, six, and twelve months. Transcatheter closure of the atrial septal defect proved successful in all without any relevant residual shunts. In particular, complete closure was achieved in all patients after presumed paradoxical embolism. The mean period of follow-up is 9.5 months, with a range from 0.4 to 23.5 months, giving a total of 1898 patient months. The occlusion rate after three month was 98.1%. A trivial haemodynamically insignificant residual shunt remained in 1.9% of the patients. Fluoroscopy times ranged from 0 to 43.5 minutes, with a median of 8.7 minutes. The excellent results in the short and medium term in children and adults have resulted in using this device routinely at the present time for closure of central atrial septal defects up to a diameter of 28 mm. Final judgement, however, is only possible after long-term follow-up. PMID- 10535826 TI - Reduced frequency of occlusion of aorto-pulmonary shunts in infants receiving aspirin. AB - OBJECTIVE: Infants with severely reduced pulmonary perfusion due to complex congenital cardiac malformations are in need of an improved flow of blood to the lungs. One option for treatment is to construct a systemic-to-pulmonary arterial shunt. Although such shunts have been used since 1945, their spontaneous occlusion remains a major problem in the long-term. DESIGN: We studied all infants in whom a systemic-to-pulmonary arterial shunt had been constructed using a Gore-Tex tube graft between December 1989 and March 1996. PATIENTS: Of 46 infants undergoing construction of a shunt, 7 (15%) died within 30 days of surgery. The shunts had to be taken down in 2 infants. Thus, 37 infants were included in the study. All but three infants received Aspirin. Aspirin was discontinued on the personal decision of individual physicians. Of 22 infants, 3 never received Aspirin, and in 19 it was stopped well before undertaking subsequent surgery. Aspirin was administered continuously to 15 infants until further surgery. RESULTS: Those in whom Aspirin was discontinued, or not given, and those receiving Aspirin until further surgery, were comparable concerning their age, time of follow-up, severity of the cardiac lesions, and size and type of shunt. Partial or complete occlusion of the shunt occurred in 2 of 15 (13%) infants taking Aspirin, but was seen in 12 of 22 (54%) infants in whom Aspirin was discontinued. Of these, 3 died due to acute occlusion of the shunt. CONCLUSIONS: Aspirin reduced effectively the rate of occlusion of systemic-to pulmonary arterial shunts, and should be continued as long as the shunt is in place. PMID- 10535827 TI - Unguarded mitral orifice, mirror-imaged atrial arrangement, and discordant atrioventricular connections. AB - We report two autopsy proven cases of unguarded mitral orifice associated with mirror-imaged atrial arrangement, discordant atrioventricular connections, double outlet right ventricle, pulmonary valvar stenosis or atresia, and atrialisation of the morphologically left ventricle. The morphologically left atrioventricular junction was devoid of valvar leaflets, and there was no tension apparatus within the ventricle. To the best of our knowledge, this is the first description of this rare cardiac malformation. PMID- 10535828 TI - Comparison of acute effects on right ventricular haemodynamics of surgical versus interventional closure of atrial septal defects. AB - OBJECTIVES: We compared the acute effects on right ventricular haemodynamics of surgical versus transcatheter closure of medium-sized atrial septal defects. METHODS: We studied 47 consecutive patients with a defect in the oval fossa and a ratio of pulmonary to systemic flows between 1.5: 1 and 2: 1. They were divided into two groups according to whether the defects were closed by surgery, performed in 23 patients, or by interventional catheterization, achieved in 24 patients. By means of transthoracic cross-sectional echocardiography, we measured right ventricular end-diastolic and endsystolic volumes and calculated ejection fractions. These calculations were performed before, and between 1 and 7 days after closure of the defect. RESULTS: Before closure of the defect, all patients had an enlarged right ventricle with normal function. After closure by either method, there was no difference in the rate of normalization of end-diastolic volume, but endsystolic volume remained enlarged. Thus, the calculated ejection fraction was lower than before closure. CONCLUSIONS: There was no difference in right ventricular volumes or function early after closure of atrial septal defects, irrespective of whether this was achieved surgically or via transcatheter closure. PMID- 10535829 TI - The aortic valve with two leaflets--a study in 2,000 autopsies. AB - Our study is based on the examination of 2,000 aortic valves obtained from fresh cadavers (1,499 males, 501 females) at the Institute of Forensic Medicine in Rio de Janeiro. We discovered 13 valves having two leaflets, giving a prevalence of 0.65%, much lower than generally reported in the literature. All 13 valves were from males, 10 from whites, three from mulattos, and none from blacks. Special attention was given to the raphe and the leaflets, the calcification of which can lead to stenosis. The only valves with normal texture and flexibility were two obtained from children less than one year old. All the other valves were thickened, and five of them had some degree of calcification. Six valves were judged to be functioning normally, while 7 valves were abnormal, 5 being stenotic and two showing evidence of insufficiency. The insufficiency in one was due to endocarditis, but in the other was due to redundancy of the leaflets. The only other cardiac anomaly discovered in these 13 cases was one patient with aortic coarctation. Bifoliate aortic valve, therefore, is probably the most common cardiac anomaly, although its prevalence as discovered in Brazil is lower than that reported in the literature. It affects mainly white males. After the fourth decade of life, most valves present some thickening, with stenosis being the most common complication. As is well recognised, infective endocarditis and aortic insufficiency are the other frequent complications. It is in general, nonetheless, an isolated anomaly. PMID- 10535830 TI - The prevalence of bifoliate pulmonary valves. PMID- 10535831 TI - Spontaneous closure and thrombosis of a ductal aneurysm in a neonate. AB - The natural history of a ductal aneurysm detected prenatally is presented. Neurologic concerns in the neonate and absence of cardio-respiratory compromise permitted serial echo demonstration of constriction, thickening and subsequent closure with formation of thrombus in the aneurysm. PMID- 10535833 TI - Obstruction of the right ventricular outflow tract caused by a tuberculoma in a patient with ventricular septal defect and aneurysm of the membranous septum. AB - Reported here is an obstruction of the right ventricular outflow tract caused by a tuberculoma in a 15-year-old boy who presented with a ventricular septal defect. The obstruction was discovered at surgery and the tuberculous aetiology was only demonstrated histologically. To the authors' knowledge, this is the first report of a tuberculoma of the heart associated with congenital heart disease. PMID- 10535832 TI - Cyanosis due to diastolic right-to-left shunting across a ventricular septal defect in a patient with repaired tetralogy of Fallot and pulmonary atresia. AB - Cyanosis as a result of right-to-left shunting across a ventricular septal defect is commonly encountered in patients with congenital heart disease when systolic pressure in the right ventricle exceeds that in the left ventricle. Reported is the case of a child who remained cyanosed after surgical correction of pulmonary atresia despite right ventricular systolic pressure being lower than left ventricular pressure. Colour-flow Doppler showed a residual ventricular septal defect, with right-to-left shunting in diastole alone. PMID- 10535834 TI - Pulmonary atresia with intact ventricular septum, right-sided aortic arch, and an aorto-pulmonary collateral artery. AB - Described is a rare association in a patient with the heart in the left chest, namely pulmonary atresia with intact ventricular septum, fistulous coronary arterial connections, a right-sided aortic arch and an aorto-pulmonary collateral artery feeding one lung. The pulmonary arteries were non-confluent, with the right lung supplied by the right arterial duct originating from the under surface of the right-sided aortic arch, and the left lung supplied through the aorto pulmonary collateral artery arising from the descending aorta. The surgical management is different in the setting of non-confluent pulmonary arteries. PMID- 10535835 TI - Interruption of the aortic arch at the isthmus with DiGeorge syndrome and 22q11.2 deletion. AB - A 6-day-old male with interruption of the aortic arch at the isthmus (type A) had the typical phenotype of DiGeorge syndrome. There was also a doubly committed juxta-arterial ventricular septal defect and an unobstructed left ventricular outflow tract. Hypoplasia of the thymus was confirmed during a modified Blalock Park operation. He had persistent hypocalcemia, and was susceptible to infection. He was subsequently revealed by the use of fluorescence in situ hybridization analysis to have 22q11.2 deletion. Interruption of the aortic arch at the isthmus is presumed to reflect abnormal fetal hemodynamics, and is considered a distinct pathogenetic entity from interruption between the left common carotid and subclavian arteries, the latter being the variant more frequently associated with DiGeorge syndrome. In our case, the 22q11.2 deletion likely played a major role in the etiology of the interrupted aortic arch. PMID- 10535836 TI - Rapid reversal of dilated cardiomyopathy following removal of neuroblastoma. AB - Reported is a child with dilated cardiomyopathy, in whom medical therapy resulted in a mild improvement of cardiac function. Metabolic studies suggested the presence of a catecholamine-secreting tumour; and an adrenal neuroblastoma was identified and surgically removed. Following surgery, there was progressive and complete normalization of cardiac function. Although very rare, neurogenic tumours may be involved in the development of a dilated cardiomyopathy in the infant and child. PMID- 10535837 TI - Cardiac anomalies associated with congenital absence of the portal vein. AB - The authors discovered congenital absence of the portal vein, with visceral venous return to the right atrium, in a 5-year-old girl with aortic valvar stenosis. Interestingly, of the 19 patients, it was discovered that 11 reported with portal venous agenesis also had cardiac defects. We have, therefore, investigated the hypothesis that the congenital absence of the portal vein and the associated cardiac malformations may result from a similar embryologic insult, and that cardiac development may be affected by the systemic diversion of portal venous flow. PMID- 10535838 TI - The syndrome of pericarditis, arthritis, and camptodactyly: an under-recognized cause of pericardial constriction in children? AB - The syndrome encompassing the combination of pericarditis, arthritis, and camptodactyly is a rarely described cause of pericardial constriction in children. It is likely that this association is being under-recognized. We report a new case in which the skeletal abnormalities were subtle. The syndrome should be included in the differential diagnosis of any child with persistent non inflammatory pericardial effusion. A careful search at the bedside for the associated skeletal abnormalities should lead to the correct diagnosis. Pericardiectomy is the treatment of choice. PMID- 10535839 TI - Magnetic resonance imaging comes of age. PMID- 10535840 TI - Unusual malformation of the aortic arch in a patient with a large facial hemangioma. PMID- 10535841 TI - Impact of new technologies in ophthalmology. PMID- 10535842 TI - Transpupillary thermotherapy of choroidal melanoma with or without brachytherapy: a dilemma. PMID- 10535843 TI - Ophthalmology in the post-genomic era. PMID- 10535844 TI - Molecular therapy in ocular wound healing. PMID- 10535845 TI - New developments in sustained release drug delivery for the treatment of intraocular disease. PMID- 10535847 TI - Closer to nature: new biomaterials and tissue engineering in ophthalmology. PMID- 10535848 TI - Laser imaging of the retina. PMID- 10535846 TI - Immunomodulation of autoimmune responses with monoclonal antibodies and immunoadhesins: treatment of ocular inflammatory disease in the next millennium. PMID- 10535849 TI - New applications in ultrasound technology. PMID- 10535850 TI - Scanning laser ophthalmoscopy and fundus fluorescent leucocyte angiography. PMID- 10535851 TI - Telemedicine. PMID- 10535852 TI - Refractive surgery. PMID- 10535854 TI - Information technology in ophthalmology-experience with an electronic patient record. PMID- 10535853 TI - Eradication of trachoma worldwide. PMID- 10535855 TI - Mycophenolate mofetil versus cyclosporin A in high risk keratoplasty patients: a prospectively randomised clinical trial. AB - BACKGROUND/AIMS: The requirement for an effective, minimally toxic immunosuppressive agent remains a major obstacle to performing high risk corneal transplantation. Although therapy with cyclosporin A (CSA) allows superior graft survival, its use is limited because of a wide range of side effects. Mycophenolate mofetil (MMF) has been shown to be a safe and effective immunosuppressive agent following renal transplantation. This prospective, randomised clinical trial was carried out to investigate the efficacy and safety of MMF in preventing corneal allograft rejection. METHODS: Recipients of corneal transplants who were at high risk for graft failure were randomly assigned to either CSA or MMF immunosuppressive therapy. CSA was given in doses to achieve whole blood trough levels of 120-150 ng/ml. MMF was given in a daily dose of 2 g. Both therapy groups additionally received oral corticosteroids (fluocortolone 1 mg/kg) which were tapered and discontinued within the first 3 postoperative weeks. Patients were monitored closely for evidence of corneal graft rejection and adverse side effects. Drug efficacy was measured, primarily, by the number of patients who experienced at least one episode of clinical graft rejection. Safety analysis focused on reported adverse side effects and laboratory measurements. RESULTS: 41 patients were enrolled in the study. There was no statistically significant difference between the two groups. 20 patients received CSA and 21 patients received MMF. Two patients in each group showed evidence of acute graft rejection which could be treated effectively by corticosteroids. All corneal grafts remained clear throughout the follow up. CONCLUSIONS: In this study it was shown that MMF is just as effective as CSA in preventing acute rejection following high risk corneal transplantation. Mycophenolate mofetil represents a promising alternative therapeutic option in patients who are at high risk for corneal graft failure. PMID- 10535856 TI - Outcome of cataract surgery considering the preoperative situation: a study of possible predictors of the functional outcome. AB - AIM: To analyse possible predictors of the self assessed functional outcome of a cataract extraction. METHODS: The patients' self assessed visual function was studied by use of a questionnaire, the "Catquest", before and 6 months after surgery. All patients (n=1933, mean age 75.5 years, 66.8% women) who were undergoing cataract surgery in March 1995, in 35 different departments of ophthalmology participating in the National Swedish Cataract Register, were included in the study. A routine ophthalmic examination was performed before and after surgery. The following preoperative variables were studied with regard to a possible relation to the outcome: age, sex, ocular comorbidity, best corrected preoperative vision (better eye), first or second eye surgery, other diseases with a need for long term medication, need for home help, need for subsidised travel by taxi. RESULTS: Ocular comorbidity was strongly related to a "no benefit" outcome after surgery (p= 0.005). Second eye surgery and young age was related to a "very good benefit" outcome after surgery (p=0.0001 and p<0.0001 respectively). CONCLUSIONS: Patients with an ocular comorbidity in the eye undergoing a cataract extraction were characterised by a significantly higher frequency of deteriorated self assessed visual function after surgery than patients with no ocular comorbidity. The highest degree of improvement was most frequently found in younger patients undergoing second eye surgery. PMID- 10535857 TI - Oral vitamins C and E as additional treatment in patients with acute anterior uveitis: a randomised double masked study in 145 patients. AB - AIM: To investigate the effect of additional oral vitamins C and E on acute anterior uveitis. METHODS: A placebo controlled double masked study on the effect of vitamin C 500 mg in combination with vitamin E 100 mg twice daily in 145 patients with acute anterior uveitis. As a primary end point variable, laser cell/flare measurements were performed. Best corrected and stenopeic visual acuity (VA) testing and clinical variable scores were measured. RESULTS: Laser flare measurements (ph/s) before treatment were 207.1 (SD 258) in the vitamin group and 143.6 (156) in the placebo group. After 3 days corresponding values were 80.2 (129) and 54.7 (82), after 7 days 89.2 (187) (12.5) and 85.8 (208), after 14 days 47.1 (109.5) and 40.5 (116) after 28 days 23.1 (53.6) and 23.1 (48), and after 56 days 15.6 (26) and 15.3 (17). There was no significant difference in time trend between the two treatment groups (RMANOVA; p = 0.53). Baseline VA (logMAR) was 0.106 (0.241) in the vitamin group and 0.128 (0.456) in the placebo group. VA after 3 days was 0. 236 (0.293) and 0.344 ( 0.489), after 7 days 0.204 (0.292) and 0.292 (0.479), after 14 days 0.162 (0.274) and 0.193 (0.454), after 28 days 0.096 (0.232) and 0.158 (0.436), and 0.026 (0.213) and 0.106 (0. 437) after 56 days. Although no significant difference in time trend was detected, evaluation of the VA data of the last time point (56 days) by means of the Mann-Whitney test showed a significantly better VA in the vitamin group (p = 0.01). CONCLUSIONS: There was no significant effect of vitamins C and E on laser flare measurements. The significant effect of the oral vitamins on visual acuity at 8 weeks after start of the oral vitamins C and E may indicate a protective effect in patients with acute anterior uveitis. PMID- 10535858 TI - Measuring colour rivalry suppression in amblyopia. AB - AIMS: To determine if the colour rivalry suppression is an index of the visual impairment in amblyopia and if the stereopsis and fusion evaluator (SAFE) instrument is a reliable indicator of the difference in visual input from the two eyes. METHODS: To test the accuracy of the SAFE instrument for measuring the visual input from the two eyes, colour rivalry suppression was measured in six normal subjects. A test neutral density filter (NDF) was placed before one eye to induce a temporary relative afferent defect and the subject selected the NDF before the fellow eye to neutralise the test NDF. In a non-paediatric private practice, 24 consecutive patients diagnosed with unilateral amblyopia were tested with the SAFE. Of the 24 amblyopes, 14 qualified for the study because they were able to fuse images and had no comorbid disease. The relation between depth of colour rivalry suppression, stereoacuity, and interocular difference in logMAR acuity was analysed. RESULTS: In normal subjects, the SAFE instrument reversed temporary defects of 0.3 to 1. 8 log units to within 0.6 log units. In amblyopes, the NDF to reverse colour rivalry suppression was positively related to interocular difference in logMAR acuity (beta=1.21, p<0.0001), and negatively related to stereoacuity (beta=-0.16, p=0.019). The interocular difference in logMAR acuity was negatively related to stereoacuity (beta=-0.13, p=0.009). CONCLUSIONS: Colour rivalry suppression as measured with the SAFE was found to agree closely with the degree of visual acuity impairment in non-paediatric patients with amblyopia. PMID- 10535859 TI - Hyperhomocysteinaemia in young patients with non-arteritic anterior ischaemic optic neuropathy. AB - BACKGROUND/AIM: Elevated plasma homocysteine is a newly identified vascular risk factor among patients under age 55 years with cerebrovascular, cardiovascular, or peripheral vascular disease. This study sought to evaluate retrospectively the plasma homocysteine status among healthy younger patients with ischaemic optic disc disease. METHODS: 12 non-diabetic patients who had been diagnosed with non arteritic anterior ischaemic optic neuropathy (NAION) before the age of 50 years were identified from chart review. None had experienced previous ischaemic cerebrovascular, cardiovascular, or peripheral vascular events. Plasma homocysteine, CBC, renal function, vitamin B6, vitamin B12, and folate levels were sampled in the fasting state. RESULTS: Two of 12 patients (17%) had hyperhomocysteinaemia. Both had experienced NAION in both eyes with recurrent episodes. Neither patient was hypertensive nor had a smoking history. One of these two patients had mild hypercholesterolaemia which did not warrant medication. CONCLUSIONS: Elevated plasma homocysteine may be associated with NAION. An evaluation for hyperhomocysteinaemia should be considered in patients with NAION who do not have the typical risk factor such as older age, diabetes, hypertension, or tobacco use. It should also be considered in young patients with bilateral or recurrent attacks of NAION. PMID- 10535860 TI - Localised retinal nerve fibre layer defects in chronic experimental high pressure glaucoma in rhesus monkeys. AB - AIM: To evaluate prospectively in an experimental model of chronic high pressure glaucoma whether the concept of a mainly diffuse pattern of optic nerve damage holds true for high pressure glaucoma. METHODS: The study comprised nine eyes of nine rhesus monkeys (Macaca mulatta) with a mean age of 17.7 (SD 3.1) years (range 13-23 years). Experimental glaucoma was produced by multiple applications of argon laser to the trabecular meshwork. Applanation tonometry was regularly performed and fundus photographs, which were taken serially, were used for retinal nerve fibre layer (RNFL) assessment and morphometric optic disc analysis. Six monkeys, in which arterial hypertension and atherosclerosis had additionally been induced several years before elevation of intraocular pressure, did not show any sign of diffuse loss or localised defects of the RNFL before initiation of glaucoma. RESULTS: Compared with the same eyes at baseline, localised RNFL defects had developed in eight (89%) eyes. It included all three eyes (100%) of the monkeys without arterial hypertension/arteriosclerosis, and five of the six monkeys (83%) with arterial hypertension/arteriosclerosis. Four eyes had multiple localised RNFL defects. In all eyes, diffuse RNFL loss was additionally present. CONCLUSIONS: Besides diffuse loss of RNFL, localised RNFL defects were present in almost all eyes of monkeys with chronic experimental high pressure glaucoma. Challenging the concept that a mostly diffuse type of optic neuropathy occurs in high pressure glaucoma, the results suggest that, in high pressure glaucoma, at least a mixture of localised and diffuse pattern of optic nerve damage prevails. PMID- 10535861 TI - Detection of cytokine mRNA production in infiltrating cells in proliferative vitreoretinopathy using reverse transcription polymerase chain reaction. AB - AIMS: To determine whether the infiltrating cells in the vitreous and subretinal fluid of patients with proliferative vitreoretinopathy (PVR) express messenger RNA for various cytokines found in this condition. METHODS: The presence of mRNA coding for HPRT, IL-6, IL-1beta, IL-8, and TNFalpha was investigated in 20 vitreous and subretinal fluid (SRF) samples from patients with PVR by reverse transcriptase polymerase chain reaction (RT-PCR). 16 samples from patients with retinal detachment and macular holes were used as controls. RESULTS: HPRT was detected in all samples of PVR and in 11 (69%) control cases. Patients with PVR demonstrated mRNA for the cytokines tested more often than controls. The difference was statistically significant. CONCLUSION: The presence of mRNA encoding for IL-6, IL-1beta, IL-8, and TNFalpha is significantly detected by RT PCR in vitreous and SRF samples of patients with PVR, indicating local production of these cytokines by vitreous and SRF cells. PMID- 10535862 TI - Rhinostomies: an open and shut case? AB - AIMS: To analyse bone fragments from rhinostomies of patients undergoing revisional dacryocystorhinostomy, looking for evidence of new bone formation. METHODS: 14 consecutive patients undergoing secondary lacrimal surgery were included in this study. In each case the existing rhinostomy was enlarged with bone punches, care being taken to use the punches with the jaws cutting perpendicularly to the edge of the rhinostomy, to allow accurate orientation of the specimens. The fragments were examined histologically for evidence of new bone formation. RESULTS: Histological sections showed fragments of bone with variable fibrosis at the edge of the rhinostomy. There was evidence of only very little new bone formation. CONCLUSION: This study has clearly shown that, at the edge of a rhinostomy, healing is predominantly by fibrosis and there is only very limited new bone formation. PMID- 10535863 TI - Corneal wound healing following laser in situ keratomileusis (LASIK): a histopathological study in rabbits. AB - AIMS: To investigate the histopathological changes of rabbit corneas after laser in situ keratomileusis (LASIK) and to evaluate the corneal wound healing process. METHODS: A LASIK was performed on white rabbit eyes. Postoperatively, rabbits were killed on days 1 and 7, and at 1, 3, and 9 months. RESULTS: Periodic acid Schiff (PAS) positive material and disorganised collagen fibre were seen along the interface of the corneal flap even 9 months after operation. CONCLUSIONS: The wound healing process still continued at 9 months after LASIK indicating that a much longer time than expected was required for corneal wound healing following LASIK. PMID- 10535864 TI - Recurrent corneal erosion syndrome. PMID- 10535865 TI - Complementary and alternative medicine: the roots of healing. PMID- 10535866 TI - Long-term survival and parenteral nutrition dependence in adult patients with the short bowel syndrome. AB - BACKGROUND & AIMS: The short bowel syndrome (SBS) may be associated with either transient or permanent intestinal failure, presently treated by parenteral nutrition (PN). Survival and PN-dependence probabilities, taking into account both small bowel remnant length and the type of the digestive circuit of anastomosis, are not known in adult SBS patients. The aim of this study was to assess such prognostic factors. METHODS: A total of 124 consecutive adults with nonmalignant SBS were enrolled from 1980 to 1992 at 2 home PN centers. They were analyzed for survival and PN-dependence probabilities using the Cox model and for PN dependence using linear discriminant analysis. Data were updated in April 1996. RESULTS: Survival and PN-dependence probabilities were 86% and 49% and 75% and 45% at 2 and 5 years, respectively. In multivariate analysis, survival was related negatively to end-enterostomy, to small bowel length of <50 cm, and to arterial infarction as a cause of SBS, but not to PN dependence. The latter was related negatively to postduodenal small bowel lengths of <50 and 50-99 cm and to absence of terminal ileum and/or colon in continuity. Cutoff values of small bowel lengths separating transient and permanent intestinal failure were 100, 65, and 30 cm in end-enterostomy, jejunocolic, and jejunoileocolic type of anastomosis, respectively. CONCLUSIONS: In adult SBS patients, small bowel length of <100 cm is highly predictive of permanent intestinal failure. Presence of terminal ileum and/or colon in continuity enhances both weaning off PN and survival probabilities. After 2 years of PN, probability of permanent intestinal failure is 94%. These rates may lead to selection of other treatments, especially intestinal transplantation, instead of PN, for permanent intestinal failure caused by SBS. PMID- 10535867 TI - Esophagopharyngeal acid regurgitation: dual pH monitoring criteria for its detection and insights into mechanisms. AB - BACKGROUND & AIMS: A valid technique for the detection of esophagopharyngeal acid regurgitation would be valuable to evaluate suspected reflux-related otolaryngologic and respiratory disorders. The aim of this study was to derive pH criteria that optimally define esophagopharyngeal acid regurgitation and to examine patterns of regurgitation. METHODS: In 19 healthy controls and 15 patients with suspected regurgitation, dual or quadruple pH sensors were used to monitor pharyngeal and esophageal pH. For each combination of the 2 variables, DeltapH and nadir pH, proportions of pH decreases that occurred during or independent of esophageal acidification were calculated to determine the likelihood that an individual pharyngeal pH decrease was a candidate regurgitation event or a definite artifact. RESULTS: Overall, 92% of pharyngeal pH decreases of 1-2 pH units and 66% of pH decreases of this magnitude reaching a nadir pH of <4 were artifactual. Optimal criteria defining a pharyngeal acid regurgitation event were a pH decrease that occurred during esophageal acidification, had a DeltapH of >2 units, and reached a nadir of <4 units in less than 30 seconds. Regurgitation occurred more frequently in subjects in an upright (32 of 35) than in a supine (3 of 35 events; P 25, diabetes, or hyperlipidemia); serum iron tests and liver iron concentration (LIC; reference value, <36 micromol/g); liver function test results; C282Y and H63D HFE mutations; and liver histological status. RESULTS: Patients were predominantly male and middle-aged. Most (94%) had IRS. Transferrin saturation was increased in 35% (median, 42%; range, 13%-94%). LIC ranged from 38 to 332 micromol/g (median, 90 micromol/g), and LIC/age ratio ranged from 0.5 to 4.8 (median, 1.8). Allelic frequencies of both HFE mutations were significantly increased compared with values in normal controls (C282Y, 20% vs. 9%; H63D, 30% vs. 17%), only because of a higher prevalence of compound heterozygotes. Patients with no HFE mutations had similar degrees of iron overload as those with other genotypes, except for compound heterozygotes, who had slightly more iron burden. Steatosis was present in 25% of patients and NASH in 27%. Portal fibrosis (grades 0-3) was present in 62% of patients (grade 2 or 3 in 12%) in association with steatosis, inflammation, and increased age. Sex ratio, IRS, transferrin saturation, and LIC did not vary with liver damage. Serum ferritin concentration, liver function test results, and fibrosis grade were more elevated in patients with steatosis and NASH than in others, but LIC and allelic frequencies of HFE mutations were similar. CONCLUSIONS: This study shows that patients with unexplained hepatic iron overload are characterized by a mild to moderate iron burden and the nearly constant association of an IRS irrespective of liver damage. PMID- 10535880 TI - A randomized, controlled trial of maintenance interferon therapy for patients with chronic hepatitis C virus and persistent viremia. AB - BACKGROUND & AIMS: : At least half of patients with chronic hepatitis C virus (HCV) fail to respond to interferon or interferon/ribavirin therapy. Histological improvement is observed in some nonresponders. We conducted a randomized, controlled trial to determine if maintenance interferon therapy could prevent histological progression in this subset of nonresponders. METHODS: Fifty-three patients with chronic HCV were enrolled. All were HCV-RNA positive after 6 months of treatment with interferon alfa-2b but had a histological response. Twenty seven of the patients were randomly assigned to continue interferon (3 MU 3 times weekly) for 24 months; 26 patients discontinued treatment and were observed prospectively. Alanine aminotransferase (ALT) level and HCV-RNA titer were monitored, and liver biopsy was repeated every 12 months. RESULTS: Before interferon therapy, the 2 groups were well matched for all demographic factors, serum ALT (94.0 +/- 15.6), log HCV-RNA titer (5. 85 +/- 0.15 copies/mL), histology score (9.5 +/- 0.2), and percentage with cirrhosis (25%). After 6 months of treatment, significant reductions (P < 0.05) in serum ALT level (62.6 +/- 9.6), log HCV-RNA titer (4.79 +/- 0.13 copies/mL), and hepatic inflammation (4.0 +/- 0.2) were observed. These improvements were maintained in the patients randomized to continue interferon. Stopping treatment was associated with an increase in serum ALT, log HCV-RNA, and hepatic inflammation back to baseline. After 30 months of treatment, mean fibrosis score declined from 2.5 to 1.7 and 80% of patients had histological improvement (P < 0.03). Discontinuation of interferon was associated with an increase in mean fibrosis score from 2.2 to 2.4 and worsening of hepatic histology in 30% of patients (P < 0.01). CONCLUSIONS: These data support the hypothesis that maintenance interferon may prevent histological progression of chronic HCV in patients who remain viremic. PMID- 10535882 TI - HLA association of amoxicillin-clavulanate--induced hepatitis. AB - BACKGROUND & AIMS: Drug-induced immunoallergic hepatitis typically affects a minority of patients exposed to a particular drug. Its rarity is believed to be due to metabolic or immunologic idiosyncrasy. The presence of an immunologic idiosyncrasy might imply an HLA association. Previous studies reporting an HLA association of drug-induced hepatitis included only small numbers of patients and used serological HLA typing. METHODS: We studied 35 patients with biopsy documented amoxicillin-clavulanate-induced hepatitis. HLA-A and -B were typed using alloantisera and compared with those of 300 controls (volunteer bone marrow donors). HLA-DRB and -DWB were typed by polymerase chain reaction-line probe assay, with 60 volunteer bone marrow donors serving as controls. RESULTS: The study group was characterized by a higher frequency of DRB1*1501-DRB5*0101 DQB1*0602 haplotype (57.1% vs. 11.7% in controls, P < 0.000005; after correction for the large number of comparisons, P < 0.0002). Patients with DRB1*1501 DRB5*0101-DQB1*0602 haplotype were more likely than patients without it to have a cholestatic (70% vs. 60%) or mixed (30% vs. 13%) than a hepatocellular pattern of hepatitis (0% vs. 27%) (P < 0.05). CONCLUSIONS: Amoxicillin-clavulanate-induced hepatitis is associated with the DRB1*1501-DRB5*0101-DQB1*0602 haplotype. The data support the view that an immunologic idiosyncrasy, mediated through HLA class II antigens, plays a role in the pathogenesis of drug-induced immunoallergic hepatitis. HLA association has a limited impact on the expression of hepatitis. PMID- 10535881 TI - A prospective trial of colchicine and methotrexate in the treatment of primary biliary cirrhosis. AB - BACKGROUND & AIMS: The aim of this study was to determine if colchicine or methotrexate improves blood test results, symptoms, and/or liver histology in patients with primary biliary cirrhosis. METHODS: Patients with histologically confirmed primary biliary cirrhosis whose serum alkaline phosphatase (ALP) levels were at least 2 times above normal and who were not yet candidates for liver transplantation received colchicine or methotrexate and were followed up for 2 years. RESULTS: In patients receiving colchicine (n = 43), mean pruritus score decreased from 1.63 to 1.12 (P = 0.04), ALP level from 494 to 355 U/L (P < 0.0001), and alanine aminotransferase (ALT) level from 79 to 61 U/L (P < 0.0001). In patients receiving methotrexate (n = 42), pruritus score decreased from 1.25 to 0.44 (P = 0.0001), ALP from 478 to 235 U/L (P < 0.0001), and ALT from 96 to 61 U/L (P = 0.0001). Methotrexate but not colchicine significantly improved liver histology (P = 0.005) and serum immunoglobulin G levels (P = 0.0002). Methotrexate improved most blood test results more than colchicine. Serum bilirubin levels increased slightly with each drug, and albumin levels decreased slightly. CONCLUSIONS: Both colchicine and methotrexate improved biochemical test results and symptoms in primary biliary cirrhosis, but the response to methotrexate was greater. PMID- 10535883 TI - Hepatitis C virus NS5A protein inhibits interferon antiviral activity, but the effects do not correlate with clinical response. AB - BACKGROUND & AIMS: Patients with chronic hepatitis C virus infection are commonly treated with interferon alfa (IFN-alpha), but the long-term response rate is poor. A region of NS5A of hepatitis C virus genotype 1 (the ISDR) has been associated with treatment outcome in some patients. NS5A binds to and inhibits PKR in vitro and inhibits IFN-alpha in human cells. We examined the effects of the NS5A protein from patients who did or did not respond to IFN-alpha to determine whether NS5A from IFN-alpha nonresponders inhibited the effects of IFN alpha in vitro. METHODS: We cloned NS5A from patients who had well-characterized responses to IFN-alpha and expressed them in a human fibroblast cell line under the control of an inducible promoter. The NS5A expression levels were controlled, and the effects of different proteins on the protective actions of IFN-alpha against encephalomyocarditis virus were investigated. RESULTS: NS5A expression blocked the antiviral effects of IFN-alpha in human cells. This inhibition was dependent on the level of NS5A expression. Although ISDR changes gave only small differences in IFN-alpha inhibition, clones derived from a patient who did not respond to IFN-alpha and one who did respond to treatment differed greatly: the clones from a patient with response to IFN-alpha were much more inhibitory than those derived from the patient with no response. CONCLUSIONS: The inhibition of the antiviral effects of IFN-alpha by NS5A is not regulated exclusively by the ISDR, and the effects of NS5A in vitro do not correlate with treatment outcomes. PMID- 10535884 TI - Rapamycin inhibits hepatic stellate cell proliferation in vitro and limits fibrogenesis in an in vivo model of liver fibrosis. AB - BACKGROUND & AIMS: The accelerated course of hepatic fibrosis that occurs in some patients after liver transplantation is a major clinical problem. This response may be caused by the antirejection therapeutics, and in an earlier report we showed that FK-506 enhanced the fibrogenic process in in vivo and in vitro models of liver fibrosis. In the present study, the aim was to determine whether a new immunosuppressive agent, rapamycin, enhances or inhibits liver fibrosis. METHODS: Effects of rapamycin were investigated in a carbon tetrachloride model of hepatic fibrosis in rats and on hepatic stellate proliferation in vitro. RESULTS: Rapamycin inhibited extracellular matrix deposition in the rat model of fibrogenesis as determined by histological analysis, collagen content, messenger RNA levels of procollagen and transforming growth factor beta1, and tissue transglutaminase activity. Moreover, rapamycin decreased platelet growth factor induced proliferation of hepatic stellate cells. CONCLUSIONS: These findings indicate that the new antirejection agent rapamycin inhibits hepatic fibrosis and thus may become a valuable addition to the immunosuppression armamentarium. PMID- 10535885 TI - Rat liver myofibroblasts and hepatic stellate cells: different cell populations of the fibroblast lineage with fibrogenic potential. AB - BACKGROUND & AIMS: Hepatic stellate cells (HSCs) are considered the principal matrix-producing cells of the damaged liver. However, other cell types of the fibroblast lineage that have not yet been characterized are also involved in liver tissue repair and fibrogenesis. METHODS: We established cultures of cells of the fibroblast lineage, termed rat liver myofibroblasts, and analyzed their phenotypical and functional properties in comparison with HSCs. RESULTS: HSCs and rat liver myofibroblasts were discernible by morphological criteria and growth behavior. Prolonged subcultivation of rat liver myofibroblasts was achieved, but HSCs were maintained in culture at maximum until second passage. HSCs were characterized by expression of glial fibrillary acidic protein, desmin, and vascular cell adhesion molecule 1, which were almost completely absent in rat liver myofibroblasts. For synthetic properties, HSCs and rat liver myofibroblasts displayed mostly overlapping properties with 4 striking differences. The complement-activating protease P100 and the protease inhibitor alpha(2) macroglobulin were preferentially expressed by HSCs, whereas interleukin 6-coding messenger RNAs and the extracellular matrix protein fibulin 2 were almost exclusively detectable in rat liver myofibroblasts. CONCLUSIONS: The data show that morphologically and functionally different fibroblastic populations, HSCs and rat liver myofibroblasts, can be derived from liver tissue. HSCs may not represent the single matrix-producing cell type of the fibroblast lineage in the liver. PMID- 10535886 TI - Impaired endothelial nitric oxide synthase activity associated with enhanced caveolin binding in experimental cirrhosis in the rat. AB - BACKGROUND & AIMS: A reduction in nitric oxide (NO) has been implicated as a cause of intrahepatic vasoconstriction in cirrhosis, but the regulatory mechanisms remain undefined. The aim of this study was to examine a contributory role for caveolin-1, a putative negative regulator of endothelial NO synthase, in mediating deficient intrahepatic NO production in the intact cirrhotic liver. METHODS: Cirrhosis was induced by carbon tetrachloride inhalation. Flow regulation of NO production and perfusion pressure was examined in the perfused rat liver. Protein expression of endothelial NO synthase (eNOS), caveolin, and calmodulin was examined by Western blotting and immunohistochemistry. NOS activity and NO production were assessed by citrulline generation and chemiluminescence, respectively. Protein-protein interactions were examined using whole tissue protein immunoprecipitation. RESULTS: In response to incremental increases in flow, cirrhotic animals produced significantly less NO(x) than control animals. NOS activity was significantly reduced in liver tissue from cirrhotic animals compared with control animals in the presence of similar eNOS protein levels. Deficient eNOS activity was associated with a severalfold increase in binding of eNOS with caveolin. Protein levels of caveolin-1 were markedly increased in the cirrhotic liver. CONCLUSIONS: These studies provide evidence that enhanced expression and interaction of caveolin with eNOS contribute to impaired NO production, reduced NOS activity, and vasoconstriction in the intact cirrhotic liver. PMID- 10535887 TI - Resolution of chronic delta hepatitis after 12 years of interferon alfa therapy. AB - Chronic delta hepatitis is an uncommon but severe form of chronic viral hepatitis for which there is currently no satisfactory therapy. A patient with chronic delta hepatitis was treated with interferon alfa, 5 million units daily for 12 years. Serial serum samples were tested for routine liver tests and selected samples for quantitative levels of hepatitis B surface antigen (HBsAg) and hepatitis delta virus RNA. Liver biopsies were performed before, during, and after an initial 1-year course of therapy and again after 3 and 10 years of continuous therapy. With initiation of interferon therapy, serum aminotransferase levels decreased to normal range, became abnormal again when the dose was reduced, and increased to pretreatment levels when therapy was stopped. With reinstitution and prolonged therapy, aminotransferase levels became persistently normal; after several years, both hepatitis delta virus RNA and serum HBsAg became undetectable. Liver biopsy, which initially revealed cirrhosis, showed gradual improvement in inflammatory and fibrosis scores and, after 10 years, no abnormalities or fibrosis. Therapy was stopped, and the patient remained free of evidence of infection. In conclusion, long-term therapy with interferon alfa in high doses led to resolution of chronic delta hepatitis, disappearance of hepatitis delta and B virus markers, and improvement in fibrosis. PMID- 10535888 TI - Acute hepatitis induced by greater celandine (Chelidonium majus). AB - The hepatotoxic potential of conventional drugs is well known, but herbal medicines are often assumed to be harmless. In the last 2 years, we have observed 10 cases of acute hepatitis induced by preparations of greater celandine (Chelidonium majus), which are frequently prescribed to treat gastric and biliary disorders. The course of hepatitis was mild to severe. Marked cholestasis was observed in 5 patients, but liver failure did not occur. Other possible causes of liver disease (viral, autoimmune, hereditary, alcohol, and secondary biliary) were excluded by laboratory tests and imaging procedures, and liver biopsy specimens were consistent with drug-induced damage. After discontinuation of greater celandine, rapid recovery was observed in all patients and liver enzyme levels returned to normal in 2-6 months. Unintentional rechallenge led to a second flare of hepatic inflammation in 1 patient. Greater celandine has to be added to the list of herbs capable of inducing acute (cholestatic) hepatitis. A significant proportion of unexplained cases of hepatitis may be caused by greater celandine. PMID- 10535889 TI - From immune activation to gut tissue injury: the pieces of the puzzle are coming together. PMID- 10535890 TI - More clues to the relationship between hepatic iron and steatosis: An association with insulin resistance? PMID- 10535892 TI - Infliximab for the treatment of fistulas in patients with Crohn'S disease. PMID- 10535891 TI - The virtuosity of hepatic stellate cells. PMID- 10535893 TI - Garlic breath explained: why brushing your teeth won't help. PMID- 10535894 TI - Herbal remedies for the liver: myths, proofs, and treatments. PMID- 10535895 TI - Use of hepatitis C-infected donors for hepatitis C-positive OLT recipients. PMID- 10535896 TI - Long terminal repeat retrotransposons jump between species. PMID- 10535897 TI - Calcium from internal stores modifies dendritic spine shape. PMID- 10535898 TI - Cancer prevention and diet: help from single nucleotide polymorphisms. PMID- 10535899 TI - Making sense of polymer-based biosensors. PMID- 10535900 TI - Delineating muscarinic receptor functions. PMID- 10535901 TI - Bridging the gap: a family of novel DNA polymerases that replicate faulty DNA. PMID- 10535902 TI - SNARE proteins mediate lipid bilayer fusion. PMID- 10535903 TI - VHL: a very hip ligase. PMID- 10535905 TI - Energetic considerations of ciliary beating and the advantage of metachronal coordination. AB - The internal mechanism of cilia is among the most ancient biological motors on an evolutionary scale. It produces beat patterns that consist of two phases: during the effective stroke, the cilium moves approximately as a straight rod, and during the recovery stroke, it rolls close to the surface in a tangential motion. It is commonly agreed that these two phases are designed for efficient functioning: the effective stroke encounters strong viscous resistance and generates thrust, whereas the recovery stroke returns the cilium to starting position while avoiding viscous resistance. Metachronal coordination between cilia, which occurs when many of them beat close to each other, is believed to be an autonomous result of the hydrodynamical interactions in the system. Qualitatively, metachronism is perceived as a way for reducing the energy expenditure required for beating. This paper presents a quantitative study of the energy expenditure of beating cilia, and of the energetic significance of metachronism. We develop a method for computing the work done by model cilia that beat in a viscous fluid. We demonstrate that for a single cilium, beating in water, the mechanical work done during the effective stroke is approximately five times the amount of work done during the recovery stroke. Investigation of multicilia configurations shows that having neighboring cilia beat metachronally is energetically advantageous and perhaps even crucial for multiciliary functioning. Finally, the model is used to approximate the number of dynein arm attachments that are likely to occur during the effective and recovery strokes of a beat cycle, predicting that almost all of the available dynein arms should participate in generating the motion. PMID- 10535906 TI - Reptilian chemistry: characterization of dianeackerone, a secretory product from a crocodile. AB - The major volatile component in the paracloacal glandular secretion of the adult African dwarf crocodile (Osteolaemus tetraspis) was isolated and characterized as a 19-carbon aromatic ketone, dianeackerone (3,7-diethyl-9-phenyl-2-nonanone). This ketone is absent from the secretion of immatures. Careful examination of dianeackerone samples isolated from individual adults revealed that this ketone occurs as both the (3S, 7S) and (3S, 7R) stereoisomers, with different individuals presenting strikingly different ratios of the isomeric forms. Our initial suspicion that the stereoisomeric dianeackerones might be indicators of gender proved untenable, leaving the role of these glandular constituents a challenge for future study. PMID- 10535904 TI - The metabolic implications of intracellular circulation. AB - Two views currently dominate research into cell function and regulation. Model I assumes that cell behavior is quite similar to that expected for a watery bag of enzymes and ligands. Model II assumes that three-dimensional order and structure constrain and determine metabolite behavior. A major problem in cell metabolism is determining why essentially all metabolite concentrations are remarkably stable (are homeostatic) over large changes in pathway fluxes-for convenience, this is termed the [s] stability paradox. For muscle cells, ATP and O(2) are the most perfectly homeostatic, even though O(2) delivery and metabolic rate correlate in a 1:1 fashion. In total, more than 60 metabolites are known to be remarkably homeostatic in differing metabolic states. Several explanations of [s] stability are usually given by traditional model I studies-none of which apply to all enzymes in a pathway, and all of which require diffusion as the means for changing enzyme-substrate encounter rates. In contrast, recent developments in our understanding of intracellular myosin, kinesin, and dyenin motors running on actin and tubulin tracks or cables supply a mechanistic basis for regulated intracellular circulation systems with cytoplasmic streaming rates varying over an approximately 80-fold range (from 1 to >80 micrometer x sec(-1)). These new studies raise a model II hypothesis of intracellular perfusion or convection as a primary means for bringing enzymes and substrates together under variable metabolic conditions. In this view, change in intracellular perfusion rates cause change in enzyme-substrate encounter rates and thus change in pathway fluxes with no requirement for large simultaneous changes in substrate concentrations. The ease with which this hypothesis explains the [s] stability paradox is one of its most compelling features. PMID- 10535907 TI - Reptilian chemistry: characterization of a family of dianeackerone-related steroidal esters from a crocodile secretion. AB - The African dwarf crocodile, Osteolaemus tetraspis (Crocodilidae, Reptilia), possesses a pair of skin glands, the paracloacal glands, the secretion of which is thought to be used to mark nest sites or attract mates. Ten aromatic steroidal esters were isolated from this secretion and characterized on the basis of NMR spectroscopic investigations, electrospray ionization-MS analyses, and chemical degradation. These esters, which account for more than 90% of the paracloacal glandular secretion, are derived from either cholesterol or cholestanol, esterified with a C-20 or C-22 acid closely related to dianeackerone, the only significant volatile compound found in this secretion. PMID- 10535909 TI - Solution of the pulse width modulation problem using orthogonal polynomials and Korteweg-de Vries equations. AB - The mathematical underpinning of the pulse width modulation (PWM) technique lies in the attempt to represent "accurately" harmonic waveforms using only square forms of a fixed height. The accuracy can be measured using many norms, but the quality of the approximation of the analog signal (a harmonic form) by a digital one (simple pulses of a fixed high voltage level) requires the elimination of high order harmonics in the error term. The most important practical problem is in "accurate" reproduction of sine-wave using the same number of pulses as the number of high harmonics eliminated. We describe in this paper a complete solution of the PWM problem using Pade approximations, orthogonal polynomials, and solitons. The main result of the paper is the characterization of discrete pulses answering the general PWM problem in terms of the manifold of all rational solutions to Korteweg-de Vries equations. PMID- 10535908 TI - Design, synthesis, and biological characterization of a peptide-mimetic antagonist for a tethered-ligand receptor. AB - Protease-activated receptors (PARs) represent a unique family of seven transmembrane G protein-coupled receptors, which are enzymatically cleaved to expose a truncated extracellular N terminus that acts as a tethered activating ligand. PAR-1 is cleaved and activated by the serine protease alpha-thrombin, is expressed in various tissues (e.g., platelets and vascular cells), and is involved in cellular responses associated with hemostasis, proliferation, and tissue injury. We have discovered a series of potent peptide-mimetic antagonists of PAR-1, exemplified by RWJ-56110. Spatial relationships between important functional groups of the PAR-1 agonist peptide epitope SFLLRN were employed to design and synthesize candidate ligands with appropriate groups attached to a rigid molecular scaffold. Prototype RWJ-53052 was identified and optimized via solid-phase parallel synthesis of chemical libraries. RWJ-56110 emerged as a potent, selective PAR-1 antagonist, devoid of PAR-1 agonist and thrombin inhibitory activity. It binds to PAR-1, interferes with PAR-1 calcium mobilization and cellular function (platelet aggregation; cell proliferation), and has no effect on PAR-2, PAR-3, or PAR-4. By flow cytometry, RWJ-56110 was confirmed as a direct inhibitor of PAR-1 activation and internalization, without affecting N-terminal cleavage. At high concentrations of alpha-thrombin, RWJ 56110 fully blocked activation responses in human vascular cells, albeit not in human platelets; whereas, at high concentrations of SFLLRN-NH(2), RWJ-56110 blocked activation responses in both cell types. Thus, thrombin activates human platelets independently of PAR-1, i.e., through PAR-4, which we confirmed by PCR analysis. Selective PAR-1 antagonists, such as RWJ-56110, should serve as useful tools to study PARs and may have therapeutic potential for treating thrombosis and restenosis. PMID- 10535910 TI - Pressure-enhanced ortho-para conversion in solid hydrogen up to 58 GPa. AB - We measured the ortho-para conversion rate in solid hydrogen by using Raman scattering in a diamond-anvil cell, extending previous measurements by a factor of 60 in pressure. We confirm previous experiments that suggested a decrease in the conversion rate above about 0.5 GPa. We observe a distinct minimum at 3 GPa followed by a drastic increase in the conversion rate to our maximum pressure of 58 GPa. This pressure enhancement of conversion is not predicted by previous theoretical treatments and must be due to a new conversion pathway. PMID- 10535911 TI - Derived variables for longitudinal studies. AB - Suppose that for each individual a vector of features is measured at a number of time points. We look for a transformation of the features, the same at all time points, that will induce a simple dependency structure. In the simplest situation this requires that a certain asymmetric matrix has real nonzero eigenvalues. Extensions are considered. PMID- 10535912 TI - Identification of lumichrome as a sinorhizobium enhancer of alfalfa root respiration and shoot growth. AB - Sinorhizobium meliloti bacteria produce a signal molecule that enhances root respiration in alfalfa (Medicago sativa L.) and also triggers a compensatory increase in whole-plant net carbon assimilation. Nuclear magnetic resonance, mass spectrometry, and ultraviolet-visible absorption identify the enhancer as lumichrome, a common breakdown product of riboflavin. Treating alfalfa roots with 3 nM lumichrome increased root respiration 21% (P < 0.05) within 48 h. A closely linked increase in net carbon assimilation by the shoot compensated for the enhanced root respiration. For example, applying 5 nM lumichrome to young alfalfa roots increased plant growth by 8% (P < 0.05) after 12 days. Soaking alfalfa seeds in 5 nM lumichrome before germination increased growth by 18% (P < 0.01) over the same period. In both cases, significant growth enhancement (P < 0.05) was evident only in the shoot. S. meliloti requires exogenous CO2 for growth and may benefit directly from the enhanced root respiration that is triggered by lumichrome. Thus Sinorhizobium-alfalfa associations, which ultimately form symbiotic N2-reducing root nodules, may be favored at an early developmental stage by lumichrome, a previously unrecognized mutualistic signal. The rapid degradation of riboflavin to lumichrome under many physiological conditions and the prevalence of riboflavin release by rhizosphere bacteria suggest that events demonstrated here in the S. meliloti-alfalfa association may be widely important across many plant-microbe interactions. PMID- 10535913 TI - Direct radiocarbon dates for Vindija G(1) and Velika Pecina late Pleistocene hominid remains. AB - New accelerator mass spectrometry radiocarbon dates taken directly on human remains from the Late Pleistocene sites of Vindija and Velika Pecina in the Hrvatsko Zagorje of Croatia are presented. Hominid specimens from both sites have played critical roles in the development of current perspectives on modern human evolutionary emergence in Europe. Dates of approximately 28 thousand years (ka) before the present (B.P.) and approximately 29 ka B.P. for two specimens from Vindija G(1) establish them as the most recent dated Neandertals in the Eurasian range of these archaic humans. The human frontal bone from Velika Pecina, generally considered one of the earliest representatives of modern humans in Europe, dated to approximately 5 ka B.P., rendering it no longer pertinent to discussions of modern human origins. Apart from invalidating the only radiometrically based example of temporal overlap between late Neandertal and early modern human fossil remains from within any region of Europe, these dates raise the question of when early modern humans first dispersed into Europe and have implications for the nature and geographic patterning of biological and cultural interactions between these populations and the Neandertals. PMID- 10535914 TI - Highly sensitive biological and chemical sensors based on reversible fluorescence quenching in a conjugated polymer. AB - The fluorescence of a polyanionic conjugated polymer can be quenched by extremely low concentrations of cationic electron acceptors in aqueous solutions. We report a greater than million-fold amplification of the sensitivity to fluorescence quenching compared with corresponding "molecular excited states." Using a combination of steady-state and ultrafast spectroscopy, we have established that the dramatic quenching results from weak complex formation [polymer( )/quencher(+)], followed by ultrafast electron transfer from excitations on the entire polymer chain to the quencher, with a time constant of 650 fs. Because of the weak complex formation, the quenching can be selectively reversed by using a quencher-recognition diad. We have constructed such a diad and demonstrate that the fluorescence is fully recovered on binding between the recognition site and a specific analyte protein. In both solutions and thin films, this reversible fluorescence quenching provides the basis for a new class of highly sensitive biological and chemical sensors. PMID- 10535915 TI - DNA packing in stable lipid complexes designed for gene transfer imitates DNA compaction in bacteriophage. AB - The structure of complexes made from DNA and suitable lipids (lipoplex, Lx) was examined by cryo-electron microscopy (cryoEM). We observed a distinct concentric ring-like pattern with striated shells when using plasmid DNA. These spherical multilamellar particles have a mean diameter of 254 nm with repetitive spacing of 7.5 nm with striation of 5.3 nm width. Small angle x-ray scattering revealed repetitive ordering of 6.9 nm, suggesting a lamellar structure containing at least 12 layers. This concentric and lamellar structure with different packing regimes also was observed by cryoEM when using linear double-stranded DNA, single stranded DNA, and oligodeoxynucleotides. DNA chains could be visualized in DNA/lipid complexes. Such specific supramolecular organization is the result of thermodynamic forces, which cause compaction to occur through concentric winding of DNA in a liquid crystalline phase. CryoEM examination of T4 phage DNA packed either in T4 capsides or in lipidic particles showed similar patterns. Small angle x-ray scattering suggested an hexagonal phase in Lx-T4 DNA. Our results indicate that both lamellar and hexagonal phases may coexist in the same Lx preparation or particle and that transition between both phases may depend on equilibrium influenced by type and length of the DNA used. PMID- 10535916 TI - Three metal ions at the active site of the Tetrahymena group I ribozyme. AB - Metal ions are critical for catalysis by many RNA and protein enzymes. To understand how these enzymes use metal ions for catalysis, it is crucial to determine how many metal ions are positioned at the active site. We report here an approach, combining atomic mutagenesis with quantitative determination of metal ion affinities, that allows individual metal ions to be distinguished. Using this approach, we show that at the active site of the Tetrahymena group I ribozyme the previously identified metal ion interactions with three substrate atoms, the 3'-oxygen of the oligonucleotide substrate and the 3'- and 2'-moieties of the guanosine nucleophile, are mediated by three distinct metal ions. This approach provides a general tool for distinguishing active site metal ions and allows the properties and roles of individual metal ions to be probed, even within the sea of metal ions bound to RNA. PMID- 10535917 TI - A structural view of evolutionary divergence. AB - Two directed evolution experiments on p-nitrobenzyl esterase yielded one enzyme with a 100-fold increased activity in aqueous-organic solvents and another with a 17 degrees C increase in thermostability. Structures of the wild type and its organophilic and thermophilic counterparts are presented at resolutions of 1.5 A, 1.6 A, and 2.0 A, respectively. These structures identify groups of interacting mutations and demonstrate how directed evolution can traverse complex fitness landscapes. Early-generation mutations stabilize flexible loops not visible in the wild-type structure and set the stage for further beneficial mutations in later generations. The mutations exert their influence on the esterase structure over large distances, in a manner that would be difficult to predict. The loops with the largest structural changes generally are not the sites of mutations. Similarly, none of the seven amino acid substitutions in the organophile are in the active site, even though the enzyme experiences significant changes in the organization of this site. In addition to reduction of surface loop flexibility, thermostability in the evolved esterase results from altered core packing, helix stabilization, and the acquisition of surface salt bridges, in agreement with other comparative studies of mesophilic and thermophilic enzymes. Crystallographic analysis of the wild type and its evolved counterparts reveals networks of mutations that collectively reorganize the active site. Interestingly, the changes that led to diversity within the alpha/beta hydrolase enzyme family and the reorganization seen in this study result from main-chain movements. PMID- 10535918 TI - Targeting a SWI/SNF-related chromatin remodeling complex to the beta-globin promoter in erythroid cells. AB - Chromatin remodeling complexes such as the SWI/SNF complex make DNA accessible to transcription factors by disrupting nucleosomes. However, it is not known how such complexes are targeted to the promoter. For example, a SWI/SNF1-like chromatin remodeling complex erythroid Kruppel-like factor (EKLF) coactivator remodeling complex 1 (E-RC1) disrupts the nucleosomes over the human beta-globin promoter in an EKLF-dependent manner. However, it is not known whether E-RC1 is targeted specifically to the beta-globin promoter or whether E-RC1 is randomly targeted, but its activity is evident only at the beta-globin promoter. Because E RC1 cannot remodel chromatin over the beta-globin promoter without EKLF in vitro, it has been proposed that SWI/SNF1-like complexes such as E-RC1 are targeted specifically to the promoter by selectively interacting with promoter-associated transcription factors such as EKLF. In this report, we test this hypothesis in the cellular context by using the ProteIN POsition Identification with Nuclease Tail (PIN*POINT) assay. We find that the Brahma-related gene (BRG) 1 and BRG1 associated factor (BAF) 170 subunits of E-RC1 are both recruited near the transcription initiation site of the beta-globin promoter. On transiently transfected templates, both the locus control region and the EKLF-binding site are important for their recruitment to the beta-globin promoter in mouse erythroleukemia cells. When the beta-globin promoter was linked to the cytomegalovirus enhancer, the E-RC1 complex was not recruited, suggesting that recruitment of the E-RC1 complex is not a general property of enhancers. PMID- 10535919 TI - Assembly of a catalytic unit for RNA microhelix aminoacylation using nonspecific RNA binding domains. AB - An assembly of a catalytic unit for aminoacylation of an RNA microhelix is demonstrated here. This assembly may recapitulate a step in the historical development of tRNA synthetases. The class-defining domain of a tRNA synthetase is closely related to the primordial enzyme that catalyzed synthesis of aminoacyl adenylate. RNA binding elements are imagined to have been added so that early RNA substrates could be docked proximal to the activated amino acid. RNA microhelices that recapitulate the acceptor stem of modern tRNAs are potential examples of early substrates. In this work, we examined a fragment of Escherichia coli alanyl tRNA synthetase, which catalyzes aminoacyl adenylate formation but is virtually inactive for catalysis of RNA microhelix aminoacylation. Fusion to the fragment of either of two unrelated nonspecific RNA binding domains activated microhelix aminoacylation. Although the fusion proteins lacked the RNA sequence specificity of the natural enzyme, their activity was within 1-2 kcal.mol(-1) of a truncated alanyl-tRNA synthetase that has aminoacylation activity sufficient to sustain cell growth. These results show that, starting with an activity for adenylate synthesis, barriers are relatively low for building catalytic units for aminoacylation of RNA helices. PMID- 10535920 TI - Conversion of agonist site to metal-ion chelator site in the beta(2)-adrenergic receptor. AB - Previously metal-ion sites have been used as structural and functional probes in seven transmembrane receptors (7TM), but as yet all the engineered sites have been inactivating. Based on presumed agonist interaction points in transmembrane III (TM-III) and -VII of the beta(2)-adrenergic receptor, in this paper we construct an activating metal-ion site between the amine-binding Asp-113 in TM III-or a His residue introduced at this position-and a Cys residue substituted for Asn-312 in TM-VII. No increase in constitutive activity was observed in the mutant receptors. Signal transduction was activated in the mutant receptors not by normal catecholamine ligands but instead either by free zinc ions or by zinc or copper ions in complex with small hydrophobic metal-ion chelators. Chelation of the metal ions by small hydrophobic chelators such as phenanthroline or bipyridine protected the cells from the toxic effect of, for example Cu(2+), and in several cases increased the affinity of the ions for the agonistic site. Wash out experiments and structure-activity analysis indicated, that the high-affinity chelators and the metal ions bind and activate the mutant receptor as metal ion guided ligand complexes. Because of the well-understood binding geometry of the small metal ions, an important distance constraint has here been imposed between TM-III and -VII in the active, signaling conformation of 7TM receptors. It is suggested that atoxic metal-ion chelator complexes could possibly in the future be used as generic, pharmacologic tools to switch 7TM receptors with engineered metal-ion sites on or off at will. PMID- 10535921 TI - NMR characterization of lignins in Arabidopsis altered in the activity of ferulate 5-hydroxylase. AB - Nuclear magnetic resonance (NMR) of isolated lignins from an Arabidopsis mutant deficient in ferulate 5-hydroxylase (F5H) and transgenic plants derived from the mutant by overexpressing the F5H gene has provided detailed insight into the compositional and structural differences between these lignins. Wild-type Arabidopsis has a guaiacyl-rich, syringyl-guaiacyl lignin typical of other dicots, with prominent beta-aryl ether (beta-O-4), phenylcoumaran (beta-5), resinol (beta-beta), biphenyl/dibenzodioxocin (5-5), and cinnamyl alcohol end group structures. The lignin isolated from the F5H-deficient fah1-2 mutant contained only traces of syringyl units and consequently enhanced phenylcoumaran and dibenzodioxocin levels. In fah1-2 transgenics in which the F5H gene was overexpressed under the control of the cauliflower mosaic virus 35S promoter, a guaiacyl-rich, syringyl/guaiacyl lignin similar to the wild type was produced. In contrast, the isolated lignin from the fah1-2 transgenics in which F5H expression was driven by the cinnamate 4-hydroxylase promoter was almost entirely syringyl in nature. This simple lignin contained predominantly beta-aryl ether units, mainly with erythro-stereochemistry, with some resinol structures. No phenylcoumaran or dibenzodioxocin structures (which require guaiacyl units) were detectable. The overexpression of syringyl units in this transgenic resulted in a lignin with a higher syringyl content than that in any other plant we have seen reported. PMID- 10535922 TI - Crystal structure of a multifunctional 2-Cys peroxiredoxin heme-binding protein 23 kDa/proliferation-associated gene product. AB - Heme-binding protein 23 kDa (HBP23), a rat isoform of human proliferation associated gene product (PAG), is a member of the peroxiredoxin family of peroxidases, having two conserved cysteine residues. Recent biochemical studies have shown that HBP23/PAG is an oxidative stress-induced and proliferation coupled multifunctional protein that exhibits specific bindings to c-Abl protein tyrosine kinase and heme, as well as a peroxidase activity. A 2.6-A resolution crystal structure of rat HBP23 in oxidized form revealed an unusual dimer structure in which the active residue Cys-52 forms a disulfide bond with conserved Cys-173 from another subunit by C-terminal tail swapping. The active site is largely hydrophobic with partially exposed Cys-173, suggesting a reduction mechanism of oxidized HBP23 by thioredoxin. Thus, the unusual cysteine disulfide bond is involved in peroxidation catalysis by using thioredoxin as the source of reducing equivalents. The structure also provides a clue to possible interaction surfaces for c-Abl and heme. Several significant structural differences have been found from a 1-Cys peroxiredoxin, ORF6, which lacks the C terminal conserved cysteine corresponding to Cys-173 of HBP23. PMID- 10535923 TI - Purification of ribonucleotide reductase subunits Y1, Y2, Y3, and Y4 from yeast: Y4 plays a key role in diiron cluster assembly. AB - Ribonucleotide reductases (RNRs) catalyze the conversion of nucleotides to deoxynucleotides. Class I RNRs are composed of two types of subunits: RNR1 contains the active site for reduction and the binding sites for the nucleotide allosteric effectors. RNR2 contains the diiron-tyrosyl radical (Y.) cofactor essential for the reduction process. Studies in yeast have recently identified four RNR subunits: Y1 and Y3, Y2 and Y4. These proteins have been expressed in Saccharomyces cerevisiae and in Escherichia coli and purified to approximately 90% homogeneity. The specific activity of Y1 isolated from yeast and E. coli is 0.03 micromol.min(-1).mg(-1) and of (His)(6)-Y2 [(His)(6)-Y2-K387N] from yeast is 0.037 micromol. min(-1).mg(-1) (0.125 micromol.min(-1).mg(-1)). Y2, Y3, and Y4 isolated from E. coli have no measurable activity. Efforts to generate Y. in Y2 or Y4 using Fe(2+), O(2), and reductant have been unsuccessful. However, preliminary studies show that incubation of Y4 and Fe(2+) with inactive E. coli Y2 followed by addition of O(2) generates Y2 with a specific activity of 0.069 micromol.min(-1). mg(-1) and a Y. A similar experiment with (His)(6)-Y2-K387N, Y4, O(2), and Fe(2+) results in an increase in its specific activity to 0.30 micromol.min(-1).mg(-1). Studies with antibodies to Y4 and Y2 reveal that they can form a complex in vivo. Y4 appears to play an important role in diiron-Y. assembly of Y2. PMID- 10535924 TI - A protein residing at the subunit interface of the bacterial ribosome. AB - Surface labeling of Escherichia coli ribosomes with the use of the tritium bombardment technique has revealed a minor unidentified ribosome-bound protein (spot Y) that is hidden in the 70S ribosome and becomes highly labeled on dissociation of the 70S ribosome into subunits. In the present work, the N terminal sequence of the protein Y was determined and its gene was identified as yfia, an ORF located upstream the phe operon of E. coli. This 12.7-kDa protein was isolated and characterized. An affinity of the purified protein Y for the 30S subunit, but not for the 50S ribosomal subunit, was shown. The protein proved to be exposed on the surface of the 30S subunit. The attachment of the 50S subunit resulted in hiding the protein Y, thus suggesting the protein location at the subunit interface in the 70S ribosome. The protein was shown to stabilize ribosomes against dissociation. The possible role of the protein Y as ribosome association factor in translation is discussed. PMID- 10535925 TI - Covalent modification of the homeodomain-interacting protein kinase 2 (HIPK2) by the ubiquitin-like protein SUMO-1. AB - Posttranslational modifications such as ubiquitination and phosphorylation play an important role in the regulation of cellular protein function. Homeodomain interacting protein kinase 2 (HIPK2) is a member of the recently identified family of nuclear protein kinases that act as corepressors for homeodomain transcription factors. Here, we show that HIPK2 is regulated by a ubiquitin-like protein, SUMO-1. We demonstrate that HIPK2 localizes to nuclear speckles (dots) by means of a speckle-retention signal. This speckle-retention signal contains a domain that interacts with a mouse ubiquitin-like protein conjugating (E2) enzyme, mUBC9. In cultured cells, HIPK2 is covalently modified by SUMO-1, and the SUMO-1 modification of HIPK2 correlates with its localization to nuclear speckles (dots). Thus, our results provide firm evidence that the nuclear protein kinase HIPK2 can be covalently modified by SUMO-1, which directs its localization to nuclear speckles (dots). PMID- 10535926 TI - Yeast HOS3 forms a novel trichostatin A-insensitive homodimer with intrinsic histone deacetylase activity. AB - Histone deacetylases such as human HDAC1 and yeast RPD3 are trichostatin A (TSA) sensitive enzymes that are members of large, multiprotein complexes. These contain specialized subunits that help target the catalytic protein to histones at the appropriate DNA regulatory element, where the enzyme represses transcription. To date, no deacetylase catalytic subunits have been shown to have intrinsic activity, suggesting that noncatalytic subunits of the deacetylase complex are required for their enzymatic function. In this paper we describe a novel yeast histone deacetylase HOS3 that is relatively insensitive to the histone deacetylase inhibitor TSA, forms a homodimer when expressed ectopically both in yeast and Escherichia coli, and has intrinsic activity when produced in the bacterium. Most HOS3 protein can be found associated with a larger complex in partially purified yeast nuclear extracts, arguing that the HOS3 homodimer may be dissociated from a very large nuclear structure during purification. We also demonstrate, using a combination of mass spectrometry, tandem mass spectrometry, and proteolytic digestion, that recombinant HOS3 has a distinct specificity in vitro for histone H4 sites K5 and K8, H3 sites K14 and K23, H2A site K7, and H2B site K11. We propose that while factors that interact with HOS3 may sequester the catalytic subunit at specific cellular sites, they are not required for HOS3 histone deacetylase activity. PMID- 10535927 TI - The protein kinase CK2 is involved in regulation of circadian rhythms in Arabidopsis. AB - A wide range of processes in plants, including expression of certain genes, is regulated by endogenous circadian rhythms. The circadian clock-associated 1 (CCA1) and the late elongated hypocotyl (LHY) proteins have been shown to be closely associated with clock function in Arabidopsis thaliana. The protein kinase CK2 can interact with and phosphorylate CCA1, but its role in the regulation of the circadian clock remains unknown. Here we show that plants overexpressing CKB3, a regulatory subunit of CK2, display increased CK2 activity and shorter periods of rhythmic expression of CCA1 and LHY. CK2 is also able to interact with and phosphorylate LHY in vitro. Additionally, overexpression of CKB3 shortened the periods of four known circadian clock-controlled genes with different phase angles, demonstrating that many clock outputs are affected. This overexpression also reduced phytochrome induction of an Lhcb gene. Finally, we found that the photoperiodic flowering response, which is influenced by circadian rhythms, was diminished in the transgenic lines, and that the plants flowered earlier on both long-day and short-day photoperiods. These data demonstrate that CK2 is involved in regulation of the circadian clock in Arabidopsis. PMID- 10535928 TI - Identification and reconstitution of the origin recognition complex from Schizosaccharomyces pombe. AB - The origin recognition complex (ORC), first identified in Saccharomyces cerevisiae (sc), is a six-subunit protein complex that binds to DNA origins. Here, we report the identification and cloning of cDNAs encoding the six subunits of the ORC of Schizosaccharomyces pombe (sp). Sequence analyses revealed that spOrc1, 2, and 5 subunits are highly conserved compared with their counterparts from S. cerevisiae, Xenopus, Drosophila, and human. In contrast, both spOrc3 and spOrc6 subunits are poorly conserved. As reported by Chuang and Kelly [(1999) Proc. Natl. Acad. Sci. USA 96, 2656-2661], the C-terminal region of spOrc4 is also conserved whereas the N terminus uniquely contains repeats of a sequence that binds strongly to AT-rich DNA regions. Consistent with this, extraction of S. pombe chromatin with 1 M NaCl, or after DNase I treatment, yielded the six subunit ORC, whereas extraction with 0.3 M resulted in five-subunit ORC lacking spOrc4p. The spORC can be reconstituted in vitro with all six recombinant subunits expressed in the rabbit reticulocyte system. The association of spOrc4p with the other subunits required the removal of DNA from reaction mixture by DNase I. This suggests that a strong interaction between spOrc4p and DNA can prevent the isolation of the six-subunit ORC. The unique DNA-binding properties of the spORC may contribute to our understanding of the sequence-specific recognition required for the initiation of DNA replication in S. pombe. PMID- 10535929 TI - The Escherichia coli SOS mutagenesis proteins UmuD and UmuD' interact physically with the replicative DNA polymerase. AB - The Escherichia coli umuDC operon is induced in response to replication-blocking DNA lesions as part of the SOS response. UmuD protein then undergoes an RecA facilitated self-cleavage reaction that removes its N-terminal 24 residues to yield UmuD'. UmuD', UmuC, RecA, and some form of the E. coli replicative DNA polymerase, DNA polymerase III holoenzyme, function in translesion synthesis, the potentially mutagenic process of replication over otherwise blocking lesions. Furthermore, it has been proposed that, before cleavage, UmuD together with UmuC acts as a DNA damage checkpoint system that regulates the rate of DNA synthesis in response to DNA damage, thereby allowing time for accurate repair to take place. Here we provide direct evidence that both uncleaved UmuD and UmuD' interact physically with the catalytic, proofreading, and processivity subunits of the E. coli replicative polymerase. Consistent with our model proposing that uncleaved UmuD and UmuD' promote different events, UmuD and UmuD' interact differently with DNA polymerase III: whereas uncleaved UmuD interacts more strongly with beta than it does with alpha, UmuD' interacts more strongly with alpha than it does with beta. We propose that the protein-protein interactions we have characterized are part of a higher-order regulatory system of replication fork management that controls when the umuDC gene products can gain access to the replication fork. PMID- 10535930 TI - Structure of a soluble secreted chemokine inhibitor vCCI (p35) from cowpox virus. AB - Most poxviruses, including variola, the causative agent of smallpox, express a secreted protein of 35 kDa, vCCI, which binds CC-chemokines with high affinity. This viral protein competes with the host cellular CC-chemokine receptors (CCRs), reducing inflammation and interfering with the host immune response. Such proteins or derivatives may have therapeutic uses as anti-inflammatory agents. We have determined the crystal structure to 1.85-A resolution of vCCI from cowpox virus, the prototype of this poxvirus virulence factor. The molecule is a beta sandwich of topology not previously described. A patch of conserved residues on the exposed face of a beta-sheet that is strongly negatively charged might have a role in binding of CC-chemokines, which are positively charged. PMID- 10535931 TI - hMutSalpha- and hMutLalpha-dependent phosphorylation of p53 in response to DNA methylator damage. AB - hMSH2.hMSH6 heterodimer (hMutSalpha) and hMLH1.hPMS2 complex (hMutLalpha) have been implicated in the cytotoxic response of mammalian cells to a number of DNA damaging compounds, including methylating agents that produce O(6)-methylguanine (O(6)MeG) adducts. This study demonstrates that O(6)MeG lesions, in which the damaged base is paired with either T or C, are subject to excision repair in a reaction that depends on a functional mismatch repair system. Furthermore, treatment of human cells with the S(N)1 DNA methylators N-methyl-N-nitrosourea or N-methyl-N'-nitro-N-nitrosoguanidine results in p53 phosphorylation on serine residues 15 and 392, and these phosphorylation events depend on the presence of functional hMutSalpha and hMutLalpha. Coupled with the previous demonstration that O(6)MeG.T and O(6)MeG.C pairs are recognized by hMutSalpha, these results implicate action of the mismatch repair system in the initial step of a damage signaling cascade that can lead to cell-cycle checkpoint activation or cell death in response to DNA methylator damage. PMID- 10535933 TI - Mobilization of the A-kinase N-myristate through an isoform-specific intermolecular switch. AB - Although the catalytic (C) subunit of cAMP-dependent protein kinase is N myristylated, it is a soluble protein, and no physiological role has been identified for its myristyl moiety. To determine whether the interaction of the two regulatory (R) subunit isoforms (R(I) and R(II)) with the N-myristylated C subunit affects its ability to target membranes, the effect of N-myristylation and the R(I) and R(II) subunit isoforms on C subunit binding to phosphatidylcholine/phosphatidylserine liposomes was examined. Only the combination of N-myristylation and R(II) subunit interaction produced a dramatic increase in the rate of liposomal binding. To assess whether the R(II) subunit also increased the conformational flexibility of the C subunit N terminus, the effect of N-myristylation and the R(I) and R(II) subunits on the rotational freedom of the C subunit N terminus was measured. Specifically, fluorescein maleimide was conjugated to Cys-16 in the N-terminal domain of a K16C mutant of the C subunit, and the time-resolved emission anisotropy was determined. The interaction of the R(II) subunit, but not the R(I) subunit, significantly increased the backbone flexibility around the site of mutation and labeling, strongly suggesting that R(II) subunit binding to the myristylated C subunit induced a unique conformation of the C subunit that is associated with an increase in both the N-terminal flexibility and the exposure of the N-myristate. R(II) subunit thus appears to serve as an intermolecular switch that disrupts of the link between the N-terminal and core catalytic domains of the C subunit to expose the N-myristate and poise the holoenzyme for interaction with membranes. PMID- 10535932 TI - In Azotobacter vinelandii, the E1 subunit of the pyruvate dehydrogenase complex binds fpr promoter region DNA and ferredoxin I. AB - In Azotobacter vinelandii, deletion of the fdxA gene that encodes a well characterized seven-iron ferredoxin (FdI) is known to lead to overexpression of the FdI redox partner, NADPH:ferredoxin reductase (FPR). Previous studies have established that this is an oxidative stress response in which the fpr gene is transcriptionally activated to the same extent in response to either addition of the superoxide propagator paraquat to the cells or to fdxA deletion. In both cases, the activation occurs through a specific DNA sequence located upstream of the fpr gene. Here, we report the identification of the A. vinelandii protein that binds specifically to the paraquat activatable fpr promoter region as the E1 subunit of the pyruvate dehydrogenase complex (PDHE1), a central enzyme in aerobic respiration. Sequence analysis shows that PDHE1, which was not previously suspected to be a DNA-binding protein, has a helix-turn-helix motif. The data presented here further show that FdI binds specifically to the DNA-bound PDHE1. PMID- 10535935 TI - RNase E is required for the maturation of ssrA RNA and normal ssrA RNA peptide tagging activity. AB - During recent studies of ribonucleolytic "degradosome" complexes of Escherichia coli, we found that degradosomes contain certain RNAs as well as RNase E and other protein components. One of these RNAs is ssrA (for small stable RNA) RNA (also known as tm RNA or 10Sa RNA), which functions as both a tRNA and mRNA to tag the C-terminal ends of truncated proteins with a short peptide and target them for degradation. Here, we show that mature 363-nt ssrA RNA is generated by RNase E cleavage at the CCA-3' terminus of a 457-nt ssrA RNA precursor and that interference with this cleavage in vivo leads to accumulation of the precursor and blockage of SsrA-mediated proteolysis. These results demonstrate that RNase E is required to produce mature ssrA RNA and for normal ssrA RNA peptide-tagging activity. Our findings indicate that RNase E, an enzyme already known to have a central role in RNA processing and decay in E. coli, also has the previously unsuspected ability to affect protein degradation through its role in maturation of the 3' end of ssrA RNA. PMID- 10535934 TI - Isolation of a Chinese hamster ovary cell mutant defective in intramitochondrial transport of phosphatidylserine. AB - A CHO-K1 cell mutant with a specific decrease in cellular phosphatidylethanolamine (PE) level was isolated as a variant resistant to Ro09 0198, a PE-directed antibiotic peptide. The mutant was defective in the phosphatidylserine (PS) decarboxylation pathway for PE formation, in which PS produced in the endoplasmic reticulum is transported to mitochondria and then decarboxylated by an inner mitochondrial membrane enzyme, PS decarboxylase. Neither PS formation nor PS decarboxylase activity was reduced in the mutant, implying that the mutant is defective in some step of PS transport. The transport processes of phospholipids between the outer and inner mitochondrial membrane were analyzed by use of isolated mitochondria and two fluorescence-labeled phospholipid analogs, 1-palmitoyl-2-[N-[6(7-nitrobenz-2-oxa-1, 3-diazol-4 yl)amino]caproyl]-PS (C6-NBD-PS) and C6-NBD-phosphatidylcholine (C6-NBD-PC). On incubation with the CHO-K1 mitochondria, C6-NBD-PS was readily decarboxylated to C6-NBD-PE, suggesting that the PS analog was partitioned into the outer leaflet of mitochondria and then translocated to the inner mitochondrial membrane. The rate of decarboxylation of C6-NBD-PS in the mutant mitochondria was reduced to approximately 40% of that in the CHO-K1 mitochondria. The quantity of phospholipid analogs translocated from the outer leaflet of mitochondria into inner mitochondrial membranes was further examined by selective extraction of the analogs from the outer leaflet of mitochondria. In the mutant mitochondria, the translocation of C6-NBD-PS was significantly reduced, whereas the translocation of C6-NBD-PC was not affected. These results indicate that the mutant is defective in PS transport between the outer and inner mitochondrial membrane and provide genetic evidence for the existence of a specific mechanism for intramitochondrial transport of PS. PMID- 10535936 TI - Replacement of the proximal heme thiolate ligand in chloroperoxidase with a histidine residue. AB - Chloroperoxidase is a versatile heme enzyme which can cross over the catalytic boundaries of other oxidative hemoproteins and perform multiple functions. Chloroperoxidase, in addition to catalyzing classical peroxidative reactions, also acts as a P450 cytochrome and a potent catalase. The multiple functions of chloroperoxidase must be derived from its unique active site structure. Chloroperoxidase possesses a proximal cysteine thiolate heme iron ligand analogous to the P450 cytochromes; however, unlike the P450 enzymes, chloroperoxidase possesses a very polar environment distal to its heme prosthetic group and contains a glutamic acid residue in close proximity to the heme iron. The presence of a thiolate ligand in chloroperoxidase has long been thought to play an essential role in its chlorination and epoxidation activities; however, the research reported in this paper proves that hypothesis to be invalid. To explore the role of Cys-29, the amino acid residue supplying the thiolate ligand in chloroperoxidase, Cys-29 has been replaced with a histidine residue. Mutant clones of the chloroperoxidase genome have been expressed in a Caldariomyces fumago expression system by using gene replacement rather than gene insertion technology. C. fumago produces wild-type chloroperoxidase, thus requiring gene replacement of the wild type by the mutant gene. To the best of our knowledge, this is the first time that gene replacement has been reported for this type of fungus. The recombinant histidine mutants retain most of their chlorination, peroxidation, epoxidation, and catalase activities. These results downplay the importance of a thiolate ligand in chloroperoxidase and suggest that the distal environment of the heme active site plays the major role in maintaining the diverse activities of this enzyme. PMID- 10535937 TI - Mechanism-based inhibition of the melatonin rhythm enzyme: pharmacologic exploitation of active site functional plasticity. AB - Serotonin N-acetyltransferase is the enzyme responsible for the diurnal rhythm of melatonin production in the pineal gland of animals and humans. Inhibitors of this enzyme active in cell culture have not been reported previously. The compound N-bromoacetyltryptamine was shown to be a potent inhibitor of this enzyme in vitro and in a pineal cell culture assay (IC(50) approximately 500 nM). The mechanism of inhibition is suggested to involve a serotonin N acetyltransferase-catalyzed alkylation reaction between N-bromoacetyltryptamine and reduced CoA, resulting in the production of a tight-binding bisubstrate analog inhibitor. This alkyltransferase activity is apparently catalyzed at a functionally distinct site compared with the acetyltransferase activity active site on serotonin N-acetyltransferase. Such active site plasticity is suggested to result from a subtle conformational alteration in the protein. This plasticity allows for an unusual form of mechanism-based inhibition with multiple turnovers, resulting in "molecular fratricide." N-bromoacetyltryptamine should serve as a useful tool for dissecting the role of melatonin in circadian rhythm as well as a potential lead compound for therapeutic use in mood and sleep disorders. PMID- 10535938 TI - Purification and characterization of sortase, the transpeptidase that cleaves surface proteins of Staphylococcus aureus at the LPXTG motif. AB - Surface proteins of Staphylococcus aureus are linked to the bacterial cell wall by sortase, an enzyme that cleaves polypeptides at the threonine of the LPXTG motif. Surface proteins can be released from staphylococci by treatment with hydroxylamine, resulting in the formation of threonine hydroxamate. Staphylococcal extracts, as well as purified sortase, catalyze the hydroxylaminolysis of peptides bearing an LPXTG motif, a reaction that can be inhibited with sulfhydryl-modifying reagents. Replacement of the single conserved cysteine at position 184 of sortase with alanine abolishes enzyme activity. Thus, sortase appears to catalyze surface-protein anchoring by means of a transpeptidation reaction that captures cleaved polypeptides as thioester enzyme intermediates. PMID- 10535939 TI - Translocation and specific cleavage of bacteriophage T7 DNA in vivo by EcoKI. AB - Infection of Escherichia coli containing the type I restriction enzyme EcoKI by bacteriophage T7 0.3 mutants leads to restriction during the late stages of genome entry and during DNA replication. Patterns of cleavage in vivo suggest that some cutting occurs near the midpoint of two recognition sites, consistent with the idea that EcoKI translocates DNA bidirectionally through itself and cuts when two enzyme molecules collide. Rapid ejection of a 0.3(+) T7 genome from a bacteriophage lambda particle results in degradation of the infecting DNA by EcoKI, showing that the normal T7 DNA translocation process delays restriction. A unique recognition site inserted at the genomic left end allows EcoKI to function as a molecular motor and to translocate the remaining 39 kilobases of T7 DNA into the cell. PMID- 10535940 TI - Identification of the von Hippel-lindau tumor-suppressor protein as part of an active E3 ubiquitin ligase complex. AB - Mutations of von Hippel-Lindau disease (VHL) tumor-suppressor gene product (pVHL) are found in patients with dominant inherited VHL syndrome and in the vast majority of sporadic clear cell renal carcinomas. The function of the pVHL protein has not been clarified. pVHL has been shown to form a complex with elongin B and elongin C (VBC) and with cullin (CUL)-2. In light of the structural analogy of VBC-CUL-2 to SKP1-CUL-1-F-box ubiquitin ligases, the ubiquitin ligase activity of VBC-CUL-2 was examined in this study. We show that VBC-CUL-2 exhibits ubiquitin ligase activity, and we identified UbcH5a, b, and c, but not CDC34, as the ubiquitin-conjugating enzymes of the VBC-CUL-2 ubiquitin ligase. The protein Rbx1/ROC1 enhances ligase activity of VBC-CUL-2 as it does in the SKP1-CUL-1-F box protein ligase complex. We also found that pVHL associates with two proteins, p100 and p220, which migrate at a similar molecular weight as two major bands in the ubiquitination assay. Furthermore, naturally occurring pVHL missense mutations, including mutants capable of forming a complex with elongin B-elongin C-CUL-2, fail to associate with p100 and p220 and cannot exhibit the E3 ligase activity. These results suggest that pVHL might be the substrate recognition subunit of the VBC-CUL-2 E3 ligase. This is also, to our knowledge, the first example of a human tumor-suppressor protein being directly involved in the ubiquitin conjugation system which leads to the targeted degradation of substrate proteins. PMID- 10535941 TI - SnoN and Ski protooncoproteins are rapidly degraded in response to transforming growth factor beta signaling. AB - Transforming growth factor beta (TGF-beta) regulates a variety of physiologic processes, including growth inhibition, differentiation, and induction of apoptosis. Some TGF-beta-initiated signals are conveyed through Smad3; TGF-beta binding to its receptors induces phosphorylation of Smad3, which then migrates to the nucleus where it functions as a transcription factor. We describe here the association of Smad3 with the nuclear protooncogene protein SnoN. Overexpression of SnoN represses transcriptional activation by Smad3. Activation of TGF-beta signaling leads to rapid degradation of SnoN and, to a lesser extent, of the related Ski protein, and this degradation is likely mediated by cellular proteasomes. These results demonstrate the existence of a cascade of the TGF-beta signaling pathway, which, upon TGF-beta stimulation, leads to the destruction of protooncoproteins that antagonize the activation of the TGF-beta signaling. PMID- 10535942 TI - Opening of a monomer-monomer interface of the trimeric bacteriophage T4-coded GP45 sliding clamp is required for clamp loading onto DNA. AB - The replication system of bacteriophage T4 uses a trimeric ring-shaped processivity clamp (gp45) to tether the replication polymerase (gp43) to the template-primer DNA. This ring is placed onto the DNA by an ATPase-driven clamp loading complex (gp44/62) where it then transfers, in closed form, to the polymerase. It generally has been assumed that one of the functions of the loading machinery is to open the clamp to place it around the DNA. However, the mechanism by which this occurs has not been fully defined. In this study we design and characterize a double-mutant gp45 protein that contains pairs of cysteine residues located at each monomer-monomer interface of the trimeric clamp. This mutant protein is functionally equivalent to wild-type gp45. However, when all three monomer-monomer interfaces are tethered by covalent crosslinks formed (reversibly or irreversibly) between the cysteine pairs these closed clamps can no longer be loaded onto the DNA nor onto the polymerase, effectively eliminating processive strand-displacement DNA synthesis. Analysis of the individual steps of the clamp-loading process shows that the ATPase-dependent interactions between the clamp and the clamp loader that precede DNA binding are hyperstimulated by the covalently crosslinked ring, suggesting that binding of the closed ring induces a futile, ATP-driven, ring-opening cycle. These findings and others permit further characterization and ordering of the steps involved in the T4 clamp-loading process. PMID- 10535943 TI - Ku is associated with the telomere in mammals. AB - Telomeres are specialized DNA/protein complexes that comprise the ends of eukaryotic chromosomes. The highly expressed Ku heterodimer, composed of 70 and 80 K(d) subunits (Ku70 and Ku80), is the high-affinity DNA binding component of the DNA-dependent protein kinase. Ku is critical for nonhomologous DNA double stranded break repair and site-specific recombination of V(D)J gene segments. Ku also plays an important role in telomere maintenance in yeast. Herein, we report, using an in vivo crosslinking method, that human and hamster telomeric DNAs specifically coimmunoprecipitate with human Ku80 after crosslinking. Localization of Ku to the telomere does not depend on the DNA-dependent protein kinase catalytic component. These findings suggest a direct link between Ku and the telomere in mammalian cells. PMID- 10535944 TI - Rapid treadmilling of brain microtubules free of microtubule-associated proteins in vitro and its suppression by tau. AB - We have determined the treadmilling rate of brain microtubules (MTs) free of MT associated proteins (MAPs) at polymer mass steady state in vitro by using [(3)H]GTP-exchange. We developed buffer conditions that suppressed dynamic instability behavior by approximately 10-fold to minimize the contribution of dynamic instability to total tubulin-GTP exchange. The MTs treadmilled rapidly under the suppressed dynamic instability conditions, at a minimum rate of 0.2 micrometer/min. Thus, rapid treadmilling is an intrinsic property of MAP-free MTs. Further, we show that tau, an axonal stabilizing MAP involved in Alzheimer's disease, strongly suppresses the treadmilling rate. These results indicate that tau's function in axons might involve suppression of axonal MT treadmilling. We describe mathematically how treadmilling and dynamic instability are mechanistically distinct MT behaviors. Finally, we present a model that explains how small changes in the critical tubulin subunit concentration at MT minus ends, caused by intrinsic differences in rate constants or regulatory proteins, could produce large changes in the treadmilling rate. PMID- 10535945 TI - The solution structure of the Zalpha domain of the human RNA editing enzyme ADAR1 reveals a prepositioned binding surface for Z-DNA. AB - Double-stranded RNA deaminase I (ADAR1) contains the Z-DNA binding domain Zalpha. Here we report the solution structure of free Zalpha and map the interaction surface with Z-DNA, confirming roles previously assigned to residues by mutagenesis. Comparison with the crystal structure of the (Zalpha)(2)/Z-DNA complex shows that most Z-DNA contacting residues in free Zalpha are prepositioned to bind Z-DNA, thus minimizing the entropic cost of binding. Comparison with homologous (alpha+beta)helix-turn-helix/B-DNA complexes suggests that binding of Zalpha to B-DNA is disfavored by steric hindrance, but does not eliminate the possibility that related domains may bind to both B- and Z-DNA. PMID- 10535946 TI - An optimal Mg(2+) concentration for kinetic folding of the tetrahymena ribozyme. AB - Divalent metal ions, such as Mg(2+), are generally required for tertiary structure formation in RNA. Although the role of Mg(2+) binding in RNA-folding equilibria has been studied extensively, little is known about the role of Mg(2+) in RNA-folding kinetics. In this paper, we explore the effect of Mg(2+) on the rate-limiting step in the kinetic folding pathway of the Tetrahymena ribozyme. Analysis of these data reveals the presence of a Mg(2+)-stabilized kinetic trap that slows folding at higher Mg(2+) concentrations. Thus, the Tetrahymena ribozyme folds with an optimal rate at 2 mM Mg(2+), just above the concentration required for stable structure formation. These results suggest that thermodynamic and kinetic folding of RNA are cooptimized at a Mg(2+) concentration that is sufficient to stabilize the folded form but low enough to avoid kinetic traps and misfolding. PMID- 10535947 TI - The use of site-directed fluorophore labeling and donor-donor energy migration to investigate solution structure and dynamics in proteins. AB - The use of molecular genetics for introducing fluorescent molecules enables the use of donor-donor energy migration to determine intramolecular distances in a variety of proteins. This approach can be applied to examine the overall molecular dimensions of proteins and to investigate structural changes upon interactions with specific target molecules. In this report, the donor-donor energy migration method is demonstrated by experiments with the latent form of plasminogen activator inhibitor type 1. Based on the known x-ray structure of plasminogen activator inhibitor type 1, three positions forming the corners of a triangle were chosen. Double Cys substitution mutants (V106C-H185C, H185C-M266C, and M266C-V106C) and corresponding single substitution mutants (V106C, H185C, and M266C) were created and labeled with a sulfhydryl specific derivative of BODIPY (=the D molecule). The side lengths of this triangle were obtained from analyses of the experimental data. The analyses account for the local anisotropic order and rotational motions of the D molecules, as well as for the influence of a partial DD-labeling. The distances, as determined from x-ray diffraction, between the C(alpha)-atoms of the positions V106C-H185C, H185C-M266C, and M266C-V106C were 60.9, 30.8, and 55.1 A, respectively. These are in good agreement with the distances of 54 +/- 4, 38 +/- 3, and 55 +/- 3 A, as determined between the BODIPY groups attached via linkers to the same residues. Although the positions of the D molecules and the C(alpha)-atoms physically cannot coincide, there is a reasonable agreement between the methods. PMID- 10535948 TI - Folding protein models with a simple hydrophobic energy function: the fundamental importance of monomer inside/outside segregation. AB - The present study explores a "hydrophobic" energy function for folding simulations of the protein lattice model. The contribution of each monomer to conformational energy is the product of its "hydrophobicity" and the number of contacts it makes, i.e., E(h,c) = -Sigma N/i=1 c(i)h(i) = -(h.c) is the negative scalar product between two vectors in N-dimensional cartesian space: h = (h1,., hN), which represents monomer hydrophobicities and is sequence-dependent; and c = (c(1),., c(N)), which represents the number of contacts made by each monomer and is conformation-dependent. A simple theoretical analysis shows that restrictions are imposed concomitantly on both sequences and native structures if the stability criterion for protein-like behavior is to be satisfied. Given a conformation with vector c, the best sequence is a vector h on the direction upon which the projection of c - c is maximal, where c is the diagonal vector with components equal to c, the average number of contacts per monomer in the unfolded state. Best native conformations are suggested to be not maximally compact, as assumed in many studies, but the ones with largest variance of contacts among its monomers, i.e., with monomers tending to occupy completely buried or completely exposed positions. This inside/outside segregation is reflected on an apolar/polar distribution on the corresponding sequence. Monte Carlo simulations in two dimensions corroborate this general scheme. Sequences targeted to conformations with large contact variances folded cooperatively with thermodynamics of a two-state transition. Sequences targeted to maximally compact conformations, which have lower contact variance, were either found to have degenerate ground state or to fold with much lower cooperativity. PMID- 10535950 TI - Measures of residue density in protein structures. AB - A hierarchy of residue density assessments and packing properties in protein structures are contrasted, including a regular density, a variety of charge densities, a hydrophobic density, a polar density, and an aromatic density. These densities are investigated by alternative distance measures and also at the interface of multiunit structures. Amino acids are divided into nine structural categories according to three secondary structure states and three solvent accessibility levels. To take account of amino acid abundance differences across protein structures, we normalize the observed density by the expected density defining a density index. Solvent accessibility levels exert the predominant influence in determinations of the regular residue density. Explicitly, the regular density values vary approximately linearly with respect to solvent accessibility levels, the linearity parameters depending on the amino acid. The charge index reveals pronounced inequalities between lysine and arginine in their interactions with acidic residues. The aromatic density calculations in all structural categories parallel the regular density calculations, indicating that the aromatic residues are distributed as a random sample of all residues. Moreover, aromatic residues are found to be over-represented in the neighborhood of all amino acids. This result might be attributed to nucleation sites and protein stability being substantially associated with aromatic residues. PMID- 10535949 TI - Tropomyosin directly modulates actomyosin mechanical performance at the level of a single actin filament. AB - Muscle contraction is the result of myosin cross-bridges (XBs) cyclically interacting with the actin-containing thin filament. This interaction is modulated by the thin filament regulatory proteins, troponin and tropomyosin (Tm). With the use of an in vitro motility assay, the role of Tm in myosin's ability to generate force and motion was assessed. At saturating myosin surface densities, Tm had no effect on thin filament velocity. However, below 50% myosin saturation, a significant reduction in actin-Tm filament velocity was observed, with complete inhibition of movement occurring at 12. 5% of saturating surface densities. Under similar conditions, actin filaments alone demonstrated no reduction in velocity. The effect of Tm on force generation was assessed at the level of a single thin filament. In the absence of Tm, isometric force was a linear function of the density of myosin on the motility surface. At 50% myosin surface saturation, the presence of Tm resulted in a 2-fold enhancement of force relative to actin alone. However, no further potentiation of force was observed with Tm at saturating myosin surface densities. These results indicate that, in the presence of Tm, the strong binding of myosin cooperatively activates the thin filament. The inhibition of velocity at low myosin densities and the potentiation of force at higher myosin densities suggest that Tm can directly modulate the kinetics of a single myosin XB and the recruitment of a population of XBs, respectively. At saturating myosin conditions, Tm does not appear to affect the recruitment or the kinetics of myosin XBs. PMID- 10535951 TI - Atom density in protein structures. AB - The residue environment in protein structures is studied with respect to the density of carbon (C), oxygen (O), and nitrogen (N) atoms within a certain distance (say 5 A) of each residue. Two types of environments are evaluated: one based on side-chain atom contacts (abbreviated S-S) and the other based on all atom (side-chain + backbone) contacts (abbreviated A-A). Different atom counts are observed about nine-residue structural categories defined by three solvent accessibility levels and three secondary structure states. Among the structural categories, the S-S atom count ratios generally vary more than the A-A atom count ratios because of the fact that the backbone (O) and (N) atoms contribute equal counts. Secondary structure affects the (C) density for the A-A contacts whereas secondary structure has little influence on the (C) density for the S-S contacts. For S-S contacts, a greater density of (O) over (N) atom neighbors stands out in the environment of most amino acid types. By contrast, for A-A contacts, independent of the solvent accessibility levels, the ratio (O)/(N) is approximately 1 in helical states, consistent with the geometry of alpha-helical residues whose side-chains tilt oppositely to the amino to carboxy alpha-helical axis. The highest ratio of neighbor (O)/(N) is achieved under solvent exposed conditions. This (O) vs. (N) prevalence is advantageous at the protein surface that generally exhibits an acid excess that helps to enhance protein solubility in the cell and to avoid nonspecific interactions with phosphate groups of DNA, RNA, and other plasma constituents. PMID- 10535952 TI - Odor space and olfactory processing: collective algorithms and neural implementation. AB - Several basic olfactory tasks must be solved by highly olfactory animals, including background suppression, multiple object separation, mixture separation, and source identification. The large number N of classes of olfactory receptor cells-hundreds or thousands-permits the use of computational strategies and algorithms that would not be effective in a stimulus space of low dimension. A model of the patterns of olfactory receptor responses, based on the broad distribution of olfactory thresholds, is constructed. Representing one odor from the viewpoint of another then allows a common description of the most important basic problems and shows how to solve them when N is large. One possible biological implementation of these algorithms uses action potential timing and adaptation as the "hardware" features that are responsible for effective neural computation. PMID- 10535954 TI - Self-assembly of fibronectin into fibrillar networks underneath dipalmitoyl phosphatidylcholine monolayers: role of lipid matrix and tensile forces. AB - The cell-mediated assembly of fibronectin (Fn) into fibrillar matrices is a complex multistep process that is incompletely understood because of the chemical complexity of the extracellular matrix and a lack of experimental control over molecular interactions and dynamic events. We have identified conditions under which Fn assembles into extended fibrillar networks after adsorption to a dipalmitoyl phosphatidylcholine (DPPC) monolayer in contact with physiological buffer. We propose a sequential model for the Fn assembly pathway, which involves the orientation of Fn underneath the lipid monolayer by insertion into the liquid expanded (LE) phase of DPPC. Attractive interactions between these surface anchored proteins and the liquid condensed (LC) domains leads to Fn enrichment at domain edges. Spontaneous self-assembly into fibrillar networks, however, occurs only after expansion of the DPPC monolayer from the LC phase though the LC/LE phase coexistence. Upon monolayer expansion, the domain boundaries move apart while attractive interactions among Fn molecules and between Fn and domain edges produce a tensile force on the proteins that initiates fibril assembly. The resulting fibrils have been characterized in situ by using fluorescence and light scattering microscopy. We have found striking similarities between fibrils produced under DPPC monolayers and those found on cellular surfaces, including their assembly pathways. PMID- 10535953 TI - Exploring the origins of topological frustration: design of a minimally frustrated model of fragment B of protein A. AB - Topological frustration in an energetically unfrustrated off-lattice model of the helical protein fragment B of protein A from Staphylococcus aureus was investigated. This G-type model exhibited thermodynamic and kinetic signatures of a well-designed two-state folder with concurrent collapse and folding transitions and single exponential kinetics at the transition temperature. Topological frustration is determined in the absence of energetic frustration by the distribution of Fersht phi values. Topologically unfrustrated systems present a unimodal distribution sharply peaked at intermediate phi, whereas highly frustrated systems display a bimodal distribution peaked at low and high phi values. The distribution of phi values in protein A was determined both thermodynamically and kinetically. Both methods yielded a unimodal distribution centered at phi = 0.3 with tails extending to low and high phi values, indicating the presence of a small amount of topological frustration. The contacts with high phi values were located in the turn regions between helices I and II and II and III, intimating that these hairpins are in large part required in the transition state. Our results are in good agreement with all-atom simulations of protein A, as well as lattice simulations of a three- letter code 27-mer (which can be compared with a 60-residue helical protein). The relatively broad unimodal distribution of phi values obtained from the all-atom simulations and that from the minimalist model for the same native fold suggest that the structure of the transition state ensemble is determined mostly by the protein topology and not energetic frustration. PMID- 10535955 TI - Structure-based conformational preferences of amino acids. AB - Proteins can be very tolerant to amino acid substitution, even within their core. Understanding the factors responsible for this behavior is of critical importance for protein engineering and design. Mutations in proteins have been quantified in terms of the changes in stability they induce. For example, guest residues in specific secondary structures have been used as probes of conformational preferences of amino acids, yielding propensity scales. Predicting these amino acid propensities would be a good test of any new potential energy functions used to mimic protein stability. We have recently developed a protein design procedure that optimizes whole sequences for a given target conformation based on the knowledge of the template backbone and on a semiempirical potential energy function. This energy function is purely physical, including steric interactions based on a Lennard-Jones potential, electrostatics based on a Coulomb potential, and hydrophobicity in the form of an environment free energy based on accessible surface area and interatomic contact areas. Sequences designed by this procedure for 10 different proteins were analyzed to extract conformational preferences for amino acids. The resulting structure-based propensity scales show significant agreements with experimental propensity scale values, both for alpha-helices and beta-sheets. These results indicate that amino acid conformational preferences are a natural consequence of the potential energy we use. This confirms the accuracy of our potential and indicates that such preferences should not be added as a design criterion. PMID- 10535956 TI - Effectors of a developmental mitogen-activated protein kinase cascade revealed by expression signatures of signaling mutants. AB - Despite the importance of mitogen-activated protein kinase (MAPK) signaling in eukaryotic biology, the mechanisms by which signaling yields phenotypic changes are poorly understood. We have combined transcriptional profiling with genetics to determine how the Kss1 MAPK signaling pathway controls dimorphic development in Saccharomyces cerevisiae. This analysis identified dozens of transcripts that are regulated by the pathway, whereas previous work had identified only a single downstream target, FLO11. One of the MAPK-regulated genes is PGU1, which encodes a secreted enzyme that hydrolyzes polygalacturonic acid, a structural barrier to microbial invasion present in the natural plant substrate of S. cerevisiae. A third key transcriptional target is the G(1) cyclin gene CLN1, a morphogenetic regulator that we show to be essential for pseudohyphal growth. In contrast, the homologous CLN2 cyclin gene is dispensable for development. Thus, the Kss1 MAPK cascade programs development by coordinately modulating a cell adhesion factor, a secreted host-destroying activity, and a specialized subunit of the Cdc28 cyclin dependent kinase. PMID- 10535957 TI - A cytomegalovirus-encoded mitochondria-localized inhibitor of apoptosis structurally unrelated to Bcl-2. AB - Human cytomegalovirus (CMV), a herpesvirus that causes congenital disease and opportunistic infections in immunocompromised individuals, encodes functions that facilitate efficient viral propagation by altering host cell behavior. Here we show that CMV blocks apoptosis mediated by death receptors and encodes a mitochondria-localized inhibitor of apoptosis, denoted vMIA, capable of suppressing apoptosis induced by diverse stimuli. vMIA, a product of the viral UL37 gene, inhibits Fas-mediated apoptosis at a point downstream of caspase-8 activation and Bid cleavage but upstream of cytochrome c release, while residing in mitochondria and associating with adenine nucleotide translocator. These functional properties resemble those ascribed to Bcl-2; however, the absence of sequence similarity to Bcl-2 or any other known cell death suppressors suggests that vMIA defines a previously undescribed class of anti-apoptotic proteins. PMID- 10535958 TI - The importin/karyopherin Kap114 mediates the nuclear import of TATA-binding protein. AB - Two high copy suppressors of temperature-sensitive TATA-binding protein (TBP) mutants were isolated. One suppressor was TIF51A, which encodes eukaryotic translation initiation factor 5A. The other high copy suppressor, YGL241W, also known as KAP114, is one of 14 importin/karyopherin proteins in yeast. These proteins mediate the transport of specific macromolecules into and out of the nucleus. Cells lacking Kap114 partially mislocalize TBP to the cytoplasm. Kap114 binds TBP in vitro, and binding is disrupted in the presence of GTPgammaS. Therefore, Kap114 is an importer of TBP into the nucleus, but alternative import pathways must also exist. PMID- 10535959 TI - Casein kinase iepsilon in the wnt pathway: regulation of beta-catenin function. AB - Wnt and its intracellular effector beta-catenin regulate developmental and oncogenic processes. Using expression cloning to identify novel components of the Wnt pathway, we isolated casein kinase Iepsilon (CKIepsilon). CKIepsilon mimicked Wnt in inducing a secondary axis in Xenopus, stabilizing beta-catenin, and stimulating gene transcription in cells. Inhibition of endogenous CKIepsilon by kinase-defective CKIepsilon or CKIepsilon antisense-oligonucleotides attenuated Wnt signaling. CKIepsilon was in a complex with axin and other downstream components of the Wnt pathway, including Dishevelled. CKIepsilon appears to be a positive regulator of the pathway and a link between upstream signals and the complexes that regulate beta-catenin. PMID- 10535960 TI - Tumor burden and clonality in multiple intestinal neoplasia mouse/normal mouse aggregation chimeras. AB - Aggregation chimeras were formed between C57BL/6 mice heterozygous for the Apc(min) (Min) mutation and wild-type SWR mice, that differ in their Pla2g2a status, a modifier of Apc(min), and also in their resistance to intestinal polyp formation. Variation in the dolichos biflorus agglutinin-staining patterns of the intestines of these mouse strains was used to determine the chimeric composition of the intestine in individual mice and to examine the clonal composition of adenomas. Macroscopic adenoma numbers in chimeric mice were compared with the expected adenoma numbers based on the percentage of C57BL/6J-Apc(min/+) epithelium in individual mice. These results unexpectedly show that there was no apparent inhibitory effect of the SWR-derived (Pla2g2a wild-type) tissue on adenoma formation in the C57BL/6J-Apc(min/+) epithelium. This suggests that the main genetic modifiers of the Min phenotype act at a cellular or crypt-restricted level with no discernable systemic effect. All adenomas were seen to contain C57BL/6J-Apc(min/+)-derived epithelium, confirming that the germ-line mutation of the mApc gene is necessary to initiate tumorigenesis in this model system, and that the mApc gene acts in a cell autonomous fashion. PMID- 10535961 TI - Identification of the endophilins (SH3p4/p8/p13) as novel binding partners for the beta1-adrenergic receptor. AB - Several G-protein coupled receptors, such as the beta1-adrenergic receptor (beta1 AR), contain polyproline motifs within their intracellular domains. Such motifs in other proteins are known to mediate protein-protein interactions such as with Src homology (SH)3 domains. Accordingly, we used the proline-rich third intracellular loop of the beta1-AR either as a glutathione S-transferase fusion protein in biochemical "pull-down" assays or as bait in the yeast two-hybrid system to search for interacting proteins. Both approaches identified SH3p4/p8/p13 (also referred to as endophilin 1/2/3), a SH3 domain-containing protein family, as binding partners for the beta1-AR. In vitro and in human embryonic kidney (HEK) 293 cells, SH3p4 specifically binds to the third intracellular loop of the beta1-AR but not to that of the beta2-AR. Moreover, this interaction is mediated by the C-terminal SH3 domain of SH3p4. Functionally, overexpression of SH3p4 promotes agonist-induced internalization and modestly decreases the Gs coupling efficacy of beta1-ARs in HEK293 cells while having no effect on beta2-ARs. Thus, our studies demonstrate a role of the SH3p4/p8/p13 protein family in beta1-AR signaling and suggest that interaction between proline rich motifs and SH3-containing proteins may represent a previously underappreciated aspect of G-protein coupled receptor signaling. PMID- 10535962 TI - Rapid and efficient fusion of phospholipid vesicles by the alpha-helical core of a SNARE complex in the absence of an N-terminal regulatory domain. AB - A protease-resistant core domain of the neuronal SNARE complex consists of an alpha-helical bundle similar to the proposed fusogenic core of viral fusion proteins [Skehel, J. J. & Wiley, D. C. (1998) Cell 95, 871-874]. We find that the isolated core of a SNARE complex efficiently fuses artificial bilayers and does so faster than full length SNAREs. Unexpectedly, a dramatic increase in speed results from removal of the N-terminal domain of the t-SNARE syntaxin, which does not affect the rate of assembly of v-t SNARES. In the absence of this negative regulatory domain, the half-time for fusion of an entire population of lipid vesicles by isolated SNARE cores ( approximately 10 min) is compatible with the kinetics of fusion in many cell types. PMID- 10535964 TI - The active digestion of uniparental chloroplast DNA in a single zygote of Chlamydomonas reinhardtii is revealed by using the optical tweezer. AB - The non-Mendelian inheritance of organelle genes is a phenomenon common to almost all eukaryotes, and in the isogamous alga Chlamydomonas reinhardtii, chloroplast (cp) genes are transmitted from the mating type positive (mt(+)) parent. In this study, the preferential disappearance of the fluorescent cp nucleoids of the mating type negative (mt(-)) parent was observed in living young zygotes. To study the change in cpDNA molecules during the preferential disappearance, the cpDNA of mt(+) or mt(-) origin was labeled separately with bacterial aadA gene sequences. Then, a single zygote with or without cp nucleoids was isolated under direct observation by using optical tweezers and investigated by nested PCR analysis of the aadA sequences. This demonstrated that cpDNA molecules are digested completely during the preferential disappearance of mt(-) cp nucleoids within 10 min, whereas mt(+) cpDNA and mitochondrial DNA are protected from the digestion. These results indicate that the non-Mendelian transmission pattern of organelle genes is determined immediately after zygote formation. PMID- 10535963 TI - Content mixing and membrane integrity during membrane fusion driven by pairing of isolated v-SNAREs and t-SNAREs. AB - Membrane bilayer fusion has been shown to be mediated by v- and t-SNAREs initially present in separate populations of liposomes and to occur with high efficiency at a physiologically meaningful rate. Lipid mixing was demonstrated to involve both the inner and the outer leaflets of the membrane bilayer. Here, we use a fusion assay that relies on duplex formation of oligonucleotides introduced in separate liposome populations and report that SNARE proteins suffice to mediate complete membrane fusion accompanied by mixing of luminal content. We also find that SNARE-mediated membrane fusion does not compromise the integrity of liposomes. PMID- 10535965 TI - Calcium-independent activation of endothelial nitric oxide synthase by ceramide. AB - The endothelial isoform of NO synthase (eNOS) is targeted to sphingolipid enriched signal-transducing microdomains in the plasma membrane termed caveolae. Among the caveolae-targeted sphingolipids are the ceramides, a class of acylated sphingosine compounds that have been implicated in diverse cellular responses. We have explored the role of ceramide analogues in eNOS signaling in cultured bovine aortic endothelial cells (BAEC). Addition of the ceramide analogue N acetylsphingosine (C(2)-ceramide; 5 microM) to intact BAEC leads to a significant increase in NO synthase activity (assayed by using the fluorescent indicator 4,5 diaminofluorescein) and translocation of eNOS from the endothelial cell membrane to intracellular sites (measured by using quantitative immunofluorescence techniques); the biologically inactive ceramide N-acetyldihydrosphingosine is entirely without effect. C(2)-ceramide-induced eNOS activation and translocation are unaffected by the intracellular calcium chelator 1, 2-bis-o aminophenoxyethane-N,N,N',N'-tetraacetic acid (BAPTA). Using the calcium-specific fluorescent indicator fluo-3, we also found that C(2)-ceramide activation of eNOS is unaccompanied by a drug-induced increase in intracellular calcium. These findings stand in sharp contrast to the mechanism by which bradykinin, estradiol, and other mediators acutely activate eNOS, in which a rapid, agonist-promoted increase in intracellular calcium is required. Finally, we show that treatment of BAEC with bradykinin causes a significant increase in cellular ceramide content; the response to bradykinin has an EC(50) of 3 nM and is blocked by the bradykinin B(2)-receptor antagonist HOE140. Bradykinin-induced ceramide generation could represent a mechanism for longer-term regulation of eNOS activity. Our results suggest that ceramide functions independently of Ca(2+)-regulated pathways to promote activation and translocation of eNOS, and that this lipid mediator may represent a physiological regulator of eNOS in vascular endothelial cells. PMID- 10535966 TI - Proteomic definition of normal human luminal and myoepithelial breast cells purified from reduction mammoplasties. AB - Normal human luminal and myoepithelial breast cells separately purified from a set of 10 reduction mammoplasties by using a double antibody magnetic affinity cell sorting and Dynabead immunomagnetic technique were used in two-dimensional gel proteome studies. A total of 43,302 proteins were detected across the 20 samples, and a master image for each cell type comprising a total of 1,738 unique proteins was derived. Differential analysis identified 170 proteins that were elevated 2-fold or more between the two breast cell types, and 51 of these were annotated by tandem mass spectrometry. Muscle-specific enzyme isoforms and contractile intermediate filaments including tropomyosin and smooth muscle (SM22) alpha protein were detected in the myoepithelial cells, and a large number of cytokeratin subclasses and isoforms characteristic of luminal cells were detected in this cell type. A further 134 nondifferentially regulated proteins were also annotated from the two breast cell types, making this the most extensive study to date of the protein expression map of the normal human breast and the basis for future studies of purified breast cancer cells. PMID- 10535967 TI - Postgastrulation Smad2-deficient embryos show defects in embryo turning and anterior morphogenesis. AB - SMAD2 is a member of the transforming growth factor beta and activin-signaling pathway. To examine the role of Smad2 in postgastrulation development, we independently generated mice with a null mutation in this gene. Smad2-deficient embryos die around day 7.5 of gestation because of failure of gastrulation and failure to establish an anterior-posterior (A-P) axis. Expression of the homeobox gene Hex (the earliest known marker of the A-P polarity and the prospective head organizer) was found to be missing in Smad2-deficient embryos. Homozygous mutant embryos and embryonic stem cells formed mesoderm derivatives revealing that mesoderm induction is SMAD2 independent. In the presence of wild-type extraembryonic tissues, Smad2-deficient embryos developed beyond 7.5 and up to 10.5 days postcoitum, demonstrating a requirement for SMAD2 in extraembryonic tissues for the generation of an A-P axis and gastrulation. The rescued postgastrulation embryos showed malformation of head structures, abnormal embryo turning, and cyclopia. Our results show that Smad2 expression is required at several stages during embryogenesis. PMID- 10535968 TI - Developmental exposure to vasopressin increases aggression in adult prairie voles. AB - Although the biological roots of aggression have been the source of intense debate, the precise physiological mechanisms responsible for aggression remain poorly understood. In most species, aggression is more common in males than females; thus, gonadal hormones have been a focal point for research in this field. Although gonadal hormones have been shown to influence the expression of aggression, in many cases aggression can continue after castration, indicating that testicular steroids are not completely essential for the expression of aggression. Recently, the mammalian neuropeptide arginine vasopressin (AVP) has been implicated in aggression. AVP plays a particularly important role in social behavior in monogamous mammals, such as prairie voles (Microtus ochrogaster). In turn, the effects of social experiences may be mediated by neuropeptides, including AVP. For example, sexually naive prairie voles are rarely aggressive. However, 24 h after the onset of mating, males of this species become significantly aggressive toward strangers. Likewise, in adult male prairie voles, central (intracerebroventricular) injections of AVP can significantly increase intermale aggression, suggesting a role for AVP in the expression of postcopulatory aggression in adult male prairie voles. In this paper, we demonstrate that early postnatal exposure to AVP can have long-lasting effects on the tendency to show aggression, producing levels of aggression in sexually naive, adult male prairie voles that are comparable to those levels observed after mating. Females showed less aggression and were less responsive to exogenous AVP, but the capacity of an AVP V(1a) receptor antagonist to block female aggression also implicates AVP in the development of female aggression. PMID- 10535970 TI - Self-organized defensive behavior in honeybees. AB - We investigated the defensive behavior of honeybees under controlled experimental conditions. During an attack on two identical targets, the spatial distribution of stings varied as a function of the total number of stings, evincing the classic "pitchfork bifurcation" phenomenon of nonlinear dynamics. The experimental results support a model of defensive behavior based on a self organizing mechanism. The model helps to explain several of the characteristic features of the honeybee defensive response: (i) the ability of the colony to localize and focus its attack, (ii) the strong variability between different hives in the intensity of attack, as well as (iii) the variability observed within the same hive, and (iv) the ability of the colony to amplify small differences between the targets. PMID- 10535969 TI - The STAR protein QKI-6 is a translational repressor. AB - The signal transduction and activation of RNA (STAR) family of RNA-binding proteins, whose members are evolutionarily conserved from yeast to humans, are important for a number of developmental decisions. For example, in the mouse, quaking proteins (QKI-5, QKI-6, and QKI-7) are essential for embryogenesis and myelination, whereas a closely related protein in Caenorhabditis elegans, germline defective-1 (GLD-1), is necessary for germ-line development. Recently, GLD-1 was found to be a translational repressor that acts through regulatory elements, called TGEs (for tra-2 and GLI elements), present in the 3' untranslated region of the sex-determining gene tra-2. This gene promotes female development, and repression of tra-2 translation by TGEs is necessary for the male cell fates. The finding that GLD-1 inhibits tra-2 translation raises the possibility that other STAR family members act by a similar mechanism to control gene activity. Here we demonstrate, both in vitro and in vivo, that QKI-6 functions in the same manner as GLD-1 and can specifically bind to TGEs to repress translation of reporter constructs containing TGEs. In addition, expression of QKI-6 in C. elegans wild-type hermaphrodites or in hermaphrodites that are partially masculinized by a loss-of-function mutation in the sex determining gene tra-3 results in masculinization of somatic tissues, consistent with QKI-6 repressing the activity of tra-2. These results strongly suggest that QKI-6 may control gene activity by operating through TGEs to regulate translation. In addition, our data support the hypothesis that other STAR family members may also be TGE-dependent translational regulators. PMID- 10535971 TI - Two living species of coelacanths? AB - During the period of September 1997 through July 1998, two coelacanth fishes were captured off Manado Tua Island, Sulawesi, Indonesia. These specimens were caught almost 10,000 km from the only other known population of living coelacanths, Latimeria chalumnae, near the Comores. The Indonesian fish was described recently as a new species, Latimeria menadoensis, based on morphological differentiation and DNA sequence divergence in fragments of the cytochrome b and 12S rRNA genes. We have obtained the sequence of 4,823 bp of mitochondrial DNA from the same specimen, including the entire genes for cytochrome b, 12S rRNA, 16S rRNA, four tRNAs, and the control region. The sequence is 4.1% different from the published sequence of an animal captured from the Comores, indicating substantial divergence between the Indonesian and Comorean populations. Nine morphological and meristic differences are purported to distinguish L. menadoensis and L. chalumnae, based on comparison of a single specimen of L. menadoensis to a description of five individuals of L. chalumnae from the Comores. A survey of the literature provided data on 4 of the characters used to distinguish L. menadoensis from L. chalumnae from an additional 16 African coelacanths; for all 4 characters, the Indonesian sample was within the range of variation reported for the African specimens. Nonetheless, L. chalumnae and L. menadoensis appear to be separate species based on divergence of mitochondrial DNA. PMID- 10535973 TI - Macroevolution of insect-plant associations: the relevance of host biogeography to host affiliation. AB - Identifying the factors that have promoted host shifts by phytophagous insects at a macroevolutionary scale is critical to understanding the associations between plants and insects. We used molecular phylogenies of the beetle genus Blepharida and its host genus Bursera to test whether these insects have been using hosts with widely overlapping ranges over evolutionary time. We also quantified the importance of host range coincidence relative to host chemistry and host phylogenetic relatedness. Overall, the evolution of host use of these insects has not been among hosts that are geographically similar. Host chemistry is the factor that best explains their macroevolutionary patterns of host use. Interestingly, one exceptional polyphagous species has shifted among geographically close chemically dissimilar plants. PMID- 10535972 TI - Evidence for the recent horizontal transfer of long terminal repeat retrotransposon. AB - The evolutionary dynamics existing between transposable elements (TEs) and their host genomes have been likened to an "arms race." The selfish drive of TEs to replicate, in turn, elicits the evolution of host-mediated regulatory mechanisms aimed at repressing transpositional activity. It has been postulated that horizontal (cross-species) transfer may be one effective strategy by which TEs and other selfish genes can escape host-mediated silencing mechanisms over evolutionary time; however, to date, the most definitive evidence that TEs horizontally transfer between species has been limited to class II or DNA-type elements. Evidence that the more numerous and widely distributed retroelements may also be horizontally transferred between species has been more ambiguous. In this paper, we report definitive evidence for a recent horizontal transfer of the copia long terminal repeat retrotransposon between Drosophila melanogaster and Drosophila willistoni. PMID- 10535975 TI - Calibrating bacterial evolution. AB - Attempts to calibrate bacterial evolution have relied on the assumption that rates of molecular sequence divergence in bacteria are similar to those of higher eukaryotes, or to those of the few bacterial taxa for which ancestors can be reliably dated from ecological or geological evidence. Despite similarities in the substitution rates estimated for some lineages, comparisons of the relative rates of evolution at different classes of nucleotide sites indicate no basis for their universal application to all bacteria. However, there is evidence that bacteria have a constant genome-wide mutation rate on an evolutionary time scale but that this rate differs dramatically from the rate estimated by experimental methods. PMID- 10535974 TI - All males are not created equal: fertility differences depend on gamete recognition polymorphisms in sea urchins. AB - Behaviors, morphologies, and genetic loci directly involved in reproduction have been increasingly shown to be polymorphic within populations. Explaining how such variants are maintained by selection is crucial to understanding the genetic basis of fertility differences, but direct tests of how alleles at reproductive loci affect fertility are rare. In the sea urchin genus Echinometra, the protein bindin mediates sperm attachment to eggs, evolves quickly, and is polymorphic within species. Eggs exposed to experimental sperm mixtures show strong discrimination on the basis of the males' bindin genotype. Different females produce eggs that nonrandomly select sperm from different males, showing that variable egg-sperm interactions determine fertility. Eggs select sperm with a bindin genotype similar to their own, suggesting strong linkage between female choice and male trait loci. These experiments demonstrate that alleles at a single locus can have a strong effect on fertilization and that reproductive loci may retain functional polymorphisms through epistatic interactions between male and female traits. They also suggest that positive selection at gamete recognition loci like bindin involves strong selection within species on mate choice interactions. PMID- 10535976 TI - Signaling of need, sibling competition, and the cost of honesty. AB - Young birds and mammals frequently solicit food by means of extravagant and apparently costly begging displays. Much attention has been devoted to the idea that these displays are honest signals of need, and that their apparent cost serves to maintain their honesty. Recent analyses, however, have shown that the cost needed to maintain a fully informative, honest signal may often be so great that both offspring (signaler) and parent (receiver) would do better to refrain from communication. This apparently calls into question the relevance of the costly signaling hypothesis. Here, I show that this argument overlooks the impact of sibling competition. When multiple signalers must compete for the attention of a receiver (as is commonly the case in parent-offspring interactions), I show that (all other things being equal) individual equilibrium signal costs will typically be lower. The greater the number of competitors, the smaller the mean cost, though the maximum level of signal intensity employed by very needy signalers may actually increase with the number of competitors. At the same time, costs become increasingly sensitive to relatedness among signalers as opposed to relatedness between signalers and receivers. As a result of these trends, signaling proves profitable for signalers under a much wider range of conditions when there is competition (though it is still likely to be unprofitable for receivers). PMID- 10535977 TI - Profound misregulation of muscle-specific gene expression in facioscapulohumeral muscular dystrophy. AB - Facioscapulohumeral muscular dystrophy (FSHD) is a neuromuscular disorder characterized by an insidious onset and progressive course. The disease has a frequency of about 1 in 20,000 and is transmitted in an autosomal dominant fashion with almost complete penetrance. Deletion of an integral number of tandemly arrayed 3.3-kb repeat units (D4Z4) on chromosome 4q35 is associated with FSHD but otherwise the molecular basis of the disease and its pathophysiology remain obscure. Comparison of mRNA populations between appropriate cell types can facilitate identification of genes relevant to a particular biological or pathological process. In this report, we have compared mRNA populations of FSHD and normal muscle. Unexpectedly, the dystrophic muscle displayed profound alterations in gene expression characterized by severe underexpression or overexpression of specific mRNAs. Intriguingly, many of the deregulated mRNAs are muscle specific. Our results suggest that a global misregulation of gene expression is the underlying basis for FSHD, distinguishing it from other forms of muscular dystrophy. The experimental approach used here is applicable to any genetic disorder whose pathogenic mechanism is incompletely understood. PMID- 10535978 TI - Characterization of the Saccharomyces cerevisiae ERG27 gene encoding the 3-keto reductase involved in C-4 sterol demethylation. AB - The last unidentified gene encoding an enzyme involved in ergosterol biosynthesis in Saccharomyces cerevisiae has been cloned. This gene, designated ERG27, encodes the 3-keto sterol reductase, which, in concert with the C-4 sterol methyloxidase (ERG25) and the C-3 sterol dehydrogenase (ERG26), catalyzes the sequential removal of the two methyl groups at the sterol C-4 position. We developed a strategy to isolate a mutant deficient in converting 3-keto to 3-hydroxy-sterols. An ergosterol auxotroph unable to synthesize sterol or grow without sterol supplementation was mutagenized. Colonies were then selected that were nystatin resistant in the presence of 3-ketoergostadiene and cholesterol. A new ergosterol auxotroph unable to grow on 3-ketosterols without the addition of cholesterol was isolated. The gene (YLR100w) was identified by complementation. Segregants containing the YLR100w disruption failed to grow on various types of 3-keto sterol substrates. Surprisingly, when erg27 was grown on cholesterol- or ergosterol-supplemented media, the endogenous compounds that accumulated were noncyclic sterol intermediates (squalene, squalene epoxide, and squalene dioxide), and there was little or no accumulation of lanosterol or 3-ketosterols. Feeding experiments in which erg27 strains were supplemented with lanosterol (an upstream intermediate of the C-4 demethylation process) and cholesterol (an end product sterol) demonstrated accumulation of four types of 3-keto sterols identified by GC/MS and chromatographic properties: 4-methyl-zymosterone, zymosterone, 4-methyl-fecosterone, and ergosta-7,24 (28)-dien-3-one. In addition, a fifth intermediate was isolated and identified by (1)H NMR as a 4-methyl-24, 25 epoxy-cholesta-7-en-3-one. Implications of these results are discussed. PMID- 10535979 TI - Epigenetic phenotypes distinguish microsatellite-stable and -unstable colorectal cancers. AB - Aberrant DNA methylation is a common phenomenon in human cancer, but its patterns, causes, and consequences are poorly defined. Promoter methylation of the DNA mismatch repair gene MutL homologue (MLH1) has been implicated in the subset of colorectal cancers that shows microsatellite instability (MSI). The present analysis of four MspI/HpaII sites at the MLH1 promoter region in a series of 89 sporadic colorectal cancers revealed two main methylation patterns that closely correlated with the MSI status of the tumors. These sites were hypermethylated in tumor tissue relative to normal mucosa in most MSI(+) cases (31/51, 61%). By contrast, in the majority of MSI(-) cases (20/38, 53%) the same sites showed methylation in normal mucosa and hypomethylation in tumor tissue. Hypermethylation displayed a direct correlation with increasing age and proximal location in the bowel and was accompanied by immunohistochemically documented loss of MLH1 protein both in tumors and in normal tissue. Similar patterns of methylation were observed in the promoter region of the calcitonin gene that does not have a known functional role in tumorigenesis. We propose a model of carcinogenesis where different epigenetic phenotypes distinguish the colonic mucosa in individuals who develop MSI(+) and MSI(-) tumors. These phenotypes may underlie the different developmental pathways that are known to occur in these tumors. PMID- 10535980 TI - LFG: an anti-apoptotic gene that provides protection from Fas-mediated cell death. AB - Programmed cell death regulates a number of biological phenomena, and the apoptotic signal must itself be tightly controlled to avoid inappropriate cell death. We established a genetic screen to search for molecules that inhibit the apoptotic signal from the Fas receptor. Here we report the isolation of a gene, LFG, that protects cells uniquely from Fas but not from the mechanistically related tumor necrosis factor alpha death signal. LFG is widely distributed, but remarkably is highly expressed in the hippocampus. LFG can bind to the Fas receptor, but does not regulate Fas expression or interfere with binding of an agonist antibody. Furthermore LFG does not inhibit binding of FADD to Fas. PMID- 10535981 TI - A genetic method for dissecting the mechanism of transcriptional activator synergy by identical activators. AB - Pairs of transcriptional activators in prokaryotes have been shown to activate transcription synergistically from promoters with two activator binding sites. In some cases, such synergistic effects result from cooperative binding, but in other cases each DNA-bound activator plays a direct role in the activation process by interacting simultaneously with separate surfaces of RNA polymerase. In such cases, each DNA-bound activator must possess a functional activating region, the surface that mediates the interaction with RNA polymerase. When transcriptional activation depends on two or more identical activators, it is not straightforward to test the requirement of each activator for a functional activating region. Here we describe a method for directing a mutationally altered activator to either one or the other binding site, and we demonstrate the use of this method to examine the mechanism of transcriptional activator synergy by the Escherichia coli cyclic AMP receptor protein (CRP) working at an artificial promoter bearing two CRP-binding sites. PMID- 10535982 TI - The MAPKKK Ste11 regulates vegetative growth through a kinase cascade of shared signaling components. AB - In haploid Saccharomyces cerevisiae, the mating and invasive growth (IG) pathways use the same mitogen-activated protein kinase kinase kinase kinase (MAPKKKK, Ste20), MAPKKK (Ste11), MAPKK (Ste7), and transcription factor (Ste12) to promote either G(1) arrest and fusion or foraging in response to distinct stimuli. This exquisite specificity is the result of pathway-specific receptors, G proteins, scaffold protein, and MAPKs. It is currently not thought that the shared signaling components function under the basal conditions of vegetative growth. We tested this hypothesis by searching for mutations that cause lethality when the STE11 gene is deleted. Strikingly, we found that Ste11, together with Ste20, Ste7, Ste12, and the IG MAPK Kss1, functions in a third pathway that promotes vegetative growth and is essential in an och1 mutant that does not synthesize mannoproteins. We term this pathway the STE vegetative growth (SVG) pathway. The SVG pathway functions, in part, to promote cell wall integrity in parallel with the protein kinase C pathway. During vegetative growth, the SVG pathway is inhibited by the mating MAPK Fus3. By contrast, the SVG pathway is constitutively activated in an och1 mutant, suggesting that it senses intracellular changes arising from the loss of mannoproteins. We predict that general proliferative functions may also exist for other MAPK cascades thought only to perform specialized functions. PMID- 10535983 TI - Human ATP-binding cassette transporter 1 (ABC1): genomic organization and identification of the genetic defect in the original Tangier disease kindred. AB - Tangier disease is characterized by low serum high density lipoproteins and a biochemical defect in the cellular efflux of lipids to high density lipoproteins. ABC1, a member of the ATP-binding cassette family, recently has been identified as the defective gene in Tangier disease. We report here the organization of the human ABC1 gene and the identification of a mutation in the ABC1 gene from the original Tangier disease kindred. The organization of the human ABC1 gene is similar to that of the mouse ABC1 gene and other related ABC genes. The ABC1 gene contains 49 exons that range in size from 33 to 249 bp and is over 70 kb in length. Sequence analysis of the ABC1 gene revealed that the proband for Tangier disease was homozygous for a deletion of nucleotides 3283 and 3284 (TC) in exon 22. The deletion results in a frameshift mutation and a premature stop codon starting at nucleotide 3375. The product is predicted to encode a nonfunctional protein of 1,084 aa, which is approximately half the size of the full-length ABC1 protein. The loss of a Mnl1 restriction site, which results from the deletion, was used to establish the genotype of the rest of the kindred. In summary, we report on the genomic organization of the human ABC1 gene and identify a frameshift mutation in the ABC1 gene of the index case of Tangier disease. These results will be useful in the future characterization of the structure and function of the ABC1 gene and the analysis of additional ABC1 mutations in patients with Tangier disease. PMID- 10535984 TI - Activation changes the spectrum but not the diversity of genes expressed by T cells. AB - During activation T cells are thought to change their patterns of gene expression dramatically. To find out whether this is true for T cells activated in animals, the patterns of genes expressed in resting T cells and T cells 8 and 48 hr after activation were examined by using Affymetrix gene arrays. Gene arrays gave accurate comparisons of gene expression in the different cell types because the expression of genes known to vary during activation changed as expected. Of the approximately 6,300 genes assessed by the arrays, about one-third were expressed to appreciable extents in any of the T cells tested. Thus, resting T cells express a surprisingly large diversity of genes. The patterns of gene expression changed considerably within 8 hr of T cell activation but returned to a disposition more like that of resting T cells within 48 hr of exposure to antigen. Not unexpectedly, the activated T cells expressed genes associated with cell division at higher levels than resting T cells. The resting T cells expressed a number of cytokine receptor genes and some genes thought to suppress cell division, suggesting that the state of resting T cells is not a passive failure to respond to extant external stimuli. PMID- 10535985 TI - BAC-VAC, a novel generation of (DNA) vaccines: A bacterial artificial chromosome (BAC) containing a replication-competent, packaging-defective virus genome induces protective immunity against herpes simplex virus 1. AB - This study aimed to exploit bacterial artificial chromosomes (BAC) as large antigen-capacity DNA vaccines (BAC-VAC) against complex pathogens, such as herpes simplex virus 1 (HSV-1). The 152-kbp HSV-1 genome recently has been cloned as an F-plasmid-based BAC in Escherichia coli (fHSV), which can efficiently produce infectious virus progeny upon transfection into mammalian cells. A safe modification of fHSV, fHSVDeltapac, does not give rise to progeny virus because the signals necessary to package DNA into virions have been excluded. However, in mammalian cells fHSVDeltapac DNA can still replicate, express the HSV-1 genes, cause cytotoxic effects, and produce virus-like particles. Because these functions mimic the lytic cycle of the HSV-1 infection, fHSVDeltapac was expected to stimulate the immune system as efficiently as a modified live virus vaccine. To test this hypothesis, mice were immunized with fHSVDeltapac DNA applied intradermally by gold-particle bombardment, and the immune responses were compared with those induced by infection with disabled infectious single cycle HSV-1. Immunization with either fHSVDeltapac or disabled infectious single cycle HSV-1 induced the priming of HSV-1-specific cytotoxic T cells and the production of virus-specific antibodies and conferred protection against intracerebral injection of wild-type HSV-1 at a dose of 200 LD(50). Protection probably was cell-mediated, as transfer of serum from immunized mice did not protect naive animals. We conclude that BAC-VACs per se, or in combination with genetic elements that support replicative amplification of the DNA in the cell nucleus, represent a useful new generation of DNA-based vaccination strategies for many viral and nonviral antigens. PMID- 10535986 TI - CD8 T cell ignorance or tolerance to islet antigens depends on antigen dose. AB - There are two major mechanisms reported to prevent the autoreactivity of islet specific CD8(+) T cells: ignorance and tolerance. When ignorance is operative, naive autoreactive CD8(+) T cells ignore islet antigens and recirculate without causing damage, unless activated by an external stimulus. In the case of tolerance, CD8(+) T cells are deleted. Which factor(s) contributes to each particular outcome was previously unknown. Here, we demonstrate that the concentration of self antigen determines which mechanism operates. When ovalbumin (OVA) was expressed at a relatively low concentration in the pancreatic islets of transgenic mice, there was no detectable cross-presentation, and the CD8(+) T cell compartment remained ignorant of OVA. In mice expressing higher doses of OVA, cross-presentation was detectable and led to peripheral deletion of OVA specific CD8(+) T cells. When cross-presentation was prevented by reconstituting the bone marrow compartment with cells incapable of presenting OVA, deletional tolerance was converted to ignorance. Thus, the immune system uses two strategies to avoid CD8(+) T cell-mediated autoimmunity: for high dose antigens, it deletes autoreactive T cells, whereas for lower dose antigens, it relies on ignorance. PMID- 10535987 TI - Humoral response to herpes simplex virus is complement-dependent. AB - The complement system represents a cascade of serum proteins, which provide a major effector function in innate immunity. Recent studies have revealed that complement links innate and adaptive immunity via complement receptors CD21/CD35 in that it enhances the B cell memory response to noninfectious protein antigens introduced i.v. To examine the importance of complement for immune responses to virus infection in a peripheral tissue, we compared the B cell memory response of mice deficient in complement C3, C4, or CD21/CD35 with wild-type controls. We found that the deficient mice failed to generate a normal memory response, which is characterized by a reduction in IgG antibody and germinal centers. Thus, complement is important not only in the effector function of innate immunity but also in the stimulation of memory B cell responses to viral-infected cell antigens in both blood and peripheral tissues. PMID- 10535988 TI - RAG2 is regulated differentially in B and T cells by elements 5' of the promoter. AB - To study RAG2 gene regulation in vivo, we developed a blastocyst complementation method in which RAG2-deficient embryonic stem cells were transfected with genomic clones containing RAG2 and then assessed for their ability to generate lymphocytes. A RAG2 genomic clone that contained only the RAG2 promoter sequences rescued V(D)J recombination in RAG2-deficient pro-B cell lines, but did not rescue development of RAG2-deficient lymphocytes in vivo. However, inclusion of varying lengths of sequences 5' of the RAG2 promoter generated constructs capable of rescuing only in vivo B cell development, as well as other constructs that rescued both B and T cell development. In particular, the 2-kb 5' region starting just upstream of the RAG2 promoter, as well as the region from 2-7 kb 5', could independently drive B cell development, but not efficient T cell development. Deletion of the 2-kb 5' region from the murine germ line demonstrated that this region was not required for RAG expression sufficient to generate normal B or T cell numbers, implying redundancy among 5' elements. We conclude that RAG2 expression in vivo requires elements beyond the core promoter, that such elements contribute to differential regulation in the B vs. T lineages, and that sequences sufficient to direct B cell expression are located in the promoter-proximal 5' region. PMID- 10535990 TI - Coexpression of factor VIII heavy and light chain adeno-associated viral vectors produces biologically active protein. AB - We are interested in using recombinant adeno-associated viral vectors in the treatment of hemophilia A. Because of the size constraints of recombinant adeno associated viral vectors, we delivered the heavy and light chains of the human factor 8 (hFVIII) cDNA independently by using two separate vectors. Recombinant AAV vectors were constructed that utilized the human elongation factor 1alpha promoter, a human growth factor polyadenylation signal, and the cDNA sequences encoding either the heavy or light chain of hFVIII. Portal vein injections of each vector alone, a combination of both vectors, or a hFIX control vector were performed in C57BL/6 mice. An ELISA specific for the light chain of hFVIII demonstrated very high levels (2-10 microgram/ml) of protein expression in animals injected with the light chain vector alone or with both vectors. We utilized a chromogenic assay in combination with an antibody specific to hFVIII to determine the amount of biologically active hFVIII in mouse plasma. In animals injected with both the heavy and light chain vectors, greater than physiological levels (200-400 ng/ml) of biologically active hFVIII were produced. This suggests that coexpression of the heavy and light chains of hFVIII may be a feasible approach for treatment of hemophilia A. PMID- 10535989 TI - In vivo inhibition of rat stellate cell activation by soluble transforming growth factor beta type II receptor: a potential new therapy for hepatic fibrosis. AB - Transforming growth factor beta (TGF-beta) is a well characterized cytokine that appears to play a major role in directing the cellular response to injury, driving fibrogenesis, and, thus, potentially underlying the progression of chronic injury to fibrosis. In this study, we report the use of a novel TGF-beta receptor antagonist to block fibrogenesis induced by ligation of the common bile duct in rats. The antagonist consisted of a chimeric IgG containing the extracellular portion of the TGF-beta type II receptor. This "soluble receptor" was infused at the time of injury; in some experiments it was given at 4 days after injury, as a test of its ability to reverse fibrogenesis. The latter was assessed by expression of collagen, both as the mRNA in stellate cells isolated from control or injured liver and also by quantitative histochemistry of tissue sections. When the soluble receptor was administered at the time of injury, collagen I mRNA in stellate cells from the injured liver was 26% of that from animals receiving control IgG (P < 0.0002); when soluble receptor was given after injury induction, collagen I expression was 35% of that in control stellate cells (P < 0.0001). By quantitative histochemistry, hepatic fibrosis in treated animals was 55% of that in controls. We conclude that soluble TGF-beta receptor is an effective inhibitor of experimental fibrogenesis in vivo and merits clinical evaluation as a novel agent for controlling hepatic fibrosis in chronic liver injury. PMID- 10535991 TI - Nrf2 is essential for protection against acute pulmonary injury in mice. AB - Nrf2 is a member of the "cap 'n' collar" family of transcription factors. These transcription factors bind to the NF-E2 binding sites (GCTGAGTCA) that are essential for the regulation of erythroid-specific genes. Nrf2 is expressed in a wide range of tissues, many of which are sites of expression for phase 2 detoxification genes. Nrf2(-/-) mice are viable and have a normal phenotype under normal laboratory conditions. The NF-E2 binding site is a subset of the antioxidant response elements that have the sequence GCNNNGTCA. The antioxidant response elements are regulatory sequences found on promoters of several phase 2 detoxification genes that are inducible by xenobiotics and antioxidants. We report here that Nrf2(-/-) mice are extremely susceptible to the administration of the antioxidant butylated hydroxytoluene. With doses of butylated hydroxytoluene that are tolerated by wild-type mice, the Nrf2(-/-) mice succumb from acute respiratory distress syndrome. Gene expression studies show that the expression of several detoxification enzymes is altered in the Nrf2(-/-) mice. The Nrf2(-/-) mice may prove to be a good in vivo model for toxicological studies. As oxidative damage causes DNA breakage, these mice may also be useful for testing carcinogenic agents. PMID- 10535992 TI - Sterol regulatory element binding protein-1c is a major mediator of insulin action on the hepatic expression of glucokinase and lipogenesis-related genes. AB - Hepatic glucokinase plays a key role in glucose metabolism as underlined by the anomalies associated with glucokinase mutations and the consequences of tissue specific knock-out. In the liver, glucokinase transcription is absolutely dependent on the presence of insulin. The cis-elements and trans-acting factors that mediate the insulin effect are presently unknown; this is also the case for most insulin-responsive genes. We have shown previously that the hepatic expression of the transcription factor sterol regulatory element binding protein 1c (SREBP-1c) is activated by insulin. We show here in primary cultures of hepatocytes that the adenovirus-mediated transduction of a dominant negative form of SREBP-1c inhibits the insulin effect on endogenous glucokinase expression. Conversely, in the absence of insulin, the adenovirus-mediated transduction of a dominant positive form of SREBP-1c overcomes the insulin dependency of glucokinase expression. Hepatic fatty acid synthase and Spot-14 are insulin/glucose-dependent genes. For this latter class of genes, the dominant positive form of SREBP-1c obviates the necessity for the presence of insulin, whereas glucose potentiates the effect of SREBP-1c on their expression. In addition, the insulin dependency of lipid accumulation in cultured hepatocytes is overcome by the dominant positive form of SREBP-1c. We propose that SREBP-1c is a major mediator of insulin action on hepatic gene expression and a key regulator of hepatic glucose/lipid metabolism. PMID- 10535993 TI - Plasmodium falciparum domain mediating adhesion to chondroitin sulfate A: a receptor for human placental infection. AB - Malaria during the first pregnancy causes a high rate of fetal and neonatal death. The decreasing susceptibility during subsequent pregnancies correlates with acquisition of antibodies that block binding of infected red cells to chondroitin sulfate A (CSA), a receptor for parasites in the placenta. Here we identify a domain within a particular Plasmodium falciparum erythrocyte membrane protein 1 that binds CSA. We cloned a var gene expressed in CSA-binding parasitized red blood cells (PRBCs). The gene had eight receptor-like domains, each of which was expressed on the surface of Chinese hamster ovary cells and was tested for CSA binding. CSA linked to biotin used as a probe demonstrated that two Duffy-binding-like (DBL) domains (DBL3 and DBL7) bound CSA. DBL7, but not DBL3, also bound chondroitin sulfate C (CSC) linked to biotin, a negatively charged sugar that does not support PRBC adhesion. Furthermore, CSA, but not CSC, blocked the interaction with DBL3; both CSA and CSC blocked binding to DBL7. Thus, only the DBL3 domain displays the same binding specificity as PRBCs. Because protective antibodies present after pregnancy block binding to CSA of parasites from different parts of the world, DBL-3, although variant, may induce cross-reactive immunity that will protect pregnant women and their fetuses. PMID- 10535994 TI - Dose-response resistance to HIV-1/MuLV pseudotype virus ex vivo in a hairpin ribozyme transgenic mouse model. AB - We have investigated the efficacy of a hairpin ribozyme targeting the 5' leader sequence of HIV-1 RNA in a transgenic model system. Primary spleen cells derived from transgenic or control mice were infected with HIV-1/MuLV pseudotype virus. A significantly reduced susceptibility to infection in ribozyme-expressing transgenic spleen cells (P = 0.01) was shown. Variation of transgene-expression levels between littermates revealed a dose response between ribozyme expression and viral resistance, with an estimated cut off value below 0.2 copies of hairpin ribozyme per cell. These findings open up possibilities for studies on ribozyme efficacy and anti-HIV-1 gene therapy. PMID- 10535995 TI - In situ detection of the hypermethylation-induced inactivation of the p16 gene as an early event in oncogenesis. AB - We have developed a technique, methylation-specific PCR in situ hybridization (MSP-ISH), which allows for the methylation status of specific DNA sequences to be visualized in individual cells. We use MSP-ISH to monitor the timing and consequences of aberrant hypermethylation of the p16 tumor suppresser gene during the progression of cancers of the lung and cervix. Hypermethylation of p16 was localized only to the neoplastic cells in both in situ lesions and invasive cancers, and was associated with loss of p16 protein expression. MSP-ISH allowed us to dissect the surprising finding that p16 hypermethylation occurs in cervical carcinoma. This tumor is associated with infection of the oncogenic human papillomavirus, which expresses a protein, E7, that inactivates the retinoblastoma (Rb) protein. Thus, simultaneous Rb and p16 inactivation would not be needed to abrogate the critical cyclin D-Rb pathway. MSP-ISH reveals that p16 hypermethylation occurs heterogeneously within early cervical tumor cell populations that are separate from those expressing viral E7 transcripts. In advanced cervical cancers, the majority of cells have a hypermethylated p16, lack p16 protein, but no longer express E7. These data suggest that p16 inactivation is selected as the most effective mechanism of blocking the cyclin D-Rb pathway during the evolution of an invasive cancer from precursor lesions. These studies demonstrate that MSP-ISH is a powerful approach for studying the dynamics of aberrant methylation of critical tumor suppressor genes during tumor evolution. PMID- 10535996 TI - A mechanism of paraquat toxicity involving nitric oxide synthase. AB - Paraquat (PQ) is a well described pneumotoxicant that produces toxicity by redox cycling with cellular diaphorases, thereby elevating intracellular levels of superoxide (O-(2)). NO synthase (NOS) has been shown to participate in PQ-induced lung injury. Current theory holds that NO reacts with O-(2) generated by PQ to produce the toxin peroxynitrite. We asked whether NOS might alternatively function as a PQ diaphorase and reexamined the question of whether NO/O-(2) reactions were toxic or protective. Here, we show that: (i) neuronal NOS has PQ diaphorase activity that inversely correlates with NO formation; (ii) PQ-induced endothelial cell toxicity is attenuated by inhibitors of NOS that prevent NADPH oxidation, but is not attenuated by those that do not; (iii) PQ inhibits endothelium-derived, but not NO-induced, relaxations of aortic rings; and (iv) PQ induced cytotoxicity is potentiated in cytokine-activated macrophages in a manner that correlates with its ability to block NO formation. These data indicate that NOS is a PQ diaphorase and that toxicity of such redox-active compounds involves a loss of NO-related activity. PMID- 10535997 TI - Hepatitis C virus and other flaviviridae viruses enter cells via low density lipoprotein receptor. AB - Endocytosis of the Flaviviridae viruses, hepatitis C virus, GB virus C/hepatitis G virus, and bovine viral diarrheal virus (BVDV) was shown to be mediated by low density lipoprotein (LDL) receptors on cultured cells by several lines of evidence: by the demonstration that endocytosis of these virus correlated with LDL receptor activity, by complete inhibition of detectable endocytosis by anti LDL receptor antibody, by inhibition with anti-apolipoprotein E and apolipoprotein B antibodies, by chemical methods abrogating lipoprotein/LDL receptor interactions, and by inhibition with the endocytosis inhibitor phenylarsine oxide. Confirmatory evidence was provided by the lack of detectable LDL receptor on cells known to be resistant to BVDV infection. Endocytosis via the LDL receptor was shown to be mediated by complexing of the virus to very low density lipoprotein or LDL but not high density lipoprotein. Studies using LDL receptor-deficient cells or a cytolytic BVDV system indicated that the LDL receptor may be the main but not exclusive means of cell entry of these viruses. Studies on other types of viruses indicated that this mechanism may not be exclusive to Flaviviridae but may be used by viruses that associate with lipoprotein in the blood. These findings provide evidence that the family of LDL receptors may serve as viral receptors. PMID- 10535998 TI - Molecular cloning and functional analysis of SUT-1, a sulfate transporter from human high endothelial venules. AB - High endothelial venules (HEV) are specialized postcapillary venules found in lymphoid organs and chronically inflamed tissues that support high levels of lymphocyte extravasation from the blood. One of the major characteristics of HEV endothelial cells (HEVEC) is their capacity to incorporate large amounts of sulfate into sialomucin-type counter-receptors for the lymphocyte homing receptor L-selectin. Here, we show that HEVEC express two functional classes of sulfate transporters defined by their differential sensitivity to the anion-exchanger inhibitor 4,4'-diisothiocyanostilbene-2, 2'-disulfonic acid (DIDS), and we report the molecular characterization of a DIDS-resistant sulfate transporter from human HEVEC, designated SUT-1. SUT-1 belongs to the family of Na(+)-coupled anion transporters and exhibits 40-50% amino acid identity with the rat renal Na(+)/sulfate cotransporter, NaSi-1, as well as with the human and rat Na(+)/dicarboxylate cotransporters, NaDC-1/SDCT1 and NaDC-3/SDCT2. Functional expression studies in cRNA-injected Xenopus laevis oocytes showed that SUT-1 mediates high levels of Na(+)-dependent sulfate transport, which is resistant to DIDS inhibition. The SUT-1 gene mapped to human chromosome 7q33. Northern blotting analysis revealed that SUT-1 exhibits a highly restricted tissue distribution, with abundant expression in placenta. Reverse transcription-PCR analysis indicated that SUT-1 and the diastrophic dysplasia sulfate transporter (DTD), one of the two known human DIDS-sensitive sulfate transporters, are coexpressed in HEVEC. SUT-1 and DTD could correspond, respectively, to the DIDS resistant and DIDS-sensitive components of sulfate uptake in HEVEC. Together, these results demonstrate that SUT-1 is a distinct human Na(+)-coupled sulfate transporter, likely to play a major role in sulfate incorporation in HEV. PMID- 10535999 TI - Clinical relevance of the Helicobacter pylori gene for blood-group antigen binding adhesin. AB - Infection with Helicobacter pylori is associated with different human gastric diseases. Biochemical studies, in vitro adherence assays, and in vivo animal models revealed that epithelial attachment of H. pylori can be mediated by the blood-group antigen-binding adhesin (BabA) targeting human Lewis(b) surface epitopes. Studies with transgenic mice expressing the Lewis(b) epitope have shown that such attachment can alter disease outcome. In the current study, the presence of the babA2 gene encoding the adhesin was investigated in clinical isolates from a German population by using PCR and reverse transcription-PCR. A positive genotype was correlated to allelic variations in the genes encoding VacA and CagA and also to the prevalence of duodenal ulcer, distal gastric adenocarcinoma, mucosa-associated lymphoid tissue lymphoma, and antral gastritis. The presence of babA2 was significantly associated with duodenal ulcer (P = 0.0002) and adenocarcinoma (P = 0.033). In contrast, type 1 strains (vacAs1- and cagA-positive) were associated with only duodenal ulcer (P = 0.004) but not adenocarcinoma (P = 0.235). Genotype presence of babA2, vacAs1, and cagA ("triple positive" strains) showed a highly significant correlation to the prevalence of ulcer (P = 0.000002) and adenocarcinoma (P = 0.014) and discriminated significantly better between disease outcome than did the current type 1 classification. These results indicate that the babA2 gene is of high clinical relevance and would be a useful marker to identify patients who are at higher risk for specific H. pylori-related diseases. PMID- 10536000 TI - Cardioprotection from ischemia by a brief exposure to physiological levels of ethanol: role of epsilon protein kinase C. AB - Recent epidemiological studies indicate beneficial effects of moderate ethanol consumption in ischemic heart disease. Most studies, however, focus on the effect of long-term consumption of ethanol. In this study, we determined whether brief exposure to ethanol immediately before ischemia also produces cardioprotection. In addition, because protein kinase C (PKC) has been shown to mediate protection of the heart from ischemia, we determined the role of specific PKC isozymes in ethanol-induced protection. We demonstrated that (i) brief exposure of isolated adult rat cardiac myocytes to 10-50 mM ethanol protected against damage induced by prolonged ischemia; (ii) an isozyme-selective epsilonPKC inhibitor developed in our laboratory inhibited the cardioprotective effect of acute ethanol exposure; (iii) protection of isolated intact adult rat heart also occurred after incubation with 10 mM ethanol 20 min before global ischemia; and (iv) ethanol induced cardioprotection depended on PKC activation because it was blocked by chelerythrine and GF109203X, two PKC inhibitors. Consumption of 1-2 alcoholic beverages in humans leads to blood alcohol levels of approximately 10 mM. Therefore, our work demonstrates that exposure to physiologically attainable ethanol levels minutes before ischemia provides cardioprotection that is mediated by direct activation of epsilonPKC in the cardiac myocytes. The potential clinical implications of our findings are discussed. PMID- 10536001 TI - Steroid disorders in children: congenital adrenal hyperplasia and apparent mineralocorticoid excess. AB - Our research team and laboratories have concentrated on two inherited endocrine disorders, congenital adrenal hyperplasia (CAH) and apparent mineralocorticoid excess, in thier investigations of the pathophysiology of adrenal steroid hormone disorders in children. CAH refers to a family of inherited disorders in which defects occur in one of the enzymatic steps required to synthesize cortisol from cholesterol in the adrenal gland. Because of the impaired cortisol secretion, adrenocorticotropic hormone levels rise due to impairment of a negative feedback system, which results in hyperplasia of the adrenal cortex. The majority of cases is due to 21-hydroxylase deficiency (21-OHD). Owing to the blocked enzymatic step, cortisol precursors accumulate in excess and are converted to potent androgens, which are secreted and cause in utero virilization of the affected female fetus genitalia in the classical form of CAH. A mild form of the 21-OHD, termed nonclassical 21-OHD, is the most common autosomal recessive disorder in humans, and occurs in 1/27 Ashkenazic Jews. Mutations in the CYP21 gene have been identified that cause both classical and nonclassical CAH. Apparent mineralocorticoid excess is a potentially fatal genetic disorder causing severe juvenile hypertension, pre- and postnatal growth failure, and low to undetectable levels of potassium, renin, and aldosterone. It is caused by autosomal recessive mutations in the HSD11B2 gene, which result in a deficiency of 11beta hydroxysteroid dehydrogenase type 2. In 1998, we reported a mild form of this disease, which may represent an important cause of low-renin hypertension. PMID- 10536002 TI - Sustained in vivo cardiac protection by a rationally designed peptide that causes epsilon protein kinase C translocation. AB - Brief periods of cardiac ischemia trigger protection from subsequent prolonged ischemia (preconditioning). epsilon Protein kinase C (epsilonPKC) has been suggested to mediate preconditioning. Here, we describe an epsilonPKC-selective agonist octapeptide, psiepsilon receptor for activated C-kinase (psiepsilonRACK), derived from an epsilonPKC sequence homologous to its anchoring protein, epsilonRACK. Introduction of psiepsilonRACK into isolated cardiomyocytes, or its postnatal expression as a transgene in mouse hearts, increased epsilonPKC translocation and caused cardio-protection from ischemia without any deleterious effects. Our data demonstrate that epsilonPKC activation is required for protection from ischemic insult and suggest that small molecules that mimic this epsilonPKC agonist octapeptide provide a powerful therapeutic approach to protect hearts at risk for ischemia. PMID- 10536003 TI - Autocrine production and action of IL-3 and granulocyte colony-stimulating factor in chronic myeloid leukemia. AB - Primitive subsets of leukemic cells isolated by using fluorescence-activated cell sorting from patients with newly diagnosed Ph(+)/BCR-ABL(+) chronic myeloid leukemia display an abnormal ability to proliferate in vitro in the absence of added growth factors. We now show from analyses of growth-factor gene expression, protein production, and antibody inhibition studies that this deregulated growth can be explained, at least in part, by a novel differentiation-controlled autocrine mechanism. This mechanism involves the consistent and selective activation of IL-3 and granulocyte colony-stimulating factor (G-CSF) production and a stimulation of STAT5 phosphorylation in CD34(+) leukemic cells. When these cells differentiate into CD34(-) cells in vivo, IL-3 and G-CSF production declines, and the cells concomitantly lose their capacity for autonomous growth in vitro despite their continued expression of BCR-ABL. Based on previous studies of normal cells, excessive exposure of the most primitive chronic myeloid leukemia cells to IL-3 and G-CSF through an autocrine mechanism could explain their paradoxically decreased self-renewal in vitro and slow accumulation in vivo, in spite of an increased cycling activity and selective expansion of later compartments. PMID- 10536004 TI - Polymorphisms in the methylenetetrahydrofolate reductase gene are associated with susceptibility to acute leukemia in adults. AB - Reduction of 5,10-methylenetetrahydrofolate (methyleneTHF), a donor for methylating dUMP to dTMP in DNA synthesis, to 5-methyltetrahydrofolate (methylTHF), the primary methyl donor for methionine synthesis, is catalyzed by 5,10-methylenetetrahydrofolate reductase (MTHFR). A common 677 C --> T polymorphism in the MTHFR gene results in thermolability and reduced MTHFR activity that decreases the pool of methylTHF and increases the pool of methyleneTHF. Recently, another polymorphism in MTHFR (1298 A --> C) has been identified that also results in diminished enzyme activity. We tested whether carriers of these variant alleles are protected from adult acute leukemia. We analyzed DNA from a case-control study in the United Kingdom of 308 adult acute leukemia patients and 491 age- and sex-matched controls. MTHFR variant alleles were determined by a PCR-restriction fragment length polymorphism assay. The MTHFR 677TT genotype was lower among 71 acute lymphocytic leukemia (ALL) cases compared with 114 controls, conferring a 4.3-fold decrease in risk of ALL [odds ratio (OR = 0.23; 95% CI = 0.06-0.81]. We observed a 3-fold reduction in risk of ALL in individuals with the MTHFR 1298AC polymorphism (OR = 0.33; 95% CI = 0.15 0.73) and a 14-fold decreased risk of ALL in those with the MTHFR 1298CC variant allele (OR = 0.07; 95% CI = 0.00-1.77). In acute myeloid leukemia, no significant difference in MTHFR 677 and 1298 genotype frequencies was observed between 237 cases and 377 controls. Individuals with the MTHFR 677TT, 1298AC, and 1298CC genotypes have a decreased risk of adult ALL, but not acute myeloid leukemia, which suggests that folate inadequacy may play a key role in the development of ALL. PMID- 10536005 TI - Administration of helper-dependent adenoviral vectors and sequential delivery of different vector serotype for long-term liver-directed gene transfer in baboons. AB - The efficiency of first-generation adenoviral vectors as gene delivery tools is often limited by the short duration of transgene expression, which can be related to immune responses and to toxic effects of viral proteins. In addition, readministration is usually ineffective unless the animals are immunocompromised or a different adenovirus serotype is used. Recently, adenoviral vectors devoid of all viral coding sequences (helper-dependent or gutless vectors) have been developed to avoid expression of viral proteins. In mice, liver-directed gene transfer with AdSTK109, a helper-dependent adenoviral (Ad) vector containing the human alpha(1)-antitrypsin (hAAT) gene, resulted in sustained expression for longer than 10 months with negligible toxicity to the liver. In the present report, we have examined the duration of expression of AdSTK109 in the liver of baboons and compared it to first-generation vectors expressing hAAT. Transgene expression was limited to approximately 3-5 months with the first-generation vectors. In contrast, administration of AdSTK109 resulted in transgene expression for longer than a year in two of three baboons. We have also investigated the feasibility of circumventing the humoral response to the virus by sequential administration of vectors of different serotypes. We found that the ineffectiveness of readministration due to the humoral response to an Ad5 first generation vector was overcome by use of an Ad2-based vector expressing hAAT. These data suggest that long-term expression of transgenes should be possible by combining the reduced immunogenicity and toxicity of helper-dependent vectors with sequential delivery of vectors of different serotypes. PMID- 10536006 TI - The inv(16) encodes an acute myeloid leukemia 1 transcriptional corepressor. AB - The inv(16) is one of the most frequent chromosomal translocations associated with acute myeloid leukemia (AML). The inv(16) fusion protein acts by dominantly interfering with AML-1/core binding factor beta-dependent transcriptional regulation. Here we demonstrate that the inv(16) fusion protein cooperates with AML-1B to repress transcription. This cooperativity requires the ability of the translocation fusion protein to bind to AML-1B. Mutational analysis and cell fractionation experiments indicated that the inv(16) fusion protein acts in the nucleus and that repression occurs when the complex is bound to DNA. We also found that the inv(16) fusion protein binds to AML-1B when it is associated with the mSin3A corepressor. An AML-1B mutant that fails to bind mSin3A was impaired in cooperative repression, suggesting that the inv(16) fusion protein acts through mSin3 and possibly other corepressors. Finally, we demonstrate that the C terminal portion of the inv(16) fusion protein contains a repression domain, suggesting a molecular mechanism for AML-1-mediated repression. PMID- 10536007 TI - A conserved residue in the tip proteins of CS1 and CFA/I pili of enterotoxigenic Escherichia coli that is essential for adherence. AB - Enterotoxigenic Escherichia coli associated with human diarrheal disease utilize any of a limited group of serologically distinguishable pili for attachment to intestinal cells. These include CS1 and CFA/I pili. We show here that chemical modification of arginyl residues in CS1 pili abolishes CS1-mediated agglutination of bovine erythrocytes, which serves as a model system for attachment. Alanine substitution of the single arginyl residue in CooA, the major pilin, had no effect on the assembly of pili or on hemagglutination. In contrast, substitution of alanine for R181 in CooD, the minor pilin associated with the pilus tip, abolished hemagglutination, and substitution of R20 reduced hemagglutination. Neither of these substitutions affected CS1 pilus assembly. This shows that CooD is essential for CS1-mediated attachment and identifies specific residues that are involved in receptor binding but not in pilus assembly. In addition to mediating agglutination of bovine erythrocytes, CFA/I also mediates agglutination of human erythrocytes. Substitution of R181 by alanine in the CooD homolog, CfaE, abolished both of these reactions. We conclude that the same region of the pilus tip protein is involved in adherence of CS1 and CFA/I pili, although their receptor specificities differ. This suggests that the region of the pilus tip adhesin protein that includes R181 might be an appropriate target for therapeutic intervention or for a vaccine to protect against human diarrhea caused by enterotoxigenic E. coli strains that have serologically different pili. PMID- 10536008 TI - Exploring drug-induced alterations in gene expression in Mycobacterium tuberculosis by microarray hybridization. AB - Tuberculosis is a chronic infectious disease that is transmitted by cough propelled droplets that carry the etiologic bacterium, Mycobacterium tuberculosis. Although currently available drugs kill most isolates of M. tuberculosis, strains resistant to each of these have emerged, and multiply resistant strains are increasingly widespread. The growing problem of drug resistance combined with a global incidence of seven million new cases per year underscore the urgent need for new antituberculosis therapies. The recent publication of the complete sequence of the M. tuberculosis genome has made possible, for the first time, a comprehensive genomic approach to the biology of this organism and to the drug discovery process. We used a DNA microarray containing 97% of the ORFs predicted from this sequence to monitor changes in M. tuberculosis gene expression in response to the antituberculous drug isoniazid. Here we show that isoniazid induced several genes that encode proteins physiologically relevant to the drug's mode of action, including an operonic cluster of five genes encoding type II fatty acid synthase enzymes and fbpC, which encodes trehalose dimycolyl transferase. Other genes, not apparently within directly affected biosynthetic pathways, also were induced. These genes, efpA, fadE23, fadE24, and ahpC, likely mediate processes that are linked to the toxic consequences of the drug. Insights gained from this approach may define new drug targets and suggest new methods for identifying compounds that inhibit those targets. PMID- 10536009 TI - Reciprocal secretion of proteins by the bacterial type III machines of plant and animal pathogens suggests universal recognition of mRNA targeting signals. AB - Bacterial pathogens of both animals and plants use type III secretion machines to inject virulence proteins into host cells. Although many components of the secretion machinery are conserved among different bacterial species, the substrates for their type III pathways are not. The Yersinia type III machinery recognizes some secretion substrates via a signal that is encoded within the first 15 codons of yop mRNA. These signals can be altered by frameshift mutations without affecting secretion of the encoded polypeptides, suggesting a mechanism whereby translation of yop mRNA is coupled to the translocation of newly synthesized polypeptide. We report that the type III machinery of Erwinia chrysanthemi cloned in Escherichia coli recognizes the secretion signals of yopE and yopQ. Pseudomonas syringae AvrB and AvrPto, two proteins exported by the recombinant Erwinia machine, can also be secreted by the Yersinia type III pathway. Mapping AvrPto sequences sufficient for the secretion of reporter fusions in Yersinia revealed the presence of an mRNA secretion signal. We propose that 11 conserved components of type III secretion machines may recognize signals that couple mRNA translation to polypeptide secretion. PMID- 10536010 TI - Attenuation of virulence in Mycobacterium tuberculosis expressing a constitutively active iron repressor. AB - Iron is an essential nutrient for the survival of most organisms and has played a central role in the virulence of many infectious disease pathogens. Mycobacterial IdeR is an iron-dependent repressor that shows 80% identity in the functional domains with its corynebacterial homologue, DtxR (diphtheria toxin repressor). We have transformed Mycobacterium tuberculosis with a vector expressing an iron independent, positive dominant, corynebacterial dtxR hyperrepressor, DtxR(E175K). Western blots of whole-cell lysates of M. tuberculosis expressing the dtxR(E175K) gene revealed the stable expression of the mutant protein in mycobacteria. BALB/c mice were infected by tail vein injection with 2 x 10(5) organisms of wild type or M. tuberculosis transformed with the dtxR mutant. At 16 weeks, there was a 1.2 log reduction in bacterial survivors in both spleen (P = 0.0002) and lungs (P = 0.006) with M. tuberculosis DtxR(E175K). A phenotypic difference in colonial morphology between the two strains also was noted. A computerized search of the M. tuberculosis genome for the palindromic consensus sequence to which DtxR and IdeR bind revealed six putative "iron boxes" within 200 bp of an ORF. Using a gel shift assay we showed that purified DtxR binds to the operator region of five of these boxes. Attenuation of M. tuberculosis can be achieved by the insertion of a plasmid containing a constitutively active, iron-insensitive repressor, DtxR(E175K), which is a homologue of IdeR. Our results strongly suggest that IdeR controls genes essential for virulence in M. tuberculosis. PMID- 10536011 TI - Adeno-associated virus (AAV) site-specific integration: formation of AAV-AAVS1 junctions in an in vitro system. AB - An in vitro system to study the mechanism of site-specific integration of adeno associated virus (AAV) was developed. This system is based on two substrates, a linear or circular AAV donor and a circular acceptor containing the preintegration locus AAVS1. In the presence of HeLa extract and the His-Tag purified Rep68 protein, specific covalent junctions between AAV and AAVS1 were formed and detected by PCR. The majority of the junctions were located within the Rep binding site of both the AAV and the AAVS1 substrates, underlining the involvement of the Rep protein. A limited amount of replication and the presence of nuclear factors promoted the efficiency of the reaction. The process was ATP dependent, indicating that the helicase activity of Rep may be important in the formation of the junctions. According to current models of integration, the formation of the junctions would represent a first step in the process of AAV integration. This step could be crucial for the site specificity of the recombination event that leads to the integration of AAV into human chromosome 19 in vivo. PMID- 10536012 TI - The neuroprotective agent SR 57746A abrogates experimental autoimmune encephalomyelitis and impairs associated blood-brain barrier disruption: implications for multiple sclerosis treatment. AB - Experimental autoimmune encephalomyelitis (EAE) is a T cell autoimmune disorder that is a widely used animal model for multiple sclerosis (MS) and, as in MS, clinical signs of EAE are associated with blood-brain barrier (BBB) disruption. SR 57746A, a nonpeptide drug without classical immunosuppressive properties, efficiently protected the BBB and impaired intrathecal IgG synthesis (two conventional markers of MS exacerbation) and consequently suppressed EAE clinical signs. This compound inhibited EAE-induced spinal cord mononuclear cell invasion and normalized tumor necrosis factor alpha and IFN-gamma mRNA expression within the spinal cord. These data suggested that pharmacological intervention aimed at inhibiting proinflammatory cytokine expression within the central nervous system provided protection against BBB disruption, the first clinical sign of EAE and probably the key point of acute MS attacks. This finding could lead to the development of a new class of compounds for oral therapy of MS, as a supplement to immunosuppressive agents. PMID- 10536013 TI - Postsynaptic clustering of gamma-aminobutyric acid type A receptors by the gamma3 subunit in vivo. AB - Synaptic localization of gamma-aminobutyric acid type A (GABA(A)) receptors is a prerequisite for synaptic inhibitory function, but the mechanism by which different receptor subtypes are localized to postsynaptic sites is poorly understood. The gamma2 subunit and the postsynaptic clustering protein gephyrin are required for synaptic localization and function of major GABA(A) receptor subtypes. We now show that transgenic overexpression of the gamma3 subunit in gamma2 subunit-deficient mice restores benzodiazepine binding sites, benzodiazepine-modulated whole cell currents, and postsynaptic miniature currents, suggesting the formation of functional, postsynaptic receptors. Moreover, the gamma3 subunit can substitute for gamma2 in the formation of GABA(A) receptors that are synaptically clustered and colocalized with gephyrin in vivo. These clusters were formed even in brain regions devoid of endogenous gamma3 subunit, indicating that the factors present for clustering of gamma2 subunit-containing receptors are sufficient to cluster gamma3 subunit-containing receptors. The GABA(A) receptor and gephyrin-clustering properties of the ectopic gamma3 subunit were also observed for the endogenous gamma3 subunit, but only in the absence of the gamma2 subunit, suggesting that the gamma3 subunit is at a competitive disadvantage with the gamma2 subunit for clustering of postsynaptic GABA(A) receptors in wild-type mice. PMID- 10536014 TI - MEK1 protein kinase inhibition protects against damage resulting from focal cerebral ischemia. AB - The MEK1 (MAP kinase/ERK kinase)/ERK (extracellular-signal-responsive kinase) pathway has been implicated in cell growth and differentiation [Seger, R. & Krebs, E. G. (1995) FASEB J. 9, 726-735]. Here we show that the MEK/ERK pathway is activated during focal cerebral ischemia and may play a role in inducing damage. Treatment of mice 30 min before ischemia with the MEK1-specific inhibitor PD98059 [Alessi, D. R., Cuenda, A., Cohen, P. , Dudley, D. T. & Saltiel, A. R. (1995) J. Biol. Chem. 270, 27489-27494] reduces focal infarct volume at 22 hr after ischemia by 55% after transient occlusion of the middle cerebral artery. This is accompanied by a reduction in phospho-ERK1/2 immunohistochemical staining. MEK1 inhibition also results in reduced brain damage 72 hr after ischemia, with focal infarct volume reduced by 36%. This study indicates that the MEK1/ERK pathway contributes to brain injury during focal cerebral ischemia and that PD98059, a MEK1-specific antagonist, is a potent neuroprotective agent. PMID- 10536015 TI - Evidence for a protective role of metallothionein-1 in focal cerebral ischemia. AB - Metallothioneins (MTs) are a family of metal binding proteins that have been proposed to participate in a cellular defense against zinc toxicity and free radicals. In the present study, we investigated whether increased expression of MT in MT-1 isoform-overexpressing transgenic mice (MT-TG) affords protection against mild focal cerebral ischemia and reperfusion. Transient focal ischemia was induced in control (wild type) and MT-TG mice by occluding the right middle cerebral artery for 45 min. Upon reperfusion, cerebral edema slowly developed and peaked at 24 hr as shown by T2-weighted MRI. The volume of affected tissue was on the average 42% smaller in MT-TG mice compared with control mice at 6, 9, 24, and 72 hr and 14 days postreperfusion (P < 0.01). In addition, functional studies showed that 3 weeks after reperfusion MT-TG mice showed a significantly better motor performance compared with control mice (P = 0.011). Although cortical baseline levels of MT-1 mRNA were similar in control and MT-TG mice, there was an increase in MT-1 mRNA levels in the ischemic cortex of MT-TG mice to 7.5 times baseline levels compared with an increase to 2.3 times baseline levels in control mice 24 hr after reperfusion. In addition, MT-TG mice showed an increased MT immunoreactivity in astrocytes, vascular endothelial cells, and neurons 24 hr after reperfusion whereas in control mice MT immunoreactivity was restricted mainly to astrocytes and decreased in the infarcted tissue. These results provide evidence that increased expression of MT-1 protects against focal cerebral ischemia and reperfusion. PMID- 10536016 TI - Bidirectional, experience-dependent regulation of N-methyl-D-aspartate receptor subunit composition in the rat visual cortex during postnatal development. AB - In the visual cortex, as elsewhere, N-methyl-D-aspartate receptors (NMDARs) play a critical role in triggering long-term, experience-dependent synaptic plasticity. Modifications of NMDAR subunit composition alter receptor function, and could have a large impact on the properties of synaptic plasticity. We have used immunoblot analysis to investigate the effects of age and visual experience on the expression of different NMDAR subunits in synaptoneurosomes prepared from rat visual cortices. NMDARs at birth are comprised of NR2B and NR1 subunits, and, over the first 5 postnatal weeks, there is a progressive inclusion of the NR2A subunit. Dark rearing from birth attenuates the developmental increase in NR2A. Levels of NR2A increase rapidly (in <2 hr) when dark-reared animals are exposed to light, and decrease gradually over the course of 3 to 4 days when animals are deprived of light. These data reveal that NMDAR subunit composition in the visual cortex is remarkably dynamic and bidirectionally regulated by sensory experience. We propose that NMDAR subunit regulation is a mechanism for experience-dependent modulation of synaptic plasticity in the visual cortex, and serves to maintain synaptic strength within an optimal dynamic range. PMID- 10536017 TI - Glutamate infused posttraining into the hippocampus or caudate-putamen differentially strengthens place and response learning. AB - A cross-maze task that can be acquired through either place or response learning was used to examine the hypothesis that posttraining neurochemical manipulation of the hippocampus or caudate-putamen can bias an animal toward the use of a specific memory system. Male Long-Evans rats received four trials per day for 7 days, a probe trial on day 8, further training on days 9-15, and an additional probe trial on day 16. Training occurred in a cross-maze task in which rats started from a consistent start-box (south), and obtained food from a consistent goal-arm (west). On days 4-6 of training, rats received posttraining intrahippocampal (1 microgram/0.5 microliter) or intracaudate (2 microgram/0.5 microliter) injections of either glutamate or saline (0.5 microliter). On days 8 and 16, a probe trial was given in which rats were placed in a novel start-box (north). Rats selecting the west goal-arm were designated "place" learners, and those selecting the east goal-arm were designated "response" learners. Saline treated rats predominantly displayed place learning on day 8 and response learning on day 16, indicating a shift in control of learned behavior with extended training. Rats receiving intrahippocampal injections of glutamate predominantly displayed place learning on days 8 and 16, indicating that manipulation of the hippocampus produced a blockade of the shift to response learning. Rats receiving intracaudate injections of glutamate displayed response learning on days 8 and 16, indicating an accelerated shift to response learning. The findings suggest that posttraining intracerebral glutamate infusions can (i) modulate the distinct memory processes mediated by the hippocampus and caudate putamen and (ii) bias the brain toward the use of a specific memory system to control learned behavior and thereby influence the timing of the switch from the use of cognitive memory to habit learning to guide behavior. PMID- 10536018 TI - Survival of reproductive behaviors in estrogen receptor beta gene-deficient (betaERKO) male and female mice. AB - Previously, it was shown that the lack of a functional estrogen receptor (ER) alpha gene (ERalpha) greatly affects reproduction-related behaviors in both female and male mice. However, widespread expression of a novel second ER gene, ERbeta, demanded that we examine the possible participation of ERbeta in regulation of these behaviors. In dramatic contrast to our results with ERalpha knockout (alphaERKO) males, betaERKO males performed at least as well as wild type controls in sexual behavior tests. Moreover, not only did betaERKO males exhibit normal male-typical aggressive behavior, including offensive attacks, but they also showed higher levels of aggression than wild-type mice under certain conditions of social experience. These data revealed a significant interaction between genotype and social experience with respect to aggressive behavior. Finally, females lacking a functional beta isoform of the ER gene showed normal lordosis and courtship behaviors, extending in some cases beyond the day of behavioral estrus. These results highlight the importance of ERalpha for the normal expression of natural reproductive behaviors in both sexes and also provide a background for future studies evaluating ERbeta gene contributions to other, nonreproductive behaviors. PMID- 10536020 TI - Vocal imitation in zebra finches is inversely related to model abundance. AB - A juvenile male zebra finch, Taeniopygia guttata, kept singly with its father develops a fairly complete imitation of the father's song. The imitation is less complete when other male siblings are present, possibly because as imitation commences, model abundance increases. Here we examine the consequences of allowing more or less access to a song model. Young males heard a brief song playback when they pecked at a key, but different males were allowed to hear different numbers of playbacks per day. Using an automated procedure that scored the similarity between model and pupil songs, we discovered that 40 playbacks of the song motif per day, lasting a total of 30 sec, resulted in a fairly complete imitation. More exposure led to less complete imitation. Vocal imitation often may reflect the interaction of diverse influences. Among these, we should now include the possible inhibitory effect of model overabundance, which may foster individual identity and explain the vocal diversity found in zebra finches and other songbirds. PMID- 10536019 TI - Activity-dependent regulation of synaptic clustering in a hippocampal culture system. AB - Currently, there is a limited understanding of the factors that influence the localization and density of individual synapses in the central nervous system. Here we have studied the effects of activity on synapse formation between hippocampal dentate granule cells and CA3 pyramidal neurons in culture, taking advantage of FM1-43 as a fluorescent marker of synaptic boutons. We observed an early tendency for synapses to group together, quickly followed by the appearance of synaptic clusters on dendritic processes. These events were strongly influenced by N-methyl-D-aspartic acid receptor- and cyclic AMP-dependent signaling. The microstructure and localization of the synaptic clusters resembled that found in hippocampus, at mossy fiber synapses of stratum lucidum. Activity dependent clustering of synapses represents a means for synaptic targeting that might contribute to synaptic organization in the brain. PMID- 10536021 TI - Attenuated sensitivity to neuroactive steroids in gamma-aminobutyrate type A receptor delta subunit knockout mice. AB - gamma-Aminobutyric acid (GABA) type A receptors mediate fast inhibitory synaptic transmission and have been implicated in responses to sedative/hypnotic agents (including neuroactive steroids), anxiety, and learning and memory. Using gene targeting technology, we generated a strain of mice deficient in the delta subunit of the GABA type A receptors. In vivo testing of various behavioral responses revealed a strikingly selective attenuation of responses to neuroactive steroids, but not to other modulatory drugs. Electrophysiological recordings from hippocampal slices revealed a significantly faster miniature inhibitory postsynaptic current decay time in null mice, with no change in miniature inhibitory postsynaptic current amplitude or frequency. Learning and memory assessed with fear conditioning were normal. These results begin to illuminate the novel contributions of the delta subunit to GABA pharmacology and sedative/hypnotic responses and behavior and provide insights into the physiology of neurosteroids. PMID- 10536023 TI - Mechanisms underlying kainate receptor-mediated disinhibition in the hippocampus. AB - Kainate (KA) receptor activation depresses stimulus-evoked gamma-aminobutyric acid (GABA-mediated) synaptic transmission onto CA1 pyramidal cells of the hippocampus and simultaneously increases the frequency of spontaneous GABA release through an increase in interneuronal spiking. To determine whether these two effects are independent, we examined the mechanism by which KA receptor activation depresses the stimulus-evoked, inhibitory postsynaptic current (IPSC). Bath application of the alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (AMPA)/KA receptor agonist KA in the presence of the AMPA receptor antagonist GYKI 53655 caused a large increase in spontaneous GABA release and a coincident depression of the evoked IPSC. The depressant action on the evoked IPSC was reduced, but not abolished, by the GABA(B) receptor antagonist SCH 50911, suggesting that the KA-induced increase in spontaneous GABA release depresses the evoked IPSC through activation of presynaptic GABA(B) receptors. KA had no resolvable effect on the potassium-induced increase in miniature IPSC frequency, suggesting that KA does not act through a direct effect on the release machinery or presynaptic calcium influx. KA caused a decrease in pyramidal cell input resistance, which was reduced by GABA(A) receptor antagonists. KA also caused a reduction in the size of responses to iontophoretically applied GABA, which was indistinguishable from the SCH 50911-resistant, residual depression of the evoked IPSC. These results suggest that KA receptor activation depresses the evoked IPSC indirectly by increasing interneuronal spiking and GABA release, leading to activation of presynaptic GABA(B) receptors, which depress GABA release, and postsynaptic GABA(A) receptors, which increase passive shunting. PMID- 10536022 TI - The role of the synthetic enzyme GAD65 in the control of neuronal gamma aminobutyric acid release. AB - We have studied GABAergic synaptic transmission in retinal ganglion cells and hippocampal pyramidal cells to determine, at a cellular level, what is the effect of the targeted disruption of the gene encoding the synthetic enzyme GAD65 on the synaptic release of gamma-aminobutyric acid (GABA). Neither the size nor the frequency of GABA-mediated spontaneous inhibitory postsynaptic currents (IPSCs) were reduced in retina or hippocampus in GAD65-/- mice. However, the release of GABA during sustained synaptic activation was substantially reduced. In the retina both electrical- and K(+)-induced increases in IPSC frequency were depressed without a change in IPSC amplitude. In the hippocampus the transient increase in the probability of inhibitory transmitter release associated with posttetanic potentiation was absent in the GAD65-/- mice. These results indicate that during and immediately after sustained stimulation the increase in the probability of transmitter release is not maintained in GAD65-/- mice. Such a finding suggests a decrease in the size or refilling kinetics of the releasable pool of vesicles, and various mechanisms are discussed that could account for such a defect. PMID- 10536024 TI - Compromised disease resistance in saponin-deficient plants. AB - Saponins are glycosylated plant secondary metabolites found in many major food crops [Price, K. R., Johnson, I. T. & Fenwick, G. R. (1987) CRC Crit. Rev. Food Sci. Nutr. 26, 27-133]. Because many saponins have potent antifungal properties and are present in healthy plants in high concentrations, these molecules may act as preformed chemical barriers to fungal attack. The isolation of plant mutants defective in saponin biosynthesis represents a powerful strategy for evaluating the importance of these compounds in plant defense. The oat root saponin avenacin A-1 fluoresces under ultraviolet illumination [Crombie, L., Crombie, W. M. L. & Whiting, D. A. (1986) J. Chem. Soc. Perkins 1, 1917-1922], a property that is extremely rare among saponins. Here we have exploited this fluorescence to isolate saponin-deficient (sad) mutants of a diploid oat species, Avena strigosa. These sad mutants are compromised in their resistance to a variety of fungal pathogens, and a number of lines of evidence suggest that this compromised disease resistance is a direct consequence of saponin deficiency. Because saponins are widespread throughout the plant kingdom, this group of secondary metabolites may have general significance as antimicrobial phytoprotectants. PMID- 10536025 TI - Interactions among enzymes of the Arabidopsis flavonoid biosynthetic pathway. AB - Flavonoids are secondary metabolites derived from phenylalanine and acetate metabolism that perform a variety of essential functions in higher plants. Studies over the past 30 years have supported a model in which flavonoid metabolism is catalyzed by an enzyme complex localized to the endoplasmic reticulum [Hrazdina, G. & Wagner, G. J. (1985) Arch. Biochem. Biophys. 237, 88 100]. To test this model further we assayed for direct interactions between several key flavonoid biosynthetic enzymes in developing Arabidopsis seedlings. Two-hybrid assays indicated that chalcone synthase, chalcone isomerase (CHI), and dihydroflavonol 4-reductase interact in an orientation-dependent manner. Affinity chromatography and immunoprecipitation assays further demonstrated interactions between chalcone synthase, CHI, and flavonol 3-hydroxylase in lysates from Arabidopsis seedlings. These results support the hypothesis that the flavonoid enzymes assemble as a macromolecular complex with contacts between multiple proteins. Evidence was also found for posttranslational modification of CHI. The importance of understanding the subcellular organization of elaborate enzyme systems is discussed in the context of metabolic engineering. PMID- 10536026 TI - Biosynthetic origin of conjugated double bonds: production of fatty acid components of high-value drying oils in transgenic soybean embryos. AB - Vegetable oils that contain fatty acids with conjugated double bonds, such as tung oil, are valuable drying agents in paints, varnishes, and inks. Although several reaction mechanisms have been proposed, little is known of the biosynthetic origin of conjugated double bonds in plant fatty acids. An expressed sequence tag (EST) approach was undertaken to characterize the enzymatic basis for the formation of the conjugated double bonds of alpha-eleostearic (18:3Delta(9cis, 11trans,13trans)) and alpha-parinaric (18:4Delta(9cis,11trans, 13trans,15cis)) acids. Approximately 3,000 ESTs were generated from cDNA libraries prepared from developing seeds of Momordica charantia and Impatiens balsamina, tissues that accumulate large amounts of alpha-eleostearic and alpha parinaric acids, respectively. From ESTs of both species, a class of cDNAs encoding a diverged form of the Delta(12)-oleic acid desaturase was identified. Expression of full-length cDNAs for the Momordica (MomoFadX) and Impatiens (ImpFadX) enzymes in somatic soybean embryos resulted in the accumulation of alpha-eleostearic and alpha-parinaric acids, neither of which is present in untransformed soybean embryos. alpha-Eleostearic and alpha-parinaric acids together accounted for as much as 17% (wt/wt) of the total fatty acids of embryos expressing MomoFadX. These results demonstrate the ability to produce fatty acid components of high-value drying oils in transgenic plants. These findings also demonstrate a previously uncharacterized activity for Delta(12)-oleic acid desaturase-type enzymes that we have termed "conjugase." PMID- 10536028 TI - Proteinase inhibitor-inducing activity of the prohormone prosystemin resides exclusively in the C-terminal systemin domain. AB - Prosystemin is the 200-amino acid precursor of the 18-amino acid polypeptide defense hormone, systemin. Herein, we report that prosystemin was found to be as biologically active as systemin when assayed for proteinase inhibitor induction in young tomato plants and nearly as active in the alkalinization response in Lycopersicon esculentum suspension-cultured cells. Similar to many animal prohormones that harbor multiple signals, the systemin precursor contains five imperfect repetitive domains N-terminal to a single systemin domain. Whether the five repetitive domains contain defense signals has not been established. N terminal deletions of prosystemin had little effect on its activity in tomato plants or suspension-cultured cells. Deletion of the C-terminal region of prosystemin containing the 18-amino acid systemin domain completely abolished its proteinase inhibitor induction and alkalinization activities. The apoplastic fluid from tomato leaves and the medium of cultured cells were analyzed for proteolytic activity that could process prosystemin to systemin. These experiments showed that proteolytic enzymes present in the apoplasm and medium could cleave prosystemin into large fragments, but the enzymes did not produce detectable levels of systemin. Additionally, inhibitors of these proteolytic enzymes did not affect the biological activity of prosystemin. The cumulative data indicated that prosystemin and/or large fragments of prosystemin can be active inducers of defense responses in both tomato leaves and suspension cultured cells and that the only region of prosystemin that is responsible for activating the defense response resides in the systemin domain. PMID- 10536027 TI - Genetic ablation of root cap cells in Arabidopsis. AB - The root cap is increasingly appreciated as a complex and dynamic plant organ. Root caps sense and transmit environmental signals, synthesize and secrete small molecules and macromolecules, and in some species shed metabolically active cells. However, it is not known whether root caps are essential for normal shoot and root development. We report the identification of a root cap-specific promoter and describe its use to genetically ablate root caps by directing root cap-specific expression of a diphtheria toxin A-chain gene. Transgenic toxin expressing plants are viable and have normal aerial parts but agravitropic roots, implying loss of root cap function. Several cell layers are missing from the transgenic root caps, and the remaining cells are abnormal. Although the radial organization of the roots is normal in toxin-expressing plants, the root tips have fewer cytoplasmically dense cells than do wild-type root tips, suggesting that root meristematic activity is lower in transgenic than in wild-type plants. The roots of transgenic plants have more lateral roots and these are, in turn, more highly branched than those of wild-type plants. Thus, root cap ablation alters root architecture both by inhibiting root meristematic activity and by stimulating lateral root initiation. These observations imply that the root caps contain essential components of the signaling system that determines root architecture. PMID- 10536029 TI - Invariance between subjects of brain wave representations of language. AB - In three experiments, electric brain waves of 19 subjects were recorded under several different experimental conditions for two purposes. One was to test how well we could recognize which sentence, from a set of 24 or 48 sentences, was being processed in the cortex. The other was to study the invariance of brain waves between subjects. As in our earlier work, the analysis consisted of averaging over trials to create prototypes and test samples, to both of which Fourier transforms were applied, followed by filtering and an inverse transformation to the time domain. A least-squares criterion of fit between prototypes and test samples was used for classification. In all three experiments, averaging over subjects improved the recognition rates. The most significant finding was the following. When brain waves were averaged separately for two nonoverlapping groups of subjects, one for prototypes and the other for test samples, we were able to recognize correctly 90% of the brain waves generated by 48 different sentences about European geography. PMID- 10536031 TI - Low-status monkeys "play dumb" when learning in mixed social groups. AB - Many primates, including humans, live in complex hierarchical societies where social context and status affect daily life. Nevertheless, primate learning studies typically test single animals in limited laboratory settings where the important effects of social interactions and relationships cannot be studied. To investigate the impact of sociality on associative learning, we compared the individual performances of group-tested rhesus monkeys (Macaca mulatta) across various social contexts. We used a traditional discrimination paradigm that measures an animal's ability to form associations between cues and the obtaining of food in choice situations; but we adapted the task for group testing. After training a 55-member colony to separate on command into two subgroups, composed of either high- or low-status families, we exposed animals to two color discrimination problems, one with all monkeys present (combined condition), the other in their "dominant" and "subordinate" cohorts (split condition). Next, we manipulated learning history by testing animals on the same problems, but with the social contexts reversed. Monkeys from dominant families excelled in all conditions, but subordinates performed well in the split condition only, regardless of learning history. Subordinate animals had learned the associations, but expressed their knowledge only when segregated from higher-ranking animals. Because aggressive behavior was rare, performance deficits probably reflected voluntary inhibition. This experimental evidence of rank-related, social modulation of performance calls for greater consideration of social factors when assessing learning and may also have relevance for the evaluation of human scholastic achievement. PMID- 10536030 TI - Functional neuroanatomical double dissociation of mnemonic and executive control processes contributing to working memory performance. AB - We used event-related functional MRI to investigate the neural bases of two categories of mental processes believed to contribute to performance of an alphabetization working memory task: memory storage and memory manipulation. Our delayed-response tasks required memory for the identity and position-in-the display of items in two- or five-letter memory sets (to identify load-sensitive regions) or memory for the identity and relative position-in-the-alphabet of items in five-letter memory sets (to identify manipulation-sensitive regions). Results revealed voxels in the left perisylvian cortex of five of five subjects showing load sensitivity (as contrasted with alphabetization-sensitive voxels in this region in only one subject) and voxels of dorsolateral prefrontal cortex in all subjects showing alphabetization sensitivity (as contrasted with load sensitive voxels in this region in two subjects). This double dissociation was reliable at the group level. These data are consistent with the hypothesis that the nonmnemonic executive control processes that can contribute to working memory function are primarily prefrontal cortex-mediated whereas mnemonic processes necessary for working memory storage are primarily posteriorly mediated. More broadly, they support the view that working memory is a faculty that arises from the coordinated interaction of computationally and neuroanatomically dissociable processes. PMID- 10536032 TI - Further characterization of the combining sites of Bandeiraea (Griffonia) simplicifolia lectin-I, isolectin A(4). AB - Bandeiraea (Griffonia) simplicifolia lectin-I, isolectin A(4)(GS I-A(4)), which is cytotoxic to the human colon cancer cell lines, is one of two lectin families derived from its seed extract. It contains only a homo-oligomer of subunit A, and is most specific for GalNAcalpha1-->. In order to elucidate the GS I-A(4) glycoconjugate interactions in greater detail, the combining site of this lectin was further characterized by enzyme linked lectino-sorbent assay (ELLSA) and by inhibition of lectin-glycoprotein interactions. This study has demonstrated that the Tn-containing glycoproteins tested, consisting of mammalian salivary glycoproteins (armadillo, asialo-hamster sublingual, asialo-ovine, -bovine, and porcine submandibular), are bound strongly by GS I-A(4.)Among monovalent inhibitors so far tested, p-NO2-phenylalphaGalNAc is the most potent, suggesting that hydrophobic forces are important in the interaction of this lectin. GS I A(4)is able to accommodate the monosaccharide GalNAc at the nonreducing end of oligosaccharides. This suggests that the combining site of the lectin is a shallow cavity. Among oligosaccharides and monosaccharides tested as inhibitors of the binding of GS I-A(4), the hierarchy of potencies are: GalNAcalpha1- >3GalNAcbeta1-->3Galalpha1-->4Galbeta 1-->4Glc (Forssman pentasaccharide) > GalNAcalpha1-->3(LFucalpha1-->2)Gal (blood group A)()> GalNAc > Galalpha1-->4Gal > Galalpha1-->3Gal (blood group B-like)> Gal. PMID- 10536034 TI - Secreted human conjunctival mucus contains MUC5AC glycoforms. AB - This study addresses the extent of variation in secreted end-product mucins in human conjunctival mucus. The aim was to determine whether the variety of mucin species found was encompassed by the mucin genes which have been cloned to date. Extraction into guanidine hydrochloride and separation of mucin constituents, by a combination of cesium chloride density gradient centrifugation, size separation on Sepharose CL-2B, MonoQ ion exchange chromatography and agarose gel electrophoresis, demonstrates a complex mixture of mucins. Sample size limitations precluded compositional amino acid analysis. MUC 5AC and MUC1, 2, and 4 are all detected in the buoyant density range 1.3-1.5 g/ml by antibody binding. The mucins vary in size from >40 x 10(6)to <97 x 10(3)Da. A wide range of molecular size was confirmed using rate zonal centrifugation. The presence of smaller species contrasts with other mucous secretions similarly studied. In each size range are low, medium, and high charge mucins. Sialylation predominates in the medium charge and sulfate in the high charge. Only MUC5AC cross-reactivity is maintained throughout the analysis. It is detected in large and medium sized mucins but accounts for only the least mobile mucins within copurified species of similar density, size, and charge resolved using agarose electrophoresis. MUC5AC cross-reactivity is also detected in both medium and high charge species, indicating the presence of glycoforms. PMID- 10536033 TI - Fucose in alpha(1-6)-linkage regulates proliferation and histogenesis in reaggregated retinal spheroids of the chick embryo. AB - We have used the lectin from Aleuria aurantia (AAL) which is highly specific for alpha(1-6)-linked fucose, to examine its effect on chicken retinogenesis in a reaggregation culture system. When dispersed cells of the embryonic chick retina are reaggregated to form histotypic retinospheroids, AAL elicits strong inhibition of spheroid growth. The action of AAL is specific, since its effect is dose-dependent, saturable, and inhibited by an excess of fucose. Fucosidase treatment entirely abolishes reaggregation. In contrast, Anguilla anguilla agglutinin (AAA) binding to fucose in alpha(1-2)-linkage does not show any effects. Incubation with CAB4-a specific monoclonal antibody for fucose in alpha(1-6)-linkage-reduces spheroid size and shape. AAL does not much affect primary aggregation, but rather subsequent processes of cell proliferation and histogenesis. In particular, AAL inhibits uptake of bromo-desoxyuridine (BrdU), most efficiently so during days in vitro 2 (div2) and div3. As a consequence, the histological differentiation is entirely disturbed, as evidenced by vimentin immunostaining; particularly, rosettes are not forming and the radial glia scaffold is disorganized. We conclude that glycoproteins exhibiting fucose in alpha(1-6)-linkage may play major roles in early processes of retinal tissue formation. PMID- 10536035 TI - Specialized expression of simple O-glycans along the rat kidney nephron. AB - Glycosyltransferases can exhibit tissue-specific expression. By histochemistry glycosyltransferases and their products can be localized to specific cell types in organs of complex cellular composition. We have applied the lectin Amaranthin, having a nominal specificity for Galbeta1,3GalNAcR and Neu5Ac2,3Galbeta1, 3GalNAcalpha-R, and a monoclonal antibody raised against Galbeta1, 3GalNAcalphaR to examine the distribution of these simple O-glycans in adult rat kidney. The monoclonal antibody stained ascending thin limbs of Henle, distal convoluted tubules, and collecting ducts of cortex and outer medulla. Remarkably, the ascending thick limb of Henle, located between ascending thin limb and distal convoluted tubules, was unreactive. However, Amaranthin staining was detectable in ascending thick limbs of Henle, in addition to the structures positive with the monoclonal antibody. In kidney extracts, two bands of approximately 160 kDa and >210 kDa were reactive with both Amaranthin and the monoclonal antibody. One band at approximately 200 kDa, and a smear at approximately 100 kDa, were reactive only with Amaranthin. Our data show that in rat kidney simple O-linked glycans are expressed in a highly specialized manner along the renal tubule and can be detected only on a few glycoproteins. This may reflect a cell-type specific expression of the corresponding glycosyltransferases. PMID- 10536036 TI - Sequential biosynthesis of sulfated and/or sialylated Lewis x determinants by transferases of the human bronchial mucosa. AB - The structural determination of sulfated carbohydrate chains from a cystic fibrosis patient respiratory mucins has shown that sulfation may occur either on the C-3 of the terminal Gal, or on the C-6 of the GlcNAc residue of a terminal N acetyllactosamine unit. The two enzymes responsible for the transfer of sulfate from PAPS to the C-3 of Gal or to the C-6 of GlcNAc residues have been characterized in human respiratory mucosa. These two enzymes, in conjunction with fucosyl- and sialyltransferases, allow the synthesis of different sulfated epitopes such as 3-sulfo Lewis x (with a 3- O -sulfated Gal), 6-sulfo Lewis x and 6-sulfo-sialyl Lewis x (with a 6- O -sulfated GlcNAc). In the present study, the sequential biosynthesis of these epitopes has been investigated using microsomal fractions from human respiratory mucosa incubated with radiolabeled nucleotide sugars or PAPS, and oligosaccharide acceptors, mostly prepared from human respiratory mucins. The structures of the radiolabeled products have been determined by their coelution in HPAEC with known oligosaccharidic standards. In the biosynthesis of 6- O -sulfated carbohydrate chains by the human respiratory mucosa, the 6- O -sulfation of a terminal nonreducing GlcNAc residue precedes beta1-4-galactosylation, alpha2-3-sialylation (to generate 6-sulfo-sialyl- N acetyllactosamine), and alpha1-3-fucosylation (to generate the 6-sulfo-sialyl Lewis x determinant). The 3- O -sulfation of a terminal N -acetyllactosamine may occur if this carbohydrate unit is not substituted. Once an N -acetyllactosamine unit is synthesized, alpha1-3-fucosylation of the GlcNAc residue to generate a Lewis x structure blocks any further substitution. Therefore, the present study defines the pathways for the biosynthesis of Lewis x, sialyl Lewis x, sulfo Lewis x, and 6-sulfo-sialyl Lewis x determinants in the human bronchial mucosa. PMID- 10536037 TI - Cloning and expression of a human gene encoding an N-acetylgalactosamine-alpha2,6 sialyltransferase (ST6GalNAc I): a candidate for synthesis of cancer-associated sialyl-Tn antigens. AB - The sialyl-Tn (sTn) antigen is a well known cancer-associated antigen, the expression of which is related to the prognosis of cancer patients. We aimed to isolate a human gene encoding an N -acetylgalactosamine alpha2,6 sialyltransferase which synthesizes sTn antigen, and to characterize the enzyme. Degenerate primers encoding sialyl motifs were used for the polymerase chain reaction to amplify complementary DNAs prepared from RNAs of human pyloric mucosae with intestinal metaplasia, which abundantly expressed sTn antigen, followed by screening of full-length cDNAs using the amplified DNA fragment as a probe. We isolated two human cDNA clones, long-form (2.46 kb) and short-form (2.23 kb) cDNAs. The former encodes an active enzyme with a predicted 600 amino acid sequence. The latter, a splice-variant of the long-form, encodes an inactive enzyme. HCT15 human colorectal cancer cells stably expressing the long-form cDNA expressed sTn epitopes on O -glycans. The long form cDNA was considered to encode a human homologue of chick ST6GalNAc I for the following reasons: (1) the putative amino acid sequence showed greater homology to that of chick ST6GalNAc I (55%) compared to other sialyltransferases, (2) it encodes the extraordinarily long stem region that is a typical feature of chick ST6GalNAc I, and (3) the substrate specificity was very similar to that of chick ST6GalNAc I. In situ hybridization demonstrated that the localization of transcripts correlated well with that of sTn antigen in gastric cancer cells and Goblet cells in intestinal metaplastic glands. Thus, we determined that the long-form cDNA of the human ST6GalNAc I gene encodes the probable candidate for the human sTn synthase(s). PMID- 10536038 TI - Cryptic sialic acid binding lectins on human blood leukocytes can be unmasked by sialidase treatment or cellular activation. AB - We recently reported that the sialic acid-specific binding sites of CD22 molecules on B cells are masked by endogenous ligands, and can be unmasked by sialidase treatment or cellular activation. Here, we show that many other human blood leukocyte types have endogenous sialic acid binding sites that can be unmasked by sialidase treatment. Truncation of sialic acid side chains on the soluble probes used for detection abolishes all binding, indicating the specificity of the interaction for the details of sialic acid structure. There is limited overlap between alpha2-6- and alpha2-3-sialic acid-specific binding sites, which are unmasked on monocytes, natural killer cells, a minority of mature T cells, neutrophils, and some cultured human leukemic cell lines. Activation with phorbol ester and calcium ionophore causes spontaneous exposure of some of the binding sites, occurring over a period of minutes on neutrophils and several hours on monocytes and U937 leukemia cells. Activation is accompanied by some evidence for desialylation of cell surface molecules. Thus, many human blood cells have specific binding sites for sialic acids, masked by endogenous sialylated ligands. Cellular activation can unmask these sites, possibly by the action of an endogenous sialidase. The nearly universal masking of such sites in unactivated blood cells could explain why many of these sialic acid-binding lectins have not been previously discovered. Similar considerations may apply to sialic acid binding lectins of other cell types and tissues. PMID- 10536039 TI - A structural comparison of lipopolysaccharides from two strains of Helicobacter pylori, of which one strain (442) does and the other strain (471) does not stimulate pepsinogen secretion. AB - Lipopolysaccharides (LPSs) from strains of Helicobacter pylori (442 and 471), which differed in stimulation of pepsinogen secretion, were isolated as water soluble material of high-M(r), and as water-insoluble gels of low-M(r). Chemical and spectroscopic analyses of soluble LPS and oligosaccharides liberated from the gels led to proposed structures with Lewis (Le) antigen termini connected to N acetyllacto-saminoglycans of alternating 3-linked beta-D-Gal and 4-linked beta-D GlcNAc residues with various laterally attached glycosyl substituents. The LPS of H.pylori 442 was similar to previously examined strains (NCTC 11637 and P466) in having partially glycosylated chains with alpha-L-Fuc units attached to O-3 of the majority of GlcNAc residues in Le(x)units, and in chain termination with Le(x)or Le(y)determinants. In contrast, terminal Le(y)units occurred in LPS of H.pylori 471 and glycosaminoglycan chains carried a smaller proportion of alpha-L Fuc units, but at O-6 of a majority of nonfucosylated GlcNAc residues, there was a novel type of branching with alpha-D-Gal substituents. Evidence for the branched regions was obtained from(1)H-NMR spectra and from characterization of oligosaccharides formed by the action of endo-beta-galactosidase. Examination of oligosaccharides liberated from water-insoluble LPS gels of H.pylori 442 and 471 provided evidence for similar core OS structures to those from other H.pylori strains but interesting differences were observed. PMID- 10536040 TI - Prevention of the death of the rat axotomized hypoglossal nerve and promotion of its regeneration by bovine brain gangliosides. AB - We have examined the time course of the neuronal death and regeneration of rat axotomized hypoglossal nerve with various conditions of the nerve resection, and established a useful system to measure neurotrophic activities of bioactive substances. In this system, neuronal death can be evaluated by counting surviving neurons in the nucleus of hypoglossal neuron at the brain stem, and the degree of the regeneration can be measured by counting horseradish peroxidase-positive cells at the same region after injection of horseradish peroxidase into tongue. Using this system, the effects of brain gangliosides on rat hypoglossal nerve regeneration following 5 mm transection were examined. The addition of a ganglioside mixture from bovine brain as well as the autograft strongly prevented the death of neurons and promoted the regeneration of the lesioned nerve at 10 weeks after the operation. Further analyses on the dose effects and injection sites of gangliosides were performed. Although the mechanisms of the neurotrophic effects of the gangliosides are unknown, the therapeutic application of gangliosides for neuronal degeneration is a promising approach. PMID- 10536041 TI - Lactose-containing starburst dendrimers: influence of dendrimer generation and binding-site orientation of receptors (plant/animal lectins and immunoglobulins) on binding properties. AB - Starburst glycodendrimers offer the potential to serve as high-affinity ligands for clinically relevant sugar receptors. In order to define areas of application, their binding behavior towards sugar receptors with differential binding-site orientation but identical monosaccharide specificity must be evaluated. Using poly(amidoamine) starburst dendrimers of five generations, which contain the p isothiocyanato derivative of p-aminophenyl-beta-D-lactoside as ligand group, four different types of galactoside-binding proteins were chosen for this purpose, i.e., the (AB)(2)-toxic agglutinin from mistletoe, a human immunoglobulin G fraction, the homodimeric galectin-1 with its two binding sites at opposite ends of the jelly-roll-motif-harboring protein and monomeric galectin-3. Direct solid phase assays with surface-immobilized glycodendrimers resulted in obvious affinity enhancements by progressive core branching for the plant agglutinin and less pronounced for the antibody and galectin-1. High density of binding of galectin-3 with modest affinity increases only from the level of the 32-mer onwards points to favorable protein-protein interactions of the monomeric lectin and a spherical display of the end groups without a major share of backfolding. When the inhibitory potency of these probes was evaluated as competitor of receptor binding to an immobilized neoglycoprotein or to asialofetuin, a marked selectivity was detected. The 32- and 64-mers were second to none as inhibitors for the plant agglutinin against both ligand-exposing matrices and for galectin-1 on the matrix with a heterogeneous array of interglycoside distances even on the per-sugar basis. In contrast, a neoglycoprotein with the same end group was superior in the case of the antibody and, less pronounced, monomeric galectin-3. Intimate details of topological binding-site presentation and the ligand display on different generations of core assembly are major operative factors which determine the potential of dendrimers for applications as lectin-targeting device, as attested by these observations. PMID- 10536042 TI - A recessive deletion in the GlcNAc-1-phosphotransferase gene results in peri implantation embryonic lethality. AB - Formation of the dolichol oligosaccharide precursor is essential for the production of asparagine- (N-) linked oligosaccharides (N-glycans) in eukaryotic cells. The first step in precursor biosynthesis requires the enzyme UDP-GlcNAc: dolichol phosphate N-acetylglucosamine-1-phosphate transferase (GPT). Without GPT activity, subsequent steps necessary in constructing the oligosaccharide precursor cannot occur. Inhibition of this biosynthetic step using tunicamycin, a GlcNAc analog, produces a deficiency in N-glycosylation in cell lines and embryonic lethality during preimplantation development in vitro, suggesting that N-glycan formation is essential in early embryogenesis. In exploring structure function relationships among N-glycans, and since tunicamycin has various reported biochemical activities; we have generated a germline deletion in the mouse GPT gene. GPT mutant embryos were analyzed and the phenotypes obtained were compared with previous studies using tunicamycin. We find that embryos homozygous for a deletion in the GPT gene complete preimplantation development and also implant in the uterine epithelium, but die shortly thereafter between days 4-5 postfertilization with cell degeneration apparent among both embryonic and extraembryonic cell types. Of cells derived from these early embryos, neither trophoblast nor embryonic endodermal lineages are able to survive in culture in vitro. These results indicate that GPT function is essential in early embryogenesis and suggest that N-glycosylation is needed for the viability of cells comprising the peri-implantation stage embryo. PMID- 10536043 TI - alpha1,3Fucosyltransferase VI is expressed in HepG2 cells and codistributed with beta1,4galactosyltransferase I in the golgi apparatus and monensin-induced swollen vesicles. AB - The major alpha1,3fucosyltransferase activity in plasma, liver, and kidney is related to fucosyltransferase VI which is encoded by the FUT6 gene. Here we demonstrate the presence of alpha1, 3fucosyltransferase VI (alpha3-FucT VI) in the human HepG2 hepatoma cell line by specific activity assays, detection of transcripts, and the use of specific antibodies. First, FucT activity in HepG2 cell lysates was shown to prefer sialyl-N-acetyllactosamine as acceptor substrate indicating expression of alpha3-FucT VI. RT-PCR analysis further confirmed the exclusive presence of the alpha3-FucT VI transcripts among the five human alpha3 FucTs cloned to date. alpha3-FucT VI was colocalized with beta1,4galactosyltransferase I (beta4-GalT I) to the Golgi apparatus by dual confocal immunostaining. Pulse/chase analysis of metabolically labeled alpha3 FucT VI showed maturation of alpha3-FucT VI from the early 43 kDa form to the mature, endoglycosidase H-resistant form of 47 kDa which was detected after 2 h of chase. alpha3-FucT VI was released to the medium and accounted for 50% of overall cell-associated and released enzyme activity. Release occurred by proteolytical cleavage which produced a soluble form of 43 kDa. Monensin treatment segregated alpha3-FucT VI from the Golgi apparatus to swollen peripheral vesicles where it was colocalized with beta4-GalT I while alpha2,6(N)sialyltransferase remained associated with the Golgi apparatus. Both constitutive secretion of alpha3-FucT VI and its monensin-induced relocation to vesicles analogous to beta4-GalT I suggest a similar post-Golgi pathway of both alpha3-FucT VI and beta4-GalT I. PMID- 10536044 TI - Differences in the acid-labile component of Candida albicans mannan from hydrophobic and hydrophilic yeast cells. AB - Cell surface hydrophobicity of the opportunistic fungal pathogen Candida albicans has been linked to the level of cell wall protein glycosylation. Previous work demonstrated that outer chain mannosylation, rather than overall glycosylation, correlated with cell surface hydrophobicity. These studies further suggested that the phosphodiester-linked, acid-labile beta-1,2-mannan was the correlating element. The present work tests this hypothesis and extends the previous results. The composition of bulk mannan from hydrophobic and hydrophilic yeast cells, and the acid-labile mannan from both cell types are compared. Compositional analysis shows that the protein, hexose, and phosphorus content of bulk mannan is similar between the two phenotypes. Electrophoretic separation of acid-released and fluorophore-labeled mannan shows that the acid-labile oligomannosides from hydrophobic cells are longer and potentially in greater abundance than those from hydrophilic cells. These results suggest that regulation of a single step in cell wall protein outer chain mannosylation affects the cell surface ultrastructure and phenotype of C.albicans. PMID- 10536045 TI - Proregion of Bombyx mori cysteine proteinase functions as an intramolecular chaperone to promote proper folding of the mature enzyme. AB - A cDNA encoding the proform of Bombyx cysteine proteinase (BCP) was expressed at a high level in Escherichia coli using the T7 polymerase expression system. The insoluble recombinant zymogen was solubilized and renatured by modifying a method applied to human pro-cathepsin L. Like the natural BCP precursor, the recombinant proenzyme was spontaneously converted to an active proteinase at pH 3.75. A deletion in the central region of the propeptide resulted in much loss of the activity, suggesting that the propeptide is essential for proper folding during renaturation. In contrast, the renatured mature form of recombinant BCP was not active but regained activity by including the propeptide in the renaturing buffer, suggesting that the propeptide, acting as an intramolecular chaperone, promotes refolding of the associated proteinase domain into an active conformation. The mature form of natural BCP rapidly lost its activity at neutral pH, whereas its proform was stable. The mature enzyme retained some activity in the presence of the propeptide. Arch. PMID- 10536046 TI - In vitro secretion of digestive phospholipase A(2) by midguts isolated from tobacco hornworms, manduca sexta AB - We report on secretion of phospholipase A(2) (PLA(2)) by in vitro preparations of midguts isolated from tobacco hornworms, Manduca sexta. This enzyme is responsible for hydrolysis of fatty acids from the sn-2 position of phospholipids, a necessary step in fatty acid absorption. The in vitro midgut preparations are competent to secrete PLA(2) into incubation buffer. Secretion began within the first 30 min of incubation and increased to a maximum at 8 h. We selected 2 h incubations because substantial loss of tissue integrity was observed after 8 h incubations. Using 2 h incubations, we recorded increased secretion of digestive PLA(2) from midguts incubated in buffer amended with diet or with yeast as a component of the diet. We also recorded small increases in secretion of PLA(2) from midguts incubated in buffer amended with a specific phospholipid, phosphatidylcholine. Midguts incubated in buffer amended with increased concentrations of phospholipid did not yield higher levels of PLA(2) activity. Lepidopteran midguts can be divided into three regions, and we recorded the highest secretion of PLA(2) from the middle region and lowest secretion from the anterior region. Because isolated midguts responded to food chemicals with increased secretion of digestive PLA(2), we suggest that secretion of digestive enzymes in tobacco hornworms is regulated by a prandial and/or paracrine mechanism, as suggested for digestive proteases in other insect species. Arch. Insect Biochem. Physiol. 42:179-187, 1999.Copyright 1999 Wiley-Liss, Inc. PMID- 10536047 TI - Relationships among food and contact signals in experimental group-living young of tegenaria atrica AB - Relationships between the quantity of diet, and group vs. individual rearing toward ontogenesis on degree of cannibalism and cuticular lipid profile of young Tegenaria atrica Araneae, Agelenidae were studied. Ad libitum diet increases the development rate and decreases cannibalism between conspecifics compared to a reduced diet. Individually reared young on the two diets showed no quantitative differences in cuticular lipid profile, but differences were observed in individual vs. group rearing modes. The ad libitum-fed grouped young spiders had notably increased quantities of palmitic acid, 13,17-,11,17-, and 9,17 dimethylhentriacontane, methyl oleate, and n-heptatriacontane over the reduced diet, cannibalistic group. Arch. Copyright 1999 Wiley-Liss, Inc. PMID- 10536048 TI - Developmental expression of Manduca sexta hemolin. AB - Hemolin is hemolymph protein that is a member of the immunoglobulin superfamily. Its induced expression after bacterial infection suggests that it functions in the immune response. In this paper, we describe the expression of the Manduca sexta hemolin gene at certain developmental stages in the absence of microbial challenge. Hemolin was present at a very low level in hemolymph of naive larvae until the beginning of the wandering stage prior to pupation, when its concentration in hemolymph increased dramatically. At the same time, hemolin could be found in the fluid contained in the midgut lumen. The appearance of hemolin mRNA in fat body and midgut at the beginning of the wandering stage correlated with the presence of hemolin in the hemolymph and midgut lumen. Hemolin was present in hemolymph through the pupal and adult stages. Hemolin was also present in newly deposited eggs, and persisted in eggs throughout embryonic development. A hemolin cDNA isolated from an adult fat body library had the same sequence as those previously obtained from larval libraries. Hemolin purified from hemolymph of bacteria-injected larvae, from hemolymph of naive wandering stage larvae and adult moths, and from midgut fluid of wandering stage larvae had the same apparent mass, which was consistent with the mass predicted from the hemolin cDNA sequence. Hemolin from hemolymph of wandering stage larvae did not contain any detectable carbohydrate, but hemolin from the hemolymph of bacteria injected larvae and from naive adult moths was associated with carbohydrate, although of different amounts and composition. These results suggest that a single hemolin gene is developmentally regulated and is also induced when insects are exposed to microbial infection. M. sexta hemolin apparently lacks post translational covalent glycosylation, but instead is associated under some conditions with non-covalently bound carbohydrates. Arch. PMID- 10536049 TI - Plasmatocyte spreading peptide induces spreading of plasmatocytes but represses spreading of granulocytes. AB - In most Lepidoptera, plasmatocytes and granulocytes are the two hemocyte classes capable of adhering to foreign targets. Previously, we identified plasmatocyte spreading peptide (PSP1) from the moth Pseudoplusia includens and reported that it induced plasmatocytes to rapidly spread on foreign surfaces. Here we examine whether the response of plasmatocytes to PSP1 was influenced by cell density or culture conditions, and whether PSP1 affected the adhesive state of granulocytes. Plasmatocyte spreading rates were clearly affected by cell density in the absence of PSP1 but spreading was density independent in the presence of PSP1. PSP1 also induced plasmatocytes in agarose-coated culture wells to form homotypic aggregations rather than spread on the surface of culture wells. In contrast, granulocytes rapidly spread in a density independent manner in the absence of PSP1, but were dose-dependently inhibited from spreading by the addition of peptide. An anti-PSP1 polyclonal antibody neutralized the spreading activity of synthetic PSP1. This antibody also neutralized the plasmatocyte spreading activity of granulocyte-conditioned medium, and significantly delayed plasmatocyte spreading when cells were cultured at a high density in unconditioned medium. These results suggested that the spreading activity derived from granulocytes is due in part to PSP1. Pretreatment of plasmatocytes with trypsin had no effect on PSP1-induced aggregation but PSP1-induced aggregations were readily dissociated by trypsin. This suggested that PSP1 is not an adhesion factor but induces adhesion by stimulating a change in the cell surface of plasmatocytes. Synthetic PSP1 also induced aggregation of plasmatocytes from other Lepidoptera indicating that regulation of hemocyte activity by PSP1-related peptides may be widespread. Arch. PMID- 10536051 TI - Differentiation of avian craniofacial muscles: I. Patterns of early regulatory gene expression and myosin heavy chain synthesis. AB - Myogenic populations of the avian head arise within both epithelial (somitic) and mesenchymal (unsegmented) mesodermal populations. The former, which gives rise to neck, tongue, laryngeal, and diaphragmatic muscles, show many similarities to trunk axial, body wall, and appendicular muscles. However, muscle progenitors originating within unsegmented head mesoderm exhibit several distinct features, including multiple ancestries, the absence of several somite lineage-determining regulatory gene products, diverse locations relative to neuraxial and pharyngeal tissues, and a prolonged and necessary interaction with neural crest cells. The object of this study has been to characterize the spatial and temporal patterns of early muscle regulatory gene expression and subsequent myosin heavy chain isoform appearance in avian mesenchyme-derived extraocular and branchial muscles, and compare these with expression patterns in myotome-derived neck and tongue muscles. Myf5 and myoD transcripts are detected in the dorsomedial (epaxial) region of the occipital somites before stage 12, but are not evident in the ventrolateral domain until stage 14. Within unsegmented head mesoderm, myf5 expression begins at stage 13.5 in the second branchial arch, followed within a few hours in the lateral rectus and first branchial arch myoblasts, then other eye and branchial arch muscles. Expression of myoD is detected initially in the first branchial arch beginning at stage 14.5, followed quickly by its appearance in other arches and eye muscles. Multiple foci of myoblasts expressing these transcripts are evident during the early stages of myogenesis in the first and third branchial arches and the lateral rectus-pyramidalis/quadratus complex, suggesting an early patterned segregation of muscle precursors within head mesoderm. Myf5-positive myoblasts forming the hypoglossal cord emerge from the lateral borders of somites 4 and 5 by stage 15 and move ventrally as a cohort. Myosin heavy chain (MyHC) is first immunologically detectable in several eye and branchial arch myofibers between stages 21 and 22, although many tongue and laryngeal muscles do not initiate myosin production until stage 24 or later. Detectable synthesis of the MyHC-S3 isoform, which characterizes myofibers as having "slow" contraction properties, occurs within 1-2 stages of the onset of MyHC synthesis in most head muscles, with tongue and laryngeal muscles being substantially delayed. Such a prolonged, 2- to 3-day period of regulatory gene expression preceding the onset of myosin production contrasts with the interval seen in muscles developing in axial (approximately 18 hr) and wing (approximately 1-1.5 days) locations, and is unique to head muscles. This finding suggests that ongoing interactions between head myoblasts and their surroundings, most likely neural crest cells, delay myoblast withdrawal from the mitotic pool. These descriptions define a spatiotemporal pattern of muscle regulatory gene and myosin heavy chain expression unique to head muscles. This pattern is independent of origin (somitic vs. unsegmented paraxial vs. prechordal mesoderm), position (extraocular vs. branchial vs. subpharyngeal), and fiber type (fast vs. slow) and is shared among all muscles whose precursors interact with cephalic neural crest populations. Dev Dyn 1999;216:96-112. PMID- 10536050 TI - Expression of cathepsin K in the human embryo and fetus. AB - Cathepsin K is a protease with high collagenolytic and elastinolytic activity. Its cellular expression was previously thought to be restricted to osteoclasts and osteoclast-mediated bone resorption. In this study, the expression of cathepsin K in the human embryo and fetus was demonstrated by immunohistochemistry, in situ hybridization, and by Northern blotting of fetal tissue extracts. Besides osteoclasts and chondroclasts and their precursors, epithelial cells of various organ systems expressed significant amounts of this enzyme. Respiratory and gastrointestinal mucosa, including bile duct epithelia and urothelia, showed high levels of cathepsin K expression. With the exception of the urothelium, showing a more homogenous expression pattern, the protease was usually accentuated in the surface cell layers of pithelia. In summary, these findings in the human embryo and early fetus demonstrated a significant expression of cathepsin K in different epithelial cell types besides osteoclasts. The functional aspects of cathepsin K expression in nonosteoclastic cells and potential conclusions on physiological and pathological conditions in the embryo fetal or adult organism remain to be investigated. Dev Dyn 1999;216:89-95. PMID- 10536052 TI - The hairless gene of the mouse: relationship of phenotypic effects with expression profile and genotype. AB - Various mutations of the hairless (hr) gene of mice result in hair loss and other integument defects. To examine the role of the hr gene in mouse development, the expression profile of hr has been determined by in situ hybridisation and correlated to the nature of genetic changes and morphological abnormalities in different mutant animals. Four variant alleles have been characterised at the molecular level. hr/hr mice produce reduced, but significant, levels of hr mRNA whereas other alleles contain mutations which would be expected to preclude the synthesis of functional product, demonstrating a correlation between allelic variation at the hr locus and phenotypic severity. hr expression was shown to be widespread and temporally regulated. It was identified in novel tissues such as cartilage, developing tooth, inner ear, retina, and colon as well as in skin and brain. Analysis of mice homozygous for the rhino allele of hairless revealed that, although no morphological defects were detectable in many tissues normally expressing hr, previously undescribed abnormalities were present in several tissues including inner ear, retina, and colon. These findings indicate that the hairless gene product plays a wider role in development than previously suspected. Dev Dyn 1999;216:113-126. PMID- 10536053 TI - Transitory expression of the A2b adenosine receptor during implantation chamber development. AB - Adenosine is a short-range signal molecule that surges in the mouse uterus immediately after blastocyst implantation (Blackburn et al. [1992] Dev. Dyn. 194:155-168). The present study has investigated patterns of uterine adenosine receptor expression during early post-implantation development. Strong expression of the A2b adenosine receptor was observed. Utilizing northern blot analysis, in situ hybridization, and immunostaining, the source of expression was mapped to the primary and secondary decidua of the antimesometrial region, between days 4-8 of gestation. Distribution of the A2b receptor protein followed that of the corresponding transcript by about one gestational day and reflected the dynamics of antimesometrial tissue organization during implantation chamber development. Uterine adenosine surges to levels sufficient for A2b receptor engagement during a defined period (i.e., days 4-6) after blastocyst implantation. Decidual A2b receptor expression thus defines a transitory window of murine gestation that corresponds to a period of human gestation encompassing most spontaneous pregnancy losses. Because adenosine receptors are sensitive to metabolically stable adenosine analogues, their differential expression during implantation chamber development may hold therapeutic potential in the prevention of early pregnancy loss. Dev Dyn 1999;216:127-136. PMID- 10536054 TI - Expression of Ret-, p75(NTR)-, Phox2a-, Phox2b-, and tyrosine hydroxylase immunoreactivity by undifferentiated neural crest-derived cells and different classes of enteric neurons in the embryonic mouse gut. AB - Cells of the enteric nervous system are derived from the neural crest. Probes to a number of molecules identify neural crest-derived cells within the gastrointestinal tract of embryonic mice prior to their differentiation into neurons and glial cells. However, it is unclear whether the different markers are identifying all neural crest-derived cells. In this study the distribution of p75(NTR)-immunoreactivity was compared with that of Ret-, Phox2a-, Phox2b-, and tyrosine hydroxylase (TH) in undifferentiated neural crest-derived cells in the E10.5-E13.5 mouse intestine. Neural crest-derived cells colonise the embryonic mouse gut in a rostral-to-caudal wave between E9.5-E14, and differentiation into enteric neurons also occurs in a rostral-to-caudal wave. Thus, the most caudal neural crest-derived cells within the gut are undifferentiated. These most caudal neural crest-derived cells co-expressed p75(NTR)-, Phox2b- and Ret immunoreactivity; at E10.5 a sub-population was also TH-positive. The most caudal cells did not show Phox2a-immunoreactivity at any stage. However, a sub population of cells, which was rostral to the undifferentiated neural crest derived cells, was Phox2a-positive, and these are likely to be cells beginning to differentiate along a neuronal lineage. The expression of Ret-, Phox2a-, Phox2b- and p75(NTR)-immunoreactivity by two classes of enteric neurons that differentiate prior to birth was also examined. Nitric oxide synthase (NOS) neurons showed Phox2b and Ret immunoreactivity at all ages, and Phox2a and p75(NTR) immunoreactivity only transiently. Calcitonin gene-related peptide (CGRP) neurons showed Phox2b and Ret-immunoreactivity, but not Phox2a immunoreactivity. It is concluded that all undifferentiated neural crest-derived cells initially express Phox2b, Ret, and p75(NTR); a sub-population of these cells also expresses TH transiently. Those cells that are beginning to differentiate along a neuronal lineage maintain their expression of Phox2b and Ret, and they start to express Phox2a, but down-regulate p75(NTR); those cells that differentiate along a glial lineage down-regulate Ret and maintain their expression of p75(NTR). Dev Dyn 1999;216:137-152. PMID- 10536056 TI - Beta-catenin is a major tyrosine-phosphorylated protein during mouse oocyte maturation and preimplantation development. AB - During mouse preimplantation development, the components of the E-cadherin catenin complex are derived from both maternal and zygotic gene activity and the adhesion complex is increasingly accumulated and stored in a nonfunctional form, ready to be used for compaction and the formation of the trophectoderm cell layer (Ohsugi et al., Dev. Dyn. 206:391-402, 1996). Here, we show that beta-catenin is a major tyrosine-phosphorylated protein in oocytes and early cleavage-stage embryos and that the relative amount of phosphorylated beta-catenin is greatly reduced during the morula-blastocyst transition. Peptide-specific antibodies indicate that beta-catenin undergoes conformational changes and/or that the carboxy-terminal region of beta-catenin is blocked during preimplantation development. Moreover, the availability of a carboxy-terminal epitope seems to depend on the tyrosine phosphorylation state of beta-catenin and becomes unmasked when oocytes are treated with the tyrosine kinase inhibitor genistein. Our results suggest that tyrosine phosphorylation of beta-catenin represents a molecular mechanism to keep the accumulating E-cadherin adhesion complex in a nonfunctional form. Dev Dyn 1999;216:168-176. PMID- 10536055 TI - Characterization of zebrafish primordial germ cells: morphology and early distribution of vasa RNA. AB - Research into germ line development is of conceptual and biotechnologic importance. In this study, we used morphology at the level of light and electron microscope to characterize the primordial germ cells (PGCs) of the zebrafish throughout embryonic and larval development. The study was complemented by the detailed analysis of mRNA expression of a putative germ line marker vasa. By morphology alone PGCs were identified at the earliest at the 5-somite stage in the peripheral endoderm in contact with the yolk syncytial layer. Subsequently, they move from lateral to medial positions into the median mesoderm and from there by means of the dorsal mesentery into the gonadal anlage at day 5 postfertilization (pf), to establish gonads with mesenchymal cells by day 9 pf. Ultrastructural analysis of the 4-day-old zebrafish larvae demonstrates the presence of the germ line-specific structures, nuage, and annulate lamellae. vasa RNA-positive cells can be followed during zebrafish embryogenesis from the 32 cell stage onward (Yoon et al., 1997). Upon completion of gastrulation, the RNA is exclusively present in the cells of the hypoblast, which as a consequence of convergence and extension movements first arrange themselves in a V-shaped string like conformation to end up, by late somitogenesis, as a string of cells on each side of the midline. We show that the localization of maternal vasa RNA in the ovary changes from cytoplasmic, in the previtellogenic oocytes, to cortical in the vitellogenic oocytes, to concentrate at the boundary of the yolk and cytoplasm in the one cell stage zygote. These results demonstrate that the cortical vasa RNA localization precedes its cleavage furrow-associated localization in the embryos and is presumably cytoskeleton dependent. vasa RNA localization changes from asymmetric subcellular at the sphere stage, to become entirely cytoplasmic at the dome stage. These data suggest a close resemblance in modes of segregation of the germ plasma in the frog and vasa mRNA in the fish during cleavage stages. Based on the significantly larger size and the stereotype and similar position of morphologically distinct cells, presumed to be PGCs, and their vasa RNA-positive counterparts, we conclude that vasa RNA-positive cells are the PGCs and vasa RNA represents a definitive germ line marker in the fish. Dev Dyn 1999;216:153-167. PMID- 10536057 TI - Protein kinase C activity regulates slow myosin heavy chain 2 gene expression in slow lineage skeletal muscle fibers. AB - Expression of the slow myosin heavy chain (MyHC) 2 gene defines slow versus fast avian skeletal muscle fiber types. Fetal, or secondary, skeletal muscle fibers express slow MyHC isoform genes in developmentally regulated patterns within the embryo, and this patterning is at least partly dependent on innervation in vivo. We have previously shown that slow MyHC 2 gene expression in vitro is regulated by a combination of innervation and cell lineage. This pattern of gene expression was indistinguishable from the pattern observed in vivo in that it was restricted to innervated muscle fibers of slow muscle origin. We show here that slow MyHC 2 gene expression in the slow muscle fiber lineage is regulated by protein kinase C (PKC) activity. Inhibition of PKC activity induced slow MyHC 2 gene expression, and the capacity to express the slow MyHC 2 gene was restricted to muscle fibers of slow muscle (medial adductor) origin. Fast muscle fibers derived from the pectoralis major did not express significant levels of slow MyHC 2 with or without inhibitors of PKC activity. This differential expression pattern coincided with different inherent PKC activities in fast versus slow muscle fiber types. Furthermore, over-expression of an unregulated PKCalpha mutant suppressed slow MyHC 2 gene expression in muscle fibers of the slow lineage. Lastly, denervation of skeletal muscles caused an increase in PKC activity, particularly in the slow medial adductor muscle. This increase in PKC activity was associated with lack of slow MyHC 2 gene expression in vivo. These results provide a mechanistic link between innervation, an intracellular signaling pathway mediated by PKC, and expression of a muscle fiber type-specific contractile protein gene. Dev Dyn 1999;216:177-189. PMID- 10536058 TI - Murine prenatal expression of cholecystokinin in neural crest, enteric neurons, and enteroendocrine cells. AB - Cholecystokinin (CCK) is a regulatory peptide that is primarily expressed in two adult cell types: endocrine cells of the intestine and neurons of the central nervous system. To determine the ontogeny of CCK expression during intestinal organogenesis, we created a mouse strain in which the CCK gene was replaced by a lacZ reporter cassette using homologous recombination in embryonic stem cells. Initially, CCK expression in the developing intestine was limited to the myenteric plexus of the enteric nervous system. This expression pattern was widespread, extending from the proximal stomach into the colon, yet transient, being detected soon after gut tube closure [embryonic day 10.5 (E10.5)] through E15.5. Since enteric neurons are derived from the neural crest, we examined earlier (E8.5-9.5) embryos and concluded that lacZ was expressed in subpopulations of neural tube and neural crest cells. Endocrine cell expression in the intestinal epithelium occurred later, beginning at E15.5 as enteric neuronal expression was dwindling. This expression persisted to yield the adult pattern of scattered single endocrine cells in the upper small intestine. The data show that CCK is a very early marker of both neuronal and endocrine cell lineages in the developing gastrointestinal tract. Furthermore, reverse transcriptase polymerase chain reaction (RT-PCR) analysis showed that CCK receptor transcripts were detected in embryos as early as E10.5, suggesting that CCK signaling is established early in mouse development. Dev Dyn 1999;216:190 200. PMID- 10536060 TI - Adaptations to neonatal anoxia and control of ventilation. PMID- 10536059 TI - Differential expression of Hoxa-2 protein along the dorsal-ventral axis of the developing and adult mouse spinal cord. AB - We have used synthetic oligopeptides derived from the coding sequence of the murine Hoxa-2 protein to produce polyclonal antibodies that specifically recognize the Hoxa-2 recombinant protein. Immunohistochemical studies reveal a distinct pattern of spatial and temporal expression of Hoxa-2 protein within the mouse spinal cord which is concomitant with the cytoarchitectural changes occurring in the developing cord. Hoxa-2 protein is predominantly detected in the nuclei of cells in the ventral mantle region of 10-day-old mouse embryos. Islet 1, a marker for motor neurons was also shown to be co-localized with Hoxa-2 in nuclei of cells in this region. As development progresses from 10-days to 14-days of gestation, Hoxa-2 protein expression gradually extends to the dorsal regions of the mantle layer. The Hoxa-2 protein expression pattern changes at 16-days of embryonic development with strong expression visible throughout the dorsal mantle layer. In 18-day-old and adult mouse spinal cords, Hoxa-2 protein was expressed predominantly by cells of the dorsal horn and only by a few cells of the ventral horn. Double labeling studies with an antibody against glial fibrillary acidic protein (GFAP, an astrocyte-specific intermediate filament protein) showed that within the adult spinal cord, astrocytes rarely expressed the Hoxa-2 protein. However, Hoxa-2 and GFAP double-labeled astrocytes were found in the neopallial cultures, although not all astrocytes expressed Hoxa-2. Hoxa-2 expressing oligodendrocyte progenitor cells were also identified after double-labeling with O4 and Hoxa-2 antibodies; although cells in this lineage that have begun to develop a more extensive array of cytoplasmic processes were less likely to be Hoxa-2 positive. The early pattern of Hoxa-2 protein expression across transverse sections of the neural tube is temporally and spatially modified as each major class of neuron is generated. This congruence in the expression of the Hoxa-2 protein and the generation of neurons in the cord suggests that the Hoxa-2 protein may contribute to dorsal-ventral patterning and/or to the specification of neuronal phenotype. Dev Dyn 1999;216:201-217. PMID- 10536061 TI - Effect of neonatal anoxia on the ventilatory response to hypoxia in developing rats. AB - The purpose of the study was to determine whether an episode of anoxia in the neonatal period affects the hypoxic ventilatory response in developing rats. Three to 4 day old rats (EXP rats) (day 0 = day of birth) were exposed to 100% N(2) for about 20 min. In anoxia, animals changed their respiratory movements to "gasping," and were successfully autoresuscitated by placing them back into air. None of the EXP rats died after the resuscitation, and no differences were found between the EXP and control (CONT) rats in the somatic growth. The CONT rats were not exposed to anoxia during the neonatal period. On day 9-10(mean day 9) and days 23-29(mean day 25), ventilatory and metabolic responses to hypoxia (inspiratory oxygen fraction F(i)O(2 )= 10% on day 9; 15% and 10% on day 25) were compared in the EXP and CONT groups. On day 9, there were no differences in ventilatory and metabolic responses to hypoxia between the EXP and CONT groups. On day 25, 10% O(2)hypoxic ventilatory response (by the barometric method) was significantly higher in the EXP group than in the CONT group, as indicated by increases in both tidal volume (V(T)) and mean inspiratory flow (V(T)/T(I)). On the other hand, the metabolic effects were small, because metabolic rates (V'O(2) and V'CO(2), by a flow-through method) were similar in both EXP and CONT groups under each gaseous condition. These results suggest that an episode of anoxia during the neonatal period has long-lasting effects on the control of ventilation, including chemoreflexes, in rats. PMID- 10536062 TI - Diagnostic accuracy of oropharyngeal cultures in infants and young children with cystic fibrosis. AB - The objective of this study was to assess the diagnostic accuracy of oropharyngeal (OP) cultures relative to simultaneous bronchoalveolar lavage (BAL) cultures in very young children with CF, and to examine the effects of bacterial density, age, and study cohort on diagnostic accuracy. Respiratory culture data were analyzed from three independent, prospective studies involving simultaneous collection of 286 OP and BAL cultures from 141 children with CF <5 years of age. For predicting any growth of Pseudomonas aeruginosa (Pa) from the lower airway in subjects /=10(3) or >/=10(5) cfu/mL. Specificity for Pa declined significantly with increasing age. In children with CF <5 years of age, the specificity and negative predictive value of OP cultures for Pa are high, while the sensitivity and positive predictive value are poor. Thus, in this age range, a negative throat culture is helpful in "ruling out" lower airway infection with Pa. However, a positive culture does not reliably "rule in" the presence of Pa in the lower respiratory tract. These findings may have implications for study design and interpretation as well as clinical management of young children with CF. PMID- 10536063 TI - Bronchial casts in children with cardiopathies: the role of pulmonary lymphatic abnormalities. AB - Expectoration of bronchial casts, a condition also called plastic bronchitis, is very rare in children. Bronchial casts may be associated with bronchopulmonary diseases associated with mucus hypersecretion, bronchopulmonary bacterial infections, congenital and acquired cardiopathies, or pulmonary lymphatic abnormalities. A classification based on anatomy and pathology has been proposed which identifies an "acellular" group associated with congenital cardiopathies and palliative surgery. We report on 3 cases with bronchial casts associated with cardiopathy. Observations suggest that the formation of bronchial casts may result from lymphatic leakage into the bronchi. The 3 cases on which we report were immunodeficient and had pulmonary lymphatic abnormalities. The bronchial casts contained lymphocytes and lipids, as determined by histologic examination. In the absence of congenital pulmonary or diffuse lymphatic dysplasia associated with cardiopathy, the principal factors resulting in the formation of bronchial casts appear to be surgical trauma to the lymphatic channels surrounding the bronchi, pleural adhesions, and high systemic venous blood pressure. The prognosis for these patients is poor, and possibilities for treatment are limited. PMID- 10536064 TI - Efficacy of once-daily versus twice-daily administration of budesonide by Turbuhaler(R) in children with stable asthma. AB - We evaluated the efficacy of once-daily versus twice-daily treatment with budesonide, delivered by a Turbuhaler(R), in the management of children with stable asthma in a randomized, double-blind, parallel-group study involving 206 children (age 5-15 years). After a 2-week run-in period during which the children were maintained on their usual dose of budesonide (200 microg or 400 microg/day), patients were randomized to receive the same daily dose in either two daily administrations (morning and evening) or as a single dose in the morning over a period of 12 weeks. The primary efficacy variable was morning peak expiratory flow (PEF). The mean morning PEF during the run-in phase was 271 L/min in patients randomized to once-daily treatment and 264 L/min in those randomized to twice-daily treatment. The mean change from baseline to the last 2 weeks of the treatment period in the two groups was -0.3 L/min (95% confidence limits -6.6 to +6.0) and 2.5 L/min (-4.3 to +9.3). The estimated difference between the groups was -2.8 L/min, with 90% confidence limits of -10.4 + 4.5; these were close to the limits regarded as indicative of equivalence (-10 to +10), and hence the difference was not regarded as clinically relevant. Similarly, there were no significant differences between the groups in regard to secondary efficacy measures such as spirometric tests and symptom scores. Both treatments were well tolerated. We conclude that once-daily administration of budesonide by Turbuhaler(R) is as effective as twice-daily treatment in the management of stable asthma in children treated with inhaled steroids at doses of 200-400 microg/day. PMID- 10536065 TI - Diagnosis of tuberculosis in children using a polymerase chain reaction. AB - We investigated the value of the polymerase chain reaction (PCR) in the diagnosis of active tuberculosis in children and evaluated the relationship between PCR results in children with tuberculous infections and mediastinal adenopathies detected by computerized tomography (CT-Scan). This was a controlled, blinded, prospective study comparing nested PCR, mycobacterial cultures and the clinical diagnosis based on 350 clinical specimens from 117 children referred for evaluation of suspected pulmonary tuberculosis. All children with tuberculous infection but without active disease underwent a chest CT-scan to detect the presence of mediastinal adenopathies not evident on chest x-ray. The sensitivity of PCR was 56.8% in children with clinically active disease (culture: 37.8%; smears: 13.5%). A major advantage of PCR over cultures was noted when there was no parenchymal involvement on chest radiograph and when the patient was undergoing anti-tuberculous treatment. There were nine specimens with false negative PCR results due to the presence of amplification reaction inhibitors. PCR was positive in five children with tuberculous infection without active disease and these children presented mediastinal adenopathies on the CT-scan that were not evident on chest radiography. There were no false-positive PCR results in the control groups of children. We conclude that nested PCR is a rapid and sensitive method for the early diagnosis of tuberculosis in children. It is especially useful when the diagnosis of active tuberculosis is difficult. In our study children with tuberculous infection without apparent disease who have positive PCR results have mediastinal adenopathies on CT-scan. PMID- 10536066 TI - Noninvasive diagnosis of P. carinii pneumonia on oral washes in an HIV-infected child. PMID- 10536068 TI - Serine proteinase inhibitor therapy in alpha(1)-antitrypsin inhibitor deficiency and cystic fibrosis. AB - Proteinase-antiproteinase imbalances are recognized in several diseases including the two most common lethal hereditary disorders of white populations, alpha(1) antitrypsin (alpha(1)-AT) deficiency and cystic fibrosis (CF). In alpha(1)-AT deficiency, the type Z variant of alpha(1)-AT forms polymers in the endoplasmic reticulum of hepatocytes resulting in liver disease in childhood. The block in alpha(1)-AT processing in hepatocytes significantly reduces levels of circulating alpha(1)-AT. This may lead in young adults to panacinar emphysema due to insufficient protection of the lower respiratory tract from neutrophil elastase, permitting progressive destruction of the alveoli. In CF, chronic bacterial lung infections due to impaired mucociliary clearance lead to a vigorous influx of neutrophils in the airways. Released levels of neutrophil serine proteinases, particularly elastase, exceed the antiproteinase capacity of endogenous serine proteinase inhibitors in the airways. Progressive proteolytic impairment of multiple defense pathways in addition to endobronchial obstruction and airway wall destruction are thought to be responsible for the reduced life expectancy in CF patients. Strategies to augment the antiproteinase defenses in the airways of patients with severe alpha(1)-AT deficiency or CF include the intravenous or aerosol administration of serine proteinase inhibitors. Studies in both patient groups using plasma-derived or transgenic alpha(1)-AT, recombinant secretory leukoprotease inhibitor or synthetic elastase inhibitors show promising results concerning drug safety and efficacy. PMID- 10536067 TI - Intraindividual variability of infant whole-body plethysmographic measurements: effects of age and disease. AB - The intraindividual variability of whole-body plethysmographic measurements was studied in a large series of consecutive infants (N = 144), divided into two groups: a group of infants born very prematurely (PM, N = 63), with (N = 28) or without (N = 35) a history of bronchopulmonary dysplasia (BPD), and a group of infants with persistent respiratory symptoms (PRS, N = 81), i.e., wheezing (N = 53) or cough (N = 28). The intraindividual variability was determined within each test and between tests, separated by a 10-min interval. In both study groups, the between-test variability was significantly larger than that within tests. Expressed as the median coefficient of variation (CV), the between-test repeatabilities in the PRS group were 8.0% for thoracic gas volume (TGV), 17.5% for airway resistance (Raw), and 18.4% for specific airway conductance (sGaw), and in the PM group, 8.9% for TGV, 20.4% for Raw, and 20.7% for sGaw. However, the individual range of CVs was large, ranging from 3 to 19% for TGV and from 5 to 55% for sGaw. With respect to TGV, the difference between the groups was statistically significant (P = 0.03). In infants with a history of BPD, there was also a significant negative age dependency in CVs of sGaw (r = -0.50, P = 0. 009), showing larger variation among younger individuals. The presenting symptom (wheezing or cough) in the PRS group did not influence the measurement variability significantly, and neither did the degree of bronchial obstruction. We conclude that on a group basis, the repeatability of infant body plethysmographic measurements may be satisfactory for scientific studies demonstrating pharmacodynamic effects; however, the intraindividual measurement variability should be reported for each test conditions and for infant groups in each study. Due to the large range in individual variation and the influence of age and disease processes on the variation, for an individual child there is only questionable benefit from a given measurement, unless the intrasubject, between test variability is assessed individually before interventions, such as a bronchodilation test. PMID- 10536069 TI - Pneumonectomy in cystic fibrosis. AB - A 17-year-old boy and a 12-year-old girl with cystic fibrosis (forced expiratory volume in 1 sec, 36% and 14% of predicted values, respectively) developed severe right-sided lung infections with abscess formations and complete atelectases unresponsive to medical therapy. In both patients, unilateral emergency pneumonectomy resulted in rapid clinical improvement. Despite her severe underlying lung disease, the girl experienced a remarkable increase in quality of life; 2 years after surgery, she died from respiratory failure. The male patient has now survived for 4 years, and lung transplantation still remains a therapeutic option for him. We believe that pneumonectomy is a valuable rescue therapy for patients with cystic fibrosis and intractable unilateral lung infections who are at high risk of dying while waiting for lung transplantation. PMID- 10536070 TI - Treatment of anorexia and weight loss with megestrol acetate in patients with cystic fibrosis. AB - Four patients with severe cystic fibrosis lung disease, anorexia and weight loss, received Megestrol Acetate (MA), as an appetite stimulant. The initial dose was 400-800 mg daily and was continued for 6-15 months. Appetite was improved, with significant weight gain in all patients and an increase in their weight for age percentile from <5% at the start of the study to approximately the 25(th) percentile after 6 months of use and improvement in quality of life. One patient discontinued MA after 6 months, and subsequently appetite and weight were depressed. PMID- 10536072 TI - Selected abstracts PMID- 10536071 TI - Reduced asthma morbidity following pediatric pulmonary consultation. PMID- 10536082 TI - Working memory(s). AB - Working memory is variously defined as a set of linked and interacting information processing components that maintain information in a short-term store (or retrieve information into that store) for the purpose of the active manipulation of the stored items. The purpose of the this Special Issue is to present data relevant to the question of the functional organization of working memory. In this Introduction we review the two models of working memory and suggest that some of the similarities may be more apparent than real. We further suggest that the two models describe different systems that are specialized for different kinds of stimuli and for different kinds of information processing. PMID- 10536083 TI - Spatial working memory in Asperger's syndrome and in patients with focal frontal and temporal lobe lesions. AB - Spatial working memory (SWM) was investigated in 15 patients with Asperger's syndrome (AS) comparing their performance to 18 age- and IQ-matched control subjects. An additional comparison was made with 20 unilateral frontal excision patients [9 right (RFL); 11 left (LFL)] and with 38 unilateral temporal lobectomy patients [18 right (RTL); 18 left (LTL)], the frontal and temporal lobe patients having separate matched control groups. SWM was tested using the Executive Golf Task, a test that also measures spatial strategy formation. The AS group showed a substantial deficit on SWM, but no impairment in strategy formation. The LFL showed the same pattern of impairment, but with a less substantial deficit. The RFL group showed a large deficit, but some of this was accounted for by a strategy formation impairment. Of the temporal lobe lesions groups, only the RTL group was impaired on SWM, but this group showed normal strategy formation. It was concluded that the SWM deficit in AS may reflect a more general difficulty in accessing different types of representations in order to guide voluntary behavior, providing at least a partial explanation for the executive deficits found in AS. PMID- 10536084 TI - A comment on the functional localization of the phonological storage subsystem of working memory. AB - Working memory, as defined by Baddeley and Hitch, is an interactive set of cognitive processes responsible for holding information online and available for analysis. Part of that system is a specialized subsystem devoted to maintaining verbal information; the verbal "slave" subsystem has as a core component a phonological store. For many years, the anatomical locus of the phonological store has been thought to be in the supramarginal and angular gyri of the speech dominant hemisphere, and functional neuroimaging studies provide broad support for this localization. However, a finer grained analysis of published experiments reveals two possible foci for the phonological store within the parietal lobe, neither of which has a pattern of functional activation that is fully consistent with the Baddeley and Hitch model. The purpose of the present paper is to review several studies relevant to the question of the localization of the phonological store and to suggest possible reasons the results are discrepant. PMID- 10536085 TI - Hippocampal involvement in spatial and working memory: a structural MRI analysis of patients with unilateral mesial temporal lobe sclerosis. AB - Forty-seven patients with unilateral temporal lobe epilepsy (TLE) were investigated on the Nine-Box Maze. The task was designed to compare working memory and spatial mapping theories of the functions of the hippocampus and provide measures of spatial, object, working, and reference memory. The results extended our previous findings in a larger group of patients. Spatial memory deficits across both working and reference memory conditions were found in patients with a right epileptogenic focus. There was no evidence of an object working memory deficit, but a nonlateralized impairment in object reference memory was revealed, which is consistent with our previous findings. The pattern of results was confirmed in a subgroup of 33 patients with unilateral atrophy localized to the hippocampus and parahippocampal gyrus, as verified by volumetric analysis of magnetic resonance images. In addition spatial memory errors significantly correlated with volumetric measures of mesial temporal lobe structures and not with measures of the remaining temporal cortex. In contrast, object reference memory errors correlated with volumetric measures of the temporal cortex and not with mesial temporal lobe structures. These findings support a specialized role for the right hippocampal region in spatial memory. PMID- 10536086 TI - Maintenance versus manipulation of information held in working memory: an event related fMRI study. AB - One model of the functional organization of lateral prefrontal cortex (PFC) in primates posits that this region is organized in a dorsal/ventral fashion subserving spatial and object working memory, respectively. Alternatively, it has been proposed that a dorsal/ventral subdivision of lateral PFC instead reflects the type of processing performed upon information held in working memory. We tested this hypothesis using an event-related fMRI method that can discriminate among functional changes occurring during temporally separated behavioral subcomponents of a single trial. Subjects performed a delayed-response task with two types of trials in which they were required to: (1) retain a sequence of letters across the delay period (maintenance) or (2) reorder the sequence into alphabetical order across the delay period (manipulation). In each subject, activity during the delay period was found in both dorsolateral and ventrolateral PFC in both types of trials. However, dorsolateral PFC activity was greater in manipulation trials. These findings are consistent with the processing model of the functional organization of working memory in PFC. PMID- 10536087 TI - Working memory and vigilance: evidence from normal aging and Alzheimer's disease. AB - Both single unit recording and neuroradiological studies suggest that frontal and executive processes are necessary for visual maintenance rehearsal. This observation is linked to the classic vigilance literature by the proposal that vigilance decrement is found when the subject is required to maintain a representation over a brief delay. Vigilance performance was therefore studied in a sample of elderly subjects who were tested over a 40-min period involving perceptual or memory-based tasks which were matched for initial level of performance. There was a significant interaction between task and delay, with only the memory-based task showing decrement. A second study used the same two tasks to investigate vigilance performance in patients suffering from probable Alzheimer's Disease. Over a 15-min delay period, an equivalent interaction effect occurred, again indicating substantially greater decrement for the memory-based task. The results are interpreted as consistent with a role for the executive processes of working memory in both visual rehearsal and vigilance performance. PMID- 10536088 TI - The scaling of basicranial flexion and length. AB - Based on correlations between the cranial base angle (CBA) and the index of relative encephalization (IRE, calculated as the cubed root of brain volume divided by basicranial length), several recent studies have identified relative brain size as the factor most responsible for determining basicranial flexion in primates. IRE, however, scales with positive allometry relative to body mass, unlike the negatively allometric relationship between brain volume and body mass. This poses new questions concerning the factors underlying the correlation between IRE and CBA. Specifically, if basicranial flexion represents a spatial solution to the problem of housing a large brain within a neurocranium of limited size, then why is it that the problem is greatest in those species whose brains are smallest relative to body mass? To address this question, the scaling relationships of IRE and the measurements used to calculate it were examined in 87 primate species. It was found that the positive allometry of IRE is due to the fact that its denominator, basicranial length (BL), scales with very strong negative allometry relative to body mass. The scaling relationship of BL may reflect the fact that the noncortical components of the brain (i.e., diencephalon, mesencephalon, medulla) also scale with strong negative allometry relative to body mass, perhaps because of energetic constraints. Importantly, BL and these three brain components scale isometrically against each other. Thus, although cranial base flexion may be an adaptation to accommodate the size of the brain relative to basicranial length, the reason why that adaptation is necessary is not the evolution of a large brain, but rather the evolution of a short cranial base. In so far as basicranial length is affected by the strong negative allometry of the diencephalon, mesencephalon and medulla, the scaling relationships of these brain components are therefore indirectly responsible for the evolution of basicranial flexion. PMID- 10536089 TI - Early pleistocene habitat in member 1 Olorgesailie based on paleosol stable isotopes. AB - Stable carbon and oxygen isotope values from soil carbonates were used to determine the vegetation context of archaeological sites and local climatic conditions represented in a approximately 0.99 Ma paleosol that is exposed laterally in the Olorgesailie basin, southern Kenya rift valley. As part of this landscape-scale project, samples of an upper Member 1 paleosol were analyzed along nearly 4 km of outcrop in three adjacent parts of the basin. Modern East African soil and plant community analogs are used to interpret the isotope ratios. The carbon isotopic composition of the paleosol carbonates indicates that the area supported a local biomass of about 75-100% C(4)plants during the period of soil formation. After averaging the data for each trench, an open C(4)grassland is represented by half of the carbon values, with wooded grassland more abundant across the paleolandscape than it is in the area now. This vegetational reconstruction is supported by the mammalian faunal assemblage, which has a high percentage of grazers. Although the relatively small sample size outside the main excavation area precludes firm characterization of vegetational diversity across the basin in upper Member 1 times, eastern and western localities in the study area may have had more woody C(3)plants than the widely sampled zone in between. Oxygen isotopes indicate that the lowland basin was slightly cooler and moister than today's semi-arid climate, with greater annual rainfall. Archaeological traces have a virtually continuous distribution across the paleolandscape, but vary in density of occurrence. With the strong evidence for C(4)grassland as the primary vegetation context across most of the study area, no habitat preference by the Acheulean toolmakers at Olorgesailie is shown in our initial comparison between carbon isotope values and stone/bone densities. PMID- 10536090 TI - Paleolandscape variation and early pleistocene hominid activities: members 1 and 7, Olorgesailie formation, Kenya. AB - Paleolandscape research tests for variation in the spatial distribution of hominid artefacts and establishes the association of hominid activities with paleoenvironmental features over distances of 100s to 1000s of meters. This approach requires (1) precise definition of narrow stratigraphic intervals based on sedimentary criteria that can be documented over a broad area, and (2) excavation of these intervals in order to establish taphonomic and paleoenvironmental contexts. In this report, excavations of three target intervals within the early Pleistocene deposits (992 to 780 ka) of the Olorgesailie basin are described. Assessment of time-averaging and paleolandscape structure shows that each target interval represents a relatively brief period (50%) inhibition of ICAM-1 expression in mammalian cells. PMID- 10536140 TI - Multi-forms of human MTH1 polypeptides produced by alternative translation initiation and single nucleotide polymorphism. AB - The human MTH1 gene for 8-oxo-7,8-dihydrodeoxyguanosine triphosphatase, produces seven types (types 1, 2A, 2B, 3A, 3B, 4A and 4B) of mRNAs. The B-type mRNAs with exon 2b-2c segments have three additional in-frame AUGs in their 5' regions. We report here that these transcripts produce three forms of MTH1 polypeptides (p22, p21 and p18) in in vitro translation reactions. Three polypeptides were also detected in extracts of human cells, using western blotting. B-type mRNAs with a polymorphic alteration (GU-->GC) at the beginning of exon 2c that converts an in frame UGA to CGA yielding another in-frame AUG further upstream, produced an additional polypeptide (p26) in vitro. Substitution of each AUG abolished the production of each corresponding polypeptide. Cell lines from individuals with the GC allele contain more B-type mRNAs than do those of GT homozygotes, and the former produce all of four polypeptides but the latter lack p26. Amounts of each polypeptide reflected copy number of the GC allele in each cell line. There is an apparent linkage dis-equilibrium between the two polymorphic sites, GT/GC at exon 2c and Val83/Met83 at codon 83 for p18. PMID- 10536141 TI - Cloning and analysis of a Toxoplasma gondii histone acetyltransferase: a novel chromatin remodelling factor in Apicomplexan parasites. AB - The yeast transcriptional adaptor GCN5 functions as a histone acetyltransferase, directly linking chromatin modification to transcriptional regulation. Homologues of yeast GCN5 have been found in Tetrahymena, Drosophila, Arabidopsis and human, suggesting that this pathway of chromatin remodelling is evolutionarily conserved. Consistent with this view, we have identified the Toxoplasma gondii homologue, referred to here as TgGCN5. The gene codes for a protein of 474 amino acids with an estimated molecular mass of 53 kDa. The protein reveals two regions of close similarity with the GCN5 family members, the HAT domain and the bromodomain. Tg GCN5 occurs in a single copy in the T.gondii genome. The introduction of a second copy of TgGCN5 in T.gondii tachyzoites is toxic unless the HAT activity is disrupted by a single point mutation. Full TgGCN5 does not complement the growth defect in a yeast gcn5 (-)mutant strain, but a chimera comprising the T.gondii HAT domain fused to the remainder of yGCN5 does. These data show that T.gondii GNC5 is a histone acetyltransferase attesting to the significance of chromatin remodelling in gene regulation of Apicomplexa. PMID- 10536142 TI - Characterization of transient RNA-RNA interactions important for the facilitated structure formation of bacterial ribosomal 16S RNA. AB - The co-transcribed leader sequences of bacterial rRNA are known to affect the structure and function of the small ribosomal subunits. Base changes in the leader nut -like sequence elements have been shown to cause misfolded but correctly processed 16S rRNA structures at low growth temperature. Transient interactions of leader sequences with the nascent 16S rRNA are considered to guide rRNA folding and to facilitate correct structure formation. In order to understand this chaperone-like activity of the leader RNA we have analyzed the thermodynamic stabilities of wild-type and mutant leader transcripts. We show here that base changes cause subtle differences in the melting profiles of the corresponding leader transcripts. Furthermore, we show that direct interaction between leader transcripts and the 16S rRNA is limited to the 5'-domain of the 16S rRNA for both wild-type and mutant leaders. Binding studies of mutant and wild-type leader transcripts to 16S rRNA revealed small changes in the affinities and the thermal stabilities as a consequence of the base changes. Different complex stabilities as a function of the Mg(2+) ion concentration indicated that mutant and wild-type leader transcripts interact differently with the 16S rRNA, consistent with a less stable and tightly folded structure of the mutant leader. Employing time-resolved oligonucleotide hybridization assays we could show different folding kinetics for 16S rRNA molecules when linked to wild-type leader, mutant leader or in the absence of leader RNA. The studies help to understand how bacterial rRNA leader transcripts may affect the folding of the small subunit rRNA. PMID- 10536143 TI - Localisation of a reporter transcript by the c-myc 3'-UTR is linked to translation. AB - The 3'-untranslated region of c-myc mRNA contains a perinuclear localisation signal which is sufficient to target beta-globin coding sequences. The link between perinuclear mRNA localisation and translation has been investigated using cells transfected with chimeric gene constructs in which globin reporter sequences were linked to the c-myc 3'-untranslated region and the iron-responsive element from ferritin mRNA. Iron supplementation of the medium promoted translation of the chimeric mRNA as assessed by its presence in polysomes; in situ hybridisation showed that the mRNA was localised around the nucleus. Treatment with the iron chelator desferrioxamine for 16 h prevented both translation and mRNA localisation. In controls where the expressed mRNA lacked the iron-responsive element desferrioxamine had no effect upon localisation. In contrast, arrest of on-going global translation by puromycin treatment had no effect on mRNA localisation. The data suggest that if initiation of translation of a mRNA containing the c-myc localisation signal is prevented in some way then localisation does not occur, whereas once the mRNA has been localised further translation is not required to maintain mRNA localisation. PMID- 10536144 TI - The antiviral enzymes PKR and RNase L suppress gene expression from viral and non viral based vectors. AB - Expression of transfected genes is shown to be suppressed by two intracellular enzymes, RNase L and protein kinase PKR, which function in interferon-treated cells to restrict viral replication. RNase L(-/-) or PKR(-/-) murine embryonic fibroblasts produced enhanced levels of protein from transfected genes compared with wild-type cells. Increased expression of exogenous genes in RNase L(-/-) cells correlated with elevated levels of mRNA and thus appeared to be due to enhanced mRNA stability. Plasmid encoding adenovirus VA RNAs was able to further enhance accumulation of the exogenous gene transcript and protein, even in cells lacking PKR. In contrast to the increased expression of transfected genes in cells lacking RNase L or PKR, expression of endogenous host genes was unaffected by the absence of these enzymes. In addition, a dominant-negative PKR mutant improved expression from a conventional plasmid vector and from a Semliki Forest virus derived, self-replicating vector. These results indicate that viral infections and transfections produce similar stress responses in mammalian cells and suggest strategies for selectively increasing expression of exogenous genes. PMID- 10536145 TI - Identification of 23S rRNA nucleotides neighboring the P-loop in the Escherichia coli 50S subunit. AB - We report the synthesis of a radioactive, photolabile 2'-O-methyloligoRNA probe, 2258-53/52(SAz)-48, PHONT1, and its exploitation in identifying 23S rRNA nucleotides neighboring the so-called 'P-loop'. The probe is complementary to nt 2248-2258 in Escherichia coli 50S subunits. PHONT1 contains a p-azidophenacyl group attached to a phosphorothioate bridge between the nucleotides complementary to the positions 2252-2253, such that the photogenerated nitrene is maximally 17 19 A from 23S RNA nucleotides G2252 and G2253. PHONT1 binds to the 50S subunit, and photoincorporates within or immediately adjacent to its target site, as well as into several nucleotides falling between G2357 and A2430. The significance of these results for the structure of the peptidyl transferase center is considered. The PHONT approach is generally applicable to studies of complex RNA-containing molecules. PMID- 10536146 TI - Formation of 5-formyl-2'-deoxycytidine from 5-methyl-2'-deoxycytidine in duplex DNA by Fenton-type reactions and gamma-irradiation. AB - 5-methyl-2'-deoxycytidine (5-Me-dC) is formed by the enzymatic methylation of dC, primarily in CpG sequences in DNA, and is involved in the regulation of gene expression. In the present study, 5-Me-dC and double-stranded DNA fragments containing 5-Me-dC were either gamma-irradiated or aerobically treated with Fenton-type reagents, Fe(II)-EDTA, Fe(II)-nitrilotriacetic acid, Fe(III)-EDTA H(2)O(2)-catechol or ascorbic acid-H(2)O(2) under neutral conditions. The formation of 5-formyl-2'-deoxycytidine (5-CHO-dC) was observed upon treatment of both 5-Me-dC and DNA fragments containing 5-Me-dC. The yields of 5-CHO-dC from 5 Me-dC and those of 5-formyl-2'-deoxyuridine from dT were comparable. These results suggest that 5-Me-dC in DNA is as susceptible to oxidation as dT in cells, and raise the possibility that 5-CHO-dC may contribute to the high mutagenic rate observed in CpG sequences in genomic DNA. PMID- 10536147 TI - Expression of GUT1, which encodes glycerol kinase in Saccharomyces cerevisiae, is controlled by the positive regulators Adr1p, Ino2p and Ino4p and the negative regulator Opi1p in a carbon source-dependent fashion. AB - In Saccharomyces cerevisiae glycerol utilization is mediated by two enzymes, glycerol kinase (Gut1p) and mitochondrial glycerol-3-phosphate dehydrogenase (Gut2p). The carbon source regulation of GUT1 was studied using promoter-reporter gene fusions. The promoter activity was lowest during growth on glucose and highest on the non-fermentable carbon sources, glycerol, ethanol, lactate, acetate and oleic acid. Mutational analysis of the GUT1 promoter region showed that two upstream activation sequences, UAS(INO) and UAS(ADR1), are responsible for approximately 90% of the expression during growth on glycerol. UAS(ADR1) is a presumed binding site for the zinc finger transcription factor Adr1p and UAS(INO) is a presumed binding site for the basic helix-loop-helix transcription factors Ino2p and Ino4p. In vitro experiments showed Adr1 and Ino2/Ino4 protein-dependent binding to UAS(ADR1) and UAS(INO). The negative regulator Opi1p mediates repression of the GUT1 promoter, whereas the effects of the glucose repressors Mig1p and Mig2p are minor. Together, the experiments show that GUT1 is carbon source regulated by different activation and repression systems. PMID- 10536148 TI - Transcription-independent phosphorylation of the RNA polymerase II C-terminal domain (CTD) involves ERK kinases (MEK1/2). AB - The largest subunit of the mammalian RNA polymerase II possesses a C-terminal domain (CTD) consisting of 52 repeats of the consensus sequence, Tyr(1)-Ser(2) Pro(3)-Thr(4)-Ser(5)-Pro(6)-Ser(7). Phosphorylation of the CTD is known to play a key role in gene expression. We now show that treatments such as osmotic and oxidative shocks or serum stimulation generate a new type of phosphorylated subunit, the IIm form. This IIm form might be generated in vivo by ERK-type MAP kinase phosphorylation as: (i) ERK1/2 are major CTD kinases found in cell extracts; (ii) the immunoreactivity of the IIm form against a panel of monoclonal antibodies indicates that the CTD is exclusively phosphorylated on Ser-5 in the repeats, like RNA polymerase II phosphorylated in vitro by an ERK1/2; and (iii) the IIm form does not appear when ERK activation is prevented by treating cells with low concentrations of highly specific inhibitors of MEK1/2. Since the IIm subunit is not affected by inhibition of transcription and is not bound to chromatin, it does not participate in transcription. PMID- 10536149 TI - Novel coding regions in four complete archaeal genomes. AB - In the process of analysing the four available complete archaeal genomes, we have noted that certain regions characterised as 'non-coding' exhibit significant sequence similarity to other protein sequences from Archaea and other species. Using established technology, we have identified a number of potential protein coding regions in these putative 'non-coding' regions. We have detected 524 such cases, of which 113 regions appear to code for proteins present in archaeal or other species, while the remaining 411 regions are mostly start/stop definition conflicts. Of the 113 protein coding regions, only 21 code for proteins with homologues of known function. The number of novel coding sequences identified herein amounts to 1. 5% of the total genome entries, while the conflicting cases represent an additional 5%. The observed differences between the four complete archaeal genomes seem to reflect disparate approaches to genome annotation. Genome sequence collections should be regularly checked to improve gene prediction by sequence similarity and greater effort is required to make gene definitions consistent across related species. PMID- 10536150 TI - Markerless gene replacement in Escherichia coli stimulated by a double-strand break in the chromosome. AB - A simple and efficient gene replacement method, based on the recombination and repair activities of the cell, was developed. The method permits the targeted construction of markerless deletions, insertions and point mutations in the Escherichia coli chromosome. A suicide plasmid, carrying the mutant allele and the recognition site of meganuclease I- Sce I, is inserted into the genome by homologous recombination between the mutant and the wild-type (wt) alleles. Resolution of this cointegrate by intramolecular recombination of the allele pair results in either a mutant or a wt chromosome which can be distinguished by allele-specific PCR screening. The resolution process is stimulated by introducing a unique double-strand break (DSB) into the chromosome at the I- Sce I site. Cleavage by the nuclease not only enhances the frequency of resolution by two to three orders of magnitude, but also selects for the resolved products. The DSB-stimulated gene replacement method can be used in recombination-proficient E.coli cells, does not require specific growth conditions, and is potentially applicable in other microorganisms. Use of the method was demonstrated by constructing a 17-bp and a 62-kb deletion in the MG1655 chromosome. Cleavage of the chromosome induces the SOS response but does not lead to an increased mutation rate. PMID- 10536151 TI - Synthesis of polyacrylamides N-substituted with PNA-like oligonucleotide mimics for molecular diagnostic applications. AB - Two types of oligonucleotide mimics relative to peptide nucleic acids (PNAs) were tested as probes in nucleic acid hybridisation assays based on polyacrylamide technology. One type of mimic oligomers represented a chimera constructed of PNA and phosphono-PNA (pPNA) monomers, and the other one contained pPNA residues alternating with PNA-like monomers on the base of trans -4-hydroxy-L-proline (HypNA). A chemistry providing efficient and specific covalent attachment of these DNA mimics to acrylamide polymers using a convenient approach based on the co-polymerisation of acrylamide and some reactive acrylic acid derivatives with oligomers bearing 5'- or 3'-terminal acrylamide groups has been developed. A comparative study of polyacrylamide conjugates with oligonucleotides and mimic oligomers demonstrated the suitability and high potential of PNA-pPNA and HypNA pPNA chimeras as sequence-specific probes in capture and detection of target nucleic acid fragments to serve current forms of DNA arrays. PMID- 10536153 TI - Analysis of chemical modification of RNA from formalin-fixed samples and optimization of molecular biology applications for such samples. AB - Formalin-fixed archival samples are known to be poor materials for molecular biological applications. We conducted a series of experiments to understand the alterations in RNA in fixed tissue. We found that formalin-fixed tissue was resistant to solubilization by chaotropic agents. However, proteinase K completely solubilized the fixed tissue and enabled the extraction of almost the same amount of RNA as from a fresh sample. The extracted RNA did not show apparent degradation. However, as reported, successful PCR amplification was limited to short targets. The nature of such 'fixed' RNA was analyzed using synthetic homo-oligo RNAs. The heterogeneous increase in molecular weight of the RNAs, measured by MALDI-TOF mass spectrometry, showed that all four bases showed addition of mono-methylol (-CH(2)OH) groups at various rates. The modification rate varied from 40% for adenine to 4% for uracil. In addition, some adenines underwent dimerization through methylene bridging. The majority of the methylol groups, however, could be removed from bases by simply elevating the temperature in formalin-free buffer. This demodification proved effective in restoring the template activity of RNA from fixed tissue. The improvement in PCR results suggested that more than half of the modification was removed by this demodification. PMID- 10536152 TI - Analysis of the histone acetyltransferase B complex of maize embryos. AB - Purified histone acetyltransferase B (HAT-B) from maize consists of two subunits, p50 and p45. Cloning of the cDNA and genomic DNA encoding the catalytic subunit p50 revealed a consensus motif reminiscent of other acetyltransferases. Internal peptide sequences and immunological studies identified p45 as a protein related to the Retinoblastoma associated protein Rbap. Antibodies against recombinant p50 were able to immunoprecipitate the enzymatic activity of p50 as well as p45. Consistent with the idea that HAT-B is involved in acetylation of newly synthesized histone H4 during DNA replication, mRNA and protein levels are correlated with S-phases during embryo germination. Inhibition of histone deacetylases by HC toxin or Trichostatin A caused a decrease of the in vivo expression of HAT-B mRNA. Regardless of its predominant cytoplasmic localization, a significant proportion of HAT-B-p50 is present in nuclei, irrespective of the cell cycle stage, suggesting an additional nuclear function. PMID- 10536154 TI - Identification and characterization of a DNA primase from the hyperthermophilic archaeon Methanococcus jannaschii. AB - We report the identification and characterisation of a DNA primase from the thermophilic methanogenic archaeon Methanococcus jannaschii (Mjpri). The analysis of the complete genome sequence of this organism has identified an open reading frame coding for a protein with sequence similarity to the small subunit of the eukaryotic DNA primase (the p50 subunit of the polymerase alpha-primase complex). This protein has been overexpressed in Escherichia coli and purified to near homogeneity. Recombinant Mjpri is able to synthesise oligoribonucleotides on various pyrimidine single-stranded DNA templates [poly(dT) and poly(dC)]. This activity requires divalent cations such Mg(2+), Mn(2+)or Zn(2+), and is additionally stimulated by the monovalent cation K(+). A multiple sequence alignment has revealed that most of the regions that are conserved in eukaryotic p50 subunits are also present in the archaeal primases, including the conserved negatively charged residues, which have been shown to be essential for catalysis in the mouse primase. Of the four cysteine residues that have been postulated to make up a putative Zn-binding motif, two are not present in the archaeal homologue. This is the first report on the biochemical characterisation of an archaeal DNA primase. PMID- 10536155 TI - Pyrophosphate mediates the effect of certain tRNA mutations on aminoacylation of yeast tRNA(Phe). AB - The influence of pyrophosphate hydrolysis by inorganic pyrophosphatase on homologous aminoacylation of different yeast tRNA(Phe) mutants was studied. The addition of pyrophosphatase significantly improved the aminoacylation efficiency of tRNA(Phe) structural mutants as well as the mutant with substitution at position 20, while having no effect on the charge of wild-type tRNA(Phe). Aminoacylation of tRNA(Phe) anticodon and discriminator base (N(73)) mutants was not affected by pyrophosphatase. Activation of wild-type tRNA(Phe) transcript aminoacylation by inorganic pyrophosphatase was observed only at low Mg(2+) concentrations due to distortion of the tRNA(Phe) structure under these conditions. Our results demonstrate that pyrophosphate dissociation becomes a rate-limiting step of the reaction in yeast phenylalanyl-tRNA synthetase catalyzed aminoacylation of tRNA(Phe) variants with altered tertiary structure. A possible mechanism of pyrophosphate-mediated inhibition of tRNA mutants aminoacylation is discussed. PMID- 10536156 TI - The distribution of RNA motifs in natural sequences. AB - Functional analysis of genome sequences has largely ignored RNA genes and their structures. We introduce here the notion of 'ribonomics' to describe the search for the distribution of and eventually the determination of the physiological roles of these RNA structures found in the sequence databases. The utility of this approach is illustrated here by the identification in the GenBank database of RNA motifs having known binding or chemical activity. The frequency of these motifs indicates that most have originated from evolutionary drift and are selectively neutral. On the other hand, their distribution among species and their location within genes suggest that the destiny of these motifs may be more elaborate. For example, the hammerhead motif has a skewed organismal presence, is phylogenetically stable and recent work on a schistosome version confirms its in vivo biological activity. The under-representation of the valine-binding motif and the Rev-binding element in GenBank hints at a detrimental effect on cell growth or viability. Data on the presence and the location of these motifs may provide critical guidance in the design of experiments directed towards the understanding and the manipulation of RNA complexes and activities in vivo. PMID- 10536158 TI - Oxidative damage-induced PCNA complex formation is efficient in xeroderma pigmentosum group A but reduced in Cockayne syndrome group B cells. AB - Proliferating cell nuclear antigen (PCNA), a processivity factor for DNA polymerases delta and epsilon, is essential for both DNA replication and repair. PCNA is required in the resynthesis step of nucleotide excision repair (NER). After UV irradiation, PCNA translocates into an insoluble protein complex, most likely associated with the nuclear matrix. It has not previously been investigated in vivo whether PCNA complex formation also takes place after oxidative stress. In this study, we have examined the involvement of PCNA in the repair of oxidative DNA damage. PCNA complex formation was studied in normal human cells after treatment with hydrogen peroxide, which generates a variety of oxidative DNA lesions. PCNA was detected by two assays, immunofluorescence and western blot analyses. We observed that PCNA redistributes from a soluble to a DNA-bound form during the repair of oxidative DNA damage. PCNA complex formation was analyzed in two human natural mutant cell lines defective in DNA repair: xeroderma pigmentosum group A (XP-A) and Cockayne syndrome group B (CS-B). XP-A cells are defective in overall genome NER while CS-B cells are defective only in the preferential repair of active genes. Immunofluorescent detection of PCNA complex formation was similar in normal and XP-A cells, but was reduced in CS-B cells. Consistent with this observation, western blot analysis in CS-B cells showed a reduction in the ratio of PCNA relocated as compared to normal and XP-A cells. The efficient PCNA complex formation observed in XP-A cells following oxidative damage suggests that formation of PCNA-dependent repair foci may not require the XPA gene product. The reduced PCNA complex formation observed in CS-B cells suggests that these cells are defective in the processing of oxidative DNA damage. PMID- 10536157 TI - The human REV1 gene codes for a DNA template-dependent dCMP transferase. AB - DNA is frequently damaged by various physical and chemical agents. DNA damage can lead to mutations during replication. In the yeast Saccharomyces cerevisiae, the damage-induced mutagenesis pathway requires the Rev1 protein. We have isolated a human cDNA homologous to the yeast REV1 gene. The human REV1 cDNA consists of 4255 bp and codes for a protein of 1251 amino acid residues with a calculated molecular weight of 138 248 Da. The human REV1 gene is localized between 2q11.1 and 2q11.2. We show that the human REV1 protein is a dCMP transferase that specifically inserts a dCMP residue opposite a DNA template G. In addition, the human REV1 transferase is able to efficiently and specifically insert a dCMP opposite a DNA template apurinic/apyrimidinic (AP) site or a uracil residue. These results suggest that the REV1 transferase may play a critical role during mutagenic translesion DNA synthesis bypassing a template AP site in human cells. Consistent with its role as a fundamental mutagenic protein, the REV1 gene is ubiquitously expressed in various human tissues. PMID- 10536159 TI - Interleukin-6 repression is associated with a distinctive chromatin structure of the gene. AB - Expression of the interleukin-6 (IL-6) gene is usually tightly controlled and may be induced in specific tissues only after treatment with appropriate stimuli. The molecular mechanisms responsible for IL-6 gene repression in specific tissues or cell lines remain poorly defined. In order to address this question we have studied two human breast carcinoma cell lines, MDA-MB-231, in which the IL-6 gene is expressed, and MCF-7, in which it is not. The promoter region of the IL-6 gene was analysed in both cell lines with reference to two different parameters: (i) DNase I hypersensitivity; (ii) the in vivo pattern of DNA-protein interactions. We show herein that the mechanism responsible for silencing IL-6 gene expression in MCF-7 cells most probably involves a modification of chromatin structure, as suggested by a decreased sensitivity of the IL-6 promoter to DNase I relative to the IL-6-expressing cell line MDA-MB-231. Moreover, we show that a 'closed' nucleosomal structure in MCF-7 cells does not inhibit the binding of nuclear proteins to IL-6 gene regulatory sequences in vivo. We suggest, therefore, that, in non-expressing cells, local chromatin remodelling at the proximal promoter is inhibited by negative regulators, as suggested by two specific hallmarks of nuclear factor binding that are not observed in expressing cells: an additional in vivo footprint spanning positions -135/-119 and an additional DNase I hypersensitive site far upstream, around position -1400. Furthermore, a specific factor binding in vitro to the -140/-116 region of the IL-6 promoter is found in MCF-7 cells. PMID- 10536160 TI - Long W tracts are over-represented in the Escherichia coli and Haemophilus influenzae genomes. AB - The occurrence of DNA tracts of the three binary base combinations: R.Y, K.M and W;S has been mapped in the complete genomes of Haemophilus influenzae and Escherichia coli. A highly significant over-representation of W tracts is observed in both bacteria. The excess of W tracts is particularly striking in the 10% intercoding regions. Subdivision of intercoding regions into divergent (promoting), convergent (terminating) and sequential subregions shows that the excess of W tracts is most concentrated in the promoter regions. A particularly high excess of W tracts is observed in the first 200 bases 5' upstream of coding start sites. The data suggest that W tracts have a role in promoter function. A function as unwinding centers, analogous to the role of R.Y tracts in eukaryotes, is proposed. R.Y and K.M tracts are only modestly over-represented in the two bacteria. PMID- 10536161 TI - Molecular evolution of DNA-(cytosine-N4) methyltransferases: evidence for their polyphyletic origin. AB - DNA N4-cytosine methyltransferases (N4mC MTases) are a family of S-adenosyl-L methionine (AdoMet)-dependent MTases. Members of this family were previously found to share nine conserved sequence motifs, but the evolutionary basis of these similarities has never been studied in detail. We performed phylogenetic analysis of 37 known and potential new family members from the multiple sequence alignment using distance matrix, parsimony and maximum likelihood approaches to infer the evolutionary relationship among the N4mC MTases and classify them into groups of orthologs. All the treeing algorithms employed as well as results of exhaustive sequence database searching support a scenario, in which the majority of N4mC MTases, except for M. Bal I and M. Bam HI, arose by divergence from a common ancestor. Interestingly, MTases M. Bal I and M. Bam HI apparently originated from N6-adenine MTases and represent the most recent addendum to the N4mC MTase family. In addition to the previously reported nine sequence motifs, two more conserved sequence patches were detected. Phylogenetic analysis also provided the evidence for massive horizontal transfer of MTase genes, presumably with the whole restriction-modification systems, between Bacteria and Archaea. PMID- 10536162 TI - Nitric oxide-induced damage to mtDNA and its subsequent repair. AB - Mutations in mitochondrial DNA (mtDNA) have recently been associated with a variety of human diseases. One potential DNA-damaging agent to which cells are continually exposed that could be responsible for some of these mutations is nitric oxide (NO). To date, little information has been forthcoming concerning the damage caused by this gas to mtDNA. Therefore, this study was designed to investigate damage to mtDNA induced by NO and to evaluate its subsequent repair. Normal human fibroblasts were exposed to NO produced by the rapid decomposition of 1-propanamine, 3-(2-hydroxy-2-nitroso-1-propylhydrazino) (PAPA NONOate) and the resultant damage to mtDNA was determined by quantitative Southern blot analysis. This gas was found to cause damage to mtDNA that was alkali-sensitive. Treatment of the DNA with uracil-DNA glycosylase or 3-methyladenine DNA glycosylase failed to reveal additional damage, indicating that most of the lesions produced were caused by the deamination of guanine to xanthine. Studies using ligation-mediated PCR supported this finding. When a 200 bp sequence of mtDNA from cells exposed to NO was analyzed, guanine was found to be the predominantly damaged base. However, there also was damage to specific adenines. No lesions were observed at pyrimidine sites. The nucleotide pattern of damage induced by NO was different from that produced by either a reactive oxygen species generator or the methylating chemical, methylnitrosourea. Most of the lesions produced by NO were repaired rapidly. However, there appeared to be a subset of lesions which were repaired either slowly or not at all by the mitochondria. PMID- 10536164 TI - Inhibition of poly(ADP-ribose)polymerase stimulates extrachromosomal homologous recombination in mouse Ltk-fibroblasts. AB - Poly(ADP-ribose)polymerase (PARP) is an abundant nuclear enzyme activated by DNA breaks. PARP is generally believed to play a role in maintaining the integrity of the genome in eukaryote cells via anti-recombinogenic activity by preventing inappropriate homologous recombination reactions at DNA double-strand breaks. While inhibition of PARP reduces non-homologous recombination, at the same time it stimulates sister chromatid exchange and intrachromosomal homologous recombination. Here we report that the inhibition of PARP with 100 microg/ml (0.622 mM) 1,5-isoquinolinediol results in an average 4.6-fold increase in the frequency of extrachromosomal homologous recombination between two linearized plasmids carrying herpes simplex virus thymidine kinase genes inactivated by non overlapping mutations, in mouse Ltk-fibroblasts. These results are in disagreement with the previously reported observation that PARP inhibition had no effect on extrachromosomal homologous recombination in Ltk-cells. PMID- 10536163 TI - CUG repeat binding protein (CUGBP1) interacts with the 5' region of C/EBPbeta mRNA and regulates translation of C/EBPbeta isoforms. AB - The transcription factor CCAAT/enhancer binding protein beta, C/EBPbeta, plays a significant role in the regulation of hepatocyte growth and differentiation. A single mRNA coding for C/EBPbeta produces several protein isoforms. Two pathways for generation of low molecular weight C/EBPbeta isoforms have been described: specific proteolytic cleavage and initiation of translation from different AUG codons of C/EBPbeta mRNA. A truncated C/EBPbeta isoform, LIP, is induced in rat livers in response to partial hepatectomy (PH) via the alternative translation mechanism. Here we present evidence that CUG repeat binding protein, CUGBP1, interacts with the 5' region of C/EBPbeta mRNA and regulates translation of C/EBPbeta isoforms. Two binding sites for CUGBP1 are located side by side between the first and second AUG codons of C/EBPbeta mRNA. One binding site is observed in an out of frame short open reading frame (sORF) that has been previously shown to regulate initiation of translation from different AUG codons of C/EBPbeta mRNA. Analysis of cytoplasmic and polysomal proteins from rat liver after PH showed that CUGBP1 is associated with polysomes that translate low molecular weight isoforms of C/EBPbeta. The binding activity of CUGBP1 to the 5' region of C/EBPbeta mRNA shows increased association with these polysomal fractions after PH. Addition of CUGBP1 into a cell-free translation system leads to increased translation of low molecular weight isoforms of C/EBPbeta. Our data demonstrate that CUGBP1 protein is an important component for the regulation of initiation from different AUG codons of C/EBPbeta mRNA. PMID- 10536165 TI - Analysis of chromatin in limited numbers of cells: a PCR-SSCP based assay of allele-specific nuclease sensitivity. AB - Chromatin can be analysed by assaying its sensitivity to DNase I or other nucleases in purified nuclei. Usually, this is performed by Southern analysis of genomic DNA extracted from nuclease-treated nuclei, a methodology that requires many cells. Applying restriction fragment length polymorphisms (RFLPs), this methodology has been used for parental allele-specific chromatin studies on imprinted mammalian genes. However, such allelic studies are limited by the availability of suitable RFLPs. We therefore developed an alternative, PCR and single strand conformation polymorphism (SSCP)-based assay with which allelic sensitivity to nucleases can be determined in virtually all localised regions that have nucleotide polymorphisms. We also demonstrate that analysis of DNase I sensitivity can be performed on permeabilised cells. Combining the two approaches, in the imprinted mouse U2af1-rs1 gene we analysed parental allele specific chromatin conformation in limited numbers of cultured cells. We also applied the PCR-SSCP approach to assay allelic DNA methylation at specific restriction enzyme sites. In summary, we developed an allele-specific assay that should be useful for biochemical and developmental investigation of chromatin, in particular for studies on genomic imprinting and X-chromosome inactivation. PMID- 10536166 TI - Normalization of array hybridization experiments in differential gene expression analysis. AB - For detecting and confirming differentially expressed genes it is necessary to have a trustworthy reference. So called 'housekeeping genes' are frequently used for this purpose as internal standard. However, if the influence of new experimental conditions is to be analyzed it is not safe to assume a priori that the expression of these genes is not affected. Therefore two synthetic poly(A) RNAs were generated by PCR and in vitro transcription. They were used as external standards for normalization of northern blots and cDNA arrays where non-regulated genes as internal reference were not available. PMID- 10536167 TI - Differences in induction of p53, p21WAF1 and apoptosis in relation to cell cycle phase of MCF-7 cells treated with camptothecin. AB - The DNA topoisomerase I (topI) inhibitor camptothecin (CPT), stabilizes so-called cleavable complexes which consist of topI covalently attached to 3' OH ends of DNA nicks. Collisions between the progressing DNA replication forks (occurring in S phase cells) or between the transcription driven RNA polymerase molecules (occurring in G1, S and G2 cells) and these complexes convert the latter into secondary DNA lesions which are unrepairable and lethal to the cell. Changes induced by CPT in the level of the tumor suppressor p53, cyclin-dependent kinase inhibitor p21WAF1 and proapoptotic protein Bax (all detected immunocytochemically), were measured separately in the nucleus and cytoplasm of individual human breast carcinoma MCF-7 cells by laser scanning cytometry (LSC) in relation to cell cycle position and induction of apoptosis. The initial transient cell arrest at the G1 checkpoint seen at 8-16 h of treatment with 0.15 microM CPT was accompanied by the rapid accumulation of p53 (preventable by cycloheximide) in the nucleus; the rise (>20-fold) in p53 was maximal for S phase cells. The magnitude of the nuclear p53 increase induced by CPT, at maximum, was 2-fold higher than that induced by the proteasome inhibitor N-acetyl-Leu-Leu norleucinal (LLnL). While the accumulation of p53 was seen in all phases of the cycle, only G1 cells responded by induction ( approximately 60-fold increase) of p21WAF1. Inhibition of DNA replication by aphidicolin prevented the accumulation of p53 in S and G2/M but had no effect on its induction in G1 cells. Perturbation of cell progression through S phase was seen between 24-72 h of treatment, and it coincided with induction of Bax and apoptosis (both maximal in S phase cells). Thus, the changes observed in S phase cells (nuclear accumulation of p53 preventable by aphidicolin, induction of Bax, apoptosis), triggered by the collisions of DNA replication forks with the CPT-induced lesions, were distinct from the changes in G1 (nuclear p53 accumulation unaffected by aphidicolin, induction of p21WAF1) presumably triggered by collisions of RNA polymerase with the CPT-lesions. Great heterogeneity in expression of p53 and p21WAF1 of the G1 cell population in response to CPT was observed, which may reflect the intercellular variability in the rate of transcription (i.e., frequencies of collisions of RNA polymerase with the lesions). Thus, differences in the transcriptional activity of G1 cells may play a role in their sensitivity to CPT and similar topI inhibitors. PMID- 10536168 TI - Treatment with tumor-reactive Fab-IL-2 and Fab-staphylococcal enterotoxin A fusion proteins leads to sustained T cell activation, and long-term survival of mice with established tumors. AB - C215Fab-IL-2 fusion protein, with full IL-2 and antigen binding activity, was produced in E. coli at high level (>50 mg/l). When co-administered with Fab superantigen fusion protein (C215Fab-SEA) in mice strong and sustained T cell activation was observed. Combination treatment of mice carrying B16 melanoma transfected with C215 antigen was also more efficient than using C215Fab-SEA (p<0.01) or C215Fab-IL-2 alone (p<0.001). In a long-term survival experiment 5/12 mice having received combination treatment 5 days after i.v. inoculation of B16 cells survived >85 days. Improved therapeutic efficacy correlated with increased tumor infiltration by activated CD25+ T cells, indicating a T cell mediated mechanism. PMID- 10536169 TI - Constitutive expression of Heregulin induces apoptosis in an erbB-2 overexpressing breast cancer cell line SKBr-3. AB - We have previously reported that Heregulin (HRG)/neu differentiation factor (NDF) induced growth arrest and cellular differentiation in breast cancer cells overexpressing erbB-2 receptor. To elucidate the cellular mechanisms underlying the growth inhibition by HRG, we developed an in vitro model by transfection of HRG cDNA into the erbB-2 overexpressing breast cancer cell line, SKBr-3. We showed that the enforced expression of HRG in SKBr-3 cells induces dramatic morphological changes and pronounced inhibition of anchorage-dependent and independent growth. Most SKBr-3/HRG-transfected (SK/HRG) cells exhibited about 15 fold increase in size and persisted as multinucleated cells with extended flattened vacuole-filled cytoplasm with reduced cell attachment. The growth suppression of SK/HRG cells was accompanied by a reduction in S phase, the presence of a G2-M cell cycle delay, and an increase in DNA aneuploid components. In addition, DNA fragmentation assays showed that HRG induced apoptosis of SKBr-3 cells. In contrast, while HRG treatment of other erbB-2 overexpressing breast cancer cell lines led to growth arrest and cell detachment, it did not induce apoptotic features. Thus, this study demonstrates that while growth arrest and cell detachment are general effects of HRG towards erbB-2 overexpressing cells, the ability of HRG to induce apoptosis is a phenomenon confined to selective cells including SKBr-3 cells. PMID- 10536170 TI - Co-overexpression of p53 protein and epidermal growth factor receptor in human papillary thyroid carcinomas correlated with lymph node metastasis, tumor size and clinicopathologic stage. AB - Expressions of p53 protein and epidermal growth factor receptor (EGFR) were immunohistochemically investigated in 111 patients with papillary thyroid carcinomas (PTC) in order to evaluate their co-expression in relation to lymph node metastases (LNM), tumor size and clinicopathologic stage. In PTC, positive staining for p53 in dewaxed sections was present in nuclei or cytoplasm, or in both, whereas surface linear or cytoplasmic staining for EGFR was observed with varying degrees of extent and intensity. Positive reaction (more than 10% of tumor cells positive) was observed in 65 cases (58. 5%) for p53, and in 87 cases (78.4%) for EGFR. A significant correlation was found between p53 protein and EGFR overexpressions (p<0.01). Notably, p53-positive cases always exhibited positive staining for EGFR. Forty-four patients (39.6%) exhibited concomitant LNM, most of whom had both p53 and EGFR expression in primary foci. Statistical analysis revealed that co-expression of p53 protein and EGFR was significantly correlated with LNM, tumor size and clinicopathologic stage, but no correlation was found between their co-expression and age or sex. Our findings suggest that overexpression of p53 protein or EGFR in PTC tends to be associated with a high frequency of LNM, increased tumor size and advanced clinicopathologic stage, and that co-expression of both p53 protein and EGFR may predispose to growth and progression of PTC. Our findings also suggest that p53 protein and EGFR expressions may be clinicopathologic and prognostic indicators of PTC. PMID- 10536171 TI - Prospective long-term endoscopic and histologic follow-up of gastric lymphoproliferative disease of early stage IE low-grade B-cell mucosa-associated lymphoid tissue type following Helicobacter pylori eradication treatment. AB - Long-term endoscopic and histologic follow-up of Stage IE gastric lymphoproliferative disease of the low-grade B-cell mucosa-associated lymphoid tissue (MALT) type following cure of H. pylori was undertaken. Clinical and endoscopic features (age, race, endoscopic appearance, cure of H. pylori and duration of follow-up) were also evaluated as potential prognostic indicators for complete or near-complete regression of low-grade MALT lymphoma. Sixty-eight MALT lymphoma patients prospectively underwent H. pylori eradication. follow-up at periodic intervals with endoscopy and extensive mucosal biopsy protocol on 65 patients ranging from 12 weeks up to 73 months (mean +/- SD of 22.5+/-15.8 months) has been completed. H. pylori was eradicated in 89.2% of MALT lymphoma patients with complete histologic regression noted in 58.5%, near-complete regression in 18.5%, partial in 4.6%, and no change in 18.5%. Univariate analysis revealed two factors predictive of complete and/or near complete MALT lymphoma regression, H. pylori cure (p=0. 001) and duration of follow-up (p=0.001). Stepwise logistic regression also demonstrated that both H. pylori cure (p<0.0001) and duration of follow-up (p<0.02) were independently associated with complete and near complete MALT lymphoma regression. Age, race, and endoscopic appearance were not predictive of regression. We conclude that this lymphoproliferative disease predictably undergoes complete to near-complete histologic regression at variable rates following cure of H. pylori in a majority of patients. PMID- 10536172 TI - NK binding capacity and lytic activity depend on the expression of ICAM-1 on target bone tumours. AB - Osteosarcoma cell lines are differently lysed by natural killer (NK) lymphocytes. A critical step in the lytic process is the recognition and attachment of effector to target cells. To determine binding capacity and lytic activity of NK cells, we investigated the distribution and role of ICAM-1, 2 and 3 on two osteosarcoma cell lines (HOS and Saos-2) in basal conditions and after TNFalpha treatment. Modulation of ICAM-1 after TNFalpha treatment modified the binding capacity of NK cells to osteosarcoma target cells. This modulation process appears to play a critical role in determining the susceptibility of these cells to NK-mediated lysis. PMID- 10536173 TI - EXTL3/EXTR1 alterations in colorectal cancer cell lines. AB - We previously demonstrated that metastasis-related tumor suppressor gene(s) may exist on chromosome 8p21-22 on allelotype analysis of early colorectal carcinomas (CRC) with lymph node metastasis. Here, we searched for target gene(s) in this chromosomal region in the UniGene database. The EXTL3 (also called EXTR1) gene was selected as a candidate because of its homology to EXT1 and EXT2, putative tumor suppressor genes. We screened 12 CRC cell lines for mutations by means of polymerase chain reaction (PCR)-single strand conformation polymorphism. Three cell lines showed EXTL3 mutations, all of which were located within exon 3 and caused amino acid substitutions. Reverse transcription-PCR analysis showed that the EXTL3 expression was lacking in 1 of the 12 colorectal cancer cell lines. Although there is still no definitive evidence that EXTL3 is a tumor suppressor gene for CRC, these data suggest that inactivation of the EXTL3 gene may at least offer a selective growth advantage for some CRC cell lines. PMID- 10536175 TI - Comprehensive analysis of p53 gene mutation characteristics in lung carcinoma with special reference to histological subtypes. AB - To date, the characteristics of p53 gene mutations in lung cancer have been extensively investigated. However, current estimates of p53 alterations are inaccurate, since most investigators have limited their analyses to exons 5 to 8 of the p53 gene. We examined 52 lung carcinoma cell lines and 106 primary non small cell lung carcinomas (NSCLC) for mutations in the entire coding region of the p53 gene, from exons 2 to 11. High resolution single strand conformation polymorphism analysis was performed using a modified electrophoretic apparatus with a high concentration gel (14%) and accurate temperature control. The prevalence of mutations was high (more than 80%) in both small cell lung carcinoma (SCLC) (15 of 18) and NSCLC cell lines (28 of 34), and 9 of 45 mutations (20%) were detected outside the region of exons 5 to 8. The frequency of the mutations in primary NSCLC was 48% (51 of 106) and was significantly different (p=0.01) between adenocarcinoma (39%) and squamous cell carcinoma (67%). A-->G transitions (14%, 6 of 43 cases) as well as G-->T transversions (26%, 11 of 43 cases) were frequently detected with significant strand bias in smoking patients, suggesting that carcinogens causing these mutations are involved in smoking associated lung carcinogenesis. PMID- 10536174 TI - A phase II trial of low dose administration of 5-fluorouracil and cisplatin in patients with advanced and recurrent gastric cancer. AB - The clinical efficacy of chemotherapy for patients with advanced gastric cancer has not been established. We investigated in a phase II study the combination chemotherapy of low dose 5-fluorouracil (5-FU) and cisplatin (low dose FP) to evaluate its clinical efficacy in terms of response, quality of life and survival in 43 gastric cancer patients including 29 advanced and 14 recurrent cases. The administration of 5-FU was done by continuous intravenous infusion for 28 consecutive days with a dose of 250 mg/m2, while the administration of cisplatin was done by 1-h intravenous drip-infusion for 5 consecutive days and 2-day intervals per week with a dose of 3.5 mg/m2, that was repeated for 4 weeks in one cycle. The response rate in advanced cases was 48.3%, evaluated in 14 cases of partial response (PR), whereas its response rate in recurrent cases was 35.7%, evaluated in 5 cases of PR. The most effective lesions for low dose FP chemotherapy were primary lesion and lymph node metastasis. The quality of life assessed by performance status and oral intake was improved in 13.8% and 37.9% of advanced cases, and 21.4% and 28.6% of recurrent cases, respectively, as compared to those of pretreatment. The median survival time and 1-year survival rate were 6 months, 21.6% in advanced cases and 10 months, 28.8% in recurrent cases, respectively. The major adverse effect was observed in gastrointestinal toxicity and leukopenia, and the all toxicities were less than grade 2, that were controllable during the treatment. These results indicated that the combination chemotherapy of low dose administration of 5-FU and cisplatin might have therapeutic efficacy in tumor response and offer improvement in quality of life in patients with advanced and recurrent gastric cancer. PMID- 10536176 TI - Wild-type p53 gene transfer resulted in cell cycle arrest, but not apoptosis of newly established human malignant fibrous histiocytoma cell line. AB - We established a human malignant fibrous histiocytoma (MFH) cell line, MFH-ToE, from a tumor originally developed in the right thigh of a 78-year-old woman. The original tumor histologically consisted of histiocytic, fibroblastic and giant cells. The tumor cells showed immunoreactivity for vimentin and alpha-1 antichymotrypsin, and were positive for acid phosphatase and non-specific esterase, being compatible with MFH. Although the histology of the heterotransplanted tumor into nude mice was similar to that of the primary MFH, the population of giant cells gradually decreased along with the culture passages. Cytogenetic analysis revealed a highly aneuploid nature with varying numbers of chromosomes from 71 to 140. Chromosome 17 showed monosomy and exon 6 to 8 of p53 gene was not amplified by PCR, implying absence of p53 function. Adenovirus vector-mediated wild-type p53 gene was successfully transfected into the MFH-ToE, which showed up-regulation of P53 and P21, as well as gradual up regulation of Bcl-2 protein. The transfection resulted in cell cycle arrest, but not apoptosis of the MFH-ToE cells. These results revealed unique properties of the MFH-ToE, which might be useful in further studies analyzing pathological and biological characteristics of MFH. PMID- 10536177 TI - Loss of tumorigenicity of human breast cancer cells engineered to produce IL-2, IL-4 or GM-CSF in nude mice. AB - Human breast cancer cells (OCUB-M), retrovirally transduced with granulocyte macrophage-colony stimulating factor (GM-CSF), interleukin-2 (IL-2) or IL-4 gene were examined for their antitumor activities in nude mice. Although cell proliferation rates in vitro of these cytokine-producing cells were not significantly different from that of wild-type cells, nude mice that were subcutaneously inoculated with cytokine-producing cells did not develop tumors in contrast to mice that were injected with wild-type cells. Injection of GM-CSF producing cells into the vicinity of growing wild-type tumors retarded subsequent growth of wild-type tumors. Histological examination of tumors which received GM CSF-producing cells revealed marked infiltration of mononuclear cells around the tumors. Irradiation of cytokine-producing cells diminished their proliferation capacity but production of cytokine(s) was retained. Therefore, inoculation of irradiated cytokine producer cells into growing tumors can be used as a therapeutic maneuver for breast cancer. PMID- 10536178 TI - Mutation analysis of the Smad3 gene in human ovarian cancers. AB - The Smad3 gene is a member of the Smad family, vertebrate homologues of Drosophila Mad, and its gene product is a cytoplasmic element in the transforming growth factor-beta (TGF-beta) signaling pathway. Mutations in TGF-beta receptors and their cytoplasmic elements of transduction signals commonly accompany various cancers. Using PCR-SSCP analysis we searched for the presence of Smad3 gene mutations in 36 human ovarian cancers, and found that 15 cases (41. 7%) had a polymorphism at codon 103. Because this mutation was not accompanied by amino acid replacement, the present results show that the mutations in the Smad3 gene are unlikely to be involved in human ovarian cancers. PMID- 10536179 TI - Resveratrol pretreatment desensitizes AHTO-7 human osteoblasts to growth stimulation in response to carcinoma cell supernatants. AB - Resveratrol, a natural phytoestrogen, has been reported to promote differentiation of murine MC3T3-E1 osteoblasts and to inhibit proliferation of prostate cancer cell lines. In the present study we tested the effects of resveratrol on the increased proliferation of human AHTO-7 osteoblastic cell line induced by conditioned media (CM) from a panel of carcinoma cell lines. This compound was found to modulate AHTO-7 proliferation in a tamoxifen-sensitive mechanism at lower concentrations, but failed to induce the osteoblast differentiation marker alkaline phosphatase (ALP) in contrast to vitamin D3. The proliferative response of AHTO-7 cells to conditioned media from carcinoma cell lines was diminished (30-71.4% inhibition) upon pretreatment with 0.5 microM resveratrol. Highest inhibition was demonstrated for pancreas (BxPC3, Panc-1), breast (ZR75-1) and renal (ACHN) carcinoma cell line supernatants whereas the effect on colon carcinoma (SW620, Colo320DM) cell CM and prostate cancer (PC3, DU145 and LNCaP) CM was less pronounced. Direct addition of resveratrol affected only supernatants of cell lines (<25% inhibition) exhibiting growth stimulatory activity for normal WI-38 lung fibroblasts. Resveratrol inhibited proliferation of DU145 and LNCaP cells in concentrations exceeding 5 microM, altered cell cycle distribution of all prostate cancer cell lines in concentrations as low as 0.5 microM, but did not inhibit the production of osteoblastic factors by these lines. In conclusion, resveratrol failed to induce ALP activity as marker of osteoblast differentiation in human osteoblastic AHTO-7 cells, however, inhibited their response to osteoblastic carcinoma-derived growth factors in concentrations significantly lower than those to reduce growth of cancer cells, thus effectively modulating tumor - osteoblast interaction. PMID- 10536180 TI - Telomerase activity in non-small cell lung carcinomas correlates with smoking status. AB - Human telomerase is a ribonucleoprotein DNA polymerase which maintains the telomeric region of human chromosomes and has been detected in all types of human cancer tested. We used the telomeric repeat amplification protocol (TRAP) assay to examine 71 non-small cell lung carcinomas (NSCLC) and their adjacent normal tissue. Telomerase activity was detected in 61 (86%) of the 71 NSCLC examined but not in any of the matched normal lung tissues. A significant correlation was found between the presence of telomerase activity and current smoking status at the time of diagnosis (p=0. 0076). In addition, a trend was found between telomerase activity and smoking exposure (p=0.06). Our findings demonstrate that telomerase activity is a common phenomenon in NSCLC cases but not in the normal lung. However, certain cases in former smokers may follow a telomerase independent pathway. PMID- 10536182 TI - The rate of homozygous CDKN2A/p16 deletions in glioma cell lines and in primary tumors. AB - The rate of homozygous deletions of CDKN2A/p16 is variable between different tumor entities, and in addition it is higher in established cell lines in comparison with primary tumors. Such incongruencies may reflect statistical sampling errors, true differences depending on tissue derivatisation and CDKN2A/p16 loss under selective pressure in tissue culture. Clarification of these issues is warranted in the context of defining tumor suppressor genes such as CDKN2A/p16 as targets for gene replacement therapies. We therefore compared established cell lines derived from human glioblastomas and their corresponding primary tumors by multiplex PCR methodology. Archival early passages were included to determine the time point at which the p16 status of a cell line changes if it is different from the original tumor. It was found that in 2 of 11 cases (18%) the primary tumor had no p16 alteration whereas the corresponding cell lines had a homozygous p16 deletion. Tracking the in vitro evolution of these two cell lines we found that CDKN2A/p16 was lost already in the earliest passages. This suggests a clonal outgrowth advantage of a subpopulation of p16 deleted tumor cells rather than instability of the CDKN2A/p16 genotype in vitro. Including 20 additional glioblastoma-derived cell lines we detected that in 19 of the total 31 lines at least one exon was lost bringing the rate of p16 loss in the whole panel to 61%. This compares to a rate of 49% which was found in original glioma tissue from 47 unselected other patients. It is concluded, that in cell culture selective pressure favours the outgrowth of pre-existing CDKN2A/p16 negative clones, which account for the difference of CDKN2A/p16 status between cell lines and tumors. In no case did we see a change of the CDKN2A/p16 status during prolonged tissue culture periods of up to 8 years. PMID- 10536181 TI - Peroxisome proliferator-activated receptor gamma activation in human breast cancer. AB - The peroxisome proliferator-activated receptor gamma (PPARgamma) is a member of the nuclear receptor family of ligand-activated transcription factors. This study was designed to evaluate ligand activation of PPARgamma in human breast cancer cells. DNA binding by endogenous PPARgamma in gel shift assays and activation of PPARgamma by prostanoid and thiazolidinedione ligands in reporter gene assays differed between the cell lines. The PPARgamma ligands elicited an anti proliferative effect in MTT proliferation assays. Our data point to a variable, cell-specific response to different gamma-ligands, which holds significance for further studies on the role of PPARgamma in mediating breast cancer growth and progression. PMID- 10536184 TI - Antisense oligodeoxynucleotide against c-myc mRNA induces differentiation of human hepatocellular carcinoma cells. AB - We have previously demonstrated that the human liver-specific antigen (HLSA) expression was enhanced and c-myc levels were reduced during sodium butyrate induced differentiation in human hepatoma cells. To further elucidate a linkage between the reduction of c-myc levels and an increase in the HLSA expression, antisense oligodeoxynucleotide against c-myc mRNA was transferred into human hepatoma cells. Human hepatoma cell lines, HCC-M, HCC-T and PLC/PRF/5 were transfected with antisense oligodeoxynucleotide and changes in the cell cycle, expression of the HLSA, albumin, and alpha-fetoprotein were examined. Antisense oligodeoxynucleotide was successfully induced into cells visualized by a confocal microscope using fluorescein-labeled oligodeoxynucleotides, and Northern blot analysis revealed that c-myc expression was reduced three and six hours after the transfection. Following these changes, cell proliferation was inhibited and flow cytometric analysis showed that cell number in the G1 phase significantly increased. Increased expression of the HLSA and albumin, and decreased expression of alpha-fetoprotein was observed by flow cytometry in accordance with those changes. These results showed similar changes to those induced by butyrate treatment obtained in our previous studies. The present study indicates that the reduction of c-myc transcription increases HLSA expression levels through intracellular changes similar to those induced by butyrate, a differentiation inducer. PMID- 10536183 TI - Homozygous deletion of the p16/MTS-1/CDKN2 gene in malignant gliomas is infrequent among Japanese patients. AB - We have analyzed the status of the p16/MST-1/CDKN2 gene in 63 brain tumors from Japanese patients. With quantitative multiplex polymerase chain reaction (PCR) assay using the exon 2 primers of the p16 gene and control chromosome 9qSTS primers, we found homozygous deletion of the p16 gene in 7 cases; in 1 out of 10 cases of anaplastic astrocytomas (WHO grade III), 6 out of 35 cases of glioblastoma multiformes (grade IV) but in none of the tumors of grade I or II. We also found mobility-shifted PCR products in 8 cases using the single-strand conformation polymorphism technique. DNA sequencing of the aberrantly migrated products revealed that 5 cases of glioblastoma multiforme had mutations which caused amino acid substitutions. We found one case with silent mutations and two cases with nucleotide changes in the non-coding region. The frequency of the alteration of the p16 gene, either homozygous deletion or mutation accompanied with amino acid substitutions, increased in malignant brain tumors (grade III and IV) compared with that in low grade tumors (grade I and II) (p=0.0275), suggesting possible role(s) of the gene in the progression of brain tumors. In addition, the low frequency of homozygous deletions shown in this study is quite different from previous reports that demonstrated frequently deleted p16 gene in malignant gliomas from Caucasian patients. We have also shown the presence of heterogeneous cell populations within the glioblastoma masses based on the variety of the mutated p16 sequences. The present study, therefore, suggests a possible racial difference in the mechanism of the tumorigenesis and a heterogeneity of malignant gliomas developed during the tumor progression. PMID- 10536185 TI - Bovine seminal ribonuclease selectively kills human multidrug-resistant neuroblastoma cells via induction of apoptosis. AB - Bovine seminal ribonuclease (BS-RNase) is a homologue of RNase A with specific antitumor activity. The cytotoxic action of this agent was examined in human neuroblastoma (NB) cell lines (SK-N-SH and UKF-NB-4) possessing the multidrug resistance (MDR) phenotype and NB cell lines (IMR-32, UKF-NB-1, UKF-NB-2 and UKF NB-3) without MDR. Although MDR cells expressed large amounts of mdr-1 mRNA, contained functional P-glycoprotein and had 20- to 105-fold lower sensitivities to doxorubicin and vincristine than cells with non-MDR phenotypes, BS-RNase was equally toxic to all NB cells at concentrations employed (0.2 to 100 microg/ml). BS-RNase showed high selectivity for NB cells and was non-toxic to normal fibroblasts and epithelial cells. Ultrastructural investigation and annexin V assay showed that BS-RNase is a powerful inductor of apoptosis. The antitumoral effects of BS-RNase were also demonstrated in vivo using established subcutaneous xenografts in athymic (nude) mice of the MDR-1-bearing UKF-NB-4 cell line. Intratumoral injections (12.5 mg/kg) of BS-RNase over four weeks resulted in complete tumor regression and absence of tumor regrowth over a two-week observation period after cessation of treatment. The results show that BS-RNase selectively kills NB cells by inducing apoptosis and that this agent is active against mdr-1 expressing cells both in vitro and in vivo. BS-RNase fulfills important criteria for a candidate antitumor agent in NB patients with advanced disease. PMID- 10536187 TI - Preclinical evaluation of 90Y-labeled anti-CD20 monoclonal antibody for treatment of non-Hodgkin's lymphoma. AB - A high-affinity IgG1 kappa murine monoclonal anti-CD20 antibody (IDEC-2B8) was developed for radioimmunotherapy of non-Hodgkin's B-cell lymphoma. A stable immunoconjugate (Zevalintrade mark) was prepared by reacting IDEC-2B8 with a derivative of diethylenetriaminepentaacetic acid, designated MX-DTPA, a chelator exhibiting high affinity and retention for 90Y. Zevalin exhibited antigen specificity, human tissue reactivity, and preclinical safety profile comparable to the native antibody. The conjugate radiolabeled with 90Y (90Y-Zevalin) or 111In (111In-Zevalin) exhibited excellent retention of immunoreactivity with radioincorporations >95%. The radiolabeled conjugates formulated in PBS containing human serum albumin were stable in vitro at 4 degrees C for 48 h as indicated by negligible loss of radioisotope and retention of binding to CD20+ cells. In vitro human serum stability studies at 37 degrees C with 90Y-Zevalin indicated that loss of 90Y from the conjugate was minimal, averaging 1% per day. Biodistribution studies in BALB/c mice confirmed the in vitro stability of 90Y Zevalin and 111In-Zevalin. In particular, excellent in vivo retention of 90Y by the conjugate was demonstrated by minimal bone accumulation (4h) (the area below basal glucose level) and C(nadir) (the plasma glucose levels at the nadir) than did the solid and liquid suppositories without sodium salicylate in rats, indicating that the insulin from liquid suppository with sodium salicylate could be well absorbed in rats due to the absorption enhancing effect of sodium salicylate. It is concluded that thermo-reversible insulin liquid suppository [insulin/P 407/P 188/polycarbophil/sodium salicylate (100 (IU/g)/15/20/0.2/10%)], which was easy to administer without any pain during insertion and remained at the administered sites, could have a potential to be developed as a more convenient, safe and effective rectal delivery system of insulin. PMID- 10536243 TI - Correlation of in vitro and in vivo paracetamol availability from layered excipient suppositories. AB - An in vivo investigation of paracetamol availability was carried out on eight healthy volunteers, comparing two paracetamol suppository formulations prepared using two different gliceride bases, a fast drug-releasing one and a slow drug releasing one, i.e. Witepsol H15 and W35, respectively. The formulations were selected on the basis of a previous in vitro drug release study, which showed that, by superimposing the excipients in two layers within the same suppository, the drug release kinetics could be modulated using different ratios between the two layers. The comparison between the two different formulations in terms of plasma profiles and total amounts of drug excreted in urine revealed an increase in the extent of drug absorption from the layered excipient suppository. As the W35 has a higher monoglyceride content than the H15, this improved paracetamol availability could be ascribed to the absorption-enhancing effect of the monoglycerides. Moreover, the W35 has also a higher viscosity, which could possibly cause the suppository to be retained for a longer time in the lower part of the rectum, where the blood is drained directly to the systemic circulation. It was therefore hypothesized that the enhanced paracetamol availability could be also due to a liver bypass mechanism. For a further examination of the paracetamol absorption kinetics after rectal administration, a one-compartment model was fitted to the drug plasma concentration data. This approach allowed to draw absorption versus time profiles, which showed that a retardation actually occurred in paracetamol absorption when using suppositories containing the slow drug releasing excipient W35. These absorption data were then employed for an A level in vitro-in vivo correlation testing, and a linear relationship was found between in vitro release rate and in vivo absorption rate, both for fast releasing and for the layered excipient suppositories. PMID- 10536244 TI - Preparation and in vitro characterization of HSA-mPEG nanoparticles. AB - Surface modified human serum albumin (HSA) nanoparticles with a size of approximately 150 nm in diameter were prepared from a PEG-HSA conjugate, methoxy polyethylene glycol modified human serum albumin (HSA-mPEG) using a coacervation method and crosslinked with glutaraldehyde. The zeta-potential of the surface modified nanoparticles was significantly lower than that of unmodified HSA nanoparticles. The existence of a hydrated steric barrier surrounding the nanoparticles was confirmed by electrolyte and pH induced flocculation tests. The surface modified nanoparticles showed a reduced plasma protein adsorption on the particle surface compared with unmodified particles. PMID- 10536245 TI - Biodegradable cross-linked starch/protein microcapsules containing proteinase inhibitor for oral protein administration. AB - The objective of this study is to demonstrate the feasibility of microcapsules containing a protein and a proteinase inhibitor in order to allow the oral administration of proteic or peptidic drug. Starch/bovine serum albumin mixed walled microcapsules were prepared using interfacial cross-linking with terephthaloyl chloride. The microcapsules were loaded with native or amino protected aprotinin by incorporating protease inhibitors in the aqueous phase during the cross-linking process. Microcapsules can be degraded in the presence of alpha-amylase. The influence of the formulation parameters on the in vitro release of the inhibitor activity and the protein was studied. The protective effect of microcapsules with aprotinin for bovine serum albumin was revealed in vitro. The presence of the native bovine serum albumin was demonstrated after incubation of the microcapsules with aprotinin in a mixture of alpha-amylase (5.4 U/ml) and trypsin (900 spectrophotometric BAEE units/ml) for 3 h at 37 degrees C, whereas the protein was completely degraded in the release medium of the microcapsules without aprotinin. PMID- 10536246 TI - Morning versus evening dosing of ibuprofen using conventional and time-controlled release formulations. AB - Many functions of the human body vary considerably during a day. These variations can lead to changes in drug plasma concentrations. In the study on healthy volunteers described here it was determined whether ibuprofen plasma levels following single oral doses of immediate-release and press-coated time-controlled release tablet formulations depend on time of drug administration (08:00 or 22:00 h). The difference between morning and evening dosing of the immediate-release formulation was minimal. The results with the press-coated formulation were unexpected having regard to results of previous studies on non-steroidal anti inflammatory analgesics. Time to peak concentration was 6 h after morning administration, 4 h after evening administration. Both the rate and extent of bioavailability of ibuprofen were lower when dosing took place at 08:00 h than when dosing took place at 22:00 h. The influence of food on the pharmacokinetic profile of an evening dose of the press-coated formulation was also studied. When tablets were administered with a meal the ratio C(max)/AUC and t(max) and AUC values indicated that bioavailability was reduced. The main conclusion was that the chronopharmacokinetic behaviour of the press-coated ibuprofen tablet is related to the formulation, not the drug substance as such. PMID- 10536247 TI - Percolation theory, conductivity and dissolution of hydrophilic suppository bases (PEG systems). AB - We investigated the conductivity of binary mixtures of different volume to volume ratios of liquid polyethylene glycol 200 (PEG 200) and solid PEG 6000. The conductivity was measured using a specially designed cell, which could be filled with the sample to be analysed. The results were analysed by the help of the percolation theory and its basic equation: X=S(p-p(c))(q) with X=gamma the conductivity and X=k the dissolution rate constant of binary mixtures with p the concentration of PEG 200 including Mepyraminmaleat as a marker substance. The intrinsic dissolution rate was determined spectrophotometrically. The simultaneous determination of the critical exponent q and the percolation threshold p(c) of the conductivity experiment yielded the following values q=1.83+/-0.05 and p(c)=11.6+/-1.1% with a squared correlation coefficient R(2)=0. 9986. For q=mu=2.00 fixed the percolation threshold is equal to p(c)=8.4+/-1.3% with R(2)=0.9984. Thus if the same percolation threshold p(c)=8.4+/-1.3% is adopted for the dependence of the dissolution process q becomes equal to q approximately mu=1. 94+/-0.045. R(2)=0.9991. This result is in excellent agreement with the theoretical prediction that the permeability of a porous network scales in the same way as the conductivity. PMID- 10536248 TI - Influence of the technological parameters of ultrasonic nebulisation on the nebulisation quality of alpha1 protease inhibitor (alpha1PI). AB - The principle of an ultrasonic nebuliser is based on the vibrations of a piezo electric crystal driven by an alternating electrical field. These periodical vibrations are characterised by their frequency, their amplitude and their intensity which corresponds to the energy transmitted per surface unit. When the vibration intensity is sufficient, cavitation appears and generates droplets. Ventilation enables an airflow to cross the nebuliser and to expulse the aerosol droplets. For a given nebuliser, the vibration frequency of the piezo-electric crystal is fixed and is often in the range of 1-2.5 MHz. In most cases, an adjustment in vibration intensity is possible by modifying vibration amplitude. The ventilation level is adjustable. The influence of these two parameters on the efficiency of ultrasonic nebulisation is studied. The study was carried out with a protein solution that had to be administered into the lungs. The solution used presented a viscosity of 1.25 mPa and a surface tension of 53 mN/m. The integrity of the protein was checked which was submitted to different vibration conditions. Nebulisation efficiency was evaluated by determining droplet size, the percentage of drug nebulised and nebulisation time. An increase in vibration intensity does not modify the size of droplets emitted, but decreases nebulisation time and raises the quantity of protein nebulised, thus improving performance. On the other hand, an increase in ventilation increases the size of droplets emitted, decreases nebulisation time and the quantity of protein nebulised because more drug is lost on the walls of the nebuliser. High intensity associated with low ventilation favours drug delivery deep into the lungs. PMID- 10536249 TI - Inhalation of tobramycin in cystic fibrosis. Part 1: the choice of a nebulizer. AB - Forteen commercially available jet and ultrasonic nebulizers were investigated with the aim to select the most suitable type of apparatus for the inhalation of a 10% tobramycin solution. Two different techniques for measurement of particle size distribution were evaluated: laser diffraction and cascade impactor analysis. The final selection of the nebulizers is based on particle size distribution, output and stable performance during nebulization. All 14 nebulizers (eight jet and six ultrasonic) were filled with a solution of 10% m/v tobramycin (as sulphate) in water. The volume in the tested devices ranged from 4.5 to 10 ml (=450-1000 mg tobramycin) in accordance with the prescribed usage by the suppliers. The nebulizers were connected with a special designed adapter to a laser diffraction analyser in order to measure particle size distribution of the aerosol. Inhalation was simulated with a static flow of 40 l/min. The particle size distribution (expressed as X(10), X(50), and X(90)) was determined after 10 s, 1.5, 3, 4.5, 6, 9 and 12 min of nebulization. Furthermore, the tobramycin solutions were assayed for tobramycin content before and after nebulization. For all nebulizers, the mean particle size distribution, depicted as X(50), was within the range of 1-5 mm. There were no relevant differences between the nebulizers in concentration or particle size distribution during nebulization. The output of the nebulizers is a result of both nebulization and evaporation. The output, expressed as volume of tobramycin solution, ranged from 0.06 to 0.50 ml/min. The output of tobramycin ranged from 1.2 to 39.5 mg/min. For clinical practice 300-600 mg have to be nebulized within 20-30 min. It was concluded that only three jet nebulizers [Porta-Neb Sidestream (PNS), Porta-Neb Ventstream (PNV) and Pariboy Pari LC+ (PLC)] have a reasonable output and an acceptable particle size distribution for the administration of a 10% tobramycin solution in the therapeutic dosage range. PMID- 10536250 TI - Inhalation of tobramycin in cystic fibrosis. Part 2: optimization of the tobramycin solution for a jet and an ultrasonic nebulizer. AB - The inhalation of tobramycin is part of current cystic fibrosis (CF) therapy. Local therapy with inhaled antibiotics has demonstrated improvements in pulmonary function. Current inhalation therapy is limited by the available drug formulations in combination with the nebulization time. The aim of this study is to develop a highly concentrated tobramycin solution for inhalation. Several tobramycin solutions, ranging from 5 to 30% (m/v), were compared after aerosolation with a jet and with an ultrasonic nebulizer. Laser diffraction and cascade impactor analysis were used for characterization of the aerosolized solutions. The output rate was determined in volume and mass output per minute. From the output rate measurements, it was concluded that a 20% tobramycin solution is the optimal and maximal concentration to be aerosolized. The jet nebulizer was most suitable. Using the jet nebulizer and the 20% solution, it is possible to administer a dosage of 1000 mg tobramycin by inhalation within 30 min. PMID- 10536251 TI - New injectable melphalan formulations utilizing (SBE)(7m)-beta-CD or HP-beta-CD. AB - The objective of this work was to evaluate the potential of using (SBE)(7m)-beta CD and HP-beta-CD as enabling excipients to improve on the current melphalan injectable formulation. Melphalan is an anti-neoplastic agent formulated for parenteral use as a sterile, non-pyrogenic, freeze-dried powder. It is marketed by Glaxo-Wellcome as ALKERAN((R)) for Injection (Alkeran). A major concern with melphalan therapy, other than its intrinsic cytotoxicity and biocompatibility, arises from its marginal aqueous solubility and chemical stability; thus, co solvents are used in the current two-vial formulation. Because of the two-vial system, the product is also inconvenient to use. Two approaches to improve melphalan's formulation utilizing cyclodextrins, including the use of aqueous (SBE)(7m)-beta-CD or HP-beta-CD solutions as the reconstitution diluents, and/or the use of (SBE)(7m)-beta-CD as a freeze-drying excipient in a melphalan formulation, are presented. Results showed that, when the cyclodextrins were used as diluents, the use of organic co-solvents can be eliminated and the shelf-life of the reconstituted melphalan greatly enhanced. When the freeze-dried melphalan/(SBE)(7m)-beta-CD formulation was prepared, the formulation was found to be stable; and a simplified one-vial delivery system was achieved. In conclusion, the parenterally safe beta-cyclodextrins derivatives can provide promising alternatives and improved formulations for melphalan injectable and perhaps similar problematic drugs. PMID- 10536252 TI - Comparative study of antibacterial activity of cefazolin sodium and of its crystalline modifications. AB - Antibacterial activity of four crystalline modifications (ground, solvated and two desolvated) of cefazolin sodium (CZS) have been studied. Their antibacterial properties have been determined by means of minimal inhibitory concentrations (MIC(50) and MIC(90)) and of disk diffusion inhibition zone values. The above mentioned crystalline modifications (Mod.) of CZS-I, II, III and IV, respectively, have been shown to have differences in their antibacterial properties in comparison with the trade CZS as well as between themselves. Mod. I has higher or the same activity as CZS. The obtained results we have explained with the formation of different intermolecular H-bonds in the modifications could be due to the rotation about C(alpha)- C' and C-N bonds in the amide groups in their molecules. PMID- 10536253 TI - Detection of the principal synthetic route indicative impurity in lamotrigine. AB - An analytical method has been developed for the detection of trace amounts of the principal synthetic route indicative impurity in lamotrigine (3,5-diamino-6-(2,3 dichlorophenyl)-1,2,4-triazine). A sample extract was preconcentrated by normal phase high-performance liquid chromatography (HPLC) and analysed by subsequent on line reversed-phase HPLC-thermospray mass spectrometry (TSP-MS). During the sample extraction and concentration step, carried out by semipreparative normal phase chromatography, the preliminary separation of the impurity from the lamotrigine takes place. The organic solvent (dichloroethane-methanol, 90:10, v/v) is evaporated from the collected fraction and the material is redissolved in a smaller volume of the reversed-phase mobile phase. The collected fraction is then subjected to reversed-phase HPLC-TSP-MS. The influence of an ultrasonic extraction step has been examined. When the method was applied to lamotrigine tablets, a shake flask partitioning step using 1 mg/ml EDTA in water dichloroethane was used instead of the ultrasonic extraction. Detection limit and recovery measurements showed that the route indicative impurity formed during the synthesis could be detected in the 50-100 ppb (w/w) range. PMID- 10536254 TI - Quantitative HPLC analysis of sunscreens and caffeine during in vitro percutaneous penetration studies. AB - This report describes rapid analytical HPLC for the quantification of five UV filters (octyl methoxycinnamate, benzophenone-3, benzophenone-4, octyl triazone and octocrylene) and of caffeine in various skin layers (stratum corneum, dermis, epidermis and receptor fluid) and in cosmetic preparations. The predominant purpose of the study was to establish standard operating procedures for rapid analysis of the compounds in various skin samples. Particular attention was paid to the preparation of biological samples whose natural constitution could interfere with the quantitative analysis. Our methods used the isocratic chromatographic mode in an RP-HPLC with UV detection and did not involve centrifugation or evaporation. Our results were validated in terms of specificity, linearity, precision, accuracy and limits of detection and quantification. The first results, obtained after in vitro experiments, are presented in this report. PMID- 10536255 TI - Hyposmotically induced amino acid release from the rat cerebral cortex: role of phospholipases and protein kinases. AB - In an evaluation of the contribution of swelling-induced amino acid release, through the regulatory volume decrease (RVD) process, to cerebral ischemic injury, studies of the role of phospholipases and protein kinases in the response to hyposmotic stress were undertaken using an in vivo rat cortical cup model. Hyposmotic stress induced significant releases of aspartate, glutamate, glycine, phosphoethanolamine, taurine and GABA from the rat cerebral cortex. Taurine release was most affected, exhibiting a greater than 9-fold increase during the hyposmotic stimulus. The phospholipase A2 (PLA2) inhibitors 4-bromophenacyl bromide (1 microM) and 7,7-dimethyleicosadienoic acid (5 microM) had no significant effects on hyposmotically induced amino acid release. AACOCF3 (50 microM), an inhibitor of cytosolic PLA2 decreased taurine release to 84% of DMSO controls. The release of the other amino acids was not affected. The phospholipase C inhibitor U73122 (5 microM) had no significant effects on amino acid release. The protein kinase C (PKC) inhibitor chelerythrine (5 microM) significantly reduced hyposmotically induced taurine release to 72% of saline controls but had no significant effects on the other amino acids. Stimulation of PKC with phorbol 12-myristate, 13-acetate (10 microM) did not significantly change taurine, glutamate, glycine or phosphethanolamine release. The releases of aspartate and GABA were enhanced 2 to 3 fold. Phorbol 12,13-didecanoate (10 microM), another potent stimulator of PKC, significantly increased taurine release to 122% of DMSO controls. The releases of aspartate, glutamate and glycine were enhanced 2.5 to 3.5 fold. Similarly, stimulation of protein kinase A with forskolin (100 microM) significantly increased taurine, aspartate, and glycine release 1.5- to 2-fold compared to DMSO controls. In summary, phospholipases may play a minor role in volume regulation. These studies also support the hypothesis that protein kinases play a modulatory role in the RVD response. The results show that although RVD may play a role, additional mechanisms, including phospholipase activation, must be involved in the ischemia evoked release of excitotoxic amino acids. PMID- 10536256 TI - N-methyl-D-aspartate receptor binding is altered and seizure potential reduced in pregnant rats. AB - The objective of this study was to determine if a change in brain tissue excitatory amino acid receptor binding occurs during pregnancy using in vitro quantitative autoradiography and to examine seizure potential during pregnancy via central injection of N-methyl-D-aspartate (NMDA). For the receptor autoradiography studies, eight pregnant rats (day 21) and eight non-pregnant rats were euthanized with carbon dioxide, perfused, their brains dissected and frozen. Cryostat sections were taken and labeled in vitro by one of the following ligands: [3H]-CGP 39653, [3H]-glycine, [3H]-MK-801, [3H]-2-amino-3-hydroxy-5 methyl-4-isoxazole propionate (AMPA) or [3H]-kainate. Optical density measurements of binding in 11 brain regions were performed using image analysis. To test seizure susceptibility, 74 rats were surgically implanted with an electrode into the hippocampus and a cannula into the lateral ventricle. Rats were mated; others served as non-pregnant controls. On gestational day 20, rats were randomized to receive no drug or an injection of NMDA (34, 68 or 136 nmol) through their indwelling cannulae. Seizures were assessed for 20 min. During pregnancy, the density of the NMDA competitive antagonist site measured by [3H] CGP 39653 was decreased in the hippocampus, thalamus and hypothalamus (P<0.01), while the glycine modulation site was decreased in the cortex, hippocampus, thalamus, caudate and cerebellum (P<0.01). Kainate binding was significantly decreased in the hippocampus (P<0. 05). Total seizure duration and total number of seizures were significantly reduced in pregnant vs. non-pregnant rats (P<0.05). Pregnancy is associated with a significant alteration of NMDA and non NMDA receptor binding in rats. These findings suggest that pregnancy affords some protection against seizures induced by an activation of NMDA receptors in the brain. PMID- 10536257 TI - Treatment with estrogen and progesterone affects relative levels of brain-derived neurotrophic factor mRNA and protein in different regions of the adult rat brain. AB - Quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and enzyme linked immunosorbent assay (ELISA) were used to examine the effects of acute estrogen and progesterone replacement on relative levels of brain-derived neurotrophic factor (BDNF) mRNA and protein in different regions of the adult rat brain. Adult ovariectomized animals were killed 53 h after receiving estrogen (E53), 53 h after receiving estrogen and 5 h after receiving progesterone (E53P), or 72 h after receiving estrogen and 24 h after receiving progesterone (E72P). Ovariectomized controls were killed 53 and 72 h after receiving vehicle. Tissues from the hippocampus, pyriform cortex, olfactory bulbs, septum, and nucleus basalis/ventral pallidum were dissected. Tissues from the right hemisphere were processed for quantitative RT-PCR analysis of BDNF mRNA, and tissues from the left hemisphere were processed for the detection and quantification of BDNF protein by ELISA. The results demonstrate significant increases in BDNF mRNA in the pyriform cortex of E53- and E53P-treated animals, as well as an increase in BDNF protein in the pyriform cortex of E72P-treated animals, relative to controls. Significant increases in BDNF mRNA were likewise detected in the hippocampus of E53- and E72P-treated animals, but were accompanied by a significant decrease in BDNF protein in the hippocampus of E53P- and E72P-treated animals relative to controls. No significant changes in BDNF mRNA or protein were detected in the olfactory bulbs, frontal cortex, or nucleus basalis/ventral pallidum following hormone treatment; however, an increase in BDNF protein was detected in the septum of E53-treated animals. This may indicate an increase in the retrograde transport of BDNF from the hippocampus to the septum, which could help account for the decrease in BDNF protein detected in the hippocampus following hormone treatment. These findings demonstrate that hormone replacement significantly affects relative levels of BDNF mRNA and protein within specific regions of the brain. These effects may, in turn, contribute to the effects of estrogen replacement on hippocampal connectivity and cognitive processes that have recently been reported. PMID- 10536258 TI - Mossy fiber sprouting induced by repeated electroconvulsive shock seizures. AB - The elicitation of repeated focal seizures (kindling) induces mossy fiber sprouting in the hippocampus of the rat. The present study investigated whether repeated generalized seizures also induce mossy fiber sprouting. Human psychiatric patients receive repeated generalized seizures during electroconvulsive therapy (ECT). Male Long-Evans rats received a course of eight electroconvulsive shock (ECS) seizures administered on a 48-h schedule over a course of 2 1/2 weeks. Control subjects received matched handling, but no stimulation. Fourteen days after the last ECS trial, all subjects were sacrificed and their brains subjected to Timm staining. Cell counts and area measures were also taken in the hilus. Significant sprouting, but not significant cell loss, was seen in the fascia dentata of the subjects that had received ECS. PMID- 10536259 TI - Estrogen decreases hypothalamic angiotensin II AT1 receptor binding and mRNA in the female rat. AB - Estrogen has been shown to modulate angiotensin II (AngII)-regulated behaviors, such as thirst, and may do so by influencing the central renin-angiotensin system (RAS). While numerous studies have attempted to correlate changes in AngII receptors or other components of the RAS with estrogen treatment, the low abundance of these genes has made comparisons difficult. Generally, such experiments have relied on traditional approaches to analyze gene expression that often restrict the experimenter to studying only a few mRNA species, whereas a behavior as complex as thirst may be influenced by changes in multiple genes. The present experiments utilized quantitative receptor autoradiography and mRNA expression profiling to identify and compare AngII receptors and their mRNA levels as well as other components of the RAS in female rat pituitary and hypothalamic-thalamic-septal (HTS) tissue samples. This relatively new approach to the study of gene expression permits the simultaneous comparison of multiple genes from a single tissue sample. These studies revealed that ovariectomized (OVX) female rats treated with estradiol benzoate (EB) had a 30%-40% reduction in the levels of AT(1) receptor mRNA in pituitary and HTS samples as compared to OVX, control animals. In the pituitary, the mRNA levels for angiotensinogen (AGT) were increased by 45% following estrogen administration. In addition, a reduction in [125I]-AngII binding to AT(1) receptors in the pituitary and the subfornical organ was measured following estrogen treatment. These results suggest that estrogen may modulate the pituitary and central RAS through a coordinate regulation of the angiotensin receptors and the levels of newly synthesized AngII. PMID- 10536260 TI - Age-related changes in ultrastructural features of cathepsin B- and D-containing neurons in rat cerebral cortex. AB - The present study examines age-related changes in the subcellular localization of cathepsin B (cath B) and cathepsin D (cath D), as well as morphological features of the cathepsin-immunoreactive (ir) neurons in rat cerebral cortex. Sprague Dawley rats were studied at 3 and 26 months. By immunoelectron microscopy cath B- or cath D-immunoreactivities were found in many, but not all, pyramidal neurons. In young rat cerebral cortical neurons, cath B was observed not only in lysosomal systems such as multivesicular bodies, dense bodies, and lipofuscin granules, but also in extralysosomal sites. By contrast, cath D was confined mainly to lysosomal systems in young rats. In aged rats, cath B showed a similar pattern in its subcellular localization compared to the young control, but some of the dense bodies containing cath B was closely apposed to the outer nuclear envelope. These cells exhibited a relatively normal appearance. Regardless of subcellular localization, approximately 10% of cath B-ir neurons displayed ultrastructural disturbances presumed to indicate an early stage of degeneration. The nucleus was indented, nuclear boundary was indistinct, nuclear pore structures appeared separately with high frequency, and the endoplasmic reticulum appeared to be affected. In addition to its presence in lysosomal structures, cath D immunoreactivity in aged cerebral cortex was noted prominently in the cytosol as diffuse granules. About 37% of cath D-ir cells showed this age-related change. Among the neurons with the diffusely scattered form of cath D, approximately 70% of cells exhibited the degenerating features. These cells were characterized by large amounts of diffuse cath D, reduced cellular size, loss of the nuclear boundary, scattered nuclear pore structures, an often fragmentation of the nucleus, disturbances of endoplasmic reticular system, and in advanced stages, condensed nucleus and poor preservation of almost cytoplasmic organelles. Though some of these features were also found in cath B-ir neurons, findings of overt degeneration, such as fragmented and condensed nuclei and impaired almost cytoplasmic organelles, were generally not observed in cath B-ir neurons. In addition, lipofuscin aggregates containing cath D were observed frequently in the extracellular space close to sites of ruptured plasma membrane, whereas in the sections stained with anti-cath B antibodies, large-sized lipofuscin aggregates showed only very weak or no cath B-immunoreactivity at all. Taken together, the present results suggest that cath D and cath B may be regulated differently and play their specific roles in the aging of the brain, especially, the change in location of cath D from the lysosomal system to the cytosol in the aged brain may play an important role in age-related cell death. PMID- 10536261 TI - Three-dimensional measurement of cerebral microvascular plasma perfusion, glial fibrillary acidic protein and microtubule associated protein-2 immunoreactivity after embolic stroke in rats: a double fluorescent labeled laser-scanning confocal microscopic study. AB - Early astroglial response to post-ischemic microvascular hypoperfusion may contribute to progressive cerebral microcirculatory impairment and ischemic neuronal injury. Using laser-scanning confocal microscopy and three fluorescent probes, we measured in three-dimensions cerebral microvascular plasma perfusion, astrocytic reactivity, and neuronal injury assessed by fluorescein isothiocyanate (FITC)-dextran, GFAP immunoreactivity, and microtubule associated protein-2 (MAP2) immunoreactivity, respectively, in rats subjected to 2 h of middle cerebral artery occlusion. Three-dimensional quantitative analysis revealed that 2 h of embolic ischemia resulted in a significant (P<0.05) reduction of cerebral microvascular plasma perfusion in the ipsilateral cortex and subcortex. Tissue within the ipsilateral cortex and subcortex with low plasma perfusion exhibited a significant (P<0.05) increase in GFAP immunoreactivity compared with the homologous contralateral tissue. Three-dimensional re-constructed images showed that prominent GFAP immunoreactive astrocytes surrounded large vessels with decreased plasma perfusion in downstream capillaries in the ipsilateral MCA territory when compared to the vessels in the contralateral homologous tissue. Triple fluorescence probe-stained sections showed that tissue with decreased plasma perfusion and with increased GFAP immunoreactivity was accompanied by a reduction of MAP2 immunoreactivity. The present study demonstrates that an impairment of microvascular perfusion induces an early increase in GFAP immunoreactivity, and reactive astrocytes may contribute to a further reduction of cerebral microvascular plasma perfusion. The three-dimensional quantitative imaging analysis used in the present study provides a means to investigate parenchymal cellular responses to changes of cerebral microvascular plasma perfusion after MCA occlusion. PMID- 10536263 TI - Does sex or photoperiodic condition influence ZENK induction in response to song in European starlings? AB - Variables such as the photoperiod a bird experiences, or its sex, affect behavioral responses to song. The present experiment investigated whether song induced expression of the immediate-early gene ZENK is also influenced by sex or photoperiod. We examined the expression of the protein product of ZENK in wild caught male and female European starlings (Sturnus vulgaris) in different photoperiodic conditions. In the first experiment, adult reproductively active male and female starlings were presented with either conspecific male song or no song in experimental chambers. In the second experiment, conspecific male song was presented to reproductively active and reproductively inactive adult female starlings. Localization of the ZENK protein product revealed that song stimulation resulted in a significant increase in the number of ZENK immunoreactive (-ir) cells in the caudomedial neostriatum (NCM) and the caudomedial hyperstriatum ventrale (cmHV) compared with unstimulated birds. No differences in the number of ZENK-ir neurons were observed between males and females or between reproductively active and inactive females. Thus, the present data indicate that the number of cells expressing ZENK in NCM and cmHV following song playback does not vary with sex or photoperiod in starlings. PMID- 10536262 TI - Calbindin-D 28kD immunofluorescence in ventral mesencephalic neurons labeled following injections of Fluoro-Gold in nucleus accumbens subterritories: inverse relationship relative to known neurotoxin vulnerabilities. AB - The shell and core of the nucleus accumbens exhibit different vulnerabilities to neurotoxins. Calcium binding proteins are reported to offer some neuroprotection against excitotoxicity by suppressing or buffering intracellular calcium. Differences in the distributions of the calbindin-D 28kD (CB) and calretinin (CR) might be related to the different vulnerabilities to neurotoxins of dopaminergic neurons in the ventral mesencephalon that project to the core and medial shell of the nucleus accumbens. To address this possibility, Fluoro-Gold (FG) was injected into accumbens subterritories and numbers of retrogradely labeled neurons in the ventral tegmental area containing CB and CR immunoreactivities (ir) were expressed as a percentage of total numbers of labeled neurons. The perikaryal diameters and lengths of the immunoreactive dendrites of FG labeled neurons were also measured. About 70% and 35% of retrogradely labeled cells observed following core and medial shell injections, respectively, exhibited CB immunoreactivity. Differences were not observed in the percentages of FG labeled cells exhibiting CR immunoreactivity following medial shell (13%) and core (15%) injections. The mean perikaryal diameters and median summed lengths of dendrites of retrogradely labeled neurons containing CB were smaller than in labeled neurons lacking CB following injections in both core and medial shell of the nucleus accumbens. The data indicate that the different 6-hydroxydopamine (6-OHDA) vulnerabilities of ventral mesencephalic dopaminergic neurons are not obviously related to the presence of CB and CR. PMID- 10536264 TI - Neurogenic calcitonin gene-related peptide: a neurotrophic factor in the maintenance of acetylcholinesterase molecular forms in adult skeletal muscles. AB - This work addresses the role of calcitonin gene-related peptide (CGRP) in the physiological maintenance of acetylcholinesterase (AChE) molecular forms in motor endplate regions of adult Sprague-Dawley rat fast-twitch anterior gracilis muscles. Results show that: (a) CGRP is present in obturator nerve motor neurons which supply the gracilis muscle, as well as in the corresponding motor endplate regions where high levels of both AChE activity and acetylcholine receptors (AChRs) are detected; (b) endplate-associated CGRP declines with muscle denervation several hours before any changes in AChE forms are detected; (c) a single subcutaneous injection of CGRP reversibly reduces the activities of all AChE forms in endplate regions of normally innervated and otherwise untreated gracilis muscles; and (d) similar treatment with hCGRP(8-37), a potent and selective CGRP antagonist, produces the opposite effects, i.e., it reversibly elevates the activities of all AChE forms. These and other findings indicate that CGRP and hCGRP(8-37) influence the mechanism(s) by which AChE forms are maintained in intact adult gracilis muscles. Indeed, the findings lend strong support to the hypothesis that nerve-derived CGRP plays a key role in the trophic regulation of AChE forms at the neuromuscular junction. PMID- 10536265 TI - Involvement of peripheral NMDA and non-NMDA receptors in development of persistent firing of spinal wide-dynamic-range neurons induced by subcutaneous bee venom injection in the cat. AB - To study the roles of peripheral excitatory amino acids receptor subtypes N methyl-D-aspartate (NMDA) and non-NMDA receptors in persistent nociception, extracellular single unit recording technique was used to assess the effects of a single dose NMDA and non-NMDA receptor antagonists, AP(5) (5 aminophosphonovaleric acid) and CNQX (6-cyano-7-nitroquinoxaline-2,3-dione) or DNQX (6,7-dinitroquinoxaline-2,3-dione), on s.c. bee venom-induced increase in firing of wide-dynamic-range (WDR) neurons in the spinal dorsal horn of the urethane-chloralose anesthetized cats. Subcutaneous bee venom injection into the cutaneous receptive field resulted in a single phase of increased firing of WDR neurons over the background activity for more than 1 h. Local pre-administration of AP(5) (200 microg/100 microl) or CNQX (8.3 microg/100 microl) into the bee venom injection site produced 94% (1.01+/-0.96 spikes/s, n=5) or 76% (2.97+/-0.58 spikes/s, n=4) suppression of the increased neuronal firing when compared with local saline (16.32+/-4.55 spikes/s, n=10) or dimethyl sulfoxide (DMSO) (12.37+/ 6.36 spikes/s, n=4) pre-treated group, respectively. Local post-administration of the same dose of AP(5) produced a similar result to the pre-treatment group with a 67% inhibition of the mean firing rate, however, the same treatment with CNQX and even a higher dose of DNQX (100 microg/100 microl) did not produce any inhibition of the neuronal firing induced by s.c. bee venom injection (DNQX vs. DMSO: 23.91+/-0. 25 vs. 22.14+/-0.04 spikes/s, P=0.0298, n=5). In the control experiments, local pre-administration of the same dose of AP(5) or CNQX into a region on the contralateral hindpaw symmetrical to the bee venom injection site produced no significant influence on the increased firing of the WDR neurons [contralateral AP(5) vs. saline: 14.17+/-6.27 spikes/s (n=5) vs. 16.32+/-4.55 spikes/s (n=10), P0.05; contralateral CNQX vs. DMSO: 12.85+/-6.38 spikes/s (n=4) vs. 12. 37+/-6.36 spikes/s (n=4), P0.05], implicating that the suppressive action of local AP(5) or CNQX was not the result of systemic effects. The present results suggest that activation of the peripheral NMDA receptors is involved in both induction and maintenance, while activation of non-NMDA receptors is only involved in induction, but not in the maintenance of persistent firing of the dorsal horn WDR neurons induced by s.c. bee venom injection. PMID- 10536266 TI - Regulation of NMDA-stimulated [14C]GABA and [3H]acetylcholine release by striatal glutamate and dopamine receptors. AB - Striatal function is heavily influenced by glutamatergic and dopaminergic afferent input. To ultimately better understand how the N-methyl-D-aspartate (NMDA) antagonist, phencyclidine (PCP), alters striatal function, we sought to determine how NMDA receptor function is influenced by activation of other glutamatergic receptors and by dopaminergic receptors. To this end, we used NMDA stimulated efflux of [14C]GABA and [3H]acetylcholine (ACh) from striatal slices to assess the influence of these receptors on NMDA function. NMDA-stimulated [14C]GABA release was more sensitive to NMDA and glycine antagonists than was [3H]ACh release, suggesting that different NMDA receptors regulate the release of these neurotransmitters. Furthermore, NMDA-stimulated [3H]ACh release was inhibited by a D2 receptor mechanism whereas NMDA-stimulated [14C]GABA release was enhanced by D1 receptor activation. NMDA and (+/-)-alpha-amino-3-hydroxy-5 methylisoxazole-4-propionic acid hydrobromide (AMPA) interact additively to evoke [3H]ACh release, and synergistically to evoke [14C]GABA release. An additive effect of NMDA and kainate (KA) was found on [14C]GABA release, but NMDA and KA acted in a less than additive manner in evoking [3H]ACh release. KA-stimulated [3H]ACh release was largely blocked by NMDA antagonists, suggesting mediation through activation of NMDA receptors, probably secondary to KA-induced glutamate release. A selective group II metabotropic receptor agonist inhibited NMDA stimulated [14C]GABA and [3H]ACh release. On the other hand, NMDA-stimulated [14C]GABA release was potentiated by activation of group I metabotropic receptors. Thus, in addition to the differential modulation by D1- and D2-like receptors, the release of striatal neurotransmitters by NMDA receptor activation depends on the extent to which the other glutamate receptors, both ionotropic and metabotropic, are activated. PMID- 10536267 TI - An in vitro electrophysiological and Co2+-uptake study on the effect of infraorbital nerve transection on the cortical and thalamic neuronal activity. AB - Changes of neuronal membrane characteristics in somatosensory barrel cortex and barreloid thalamus were investigated in rats following unilateral transection of the infraorbital nerve. Kainate induced Co2+-uptake method and image analysis were used to assess the Ca2+ permeability of non-NMDA (N-methyl-D-aspartate) glutamate receptors. Changes in some biophysical parameters of the affected cortical neurons were also investigated by intracellular recording in slice experiments. The altered neuronal activity was measured on days 1, 5 and 14 after surgery. Kainate induced Co2+ uptake increased markedly reflecting enhanced Ca2+ permeability of alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionate/kainate (AMPA/KAIN)-type receptors. Changes were more pronounced in the cortex than in the thalamus and peaked on the first day following nerve transection. After that, parameters gradually returned to the normal level. However, a small enhancement was still detectable in the cortex at the end of the 2-week-long observation period. In parallel with the increased Co2+-uptake, moderate membrane potential changes, stronger spiking activity and enhanced excitability were characteristic for cortical neurons. The observed alterations in neuronal characteristics underlie the reorganization and regeneration processes following injuries or surgeries. We can conclude that immediate change of the receptive field in the barrel cortex following unilateral nerve transection is based on changes in biophysical parameters of the neurons. Altered peripheral activation evokes changes in the neuronal activity, thus providing opportunity for a quick synaptic rearrangement. AMPA/KAIN-type glutamate receptors have a decisive role in the regulation of these processes. This kind of synaptic plasticity is more significant in the cortex than in the thalamus. PMID- 10536269 TI - Effects of fasting and insulin-induced hypoglycemia on brain cell membrane function and energy metabolism during hypoxia-ischemia in newborn piglets. AB - This study was done to determine the effects of 12 h fasting-induced mild hypoglycemia (blood glucose 60 mg/dl) and insulin-induced moderate hypoglycemia (blood glucose 35 mg/dl) on brain cell membrane function and energy metabolism during hypoxia-ischemia in newborn piglets. Sixty-three ventilated piglets were divided into six groups; normoglycemic control (NC, n=8), fasting-induced mildly hypoglycemic control (FC, n=10), insulin-induced moderately hypoglycemic control (IC, n=10), normoglycemic/hypoxic-ischemic (NH, n=11), fasting-induced mildly hypoglycemic/hypoxic-ischemic (FH, n=12) and insulin-induced moderately hypoglycemic/hypoxic-ischemic (IH, n=12) group. Cerebral hypoxia-ischemia was induced by occlusion of bilateral common carotid arteries and simultaneous breathing with 8% oxygen for 30 min. The brain lactate level was elevated in NH group and this change was attenuated in FH and IH groups. The extent of cerebral lactic acidosis during hypoxic-ischemic insult showed significant positive correlation with blood glucose level (r=0.55, p<0.001). Cerebral Na+, K+-ATPase activity and concentrations of high-energy phosphate compounds were reduced in NH group and these changes were not ameliorated in FH or IH group. Cortical levels of conjugated dienes, measured as an index of lipid peroxidation of brain cell membrane, were significantly elevated in NH, FH and IH groups compared with NC, FC and IC groups and these increases were more profound in FH and IH with respect to NH. Blood glucose concentration showed significant inverse correlation with levels of conjugated dienes (r=-0.35, p<0.05). These findings suggest that, unlike in adults, mild or moderate hypoglycemia, regardless of methods of induction such as fasting or insulin-induced, during cerebral hypoxia-ischemia is not beneficial and may even be harmful in neonates. PMID- 10536268 TI - Screening in a cell-based assay for inhibitors of microglial nitric oxide production reveals calmodulin-regulated protein kinases as potential drug discovery targets. AB - A high-throughput screening (HTS) assay for inhibitors of nitric oxide (NO) production by activated microglia was developed and used to compare the relative activities of various anti-inflammatory compounds and cell-permeable protein kinase inhibitors. BV-2 cells, an immortalized line that retains phenotypic features of microglia and produces NO in response to lipopolysaccharide (LPS), were used in the activation paradigm for the HTS assay. A characteristic feature of the compounds that were the most potent dose-dependent inhibitors of NO production is their ability to modulate serine/threonine protein kinases. The anti-inflammatory compound K252a, an inhibitor of calmodulin (CaM)-regulated protein kinases, had one of the highest potencies in the assay. Other classes of kinase inhibitors, including the protein kinase A inhibitor H-89, the mitogen activated protein kinase inhibitors PD98059 and SB203580, and the tyrosine kinase inhibitor genistein, were less potent and efficacious than K252a or the general serine/threonine/tyrosine kinase inhibitor staurosporine. K252a suppresses production of the inducible nitric-oxide synthase (iNOS). The inhibitory effect of K252a is not due to cell toxicity and does not correlate with inhibition of NFkappaB nuclear translocation. The mechanism of action appears to involve inhibition of phosphorylation of the transcription factor CREB, a protein whose activity is modulated by phosphorylation by CaM-dependent protein kinases. These data suggest that signal transduction pathways mediated by CaM-dependent protein kinases warrant future study as potential drug discovery targets. PMID- 10536270 TI - Effect of small changes in temperature on CA1 pyramidal cells from rat hippocampal slices during hypoxia: implications about the mechanism of hypothermic protection against neuronal damage. AB - Small reductions in temperature have been shown to improve neurologic recovery after ischemia. We have examined the effect of temperature on biochemical and physiological changes during hypoxia using rat hippocampal slices as a model system. The postsynaptic population spike recorded from the CA1 pyramidal cell region of slices subjected to 7 min of hypoxia with hypothermia (34 degrees C) recovered to 73% of its prehypoxic level; slices subjected to the same period of hypoxia at 37 degrees C did not recover. After 7 min of hypoxia ATP fell to 48% of its prehypoxic concentration at 34 degrees C and 30% at 37 degrees C. Potassium fell to 86% during 7 min of hypoxia with hypothermia, this compares to a fall to 58% at 37 degrees C. The increase in sodium after 7 min of hypoxia was also attenuated by hypothermia (133% vs. 163% of its prehypoxic concentration). When the hypoxic period was shortened to 3 min (37 degrees C) the population spike recovered to 94%. If the temperature was increased to 40 degrees C there was only 7% recovery of the population spike after 3 min of hypoxia. With hyperthermia (40 degrees C), ATP fell to 33% after 3 min of hypoxia, this compares to 81% at normothermia. Potassium fell to 76% after 3 min of hypoxia with hyperthermia, this compares to 91% at 37 degrees C. Sodium concentrations increased with hyperthermia before hypoxia, at 3 min of hypoxia there was no significant difference between the hyperthermic and normothermic tissue; there was a large increase in sodium with hyperthermia after 5 min of hypoxia (209% vs. 146%). We conclude that the improved recovery after hypothermic hypoxia is at least in part due to the attenuated changes in ATP, potassium and sodium during hypoxia and that the worsened recovery with hyperthermia is due to an exacerbation of the change in ATP, potassium and sodium concentrations during hypoxia. PMID- 10536271 TI - [3H]Spermine binding to synaptosomal membranes from the chick retina. AB - The binding of [3H]spermine to synaptosomal membranes from chick retina was examined. Saturable specific binding of [3H]spermine to synaptosomal membranes from plexiform layers of retina (P1 and P2) has been characterized, and found to concentrate in the inner plexiform layer compared to the outer plexiform layer (Bmax=9.3 and 37 pmol/mg protein for P1 and P2, respectively). Kinetics of specific [3H]spermine binding yield a sigmoidal saturation curve, indicating positive cooperativity (nH: 2.4 and 3.2 for P1 and P2, respectively) with high affinity: Kapp=61 and 67 nM for P1 and P2. The time required to attain equilibrium at room temperature was less than 5 min in both fractions. Dose response curves for spermine, spermidine, and diethylene-triamine (DET) show different potencies for inhibiting [3HDET. Our results support a role for polyamines (PA) as neurotransmitters or neuromodulators in the vertebrate retina. PMID- 10536272 TI - Vagotomy does not affect thermal responsiveness to intrabrain prostaglandin E2 and cholecystokinin octapeptide. AB - Subdiaphragmatic vagotomy has been repeatedly shown to attenuate the febrile response to peripherally injected pyrogens. In the present study, we investigated whether vagotomy-induced attenuation of febrile responsiveness reflects a decreased sensitivity of the brain to central fever mediators, prostaglandin E2 (PGE2) and cholecystokinin octapeptide (CCK-8). Male rats were subjected to subdiaphragmatic vagotomy (or sham surgery) on day 0 and had a cannula implanted into the lateral cerebral ventricle on day 24. On day 30-36, the thermal responsiveness of the rats to PGE2 or CCK-8 was tested. Each animal was injected in the ventricle with either PGE2 (0, 10, 100, or 500 ng) in pyrogen-free saline with 0.5% ethanol (5 microl) or CCK-8 (0 or 1.6 microg) in artificial cerebro spinal fluid (5 microl). While the 0-dose of either PGE2 or CCK-8 (vehicle alone) induced no thermal response, all the higher doses of either agent caused a body temperature rise preceded by tail skin vasoconstriction. The vagotomized rats did not respond differently than their sham-operated counterparts to any dose of either drug. It is concluded that subdiaphragmatic vagotomy does not change the rat's thermal responsiveness to intrabrain PGE(2) and CCK-8. PMID- 10536273 TI - Changes in extracellular glutamate and pressor response during muscle contraction following AMPA-receptor blockade in the RVLM and CVLM. AB - We examined whether modulation of cardiovascular responses by administering 6 cyano-7-nitroquinoxaline-2,3-dione (CNQX, an AMPA-receptor antagonist) into the rostral (RVLM) or caudal (CVLM) ventrolateral medulla are mediated via changes in extracellular levels of glutamate. Microdialysis probes were inserted bilaterally into the RVLM or the CVLM. For the RVLM experiments (n=8), muscle contraction for 2 min increased mean arterial pressure (MAP) and heart rate (HR) by 18+/-3 mmHg and 24+/-5 bpm, respectively. Extracellular glutamate concentrations increased from 1.5+/-0.3 to 4.3+/-0.9 ng/5 microl during the contraction. Microdialysis of CNQX (1.0 microM) for 30 min into the RVLM attenuated the increases in MAP, HR, and glutamate concentration in response to a muscle contraction (8+/-2 mmHg, 11+/ 3 bpm, and 2.2+/-0.7 ng/5 microl, respectively). Developed tensions did not change during contractions before and after CNQX. Microdialysis of CNQX into the CVLM (n=8) potentiated the contraction-evoked responses in MAP (19+/-3 vs. 34+/-3 mmHg) and HR (25+/-4 vs. 49+/-5 bpm) without a change in developed tension. Following CNQX perfusion into the CVLM, the levels of extracellular glutamate in the CVLM were also augmented during the contraction. Results suggests that AMPA receptors within the RVLM and CVLM differentially modulate cardiovascular responses during static muscle contraction via increasing and decreasing, respectively, extracellular glutamate concentrations. PMID- 10536274 TI - Increased expression of growth-associated protein (GAP-43) in lower urinary tract pathways following cyclophosphamide (CYP)-induced cystitis. AB - Alterations in the expression of growth-associated protein 43 (GAP-43) were examined in lower urinary tract micturition reflex pathways in a chronic model of cyclophosphamide (CYP)-induced cystitis. In control animals, expression of GAP-43 was present in specific regions of the gray matter in the rostral lumbar and caudal lumbosacral spinal cord, including: (1) the dorsal commissure; (2) the dorsal horn and (3) the regions of the intermediolateral cell column (L1-L2) and the sacral parasympathetic nucleus (L6-S1) and (4) in the lateral collateral pathway of Lissauer in L6-S1 spinal segments. Densitometry analysis has demonstrated significant increases (p/=0.1 microm moving toward a cold surface or away from a hot surface at a given temperature gradient. The electrophoresis effect dominates the deposition of submicron particles. PMID- 10536288 TI - Hydrogen peroxide decomposition in model subsurface systems. AB - Rates of hydrogen peroxide decomposition, hydroxyl radical production, and oxygen evolution were investigated in silica sand-goethite slurries using unstabilized and stabilized hydrogen peroxide formulations. The goethite-catalyzed decomposition of unstabilized hydrogen peroxide formulations resulted in more rapid hydrogen peroxide loss and oxygen evolution relative to systems containing a highly stabilized hydrogen peroxide formulation. Systems at neutral pH and those containing higher goethite concentrations were characterized by higher rates of hydrogen peroxide decomposition and by more oxygen evolution. The stabilized hydrogen peroxide formulation showed greater hydroxyl radical production relative to the unstabilized formulations. Furthermore, hydroxyl radical production rates were greater at neutral pH than at the acidic pH regimes. The results suggest that when stabilized hydrogen peroxide is injected into the subsurface during in situ bioremediation, naturally occurring minerals such as goethite may initiate Fenton-like reactions. While these reactions may prove to be toxic to microorganisms, they have the potential to chemically oxidize contaminants in soils and groundwater. PMID- 10536289 TI - The formation of explosive compounds in bitumen/nitrate mixtures AB - Ignition of various bitumen/nitrate mixtures was examined using a new method of thermal analysis, developed in the present study, in order to determine the self ignition temperatures (SIT) of the sample materials. The effects on the SIT of transition elements, particularly silver, and the heating rates were examined. The same bitumen/nitrate mixtures were prepared by mixing bitumen, NaNO(3), Ni(NO(3))(2) and either AgNO(3) or AgI. The sample heating rates were in the range from 0.5 to 4.0 degrees C/min. In the present work, SIT was found to be independent on the heating rate when AgI is added, while SIT was found to depend on the heating rate when AgNO(3) was added. The activation energy E(a) for the ignition of bitumen/nitrate mixtures has been calculated to be 30 kJ/mol. The suspected initiator of the ignition is thought to be silver fulminates or related compounds. PMID- 10536290 TI - Removal and degradation of phenol in a saturated flow by in-situ electrokinetic remediation and Fenton-like process. AB - In this laboratory study, a sandy loam soil saturated with phenol solution was treated by in-situ electrokinetics-Fenton process incorporated with a permeable reactive wall of scrap iron powder (SIP). The soil was contaminated and saturated with aqueous phenol solution of 90-115 mg/kg in concentration. It was then placed in a soil cell. The soil cell was assembled with an anode reservoir and a cathode reservoir at its ends. A bed of SIP (1.05-32.69 g) was inserted in the soil cell at a distance of 5 cm from the anode reservoir compartment. For the test runs, 0.3% H(2)O(2) was used as the anode reservoir fluid, whereas de-ionized water was used as the cathode reservoir fluid. An electric gradient of 1 V/cm was applied to enhance the saturated flow in the soil cell for a period of 10 days. Experimental results have shown that the electroosmotic (EO) flow quantity decreased as the amount of SIP increased. This phenomenon was in good agreement with the results showing the value of EO permeability increased with a decreasing amount of SIP. Results also showed that throughout the test period the cumulative, consumed mass of H(2)O(2) in the anode reservoir increased as the amount of SIP decreased. On the other hand, the cumulative, increased mass of phenol in the cathode reservoir was found to increase with a decreasing amount of SIP. Meanwhile, the residual phenol concentration in the soil cell was found to decrease with a decreasing amount of SIP. When 1.05 g scrap iron powder was used, an overall removal and destruction efficiency of phenol of 99.7% was obtained. Therefore, it is evident that an in-situ combined technology of electrokinetic remediation and Fenton-like process is capable of simultaneously removing and degrading the phenol in a saturated flow. PMID- 10536292 TI - Pore structure and adsorption performance of the activated carbons prepared from plum kernels. AB - According to iodine number, amount of methylene blue adsorption, the BET specific surface area, and the yield, the conditions for preparing activated carbons as adsorbents from plum kernels were optimized. The activation temperature and time tested were in the ranges 750-900 degrees C and 1-4 h, respectively. Adsorption isotherms of two commercial dyes and phenol from water on such activated carbons were measured at 30 degrees C. It was shown that the optimal activation temperature and time depended on the molar mass of the solutes, and all equilibrium isotherms could be fitted by the Langmuir equation. The experimental results indicated that the prepared activated carbons were economically promising for adsorption removal of dyes and phenol, in contrast to other commercial adsorbents. PMID- 10536291 TI - Recycling of lead-contaminated EDTA wastewater. AB - Ethylene diaminetetraacetic acid (EDTA) is one of the chelating agents used in the soil washing process for the decontamination of lead-contaminated soil. Lead EDTA complexes in the wastewater from the soil washing process must be removed before the wastewater can be safely discharged. This study outlines a method to recycle Pb-EDTA wastewater by substituting the Pb complexed with EDTA with Fe(III) ions at low pH, followed by precipitation of Pb ions with phosphate or sulfate ions. Fe(III) ions complexed with EDTA were then precipitated at high pH using sodium hydroxide. The resulting solution (Fe-precipitated solution) was tested on three lead-contaminated soils. The Fe-precipitated EDTA solution was found to have similar extraction capabilities as fresh EDTA solution. Experimental results showed that the recycled EDTA solution may be recycled several times without losing its extractive power. Recycled EDTA wastewater with phosphate precipitation was found to be slightly more effective than recycled EDTA solution using sulfate precipitation. The recycling procedure may be applied to wastewater generated during soil washing of lead-contaminated soil, resulting in a reduction in wastewater generated and savings in the amount of EDTA used. PMID- 10536293 TI - Ozonation of p-chlorophenol in aqueous solution. AB - The ozonation of p-chlorophenol (CHP) in aqueous solution has been studied in the pH range 2.0-8.0, in the presence of tert-butyl alcohol, which prevents the activation of the radical mechanism of oxidation. Results indicate that the pH influences the system reactivity and that only a partial chlorine release is observed for lengthy ozonation too, after the complete substrate disappearance. For adopted experimental conditions the oxidation process develops under a quasi diffusional regime of absorption with reaction, a transition domain between kinetic and diffusional regimes in which ozone and dissolved substances react exclusively in the liquid film. A proper mathematical model has been developed and used to simulate the system behaviour PMID- 10536294 TI - Thermodynamics of ions precipitation in mixed-solvent mixtures AB - A framework derived from thermodynamic principles of solid-liquid equilibrium criteria was formulated to correlate and predict the precipitation of salts from aqueous solutions using organic solvents. The activity coefficient of a given salt in a mixed-solvent mixture was related to the activity coefficients of such a salt in each of the pure solvents (water and organic) using the excess Henry's constant approach (H(i)(E)). The Wohl's expansion was then employed to model the excess Gibbs free energy (g(E)) function. Two equations were provided; the two suffix equation, and the three-suffix equation. A previously acquired precipitation database was used to evaluate the correlative ability of the framework equations. The precipitation measurements were adequately correlated and predicted by the two interaction parameters equation; the two-suffix equation. However, the three-suffix equation, with three interaction parameters, was more accurate. The regressed interaction parameters can be used as predictive tools to estimate the precipitation fractions (P) for the tested systems for which no experimental data are available. Furthermore, such parameters can be employed to predict the solubilities of the tested salts in the organic solvent. PMID- 10536295 TI - Chemotherapy for enterohemorrhagic Escherichia coli O157:H infection in a mouse model. AB - The aim of this study is to evaluate the therapeutic effect of the antimicrobial agents, fosfomycin (FOM), minocycline (MINO), kanamycin (KM) and norfloxacin (NFLX) in the enterohemorrhagic Escherichia coli (EHEC) infected mouse model which we established previously (Infect. Immun. 62 (1994) 3447-3453). Each of the antimicrobial agents, 1/16 LD(50), was given to the mice per os (p.o. ) or intraperitoneally (i.p.) for 3 days after bacterial inoculation and then we observed their mortality rate for 2 weeks. The mortality rates of mice administered with MINO (p.o./i.p.), KM (p.o.), NFLX (p. o./i.p.) were significantly lower than those of the control group. Both the bacterial number and VT2c level in the feces of the FOM group were lower than those of the NFLX group on day 1, but reversed on day 3. In an in vitro experiment, each of the four drugs in combination with mitomycin C (MMC) caused a more significant decrease in the bacterial number than sole MMC, and they consequently indicated the suppressive effect on the release of VT2c. PMID- 10536296 TI - A comparison of the effects of two dinitroanilines against Cryptosporidium parvum in vitro and in vivo in neonatal mice and rats. AB - The effects of two dinitroanilines, oryzalin and trifluralin, were compared against Cryptosporidium parvum, in vitro using HCT-8 cells and in vivo using neonatal Swiss ARC mice and Wistar neonatal rats. In vitro, oryzalin and trifluralin exhibited IC(50) values (concentration necessary to cause a 50% inhibition) of 750 and 800 nM, respectively. A viability assay showed that neither compound produced a cytotoxic effect on the host cells at concentrations as high as 1 microM. The in vivo component of this study consisted of inoculation of neonatal mice and neonatal rats with 10(5) viable oocysts of C. parvum per animal and the subsequent treatment of this infection with trifluralin and oryzalin administered via gastric intubation. At doses of 100 mg kg(-1) body weight administered twice daily for 3 consecutive days, trifluralin had no statistically significant effect on the number of oocysts recovered from the gut of either rats or mice compared with controls, whereas at the same concentration, oryzalin caused 90 and 79% inhibition of oocysts recovered from mice and rats, respectively. PMID- 10536297 TI - Cytokine-induced fungicidal activity of human polymorphonuclear leukocytes against Penicillium marneffei. AB - We investigated the fungicidal activity of human polymorphonuclear leukocytes (PMN) against Penicillium marneffei. The yeast cells were cocultured in vitro with PMN for 24 h. Microscopic examination was also performed to examine the germination of yeast cells and their transformation to hyphal form during culture. Unstimulated PMN inhibited fungal growth when used at a higher effector/target (E/T) ratio but inhibited germination at a lower E/T ratio. We also examined the effects of various PMN-activating cytokines, including granulocyte-macrophage colony stimulating factor (GM-CSF), G-CSF, interleukin (IL)-8, interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha on the activity of PMN. Among these, GM-CSF, G-CSF and IFN-gamma enhanced PMN activity from being fungistatic to fungicidal. However, the other cytokines had little or no effect on PMN activity. In contrast, all tested cytokines enhanced PMN inhibitory effects on germination and morphological changes of P. marneffei. These antifungal activities were most strongly induced by GM-CSF. The combined use of any of the above cytokines failed to synergistically enhance antifungal PMN activity. Our results demonstrated that cytokine-activated PMN exert a significant antifungal activity, by suppressing the growth and germination of P. marneffei. Our results suggest that PMN may contribute to host resistance to infection against this fungal pathogen. PMID- 10536298 TI - Experimental murine mycotic mastitis: a sensitive and lenient model for studies of antifungal chemotherapy. AB - The murine model of mycotic mastitis was used to study the efficacy of amphotericin B (AmB). Twenty-four BALB/cJ mice at the fifth day of lactation were anesthetized and inoculated through the teat canal (two glands) with 50 microl suspension containing 5.0 x 10(7) cfu ml(-1) Candida albicans blastospores. Mice were randomly divided into two groups: untreated controls and AmB treated. Animals were euthanized 3 and 6 days after infection and treatment (4 mg kg(-1) per day intraperitoneally). The fungal burden of the mammary gland was determined by quantitative cultures. The number of C. albicans cells recovered from mammary gland homogenates were significantly lower in the AmB treated animals (both 3 and 6 days post-infection) than in the untreated controls (P<0.007 and P<0.003, respectively). The mammary glands of all untreated control animals showed marked neutrophilic infiltration, severe necrosis, and presence of blastospores, hyphae and pseudohyphae. In contrast, 10 of 12 animals treated with AmB showed only a mild neutrophilic infiltration which was restricted to alveoli and excretory ducts. All extra-mammary organs were free of infection in both groups. The results demonstrate that the murine mycotic mastitis model is suitable for investigations of new antifungal compounds. In addition, this model is more lenient than the systemic candidiasis models. PMID- 10536300 TI - The effect of probiotic bacteria on the adhesion of pathogens to human intestinal mucus. AB - Human intestinal glycoproteins extracted from faeces were used as a model for intestinal mucus to investigate adhesion of pathogenic Escherichia coli and Salmonella strains, and the effect of probiotics on this adhesion. S-fimbriated E. coli expressed relatively high adhesion in the mucus model, but the other tested pathogens adhered less effectively. Probiotic strains Lactobacillus GG and L. rhamnosus LC-705 as well as a L. rhamnosus isolated from human faeces were able to slightly reduce S-fimbria-mediated adhesion. Adhesion of S. typhimurium was significantly inhibited by probiotic L. johnsonii LJ1 and L. casei Shirota. Lactobacillus GG and L. rhamnosus (human isolate) increased the adhesion of S. typhimurium suggesting that the pathogen interacts with the probiotic. PMID- 10536299 TI - The effect of orally administered viable probiotic and dairy lactobacilli on mouse lymphocyte proliferation. AB - Four common Lactobacillus strains were screened for their effects on proliferation of mouse splenic lymphocytes. Mice received perorally 10(9) viable bacteria kg(-1) body weight for 7 days. Lactobacillus acidophilus treatment enhanced ex vivo basal proliferation (by 43%) and B-cell response at suboptimal and optimal concentrations of lipopolysaccharide (LPS) (by 27-28%). Conversely, Lactobacillus casei, Lactobacillus gasseri and Lactobacillus rhamnosus inhibited both basal proliferation (by 14-51%) and mitogen-stimulated lymphoproliferation, particularly at supra-optimal concentrations of concanavalin A (by 43-68%) and LPS (by 23-62%). Therefore, these Lactobacillus strains demonstrate strain specific effects on B- and T-cells and may also alter the splenocyte sensitivity to the cytotoxic effects of mitogens. PMID- 10536301 TI - Secretory proteins of Mycobacterium habana induce a protective immune response against experimental tuberculosis. AB - The capacity of proteins of Mycobacterium habana TMC 5135 secreted into culture medium during the mid-exponential growth phase (secretory proteins, SPs) to induce protective immunity against Mycobacterium tuberculosis H37Rv was studied in the mouse model. Mice immunized with SPs followed by a challenge with M. tuberculosis H37Rv showed lesser M. tuberculosis bacilli in their lung and spleen and survived longer than unimmunized controls. The findings suggest that SP antigens of M. habana are protective against tuberculosis infection. PMID- 10536302 TI - Immunological cross-reactivity between the vaccine and other isolates of Streptococcus bovis and Lactobacillus. AB - Recent studies have showed that immunisation with Streptococcus bovis (Sb-5) and Lactobacillus (LB-27) may confer protection against lactic acidosis in sheep and cattle. The present study was designed to determine the degree of immunological cross-reactivity between Sb-5 and eight other strains of Streptococcus bovis; and between LB-27 and four other isolates of Lactobacillus. The cross-reactivity index (CRI, a low CRI indicates a high degree of immunological cross-reactivity) ranged from 7.3 to 56.1% between the strains of S. bovis (the encapsulated strains with CRIs ranging from 7.3 to 12.4%). For isolates of Lactobacillus the CRIs ranged from 11.5 to 72.2%. The results indicate that all the isolates tested have a certain degree of immunological homology with Sb-5 and LB-27, and suggest that the vaccine may cross-react with a large number of strains of S. bovis and Lactobacillus which may cause lactic acidosis. As most of the S. bovis strains in the rumen are encapsulated, the high degree of homology between Sb-5 and encapsulated S. bovis strains further suggests that the vaccine containing Sb-5 may be effective against a wide range of strains of S. bovis in sheep and cattle. PMID- 10536303 TI - Purification and characterization of outer membrane protein P6, a vaccine antigen of non-typeable Haemophilus influenzae. AB - Outer membrane protein P6 is a promising vaccine antigen with potential to prevent infections caused by non-typeable Haemophilus influenzae. A convenient and reliable method for the purification of P6 and an assessment of the purity, yield, protein structure, antigenicity and immunogenicity of the purified protein are described. The method begins with intact H. influenzae and utilizes a series of incubations and centrifugations using a single buffer to remove all cell components with the exception of the peptidoglycan to which the P6 is associated. P6 is dissociated from the complex with heat and the insoluble peptidoglycan is removed by centrifugation. The procedure yields highly purified P6. Contamination with lipooligosaccharide is less than 0.025 endotoxin U per microgr P6. The yield of P6 is approximately 2 mg of P6 per l H. influenzae culture. The purified P6 retains both the secondary and tertiary structure as measured by circular dichroism and analysis with monoclonal antibodies. The purified P6 is immunogenic in animals. A convenient method for purifying P6 which retains antigenicity and immunogenicity will be an important tool for future studies of the vaccine potential of P6. PMID- 10536304 TI - In vivo induction of CTL responses by recombinant adenylate cyclase of Bordetella pertussis carrying multiple copies of a viral CD8(+) T-cell epitope. AB - Bordetella pertussis adenylate cyclase toxin (ACT) is one of the few known protein toxins penetrating directly into the cytosol of target cells across their cytoplasmic membrane without the need for endocytosis. This capacity of ACT was recently exploited for in vivo delivery of single viral CD8(+) T-epitopes into MHC class I-presenting cells and induction of protective antiviral cytotoxic T cell (CTL) responses. Here, we have explored the potential of the cell-invasive adenylate cyclase domain of the toxin to deliver larger antigens by evaluating the epitope-specific CTL responses induced by constructs bearing one to four copies of the CD8(+) T-epitope from the nucleoprotein of the lymphocytic choriomeningitis virus. The increase in the number of copies of the epitope was accompanied by a moderate decrease of the specific cell invasiveness of the ACT protein and did not lead to further enhancement of the level of induced epitope specific CTL cells in mice, as compared to ACT with a single copy of the epitope. These results demonstrate the capacity of ACT to deliver larger heterologous antigens comprising several epitopes for antigenic presentation in vivo. PMID- 10536305 TI - Hamycin treatment of candidiasis in normal and diabetic rats. AB - Hamycin, a heptaene antifungal antibiotic was compared with amphotericin B in the treatment of established systemic infection with Candida albicans in normal and diabetic rats. In normal rats, orally administered hamycin at 10 mg kg(-1) per day for 7 days reduced Candida colony counts in the kidneys and livers as well as amphotericin B did and was nearly as effective as amphotericin B in a 21-day treatment trial. There was no further reduction in Candida colony counts when normal rats were treated with hamycin at 25 mg kg(-1) twice a day for 7 days. In streptozotocin induced diabetic rats, hamycin at 20 mg kg(-1) per day for either 7 or 21 days compared favourably with amphotericin B in efficacy. Results of the present study suggest that oral hamycin may be useful in the treatment of established disseminated candidiasis in normal as well as diabetic hosts. PMID- 10536306 TI - Molecular and biochemical mechanisms of drug resistance in fungi. AB - This paper reviews the current status of our understanding of resistance mechanisms of three major classes of antifungal drugs for systemic use, amphotericin B (AMPH), flucytosine (5-FC) and several azole antifungals, in particular fluconazole (FLCZ), at the molecular and cellular levels. Although the number of reports of AMPH- or 5-FC-resistant fungal species and strains is limited, several mechanisms of resistance have been described. AMPH-resistant Candida have a marked decrease in ergosterol content compared with AMPH susceptible control isolates. A lesion in the UMP-pyrophosphorylase is the most frequent determinant of 5-FC resistance in C. albicans. Recently resistance of C. albicans to azoles has become an increasing problem. Extensive biochemical studies have highlighted a significant diversity in mechanisms conferring resistance to FLCZ and other azoles, which include alterations in sterol biosynthesis, target site, uptake and efflux. Among them, the most important mechanism clinically is reduced access of the drug to the intracellular P450 14 DM target, probably because of the action of a multidrug resistance efflux pump, and overproduction of that target. However, other possible resistance mechanisms for azoles remain to be identified. PMID- 10536307 TI - Antifungal activities of D0870 against fluconazole-resistant Candida albicans. AB - We compared the in vitro and in vivo antifungal activities of D0870, a new triazole antifungal agent, with those of other antifungal agents against 8 clinical isolates of fluconazole-resistant Candida albicans. Microdilution testing was performed according to National Committee for Clinical Laboratory Standards (NCCLS) document M27-T. Minimal inhibitory concentration of D0870 (<0.004-1.0 micro g/ml) was lower than those of fluconazole (2->64 microEg/ml) and itraconazole (0.031-8.0 microEg/ml). In systemic infection models with C. albicans in normal and immunosuppressed mice, D0870 at 0. 3-30 mg/kg/day for 5 days after infection prolonged survival of the animals and showed the highest efficacy among the triazole antifungal agents. At pH 7 and 37C in Sabouraud dextrose broth (SDB), D0870 inhibited the growth of C. albicans and acted cytocidally against one of the middle-resistant strains. In an in vivo study against this strain, D0870 at 10 mg/kg/day for 5 days after infection significantly reduced kidney colony counts (2850+406-997+537 CFU/kidney, P<0.05) on day 7 after infection in comparison with those of the control mice at 24 h after infection. Plasma concentration of D0870 after a single oral administration at 10 mg/kg maintained a sufficient level for interpretation of in vivo antifungal activities. These results suggest that D0870 has strong antifungal activities against clinical isolates of fluconazole-resistant C. albicans in vitro and in vivo, and that these strong activities are at least partially concerned with the fungicidal action. PMID- 10536308 TI - Cytotoxic activity and cytokine gene induction of Asp-hemolysin to murine macrophages. AB - We examined the effects of Asp-hemolysin from Aspergillus fumigatus Fresenius Muramatsu strain on the viability and cytokine gene expression of mouse peritoneal macrophages. The cytotoxic activity of Asp-hemolysin to macrophages cultured in FCS-RPMI medium was increased in a dose-dependent manner. Treatment of Asp-hemolysin with N-ethylmaleimide or sulfo-N-hydroxy-sulfosuccinimide acetate caused a remarkable loss of the cytotoxic activity, however, the cytotoxic activity of Asp-hemolysin to macrophages cultured in serum-free medium was significantly increased as compared with that in FCS-RPMI medium. As other biological activities of Asp-hemolysin, tumor necrosis factor-alfa (TNF-alfa), interleukin-1 beta(IL-l beta) and interleukin-1 alfa (IL-l alfa) mRNA expression were observed in macrophages cultured with 1 micro g/ml of Asp-hemolysin. PMID- 10536309 TI - Clinical evaluation of diagnostic methods using plasma and/or serum for three mycoses: aspergillosis, candidosis, and pneumocystosis. AB - Clinical evaluation was retrospectively made of the results of serological diagnostic methods using plasma and/or sera of patients for the diagnosis of aspergillosis, candidosis, and pneumocystosis. Specimens were drawn from 8 patients with invasive aspergillosis, 3 with aspergilloma, 9 with candidosis, 4 with pneumocystosis, and 15 with no fungal infections. In invasive aspergillosis, the sensitivities of the (1-3)-beta-D-glucan measurement test using chromogenic and turbidimetric methods were 78.6% and 82.1%, with specificities of 75% and 87.5%, respectively. The sensitivity of the Pastorex Aspergillus test for invasive aspergillosis was 16.7%, with a specificity of 92.3%. In candidosis, the sensitivities of the (1-3)-bata-D-glucan test using the above two methods were 84.2% and 100%, with specificities of 75% and 87.5%, respectively. The sensitivity of the CAND-TEC test and the Pastorex Candida test for candidosis were 68.8% and 16.7%, with specificities of 57.1% and 100%, respectively. These results indicate that the (1-3)-bata-D-glucan measurement methods are more reliable in clinical application than the other antigen detection methods, but they still lack efficiency in differentiating fungal infections such as aspergillosis, candidosis and pneumocystosis. For a more exact diagnosis of systemic fungal infections, detailed studies on the clinical symptoms are considered essential. PMID- 10536310 TI - A case of melanonychia caused by Exophiala dermatitidis. AB - We report a case of a healthy 61-year-old woman with discoloration of the nail on her right big toe. We first treated her with topical steroid and urea under suspected diagnosis of nail eczema, but the lesion remained. In culture, black, shiny, pasty and yeast-like colonies grew repeatedly. Examination of debris from her nail showed dematiaceous spherical cells and hyphal elements. Microscopically, annelloconidia were produced at the apical ends of anellidic conidiogenous cells. This colony grew at 40C. Mitochondrial DNA restriction fragment length polymorphism was analysed in this strain and its restriction pattern confirmed the isolate to be Exophiala dermatitidis. Based on these findings, we diagnosed this nail deformity as fungal melanonychia due to Exophiala dermatitidis. This is the third reported case of this disease. PMID- 10536311 TI - The effect of an Aspergillus fumigatus derived elastolytic proteinase on human neutrophils. AB - The effect of elastolytic proteinase on quantitative nitroblue tetrazolium (NBT) dye reduction and chemotaxis of human neutrophil was examined. Elastolytic proteinase is derived from Aspergillus fumigatus 9409. NBT dye reduction was inhibited significantly by 133microEg/ml of elastolytic proteinase (p<0.05). Chemotaxis was also inhibited significantly by 400microEg/ml of the enzyme, compared to the control group (p<0.05). These findings suggest that elastolytic proteinase, depending on its concentration, acts as an inhibitory agent to both NBT dye reduction and chemotaxis activities in the human neutrophil. The findings that this enzyme suppressed neutrophil function in A. fumigatus infection is of importance, because it is now suspected that the enzyme has the potential to cause infection. PMID- 10536312 TI - Site specificity of colorectal neoplasms in families without an inherited syndrome. AB - BACKGROUND: Relatives of patients with bowel neoplasia have an increased risk of bowel neoplasia. If there were concordance in location of neoplasia between relatives, then location-specific screening could be used. Such concordance might also assist in the understanding of the etiology of neoplasia within individual families. METHODS: We have investigated the concordance in anatomic location of colonic neoplasia between first-degree relatives using a new statistical technique for paired data called alternating logistic regression. RESULTS: A total of 146 families were ascertained, none of which had clinical evidence of a hereditary predisposition to edon neoplasia. Among those with neoplasia, there was an increased risk for right-sided disease with older age (40% for less than age 60 vs. 58% for at least 70 years of age, p = 0.008). As assessed by the odds ratio, we found no significant concordance within families for location of neoplasia (odds ratio = 1.2: CI [0.7, 2.2]), although there was a suggestion that location in family members of the same generation was more strongly associated (odds ratio 1.87: CI [0.82, 4.25]). CONCLUSIONS: The lack of concordance within families for location argues against considering family-specific bowel screening protocols and indicates that the most important causes of bowel neoplasia are not sufficiently focused on one anatomic site to facilitate etiologic research. PMID- 10536313 TI - Endoscopic mucosal resection with a cap-fitted endoscope versus freehand gastric mucosectomy in an animal model. AB - BACKGROUND: Endoscopic mucosal resection with a cap-fitted endoscope appears to be as effective but easier to perform than freehand mucosectomy. However, experience with this technique has been limited to small case series and there is a lack of data from direct comparison trials. METHODS: Nine pigs were randomized to mucosectomy using a cap-fitted endoscope or the freehand technique. Five mucosal resections were performed at five different sites in the gastric body in each pig. Eight to ten milliliters of a saline, epinephrine, and methylene blue solution were injected to raise a bleb to simulate a mucosal lesion. Endoscopic mucosal resection with a cap-fitted endoscope was performed by suctioning the bleb into the cap device pre-looped with an oval snare; mucosectomy was performed electrosurgically. Freehand mucosectomy was performed by encircling and then resecting the bleb using an oval snare. The ease of procedure (1 = "very easy" to 5 = "unable to complete") was assessed after each resection. The animals were recovered, maintained, and then humanely sacrificed after 2 weeks. RESULTS: Five pigs underwent endoscopic mucosal resection with a cap-fitted endoscope and 4 underwent freehand mucosectomy. Eight animals survived for 2 weeks without complications and one animal from the freehand group died of massive hemorrhage within 48 hours of endoscopy. Both methods produced rounded resection specimens measuring 9 to 12 mm in diameter of the full thickness of the mucosa and the submucosa. Overall ease of resection was 1.84 +/- 0.52 for the cap-fitted group and 2.98 +/- 0.86 for the freehand group (p < 0.0001). All of the sites identified at autopsy were completely re-epithelialized, except for the five sites found in the pig that died prematurely. CONCLUSIONS: Endoscopic mucosal resection with a cap-fitted endoscope is as effective, safe, but easier to perform compared with freehand mucosectomy. PMID- 10536314 TI - Push enteroscopy in celiac sprue and refractory sprue. AB - BACKGROUND: The aim of this study was to determine in patients with sprue whether jejunal endoscopy improves the diagnostic yield or provides information that may modify management, when compared with evaluation limited to the duodenum. METHODS: From January 1994 to June 1998, a total of 31 patients (6 men, 25 women, mean age 41 years) were prospectively evaluated by push enteroscopy. They were divided into two groups: (1) celiac disease at different stages of activity (n = 23) and (2) refractory sprue (n = 8). The endoscopic and histologic findings in the duodenum and in the jejunum were compared. RESULTS: Celiac disease: In 19 patients, endoscopic and histologic findings in the duodenum and jejunum were similar; in four patients villous atrophy was more severe in the duodenum than in the jejunum. Refractory sprue: In 5 of 8 patients, enteroscopy revealed ulcerative jejunitis, whereas ulcerations were found in the duodenum in only one case. CONCLUSION: In refractory sprue, push enteroscopy with jejunal biopsies was of diagnostic value in 50% of cases demonstrating ulcerative jejunitis, whereas it did not modify the management of patients with responsive celiac disease. PMID- 10536316 TI - Endoscopic therapy in anomalous pancreatobiliary duct junction. AB - BACKGROUND: Anomalous pancreaticobiliary duct junction is associated with bile duct strictures, pancreatitis, choledochal cysts, and biliary carcinoma. Limited data are available for outcomes of endoscopic therapy. METHODS: Review of 7537 patients undergoing endoscopic retrograde cholangiopancreatography from 1988 to 1997 yielded 18 patients with anomalous pancreaticobiliary duct junction. Therapeutic responses were tallied by chart review and phone calls. RESULTS: There were 13 women and 5 men, with a mean age of 36 years. Twelve patients had no ductographic evidence of pancreatitis and 6 had chronic pancreatitis. Seven had choledochal cysts. Fifteen patients (83%) underwent endoscopic biliary sphincterotomy, two of whom underwent repeat endoscopic biliary sphincterotomy for recurrence of symptoms. The other therapies included stent placement for benign biliary strictures in 5 patients, lithotripsy of pancreatic stones in 1 patient, and choledochal cyst removal in 4 patients. Three cases with malignant biliary strictures are excluded from endoscopic outcome studies. The 12 patients with pancreatitis had a mean of 2.0 episodes per year before any treatment. After endoscopic therapy 7 patients had no further episodes of pancreatitis, whereas 5 patients had further episodes, with a mean of one additional attack per year, over 3 years mean follow-up. CONCLUSIONS: Patients with anomalous pancreaticobiliary duct junction have complex pathology associated with strictures, choledochal cysts, pancreatitis, and malignancies. Endoscopic therapy appeared to benefit 13 of 15 patients without malignant disease with elimination of or decreased frequency of pancreatitis. Endoscopic therapy appears to be a logical first step in the management of most symptomatic patients with anomalous pancreaticobiliary duct junction. PMID- 10536315 TI - Relationship of low-dose aspirin to GI injury and occult bleeding: a pilot study. AB - BACKGROUND: Low-dose aspirin is commonly used in patients with cardiovascular disease. However, little is known about the effect of aspirin on occult blood loss caused by gastrointestinal injury. Therefore, we studied endoscopic injury and fecal occult blood loss in patients ingesting different quantities of low dose aspirin. METHODS: Forty healthy volunteers were enrolled in a randomized, double-blind, prospective, pilot endoscopic study. Each volunteer underwent 30 days of treatment with daily aspirin 30 mg, 81 mg, 325 mg, or placebo. Subjects completed fecal occult blood test cards before and at the end of treatment of two types: guaiac impregnated (Hemoccult and Hemoccult SENSA) and immunochemical (HemeSelect and FlexSure OBT). Each volunteer underwent upper endoscopy at baseline and after completing 30 days of study medication. RESULTS: Aspirin did not induce significant injury as determined by endoscopy when compared with placebo. Six of 30 volunteers taking aspirin developed erosions, whereas 1 of 10 subjects on placebo developed a new erosion (p = 0.66). Aspirin (325 mg) was associated with a higher mean symptom score than the lower aspirin dosages and the placebo group (p = 0.12). Only one subject taking aspirin (325 mg) had fecal occult blood on a single HemeSelect card. No subject had a positive fecal occult blood test with Hemoccult II, Hemoccult II SENSA, or FlexSure OBT cards. CONCLUSIONS: Aspirin in dosages commonly used for cardiovascular prophylaxis does not generally cause significant gastric or duodenal mucosal endoscopic lesions. In the absence of frank ulceration, low-dose aspirin does not result in positive fecal occult blood tests. Low-dose aspirin should not interfere with fecal occult blood testing and probably should not be stopped during stool collection. PMID- 10536317 TI - Impact of skill and experience of the endoscopist on the outcome of endoscopic sphincterotomy techniques. AB - BACKGROUND: Our aim was to assess the influence of the skill and experience of the endoscopist on the success and risk of endoscopic sphincterotomy techniques. METHODS: The outcome of all endoscopic sphincterotomies (n = 1335) carried out between 1988 and 1995 were retrospectively analyzed with respect to the endoscopist performing the procedure. Endoscopists were differentiated according to whether they had previous experience with endoscopic sphincterotomy techniques (n > 100) and the frequency of endoscopic sphincterotomy during the study period (>40, 26 to 40, 10 to 25, <10 per year). RESULTS: Indications for endoscopic sphincterotomy techniques and technical execution had only a minor influence on the results of endoscopic sphincterotomy and were comparable for the individual endoscopists. The overall success rate of endoscopic sphincterotomy was 94.4% and did not significantly differ among the endoscopists. The overall complication rate of endoscopic sphincterotomy was 7.3%. Endoscopists learning endoscopic sphincterotomy techniques with a case frequency of less than 10 procedures per year had a consistently high complication rate (10.5%). Those learning endoscopic sphincterotomy techniques with a case frequency of more than 25 procedures per year had an above-average complication rate for their first 40 endoscopic sphincterotomy procedures and a significant decrease in complication rate as the number of procedures increased. The complication rate for experienced endoscopists was 7.7%. There were distinct and, in one case, significant differences in complication rates between individual endoscopists (11.5% vs. 4.8%, p = 0.01). However, when corrected for multiple testing, there were no significant differences at the p < 0. 05 level. The endoscopic sphincterotomy frequency of the endoscopist was the only significant risk factor for complications. Endoscopists with a frequency of more than 40 procedures per year had a significantly lower complication rate (5.6%) than endoscopists with a lower case frequency (9.3%, p < 0.05). CONCLUSIONS: A low endoscopic sphincterotomy frequency is, even for endoscopists with previous experience with the procedure, a risk factor for complications after endoscopic sphincterotomy. The learning of endoscopic sphincterotomy techniques requires a minimum of 40 procedures, but also after 100 procedures a further decrease of the complication rate can be expected. PMID- 10536318 TI - Frequency of abnormal pancreatic and biliary sphincter manometry compared with clinical suspicion of sphincter of Oddi dysfunction. AB - BACKGROUND: Sphincter of Oddi manometry as performed at ERCP is the most accepted method to evaluate for sphincter of Oddi dysfunction. To fully assess for sphincter of Oddi dysfunction, both the pancreatic and the bile ducts must be evaluated. We assessed the frequency of pancreatic and biliary sphincter abnormalities in a large series of patients. METHODS: A total of 593 patients underwent manometry of the biliary and pancreatic ducts at one endoscopic retrograde cholangiopancreatography session. Basal sphincter pressure greater than or equal to 40 mm Hg was considered abnormal. Phasic waves were not evaluated. Manometric abnormalities were correlated with the clinical presentation as categorized using a modified Geenen/Hogan classification. RESULTS: Of 360 patients with intact sphincters, 18.9% had abnormal pancreatic sphincter basal pressure alone, 11.4% had abnormal biliary basal sphincter pressure alone, and in 31.4% the basal pressure was abnormal for both sphincters; thus, 60.1% of the patients had sphincter dysfunction. The frequency of sphincter of Oddi dysfunction did not differ whether typed by biliary or pancreatic criteria: approximately 65% type II and 59% type III. Of patients without pancreatitis, 55.9% had an abnormal basal sphincter pressure, whereas sphincter dysfunction was present in 72.3% of those with idiopathic pancreatitis and 53.9% of patients with chronic pancreatitis. Of patients with an ablated biliary sphincter, 45.9% had abnormal basal pancreatic sphincter pressure and only 0.6% had an abnormal biliary sphincter pressure alone. Abnormal pressure in both sphincters was found in 9.3%. CONCLUSION: If both portions of the sphincter of Oddi are studied simultaneously, abnormalities are found very commonly (55% to 72%). Assessment of both sides of the sphincter is necessary. Classifying patients according to both pancreatic and biliary sphincter segments is cumbersome, and may be replaced by an overall type. Using this modified classification, the frequency of sphincter of Oddi dysfunction is similar in both type II and type III patients (59% to 67%). PMID- 10536319 TI - Long-term experience with the transpancreatic sphincter pre-cut approach to biliary sphincterotomy. AB - BACKGROUND: The transpancreatic duct pre-cut to gain access to the bile duct for diagnostic and therapeutic maneuvers has been described as useful, but questions of efficacy and safety remain to be resolved. METHODS: To further evaluate this technique, we performed a review on 200 consecutive endoscopic sphincterotomies. Standard direct biliary sphincterotomy was performed in 143 patients and transpancreatic duct pre-cut in 51 patients. RESULTS: The overall complication rate for the standard sphincterotomy was 2.1%; that for the transpancreatic approach was 1.96%. There were no cases of post-ERCP pancreatitis after transpancreatic duct pre-cut sphincterotomy. The length of hospital stay was 1 day or less for 192 patients, 2 days for 5 patients, 4 days for 1 patient and 7 days for 2 patients. In 2 patients there was failure to enter the bile duct despite the pre-cut. In one, the procedure was successful at a second attempt 48 hours later. CONCLUSIONS: Transpancreatic duct pre-cut is a safe and effective method for gaining quick access to the bile duct in patients in whom cannulation is difficult. PMID- 10536320 TI - Biliary changes in extrahepatic portal venous obstruction: compression by collaterals or ischemic? AB - BACKGROUND: The postulated mechanisms of biliary abnormalities in extrahepatic portal venous obstruction (EHPVO) are either extrinsic compression by collaterals or ischemic injury due to venous thrombosis. If the former hypothesis is correct, then biliary changes should revert to normal after portasystemic shunt surgery. METHODS: Five patients with EHPVO who underwent portasystemic shunt surgery were studied. One of these patients had obstructive jaundice due to portal cavernoma. Endoscopic retrograde cholangiography (ERC) was performed before as well as after the shunt surgery. Doppler ultrasound and splenoportovenography were obtained to confirm the diagnosis of EHPVO as well as shunt patency. RESULTS: All patients had biliary abnormalities on pre-shunt ERC. The post-shunt ERC showed partial reversal of biliary abnormalities in 3 patients, complete reversal in 1 patient, and no reversal in 1 patient. Smooth strictures opened after shunt surgery and proximal dilatation disappeared in most patients. The indentations and caliber irregularities disappeared after shunt surgery, whereas angulations and ectasias of biliary ducts persisted. CONCLUSION: Shunt surgery results in regression of some of the biliary abnormalities and relieves biliary obstruction, suggesting that mechanical compression by collaterals is the mechanism behind biliary abnormalities in EHPVO. However, some biliary changes persist after shunt surgery signifying fixed obstruction due to ischemia or fibrous scarring. Thus, the two theories are not mutually exclusive. PMID- 10536321 TI - How many biopsies should be performed during percutaneous transhepatic cholangioscopy to diagnose biliary tract cancer? AB - BACKGROUND: The sensitivity of biopsy in the diagnosis of cholangiocarcinoma using percutaneous transhepatic cholangioscopy is not well defined. METHODS: Patients with a biliary tract malignancy (n = 52) underwent directed biopsy during percutaneous transhepatic cholangioscopy using a 1.8 mm diameter forceps. Histologic findings were correlated with endoscopic appearance. RESULTS: A diagnosis of carcinoma was made in all four patients with a tumor of the major duodenal papilla and in all 15 patients with a polypoid bile duct tumor with two biopsies from the mass. In 19 patients with stenotic bile duct cancer, a positive diagnosis was made in 95% of cases when three biopsies were taken from the margin of the stenotic area. When cholangioscopy showed a tortuous, dilated vessel (n = 10), the diagnosis of cancer was made with two biopsies taken from the margin of the stenosis. In 14 patients with metastatic bile duct cancer, the diagnosis was made in only 43% of cases when three biopsies were taken from the margin of the stenosis. When combined with results from the three biopsies taken from within the area of stenosis, the sensitivity for diagnosing pancreatic cancer improved from 20% to 60%. CONCLUSIONS: Directed cholangioscopic biopsies are highly sensitive for the diagnosis of cholangiocarcinoma but less sensitive for cancer metastatic to the bile duct. The numbers and locations of the biopsies required to make a diagnosis of carcinoma depend on the origin and cholangioscopic appearance of the tumor. PMID- 10536322 TI - Diagnosis of the depth of invasion of gallbladder carcinoma by EUS. AB - BACKGROUND: The prognosis of gallbladder carcinoma is dismal and relates to the depth of invasion as expressed by the T factor in TNM staging. We evaluated the utility of endoscopic ultrasound (EUS) in the diagnosis of the depth of invasion of gallbladder cancer. METHODS: Thirty-nine patients who underwent both EUS and surgery were included in this study. The EUS images were classified according to the relation between tumor echo pattern and gallbladder-wall structure, and the resulting types were compared with depth of invasion as determined histologically. Based on the results, a set of diagnostic criteria is proposed. RESULTS: The EUS images were classified into four categories. Type A is a pedunculated mass with a fine-nodular surface and intact neighboring wall. Type B is a broad-based mass with an irregular surface and intact outer hyperechoic layer of the adjacent wall. In type C, the outer hyperechoic layer is irregular due to a mass echo, whereas, in type D, the outer hyperechoic layer is disrupted by a mass echo. Each of the four categories of EUS images correlated well with the histologic depth of invasion. CONCLUSION: EUS is useful in the T staging of gallbladder cancer. PMID- 10536323 TI - Laparoscopic observations of hepatic capsular abnormalities: non-postoperative adhesions and hepatic capsular thickening. AB - BACKGROUND: Hepatic capsular abnormalities (adhesions or thickening) are often striking at laparoscopy. However, their diagnosis is difficult because capsular abnormalities can also be caused by several pathologic conditions. The aim of this study was to systematically investigate the associated factors and prevalence of laparoscopically observed non-postoperative adhesions and hepatic capsular thickening. METHODS: We reviewed all data and studied laparoscopically observed hepatic capsular abnormalities (non-postoperative adhesions and thickening) in 2500 consecutive patients who underwent laparoscopy from 1981 to 1997. RESULTS: Non-postoperative adhesions were observed in 14.6% of cases and their frequency increased with age. Although several types of adhesions, from band-like to membrane-like, were seen, there were no correlations between type and underlying pathologic conditions, except tuberculous peritonitis with membrane-like adhesions and Fitz-Hugh-Curtis syndrome with violin string-like adhesions. Hepatic capsular thickening was observed in 9.7% of cases. The main associated factor was viral hepatitis followed by other liver diseases. CONCLUSIONS: Hepatic capsular abnormalities are observed relatively frequently (21.5%) during laparoscopy. Initial laparoscopic diagnosis of non-postoperative adhesions may help in selecting patients with tuberculous peritonitis and Fitz Hugh-Curtis syndrome for appropriate treatment. PMID- 10536324 TI - Endoscopic and histologic healing of Crohn's (ileo-) colitis with azathioprine. AB - BACKGROUND: The correlation between disease activity and endoscopic findings in Crohn's disease is poor. Corticosteroids induce symptom relief without consistent improvement of endoscopic lesions. Our aim was to examine the effect of azathioprine therapy on healing of inflammatory lesions in patients with severe Crohn's disease. METHODS: The study included 20 consecutive patients with Crohn's colitis or ileocolitis in clinical remission while taking azathioprine for at least 9 months and no corticosteroids for at least 3 months who had had an ileocolonoscopy less than 1 year before the start of azathioprine. All 20 patients underwent a new ileocolonoscopy with biopsies. RESULTS: The duration of azathioprine therapy was 24.4 +/- 13.7 months. In the colon, we observed complete healing in 14 of 20 patients (70%), near-complete healing in 2 of 20 (10%), partial healing in 3 of 20 (15%), and no healing in 1 of 20 (5%). In the ileum complete healing was seen in 7 of 13 (54%) patients with ileitis, near-complete healing in 2 of 13 (15%), partial healing in 1 of 13 (8%), and no healing in 2 of 13 (15%); the ileum was not reached in 1 patient. Histologic examination revealed disappearance of the inflammatory infiltrate, with a certain degree of architectural disturbance remaining. CONCLUSIONS: Successful azathioprine therapy is accompanied by mucosal healing and disappearance of the inflammatory infiltrate. PMID- 10536325 TI - Necrotizing acute pancreatitis caused by tiny carcinoma in adenoma in Vater's papilla. PMID- 10536326 TI - Mini-detachable snare ligation for the treatment of esophageal varices. AB - BACKGROUND: We manufactured and studied the usefulness of a newly designed mini detachable snare in the treatment of esophageal varices. The use of a multiple rubber band ligator, although generally effective and well tolerated, has certain limitations, including high cost, reduced visual field, and inadvertent band release. METHODS: We performed a prospective randomized controlled trial of the use of mini-detachable snare ligation vs. multiple band ligation in patients with recent or active esophageal variceal bleeding. The outcomes assessed were immediate hemostasis and rates of recurrent bleeding, eradication, and recurrence. RESULTS: From March 1997 to October 1998, 103 patients were entered into this trial; 46 underwent mini-detachable snare ligations and 57 multiple band ligations. Among patients with active bleeding, hemostasis was achieved in 6 of 7 (86%) in the mini-detachable snare ligation group and 11 of 13 (85%) in multiple band ligation group. Recurrent bleeding after initial treatment occurred in 2 of 46 (5.5%) in the mini-detachable snare ligation group and 3 of 57 (5.3%) in the multiple band ligation group. Esophageal varices were eradicated or reduced to grade I in 4.8 +/- 2.1 and 4.5 +/- 1.9 sessions in the mini-detachable snare ligation group and multiple band ligation group, respectively. The recurrence rate was 5 of 46 (11%) and 6 of 57 (11%) in the mini-detachable snare ligation group and multiple band ligation group during a follow-up period of 6 and 16 months, respectively. No serious complication occurred in either group. CONCLUSION: The mini-detachable snare is a new device that provides safe and effective treatment for esophageal varices that is comparable to multiple band ligation. PMID- 10536328 TI - Detection of the fluorescence of GI vessels in rats using a CCD camera or a near infrared video endoscope. AB - BACKGROUND: Choroidal near-infrared fluorescent angiography can detect vessels in the eye with high resolution. Observation of fluorescent gastrointestinal (GI) vessels by endoscopy may be useful in portal hypertension and bleeding ulcer. We here describe a technique for the detection of fluorescent GI vessels with a CCD camera or a near-infrared video endoscope. METHODS: Laparotomy was performed on rats. A tissue target was excited by means of a laser diode. We took pictures of tissue under both white and near-infrared light, both before and after intravenous injection of indocyanine green. Fluorescent light was selected by means of filters placed in front of the lens of a CCD camera or a near-infrared video endoscope. RESULTS: Under near-infrared light and after dye injection, we observed fluorescent vessels in real time and distinguished arterial from venous fluorescence. CONCLUSIONS: This device permits visualization of GI vessels, which could be useful for diagnosis of vascular abnormalities during endoscopy and surgery. PMID- 10536327 TI - Development and application of endoloops for the treatment of bleeding esophageal varices. AB - BACKGROUND: Endoloops are detachable nylon snares. The aims of this study were to develop an endoscopic method for repeated delivery of endoloops to arrest variceal bleeding, to compare efficacy of endoloop hemostasis with injection and band ligation in experimental models of bleeding, and to test the reliability and safety of endoloops in a pilot study in patients with varices. METHODS: Technical modifications including ridged endcaps and alterations in angulation of endoloops were developed to speed delivery and improve efficacy. Hemostatic efficacy of endoloops was compared with sclerotherapy and band ligation in animal studies before studies in patients. RESULTS: Modified endcap and endoloops allowed repeated applications without withdrawal of the endoscope. Right-angled endoloops ensnared more (p < 0.0001) gastric tissue and were more reliable (p < 0.05) than straight endoloops. Injection therapy and prestretched bands appeared ineffective, whereas band ligation was only effective on vessels up to 2 mm in diameter. Only endoloops achieved hemostasis on vessels of 3 to 5 mm (p < 0.05). No significant complications occurred using endoloops in animal (esophagus n = 20, stomach n = 20) or human (n = 11) studies. CONCLUSIONS: Endcap and endoloop modifications simplified repeated application to varices. Endoloops were more effective than injection or band ligation in experimental hemostasis and appeared safe and effective in patients. PMID- 10536330 TI - A case of multiple gastric carcinoids associated with multiple endocrine neoplasia type 1 without hypergastrinemia. PMID- 10536329 TI - Cronkhite-Canada syndrome associated with triple gastric cancers: a case report. PMID- 10536331 TI - Biliary strictures and cholangitis secondary to ascariasis: endoscopic management. PMID- 10536332 TI - Cytomegalovirus colitis presenting with the endoscopic appearance of pseudomembranous colitis. PMID- 10536333 TI - A successful single-step endoscopic resection of a 40 millimeter flat-elevated tumor in the rectum: endoscopic mucosal resection using sodium hyaluronate. PMID- 10536334 TI - New endoscopic finding associated with hyperplastic polyps. PMID- 10536335 TI - Superior mesenteric artery thrombosis after colonoscopy. PMID- 10536336 TI - Toothpick impaction with localized sigmoid perforation: successful colonoscopic management. PMID- 10536338 TI - Intrahepatic biloma: an unusual complication of cholangiocarcinoma treated endoscopically. PMID- 10536337 TI - Left hepatic duct cutaneous fistula after right hepatic lobe hydatid cyst operation treated with nasobiliary tube. PMID- 10536339 TI - Impact of endoscopic sphincter dilation on papillary structure: a case report. PMID- 10536340 TI - Endoscopic therapy for multiple mucosal bridges in the esophagus of a patient with Crohn's disease. PMID- 10536341 TI - Endoscopic technology: is this as good as it gets? PMID- 10536342 TI - Gastrointestinal endoscopy at the dawn of the new millennium. PMID- 10536343 TI - Upper GI bleeding. PMID- 10536344 TI - Comment PMID- 10536345 TI - Utilization of naloxone plus heater probe jet washing to localize friable colonic submucosal vascular ectasia. PMID- 10536346 TI - ERCP for intradiverticular papilla continues to be a challenge. PMID- 10536347 TI - Agonizing over AGUS. PMID- 10536348 TI - Cytologic and cytochemical features of adenoma malignum of the uterine cervix. AB - BACKGROUND: We previously reported that adenoma malignum (the mucinous type of minimal deviation adenocarcinoma [mucinous MDA]) of the uterine cervix expresses gastric phenotypes. The application of immunocytochemistry using HIK1083, a monoclonal antibody for gastric gland mucous cell mucin, enabled us to identify cellular clusters derived not only from typical lesions of mucinous MDA and less well differentiated lesions but also from gastric metaplasia. In this study, we tried to clarify the cellular features of such clusters in cervical and endometrial smears. METHODS: Twelve cases of mucinous MDA were studied. Papanicolaou stain and immunostaining with HIK1083 were performed on histologic and cytologic slides, including cervical and endometrial smears, decolorized smear slides, imprint slides of the tumors, and imprint slides of normal endocervical and gastric mucosa. RESULTS: Cellular clusters derived from metaplastic lesions, typical lesions, and less well differentiated lesions were found in 9, 4, and 2 cases, respectively. The cellular clusters derived from the metaplastic lesions lacked atypia but resembled those from gastric mucosa. Those derived from the typical lesions showed slight atypia. The cytoplasmic mucins of these cellular clusters, which were positive for HIK1083, were stained yellowish orange by Papanicolaou stain, whereas those of normal endocervical cells were negative for HIK1083 and were stained pinkish by Papanicolaou stain. CONCLUSIONS: Yellowish-orange staining of cytoplasmic mucins by the Papanicolaou method is an important diagnostic clue aiding identification of mucinous MDA and related lesions by cytology. Moreover, immunostaining with HIK1083 is a useful tool in the diagnosis of these lesions by cytology as well as by histology. Cancer (Cancer Cytopathol) PMID- 10536349 TI - Adenocarcinoma in situ with a small cell (endometrioid) pattern in cervical smears: a test of the distinction from benign mimics using specific criteria. AB - BACKGROUND: Papanicolaou smears have been less effective in preventing cervical adenocarcinoma than in preventing squamous carcinoma. One reason may be a lack of awareness of certain smear patterns of adenocarcinoma in situ (AIS) such as those with crowded small cells (endometrioid pattern). METHODS: A test set of 29 smears (17 AIS with an endometrioid pattern, 12 benign mimics) was reviewed by 11 cytologists (4 experienced cytotechnologists, 3 cytopathology fellows, and 4 cytopathologists with varying levels of experience). Participants were blinded as to the actual diagnosis and the number of cases in each category and were instructed to diagnose either AIS or a benign lesion. Results of this review were not disclosed before a second review conducted after instruction in specific criteria for "endometrioid" AIS. Results were compiled using kappa statistics. RESULTS: In the first round, the ability to distinguish these lesions was poor for 8 of the 11 reviewers, and no reviewer was in excellent agreement with the actual diagnosis. In the second round, only 1 reviewer had a poor rating, and 4 of 11 were in the excellent category. Misdiagnoses in both rounds were more commonly the result of underdiagnosis of AIS than overdiagnosis of benign cases. CONCLUSIONS: The presentation of AIS in smears as groups of crowded small cells is prone to underdiagnosis. Awareness of this problem and use of criteria improves sensitivity. [See editorial on pages 243-4, this issue.] Cancer (Cancer Cytopathol) PMID- 10536350 TI - Peritoneal washing cytology is unnecessary in gynecologic surgery for benign diseases. AB - BACKGROUND: The presence of tumor cells in peritoneal washing cytology specimens taken during surgery affects the staging of many gynecologic malignancies. Peritoneal washings are often collected routinely, even in cases of presumed benign disease. This study was designed to address whether evaluation of these specimens is justified. METHODS: We reviewed diagnostic reports from all peritoneal washings and the corresponding surgical pathology specimens from patients undergoing gynecologic surgery during a 1-year period in one institution and a 20-month period in the other. Cases were divided into benign and malignant categories based on the surgical pathology diagnosis. RESULTS: Three hundred forty-six patients had peritoneal washings collected during the study period. The proportion of cases with malignancy was 30% in one institution and 49% in the other. Of these, 119 had an endometrial or ovarian malignancy, including 16 ovarian tumors of low malignant potential. Malignant cells were detected in 19 cases. In 10 of these 19, grossly apparent peritoneal tumor implants were present at the time of surgery. The remaining 227 were found to have benign disease, and the peritoneal washing cytology diagnosis was negative in all cases. Potential savings of $13,000 to $17,000 based on current insurance reimbursement could have been realized for these 227 patients without compromising patient care. CONCLUSIONS: These data suggest that peritoneal washing cytology specimens collected at the time of gynecologic surgery for presumed benign disease can be held and processed later if an unsuspected malignancy is discovered. This practice can result in cost savings without compromising patient care. Cancer (Cancer Cytopathol) PMID- 10536351 TI - Quick-staining urinary cytology and bladder wash image analysis with an integrated risk classification: a worthwhile improvement in the follow-up of bladder cancer? AB - BACKGROUND: With an end toward an increase in patient quality of life, morphologic methods were tested for their combinatory value in expanding the effectiveness of follow-up appointments and finding a more specific supervision of patients with bladder cancer. METHODS: Voided urine and bladder washing specimens were gathered in 223 follow-up sessions of 124 patients with a history of bladder cancer. Hemacolor (Merck, Darmstadt, Germany)-stained cytospin preparations of voided urine specimens were ready for diagnosis within 15 minutes, and results were available shortly before cystoscopy. Feulgen-Schiff stained cytospin preparations of bladder washings entered the image analysis system. A special software was used to classify the DNA histogram by a risk factor for bladder cancer. RESULTS: Follow-up of patients revealed 83 tumor recurrences. Depending on the grade of the underlying tumor, the sensitivity of quick-staining cytology was 86.4%, 46.2%, or 13.6% for grade 3 to grade 1 TCC, respectively. Cytology and image analysis data demonstrated complementary potency. The combination of methods increased sensitivity to 90.9%, 66.7%, and 31.8%, respectively. Although 24 of 140 image analyses denoted high risk for bladder cancer without simultaneously visible tumor, correct evidence of high risk could be found for 92.2%. CONCLUSIONS: The combinatory use of quick-staining urinary cytology and bladder wash image analysis was demonstrated to be most valuable in diagnosing recurrent bladder cancer and selecting patients needing more intensive follow-up. At a minimum of patients discomfort, the tested combination also seems helpful to surpass diagnostic limits in cystoscopy and cytology caused by therapeutic effects on the bladder epithelium. Cancer (Cancer Cytopathol) PMID- 10536352 TI - Cytodiagnosis of well differentiated hepatocellular carcinoma: can indeterminate diagnoses be reduced? AB - BACKGROUND: Distinction of well differentiated hepatocellular carcinoma (HCC) from benign hepatocellular lesions is a well recognized problem in fine-needle aspiration (FNA) cytology, sometimes leading to indeterminate reports. The aim of this study was to critically examine criteria that might allow definitive diagnosis in these cases. METHODS: FNA smears and cell blocks from 65 patients with primary hepatocellular lesions were reviewed. Seventy separate samples had been obtained. The initial reports in these samples were: HCC in 34, benign findings in 27, and indeterminate findings in 9. We defined architectural and cytological features seen in the malignant cases but not seen in the benign cases, including an assessment of reticulin in cell blocks. These criteria were then applied to the indeterminate cases. RESULTS: The most specific cytologic criteria of malignancy in well differentiated HCC were (i) numerous stripped atypical nuclei, (ii) macronucleoli, (iii) increased mitoses, and (iv) multinucleation. The most specific architectural criteria in smears were (i) widened trabeculae, (ii) well defined capillaries traversing tissue fragments, and (iii) solid islands of hepatocytes rimmed by endothelial cells. The most valuable architectural criteria in cell blocks were (i) trabeculae greater than two cells thick and (ii) reduced or absent reticulin framework. Using the above criteria a retrospective diagnosis of HCC was possible in eight of the nine indeterminate cases, all but one of which have subsequently been confirmed as malignant. CONCLUSIONS: Close attention to architectural features in both smears and cell blocks should allow most well differentiated HCCs to be diagnosed by FNA cytology. A reticulin stain should be part of the routine assessment of cell blocks. Cancer (Cancer Cytopathol) PMID- 10536353 TI - Fine needle aspiration (FNA) biopsy of palpable breast masses: comparison of conventional smears with the Cyto-Tek MonoPrep system. AB - BACKGROUND: One of the limitations preventing the widespread use of fine-needle aspiration (FNA) is that it requires skill to obtain an adequate sample and well prepared smears. In this study, a new monolayer technique, the Cyto-Tek MonoPrep (MP) system, which obviates the need for smear preparation, was evaluated against conventional smear (CS) preparation for palpable breast lesions. METHODS: A total of 44 paired CS/MP breast FNA specimens were studied. The authors blindly analyzed the CS and the MP slides separately, then by a side-by-side evaluation. The two methods were compared with respect to diagnostic concordance, cellularity, cell preservation, background debris, and time needed to prepare and diagnose each case. RESULTS: An exact diagnostic correlation was present in 34 of 44 (77%) cases. The 10 noncorrelating cases were caused by decreased cellularity in the MP cases; nonetheless, 7 of these were correctly assigned to the right general diagnostic category, whereas the remaining 3 cases had insufficient cells. In addition to overall lesser cellularity on MP, fibroadenoma cases had smaller epithelial sheets and absence of stroma compared with CS. Both methods had comparable cellular preservation and diagnostic evaluation time, but background debris and preparation time were greater for MP. CONCLUSION: CS are favored over MP for the preparation of breast FNA specimens in centers with specialized FNA services because of their higher diagnostic yield, ease of preparation, and availability for immediate cytologic evaluation. However, in settings where specimens are collected sporadically by unskilled clinicians, the MP system may prove to be useful as an alternative or an adjunct to CS. Cancer (Cancer Cytopathol) PMID- 10536354 TI - Role of fine-needle aspiration in the clinical management of solid organ transplant recipients: a review. AB - BACKGROUND: We evaluated the clinical course of the solid-organ transplant population at our institutions to determine the role of fine-needle aspiration (FNA) in the clinical management of this subgroup of patients. METHODS: 1196 allograft recipients (522 liver, 288 cardiac, 250 renal, 131 lung, 5 heart and lung) were reviewed. A total of 62 (5.2%) (32 liver, 23 heart, 6 lung, and 1 renal) transplant patients underwent an FNA procedure. Thirty-seven males and 25 females were included, ranging in age from 18 to 71 years (mean 50 years). RESULTS: Of the 62 fine-needle aspirates, 29 (47%) were neoplastic. The most common malignancies aspirated were malignant solid tumors (15 cases)-including 8 epithelial malignancies, 5 hepatocellular carcinomas, and 2 mesenchymal neoplasms followed by posttransplant lymphoproliferative disorders (14 cases). Thirteen (21%) aspirates were inflammatory. The remaining 20 (32%) cases were benign aspirates from various sites (9 liver, 3 breast, 2 thyroid, 2 soft tissue, 2 lung, and 2 vertebral body). Surgical and/or autopsy material was available in 34 cases (55%). There was agreement between the tissue diagnosis and FNA material in 33 cases (97%). One case (3%) was a false negative. No false-positive cases were recorded. CONCLUSIONS: This study showed that over 50% of the aspirates were benign, justifying a conservative approach in the clinical management of these patients. Histologic correlation was available in 54% of the cases with an overall specificity of 100% and a sensitivity of 97%. We conclude that FNA is a highly sensitive and specific technique in the evaluation of lesions occurring in posttransplant patients. Cancer (Cancer Cytopathol) PMID- 10536355 TI - MOC-31 aids in the differentiation of metastatic adenocarcinoma from hepatocellular carcinoma. AB - BACKGROUND: Fine-needle aspiration biopsy (FNAB) is frequently used to diagnose mass lesions in the liver. Differentiating metastatic adenocarcinoma from primary hepatocellular carcinoma can be difficult. Despite a number of morphologic criteria, there remain occasional cases in which the cytologic features fail to resolve this differential reliably; in these cases ancillary studies may be useful. Recently, it has been reported that the antibody MOC-31 reliably separates metastatic adenocarcinoma from hepatocellular carcinoma. In this study we examine the utility of MOC-31 in liver FNAB material. METHODS: Thirty-three archival, alcohol-fixed, paraffin-embedded cell blocks representing 17 cases of hepatocellular carcinomas and 16 cases of metastatic adenocarcinoma were retreived. After protease digestion, the sections were immunostained with the antibody MOC-31 (Dako, Carpinteria, CA) utilizing a modified avidin-biotin complex technique. Only membrane-based reactivity was considered positive. RESULTS: In five cases there was insufficient diagnostic material remaining in the cell block for immunohistochemical staining. Among the remaining cases, MOC 31 reactivity was observed in 10 of 12 metastatic adenocarcinomas and 2 of 16 hepatocellular carcinomas. For metastatic adenocarcinoma the presence of MOC-31 reactivity yields a sensitivity of 83%, a specificity of 87%, a positive predictive value of 83%, and a negative predicitive value of 87%. CONCLUSIONS: MOC-31 is useful in separating metastatic adenocarcinoma from hepatocellular carcinoma in FNAB cell block material. Cancer (Cancer Cytopathol) PMID- 10536356 TI - Ultrasound-guided fine-needle aspiration biopsy of the thyroid. AB - BACKGROUND: We reviewed the Massachusetts General Hospital experience with ultrasound-guided fine-needle aspiration biopsies (FNABs) of the thyroid to determine the indications, rate of unsatisfactory smears, correlation with excisional biopsy results, and verification of efficient use of personnel time. METHODS: All radiologically guided FNABs of the thyroid from January 1993 through June 1997 were reviewed. As a measure of efficient use of technologist time, a sample of times spent by the technologist during the procedure for 20 cases in 1993 and 1997 was compared with that of an equal number of random nonthyroid image guided FNABs. RESULTS: Two hundred-ninety FNABs were identified in 251 patients, representing 12% of all thyroid FNABs and 11% of all radiologically guided FNABs. Indications in the 251 patients included multiple nodules (78), solitary nodules (61), complex nodules (39), prior failed FNAB (39), thyroid bed abnormalities post-thyroidectomy (21), difficult access (7), and investigation of recurrent tumor in residual thyroid lobe (6). Available records indicated 118 lesions were palpable and 45 were nonpalpable; the physical examination characteristics of the remainder (88) were not stated. Diagnoses included 44 unsatisfactory cases (15%), 103 macrofollicular lesions, 20 microfollicular lesions, 26 mixed macro/microfollicular lesions, 5 oxyphilic lesions, 1 trabecular pattern, 15 nonspecific follicular cell pattern, 9 follicular cell atypia, 30 cysts, 11 thyroiditis, 23 malignant tumors, and 3 other (1 parathyroid, 2 lymph node). Eighty-nine FNABs from 76 patients had subsequent surgical biopsy. Excisional biopsies in 14 unsatisfactory FNABs were benign. In the remaining 75 FNABs from 67 patients, 18 malignancies on FNAB were correctly diagnosed, but 3 other papillary carcinomas were only qualified as atypical follicular cells on cytology. No false-positive cases occurred. Of 15 macrofollicular lesions on cytology, 10 were adenomas on excision, only 2 of which were microfollicular adenomas, and 4 were adenomatous nodules. An aspirate of a parathyroid adenoma was misinterpreted as a macrofollicular lesion of the thyroid. Three microfollicular lesions on FNAB proved to be nodular hyperplasia on excision, and the other 11 were adenomas, 5 of them microfollicular. Average technologist time was significantly longer for thyroid FNABs than nonthyroid FNABs in 1993, but in the 1997 sample no significant difference was identified. CONCLUSIONS: Radiologically guided FNAB of the thyroid is a clinically useful procedure with a high correlation between benign lesions not needing excision (macrofollicular), and lesions that need excision (microfollicular/oxyphilic cell or malignant). Technologist time needed for immediate evaluation tends to decrease with increasing operator experience. Cancer (Cancer Cytopathol) PMID- 10536357 TI - Diagnostic p53 immunostaining of endobiliary brush cytology: preoperative cytology compared with the surgical specimen. AB - BACKGROUND: Endobiliary brush cytology is important in the distinction of malignant and benign causes of extrahepatic bile duct obstruction. The additional diagnostic value of p53 immunostaining on these cytology specimens was assessed. METHODS: All patients with extrahepatic bile duct obstruction who underwent endoscopic retrograde cholangiopancreatography (ERCP) with endobiliary brush cytology and subsequent surgery at the Academic Medical Center in Amsterdam during a 3-year period were studied. p53 Immunocytology was compared with the corresponding conventional light microscopic cytology and p53 immunostaining of the subsequent surgical specimen. RESULTS: Fifty-three patients with the following diagnoses were included: pancreatic carcinoma (23), bile duct carcinoma (15), ampullary carcinoma (5), lymph node metastases (2), carcinoma of unknown origin (4), chronic pancreatitis (3), and primary sclerosing cholangitis (1). Fifty-one percent of the carcinomas showed positive p53 immunostaining; all four surgical specimens without carcinoma were negative. The sensitivities of conventional light microscopic cytology, p53 immunocytology, and both tests combined were 29%, 24%, and 43%, respectively. These sensitivities were higher in cases of bile duct carcinoma (46%, 40%, and 66%) compared with cases of pancreatic carcinoma (13%, 9%, and 22%). Specificities of both tests were 100%. CONCLUSIONS: p53 Immunostaining on endobiliary brush cytology may be helpful in the diagnosis of malignant extrahepatic bile duct stenosis, especially in patients with bile duct carcinoma. Cancer (Cancer Cytopathol) PMID- 10536358 TI - Detection and quantitation by fluorescence in situ hybridization (FISH) and image analysis of HER-2/neu gene amplification in breast cancer fine-needle samples. AB - BACKGROUND: Fine-needle sampling, although a practical and noninvasive method of tissue acquisition, has rarely been used for HER-2/neu fluorescent in situ hybridization (FISH). To assess HER-2/neu gene amplification in mammary carcinoma, FISH signals on cytology and corresponding tissue biopsies were detected visually and measured by image analysis. The results were correlated with patient and tumor characteristics. METHODS: In situ HER-2/neu DNA probe hybridization was performed on 61 cytology specimens and on 47 corresponding frozen sections of breast carcinomas. Tumors were classified by visual evaluation as unamplified, moderately amplified, or highly amplified. Multiparametric image analysis was performed using the Discovery automated image analyzer (Becton Dickinson, Leiden, Netherlands). The integrated fluorescence ratio (IFR) was calculated for each sample as the integrated FISH fluorescence of the tumor cells divided by the integrated FISH fluorescence of internal control cells containing two spots. The percentage positive nuclear area (PPN), calculated as the area of FISH fluorescence divided by the area of nuclear DNA fluorescence, and the PPR, ratio of the PPN of the tumor cells divided by the control cells, were also calculated for each sample. RESULTS: Visual analysis yielded 46 unamplified and 15 (24.6%) amplified (seven moderately amplified and eight highly-amplified) tumors. Strong (P < 0.001) correlation between results on cytological and histological materials was obtained. The FISH spots on the cytological preparations were more easily visualized and scored than those on the corresponding tissue sections. Visual HER-2/neu signal scoring was strongly correlated with IFR (P = 0.0001) and PPR (P = 0.0001). Within the tumors classified as highly amplified by visual examination, quantitation of the degree of amplification fluorescence signal was possible using image analysis. CONCLUSIONS: Cytologic specimens were a suitable and representative source of materials for detection and quantitation of HER-2/neu gene amplification by FISH and image analysis. Cancer (Cancer Cytopathol) PMID- 10536359 TI - Regulatory functions of Cdk9 and of cyclin T1 in HIV tat transactivation pathway gene expression. AB - HIV-1 gene expression relies upon a complex machinery that is primarily controlled by two viral regulatory proteins, Tat and Rev. Rev is involved in regulating post-transcriptional events of HIV-1 gene expression. The Tat protein transactivates transcription from the HIV-1 5' long terminal repeat (LTR) and acts in synergy with specific cellular factors. Recently, it has been shown that one set of these cellular factors is a protein kinase activity termed TAK (Tat associated kinase), which activates transcription by hyperphosphorylation of the carboxyl-terminal domain (CTD) of the large subunit of RNA polymerase II. TAK also enhances transcription of HIV-2, together with the retroviral transactivator, Tat-2. The TAK activity appears to be related to the CTD kinase P TEFb, which stabilizes transcription elongation of many genes and was originally isolated from Drosophila extracts. Both TAK and P-TEFb contain at least two subunits: the cyclin-dependent kinase, CDK9 (PITALRE), the catalytic subunit, and the regulatory subunit, cyclin T1. CDK9 and cyclin T1 are ubiquitous factors that affects many cellular processes, including cell differentiation and apoptosis. The involvement of TAK in HIV-1 and HIV-2 gene expression is an important aspect in the biology of these two retroviruses, and may lead to the development of novel antiretroviral drugs and/or gene therapy approaches for the treatment of patients with AIDS. PMID- 10536360 TI - Electromagnetic fields may act directly on DNA. AB - A wide variety of environmental stimuli induce the expression of stress response genes, including high temperatures, hypoxia, heavy metal ions, and amino acid analogs. Stress genes are also induced by low frequency magnetic fields. The cellular response to magnetic fields is activated by unusually weak stimuli, and involves pathways only partially associated with heat shock stress. Since magnetic fields interact with moving charges, as we have shown in enzymes, it is possible that magnetic fields stimulate the stress response by interacting directly with moving electrons in DNA. In this paper, we review several lines of evidence that support this hypothesis. PMID- 10536361 TI - Modulation of glutathione transferase P1-1 activity by retinoic acid in neuroblastoma cells. AB - The ability of retinoic acid to modulate glutathione S-transferase P1-1 (GSTP1-1) activity has important implications both for cancer prevention and for anticancer therapy. We investigated GSTP1-1 expression and activity in the human neuroblastoma cell line SK-N-BE(2) (genotype A*/B*) under basal conditions and during 48-h incubation with 0.1 microM all-trans-retinoic acid. The steady-state levels of glutathione transferase P1-1 mRNA and protein during 48-h incubation with all-trans-retinoic acid did not increase substantially, but we detected a significant reduction of GSTP1-1 specific activity. This reduction in enzymatic activity could not be ascribed to a differential action of retinoic acid on the gene variants A* and B*; indeed, the two GSTP1-1 isoforms have different affinities toward 1-chloro-2,4-dinitrobenzene (CDNB), while we found a substantial invariance of the K(m) (CDNB) in the cytosol during retinoid treatment. A modulatory effect of retinoic acid on other enzymes involved in glutathione transferase P1-1 metabolism, such as the retinoic acid-induced tissue trans-glutaminase, might be hypothesized, as well as a direct inactivation of GSTP1-1 by the oxidative stress that characterizes the early phases of apoptosis. PMID- 10536363 TI - Coiled bodies are predisposed to a spatial association with genes that contain snoRNA sequences in their introns. AB - Small nucleolar RNAs (snoRNAs) constitute a group of more than 100 different stable RNA molecules that are found concentrated in the nucleolus where they are involved in the maturation of ribosomal RNA. Most snoRNAs are not produced from their own genes but are encoded in the introns of other genes, referred to as snoRNA host genes. Little is known about the mechanisms by which the snoRNAs are produced from introns and how the snoRNAs mature to functional snoRNP complexes in the nucleolus. One class of intron-encoded snoRNAs binds with high specificity to the protein fibrillarin which is found concentrated in the nucleolus, but also in small nuclear domains known as coiled bodies. It has become clear that genes that code for small stable RNAs, e.g., U1, U2 snRNA, and the U3 snoRNA, are often found adjacent to coiled bodies. High concentrations of transcription factors and RNA processing factors in and around coiled bodies indicate that they may be involved in the expression of the adjacent genes. In order to investigate whether coiled bodies could play a role in the synthesis of intron-encoded snoRNAs the distribution of coiled bodies was studied relative to three different snoRNA host genes, i.e., hsc70, RPS3, UHG. All three were found adjacent to coiled bodies at significantly high frequencies (11-19%), compared to control sequences (0-2%), to conclude a preferential association between the snoRNA host genes and coiled bodies. This association could point to a possible role for coiled bodies in the synthesis and/or maturation of snoRNAs. An involvement in snoRNA production could explain the presence of transcription factors, splicing factors, and fibrillarin in coiled bodies. PMID- 10536362 TI - Human bone marrow osteoprogenitors express estrogen receptor-alpha and bone morphogenetic proteins 2 and 4 mRNA during osteoblastic differentiation. AB - Understanding the mechanisms that control the proliferation and commitment of human stem cells into cells of the osteogenic lineage for the preservation of skeletal structure is of basic importance in bone physiology. This study examines some aspects of the differentiation in vitro of human bone marrow fibroblastic cells cultured in the absence (basal media) or presence of 1nM dexamethasone and 50 micrograms/ml ascorbate for 6, 10, 14, and 21 days. Northern blot analysis and in situ hybridisation with digoxygenin-labelled riboprobes for Type I collagen, osteocalcin, bone morphogenetic proteins 2 (BMP-2), and 4 (BMP-4) and the estrogen receptor alpha (ERalpha), together with immunocytochemical analysis of ERalpha expression and histochemical staining of alkaline phosphatase was performed. In basal media, alkaline phosphatase activity and collagen expressions were detected at day 6, ERalpha from day 10 and osteocalcin from day 10. In the presence of dexamethasone and ascorbate, cell proliferation and alkaline phosphatase were markedly stimulated over 10 to 14 days with a dramatic increase in the temporal expression of Type I collagen, ERalpha, and osteocalcin mRNAs in these cultures. Northern blot analysis showed cells cultured in basal media, expressed the highest levels of the mRNA for each marker protein at day 14, whereas in the presence of ascorbate and dexamethasone, the highest levels for alkaline phosphatase, ERalpha, osteocalcin, BMP-2, and BMP-4 were observed at day 21. ERalpha, BMP-2, and BMP-4 expression were found to correlate temporally with induction of the osteoblast phenotype as determined by alkaline phosphatase, collagen, and osteocalcin expression. These results give additional information on the development of the osteoblast phenotype from early fibroblastic stem cells and on the biological factors involved in this process. These studies suggest a role for estrogen and BMP-2 and -4 in the differentiation of osteoprogenitor cells. PMID- 10536364 TI - CREB and COUP-TF mediate transcriptional activation of the human immunodeficiency virus type 1 genome in Jurkat T cells in response to cyclic AMP and dopamine. AB - Infection of lymphocytes by the human immunodeficiency virus type 1 (HIV-1) is associated with an increase in intracellular cAMP levels. Recent studies have shown that lymphocytes are able to synthesize and bind the dopamine, known to affect multiple cellular pathways, such as the cAMP pathway. Here we have investigated the molecular mechanisms by which cAMP and dopamine regulate HIV-1 gene transcription in Jurkat T cells. Transient expression experiments revealed that dopamine and forskolin lead to a synergistic stimulation of long terminal repeat (LTR)-driven transcription. This action is mediated through the cAMP response element binding (CREB) protein and chicken ovalbumin upstream promoter transcription factor (COUP-TF). CREB and COUP-TF act indirectly through the minimal -40/+80 and -68/+80 LTR region, respectively. We have previously demonstrated that COUP-TF stimulates HIV-1 transcription via the -68/+29 LTR region without direct DNA binding. Here, gel supershift experiments show that CREB does not directly bind to the -45/+85 proximal LTR sequences. Moreover, our data reveal novel functional interactions between COUP-TF and CREB, which lead to synergistic cAMP- and dopamine-induced transactivation of the HIV-1 LTR. These findings reveal that dopamine-induced signals and the cAMP pathway stimulate HIV 1 gene transcription in lymphocytes by converging to the minimal -68/+80 LTR region, through the transcription factors CREB and COUP-TF. PMID- 10536365 TI - Abnormal osteogenesis in osteoporotic patients is reflected by altered mesenchymal stem cells dynamics. AB - Bone marrow contains a population of mesenchymal stem cells with the ability to differentiate into cells that form bone, cartilage, adipose, and other connective tissues. Stem cells can be isolated from bone marrow aspirates and expanded in vitro. Presently, most stem cells studies have been performed in cells obtained from "healthy" control subjects. The goal of this study was to compare the functional characteristics of mesenchymal stem cells derived from "healthy" control and osteoporotic postmenopausal women to better understand the mechanisms involved in the pathogenesis of this disease. Osteoporotic and control stem cells have similar morphology and size and express similar cell surface antigens as evidenced by their reactivity with cell specific monoclonal antibodies. Mesenchymal stem cells from osteoporotic women differ from controls in having a lower growth rate than control cells, being refractory to the mitogenic effect of IGF-1, and exhibiting a deficient ability to differentiate into the osteogenic linage as evidenced by the alkaline phosphatase activity and calcium phosphate deposition. We conclude that in osteoporosis stem cell growth, proliferative response and osteogenic differentiation are significantly affected. Also, the study of mesenchymal stem cells from osteoporotic postmenopausal women may provide a better understanding of the mechanisms involved in the pathogenesis of the osteoporosis. It may also serve to test in vitro in rapid manner novel new therapeutic strategies. PMID- 10536366 TI - Donor variation in the growth properties and osteogenic potential of human marrow stromal cells. AB - Human marrow stromal cells (MSCs) were isolated from posterior illiac crest marrow aspirates obtained from 17 healthy donors, ages 19-45 years, with no apparent physical disability. First passage hMSCs exhibited growth rates in vitro that varied up to 12-fold between donors. No correlation between growth rate and the age or gender of the donor was evident (P 1000 pregnancies. Weekly measurements of the cervical canal by transvaginal ultrasound probe. RESULTS: Inverse relationship between sonographic measurements and advancing gestational age. At 38 weeks of gestation, significant difference in the ultrasonic recordings between patients delivered before and beyond 41 completed weeks. CONCLUSIONS: It is possible to identify as early as 38 weeks of gestation those women that will deliver beyond 41 weeks. There is a subgroup of nulliparous patients that will probably be candidates to induction of labour. PMID- 10536422 TI - [Total laparoscopic hysterectomy. Cost analysis]. AB - BACKGROUND: Hysterectomy performed totally by laparoscopy is still one of the most advanced procedure in endoscopic gynecological surgery. The aim of this study has been to evaluate the cost and the hospital charges of total laparoscopic hysterectomy as compared with those for total abdominal and vaginal hysterectomy. METHODS: In particular, as far as laparoscopic technique is concerned, the cost of linear staplers have been analyzed as compared to disposable and non-disposable instruments for bipolar coagulation. RESULTS: After analysing the data, it has been found that the use of linear staplers involves very high median costs of Lire 2,932,304 ($ 1,650) versus the use of non disposable instruments of Lire 936,488 ($ 526). The median total hospital charges was of Lire 6,014,448 ($ 3,380) for abdominal hysterectomy, of Lire 4,449,617 ($ 2,500) for vaginal hysterectomy and of Lire 4,078,000 ($ 2,291) for laparoscopic technique with non-disposable supplies. CONCLUSIONS: The results confirm, on the one hand, that bipolar coagulation is a reliable method of hemostasis, and on the other hand that laparoscopic surgery is a technique that can be recommended as an alternative to laparotomy for hysterectomy with unquestionable benefits both for patients and hospital. PMID- 10536423 TI - [Congenital diaphragmatic hernia. Prenatal diagnosis and neonatal outcome]. AB - BACKGROUND: The objective of this study is to determine the management and the outcome of 17 cases of congenital diaphragmatic hernia observed, and to compare the results with the literature data. METHODS: The study was made between June 1994 and June 1998 in the Department of Obstetrics and Gynecology of the University of Naples Federico II and it collected 17 pregnant women with diagnosed or suspected congenital diaphragmatic hernia, referred to our diagnostic unit from other institutes: between 18 and 37 weeks' gestation, a detailed ultrasound examination and an echocardiography were performed to confirm the diagnosis, to establish the site and contents of the hernia exactly and to detect associated structural malformations. Fetal kariotyping was made in 7 cases. When the women decided to continue pregnancy, or the legal limit for termination of pregnancy was exceeded, the pregnancy was monitored with ultrasound examination, delivery took place in our department and the baby was transferred to an intensive care unit. RESULTS: Ultrasound examination led to the diagnosis of 14 postero-lateral left-sided diaphragmatic hernias (82.3%), 1 antero-lateral left-sided (5.9%), 1 bilateral postero-lateral sided (5.9%) and 1 diaphragmatic eventration (5.9%). Associated structural malformation were diagnosed in 4 fetuses (27%). In 1 case only (14.3% of examinated kariotyping) an abnormal result was found. Five pregnancies (29.4%) were terminated, 2 (11.7%) are still going on and 10 fetuses (58.9%) were born alive: 5 fetuses (50%) died in the first days of life before surgical intervention, 2 (40% of operated children and 20% of born alive) died after the operation and 3 (60% of operated children and 30% of born alive) are actually alive and in good health. The total postnatal mortality was 70% and 40% after operation. CONCLUSIONS: From the analysis of these data and from the international literature the conclusion is drawn that congenital diaphragmatic hernia is associated with a high postnatal mortality, although potentially it can be corrected with surgery: a better postnatal management and a better knowledge of evaluable in uterus prognostic factors are necessary to improve postnatal outcome. PMID- 10536424 TI - [Automation of cytological analysis of cervical smears]. AB - Quality Control (QC) is essential in cytologic laboratories. To reduce or avoid false negatives due to screening or interpretation errors, all cervical smears may be analyzed twice by two different cytopathologists (CP). During rescreening or Quality Control (QC), the second CP may be assisted by an automated device. Today there are different instruments: computerized microscope Ac Cell Series 2000 Pathfinder System), semiautomated or interactive systems (PAPNET, CytoRich or AutoCyte, ACCESS), automated systems (AutoPap 300). These devices, except computerized microscopes, utilize algorithmic image analysis that values single cells using morphological features but has difficulty in analyzing clusters of overlapping cells. For this reason it is better to use thin or monolayer preparations. This approach results in the loss of background and cells useful for the diagnosis and modifies sampling and processing methods. However, the techniques to obtain thin or monolayer preparations may process a higher number of cells than conventional method; moreover, the samples obtained may be valued also by the optic microscope, making cytologic analysis easier. The only device that combines the use of algorithmic image analysis with neural networks is the PAPNET system. Neural networks was inspired by neurobiology and may identify different cellular morphology and overlapping cells. In our laboratory, the PAPNET was proposed in 1995. In the present study, the last rescreening results of 1958 negative cervical smears are reported, analyzed during primary screening from July 1997 to February 1998. During the QC assisted by the PAPNET, 6 false negatives (0.31%), due to cytologic errors in the primary screening were detected. These results confirm the usefulness and the effectiveness of QC assisted by automated devices. However, only the CP evaluates abnormal cells detected by semiautomated systems or analyzes more atypical smears identified by the instruments. The work of CP is difficult: therefore a strict collaboration between clinician and CP to formulate the correct diagnosis is essential. PMID- 10536425 TI - Congenital heart disease in pregnancy. AB - The clinical cases of three patients affected respectively by Eisenmerger's syndrome, Marfan syndrome, coarctation of the aorta are described. All patients belonged to NYHA class I or II. During pregnancy contact with cardiologists, anaesthetists, neonatologists was maintained and this allowed accurate management. Both pregnancy and delivery evolved without any complication and with a positive outcome for mother and newborn. PMID- 10536426 TI - [Proposal for an informed consent form for hormonal replacement therapy in ambulatory care]. AB - Hormonal Replacement Therapy (HRT) represents a valid therapeutic approach for menopausal symptoms and for the prevention of cardiovascular disease and osteoporosis. Nevertheless, an informed consent, after a complete information, must be obtained from the patient. This procedure is generally adopted in any medical activity, but the modality of the consent in the HRT administration is not well established (verbal or written?, timing of administration?). The authors propose an informed written consent model to be utilized in menopausal centers; this model synthetically informs about HRT benefits and risks and must be red and signed by the patient. The written consent should be explained through a verbal detailed discussion about it, during which the patient's comprehension must be assured. The informed consent procedure should be renewed every year in long term users. The influence of the HRT informed consent in menopausal centers must be analyzed in particular as far as women compliance is concerned. PMID- 10536427 TI - [Problems of nosologic classification of polycystic ovary syndrome and metabolic syndrome X]. PMID- 10536428 TI - The past, present and future of antimycotic combination therapy. PMID- 10536429 TI - Molecular epidemiology of Candida albicans isolates from AIDS and cancer patients using a novel standardized CARE-2 DNA fingerprinting technique. AB - A total of 277 Candida isolates from various body sites of 149 AIDS and cancer patients treated in four different university clinics in Wurzburg, Germany were collected over a period of 27 months and phenotypically and genotypically characterized. The fingerprinting patterns of 194 Candida albicans isolates obtained with the moderately repetitive, C. albicans-specific DNA fragment CARE-2 were digitized and retrospectively compared with a highly accurate computer assisted standardization method. A total of 168 different genotypic patterns (< 100% identity) could be differentiated using this technique. Although comparative analysis of C. albicans subsets revealed a pronounced tendency of C. albicans isolates from HIV patients to form clusters, the mean genetic variability in HIV and cancer patient isolates was virtually identical. Patients with a specific disease condition or in a certain age group were not found to harbour C. albicans isolates displaying a characteristic "signature genotype". Micro-evolutionary changes were detected by CARE-2 fingerprinting in temporal successive isolates of one patient, but nosocomial transmission of identical isolates between unrelated patients was never seen. Genotyping showed that patient isolates can replace one another; occasionally also species switches were observed. Secreted aspartic protease (SAP) production was not correlated with a specific C. albicans banding pattern; isolates obtained from HIV patients and from an internal control group secreted comparable amounts of SAP. Candida dubliniensis isolates in this study showed an elevated level of SAP production. When used under standardized conditions, CARE-2 fingerprinting is an efficient, reproducible and sensitive technique to characterize C. albicans isolates. PMID- 10536430 TI - Genotypic relatedness of yeast isolates from women infected with human immunodeficiency virus and their children. AB - The objective of this study was to compare polymerase chain reaction (PCR) fingerprinting with other molecular typing methods as an epidemiologic tool to investigate the transmission of Candida strains between HIV-positive mothers and their children. Forty-nine yeast strains (including Candida albicans, Candida glabrata, Rhodotorula rubra, Candida tropicalis, Candida famata, Candida dubliniensis, Saccharomyces cerevisiae) from 30 individuals (15 children and 15 HIV-infected mothers or accompanying person) were isolated. Colonization/infection with yeast was observed in 80% of all individuals in the oral cavity, and in 33% from hand cultures, respectively. Thirteen out of 15 children (86%) and 12 out of 15 adults (80%) were colonized/infected with yeasts. Candida dubliniensis strains were found in four HIV-infected women but not in children. The results with an arbitrarily primed (AP)-PCR mediated genotyping assay using phage M13 core sequence were compared with the hybridization patterns using the species-specific DNA probe CARE-2 for the C. albicans isolates. Typing of non-C. albicans strains was done using AP-PCR in comparison with pulsed-field gel electrophoresis (PFGE). Twenty-six C. albicans strains gave two different genotypes by AP-PCR but 16 genotypes by CARE-2 hybridization. The CARE-2 probe appeared to have a higher discriminatory power compared with the primer 'M13' mediated AP-PCR in typing C. albicans isolates. PMID- 10536432 TI - Early investigation and initiation of therapy for invasive pulmonary aspergillosis in leukaemic and bone marrow transplant patients. AB - Invasive fungal infections are an increasingly common problem in cancer patients and in other vulnerable groups. Invasive pulmonary aspergillosis (IPA) in the neutropenic host presents particular challenges in terms of diagnosis and therapy. Against the background of a recognized problem of invasive aspergillosis in haematology/oncology patients treated at the Christie Hospital, we undertook a prospective study in patients at risk for IPA. The aim of the study was to improve outcome by using the linked strategies of first, early diagnosis, and secondly, early aggressive therapy with a lipid-associated formulation of amphotericin B, amphotericin B colloidal dispersion ('Amphocil'). Early investigation comprised the use of high-resolution computerized tomography scanning of the thorax and fibreoptic bronchoscopy to obtain bronchoalveolar lavage specimens, processed using conventional detection and culture methods. Using this approach, the incidence of proven or probable IPA in patients with acute leukaemia was 9%. Prompt initiation of amphotericin B colloidal dispersion therapy led to a successful outcome in 11 of 13 patients, compared with a mortality of 100% in historical controls. PMID- 10536431 TI - Fungal surveillance cultures during antifungal prophylaxis with itraconazole in neutropenic patients with acute leukaemia. AB - Fungal colonization has been associated with an increased rate of invasive fungal infections in neutropenic patients. This study evaluates weekly fungal surveillance cultures from the oropharyngeal and perianal space as well as other suspected sites in 219 courses of myelosuppressive chemotherapy with itraconazole antifungal prophylaxis in 116 neutropenic patients with acute leukaemia. Itraconazole was given from the start of chemotherapy in one of six different dosing regimens. Fungal colonization occurred in 68 (31%) of courses, which was lower than in a historical control group without prophylaxis (53%, P = 0.004). Twenty-six per cent of these 116 isolates had a growth rate of more than 50 colony forming units (CFU) per culture. Candida glabrata (51%), Candida albicans (18%) and Candida krusei (4%) were the most frequently isolated species. Higher median itraconazole trough concentrations were associated with a lower growth rate in the cultures (< or = 50 CFU/culture versus > 50 CFU/culture): 710 (430 1180) ng ml-1 versus 900 (560-1650) ng ml-1 (P = 0.015). The use of itraconazole solution--compared with capsules--led to a reduced growth rate (P = 0.035). In conclusion, compared with historical controls itraconazole antifungal prophylaxis reduces the incidence and the extent of fungal colonization during neutropenia in patients with acute leukaemia. PMID- 10536433 TI - Characterization of Malassezia species by means of phenotypic characteristics and detection of electrophoretic karyotypes by pulsed-field gel electrophoresis (PFGE). AB - A total of 220 isolates of the genus Malassezia obtained from affected skin of animals and humans were included in this study. The isolates were examined for their belonging to the known species Malassezia pachydermatis, Malassezia sympodialis, Malassezia furfur, Malassezia obtusa, Malassezia restricta, Malassezia globosa and Malassezia slooffiae by means of phenotypic and biochemical characteristics as well as their ability to assimilate polyethylen(20)-sorbitan-esters (Tween 20, 40, 60, 80). The electrophoretic karyotypes of all isolates were studied by pulsed-field gel electrophoresis. Of the 220 Malassezia isolates examined, 217 were identified as M. pachydermatis and three were classified as M. sympodialis. Moreover, the proof of limited lipid dependence of M. pachydermatis indicated that the commonly used criterion 'lipid independence' is not sufficient to differentiate this species from the other known Malassezia species. PMID- 10536435 TI - Case reports. Four paediatric cases of tinea capitis due to unusual agents. PMID- 10536434 TI - Efficacy and tolerability of terbinafine 1% topical solution used for 1 week compared with 4 weeks clotrimazole 1% topical solution in the treatment of interdigital tinea pedis: a randomized, double-blind, multi-centre, 8-week clinical trial. AB - In this double-blind clinical trial 429 patients (217 terbinafine and 212 clotrimazole) were randomized to receive twice daily terbinafine 1% topical solution for 1 week followed by a vehicle application for 3 weeks, or 1% clotrimazole solution for 4 weeks. Patients were evaluated clinically and mycologically at baseline and then at weeks one, two, four (end of treatment), and eight (end of follow-up). To be evaluable the patient needed to have a positive culture for a dermatophyte and positive KOH microscopy and a clinical diagnosis of tinea pedis (interdigital type) at baseline. Effective treatment of tinea pedis was recorded in 181 of 217 (83%) of patients treated for 1 week with terbinafine 1% solution and 174 of 212 (82%) of patients treated for 4 weeks with clotrimazole 1% solution. Mycological cure and disappearance of signs and symptoms were similar at each assessment visit in the two groups. In the subgroup of patients without any protocol violation the mycological cure rate was 95% (164 of 173) with terbinafine solution and 91% (159 of 174) with clotrimazole solution (P = 0.05). Adverse events believed to be drug-related occurred in 13 patients in the terbinafine group and 11 in the clotrimazole group (4 to 5% in each group). The events were primarily local skin reactions of mild to moderate intensity. It can be concluded that terbinafine 1% solution used for 1 week to treat tinea pedis is well tolerated and at least as effective as clotrimazole 1% solution used for 4 weeks. PMID- 10536436 TI - ASM--conference 'Candida and Candidiasis'. March 1-4, 1999. Charleston SC, USA. PMID- 10536437 TI - [Proteome analysis: the state of the art of the methodology]. AB - The proteome is the protein complement of a genome. Proteome analysis has been progressing worldwide. Two-dimensional electrophoresis (2-DE), a key technique in proteome analysis, separates proteins on a polyacrylamide gel according to the isoelectric point and molecular mass. A total of 1,000-1,500 protein spots can be separated and detected on a polyacrylamide gel using silver-staining. It is important to identify individual protein spots in order to correlate the information of the genome to that of the corresponding proteome. By automatic amino-terminal sequencing, around 15 amino acid residues from the amino terminus can be determined from 1 pmole of a protein sample. The homology search of the obtained sequences against a protein sequence database can clarify the protein unambigously. Recently, a carboxyl-terminal sequencing method using a vapor from a high concentration of an organic acid has been developed. Peptide-mass fingerprinting is a new method for protein identification using residue-specific proteases and mass spectrometry. Two types of chemical cleavage methods, the carboxyl side cleavage of the aspartyl peptide bond and the amino terminal cleavage of serine/threonine peptide bonds would be more suitable for peptide mass-fingerprinting of micro amount protein because of no contamination from the gel matrix or the enzyme used. It would be possible to analyse less amount protein sample (100 femtomole) more rapidly according to advancement of mass spectrometry. PMID- 10536438 TI - [Polymorphism of hypervariable region in D-loop of mitochondrial DNA]. AB - DNA sequences of PCR products from mitochondrial DNA (mtDNA) of 80 healthy Japanese volunteers (40 pairs, mother and child) were determined by the direct sequencing method for polymorphism. Thirty (15 pairs) of 80 samples analyzed showed a T-to-C transition at position 16189 (T16189C) of the C-stretch region in the hyper-variable region of mtDNA. For seven pairs randomly selected from the 15 T16189C pairs (C-stretch) and a single pair without the transition (non C stretch), PCR products from the D-loop region were cloned and then sequenced. The repeat number of C in the C-stretch region was found to show heteroplasmy by sequencing multiples clones from each mtDNA. Statistical analyses of the distribution patterns of the repeat number revealed no significant differences between the mother and child in each lineage but significant differences between the lineages. The seven lineages could be then classified into four groups. The result of our data confirmed the existence of heteroplasmic polymorphism in the C stretch region and the inheritance of the heteroplasmy from mother to child. Therefore, the analysis of heteroplasmy is applicable to individual identification. PMID- 10536439 TI - [Histological study of early postmortem changes in various organs: comparison of the paraffin embedding method and the epoxy resin embedding method]. AB - For the purpose of morphological assessment of the early postmortem interval, Wistar rats were killed by cervical dislocation and left at 23 degrees C for 1, 3, 5, 10, 15 and 24 hours. After a given postmortem interval, tissue samples taken from kidney, pancreas, liver, heart and skeletal muscle were embedded in paraffin or epoxy resin and examined by light microscopy. Specimens obtained from the paraffin block did not show a good correlation between histological changes and postmortem interval, because the postmortem changes continued during the fixation period. On the other hand, the time course of histological changes in specimens obtained from the epoxy block, particularly the development of clumping of nuclear chromatin and cytoplasmic vacuolation in each organ, reflected the postmortem interval because of the rapid fixation by glutaraldehyde. These histological changes were characteristic of each organ up to 24 hours after death. In addition, the semithin epoxy resin section made high-resolution light microscopy possible. Therefore, the epoxy resin embedding method is superior to the paraffin embedding method for the purpose of estimation of the time of death. The morphological changes characterising time after death are as follows: at 1 hour after death, cytoplasmic vacuolization and slight clumping of nuclear chromatin in pancreatic acinar cells; at 3 hours after death, slight clumping of nuclear chromatin in distal tubules, cytoplasmic vacuolization in skeletal muscle, and edema in cardiac muscle; at 5 hours after death, clumping of nuclear chromatin in proximal tubules as well as distal tubules, and cytoplasmic vacuolization in hepatocytes; at 10 hours after death, edema in proximal tubules, condensation of nuclear chromatin (apoptosis) and edema in distal tubules, and atrophy of acinar cells in the pancreas; at 15 hours after death, cytolysis of distal tubules; at 24 hours after death, cytolysis of hepatocytes and clumping of chromatin in skeletal muscle. Thus we can conclude that the time course of histological changes is useful for the estimation of postmortem interval. PMID- 10536440 TI - [Tests for Hardy-Weinberg equilibrium in Japanese population]. AB - In population genetics, the absence of the departure from Hardy-Weinberg equilibrium (HWE) is usually tested when a population study of a certain DNA marker is performed to show the observed allele frequencies represent those of the whole population. The goodness-of-fit test (chi 2 test) assuming chi 2 approximation has frequently been used with classical blood type markers having a few alleles. However, new tests suitable for DNA markers having many alleles, such as homozygosity test, likelihood ratio test and Guo-Thompson's (G-T') exact test, have recently been devised. In the present study, appropriate tests for HWE was studied using population data of 206 Japanese individuals with 9 different short tandem repeat loci. Firstly, we found that the recommendation of NRC II for the treatment of rare allele frequencies (If a bin in the database contains fewer than five entries, it is pooled with adjacent bins so that no bin has fewer than five) is quite reasonable for personal identification in forensic sciences. Secondly, we proposed that homozygosity test, likelihood ratio test and G-T's exact test should be applied altogether and HWE of the sample population should be valid only when all of the three tests were cleared. PMID- 10536441 TI - [The structural polymorphism in D8S580-STR region]. AB - The STR structure of the D8S580 locus was analyzed by the base sequencing technique. Alleles were collected separately by urea denaturing polyacrylamide gel electrophoresis of the PCR amplification product, followed by cutting of the target DNA band from the gel, and reamplification. As a result, this locus was shown to have a complex STR structure consisting of four types of repeat units, i.e. GGAA (II), GAAA (III), GAAAA (IV), and GCAA (V). Of 53 unrelated Japanese individuals, 51 were heterozygous, and the most frequent allele was type IV 8-II 7 III 6 III 3 (16%). Forty-four alleles with distinct structures appeared once the samples tested. As 57 different alleles were detected in this study, this region is considered to be one of the locus that have a high degree of repetitive structural polymorphism and therefore useful in forensic science. PMID- 10536442 TI - [A sudden death case in the bath with peculiar scald]. AB - A 60-year-old man was found dead in the bathtub of his house on the 2nd of February. Erythema with a clear border was observed on the right side of the face, trunk and around the knees, though the left side of the face, the hips and the feet were normal. The water heater was set outside of the bath, and the heated water flowed from the heater to the bath through the upper pipe. In order to determine the mechanism of the peculiar scalding, changes in the water temperature in the bathtub were measured under the same conditions. An hour after turning on the water heater, the surface water temperature of the bath was 73 degrees C. Zonal gradation of the temperature was observed. At the level of the opening for the lower intake pipe, it was 50 degrees C. The border of the scalding was consistent with the water layer above 50 degrees C. PMID- 10536443 TI - [MR findings of intramedullary tumors]. AB - Intramedullary tumors are relatively rare. In this review article, we describe the characteristic MR findings and differential diagnosis of three common intramedullary tumors: astrocytoma, ependymoma and hemangioblastoma. It is important preoperatively to differentiate ependymoma from astrocytoma, because ependymoma has a clear tumor margin, and therefore complete removal of the tumor can be achieved. Other intramedullary lesions that need to be differentiated from intramedullary tumors are also described. PMID- 10536444 TI - [CT diagnosis of the gastrointestinal tract]. AB - Five topics regarding CT diagnosis of the gastrointestinal tract are discussed with illustrative cases: 1) Fat-containing tumors can be seen in the esophageal lumen, and their differential diagnosis includes not only lipoma and liposarcoma (sometimes teratoma) but also fibrovascular polyp and carcinosarcoma. 2) The multi-layered pattern of the gastric wall observed on dynamic CT is important for the differential diagnosis of gastric lesions and the staging of gastric cancer. 3) Swelling of the sentinel lymph nodes is an important finding for correct staging of cancer, incidental detection of occult cancer, and diagnosis of the origin of abdominal tumors. 4) Air-inflated CT and virtual CT endoscopy of the colon are necessary for the detection of small colorectal cancers, and multi planar reconstruction (MPR) images vertical to the lesions are helpful for cancer staging. 5) MRI and CT with MPR images are useful for the evaluation of anorectal fistula. PMID- 10536445 TI - [MR angiography of steno-occlusive lesions of intracranial arteries: a comparative study between turbo MRA and conventional MRA]. AB - PURPOSE: To compare the quality and diagnostic accuracy of images of intracranial steno-occlusive lesions obtained by conventional MRA and turbo MRA reconstructed using the zero-filled interpolation technique in the slice-select direction. MATERIALS AND METHODS: Eighteen patients with suspected steno-occlusive lesions of the intracranial arteries were studied with two types of three-dimensional time-of-flight angiography and conventional digital subtraction angiography. In total, 45 steno-occlusive lesions were quantitatively measured using calipers and correlated with DSA stenosis. A phantom that simulated vessels with stenosis was also imaged using the two types of MRA under the same conditions as those employed in the clinical study. RESULTS: Compared with conventional MRA, turbo MRA reduced the jaggedness of vessels and offered appearances more similar to those of DSA in the antero-posterior and lateral views. The severity of stenosis was classified into five grades based on the percentage of occlusion: not significant (0-24%), mild (25-49%), moderate (50-74%), severe (75-99%), and occlusive (100%). Neither turbo MRA nor conventional MRA showed any discrepancy from DSA above grade-1 stenosis. CONCLUSION: The advantage of turbo MRA is its ability to reduce the jaggedness of vessels on conventional MRA, and to simplify the recognition of vessel contours without prolonging acquisition time. Turbo MRA and conventional MRA have equally high diagnostic accuracy for steno-occlusive lesions. PMID- 10536446 TI - [CT findings of non-traumatic colorectal perforation]. AB - We retrospectively analyzed CT scans from 23 patients with non-traumatic colorectal perforation. We compared the sensitivity of CT and plain film radiography in the detection of free gas. Free gas was observed in 7 of the 23 cases (30.4%) on plain film radiography and 16 of the 23 cases (69.6%) on CT. Retroperitoneal abscess was demonstrated in 6 of the 7 patients without free gas on CT. Extraluminal air and abscess covered by the omentum and mesenterium were demonstrated in the remaining one patient. The site of perforation was identified in 19 of the 23 patients (82.6%) on CT. CT was useful for demonstrating retroperitoneal free gas, changes in mesenteric fat, extraluminal feces, and tumors. We conclude that CT is indicated in cases of suspected colorectal perforation. PMID- 10536447 TI - [Factors influencing treatment results of definitive radiotherapy following transurethral surgery for muscle-invasive bladder cancer]. AB - PURPOSE: To determine the prognostic factors influencing the outcome of bladder cancer patients treated with definitive radiotherapy following transurethral tumor resection (TURBT). MATERIALS AND METHODS: From March 1977 through August 1991, 83 patients with muscle-invasive bladder cancer were treated with TURBT (as thoroughly as possible) and definitive radiotherapy (median total dose: 64 Gy, median fractional dose: 2 Gy). Cystectomy was performed when possible for the residual or recurrent invasive cancer following radiotherapy. The median follow up period was 76 months. RESULTS: The overall survival (OS) and bladder preserving survival (BPS) rates at 5 years were 38% and 28%, respectively. Univariate analysis indicated that depth of invasion (T2 vs. T3), tumor diameter (< 3 cm vs. > or = 3 cm), and visible (R1) or not visible (R0) residual tumor after TURBT influenced both OS and BPS. In multivariate analysis, absence of visible residual tumor after TURBT was the only significant prognostic factor related to OS (p < 0.001) and BPS (p = 0.002). Five-year OS and BPS were 54% and 43% in T2-3R0 and 14% and 7% in T2-3R1, respectively. CONCLUSIONS: Absence of visible residual tumor after TURBT was significantly associated with better overall survival and bladder-preserving survival for muscle-invasive bladder cancer patients treated with definitive radiotherapy following TURBT. PMID- 10536449 TI - [Usefulness of visualized highly sensitive new chemical dosimeter: application to blood irradiation]. AB - In the prophylaxis of post-transfusion GVHD, it is recommended to irradiate blood components with an absorbed dose ranging from 15 Gy to 50 Gy. We applied a new chemical dosimeter to measure absorbed dose for blood irradiation. This system is composed of diphenyliodoniumchloride (DICI) as an oxidizer and crystal violet lactone as a pH indicator. Protons are released from DICI after irradiation. The maximum absorption wavelength of this indicator is 610 nm, and absorbance is linearly related with a given dose from 1 to 50 Gy. Coloration is stable for at least 2 weeks when stored in the dark at 4 degrees C. This system may be used as a chemical dosimeter for blood irradiation. PMID- 10536448 TI - [Hard-copy (film) versus soft-copy (CRT) reading performance between compressed and uncompressed images: SOLs in abdominal CT images]. AB - The objective of this study was to examine the feasibility of soft-copy (CRT) reading and suitable compression. Forty abdominal CT images with a space occupying lesion (SOL) in liver and 40 normal images were selected for receiver operating-curve (ROC) analysis. Each image was compressed by JPEG extended mode into 1/10 its original capacity, and then an expanded image was printed on film. Ten radiologists evaluated the presence of liver SOLs (primary and secondary tumors) on soft-copy (CRT) and hard-copy (film) images. Each radiologist reviewed four types of images (original and compressed hard-copy and original and compressed soft-copy images). Values of the area under the curve in the various ROC analyses were 0.858 (FILM) and 0.842 (CRT) for original images and 0.879 (FILM) and 0.846 (CRT) for compressed images. The results of ROC analysis showed better reading performance with hard-copy than soft-copy images, but the difference was not statistically significant. Compressed images showed a higher value (0.879) than original images (0.858), a difference that was statistically significant (p < 0.029) by the paired t-test but not by the jackknife method. The results indicate that soft-copy reading is a clinically acceptable alternative to hard-copy reading. We have had no difficulty in reading abdominal CT images compressed to 1/10 of the original size by the JPEG method. This study was supported in part by a grant from the Japanese Ministry of Health and Welfare. PMID- 10536450 TI - [Development of a multiple side-hole sheath introducer for use in aortography]. AB - We developed a multiple side-holed sheath introducer that can also be used as a catheter for aortography before selective arteriography of the pelvic organs or lower extremities, as an alternative to straight or pig-tail catheters, which were formerly used for aortography. Aortography using the multiple side holed sheath introducer provided good detection of arteries in 100% of 10 cases, without any complications or difficulties. Thus, the present sheath introducer can replace straight or pig-tail catheters, resulting in the more economical use of one catheter prior to selective arteriography. PMID- 10536451 TI - [Consideration of normal or reference values in the clinical laboratory tests for aged people--investigation in 27 biochemical tests]. PMID- 10536452 TI - [PDGF and PDGF receptors]. PMID- 10536453 TI - [Separate measurement of radial trabecular and cortical bone mineral density by peripheral quantitative computed tomography (pQCT) in degenerative joint disease]. AB - Degenerative joint disease and osteoporosis, both of which tend to increase with advance in age, were thought to be essentially different mainly because of the artifactually high lumbar spine bone mineral density (BMD) in the former unlike osteoporosis characterized by persistent decrease of BMD. To clarify the relationship between these two diseases, three-dimensional BMD of the trabecular and cortical bone was measured separately in an area of the radius relatively free of degenerative changes by peripheral computed tomography (pQCT). In addition to radiological assessment of spondylosis deformans, quantification of vertebral deformity was attempted by calculation of standard deviation, coefficient of variation, difference between maximum and minimum density divided by the mean of L1-L4. With advance in age and progress of spondylosis deformans, the standard deviation, coefficient of variation difference between the maximum and minimum density and this difference divided by the mean of L1-L4 increased, but radial trabecular bone density, cortcal density and relative cortical volume decreased, suggesting parallel advance of degenerative joint disease and osteoporosis. Sodium etidronate, an antiresorber commonly used in the treatment of osteoporosis, increased mean lumbar spine BMD and markedly decreased the standard deviation, coefficient of variation, difference between the maximum and minimum density and this difference divided by the mean of L1 and L2 but maintained maximum BMD constant, decreasing vertebral deformity due to spondylosis deformans. It is conceivable that calcium release from bone on increased resorption leads to osteoporosis, and calcium entrance into cartilage, causing its hardening, disappearance and degeneration, direct contact between bones, osteoarthritis and subsequent deformity in a single sequence of events. PMID- 10536454 TI - [Association of physical performance and falls among the community elderly in Japan in a five year follow-up study]. AB - Since falling is one of the main causes of morbidity and mortality among the elderly, attention should be paid to prevention. The purpose of the present study was to investigate the risk factors for falls among the elderly by a 5-year longitudinal observation. The subjects were the elderly aged 65 years and over living in a rural community in the eastern part of Honshu, Japan. Of the 852 eligible subjects, 685 persons (278 males and 407 females) received the multidimensional medical examination including physical performance as the baseline of TMIG-LISA (Tokyo Metropolitan Institute of Gerontology, Longitudinal and Interdisciplinary Study on Aging) in 1992. From baseline medical examinations, the following variables were adopted to investigate risk factors for falls (2 or more times during the 5-year follow-up): self-rated health, experience of falling, TMIG-index of competence, visual deficit, anthropometric measurements (height, weight, BML total skinfold thickness), lumbar bone mineral density, and physical performance tests (grip strength, finger-tapping rate, one leg standing time with eyes open or closed, preferred and maximum speed walking for 5 meters). A total of 108 subjects experienced falls two or more times during the 5-year follow-up period (43 men and 65 women). There were significant differences in grip strength, preferred and maximum walking speeds between those who fell and those who did not. Multiple logistic regression analysis controlled for age, sex and other variables adopted in this study revealed the following variables were significantly and independently related to falling: experience of falling within one year preceding the baseline survey (directly, p < 0.0001), preferred walking speed (inversely, p < 0.005) and total skinfold thickness (inversely, p < 0.01). These results suggest that the walking test as well as the experience of falling have discriminant and predictive validity for assessing the future occurrence of falls among the elderly. PMID- 10536455 TI - [Factors influencing clients' return home on discharge from geriatric intermediate care facilities in a metropolitan suburb]. AB - The present study investigated factors influencing the destination following discharge from geriatric intermediate care facilities (GICF) located in a suburb of a metropolis. A set of questionnaires was prepared, and a survey was conducted involving 204 subjects (46 men, 158 women, average age 84 years) of three GICFs and their families. Seventy-two percent of clients were termed bedridden and 84% as having dementia. Furthermore, 63% of them were admitted to GICFs from hospitals, and about half used the GICFs more than twice. Most families cared for the subjects before admission into the GICFs, and were feeling the burden of daily care. The families desired them to stay in the GICFs for as long as possible. Forty-five percent of the families wanted the subject to go back home after discharge. About 30% of subjects returned home, 31% were admitted to a hospital and 40% were placed in GICFs. The following factors were related to discharge and return home: the family wanted the subject to return home, subjects admitted to GICFs from home could return home, good ability to walk, families consulted subjects about the destination following discharge, the cost of staying at the GICF was covered by the subjects' pension, and confirmed administration of drugs was not necessary. These findings suggest that in order for clients at GICFs to successfully return home, the facilities should consider where the family wants the clients to go. PMID- 10536456 TI - [Bladder dysfunction in dementia patients showing urinary incontinence: evaluation with cystometry and treatment with propiverine hydrochloride]. AB - In a total of 46 dementia patients with urinary incontinence, filling cystometry was performed to evaluate bladder function, followed by peroral administration of propiverine hydrochloride. Therapeutic effects were evaluated in terms of changes in cystometric bladder capacity and urinary incontinence. Based on cystometric findings, 58.1%, 35.3% and 6.4% of Alzheimer dementia patients proved to have overactive bladder, normoactive bladder and low compliant bladder, respectively. Similarly, 90.9% and 9.1% of vascular dementia patients had overactive detrusor and low compliance, respectively. As a result of propiverine hydrochloride administration for 2 weeks, both bladder capacity and incontinence improved in around 40%, irrespective of the dementia type (Alzheimer vs vascular). It is of interest, however, that patients with overactive bladder demonstrated more satisfactory response to the treatment than those with normoactive bladder. It is accordingly suggested that propiveline hydrochrolide is a promising treatment option for urinary incontinence in demented elderly and cystometry could be useful in predicting treatment outcome. PMID- 10536457 TI - [An elderly case of chronic inflammatory demyelinating polyneuropathy with acute onset in the course of diabetes mellitus]. AB - A 77-year-old man was admitted because of muscle weakness in both upper and lower extremities. Diabetes mellitus was diagnosed in 1988 and he had been treated by oral hypoglycemic agents. He had a common cold at the end of January, 1997. Muscle weakness appeared in the upper extremities, followed by the lower extremities at the end of February. No sensory disturbance or dysuria was recognized. Nerve conduction study revealed distally dominant demyelinating polyneuropathy. Guillain-Barre's syndrome was diagnosed and he recovered completely following immunological absorption therapy (IAT). However, he had quadriplegia again at the end of April. He was treated by IAT combined with corticosteroid and has shown no relapse. In June, 1997, gastric cancer was detected by upper gastrointestinal fiberscopy and subtotal gastrectomy was performed. Judging from this clinical course, this case seems to be chronic inflammatory demyelinating polyneuropathy (CIDP) with acute onset. Many kinds of causes often contribute to the pathogenesis of neuropathy in the elderly. So in cases of progression or worsening, we should consider such possibilities and it is necessary not to exclude treatable causes of neuropathy. PMID- 10536458 TI - [A case of successful treatment of postprandial syncope with combined use of amezinium metilsulfate and dihydroergotamine mesylate]. AB - An 86-year-old man, with a history of old cerebral hemorrhage, experienced repeated syncope after the meal. The blood pressure significantly decreased at the onset of syncope, and postprandial hypotension (PPH) was diagnosed. In order to treat the syncope, we administered midodrine hydrochloride, amezinium metilsulfate and dihydroergotamine mesylate. However, each of these drugs failed to prevent PPH and the syncope persisted. Then we administered a combination of amezinium metilsulfate and dihydroergotamine mesylate. This led to the disappearance of the syncope together with PPH. Combined use of these drugs should be further evaluated in the elderly patients with PPH. PMID- 10536459 TI - [Cellular kinetics for a biological classification of lymphoma]. PMID- 10536460 TI - [Quo vadis, cardiopathology?]. PMID- 10536461 TI - CATCH 22 syndrome: report of 7 infants with follow-up data and review of the recent advancements in the genetic knowledge of the locus 22q11. AB - CATCH 22 is a medical acronym for Cardiac defects, Abnormal facies, Thymic hypoplasia, Cleft palate, and Hypocalcemia, and a variable deletion on chromosome 22. The deletion within the chromosome region of 22q11 may occur in patients with three well-described dysmorphologic+ cardiological syndromes: DiGeorge syndrome (DGS), velocardiofacial syndrome (VCFS), and conotruncal anomaly face syndrome (CTAFS). We report in detail seven infants with a deletion of the locus 22q11 showing overlapping clinical features of DGS and CTAFS with complex congenital heart defects (double outlet right ventricle, atresia or stenosis of the pulmonary valve, atrial and ventricular septal defects, patent ductus arteriosus, tetralogy of Fallot, major aortopulmonary collateral arteries, arcus aortae dexter, and persistence of the left superior vena cava). A homograft was implanted between the right ventricle and the main stem of the pulmonary artery in 2 patients, while a balloon valvuloplastic of the pulmonary valve was performed in one patient only. Pulmonary hemorrhage, acute hypoxia, and Aspergillus pneumonia were the complications. Death occurred in three out of seven patients. Recent advancements in the genetic knowledge of the locus 22q11 are described. Since the locus 22q11 is highly heterogeneous, the CATCH 22 acronym should be used and temporarily the old eponyms should be abandoned waiting for the identification of the different genes. PMID- 10536463 TI - [Microcalcifications associated with in situ ductal breast carcinoma. Mammographic and histological correlations with morphometric evaluation]. AB - This study includes 33 cases of ductal carcinoma in situ and its aim is to detect and classify microcalcifications according to their appearance, mammographically, histologically and morphometrically. From the histological point of view, intraductal carcinomas are classified following the criteria proposed by Holland et al. The types of carcinoma examined reveal the presence of different microcalcifications. The calcifications associated with G1 carcinomas appear as highly compact morphometrically, fine or mainly so mammographically and laminated or mixed histologically; no granular calcifications are observed. On the other hand, calcifications associated with type G2 appear as coarse mammographically, granular histologically and scarcely compact morphometrically. Finally, G3 carcinomas mainly reveal by mammography vermicular calcifications (in a few cases associated with fine ones). Granular calcifications are predominant on histological examination while morphometry shows the poorest level of compactness. PMID- 10536462 TI - [Computerized histological analysis in chronic viral hepatitis]. AB - Chronic liver diseases evolve towards cirrhosis by means of the progressive substitution of hepatic tissue with fibrotic areas. In this work we measured the percentage of fibrotic areas with respect to the total area of the bioptic specimens, utilizing a histomorphometric computerized evaluation. The specimens were obtained by 39 patients affected to chronic viral liver diseases, before and after a therapy with alpha interferon. The aim of this study is to determine the architectural sprain degree at the different stages of chronic liver disease, and to verify if the response to the therapy with interferon is associated with a reduction of the fibrotic areas within the hepatic tissue. The comparison between our computerized index and the Knodell index indicates that the first can be considered an useful system for the evaluation of hepatic biopsies from patients affected by chronic viral hepatitis. PMID- 10536464 TI - [Angiodysplasia of the duodenum with arteriovenous shunts, sub-serosal prevalence and interstitial elastosis; peritonitis and rupture of sub-serous dysplasic vessels and hemoperitoneum. Description of a case and review of the literature]. AB - We present a case report of a 74 year old man, affected by angiodysplasia of the duodenum, which developed in peritonitis and hemoperitoneum. After undergoing two surgical interventions, it is 14 days since he died. First of all, the importance of this case is due to the extreme rarity of the angiodysplasia of the duodenum, specially when it becomes worse in peritonitis and hemoperitoneum, this case having been described only one time till now, then the localization mainly sub serousal and lastly the presence of a component, characterized by an elastotic nature, disposed in islets and interstitial widespread also among the glands, which we have first described in literature. PMID- 10536467 TI - [Breast cytology guidelines for a mammographic screening program. Report edited by the working group on "Breast Pathology in the Mammographic Screening Program" of the European Union]. PMID- 10536465 TI - [Nephrogenic adenoma of the bladder. Morphological and immunophenotypic study with particular attention to differential diagnosis]. AB - BACKGROUND: Nephrogenic Adenoma (NA) is a rare lesion of the urinary tract, considered a metaplastic response to chronic inflammation, trauma or immunosuppression. METHODS AND RESULTS: We report two cases of NA arising in the urinary bladder of patients with previous history of recurrent urinary tract infections due to neuropsychiatric disease. Pathological examination of the lesions, resected by transurethral (TUR) management, revealed a papillary proliferation of tubules and cysts lined by cuboidal to low-columnar cells without atypia. Immunohistochemistry showed positivity for Cam 5.2, CK7 and EMA. MIB 1 count demonstrated a positivity in 12/200 cells in case 1 and < 2/200 in case 2. No expression of nuclear p53 was evident. CONCLUSION: NA is a benign unusual neoplasm which might be misdiagnosed as clear cell adenocarcinoma of the bladder or prostatic adenocarcinoma. Its recognition is important because it is a benign lesion cured by a conservative resection and no additional therapy is generally required. PMID- 10536468 TI - [Analytical diagnosis of melanocytic nevi]. PMID- 10536466 TI - [Rhabdoid tumor of the kidney arising in a newborn infant: description of a case with ultrastructural observations]. AB - INTRODUCTION: Rhabdoid tumour of the kidney is a new independent entity. Before it was considered a variant of Wilms tumor of the kidney. Now we have enough parametres to define the rhabdoid tumor: immunohistochemistry positive by the vimentin, special histological features and behaviour. CASE: We report a very aggressive case of rhabdoid tumor found in a two week old infant. RESULTS: We studied aspects of histology and histochemistry. We found a positivity for vimentin and many cells in apoptosis. DISCUSSION: The mild positivity for vimentin and the high number of cells in apoptosis suggest a relationship between apoptosis and behaviour. PMID- 10536469 TI - Relationship between radionuclide right ventricular ejection fraction and clinical status in patients with left ventricular dysfunction after myocardial infarction. AB - OBJECTIVE: To evaluate the influence of right ventricular (RV) function, determined by RV ejection fraction, on the clinical status of patients with ischemic heart disease and left ventricular (LV) EF under 40%. BACKGROUND: The role of RV function as a marker of prognosis in heart failure has been debated. We hypothesized that the degree of RV dysfunction is a determinant of the clinical status and outcome of patients with LV dysfunction after myocardial infarction. PATIENTS AND METHODS: 30 patients, 25 male, with previous myocardial infarction, more than 6 months age, were studied by equilibrium radionuclide angiography. Functional capacity was evaluated by cardiopulmonary exercise test with Naughton protocol. Patients were followed during a 12 month period for major clinical events: death or hospitalisation for congestive heart failure. Two groups of patients were considered according the value of RVEF (< or = 30% and > 30%). RESULTS: The values of EF were: LV = 25 +/- 7% and RV = 35 +/- 9%. Maximum oxygen consumption correlated with RVEF (r = 0.78, p < 0.001) but not with LVEF (r = 0.12, NS). The group of patients with RVEF > 30% had a greater exercise time (712 +/- 229 versus 441 +/- 208 seconds, p = 0.003), higher oxygen consumption (19.8 +/- 5.3 versus 13.5 +/- 3.3 ml/kg/min, p = 0.001) and oxygen consumption in relation to the maximum predicted for age and sex (71 +/- 19 versus 50 +/- 13%, p = 0.002). Cumulative frequency of major clinical events was greater in the group with RVEF < or = 30% (58% vs 6%, relative risk 3.14, 95% CI 1.23 to 5.05). There was no correlation between the values of LVEF and outcome. CONCLUSIONS: In this setting of ischemic LV dysfunction, the RVEF correlates with functional capacity in cardio-pulmonary exercise test and the presence of RV dysfunction is associated to a higher incidence of clinical events. PMID- 10536470 TI - [Primary hyperaldosteronism--12 clinical cases]. AB - STUDY OBJECTIVES: To show clinical, biochemical, and morphological data of 12 patients with primary hyperaldosteronism: eight with an aldosterone-producing adenoma and four with adrenal hyperplasia. To compare clinical and biochemical parameters of the patients with adenoma and hyperplasia. For those with adenoma, to verify clinical and biochemical modifications after adrenalectomy. PATIENTS AND METHODS: In the 12 patients with hyperaldosteronism, retrospective analysis of clinical (age, sex, blood pressure), biochemical (plasmatic and urinary potassium, plasmatic aldosterone, plasma renin activity, and plasmatic aldosterone/renin activity ratio), and morphological (computed tomography, magnetic resonance, and norcholesterol scintigraphy) data was performed. RESULTS: 1--In the 12 patients with hyperaldosteronism (seven female), the age was 51.0 +/ 10.2 years (mean +/- standard deviation), the systolic pressure 200.9 +/- 34.5 mm Hg and the diastolic pressure 120.0 +/- 12.3 mm Hg. Hypertension was diagnosed 12.0 +/- 10.1 years before. As biochemical evidence, we found kalaemia of 3.06 +/ 0.28 and urinary potassium of 63.4 +/- 16.5 mEq/l, renin activity 0.98 +/- 1.02 ng/ml/h, plasmatic aldosterone of 49.4 +/- 36.0 ng/dl, aldosterone/renin activity > 30 in 83% of the cases. As morphological evidence, computed tomography allowed diagnosis in nine patients, suggested it in two, being doubtful in one. Performed on four patients, resonance confirmed the tomography in three and was not contributive in one. The scintigraphy performed in four patients visualized two adenomas, was negative in one adenoma and in one hyperplasia. 2--In the eight patients with adenoma (six female), the youngest age and the highest diastolic pressure compared with patients with hyperplasia were statistically significant (p < 0.01 and 0.05). In the adenomas, the biochemical changes were more pronounced, but not statistically significant. The plasmatic aldosterone/renin activity ratio was also higher in the adenoma cases. 3--After the adrenalectomy, blood pressure became normal in five patients and was more easily therapeutically controlled in three. The average systolic and diastolic pressures decreased and the biochemical parameters became normal in all patients. The pre/post surgical modification of these parameters had statistical significance (systolic pressure decrease, p < 0.01; diastolic pressure decrease, p < 0.01; kalaemia increase, p < 0.001; renin activity increase, p < 0.01; aldosterone decrease, p < 0.02). The plasmatic aldosterone/renine activity ratio normalized in all patients. CONCLUSIONS: In diagnosing primary hyperaldosteronism, biochemical (kalaemia, urinary potassium, plasmatic aldosterone, renin activity, aldosterone plasmatic/renin activity) and tomography studies were important. On comparing the patients with hyperplasia with those with adenoma, we found that the latter are younger and exhibit higher diastolic pressure, both findings with statistical significance. After adenoma surgery, blood pressure became normal in five patients and improved in three, these findings, and the improvement of the kalaemia, plasmatic aldosterone, and renin activity parameters were statistically significant. PMID- 10536471 TI - [The use of Abciximab in patients with acute myocardial infarction undergoing direct percutaneous coronary revascularization]. AB - OBJECTIVE: To evaluate the initial experience, in our Centre, with Abciximab in patients with acute myocardial infarction undergoing direct percutaneous transluminal coronary angioplasty (PTCA). METHODS: Between October 1996 and May 1998, 65 patients (51 males, mean age 56.9 +/- 11 years) underwent direct PTCA for acute myocardial infarction. In thirty-seven patients the myocardial infarction was anterior and 40 had multivessels disease. Mainly to compare the incidence of bleeding complications we considered 2 groups: Group A--17 patients submitted to PTCA without the use of Abciximab, and Group B--48 patients submitted to PTCA and to a bolus followed by a 12 hour infusion of Abciximab. All the patients were treated with aspirin and heparin (5,000 to 15,000 U according to ACT) and ticlopidine in case of stent implantation. RESULTS: Percutaneous coronary revascularization was successfully achieved in 92.3% of the patients. The total number of bleeding complications was ten cases (20.8%) in Group B and 1 case (5.8%) in Group A. Most of the bleeding complications in the Abciximab Group were minor and related to the femoral vascular access site (9 cases--18.7%) and were easily resolved with local measures (8 cases). There were also 3 cases of hematemesis and one of oral bleeding, all well tolerated. Major bleeding complications were identified in only one patient of the Abciximab Group related to the vascular access site, however there was an absolutely similar case in Group A (2% versus 5.8%). CONCLUSIONS: Although bleeding complications were more frequent in patients receiving Abciximab, mostly related to the vascular access site, they were transient and well tolerated. PMID- 10536472 TI - [Atrial fibrillation in Wolff-Parkinson-White syndrome]. AB - The authors describe the main etiopathogenic factors and clinical importance of atrial fibrillation and analyse the results of catheter ablation of atrioventricular accessory pathways in Wolff-Parkinson-White syndrome. Atrial vulnerability is the principal mechanism and radiofrequency catheter ablation of atrioventricular accessory pathways seems to be useless to prevent atrial fibrillation. PMID- 10536473 TI - [IX Forum of the Portuguese Society of Cardiology. Ethical problems in the practice of cardiology. Opening remarks]. PMID- 10536474 TI - [Ethical controls of drug trials]. PMID- 10536475 TI - [Cost containment, distribution of resources and the beginning of equity]. PMID- 10536476 TI - [Ethical problems of care provided to terminal cardiological patients]. PMID- 10536477 TI - [Isolated supravalvular aortic stenosis]. PMID- 10536478 TI - Prediction of hospital readmission for heart failure: development of a simple risk score based on administrative data AB - OBJECTIVES: The purpose of this study was to develop a convenient and inexpensive method for identifying and individual's risk for hospital readmission for congestive heart failure (CHF) using information derived exclusively from administrative data sources and available at the time of an index hospital discharge. BACKGROUND: Rates of readmission are higher after hospitalization for CHF. The significant determinants of rehospitalization are debated. METHODS: Administrative information on all 1995 hospital discharges in New York State which were assigned International Classification of Diseases--9--Clinical Modification codes indicative of CHF in the principal diagnosis position were obtained. The following were compared among hospital survivors who did and did not experience readmission: demographics, comorbid illness, hospital type and location, processes of care, length of stay and hospital charges. RESULTS: A total of 42,731 black or white patients were identified. The subgroup of 9,112 (21.3%) who were readmitted were distinguished by a greater proportion of blacks, a higher prevalence of Medicare and Medicaid insurance, more comorbid illnesses and the use of telemetry monitoring during their index hospitalization. Patients treated at rural hospitals, those discharged to skilled nursing facilities and those having echocardiograms or cardiac catheterization were less likely to be readmitted. Using multiple regression methods, a simple methodology was devised that segregated patients into low, intermediate and high risk for readmission. CONCLUSIONS: Patient characteristics, hospital features, processes of care and clinical outcomes may be used to estimate the risk of hospital readmission for CHF. However, some of the variation in rehospitalization risk remains unexplained and may be the result of discretionary behavior by physicians and patients. PMID- 10536479 TI - [Does clinical and medical science for rheumatology hold true as pure life science?]. PMID- 10536480 TI - [Investigation of plasma levels of etodolac and urine PGE2 in patients with rheumatoid arthritis]. AB - 13 women patients (containing high-aged, mean age 71.9 years) with rheumatoid arthritis (RA) (10 with normal renal function and 3 with moderate renal insufficiency) participated in a 5-day study to assess the effects of etodolac on renal function and the necessity of dose adjustment. After no drug-free day, etodolac, 200 mg b.i.d. was started. The plasma levels of etodolac were similar in both normal control in phase I studies and throughout this study. Although the mean urine PGE2 concentration in renal insufficient group was significantly lower than that of normal renal function group, the mean urine PGE2 concentration in after etodolac administration was not different from that in before its administration in each group. This result suggested that renal adverse reactions with etodolac were low in incident. Moreover, it was required to consider that glucocorticoid might influence renal and/or hepatic excretion of etodolac. In this study the glucocorticoid was tend to be administrated in the patients with moderate renal insufficiency. Those glucocorticoid group showed lower levels of etodolac in blood serum (monitored by AUC0-8, day 4 Cmin and day 5 Cmin) than non glucocorticoid group did, but not significantly. Interestingly, there is a negative correlation (r = -0.442) between AUC0-8 at day 1 and urine PGE2 at day 5 in glucocorticoid group. The levels of etodolac in blood serum in normal renal function group were also not significant in difference from that in the moderate renal insufficient group. These results suggest that the dose adjustment of etodolac in high-aged RA patients with moderate renal insufficiency can be excluded. PMID- 10536481 TI - [A case of spontaneous osteonecrosis of the grossly depressed medial tibial plateau]. AB - The patient of this case was a 64 year-old woman who visited our hospital with a complaint of arthralgia in the left knee lasting from nearly seven months ago. The radiogram taken at the first visit revealed no abnormalities in the knee, but she had persistent arthralgia, and so prednisolone was intraarticularly injected at a dose of 25 mg twice with an one-week interval. One month later, varus deformity rapidly progressed, resulting to cause gait disturbance. Since comminution and depression of the tibial plateau were revealed by radiography, steroid arthropathy was suspected. Histological examination showed normal articular cartilage and osteonecrosis in the subchondral bone. In addition, new bone formation was observed in the region around the tibial intercondylar eminence apart from the necrotic area, indicating extensive bone remodeling. On these findings, the patient was diagnosed as the spontaneous osteonecrosis. At that time unicompartmental knee arthroplasty (UKA) was done. Thereafter five years have passed and a revision TKA with bone graft was made because of the loosening of the prosthesis. Though the postoperative course is satisfactory, osteonecrosis developed in the contralateral medial femoral condyle, so that she is receiving a conservative treatment. PMID- 10536482 TI - [An infant of autoimmune hepatitis (type I) with cirrhosis]. AB - A 6 year-old boy with autoimmune hepatitis accompanied with cirrhosis was reported. He was admitted to our hospital because of abdominal distention, high fever, and diarrhea. Laboratory examination revealed abnormalities in hepatic function, cholestasis, anemia, thrombocytopenia, hypoalbuminemia, hypocomplementemia, and low concentration of coagulation factors. Abdominal MRI, and asialoglycoprotein receptor-mediated liver scintigraphy strongly indicated liver cirrhosis. Viral hepatitis, Wilson's disease, and antitrypsin deficiency were excluded serologically. Instead, hypergammaglobulinemia, and positive antinuclear antibody suggested autoimmune hepatitis, and the survey of anti mitochondrial antibody, anti-smooth muscle antibody, and anti-LKM-1 antibody was negative, indicating type I autoimmune hepatitis. Finally, the histology of liver biopsy specimen indicating the destruction of hepatic lobular architecture, dense mononuclear cell infiltrates, and severe fibrosis confirmed the diagnosis. He was treated firstly with methylprednisolone pulses, and then prednisolone p.o. + azathioprine p.o. All of the abnormal laboratory parameters improved to normal levels, indicating that the immunosuppressive therapy will be effective for the severe AIH with cirrhosis. PMID- 10536484 TI - [A child of microscopic polyarteritis nodosa effectively treated with intravenous methylprednisolone pulses and serial cyclophosphamide pulse therapy]. AB - Microscopic polyarteritis nodosa (mPAN) is a rare disorder in pediatric field of systemic small vessel vasculitis, and affects skin and musculoskeletal system accompanied by progressive necrotizing glomerulonephritis. We described here a 15 year-old boy with positive anti-neutrophil cytoplasmic antibody (MPO-ANCA) and severe mPAN, who was effectively treated with intravenous methylprednisolone pulse therapy followed by monthly cyclophosphamide pulses for 1 year. The histologic examination of renal biopsy specimen showed a severely desolated disease characterized by fibrinoid necrosis, crescent formation of most glomeruli and interstitial infiltration of inflammatory cells. The elevated titers of MPO ANCA were useful markers for diagnosis, and the serial determinations of the antibody titers were indicative of disease activity. Moreover, dramatic clinical improvement after the induction of the combinatorial therapy and the disappearance of MPO-ANCA was correlated in the disease course. In this report, the serial determination of MPO-ANCA constituted a useful diagnostic tool and a sensitive marker of disease activity. PMID- 10536483 TI - [Portal and pulmonary hypertension in a patient with MCTD]. AB - A 42-year-old woman with mixed connective tissue disease (MCTD) died due to the rupture of esophageal varices. The autopsy revealed fresh thrombi in the main trunk of the portal vein. Microscopic examination disclosed wide-spread periportal fibrosis and stenosis of peripheral portal veins without remodeling of hepatic lobular architecture, which was compatible to idiopathic portal hypertension (IPH). Anti-phospholipid antibody was negative. Accordingly it is likely that portal vein thrombosis developed secondary to IPH. In the literature 6 (37.5%) of 16 collagen vascular disease patients with IPH died, and three of them were due to rupture of esophageal varices. Therefore IPH should be considered to be one of the most important complications affecting its grave prognosis in patients with collagen vascular disease. The patient also had had pulmonary hypertension (PH) when the diagnosis of portal hypertension was made. In the literature we found 5 collagen vascular disease patients with both PH and IPH like this case. The most outstanding common clinical feature among these 6 patients was Raynaud's phenomenon associated with positive anti-RNP antibody. Moreover 5 of 6 cases including this case simultaneously developed both PH and IPH. The clinical course of these patients suggests there may be a common pathogenetic factor for these two lesions. A possible candidate involved in the pathogenesis of PH and IPH may be endothelin, one of the vasoactive substances, since its receptor is said to be expressed abundantly in pulmonary and portal vasculatures. However, further investigation is necessary to elucidate the mechanism of PH and IPH in collagen vascular diseases. PMID- 10536485 TI - [Treatment for polymyositis/dermatomyositis]. PMID- 10536486 TI - [Systemic lupus erythematosus]. PMID- 10536487 TI - Increase of soluble cytoadhesive molecules sE-selectin and sICAM-1 and hyperfibrinogenaemia in patients with polytrauma without septicaemia. AB - Multiple organ failure with thrombophilia is suggested to occur in the course of polytrauma with septicaemia. The aim of the study was to investigate fibrinogen level, other proteins of acute phase response: positive-orosomucoid and negative transferrin and soluble cytoadhesive molecules in plasma of patients (n = 28) with polytrauma (I-II. stage according to Hannover score) without detectable bacteraemia until 36 hours post injury. The patients treated by massive blood transfusion were excluded. An increase of fibrinogen (Fbg pts 4.34 +/- 2.5 g/l versus control 2.55 +/- 0.55 g/l, p < 0.01), orosomucoid (ORM pts 1.47 +/- 0.8 g/l versus control 0.54 +/- 0.18 g/l, p < 0.01), SE-selectin (sE-sel. pts 92.11 +/- 79.4 g/l versus control 46.6 +/- 29.6 g/l, p < 0.05), sICAM-1 (sICAM pts. 698.3 +/- 54.4 versus control 255.6 +/- 58.0 g/l, p < 0.01) and a decrease of transferrin (Trf pts. 1.77 +/- 0.82 versus control 2.83 +/- 0.71 g/l, p < 0.01) were observed in this patients with polytrauma. We suppose that an increase of fibrinogen and cytoadhesive molecules sE-selectin and sICAM-1 may be induced in patients with polytrauma due to a "non-septic" inflammatory acute phase response in the course of wound healing process after tissue injury too. PMID- 10536488 TI - Haemostasis, cytoadhesive molecules (sE-selectin and sICAM-1) and inflammatory markers in non-insulin dependent diabetes mellitus (NIDDM). AB - Non-insulin dependent diabetes mellitus (NIDDM) is connected with a higher incidence of macrovascular atherosclerotic disorders. The aim of the study was to detect any difference in levels of "cardiovascular risk factors"--fibrinogen, PAI 1 and inflammation response (documented by an increase of protein of acute phase orosomucoid) and of soluble cytoadhesive molecule sE-selectin and sICAM-1 (as markers of endothelial dysfunction) in blood plasma of 118 patients with NIDDM in comparison to the levels in blood plasma of 59 healthy persons as a control group. We observed higher levels of fibrinogen (fibrinogen level was 3.44 +/- 1.02 g/l in NIDDM pts versus 2.44 +/- 0.55 g/l in control group, p < 0.01) and PAI-1 Ag concentration was 159.7 +/- 110.3 ng/ml in NIDDM pts versus 51.43 +/- 24.64 ng/ml in control group, p < 0.01) together with an increase of acute phase protein orosomucoid as a "inflammatory response marker" (orosomucoid concentration was 0.85 +/- 0.23 g/l in NIDDM pts versus 0.54 +/- 0.18 g/l in control group, p < 0.01) in patients with NIDDM. The increase of these "cardiovascular risk factors" levels will be probably induced by higher activity of inflammatory cytokines IL-1 beta and/or TNF alpha in NIDDM patients, because both are inducers of orosomucoid fibrinogen and PAI-1 synthesis. This hypothesis is also supported by observation of higher levels of soluble cytoadhesive molecules sE-selectin (sE-selectin level was 64.25 +/- 26.8 ng/ml in NIDDM pts versus 46.64 +/- 29.57 ng/ml in control group, p < 0.01) and sICAM-1 (sICAM-1 level was 307.71 +/- 86.2 ng/ml in NIDDM pts versus 255.6 +/- 58.0 ng/ml in control group, p < 0.01) in patients with NIDDM. Both cytoadhesive molecules are produced by endothelial cells which are influenced by IL-1 beta and/or TNF alpha. According to these findings we suppose that an "inflammation" plays an important role in the evolution of atherosclerotic process at NIDDM together with the known influence of glucose and lipid metabolism pathology. PMID- 10536489 TI - The effect of glycosaminoglycan sulodexide on oxidative stress and fibrinolysis in diabetes mellitus. AB - Glycosaminoglycan sulodexide may influence morphology and functional properties of the basement membranes in microvessels. The aim of this study was to evaluate the effect of sulodexide administration on albuminuria and on different biochemical variables indicating endothelial dysfunction, oxidative stress and fibrinolysis in diabetic patients. Twenty diabetic patients of both types with micro- or macroalbuminuria were selected for sulodexide treatment. Daily dose of 600 U (60 mg) was injected intramuscularly five days a week. Fifteen doses were applied during 3 weeks. The patients were examined before and after treatment as well as 6 months later. No changes of diabetes control were observed during the study and after 6 months of wash-out period. Significant decrease of albuminuria (p < 0.001) was observed during the sulodexide administration with the following increase to pretreated values during the wash-out period. A decrease of serum N acetyl-beta-glucosaminidase (NAG) activity (p < 0.03) at the end of treatment as compared to pretreated values was found in the whole group of diabetic patients. Slight reduction of oxidative stress expressed by malondialdehyde and superoxide dismutase was apparent after treatment but no simultaneous change in fibrinolysis was observed. Sulodexide may have some protective effects influencing functional properties of the basement membrane as manifested by lowered albuminuria. In addition, it may slightly decrease oxidative stress in diabetic patients and it could stabilize endothelial cells. PMID- 10536490 TI - [Complications of prevention and treatment of erythrocyte alloimmunization (case reports)]. AB - OBJECTIVES: The treatment of late recognized alloimunization with intraumbilical transfusions is more difficult and more often connected with complications. MATERIAL AND METHOD: Between 1991-1997 we performed 70 intraumbilical transfusions in 25 fetuses for erythrocyte alloimunization. Six fetuses (24%) were hydropic in the beginning of the treatment. Eleven fetuses were delivered before 36 weeks of pregnancy. Two immature neonates (660 g and 1320 g) had intraventricular hemorrhage with neurologic complications. In six cases the transfusion was complicated by severe bradycardia of the fetus, but only twice the pregnancy was to be terminated by cesarean sectio during 24 hours after the procedure. Two of the 25 fetuses died antenataly and one postnataly, all of them primary hydropic. Two neonates had severe late onset anemia. CONCLUSION: Fetal alloimune anemia should be treated before onset of hydrops. The study was supported by the grant of IGA Ministry of Health CR No. 3200-3. PMID- 10536491 TI - [Changes in strategy and technics in the surgical treatment of hernias]. AB - During last five years, 1,271 patients with the diagnosis of hernia were operated at the IIIrd Surgical clinic, 1st Faculty of Medicine, Charles University in Prague. Types of hernia were not differentiated. The patients group includes primary inguinal hernias and their relapses, hernias in other localizations as well as hernias in cicatrices. Ligation of the hernia sac as a separated intervention was used in no case. Herniorrhaphy was performed in 52% of cases, hernioplasty in 48%. The hernioplasty/herniorrhaphy ratio increases every year. While the hernioplasty was performed in 34% of patients in 1993, in 1997 was used already in 53% of cases. During that period of time, prolene mesh fixed in the defect with prolene suture was almost exclusively used as implant. In very complicated cases, Goretex was applied with a very good result. Polytetrafluoroethylene is the best implant material at the present time, but its price restrains its general use. Prolene is available for a moderate price and complies sufficiently with requirements on mechanical strength and non irritability. PMID- 10536492 TI - [The professional style of written and verbal communication and its characteristics]. PMID- 10536493 TI - [A practical classification of medical sciences for bibliographic purposes in a university library]. PMID- 10536494 TI - [Study of disorders of the GH-IGF-I axis by monitoring levels of IGF-I binding protein]. AB - Measurement of IGF I (Insulin Like Growth Factor I) and its binding proteins (IGFBP) in serum has become a principal diagnostic tool at the determination of growth hormone (GH) status. These data could, at the correct interpretation, fully replace the stimulation tests necessary for the GH determination. A lot of firms were interested in the analysis of IGFBPs, most frequently based on the immunological principle between IGFBP and its antibody. The assessment of IGFBP-3 is the most spread among all IGFBPs in the studies of GH-IGF I axis. It is used both at the GH deficiency, and at the GH access, it has become a valuable marker at the therapy of children suffering from GH insufficiency. IGFBPs analysis based on electrophoresis and Western blotting with the following detection of ligan IGFBP by radioisotop or by its immunological reaction with antibody plays an important role at the clinical application research. The changes in IGFBP profile or in their fragments can be an important markers (even specific one) at different disease. The contemporary research in IGFBPs domain is fully concentrated to the IGFBPs action in cells. It has been found f.e., that IGFBPs inhibit the growth of carcinomic cells. PMID- 10536495 TI - [Electronic information sources in medicine]. AB - This article is a brief introduction into electronic information resources in the medical field. Several commonly believed myths claiming the Internet to be the best universal information source are discussed and reviewed of both currently used professional methods of access to information: CD-ROM and online, is provided. The closing part of the article provides descriptions for the most important electronic databases and their types. PMID- 10536497 TI - [Some aspects of sensorial asymmetries]. AB - The present review shows the variability assessing sensoric asymmetries. There are preference and performance tests. Sensoric asymmetries are strongest and most manifest for eyedness (eye preference and performance), descending through earedness and other less known asymmetries. A vast range of testing techniques have been used to assess eyedness. Sighting dominance and self-report are two of the most popular techniques. Other preference measures include observation of how people use tools and questionnaires. Performance test assess speed and accuracy in tasks stressing sensoric dexterity. Although questionnaires are generally thought to be reliable and valid instruments, there is a disagreement as to the nature, the number and weighting of the items to be included. PMID- 10536496 TI - [Effect of alcohol on circadian blood pressure]. AB - The effects of alcohol on blood pressure have been studied extensively. Abstention is recommended in high blood pressure as basic non pharmacological treatment. On the other hand short term lowering of blood pressure by alcohol is known. Blood pressure effects of alcohol vary according to chronicity and amount of intake. It is not known how alcohol affects the 24 hour profile of blood pressure, in particular day- and night-time differences. This explorative study investigates the effects of a single dose of alcohol in the evening on the 24 hour blood pressure profile. Nine individuals with essential hypertension (mean age 65.4 +/- 8.7 years) were compared to 10 normotensives (29.6 +/- 3.0 years). Blood pressure was followed on 2 consecutive days by means of a 24 hour ABPM. On one evening the test persons consumed 0.6 g/kg ethanol before bed time. Apart from the direct comparison of the two groups, effects of body weight and daily alcohol consumption were also considered. For analysis of the 24 hour recording the mean 24 hour values, the mean difference between day and night and loads (fraction of blood pressure > 140/90 mm Hg) as well as heart rate were used. Ethanol led to nocturnal drops of blood pressure in normotensives and hypertensives alike and thus to an increased day/night difference. The latter increased by 2 +/- 4 mm Hg for the systolic and 2 +/- 1 mm Hg for the diastole values in normotensives and by 6 +/- 2 mm Hg and 3 +/- 1 mm Hg, respectively, in hypertensives on the day of alcohol intake. This trend was more marked in individuals with smaller daily alcohol consumption as well as in obese hypertensives. The blood pressure differences were not significant in our test sample because of a large variance in the response. Two normotensives were found to be borderline hypertensives. They exhibited a marked increase of nocturnal blood pressure values above 140/90 mm Hg when compared to the control night. Our study indicates that alcohol consumption should be considered when evaluating 24 hour blood pressure recordings. The blood pressure lowering effect of alcohol during the night is able to modulate the day/night difference can also be useful as diagnostic criterion our data may be clinically relevant. PMID- 10536498 TI - ["I have swollen lymph nodes"]. PMID- 10536499 TI - [Enchondromatosis: Ollier's disease]. PMID- 10536500 TI - [Acquired amyloidosis. Patient: 38-year old house-wife, originally from Turkey]. PMID- 10536501 TI - [Working title: Will reducing alendronate increase the risk of bone fractures by a decrease in bone density? Benefit from alendronate therapy for women without prior fractures remains uncertain]. PMID- 10536502 TI - [trans-translation mediated by tmRNA]. PMID- 10536503 TI - [Crystallization of Michaelis complex and its further advancements]. PMID- 10536504 TI - [Is threonine aldolase identical to serine hydroxymethyltransferase?]. PMID- 10536505 TI - [Structural feature, putative function, and production of free N-glycan occurring in plant cells]. PMID- 10536506 TI - [Structure and regulation of phospholipase C-delta]. PMID- 10536507 TI - [Erythropoietin gene regulation]. PMID- 10536508 TI - [The novel immune system: V alpha 14NKT cells]. PMID- 10536509 TI - [Nordic Medicine. Continuing cooperation between the editorial boards of the Nordic Journals]. PMID- 10536510 TI - [Why more about hypertension?]. PMID- 10536511 TI - [On angiotensin II receptor antagonists in the treatment of hypertension]. PMID- 10536512 TI - [Cell biology of the small arterial vessels]. PMID- 10536513 TI - [Cholesterol ester transfer protein. A major player in cholesterol metabolism]. AB - Plasma cholesterol transfer protein (P-CETP) plays a central role in cholesterol metabolism by pronroting transfer of cholesteryl esters from high density lipoprotein (HDL) to very low density lipoproteins in exchange for triglycerides. The CETP-reaction may hereby be a critical factor in reverse cholesterol transport, i.e., the transfer of cholesterol from peripheral tissues to the river for catabolism. Human genetic CETP deficiency is associated with increased P-HDL cholesterol, whereas CEPT transgenic mice display decreased P-HDL-cholesterol. The effects of CETP on atherogenesis are currently unpredictable, and they are likely to depend on the metabolic context. However, the presence of specific polymorphisms in the CETP gene may be of clinical importance, and CETP should be considered among the factors relevant for differentiation of dyslipidemic syndromes associated with susceptibility to and protection from atherosclerotic disease. PMID- 10536514 TI - [Diagnostic imaging in lumbago and sciatica]. AB - Low back pain and sciatica are among the most common medical problems in Western countries, affecting up to 80% of the population at some time during their lives. Plain radiography is still a sensitive method in degenerative spinal disease and for the identification of spondylolysis and destructions as well as transitional vertebra and other anomalies in the lumbosacral region. In lumbar disk herniation, CT and MR have higher sensitivity than lumbar myelography, and should be used as the primary imaging methods. Myelography is still the method of choice in lumbar spinal stenosis. Myelography should also be considered in patients with poor consistency between CT or MR findings and the clinical presentation. Postoperatively, MR is superior to CT and myelography for distinguishing between scar tissue and recurrent disk herniation. PMID- 10536515 TI - [Referral from general practice to diagnostic evaluation of dementia]. AB - The study objective was to describe the GP's referrals of patients to diagnostic evaluation of dementia and the GP's perception of the organization of this process++. The study is based on postal questionnaire mailed to all GPs in Denmark, spring 1998. Seventy-five percent of 3379 GPs answered the questionnaire. Seventy-one percent (1799) of the GPs had referred a patient to diagnostic evaluation of dementia within the last 12 months. Thirty-nine percent had referred to a geriatric psychiatric service, 36% to a neurologic service, 18% to a psychiatric service, 11% to a geriatric service, and 16% had referred to other services. Fifteen percent of the GPs had referred to two or more of the services. The study concludes that there is considerable regional variation as to where the GPs refer patients for diagnostic evaluation of dementia as well as to the GPs perception of the possibilities of referrals in Denmark. The implications are discussed. PMID- 10536516 TI - [Dietary guidelines on obesity at Danish pharmacies. Results of a 12-week course with a 1-year follow-up. Research Group on Human Nutrition, Frederiksberg]. AB - The increasing prevalence of obesity has created a need for alternative counselling sites. This retrospective study evaluates the results of a 12 week slimming course for obese subjects held at Danish pharmacies at one year follow up. Two hundred and sixty-nine obese (BMI > 25 kg/m2, 32 +/- 4.5 [mean +/- SD]) paid 550 Dkr each for a 12 weeks slimming course held at 19 Danish pharmacies with groups of 8-20 subjects each. The age was between 18 to 81 years, 259 were females. The course included eight sessions of 1 1/2 hour education in nutrition and physiology aiming for a dietary change toward a low-fat, high carbohydrate diet. Self-reported body weight was assessed on at the pharmacy scale before and after the course and again after three, six and 12 months follow-up. One hundred and ninety-one or 71% of subjects completed the 12 week slimming programme. The average weight loss was 5.3 and 6.2 kg among females and males, respectively. The weight loss maintenance was assessed at one year follow-up in 122 (45%) of the subjects who entered the course and was 4.0 and 6.7 kg in 118 females and four males, respectively. At one year follow-up 40 subjects (20%) of the subjects who completed the course had maintained a weight loss > 5 kg. In conclusion, the initial weight loss, and maintenance and drop-out rate are comparable with results from general practitioners and hospital out-patient clinics, but the costs are substantially lower. PMID- 10536517 TI - [Dementia and psychiatric service]. AB - The study objective was to describe the service provision in geriatric psychiatry in 1997 and the trends in admission patterns for the elderly to psychiatric hospitals in Denmark from 1988 to 1996. Information concerning admission pattern was obtained from the Danish Psychiatric Case Register. The information on the supply of geriatric psychiatric services was collected by a questionnaire to the individual geriatric psychiatric departments. All geriatric psychiatric departments in Denmark have been identified. The number of demented patients admitted to psychiatric hospitals decreased considerably as did the length of stay for demented patients admitted from 1988 to 1996. For all other diagnoses the number of admissions increased in the same period. Four counties out of 14 did not have a special unit for geriatric psychiatry. There were considerable geographical variations in supply as well as target groups in the counties that supplied geriatric psychiatric service. The unequal access to geriatric psychiatric services and variations in target groups underlines the need for a discussion of future directions for this service provision. PMID- 10536518 TI - [Cost analysis of inguinal hernia surgery in Denmark]. PMID- 10536519 TI - [Percutaneous transluminal embolization in arterial retroperitoneal traumatic hemorrhage]. AB - This study presents three cases of uncontrollable retroperitoneal arterial haemorrhage diagnosed by angiography and treated by transluminal embolization with coils. The haemorrhage originated in one patient from a branch of the internal iliac artery and in the other two patients from a renal artery. The circulation was stabilized in all three patients after transluminal coil embolization proximal to the arterial lesions. One patient died of multiorgan failure due to prolonged uncontrolled haemorrhage before embolization and one patient died as a consequence of low haemoglobin due to religiously based rejection of blood transfusion. Trauma patients with uncontrollable haemorrhage and retroperitoneal haematoma should be suspected of an arterial lesion and as early as possible be transferred to a center with experience in catheterisation and transluminal embolization. PMID- 10536520 TI - [Mathematical coupling and "regression to the mean". A statistical case history]. AB - "Regression to the mean" (RTM) is a statistical phenomenon originating from the random variation of variables rather than a true association between the variables. RTM is closely linked to mathematical coupling; unawareness of these phenomena may lead to misinterpretation of data. A study examining intracranial pressure before and after infusion of manitol is presented, in which mathematical coupling caused a highly significant (but spurious) correlation between the analysed data. The significance of RTM and mathematical coupling is demonstrated by analysis of a data set derived from random numbers. PMID- 10536522 TI - [Tuberculosis in Greenland and the rest of Denmark]. PMID- 10536521 TI - [Economic expenditures of different intervention regimes in the prevention of hip fractures]. PMID- 10536523 TI - [The benefit of new clinical information]. PMID- 10536524 TI - [Spontaneous remission of breast cancer]. PMID- 10536525 TI - [Angiotensin II receptor antagonists. Clinically significant differences]. PMID- 10536526 TI - Control of transgenesis in higher cells: the procell transposon Tn10 TetR mRNA has several major hairpins and can be unstable in eucells. AB - The TetR regulatory gene from the transposon Tn10 has some excellent characteristics for transgenic control in higher cells. However, we experienced severe problems with mRNA instability for this gene in eucells (CHO cells). This may be connected with the existence within the Tn10 TetR mRNA of several sizeable hairpins. They resemble canonical RNase E sites for mRNA destabilisation in procells and possibly also in eucells. Two of the hairpins also included sequences resembling eucell hnRNA polyadenylation or processing signals. The TetR counterpart from the plasmid RA1 appears to have less of the hairpin secondary structure; perhaps because of this, it did not present these mRNA instability problems in CHO cells, and it may prove a valuable alternative for transgene control in gene therapy and biotechnology. PMID- 10536527 TI - Grasping essential tremor. PMID- 10536528 TI - Fewer fractures with raloxifene. PMID- 10536529 TI - A new option for insomnia. PMID- 10536530 TI - "Thawing" frozen shoulders. PMID- 10536531 TI - High-protein diet may boost breast-cancer survival. PMID- 10536532 TI - Nocturnal noshing is no joke for some. PMID- 10536534 TI - Should age affect heart surgery decisions? PMID- 10536533 TI - Diuretics may forestall dementia. PMID- 10536535 TI - Walking away from heart disease. PMID- 10536536 TI - Echinacea: treatment or prevention? PMID- 10536537 TI - The placebo effect is alive and well. PMID- 10536538 TI - Heart rate variability, BIS and 'depth of anaesthesia'. PMID- 10536539 TI - 'The only man to have all his work done by Friday was Robinson Crusoe' (anon) PMID- 10536540 TI - Comparison of bispectral index, 95% spectral edge frequency and approximate entropy of the EEG, with changes in heart rate variability during induction of general anaesthesia. AB - We have compared bispectral index (BIS), 95% spectral edge frequency (SEF) and approximate entropy (ApEn) in 37 patients during induction and recovery from a short general anaesthetic. Heart rate variability (HRV) was also compared during induction only. These indices were noted at the start of induction, when a syringe held between the thumb and fingertips was dropped, at insertion of a laryngeal mask or tracheal tube (tube insertion), at incision, at the end of surgery, on return of the gag reflex and when the patient could follow a verbal command. When indices at the start of induction were compared with those at tube insertion, all four decreased significantly. BIS decreased from a mean of 95.38 (SEM 1.02) to 44.22 (1.05), mean SEF from 20.91 (1.19) to 14.14 (0.70) Hz, mean HRV from 37.1 (7.75) to 17.9 (3.6) bpm2 and ApEn from 0.90 (0.06) to 0.65 (0.04). Using logistic regression, the indices were compared both individually and in combination as to the power of distinguishing awake (at pre-induction) from asleep (at tube insertion) states. BIS had the best predictive power, with a sensitivity of 97.3%, specificity 94.4%, positive predictive value 94.7% and negative predictive value 97.1%. A combination of the indices conferred no additional predictive advantage. PMID- 10536541 TI - Comparison of bispectral EEG analysis and auditory evoked potentials for monitoring depth of anaesthesia during propofol anaesthesia. AB - We have compared the auditory evoked potential index (AEPIndex) and bispectral index (BIS) for monitoring depth of anaesthesia in spontaneously breathing surgical patients. Twenty patients (aged 17-49 yr) undergoing day surgery were anaesthetized with computer-controlled infusions of propofol. The mean (SD and range) of each measurement was determined during consciousness and unconsciousness and at specific times during the perioperative period. Mean values for AEPIndex during consciousness and unconsciousness were 74.5 (SD 14.7) 36.7 (7.1), respectively. BIS had mean values of 89.5 (SD 4.6) during consciousness and 48.8 (16.4) during unconsciousness. AEPIndex and BIS were greater during consciousness compared with during unconsciousness. The average awake values of AEPIndex were significantly higher than all average values during unconsciousness but this was not the case for BIS. BIS increased gradually during emergence from anaesthesia and may therefore be able to predict recovery of consciousness at the end of anaesthesia. AEPIndex was more able to detect the transition from unconsciousness to consciousness. PMID- 10536542 TI - Risk factors for cardiovascular death within 30 days after anaesthesia and urgent or emergency surgery: a nested case-control study. AB - The Oxford Record Linkage Study (an epidemiological database) was used to identify patients who died from a cardiovascular cause within 30 days of emergency or urgent surgery under general anaesthesia. Each case was paired with a control patient (matched for age within 10 yr of the patient, operation and consultant). Additional clinical information was sought from the patient's case notes. Cases and controls were compared for cardiovascular risk factors using conditional logistic regression analysis and a prognostic model was generated. Only one significant risk factor was identified in the final model: a history of cardiac failure (odds ratio 14.84; 95% confidence intervals 2.53-87.13; P = 0.003). Associations between a history of cerebrovascular accident or renal impairment and cardiovascular mortality were seen using univariate analysis but not after adjustment for confounding factors. PMID- 10536543 TI - Cardioventilatory coupling in atrial fibrillation. AB - Cardioventilatory coupling is an entrainment phenomena, distinct from respiratory sinus arrhythmia, whereby heart and breathing rhythms show temporal coherence. Coupling is commonly observed during rest, sleep and anaesthesia. Five graphical methods, each with different underlying mechanistic assumptions, have been suggested for studying this entrainment relationship: (a) time relationship between inspiration and a preceding heart beat, (b) time relationship between inspiration and a following heart beat, (c) phase of the cardiac cycle at which inspiration occurs, (d) phases of the ventilatory cycle at which heart beats occur and (e) 'relative phases' over multiple ventilatory cycles at which heart beats occur. In eight elderly human subjects with atrial fibrillation, breathing spontaneously during general anaesthesia, we recorded heart period and ventilatory time series and compared each of the graphical methods used for demonstration of coupling. We observed cardioventilatory coupling in seven of eight subjects. In each of these seven subjects, coupling was best described, both qualitatively and quantitatively, in terms of the relationship between inspiration and a preceding heart beat. The variation of the interval between inspiration and a preceding heart beat was less than for any other phase or time relationship. These data support a model of cardioventilatory coupling in which a heart beat triggers the onset of inspiration, rather than modulation of cardiac timing by ventilation or a phase relationship between the two systems. PMID- 10536544 TI - Haemodynamic changes associated with portal triad clamping are suppressed by prior hepatic pedicle infiltration with lidocaine in humans. AB - Portal triad clamping (PTC) reduces venous return of blood to the heart. However, the decrease in cardiac index (CI) is associated with an unexpected increase in mean arterial pressure (MAP) and the 40% increase in systemic vascular resistance is greater than anticipated in compensation for the 10% decrease in CI. We hypothesized that a reflex elicited in the peritoneum accounted for this unanticipated haemodynamic response. Twenty patients undergoing liver resection were allocated randomly to have hepatic pedicle infiltration before PTC with either lidocaine 200 mg or placebo. MAP was recorded, and plasma osmolality and plasma concentrations of vasopressin, epinephrine, norepinephrine, dopamine, renin and endothelin were measured. After PTC, MAP increased significantly in the placebo group but decreased significantly in the lidocaine group. Plasma concentrations of vasopressin, epinephrine and norepinephrine increased significantly in the placebo group. Plasma concentrations of vasopressin decreased significantly in the lidocaine group, while plasma concentrations of epinephrine and norepinephrine were unchanged. A subsequent study in eight patients found that neither haemodynamic nor hormonal changes associated with PTC in the placebo group were altered by administration of lidocaine 200 mg i.m. before PTC. PMID- 10536545 TI - Flumazenil antagonizes midazolam-induced airway narrowing during nasal breathing in humans. AB - We measured nasal resistance (Rn) while awake, during midazolam-induced sedation and after antagonism with flumazenil (n = 9). Nasal and oral airflow were measured. Rn was calculated by dividing the difference between maximal nasal mask and oropharyngeal pressures by inspiratory airflow at minimum pharyngeal pressure. During sedation, two subjects developed obstructive apnoeic events and four subjects had snoring events. Each apnoea was ended by mechanisms other than a change in breathing route. After antagonism with flumazenil, apnoeic and snoring events were abolished. Rn during midazolam sedation (median 1.46 (25th percentile 1.00, 75th 2.61) kPa litre-1 s) was significantly greater than before midazolam (0.29 (0.25, 0.48) kPa litre-1 s) and after flumazenil (0.41 (0.25, 0.58) kPa litre-1 s) (P < 0.01 in each subject). We conclude that midazolam increased Rn, sometimes leading to obstruction, and flumazenil abolished this increase in Rn. PMID- 10536546 TI - Laryngeal mask airway size selection in males and females: ease of insertion, oropharyngeal leak pressure, pharyngeal mucosal pressures and anatomical position. AB - We have compared ease of insertion, oropharyngeal leak pressure, directly measured pharyngeal mucosal pressure and anatomical position (assessed fibreoptically) for the size 4 and size 5 laryngeal mask airway (LMA) in 20 male and 20 female patients. Microchip pressure sensors were attached to the LMA at locations corresponding to the piriform fossa, hypopharynx, base of the tongue, lateral and posterior pharynx, and the oropharynx. Oropharyngeal leak pressure, mucosal pressure and fibreoptic position were recorded during inflation of the cuff from 0 to 30 ml in 10-ml increments. In males, oropharyngeal leak pressure over the inflation range was higher for size 5 (21 vs 17 cm H2O; P = 0.01); mucosal pressure over the inflation range was higher in the posterior pharynx for size 4 (7 vs 2 cm H2O; P = 0.007), and higher in the piriform fossa (8 vs 5 cm H2O; P = 0.003) and hypopharynx (9 vs 5 cm H2O; P = 0.003) for size 5. In females, oropharyngeal leak pressure over the inflation range was the same (21 vs 21 cm H2O), but mucosal pressure over the inflation range was higher in the piriform fossa (21 vs 8 cm H2O; P = 0.003) and posterior pharynx (4 vs 2 cm H2O; P = 0.004) for size 4, and higher in the lateral pharynx (5 vs 1 cm H2O; P = 0.01) and oropharynx (11 vs 5 cm H2O; P = 0.009) for size 5. The distribution of mucosal pressure was different for size 4 between males and females, but not for size 5. For both males and females, fibreoptic position was similar. We conclude that the size 5 LMA is optimal in males, but either size is suitable for females. The shape of the pharynx may be different between males and females. PMID- 10536547 TI - Direct measurement of mucosal pressures exerted by cuff and non-cuff portions of tracheal tubes with different cuff volumes and head and neck positions. AB - We measured directly mucosal pressures against the cuff and non-cuff portions of the tracheal tube in different head-neck positions and tested the reliability of calculated mucosal pressures, in vivo intracuff pressures and cuff volume as determinants of directly measured mucosal pressures. We studied 10 anaesthetized, paralysed adult patients. An 8.5-mm, high volume, low pressure PVC tracheal tube was used. Microchip sensors were attached to three cuff locations (anterior, lateral and posterior) and two non-cuff locations (anterior tip and anterior aspect of the tube, 5 cm proximal to the cuff). Directly measured mucosal pressures, in vivo intracuff pressures and calculated mucosal pressures (in vivo minus in vitro intracuff pressures) were determined after brief inflation (< 15 s) to 0, 5, 10 and 15 ml. In vivo intracuff pressures were then set at 30 mm Hg and the measurements repeated, first in the neutral position and then with the head-neck extended, flexed and rotated. Cuff mucosal pressures were highest anteriorly and lowest posteriorly. Non-cuff mucosal pressures did not vary with cuff volume and were approximately 15 mm Hg. Compared with the neutral position, in vivo intracuff pressures were higher in the rotated, extended and flexed positions. Compared with the neutral position, mucosal pressure increased on the anterior aspect of the tube in the flexed position by 22 mm Hg (P = 0.003), at the anterior tip in the extended position by 11 mm Hg (P = 0.002) and at the anterior tip (5 mm Hg, P = 0.05) and lateral aspect of the cuff (5 mm Hg, P = 0.03) in the rotated position. In vivo intracuff pressures and calculated mucosal pressures were moderate predictors of measured mucosal pressures; cuff volume was a poor predictor. We conclude that tracheal mucosal pressures were highest anteriorly, that non-cuff portions of the tube exerted substantial mucosal pressures and that the rotated position caused a greater increase in tracheal mucosal pressure than the extended or flexed position. Indirect methods of measuring mucosal pressure were of moderate predictive value. PMID- 10536548 TI - Placement of the intubating laryngeal mask is easier than the laryngeal mask during manual in-line neck stabilization. AB - We have compared in 25 patients ease of placement of the conventional and intubating laryngeal masks while the patient's head and neck were stabilized by a manual in-line method, in a randomized, crossover study. After induction of anaesthesia and neuromuscular block, the masks were placed in turn. Adequacy of ventilation and ease of placement (using a 10-cm visual analogue scale (VAS)) were assessed; time for placement between removal of the face mask and connection of the laryngeal mask to the breathing system was measured. Adequate ventilation was always obtained after placement of the intubating laryngeal mask, whereas ventilation was adequate in 22 of 25 patients after placement of the conventional laryngeal mask. Placement of the intubating laryngeal mask was significantly easier (P < 0.001; 95% confidence intervals (CI) for median difference 8-31 mm in VAS) and faster (P << 0.001; 95% CI for mean difference 3.2-6.2 s) than that of the conventional mask. PMID- 10536549 TI - Analgesic efficacy of paracetamol and diclofenac in children receiving PCA morphine. AB - We studied 80 children, aged 5-13 yr, who received PCA with morphine after appendicectomy using a standardized tracheal general anaesthetic. All patients received morphine 0.1 mg kg-1 before surgical incision and all had wound infiltration with bupivacaine 1 mg kg-1 at the end of surgery. Patients were allocated randomly to receive postoperative analgesia with PCA morphine alone, morphine plus diclofenac 1 mg kg-1, morphine plus paracetamol 15-20 mg kg-1 or morphine plus a combination of both diclofenac and paracetamol. Cumulative morphine consumption was significantly reduced by concurrent administration of diclofenac but no additive effect of paracetamol was demonstrable with the doses used in the study. Analgesia, as assessed by movement pain scoring, was significantly improved by the addition of diclofenac despite lower morphine consumption. Adverse effects and duration of PCA were comparable in the four groups. PMID- 10536550 TI - Effect of riluzole on acute pain and hyperalgesia in humans. AB - Riluzole modulates several transmitter systems which may be involved in nociception. Antinociceptive effects have been shown in animal studies, but there are no human data. Therefore, we have examined the acute analgesic effect of riluzole in a human model of inflammatory pain induced by a thermal injury on the distal leg (47 degrees C, 7 min, 12.5 cm2) in 20 healthy volunteers. Hyperalgesia to mechanical and heat stimuli were examined by von Frey hairs and thermodes. We used a randomized, double-blind, placebo-controlled design, and subjects received riluzole 100 mg or placebo for 2 days with a 14-day interval. The burns produced significant hyperalgesia, but riluzole had no acute analgesic effects in normal or hyperalgesic skin. PMID- 10536551 TI - Concentration-dependent inotropic effects of halothane, isoflurane and sevoflurane on rat ventricular myocytes. AB - We have described the concentration-dependent inotropic effects of halothane, isoflurane and sevoflurane on rat ventricular cells and investigated the role of the sarcoplasmic reticulum (SR) in these inotropic actions. Single ventricular myocytes, isolated from rat hearts, were stimulated electrically at 1 Hz and contractions recorded optically. Cells were exposed to a range of concentrations of halothane, isoflurane or sevoflurane for a period of 1 min to determine the concentration-dependency of their inotropic actions. For each anaesthetic, the peak negative inotropic action was determined early during an exposure, and sustained negative inotropic action was measured at steady-state just before wash off. In some experiments, cells were equilibrated with ryanodine 1 mumol litre-1 to investigate the role of the SR in these intropic effects. Halothane caused a concentration-dependent initial increase in contractions (to mean 130 (SEM 28)% at 10 mmol litre-1) followed by rapid onset of a negative inotropic effect (K0.5 0.34 mmol litre-1 for peak effect; K0.5 0.46 mmol litre-1 for sustained effect). Exposure to isoflurane induced a small potentiation of contractions in some cells, followed by a concentration-dependent decrease in contraction in all cells (K0.5 0.85 mmol litre-1 for peak effect; K0.5 1.92 mmol litre-1 for sustained effect); contractions recovered partially during a 1-min exposure. On wash-off, contractions were increased transiently above control. Sevoflurane caused a large initial decrease in contraction which then returned rapidly towards control (K0.5 0.2 mmol litre-1 for peak effect; K0.5 2.57 mmol litre-1 for sustained effect). In common with isoflurane, removal of sevoflurane caused a transient increase in contractions above control. After exposure to ryanodine, the positive inotropic effects of halothane and isoflurane did not occur, and recovery of contractions during exposure to isoflurane and sevoflurane was abolished as was the transient increase in contractions seen on wash-off, indicating that these effects were mediated via the SR. Halothane had the most potent sustained negative inotropic effect but there was little difference between the negative inotropic effects of isoflurane and sevoflurane at clinically relevant concentrations. At higher concentrations, sevoflurane caused a less potent negative inotropic effect than isoflurane. The SR plays a major role in the effects of all three anaesthetics. One possible mechanism underlying the initial potentiation of contraction by halothane (and isoflurane) may be sensitization of the Ca(2+)-induced Ca(2+) release process of the SR. PMID- 10536552 TI - Anaesthetic effects of pregnanolone in combination with allopregnanolone, thiopental, hexobarbital and flurazepam: an EEG study in the rat. AB - The anaesthetic interactions of the steroid, 3 alpha-hydroxy-5 beta-pregnan-20 one, in male rats were investigated in different fixed binary combinations with the steroid allopregnanolone (3 alpha-hydroxy-5 alpha-pregnan-20-one), two barbiturates (thiopental and hexobarbital) and the benzodiazepine, flurazepam. Anaesthetic effects were determined using an EEG threshold method. Interactions were assessed using an isobolographic method. The interaction between the two steroids, pregnanolone and allopregnanolone, showed an anaesthetic effect significantly less than additive (antagonistic). The interactions between pregnanolone and the two barbiturates and the benzodiazepine showed an anaesthetic effect significantly greater than additive (potentiation) in all tests performed. These results could be explained by a pharmacodynamic interaction at the hypothetical GABA-benzodiazepine-barbiturate-steroid complex in the CNS. PMID- 10536553 TI - Mucosal tissue oxygenation of the porcine jejunum during normothermic cardiopulmonary bypass. AB - Cardiopulmonary bypass (CPB) has been associated with intestinal tissue hypoxia, but direct measurements of mucosal oxygenation have not been performed. In anaesthetized pigs, jejunal mucosal oxygen tension and microvascular haemoglobin oxygen saturation were measured by a Clark-type electrode and tissue reflectance spectrophotometry. In pigs, normothermic CPB with systemic oxygen transport equivalent to baseline values was performed. In control animals, mucosal oxygen tension and mucosal haemoglobin oxygen saturation were mean 5.01 (SD 1.08) kPa and 38.0 (2.3)%, respectively. CPB was associated with a decrease in mucosal oxygen tension to 2.26 (1.21) kPa, decrease in mucosal microvascular haemoglobin oxygen saturation to 26.0 (3.9)% and appearance of oscillations in mucosal microvascular haemoglobin oxygen saturation. With CPB, arterial lactate concentrations increased from 1.77 (1.37) to 3.52 (1.58) mmol litre-1, but transvisceral lactate and splanchnic venous-arterial carbon dioxide tension gradients remained unchanged. Our results support the concept that CPB is associated with diminished oxygenation of intestinal mucosa that is probably caused by regional redistribution. PMID- 10536554 TI - Anaesthetic management of epidermolysis bullosa. PMID- 10536555 TI - Gastric pressure during emergency caesarean section under general anaesthesia. AB - Gastric pressure and volume were measured in 20 pregnant women during emergency Caesarean section under general anaesthesia with neuromuscular block. Mean gastric pressure was 11 (range 4-19) mm Hg and we can predict that 99% of women undergoing emergency Caesarean section with neuromuscular block are likely to have gastric pressures of less than 25 mm Hg (mean + 3 SD). This has implications for the amount of cricoid pressure required during induction of anaesthesia. Gastric pressure increased during delivery to 19 mm Hg and fundal pressure caused a gastric pressure of 65 mm Hg in one woman. Gastric pressure decreased significantly after delivery (P < 0.001) to 8 mm Hg. Although we measured large gastric volumes (mean 112 (range 20-350) ml), there was no correlation between gastric volume and gastric pressure. PMID- 10536556 TI - Spontaneous or neostigmine-induced recovery after maintenance of neuromuscular block with Org 9487 (rapacuronium) or rocuronium following an initial dose of Org 9487. AB - We have examined spontaneous and neostigmine-induced recovery after an initial dose of Org 9487 1.5 mg kg-1 followed by three repeat doses of Org 9487, a 30-min infusion of Org 9487 or two incremental doses of rocuronium. Mean clinical duration after incremental doses of Org 9487 0.5 mg kg-1 increased from 12.3 (SD 3.4) min to 14.0 (4.0) and 15.9 (5.9) min (P < 0.01), and after rocuronium from 14.4 (5.2) min to 19.2 (5.9) min (P < 0.01). Times for spontaneous recovery from a T1 of 25% to a TOF ratio of 0.8 after the last bolus dose of Org 9487 and after a 30-min infusion were 72.4 (16.5) and 66.1 (26.9) min compared with 36.7 (15.8) min in the group receiving reocuronium. These times were significantly reduced to 9.9 (4.5), 8.6 (6.1) and 5.7 (2.5) min, respectively, after neostigmine administration at a T1 of 25% (P < 0.05). We conclude that administration of Org 9487 by repeat bolus doses or infusion was associated with slow spontaneous recovery but neostigmine administration resulted in adequate recovery in less than 10 min. PMID- 10536558 TI - Anaesthesia for strabismus surgery: a regional survey. AB - An increase in the demand by local surgeons for neuromuscular block during strabismus surgery, and the forced duction test in particular, led us to review the literature and conduct a regional survey of anaesthetic techniques used. A questionnaire was distributed to 379 anaesthetists in the region and 264 responses were received. The results demonstrated that 55% of paediatric patients and 66% of adult patients may have been operated on under suboptimal conditions; residual tone may have been present in the extraocular muscles during forced duction testing and strabismus correction. PMID- 10536557 TI - Effect of rocuronium compared with succinylcholine on intraocular pressure during rapid sequence induction of anaesthesia. AB - We have compared the effect of rocuronium and succinylcholine on intraocular pressure (IOP) during rapid sequence induction of anaesthesia using propofol and fentanyl, in a randomized double-blind study. We studied 30 adult patients, allocated to one of two groups. Anaesthesia was induced with fentanyl 2 micrograms kg-1 and propofol until loss of verbal response. This was followed by succinylcholine 1.5 mg kg-1 (group S; n = 15) or rocuronium 0.9 mg kg-1 (group R; n = 15). Laryngoscopy was performed 60 s later. IOP, mean arterial pressure (MAP) and heart rate (HR) were measured before induction, immediately before intubation and every minute after intubation for 5 min. A Keeler Pulsair air impulse tonometer was used to measure IOP and the mean of two readings obtained in the right eye at each measurement time was recorded. Intubating conditions were evaluated according to a simple scoring system. IOP in the succinylcholine group was significantly greater than that in the rocuronium group (mean 21.6 (SEM 1.4) mm Hg vs 13.3 (1.4) mm Hg; P < 0.001). Intubating conditions were equally good in both groups. We conclude that with rapid sequence induction of anaesthesia using propofol and fentanyl, rocuronium did not cause as great an increase in IOP as succinylcholine and may be an alternative in open eye injury cases. PMID- 10536559 TI - Waste anaesthetic gases induce sister chromatid exchanges in lymphocytes of operating room personnel. AB - Genotoxicity related to waste anaesthetic gas exposure is controversial. We have investigated the frequency of sister chromatid exchanges in peripheral lymphocytes of operating room personnel exposed to trace concentrations of isoflurane and nitrous oxide. Occupational exposure was recorded using a direct reading instrument. Frequencies of sister chromatid exchanges were measured in lymphocyte cultures of 27 non-smokers working in the operating room and 27 non smoking controls. Personnel were exposed to an 8-h time-weighted average of nitrous oxide 11.8 ppm and isoflurane 0.5 ppm. After exposure, sister chromatid exchange frequency was increased significantly (mean 9.0 (SD 1.3) vs 8.0 (1.4) in exposed and control personnel, respectively) (P < 0.05). We conclude that exposure to even trace concentrations of waste anaesthetic gases may cause genetic damage comparable with smoking 11-20 cigarettes per day. PMID- 10536561 TI - Failure to prevent an anaphylactic reaction to a second neuromuscular blocking drug during anaesthesia. AB - Skin testing is used widely to determine the drug responsible for an anaphylactic reaction during anaesthesia. When a neuromuscular blocking drug in incriminated as the cause of a reaction, it is usual for neuromuscular blocking drugs which do not produce positive skin tests to be considered safe for subsequent use during anaesthesia. We describe three patients in whom false negative skin tests led to a second severe anaphylactic reaction to another neuromuscular blocking drug. PMID- 10536560 TI - Differential effects of isoflurane and i.v. anaesthetic agents on metabolism of alveolar type II cells. AB - Alveolar type II (ATII) cells perform many important functions within the lung, including surfactant metabolism. We have investigated the effects of isoflurane and different i.v. anaesthetics on cell metabolism in primary cultures of rat ATII cells. Biosynthesis of phosphatidylcholine, the main phospholipid component of surfactant, was decreased in cells exposed to isoflurane in a dose- and time related manner. This effect was fully reversible within 2 h after isoflurane removal. Lactate dehydrogenase (LDH) release, an index of cell damage, was increased with isoflurane exposure (1%) over a long incubation period (8-12 h). Enhanced lactate production, reflecting an increase in glycolytic metabolism, was also observed in isoflurane exposed cells. In contrast, metabolism of ATII cells was moderately affected by i.v. anaesthetics. Our data suggest differential metabolism of alveolar homeostasis depending on the anaesthetic agent used. PMID- 10536562 TI - Bladder exstrophy in a neonate at risk of transient myasthenia gravis: a role for remifentanil and epidural analgesia. AB - Infants born to mothers with myasthenia gravis may exhibit a transient form of the disease, with similar sensitivity to non-depolarizing neuromuscular blocking drugs. We report the case of an infant at risk who required major surgery when 48 h old for closure of bladder exstrophy. A combined epidural-general anaesthetic technique, with remifentanil supplementation, enabled us to avoid unnecessary neuromuscular blocking drugs and prolonged intensive care, which had been anticipated. The potential benefits of remifentanil and epidural analgesia in neonates are discussed. PMID- 10536563 TI - Pulmonary oedema after peribulbar block. AB - Local anaesthesia is now preferred for cataract surgery. Respiratory distress caused by pulmonary oedema is a rare, if well recognized, complication of the technique of retrobulbar block. We report this complication after the increasingly favoured peribulbar approach. PMID- 10536564 TI - Comparison of a new piezoelectric train-of-four neuromuscular monitor, the ParaGraph, and the Relaxometer mechanomyograph. AB - The ParaGraph is a new device for monitoring neuromuscular function using a piezoelectric motion sensor. In 20 patients, monitoring of neuromuscular block produced by cisatracurium 0.1 mg kg-1 was compared using the ParaGraph and a Relaxometer 2 mechanomyograph. The ParaGraph was quick to set up, and easy to operate and interpret. There were no significant differences in the time to 100% depression of T1/T0, time to 25% recovery of T1/T0 or time to recovery of T1/T0 from 25% to 75%, measured by the two monitors. When the difference between the two monitors was plotted against the average of the two measurements, the limits of agreement for T1/T0 (-42.95, +53.98%) and the train-of-four ratio, T4/T1 ( 0.28, +0.21) were too wide to allow the values given by the two monitors for individual patients to be used interchangeably. PMID- 10536565 TI - Autologous transfusion, 3 yr on--what is new? What has happened? Second Consensus Conference on Autologous Transfusion, held at the Royal College of Physicians, Edinburgh, November 11, 1998. PMID- 10536566 TI - Mortality of medical oncological patients admitted to intensive care. PMID- 10536567 TI - Sudden airway obstruction following inhalation drug abuse. PMID- 10536568 TI - Anaesthesia for caesarean section in severe pulmonary hypertension. PMID- 10536569 TI - Anaesthesia for caesarean section in severe pulmonary hypertension. PMID- 10536570 TI - The intubating laryngeal mask and distorted airway anatomy. PMID- 10536571 TI - The intubating laryngeal mask and distorted airway anatomy. PMID- 10536572 TI - Effect of nitrous oxide on dyspnoea. PMID- 10536573 TI - Inadvertent inhalation anaesthesia during surgery under retrobulbar eye block. PMID- 10536574 TI - Postoperative cognitive dysfunction in the elderly surgical patient. PMID- 10536575 TI - Logbooks and the 'millennium bug'. PMID- 10536576 TI - Resident and faculty perceptions of continuity practice experience in two teaching models. PMID- 10536577 TI - A comparative study of problem-based and conventional undergraduate curricula in preparing students for graduate medical education. PMID- 10536578 TI - The effects of basic science pathway on USMLE Step 1 scores. PMID- 10536579 TI - Measuring the promotion of thinking during precepting encounters in outpatient settings. PMID- 10536580 TI - Can diagnostic semantic competence be assessed from the medical record? PMID- 10536581 TI - The effects of immediate online feedback upon diagnostic performance. PMID- 10536582 TI - The dependability of students' ratings of preceptors. PMID- 10536583 TI - A comparison of student evaluations and faculty peer evaluations of faculty lectures. PMID- 10536584 TI - Factorial validation of an educational framework using residents' evaluations of clinician-educators. PMID- 10536585 TI - Recertification: is there a link between take-home and proctored examinations? PMID- 10536586 TI - Physician outcomes and implications for planning an intensive educational experience on attention-deficit hyperactivity disorder. PMID- 10536587 TI - A study of the factors that influence physicians' commitments to change their practices using learning diaries. PMID- 10536588 TI - Does sex make a difference? Sometimes it does and sometimes it doesn't. PMID- 10536589 TI - Validity of MCAT scores for predicting clerkship performance of medical students grouped by sex and ethnicity. PMID- 10536590 TI - Identifying students at risk for poor performance on the USMLE Step 2. PMID- 10536591 TI - Modeling the effects of security breaches on students' performances on a large scale standardized patient examination. PMID- 10536592 TI - Assessing the impacts of intra-site and inter-site checklist recording discrepancies on the reliability of scores obtained in a nationally administered standardized patient examination. PMID- 10536593 TI - Using tagged items to detect threats to security in a nationally administered standardized patient examination. PMID- 10536594 TI - A retrospective review of medical school admission files of academically at-risk matriculants. PMID- 10536595 TI - "Watch me do it": three trajectories toward medical school admission in a post baccalaureate, premedical program. PMID- 10536596 TI - Academic and noncognitive factors affecting placement of medical school applicants on an alternate list. PMID- 10536597 TI - Outcomes assessment of a faculty development program in medicine and pediatrics. PMID- 10536598 TI - Academic careers in medical education: perceptions of the effects of a faculty development program. PMID- 10536599 TI - Improvement of tutorial skills: an effect of workshops or experience? PMID- 10536601 TI - Incoming primary care interns' attitudes toward and knowledge of managed care. PMID- 10536600 TI - A brief instrument to measure attitudes of medical students toward changes in the health care system. PMID- 10536602 TI - Evaluating the serial migration of an existing required course to the World Wide Web. PMID- 10536603 TI - Do virtual computer models hinder anatomy learning? PMID- 10536604 TI - The teaching of chest auscultation in U.S. internal medicine and family practice medicine residencies. PMID- 10536605 TI - Effectiveness of a computer-based system to teach bedside cardiology. PMID- 10536606 TI - Meeting the challenge of MSOP: comprehensive measurement of profiles of student characteristics across medical schools. PMID- 10536607 TI - Patterns of medical student abuse during the internal medicine clerkship: perspectives of students at 11 medical schools. PMID- 10536608 TI - Education in end-of-life care during medical school and residency training. PMID- 10536609 TI - Community-based palliative care education: can it improve care of the terminally ill? PMID- 10536610 TI - Can resident evaluations demonstrate increases in residents' skills over time? PMID- 10536611 TI - Development of a rating scale to evaluate written communication skills of residents. PMID- 10536612 TI - Dissecting the medical training-to-practice continuum: factors associated with choosing in-state graduate medical education. PMID- 10536613 TI - In-training evaluation during an internal medicine clerkship. PMID- 10536614 TI - Influences on specialty choice: are they different for switchers and non switchers? PMID- 10536615 TI - What do we mean by "relevance"? A clinical and rhetorical definition with implications for teaching and learning the case-presentation format. PMID- 10536616 TI - Predictors of student self-assessment accuracy during a clinical performance exam: comparisons between over-estimators and under-estimators of SP-evaluated performance. PMID- 10536617 TI - Effect of multiple standardized patients on case and examination means and passing rates. PMID- 10536618 TI - OSCE performance evaluations made by standardized patients: comparing checklist and global rating scores. PMID- 10536619 TI - Why did I miss the diagnosis? Some cognitive explanations and educational implications. PMID- 10536620 TI - Learning by tutoring. PMID- 10536621 TI - An Internet-based law and medicine curriculum for residents. PMID- 10536622 TI - Clinical service-line structures can better carry out the missions of traditional clinical departments. PMID- 10536623 TI - Violence against people with disabilities: barriers to diagnosis, prevention, and treatment. PMID- 10536624 TI - The liberal arts physician. AB - The United States is in the midst of the second revolution in American health care to occur during this century, as Kenneth Ludmerer makes clear in his book Time to Heal: American Medical Education from the Turn of the Century to the Era of Managed Care. The "Flexnerian revolution" eventually led to the closing of a third of the medical schools. Although such closures are not likely this time, familiar arrangements are collapsing, without a clear picture of the shape of things to come. Whatever the outcome of the current revolution, well-trained physicians will be needed to care for the sick. Academic medical centers truly are at risk and increasingly require public support to flourish or even to survive, but medical schools and their teaching hospitals must demonstrate that they deserve this support. These institutions have responded by focusing on the business aspects of medicine, perhaps to the detriment of medical education. Lost in this focus is teaching time, and perhaps even more important, the time for mentoring. Often lacking too is a clear vision of the preparation needed by the student to practice medicine successfully in the future: different specialty mixes, interdisciplinary group practice; vastly increased use of information technologies, and overwhelming amounts of relevant and interrelated information. Yet the answer is the same as it was 75 years ago when Yale introduced the first radical medical curricular reform--the "liberal arts physician," trained in science, the values of medicine, and particularly for uncertainly and with the capacity to adapt. PMID- 10536625 TI - Crushing the commercial spirit in academic medicine: a crusade that failed. AB - The struggle between academic values and the practice opportunities in clinical medicine has continued throughout the present century. The reformers who prevailed in bringing clinical teaching into the university as a full-time occupation were persuaded that only university ideals--academic rigor, high professionalism, and full-time service in teaching and research--could create the kind of environment in which clinical science and effective clinical teaching could flourish. Their victory was never complete, and much of America's clinical establishment resisted the change, arguing that it was not commercial gain but concerns over teaching medicine in a narrowly academic enclave that motivated them. For the first two thirds of the century, the commercial spirit in academic medicine, while never completely crushed, gave way to an academic ethos that honored academic recognition and research honors over making money. Events of the past 30 years have reawakened the commercial spirit with a vengeance. In the years since Medicare, managed care, and HMOs have become prominent, faculty practice has become a principal means of maintaining teaching hospitals, high professional salaries, and medical teaching. In the present crisis, the author believes, only an unprecedented, all-out effort on the part of medical faculties and their allies to separate out medical education from other health care concerns and secure strong support from government offers any long-range hope for success. PMID- 10536627 TI - Transformation of medical students' education: work in progress and continuing challenges. AB - In his book Time to Heal: American Medical Education from the Turn of the Century to the Era of Managed Care, Ludmerer expresses concern about the erosion of the environment in which medical students and residents learn the clinical skills, attitudes, and behaviors that they will need to practice high-quality medicine. Importantly, while he attributes the erosion of the clinical environment largely to the impact of managed care, he also places some responsibility within academic medicine itself, primarily the redirection of the clinical faculty's efforts away from traditional academic pursuits to the generation of clinical revenues. The Association of American Medical Colleges has information about the kinds of changes already occurring. In the preclinical curriculum, schools have introduced a wide range of new courses and topics, and there is more attention on professionalism and values. Schools are making fundamental changes in the design and conduct of the curriculum, primarily by adopting more integrated (non departmental) approaches to course design and management. The clinical curriculum is changing primarily through the greatly expanded use of ambulatory care sites, and medical schools are developing new approaches to managing dispersed and varied instruction. Also, faculty are paying more attention to the role of residents as teachers and role models. These changes speak well for medical education. Nonetheless, substantial and sustained work remains to be done despite the present uncertainty about the future of academic medical centers. This work is essential--a challenge that the leaders of academic medicine must not fail. PMID- 10536626 TI - Time to heal medical education? AB - In Time to Heal: American Medical Education from the Turn of the Century to the Era of Managed Care, Kenneth Ludmerer presents a penetrating narrative and analytic account of the daunting economic, organizational, social, and value problems facing medical education that are threatening its excellence and its sense of purpose. The new conditions have the potential not only to seriously undermine the scientific and clinical quality and vibrancy of the education that medical students and housestaff receive, but also to subvert the development of core elements of a "Samaritan" concern for the care, the suffering, and the well being of patients and of dedication to the public good. American medical schools have a long tradition of trying to improve medical education through periodic curriculum reforms. Since the famous and influential Flexner report in 1910, at least 24 other major reports advocating such reform have been issued, and they have been strikingly similar in their prescriptions. The resulting modifications have, however, been "reform without change" because of medical educators' failure to address the basic social, organizational, and financial problems with which medical schools have been progressively confronted. Ludmerer considers that today's problems, however formidable they may be, are not as profound as those that faced medical educators a century ago--and that there is time and opportunity for visionaries and leaders to act. This may be true but only if they can properly diagnose the present problems of medical education, amply understand their roots, identify effective means to remedy them, and galvanize medical educators to make the systemic, institutional changes, locally and nationally, that are called for. PMID- 10536628 TI - Understanding the value added to clinical care by educational activities. Value of Education Research Group. AB - In an era of competition in health care delivery, those who pay for care are interested in supporting primarily those activities that add value to the clinical enterprise. The authors report on their 1998 project to develop a conceptual model for assessing the value added to clinical care by educational activities. Through interviews, nine key stakeholders in patient care identified five ways in which education might add value to clinical care: education can foster higher-quality care, improve work satisfaction of clinicians, have trainees provide direct clinical services, improve recruitment and retention of clinicians, and contribute to the future of health care. With this as a base, an expert panel of 13 clinical educators and investigators defined six perspectives from which the value of education in clinical care might be studied: the perspectives of health-care-oriented organizations, clinician-teachers, patients, education organizations, learners, and the community. The panel adapted an existing model to create the "Education Compass" to portray education's effects on clinical care, and developed a new set of definitions and research questions for each of the four major aspects of the model (clinical, functional, satisfaction, and cost). Working groups next drafted proposals to address empirically those questions, which were critiqued at a national conference on the topic of education's value in clinical care. The next step is to use the methods developed in this project to empirically assess the value added by educational activities to clinical care. PMID- 10536629 TI - Standards and reliability in evaluation: when rules of thumb don't apply. AB - The purpose of this paper is to identify situations in which two rules of thumb in evaluation do not apply. The first rule is that all standards should be absolute. When selection decisions are being made or when classroom tests are given, however, relative standards may be better. The second rule of thumb is that every test should have a reliability of .80 or better. Depending on the circumstances, though, the standard error of measurement, the consistency of pass/fail classifications, and the domain-referenced reliability coefficients may be better indicators of reproducibility. PMID- 10536630 TI - Avoiding the great train wreck: standardizing the architecture for online curricula. AB - The advent of the World Wide Web and related technologies has encouraged medical schools to generate a wealth of online curricular materials. Unfortunately, the diversity and abundance of user-friendly authoring tools have enabled individual instructors within and among medical schools to proliferate educational resources with often widely dissimilar architectures. Because there are no universal standards for educational technology, the resulting information islands cannot be easily managed, reused, or shared with other educators and institutions. Thoughtful planning and reuse of online educational materials could allow instructors to present subject matter more consistently across the curriculum, discover new teaching resources, easily create new materials, reduce unnecessary duplication, and better integrate the basic and clinical sciences. To make this possible, standardized, object-oriented architecture is needed. While several excellent HTML authoring tools and integrated courseware packages are available, differences in their technologies and methods impede inter-operability between the various products and the materials they create. Through the National Learning Infrastructure Initiative, the Instructional Management Systems (IMS) specification has emerged as a global standard for online teaching and learning environments. This proposed standard defines a common architecture for content management and delivery. PMID- 10536631 TI - Words, art, body and memory: readings from a writer. AB - This essay considers the consequences of childhood experiences with family illness on future adult sensibilities. Novelist and memoirist Mary Gordon describes her father's early death from heart disease and her childhood responsibilities toward her mother who was chronically ill with polio. She examines the relations between the ill body and the rituals and teachings of Catholicism and, by implication, all religious imagings of the body. By detailing her current caregiving responsibilities toward her mother, who is now homebound with Alzheimer disease, she scrutinizes the responses of the healthy toward the ill. She cites passages from her own novels--The Other Side, The Shadow Man, and Spending--in which characters' bodies fail and sicken. By examining these retrospective and prospective experiences with other people's ailments, Gordon exhorts medical students and doctors to tend to the bodies in their care with skill, with vision, and with words. She joins fellow writer Joseph Conrad in his task: "by the power of the written word to make you hear, to make you feel; it is, before all, to make you see." PMID- 10536632 TI - Women's health research: progress and future directions. PMID- 10536633 TI - How doctors learn: physicians' self-directed learning episodes. AB - PURPOSE: To qualitatively examine the self-directed learning activities of physicians in light of several lines of research on how doctors learn. METHOD: Under the auspices of the Royal College of Physicians and Surgeons of Canada, the author elicited from physicians narratives about past learning experiences. He analyzed the narratives (1) seeking themes among the doctors' approaches and (2) examining those themes in light of the existing literature. RESULTS: The 32 physicians interviewed described learning experiences, confirming earlier research that two varieties of problems (specific and general) precipitate learning and that learning episodes follow definite stages: scanning for problems, deciding whether to pursue the learning task, acquiring new knowledge and skill, and gaining experience with what has been learned. The latter three stages have been described previously and are expanded upon here. CONCLUSION: This study produced an integrated and elaborated theory of learning in clinical practice with implications for both the education of physicians in training and physicians' continuing professional development. In particular, the theory points to problem areas in teaching medical students and residents to learn in clinical practice, and in matching the learning needs of physicians to organized continuing medical education activities. PMID- 10536634 TI - The Patient Findings Questionnaire: one solution to an important standardized patient examination problem. AB - PURPOSE: To describe the Patient Findings Questionnaire (PFQ) and compare its scores and pass/fail decisions with those obtained from standardized patient (SP) examination checklists. METHOD: Checklists and PFQs were used to assess data acquisition by 790 second-year medical students. PFQs were composed of multiple choice items designed to determine whether examines had acquired key historical patient information. RESULTS: At the item level, the two measurement methods yielded the same decisions about data acquisition on 88% of observed occasions. Most discrepancies (74%) involved SPs rating examinees as having elicited information when the examinee was unable to answer the associated PFQ item. At the test level, the two instruments yielded the same pass/fail decision on a large majority of occasions. CONCLUSIONS: The PFQ and checklist yielded similar data acquisition scores and decisions at the item and test levels. Replacement of the checklist with the PFQ should result in examinees' behaving in a way more consistent with recommended interviewing practices. PMID- 10536635 TI - Evaluating the usefulness of computerized adaptive testing for medical in-course assessment. AB - PURPOSE: This study investigated the feasibility of converting an existing computer-administered, in-course internal medicine test to an adaptive format. METHOD: A 200-item internal medicine extended matching test was used for this research. Parameters were estimated with commercially available software with responses from 621 examinees. A specially developed simulation program was used to retrospectively estimate the efficiency of the computer-adaptive exam format. RESULTS: It was found that the average test length could be shortened by almost half with measurement precision approximately equal to that of the full 200-item paper-and-pencil test. However, computer-adaptive testing with this item bank provided little advantage for examinees at the upper end of the ability continuum. An examination of classical item statistics and IRT item statistics suggested that adding more difficult items might extend the advantage to this group of examinees. CONCLUSIONS: Medical item banks presently used for incourse assessment might be advantageously employed in adaptive testing. However, it is important to evaluate the match between the items and the measurement objective of the test before implementing this format. PMID- 10536636 TI - OSCE checklists do not capture increasing levels of expertise. AB - PURPOSE: To evaluate the effectiveness of binary content checklists in measuring increasing levels of clinical competence. METHOD: Fourteen clinical clerks, 14 family practice residents, and 14 family physicians participated in two 15-minute standardized patient interviews. An examiner rated each participant's performance using a binary content checklist and a global process rating. The participants provided a diagnosis two minutes into and at the end of the interview. RESULTS: On global scales, the experienced clinicians scored significantly better than did the residents and clerks, but on checklists, the experienced clinicians scored significantly worse than did the residents and clerks. Diagnostic accuracy increased for all groups between the two-minute and 15-minute marks without significant differences between the groups. CONCLUSION: These findings are consistent with the hypothesis that binary checklists may not be valid measures of increasing clinical competence. PMID- 10536637 TI - Undergraduate university students' views of the use of animals in biomedical research. AB - PURPOSE: To investigate the influences of gender, discipline, and level on undergraduate students' views of the use of animals in research. METHOD: In 1998, 888 university undergraduate students from six different programs were surveyed at Uppsala University for their views of animal use in biomedical research. Statistical analysis involved chi-square tests. RESULTS: Most students found animal use morally acceptable and believed it plays a significant role in the treatment of human diseases. Engineering, law, and medical students were the most supportive, whereas pre-school-teaching students were the least supportive. Men were more supportive than were women. Sixth-term medical and nursing students had a more positive view than did their inexperienced first-term peers. CONCLUSION: The results of this study show that university students, who will be tomorrow's decision makers, are likely to continue supporting the use of animals in biomedical research and teaching. PMID- 10536638 TI - Attitudes of faculty, housestaff, and medical students toward clinical practice guidelines. AB - PURPOSE: To examine attitudes of faculty, housestaff, and medical students toward clinical practice guidelines. METHOD: In a 1997 cross-sectional survey, a two part, 26-item, self-administered questionnaire was mailed to all faculty, housestaff, and medical students in the department of internal medicine at Case Western Reserve University School of Medicine. The questionnaire asked for demographic information and attitudes toward clinical guidelines. RESULTS: Of 379 persons surveyed, 254 (67%) returned usable questionnaires: 56% of the medical students, 70% of the housestaff, and 73% of the full-time faculty. Medical students reported learning about guidelines predominantly during clerkships in internal medicine (71%) and pediatrics (68%). Overall, the respondents agreed most strongly that guidelines are "useful for the care of common problems," and least strongly that guidelines are "difficult to apply to individual patients" and "reduce physician options in patient care." Faculty were more likely to consider guidelines a "good educational tool" and less likely than were medical students and housestaff to agree that they promote "cookbook medicine." Of 11 influences on clinical decision making, the three groups together rated practice guidelines eighth or ninth. The use of guidelines for academic investigations was rated most appropriate, overall. In terms of their appropriateness, faculty consistently rated the use of guidelines more favorably except for use in malpractice suits. CONCLUSION: Faculty, housestaff, and medical students have significantly different perceptions of and attitudes toward clinical practice guidelines. Further studies are needed to explain the reasons for these differences. Considerable education and involvement must occur at all levels for practice guidelines to be successfully implemented and understood. PMID- 10536639 TI - Educational technology to facilitate medical students' learning: background paper 2 of the medical school objectives project. AB - The present article is the second in a series of Background Papers prepared as part of the AAMC's Medical School Objectives Project (MSOP). This report provides information about and insight into U.S. medical schools' use of educational technology in 1998. The authors define educational technology as the use of information technology to facilitate students' learning. They note that in the last two decades, a number of reports have recommended that medical schools incorporate educational technology into their teaching programs. To gain insight into the effects of these recommendations, particularly those of the ACME-TRI Report in 1992, the authors analyzed the responses of administrators at 125 U.S. medical schools to relevant items of the 1997-98 Liaison Committee on Medical Education Part II Medical School Questionnaire and students' responses to relevant items of the 1998 AAMC Medical Student Graduation Questionnaire. In addition, site visits were made to six medical schools believed to be among the more advanced ones in the use of educational technology, to see what was happening on the "cutting edge" of educational technology applications. Data from 20 other schools were also used. The authors found that by 1998, medical schools as a group had made limited progress in accomplishing the recommended educational technology goals, and that there was a much greater use of such technology in basic sciences courses than in clinical clerkships. However, great variability existed across schools in the use of such technology and in the administrative arrangements for it. They observe that the use of educational technology in medical schools is increasing rapidly, and recommend that each school develop a strategic approach that will guarantee that it can meet the future educational technology needs of its students. PMID- 10536640 TI - Neuronal loss in the brainstem and cerebellum--part of the normal aging process? A morphometric study of the vermis cerebelli and inferior olivary nucleus. AB - Based on the known age-related loss of Purkinje cells (PC) in the cerebellum, this study focuses on whether a marked loss of PC occurs in individuals of very high age. The inferior olive, which is intimately connected with the cerebellum anatomically as well as functionally, was also studied. The study group included 15 nondemented and basically healthy cases aged 32-104 years. Linear neuronal density was expressed as number of PC per millimeter tissue measured in the vermis and as neuronal numbers per square millimeter tissue in the inferior olive. The linear PC density clearly decreased with increasing age, R2 = 0.82 and p < .001. The inferior olive showed a small but insignificant age-related neuronal loss. We conclude that aging results in reduced PC density in the vermis cerebelli, further accentuated in the very late stages of life. PMID- 10536642 TI - Age-related changes in alpha-tocopherol dynamics with relation to lipid hydroperoxide content and fluidity of rat erythrocyte membrane. AB - Age-related changes in alpha-tocopherol dynamics in plasma and erythrocyte membranes of 10- to 120-week-old rats were investigated by high-performance liquid chromatography (HPLC) with redox detection mode. Furthermore, changes in lipid hydroperoxide content and fluidity of erythrocyte membrane with age were assessed using chemiluminescence-HPLC and Fourier transform infrared spectrophotometer, respectively. A slight increase in the alpha tocopherolquinone/alpha-tocopherol ratio in erythrocyte membrane and a decrease in the alpha-tocopherol in erythrocyte membrane/alpha-tocopherol in plasma ratio were observed. A significant increase in lipid hydroperoxide content and a marked decrease in the fluidity of erythrocyte membrane were seen with age. These findings suggest that alpha-tocopherol uptake in erythrocyte membrane declines, and utilization rate of alpha-tocopherol in erythrocyte membrane increases age dependently. These changes, which enhanced lipid peroxidation and consequently reduced membrane fluidity, may be caused by the impairment of this transfer mechanism. PMID- 10536641 TI - Potassium depletion: renal membrane lipid metabolism and Na/H exchanger abundance in aged F344 x BNF1 rats. AB - Potassium depletion (-K) is a common electrolyte abnormality in elderly humans, occurring after diuretic use or poor oral intake. We hypothesized that aging would result in an increase in renal membrane lipid metabolism in both control and -K, and that the Na/H exchanger's protein abundance to -K would be blunted. Young and senescent non-obese male Fisher 344 x Brown-Norway F1 rats (F344 x BNF1) were fed either a normal or a K-deficient diet for 7 days. At 24-h, 32P incorporation was measured for renal cortical brush-border (BBM) and basolateral membrane (BLM) lipid metabolism. All -K animals showed a reduction in total body potassium stores, a fall in plasma aldosterone, a urinary concentrating defect, and an increase in plasma cholesterol and urine ammonium excretion (p < .001). In BBM of both age groups, -K increased phosphatidylserine, sphingomyelin, phosphatidylcholine, and phosphatidylethanolamine concentrations, but 32P incorporation fell. In BLM of young K-depleted rats, however, only phosphatidylcholine concentration increased. In the hypokalemic aged rats, the concentration of all BLM phospholipids rose, whereas 32P incorporation fell. In both membranes, cholesterol concentration and the molar ratio of cholesterol to total phospholipid increased with -K (p < .05). Potassium depletion caused brush border membrane NHE-3 protein abundance to rise, but only in the young rats. Neither NHE-3 nor basolateral NHE-1 protein abundance was affected in aged animals with -K. These results provide the first evidence, in non-obese aged rats, that selective age-associated modifications occur in membrane lipid metabolism and membrane transporter protein abundance during -K. That aging causes a maladaptive response in brush-border NHE-3 protein expression may have important implications for elderly humans, particularly if they are given diuretics and become potassium depleted. PMID- 10536643 TI - Dietary restriction alters retinol and retinol-binding protein metabolism in aging rats. AB - Recent studies reported that retinoid metabolism was influenced by long-term dietary restriction (DR) in rats. Because plasma retinol was decreased in rats subjected to DR, it was thought that this dietary manipulation may have an effect on retinol-binding protein (RBP) metabolism. Thus, the aim of this study was to assess retinoids, RBP, and transthyretin (TTR) levels in plasma and liver of young (3 months), adult (12 months), and old (22 months) female Sprague-Dawley rats fed ad libitum (AL) or subjected to a 40% DR, enriched (DR+), or not (DR), with vitamins and minerals. Results indicate that hepatic total retinoid concentrations and content increased with age in all the groups. DR+ rats showed higher hepatic retinoid concentrations than age-matched AL and DR rats. Adult and old DR and DR+ rats exhibited significantly lower plasma RBP-retinol and higher total retinoic acid levels than corresponding controls, although these parameters were not influenced by aging. Liver RBP levels were also decreased in DR and DR+ rats when compared to respective AL controls. There was a slight age-related decline in plasma TTR levels in DR and DR+ rats which was not associated with modifications in liver TTR levels. Hepatic gene expression of RBP and TTR, as evaluated by Northern blot hybridization, did not change with age or diet, suggesting that the lower levels of plasma RBP-retinol and liver RBP in vitamin A sufficient rats subjected to DR may reflect post-transcriptional alterations and/or accelerated degradation of hepatic RBP. The elevated plasma levels of retinoic acid may represent an adaptive mechanism developed by DR rats to maintain retinoid-dependent functions. PMID- 10536644 TI - Effects of age and gender on the cardiovascular responses to isoproterenol. AB - We studied the effects of age and gender on cardiovascular responses to beta adrenergic stimulation with the use of two-dimensional echocardiography in 16 young (aged 20-31) and 20 older (aged 60-75) healthy individuals. Following administration of atropine, each subject was given an infusion of isoproterenol at incremental doses from 0.010 to 0.030 microgram kg-1 min-1. The slopes of the fractional shortening-end-systolic wall stress (FS-sigma es) relationships were steeper in the young men (-0.87 +/- 0.28, n = 8) compared to the older men (-0.41 +/- 0.13, n = 10), and in the young women (-0.55 +/- 0.14, n = 8) compared to the older women (-0.38 +/- 0.13, n = 10). Furthermore, the magnitude of the age associated differences in these slopes was larger in the men (old vs young) than in the women (old vs young) which, in the absence of changes in preload, suggests a greater decline in the contractile response to isoproterenol with advancing age in men compared to women. Furthermore, the men exhibited a greater attenuation of chronotropic response to isoproterenol than did the women. These observations suggest that gender plays a significant role in the age-associated decline in inotropic and chronotropic responses to beta-adrenergic stimulation, with men exhibiting a greater decline with aging than women. PMID- 10536645 TI - Life stress, mood disturbance, and elevated interleukin-6 in healthy older women. AB - BACKGROUND: Although adverse effects of severe chronic stress on immunocompetence and physical well-being in older adults have been reported, the immune response to less severe life stress among healthy older adults, particularly among women, is not well understood. Interleukin-6 (IL-6) has been considered a good overall indicator of immune functioning in older adults because of its contribution to the pathogenesis of several age-related conditions such as osteoporosis. Regulation of IL-6 is impaired in elderly adults, and levels of IL-6 increase with stress and depression. This research cross-sectionally examined levels of IL 6 in three groups of healthy older women with varying levels of life stress and mood disturbance and a healthy group of young women. METHODS: Subjects included 18 caregivers of Alzheimer's patients, 17 older women assessed one month before relocation of their residence, 15 nonmoving and noncaregiving older women, and 20 younger women. Subjects completed the Profile of Mood States (POMS) and had early morning blood draws. RESULTS: Alzheimer's caregivers reported significantly greater distress than women of all other groups. IL-6 levels in caregivers were significantly higher than those of all other women. The older women had significantly higher IL-6 than young controls, but there were no significant differences in IL-6 between movers and older controls. Among all women, greater depression and distress were related to higher levels of IL-6. CONCLUSIONS: These findings suggest that in older women, chronic stressors are associated with significant elevations in IL-6 over and above the elevations associated with normal aging, but that moderate stressors may not be related to appreciable elevations in IL-6. PMID- 10536646 TI - Weight loss and mortality among free-living frail elders: a prospective study. AB - BACKGROUND: Numerous studies of the elderly population have indicated that body weight and weight changes are related to mortality, but the one group at particularly high risk of nutritional inadequacies--frail elders receiving home help services--has not been studied. METHODS: A prospective cohort of 288 frail elders (81 men; 207 women; mean age: 78.2 +/- 7.6 yrs) receiving home support services was followed for 3-5 years. Nutritional variables included baseline body mass index (BMI), weight loss prior to baseline, and energy and protein intake. Covariates included age, gender, smoking, and health and functional status. Cox's multivariate survival analysis was used to identify independent predictors of mortality. RESULTS: There were 102 deaths (35.4%) over the follow-up period. Univariate predictors included age, sex, BMI, weight loss, and functional status. In multivariate analysis, weight loss at baseline was a significant predictor of mortality, RR = 1.76 (95% CI: 1.15-2.71), as was male gender, RR = 2.71 (95% CI: 1.73-4.24), and age at baseline, RR = 1.40 (95% CI: 1.06-1.86). CONCLUSION: Among free-living frail elders, weight loss is a predictor of early mortality after controlling for smoking, and functional and health status indicators. From our observations, however, we cannot conclude that prevention of weight loss would lead to increased survival. This needs to be explored in an intervention study. PMID- 10536647 TI - Implications of body fat distribution in an older twin population. AB - BACKGROUND: As people age, fat becomes preferentially deposited in the abdominal region over the periphery, and such changes are thought to be associated with adverse metabolic outcomes. We were interested in whether body mass index (BMI) and waist-hip ratio (WHR) are differentially associated with fasting insulin levels, triglycerides, and blood pressure (systolic and diastolic) in an older population. We were also interested in whether these associations change after controlling for genetic influences. METHODS: Data were obtained as part of the Swedish Adoption/Twin Study of Aging. All blood samples and anthropometric measures were assessed from 1989-1991 except insulin, which was assessed from 1986-1988. The sample contains 263 twin pairs (97 monozygotic and 166 dizygotic), 56% women, average age 65 years. RESULTS: In men and women, WHR and BMI were significantly associated with all the metabolic variables except for diastolic blood pressure. When BMI's association with the metabolic variables was assessed independent of WHR, it remained significantly associated with all metabolic variables except diastolic blood pressure in men and triglycerides in women. When WHR's association with the metabolic variables was assessed independent of BMI, it did not remain significantly associated with any of the metabolic variables in men and remained significantly associated with insulin and diastolic pressure in women. After controlling for genetic effects, the relationship between WHR and the metabolic variables became nonsignificant. However, BMI remained significantly associated with systolic blood pressure and triglycerides in men, independent of WHR. CONCLUSION: The results suggest that overall body fat is important to consider in relation to these metabolic parameters in older individuals. The results also suggest that BMI may share associations with blood pressure and triglycerides beyond those that can be attributed to familial influences. PMID- 10536648 TI - The inability of hormone replacement therapy or chronic running to maintain bone mass in master athletes. AB - BACKGROUND: Previous studies have demonstrated equivocal findings on the effect of chronic running on bone mass in post-menopausal women. The purpose of this study was to determine the effect of chronic running alone and in conjunction with hormone replacement therapy (HRT) on bone mineral density (BMD) in postmenopausal women. METHODS: Forty-three women [15 premenopausal 48.1 +/- .4 yrs (Pre); 13 postmenopausal 57.3 +/- 2.3 yrs (Post); and 15 HRT-treated postmenopausal 56.8 +/- 1.5 yrs (PostE)] served as subjects. All were chronic runners (duration > 5 yrs, > 10 miles per week). BMD was determined by dual energy x-ray absorptiometry, VO2 max on a treadmill, body composition by hydrostatic weighing, knee strength by KinCom dynamometer, and training and menstrual history by questionnaire. Analysis of covariance with Tukey post hoc tests was utilized to compare the groups. RESULTS: The groups were similar in body weight, VO2 max, years training, and miles run per week. Pre and PostE did not differ in total or spine BMD. However, Pre had greater hip BMD than PostE (.973 +/- .03 vs .876 +/- .03 g/cm2; p < .05). As well, Pre had greater BMD of the hip (.973 +/- .03 vs .805 +/- .03 g/cm2; p < .05), spine (1.047 +/- .04 vs .870 +/- .04 g/cm2; p < .05), and total body (1.115 +/- .02 vs .996 +/- .03 g/cm2; p < .05) than Post. CONCLUSIONS: These results suggest that (a) chronic running + HRT is insufficient to protect hip BMD and (b) chronic running alone provides no protection for bone mass in postmenopausal women. PMID- 10536649 TI - Measuring accumulated health-related benefits of exercise participation for older adults: the Vitality Plus Scale. AB - BACKGROUND: Existing measures fail to capture the perceived benefits attributed to exercise participation by older adults themselves. Noticeable improvements in sleep, energy level, bodily aches and pains, constipation, and other psychophysical aspects of "feeling good" may represent ongoing sources of motivation for continued participation. The Vitality Plus Scale (VPS) was developed to measure these potential health-related benefits of exercising. METHODS: The 10-item VPS was developed using an inductive approach, in collaboration with regularly exercising older adults and their instructors. Multiple samples of exercises and nonexercisers ranging in age from 40 to 94 were used to examine the reliability and validity of the new scale. RESULTS: The VPS showed good internal consistency and test-retest reliability over one week. Scores were able to discriminate on the basis of various indicators of health status and self-reported level of physical activity, and were related to two measures of functional mobility. Convergence was found with several subscales of the SF-36, whereas low correlations emerged with a measure of episode-specific sensations. Responsiveness to change was found with various types of exercise for individuals with low to moderate scores prior to participation. CONCLUSIONS: Improvements in sleep, energy level, mood, and generally feeling good appear to be the most noticeable benefits of exercising for many adults. These associations are reinforced by sustained exercise participation. Capturing these interrelated psychophysical constructs in a single, short measure will enable exercise researchers and instructors to measure incremental improvements previously reported only anecdotally. PMID- 10536650 TI - Serum concentrations of steroids, parathyroid hormone, and calcitonin in postmenopausal women during the year following hip fracture: effect of location of fracture and age. AB - BACKGROUND: Hip fracture in the aged is a major health problem, especially considering the increasing proportion of the elderly in the population. This study examines changes in circulating levels of hormones, which are purported to affect bone metabolism, in response to hip fracture in postmenopausal women. METHODS: Patients consisted of women ages 65 and older who had surgery within 2 days of fracture. Serum samples were obtained at 3, 10, 60, 180, and 360 days postfracture. Healthy women without hip fractures from the same age range served as a control group (n = 17). Hormones were determined by radioimmunoassay. Subjects with fractures in the neck region of the femur (n = 78) were compared to subjects with fractures in the trochanteric region (n = 88). RESULTS: Estrone concentration (47.6 +/- 5.7 pg/mL; mean +/- SEM) at 3 days postfracture was elevated (p < .001) compared to control levels of 20.7 +/- 4.6 pg/mL. By 2 months, levels had declined to control levels. Androstenedione and the adrenal hormones, DHEAS and cortisol, displayed similar responses. Parathyroid hormone (PTH) levels were not significantly different from the control concentration at 3 days following fracture, but increased (p < .001) during the year following fracture. Calcitonin concentrations were much higher (p < .001) 3 days postfracture (42.1 +/- 3.7 pg/mL) compared to controls without fracture (9.8 +/- 3 pg/mL). Except for testosterone, no differences could be attributed to fracture location. Only PTH, with concentrations higher in the older age groups (p < .001), showed an age-related response. CONCLUSIONS: Following hip fracture, there are some dramatic responses in hormones that purportedly are mechanistically important in bone metabolism. These changes include transient increases in steroid hormones, chronic elevations in calcitonin, and rising levels of PTH during the year after fracture. PMID- 10536651 TI - Temporal and regional variation in the use of breast-conserving surgery and radiotherapy for older women with early-stage breast cancer from 1983 to 1995. AB - BACKGROUND: Authorities recommend radiation therapy after breast-conserving surgery for breast cancer. Numerous studies have reported that older women diagnosed with breast cancer are less likely to receive radiation after breast conserving surgery. It is unclear how care of older women with breast cancer has changed over time. METHODS: Women with local or regional stage breast cancer diagnosed between 1983-1995 were identified from the Surveillance, Epidemiology, and End Results (SEER) Cancer Registries. The treatment information in SEER includes type of surgical procedures and receipt of radiation therapy. RESULTS: There were small increases in the percentage of women receiving breast-conserving surgery during the 1980s followed by substantial increases in the 1990s. Age was a major factor in determining receipt of radiation therapy after breast conserving surgery. A large increase in use of radiotherapy after surgery was observed in women aged > or = 75, from below 30% in 1983 to over 50% in 1995. Women aged > or = 75 diagnosed in 1992-1995 were 1.76 and 2.34 times more likely to receive radiation for local and regional stage respectively, as compared to those in 1983-1987. There was no increase in use of radiation for women aged < 65. CONCLUSIONS: There has been a substantial increase in use of breast conserving surgery and in receipt of radiation therapy after breast-conserving surgery in older women. However, there was a net increase in the percentage of all women with breast cancer who received this surgery without radiotherapy, due to the large increase in the overall percentage of women receiving this surgery. PMID- 10536652 TI - Elevated salivary cortisol in the evening in healthy elderly men and women: correlation with bone mineral density. AB - BACKGROUND: Aging is associated with a loss of bone mineral density (BMD) in men and women. Loss of BMD can also be caused by hypercortisolemia in men or women at any age. This study measured salivary cortisol at 2300 h and 0700 h as indices of cortisol secretory activity in 228 elderly, community-dwelling subjects. Salivary cortisol results were correlated with BMD. We hypothesized that salivary cortisol is elevated at 2300 h in elderly people, and that salivary cortisol will correlate negatively with BMD. METHODS: Saliva was sampled at 2300 h (nadir in circadian rhythm) and 0700 h (peak in circadian rhythm) in 130 men (70.7 +/- 0.4 years old) and 98 women (70.0 +/- 0.4 years old); approximately half of the women were receiving hormone replacement therapy (HRT). BMD was measured by dual energy x-ray absorptiometry. RESULTS: Salivary cortisol at 2300 h was significantly elevated in men (2.3 +/- 0.1 nmol/L) and women (2.1 +/- 0.1 nmol/L) as compared to 73 younger controls (1.2 +/- 0.1 nmol/L; 37 +/- 1 year old). Salivary cortisol at 0700 h was not different between older subjects and younger controls. There was a significant negative correlation of lumbar (L2-4) BMD and 2300 h salivary cortisol in older women (r = -0.20, p = .05; n = 98); this correlation was significant only in women not on HRT. There was a highly significant negative correlation of lumbar (L2-4) BMD and 0700 h salivary cortisol in older men (r = 0.31, p = .0003). CONCLUSIONS: Salivary cortisol is a simple, nonstressful method for assessing activity of the hypothalamic-pituitary-adrenal (HPA) axis in the elderly population. A major finding was an elevation in the late night nadir in cortisol secretion. We also suggest that elevated cortisol secretion in elderly people may contribute to the age-related loss in bone mineral density and that this effect is prevented by HRT. PMID- 10536653 TI - A vicious backhand. PMID- 10536654 TI - Tetrahydrobiopterin and endothelial nitric oxide synthase activity. PMID- 10536655 TI - Heterogeneous transmural gene expression of calcium-handling proteins and natriuretic peptides in the failing human heart. PMID- 10536656 TI - Sarcoplasmic reticulum Ca2+ ATPase gene expression to the rescue of myocardial contractility in hypothyroid associated heart failure. PMID- 10536657 TI - Understanding the temporal relationship of ATP loss, calcium loading, and rigor contracture during anoxia, and hypercontracture after anoxia in cardiac myocytes. PMID- 10536658 TI - Measure is treasure. PMID- 10536659 TI - Inflammation in ischemic heart disease: response to tissue injury or a pathogenetic villain? PMID- 10536660 TI - Interactions between endothelin-1 and the renin-angiotensin-aldosterone system. AB - The renin-angiotensin-aldosterone (RAA) system and the endothelin (ET) system entail the most potent vasopressor mechanisms identified to date. Although they were studied in depth in relation to arterial hypertension and cardiovascular diseases, limited information on their interrelationships in causing hypertension and related target organ damage exists. The identification of consensus sequences for jun in the regulatory region of the preproendothelin-1 (ppET-1) gene raised the possibility of its transcriptional regulation by angiotensin II (Ang II). This was confirmed by the finding that stimulation with Ang II of cultured vascular smooth muscle cells (VSMCs) and endothelial cells (ECs) induced expression of the ppET-1 gene and synthesis of ET-1. Endogenously produced ET-1 was found to contribute to the hypertrophic response of cardiomyocytes to Ang II and thereby to cardiac hypertrophy. Furthermore, ET-1 exerts multifaceted effects on the RAA system, such as dose-dependent inhibition of renin synthesis, and stimulation of aldosterone secretion. The finding of abundant specific ET-1 receptors in the adrenocortical zona glomerulosa (ZG) suggested a direct secretagogue effect of ET-1. In rats, ETB receptors mediate such an effect, whilst in humans, both ETA and ETB receptor subtypes intervene in regulating the transcription of the aldosterone synthase gene. In addition, ET-1 stimulates DNA synthesis and proliferation of ZG cells via ETA receptors and, therefore, might play a role in cell turnover of the normal adrenal cortex and in the onset of adrenal tumours. Studies on the in vivo interactions between ETs and the RAA system have given conflicting results, insofar as some suggested a participation of ET-1 in the pressor and cellular effects of exogenously administered Ang II, whereas others did not in the transgenic TGR(Ren 2m)27 rats and in the two kidney, one clip. PMID- 10536661 TI - Lipids and the endothelium. AB - The normal endothelium is characterised by the production of a number of molecules which affect the contractile state of adjacent myocytes and the behavior of formed elements within the blood stream, and by the absence of cell surface adhesion molecules. In addition, endothelial cells are important modulators of coagulation and fibrinolysis. Whilst effects of lipids have been documented on many of these endothelial processes, there is particularly strong evidence for effects on the vasodilatation mediated by endothelium derived nitric oxide and on the interaction between leukocytes and the endothelial surface. Both LDL cholesterol and triglyceride rich lipoproteins impair endothelium dependent vasodilatation. The effects of LDL cholesterol are primarily evident for lipoprotein particles that have been oxidised with evidence for effects of specific constituents of oxidised LDL, such as lysophosphatidylcholine (LPC). LDL effects have been demonstrated at numerous sites of the nitric oxide signaling pathway including receptor-G protein coupling, nitric oxide synthase and NO bioactivity, with evidence for enhanced superoxide formation and the consequent production of the less potent dilator peroxynitrite. The effects of lipids on endothelium dependent vasodilatation can be reversed not only by reducing the level of elevated lipids levels but also by provision of the NOS substrate, L arginine and by the provision of antioxidants, although the mechanism for these effects are not fully elucidated. The adhesion of leukocytes to the endothelial surface is stimulated by low density and triglyceride rich lipoproteins. As with endothelium dependent vasodilatation, the effects of LDL cholesterol are primarily evident for low-density lipoprotein particles that have been oxidised, and many of the effects of oxidised LDL can be mimicked by LPC. HDL can overcome pro-adhesive effects of oxidised LDL. The effects of LDL on leukocyte adhesion are secondary to the expression of adhesion molecules on the luminal surfaces of endothelial cells. In addition to the likely deleterious effects of lipids on endothelium-mediated vasodilatation and leukocyte-endothelial cell interaction, lipids have been shown to affect a number of other endothelial processes and function. Thus, oxidised LDL affects endothelial ET1 and PGI2 release. Although effects have been shown on endothelial cell growth and apoptosis and on endothelial processes related to thrombosis and fibrinolysis, these effects have been less extensively studied than endothelial dependent vasodilatation and leukocyte-endothelial cell interaction. Many of the effects of elevated or modified low density and TG rich lipoproteins on endothelial cells and endothelial cell processes could be expected to contribute to the development of atherosclerosis and therefore, to the association between lipids and atherosclerotic, particularly coronary, vascular disease. However, the extent to which "endothelial dysfunction" accounts for the known relationships between serum lipid concentrations and CHD is yet to be established. PMID- 10536662 TI - Heterogeneous transmural gene expression of calcium-handling proteins and natriuretic peptides in the failing human heart. AB - OBJECTIVE: Human heart failure is associated with a disturbed intracellular calcium (Ca2+) homeostasis. In this regard, ventricular wall stress is considered to be a determinant for expression of sarcoplasmic reticulum Ca(2+)-ATPase (SERCA2a). In the present study, we analyzed the transmural protein and/or mRNA levels of SERCA2a, other Ca(2+)-handling proteins, and of atrial and brain natriuretic peptides (ANP and BNP) in the human heart. METHODS: Subepicardial (epi), midmyocardial (mid), and subendocardial (endo) sections of the left ventricular free wall from end-stage failing (n = 17) and nonfailing (n = 5) human hearts were analyzed by Western blot for immunoreactive protein levels of SERCA2a, phospholamban (PLN), and calsequestrin (CS). Subepi- and subendocardial sections were analyzed by Northern blot for steady-state mRNA levels of SERCA2a, Na(+)-Ca2+ exchanger (NCX1), ANP, and BNP. RESULTS: SERCA2a protein and mRNA levels were reduced by 40 +/- 5% (P < 0.01) and 25 +/- 7% (P < 0.05) in endo compared to epi in the failing heart and by 27 +/- 14% and 16 +/- 12% (non significant) in the nonfailing heart, respectively. PLN protein levels were reduced by 23 +/- 6% (P < 0.05) in endo compared to epi in the failing heart and by 17 +/- 25% (non-significant) in the nonfailing heart, whereas CS protein levels and NCX1 mRNA levels were similar across the left ventricular wall. Strikingly, in the failing heart, both BNP and ANP mRNA levels were upregulated predominantly in endo. CONCLUSIONS: In the failing human heart, SERCA2a and PLN, as well as natriuretic peptides but not CS and NCX1 are differentially expressed across the left ventricular wall, implicating (1) different susceptibility of subendocardium and subepicardium to factors affecting expression of these proteins and (2) differences in regulation of the distinct calcium-cycling proteins. PMID- 10536663 TI - Repolarizing currents in ventricular myocytes from young patients with tetralogy of Fallot. AB - OBJECTIVE: It was the aim of our study to describe repolarizing currents in ventricular myocytes isolated from children with tetralogy of Fallot. This is the first report on outward currents in ventricular myocytes from children. METHODS: Ventricular myocytes were isolated from tissue samples of the outflow tract of the right ventricle which were obtained during corrective surgery of tetralogy of Fallot. Action potentials and whole-cell currents were recorded with the patch clamp technique at a temperature of 36-37 degrees C. RESULTS: The mean resting potential was -71.7 +/- 1.92 mV, action potential amplitude was 110 +/- 2.96 mV and action potential duration at 90% repolarization was 794 +/- 99.5 ms (n = 12). In four out of 12 myocytes early afterdepolarizations (EADs) were observed. Upon hyperpolarization Ba(2+)-sensitive inward currents similar to the inward rectifier current (IKl) could be observed. The current density at -120 mV was 22.8 +/- 2.47 pA/pF (n = 14). A transient outward current (Itol) could be recorded in all myocytes studied, the current density varied from 0.3 to 8.6 pA/pF with a mean of 3.77 +/- 0.47 pA/pF at +40 mV (n = 38). Recovery of Itol from inactivation was fast (70% recovery within 100 ms), rate-dependent reduction amounted to 38.2% at 4 Hz. A delayed rectifier current was seen in only two out of 38 myocytes (rapid component IKr). CONCLUSIONS: The electrophysiological characteristics of right ventricular myocytes isolated from children with tetralogy of Fallot resemble in most cases subendocardial myocytes from adults. The most prominent difference is a fast recovery from inactivation as well as a small rate dependent reduction of Itol. The observed EADs may have clinical implications. PMID- 10536664 TI - Afterload induced changes in myocardial relaxation: a mechanism for diastolic dysfunction. AB - BACKGROUND: Diastolic left ventricular (LV) dysfunction manifests as an upward shift of the diastolic pressure-volume relation. One of the possible causes of diastolic LV dysfunction is incomplete myocardial relaxation. It is well known that high afterload slows myocardial relaxation. This contribution investigated to what extent afterload elevation could also affect LV filling pressures including end-diastolic LV pressure (LVP). METHODS: Selective, beat-to-beat elevations of afterload were induced in anaesthetised open-chest rabbits (n = 9) by abrupt narrowing of the ascending aorta during the diastole of the preceding heartbeat. This was performed with physiological heart rate and blood pressure. RESULTS: These interventions increased systolic LVP from 90 +/- 3 mm Hg at baseline to 103 +/- 4, 123 +/- 5, 139 +/- 5 and 154 +/- 6 mm Hg. The last intervention was a total aortic occlusion inducing a first beat isovolumetric contraction. Smaller afterload elevations decreased tau (accelerated LVP fall) and did not elevate diastolic pressure-internal diameter relation (P-ID). Larger afterload elevations increased tau (decelerated LVP fall), induced an upward shift of the diastolic P-ID and increased end-diastolic LVP. Effects of afterload on end-diastolic LVP were correlated with effects on tau (r = 0.89; P < 0.01). Incomplete relaxation or load-dependent residual active state appeared to be the mechanism for this diastolic dysfunction. Similar findings were made retrospectively in dogs instrumented with circumferential segment length gauges (n = 16). CONCLUSIONS: Diastolic LV dysfunction was induced by elevated afterload in healthy hearts of rabbits and dogs. If this mechanism could be shown to be operative in the failing heart, reversal of diastolic dysfunction should contribute to the beneficial effects of vasodilating and inotropic therapy on pulmonary congestion. PMID- 10536665 TI - Load sensitivity of left ventricular relaxation in normal and failing hearts: evidence of a nonlinear biphasic response. AB - OBJECTIVE: This investigation sought to define the effect of heart failure (HF) on the load sensitivity of left ventricular (LV) relaxation and to correlate alterations in load sensitivity with the variables of ejection timing, systolic load profile and elastic recoil. METHODS: Nine dogs instrumented with LV micromanometers and piezoelectric crystals were studied before and after HF produced by prolonged rapid LV pacing. After pharmacologic autonomic blockade and atrial pacing at 160 bpm, hemodynamic measurements were recorded at steady-state and during vena caval occlusion. LV relaxation for individual beats during caval occlusion was assessed using tau, the monoexponential time constant, and systolic load was estimated using end-systolic circumferential force (ESF). RESULTS: The tau-ESF relation was nonlinear and biphasic, with an initial decrease in tau followed by a delayed increase, and was described by a parabolic equation with a curvilinearity coefficient a. Examination of ejection variables and systolic load profile indicated that the initial acceleration of relaxation reflected the influence of increased elastic recoil, whereas the late slowing reflected the influence of earlier end-ejection and delayed systolic loading. HF produced significant baseline prolongations of tau (P < 0.005), time to relaxation onset (P < 0.001) and time to peak force (P < 0.015) compared to control. The curvilinearity coefficient of the tau-ESF relation was significantly increased (18.1 +/- 20.1.10(-5) vs. 3.99 +/- 2.89.10(-5) g-2, P = 0.048), indicating increased load sensitivity of relaxation. This increased load sensitivity correlated with delayed onset but increased overall magnitude of the effects of earlier end-ejection and late systolic loading on relaxation. CONCLUSIONS: The tau-ESF relationship during transient load reduction is nonlinear and biphasic with an initial acceleration of relaxation, reflecting the impact of elastic recoil, and delayed slowing, reflecting changes in ejection timing and systolic loading sequence. This relation is more curvilinear in the failing heart, indicating increased load sensitivity of LV relaxation. These changes primarily occur due to alterations in the impact of ejection timing and systolic load profile rather than increased elastic recoil. PMID- 10536666 TI - Electrophysiological responses of canine atrial endocardium and epicardium to acetylcholine and 4-aminopyridine. AB - OBJECTIVES: Prior studies demonstrated marked electrophysiological and pharmacological differences between canine ventricular epicardium and endocardium. For atrium, however, it has been assumed that, because of the thin wall, electrical properties of epicardium and endocardium are similar. The aim of the present study was to compare the action potential (AP) characteristics in epicardial and endocardial atrial cells before and following addition of acetylcholine (ACh) and 4-aminopyridine (4-AP). METHODS AND RESULTS: Microelectrode techniques were used to study the effects of ACh (10(-7)-10(-5) M) and 4-AP (0.5 mM) on epicardial and endocardial AP of canine right atrial free wall at cycle lengths (CL) of 250 to 2000 ms. ACh hyperpolarized epicardial and endocardial cells (by 5-8 mV at 10(-5) M). In control, AP duration to 90% repolarization (APD90) was longer in endocardium at all CL. ACh shortened APD90 in either tissue with more prominent effect in endocardium (at 10(-5) M and CL = 2000 ms, from 179 +/- 10 to 90 +/- 11 ms in epicardium and from 209 +/- 10 to 65 +/- 6 ms in endocardium, P < 0.05). As a result, at 10(-5) M, APD90 in endocardium was shorter than in epicardium at all CL 4-AP effects on AP duration were similar in both tissue types. No effects of 4-AP was seen at CL = 250 ms and at long CL, the compound shortened APD90 and prolonged AP duration to 50% repolarization. CONCLUSIONS: (1) ACh exerts direct effects on atrial epicardial and endocardial AP; (2) 4-AP-sensitive transient outward current (Itol) is expressed both in canine atrial epicardial and endocardial cells; (3) differential response of epicardial and endocardial APD to ACh may alter the gradient of repolarization across the atrial wall and contribute to vagally induced atrial flutter and fibrillation. PMID- 10536667 TI - Differential expression of alpha 1, alpha 3 and alpha 5 integrin subunits in acute and chronic stages of myocardial infarction in rats. AB - OBJECTIVE: Anchoring cardiac myocytes to extracellular matrix, which is mediated mainly by integrins on their surfaces, is important for maintaining the architecture of myocardial tissues and transmitting mechanical force. We evaluated the expression of alpha integrin subunits on myocytes and the accumulation of interstitial collagen and fibronectin at acute and chronic stages after myocardial infarction. METHODS: Myocardial infarction was induced by ligation of left coronary arteries in rats. The expression of alpha 1, alpha 3 and alpha 5 integrin subunits, and accumulation of collagen and fibronectin were analyzed with immunohistochemistry or sirius-red staining. RESULTS: In hearts without infarction, moderate expression of the alpha 3 subunit and only slight expression of the alpha 5 subunit were observed on myocytes. In the first week after infarction, the alpha 1 subunit, collagen and fibronectin were increased only in the peri-infarcted area, while the alpha 5 subunit was increased both in peri-infarcted and non-infarcted areas. At day 42, the expression of the alpha 1 subunit and collagen were still increased, although the alpha 5 subunit and fibronectin were decreased. The expression of the alpha 3 subunit was not altered throughout the experimental period. CONCLUSION: These data suggest that integrin subunits play an important role in healing and remodeling processes after myocardial infarction. PMID- 10536668 TI - Overexpression of sarcoplasmic reticulum Ca(2+)-ATPase improves cardiac contractile function in hypothyroid mice. AB - OBJECTIVE: Prolonged cardiac contraction and relaxation in hypothyroidism are in part related to diminished expression of the gene coding for the calcium pump of the sarcoplasmic reticulum (SERCA2a). Therefore, we examined whether or not transgenic SERCA2a gene expression in mice may compensate for the cardiac effects of hypothyroidism. METHODS: SERCA2a mRNA and protein were analyzed from hearts of euthyroid and hypothyroid mice of wild-type or SERCA2a transgene status. Contractile function was studied in isolated left ventricular papillary muscles. RESULTS: We found significant decreases of SERCA2a mRNA and protein levels in hearts of hypothyroid wild-type mice in comparison with euthyroid wild-type mice (controls). Papillary muscles from hypothyroid wild-type mice showed significant increases in time to peak contraction and relaxation times compared with controls. In contrast, SERCA2a mRNA and protein levels were significantly higher in hypothyroid SERCA2a transgenic mice than in hypothyroid wild-type mice. The transgene led to a functional improvement by compensating for the prolonged contraction and relaxation of papillary muscles. CONCLUSIONS: Our murine model of hypothyroidism revealed decreases in SERCA2a gene expression accompanied by prolonged contraction and relaxation of papillary muscles, and an improvement of the contractile phenotype due to compensated SERCA2a gene expression in SERCA2a transgenic mice. PMID- 10536669 TI - Inhibitory effects of vesnarinone in the progression of myocardial damage in experimental autoimmune myocarditis in rats. AB - OBJECTIVES: Vesnarinone, a positive inotropic and immunomodulatory agent, diminishes nitric oxide (NO) levels by suppressing the induction of inducible NO synthase (iNOS) expressed in cytokine-stimulated macrophages and cardiomyocytes. We examined whether vesnarinone exerts inhibitory effects on the progression of myocardial damage in experimental autoimmune myocarditis in rats through suppression of iNOS. METHODS: Myocarditis was induced in 30 Lewis rats by injection of porcine cardiac myosin and vesnarinone was orally administered to 20 of the 30 rats. On day 21 after immunization (the climax of inflammation), the hemodynamics were examined and the severity of myocarditis was evaluated by determining the area ratio (%) [affected/entire area] of myocardial lesions in histological sections. Levels of serum CK-MB, NOx (NO2(-)+NO3-), TNF-alpha and IL 1 beta, and cyclic GMP, iNOS mRNA, TNF-alpha and IL-1 beta in heart tissues were determined. Expression of iNOS and TNF-alpha protein were examined by immunohistochemical methods. RESULTS: Histopathological examination revealed extensive myocardial destruction and massive infiltration of inflammatory cells in the vesnarinone-untreated rats. The area ratio of the lesions in the treated rats was significantly lower than that in the untreated ones. Levels of CK-MB, NOx, cyclic GMP, cytokines and iNOS mRNA were significantly lower in the vesnarinone-treated rats. Infiltrating macrophages and cardiomyocytes in the untreated rats showed much higher levels of expression of iNOS and TNF-alpha than those in the vesnarinone-treated rats. CONCLUSIONS: Vesnarinone may prove to be useful in the treatment of myocarditis by attenuating NO production through suppression of iNOS induced by cytokines. PMID- 10536670 TI - Doxorubicin induces slow ceramide accumulation and late apoptosis in cultured adult rat ventricular myocytes. AB - OBJECTIVES: Anthracyclines cause apoptotic death in many cell types through activation of the ceramide pathway. We tested the hypothesis that doxorubicin induces cardiac myocyte apoptosis through ceramide generation. METHODS: Adult rat ventricular myocytes were grown in the presence of 10% fetal calf serum, and exposed to 0.5 microM doxorubicin (Dox) for 1 h on the day of cell isolation (day 0). We used the membrane-permeant ceramide analog C2-ceramide (C2-cer) to mimic the effects of endogenous ceramide and PDMP to induce endogenous ceramide accumulation. Apoptosis was assessed upon morphological criteria and DNA fragmentation by the TUNEL method and agarose gel electrophoresis. Ceramide concentration was assessed using the DAG kinase assay. RESULTS: Myocyte exposure to Dox was associated with cellular and nuclear alterations typical of apoptosis on day 7 but not on day 3. At day 7, the percentage of TUNEL-positive myocytes was markedly increased in Dox-treated cultures compared to control (Cl) cultures (82 +/- 3 vs. 12 +/- 1%, n = 7; p < 0.001); internucleosomal DNA fragmentation was confirmed by the observation of DNA ladders. These alterations were associated with an increase in the intracellular ceramide concentration (1715 +/- 243 vs. 785 +/- 99 pmol/mg prot, n = 5; p < 0.01), a phenomenon also detected on day 3 (731 +/- 59 vs. 259 +/- 37 pmol/mg prot, n = 5; p < 0.001). Incubation of myocytes at day 0 with 50 microM C2-cer induced rapid cell shrinkage and DNA fragmentation (45 +/- 3 vs. 10 +/- 1% TUNEL-positive myocytes on day 1 in C2-cer treated and Cl cultures, respectively; n = 6, p < 0.001). Myocyte exposure to 10 microM PDMP for 7 days (n = 5), caused ceramide accumulation (1.7-fold increase vs. Cl, p < 0.01), and a marked increase in the percentage of TUNEL-positive myocytes (62 +/- 6 vs. 11 +/- 3% in Cl cultures, p < 0.001). Ventricles of rats injected intraperitoneally with a cumulative dose of 14 mg/kg Dox over a period of 2 weeks also showed an increased ceramide concentration 2 weeks later (11.01 +/- 0.64 vs. 5.24 +/- 0.88 pmol/mg prot, n = 6; p < 0.001). CONCLUSION: Our study confirms the existence of a functional ceramide pathway related to apoptosis in cardiac myocytes, and points to its possible involvement in doxorubicin-induced cardiomyopathy. PMID- 10536671 TI - Pretreatment with PKC activator protects cardiomyocytes against reoxygenation induced hypercontracture independently of Ca2+ overload. AB - OBJECTIVE: Although several studies have shown that activation of protein kinase C (PKC) plays an important role in protection through ischemic preconditioning, little is known about the effects of direct PKC activation on the course of ischemia-reperfusion injury. The aim of this study was to analyse the effects of a pretreatment with the PKC activator 1,2-dioctanoyl-sn-glycerol (1,2DOG). METHODS: Isolated adult Wistar rat cardiomyocytes were exposed to 80 min of simulated ischemia (anoxia, pHo 6.4) and 20 min of reoxygenation (pHo 7.4). Cytosolic Ca2+ (fura-2), cytosolic pH (BCECF), Mg2+ (Mg-fura-2), lactate and cell length were measured and compared between control cells and cells treated with 20 mumol/l 1,2DOG before anoxia (10 min treatment and 10 min wash out). RESULTS: 1,2DOG-pretreatment delayed the time to extreme ATP depletion, but had no effect on lactate production and cytosolic pH. The accumulation of cytosolic Ca2+ was markedly accelerated in pretreated cells that developed rigor shortening, but reoxygenation-induced hypercontracture was significantly reduced. 1,2DOG, therefore, completely abolished Ca(2+)-dependence of hypercontracture. The effects of pretreatment were fully abolished with 1 mumol/l bisindolylmalcimide (PKC inhibitor). We conclude that PKC preactivation leads to (1) reduction of energy demand, (2) acceleration of Ca2+ overload during anoxia and (3) prevention of reoxygenation-induced hypercontracture independent of anoxic changes in cytosolic Ca2+ and pH. PMID- 10536672 TI - Sodium influx via a non-selective pathway activated by the removal of extracellular divalent cations: possible role in the calcium paradox. AB - OBJECTIVE: Cation non-selective conductances which are induced upon removal of extracellular divalent cations have been identified in cardiac and other cells. We have examined whether the conductance identified in cardiac myocytes mediates an increase in intracellular Na+ (Nai+) and have tested the ability of drugs to prevent this influx. METHODS: Rat single ventricular myocytes at 22 degrees C were voltage-clamped in whole-cell mode to measure membrane currents or were loaded with SBFI to measure Nai+. RESULTS: Removal of extracellular Ca2+ (Cao2+) and Mg2+ (Mgo2+), which induced a current with reversal potential of -10 mV, also caused an increase in SBFI fluorescence ratio (340/380 nm). These changes were reversible on repletion of Cao2+ and/or Mgo2+. They could not be prevented by nifedipine, indicating that they were not mediated by L-type Ca2+ channels. Both increases in non-selective conductance and in Nai+ were prevented by trivalent cations (Dyo3+, Gdo3+ or Lao3+; 100 microM) or reduced by the aminoglycoside gentamicin. CONCLUSION: A cation non-selective conductance, different from L-type Ca2+ channels, contributes to the Nai+ accumulation obtained during perfusion with Ca2+/Mg(2+)-free media, hence also to the Cai2+ overload and cellular damage upon Cao2+ repletion (the Ca2+ paradox). PMID- 10536673 TI - Sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA2) gene products are regulated post-transcriptionally during rat cardiac development. AB - OBJECTIVE: The Sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA2) plays a major role in the contraction-relaxation cycle and is responsible for transporting calcium into the lumen of the sarcoplasmic reticulum. This study was performed to determine if the increase in SERCA2 messenger RNA (mRNA) abundance during the perinatal period is regulated transcriptionally. METHODS: Transcriptional activity was determined by nuclear run-on assays and mRNA and protein abundances were determined during late fetal and early neonatal cardiac development in rat. RESULTS: From nuclear run-on assays, SERCA2 gene transcription at 17/18 embryonic days (139 +/- 41 parts per million (ppm), n = 7) did not differ from that at 20 neonatal days (139 +/- 37 ppm, n = 6) after birth. No increase in transcriptional activity could be demonstrated during the time frame examined. In contrast, both alpha and beta myosin heavy chains showed significant changes in measured transcriptional activity. SERCA2 mRNA normalized to 18S RNA levels are very low in the fetus (9.8 +/- 1.9 to 13.4 +/- 4.9 arbitrary units (A.U.) from 17/18 to 19/20 embryonic days) and significantly increase from birth (15 +/- 3.8 A.U.) to reach a maximum at 20 days of age (29.1 +/- 9.5 to 48.3 +/- 7.0 in 15 to 20 neonatal days rats respectively). Similarly, SR Ca(2+)-ATPase protein levels are less abundant in the fetus (0.82 +/- 0.08 to 1.13 +/- 0.13 A.U./microgram total protein) and reach a maximum at 15-20 neonatal days (3.08 +/- 0.58 to 2.98 +/- 0.17). Ca2+ uptake in the fetal heart is about one sixth the level seen in the adult, reaches the highest observed value at 5 days after birth (6.05 +/- 0.77 pmole Ca2+ per microgram/min) and remains relatively constant over the next 15 days. The activity increases even though phospholamban protein increases in abundance. CONCLUSIONS: Since the transcriptional activity of this gene is unchanged whereas the mRNA, protein abundance and activity increase, we conclude that the abundance of SERCA2 gene products is regulated primarily through post transcriptional mechanisms during the perinatal period. PMID- 10536674 TI - Chronic regulation of arterial blood pressure in ANP transgenic and knockout mice: role of cardiovascular sympathetic tone. AB - OBJECTIVE: Atrial natriuretic peptide (ANP) lowers arterial blood pressure (ABP) chronically, in association with vasodilation of the resistance vasculature. The mechanism mediating the chronic relaxant effect of ANP is likely indirectly mediated by interactions with tonic vasoeffector mechanisms, inasmuch as the resistance vasculature is relatively insensitive to direct cGMP-mediated relaxation by ANP. On the basis of evidence that ANP has widespread sympatholytic activity, the current study investigated whether the chronic hypotensive effect of ANP is mediated by attenuation of tonic cardiovascular sympathetic tone. METHODS: Total plasma catecholamine concentration and changes in basal ABP and heart rate (HR) following autonomic ganglionic blockade were measured as indices of underlying sympathetic nerve activity in hypotensive ANP-overexpressing transgenic mice (TTR-ANP), hypertensive ANP knockout mice (-/-) and the genetically-matched wild type (NT and +/+, respectively) control mice. Pressor and chronotropic responses to norepinephrine infusion were measured in ganglion blocked mice of all genotypes, and norepinephrine receptor binding was assessed in representative tissues of -/- and +/+ mice, in order to determine whether peripheral adrenergic receptor responsiveness is altered by ANP-genotype. RESULTS: Basal ABP was significantly lower in TTR-ANP and higher in -/- compared to their wild-type controls. Basal HR did not differ significantly between mutant and control mice. Autonomic ganglionic blockade reduced ABP and HR in all genotypes, however, the relative decrease in ABP was significantly smaller in TTR ANP and greater in -/- mice than in their respective controls. Total plasma catecholamine was significantly higher in -/- than in +/+ mice but did not differ significantly between TTR-ANP and NT mice. Norepinephrine infusion during ganglionic blockade elicited quantitatively similar pressor and chronotropic responses in mutant and control mice. Tissue norepinephrine binding did not differ significantly between -/- and +/+ mice. CONCLUSIONS: The present study shows that differences in endogenous ANP activity in mice, resulting in chronic alterations in ABP are accompanied by directional changes in underlying cardiovascular sympathetic tone, and suggests that the chronic vasodilator effect of ANP is, at least partially, dependent on attenuation of vascular sympathetic tone, possibly at a prejunctional site(s). PMID- 10536675 TI - Altered endothelin-1 binding following balloon angioplasty of pig coronary arteries: effect of the ETA receptor antagonist, LU 135252. AB - OBJECTIVE: Since raised levels of endothelin-1 (ET-1) have been detected in the human coronary sinus following percutaneous transluminal angioplasty (PTCA) we investigated the role of ET-1 in the etiology of vascular restenosis. METHODS: Balloon angioplasty of coronary arteries was performed in pigs and the animals were treated with placebo or the endothelin (ETA) receptor antagonist LU 135252 (30 mg/kg/day). After 4 weeks vascular stenosis and the distribution of endothelin and its receptors was evaluated. RESULTS: The pronounced neointima formation in the control group (neointima:media ratio = 0.87 +/- 0.36) was significantly reduced by LU 135252 (0.43 +/- 0.30, P < 0.001). Angioplasty caused a significant increase in medial ETA (approximately 275%, P < 0.026) and ETB (approximately 250%, P < 0.001) binding to injured, compared with non-injured segments, an effect that was also reduced by LU 135252 (ETA = 11.5% increase; ETB = 14% increase). The neointima of control animals exhibited ET-1 like immunoreactivity as well as ETA and ETB binding sites. CONCLUSION: These data indicate that endothelin is locally-released from endothelial and vascular smooth muscle cells following angioplasty which binds to ETA and ETB receptor sites in the neointima and media. Since administration of the ETA antagonist LU 135252 markedly reduces neointima formation and medial ET binding, we conclude that vascular smooth muscle cell proliferation and subsequent neointima formation is mediated predominantly via ETA receptors. These data underscore the therapeutic potential of ETA antagonists in reducing the degree of restenosis following vascular injury. PMID- 10536676 TI - Impairment of G-protein-mediated signal transduction in the porcine coronary endothelium during rejection after heart transplantation. AB - BACKGROUND: Endothelial dysfunction is an early event leading to atherosclerosis. It also occurs after orthotopic heart transplantation and can be used to predict the development of intimal hyperplasia in the coronary artery wall. The present study was designed to assess the time course and specific alterations underlying endothelial dysfunction due to rejection after heart transplantation. METHODS: A porcine model of heterotopic heart transplantation was used. Preoperative serum typing for the class I antigen of the swine lymphocyte alloantigen was performed to ensure compatibility for this antigen. This permitted survival of the graft with a low grade rejection without immunosuppression. Rings (with or without endothelium) of epicardial coronary arteries of native and transplanted hearts were studied in organ chambers filled with modified Krebs-Ringer bicarbonate solution and compared 1, 30 and 60 days after transplantation. RESULTS: Myocardial contractility was normal in all grafts studied at 60 days after transplantation and all coronary arteries were patent. Myocardial biopsies showed the progression of rejection from day 1 to day 60 after implantation. All endothelium-dependent relaxations were normal one day after transplantation. Endothelium-dependent relaxations to serotonin and to the alpha 2-adrenergic agonist UK14304 (which both activate receptors coupled to Gi-proteins) and to sodium fluoride (a direct activator of G-proteins) were decreased 30 days after transplantation, while those to the calcium ionophore, A23187, and bradykinin were shifted to the right and those to ADP were normal. At 60 days, endothelium dependent relaxations mediated by the Gi-protein pathway were decreased further while the concentration-relaxation curves to the other agonists were further shifted to the right. Endothelium-independent relaxations to the nitric oxide donor, Sin-1, were progressively reduced at 30 and 60 days, but maximal relaxations were maintained at 60 days. Histomorphometric studies showed a progressive increase in the percentage of coronary rings with intimal thickening from day 1 to day 60 after transplantation. CONCLUSIONS: The progressive endothelial dysfunction reported in this model of accelerated coronary atherosclerosis after transplantation without immunosuppression involves preferentially the pertussis-toxin-sensitive Gi-protein-mediated pathway. Endothelium-independent relaxations are decreased at 60 days, as are all other endothelium-dependent relaxations. Decreased endothelium-dependent vasodilatation may contribute to the development of coronary graft vasculopathy. PMID- 10536677 TI - The influence of aging and aortic stiffness on permanent dilation and breaking stress of the thoracic descending aorta. AB - OBJECTIVE: To assess the influence of aging and aortic stiffness on the extent of irreversible deformation and breaking stress of the human thoracic aorta. METHODS: From 14 human heart valve donors without aortic disease (mean age 35 years, range 8-59 years), 14 intact segments of the thoracic descending aorta were studied within 48 h after cardiac arrest. In an experimental setup, the segments were submitted to increasing hydrostatic pressure loads, both statically and dynamically, while radius and wall thickness were monitored echocardiographically. Pressure-radius curves were constructed. Radius and wall thickness were determined at a pressure of 100 mmHg. Radius at elastin resting length and collagen recruitment pressure (Pcol, mmHg) were derived from the pressure-radius relationship and stress-strain curves were constructed to yield Young's moduli of elastin and collagen. Distensibility (D, mmHg-1) was determined while loading the segment with a sinusoidal pressure wave of 120/50 mmHg at both 0.5 and 1 Hz. Subsequently increasing static pressure loads of 400, 800, 1200 and 1600 mmHg were applied. After each pressure load, the increase in aortic radius at a pressure of 100 mmHg (Rinc) was determined. The experiment continued until rupture occurred and breaking stress (sigma break, N m-2) was calculated, donor age and aortic stiffness were correlated with Rinc and sigma break of the aortic segments. RESULTS: Mean breaking stress of the 14 segments was 2.7 x 10(6) N m-2. Breaking stress was negatively correlated with age (r2 = 0.66) and positively with D (r2 = 0.44) and with Pcol (r2 = 0.18). Seven segments survived a pressure load of 800 mmHg, in these vessels, the extent of irreversible dilation was positively correlated with age (r2 = 0.42) and negatively with D (r2 = 0.40) and Pcol (r2 = 0.40). CONCLUSION: Permanent deformation and rupture of the human thoracic aorta following pressure overload are influenced by age, distensibility and collagen recruitment pressure. PMID- 10536678 TI - Prostacyclin synthase gene transfer inhibits neointimal formation in rat balloon injured arteries without bleeding complications. AB - OBJECTIVE: This study was designed to compare the effects of prostacyclin synthase (PCS) gene transfer with those of a systemic infusion of beraprost sodium (BPS), a prostacyclin analogue, on vascular smooth muscle cell (VSMC) proliferation and neointimal formation after arterial injury. METHODS: PCS gene (3 or 30 micrograms) was transfected into rat balloon-injured carotid arteries by a non-viral lipotransfection method. BPS (100 or 300 micrograms/kg/day) was subcutaneously infused with osmotic pumps after the injury. LacZ gene (30 micrograms) was used as a control. VSMC proliferation was estimated by the bromodeoxyuridine (BrdU) index (BrdU-positive nuclei/total nuclei) at day 7. Neointimal formation was evaluated at day 14. Each treatment group had six rats. RESULTS: PCS gene transfer prevented the increase in intimal/medial area ratio (3 micrograms: 46.6%, 30 micrograms: 61.1% reduction; P < 0.05, P < 0.01, respectively), as did BPS 300 micrograms/kg/day (49.8% reduction; P < 0.05). BPS 100 micrograms/kg/day, however, had no effects on the ratio. PCS gene transfer and BPS 300 micrograms/kg/day significantly suppressed the BrdU index. BPS 300 micrograms/kg/day group had more frequent hematoma and longer bleeding time. There were no significant differences in blood pressure, heart rate, or urinary volume among all groups. CONCLUSION: Both PCS gene transfer and BPS 300 micrograms/kg/day reduced neointimal formation after arterial injury by inhibiting VSMC proliferation. PCS gene transfer may be a safer therapeutic modality against neointimal formation than a systemic infusion of BPS because the former method resulted in fewer bleeding complications. PMID- 10536679 TI - The isoprostane, 8-epi-PGF2 alpha, is accumulated in coronary arteries isolated from patients with coronary heart disease. AB - OBJECTIVE: In the present study we wanted to know whether 8-epi-PGF2 alpha, which belongs to the class of isoprostanes formed by free radical-mediated peroxidation of arachidonic acid and arachidonyl-containing phospholipids, is enriched in isolated coronary arteries of patients suffering from coronary heart disease (CHD, n = 23) who received allograft heart transplants as compared to vessels derived from patients with dilative cardiomyopathy (CMP, n = 19) or from healthy heart donors (controls, n = 6). METHODS: Sections from the isolated coronary arteries were analysed by semiquantitative immunohistochemistry by determining the area and intensity of positive reaction for 8-epi-PGF2 alpha in the vascular intima and media. In addition, the 8-epi-PGF2 alpha content was determined using a specific immunoassay after extraction and purification. RESULTS: The immunohistochemical results indicated that 8-epi-PGF2 alpha is significantly enriched in arteries from patients suffering from CHD as compared to CMP (P < 0.0001). In controls, significantly less immunostaining was observed. Furthermore, a significant positive correlation between semiquantitative immunohistochemistry and radioimmunological determination was observed too. CONCLUSIONS: From our findings we conclude that 8-epi-PGF2 alpha is especially accumulated in coronary arteries from CHD patients and therefore is likely to be involved in atherogenesis. PMID- 10536680 TI - Comment on "Effects of long-term angiotensin II AT-1 receptor blockade on survival, hemodynamics and cardiac remodeling in chronic heart failure in rats". PMID- 10536681 TI - A dilemma on Orchid Island. PMID- 10536682 TI - Aldosterone and the heart: towards a physiological function? PMID- 10536684 TI - A new, sympathetic look at KATP channels in the heart. PMID- 10536683 TI - Opioid peptides and the heart. AB - The heart is a complex neuroendocrine (opioids, NPY, VIP) or mechanoendocrine (ANP) organ. Opioid peptides and receptors are present in the heart and nerves such that they can easily modulate cardiac function. Cardiac opioids may have autocrine, paracrine and endocrine function. The challenge is to find the signal that turns them on and then what they do in the face of an overwhelmingly redundant system making knock-out technology hard to interpret. Determining the cardiac release of opioids in an intact system still requires a larger animal model. PMID- 10536685 TI - Implications of inhomogeneous distribution of IKS and IKr channels in ventricle with respect to effects of class III agents and beta-agonists. PMID- 10536686 TI - The ACE gene polymorphism: the good, the bad and the ugly. PMID- 10536687 TI - New look at myocardial infarction: toward a better aspirin. AB - The evidence for the formation of NO and of its oxidation products, as well as of prostacyclin and thromboxane by the infarcted heart muscle is reviewed. The importance of inflammatory cells, primarily macrophages of cardiac origin is documented. Because of its side effects on gastric mucosa and kidney by aspirin, several modifications of aspirin are currently being developed. These are based on eliminating their inflammatory effect by selective inhibition of COX-2, or by attaching an NO-delivering side chain to the aspirin molecule (NO-aspirin), or by combining two preparations, an NO donor with aspirin. NO-aspirins and the combination of an NO-donor with aspirin promise to be beneficial in the early stages of myocardial infarction. Unfortunately, the main beneficial effect of aspirin, that of inhibition of thrombus formation, is also the cause for its most dreaded complication, hemorrhagic stroke. None of the new aspirins is able to prevent this complication. PMID- 10536688 TI - Role of energy metabolism in the preconditioned heart--a possible contribution of mitochondria. AB - A brief period of ischemia and reperfusion has been shown to protect the myocardium against subsequent sustained ischemia and reperfusion injury, which is called "preconditioning". A great number of investigators have explored the mechanisms underlying this preconditioning-induced cardioprotection. This article dealt with possible mechanisms of energy metabolism and mitochondrial activity for preconditioning-induced cardioprotection. Particularly, the contribution of energy metabolites produced during a brief period of ischemia and reperfusion injury, as well as mitochondrial function that is modified by changes in mitochondrial ATPase activity, opening of mitochondrial ATP-dependent potassium channels and production of free radicals in mitochondria, to ischemic preconditioning is discussed. PMID- 10536689 TI - Multifactorial basis for coronary collateralization: a complex adaptive response to ischemia. AB - Angiogenesis and vasculogenesis are adaptive responses of the coronary collateral circulation to myocardial ischemia. This review focuses on the concerted action of growth factors, growth factor receptors, extracellular matrix, and inflammatory cellular responses to regulate angiogenesis and vasculogenesis in response to myocardial ischemia and alterations in shear stress. Therapeutic angiogenesis represents a novel approach to increase myocardial perfusion in patients with coronary artery disease and provides an opportunity to further clarify the mechanisms that regulate collateral development. Impairment of angiogenic adaptive responses to ischemia during disease states is an important subject for future investigation. PMID- 10536690 TI - Assessment of the functional status of heart failure in non ischemic dilated cardiomyopathy: an echo-dobutamine study. AB - BACKGROUND: The functional status of heart failure (HF) is conventionally evaluated by peak exercise oxygen consumption (VO2 max). Dobutamine echocardiography can be used to evaluate myocardial reserve. The aim of this study was to estimate the functional status of chronic HF in patients with dilated cardiomyopathy, by investigating the changes in echo-variables, as assessed by echo-dobutamine, in relation with VO2 max. METHODS AND RESULTS: A low infusion rate echo-dobutamine test (10 micrograms/kg/min) was performed in 30 patients with dilated cardiomyopathy and 1 h later VO2 max was measured. VO2 max (ranging from 7.6 to 23 ml/kg/min, mean 14.06 +/- 0.64 ml/kg/min) was correlated with the changes (values obtained after inotropic stimulation minus those obtained at baseline) in left ventricular end-systolic diameter (r:0.80, p:0.001), in left ventricular end-systolic posterior wall thickness (r:0.73, p:0.001) and in left ventricular heart-rate corrected mean velocity of circumferential fiber shortening (Vcfc)/end-systolic meridional wall stress ratio (r:0.64, p:0.0001). A negative correlation was found between VO2 max and the changes in end-systolic meridional wall stress (r: -0.76, p:0.001). After dobutamine infusion Vcfc/systolic meridional wall stress ratio increased in patients with VO2 max > 14 ml/kg/min but decreased in patients with VO2 max < 14 ml/kg/min (0.0001 +/- 0.0001 vs -0.0002 +/- 0.0003 circ x cm2/g x s, p:0.0001). End-systolic meridional wall stress was decreased in patients with VO2 max > 14 ml/kg/min but increased in patients with VO2 max < 14 ml/kg/min (-126.97 +/- 34.24 vs 205.77 +/- 56.71 g/cm2, p:0.0001). CONCLUSION: The changes in echo variables assessed by echo-dobutamine are well correlated with VO2 max and seem to be accurate for evaluating the functional status of chronic HF in patients with dilated cardiomyopathy. PMID- 10536691 TI - Regional expression of phospholamban in the human heart. AB - BACKGROUND: Several independent lines of evidence indicate that phospholamban (PLB) expression correlates positively with depression of force of contraction and duration of contraction in isolated cardiac preparations of several animal species. Here, we studied whether PLB levels correlate with attenuation of contractility and enhancement of contractile time parameters in different parts of the human heart. METHODS: Force of contraction was measured in isolated electrically driven atrial and ventricular preparations from human hearts. Ca(2+) uptake by human atrial and ventricular homogenates was assayed at different ionized Ca(2+)-concentrations. Protein expression of PLB and the sarcoplasmic Ca(2+)-ATPase (SERCA) was measured in homogenates by quantitative immunoblotting using specific antibodies. PLB mRNA expression was quantified in human cardiac preparations by Northern blot analysis. RESULTS: The duration of contraction in isolated preparations of human right ventricle (RV) was double that found in right atrial preparations (RA) (620 +/- 25 ms versus 308 +/- 15 ms). In RA, PLB expression was reduced by 44% at the protein level and by 34% at the mRNA level compared to RV. In contrast, the SERCA protein content was increased by 104% in RA compared to RV. Ca(2+)-uptake at low ionized Ca(2+)-concentration, where the inhibiting effect of PLB is maximal, amounted to 1.39 +/- 0.28 nmol Ca2+/mg protein in RA and to 0.62 +/- 0.09 nmol Ca2+/mg protein in RV (n = 6 both). CONCLUSIONS: It is suggested that duration of contraction is shorter in human atrium versus ventricle due to the combined effect of decreased PLB levels (which inhibits SERCA function) and increased SERCA levels. The lower relative ratio of PLB to SERCA leads to less inhibition of SERCA and increased Ca(2+)-uptake which enhances relaxation and contraction in human atrium. PMID- 10536692 TI - Chronic AT1 receptor blockade and angiotensin-converting enzyme (ACE) inhibition in (CHF 146) cardiomyopathic hamsters: effects on cardiac hypertrophy and survival. AB - OBJECTIVE: We have reported that angiotensin II AT1 receptors are upregulated and that there are no AT2 receptors in the ventricles of cardiomyopathic hamsters. Since the upregulation was present even when no histological lesions were detectable, these results suggested that angiotensin II plays a role in the genesis/maintenance of this pathology. A survival study was conducted to compare the effects of an angiotensin II AT1 receptor antagonist, losartan (L), to those of a placebo (P). Since the angiotensin-converting enzyme (ACE) inhibitor quinapril (Q) has been shown to have beneficial effects in this animal model, a Q group was included. METHODS: Male Syrian cardiomyopathic hamsters (CHF 146, n = 360) were orally administered P, low- (30 mg/kg/day) or high-dose (100 mg/kg/day) L, or Q (100 mg/kg/day), starting at day 50 of life. Inbred control hamsters (CHF 148, n = 180) were treated with P or L (100 mg/kg/day) as controls. Animals were sacrificed at intervals to evaluate cardiac hypertrophy. Kaplan-Meier analysis was performed to assess differences in survival. RESULTS: High-dose L had no effects on the survival of control hamsters. There was an unexpected dose dependent decrease in the survival of cardiomyopathics treated with L (low-dose, P = 0.14; high-dose, P = 0.0015) compared to an increase with Q (P = 0.0003). Cardiac hypertrophy compared to P was increased with L but significantly decreased with Q in cardiomyopathics. CONCLUSIONS: In this model, losartan did not improve survival compared to placebo and quinapril and, if anything, increased mortality. Our results suggest that AT1 receptor antagonists and ACE inhibitors are not necessarily equivalent or interchangeable in terms of their effects on cardiac hypertrophy and survival in selected progressive heart failure models. PMID- 10536693 TI - Right ventricular systolic function and ventricular interaction during acute embolisation of the left anterior descending coronary artery in sheep. AB - OBJECTIVE: Regional LV ischemia involving the septum affects LV systolic function and geometry. We investigated the effects of these changes on RV function and geometry. METHODS: In six closed-chest sheep end-systolic pressure-volume relationships (ESPVRs) were constructed from ventricular volumes, measured with magnetic resonance imaging (MRI) and matching intraventricular pressures, before and after selective embolisation of the left anterior descending coronary artery (LAD). The extent of myocardial ischemia was assessed post-mortem by coronary perfusion with Evans-Blue. Alterations in septal geometry were studied by measuring the curvature, segmental length and thickness of the septum in two midventricular (short-axis) MRI slices before and during ischemia. From these data, changes in LV and RV free wall segmental lengths were calculated. RESULTS: Selective embolisation of the LAD resulted in left ventricular ischemia (15 +/- 2.1% of the total LV) with 23% of the septum involved. Stroke volume did not change significantly, while LV systolic pressure decreased by 24 mmHg (p < 0.05). Although RV systolic function decreased to a significantly lesser extent than LV function (p < 0.01), systolic function of both ventricles diminished significantly as indicated by substantial rightward shifts of the ESPVRs: 121% for LV and 41% for RV (both p < 0.01). At mid-ventricular level and end-systole, the septum showed significant increases in its radius of curvature and segmental length (both p < 0.05), and a significant wall thinning (p < 0.01). Calculated end-systolic lengths of LV and RV free walls also increased, by 57 and 14% respectively. CONCLUSIONS: LAD embolisation not only results in a significantly diminished LV systolic function but also causes RV systolic function to decline significantly. Regional dysfunction by necessity entails global dysfunction as well. Analysis of ventricular geometry reveals that both the septum and the RV free wall increase their length, which plays an important role in the pathophysiology of diminished RV systolic function concomitant with reduced LV function. PMID- 10536694 TI - Preferential inhibition of lactate oxidation relative to glucose oxidation in the rat heart following diabetes. AB - OBJECTIVE: Alterations in myocardial metabolism occur early after the onset of diabetes suggesting that they may play a role in the development of cardiac dysfunction. Inhibition of myocardial pyruvate dehydrogenase (PDH), glucose transport and glycolysis have all been reported following diabetes. In vivo lactate is also a potential source of energy for the heart and its oxidation should not be affected by changes in glucose transport and glycolysis. Therefore, the objective of this study, was to test the hypothesis that following diabetes the inhibition of glucose oxidation would be greater than the inhibition of lactate oxidation. METHODS: Hearts from control and one-week-old diabetic rats were perfused with [1-13C]glucose (11 mmol/l) alone, [1-13C]glucose plus lactate (0.5 mmol/l) or glucose plus [3-13C]lactate (0.5 or 1.0 mmol/l) as substrates. Glucose and lactate oxidation rates were determined by combining 13C-NMR glutamate isotopomer analysis of tissue extracts with measurements of oxygen consumption. RESULTS: In diabetic hearts perfused with glucose alone, glucose oxidation was decreased compared to controls (0.31 +/- 0.08 vs. 0.71 +/- 0.11 mumoles/min/g wet weight; p < 0.05). Surprisingly, in hearts perfused with glucose plus 0.5 mmol/l lactate, there was no difference in glucose oxidation between control and diabetic groups (0.20 +/- 0.05 vs. 0.16 +/- 0.04 mumoles/min/g wet weight respectively). However, under these conditions lactate oxidation was markedly reduced in the diabetic group (0.89 +/- 0.18 vs. 0.24 +/- 0.05 mumoles/min/g wet weight; p < 0.05). At 1.0 mmol/l lactate oxidation was still significantly depressed in the diabetic group. CONCLUSION: There was a greater decrease in lactate oxidation relative to glucose oxidation in hearts from diabetic animals. These results demonstrate that diabetes leads to a specific inhibition of lactate oxidation independent of its effects on pyruvate dehydrogenase. PMID- 10536695 TI - Localization of pro-opiomelanocortin mRNA transcripts and peptide immunoreactivity in rat heart. AB - OBJECTIVE: alpha-Melanocyte-stimulating hormone (alpha-MSH), beta-endorphin and other pro-opiomelanocortin-(POMC) derived peptides have been detected in the heart, but it is uncertain whether they are synthesized by cardiomyocytes or by cardiac nerves innervating the heart. The objective of this study was to determine whether POMC peptides are synthesized by cardiomyocytes. METHODS: Pro opiomelanocortin peptides were localized in rat heart by immunohistochemistry using antisera against alpha-MSH, beta-endorphin and alpha N-acetyl-beta endorphin, the predominant POMC peptides found in heart. Pro-opiomelanocortin mRNA was investigated by reverse transcription polymerase chain reaction (RT-PCR) using primers that discriminate between full-length POMC mRNA and a 5' truncated POMC transcript that is presumed to be non-functional. RESULTS: alpha-Melanocyte stimulating hormone, beta-endorphin and alpha N-acetyl-beta-endorphin immunoreactivities were localized in atrial myocytes, particularly in the atrial appendages, but not to a significant extent in ventricular myocytes. Cardiac nerves were not immunostained. Atrial natriuretic peptide (ANP) immunoreactivity was similarly distributed in the adult heart. In neonatal heart, POMC-peptide and ANP immunoreactivities were present in both atrial and ventricular myocytes. RT PCR amplification showed that full-length POMC mRNA transcripts were present in both atrial and ventricular tissue and provide evidence that 5' truncated POMC mRNA is expressed in heart. CONCLUSIONS: These results support the hypothesis that cardiomyocytes synthesize POMC peptides. PMID- 10536696 TI - Chronic high-dose creatine feeding does not attenuate left ventricular remodeling in rat hearts post-myocardial infarction. AB - OBJECTIVE: In heart failure, cardiac energy metabolism is compromised. The failing myocardium is characterized by reduced contents of both phosphorylated (phosphocreatine) and non-phosphorylated (free) creatine content as well as decreased energy reserve via creatine kinase (creatine kinase reaction velocity). These changes may contribute to cardiac dysfunction. The purpose of the present study was to determine whether chronic feeding with high-dose dietary creatine prevents the derangement of energy metabolism and the development of left ventricular remodeling in a rat model of heart failure, i.e. post-myocardial infarction (MI). METHODS AND RESULTS: Rats were subjected to sham operation or left coronary artery ligation. Surviving rats were fed with 0% (untreated) or 3% creatine (related to weight of diet) for 8 weeks. Creatine feeding increased serum creatine levels significantly approximately 2-fold. Thereafter, hearts were isolated, perfused and left ventricular pressure-volume curves obtained. Steady state and dynamic (CK reaction velocity) high-energy phosphate metabolism was determined with 31P NMR spectroscopy. In both MI groups (treated n = 8, untreated n = 7), pressure-volume curves were shifted right- and downward compared to both sham groups (treated n = 5, untreated n = 7), i.e. creatine had no effect on left ventricular remodeling. Likewise, similar reductions of phosphocreatine, free creatine and creatine kinase reaction velocity (untreated sham 12.0 +/- 0.7 mmol/lxs; untreated MI 7.8 +/- 0.7*; treated sham 13.6 +/- 1.0; treated MI 7.2 +/ 1.1*; *p < 0.025 sham vs. MI) were found in both MI groups. CONCLUSIONS: Chronic creatine feeding of post-MI rats is ineffective in preventing the functional and energetic derangements occurring post-MI. Inspite of increased serum creatine levels, neither the normal nor the failing heart accumulates additional creatine. PMID- 10536697 TI - Modulation of norepinephrine release by ATP-dependent K(+)-channel activators and inhibitors in guinea-pig and human isolated right atrium. AB - OBJECTIVE: The aim of this study was to show, whether ATP sensitive K+ channels (KATP channels), are involved in the modulation of norepinephrine (NE) release from the sympathetic nerves innervating the guinea-pig and human right atrium. METHODS: The resting and stimulation-evoked release of [3H]norepinephrine ([3H]NE) was measured from the isolated guinea-pig and human right atrium and the effect of activators and inhibitors of ATP sensitive K+ channels was studied. RESULTS: Cromakalim (30-300 microM), a KATP channel-agonist decreased concentration-dependently the stimulation-evoked release of NE from the guinea pig atrium, an effect, antagonized by glibenclamide, a KATP channel-antagonist (30 microM). Diazoxide (30-300 microM), another activator of the KATP channels reduced the resting release of NE, and also attenuated the evoked release at a single concentration (100 microM), and this latter action was also counteracted by glibenclamide (30 microM). Pinacidil, increased dose-dependently the resting and stimulation-evoked release of NE in a glibenclamide-sensitive manner and reversed the inhibitory effect of cromakalim (100 microM), suggesting that it acts as an antagonist. Glibenclamide (30-300 microM), by itself enhanced the stimulation-evoked release of [3H]NE, and also increased the resting release of NE. On the other hand, 5-hydroxydecanoate, an ischemia-selective inhibitor of cardiac KATP channels did not change NE release. Adenosine, (30-300 microM), an A1-receptor agonist, clonidine (3 microM), an alpha 2-adrenoceptor agonist and oxotremorine, a muscarinic receptor agonist (30 microM) all reduced the evoked release of [3H]NE, but these effects were not modified by glibenclamide (300 microM), indicating that neuronal adenosine (A1), adrenergic (alpha 2) and muscarinic (M3) receptors do not act on KATP channels. In the human right atrium, cromakalim, and diazoxide did not affect significantly the release of [3H]NE. However, glibenclamide (30-300 microM) and pinacidil (30-300 microM) enhanced dose-dependently the evoked-release of NE, and pinacidil also augmented the resting release. CONCLUSIONS: Our results indicate that sympathetic nerve endings of the human and guinea-pig atrium are endowed with ATP-sensitive K+ channels. These channels responded to agonists and antagonists under the experimental conditions applied and they could modulate the release of NE thereby affecting the autonomic control of cardiac function under various physiological and pathophysiological conditions. PMID- 10536698 TI - Heterogeneous distribution of the two components of delayed rectifier K+ current: a potential mechanism of the proarrhythmic effects of methanesulfonanilideclass III agents. AB - OBJECTIVE: To elucidate the regional difference of the K+ current blocking effects of methanesulfonanilide class III agents. METHODS: Regional differences in action potential duration (APD) and E-4031-sensitive component (IKr) as well as -insensitive component (IKs) of the delayed rectifier K+ current (IK) were investigated in enzymatically isolated myocytes from apical and basal regions of the rabbit left ventricle using the whole-cell clamp technique. RESULTS: At 1 Hz stimulation, APD was significantly longer in the apex than in the base (223.1 +/- 10.6 vs. 182.7 +/- 14.5 ms, p < 0.05); application of 1 microM E-4031 caused more significant APD prolongation in the apex than in the base (32.5 +/- 6.4% vs. 21.0 +/- 8.8%, p < 0.05), resulting in an augmentation of regional dispersion of APD. In response to a 3-s depolarization pulse to +40 mV from a holding potential of 50 mV, both IK tail and IKs tail densities were significantly smaller in apical than in basal myocytes (IK: 1.56 +/- 0.13 vs. 2.09 +/- 0.21 pA/pF, p < 0.05; IKs: 0.40 +/- 0.15 vs. 1.43 +/- 0.23, p < 0.01), whereas IKr tail density was significantly greater in the apex than in the base (1.15 +/- 0.13 vs. 0.66 +/- 0.11 pA/pF, p < 0.01). The ratio of IKs/IKr for the tail current in the apex was significantly smaller than that in the base (0.51 +/- 0.21 vs. 3.09 +/- 0.89; p < 0.05). No statistical difference was observed in the voltage dependence as well as activation and deactivation kinetics of IKr and IKs between the apex and base. Isoproterenol (1 microM) increased the time-dependent outward current of IKs by 111 +/- 8% during the 3-s depolarizing step at +40 mV and its tail current by 120 +/- 9% on repolarization to the holding potential of -50 mV, whereas it did not affect IKr. CONCLUSIONS: The regional differences in IK, in particular differences in its two components may underlie the regional disparity in APD, and that methanesulfonanilide class III antiarrhythmic agents such as E-4031 may cause a greater spatial inhomogeneity of ventricular repolarization, leading to re-entrant arrhythmias. PMID- 10536699 TI - Cultured neonatal rat cardiac myocytes and fibroblasts do not synthesize renin or angiotensinogen: evidence for stretch-induced cardiomyocyte hypertrophy independent of angiotensin II. AB - OBJECTIVE: The hypertrophic response of cardiomyocytes exposed to mechanical stretch is assumed to depend on the release of angiotensin (Ang) II from these cells. Here we studied the synthesis of renin-angiotensin system (RAS) components by cardiac cells under basal conditions and after stretch. METHODS: Myocytes and fibroblasts were isolated by enzymatic dissociation from hearts of 1-3-day-old Wistar rat strain pups, grown for 1 day in serum-supplemented medium and then cultured in a chemically defined, serum-free medium. Medium and cell lysate were collected 5 days later or after exposure of the cells to cyclic stretch for 24 h. Prorenin, renin and angiotensinogen were measured by enzyme-kinetic assay; Ang I and Ang II were measured by radioimmunoassay after SepPak extraction and HPLC separation. RESULTS: Prorenin, but none of the other RAS components, could be detected in the medium of both cell types. However, its levels were low and the Ang I-generating activity corresponding with these low prorenin levels could not be inhibited by the specific rat renin inhibitor CH-732, suggesting that it was most likely due to bovine and/or horse prorenin sequestered from the serum containing medium to which the cells had been exposed prior to the serum-free period. When incubated with Ang I, both myocytes and fibroblasts generated Ang II in a captopril-inhibitable manner. Myocyte and fibroblast cell lysates did not contain prorenin, renin, angiotensinogen, Ang I or Ang II in detectable quantities. Stretch increased myocyte protein synthesis by 20%, but was not accompanied by Ang II release into the medium. CONCLUSION: Cardiac myocytes and fibroblasts do not synthesize renin, prorenin or angiotensinogen in concentrations that are detectable or, it not detectable, high enough to result in Ang II concentrations of physiological relevance. These cells do synthesize ACE, thereby allowing the synthesis of Ang II at cardiac tissue sites when renin and angiotensinogen are provided via the circulation. Ang II is not a prerequisite to observe a hypertrophic response of cardiomyocytes following stretch. PMID- 10536701 TI - Angiotensin II receptor blockade during gestation attenuates collagen formation in the developing rat heart. AB - OBJECTIVE: Fetal cardiac development includes rapid formation of a three dimensional collagen network, composed mainly of type I and III fibrillar collagens. Collagen fibrils have been found in cardiac jelly at very early stages of cardiac development and are thought to have structural and functional properties. In adult rat cardiac tissue, angiotensin II (AngII) via AT1 receptor binding and AngII-regulated expression of transforming growth factor beta-1 (TGF beta 1) each upregulate collagen transcription. AT1 and AT2 receptor subtypes are developmentally regulated; both have been localized in fetal tissue where the AT2 receptor is considered a determinant of morphogenesis. We sought to determine whether blockade of either receptor would result in attenuation of collagen mRNA expression and fibrillar collagen accumulation and alter TGF-beta 1 mRNA expression in the developing fetal heart examined at birth. METHODS: Pregnant rats were treated either with an AT1 receptor antagonist losartan or an AT2 receptor antagonist PD123319 and compared with untreated age-matched controls. Offspring were studied within 24 h of birth. Type I and type III collagen mRNA expression, as well as TGF-beta 1 mRNA expression, were examined by in situ hybridization. Collagen concentration was determined spectrophotometrically by picrosirius red staining and type I and III collagens were detected by immunoblotting. RESULTS: We found: (1) comparable birth weights in control and PD123319-treated animals, but reduced body weight in newborn losartan-treated animals; (2) compared to untreated animals, type I collagen and TGF-beta 1 mRNA expression in cardiac tissue were each equally reduced in both losartan and PD123319-treated animals; (3) increased type III collagen mRNA expression in both PD123319- and losartan-treated groups; and (4) a significant decrease in total soluble cardiac collagen concentration in both losartan and PD123319-treated groups, confirmed by attenuated immunoreactivity of type I and III collagens in whole heart extracts by Western blotting. CONCLUSIONS: The results of these pharmacologic interventions suggest AngII receptors are expressed in cardiac tissue during gestation, where both AT1 and AT2 receptors are involved in the regulation of type I and III collagen expression and structural protein accumulation. These effects appear to be mediated, in part, by attenuated cardiac TGF-beta 1 levels. The marked decrease in newborn cardiac collagen content has yet undefined functional consequences. PMID- 10536700 TI - Positive inotropic effect of insulin-like growth factor-1 on normal and failing cardiac myocytes. AB - OBJECTIVE: The acute administration of growth hormone (GH) or insulin-like growth factor-1 (IGF-1) improves cardiac performance, possibly contributing to the beneficial effects of GH therapy on heart failure (HF). GH can induce the production of IGF-1 and thus the actions of GH may be mediated through its IGF-1 induction. However, these effects have not yet been demonstrated in failing hearts and the cellular basis of GH or IGF-1-induced inotropic effects remains unknown. We examined the direct effects of GH and IGF-1 on the contractile function and intracellular calcium ([Ca2+]i) homeostasis in normal and failing myocytes. METHODS: To determine whether GH and IGF-1 have a direct effect on myocardial contractility and whether the GH/IGF-1-induced effect was the results of changes in Ca2+ activation, cell shortening and [Ca2+]i transient were simultaneously measured in the left ventricular myocyte preparations, isolated from normal and rapid pacing-induced HF dogs. RESULTS: Basal shortening of HF myocytes was reduced by 64% (p < 0.01). In normal and HF myocytes, GH (0.4-40 x 10(-3) IU/ml) had no effect on either cell shortening or [Ca2+]i transients. In normal myocytes, IGF-1 exerted a positive inotropic effect in a time- and dose dependent manner (25-500 ng/ml), associated with a parallel increase of [Ca2+]i transient amplitude. IGF-1 increased the shortening magnitude in normal (121 +/- 5% increase from baseline, p < 0.05) and HF (118 +/- 4% increase from baseline, p < 0.05) myocytes. It also increased [Ca2+]i transient amplitude in normal and HF cells by 124 +/- 4 and 125 +/- 7%, respectively. The percent increase of cell shortening and [Ca2+]i transient amplitude was comparable between normal and HF myocytes. Furthermore, IGF-1 did not shift the trajectory of the relaxation phase in the phase-plane plots of cell length vs. [Ca2+]i, indicating that it did not change myofilament Ca2+ sensitivity. CONCLUSIONS: In both normal and HF conditions, IGF-1 exerted an acute direct positive inotropic effect in adult cardiac myocytes by increasing the availability of [Ca2+]i to the myofilaments, possibly explaining the beneficial effect of GH on HF. PMID- 10536702 TI - Role of autacoids in cardiovascular colapse in anaphylactic shock in anaesthetized dogs. AB - OBJECTIVE: In anaphylactic shock (AS), the relative effects of the autacoids including histamine, prostaglandins (prost), and leukotrienes (leuk) on causing cardiovascular collapse and the extent to which receptor blocking agents and pathway inhibitors may prevent this collapse are not clear. METHODS: In randomized design, we investigated whether blockade of histamine H1, H2, and H3 receptors or inhibition of the cyclooxygenase (cyclo) and lipoxygenase pathways (lipox) prevented AS in ragweed sensitized dogs. Seven dogs were studied under pentobarbital anesthesia in which the treatment studies were approximately 2 weeks apart. RESULTS: During H1 receptor blockade, the decreases in blood pressure and cardiac output otherwise observed in AS were attenuated (P < 0.05) and the release of prost, thromboxanes, and leuk were reduced as compared with nontreatment studies. Cyclo inhibition also attenuated cardiovascular collapse and mediator release in AS, but the other treatments showed no effects. CONCLUSION: H1 receptor blockade and cyclo may attenuate cardiovascular shock in AS. These agents inhibit autacoid release from mast cells in addition to any specific receptor blocking and pathway inhibition effects. PMID- 10536704 TI - Hypertensive left ventricular remodeling and ACE-gene polymorphism. AB - OBJECTIVE: To evaluate the relationship between ACE-gene polymorphism and left ventricular geometry in never treated hypertensives. METHODS: We enrolled 200 hypertensive outpatients that underwent clinical and ambulatory blood pressure measurements, echocardiographic evaluation and analysis for insertion (I)/deletion (D) polymorphism by PCR. Patients with normal or increased (> 125 g/m2 in males and > 110 g/m2 in females) left ventricular mass were considered to have concentric remodeling or concentric left ventricular hypertrophy if their relative wall thickness was > or = 0.45. RESULTS: The left ventricular mass index values (g/m2) were 136 +/- 30 in DD genotype, 124 +/- 26 in ID genotype, and 116 +/- 20 in II genotype (DD vs. ID P < 0.005; DD vs. II P < 0.05), and were unrelated to blood pressure. Ninety-six patients presented left ventricular hypertrophy (48.0%): 51 with concentric and 45 with eccentric hypertrophy. The eccentric left ventricular hypertrophy was detected in 32 (36.8%) DD patients, in ten (10.5%) ID patients (P < 0.05), and in three (16.6%) II patients. The relative septal thickness was 0.43 +/- 0.09 in DD genotype, 0.45 +/- 0.08 in ID genotype, and 0.43 +/- 0.10 in II genotype. In DD and ID genotypes, the relative posterior wall thickness (0.37 +/- 0.07 vs. 0.41 +/- 0.07; P < 0.0001) and the end-diastolic left ventricular internal dimension (52.8 +/- 3.3 mm vs. 48.3 +/- 2.8 mm; P < 0.0001) were statistically different. CONCLUSIONS: The DD genotype of the ACE-gene is associated with an increased left ventricular mass and with a significantly higher prevalence of eccentric left ventricular hypertrophy, when compared to ID genotype. PMID- 10536703 TI - Platelet aggregatory response to platelet activating factor (PAF), ex vivo, and PAF-acetylhydrolase activity in patients with unstable angina: effect of c7E3 Fab (abciximab) therapy. AB - OBJECTIVE: Platelet activation and aggregation is a dominant feature in the pathophysiology of unstable angina. The final step of platelet aggregation is mediated through the platelet integrin glycoprotein IIb/IIIa (GP IIb/IIIa), while abciximab (ReoPro) is one of the most potent inhibitors of this receptor. Platelet-activating factor (PAF) is a potent platelet agonist which is degraded and inactivated by PAF-acetylhydrolase (PAF-AH). The plasma form of PAF-AH is associated with lipoproteins. We studied the platelet response to the aggregatory effect of PAF, ex vivo, in relation to the plasma PAF-AH activity in 32 patients with unstable angina, as well as the effect of abciximab therapy on the above parameters. METHODS: Thirty two patients with unstable angina and 25 sex- and age matched healthy controls participated in the study. On the day of admission (day 1) 17 patients received a bolus of abciximab (0.25 mg/kg) followed by a 12-h infusion (10 micrograms/min). Platelet aggregation to both PAF and ADP, in platelet rich plasma, was successively studied in both patients receiving abciximab or remaining untreated. The plasma and HDL-associated PAF-AH activity was also determined at the same times. RESULTS: In the untreated patients, the PAF EC50 values were significantly lower on the day of admission, whereas the maximal percentage of aggregation was significantly higher compared to controls (p < 0.01 for both comparisons). Similar behaviour of the platelets was observed in the aggregatory effect of ADP. This aggregatory response was not significantly altered 4 days, 7 days or 1 month afterwards. In the 17 patients who received abciximab, platelet aggregation to both PAF and ADP was inhibited by 90 +/- 5 and 96 +/- 3%, respectively, 1 h after bolus. At 2 and 3 days after treatment, platelet aggregation to both agonists was significantly recovered being similar to controls. However, it was fully restored 6 days after bolus, still being significantly higher compared to controls (p < 0.01 for PAF and p < 0.003 for ADP). The total plasma PAF-AH activity in both patient groups was not different from that of controls, whereas the HDL-associated PAF-AH activity was significantly lower. The total plasma or HDL-associated enzyme activity was not altered at any time interval studied, and it was not influenced by abciximab. CONCLUSIONS: The increased aggregatory response of platelets to PAF and the low plasma levels of HDL-cholesterol and HDL-associated PAF-AH activity in patients with unstable angina may contribute to the severe atherosclerosis and to acute thrombosis found in these patients. Abciximab therapy may protect platelets from PAF action in vivo the first days after drug administration, but it fails to permanently restore the enhanced aggregatory response observed. PMID- 10536705 TI - Enhanced acetylcholine and P2Y-receptor stimulated vascular EDHF-dilatation in congestive heart failure. AB - OBJECTIVE: Congestive heart failure (CHF) is accompanied by impaired peripheral blood flow and endothelial dysfunction with decreased release of nitric oxide (NO). Strong evidence supports the existence of another vasodilatory substance, endothelium derived hyperpolarising factor (EDHF), which has not previously been studied in CHF. METHOD: CHF was induced by left coronary artery ligation resulting in a reproducible myocardial infarction in Sprague Dawley rats. Vasodilatory responses to acetylcholine and extracellular nucleotides (ATP, ADP beta S, ADP and UTP) were examined in cylindrical segments of the mesenteric artery, precontracted with noradrenaline. The combined NO- and EDHF-dilatation (after inhibition of cyclo-oxygenase pathways) was called "total dilatation", as indomethacin had only minor effects in this system. NO-dilatation was studied in segments pretreated with indomethacin and the potassium channel inhibitors charybdotoxin (10(-7.5) M) and apamin (10(-6) M), while EDHF-dilatations were studied in the presence of indomethacin (10(-5) M) and L-NOARG (10(-3.5) M). RESULTS: EDHF-dilatations in CHF were strongly up-regulated for ACh (36% vs. 73%; sham vs. CHF operated rats), ADP beta S (10% vs. 42%), ADP (0% vs. 21%) and UTP (3% vs. 35%). These dilatations were abolished by a combination of charybdotoxin and apamin, confirming that they were mediated by EDHF. The NO-dilatations on the other hand were down-regulated in CHF as compared to sham operated rats for ACh (93% vs. 76%; sham vs. CHF operated rats), ADP beta S (61% vs. 37%). ADP (60% vs. 30%), ATP (49% vs. 34%) and UTP (65% vs. 47%), while a minor decrease was seen in the total dilatation for ACh (87% vs. 75%; sham vs. CHF operated rats), ADP beta S (47% vs. 42%), ADP (59% vs. 39%), ATP (52% vs. 39%) and UTP (59% vs. 44%). CONCLUSION: In this model of non-atherosclerotic CHF there was a minor decrease in the total dilatation and a marked down-regulation of the NO-mediated dilatation, while the EDHF-dilatation was up-regulated. Increased EDHF-activity in CHF may represent a compensatory response to decreased NO-activity to preserve endothelial function and tissue perfusion. PMID- 10536706 TI - Activated platelets and leucocytes cooperatively stimulate smooth muscle cell proliferation and proto-oncogene expression via release of soluble growth factors. AB - BACKGROUND: Previous studies indicate that platelets and leucocytes might contribute to the development of neointimal hyperplasia following arterial injury. The present study was aimed at further investigating the role of platelets and leucocytes, alone or in combination, in promoting vascular smooth muscle cell (SMC) proliferation in vitro, focusing on the relative contribution of different soluble growth factors released by these cells, and on the ability to induce proto-oncogene expression, such as c-fos. METHODS: SMCs from rabbit aortas, made quiescent by serum deprivation, were stimulated with either activated platelets, leucocytes, or both, separated from SMCs by a membrane insert. SMC proliferation was evaluated by measuring the incorporation of 3H thymidine. The relative contribution of different platelet-derived mediators to SMC growth was evaluated by adding either ketanserin, a 5-HT2 receptor antagonist, R68070, a TxA2 receptor antagonist, BN52021, a platelet activating factor (PAF) receptor antagonist, and trapidil, a platelet derived growth factor (PDGF) receptor antagonist. The role of different leucocyte sub-populations (neutrophils and monocytes + lymphocytes) was also determined in additional experiments. RESULTS: SMC proliferation was significantly increased by activated platelets to 360 +/- 9% of control values (P < 0.05). This effect was reduced by ketanserin, R68070, BN 52021 or trapidil. Whole leucocytes, neutrophils or lymphocytes + monocytes also increased SMC proliferation with respect to control experiments. Simultaneous stimulation of SMCs by platelets and whole leucocytes was associated with a significant greater increase in SMC proliferation as compared to SMC stimulated with platelets or leucocytes alone. c-fos expression, almost undetectable in unstimulated SMCs, was markedly increased by activated platelets or leucocytes. CONCLUSIONS: Activated platelets promote SMC proliferation in vitro via release of soluble mediators, including serotonin, thromboxane A2 PAF and PDGF; activated leucocytes also induce a significant SMC proliferation and exert an additive effect when activated together with platelets; SMCs stimulated with activated platelets and leucocytes show an early expression of the proto-oncogene c-fos. PMID- 10536707 TI - Congestive heart failure induces downregulation of P2X1-receptors in resistance arteries. AB - OBJECTIVE: Congestive heart failure (CHF) is accompanied by enhanced peripheral sympathetic nerve activity, increased vascular resistance and impaired peripheral blood flow. Besides noradrenaline and neuropeptide Y, the sympathetic nervous system also releases ATP, which has contractile effects mediated by different subtypes of P2-receptors on the vascular smooth muscle cells. The present study was designed to examine postsynaptic changes of the contractile responses to ATP and other extracellular nucleotides in CHF. METHODS: CHF was induced by left coronary artery ligation resulting in a reproducible myocardial infarction in Sprague-Dawley rats. Contractile responses were examined in cylindrical segments of aorta and the mesenteric artery after endothelium removal. To determine if an altered response was regulated on the transcriptional level, competitive reverse transcription polymerase chain reaction (RT-PCR) was used to estimate the amount of P2X1-receptor mRNA. RESULTS: ATP, which is both a P2X1- and a P2Y-receptor agonist, induced a weaker contraction in the mesenteric artery from CHF as compared to sham operated rats. A decrease in both potency and maximum contraction was shown for the selective P2X1-receptor agonist, alpha beta-MeATP, in the mesenteric artery (pEC50 = 6.04 vs. 5.76, Cmax = 57% vs. 33%, sham vs. CHF operated rats), but not in the aorta. Competitive RT-PCR also revealed decreased P2X1-receptor mRNA levels in CHF operated rats in the mesenteric artery (9106 x 10(3) vs. 714 x 10(3) molecules/microgram, sham vs. CHF operated rats), while it remained unaltered in the aorta. To study the P2Y-receptor induced contractile effects, the P2X1-receptors were first desensitised with alpha beta-MeATP (10(-5) M for 8 min). After P2X1-receptors desensitisation, UTP and UDP induced strong contractions in both the mesenteric artery and in the aorta, while ATP and ADP were much less effective. These contractions were not altered by CHF, indicating that vascular contraction mediated by P2Y-receptors are unaffected by CHF. CONCLUSION: CHF induces downregulation of P2X1-receptor stimulated contraction in the mesenteric artery depending on decreased mRNA synthesis for the receptor, while the P2Y-receptor activity remains unchanged. Downregulation of P2X1 receptors appears to be specific for peripheral resistance arteries. This may represent a compensatory response to enhanced peripheral sympathetic nerve activity and increased vascular resistance in CHF. PMID- 10536708 TI - Adenosine plasma concentration in pulmonary hypertension. AB - OBJECTIVE: In this study, we sought to appreciate the role of adenosine in the regulation of pulmonary vascular tone, especially in the case of clinical pulmonary hypertension, by investigating the relationship between endogenous plasma adenosine levels and pulmonary artery vasoconstriction. METHODS: Adenosine plasma concentrations, were measured simultaneously in the distal right pulmonary artery and in the femoral artery, both at steady state (room air) and during pure oxygen inhalation. Three clinical situations were considered: (1) normal hemodynamics [7 control subjects, mean pulmonary artery pressure (MPAP) = 18.5 +/ 1 mm Hg], (2) moderate pulmonary hypertension secondary to chronic obstructive pulmonary disease (COPD), (8 patients, MPAP = 31 +/- 3 mm Hg), (3) severe primary pulmonary hypertension (PPH), (8 patients, MPAP = 70 +/- 5 mm Hg). RESULTS: In every instance, adenosine evaluated by HPLC was higher in the pulmonary than in the systemic circulation. For room air, adenosine plasma concentrations were significantly lower in COPD (0.49 +/- 0.16 mumol l-1) and PPH patients (0.45 +/- 0.14 mumol l-1) than in controls (1.26 +/- 0.12 mumol l-1). During O2 administration, adenosine plasma concentrations all decreased but more so in COPD and PPH patients. The significant correlations between adenosine plasma concentrations and both pulmonary vascular resistance and PvO2, in controls, were not found in COPD or PPH patients. CONCLUSION: The adenosine plasma concentrations in the pulmonary circulation of PPH and COPD patients are low, and may contribute to pulmonary artery hypertension. PMID- 10536709 TI - Opposing adrenergic actions of intravenous metformin on arterial pressure in female spontaneously hypertensive rats. AB - OBJECTIVE: Intravenous (i.v.) injection of the antidiabetic drug metformin rapidly lowers mean arterial pressure (MAP) in spontaneously hypertensive rats (SHR). However, if autonomic ganglia or alpha-adrenoceptors are first blocked then metformin rapidly raises MAP in SHR. This study was conducted to further characterize the adrenergic mechanisms of these opposing i.v. actions of the drug. METHODS: Conscious, undisturbed female SHR with indwelling vascular catheters were used to measure acute effects of i.v. metformin (100 mg/kg; before and after sustained ganglionic blockade, GB, with chlorisondamine, 5 mg/kg) on: (1) circulating levels of catecholamines, (2) MAP after pharmacologic modulation of beta- as well as alpha-adrenoceptors and (3) all the above in the absence as well as presence of the adrenal medulla. RESULTS: Plasma norepinephrine (NE) and epinephrine (E) levels (pg/ml) were rapidly increased by i.v. metformin (8 SHR, p < 0.05) both before GB (delta NE = +146 +/- 41; delta E = +119 +/- 31) and after GB (delta NE = +79 +/- 24; delta E = +120 +/- 32). Similar increases in plasma NE (though not E) were seen in SHR without adrenal medullae. Blockade of beta adrenoceptors with propranolol (pro; 3 mg/kg, 8 SHR) enhanced the rapid depressor response to i.v. metformin before GB (delta MAP, mmHg: -38 +/- 4 with pro vs -17 +/- 3 without pro; p < 0.05) and attenuated the rapid pressor response to i.v. metformin after GB (delta MAP, mmHg: +8 +/- 3 with pro vs +30 +/- 4 without pro; p < 0.05). Results were similar in SHR without adrenal medullae. Finally, if baseline MAP under GB was raised back to hypertensive levels with i.v. infusion of either NE or phenylephrine then i.v. metformin did not raise but rather reduced MAP in SHR. CONCLUSION(S): The acute depressor action of i.v. metformin in female SHR (1) is most likely due to a direct vasodilator action which includes inhibition of alpha-receptor-mediated vasoconstriction and (2) is buffered by an acute beta-receptor-mediated pressor action likely due to a direct metformin-induced release of NE from postganglionic sympathetic nerve endings. PMID- 10536710 TI - Cardiovascular diseases, oxidative stress and antioxidants: the decisive role of coenzyme Q10. PMID- 10536711 TI - On the role of coenzyme Q10 in cardiovascular diseases. PMID- 10536712 TI - Does growth hormone reduce fibrosis? PMID- 10536713 TI - Growth hormone and interstitial fibrosis. PMID- 10536714 TI - World Federation of Neurosurgical Societies and European Association of Neurosurgical Societies (final version: August 1998). Good Practice: A Guide for Neurosurgeons. PMID- 10536715 TI - Surgical treatment of pineal region tumours through the occipital transtentorial approach: evaluation of the effectiveness of intra-operative micro-endoscopy combined with neuronavigation. AB - OBJECT: To determine the efficacy and accuracy of surgically-assisted systems including endoscopy combined with neuronavigation in the treatment of pineal region tumours through the occipital transtentorial approach, an evaluation of thirty-one patients undergoing surgery was performed over a 10-year period. METHOD: The study was performed in 2 parts. The surgical approach to the pineal region was the same in the two parts, but in part 2 a smaller craniotomy window was used. Part 1 (from March 1989 to March 1997) included 15 patients who underwent surgical removal of pineal region tumours without using assisted systems; four out of the fifteen patients had surgery-related complications, including seizure and hemianopsia. Part 2 (from April 1997 to February 1999) included 16 patients who underwent surgical treatment by the same surgical team and with assisted systems; all 16 patients had excellent outcomes, with no complications. CONCLUSIONS: Although this study was the first specifically to examine the efficacy of endoscopy combined with neuronavigation in the treatment of pineal region tumours, our findings suggest that these systems are very useful, safe, and accurate in evaluating the primary tumour and surrounding anatomy as well as in determining operative strategy, such as the location and size of the scalp incision, craniotomy, and the extent of surgical removal. Therefore, we conclude that the addition of endoscopy combined with neuronavigation to standard surgical procedures can improve the outcome of surgical treatment of pineal region tumours. PMID- 10536716 TI - Gamma knife radiosurgery for glomus jugulare tumours. AB - The aim of this clinical study was to determine the tumour control rate, clinical outcome and complication rate following gamma knife treatment for glomus jugulare tumours. Between May 1992 and May 1998, 13 patients with glomus tumours underwent stereotactic radiosurgical treatment in our department. The age of these patients ranged from 21 to 80 years. The male:female ratio was 2:11. Six patients had primary open surgery for partial removal or recurrent growth and subsequent radiosurgical therapy. Radiosurgery was performed as primary treatment in 7 cases. The median tumour volume was 6.4 cm3 (range: 4.6-13.7 cm3). The median marginal dose applied to an average isodose volume of 50% (30-50%) was 13.5 Gy (12-20 Gy). In 10 patients, a total of 48 MRI and CT follow-up scans were available. The remaining three patients have been excluded from the postradiosurgical evaluation since the observation time (t < 12 months) was too short or patients were lost to follow up. The median interval from Gamma Knife treatment to the last radiological follow-up was 37.6 months (5-68 months). In 4 patients (40%) decreased tumour volumes were observed and in 6 cases (60%) the tumour size remained unchanged. Neurological follow-up examinations revealed improved clinical status in 5 patients (50%), a stable neurological status in 5 patients (50%) and no complications occurred. According to our preliminary experience Gamma Knife radiosurgery represents an effective treatment option for glomus jugulare tumours. PMID- 10536717 TI - Factors of surgical outcome in tumoural epilepsy. AB - OBJECTIVES: The purposes of the study were the assessment of the role of surgery in the suppression of epilepsy due to low-grade primitive cerebral tumours and the search for factors relevant to the surgical outcome. PATIENTS AND METHODS: Forty-eight patients with epilepsy due to low-grade supratentorial cerebral tumours were considered. They presented drug-resistant daily to monthly seizures since for least one year (mean 7 yrs). Twenty-four patients underwent a combined tumour and epileptogenic zone resection ("epilepsy surgery") and 24 tumour resection alone ("lesionectomy"). The surgical outcome was evaluated two years after surgery. Several variables related to the characteristics of the epilepsy, the tumour and surgery, were considered for a possible association with the outcome. Statistical analyses were performed. RESULTS: Seizure freedom, including aura, was obtained in 35 patients (72.9%). Mild permanent complications occurred in 6 cases. Seizure suppression was significantly associated with complete tumour resection (post-surgical CT or MRI) and relatively low presurgical seizure frequency; it was also related, though not significantly, to small tumour size and histological grade I. The surgical outcome was only slightly better following "epilepsy surgery" than "lesionectomy". However: i) the extent of tumour resection was not relevant regarding the "epilepsy surgery" outcome, while significantly influencing the outcome after "lesionectomy"; ii) the presurgical frequency of seizures and, to a less extent, the tumour size, had a higher influence on the outcome after "lesionectomy". CONCLUSION: Long-lasting and drug resistant epilepsy due to cerebral tumours can be suppressed surgically in the majority of cases. The extent of tumour resection and the frequency of the seizures are the most relevant prognostic factors. Both "epilepsy surgery" and "lesionectomy" can provide good results. However, the two approaches should not be regarded as interchangeable: a choice of the approach based on the characteristics of seizures and of the tumour appears relevant to improve the surgical prognosis. PMID- 10536719 TI - The application of transcranial Doppler sonography in patients with chronic subdural haematoma. AB - The aim of this study was to evaluate the haemodynamic changes of the middle cerebral artery (MCA) and their clinical significance before and after surgical aspiration in patients with chronic subdural haematoma (CSDH). Nineteen patients with CSDH (17 unilateral and 2 bilateral) received transcranial Doppler sonography (TCD) examinations for cerebral blood flow velocity (CBFv) of the MCA prior to and 5 days after neurosurgical treatment. A total of 21 lesion and 10 non-lesion hemispheres were included. Cranial computerized tomography (CT) and clinical assessments were performed before and 3 months following surgery. The preoperative TCD study revealed that the lesion hemisphere had a modest decrease in CBFv in the MCA as compared to the non-lesion hemisphere. Postoperatively, the CBFv significantly improved in the lesion hemisphere, but not in the non-lesion hemisphere, compared to the preoperative data (P < 0.005). The improvement in CBFv showed no significant correlation with brain shift and haematoma volume of the initial cranial CT. Additionally, two patients, who were proved to have a postoperative complication of subdural pneumocephalus, failed to attend follow-up examinations of TCD. Our results support TCD as an alternative follow-up examination for patients with CSDH, although it may not be sensitive enough as a preoperative screening tool. Postoperatively, improvements in the CBFv of the lesion hemisphere are characteristic. An unexplained difficulty of accessing cerebral basal arteries in follow-up TCD examinations should suggest pneumocephalus in the primary differential diagnosis. PMID- 10536718 TI - Correlation between jugular bulb oxygen saturation and partial pressure of brain tissue oxygen during CO2 and O2 reactivity tests in severely head-injured patients. AB - PURPOSE: To correlate the jugular bulb oxygen saturation (SjvO2) and brain tissue oxygen pressure (PbtO2) during carbon dioxide (CO2) and oxygen (O2) reactivity tests in severely head-injured patients. METHODS AND RESULTS: In nine patients (7 men, 2 women, age: 26 +/- 6.5 years, GCS of 6.5 +/- 2.9), a polarographic microcatheter (Clark-type) was inserted into nonlesioned white matter (frontal lobe). PbtO2 and SjvO2 were monitored simultaneously and cerebral vasoreactivity to CO2 and O2 was tested on days three, five and seven after injury. Simultaneous measurements of vasoreactivity by transcranial Doppler (TCD) were undertaken. A total of twenty-one CO2 and O2 reactivity tests were performed. Critical values of PbtO2 (< 15 mm Hg) during induced hyperventilation could be observed four times in two patients. High PbtO2 values up to 80 mm Hg were observed during hyperoxygenation (FiO2 100%). CO2 vasoreactivity by means of PbtO2 was absent in four tests in which measurements by TCD showed intact responses. A stronger correlation between SjvO2 and PbtO2 during the O2 reactivity tests was observed (r = 0.6, p < 0.001), in comparison to values obtained during the CO2 reactivity tests (r = 0.33, p < 0.001). In addition, there was no statistically significant correlation (r = 0.22, p = 0.26) between CO2 reactivity values measured by TCD (4.5 +/- 5.7%) and PbtO2 (3 +/- 2.8%). CONCLUSIONS: Correlation between SjvO2 and PbtO2 during CO2 reactivity test is low, even if significant differences between normo- and hyperventilation values are present. In comparison to SjvO2, monitoring of PbtO2 might more accurately detect possible focal ischaemic events during rapidly induced hyperventilation in severely head-injured patients. The CO2 vasoreactivity by means of changes in Vm MCA seems to be higher in comparison to changes of PbtO2. These observations lead to the hypothesis that vasoreactivity measured by TCD overestimates the cerebrovascular response to CO2. PMID- 10536720 TI - Prediction of the post-comatose motor function by motor evoked potentials obtained in the acute phase of traumatic and non-traumatic coma. AB - OBJECTIVE: To define the value of electrically elicited motor evoked potentials (MEP), obtained during the initial phase of the coma, for correct prediction of the post-coma motor status. METHODS: Fifty-two patients were investigated by MEP within 72 hours after onset of the coma. It was the aim to correlate the MEP findings to the motor function two months after coma onset. RESULTS: Three patients with normal MEP showed no post-coma motor deficit. In 21 patients, a bilateral, symmetric prolongation of the central motor conduction time (CMCT) was registered. Eighteen of these 21 patients (86%) showed a normal post-coma motor status. In 28 patients, unilaterally absent evoked potential, or unilaterally prolonged CMCT, or bilaterally prolonged CMCT with significant difference in each hemisphere were observed. A post-coma contralateral paresis was found in 25 of these 28 patients (89%). That paresis was functionally important in 15 patients (54%) and functionally unimportant in 13 patients (46%). CONCLUSION: We identified certain MEP patterns (unilateral extinction of the evoked potential, unilateral, bilateral prolongation of the CMCT with significant "side" difference), which indicated a pyramidal tract lesion and a post-coma motor deficit with an accuracy of 89%. This refers to the motor results, which may not be the final post-coma motor results which are usually assessed six months after the coma onset. The MEP changes did not allow one to predict the severity of the paresis. The accuracy of prediction of a motor deficit increased from the MEP finding of unilaterally prolonged CMCT to the MEP finding of unilateral extinction of the potential. The most common finding, bilateral central motor slowing without significant "side" difference, did not indicate a post-coma paresis in 86%, leading to the assumption, that bilateral, symmetrical prolongation of the CMCT was not caused by lesions of the descending motor pathways, but by the drugs administered for treating the comatose patient. In conclusion, MEP allows one to predict the presence of a post-coma motor deficit with a high degree of accuracy already in the initial phase of coma, but MEP fails to predict the severity of that deficit. PMID- 10536721 TI - Prevalence of Alzheimer's disease in patients investigated for presumed normal pressure hydrocephalus: a clinical and neuropathological study. AB - During 1991-1995, 223 patients were investigated in the Department of Neurosurgery, Kuopio University Hospital because of a clinical and CT diagnosis of NPH. All patients underwent intracranial pressure measurements and were formed into 3 biopsy groups. Group A included incidentally biopsied patients (104 patients, 34 biopsies) seen during 1991-1992; Group B was a prospective study group from 1993-1995 (all 51 patients biopsied); and Group C patients excluded from Group B (68 patients, 34 biopsies) by age and concomitant diseases. A cortical biopsy was taken before intracranial pressure recording altogether in 118 of the 223 patients. The biopsy revealed normal brain tissue in 66 patients. Prevalence of Alzheimer's disease (AD) in biopsied patients was 42% in Group A, 31.3% in Group B and 50% in Group C. A shunt was placed according to pressure measurement in 110 patients; of these, 8 had both AD and raised ICP. Two patients with both AD and raised ICP improved after shunt placement during the first follow-up year, 4 patients deteriorated and the condition of 2 was similar to that before shunting. The frequency of haematomas after biopsy was 2.9% in groups A and C; in Group B patients had no postoperative haematomas. There was no difference in the incidence of complications in patients who had or did not have a biopsy. The relatively high prevalence of AD in patients with NPH may explain the unsuccessful recovery of many patients after shunt placement. Cortical biopsy is an effective and safe method for finding the co-existence of AD and thus improving the diagnosis of NPH and may prevent unnecessary shunt surgery. PMID- 10536722 TI - Sensitivity, specificity and predictive value of intra-operative elevation of hand temperature to ensure a successful T2-sympathectomy in patients with palmar hyperhidrosis. AB - To appraise the validity, intra-operative elevation of hand temperature ensuring a successful T2-sympathectomy, we conducted a randomized, self-compared, case control study on 40 consecutive patients with palmar hyperhidrosis. All patients had a postoperative follow-up of at least 18 months without recurrence. During operation, dynamic temperature changes on their thenar eminence of both the surgically treated and non-surgically treated hands were simultaneously measured just before (baseline) and after completion of T2-sympathectomy, and again 5 and 10 minutes later. An elevation of the temperature by at least 0.5 degree C from the baseline temperature was recognized as an "elevated" temperature. The relationship between sensitivity and specificity of temperature changes was compared using receiver operator characteristic (ROC) analysis. Sensitivity was defined as the proportion of temperature-elevating procedures in the group of operated hands. As a whole, post-sympathectomy elevation of hand temperature is a useful, but not an ideal, indicator for assuring a successful T2-sympathectomy due to its low sensitivity. At the 5-minute point, if the hand temperature was elevated by 1 degree C, its sensitivity, specificity and positive predictive value were 40%, 80% and 66.7%. In comparison, a 2 degrees C elevation at the 10 minute point had a sensitivity, specificity and positive predictive value of 30%, 90% and 75% (p < 0.05). We suggest that correct localization of the T2 ganglion followed by adequate ablation should be the prerequisite for use of this monitoring system. PMID- 10536723 TI - Platelet-derived growth factor (PDGF-AB) like immune reactivity in serum and in cerebral spinal fluid following experimental subarachnoid haemorrhage in dogs. AB - The aim of this study was to evaluate the following questions: Can the platelet derived growth factor (PDGF-AB) be identified in the serum and cerebro spinal fluid (CSF) of dogs? Is there an increase in the concentration of PDGF-AB following experimental subarachnoid haemorrhage (SAH)? Is the increase in concentration related to the angiographic cerebral vasospasm of the basilar artery. The "double haemorrhage" model was applied in seven dogs to produce experimental SAH with determination of angiographic vasospasm in the basilar artery. Blood and CSF samples were taken on the first, third and eighth days. The analyses were performed with an ELISA human PDGF-AB antibody kit (quantikine human PDGF-AB, R&D Systems, Minneapolis, USA). The average PDGF-AB base value in the serum on the day before the SAH was 410.77 +/- 177.56 pg/ml, in the CSF it was 6.43 +/- 3.19 pg/ml. There was a significant (p = 0.05) increase in the concentration of PDGF-AB (third day 717.35 pg/ml, eighth day 918.07 pg/ml) in the serum of all animals. No significant increase was found in the CSF samples of any animal. In summary, a PDGF-AB like immune reactivity was found in the serum of dogs with the human PDGF-AB ELISA kit and the concentration of PDGF-AB in the serum increased after experimental SAH but not in CSF, but there was no relationship between the increase in PDGF-AB serum concentration and angiographic vasospasm. PMID- 10536724 TI - Low efficacy of gene therapy for rat BT4C malignant glioma using intra-tumoural transduction with thymidine kinase retrovirus packaging cell injections and ganciclovir treatment. AB - BACKGROUND: The purpose of this study was to test the use of Herpes Simplex virus thymidine kinase (HSVtk) retrovirus packaging cell injections in the treatment of malignant brain tumours. METHODS: Therapeutic effect and tissue responses were examined in vivo in a syngeneic BT4C rat glioma model after HSVtk-producing PA317 packaging cell injections and intraperitoneal ganciclovir (GCV) medication. MRI was used to visualise the tumours before and after the treatment. Immunohistochemical stainings were performed to study astroglia and microglia responses and apoptosis-mediated cell death. RESULTS: The results suggest that only a limited treatment effect can be achieved with HSVtk packaging cell injections with no prolonged survival rates. Histological examination showed a strong astroglia response but only a modest microglia response after the treatment. HSVtk and GCV-induced cell death was at least partially mediated by apoptosis. It is concluded that HSVtk packaging cell injections and GCV treatment do not lead to eradication of malignant cells in a syngeneic BT4C rat glioma model. The lack of efficacy is most likely due to low gene transfer efficiency and limited life span of the injected packaging cell inside the tumours. CONCLUSIONS: Improvements in gene transfer efficiency, and stimulation of immunoresponse against tumour cells might lead to a more effective therapeutic response in vivo. PMID- 10536725 TI - Evaluation of functional nerve recovery shows that allogeneic nerve graft treated with ICAM-1 and LFA-1 mAbs can be good alternative to syngeneic graft. AB - The objective of the study is to establish recovery results of tibial nerve defects reconstructed using allogeneic and xenogeneic graft, in host immunosuppressed with Intercellular Adhesion Molecule-1 (ICAM-1) and Lymphocyte Function Antigen-1 (LFA-1) monoclonal antibodies (mAbs). A pilot study was conducted in fifteen Fischer rats by forming a 1 cm right tibial nerve gap, then reconstructing it with 1.2 cm long grafts, namely, Wistar allogeneic, Black mouse xenogeneic, and syngeneic (n = 5/group). The main study included forty-eight rats allocated to the following groups (n = 12/group): 1) Allograft without treatment as control group. 2) Allograft with intraperitoneal ICAM-1 and LFA-1 mAbs treatment. 3) Allograft preserved in Belzers' solution including ICAM-1 mAbs plus standard intraperitoneal treatment. 4) Syngraft as benchmark. At 3, 6 and 9 weeks postengraftment walking track analysis was performed and expressed as Tibial Functional Index (TFI). Motor and compound nerve action potential across the graft conduction velocities were measured at week 10. Xenograft did not show any functional recovery and was therefore excluded from main study. However, pilot and main study results showed recovery results in both treated allogeneic groups and were comparable to benchmark syngraft. Therefore, allogeneic nerve graft could be an alternative in peripheral nerve reconstruction and spinal cord grafting. PMID- 10536726 TI - Punctate midline myelotomy. A new approach in the management of visceral pain. AB - Nauta et al. reported on a successful punctate midline myelotomy (PMM) for the treatment of intractable pelvic pain. The authors describe an other case history of a patient with multiple anaplastic carcinomas of the small intestine, peritoneal carcinosis and retroperitoneal lymphomas, suffering from severe visceral pain in the hypo-, meso-, and epigastrium. Nauta's PMM was successfully performed at the Th4 level. Narcotic medication was tapered from 30 mg i.v. morphine per hour pre-operatively to 5 mg per hour within 5 days postoperatively. Intensity of pain decreased from 10 to 2-3 on the visual analog scale. Only minor transient side effects appeared and the patient was discharged 5 days postoperatively. The pain reduction was maintained until the patient died from the extended disease five weeks later. We conclude that punctate midline myelotomy sufficiently controls not only pelvic visceral pain, but also visceral pain generated in the meso- and epigastrium. The findings support the concept of a midline dorsal column visceral pain pathway. PMID- 10536727 TI - Demonstration of cerebral plasticity by intra-operative neurophysiological monitoring: report of an uncommon case. AB - It has been postulated long ago that "eloquent" areas shift their location in patients with arteriovenous malformations (AVM). Obviously the "motor region" in not located in the precentral gyrus in a patient with an AVM in the "motor region". We report on the case of a 15-year old boy with an AVM in the left sensorimotor cortex, in whom intra-operative mapping showed an inexcitability of the precentral gyrus, while stimulation of the cortex anterior to the primary motor cortex elicited motor responses. This indicates that motor function was translocated from the primary to the supplementary motor cortex. Surgery was performed under general anaesthesia. Neurophysiological monitoring was performed throughout surgery. The central sulcus was identified by phase reversal of the somatosensory evoked potentials. The motor cortex was mapped by direct high frequency (500 Hz) monopolar anodal stimulation. In the patient herein reported, stimulation of the "anatomically" defined primary motor cortex induced no motor response, as expected. Motor response was elicited only by stimulation of the cortex anterior to the precentral gyrus. There was no postoperative deterioration of motor function. These observations indicate that the precentral gyrus was functionally "useless". The motor region was relocated into more rostral areas in the supplementary motor cortex. This translocation of function in the presence of an AVM indicates cerebral plasticity. PMID- 10536728 TI - Ophthalmic artery aneurysm associated with Horner's syndrome. PMID- 10536729 TI - Trilateral retinoblastoma. PMID- 10536730 TI - Bow Hunter's stroke caused by simultaneous occlusion of both vertebral arteries. PMID- 10536731 TI - Vagoglossopharyngeal neuralgia caused by a neuroma of vagal rootlets. PMID- 10536732 TI - Transient mutism after brain stem infarction (ref. Acta Neurochirurgia 1999, 141: 209-213) PMID- 10536733 TI - Referring to the posterior fossa cranioectomy and tonsillar resection in order to treat Chiari I malformation with syringomyelia. PMID- 10536734 TI - NIMH symptom challenge and medication discontinuation workshop. PMID- 10536735 TI - The National Bioethics Advisory Commission (NBAC) report. PMID- 10536736 TI - An NIMH commentary on the NBAC report. PMID- 10536737 TI - Medication discontinuation and symptom provocation in research: a consumer and family perspective. AB - Research involving symptom provocation and medication discontinuation, although common throughout medicine, raises some significant issues for consumers and family members facing severe mental illnesses. Reasons for our attention to these research approaches include the apparent prevalence of this research; the prospect of significant distress, illness, or disability, especially in the event of medication discontinuation; questions about the scientific strength of at least some challenge studies; the changing nature and pressures on research, including its "deinstitutionalization," and the increasing role of private funders; early evidence of consumer concerns about medication discontinuation; the possibility of long-term damage as a result of delayed treatment; concerns about subject recruitment methods and the adequacy of informed consent for these studies; and the lack of data concerning consumer experience and outcome from this research. Because research involving symptom provocation and medication discontinuation can present significant risks to the consumer participating in the study, we must take steps to make sure of the following: that the use of this research methodology be minimized as much as possible; that it meets the very highest standards of scientific merit and necessity; that it includes the strongest possible protections for human subjects, including optimal informed consent about the nature and risks of the study and communication with care giving family members and others; that in discontinuation studies research protocols precisely spell out in advance the clinical situations that will trigger intervention and what the intervention will be, and furthermore, that discontinuation studies should assure adequate clinical monitoring for early identification and treatment of signs of clinical distress or deterioration; and that researchers should begin systematically collecting and publishing information on the outcomes for and experiences of consumers in these types of studies. Enhanced oversight of these research protocols, by funders and IRBs is essential. Finally, our consideration of these issues has raised concerns for us about recruitment methods, which warrant examination; and we would like to see attention to placebo control use, insofar as it is necessary to show new medications are effective. PMID- 10536738 TI - The controversy over challenge and discontinuation studies: perspective from a consumer-psychiatrist. PMID- 10536739 TI - Psychiatric research ethics: an overview of evolving guidelines and current ethical dilemmas in the study of mental illness. AB - The field of psychiatry has an opportunity to construct a more refined, perhaps more enduring understanding of the ethical basis of mental illness research. The aim of this paper is to help advance this understanding by 1) tracing the evolution of the emerging ethic for biomedical experimentation, including recent recommendations of the President's National Bioethics Advisory Commission, and 2) reviewing data and concepts related to compelling ethical questions now faced in the study of mental disorders. Empirical findings on informed consent, the ethical safeguards of institutional review and surrogate decision making, and the relationship between scientific and ethical imperatives are outlined. Psychiatric researchers will increasingly be called upon to justify their scientific approaches and to seek ways of safeguarding the well-being of people with mental illness who participate in experiments. Most importantly, psychiatric investigators will need to demonstrate their appreciation and respect for ethical dimensions of investigation with special populations. Further empirical study and greater sophistication with respect to the distinct ethical issues in psychiatric research are needed. Although such measures present many challenges, they should not interfere with progress in neuropsychiatric science so long as researchers in our field seek to guide the process of reflection and implementation. PMID- 10536740 TI - IRB review of psychiatric medication discontinuation and symptom-provoking studies. AB - Federal regulations governing human subjects research call for additional protections for the "mentally disabled." However, there is currently no consensus definition of mental disability or guidelines for how these research subjects should be protected. This ambiguity complicates the work of institutional review boards (IRBs) charged with the review and approval of protocols involving psychiatric medication discontinuation and symptom provocation. It is particularly important for these studies to be reviewed within the larger context of the research program in which they are conducted. The author proposes a process for IRB review of these studies, which includes the implementation of additional safeguards for subjects determined by the IRB to be vulnerable. Recommendations also are made for training psychiatric clinical investigators in issues related to research bioethics. PMID- 10536741 TI - Integrating science and ethics in child and adolescent psychiatry research. AB - Research to elucidate the biological bases of psychopathology in children and adolescents is needed to understand pathogenesis and to develop effective and safe treatment and preventive interventions. Because of the effect of development, data collected in adults are not always applicable to youth, and direct participation of children in research is necessary. Many medications are currently used in the community to treat children and adolescents with neuropsychiatric disorders without adequate data about their safety and efficacy. Conducting research in children requires attention to specific ethical and regulatory factors. In deciding whether minors can participate in a study with potential direct benefit to the research subjects, the most important variable to consider is the balance between risks and potential benefit, in the context of the severity of the child's condition and the available alternatives. Research with no potential benefit to the participants is guided by the concepts of minimal risk (which may apply more to normal children) and minor increase over minimal risk (perhaps more relevant to children affected by psychopathology). Recently conducted studies relevant to this issue are reviewed. Of paramount importance is the ratio of risk/scientific value of the proposed experiment. In fact, no research is justifiable, no matter how low the risk may be, unless the potential yield of the study is important and may help advance our understanding of normal functioning and mental illness. PMID- 10536743 TI - Symptom provocation studies in psychiatric disorders: scientific value, risks, and future. AB - Several lines of investigation have contributed to the increasing recognition of the biological basis of psychiatric disorders. Symptom provocation studies have made important contributions toward this. With the emergence of novel methodologies, the role of symptom provocation studies has come under increasing scrutiny and debate. The scientific contributions and risks of symptom provocation studies are discussed using the psychostimulant paradigm in schizophrenia research as the prototypical study. The application of studies in other areas of medicine that carry risks similar to those associated with symptom provocation studies, are also reviewed. The authors draw on the parallel of cardiac stress testing to highlight risks: benefits issues. Finally, the authors discuss the future of symptom provocation studies and emphasize that these studies will need to meet the highest scientific standards, ethical standards and safeguards. PMID- 10536742 TI - Neuroleptic discontinuation in clinical and research settings: scientific issues and ethical dilemmas. AB - The ethics of neuroleptic discontinuation in clinical and research settings are currently a topic of much discussion. The issues underlying this debate are complicated by the fact that these medications can be fairly effective in managing the symptoms and preventing relapse in schizophrenia and other psychotic disorders, yet these drugs have therapeutic limitations and their prolonged use is associated with a risk of serious, potentially persistent side-effects such as tardive dyskinesia. Over the past 47 years, the public perception about the value of neuroleptics has undergone dramatic shifts, based partly on the data available at different time periods. The risk-benefit ratio is better for the atypical antipsychotics compared to the conventional ones, but long-term experience with the newer agents has been limited. At present, a prudent strategy for most clinical and research purposes is to gradually taper the medications in clinically stable, carefully selected, consenting subjects to the lowest doses on which individual patients can be effectively maintained. In this article we discuss clinical, research, and ethical aspects of neuroleptic discontinuation. It is critical to protect potentially vulnerable patients with serious mental illnesses, while allowing them to benefit from appropriate investigations. PMID- 10536744 TI - The schizophrenia ketamine challenge study debate. AB - Protection of subjects in psychiatric research is an issue of considerable public and professional interest. Perhaps the most hotly debated issue concerns challenge study protocols where symptoms of illness are increased in a bioassay designed to gain knowledge of pathophysiology. Although widely used in biomedical research, the ethics of this application in mental illness research are contested. At issue is whether acute distress and lasting harm are caused without direct benefit in vulnerable subjects without valid informed consent. The ketamine challenge study in schizophrenia subjects is at the vortex of the current debate. This report presents background data on ketamine safety and a qualitative and quantitative analysis of data from all schizophrenia subjects in North American ketamine studies. Duration and severity of change in psychosis and anxiety, "worst case" experiences, and information on prolonged adverse effects are detailed. The vulnerable population and informed consent issue is discussed. Group results show that psychosis increase is mild to moderate and brief, anxiety is mild and brief, and no evidence of prolonged adverse effects is found. Few "worst case" incidents were identified, and these were clinically managed successfully in a short time period. Although more difficult to evaluate, informed consent procedures seem adequate, and consent was voluntary in subjects judged to have decisional capacity for this purpose and in circumstances where alternative clinical care could be freely chosen. The author concludes that ketamine challenge studies meet ethical standards, have been conducted without lasting adverse effects, that discomfort is modest and brief, and important new knowledge has been gained of potential benefit to the class from which subjects were drawn. PMID- 10536746 TI - Ethical dimensions of psychiatric research: a constructive, criterion-based approach to protocol preparation. The Research Protocol Ethics Assessment Tool (RePEAT). AB - Preparing experimental protocols that are ethically sound, possess scientific merit, and meet institutional and national standards for human subject protections is a key responsibility of psychiatric investigators. This task has become increasingly complex due to developments in biomedical science, bioethics, and society at large. Practical and constructive approaches to help investigators in their efforts to create protocols that are ethically acceptable have nevertheless received little attention. To better address this gap, the Research Protocol Ethics Assessment Tool (RePEAT) was developed as an educational instrument to help assure that ethically important elements, including scientific design features, are explicitly addressed by investigators in their work with protocols involving human participants. The RePEAT is a brief evaluative checklist that reflects rigorous ethical standards, particularly with respect to criteria for studies that may involve individuals with compromised decisional abilities. For this reason, it may be especially beneficial as a self-assessment tool for investigators and protocol reviewers in psychiatry. To stimulate education and dialogue, this report presents the RePEAT and outlines its content, format, use, and limitations. PMID- 10536745 TI - The long-term effects of placebo in patients with chronic schizophrenia. AB - BACKGROUND: It has been hypothesized that placebo periods may increase long-term morbidity for patients with schizophrenia. In this study, the long-term effect of a placebo period was evaluated in a group of relatively treatment-refractory patients with chronic schizophrenia. METHODS: This retrospective study examined behavioral rating scores for 127 patients with chronic schizophrenia who were placed in a double-blind placebo study on the inpatient units of the National Institute of Mental Health Neuropsychiatric Research Hospital. Patients were rated daily with the Psychiatric Symptom Assessment Scale (PSAS), an extended and anchored version of the Brief Psychiatric Rating Scale (BPRS). At the end of the placebo phase, most patients were placed on haloperidol. Pre-placebo baseline PSAS ratings were compared with, first, discharge ratings and second, post placebo ratings. To determine expected variability in the course of illness, patients in the placebo group were compared with patients hospitalized during the same time period, but who did not enter the placebo study. RESULTS: By discharge, ratings for placebo patients had returned to baseline. Post-placebo ratings were quite variable. Although many of the placebo patients had returned to baseline by day 3 of the post-placebo phase, others had not returned to baseline by post placebo day 42. PSAS Total Scores for patients who left the study early were no different at baseline, placebo, or through post-placebo day 35 compared with patients who completed the study. CONCLUSIONS: The results indicate that given a sufficiently lengthy recovery period, patients with chronic schizophrenia who go through a placebo phase return to baseline, but that the speed with which they attain that recovery is highly variable. PMID- 10536747 TI - The role of surgery and chemoradiation therapy for cancer of the rectum. PMID- 10536748 TI - Child abuse--controversies and imposters. PMID- 10536749 TI - Developmental toxicity study of glycolic acid in rats. AB - The developmental toxicity of glycolic acid was assessed in rats by orally administering solutions of the test material in water over days 7-21 of gestation (the day of copulation plug detection was defined as day 1 of gestation). Groups of 25 mated female Crl: CD BR rats were gavaged at daily dose levels of 0, 75, 150, 300 or 600 mg/kg. The dams were euthanized on day 22 and the offspring were weighed, sexed, and examined for external, visceral, and skeletal alterations. Clear evidence of maternal toxicity was demonstrated at 600 mg/kg; adverse clinical observations were statistically significantly increased (wheezing/lung noise, abnormal gait/staggering, lethargy). In addition, maternal body weights, weight changes, and food consumption were statistically significantly reduced at this dose level. Marginal evidence of maternal toxicity was demonstrated at 300 mg/kg; wheezing/lung noise similar to that seen at 600 mg/kg was observed in 2 of 25 dams. This increase approached statistical significance (p = 0.0553). There was marked evidence of developmental toxicity at 600 mg/kg. Mean fetal weight was statistically significantly reduced while the incidences of skeletal (ribs, vertebra, and sternebra) malformations and variations were statistically significantly increased. At 300 mg/kg/day, there was a slight (2 affected fetuses from 2 litters) increase in the incidence of two skeletal malformations: fused ribs and fused vertebra. Although these increases were not statistically significant (p = 0.0555), they were consistent with findings seen at 600 mg/kg/day and thus were considered relevant. There was no other evidence of developmental toxicity at 300 mg/kg/day nor was any developmental toxicity seen at 150 or 75 mg/kg/day. Thus, the maternal and developmental no-observed-effect level (NOEL) was considered 150 mg/kg. PMID- 10536750 TI - Occupational hazard evaluation of p-bromobenzyl bromide from tests for genotoxicity. AB - As part of an occupational hazard evaluation, p-bromobenzyl bromide (p-BBB) was evaluated for genotoxic activity in the Ames microbial mutagenicity assay, the alkaline elution assay for DNA strand breaks in rat hepatocytes and the in vitro chromosome aberration assay in Chinese hamster ovary cells. The compound produced equivocal results in the microbial mutagenicity assay but was negative in the alkaline elution assay for DNA strand breaks in rat hepatocytes. The compound produced weakly positive results in the in vitro chromosome aberration assay. There was substantial cytotoxicity in all three assays. It is concluded that p BBB is weakly genotoxic. PMID- 10536751 TI - Effects of inhalation exposures to an M1-receptor agonist on ventilation in rhesus monkeys. AB - Information was needed on effects of possible occupational inhalation exposure to an M1-receptor agonist (xanomeline) such as might occur during the manufacturing process. Both acute and repeated inhalation exposures to xanomeline were carried out in six male rhesus monkeys using a head-dome exposure system. Exposure concentrations ranged from 0.3 to 10 mg/m3. The exposure durations were up to 2 weeks. Decreases in tidal volume and increases in respiratory frequency were both time and concentration related during acute exposures. These effects were blocked with atropine pre-treatment. Correlation with pulmonary resistance measurements in two monkeys suggested that these were bronchoconstrictive changes that increased with severity with time at a given concentration and with concentration when measured after a constant exposure time. The dose-response was relatively steep with 10 mg/m3 becoming intolerable to the monkeys after approximately 15 minutes, but no measurable effects were observed at 0.3 mg/m3 after up to 4 hours of exposure. To investigate the effects of repeated exposures, monkeys were exposed for 4 hr/day, 5 days/wk for 2 weeks to 0.0 (air only), 0.3, and 1.2 mg xanomeline/m3 of air. When compared to the air-only exposure, 0.3 mg/m3 caused no significant changes in tidal volume. In contrast, 1.2 mg/m3 caused a rapid and significant decrease in tidal volume that was sustained throughout the 4-hr exposure. A slower rise in breathing frequency also occurred. Repeated exposures did not alter the effects seen after a single exposure. It is concluded that xanomeline, a M1-receptor agonist, can acutely alter normal ventilation in non human primates at airborne concentrations > or = 0.6 mg/m3 and should be carefully controlled in a manufacturing environment. The no-observed-effect concentration was 0.3 mg/m3. PMID- 10536752 TI - In vitro methylation of arsenite by rabbit liver cytosol: effect of metal ions, metal chelating agents, methyltransferase inhibitors and uremic toxins. AB - The methylation of carrier-free 74As-arsenite by liver cytosol of Flemish Giant rabbits is highly susceptible to additions of trace elements. In vitro supplementation of essential trace elements like zinc (Zn2+), vanadium (V5+), iron (Fe2+), copper (Cu2+) and selenate was shown to increase the methylation efficiency. Trivalent metal ions (e.g. Al3+, Cr3+ and Fe3+), Hg2+, Tl+ and SeO3(2 ) had a deleterious effect. The inhibitory effect of EDTA, oxime and many divalent cations (Ca2+, Mg2+, Sr2+, ...) suggest a co-factor role for a specific divalent metal ion, possibly Zn2+. Chelating agents used in clinical treatment of acute and chronic inorganic arsenic poisoning lower the methylation capacity of cytosol by rendering the trivalent arsenic unavailable for the methyltransferase enzymes. S-adenosylhomocysteine and periodate-oxidized adenosine, inhibitors of s adenosylmethionine dependent methylation pathways, inhibit the methylation of arsenite. Pyrogallol, a catechol-O-methyltransferase inhibitor, blocks the action of arsenite- and monomethylarsonic methyltransferase enzymes, suggesting a close structural relationship between the active sites of the different enzymes. Some uraemic toxins, namely oxalate, p-cresol, hypoxanthine, homocysteine and myo inositol, inhibit arsenic methylation. PMID- 10536753 TI - Some studies on the rodenticidal action of indomethacin. AB - The oral LD50 of indomethacin for a seven-day observation was found to be 12.58 +/- 1.15 mg/kg. At LD10 of 6.61 mg/kg, a dose to weight ratio of 28 was obtained for a 240 g rat, while at a maximum single dose of 3 mg/kg in man it is only 0.04. Neither diazepam nor phenobarbital influenced death at the doses of both drugs used. However, cholestyramine 2 g/kg/day was found to protect by 50% from the LD100 of indomethacin. Gross pathological studies showed dose-dependent ulceration and perforation (P < 0.001, 12 vs 24 mg/kg) and such lesions occurred in starved rats, were low in bile duct-ligated compared to sham-operated rats (P < 0.001) and were also low in cholestyramine-treated rats. Indomethacin-induced lethality in rats was found to be dose-dependent. PMID- 10536754 TI - Protective effect of Cassia occidentalis extract on chemical-induced chromosomal aberrations in mice. AB - This study was conducted to determine the antimutagenic potential of aqueous extract of Cassia occidentalis against the chromosomal aberrations (CA) produced in vivo by benzo[a]pyrene (B[a]P) and cyclophosphamide (CP) in mice. Animals (male mice) were treated with three doses of plant extract (50 mg/kg, 250 mg/kg and 500 mg/kg) for 7 days prior to the administration of single dose of mutagens (B[a]P 125 mg/kg oral; CP 40 mg/kg i.p.). The results indicated that C. occidentalis was not genotoxic per se and exerted no other toxic signs and symptoms in treated animals. The chromosomal aberrations produced by B[a]P and CP were significantly reduced (p < 0.001) by C. occidentalis pre-treatment. Furthermore, animals treated with plant extract showed a reduced level of cytochrome P 450 (Cyt P 450) and elevated levels of glutathione S-transferase (GST) activity and glutathione content in the liver. It seems that C. occidentalis exerts its antimutagenic activity by modulating the xenobiotic activation and detoxification mechanisms. PMID- 10536755 TI - Toxicology and humoral immunity assessment of octamethylcyclotetrasiloxane (D4) following a 28-day whole body vapor inhalation exposure in Fischer 344 rats. AB - Octamethylcyclotetrasiloxane, D4, is a low viscosity, silicone fluid consisting of four dimethyl-siloxy units ((CH3)2SiO)4 in a cyclic structure. It is primarily used as a building block in the industrial synthesis of long chain silicone polymers. The combination of D4 with decamethylcyclopentasiloxane (D5) is commonly referred to as cyclomethicone which has a wide range of applications as a formulation aid in personal care products. To extend the existing database regarding the biological activities of D4, a 28 day whole body vapor inhalation study was conducted using Fischer 344 rats at 0 (room air), 7, 20, 60, 180 and 540 ppm for 6 hours/day, 5 days/week. Parameters measured included body weights, organ weights, gross pathology, histopathology, serum chemistries, and urinalysis. In addition to these standard toxicological endpoints, the ability of D4 exposed animals to mount an IgM antibody response was evaluated by a splenic antibody forming cell (AFC) assay and a serum enzyme-linked immunosorbant assay (ELISA). The results of this 28-day inhalation study indicate that D4 exposure caused no adverse effects on body weight, food consumption, or urinalysis parameters. In addition, there were no exposure related histopathological alterations at any site for any exposure group. A statistically significant increase in liver weight and the liver to body weight ratio was observed in both male (180-540 ppm) and female (20-540 ppm) rats, which was not observed in the 14 day recovery group animals. There were no other significant organ weight changes. Although statistically significant changes were observed in several hematological and serum chemistry parameters in both the terminal and 14-day recovery animals, the changes were marginal and within the normal range of values for the rat. Under these experimental conditions, there were no alterations noted in immune system function at any of the D4 exposure levels. PMID- 10536756 TI - Time course of chronic oral cadmium nephrotoxicity in Wistar rats: excretion of urinary enzymes. AB - Twelve male and female Wistar rats each received cadmium (as CdCl2) in their diet at concentrations of 0, 10, 50, and 250 ppm for 72 weeks. After 1, 4, 8, 13, 18, 26, 32, 45, 57, and 68 weeks a total of 8 enzymes from different cellular compartments of the nephron were measured. At the end of the study period, the kidneys were examined histopathologically. Concentrations up to and including 50 ppm did not induce any adverse effect. At 250 ppm, growth of male and female animals was markedly retarded. Significantly increased activities of the cytosolic phosphohexose isomerase were excreted by males and females receiving 250 ppm at all timepoints from week 13. The values of the mitochondrial glutamate dehydrogenase were mostly elevated from week 1 to 57, however, due to a wide scatter range, were only occasionally significantly different from control values. The brush border enzymes (gamma-glutamyl transferase, alkaline phosphatase and leucine arylamidase) were not changed in a relevant manner in female rats, while in 250 ppm males the excreted activity of ALP and LAP from week 1 to week 18, and that of GGT during the entire study period were significantly lower than the control values. Excretion of the lysosomal enzymes aryl sulfatase A, beta-galactosidase, and beta-N-acetyl-D-glucosaminidase was at no time influenced in a noteworthy manner. Histopathology after 72 weeks revealed chronic but also acute degenerative changes in the kidneys of 250 ppm males and females. A comparison of published data on persons having undergone high cadmium exposure with the results presented here shows remarkable differences. PMID- 10536757 TI - Reforming the 1983 Mental Health Act. PMID- 10536758 TI - Presumed consent or contracting out. PMID- 10536759 TI - Evaluating ethics competence in medical education. AB - We critically evaluate the ways in which competence in medical ethics has been evaluated. We report the initial stage in the development of a relevant, reliable and valid instrument to evaluate core critical thinking skills in medical ethics. This instrument can be used to evaluate the impact of medical ethics education programmes and to assess whether medical students have achieved a satisfactory level of performance of core skills and knowledge in medical ethics, within and across institutions. PMID- 10536760 TI - Resurrecting autonomy during resuscitation--the concept of professional substituted judgment. AB - The urgency of the resuscitation and the impaired ability of the patient to make a reasonable autonomous decision both conspire against adequate consideration of the principles of medical ethics. Informed consent is usually not possible for these reasons and this leads many to consider that consent is not required for resuscitation, because resuscitation brings benefit and prevents harm and because the patient is not in a position to give or withhold consent. However, consent for resuscitation is required and the common models employed for this purpose are presumed consent or consent from a patient proxy. However, if we are to honour the principles of respect for patient autonomy, as well as beneficence and non maleficence, when starting and continuing resuscitation we must try and achieve the best balance between benefit and harm from the patient's perspective. The concept of professional substituted judgment involves the resuscitators gathering as much information about the patient as they possibly can, including any previously expressed attitudes towards such a situation, and combining this with their acquired professional knowledge of the likely benefits and harms of the resuscitation endeavour and then exercising their moral imagination, imagining themselves as the patient, and asking "would I want this treatment?" By employing professional substituted judgment resuscitators should recognise when the balance of benefit and harm becomes unfavourable from the patient's perspective and at this point they have a moral obligation to withdraw resuscitation as they can no longer presume the patient's consent. In this way the principles of beneficence, non-maleficence and respect for patient autonomy are more favourably balanced than under other resuscitation decision making processes. PMID- 10536761 TI - Parental consent to publicity. AB - The problems presented by the use of named child patients and their medical histories in television, radio and newspapers is discussed. It is suggested that it is not acceptable to regard this as comparable to their participation in non therapeutic research, and that no one, not even the parent has the authority to give consent to such use. PMID- 10536762 TI - Managed care and ethical conflicts: anything new? AB - Does managed care represent the death knell for the ethical provision of medical care? Much of the current literature suggests as much. In this essay I argue that the types of ethical conflicts brought on by managed care are, in fact, similar to those long faced by physicians and by other professionals. Managed care presents new, but not fundamentally different, factors to be considered in medical decision making. I also suggest ways of better understanding and resolving these conflicts, in part by distinguishing among conflicts of interest, of bias and of obligation. PMID- 10536763 TI - Beneficence in general practice: an empirical investigation. AB - OBJECTIVES: To study and report the attitudes of patients and general practitioners (GPs) concerning the obligation of doctors to act for the good of their patients, and to provide a practical account of beneficence in general practice. DESIGN: Semi-structured interviews administered to GPs and patients. SETTING AND SAMPLE: Participants randomly recruited from an age and gender stratified list of GPs in a geographically defined region of South Australia. The sample comprised twenty-one general practitioners and seventeen patients recruited by participating GPs. RESULTS: In practice, acting for the good of the patient not only accommodates the views of patients and GPs on expertise and knowing best, but also responds to the particular details of the clinical situation. Patients had a complex understanding of the expertise necessary for medical practice, describing a contextual domain in which they were expert, and which complemented the scientific expertise of their GPs. General practitioners identified multiple sources for their expertise, of which experience was the most significant. The role of the GP included responding to individual patients and particular clinical problems and ranged from the assumption of responsibility through to the proffering of medical advice. CONCLUSION: This study found that GPs acting for the good of their patients covered a variety of GP actions and patient preferences. Beneficence was not justified by presumed patient vulnerability or the inability of patients to understand medical problems, but furthered through a recognition of the different areas of expertise contributed by both parties to the consultation. PMID- 10536764 TI - The role of ethical principles in health care and the implications for ethical codes. AB - A common ethical code for everybody involved in health care is desirable, but there are important limitations to the role such a code could play. In order to understand these limitations the approach to ethics using principles and their application to medicine is discussed, and in particular the implications of their being prima facie. The expectation of what an ethical code can do changes depending on how ethical properties in general are understood. The difficulties encountered when ethical values are applied reactively to an objective world can be avoided by seeing them as a more integral part of our understanding of the world. It is concluded that an ethical code can establish important values and describe a common ethical context for health care but is of limited use in solving new and complex ethical problems. PMID- 10536765 TI - Attitudes of the Lebanese public regarding disclosure of serious illness. AB - OBJECTIVES: To measure the preference regarding disclosure of a serious diagnosis, and its determinants, of the Lebanese public. DESIGN AND SETTING: Non random sample survey of 400 persons interviewed in health care facilities in Beirut in 1995. RESULTS: Forty-two per cent of respondents generally preferred truth not to be disclosed directly to patients. Preference for disclosure was associated with younger age, better education and tendency to rapport-building with physicians. There were no meaningful associations between place of residence (urban/rural), level of religious practice, or religious affiliation, and preference for disclosure. CONCLUSIONS: Under one plausible interpretation, this survey suggests that the expectation for concealment will decrease as the advantage of knowledge in better coping with disease is understood by an increasingly better educated public, and that the Lebanese public will increasingly come to expect direct and full disclosure of serious diagnoses. PMID- 10536766 TI - Bioethics regulations in Turkey. AB - Although modern technical and scientific developments in medicine are followed closely in Turkey, it cannot be claimed that the same is true in the field of bioethics. Yet, more and more attention is now being paid to bioethics and ethics training in health sciences. In addition, there are also legal regulations in bioethics, some of which are not so new. The objective of these regulations is to provide technical and administrative control. Ethical concerns are rather few. What attracts our attention most in these regulations is the presence of the idea of "consent". PMID- 10536767 TI - Hospice and euthanasia in The Netherlands: an ethical point of view. AB - This contribution is a report of a two months' participant observation in a Dutch hospice. The goal of the observation was to gain an overview of moral decisions in a hospice in which euthanasia, a tolerated practice in the Netherlands, is not accepted as an option. In an introduction, the development of palliative care in the Netherlands will be briefly presented. Subsequently, various moral decisions that were taken during the participant observation are presented and analysed by means of case reports. Attention is especially drawn to decisions that directly or indirectly relate to euthanasia. These moral decisions will be clarified in the light of the philosophy behind the concept of palliative care as it has evolved since the foundation of St Christopher's Hospice, London in 1967. PMID- 10536768 TI - League tables, institutional success and professional ethics. AB - League tables are just one example of the growing importance of "institutional success" in the health service. What are the implications of attaching importance to institutional success, and what impact might this have on professional ethics? This paper considers these issues and argues that public policy processes which centre on institutional performance, and which co-opt professional loyalties to this end, shift the balance between person-centred and impersonal standpoints in health care (from the former and towards the latter). There is no attempt to make a global ethical appraisal of this putative shift but rather to raise a matter of concern for those committed to a person-centred conception of professional ethics. PMID- 10536769 TI - 'He is too young to die ... and you too, doctor'. AB - Sometimes caregiving can be a matter of fear rather than of love. PMID- 10536770 TI - Smokers and taxes. PMID- 10536771 TI - A clinical ethics committee in a small health service trust. PMID- 10536772 TI - The recruitment of non-English speaking subjects into human research. PMID- 10536773 TI - Medicine and literature: imagine a third way. PMID- 10536774 TI - Changing concepts in the management of pediatric urinary tract infections. PMID- 10536775 TI - Urinary tract infections: from pathogenesis to outcome. PMID- 10536776 TI - The pathogenesis of urinary tract infections. PMID- 10536777 TI - The epidemiology and clinical presentation of urinary tract infections in children younger than 2 years of age. AB - UTI in young infants generally presents with fever. Among the youngest infants, boys and girls are equally affected. The incidence of UTI in uncircumcised boys is comparable with that in girls, whereas the rate in circumcised boys is much lower. Based on gender and race, white girls have the highest incidence of UTI. A full understanding of the epidemiology of UTI is complicated by the presence of asymptomatic bacteriuria and by incomplete evidence regarding the significance of scarring and the risk of sequelae. PMID- 10536778 TI - The epidemiology and clinical presentation of urinary tract infections in children 2 years of age through adolescence. PMID- 10536779 TI - The effects of naked neck genotypes, ambient temperature, and feeding status and their interactions on body temperature and performance of broilers. AB - The effect of ambient temperature (AT) and feeding status on body temperature (BT) were investigated in broilers of the three naked neck genotypes (Na/Na, Na, na, and na/na). From 29 to 49 d of age, chicks were reared in a temperature controlled chamber, where AT alternated daily between 24 and 32 C. At Day 47, all birds were deprived of feed for 12 h at 32 C, followed by 12 h of ad libitum intake at 24 C, then 12 h of ad libitum intake at 32 C, and finally feed deprivation for 12 h at 24 C. Body temperature was measured at the end of each of these 12-h periods. Body weight, feed consumption, feather coverage, and breast yield were determined. The Na/na and Na/Na birds had 20 and 40% less feather mass than the na/na birds. The three genotypes had similar BW at Day 49, but the naked neck birds had a higher breast yield. At high AT, BT was positively associated with feather mass of the three naked neck genotypes. The highest BT was exhibited by the fully feathered birds, and the lowest by the homozygous naked neck birds. The feeding status also affected BT of all birds, but to a larger extent in the normally feathered than in the naked neck birds. It appears that the lower negative effects of high AT on growth rate and meat yield in naked neck broilers can be attributed to their lower BT. Thus, it is suggested that measuring BT of broilers can be used as an indicator of the level of stress imposed on them by high AT. PMID- 10536780 TI - The effects of the naked neck (Na) and frizzle (F) genes on growth and meat yield of broilers and their interactions with ambient temperatures and potential growth rate. AB - High ambient temperatures (AT) decrease the growth of broilers because of difficulty in dissipating heat through the feather coverage. Broilers selected for higher growth rate eat more and generate more heat per unit of time; hence, they may become more sensitive to high AT. Reduced feather coverage, either by decreased number or by modified shape, may help birds to dissipate internal heat more efficiently. Two broiler stocks were studied; each was segregated for four genotypes with regard to the genes for naked neck (Na) and frizzled feathers (F): heterozygous naked neck (Na/na f/f), heterozygous frizzle (na/na F/f), double heterozygous (Na/na F/f), and normally feathered (na/na f/f). One stock had a high growth rate (GR) similar to current commercial broilers, whereas the second stock had a lower GR. Birds of each stock, genotype, and sex were reared under constant standard AT (24 C) or high AT (32 C). Body weight at 4 and 7 wk, weight gain (WG) from 4 to 7 wk, breast meat yield, and feather weight were recorded. Reduction in WG from 4 to 7 wk because of high AT was greater in high-GR birds than in low-GR birds, but, in both stocks, the high AT effect was greater on normally feathered birds than on the other three genotypes. AT 32 C, in low- and high-GR stocks, the F allele increased WG from 4 to 7 wk and increased the BW at 7 wk of fully feathered (na/na) broilers but had no effect on meat yield. The effects of the Na allele were similar to or greater than those of the F allele. The Na allele did not affect breast meat yield of low-GR broilers but increased it significantly in high-GR broilers. Combining the two allles resulted in an additive effect, which was more pronounced in the high-GR stock. PMID- 10536781 TI - Season by genotype interaction related to broiler growth rate and heat tolerance. AB - A study was conducted to evaluate the effect of genotype by environment (G x E) interaction on the performance of commercial broilers. Temperate and hot environments were established by making use of the natural climatic differences between spring and summer in western Turkey. The experimental population was produced by a full-pedigree, randomly assigned mating scheme consisting of 29 sires and five dams per sire. The sires were considered genotypes, and the G x E interaction was evaluated by regressing sire breeding values in summer on those estimated from their spring offspring. The correlation between the two seasons for weight gain from 0 to 4 wk of age was r = 0.26, significantly lower than rho = 1 (the expectation when there is no G x E interaction). This correlation was even negative (although not significantly lower than rho = 0) for weight gain (WG) from 4 to 7 wk of age and BW at 7 wk of age. Genotype by season ANOVA also revealed highly significant G x E interaction effects on all traits. These interactions suggest the presence of substantial genetic variation in the magnitude of heat tolerance. It appeared that this variation was not random, but rather related to growth potential, where genotypes that gain more weight in the spring tended to gain less weight under the hot conditions of summer. PMID- 10536782 TI - The natural history of the obese strain of chickens--an animal model for spontaneous autoimmune thyroiditis. AB - Chickens of the Obese strain (OS) are hereditarily affected with spontaneous autoimmune thyroiditis that resembles Hashimoto's thyroiditis of humans in clinical, histopathological, serological, and endocrinological aspects. In this review, the natural history of the OS, reflecting the development and maintenance of the stock and its improvement of productivity over many years at Cornell University, is summarized. To underline the value and usefulness of this animal model, the concept of the multifactorial pathogenesis of autoimmune diseases, which was mainly established at the University of Innsbruck, Austria, is briefly outlined. This detailed analysis on the natural history of the OS was only made feasible by the availability of four decades of records on this chicken line at Cornell University. The report starts with the initial occurrence of a few pullets within the Cornell C-strain (CS) flock that showed obesity, long and silky feathers, and small body size, which, therefore, caused these birds to look notably different than the other CS birds. Experimental findings indicated an autoimmune basis for these characteristics, and the objectives of matings were initially focused on increasing the number of obese individuals for studies on the etiopathology and the mode of inheritance. In subsequent years, matings were directed toward increasing the penetrance and severity of the obese trait in the population. In recent years, the reproductive capability of the obese stock was improved by mating only the best breeders of the population in terms of body weight, egg weight, egg production, fertility, hatchability, and the expression of the obese phenotype. Housing conditions for OS chickens at Cornell and the qualification standards and selection procedures for breeders are described in detail, and results of blood typings are shown. A specific recent finding in OS hens is the high incidence of residues of the right Mullerian duct with cyst formation. PMID- 10536783 TI - Genetic relationship among lines and smooth muscle and ovarian follicular development within lines of Japanese quail in two long-term selection studies. AB - Smooth muscle tumor and ovarian follicular development were studied in lines of Japanese quail selected for increased 4-wk BW (HW, P, and T) and their randombred controls (C and R1). The lines studied were from long-term selection studies at The Ohio State University (HW and R1) and The University of Georgia (P, T, and C). To study the genetic relationship among the lines in the two selection studies, the C, P, HW, and R1 lines were DNA-fingerprinted by digestion of the DNA with the HaeIII restriction enzyme and using Jeffreys' 33.6 probe. The BW of females at 4 wk of age and at the end of a 240-d egg production period were similar for the C and R1 lines. The BW of the selected lines was ranked P > T > HW for both measurements. Smooth muscle tumors were found in the oviducal ligaments adjacent to the magnum. A greater percentage of hens from the BW selected lines had smooth muscle tumors of greater weight than the randombred control lines, which did not differ in tumor incidence or weight. The P and T lines had a greater incidence of multiple-lobed tumors than the HW line. Based on bandsharing (BS) of DNA fingerprints, the Georgia and Ohio lines did not appear to be closely related, suggesting that, perhaps, the smooth muscle tumors in the BW-selected lines in the two studies might have resulted from pleiotrophic effects of genes affecting growth or to genes closely linked to the growth genes. The BW-selected lines in both selection studies had more ovarian follicles in rapid development, which were of greater weight, than the randombred control lines. The HW line had a larger number of ovarian follicles in rapid development than the P and T lines. The percentage of hens with atretic follicles was greater in the BW-selected lines. The results of the present study suggest that the effect of BW selection on ovarian follicular development may occur early in selection (within the first 30 generations) and is not influenced by additional genetic changes in BW. PMID- 10536784 TI - Variation in resistance to Pasteurella multocida among turkey lines. AB - Previous research has shown that a line (F) of turkeys selected long-term for increased 16-wk body weight was more susceptible to challenge with washed Pasteurella multocida than a randombred control line (RBC2), the base population of F. A previous study also indicated that the mortality of the F line following challenge with P. multocida was similar to that of sire lines from two of the three major primary breeders. The purpose of the present study was to compare the resistance of the sire line from the third major primary turkey breeder (C) not previously studied with that of the F and RBC2 lines to determine whether there is variation in resistance among the sire lines from three major primary breeders. The sire lines from all three primary breeders were used in the production of commercial turkeys. Body weights of the F line were greater than those of the C line at the time of challenge with P. multocida. Both the C and F lines were heavier than the RBC2 line. The birds were challenged at 6 wk of age with a field isolate of washed P. multocida (1.2 x 10(7) organism per bird of capsular serogroup A and somatic serotype 3,4). Mortality was recorded daily for 14 d. Mortality following challenge with P. multocida was higher in the F line than in the C line, and both large-bodied lines had higher mortality than the RBC2 line. Based on the present results and those published in the literature, there may be variation in resistance among commercial sire lines from the three major primary breeders. PMID- 10536785 TI - Toxicological evaluations of cyclopiazonic acid and ochratoxin A in broilers. AB - The individual and combined effects of ochratoxin A (OA) and cyclopiazonic acid (CPA) were evaluated in Petersen x Hubbard broiler chickens from 1 d to 3 wk of age. The experimental design was a 2 x 2 factorial with treatments of 0 and 2.5 mg OA/kg feed and 0 and 34 mg CPA/kg feed. Production performance, serum biochemistry, and gross pathological observations were evaluated. Body weight gain was reduced (P < 0.05) by OA, CPA, and OA-CPA in combination at the end of 3 wk. Ochratoxin A significantly increased the relative weight of the kidney and serum concentrations of uric acid and triglycerides and decreased total protein, albumin, and cholesterol. The toxicity of CPA was expressed primarily through increased relative weights of the proventriculus and increased activity of creatine kinase. Exposure to OA-CPA was characterized by increased relative weights of the liver, kidney, pancreas, and proventriculus; decreased concentrations of serum albumin, total protein, and cholesterol; increased activity of creatine kinase; and increased concentrations of triglycerides and uric acid. Postmortem examination revealed that the chickens fed CPA or OA-CPA had thickened mucosa and dilated proventricular lumen. Data from this study demonstrate that OA, CPA, and the OA-CPA combination can limit broiler performance and adversely affect broiler health. The interaction of the compounds was primarily additive or less than additive in the parameter in which the interaction occurred. PMID- 10536786 TI - Effects of dietary protein on restrict-fed broiler breeder pullets during a coccidial challenge. AB - In each of two experiments with young, feed-restricted broiler breeder pullets, the effects of differences in dietary protein intake on intestinal development and growth were studied. All pullets were restrict-fed either a 15 or 19% CP diet to see whether differences in dietary protein would influence intestinal growth in the face of controlled exposure to coccidiosis. In each experiment, pullets were vaccinated with one of three dilutions of Coccivac (control, 1X, 4X), each level representing a different proportion of the manufacturers' suggested dosage level. Experiment 1 was conducted in battery cages with wire floors, and no infection was established, most likely because of a lack of oocyst recycling. The pullets that were restrict-fed the 19% CP diet had a significantly heavier Pectoralis major breast muscle weight at 14 and 21 d postvaccination (PV) and heavier BW at 21 d PV. Experiment 2 was conducted in floor pens with litter. In this experiment, coccidiosis was successfully established as coccidial oocysts invaded the mucosal cells of the villi in the upper small intestine. Pullets fed the 19% CP diet had significantly heavier BW at 14, 28, and 35 d of age. There were, however, no significant effects caused by level of dietary protein or vaccination dose on intestinal development (villus height and crypt depth). In conclusion, mild coccidial infections induced via the administration of commercial anticoccidial vaccines do not warrant changes in dietary protein during the onset of feed restriction in young broiler breeder pullets. PMID- 10536787 TI - Evaluation of sorghum ergot toxicity in broilers. AB - Three experiments evaluated the performance of broilers fed sorghum ergot consisting of sphacelia/sclerotia of Claviceps africana present in tailings removed by conditioning of seed from grain sorghum hybrid seed production fields near Uvalde (Experiments 1 and 2) and Dumas (Experiment 3), Texas. Percentage sphacelia/sclerotia and total alkaloid content, respectively, in sorghum ergot tailings were 8% and 11.3 ppm for Uvalde and 75% and 235 ppm for Dumas. Sorghum ergot and control sorghum diets were based on the NRC (1994) requirements for starting broilers. In Experiment 1, neither growth nor feed efficiency were significantly reduced in male broilers fed sorghum ergot from hatch to 3 wk of age, but liver weights were significantly greater than those in the control. In Experiment 2, straight-run broilers were raised to 6 wk of age in floor pens using a three-phase feeding program. Sorghum ergot significantly reduced gain in 4-wk-old broilers and cumulative body weight at 5 wk. Feed conversion was significantly reduced during all three phases of feeding. In Experiment 3, control sorghum and the 75% ergot tailings were added to corn-soy basal diets at rates of 2.5, 5, and 10% by weight and fed to male broilers from hatch to 3 wk of age. Sorghum ergot did not significantly reduce growth, but, during Weeks 2 and 3, feed-to-gain ratios were higher. Neither type nor concentration of sorghum ergot significantly affected relative liver weights. We did not observe significant mortality or obvious symptoms of ergot toxicity, such as necrotic lesions of the feet or vesicular dermatitis of the comb, in any of the three experiments. PMID- 10536788 TI - Differential channelling of liver lipids in relation to susceptibility to hepatic steatosis in the goose. AB - In response to overfeeding for the production of "foie gras," the Poland goose differs from the Landes goose by a lesser susceptibility to hepatic steatosis, resulting in a lower accumulation of hepatic triacylglycerol (TG), together with a greater exportation of hepatic phospholipid (PL) in very low density lipoproteins (VLDL) and high density lipoproteins (HDL) (Fournier et al., 1997). A study was designed 1) to compare the liver composition in overfed and nonoverfed geese of the two breeds of geese and 2) to determine whether the differential channelling of lipids in response to overfeeding is reflected in the PL and fatty acid profiles of the different hepatic lipids, whether stored or secreted. In nonoverfed geese, there were no breed-related differences in liver weight (approximately 90 to 100 g), hepatic lipid content (3 to 4%), and lipid and PL composition. However, plasma VLDL and HDL of the Landes breed contained a higher phosphatidylcholine (PC) to phosphatidylethanolamine (PE) ratio than those of the Poland breed (20.7 and 33.8 vs 12.6 and 25.6 in VLDL and HDL, respectively). After 14 d of overfeeding, hepatic PL profiles were identical in the two breeds and similar to that in control livers; choline-containing PL accounted for 95% of total PL. In contrast, plasma HDL concentrations of the Landes geese were lower than those of the Poland geese (9.4 vs 12.9 g/L) and their PC:PE (13.6%) and PL-polyunsaturated fatty acids (PUFA) content (25%) were decreased compared with the Poland geese (21.2 and 30%). It is likely that the higher susceptibility to fatty liver of the Landes breed involves a differential channelling of PL, resulting in a greater hepatic retention of PC and PUFA that are necessary for plasma membrane growth and cell hypertrophy. PMID- 10536789 TI - Influence of supplemental microbial phytase on first cycle laying hens fed phosphorus-deficient diets from day one of age. AB - Pullets (19 wk of age) previously fed varying levels of aP (available P) with and without phytase (Natuphos) from Day 1 of age were used to determine the influence of 300 FTU phytase/kg diet on hen performance during Phase 1 (Week 21 to 36) and 2 (Week 37 to 48). At 19 wk of age, pullets were switched from developer diets to layer diets. The aP levels used in this portion of the study remained at 0.1, 0.2, 0.3, and 0.4% with and without phytase. These were the same levels fed in the starter and developer diets. Feed consumption, egg production, egg weight, egg specific gravity, and mortality were the criteria used. Reduction of aP from 0.4 to 0.2% had no effect on feed intake, egg production, egg weight, or eggshell quality (P > 0.05). However, hens fed 0.1% aP showed reduced feed intake, egg production, and bone mineral density (P < 0.001) and increased mortality (P < 0.001), but 300 FTU/kg phytase supplementation completely prevented these deficiencies. Eggs from hens fed 0.2, 0.3 and 0.4% aP diets were heavier than those fed 0.1% aP. Interactions between aP and phytase for feed consumption (P < 0.003), egg production (P < 0.001), and egg weight (P < 0.04) indicated that phytase corrected all deficiency symptoms in hens consuming 0.1% aP but showed no influence on hens fed aP levels > 0.2%. During Phase 2, aP by phytase interactions for feed consumption and egg production demonstrated that dietary phytase-corrected reductions related to P deficiency in hens consuming 0.2% aP. Results indicate that the addition of phytase in pullet diets from Day 1 of age through the first lay cycle can prevent reductions in performance of pullets fed low P diets. PMID- 10536790 TI - Maintenance lysine requirement and efficiency of using lysine for accretion of whole-body lysine and protein in young chicks. AB - Two bioassays were conducted to determine the maintenance requirement and efficiency of utilization of dietary Lys in young chicks. New Hampshire x Columbian males were used in Assay 1 and Avian x Avian males were used in Assay 2. In each assay, chicks were given free access for 10 d to crystalline amino acid (AA) diets containing graded levels of L-Lys.HCl. Doses of Lys represented 5, 40, 55, 70, and 95% of its ideal level in Assay 1; all other AA were set at 100% of their ideal levels, except for the lowest Lys level, in which the other AA were maintained at a 15% excess. In Assay 2, doses of Lys represented 5, 10, 40, 55, 70, and 95% of ideal; all other AA were maintained at minimized excess levels that were 15% (of ideal) above the various doses of Lys. After 24 h of feed deprivation, chicks were killed for whole-body protein and AA analysis. In Assay 1, protein accretion (r2 = 0.95) and Lys accretion (r2 = 0.98) were linear (P < 0.01) functions of Lys intake. Slope of the Lys accretion regression line indicated that 75.8% of the crystalline Lys ingested (above maintenance) was retained. The Lys required for zero protein accretion was 12 mg/d or 45 mg/d per kg3/4, whereas the Lys required for zero Lys accretion was 30.3 mg/d or 114 mg/d per kg3/4. With Avian x Avian chicks, protein accretion (r2 = 0.99) and Lys accretion (r2 = 0.99) were linear (P < 0.01) functions of Lys intake. Slope of the Lys regression line indicated that 79.3% of the Lys ingested was retained. The Lys requirement for zero protein accretion was 2.5 mg/d or 6.9 mg/d per kg3/4. The Lys maintenance requirement for zero Lys accretion, however, was 32.3 mg/d or 89.1 mg/d per kg3/4. The data demonstrated that at nitrogen equilibrium, chicks are in negative Lys balance but are in positive balance of glycine and proline. PMID- 10536791 TI - Maintenance sulfur amino acid requirements of young chicks and efficiency of their use for accretion of whole-body sulfur amino acids and protein. AB - Peterson x Hubbard male chicks were used in two bioassays conducted to determine the maintenance requirement and efficiency of utilization of dietary Met and Cys in young chicks. In each assay, chicks were given free access for 10 d to crystalline amino acid (AA) diets containing graded levels of DL-Met (Assay 1) or graded equal levels of DL-Met and L-Cys (Assay 2). Doses of Met represented 5, 10, 40, 55, 70, and 95% of its ideal level in Assay 1, with all other AA maintained at minimized excess levels that were 15% (of ideal) above the various doses of Met, except for Cys, which was maintained at 100% of its ideal level for all treatments. For example, when Met was fed at 40% of its ideal level, all other AA were fed at 55% of their ideal levels, and Cys was fed at 100%. In Assay 2, Met and Cys were fed at equal levels representing 5, 10, 40, 55, 70, and 95% of ideal with all other AA maintained at minimized excess levels that were 15% (of ideal) above the various doses of Met + Cys. After 24 h of feed deprivation, chicks were killed for whole-body protein and AA analysis. In Assay 1, Met accretion was a linear (P < 0.01) function of Met intake (r2 = 0.97). The slope of the Met accretion regression line indicated that 68% of the crystalline Met ingested (above maintenance) was retained. In Assay 2, increases in whole-body protein and whole-body TSAA were linear (P < 0.01) between dosage levels of 5 and 70% of the ideal TSAA level. Slope of the TSAA accretion line between these dose levels indicated that 52% of the TSAA was retained. The TSAA requirement for zero protein accretion was calculated to be 3.2 mg/d or 9.4 mg/d per kg3/4, whereas the TSAA required for zero TSAA accretion was 5.3 mg/d or 15.3 mg/d per kg3/4. PMID- 10536792 TI - Effects of timing and duration of feed restriction during rearing on reproductive characteristics in broiler breeder females. AB - The objective of this study was to determine whether there exists a critical period during rearing when restricted feeding of broiler breeder hens can be most beneficial on subsequent egg production. Broiler breeder pullets were subjected to combinations of either ad libitum or restricted feeding during three periods before sexual maturity. Body weight gain, feed intake, and egg production were recorded. Ovary and oviduct weights were noted at age of first oviposition. At 15 and 18 wk of age and at age of first oviposition, chicken luteinizing hormone releasing hormone-I (cLHRH-I) in the median eminence and the levels of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in the pituitary and plasma were determined. The results demonstrated that alternation between ad libitum and restricted feeding during rearing changed the growth curves of the birds. Feed restriction from 7 to 15 wk followed by either ad libitum or restricted feeding led to improved reproductive performance, suggesting that long term feed restriction may not be necessary to attain good reproductive performance. The birds restricted from 7 to 15 wk of age had higher proportional weights of ovary (> or = 1.7%) and oviduct (> or = 1.58%) at age of sexual maturity. The cLHRH-I levels in the median eminence and gonadotrophin contents in the pituitary followed that of growth in response to feeding levels and timing of feeding and could be related to the timing of the onset of lay. At age of first egg, all groups had similar levels of cLHRH-I, LH, and FSH. However, no clear effect of level of feed intake or time period of restriction could be observed for plasma LH or FSH concentrations. PMID- 10536793 TI - Embryo and yolk compositional relationships in broiler hatching eggs during incubation. AB - Developmental relationships between yolk, embryo body, and embryo liver compositions during incubation were determined in two trials. In Trial 1, embryo body moisture, fat, and CP contents and embryo liver moisture and fat contents were determined. In Trial 2, relative yolk weights, moisture, fat, and fatty acid contents, relative wet and dry embryo weights and moisture contents, and relative wet and dry liver weights and moisture contents were determined. In Trial 1, embryo moisture decreased sigmoidally between Days 6 and 21, whereas embryo fat increased between Days 12 and 21 of incubation; embryo CP displayed sequential fluctuations throughout incubation. However, an overall significant decrease in embryo CP occurred between Days 6 and 21. Liver fat content increased between Days 12 and 21, whereas liver moisture decreased through Day 18, with a subsequent increase by Day 21. In Trial 2, relative yolk weight and moisture content decreased, whereas percentage yolk lipid content increased between Days 6 and 15. Relative wet and dry embryo weights changed in a similar manner, with rapid increases between Days 12 and 18 of incubation. Embryo moisture and CP were negatively correlated to embryo fat content. Furthermore, relative embryo and liver DM were related to yolk palmitic acid concentration, whereas yolk oleic acid was correlated only with liver DM. In conclusion, embryos and their livers displayed differential accumulations of moisture and DM during incubation, and these differences exhibited distinctive associations with various yolk fatty acids. PMID- 10536795 TI - Effect of initial product temperature and initial pH on foaming time during vacuum evaporation of liquid whole eggs. AB - During earlier studies on vacuum concentration of liquid egg white, the phenomena of foaming during the initial stages of the process were reported. In these studies, it was also shown that no concentration took place during the foaming period that varied from test to test. To minimize the total process time, this present study was undertaken to investigate what variables contributed to foaming, how they could be controlled, and what effect they had on product quality and functional properties. This study investigated the relationships among initial product temperature, initial pH, and foaming of liquid whole eggs. Two temperatures (9 and 20 C) and three pH levels (6.5, 7.3, and 8.5) were studied using a vacuum evaporation system with a maximum vacuum of 5 kPa. Tests showed that higher initial pH levels had decreased foaming times. At the end of foaming experiments, the liquid whole egg was evaluated to determine the extent of functional property change during foaming. A decrease in foaming time resulted in a decrease in whip time. The cakes made from the processed liquid whole egg had larger volumes than those from the unprocessed control. Furthermore, the liquid whole egg, which foamed the longest, had higher (P < 0.05) cake volumes. Our current experiments also verified our earlier findings that no product concentration takes place during the foaming process. PMID- 10536794 TI - Venous blood pressure in broilers during acute inhalation of five percent carbon dioxide or unilateral pulmonary artery occlusion. AB - We evaluated the hypothesis that venous congestion (increased venous volume), as reflected by venous hypertension (increased venous pressure), can arise when the right ventricle is unable to elevate the pulmonary arterial pressure sufficiently to propel the cardiac output through an anatomically inadequate or inappropriately constricted pulmonary vasculature. Changes in venous pressure were evaluated in clinically healthy broilers during modest increases in pulmonary vascular resistance induced by inhalation of 5% CO2 and during large increases in pulmonary vascular resistance accomplished by acutely tightening a snare around one pulmonary artery. Inhalation of 5% CO2 induced a pronounced respiratory acidosis, as reflected by increases the partial pressure of CO2 and the hydrogen ion concentration in arterial blood. Inhalation of 5% CO2 also increased pulmonary arterial pressure by approximately 3 mm Hg and increased venous pressure by approximately 1 mm Hg when compared with the pre-inhalation venous pressure. Tightening the pulmonary artery snare increased the pulmonary arterial pressure by approximately 10 mm Hg, and this degree of pulmonary hypertension was sustained until the snare was released. When compared with the pre- and post-snare intervals, tightening of the pulmonary artery snare induced a sustained increase in venous pressure of > or = 1 mm Hg. Veins have highly compliant walls that permit an approximate doubling in volume with only small (4 to 6 mm Hg) increases in central venous pressure. Presumably the apparently modest 1 mm Hg increase in venous pressure measured after CO2 inhalation or unilateral pulmonary artery occlusion reflects a large increase in venous volume and, thus, substantial venous congestion. These observations support the hypothesis that increases in pulmonary vascular resistance can initiate increases in venous pressure by challenging the capacity of the right ventricle to propel all of the returning venous blood through the lungs. Central venous congestion predisposes broilers to the onset of cirrhosis and ascites by impeding the outflow of hepatic venous blood and increasing the hydrostatic pressure within hepatic sinusoids. PMID- 10536796 TI - Mixing of dye in a model scald tank. AB - A model scald tank was constructed to study the mixing pattern of water in a poultry scalding system. Tank dimensions were approximately 6 m long by 10.5 cm wide with a water depth of 18 cm. Water was vigorously agitated with compressed air delivered through a 1.9-cm polyvinyl chloride pipe on the bottom of the tank. Food coloring was added to the tank at a single point, and water samples were taken at distances of 0, 0.5, 1.0, 1.5, and 2.5 m every 30 s for 10 min, with 0 or 10 L/min water flow through the tank. Dye concentration was determined spectrophotometrically. A chain drive was then installed above the tank with aluminum paddles (area about 25% of tank cross-sectional area) attached to the chain every 15.2 cm to simulate the movement of carcasses through the water at 140 carcasses per minute. Food coloring was added to the tank, and water samples were taken every 15 s for 2.5 min, with 0 or 13.5 L/min water flow through the tank. A computer program based on perfect mixing of water in small slices or cells within the tank was adjusted until predicted dye movement matched sampling data, with correlations of 0.91 or better at all sampling points. For scalder designs with uniform mixing of water, the computer model can predict mixing patterns, including counterflow conditions in a single tank, well enough to yield realistic residence time patterns for bacteria suspended in scald water. PMID- 10536797 TI - Effect of ingredients and processing parameters on pellet quality. AB - Rations containing varying ratios of corn, high-oil corn, soybean meal, and mechanically expelled soybean meal were pelleted. The effects of ingredients, conditioning steam pressure, and mixing paddle configuration inside the conditioner on pellet quality were investigated. Ration ingredients strongly affected pellet quality. Increasing the protein content increased the pellet durability, whereas increasing the oil content above 7.5% greatly decreased pellet durability. High-oil corn and mechanically expelled soybean meal produced acceptable pellets when combined with soybean meal and regular corn, respectively. However, poor pellet quality resulted when rations containing high oil corn and mechanically expelled soybean meal were processed. Increasing the residence time in the conditioner by changing mixing paddle pitch resulted in an average 4.5-point increase in pellet durability indices among 65:35 (wt) corn:soybean meal and 65:35 high-oil corn:soybean meal rations. PMID- 10536799 TI - [Quality assurance: thoughts on the change of Article 58]. PMID- 10536798 TI - Meat quality and rigor mortis development in broiler chickens with gas-induced anoxia and postmortem electrical stimulation. AB - This study was conducted to evaluate the combined rigor-accelerating effects of postmortem electrical stimulation (ES) and argon-induced anoxia (Ar) of broiler chickens. One hundred broilers were processed in the following treatments: untreated controls, ES, Ar, or Ar with ES (Ar + ES). Breast fillets were harvested at 1 h postmortem for all treatments or at 1 and 6 h postmortem for the control carcasses. Fillets were sampled for pH and ratio of inosine to adenosine (R-value) and were then individually quick frozen (IQF) or aged on ice (AOI) until 24 h postmortem. Color was measured in the AOI fillets at 24 h postmortem. All fillets were then cooked and evaluated for Allo-Kramer shear value. The Ar treatment accelerated the normal pH decline, whereas the ES and AR + ES treatments yielded even lower pH values at 1 h postmortem. The Ar + ES treatment had a greater R-value than the ES treatment, which was greater than either the Ar or 1-h controls, which, in turn, were not different from each other. The ES treatment had the lowest L* value, and ES, Ar, and Ar + ES produced significantly higher a* values than the 1-h controls. For the IQF fillets, the ES and Ar + ES treatments were not different in shear value but were lower than Ar, which was lower than the 1-h controls. The same was true for the AOI fillets except that the ES and the Ar treatments were not different. These results indicated that although ES and Ar had rigor-accelerating and tenderizing effects, ES seemed to be more effective than Ar; there was little enhancement when Ar was added to the ES treatment and fillets were deboned at 1 h postmortem. PMID- 10536800 TI - [Quality management in histopathology. "From biopsy to diagnosis"]. AB - The University Institute of Pathology (IUP), Lausanne, has established a system designed to continuously document and manage the quality of its diagnostic activity in surgical pathology. To realize the system a few tools were created. Several "free codes" were introduced into the general computer system in order to register and evaluate (1) the quality of frozen section diagnoses, (2) the relevance of the use of this type of examination and (3) the frequency of spontaneous "official" internal consultations in search of a second opinion. Further, an evaluation checklist including all steps leading to the diagnosis (some of which were facultative*) was established. Eleven items were evaluated and registered in this way: quality of slides and staining, reporting time, selection of tissue blocks*, macro- and microscopic descriptions*, appropriateness of special stains and/or techniques and their quality*, final diagnosis, accuracy of codes for invoicing and diagnoses, quality of the final report. The relevance of the furnished clinical data was also evaluated and recorded. RESULTS: Of all registered frozen section examinations (397), 86.6% showed identity of provisional and definitive diagnoses. In 11.6% of cases there was a difference between the two diagnoses without clinical relevance, and in 1.8% the discrepancy was clinically significant. These percentages are comparable with those published in the literature. Internal consultation for second opinion is a current habit in our institution, and 980 cases (4%) were registered as such. Among the 25,249 cases received for histopathological examination during 1998 (each case may concern more than one specimen from the same patient), 495 (2% = 2-3 cases/day) were evaluated in detail by two reviewers throughout the year. The statistical results obtained demonstrate that our diagnostic performance in histopathology is very satisfactory: 43% of the analysed cases bear no deficit and 67.3% present two or less points in deficit; no diagnostic error was evidenced; 70% of the reports are signed-out within 3 days after reception of the material (mean delay < 3 days). CONCLUSIONS: The adopted system represents an excellent tool by which to continuously document and manage the quality of our diagnostic performances. It is reliable, efficient, sensitive and also feasible, as it does not require significant or exaggerated additional investments. The results obtained after one year confirm the high quality of the IUP's diagnostic activity. PMID- 10536801 TI - [Tenascin: a simple tool in the diagnosis of collagenous colitis]. AB - BACKGROUND: Collagenous colitis is characterised clinically by chronic, voluminous, watery diarrhoea, and histopathologically by a thickened subepithelial collagen layer and infiltration of the lamina propria with inflammatory cells. In practice, the exact measurement of this subepithelial collagen layer is complicated by its blunt limit, a nonorthogradely affected cell layer and often only focal findings. Immunohistochemically, the subepithelial collagen layer stains positively with the glycoprotein tenascin, a marker for extracellular matrix remodeling. AIM: To assess if immunohistochemical staining with tenascin could be used as a simple and practical tool in the diagnosis of collagenous colitis. METHODS: 86 of the routinely examined colon biopsy specimens with the following initial diagnosis: 33 collagenous colitis, 21 lymphocytic colitis, 15 inflammatory bowel disease (ulcerative colitis or Crohn's disease), 8 unspecific inflammation and 9 without histopathological findings, were incubated with tenascin. They were then evaluated blindly by 5 different investigators (2 senior pathologists, 2 assistant doctors and 1 chief assistant of the laboratory experienced in immunohistochemistry). Each specimen was evaluated semiquantitatively from 0 (for no tenascin-positive subepithelial collagen layer) to 3 (wide, clearly tenascin-positive collagen layer). RESULTS: In specimens with the initial diagnosis "collagenous colitis" the semiquantitatively assessed subepithelial collagen layer was significantly wider than in specimens with other initial diagnosis. Experience and educational level of the investigators did not influence the results. CONCLUSIONS: Immunohistochemical incubation with tenascin is a simple, economical and rapid tool in the diagnosis of collagenous colitis. With the aid of tenascin staining collagenous colitis can be differentiated histopathologically with sufficient accuracy from other colitis. PMID- 10536802 TI - [Whiplash-associated disorders]. AB - Whiplash-associated disorders (WAD) represent a class of clinical complaints which commonly result from rear-end car accidents. An automobile collision can generate major forces which are transferred to the neck by an acceleration deceleration mechanism (whiplash), resulting in bony or soft-tissue injuries (whiplash injury). Incidence of WAD is estimated to be 0.1 to 3.8/1000/year; WAD cost $29 billion a year in the USA. They can be classified clinically into 5 degrees of severity, namely WAD grades 0 to IV. Signs and symptoms typically crescendo during the first few days after an accident. Pathological findings (especially of musculo-skeletal or neurological types) must often be sought actively and should be documented at the earliest stage. Prevention of possible chronicity is the most important goal in clinical management of WAD. WAD grade IV patients are treated in the way their fracture or dislocation demands. Therapy of WAD grades I to III has three main aspects: non-narcotic analgesics, early active mobilisation (to the extent possible consistent with pain) and education of the patient. Soft collars should not be used (or only temporarily and sparingly). Most patients with WAD grades I-III feel well again relatively soon. Symptoms and signs that persist for longer than two months are important warning signs for imminent chronicity, which occurs at rates of 14-42%. In such cases, an interdisciplinary approach is recommended. Risk factors are accident severity, head position at the time of accident, age and pretraumatic existence of headache. Patients with chronic complaints can develop additional psychic and cognitive problems, which are caused by--and not the cause of--their chronic disorder. Therapy of chronic whiplash-associated disorders involves all the problems inherent in therapies of chronic pain. There are many therapeutic concepts, but little evidence that anything helps. Prevention of whiplash injuries is therefore very important in view of the lack of powerful treatment options. Although there is a substantial body of scientific literature about WAD, many unanswered questions remain. In particular the most important questions (how can patients with acute and chronic disorders be helped best) have no clear answer yet. Furthermore, there are many opinions and prejudices (especially concerning psycho-social factors of WAD) which have no scientific basis. Therefore, an intensive exchange of information between health care professionals, patients and the general public appears to be very important. PMID- 10536803 TI - [Diagnostic error: bronchial asthma]. PMID- 10536804 TI - [Benign myoclonic epilepsy in infancy]. AB - Benign myoclonic epilepsy in infancy (NMEI) is one of rare epileptic syndromes. 5 patients (all female sex) aged 4-16 years were observed. NMEI debuted at the age from 7 months till 2.5 years (mean age 1.3 years). Pathology of pregnancy and labor, disorders in both psychomotor development and genetic predisposition were not found. In all the cases the disease began with typical transitory repeated myoclonic paroxysms of different intensity and frequency, without loss of consciousness and with primary involvement of the muscles of the neck and the upper extremities. Most patients had muscular hypotension, mild coordinatory disorders, delayed psycho-speech development, mental retardation, EEG signs of generalized epileptic activity. Valproates, suxilep, clonazepam and lamotrigin (lamiktal) were used for treatment. The most pronounced effect was achieved using either monotherapy with valproates (depakin) or a combination depakin + lamiktal. A stable clinical-encephalographic remission was achieved in all the patients, but during puberty in 2 patients (15 and 16 years old) rare generalized convulsive fits debuted. High frequency of intellectual-mnestic disorders were found even after a complete remission. So benign definition concerns only a course of the fits, but not NMEI prognosis. PMID- 10536805 TI - [Psychopathology-like states after craniocerebral trauma]. AB - The presence of stable personal disharmony in the form of a reduced organic mental syndrome served as a criterion for diagnosis of personality changes after traumatic head injuries. The patients were divided into a group with prevalence of personality changes (81 patients) and a group with predominance of pronounced organic mental syndrome (141 patients). Personality disorders prevailed in the patients who had got mild and moderate traumas at the age of 13-25 years, with less number of additional pathogenic factors of exogenic organic spectrum in anamnesis. 3 variants of the course of personality disorders were recognized: with a tendency toward reduction of emotional and drive pathology; with a tendency to intensification of intellectual and mnestic disorders and transformation into large-scale organic mental syndrome; with a tendency toward an increase of pathological traits of personality in conditions of growing influence of the environmental and psychogenic factors. PMID- 10536806 TI - [The psychological characteristics of patients with drug addictions]. AB - The paper presents results of the examination of 70 patients with different forms of drug addiction (opium, heroin, pervitin addictions, including patients with polynarcomania). Psychologic tests were used to evaluate personal traits of the patients (MMPI variation), peculiarities of the emotional sphere (test of Lusher) and the intellectual level (Progressive Matrices of Raven). It was revealed that chronic drug dependence resulted in considerable disorders of personality- psychopathization and schizoidization. Intellectual abilities of the patients were on the low normal level. PMID- 10536807 TI - [Depakin 300 and Depakin-chrono in the therapy of epilepsy]. AB - The results of administration of Depakin 300 and Depakin-chrono during 4-year period were analysed in 137 patients aged 14-36 years with idiopathic generalized, cryptogenic, symptomatic partial and nonclassified epilepsy. Depakin was prescribed as basic medicine in idiopathic generalized epilepsy, while in partial and nonclassified epilepsy this drug was more frequently administered in combination with some other anticonvulsants. Early administration of Depakin was highly effective: a remission was observed in 91.2% of the patients during a year. During 3 years a remission was observed in 75.4%, frequency of the fits decreased more than by 50%--in 14.1% of the cases, no response was found in 10.5% of the patients (in cases of gross structural changes of the brain, inefficiency of the preceding therapy during 5 years). Frequency of side-effects was 1.7 12.3%. Indications to treatment are discussed: in addition to generally accepted recommendations (absence, myoclonic primary generalized, tonic-clonic forms of epilepsy, of reading, photosensitive epilepsy), application of Depakin is also recommended in nonclassified epilepsy with generalized epileptic EEG-patterns as well as a drug of choice in all cases of nonclassified fits. Depakin is also quite efficient as an additional drug in cases of partial epilepsy resistant to treatment. PMID- 10536808 TI - [Transcranial magnetic stimulation in neurotic depression]. AB - Transcranial magnetic stimulation (TMS) was applied in combination with psychotherapy in patients with neurotic depression, including 15 patients of the experimental group and 14 patients of the control one. 10 sessions of daily TMS for the patients from the experimental group (0.015 T, 40 pulses per sec) were performed at the same time for 20 min (twice for 10 min with 5-min interval) in a room which excluded any external stimulation. TMS was performed by contact method: 5 cm coil was applied to the left prefrontal area. The control group received the imitation of TMS-procedure stimulation. The improvement of mental state was in 13 patients of experimental group and in 3 of control one. The course of TMS resulted in a significant attenuation of depression by the Hamilton Depression Rating scale (from 22.9 to 8.6) and the Anxiety Inventory (from 39.4 to 26.6), that was significantly higher in comparison with the control. There weren't found any TMS-related changes in blood pressure and pulse rate as well as any pathological EEG symptoms. PMID- 10536809 TI - [The use of sirdalud in cerebral palsy in children]. AB - 30 diplegic children (mean age 11.3 +/- 2.8 years old) with severe form of cerebral palsy received sirdalud monotherapy during 2-6 weeks (1 mg for children under 10 years old and 2 mg for older children, 3 times daily). Positive effects were determined in motor, autonomic and mental (emotional) spheres. Sirdalud was also very effective in patients after orthopedic-surgical treatment. Electroneuromyographic analysis showed the decrease of the synergic tonic activity, as well as the improvement of the supraspinal influences and the segmental interaction. Thus, the small doses of sirdalud are effective without side effects in children with cerebral palsy. PMID- 10536810 TI - [The use of the clinical biochemical Epitest kit in children with epilepsy]. AB - 57 children aged 3-15 years were examined: children with epilepsy (n = 26), children with epileptic syndrome (n = 16) and children with other neurologic pathology (n = 15). Control group included 25 healthy children. Diagnostic efficiency of the epileptic test was investigated on the basis of the detection of autoantibodies (aAb) to glutamate receptor of AMPA type in blood serum of the patients. It was shown that children with epilepsy had elevated level of aAb to AMPA receptors in blood in comparison with that of the healthy children. The level of aAb to AMPA receptors correlated with severity of the disease and location of the epileptic focus. The authors recommend the "Epitest" as an additional biochemical tool for differential diagnosis of children's epilepsy. PMID- 10536811 TI - [The mechanisms of the occurrence and development of the epileptic range of disorders]. AB - An attempt was made to approach to the causes of the development of epileptic disorders from general biologic positions and to determine the signs of a normal organism, that might help to predict a risk of the disease manifestation. A pattern of electric activity of the brain of the animals, both sensitive and resistant to convulsive agents' action in terms of the influence of convulsive and anticonvulsive preparations on EEG are presented. A role of the brain's mass and of the peculiarities of dermatoglyphics as the predictors of the development of epileptic disorders are also discussed. PMID- 10536812 TI - [The role of genetic factors in the manifestation of delirium tremens]. AB - Among 27,692 patients suffering from alcoholism, delirium tremens occurs only in 8.1% of the cases. From 2417 patients with this psychosis repeated delirium tremens was noted in 12.9%. In the studied group (n = 2417) delirium tremens was observed in men (84.2%) 5.3 times more often than in women (15.8%). The mean age of the onset of the delirium was 43.2 years in women and was older than in the group of men (42.0 years). Age distribution of the disease onset in men and women follows a curve of normal distribution. In 125 patients with schizophrenia combined with delirium tremens there was a high rate of repeated alcoholic delirium (22.4%), epileptic seizures (21.6%), diabetes mellitus of type II (10.4%). These facts confirm the role of hereditary predisposition to development of delirium tremens and allow to relate alcoholic delirium to multifactorial diseases. PMID- 10536813 TI - [An epidemiological study of epilepsy in Moscow]. PMID- 10536814 TI - [Paraneoplastic syndromes]. PMID- 10536815 TI - [The shaping of priorities in neurological science on the eve of the 21st century]. PMID- 10536816 TI - Dissociation energies of deoxyribose nucleotide dimer anions measured using blackbody infrared radiative dissociation. AB - The dissociation kinetics of deprotonated deoxyribose nucleotide dimers were measured using blackbody infrared radiative dissociation. Experiments were performed with noncovalently bound dimers of phosphate, adenosine (dAMP), cytosine (dCMP), guanosine (dGMP), thymidine (dTMP), and the mixed dimers dAMP.dTMP and dGMP.dCMP. The nucleotide dimers fragment through two parallel pathways, resulting in formation of the individual nucleotide or nucleotide + HPO3 ion. Master equation modeling of this kinetic data was used to determine threshold dissociation energies. The dissociation energy of (dGMP.dCMP-H)- is much higher than that for the other nucleotide dimers. This indicates that there is a strong interaction between the nucleobases in this dimer, consistent with the existence of Watson-Crick hydrogen bonding between the base pairs. Molecular mechanics simulations indicate that Watson-Crick hydrogen bonding occurs in the lowest energy structures of (dGMP.dCMP-H)-, but not in (dAMP.dTMP-H)-. The trend in gas phase dissociation energies is similar to the trend in binding energies measured in nonaqueous solutions within experimental error. Finally, the acidity ordering of the nucleotides is determined to be dTMP < dGMP < dCMP < dAMP, where dAMP has the highest acidity (largest delta Gacid). PMID- 10536817 TI - Laser desorption in transmission geometry inside a Fourier-transform ion cyclotron resonance mass spectrometer. AB - We report here the first application of laser desorption (LD) in transmission geometry (backside irradiation of the sample through a transparent support) inside a Fourier-transform ion cyclotron resonance mass spectrometer (FT-ICR). A probe-mounted fiber optic assembly was used to simplify the implementation of this LD technique. This setup requires little or no instrument modifications, has minimum maintenance requirements, and is relatively inexpensive to build. The performance of the probe was tested by determining the molecular weight of a commercial polystyrene standard from its matrix-assisted laser desorption/ionization (MALDI) spectrum. The measured average molecular weight is comparable to that obtained for the same sample by MALDI in the conventional top illumination arrangement (reflection geometry) and by the manufacturer of the sample by gel permeation chromatography. The average velocities measured for ions evaporated by transmission mode LD of several neat samples are about half the velocity of those obtained by using the reflection geometry. Therefore, transmission mode irradiation of the sample holds promise to desorb ions that are easier to trap in an ICR cell. An oscillating capillary nebulizer was adapted for the deposition of analytes to improve sampling reproducibility. PMID- 10536818 TI - Calcium-induced noncovalently linked tetramers of MRP8 and MRP14 detected by ultraviolet matrix-assisted laser desorption/ionization mass spectrometry. AB - MRP8 and MRP14 are members of the S100 family of calcium-binding proteins which play an important role during calcium-induced activation of phagocytes. Both proteins form noncovalently associated complexes as a prerequisite for biological functions. The exact stoichiometric composition of these complexes, however, has not been completely clarified yet. In the present study we show for the first time by ultraviolet matrix-assisted laser desorption/ionization mass spectrometry (UV-MALDI-MS) the calcium-induced formation of noncovalently associated (MRP8/MRP14)2 tetramers. Furthermore, we could determine posttranslational modifications of MRP8 and MRP14, the stoichiometric proportion of the two known MRP14 isoforms in the complexes as well as the number of calcium ions bound to the single MRP8 and MRP14 monomers and tetramers. MRP14 showed a higher affinity for calcium than MRP8. Upon complex formation the calcium binding increased to maximal saturation of the known EF hands in the complexed forms. Calcium-induced stabilization of the MRP8/MRP14 complexes was confirmed by DSC studies. Our results extend scope and application of UV-MALDI-MS by allowing identification of noncovalent protein complexes, the identification of minor alterations of subunits in such complexes as well as the determination of bound calcium ions. PMID- 10536819 TI - Use of an internal control for matrix-assisted laser desorption/ionization time of-flight mass spectrometry analysis of bacteria. AB - A method to aid in the analysis of bacterial samples of unknown concentration by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry is demonstrated. It is shown that in MALDI analysis of bacteria, the intensities of resulting peaks in spectra are sensitive to the microbial concentration. At the high and low ends of the concentration range, no signal can be obtained, leaving very concentrated or very dilute samples indistinguishable. The addition of cytochrome c as an internal control allows the differentiation of these concentrated and dilute samples. The presence of the internal control causes only a 20% to 30% decrease in signal intensity when the bacterial concentration is optimum. However, the signal quality is improved when the internal control is added to some low concentrations of bacteria. PMID- 10536820 TI - Electrospray ionization mass spectrometric analysis of microcystins, cyclic heptapeptide hepatotoxins: modulation of charge states and [M + H]+ to [M + Na]+ ratio. AB - Electrospray ionization mass spectrometry was used to develop a rapid, sensitive, and accurate method for determination and identification of hepatotoxic microcystins, cyanobacterial cyclic heptapeptides. To optimize the electrospray ionization conditions, factors affecting charge state distribution, such as amino acid components of sample, proton affinity of the additives, and additive concentration, were investigated in detail and a method for controlling charge states was developed to provide molecular-related ions for assignment of molecular weight and reasonably abundant precursor ions for MS/MS analysis. A procedure for identification of microcystins consisting of known amino acids was proposed: for microcystins giving abundant [M + 2H]2+ ions, the addition of nitrogen-containing bases to the aqueous sample solution is effective to obtain an increased intensity of [M + H]+ ions, whereas the addition of Lewis acids containing nitrogen can produce increased abundances of [M + 2H]2+ ions for microcystins giving weak [M + 2H]2+ ions. Microcystins possessing no arginine residue always give sodium adduct ions [M + Na]+ as the base peak, and these are difficult to fragment via low energy collision-induced dissociation to yield structurally informative products; the addition of oxalic acid increases [M + H]+ ion abundances, and these fragment readily. PMID- 10536821 TI - Determination of 3,3'-dichlorobenzidine and its degradation products in environmental samples with a small low-field Fourier transform ion cyclotron resonance mass spectrometer. AB - 3,3'-Dichlorobenzidine (DCB) and its degradation products, 3-chlorobenzidine (MCB) and benzidine, are of environmental concern because of their carcinogenic nature. The suitability of a small Fourier transform ion cyclotron resonance (FT ICR) mass spectrometer for the analysis of these environmental contaminants in different types of matrices was explored. All the measurements were carried out by depositing the sample solution directly on a disk that was introduced into the mass spectrometer. This approach is very fast and simple because it requires no prior chromatographic separation or derivatization. Calibration curves determined by collecting 70-eV electron ionization mass spectra of neat samples yielded lower limits of detection of 29 and 17 pg (total amount on the solids probe) for DCB and benzidine, respectively (based on a signal to noise ratio of > or = 2:1), while chemical ionization with ammonia resulted in lower limits of detection of 21 pg for DCB and 9 pg for benzidine (total amount on the solids probe). FT-ICR analysis of sediments collected from Lake Macatawa (Holland, MI) verified the presence of DCB in this complex, environmentally significant sample matrix. Laboratory experiments designed to probe biodegradation and photodegradation pathways showed that DCB undergoes sequential dehalogenation to yield MCB and then benzidine under exposure to microorganisms and under simulated tropospheric solar radiation. The ability of the FT-ICR to determine elemental compositions of compounds introduced as described above was demonstrated for one of the degradation products. PMID- 10536822 TI - A database of 660 peptide ion cross sections: use of intrinsic size parameters for bona fide predictions of cross sections. AB - An ion trap/ion mobility/time-of-flight mass spectrometry technique has been used to measure collision cross sections for 660 peptide ions generated by tryptic digestion of 34 common proteins. Measured cross sections have been compiled into a database that contains peptide molecular weight and sequence information. The database is used to generate average intrinsic contributions to cross section (size parameters) for different amino acid residues by solving systems of equations that relate the unknown contributions of individual residues to the sequences and cross sections of database peptides. Size parameters are combined with information about amino acid composition to calculate cross sections for database peptides. Bona fide cross section predictions (made prior to measurement) for peptides observed in tryptic digests of sperm whale myoglobin and yeast enolase are made. Eight of 10 predicted cross sections are within 2% of the experimental values and all 10 are within 3.2%. The utility of size parameters for cross section prediction is explored and discussed. PMID- 10536823 TI - Reversed-phase liquid chromatographic separation of complex samples by optimizing temperature and gradient time I. Peak capacity limitations. AB - The separation of samples that contain more than 15 to 20 analytes (n > 15-20) is typically difficult and usually requires gradient elution. We have examined the reversed-phase liquid chromatographic separation of 24 samples with 8 < or = n < or = 48 as a function of temperature T and gradient time tG. The required peak capacity was determined for each sample, after selecting T and tG for optimum selectivity and maximum sample resolution. Comparison of these results with estimates of the maximum possible peak capacity in reversed-phase gradient elution was used to quantify the maximum value of n for some required sample resolution (when T and tG have been optimized). These results were also compared with literature studies of similar isocratic separations as a function of ternary solvent mobile phase composition, where the proportions of methanol (MeOH), tetrahydrofuran (THF) and water were varied simultaneously. This in turn provides information on the relative effectiveness of these two different method development procedures (optimization of T and tG vs. % MeOH and % THF) for changing selectivity and achieving maximum resolution. PMID- 10536824 TI - Reversed-phase liquid chromatographic separation of complex samples by optimizing temperature and gradient time II. Two-run assay procedures. AB - By optimizing column temperature T and gradient time tG, complex samples can often be separated by means of reversed-phase high-performance liquid chromatography (RP-LC). Conclusions reached in Part I suggest that the complete separation of such samples will be difficult, however, when more than 15-20 components are present in the sample. An alternative approach is to carry out two separations with different conditions (T, tG) in each run. The combination of results from these two runs then allows the total analysis of the sample, providing that every sample component is adequately resolved in one run or the other. Examples of this approach, carried out by means of computer simulation, are shown here for several samples of varying complexity. Also considered is the ability of a single separation where T and tG are optimized to enable the separation and analysis of one or more individual sample components from complex mixtures (e.g., drugs in animal plasma), including the resolution of isomeric compounds from each other. PMID- 10536825 TI - Reversed-phase liquid chromatographic separation of complex samples by optimizing temperature and gradient time III. Improving the accuracy of computer simulation. AB - Previous studies have shown that four experimental runs, where both temperature T and gradient time tG are varied, can be used for the reliable prediction of separation as a function of these two variables (two-dimensional optimization). Computer simulation (e.g., DryLab) can then be used to predict "optimized" conditions for maximum sample resolution using either isocratic or gradient elution. Samples that contain a large number of components (e.g., n>15-20) present a greater challenge. Resolution for these more complex samples is often quite sensitive to small changes in T or tG in turn requiring greater accuracy in predictions that result from computer simulation. In the present study of several samples, we have examined computer simulation errors that can arise from inexact expressions for retention time as a function of T, tG or isocratic %B. Resulting conclusions are applicable to both complex and simpler samples, in either one- or two-dimensional optimization. Means to anticipate and minimize the impact of these predictive errors are examined. PMID- 10536826 TI - Influence of column radial heterogeneity on peak fronting in linear chromatography. AB - Using numerical calculations of elution peak profiles, an explanation of the fronting behavior of elution peaks in linear chromatography was found in certain radial distributions of the mobile phase flow velocity and local bed efficiency. Fronting peaks are observed only if the flow velocity is higher in the wall region than in the center part of the column and the local efficiency is lower near the wall than in the center. By contrast, tailing or symmetrical peaks are observed if only the flow velocity or the local efficiency are radially heterogeneous. The degree of peak fronting increases with increasing amplitude of the radial distributions. The influence of the radial heterogeneity of the flow velocity on the degree of peak fronting is more severe for high than for low efficiency columns. An equation is suggested to correlate peak fronting behavior for columns of different efficiencies and a procedure proposed for the estimation of the radial distributions of the flow velocity and the local efficiency by analyzing some characteristics of asymmetric peaks. PMID- 10536827 TI - Size-exclusion chromatography with on-line ultraviolet, proton nuclear magnetic resonance and mass spectrometric detection and on-line collection for off-line Fourier transform infrared spectroscopy. AB - The coupling of HPLC with UV detection and on-line NMR spectroscopy and mass spectrometry combined with a dedicated interface for the collection of the chromatographic eluent for subsequent Fourier transform (FT) IR has been investigated using a number of polymer additives as model compounds. Size exclusion chromatography was performed using deuterated chloroform as eluent with the separation monitored on-line by UV detection at 254 nm and on-flow 1H-NMR and MS. The effluent from the NMR probe was directed to a dedicated HPLC interface where it was deposited on a germanium plate for subsequent FT-IR. NMR and MS spectra were successfully obtained for 2,6-di-tert.-butyl-4-methylphenol, octadecyl-3-(3,5-di-tert.-butyl-4-hydroxyphenyl) propionate (Irganox 1076) and diisooctyl phthalate on-line and FT-IR spectra for all three compounds were obtained off-line. Practical problems encountered with this multiple hyphenation are described. PMID- 10536828 TI - Efficient solid-phase extraction procedures for trace enrichment of priority phenols from industrial effluents with high total organic carbon content. AB - Polymeric solid-phase extraction (SPE) cartridges and Speedisks were used to extract 17 phenols from HPLC-grade water, tap water, river water and industrial effluents. With SPE cartridges, recoveries between 60 and 120% were obtained for waters with a total organic carbon (TOC) content below 20 mg C/l. However, when extracting industrial effluent waters with higher TOC values (75 mg C/l), only the polar phenols were recovered from the water fraction. Nonpolar compounds (di , tri- and tetrachlorophenols) remained attached to the particulate matter and were recovered from the 0.45-microm filter membrane disks by Soxhlet extraction. Speedisks offered a high efficiency and permitted one to extract phenols without a prior filtration step. Acceptable recoveries were obtained when processing heavily charged industrial effluents with a TOC of 505 mg C/l. Liquid chromatography with electrochemical detection was used for the routine determination of 17 priority phenols. PMID- 10536829 TI - Discrimination between enantioselective and non-selective binding sites on cellobiohydrolase-based stationary phases by site specific competing ligands. AB - A systematic study was performed to investigate the influence of cellobiose or lactose on the enantioselective retention behaviour of some beta-blockers in liquid chromatography using Cellobiohydrolase (CHB) I from Trichoderma reesei or Cellobiohydrolase 58 from Phanerochaete chrysosporium immobilized on silica as stationary phases. The results revealed that the retention could be described by the function [equation; see text] where the observed capacity factor corresponds to the sum of an enantioselective mode being influenced by a site specific competing ligand (competitor) and a non-selective mode unaffected by the competitor. A non-constrained non-linear least-square regression gave in all cases virtually identical nondisplacable capacity factors (k'ns) for both enantiomers of the same drug. The experimental capacity factors (k'(x,C)) of the enantiomers all show a close fit to the adapted function. The Kd values calculated for the competitor were also virtually identical for each pair of enantiomers and were in accordance with Ki data determined for the competitors in classical enzyme kinetics experiments, demonstrating that one unique site; namely, the catalytic site, was responsible for the enantioselective binding. Similar results were obtained with the resolution of rac-alprenolol and rac metoprolol on CBH I phase. PMID- 10536830 TI - Uniform-sized molecularly imprinted polymer for (S)-ibuprofen retention properties in aqueous mobile phases. AB - A uniform-sized molecularly imprinted polymer for (S)-ibuprofen has been prepared by a multi-step swelling and thermal polymerization method using 4-vinylpyridine (4-VPY) and ethylene glycol dimethacrylate (EDMA) as a host functional monomer and cross-linker, respectively. The obtained (S)-ibuprofen imprinted 4-VPY-EDMA materials were evaluated using aqueous eluents by HPLC. Hydrophobic and hydrogen bonding interactions between ibuprofen enantiomers and 4-VPY-EDMA materials could play an important role in the retentivity and enantioselectivity. Further, partial resolution of the enantiomers of ibuprofen metabolites, 2-hydroxy- and 2 carboxyibuprofen, was attained with the (S)-ibuprofen imprinted 4-VPY-EDMA materials. PMID- 10536831 TI - Enantiomeric purity determination of acetyl-L-carnitine by reversed-phase high performance liquid chromatography using chiral derivatization. AB - An indirect HPLC enantioseparation method for the determination of acetyl-D carnitine (D-AC) in acetyl-L-carnitine (L-AC) was developed. L-AC was derivatized with a chiral amino compound which has a chromophore for UV detection. Six chiral amino compounds were examined as chiral derivatization reagents. Among them, enantiomers of acetylcarnitine derivatized with L-alanine-beta-naphthylamide (L Ala-beta-NA) were successfully separated on an ODS column within 10 min with Rs = 1.94 and alpha = 1.10. Quantitation was achieved through UV detection at 254 nm. The derivatization reaction of L-AC with L-Ala-beta-NA was completed in less than 10 min at room temperature (ca. 20 degrees C). Validation data such as linearity, detection limit, and precision are also presented. The detection limit of D-AC in L-AC in this method was below 0.05% (visual evaluation). This method was found to be applicable as a practical quality control method for the enantiomeric purity determination of L-AC. PMID- 10536832 TI - Continuous-bed chromatography for the analysis and purification of recombinant human basic fibroblast growth factor. AB - The chromatographic properties of the commercial cation exchanger UNO-S1 (35x7 mm) was investigated using lysozyme from hen egg white as model protein and recombinant human basic fibroblast growth factor (rh-bFGF) from a high cell density cultivation of E. coli. The dynamic capacity for lysozyme (c(o) = 1 mg/ml) in 100 mM acetate buffer, pH 5 was 27 mg per ml sorbent. It was found independent of the flow-rate from 78 to 935 cm/h owing to the absence of mass transfer restrictions with this column concept. Regarding the selectivity for rh bFGF and the capacity for lysozyme, no changes were apparent after cleaning-in place (CIP) procedures with 0.5 M NaOH. Clogging of the column by a clarified crude cell homogenate of E. coli was not critical as precipitates were removed by reversal of the flow during CIP. Rh-bFGF elutes in three consequent peaks from the UNO-S1 column, which could be attributed to soluble rh-bFGF aggregates of different size. The dynamics of rh-bFGF aggregation and reaggregation in the crude feedstock was monitored by fast gradient elution chromatography. PMID- 10536833 TI - Direct chromatographic resolution of carnitine and O-acylcarnitine enantiomers on a teicoplanin-bonded chiral stationary phase. AB - R-(-)-Carnitine (vitamin B(T)) plays an important role in human energy metabolism, by facilitating the transport of long-chained fatty acids across the mitochondrial membranes. Its (S)-enantiomer acts as a competitive inhibitor of carnitine acetyltransferase, causing depletion of the body R-(-)-carnitine stock. Consequently, the separation of carnitine enantiomers is very important both to study their biological activities and to control the enantiomeric purity of pharmaceutical formulations. In the present paper we describe an easy, fast and convenient procedure for the separation of the enantiomers of carnitine and O acylcarnitines by enantioselective HPLC on a laboratory-made chiral column containing covalently bonded teicoplanin as selector. High enantioselectivity factors (alpha values ranging from 1.31 to 3.02) and short-time analyses characterize the analytical procedure; in addition, analytes are easily detected by evaporative light scattering with no need for preliminary derivatization. The effects of pH and ionic strength of the mobile phase and of the nature of the organic modifier on the enantioselective separations were also investigated. PMID- 10536834 TI - Ultra-trace-level determination of polar pesticides and their transformation products in surface and estuarine water samples using column liquid chromatography-electrospray tandem mass spectrometry. AB - A method is developed for the determination of polar pesticides and their transformation products [atrazine, deethylatrazine, deisopropylatrazine, hydroxyatrazine, diuron, 3,4-dichlorophenylmethylurea, 3,4-dichlorophenylurea (DPU), monuron, bentazone, anthranil-isopropylamide, chloridazon, metolachlor] in surface, estuarine and sea water samples at the low ng/l level. Solid-phase extraction is combined off-line with column liquid chromatography-electrospray ionization tandem mass spectrometric detection (LC-ESI-MS-MS). The applicability of two solid-phase materials, i.e., LiChrolut EN cartridges and graphitized carbon black extraction disks, is evaluated. The influence of the organic solvent used in gradient LC, as well as the amount of co-extracted humic material on the ESI process is studied. The eluotropic strength of the organic solvent was found to have a distinct effect on the sensitivity of ESI-MS if coupled with LC gradient separations. Methanol gave much better results than acetonitrile and phenylurea compounds are more susceptible to solvent changes than triazines. Co extracted humic material causes signal suppression in ESI-MS-MS detection. The degree of suppression depends upon the sample pH and the nature of the samples, i.e., surface or estuarine water. Detection limits in LC-ESI-MS-MS ranged from 0.2 to 2 ng/l, with the exception of DPU (8 ng/l). The applicability of the procedure was demonstrated by analyzing surface and estuarine water. PMID- 10536835 TI - Computer-aided development of a high-performance liquid chromatographic method for the determination of hydroxyanthraquinone derivatives in Chinese herb medicine rhubarb. AB - With computer simulation predicting separation in reversed-phase gradient elution, a method to separate and determine five hydroxyanthraquinone derivatives having a wide range of polarity in extract of Chinese herbal medicine rhubarb has been developed. The software DryLab was used to model the retention behavior of the compounds as a function of gradient conditions, based on data from two scouting gradient runs. Under the optimized conditions, i.e. column, Zorbax RX C18, 5 microm, 15x0.46 cm; solvent A, 36 mM triethylamine phosphate (TEAP), pH=2.5; solvent B, ACN; gradient, 36/36/80/80% B at 0/5.5/20.5/25.5 min; flow rate, 1.00 ml/min; temperature, ambient, the method was successfully applied to monitor the quality of rhubarb from different sources. The effect of sample preparation procedures on extraction efficiency was also examined. PMID- 10536836 TI - Ultra-short columns for low-pressure ion chromatography. AB - Since the development of low-pressure ion chromatography (LPIC), many inorganic cations and anions as well as organic acids could be analyzed at a low-pressure of 1.96 x 10(5)-2.94 x 10(5) Pa. And the ultra-short columns took the place of the common long chromatographic columns. Furthermore, the ultra-short columns of LPIC not only reduced the system pressure appreciably, but also achieved high sensitivity and precision. In order to assess the characteristics of the super short columns, much analysis, i.e., the analysis of alkali metals, alkaline earth metals, transition-metals, rare-earth elements, inorganic anions and organic acids as well as the common amino acids, were conducted. Moreover, excellent results were obtained from the LPIC columns. PMID- 10536837 TI - Integrated approach to the multidimensional analysis of complex biological samples by microseparation techniques. Analysis of glycoprotein factor associated with cancer cachexia. AB - Microanalytical separation techniques including capillary liquid chromatography, capillary electrophoresis and capillary electrochromatography are suitable for detection of diagnostically important changes in the metabolic profiles of biological fluids. A prototype instrument was employed to serve as an integrated platform for the analysis of urine sample from patients suffering from cancer cachexia. The instrument provides for convenient, rapid and efficient multidimensional approach towards method development which would facilitate simultaneous analysis of complex biological mixtures by the above techniques. PMID- 10536838 TI - Magnetic split-flow thin fractionation of magnetically susceptible particles. AB - We recently built a magnetic separation system to extend the applications of split-flow thin (SPLITT) fractionation to magnetically susceptible particles. Here, we characterize the magnetic SPLITT system using magnetically susceptible particles and ion-labeled particles. The flow axis of separation channel was orientated parallel and perpendicular to gravitational forces to exclude and include, respectively, gravitational effects on separation. Both operating modes were used to test the theory experimentally, with emphasis on the parallel mode. The magnetic susceptibilities of carrier and ion-labeled particles were varied, and various ion-labeled and unlabeled particles were studied experimentally, resulting in successful separation of labeled particles, yeasts, and cells from unlabeled ones. The minimal difference in magnetic susceptibility (delta(chi)) required for complete particle separation was about 1.75 x 10(-5) [cgs], corresponding to about 10(9) labeling ions per particle in this study. The throughput was around 7.2 x 10(8) particles/h using the present setup. Magnetic SPLITT fractionation shows good potential for use in obtaining particles magnetic susceptibilities from a simple theoretical treatment. PMID- 10536839 TI - Analysis of methyl tert.-butyl ether and its degradation products by direct aqueous injection onto gas chromatography with mass spectrometry or flame ionization detection systems. AB - A method was developed to analyze methyl tert.-butyl ether (MTBE) and its degradation products by gas chromatography with mass spectrometry (GC-MS) or flame ionization detection (FID) with direct aqueous injection. The column had dimensions of 30 m x 0.25 mm with film thickness 0.25 microm and a stationary phase of FFAP (nitroterephthalic acid-modified polyethylene glycol). The optimized GC conditions for non-acid components were as follows: carrier gas flow rate,l mL/min; oven temperature, 35 degrees C for 5.5 min, ramped to 90 degrees C at 25 degrees C/min, then ramped to 200 degrees C at 40 degrees C/min and held at 200 degrees C for 8 min. The conditions for the acid components were: carrier gas flow-rate, 1 mL/min; oven temperature, 110 degrees C for 2 min, ramped to 150 degrees C at 10 degrees C/min, then ramped to 200 degrees C at 40 degrees C/min. The injection port contained a silanized-glass reverse-cup liner filled with Carbofrit. The minimum concentrations for the linear range for the selective ion monitoring mode were 30 to 100 microg/L, depending on the analytes. The minimum detection limit was 1 mg/L for MTBE and tert.-butanol when using FID. More components could be analyzed with the FFAP-type column than with the cyanopropylphenyl-dimethyl polysiloxane-type column. PMID- 10536840 TI - Simultaneous determination of halogenated neutral and acidic disinfection by products in drinking water by closed-loop stripping extraction and capillary gas chromatography. AB - The analytical capabilities of Grob closed-loop stripping analysis technique were evaluated for the determination, in drinking water, of trihalomethanes, haloacetonitriles, haloacetic acids, chloropicrin, halogenated ketones and chloral hydrate, reported as chlorination disinfection by-products. Thus by one step enrichment and isolation procedure and subsequent analysis by capillary gas chromatography with electron-capture detection, organic polar and non-polar disinfection by-products could be analyzed at levels as low as 0.5 ng/l for trihalomethanes, 1 ng/l for haloacetonitriles and 45-72 ng/l for haloacetic acids. PMID- 10536841 TI - Determination of tocopherols and sterols in vegetable oils by solid-phase extraction and subsequent capillary gas chromatographic analysis. AB - In general, analyses of tocopherols and sterols are performed separately in vegetable oils. By applying solid-phase extraction (SPE) prior to capillary gas chromatography, a simple and reliable procedure for the quantification of both tocopherols and sterols in a single analytical run has been developed. SPE was used as sample clean up procedure for the separation of these minor components from the triacylglycerol matrix, replacing time consuming saponification or on line LC-GC. The analysis of tocopherols and free sterols in five different vegetable oils illustrates robustness and reliability of this method outlined. Quantification of the analytes was performed by external calibration with reference substances and internal standardization. The recovery of the procedure as well as the repeatability of the quantitative results have been evaluated. PMID- 10536842 TI - Determination of oak lactones in barrel-aged wines and in oak extracts by stable isotope dilution analysis. AB - The cis- and trans-isomers of 5-butyl-4-methyl-4,5-dihydro-2(3H)-furanone, the so called oak lactones, are derived from oakwood, and the cis-isomer is an important contributor to wine flavour. Their deuterium-labelled forms, [2H4]cis-oak lactone and [2H4]trans-oak lactone, were synthesised from the unlabelled analogues, and were utilised in a new method employing gas chromatography-mass spectrometry to determine the concentration of these compounds in wine or extracts of oak shavings in a single analysis. The method can employ either liquid-liquid extraction or solid-phase microextraction, and is both rapid and accurate. There was some artefactual generation of cis-oak lactone during the analysis of model wine extracts of unheated oak shavings when diethyl ether extraction and injector block temperatures at or above 225 degrees C were employed. PMID- 10536843 TI - Study of applicability of various solid-phase extraction materials for sample handling in screening analysis of organic micropollutants in water. AB - At present, solid-phase extraction (SPE) has become an often preferred preconcentration technique in the screening for a wide range of organic micropollutants in water. A wide choice of materials available on the market makes SPE a suitable tool to cope with an increasing variability of organic compounds entering the hydrosphere. However, the interactions of various sorbent materials with compounds having different physico-chemical properties leads inevitably to large differences in preconcentration efficiency. The aim of this paper was to investigate the efficiency of preconcentration of selected organic compounds from aqueous solutions on various SPE materials. Simultaneously, the potential of newly emerging SPE procedures was compared to results of traditional liquid-liquid extraction methods. The group of 19 tested analytes was selected so as to represent different classes of organic compounds which may occur in waters. The results obtained showed that most of the tested materials were suitable for sufficient preconcentration of a substantial part of the tested analytes. However, specific differences in recovery of one or more analytes were found for almost each sorbent even in the case when the materials had similar composition. This behaviour clearly indicates the need for a thorough testing of capabilities of any SPE material intended for the use in a wide range screening method for the identification of unknown organic micropollutants in water. PMID- 10536844 TI - Retention and separation studies of cholesterol and bile acids using thermostated thin-layer chromatography. AB - The influence of temperature on retention and separation of cholesterol and bile acids, using reversed-phase thin-layer chromatography, was studied. As mobile phases methanol-water mixtures of various compositions were used. Chromatographic experiments were performed using vapor-saturated chambers at temperatures ranging from 5 to 60 degrees C. A linear relationship between R(M) values and temperature (1/T) as well as mobile phase composition was observed. The elution order of steroids under the conditions investigated was discussed. Each chromatogram was evaluated using simple optimization parameters and the best chromatographic conditions for the separation of multicomponent samples were chosen. PMID- 10536845 TI - Studies on the relationship between structure and electrophoretic mobility of alpha-helical and beta-sheet peptides using capillary zone electrophoresis. AB - The electrophoretic behaviour of a series of 33 different synthetic peptides has been investigated using free solution high-performance capillary zonal electrophoretic (HPCZE) methods. The dependency of the electrophoretic mobility, mu(em), on the peptide charge, q, and on the charge-to-size ratio parameter, zeta, determined according to several electromobility models, have been examined. Significant divergences from linearity in the mu(em) vs. q or the mu(em) vs. zeta plots were noted for several peptides, possibly due to the proclivity of specific arrangements of their amino acid sequences to assume preferred alpha-helical or beta-sheet conformational features rather than random coil structures under the HPCZE conditions. These results provide further demonstration of the facility of HPCZE procedures to probe the effects of compositional, sequential and conformational differences of closely-related peptides and their consequences on their physicochemical behaviour in solution. PMID- 10536846 TI - Measurement of electroosmotic flow in plastic imprinted microfluid devices and the effect of protein adsorption on flow rate. AB - Several commercially available plastic materials were used as substrates in the fabrication of microfluid channels for biochemical analysis. Protocols for fabrication using the wire-imprinting method are reported for polystyrene, polymethylmethacrylate and a copolyester material. Channel sealing was accomplished by low-temperature bonding of a substrate of similar material; therefore, each channel was composed of a single material on all sides. The electroosmotic flow in 25-microm imprinted channels was evaluated for each substrate material. The copolyester material exhibited the highest electroosmotic flow mobility of 4.3 x 10(-4) cm2 V(-1) s(-1) which is similar to that previously reported for fused-silica capillaries. Polystyrene exhibited the lowest electroosmotic flow mobility of 1.8 x 10(-4) cm2 V(-1) s(-1). Plots of linear velocity versus applied electric field strength were linear from 100 V cm(-1) to 500 V cm(-1) indicating that heat dissipation is effective for all substrates in this range. Electroosmotic flow was reevaluated in the plastic channels following incubation in antibody solution to access the non-specific binding characteristics of a common biochemical reagent onto the substrate materials. All materials tested showed a high degree of non-specific adsorption of IgG as indicated by a decrease in the electroosmotic flow mobility in post-incubation testing. PMID- 10536847 TI - Chiral selectors from fruit: application of citrus pectins to enantiomer separations in capillary electrophoresis. AB - Pectins were investigated as chiral selective agents in capillary electrophoresis. Successful enantioresolution of antihistaminic and antimalarial compounds, as well as others, was achieved by utilizing potassium polypectate as the chiral selector. Changes in pH, chiral additive concentration and capillary type were studied in relation to chiral resolution. The effect of degree of esterification of pectin materials on chiral recognition was also evaluated. PMID- 10536848 TI - Analysis of beta-N-oxalyl-L-alpha,beta-diaminopropionic acid and homoarginine in Lathyrus sativus by capillary zone electrophoresis. AB - A simple capillary zone electrophoresis (CZE) method has been developed for the simultaneous quantitative determination of beta-N-oxalyl-L-alpha,beta diaminopropionic acid (beta-ODAP) and homoarginine in Lathyrus sativus (LS; grass pea). A new Na2B4O7-Na2SO4 run buffer was used and the pH was 9.20, contents of beta-ODAP and homoarginine in crude extracts of LS plant material were determined with this method, the RSDs of peak areas of beta-ODAP and homoarginine were 2.62% and 3.61%, respectively. It was found that the equilibrium concentration ratio of alpha- and beta-ODAP decreased from 34.5/65.5 to 28.6/71.4 when the pH of the solution increased from pH 3.0 to pH 11.0. PMID- 10536849 TI - Determination of quinolizidine alkaloids in traditional Chinese herbal drugs by nonaqueous capillary electrophoresis. AB - A rapid method for the determination of quinolizidine alkaloids by nonaqueous capillary electrophoresis was developed. A total of 10 alkaloids (matrine, sophocarpine, oxymatrine, oxysophocarpine, sophoridine, cytisine, sophoramine, aloperine, lehmannine and dauricine) could be easily separated within 18 min. A running buffer composed of 50 mM ammonium acetate, 10% tetrahydrofuran and 0.5% acetic acid in methanol was found to be the most suitable for this separation. Five of these alkaloids were selected for further studies. The linear calibration ranges were 2.51-50.1 microg/ml for sophoridine and sophocarpine, 2.71-54.2 microg/ml for matrine, 3.30-65.9 microg/ml for oxymatrine, and 3.10-62.0 microg/ml for oxysophocarpine. The recovery of the five alkaloids was 98.0-101.3% with relative standard deviations from 1.03 to 2.68% (n=5). The limits of detection for all 10 alkaloids were over the range 0.93-2.31 microg/ml. The method was successfully applied to the phytochemical analysis of alkaloid extracts from three commonly used traditional Chinese herbal drugs: Sophora flavescens Ait. (Kushen), S. alopecuroides L. (Kudouzi or Kugancao) and S. tonkinensis Gapnep (Shandougen). PMID- 10536851 TI - Direct determination of small cations in proteinaceous samples using a flow injection-capillary electrophoresis system. AB - A method is described for the direct determination of small inorganic cations in samples containing large amounts of proteins, such as milk or blood plasma. The method is based on electrokinetic injection in a flow injection analysis capillary electrophoresis (CE) system. The selected CE-electrolyte, containing 5 mM 4-aminopyridine and 7 microM cetyltrimethylammonium bromide at pH 4.5, prevents detrimental protein adsorption on the capillary walls. Therefore, no sample pretreatment, except for dilution, is required. Up to 30 repeated injections in one electrophoretic run can be performed, yielding RSD values of the migration time of less than 1 and 2.5% (n=30) for milk and blood plasma samples, respectively. PMID- 10536850 TI - Micellar electrokinetic capillary chromatography of macrolide antibiotics. Separation of tylosin, erythromycin and their related substances. AB - The separation of tylosin by micellar electrokinetic capillary chromatography with a mixed micelle system is described. Good selectivity was obtained with sodium phosphate buffer (80 mM, pH 7.5) containing 20 mM sodium cholate and 7 mM cetyltrimethylammonium bromide (CTAB). This method permits tylosin to be separated from its closely related substances within 15 min. The influences of type of buffer, buffer pH, the concentrations of sodium cholate and CTAB were investigated. The robustness of the method was examined for tylosin by means of a full-fraction factorial design. Quantitative results are presented. Using a similar buffer system (80 mM sodium phosphate, pH 6.0, 20 mM sodium cholate and 5 mM CTAB), separation of erythromycin and its main related substances was also obtained. However, detection sensitivity and resolution are not sufficient for analysis of related substances in erythromycin commercial samples. PMID- 10536852 TI - Liquid chromatographic determination of imazosulfuron in drinking water and in soil using ultraviolet detection. AB - Reversed-phase liquid chromatography (LC) is used to determine a relatively new sulfonylureic herbicide, imazosulfuron, 1-(2-chloroimidazo-[1,2-a] pyridin-3 ylsulfonyl)-3-(4,6-dimethoxy-2-pyrimidinyl)-urea (TH-913), in drinking water and in soil. TH-913 is extracted from water using solid-phase extraction on C18 bonded silica. Soil samples (20 g) are extracted with 300 ml of methanol-water (50:50) and the acidified extracts are transferred onto Sep-Pak C18 and processed as described for water samples. Off-line desorption is done with 20 ml of methanol-water (50:50). The eluate is evaporated to dryness, the residue dissolved in acetonitrile and analysed by LC with UV detection at 238 nm. The recoveries of TH-913 from water were over 95% (at 0.05 microg/l level) and from soil over 90% (at 0.005 mg/kg level). PMID- 10536853 TI - Determination of betaine in Lycium chinense fruits by liquid chromatography electrospray ionization mass spectrometry. AB - A rapid and sensitive high-performance liquid chromatography-electrospray mass spectrometric method has been developed for the determination of betaine in Lycium chinense fruits. Betaine was analyzed on a system consisting of a NH2 stationary phase and a mobile phase of water-acetonitrile (25:75) by isocratic elution for 40 min. Betaine was identified and quantitated by electrospray ionization mass spectrometry with selected ion monitoring of the protonated ion [Betaine+H]+ and clustered ions [nBetaines+H]+. The limit of detection for betaine by this method was ca. 0.2 ng/ml and the relative standard deviations of the assay (intra- and inter-day) were less than 8.1%. PMID- 10536854 TI - Potential for chlorate interference in ion chromatographic determination of total nitrogen in natural waters following alkaline persulfate digestion. AB - Determination of total nitrogen in aqueous samples after thermal potassium peroxydisulfate (persulfate) digestion is a commonly used alternative to the tedious Kjeldahl procedure. When ion chromatography is used to quantify the nitrate formed during digestion, there is a potential for interference from a chlorate peak if the digested sample initially contained chloride in concentrations close to or greater than the concentration of nitrogen. It was determined that this interference can be avoided either by using chromatographic conditions which cleanly resolve the nitrate and chlorate peaks (e.g., the Dionex AG9-HG column) or by using digestion reagent concentrations chosen to maintain a high pH throughout the digestion. The second alternative is not a viable option for investigators using a single digestion for both total nitrogen (TN) and total phosphorus (TP) analysis. PMID- 10536855 TI - Use of ion chromatography for the determination of heavy and transition metals in biochemical samples. AB - A novel, highly sensitive method for simultaneous separation and determination of Cu2+, Ni2+, Zn2+, Cd2+, Co2+, Mn2+ and Pb2+ in biochemical samples was developed and evaluated by ion chromatography. All of these metals were well separated on a bifunctional ion-exchange column by a concentration gradient of oxalic acid and sodium chloride eluents, coupled with spectrophotometric detection after post column derivatization with 2-[(5-bromo-2-pyridyl)azo]-5-diethylaminophenol at 560 nm. The method detection limits (signal-to-noise 3:1) were at microg l(-1) levels. The calibration graphs were linear (r2>0.999) over two-orders of magnitude. This technique was optimized and validated by analyzing five standard biochemical references. PMID- 10536856 TI - Quantitative analysis of manganese, chromium and molybdenum by ion-pair reversed phase high-performance liquid chromatography with pre-column derivatization and UV-visible detection. AB - An ion-pair reversed-phase high-performance liquid chromatographic method with UV visible spectrophotometric detection is proposed for the simultaneous determination of manganese, chromium and molybdenum. By using a C18-bonded silica column, 4-(2-pyridylazo)resorcinol (PAR) chelates of Mn(II), Cr(VI) and Mo(VI) were successfully separated and accurately determined at 480 nm. Tetrabutylammonium bromide (TBAB) was used as the ion-pair reagent. Effects of pH, the buffer system, the concentration of buffer, the color developing time, the concentration of chelating reagent and the ion-pair reagent on the resolution were investigated. PAR chelates were eluted within 20 min at a flow-rate of 1.0 ml min(-1) with a methanol aqueous mobile phase, CH3OH-water (20:80, v/v), containing 1.0 x 10(-3) mol l(-1) acetate buffer (pH 6.5), 1.8 x 10(-2) mol l(-1) TBAB and 2.0 x 10(-4) mol l(-1) PAR. The feasibility of the proposed method was verified with the standard reference materials of nickel-based alloys. The nickel based alloys were analyzed chromatographically after ammonium pretreatment. Under the optimum conditions, the detection limits for the chelates of Mn(II), Cr(VI) and Mo(VI) were 0.31, 4.2 and 4.6 ng with 100 microl injection, respectively. The accuracy of the proposed chromatographic method was verified by good agreement between the values obtained by this method and certified values. PMID- 10536857 TI - Analysis of linear alkylbenzenesulfonates in water samples by large-volume injection-port derivatization and gas chromatography-mass spectrometry. AB - This work presents a modified method to analyze linear alkylbenzenesulfonates (LASs) in water samples. The method involves extraction of samples by a graphitized carbon black (GCB) cartridge, and direct derivatization in the GC injection port using a large-volume (10-20 microl) direct sample introduction (DSI) device with tetraalkylammonium (TAA) salts. The analytes were then identified and quantitated by ion-trap GC-MS. The large-volume DSI injection-port derivatization technique provides sensitivity, fast and reproducible results for LAS residues, to quantitation at 0.1 microg/l in 200 ml of water samples. The retention effect of TAA salts in the injection port was not detected. Enhanced selected mass chromatograms of [M-55]+ ions of butylated C10-C13 LASs by electron impact ionization MS allows one to determine LAS residues at trace levels in environmental samples. Recovery of total LASs in spiked variety water samples ranged from 89 to 112% while RSDs ranged from 2 to 13%. PMID- 10536858 TI - Capillary electrophoretic determination of ferric dimethyldithiocarbamate as iron(III) chelate of EDTA. AB - A simple and sensitive capillary electrophoretic method with UV detection has been developed for the determination of Ferbam (ferric dimethyldithiocarbamate) in boric acid buffer after its acidic decomposition and complexation with EDTA as Fe-EDTA- complex. The determination is dependent on the pH and the nature of the buffer solutions. In this method the detection limit (S/N = 3) is 1.8 x 10(-6) mol/L (0.7 mg/kg) of Ferbam. The relative standard deviation for the analysis of 50 microg/ml was found to be 2.9%. The method was successfully applied for the analysis of wheat grain samples spiked with Ferbam. The applicability of capillary electrophoresis as a useful tool for the analysis of Ferbam is demonstrated. PMID- 10536859 TI - PCNA positivity in the telencephalic matrix areas in the adult of a lizard, Podarcis sicula. AB - In the present paper a reappraisal is made of the persistence and size of telencephalic matrix areas in normal adult specimens of a lizard, Podarcis sicula, using a recent in situ marker of cells in the S phase, the Proliferating Cell Nuclear Antigen (PCNA), belonging to the cyclin family, proteins that regulate the cell cycle and that can be immunocytochemically evidenced using monoclonal antibodies. Only a small number of positive PCNA elements, randomly arranged in the dorsal (medial and lateral) matrix areas, were found in the telencephalic ependymal tissue. These elements were found in larger numbers and distributed more uniformly in the ventral matrix areas, both opposite the ventricular lumen and outside the ependyma. PMID- 10536860 TI - Substance P receptor-like immunoreactive neurons in the caudal spinal trigeminal nucleus send axons to the gelatinosus thalamic nucleus in the rat. AB - By means of substance P receptor (SPR) immunofluorescence histochemistry combined with Fluoro-Gold (FG) fluorescent retrograde labeling, SPR-like immunoreactive neurons in the caudal spinal trigeminal nucleus of the rat were observed to send their axons to the gelatinosus thalamic nucleus with a clear ipsilateral dominance. FG/SPR double-labeled neurons were distributed mainly in the ventral part of lamina I at the rostral level of the caudal spinal trigeminal nucleus. The percentages of FG/SPR-LI neurons in the total number of SPR-LI neurons and FG labeled neurons are 10.5% and 31.1%, respectively. The present results suggest that trigemino-gelatinosus thalamic projection neurons with SPR-LI in the caudal spinal trigeminal nucleus might receive SP-containing, nociceptive primary afferent fibers from the orofacial region and transmit nociception to the gelatinosus thalamic nucleus. PMID- 10536861 TI - Expression and distribution of various antigens of developing microglial cells in the rat telencephalon. AB - The distribution of microglia during the early stages of postnatal development in the rat was studied on rat brain from day of birth to postnatal day 90 (P90), using immunohistochemical methods with a panel of monoclonal antibodies that recognized the complement type 3 receptor (OX-42), macrophage antigen of unknown function (ED1), and the major histocompatibility complex (MHC) class I (OX-18) or class II (OX-6) antigens. Starting from the day of birth, ameboid microglia can be differentiated with positive immunoreactivity to OX-42, OX-18, and ED1. Labeled cells were localized mainly in the developing white matter. After P21, only positive reaction to OX-42 was present, and those cells had the typical morphology of the resting microglial cells that were located either in the white or grey matter. The changes in the appearance of different antigens are correlated with the morphological differentiation and transformation of ameboid microglial cells that are to become ramified microglia, present in the adult animals. PMID- 10536862 TI - Formation of the corticopontine projection in a long-term culture system. AB - The reestablishment in vitro of the corticopontine projection was studied in organotypic co-cultures of cortex and pons of rats 0 (day of birth) - 3 days old. After 2-3 weeks in vitro, application of the lipophilic tracer DiI in the pontine explant retrogradely stained in layer V of the cortical explant pyramidal neurons which were characterized by large somata and spiny dendrites with an apical dendrite that reached upper cortical layers II/III. The projection developed only in co-cultures from rats 0-2 days old when the pontine explant was placed in close vicinity either to the white matter or the pial surface of the cortical explant. Control experiments demonstrated the specificity of the corticopontine projection in vitro by showing that the projections of the layer V pyramidal cells to the pons did not reflect a non-directed outgrowth pattern with subsequent survival of axons contacting the target explant. Our findings demonstrate that morphology and laminar position of corticopontine projection neurons in vitro are similar to those in vivo and thus support the "organotypic" nature of the explant co-culture system. PMID- 10536863 TI - Projections from subnucleus oralis of the spinal trigeminal nucleus to contralateral thalamus via the relay of juxtatrigeminal nucleus and dorsomedial part of the principal sensory trigeminal nucleus in the rat. AB - Following injection of HRP into contralateral thalamus, retrogradely labeled cells were observed in principal sensory trigeminal nucleus (Vp) and an area of juxtatrigeminal nucleus (JX) formerly described by John and Tracey (1987). When PHA-L was delivered to dorsomedial part of the subnucleus oralis (Vodm), PHA-L labeled terminals were seen in dorsomedial part of the Vp (Vpdm) and in the JX region. Comparing the distribution of PHA-L labeled terminal field with that of HRP labeled JX neurons showed that the labeled terminals and neurons were overlapped closely in the JX. The distribution patterns of the labeled terminals and JX neurons were also the same: viewed on the coronal planes caudal-rostrally, both of the labelings began to appear at the levels where the facial nerve root was just broken. Rostrally, at middle levels of the motor trigeminal nucleus (Vmo), the labelings showed their typical view covering dorsal and ventral JX (dJX, vJX). The labelings disappeared at rostral poles of the Vmo and Vp. When injections of PHA-L into the Vodm and HRP into the contralateral thalamus was made in one rat, the contacts between Vodm projecting terminals labeled with PHA L and HRP labeled trigemino-thalamic neurons were seen in the JX and also in the Vpdm. Then, electron microscopic (EM) study was done, injections of kainic acid into the Vodm and HRP into the contralateral thalamus was performed simultaneously. After EM embedding, the JX and Vpdm regions were selected, ultrathin sections were cut and observed with EM. In both areas, axo-somatic and axo-dendritic synapses were seen between degenerated boutons and HRP labeled somata or dendrites. Namely, the Vodm projecting terminals synapsed on trigemino thalamic neurons in the JX and Vpdm. Anyway, axo-dendritic synapses was the main type of observed synapses. Thus, the present work demonstrated 1. the JX containing a group of trigemno-thalamic neurons was a target of special projections froin the Vodm; 2. The Vodm neurons projected to the contralateral thalamus through the relay of JX and Vpdm neurons. PMID- 10536864 TI - The pigmentarchitectonic divisions and neuronal types of the central nucleus and intercalated masses of the human amygdala. AB - The central nucleus of the amygdala is a high level limbic center which controls vegetative functions and is involved in neurodegenerative diseases. In the present study, the central nucleus and intercalated masses of the human amygdala have been investigated with the pigment Nissl technique. The central nucleus of the amygdala consists of a main body surrounded by fiber tracts and accessory islands located dorsal to the main body. In the main body, one can distinguish the medial central nucleus with heavily pigmented neuronal types and the lateral central nucleus, which is composed of three subnuclei: the centro-lateral central nucleus has heavily and sparsely pigmented neuronal types, the apico-lateral central nucleus sparsely pigmented neuronal types and the capsular-lateral central nucleus heavily pigmented neuronal types. Based on the pigmentarchitecture, the accessory islands of the central nucleus are part of the apico-lateral and capsular-lateral central nuclei. Altogether, there are eight neuronal types in the medial central nucleus, while four to six neuronal types are found in the subnuclei of the lateral central nucleus. In the intercalated masses, there are four neuronal types, and the predominating cell types are small and sparsely pigmented. In conclusion, the pigment Nissl stain shows that the lateral central nucleus of the human amygdala has three subnuclei, and the accessory islands are part of two of these subnuclei. Furthermore, numerous neuronal types are identified within the central nucleus and intercalated masses of the amygdala, which may reflect the variability in their neurochemical characteristics. PMID- 10536865 TI - Axon collaterals projection from nucleus reticularis tegmenti pontis onto the cerebellar paramedian lobule in the rabbit: a fluorescent double labelling study. AB - Double labelling method with retrograde transport of fluorescent tracers (Fast Blue; FB and Diamidino Yellow; DY) was employed in the rabbit to investigate whether neurones of the nucleus reticularis tegmenti pontis (NRTP) give off axon collaterals to the cerebellar paramedian lobule (PML) of both sides. Following injections to various regions of the homotopic or heterotopic sublobules of the left (FB) and right (DY) PML cortex, distribution of double labelled neurones within NRTP was analyzed. NRTP of the rabbit consists of a medial principal part (the nucleus papillioformis: PLF) and smaller lateral part (the processus tegmentosus lateralis: PTL). Within PLF three subdivisions are to be distinguished: the dorsomedial part -- zone A, the main part -- zone B and the ventrolateral part -- zone C. The present study in the rabbit indicated collateral projections from neurones in some NRTP regions to the both PML. The cells of origin of these projections were located prominently through the rostrocaudal extent of zone B. Projections from zone A were sparse and those from zone C were absent. Moreover, a weak projection arose mainly from the caudal aspect of PTL. It is concluded that the rostral (e and f) and middle (c and d) sublobules are the main targets for the NRTP-PML branching projections. PMID- 10536866 TI - Small brain, large brain -- a quest for nature's scale-up rules. AB - A scaling model of the gyrencephalic mammalian brain, invoking identical repeating cortical units, whose number and size both increase with increasing brain size, was described previously (Prothero, 1997a,b). Each repeating unit, of microscopic dimensions, is viewed as extending from the pial membrane to the underlying white matter. The model predicts discrete scaling exponents, all integral multiples of 1/9, as a function of brain size, as follows: cortical thickness (1/9), outer cortical surface area (6/9), total (folded) cortical surface area (8/9), cortical volume (9/9), white matter volume (9/9), neuron line count (0/9) and neuron volume density (-3/9). In the present paper extensions to the model give discrete exponents for the scaling of the cross-sectional area of the corpus callosum (6/9), for mean foliar and gyral width (each 1/9), for foliar number (2/9), for mean foliar length (3/9), for mean foliar and gyral perimeter (each 3/9) and for total foliar and total gyral length (each 5/9). The predicted scaling exponent for cross-sectional area of the corpus callosum is in reasonable agreement with the empirical observations. Needed are more precise and more extensive data on cortical folding to stringently test the model predictions relating to cortical folding. On the whole, the model is in good agreement with a diverse body of empirical data bearing on the scaling of the gyrencephalic mammalian brain. PMID- 10536867 TI - Establishment of the FMRFamide-immunoreactive olfacto-retinalis pathway during ontogeny of the rainbow trout (Oncorhynchus mykiss). AB - Migration of neurons is one of the mechanisms establishing normal central nervous system connectivity during ontogeny. Proper timing of axonal sprouting is relevant in the same context. In the present study, we used the immunoreactivity of the tetrapeptide FMRFamide (Phe-Met-Arg-Phe-NH2) to visualize the olfacto retinalis projection during trout ontogeny. It starts to innervate the retina two to four weeks after hatching, in contrast to reports on salmon where it only appears after the fish are imprinted on their natal stream. PMID- 10536868 TI - Cornu Ammonis of the dog: a rudimentary CA2 field is only present in a small part of the dorsal division, and is absent in the ventral division of the cornu Ammonis. AB - The relationship between mossy fiber projections and cytoarchitecture was studied in the cornu Ammonis (CA) of three mongrel dogs and two Japanese white rabbits. Precise comparisons were accomplished by examining adjoining sections that were stained by two methods: 1) the Timm-Danscher method which stains for zinc, which is contained in high concentrations in the mossy fiber terminals, and 2) thionin staining of the Nissl substance, so that the mossy fiber projections and cytoarchitecture could be closely compared in each subdivision of the CA. There was a cytoarchitectonically transitional zone (TZ) between CA3 and CA1 in both dogs and rabbits. The TZ was composed of an intermingling of CA3-like large pyramidal cells and CA1-like small pyramidal cells. Across the dorso-ventral length of the rabbit CA, the mossy fiber projections emitted from the dentate granular cell layer extended as far as the distal border of CA3 toward the TZ but did not enter the TZ or CA1. The TZ of the rabbit was equivalent to CA2 of Lorente de No (1934), that was characterized by CA3-like large pyramidal cells which did not receive mossy fiber inputs. In the dorsal CA of the dog, the mossy fiber projections extended across the CA3/TZ border, as far as the most distal part of the TZ in one dog but only for a short distance in the other two dogs. That part of the TZ which did not receive mossy fiber projections corresponded with CA2. There were no mossy fiber projections to CA1. In the ventral CA of the dog, however, mossy fiber projections were more extensive than in the dorsal CA, extending across CA3/TZ as well as TZ/CA1 borders to enter the proximal part of CA1. These observations indicate that there is a distinct CA2 in the rabbit CA, and that a rudimentary CA2 is present only in a small part of the dorsal CA of the dog, whereas it is absent in the ventral CA. PMID- 10536869 TI - Neuronal connections through the posterior commissure in the frog Rana esculenta. AB - The cobalt-labelling technique was used. After iontophoretic injections of cobaltic-lysine complex into the posterior commissure, fibres and neurons were bilaterally labelled in the posterior thalamic nucleus, three other pretectal nuclei, the optic tectum, the nucleus of the medial longitudinal fasciculus, and in the basal optic nucleus. Unilateral cobalt injections into the terminal areas of the posterior commissure in the mesencephalic tegmentum, labelled neurons in the above-mentioned nuclei ipsilaterally, and some cells contralaterally. The connections through the frog's posterior commissure are very similar to those in mammals. PMID- 10536870 TI - Localization of Barrington's nucleus in the pontine dorsolateral tegmentum of the rabbit. AB - The localization of Barrington's nucleus in the dorsolateral pons of the rabbit and its projections to the sacral spinal cord were examined by using retrograde and anterograde labeling methods combined with immunohistochemistry. After injection of wheat germ agglutinin-horseradish peroxidase (WGA-HRP) or a fluorescence tracer, tetramethylrhodamine-dextran amine (TMR), into the sacral spinal cord segments, a cluster of neurons labeled with WGA-HRP or TMR were seen in the pontine dorsolateral tegmentum. To identify whether the retrogradely labeled neurons were situated within the locus coeruleus, the sections containing TMR-labeled neurons through the pons were incubated with anti-tyrosine hydroxylase (TH) antibody and observed under epifluorescence microscope. It was shown that the cluster of TMR-labeled neurons in the dorsolateral tegmentum were surrounded by TH-positive neurons, but they were negatively immunostained with TH like immunoreactivity. In anterograde experiment, injection of WGA-HRP into the dorsolateral tegmentum resulted in many anterogradely labeled nerve fibers and terminals in the sacral spinal cord, including the sacral parasympathetic nucleus. The present results suggest that the cluster of neurons in the dorsolateral tegmentum of the rabbit may correspond to Barrington's nucleus revealed in the rat and cat, and thus may be involved in micturtion reflex of the rabbit. PMID- 10536871 TI - The central distribution pattern of primary afferent fibers innervating the thigh muscle posterior iliotibialis in the chicken. AB - The anatomical relationship between motoneuron dendrites and primary afferent fibers innervating the same muscle were examined by using a modified cholera toxin tracing method. The cholera toxin B subunit conjugated with latex beads was injected into the posterior iliotibial muscle (PIT) of chicken. Both motoneuron dendrites and primary afferent fibers were intensely labeled. Labeled primary afferent fibers innervating the PIT were mainly located in laminae I (ventral part), V (lateral part), VI, VII (except ventromedial part) and lateral motor column (LMC, lamina IX) of the lumbar spinal cord. Numerous labeled primary afferent fibers were observed in proximity to the motoneuron pool of the PIT located in the dorsolateral region of the LMC; primary afferent fibers were closely apposed to somatic profiles of the PIT motoneurons. PMID- 10536872 TI - Distribution of cytoplasmic phospholipase A2 in the normal rat brain. AB - The calcium-dependent cytoplasmic PLA2 (cPLA2) is an 85-kDa cytosolic enzyme that has been detected in cytosolic fractions from rat brain. With immunocytochemical methods, this cPLA2 is distributed throughout rat brain. Very dense immunostaining is observed in the superior olivary nucleus, periolivary nucleus, facial motor nucleus and dorsal cochlear nucleus in hindbrain whereas light immunostaining is seen in forebrain and midbrain areas. Assays of cPLA2 activity in forebrain, midbrain and hindbrain show the highest specific activity in the hindbrain. The distribution of cPLA2 coincides with that of protein kinase C activity in rat brain. The presence of cPLA2 and PKC in hindbrain suggests that these enzymes play a central role in neurotransmitter release, long-term potentiation and neuritogenesis in this area under normal conditions. PMID- 10536873 TI - Eye muscle nerves nuclear size in a breed of domestic rabbits with floppy ears. AB - In many wild species there is a correlation between the capacity for directional hearing and directional seeing, which is associated with the mobility of the eye balls. Among the breeds of domestic rabbits, there are some with pendulous, floppy external ears (e.g., Englische Widder, EW) that might limit directional hearing. The size of the brain stem nuclei in EWs has been determined and compared to rabbits with upright ears. In EW, the oculomotor nuclei are relatively larger than in the other breeds. Possibly, this indicates a compensation of a loss in directional hearing ability achieved through greater mobility of the eyes. At the same time, a variability of brain composition between the breeds, which is an intraspecific variability, is obvious. PMID- 10536874 TI - Neuronal organization of the optic tectum in the river lamprey, Lampetra japonica: a Golgi study. AB - The neuronal and laminar organization of the optic tectum (OT) in the river lamprey was studied using the rapid Golgi method. Based primarily on the distribution pattern of the dendrites, the OT neurons were divided into vertical, horizontal and stellate neurons. The river lamprey OT shows a laminar structure consisting of eight concentric strata. The stratum ependymale consists of several rows of ependymal cells. The stratum cellulare periventriculare contains one to two rows of vertical neurons. The stratum fibrosum periventriculare is thin and contains a few vertical neurons. The stratum cellulare et fibrosum internum consists of several alternating cellular and fibrous layers: a large variety of vertical and horizontal neurons are distributed in this stratum. The stratum fibrosum centrale consists of compact horizontal fiber bundles, among which a few horizontal neurons are disseminated. In the stratum cellulare et fibrosum externum, numerous fibers run horizontally in a loosely organized plexus; various types of vertical, horizontal and stellate neurons are distributed among these fibers. The stratum opticum is the main terminal area of the optic nerve, and contains stellate and horizontal neurons. The stratum marginale is a thin layer and consists of sparse populations of vertical and horizontal neurons. Besides the above outer to inner laminar structure, the OT is divided into medial (m-OT) and lateral parts (1-OT), based primarily on the distribution pattern of the dendrites. The dendrites of neurons in the m-OT are distributed almost exclusively within the OT. On the other hand, the dendrites of some neurons in the 1-OT extended into the confines of the torus semicircularis (TS), and conversely, the dendrites of some neurons in the TS are distributed in the 1-OT. These findings are discussed in relation to the neuronal and laminar organization of the OT in other lamprey species and to recent hodological studies. PMID- 10536875 TI - Carbonic anhydrase inhibitors. Schiff bases of some aromatic sulfonamides and their metal complexes: towards more selective inhibitors of carbonic anhydrase isozyme IV. AB - Reaction of three aromatic sulfonamides possessing a primary amino group, i.e., sulfanilamide, homosulfanilamide and p-aminoethyl-benzenesulfonamide with heterocyclic and aromatic aldehydes afforded a series of Schiff bases. Metal complexes of some of these Schiff bases with divalent transition ions such as Zn(II), Cu(II), Co(II) and Ni(II) have also been obtained. The new compounds were assayed as inhibitors of three isozymes of carbonic anhydrase (CA). Several of the new compounds showed a modest selectivity for the membrane-bound (bovine) isozyme CA IV (bCA IV) as compared to the cytosolic human isozymes hCA I and II, in contrast to classical inhibitors which generally possess a 17-33 times lower affinity for bCA IV. This greater selectivity toward bCA IV is due mainly to a slightly decreased potency against hCA II relative to classical inhibitors. However, metal complexes of these Schiff bases possessed an increased affinity for hCA II, being less inhibitory against bCA IV. The first type of compounds reported here (i.e., the Schiff bases of aromatic sulfonamides with heterocyclic aldehydes) might thus lead to the development of low molecular weight isozyme specific CA IV inhibitors. The difference in affinity for the three isozymes of the inhibitors reported by us here is tentatively explained on the basis of recent X-ray crystallographic studies of these isozymes and their adducts with substrates/inhibitors. PMID- 10536876 TI - Evaluation of the inhibition of other metalloproteinases by matrix metalloproteinase inhibitors. AB - Two series of compounds synthesized as specific matrix metalloproteinase (MMP) inhibitors have been evaluated for their inhibition of non-MMPs. In a series of substituted succinyl hydroxamic acids, some were found to be significant (IC50 < 1 microM) inhibitors of leucine (microsomal) aminopeptidase, neprilysin (3.4.24.11), and thermolysin. Macrocyclic compounds in which the alpha carbon of the succinyl hydroxamate is linked to the side chain of the P2' amino acid were found to be good inhibitors of aminopeptidase, but not of neprilysin or thermolysin. Compounds of neither series were found to be significant inhibitors of angiotensin converting enzyme or carboxypeptidase A. PMID- 10536877 TI - The absolute configuration of belactin A, a beta-lactone-containing serine carboxypeptidase inhibitor: importance of the beta-lactone structure for serine carboxypeptidase inhibition. AB - The absolute configuration of belactin A, a beta-lactone-containing serine carboxypeptidase inhibitor was studied by a crystal X-ray diffraction analysis and its absolute structure was determined to be (2R,3S)-2-?(3S)-3-[(2-amino-5 chlorophenyl)carboxamido]-1,1-dimethyl-2-o xobutyl?-3-methyl-4-oxooxetane. The importance of the beta-lactone structure for inhibitory activity was found by preparing several derivatives of belactin A. PMID- 10536878 TI - Mixed-type inhibition of pulmonary angiotensin I-converting enzyme by captopril, enalaprilat and ramiprilat. AB - We have compared at the enzymological level pulmonary angiotensin I-converting enzymes (ACE) purified to electrophoretic homogeneity from four mammalians species: pig, rat, monkey and human. Using both substrates hippuryl-histidyl leucine and furylacryloyl-phenylalanyl-glycyl-glycine in steady-state conditions, all the ACEs exhibited Michaelis kinetics with identical Michaelis constants, maximal velocities, optimal pH and optimal activating chloride-concentrations. The apparent inhibitory constant was higher for Captopril than for Enalaprilat and even more so for Ramiprilat irrespective of the origin of ACE and the substrate used. Although these inhibitors have been described as competitive inhibitors, Lineweaver-Burk plots were not in accordance with a simple competitive model; moreover, Dixon plots were rather characteristic of non competitive inhibition. These data emphasize the hypothesis that ACE inhibitors act with mixed-type inhibition, which is consistent with their slow-tight binding to the ACE active center, also with binding of chloride on a critical lysine residue leading to a potential conformational change, and finally with the fact that ACE has two domains, each bearing one catalytic site. On the other hand, as identical kinetic parameters were obtained on the different ACE preparations, results from animal models should allow the extrapolation to humans, in particular for investigations on both renin-angiotensin and kallikrein-kinin systems, and on their inhibition. PMID- 10536880 TI - The role of mIgE during thymus-dependent and thymus-independent immune responses. PMID- 10536879 TI - Carbonic anhydrase inhibitors. Water-soluble, topically effective intraocular pressure lowering agents derived from isonicotinic acid and aromatic/heterocyclic sulfonamides: is the tail more important than the ring? AB - Reaction of twenty aromatic/heterocyclic sulfonamides containing a free amino, imino, hydrazino or hydroxyl group, with isonicotinoyl chloride afforded a series of water-soluble compounds (as hydrochloride or triflate salts). The new derivatives were examined as inhibitors of three carbonic anhydrase (CA) isozymes, CA I, II (cytosolic forms) and IV (membrane-bound form). Efficient inhibition was observed against all three isozymes, but especially against CA II and CA IV (K(I) in the nanomolar range), the two isozymes known to play a critical role in aqueous humor secretion within the ciliary processes of the eye. Some of the most potent inhibitors synthesized were applied as 2% water solutions directly into the eye of normotensive or glaucomatous albino rabbits. Very strong intraocular pressure (IOP) lowering was observed for many of them, and the active drug was detected in eye tissues and fluids. According to others the IOP lowering effect of topically effective antiglaucoma sulfonamides is due to the intrinsic nature of the specific heterocyclic sulfonamide considered, among which the thienothiopyran-2-sulfonamide derivatives represent the best studied case e.g. dorzolamide. In order to prove that the tail (in this case the isonicotinoyl moiety) conferring water solubility to a sulfonamide CA inhibitor is more important than the ring to which the sulfonamido group is grafted a dorzolamide derivative to which the isonicotinoyl moiety was attached was also prepared. This new compound is more water soluble than dorzolamide (as hydrochloride salt), behaves as a strong CA II inhibitor and acts similarly to the parent derivative in lowering IOP in experimental animals. Thus, it seems that the tail conferring water solubility is more important for topical activity as an antiglaucoma drug than the heterocyclic/aromatic ring to which the sulfonamido moiety is attached. PMID- 10536881 TI - Immunotherapy in asthma: an updated systematic review. PMID- 10536882 TI - Asthma and allergy in Albania. AB - BACKGROUND: The risk of allergic disease may be influenced by the degree of "westernization". A survey was conducted to ascertain whether the prevalence of allergy was lower in Albania than elsewhere in Europe, as it has been the most isolated European country. METHODS: The subjects were residents of Tirana aged 20 44 years. A screening questionnaire was completed by 2653 subjects. A more detailed questionnaire was administered to a random sample of 564 respondents, together with skin prick tests and serum IgE assay. RESULTS: The prevalence of wheeze in the last year, and of wheeze without a cold, was lower in Albania than in any country that participated in the European Community Respiratory Health Survey. Nasal allergy and atopy (as indicated by serum specific IgE) were also uncommon in Albania, although serum total IgE concentrations were high. CONCLUSIONS: The findings confirmed the hypothesis of a low prevalence of allergy in Albania. Possible reasons include the recent economic isolation of Albania, the infrequency of smoking by women, the lack of domestic pets, and the high incidence of childhood infection and parasitic infestation. The prevalence of allergy and its potential determinants should be monitored in Albania as that country acquires the characteristics of other parts of Europe. PMID- 10536883 TI - Characterization of a dodecapeptide containing a dominant epitope of Par j 1 and Par o 1, the major allergens of P. judaica and P. officinalis pollen. AB - The pollen of Parietaria, a weed of the Urticaceae family, is a major cause of respiratory allergy in Europe, where the most common species are P. judaica and P. officinalis. Previously, we reported that a beta-galactosidase fusion protein (6a-BG) expressing a 26-bp cDNA fragment (6a cDNA) contained a dominant IgE binding epitope (6a epitope) of the major allergens Par o 1 and Par j 1. The present study aimed to define the amino-acid sequence containing the 6a epitope. We analyzed the reactivity of anti-Par o 1 antibodies affinity purified from allergic patient sera with: 1) a panel of synthetic peptides deduced from the 6a nucleotide sequence using different reading frames 2) glutathione S-transferase (GST) fusion proteins containing selected peptides. The peptide NSARARADSCRI (p102) specifically bound anti-Par o 1 antibodies affinity purified from allergic patient sera or from rabbit anti-Par o 1 antiserum (ELISA). The related peptide NSARAGTSSCRI (p101) reacted to human but not to rabbit, anti-Par o 1 antibodies. GST fusion proteins containing p101 (GST 3.5) or p102 (GST 3.2) extensively inhibited the binding between Par o 1 and IgE or IgG antibodies from an allergic patient serum pool according to a dose-response curve. Percent inhibition of IgE antibodies binding obtained by absorbing a solution (50 microl) of affinity purified antibodies with 5 microg of GST 3.2 or with 1.2 mg of GST 3.5 was 69% and 66%, respectively. In conclusion, the results of the present study indicate that the amino-acid sequences NSARARADSCRI (p102) and NSARAGTSSCRI (p101) contain the dominant epitope of Par o 1 and Par j 1 for human IgE and IgG antibodies indicated as 6a epitope. Moreover, the study shows that the epitope is conserved in recombinant molecules containing these peptides, irrespective of the fused polypeptide (beta-galactosidase or GST). The knowledge of the amino-acid sequence of this dominant epitope is important in therapeutic approaches to the development of allergen-derived haptens. PMID- 10536884 TI - Monthly measurements of indoor allergens and the influence of housing type in a northeastern US city. AB - BACKGROUND: We examined seasonal variation of dust-mite (Der f 1 and Der p 1), cat (Fel d 1), and cockroach (Bla g 1) allergens in Boston, while adjusting for other covariates. Limited data are available on seasonal patterns of indoor allergen concentrations for different geographic regions in the USA. Understanding within-home seasonal variation of allergens is important epidemiologically and clinically. METHODS: From June 1995 to June 1996, dust samples were vacuumed monthly from the bed, bedroom floor, and kitchen of 20 homes. Indoor temperatures were measured monthly and used in calculating relative and absolute humidity. Monthly home characteristics questionnaires were completed by an adult resident of each home. Dust samples were assayed by enzyme-linked immunosorbent assays. RESULTS: Der f 1 and Der p 1 in beds and floors peaked in the autumn months, Fel d 1 peaked in winter and spring, and Bla g 1 was highest in summer. Dust-mite allergen concentrations were 1.9-2.4 times higher in autumn than spring, but the levels in beds were 19-31 times higher in houses than those in apartments. Although Fel d 1 levels in beds were 2.4 times higher in spring than summer, homes with cats had levels 224 times higher than those without cats. Similarly, Bla g 1 levels in kitchens were 2.1 times higher in summer than winter, but apartments had levels five times higher than those of houses. CONCLUSIONS: Sampling season is a source of within-home dust-mite, cat, and cockroach allergen variation in the northeastern USA. However, the influence of housing type and owning a cat far outweighed the seasonal variation of these indoor allergens. PMID- 10536885 TI - Cross-reacting IgE and IgG antibodies to Pityrosporum ovale mannan and other yeasts in atopic dermatitis. AB - Atopic dermatitis (AD) patients often demonstrate positive skin prick test results and serum IgE antibodies to a range of different yeasts. This has been thought to be due to cross-reactivity. In this study, the cross-reactivity of IgE and IgG antibodies between mannan and crude antigens of Pityrosporum ovale, Candida albicans, and Saccharomyces cerevisiae and crude antigens of Cryptococcus albidus and Rhodotorula rubra was examined by RAST and ELISA inhibition with two serum pools of AD patients. We found cross-reacting IgE and IgG antibodies. In the IgE response, the main cross-reacting pattern was the mannan region, although inhibition could be achieved also with crude antigens of C. albicans, S. cerevisiae, and, to some extent, C. albidus. P. ovale was the most potent inhibitor of IgE-binding components, and against it the highest IgE antibody levels were detected in AD serum pools. In contrast, C. albicans was found to be the most important inducer of IgG antibodies, since the IgG level against P. ovale mannan in both AD serum pools was very low. Cross-reacting antibodies were also seen in ELISA inhibition with both crude and mannan antigens, but since the IgG antibody level of P. ovale mannan in AD serum pools was low, further studies are needed to confirm the IgG results. PMID- 10536886 TI - Bronchial hyperresponsiveness and airway wall remodelling induced by exposure to allergen for 9 weeks. AB - Prolonged exposure to allergen has been proposed to be important for the development of bronchial hyperresponsiveness and airway remodelling in asthma. The present study was designed to examine the effect of chronic allergen exposure on bronchial responsiveness, eosinophil infiltration, and airway remodelling. We sensitized brown Norway rats with the occupational allergen trimellitic anhydride (TMA) and exposed the animals to TMA conjugated to rat serum albumin (TMA-RSA) on 5 consecutive days each week for 9 weeks, starting 4 weeks after sensitization. IgE and IgG anti-TMA antibodies in serum and bronchial responsiveness to acetylcholine were evaluated before and at weeks 3, 6, and 9 of allergen exposure. Thickness of the airway wall, airway luminal narrowing, and the number of goblet cells and eosinophils in the airway wall were evaluated with an image analysis system in lungs resected after the last assessment of bronchial responsiveness, at the end of the 9-week allergen exposure. All rats developed IgE and IgG anti-TMA antibodies after sensitization. The levels of antibodies increased with allergen exposure until week 6, and then declined. Bronchial hyperresponsiveness to acetylcholine was induced in allergen-exposed rats without ongoing airway eosinophilia. Bronchial hyperresponsiveness induced by chronic allergen exposure via inhalation was accompanied by significantly increased thickness of smooth muscle and airway narrowing in the small airways, and goblet cell hyperplasia in the large airways. We conclude that chronic exposure to allergen can induce bronchial hyperresponsiveness and airway wall remodelling. Airway wall remodelling may contribute to bronchial hyperresponsiveness. PMID- 10536887 TI - Cytokine profiles of BAL T cells and T-cell clones obtained from human asthmatic airways after local allergen challenge. AB - BACKGROUND: This study assessed the heterogeneity of cytokine expression in asthma before and after local allergen challenge. METHODS: BAL T cells were obtained 10 min or 24 h after local endobronchial allergen challenge in atopic asthmatic subjects. T cells were cloned by direct limiting dilution. mRNA expression was assessed by RT-PCR, and cytokine protein production by ELISA. RESULTS: Unstimulated baseline BAL T cells expressed mRNA for IFN-gamma, IL-13, and TNF-alpha. A minority of samples expressed IL-4 and IL-5, but no IL-3 mRNA was detected. PHA stimulation increased expression of IL-3, IL-4, and IL-5 mRNA in 4/6 samples. IL-13 and GM-CSF mRNA were found in BAL cells after allergen challenge, but expression of IFN-gamma was reduced. Both IL-4 and IL-3 were strongly upregulated after PHA stimulation, while the expression of TNF-alpha and IFN-gamma was reduced, compared to equivalent baseline samples. Seventeen panels of BAL T-cell clones were derived (average cloning efficiency 1/40 T cells). Seven panels survived to 8 weeks for analysis. Clones derived 4 h after saline challenge showed strong mRNA signals for IL-13, IL-4, and IFN-gamma, whereas clones derived 24 h after allergen challenge expressed IL-13, GM-CSF, IL-3, IL-4, and often IL-5 (i.e., closer to the Th2 profile). There was considerable heterogeneity in the patterns of cytokine mRNA and protein production by different clones. CONCLUSIONS: T cells from asthmatic airways produce IL-13, IFN gamma, and TNF-alpha, but after allergen challenge, type 2 cytokines are upregulated. mRNA and protein analysis provide complementary information on airways T-cell cytokine profiles. PMID- 10536888 TI - Major basic protein, but not eosinophil cationic protein or eosinophil protein X, is related to atopy in cystic fibrosis. AB - Increased eosinophil granule proteins have been described in serum and sputum samples of patients with cystic fibrosis (CF). It has been assumed that eosinophil degranulation is enhanced in atopic subjects - as in asthmatics. Since in CF no differences in eosinophil cationic protein (ECP), eosinophil protein X (EPX), and eosinophil peroxidase between atopic and nonatopic subjects have been detected, we investigated whether major basic protein (MBP) is increased in serum and sputum samples derived from atopic (n = 14) compared with nonatopic CF subjects (n = 26). In CF patients, high mean serum (sputum) levels of ECP 29.7 microg/l (2.7 mg/l), EPX 53.7 microg/l (7.9 mg/l), and MBP 984.6 microg/l but low sputum MBP levels (57.4 microg/l) were measured. In addition, in serum and in sputum samples, a significant correlation between MBP and ECP (P<0.03 and P<0.0001, respectively) or EPX (P<0.05 and P<0.0004, respectively) was detected. By subdivision of the patients into allergic and nonallergic subjects, significant differences were found for serum MBP values only(mean 1382.2 microg/l vs. 770.5 microg/l; P<0.0001), but not for ECP or EPX serum levels or for eosinophil proteins in sputum. Although no differences between atopic and nonatopic CF patients in ECP and EPX were found, serum MBP levels were higher in patients sensitized to inhalant allergens than in nonsensitized subjects. These results indicate differential release of eosinophil granule proteins in peripheral blood from eosinophils, and they also indicate that MBP in serum likely is to be a better discriminator of atopy in CF. PMID- 10536889 TI - Prevalence of pollinosis in the Basque Country. AB - BACKGROUND: The prevalence of allergic diseases, mainly pollinosis, has increased within the last decades. Our study aimed to determine the prevalence of sensitization to Poaceae pollen in the Basque Country. This is a region of northern Spain, with an area of 7261 km2 and a population of 2109009 inhabitants. Two different climatic regions may be distinguished in the Basque Country (the Atlantic and the Oceanic). METHODS: A transversal study was carried out on 2216 subjects, aged 10-40 years. A personal interviewwas carried out in order to compile study data, by means of a questionnaire that had been previously validated with a clinical history and an allergy study. RESULTS AND CONCLUSIONS: Our results show the prevalence of pollinosis in the Basque Country to be 10.6% (C.I. 95% 9.35-11.96%) without significant differences between men and women. The prevalence in the Atlantic climate area (9.71%) was lower than in the Oceanic climate area (13.61%). There were no differences between persons living in a rural environment (10.87%) and those living in an urban setting (10.51%). Pollinosis was more frequent in individuals aged 10-20 (11.41%) and 20-30 (12.54%) than those aged 30-40 years (7.43%). Three features significantly distinguished pollinic from nonpollinic patients: 1) a more common complaint of symptoms after ingestion of Rosaceae fruits or nuts (10.2%) 2) a family history of atopy (8.8%) 3) a greater occurrence of bronchial asthmatic symptoms (23%). PMID- 10536890 TI - Glutaraldehyde: an occupational hazard in the hospital setting. AB - BACKGROUND: We report a series of 24 health-care workers with respiratory symptoms suggestive of occupational asthma due to glutaraldehyde exposure. METHODS: The history of asthmatic symptoms was investigated with peak expiratory flow rate (PEFR) monitoring, and in eight of the subjects, the specific bronchial provocation test (SBPT) was applied as reference standard for diagnosis of occupational asthma. Levels of glutaraldehyde were monitored in the challenge chamber during the SBPT. Work environmental levels of glutaraldehyde were measured from air samples collected at least once during the PEFR monitoring of endoscopy and theatre nurses. Specific IgE antibodies to glutaraldehyde were measured with a series of glutaraldehyde modified proteins. RESULTS: In the eight workers who underwent SBPT, the diagnosis of occupational asthma was confirmed by a positive reaction (late and dual reaction in five and in three subjects, respectively). The mean level of glutaraldehyde observed during the challenge tests was 0.075 mg/m3 (range 0.065-0.084 mg/m3). In 13 out of the 16 remaining workers, the serial PEFR monitoring showed a work-related effect. In three workers, there was no physiological confirmation of occupational asthma. Levels of glutaraldehyde from the air samples collected in the workplace were as follows: personal short-term samples (mean 0.208 mg/m3; median 0.14 mg/m3; range 0.06-0.84 mg/m3), personal long-term samples (mean 0.071 mg/m3; median 0.07 mg/m3; range 0.003-0.28 mg/m3). Measurements of specific IgE antibodies to glutaraldehyde-modified proteins were positive in seven patients (29.1%) according to a cutoff value of 0.88% RAST binding. The presence of atopy to common environmental allergens and smoking was not associated with specific IgE positivity (P>0.05; Fisher's exact test). CONCLUSIONS: Our report indicates the importance of glutaraldehyde as an occupational hazard among exposed health-care workers. Intervention in the workplace, training of personnel handling this chemical, and accurate health surveillance may reduce the risk of developing occupational asthma due to glutaraldehyde. PMID- 10536891 TI - Post-marketing surveillance study on the safety of sublingual immunotherapy in pediatric patients. AB - BACKGROUND: Immunotherapy (IT) is the only causal treatment for allergic subjects recognized to be effective and to offer long-lasting efficacy. The noninjective routes, aimed at improving the safety of the treatment, have been validated as effective in adults, but documentation of their safety in children is still poor. The aim of the present survey study was to assess the safety of sublingual immunotherapy in pediatric patients, by evaluating a large population. METHODS: A total of 268 children (aged 2-15 years), receiving sublingual IT for respiratory allergy, were followed-up over a period ranging from 3 months to 7 years (mean 34 months). The side-effects possibly due to the treatment were recorded on a proper diary card; self-assessment of the clinical outcome was also evaluated. RESULTS: About 96000 doses of extract were globally administered. Local side-effects were of no clinical relevance. Eight side-effects were reported (3% of patients; 0.083 per 1000 doses). Seven systemic side-effects (abdominal pain, conjunctival itching, and rhinitis) were mild and required no treatment. One case of urticaria was well controlled with oral antihistamines. No life-threatening event occurred. The clinical outcome was judged excellent or good by 80% of the patients. CONCLUSIONS: The sublingual IT herein investigated appeared to be well tolerated and safe in pediatric patients. The risk/benefit ratio was therefore favorable. PMID- 10536892 TI - Varicella may cause asthma. PMID- 10536893 TI - Acetaminophen-induced rhabdomyolysis. PMID- 10536894 TI - Food allergy to egg and soy lecithins. PMID- 10536895 TI - Anaphylactic reaction to castor bean seeds. PMID- 10536896 TI - Unconventional medicine: a risk of undertreatment of allergic patients. PMID- 10536897 TI - Long-term treatment of asthma with beta2-agonists. PMID- 10536898 TI - Genetic studies on atopy and helminthiasis. PMID- 10536899 TI - Glutaraldehyde-induced asthma. PMID- 10536900 TI - Culture positive tuberculous meningitis: clinical indicators of poor prognosis. AB - Few studies have evaluated culture positive tuberculous meningitis (TBM) as a group. We evaluated certain clinical factors in culture positive TBM which could be associated with a poorer outcome. Out of 40 consecutive TBM patients seen over a period of 4 years in a tertiary referral hospital, 18 culture positive and non human immunodeficiency virus (HIV) related cases were studied. The mean age was 37.9 +/- 14.9 years (range 9-63); five were males and 13 females. None had any associated active chronic medical illness. Patients (44.4%) started on antituberculous treatment within 24 h of admission. Treatment was initiated at a median time of 48 h upon admission in hospital. Univariate analysis revealed a significant correlation between hydrocephalus (P = 0.007) and poor morbidity and mortality. The other clinical factors were not statistically significant: age (P = 0.36): sex (P = 0.49); symptom duration (P = 0.69); BCG vaccination (P = 0.65); cerebral infarct (P = 0.63); extrameningeal spread (P = 1.00); steroids (P = 1.00); time to treatment (P = 0.94) and stage of disease (P = 0.11). Hydrocephalus was the only significant factor predisposing culture positive TBM patients to a poorer outcome. There was also a trend towards a poorer prognosis in those with advanced stage of the disease. PMID- 10536901 TI - Angelman syndrome: a review of clinical and genetic aspects. AB - This paper reviews Angelman syndrome (AS) with regard to the clinical features in childhood and adulthood, epileptic seizures and EEG findings, neuroimaging studies and the present knowledge on the genetic mechanisms underlying this syndrome. Different clinical phenotypes and genotypes of AS are described, including chromosome 15q11-13 deletion, uniparental disomy, methylation imprinting abnormalities and mutations in the UBE3A gene. PMID- 10536902 TI - Excitatory amino acids and taurine levels in cerebrospinal fluid of hypoxic ischemic encephalopathy in newborn. AB - The recent studies indicating the transiently enhanced expression of excitatory amino acid receptors in hypoxia vulnerable brain regions and the elevated concentration of aspartate and glutamate in cerebrospinal fluid of asphyxiated newborns strongly suggest the role of excitatory amino acids in hypoxic ischemic brain damage in the developing human brain. In this study, we compared the concentrations of glutamate, aspartate, taurine and glycine in the cerebrospinal fluid of asphyxiated infants with values of a healthy control group. The concentrations of aspartate (5.82 +/- 3.36), glutamate (1.76 +/- 1.0) and taurine (9.32 +/- 9.1) were significantly elevated in cerebrospinal fluid of asphyxiated infants (P < 0.05). When compared to the control group, the high levels of aspartate was correlated with the degrees of hypoxic-ischemic encephalopathy (HIE) and the varying outcome. The high levels of aspartate and glutamate in the asphyxiated patients adds further evidence to the role of excitotoxicity in hypoxic ischemic encephalopathy. The mental and motor development of the patients in asphyxiated group was followed for 3 years. PMID- 10536903 TI - Penetrating craniocerebral injuries in a civilian population in mid-Europe. AB - Our current neurosurgical understanding of civilian penetrating craniocerebral injuries is based on US metropolitan series. It is unknown whether all principles applied to these patients are relevant in the Mid-European setting with its distinct epidemiology. The objective of this study was to characterize our patients with penetrating craniocerebral injuries, to analyze their outcome, and to identify relevant prognostic factors. Thirty-two patients with penetrating craniocerebral injuries were entered into the study. Patient evaluation comprised neurological, laboratory and radiographic analyses. Motivating factors were suicide (75%), assault (13%), and accident (9%). Initial GCS score, coagulopathy on admission, and radiographic extent of injury could be identified as outcome predictors (P < 0.001). An aggressive therapeutic approach to patients with GCS 3 7 reduced mortality when compared to a conservative management (67 vs. 91%). Due to major differences in epidemiology and outcome of our penetrating craniocerebral injury patients when compared to major US metropolitan series, current therapeutic strategies applied to this patient population in mid-Europe should be reconsidered. The results of our study justify an aggressive neurosurgical approach even in those patients that are thought to have a deleterious prognosis. Predictive variables identified in this study and a novel CT-grading algorithm may help in decision making. PMID- 10536904 TI - Cognitive outcome following staged bilateral pallidal stimulation for the treatment of Parkinson's disease. AB - As neurosurgical treatment of parkinsonian symptoms has become increasingly popular, concern about the cognitive morbidity which may result from such interventions has risen proportionately. Previous reports of cognitive difficulties associated with pallidotomy and thalamotomy, especially in bilateral cases, have provided the impetus for research into chronic electrical deep brain stimulation procedures which are believed to be safer than ablation. Given the lack of neurobehavioral research following bilateral deep brain stimulation procedures, this preliminary study of six Parkinson's disease patients undergoing staged bilateral pallidal stimulation was undertaken. A battery of tests assessing attention, executive function, visuomotor coordination, language, visuoperceptual function, learning memory and mood revealed no significant change in overall level of cognitive functioning after either unilateral or bilateral pallidal deep brain stimulation. No significant declines were observed about three months following bilateral stimulation, and in fact, significant gains in delayed recall and relief of anxiety symptoms were noted. It was concluded from this preliminary data that bilateral pallidal stimulation for the treatment of Parkinson's disease, at least in the absence of operative complications, offers a cognitively safe alternative to ablation. PMID- 10536905 TI - Progression of optic neuritis to multiple sclerosis: an 8-year follow-up study. AB - OBJECTIVE: The relationship between acute monosymptomatic optic neuritis (AON) and subsequent multiple sclerosis (MS) is still doubtful. We investigated the risk of developing MS in patients from North Bavaria, who were suffering from AON. PATIENTS/METHODS: Twenty-nine patients with clinical evidence of AON were included in the study. Initial evaluation included brain resonance imaging (MRI) and a clinical neurological examination. Follow-up examinations were performed after 72-108 months (mean: 96 months) in 26 patients (three patients were lost to follow-up) and consisted of a second complete neurological examination. Diagnosis of MS was established according to the criteria of Poser CM, Paty DW, Scheinberg L. New diagnostic criteria for multiple sclerosis: guidelines for research protocols. Ann Neurol 1983:13:227-231. RESULTS: At follow-up, 14 of 26 patients (54%) had converted to clinically definite MS. Nine patients (64%) had developed MS within 2 years after the onset of AON. CONCLUSION: We observed the development of MS in 54% of the 26 investigated AON patients. The exceptional feature of the present study was the relatively long follow-up period of 8 years. PMID- 10536906 TI - Migraine associated bilateral intracerebral haemorrhages. AB - The authors report a case of bilateral basal ganglionic haemorrhages which occurred during an attack of classical migraine. The patient had a history of migraine associated with aura of neurological deficit for 10 years and a history of arterial hypertension for 20 years, which was treated with propranolol. Intracerebral haemorrhage during an attack of migraine is very rare and up to now the existence of true migraine-induced intracerebral haemorrhage has been controversial. Our case of bilateral occurrence of the haemorrhages supports the theory of the existence of migraine-induced damage of the wall of intraparenchymal vessels during vasoconstriction and focal ischaemia at the beginning of a migraine attack. Subsequent vessel rupture may occur during the following period of increased cerebral blood flow especially with coexisting arterial hypertension. The terminology of the syndrome of migraine associated with intracerebral haemorrhage is reviewed. PMID- 10536907 TI - Ondine's curse in association with diabetes insipidus following transient vertebrobasilar ischemia. AB - Ischemic lesions of the brainstem can lead to complex neurologic deficits. Failure of the automatic control of ventilation (Ondine's curse syndrome) is a possible but rare syndrome following localized brainstem dysfunction. We report on a 49-year-old man with intermittent bradycardia, cranial nerves' dysfunctions and a slight right-sided hemiparesis. An acute brainstem ischemia was diagnosed and treated immediately with high-dose heparin. Cerebral angiography revealed a proximal occlusion of the left vertebral artery but a normal right vertebral artery and a hyperplastic right posterior inferior cerebellar artery. Cranial Computed Tomography and MRI scan demonstrated multiple ischemic lesions in the posterior circulation. During a 4-week treatment course the patient underwent six episodes of acute severe hypoxia and hypercapnia requiring orotracheal intubation twice and manual ventilation by air mask over a few minutes for four times after a tracheostomy had been performed. Twice a short-term episode of hypothalamic Diabetes insipidus was observed following hypoventilation. We conclude that both Ondine's curse syndrome and diabetes insipidus were due to transient vertebrobasilar ischemia. PMID- 10536908 TI - Encephalomyeloradiculoneuropathy following exposure to an industrial solvent. AB - A 19-year-old male developed complaints including weakness of the lower extremities and right hand, numbness, dysphagia and urinary difficulties following a 2 month exposure to an industrial solvent constituted mainly of 1 bromopropane, but also containing butylene oxide, 1,3 dioxolane, nitromethane, and other components. Nerve conduction studies revealed evidence of a primary, symmetric demyelinating polyneuropathy. Evidence of CNS involvement came from gadolinium enhanced MRI scans of the brain, showing patchy areas of increased T2 signal in the periventricular white matter, similar scans of the spinal cord revealing root enhancement at several lumbar levels, and SSEP studies. The patient's symptoms had started to resolve following the discontinuation of the exposure, before he was lost to follow-up. Similar findings have been reported following 1-bromopropane exposure in rats. I hypothesize that this patient's symptoms may have been due to 1-bromopropane-induced neurotoxicity. PMID- 10536909 TI - Intracranial dislocation of a lumbo-peritoneal shunt-catheter: case report and review of the literature. AB - We report on the dislocation of the tip of a lumbo-peritoneal shunting catheter into the cerebral parenchyma 10 months after insertion. The progressive migration towards the deep structures of the brain, once the catheter had left the peritoneal cavity, might have been caused by CSF-flow. Such hypothesis is supported by modern MRI technology visualizing CSF-flow in a spino-cerebral direction. PMID- 10536910 TI - Neuromyotonia, myocloni, sensory neuropathy and cerebellar symptoms in a patient with antibodies to neuronal nucleoproteins (anti-Hu-antibodies). AB - A middle-aged patient presented with subacute muscular stiffness, myocloni of both extremity and facial muscles, gait ataxia and symmetrical distal painful paraesthesias. Electrophysiologically, neuromyotonia was confirmed. High titer anti-Hu antibodies were detected, but no other paraneoplastic antibodies were found. Small-cell lung cancer was diagnosed. Under chemotherapy tumor remission was achieved and, except for minor sensory deficits, neurological symptoms disappeared. This report shows that paraneoplastic syndromes associated with antibodies to neuronal nucleoproteins (anti-Hu antibodies) may be associated with a syndrome including neuromyotonia, sensory neuropathy, cerebellar symptoms and myocloni. PMID- 10536911 TI - A case of adult moyamoya disease showing progressive angiopathy on cerebral angiography. AB - In moyamoya disease, progression of carotid occlusive lesion in an adult patient is very rare. We report a case of adult moyamoya disease with acute angiographical stage progression and hemodynamic deterioration. A 56-year-old female complaining of left motor weakness visited our department. On MRI, infarct lesion was found in the right white matter. On cerebral angiography, occlusive lesions in the bilateral internal carotid arterial siphons and moyamoya vessels were found. The right side was stage V and the left side was stage III. On IMP SPECT, decreased cerebral hemodynamic reserve of the right cerebral hemisphere was found. In this patient, right STA-MCA anastomosis was performed. After operation, she became possible to walk and discharged to home. Five months after operation, good collateral formation and improvement of hemodynamic reserve in the right hemisphere were found. However, on left carotid arteriography, the anterior and middle cerebral arteries were only slightly imaged, and the disease state progressed to stage IV. In addition, decreased blood flow and hemodynamic reserve were appeared in the left hemisphere. PMID- 10536912 TI - Frontal intraosseous cryptic hemangioma presenting with supraorbital neuralgia. AB - Primary intraosseous cranial hemangiomas are rare benign tumors comprising 0.2% of all osseous neoplasms. Symptomatic cranial cryptic hemangiomas are extremely rare. We report the case of a 43-year-old man with a cryptic hemangioma of the superior orbital rim. Radiological investigations revealed it to be an intraosseous cryptic mass which was totally excised and the supraorbital nerve was decompressed, relieving the patient of his symptoms. Histopathology showed features of an intraosseous hemangioma. PMID- 10536913 TI - Natural interferon-beta treatment of relapsing-remitting and secondary progressive multiple sclerosis patients. A two-year study. AB - OBJECTIVES: To evaluate clinical and MRI effects of natural interferon beta treatment in both relapsing-remitting (RR) and secondary-progressive (SP) multiple sclerosis patients. MATERIAL AND METHODS: A double-blind, randomized trial of natural interferon beta (nIFN-beta) in 58 ambulatory patients with RR and 40 with SP multiple sclerosis. Forty-nine patients (29 RR and 20 SP) were treated with intramuscular nIFN-beta6 MIU three times a week for 24 months and 49 control patients were treated with placebo. RESULTS: Primary clinical endpoints were differences in exacerbation rates and proportion of patients remaining exacerbation-free. There were no significant baseline differences between the treated and placebo groups. In the treated RR group a significant reduction in exacerbation rate, an increase in the probability of remaining exacerbation-free, and an improvement in mean EDSS were found at 24 months. MRI activity and total lesion burden were significantly reduced in treated RR patients. In the SP group, nIFN-beta produced a significant reduction in EDSS score, a significant reduction in active lesion number, a marginally significant favourable difference in total lesion burden but no significant effect on the number of gadolinium-enhancing lesions. Side effects were transient and mild in treated patients. CONCLUSIONS: These observations confirm that nIFN-beta is a promising and well-tolerated treatment for either RR or SP MS patients. PMID- 10536915 TI - The deterioration of cardiovascular reflexes in Parkinson's disease. AB - OBJECTIVES: We have investigated the deterioration of cardiovascular reflexes in Parkinson's disease. PATIENTS AND METHODS: In a group of 20 patients with Parkinson's disease (PD) and in a group of 10 age-matched control subjects a battery of cardiovascular reflex (CVR) tests (Valsalva manoeuvre, deep breathing test, handgrip test, orthostatic test) was repeated after 3 years. RESULTS: In PD patients at deep breathing test, at Valsalva manoeuvre and at orthostatic test the heart rate response decreased significantly (P<0.05). The impairment of systolic and diastolic blood pressure response at orthostatic test was significant as well (P < 0.005). At hand grip test no significant changes in heart rate and blood pressure response were found. In control subjects only heart rate response at orthostatic test decreased significantly after 3 years. CONCLUSION: Our results show progression of an impairment of sympathetic and parasympathetic control of the cardiovascular functions in patients with PD over a period of 3 years. PMID- 10536914 TI - APOE and risk of cognitive impairment in multiple sclerosis. AB - OBJECTIVES: The APOE gene polymorphism and the -491 A/T polymorphism in its regulatory region have been associated with an increased risk for developing Alzheimer's disease. We examined these polymorphisms in multiple sclerosis (MS) patients, to determine if a genetic predisposition may explain the risk for developing cognitive decline in MS. MATERIAL AND METHODS: Eighty-nine relapsing remitting and secondary progressive MS patients underwent to a full neuropsychological battery as well as to determination of APOE and -491 A/T polymorphisms. Genetic analysis was also performed in 107 population controls. RESULTS: The APOE polymorphism was not associated with the risk of cognitive impairment in MS patients. The AA genotype of the -491 A/T polymorphism in the APOE regulatory region was more frequent in cognitively impaired than in cognitively preserved MS subjects. CONCLUSION: The AA homozygous state of the 491 A/T polymorphism of the APOE regulatory region is associated with cognitive impairment in patients with MS. PMID- 10536916 TI - Polymorphonuclear leukocyte nitrite content and antioxidant enzymes in Parkinson's disease patients. AB - OBJECTIVE: The present study was undertaken to evaluate the alteration in the peripheral neuronal nitric oxide synthase (NOS) activity in Parkinson's disease patients. Therefore, basal nitrite content in PMNs, platelets and in the plasma of PD and control Indian population were evaluated. MATERIALS AND METHODS: We estimated nitrite, the nitric oxide (NO) metabolite, in neutrophils (PMNs), platelets and in plasma of control and in L-dopa treated Parkinson's disease (PD) patients. We also measured the activity of catalase, superoxide dismutase (SOD) and glutathione peroxidase (GPx) in the PMNs. RESULTS: We observed a significant increase in the basal nitrite content in PMNs of PD patients without any alteration in the plasma and platelets. Thus, the change was specific to PMNs. Catalase activity was significantly less in the PMNs of PD patients, but SOD and GPx remained unaltered. CONCLUSION: Results obtained in the PD patients exhibit an increase in the NOS activity in PMNs. Thus, involvement of NO is suggested in PD. PMID- 10536917 TI - Double filtration plasmapheresis in myasthenia gravis--analysis of clinical efficacy and prognostic parameters. AB - OBJECTIVES: The aim of this study was to evaluate the efficacy of double filtration plasmapheresis (DFP) in the treatment of patients with myasthenia gravis (MG) and to analyze the possible prognostic factors related to responsiveness to DFP. MATERIALS AND METHODS: We treated 45 MG patients, 26 women and 19 men aged 21-72 years, with DFP for 5 consecutive sessions. All were affected by severe generalized or respiratory weakness with an Osserman's classification of group 2 or 3 and had not responded to previous treatments. RESULTS: Thirty-eight out of 45 patients (84%) achieved significant improvements after DFP. The baseline MG score and removal rate for immunoglobulin G (IgG) were significantly higher in the patients with good response than in the other response groups. Poor responders were more likely to have thymoma and a longer interval among sessions of DFP. Better response in patients with age at onset of less than 40 years was associated with higher MG score. Serum concentration of all proteins tested fell as follows (mean +/- SD): IgM, 88+/-7%; IgA, 71+/-11%; IgG, 59+/-14%; globulin, 52+/-11%; AchRAb, 47+/-14%; and albumin, 27+/-10%. All the patients tolerated plasmapheresis well except for 2.2% who experienced hypotension. CONCLUSION: In this study, DFP was effective and safe in the treatment of patients with severe generalized MG. The factors correlating with the better clinical response were high MG score, a thymic pathology of non thymoma, daily apheresis, young age at onset, and high removal rate for IgG. PMID- 10536918 TI - Prolonged P3 latency and decreased brain perfusion in cerebellar degeneration. AB - OBJECTIVES: To clarify the underlying mechanism of cognitive impairments in patients with cerebellar degeneration using event-related potentials (ERPs) and regional brain perfusion measurement. MATERIALS AND METHODS: The P3 latency of ERPs was measured during a visual discrimination task in 15 patients with idiopathic late-onset cerebellar ataxia (ILOCA) and 10 age-matched control subjects. Regional brain perfusion was measured by single photon emission computed tomography using N-isopropyl-p[123I] iodoamphetamine. RESULTS: The mean P3 latency was longer in the patients than in controls. The patients showed lower perfusion in the frontal cortex and in the cerebellum than control subjects. There was a significant correlation between perfusion of the frontal cortex and cerebellum. CONCLUSIONS: The results indicate that patients with ILOCA exhibit slowing of cognitive information processing and suggest that its cognitive slowing may be due to a disruption of neural circuits between the cerebellum and the frontal cortex. PMID- 10536919 TI - Upregulation of Bax protein and increased DNA degradation in ALS spinal cord motor neurons. AB - OBJECTIVES: To investigate if degeneration of motor neurons in amyotrophic lateral sclerosis (ALS) is related to altered levels of the apoptosis regulating proteins Bcl-2 and Bax. In addition, immunoreactivity of the cysteine protease ICH-IL and detection of motor neurons with DNA fragmentation, indicative of apoptosis, was also studied. MATERIAL AND METHODS: The immunoreactivity of Bcl-2, Bax and ICH-1L were compared in ALS and control spinal cord motor neurons by immunohistochemical analysis and motor neurons with DNA fragmentation were identified by the TUNEL-method. RESULTS: The results demonstrate an increased expression of Bax in the ALS material as compared to controls but no change in Bcl-2 and ICH-1L expressions. Moreover, a larger proportion of motor neurons stained positive for TUNEL in ALS spinal cords. CONCLUSION: Present study suggest an upregulation of the cell death promoting protein Bax and increased DNA degradation, indicative of apoptosis, in spinal motor neurons of ALS patients. PMID- 10536920 TI - Neuroacanthocytosis--the variability of presenting symptoms in two siblings. AB - Neuroacanthocytosis is a progressive multisystem disease with a wide range of symptoms. The involuntary movements mainly include chorea and orofaciolingual dyskinesias. The descriptive name of the disease refers to the presence of abnormal erythrocytes in peripheral blood. Two siblings are presented. One young female had dystonia, self-mutilating behaviour, lip biting and eating difficulties. Her brother had repeated generalized epileptic seizures several years before developing choreatic movements and neuropsychiatric symptoms. Both had clinical signs of sensorimotor axonal polyneuropathy. Fresh blood smears in each patient contained between 15 and 20% acanthocytes compared to less than 2% in normal controls. Neuroacanthocytosis must be kept in mind in young adult patients without heredity for Huntington's disease and the diagnosis is easily confirmed when making a fresh blood smear. PMID- 10536921 TI - Methylation of cortical brain proteins from patients with HIV infection. AB - OBJECTIVES: Experimental models of vitamin B12 deficient-neuropathy are characterized by central nervous system protein hypomethylation. The encephalitis/vacuolar myelopathy complicating HIV infection and subacute combined degeneration of the cord due to vitamin B12 deficiency share similar biochemical and pathological abnormalities. Altered central nervous system methylation may be important in the pathogenesis of HIV encephalitis. To test this hypothesis we compared brain protein methylation of HIV-positive, and control, subjects. MATERIALS AND METHODS: Carboxymethyltransferase activity was assayed in postmortem cortical brain samples obtained from 16 control patients (9 males); mean age (59+/-5.1 years, range 21-87 years), 9 HIV-positive patients (7 males, 6 IVDA, 3 homosexual, 4 with HIV encephalitis, mean age 37, range 23-45), and 3 patients with Alzheimer's disease (mean age 78 years). RESULTS: The amount of radiolabelled SAM (S-adenosylmethionine) incorporated into carboxymethyl, and N methylation sites within brain proteins from cortical white matter in vitro was significantly lower (P<0.05) in the HIV+ group vs controls. Carboxymethyltransferase activity was similar in the HIV-infected brains irrespective of the presence or absence of HIV encephalitis. Mean cortical methyl group incorporation was also lower in the Alzheimer's disease group compared to controls. CONCLUSION: The observation of reduced in vitro methylation of brain proteins from patients with HIV infection and Alzheimer's disease suggests that fewer unmethylated sites exist due to relative protein hypermethylation in vivo. The absence of hypomethylation in the brains of patients with HIV encephalitis suggests that hypomethylation is not necessary for the development of HIV encephalitis. PMID- 10536922 TI - Chorea in patients with AIDS. AB - OBJECTIVE: To describe differing etiologies and possible anatomoclinical correlates of choreic movements in a series of AIDS patients. METHODS: We analyzed the clinical records and neuroimaging data of 5 consecutive AIDS patients who developed choreic movements at our center from January, 1994 to December, 1996. RESULTS: There were 2 cases of focal choreic dyskinesias, 1 of right hemichorea, and 2 of generalized chorea. Onset was acute and febrile in 1 case, and subacute in the other 4. In 1 patient the chorea was the AIDS onset symptom; in another choreic movements were the first neurological symptom following AIDS diagnosis; in 2 patients AIDS had a neurological onset other than chorea; and in the fifth patient buccofacial dyskinesias appeared following the development of bacterial encephalitis. CONCLUSION: Chorea was associated with cerebral toxoplasmosis in 2 patients, progressive multifocal leukoencephalopathy in 1, subacute HIV encephalopathy in another, and was probably iatrogenic in the last. Chorea is not unusual in AIDS, however the causes are variable and careful neuroradiological and clinical evaluation is required to identify them. AIDS related disease should be considered in young patients presenting with chorea without a family history of movement disorders. PMID- 10536923 TI - Muscle fiber conduction velocity in situ (MFCV) in denervation, reinnervation and disuse atrophy. AB - OBJECTIVES: Muscle fiber conduction velocity (MFCV) was performed in disuse atrophy, in denervated muscle and during reinnervation as a possible index of muscle atrophy, and to clarify the evolution of the fiber size. MATERIAL AND METHODS: MFCV was performed in 12 patients with complete denervation of biceps brachii muscle and during various stages of reinnervation. Twenty-one patients with disuse quadriceps atrophy were also tested. Invasive MFCV was performed according to the method reported elsewhere (2). RESULTS: MFCV decreased significantly in denervated muscles. Reduction of MFCV was found during the first weeks and was progressive. Peak frequency in histograms decreased and the normal Gaussian distribution was lost. MFCV increased progressively after reinnervation with coexistence of slow and significant increase of faster MFCV. MFCV decreased significantly also during the first weeks after immobilization and improved by rehabilitation therapy. CONCLUSION: MFCV is a reliable method to test the muscle fiber size after denervation and immobilization, and its evolution by reinnervation and therapy. PMID- 10536924 TI - Fronto-temporal dementia and motor neuron disease: a neuropsychological study. AB - The neuropsychological follow-up study of a 58-year-old man suffering from Motor Neuron Disease (ALS/MND) and Fronto-Temporal Dementia (FTD) is reported. Neuromuscular signs first appeared at the age of 51 and slowly progressed to late bulbar involvement; behavioural symptoms of the frontal type first appeared around age 53; lastly, several neuropsychological symptoms suggestive of worsening temporal involvement supervened at age 57. Our patient died at 59 of respiratory failure with the classic clinical and neuroradiological picture of FTD. A short discussion addresses the controversial issue of the coupling of ALS/MND with Dementia and its possible interpretation as the expression of a chance association of relatively common diseases, versus that of a single multifaceted disease. The role of a detailed neuropsychological assessment is highlighted, within the context of increasingly specific diagnostic criteria for FTD. PMID- 10536925 TI - Malignant optic glioma of adulthood. Case report and review of the literature. AB - INTRODUCTION: The malignant optic glioma in adulthood is a rare tumour of middle age which causes an early loss of vision and always leads to death within a year. CASE REPORT: The authors report a case of this disease in a 68-year-old woman with a history of rapidly deteriorating vision and death 6 months after surgery. CONCLUSION: A review of the previous cases showed the accordance of these with the syndrome defined first by Hoyt et al. in 1973; a statistical analysis reveals that the radiotherapy improves the survival whereas the role of chemotherapy is still not definite. PMID- 10536926 TI - Oxidized low density lipoprotein: atherogenic and proinflammatory characteristics during macrophage foam cell formation. An inhibitory role for nutritional antioxidants and serum paraoxonase. AB - Oxidative stress and inflammatory processes are of major importance in atherogenesis because they stimulate oxidized LDL (Ox-LDL)-induced macrophage cholesterol accumulation and foam cell formation, the hallmark of early atherosclerosis. Under oxidative stress, both blood monocytes and plasma lipoproteins invade the arterial wall, where they are exposed to atherogenic modifications. Oxidative stress stimulates endothelial secretion of monocyte chemoattractant protein 1 (MCP-1) and of macrophage colony stimulating factor (M CSF), leading to monocyte adhesion and differentiation, respectively. LDL binds to extracellular matrix (ECM secreted by endothelial cells, smooth muscle cells and macrophages) proteoglycans, in a process that contributes to the enhanced susceptibility of the lipoprotein to oxidation by arterial wall macrophages. ECM retained Ox-LDL is taken up by activated macrophages via their scavenger receptors. This leads to cellular cholesterol accumulation and enhanced atherogenesis. Protection of LDL against oxidation by antioxidants that can act directly on the LDL, or indirectly on the cellular oxidative machinery, or conversion of Ox-LDL to a non-atherogenic particle by HDL-associated paraoxonase (PON-1), can contribute to attenuation of atherosclerosis. PMID- 10536927 TI - Molecular aspects and natural source of procalcitonin. AB - The search for sensitive and specific markers of systemic infection has shown that procalcitonin levels are increased in sepsis, and, consequently, this plasma protein has come into the focus of clinical research. Human procalcitonin is encoded by the Calc-l gene, which gives rise to two alternatively spliced transcripts. Despite systemic investigation of the Calc-l gene and mechanisms of the tissue-specific regulation and alternative splicing, little is known about the biology of procalcitonin and the cells which express this protein during inflammation. Here we focus on the molecular and biochemical properties of the molecule and summarize the known biological functions of procalcitonin. We report on the structure of the Calc-l gene, the amino acid conservation of procalcitonin in different species, and the consensus sequences of the protein with regard to sites relevant for posttranslational modification, spatial distribution, and homologies to other cytokines. We discuss aspects of intracellular location of procalcitonin and demonstrate that it has the characteristics of a secreted protein. PMID- 10536928 TI - Urinary porphyrin excretion in hepatitis C infection. AB - A high prevalence of hepatitis C virus infection in porphyria cutanea tarda in some populations suggests a close link between viral hepatitis and alteration of porphyrin metabolism. Moreover, there is evidence of a role of porphyrinopathies in hepatocarcinogenesis. The aim of our study was to obtain data on the prevalence and patterns of heme metabolism alterations in patients with chronic hepatitis C virus infection. Urinary porphyrin excretion was prospectively studied in 100 consecutive outpatients with chronic hepatitis C infection without signs of photosensitivity, using an ion-pair high-performance liquid chromatography method. Increased total porphyrin excretion was found in 41 patients, with predominant excretion of coproporphyrins (whole study group: mean 146 microg/g creatinine, interquartile range 76-186; normal < 150), in 10 patients excretion exceeded 300 microg/g creatinine. In the majority of all patients studied (75/100) an increased ratio of the relatively hydrophobic coproporphyrin isomer I to isomer III was found. In just one case, urinary porphyrin pattern characteristic for chronic hepatic porphyria was present (uroporphyrin > coproporphyrin, heptacarboxyporphyrin III increased) but the total porphyrin excretion was only slightly elevated in this case. In the whole group, total urinary porphyrin excretion correlated well with serum bilirubin and was inversely correlated with albumin and thrombin time. In conclusion, secondary coproporphyrinuria occurs frequently in heptatitis C infection, whereas in Germany, preclinical porphyria cutanea tarda seems to be rare in these patients. PMID- 10536929 TI - Quantification of low abundance natriuretic peptide receptor mRNA in rat tissues. AB - Natriuretic peptides are important regulators of vascular resistance and volume and electrolyte homeostasis. The quantification of natriuretic peptide receptor (NPR) mRNA is important for the understanding of the regulation of this humoral system, but is difficult due to low expression of the NPR mRNA. We report here on the evaluation of a polymerase chain reaction (PCR)-aided transcript titration assay for quantification of all three NPR subtypes (NPR-A, NPR-B, and NPR-C) mRNA. A multispecific internal standard RNA with parts of NPR-A, NPR-B, NPR-C and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) nucleotide sequences was constructed and reverse transcription of standard and sample RNA (400 ng) was performed in parallel for all three NPRs and GAPDH. The specific PCR yielded differently sized products, which were quantified by high performance liquid chromatography (HPLC). The determination of specific mRNA concentrations was not influenced by cDNA input and did not depend on the PCR cycle number. Linearity between sample RNA input and mRNA concentration was demonstrated. Application of the evaluated method showed that the NPR-A mRNA expression was the most abundant of the three natriuretic peptide receptor mRNAs in rat lungs, glomeruli and left ventricles, followed by the NPR-C mRNA and the NPR-B mRNA expression. Thus, the described method allows the reliable quantification of the specific mRNA expression of all three NPRs with small amounts of RNA. The presented method might foster future research on the regulation of this humoral system in cardiovascular and kidney diseases. PMID- 10536930 TI - Rat tissue collagen modified by advanced glycation: correlation with duration of diabetes and glycemic control. AB - Collagenous proteins are especially prone to nonenzymatic glycation, because they contain several dibasic amino acid residues with free amino groups, have a very slow turnover rate, and are exposed to ambient levels of glucose. The aim of this study was to determine the time-dependent course of advanced glycation process in diabetic rats in relation to glycemic control and duration of diabetes, compared to age-matched controls. Immunochemical assay with antibodies to advanced glycation end products (AGE) was first developed to qualitatively detect and quantify the AGE formed in rat tendon and aortic collagen. Individual collagen samples were extracted by extensive pepsin and collagenase digestion. The amount of AGE was measured by competitive ELISA and results were expressed as AGE U/mg collagen. Diabetic rats showed a significant increase in AGE content in aortic collagen at 20 weeks (n = 6, 206.6 +/- 16.7 U/mg collagen) compared with that measured at 4 and 12 weeks (n = 6, 110 +/- 12.8 U/mg collagen, and n = 13, 184.9 +/- 12.3 U/mg collagen at 4 and 12 weeks, respectively; p < 0.001 between 20 weeks and 4 weeks; p < 0.01 between 20 weeks and 12 weeks). The amount of AGE in tendon collagen of diabetic rats increased from 1.9 +/- 0.38 U/mg at 4 weeks to 11.2 +/- 6.1 U/mg collagen at 20 weeks, p < 0.001. Considerable disparity was observed in the intensity of glycation between aortic and tendon collagen. AGE content per mg of aortic collagen was several-fold to that found in tendon collagen (p < 0.001). To investigate the effect of glycemic control on the advanced glycation process, total aortic AGE-collagen content was compared between untreated diabetic rats (D; n = 13, 184.9 +/- 12.3 U/mg) and diabetic rats treated for 12 weeks with insulin (DI; n = 6, 133.9 +/- 10.7 U/mg), or phlorizin (DP; n = 6, 132.4 +/- 8.9 U/mg), or by a combination of insulin/phlorizin (DIP; n = 6, 124.3 +/- 6.5 U/mg). In spite of therapy used, all groups of diabetic animals had a significantly higher aortic AGE-collagen content than those in the nondiabetic control group (C: n = 8, 104.6 +/- 14.9 U/mg) of the same age (D, DI, DP, DIP vs. C, p < 0.001). Comparison between the mean levels of glycated hemoglobin (D: 5.62 +/- 0.38 % vs. C: 1.7 +/- 0.05%) and mean AGE levels in the studied group of animals yielded a very good exponential correlation (r = 0.89, p < 0.001). Glycation-derived late-stage collagen modification was detected by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and by immunoblotting confirmed to contain (an) AGE structure(s). Our study provides strong immunochemical evidence of AGE formation in vivo during hyperglycemia, and of their temporal association with structural alterations of extracellular matrix proteins. The advanced glycation process is retarded and reduced in intensity, but not completely abolished, by glycemia regulation with, or independently of, insulin. PMID- 10536931 TI - Reference values for a homogeneous Ferritin assay and traceability to the 3rd International Recombinant Standard for Ferritin (NIBSC code 94/572). AB - Reference values for two ferritin assays (Tina-quanta Ferritin, Enzymun, both Roche Diagnostics, Mannheim, Germany) were established (136 males and 139 females). To rule out inflammation as well as iron deficiency in the reference population, subjects with the C-reactive protein concentration < 5 mg/l, and zinc protoporphyrin < 40 micromol/mol heme and the soluble transferrin receptor < 3 mg/l were selected. Taking into account latent iron deficiency as well as hereditary hemochromatosis the 5-95 percentile range was as follows: male, 27-365 microg/l; female, 13-148 microg/l for Tina-quanta and 12-151 microg/l for Enzymun. The Tina-quanta Ferritin assay showed a very good correlation (r > or = 0.990) to Enzymun ferritin, Ferritin Abbott (Abbott Diagnostics, Delkenheim, Germany), N Latex Ferritin (Dade Behring, Marburg, Germany) and the Ferritin Chiron (Chiron Diagnostics, Fernwald, Germany). However, the slopes of the standard principal component method analysis were calculated to be between 1.03 (Enzymun) and 1.41 (N Latex Ferritin). For four assays the median recovery of the 3rd International Recombinant Ferritin Standard (NIBSC 94/572) measured by serial dilution was 89-109%. The N Latex Ferritin assay recovered half of the target values. Because of the good correlation with other assays, a matrix effect is likely. The question arises whether the manufacturers' agreement on the recombinant ferritin standard would harmonize ferritin measurement. PMID- 10536932 TI - Iron saturation of ferritin in the course of phlebotomy treatment in patients with haemochromatosis. AB - This paper describes the iron saturation of ferritin in haemochromatosis patients during phlebotomy therapy. The iron saturation of ferritin does not change during therapy and cannot be used as a parameter to follow therapy. Furthermore, the iron saturation seems to be a constant characteristic of a given person. It does not vary with the body iron stores in patients with haemochromatosis. PMID- 10536933 TI - Neural networks for the biochemical prediction of bone mass loss. AB - Neural networks are specialized artificial intelligence techniques that have shown high efficiency in dealing with complex problems. Paradigms such as backpropagation have been successfully applied in a number of biomedical applications, but not in attempts to identify women at risk of postmenopausal osteoporotic complications. In this paper, several neural networks were trained using different combinations of biochemical variables as inputs. Bone densitometric measurements in Ward's triangle and in the spinal column were used as separate classification criteria (outputs) between slow and fast bone mass losers. The most parsimonious model with the best performance included plasma concentrations of estrone, estradiol, osteocalcin, parathyrin and urine concentrations of calcium and hydroxyproline (expressed as ratio to creatinine excretion) as input neurons; ten neurons in a single hidden layer; and one neuron in the output layer. Diagnostic efficiency was 76% in Ward's triangle and 74% in the spinal column; sensitivity was 70 and 81%, and specificity was 77 and 65%, respectively. Linear discriminant analysis showed a diagnostic efficiency of 66% in Ward's triangle and 64% in the spinal column, sensitivity was 55 and 86%, and specificity was 75 and 13%, respectively. We conclude that performance of the stepwise discriminant analysis was not superior to the neural networks. PMID- 10536934 TI - Serum dipeptidyl peptidase IV activity correlates with the T-cell CD26 antigen. PMID- 10536935 TI - The Baltic Atherosclerosis Journal. PMID- 10536936 TI - Forging a link. PMID- 10536938 TI - Is radical lymphadenectomy a valid oncologic procedure? PMID- 10536937 TI - Surgical education in changing times. PMID- 10536939 TI - Pulmonary metastases: biologic and historical justification for VATS. Video assisted thoracic surgery. PMID- 10536940 TI - Lymph node sampling in lung cancer: how should it be done? AB - OBJECTIVES: Systematic lymph node dissection in radical operation for lung cancer is recognized as an operative procedure which is expected to improve local control. We investigate the most effective method of lymph node dissection or sampling. METHODS: A retrospectrive study was carried out on 1815 patients who underwent systematic lymph node dissection and complete resection. The lymphatic route of metastatis from each lobe was investigated by examining which nodes had the most likelihood of metastasis, or to find out which is the sentinel lymph node in the case of small sized tumor, suitable for the video assisted thoracic surgery (VATS) approach. RESULTS: At N2 level, distribution of major metastases from each lobe are as follows: right upper lobe tumor, #3 - 12.3% (80/648) and/or #4 - 8% (52/648); right middle lobe tumor, #3 and/or #7 - 16.4% (13/79); right lower lobe tumor, #7 - 13.7% (52/380); left upper lobe tumor, #5 - 12.3% (60/489) and/or #6 - 6.7% (33/489); and left lower lobe tumor, #7 - 11.9% (26/219). Small sized tumor requires lymph node sampling upon staging, and the lymph node most likely to become the first metastasis, i.e. sentinel node, are as follows: regardless of the location of tumor, #12, #11, and/or #10 in N1 level, which means dissection or sampling within these locations of lymph nodes are prerequisite. In N2 level, #3 and/or #4 in right upper lobe tumor, #3 and/or #7 in right middle lobe tumor, #7 in right lower lobe tumor, #5 and/or #6 in left upper lobe tumor, and, #7 in left lower lobe tumor. CONCLUSIONS: In clinical T1NO lung cancer, sentinel lymph node sampling should be done first, if the nodes are negative, complete mediastinal lymph node dissection might be omitted. On the other hand, if the sentinel nodes are positive for pathology, complete medistinal lymph node dissection is required for curative resection. PMID- 10536941 TI - F-18 fluorodeoxyglucose positron emission tomography in the non-invasive staging of non-small cell lung cancer. AB - OBJECTIVE: Positron emission tomography (PET) using F-18 fluorodeoxyglucose (FDG), a glucose analogue, as a metabolic tumour marker, has been proposed for the non-invasive staging of oncological disease. Tumours demonstrate increased glycolytic activity and thereby, FDG PET can differentiate benign from malignant lesions. To determine its role in the mediastinal staging of patients with suspected non-small cell lung cancer, a prospective study of FDG PET and computed tomography (CT) compared to surgery and pathology was performed. The analysis group consists of 50 patients, 37 men and 13 women, mean age 64 years (range, 41 78 years). METHODS: A nuclear physician, blind to the clinical and CT data, graded the FDG PET studies qualitatively on a five-point scale, based on the intensity of glucose uptake, for the presence of mediastinal nodal tumour involvement. Scores of four or greater were considered positive for tumour. An experienced radiologist interpreted the patients' CT scans blind to the other data. The CT criterion for tumour involvement was a nodal long axis diameter of 10 mm or greater. All patients underwent either thoracotomy or mediastinoscopy to obtain surgical specimens. The PET, CT, surgery and pathology were mapped according to the American Thoracic Society nodal classification resulting in 201 nodal stations evaluated. The imaging studies were analysed for N2 or N3 tumour involvement compared to histology or dissection of nodal stations. RESULTS: All patients had proven non-small cell lung carcinoma. PET excluded tumour in 175 of 181 nodal stations (specificity 97%) compared to 162 of 181 (specificity 90%) by CT. PET correctly identified 16 of 20 (sensitivity 80%) nodal stations with tumour compared to 13 of 20 by CT (sensitivity 65%). Overall, PET correctly staged 191 of 201 nodal stations (accuracy 95%) compared to 175 of 201 by CT (accuracy 87%). By the McNemar test, PET was significantly more specific than CT in excluding nodal tumour involvement (X2 = 5.5, P < 0.05). CONCLUSIONS: FDG PET is more specific than computed tomography in the non-invasive mediastinal staging of non-small cell lung cancer and has an important clinical role in the pre operative staging of lung cancer patients. PMID- 10536942 TI - The role of thoracoscopic staging of esophageal cancer patients. AB - INTRODUCTION: This study was designed to compare thoracoscopy/laparoscopy (TS/LS) staging with non-invasive clinical staging by CT and EUS for patients with esophageal carcinoma. METHODS AND RESULTS: CT and EUS followed by TS/LS were used to stage 88 patients with EGD proven esophageal carcinoma. Thoracoscopic staging was done in 82 patients and found N1 in 11 patients. Fifty-four patients had laparoscopy which detected N1 in 21 patients. Thirty-four cases had chemoradiation followed by surgery. Esophagectomy was performed in 47 patients after thoracoscopic staging and 33 with laparoscopic staging. Of these 47 resected patients, thoracoscopic staging showed N0 in 42 patients and N1 in five patients with an accuracy of 93.6%. Laparoscopic staging detected normal celiac lymph nodes in 20 patients and diseased LN in 11 patients with an accuracy of 93.9%. Comparing with final resection pathology, the sensitivity, specificity and positive predictive value of staging for N1 disease in the chest was 62.5, 100.0 and 100.0% by TS; 75.0, 75.6, and 23.1% by CT and 0.0, 51.4 and 5.5% by EUS, respectively. For N1 disease in the abdomen it was 84.6, 100.0 and 100.0% by Ls; 0.0, 97.1 and 0.0% by CT and 22.2, 81.5 and 28.6% by EUS, respectively. CONCLUSION: TS/LS staging of esophageal cancer patients with or without preoperative chemoradiation has a higher specificity and accuracy than CT and EUS, especially for N1 disease in the chest. It also allows individualization of preoperative radiotherapy fields. PMID- 10536943 TI - Comparison of thoracoscopic and laproscopic esophagomyotomy with fundoplication for primary motility disorders. AB - OBJECTIVES: With the introduction of videoscopic techniques, controversy has arisen whether a thoracoscopic or laproscopic approach is indicated for the surgical management of symptomatic primary motility disorders. The aim of this study was to compare the outcomes of the two techniques performed by one group. METHODS: Between 1995 and 1997, 78 patients (42 female, 36 males: ages 21-86; mean 53 years) underwent a videoscopic esophagomyotomy with fundoplication via a thoracic (12) or abdominal (66) approach for dysphagia or chest pain. Pre operative evaluation with esophagogastroscopy and manometry revealed a primary motility disorder in 64 and primary motility disorder with stricture in 14. Primary motility disorders exhibited were hypertensive LES (25), nutcracker (26), achalasia (14), and diffuse esophageal spasm (13). Associated fundoplications to prevent reflux included abdominal Toupet partial fundoplicatio (52), abdominal Nissen (14) and thoracic Belsey (12). Significance of variation in outcomes was determined by Mann-Whitney U-test. RESULTS: There was no mortality. Follow-up ranged from 6-40 months (mean = 18). Early morbidity included dyshagia--chest pain greater than 6 weeks in 16 patients. (5 Belsey 41%, 10 Toupet 19%, 1 Nissen 7%) Late morbidity included three recurrent strictures requiring dilatation (Belsey 2/5, Toupet 1/7). Two patients (3.1%) experienced a recurrent motility disorder after abdominal short myotomy--Toupet. Five patients experienced postoperative gastroesophageal reflux after partial fundoplication (two Belsey = 16.6%, three Toupet = 5.7%). Overall 63 patients (81%) were completely relieved of dysphagia--chest pain. CONCLUSIONS: Thoracoscopic esophagomyotomy with Belsey fundoplication was associated with a significantly higher incidence of post operative dysphagia--chest pain (P = 0.05) and recurrent stricture (P = 0.01 ) than laproscopic esophagomyotomy with partial or total fundoplication, however, there was no significant difference in the incidence of recurrent motility disorders (P = 0.54) or gastroesophageal reflux disease (P = 0.12) between the techniques. Our results support utilization of a laproscopic approach for primary motility disorders. PMID- 10536944 TI - Staged axillary thoracotomy for bilateral lung metastases: an effective and minimally invasive approach. AB - OBJECTIVE: We describe our experience with the staged axillary thoracotomy (SAT), for the treatment of bilateral lung metastases. MATERIALS AND METHODS: Between January 1995 and June 1998, 75 lung metastasectomies were carried out in our institution, 49 (65%) monolateral, and 26 (35%) bilateral. In the latter group of patients we adopted a staged axillary thoracotomy. RESULTS: All wedge resections and two lobectomies (1 LUL and 1 RLL) were performed through this approach. Resection has been complete in all patients. Histology was epithelial in 15 (57%), sarcoma in nine (35%) and germ cell in two (8%). Two to three metastases have been resected in 10 patients (38%); four to 10 in 12 patients (46%) and over 10 in four patients (15%). The radiological pre-operative assessment was accurate in 15 patients (57%), underestimated in nine (35%) and overestimated in two (8%). The average interval between the two procedures has been 24 +/- 6 days. The average operation duration time was 50 min (range 36-67). We do not report any post-operative death or major complication. The average hospitalization was 3.2 days (range 2-6) for each single procedure and 6.2 days (range 4-10) for both procedures. CONCLUSION: This technique is adequate, fast and safe and did not affect the shoulder girdle motion at all providing an excellent cosmetic outcome. The operative trauma is limited and a minor post-operative pain is present. A shortening of the interval between the two operations is allowed. PMID- 10536945 TI - Emphysema surgery--loop ligation approach. AB - OBJECTIVES: To demonstrate the efficacy of using thoracoscopic endoloop ligation of bullae in patients with bullous emphysema. METHODS: From 1992 to 1997, 93 advanced age (mean age, 66 years) and oxygen dependency patients underwent thoracoscopic procedure using endoloop ligation for treatment of bullous emphysema. Clinical data were collected from chart review. Thoracoscopic loop ligation of bulla was carried out under general anesthesia with double lumen endotracheal tube and single lung ventilation. RESULTS: Eighty-two patients (88%) exhibited subjective improvement in their symptom status at 3-month follow-up (from grade 2 or 3 to grade 1 or 2) according to the modified Medical Research Council dyspnea scale. The mean duration of chest drainage was 7.5 days (range, 4 19 days). Average hospital stay was 9.5 (range, 5-26) days. There was no post operative death. A comparison of pre-operative and post-operative functional evaluation was available in 27 patients who showed an average increase in FEV1 (from 0.89 to 1.12 l) and declined in residual volume after operation. Complications include persistent airleak over 10 days in nine patients (9.7%), wound infection in three patients and localized empyema in five patients. There was no recurrent after a mean follow-up of 37 months. CONCLUSION: Thoracoscopic loop ligation of bulla has proven to be a safe, reliable and cost effective means of technique for bullous emphysema. PMID- 10536946 TI - Is there a role for radical esophagectomy. AB - The aim of primary surgery in the treatment of carcinoma of the esophagus and gastroesophageal junction (GEJ) is definite cure. To obtain this goal R0 resection, i.e. complete macroscopic and microscopic removal is of paramount importance. However, one of the most controversial questions remains the extent of lymph node dissection, in particular the value of cervical lymph node dissection (the so called third field). Three arguments are believed to favour more extended lymphadenectomy: optimal staging, prolonged tumour control, improved cure rate. (a) Optimal staging: available data indicate that unforeseen lymph node involvement in the neck is encountered in approximately 30% of the patients after 3-field lymphadenectomy. Even in tumours of the GEJ up to 20% of the patients in the T3N+ setting have unforeseen positive nodes in the neck. (b) Prolonged tumour control: radical esophagectomy and extensive lymphadenectomy is decreasing locoregional recurrence substantially, below 10%, in several published reports. More over extended lymphadenectomy seems to defer onset of locoregional recurrence and generalised metastasis for up to 3 years or more. (c) Improved cure rate: despite a lack of prospective randomised study many studies indicate a distinct survival benefit after radical esophagectomy and extensive lymphadenectomy. From the available data it becomes clear that radical surgery and extensive lymphadenectomy offers the best chances for prolonged survival or cure. This can be done without increasing hospital mortality and morbidity. Survival figures obtained by this technique are a gold standard to which survival obtained by other techniques (e.g. multimodality treatment forms, VATS resections) have to be compared. PMID- 10536947 TI - Extended resections for lung cancer. How far is too far? AB - Pulmonary resection is the preferred treatment for patients with lung cancer. Half of all patients, however, have signs of unresectability at the time of diagnosis. Contraindication to pulmonary resection is based on cell type, the extent of the disease, and the patient's overall general medical condition. Invasion of the chest wall by bronchogenic carcinoma is not rare. The diagnostic importance of this findings, however, has been controversial. Early reports had regarded thoracic wall invasion as a uniformly ominous sign, while recent reports have been more optimistic, especially when lymph nodes were not metastatically involved. Chest wall resection should be preceeded by mediastinoscopy. If lymph nodes are negative, excision is generally performed en bloc with pulmonary resection. After the thorax is entered and the cancer is found to be invading the chest wall, wide resection of the chest wall with attached lung is performed. Generally, the line of resection should encompass the area of invasion by several centimeters. The lung with attached chest wall is then allowed to fall back into the pleural cavity, where the appropriate pulmonary resection is performed. If the chest wall defect is less than 5 cm in diameter, no reconstruction of the defect is required. If, however the defect is larger and structural, stability is required. The defect should be reconstructed with a prosthetic material, such as the various meshes, metals, or soft tissue patches, and reinforced with a muscle flap. If the wound is contaminated from an intrathoracic source, prosthetic material should be avoided and reconstruction with a muscle flap alone is preferred. Muscles commonly used include serratus anterior, pectoralis major, latissimus dorsi, and occasionaly, rectus abdominus. Because the omentum lacks structural stabilitly, it should be considered a back-up alternative procedure. Operative mortality is usually related to the extent of pulmonary resection rather than the extent of chest wall resection. Five-year survival approaches 50% for patients with T3N0M0 lesions. For patients with either N1 or N2 neoplasms, 5 year survial is less than 10%. Postoperative radiation therapy appears to have no effect on surival. PMID- 10536948 TI - The preoperative selection of patients for emphysema surgery. AB - Lung volume reduction surgery for emphysema is evolving rapidly since its re introduction in 1993. Lung transplantation remains a viable option for others with emphysema. The major difficulty facing surgeons lies in appropriate selection of patients for either procedure. The following paper represents an attempt by review of the literature and personal experience to describe some of the important features involved in patient selection. The current literature on patient selection for lung volume reduction surgery and transplantation for emphysema was reviewed, and the results within the University of Toronto Lung Volume Reduction Program were analyzed. The review suggests that the most reliable predictors of success are heterogeneous distribution of emphysematous change as reflected by the CAT scan and the quantitative ventilation perfusion scan with new emphasis being placed on the ventilation portion of the latter. Poor prognostic indicators are hypercarbia and pulmonary hypertension. It was felt that an algorithm could be established for determination of whether lung volume reduction or transplantation should be offered to patients for emphysema surgery. The algorithm is described. PMID- 10536949 TI - Surgical strategy for lung volume reduction surgery. AB - Lung volume reduction surgery (LVRS) has been a popular procedure since the early 1990s. It appears that there has developed a consensus in the literature that the ideal patient is one with evidence of marked hyperinflation and heterogenous disease. In this patient profile, LVRS has produced excellent results with respect to lung function and improved exercise tolerance. General areas of controversy are discussed which include the role of lasers; unilateral versus bilateral procedures; the role of a staged unilateral procedure; and which surgical route is best for patients. The existing literature is reviewed on these issues. PMID- 10536950 TI - Sublobar resection for lung cancer. AB - The role of limited lung resection 'segmentectomy and wedge resection' in the treatment of lung cancer has been reviewed. Survival for patients with stage I lung cancer and lesions less than 2 cm is comparable to that of major resections such as lobectomy. The theoretical advantage of limited resection is the simplicity of the procedure and the potential for performing it through lesser invasive techniques. The major drawback at this time which should render it a compromise rather than a choice operation is the increased risk of locoregional recurrence. Until properly conducted clinical trials validate its efficacy in peripheral T1 lung cancer with or without adjuvent therapy, sublobar resection should be limited to patients that are at poor risk of tolerating major lung resection. Sublobar resections however may also play a useful role in treatment of metachronous or synchronous lung cancer. PMID- 10536951 TI - Minimally invasive valve surgery: trends for the future. PMID- 10536952 TI - Complete revascularization on cardiopulmonary bypass: a closer look at existing technology. PMID- 10536953 TI - Current results in off pump surgery. AB - OBJECTIVE: We reviewed our experience with myocardial revascularization without cardiopulmonary by-pass (CPB) to evaluate early- and mid-term results compared with those obtained using CPB. METHODS: From May 21, 1997 to November 1998, 747 patients had isolated myocardial revascularization, 480 without CPB (Group A) and 267 with CPB (Group B). Exposure of the target vessels was obtained with four slings (two passed through the transverse sinus and two behind the inferior vena cava) and four deep pericardial sutures on the mobile pericardium around the left atrium (Lima stitches). The number of anastomoses/patient (when two or more conduits were used) was higher in Group B (3.1 +/- 1.0 vs 2.6 +/- 0.7, P < 0.001). More marginal branches were grafted in Group A (258 vs 239), but the percentage was higher in Group B (P < 0.001). Crude and risk adjusted mortality was similar in both groups, as well as cerebrovascular accident (CVA) and acute myocardial infarction incidences. Patients in Group A woke earlier, had less inotropes, lower creatinkinase myocardial band (CK-MB) peak, lower bleeding and less transfusion, shorter Intensive Care Unit (ICU) and postoperative stay in hospital than patients in Group B. 266 anastomoses were checked; of these 98.5% were patent and 97.0% were patent and not restrictive. CONCLUSIONS: Myocardial revascularization without CPB can provide good early- and mid-term results in selected patients. Primary endpoints (death and acute myocardial infarction) were similarly independent from the technique used. Some of the secondary endpoints were favorable in Group A: however their importance is minor. Even if we feel that some high risk patients with severe comorbidities can benefit from CPB surgery; this aspect is difficult to demonstrate scientifically. PMID- 10536954 TI - Surgical treatment of dilated cardiomyopathy with conventional techniques. AB - OBJECTIVE: We review our surgical experience using different conventional surgical techniques in the surgical treatment of the dilated cardiomyopathy (DCMP) in non-transplant eligible patients. METHODS: In this series we included patients who fit the following criteria: ejection fraction < 35%; end diastolic volume > or = 110 ml/m2; enlargement of the base of the heart (maximal mitral diameter > or = 22 mm/m2) with functional mitral regurgitation; mitral surgery to be performed in every case. Moreover, two groups were considered. (A) Normal or moderately impaired right ventricular function; PAP < 45 mmHg; elective or semielective surgery. (B) Severely impaired right ventricular function; PAP > or = 45 mmHg; severe organ failure; dependency on IABP and/or inotropes; need of ICU stay. From January 1990 to September 1998, 66 patients underwent isolated mitral valve surgery (n = 30); in the remaining 36 the Batista operation (n = 21) or exclusion of akinetic areas (n = 15) were associated. The etiology was ischemic in 42, idiopathic in 23 and post-valvular in one. RESULTS: When isolated mitral valve surgery was performed, early mortality in group A (n = 22) was 0, in group B (n = 8) 50%. Overall 5-year survival was 70.0 +/- 8.4. in group A 81.8 +/- 8.2, and in group B 37.5 +/- 17.1. When the Batista operation was performed, early mortality in group A (n = 13) was 23.1%, in group B (n = 8) 75%. Overall 2-year survival was 42.9 +/- 10.8 in group A 61.5 +/- 13.5 and in group B 25.0 +/- 15.3. When akinetic areas were excluded, early mortality in group A (n = 11) was 18.2% and in group B (n = 4) 100%. Overall 1-year survival was 53.3 +/- 12.9, in group A 72.7 +/- 13.4. CONCLUSION: Group A patients have better results in every cohort of patients considered. Even if patients selection seems to be the most important variable for early mortality and late survival, isolated mitral valve surgery, when feasible, provides the best early and late results. PMID- 10536956 TI - Spectral analysis of graft flow for anastomotic error detection in off-pump CABG. AB - OBJECTIVE: Flow probes have been introduced as a non-invasive means of anastomotic quality assessment in off-pump coronary artery bypass graft (CABG). Flow waveform morphology cannot reliably be assessed visually unless severe anastomotic stenosis is present ( > 90%). We applied spectral analysis techniques to determine whether the frequency content of graft flow can improve the surgeon's ability to detect anastomotic errors. METHODS: Forty-six mammary to left anterior descending artery (LAD) anastomoses were created in mongrel dogs during off-pump CABG surgery. Graft flow was measured using transit-time flow probes with the LAD closed, and the mammary graft patent and with varying degrees of stenosis. The degree of anastomotic stenosis was created by an artificial stitch and verified by random postoperative angiography. Spectral analysis of the graft flow waveforms was performed. Differences in the magnitude and phase components of the graft flow for the first five harmonics were determined for the varying anastomosis test conditions. Differences were determined using analysis of variance and least square means techniques. RESULTS: The magnitude of the fundamental (zeroth) harmonic was statistically different in the internal mammary artery (IMA) with 0-25% stenosis compared to IMA with 50-75% stenosis (P < 0.01 ). Further, the magnitude of the first, second, and fourth harmonics were statistically different in IMA with 0-25% compared to IMA with 75% (P < 0.01). The phase of the first harmonic was statistically different in IMA with 25% stenosis than IMA with 50% stenosis (P < 0.01 ). No differences in interaction between the LAD and IMA for all ranges of stenosis were detected (P > 0.50). CONCLUSION: Spectral analysis of graft flow waveforms may be beneficial in detecting lesser degrees of anastomotic stenosis (i.e. < 90%) compared to traditional visual assessment of mean graft flow and/or graft flow waveform morphology. PMID- 10536955 TI - Indication and patient selection in minimally invasive and off-pump' coronary artery bypass grafting. AB - BACKGROUND: The selection criteria to perform 'off-pump' coronary bypass (OPCAB) grafting are not well defined. The aim of this presentation is to outline the indications and the patient selection on the basis of 2 years experience with 572 OPCAB procedures. MATERIALS AND METHODS: From November 1996 minimally invasive coronary bypass grafting was performed in 406 patients using a limited minithoracotomy for single left anterior descending artery (LAD) revascularization (group A). In 166 patients full sternotomy and OPCAB grafting for single or multiple vessel revascularization was performed (group B). RESULTS: In group A the procedure could be performed 'off-pump' together with a limited thoracotomy in 406 out of 457 patients (88.8%) who were scheduled for single graft revascularization to LAD. Exposure and quality of the LAD was good in 308/406 (76.0%) of the patients. The decision for sternotomy was made for different preoperative characteristics of these patients: Obese female patients 16/457 (3.5%), angiographic evidence of an intramyocardial running LAD 6/457% (1.4%), diffusely diseased and small LAD 11/457 (2.4%) severe COPD 3/457 (0.7%), unstable angina 11/457 (2.4%), emergency revascularization after failed PTCA 4/457 (0.8%). In 315/406 (77.8%) of the minimally invasive direct coronary artery bypass (MIDCAB)-patients exposure and quality of the LAD was good, in 97/406 (22.2%) moderate or even bad. In the latter subgroup stenosis free anastomosis was reduced (86.5%) compared to the subgroup of good exposure and quality with 98.3%. In group B selection for sternotomy and 'off-pump' procedure was made in 117/166 (70.4%) patients with a normal preoperative status (stable angina, ejection fraction > 35%) and with coronary lesions amenable for beating heart surgery (proximal RCA lesion > 80%, not calcified and well defined POD and marginal branches). In 49/166 (29.5%) decision for 'off-pump' procedure was made on the basis of a potential risk for cardiopulmonary bypass (CPB) such as acute myocardial infarction in 10/166 (6.0%), reduced ventricular function with EF < 35 in 28/166 (16.9%), calcified ascending aorta 4/166 (2.4%) or concomitant diseases 7/166 (2.5). CONCLUSION: To maintain excellent results after single LAD revascularization using the MIDCAB-approach, appropriate patient selection is crucial. Indication for sternotomy and 'off-pump' single LAD revascularization should made in those patients excluded for MIDCAB and in patients scheduled for multiple vessel-CABG who are at high risk for CPB (concomitant pulmonary, renal, neurological diseases or severely impaired left ventricular dysfunction) and have suitable target coronary arteries in term of location and quality. PMID- 10536957 TI - Off-pump arterial grafting: 125 cases using the Medtronic-Utrecht Octopus. AB - OBJECTIVES: The use of arterial grafts in coronary bypass surgery requires a high degree of cardiac stabilization, traditionally achieved with cardiopulmonary bypass and cardioplegic arrest. The Medtronic-Utrecht Octopus has recently been developed as an advanced cardiac stabilization device, based on its unique suction method for regional epicardial immobilization and retraction. The objective of this study was to investigate the feasibility of using this device to enable total arterial revascularization on the beating, working heart. METHODS: From May 1997 to November 1998, off-pump coronary artery bypass using exclusively arterial grafts was performed in 125 selected patients (108 males), aged 2682 years (mean 61.1 +/- 10.5 years). Coronary artery immobilization was achieved with the Octopus, which uses local epicardial suction and avoids cardiac compression. Aortic anastomoses were avoided: both internal thoracic arteries and the right gastroepiploic artery were used as pedicle grafts in all but one case. All radial artery grafts and one right internal thoracic artery were used as Y grafts from the left internal thoracic artery. There were four surgical approaches: sternotomy (98 patients), left anterior small thoracotomy (20 patients), anterolateral thoracotomy (six patients) and a subxiphoid approach in one patient. RESULTS: Sternotomy: 187 grafts were performed in 98 patients (mean 1.9 grafts per patient). There were 99 grafts to anterior wall vessels, 47 grafts to posterior wall vessels and 41 grafts to lateral wall vessels. Left anterior thoracotomy: 20 patients had a single graft to the left anterior descending artery (LAD). Left anterolateral thoracotomy: three patients had a single graft to a circumflex branch, while three had composite grafts to the LAD and circumflex systems. Subxiphoid: one patient had a single graft to the posterior descending branch of the right coronary artery. There were no peri-operative deaths in any group. No patient required conversion to cardiopulmonary bypass. Three patients required conversion from a limited-access approach to sternotomy. There was one re-operation for bleeding. Postoperative stay was 27 days (mean 3.6 +/- 1.1; median 3 days) for anterior thoracotomy, 3-4 days (mean 3.5 +/- 0.6) for anterolateral thoracotomy, and 378 days (mean 6.6 +/- 8.7; median 4 days) for sternotomy. There were two late deaths in salvage patients; no patient has required cardiac intervention or re-operation. CONCLUSIONS: The Octopus maintains excellent local cardiac immobilization--enabling the routine use of arterial grafts in off-pump coronary surgery. It allows easy access to anterior wall vessels on the heart, and relatively straightforward access to the posterior wall. Circumflex branches are graftable with careful case selection and adjunctive technical maneuvers. PMID- 10536959 TI - Evaluation of two new heart valve surgery techniques: partial sternotomy and port access approaches. AB - OBJECTIVES: This review attempts to compare the port-access and partial sternotomy approaches of minimally invasive valve surgery. METHODS: Our brief experiences of the two techniques are summarized with an attempt to compare safety, cost-effectiveness of the procedure and post discharge follow-up. One hundred and two patients undergoing the procedures between May 1996 and October 1998 were analyzed. There were 65 patients in the partial sternotomy (MIV) group and 37 patients in the port-access (PAV) group. With the exception of a higher incidence of COPD in the MIV patients, there was no significant difference in pre operative variables between these two groups. RESULTS: Total operating room time, surgery time and cross-clamp time were significantly increased in the PAV group. The operative mortality of patients with MIV was 3% (n = 2) while the PAV group was 8% (n = 3) (P = ns). More new atrial fibrillation was found in the MIV (26% versus 5%, P = 0.009). Otherwise, there was no significant complications observed in either group. During the 4-6 week follow-up, of those who were employed, 76% of MIV and 69% of PAV patients had returned to work. Of the retired patients more than 95% of the patients in both groups had resumed their daily routine activity. Importantly, the study showed PAV patients returned to work about 4 weeks sooner than MIV patients. CONCLUSIONS: MIV approach is more 'surgeon friendly' and can be carried out without increased intra-operative resource utilization. The PAV approach requires formal training and capital outlay for unique equipment, disposable and ancillary procedures. From a financial perspective, if the PAV technique is to become widely accepted intra-operative efficiencies must be maximized, post-operative fast-tract protocol must be utilized, financial expenditures for disposable equipment must decrease and requirement of ancillary procedures must be reduced. PMID- 10536958 TI - Video-assisted thoracoscopic sympathectomy for severe intractable angina. AB - OBJECTIVE: Endoscopic trans-thoracic sympathectomy is a well documented, safe and successful treatment for palmar and axillary hyperhidrosis. This may also be helpful in the management of patients with intractable angina and advanced coronary disease unsuitable for coronary artery bypass graft (CABG) or percutaneous transluminal coronary angioplasty (PTCA). We evaluated video assisted thoracoscopic sympathectomy (VATS) in such patients with the aim of improving symptoms and quality of life. METHODS: Video assisted thoracoscopic sympathectomy, a minimally invasive procedure, was performed under general anaesthesia with alternating single lung ventilation. Three stab incisions were made at the level of the fourth intercostal space in the anterior and posterior axillary lines, and at the fifth intercostal space in the mid-axillary line through which an extensive thoracic sympathectomy was performed to include second to the fourth ganglia, bilaterally. RESULTS: A total of 16 patients aged 46-76 (mean 61) years were assessed for VATS. Of these 10 patients had the procedure performed; nine with previous CABG and one with diffuse coronary disease. Six patients were excluded because of an evolving MI (n = 1), left ventricular ejection fraction (LVEF) < 30% (n = 2), and chronic stable angina with no objective evidence of ischaemia (n = 3). All 10 patients had marked symptomatic improvement with reduction of both angina frequency and intensity of attacks. Mean follow-up period 11.5 months. Exercise tolerance and time to onset of angina measured on exercise treadmill was significantly increased post-VATS (P = 0.028) and maintained 1 year post-operative. CONCLUSION: VATS was associated with both reduction in angina symptoms and an increase in exercise time to onset of angina. An improved quality of life was evident. PMID- 10536960 TI - The clinical and financial impact of port-access coronary revascularization. AB - OBJECTIVE: Port-access coronary bypass grafting (CABG)was performed in an attempt to impact the clinical course of patients with coronary artery disease. METHODS: One hundred patients (56 men and 44 women) with a median age of 61 years underwent port-access coronary revascularization. The clinical and financial profiles of these patients were compared with fiscal year 1997 patients (n = 531) who underwent standard median sternotomy coronary bypass. RESULTS: Preoperative clinical demographics were similar in both groups of patients. Among the port access population there were no incidences of aortic dissection, deep vein thrombosis, conversion to median sternotomy, or death. Total time in the Intensive Care Unit (ICU), incidence of atrial fibrillation, transfusion requirements, and (subjective) pain rating at 28 days postoperatively were less in the port-access group. The average hospital cost per case was $2703.00 (US dollars) more in the port-access patients, despite a similar length of stay versus conventional sternotomy patients. CONCLUSIONS: Coronary bypass surgery can be performed safely with port-access technology with significant clinical benefits in selected patients. Currently these benefits are attained at a significant cost to the institution. PMID- 10536961 TI - Cytokines in myocardial injury: impact on cardiac surgical approach. AB - Myocardial ischemia-reperfusion injury associated with cardiac surgery is an acute inflammatory process in which activated leukocytes and endothelial cells play a critical role. Recent data indicate that the release of cytokines is crucial in inducing leukocytes and endothelial cells activation during cardiopulmonary bypass (CPB). Some inflammatory cytokines can be produced locally from the heart, particularly interleukin (IL)-8, which may further enhance leukocyte activation and accumulation in the injured myocardium. In fact, postoperative levels of cardiac troponin-I, a highly specific marker of myocardial injury, correlated strongly with IL-8 values in patients undergoing coronary artery bypass grafting (CABG). Off-pump CABG is associated with less IL 8 production compared with conventional procedure, which may in turn reduce the degree of myocardial injury. On the other hand, reduced release of IL-8 and cardiac troponin-I has also been discovered following the use of heparin-coated CPB circuits. In addition, the balance between pro- and anti-inflammatory mediators may be even more crucial in determining the extent of injury. Hence, avoiding the use of CPB or improving the biocompatibility of CPB may lead to better myocardial preservation. Research along these lines is expected to help in the development of ideal therapeutic strategies to minimize the inflammatory response and subsequent myocardial injury associated with cardiac surgery. PMID- 10536962 TI - Evaluation of myocardial metabolism and function during beating heart coronary surgery. AB - Coronary artery bypass surgery on the beating heart either via a left anterior small thoracotomy (LAST) or a median sternotomy is becoming increasing popular world-wide. Concern still remains about the potential for a temporary regional myocardial ischaemia associated with the stabilisation and occlusion of the coronary during construction of the anastomosis. This review summarises the results of a series of studies intended to evaluate the effect of beating heart coronary revascularization on myocardial function, myocardial tissue injury and clinical outcome. PMID- 10536963 TI - Alopecia areata: an autoimmune disease? AB - A wide range of hypotheses such as focal infection, trophoneuroses, and endocrine dysfunction, have been previously proposed to explain the pathogenesis of alopecia areata (AA). Currently, the most widely held belief is that AA is an autoimmune disease with cellular and/or humoral immunity directed against anagen hair follicle antigen(s). However, until recently evidence in support of an autoimmune mechanism of AA has been largely circumstantial. More fundamental evidence has recently been amassed in support of AA as an autoimmune disease by using animal models. These data include: 1) identification of cross-species hair follicle specific IgG autoantibodies, 2) The ability to induce AA in an animal model with transfer of skin from affected to naive individuals, and 3) the induction of disease by transfer of lymphocytes to human skin grafted to severe combined immunodeficiency mutant mice. A review of the previous and current data related to the autoimmune basis of AA is provided to put into perspective the future studies needed to definitively determine whether AA is an autoimmune disease. PMID- 10536964 TI - Alterations in ganglioside expression during the differentiation of human mast cells. AB - Gangliosides are physiological components of the outer cell membrane. In the present study, the role of ganglioside expression during differentiation of human mast cells was evaluated. After 11 days of culture in medium known to induce mast cell differentiation, 70% of peripheral blood mononuclear cells (PBMC) showed positive staining for the high affinity IgE receptor and tryptase on immunocytochemistry and an associated 20-fold increase of ganglioside GM3 expression. Furthermore, exogenous addition of GM3 during cultivation of PBMC in medium containing low levels of growth factors induced an increase of mast cell specific tryptase. The association of ganglioside expression with mast cell differentiation was confirmed by experiments with the human mast cell line HMC-1. FcepsilonRI-positive cultured cells enriched with immunobeads exhibited a 3-fold higher expression of GM3, compared to FcepsilonRI negative HMC-1 cells. Furthermore, measurable amounts of the gangliosides GM2, GM1 and GD1a were found only in the FcepsilonRI positive cells. A corresponding transient increase of mRNA for GalNAcT, the key enzyme in the production of these latter gangliosides, could be detected preceding the expression of these gangliosides and the FcepsilonRI by RT-PCR. Taken together, these data point to a functional role of gangliosides in the differentiation of human mast cells. PMID- 10536965 TI - Fine genetic mapping of diffuse non-epidermolytic palmoplantar keratoderma to chromosome 12q11-q13: exclusion of the mapped type II keratins. AB - Diffuse non-epidermolytic palmoplantar keratoderma (NEPPK) belongs to the heterogeneous group of skin diseases characterized by thickening of the stratum corneum of the palms and soles (1). This autosomal dominant PPK is characterized by a diffuse pattern of palmar and plantar hyperkeratosis giving the affected areas a thickened yellowish appearance with a marked erythematous edge. Linkage of diffuse NEPPK to chromosome 12q11-q13 has been demonstrated in two independent reports (2, 3). In this study, we describe detailed haplotyping with microsatellite markers mapping to this chromosomal region in three diffuse NEPPK pedigrees from the south of England. Fine mapping of a previously identified recombination event and the identification of a common disease haplotype segregating in the three pedigrees places the diffuse NEPPK locus proximal to the type II keratin gene cluster. PMID- 10536966 TI - Identification of citrulline residues in the V subdomains of keratin K1 derived from the cornified layer of newborn mouse epidermis. AB - Citrulline residues are detected in keratins and filaggrin in the cornified layers of mammalian epidermis. Such citrulline residues are formed by the enzymatic deimination of arginine residues by peptidylarginine deiminase (EC 3.5.3.15). Major deiminated keratins are thought to be partially degraded/disulfide-cross-linked keratin K1 based on the immunoblotting profiles. In order to obtain more definitive evidence of the deimination of keratin K1 and also to investigate its functional significance, we attempted to identify its preferred acting sites of peptidylarginine deiminase. A partially degraded keratin K1 fraction obtained from the cornified layer of newborn mouse epidermis was subjected to limited proteolytic cleavages, and the resulting deiminated peptides were fractionated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis or reverse-phase high-performance liquid chromatography for N terminal sequencing and/or amino acid analysis. At least two sites were identified, one in the V1 and the other in the V2 subdomains of keratin K1. An undecapeptide sequence covering the latter shows about 70% homology with an undecapeptide sequence in the V2 subdomain of human K1, a presumptive site of deimination. We speculated that the deimination of arginine residues in these subdomains might modulate their interactions with epidermal proteins other than keratins and filaggrin during the terminal stage of epidermal differentiation. PMID- 10536967 TI - Antigen retrieval of loricrin epitopes at desmosomal areas of cornified cell envelopes: an immunoelectron microscopic analysis. AB - Cell envelopes (CEs) are insoluble, chemically and mechanically tough structures formed during terminal differentiation of keratinocytes, providing skin with a protective barrier against the environment. They are 15 to 20 nm thick structures beneath the plasma membrane and continuous with desmosomal attachment plaques. Sequential deposition of several proteins including involucrin and loricrin leads to a gradual increase in envelope thickness and rigidity. Cross-linking of desmosomal components to other CE-proteins has been demonstrated and desmosomes in the cornified cells have been regarded as a part of CEs. Our previous immunoelectron microscopy studies showed that desmosomal areas of granular cells were loricrin-positive, but those in cornified cells were negative. We asked whether this is due to epitope masking and applied trypsin digestion of the electron microscopy sections to retrieve the possibly masked epitopes. Since this treatment made desmosomal structures obscure, one side of the sections was stained with anti-desmoglein antibody as an indicator of desmosomes. Trypsin was applied on the other side followed by immunolabeling with anti-loricrin antibody. Trypsin digestion indeed unmasked the loricrin epitopes in the desmoglein positive desmosomal areas of CEs. It seems therefore that loricrin is first accumulated at the desmosomes before the CE-assembly and cross-linking of loricrin occurs at the desmosomal areas of CEs as well as at the non-desmosomal areas. PMID- 10536969 TI - Purification of mast cell proteases from murine skin. AB - Different subpopulations of mast cells are characterized by their abundant contents of either tryptase or in addition chymase. These two neutral proteases are found in mast cells and may thus hold a key to the understanding of mast cell dependent reactions. Such studies are however hampered by the lack of readily available supplies of chymase. We have therefore studied the simultaneous purification of both proteases from hairless moro hr/hr mouse skin, using a sequence of salt extractions and chromatographic separations. After three steps of extraction, a 13-fold purification with an 82% yield was obtained for tryptase and a 15-fold purification with a 90% yield for chymase. Further one step purification on conventional sephadex, sephacryl and octyl sepharose columns was unsatisfactory because of further protein contamination of the fractions. Heparin affinity chromatography caused a high loss of tryptase and residual protein contamination. Gradient elution on a benzamidine sepharose 6B column resulted however in a single, low yield (17.9%) tryptase peak and a broader, high yield (>90%) chymase peak, and a 34% yield high purity fraction. The proteases thus purified exhibited their typical inhibitor profile. On Western blot analysis and on autoradiography in the presence of the serine protease inhibitor diisopropylfluorophosphate (DFP), only one 28 kD molecule with chymase activity was identified, whereas a broad 32-38 kD band of tryptase monomers was noted. Taken together, these data show that, after salt extraction and a single benzamidine affinity chromatography step, both mast cell chymase and tryptase can be separated and in case of chymase also highly purified, allowing thus for the study of biological activities of this molecule. PMID- 10536968 TI - Cytokine modulation of type XV collagen gene expression in human dermal fibroblast cultures. AB - The expression of type XV collagen was studied in cultured human dermal fibroblasts exposed to transforming growth factor-beta (TGF-beta), tumor necrosis factor-alpha (TNF-alpha) or interleukin-1beta (IL-1beta), cytokines which have been shown previously to alter the expression of several extracellular matrix genes. TGF-beta enhanced the expression of the type XV collagen gene (COL15A1) in a time-dependent manner, up to 4.3-fold after 24 h of incubation, whereas TNF alpha and IL-1beta reduced the mRNA steady-state levels by 32 and 80%, respectively. When TGF-beta and TNF-alpha were added together to the cultures, the stimulatory effect of TGF-beta on type XV collagen gene expression was abrogated, indicating antagonistic modulation by these 2 cytokines. These data suggest that the cytokines tested in this study may contribute to the regulation of type XV collagen synthesis in a variety of tissues which have recently been shown to express this particular collagen gene. PMID- 10536970 TI - How realistic is cutaneous gene therapy? AB - Recent progress with innovative, experimental gene therapy approaches in animals, and recent improvements in our understanding and manipulation of stem cells, gene expression and gene delivery systems, have raised plenty of hopes in essentially all branches of clinical medicine that hitherto untreatable or poorly manageable diseases will soon become amenable to treatment. Few other organ systems have received such enthusiastic reviews in recent years as to the chances and prospects of gene therapy as the skin, with its excellent accessibility and its pools of--seemingly--readily manipulated epithelial stem cells (cf. Cotsarelis et al., Exp Dermatol 1999: 8: 80-88). However, as in other sectors of clinical medicine, the actual implementation of general gene therapy strategies in clinical practice has been faced with a range of serious difficulties (cf. Smith, Lancet 1999: 354 (suppl 1): 1-4; Lattime & Gerson (eds.), Gene Therapy of Cancer, Academic Press, San Diego, 1999). Thus, it is critically important to carefully distinguish unfounded hype from justified hope in this embryonal area of dermatologic therapy, to discuss in detail what can be realistically expected from cutaneous gene therapy approaches in the next few years, and importantly, what kind of promises should not be made to our patients at this time. PMID- 10536971 TI - Consumer preferences: path to improvement? PMID- 10536973 TI - Eliciting consumer preferences for health plans. AB - OBJECTIVE: To examine (1) what people say is important to them in choosing a health plan; (2) the effect, if any, that giving health plan information has on what people say is important to them; and (3) the effect of preference elicitation methods on what people say is important. DATA SOURCES/STUDY SETTINGS: A random sample of 201 Wisconsin state employees who participated in a health plan choice experiment during the 1995 open enrollment period. STUDY DESIGN: We designed a computer system to guide subjects through the review of information about health plan options. The system began by eliciting the stated preferences of the subjects before they viewed the information, at time 0. Subjects were given an opportunity to revise their preference structures first after viewing summary information about four health plans (time 1) and then after viewing more extensive, detailed information about the same options (time 2). At time 2, these individuals were also asked to rate the relative importance of a predefined list of health plan features presented to them. DATA COLLECTION/EXTRACTION METHODS: Data were collected on the number of attributes listed at each point in time and the importance weightings assigned to each attribute. In addition, each item on the attribute list was content analyzed. PRINCIPAL FINDINGS: The provision of information changes the preference structures of individuals. Costs (price) and coverage dominated the attributes cited both before and after looking at health plan information. When presented with information on costs, quality, and how plans work, many of these relatively well educated consumers revised their preference structures; yet coverage and costs remained the primary cited attributes. CONCLUSIONS: Although efforts to provide health plan information should continue, decisions on the information to provide and on making it available are not enough. Individuals need help in understanding, processing, and using the information to construct their preferences and make better decisions. PMID- 10536972 TI - Simulating the effects of employer contributions on adverse selection and health plan choice. AB - OBJECTIVE: To investigate the effect of employer contribution policy and adverse selection on employees' health plan choices. STUDY DESIGN: Microsimulation methods to predict employees' choices between two health plan options and to track changes in those choices over time. The simulation predicts choice given premiums, healthcare spending by enrollees in each plan, and premiums for the next period. DATA SOURCES: The simulation model is based on behavioral relationships originally estimated from the RAND Health Insurance Experiment (HIE). The model has been updated and recalibrated. The data processed in the simulation are from the 1993 Current Population Employee Benefits Supplement sample. PRINCIPAL FINDINGS: A higher fraction of employees choose a high-cost, high-benefit plan if employers contribute a proportional share of the premium or adjust their contribution for risk selection than if employees pay the full cost difference out-of-pocket. When employees pay the full cost difference, the extent of adverse selection can be substantial, which leads to a collapse in the market for the high-cost plan. CONCLUSIONS: Adverse selection can undermine the managed competition strategy, indicating the importance of good risk adjusters. A fixed employer contribution policy can encourage selection of more efficient plans. Ironically, however, it can also further adverse selection in the absence of risk adjusters. PMID- 10536974 TI - Benchmarking organ procurement organizations: a national study. AB - OBJECTIVE: An exploratory examination of the technical efficiency of organ procurement organizations (OPOs) relative to optimal patterns of production in the population of OPOs in the United States. DATA SOURCES: A composite data set with the OPO as the unit of analysis, constructed from a 1995 national survey of OPOs (n = 64), plus secondary data from the Association of Organ Procurement Organizations and the United Network for Organ Sharing. STUDY DESIGN: The study uses data envelopment analysis (DEA) to evaluate the technical efficiency of all OPOs. PRINCIPAL FINDINGS: Overall, six of the 22 larger OPOs (27 percent) are classified as inefficient, while 23 of the 42 smaller OPOs (55 percent) are classified as inefficient. Efficient OPOs recover significantly more kidneys and extrarenal organs; have higher operating expenses; and have more referrals, donors, extrarenal transplants, and kidney transplants. The quantities of hospital development personnel and other personnel, and formalization of hospital development activities in both small and large OPOs, do not significantly differ. CONCLUSIONS: Indications that larger OPOs are able to operate more efficiently relative to their peers suggest that smaller OPOs are more likely to benefit from technical assistance. More detailed information on the activities of OPO staff would help pinpoint activities that can increase OPO efficiency and referrals, and potentially improve outcomes for large numbers of patients awaiting transplants. PMID- 10536975 TI - Unequal access to cadaveric kidney transplantation in California based on insurance status. AB - OBJECTIVE: To assess the impact of insurance status on access to kidney transplantation among California dialysis patients. STUDY SETTING: California Medicare and Medicaid dialysis populations. STUDY DESIGN: All California ESRD dialysis patients under age 65 eligible for Medicare or Medicaid in 1991 (n = 9,102) took part in this cohort analytic study. DATA COLLECTION: Medicare and California Medicaid Program data were matched to the Organ Procurement and Transplantation Network Kidney Wait List files. PRINCIPAL FINDINGS: Only 31.4 percent of California Medicaid dialysis patients were placed on the kidney transplant waiting list compared to 38.8 percent and 45.0 percent of dually eligible Medicate/Medicaid and Medicare patients, respectively. Compared to the Medicaid population, Medicare enrollees were more likely to be placed on the kidney transplant waiting list (adjusted Relative Risk [RR] = 2.10, Confidence Interval [CI] 1.68, 2.62) as were dually eligible patients (RR = 1.54, CI 1.24, 1.91). Once on the waiting list, however, Medicare enrollment did not influence the adjusted median waiting time to acquire a first cadaveric transplant (p > .05). CONCLUSIONS: California dialysis patients excluded from Medicare coverage, who are disproportionately minority, female, and poor, are much less likely to enter the U.S. transplant system. We hypothesize that patient concerns with potential subsequent loss of insurance coverage as well as cultural and educational barriers are possible explanatory factors. Once in the system, however, insurance status does not influence receipt of a cadaveric renal transplant. PMID- 10536976 TI - The effects of fee bundling on dental utilization. AB - OBJECTIVE: To examine dental utilization following an adjustment to the provincial fee schedule in which preventive maintenance (recall) services were bundled at lower fees. DATA SOURCES/STUDY SETTING: Blue Cross dental insurance claims for claimants associated with four major Ontario employers using a common insurance plan over the period 1987-1990. STUDY DESIGN: This before-and-after design analyzes the dental claims experience over a four-year period for 4,455 individuals 18 years of age and older one year prior to the bundling of services, one year concurrent with the change, and two years after the introduction of bundling. The dependent variable is the annual adjusted payment per user. DATA COLLECTION/EXTRACTION METHODS: The analysis was based on all claims submitted by adult users for services received at recall visits and who reported at least one visit of this type between 1987 and 1990. In these data, 26,177 services were provided by 1,214 dentists and represent 41 percent of all adult service claims submitted over the four years of observation. PRINCIPAL FINDINGS: Real per capita payment for adult recall services decreased by 0.3 percent in the year bundling was implemented (1988), but by the end of the study period such payments had increased 4.8 percent relative to pre-bundling levels. Multiple regression analysis assessed the role of patient and provider variables in the upward trend of per capita payments. The following variables were significant in explaining 37 percent of the variation in utilization over the period of observation: subscriber employment location; ever having received periodontal scaling or ever having received restorative services; regular user; dentist's school of graduation; and interactions involving year, service type, and regular user status. CONCLUSIONS: The volume and intensity of services received by adult patients increased when fee constraints were imposed on dentists. Future efforts to contain dental expenditures through fee schedule design will need to take this into consideration. Issues for future dental services research include provider billing practices, utilization among frequent attenders, and outcomes evaluation particularly with regard to periodontal care and replacement of restorations. PMID- 10536977 TI - Managing hospitals in turbulent times: do organizational changes improve hospital survival? AB - OBJECTIVE: To examine (1) the degree to which organizational changes affected hospital survival; (2) whether core and peripheral organizational changes affected hospital survival differently; and (3) how simultaneous organizational changes affected hospital survival. DATA SOURCES: AHA Hospital Surveys, the Area Resource File, and the AHA Hospital Guides, Part B: Multihospital Systems. STUDY DESIGN: The study employed a longitudinal panel design. We followed changes in all community hospitals in the continental United States from 1981 through 1994. The dependent variable, hospital closure, was examined as a function of multiple changes in a hospital's core and peripheral structures as well as the hospital's organizational and environmental characteristics. Cox regression models were used to test the expectations that core changes increased closure risk while peripheral changes decreased such risk, and that simultaneous core and peripheral changes would lead to higher risk of closure. PRINCIPAL FINDINGS: Results indicated more peripheral than core changes in community hospitals. Overall, findings contradicted our expectations. Change in specialty, a core change, was beneficial for hospitals, because it reduced closure risk. The two most frequent peripheral changes, downsizing and leadership change, were positively associated with closure. Simultaneous organizational changes displayed a similar pattern: multiple core changes reduced closure risk, while multiple peripheral changes increased the risk. These patterns held regardless of the level of uncertainty in hospital environments. CONCLUSIONS: Organizational changes are not all beneficial for hospitals, suggesting that hospital leaders should be both cautious and selective in their efforts to turn their hospitals around. PMID- 10536978 TI - Dose confirmation of moxidectin pour-on against natural nematode infections in lactating dairy cows. AB - The nematocidal effectiveness of moxidectin, administered topically at the rate of 500 mcg/kg BW, was determined for lactating dairy cows. Naturally infected animals were given either topical vehicle or moxidectin (Cydectin Pour-On Fort Dodge Animal Health) at the rate of 1 ml/10 kg BW (10 animals per treatment group), and sacrificed 14-18 days post-treatment for nematode enumeration. 100% efficacies were recorded for Ostertagia lyrata males, Cooperia punctata males and Oesophagostomum radiatum L4, with treatment group differences in geometric means significant (P < 0.05) for all. Populations of Trichostrongylus L4 and adult O. radiatum were also reduced by 100%, but low prevalence rates in the control animals precluded meaningful statistical inference. Nematode populations for which efficacies ranged from 96.7 to 99.6% (based on geometric means) and for which treatment group differences were significant (P < 0.05) included Ostertagia spp. adult females, inhibited L4 and developing L4, O. ostertagi adult males, Trichostrongylus axei adults and Cooperia spp. adult females. For all nematodes combined, moxidectin was 98.9% efficacious. In addition to exhibiting excellent nematocidal effectiveness, topical moxidectin was demonstrated to be safe, with animal health and milk production unaffected during the study. PMID- 10536979 TI - The effect of simulated rainfall on the efficacy of doramectin pour-on against nematode parasites of cattle. AB - Two studies were conducted with doramectin topically administered at 500 microg/kg body weight to assess retention of therapeutic efficacy against nematode infections of cattle before, and after, simulated rainfall. In the first study, 50 heifers, with patent nematode infections, were allocated to one of five treatment groups. An untreated control group and one doramectin-treated group were not exposed to simulated rainfall. Simulated rainfall was applied at a rate of 25.4 mm of water in 35 min to three of the five groups: one group immediately before treatment, the second group 90 min after treatment, and the third group 24 h after treatment. Fecal samples were collected for determining egg counts 14 days after treatment. Percentage efficacy ranged from 97.3% to 100% in all treated calves, regardless of exposure to simulated rainfall. The second study involved 40 mixed-sex cattle that were allocated to one of four treatment groups (one saline control and three doramectin-treated groups). All cattle were confirmed to be free of nematode infections prior to oral gavage with infective larvae of Dictyocaulus viviparus, Cooperia oncophora, and Ostertagia ostertagi. Twenty-six days after infection, three groups were treated with doramectin pour on and exposed to 20 mm of simulated rainfall over 40 min: one group 60 min before treatment, the second 20 min after treatment, and the third 40 min after treatment. Approximately two weeks after treatment, all cattle were necropsied for worm counts. In all treated groups, the percentage efficacy against O. ostertagi and D. viviparus was >99% to 100%. Percentage efficacy against Cooperia ranged from 97% to 98%. Results indicated that doramectin pour-on remains efficacious against nematodes of cattle when administered immediately before or after rainfall. PMID- 10536980 TI - Serologic prevalence of Toxoplasma gondii in horses slaughtered for food in North America. AB - Serum samples from 1788 horses slaughtered for food in North America were tested for antibodies to Toxoplasma gondii using the modified direct agglutination test (MAT). Antibodies to T. gondii were found by the MAT in 124 (6.9%) of 1788 sera; the titers were 1:20 (69 horses), 1:40 (37 horses), 1:80 (9 horses), and > or =1:160 (9 horses). A total of 339 selected horses were also tested by the Sabin Feldman dye test (DT). Dye test antibodies were found in 54 horses with titers of 1:10 (29 horses) 1:20 (12 horses), 1:40 (4 horses) and 1:80 (9 horses). There was no correlation between the DT and the MAT. PMID- 10536981 TI - Isolation of Sarcocystis falcatula from the South American opossum (Didelphis albiventris) from Argentina. AB - Sarcocystis sporocysts from the intestines of four opossums (Didelphis albiventris) from Argentina were identified as Sarcocystis falcatula based on schizogonic stages and pathogenicity to budgerigars (Melopsittacus undulatus). Seven budgerigars fed sporocysts from the opossum feces died of acute sarcocystosis 8, 9, 11, 12, and 14 days after inoculation. Schizonts and merozoites found in the lungs and other organs of the budgerigars were identified as S. falcatula based on structure and immunoreactivity with S. falcatula specific antibody. Sarcocystis falcatula was also isolated in bovine monocyte cell cultures inoculated with lung tissue from a budgerigar that died nine days after ingesting sporocysts. Two budgerigars inoculated subcutaneously with 1,000,000 culture-derived S. falcatula died 11 and 12 days post-inoculation. This is the first report of S. falcatula infection in South America. PMID- 10536982 TI - Implications of isoform multiplicity of microphthalmia-associated transcription factor in the pathogenesis of auditory-pigmentary syndromes. AB - Microphthalmia-associated transcription factor (MITF) is the human homolog of a basic helix-loop-helix-leucine zipper protein (Mitf), encoded by the mouse microphthalmia locus. Mutations in the MITF gene have been identified in some patients with Waardenburg syndrome type 2 (WS2), which is a dominantly inherited disorder, characterized by varying combinations of sensorineural hearing loss and pigmentary disturbances. Furthermore, mice with mutations at the Mitf locus are associated with various phenotypes, such as white coat color, small eyes, a deficiency in mast cells, and osteopetrosis. Thus, MITF/Mitf may play an important role in differentiation of melanocytes and some other cell types. Recently we have identified two MITF isoforms with extended amino-termini, MITF-A and MITF-H. Both isoforms possess unique amino-termini that are different from the amino-terminus of the originally identified melanocyte-specific MITF (MITF M). MITF-M mRNA is exclusively expressed in melanocytes and pigmented melanoma cells, whereas MITF-A and MITF-H mRNA are widely expressed in many cell types, including retinal pigment epithelium. Transient transfection assays suggested that these isoforms possess differential transactivation capacity. It is therefore conceivable that the previously identified mutations may alter the functions of not only MITF-M but also MITF-A and MITF-H. Possible implications of the MITF isoform multiplicity in the pathogenesis of auditory-pigmentary disorders are discussed. PMID- 10536983 TI - Expression of proopiomelanocortin, corticotropin-releasing hormone (CRH), and CRH receptor in melanoma cells, nevus cells, and normal human melanocytes. AB - Proopiomelanocortin (POMC) is a 31 kDa prohormone that is processed to various bioactive peptides, including adrenocorticotropin (ACTH), melanotropins (alpha, beta, gamma-MSH), lipotropins, and endorphins. POMC is expressed not only in the pituitary gland but also in a variety of nonpituitary organs and tumors, including melanomas. We previously showed that normal human melanocytes produce and secrete alpha-MSH and ACTH, and furthermore, that advanced melanoma cells generally produce higher amounts of POMC peptides that correlate with tumor progression. To elucidate the mechanism of this upregulation, the expression of genes encoding corticotropin-releasing hormone (CRH) and its receptor, CRH-R, as well as POMC and the MSH receptor (MC1-R), was evaluated by reverse transcriptase polymerase chain reaction using cultured human melanoma cells, nevus cells, and normal melanocytes. Our results show that all melanocytic cells express CRH, CRH R, POMC, and MC1-R, with highest intensities in melanoma cells. Furthermore, immunohistochemistry shows that CRH as well as POMC is strongly expressed in advanced melanomas, such as vertically growing lesions of acral lentiginous, nodular and metastatic melanomas, in contrast to negative expression in nevus cells. These results indicate that tumor progression accentuates CRH, CRH-R, and POMC expression by melanoma cells. PMID- 10536984 TI - Keratin subunit expression in human cultured melanocytes and mouse neural crest cells without formation of filamentous structures. AB - The synthesis of keratin is considered to occur in epithelial and epidermal cells. Previous studies have not reported on keratin synthesis within melanocytes that derive from neural crest cells. Epithelial and neural crest cells originally develop from ectodermal tissue. We previously reported that the expression of keratin is a universal phenomenon seen in cultured melanoma cell lines, as demonstrated by two-dimensional polyacrylamide gel electrophoresis, western blot, and electron microscopy analyses. To further investigate the specificity of keratin function in melanocytic cells, we first examined the presence of keratin proteins in cultured human melanocytes, and unexpectedly found keratin subunits in melanocytes by the above-mentioned procedures. The keratin (K) subunits were composed of K1, K5, K8, K10, K14, K16, and K18, together with vimentin. Neural crest cells, which contain immature embryonic melanocytes developing from ectoderm, already expressed keratins; however, under electron microscopy, the expressed keratin did not form filamentous structures. Although the ATP synthase alpha-chain, which is expressed universally in cultured epidermal tumor cell lines, was also expressed in cultured melanocytes and neural crest cells, a novel malignant melanoma-related protein (MMRP) was absent in melanocytes and neural crest cells. We concluded that keratin subunits are present in both cells, but do not construct keratin filaments. PMID- 10536985 TI - Calcitonin gene-related peptide upregulates melanogenesis and enhances melanocyte dendricity via induction of keratinocyte-derived melanotrophic factors. AB - It has recently been shown that cutaneous axon terminals and epidermal melanocytes make contact via chemical synapses in human skin and that calcitonin gene-related peptide (CGRP) induces melanocyte proliferation. To further clarify the effect of neuropeptides on the biology and morphology of melanocytes, especially with respect to melanogenesis and melanocyte dendricity, organ cultures of normal human skin and cultured melanocytes were exposed to various neuropeptides present in intraepidermal nerve endings. Of the neuropeptides examined, skin exposed to CGRP in organ culture showed increases in melanocyte number, epidermal melanin content, melanosome number, and degree of melanization. CGRP alone had no significant effect on melanogenesis of cultured melanocytes, whereas the addition of medium conditioned by CGRP-stimulated keratinocytes (CGRP KCM) induced melanogenesis as indicated by biochemical assays of tyrosinase activity and melanin content. Furthermore, CGRP-KCM significantly enhanced melanocyte dendricity, a crucial factor affecting epidermal pigmentation. These findings suggest that keratinocytes produce and secrete some melanotrophic factors following stimulation with CGRP, which modulate growth, melanin synthesis, and dendricity of melanocytes. These data demonstrate intimate interactions between the cutaneous nervous system and melanocytes within the epidermal environment. PMID- 10536986 TI - A cascade of genes related to Waardenburg syndrome. AB - On some occasions, mutations of a gene cause different syndromes that may have similar phenotypes. For example, mutations of the MITF gene cause Waardenburg syndrome type 2 (Tassabehji et al, 1994; Nobukuni et al, 1996) as well as Tietz syndrome (Smith et al, 1997). On other occasions, mutations of different genes cause an identical syndrome. Molecular analyses of these genes may provide a good opportunity to not only understand such syndromes themselves but also the biologic aspects of cells relevant to these syndromes. By analyzing the genes for Waardenburg syndrome, we showed that PAX3, the gene responsible for Waardenburg syndrome type 1, regulates MITF, the gene responsible for Waardenburg syndrome type 2. Such epistatic relationships have been shown between other genes related to Waardenburg syndrome, and likely to construct a cascade. This paper proposes such a cascade, one that involves genes for PAX3, MITF, human MyoD, MYF5, c-MET, c-KIT, tyrosinase, TRP-1, human QNR-71, SOX10, EDNRB, and EDN3. PMID- 10536987 TI - Sulfur containing tyrosine analogs can cause selective melanocytotoxicity involving tyrosinase-mediated apoptosis. AB - Sulfur-containing tyrosine analogs such as 4-S-cysteaminylphenol (4-S-CAP) and its N-acetyl derivative, N-acetyl-4-S-CAP, are tyrosinase substrates and can cause selective cytotoxicity or cell death of melanocytes and melanoma cells. It is not clear, however, if the cytotoxicity derives from a cytostatic or cytocidal effect. The latter can also be either apoptotic or necrotic. This paper summarizes our attempt to clarify the nature of melanocytotoxicity and cell death by using a new derivative of 4-S-CAP, N-propionyl-4-S-CAP (NPr-CAP). The i.p. administration of NPr-CAP caused marked depigmentation of black hair follicles in C57 mice. At 12 h postadministration of NPr-CAP, follicular melanocytes showed histochemical and morphologic features indicative of apoptosis by TdT-mediated dUTP-biotin nick end labeling (TUNEL) staining and electron microscopy. The agarose gel electrophoresis of DNA from drug-treated melan a2 cells, an immortal melanocyte line of C57 black mice, showed the nucleosomal DNA ladder pattern. NPr CAP caused irreversible cytotoxicity in melan a2 and the effect was inhibited by a tyrosinase inhibitor, phenylthiocarbamide. The tyrosinase-mediated cytotoxicity of NPr-CAP was further confirmed by the decreased viability of COS 7 monkey kidney cells, which expressed a level of high tyrosinase activity through transfection of human tyrosinase cDNA. NPr-CAP, however, also transiently inhibited the proliferation of melan c cells, a control tyrosinase-negative albino melanocyte line, and vector-transfected COS 7 cells. Thus, the major process of NPr-CAP-mediated melanocytotoxicity involves cytocidal apoptosis associated with active tyrosinase. In addition, there is transient, nontyrosinase mediated cytostatic cytotoxicity. PMID- 10536988 TI - Transmembrane signaling for adhesive regulation of desmosomes and hemidesmosomes, and for cell-cell datachment induced by pemphigus IgG in cultured keratinocytes: involvement of protein kinase C. AB - We have investigated transmembrane signaling for the regulation of desmosomes and hemidesmosomes, using a human squamous cell carcinoma cell line (DJM-1) and normal human keratinocytes. This review discusses the involvement of protein kinase C (PKC) in regulation of these junctions, and signaling pathways involved in cell-cell detachment induced by pemphigus vulgaris (PV) IgG in a culture system. Cells grown in low-Ca++ conditions, which lack desmosomes, rapidly form desmosomes upon a low-normal Ca++-shift in association with PKC-activation and, in turn, PKC-activation by 12-O-tetradecanoylphorbol-13-acetate (TPA) induces desmosome formation even in low-Ca++ conditions. TPA induces serine phosphorylation of the 180 kDa-bullous pemphigoid antigen (BPAG2), generating 190 kDa-phosphorylated BPAG2, and dissociates BPAG2 from hemidesmosomes. TPA treatment also causes secretion of urokinase-type plasminogen activator (uPA) and expression of its receptor (uPAR), which activates plasminogen to plasmin and may digest extracellular domains of desmosomes and hemidesmosomes. These results suggest that PKC may play a role in activation of desmosome turnover and dysfunction of hemidesmossomes, and thus a role in up-migration of keratinocytes. Binding of PV-IgG to Dsg3 induces activation of diverse isoenzymes of PKC, linked to uPA secretion and uPAR expression. Furthermore, PV-IgG binding alone induces the serine-phosphorylation of Dsg 3, associated with its dissociation from plakoglobin and its deletion from desmosomes. This PV-IgG-induced Dsg 3 phosphorylation and Dsg 3-deletion from desmosomes may impair desmosome formation, whereas PV-IgG-induced PKC signaling mediates the uPA secretion and uPAR expression leading to digestion of preexisting desmosomes from the outside of the cell. These two different PV-IgG-activated signaling pathways may play a key role in acantholysis in PV. PMID- 10536989 TI - Programmed cell death in normal epidermis and loricrin keratoderma. Multiple functions of profilaggrin in keratinization. AB - The terminal differentiation of epidermal keratinocytes has been regarded as an example of programmed cell death. Among the proteins specifically expressed in this process is profilaggrin, which consists offilaggrin repeats and N- and C terminal domains. Profilaggrin is proteolytically processed into individual domains during the terminal differentiation. Filaggrin released from profilaggrin aggregates keratin filaments to form compacted cornified cells with a keratin pattern. A recent transfection experiment has indicated initiation of cell death by filaggrin expression constructs. The transitional cells between the granular and cornified cells show morphologic characteristics of apoptotic cells, and their nuclei contain fragmented DNA and profilaggrin N-terminal domains. This suggests that the N-terminus of profilaggrin may participate in nuclear events accompanying programmed cell death. Among inherited skin disorders with abnormal keratinization, progressive symmetric erythrokeratoderma is caused by loricrin mutation (loricrin keratoderma). In this disease, profilaggrin N-terminal domains are aggregated with mutant loricrin within condensed nuclei. These nuclei persist in the cornified layer as parakeratosis. Loricrin keratoderma could therefore be regarded as a representative form of disrupted cell death. PMID- 10536990 TI - R162W mutation of keratin 9 in a family with autosomal dominant palmoplantar keratoderma with unique histologic features. AB - Recurrent R162W mutation ofkeratin 9 has been reported in multiple families with epidermolytic hyperkeratosis (EHK)-type hereditary palmoplantar keratoderma (PPK). Recently, we have observed a family whose members showed autosomal dominant PPK with unique histologic features such as rounded, dissociated, and slightly eosinophilic keratinocytes at the middle spinous and granular layers of epidermis, but without the distinct EHK phenotype. To investigate the genotype phenotype correlation in this family, we searched for a mutation of keratin 9 and found R162W substitution in the coiled 1A region. This mutation was not detected in 50 control individuals. These results may further our understanding of the pathogenesis of EHK. PMID- 10536991 TI - Adenovirus-mediated gene transfer to keratinocytes--a review. AB - The introduction and expression of a foreign gene provide a powerful tool for investigating functions and regulation of a gene of interest; however, keratinocytes have a major drawback in that foreign genes are hardly transfected by conventional methods and stable transformants are most difficult to establish in normal keratinocytes with a limited short life span. To overcome these problems, we used an adenovirus vector, Ax, developed by Saito et al, which yields desired recombinant viruses at an efficiency about 100-fold that of conventional methods, and by which genes are expressed at a high level under the control of a composite CAG promoter. We established Ax vectors carrying various isoforms of protein kinase C (PKC). Using these vectors, we found that the eta and delta isoforms of PKC, but not the alpha and zeta isoforms, mediate terminal differentiation in normal human keratinocytes. These Ax-vectors are also applicable to organ culture of mouse embryos. Advantages and disadvantages of adenovirus vectors and their use for keratinocyte biology are reviewed. PMID- 10536992 TI - Dendritic cells play a crucial role in innate immunity to simple chemicals. AB - Recently, it has been demonstrated that immunity to infectious agents is composed of innate immunity and acquired immunity, and that dendritic cells (DC) and macrophages, both of which are the participants in the innate immunity, play a crucial role in acquired immune responses, via their expression of several costimulatory molecules and production of cytokines. It is clear that the immune system responds not only to infectious organisms but also to simple chemicals. Allergic contact hypersensitivity reaction is a good example of the immune response to simple chemicals. In contrast to the immunity to microorganisms, however, the role of the innate immune system in responses to simple chemicals still remains unclear. This paper demonstrates that the activation and apoptosis of DC are directly induced by certain simple chemicals, and we suggest that DC, as cells involved in the innate immune system, play a crucial role in the immunity to simple chemicals. PMID- 10536994 TI - Molecular mechanisms involved in the migration of epidermal dendritic cells in the skin. AB - The murine epidermis contains two types of dendritic cells (DC), Thy-1+ dendritic epidermal T cells (DETC) and Langerhans cells. In this review, we introduce our data obtained using a skin organ culture system to examine the migratory capacity of DETC and Langerhans cells into the epidermis. DETC or Langerhans cells were depleted by topical application ofclobetazole propionate (CP) solution onto the murine ears. CP-treated or untreated ear skin was co-cultured with syngeneic (semi-syngeneic, or allogenic, in experiments with Langerhans cells) epidermal cell suspension. We found (i) that donor DETC or Langerhans cells migrated into the CP-treated epidermis as well as into untreated epidermis, (ii) that leukosialin Ly48 recognized by monoclonal antibody S11 and TNF-alpha strongly inhibited donor Langerhans cell migration into the epidermis. We mention other molecules that may participate in the migration of Langerhans cells such as chemotactic cytokines, monocyte chemoattractant protein (MCP)-1, TGF-beta and skin-homing molecule, cutaneous lymphocyte-associated antigen (CLA) on Langerhans cells. PMID- 10536993 TI - Activation pattern of Langerhans cells in the afferent and efferent phases of contact hypersensitivity. AB - Langerhans cells are MHC class II (Ia) positive antigen-presenting cells that play a crucial role in the induction of contact hypersensitivity (CHS). The topical application of a hapten modifies the cell surface moieties of Langerhans cells, and activates Langerhans cells to increase their size and Ia intensity. The haptenated and activated Langerhans cells emigrate from the epidermis and thus the in situ density of Langerhans cells usually decreases during 24-48 h after the hapten application in CHS. To determine whether the early activation pattern of Langerhans cells is different between the afferent phase and the efferent phase of CHS, we compared the density and morphologic changes of Langerhans cells in CHS to trinitrochlorobenzene using nonsensitized and sensitized mice. We found that the application of a hapten induces more significant enlargement of Langerhans cell size in the afferent phase than in the efferent phase, whereas the reduction of Langerhans cell density is more marked in the efferent than in the afferent phase of CHS. Moreover, topical immunosuppressive drugs inhibit the in situ activation of Langerhans cells. PMID- 10536995 TI - Biologic roles of gangliosides G(M3) and G(D3) in the attachment of human melanoma cells to extracellular matrix proteins. AB - The biologic functions of gangliosides G(M3) and G(D3) in the attachment of human melanoma cells to extracellular matrix proteins (type I and IV collagens, fibronectin, and laminin) were investigated by using the G(D3)-deficient mutant clone (SK-MEL-28-N1) and the parent cell line SK-MEL28. SK-MEL-28-N1 (N1) (high G(M3) expression: G(M3), 97.3%; G(D3), 0%) was selected by treating SK-MEL-28 (high G(D3) but low G(M3): G(M3), 6.5%, G(D3), 93.5%) with an anti-G(D3) monoclonal antibody (R24) and rabbit complement and subsequent subcloning of the surviving cells. The N1 clone showed significantly higher ability to adhere to type I and IV collagens and laminin than the parent clone SK-MEL-28. In the N1 clone, the expression of alpha2beta1 and alpha3beta1 integrin receptors was increased, whereas in SK-MEL-28, their expression was very low or undetectable. The treatment with monoclonal antibodies directed specifically to G(D3) expressed on SK-MEL-28 inhibited the cell attachment to type IV collagen (33% inhibition of control), fibronectin (59%), and laminin (71%). These findings suggest that gangliosides G(M3) (by influencing integrin receptor levels) and G(D3) (by interacting directly with matrix proteins) might play some functional roles in attachment to extracellular matrix proteins and thereby enhance the metastatic potency of melanoma cells. PMID- 10536996 TI - The roles of keratinocyte-derived cytokines in the epidermis and their possible responses to UVA-irradiation. AB - Skin is the largest organ, covering the entire body surface. Keratinocytes (KC) are its major component. The KC, by making keratin protein, function as a protective barrier against exogenous stimuli. As KC have been demonstrated to produce various kinds of cytokines, skin plays an important role in immunologic and inflammatory responses of the body. Cytokines affect other cells and organs, mediating cellular growth and differentiation as well as inflammation and immune reactions. Thus, cytokines maintain the cellular and intercellular homeostasis. Dysregulation and abnormal production of cytokines are detected in various skin diseases. Evidence is accumulating to show the significant contribution of cytokines to the pathogenesis or severity of certain diseases. In this report, the effects of KC-derived cytokines on various components in the skin are briefly summarized. We further demonstrate that ultraviolet (UV) light has a distinct effect on the production and secretion of cytokines from KC, depending upon its wavelength. Although some KC-derived cytokines were induced both by UVA and by UVB, suggesting augmentative effects of UVA on UVB-induced cutaneous responses such as sunburn and suntan, other cytokines, including IL-10 and IL-12, were found to be differentially regulated by UVA and UVB. UVA (less than 20 kJ per m2) was found to induce IL-12 but not IL-10 in normal human KC. Our results suggest an antagonistic effect of UVA against UVB, indicating the contribution of UV irradiation to the balance between Th1 and Th2 cytokines in the in situ skin. PMID- 10536997 TI - Modulation of T-lymphocyte proliferation by exogenous natural ceramides and sphingosylphosphorylcholine. AB - Sphingolipids such as ceramide and sphingosine are abundantly present in the stratum corneum of epidermis. In atopic stratum corneum, sphingosylphosphorylcholine (SPC) is present in association with a reduction in the amount of ceramides. We have previously shown that the cellular kinetics of T cells are affected by exogenous addition of sphingosine and synthetic ceramides, raising the possibility that sphingolipids diffusing from the stratum corneum modulate skin-infiltrating T cells. By using two natural ceramides and murine T cells, this study further clarified the conditions under which exogenous ceramides enhance the proliferation of T cells. KLH-specific T cell clones 28-4 and 24-2 proliferated in response to natural ceramides when cultured for 44-48 h in the presence of concanavalin A at 1 microg per ml. Elongation of culture periods adversely inhibited the T cell proliferation, suggesting the existence of an optimal exposure time. Augmentation of DNA synthesis by natural ceramides was more pronounced in tumor necrosis factor alpha (TNFalpha)-sensitive 28-4 cells than in less sensitive 24-2 cells, and TNFalpha-induced proliferation of 28-4 cells was suppressed by the concomitant addition of natural ceramides. Similar to ceramides, SPC augmented the proliferation of resting spleen cells. Our study suggests that ceramide modulation of T cell proliferation depends on the TNFalpha sensitivity and activation level of T cells and that SPC also has a mitogenic potential for T cells. PMID- 10536998 TI - The interaction of cellular fibronectin with collagen during fibroblast-mediated contraction of collagen gels. AB - In the first instance highly hydrated collagen gels contract to dense and compact gels when populated by fibroblasts. We previously reported the involvement of fibronectin (FN) in an early process of the collagen gel contraction, utilizing a specific monoclonal antibody dubbed A3A5 (MoAb-A3A5) that inhibits the gel contraction. This study was performed to further characterize the role of the epitope for MoAb-A3A5 in the interaction between fibroblasts and collagen fibrils. Although both cellular FN (cFN) and plasma FN (pFN) were reactive with MoAb-A3A5, the FN that actually participates in a process of the gel contraction was shown to be cFN. The gel contraction was significantly accelerated when fibroblasts were pretreated with excess amounts of cFN and was significantly inhibited when the collagen molecules were pretreated with excess cFN. Such effects of the pretreatments were not observed for pFN. The involvement of cFN, but not pFN, in the interaction of fibroblasts with collagen fibrils was additionally shown by the similar inhibitory action of cFN, but not pFN, on the spreading and elongation of fibroblasts on collagen fibrils. The epitope for MoAb A3A5 was strongly suggested to be a new functional domain responsible for the interactions between fibroblasts and native collagen molecules. This was not the case for those with denatured one, because fibroblasts on collagen fibrils were not stainable with MoAb-A3A5, whereas the interactions on gelatin were stainable. The lack of the reactivity of fibroblasts on collagen fibrils toward MoAb-A3A5 was not a result of the absence of FN on the cell membrane, but seemed to be a steric hindrance to the access of the antibody. PMID- 10536999 TI - DNA repair, DNA replication, and UV mutagenesis. AB - Cells that have been irradiated with ultraviolet light (UV) suffer damage to their DNA, primarily in the form of covalent linkage between adjacent pyrimidines. Such photoproducts represent blocks to RNA and DNA polymerases and are potentially mutagenic. Blockage of RNA polymerase II by a photoproduct in the transcribed strand of an active gene leads to induction of the p53 protein, which induces pleiotropic responses that may include apoptotic cell death. If a cell survives, the blocked polymerase targets the nucleotide excision repair machinery to the site of the lesion, which is repaired in an error-free manner. Repair coupled to transcription in this manner strongly influences the mutation spectrum induced by UV, reducing the proportion of base substitutions that arise from photoproducts on the transcribed strand. If the damage persists when the DNA is replicated in S-phase, either because the cell is unable to repair the damage or because there is insufficient time between the induction of damage and the onset of S-phase. To do so, the replicative DNA polymerase complex may be blocked. In this situation, lesion bypass can be accomplished using an error-free mechanism, or using an error-prone mechanism that involves the newly described, non processive DNA polymerase zeta encoded by the human homolog of the yeast REV3 gene. PMID- 10537000 TI - Ultraviolet radiation induced signature mutations in photocarcinogenesis. AB - The photons of sunlight begin a series of genetic events in skin leading to cancer. UV signature mutations provide an alternative to inherited mutations as a way of identifying genes that are involved in cancer development. They augment epidemiologic and clinical data by serving as molecular evidence for the role of UV radiation in skin carcinogenesis. Signature mutations are present in TP53 and PTCH, two tumor suppressor genes responsible for non-melanoma skin cancer. We review evidence that clones of TP53-mutated cells are present in normal human and murine epidermis exposed to UVB and conclude that, in addition to being a tumorigenic mutagen, sunlight acts as a tumor promoter by favoring the clonal expansion of TP53 mutated cells. These combined actions of sunlight result in normal individuals' carrying a substantial burden of keratinocytes predisposed to cancer. Thus cancer involves both a single-cell problem and a multi-cell problem; in skin cancer, sunlight appears to drive both. PMID- 10537002 TI - Light and death: photons and apoptosis. AB - Phototherapies like photodynamic therapy (PDT), UVA1, UVB, and PUVA treat skin diseases. These phototherapies work because they alter cytokine profiles, change immune cytotoxicity in the skin, and directly kill diseased cells by apoptosis. Apoptosis is a term that only describes the morphologic changes a cell undergoes during this mode of cell death. The terms "immediate", "intermediate", and "delayed" apoptosis segregate the different apoptotic mechanisms into three kinetic categories, whereas the terms preprogrammed cell death (pre-PCD) and programmed cell death (PCD) describe the underlying mechanisms. Immediate apoptosis (T< or =0.5 h post-exposure) is triggered by singlet-oxygen damage that opens the mitochondrial megachannel, which can be mediated by PDT or UVA1 radiation. It is a pre-PCD mechanism of apoptosis, i.e., protein synthesis is not required post-insult, because all the necessary components are constitutively synthesized and only need to be activated. Intermediate apoptosis (T< or =4 h>0.5 h) is initiated by receptor cross-linking on the plasma membrane, which can be achieved using high doses of UVB or UVC radiation. It is also a pre-PCD mechanism. Delayed apoptosis (T>4 h) is induced by DNA damage that can be caused by X-rays, PUVA, UVC, UVB, UVA, and PDT. It is a PCD mechanism of apoptosis, i.e., protein synthesis is required post-insult. These three apoptotic mechanisms each access one of two "points-of-no-return" located on the mitochondrial membrane, which activate different, but not mutually exclusive, final pathways of apoptosis. This review discusses the latest findings on these apoptotic mechanisms and their implications in phototherapies. PMID- 10537001 TI - Psoriasis, PUVA, and skin cancer--molecular epidemiology: the curious question of T-->A transversions. AB - Photochemotherapy with 8-methoxypsoralen and long wavelength ultraviolet radiation (PUVA) is commonly used to treat psoriasis and vitiligo. These vastly different diseases respond to the therapy by different mechanisms even though the immediate effects of the therapy - photoadduct formation - is the same for both. Because psoriasis is not cured by PUVA, patients receive many treatments over their lifetime and develop a significant risk for the development of skin cancers (primarily squamous cell carcinomas). In this review the basic aspects of psoralen photobiology are reviewed briefly. In addition the impact of the analysis of mutations in the tumor suppressor gene, p53, are summarized. An unexpected mutation spectrum (very few T-->A transversions and frequent UVB signature C-->T transitions) suggest that effects other than direct DNA photoadduct formation may be at play. The roles of reactive oxygen species induced base changes as well as other clastogenic factors are discussed. This analysis suggests that it may be possible to improve the therapeutic efficacy of PUVA by a careful evaluation of the mode of delivery. PMID- 10537003 TI - Biochemical control of melanogenesis and melanosomal organization. AB - Current knowledge on the regulation of mammalian pigmentation at the genetic and biochemical level, and constituents that participate in melanosomal organization, is summarized. Approximately 25% of the more than 80 genes known to regulate pigmentation in mammals have been cloned and characterized to date. Almost half of those encode proteins that localize, either specifically or nonspecifically, to melanosomes; mutations in those genes generally lead to phenotypic changes in pigmentation as well as in other pleiotropic changes. The expression and function of these proteins not only affects phenotypic appearance, but also the properties of melanins, especially their photoprotective characteristics. Because many of those melanosomal proteins also serve as melanoma-specific targets, regulation of their expression has dramatic implications for immune targeting of malignant melanoma. PMID- 10537004 TI - Participation of the melanocortin-1 receptor in the UV control of pigmentation. AB - The cloning of the melanocortin-1 receptor (MC1R) gene from human melanocytes and the demonstration that these cells respond to the melanocortins alpha-melanocyte stimulating hormone (alpha-MSH) and adrenocorticotropic hormone (ACTH) with increased proliferation and melanogenesis have renewed the interest in investigation the physiological role of these hormones in regulating human pigmentation. Alpha-melanocyte stimulating hormone and ACTH are both synthesized in the human epidermis, and their synthesis is upregulated by exposure to ultraviolet radiation (UVR). Activation of the MC1R by ligand binding results in stimulation of cAMP formation, which is a principal mechanism for inducing melanogenesis. The increase in cAMP is required for the pigmentary response of human melanocytes to UVR, and for allowing them to overcome the UVR-induced G1 arrest. Treatment of human melanocytes with alpha-MSH increases eumelanin synthesis, an effect that is expected to enhance photoprotection of the skin. Population studies have revealed more than 20 allelic variants of the MC1R gene. Some of these variants are overexpressed in individuals with skin type I or II, red hair, and poor tanning ability. Future studies will aim at further exploration of the role of these variants in MC1R function, and in determining constitutive human pigmentation, the response to sun exposure, and possibly the susceptibility to skin cancer. PMID- 10537005 TI - DNA photodamage stimulates melanogenesis and other photoprotective responses. AB - Ultraviolet (UV) irradiation is a major source of environmental damage to skin. Melanin pigmentation protects against this damage by absorbing UV photons and UV generated free radicals before they can react with DNA and other critical cellular components; and UV-induced melanogenesis or tanning is widely recognized as exposed skin's major defense against further UV damage. This article reviews extensive data suggesting DNA damage or DNA repair intermediates directly triggers tanning and other photoprotective responses. Evidence includes the observations that tanning is enhanced in cultured pigment cells by accelerating repair of UV-induced cyclobutane pyrimidine dimers or by treating the cells with UV-mimetic DNA-damaging chemicals. Moreover, small single stranded DNA fragments such as thymidine dinucleotides (pTpT), the substrate for almost all DNA photoproducts, also stimulates tanning when added to cultured pigment cells or applied topically to intact skin. In bacteria, single stranded DNA generated by DNA damage or its repair activates a protease that in turn derepresses over 20 genes whose protein products enhance DNA repair and otherwise promote cell survival, a phenomenon termed the SOS response. Interestingly, pTpT also enhances repair of UV-induced DNA damage in human cells and animal skin, at least in part by activating the tumor suppressor protein and transcription factor p53 and thus upregulating a variety of gene products involved in DNA repair and cell cycle regulation. Together, these data suggest that human cells have an evolutionarily conserved SOS-like response in which UV-induced DNA damage serves as signal to induce photoprotective responses such as tanning and increased DNA repair capacity. The responses can also be triggered in the absence of DNA damage by addition of small single-stranded DNA fragments such as pTpT. PMID- 10537006 TI - Ultraviolet radiation mutagenesis of hedgehog pathway genes in basal cell carcinomas. AB - The identification of mutations in Hedgehog (HH) pathway genes in some basal cell carcinomas (BCC) and the detection of HH pathway dysregulation in almost all BCC confirms the importance of this developmental regulatory pathway in human BCC tumorigenesis. Moreover, the occurrence of UVB signature mutations in key HH pathway genes in BCC provides the first genetic evidence that UV radiation (UVR) may be the principal mutagen involved in BCC tumorigenesis. We review herein current advances in the understanding of the role of the HH pathway in BCC tumorigenesis including transgenic and knock-out animal models of HH pathway dysregulation. Furthermore, we summarize abnormalities in other tumor suppressors and oncogenes including ras and p53 and evidence for interactions between these regulatory genes and the HH pathway. PMID- 10537007 TI - Spectral regions contributing to melanoma: a personal view. AB - Although human cutaneous melanoma is a complicated disease, the principal etiologic agent for its incidence in fair skin individuals is exposure to sunlight. In order to understand the epidemiology of melanoma - temporal effects, latitude effects, sunscreen effects, albino susceptibility, and differences from nonmelanoma skin cancer -one must approach the problem by obtaining clues indicating which wavelengths in sunlight are effective in inducing melanomas. One way is to use an animal model. At present, the only suitable model is a backcross hybrid of small tropical fish of the genus Xiphophorus, bred to have only one tumor suppressor gene. Single UV exposures to 7-d-old fish induce melanomas readily observable by 4 mo. The initial slopes of dose-response curves for exposures at 302, 313, 365, 405, 436, and 547 nm yield sensitivity as a function of wavelength. This action spectrum does not look like the spectrum for light absorption by DNA (mostly in the UVB), but has appreciable sensitivities in the UVA and visible regions, and looks like a direct effect of light on DNA plus a large indirect effect on DNA by absorption of light by the intracellular melanin. Because the UVB is only a fraction of solar irradiance, one may calculate that 90% of melanoma induction in humans arises from UVA and visible, assuming the human spectrum is similar to the fish spectrum. The implications of this calculation are that (i) depletion of stratospheric ozone will not affect melanoma incidence, (ii) an increase in sun exposure time as a result of using UVB sunscreens could increase the risk of melanoma, and (iii) the use of high UVA sun tanning devices could increase the risk of melanoma. PMID- 10537008 TI - Familial melanoma; CDKN2A and beyond. AB - The most common hereditary melanoma susceptibility disorder is the familial atypical multiple mole-melanoma (FAMMM) syndrome. FAMMM is regarded as an ideal natural model to study the very complex pathologic mechanism of melanoma. In 1994, cloning of the melanoma susceptibility gene CDKN2A was thought to give answers to many questions on genotype-phenotype correlations in familial melanoma. Today, 4 y later, germline mutations cosegregate with melanoma in only 40%-50% of the families predisposed to this disease. The hunt for genes and modifying genes is on again. Through the years the very well-characterized Dutch FAMMM families have proven to be valuable study subjects in melanoma research. This paper describes over 10 y of melanoma research illustrated by research performed in the Dutch FAMMM families. PMID- 10537009 TI - Effect of ultraviolet light on the release of neuropeptides and neuroendocrine hormones in the skin: mediators of photodermatitis and cutaneous inflammation. AB - Ultraviolet (UV) irradiation of the skin causes both inflammation and alterations in the skin immune system. There is increasing experimental evidence that UV induced skin inflammation is influenced by the sensory nervous system and the neuroendocrine system in the skin. The resulting complex network of cytokines, chemokines, neuropeptides, neuropeptide-degrading enzymes, neurohormones, and other inflammatory mediators mediate photodermatitis and cutaneous inflammation. Neuropeptides such as substance P (SP) and calcitonin gene-related peptide (CGRP) are released from sensory nerves innervating the skin upon UV exposure. In addition, a variety of cells in the skin produce increased neuroendocrine hormones such as proopiomelanocortin (POMC) peptides and their receptors as well as neurotrophins after UV exposure. Neuropeptides and neurohormones are capable of directly or indirectly mediating UV-induced cutaneous neurogenic inflammation by the induction of vasodilatation, plasma extravasation, and augmentation of UV induced cytokine, chemokine, or cellular adhesion molecule expression required for activation and trafficking of inflammatory cells into the inflamed tissue. Neuropeptides and neurotrophins may also play a role in the repair of cutaneous UV injury. In addition to proinflammatory effects, UV-induced neuropeptides and neurohormones such as CGRP and alpha-melanocyte-stimulating hormone may have immunosuppressive effects in the skin. This review will focus on the role that SP, CGRP, POMC peptides, and their receptors may play in modulating UV-induced inflammation in the skin. PMID- 10537010 TI - Mechanisms involved in ultraviolet light-induced immunosuppression. AB - Ultraviolet light (UV) represents one of the most relevant environmental factors influencing humans, especially with regard to its hazardous health effects, which include premature skin aging, skin cancer, and exacerbation of infectious diseases. Several of these effects are mediated by the immunosuppressive properties of UV. UV can compromise the immune system in several ways, e.g., by affecting the function of antigen-presenting cells, inducing the release of cytokines, and modulating the expression of surface molecules. Recently a link between UV-induced immunosuppression and apoptosis was recognized. In the following, the basic mechanisms underlying UV-induced immunosuppression will be discussed. PMID- 10537011 TI - Sunscreen effects on UV-induced immune suppression. AB - In order to protect the public against the adverse effects of sunlight, the scientific, medical, and particularly the dermatologic community has promoted "safe sun exposure." This strategy includes sun avoidance whenever possible, wearing hats and other protective clothing and/or devices, such as sunglasses, and extensive use of sunscreens. Sunscreen efficacy is determined by measuring the ability of the sunscreen to block ultraviolet (UV)-induced erythema (sun protection factor or SPF), and most sunscreen formulations on the market, if used properly, are very good at preventing erythema and sunburn. How well sunscreens protect against the other adverse effects of sunlight, such as immune suppression, is not as clear. The purpose of this paper is to review and discuss the literature in this area, concentrating on some of the complications of determining how well sunscreens protect against UV-induced immune suppression. PMID- 10537012 TI - Mechanisms of ultraviolet (UV) B and UVA phototherapy. AB - Ultraviolet (UV) radiation has been used for decades with great success and at a constantly increasing rate in the management of skin diseases, becoming an essential part of modern dermatologic therapy (Krutmann et al, 1999). For phototherapy, irradiation devices emitting either predominantly middlewave UV (UVB, 290-315 nm) or longwave UV (UVA, 315-400 nm) radiation are employed. In former years, patients were treated with broad-band UVB, broad-band UVA, or combination regimens. Broad-band UV phototherapy, however, is being replaced more frequently by the use of irradiation devices that allow treatment of patients' skin with selected emission spectra. Two such modalities which have their origin in European Photodermatology are 311 nm UVB phototherapy (which uses long-wave UVB radiation above 300 nm rather than broadband UVB) and high-dose UVA1 therapy (which selective employs long-wave UVA radiation above 340 nm). In Europe, 311 nm UVB phototherapy has almost replaced classical broad-band UVB phototherapy and has significantly improved therapeutic efficacy and safety of UVB phototherapy (van Welden et al, 1988; Krutmann et al, 1999). The constantly increasing use of UVA-1 phototherapy has not only improved UVA phototherapy for established indications such as atopic dermatitis (Krutmann et al, 1992a, 1998; Krutmann, 1996), but has also provided dermatologists with the opportunity to successfully treat previously untractable skin diseases, e.g., connective tissue diseases (Stege et al, 1997; Krutmann, 1997). These clinical developments have stimulated studies about the mechanisms by which UVB and UVA phototherapy work. The knowledge obtained from this work is an indispensable prerequisite to make treatment decisions on a rationale rather than an empirical basis. Modern dermatologic phototherapy has started to profit from this knowledge, and it is very likely that this development will continue and provide dermatologists with improved phototherapeutic modalities and regimens for established and new indications. This review aims to provide an overview about current concepts of the mode of action of dermatologic phototherapy. Special emphasis will be given on studies that have identified previously unrecognized immunosuppressive/anti inflammatory principles of UV phototherapy. PMID- 10537013 TI - Photosensitivity diseases: cutaneous lupus erythematosus. AB - Ultraviolet radiation plays an important role in the induction of lesions in many patients with cutaneous lupus. In the photosensitive subset of lupus, subacute cutaneous lupus, the effects of ultraviolet radiation likely act in concert with specific autoantibodies, particularly anti-Ro-related autoantibodies, to produce lesions. Potential effects of ultraviolet radiation on the induction of cutaneous lupus, and the potential interplay of specific autoantibodies with ultraviolet radiation are discussed. The steps involved in the induction of cutaneous lupus lesions by ultraviolet radiation have not been fully elucidated. Recent advances in phototesting and analysis of the genetics of lupus should clarify the events leading to photosensitive cutaneous lupus lesions. PMID- 10537014 TI - Ultraviolet A1 (340-400 nm) irradiation and systemic lupus erythematosus. AB - Short wavelengths of ultraviolet (UV) light are clearly harmful in systemic lupus erythematosus (SLE), but the action of long UV wavelengths in SLE is more enigmatic. In a series of animal and human studies, long-wavelength UV radiation, i.e., radiation in the ultraviolet-A1 (UVA1) range (340-400 nm), has proven effective in the treatment of SLE. Disease amelioration and a marked decrease in mortality followed ultraviolet-A (UVA) radiation (320-400 nm) of the New Zealand White/New Zealand Black mouse model of lupus. A follow-up study in the same animal suggested that the longer wavelengths (UVA1, 340-400 nm) in the UVA wave band were primarily responsible. There followed four human studies. The first three of these provided data indicating that low-dose UVA1 radiation significantly reduced constitutional symptoms, joint pain, rashes, and the systemic lupus activity measures, a validated gauge of disease activity in SLE. The fourth human study showed that the therapeutic action of low-dose UVA1 action persisted or progressed long term, a period averaging 3.4 y. UVA1 effects on DNA repair, cell-mediated immunosuppression, tumor necrosis factor alpha release, and apoptosis contrast markedly with those of ultraviolet B (UVB, 280-320 nm) radiation and afford a possible basis for the salutary action of this modality of treatment. The unique features of UVA1 wavelengths may be suited to further therapeutic use, not only in SLE but also in other immunologic disorders. PMID- 10537015 TI - Photopheresis: clinical applications and mechanism of action. AB - Photopheresis is a leukapheresis-based therapy that utilizes 8-methoxypsoralen and ultraviolet A irradiation. Photopheresis is currently available at approximately 150 medical centers worldwide. Recent evidence suggests that this therapy used as a single agent may significantly prolong life, as well as induce a 50%-75% response rate among individuals with advanced cutaneous T cell lymphoma (CTCL). Furthermore, a 20%-25% complete response rate with photopheresis alone, or in combination with other biologic response modifiers, has been obtained at our institution among patients with Sezary syndrome. These complete responses have been characterized by the complete disappearance of morphologically atypical cells from the skin and blood. The use of sensitive molecular techniques has also confirmed the sustained disappearance of the malignant T cell clone from the blood of patients with complete responses. In addition to the treatment of CTCL, numerous reports indicate that photopheresis is a potent agent in the therapy of acute allograft rejection among cardiac, lung, and renal transplant recipients. Chronic graft versus host disease also appears to be quite responsive to photopheresis therapy. Likewise, there may also be a potential role for photopheresis in the therapy of certain autoimmune diseases that are poorly responsive to conventional therapy. The immunologic basis for the responses of patients with these conditions is likely due to the induction of anticlonotypic immunity directed against pathogenic clones of T lymphocytes. Treatment-induced apoptotic death of pathogenic T cells and activation of antigen presenting cells are postulated to have important effects in this therapeutic process. PMID- 10537016 TI - In vivo and in vitro evidence for hydrogen peroxide (H2O2) accumulation in the epidermis of patients with vitiligo and its successful removal by a UVB-activated pseudocatalase. AB - To date there is compelling in vitro and in vivo evidence for epidermal H2O2 accumulation in vitiligo. This paper reviews the literature and presents new data on oxidative stress in the epidermal compartment of this disorder. Elevated H2O2 levels can be demonstrated in vivo in patients compared with healthy controls by utilizing Fourier-Transform Raman spectroscopy. H2O2 accumulation is associated with low epidermal catalase levels. So far, four potential sources for epidermal H2O2 generation in vitiligo have been identified: (i) perturbed (6R)-L-erythro 5,6,7,8 tetrahydrobiopterin (6BH4) de novo synthesis/recycling/regulation; (ii) impaired catecholamine synthesis with increased monoamine oxidase A activities; (iii) low glutathione peroxidase activities; and (iv) "oxygen burst" via NADPH oxidase from a cellular infiltrate. H2O2 overproduction can cause inactivation of catalase as well as vacuolation in epidermal melanocytes and keratinocytes. Vacuolation was also observed in vitro in melanocytes established from lesional and nonlesional epidermis of patients (n = 10) but was reversible upon addition of catalase. H2O2 can directly oxidize 6BH4 to 6-biopterin, which is cytotoxic to melanocytes in vitro. Therefore, we substituted the impaired catalase with a "pseudocatalase". Pseudocatalase is a bis-manganese III-EDTA-(HCO3-)2 complex activated by UVB or natural sun. This complex has been used in a pilot study on 33 patients, showing remarkable repigmentation even in long lasting disease. Currently this approach is under worldwide clinical investigation in an open trial. In conclusion, there are several lines of evidence that the entire epidermis of patients with vitiligo is involved in the disease process and that correction of the epidermal redox status is mandatory for repigmentation. PMID- 10537017 TI - Do sunscreens increase or decrease melanoma risk: an epidemiologic evaluation. AB - Ultraviolet adiation is an important cause of melanoma, so the use of sunscreen lotions has been advocated for melanoma prevention. Several arguments have been raised in opposition to this inference. Sunscreen use may interfere with cutaneous vitamin D synthesis, which some have hypothesized may lower melanoma risk. Sunscreen users may compensate for their sunscreen use by staying out much longer in the sun, or may use sunscreen lotions inconsistantly. Published melanoma case-control studies have not consistantly demonstrated a protective effect of sunscreens; however, these studies do not provide strong evidence, ultraviolet radiation is a known cause of melanoma, and ultraviolet B may be particularly potent, so on balance the evidence supports continued advocacy of sunscreen lotion use as part of an overall sun-protection regimen. Uncertainty will remain, however, until the action spectrum of melanoma is convincingly demonstrated or the methodologic limitations of existing epidemiologic evidence are overcome. The latter may require another decade or more of experience with sunscreen use. PMID- 10537018 TI - Genes as gerontological variables: uniform genotypes. AB - Genetic conceptualizations and procedures have become integral to the conduct of research across the spectrum of life sciences, including gerontology, even when genetics is not the focus of inquiry. Among the research tools thus provided, one of the most basic is that of inbred strains. A close approximation to genetic uniformity is achieved by a sufficient number of successive generations of matings of relatives, and, once this near-uniformity is attained, the members of an inbred strain constitute a reference group relatively stable over time and available to diverse investigators. Different inbred strains possess different genotypes, so that numerous distinctive reference groups are available. The stability of these groups enhances prospects of replication-testing, and makes possible the focused accumulation of pertinent data. Phenotypic differences among strains identify particular groups that can be most appropriate for particular subsequent research objectives (and also provide ipso facto evidence of genetic influence on the phenotype). The very substantial advantages of the uniform genotypes provided by inbred strains (and by their F1 offspring) are purchased at the cost of limited generalizability of results and constraints on assessment of co-variation among variables. Uniform genotypes are, thus, not a tool for all purposes but must be seen as a powerful basic tool within an abundant genetic tool-kit. Particular research purposes will require use of more than one tool from the kit. PMID- 10537019 TI - Senescence-accelerated mouse (SAM): a biogerontological resource in aging research. AB - The senescence-accelerated mouse (SAM), consisting of 14 senescence-prone inbred strains (SAMP) and 4 senescence-resistant inbred strains (SAMR) has been under development since 1970 through the selective inbreeding of AKR/J strain mice donated by the Jackson laboratory in 1968, based on the data of the grading score of senescence, life span, and pathologic phenotypes. The characteristic feature of aging common to all SAMP and SAMR mice is accelerated senescence and normal aging, respectively. Furthermore, SAMP and SAMR strains manifest various pathobiological phenotypes which include such neurobiological phenotypes as deficits in learning and memory, emotional disorders, abnormal circadian rhythms, brain atrophy, hearing impairment, etc., and are often characteristic enough to differentiate the strains. Various efforts are currently being made using the SAM model to clarify the underlying mechanisms in accelerated senescence as well as the etiopathogenic mechanisms in age-associated pathobiologies. Genetic background and significance of SAM development are discussed. PMID- 10537020 TI - Age-related defects in lifespan and learning ability in SAMP8 mice. AB - Pertinent animal models of age-related learning deficiencies are required to elucidate the mechanism of age-related learning deficiencies and to develop novel therapeutic drugs for age-related diseases such as learning defects. Among many strains of accelerated senescence prone, senescence-accelerated mouse (SAM), SAMP8 mice have age-related defects in learning and cognitive abilities. We review recent findings on alterations in SAMP8 brain in neurochemical parameters related to neurotransmission and synaptic plasticity compared to those in SAMR1 brain as the control. In addition, we report the preventive effects of drugs on learning deficiencies in SAMP8 and discuss the usefulness of SAMP8 as an animal model of age-related learning deficiencies. PMID- 10537021 TI - Aging of blood-brain barrier and neuronal cells of eye and ear in SAM mice. AB - The SAMP, Senescence-Accelerated Mouse strains show senescence acceleration and age-associated pathological phenotypes similar to geriatric disorders seen in humans. Among these strains, SAMP8 mice show age-associated deficits in learning and memory. Histopathological studies revealed various neurodegenerative changes in the brain, including age-associated appearance of spongiform degeneration in the brain stem and of PAS-positive granular structures in the hippocampus. The blood-brain barrier (BBB) function of SAMP8 mice was also impaired with advancing age. The compromised BBB function in the olfactory bulb, the hippocampus and the pons of SAMP8 mice coincided with and might have been the cause of some morphological changes. Age-associated degeneration of receptor cells and ganglion neurons in the retina and cochlea also occurred in the SAM mice. Oxidative stress partly caused by mitochondrial dysfunction was detected and may be a cause of the neuronal cell degeneration. The SAM strains are useful tool in the attempt to understand the mechanisms of age-dependent neurodegeneration and to develop clinical interventions. PMID- 10537022 TI - Age-dependent cerebral atrophy and cognitive dysfunction in SAMP10 mice. AB - Findings obtained from a series of studies to characterize age-dependent changes in brain morphology and behavior of inbred SAMP10 mice are reviewed. Following apparently normal development, SAMP10 mice developed brain atrophy with advancing age. The neocortex was diffusely atrophic in aged SAMP10 mice, with the frontal cortex being most affected. The entorhinal cortex, amygdala, and nucleus accumbens were also atrophic. Other subcortical structures were mildly atrophic, but the hippocampus was not atrophic. Mild to moderate hypertrophic astrocytosis was seen in the atrophied regions. Alzheimer's type pathology was not seen. The cortical atrophy was due to both loss and shrinkage of neurons. Brain atrophy was not remarkable in normal aging control SAMR10 mice. In accordance with above morphological changes, SAMP10 mice developed cognitive impairments with advancing age that were demonstrated by poor performance in passive avoidance and conditional avoidance tasks. All of these features of SAMP10 mice were inherited. SAMP10, therefore, is a unique model of age-dependent, inherited cerebral atrophy with cognitive dysfunction. PMID- 10537023 TI - Developing mouse models of aging: a consideration of strain differences in age related behavioral and neural parameters. AB - Increased interest is emerging for using mouse models to assess the genetics of brain aging and age-related neurodegenerative diseases. Despite this demand, relatively little information is available on aging in behavioral or neuromorphological parameters in various mouse strains that are being used to create transgenic and null mutant mice. We review several issues regarding selection of appropriate strains as follows: (1) Does the behavioral parameter exhibit a significant age by strain interaction? (2) Do the strains differ in lifespan? (3) Are there potential intervening variables, such as strain-specific performance strategies or disease, in the behavioral task being investigated that would confound the desired conclusions? (4) Does the behavioral difference have an underlying neural correlate? In this context we present a conceptual model pertaining to the selection of mouse strains and behavioral parameters for genetic analyses. We also review the importance of applying stereological techniques for determining age-related structural changes in the mouse brain as well as the potential value of a database that would catalog this information. Thus, our intention is to underscore the growing importance of mouse models of brain aging and the concomitant need for additional information about mouse aging in general. PMID- 10537024 TI - Genes as gerontological variables: genetically heterogeneous stocks and complex systems. AB - In gerontological research utilizing animal models, a major general strategy has been the use of uniform genotypes of inbred strains or their F1 hybrids. These animal models provide standard reference groups that are of major importance in establishing a reliable data base on aging phenomena. There are limitations to their usage, however, particularly in respect to descriptions or evaluations of variances or of covariance relationships. For these purposes, genetically heterogeneous stocks have the advantage that phenotypic variance (and covariance) has a genetic as well as an environmental component. The advantages of genetic heterogeneity are best realized when the stock has been systematically derived (usually by intercrossing of inbred strains) and maintained by a mating scheme of sufficient size to minimize inbreeding. Genetically heterogeneous stocks are of particularly high potential value in the study of complex systems. Some examples of their use in a gerontological context are provided. PMID- 10537025 TI - Controlling caloric consumption: protocols for rodents and rhesus monkeys. AB - One approach for investigating biological aging is to compare control-fed animals with others restricted in calorie intake by 20% or more. Caloric restriction (CR) is the only intervention shown to extend the maximum lifespan of several invertebrates and vertebrates including spiders, fish, rats and mice. The capacity of CR to retard aging in nonhuman primates is now being explored. The rodent studies show that CR opposes the development of many age-associated pathophysiological changes, including changes to the brain and changes in learning and behavior. One goal of studying CR in rodent is to determine the mechanisms by which it retards aging to design interventions that duplicate those effects. The methods that we use for conducting CR studies on mice and rhesus monkeys are described. We employ procedures designed to achieve a high degree of caloric control for all animals in the study. As used in our studies, this control includes the following features: 1) animals are individually housed, and 2) all individuals in the control group eat the same number of calories (i.e., they are not fed ad lib). Although this method results in strict caloric control for all animals, there seems to be considerable procedural flexibility for the successful conduct of CR studies. PMID- 10537026 TI - Estimating age-related changes in psychomotor function: influence of practice and of level of caloric intake in different genotypes. AB - This article presents a discussion of some key considerations in the measurement of age-related changes in psychomotor function of mice. We illustrate that "standard" measures of psychomotor performance, such as running speed on a rotorod task, are highly sensitive to practice effects. Examples are cited in which failure to assess practice effects can influence conclusions regarding the magnitude and rate of change in psychomotor capacity as a function of age. A second set of examples is focused on estimating the effect of an experimental intervention, caloric restriction, on age-related changes in psychomotor performance. These examples show that psychomotor performance at a given age may vary directly, and reversibly, with the level of caloric intake. Independent of such reversible effects, the level of caloric intake can also modulate the rate of change in capacity as a function of age. It is concluded that reversible, short-term effects must be considered in estimating the effect of an experimental intervention on the rate of age-associated change in psychomotor function. PMID- 10537027 TI - Life-long diet restriction failed to retard cognitive aging in Fischer-344 rats. AB - Although the beneficial effects of diet restriction on longevity and the retardation of many somatic age-related processes are well established, the answer to the question of whether anti-aging effects of diet restriction extend to the brain and cognitive function remains unclear. In the present study, the effects of long-term dietary restriction (60% of ad-libitum calories) on an age related alteration of memory and sensorimotor function have been investigated in Fischer 344 male rats at four different ages: 6 months, 12 months, 18 months, and 24 months. A major drop in reference memory of DR and AL rats occurred at the age of 18 months. The performance deficits in working memory tasks were observed in both diet groups at the age of 24 months. These results indicate that diet restriction failed to provide protection against age-related deficits in memory. Although DR rats outperformed AL rats in sensorimotor tasks throughout the life span, the slope of the declining function in DR rats paralleled those of AL rats, suggesting that diet restriction failed to alter the rate of aging in sensorimotor performance, as well. PMID- 10537028 TI - Effects of moderate caloric restriction on cortical microvascular density and local cerebral blood flow in aged rats. AB - The present study was designed to assess the impact of moderate caloric restriction (60% of ad libitum fed animals) on cerebral vascular density and local cerebral blood flow. Vascular density was assessed in male Brown-Norway rats from 7-35 months of age using a cranial window technique. Arteriolar density, arteriole-arteriole anastomoses, and venular density decreased with age and these effects were attenuated by moderate caloric restriction. Analysis of local cerebral blood using [14C]iodoantipyrine indicated that basal blood flow decreased with age in CA1, CA3 and dentate gyrus of hippocampus; similar trends were evident in cingulate, retrosplenal, and motor cortex. Basal blood flow was increased in all brain regions of moderate caloric restricted old animals (compared to old ad libitum fed animals) and no differences were observed between ad libitum fed young and caloric restricted older animals. In response to a CO2 challenge to maximally dilate vessels, blood flow increased in young and old ad libitum fed animals, but a similar increase was not observed in caloric restricted old animals. We conclude that a decrease in cerebral vasculature is an important contributing factor in the reduction in blood flow with age. Nevertheless, vessels from young and old animals have the capacity to dilate in response to a CO2 challenge and, after CO2, no differences are observed between the two age-groups. These results are consistent with the hypothesis that aged animals fail to adequately regulate local cerebral blood flow in response to physiological stimuli. Moderate caloric restriction increases microvascular density and cerebral blood flow in aged animals but tissues exhibit little or no increase in blood flow in response to CO2 challenge. The cause of this deficient response may indicate that vessels are maximally dilated in aged calorically restricted animals or that they fail to exhibit normal regulatory control. PMID- 10537029 TI - Progress toward valid transgenic mouse models for Alzheimer's disease. AB - A transgenic mouse model for Alzheimer's disease (AD) should mimic the age dependent accumulation of beta-amyloid plaques, neurofibrillary tangles, neuronal cell death as well as display memory loss and behavioral deficits. Age-dependent accumulation of A beta deposits in mouse brain has been achieved in mice overexpressing mutant alleles of the amyloid precursor protein (APP). In contrast, mice bearing mutant alleles of the presenilin genes show increased production of the A beta42 peptide, but do not form amyloid deposits unless mutant alleles of APP are also overproduced. Furthermore, the onset of A beta deposition is greatly accelerated, paralleling the involvement of presenilins in early onset AD. Studies of APP and presenilin transgenic mice have shown 1) the absence of a requirement for a maturation step in dense core plaque formation, 2) evidence that beta-amyloid deposition is directed by regional factors, and 3) behavioral deficits are observed before A beta deposition. Crosses of APP transgenic mice with mice modified for known AD risk factors and "humanizing" the mouse may be necessary for complete replication of AD. PMID- 10537030 TI - Infrastructure for research on aging rodents: need for regional facilities to support transgenic studies on aging. AB - I propose that North American investigators who depend largely on individual external grants would benefit from the creation of regional facilities designed to support studies of aging in transgenic models. In general, academic institutions cannot achieve the most cost effectiveness because their facilities are not designed for industrial-scale cost efficiencies. Regional facilities would also increase the reproducibility of experiments. Transgenic studies of prospective genes that modify the risk of Alzheimer and other outcomes of aging could be carried out under standard conditions of nutrition and caging density, which can influence brain structure and behavior. The regional labs should have a strong Pathology Core, which would go far beyond the resources usually available to academic researchers. A national system of regional facilities with Core labs for characterizing genetics, hormones, and pathology would greatly accelerate progress. PMID- 10537031 TI - Exotic mice as models for aging research: polemic and prospectus. AB - Most gerontological research using rodent models employs inbred strains, or F1 hybrids derived from them, rather than populations of genetically heterogeneous individuals. This study presents the argument that reliance on genetically homogeneous rodents, though sanctioned by tradition, may not be ideal for many sorts of investigations, and that use of heterogeneous mice and rats would allow researchers to reach robust conclusions that were less likely to reflect strain specific idiosyncrasies. Segregating stocks, bred by backcross, F2 cross, or four way cross procedures, would be an improvement over inbred and F1 stocks, providing inexpensive, arbitrarily large, and reproducible populations of genetically diverse test subjects. These stocks would not, however, recapture allelic variations that are likely to have been lost when wild-trapped mice and rats are selected inadvertently over dozens of generations for breeding success in laboratory conditions. Development of specific pathogen free stocks from wild trapped progenitors particularly from populations selected for relevant evolutionary history and physiological characteristics, may be of great value for analysis of aging and late-life pathophysiology. PMID- 10537032 TI - Exotic mice as models for aging research: polemic and prospectus by R. Miller et al. AB - Miller and colleagues present a polemic against uniform genotypes in gerontological research and a paean for genetically heterogeneous populations. Both on simple sampling considerations, and because of selective loss of alleles through inbreeding depression, inbred strains are idiosyncratic, and the lack of genetic variance within strains limits their utility in examining relationships among phenotypes. Heterogeneous stocks, by contrast, are not impaired by inbreeding depression, and the presence of genetic and environmental variance provides for effective assessment of correlations among variables. Those assembled by intermating of inbred strains are replicable in terms of gene pool parameters, even if no individual can ever be replicated, and those derived from wild-trapping may include alleles absent for whatever reason from current laboratory strains. This author concurs that there are limitations on research with uniform genotypes and advantages of heterogeneous populations. However, for specific purposes, the uniformity and stability of inbred strains are extremely valuable attributes, and heterogeneous stocks have their own limitations. Researchers should select the animal model most appropriate for their specific purpose. PMID- 10537033 TI - Warning: the use of heterogeneous mice may seriously damage your research. AB - Genetically heterogeneous (GH) mice and rats continue to be widely used in research even though the case for using isogenic strains has been argued repeatedly. The paper by Miller et al. in this issue appears to be the only one in the last 22 to attempt a scientific justification for the continued use of (a limited subset of) GH stocks. However, although they are to be commended for bravery, they fail to make their case. GH stocks represent poor material for controlled studies because genetic heterogeneity normally leads to phenotypic variability and a decline in experimental sensitivity. To counter this argument, Miller et al. claim that phenotypic variability may actually be smaller in GH animals than in their isogenic parents. Were this so (e.g., all mice being short lived, small, and aggressive), it is difficult to see how the use of such a stock could increase the generality of research results based on it, as claimed by Miller et al. Isogenic strains are a vital, proven, and powerful resource for biomedical research, and should be used in preference to GH stocks by all scientists who use laboratory rodents. PMID- 10537034 TI - Consensus report of the Working Group on: molecular and biochemical markers of Alzheimer's disease. PMID- 10537035 TI - Selective reduction in A2 adenosine receptor desensitization following antisense induced suppression of G protein-coupled receptor kinase 2 expression. AB - Phosphorylation of G protein-coupled receptors (GPCRs) by GPCR kinases (GRKs) is considered to play a critical role in the desensitization of responses mediated by these receptors. To explore the role of GRK2 in A2 adenosine receptor desensitization, we attempted to reduce specifically GRK2 expression in NG108-15 cells by stable transfection with an antisense rat GRK2 cDNA sequence. This yielded up to a 69% loss of GRK2 when compared with plasmid-transfected control cells, which correlated with a reduction in kinase activity when measured by the ability of cell lysates to promote light-dependent phosphorylation of rhodopsin. Levels of GRK3 were the same in antisense and plasmid-transfected controls. On addition of the A2 adenosine receptor agonist 5'-(N-ethylcarboxamido)adenosine, cyclic AMP accumulation was greater in GRK2 antisense cells as compared with plasmid control cells. In contrast, cyclic AMP accumulation via agonist stimulation of either IP-prostanoid or secretin receptors or by addition of forskolin was not significantly different among all clones examined. The increase in A2 adenosine receptor response could not be explained by changes in A2A adenosine receptor expression, as assessed by ligand binding experiments with the radioligand 2-3H-labelled 4-[2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3 a][1,3,5]triazin-5-++ +ylamino]ethyl]phenol ([3H]ZM241385). These data show for the first time a direct correlation between expression of GRK2 and desensitization of natively expressed A2 adenosine receptors in intact cells, suggesting that GRK2 plays a major role in the regulation of these receptors. Key Words: G protein-coupled receptor kinase-G protein-coupled receptor-Antisense NG108-15 cells-A2 adenosine receptors-Desensitization. PMID- 10537036 TI - The 5' untranslated regions of the rat A2A adenosine receptor gene function as negative translational regulators. AB - The rat A2A adenosine receptor (A2A-R) gene contains two promoters, P1 and P2, which produce transcript 1 and transcript 2, respectively. These transcripts differ in the lengths of their 5' untranslated regions (5'UTR1: 514 bp, initiated from P1; 5'UTR2: 221 bp, initiated from P2) but encode the same protein. In the present study, we demonstrate that transcript 2 is present in various tissues at different levels, whereas transcript 1 is found only in the striatum. In the striatum, the level of transcript 2 is approximately 300-fold higher than that of transcript 1. The 5'UTR of both transcripts suppresses the expression of A2A-R and a firefly luciferase reporter gene at the translational level; this suppression is not observed after mutational inactivation of an "out-of-frame" upstream AUG codon. Translational suppression by the 5'UTR was also confirmed in cells using a bicistronic strategy. Collectively, these data suggest that P2 is the major promoter of the rat A2A-R gene. The 5'UTR of the rat A2A-R gene exerts an inhibitory effect on translation by an upstream open reading frame. Because the 5'UTR of the A2A-R gene possesses strong interspecies homology, translational suppression may be a general mechanism by which the expression of the A2A-R gene is regulated. PMID- 10537037 TI - Effects of cyclic AMP on components of the cell cycle machinery regulating DNA synthesis in cultured astrocytes. AB - Cyclic AMP is a second messenger for various hormones that inhibits cell multiplication and DNA synthesis in cultured astrocytes. We examined the effects of increasing intracellular cyclic AMP on the catalytic (cdks) and regulatory (cyclins and ckis) components of cyclin-dependent protein kinases, which regulate progression of the cell cycle before completion of DNA synthesis, in primary cultured astrocytes and in an astrocytic cell line C.LT.T.1.1. The amount of cdk4 changed little during the cell cycle and was not affected by cyclic AMP. There was little cdk1 and cdk2 in quiescent cells, and their expression increased during the G1-S phases. Cyclic AMP strongly inhibited cdk1 and cdk2 expression. Transforming growth factor beta also inhibited cdk1 expression in primary astrocytes. Cyclic AMP did not affect the two ckis p27KIP1 and p21CIP1. There was little cyclin D1 in quiescent cells, but it increased during the G1 phase and was reduced by cyclic AMP. Cyclin E increased during the G1-S phases and was not affected by cyclic AMP in primary astrocytes. The amount of cyclin A was low in quiescent cells and increased during the G1-S phases. Expression of its mRNA and protein was inhibited by cyclic AMP. The protein kinase activities associated with complexes of cyclins and cdks were increased by growth factors and prevented by cyclic AMP. We conclude that cyclic AMP inhibits progression of the cell cycle in astrocytes at least by preventing the expression of the regulatory subunits, cyclins D1 and A, and catalytic subunits, cdk1 and cdk2, of cyclin-regulated protein kinases. Key Words: Cyclin-dependent protein kinases-Glial cells-Cyclic AMP. PMID- 10537038 TI - Involvement of intronic sequences in cell-specific expression of the peripherin gene. AB - Peripherin is an intermediate filament protein expressed in restricted populations of neurons. Our previous study of the chromatin structure of the mouse peripherin gene in cells that do or do not express peripherin suggested that the region located between -1,500 and +800 bp of the gene could be involved in its cell specificity. In the present work, we performed an in vitro functional analysis of the 5' flanking region of the mouse peripherin gene and observed that this region up to 9 kb contained both enhancer and inhibiting activities; however, it was insufficient to achieve a complete extinction of reporter gene expression in peripherin-negative cells. Furthermore, analysis of the first three introns with the 5' flanking sequences of the gene showed that intron I greatly increased specificity of the gene expression. Intron I also conferred the same properties to thymidine kinase heterologous promoter. DNase I footprinting experiments performed with intron I revealed at least two protected regions (Inl A and Inl B). Inl A encompasses an AP-2-like binding site that interacted with both neuroblast and fibroblast nuclear factors, as well as with the recombinant AP-2alpha protein. However, gel shift experiments suggested that the interacting nuclear factors are distinct from AP-2alpha itself and probably belong to the AP 2 family. Inl B perfectly matched the consensus binding site for Sp1 and specifically interacted with nuclear protein factors that showed the same binding properties as the Sp1 family members. Fine deletion analysis of intron I indicated that the Inl A element alone is responsible for its enhancing properties, whereas a region located between +789 and +832 gives to intron I its silencer activity. PMID- 10537040 TI - Antisense knockdown of glutamate transporters alters the subfield selectivity of kainate-induced cell death in rat hippocampal slice cultures. AB - Organotypic rat hippocampal slice cultures were used to study the role of excitatory amino acid transporters (EAATs) in kainate-induced cell death. Expression of the neuronal (EAAT3) or glial (EAAT2) transporters was inhibited with antisense phosphothioate oligonucleotides, and cytotoxicity was assessed with propidium iodide uptake. In control cultures, a concentration of 10 microM kainate was more cytotoxic in CA3 than in CA1. Treatment for 24 h with EAAT3 antisense oligonucleotide decreased kainate toxicity in CA1 but had an opposite effect in CA3. Neither antisense oligonucleotide to EAAT2 nor mismatch oligonucleotide to EAAT3 decreased kainate toxicity in CA1. Immunoblotting with affinity-purified antibodies showed that EAAT3 antisense oligonucleotide decreased selectively EAAT3 but not EAAT2 protein levels, and vice versa. NMDA was more cytotoxic in CA1 than in CA3, and antisense oligonucleotides to either EAAT3 or EAAT2 did not decrease the NMDA effect in CA1 or CA3. Dihydrokainate and DL-threo-beta-hydroxyaspartic acid were more cytotoxic in CA1 than in CA3, suggesting that the higher vulnerability of CA3 to kainate was not the result of its activity as transporter blocker. We conclude that glutamate transporters differentially regulate excitotoxicity in different hippocampal subfields. PMID- 10537039 TI - Growth arrest and spontaneous differentiation are initiated through an autocrine loop in clonally derived Schwann cells by alpha1-procollagen I C-propeptide. AB - Schwann cells cloned from rat sciatic nerve survive and display self-induced growth suppression, or undergo spontaneous apoptosis, on long-term serum-free subconfluent culture. Strain SCL4.1/F7 sustained the capacity to growth arrest for up to 40 generations. A soluble activity transmitted between neighbouring cells of this strain suppresses DNA synthesis within three cell cycles. Autocrine Schwann cell growth-inhibitory factor (SGIF) operates during the G1 phase of the cell cycle, overcomes the mitogenic action of Schwann cell/serum-associated (platelet-derived growth factor-BB) and axon-associated (axolemma-enriched fraction) stimuli in serum-free conditions, and suppresses DNA synthesis in sciatic nerve Schwann cell cultures in a stage-specific manner. A 35-kDa protein with N-terminal sequence and approximate molecular mass of the C-propeptide of rat alpha1-procollagen I makes a major contribution to SGIF. Growth suppression in the SCL4.1/F7 strain is mediated by the ras/extracellular signal-regulated kinase pathway, is accompanied by down-regulation of erbB2/erbB3 and of tetraethylammonium-sensitive K+ currents, and is followed by transition of cells within 5-10 days from O4+, p75 nerve growth factor receptor (p75NGF-R)+, glial fibrillary acidic protein (GFAP)+ to O4+, p75NGF-R-, GFAP-, periaxin+ phenotypes. Oct-6/SCIP mRNA is present in both proliferating and growth-arrested SCL4.1/F7 cells. These results demonstrate an autocrine/ paracrine loop for the growth arrest of clonally derived Schwann cells in the absence of axons linked in part to the metabolism of collagen. Schwann cells thus appear to self-regulate growth in a negative as well as a positive direction through characterized molecular mechanisms and signal pathways. PMID- 10537041 TI - CREB phosphorylation promotes nerve cell survival. AB - The cyclic AMP-responsive element binding protein (CREB) is a posttranslationally activated transcription factor that has been implicated in numerous brain functions including cell survival. In this study we investigated whether CREB overexpression using transient transfection of a pAAV/CMV-CREB plasmid altered neuronal cells' susceptibility to apoptosis. We found that elevated CREB protein inhibited apoptosis induced by okadaic acid. At least part of this effect is critically dependent on prolonged Ser133 phosphorylation, as a directed mutation at this site decreased CREB-induced protection. These results suggest that CREB is a survival factor for neuronal cells and that treatments aimed at augmenting CREB phosphorylation in the brain may be neuroprotective. PMID- 10537042 TI - Modulation of tau phosphorylation within its microtubule-binding domain by cellular thiols. AB - Tau is a microtubule-stabilizing protein that is functionally modulated by alterations in its phosphorylation state. Because phosphorylation regulates both normal and pathological tau functioning, it is of importance to identify the signaling pathways that regulate tau phosphorylation in vivo. The present study examined changes in tau phosphorylation and function in response to modulation of cellular thiol content. Treatment of cells with phenylarsine oxide, which reacts with vicinal thiols, selectively increased tau phosphorylation within its microtubule-binding domain, at the non-Ser/Thr-Pro sites Ser262/356, while decreasing tau phosphorylation at Ser/ Thr-Pro sites outside this region. This increase in tau phosphorylation correlated with a decrease in the amount of tau associated with the cytoskeleton and decreased microtubule stability. Phenylarsine oxide-induced tau phosphorylation was inhibited by oxidants and by the protein kinase inhibitor staurosporine. Although staurosporine completely eliminated the increase in tau phosphorylation at Ser262/356, as detected by immunostaining with 12E8, it had a comparatively minor effect on the changes in tau localization induced by phenylarsine oxide. The results suggest that regulation of cellular thiols is important for modulating tau phosphorylation and function in situ. Additionally, although phosphorylation of Ser262/356 decreases tau's interaction with the cytoskeleton, phosphorylation of these residues alone is not sufficient for the phenylarsine oxide-induced changes in tau localization. PMID- 10537043 TI - Inhibition of the activity of a neuronal kappaB-binding factor by glutamate. AB - Activation of transcription factors with affinity for kappaB enhancers is generally correlated with enhanced survival of neurons. In an apparent exception, excitotoxic concentrations of glutamate have been reported to elevate the activity of one such factor, nuclear factor-kappaB (NF-kappaB). Our data indicate that the constitutive neuronal kappaB-binding factor (NKBF) is distinct from bona fide NF-kappaB (RelA/p50 heterodimer). Therefore, we analyzed glutamate's effects on KB-binding activity in highly enriched primary neuronal cultures and in mixed neuron/glia cocultures. Electrophoretic mobility shift assays indicated that a 30 60-min exposure to 50-500 microM glutamate reduced NKBF activity by as much as 70%. Subtoxic doses of glutamate had little or no effect on this DNA-binding activity. Selective antagonists of either NMDA or AMPA [(RS)-alpha-amino-3 hydroxy-5-methyl-4-isoxazolepropionate]/kai nat e receptors inhibited the influence of glutamate on NKBF activity. The effect of glutamate was mimicked by calcium ionophore, and it was blocked by lowering extracellular calcium concentrations or by cyclosporin A. Bona fide NF-kappaB was found only in cocultures containing significant numbers of glia, where it could be activated by glutamate. These data suggest that the primary influence of excitatory amino acids on neuronal KB-binding activity is an inhibitory one, strengthening the correlation between this transcriptional parameter and neuronal survival. PMID- 10537044 TI - Isolation of a potential neural stem cell line from the internal capsule of an adult transgenic rat brain. AB - A thermosensitive mutation of simian virus 40 large T antigen (LTA) gene, the tsA58 gene, was cloned downstream of the 6-kbp neurofilament light chain promoter in pPOLYIII and injected into the pronucleus of fertilised oocytes of Sprague Dawley rats to develop a strain harbouring six copies of the transgene. Immunocytochemical staining of hemizygous adult tissues with antibodies to the C terminus of LTA showed that the inactive form of LTA was expressed only in the fibres of the internal capsule and in the choroid plexus of the brain. Culturing the former region at 33 degrees C, the permissive temperature for LTA, yielded a cell line, NF2C, which produced active LTA and grew at 33 degrees C but which produced only inactive LTA and eventually died at the non-permissive temperature of 39 degrees C. This clonal cell line was heterogeneous at 33 degrees C, producing the precursor neuronal cell marker nestin and the glial-specific markers glial fibrillary acidic protein, vimentin and S100A1, as well as weakly producing the neuronal cell markers 68-kDa neurofilament protein (NF68) and microtubule-associated protein 2 (MAP2) in different subpopulations of cells. However, at 39 degrees C, the cells produced dendritic, neuronal-like processes and elevated levels of NF68 and MAP2, as well as the neuronal markers synaptophysin, neurone-specific enolase, and low levels of tau, all determined by western blotting and immunofluorescent staining. Basic fibroblast growth factor enhanced the growth of the cells at 33 degrees C but also enhanced the formation of dendritic neuronal-like processes at 39 degrees C. It is suggested that NF2C represents a potential stem cell line from adult brain that expresses precursor and glial cell markers at 33 degrees C but undergoes partial differentiation to a neuronal cell phenotype at 39 degrees C. PMID- 10537045 TI - Tau is modified by tissue transglutaminase in situ: possible functional and metabolic effects of polyamination. AB - Tissue transglutaminase (tTG) is up-regulated in Alzheimer's disease brain and localizes to neurofibrillary tangles with the tau protein. Tau is an in vitro tTG substrate, being cross-linked and/or polyaminated. Further, the Gln and Lys residues in tau that are modified by tTG in vitro are located primarily within or adjacent to the microtubule-binding domains. Considering these and other previous findings, this study was carried out to determine if tau is modified in situ by tTG in human neuroblastoma SH-SY5Y cells, and whether tTG-catalyzed tau polyamination modulates the function and/or metabolism of tau in vitro. For these studies, SH-SY5Y cells stably overexpressing tTG were used. tTG coimmunoprecipitated with tau, and elevating intracellular calcium levels with maitotoxin resulted in a 52 +/- 4% increase in the amount of tTG that coimmunoprecipitated with tau. The increase in association of tTG with tau after treatment with maitotoxin corresponded to a coimmunolocalization of tTG, tTG activity, and tau in the cells. Further, tau was modified by tTG in situ in response to maitotoxin treatment. In vitro polyaminated tau was significantly less susceptible to micro-calpain proteolysis; however, tTG-mediated polyamination of tau did not significantly alter the microtubule-binding capacity of tau. Thus, tau interacts with and is modified by tTG in situ, and modification of tau by tTG alters its metabolism. These data indicate that tau is likely to be modified physiologically and pathophysiologically by tTG, and tTG may play a role in Alzheimer's disease. PMID- 10537046 TI - Expression of the vesicular acetylcholine transporter, proteins involved in exocytosis, and functional calcium signaling in varicosities and soma of a murine septal cell line. AB - The expression and localization of the vesicular acetylcholine transporter in a septal cell line, SN56, were investigated. Immunoprecipitation and immunoblot analysis of postnuclear supernatants indicated that this cell line expresses reasonable amounts of the transporter. Immunofluorescence and confocal microscopy experiments showed that the vesicular transporter is present in varicosities and also in the cell body of differentiated cells. Varicosities have the potential to be functional sites of transmitter release because they responded to depolarization with calcium influx through voltage-gated calcium channels and expressed the synaptic proteins synaptotagmin, SV2, synaptophysin, and a subunit of P/Q calcium channels. In the soma of SN56 cells, the transporter immunoreactivity was similar to that for synaptotagmin, and it colocalized with synaptophysin, but it did not colocalize with SV2. Labeling for SV2 appeared prominently in a defined perinuclear structure, whereas the two former proteins were widely distributed in the soma, where several endocytic compartments could be identified with the vital dye FM4-64. These data suggest that distinct synaptic vesicle proteins exist in different subcellular compartments, and consequently they may follow distinct pathways in neurites before reaching sites of transmitter storage and release in SN56 cells. PMID- 10537047 TI - Serotonin N-acetyltransferase mRNA levels in photoreceptor-enriched chicken retinal cell cultures: elevation by cyclic AMP. AB - Serotonin N-acetyltransferase (AA-NAT; arylalkylamine N-acetyltransferase; EC 2.3.1.87) is a key regulatory enzyme in the biosynthesis of melatonin. Previous studies have shown that the activity of this enzyme in the chicken retina is regulated by a cyclic AMP-dependent mechanism. In the present report, we investigated whether cyclic AMP can regulate the levels of AA-NAT mRNA in photoreceptor-enriched chick retinal cell cultures. AA-NAT mRNA levels were elevated by acute treatment with cyclic AMP protagonists, including forskolin; this response was blocked by H-89, a selective inhibitor of cyclic AMP-dependent protein kinase. Forskolin did not alter the rate of disappearance of AA-NAT mRNA in actinomycin D-treated cells, suggesting that cyclic AMP enhances transcription of the AA-NAT gene. Forskolin-induced elevation of AA-NAT mRNA levels was enhanced by cycloheximide, which decreased the degradation of the transcript in cells treated with actinomycin D. These studies indicate that the abundance of AA NAT mRNA is regulated in part through a cyclic AMP-dependent mechanism. PMID- 10537048 TI - CEP-1347 (KT7515), an inhibitor of JNK activation, rescues sympathetic neurons and neuronally differentiated PC12 cells from death evoked by three distinct insults. AB - The c-Jun N-terminal kinase signaling cascade appears to play a role in some cases of cell death, including neuronal apoptosis. CEP-1347 (KT7515), an indolocarbazole of the K252a family, blocks this stress signaling cascade and promotes survival. Here, we used CEP-1347 to probe whether neuronal death pathways activated by distinct insults also possess elements in common. Cultured rat sympathetic neurons and neuronally differentiated PC12 cells were induced to die by withdrawal of nerve growth factor, exposure to ultraviolet irradiation, or subjection to oxidative stress. In each case, death was prevented by 100-200 nM CEP-1347. Moreover, in each of these death paradigms, c-Jun N-terminal kinase 1 activity in neuronally differentiated PC12 cells was elevated by two- or threefold, and this increase was totally blocked by CEP-1347 at concentrations that promoted survival. In contrast, 200 nM CEP-1347 did not block death due to serum withdrawal from undifferentiated PC12 cells or to activation of Fas in Jurkat T cell cultures, even though in each case c-Jun N-terminal kinase 1 activation occurred and was inhibited by CEP-1347. These observations suggest that some but not all death pathways triggered by different insults can include a common mechanistic component, a likely candidate for which is activation of the c Jun N-terminal kinase signaling cascade. PMID- 10537049 TI - Myelin-associated oligodendrocytic basic protein mRNAs reside at different subcellular locations. AB - The mRNAs for two myelin proteins, myelin basic protein (MBP) and myelin associated oligodendrocytic basic protein (MOBP)-81A, are uniquely located at sites where myelin sheaths are assembled. Here, we use subcellular fractionation to show that four MOBP mRNAs, like MBP mRNA, are located at sites of myelin sheath assembly, and that three other MOBP mRNAs are located in oligodendrocyte soma. The MOBP-81 protein is found in myelin and in another subcellular fraction, whereas other myelin proteins, including MBP, 2',3'-cyclic nucleotide 3' phosphodiesterase, and myelin-associated glycoprotein, are largely restricted to myelin. Different MBP mRNAs are generated by alternative splicing. All of them contain an RNA transport sequence (RTS) that directs them to sites in oligodendrocytes, where myelin sheaths are assembled. Consequently, all are enriched in myelin. After fractionation, four MOBP mRNAs, MOBP-71, MOBP-81A, MOBP 99, and MOBP-169 (identified in this study), are enriched in myelin. These mRNAs contain a common exon, exon 8b, which has a nucleotide sequence that is similar to MBP mRNA RTS. This sequence likely directs these mRNAs to sites of myelin sheath assembly. Three other MOBP mRNAs, MOBP-69, MOBP-81B, and MOBP-170, lack this exon. Their subcellular distribution indicates that they are largely retained in oligodendrocyte soma. We conclude that the distribution of MOBPs in oligodendrocytes is strongly influenced by alternative splicing of the corresponding mRNAs. PMID- 10537050 TI - Phosphorylation and desensitization of neurokinin-1 receptor expressed in epithelial cells. AB - A rat kidney epithelial cell line expressing the rat neurokinin-1 receptor (NK-1 R) was used to investigate the relationship between receptor phosphorylation and desensitization. Substance P (SP) maximally stimulated cellular inositol 1,4,5 trisphosphate (IP3) production 14-fold within 3 s, after which cellular IP3 levels rapidly diminished to near basal levels in the continuing presence of SP. SP also caused concentration-dependent phosphorylation of the NK-1R, and this effect was blocked by a receptor antagonist. Stimulation with 100 nM SP for as little as 2 s resulted in 90% desensitization of the receptor to restimulation by SP, and near-maximal receptor phosphorylation was observed at 5 s. Receptor desensitization was not affected by agents that affect protein kinase A. Phorbol 12-myristate 13-acetate (PMA) also caused phosphorylation and desensitization of the receptor but with slower kinetics and to a lesser extent than SP. PMA- but not SP-induced NK-1 R desensitization and phosphorylation were abolished by the protein kinase C inhibitor bisindolylmaleimide 1. The concentration-response curves for SP-stimulated IP3 signaling and desensitization were similar, but the curve for NK-1R phosphorylation was shifted to the right and was steeper, suggesting that the relationship between desensitization and phosphorylation is complex. These results show that both rapid homologous and rapid heterologous NK 1R desensitizations may be mediated by receptor phosphorylation but occur via distinct mechanisms with different kinetics and efficacies. PMID- 10537051 TI - Attenuation of focal adhesion kinase signaling following depletion of agonist sensitive pools of phosphatidylinositol 4,5-bisphosphate. AB - The effect of phosphoinositide depletion on focal adhesion kinase (FAK) signaling was investigated in two neuronal cell lines. Treatment of either SH-SY5Y neuroblastoma cells or PC12 cells with wortmannin, at a concentration that inhibits phosphatidylinositol 4-kinase activity, led to a selective depletion of phosphatidylinositol 4-phosphate without significantly altering phosphatidylinositol 4,5-bisphosphate (PIP2) content. An enhanced tyrosine phosphorylation of FAK elicited by agonist occupancy of phospholipase C-coupled receptors (muscarinic cholinergic in SH-SY5Y neuroblastoma or bradykinin in PC12 cells) was blocked completely by wortmannin. Under the above conditions, phosphoinositide resynthesis was prevented, and as a consequence, receptor stimulation led to a marked depletion of PIP2. In contrast, the increased tyrosine phosphorylation of FAK elicited by agents that do not activate phospholipase C (phenylarsine oxide, lysophosphatidic acid, or phorbol ester) persisted in the presence of wortmannin. However, the ability of these agents to elicit an increase in FAK phosphorylation was also prevented if PIP2 was depleted by activation of a phospholipase C-coupled receptor in the presence of wortmannin. The results suggest that agonist-sensitive pools of PIP2 must be maintained for FAK signaling to occur in response to a mechanistically diverse range of stimuli. PMID- 10537052 TI - Regulation of growth inhibitory factor expression by epidermal growth factor and interleukin-1beta in cultured rat astrocytes. AB - Growth inhibitory factor (GIF) is highly expressed in the CNS under physiological conditions, but its expression is reduced in neurodegenerative diseases, such as Alzheimer's disease. The results of this study show that the levels of GIF and GIF mRNA were not influenced by neuroglial interactions. GIF was highly expressed in confluent astrocytes, but the expression was down-regulated in low-density growing astrocytes. A high level of GIF was not observed in serum-starved low density cultures. These findings suggest that GIF is a quiescent state-specific protein and that two different mechanisms may exist for the cells to enter the quiescent state. Among interleukin-1beta (IL-1beta), fibroblast growth factor-2, epidermal growth factor (EGF), amyloid beta1-42, and 50% O2, only EGF and IL 1beta altered the level of GIF in confluent astrocytes: EGF increased both GIF mRNA and protein, and IL-1beta decreased GIF mRNA, but did not alter GIF protein. Kinetic analysis of the GIF mRNA level revealed the biphasic regulation of GIF mRNA expression by IL-1beta, i.e., a transient up-regulation followed subsequently by down-regulation, explaining in part the discrepancy between the levels of GIF mRNA and protein in astrocytes treated with IL-1beta. PMID- 10537053 TI - Dexamethasone regulation of P-glycoprotein activity in an immortalized rat brain endothelial cell line, GPNT. AB - The blood-brain barrier (BBB) plays an important role in controlling the passage of molecules from the blood to the extracellular fluid environment of the brain. The multidrug efflux pump P-glycoprotein (P-gp) is highly expressed in the luminal membrane of brain capillary endothelial cells, thus forming a functional barrier to lipid-soluble drugs, notably, antitumor agents. It is of interest to develop an in vitro BBB model that stably expresses P-gp to investigate the mechanisms of regulation in expression and activity. The rat brain endothelial cell line, GPNT, was derived from a previously characterized rat brain endothelial cell line. A strong expression of P-gp was found in GPNT monocultures, whereas the multidrug resistance-associated pump Mrp1 was not expressed. The transendothelial permeability coefficient of the P-gp substrate vincristine across GPNT monolayers was close to the permeability coefficient of bovine brain endothelial cells cocultured with astrocytes, a previously documented in vitro BBB model. Furthermore, the P-gp blocker cyclosporin A induced a large increase in apical to basal permeability of vincristine. Thus, P gp is highly functional in GPNT cells. A 1-h treatment of GPNT cells with dexamethasone resulted in decreased uptake of vincristine without any increase in P-gp expression. This effect could be mimicked by protein kinase C (PKC) activation and prevented by PKC inhibition, strongly suggesting that activation of P-gp function may involve a PKC-dependent pathway. These results document the GPNT cell line as a valuable in vitro model for studying drug transport and P-gp function at the BBB and suggest that activation of P-gp activity at the BBB might be considered in chemotherapeutic treatment of cancer patients. PMID- 10537055 TI - Effects of cholecystokinin peptides and GV 150013, a selective cholecystokininB receptor antagonist, on electrically evoked endogenous GABA release from rat cortical slices. AB - The effects of cholecystokinin (CCK) agonists and antagonists on spontaneous and electrically evoked endogenous GABA release from rat cerebral cortex slices were evaluated. Neither the nonselective and CCK(B)-selective receptor agonists CCK-8S (3-1,000 nM) and CCK-4 (3-1,000 nM), respectively, nor the selective CCK(B) and CCK(A) receptor antagonists GV 150013 (3-30 nM) and L-364,718 (10-100 nM), respectively, significantly affected spontaneous GABA release. CCK-8S (1-1,000 nM) and CCK-4 (1-1,000 nM) increased the electrically (5 and 10 Hz)-evoked GABA release. On the contrary, GV 150013 (10 and 30 nM) significantly decreased the electrically evoked GABA release only when the slices were stimulated at the higher 10 Hz frequency. The CCK-8S- and CCK-4-induced increases in electrically evoked GABA release were counteracted by GV 150013, but not by L-364,718. Furthermore, GV 150013 at 3 nM shifted to the right the CCK-4 concentration response curve, whereas at the higher 10 nM concentration it dramatically flattened the curve. Finally, in cortical slices obtained from rats chronically treated with GV 150013, the concentration-response curve of CCK-4 was shifted to the left and the peak effect of the peptide was significantly higher than that observed in naive animals. These results suggest that CCK increases electrically evoked, but not spontaneous, endogenous GABA release from rat cortical slices, possibly by activating local CCK(B) receptors. In addition, chronic treatment with the novel CCK(B) receptor antagonist GV 150013 leads to an enhanced responsiveness of cortical slices to CCK-4 application. PMID- 10537054 TI - Thyroid hormone-induced maturation of astrocytes is associated with the expression of new variants of vimentin and their phosphorylation. AB - The role of thyroid hormone (TH)-induced vimentin expression in promoting the maturation of astrocytes has been investigated. Comparative immunocytochemical staining with vimentin antibody demonstrated distinctly different patterns of expression of vimentin during the progressive maturation of astrocytes of normal and TH-deficient astrocyte cultures. [35S]Methionine labeling of cells showed a significant decline in the level of vimentin in the hypothyroid cultures at all ages from day 5 to 20. Western blot analysis suggested that the maturation of normal astrocytes was associated with the appearance of several acidic and basic variants of vimentin, whose expression was significantly delayed or reduced in the TH-deficient astrocytes. In normal astrocyte cultures, two acidic variants of vimentin, one of similar molecular weight and the other of lower molecular weight, were found to be phosphorylated during the final transformation of the epithelioid form into the mature stellate form. None of these phosphorylated forms was evident in the TH-deficient astrocytes. This was further confirmed by negative immunocytochemical staining of 5- to 18-day-old cultures of TH-deficient astrocytes with antibody TM71, specific to phosphorylated vimentin, compared with age-matched normal astrocytes, which displayed consistent positive staining. The overall results indicate that TH-induced expression of phosphorylated forms of vimentin may be essential for the intracellular organization of the cytoskeleton in a way conducive to the morphological maturation of astrocytes. PMID- 10537057 TI - Hypoxia-ischemia in perinatal rat brain induces the formation of a low molecular weight isoform of striatal enriched tyrosine phosphatase (STEP). AB - Protein tyrosine phosphatases play a critical role in controlling tyrosine phosphorylation levels of proteins. Ischemia induces changes in tyrosine phosphorylation. As part of our investigations of the mechanisms responsible for these changes, we studied the effects of cerebral hypoxia-ischemia in 7-day-old (P7) and P21 rat brains on expression of the STEP (striatal enriched phosphatase) family of protein tyrosine phosphatases. P7 and P21 rats were subjected to unilateral hypoxia-ischemia, and brains were analyzed at various intervals of recovery for the presence of STEP. Hypoxia-ischemia induced the formation of a low Mr isoform of STEP, STEP33, in the ipsilateral (damaged) hemisphere but not in the contralateral (undamaged) side. STEP33 produced as a result of ischemia was located exclusively in the cell soluble fraction. In P21 rats, the ischemia induced elevation in STEP33 was delayed relative to P7 rats. STEP33 was produced by digestion of postsynaptic densities with calpain I and by exposure of NT2/D1 cells expressing STEP to the calcium ionophore A23187. The results suggest that ischemia-induced calcium influx results in the calcium-dependent proteolysis of membrane-associated STEP61 and the concomitant release of STEP33 into the cytoplasm. PMID- 10537056 TI - Regulation of rat dopamine transporter mRNA and protein by chronic cocaine administration. AB - This study describes a direct comparison of dopamine transporter (DAT) mRNA and protein, as well as its binding sites, in tissue from the same animals after chronic cocaine administration. Rats were treated twice daily with 25 mg/kg cocaine or with saline. After 8 days of cocaine administration, changes in DAT mRNA levels in the substantia nigra pars compacta and ventral tegmental area were measured by in situ hybridization, and DAT protein in the striatum was quantified by immunoblotting. Whereas chronic cocaine treatment significantly reduced levels of DAT mRNA in the substantia nigra pars compacta and ventral tegmental area as compared with vehicle-treated controls, cocaine treatment did not alter DAT protein levels in the striatum. Furthermore, the density of DAT binding sites was also measured in the striatum by quantitative autoradiography using two DAT radioligands, 33-(4-[125I]iodophenyl)tropane-2-carboxylic acid methyl ester ([125I]RTI-55) and [3H]propanoyl-3beta-(4-tolyl)tropane ([3H]PTT). Similar to the results of immunoblotting of DAT protein, [1251]RTI-55 and [3H]PTT binding site levels also remained unaltered. These results indicate a dissociation in the regulation of DAT mRNA and its protein levels as a result of cocaine administration in rats. This study also indicates that the DAT ligands [3H]PTT and [125I]RTI-55 provide an accurate assessment of DAT protein levels. PMID- 10537058 TI - Calcium-dependent cleavage of striatal enriched tyrosine phosphatase (STEP). AB - Striatal enriched phosphatase (STEP) is a family of protein tyrosine phosphatases enriched within the CNS. A member of this family, STEP61, is a membrane associated protein located in postsynaptic densities of striatal neurons. In this study, we demonstrate that STEP61, is cleaved into smaller isoforms. To clarify the mechanism of cleavage, STEP61 was transiently expressed in NT2/D1 neuronal precursor cells. Exposure of transfected cells to the calcium ionophore, A23187, or to thapsigargin resulted in the rapid cleavage of STEP61. Pretreatment with the calpain inhibitor, calpeptin, or EGTA prevented proteolysis. One of the cleavage products has a relative molecular mass of 33 kDa (STEP33). A protein with the identical mobility is detected following calpain treatment of STEP61 fusion protein or postsynaptic densities purified from rat striatum. Exposure of primary neuronal cultures to glutamate also led to a significant increase in the concentration of a low molecular weight form of STEP. Taken together, these results suggest that in response to a rapid influx of calcium, STEP61, is proteolytically cleaved by calpain, leading to the release of a smaller isoform. This model may explain the rapid appearance of STEP33 in response to transient hypoxia-ischemia in the brain as cells attempt to counter the increase in tyrosine phosphorylation levels following neuronal insults. PMID- 10537059 TI - Transport of cationized anti-tetanus Fab'2 fragments across an in vitro blood brain barrier model: involvement of the transcytosis pathway. AB - Tetanus neurotoxin reaches the CNS by axonal retrograde transport and thus becomes inaccessible to current treatments. A possible strategy to improve current therapy for tetanus disease would be the vectorization of Fab'2 fragments, allowing their delivery into the CNS. The purpose of this study was to investigate whether after cationization anti-tetanus Fab'2 fragments are able to cross the blood-brain barrier, the first obstacle to CNS delivery. We used primary cocultures of bovine brain capillary endothelial cells and newborn rat astrocytes as an in vitro model to study the binding and transport of cationized Fab'2 (cFab'2) fragments across the brain endothelium. We first show that cationization does not alter Fab'2 affinity for tetanus toxin. Then we demonstrate that after cationization Fab'2 fragments are able to bind to the negative charges on the surface of endothelial cells and subsequently to be transported across the endothelial cell monolayer without any modification of affinity. Finally, using fluorescence microscopy, we show that cFab'2 fragments are transported through endocytotic vesicles. The present study demonstrates that cationization allows Fab'2 directed against tetanus toxin to be transported through brain endothelium by adsorptive-mediated transcytosis. We suggest that this vectorization way could be a promising delivery strategy for carrying anti tetanic immunoglobulin fragments across the blood-brain barrier to improve tetanus treatment. PMID- 10537060 TI - Interleukin-6 activates signal transducer and activator of transcription and mitogen-activated protein kinase signal transduction pathways and induces de novo protein synthesis in human neuronal cells. AB - Interleukin-6 (IL-6) is involved in the pathophysiology of various diseases of the CNS. Because the molecular mechanism of action of this cytokine in human neurons is not well understood, we were interested in characterizing and defining a model system for IL-6-induced activation of signal transduction cascades, transcriptional activation, and protein synthesis in human neuronal cells. We show that IL-6 leads to transcriptional activation of signal transducer and activator of transcription 3 (STAT3) in human SH-SY5Y neuroblastoma cells. IL-6 induced activation and translocation of STAT3 and to a lesser degree STAT1 but not STAT5 are demonstrated. STAT3 is phosphorylated on Tyr705 and Ser727 residues on stimulation with IL-6, suggesting maximal activation of transcription. We also show IL-6-induced phosphorylation of p42/44 mitogen-activated protein (MAP) kinase, providing evidence for MAP kinase pathway activation. The physiological relevance of our results is confirmed by IL-6-induced phosphorylation of key signaling proteins of both STAT and MAP kinase pathway in rat primary hippocampal neurons. Furthermore, de novo protein synthesis on IL-6 activation is demonstrated. PMID- 10537061 TI - Tissue transglutaminase is increased in Huntington's disease brain. AB - The polyglutamine-expanded N-terminal region of mutant huntingtin causes neurodegeneration in Huntington's disease (HD). Neuronal intranuclear and cytosolic inclusions composed of mutant huntingtin are found in brains of HD patients. Because tissue transglutaminase cross-links proteins into filamentous aggregates and polypeptide-bound glutamines are primary determining factors for tissue transglutaminase-catalyzed reactions, it has been hypothesized that tissue transglutaminase may contribute to the formation of these aggregates. In this report immunohistochemical and biochemical methods were used to demonstrate that tissue transglutaminase expression and transglutaminase activity are elevated in HD brains in a grade-dependent manner. In the striatum, tissue transglutaminase activity was significantly increased in the grade 3 HD cases compared with controls. When normalized to the neuronal marker calbindin D28k, immunoblot analysis revealed that in the striatum the levels of tissue transglutaminase were significantly increased in all HD cases compared with controls. Immunohistochemical staining of the HD striatum revealed that tissue transglutaminase immunoreactivity was markedly increased in all grades as compared with controls. In the superior frontal cortex, tissue transglutaminase activity was significantly higher in all HD cases as compared with controls. Quantitative analysis of immunoblots demonstrated that tissue transglutaminase levels were elevated in HD grades 2 and 3 cases. Tissue transglutaminase immunoreactivity within the superior frontal neocortex was also greater in all the HD cases compared with controls. These data clearly indicate that tissue transglutaminase is elevated in HD brain and may play a role in the disease process. PMID- 10537062 TI - Preservation of noradrenergic neurons in the locus ceruleus that coexpress galanin mRNA in Alzheimer's disease. AB - Galanin (GAL) innervation is hypertrophied in the basal forebrain and cortex of patients with Alzheimer's disease (AD). Increased GAL could exacerbate the cognitive and behavioral deficits of AD because GAL acts as an inhibitory modulator of cholinergic and noradren-ergic neurotransmission. The locus ceruleus (LC) may be a source of increased GAL in AD because (a) GAL is coexpressed in a subset of LC neurons, (b) GAL expression is up-regulated with neuronal injury, and (c) the LC undergoes extensive degeneration in AD. Therefore, we have used in situ hybridization histochemistry to measure GAL gene expression in the LC of AD patients and sex- and age-matched nondemented controls. Despite the extensive loss of norepinephrine neurons with AD, GAL mRNA-expressing neurons in the LC did not differ between groups. This resulted in a significant increase in the percentage of neuromelanin-pigmented cells that coexpressed GAL in AD patients compared with controls. These findings raise the possibility that the increased incidence of GAL expression among remaining LC neurons contributes to the hyperinnervation of GAL fibers in AD. Furthermore, GAL may be neuroprotective in the LC. PMID- 10537063 TI - Activation of Akt kinase inhibits apoptosis and changes in Bcl-2 and Bax expression induced by nitric oxide in primary hippocampal neurons. AB - Emerging data indicate that growth factors such as insulin-like growth factor-1 (IGF-1) prevent neuronal death due to nitric oxide (NO) toxicity. On the other hand, growth factors can promote cell survival by acting on phosphatidylinositol 3-kinase (PI3-kinase) and its downstream target, serine-threonine kinase Akt, in various types of cells. Here, we examined the mechanism by which IGF-1 inhibits neuronal apoptosis induced by NO in primary hippocampal neurons. IGF-1 was capable of preventing apoptosis and caspase-3-like activation induced by a NO donor, sodium nitroprusside or 3-morpholin-osydnonimine. Incubation of neurons with a P13-kinase inhibitor, wortmannin or LY294002, blocked the effects of IGF-1 on NO-induced neurotoxicity and caspase-3-like activation. In addition, the P13 kinase inhibitors blocked the effect of IGF-1 on down-regulation in Bcl-2 and upregulation in Bax expression induced by NO. Adenovirus-mediated overexpression of the activated form of Akt significantly inhibited NO-induced cell death, caspase-3-like activation, and changes in Bcl-2 and Bax expression. Moreover, expression of the kinase-defective form of Akt almost completely blocked the effects of IGF-1. These findings suggest that activation of Akt is necessary and sufficient for the effect of IGF-1 and is capable of preventing NO-induced apoptosis by modulating the NO-induced changes in Bcl-2 and Bax expression. PMID- 10537064 TI - Kinetic modeling of 52Fe/52mMn-citrate at the blood-brain barrier by positron emission tomography. AB - The kinetics of iron at the blood-brain barrier of the monkey were studied in vivo using positron emission tomography (PET) and the tracer 52Fe/52mMn-citrate. 52mMn is the beta(+)-emitting daughter nuclide of 52Fe and therefore contributes to the observed signal and background in the PET images and may influence the quantification of physiological relevant iron parameters. The kinetics of pure (52m)Mn-citrate at the blood-brain barrier of the monkey were studied experimentally, and the analysis of the data with a reasonable compartment model led to equal efflux and influx parameters for Mn (1.35 +/- 0.3 x 10(-2) min(-1)). By using complexes between Mn and diethylenetriaminepentaacetic acid, the validity of the proposed model could be confirmed. To describe the observed kinetics of 52Fe/(52m)Mn-citrate, the manganese model was coupled to an iron model, which finally allowed the quantification of two iron-specific parameters: an input rate into global brain tissue of 7.15 +/- 2.6 x 10(-4) min(-1) and a time delay of roughly 24 min to account for the observed activities. The simpler linearization procedure has been proposed and could be applied to all our data sets and is able to replace the complicated nonlinear iron/manganese tracer kinetic model neglecting any influence of manganese on the analysis. PMID- 10537065 TI - Proprotein convertase activity contributes to the processing of the Alzheimer's beta-amyloid precursor protein in human cells: evidence for a role of the prohormone convertase PC7 in the constitutive alpha-secretase pathway. AB - The physiological maturation of the beta-amyloid precursor protein (betaAPP) leads to the secretion of a fragment termed APPalpha, after cleavage by a proteolytic enzyme called-secretase. In Alzheimer's disease, betaAPP undergoes exacerbated proteolytic attacks by beta- and gamma-secretases, which liberate a readily aggregatable 40-42-amino acid peptide called AP. We show here that overexpression of the prohormone convertase PC7 triggers increased secretion of APPalpha and lowers both Abeta40 and Abeta42 recoveries. Overexpression of alpha1 antitrypsin Portland (alpha1-PDX), which blocks mammalian precursor convertases of the constitutive secretory pathway, reverses the PC7-induced APPalpha increase as well as the decrease of Abeta40/42 in HEK293 cells. It is interesting that alpha1-PDX also lowers the level of APPalpha endogenously produced by mock transfected HEK293 cells. Finally, a Jurkat clone stably expressing alpha1-PDX produces noticeably lower amounts of APPalpha. Therefore, this serpin affects endogenous a-secretase activity/pathway in distinct cell types. By contrast, alpha1-PDX does not alter the processing of presenilin 1 or its mutated congeners linked to some familial forms of Alzheimer's disease. Altogether, we demonstrate that a prohormone convertase participates in the alpha-secretase pathway of betaAPP maturation in human cells and concomitantly contributes to slowing the pathogenic route leading to the formation of Abeta. Our data strongly suggest that PC7 could fulfill such a role. PMID- 10537066 TI - Effects of aging on the interaction between glutamate, dopamine, and GABA in striatum and nucleus accumbens of the awake rat. AB - The aim of the present study was to investigate, using microdialysis, the effects of aging on the glutamate/dopamine/GABA interaction in striatum and nucleus accumbens of the awake rat. For that, the effects of an increase of the endogenous concentration of glutamate on the extracellular concentration of dopamine and GABA in striatum and nucleus accumbens of young (2-4 months), middle aged (12-14 months), aged (27-33 months), and very aged (37 months) male Wistar rats were studied. Endogenous extracellular glutamate was selectively increased by perfusing the glutamate uptake inhibitor L-trans-pyrrolidine-3,4-dicarboxylic acid (PDC) through the microdialysis probe. In young rats, PDC (1, 2, and 4 mM) produced a dose-related increase of dialysate concentrations of glutamate in both striatum and nucleus accumbens. PDC also increased dialysate dopamine and GABA in both structures. These increases were significantly correlated with the increases of glutamate but not with the PDC dose used, which strongly suggests that the increases of dopamine and GABA were produced by glutamate. In striatum, there were no significant differences in the dopamine/glutamate and GABA/glutamate correlations between young and aged rats. This means that the effects of glutamate on dopamine and GABA do not change during aging. On the contrary, in the nucleus accumbens of aged rats, the increases of dopamine, when correlated with the increases of glutamate, were significantly lower than in young rats. Moreover, the ratio of dopamine to glutamate increases at maximal increases of glutamate was negatively correlated with aging. On the contrary, the ratio of GABA to glutamate increases in nucleus accumbens was positively correlated with aging, which suggests that the effects of endogenous glutamate on GABA tend to be higher in the nucleus accumbens of aged rats. The findings of this study suggest that aging changes the interaction between endogenous glutamate, dopamine, and GABA in nucleus accumbens, but not in striatum, of the awake rat. PMID- 10537067 TI - Lithium inhibits neurite growth and tau protein kinase I/glycogen synthase kinase 3beta-dependent phosphorylation of juvenile tau in cultured hippocampal neurons. AB - Tau protein kinase I(TPKI)/glycogen synthase kinase (GSK)-3beta is abundant in the developing rat brain. The highly phosphorylated juvenile form of tau is present during the same developmental period. To study the role of TPKI/ GSK 3beta in neuronal growth, we examined the effects of lithium, a direct inhibitor of TPKI/GSK-3beta, using primary cultures of rat hippocampal neurons. Immunohistochemical staining of the neurons indicates that in the presence of lithium (2-10 mM), neurite growth becomes inhibited in a dose-dependent manner. Western blot analyses of the cell extracts revealed that the presence of lithium in the culture medium increased the amount of dephosphorylated tau while decreasing phosphorylation at Ser199 and Ser396, both of which are TPKI/GSK-3beta phosphorylation sites on tau. The inhibition by lithium is reversible. Although the amount of TPKI/GSK-3beta remained constant, the amount of tau decreased in a dose-dependent manner in the presence of lithium. TPKI/GSK-3beta was distributed in the somata and proximal neurites of the cultured hippocampal neurons. These results therefore suggest that TPKI/GSK-3beta plays an important role in the axonal growth of neurons during synapse formation in the developing brain. PMID- 10537068 TI - Midkine inhibits caspase-dependent apoptosis via the activation of mitogen activated protein kinase and phosphatidylinositol 3-kinase in cultured neurons. AB - Midkine (MK) is a new member of the heparin-binding neurotrophic factor family. MK plays important roles in development and carcinogenesis and has several important biological effects, including promotion of neurite extension and neuronal survival. However, the mechanism by which MK exerts its neurotrophic actions on neurons has not been elucidated to date. We have established an apoptosis induction system by serum deprivation in primary neuronal cultures isolated from mouse cerebral cortices. Neuronal apoptosis induced by serum deprivation was accompanied by the activation of caspase-3. MK, when added into the culture medium, inhibited the induction of apoptosis and activation of caspase-3 in a dose-dependent manner. Extracellular signal-regulated kinase (ERK) and Akt were not activated by serum deprivation, whereas ERK and Akt were rapidly activated by addition of MK. In addition, the trophic actions of MK of suppressing apoptosis and suppressing the activation of caspase-3 were abolished by concomitant treatment with PD98059, a specific inhibitor of mitogen-activated protein kinase kinase, and with wort-mannin or LY294002, specific inhibitors of phosphatidyl-inositol 3-kinase (PI 3-kinase). These PI 3-kinase inhibitors also inhibited the activation of ERK in response to MK, demonstrating a link between ERK and the caspase-3 pathway that is modulated by the PI 3-kinase activation. These results indicate that the ERK cascade plays a central role in MK-mediated neuronal survival via inhibition of caspase-3 activation. PMID- 10537069 TI - Alpha-synuclein immunoisolation of glial inclusions from multiple system atrophy brain tissue reveals multiprotein components. AB - Immunohistochemical studies have shown that oligodendroglial inclusions in multiple system atrophy contain alpha-synuclein, a synaptic protein also found in Lewy bodies in Parkinson's disease. We have now used density gradient enrichment and an anti-alpha-synuclein immunomagnetic technique to isolate pure and morphologically intact oligodendroglial inclusions from brain white matter of patients dying with multiple system atrophy. Filamentous inclusion structures were obtained only from multiple system atrophy tissue, but not from normal brain tissues, or from multiple system atrophy tissue processed without anti-alpha synuclein antibody. We confirmed the purity and morphology of isolated inclusions by electron microscopy. The inclusions comprised multiple protein bands after separation by polyacrylamide gel electrophoresis. Immunoblotting demonstrated that these proteins included alpha-synuclein, alphaB-crystallin, tubulins, ubiquitin, and prominent, possibly truncated alpha-synuclein species as high molecular-weight aggregates. Our study provides the first biochemical evidence that oligodendroglial inclusion filaments consist of multiple protein components, suggesting that these inclusions may form as a result of multiprotein interactions with alpha-synuclein. PMID- 10537070 TI - Disruption of receptor-mediated activation of G protein by mutating a conserved arginine residue in the switch II region of the alpha subunit. AB - A naturally occurring point mutation (R231H) within one of the major 3gamma binding surface (switch II region) on the a subunit of Gs (alpha(s)) has previously been found to disrupt receptor-mediated activation of Gs. The disruption caused by mutating this conserved residue may be a general phenomenon for all a subunits. Homologous mutants of the alpha subunit of Gz [alpha(z); a negative regulator of adenylyl cyclase (AC)] and G16 (alpha16; a stimulator of phospholipase C) were constructed and examined for receptor-mediated regulation of their corresponding effectors. The mutant alphazR209H cannot be fully activated by the delta-opioid receptor, as indicated by the impairment of the inhibition of alpha(s)-stimulated AC and betagamma-mediated stimulation of AC type II (AC2). Similarly, the mutant alpha16R216H lost the ability to mediate receptor-induced activation of phospholipase C and AC2. The receptor coupling efficacy and promiscuity of alpha16R216H were eradicated. The mutation of the conserved arginine has no observable effect on the constitutive activities of the GTPase-deficient derivatives of both alpha(z) and alpha16. The alpha subunit of Gt1 (transducin; alphat1) attenuated betagamma-mediated stimulation of AC2 by sequestrating free betagamma subunits, but the mutant alphat1R204H showed reduced ability to scavenge betagamma-mediated AC2 activation. Presumably, mutation of the conserved arginine disrupted the subunit interactions in addition to the impairment of receptor interaction. PMID- 10537071 TI - The small myelin-associated glycoprotein is a zinc-binding protein. AB - The myelin-associated glycoprotein is a transmembrane cell adhesion molecule expressed specifically by myelinating glial cells of the nervous system. Its two isoforms, whose amino acid sequences differ only by their respective cytoplasmic carboxy-terminal domains, are important for the formation and maintenance of a normal functional myelin sheath. In this study, by using recombinant proteins, we identify the cytoplasmic domain of the small isoform of the myelin-associated glycoprotein as a zinc-binding protein. The observed dissociation constant lies in the low micromolar range (K(D) = 6-7 microM). The binding of zinc by the small myelin-associated glycoprotein induces a conformational change that enables the protein to reversibly bind to a hydrophobic phenyl-Sepharose matrix. Our results also suggest that zinc may induce dimerization of the small myelin-associated glycoprotein. We suggest roles for zinc in the stabilization of the structure of the cytoplasmic domain of the small myelin-associated glycoprotein and in protein protein interactions that involve this short domain. PMID- 10537073 TI - Interprotein disulfide bonds formed during isolation process tighten the structure of the postsynaptic density. AB - Postsynaptic densities (PSDs) isolated by biochemical means consist of a complex mixture of proteins that tightly bond to each other. The purpose of this report is to study whether the numerous interprotein disulfides found in the isolated PSDs contribute to the tight structure of the PSDs and whether these interprotein disulfides exist in vivo. PSDs were isolated from pig cerebral cortex by conventional methods except that iodoacetic acid (IAA) was added to all solutions to curtail the formation of disulfides during the isolation process. The PSDs thus isolated were fragmented easily by treatment with chaotropic reagents or ionic detergents, whereas the PSDs isolated in IAA-free solutions were resistant to these treatments. Electron microscopy revealed that the PSDs isolated in IAA containing solutions were more fragmented than those isolated in IAA-free solutions. Furthermore, the PSD sample isolated in IAA-free solutions contained very large disulfide-linked aggregates that were virtually absent from the PSDs isolated in IAA-containing solutions. Our results suggest that the exceptionally tight structure of the PSDs isolated by conventional methods is due largely to the new disulfides formed during the isolation process and that the PSD proteins under in vivo conditions are held together primarily by noncovalent interactions. PMID- 10537072 TI - Distribution and intracellular localization of a mouse homologue of Ca2+/calmodulin-dependent protein kinase Ibeta2 in the nervous system. AB - Ca2+/calmodulin-dependent protein kinases (CaMKs) are believed to play important roles in the development and function of the nervous system. We report here the identification and expression of mouse CaMKIbeta (mCaMKIbeta), in particular mCaMKIbeta2, an isoform of mCaMKIbeta. During embryogenesis, the mCaMKIbeta2 gene is expressed mainly in the nervous system, including brain, spinal cord, trigeminal ganglion, and retina. Within the CNS, the expression of mCaMKIbeta2 is detected in the mantle zone, but not in the ventricular zone, suggesting its possible involvement in the differentiation of neurons. In the adult brain, mCaMKIbeta2 transcripts are detected at high levels in the anterior olfactory nuclei, piriform cortex, septal nuclei, bed nuclei of the stria terminalis, hippocampal pyramidal cells, dentate granule cells, amygdala, hypothalamic nuclei, parabrachial nucleus, and nucleus of the solitary tract. The distinct gene expression pattern suggests that mCaMKIbeta2 may also be involved in different mature neuronal functions from other CaMKs. In addition, mCaMKI/beta2 proteins are localized to the cytoplasm and nuclei, but not to nucleoli, suggesting that mCaMKIbeta2 proteins might be involved in the cytoplasmic and nuclear signal transduction of the nervous system. PMID- 10537074 TI - Hypoxic response of synaptosomal proteins in term guinea pig fetuses. AB - Early events in the hypoxia-induced response trigger tyrosine phosphorylation cascades involving a large number of enzymes and substrates. The resolving power of advanced two-dimensional gel electrophoresis, followed by immunoblotting with specific antibodies to phosphotyrosine, has been used to analyze hypoxia-induced modifications in guinea pig brain synaptosomes. These procedures, in conjunction with computer-aided image analysis, are useful in the differential display of gene products, providing comparison at the level of posttranslationally modified products. Studies were performed in cerebral cortical synaptosomes from three normoxic and three hypoxic newborn guinea pigs. To filter off background noise consisting of nonreproducible migrating protein spots, only reproducible features of electrophoretic patterns were considered. Immunoreactivity patterns obtained with anti-phosphotyrosine antibodies proved to be different in normoxic and hypoxic synaptosomes: of a total of 130 immunoreactive spots, 49 were tyrosine phosphorylated in hypoxic synaptosomes only and 20 in the normoxic ones only. Our data suggest that hypoxia extensively remodels the signaling pathway by switching off tyrosine phosphorylation of some cellular components (i.e., alpha-internexin) and switching on tyrosine phosphorylation of some other proteins (i.e., heat shock cognate 70, aconitase, 2',3'-cyclic nucleotide 3'-phosphodiesterase, and pyruvate kinase). PMID- 10537075 TI - Nitric oxide-independent down-regulation of soluble guanylyl cyclase by bacterial endotoxin in astroglial cells. AB - Induction of nitric oxide (NO) synthase (NOS) type 2 (NOS-2) in glial cells after exposure to bacterial endotoxin [lipopolysaccharide (LPS)] or inflammatory cytokines has been repeatedly demonstrated both in vitro and in vivo. However, little is known about effects of these agents on NO-dependent cyclic GMP (cGMP) formation. In this work, we show that treatment of rat cerebellar astrocyte enriched primary cultures with LPS decreases NO donor-stimulated cGMP formation with a similar initial time course (up to 9-12 h) and concentration dependency (0.1-1 ng/ml) as for induction of NOS-2. This effect appears to be due to a down regulation of soluble guanylyl cyclase (sGC) because LPS treatment decreases sGC activity and sGC beta1 subunit levels. In contrast, cGMP phosphodiesterase activity and stimulation of the particulate guanylyl cyclase by atrial natriuretic peptide are not affected. Incubation of astroglial cultures with a transcription inhibitor (actinomycin D) or a protein synthesis inhibitor (cycloheximide) for 18-20 h does not decrease sGC activity but totally prevents LPS-induced desensitization of sGC. Inhibition of NOS-2 activity with N(G) monomethyl-L-arginine or inhibition of NOS-2 induction with the synthetic glucocorticoid dexamethasone failed to prevent the inhibitory effect of LPS on sGC, indicating that NO production is not involved. Moreover, after removal of LPS the time for recovery of sGC responsiveness is much longer than that for NOS 2 return to basal levels. LPS impairment of cGMP formation also occurs in cortical astrocytes but not in cerebellar granule neurons. The decreased responsiveness of sGC to NO stimulation following LPS challenge may prevent inappropriate astroglial cGMP signaling caused by excess production of NO by adjacent activated glial cells. Key Words: Astroglia-Neurons-Nitric oxide-Soluble guanylyl cyclase-Lipopolysaccharide. PMID- 10537076 TI - Identification of a new ligand binding domain in the alpha1 subunit of the inhibitory glycine receptor. AB - Four discontinuous extracellular sequence domains have been proposed to form the ligand binding sites of the ligand-gated ion channel receptor superfamily. In this study, we investigated the role of 12 contiguous residues of the inhibitory glycine receptor that define the proposed "loop A" ligand binding domain. Using the techniques of site-directed mutagenesis and patch-clamp electrophysiology, four of the 12 residues were shown to have impaired ligand binding. Three mutants, 193A, A101H, and N102A, resulted in significant (17-44-fold) increases in the agonist EC50 values as compared with the wild-type glycine receptor, whereas Hill coefficients, ImaX values, and antagonist affinity remained largely unaffected. Consideration of receptor efficacy values indicates that these residues are involved in ligand binding rather than channel activation. A fourth mutant, W94A, failed to give rise to any glycine-activated currents, although cell-surface expression was observed, suggesting that this residue may also be involved in agonist binding. These data provide the most extensive characterization of the loop A ligand binding domain available to date and define two new residue locations, Ile93 and Asn102, as contributing to the four-loop model of ligand binding. PMID- 10537077 TI - Prostaglandin E2 induces Ca2+ release from ryanodine/caffeine-sensitive stores in bovine adrenal medullary cells via EP1-like receptors. AB - Prostaglandin E2 (PGE2) causes Ca2+ release from intracellular Ca2+ stores and stimulates phosphoinositide metabolism in bovine adrenal medullary cells. These results have been interpreted as PGE2 induces Ca2+ release from inositol trisphosphate (IP3)-sensitive stores. However, we have recently shown that pituitary adenylate cyclase-activating polypeptide (PACAP), bradykinin, and angiotensin II release Ca2+ from caffeine/ryanodine-sensitive stores, although they cause a concomitant increase of intracellular IP3. In light of these results, the mechanism of PGE2-induced Ca2+ release was investigated in the present study. PGE2 dose-dependently caused a transient but consistent Ca2+ release from internal Ca2+ stores. The PGE2-induced Ca2+ release was unaffected by cinnarizine, a blocker of IP3-induced Ca2+ release. By contrast, it was potently inhibited by prior application of caffeine and ryanodine. Although IP3 production in response to PGE2 was abolished by the phospholipase C inhibitor U 73122, Ca2+ release in response to PGE2 was unaffected by U-73122. The PGE2 induced Ca2+ release was unaffected by Rp-adenosine 3',5'-cyclic monophosphothioate, an inhibitor of protein kinase A, and forskolin, a cyclic AMP (cAMP)-elevating agent, did not cause Ca2+ release. The EP1 agonist 17-phenyl trinorPGE2 and the EP1/EP3 agonist sulprostone mimicked the Ca(2+)-releasing effects of PGE2, whereas the EP2 agonist butaprost or the EP2/EP3 agonist misoprostol caused little or no Ca2+ release. The EP1 antagonist SC-51322 significantly suppressed the Ca2+ release response induced by PGE2, whereas the EP4 antagonist AH-23828B had little effect. These results suggest that PGE2, acting on EP1-like receptors, induces Ca2+ release from ryanodine/caffeine sensitive stores through a mechanism independent of IP3 and cAMP and that PGE2 may share the same mechanism with PACAP and the other peptide ligands in causing Ca2+ release in bovine adrenal medullary cells. PMID- 10537078 TI - Differential changes in the phosphorylation of the protein kinase C substrates myristoylated alanine-rich C kinase substrate and growth-associated protein-43/B 50 following Schaffer collateral long-term potentiation and long-term depression. AB - Activation of protein kinase C (PKC) is one of the biochemical pathways thought to be activated during activity-dependent synaptic plasticity in the brain, and long-term potentiation (LTP) and long-term depression (LTD) are two of the most extensively studied models of synaptic plasticity. Here we have examined changes in the in situ phosphorylation level of two major PKC substrates, myristoylated alanine-rich C kinase substrate (MARCKS) and growth-associated protein (GAP)-43/B 50, after pharmacological stimulation or induction of LTP or LTD in the CA1 field of the hippocampus. We find that direct PKC activation with phorbol esters, K+ induced depolarization, and activation of metabotropic glutamate receptors increase the in situ phosphorylation of both MARCKS and GAP-43/B-50. The induction of LTP increased the in situ phosphorylation of both MARCKS and GAP 43/B-50 at 10 min following high-frequency stimulation, but only GAP-43/B-50 phosphorylation remained elevated 60 min after LTP induction. Furthermore, blockade of LTP induction with the NMDA receptor antagonist D-2-amino-5 phosphonopentanoic acid prevented elevations in GAP-43/B-50 phosphorylation but did not prevent the elevation in MARCKS phosphorylation 10 min following LTP induction. The induction of LTD resulted in a reduction in GAP-43/B-50 phosphorylation but did not affect MARCKS phosphorylation. Together these findings show that activity-dependent synaptic plasticity elicits PKC-mediated phosphorylation of substrate proteins in a highly selective and coordinated manner and demonstrate the compartmentalization of PKC-substrate interactions. Key Words: Protein kinase C-Myristoylated alanine-rich C kinase substrate-Growth associated protein-43-Long-term potentiation-Long-term depression-(RS)-alpha Methyl-4-carboxyphenylglycine-D-2-Amino-5-ph osphonopentanoic acid-Glutamate. PMID- 10537079 TI - The astroglial ASCT2 amino acid transporter as a mediator of glutamine efflux. AB - Glutamine release from astrocytes is an essential part of the glutamate-glutamine cycle in the brain. Uptake of glutamine into cultured rat astrocytes occurs by at least four different routes. In agreement with earlier studies, a significant contribution of amino acid transport systems ASC, A, L, and N was detected. It has not been determined whether these systems are also involved in glutamine efflux or whether specific efflux transporters exist. We show here that ASCT2, a variant of transport system ASC, is strongly expressed in rat astroglia-rich primary cultures but not in neuron-rich primary cultures. The amino acid sequence of rat astroglial ASCT2 is 83% identical to that of mouse ASCT2. In Xenopus laevis oocytes expressing rat ASCT2, we observed high-affinity uptake of [U 14C]glutamine (Km = 70 microM) that was Na(+)-dependent, concentrative, and unaffected by membrane depolarization. When oocytes were preloaded with [U 14C]glutamine, no glutamine efflux was detected in the absence of extracellular amino acids. Neither lowering intracellular pH nor raising the temperature elicited efflux. However, addition of 0.1 mM unlabeled alanine, serine, cysteine, threonine, glutamine, or leucine to the extracellular solution resulted in a rapid release of glutamine from the ASCT2-expressing oocytes. Amino acids that are not recognized as substrates by ASCT2 were ineffective in this role. Extracellular glutamate stimulated glutamine release weakly at pH 7.5 but was more effective on lowering pH to 5.5, consistent with the pH dependence of ASCT2 affinity for glutamate. Our findings suggest a significant role of ASCT2 in glutamine efflux from astrocytes by obligatory exchange with extracellular amino acids. However, the relative contribution of this pathway to glutamine release from cells in vivo or in vitro remains to be determined. PMID- 10537080 TI - Regional distribution and developmental changes of GluR1-flop protein revealed by monoclonal antibody in rat brain. AB - From immunizations of mice with a glutathione S-transferase fusion protein containing residues 724-781 of the alpha-amino-3-hydroxy-5-methyl-4 isoxazolepropionic acid receptor subunit GluR1-flop, two monoclonal antibodies (mAbs) were developed that differed widely in their ranges of specificity. In immunocytochemical and immunoblotting assays performed on COS-7 cells transfected with one of the eight GluR1-4-flip/flop forms, mAb 19B10 recognized all eight forms, whereas mAb 8E11 was specific for GluR1-flop. By means of synthetic peptides, the epitopes were determined to be NKWWYDKG (GluR1-flop760-767) for mAb 19B10 but GSALRNPVN (GluR1-flop740-748) plus a partial epitope, QGLL (GluR1 flop757-760), for mAb 8E11. Further analysis on synthetic peptides pointed to a potential cross-reactivity of mAb 8E11 with GluR2-4-flop variants that lacked editing at the R/G site. The contribution of such cross-reactivities in histoblot labeling patterns on adult rat brain material, however, was judged to be negligible. Histoblot patterns with mAb 8E11 were dominated by strong immunoreactivity in CA1 strata radiatum and oriens and in the dentate molecular layer, whereas the CA3 region was virtually free of labeling. This pattern and those observed at different stages of postnatal development were generally similar to regional distribution patterns previously reported in the literature for GluR1-flop transcripts. Key Words: Glutamate receptors-AMPA receptor subunits Alternative splicing-Developmentally regulated expression-R/G site. PMID- 10537081 TI - Interleukin-1beta induces substance P release from primary afferent neurons through the cyclooxygenase-2 system. AB - Substance P (SP) is synthesized in the dorsal root ganglion (DRG) and released from primary afferent neurons to convey information regarding noxious stimuli. The effects of the proinflammatory cytokine interleukin-1 (IL-1) beta on the release of SP were investigated using primary cultured rat DRG cells. Recombinant mouse IL-1beta added to the cells at 0.1 ng/ml increased the SP-like immunoreactivity (SPLI) in the culture medium after incubation for 6 h by approximately 50% as compared with that of nontreated DRG cells. The effect of IL 1beta was Ca(2+)-dependent and significantly inhibited by 100 ng/ml IL-1 receptor specific antagonist (IL-1r antagonist), cyclooxygenase (COX) inhibitors such as 0.1 mM aspirin, 1 microg/ml indomethacin, and 1 microM NS-398 (specific for COX 2), and 1 microM dexamethasone. Furthermore, a 1-h incubation with IL-1beta markedly increased the inducible COX-2 mRNA level, which was inhibited by an IL 1r antagonist and dexamethasone, whereas IL-1beta showed no effect on the level of constitutive COX-1 mRNA. These observations indicated that IL-1beta induced the release of SP from the DRG cells via specific IL-1 receptors, the mechanism of which might involve prostanoid systems produced by COX-2. This could be responsible for the hyperalgesic action with reference to inflammatory pain in the primary afferent neuron to spinal cord pathway. PMID- 10537082 TI - Cyclosporin A-sensitive changes in mitochondrial glutathione are an early response to intrastiatal NMDA or forebrain ischemia in rats. AB - An intrastriatal injection of NMDA produced an increase in glutathione to 152% of control values in mitochondria isolated from striatum at 1 h later. Total tissue glutathione was not changed. The mitochondrial increase was largely reversed by 2 h. Glutathione content was not significantly affected in mitochondria from a part of the cerebral cortex that did not exhibit damage following intrastriatal NMDA. Glutathione was similarly increased in mitochondria from both cortex and striatum at 1 h after a short period of forebrain ischemia, confirming our previous findings. The increases in mitochondrial glutathione developed shortly after accumulations of mitochondrial calcium that have been observed previously. Intravenous injection of cyclosporin A immediately following either the NMDA treatment or reversal of the ischemic period partially inhibited the increases in glutathione in mitochondria from the affected brain subregions. These studies provide evidence that early changes sensitive to cyclosporin A develop in mitochondria under pathological conditions in the intact brain. These glutathione increases are consistent with an induction of the mitochondrial permeability transition in the affected tissue. PMID- 10537083 TI - Reduced Nurr1 expression increases the vulnerability of mesencephalic dopamine neurons to MPTP-induced injury. AB - Mutation in the Nurr1 gene, a member of the nuclear receptor superfamily, causes selective agenesis of dopaminergic neurons in the midbrain of null mice. Homozygous Nurr1 knockout mice (Nurr1-/-) die 1 day after birth, but heterozygous mice (Nurr1 +/-) survive postnatally without obvious locomotor deficits. Although adult Nurr1 +/- mice show significantly reduced Nurr1 protein levels in the substantia nigra (SN), they display a normal range of tyrosine hydroxylase positive neuron numbers in the SN and normal levels of dopamine in the striatum. The reduction in Nurr1 expression in Nurr1 +/- mice, however, confers increased vulnerability to the selective dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine (MPTP) compared with wild-type (Nurr1 +/+) mice. This study suggests that Nurr1 may play an important role in maintaining mature mesencephalic dopaminergic neuron function and that a defect in Nurr1 may increase susceptibility to SN injury. PMID- 10537084 TI - Extracellular glucose concentrations in the brain. PMID- 10537085 TI - Caution: traditional knowledge. PMID- 10537086 TI - Medicine Nobel goes to pioneer of protein guidance mechanisms. PMID- 10537087 TI - Publishers agree on a 'seamless web'. PMID- 10537088 TI - Split-second chemistry is rewarded. PMID- 10537089 TI - Varmus announces decision to quit NIH for cancer centre. PMID- 10537090 TI - Anger at Israeli sex crimes DNA bank. PMID- 10537091 TI - German garlic study under scrutiny. PMID- 10537092 TI - Berkeley puts $500m into multidisciplinary approach to disease. PMID- 10537093 TI - Joint institute set to boost Austria's genome research. PMID- 10537094 TI - Institut Pasteur names Kourilsky as new director. PMID- 10537095 TI - ICSU seeks to classify 'traditional knowledge'. International Council of Scientific Unions. PMID- 10537096 TI - UK government not convinced by claims for flu drug. PMID- 10537097 TI - Conventional crops are the test of GM prejudice. PMID- 10537098 TI - No GM conspiracy. PMID- 10537099 TI - Putting transparency into ethical balance. PMID- 10537100 TI - Neurobiology. Straight from the top. PMID- 10537101 TI - Developmental biology. Controlling the cellular brakes. PMID- 10537102 TI - Dual personality of memory T cells. PMID- 10537103 TI - Familiarity breeds contempt in guppies. PMID- 10537104 TI - Apoptosis. Searching for FLASH domains. PMID- 10537105 TI - Id1 and Id3 are required for neurogenesis, angiogenesis and vascularization of tumour xenografts. AB - Id proteins may control cell differentiation by interfering with DNA binding of transcription factors. Here we show that targeted disruption of the dominant negative helix-loop-helix proteins Id1 and Id3 in mice results in premature withdrawal of neuroblasts from the cell cycle and expression of neural-specific differentiation markers. The Id1-Id3 double knockout mice also display vascular malformations in the forebrain and an absence of branching and sprouting of blood vessels into the neuroectoderm. As angiogenesis both in the brain and in tumours requires invasion of avascular tissue by endothelial cells, we examined the Id knockout mice for their ability to support the growth of tumour xenografts. Three different tumours failed to grow and/or metastasize in Id1+/- Id3-/- mice, and any tumour growth present showed poor vascularization and extensive necrosis. Thus, the Id genes are required to maintain the timing of neuronal differentiation in the embryo and invasiveness of the vasculature. Because the Id genes are expressed at very low levels in adults, they make attractive new targets for anti-angiogenic drug design. PMID- 10537106 TI - Symmetry in locomotor central pattern generators and animal gaits. AB - Animal locomotion is controlled, in part, by a central pattern generator (CPG), which is an intraspinal network of neurons capable of generating a rhythmic output. The spatio-temporal symmetries of the quadrupedal gaits walk, trot and pace lead to plausible assumptions about the symmetries of locomotor CPGs. These assumptions imply that the CPG of a quadruped should consist of eight nominally identical subcircuits, arranged in an essentially unique matter. Here we apply analogous arguments to myriapod CPGs. Analyses based on symmetry applied to these networks lead to testable predictions, including a distinction between primary and secondary gaits, the existence of a new primary gait called 'jump', and the occurrence of half-integer wave numbers in myriapod gaits. For bipeds, our analysis also predicts two gaits with the out-of-phase symmetry of the walk and two gaits with the in-phase symmetry of the hop. We present data that support each of these predictions. This work suggests that symmetry can be used to infer a plausible class of CPG network architectures from observed patterns of animal gaits. PMID- 10537107 TI - Probing the human stereoscopic system with reverse correlation. AB - Our two eyes obtain slightly different views of the world. The resulting differences in the two retinal images, called binocular disparities, provide us with a stereoscopic sense of depth. The primary visual cortex (V1) contains neurons that are selective for the disparity of individual elements in an image, but this information must be further analysed to complete the stereoscopic process. Here we apply the psychophysical technique of reverse correlation to investigate disparity processing in human vision. Observers viewed binocular random-dot patterns, with 'signal' dots in a specific depth plane plus 'noise' dots with randomly assigned disparities. By examining the correlation between the observers' ability to detect the plane and the particular sample of 'noise' disparities presented on each trial, we revealed detection 'filters', whose disparity selectivity was remarkably similar to that of individual neurons in monkey V1. Moreover, if the noise dots were of opposite contrast in the two eyes, the tuning inverted, just like the response patterns of V1 neurons. Reverse correlation appears to probe disparity processing at the earliest stages of binocular combination, prior to the generation of a full stereoscopic depth percept. PMID- 10537108 TI - Top-down signal from prefrontal cortex in executive control of memory retrieval. AB - Knowledge or experience is voluntarily recalled from memory by reactivation of the neural representations in the cerebral association cortex. In inferior temporal cortex, which serves as the storehouse of visual long-term memory, activation of mnemonic engrams through electric stimulation results in imagery recall in humans, and neurons can be dynamically activated by the necessity for memory recall in monkeys. Neuropsychological studies and previous split-brain experiments predicted that prefrontal cortex exerts executive control upon inferior temporal cortex in memory retrieval; however, no neuronal correlate of this process has ever been detected. Here we show evidence of the top-down signal from prefrontal cortex. In the absence of bottom-up visual inputs, single inferior temporal neurons were activated by the top-down signal, which conveyed information on semantic categorization imposed by visual stimulus-stimulus association. Behavioural performance was severely impaired with loss of the top down signal. Control experiments confirmed that the signal was transmitted not through a subcortical but through a fronto-temporal cortical pathway. Thus, feedback projections from prefrontal cortex to the posterior association cortex appear to serve the executive control of voluntary recall. PMID- 10537109 TI - L-type calcium channels and GSK-3 regulate the activity of NF-ATc4 in hippocampal neurons. AB - The molecular basis of learning and memory has been the object of several recent advances, which have focused attention on calcium-regulated pathways controlling transcription. One of the molecules implicated by pharmacological, biochemical and genetic approaches is the calcium/calmodulin-regulated phosphatase, calcineurin. In lymphocytes, calcineurin responds to specific calcium signals and regulates expression of several immediate early genes by controlling the nuclear import of the NF-ATc family of transcription factors. Here we show that NF ATc4/NF-AT3 in hippocampal neurons can rapidly translocate from cytoplasm to nucleus and activate NF-AT-dependent transcription in response to electrical activity or potassium depolarization. The calcineurin-mediated translocation is critically dependent on calcium entry through L-type voltage-gated calcium channels. GSK-3 can phosphorylate NF-ATc4, promoting its export from the nucleus and antagonizing NF-ATc4-dependent transcription. Furthermore, we show that induction of the inositol 1,4,5-trisphosphate receptor type 1 is controlled by the calcium/calcineurin/NF-ATc pathway. This provides a new perspective on the function of calcineurin in the central nervous system and indicates that NF-AT mediated gene expression may be involved in the induction of hippocampal synaptic plasticity and memory formation. PMID- 10537110 TI - Two subsets of memory T lymphocytes with distinct homing potentials and effector functions. AB - Naive T lymphocytes travel to T-cell areas of secondary lymphoid organs in search of antigen presented by dendritic cells. Once activated, they proliferate vigorously, generating effector cells that can migrate to B-cell areas or to inflamed tissues. A fraction of primed T lymphocytes persists as circulating memory cells that can confer protection and give, upon secondary challenge, a qualitatively different and quantitatively enhanced response. The nature of the cells that mediate the different facets of immunological memory remains unresolved. Here we show that expression of CCR7, a chemokine receptor that controls homing to secondary lymphoid organs, divides human memory T cells into two functionally distinct subsets. CCR7- memory cells express receptors for migration to inflamed tissues and display immediate effector function. In contrast, CCR7+ memory cells express lymph-node homing receptors and lack immediate effector function, but efficiently stimulate dendritic cells and differentiate into CCR7- effector cells upon secondary stimulation. The CCR7+ and CCR7- T cells, which we have named central memory (TCM) and effector memory (TEM), differentiate in a step-wise fashion from naive T cells, persist for years after immunization and allow a division of labour in the memory response. PMID- 10537111 TI - Atomic structure of the GCSF-receptor complex showing a new cytokine-receptor recognition scheme. AB - Granulocyte colony-stimulating factor (GCSF) is the principal growth factor regulating the maturation, proliferation and differentiation of the precursor cells of neutrophilic granulocytes and is used to treat neutropenia. GCSF is a member of the long-chain subtype of the class 1 cytokine superfamily, which includes growth hormone, erythropoietin, interleukin 6 and oncostatin M. Here we have determined the crystal structure of GCSF complexed to the BN-BC domains, the principal ligand-binding region of the GCSF receptor (GCSFR). The two receptor domains form a complex in a 2:2 ratio with the ligand, with a non crystallographic pseudo-twofold axis through primarily the interdomain region and secondarily the BC domain. This structural view of a gp130-type receptor-ligand complex presents a new molecular basis for cytokine-receptor recognition. PMID- 10537112 TI - Structural evidence for dimerization-regulated activation of an integral membrane phospholipase. AB - Dimerization is a biological regulatory mechanism employed by both soluble and membrane proteins. However, there are few structural data on the factors that govern dimerization of membrane proteins. Outer membrane phospholipase A (OMPLA) is an integral membrane enzyme which participates in secretion of colicins in Escherichia coli. In Campilobacter and Helicobacter pylori strains, OMPLA is implied in virulence. Its activity is regulated by reversible dimerization. Here we report X-ray structures of monomeric and dimeric OMPLA from E. coli. Dimer interactions occur almost exclusively in the apolar membrane-embedded parts, with two hydrogen bonds within the hydrophobic membrane area being key interactions. Dimerization results in functional oxyanion holes and substrate-binding pockets, which are absent in monomeric OMPLA. These results provide a detailed view of activation by dimerization of a membrane protein. PMID- 10537113 TI - The reaction cycle of isopenicillin N synthase observed by X-ray diffraction. AB - Isopenicillin N synthase (IPNS), a non-haem iron-dependent oxidase, catalyses the biosynthesis of isopenicillin N (IPN), the precursor of all penicillins and cephalosporins. The key steps in this reaction are the two iron-dioxygen-mediated ring closures of the tripeptide delta-(L-alpha-aminoadipoyl)-L-cysteinyl-D-valine (ACV). It has been proposed that the four-membered beta-lactam ring forms initially, associated with a highly oxidized iron(iv)-oxo (ferryl) moiety, which subsequently mediates closure of the five-membered thiazolidine ring. Here we describe observation of the IPNS reaction in crystals by X-ray crystallography. IPNS Fe2+ substrate crystals were grown anaerobically, exposed to high pressures of oxygen to promote reaction and frozen, and their structures were elucidated by X-ray diffraction. Using the natural substrate ACV, this resulted in the IPNS x Fe2+ x IPN product complex. With the substrate analogue, delta-(L-alpha aminoadipoyl)-L-cysteinyl-L-S-methylcysteine (ACmC) in the crystal, the reaction cycle was interrupted at the monocyclic stage. These mono- and bicyclic structures support our hypothesis of a two-stage reaction sequence leading to penicillin. Furthermore, the formation of a monocyclic sulphoxide product from ACmC is most simply explained by the interception of a high-valency iron-oxo species. PMID- 10537114 TI - The ups and downs of leptin action. PMID- 10537115 TI - Distinct physiologic and neuronal responses to decreased leptin and mild hyperleptinemia. AB - Leptin acts on specific populations of hypothalamic neurons to regulate feeding behavior, energy expenditure, and neuroendocrine function. It is not known, however, whether the same neural circuits mediate leptin action across its full biologic dose-response curve, which extends over a broad range, from low levels seen during starvation to high levels characteristic of obesity. Here, we show that the characteristic fall in leptin with fasting causes a rise in neuropeptide Y (NPY) messenger RNA (mRNA), as well as a fall in POMC and cocaine and amphetamine-regulated transcript (CART) mRNAs. Sc infusion of leptin sufficient to maintain plasma levels within the physiologic range during the fast prevents changes in the expression of these peptides, as well as changes in neuroendocrine function, demonstrating that multiple neural circuits are highly sensitive to small changes in leptin within its low physiologic range. In contrast, a modest elevation of plasma leptin above the normal fed range by constant sc infusion, which produced marked reduction in food intake and body weight, decreased NPY mRNA in the arcuate hypothalamic nucleus but did not affect the levels of mRNAs encoding the anorexigenic peptides alpha-MSH, CART or CRH. These results suggest that the dose response characteristics of leptin on hypothalamic target neurons at the level of mRNA expression are variable, with some neurons (e.g. NPY) responding across a broad dose range and others (e.g. POMC and CART) showing a limited response within the low range. These results further suggest that the central targets of leptin that mediate the transition from starvation to the fed state may be distinct from those that mediate the response to overfeeding and obesity. PMID- 10537116 TI - Susceptibility and resistance to experimental allergic encephalomyelitis: relationship with hypothalamic-pituitary-adrenocortical axis responsiveness in the rat. AB - Susceptibility to experimental allergic encephalomyelitis (EAE) may be influenced by variations in the production of endogenous glucocorticoids. We investigated whether this concept is consistent across different genotypes and paradigms of EAE. In the major histocompatibility complex-disparate rat strains, Lewis (LEW), Brown Norway (BN), and Dark Agouti (DA), inflammatory and inflammatory demyelinating variants of EAE were induced by immunization with myelin basic protein and myelin oligodendrocyte glycoprotein, respectively. We analyzed hormone production in EAE and after exposure to novel environment. DA and BN rats showed a robust hypothalamic-pituitary-adrenocortical (HPA) axis response to novelty stress and produced significantly higher ACTH and corticosterone plasma levels compared with LEW rats. However, HPA axis responsiveness was not associated with a generalized resistance to EAE, as both DA and LEW rats were susceptible to myelin basic protein-induced EAE. Moreover, both robust HPA responder strains, DA and the EAE-resistant BN rat, were highly susceptible to myelin oligodendrocyte glycoprotein-induced EAE. In animals of all strains, clinical disease was associated with significantly elevated plasma levels of corticosterone, and no differences in brain glucocorticoid-binding receptors were detected. Therefore, HPA axis characteristics are not a predictor of disease susceptibility in EAE. PMID- 10537117 TI - Expression of functional leptin receptors in rodent Leydig cells. AB - Several studies indicate that the size of body fat stores and the circulating levels of the adipocyte-derived hormone leptin are able to influence the activity of the hypothalamic-pituitary-gonadal axis. The leptin-hypothalamic-pituitary gonadal interactions have been mainly studied at the level of the central nervous system. In this study, we investigated the possibility that leptin may have direct effects on the rodent Leydig cell function. To probe this hypothesis, we first analyzed the expression of leptin receptors (OB-R) in rodent Leydig cells in culture. RT-PCR studies showed that rat Leydig cells express both the long (OB Rb) and short isoform (OB-Ra) of leptin receptor, whereas MLTC-1 cells (a murine Leydig tumor cell line) express only the long isoform. Short-term (30-90 min) incubation of rat Leydig cells with increasing concentrations ofleptin (2-500 ng/ml) led to a significant and dose-dependent inhibition of human (h)CG stimulated testosterone (T) production (approximately 60% reduction, IC50 = 20 ng/ml) but no change in basal androgen release. Also, leptin (150 ng/ml) amplified hCG-induced intracellular cAMP formation (1- to 2-fold) without modifying basal cAMP levels. Subsequent experiments showed that leptin inhibited 8Br-cAMP-stimulated T production, indicating that leptin's effect is exerted beyond cAMP. The inhibitory effect of leptin on hCG-induced T secretion was accompanied by a significant reduction of androstenedione and a concomitant rise of the precursor metabolites pregnenolone, progesterone, and 17-OH-progesterone, conceivable with a leptin-induced lesion of 17,20 lyase activity. Separate experiments performed with the MLTC-1 cells (not expressing cytochrome P450 17alpha) showed that leptin, though amplifying hCG-stimulated cAMP production, did not modify hCG-stimulated pregnenolone and progesterone release. These results further indicate that leptin action on steroidogenesis occurs downstream of progesterone synthesis. Northern Blot experiments showed no acute effect of leptin on cytochrome P450-17alpha messenger RNA accumulation in rat Leydig cells in basal and hCG-stimulated conditions, excluding that the rapid changes observed were caused by messenger RNA degradation. In conclusion, these findings, for the first time, show that leptin has direct, receptor-mediated actions on rodent Leydig cells in culture, at concentrations within the range of obese men. PMID- 10537119 TI - Targeted overexpression of galanin in lactotrophs of transgenic mice induces hyperprolactinemia and pituitary hyperplasia. AB - We generated transgenic mice that carry 4.6 kb of the mouse galanin gene fused to 2.5 kb of the rat PRL promoter. In all transgenic lines that carried and transmitted the transgene, there were significant increases in galanin messenger RNA and peptide levels in the anterior pituitary in both male and female transgenic mice, and the elevation of galanin was restricted to the anterior lobe. Furthermore, galanin release from pituitary cells in vitro of both male and female transgenic mice was dramatically increased compared with that in control mice. At 2-4 months of age, pituitary PRL contents in female transgenic mice were increased compared with those in normal controls. Moreover, PRL messenger RNA levels were increased in female transgenic mice. However, plasma levels of PRL in female transgenic mice were not significantly higher until 6 months of age. By 11 months of age, cell numbers in the anterior pituitary were increased in female, but not male, transgenic mice. The percentage of lactotrophs in female transgenic mice as well as PRL gene expression per cell were significantly higher. No differences were detected in PRL content, gene expression, or release between normal and transgenic male mice. Six weeks of estrogen treatment significantly increased anterior pituitary weights and PRL secretion in male transgenic mice compared with that in normal male mice. In addition, anterior pituitary weights and PRL secretion were decreased in female transgenic mice compared with controls 6 weeks after ovariectomy. We conclude that overexpression of galanin in lactotrophs stimulates PRL synthesis and secretion and acts as a growth factor resulting in the formation of pituitary hyperplasia and hyperprolactinemia. Furthermore, estrogen appears critical for these galanin-mediated events. PMID- 10537118 TI - Thyroid hormone export in rat FRTL-5 thyroid cells and mouse NIH-3T3 cells is carrier-mediated, verapamil-sensitive, and stereospecific. AB - Export of L-T3 out of the cell is one factor governing the cellular T3 content and response. We previously observed in liver-derived cells that T3 export was inhibited by verapamil, suggesting that it is due to either ATP-binding cassette/multidrug resistance (MDR1/mdr1b) or multidrug resistance-related (MRP1/mrp1) proteins. To test this hypothesis we measured T3 export in FRTL-5, NIH-3T3, and rat hepatoma (HTC) cells that varied in expression of these proteins. FRTL-5 and NIH-3T3 cells were found to contain a T3 efflux mechanism that is verapamil inhibitable, saturable, and stereospecific. By contrast, T3 efflux in HTC cells was slow and unaffected by verapamil. Neither FRTL-5 nor NIH 3T3 cells express mdrlb, but all three cell types express mrpl, as assessed by immunoblotting. Overexpression of MDR1 in NIH-3T3 cells did not enhance verapamil inhibitable T3 efflux. Photoaffinity labeling of FRTL-5 and NIH-3T3 cells with [125I]L-T3 revealed a labeled 90- to 100-kDa protein that was not present in HTC cells. Verapamil and excess nonradioactive L-T3, but not D-T3, inhibited labeling of this protein. The lack of correlation between T3 efflux and MDR1 and mrpl expression and the finding of a photoaffinity-labeled putative transport protein smaller than MDR1 or mrp1 protein (approximately 170 kDa) suggest that a novel protein is involved in the transport of T3 out of cells. PMID- 10537120 TI - A role of insulin-like growth factor I in luteinizing hormone receptor expression in granulosa cells. AB - The present study was undertaken to identify the mechanisms underlying the effect of insulin-like growth factor (IGF-I) on LH receptor in rat granulosa cells. Treatment with FSH, as expected, produced a substantial increase in LH receptor messenger RNA (mRNA) level, and concurrent treatment with increasing concentrations of IGF-I brought about dose-dependent increases in FSH-induced LH receptor mRNA, with a maximal response 2.5-fold greater than that induced by FSH alone. IGF-I, either alone or in combination with FSH, did not affect intracellular cAMP levels, whereas it enhanced the effect of 8-bromo-cAMP on LH receptor mRNA production. We then investigated whether the effects of IGF-I and FSH on LH receptor mRNA levels are the results of increased transcription and/or altered mRNA stability. To determine whether the LH receptor 5'-flanking region plays a role in directing LH receptor mRNA expression, the proximal area of the LH receptor 5'-flanking regions were inserted into a transient expression vector, pGL-Basic, which contains luciferase as the reporter gene, and the resulting plasmids were transiently transfected into rat granulosa cells. Our studies show that the FSH-induced luciferase activity varied dependent upon the length of the 5'-flanking region sequence in the reporter gene. In addition, FSH (30 ng/ml) significantly enhanced the activity of 1379 bp of the LH receptor 5'-flanking region, but treatment with 10 ng/ml IGF-I alone did not significantly influence the activity of the LH receptor promoter or affect the increased promoter activity induced by FSH. The rates of LH receptor mRNA gene transcription, assessed by nuclear run-on transcription assay, were not increased by the addition of IGF-I. On the other hand, the decay curves for LH receptor mRNA transcript in primary granulosa cells showed a significant increase in the half life after the addition of IGF-I. These data suggest a possible role for changes in LH receptor mRNA stability in the IGF-I-induced regulation of LH receptor in rat granulosa cells. This interface between circulating hormones and paracrine/autocrine systems could provide an important mechanism to amplify the effects of gonadotropic hormones at the local level. PMID- 10537121 TI - Studies of the N-terminal region of a parathyroid hormone-related peptide (1-36) analog: receptor subtype-selective agonists, antagonists, and photochemical cross linking agents. AB - The N-terminal regions of PTH and PTH-related peptide (PTHrP) are involved in receptor-mediated signaling and subtype selectivity. To better understand the molecular basis for these processes, we first prepared a series of [I5,W23,Y36] PTHrP(1-36)NH2 analogs having stepwise deletions of residues 1-4 and characterized them with the human (h)PTH-1 and hPTH-2 receptor subtypes stably transfected in LLC-PK1 cells. Deletions beyond residue 2 caused progressive and severe losses in cAMP-signaling efficacy without dramatically diminishing receptor-binding affinity; consistent with this, [I5,W23]-PTHrP(5-36) was a potent antagonist for both PTH receptor subtypes. We then prepared and characterized photolabile analogs of [I5,W23,Y36]-PTHrP(1-36)NH2 that were singly modified with parabenzoyl-L-phenylalanine (Bpa) along the first six residues. These full-length analogs exhibited receptor subtype-selective agonism, antagonism, and photochemical cross-linking profiles. In particular, the [Bpa2]- and [Bpa4]-substituted analogs selectively antagonized and preferentially cross linked to the PTH-1 receptor and PTH-2 receptor, respectively. These results demonstrate that the 1-5 region of [I5,W23]-PTHrP(1-36) is critical for activating the PTH-1 and PTH-2 receptors and suggest that the individual residues in this region play distinct roles in modulating the activation states of the two receptors. The cross-linking of both agonist and antagonist ligands to these PTH receptors lays the groundwork for identifying critical signaling determinants in the ligand binding pocket of the receptor. PMID- 10537122 TI - Rescue of the skeletal phenotype of vitamin D receptor-ablated mice in the setting of normal mineral ion homeostasis: formal histomorphometric and biomechanical analyses. AB - 1,25-Dihydroxyvitamin D3 has been shown to play an important role in vitro in regulating osteoblast gene transcription and promoting osteoclast differentiation. To address the role of the vitamin D receptor (VDR) in skeletal homeostasis, formal histomorphometric analyses were performed in VDR null mice in the setting of impaired mineral ion homeostasis as well as in VDR null mice in whom normal mineral ion homeostasis had been preserved. In hypocalcemic VDR null mice, there was an increase in bone volume as a result of a dramatic increase in osteoid. There was also an increase in the number of osteoblasts without a significant change in the number of osteoclasts. Examination of the growth plate revealed marked disorganization, with an increase in vascularity and matrix. Biomechanical parameters demonstrated increased bone fragility in the hypocalcemic VDR null mice. In the VDR ablated mice in whom normal mineral ion homeostasis had been preserved, none of these measurements was significantly different from those in wild-type littermates raised under identical conditions. Notably, the morphology and width of the growth plate were indistinguishable from those in wild-type controls, demonstrating that a calcium/phosphorus/lactose enriched diet started at 16 days of age in the VDR null mice permits the development of both normal morphology in the growth cartilage and adjacent metaphysis and normal biomechanical competence of cortical bone. Thus, the principle action of the VDR in skeletal growth, maturation, and remodeling is its role in intestinal calcium absorption. The skeletal consequences of VDR ablation are a result of impaired intestinal calcium absorption and/or the resultant secondary hyperparathyroidism and hypophosphatemia. PMID- 10537123 TI - Transcriptional and translational regulation of angiotensin II type 2 receptor by angiotensin II and growth factors. AB - The regulatory effects of angiotensin-II (AngII) and several growth factors, including insulin-like growth factor 1 (IGF-1), basic fibroblast growth factor (bFGF), and transforming growth factor beta1 (TGFbeta1) on the AngII subtype 2 (AT2) receptor were studied using R3T3 cells, a mouse fibroblast cell line that expresses only AT2 receptors. AngII increased (in a time- and dose-dependent manner) AT2 binding sites but had no effects on AT2 messenger RNA (mRNA) levels. At maximal concentration (10(-7) M) AngII caused a 4-fold increase of AT2 receptor number. In contrast, IGF-1 increased (3- to 4-fold), whereas bFGF and TGFbeta1 decreased (by about 90% and 80%, respectively) AT2 receptor and mRNA levels. Moreover, AngII potentiated the effect of IGF-1 on receptor number, but not on AT2 mRNA levels, and significantly reduced the inhibitory action of bFGF and TGFbeta1 on AT2 binding sites but not on AT2 mRNA levels. None of these factors modified AT2 mRNA half-life. The potential effects of these factors on transcription of the AT2 gene were measured by means of nuclear run-on assays. IGF-1 increased the rate of transcription by about 2.5-fold, whereas bFGF and TGFbeta1 reduced it by 90 and 80%, respectively. In contrast, AngII did not modify either the basal or IGF-1-stimulated transcription rate. Finally, AngII alone or together with IGF-1, but not IGF-1 alone, increased the attachment of AT2 mRNA to polysomal fractions. The present findings demonstrate that the main mechanism by which AngII regulates the AT2 receptor is by increasing the rate of AT2 mRNA translation, whereas the stimulatory (IGF-1) or inhibitory (bFGF and TGFbeta1) effects of these growth factors on AT2 expression are exerted at the transcriptional level. PMID- 10537124 TI - Colocalization of neurotensin messenger ribonucleic acid (mRNA) and progesterone receptor mRNA in rat arcuate neurons under estrogen-stimulated conditions. AB - In the female rat, estrogen and progesterone directly or indirectly regulate the activity of neurotensin (NT)-synthesizing neurosecretory cells located in the hypothalamic arcuate nucleus (ARC). To determine whether these NT neurons are subject to direct regulation by ovarian steroids, estrogen-inducible messenger RNA (mRNA) encoding nuclear progesterone receptor (PR) was used as a cellular marker for nuclear estrogen receptor (ER) as well as PR, and double label in situ hybridization was employed to determine the extent to which NT/neuromedin N mRNA and PR mRNA are colocalized in ARC neurons under estrogen-stimulated conditions. In estradiol-treated ovariectomized rats, approximately 80% of NT/neuromedin N mRNA-expressing cells in sections through the dorsomedial division of the ARC and approximately 60% of such cells in sections through the ventrolateral division of the ARC were found to contain PR mRNA. Depending on the ARC division and rostrocaudal level, double labeled cells accounted for approximately 20-50% of PR mRNA-containing cells. These results indicate that under estrogen-stimulated conditions the majority of NT neurons in the ARC express both PR and ER, as previous studies of this region indicate that estrogen-inducible PR occurs only in cells that also express ER. In the rat, NT neurons appear to be a major ARC cell type subject to direct regulation by estrogen and progesterone. PMID- 10537125 TI - Regulation of the mouse preprothyrotropin-releasing hormone gene by retinoic acid receptor. AB - Retinoic acid (RA) has been reported to inhibit the secretion and synthesis of the pituitary TSH in vivo and in vitro. However, little is known about the influence of RA on the expression of the prepro-TRH gene. We therefore investigated whether the promoter activity of the mouse TRH gene is directly regulated by RA using a transient transfection assay into CV-1 cells. In the absence of cotransfected RA receptor (RAR), all-trans-RA did not affect the promoter activity. In contrast, the cotransfected RARalpha significantly stimulated promoter activity in the absence of ligand, and all-trans-RA reversed basal promoter activation. The cotransfected thyroid hormone receptor-beta (TRbeta), but not 9-cis-RA receptor (RXR), had an additive effect on the RAR dependent stimulation. TR and RAR can similarly interact with the corepressor proteins, and the cotransfected nuclear receptor corepressor (N-CoR) has been demonstrated to augment the transcriptional stimulation of the TRH gene by unliganded TR. As observed with TR, the coexpression of a N-CoR variant significantly enhanced the ligand-independent stimulation by RAR. A mutant RAR (RAR403) lacking the C-terminal activation function-2 (AF-2) activation domain that was essential for ligand-induced corepressor release constitutively stimulated the promoter activity. The constitutive stimulation by RAR403 was augmented by the cotransfected N-CoR variant. A deletion analysis of the 5' flanking region of the TRH gene revealed that the minimal promoter region for the regulation by RAR was -83 to +53, with a consensus half-site motif for the thyroid hormone response element at -57. In contrast to the strong binding of TR to the thyroid hormone response element half-site in gel retardation assays, no binding of RAR homodimer, RAR/ RXR heterodimer, or RAR/TR heterodimer was observed to the minimal promoter region. These results collectively suggest that RAR without heterodimerization with RXR and TR regulates transcription of the mouse TRH gene in cooperation with the corepressor, and that the DNA binding of RAR appeared to be unnecessary for regulation of the TRH gene promoter. PMID- 10537126 TI - Thyroid hormone regulates the extracellular organization of laminin on astrocytes. AB - Astrocytes produce laminin, a key extracellular matrix guidance molecule in the developing brain. Laminin is bound to transmembrane receptors on the surface of astrocytes known as integrins, which are, in turn, bound to the microfilament meshwork inside the astrocyte. Previous studies have shown that T4 regulates the pattern of integrin distribution in astrocytes by modulating the organization of the microfilaments. In this study, the effect of thyroid hormone on the secretion and topology of laminin in astrocytes was examined. Linear arrays of secreted laminin were observed on the surface of the T4-treated astrocytes within 10 h after seeding the cells onto poly-D-lysine-coated coverslips and became an organized meshwork by 24 h. In contrast, little if any laminin was identified on the surface of either hormone-deficient or T3-treated cells until 36 h after seeding and then was restricted to punctate deposits. Secretion of laminin into the medium by hormone-deficient and T3-treated cells was significantly greater than that by T4-treated cells. Conversely, deposition of laminin into the extracellular matrix was significantly greater in T4-treated cells than in hormone-deficient and T3-treated cells. Thyroid hormone had no effect on the production of laminin by astrocytes. These data show that T4 regulates the extracellular deposition and organization of laminin on the surface of astrocytes and provide a mechanism by which this morphogenic hormone can influence neuronal migration and axonal projection in the developing brain. PMID- 10537127 TI - Regulated, side-directed secretion of proguanylin from isolated rat colonic mucosa. AB - Guanylin, an activator of the guanylyl cyclase C receptor in the apical membrane of intestinal epithelium, modulates intestinal fluid and electrolyte transport. The bioactive 15-amino acid peptide originally isolated from rat intestine represents the C-terminal part of a longer, 115-residue prepropeptide. The aim of the present study was to characterize the direction and molecular form in which guanylin is secreted from the colonic mucosa, as well as the mechanisms that trigger its secretion. Isolated rat colonic mucosa was mounted in Ussing chambers, allowing the separate determination of apical and basolateral release. After HPLC purification, two different molecular forms of guanylin were identified in the apical incubation media by combining a bioassay for guanylyl cyclase C activation, a specific guanylin enzyme-linked immunosorbent assay and mass spectrometry, as well as sequence analysis: a bioactive form coeluting with synthetic 15-residue guanylin and the 94-residue propeptide, guanylin-22-115. The basal concentration of proguanylin at the apical side of epithelia was about 15 fold higher, compared with that of the small, bioactive peptide. In the basolateral incubation media, no proguanylin and only very low amounts of bioactive guanylin were detected. Incubation with carbachol led to a significant increase of about 7-fold in the release of proguanylin to both sides of the isolated epithelia. On the apical side, a concomitant increase of the small, bioactive peptide was observed; whereas, on the basolateral side, its concentration remained unchanged. Vasoactive intestinal peptide or the NO-donor S nitroso-N-acetylpenicillamine did not affect guanylin secretion. Our results suggest that, in the intestine, guanylin is secreted mainly to the luminal side of the epithelium. The peptide is released as a 94-residue propeptide, which is then processed to a smaller, bioactive form (luminocrine secretion). Carbachol stimulates the release of proguanylin to both sides of the intestinal mucosa, but a parallel increase in the bioactive C-terminal derivative only occurs on the apical side. In vivo, the basolateral release could be a source of circulating proguanylin, which might be processed proteolytically to the active peptide in distant target tissues (endocrine secretion). PMID- 10537128 TI - Ovulation in plasminogen-deficient mice. AB - Many different studies suggest that plasmin generated from plasminogen plays a crucial role in the degradation of the follicular wall at the time of ovulation. We have assessed the physiological relevance of plasmin on ovulation by studying plasminogen-deficient mice. Ovulation efficiency (mean number of ova released per mouse) was determined both in a standardized ovulation model in which 25-day-old immature mice were injected with finite amounts of gonadotropins to induce ovulation and during physiological ovulation using adult normally cycling mice. Our results revealed that the temporal onset of follicular wall rupture (first ova observed in bursa or oviduct) was not delayed in plasminogen-deficient mice during gonadotropin-induced ovulation. However, there was a trend toward slightly reduced ovulation efficiency in the plasminogen-deficient mice. This reduction was only 13% and not statistically significant (P = 0.084) and may be connected to a delayed maturation of these mice manifested in reduced body and ovary weights. During physiological ovulation adult plasminogen-deficient mice had normal ovulation efficiency compared with plasminogen wild-type mice. Taken together our results indicate that under the conditions used in this study plasmin is not required for efficient follicular rupture or for activation of other proteases involved in this process. Alternatively, the role of plasmin may be effectively compensated for by other mechanisms in the absence of plasmin. PMID- 10537129 TI - Response of bipotential human marrow stromal cells to insulin-like growth factors: effect on binding protein production, proliferation, and commitment to osteoblasts and adipocytes. AB - Insulin-like growth factors (IGFs) are important regulators of the activity of mature osteoblasts, but their effects on osteoprogenitor cells in human bone marrow stroma are unclear. In this study, we assessed the effects of IGFs on a conditionally immortalized human marrow stromal cell line, hMS(3-4), which has the ability to differentiate to either mature osteoblasts or adipocytes. hMS(3-4) cells expressed functional receptors for IGFs as well as specific IGF-binding proteins (IGFBP-3, -4, -5, and -6). IGF treatment of hMS(3-4) cells did not alter IGFBP expression, but resulted in distinct posttranslational modifications of secreted IGFBP-3 and IGFBP-4 proteins. IGF-I, IGF-II, and their receptor activating analogs significantly increased by 2-fold the proliferation rate of the hMS(3-4) cells, but had a more complex effect on hMS(3-4) cell differentiation. Treatment with IGFs did not affect gene expression of Cbfa1 or peroxisome proliferator-activated receptor gamma2 (transcription factors involved in commitment to osteoblast and adipocyte pathways, respectively), alkaline phosphatase, type I collagen, and osteocalcin (markers of the osteoblast lineage), or lipoprotein lipase and adipsin (markers of the adipocyte lineage) and did not change alkaline phosphatase activity or type I collagen and osteocalcin protein relative to total protein production. In contrast, IGFs significantly increased type I collagen expression in differentiated hMS(3-4) cells as well as mature osteoblasts and promoted lipid accumulation in differentiated adipocytes. In summary, hMS(3-4) cells express essential components of the IGF system and respond to IGF treatment with increased proliferation. There was no evidence for IGFs directly modulating the commitment of hMS(3-4) cells to either osteoblast or adipocyte pathways, and their effects on differentiation within these lineages were dependent on the stage of cell maturation. PMID- 10537130 TI - Estrogen directly respresses gonadotropin-releasing hormone (GnRH) gene expression in estrogen receptor-alpha (ERalpha)- and ERbeta-expressing GT1-7 GnRH neurons. AB - Estrogen has wide-ranging and complex effects on the reproductive axis, which are often difficult to interpret from in vivo studies. Estrogen negatively regulates tonic GnRH synthesis and also plays a pivotal role in the positive regulation of GnRH necessary for the preovulatory surge. To dissect the mechanisms by which these divergent effects occur, we attempted to observe the direct action of estrogen on the regulation of GnRH messenger RNA (mRNA) levels using the well characterized, GnRH-secreting, hypothalamic cell line, GT1-7. Using RT-PCR, we first investigated estrogen receptor transcript expression in GT1-7 neurons. We found that the GT1-7 cells express both estrogen receptor-alpha (ERalpha) and the recently described ERbeta mRNAs. We also detected the presence of both receptor subtypes in the GT1-7 neurons by Western blot analysis using specific ER antibodies. By Northern blot analysis of total GT1-7 RNA, we found that 17beta estradiol (1 nM) down-regulates GnRH mRNA levels to approximately 55% of basal levels over a 48-h time course. This effect appears to occur specifically through an ER-mediated mechanism, as ICI 182,780, a complete ER antagonist, blocks the repression of GnRH mRNA levels by estradiol. The recently reported ERalpha specific agonist/ERbeta-specific antagonist 2,2-bis-(p-hydroxyphenyl-1,1,1 trichloroethane (HPTE), a methoxychlor metabolite, also down-regulated GnRH gene expression. The repression of GnRH mRNA levels appears to occur at the transcriptional level, as simian virus 40 T antigen mRNA expression, which is under the control of 2.3 kb of the rat GnRH 5'-regulatory region, mimics the down regulation of GnRH after treatment with estradiol. As the rat GnRH regulatory region in GT1-7 neurons does not appear to harbor a classic estrogen response element, the mechanism involved in the repression of GnRH has yet to be determined. These results suggest that estradiol directly regulates GnRH gene expression at the level of the GnRH neuron and may exert its neuroendocrine control through direct interaction with specific receptors expressed in these cells. PMID- 10537131 TI - Molecular mechanisms of androgen-independent growth of human prostate cancer LNCaP-AI cells. AB - The goal of this study is to investigate the molecular mechanisms of androgen independent growth in prostate cancer. We have established an androgen independent prostatic carcinoma LNCaP-AI (defined as a LNCaP cell line that is capable of growing in charcoal-stripped serum) from the androgen-dependent LNCaP FGC cells. In contrast to the androgen-independent PC-3 human prostate cancer cells, LNCaP-AI cells still express a similar level of androgen receptor as their parental cells and are sensitive to androgen stimulation. Compared with the parental LNCaP-FGC cells, LNCaP-AI cells are more resistant to apoptosis induced by 12-O-tetradecanoylphorbol-13-acetate and express a much higher level of antiapoptotic gene bcl-2 and cyclin-dependent kinase inhibitor p21, which may confer an enhanced antiapoptosis phenotype. On the other hand, expression of cyclin-dependent kinase inhibitor p16 is significantly reduced in the LNCaP-AI cells, implying the release of an inhibitory effect of p16 on cell cycle progression. Taken together, our results suggest that multiple factors contribute to the development of androgen-independent growth of prostatic carcinoma cells, including enhancement of cell antiapoptosis function, release of cell cycle inhibition, and stimulation of cell proliferation by alternative signaling pathways. PMID- 10537132 TI - Thiazolidinediones inhibit osteoclast-like cell formation and bone resorption in vitro. AB - Osteoblasts and adipocytes are derived from common bone marrow stromal cells that play crucial roles in the generation of osteoclasts. Activation of peroxisome proliferator-activated receptor-gamma (PPARgamma) induces adipogenic differentiation of stromal cells; however, whether this would affect osteoblast/osteoclast differentiation is unknown. Thus, we examined the effects of the thiazolidinedione (TZD) class of antidiabetic agents that activate PPARgamma on osteoblast/osteoclast differentiation using mouse whole bone marrow cell culture. As reported, all TZDs we tested (troglitazone, pioglitazone, and BRL 49653) markedly increased the number of Oil Red O-positive adipocytes and the expression of adipsin and PPARgamma 2. 1alpha,25-Dihydroxyvitamin D3 [1,25 (OH)2D3] did not affect adipogenic differentiation induced by TZDs. TZDs did not affect alkaline phosphatase activity, an early marker of osteoblastic differentiation, despite their marked adipogenic effects. TZDs decreased the number of tartrate-resistant acid phosphatase-positive multinucleated osteoclast like cells induced by 1,25-(OH)2D3 or PTH. Troglitazone dose dependently inhibited basal and 1,25-(OH)2D3- and PTH-induced bone resorption as assessed by pit formation assay. Interleukin-11 blocked the induction by troglitazone of adipogenesis, but had no effect on the inhibition of osteoclast-like cell formation. These results indicate that TZDs are potent inhibitors of bone resorption in vitro. Inhibitory effects of TZDs on osteoclastic bone resorption was not osteotropic factor specific and did not appear to be related to their adipogenic effects. Thus, TZDs may suppress bone resorption in diabetic patients and prevent bone loss. PMID- 10537133 TI - The mutant growth hormone-releasing hormone (GHRH) receptor of the little mouse does not bind GHRH. AB - The little mouse is a dwarf strain characterized by low levels of GH, pituitary hypoplasia, and an unresponsiveness to treatment with exogenous GHRH. The defect has been mapped to a missense mutation in the GHRH receptor gene that abolishes the function of the receptor, but the mechanism of this inactivation is unknown. Receptor function might be affected at the level of protein expression, maturation and processing, localization to the cell surface, ligand binding, or signaling. In this study, Western blots, using antiserum raised against the GHRH receptor and immunoprecipitation analysis of epitope-tagged receptors, demonstrate that both wild-type and mutant receptor proteins are expressed at equivalent levels in transfected cells. Immunofluorescence analysis of intact and permeabilized cells expressing the epitope-tagged receptors suggests that wild type and little mouse receptors are similarly localized to the cell surface. A species homologous binding assay was developed and used to show that 125I-mouse GHRH binds with high affinity to the wild-type mouse receptor but not to the little mutant receptor. Consistent with this, the mutant receptor fails to stimulate intracellular cAMP accumulation. Our results demonstrate that the little mutation does not dramatically affect the expression level, glycosylation, or cellular localization of the receptor protein but that it blocks specific GHRH binding, and therefore, signaling does not take place. PMID- 10537134 TI - Insulin-like growth factor I is essential for terminal end bud formation and ductal morphogenesis during mammary development. AB - Previous studies from this laboratory have emphasized the essential role of GH in pubertal mammary development and shown that insulin-like growth factor I (IGF-I) was capable of substituting for GH in this process in rats and mice. The present study shows that, even when GH is present, no mammary development is possible unless IGF-I is present. IGF-I(-/-) null female animals were found to have significantly less mammary development than age matched wild-type controls (P <0.006) using several endpoints including the number of terminal end buds or TEBs (1.3 vs. 7.3), percent of the fat pad occupied by glandular elements (6.5 vs. 100), and number of ducts (15 vs. too numerous to count). That the deficiency in mammary gland development was related to the absence of IGF-I was underscored by the observation that des (1-3) IGF-I administration to IGF-I(-/-) null animals for 5 days caused significant mammary gland development as measured by TEB formation and branching of ducts. The number of TEBs rose from a mean of 1.3 in controls to 20.5 without added E2 (P < 0.009), and from 1.7 to 21 when des (1-3) IGF-I was given together with E2 (P < 0.006). The number of ducts increased significantly from a mean of 12 to 27 in response to IGF-I and E2, and from 15 to 24.5 with IGF-I alone. In contrast, administration of human GH with E2 had no stimulatory effect on mammary development in these animals, indicating that the full effect of GH in this process is mediated by IGF-I. To determine whether IGF I was also responsible for further ductal morphogenesis, we administered des (1 3) IGF-I + E2 to the knockout animals for 14 days and compared the effects of this combination of hormones on mammary development with those observed after 5 days. We found that there was a significant increase from 5 to 14 days in the number of TEBs (mean: 21 vs. 41) and the area of the mammary fat pad occupied by glands (mean: 10 vs. 20%). There was elongation and thickening of the ducts which accounted for the increased area that was occupied by ductal structures. There was no significant increase in the number of ducts. However, there was the appearance of a large number of buds along the length of the ductal structures, suggesting the beginning of side branching. These results suggest that IGF-I, when given along with E2, is responsible for ductal morphogenesis. PMID- 10537135 TI - Water drinking in rats resulting from intravenous relaxin and its modification by other dipsogenic factors. AB - The purpose of the study was to determine whether iv infusion of relaxin would acutely stimulate water drinking in rats and, if it did, whether such drinking is affected by other dipsogenic stimuli or is blocked by centrally administered losartan. iv infusions of human gene 2 relaxin at doses of 25, 40, 55, or 80 microg/kg x h for 1 h induced dose-dependent water drinking in both male and female rats within 15-30 min of commencement of infusions. iv infusion of a nondipsogenic dose of angiotensin II (0.5 microg/h), combined with relaxin (40 microg/kg x h), almost tripled the relaxin-induced water intake. iv infusion of hypertonic (1 M) NaCl did not potentiate relaxin-induced drinking. Intracerebroventricular injection of the angiotensin AT1 antagonist losartan (10 microg) reduced water drinking induced by iv infusion of relaxin. The water drinking induced by iv infusion of relaxin in the rat suggests that blood-borne relaxin may be a dipsogenic hormone. Potentiation of this relaxin-induced drinking by moderate levels of circulating angiotensin II is additional evidence in support of this view. The results also indicate that a central angiotensinergic neural pathway, utilizing AT1 receptors, subserves relaxin induced drinking. PMID- 10537136 TI - Dissociation of Janus kinase 2 and signal transducer and activator of transcription 5 activation after treatment of Nb2 cells with a molecular mimic of phosphorylated prolactin. AB - We have previously demonstrated that phosphorylated PRL acts as an antagonist to the Nb2 proliferative activities of unmodified PRL. A molecular mimic of phosphorylated PRL, which substitutes an aspartate residue for the normally phosphorylated serine (serine 179), has the same properties. Because it takes less than one fourth the amount of phosphorylated hormone, or the aspartate mutant, to block the proliferative activity of unmodified hormone, we have investigated whether the high potency of the aspartate mutant is achieved by the production of an alternate and interfering intracellular signal cascade. Nb2 cells were exposed to 5 or 500 ng/ml human NIDDK PRL, wild-type recombinant PRL (unmodified PRL), or aspartate mutant PRL (pseudophosphorylated PRL) for 1, 5, or 10 min at 37 C. At 5 ng/ml and 10 min, wild-type recombinant PRL showed greater activation of Janus kinase 2 (JAK 2) than the NIDDK preparation. This is consistent with a previous report of higher proliferative activity for the wild type hormone and is primarily a reflection of the presence of some phosphorylated hormone in the NIDDK preparation. At 500 ng/ml and 10 min, saturation eliminated any differences between responses to the two preparations. JAK 2 activation was not seen in response to the aspartate mutant at either concentration. Signal transducer and activator of transcription 5 (STAT 5) activation was, however, just as robust for the aspartate-treated cells as for the other two groups. Time course experiments eliminated the possibility that STAT 5 phosphorylation in response to the aspartate mutant was the result of JAK 2 activation at earlier time points. Experiments in the present study also interestingly showed preassociation of JAK 2 and STAT 5 in the absence of PRL and the absence of detectable phosphorylation of either JAK 2 or STAT 5. Like JAK 2, receptor phosphorylation was absent with the aspartate mutant. A comparison between STAT 5a and STAT 5b activation showed a marked reduction in STAT 5b phosphorylation in response to the aspartate mutant, with concomitant reduction in STAT 5a-STAT 5b heterodimers. STAT 5a activation, however, was indistinguishable between the wild type and aspartate mutant. We conclude that the nonproliferative aspartate mutant signals and activates STAT 5 without, or with minimal, use of JAK 2 or receptor phosphorylation. The wild-type proliferative PRL, on the other hand, uses receptor phosphorylation and JAK 2 activation. PMID- 10537137 TI - Prolactin (PRL)-like protein J, a novel member of the PRL/growth hormone family, is exclusively expressed in maternal decidua. AB - A search of a nonmouse, nonhuman, expressed sequence tag database for messenger RNAs in the PRL/GH family has identified a novel rat complementary DNA clone. The encoded protein, designated PRL-like protein J (PLP-J), is predicted to be synthesized as a precursor of 211 amino acids, modified by N-linked glycosylation, and secreted as a mature glycoprotein of 182 residues. PLP-J messenger RNA synthesis is limited to early pregnancy with abundant expression on day 7, slightly declining expression on day 9, and no detectable expression by day 11. Unlike most other PRL family members, PLP-J does not appear to be synthesized by placental trophoblasts but, rather, by decidual cells surrounding the implantation site. By sequence similarity to rat PLP-J, a murine clone was identified in a mouse expressed sequence tag database. Mouse PLP-J was used to map the gene to a 700-kb region of mouse chromosome 13 that includes other members of the PRL/GH family. PMID- 10537138 TI - Role of N-linked glycosylation on the function and expression of the human secretin receptor. AB - Secretin is a 27-amino acid long peptide hormone that regulates pancreatic water, bicarbonate, enzymes, and potassium ion secretion. The human secretin receptor (hSR) is a glycoprotein consisting of 440 amino acids, of which there are 5 putative N-linked glycosylation sites at positions Asn72, Asn100, Asn106, Asn128 (N-terminal ectodomain), and Asn291 (second exoloop). Through functional analysis of the hSR-transfected cells cultured in the presence of various glycosylation inhibitors, it was found that tunicamycin and castanospermine were able to significantly reduce the secretin-stimulated cAMP response. On the other hand, the effects of other inhibitors, swainsonine and deoxymannojirimycin, were much lower, suggesting that the high mannose-type carbohydrate side-chain is essential to the expression of a fully functional hSR. The role of individual N-linked glycosylation sites was studied by mutation analysis (Asn to Leu or Ser to Ala) coupled to measurements of cAMP accumulation and extracellular acidification rate. The ED50 values of the wild-type receptor in these two assay systems were 0.25 and 0.11 nM, respectively, and mutation at position 100, 106, or 291 did not affect either the ED50 values or the maximal responses in the two assays. However, the Asn72Leu and Ser74Ala mutations reduced the maximal responses and increased the ED50 values in both assays, suggesting that this site is a true glycosylation signal. This hypothesis was further supported by competitive binding studies, the same mutants were found to be defective in binding with [125I]secretin. To evaluate whether the change in receptor function of the mutants is caused by the change in the process of presenting the receptor to the cell surface, the mutants and the wild-type receptor were tagged with a c-Myc epitope at the C-termini. Using an anti-c-Myc monoclonal antibody and confocal microscopy, all of the mutant receptors were found to be expressed and delivered to the plasma membrane. PMID- 10537139 TI - Hypertension associated with decreased testosterone levels in natriuretic peptide receptor-A gene-knockout and gene-duplicated mutant mouse models. AB - Mice lacking the gene (Npr1) encoding the natriuretic peptide receptor A (NPRA) have hypertension with elevated blood pressure and cardiac hypertrophy. In particular, Npr1 gene-deficient male mice exhibit lethal vascular events similar to those seen in untreated human hypertensive patients. Serum testosterone levels tend to be lower in hypertensive male humans than in normal males without hypertension, but the genetic basis for this tendency remains unknown. To determine whether Npr1 gene function affects the testosterone level, we measured serum testosterone in male hypertensive mice lacking a functional Npr1 gene, wild type animals with two copies, and the gene-duplicated littermates expressing four copies of the gene. In the Npr1 gene-knockout (zero-copy) mice, the serum testosterone level was 62% lower than that in the two-copy control mice (80+/-10 ts. 120+/-14 ng/ml, respectively; P < 0.005). Serum testosterone in the four-copy mice was 144% (P < 0.005) of that in the two-copy wild-type control mice. To investigate the role of NPRA in testicular steroidogenesis, we analyzed atrial natriuretic peptide (ANP)-dependent guanylyl cyclase activation, accumulation of intracellular cGMP, and testosterone production in purified primary Leydig cells from animals with zero, two, or four copies of the Npr1 gene. Leydig cells lacking the Npr1 gene did not show ANP-stimulated guanylyl cyclase activation or cGMP accumulation and had no ANP-dependent testosterone production. ANP stimulation of Leydig cells from the four-copy males elicited a 2-fold greater production of cGMP compared to that in the two-copy wild-type counterparts (260+/ 12 vs. 126+/-7 pmol/l x 10(6) cells; P < 0.001). Similarly, ANP-dependent testosterone production in Leydig cells was nearly twice as high in four-copy mice as in two-copy wild-type controls (561+/-18 vs. 325+/-11 ng/l x 10(6) cells; P < 0.001). ANP-dependent guanylyl cyclase activation and production of cGMP in Leydig cells increased progressively with the number of Npr1 gene copies. Our results establish the existence of an alternate mechanism for testicular steroidogenesis that is stimulated by NPRA-dependent cGMP signaling, in addition to that mediated by gonadotropins, via a cAMP pathway. These findings demonstrate the role of Npr1 gene function in the maintenance of serum testosterone levels and testicular steroidogenesis and provide a genetic link between hypertension associated with decreased NPRA and low testosterone levels. PMID- 10537140 TI - Mitogen-activated protein (MAP) kinases are involved in interleukin-1 (IL-1) induced IL-6 synthesis in osteoblasts: modulation not of p38 MAP kinase, but of p42/p44 MAP kinase by IL-1-activated protein kinase C. AB - We previously reported that interleukin-1alpha (IL-1alpha)-induced activation of protein kinase C (PKC) via phosphatidylcholine-specific phospholipase C (PC-PLC) limits IL-6 synthesis induced by IL-1alpha itself in osteoblast-like MC3T3-E1 cells. In the present study, we further investigated the mechanism behind IL 1alpha-induced IL-6 synthesis in MC3T3-E1 cells. IL-1alpha time-dependently stimulated the phosphorylation of both p42/p44 mitogen-activated protein (MAP) kinase and p38 MAP kinase. PD98059, a specific inhibitor of the upstream kinase that activates p42/p44 MAP kinase, inhibited the IL-1alpha-induced IL-6 synthesis as well as the phosphorylation of p42/p44 MAP kinase induced by IL-1alpha. SB203580, a specific inhibitor of p38 MAP kinase, also reduced both the phosphorylation of p38 MAP kinase and the IL-6 synthesis. 1-Oleoyl-2 acetylglycerol, an activator of PKC, suppressed the IL-1alpha-induced IL-6 synthesis. Calphostin C, a specific inhibitor of PKC, or D-609, a specific inhibitor of PC-PLC, significantly enhanced the IL-1alpha-induced phosphorylation of p42/p44 MAP kinase without affecting the phosphorylation of p38 MAP kinase. The phosphorylation of p42/p44 MAP kinase by IL-1alpha was markedly increased in PKC-down-regulated MC3T3-E1 cells. Neither 12-O-tetradecanoylphorbol-13-acetate, known to be an activator of PKC, nor 1-oleoyl-2-acetylglycerol affected the phosphorylation of p38 MAP kinase induced by IL-1alpha. These results strongly suggest that IL-1alpha-induced IL-6 synthesis is mediated via activations of both p42/p44 MAP kinase and p38 MAP kinase in osteoblasts, and that PKC activated by IL-1alpha itself negatively regulates IL-6 synthesis at a point upstream from p42/p44 MAP kinase. PMID- 10537141 TI - Growth hormone, but not prolactin, maintains, low-level activation of STAT5a and STAT5b in female rat liver. AB - STAT5b, a member of the signal transducers and activators of transcription family, is activated in rat liver in response to the intermittent (pulsatile) plasma pattern of GH that is characteristic of adult males. Previous studies have shown that the near-continuous plasma GH pattern of adult female rats is associated with a dramatic down-regulation of the STAT5 activation pathway. The present study demonstrates the presence of a low-level STAT5 DNA-binding activity in adult female rat liver and investigates the hormonal factors required for its maintenance. PRL is not responsible for this low-level STAT5 activity, as demonstrated in experiments involving estrus cycle monitoring (to investigate a possible role of the proestrus PRL surge), implantation of bromocriptine pellets (to eliminate PRL release by the pituitary), and direct injection of purified PRL. Rather, the low-level STAT5 activity is shown to result from chronic plasma GH stimulation, as demonstrated by GH infusion studies carried out in hypophysectomized rats. Furthermore, gel mobility supershift experiments demonstrate that the same STAT5-containing DNA-binding complexes are formed by both male and female adult rat liver extracts, albeit at approximately 10- to 20 fold lower levels by the female extracts. This DNA-binding activity is primarily comprised of STAT5b but also contains STAT5a, which is shown to be preferentially activated by the male plasma GH pattern in a manner similar to STAT5b. Thus, the dominance of activated STAT5b, compared with STAT5a, in the strong DNA-binding complexes formed in adult male rat liver nuclear extracts, is a reflection of the relative abundance in liver of the two STAT5 forms and is not attributable to an intrinsic, preferential activation of STAT5b by plasma GH pulses. The physiological significance of the low-level activated STAT5a and STAT5b seen in female rat liver and its effects on liver gene expression are uncertain but may involve the activation of female-expressed cytochromes P450 and other liver genes. PMID- 10537142 TI - DOTA-lanreotide: a novel somatostatin analog for tumor diagnosis and therapy. AB - Long acting somatostatin-14 (SST) analogs are used clinically to inhibit tumor growth and proliferation of various tumor types via binding to specific receptors (R). We have developed a 111In-/90Y-labeled SST analog, DOTA-(D)betaNal1 lanreotide (DOTALAN), for tumor diagnosis and therapy. 111In-/90Y-DOTALAN bound with high affinity (dissociation constant, Kd, 1-12 nM) to a number of primary human tumors (n = 31) such as intestinal adenocarcinoma (n = 17; 150-4000 fmol/mg protein) or breast cancer (n = 4; 250-9000 fmol/mg protein). 111In-/90Y-DOTALAN exhibited a similar high binding affinity (Kd, 1-15 nM) for the human breast cancer cell lines T47D and ZR75-1, the prostate cancer cell lines PC3 and DU145, the colonic adenocarcinoma cell line HT29, the pancreatic adenocarcinoma cell line PANC1, and the melanoma cell line 518A2. When expressed in COS7 cells, 111In DOTALAN bound with high affinity to hsst2 (Kd, 4.3 nM), hsst3 (Kd, 5.1 nM), hsst4 (Kd, 3.8 nM), and hsst5 (Kd, 10 nM) and with lower affinity to hsst1 (Kd, approximately 200 nM). The rank order of displacement of [125I]Tyr11-SST binding to hsst1 was: SST (IC50, 0.5 nM) >> DOTALAN (IC50, 154 nM) > lanreotide (LAN) approximate to Tyr3-octreotide (TOCT) approximate to DOTA-Tyr3-octreotide (DOTATOCT) approximate to DOTA-vapreotide (DOTAVAP; IC50, >1000 nM); that to hsst2 was: DOTATOCT approximate to TOCT approximate to DOTALAN approximate to SST approximately LAN approximate to DOTAVAP (IC50, 1.4 nM); that to hsst3 was: SST (IC50, 1.2 nM) > DOTALAN = LAN (IC50, 15 nM) approximate to TOCT (IC50, 20 nM) approximate to DOTAVAP (IC50, 28 nM) > DOTATOCT (IC50, 73 nM); that to hsst4 was: SST (IC50, 1.8 nM) approximate to DOTALAN (IC50, 2.5 nM) > LAN (IC50, 22 nM) >> DOTATOCT approximate to DOTAVAP approximate to TOCT (IC50, >500 nM); and that to hsst5 was: DOTALAN (IC50, 0.45 nM) > SST (IC50, 0.9 nM) > TOCT (IC50, 1.5 nM) > DOTAVAP (IC50, 5.4 nM) >> LAN (IC50, 21 nM) > DOTATOCT (IC50 260 nM). In Sprague Dawley rats (n = 10), 90Y-DOTALAN was rapidly cleared from the circulation and concentrated in hsst-positive tissues such as pancreas or pituitary. Taken together, our results indicate that 111In-/90Y-DOTALAN binds to a broad range of primary human tumors and tumor cell lines, probably via binding to hsst2-5. We conclude that this radiolabeled peptide can be used for hsst-mediated diagnosis (111In-DOTALAN) as well as systemic radiotherapy (90Y-DOTALAN) of human tumors. PMID- 10537143 TI - Prolactin stimulates leptin secretion by rat white adipose tissue. AB - Leptin, the obese (Ob) gene product, is an adipocyte-derived satiety factor that is involved in the regulation of food intake and body weight. Leptin signals nutritional status to several other physiological systems and modulates their function. As PRL is involved in energy and lipid metabolism, this study was undertaken to investigate the role of PRL on in vivo regulation of leptin serum concentration and Ob messenger RNA expression in white adipose tissue in rats. It was found that increased serum PRL levels, obtained by pituitary graft or exogenous injected ovine PRL (oPRL, 5 mg/kg), significantly stimulate serum leptin concentration. A significant increase (P < 0.01) in serum leptin concentration was present in hyperprolactinemic animals (4.7+/-0.4 microg/liter) in comparison to controls (1.2+/-0.1 microg/liter and 1.09+/-0.09 microg/liter of intact sham operated and ovariectomized rats, respectively). Similar results were obtained in oPRL-treated animals where leptin levels were 5.4+/-0.1 microg/liter vs. 1.1+/-0.1 microg/liter and 0.8+/-0.08 microg/liter of intact sham operated rats and ovariectomized, respectively (P < 0.001). This stimulatory effect of PRL on serum leptin levels was significantly reduced by food deprivation (P < 0.01) where serum leptin levels were 12.5+/-0.65 microg/liter in grafted animals vs. 3.2+/-0.36 microg/liter of grafted animals subjected to 48 h of food deprivation. Moreover, in vivo, PRL was able to induce leptin messenger RNA levels in several areas of rat white adipose tissue. The data demonstrate that PRL acts on the adipose tissue increasing leptin synthesis and secretion, suggesting a new role for this lactogenic hormone in the regulation of food intake. PMID- 10537144 TI - The in vivo effects of adrenocorticotropin and sodium restriction on the formation of the different species of steroidogenic acute regulatory protein in rat adrenal. AB - We have studied the in vivo expression of steroidogenic acute regulatory protein (StAR) in adrenals of control, ACTH-treated, and Na+-restricted rats. Indirect immunofluorescence by microscopy revealed the presence of StAR in the zonae glomerulosa (ZG) and fasciculata-reticularis (ZFR). An increased signal was observed in the ZG and zona fasciculata, 5 h after ACTH injection; a few cells of the medulla were also positive. Immunogold electron microscopy showed that StAR was mainly located over mitochondria (MT). By immunoblotting, a major 29-kDa and other minor StAR bands migrating between 30 and 39 kDa were increased 5 h after ACTH treatment but remained unchanged after 1 h. By two-dimensional-PAGE, four StAR species were revealed in homogenates of control ZG, and their intensity was increased 5 h after ACTH treatment but not after 1 h. Also, additional acidic species were seen 5 h after treatment. Other bands with basic isoelectric point were revealed between 29 and 37 kDa. Analyses on whole gland MT and supernatant (SN) revealed four bands in the control SN and five in ACTH SN; the intensity of one band was increased, and that of another one was decreased, in SN of treated rats. ACTH treatment resulted in the localization of many low-isoelectric point StARs in MT. After two-dimensional-PAGE, differences were found in the mobility of some StAR species in the ZG between controls and Na+-restricted rats. In MT, four bands were revealed in the ZG preparations of Na+-restricted and two bands in controls. Four bands were revealed in the ZG SNs of control and Na+-restricted rats; an additional band was observed only in the SN of treated animals, whereas the intensity of another band decreased. Na+ restriction did not affect StAR in the ZFR. In conclusion, StAR was present in the rat adrenal cortex ZG and ZFR and was mainly located in MT. StAR expression was inducible in the ZG and the ZF by ACTH, resulting in the formation of many StAR acidic species; interestingly, such changes were detectable 5 h, but not 1 h, after ACTH administration, suggesting that steroidogenesis stimulation by StAR might occur mainly outside MT. Although less spectacular than for ACTH, Na+ restriction also affected StAR expression in the ZG but not in the ZFR, by increasing two mitochondrial and one SN species, implying that StAR is involved in the mechanism of action of Na+ restriction in promoting aldosterone formation. These results suggest that differential processing and/or changes in phosphorylation may occur in vivo upon ACTH treatment and Na+ restriction. We hypothesize that modification of a relatively small quantity of StAR, mainly located outside MT, is necessary to increase adrenal steroidogenesis challenged either by ACTH or Na+ restriction. PMID- 10537145 TI - Protein kinase A anchoring to the myometrial plasma membrane is required for cyclic adenosine 3',5'-monophosphate regulation of phosphatidylinositide turnover. AB - The importance of the localization of protein kinase A (PKA) to the plasma membrane for cAMP-mediated inhibition of phosphatidylinositide turnover was tested in an immortalized pregnant human myometrial (PHM1-41) cell line, and the putative A kinase anchoring protein (AKAP) involved was identified. Preincubation in PHM1-41 cells with chlorophenylthio-cAMP (CPT-cAMP), forskolin, or relaxin inhibited the ability of oxytocin to stimulate phosphatidylinositide turnover. The addition of a peptide that specifically disrupts interactions of PKA RII subunits with AKAPs (S-Ht31) reversed the effects of these agents, whereas a control peptide was ineffective. The pharmacology of S-Ht31 on this particular membrane event was further characterized. A 10-min incubation with S-Ht31 at a concentration of 1 microM completely reversed the inhibitory effect of relaxin on phosphatidylinositide turnover. S-Ht31 inhibited cAMP-stimulated PKA activity in PHM1-41 cell plasma membranes and decreased the concentration of PKA. Overlay analysis detected a single AKAP of approximately 86 kDa associated with the plasma membrane of PHM1-41 cells, suggesting that the association of PKA with this AKAP is important for the cAMP inhibitory mechanism. The mol wt of this AKAP was similar to that of an AKAP associated with the plasma membrane in the human brain, AKAP79. Antibodies against AKAP79 recognized a band at 86 kDa in purified plasma membranes from the PHM1-41 cells, indicating similar determinants in these proteins. These data suggest that PKA is anchored to the myometrial plasma membrane through association with an AKAP similar to AKAP79, and that this anchoring is required for the cAMP-mediated inhibition of phosphatidylinositide turnover in PHM1-41 cells. PMID- 10537146 TI - Evidence that stimulation of two modalities of pituitary luteinizing hormone release in ovarian steroid-primed ovariectomized rats may involve neuropeptide Y Y1 and Y4 receptors. AB - A large body of evidence indicates that neuropeptide Y (NPY) is involved in stimulation of basal and cyclic release of hypothalamic LHRH and pituitary LH. To identify the NPY receptor subtypes that mediate the excitatory effects of NPY in these two modalities of LH release, we studied the effects of 1229U91, a selective Y1 receptor antagonist and Y4 receptor agonist, in two experimental paradigms that reproduce the two modalities of LH secretion in steroid-primed ovariectomized (OVX) rats. Rats were ovariectomized and implanted with a permanent cannula into the lateral cerebroventricle. In the first experiment, rats received estradiol benzoate (EB, 30 microg/rat) on day 5, followed 2 days later with progesterone (2 mg/rat) at 1000 h to induce an afternoon LH surge. 1229U91 (30 microg/3 microl) or vehicle (control) was injected intracerebroventricularly into these rats either once at 1300 h or twice (15 microg/injection) at 1100 and 1200 h. Blood samples were collected before progesterone injection at 1000 h and at hourly intervals from 1300 -1800 h via an intrajugular cannula implanted on the previous day. In control rats, serum LH levels rose significantly at 1400 h, and these high levels were maintained until 1700 h. After two injections of 1229U91, LH levels displayed a tendency to rise at 1300-1400 h, as in controls, but thereafter, decreased rapidly below the control range. In the second experiment, the acute effect of 1229U91 on LH release was evaluated in OVX rats pretreated with EB alone. Saline alone or saline containing 1, 3, 10, or 30 microg 1229U91 was injected intracerebroventricularly at 1000 h, and the effects on LH release were analyzed at 10, 20, 30, and 60 min. 1229U91 elicited a dose-dependent stimulation of LH release, with maximal response (950% of basal levels) occurring at 10 min after the 30-microg dose; elevated levels were maintained for 1 h. Because 1229U91 is a potent Y4 agonist with some affinity for Y5 receptors, these results raised the possibility that activation of Y4/Y5 receptors by 1229U91 may augment LH release. Therefore, we examined the effects of icv administration of rat pancreatic polypeptide, a Y4-selective agonist, and [D-Trp32]-NPY, a Y5 agonist on LH release in EB-primed rats. Rat pancreatic polypeptide (0.5-2 microg/rat) stimulated LH release in a dose-related manner, and peak levels (280% of basal levels) were seen at 10-20 min; the response evoked by a higher dose (10 microg) was smaller than that induced by 0.5 or 2 microg. [D-Trp32]-NPY was relatively less effective, because only the highest (10-microg) dose elicited a modest stimulation (244% of basal levels). These results demonstrate that 1229U91, a Y1 antagonist and Y4 agonist, evokes two types of responses; it suppresses the protracted ovarian steroid-induced LH surge, and acutely, it also stimulates LH. These results imply that normally two different types of NPY receptors may mediate the stimulation of LH release. Because 1229U91 is a Y1 receptor antagonist, inhibition of the steroid-induced LH surge by 1229U91 suggests that Y1 receptors may mediate the cyclic release of LH. On the other hand, acute stimulation of LH by 1229U91 implies that the Y4 agonist-like activity of 1229U91 may mediate the basal release of LH and that either NPY or a yet-to-be-identified endogenous Y4 receptor agonist may activate Y4 receptors in the hypothalamus to stimulate LH release. PMID- 10537147 TI - Insulin-like growth factor I is essential for postnatal growth in response to growth hormone. AB - Insulin-like growth factor I (IGF-I) is essential for cell growth and intrauterine development while both IGF-I and GH are required for postnatal growth. To explore the possibility of direct GH action on body growth, independent of IGF-I production, we have studied the effects of GH in an IGF-I deficient mouse line created by the Cre/loxP system. The IGF-I null mice are born with 35% growth retardation and show delayed onset of peripubertal growth, grow significantly slower, and do not attain puberty. Their adult body weight was approximately one third and body length about two thirds that of their wild-type litter mates. Injection of recombinant human GH (rhGH, 3 mg/kg, twice daily, sc) between postnatal day 14 (P14) to P56 failed to stimulate their growth as measured as both body weight and length. In contrast, wild-type mice receiving the same doses of rhGH exhibited accelerated growth starting at P21 that continued until P56, when their body weight was increased by 30% and length by 12% compared with control mice treated with diluent. Despite the lack of response in growth, IGF-I null mice have normal levels of GH receptor expression in the liver and increased liver Jun B expression and liver size in response to rhGH treatment. Our results support an essential role for IGF-I in GH-induced postnatal body growth in mice. PMID- 10537149 TI - Detection of estrogen receptor alpha and beta messenger ribonucleic acids in adult gonadotropin-releasing hormone neurons. AB - The behavior of the gonadotropin-releasing hormones (GnRH) neurons controlling fertility is dependent upon cyclic fluctuations in circulating concentrations of estrogen. However, the nature of estrogen action upon these cells has remained controversial due to their dispersed distribution within the brain, and evidence indicating that they do not express nuclear estrogen receptors (ERs) in vivo. We report here an acute brain slice preparation that enables individual living GnRH neurons to be identified within the mouse brain and show, using single cell multiplex RT-PCR, that the greater than 50% of GnRH neurons in adult and prepubertal females contain ERalpha messenger RNA. Approximately 10% of GnRH neurons contained ERbeta transcripts that were always coexistent with ERalpha. Single cell RT-PCR analysis of nonGnRH cells located in the medial preoptic area revealed a similar coexpression pattern of ERalpha and ERbeta transcripts. In contrast, single striatal cells were not found to contain ERbeta despite ERalpha being present in approximately 25% of cells. The analysis of single GnRH neurons in cycling female mice revealed that the detection of ERalpha and ERbeta transcripts was lowest on proestrus (ERalpha, 18% of all GnRH neurons; ERbeta, 0%) compared with diestrus (44% and 6%) and estrus (75% and 19%, respectively). Using a novel approach that enables single cell RT-PCR analysis of GnRH neurons, we present here evidence for the cyclic expression of ERalpha and ERbeta messenger RNAs within prepubertal and adult female GnRH neurons. PMID- 10537148 TI - Role of progesterone receptor activation in pituitary adenylate cyclase activating polypeptide gene expression in rat ovary. AB - It is well known that the pituitary gonadotropin surge induces progesterone receptor (PR) gene expression in luteinizing granulosa cells and that PR activation is critical for successful ovulation. To further understand the molecular mechanism(s) by which PR plays a role critical for granulosa cell functions, we wanted to identify progesterone-induced genes in granulosa cells. We employed a PCR-based subtraction cloning strategy to screen for genes expressed differentially in granulosa cells that were challenged with forskolin in the presence of progesterone or ZK98299. One such differentially expressed clone was identified as the pituitary adenylate cyclase activating polypeptide (PACAP). To begin to understand the relationship between PR activation and PACAP gene expression in luteinizing granulosa cells, we examined whether PR and PACAP messenger RNA (mRNA) expression is temporally correlated. In cultured granulosa cells, both human CG and forskolin induced PR and PACAP mRNA levels in a dose dependent manner, as determined by semi-quantitative RT-PCR assays. However, the peak expression for PR and PACAP mRNAs was observed at 3 h and 6 h after hormone treatment, respectively. This time difference in cAMP-responsive expression of the PR and PACAP genes is due, at least in part, to the requirement of ongoing protein synthesis for PACAP expression, as demonstrated by the inhibitory effect of cycloheximide on cAMP-induced PACAP, but not PR, mRNA levels. To determine whether PR synthesis is prerequisite for PACAP expression, we examined the effect of ZK98299, a specific PR antagonist, on cAMP-induced PACAP mRNA expression. This compound blocked cAMP-induced PACAP mRNA expression in a dose-dependent manner, indicating that PR activation is required for PACAP gene expression in granulosa cells. We then compared cellular localization and hormonal regulation of ovarian PR and PACAP gene expression in immature rats treated with gonadotropins as well as in adult rats during the preovulatory period by using in situ hybridization and semiquantitative RT-PCR assays. Results show that both PR and PACAP mRNAs are induced in granulosa cells of preovulatory follicles by human CG, but that the PR gene is expressed before the PACAP gene. Taken together, these results demonstrate that PRs mediate the LH-induced PACAP gene expression in rat granulosa cells. PMID- 10537150 TI - Changes in testosterone metabolism associated with the evolution of placental and gonadal isozymes of porcine aromatase cytochrome P450. AB - Differences in the catalytic activity of the placental and gonadal isozymes of porcine aromatase cytochrome P450 (P450arom) were examined in cell lines exhibiting stable expression of recombinant enzyme. Cell lines were selected that expressed high, but similar, immunodetectable levels of each isozyme based on Western analysis. Aromatase activity varied with growth in culture, decreasing at confluence from a peak reached between 50-80% cell density. Cells expressing the placental isozyme had 3-5 times higher catalytic activity (per mg protein) than those expressing the gonadal isozyme. The P450arom inhibitor fadrazole (1 microM) inhibited more than 97% of this activity, whereas another imidazole, etomidate (1 microM), selectively inhibited gonadal P450arom activity by 92%. Estrogen synthesis from androstenedione and testosterone was determined by RIA and confirmed by HPLC analysis, which also identified the accumulation of the 19 hydroxy and 19-oxo intermediates of the respective substrates. There was no evidence of other steroid metabolites accumulating in the media of cell lines expressing either isozyme. Tritiated water formed during aromatization of substrates 3H labeled at the C1 and C2 positions was stereo-selective for the beta orientation, but less so for testosterone than androstenedione during metabolism by the porcine placental (and human) isozyme than the gonadal isozyme. Testosterone showed a higher affinity for the porcine placental P450arom than the gonadal P450arom, but both isozymes had similar affinities for androstenedione. Testosterone was also aromatized more slowly than androstenedione by the porcine gonadal P450arom. These data suggest that catalytic differences have arisen in the substrate binding pocket during the evolution of isozymes of porcine P450arom that affect androgen metabolism, particularly the aromatization of testosterone. The physiological significance of these differences to the reproductive biology of the pig remains to be determined. PMID- 10537151 TI - Molecular cloning and expression of two type one somatostatin receptors in goldfish brain. AB - Somatostatin (SRIF or SS) exerts diverse inhibitory actions through binding to specific receptors. In this study, two SRIF receptor complementary DNAs (cDNAs) were cloned and sequenced from goldfish brain using PCR and cDNA library screening. The two cDNAs share 92% similarity in nucleotide sequence and 98% similarity in the deduced amino acid sequences and are presumably derived from duplicate genes, as goldfish are tetraploid. Two cDNAs encode two 367-amino acid goldfish type one SRIF receptors (designated as sst1A and sst1B, respectively), with seven putative transmembrane domains (TMD) and YANSCANP motif in the 7th TMD, a signature sequence for mammalian SRIF receptor (sst) family. In addition, the amino acid sequences of two receptors have 76% and 75% similarity to human or rat sst1, respectively, and 39-55% similarities to other mammalian sst subtypes (sst2-5), suggesting that the two receptors could be the goldfish homologs of mammalian sst1. The difference between goldfish and mammalian sst1 is mainly reflected by the extreme divergence in their extracellular N termini. Both SRIF 14 and [Pro2]SRIF-14, two of the native goldfish SRIF forms, significantly inhibited forskolin-stimulated cAMP release in COS-7 cells transiently expressing goldfish sSt1A or sst1B, suggesting functional coupling of the two receptors to adenylate cyclase. Northern blot and RT-PCR showed that messenger RNAs (mRNAs) for both receptors are widely distributed throughout goldfish brain, whereas only one receptor mRNA is expressed in the pituitary. RT-PCR analysis also detected sst1 receptor mRNAs in several peripheral tissues. These findings provide fundamental information for studying the mechanism of SRIF actions in vertebrates and structural analysis of mammalian sst receptors. PMID- 10537152 TI - Expression of steroidogenic genes in maternal and extraembryonic cells during early pregnancy in mice. AB - The ontogeny and functional role of steroidogenesis during early gestation in rodents is poorly understood. In previous studies, we have shown that expression of messenger RNAs (mRNAs) encoding two key enzymes indispensable for de novo synthesis of steroid hormones, i.e. cholesterol side chain cleavage cytochrome P450 (P450scc) and a newly identified isoform of murine 3beta-hydroxysteroid dehydrogenase/isomerase type VI (3betaHSD VI), is initiated upon decidualization of the uterine wall induced by implantation. In situ hybridization and immunohistochemical visualization of 3betaHSD VI mRNA and protein shows high expression of this enzyme in the antimesometrial cells of the decidua of days 6.5 7.5 post coitum (p.c.). Thereafter, expression of 3betaHSD VI in the decidual zones disappears and is replaced by a high expression of mRNA and protein in the embryonal giant trophoblast cells. At the peak of their development on day 9.5 p.c., the mouse giant trophoblast cells also express Steroidogenic Acute Regulatory (StAR) protein, which is required for steroidogenesis in the gonads and adrenal cortex. Our findings also suggest that the declining levels of P450scc, 3betaHSD VI, and StAR proteins between days 10.5-14.5 p.c. in the developing placenta is consistent with previous reports that the mouse placenta is not involved in de novo synthesis of steroids during the second half of pregnancy. Collectively, the results of the present study suggest that, during early phases of pregnancy, local progesterone synthesis in the maternal decidua and the trophoblast layers surrounding the embryonal cavity is important for successful implantation and/or maintenance of pregnancy. We propose that the local production of progesterone acts as an immunosuppressant at the fetal maternal interface preventing the rejection of the fetal allograft. PMID- 10537153 TI - Development changes in long-form leptin receptor expression and localization in rat brain. AB - The expression and localization of long-form leptin receptor (OB-Rb) were studied immunocytochemically in the brain of fetal and adult rats using a polyclonal antibody that specifically recognized OB-Rb. At 14 days of gestation, immunoreactive cells were observed in the ventricular layer, which contains premature neuronal cells. At 18 days of gestation, they were weakly stained but obvious in the paraventricular nucleus (PVN), and ependymal cells also showed immunoreactivity. At birth, the immunoreactivity of OB-Rb in the PVN seemed to be much lower than that in adult rats and remained low during the suckling period. Considering the presence of neuroendocrine and structural neuronal abnormalities in Lepob/Lepob mice with genetic leptin deficiency, these results suggest that the expression of OB-Rb in premature neuronal cells may have some function, and that the regulation of energy balance by leptin through hypothalamic regions, such as PVN, may not yet be developed in the perinatal period. PMID- 10537154 TI - Signaling between the placenta and the uterus involving the mitogen-regulated protein/proliferins. AB - The aim of this investigation was to examine signaling between the placenta and uterus during pregnancy. To do this, we determined the tissue messenger RNA and protein levels of members of a glycopeptide hormone family known to stimulate the proliferation of uterine cells and related these levels to the growth of the uterus during pregnancy in the mouse. This hormone family is known as mitogen regulated protein (MRP); alternatively proliferin (PLF). Three mrp/plf genes, plf1, mrp3 and mrp4, are expressed by the placenta with different developmental profiles. The major increase of about 4-fold in DNA content of the uterus occurs between days 9 and 14 when MRP/PLFs are present in the placenta. By contrast, the gestational changes in estradiol-17beta levels in placental and uterine tissues and in circulation do not correlate with the period of uterine growth. The previously reported mitogenic activity of the MRP/PLFs and their gestational profiles suggest that one or more of these proteins stimulates uterine proliferation during gestation. Evidence is also presented that expression of MRP3 and/or PLF1, but not MRP4, is negatively regulated by feedback from the uterus. Our results are consistent with the hypothesis that MRP/PLFs stimulate uterine proliferation in vivo and that a uterine factor shuts off PLF1 and/or MRP3 synthesis in the latter half of gestation. PMID- 10537155 TI - The luteinizing hormone-releasing hormone receptor in human prostate cancer cells: messenger ribonucleic acid expression, molecular size, and signal transduction pathway. AB - Evidence has accumulated indicating that LHRH might behave as an autocrine/paracrine growth inhibitory factor in some peripheral tumors. However, LHRH receptors in tumor cells have not been fully characterized, so far. The present experiments were performed to analyze: 1) the messenger RNA expression; 2) the molecular size; and 3) the signal transduction pathway of LHRH receptors in prostate cancer. For these studies, the human androgen-dependent LNCaP and androgen-independent DU 145 prostate cancer cell lines were used. 1) By RT-PCR, a complementary DNA product, which hybridized with a 32P-labeled oligonucleotide probe specific for the pituitary LHRH receptor complementary DNA, was found both in LNCaP and in DU 145 cells. 2) Western blot analysis, using a monoclonal antibody raised against the human pituitary LHRH receptor, revealed the presence of a protein band of approximately 64 kDa (corresponding to the molecular mass of the pituitary receptor) in both cell lines. 3) In LNCaP and DU 145 cells, pertussis toxin completely abrogated the antiproliferative action of a LHRH agonist (LHRH-A). Moreover, LHRH-A substantially antagonized the pertussis toxin catalyzed ADP-ribosylation of a Galpha(i) protein. Finally, LHRH-A significantly counteracted the forskolin-induced increase of intracellular cAMP levels in both cell lines. These data demonstrate that the LHRH receptor, which is present in prostate cancer cells, independently of whether they are androgen-dependent or not, corresponds to the pituitary receptor, in terms of messenger RNA expression and protein molecular size. However, at variance with the receptor of the gonadotrophs, prostate cancer LHRH receptor seems to be coupled to the Galpha(i) protein-cAMP signal transduction pathway, rather than to the Galpha(q/11) phospholipase C signaling system. This might be responsible for the different actions of LHRH in anterior pituitary and in prostate cancer. PMID- 10537156 TI - Effects of pulsatile infusion of the GABA(A) receptor blocker bicuculline on the onset of puberty in female rhesus monkeys. AB - In order to test the hypothesis that GABA is an inhibitory neurotransmitter restricting the release of LHRH before puberty, we examined the effects of pulsatile infusion of the GABA(A) receptor blocker, bicuculline, on the timing of puberty. Eleven female monkeys at 14-15 months of age were implanted with a stainless steel cannula into the base of the third ventricle above the median eminence. Five monkeys received bicuculline infusion every 2 h at a dose of 1 microM with a gradual increase to 100 microM in 10 microl using a portable infusion pump. The remaining 6 monkeys received similar infusions of saline. An additional 11 colony monkeys without cannula implantation were used for controls. Results indicate that bicuculline infusion advances the timing of puberty. The age of menarche (17.8+/-0.5 months) in the bicuculline infusion animals was significantly earlier than that in the saline controls (28.2+/-2.3, P < 0.001) as well as in colony controls (30.6+/-0.9, P < 0.001). The age of first ovulation (30.5+/-3.3 months) in bicuculline-treated animals was much younger (P < 0.001) than that in both controls (44.8+/-1.8 and 44.7+/-1.2, respectively). Bicuculline also accelerated the growth curve. These results suggest that the reduction of tonic GABA inhibition of LHRH neurons advances the onset of puberty. PMID- 10537157 TI - Thyroid hormone-induced cell proliferation in GC cells is mediated by changes in G1 cyclin/cyclin-dependent kinase levels and activity. AB - The thyroid hormone, 3,3', 5-triiodo-L-thyronine (T3), is essential for growth and regulation of metabolic functions. The biological activities of T3 are mediated by its interaction with the thyroid hormone nuclear receptors (TRs). The mechanism by which TRs mediate cell growth is unknown. We found that T3 stimulated cell growth in GC cells by shortening the doubling time approximately 3-fold. Flow cytometric analysis indicated that the growth stimulatory effect was mainly due to shortening of G1 phase accompanied by increases in S and G2/M phases of the cell cycle. These changes correlated with T3-induced increases in messenger RNA and protein levels of two key regulators of G1 progression, cyclins D1 and E, as well as cdk2. Furthermore, the kinase activities associated with cyclin D1 and E were activated up to 4-fold by T3, which led to increased phosphorylation of the retinoblastoma protein (Rb), the driving force in G1 to S cell cycle progression. These results show for the first time that the growth promoting effect of T3 in GC cells is mediated, at least in part, by increases in cyclin/cdk activities and the phosphorylation state of Rb. The functional link of T3 to Rb has important implications for the understanding of the biology of normal and cancer cells. PMID- 10537158 TI - Complementary deoxyribonucleic acid cloning and enzymatic characterization of a novel 17beta/3alpha-hydroxysteroid/retinoid short chain dehydrogenase/reductase. AB - 17Beta-hydroxysteroid dehydrogenases (17betaHSDs) convert androgens and estrogens between their active and inactive forms, whereas retinol dehydrogenases catalyze the conversion between retinol and retinal. Retinol dehydrogenases function in the visual cycle, in the generation of the hormone retinoic acid, and some also act on androgens. Here we report cloning and expression of a complementary DNA that encodes a new mouse liver microsomal member of the short chain dehydrogenase/reductase (SDR) superfamily and its enzymatic characterization, i.e. 17betaHSD9. Although 17betaHSD9 shares 88% amino acid identity with rat 17betaHSD6, its closest homolog, the two differ in substrate specificity. In contrast to other 17betaHSD, 17betaHSD9 has nearly equivalent activities as a 17betaHSD (with estradiol approximately = adiol) and as a 3alphaHSD (with adiol approximately = androsterone). It also recognizes retinol as substrate and represents in part the NAD+-dependent liver microsomal dehydrogenase that uses unbound retinol, but not retinol complexed with cellular retinol-binding protein. Thus, this enzyme has catalytic properties that overlap with two subgroups of SDR, 17betaHSD and retinol dehydrogenases. Inactivation of estrogen and a variety of androgens seems to be its most probable function. Because of its apparent inability to access retinol bound with cellular retinol-binding protein, a function in the pathway of retinoic acid biosynthesis seems less obvious. These data provide additional insight into the enzymology of estrogen, androgen, and retinoid metabolism and illustrate how closely related members of the SDR superfamily can have strikingly different substrate specificities. PMID- 10537159 TI - Insulin inhibits angiotensinogen gene expression via the mitogen-activated protein kinase pathway in rat kidney proximal tubular cells. AB - The present study aimed to investigate the molecular mechanism(s) of insulin action on angiotensinogen (ANG) secretion and gene expression in kidney proximal tubular cells exposed to high levels of glucose. Immortalized rat proximal tubular cells (IRPTC) were cultured in monolayer. The levels of rat ANG and ANG messenger RNA in the IRPTC were quantified by a specific RIA for rat ANG (RIA rANG) and by an RT-PCR assay. Insulin inhibited the stimulatory effect of a high level of glucose (25 mM) and phorbol 12-myristate 13-acetate, an activator of protein kinase C) on the secretion of ANG and the expression of the ANG messenger RNA in IRPTC. This inhibitory action of insulin on the ANG secretion and gene expression was blocked by PD98059 (an inhibitor of mitogen-activated protein kinase kinase) but not by Wortmannin (an inhibitor of phosphatidylinositol-3 kinase). PD98059 was effective in inhibiting the phosphorylation of MEK 1/2 and p44/42 MAP kinase in IRPTC stimulated by insulin. These studies demonstrate that insulin prevents the stimulatory effect of high levels of glucose on the expression of the renal ANG gene in IRPTC, at least in part, via the MAPK kinase signal transduction pathway, subsequently inhibiting the activation of the local renal renin-angiotensin system. PMID- 10537160 TI - The magnitude of decrease in hepatic very low density lipoprotein apolipoprotein B secretion is determined by the extent of 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibition in miniature pigs. AB - It has been postulated that the rate of hepatic very low density lipoprotein (VLDL) apolipoprotein (apo) B secretion is dependent upon the activity of 3 hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase. To test this hypothesis in vivo, apoB kinetic studies were carried out in miniature pigs before and after 21 days treatment with high-dose (10 mg/kg/day), atorvastatin (A) or simvastatin (S) (n = 5). Pigs were fed a diet containing fat (34% of calories) and cholesterol (400 mg/day; 0.1%). Statin treatment decreased plasma total cholesterol [31 (A) vs. 20% (S)] and low density lipoprotein (LDL) cholesterol concentrations [42 (A) vs. 24% (S)]. Significant reductions in plasma total triglyceride (46%) and VLDL triglyceride (50%) concentrations were only observed with (A). Autologous [131I]VLDL, [125I]LDL, and [3H]leucine were injected simultaneously, and apoB kinetic parameters were determined by triple-isotope multicompartmental analysis using SAAM II. Statin treatment decreased the VLDL apoB pool size [49 (A) vs. 24% (S)] and the hepatic VLDL apoB secretion rate [50 (A) vs. 33% (S)], with no change in the fractional catabolic rate (FCR). LDL apoB pool size decreased [39 (A) vs. 26% (S)], due to reductions in both the total LDL apoB production rate [30 (A) vs. 21% (S)] and LDL direct synthesis [32 (A) vs. 23% (S)]. A significant increase in the LDL apoB FCR (15%) was only seen with (A). Neither plasma VLDL nor LDL lipoprotein compositions were significantly altered. Hepatic HMG-CoA reductase was inhibited to a greater extent with (A), when compared with (S), as evidenced by 1) a greater induction in hepatic mRNA abundances for HMG-CoA reductase (105%) and the LDL receptor (40%) (both P < 0.05); and 2) a greater decrease in hepatic free (9%) and esterified cholesterol (25%) (both P < 0.05). We conclude that both (A) and (S) decrease hepatic VLDL apoB secretion, in vivo, but that the magnitude is determined by the extent of HMG-CoA reductase inhibition. PMID- 10537161 TI - Expression of activin A and follistatin core proteins by human prostate tumor cell lines. AB - Activin and follistatin (FS) messenger RNA and protein are expressed and localized to human prostate tissue from men with high grade cancer and to human prostate tumor cell lines LNCaP, DU145, and PC3. Although activin A induces apoptosis and inhibits cell proliferation in LNCaP cells, PC3 cells are insensitive to the effect of exogenous addition of activin A. The results of this study show that activin A and FS are produced and can be measured by specific enzyme-linked immunosorbent assays in PC3 cells and media but are not detectable in LNCaP cells. Over 10 days in culture, the production of activin A by PC3 cells declines and is inversely correlated (r = -0.779) to FS288 production, which steadily increases and is significantly elevated compared with Day 1 of culture. The presence of FS288 and FS315 proteins was confirmed by immunocytochemistry and showed that only PC3 cells produced the FS288 isoform. Western blotting of PC3 cell media confirmed the presence of the FS288 isoform. Blockade of FS288 activity with a neutralizing antibody rendered PC3 cells responsive to activin A, as measured by inhibition of proliferation. Collectively, these results suggest that PC3 tumor cells are insensitive to activin A because they produce measurable amounts of activin ligand and FS288 protein, which is capable of blocking the autocrine response of these cells to activin A. PMID- 10537163 TI - Age- and sex-specific promoter function of a 2-kilobase 5'-flanking sequence of the murine luteinizing hormone receptor gene in transgenic mice. AB - A transgenic (TG) mouse model carrying a 2-kb murine LH receptor (LHR) promoter/beta-galactosidase (beta-GAL) fusion gene was created to study the regulatory function of the 5'-flanking region of the murine LHR gene. Of the five TG mouse lines produced, three displayed high beta-GAL expression in the testis, but none showed any expression in the ovary. In addition, all mouse lines of both sexes expressed beta-GAL consistently in the brain, most prominently in hippocampus, hypothalamus, midbrain, and cortex. Weak staining was found in a few pituitary samples. All other tissues examined were negative for transgene expression. In support of sex-specific gonadal expression of the transgene, transient transfection of the LHR/beta-GAL gene construct into immortalized mouse granulosa (KK-1) and Leydig (mLTC-1) tumor cells revealed a more than 5-fold higher expression level in the Leydig cells. Histological examination of the TG testes demonstrated strong beta-GAL expression in Leydig cells, but, unexpectedly, also in elongating spermatids of adult age and in some spermatogonia of the neonatal testis. The functional significance of the latter findings remains open. The transgene was only expressed in adult Leydig cells; no expression was found in the fetal population of these cells. Hence, these findings indicate that the immediate 2-kb fragment of the murine LHR 5'-flanking sequence is transcriptionally active only in adult Leydig cells and certain brain areas, but other promoter sequences are apparently needed for ovarian and fetal testicular expression of the LHR gene. PMID- 10537162 TI - Differential uterine expression of estrogen and progesterone receptors correlates with uterine preparation for implantation and decidualization in the mouse. AB - The present investigation examined the spatiotemporal expression of estrogen receptors (ER-alpha and ER-beta) and progesterone receptor (PR) in the periimplantation mouse uterus (days 1-8). ER-alpha messenger RNA (mRNA) was detected at much higher levels in the periimplantation uterus compared with that of ER-beta mRNA, the levels of which were very low in all uterine cells during this period. Results of in situ hybridization demonstrated expression of ER-alpha mRNA primarily in the luminal and glandular epithelia on days 1 and 2 of pregnancy. On days 3 and 4, the accumulation was localized primarily in stromal cells in addition to its presence in the epithelium. Following implantation on day 5, the accumulation of this mRNA was more condensed in the luminal and glandular epithelia, but declined in the subluminal epithelial stroma at the sites of implanting embryos. On days 6-8, the accumulation of ER-alpha mRNA was primarily localized in the secondary decidual zone (SDZ) with more intense localization in the subepithelial cells at the mesometrial pole. In contrast, signals were very low to undetectable in the primary decidual zone (PDZ), and no signals were detected in implanting embryos. The undifferentiated stroma underneath the myometrium also showed positive signals. The immunolocalization of ER-alpha protein correlated with the mRNA localization. Western blot analysis showed down-regulation of ER-alpha in day 8 decidual cell extracts consistent with the down-regulation of ER-alpha mRNA in decidual cells immediately surrounding the embryo on this day. The expression pattern of PR was also dynamic in the periimplantation uterus. On day 1, the accumulation of PR mRNA was very low to undetectable, whereas only a modest level of accumulation in the epithelium was noted on day 2. On days 3 and 4, the accumulation of this mRNA was detected in both the epithelium and stroma. In contrast, the expression was restricted only to the stroma with increased signals at the sites of implantation on day 5. On days 6-8, PR mRNA accumulation increased dramatically throughout the deciduum. The localization of immunoreactive PR correlated with the mRNA distribution in the periimplantation uterus. Taken together, the results demonstrate that the expression of ER-alpha, ER-beta, and PR is differentially regulated in the periimplantation mouse uterus. This compartmentalized expression of ER and PR provides information regarding the sites of coordinated effects ofestrogen and progesterone in the preparation of the uterus for implantation and decidualization during early pregnancy. PMID- 10537164 TI - Distinct expression of gelatinase A [matrix metalloproteinase (MMP)-2], collagenase-3 (MMP-13), membrane type MMP 1 (MMP-14), and tissue inhibitor of MMPs type 1 mediated by physiological signals during formation and regression of the rat corpus luteum. AB - The corpus luteum (CL) is a transient endocrine organ that secretes progesterone to support pregnancy. The CL is formed from an ovulated follicle in a process that involves extensive angiogenesis and tissue remodeling. If fertilization does not occur or implantation is unsuccessful, the CL will undergo regression, which involves extensive tissue degradation. Extracellular proteases, such as serine proteases and matrix metalloproteinases (MMPs), are thought to play important roles in both the formation and regression of the CL. In this study, we have examined the physiological regulation pattern and cellular distribution of messenger RNAs coding for gelatinase A (MMP-2), collagenase-3 (MMP-13), membrane type MMP 1 (MT1-MMP, MMP-14), and the major MMP inhibitor, tissue inhibitor of MMPs type 1 (TIMP-1) in the CL of adult pseudopregnant (psp) rat. Northern blot and in situ hybridization analyses revealed that gelatinase A messenger RNA was mainly expressed during luteal development, indicating that gelatinase A may be associated with the neovascularization and tissue remodeling that takes place during CL formation. Collagenase-3 had a separate expression pattern and was only expressed in the regressing CL, suggesting that this MMP may be related with luteal regression. MT1-MMP that in vitro can activate progelatinase A and procollagenase-3 was constitutively expressed during the formation, function, and regression of the CL and may therefore be involved in the activation of these MMPs. TIMP-1 was induced during both the formation and regression of the CL, suggesting that this inhibitor modulates MMP activity during these processes. To test whether the induction of collagenase-3 and TIMP-1 is coupled with luteal regression, we prolonged the luteal phase by performing hysterectomies, and induced premature luteal regression by treating the pseudopregnant rats with a PGF2alpha analog, cloprostenol. In both treatments, collagenase-3 and TIMP-1 were induced only after the serum level of progesterone had decreased, suggesting that collagenase-3 and TIMP-1 are induced by physiological signals, which initiate functional luteolysis to play a role in tissue degradation during structural luteolysis. In conclusion, our data suggest that gelatinase A, collagenase-3, and MT1-MMP may have separate functions during the CL life span: gelatinase A mainly takes part in CL formation, whereas collagenase-3 mainly takes part in luteal regression; MT1-MMP is constitutively expressed during the CL life span and may therefore serve as an in vivo activator of both gelatinase A and collagenase-3. TIMP-1 is up-regulated both during the formation and regression of the CL and may therefore regulate MMP activity during both processes. PMID- 10537165 TI - Estrogen prevents glucocorticoid-induced apoptosis in osteoblasts in vivo and in vitro. AB - The ability of estrogen to prevent glucocorticoid-induced apoptosis in osteoblasts was studied both in vitro and in vivo. Glucocorticoid treatment for 72 h produced a dose-dependent increase in the number of apoptotic cells, determined by acridine orange/ethidium bromide staining, with a maximal response of 31+/-2% and 26+/-3% with 100 nM corticosterone in primary rat and mouse osteoblasts, respectively. Simultaneous administration of varying concentrations of 17beta-estradiol and 100 nM corticosterone decreased apoptotic osteoblasts in a dose-dependent manner, with a maximal decrease of 70% with 0.01 nM 17beta estradiol. Terminal deoxynucleotidyltransferase-mediated deoxy-UTP-biotin nick end labeling also demonstrated glucocorticoid-induced DNA fragmentation that was inhibited by estrogen. Estrogen was shown to inhibit apoptosis induced by lipopolysaccharide treatment. As early as 6 h, Western blots demonstrated a dose dependent decrease in the Bcl-2/Bax ratio, which reached a minimum of 0.18 in osteoblasts treated with 1000 nM corticosterone for 72 h. This reduction in Bcl 2/Bax was abolished by treating osteoblasts simultaneously with 17beta-estradiol, but not with 17alpha-estradiol. In 7-day-old mice, administration of varying concentrations of dexamethasone for 72 h resulted in a dose-dependent increase in the number of apoptotic osteoblasts as demonstrated by in situ terminal deoxynucleotidyltransferase-mediated deoxy-UTP-biotin nick end labeling staining of calvaria. A maximum of 22+/-1% apoptotic osteoblasts on the bone surface was found with 1 mg/kg BW dexamethasone compared with 2+/-1% in vehicle-treated mice. Injection of varying concentrations of 17beta-estradiol (0.5-5 mg/kg BW), but not 17alpha-estradiol, with 1 mg/kg dexamethasone produced a dose-dependent decrease in the number of apoptotic osteoblasts to 5+/-1% with 5 mg/kg 17beta-estradiol. Thus, glucocorticoid-induced apoptosis of osteoblasts may be prevented at least in part by 17beta-estradiol. PMID- 10537166 TI - Pituitary lactotroph adenomas develop after prolonged lactotroph hyperplasia in dopamine D2 receptor-deficient mice. AB - Tuberoinfundibular dopamine tonically inhibits PRL expression and secretion from the pituitary gland by the activation of dopamine D2 receptors (D2R) localized on lactotrophs. Mutant female mice that lack D2Rs have persistent hyperprolactinemia but also develop extensive hyperplasia of pituitary lactotrophs and peliosis of the adenohypophysis at 9 to 12 months of age, while age-matched male D2R deficient mice have no morphologic adenohypophysial lesion. We now report that both female and male D2R-deficient mice 17 to 20 months of age develop pituitary lactotroph adenomas. Of 12 aged female mice examined, all developed monohormonal PRL-immunoreactive neoplasms that had a characteristic juxtanuclear Golgi pattern of PRL staining and loss of the reticulin fiber network. Several of these adenomas were 50-fold larger than normal glands with marked suprasellar extension and invasion of brain but no gross evidence of distant metastases. They also had striking peliosis that was more marked than the lesion seen in the hyperplastic pituitaries of the younger females. These findings demonstrate that a chronic loss of neurohormonal dopamine inhibition promotes the hyperplasia-neoplasia sequence in adenohypophysial lactotrophs. Our results are analogous to previous data indicating that protracted stimulation of adenohypophysial cells by hormones or growth factors results in proliferation with initial hyperplasia followed by the development of neoplasia. Six aged male D2R-deficient mice had slightly enlarged anterior pituitaries similar in size to normal female glands. However, each case exhibited multifocal, microscopic lactotroph adenomas with strong nuclear immunoreactivity for estrogen receptors and Pit-1 transcription factor. The unexpected development of adenomas in males without preexisting or concomitant hyperplasia suggests that prolonged loss of dopamine inhibition may also cause neoplasia by distinct cellular mechanisms in male and female animals. PMID- 10537167 TI - Glucagon-like peptide-1-(7-36)amide is transformed to glucagon-like peptide-1-(9 36)amide by dipeptidyl peptidase IV in the capillaries supplying the L cells of the porcine intestine. AB - The insulinotropic hormone glucagon-like peptide-1 (GLP-1) is stored in the intestinal L cell in an active form, GLP-1-(7-36)amide, but more than half of the endogenous peptide circulates in an inactive, N-terminally truncated form, GLP-1 (9-36)amide. This study examined the GLP-1 newly secreted from the porcine ileum, in vitro (isolated perfused preparation) and in vivo (anesthetized pig), to determine where this conversion occurs. Although the GLP-1 extractable from the porcine ileum is predominantly the intact peptide (94.6+/-1.7%), a large proportion of the GLP-1 that is secreted has already been degraded to the truncated form both in vitro (53.8+/-0.9% intact) and in vivo (32.9+/-10.8% intact). In the presence of a specific dipeptidyl peptidase IV (DPP IV) inhibitor (valine-pyrrolidide), the proportion of intact GLP-1 released from the perfused ileum was increased under both basal (99% intact; P < 0.05) and stimulated (86 101% intact; P < 0.05) conditions. Immunohistochemical and histochemical studies revealed specific DPP IV staining in the brush border epithelium as well as in the capillary endothelium. Double staining showed juxtapositioning of DPP IV positive capillaries and GLP-1-containing L cells. From these results, we suggest that GLP-1 is degraded as it enters the DPP IV containing blood vessels draining the intestinal mucosa. PMID- 10537168 TI - Permanent effects of neonatal estrogen exposure in rats on reproductive hormone levels, Sertoli cell number, and the efficiency of spermatogenesis in adulthood. AB - This study aimed to identify the mechanism(s) for impairment of spermatogenesis in adulthood in rats treated neonatally with estrogens. Rats were treated (days 2 12) with 10, 1, or 0.1 microg diethylstilbestrol (DES), 10 microg ethinyl estradiol (EE), 10 mg/kg of a GnRH antagonist (GnRHa), or vehicle and killed in adulthood. DES/EE caused dose-dependent reductions in testis weight, total germ cell volume per testis, and Sertoli cell volume per testis. Sertoli cell number at 18 days of age in DES-treated rats was reduced dose dependently. GnRHa treatment caused changes in these parameters similar to those in rats treated with 10 microg DES. Plasma FSH levels were elevated (P < 0.001) to similar levels in all treatment groups regardless of differences in Sertoli cell number and levels of inhibin B; the latter reflected Sertoli cell number, but levels were disproportionately reduced in animals treated with high doses of DES/EE. Neonatal estrogen treatment, but not GnRHa, caused dose-dependent reductions (40-80%) in plasma testosterone levels in adulthood, but did not alter LH levels. Preliminary evidence suggests that the decrease in testosterone levels in estrogen-treated rats is not due to reduced Leydig cell volume per testis. GnRHa-treated rats exhibited a significant increase in germ cell volume per Sertoli cell and a reduction in germ cell apoptosis, probably because of the raised FSH levels. Despite similar raised FSH levels, rats treated with DES (10 or 1 microg) or EE (10 microg) had reduced germ cell volume/Sertoli cell and increased germ cell apoptosis, especially when compared with GnRHa-treated animals. The latter changes were associated with an increase in lumen size per testis, indicative of impaired fluid resorption from the efferent ducts, resulting in fluid accumulation in the testis. Rats treated neonatally with 0.1 microg DES showed reduced germ cell apoptosis comparable to that in GnRHa-treated animals. The changes in apoptotic rate among treatment groups occurred across all stages of the spermatogenic cycle. It is concluded that 1) neonatal estrogen treatment results in dose-dependent alterations in Sertoli cell numbers, germ cell volume, efficiency of spermatogenesis, and germ cell apoptosis in adulthood; 2) the relatively poor spermatogenesis in estrogen-treated animals is most likely due to altered testis fluid dynamics and/or altered Sertoli cell function; 3) as indicated by FSH (LH) and testosterone levels, the hypothalamic-pituitary axis and Leydig cells are probably more sensitive than the Sertoli cells to reprogramming by estrogens neonatally; and 4) elevated FSH levels in adulthood may improve the efficiency of spermatogenesis. PMID- 10537169 TI - Developmental expression of the homeodomain protein IDX-1 in mice transgenic for an IDX-1 promoter/lacZ transcriptional reporter. AB - Expression of the homeodomain transcription factor IDX-1 (also known as IPF-1, STF-1, and PDX-1) is required for pancreas development, because disruption of the gene in mice and humans results in pancreatic agenesis. During embryonic development the idx-1 gene is first expressed in a localized region of foregut endoderm from which the duodenum and pancreas later develop. To more fully understand the role of IDX-1 in pancreas development, transgenic mice expressing the Escherichia coli lacZ gene under control of the 5'-proximal 4.6 kb of the idx 1 promoter were created as a reporter for the developmental expression of IDX-1. Here we show that the determinants for the developmental and tissue-specific expression of the endogenous idx-1 gene are faithfully reproduced by the 4.6-kb region of the idx-1 promoter. Expression of lacZ is detected in the development of the exocrine and endocrine pancreas in pancreatic ducts, common bile and cystic ducts, pyloric glands of the distal stomach, Brunner's glands, the intestinal epithelium of the duodenum, and the spleen. The observed spatial and temporal pattern of lacZ expression directed by the IDX-1 promoter further supports an important role of IDX-1 in specifying the development of several endodermal structures within the midsegment of the body. An unexpected finding is that IDX-1 promoter-driven (transcriptional) lacZ activity does not always coincide with the localization of IDX-1 messenger RNA by in situ hybridization and IDX-1 protein by immunocytochemistry in adult rat duodenum, suggesting the existence of regulation of IDX-1 expression at the posttranscriptional level of expression of the idx-1 gene. PMID- 10537170 TI - Morphological evidence for direct interaction between arcuate nucleus neuropeptide Y (NPY) neurons and gonadotropin-releasing hormone neurons and the possible involvement of NPY Y1 receptors. AB - Neuropeptide Y (NPY) neurons in the arcuate nucleus of the hypothalamus (ARH) have been shown to play an important role in modulating LH secretion. One mechanism by which the ARH NPY system may regulate LH secretion is by modulating GnRH neuronal function. Thus, the present study examined whether the ARH NPY system provided direct input to GnRH cell bodies in the preoptic area (POA), as well as to their nerve terminals in the median eminence (ME). The possible involvement of the NPY Y1 receptor subtype in mediating the effects of NPY was also investigated. Lactating rats were used in these studies because they have increased hypothalamic NPY content, especially in the ARH/ME areas, making it easier to detect NPY fibers and terminals. The anterograde tracer, Phaseolus vulgaris leucoagglutinin (PHA-L), was iontophoresed into the ARH of lactating rats; and triple-label immunofluorescence was performed, with the aid of confocal microscopy, to visualize NPY, PHA-L, and GnRH. GnRH cell bodies were found scattered throughout the organum vasculosum laminae terminalis (OVLT)/POA region, and NPY/ PHA-L double-labeled fibers were found in very close proximity to numerous GnRH perikarya. In the ME, double-labeled NPY/PHA-L fibers were found in the inner and external zones, and they were found in close proximity to GnRH neuronal fibers. Using a NPY Y1 specific antibody, double-label immunofluorescence was performed to examine whether the Y1 receptor subtype was expressed in GnRH neurons. No convincing Y1-positive staining was found in GnRH cell bodies in the OVLT/POA region. However, abundant Y1-positive fiber and cell staining were observed throughout the region, and Y1-positive fibers were found in close apposition to GnRH cell bodies. In contrast, numerous GnRH nerve fibers and terminals in both the OVLT and ME were colocalized with Y1-positive staining. The results of this study suggest that ARH NPY neurons come in close contact with GnRH neurons and may provide direct input to both GnRH cell bodies in the POA region and to their nerve terminals in the ME. The Y1 receptor subtype may be directly involved in NPY modulation of GnRH secretion from its nerve terminals. PMID- 10537171 TI - Single cell reverse transcription-polymerase chain reaction analysis of rat supraoptic magnocellular neurons: neuropeptide phenotypes and high voltage-gated calcium channel subtypes. AB - Magnocellular neurosecretory cells (MNCs) in the hypothalamo-neurohypophysial system that express and secrete the nonapeptides oxytocin (OT) and vasopressin (VP) were evaluated for the expression of multiple genes in single magnocellular neurons from the rat supraoptic nucleus using a single cell RT-PCR protocol. We found that all cells representing the two major phenotypes, the OT and VP MNCs, express a small, but significant, amount of the other nonapeptide's messenger RNA (mRNA). In situ hybridization histochemical analyses confirmed this observation. A third phenotype, containing equivalent amounts of OT and VP mRNA, was detected in about 19% of the MNCs from lactating female supraoptic nuclei. Analyses of these phenotypes for other coexisting peptide mRNAs (e.g. CRH, cholecystokinin, galanin, dynorphin, and the calcium-binding protein, calbindin) generally confirmed expectations from the literature, but revealed cell to cell variation in their coexpression. Our results also show that the high voltage-activated calcium channel subunit genes, alpha1A-D, alpha2, and beta1-4 are expressed in virtually all MNCs. However, the alpha1E subunit gene is not expressed at detectable levels in these cells. The expression of all of the beta-subunit genes in each MNC may account for the variations in physiological and pharmacological properties of the high voltage-activated channels found in these neurons. (Endocrinology 140: 5391-5401, 1999) PMID- 10537172 TI - Underlying disease stress augments plasma and tissue adrenomedullin (AM) responses to endotoxin: colocalized increases in AM and inducible nitric oxide synthase within pancreatic islets. AB - Rapid onset metabolic impairments accompany the initiation of the acute phase response to many disease stresses, whereas more chronic metabolic perturbations may prolong the recovery period. In the present experiment the application of a mild endotoxin challenge [lipopolysaccharide (LPS)] alone or additive to a chronic subclinical parasitic infection (Sarcocystis cruzi; LPS + PI) in calves was used as a model to investigate and define a dynamic axis coordinated between adrenomedullin (AM) and nitric oxide in response to immune challenge. Plasma AM and NO2/NO3 concentration responses after LPS (0.45 microg/kg, iv) were rapid in onset and of higher magnitude and longer duration in PI + LPS calves than in those challenged with LPS alone. The post-LPS increase in plasma insulin was significantly greater in PI + LPS than in LPS; following refeeding of calves, insulin secretion was most blunted in PI + LPS calves, consistent with the inhibitory effects of NO and AM on insulin secretion. A more chronic response to the immune challenge (organ specific) was in evidence in tissues harvested 24 h after LPS challenge. Where lung and liver showed no immunostaining for inducible nitric oxide (iNOS), iNOS immunostaining was present in the pancreas, localized to islets only. The percentages of iNOS-immunopositive cells in islets were 1.7%, 21%, 6.7%, and 24% for control (C; saline infused), PI, LPS, and PI + LPS calves, respectively. AM immunostaining was not evident in the liver and was present, but not differentially affected by treatment, in airway epithelium in the lung. The number of islet cells with positive immunostaining for AM was increased in LPS, PI, and PI + LPS calves. The percentages ofAM-immunopositive cells in islets were 8%, 27%, 20%, and 33% for C, PI, LPS, and PI + LPS, respectively. Immunostaining for AM and iNOS was colocalized with cells positive for pancreatic polypeptide. By triple label confocal fluorescence immunocytochemistry, colocalization of intense AM and iNOS immunostaining was confirmed in peripheral islet cells. A weaker, more diffuse iNOS signal was also apparent in insulin-containing cells in PI + LPS. We conclude that chronic low level infection potentiates the severity of metabolic perturbations that arise with additive sudden onset immune challenge, as can occur with bacterial toxins. These metabolic disturbances are reflected in and possibly mediated by early onset increases in plasma tumor necrosis factor-alpha, insulin, and AM and up-regulated iNOS activity. These acute complications rapidly progress into a more chronic state characterized by diminished insulin response to feeding stimulus and colocalized increases in pancreatic islet AM and iNOS. The pancreatic responses in AM and iNOS may play a major role in mediating prolonged disturbances in nutrient use by tissues through their influences on temporal patterns of pancreatic hormone secretion during chronic illness. PMID- 10537173 TI - Prolactin is a survival factor for androgen-deprived rat dorsal and lateral prostate epithelium in organ culture. AB - PRL is one of several polypeptide factors that regulate growth and differentiation of prostate epithelium besides steroid hormones. This hormone may also participate in the development of pathologic changes of the prostate, as evidenced by marked prostate hyperplasia in hyperprolactinemic mice. We have previously demonstrated expression of PRL receptors and androgen-dependent local production of PRL in rat and human prostate epithelium, suggesting the existence of an autocrine loop. We now show that PRL acts as a survival factor for epithelial cells of rat dorsal and lateral prostate but not ventral prostate, using long-term organ cultures as an in vitro model. Culture of prostate explants in androgen-free medium was associated with a transient surge of apoptosis during the first 2-4 days of culture in rat ventral, dorsal, and lateral prostate tissues, as quantified by either nuclear morphology or in situ DNA fragmentation analysis. PRL significantly inhibited apoptosis in androgen-deprived dorsal and lateral prostate cultures, by 40-60%, as determined by the two methods. The present study has established conditions and methodology for analysis of apoptosis in organ cultures of rat prostate and suggests a physiological role for PRL as a survival factor for prostate epithelium. PMID- 10537174 TI - Follicular thyroglobulin suppresses iodide uptake by suppressing expression of the sodium/iodide symporter gene. AB - A major function of the thyrocyte is to take up and concentrate iodide. This is needed for thyroid hormone synthesis and is accomplished by the sodium iodide symporter (NIS), whose expression and activity are up-regulated by TSH. Recently, we reported that follicular thyroglobulin (TG) is a potent suppressor ofthyroid specific gene expression and can overcome TSH-increased gene expression. We suggested this might be a negative feedback, autoregulatory mechanism that counterbalanced TSH stimulation of follicular function. In this report, we support this hypothesis by coordinately evaluating TG regulation of NIS gene expression and iodide transport. We show that physiological concentrations of TG similarly and significantly suppress TSH-increased NIS promoter activity, NIS protein, and NIS-dependent iodide uptake as well as RNA levels. We show, in vivo, that TG accumulation at the apical membrane of a thyrocyte facing the follicular lumen is associated with decreased uptake ofradioiodide. It is likely, therefore, that TG suppresses NIS-dependent iodide uptake and NIS gene expression in vivo, as is the case in vitro. RNA levels of NIS and vascular endothelial growth factor/vascular permeability factor, which has been reported to be TSH regulated and possibly associated with TSH-increased iodide uptake, are coordinately decreased by follicular TG as a function of concentration and time. Also, removal of follicular TG from the medium, but not TSH, coordinately returns NIS and vascular endothelial growth factor/vascular permeability factor RNA levels to their TSH-stimulated state. TG accumulated in the follicular lumen appears, therefore, to be a negative feedback regulator of critical TSH-increased follicular functions, iodide uptake, and vascular permeability. PMID- 10537175 TI - Characterization of FAP-1 expression and function in thyroid follicular cells. AB - Human thyrocytes are resistant to Fas-mediated programmed cell death (PCD). It has been reported that a labile protein inhibitor is involved in the protection of thyrocytes from PCD, and its action can be reversed by incubation of thyrocytes with cycloheximide (CHX) during treatment with agonist anti-Fas Ab. Fas-associated phosphatase-1 (FAP-1) is a protein that has been shown to interact with the negative regulatory domain of Fas and block Fas-mediated apoptosis in FAP-1 transfected Jurkat cells. We investigated the possibility that FAP-1 might be involved in protection against Fas-mediated PCD in human thyrocytes. FAP-1 mRNA was detected in primary thyrocytes using a ribonuclease protection assay. The presence of FAP-1 protein was confirmed by immunohistochemical staining and flow cytometry using a polyclonal anti-FAP-1 Ab. FAP-1 protein also disappeared from thyroid cells in response to CHX. To determine whether FAP-1 is a functional inhibitor of PCD in thyrocytes, we incubated thyrocytes with synthetic SLV (Ac SLV) tripeptide to compete with Fas for interaction with FAP-1. Thyrocytes treated with Ac-SLV tripeptide showed significantly increased cell death as compared to cells treated with control tripeptide. In addition, in the presence of a suboptimal concentration of CHX, the Ac-SLV tripeptide yielded a strong, synergistic increase in Fas-mediated PCD as compared to thyrocytes treated with control tripeptide. These results implicate FAP-1 as a regulator of Fas-induced PCD in thyrocytes. PMID- 10537176 TI - Estrogen regulates angiotensin AT1 receptor expression via cytosolic proteins that bind to the 5' leader sequence of the receptor mRNA. AB - Two of the most highly recognized factors implicated in the pathogenesis of hypertension, atherosclerosis, congestive heart failure and associated cardiovascular disease are the renin angiotensin system (RAS) and estrogen. A major effect of estrogen results from its influence on the RAS. Beta-estradiol (E2) replacement in ovariectomized (OVX) rats significantly decreased type 1 angiotensin (AT1) receptor expression in the pituitary and adrenal, whereas it significantly increased receptor expression in the uterus when compared to OVX controls. Additional evidence demonstrated an important influence of estrogen on a recently discovered post-transcriptional mechanism for regulating expression of the AT1 receptor. This mechanism consists of cytosolic RNA binding proteins (BPs) that recognize the 5' leader sequence (5'LS) of the receptor mRNA. The activities of these 5'LS BPs were modulated by estrogen in an inverse manner to AT1 receptor regulation. Moreover, in vitro translation assays in wheat germ lysates suggested that the 5'LS BPs inhibited AT1 receptor translation. Our data therefore indicate that hormonal regulation of AT1 receptors involves modulation of 5'LS BPs by estrogen. These findings may in part account for the observed protective effects of estrogen on cardiovascular disease. PMID- 10537178 TI - Type 3 iodothyronine deiodinase is selectively expressed in areas related to sexual differentiation in the newborn rat brain. AB - Thyroid hormone (T4 and T3) concentrations in target tissues are greatly influenced by the activity of iodothyronine deiodinases. Type 1 and 2 deiodinases generate T3 from T4, while deiodinase type 3 (D3) transforms T4 and T3 to inactive metabolites. Coordination of the expression and activity of these enzymes is postulated to play an important role in physiology and development, making it possible that individual cells and tissues regulate the concentrations of the active hormone according to specific needs. We have analyzed the expression of D3 in the neonatal rat brain by in situ hybridization using a specific 35S-labelled riboprobe. At postnatal day 0 D3 transcripts were unexpectedly found to be selectively expressed in areas involved in sexual differentiation of the brain such as the bed nucleus of the stria terminalis and preoptic nuclei. Expression in these areas was transient and was no longer observed at postnatal day 10. These observations suggest that D3 expression is linked to the early mechanism determining sexual function and behavior. PMID- 10537177 TI - Expression of adrenomedullin in feto-placental circulation of human normotensive pregnant women and pregnancy-induced hypertensive women. AB - The adrenomedullin (AM) peptide and the expression of AM messenger RNA (mRNA) from feto-maternal tissues of 22 normotensive pregnant women and from 7 women with pregnancy-induced hypertension (PIH) during third-trimester were examined to clarify the pathophysiological features of PIH. Samples of the placenta, uterine muscle, umbilical artery, and fetal membranes were obtained from each patients under informed consent. The AM peptide was measured by a radioimmunoassay and the AM mRNA level was analyzed by Northern hybridization. The total immunoreactive AM (fmol/mg wet tissue) was significantly increased in the fetal membranes (1.95+/ 0.20 vs. 3.03+/-0.44) and the umbilical artery (0.11+/-0.01 vs. 0.15+/-0.02) of the patients with PIH. On the other hand, the AM mRNA level was higher in the umbilical artery, and lower in the fetal membranes in the patients with PIH. The present results thus suggest that the changes in the expression of AM in fetal membrane and umbilical artery in PIH may play an important role in the fetal and maternal circulation. PMID- 10537179 TI - Prolactin as a chemoattractant for human breast carcinoma. AB - Prolactin (PRL) is recognized as a growth and differentiating hormone in the human breast. These effects are mediated by the PRL receptor (PRLr); when stimulated the PRL-PRLr complex activates several signaling cascades, including those involving the GTP-binding proteins Ras and Rac. The activation of these signaling pathways has been associated with cytoskeletal alterations and increased cellular motility. We hypothesized that such changes could occur in PRL stimulated human breast cancer cells. To test this hypothesis, complementary studies, including wound closure, time-lapse video microscopy (TLVM), and Boyden chamber assay were performed. These studies revealed that PRL significantly enhanced the migration of the breast cancer cell lines T47D, MCF7, and MDA23 1. Co-stimulation with PRL was noted to potentiate epidermal growth factor (EGF) induced cell motility. IF microscopy of filamentous actin using rhodamine conjugated phalloidin revealed a significant and rapid generation of both membrane ruffling and stress fibers in response to PRL, an effect inhibited by the phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002. In sum, these data reveal that PRL stimulation modulates the cytoskeleton and induces the motility of human breast cancer cells in vitro, events that have been associated with the progression of mammary carcinoma in vivo. Given the recently delineated autocrine paracrine role for PRL in human breast cancer, these findings could be of appreciable clinical significance. PMID- 10537180 TI - A nonsteroidal anti-inflammatory drug, flufenamic acid, inhibits the expression of the androgen receptor in LNCaP cells. AB - Nonsteroidal anti-inflammatory drugs (NSAIDs) play potential roles in chemoprevention of colon cancer and others by inhibiting prostaglandin synthesis. In this report, we used LNCaP cells, an androgen-responsive human prostate carcinoma cell line, to study the effects of two NSAIDs, flufenamic acid (FA) and piroxicam (PXM), on the cancer cell growth stimulated by androgens. We found that FA had much higher potency to inhibit LNCaP cell growth than PXM. FA dramatically reduced the expression of androgen inducible genes, such as prostate-specific antigen (PSA) and the homeo-domain transcription factor Nkx3.1, but PXM did not. In vitro transfection experiments showed that FA down regulated the PSA expression at the transcription level. Western and northern blot analyses demonstrated that FA inhibited the androgen receptor (AR) expression at mRNA and protein levels. Suppressed AR expression may be the cause of FA-mediated inhibition of the androgen inducible gene expression. Our data also showed that FA significantly reduced the AR promoter-mediated transcription activities. This study indicated that AR might be a target for FA to inhibit LNCaP cell growth. FA and other similar NSAIDs may be potential candidates for chemoprevention of human prostate cancer by modulating the expression of AR. PMID- 10537181 TI - Differential effects of estradiol and estradiol-BSA conjugates. AB - The steroid 17beta-estradiol (E2) acts to modulate transcription through classical nuclear estrogen receptors (ER-alpha and ER-beta). However, E2 also induces a number of rapid responses (<10 min) within cells, including cells devoid of classical ERs, consistent with the presence of a membrane receptor for E2. Membrane impermeable steroids, typically bovine serum albumin (BSA) conjugates, are commonly used to characterize these non-genomic actions of E2 to exclude the involvement of nuclear ERs. Here we report that E2-BSA conjugate preparations, but not unconjugated E2, activate extracellular signal-regulated protein kinases (ERK1 and ERK2) in the SK-N-SH neuroblastoma cell line, raising concerns regarding the use of these reagents as E2 mimics. Freshly prepared solutions of E2-BSA were found to contain free immunoassayable E2 (iE2), which could be removed by filtration. E2-BSA solutions devoid of free iE2 failed to compete for binding of 125I16alpha-iodo-E2 to ER-alpha or ER-beta. Furthermore, in contrast to E2, E2-BSA conjugates did not bind to ER-alpha or ER-beta as assessed by electrophoretic mobility shift analyses. Protein analysis demonstrated that certain E2-BSA preparations were of very high molecular weight, suggesting extreme protein cross-linking. These findings suggest that E2-BSA does not mimic E2 and is not an appropriate ligand for investigating estrogen receptors. This underscores the need to design stable, cell impermeable analogs of estrogen for the characterization of membrane estrogen receptors. PMID- 10537182 TI - Agouti related peptide (Agrp) stimulates the hypothalamo pituitary gonadal axis in vivo & in vitro in male rats. PMID- 10537183 TI - Nanobacteria and associated 'elementary bodies' in human disease and cancer. PMID- 10537184 TI - Cloning and assembly strategies in microbial genome projects. PMID- 10537185 TI - Microevolutionary changes in Candida albicans identified by the complex Ca3 fingerprinting probe involve insertions and deletions of the full-length repetitive sequence RPS at specific genomic sites. AB - The 11 kb complex DNA fingerprinting probe Ca3 is effective both in cluster analyses of Candida albicans isolates and in identifying microevolutionary changes in the size of hypervariable genomic fragments. A 2.6 kb EcoRI fragment of Ca3, the C fragment, retains the capacity to identify these microevolutionary changes, and when the C fragment is cleaved with SacI, the capacity is retained exclusively by a 1 kb subfragment, C1, which contains a partial RPS repeat element. The microevolutionary changes identified by Ca3, therefore, may involve reorganization of RPS elements dispersed throughout the genome. To test this possibility, hypervariable fragments from several strains of C. albicans were sequenced and compared. The results demonstrate that the microevolutionary changes identified by Ca3 are due to the insertion and deletion of full-length tandem RPS elements at specific genomic sites dispersed throughout the C. albicans genome. The RPS elements at these dispersed sites are bordered by the same upstream and downstream sequences. The frequency of recombination was estimated to be one recombination per 1000 cell divisions by following RPS reorganization in vitro. The results are inconsistent with unequal recombination between homologous or heterologous chromosomes, but consistent with intrachromosomal recombination. Two alternative models of intrachromosomal recombination are proposed: unequal sister-chromatid exchange and slipped misalignment at the replication fork. PMID- 10537186 TI - Repeated extragenic sequences in prokaryotic genomes: a proposal for the origin and dynamics of the RUP element in Streptococcus pneumoniae. AB - A survey of all Streptococcus pneumoniae GenBank/EMBL DNA sequence entries and of the public domain sequence (representing more than 90% of the genome) of an S. pneumoniae type 4 strain allowed identification of 108 copies of a 107-bp-long highly repeated intergenic element called RUP (for repeat unit of pneumococcus). Several features of the element, revealed in this study, led to the proposal that RUP is an insertion sequence (IS)-derivative that could still be mobile. Among these features are: (1) a highly significant homology between the terminal inverted repeats (IRs) of RUPs and of IS630-Spn1, a new putative IS of S. pneumoniae; and (2) insertion at a TA dinucleotide, a characteristic target of several members of the IS630 family. Trans-mobilization of RUP is therefore proposed to be mediated by the transposase of IS630-Spn1. To account for the observation that RUPs are distributed among four subtypes which exhibit different degrees of sequence homogeneity, a scenario is invoked based on successive stages of RUP mobility and non-mobility, depending on whether an active transposase is present or absent. In the latter situation, an active transposase could be reintroduced into the species through natural transformation. Examination of sequences flanking RUP revealed a preferential association with ISs. It also provided evidence that RUPs promote sequence rearrangements, thereby contributing to genome flexibility. The possibility that RUP preferentially targets transforming DNA of foreign origin and subsequently favours disruption/rearrangement of exogenous sequences is discussed. PMID- 10537187 TI - Expression of leading region genes on IncI1 plasmid ColIb-P9: genetic evidence for single-stranded DNA transcription. AB - The leading region of a plasmid is the first sector to enter the recipient cell in bacterial conjugation. This sector of IncI1 plasmid ColIb-P9 includes genes that are transcribed in a transient pulse early in the conjugatively infected cell to promote establishment of the immigrant plasmid. Evidence is presented that the burst of gene expression is regulated by a process which is independent of a repressor but dependent on the orientation of the genes on the unique plasmid strand transferred in conjugation. The nucleotide sequence of 11.7 kb of the leading region was determined and found to contain 10 ORFs; all are orientated such that the template strand for transcription corresponds to the transferred strand. The leading region contains three dispersed repeats of a sequence homologous to a novel promoter in ssDNA described by H. Masai & K. Arai (1997, Cell 89, 897-907). It is proposed that the repeats are promoters that form in the transferring strand of ColIb to support transient transcription of genes transferred early in conjugation. PMID- 10537188 TI - The distribution of enteric bacteria from Australian mammals: host and geographical effects. AB - Bacteria of the family Enterobacteriaceae were isolated from 642 mammalian hosts, representing 16 families and 79 species, collected from throughout Australia. Escherichia coli was the most common of the 24 enteric species recovered and represented almost half of the isolates. Association analysis revealed that most other species of bacteria were less likely to be recovered from hosts in which E. coli was present. The composition of the enteric community of a host was found to be determined by both the taxonomic family to which the host belonged and the geographical area from which the host was collected. Hosts collected from the northern areas of Queensland and the Northern Territory had more diverse enteric communities than hosts collected from New South Wales or Western Australia. Hosts of the families Petauridae and Vespertilionidae had more diverse enteric communities than did members of the Macropodidae or Phalangeridae. The probability of occurrence of Citrobacter freundii, Enterobacter cloacae, Escherichia coli, Hafnia alvei, Klebsiella oxytoca and K. pneumoniae in a host was found to vary with respect to host family and/or host locality. The non random distribution of these species demonstrates the presence of extensive population structure and may suggest the existence of adaptations specific to both the primary and secondary habitats of these enteric bacteria. PMID- 10537189 TI - The genetic structure of enteric bacteria from Australian mammals. AB - A total of 246 isolates representing five species of the family Enterobacteriaceae, taken from a variety of Australian mammal species, were characterized using multi-locus enzyme electrophoresis. Genome diversity estimates varied significantly among species, with the Klebsiella pneumoniae sample exhibiting the lowest diversity and the Citrobacter freundii sample the highest. Multi-locus linkage disequilibrium estimates revealed that alleles were non-randomly associated in all five species samples, but the magnitude of the estimates differed significantly among species. Escherichia coli had the lowest linkage disequilibrium estimate and Klebisella oxytoca the largest. Molecular analyis of variance was used to determine the extent to which population structure explained the observed genetic variation in a species. Two population levels were defined: the taxonomic family of the host from which the isolate was collected and the geographical locality where the host was collected. The amount of explained variation varied from 0% for K. oxytoca to 22% for K. pneumoniae. Host locality explained a significant amount of the genetic variation in the C. freundii (12%), E. coli (5%), Hafnia alvei (17%) and K. pneumoniae (22%) samples. Host family explained a significant fraction of the variation in E. coli (6%) H. alvei (7%) and K. pneumoniae (20%). Estimates of effective population size for all five species, based on the probability that two randomly chosen isolates will be identical, failed to reveal any relationship between the effective population size and the genetic diversity of a species. PMID- 10537190 TI - In vivo detection of Escherichia coli type 1 fimbrial expression and phase variation during experimental urinary tract infection. AB - Adhesion mediated by fimbriae is thought to play an important role in the pathogenesis of urinary tract infections (UTI) by Escherichia coli. The majority of clinical isolates of E. coli from UTI are able to express type 1 fimbriae. However, the importance of these fimbriae as a virulence factor has been controversial. To investigate the expression of type 1 fimbriae in vivo during UTI, mice were transurethrally infected with uropathogenic E. coli C175-94 and type 1 fimbrial expression was determined directly by two independent methods at 2 h, 1 d and 3 d after infection. By use of an assay combining in situ rRNA hybridization and immunofluorescence, all bacterial cells detected in urine, bladders and kidneys from mice sacrificed 1 and 3 d after onset of infection were found to express type 1 fimbriae. In contrast, the majority of cells in the suspension used for infection of mice and specimens from mice sacrificed 2 h after inoculation were found to be non-fimbriated. Similar results were obtained with a PCR assay revealing the orientation of the invertible promoter driving the transcription of type 1 fimbrial genes. Whilst the promoter in both ON and OFF positions could be amplified from the suspension used for infection and specimens from mice sacrificed 2 h after inoculation, at 1 and 3 d after onset of infection only the promoter in the ON orientation could be amplified. These results show that introduction of E. coli C175-94 into the mouse urinary tract leads to markedly enhanced expression of type 1 fimbriae. PMID- 10537191 TI - The entomopathogenic fungus Metarhizium anisopliae alters ambient pH, allowing extracellular protease production and activity. AB - Ambient pH regulates the expression of virulence genes of Metarhizium anisopliae, but it was unknown if M. anisopliae can regulate ambient pH. Mutants of M. anisopliae altered in production of oxalic acid were evaluated for the interrelationship of ambient pH, buffering capacity added to media, growth, and generation of extracellular proteases and ammonia. Wild-type and acid overproducing mutants [Acid(+)] grew almost as well at pH 8 as at pH 6, but acid non-producing [Acid(-)] mutants showed limited growth at pH 8, indicating that acid production is linked to the ability to grow at higher pH. Production of ammonia by M. anisopliae was strongly stimulated by low levels of amino acids in the medium when cells were derepressed for nitrogen and carbon. Likewise, although Aspergillus fumigatus and Neurospora crassa produced some ammonia in minimal media, addition of low levels of amino acids enhanced production. Ammonia production by A. fumigatus, N. crassa and M. anisopliae increased the pH of the medium and allowed production of subtilisin proteases, whose activities are observed only at basic pH. In contrast, protease production by the Acid(+) mutants of M. anisopliae was greatly reduced because of the acidification of the medium. This suggests that alkalinization by ammonia production is adaptive by facilitating the utilization of proteinaceous nutrients. Collectively, the data imply that ammonia may have functions related to regulation of the microenvironment and that it represents a previously unconsidered virulence factor in diverse fungi with the potential to harm tissues and disturb the host's immune system. PMID- 10537192 TI - Contribution of mutations in the cytochrome P450 14alpha-demethylase (Erg11p, Cyp51p) to azole resistance in Candida albicans. AB - The cytochrome P450 14alpha-demethylase, encoded by the ERG11 (CYP51) gene, is the primary target for the azole class of antifungals. Changes in the azole affinity of this enzyme caused by amino acid substitutions have been reported as a resistance mechanism. Nine Candida albicans strains were used in this study. The ERG11 base sequence of seven isolates, of which only two were azole sensitive, were determined. The ERG11 base sequences of the other two strains have been published previously. In these seven isolates, 12 different amino acid substitutions were identified, of which six have not been described previously (A149V, D153E, E165Y, S279F, V452A and G4655). In addition, 16 silent mutations were found. Two different biochemical assays, subcellular sterol biosynthesis and CO binding to reduced microsomal fractions, were used to evaluate the sensitivity of the cytochromes for fluconazole and itraconazole. Enzyme preparations from four isolates showed reduced itraconazole susceptibility, whereas more pronounced resistance to fluconazole was observed in five isolates. A three-dimensional model of C. albicans Cyp51p was used to position all 29 reported substitutions, 98 in total identified in 53 sequences. These 29 substitutions were not randomly distributed over the sequence but clustered in three regions from amino acids 105 to 165, from 266 to 287 and from 405 to 488, suggesting the existence of hotspot regions. Of the mutations found in the two N-terminal regions only Y132H was demonstrated to be of importance for azole resistance. In the C-terminal region three mutations are associated with resistance, suggesting that the non characterized substitutions found in this region should be prioritized for further analysis. PMID- 10537193 TI - Multiple amino acid substitutions in lanosterol 14alpha-demethylase contribute to azole resistance in Candida albicans. AB - Lanosterol 14alpha-demethylase (14DM) is the target of the azole antifungals, and alteration of the 14DM sequence leading to a decreased affinity of the enzyme for azoles is one of several potential mechanisms for resistance to these drugs in Candida albicans. In order to identify such alterations the authors investigated a collection of 19 C. albicans clinical isolates demonstrating either frank resistance (MICs > or = 32 microg ml(-1)) or dose-dependent resistance (MICs 8-16 microg ml(-1)) to fluconazole. In cell-free extracts from four isolates, including the Darlington strain ATCC 64124, sensitivity of sterol biosynthesis to inhibition by fluconazole was greatly reduced, suggesting that alterations in the activity or affinity of the 14DM could contribute to resistance. Cloning and sequencing of the 14DM gene from these isolates revealed 12 different alterations (two to four per isolate) leading to changes in the deduced amino acid sequence. Five of these mutations have not previously been reported. To demonstrate that these alterations could affect fungal susceptibility to azoles, the 14DM genes from one sensitive and three resistant C. albicans strains were tagged at the carboxyl terminus with a c-myc epitope and expressed in Saccharomyces cerevisiae under control of the endogenous promoter. Transformants receiving 14DM genes from resistant strains had fluconazole MICs up to 32-fold higher than those of transformants receiving 14DM from a sensitive strain, although Western blot analysis indicated that the level of expressed 14DM was similar in all transformants. Amino acid substitutions in the 14DM gene from the Darlington strain also conferred a strong cross-resistance to ketoconazole. In conclusion, multiple genetic alterations in C. albicans 14DM, including several not previously reported, can affect the affinity of the enzyme for azoles and contribute to resistance of clinical isolates. PMID- 10537194 TI - Candida albicans and Yarrowia lipolytica as alternative models for analysing budding patterns and germ tube formation in dimorphic fungi. AB - The site for bud selection and germ tube emission in two yeasts, Candida albicans and Yarrowia lipolytica, was analysed. Both dimorphic organisms display different patterns of budding, which also differ from those described for Saccharomyces cerevisiae. C. albicans, which is diploid and (until now) lacks a known sexual cycle, buds in an axial budding pattern. During the yeast-hypha transition induced by pH, serum, N-acetylglucosamine (GlcNAc) or temperature, germ tube emergence occurs at approximately 50% in a polar manner, while the other 50% of cells show non-polar germ tube emission. Y. lipolytica, in which most of the natural isolates are haploid and which has a well characterized sexual cycle, buds with a polar budding pattern independently of the degree of ploidy. Germ tube emission during the yeast-hypha transition in both haploid and diploid cells generally occurs at the pole distal from the division site (bipolar). The addition of hydroxyurea (HU), an inhibitor of DNA synthesis, also produces different effects. In its presence, and therefore in the absence of DNA synthesis, the yeast-hypha transition is completely abolished in Y. lipolytica. By contrast, in C. albicans germ tube emission in the presence of HU is similar to that observed in control cultures for at least 90 min under induction conditions. These results demonstrate that, rather than a single developmental model, several models of development should be invoked to account for the processes involved in the morphological switch in yeasts (the yeast-hypha transition). PMID- 10537195 TI - Involvement of glutathione in the regulation of respiratory oscillation during a continuous culture of Saccharomyces cerevisiae. AB - Respiratory oscillation occurred during aerobic continuous culture of Saccharomyces cerevisiae. During oscillation, phase-related changes in NAD(P)H and GSH levels occur. Perturbation of oscillation and inhibition of respiration occurred when GSH or GSSG was injected; however, there was a phase delay in perturbation in the case of an injection during high respiration. The perturbation phase delay was not apparent when a combination of DL-buthionine (S,R)-sulphoximine, GSH and 5-nitro-2-furaldehyde was injected. Perturbation by GSH injection caused the intracellular GSH concentration to increase, the GSSG concentration to decrease and the cessation of ethanol uptake. NAD(P)H during perturbation was inversely related to dissolved oxygen. Perturbation by calcium pantothenate and pyridoxal-HCl caused a period of enhanced respiration before oscillation returned. These results suggest that the NAD+/NADH redox is not directly involved in oscillation control and regulation involves glutathione metabolism. Possible regulation points include alcohol dehydrogenase inhibition and/or respiratory-chain inhibition. PMID- 10537196 TI - Morphogenesis is coordinated with nuclear division in germinating Aspergillus nidulans conidiospores. AB - Germinating Aspergillus nidulans conidiospores switch to polarized apical growth following an initial period of isotropic expansion. At the same time, they re enter the nuclear division cycle. The relationship between spore polarization and nuclear division was investigated by testing the effect of cell cycle inhibitors and temperature-sensitive cell cycle mutations on spore morphogenesis. On rich media, it was found that spore polarization is delayed if completion of the first mitosis is blocked. The observed delay may be dependent upon the activity of the mitosis-promoting NIMA kinase. An additional mechanism appears to prevent polarization as the spore progresses through its first S phase. In contrast, on poor media, spore polarization does not require completion of the first mitosis. These observations suggest that spore morphogenesis is influenced by cell cycle signals in a growth-dependent manner. PMID- 10537197 TI - The diversity of gas vesicle genes in Planktothrix rubescens from Lake Zurich. AB - Part of the gas vesicle gene cluster was amplified by PCR from three strains of Planktothrix rubescens isolated from Lake Zurich, Switzerland. Each contains multiple alternating copies of gvpA and gvpC. All of the gvpA sequences in the different strains are identical. There are two types of gvpC: gvpC20, of length 516 bp, encodes a 20 kDa protein of 172 amino acid residues (whose N-terminal amino acid sequence is homologous with the sequence of GvpC in Planktothrix [Oscillatoria] agardhii); gvpC16, of length 417 bp, encodes a 16 kDa protein of 139 amino acid residues that differs in lacking an internal 33-residue section. An untranslated 72 bp fragment from the 3' end of gvpC, designated omegaC, is also present in some strains. The two types of gvpC and presence of omegaC could be distinguished by the different lengths of PCR amplification products obtained using pairs of oligonucleotide primers homologous to internal sequences in gvpC and gvpA. Three genotype classes were found: GV1, containing only gvpC20; GV2, containing gvpC20 and omegaC; and GV3, containing gvpC16, gvpC20 and omegaC. Subclasses of GV2 and GV3 contained either one or two copies of omegaC. The accompanying paper by D. I. Bright & A. E. Walsby (Microbiology 145, 2769-2775) shows that strains of the GV3 genotype produce gas vesicles with a higher critical pressure than those of GV1 and GV2. A PCR survey of 185 clonal cultures of P. rubescens isolated from Lake Zurich revealed that 3 isolates were of genotype GV1, 73 were of GV2 and 109 were of GV3. The PCR technique was used to distinguish the gas vesicle genotype, and thence the associated critical-pressure phenotype, of single filaments selected from lakewater samples. Sequence analysis of the 16S rDNA and of regions within the operons encoding phycoerythrin, phycocyanin and Rubisco confirmed that these strains of Planktothrix form a tight phylogenetic group. PMID- 10537198 TI - The relationship between critical pressure and width of gas vesicles in isolates of Planktothrix rubescens from Lake Zurich. AB - The mean critical collapse pressure (p(c)) of gas vesicles in 81 strains of the cyanobacterium Planktothrix rubescens from Lake Zurich, Switzerland, was bimodally distributed between a minimum of 0.86 MPa and a maximum of 1.17 MPa. Measurements were made of the cylinder diameter (d) of gas vesicles isolated from seven of the strains. The mean diameter, which varied from 48 to 61 nm, was inversely related to p(c), in keeping with the theory of strength of thin-walled rigid cylinders. These measurements extended the range of p(c)-width relationship of gas vesicles, which can be described by the expression p(c) = 461(d/nm)(-1.53) MPa. p(c) was correlated with gas vesicle genotype (see the accompanying paper by S. J. Beard, B. A. Handley, P. K. Hayes & A. E. Walsby, Microbiology 145, 2757 2768): of the 81 strains investigated, all those with the gas vesicle genotype GV2 produced gas vesicles with a mean p(c) of less than 1.0 MPa, whereas those of GV3 had a mean p(c) of greater than 1.0 MPa. It is suggested that gas vesicles of the GV3 strains, which are narrower and stronger than any previously recorded in freshwater cyanobacteria, have evolved to withstand the high hydrostatic pressures during deep winter mixing in Lake Zurich. PMID- 10537199 TI - Antibodies to a synthetic 1-9-N-terminal amino acid fragment of mature pediocin PA-1: sensitivity and specificity for pediocin PA-1 and cross-reactivity against Class IIa bacteriocins. AB - Polyclonal antibodies specific for pediocin PA-1 (PedA1) were generated by immunization of rabbits with a chemically synthesized 1-9-N-terminal amino acid fragment of this bacteriocin (PH1) conjugated to the carrier protein keyhole limpet haemocyanin (KLH). The PH1 fragment holds a highly conserved amino acid sequence with closely related Class IIa bacteriocins. The sensitivity and specificity of the PH1-KLH-generated rabbit polyclonal antibodies were evaluated by the development of various ELISAs, such as a non-competitive indirect ELISA (NCI-ELISA), a competitive indirect ELISA (CI-ELISA), a competitive direct ELISA (CD-ELISA) and a sandwich ELISA (S-ELISA), and by protein slot-blotting and Western blotting. NCI- and CI-ELISA were valuable for detecting the existence of PedA1-specific antibodies in the sera of immunized rabbits. The limit of detection of PedA1 in MRS medium was found to be 0.5 microg ml(-1) in NCI-ELISA, while CI-ELISA on plates coated with purified PedA1 increased the affinity of the PH1-KLH-generated antibodies for PedA1; the limit of detection of PedA1 was less than 0.01 microg ml(-1) and 50% binding inhibition was achieved with 0.1 microg PedA1 ml(-1). Similarly, the limits of detection of PedA1 in MRS medium were found to be 5 microg ml(-1) by protein slot-blotting and 0.01 microg ml(-1) by Western blotting. Most importantly, PH1-KLH-generated polyclonal antibodies detected the presence of PedA1 in the supernatants of the producing strains of Pediococcus acidilactici 347, Z102, A172, X13 and P20, with no reactivity or negligible immunoreactivity with the supernatants of other lactic acid bacteria producing or not producing closely related or different bacteriocins. The approaches taken for the selection of the bacteriocin peptide fragment, the generation of antibodies and the development of immunoassays could prove useful for the generation and evaluation of antibodies of adequate specificity for other bacteriocins of interest in the food industry. PMID- 10537200 TI - Galactomannans from the cell walls of species of Paecilomyces sect. Paecilomyces and their teleomorphs as immunotaxonomic markers. AB - An alkali-extractable and water-soluble fraction (F1S) was obtained from cell walls of Paecilomyces variotii and species of the related genera Talaromyces, Byssochlamys and Thermoascus. The structure of the main polysaccharide of these fractions was studied and found to consist of a core of (1 --> 6)-alpha mannopyranose partially substituted at 0.2 by chains of galactofuranose and shorter chains of mannopyranose. The differences in the regularity of the branching points and the length of the galactofuranose side chains are useful to distinguish between species. These differences were detected by immunological methods, since highly specific polyclonal antibodies were raised against these polysaccharides. Mycelium of P. variotii CBS 990.73A was stained by indirect immunofluorescence. The polysaccharides studied in this work differ from the one described for species from section Isarioidea, and this is another indication of the heterogeneity of the genus Paecilomyces. PMID- 10537201 TI - The effect of motility and cell-surface polymers on bacterial attachment. AB - Recently it was shown that motility of Vibrio alginolyticus facilitated cell attachment to glass surfaces. In the present study the same relationship between motility and cell attachment was confirmed for Alcaligenes and Alteromonas spp. These findings clearly answer a long-standing question: does motility facilitate attachment? However, they are contradictory to a general view on cell attachment that the energy barrier due to electrostatic repulsion between negatively charged bacterial cells and a glass surface is much greater than both the thermal kinetic energy of the bacterial cell and the bacterial swimming energy. It is shown that the energy barrier becomes far less than that usually estimated when bacterial cells are rich in polymers at their surfaces. This finding reasonably explains the dependence of bacterial attachment rate on cell motility and demands reconsideration of the mechanism of bacterial attachment. PMID- 10537202 TI - The relationship between Helicobacter pylori motility, morphology and phase of growth: implications for gastric colonization and pathology. AB - To explore the relationship between Helicobacter pylori motility, morphology and phase of growth, bacteria were isolated from antral biopsies of patients with duodenal ulcer or non-ulcer dyspepsia, and grown in liquid medium in batch and continuous culture systems. Motilities and morphologies of H. pylori in different phases of growth were examined with a Hobson BackTracker and by transmission electron microscopy. Morphologies of bacteria grown in vitro were also compared with those of bacteria in antral biopsies from patients with non-autoimmune gastritis. H. pylori had poor motility in lag phase, became highly motile in mid exponential phase and lost motility in the decline phase of growth. Motilities of bacteria in the same phase of growth from patients with duodenal ulcer or non ulcer dyspepsia were not significantly different. In the mid/late-exponential phase of growth bacteria had helical morphologies and multiple polar flagella, typical of H. pylori in the gastric mucus layer. In the decline phase of growth bacteria shed flagella, and had precoccoidal or coccoidal morphologies. These findings support the view that helical and coccoidal H. pylori are in different phases of growth with different roles in gastric colonization, indicate that bacterial motility per se is unlikely to be a determinant of H. pylori pathology, and suggest that H. pylori in the antral mucus layer is in a state of continuous (exponential phase) growth. PMID- 10537203 TI - Genetic organization and characteristics of the 3-(3-hydroxyphenyl)propionic acid degradation pathway of Comamonas testosteroni TA441. AB - Comamonas testosteroni TA441 degrades 3-(3-hydroxyphenyl)propionate (3HPP) via the meta pathway. A gene cluster required for degradation of 3HPP was cloned from strain TA441 and sequenced. The genes encoding six catabolic enzymes, a flavin type hydroxylase (mhpA), extradiol dioxygenase (mhpB), 2-keto-4-pentenoate hydratase (mhpD), acetaldehyde dehydrogenase (acylating) (mhpF), 4-hydroxy-2 ketovalerate aldolase (mhpE) and the meta cleavage compound hydrolase (mhpC), were found in this cluster, encoded in this order. mhpD and mhpF were separated by two genes, orf4 and orf5, which were not necessary for growth on 3HPP. The gene mhpR, encoding a putative transcriptional activator of the IcIR family, was located adjacent to mhpA in the opposite orientation. Disruption of the mhpB or mhpR genes affected growth on 3HPP or trans-3-hydroxycinnamate. The mhpB and mhpC gene products showed high specificity for 3-(2,3-dihydroxyphenyl)propionate (DHPP) and the meta cleavage compound produced from DHPP, respectively. PMID- 10537204 TI - Occurrence and expression of glutathione-S-transferase-encoding bphK genes in Burkholderia sp. strain LB400 and other biphenyl-utilizing bacteria. AB - The gene bphK of Burkholderia sp. strain LB400 has previously been shown to be located within the bph locus, which specifies the degradation of biphenyl (BP) and chlorobiphenyls, and to encode a glutathione S-transferase (GST) which accepts 1-chloro-2,4-dinitrobenzene (CDNB) as substrate. The specific physiological role of this gene is not known. It is now shown that the gene is expressed in the parental organism and that GST activity is induced more than 20 fold by growth of the strain on BP relative to succinate when these compounds serve as sole carbon source. Approximately the same induction factor was observed for 2,3-dihydroxybiphenyl 1,2-dioxygenase activity, which is encoded by the 5' adjacent bphC gene. This suggests that the expression of bphK is coregulated with the expression of genes responsible for the catabolism of BP. A bphK probe detected only a single copy of the gene in strain LB400. A spontaneous BP- mutant of the organism neither gave a signal with the bphK probe nor showed CDNB accepting GST activity, suggesting that this activity is solely encoded by bphK. Complementation of the mutant with a bph gene cluster devoid of bphK restored the ability to grow on BP, indicating that bphK is not essential for utilization of this carbon source. BphK activity proved to be almost unaffected by up to 100 fold differences in proton concentration or ionic strength. The enzyme showed a narrow range with respect to a variety of widely used electrophilic GST substrates, accepting only CDNB. A number of established laboratory strains as well as novel isolates able to grow on BP as sole carbon and energy source were examined for BphK activity and the presence of a bphK analogue. CDNB assays, probe hybridizations and PCR showed that several, but not all, BP degraders possess this type of GST activity and/or a closely related gene. In all bacteria showing BphK activity, this was induced by growth on BP as sole carbon source, although activity levels differed by up to 10-fold after growth on BP and by up to 60-fold after growth on succinate. This resulted in a variation of induction factors between 2 and 30. In the majority of bphK+ bacteria examined, the gene appeared to be part of LB400-like bph gene clusters. DNA sequencing revealed almost complete identity of bphK genes from five different bph gene clusters. These results suggest that bphK genes, although not essential, fulfill a strain specific function related to the utilization of BPs by their host organisms. The usefulness of BphK as a reporter enzyme for monitoring the expression of catabolic pathways is discussed. PMID- 10537205 TI - Trichomonas vaginalis interactions with fibronectin and laminin. AB - The sexually transmitted protozoan Trichomonas vaginalis cytoadheres to vaginal epithelial cells and causes contact-dependent cytotoxicity which, when combined with the normal exfoliation process, leads to erosion of the epithelium, which may allow trichomonads into extracellular matrix and basement membrane sites. Therefore, the association of T. vaginalis with immobilized fibronectin (FN) and laminin (LM) on cover-slips was examined. Binding of live parasites to coated cover-slips was time- and parasite-density-dependent. Coincubation with an inhibitor of trichomonad cysteine proteinases resulted in an increased attachment of parasites to FN but had no effect on binding to LM, denoting that protease activity influenced optimal FN associations. Further, 20 h mid-exponential phase trichomonads placed in fresh culture medium for 3 h gave higher levels of binding to FN, suggesting that changes during growth in vitro to T. vaginalis organisms affect maximal levels of binding to FN. Extended incubation with substrates diminished the capacity of parasites to bind FN and LM. Treatment of live organisms with periodate reduced binding to LM but not FN, suggesting a role for carbohydrates. In addition, trypsinization of live parasites decreased numbers bound to both substrates. Placement of trypsinized parasites in medium for 2 h fully regenerated binding to FN but not LM. Incubation of trypsinized parasites with cycloheximide abrogated regeneration of attachment to FN, affirming a role for synthesized surface proteins in FN binding. Importantly, the T. vaginalis adhesin proteins that mediate cytoadherence, and iron, a factor that regulates adhesin synthesis, were not involved in FN and LM recognition. These results suggest a role for surface proteins and carbohydrates in trichomonal associations with FN and LM, respectively. PMID- 10537206 TI - The site-specific integration of genetic elements may modulate thermostable protease production, a virulence factor in Dichelobacter nodosus, the causative agent of ovine footrot. AB - The gram-negative anaerobe Dichelobacter nodosus is the causative agent of footrot in sheep. The authors have previously characterized two genetic elements, the intA (vap) and intB elements, which integrate into the genome of D. nodosus. In the virulent strain A198 there are two copies of the intA element. One copy is integrated into the 3' end of the tRNA-serGCU gene, close to the aspartokinase (askA) gene, and the second copy is integrated into the 3' end of the tRNA-serGGA gene, next to the polynucleotide phosphorylase (pnpA) gene. In this study, a new genetic element was identified in the benign strain C305, the intC element, integrated into the 3' end of the tRNA-serGCU gene, next to askA. The intC element was found in most D. nodosus strains, both benign and virulent, which were examined, and was integrated into tRNA-serGCU in most strains. Between the askA and tRNA-serGCU genes, a gene (designated glpA), was identified whose predicted protein product has very high amino acid identity with RsmA from the plant pathogen Erwinia carotovora. RsmA acts as a global repressor of pathogenicity in E. carotovora, by repressing the production of extracellular enzymes. In virulent strains of D. nodosus the intA element was found to be integrated next to pnpA, and either the intA or intC element was integrated next to glpA. By contrast, all but one of the benign strains had intB at one or both of these two positions, and the one exception had neither intA, intB nor intC at one position. The loss of the intC element from the virulent strain 1311 resulted in loss of thermostable protease activity, a virulence factor in D. nodosus. A model for virulence is proposed whereby integration of the intA and intC genetic elements modulates virulence by altering the expression of glpA, pnpA, tRNA serGCU and tRNA-serGGA. PMID- 10537207 TI - Emergence of multidrug-resistant mutants is increased under antibiotic selective pressure in Pseudomonas aeruginosa. AB - Pseudomonas aeruginosa is one of the most important opportunistic pathogens involved in nosocomial infections, cystic fibrosis patients included. Hospital isolates frequently present multidrug-resistance (MDR) phenotypes as the consequence of constant antibiotic selective pressure. The kinetics of emergence of P. aeruginosa MDR mutants under antibiotic selective pressure indicated that long-term incubation in the presence of the bacteriostatic antibiotic tetracycline increases the mutation rate per cell per day of P. aeruginosa PAO1 by several orders of magnitude. The tetracycline-resistant mutants obtained were stable, showed decreased susceptibility to antibiotics belonging to different structural families, and contained an outer-membrane protein not present in the wild-type P. aeruginosa strain PAO1. These data are consistent with the hypothesis that incubation in the presence of tetracycline favours the emergence of MDR mutants in P. aeruginosa. The results are relevant for understanding the rapid emergence of antibiotic-resistant mutants among bacterial populations during infections. Their relationship to other models of increased mutagenesis under stress is discussed with respect to the adaptive mutation phenomenon. PMID- 10537208 TI - A re-examination of twitching motility in Pseudomonas aeruginosa. AB - Twitching motility is a form of solid surface translocation which occurs in a wide range of bacteria and which is dependent on the presence of functional type IV fimbriae or pili. A detailed examination of twitching motility in Pseudomonas aeruginosa under optimal conditions in vitro was carried out. Under these conditions (at the smooth surface formed between semi-solid growth media and plastic or glass surfaces) twitching motility is extremely rapid, leading to an overall radial rate of colony expansion of 0.6 mm h(-1) or greater. The zones of colony expansion due to twitching motility are very thin and are best visualized by staining. These zones exhibit concentric rings in which there is a high density of microcolonies, which may reflect periods of expansion and consolidation/cell division. Video microscopic analysis showed that twitching motility involves the initial formation of large projections or rafts of aggregated cells which move away from the colony edge. Behind the rafts, individual cells move rapidly up and down trails which thin and branch out, ultimately forming a fine lattice-like network of cells. The bacteria in the lattice network then appear to settle and divide to fill out the colonized space. Our observations redefine twitching motility as a rapid, highly organized mechanism of bacterial translocation by which P. aeruginosa can disperse itself over large areas to colonize new territories. It is also now clear, both morphologically and genetically, that twitching motility and social gliding motility, such as occurs in Myxococcus xanthus, are essentially the same process. PMID- 10537209 TI - A low-Mr lipase activation factor cooperating with lipase modulator protein LimL in Pseudomonas sp. strain 109. AB - Pseudomonas sp. strain 109 produces a unique lipase (LipL) which efficiently catalyses intramolecular transesterification of omega-hydroxyesters to form macrocyclic lactones. In vivo production of enzymically active LipL requires lipase modulator protein (LimL), which functions as a molecular chaperone for the correct folding of LipL. However, previous work has shown that LipL forms a tight complex with LimL in vitro and the resulting LipL-LimL complex is only partially active, suggesting an additional mechanism that facilitates the dissociation of the complex to form enzymically active LipL. In the present work, a low-Mr compound (lipase activation factor, LAF) was found in Pseudomonas sp. strain 109 that when added to the LipL-LimL complex resulted in the activation of LipL. Ca2+ ions also enhanced lipase activity, but the instantaneous activation by Ca2+ was different from the gradual and time-dependent activation by LAF, indicating the novel nature of this compound. LAF passed through an ultrafiltration membrane with an Mr cut-off of 3000 and showed an apparent Mr of 330+/-30 on Superdex Peptide gel-filtration chromatography. Treatment of the LipL-LimL complex with LAF liberated free active LipL, indicating that LAF was necessary to dissociate the LipL-LimL complex. PMID- 10537210 TI - Staphylococcal phosphoenolpyruvate-dependent phosphotransferase system--two highly similar glucose permeases in Staphylococcus carnosus with different glucoside specificity: protein engineering in vivo? AB - Previous sequence analysis of the glucose-specific PTS gene locus from Staphylococcus carnosus revealed the unexpected finding of two adjacent, highly similar ORFs, glcA and glcB, each encoding a glucose-specific membrane permease EIICBA(Glc). glcA and glcB show 73% identity at the nucleotide level and glcB is located 131 bp downstream from glcA. Each of the genes is flanked by putative regulatory elements such as a termination stem-loop, promoter and ribosome binding site, suggesting independent regulation. The finding of putative cis active operator sequences, CRE (catabolite-responsive elements) suggests additional regulation by carbon catabolite repression. As described previously by the authors, both genes can be expressed in Escherichia coli under control of their own promoters. Two putative promoters are located upstream of glcA, and both were found to initiate transcription in E. coli. Although the two permeases EIICBA(Glc)1 and EIICBA(Glc)2 show 69% identity at the protein level, and despite the common primary substrate glucose, they have different specificities towards glucosides as substrate. EIICBA(Glc)1 phosphorylates glucose in a PEP-dependent reaction with a Km of 12 microM; the reaction can be inhibited by 2-deoxyglucose and methyl beta-D-glucoside. EIICBA(Glc)2 phosphorylates glucose with a Km of 19 microM and this reaction is inhibited by methyl alpha-D-glucoside, methyl beta-D glucoside, p-nitrophenyl alpha-D-glucoside, o-nitrophenyl beta-D-glucoside and salicin, but unlike other glucose permeases, including EIICBA(Glc)1, not by 2 deoxyglucose. Natural mono- or disaccharides, such as mannose or N acetylglucosamine, that are transported by other glucose transporters are not phosphorylated by either EIICBA(Glc)1 nor EIICBA(Glc)2, indicating a high specificity for glucose. Together, these findings support the suggestion of evolutionary development of different members of a protein family, by gene duplication and subsequent differentiation. C-terminal fusion of a histidine hexapeptide to both gene products did not affect the activity of the enzymes and allowed their purification by Ni2+-NTA affinity chromatography after expression in a ptsG (EIICB(Glc)) deletion mutant of E. coli. Upstream of glcA, the 3' end of a further ORF encoding 138 amino acid residues of a putative antiterminator of the BglG family was found, as well as a putative target DNA sequence (RAT), which indicates a further regulation by glucose specific antitermination. PMID- 10537211 TI - Substrate specificity of the periplasmic dipeptide-binding protein from Escherichia coli: experimental basis for the design of peptide prodrugs. AB - Pure dipeptide-binding protein (DppA) from Escherichia coli was studied in a filter binding assay to determine its binding specificity. A substrate:DppA stoichiometry of 1:1 was found with both [14C]AlaAla and Ala[14C]Phe. Surprisingly, substrate binding did not vary over the pH range pH 3-9.5. Different dipeptides yielded liganded protein with various pI values, implying that DppA can undergo subtly different conformational changes to accommodate different substrates. Using [125I]Tyr-peptides as substrates in competition assays, the relative binding affinities for a range of dipeptides were found to parallel their overall transport rates into E. coli through the dipeptide permease (Dpp), showing that DppA alone controls the specificity of Dpp. With a series of substituted glycyl peptides, binding affinity was progressively enhanced by alkylation (with methyl to butyl) of the N-terminal alpha-amino group. Thus, results from this approach provide an essential experimental basis, which complements the information from the crystal structure of DppA, for the design of peptidomimetic antibacterials targeted for transport through Dpp. PMID- 10537212 TI - Transcriptional regulation in response to oxygen and nitrate of the operons encoding the [NiFe] hydrogenases 1 and 2 of Escherichia coli. AB - Synthesis of the [NiFe] hydrogenases 1 and 2 of Escherichia coli is induced in response to anaerobiosis and is repressed when nitrate is present in the growth medium. The hydrogenase 1 and hydrogenase 2 enzymes are encoded by the polycistronic hyaABCDEF and hybOABCDEFG operons, respectively. Primer extension analysis was used to determine the initiation site of transcription of both operons. This permitted the construction of single-copy lacZ operon fusions, which were used to examine the transcriptional regulation of the two operons. Expression of both was induced by anaerobiosis and repressed by nitrate, which is in complete accord with earlier biochemical studies. Anaerobic induction of the hyb operon was only partially dependent on the FNR protein and, surprisingly, was enhanced by an arcA mutation. This latter result indicated that ArcA suppresses anaerobic hyb expression and that a further factor, which remains to be identified, is involved in controlling anaerobic induction of operon expression. Nitrate repression of hyb expression was mediated by the NarL/NarX and NarP/NarQ two-component regulatory systems. Remarkably, a narP mutant lacked anaerobic induction of hyb expression, even in the absence of added nitrate. Anaerobic induction of hya expression was dependent on the ArcA and AppY regulators, which confirms earlier observations by other authors. Nitrate repression of the hya operon was mediated by both NarL and NarP. Taken together, these data indicate that although the hya and hyb operons share common regulators, there are important differences in the control of expression of the individual operons. PMID- 10537213 TI - The dnaA gene region of Mycobacterium avium and the autonomous replication activities of its 5' and 3' flanking regions. AB - A 3.9 kb DNA fragment containing the dnaA gene region of Mycobacterium avium was cloned and its nucleotide sequence was determined. Nucleotide sequence analyses indicated that this region encodes three genes in the order rpmH (ribosomal protein L34), dnaA (the putative initiator protein) and dnaN (the beta subunit of DNA polymerase III). The intergenic regions between the rpmH-dnaA and dnaA-dnaN genes were found to contain several putative DnaA boxes, 9 nt long DnaA protein recognition sequences. A DNA fragment containing the 3' but not the 5' flanking region of the M. avium dnaA gene when cloned in Escherichia coli plasmids, which are otherwise non-replicative in mycobacteria, exhibited autonomous replication activity in M. avium but not in Mycobacterium bovis BCG and Mycobacterium smegmatis. The 5' flanking region of dnaA, on the other hand, exhibited autonomous replication activity in M. bovis BCG but not in M. avium and M. smegmatis. The implications of these results for the understanding of the M. avium oriC replication initiation process are discussed. PMID- 10537214 TI - Use of fluorescence induction and sucrose counterselection to identify Mycobacterium tuberculosis genes expressed within host cells. AB - The identification of Mycobacterium tuberculosis genes expressed within host cells would contribute greatly to the development of new strategies to combat tuberculosis. By combining the natural fluorescence of the Aequoria victoria green fluorescent protein (GFP) with the counterselectable property of the Bacillus subtilis SacB protein, M. tuberculosis promoters displaying enhanced in vivo activity have been isolated. Macrophages were infected with recombinant Mycobacterium bovis bacille Calmette-Guerin containing a library of M. tuberculosis promoters controlling gfp and sacB expression, and fluorescent bacteria recovered by fluorescence-activated cell sorting. The expression of sacB was used to eliminate clones with strong promoter activity outside the macrophage, resulting in the isolation of seven clones containing M. tuberculosis promoters with greater activity intracellularly. The gene products identified displayed similarity to proteins from other organisms whose functions include nutrient utilization, protection from oxidative stress and defence against xenobiotics. These proposed functions are consistent with conditions encountered within the host cell and thus suggest that the augmented activity of the isolated promoters/genes may represent strategies employed by M. tuberculosis to enhance intracellular survival and promote infection. PMID- 10537215 TI - Regulation of polyphosphate kinase gene expression in Acinetobacter baumannii 252. AB - A strain of Acinetobacter baumannii cultured in butyric acid media was found to take up phosphate following a period of phosphate release. PCR was used to clone the polyphosphate kinase (ppk) gene from the strain. The promoter for the ppk gene was functional in the heterologous Escherichia coli host. Using RT-PCR, transcription of the ppk gene was found to be regulated by phosphate concentration. PMID- 10537216 TI - Two fatty acid delta9-desaturase genes, ole1 and ole2, from Mortierella alpina complement the yeast ole1 mutation. AB - Genes encoding two distinct fatty acid delta9-desaturases were isolated from strains of the oleaginous fungus Mortierella alpina. Two genomic sequences, delta9-1 and delta9-2, each containing a single intron, were cloned from strain CBS 528.72 while one cDNA clone, LM9, was isolated from strain CBS 210.32. The delta9-1 gene encoded a protein of 445 aa which shared 99% identity with the LM9 gene product. These proteins also showed 40-60% identity to the delta9 desaturases (Ole1p) of other fungi and contained the three conserved histidine boxes, C-terminal cytochrome b5 fusion and transmembrane domains characteristic of endoplasmic reticulum membrane-bound delta9-desaturases. LM9 and delta9-1 are therefore considered to represent the same gene (ole1). The ole1 gene was transcriptionally active in all M. alpina strains tested and its function was confirmed by complementation of the Saccharomyces cerevisiae ole1 mutation. Fatty acid analysis of yeast transformants expressing the CBS 210.32 ole1 gene showed an elevated level of oleic acid (18:1) compared to palmitoleic acid (16:1), the major fatty acid component of wild-type S. cerevisiae. This indicated that the M. alpina delta9-desaturase had a substrate preference for stearic acid (18:0) rather than palmitic acid (16:0). Genomic clone delta9-2 (ole2) also encoded a protein of 445 aa which had 86% identity to the delta9-1 and LM9 proteins and whose ORF also complemented the yeast ole1 mutation. The transcript from this gene could only be detected in one of the six M. alpina strains tested, suggesting that its expression may be strain-specific or induced under certain physiological conditions. PMID- 10537217 TI - Identification of structural and functional domains of the tetracycline efflux protein TetA(P) from Clostridium perfringens. AB - The Clostridium perfringens tetracycline-resistance protein, TetA(P), is an integral inner-membrane protein that mediates the active efflux of tetracycline from the cell. TetA(P) acts as an antiporter, presumably transporting a divalent cation-tetracycline complex in exchange for a proton, and is predicted to have 12 transmembrane domains (TMDs). Two glutamate residues that are located in predicted TMD 2 were previously shown to be required for the active efflux of tetracycline by TetA(P). To identify additional residues that are required for the structure or function of TetA(P), a random mutagenesis approach was used. Of the 61 tetracycline-susceptible mutants that were obtained in Escherichia coli, 31 different derivatives were shown to contain a single amino acid change that resulted in reduced tetracycline resistance. The stability of the mutant TetA(P) proteins was examined by immunoblotting and 19 of these strains were found to produce a detectable TetA(P) protein. The MIC of these derivatives ranged from 2 to 15 microg tetracycline ml(-1), compared to 30 microg tetracycline ml(-1) for the wild-type. The majority of these mutants clustered into three potential loop regions of the TetA(P) protein, namely the cytoplasmic loops 2-3 and 4-5, and loop 7-8, which is predicted to be located in the periplasm in E. coli. It is concluded that these regions are of functional significance in the TetA(P) mediated efflux of tetracycline from the bacterial cell. PMID- 10537218 TI - Nucleosides as a carbon source in Bacillus subtilis: characterization of the drm pupG operon. AB - In Bacillus subtilis, nucleosides are readily taken up from the growth medium and metabolized. The key enzymes in nucleoside catabolism are nucleoside phosphorylases, phosphopentomutase, and deoxyriboaldolase. The characterization of two closely linked loci, drm and pupG, which encode phosphopentomutase (Drm) and guanosine (inosine) phosphorylase (PupG), respectively, is reported here. When expressed in Escherichia coli mutant backgrounds, drm and pupG confer phosphopentomutase and purine-nucleoside phosphorylase activity. Northern blot and enzyme analyses showed that drm and pupG form a dicistronic operon. Both enzymes are induced when nucleosides are present in the growth medium. Using mutants deficient in nucleoside catabolism, it was demonstrated that the low molecular-mass effectors of this induction most likely were deoxyribose 5 phosphate and ribose 5-phosphate. Both Drm and PupG activity levels were higher when succinate rather than glucose served as the carbon source, indicating that the expression of the operon is subject to catabolite repression. Primer extension analysis identified two transcription initiation signals upstream of drm; both were utilized in induced and non-induced cells. The nucleoside catabolizing system in B. subtilis serves to utilize the base for nucleotide synthesis while the pentose moiety serves as the carbon source. When added alone, inosine barely supports growth of B. subtilis. This slow nucleoside catabolism contrasts with that of E. coli, which grows rapidly on a nucleoside as a carbon source. When inosine was added with succinate or deoxyribose, however, a significant increase in growth was observed in B. subtilis. The findings of this study therefore indicate that the B. subtilis system for nucleoside catabolism differs greatly from the well-studied system in E. coli. PMID- 10537219 TI - Neisseria gonorrhoeae bacterioferritin: structural heterogeneity, involvement in iron storage and protection against oxidative stress. AB - The iron-storage protein bacterioferritin (Bfr) from Neisseria gonorrhoeae strain F62 was identified in cell-free extracts and subsequently purified by column chromatography. Gonococcal Bfr had an estimated molecular mass of 400 kDa by gel filtration; however, analysis by SDS-PAGE revealed that it was composed of 18 kDa (BfrA) and 22 kDa (BfrB) subunits. DNA encoding BfrB was amplified by PCR using degenerate primers derived from the N-terminal amino acid sequence of BfrB and from a C-terminal amino acid sequence of Escherichia coli Bfr. The DNA sequence of bfrA was subsequently obtained by genome walking using single-specific-primer PCR. The two Bfr genes were located in tandem with an intervening gap of 27 bp. A potential Fur-binding sequence (12 of 19 bp identical to the consensus neisserial fur sequence) was located within the 5' flanking region of bfrA in front of a putative -35 hexamer. The homology between the DNA sequences of bfrA and bfrB was 55.7%; the deduced amino acid sequences of BfrA (154 residues) and BfrB (157 residues) showed 39.7% identity, and showed 41.3% and 56.1% identity, respectively, to E. coli Bfr. Expression of recombinant BfrA and BfrB in E. coli strain DH5alpha was detected on Western blots probed with polyclonal anti-E. coli Bfr antiserum. Most Bfrs are homopolymers with identical subunits; however, the evidence presented here suggests that gonococcal Bfr was composed of two similar but not identical subunits, both of which appear to be required for the formation of a functional Bfr. A Bfr-deficient mutant was constructed by inserting the omega fragment into the BfrB gene. The growth of the BfrB-deficient mutant in complex medium was reduced under iron-limited conditions. The BfrB-deficient mutant was also more sensitive to killing by H2O2 and paraquat than the isogenic parent strain. These results demonstrate that gonococcal Bfr plays an important role in iron storage and protection from iron-mediated oxidative stress. PMID- 10537220 TI - Physiological characterization of Streptococcus bovis mutants that can resist 2 deoxyglucose-induced lysis. AB - Streptococcus bovis JB1 does not normally lyse, but stationary phase lysis can be induced by including 2-deoxyglucose (2DG) in the growth medium. Isolates deficient in glucose/2DG phosphotransferase activity (PTS-) also lysed when 2DG was present (Lys+) and this result indicated that 2DG phosphorylation via the PTS was not an obligate requirement for 2DG-induced lysis. Cells and cell walls from 2DG-grown cultures lysed faster when proteinase K was added, but glucose-grown cultures and cell walls were not affected. A lipoteichoic acid (LTA) extract (aqueous phase from hot phenol treatment) from glucose-grown cells inhibited the lysis of 2DG-grown cultures, but a similar extract prepared from 2DG-grown cells was without effect. Thin-layer chromatography and differential staining indicated that wild-type and Lys+ PTS- cells incorporated 2DG into LTA, but lysis-resistant cultures (Lys- PTS+ and Lys- PTS-) did not. LTA from lysis-resistant (Lys- PTS+ and Lys- PTS-) cells grown with glucose and 2DG also prevented 2DG-dependent lysis of the wild-type. LTA could not inhibit degradation of cell walls isolated from 2DG-grown cultures, but LTA inhibited the lysis of Micrococcus lysodeikticus (Micrococcus luteus) cells that were exposed to supernatants from 2DG-grown S. bovis cultures. Group D streptococci (including S. bovis) normally have an alpha 1,2 linked glucose disaccharide (kojibiose) in their LTA, but kojibiose cannot be synthesized from 2DG. This observation suggested that the kojibiose moiety of LTA was involved in autolysin inactivation. Wild-type S. bovis had ATP- as well as PEP-dependent mechanisms of 2DG phosphorylation and one lysis-resistant phenotype (Lys- PTS-) had reduced levels of both activities. However, the Lys- PTS+ phenotype was still able to phosphorylate 2DG via ATP and PEP and this result indicated that some other step of 2DG incorporation into LTA was being inhibited. Based on these results, growth in the presence of 2DG appears to prevent synthesis of normal LTA, which is involved in the regulation of autolytic enzymes. PMID- 10537221 TI - Extracellular metal-binding activity of the sulphate-reducing bacterium Desulfococcus multivorans. AB - Polarography was used to measure the copper-binding ability of culture filtrates from a range of sulphate-reducing bacteria (SRB), including pure cultures and environmental isolates. Of those tested, Desulfococcus multivorans was shown to have the greatest copper-binding capacity and this organism was used for further experiments. Extracellular copper- and zinc-binding activities of Dc. multivorans culture filtrates from batch cultures increased over time and reached a maximum after 10 d growth. The culture filtrate was shown to bind copper reversibly and zinc irreversibly. Twelve-day-old Dc. multivorans culture filtrates were shown to have a copper-binding capacity of 3.64 +/- 0.33 micromol ml(-1) with a stability constant, log10K, of 5.68 +/- 0.64 (n=4). The metal-binding compound was partially purified from culture growth media by dichloromethane extraction followed by HPLC using an acetonitrile gradient. PMID- 10537222 TI - Lovastatin inhibits the production of gibberellins but not sterol or carotenoid biosynthesis in Gibberella fujikuroi. AB - Sterols, carotenoids and gibberellins are synthesized after the reduction of 3 hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) to mevalonate in different subcellular compartments of the fungus Gibberella fujikuroi. Lovastatin inhibits growth in many organisms, presumably because of the inhibition of the synthesis of essential terpenoids. However, in G. fujikuroi growth of the mycelia and sterol and carotenoid content were not affected by the presence of lovastatin. Nevertheless, lovastatin did inhibit the accumulation of gibberellins in the culture medium; this inhibition, however, was counteracted by the addition of mevalonate to the medium. The conversion of HMG-CoA to mevalonate in cell-free extracts was inhibited by 10 nM lovastatin. Since G. fujikuroi apparently possesses a single gene for HMG-CoA reductase, as shown by Southern hybridization and PCR amplification, it was concluded that the biosynthesis of sterols, carotenoids and gibberellins shares a single HMG-CoA reductase, but the respective subcellular compartments are differentially accessible to lovastatin. PMID- 10537223 TI - "Rational hope" in the early treatment of Parkinson's disease. AB - The drug treatment of Parkinson's disease since the original description of the malady in 1817 is described. Consideration is given to the historic use of alkaloids of the belladonna, harmala, and aporphine families, and of amphetamine. The introduction of the L-dopa treatment is described. The modes of action of the various drugs employed in the past as well as those in current use are described in the context of knowledge of the functioning of the nigrostriatal tract. PMID- 10537224 TI - Role of Na-Ca exchange in the action potential changes caused by drive in cardiac myocytes exposed to different Ca2+ loads. AB - The role of Na-Ca exchange in the membrane potential changes caused by repetitive activity ("drive") was studied in guinea pig single ventricular myocytes exposed to different [Ca2+]o. The following results were obtained. (i) In 5.4 mM [Ca2+]o, the action potentials (APs) gradually shortened during drive, and the outward current during a train of depolarizing voltage clamp steps gradually increased. (ii) The APs shortened more and were followed by a decaying voltage tail during drive in the presence of 5 mM caffeine; the outward current became larger and there was an inward tail current on repolarization during a train of depolarizing steps. (iii) These effects outlasted drive so that immediately after a train of APs, currents were already bigger and, after a train of steps, APs were already shorter. (iv) In 0.54 mM [Ca2+]o, the above effects were much smaller. (v) In high [Ca2+]o APs were shorter and outward currents larger than in low [Ca2+]o. (vi) In 10.8 mM [Ca2+]o, both outward and inward currents during long steps were exaggerated by prior drive, even with steps (+80 and +120 mV) at which there was no apparent inward current identifiable as I(Ca). (vii) In 0.54 mM [Ca2+]o, the time-dependent outward current was small and prior drive slightly increased it. (viii) During long steps, caffeine markedly increased outward and inward tail currents, and these effects were greatly decreased by low [Ca2+]o. (ix) After drive in the presence of caffeine, Ni2+ decreased the outward and inward tail currents. It is concluded that in the presence of high [Ca2+]o drive activates outward and inward Na-Ca exchange currents. During drive, the outward current participates in the plateau shortening and the inward tail current in the voltage tail after the action potential. PMID- 10537225 TI - Myocardial meta-[125l]iodobenzylguanidine uptake in awake genetically hypertensive rats at different ages: an autoradiographic study. AB - The purpose of this work was to evaluate changes in myocardial meta [125I]iodobenzylguanidine ([125I]MIBG) uptake and distribution with age in awake spontaneously hypertensive rats (SHR) with respect to Wistar-Kyoto (WKY) rats. Rats were randomly divided into two groups, one for measuring myocardial [125I]MIBG uptake and distribution 4 h after its injection and the second for evaluating myocardial catecholamine concentrations. Mean arterial blood pressure, cardiac hypertrophy index (heart/body weight ratio), and heart rate were significantly higher with increasing age in SHR compared with matched WKY rats. Myocardial catecholamine concentrations and turnover did not differ between the two strains and were significantly decreased with increasing age. Myocardial [125I]MIBG uptake determined by gamma counting was similar in WKY rats and SHR and did not vary significantly with age when expressed as uptake density. However, in both strains of rats, [125I]MIBG uptake determined by autoradiography was significantly greater at the base of the heart than at the apex and midventricular levels, and the uptake values of young rats were significantly higher than those of older rats. In 21-week-old WKY rats and SHR, the highest [125I]MIBG uptake values were found in the right ventricle. Thus, quantitative autoradiography allowed detection of significant changes in myocardial [125I]MIBG uptake and showed its heterogeneous distribution in the rat heart. PMID- 10537226 TI - Heterogeneity among beta-adrenoreceptor blockers in the modulation of energy dependent uptake of the organic cation amantadine by rat renal tubules. AB - Eight representative beta-adrenoreceptor blocking drugs, acebutolol, atenolol, labetalol, metoprolol, nadolol, pindolol, propranolol, and timolol, were studied in vitro with respect to their potential to block energy-dependent uptake of [3H]amantadine into proximal and distal rat renal tubule fragments in the presence and absence of bicarbonate. Five of the eight beta-adrenoreceptor blockers showed a dose-dependent inhibition of renal tubule accumulation of amantadine: labetalol, metoprolol, pindolol, propranolol, and timolol. Labetalol was the only beta-adrenoreceptor blocker with greater inhibitory potency in phosphate-based buffer than in bicarbonate-based buffer. Propranolol was the most potent inhibitor of renal tubule amantadine accumulation with IC50 values of 15 +/- 10 and 31 +/- 11 microM for proximal and distal tubule fragments, respectively, in a bicarbonate-based buffer environment. Inhibition in a phosphate-based buffer was less potent only in proximal tubules, with an IC50 of 76 +/- 30 microM. Kinetic studies of propranolol inhibition of amantadine uptake were consistent with both uncompetitive and competitive inhibition mechanisms in bicarbonate-based buffer in both proximal and distal renal tubule segments. There was no chiral preference between the R and S forms of propranolol for the inhibitory effects observed. These data suggest that there is potential for selection among the beta-adrenoreceptor blocking drugs to minimize or restrict the inhibition of amantadine energy-dependent uptake at the organic cation ion uptake sites characterized by amantadine in the presence and absence of bicarbonate. PMID- 10537227 TI - Postnatal changes in regional blood flow during cold-induced shivering in sow reared piglets. AB - To determine whether newborn pigs are able to display adequate cardiovascular adjustments favouring shivering thermogenesis in skeletal muscles soon after birth, regional blood flow and fractional distribution of cardiac output were determined in 1-day-old (n = 6) and 5-day-old (n = 6) conscious piglets at thermal neutrality and during cold exposure, using coloured microspheres. Five day-old piglets stayed with the sow before the experiment. The cold challenge was designed to induce a similar increase (approximately +90%) in heat production at both ages. Skeletal muscle blood flow increased with both age (p < 0.05) and cold exposure (p < 0.001), with the effect of cold being more pronounced in 5-day-old piglets than in 1-day-old piglets (+60%, p < 0.05). The difference between individual muscles increased with age, with fractional blood flow being 41% higher in rhomboideus than in longissimus thoracis muscle during cold exposure in 5-day-old piglets (p < 0.05). Cardiac output was similar at both ages and increased by 23% in the cold (p < 0.001). At 1 day of age, there was no redistribution of cardiac output among the internal organs during the cold challenge, while at 5 days of age, the increase in muscle fractional blood flow was associated with a reduction (p < 0.05) in the fraction of cardiac output reaching the skin (-24%), the small intestine (-21%), and the liver (-20%). In conclusion, these results suggest that there is a rapid postnatal improvement of cardiovascular adjustments favouring blood perfusion and probably heat production during cold-induced shivering in the most oxidative muscles studied. This cardiovascular response may play a role in the postnatal enhancement of thermoregulation in piglets. PMID- 10537228 TI - Cyclic GMP and cyclic AMP induced changes in control and hypertrophic cardiac myocyte function interact through cyclic GMP affected cyclic-AMP phosphodiesterases. AB - We tested the hypothesis that the negative functional effects of cyclic GMP (cGMP) would be greater after increasing cyclic AMP (cAMP), because of the action of cGMP-affected cAMP phosphodiesterases in cardiac myocytes and that this effect would be altered in left ventricular hypertrophy (LVH) produced by aortic valve plication. Myocyte shortening data were collected using a video edge detector, and O2 consumption was measured by O2 electrodes during stimulation (5 ms, 1 Hz, in 2 mM Ca2+) from control (n = 7) and LVH (n = 7) dog ventricular myocytes. cAMP and cGMP were determined by a competitive binding assay. cAMP was increased by forskolin and milrinone (10(-6) M). cGMP was increased with zaprinast and decreased by 1H-[1,2,4]oxadiazolo[4,3-a]quinoxilin-1-one (ODQ) both at 10(-6) and 10(-4) M, with and without forskolin or forskolin + milrinone. Zaprinast significantly decreased percent shortening in control (9 +/- 1 to 7 +/- 1%) and LVH (10 +/- 1 to 7 +/- 1%) myocytes. It increased cGMP in control (36 +/- 5 to 52 +/- 7 fmol/10(5) myocytes) and from the significantly higher baseline value in LVH (71 +/- 12 to 104 +/- 18 fmol/10(5) myocytes). ODQ increased myocyte function and decreased cGMP levels in control and LVH myocytes. Forskolin + milrinone increased cAMP levels in control (6 +/- 1 to 15 +/- 2 pmol/10(5) myocytes) and LVH (8 +/- 1 to 18 +/- 2 pmol/10(5) myocytes) myocytes, as did forskolin alone. They also significantly increased percent shortening. There were significant negative functional effects of zaprinast after forskolin + milrinone in control (15 +/- 2 to 9 +/- 1%), which were greater than zaprinast alone, and LVH (12 +/- 1 to 9 +/- 1%). This was associated with an increase in cGMP and a reduction in the increased cAMP induced by forskolin or milrinone. ODQ did not further increase function after forskolin or milrinone in control myocytes, despite lowering cGMP. However, it prevented the forskolin and milrinone induced increase in cAMP. In hypertrophy, ODQ lowered cGMP and increased function after forskolin. ODQ did not affect cAMP after forskolin and milrinone in LVH. Thus, the level of cGMP was inversely correlated with myocyte function. When cAMP levels were elevated, cGMP was still inversely correlated with myocyte function. This was, in part, related to alterations in cAMP. The interaction between cGMP and cAMP was altered in LVH myocytes. PMID- 10537229 TI - Pharmacological evidence for beta2-adrenoceptor in right atria from stressed female rats. AB - The purpose of the present study was to demonstrate a physiological response to TA2005, a potent m-adrenoceptor (beta2-AR) selective agonist, in right atria isolated from stressed female rats under the influence of the estrous cycle. We obtained concentration-response curves to the agonist in the presence and in the absence of selective antagonists in right atria isolated from female rats submitted to three daily foot-shock sessions (30 min duration, 120 pulses of 1.0 mA, 1.0 s, applied at random intervals of 5-25 s) and sacrificed at estrus or diestrus. Our results showed that the pD2 values of TA2005 were not influenced by estrous cycle phase or foot-shock stress. However, in right atria from stressed rats sacrificed during diestrus, the concentration-response curve to TA2005 was biphasic, with a response being obtained at concentrations of 0.1 nM, whereas during estrus no response was observed at doses lower than 3 nM. ICI 1118,551, a beta2-AR antagonist, abolished the response to nanomolar concentrations of TA2005 in right atria from stressed rats at diestrus, with no changes in agonist pD2 values in right atria from control rats (7.47 +/- 0.09, p > 0.05) but a 3-fold decrease in pD2 values of TA2005 in right atria from foot shock stressed rats (7.90 +/- 0.07, p < or = 0.05). Concentration-response curves to TA2005 in the presence of ICI118,551 were best fitted by a one-site model equation. The beta1 AR antagonist, CGP20712A, shifted to the right only the second part of the concentration-response curves to the agonist, unmasking the putative beta2-AR mediated response to the agonist in tissues isolated from stressed rats at diestrus. Under this condition, concentration-response curves to the agonist were best fitted by a two-site model equation. pD2 and maximum response of TA2005 interaction with beta1- and putative beta2-adrenoceptor components were calculated. Schild analyses gave a pK(B) value for CGP20712A that was typical for the interaction with beta1-AR in each experimental group. pK(B) values for ICI118,551 could not be obtained in stressed rats sacrificed at diestrus since Schild plot slopes were lower than 1.0. In right atria from control rats, ICI118,551 pK(B) values were similar to reported values for the interaction of the antagonist with beta1-AR. These results confirm that a heterogenous beta-AR population mediating the chronotropic response to catecholamines can be demonstrated in right atria from foot shock stressed female rats sacrificed at diestrus. The stress-induced response seems to be mediated by the beta2-AR subtype. Right atria from rats sacrificed during estrus are protected against stress-induced alterations on the homogeneity of beta-AR population. PMID- 10537230 TI - Ingestion of chilli pepper (Capsicum annuum) reduces salicylate bioavailability after oral asprin administration in the rat. AB - The bioavailabilities of aspirin (acetylsalicylic acid) and of salicylic acid were studied in male Wistar rats after acute and chronic administration of a Capsicum annuum extract, containing 100 mg of capsaicin per gram. With a single administration of 100 mg/kg of the extract, aspirin blood levels remained unchanged, but salicylic acid bioavailability was reduced in 44% compared with control animals. With a single administration of 300 mg/kg of the extract, aspirin blood levels were undetectable while salicylic acid bioavailability was reduced in 59%. Chronic administration once daily for 4 weeks of 100 and 300 mg/kg of the extract resulted in undetectable aspirin blood levels, while salicylic acid bioavailability was reduced in 63 and 76%, respectively, compared with controls. Results show that Capsicum ingestion reduces oral drug bioavailability, likely as a result of the gastrointestinal effects of capsaicin. PMID- 10537231 TI - Augmentation and suppression of action potentials by estradiol in the myometrium of pregnant rat. AB - The purpose of this study was to investigate the actions of estradiol on spontaneous and evoked action potentials in the isolated longitudinal smooth muscle cells of the pregnant rat. Single cells were obtained by enzymatic digestion from pregnant rat longitudinal myometrium. Action potentials and currents were recorded by whole-cell current-clamp and voltage-clamp methods, respectively. The acute effects of 17beta-estradiol on action potentials and inward and outward currents were investigated. The following results were obtained. The average resting membrane potential of single myometrial cells was 54 mV (n = 40). In many cells, an electrical stimulation evoked a membrane depolarization, and action potentials were superimposed on the depolarization. In some cells, spontaneous action potentials were observed. Estradiol (30 microM) slightly depolarized the membrane (ca. 5 mV) and attenuated the generation of action potentials by reducing the frequency and amplitude of the spikes. Afterhyperpolarization was also attenuated by estradiol (30 microM). On the other hand, in 5 of 35 cells, estradiol increased the first spike amplitude and action potential duration, while frequency of the spike generation and afterhyperpolarization were inhibited. In voltage-clamped muscle cells, estradiol inhibited both inward and outward currents. Acute inhibition or augmentation of spike generation by estradiol is due to the balance of inhibition of inward and outward currents. Inhibition of both currents also prevented afterhyperpolarization, causing potential-dependent block of Ca spikes. PMID- 10537232 TI - Effect of saline infusion on kidney and collecting duct function in atrial natriuretic peptide (ANP) gene "knockout" mice. AB - Atrial natriuretic peptide (ANP) is thought to play a role in renal regulation of salt balance by reducing tubular reabsorption of sodium and chloride. Therefore, in the chronic absence of this hormone, a defect of salt excretion should be evident. We used an ANP gene deletion model to test this premise. F2 homozygous mutant mice (-/-) and their wild-type littermates (+/+) were fed an 8% NaCl diet prior to an acute infusion of isotonic saline. Arterial blood pressures, renal excretions of salt and water, as well as collecting duct transport of fluid and electrolytes were measured. Pressures were significantly higher in -/- compared with +/+ mice (139 +/- 4 vs. 101 +/- 2 mmHg; 1 mmHg = 133.3 Pa). There was no difference in glomerular filtration rate (-/- = 0.84 +/- 0.06; +/+ = 0.81 +/- 0.04 mL x min(-1) x g(-1) kidney weight). In the collecting duct, sodium and chloride reabsorptions were significantly higher in the -/- group than in the +/+ group. As a result, natriuresis and chloruresis were relatively reduced (U(Na)V: /- = 8.6 +/- 1.1; +/+ = 14.0 +/- 1.1; U(Cl)V: -/- = 10.1 +/- 1.4; +/+ = 16.0 +/- 1.1 micromol x min(-1) x g(-1) kidney weight). We conclude that the absence of endogenous ANP activity in mice on a high-salt diet subjected to acute saline infusion causes inappropriately high reabsorption of sodium and chloride in the medullary collecting duct, resulting in a relative defect in renal excretory capacity for salt. PMID- 10537233 TI - Proteinase-activated receptor 4 (PAR4): action of PAR4-activating peptides in vascular and gastric tissue and lack of cross-reactivity with PAR1 and PAR2. AB - We studied the actions of the human and murine proteinase-activated receptor 4 (PAR4) derived receptor-activating peptides (APs), GYPGQV-NH2 (GQV-NH2) and GYPGKF-NH2 (GKF-NH2), (i) to activate-desensitize either PAR1 or PAR2 in cultured cell systems (calcium signalling in PAR1/PAR2-bearing human HEK cells and in rat KNRK cells expressing either rat or human PAR2) and (ii) to affect contractility in rat aorta (RA) and rat gastric longitudinal muscle (LM) preparations in vitro. We found that neither PAR1 nor PAR2 was affected by concentrations of the PAR4 APs (800 microM) that caused both an endothelium-dependent nitric oxide mediated relaxation of preconstricted RA tissue and a contractile response in the LM preparation. The potencies (EC50 values 300 to 400 microM) of GQV-NH2 and GKF-NH2 for causing a relaxant effect were identical and comparable with the potency of GQV-NH2 for causing a contractile effect in the LM. However, the potencies of the PAR4-APs in the RA and LM preparations were 20- to 150-fold lower than the potency of the receptor-selective PAR1-AP, TFLLR-NH2. We conclude that the PAR4 APs do not activate either PAR1 or PAR2, and we suggest that along with PAR1 and PAR2, PAR4 may also be present in rat vascular and gastric smooth muscle. PMID- 10537234 TI - Obstacles to healing in medicine and science: the interplay of science, paradigm, and culture. PMID- 10537235 TI - Learning from adverse events of acupuncture. PMID- 10537236 TI - Self-hypnosis and coronary bypass surgery. PMID- 10537237 TI - Safety of Hypericum perforatum. PMID- 10537238 TI - Research methods and CAM. PMID- 10537239 TI - Western frontier physician taking pulse. PMID- 10537240 TI - Addition of acupuncture and self-care education in the treatment of patients with severe angina pectoris may be cost beneficial: an open, prospective study. AB - OBJECTIVES: A cost-benefit analysis of acupuncture and self-care education in the treatment of patients with angina pectoris. DESIGN: An open prospective study on an unselected group of patients. For comparison of risk three control groups were used: (1) published data concerning medical and invasive treatments; (2) an age- and sex matched group obtained from a randomly selected Danish population of 14,000 people; and (3) the 211 patients in this group with angina pectoris symptoms. SETTING: The treatment was carried out on a outpatient basis in a private research clinic. SUBJECTS: 105 patients with angina pectoris, 73 candidates for invasive treatment, and 32 for whom this was rejected. INTERVENTIONS: Acupuncture and self-care education was added to the pharmaceutical treatment. OUTCOME MEASURES: Healthcare expenses, a satisfactory medical status defined as New York Heart Association (NYHA) classification 0-I and/or no use of antianginal medication, and risk measured as cardiac death or myocardial infarction. RESULTS: The estimated cost savings during 5 years were $32,000 (U.S.) per patient, mainly due to a 90% reduction in hospitalization and 70% reduction in needed surgery. Compared to 8% before treatment, 53% of the patients achieved a life without limitations (NYHA 0-I) 1 year after treatment, as did 69% after 5 years. No increased risk for myocardial infarction or cardiac death was observed. CONCLUSIONS: The addition of acupuncture and self-care education was found to be cost beneficial in patients with advanced angina pectoris. The results invite further testing in a randomized controlled trial. PMID- 10537241 TI - Zhong Yi acupuncture and low-back pain: traditional Chinese medical acupuncture differential diagnoses and treatments for chronic lumbar pain. AB - Little attention has been given to selecting treatments in clinical trials of acupuncture. Yet in order to perform objective tests of this procedure, it is crucial that the selected treatments are considered representative of the style of practice being tested. We examined 16 traditional Chinese medicine (TCM) acupuncture texts or treatment articles to determine the consistency of diagnosis and recommended treatment for chronic low-back pain. Although 24 diagnostic patterns were described by 1 or more texts, only 4 patterns were described by at least half of the texts. Most texts (12/16) described only 3 or 4 patterns. These could be categorized into 3 broad types: cold, damp, wind, heat channel obstruction patterns; kidney vacuity patterns (sometimes differentiated into yang and yin patterns); and blood (or blood and qi) stasis patterns. Several acupuncture points were recommended by most texts regardless of the diagnosis, whereas other acupoints were recommended for specific diagnostic patterns. There was, however, substantial variation between texts in recommended acupoints, with less than 20% of all acupoints recommended by half or more of the texts. This varibility will make it difficult to select TCM treatments for clinical trials of chronic low-back pain that have wide applicability. We believe that examining treatment patterns in actual clinical practice is crucial in this situation. We suggest that this method of selecting treatments should be part of the process used when selecting treatments for all clinical trials of acupuncture, regardless of the style of practice. PMID- 10537242 TI - Effects of four herbal extracts on adjuvant-induced inflammation and hyperalgesia in rats. AB - OBJECTIVE: To evaluate the effects of four herbal medicine extracts on a rat model of inflammatory hyperalgesia. DESIGN/INTERVENTIONS: Inflammation was induced by injecting complete Freund's adjuvant (CFA) into one hindpaw of each rat. Four herbs that are routinely prescribed in Traditional Chinese Medicine for treatment of pain were used: Duhuo (Radix Angelicae Pubescentis), Bai jiang cao (Patriniae Herba cum Radice), Yan hu suo (Rhizoma Corydalis) and Sanqui (Panax Notoginseng). The crude water extracts of the herbs were inected intraperitoneally following a repeated treatment profile. OUTCOME MEASURES: Thermal hyperalgesia was assessed by testing each rat's paw withdrawal response to a noxious thermal stimulus. The magnitude of edema was determined by measuring the maximal thickness of the paw with a caliper. The effect of herb extracts on motor performance was assessed by using an accelerating rotarod test. RESULT: Duhuo, Bai jiang cao, and Yan hu suo significantly attenuated CFA-induced hyperalgesia at 2 hours and facilitated the recovery from hyperalgesia (p < 0.05), when compared to saline-treated rats. The CFA-induced edema was reduced by Duhuo at 24 hours, 72 hours and 168 hours; Bai jiang cao at 24 hours, and Yan hu suo at 24 hours and 168 hours. Sanqi did not produce any significant effect on inflammation and hyperalgesia. The rotarod performance was slightly reduced by Bai jiang cao, Yan hu suo, and Sanqi (p < 0.05) but not by Duhuo treatment. CONCLUSION: The present study identified Duhuo as a selective and effective herbal agent in attenuating persistent hindpaw inflammation and hyperalgesia in rats. These results indicate that some herbal agents may provide an alternative approach to the treatment of persistant inflammatory pain and hyperalgesia. PMID- 10537243 TI - The American coneflower: a prophylactic role involving nonspecific immunity. AB - OBJECTIVE: In humans, considerable circumstantial evidence exists that indicates soluble root extracts of the American coneflower, genus Echinacea, may act to ameliorate virus-mediated afflictions, such as the common cold, influenza, and even AIDS and virus-based tumors. This study was designed to quantify, in normal mice, Echinacea-mediated, quantitative, dynamic changes, with time on both mature and precursor cells, of all the hemopoietic and immune-cell lineages in the spleen and bone marrow. DESIGN: A specific, commercially prepared potent extract of Echinacea root was provided daily in the diet for either 1 week or 2 weeks with the aim of establishing a possible mechanism of action for this herb. RESULTS: The data revealed that natural-killer (NK) cells and monocytes, both mediators of nonspecific immunity and well-demonstrated killers of virus containing cells, were numerically and significantly increased in both the bone marrow and the spleen as early as 1 week after beginning treatment with the dietary herb. In contrast to our observations with NK cells and monocytes, the sizes of all other hemopoietic and immune cell populations in these two organs remained at control levels even after 2 weeks of daily dietary Echinacea. CONCLUSIONS: The work has demonstrated the specific nature of Echinacea-derived phytochemicals in acting as stimulants of those cells responsible for nonspecific immunity, as the first line of defense against virus-infected/transformed cells. The observations that these cells were elevated in the bone marrow indicates that at least one mechanism of action of this herb, is to stimulate new cell production in situ. The significant elevation of these two fundamental immune cell populations, in normal animals, suggests a prophylactic role for this herb. PMID- 10537244 TI - Physician-patient communication about complementary and alternative medical therapies: a survey of physicians caring for patients with human immunodeficiency virus infection. AB - OBJECTIVE: To examine frequency and correlates of physicians' reports of discussions with patients with human immunodeficiency virus (HIV) about complementary and alternative medical (CAM) therapies. DESIGN: Mailed physician survey. SETTING: The setting was Eastern Massachusetts. PARTICIPANTS: Participants included 89 physicians caring for patients with HIV. MEASUREMENTS AND MAIN RESULTS: Physicians were asked how common the use of CAM therapies was among their patients, how useful these therapies were, how often they discussed the use of CAM therapies with new and follow-up patients, and whether they had used a CAM therapy themselves in the last year. We also collected information on physicians' sociodemographic and practice characteristics. Sixty-eight percent (89/130) of physicians responded, and 26% and 5% reported discussing CAM therapies with HIV-infected patients at most new and follow-up visits, respectively. Respondents' attitudes toward the use of CAM therapies were generally positive, and they believed their HIV-infected patients used CAM therapies more than their non-HIV infected patients. The majority (63%) believed that CAM therapies may be helpful for HIV-infected patients. Thirty-six percent (36%) had used a CAM therapy themselves in the last year. In multivariate analyses, only the belief that CAM therapies are helpful was correlated with discussion of CAM therapies (p = 0.006). Respondents' demographic characteristics, training, personal use of CAM therapies, reported visit length, and satisfaction with visit length were not associated with discussion of CAM therapies. CONCLUSIONS: Despite awareness that their HIV-infected patients commonly use CAM therapies and positive attitudes towards such therapies, most of these physicians did not routinely discuss CAM therapies with them. Barriers to physician-patient communication about CAM therapies merit further investigation. PMID- 10537245 TI - Guidelines for selecting a medical herbalist for consultation and referral: consulting a medical herbalist. AB - Many Americans are self-medicating with herbal medicines. Health care practitioners often need to consult with, and/or refer to a medical herbalist in order to fully address the herb information needs of their clients. This article provides a guideline for the health practitioner to use when assessing the competency and expertise of an herb practitioner. Three models for assessing mastery in a health-related discipline are used as a foundation for the guideline. PMID- 10537246 TI - Why do patients seek treatment in hospitals of complementary medicine? AB - OBJECTIVE: This exploratory study evaluated patients' reasons for entering a complementary (alternative) medicine hospital by ranking 15 medical and psychosocial factors that were thought to influence this choice. SUBJECTS AND OUTCOME MEASURES: Two hundred patients (200) from two complementary hospitals, one focusing on Traditional Chinese Medicine and one on the Western type of complementary medicine, completed an extensive questionnaire at the beginning of their inpatient treatment. The questionnaire covered personal background; disease parameters; attitude towards conventional medicine; previous experience with, and knowledge of, complementary therapies; expectations concerning the forthcoming treatment; health-related habits; personality traits; and social support. RESULTS: Optimistic attitudes towards treatment and a positive appraisal of alternative doctors were frequently stated reasons (80%), as was the disease severity (long duration: 86%; acute progression or imminent surgery: 70%). Previous successes with complementary therapies, however, ranked relatively low (53%). Negative opinions concerning conventional therapies and conventional doctors' treatments were mentioned by 68% of the patients. Many patients felt themselves to be under considerable psychologic stress (74%). A majority (73%) was well informed about complementary therapies, and 65% were curious about the forthcoming therapies. Sixty-eight percent (68%) indicated good health behaviors. Fewer patients mentioned contemplative and/or religious attitudes (44%) or lack of social support (25%). Age primarily accounted for variations in the ranking weights of the two subgroups. The specific type of complementary medicine was of minor influence. In 14 out of 21 personality dimensions, the current patient group showed significant deviations from the healthy reference, which is in good agreement with findings from conventionally treated patients. PMID- 10537247 TI - Ignorance about homeopathy. AB - Three hundred and three (303) adults selected by a research consultancy were asked six open-ended questions about homeopathy to ascertain their beliefs, knowledge, and behavior. Just over 40% said homeopathy meant to them "natural or herbal medicine"; while approximately one third said they do not know how homeopathy "works." Just under one third believed that homeopathy was available on the British National Health Service, while two thirds agreed that it was quite easy to get homeopathic treatment. Despite its popularity, people remain surprisingly ignorant about homeopathy. PMID- 10537248 TI - Unconventional medicine in Central Europe: a misuse of public health insurance? AB - OBJECTIVE: To scrutinize the presumption maintained by critics that patients seeking medical treatment at a health resort may be more motivated by the prospect of a pleasant sojourn paid for by health insurance than by the impairment caused by a disease. DESIGN: Variables for mobility (occiput-to-wall distance, cervical rotation, chest expansion, thoracic flexion, lumbar flexion, and finger-to-floor distance) and C-reactive protein were determined in 181 patients (male 134, female 47; age 52.4 +/- SEM 0.8 years) with ankylosing spondylitis (AS) whose costs were covered by their health insurance (group A) and in 77 AS patients (male 66, female 11; age 51.6 +/- 1.2 years) who paid their own costs (group B). SUBJECTS: A group of 258 patients with AS presenting for 3- or 4 week speleotherapeutic radon treatment at the Gasteiner Heilstollen Hospital, a medical institution located at Badgastein in the Austrian Alps. RESULTS: After Bonferroni correction for multiple calculations no significant difference was seen between the two groups. CONCLUSIONS: The results suggest that patients presenting for medical treatment at a health resort suffer a like degree of disease impairment, whether they pay their own costs or not. There was no evidence that seeking treatment at a health resort may be an attempt by patients to misuse the health insurance for "sponsored" holidays. PMID- 10537249 TI - Total knee arthroplasty in the presence of severe flexion contracture: a report of 37 cases. AB - From 1987 to 1994, 37 total knee arthroplasties were performed in 23 patients with severe, fixed flexion contractures averaging 78 degrees (range, 60 degrees 100 degrees). Fourteen of the knees had flexion contractures of greater than 90 degrees and 7 were fused at 90 degrees. There were 19 women and 4 men. The average age at surgery was 42 years (range, 20-57 years). The diagnoses were rheumatoid arthritis in 17 patients, juvenile rheumatoid arthritis in 3, and ankylosing spondylitis in 3. Preoperatively, all patients were Knee Society Category C, with 14 being nonambulatory and 9 minimally ambulatory. Follow-up averaged 4.3 years (range, 2-8 years). Postoperatively, patients were immobilized in extension when not in continuous passive motion or physical therapy. Flexion contractures were corrected to an average of 7 degrees postoperatively (range, 0 degrees -15 degrees). Arc of motion improved from 25 degrees preoperatively to 82 degrees postoperatively. The average Knee Society knee scores improved from 25 points preoperatively to 78 points postoperatively, and the functional scores improved from 0 points preoperatively to 71 points postoperatively. Five knees were manipulated under anesthesia postoperatively. Complications included 3 transient peroneal nerve palsies, 1 transient episode of vascular insufficiency, 6 delayed wound healings, and 1 deep infection. There were no aseptic loosenings. We conclude that although technically difficult, total knee arthroplasty can be performed successfully in this challenging and highly debilitated subset of patients, giving them marked improvement in quality of life. PMID- 10537250 TI - Wear rates of ceramic-on-ceramic bearing surfaces in total hip implants: a 12 year follow-up study. AB - A retrospective clinical and radiographic analysis was performed on 58 patients (60 hips; mean age at time of surgery, 45.2 years) at a minimum of 10-year follow up (mean, 12.7 years) after total hip replacement using a ceramic-on-ceramic hearing total hip implant (Autophor, Smith and Nephew, Memphis, TN). Mean wear rate at final follow-up was 0.21 mim, averaging 0.016 mm/y. There were no cases of periprosthetic osteolysis in the acetabuulum or femur. For the unrevised components, there were 3 (5%) cases of protrusio acetabuli and 4 (7%) cases of acetabular component loosening. On the femoral side, 78.3% had distal pedestal formation, and 83% had greater than 2 mm implant-bone radiolucencies in more than 5 Gruen zones as a result of gross motion of the stem. Despite radiographic evidence of implant loosening, this hard bearing articulation functioned well in vivo for more than 12 years with remarkably low wear--approximately one tenth the rate reported for metal-on-polyethylene total hip bearings. PMID- 10537251 TI - Acetabular reconstruction with morcellized allograft and ring support: a medium term review. AB - Acetabular bone stock deficiency is commonly encountered in revision hip surgery. A number of techniques are available to address this problem, including the use of particulate allograft with reconstruction rings in an effort to provide a stable construct and replenish bone stock. Our technique and results using such devices in complex acetabular deficiencies are described. In the setting of a large nmedial segmental or cavitary acetabular defect, morcellized bone-graft is used to reconstitute the acetabular floor. This graft is reverse reamed until its depth allows screw fixation of a metallic support ring. The screws also serve to compress the graft. A polyethylene acetabular component is then cemented into the reconstituted acetabulum with full freedom of orientation. A series of 48 patients in whom this technique was employed is presented. These cases have been clinically and radiologically reviewed with a mean follow-up of 64 months (range, 25-102 months). Good bony incorporation with stable acetabular components was seen in all but the two cases in which sepsis predominated. PMID- 10537252 TI - Indomethacin for 3 days is not effective as prophylaxis for heterotopic ossification after primary total hip arthroplasty. AB - Although indomethacin is effective in preventing heterotopic ossification (HO) after total hip arthroplasty (THA) when used for 8 to 14 days, side effects are frequently observed. We conducted a prospective, nonrandomized pilot study of prophylaxis for HO in THA using indomethacin for 3 days. We used a 2-stage design for phase 2 clinical data, based on earlier studies in our department. Our study group consisted of 19 patients, of whom 14 (74%) developed HO; 7 (37%) showed grade 1, 4 (21%) grade II, and 3 (15.8%) grade III according to the Brooker classification. We compared these results with 2 historic control groups; one group received indomethacin for 7 days, and the other group received no prophylaxis. We did not see any reduction of the ossification relative to the group that received no prophylaxis. Indomethacin for 3 days seems not to be sufficient to prevent HO. PMID- 10537253 TI - The position of the popliteal artery in the arthritic knee. AB - We studied the position of the popliteal artery in 32 patients with primary osteoarthritis of the knee. A total of 45 knees were studied using a noninvasive technique with color-flow duplex scanning. The distance between the popliteal artery and the posterior tibial cortex was measured in various positions of flexion. The distance separating them was found to be maximal between 60 degrees and 90 degrees. The study was repeated in a smaller series of 17 patients (20 knees) after knee replacement but with less conclusive results. We believe the safest position on which to operate in primary arthroplasty is with the knee in flexion, but the safety margins are not the same in revision surgery. PMID- 10537254 TI - Injury to the popliteal artery and its anatomic location in total knee arthroplasty. AB - Injury to the popliteal artery during total knee arthroplasty (TKA) is a devastating complication. Although infrequent, these injuries can result in the need for further surgery, including revascularization or possibly even amputation. Several mechanisms are capable of producing direct trauma to the popliteal artery, including the use of posterior ret ractors. We investigated the proximity of the popliteal artery to the tibial joint surface during TKA to identify crucial steps in the procedure at which the artery was at highest risk for injury. TKA was performed on cadaveric specimens, and serial intraoperative arteriograms were taken throughout the procedure, demonstrating the potential for arterial injury by the instrumentation. Additionally, 50 transverse magnetic resonance imaging scans of unrelated knees were analyzed for the position of the popliteal artery relative to the midline of the tibial plateau as well as at a level 5 to 10 mm below this, at the site of a typical resection during TKA. All of the arteriograms showed the artery to be a lateral structure at the joint line. Additionally a posterior retractor placed the artery at risk when it was placed in a position lateral to the posterior cruciate ligament or when it was injudiciously inserted more than 1 cm into the soft tissues. Hyperextension of the knee, which might occur during preparation of the patella, produced dramatic tenting of the artery over the posterior joint line. These results demonstrate that the popliteal artery is at significant risk during TKA, particularly if posterior retractors are placed in a position lateral to the midline of the joint. Both hyperflexion and especially hyperextension produced severe deformities and kinking of the artery and would particularly jeopardize an artery with atherosclerosis. Our findings suggest that the popliteal artery may be at least risk during TKA if posterior retractors are placed medial to the midline of the tibial plateau and if care is taken to avoid extremes of both flexion and extension. PMID- 10537255 TI - The outcome of trochanteric reattachment in revision total hip arthroplasty with a Cable Grip System: mean 6-year follow-up. AB - We have reviewed 251 hips that were revised by the senior authors with subsequent reattachment using the Dall-Miles Cable Grip System. Of these patients, 223 were available for follow-up. A trochanteric slide osteotomy was used for most cases (n = 170), and the remainder had conventional trochanteric osteotomy to facilitate surgical exposure. Follow-up period was 1 to 8 years. Forty-eight percent (n = 108) of the hips had a previous trochanteric osteotomy. Thirteen percent (n = 30) had a prior trochanteric nonunion. Of the 223 hips, 91% (n = 204) of the trochanters remained attached to the trochanteric bed when reapproximated by the cable grip system. The 2 multifilament cables were passed medially through drill holes in the lesser trochanter in 67% (n = 149) of cases. Of the hips, 16% (n = 35) had 2 cables passed through bone lateral to the prosthesis, and 17% (n = 39) had cables passed 1 medial and 1 lateral to the prosthesis. Cable breakage was noted in 10% (n = 23) of cases. Of those 23, 70% (n = 16) were stainless steel. Unraveling of the cable occurred in 18% (n = 41) of cases. There were 19 nonunions (9%). Of the 19 nonunions, 74% (n = 14) were stainless steel. The trochanter was reattached to bone in 9 hips, to cement in 4 hips, and to a proximal femoral allograft in 6 hips (P = .0001). Eight of the 19 hips (42%) had the cables placed lateral to the prosthesis (P = .0002). When bone to-bone apposition was achieved at surgery, the nonunion rate was 4%. In this difficult group of revision procedures, the Dall-Miles Cable Grip has provided reliable trochanteric fixation. Factors associated with successful trochanteric healing include use of vitallium cables, use of a trochanteric slide osteotomy, cables passed medially through the lesser trochanter, cerclage rather than intramedullary placement, and bone-to-bone apposition. PMID- 10537256 TI - Results and complications of the low contact stress knee prosthesis. AB - We present the midterm results and complications of 101 low contact stress total knee replacements performed in 94 patients and reviewed at an average follow-up of 5.2 years (range, 4-8 years). The mean age at the time of surgery was 66 years (range, 53-76 years). Meniscal bearings were used in 83 knees and a rotating platform in the remaining 18 cases; cemented fixation was used only for 16 tibial components. At most recent follow-up, average knee and function scores were 93 and 78 points. None of the knees was revised because of loosening. Five knees showed distal femoral stress shielding. Complications included infection, supracondylar fracture, patellar component dislodgment, meniscal dislocation (2 cases), catastrophic wear of polyethylene, and progressive osteolysis (2 cases). PMID- 10537257 TI - An in vitro comparison of the motion behavior of different bipolar endoprostheses. AB - The relative motion of 3 different bipolar endoprostheses was evaluated in vitro. A paired fresh acetabulum was frozen at 0 degrees C and defrosted 12 hours before the experiment. Three bipolar endoprostheses were evaluated: UNIQHIP system (United Orthopedics), UHR system (Osteonics), and AML system (Depuy). The surface roughness and spherical roundness of outer shells and inner heads of the bipolar prostheses were measured before the experiments. The acetabulum and outer shell of the bipolar prostheses were fixed on a Bionix 858 material testing system axially by separate fixation tools. The axial load of 1,400 N and 2,800 N was than applied on the specimen. The axis was rotated from 0 degrees to 90 degrees at the speed of 1 degree/s. All 3 outer shells were tested to this paired acetabulum randomly and separately. The frictional torque on the outer bearing surface of the different prostheses was recorded by the material testing system. The frictional torque on the inner bearing surface was also measured by the same procedure as was done for the outer bearing. The final results were statistically compared by the 1-way analysis of variance test method. Bipolar prostheses of the UHR system showed the largest frictional torque on outer bearing when it was loaded with 1,400 N and 2,800 N. The final results showed that all the bipolar prostheses had ideal motion behavior when functioning under the loading of 1,400 N. The frictional torque on the inner bearing was found to be larger than the frictional torque on the outer bearing in some prostheses when the loading was increased to 2,800 N. Thus, the bipolar endoprostheses functioned as unipolar prostheses. The only relative motion remained between the outer bearing surface and the acetabulum. This effect causes complications, such as implant protrusion in the acetabulum. PMID- 10537258 TI - Total hip replacements done without cement after acetabular fractures: a 4- to 8 year follow-up study. AB - Twenty-one patients (21 hips) underwent cementless total hip replacement surgeries for previous acetabular fractures. The mean age at the time of hip replacement was 52 years (range, 23-78 years). The mean follow-up was 65 months (range, 48-104 months). One hip required revision of the stem secondary to a periprosthetic femur fracture from a fall at 3 months after surgery. Good to excellent clinical rating was achieved and maintained in 19 hips. Radiographic evaluation demonstrated stable cup and stem fixation in 17 and 15 hips. Only 1 patient with radiographic loosening of the components was sufficiently symptomatic. The results in this series appeared slightly better than those reported previously in hip replacements done with cement at comparable medium term follow-up. The mechanical failure rates remained high in this patient population: 19% for the cups and 29% for the stems. PMID- 10537259 TI - Posterior distal cement extrusion during primary total hip arthroplasty: a cause for concern? AB - Manually operated injection systems are routinely used to deliver polymethyl methacrylate during cemented femoral component primary total hip arthroplasty (THA). The goal of cement delivery is to achieve sufficient intrusion of cement into the trabecular bone of the prepared femur so that the femoral component is securely bonded to the femur. We have observed posterior distal cement extrusion (PDCE), which appears to be secondary to too-successful pressurization. We sought to quantify and offer a possible explanation for this phenomenon. Eight patients with PDCE were identified, with an estimated incidence range of 0.90%, to 1.6% of primary cemented femoral component THA. All occurred in female patients of small stature. Endosteal canal diameters were also small, averaging 11 mm, 10 cm from the lesser trochanter. The PDCE occurred at an average distance of 9.8 cm from the midpoint of the lesser trochanter, and was most easily visualized on the lateral radiograph where it resided in the posterior soft tissues. Examination of 49 human femora showed 1 or more vascular channels in the posterior aspect of the femur in all specimens. The most proximal vascular channel averaged 10.1 cm distal to the lesser trochanter and had an average lumen diameter of 1 mm. The vascular channel contained an artery and 2 veins by histologic examination. We postulate that PDCE represents the escape of low-viscosity cement out of the vascular channel, and laboratory simulation supports this possibility. Because this finding has not previously been reported, we hoped that other centers will look closely for this phenomenon. PMID- 10537260 TI - Cytokine response of human macrophage-like cells after contact with polyethylene and pure titanium particles. AB - The aim of this study was to establish a human macrophage cell culture system to examine the effect of polyethylene (PE) and titanium particles on cytokine release by macrophage-like cells (MLC) and to quantify this response with respect to the nature and concentration of particles. Human monocytic leukemia cells were differentiated under standard conditions with vitamin D3 and granulocyte macrophage-colony-stimulating factor. Cells were characterized by fluorescence activated cell-sorter Scan of CD 14 expression analysis as well as a phagocytosis test exploiting fluorescence-labeled particles of bacteria] walls. To achieve a relevant contact between the floating PE particles (approximately 1 microm in size) and MLC, a rotation device was used (15 rotations/min) during incubation. The same was done with the titanium particles. Cell culture supernatants were then analyzed for interleukin (IL)-1beta, IL-8, and tumor necrosis factor (TNF) alpha using the enzyme-linked immunosorbent assay technique in the absence or presence of particles. Rotation of incubated MLC alone did not influence the secretion of TNF-alpha, but it enhanced secretion of IL-1beta and IL-8 about 30 fold compared to background levels. Both PE and titanium particles significantly enhanced MLC cytokine release, the amount of which depended on the concentration of particles. Using 40 X 10(8) PE particles (0.7 x 10(8) titanium particles) and 10(6) MLC, the maximal release of IL-1beta was about 20-fold (7-fold titanium particles) higher than that of the rotating control sample. The stimulation of IL 8 release was 4-fold (3-fold titanium particles) and of TNF-alpha. 300-fold (170 fold titanium particles) compared to controls. MLC were viable (>90% cell survival) at concentrations less than 108 x 10(8) polyethylene particles per 10(6) MLC and 16 x 10(8) titanium particles per 10(6) MLC. Rotation per se as well as exposure to increasing concentrations of PE and titanium particles stimulates cytokine release (TNF-alpha, IL-1beta, IL-8) by macrophages in vitro. This in vitro model resembles the in vivo situation near arthroplasties, where implant particles make contact with inflammatory cells, such as macrophages. Cytokine release by macrophages may impair osteoblast function as well as stimulate bone resorption by osteoclasts and macrophages, thereby causing aseptic loosening of arthroplasties. Our in vitro model provides a reproducible human cell system that might shed light on the pathogenesis of particle disease and might serve as a reproducible in vitro test system for the biocompatibility of foreign materials. PMID- 10537261 TI - Femoral bone regeneration subsequent to impaction grafting during hip revision: histologic analysis of a human biopsy specimen. AB - Cemented revision with impaction grafting shows encouraging early clinical results; postoperative biopsy specimens taken from the proximal femur in humans have demonstrated viable trabecular and cortical bone. Human radiographic studies also illustrate density changes within the proximal femur, consistent with remodeling of bone-graft. In an animal experiment, bone incorporation was shown in the proximal femur, but graft lysis was reported around the distal portion of the implant. We report on a patient who sustained a traumatic femoral fracture at the level of the tip of the femoral component 27 months after revision with impaction grafting and a collarless polished taper stem. At the time of open reduction and internal fixation of the fracture, we obtained circumferential biopsy specimens from the fracture site. Three distinct zones could be identified histologically: i) an inner zone consisting of bone-cement, fibrous tissue, and partially necrotic trabeculae with evidence of bone remodeling; ii) a middle zone consisting of viable trabecular bone and probable neocortex formation with fewer particles of bone-cement; and iii) an outer zone with viable cortex. Fibrous tissue was present around some of the incorporating bone-graft fragments, but no continuous fibrous membrane was present. Cement particles were identified, but no polyethylene debris was found by light microscopy. Biopsy specimens from the distal aspect of the prosthesis may not reflect changes seen proximally, but based on the available tissue, this case illustrated histological evidence of bone-graft remodeling after impaction grafting. These results are consistent with our expectations based on radiographic findings and clinical results. PMID- 10537262 TI - In vivo measurement of acetabular cement pressurization using a simple new design of cement pressurizer. AB - Aseptic loosening of the acetabular component remains a limiting factor in the long-term success of total hip replacement. An instrumented pressurizer has been designed to allow the intraoperative measurement of acetabular cement pressurization, which is known to contribute to implant fixation. Average intraoperative cement pressures in 16 operations performed by 2 surgeons were 49 +/- 17 kPa (6.4 +/- 2.3 psi) and 47 +/- 17 kPa (6.2 +/- 2.2 psi), and peak pressures were 76 +/- 5 kPa (10.0 +/- 0.6 psi) and 93 +/- 15.kPa (12.2 +/- 1.9 psi), comparable to previous work in vitro. The pressurization required for optimal cement penetration into cleaned low-density cancellous bone is reported to be of the order of 35 to 50 kPa (4.6-6.6 psi) for 30 to 60 seconds, and the present data show that this is attainable in vivo using a simple device. PMID- 10537263 TI - Ultra-high-molecular weight polyethylene wear: an in vitro comparison of acetabular metal types and polished surfaces. AB - The generation of debris from the wear of ultra-high-molecular weight polyethylene (UHMWPE) is a well-recognized factor in the development of osteolysis and the long-term failure of total joint arthroplasties. Wear between the articulation of the femoral head and the polyethylene has been recognized for many years, but more recently, both retrieval and in vitro studies have demonstrated that convex surface wear or backside wear also occurs and may be of significance. Currently, modular acetabular components are being designed with polished surfaces, fewer screw holes, various polyethylene locking mechanisms, and stiffer metal alloys in an attempt to reduce backside wear. The purpose of this study was to determine if differences existed in UHMWPE wear based on the metal alloy used and the surface finish in modular acetabular components. Sixteen components in 4 groups were subjected to 10 million gait cycles using an in vitro joint simulator. All components used 28-mm cobalt chrome femoral heads on cobalt chrome tapered stems. The 4 groups differed only in the type of metal backing and type of interior finished surface: polished cobalt chrome, unpolished cobalt chrome, polished titanium, and unpolished titanium. UHMWPE changes were examined in terms of articular (concave) surface wear, backside (convex) surface wear, and frictional torque. The overall linear and volumetric wear rates were 1.05 mm/10 million cycles and 325 mm3/10 million cycles. No significant differences in linear and volumetric wear rates were detected between the cobalt chrome and titanium acetabular components. Surface finish did not influence wear rates. In terms of backside wear, all specimens in the 4 groups demonstrated total loss of all sputtered gold with the exception of those areas extruded through the screw holes. Extrusion through the screw holes was on the order of 0.0004 inch for all groups, and no significant difference was seen among the groups for this parameter. The measurements of articular frictional torque demonstrated a significant difference among the polished and unpolished cobalt chrome components (17.3 N x m vs 11.5 N x m; P = .0039, 2-way analysis of variance, Student's Newman Keuls method). Some designs in modular acetabular components have favored stiffer alloys, such as cobalt chrome, with polished concave surfaces to decrease wear on both the concave and the convex surfaces. In this study, there was no significant difference in wear rates noted between cobalt chrome and titanium acetabular components, and polishing of the components had no appreciable affect in reducing backside wear. PMID- 10537264 TI - Discrepancy in the length of the tibiae in unilateral congenital dislocation of the hip. AB - We measured discrepancy in the length of the tibiae in cases of unilateral congenital dislocation of the hip (CDH) in 10 patients. The mean shortening of the tibia on the affected side was 1 cm and was not related to the severity of the dislocation. Patients who are about to undergo total hip arthroplasty for CDH should be warned that the operation is unlikely to restore normal leg length. PMID- 10537265 TI - Incomplete seating of an acetabular metal wire retaining ring during total hip arthroplasty. PMID- 10537266 TI - Uncemented total hip arthroplasty in Paget's disease of the hip: a report of 5 cases with 5-year follow-up. AB - Five patients with Paget's disease localized to the acetabulum received cementless acetabular components during total hip replacement. Three were primary surgeries, and 2 were revisions of a failed cemented acetabular component. At an average of 5.8 years (range, 4.8-8.8 years) after the operation, all acetabular components were well fixed radiographically with no migration or loosening. No patients complained of clinical symptoms referable to the acetabular component. No revisions had been performed. The ability of this inherently abnormal bone to proceed through the reparative and remodeling phases of porous ingrowth adds support to the use of uncemented components for acetabular reconstruction in Paget's disease of the hip. PMID- 10537267 TI - Failure of a metal-on-metal total hip arthroplasty from progressive osteolysis. AB - Ultra-high-molecular weight polyethylene (UHMWPE) wear, debris-induced osteolysis is a frequent cause of failure of total hip arthroplasty. Metal-on-metal total hip arthroplasty eliminates the generation of UHMWPE particulate debris. Although the volumetric wear of a metal-on-metal articulation may be lower than a metal UHMWPE articulation, the number of particles may be higher. Osteolysis can develop in response to metallic and UHMWPE debris. The following case of massive osteolysis associated with large amounts of cobalt-chrome wear debris shows adhesive and abrasive wear mechanisms, as well as wear caused by third-body cobalt-chrome debris and impingement of the femoral component against the rim of the acetabular cup, which led to failure of a metal-on-metal total hip arthroplasty. PMID- 10537268 TI - Closed reduction of dislocated total hip with S-ROM constrained acetabular component. AB - One of the major disadvantages reported for the use of the S-ROM constrained total hip arthroplasty is the need for mandatory urgent revision surgery in cases of dislocation. In patients who are medically compromised and poor surgical candidates, this disadvantage presents a difficult management dilemma. To address this problem, we have developed a technique for closed reduction of dislocated S ROM constrained hips. This technique has been used successfully on 3 patients, all of whom were medically compromised and poor surgical candidates. All patients tolerated the procedure well, and all went on to revision total hip arthroplasty on an elective basis after medical optimization. PMID- 10537269 TI - Unusual case of septic arthritis of the hip: spread from adjacent adductor pyomyositis. AB - Distinguishing intracapsular and extracapsular hip infections may be clinically difficult. Because of this difficulty in diagnosis, the spread of an extracapsular infection into the hip joint may be missed and lead to significant joint destruction. The case of a patient who suffered from the spread of adductor pyomyositis to the hip joint is reported. The delay in diagnosis of an intracapsular hip infection led to significant intra-articular destruction and ultimately necessitated a Girdlestone resection arthroplasty. The patient's hip function was salvaged with a total hip arthroplasty. The presence of an extracapsular hip infection should mandate serial physical examinations and aggressive evaluation to rule out intracapsular spread. A delay in diagnosis of an intracapsular hip infection can lead to catastrophic results. PMID- 10537270 TI - Allograft reconstruction of the extensor mechanism for progressive extensor lag after total knee arthroplasty and previous patellectomy: a 3-year follow-up. AB - Major extensor lag after total knee arthroplasty may follow operative damage to the patellar tendon or its insertion. It may also occur in a late progressive form postoperatively. A successful allograft reconstruction of the extensor mechanism for progressive extensor lag after total knee arthroplasty is described. Patellectomy had been carried out earlier on the same knee for patellofemoral osteoarthritis. PMID- 10537271 TI - An unusual complication affecting an unresurfaced patella after total knee arthroplasty. AB - The unusual clinical presentation of a complication involving an unresurfaced patella after a total knee replacement is reported. The management and possible pathology are discussed. PMID- 10537272 TI - Influence of intramedullary versus extramedullary alignment: guides on final total knee arthroplasty component position by Antonio Maestro et al. (pp 552-558) PMID- 10537273 TI - Gene therapy with dominant-negative Stat3 suppresses growth of the murine melanoma B16 tumor in vivo. AB - Whereas signal transducers and activators of transcription were originally discovered as mediators of normal cytokine signaling, constitutive activation of certain signal transducer and activator of transcription proteins, including Stat3, has been found in increasing numbers of human cancers. Recently, a causal role for Stat3 activation in oncogenesis has been demonstrated, suggesting that Stat3 represents a novel target for cancer therapy. We report here that in vitro expression of a Stat3 variant with dominant-negative properties, Stat3beta, induced cell death in murine B16 melanoma cells that harbored activated Stat3. By contrast, expression of Stat3beta had no effect on normal fibroblasts or the Stat3-negative murine tumor MethA, suggesting that only tumor cells with activated Stat3 have become dependent on this pathway for survival. Significantly, gene therapy by electroinjection of the Stat3beta expression vector into preexisting B16 tumors caused inhibition of tumor growth as well as tumor regression. This Stat3beta-induced antitumor effect is associated with apoptosis of the B16 tumor cells in vivo. These findings demonstrate for the first time that interfering with Stat3 signaling induces potent antitumor activity in vivo and thus identify Stat3 as a potential molecular target for therapy of human cancers harboring activated Stat3. PMID- 10537274 TI - Fusion of the EWS-related gene TAF2N to TEC in extraskeletal myxoid chondrosarcoma. AB - Extraskeletal myxoid chondrosarcomas (EMCs) are characterized by a recurrent t(9;22)(q22;q12) translocation, resulting in the fusion of the EWS gene in 22q12 and the TEC gene in 9q22. Here we report that a third member of the EWS, TLS/FUS gene family, TAF2N, can replace EWS as a fusion partner to TEC in EMC. Two tumors, one with a novel t(9;17)(q22;q11) variant translocation and one with an apparently normal karyotype, expressed TAF2N-TEC fusion transcripts. In both cases, the chimeric transcripts were shown to contain exon 6 of TAF2N fused to the entire coding region of TEC. This transcript is structurally and functionally very similar to the EWS-TEC fusions. The exchange of the EWS NH2-terminal part with the TAF2N NH2-terminal part in EMC further underscores the oncogenic potential of these protein domains as partners in fusion genes. PMID- 10537275 TI - Germ-line msh6 mutations in colorectal cancer families. AB - Hereditary nonpolyposis colorectal carcinoma (HNPCC) is due primarily to inherited mutations in two mismatch repair genes, MSH2 and MLH1, whereas germ line mutations in other mismatch repair genes are rare. We examined the frequency of germ-line msh6 mutations in a population-based series of 140 colorectal cancer patients, including 45 sporadic cases, 91 familial non-HNPCC cases, and 4 HNPCC cases. Among the 91 population-based familial non-HNPCC cases, germ-line msh6 mutations were found in 6 patients (7.1% of probands analyzed; median age at diagnosis, 61 years). These mutations included a splice site mutation, a frameshift mutation, two missense mutations that were demonstrated to be loss of function mutations, and two missense mutations for which functional studies were not possible. In contrast, germ-line msh6 mutations were not found in any of the 45 sporadic cases and the 4 HNPCC cases in the population-based series or in the second series of 58 clinic-based, primarily HNPCC families. Our data suggest that germ-line msh6 mutations predispose individuals to primarily late-onset, familial colorectal carcinomas that do not fulfill classic criteria for HNPCC. PMID- 10537276 TI - MDM2 and MDMX inhibit the transcriptional activity of ectopically expressed SMAD proteins. AB - Transforming growth factor-beta (TGF-beta) inhibits cell proliferation in many cell types, and acquisition of TGF-beta resistance has been linked to tumorigenesis. One class of proteins that plays a key role in the TGF-beta signal transduction pathway is the SMAD protein family. MDM2, a key negative regulator of p53, has recently been shown to suppress TGF-beta-induced growth arrest in a p53-independent manner. Here we show that MDM2 and the structurally related protein MDMX can inhibit the transcriptional activity of ectopically expressed SMAD1, SMAD2, SMAD3, and SMAD4. Immunofluorescence staining indicated that ectopically expressed SMAD4 was present in both the cytoplasm and nucleus, and MDM2 and NIDMX were localized mainly to the nucleus and cytoplasm, respectively. When SMAD4 was coexpressed with either MDM2 or MDMX, nuclear accumulation of SMAD4 was strikingly inhibited. We have no evidence that SMAD4 binds directly to MDM2 or MDMX; hence, the inactivation and nuclear exclusion of SMAD4 by MDM2/MDMX may involve other indirect mechanisms. PMID- 10537277 TI - A novel human xenograft model of inflammatory breast cancer. AB - The step of intravasation or lymphovascular invasion can be a rate-limiting step in the metastatic process. Inflammatory breast carcinoma manifests an exaggerated degree of lymphovascular invasion in situ; hence, a study of its molecular basis might shed light on the general mechanism of lymphovascular invasion exhibited by all metastasizing cancers. To this end, we have established the first human transplantable inflammatory breast carcinoma xenograft (MARY-X) in scid/nude mice. Whereas all other human xenografts grew as isolated s.c. nodules, MARY-X grew exclusively within murine lymphatics and blood vessels, and these latter elements and their supporting stroma comprised, by murine Cot-1 DNA analysis, 30% of the tumor. MARY-X, like its human counterpart, exhibited striking erythema of the overlying skin. MARY-X was estrogen receptor, progesterone receptor, Her 2/neu negative and p53, epidermal growth factor receptor positive. The primary tumor of origin of MARY-X exhibited identical markers, except that about 50% of its cells exhibited Her-2/neu amplification. Comparative studies of MARY-X with noninflammatory xenografts indicated 10-20-fold overexpression of E-cadherin and MUC1, findings that were reflected in actual cases of human inflammatory breast cancer. MARY-X should allow us to further dissect out both the upstream regulatory machinery and the downstream effector molecules responsible for the inflammatory carcinoma phenotype. PMID- 10537278 TI - Inhibition of homologue of Slimb (HOS) function sensitizes human melanoma cells for apoptosis. AB - Homologue of Slimb (HOS)/beta-transducin repeats containing proteins up-regulate nuclear factor kappaB activity by targeting its inhibitor (IkappaB) for ubiquitination and subsequent degradation. We investigated whether inhibition of HOS function may modulate apoptosis in human melanoma cells. Forced expression of the dominant negative HOSdeltaF construct inhibited IkappaB degradation and led to sensitization of melanoma cells to apoptosis induced by tumor necrosis factor alpha with cycloheximide, as well as by cisplatin and ionizing and UV irradiation. These data indicate that HOS plays an important role in controlling the IkappaB-dependent apoptotic pathways in human melanoma. PMID- 10537279 TI - Compensatory lung growth after partial pneumonectomy enhances lung tumorigenesis induced by 3-methylcholanthrene. AB - In small mammals, partial pneumonectomy (PNX) elicits rapid hyperplastic compensatory growth of the remaining lung parenchyma to restore normal lung mass, structure, and function. In BALB mice subjected to PNX, compensatory lung growth is complete within 10 days. Because cellular hyperplasia contributes to the mechanism of tumor promotion by butylated hydroxytoluene (BHT), we hypothesized that hyperplastic compensatory lung growth would promote tumor formation in carcinogen-treated animals in a manner similar to that observed after BHT. In mice subjected to PNX, within 1 week of treatment with the carcinogen 3 methylcholanthrene (MCA; 10 microg/g body weight), lung tumor multiplicity was 3 7-fold higher in animals subjected to PNX than in mice subjected to a sham operation. The increase in tumor multiplicity occurred when PNX was performed 1, 3, and 6 days before or 1 day after MCA treatment. In the absence of PNX, lung tumor multiplicity in MCA-treated mice given one injection of BHT (200 mg/kg body weight) increased significantly (P < 0.01) as compared to that in mice given MCA alone. Tumor multiplicity continued to increase linearly (R2 = 0.99) with each subsequent BHT injection. Lung tumor multiplicity and tumor size in mice given one or two injections of BHT were comparable to those in animals subjected to PNX. These data demonstrate that post-PNX compensatory lung growth stimulates tumorigenesis in MCA-treated mice and provides a novel model for investigating tumor formation. PMID- 10537280 TI - Role of the prostaglandin E receptor subtype EP1 in colon carcinogenesis. AB - Although the cyclooxygenase pathway of the arachidonic acid cascade has been suggested to play an important role in colon carcinogenesis, the molecular species of prostanoids and receptors involved have not been fully elucidated yet. We examined the development of aberrant crypt foci (ACFs), putative preneoplastic lesions of the colon, in two lines of knockout mice, each deficient in prostaglandin E receptors, EP1 and EP3, by treatment with the colon carcinogen, azoxymethane. Formation of ACFs was decreased only in the EP1-knockout mice to approximately 60% of the level in wild-type mice. Administration of 250, 500, or 1000 ppm of a novel selective EP1 antagonist, ONO-8711, in the diet to azoxymethane-treated C57BL/6J mice also resulted in a dose-dependent reduction of ACF formation. Moreover, when Min mice, having a nonsense mutation in the adenomatous polyposis coli gene, were given 500 ppm ONO-8711 in the diet, the number of intestinal polyps was significantly reduced to 57% of that in the basal diet group. These results strongly suggest that prostaglandin E2 contributes to colon carcinogenesis to some extent through its action at the EP1 receptor. Thus, EP1 antagonists may be good candidates as chemopreventive agents for colon cancer. PMID- 10537281 TI - Cell cycle regulation of menin expression. AB - The multiple endocrine neoplasia type 1 gene product, menin, interacts with Jun D. The physiological role of menin in cell cycle control and the manner in which its inactivation contributes to tumorigenesis remain unknown. In the present study, the expression of menin was examined at various cell cycle stages in GH4C1 cells, a rat pituitary cell line. Cells synchronized at the G1-S-phase boundary expressed menin at a lower level than G0-G1-synchronized cells. The expression of menin increased as the cells entered S phase, at which time Jun D expression also increased. In contrast, cells synchronized at the G2-M phase expressed lower levels of menin. At G0-G1, G1-S, and G2-M phases of the cell cycle, menin was found predominantly in the nucleus. In summary, we show that in pituitary cells, menin is a nuclear protein whose expression is cell-cycle regulated. The data suggest that menin has an important role in cell growth regulation. PMID- 10537282 TI - Antitumor cytotoxicity mediated by ligand-activated human V alpha24 NKT cells. AB - Human V alpha24 NKT cells bearing an invariant V alpha24J alphaQ antigen receptor, the counterpart of the murine V alpha14 NKT cells, are activated by the specific ligand, alpha-galactosylceramide (alpha-GalCer) in a CD1d-dependent manner. Here, we demonstrate that the alpha-GalCer-activated V alpha24 NKT cells exert a potent perforin-dependent cytotoxic activity against a wide variety of human tumor cell lines. In addition, we demonstrate that V alpha24 NKT cells and dendritic cells (DCs) from melanoma patients are functionally normal, even in the tumor-bearing status. The potential use of alpha-GalCer-activated V alpha24 NKT cells and/or DCs from patients for cancer immunotherapy is discussed. PMID- 10537283 TI - Comparative studies of a retrovirus versus a poxvirus vector in whole tumor-cell vaccines. AB - A number of experimental and clinical studies have used retroviral vectors to express transgenes in whole tumor-cell vaccines. Recently, poxvirus vectors such as vaccinia or avipox have been used toward this goal. The studies reported here compare for the first time the use of a retroviral vector versus a poxvirus vector (vaccinia) in whole tumor-cell vaccines. The transgene used was the T-cell costimulatory molecule B7-1, and the tumor was the weakly or nonimmunogenic MC38 murine colon adenocarcinoma. Recombinant retrovirus (R-B7) and the recombinant vaccinia (V-B7) induced equivalent expression of B7 on the surface of the carcinoma cell. Using live whole-tumor cells as vaccine, cells transduced via recombinant retrovirus (MC38/R-B7) and recombinant vaccinia (MC38/V-B7) equally induced protection against challenge by native MC38 cells 14 days later. Upon rechallenge with native MC38 cells 40 days later, however, the MC38/R-B7 vaccine was shown to be less effective than the MC38/V-B7 vaccine. Similar results were obtained when the tumor cells were irradiated prior to administration. When comparative studies were conducted in which X-irradiated tumor-cell vaccines were given to mice bearing experimental lung metastases, the MC38/V-B7 vaccine was shown to be significantly (P = 0.0351) more effective than the MC38/R-B7 vaccine. Additional studies were carried out in mice that had received vaccinia virus previously. Again, the X-irradiated MC38/V-B7 vaccine was statistically (P = 0.024) more effective than the MC38/R-B7 vaccine in the elimination of metastases. When the naive and vaccinia-immune mice for each vaccination group were combined for meta-analysis (n = 16), the MC38/V-B7 was significantly more effective than the MC38/R-B7 in the treatment of pulmonary metastases (P = 0.0014) in this model. These studies thus demonstrate for the first time that a whole tumor-cell vaccine (either live or X-irradiated) containing a vaccinia transgene is at least as efficient, and sometimes more efficient, in inducing antitumor effects compared with the same vaccine using a retrovirus to express the transgene. The implications for the clinical applications of such approaches are discussed. PMID- 10537284 TI - p57KIP2 expression and loss of heterozygosity during immortal conversion of cultured human mammary epithelial cells. AB - We have uncovered a novel role for the cyclin-dependent kinase inhibitor, p57KIP2, during the immortalization of cultured human mammary epithelial cells (HMECs). HMECs immortalized after chemical carcinogen exposure initially expressed little or no telomerase activity, and their telomeres continued to shorten with passage. Cell populations whose mean terminal restriction fragment (TRF) length declined to < or = 3 kb exhibited slow heterogeneous growth and contained many nonproliferative cells. These conditionally immortal HMEC cultures accumulated large quantities of p57 protein. With continued passage, the conditionally immortal cell populations very gradually converted to a fully immortal phenotype of good uniform growth, expression of high levels of telomerase activity, and stabilization of telomere length. The fully immortal HMECs that grew well did not accumulate p57 in G0 or during the cell cycle. DNA and RNA analysis of mass populations and individual subclones of conditionally immortal HMEC line 184A1 showed that continued growth of conditionally immortal cells with critically short telomeres was repeatedly accompanied by loss of the expressed p57 allele and transient expression of the allele imprinted previously. Conditionally immortal 184A1 with mean TRF > 3 kb, infected with retroviruses containing the p57 gene, exhibited premature slow heterogeneous growth. Conversely, exogenous expression of human telomerase reverse transcriptase (hTERT), the catalytic subunit of telomerase, in 184A1 with mean TRF > 3 kb prevented both the slow heterogeneous growth phase and accumulation of p57 in cycling populations. These data indicate that in HMECs that have overcome replicative senescence, p57 may provide an additional barrier against indefinite proliferation. Overcoming p57-mediated growth inhibition in these cells may be crucial for acquisition of the unlimited growth potential thought to be critical for malignant progression. PMID- 10537285 TI - Classification of small cell lung cancer and pulmonary carcinoid by gene expression profiles. AB - Small cell lung cancer is a common type of lung cancer that is generally classified within the spectrum of neuroendocrine lung neoplasms. Using high density cDNA arrays, we profiled gene expression of small cell lung cancers and compared these expression profiles to those of normal bronchial epithelial cells and pulmonary carcinoids, which are classified as benign neuroendocrine tumors. We found the overall expression profiles of two small cell lung cancer cell lines, two microdissected tissue samples of primary small cell lung cancer, and cultured bronchial epithelial cells to be relatively similar to one another, with an average Pearson correlation coefficient for these comparisons of 0.63. However, we found the expression profiles of small cell lung cancers (and bronchial epithelial cells) to be surprisingly dissimilar to those of two samples of pulmonary carcinoid tumors, with an average correlation coefficient for these comparisons of 0.20. We then compared the pulmonary carcinoid expression profiles to those of two samples of infiltrating astrocytic brain cancers (oligodendroglioma and high-grade astrocytoma) and found similarity of gene expression among these four samples (average correlation coefficient, 0.57). These gene expression profiles suggest that small cell lung cancers are closely related to (and possibly derived from) epithelial cells, and that pulmonary carcinoids are related to neural crest-derived brain tumors. More generally, our results suggest that broad profiles of gene expression may reveal similarities and differences between tumors that are not apparent by traditional morphological criteria. PMID- 10537287 TI - Angiogenic activity of human soluble intercellular adhesion molecule-1. AB - Serum levels of soluble intercellular adhesion molecule-1 (sICAM-1) are elevated in a number of pathological conditions associated with angiogenesis, including tumor growth. Because the increased levels of sICAM-1 suggested that it may be angiogenic, we tested the ability of sICAM-1 to promote angiogenesis. Human recombinant sICAM-1 stimulates chemokinetic endothelial cell migration, endothelial cell tube formation on Matrigel, and sprouting of aortic rings. sICAM 1 also mediates angiogenesis in the chick chorioallantoic membrane assay. Additionally, we found a Mr 49,000 molecule that binds to sICAM-1 that may be the surface ligand on endothelial cells. The evidence that sICAM-1 has angiogenic activity suggests a possible role linking inflammation and neovascularization. Furthermore, sICAM-1 may enhance tumor growth by promoting angiogenesis and escape from immunosurveillance. PMID- 10537286 TI - Arginine vasopressin promoter regulation is mediated by a neuron-restrictive silencer element in small cell lung cancer. AB - Arginine vasopressin (AVP) is often expressed in small cell lung cancer (SCLC), and a 65-bp AVP minimal promoter fragment is sufficient to restrict activity to SCLC in vitro. We now describe a motif with homology to the neuron-restrictive silencer element (NRSE) within this fragment. Electrophoretic mobility shift analysis demonstrated that multiple specific complexes are bound by this motif. These complexes are cross-competed with a characterized SCG10 NRSE probe and do not bind to the AVP probe with a specific mutation in the NRSE. The complexes vary in mobility between lung tumor cell lines, showing different levels of AVP expression, and some are differentially bound in SCLC. Overexpression of a neuron restrictive silencer factor expression construct can silence reporter gene expression supported by the AVP promoter in SCLC, although this was dependent on both the level of endogenous AVP expression in the cells and putative enhancer elements in larger promoter constructs. Activation of the proximal AVP promoter in SCLC is therefore proposed to, at least partially, rely on modulation of normal repressor activity at the NRSE. PMID- 10537288 TI - Role of mitogen-activated protein kinase/extracellular signal-regulated kinase cascade in gonadotropin-releasing hormone-induced growth inhibition of a human ovarian cancer cell line. AB - Although gonadotropin-releasing hormone agonists (GnRHa) have been used in the therapy of the endocrine-dependent cancers, their biological mechanism remained obscure. We have studied the roles of mitogen-activated protein kinase family in the antiproliferative effect of GnRHa on the Caov-3 human ovarian cancer cell line. Reverse transcription-PCR assays confirmed mRNA for GnRH receptor in Caov-3 cells. In the presence of 1 microM GnRHa, the proliferation of cells was significantly reduced to 76% of controls after 24 h, and the effect was sustained up to 4 days. Although GnRHa had no effect on the activation of the Jun N terminal kinase (JNK), treatment of Caov-3 cells with GnRHa activated extracellular signal-regulated protein kinase (ERK), and its effect was more than that induced by GnRH. Activation of ERK by GnRHa occurred within 5 min, with the maximum occurring at 3 h and sustained until 24 h. GnRHa also activated ERK kinase (mitogen-activated protein/ERK kinase) and resulted in an increase in phosphorylation of son of sevenless (Sos), and Shc. Furthermore, we examined the mechanism by which GnRHa induced ERK activation. Both pertussis toxin (10 ng/ml), which inactivates Gi/Go proteins, and expression of a peptide derived from the carboxyl terminus of the beta-adrenergic receptor kinase I, which specifically blocks signaling mediated by the betagamma subunits of G proteins, blocked the GnRHa-induced ERK activation. Phorbol 12-myristate 13-acetate (PMA) also induced the ERK activity, but pretreatment of the cultured cells with PMA to down regulate protein kinase C did not abolish the activation of ERK by GnRHa. Elimination of extracellular Ca2+ by EGTA also did not abolish the activation of ERK by GnRHa. To examine the role of ERK cascade in the antiproliferative effect of GnRHa, PD98059, an inhibitor of mitogen-activated protein/ERK kinase, was used. This inhibitor canceled the antiproliferative effect of GnRHa and apparently reversed the GnRH-induced dephosphorylation of the retinoblastoma protein, the hyperphosphorylation of which is a hallmark of G1-S transition in the cell cycle. These results provide evidence that GnRHa stimulation of ERK activity may be mediated by Gbetagamma protein, not by PMA-sensitive protein kinase C nor extracellular Ca2+ in the Caov-3 human ovarian cancer cell line, suggesting that this cascade may play an important role in the antiproliferative effect of GnRHa. PMID- 10537289 TI - Polyethylene-glycol suppresses colon cancer and causes dose-dependent regression of azoxymethane-induced aberrant crypt foci in rats. AB - Dietary polyethylene-glycol (PEG) 8000, a nonfermented polymer laxative, strongly suppresses azoxymethane-induced aberrant crypt foci (ACF) in the colon of rats, as shown in a previous study (D. E. Corpet et al., Carcinogenesis (Lond.), 20: 915-918, 1999). In the present study, we tested the effect of PEG administered during either initiation or postinitiation, the dose-response effect of PEG, the regressive effect of PEG on established ACF, and the preventive effect of PEG on colon cancers in rats. The general design was to initiate carcinogenesis in F344 rats by a single injection of azoxymethane (20 mg/kg) and to randomize the animals 7 days later to AIN-76 diets containing 5% PEG or no PEG (control). At termination, ACF and tumors were scored blindly by a single observer. The administration of 5% PEG for 32 days to groups of 10 female rats in either food or drinking water reduced the number of ACF by a factor of 8 (P = 0.0002) and reduced the number of large ACF by a factor of 20-30 (P = 0.002). No protection was afforded when PEG was given only during the initiation phase. Diets containing 0%, 0.5%, 2%, or 5% PEG fed for 35 days to four groups of male rats inhibited ACF in a dose-dependent manner (P < 0.0001). The administration of a 5% PEG diet for 41 days, starting 42 days after carcinogen injection, led to a 73% decrease in the number of ACF (P < 0.0001). Dietary PEG thus caused the regression of established ACF. Macroscopic tumors were evaluated by histology in rats that had been fed a high-fat diet containing cooked casein to promote tumor growth for 81 days. In this accelerated model of carcinogenesis, dietary PEG suppressed the occurrence of colon adenomas and carcinomas: the incidence of tumors decreased from 70% to 10% (P = 0.005); and the multiplicity decreased from 2.1 to 0.1 tumor(s)/rat (P = 0.003). No cancer was detected in the PEG-fed rats. Taken together, these results suggest that PEG could be a potent anticancer agent in the postinitiation phase of carcinogenesis. Because PEG is a substance that is generally recognized as safe (GRAS list, Food and Drug Administration), its cancer-preventive features could be tested in humans. PMID- 10537290 TI - Pancolonic chromosomal instability precedes dysplasia and cancer in ulcerative colitis. AB - Patients with long-standing ulcerative colitis (UC) are at increased risk for colon cancer. These cancers are thought to arise from preexisting dysplasia in a field of abnormal cells that often exhibits aneuploidy and p53 abnormalities. Using dual color fluorescence in situ hybridization with centromere probes and locus-specific arm probes for chromosomes 8, 11, 17, and 18, we demonstrate that chromosomal instability (CIN) is present throughout the colon of UC patients with high-grade dysplasia or cancer. In rectal biopsies that were negative for dysplasia, abnormalities in chromosomal arms, especially losses, were most common, whereas centromere gains were most common in dysplasia and cancer. The frequency and type of abnormalities varied between the chromosomes examined; chromosome 8 was the least affected, and 17p loss was found to be an early and frequent event. Chromosomal arm instability showed 100% sensitivity and specificity for distinguishing control biopsies from histologically negative rectal biopsies from these UC patients, raising the possibility that a screen for CIN might detect the subset of UC patients who are at greatest risk for development of dysplasia and cancer. These results suggest that dysplasia and cancer in UC arise from a process of CIN that affects the entire colon; this may provide the mutator phenotype that predisposes to loss of tumor suppressor genes and evolution of cancer. PMID- 10537291 TI - N-oxidative metabolism of 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) in humans: excretion of the N2-glucuronide conjugate of 2-hydroxyamino-MeIQx in urine. AB - 2-Amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), a major heterocyclic aromatic amine (HAA) formed in cooked meats, is metabolically transformed to mutagenic/carcinogenic intermediates. Cytochrome P4501A2 (CYP1A2)-mediated N hydroxylation followed by phase II O-esterification by N-acetyltransferase (NAT2) are generally regarded as activation processes in which MeIQx and other HAAs are converted to genotoxic species. In this study, we determined the relationship between the activities of these two enzymes and the urinary excretion level of the N2-glucuronide conjugate of 2-hydroxyamino-MeIQx--N2-(beta-1-glucosiduronyl) 2-hydroxyam ino-3,8-dimethylimidazo[4,5-f]quinoxaline (N-OH-MeIQx-N2-glucuronide) -among healthy subjects fed a uniform diet containing high-temperature cooked meat. The individuals (n = 66) in the study ate meat containing known amounts of MeIQx, and urine was collected from 0 to 12 h after the meal. After addition of the deuterium-labeled internal standard to urine, N-OH-MeIQx-N2-glucuronide was isolated using solid-phase extraction and immunoaffinity separation. The isolated conjugate was converted to the deaminated product 2-hydroxy-3,8 dimethylimidazo[4,5-f]quinoxaline (2-OH-MeIQx) by heating with acetic acid. 2-OH MeIQx and its deuterated analogue were derivatized to form the corresponding 3,5 bis(trifluoromethyl)benzyl ether derivatives and analyzed by capillary gas chromatography-negative ion chemical ionization mass spectrometry using selected ion monitoring procedures. The subjects in the study excreted an average of 9.4 +/- 3.0% (+/-SD) of an ingested dose of MeIQx as N-OH-MeIQx-N2-glucuronide in urine; the range varied from 2.2 to 17.1%. A significant correlation was found between the level of N-OH-MeIQx-N2-glucuronide in urine and the amount of MeIQx ingested (r(s) = 0.44; P = 0.0002). The excretion level of N-OH-MeIQx-N2 glucuronide in urine was not associated with the enzyme activities of NAT2 or CYP1A2. This is expected with the latter enzyme because the metabolism of MeIQx is first order and very rapid at the amounts ingested. The amount of N-OH-MeIQx N2-glucuronide in urine was not correlated with the age or sex of the individuals. Our results indicate that biotransformation of MeIQx via CYP1A2 oxidation to form the N-hydroxylamine followed by N2-glucuronidation is a general pathway of MeIQx metabolism in humans; the variability in the excreted levels of N-OH-MeIQx-N2-glucuronide is probably due to interindividual differences in UDP glucuronosyltransferase activity and/or excretion pathways. PMID- 10537292 TI - Induction of immunity to prostate cancer antigens: results of a clinical trial of vaccination with irradiated autologous prostate tumor cells engineered to secrete granulocyte-macrophage colony-stimulating factor using ex vivo gene transfer. AB - Vaccination with irradiated granulocyte-macrophage colony-stimulating factor (GM CSF)-secreting gene-transduced cancer vaccines induces tumoricidal immune responses. In a Phase I human gene therapy trial, eight immunocompetent prostate cancer (PCA) patients were treated with autologous, GM-CSF-secreting, irradiated tumor vaccines prepared from ex vivo retroviral transduction of surgically harvested cells. Expansion of primary cultures of autologous vaccine cells was successful to meet trial specifications in 8 of 11 cases (73%); the yields of the primary culture cell limited the number of courses of vaccination. Side effects were pruritus, erythema, and swelling at vaccination sites. Vaccine site biopsies manifested infiltrates of dendritic cells and macrophages among prostate tumor vaccine cells. Vaccination activated new T-cell and B-cell immune responses against PCA antigens. T-cell responses, evaluated by assessing delayed-type hypersensitivity (DTH) reactions against untransduced autologous tumor cells, were evident in two of eight patients before vaccination and in seven of eight patients after treatment. Reactive DTH site biopsies manifested infiltrates of effector cells consisting of CD45RO+ T-cells, and degranulating eosinophils consistent with activation of both Th1 and Th2 T-cell responses. A distinctive eosinophilic vasculitis was evident near autologous tumor cells at vaccine sites, and at DTH sites. B-cell responses were also induced. Sera from three of eight vaccinated men contained new antibodies recognizing polypeptides of 26, 31, and 150 kDa in protein extracts from prostate cells. The 150-kDa polypeptide was expressed by LNCaP and PC-3 PCA cells, as well as by normal prostate epithelial cells, but not by prostate stromal cells. No antibodies against prostate-specific antigen were detected. These data suggest that both T-cell and B-cell immune responses to human PCA can be generated by treatment with irradiated, GM-CSF gene transduced PCA vaccines. PMID- 10537293 TI - Sensitive detection of K-ras mutations augments diagnosis of colorectal cancer metastases in the liver. AB - Postoperative survival of colorectal cancer patients is often delineated by metastases spreading to the liver. Current clinical diagnostic procedures are unable to discover micrometastases in this organ. Our aim was to develop a diagnostic tool for detecting micrometastases that are present at the time of surgery. Therefore, a PCR-RFLP assay was set up tracking point mutations of the K ras oncogene at codons 12 and 13, based on mismatch primers and restriction enzymes BstXI and XcmI. The detection limit of this assay was one mutant in one million wild-type cells. One hundred forty-two patients with colorectal carcinoma were screened for these mutations in tissue samples from their tumor, proximally adjacent mucosa, and liver. Of these, 67 patients (46%) were positive for a K-ras mutation, of which 58 had codon 12 and 9 had codon 13 mutations. No patient without a K-ras-positive tumor showed a mutation in mucosa, but 11 patients with a K-ras-positive tumor (11 of 58; 19%) were found to bear a K-ras mutation in their mucosa, and in 21 patients (21 of 64; 33%), a K-ras mutation was detected in liver tissue. Sequencing of all mutated samples revealed a 92% confirmation of PCR-RFLP results. In summary, the assay is a useful tool for detecting K-ras codon 12 and 13 mutations and allows early proof of molecular determinants of liver metastases. Such knowledge will improve the staging of colorectal cancer patients and could beneficially influence their prognosis if followed by an effective therapy. PMID- 10537294 TI - Effects of the antiestrogen EM-800 (SCH 57050) and cyclophosphamide alone and in combination on growth of human ZR-75-1 breast cancer xenografts in nude mice. AB - Human breast cancer proliferates as heterogeneous cell populations that exhibit different sensitivities to therapeutic agents. A logical approach to control these different cancer cell populations is the use of combined treatment with agents that block cell proliferation or induce apoptosis via different mechanisms. We therefore investigated the effect of treatment with the novel pure antiestrogen EM-800, alone or in combination with chemotherapy, on the growth of ZR-75-1 human breast tumors in nude mice, a well-recognized model of human breast cancer. Mice bearing estrone-releasing silastic implants as estrogenic stimulus received EM-800 or cyclophosphamide alone or in combination for 227 days. Cyclophosphamide (256 mg/kg/2 weeks) was administered by i.p. injection in 64 mg/kg fractions over 4 consecutive days with repetition of the cycle every 14 days. EM-800 was administered p.o. once daily at the maximally effective dose of 300 microg/mouse. After 227 days of treatment, average tumor size in mice receiving estrone alone was 192% higher than pretreatment. The average tumor size of mice treated with chemotherapy was reduced by 47%, whereas on the other hand, EM-800 caused a 81% decrease of the value of the same parameter. The combined treatment (EM-800 + cyclophosphamide), on the other hand, resulted in a 95% decrease in tumor size compared with control estrogen alone. In fact, EM-800 alone decreased tumor size to 55% of the value at the start of treatment, whereas the addition of cyclophosphamide to the antiestrogen further decreased tumor size to as low as 15% of the pretreatment value. The combination of EM-800 and cyclophosphamide resulted in 95% of complete or partial responses compared with 61 and 27% with EM-800 and cyclophosphamide alone, respectively. In fact, in the combination therapy group, only one tumor remained stable, while 17 regressed >50% and four disappeared. It is noteworthy that no tumor progressed with EM-800 alone or in combination with cyclophosphamide. The present data show, for the first time, that the addition of cyclophosphamide to a pure antiestrogen used at a maximal dose causes a more potent inhibition of human breast tumor growth, thus suggesting that combined treatment using a maximal dose of a pure antiestrogen and a chemotherapeutic agent(s), two classes of compounds having different mechanisms of action, could further improve breast cancer therapy above the results achieved with a potent and pure antiestrogen alone in estrogen-sensitive breast cancer. PMID- 10537295 TI - Risk factors for Ki-ras protooncogene mutation in sporadic colorectal adenomas. AB - The Ki-ras protooncogene frequently is mutated in colorectal adenocarcinomas, but the etiology of this molecular event is uncertain. We investigated the association between variables known or suspected to be related to risk for colorectal cancer and the occurrence of Ki-ras mutations in colorectal adenomas. This study was conducted among 678 male and female participants, 40-80 years of age, enrolled in a phase III trial testing the effects of a wheat bran fiber supplement on adenoma recurrence. Exposure information on the risk factors of interest was assessed through self-administered questionnaires. Mutations in codons 12 and 13 of the Ki-ras protooncogene were analyzed in baseline adenomas 0.5 cm or larger by PCR amplification followed by direct sequencing. Eighteen percent (120 of 678) of the participants had one or more adenoma(s) with Ki-ras mutations. A higher risk of Ki-ras mutations was associated with increasing age and a lower intake of total folate. The odds ratio (OR) for Ki-ras mutations for individuals >72 years of age was 1.98 [95% confidence interval (CI) = 1.19-3.27; P for trend = 0.008] compared with those less than 65 years of age. Compared with individuals in the lower tertile of total folate, those in the upper tertile had an approximately 50% lower risk of having Ki-ras mutation-positive adenomas (OR = 0.52; 95% CI = 0.30-0.88; P for trend = 0.02). There was a suggestion of a stronger inverse association of total folate with G-->T transversions (OR = 0.41; 95% CI = 0.20-0.87) than G-->A transitions (OR = 0.61; 95% CI = 0.31-1.21), although the CIs for the associations overlap. The results of these analyses suggest that the protective effect of folate in colon cancer observed in published studies may be mediated through folate's effect on Ki-ras mutations. PMID- 10537296 TI - Imaging adenoviral-mediated herpes virus thymidine kinase gene transfer and expression in vivo. AB - The feasibility of noninvasive imaging of adenoviral-mediated herpes virus type one thymidine kinase (HSV1-tk) gene transfer and expression was assessed in a well-studied animal model of metastatic colon carcinoma of the liver. Tumors were produced in syngeneic BALB/c mice by intrahepatic injection of colon carcinoma cells (MCA-26). Seven days later, three different doses (3 x 10(8), 1 x 10(8), and 3 x 10(7) plaque-forming units (pfu) of the recombinant adenoviral vector ADV. Rous sarcoma virus (RSV)-tk bearing the HSV1-tk gene were administered by intratumoral injection in separate groups of mice. Two control groups of tumor bearing mice received intratumoral injections of the control adenoviral vector dl 312 or buffer alone, respectively. T2-weighted magnetic resonance (MR) images of mice were obtained before administering the virus and provided an anatomical reference of hepatic tumor localization. Eighteen h after the virus injection, one group of animals was given i.v. injections of 300 microCi of no-carrier-added 5-[131I]-2'-fluoro-1-beta-D-arabinofuranosyluracil (FIAU) and imaged 24 h later with a gamma camera. In some animals, the tumors were sampled and processed for histology and quantitative autoradiography (QAR). The gamma camera images demonstrated highly specific localization of [131I]FIAU-derived radioactivity to the area of ADV.RSV-tk-injected tumors in the liver, which was confirmed by coregistering the gamma camera and T2-weighted MR images. There was no accumulation of [131I]FIAU-derived radioactivity in tumors that were injected with the control vector or injection solution alone. A more precise distribution of radioactivity in the area of transfected tumor was obtained by histological and QAR comparisons. A heterogeneous pattern of radioactivity distribution in transfected tumors was observed. A punctate pattern of radioactivity distribution was observed in peritumoral liver tissue in animals given injections of 3 x 10(8) and 1 x 10(8) pfu of ADV.RSV-tk but not in animals given injections of 3 x 10(7) pfu nor in control animals. A QAR-microscopic comparison showed that the punctate areas of radioactivity colocalized with cholangial ducts. The level of [131I]FIAU derived radioactivity accumulation (HSV1-tk expression) in the transfected tumors was viral dose-dependent. The viral dose-dependency of radioactivity accumulation was more pronounced in peritumoral liver, which was confirmed by reverse transcription-PCR analysis. A separate group of tumor-bearing animals received different doses of ADV.RSV-tk vector followed by treatment with ganciclovir (GCV), 10 mg/kg i.p. b.i.d. for 6 days. The ADV.RSV-tk transfected tumors significantly regressed with GCV treatment; the control tumors continued to grow. During the GCV treatment, the levels of liver transaminases (ALT and AST) were significantly increased in animals that received injections of 3 x 10(8) and 1 x 10(8) pfu of ADV.RSV-tk but not in animals that received injections of 3 x 10(7) pfu and in control animals. The observed liver toxicity confirms the results of gamma camera and QAR imaging, which demonstrated an unwanted spread of ADV.RSV-tk vector and HSV1-tk expression in peritumoral and remote liver tissue at higher doses. These and our previous results indicate that noninvasive imaging of adenoviral-mediated HSV1-tk gene expression is feasible for monitoring cancer gene therapy in patients. PMID- 10537297 TI - Reversal of radiation resistance in LNCaP cells by targeting apoptosis through ceramide synthase. AB - Cell lines derived from human prostate cancer are regarded as relatively resistant to both radiation-induced clonogenic death and apoptosis. Here we attempted to modulate the response of LNCaP prostate cancer cells to radiation therapy (XRT) by pretreatment with 12-O-tetradecanoylphorbol acetate (TPA), a known apoptogenic agent in LNCaP cells. Using plateau-phase cultures, we investigated the response of these cells to XRT, TPA, and a combination of XRT and TPA. LNCaP irradiation did not result in ceramide generation or apoptosis. However, pretreatment with TPA enabled XRT to generate ceramide via activation of the enzyme ceramide synthase and signal apoptosis. Apoptosis was abrogated by the competitive inhibitor of ceramide synthase, fumonisin B1. Furthermore, when transplanted orthotopically into the prostate of nude mice, LNCaP cells produced tumors that recapitulated the responses of LNCaP cells in vitro. XRT or TPA failed to signal apoptosis in LNCaP tumors, whereas a combination of the two resulted in substantial (20-25%) apoptosis within 24 h. There was an additional benefit associated with this regimen because TPA pretreatment protected the adjacent rectum from radiation-induced apoptosis. This represents the first description of signaling-based therapy designed to overcome one form of radiation resistance expressed preferentially in LNCaP human prostate cancer cells. PMID- 10537298 TI - Eradication of primary murine fibrosarcomas and induction of systemic immunity by adenovirus-mediated interferon beta gene therapy. AB - We determined whether an adenoviral vector-mediated murine IFN-beta gene therapy could eradicate established s.c. tumors produced by murine UV-2237m fibrosarcoma cells. The tumor cells were highly susceptible to infection by adenoviral vectors. Cells infected with 10 or 100 multiplicity of infection of AdCIFN-beta, an adenoviral vector encoding murine IFN-beta driven by the human cytomegalovirus promoter, expressed high levels of steady-state IFN-beta mRNA and produced 500 or 7,000 units of IFN-beta activity/10(6) cells/24 h, respectively. Infection of tumor cells with 30 multiplicity of infection of AdCIFN-beta (but not control AdCLacZ vector) inhibited in vitro tumor cell proliferation by 40-45%. Intralesional injection of 5 x 10(8) plaque-forming units of AdCIFN-beta (but not AdLacZ) eradicated established s.c. fibrosarcomas in syngeneic mice but not fibrosarcomas in nude mice. Mice cured of the disease developed systemic immunity against rechallenge with UV-2237m cells but not against another syngeneic tumor, the K-1735 M2 melanoma. Immunohistochemical analysis revealed that tumors injected with AdCIFN-beta contained more macrophages and CD4+ and CD8+ cells than did tumors injected with AdCLacZ or saline. Most cells in the PBS- and AdCLacZ treated tumors stained positive for proliferating cell nuclear antigen, and few cells stained for terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling. In sharp contrast, AdCIFN-beta-treated tumors contained few proliferating cell nuclear antigen-positive cells and many terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling-positive cells. Taken together, our data demonstrate that IFN-beta gene therapy delivered by adenoviral vectors can be effective against fibrosarcomas. PMID- 10537300 TI - The ribonucleoside diphosphate reductase inhibitor (E)-2'-deoxy (fluoromethylene)cytidine as a cytotoxic radiosensitizer in vitro. AB - (E)-2'-Deoxy-(fluoromethylene)cytidine (FMdC) is known as an inhibitor of ribonucleoside diphosphate reductase, a key enzyme in the de novo pathway of DNA synthesis. FMdC was tested as a modifier of radiation response in vitro on a human colon carcinoma cell line (WiDr), and the observed radiosensitization was confirmed on two human cervix cancer cell lines (C33-A and SiHa). Using the clonogenic assay, the effect ratio (ER) at a clinically relevant dose level of 2 Gy was 2.10 (50 nM FMdC), 1.70 (30 nM FMdC), and 1.71 (40 nM FMdC) for the three cell lines WiDr, C33-A, and SiHa, respectively. A more detailed analysis of the importance of timing and concentration of FMdC was done on the WiDr cell line alone, yielding an increased ER(2Gy) with increasing concentration and duration of exposure to the drug, ranging from 1.0 (6 h) to 1.8 (72 h) at 30 nM FMdC and from 1.2 (6 h) to 3.5 (24 h) at 300 nM. We investigated the effect of FMdC on the cellular deoxynucleotide triphosphate pool in WiDr cells and demonstrated a marked depletion of dATP and a significant rise of TTP levels. Cell cycle analysis showed early S-phase accumulation induced by FMdC alone, G2-M block induced by irradiation alone, and an increased accumulation of cells in G2-M if both modalities are used. Our data suggest that FMdC is a radiation response modifier in vitro on different cancer cell lines. The observed radiosensitization may in part be explained by alteration of the deoxynucleotide triphosphate pool, which is consistent with the effect of FMdC on ribonucleoside diphosphate reductase. PMID- 10537299 TI - Antivascular endothelial growth factor receptor (fetal liver kinase 1) monoclonal antibody inhibits tumor angiogenesis and growth of several mouse and human tumors. AB - Tumor angiogenesis is mediated by tumor-secreted angiogenic growth factors that interact with their surface receptors expressed on endothelial cells. Vascular endothelial growth factor (VEGF) and its receptor [fetal liver kinase 1 (Flk 1)/kinase insert domain-containing receptor] play an important role in vascular permeability and tumor angiogenesis. Previously, we reported on the development of anti-Flk-1 and antikinase insert domain-containing receptor monoclonal antibodies (mAbs) that potently inhibit VEGF binding and receptor signaling. Here, we report the effect of anti-Flk-1 mAb (DC101) on angiogenesis and tumor growth. Angiogenesis in vivo was examined using a growth factor supplemented (basic fibroblast growth factor + VEGF) Matrigel plug and an alginate encapsulated tumor cell (Lewis lung) assay in C57BL/6 mice. Systemic administration of DC101 every 3 days markedly reduced neovascularization of Matrigel plugs and tumor-containing alginate beads in a dose-dependent fashion. Histological analysis of Matrigel plugs showed reduced numbers of endothelial cells and vessel structures. Several mouse tumors and human tumor xenografts in athymic mice were used to examine the effect of anti-Flk-1 mAb treatment on tumor angiogenesis and growth. Anti-Flk-1 mAb treatment significantly suppressed the growth of primary murine Lewis lung, 4T1 mammary, and B16 melanoma tumors and growth of Lewis lung metastases. DC101 also completely inhibited the growth of established epidermoid, glioblastoma, pancreatic, and renal human tumor xenografts. Histological examination of anti-Flk-1 mAb-treated tumors showed evidence of decreased microvessel density, tumor cell apoptosis, decreased tumor cell proliferation, and extensive tumor necrosis. These findings support the conclusion that anti-Flk-1 mAb treatment inhibits tumor growth by suppression of tumor-induced neovascularization and demonstrate the potential for therapeutic application of anti-VEGF receptor antibody in the treatment of angiogenesis dependent tumors. PMID- 10537301 TI - Shock wave permeabilization with ribosome inactivating proteins: a new approach to tumor therapy. AB - Extracorporeal shock waves are high-pressure pulses of microsecond duration clinically used for lithotripsy. Recently, shock waves been shown to cause a transient increase of the permeability of the cell membrane. We therefore hypothesized that shock waves might be able to transfer tumoricidal agents into tumor cells and examined this in vitro and in vivo. In vitro, the ribosome inactivating proteins gelonin and saporin were transferred into L1210, SSK2, and HeLa cells, and dose-response curves were established. The drug concentration that reduced the cell proliferation by 50% (IC50) was assessed by 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and the enhancement factors from shock wave application were calculated. It was found that shock waves enhanced the action of gelonin from 900-fold in L1210 cells to 40,000-fold in HeLa cells and the action of saporin from 300-fold in L1210 cells to 15,000 fold in HeLa cells. In vivo, the effect of gelonin and saporin was assessed in a murine tumor model. SSK2 fibrosarcoma tumors locally grown in C3H mice were treated with shock waves after i.p. administration of gelonin or saporin. Shock wave application delayed the tumor growth, and long-term remissions lasting >180 days were induced in 40% of the animals. In conclusion, shock waves enhanced the action of ribosome inactivating proteins and led to complete tumor remissions. The local transfer of toxic substances by shock waves into tumors constitutes a new approach to a local tumor therapy. PMID- 10537302 TI - Enzyme prodrug gene therapy: synergistic use of the herpes simplex virus-cellular thymidine kinase/ganciclovir system and thymidylate synthase inhibitors for the treatment of colon cancer. AB - The goal of this study was to improve the therapeutic index of the herpes simplex virus-thymidine kinase/ganciclovir (HSV-tk/GCV) system by the addition of thymidylate synthase (TS) inhibitors. For this, we assessed the potential of GCV to synergistically interact with 5-fluorouracil (5-FU), ZD1694 (Tomudex), and (E) 5-(2-bromovinyl)-2'-deoxyuridine in HSV-tk-expressing murine MC38 STK and human HT-29 STK colon carcinoma cell lines. Synergistic cell killing was observed in a clonogenic assay over most of the cytotoxic dose range by the median-effect principle of Chou and Talalay (T. C. Chou and P. Talalay, Adv. Enzyme Regul., 22: 27-55, 1984). In a s.c. HT-29 STK xenograft tumor model, we demonstrated that the combination of GCV and 5-FU resulted in statistically significant enhanced animal survival over single-agent treatment. Furthermore, we showed that the combination of GCV and ZD1694 in association with the HSV-tk/GCV system was at least as effective as GCV/5-FU in vitro and in vivo. The mechanism for the observed synergy is most likely attributable to the increased GCV phosphorylation in the presence of the tested TS inhibitors. Our data suggest that the HSV-tk/GCV metabolic suicide gene transfer system could serve as an adjuvant of the presently used TS inhibitors, thus potentially improving the efficacy of present cancer gene therapy approaches. PMID- 10537303 TI - Radiosensitization of human tumor cell lines induced by the adenovirus-mediated expression of an anti-Ras single-chain antibody fragment. AB - The expression of activated ras genes has been implicated as a contributing factor to the radioresistance of tumor cells. As a strategy for compromising Ras protein activity and potentially enhancing the radiosensitivity of tumor cells, we have investigated the application of the AV1Y28 adenovirus, which expresses a single-chain antibody fragment directed against p21 Ras proteins. The ability of AV1Y28 transduction to modulate radioresponse was investigated using four human tumor cell lines--U251 glioblastoma, MIA PaCa-2 pancreatic carcinoma, and the colon carcinomas SW620 and HT29. Cultures were exposed to sufficient levels of AV1Y28 to transduce more than 90% of the cells; 24 h later, cultures were exposed to ionizing radiation, and clonogenic cell survival was determined. Tumor cell survival was reduced by 40-50% when the tumor cell lines were exposed to AV1Y28 only. In addition, for each tumor cell line, AV1Y28 exposure enhanced the level of radiation-induced cell killing. Dose enhancement factors at a surviving fraction of 0.1 ranged from 1.3 to 1.5. Furthermore, for each of the cell lines, the surviving fraction at 2 Gy was significantly reduced by AV1Y28 exposure. In contrast to the results seen in tumor cells, the radiosensitivity of a normal human fibroblast cell line was not affected by AV1Y28. These data indicate that this anti-Ras adenovirus enhances the radiosensitivity of tumor cells but does not affect the radiosensitivity of normal cells. PMID- 10537304 TI - Infiltration of tumors by systemically transferred tumor-reactive T lymphocytes is required for antitumor efficacy. AB - The systemic transfer of ex vivo-activated tumor-sensitized T lymphocytes can mediate immunologically specific regression of established tumors. However, it has not been conclusively established whether the infiltration of systemically transferred T cells into metastases is required for their effector function. In this study, T cells from lymph nodes draining the murine fibrosarcoma MCA 205 cells were activated ex vivo with anti-CD3 monoclonal antibody and interleukin-2. During the final 24 h of culture, the T cells were treated with pertussis toxin (PTX) to inhibit signaling through G protein-coupled chemokine receptors required for diapedesis. Systemically transferred PTX-treated cells did not have any therapeutic efficacy against 3-day established pulmonary metastases. This lack of efficacy correlated with their failure to infiltrate the tumor parenchyma. However, PTX-treated cells responded to tumor antigen stimulation with IFN-gamma secretion in vitro. More importantly, PTX-treated effector T cells prevented tumor growth when they were admixed with tumor cells and inoculated s.c. These results demonstrate that systemically transferred tumor-reactive T lymphocytes need to infiltrate the tumor parenchyma through the endothelium to initiate tumor regression, but PTX-sensitive proteins are not required for either antigen recognition or effector functions. PMID- 10537305 TI - Successful immunotherapy of an intraocular tumor in mice. AB - Immune privilege in the eye is considered essential in the protection against local sight-threatening inflammatory responses. However, the deviant immune responses in the eye may also provide an ideal opportunity to uncontrolled growth of viruses or tumors by inhibiting intraocular immunological attack. To establish to what extent immune privilege interferes with T cell-mediated antitumor immunotherapy, we established a new ocular tumor model in the mouse and tested whether well-defined tumor-specific CTLs can eradicate an immunogenic intraocularly growing tumor. Tumor cells, transformed by human adenovirus type 5 early region 1 (Ad5E1), injected s.c. in a dose of 10(7) cells, did not induce s.c. tumor growth in C57BL/6 mice. However, an injection of 0.3 x 10(6) of these cells into the anterior chamber of the eye led to intraocular tumor growth in 95% of mice (n = 20). Tumor growth in the eye did not induce systemic tumor-specific tolerance, because 70% of the mice were able to eradicate the tumor spontaneously after 5 weeks. Mice vaccinated s.c. with irradiated tumor cells were protected against intraocular tumor challenge, indicating that preactivated memory T cells are able to protect against intraocular tumor growth. Moreover, an i.v. injection of an Ad5E1-specific CTL clone was able to eradicate established intraocular Ad5E1-transformed tumors, whereas the anatomy of the eye remained intact. These results demonstrate that tumor-specific, CTL-mediated immunity can be used successfully for the prevention and eradication of tumors growing in the immune privileged anterior chamber of the eye, without detectable destruction of the eye. PMID- 10537306 TI - A chimeric cell adhesion molecule mediates homing of lymphocytes to vascularized tumors. AB - To facilitate tumor colonization by adoptively transferred cells of the immune system, we created a chimeric cell adhesion molecule that mediates tumor-specific homing by binding to the integrin alpha(v)beta3 on angiogenic endothelial cells. A high-affinity cell adhesion molecule for integrin alpha(v)beta3 was generated by fusing the disintegrin kistrin to the transmembrane adhesion molecule CD31/PECAM-1. This chimeric cell adhesion molecule, termed KISS31, mediates adhesion of lymphoid cells to soluble recombinant integrin alpha(v)beta3 and to endotheliomal monolayers in vitro. KISS31-expressing lymphoid cells accumulate in angiogenic tumors in two in vivo models, in B16/129 melanoma xenografts on the chick chorioallantois and in s.c. growing Lewis lung carcinoma in mice. Our data indicate that expression of KISS31 on lymphoid cells confers tumor-specific homing. This is, to our knowledge, the first example of an experimental mechanism that targets living cells to tumors by redirecting their homing pattern. PMID- 10537307 TI - Regression of established B16F10 melanoma with a recombinant Listeria monocytogenes vaccine. AB - We have previously shown that Listeria monocytogenes, a gram-positive, facultative intracellular bacterium, is a potent vector for targeting tumor specific antigens to the immune system. In the present study, we extend these studies to the highly tumorigenic mouse melanoma B16F10, transduced with a model tumor antigen. We are able to induce the regression of primary tumors and established lung metastases by parenteral immunization with a L. monocytogenes recombinant that expresses the same antigen. Adjunctive therapy with granulocyte macrophage colony-stimulating factor or a vaccinia-based vaccine does not result in an improved cure rate over the L. monocytogenes vaccine alone. Tumor regression is accompanied by the expression of inflammatory cytokines in the tumor. PMID- 10537308 TI - Abnormal Fhit expression in malignant and premalignant lesions of the cervix. AB - Genetic analysis of cervical cancer has demonstrated frequent allelic loss in the 3p chromosomal region. The newly described gene FHIT is located at chromosome region 3p14.2, and its expression has been demonstrated previously by reverse transcription-PCR to be abnormal in a majority of cervical cancer cell lines. In this study, 98 different lesions of the cervix were examined for Fhit expression by immunohistochemical staining. Whereas normal cervical epithelium demonstrated diffuse, moderate to intense cytoplasmic staining, many pathological lesions of the cervix displayed reduced or absent Fhit expression. Sixty-one percent of squamous carcinomas and 40% of adenocarcinomas of the cervix had abnormal Fhit expression. Sixty-five preneoplastic lesions of the cervix were examined. Eleven of 33 high-grade squamous intraepithelial lesions and 1 of 12 low-grade squamous intraepithelial lesions had abnormal Fhit expression. In summary, Fhit expression is frequently abnormal in both glandular and squamous cervical cancers, with a higher frequency of Fhit alterations observed in squamous lesions. In addition, abnormal Fhit expression can be detected in some preneoplastic lesions of the ectocervix. Alterations in Fhit expression may be an important marker of early progression in the development of cancers of the cervix. PMID- 10537309 TI - Structure-function studies of the BTB/POZ transcriptional repression domain from the promyelocytic leukemia zinc finger oncoprotein. AB - The evolutionarily conserved BTB/POZ domain from the promyelocytic leukemia zinc finger (PLZF) oncoprotein mediates transcriptional repression through the recruitment of corepressor proteins containing histone deacetylases in acute promyelocytic leukemia. We have determined the 2.0 A crystal structure of the BTB/POZ domain from PLZF (PLZF-BTB/POZ), and have carried out biochemical analysis of PLZF-BTB/POZ harboring site-directed mutations to probe structure function relationships. The structure reveals a novel alpha/beta homodimeric fold in which dimer interactions occur along two surfaces of the protein subunits. The conservation of BTB/POZ domain residues at the core of the protomers and at the dimer interface implies an analogous fold and dimerization mode for BTB/POZ domains from otherwise functionally unrelated proteins. Unexpectedly, the BTB/POZ domain forms dimer-dimer interactions in the crystals, suggesting a mode for higher-order protein oligomerization for BTB/POZ-mediated transcriptional repression. Biochemical characterization of PLZF-BTB/POZ harboring mutations in conserved residues involved in protein dimerization reveals that the integrity of the dimer interface is exquisitely sensitive to mutation and that dimer formation is required for wild-type levels of transcriptional repression. Interestingly, similar mutational analysis of residues within a pronounced protein cleft along the dimer interface, which had been implicated previously for interaction with corepressors, has negligible effects on dimerization or transcriptional repression. Together, these studies form a structure-function framework for understanding BTB/POZ-mediated oligomerization and transcriptional repression properties. PMID- 10537310 TI - The relationship of DNA ploidy to chromosomal instability in primary human colorectal cancers. AB - The aim of this investigation was to corroborate the relationship between DNA ploidy and chromosomal variation in surgically removed colorectal cancers. For 101 specimens from 21 advanced colorectal cancers, the numerical variations in chromosomes 7, 17, and 18 among cells were measured by fluorescence in situ hybridization using DNA probes specific for centromere of each chromosome, and DNA ploidy was determined by laser scanning cytometry or flow cytometry. DNA diploidy (DNA index = 1.0) was linked with minor variation in copy number of chromosomes 7, 17 and 18, whereas DNA aneuploidy (DNA index > or = 1.2) was found exclusively in tumors with large variations in centromere copy number for all chromosomes. There was a significant difference in the degree of intercellular variations in chromosome copy number between diploid and aneuploid clones for all chromosomes examined (P < 0.001). In near-diploid clones (1.0 < DNA index < 1.2), the numerical variation of chromosome 18 was significantly different from that in diploid clones (P < 0.002), but it was not different from that in aneuploid clones. These observations support the hypothesis that chromosomal instability is associated with DNA aneuploidy in colorectal cancers. Additionally, they suggest that near-diploid tumors are also unstable at a lower level than classic aneuploid tumors and that all chromosomes are not affected equally in near diploid cases. PMID- 10537311 TI - Regulation by p38 mitogen-activated protein kinase of adenylate- and uridylate rich element-mediated urokinase-type plasminogen activator (uPA) messenger RNA stability and uPA-dependent in vitro cell invasion. AB - MDA-MB-231 cells are highly metastatic breast tumor cells. Their high invasiveness is thought to be due to constitutively high levels of urokinase-type plasminogen activator (uPA) and its receptor. Previously (R. Nanbu et al., C. Eur. J. Biochem., 247: 169-174, 1997), we showed that uPA mRNA in these cells is stable and that mRNA degradation mediated by an AU-rich element (ARE) is impaired. Here we report that treatment of MDA-MB-231 cells with SB203580, an inhibitor of the stress-activated p38 mitogen-activated protein (MAP) kinase, strongly destabilized uPA mRNA in an ARE-dependent manner. In contrast, in LLC PK1 and HeLa cells, uPA mRNA is unstable, and an ARE present in the 3' untranslated region plays a role in its degradation. Enhanced ARE-mediated mRNA destabilization induced by SB203580 was also observed in both LLC-PK1 and HeLa cells with a globin chimeric mRNA harboring two copies of the ARE (globin-2ARE) from uPA mRNA. Overexpression of constitutively active MKK6, a p38 upstream activator kinase, increased the stability of the globin-2ARE message in LLC-PK1 cells, confirming the participation of p38 in the regulation of ARE-mediated mRNA decay. Interestingly, the half-life of the uPA mRNA in the three cell lines studied correlated with the basal levels of active p38. SB203580 treatment of MDA MB-231 cells decreased cell-associated uPA activity and dramatically reduced in vitro cell invasiveness. These results suggest the participation of p38 in the control of invasiveness through regulation of the stability of uPA and uPA receptor mRNA, which is also destabilized by p38. PMID- 10537312 TI - Ataxia telangiectasia mutated deficiency affects astrocyte growth but not radiosensitivity. AB - The cancer-prone neurodegenerative disorder, ataxia telangiectasia (A-T), results from mutations of ATM (ataxia telangiectasia mutated). Individuals with A-T are also hypersensitive to ionizing radiation (IR). Cultured cells from A-T individuals or Atm-/- mice have cell cycle and growth defects and are generally considered radiosensitive. However, it has been shown recently that cell populations in the Atm-/- central nervous system are radioresistant. To define specific IR sensitivities of neural populations, we analyzed Atm-/- astrocytes. Here we show that Atm-/- astrocytes exhibit premature senescence, express constitutively high levels of p21, and have impaired p53 stabilization. However, in contrast to radiosensitive Atm-/- fibroblasts and radioresistant Atm-/- neurons, survival of Atm-/- astrocytes after IR was similar to wild-type astrocytes. Additionally, p53-null astrocytes, but not fibroblasts, were moderately more radioresistant than their wild-type counterparts, suggesting that the deficit in p53 stabilization observed in Atm-null cells is not a measure of radiation susceptibility. Thus, in astrocytes, the function of Atm in cellular growth and radiosensitivity is distinct. These data may have implications for ATM disruption strategies as a radiosensitizing treatment for brain tumors. PMID- 10537313 TI - Characterization of glycosylphosphatidylinositol-linked molecule CD55/decay accelerating factor as the receptor for antibody SC-1-induced apoptosis. AB - The human monoclonal antibody SC-1 induces apoptosis of stomach carcinoma cells and is currently used in a clinical Phase II trial. The antibody binds to a target molecule that is preferentially expressed on diffuse- and intestinal-type stomach cancer cells and shows a very restricted expression on other normal and malignant tissues. In this paper, we show that the SC-1 receptor is a stomach carcinoma-associated isoform of CD55 [membrane-bound decay-accelerating factor (DAF)-B] with a relative molecular mass of approximately 82 kDa. The antigenic site of SC-1 is an N-linked carbohydrate residue. Cross-linking of the DAF receptor increases apoptotic activity. SC-1 binding induces tyrosine phosphorylation of three proteins of approximately 60, 75, and 110 kDa, whereas a serine residue of an approximately 35-kDa protein is dephosphorylated. Expression of caspase-3 (CPP32) and caspase-8 (FLICE) is elevated, and activation of these caspases occurs. These data show that a tumor-specific variant form DAF is involved in apoptosis and can be used for adjuvant therapeutical purposes on gastric carcinoma. PMID- 10537314 TI - Urokinase receptor interacts with alpha(v)beta5 vitronectin receptor, promoting urokinase-dependent cell migration in breast cancer. AB - Perturbation of adhesive interactions at cell-substratum and cell-cell contact sites is a critical event in the multistep process of cancer invasion. Recent studies indicate that the urokinase receptor (uPAR) is associated in large molecular complexes with other molecules, such as integrins. To test the possibility that uPAR may physically and functionally interact with vitronectin (Vn) receptors, we determined the expression level of uPAR, alpha(v)beta3, and alpha(v)beta5 Vn receptors in 10 human breast carcinomas. Here, we show the ability of uPAR to physically associate with alpha(v)beta5 in the breast carcinomas examined. The functional effects of this interaction were studied using HT1080 human fibrosarcoma and MCF-7 human breast carcinoma cell lines, both exhibiting a urokinase-dependent physical association between uPAR and alpha(v)beta5. Both cell lines respond to urokinase or to its noncatalytic amino terminal fragment by exhibiting remarkable cytoskeletal rearrangements that are mediated by alpha(v)beta5 and require protein kinase C activity. On the contrary, binding of Vn to alpha(v)beta5 results in the protein kinase C-independent formation of F-actin containing microspike-type structures. Furthermore, alpha(v)beta5 is required for urokinase-directed, receptor-dependent MCF-7 and HT1080 cell migration. These data show that uPAR association with alpha(v)beta5 leads to a functional interaction of these receptors and suggest that uPAR directs cytoskeletal rearrangements and cell migration by altering alpha(v)beta5 signaling specificity. PMID- 10537315 TI - Ribozyme-mediated down-regulation of ErbB-4 in estrogen receptor-positive breast cancer cells inhibits proliferation both in vitro and in vivo. AB - ErbB-4 is a recently discovered member of the class I receptor tyrosine kinase family (ErbB). Little is known about its expression and its importance in human malignancy. To delineate the biological function of ErbB-4 receptors in breast cancer, we used a hammerhead ribozyme strategy to achieve down-regulation of ErbB 4 receptors in various breast cancer cell lines. We observed that down-regulation of ErbB-4 in estrogen receptor-positive (ER+) human breast cancer cell lines (MCF 7 and T47D), which express relatively high levels of ErbB-4, significantly inhibited colony formation. No effects were observed in estrogen receptor negative (ER-) MDA-MB-453 cells, which express low levels of endogenous ErbB4 and high levels of ErbB-2 and ErbB-3. This occurred despite the fact that fluorescence-activated cell sorter analysis of these latter cells revealed that the expression of the ErbB-4 receptor was completely abrogated by ribozyme treatment. Furthermore, down-regulation of ErbB-4 in T47D and MCF-7 cells significantly inhibited tumor formation in athymic nude mice (P < 0.03 and P < 0.001, respectively). In addition, NRG-stimulated phosphorylation of ErbB-4- and NRG-induced colony formation was significantly reduced in ribozyme-transfected T47D cells. These data provide the first evidence that elevation of ErbB-4 expression plays a role in the proliferation of some ER+ human breast cancer cell lines (T47D and MCF-7) that express high levels of ErbB-4. We have also investigated the expression of ErbB-4 in human primary breast carcinoma specimens, using immunohistochemical staining with an anti-ErbB-4 monoclonal antibody. ErbB-4 expression was found in 60% of the 50 primary breast tumors examined, and high intense immunoreactivity of ErbB-4 was detected in 18% of these primary breast tumors. ErbB-4 receptor expression appeared to correlate with ER+ primary breast tumors. A similar correlation was also observed in the human breast cancer cell lines. These results provide a better understanding of the biological significance of ErbB-4 receptor in breast cancer. Our data suggest that elevation of the ErbB-4 receptor plays a role in ER+ breast cancer cell proliferation. Moreover, ribozyme technology provides a useful tool to delineate the role of a particular gene product. PMID- 10537316 TI - Apoptosis induction of human lung cancer cell line in multicellular heterospheroids with humanized antiganglioside GM2 monoclonal antibody. AB - The chimeric antiganglioside GM2 monoclonal antibody (MAb) KM966, which showed high effector functions such as complement-dependent cytotoxicity and antibody dependent cellular cytotoxicity (ADCC), potently suppressed growth and metastases of GM2-positive human cancer cells inoculated into mice. To further improve the therapeutic efficacy of the anti-GM2 MAb in humans, we constructed a humanized anti-GM2 MAb, KM8969. The humanized KM8969 was more efficient in supporting ADCC against GM2-positive human cancer cell lines than the chimeric KM966, whereas complement-dependent cytotoxicity was slightly reduced in the humanized KM8969. In addition, the humanized KM8969 was shown to exert potent ADCC mediated by both lymphocytes and monocytes. To investigate the effect of the humanized KM8969 on the biological function of GM2 in the condition physiologically mimicking formation and growth of cancer masses, the heterospheroids composed of normal human dermal fibroblasts and GM2-positive human lung cancer cells were developed. Interestingly, the humanized KM8969 gave rise to growth inhibition of heterospheroids without dependence of the effector functions. Morphological and immunocytochemical analysis suggested that the inhibitory effect was due to the apoptosis of GM2-positive cancer cells in the heterospheroids. The result indicates that GM2 captured by the antibody on the cell surface loses its physiological function that plays a critical role in maintaining the three dimensional growth of cancer cells in contact with its own cells or other type of cells in a microenvironment. The humanized KM8969, which can destroy the cancer cells via blocking functional GM2 on the cell surface as well as the effector functions, would have extraordinary potential in human cancer therapy. PMID- 10537317 TI - Biological activities and signaling pathways of the granulin/epithelin precursor. AB - Growth-regulated cells, such as 3T3 mouse embryo fibroblasts (MEFs), require more than one growth factor for growth, usually the insulin-like growth factor I (IGF I) in combination with either platelet-derived growth factor or epidermal growth factor. Singly, these growth factors cannot sustain the growth of 3T3 cells. However, if the IGF-I receptor (IGF-IR) is even modestly overexpressed, then IGF I, by itself, stimulates the growth of MEFs in monolayer and makes them capable of forming colonies in soft agar. The granulin/epithelin precursor (GEP) has been identified as the only growth factor, thus far, that can stimulate by itself the growth of R- cells, a 3T3-like cell line in which the genes for the IGF-IR have been deleted. We have expressed GEP in R- cells and show that these cells can now grow in serum-free medium. GEP, however, cannot replace other functions of the IGF-IR, such as protection from apoptosis (anoikis) or transforming activity (colony formation in soft agar). GEP activates, in R- cells, the two signaling pathways that are known to be sufficient for IGF-I-mediated mitogenesis in cells overexpressing the IGF-IR, the mitogen-activated protein kinase and the phosphatidylinositol 3-kinase pathways. This may explain why GEP, by itself, can replace the IGF-IR for growth in monolayer cultures. It also confirms that, for transformation, other pathways must be activated besides the two pathways that are sufficient for mitogenesis. PMID- 10537318 TI - Microinjection of anti-p21 antibodies induces senescent Hs68 human fibroblasts to synthesize DNA but not to divide. AB - Replicative senescence is characterized by irreversible growth arrest and has been defined by four genetic complementation groups. One of these groups is associated with the predominance of underphosphorylated, growth-suppressive retinoblastoma tumor suppressor protein (pRb). Although certain members of the cyclin-dependent kinase (cdk)/cyclin family, some of which phosphorylate pRb, are underexpressed in senescent cells, others are expressed but inactive. This lack of cdk activity and arrest in the G1 phase of the cell cycle is likely attributable to the induction upon senescence of the G1-S cdk/cyclin inhibitors p21 (WAF1/CIP1/Sdi) and p16INK4. In fact, in early presenescent normal diploid fibroblasts in which p21 is inactivated, senescence is bypassed or postponed. Moreover, in senescent cells in which p53 function was inhibited, DNA synthesis was reinitiated, an effect likely attributable, in part, to the dependence of p21 expression on p53. We report here that the apparent inactivation of p21 in senescent human fibroblasts through the introduction of inhibitory alpha-p21 antibodies causes these cells to reenter the S-phase of the cell cycle. The disruption of p21 activity affects the p21-Rb-E2F pathway in that the expression of genes transcriptionally regulated by E2F, such as cyclin A and cdc2, were found to be up-regulated in injected cells. No evidence of cell division was observed. This suggests that p21 plays an important role in the maintenance of senescence and in the inhibition of S-phase progression, but inhibition of p21 activity is insufficient to permit cells to complete the cell cycle. PMID- 10537319 TI - A mammalian severin replaces gelsolin in transformed epithelial cells. AB - A persisting paradox in cytoskeletal regulation of cell motility is the loss of the actin filament fragmenting protein, gelsolin, in transformed epithelial cells that have gained the ability to migrate. Either actin filament severing does not occur during motility of carcinoma cells or a novel fragmentation protein is expressed during transformation. Using an antibody specific for severin, the Mr 40,000 actin filament severing protein from Dictyostelium discoideum amoebae, we have identified a mammalian form of severin in murine LL/2 carcinoma cells lacking gelsolin. Mammalian severin (M-severin) isolated from LL/2-derived Lewis lung carcinoma tumors severed F-actin in a calcium-dependent manner, mimicking the function of Dictyostelium severin. M-severin preferentially localized to the cleavage furrow of dividing LL/2 cells and to the actin-rich cortex of migratory LL/2 cells, known sites of active actin cytoskeleton rearrangement. The mammalian severing protein was fully expressed in transformed LL/2 epithelial cells but went undetected in normal mouse muscle, liver, spleen, or kidney. Normal mouse lung tissue contained minute amounts of M-severin, attributed to motile cells in pulmonary connective tissue. In striking contrast to M-severin, gelsolin was highly expressed in normal lung but disappeared in transformed LL/2 carcinoma cells. Based on prior observations of a functional role for actin filament fragmentation in cell migration, the simultaneous induction of M-severin and loss of gelsolin during epithelial transformation suggests that replacement of gelsolin by M-severin may function to achieve actin filament rearrangements necessary for active cell migration in invasive or metastatic carcinoma. Induction of M-severin in an invasive tumor was directly observed in human colon adenocarcinoma by cytoimmunohistochemistry with antibodies directed against severin isolated from both Dictyostelium amoebae and Lewis lung carcinoma cells. Because normal colon epithelium from the same patient did not express M-severin, it may serve as a sensitive marker for detection and staging of epithelial tumors. PMID- 10537320 TI - Fas ligand is expressed on human squamous cell carcinomas of the head and neck, and it promotes apoptosis of T lymphocytes. AB - Recent reports have variously described expression of Fas ligand (FasL) or its absence in human tumors. The importance of the Fas-FasL mechanism for the immune evasion by tumors provided a strong rationale for the examination of FasL expression and function in squamous cell carcinoma of the head and neck (SCCHN), which is one of the most immunosuppressive human cancers. Using immunostaining or immunoblotting, SCCHN cell lines and tumor biopsies were examined for the presence of the components of the Fas-FasL pathway and found to express Fas, as well as both the full-length and cleaved forms of FasL. By reverse transcription PCR, mRNA for FasL and Fas were detected in all SCCHN tested, and cross hybridization to radioactive Fas and FasL cDNA probes confirmed the specificity of amplification. To demonstrate that FasL expressed on cell surface of SCCHN cells was biologically active, various SCCHN lines were coincubated with the Fas sensitive Jurkat T-cell lines or activated peripheral blood mononuclear cells. Tumor-induced apoptosis of T cells was dependent on the ratio of tumor cells: lymphocytes. It was significantly but only partially inhibited by neutralizing antibodies to FasL and antagonistic antibodies to FasR. Tumor-induced apoptosis was enhanced by the pretreatment of tumor cells with metalloproteinase inhibitors, which increased expression of FasL on tumor cells. Supernatants of tumor cells transduced with FasL also induced apoptosis of Jurkat cells. Thus, coincubation of SCCHN with Fas-sensitive lymphocytes can induce apoptosis of these lymphocytes, and the Fas/FasL pathway appears to be responsible, at least in part, for tumor-induced lymphocyte death. The data suggest that the Fas/FasL pathway is potentially immunosuppressive and may be involved in the escape of human carcinoma cells from immune destruction. PMID- 10537321 TI - Response of human tumor cells of varying radiosensitivity and radiocurability to fractionated irradiation. AB - The cytotoxic effects of radiation delivered in daily fractions of 2.0 Gy were examined in plateau phase cultures of human tumor cells of varying in vitro radiosensitivity, derived from tumors of varying radiocurability. Among the eight cell lines examined, three types of responses to fractionated irradiation were observed. In the group composed of tumor cell lines that were radioresistant in culture (D0 > 2 Gy) and derived from known local radiation failures or from tumor histologies associated with radiation failure, a gradual linear reduction in surviving fraction versus total dose was observed. In a second group, composed of cell lines that were radiosensitive in culture (D0 approximately 1 Gy) but derived from known radiation failures, the surviving fraction initially declined and began to plateau after 6 Gy (three fractions of 2 Gy). In the third group, composed of radiosensitive cell lines derived from tumors associated with high radiocurability, a rapid decline in surviving fraction versus total dose was observed. The in vitro response of human tumor cells to fractionated irradiation delivered at clinically relevant doses appears to be independent of in vitro X ray sensitivity and p53 status but related to clinical radiocurability, suggesting a possible role in predicting tumor response to radiotherapy. PMID- 10537322 TI - Distinctive expression and functions of the type 4 endothelial differentiation gene-encoded G protein-coupled receptor for lysophosphatidic acid in ovarian cancer. AB - Endothelial differentiation gene (edg)-encoded G protein-coupled receptors (Edg Rs)-1, -3, and -5 bind sphingosine 1-phosphate (S1P), and Edg-2 and -4 bind lysophosphatidic acid (LPA). Edg Rs transduce signals from LPA and S1P that stimulate ras- and rho-dependent cellular proliferation, enhance cellular survival, and suppress apoptosis. That high levels of LPA in plasma and ascitic fluid of patients with ovarian cancer correlate with widespread invasion suggested the importance of investigating expression and functions of Edg Rs in ovarian cancer cells (OCCs) as compared with nonmalignant ovarian surface epithelial cells (OSEs). Analyses of Edg Rs by semiquantitative reverse transcription-PCR, a radioactively quantified variant of PCR, and Western blots developed with monoclonal antibodies showed prominent expression of Edg-4 R in primary cultures and established lines of OCCs but none in OSEs. In contrast, levels of Edg-2, -3, and -5 were higher in OSEs than OCCs. LPA stimulated proliferation and signaled a serum response element-luciferase reporter of immediate-early gene activation in OCCs but not OSEs, whereas S1P evoked similar responses in both OSEs and OCCs. Pharmacological inhibitors of Edg R signaling suppressed OCC responses to LPA. A combination of monoclonal anti-Edg-4 R antibody and phorbol myristate acetate, which were inactive separately, evoked proliferative and serum response element-luciferase responses of OCCs but not OSEs. Thus the Edg-4 R may represent a distinctive marker of OCC that transduces growth-promoting signals from the high local concentrations of LPA characteristic of aggressive ovarian cancer. PMID- 10537323 TI - Signaling pathways and structural domains required for phosphorylation of EMS1/cortactin. AB - The structural characteristics of EMS1 (human cortactin) suggest that it may link signaling events to reorganization of the actin cytoskeleton. Interestingly, the EMS1 gene is commonly amplified and overexpressed in several human cancers, which may alter their invasive or metastatic properties. An 80 to 85-kDa mobility shift of EMS1 correlates with an alteration in subcellular distribution and is likely to represent an important regulatory event. In HEK 293 cells, epidermal growth factor treatment or cell detachment induced this shift, and this was blocked by the mitogen-activated protein/extracellular signal-regulated kinase kinase (MEK) inhibitor PD98059. Furthermore, expression of a constitutively active form of MEK induced the shift, indicating that MEK activation was both sufficient and necessary for this modification. The epidermal growth factor-induced shift correlated with increased phosphorylation on serine and threonine residues of the same tryptic phosphopeptides detected under basal conditions. Deletion of the helical-proline-rich region of the protein blocked the mobility shift and EMS1 phosphorylation. In vitro kinase assays demonstrated that the extracellular signal-regulated kinases represent candidate kinases for this region, although other MEK-regulated enzymes must also participate. These data identify MEK as an important intermediate involved in EMS1 phosphorylation and highlight the helical proline-rich region as a key regulatory domain. PMID- 10537324 TI - Immunohistochemical localization of caspase-3 correlates with clinical outcome in B-cell diffuse large-cell lymphoma. AB - Although B-cell diffuse large-cell lymphoma (DLCL) can respond to chemotherapy and radiotherapy, a large number of patients are still resistant to treatment. Caspase-3 is an enzyme crucial to the apoptotic process and may be important in the clinical outcome of these patients. The pattern of caspase-3 expression was studied in 54 cases of DLCL using immunohistochemistry and quantitative reverse transcription PCR. Tumor cells displayed both a diffuse cytosolic and a punctate cytosolic staining for caspase-3. Kaplan-Meier survival curves indicated that tumor cells with a diffuse cytosolic expression of caspase-3 correlated with a poor prognosis (P > 0.0004). In addition, a punctate cellular localization was associated with complete response to treatment (P = 0.011). Cases with a small percentage of lymphoma cells expressing caspase-3 also tended to show poor survival (P > 0.09). Levels of caspase-3 mRNA were not significant (P > 0.17), although a weak trend was observed similar to the immunohistochemical analysis. The pattern of expression of caspase-3 was also assessed with respect to terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) positivity in both reactive lymph nodes and B-cell DLCL cases. Our results suggest that TUNEL positive cells are not caspase-3-positive and that there is no correlation between DLCL cases with a high degree of DNA fragmentation and caspase-3 immunostaining. Furthermore, a survival curve indicated that a high TUNEL positivity was associated with a poor survival probability (P < 0.02) and a poor response to treatment (P = 0.04). These results confirm the dynamic nature of caspase-3 expression in DLCL and suggests that the pattern of expression of the enzyme has prognostic significance. PMID- 10537325 TI - Gangliosides influence angiogenesis in an experimental mouse brain tumor. AB - Gangliosides are sialated glycosphingolipids present on the plasma membranes of all vertebrate cells. Tumors shed gangliosides into the extracellular microenvironment, which may influence tumor-host cell interactions. We have investigated the role of gangliosides on the growth and angiogenesis of the EPEN experimental mouse brain tumor. EPEN cells express only ganglioside G(M3), and the solid tumors formed in vivo are sparsely vascularized with extensive necrosis. We stably transfected the EPEN cells with the cDNA for N acetylgalactosaminyl transferase, a key enzyme for the synthesis of complex gangliosides. In addition to G(M3), the transfected cell line (EPEN-GNT) expressed complex gangliosides G(M2), G(M1), and G(D1a). The EPEN-GNT tumor was more densely vascularized with less necrosis and grew more rapidly than the nontransfected EPEN or mock-transfected (EPEN-V) control tumors. Also, VEGF gene expression was higher in the EPEN-GNT tumor than in the control tumors. The synthesis of complex gangliosides in the EPEN-GNT tumor cells also stimulated vascularization in an in vivo Matrigel assay for angiogenesis. These results indicate that the ratio of G(M3) to complex gangliosides can influence the growth and angiogenic properties of the EPEN experimental brain tumor and are consistent with previous findings in other systems. We conclude that gangliosides may be important modulators of brain tumor angiogenesis. PMID- 10537326 TI - Oral cancer chemotherapy: the promise and the pitfalls. PMID- 10537327 TI - Dihydropyrimidine dehydrogenase: its role in 5-fluorouracil clinical toxicity and tumor resistance. PMID- 10537328 TI - Prostate-specific membrane antigen is produced in tumor-associated neovasculature. AB - Prostate-specific membrane antigen (PSMA), a type II transmembrane protein, was originally thought to be strictly expressed in prostatic tissue, but recent studies have demonstrated PSMA protein expression in nonprostatic tumor neovasculature as well. Using immunohistochemistry, reverse transcription-PCR assays, and in situ hybridization, we have demonstrated PSMA mRNA transcripts and protein expression in the endothelium of tumor-associated neovasculature of multiple nonprostatic solid malignancies. In addition, we found no PSMA mRNA or protein expression in the vascular endothelial cells of corresponding benign tissue examples. Our findings expand the possible therapeutic role of PSMA and establish it as a unique biomarker specifically produced and expressed by tumor associated neovasculature but not produced or expressed by normal vessels. PMID- 10537329 TI - Differences in Ki67 and c-erbB2 expression between screen-detected and true interval breast cancers. AB - Breast cancer screening facilitates the early detection of breast cancer, although a significant number of tumors still arise in the interval between screening. The objective of this study was to measure the expression of five markers of proven prognostic significance in symptomatic breast cancer (estrogen receptor, progesterone receptor, p53, Ki67, and c-erbB2) in screen-detected and interval breast cancers to identify biological markers that may be associated with the emergence of symptomatic breast cancer in the screening interval. The expression of estrogen receptor, progesterone receptor, p53, Ki67, and c-erbB2 was assessed in a series of 51 true interval and 84 screened-detected invasive tumors by immunohistochemistry. Interval cancers tended to be of higher histological grade and were of larger pathological size than screen-detected cancers. Expression of estrogen receptor was 1.7-fold lower (P<0.001), whereas expression of p53 was 2.5-fold (P<0.01), Ki67 2.4-fold (P<0.001), and c-erbB2 3.6 fold higher (P<0.01) in true interval cancers compared with screen-detected invasive cancers. There was no significant difference in progesterone receptor expression. The most important differences identified by multiple logistic regression analysis were in the expression of Ki67 and c-erbB2. The differences in the expression of these markers were more important than clinical features such as pathological grade and size. Using the logistic regression model, 83% of the tumors analyzed in this study could be correctly assigned as interval or screen-detected tumors on the basis of Ki67 and c-erbB2 expression. The importance of high expression of Ki67 in interval cancers compared with screen detected cancers suggests that tumors may become symptomatic in the screening interval as a result of increased levels of cell proliferation. The inclusion of c-erbB2 in the regression equation suggests that this growth factor receptor may play a significant role in stimulating the rapid growth of interval cancers. PMID- 10537330 TI - Detection of microsatellite alterations in plasma DNA of non-small cell lung cancer patients: a prospect for early diagnosis. AB - A major problem in lung cancer is the lack of clinically useful tests for early diagnosis and screening of an asymptomatic population by non-invasive diagnostic procedures. Recent studies have demonstrated the possibility to detect genetic alterations in plasma or serum DNA from patients with various cancers. However, these data rely on small series of aggressive tumors with advanced-stage disease. To determine whether genetic changes in plasma are also detectable in patients with limited disease and thereby potentially useful for early detection, we looked for microsatellite instability (allele shift) and loss of heterozygosity in plasma DNA of 87 stage I-III non-small cell lung cancers and 14 controls. Combining two markers with a high rate of instability (D21S1245) and loss of heterozygosity (FHIT locus), a microsatellite alteration was observed in 49 of 87 (56%) non-small cell lung cancer tumors and in 35 of 87 (40%) plasma samples. Thirty of 49 (61%) of the cases showing tumor alterations also displayed a change in plasma DNA; in addition, 5 patients displayed alterations in plasma samples only. None of the control individuals had genetic changes in plasma. No association was found between the frequency of microsatellite alterations in plasma and tumor stage or histology. Of interest, plasma DNA abnormalities were detectable in 43% of pathological stage I cases and in 45% of tumors up to 2 cm in maximum diameter. These findings highlight new prospects for early tumor detection by noninvasive screening procedures based on the analysis of genetic changes in plasma. PMID- 10537331 TI - Myeloma cells release soluble interleukin-6Ralpha in relation to disease progression by two distinct mechanisms: alternative splicing and proteolytic cleavage. AB - Multiple myeloma (MM) is a plasma-cell malignancy characterized by the accumulation of malignant plasma cells within the bone marrow. Interleukin (IL)-6 is an essential survival and growth factor for myeloma cells that exerts its activity through a cell surface receptor composed of an 80-kDa ligand binding molecule (IL-6Ralpha) and a 130-kDa signal-transducing molecule. Of major interest, the soluble form of the IL-6Ralpha (sIL-6Ralpha) is an agonistic molecule able to potentiate IL-6 activity and a strong prognostic factor in MM. In the present study, we demonstrate that purified myeloma cells from all of the patients with MM and human myeloma cell lines release sIL-6Ralpha. The level of sIL-6Ralpha release correlates with disease activity and is clearly up-regulated during tumoral expansion in vivo and immortalization in vitro. Of note, this sIL 6Ralpha release is strongly reduced (50%) by a hydroxamate-based metalloproteinase inhibitor underlying the importance of shedding in the production of sIL-6Ralpha by myeloma cells. Using specific IL-6Ralpha primers flanking the transmembrane domain, we demonstrate by PCR the presence of two IL 6R mRNAs corresponding to the membrane IL-6Ralpha and to the sIL-6Ralpha generated through alternative splicing in myeloma cells. In conclusion, we show that: (a) native myeloma cells and human myeloma cell lines release sIL-6Ralpha by two distinct mechanisms: alternative splicing and proteolytic cleavage of the membrane IL-6Ralpha; and (b) the release of the sIL-6Ralpha, which is an agonist of IL-6, correlates with disease progression, explaining in part its strong prognostic value in vivo. PMID- 10537332 TI - Association between keratin and vimentin expression, malignant phenotype, and survival in postmenopausal breast cancer patients. AB - Pathology observational reports and experimental data suggest that keratin and vimentin intermediate filament (IF) coexpression in breast cancer confers a more aggressive "interconverted" phenotype, expressing both epithelial and mesenchymal markers. In this study, we extended previous observations by measuring the expression of keratin and vimentin, in relation to other selected biomarkers of disease progression, in postmenopausal women with breast cancer. Using immunohistochemical analysis of 54 archival, formalin-fixed, paraffin-embedded invasive breast cancers from a well-defined cohort, we examined relative IF (keratin and vimentin) expression in a semiquantitative fashion and compared these results with other biological markers and survival. By univariate analysis, we found that vimentin expression was inversely associated with keratin expression alone (P = 0.0089) and directly related to histological grade (P = 0.017), nuclear grade (P = 0.027), Ki67 growth fraction (P = 0.024), and epidermal growth factor receptor immunostaining (P = 0.019). The relative expression of keratin and vimentin in approximately similar amounts characterized tumors with the poorest prognosis, as compared with keratin-high/vimentin negative or keratin-low/vimentin-positive tumors. These latter two groups demonstrated similar Kaplan-Meier survival curves; the former group (keratin and vimentin in approximately similar amounts) demonstrated a poorer survival, with a hazard ratio of 2.1 (95% confidence interval, 0.5-9.6). These data suggest that relative keratin and vimentin IF expression is more indicative of prognosis and tumor phenotype than either IF marker detected independently. PMID- 10537333 TI - Mechanisms of inactivation of p14ARF, p15INK4b, and p16INK4a genes in human esophageal squamous cell carcinoma. AB - The 9p21 gene cluster, harboring growth suppressive genes p14ARF, p15INK4b, and p16INK4a, is one of the major aberration hotspots in human cancers. It was shown that p14ARF and p16INK4a play active roles in the p53 and Rb tumor suppressive pathways, respectively, and p15INK4b is a mediator of the extracellular growth inhibition signals. To elucidate specific targets and aberrations affecting this subchromosomal region, we constructed a detailed alteration map of the 9p21 gene cluster by analyzing homozygous deletion, hypermethylation, and mutation of the p14ARF, p15INK4b, and p16INK4a genes individually in 40 esophageal squamous cell carcinomas (ESCCs) and compared the genetic alterations with mRNA expression in 18 of these samples. We detected aberrant promoter methylation of the p16INK4a gene in 16 (40%), of p14ARF in 6 (15%), and of p15INK4b in 5 (12.5%) tumor samples. Most p16INK4a methylations were exclusive, whereas all but one of the p14ARF/p15INK4b methylations were accompanied by concomitant p16INK4a methylation. We detected homozygous deletion of p16INK4a in 7 (17.5%), of p14ARF E1beta in 13 (33%), and of p15INK4b in 16 (40%) tumor samples. Most deletions occurred exclusively on the E1beta-p15INK4b loci. Two samples contained p14ARF deletion but with p16INK4a and p15INK4b intact. No mutation was detected in the p14ARF and p16INK4a genes. Comparative RT-PCR showed good concordance between suppressed mRNA expression and genetic alteration for p15INK4b and p16INK4a genes in the 18 frozen samples, whereas 5 of the 13 cases with suppressed p14ARF mRNA expression contained no detectable E1beta alteration but aberrations in the p16INK4a locus. Our results show that in human ESCCs, p14ARF is a primary target of homozygous deletion along with p15INK4b, whereas p16INK4a is the hotspot of hypermethylation of the 9p21 gene cluster. The frequent inactivation of the p14ARF and p16INK4a genes may be an important mechanism for the dysfunction of both the Rb and p53 growth regulation pathways during ESCC development. PMID- 10537334 TI - Sensitization of tumor cells to ribotoxic stress-induced apoptotic cell death: a new therapeutic strategy. AB - We describe a procedure that sensitizes chemotherapy-and tumor necrosis factor resistant human tumor cell populations in vitro and in nude mouse transplants to the immediate triggering of high rates of cell death by anisomycin, an agent causing activation of stress-activated protein kinases [SAPKs, as defined by P. Cohen (Trends Cell Biol., 7: 353-361, 1997)] including p38/RK and c-jun NH2 terminal kinase homologues, following its binding to ribosomal 28S RNA (M. S. Iordanov et al, Mol. Cell. Biol., 17: 3373-3381, 1997). Sensitization is effected by successive application of an inhibitor of histone deacetylation (trichostatin A, butyrate) and of flavopiridol, known as an inhibitor of cyclin dependent kinases and evaluated presently in clinical trials. Effective concentrations of anisomycin, flavopiridol, and trichostatin A are in the submicromolar range. Tumor cell death can be prevented by epidermal growth factor (EGF), if added before flavopiridol or after anisomycin but not if applied between the additions of these agents, suggesting that flavopiridol interrupts an EGF-activated survival pathway and that anisomycin, besides triggering cell death, guards this pathway against the interference by flavopiridol. In contrast to EGF, dibutyryl cAMP exerts protection that is flavopiridol-insensitive. For triggering cell death, anisomycin cannot be replaced by DNA- or mitotic spindle-targeted drugs in this system. The present findings, that a combination of transcriptional and signal transduction-targeted modulators sensitizes tumor cells synergetically to stress-mediated triggering of cell death and that ribotoxic stress is more efficient in this respect than genotoxic or spindle-targeted stress, bear important implications for the therapeutic exploitation of cellular stress responses. The stepwise sensitization and triggering of cell death in the present system allow separate analysis and manipulation of processes contributing to cellular death susceptibility and of the mechanism responsible for triggering cell death, thus providing the operational basis for further development of this therapeutic approach. PMID- 10537335 TI - Interferon-alpha-mediated down-regulation of angiogenesis-related genes and therapy of bladder cancer are dependent on optimization of biological dose and schedule. AB - The purpose of this study was to identify and optimize the antiangiogenic activity of IFN-alpha against human bladder cancer cells growing in the bladder of nude mice. 253J B-V IFN(R) cells (resistant to antiproliferative effects of IFN-alpha or IFN-beta) were implanted into the bladder wall of nude mice. Three days later, the mice were treated with s.c. injections of IFN-alpha (70,000 units/week) at different dosing schedules (1, 2, 3, or 7 times/week). Daily therapy with IFN-alpha produced the most significant inhibition of tumor growth, tumor vascularization, and down-regulation of basic fibroblast growth factor and matrix metalloprotease-9 mRNA and protein expression. Changing dose and schedule of IFN-alpha administration had minimal effects on the expression of vascular endothelial growth factor or interleukin 8. The daily s.c. administrations of 5,000 or 10,000 units IFN-alpha-2a produced maximal inhibition of bFGF and MMP-9 expression (mRNA and protein), maximal reduction in tumor vessel density, and maximal reduction in serum levels of bFGF. Daily administration of higher doses of IFN-alpha failed to produce significant antiangiogenic effects. These data suggest that the antiangiogenic activity of IFN-alpha is dependent on frequent administration of optimal biological dose and not maximal tolerated dose. PMID- 10537336 TI - Selective in vivo mobilization with granulocyte macrophage colony-stimulating factor (GM-CSF)/granulocyte-CSF as compared to G-CSF alone of dendritic cell progenitors from peripheral blood progenitor cells in patients with advanced breast cancer undergoing autologous transplantation. AB - Dendritic cells (DCs) are potent antigen-presenting cells that are essential for the initiation of T cell-mediated immunity. DCs develop from myeloid progenitor populations under the influence of granulocyte macrophage colony-stimulating factor (GM-CSF) and pass through an intermediate stage of maturation that is characterized by CD14 expression. Interest has focused on generating human derived DCs for antigen-specific tumor vaccines to be used as adjuvant immunotherapy in minimal disease settings, such as after autologous transplantation. In the present study, mobilized peripheral blood progenitor cells (PBPCs) were obtained from 18 patients with locally advanced or metastatic breast cancer preparing to undergo autologous stem cell transplantation. PBPCs mobilized in 10 patients with GM-CSF for 1 week, followed by the combination of GM-CSF and G-CSF, were compared with those obtained from patients receiving G-CSF alone with respect to the presence of DC progenitors and the capacity to generate functionally active mature DCs. PBPCs mobilized with GM-CSF/G-CSF were markedly enriched for CD14+ DC progenitor cells as compared with those mobilized with G CSF alone. Consistent with an immature progenitor population, the CD14+ cells express Ki-67 antigen but not nonspecific esterase. CD14+ cells purified by fluorescence-activated cell sorting from PBPCs mobilized with either regimen and cultured for 1 week in GM-CSF and interleukin-4 generated nearly pure populations of cells with characteristic DC phenotype and function. The addition of GM-CSF to the mobilization regimen resulted in greater yields of functionally active DCs for potential use in posttransplant immunotherapy. PMID- 10537337 TI - Inter- and intrapatient variability in etoposide kinetics with oral and intravenous drug administration. AB - The objective of this study was to accurately determine the within- and between patient variability in etoposide pharmacokinetics for i.v. and p.o. administered drug. Inter-and intrapatient variability in systemic etoposide exposure was measured following i.v. and p.o. drug administration using stable isotope dilution methodology. Seven patients received 50 mg of etoposide by both p.o. and i.v. routes of administration on three separate occasions 1 month apart. Etoposide plasma concentrations following p.o. and i.v. drug administration were quantitated by liquid chromatography-mass spectrometry for each route of administration. The area under the plasma etoposide concentration versus time curve, plasma etoposide clearance, and etoposide plasma half-life were calculated for each dose of drug. Kinetic measurements following i.v. and p.o. drug administration were compared. The within-patient variation in the areas under the plasma etoposide concentration versus time curves following i.v. drug administration was minimal [coefficient of variation (CV) = 9.3%]. Within-patient variability was increased 2.4-fold with oral drug administration (intrapatient CV = 22.2%). Between-patient variability was roughly three times as great as within patient variability (interpatient i.v. CV = 28.4%; interpatient p.o. CV = 58.3%). Mean etoposide bioavailability at a dose of 50 mg was 64.6%, again with greater interpatient than intrapatient variability (34.8 versus 22.6%). Greater variation in drug toxicity is expected with p.o. compared with i.v. etoposide use. Administration of repeated doses of etoposide to the same patient should produce less variation in toxicity than between-patient dosing. PMID- 10537338 TI - A phase I trial of humanized monoclonal antibody HuM195 (anti-CD33) with low-dose interleukin 2 in acute myelogenous leukemia. AB - HuM195 is a recombinant humanized IgG1 monoclonal antibody reactive with CD33, a Mr 67,000 glycoprotein expressed on early myeloid progenitor cells and myeloid leukemia cells. HuM195 has been shown to rapidly target and saturate acute myeloid leukemia (AML) cells after i.v. infusion into patients and is capable of mediating antibody-dependent cellular cytotoxicity. This activity is enhanced in vitro when natural killer (NK) effector cells are preincubated with low concentrations of interleukin 2 (IL-2). Previous Phase I trials of HuM195 in patients with relapsed AML demonstrated safety and attainment of complete responses, but significant antileukemic activity appears limited to patients with low leukemia tumor burdens. Therefore, in the present trial, we sought to determine whether low-dose IL-2 could safely enhance the numbers of NK cells and therefore the cytotoxic capability of HuM195 via presumptive NK cell antibody dependent cellular cytotoxicity in vivo against myeloid leukemia cells. Thirteen patients with relapsed or refractory AML and one patient with advanced myelodysplastic syndrome were treated with 0.6x10(6) IU/m2 of s.c. IL-2 daily for 35 days. Starting on day 15, patients received twice weekly i.v. infusions of HuM195 (3.0 mg/m2) for 3 weeks. Immediately after the HuM195 infusion, the patients received IL-2 i.v. infusions over 2 h at one of three escalating dose levels of 0.5x10(6), 1.0x10(6), and 2.0x10(6) IU/m2. Peripheral blood mononuclear cells were quantitated and immunophenotyped by flow cytometry. Safety, tolerability, bone marrow mononuclear cell morphology, and immunophenotype, as well as responses were assessed. Of the 14 patients who entered the study, 10 were able to complete at least one cycle of therapy. Adverse effects to the s.c. IL-2 were relatively mild and included erythema and induration of the skin at the injection site and low-grade fever. Toxicity from the sequential HuM195 and i.v. IL-2 infusions included nausea, rigors, and fever. Toxicity was IL-2 dose related with dose-limiting toxicity seen at the 2.0x10(6) IU/m2 dose level. Three patients had stable disease at the completion of the first cycle and went on to receive a second cycle of treatment. CD3-positive, CD56-positive, and CD33 positive cells were generally found to significantly decrease immediately after each administration of i.v. IL-2 and HuM195. CD56-expressing cells increased in 6 of 10 patients from the beginning to the end of therapy. Among the 10 evaluable patients, 2 patients had significant decreases in the percentage of blasts in the bone marrow (one of which achieved a complete bone marrow remission), 5 patients had stable levels of bone marrow blasts, and 3 had progression of disease on therapy. The combination of IL-2 and HuM195 shows modest biological activity and clinical antileukemic activity but also produced significant toxicity. PMID- 10537339 TI - Phase I trial of a MART-1 peptide vaccine with incomplete Freund's adjuvant for resected high-risk melanoma. AB - Twenty-five patients with high-risk resected stages IIB, III, and IV melanoma were immunized with a vaccine consisting of the minimal epitope, immunodominant 9 amino acid peptide derived from the MART-1 tumor antigen (AAGIGILTV) complexed with incomplete Freund's adjuvant. The last three patients received the MART-1(27 35) peptide with incomplete Freund's adjuvant mixed with CRL 1005, a block copolymer adjuvant. Patients were immunized with increasing doses of the MART 1(27-35) peptide in a Phase I trial to evaluate the toxicity, tolerability, and immune responses to the vaccine. Immunizations were administered every 3 weeks for a total of four injections, preceded by leukapheresis to obtain peripheral blood mononuclear cells for immune analyses, followed by a post-vaccine leukapheresis 3 weeks after the fourth vaccination. Skin testing with peptide and standard delayed-type hypersensitivity skin test reagents was also performed before and after vaccinations. Local pain and granuloma formation were observed in the majority of patients, as were fevers or lethargy of grade 1 or 2. No vaccine-related grade III/IV toxicity was observed. The vaccine was felt to be well tolerated. Twelve of 25 patients were anergic to skin testing at the initiation of the trial, and 13 of 25 developed a positive skin test response to the MART-1(27-35) peptide. Immune responses were measured by release of IFN-gamma in an ELISA assay by effector cells after multiple restimulations of peripheral blood mononuclear cells in the presence of MART-1(27-35) peptide-pulsed antigen presenting cells. An ELISPOT assay was also developed to measure more quantitatively the change in numbers of peptide-specific effector cells after vaccination. Ten of 22 patients demonstrated an immune response to peptide-pulsed targets or tumor cells by ELISA assay after vaccination, as did 12 of 20 patients by ELISPOT. Nine of 25 patients have relapsed with a median of 16 months of follow-up, and 3 patients in this high-risk group have died. Immune response by ELISA correlated with prolonged relapse-free survival. These data suggest a significant proportion of patients with resected melanoma mount an antigen specific immune response against a peptide vaccine and support further development of peptide vaccines for melanoma. PMID- 10537340 TI - Phase I study of direct intralesional gene transfer of HLA-B7 into metastatic renal carcinoma lesions. AB - Renal cancer cell lines exhibit deficient expression of MHC class I antigens required for appropriate CTL stimulation. Nabel et al. (Proc. Natl. Acad. Sci. USA, 90: 11307-11311, 1993) demonstrated that direct gene transfer of the deficient class I MHC molecule into melanoma cells would stimulate their immune destruction. Stopeck et al. (J. Clin. Oncol., 15: 341-349, 1997) demonstrated the clinical use of this approach in melanoma patients. We investigated the safety and ability to bestow gene expression via intratumoral transfer of escalating amounts of lipid-formulated plasmid DNA encoding for the MHC HLA-B7 gene product Allovectin-7 into metastatic renal cancer lesions. Fifteen patients with histologically confirmed, HLA-B7-negative metastatic renal cancer received intratumoral injection of Allovectin-7 on an escalating dose schedule. Tumors were evaluated serially by computed tomography scan, ultrasound, and physical examination. Presence of the HLA-B7 gene and protein was determined via PCR, flow cytometry, and immunohistochemical staining in serial biopsy specimens. HLA-B7 gene, mRNA, or protein expression could be conclusively demonstrated in 8 of 14 patients (57%). Three patients had tumor biopsy to assess the presence of tumor infiltrating lymphocytes, and all three had higher posttreatment levels of tumor infiltrating lymphocytes. There were no significant clinical responses or toxicity at the site of injection or at other, noninjected tumor sites. This study demonstrated that intratumoral injection of Allovectin-7 is safe, feasible, and associated with minimal toxicity. This approach was capable of bestowing gene expression, possible resulting in antitumor CTL response. Despite lack of tumor regression in this series of renal cancer patients, the simplicity and low toxicity of this approach commend it for Phase II studies in renal and other cancers, as well as for transfection with other genes. PMID- 10537341 TI - Immunogenicity of a fucosyl-GM1-keyhole limpet hemocyanin conjugate vaccine in patients with small cell lung cancer. AB - Although small cell lung cancer (SCLC) is highly responsive to chemotherapy, relapses are common, and most patients die within 2 years of diagnosis. After initial therapy, standard treatment is observation alone. We have been investigating immunization against selected gangliosides as adjuvant therapy directed against residual and presumably resistant disease persisting after chemotherapy and irradiation. Previously, we reported that the presence of anti GM2 ganglioside antibodies is associated with a prolonged disease-free survival in patients with melanoma, and that SCLC patients immunized with BEC2, an anti idiotypic monoclonal antibody that mimics the ganglioside GD3, had a prolonged survival compared with historical controls. In the present trial, fucosyl-alpha1 2Galbeta1-3GalNAcbeta1-4(NeuAcalpha2-3) Galbeta1-4Glcbeta1-1Cer (Fuc-GM1), a ganglioside expressed on the SCLC cell surface, was selected as a target for active immunotherapy. Fuc-GM1 is present on most SCLCs but on few normal tissues. SCLC patients achieving a major response to initial therapy were vaccinated s.c. on weeks 1, 2, 3, 4, 8, and 16 with Fuc-GM1 (30 microg) conjugated to the carrier protein keyhole limpet hemocyanin and mixed with the adjuvant QS-21. Ten patients received at least five vaccinations and are evaluable for response. All patients demonstrated a serological response, with induction of both IgM and IgG antibodies against Fuc-GM1, despite prior treatment with chemotherapy with or without radiation. Posttreatment flow cytometry demonstrated binding of antibodies from patients' sera to tumor cells expressing Fuc-GM1. In the majority of cases, sera were also capable of complement-mediated cytotoxicity. Mild transient erythema and induration at injection sites were the only consistent toxicities. The Fuc-GM1-KLH + QS-21 vaccine is safe and immunogenic in patients with SCLC. Continued study of this and other ganglioside vaccines is ongoing. PMID- 10537342 TI - Interferon-gamma and CXC chemokine induction by interleukin 12 in renal cell carcinoma. AB - Interleukin 12 (IL-12) is known to play an important role in the development of an antitumor response. Its activity has been shown to be dependent upon the intermediate production of IFN-gamma and the influx into the tumor of CD8 lymphocytes. In a murine model, tumor regression induced by IL-12 treatment correlated with IFN-gamma, IP-10, and Mig expression in the tumor bed and was abrogated by antibodies to both chemokines. Here we examined the effects of rHuIL 12 on IFN-gamma and CXC chemokine gene expression in patients with renal cell carcinoma (RCC) in an attempt to determine whether a similar series of molecular events leading to IL-12-mediated tumor regression in mice is also detectable in humans. As in the murine RENCA model, cultured RCC cells themselves could be induced by IFN-gamma to synthesize IP-10 and Mig mRNA. Explanted RCC produced IFN gamma and IP-10 mRNA in response to IL-12 treatment, which was consistent with the finding that biopsied RCC tumors from IL-12-treated patients also variably expressed augmented levels of those molecules after therapy. Although Mig mRNA was present in the majority of biopsied tumors prior to treatment, both the Mig and IP-10 chemokines as well as IFN-gamma were induced in the peripheral blood mononuclear cells of IL-12-treated patients. Skin biopsies of IL-12-treated patients also all synthesized IP-10 mRNA. This study demonstrates that recombinant human IL-12 therapy of patients with RCC has the potential to induce the expression of gene products within the tumor bed that may contribute to the development of a successful antitumor response. PMID- 10537343 TI - Comparison of telomerase and CD44 expression as diagnostic tumor markers in lesions of the thyroid. AB - The analysis of the tissue expression patterns of both the telomerase enzyme and the adhesion molecule CD44 has highlighted these molecules as potential tumor markers. In this study, the expression of these markers was analyzed in frozen tissue samples of the same human thyroid lesions, and the data were compared to evaluate their application to tumor diagnosis. The study analyzed 12 malignant specimens, including 5 papillary, 3 follicular, 2 anaplastic, 1 medullary, and 1 low-grade Hurthle cell carcinoma and 17 specimens from benign lesions, including cases of adenoma, hyperplasia, and Graves' disease. Telomerase expression was analyzed by assay of enzyme activity using the telomeric repeat amplification protocol and by reverse transcription-PCR detection of human telomerase reverse transcriptase (hTERT) mRNA. Nine of 12 (75%) malignant samples and the two Graves' disease samples were evaluated as positive for telomerase activity by the telomeric repeat amplification protocol assay. The presence of hTERT mRNA was detected in 8 (67%) of 12 malignant tissues and in 5 (29%) of 17 benign thyroid tissue samples. The expression of CD44 transcripts containing variant exons 7, 8, and 11 was evaluated by reverse transcription-PCR/Southern blot analysis. Of the 12 malignant samples, 9 (75%) included transcripts containing exon 7, 10 (83%) included transcripts containing exon 11, and 11 (92%) included transcripts containing exon 8. However, these CD44 exons were also present in transcripts in a high proportion of benign samples. Five (28%), 10 (59%), and 6 (35%) benign samples contained CD44 transcripts, including variant exons 7, 8, and 11, respectively. The measurement of telomerase activity proved to be the most specific for the detection of thyroid carcinoma in frozen tissue samples as a single analyte, but diagnostic accuracy was increased by the combination of telomerase and CD44 analyses. PMID- 10537344 TI - Drug resistance-associated markers P-glycoprotein, multidrug resistance associated protein 1, multidrug resistance-associated protein 2, and lung resistance protein as prognostic factors in ovarian carcinoma. AB - Intrinsic and/or acquired resistance to chemotherapy is the major obstacle to overcome in the treatment of patients with ovarian carcinoma. The aim of the present study was to investigate the prognostic value of drug resistance associated proteins P-glycoprotein (P-gp), multidrug resistance-associated protein 1 (MRP1), canalicular multispecific organic anion transporter (c MOAT/MRP2), and lung resistance protein (LRP) in ovarian carcinoma. Expression of P-gp, MRP1, MRP2, and LRP was determined by immunohistochemistry of frozen tissue sections of 115 ovarian carcinoma patients and related to clinicopathological factors, response to chemotherapy, and progression-free survival. P-gp expression was observed in 20 of 115 (17%), MRP1 in 51 (44%), MRP2 in 19 (16%), and LRP in 85 (74%) tumors. Expression of MRP1 was related to MRP2 (P<0.0001) and P-gp (P<0.001) expression, whereas LRP expression was more frequently observed in patients with early stage (P<0.01), lower grade (P<0.05), and smaller residual tumor (P<0.05). Early stage (P<0.001), smaller residual tumor (P<0.001), and lower differentiation grade (P<0.05) were related to longer (progression-free) survival. P-gp, MRP1, MRP2, and LRP expression were neither related to response to first-line chemotherapy in 59 evaluable patients nor to progression-free survival in all patients. On multivariate analysis, only stage and residual tumor were independent prognostic factors for survival. In conclusion, in ovarian carcinoma, MRP1 expression is associated with MRP2 and P-gp expression, whereas LRP expression is associated with favorable clinicopathological characteristics. Assessment of P-gp, MRP1, MRP2, or LRP does not allow prediction of response to chemotherapy or survival in ovarian carcinoma. PMID- 10537345 TI - Serum concentrations of vascular endothelial growth factor in vulvar cancer. AB - The aim of the present study was to evaluate serum concentrations of vascular endothelial growth factor (VEGF) in patients with vulvar cancer and healthy female controls with respect to correlation of VEGF with clinicopathological parameters and impact on the patients' prognosis. Serum concentrations of VEGF were measured using a commercially available ELISA. Results were correlated to clinical data. Median serum concentrations of VEGF in patients with vulvar cancer (n = 41) and healthy female controls (n = 130) were 260 (range, 33-1216) pg/ml and 216 (range, 0-777) pg/ml, respectively (Mann-Whitney U test, P = 0.048). Serum concentrations of VEGF significantly correlated with tumor stage (Mann Whitney U test, P = 0.02) but not with histological grade (Mann-Whitney U test, P = 0.2). In a univariate analysis, elevated pretreatment serum concentrations of VEGF were significantly correlated with a shortened disease-free and overall survival (Wilcoxon test, P = 0.03; and Wilcoxon test, P = 0.04, respectively). A multivariate Cox regression model considering tumor stage and serum concentrations of VEGF revealed, however, that serum concentrations of VEGF did not confer additional prognostic information to that already obtained by the established prognosticator tumor stage (multivariate Cox regression model: P = 0.9 and P = 0.8, respectively). Our data indicate that angiogenesis, as reflected by serum concentrations of VEGF, plays a functional role in vulvar carcinogenesis. VEGF seems to be a mediator of vulvar tumor growth but not of tumor cell dedifferentiation. Although associated with impaired disease-free and overall survival, pretreatment serum concentrations of VEGF are not an independent predictor of outcome in patients with vulvar cancer. PMID- 10537346 TI - Cyclin D1 and p16INK4A expression predict reduced survival in carcinoma of the anterior tongue. AB - Cyclin D1 and p16INK4A are molecules with pivotal roles in cell cycle control and the development of diverse human cancers, and overexpression of cyclin D1 and loss of p16INK4A expression are common genetic events in head and neck squamous cell carcinoma. The prognostic significance of these molecular events at different sites within the head and neck, however, remains controversial. Thus, we sought to determine the relationship between cyclin D1 and/or p16INK4A expression and disease outcome in squamous cell carcinoma of the anterior tongue. Immunohistochemical detection of nuclear proteins cyclin D1, p53, and p16INK4A, and the Ki-67 labeling index was undertaken in tissue sections from 148 tongue cancers treated by surgical resection. Nuclear antigen status was analyzed in relation to pathological variables, tumor recurrence, and patient survival. Statistical significance was assessed using chi2 analysis for pathological variables and the Kaplan-Meier method, log rank test, and the Cox proportional hazards model for survival parameters. Overexpression of cyclin D1 occurred in 68% of tumors (100 of 147) and was associated with increased lymph node stage (P = 0.014), increased tumor grade (P = 0.003), and reduced disease-free (P = 0.006) and overall (P = 0.01) survival. Loss of p16INK4A expression was demonstrated in 55% of tumors (78 of 143) and was associated with reduced disease-free (P = 0.007) and overall (P = 0.014) survival. Multivariate analysis confirmed that in addition to pathological stage and regional lymph node status, cyclin D1 overexpression and loss of p16INK4A expression are independent predictors of death from tongue cancer. Loss of p16INK4A in the presence of cyclin D1 overexpression conferred a significantly worse disease-free (P = 0.011) and overall (P = 0.002) survival at 5 years. p53 nuclear accumulation and the Ki-67 labeling index were not prognostic. These data indicate that cyclin D1 overexpression and loss of p16INK4A expression predict early relapse and reduced survival in squamous cell carcinoma of the anterior tongue. Simultaneous assessment of cyclin D1 and p16INK4A protein levels define subgroups of patients at increased risk of relapse and may be of clinical utility in optimizing therapy. PMID- 10537347 TI - Cell proliferation in prostate cancer patients with lymph node metastasis: a marker for progression. AB - The biological aggressiveness of lymph node-positive prostate cancer is closely linked to cancer volume in nodal metastases. We evaluated MIB-1 (Ki-67) labeling index and bcl-2 expression in primary cancer and matched nodal metastases from 138 node-positive patients treated with radical prostatectomy and bilateral pelvic lymphadenectomy between 1987 and 1992 at the Mayo Clinic. One hundred twenty-eight patients (93%) received androgen deprivation therapy within 90 days after radical prostatectomy. Mean patient age was 66 years (range, 51-78). The median follow-up was 6.7 years (range, 0.03-11). MIB-1 (Ki-67) labeling index was determined by digital image analysis, and nodal cancer volume was determined by the grid method. Systemic progression, defined as the presence of distant metastasis documented by biopsy or radiographic examination, was used as an outcome end point in the Cox proportional hazard models. MIB-1 labeling index in nodal metastases was predictive of systemic progression-free survival (P = 0.001). The 8-year systemic progression-free survival was 100% for those with MIB 1 labeling index <3.5% compared with 78% for those with MIB-1 labeling index > or =7.8%. MIB-1 labeling index correlated with Gleason score, DNA ploidy, and nodal cancer volume (P<0.001, 0.04, and <0.001, respectively). After controlling for nodal cancer volume, MIB-1 labeling index remained significant in predicting systemic progression-free survival (P = 0.047). bcl-2 expression in the primary cancer and lymph node metastasis was associated with systemic progression-free survival in univariate analysis (P = 0.027 and 0.048, respectively) but was not significant after adjusting for nodal cancer volume (P = 0.52 and 0.17, respectively). Our data indicate that assessment of cell proliferation in nodal metastasis is predictive of clinical outcome in prostate cancer patients with regional lymph node metastasis. PMID- 10537348 TI - Relative expression of matrix metalloproteinase-2 and tissue inhibitor of metalloproteinase-2 in mouse renal cell carcinoma cells regulates their metastatic potential. AB - To clarify the significance of the balance between matrix metalloproteinase-2 (MMP-2) and tissue inhibitor of metalloproteinase-2 (TIMP-2) in the progression of renal cell carcinoma, we transfected both the MMP-2 and TIMP-2 genes simultaneously into RenCa, a mouse renal cell carcinoma cell line that does not express detectable levels of either MMP-2 or TIMP-2 mRNAs, and established several clones with various MMP-2:TIMP-2 expression ratios. On the basis of the quantitative evaluation of the MMP-2: TIMP-2 mRNA expression ratio by Northern blot analysis, we selected a clone overexpressing MMP-2 alone (RenCa/M), a clone overexpressing TIMP-2 alone (RenCa/T), and two kinds of clones overexpressing both, i.e., one with a high (RenCa/MTh) and one with a low (RenCa/MTl) MMP-2: TIMP-2 ratio, to compare the tumor cell phenotypes. In an in vitro tumor cell invasion assay, the MMP-2:TIMP-2 ratios of the RenCa sublines were directly correlated with their invasive potential. The invasive abilities of the parental RenCa cells induced by conditioned media from RenCa sublines were also correlated with the MMP-2:TIMP-2 ratios of the sublines. The cell adhesion assay showed the inverse correlation between the MMP-2 expression levels in the sublines and their cell adhesion to several extracellular matrix components. Furthermore, when injected i.v. or into the renal subcapsule in syngeneic mice, RenCa sublines formed metastatic nodules in the lungs, and the number of nodules was correlated with the MMP-2:TIMP-2 ratio of each clone. In contrast, despite the growth inhibitory effects of TIMP-2 overexpression, MMP-2 overexpression had no effect on either proliferation in vitro of RenCa sublines or on their growth as tumors in vivo. These results suggest that the MMP-2:TIMP-2 expression ratio is a critical factor in the invasion and metastasis of renal cell carcinoma. PMID- 10537349 TI - Prognostic relevance of p53 alterations and Mib-1 proliferation index in subgroups of primary liposarcomas. AB - For prognostic analyses of p53 alterations (p53 gene mutations + p53 immunopositivity) and Mib-1 proliferation index, we investigated 42 primary malignant lipomatous tumors for which complete clinical data and a long follow-up were available. p53 gene mutations were investigated by PCR-single strand conformation polymorphism-sequencing analysis, and immunohistochemistry was used to determine p53 protein expression and Mib-1 proliferation index. We found a mutation frequency of 14.3%. Nine liposarcomas (21%) were p53 immunopositive, and 11 (26.2%) had at least one p53 alteration. In myxoid liposarcomas, p53 alterations are not relevant to the presence or absence of round cell components. Pleomorphic liposarcomas showed a significantly higher proliferation index and more p53 alterations than myxoid or well-differentiated variants (P<0.001). When the Cox's regression analysis tumors of grade III histology (P = 0.005) was performed, the pleomorphic subtype (P = 0.016) and liposarcomas of retroperitoneal localization (P = 0.015) showed a significantly poorer prognosis. Moreover, we found that p53 alterations and high proliferation index correlated significantly with reduced overall survival. Their prognostic value seemed to be higher in myxoid than in pleomorphic liposarcomas. The metastasis-free survival was reduced in patients who had liposarcomas with p53 alterations (P = 0.171) or elevated proliferation index (P<0.016), reflecting a more aggressive behavior. In conclusion, the determination of p53 alterations and/or Mib-1 proliferation index is useful for assessing the prognosis of patients with liposarcomas and may especially be helpful in dividing different prognostic groups for patients with myxoid variants. PMID- 10537350 TI - Relationship between intratumoral dihydropyrimidine dehydrogenase activity and gene expression in human colorectal cancer. AB - Dihydropyrimidine dehydrogenase (DPD) is the rate-limiting enzyme for 5 fluorouracil catabolism. In this study, both the enzymatic activity and mRNA level of DPD were estimated in the tumor tissue and adjacent normal mucosa of 51 patients with colorectal cancer. Although no significant difference in enzymatic activity was observed between tumor tissue and normal mucosa (70.4 and 70.7 pmol/min/mg protein, respectively), the mRNA level in normal mucosa was significantly higher than that in tumor tissue (1.37 and 0.39, respectively; P<0.01). A linear relationship was noted between DPD activity and the DPD mRNA level in cancerous tissue (r(s) = 0.714, P<0.001). Thus, the DPD mRNA level as determined by reverse transcription-PCR can be used to indicate the DPD activity of colorectal cancers. PMID- 10537351 TI - Characterization of cytokeratin 20 expression in pancreatic and colorectal cancer. AB - Cytokeratin 20 belongs to the epithelial subgroup of the intermediate filament family. Because of its restricted range of expression in humans, it has become an important tool for detecting and identifying metastatic cancer cells by immunohistochemistry and by PCR analysis. Despite its widespread diagnostic use in colorectal cancer and occasional use in pancreatic cancer, little is known about the expression of CK 20 in these tumors in vivo. Therefore, in the present study we characterized CK 20 expression in pancreatic and colorectal cancer by comparison with its expression in the normal pancreas and colon. Tissue samples from 24 patients with pancreatic cancer and from 41 patients with colorectal cancer were examined for CK 20 expression by Northern blot analysis, immunohistochemistry, and in situ hybridization. CK 20 expression was observed in the cancer cells of both cancer types. A subgroup of the pancreatic cancers exhibited a 3.2-fold increase in CK 20 mRNA by comparison with respective normal controls. In contrast, colon cancers underexpressed CK 20 mRNA by comparison with the respective controls. In the normal tissues, CK 20 immunoreactivity was relatively faint and sparse in the pancreatic ductal cells but intense and abundant in the apical portions of the colonic mucosa. CK 20 immunoreactivity was also evident in the ductal cells from the chronic pancreatitis-like lesions adjacent to the cancer cells. Furthermore, distant metastases from pancreas carcinomas exhibited strong CK 20 immunoreactivity. It is concluded that CK 20 is overexpressed in pancreatic cancer and that it can serve as an excellent marker for metastatic pancreatic cancer. PMID- 10537352 TI - Lack of independent prognostic significance of p21 and p27 expression in advanced ovarian cancer: an immunohistochemical study. AB - The eukaryotic cell cycle is controlled by protein complexes consisting of cyclin dependent kinases and cyclins. The cyclin-dependent kinases are in turn negatively regulated by a family of cyclin-dependent kinase inhibitors, comprising, among others, the p21 and p27 proteins. p21 and p27 have been shown to be of prognostic significance in a broad array of human tumors. Using immunohistochemistry, the frequency of expression and the possible prognostic and predictive significance of these proteins were examined in a series of 185 uniformly treated patients with stage III ovarian cancer. We found p21 to be overexpressed in 48% of cases. No significant correlation was found between the expression of p21 and p53 proteins (P = 0.273). A low level of p27 was demonstrated in 48.5% of cases. p21 overexpression correlated with lower Federation Internationale des Gynaecologistes et Obstetristes substage, lower patient age, and absence of ascites, but neither p21 nor p27 expression was of prognostic significance for the whole group of patients. Only a trend toward reduced survival (P = 0.092) was noticed for the small subgroup of patients (6%), whose tumors lacked p27 expression completely. A clear positive correlation could be found between p21 and p27 protein expression (P = 0.012). Despite the suggested role of the 21 and p27 proteins in determining drug sensitivity, they were not found to be predictive for response to chemotherapy, as assessed by second-look laparotomy in this large group of patients with advanced ovarian cancer. PMID- 10537353 TI - Measurement of neovascularization is an independent prognosticator of survival in node-negative breast cancer patients with long-term follow-up. AB - We measured neovascularization, epidermal growth factor receptor, and c-erbB-2 expression in a consecutive series of 233 surgically resected axillary lymph node negative breast cancer patients with a long-term follow-up to define the usefulness of these parameters as independent prognostic indicators of overall survival (OAS). Microvessel count (MVC), as a measure of neovascularization, was determined using a monoclonal antibody against human factor VIII-related antigen. The median MVC of 20 (range, 4-76) was used as a cutoff value for discriminating between low and high vascularized tumors. Epidermal growth factor receptor and c erbB-2 expression were evaluated by immunohistochemistry. Tumors were considered positive if >10% of the cells showed specific membrane staining. OAS curves were estimated by the Kaplan-Meier method. The independent prognostic effect of each variable was determined with the Cox proportional hazards model. High MVC (P = 0.04), high nuclear grade (P = 0.005), and high S-phase (P = 0.02) significantly affected OAS at univariate analysis. In a Cox multivariate analysis, the characteristics with an independent prognostic effect on OAS were: MVC (relative hazard, 2.12; 95% confidence interval, 1.18-3.81; P = 0.01) and nuclear grade (relative hazard, 2.83; 95% confidence interval, 1.12-7.17; P = 0.01). These results demonstrate that quantification of neovascularization adds useful independent prognostic information on survival in node-negative breast cancer patients with long-term follow-up. PMID- 10537354 TI - Bcl-2, Bcl-X, Bax, and Bak expression in short- and long-lived patients with diffuse large B-cell lymphomas. AB - Long-term cure is now possible for approximately 50% of all patients with aggressive non-Hodgkin's lymphoma (NHL). Apoptosis-related proteins play an important role in the chemosensitivity or chemoresistance of tumors. We examined the role of Bcl-2 family proteins in aggressive NHL. We retrospectively selected two groups of patients by clinical outcome: 24 patients with chemoresponsive disease and long survival (median, 88 months); and 20 patients with chemoresistant disease and short survival (median, 8 months). The expression of the apoptosis-regulating proteins, Bcl-2, Bcl-X, Bax, and Bak, in the initial biopsy samples was examined with immunohistochemical methods. Specimens containing >10% immunostained tumor cells were considered immunopositive. An inverse association was found between length of patient survival and expression of Bcl-2, Bcl-X, and Bax. Bcl-2 was expressed in 75% of short-lived patients but in only 42% of the long-lived ones (P = 0.026). Bcl-X expression was also higher in the short-lived patients (40% versus 12.5%; P = 0.036). Unexpectedly, Bax expression was strongly associated with short survival (60% versus 21%; P = 0.008). Several combinations of protein expression, i.e., Bcl-2 with Bax, Bcl-2 with Bcl-X, and Bcl-X with Bax, were different between the groups: a positive expression of these proteins was found in the short-lived patients. Furthermore, a strong association was found between the expression of Bcl-2 and Bcl-X, suggesting that Bcl-X potentiates rather than replaces the effect of Bcl-2 in NHL. In diffuse large B-cell NHL, Bcl-2, Bcl-X, and Bax expression alone or in combination is associated with chemoresistance and shortterm survival. PMID- 10537355 TI - Angiogenic growth factors in preinvasive breast disease. AB - Recently, we showed that preinvasive breast pathologies, such as usual hyperplasia, atypical hyperplasia, and carcinoma in situ, have an increased vascularity when compared with normal breast tissue (S. C. Heffelfinger et al., Clinical Cancer Res., 2: 1873-1878, 1996). To understand the mechanism of this increased vascularity, we examined by immunohistochemistry each of these pathological lesions for the expression of angiogenic growth factors. These studies showed that normal breast tissue contains numerous angiogenic agents, particularly vascular endothelial cell growth factor and basic fibroblast growth factor. At the transition from normal epithelium to proliferative breast disease, insulin-like growth factor (IGF) II expression was increased, primarily in the stroma and infiltrating leukocytes. However, among proliferative tissues, IGF I decreased with increasing vascularity. Finally, both epithelial vascular endothelial growth factor and epithelial and leukocytic platelet-derived endothelial cell growth factor increased at the transition to carcinoma in situ, whereas stromal and leukocytic basic fibroblast growth factor were elevated only in invasive carcinoma. Therefore, during histological progression there is also a complex progression of angiogenic growth factors. For CIS, two forms of vascularity are found: stromal microvascular density (MVD), and vascularity associated with the epithelial basement membrane (vascular score). There was 35% discordance between these two measurement systems. Among carcinoma in situ cases, decreases in stromal IGF II were associated with increasing vascular scores but not MVD, and increases in platelet-derived endothelial cell growth factor were associated with increasing MVD but not the vascular score. The presence of discordance and differential association with specific angiogenic agents suggests that these two forms of vascularity may be differentially regulated. PMID- 10537356 TI - Expression of the c-erbB-3/HER-3 and c-erbB-4/HER-4 growth factor receptors and their ligands, neuregulin-1 alpha, neuregulin-1 beta, and betacellulin, in normal endometrium and endometrial cancer. AB - The objective of this study was to determine the immunohistochemical expression of the c-erbB-3 and c-erbB-4 growth factor receptors and their principal ligands, the neuregulins and betacellulin, in normal endometrium and determine whether there was evidence of under- or overexpression in endometrial adenocarcinoma. Immunohistochemistry was performed using well-characterized antibodies against each of the five proteins analyzed on formalin-fixed, paraffin-embedded archival material. Forty-three normal endometrial samples (16 proliferative, 19 secretory, and 8 hyperplastic) and 41 endometrial adenocarcinoma cases were analyzed. There was variable expression of the growth factor receptors and the ligands in the two principal phases of the menstrual cycle as well as in endometrial adenocarcinoma. In normal endometrium, the c-erbB-3 receptor was weakly expressed in both phases. The c-erbB-4 receptor and all of the ligands examined, neuregulin alpha, neuregulin beta, and betacellulin, were expressed at significantly higher levels in the secretory as compared with the proliferative phase of the menstrual cycle, suggesting a role for these proteins in endometrial maturation. In endometrial adenocarcinoma, overexpression of c-erbB-3, c-erbB-4, and betacellulin with underexpression of neuregulin a as compared with normal controls was observed. Neuregulin beta expression was not found to be significantly different in the two groups. These results suggest that signaling through the c-erbB-3 and c-erbB-4 receptors and the ligands neuregulin alpha, neuregulin beta, and betacellulin are important in endometrial carcinogenesis. PMID- 10537357 TI - Inverse relationship between epidermal growth factor receptor expression and radiocurability of murine carcinomas. AB - The study investigated whether a relationship exists between the extent of epidermal growth factor receptor (EGFR) expression and in vivo radiocurability of murine tumors. EGFR expression was determined in nine carcinomas (four mammary carcinomas, designated MCa-4, MCa-29, MCa-35, and MCa-K; two squamous cell carcinomas, designated SCC-IV and SCC-VII; an ovarian adenocarcinoma, OCa-I; a hepatocarcinoma, HCa-I; and an adenosquamous carcinoma, ACa-SG) syngeneic to C3Hf/Kam mice using Western blot analysis. These tumors greatly differed in their radioresponse, assessed by TCD50 assay, and in their susceptibility to radiation induced apoptosis. Likewise, the expression of EGFR greatly varied, by as much as 21-fold, and the magnitude of the EGFR expression positively correlated with increased tumor radioresistance. The levels of EGFR inversely correlated with radiation-induced apoptosis, suggesting that the lack of sensitivity to apoptosis induction was a major mechanism responsible for radioresistance of tumors with high EGFR. This correlation was highly significant only for wild-type p53 carcinomas. Radiation activated EGFR autophosphorylation and increased the activity of protein tyrosine kinase, but only in tumors with high EGFR expression. Thus, EGFR expression was a major determinant of tumor radioresponse in vivo. The pretreatment assessment of EGFR expression could predict radiotherapy outcome and may assist in selecting an effective treatment modality. PMID- 10537358 TI - Progression to androgen independence is delayed by adjuvant treatment with antisense Bcl-2 oligodeoxynucleotides after castration in the LNCaP prostate tumor model. AB - Bcl-2 has emerged as a critical regulator of apoptosis in a variety of cell systems and is up-regulated during progression to androgen independence in prostate cancer cells. The objectives of this study were to characterize changes in Bcl-2 after androgen withdrawal and during progression to androgen independence in the human prostate LNCaP tumor model and determine whether adjuvant use of antisense Bcl-2 oligodeoxynucleotides (ODNs) with androgen ablation delays progression to androgen independence. Bcl-2 expression in LNCaP cells is down-regulated to undetectable levels by androgen in vitro and up regulated after castration in vivo. Antisense Bcl-2 ODN treatment reduced LNCaP cell Bcl-2 messenger RNA and protein levels by >90% in a sequence-specific and dose-dependent manner at concentrations >50 nM. Bcl-2 mRNA levels returned to pretreatment levels by 48 h after discontinuing treatment. Athymic male mice bearing SQ LNCaP tumors were castrated and injected i.p. with 12.5 mg/kg/day with two-base mismatch ODN control, reverse polarity ODN control, or antisense Bcl-2 ODN. Tumor volume in control mice gradually increased 5-fold (range, 3-6) by 12 weeks after castration compared to a 10-50% decrease in precastrate tumor volume in mice treated with antisense Bcl-2 ODN. Changes in serum PSA paralleled changes in tumor volume, increasing 4-fold faster above nadir in controls than in mice treated with antisense Bcl-2 ODN. After decreasing 70% by 1 week after castration, PSA increased 1.6-fold above precastrate levels by 11 weeks in controls while staying 30% below precastrate levels in antisense-treated mice. In a second group of experiments, LNCaP tumor growth and serum PSA levels were 90% lower (P<0.01) in mice treated with antisense Bcl-2 ODN compared with mismatch or reverse polarity ODN controls. These results support the hypothesis that Bcl-2 helps mediate progression to androgen independence and is an appropriate target for antisense therapy. PMID- 10537359 TI - Inhibitors of topoisomerase II as pH-dependent modulators of etoposide-mediated cytotoxicity. AB - Chloroquine intercalates into DNA and protects cells against topoisomerase II (topo II) poisons such as etoposide by hindering the DNA cleavage reaction of this target enzyme. Chloroquine, in contrast to etoposide, is a weak base and therefore barely enters the cell when the extracellular fluid is acidic, as is the case in most solid tumors. Such a pH-dependent drug interaction could be useful in targeting the cytotoxicity of topo II poisons toward solid tumors. Unfortunately, antagonistic chloroquine concentrations cannot be reached in vivo because of its unacceptable toxicity. Thus, antagonists with a higher therapeutic index are needed. We report here on the structure-activity relationship of several chloroquine and acridine analogues in a clonogenic assay. There were major differences in the cytotoxicity of the different compounds, with acridines being 50-fold more toxic than the chloroquine analogues. Several compounds were, however, able to antagonize etoposide-mediated cytotoxicity in a pH-dependent manner as chloroquine. Dependency on pH was lost if the aminoalkyl side arm of chloroquine was removed or lengthened by one CH2 whereas pH dependency was strong with hydroxychloroquine. In contrast, the aminoalkyl side arm was clearly dispensable in the acridines because both quinacrine and 9-aminoacridine demonstrated profound pH dependency. The results from clonogenic assay were compared with cellular transport measurements and topo II enzyme inhibition. Compounds with the most marked pH-dependent intracellular accumulation were also the best pH-dependent protectors of etoposide cytotoxicity, clearly supporting the hypothesis that extracellular pH can be used to regulate topo II poisoning. PMID- 10537360 TI - Pharmacologic disruption of base excision repair sensitizes mismatch repair deficient and -proficient colon cancer cells to methylating agents. AB - Previously we showed that a mismatch repair (MMR)-deficient cell line, HCT116 (hMLH1 mut), unlike a MMR wild-type cell line, SW480, was more resistant to the therapeutic methylating agent, temozolomide (TMZ), because the MMR complex fails to recognize TMZ-induced O6-methylguanine DNA adduct mispairings with thymine that arise after replication. TMZ also produces N7-methylguanine and N3 methyladenine adducts that are processed efficiently by the base excision repair (BER) system. After removal of the methylated base by methylpurine glycosylase, which creates the abasic or apurinic-apyrimidinic (AP) site, the phosphodiester bond is hydrolyzed immediately by AP endonuclease, initiating the repair of the AP site. Methoxyamine (MX) reacts with the abasic site and prevents AP endonuclease cleavage, disrupting DNA repair. MX potentiated the cytotoxic effect of TMZ with a dose modification factor (DMF) of 2.3+/-0.12 in SW480 and 3.1+/ 0.16 in HCT116. When combined with O6-benzylguanine (BG), MX and TMZ dramatically increased TMZ cytotoxicity (65.8-fold) in SW480, whereas no additive effect was seen in HCT116. This suggests that N7-methylguanine and N3-methyladenine adducts are cytotoxic lesions in MMR-deficient and wild-type cells when BER is interrupted. Because poly(ADP-ribose) polymerase (PARP) aids in processing of DNA strand breaks induced during MMR and BER, we asked whether PARP inhibitors would also affect BER-mediated cell killing. We found that PARP inhibitors PD128763, 3 aminobenzimide, and 6-aminonicotinamide increased the sensitivity to TMZ in both HCT116 MMR-deficient cells and SW480 MMR wild-type cells. In HCT116 cells, PD128763 remarkably decreased resistance to TMZ, with a DMF of 4.7+/-0.2. However, the combination of PD128763, BG, and TMZ had no greater effect, indicating that persistent O6-methylguanine had no effect on cytotoxicity. In SW480, the DMF for TMZ cytotoxicity was 3.1+/-0.12 with addition of PD128763 and 36 with addition of PD128763 and BG. Synergy analysis by median effect plots indicated a high degree of synergy between TMZ and MX or PD128763. In contrast, 1,3-bis(2-chloroethyl)-1-nitrosourea combined with either MX or PD128763 showed little if any potentiation observed in the absence of BG in either cell line, suggesting that BER pathway has little impact on cytotoxic processing of 1,3 bis(2-chloroethyl)-1-nitrosourea-induced adducts. These studies indicate that targeting BER with MX or PARP inhibitors enhances the cytotoxicity of methylating agents, even in MMR-deficient cells. PMID- 10537361 TI - Rapid esterase-sensitive breakdown of polysorbate 80 and its impact on the plasma pharmacokinetics of docetaxel and metabolites in mice. AB - We have developed and validated an analytical methodology for the quantification of docetaxel and its four major human oxidation metabolites in mouse plasma. We have used this procedure to study the pharmacokinetics and metabolism of docetaxel in female FVB mice, receiving 2.5, 10, or 33 mg/kg of docetaxel by i.v. injection. We have also studied the pharmacokinetics of polysorbate 80, because it was shown previously that the vehicle substance Cremophor EL, which is used in the formulation of paclitaxel, exerts a profound effect on the pharmacokinetics of this compound. Linear pharmacokinetics of docetaxel was observed at dose levels between 2.5 and 10 mg/kg, where plasma levels corresponded to those in patients receiving the maximum tolerated dose. At the highest dose level of 33 mg/kg, a deviation from the linear kinetics was observed. Compared with humans, mice could tolerate much higher plasma levels, suggesting that the toxic side effects are related to a certain plasma threshold concentration instead of area under the curve or Cmax. At the highest dose level, three docetaxel metabolites could be detected in the plasma samples of mice for up to 4 h after drug administration. The hydroxy metabolite of the tert-butoxy group (metabolite II) was the major metabolite, followed by the two epimeric hydroxyoxazolone-type compounds (metabolites I and III). A fourth putative metabolite (e.g., the cyclic oxazolidinedione derivative) was not detected. Because of rapid degradation of polysorbate 80 by esterases in plasma, the concentration of this vehicle substance declined very rapidly. Consequently, this substance was not able to interfere in the disposition of docetaxel. PMID- 10537362 TI - Flavopiridol induces cell cycle arrest and p53-independent apoptosis in non-small cell lung cancer cell lines. AB - Flavopiridol, a synthetic flavone that inhibits tumor growth in vitro and in vivo, is a potent cyclin-dependent kinase (cdk) inhibitor presently in clinical trials. In the present study, the effect of 100-500 nM flavopiridol on a panel of non-small cell lung cancer cell lines was examined. All express a wild-type retinoblastoma susceptibility protein and lack p16INK4A, and only A549 cells are known to express wild-type p53. During 72 h of treatment, flavopiridol was shown to be cytotoxic to all seven cell lines, as measured by trypan blue exclusion, regardless of whether cells were actively cycling. In most cycling cells, cytotoxicity was preceded or accompanied by cell cycle arrest. Cell death resulted in the appearance of cells with a sub-G1 DNA content, suggestive of apoptosis, which was confirmed by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay and by demonstration of cleavage of caspase targets including poly(ADP-ribose) polymerase, p21Waf1, and p27Kip1. At doses at or below 500 nM, maximal cytotoxicity required 72 h of exposure. Although flavopiridol resulted in the accumulation of p53 in A549 cells, flavopiridol-mediated apoptosis was p53 independent because it occurred to the same degree in A549 cells in which p53 was targeted for degradation by HPV16E6 expression. The data indicate that flavopiridol has activity against non-small cell lung cancers in vitro and is worthy of continued clinical development in the treatment of this disease. PMID- 10537364 TI - X-ray irradiation induces thymidine phosphorylase and enhances the efficacy of capecitabine (Xeloda) in human cancer xenografts. AB - Thymidine phosphorylase (dThdPase) is an essential enzyme for the activation of the cytostatics capecitabine (N4-pentyloxycarbonyl-5'-deoxy-5-fluorocytidine) and its intermediate metabolite 5'-deoxy-5-fluorouridine (5'-dFUrd) to 5-fluorouracil (5-FUra) in tumors. We observed previously that several cytokines and cytostatics up-regulated dThdPase expression and consequently enhanced the efficacy of capecitabine and 5'-dFUrd. In the present study, we found that X-ray irradiation also up-regulated dThdPase expression in several human cancer xenografts. A single-dose local irradiation at 5 Gy increased dThdPase levels by up to 13-fold at 9 days after the irradiation. Whole-body irradiation also up-regulated dThdPase in a tumor, but it did not increase the enzyme level in the liver. We also observed that the irradiation increased the levels of human tumor necrosis factor alpha (TNF-alpha), which is an up-regulator of dThdPase, prior to the dThdPase up-regulation. These results indicate that X-ray irradiation might increase dThdPase levels indirectly through the human TNF-alpha in the tumor tissue. In the WiDr colon and MX-1 mammary human cancer xenograft models, the combination of a single local X-ray irradiation with either capecitabine or 5' dFUrd was much more effective than either radiation or chemotherapy alone. In contrast, treatment with X-ray irradiation and 5-FUra in combination showed no clear additive effects. Combined modality treatment of cancer patients with cape citabine and X-ray irradiation would have greater potential usefulness than conventional radiochemotherapy with 5-FUra. PMID- 10537363 TI - Doxorubicin encapsulated in sterically stabilized liposomes exhibits renal and biliary clearance properties that are independent of valspodar (PSC 833) under conditions that significantly inhibit nonencapsulated drug excretion. AB - Coadministration of anticancer drugs and multidrug resistance modulators directed against P-glycoprotein over-expressed in tumors also results in nonspecific blockade of this drug efflux pump in excretory tissues such as the liver and kidneys. These interactions often result in impaired renal and biliary clearance for anticancer agents such as doxorubicin (DOX). In the present investigation, we characterized the excretory processes associated with liposomal DOX administration to elucidate how liposome encapsulation may bypass adverse pharmacokinetic interactions between DOX and (3'-keto-Bmt1)-(Val2)-cyclosporin (Valspodar). Renal and biliary clearance properties of liposome-encapsulated DOX were compared with those for nonencapsulated DOX in the presence and absence of Valspodar using an instrumented rat model with implanted jugular vein and bile duct catheters for continuous sampling. Two types of liposomal DOX formulations were used, a drug-permeable egg phosphatidyl choline/cholesterol system and a sterically stabilized polyethylene glycol/1,2 distearoyl-sn-glycero-3 phosphocholine/cholesterol system to establish the relative roles of liposome encapsulated and released drug on the pharmacokinetic and excretion alterations induced by Valspodar. DOX and its primary metabolites were quantitated using high performance liquid chromatography. When Valspodar was coadministered with nonencapsulated DOX, 3.5- and 37.5-fold reductions in renal clearance (CLr) and biliary clearance (CLb), respectively, were observed, which resulted in increased plasma DOX concentrations and total exposure. However, Valspodar-induced alterations in CLr and CLb were less profound with egg phosphatidyl choline/cholesterol DOX (1.7- and 2.0-fold reductions, respectively) and negligible with the long-circulating polyethylene glycol-containing liposomal formulation. These results indicate that liposomes may circumvent Valspodar induced DOX pharmacokinetic changes by reducing the rate of drug excretion in liver and kidney tissue to a level that is within the renal and biliary excretion capacity in the presence of P-glycoprotein blockade. PMID- 10537365 TI - In vivo toxicity and pharmacokinetic features of the janus kinase 3 inhibitor WHI P131 [4-(4'hydroxyphenyl)-amino-6,7- dimethoxyquinazoline. AB - 4-(4'Hydroxyphenyl)-amino-6,7-dimethoxyquinazoline (WHI-P131) is a potent and selective inhibitor of the Janus kinase 3, which triggers apoptosis in human acute lymphoblastic leukemia (ALL) cells. In this preclinical study, we evaluated the pharmacokinetics and toxicity of WHI-P131 in rats, mice, and cynomolgus monkeys. Following i.v. administration, the terminal elimination half-life of WHI P131 was 73.2 min in rats, 103.4 min in mice, and 45.0 min in monkeys. The i.v. administered WHI-P131 showed a very wide tissue distribution in mice. Following i.p. administration, WHI-P131 was rapidly absorbed in both rats and mice, and the time to reach the maximum plasma concentration (tmax) was 24.8 min in rats and 10.0 min in mice. Subsequently, WHI-P131 was eliminated with a terminal elimination half-life of 51.8 min in rats and 123.6 min in mice. The estimated i.p. bioavailability was 95% for rats, as well as for mice. WHI-P131 was quickly absorbed after oral administration in mice with a tmax of 5.8 min, but its oral bioavailability was relatively low (29.6%). The elimination half-life of WHI-P131 after oral administration was 297.6 min. WHI-P131 was not acutely toxic to mice at single i.p. bolus doses ranging from 0.5-250 mg/kg. Two cynomolgus monkeys treated with 20 mg/kg WHI-P131 and one cynomolgus monkey treated with 100 mg/kg WHI-P131 experienced no side effects. Plasma samples from WHI-P131-treated monkeys exhibited potent antileukemic activity against human ALL cells in vitro. To our knowledge, this is the first preclinical toxicity and pharmacokinetic study of a Janus kinase 3 inhibitor. Further development of WHI-P131 may provide the basis for new and effective treatment programs for relapsed ALL in clinical settings. PMID- 10537366 TI - Antibodies to vascular endothelial growth factor enhance the efficacy of cancer immunotherapy by improving endogenous dendritic cell function. AB - Inadequate function of dendritic cells (DCs) in tumor-bearing hosts is one mechanism of tumor escape from immune system control and may compromise the efficacy of cancer immunotherapy. Vascular endothelial growth factor (VEGF), produced by most tumors, not only plays an important role in tumor angiogenesis but also can inhibit the maturation of DCs from hematopoietic progenitors. Here, we investigate a novel combination of antiangiogenic and immunotherapy based on this dual role of VEGF. Two s.c. mouse tumor models were used: D459 cells, expressing mutant human p53; and MethA sarcoma with point mutations in the endogenous murine p53 gene. Therapy with anti-mouse VEGF antibody (10 microg i.p. twice a week over 4 weeks) was initiated when tumors became palpable. Treatment of established tumors with anti-VEGF antibody alone did not affect the rate of tumor growth. However, anti-VEGF antibody significantly improved the number and function of lymph node and spleen DCs in these tumor-bearing animals. To investigate the possible effects of this antibody on the immunotherapy of established tumors, tumor-bearing mice were immunized with DCs pulsed with the corresponding mutation-specific p53 peptides, together with injections of anti VEGF antibody. Therapy with peptide-pulsed DCs alone resulted in considerable slowing of tumor growth but only during the period of treatment, and tumor growth resumed after the end of the therapy. Combined treatment with peptide-pulsed DCs and anti-VEGF antibody resulted in a prolonged and much more pronounced antitumor effect. This effect was associated with the induction of significant anti-p53 CTL responses only in this group of mice. These data suggest that inhibition of VEGF may be a valuable adjuvant in the immunotherapy of cancer. PMID- 10537367 TI - Tamoxifen-induced apoptosis in breast cancer cells relates to down-regulation of bcl-2, but not bax and bcl-X(L), without alteration of p53 protein levels. AB - Tamoxifen (TAM) has been shown to induce apoptosis in breast cancer cells. bcl-2 family genes, which can interact with each other, have been shown to interfere with apoptosis after various stimuli. In this study, we investigated the effects of TAM on bcl-2 family gene products bcl-2, bax, and bcl-X(L) and on p53 levels in estrogen receptor-positive MCF-7 breast cancer cells. We found that TAM induced time- and concentration-dependent down-regulation of bcl-2 at both the mRNA and protein level. Down-regulation of bcl-2 correlated with TAM-induced apoptosis. In addition, estradiol treatment significantly increased bcl-2 protein expression and blocked the reduction of bcl-2 by TAM. TAM did not, however, affect bax, bcl-X(L), or p53 expression at the mRNA or protein level. Our results demonstrate that TAM can induce apoptosis in a time- and dose-dependent manner by modulating bcl-2 levels in breast cancer cells, and down-regulation of bcl-2 induced by TAM was not accompanied by alterations in p53 levels. PMID- 10537368 TI - Correspondence re: P. Salven et al., leukocytes and platelets of patients with cancer contain high levels of vascular endothelial growth factor. Clin. Cancer Res., 5: 487-91, 1999. PMID- 10537369 TI - Isobutane. CAS# 75-28-5. PMID- 10537370 TI - 2-Methylbutane (isopentane). CAS# 78-78-4. PMID- 10537371 TI - n-Pentane. CAS# 109-66-0. PMID- 10537372 TI - Cyclopentane. CAS#287-92-3. PMID- 10537373 TI - Neopentane (2,2-dimethylpropane). CAS# 463-82-1. PMID- 10537374 TI - 2-Methylpentane (isohexane). CAS# 107-83-5. PMID- 10537375 TI - 3-Methylpentane. CAS# 96-14-0. PMID- 10537376 TI - 2,2-Dimethylbutane (neohexane). CAS 75-83-2. PMID- 10537377 TI - 2,3-Dimethylbutane. CAS# 79-29-8. PMID- 10537378 TI - Treatment of right ventricular infarction. PMID- 10537379 TI - Dyspepsia: relief not yet beyond belief. PMID- 10537380 TI - Herbal remedies. PMID- 10537381 TI - Herbal remedies. PMID- 10537382 TI - Herbal remedies. PMID- 10537383 TI - Waddell signs in the evaluation of back pain. PMID- 10537384 TI - Anterior hip pain. AB - Anterior hip pain is a common complaint with many possible causes. Apophyseal avulsion and slipped capital femoral epiphysis should not be overlooked in adolescents. Muscle and tendon strains are common in adults. Subsequent to accurate diagnosis, strains should improve with rest and directed conservative treatment. Osteoarthritis, which is diagnosed radiographically, generally occurs in middle-aged and older adults. Arthritis in younger adults should prompt consideration of an inflammatory cause. A possible femoral neck stress fracture should be evaluated urgently to prevent the potentially significant complications associated with displacement. Patients with osteitis pubis should be educated about the natural history of the condition and should undergo physical therapy to correct abnormal pelvic mechanics. "Sports hernias," nerve entrapments and labral pathologic conditions should be considered in athletic adults with characteristic presentations and chronic symptoms. Surgical intervention may allow resumption of pain-free athletic activity. PMID- 10537385 TI - Pneumocystis carinii pneumonia: a clinical review. AB - Pneumocystis carinii pneumonia (PCP) is an opportunistic infection that occurs in immunosuppressed populations, primarily patients with advanced human immunodeficiency virus infection. The classic presentation of nonproductive cough, shortness of breath, fever, bilateral interstitial infiltrates and hypoxemia does not always appear. Diagnostic methods of choice include sputum induction and bronchoalveolar lavage. The drug of choice for treatment and prophylaxis is trimethoprim-sulfamethoxazole, but alternatives are often needed because of adverse effects or, less commonly, treatment failure. Adjunctive corticosteroid therapy improves survival in moderate to severe cases. Complications such as pneumothorax and respiratory failure portend poorer survival. Prophylaxis dramatically lowers the risk of disease in susceptible populations. Although PCP has declined in incidence in the developed world as a result of prophylaxis and effective antiretroviral therapy, its diagnosis and treatment remain challenging. PMID- 10537386 TI - Drug treatment of common STDs: Part II. Vaginal infections, pelvic inflammatory disease and genital warts. AB - The Centers for Disease Control and Prevention (CDC) released new guidelines for the treatment of sexually transmitted diseases (STDs) in 1998. Several treatment advances have been made since the previous guidelines were published. Part II of this two-part series on STDs describes recommendations for the treatment of diseases characterized by vaginal discharge, pelvic inflammatory disease, epididymitis, human papillomavirus infection, proctitis, proctocolitis, enteritis and ectoparasitic diseases. Single-dose therapies are recommended for the treatment of several of these diseases. A single 1-g dose of oral azithromycin is as effective as a seven-day course of oral doxycycline, 100 mg twice a day, for the treatment of chlamydial infection. Erythromycin and ofloxacin are alternative agents. Four single-dose therapies are now recommended for the management of uncomplicated gonococcal infections, including 400 mg of cefixime, 500 mg of ciprofloxacin, 125 mg of ceftriaxone or 400 mg of ofloxacin. Advances in the treatment of bacterial vaginosis also have been made. A seven-day course of oral metronidazole is still recommended for the treatment of bacterial vaginosis in pregnant women, but intravaginal clindamycin cream and metronidazole gel are now recommended in nonpregnant women. Single-dose therapy with 150 mg of oral fluconazole is a recommended treatment for vulvovaginal candidiasis. Two new topical treatments, podofilox and imiquimod, are available for patient self administration to treat human papillomavirus infection. Permethrin cream is now the preferred agent for the treatment of pediculosis pubis and scabies. PMID- 10537387 TI - Right ventricular infarction: specific requirements of management. AB - The principal cause of right ventricular infarction is atherosclerotic proximal occlusion of the right coronary artery. Proximal occlusion of this artery leads to electrocardiographically identifiable right-heart ischemia and an increased risk of death in the presence of acute inferior infarction. Clinical recognition begins with the ventricular electrocardiographic manifestations: inferior left ventricular ischemia (ST segment elevation in leads II, III and aVF), with or without accompanying abnormal Q waves and right ventricular ischemia (ST segment elevation in right chest leads V3R through V6R and ST segment depression in anterior leads V2 through V4). Associated findings may include atrial infarction (PR segment displacement, elevation or depression in leads II, III and aVF), symptomatic sinus bradycardia, atrioventricular node block and atrial fibrillation. Hemodynamic effects of right ventricular dysfunction may include failure of the right ventricle to pump sufficient blood through the pulmonary circuit to the left ventricle, with consequent systemic hypotension. Management is directed toward recognition of right ventricular infarction, reperfusion, volume loading, rate and rhythm control, and inotropic support. PMID- 10537388 TI - In pursuit of perfection: a primary care physician's guide to body dysmorphic disorder. AB - Body dysmorphic disorder is an under-recognized chronic problem that is defined as an excessive preoccupation with an imagined or a minor defect of a localized facial feature or body part, resulting in decreased social, academic and occupational functioning. Patients who have body dysmorphic disorder are preoccupied with an ideal body image and view themselves as ugly or misshapen. Comorbid psychiatric disorders may also be present in these patients. Body dysmorphic disorder is distinguished from eating disorders such as anorexia nervosa that encompass a preoccupation with overall body shape and weight. Psychosocial and neurochemical factors, specifically serotonin dysfunction, are postulated etiologies. Treatment approaches include cognitive-behavioral psychotherapy and psychotropic medication. To relieve the symptoms of body dysmorphic disorder, selective serotonin reuptake inhibitors, in higher dosages than those typically recommended for other psychiatric disorders, may be necessary. A trusting relationship between the patient and the family physician may encourage compliance with medical treatment and bridge the transition to psychiatric intervention. PMID- 10537389 TI - Pediatric advanced life support: a review of the AHA recommendations. American Heart Association. AB - The etiologies of respiratory failure, shock, cardiopulmonary arrest and dysrhythmias in children differ from those in adults. In 1988, the American Heart Association implemented the pediatric advanced life support (PALS) program. Major revisions to the program were made in 1994, with further revisions in 1997. The PALS program teaches a systematic, organized approach for the evaluation and management of acutely ill or injured children. Early identification and treatment of respiratory failure and shock in children improve survival, from a dismal 10 percent to an encouraging 85 percent. Family physicians who care for acutely ill or injured children have a tremendous opportunity to save lives through implementation of the PALS information. PMID- 10537390 TI - Diagnosis and treatment of endometriosis. AB - Endometriosis is a progressive disease affecting 5 to 10 percent of women. It can cause dyspareunia, dysmenorrhea, low back pain and infertility. A definitive diagnosis can be made only by means of laparoscopy. Medical treatment designed to interfere with ovulation generally provides effective pain relief, but the recurrence rate following cessation of therapy is high, and this type of treatment will not resolve infertility. Surgical treatment improves pregnancy rates and is the preferred initial treatment for infertility caused by endometriosis. Surgery also appears to provide better long-term pain relief than medical treatment. Bilateral oophorectomy and hysterectomy are treatment options for patients with intractable pain, if childbearing is no longer desired. PMID- 10537391 TI - Evaluation and management of dyspepsia. AB - Dyspepsia, often defined as chronic or recurrent discomfort centered in the upper abdomen, can be caused by a variety of conditions. Common etiologies include peptic ulcers and gastroesophageal reflux. Serious causes, such as gastric and pancreatic cancers, are rare but must also be considered. Symptoms of possible causes often overlap, which can make initial diagnosis difficult. In many patients, a definite cause is never established. The initial evaluation of patients with dyspepsia includes a thorough history and physical examination, with special attention given to elements that suggest the presence of serious disease. Endoscopy should be performed promptly in patients who have "alarm symptoms" such as melena or anorexia. Optimal management remains controversial in young patients who do not have alarm symptoms. Although management should be individualized, a cost-effective initial approach is to test for Helicobacter pylori and treat the infection if the test is positive. If the H. pylori test is negative, empiric therapy with a gastric acid suppressant or prokinetic agent is recommended. If symptoms persist or recur after six to eight weeks of empiric therapy, endoscopy should be performed. PMID- 10537392 TI - AHA and ACC issue scientific statement on preventive cardiology for women. American Heart Association and American College of Cardiology. PMID- 10537393 TI - Advisory committee on immunization practices updates recommendations for the prevention of varicella. PMID- 10537394 TI - HMOs and vicarious liability. Implying control over physicians may expose HMOs. PMID- 10537395 TI - Spoliation of medical records: negligent, reckless, and intentional destruction of evidence. PMID- 10537396 TI - A balancing act. How to juggle pharmacy costs and quality care. PMID- 10537398 TI - Rewriting the rules. Prospective payment has hammered some and helped others, but nearly everyone's glad to be through the first tough year. PMID- 10537397 TI - The art of the MDS. Interview by Janet Grady Sullivan. PMID- 10537399 TI - Making it work. Five providers share their strategies for surviving under PPS. PMID- 10537400 TI - It's the money, honey. Even more than respect, CNAs need to earn a living wage. PMID- 10537401 TI - Are high rehab RUG scores really for you? PMID- 10537402 TI - The selling never stops. It takes a systematic game plan to fill a new community. PMID- 10537403 TI - When bad things happen to good providers. PMID- 10537404 TI - High acuity can mean high risk of pressure ulcers. PMID- 10537405 TI - A day in the life of a CNA. PMID- 10537406 TI - The voice of experience. Interview by Bridget DeMouy. PMID- 10537407 TI - Walk this way. Getting residents out of wheelchairs and back on their own two feet. PMID- 10537408 TI - Fiduciary duties 101. Directors and officers have never been at greater risk. PMID- 10537409 TI - Building a network of stayers. Fix your staffing woes by giving good CNAs reason to stay. PMID- 10537411 TI - The cream of the crop. Meet the 1998 EMS Gold Award winners. PMID- 10537410 TI - Novel funding for ALFs (assisted living facilities). Interview by Elise Nakhnikian. PMID- 10537412 TI - Life's little safety signs. PMID- 10537413 TI - Keeping kids safe: injury prevention programs. PMID- 10537414 TI - The bugs are back in town! Serious infections in children. PMID- 10537415 TI - Prehospital management of the seizure patient. AB - Seizures are a frequent problem confronting EMS personnel. Most seizures will terminate by themselves in less than five minutes. Careful attention must be paid to the ABCs, including glucose. Seizures lasting more than five minutes should be treated with i.v. diazepam or lorazepam. Careful history-taking to look for possible causes of seizures, as well as a detailed description of the event, are invaluable to ED staff in the evaluation and treatment of a patient with a seizure. Patients with known epilepsy who have had a single typical seizure and are otherwise back to normal do not require transport to the hospital. PMID- 10537416 TI - Bedside manners. PMID- 10537417 TI - Twist one for the nipper. Balloon animals distract pediatric patients. PMID- 10537418 TI - How to defuse compliance time bombs. Six hypothetical allegations. Panel discussion. PMID- 10537420 TI - Calif. seeks controls on not-for-profit links. PMID- 10537419 TI - Shared governance. One lab's experience. PMID- 10537421 TI - AMA: Docs working less. Study shows physicians earn a little more, put in fewer hours. PMID- 10537422 TI - Multi-unit Providers Survey. For-profits report decline in acute-care hospitals ... newcomers to top 10. AB - For-profit hospital systems cleaned house last year. After years of adding hospitals, investor-owned operators shed facilities in 1998, recording the first decline in the number of acute-care hospitals they've owned or managed since 1991, according to our 23rd annual Multi-unit Providers Survey. PMID- 10537423 TI - AHA uses hospital raid in lobbying efforts. PMID- 10537424 TI - Multi-unit Providers Survey. Healthcare systems ranked by net patient revenues. PMID- 10537425 TI - Multi-unit Providers Survey. Psychiatric sector sees slight changes. PMID- 10537426 TI - Multi-unit Providers Survey. Medicare changes shake long-term care. Survey shows anticipated payment reductions prompted mergers, caused financial damage. PMID- 10537427 TI - Multi-unit Providers Survey. Respondents list. PMID- 10537428 TI - New player enters credit rating game. PMID- 10537429 TI - Primary care purchasing. Are integrated primary care provider/purchasers the way forward? AB - Integrated primary care provider/purchaser organisations are currently being developed in the UK and other countries. In common with many other European countries and New Zealand, there has been a swing away from market-orientated models of healthcare organisation towards models which blend centralised planning and quasi market approaches. Primary care groups in the UK and independent practice organisations in New Zealand provide new opportunities to combine micromanagement techniques in the delivery of patient care and approaches to improving population health. These new organisations are building on the experience of health maintenance organisations (HMOs) in the US and physician-led purchasing in the UK and other countries. However, there are distinct differences between countries due to differences in health and social care systems and the degree of emphasis on primary physician involvement. Moreover, the continuing emphasis on a physician-led medical model may be at odds with developing a public health approach which emphasises participative and collaboration with local populations and other primary care providers--an area where physicians have little experience. PMID- 10537430 TI - Pharmacoeconomic consequences of variable patient compliance with prescribed drug regimens. AB - Variable compliance with prescribed drug regimens is a leading source of variability in drug response. Specifics differ by drug and disease. The role of variable compliance was clearly defined in 2 trials of lipid-lowering agents, cholestyramine and gemfibrozil, in which exceptionally careful measurements of compliance were made, which has not been done in later trials. Economic consequences of variable compliance are estimated by converting dose-dependent changes in absolute risk of incident coronary disease into the unicohort format, which designates how many patients must be treated to prevent, in a given time, a defined 'coronary event'. Two strong influences on the costs of treatment are: (i) the shape of the relation between drug intake and risk reduction; and (ii) the strength of the linkage between intake and prescription refills. The intake effect relation for cholestyramine is linear, making compliance-neutral the cost to prevent 1 coronary event, provided that refills match intake. If refills exceed intake, treatment costs rise. The intake-effect relation for gemfibrozil is more typically nonlinear, so poorer compliers purchase and take the drug in amounts that have little benefit, increasing the cost to prevent 1 coronary event. If refills run at a higher rate than intake, costs increase still further. A key question for future study is: do policies that encourage timely refills increase compliance enough to offset their potential to waste money in the purchasing of an untaken drug? PMID- 10537431 TI - The economics of multiple sclerosis. Distribution of costs and relationship to disease severity. AB - The introduction of expensive disease-modifying agents for the treatment of multiple sclerosis (MS) has created the potential for patients with MS to become higher contributors to healthcare spending. In an attempt to make formulary and reimbursement choices for these agents, decision-makers may look to the literature for guidance. This critical review attempts to decipher a consistent message from the available economic literature regarding the relationship between disease severity and cost in MS. In the 2 studies that have examined MS disease severity, a positive correlation with total (direct and indirect) cost, indirect cost and some, if not all, components of direct cost was reported. In studies taking the societal perspective, the majority of total costs were indirect. This paper documents the high burden of MS on society and serves to guide the decision maker in interpreting the MS economic literature such that this information can be optimally utilised to make informed resource allocation decisions. PMID- 10537433 TI - Under-utilisation of beta-blockers after acute myocardial infarction. Pharmacoeconomic implications. AB - We reviewed the literature on the efficacy and effectiveness of beta-blocker therapy and examined the economic consequences of under-utilisation. Despite the literature documenting the value of beta-blockers, the therapy is not prescribed at the appropriate rates. Approximately half of acute myocardial infarction (AMI) survivors who are eligible for the therapy do not receive it. There are 3 sources of costs that may arise from such under-utilisation: (i) increased morbidity and mortality associated with under-use; (ii) increased demand for related medical resources when the health state following an AMI is suboptimal due to under-use of beta-blocker therapy; and (iii) increased cost due to substitution of higher cost and/or less effective treatments for beta-blockers. For the first category, there is evidence suggesting that around 2900 to 5000 lives are lost in the US in the first year following an AMI due to underprescription. There is very little evidence on the second category of costs; 1 recent study does address this issue and indicates that beta-blocker therapy can lead to a 22% relative risk reduction for hospital readmission during the first year. Several studies also show a decrease in reinfarction. There is no information that addresses the third category of costs adequately (though 1 study does present evidence of substitution of calcium channel-blockers for beta-blockers). We conclude that there is a dearth of evidence on the economic consequences of the under utilisation of beta-blocker therapy. What does exist suggests that the net costs to society may be substantial. PMID- 10537434 TI - New drugs for osteoporosis. Comparison of the costs and required returns with those of other drugs intended for long-term use. AB - Specific regulatory guidelines dictate that developing a new drug for osteoporosis will be significantly more expensive and take at least 2 years longer in comparison with other long-term therapies developed using the International Committee on Harmonisation (ICH) general guidelines. Assuming similar attrition rates, the minimal additional uncapitalised cost is $US86 million for nonestrogen osteoporosis compounds following a minimum programme designed to gain indications for both treatment and prevention. The excess expenditure is created by the size requirements for phase III fracture trials in both the European Union (EU)/US and Japan. The after-tax cash flows to the point of launch discounted at 11% are $US102 million greater, reflecting the additional effect of delayed time to market. Assuming similar lifecycles, the peak sales required to return the investment on an osteoporosis drug will be a minimum of $US95 million greater per launched compound. Many ongoing osteoporosis programmes are substantially larger than the theoretical minimum. The costs of substantially increasing the sample size in phase III trials mean that blockbuster revenues will be required to break even. However, the potential cost of a delayed launch because of fracture efficacy being incompletely proved is so substantial that fracture trials need to be powered conservatively to decrease the chances of this eventuality. PMID- 10537432 TI - Patient self-reports in pharmacoeconomic studies. Their use and impact on study validity. AB - The validity of estimates of resource utilisation based on patient recall has not been firmly established. A comprehensive literature search was conducted to identify studies that used patient self-reports to derive estimates of healthcare resource utilisation, direct nonmedical costs and indirect costs. Previous work in this area was examined to determine the issues that were most important in determining whether patient self-reports lead to valid and unbiased estimates in cost-effectiveness studies. This study reviews and highlights areas where patient self-reports lead to reliable estimates and where their use may lead to erroneous conclusions. In particular, it is noted that patient recall of resource utilisation declines over time, that the salience of a treatment episode affects recall and that the perceived social acceptability of a condition and other confounding factors may influence patient reporting. Moreover, it appears that not all elements of healthcare consumption are recalled to the same degree: medication use tends to be recalled with less accuracy than hospitalisation. In terms of the potential impact on cost-effectiveness results, the main concern is with problems of validity rather than bias, although bias may occur when the results are applied to league tables. PMID- 10537435 TI - Simvastatin after orthotopic heart transplantation. Costs and consequences. AB - OBJECTIVE: Recent data indicate that the combination of a low cholesterol diet and simvastatin following heart transplantation is associated with significant reduction of serum cholesterol levels, lower incidence of graft vessel disease (GVD) and significantly superior 4-year survival rates than dietary treatment alone. On the basis of this first randomised long term study evaluating survival as the clinical end-point, we investigated the cost effectiveness of the above regimens as well as the long term consequences for the patient and for heart transplantation as a high-tech procedure. DESIGN AND SETTING: The perspective of the economic analysis was that of the German health insurance fund. Life-years gained were calculated on the basis of the Kaplan-Meier survival curves from the 4-year clinical trial and from the International Society for Heart and Lung Transplantation (ISHLT) overall survival statistics. Incremental costs and incremental cost-effectiveness ratios were determined using various sources of data, and both costs and consequences were discounted by 3% per year. Sensitivity analyses using alternative assumptions were conducted in addition to the base case analysis. PATIENTS AND PARTICIPANTS: As in the original clinical trial, the target population of the economic evaluation comprised all heart transplant recipients on standard triple immunosuppression consisting of cyclosporin, azathioprine and prednisolone, regardless of the postoperative serum lipid profile. INTERVENTIONS: The therapeutic regimens investigated in the analysis were the American Heart Association (AHA) step II diet plus simvastatin (titrated to a maximum dosage of 20 mg/day) and AHA step II diet alone. MAIN OUTCOME MEASURES AND RESULTS: Four years of treatment with simvastatin (mean dosage 8.11 mg/day) translated into an undiscounted survival benefit per patient of 2.27 life years; 0.64 life-years within the trial period and 1.63 life-years thereafter. Discounted costs per year of life gained were $US1050 (sensitivity analyses $US800 to $US15,400) for simvastatin plus diet versus diet alone and $US18,010 (sensitivity analyses $US17,130 to $US21,090) for heart transplantation plus simvastatin versus no transplantation (all costs reflect 1997 values; $US1 = 1.747 Deutschmarks). CONCLUSIONS: Prevention of GVD with simvastatin after heart transplantation was cost effective in all the scenarios examined with impressive prolongation of life expectancy for the heart recipient. Simvastatin also achieved an internationally robust 21% improvement in the cost effectiveness of heart transplantation compared with historical cost-effectiveness data. PMID- 10537436 TI - Pharmaceutical price regulation. A study on the impact of the rate-of-return regulation in the UK. AB - OBJECTIVE: This work carries out an empirical evaluation of the impact of the main mechanism for regulating the prices of medicines in the UK [the Pharmaceutical Price Regulation Scheme (PPRS)] on a variety of pharmaceutical price indices. The article also discusses to what extent the rate-of-return (ROR) regulation has encouraged UK-based pharmaceutical firms with patented products to diversify into markets in which products face strong competition. DESIGN AND SETTING: The article starts with some background on the PPRS and the way firms behave under ROR constraints. The article goes on to explain the cointegration methods used and the results obtained. Finally, it offers some discussion and some conclusions related to the evidence and the incentives of UK pharmaceutical firms under the PPRS constraint. MAIN OUTCOME MEASURES AND RESULTS: The results obtained show that, according to only some cointegration tests carried out, the aggregate price indices of medical preparations and the price index of some therapeutic areas are cointegrated with the time series of ROR caps between 1980 and 1994. Additionally, a 1% change in the ROR cap has produced only a 0.15% change on the aggregate medicine price index. CONCLUSIONS: These results suggest that changes in the ROR cap have had little or no impact on medicine prices and that, at best, the impact of the ROR has also differed significantly across major therapeutic areas. Finally, it is argues that the UK regulation of prices might have encouraged firms to diversify into competitive medicine-regulated markets and into uncontrolled markets. PMID- 10537437 TI - Costs of peripheral blood progenitor cell transplantation using whole blood mobilised by filgrastim as compared with autologous bone marrow transplantation in non-Hodgkin's lymphoma. AB - OBJECTIVE: The aim of this paper is to compare the costs of autologous bone marrow transplantation (ABMT) and whole blood transplantation in patients with relapsed or poorly responding non-Hodgkin's lymphoma. DESIGN AND SETTING: In a retrospective study, we calculated the treatment costs of 40 patients who received either ABMT or, alternatively, whole blood mobilised by filgrastim (a granulocyte colony-stimulating factor. MAIN OUTCOME MEASURES AND RESULTS: The recovery of granulocytes was markedly accelerated in the whole blood group as compared with the ABMT group. This resulted in a reduction in hospital costs, and costs for diagnostics and medical procedures, antibacterials, nutrition and blood transfusions. The average costs per patient in the whole blood group amounted to approximately $US16,890 as compared with approximately $US20,713 in the ABMT group (1995 values), implying a cost reduction of 18% when changing to whole blood reinfusion. CONCLUSIONS: With the premise that both therapies are equivalent, it seems that whole blood transplantation is more cost effective than ABMT. PMID- 10537438 TI - Health economics in HIV disease. A review of the European literature. AB - The costs of providing healthcare resources for patients with HIV disease have continued to rise during the last 2 decades since the first outbreak of AIDS. Although new and effective therapies have become available during this time, and are increasingly being used, outcome measures for, and the economic consequences of, differing models of care are poorly documented or understood. Most available economic and clinical data, especially data documenting post-therapy outcomes, are from small, short term studies. This has led to the use of modelling to estimate which determinants can improve the quality and cost effectiveness of HIV care in the future. Although the attempt to provide what is currently perceived to be the standard of care is stretching many healthcare budgets, it is an anachronism that there are few studies of health economics in HIV disease. Most published data relate only to North America, and there remains a paucity of European studies. This paper provides a review of the available data from Europe and attempts to highlight the trends that have taken place to date. PMID- 10537439 TI - Lamivudine reduces healthcare resource use when added to zidovudine-containing regimens in patients with HIV infection. AB - BACKGROUND: The impact on healthcare resource use of adding lamivudine to concurrent zidovudine-containing antiretroviral regimens was studied as a part of a 52-week multinational study [CAESAR (Canada, Australia, Europe and South Africa)] in HIV-infected patients with moderate to severe immunodeficiency (25 to 250 CD4+ cells/mm3). RESULTS: Significantly fewer lamivudine than placebo recipients required hospitalisations (p = 0.002), unscheduled outpatient visits (p = 0.013) or prescribed medications for HIV-related illness (p < 0.001). The mean number of hospitalisations and the mean duration of hospitalisation for HIV related illness were 47% and 51% lower, respectively, with lamivudine than with placebo. The mean number of unscheduled outpatient visits was 32% lower with lamivudine than with placebo. Lamivudine was also associated with a significant reduction in the number of patients who were hospitalised (p = 0.04) or required unscheduled outpatient visits (p = 0.02) as a result of adverse events. CONCLUSIONS: Notwithstanding the fact that retrospective studies have suggested that more effective antiretroviral treatments reduce healthcare use, the CAESAR study is one of the few prospective controlled trials to demonstrate that by slowing disease progression with combination therapy it is possible to reduce healthcare resource use in patients with HIV infection. Although the combination of lamivudine and zidovudine alone is not likely to be sufficient to achieve complete long term suppression of viral replication and to halt disease progression, the study demonstrates the immediate economic benefits of preventing HIV progression in HIV-infected patients with moderate to severe immunodeficiency (25 to 250 CD4+ cells/mm3). These findings suggest that treatment regimens that slow progression of HIV infection have the potential to produce savings in non drug healthcare costs, which may partly or fully offset the drug costs. PMID- 10537441 TI - A prospective evaluation of the cost effectiveness of adding lamivudine to zidovudine-containing antiretroviral treatment regimens in HIV infection. European perspective. AB - BACKGROUND: A prospective cost-effectiveness analysis undertaken as part of the CAESAR (Canada, Australia, Europe, South Africa) placebo-controlled clinical trial showed that adding lamivudine to zidovudine-containing regimens for 1 year reduced progression to AIDS or death and, in addition, significantly reduced the number of hospitalisations, unscheduled outpatient visits and the requirement for medications for HIV-related illness. Data from all 1840 patients included in the intent-to-treat population of the CAESAR trial were used in the analysis reported in this paper. Because a third-party payer perspective was adopted, possible savings associated with increased productivity (indirect costs) were not taken into account. All costs were adjusted to 1997 prices. RESULTS: The savings associated with reduced healthcare resource use in the CAESAR study were estimated to be 3045 Deutschmarks (DM) [German analysis] or 432 pounds sterling (Pound) [UK analysis] per patient for the 1-year time period. These savings partly offset the cost of lamivudine in the 2 countries. The German analysis showed that the addition of lamivudine to zidovudine-containing regimens resulted in an incremental cost-effectiveness ratio of DM22,405 [95% confidence interval (CI): -DM2199 to DM59,154] for progression to AIDS/death avoided and of DM8869 (95% CI: -DM1047 to DM23,365) for HIV-related illness avoided. The corresponding ratios for the UK analysis were 12,030 Pounds (95% CI: 6752 Pounds to 21,888 Pounds) for progressions avoided and 4762 Pounds (95% CI: 2796 Pounds to 9484 Pounds) for new and recurrent HIV-related illness avoided. CONCLUSIONS: Our findings indicate that treatments that slow the progression of HIV infection to AIDS or death have the potential to facilitate healthcare savings during the period that the treatment is effective. The results also demonstrate that it is possible to undertake economic evaluations in parallel with a major clinical end point study. PMID- 10537440 TI - A prospective cost-consequence analysis of adding lamivudine to zidovudine containing antiretroviral treatment regimens for HIV infection in the US. AB - BACKGROUND: Healthcare resource use data were collected for 1 year as part of the CAESAR (Canada, Australia, Europe, South Africa) clinical trial, which evaluated the effect of adding lamivudine to treatment regimens containing zidovudine in patients with HIV infection. This study showed that lamivudine-containing regimens reduced HIV disease progression to AIDS or death, in addition to significantly reducing the number of hospital stays, unscheduled outpatient visits, and medications for HIV-related illness. Estimates of US unit costs for each healthcare service were derived from nationally representative data sources, and were used to determine the costs of treatment during the trial period for the treatment and control groups. RESULTS: A cost-consequence analysis showed that, in addition to the health benefits associated with the lamivudine regimen, costs for treating HIV-related illness and adverse events were lower with the lamivudine regimen. The average decrease in costs per patient for the 1-year period ranged from $US1922 to $US2645, depending on the data source used to estimate hospital length of stay. The incremental cost of lamivudine therapy for the 1-year period was $US2293. The estimated difference in total costs for the 2 treatment regimens thus ranged from an increase of $US371 to a cost saving of $US353. CONCLUSIONS: Our findings indicate that treatments which slow the progression of HIV infection have the potential to reduce the monthly costs associated with HIV-related illness and adverse events during the time period that progression is slowed. PMID- 10537442 TI - An evaluation of the cost effectiveness of adding lamivudine to zidovudine containing regimens in HIV infection. Canadian perspective. AB - BACKGROUND: A prospective cost-effectiveness analysis undertaken as part of the CAESAR (Canada, Australia, Europe, South Africa) placebo-controlled clinical trial showed that the addition of lamivudine to zidovudine-containing regimens for 1 year reduced progression of HIV infection to AIDS or death, as well as significantly reducing the number of hospitalisations, outpatient visits and the requirement for medications for HIV-related illness and adverse events. Data from all 1840 patients included in the 'intent-to-treat' population of the CAESAR trial were used for the present analysis. A Canadian third-party payer perspective was adopted, and all costs were adjusted to 1997 prices. RESULTS: The savings associated with reduced healthcare resource use in the CAESAR study were estimated to be 1123 Canadian dollars ($Can) per patient, over the year. These savings partly offset the cost of lamivudine. The analysis showed that the addition of lamivudine to zidovudine-containing regimens resulted in an incremental cost-effectiveness ratio of $Can 14,225 [95% confidence interval (CI): $Can4383 to $Can29,577] for progression to AIDS/death avoided and of $Can5631 (95%CI: $Can2010 to $Can12,929) for HIV-related illness avoided. CONCLUSIONS: Our findings indicate that treatments that slow the progression of HIV infection to AIDS or death have the potential to facilitate healthcare savings, which partly offset the drug acquisition costs. The results also demonstrate that it is possible to undertake economic evaluations in parallel with a major clinical end-point study. PMID- 10537443 TI - Quality of life and treatment satisfaction after the addition of lamivudine or lamivudine plus loviride to zidovudine-containing regimens in treatment experienced patients with HIV infection. AB - BACKGROUND: Assessments of health-related quality of life and treatment satisfaction were conducted as part of a randomised, double-blind, placebo controlled 52-week trial conducted in Canada, Australia, Europe, and South Africa (CAESAR). The Medical Outcomes Study HIV Health Survey (MOS-HIV) was self administered during 3 scheduled clinic visits (baseline, week 28 and the end-of treatment/withdrawal visit). A single question was used at the end of treatment to assess patient satisfaction with study medications. METHODS: Patients were randomly allocated to receive placebo, lamivudine (150 mg twice daily) or lamivudine (150 mg twice daily) plus loviride (100 mg 3 times daily) in addition to their current treatment regimen, which could be either zidovudine monotherapy, or zidovudine in combination with didanosine or zalcitabine at standard dosages. RESULTS: Statistically significant differences across treatment groups were demonstrated for the Physical and Mental Health Summary scores, and for 5 of 10 MOS-HIV subscales (physical functioning, vitality, cognitive functioning, general health perceptions, social functioning). These differences favoured the lamivudine and lamivudine plus loviride groups over the placebo group (p < 0.05). No significant difference was found between the 3 treatment groups with regard to the percentages of patients who were satisfied with their study medication. CONCLUSION: The results suggest that, for treatment-experienced patients with HIV infection and CD4+ counts < 250 cells/mm3, the addition of lamivudine or lamivudine plus loviride to antiretroviral regimens containing zidovudine maintained patient-reported mental and physical health. PMID- 10537445 TI - M. Daniel Sloan on universal standards of measurement and performance improvement. Interview by Carole S. Guinane. PMID- 10537444 TI - Assessing the efficacy of a clinical pathway in the management of older patients hospitalized with congestive heart failure. AB - Congestive heart failure (CHF) is the most common cause for hospitalization in older patients, and the prevalence of this condition is expected to rise as the population ages. The high cost of care has resulted in an increased emphasis on cost-effective approaches in patient management. One way to achieve this is the use of clinical pathways. This article compares outcomes in a group of older hospitalized patients managed on a CHF pathway with those of a historical cohort managed in the traditional manner. The patients on the pathway had significant reductions in length of stay and cost of care as well as more effective delivery of processes of care. Mortality rates were unchanged, at 3.5%. However, readmission rates showed a significant increase, from 9.25% to 13.5%, for patients on the pathway. PMID- 10537447 TI - Esprit de corps: is work something you really believe in? AB - An employee's ability to fit into an organization's culture is a key factor in achieving excellence in the workplace. This article explores socialization, anthropological factors, and shared values and their relationship to culture and esprit de corps. Cultural performance improvement studies can provide valuable information to leaders when an organization embraces a visionary philosophy. The studies described in this article suggest that scientific methodologies should be used. PMID- 10537446 TI - Outcomes measurement and quality improvement in long-term care. AB - As healthcare providers proceed with redesigning patient care, forming centers of excellence, developing clinical pathways, focusing on best practices, and restructuring care services in their efforts to improve the structures and the processes of healthcare, outcomes research can help to determine the most effective patient care regimens. In a similar manner, outcomes are also assuming a higher priority for administrators of long-term care facilities. This article defines outcomes, discusses how they are being used in long-term care settings, presents commonly used outcomes, and describes how these outcomes can be measured and graphically displayed. PMID- 10537448 TI - Cascading data sets: putting the pieces together. AB - Just as quality programs have evolved into organization-wide performance improvement efforts, the quality professional's role has expanded, bringing new challenges and expectations. The quality services umbrella, an operational framework for a total systems quality process, helps organization leaders and quality professionals identify organizational functions that contribute to overall performance. This article describes the benefits of utilizing the quality services umbrella framework through five examples. Each example highlights different benefits of the model, such as identifying a system's quality issues, enhancing performance improvement efforts, sustaining improvements, and effecting cost savings. PMID- 10537449 TI - Development and implementation of clinical pathways for the management of four trauma diagnoses. AB - Clinical pathways are similar to the production algorithms developed by industry. They are being adapted for use in healthcare to reduce resource utilization, decrease variability, and control expenditures. At Boston Medical Center we identified four trauma diagnoses that we believed to be amenable to the design and implementation of clinical pathways: closed head injury, penetrating wound to the abdomen, penetrating wound to the chest, and penetrating wound to an extremity. Upon implementation of these pathways, appropriate nonoperative, single-system, short-stay trauma patients were enrolled in them. This article details the process by which the four diagnoses were identified and the pathways designed, implemented, and evaluated. Preliminary data demonstrate a significant decrease in resource utilization following implementation of the pathways, without an adverse impact on readmission rates, length of stay, or mortality. PMID- 10537450 TI - 'Reuse' reheats. PMID- 10537451 TI - Moving daze. Taking the kinks out of the case cart system. PMID- 10537452 TI - Sharps strategy. Start with the right tools. PMID- 10537453 TI - Flash forward. Make flash sterilization safe--and prove it! PMID- 10537454 TI - Safety costs. PMID- 10537455 TI - Staying alive ... and thriving, despite shrinking budgets. PMID- 10537456 TI - Powder-free, Louisiana. PMID- 10537457 TI - Stats. Who's afraid of Y2K. PMID- 10537458 TI - Using CQI to prevent the placebo effect from causing failed clinical trials. AB - One of the greatest obstacles to bringing drugs to market is one of the most complex--the human mind. Despite the billions invested in R&D, a fundamental issue in drug development is that most patients' responses are enhanced or affected by the "placebo effect." An ongoing methodology for reducing the impact of placebo effect should be built into the clinical study design at the outset based on a continuous quality improvement model. A CQI program that requires continuous evaluation of study participant data and rater training is a way to improve study performance and, therefore, ensure more valid studies and reliable outcomes. PMID- 10537459 TI - The clinical research triad: how can we ensure quality in out-sourced clinical trials? AB - The importance of quality within clinical trials cannot be underestimated. Built on the foundation of patient care where quality may simply be understood and expected, the business of conducting clinical trials must evolve to instill quality and ensure that quality is maintained. How that is accomplished within the drug development process is complicated by the relationship between the vendors--the sponsor, the contractor and the investigative site. This article will discuss the dynamics of the drug development triad from the perspective of the authors. Who are the players and what is quality from each of their perspectives? Communication among all parties is essential in order to ensure that quality is maintained. Unfortunately, even with optimal communication, if expectations and goals are not clearly defined, the results may be unsatisfactory. Vision and values of each player contributes to the success of the relationship and the quality of the service. PMID- 10537460 TI - Can investigator certification improve the quality of clinical research? AB - With costs and competitive pressures increasing, pharmaceutical sponsors and contract research organizations are examining the drug development process. Investigators who conduct clinical studies significantly impact research quality and, thereby, costs. This article reviews current investigator selection methods and their shortcomings. Using one organization's experience certifying clinical research coordinators and clinical research associates, the authors highlight certification as a means of assessing competency and discuss investigator certification as a potentially accurate indicator of competence for conducting research and as a predictor of inherent quality at the research site. PMID- 10537461 TI - Using information technology to improve the quality and efficiency of clinical trial research in academic medical centers. AB - In the area of clinical trial research, academic medical centers (AMCs) need to create additional capacity and improve performance on vital indicators to attract more studies, as they are currently losing their share to stand-alone research sites. Through the utilization of information technology, AMCs will be in a better position to fend off the competitive threats to their clinical research dollars. Most AMCs are in an enviable position to leverage the value of information technology because of the existing people, processes, and technologies that probably already exist throughout the AMC. The challenge, then, is to deploy these resources in a different manner to support clinical trial research. PMID- 10537462 TI - Interview with Gary L. Yingling, J.D. PMID- 10537463 TI - Continuous quality improvement in contract research organizations--the customer focus. AB - The challenge of quality improvement extends beyond traditional service delivery organizations. This is the first of a two-part series on the application of continuous quality improvement (CQI) to contract research organizations associated with the pharmaceutical and biotechnology industry. The challenges and processes of clinical trials research, and the role of CQI within that process, are presented. The importance of customer focus, which is a key element of CQI, is described here as the foundation of the CQI process among contract research organizations (CROs) and as a major contributing factor to their success in recent years. PMID- 10537464 TI - By the numbers. Examining the appropriateness of care. PMID- 10537465 TI - An invitation to debate. Medical miracles cost money. PMID- 10537467 TI - The evolution of NCQA accreditation. PMID- 10537466 TI - Can fee-for-service Medicare learn from managed care? PMID- 10537468 TI - Improving end-of-life care. PMID- 10537469 TI - Weighing the choices. PMID- 10537471 TI - A labor leader's view ... the U.S. should build on a system that can best create or support the market dynamics and health care principles that will lead to universal coverage. PMID- 10537470 TI - An employer's opinion ... important issues need to be addressed before tax credits replace employer coverage. PMID- 10537472 TI - View from a think tank ... it's time to form another health care system that could operate in parallel with employer-sponsored insurance. PMID- 10537474 TI - The making of a medical director. PMID- 10537473 TI - Getting Medicaid back on track. PMID- 10537475 TI - Sharing decisions ... women and their doctors discuss midlife issues and HRT. PMID- 10537476 TI - The technology of disease management. PMID- 10537477 TI - Working with PBMs (pharmacy benefit managers). PMID- 10537478 TI - Models of care for long-stay nursing home residents. PMID- 10537479 TI - Stats & facts. Staying out of the spotlight: PPOs flourishing in the mainstream. PMID- 10537480 TI - The Balanced Budget Act's devastating effect on home care. PMID- 10537481 TI - The transition to prospective payment changes the face of home care. AB - Home care agencies are actually looking forward to the imminent implementation of the Prospective Payment System, as mandated by the Balanced Budget Act of 1997- that is, if they can survive the implementation of the Interim Payment System. PMID- 10537482 TI - Targeting formulary pull-through strategies: the role of pharmaceutical manufacturers. PMID- 10537483 TI - Evaluating outcomes in normal populations: does it have to be nonexperimental? PMID- 10537484 TI - Assessment of the effect of asthma education on outcomes. AB - Community asthma care centers have been introduced to meet the increasing need for community-based assessment, management, and education of patients with asthma. The Asthma Education Centre at Oakville-Trafalgar Hospital, in Ontario, Canada, has implemented a comprehensive assessment, treatment, and education program with a collection of relevant patient data. The findings presented in this article suggest that these specific education and treatment programs are associated with not only significant reductions in the use of health services, but improvements in health outcomes as well. PMID- 10537485 TI - Health insurance limitations and exclusions under the Americans With Disabilities Act. AB - The Americans With Disabilities Act (ADA) provides protection for persons with disabilities in employment, public services, and certain public accommodations. It may also require employers and health insurers to provide coverage for various treatments and procedures associated with certain disabilities. The following article provides employers and health insurers with useful information surrounding the pitfalls and complexities of the ADA as it relates to health insurance coverage. PMID- 10537486 TI - Marketplace. PPOs juggling act: tighter controls without giving up their prime selling point. PMID- 10537487 TI - Perspectives. HIV survival gains peak; but high prevalence among disadvantaged means new chronic care challenge. PMID- 10537488 TI - Perspectives. Heirs to White House quality panel soldier on in quest to create measurement forum. PMID- 10537489 TI - Perspectives. Reforms don't protect group purchasing schemes from marketplace vicissitudes. PMID- 10537490 TI - Marketplace. Harvard Pilgrim looks to tighter controls to turn unexpected red ink back to black. PMID- 10537491 TI - Perspectives. The long, winding road to integration of alternative and conventional medicine. PMID- 10537492 TI - B. Frederick Becker, Chairman and Chief Executive Officer, MMI Companies, Inc.. Interview by James A. Johnson. PMID- 10537493 TI - Imperatives for leadership in hospitals and health systems. PMID- 10537494 TI - Collaboration and quality in managed care. PMID- 10537495 TI - The effects of medical group practice organizational factors on physicians' use of resources. AB - Few studies have systematically examined the influence of physician, patient, and practice characteristics on physician-directed use of resources within the overall environment of medical group practices and none have included the practice culture in the analysis. This study analyzes the effects of the structure and culture of medical group practices on the amount of resources used to manage uncomplicated hypertension episodes of care for enrollees in a Minneapolis/St. Paul HMO during 1990. Three findings emerged from this study: (1) resource use for a well-defined episode of care varies much more than one would expect in this highly competitive managed care environment; (2) the culture of the group practice appears to be more important than organizational structure in determining resource use for the treatment of hypertension; and (3) together the culture and structural variables only explain 8 percent of the variance in resource use. The study indicated that medical group practice organizations have less influence on physicians' practice styles than expected. The group practices studied are all located in a highly competitive managed care environment and these conditions should be causing them to create more standardized practice styles among their physicians. However, wide variations in individual physician practice styles account for most of the differences observed. Either much of the unexplained variance in resource use for this episode of care results from unobserved patient and illness characteristics, or managed healthcare is not yet causing medical group practices in Minnesota to challenge physicians' individualistic practice styles. PMID- 10537496 TI - Administrative competencies for physician organizations with capitation. AB - In California, it is common for HMOs to capitate physician organizations (e.g., independent practice organizations and multispecialty medical groups) for all professional and outpatient ancillary services (and to share risk for inpatient care) under professional risk capitation contracts. This arrangement exports most of the financial risk from the HMO to the physician organization. When HMOs and physician organizations contract under these arrangements, HMOs delegate many of their administrative functions to physician organizations--giving the physician organization authority to make the decisions needed to manage capitated risk. As a result, administrators of physician organizations must be competent in such areas as provider network development, financial forecasting, utilization and quality management, contract negotiation, and establishing systems for claims, reporting, authorizations, and the like. In this study four HMO and 22 physician organization administrators were interviewed concerning key administrative competencies for managing capitation contracts. The competencies were assessed as key administrative work activities that required specific knowledge, skill, or ability to perform. Identifying these competencies is important for physician organizations preparing for capitated risk and will be essential for organizations preparing for HMO or Medicare capitation. PMID- 10537497 TI - Decision making in high-velocity environments: implications for healthcare. AB - Healthcare can be considered a high-velocity environment and, as such, can benefit from research conducted in other industries regarding strategic decision making. Strategic planning is not only relevant to firms in high-velocity environments, but is also important for high performance and survival. Specifically, decision-making speed seems to be instrumental in differentiating between high and low performers; fast decision makers outperform slow decision makers. This article outlines the differences between fast and slow decision makers, identifies five paralyses that can slow decision making in healthcare, and outlines the role of a planning department in circumventing these paralyses. Executives can use the proposed planning structure to improve both the speed and quality of strategic decisions. The structure uses planning facilitators to avoid the following five paralyses: 1. Analysis. Decision makers can no longer afford the luxury of lengthy, detailed analysis but must develop real-time systems that provide appropriate, timely information. 2. Alternatives. Many alternatives (beyond the traditional two or three) need to be considered and the alternatives must be evaluated simultaneously. 3. Group Think. Decision makers must avoid limited mind-sets and autocratic leadership styles by seeking out independent, knowledgeable counselors. 4. Process. Decision makers need to resolve conflicts through "consensus with qualification," as opposed to waiting for everyone to come on board. 5. Separation. Successful implementation requires a structured process that cuts across disciplines and levels. PMID- 10537498 TI - Cost analysis for decision support: the case of comparing centralized versus distributed methods for blood gas testing. AB - Distributed testing, performed in satellite laboratories or at the bedside, is proliferating within healthcare systems. Users prefer it, and it is fast and convenient. A quick look at marginal costs, however, suggests that cost differentials between distributed and centralized testing may be prohibitive. Sound decision making on the part of health system administrators requires a broader understanding of the costs and benefits of testing options. This study illustrates an approach to cost analysis for decision support where opportunity costs (the costs associated with the next best alternative) provide the basis for decision making. Health system administrators need to understand the opportunity costs involved in their decisions to avoid being misled by analyses that omit important cost elements from consideration. We describe approaches to determining the costs of "stat" laboratory testing options. The costs of various blood gas testing options are compared among a central blood gas laboratory, two satellite laboratories, and point-of-care analysis. Opportunity costs were determined by modeling the substitution of one testing process for another. The cost analysis finds that a judicious mix of alternate-site testing methods can generate annual savings of between $250,000 and $330,000, and at the same time reduce test reporting times. In other words, technology that superficially appears more costly can deliver better service with lower costs. PMID- 10537499 TI - Unions appeal to physicians disillusioned by managed care. PMID- 10537500 TI - Physician profiling alone inadequate to trim pharmacy risk. PMID- 10537501 TI - Expanding AMAP (American Medical Accreditation Program) reduces redundancy, puts providers on same page for quality. PMID- 10537502 TI - Patient protection legislation: an opportunity for MCOs to self-regulate or be regulated? PMID- 10537503 TI - Voluntary external review part of rebuilding MCO image. PMID- 10537504 TI - Hospital system discovers people, not technology, first priority in merger. PMID- 10537505 TI - New frontiers in integrated care. AB - A combination of extended life expectancy and the graying of America is resulting in a rapidly growing industry known as assisted living. Based on a Scandinavian model for senior housing, assisted living first emerged in America during the mid 1980s. The concept is still so new that states that license these facilities do not agree on a precise definition. It is generally considered a residential alternative between independent retirement living and total institutionalized skilled care. PMID- 10537506 TI - The impact of cultural issues on home care personnel and patients. AB - The United States faces increasing cultural diversity in its aging populations. This diversity provides opportunities for home care agencies to expand services as well as to improve patient care. By looking at the demographics, the benefits of comprehensive diversity programs become obvious--for both the agencies themselves and for the clients they serve. PMID- 10537508 TI - Reauthorization of Older Americans Act gathering momentum. PMID- 10537507 TI - Retooling for the future. AB - Several trends will accelerate changes in the industry initiated by Medicare's change in payment methodology, including explosive growth fueled by changing demographics, patient preferences, and technological advances; altered customer buying incentives created by managed care organization-provider partnerships; and accelerated consolidation. Home care agencies should "take inventory" of current practices and systems to determine capability gaps for competing in the new environment. PMID- 10537509 TI - Partnering to provide home medical supplies. PMID- 10537510 TI - Recruiting qualified home care aides: new candidate pools. AB - With the demographic surge of baby boomers and the number of women aged 25-45 projected to decline, the coming decades will see a shortage of workers to care for the elderly. Home care aide agencies will only be able to retain their competitive edge if they widen the pool of candidates from which they recruit and create an attractive and decent job. Creating a decent job with adequate pay, benefits, and support is a business strategy that will attract a wider range of workers, including those with minimal experience, and have positive ramifications for health care in the future--and now. PMID- 10537512 TI - A new millennium .... a new home care. AB - Home care faces many changes--from funding to changing patient populations. Agencies need to assess what they are doing and build into their missions services that take into account these changes. How can they do that? PMID- 10537511 TI - Life without the venipuncture benefit: one agency's experience. AB - The Balanced Budget Act of 1997 included legislation that profoundly affected many residents of the community served by Visiting Health Professionals, Inc. of North Carolina. This legislation not only eliminated coverage for the nursing visit for venipuncture, but also ended coverage for all home health aide visits to these patients for personal care. PMID- 10537513 TI - Hospital employs education, security system to combat baby mix-ups. PMID- 10537515 TI - Handling octuplets media swarm: joint effort of PR and security teams. PMID- 10537514 TI - Using technology to increase safety and improve security productivity. PMID- 10537516 TI - Saving an access control system when your supplier goes out of business. PMID- 10537517 TI - New monitoring systems help hospitals protect unlocked facilities. PMID- 10537518 TI - Hurricane Georges and New Orleans hospitals: preparing for killer storm--Part II. AB - In last month's issue, in Part I, we described the experiences of three New Orleans, LA, hospitals in preparing for and coping with one of the worst potential storms in years. As will be seen, each of the three hospitals featured in Part II faced problems that were both foreseen and unforeseen--some of which required considerable ingenuity to solve, requiring rethinking of prior policies. PMID- 10537519 TI - Dan Bowers on patient movement control systems for infants, elderly. PMID- 10537520 TI - Special report: new paradigms in credentialing. Avoid peer review train wreck: build compliance into your credentialing. PMID- 10537521 TI - Special report: new paradigms in credentialing. New data bank rolls out. PMID- 10537522 TI - Special report: new paradigms in credentialing. Run a 'gap analysis' to fix problems. PMID- 10537523 TI - Special report: new paradigms in credentialing. Reporting: who, what, how much, and when? PMID- 10537524 TI - Target educated patients. PMID- 10537525 TI - Patients' bills of rights get nods and nays. PMID- 10537526 TI - Your RCA (root-cause analysis): how do you select events for analysis? PMID- 10537527 TI - Special hospitals. Models of imperfection. AB - Forensic mental healthcare needs radical change. National standards should be established. An effective model needs to integrate special hospitals, secure units and community care. PMID- 10537528 TI - Special hospitals. About the size of it. AB - Special hospitals are facing their third major upheaval in a decade with their proposed assimilation into mental health trusts. Their size is likely to continue to decrease. But the need for them remains. Patients' needs would be best served by smaller hospitals offering more specialised care, closely integrated with other forensic services. PMID- 10537529 TI - Use it or lose it. PMID- 10537530 TI - Nurse recruitment. Something to write home about. PMID- 10537531 TI - Ambulatory care. High-speed ahead. PMID- 10537532 TI - Commissioner's view. Go configure. PMID- 10537533 TI - Patient's perspective. All in the timing. PMID- 10537534 TI - Minimal access surgery. Welded bliss? PMID- 10537535 TI - Women in the NHS. Leaving the laurels behind. AB - Half the women profiled in a report of top managers in 1993 have now left the NHS. Only one is still in the same job. The number of women in chief executive posts has not increased substantially since 1993. There seems to be little appetite to crack the glass celling which still exists at the top of the NHS. PMID- 10537536 TI - Women in the NHS. That was then, this is now. PMID- 10537537 TI - Successful internal program builds employee morale; reduces turnover. Washington Hospital Center, Washington, DC. AB - The Washington Hospital Center in Washington, D.C. is an example of what good can happen when a health care institution commits to developing a first-class internal communications program. The hospital center created a program that includes one-on-one dialogue between supervisors and staff, "town hall meetings, newsletters, memos, intranet, employee recognition and special events." The program builds employee morale and reduces turnover. It also has increased the quality of care and attention that patients receive. PMID- 10537538 TI - Award-winning ad campaign drives M.D. Anderson marketing efforts. PMID- 10537539 TI - Borgess Health Alliance maintains an annual report card for its agency. PMID- 10537540 TI - Introducing Intermountain Health Care's new president and CEO "went like clockwork". PMID- 10537541 TI - Dinner with the doctors encourages consumers to check out Holy Cross. PMID- 10537542 TI - Marketing effort brings Catholic Medical Centers' forty-five facilities under one banner in New York. PMID- 10537543 TI - St. Louis health system hits home run with its marketing strategies ... BJC Health System. PMID- 10537544 TI - Newborn center selects right formula for fund-raiser ... Regional Medical Center at Memphis, TN. PMID- 10537546 TI - North Oaks Health System introduces $43 million renovation. PMID- 10537545 TI - Rethink sales force's mission; capture customer value. PMID- 10537547 TI - Health care branding plays an important role in success among consumers and patients. PMID- 10537548 TI - Colorado school massacre tested area hospitals' crisis communication plans. PMID- 10537549 TI - Complex systems and the nature of professionalism. PMID- 10537550 TI - Knowledge of and attitudes to the health outcomes approach among community mental health professionals. AB - A focus on health outcomes has significant potential for increasing health gain if adopted by health care staff as part of their professional practice. Understanding health care staff attitudes and receptiveness towards health outcomes may facilitate uptake of this workplace innovation. Community mental health clinicians (n = 101) were sent a questionnaire assessing their knowledge of and attitudes to the health outcomes approach, their expectations as to its likely impact on them, as well as features of their workplace in general. Analysis of the 65 returned questionnaires identified some pessimism about what focusing on health outcomes would achieve for community mental health clinicians or their clients. Training in practical applications of health outcomes measures, involvement in and ownership of health outcomes projects and recognition of health outcomes achievements would facilitate adoption of a health outcomes approach by community mental health clinicians. PMID- 10537551 TI - An emergency department tackles bed management and home-based care. AB - Ipswich Hospital Emergency Department played a vital role in the Post Acute Treatment in the Home Program (PATH) of West Moreton District Health Service. PATH used two strategies to reduce the district reliance on acute hospital beds: a short-stay unit for rapid assessment, treatment and early discharge of patients with simple conditions; and a hospital-in-the-home program utilising community health services to treat acute conditions. The program enhanced existing services to create a new treatment stream for acute patients and to promote a cultural shift from fragmented care to district responsibility for total episode of patient care. PMID- 10537552 TI - Relocation for treatment for leukaemia: a description of need. AB - As rural Queenslanders are isolated geographically due to dispersed population patterns, they are often required to travel long distances to access services, especially services of a specialist nature. The distress of this relocation for treatment is particularly intensified for patients with leukaemia and associated haematological disorders and their carers, as they must often relocate for long periods of time and face invasive and demanding treatments away from the comfort of their own homes. Because such treatments are now highly technical and specialised, even patients from more urbanised areas are also required to relocate for prolonged specialist treatment not available locally. Consequently, for many rural and urban patients with leukaemia, relocation for specialist treatment is a major concern. This discussion presents findings from recent research on a Queensland Government initiative, the Patient Transit Assistance Scheme, designed to address this concern. These findings indicate a high level of hardship for these patients and their families who must travel long distances, often relocate for long periods, and endure additional financial burdens at a time when a majority are dependent on government assistance. PMID- 10537553 TI - Waiting list statistics as performance indicators: observations on their use in hospital management. AB - Improvements in data collection and the types of statistics collected have enhanced the usefulness of waiting list statistics as a measure of hospital performance. But these changes are not sufficient for waiting list statistics to be used effectively for management purposes. The statistics need to be viewed alongside activity data if clinicians and managers are to identify specific areas that need improvement. This means that how the data are analysed and presented is also important. During a study into the management of waiting lists, we observed that waiting list data were typically presented in a way that made interpretation difficult. A simple but effective solution was found by using available PC-based software, but obstacles remain. These stem from limitations of current information systems and the awareness among staff of the potential of common software packages. PMID- 10537554 TI - The development and evaluation of satellite endoscopy services in Western Australia. AB - A satellite endoscopy service was formally established in late January 1997 in one peripheral hospital, a second service commencing in April 1997, and a third in December 1997. More than 500 patients underwent gastrointestinal endoscopic procedures at these satellite services during 1997. The feedback received to date indicates that the establishment of this service is supported by patients and the peripheral hospitals. It is expected that this project will achieve all its stated objectives. In addition, the implementation of this service will assist in improved waiting list management at Royal Perth Hospital. The satellite endoscopy service is a unique development in Western Australia and has demonstrated definite benefits to patients in less than six months of operation. PMID- 10537555 TI - Acute hospital medical staffing during the night shift. AB - There has been little or no attempt to define the need for 24-hour medical cover, nor its appropriateness in acute hospitals, despite the great cost implications and the question of the quality of that care. This study examined the medical activity during the 'night shift' in an acute hospital. There were an average of 2.59 calls per night, most from the emergency department (247/475) and general wards (108/475). Many calls were related to active resuscitation (88/475) and immediate treatment (83/475). Over 40% (81/286) of patients had to be transferred to a higher level of care, such as an intensive care unit within the hospital. By collecting data on the demands of health care during what amounts to over a third of the hospital's time, it was established that a high level of medical care was required. Appropriate levels of staffing, using junior doctors trained in acute medicine, was able to be provided to match need as determined by these data, and extra staff at higher costs were avoided. PMID- 10537556 TI - Casemix funding in rural NSW: exploring the effects of isolation and size. AB - The New South Wales Department of Health (NSW Health) wishes to make appropriate use of casemix data as inputs to the determination of funding levels for small rural hospitals. However, other factors such as hospital size and degree of isolation might need to be taken into account. The study reported here involved correlation of actual expenditures with those predicted by use of a casemix model alone, across 105 small public hospitals in the State. We then explored the extent to which the correlation could be increased by the addition of distance and isolation variables. It was found that actual costs were highly correlated with those predicted from the casemix data alone, and that the correlation increased when both the distance and the size variables were introduced. However, contrary to expectations, reduced size was associated with reduced costs, and reduced isolation was associated with increased costs. It was concluded that, while the predicted relationships may be present, they are likely to be relatively weak and are probably being masked by other factors not present in the model. In particular, it seems likely that there are variations in severity within the acute admitted patient category which are not fully explained by the casemix instrument used in this study (the DRG classification). We suggest that other terms be introduced to control for this possibility before any further attempt is made to test whether size and distance factors can be identified which work in the expected direction. PMID- 10537557 TI - Hospital outreach service: helping to prevent nursing home placement. AB - An outreach service from a post-acute metropolitan teaching hospital delivered an intensive, multidisciplinary and coordinated allied health service, and achieved both early hospital discharge and the prevention or delay of nursing home placement. This article reports on three types of cases which illustrate how the service assisted ward teams, families and patients to determine whether nursing home placement was essential. For a group of 20 cases, the total reduction in hospital length of stay was 556 days, and home accommodation as an alternative to nursing home accommodation was achieved for a total of 7505 days. The article outlines a matrix of advantages and disadvantages, both tangible and intangible, of home versus nursing home accommodation. It is suggested that a full costing of this matrix would inform debate on the comparative merits of long-term home and nursing home accommodation. PMID- 10537558 TI - Collaborative relationships in general practice projects. AB - This article reports on a national study of collaborative relationships between general practitioners and other health care providers in 20 Division of General Practice projects. It argues that health care organisations will need to collaborate with others in the future and that much can be learnt from the literature on collaborative networks in business and community organisations. Successful collaborations between general practitioners and others were found to be consistent with a model of collaboration in 'under-organised domains', where pre-existing links between organisations are weak. Lessons are identified from the study to assist future collaborative ventures involving general practitioners. PMID- 10537559 TI - Case management at Warringal Private Hospital: challenges of development, implementation and evaluation. AB - Case management has the potential to improve the quality of care for patients, streamline efficiencies within organisations, and ultimately lower health care expenditure. This article explores why Warringal Private Hospital embraced the concept and how the chosen model of case management was developed. It describes the implementation and evaluation of the model and how it was received, accepted, and applied by the various stakeholders. The cardiac and orthopaedic units will be cited as case studies in order to emphasise some of the challenges encountered in this process as well as the successful outcomes. It should be noted, however, that each unit within the hospital is unique and, although the principles of case management have been applied throughout the hospital, the development, implementation and evaluation has been specific to each unit. PMID- 10537560 TI - Compliance of hand washing practices: theory versus practice. AB - Hand washing remains an important preventative method for making the transmission of nosocomial infections redundant. Despite awareness by health workers of the practices required and of the legislation governing hand washing, the study reported here found that compliance to these procedures was quite poor. The results of two surveys distributed to health workers and direct observation by clinical staff in an aged care hospital found that 45% of health workers did not wash their hands and 24% did not change their gloves between patient consultation. Methods for increasing effective hand washing in clinical settings must be identified if hygienic practice is to be improved. PMID- 10537561 TI - Rationalising the ordering of blood cultures. AB - In 1996 a project was undertaken to determine the clinical impact of blood cultures taken in the emergency department. It found that 1.6% of all blood cultures taken resulted in changes in patient management. In response to these findings, guidelines were developed for more appropriate utilisation of blood cultures. It was predicted that the guidelines would result in a reduction in test ordering of approximately 40% and an annual saving of approximately $18,000. The guidelines were implemented in mid-1997. An audit of test ordering for the months of January to August 1998 shows a 53% reduction in the ordering of blood cultures. PMID- 10537562 TI - Health outcomes as an organisation-wide quality initiative. AB - Since the burgeoning of the 'health outcomes' movement there has been an ever increasing body of literature on health outcomes policy debates, directions, frameworks and tools for implementing health outcome-directed initiatives. There is a significant gap in the literature, however, in regard to translating a comprehensive health outcomes policy into practice at a local level. This paper addresses that gap. It describes the local implementation of a comprehensive health outcomes approach which works across the continuum of care. It identifies those organisation-wide structures and processes that support successful progress, thereby providing a useful guide to other organisations wishing to institutionalise the health outcomes approach. PMID- 10537563 TI - Prescribed drugs: a major cause of ill health. AB - Harm caused by ill-effects of prescribed drugs is largely unrecognised, but when looked for has been found to be an important cause of ill health. Perhaps 6% of all hospital admissions and some 800 deaths a year are caused by prescribed medication in Australia. The elderly, especially those in institutions, are particularly prone to injury by drugs. Attempts to use visiting pharmacists to influence the prescribing habits of doctors have resulted in unspectacular success. It is suggested that prescribing habits may more effectively be changed by visiting from specially trained practising doctors. PMID- 10537564 TI - International health--how Australia compares. PMID- 10537565 TI - Health policy and its impact on poverty. AB - Poverty may be defined narrowly as a lack of income, but is more usefully viewed as a multidimensional concept. I discuss some associations between poverty and health, identify groups with special needs, and describe some aspects of the government's health policy which are relevant to those needs. Finally, I note the importance of ensuring there is a more integrated approach in future. PMID- 10537566 TI - Severity variations within DRGs. PMID- 10537567 TI - Trusting the surgeon: a tornado from Bristol. PMID- 10537568 TI - The Asian currency crisis and the Australian health industry. AB - This article identifies linkages between the Australian health industry and the global economy. It discusses some of the consequences of the Asian currency crisis of 1997-98 for the Australian economy and health industry, with special emphasis upon exports. Devaluation of the Australian dollar will increase the cost of most pharmaceutical and medical imports, but may offer competitive advantages to some Australian exporters. The nascent engagement with Asia of many health industry enterprises is likely to be stifled. It is therefore important for Australian governments, as well as the Australian health industry, to provide intelligence and encouragement to those enterprises that wish to continue their engagement with Asia or resume it when economic equilibrium returns. Markets throughout the world must also be further developed. The crisis may therefore provide the stimulus for re-thinking and re-stating Australian health export policy. PMID- 10537569 TI - Indigenous health: patterns of variation in terms of disease categories. AB - While many studies investigated the higher morbidity and mortality levels of indigenous Australians in the high-density indigenous areas in the Northern Territory, Western Australia and South Australia, few examined the situation in New South Wales, where more than 28% of the indigenous population lives. Admissions to acute public and private hospitals in New South Wales for 1989-1995 are used in the study reported here to examine indigenous health and its differential patterns by disease categories. The study allowed for the monitoring of disease groups with particularly high indigenous admissions and, accordingly, pinpointed areas for improvement. Age-standardised estimates for the indigenous population are provided. Age composition of admissions for each disease category and admissions by residential area are also estimated. PMID- 10537570 TI - Outcome evaluation in nursing in Australia, 1960-1980. AB - While empirical evaluation of the outcome of patient care has come to the fore in recent years due to political initiatives, there has always been a professional interest by nurses in the end result of their care. A review of the literature shows that outcome evaluation was advocated for nursing as early as the 1860s by Florence Nightingale. This article explores the evolvement of outcome evaluation within nursing in Australia, discussing its origins during the 1960s and 1970s. The measurement of patient outcomes is more relevant than ever before, with the recent drive for an evidence-based approach to nursing care. PMID- 10537571 TI - Committing the AMA to quality in Australian health care. PMID- 10537572 TI - Using endoscopic procedures for AN-DRG (Australian National Diagnosis Related Groups) assignment: Australia leads the way. AB - The study reported in this article sought to develop Australian National Diagnosis Related Groups (AN-DRGs) using endoscopic procedures in Major Diagnostic Category (MDC) 6 (Digestive System) and MDC 7 (Hepatobiliary System and Pancreas) through statistical analysis of the Australian Casemix Clinical Committee's recommendations. Five ANOVA were undertaken on final recommendations for gastroscopy and colonoscopy in MDC 6. The Reduction in Variance (RIV) for the AN-DRGs in version 3 relative to version 2 increased by up to 14.6%, representing RIV of between 25.28% to 32.30%. For all ANOVAs, F > 100, alpha < .0001, Coefficient of Variation (CV) was generally lower in version 3 by between 0.4% to 22.9%, except for AN-DRGs for other gastroscopy for major gastro-intestinal disease, which increased by 8.7%. Two ANOVA for Endoscopic Retrograde Cholangio pancreatography Procedures (ERCP) recommendations resulted in RIV of up to 18.67%, F > 100, alpha < .0001 and CV up to 0.8091. MDC 6, in AN-DRG versions 3 and 3.1, has 11 AN-DRGs following the surgical hierarchy involving gastroscopy and colonoscopy. Patients assigned will not have an operating room procedure; they will have a non-operating room procedure that is either a complex therapeutic or other (diagnostic or therapeutic) procedure. Similar AN-DRGs are in MDC 7 for ERCP. Version 3.1 has expanded the definition of Common Bile Duct Exploration (CDE) to include ERCP. There is no separate AN-DRG for laparoscopy cholecystectomy. PMID- 10537573 TI - Time for professional and individual accountability. PMID- 10537574 TI - Future financial impact of the current health financing system. AB - Major political parties remain publicly committed to Medicare and community-rated voluntary health insurance. It is important to understand the future financial consequences of this policy in order to assist community debate about whether such a commitment is appropriate or some other policy should be developed. This paper describes development of, and results from, the APHA health financing model. It suggests that health expenditure would represent 12.9% of gross domestic product by 2021, compared to 8.5% in 1995. Increasing per capita expenditure is the major contributor to the growth, with demographic changes responsible for only 14.3%. PMID- 10537575 TI - Assisted venous drainage, venous air, and gaseous microemboli transmission into the arterial line: an in-vitro study. AB - The objective of this study was to examine the interaction of cardiopulmonary bypass venous air with assisted venous drainage, focusing on its production of gaseous microemboli in the arterial line. An in-vitro recirculating cardiopulmonary bypass circuit containing fresh whole bovine blood was monitored with a pulsed-doppler microbubble detector. Air of specific amounts was injected into the venous line and gaseous microemboli counts were obtained distal to the arterial filter. Data was recorded for unassisted drainage, vacuum-assisted drainage, and centrifugal pump-assisted drainage. Centrifugal pump-assisted drainage produced over 300 microbubbles in one minute distal to the arterial filter when venous air was introduced into the circuit. Of these, 220 were greater than 80 microns in size. Vacuum-assisted drainage produced no microbubbles when the same amount of venous air was introduced into the circuit. However, vacuum-assisted drainage did produce some microbubbles in the arterial line when a stopcock was left open on the venous line for 30 seconds. Unassisted drainage produced no microbubbles at all levels of venous air entrainment. Air becomes entrained in the venous line from a variety of sources. In a typical gravity-drained situation, the air remains whole and is dissipated in the venous reservoir by buoyancy and filtration. In an assisted-drainage situation, the air is subjected to additional forces. The air is subjected to a greater degree of negative pressure and, with centrifugal pump assisted drainage, is subjected to kinetic energy imparted by the cones or vanes of the pump. The kinetic energy from the centrifugal pump appears to break the air into small bubbles which become suspended in the blood, passing through the reservoir, oxygenator, and arterial filter. In a clinical setting, these bubbles would be passed into a patient's arterial system. PMID- 10537576 TI - Clinical evaluation of a new in-line continuous blood gas monitor. AB - Two methodologies for obtaining accurate blood gas and electrolyte values during cardiopulmonary bypass (CPB) are traditional laboratory analyzers, which use an electrochemical technology, and continuous in-line monitoring systems, which use a fluorometric and/or spectrophotometric technology. The purpose of the present study was to evaluate the accuracy of a new continuous in-line monitor, the 3M CDI Blood Parameter Monitoring System 500, which provides continuous in-line measurements of pH, PCO2, PO2, potassium (K+), oxygen saturation, hematocrit, hemoglobin, and temperature, during partial or complete CPB. Study parameters included arterial pH, PCO2, PO2, and K+ values. Overall performance was analyzed by calculating the mean difference (expressed as the bias) between the CDI system 500 and the laboratory analyzer for each parameter. The accuracy of the arterial pH, PCO2, and K+ values provided by the CDI system 500 was then evaluated using target values established in the acceptable performance standards for laboratory analyzers from the Clinical Laboratory Improvement Act of 1988 (CLIA '88). The accuracy of the PO2 value provided by the CDI system 500 was evaluated using a target value of +/- 10% of the reference, or laboratory analyzer, value. A prospective multi-center trial was conducted following Institutional Review Board approval. A total of 75 cases was included in the analyses, with over 200 data points from 4 clinical locations. Results for pH, PCO2, and K+ were within the target values established by CLIA '88. pH bias was 0.00 +/- 0.02 pH units. PCO2 bias was -0.3 +/- 3.3 mm Hg. K+ bias was approximately +0.12 +/- 0.31 mmole/l. Results for PO2 were within 10% of the reference value. PO2 bias was 7.5 +/- 13.8 mm Hg. The results of this clinical trial show that the CDI System 500 continuous in-line monitoring system provides values that meet the accuracy standards for laboratory analyzers for arterial pH, PCO2, PO2, and K+. PMID- 10537577 TI - Whole blood platelet function assay on the ICHOR point-of-care hematology analyzer. AB - The role of platelets as the initial defense against insult to the vasculature is well established. Moreover, platelets are now recognized as having a critical role in the acute care settings of cardiopulmonary bypass (CPB) procedures and cardiac catheterization. In the environment of CPB, both platelet count and function have been demonstrated as being markedly compromised during and following the procedure. Unfortunately, current assays that are used to evaluate the parameters of platelet count and function are limited in regard to their utility in a near patient format. Here, we describe a practical, rapid, and user friendly whole blood platelet function assay that has been developed for the ICHOR point-of-care hematology analyzer. This analyzer is capable of performing an eight parameter blood profile including platelet count. In comparable studies, platelet aggregation in whole blood demonstrated good correlation (for ADP the values were n = 14, r2 = 0.81, p = 0.0001; for collagen, n = 10, r2 = 0.93, p = 0.0001; for ristocetin, n = 10, r2 = 0.89, p = 0.0001; and for epinephrine, n = 10, r2 = 0.81, p = 0.0003) with traditional platelet-rich aggregometry, which uses increased light transmission as an indication of platelet aggregation. Furthermore, early feasibility studies in CPB patients demonstrated both decreased platelet count and a marked reduction in platelet function peri procedurally. This new assay of platelet function is extremely suitable for the clinical environment with rapid turnaround time and provides a full hematology profile to enhance transfusion decisions. PMID- 10537578 TI - Antithrombin III in cardiac surgery: an outcome study. AB - A retrospective study examined the impact, in heparin resistant patients (HRP), of lyophilized antithrombin III (ATIII) upon five patient outcomes: intensive care unit stay (ICU-S), 24 hour chest tube drainage (CTD in ml), blood and blood product usage (BPU), development of postoperative coagulopathy (PO-Coag), and reoperation for bleeding (Re-Op). Data was collected from the medical records of 311 patients admitted to the hospital between 12/15/95 and 10/24/96. Subjects were divided into three groups based upon heparin resistance and hemostasis medication. Group 1 (n = 109) were HRP treated with increased heparin, Group 2 (n = 100) were HRP receiving ATIII, and Group 3 (n = 102) were non-HRP and served as controls. Group 2 was also subdivided by use of aminocaproic acid and time of ATIII administration. No significant differences were found between the groups for PO-Coag. and Re-Op. However, significant reduction in CTD (p = 0.05) was seen in the aminocaproic acid patients who were treated with ATIII pre-CPB or within the first 20 minutes of CPB. The CTD in this group was (419.37, +/- 72.96) as compared to Group 1 (782.88, +/- 360.94) and Group 3 (766.67, +/- 407.56). Other Group 2 subgroups showed significant differences in BPU, ICU-S and CTD. The results of this study support the notion that early identification and treatment of HRP with ATIII and aminocaproic acid may decrease postoperative blood loss. PMID- 10537579 TI - Experimental use of an ultra-low prime neonatal cardiopulmonary bypass circuit utilizing vacuum-assisted venous drainage. AB - In adult cardiopulmonary bypass surgery, vacuum assisted venous drainage has become a popular technique to augment venous return to the bypass circuit. The application of this technique in neonatal cardiopulmonary bypass surgery could be beneficial to the further miniaturization of neonatal circuitry by coupling radical respositioning of the oxygenator and pump console with decreasing line length. This report communicates the use of an investigational, vacuum assisted venous drainage neonatal circuit that is positioned at patient level utilizing a modified pump console with elevated double head twin roller pumps. The circuit, including the oxygenator, arterial line, venous line, raceway tubing, and a functional level in the venous reservoir has a priming volume of 107 ml. Initial bench and animal tests have demonstrated that this technique may be clinically feasible in CPB applications. With vacuum assisted venous drainage, the goal of asanguinous neonatal cardiac surgery could become a reality. Safety issues must be adequately addressed to ensure that this technique does not impose unacceptable risks. PMID- 10537580 TI - Residual blood in neonatal oxygenators after drainage. AB - The objective of this investigation was to measure the quantity of residual blood remaining in neonatal cardiopulmonary bypass (CPB) circuits after they had been drained and to assess the overall significance with regards to total patient blood volume. The residual blood volume left in three infant/neonatal CPB circuits-Medtronic Minimax 3381 (Group MM; n = 5), Polystan Safe Micro (Group SM; n = 6), and Terumo Capiox 308 (Group CX; n = 3)--after they had been drained was determined. This was done by using an electronic scale to weigh the circuit before setup and after CPB when all possible blood was recovered from it. Total priming volume, estimated patient blood volume, residual blood volume, surgical blood loss in theater, and autogeneic blood usage were recorded in each case. Mean residual blood volumes measured were MM = 161 ml (SD 27 ml), SM = 103 ml (SD 19 ml), and CX = 133 ml (SD 15 ml). These volumes were significant, because calculations show that the volume of red cells lost in the circuit is equivalent to fourteen percent of the total patient blood volume. In view of this, neonatal oxygenator design should be minimized to reduce the priming volume and more consideration should be given to ease of residual blood recovery. PMID- 10537581 TI - Perfusing the Jehovah's Witness patient with heparin-induced thrombocytopenia. AB - Heparin-induced thrombocytopenia (HIT) is an uncommon, yet dangerous side-effect of heparin therapy. The problems associated with the HIT patient while undergoing cardiopulmonary bypass increase dramatically when the patient is also of Jehovah's Witness faith. This case report depicts the techniques utilized and the decisions made over the course of a simple surgical procedure for an extremely high-risk patient. PMID- 10537582 TI - Osseointegration. PMID- 10537583 TI - The bone-implant interface: a dynamic surface. AB - This study quantifies and compares bone formation on and around roughened titanium implants with roughened cobalt chromium, polished solid implants, and titanium fibermetal implants. Cylindrical rods were implanted into the medullary canal of the distal femur of rabbits. The bone-implant interface was studied 3, 6, and 12 weeks after surgery using histomorphometric methods. Roughened surface implants demonstrated significantly more bone directly apposed to the surfaces when compared to the polished or fiber/metal implants at 6 and 12 weeks after surgery. New bone formation and remodeling of bone occurred directly on roughened surfaces as late as 12 weeks after implantation, but not on the unroughened implants. These results suggest that roughening of the surfaces of both titanium and cobalt chromium implants can enhance osseointegration and may be useful clinically for the fixation of prosthetic components. PMID- 10537584 TI - Osseointegration of cemented and noncemented implants in artificial hip replacement: long-term findings in man. AB - Implant fixation is of utmost importance for pain-free function of endoprostheses. Primary fixation is achieved during implantation, whereas secondary fixation is a result of repair and bone remodelling during the healing process comparable to fracture healing. The chronologic course of the healing process follows three overlapping phases: (i) an initial phase with destruction and necroses of bone, (ii) a phase of repair with integration of the implant into the bone, and (iii) a phase of stabilization of the permanent implant bed with adaptation to load transfer and possible reactions to irritations of the interface like sepsis, mechanical instability, corrosion and degradation of implant materials, and wear products from articulating and anchoring surfaces. PMID- 10537585 TI - A review of ceramic coatings for implant fixation. AB - The present series of eight studies was performed in order to investigate the effect of various clinically relevant factors on bone ingrowth in relation to hydroxyapatite (HA) and titanium-alloy (Ti) coating when subjected to pathological and mechanical conditions mimicking the clinical situation. HA- and Ti-coated implants were inserted into the femoral condyles of mature dogs and one study was performed on humans. The observation period ranged from 4 to 52 weeks, and the results were evaluated by mechanical push-out testing, histomorphometric analysis, polarized light microscopy, UV fluorescence microscopy and collagen analysis. There were no complications related to the operative procedures, and all dogs were killed according to the original time schedule. Two studies focused on in vivo mechanisms and factors influencing resorption of HA coating. The overall conclusions from these studies are that HA coatings do resorb in vivo, that micromotion accelerates resorption, and that resorbed HA is partly replaced by newly formed bone, suggesting that implants fixation is durable. The other studies focused on the significance of mechanical stabilization and loading conditions of the implant immediately after surgery. From these studies, it can be concluded that HA-coating has a positive effect on bone-implant fixation in various situations, i.e., under stable loaded conditions and under unstable mechanical conditions. The most striking effect of HA coating was that it enhanced bone growth across a gap around the implant both during stable and unstable mechanical conditions; it even converted a motion-induced fibrous membrane to bony anchorage. PMID- 10537586 TI - Bone adaptation to a polyester fiber anterior cruciate ligament replacement. AB - A series of polyester fiber ACL implants was studied in ovine stifle joints up to 2 years postimplantation. The implants were linked to the bone-tunnel wall by oriented fibrous tissue. Cross-sections of the tunnels showed bone ingrowth among the implant fibers at 2 years. A human trial of the Apex implant yielded a series of retrievals, some associated with gross bone-tunnel enlargement. There was no evidence of bone ingrowth in the human implants. It was hypothesized that-tunnel enlargement resulted from fretting at the implant-tissue interface in response to cyclic loads in use. PMID- 10537588 TI - Woven bone formation around implants and the effect of bacterial infection. AB - Several implant materials used in dental and orthopedic surgery were placed in rat tibial bones to study their effects on mineralization. The implants consisted of bone bonding and non-bonding materials. Changes in mineralization were defined by morphometric analysis of matrix vesicle distribution at the implant interface and in normal bone healing following marrow injury. Bone-bonding materials induced an increase in matrix vesicle activity. This finding was supported by study of the biochemical changes in the same model that manifested high correlations to the morphometrical observations with regard to enhancement or delay of primary mineralization. In addition, the study of healing using nuclear methods indicated that implants alter bone healing as shown by the different uptakes of 99mTc and 32P in the different bone compartments. Decreased 32P uptake by the organic phase in the presence of bone-bonding implants suggested that cleavage of 99mTc-MD32P into its technetium and methylene diphosphonate moieties was inhibited by administration of implants. Further studies on the effect of bacterial infection on the peri-implant tissues revealed a decrease in woven bone formation due to infection. PMID- 10537587 TI - The osteogenic properties of the interface membrane at the site of orthopedic implants: the impact of underlying joint disease. AB - Osseointegration at the site of orthopedic implants is dependent on the recruitment, attachment, and differentiation of osteogenic cells. Data concerning the effect of a patient's underlying joint disease on the modulation of the cellular activity and the long-term survival of joint prostheses is limited. In this study, immunocytochemistry was used to investigate the osteogenic cell phenotype within the bone-implant interface fibrous membrane in 60 patients with different underlying joint disease. Tissue specimens were removed during revision operations performed at variable times following implantation. The results provided histological evidence of the presence of fibrocartilage tissue and calcified bone within the interface. TGF-beta, metalloproteinases (MMP1 and MMP2) and their inhibitors (TIMP1 and TIMP2) were immunolocalized within fibroblasts, chondrocytes, and osteoblasts throughout the interface, indicating that signals modulating the osteogenic cell phenotype at these sites are highly regulated. Finally, the study identified a significant difference in the histological changes elicited by implant particulate debris in patients of different diagnostic categories. Such observations imply that the activity of the original joint disorder could augment specific cellular activation/immune signals that subsequently affect the degree of the local inflammatory responses to implant wear particles. The negative balance between the rate of bone growth and resorption around the prosthetic joint is central to the pathogenesis of aseptic loosening of implants. PMID- 10537589 TI - Studies of host response to orthopedic implants and biomaterials. AB - The use of implanted biomaterials in orthopedic surgery has increased rapidly during the past two decades. Total joint replacement of the hip or knee joint has become common treatment; at the same time, an increasing number of fractures are treated with osteosynthesis. The original Charnley low-friction arthroplasty of the hip is still widely used and gives in large series excellent results. Aseptic loosening of this arthroplasty has been thought to be due to wear debris of the methylmethacrylate used for fixation of the implants, or to debris generated from wear of the polyethylene socket. To date, many different materials have been tried in order to reduce wear and generation of macrophage irritating submicron sized particles, or to provide more biocompatible components. However, trials to improve the methylmethacrylate cement or to invent better polyethylenes have often failed. Diamond coating of the metallic components seems promising: there is less wear and diamond is very biocompatible in bulk and small particulate form. Biodegradable implants have also been found useful in treating fractures. Bioactive bioabsorbable materials may also make possible a tissue engineering approach and can be used as carriers for selected drugs and cytokines. Because many promising materials and designs have failed in clinical use, extensive theoretical and experimental testing is mandatory before introducing new materials and implants in a clinical setting. PMID- 10537590 TI - Osseointegration of total hip arthroplasties: studies in humans and animals. AB - Total hip replacement is a successful, time-proven surgical procedure for reconstruction of the arthritic hip joint. The state of the bone-implant interface is crucial to the long-term integration and durability of hip replacements whether cemented or cementless. This review summarizes current clinical implant retrieval and animal research in hip-joint reconstruction. Future research must attempt to extend the longevity of hip replacements to avoid complex revision surgery. PMID- 10537591 TI - Pediatric disasters. PMID- 10537592 TI - Critical incident stress intervention after loss of an air ambulance: two-year follow up. AB - OBJECTIVE: Following an air ambulance crash with five fatalities, critical incident stress debriefing (CISD) was provided for involved paramedics, physicians, and nurses. A study was conducted to evaluate the long-term effects of a critical incident with critical incident stress debriefing according to the Mitchell model. METHODS: Six months following the incident, empirically designed questionnaires were mailed to all transport paramedics and directly involved medical staff, and a random sample of both nurses from the dispatch/receiving institution and paramedics from around the province. Twenty-four months post incident, all members of the transport paramedics completed the Impact of Events Scale and the General Health Questionnaires. RESULTS: There were no differences between groups on any scores, except for disturbed sleep patterns, bad dreams, and the need for personal counseling being greater among transport paramedics at one day. There was no correlation between how well the deceased individuals were known, amount of debriefing, and symptom severity. A trend was seen for those with pre-existing stress management routines to have less severe symptoms at six months (p = 0.07). At two years, 16% of transport paramedics still had significant abnormal behavior. CONCLUSIONS: CISD did not appear to affect the severity of stress symptoms, whereas having pre-existing stress management strategies may. These findings give justification for proceeding to a randomized, controlled trial of different levels of critical incident stress intervention. PMID- 10537594 TI - EMS: the Canadian perspective. PMID- 10537593 TI - An Israeli model of a hospital emergency information center. AB - The Emergency Information Center model developed by the social work department of Tel Aviv Sourasky Medical Center is activated in cases of mass casualties following disasters. It aims to provide reliable information to help the public cope with confusion and uncertainty, and to enable the medical staff to concentrate on treating the casualties. The Information Center is comprised of several interrelated units within the hospital, and maintains contact with a range of community organizations. The article describes the structure and activities of the various units, and discusses a number of aspects relevant to personnel organization in crisis intervention. PMID- 10537595 TI - KAMEDO--a Swedish Disaster Medicine Study organization. AB - Kamedo is a Swedish Disaster Medicine study organization that sends observers to disaster areas anywhere in the world to study recent events, collect useful information, and identify problems relative to the practice of Disaster Medicine. The results of these investigations are published in the KAMEDO Reports, and the English versions will be published in Prehospital and Disaster Medicine. Three of the recent reports follow: 1) KAMEDO Report 69: Ebolus Virus Epidemic in Zaire, 1995; 2) KAMEDO Report 70: The German Rescue and Emergency Organizations: a) Industrial Chemical Fire, Memmingen, Germany 23 January 1997; b) Fire at the Dusseldorf Airport, 01 April 1996; and c) Bus Accident on the Autobahn in Rosenheim, Germany; and 3) Terrorist Attack with Sarin, 20 March 1995. In addition, a catalog listing all of the KAMEDO Reports available in English is provided. PMID- 10537596 TI - Measures for increased nutrition and utilization of non-conventional food resources during disasters in Africa. AB - The basic causes of the poor performance of the food and agricultural sector in the different parts of Africa are external, internal, and natural. The general recession in the Continent limits the capacity of the respective countries to import food to supplement inadequate domestic production and supplies. There are a number of nutritious food resources, both cultivated and gathered in the different ecological zones of Africa, whose production and consumption can be increased to ensure adequate food security and a nutritious diet, especially during disasters. These food resources could include: cereals, legumes, fruits, vegetables, fish, and insects. These food resources already are available over wide geographical areas in Africa and are utilized or utilized to a limited extent. Therefore, strategies to increase food supply, eradicate hunger and malnutrition, and keep people alive in times of disasters should have as a priority, the cultivation and consumption of non-conventional food resources in the respective communities and countries. PMID- 10537597 TI - Flood disaster in the Czech Republic in July, 1997--operations of the emergency medical service. AB - This report is a review of the response and the activities of the Emergency Medical Services during a huge flood that devastated one-third of the territory of the Czech Republic in July 1997. The Emergency Medical Services personnel extracted by helicopter a great number of citizens who were trapped in their flats and homes. For diabetics and cardiacs who were isolated from the surface transport, the EMS personnel supplied necessary medication, and transported patients to hemodialysis. The cooperation between non-medical emergency services and the district crisis staff of the Integrated Rescue System, varied in different districts. However, in most flooded districts, the cooperation was satisfactory. In addition, a large number of volunteers helped in the first days of the flood. Unfortunately, 49 people died because of the flood. Nevertheless, since the EMS was able to manage the extraordinary needs, the number of emergencies and hospitalizations was low. PMID- 10537598 TI - National EMS curricula: guidelines or policies for practice? PMID- 10537599 TI - Who's minding the public health store? PMID- 10537600 TI - Full-time employees of U.S. local health departments, 1992-1993. AB - This article describes a study to assess the most recent data on full-time U.S. local health department (LHD) staff positions. The authors used data from the National Association of County and City Health Officials' 1992-1993 national survey of LHDs. The study concludes that nurses, environmental specialists, sanitarians, and administrators constitute the core of the public health workforce in smaller and mid-sized LHDs. Numerous vacancies in these core occupations signal a weakness in the front lines of public health and vulnerability in its ability to respond to urgent health threats. To address these problems, a renewed commitment to recruiting, retraining, and retaining the local public health worker is urgently needed. PMID- 10537601 TI - Public health workforce information: a state-level study. AB - A two-stage sample survey was used to estimate the size of Texas' professional public health workforce and to describe its composition in terms of employment settings, job characteristics, and individual characteristics. The estimated 17,700 public health professionals employed in 1995 represented approximately three percent of the state's total health workforce. About 55 percent of all these professionals worked in agencies that provide population-based public health services. An estimated seven percent had formal public health education. These findings raise issues concerning the numerical adequacy of the state's supply of public health professionals, the adequacy of their educational preparation, and the human resources capacity of the state's official public health agencies. PMID- 10537602 TI - The mystery of public health workforce development. PMID- 10537603 TI - Extended degree and continuing education preferences of California public health professionals. AB - The authors conducted a randomized mail survey of members of six public health organizations in California. The purpose of the survey was to assess public health practitioner interest and resources available to participate in public health extended degree programs and public health continuing education (CE). The response rate was 52 percent (N = 262). Three CE topics of greatest interest to public health professionals were health policy, computer applications, and community-based interventions. Respondents were interested in both distance-based and on-site learning formats. PMID- 10537604 TI - Final report on public health practice linkages between schools of public health and state health agencies: 1992-1996. AB - Since 1988 there has been a call for enhanced linkages between schools of public health and public health agencies that has prompted schools of public health to develop public health practice initiatives. The University of Illinois at Chicago School of Public Health conducted surveys of schools of public health and of state public health agencies in 1992 to collect baseline data on practice initiatives undertaken by academe and governmental public health agencies to enhance collaboration; follow-up surveys were undertaken in 1993, 1994 and 1996. This article describes the trends and implications of this survey of practice linkages involving schools of public health and state health agencies. PMID- 10537605 TI - Sources of health insurance in the U.S.: analysis of state-level data and implications for public health programs. AB - Lack of health insurance coverage is associated with lack of accessibility to preventive health care services such as mammography screening, clinical breast examination, Papanicolaou smear test, digital rectal examination, proctoscopy examination, and cholesterol screening. State and federal public health agencies must have an understanding of insurance coverage of the population to plan intervention programs aimed at early detection of medical conditions. Using data from the March Supplement of the Current Population Survey for the years 1994, 1995, and 1996, this study examines the sources of health insurance coverage in the U.S. The implications of the findings for public health programs are discussed. PMID- 10537606 TI - Who speaks for public health agencies: assessing the core functions in local health departments. AB - Several large-scale studies have attempted to measure public health agency performance of core functions by interviewing health directors. Because it is not self evident that a single respondent results in a valid performance assessment, the purpose of this study was to examine the relationship between two characteristics--position in the agency and racial/ethnic identity--and perceptions of the performance of core functions. Supervisors differed from nonsupervisors and Whites differed from African Americans in their perceptions of practices reflecting assessment and policy development. Efforts to develop surveillance measures of a complex institution such as a health department should incorporate methodologies to validate the responses. PMID- 10537607 TI - Community-based promotion of breast screening using small group education. AB - Small Group Education (SGE) to promote breast cancer screening was implemented in a community-wide program. Based on diffusion of innovations theory, SGE initially was directed toward women at higher occupation and education levels and then progressively shifted toward more vulnerable populations of women at risk of not getting screening. During the four-year intervention, 116 volunteers led SGE presentations, with 8,184 women participating in 740 groups at work sites, organizations, residences, and churches. High participation in SGE and positive participant responses suggest that delivery of SGE using a social diffusion model was an effective method for reaching women throughout the community. PMID- 10537608 TI - Medical records as an alternative to self-report for measuring mammography utilization. AB - A pilot study assessed whether medical records were a viable alternative to self report for measuring mammography use in a population-based sample. Of 98 women contacted by telephone, 62 (63.3%) ultimately provided written consent to obtain their mammogram reports. Although all physicians complied with requests for records, an average of three physician contacts per woman were required and 87 percent of mammogram reports were located; therefore, records were available for only 56 percent of women contacted initially. This, coupled with the effort associated with obtaining the records, does not support the use of medical records as an alternative to self-report to measure mammography utilization in the general population. PMID- 10537609 TI - Spatial and temporal patterns in final amendments to provisional disease counts. AB - This article examines temporal and spatial patterns in the relationship between provisional and amended reports for Hepatitis A and Hepatitis B received from each state by the Centers for Disease Control and Prevention through the U.S. National Notifiable Diseases Surveillance System from 1980 to 1992. It demonstrates that, as the 1980s unfolded, the preliminary disease reports became less representative markers of final disease counts. Practitioners at the state and community levels need to be aware of the temporal and spatial instabilities in such provisional data if they are used to provide early warning of contemporary health aberrations. PMID- 10537610 TI - Assessing the prevention effectiveness of local Lyme disease control. AB - The effectiveness of any public health intervention is determined by its theoretical efficacy and by the level of engagement of the target population. A computer simulation model and basic epidemiologic concepts were used to estimate the effectiveness of interventions for preventing Lyme disease in a hypothetical community. The process for estimating numbers of Lyme disease cases prevented by each intervention is described. This assessment compares the effectiveness of alternative community-based prevention strategies, illuminates the limitations and distributive effects of interventions, and helps clarify available prevention options for community residents. PMID- 10537611 TI - Sharing information and experiences about maternal and child health data sets through the World Wide Web. AB - Maternal and child health programs face increasing requirements to collect, analyze, and disseminate information on current health status and needs of population groups. Data sets vary according to data elements. Population groups, organizational and agency boundaries, and other situation-specific characteristics. Until recently, the major venue for sharing information about various dimensions of data sets has been informal, often word-of-mouth, communications. The World Wide Web provides an opportunity to replace these slow and incomplete information exchanges with instant and comprehensive ones. This article outlines the development of a web site that includes descriptive information about 12 MCH data sets. PMID- 10537612 TI - Implementing information technology in the NHS: the role of narrative. AB - This paper analyses how groups use narratives in social processes of sensemaking and identity construction and in the pursuit and legitimation of their selfish interests. It does so through an examination of the narrativity of the experiences reported by the developers and users of an information technology (IT) system linking a haematology laboratory and a specialist haematology ward in a large acute hospital. The research contribution the paper makes is twofold. First, it illustrates the importance of group-level narratives in enacting organizational realities and especially in the social construction of IT systems. Second, it suggests that the narrative understanding of groups is a significant domain of organizational inquiry because it is through the spread and acceptance of their narratives that groups exercise their most profound influence. PMID- 10537613 TI - Cross-national comparison of capitation funding: the American, British and Dutch experience. AB - In this paper we review the performance of the capitation payment systems of three countries--the Adjusted Average Per Capita Cost (AAPCC) system used in the United States to reimburse Health Maintenance Organizations (HMOs) for insuring Medicare recipients, a somewhat similar system in the Netherlands which reimburses third-party payers for insuring the entire population and a weighted system utilized in Britain for regional funding. Our review revealed significant problems with the current version of the AAPCC formula as there is evidence of the biased selection of beneficiaries and actual losses to Medicare through its use. Furthermore, several studies show that the demographic adjusters utilized in the AAPCC formula are extremely poor predictors of future healthcare utilization relative to the potential of direct and indirect health status measures. The Dutch experience with capitated funding has been similar to that of the United States. While Dutch researchers have built on the work of their American counterparts they acknowledge that further work is needed before a fully functional system is implemented. Britain's weighted system has fulfilled its original mandate to redistribute healthcare resources based on population need but recent changes giving increased influence to age weighting could reverse some of these gains. A number of proposed improvements to these risk adjustment problems were reviewed including the development of diagnostic cost groups, the coexisting hierarchical conditions model and the use of community-rated high-risk pooling. The findings from this study can help others narrow the alternatives they need to consider when thinking of introducing capitation funding or refining already existing systems. PMID- 10537615 TI - Socioeconomic risk factors and population-based regional allocation of healthcare funds. AB - Using the notion of professional uncertainty a population-based proxy need measure for hospital services was developed. Its relationship with socioeconomic variables and Standardized Mortality Ratios (SMR) was investigated in an attempt to develop an adjustment factor for socioeconomic risk factors beyond age-sex adjustment to be used for a population-based healthcare funding formula for Alberta. The data used are 1990, 1991, 1992 vital statistics and hospital separation abstracts, 1991 census data and Refined Diagnosis Related Group (RDRG) case weights. Geographic units studied were the 26 federal electoral districts in Alberta using postal codes as a linkage geo-code between census and hospital utilization and death data. SMRs, age-sex standardized per capita hospital utilization and proxy need rates were derived and correlated with socioeconomic variables derived from the census files. It appears that the poor, the less educated and aboriginals need more hospital services than the affluent, employed and educated, confirming previous findings. The unemployed tend to need more but use fewer services while immigrants and non-white ethnics tend to need and use fewer services. The unemployed, less educated and non-white ethnics are associated with positive correlation with premature mortality (SMR based on deaths under age 75 years), while the employed, highly educated tend to live longer. In general SMRs have positive but very low correlations with utilization and need rates suggesting that SMRs should not be used for resource allocation. Stepwise multiple regression analyses showed that the percentages of unemployed, immigrants, non-whites, aboriginals and those with education less than grade 9 explain about 90% of the variation in age-sex standardized hospital utilization rates. Percentages of unemployed, non-white ethnics, residents with education less than grade 9 and aboriginals explained 71% of variations in age-sex standardized per capita proxy hospital service need measures. Based on the results of regression analyses, a SEAM (Socio-Economic Adjustment Multiplier) scale was developed for utilization (SEAM-U) and proxy needs (SEAM-N). In essence a SEAM is a set of relative value (RV) multipliers applicable to a provincial common per age-sex adjusted capita allocation value to account for the impact of socioeconomic risk factors on hospital service needs or utilization. Finally, the resulting regression equations derived from the 26 Federal electoral district data were applied to Alberta's health regions, regional SEAMs were derived, and the impact of such adjustment was assessed. PMID- 10537614 TI - Physician response to a change in Medicaid fees. AB - This study examines the effects of a change in Medicaid fees on the volume of physician services provided to beneficiaries. The data set includes price and volume at the procedure-level for Medicaid physician services in Texas in 1991, 1993, and 1995. The empirical analysis compares the volume of services provided to Medicaid participants before and after a 1992 change in reimbursement method. The results indicate that, over the period 1991 to 1993, the change in Texas Medicaid physician fees did not have a statistically significant effect on the volume of services provided. When measured over a longer period of time (1991 1995), however, volume increased significantly when price decreased, but, when price increased, there was no significant effect on volume. The results thus provide empirical support for the behavioural offset assumption underlying the switch to Medicare's Resource-Based Relative Value Scale (RBRVS) method of physician payment. A key policy implication is that reduced fees did not lead to a lower volume of physician services provided to Medicaid patients at least over the period of analysis. However, the new Medicaid fee schedule did not have the desired effect of controlling Medicaid expenditures on physician services. PMID- 10537616 TI - Level of analysis considerations in organizational citizenship behaviour research: an empirical investigation of individual and work group effects among hospital employees. AB - An implicit assumption in previous research is that the relationship between job satisfaction and organizational citizenship behaviour is an individual level phenomenon. However, due to the use of raw score correlation-based or related analyses, previous investigations have not shown empirical support for the individual level of analysis. This study empirically tested several relationships between job satisfaction (including facets) and a specific type of citizenship behaviour in order to determine whether such relationships were relevant for individuals or for the entire work group. Results indicate an overall lack of group-based effects. Instead, individual-difference effects represent the significant relationships found in the data. Several null effects were also obtained. These results go beyond the traditional approach to organizational citizenship behaviour research because group-based relationships have been explicitly rejected rather than simply assumed to be unimportant. Evidence for individual-differences was provided by testing for levels of analysis effects in terms of individuals and work groups. Future research should assess the generalizability of these results by including tests for levels of analysis. PMID- 10537617 TI - Practice brief. Retention of health information (updated). American Health Information Management Association. PMID- 10537618 TI - NCVHS (National Committee on Vital and Health Statistics) focuses on HIPAA. PMID- 10537619 TI - Performing a manual coding audit. PMID- 10537620 TI - Medical dictation: a new generation. PMID- 10537621 TI - Post-acute prospective payment: what you should know. PMID- 10537622 TI - Post-acute PPS: changing the way we code. PMID- 10537623 TI - Clinics go electronic: two stories from the field. AB - We hear a lot about computerized record system implementations in hospitals, but other settings are making the transition as well. How does the move to an electronic record system affect day-to-day operations in clinics? Our writers tell how a student health center and a family clinic did it--and how they're working now. PMID- 10537624 TI - ORYX: opportunity gained or lost? AB - For HIM practitioners willing to extend their knowledge and scope of responsibility, the Joint Commission's ORYX initiative will provide new and exciting opportunities. Your level of involvement will depend on your willingness to expand your knowledge base and accept a leadership role within your organization. For those willing to change the way they think about data analysis/outcomes, ORYX will provide some of the most important and unlimited career opportunities that will be, or have ever been, offered to HIM professionals. PMID- 10537625 TI - When bad documentation happens to good long term care facilities. PMID- 10537626 TI - Common (and uncommon) ground. PMID- 10537627 TI - Are HIM programs going the distance? PMID- 10537628 TI - Decimating duplicates. PMID- 10537629 TI - Leading through empowerment. PMID- 10537630 TI - Sign of the changing times: CPT 1999. AB - Although CPT 1999 contains fewer changes than in past years, coders should take some time to learn them by: familiarizing themselves with the new symbols + and [symbol: see text] reviewing Appendix A for a complete list of modifiers as well as modifiers used in the ambulatory surgery center hospital outpatient setting; reviewing Appendix E for a complete list of add-on codes; reviewing Appendix F for a list of modifier-51-exempt codes; consulting the excludes note found above code 69,990 to identify procedures exempt from the use of the new operating microscope code; examining the specific codes used to identify bronchoscopic procedures; reviewing the parenthetical notes found after code 15,001, directing the coder to also assign the appropriate code for lesion excision; reviewing the changes associated with the coding of destruction of lesions understanding the changes in immunization code assignment; consulting payers for specific reimbursement guidelines. PMID- 10537631 TI - Treading the rehab road. Interview by Anne Zender. PMID- 10537632 TI - Statistical thinking applied to everyday data. AB - Despite all the talk about "improvement" and "statistical thinking," our everyday work environments are horribly contaminated with poor statistical practice (even if it is not formally called "statistics"). The statistical methods our work requires are quite simple ... but initially quite counter-intuitive. Once grasped, however, they can provide a deeper understanding that will ensure better analysis, communication and decision-making. PMID- 10537633 TI - Expanding AMAP reduces redundancy, puts providers on same page for quality care. PMID- 10537634 TI - Pharmacy risk: is it in your future? AB - This article discusses how Sutter Health's affiliated medical groups and independent practice associations (IPAs) prepared for entering a pharmacy risk agreement. It should serve as a primer for those providers contemplating pharmacy risk and is written in lay terms. Providers who expect not just to survive, but to thrive, in a capitated environment must approach pharmacy risk agreements with extreme caution. PMID- 10537635 TI - Between a rock and a hard place: ethics in managed care and the physician-patient relationship. AB - At the heart of ethics in medicine is a solid physician-patient relationship. Managed care has operated to undercut this relationship. Rather than blaming managed care, however, one should identify and modify those social and economic forces that pressure managed care organizations (MCOs) to (1) undercut the physician-patient relationship and (2) engage in other unjust or unethical practices. Identifying these pressures does not let managed care off the hook for unethical practices. In fact, MCOs have both an ethical and a practical obligation to help define and support national health care legislation--to help reinvent their role in health care. PMID- 10537636 TI - Serving people with developmental disabilities in Medicaid managed care. AB - Concurrent with the sweeping changes in health care during the past decade, particularly in Medicaid financed health care, has been the reshaping of social policy toward people with developmental disabilities. The extent to which managed care entities match the themes now driving social services for people with mental retardation and other developmental disabilities (cerebral palsy, autism, etc.) is the extent to which they will be successful in serving this unique group of consumers of managed health care. The authors suggest a number of considerations for managed care organizations that increasingly serve significant numbers of this population. PMID- 10537637 TI - Practical implementation of globally priced episodes of care: lessons learned from Oxford's specialty management program. AB - Globally priced episodes of care afford payers a more efficient method of allocating risk and simultaneously accommodating the now permanent demand for choice and access at the point of service. Foreseeing the trend away from capitated delivery systems, in April of 1996. Oxford Health Plans began to invest considerable resources to implement global fees. When Oxford ran afoul of Wall Street and New York State insurance regulators (for reasons unrelated to the subject of this article), it was forced to abandon the new program. For the growing number of payers interested in pursuing similar contracting strategies, this article may serve as a useful source of information. PMID- 10537638 TI - Credentialing alternative medicine: a new challenge for managed care organizations. AB - Alternative Medicine is becoming mainstream healthcare. Credentialing alternative medical service providers is creating a new set of challenges for managed care organizations. PMID- 10537639 TI - Learning organizations: an introduction. AB - Many believe that successful companies beyond the year 2000 will all need to operate as learning organizations. Stimulating this vision have been Dr. Peter Senge of the Massachusetts Institute of Technology and Dr. James Milojkovic of Stanford University. As a learning organization, all members of a company will learn more about other parts of the organization and more about essential processes such as: helping each other learn, sharing, cooperating, leading, and participating in organizational decisions. This learning together will change existing stratifications, such as "bosses make decisions and employees implement the decisions without being consulted." A learning organization is a group of very different people who work together closely; bosses learn to treat each person as a responsible adult and hourly employees learn to participate in decisions. Learning organizations have a strong commitment to continual change. Each participant is expected to personally learn and grow. Everyone is expected to be open and share information, all while being tactful. The question that is continually before everyone in the company is "how can we do better?" PMID- 10537640 TI - Conversations on health care quality. PMID- 10537641 TI - Medical and behavioral health benefits: can they be equal? PMID- 10537642 TI - Crossing borders: considerations in delivering health insurance products and services. PMID- 10537644 TI - Disease management: when is it the right time? PMID- 10537643 TI - TennCare: Tennessee's answer to runaway Medicaid costs. AB - TennCare was implemented in Tennessee on January 1, 1994, as the state's replacement program for Medicaid. Created through a demonstration waiver under Section 1115(a) of the Social Security Act, TennCare is unlike any other state Medicaid program in the nation because it includes coverage of uninsured and uninsurable Tennesseans. By contracting with managed care organizations (MCOs) to administer the Medicaid benefit package to TennCare enrollees, the state virtually privatized the program and dramatically shifted the state's responsibility from one of total administration to an oversight function only. Amid provider lawsuits, MCO contract controversy, and general chaos surrounding the new program, the implementation of TennCare created one of the most significant health care events in the state's history. This article provides an overview of the TennCare program and its impact on enrollees, providers, and the state budget. PMID- 10537645 TI - Genetic discrimination in health insurance. PMID- 10537646 TI - Where's the money? Paying providers should be easy, but it's not working out that way. PMID- 10537647 TI - Has the medication revolution gone too far? A politically incorrect view of depression and its treatment. PMID- 10537648 TI - Restraints: are 'best practices' good enough? PMID- 10537649 TI - Stress and the workplace. Conflict management can prevent behavioral health problems. PMID- 10537650 TI - Quality in behavioral healthcare. Where do we look for inspiration? PMID- 10537651 TI - NCQA requirements: friend or foe? AB - In this time of intensified cost-containment pressures and low funding for behavioral healthcare services, organizations are particularly sensitive to externally imposed requirements that add expense and administrative burden. As a tradeoff, the contribution of such requirements to the overall quality of behavioral health services is vitally important. The requirements of the National Committee for Quality Assurance (NCQA) have stimulated many intensely felt reactions among a wide range of managed behavioral healthcare stakeholders, undoubtedly because of the substantial impact those requirements are creating. The reactions portrayed in this dialogue section, from representatives of the payor, purchaser, provider and consumer communities and from NCQA itself, indicate the ambivalence stakeholders have toward the quality-assurance movement. PMID- 10537652 TI - The HEDIS antidepressant measure. PMID- 10537653 TI - Implementing practice guidelines. PMID- 10537654 TI - Using evidence-based methods in a private practice. PMID- 10537655 TI - Selecting a clinical outcomes system. PMID- 10537656 TI - Science and the success of behavioral healthcare. PMID- 10537657 TI - Is there a middle ground? Challenges to the continuum of care threaten the value of services. PMID- 10537658 TI - A continuum of HIV services: focusing on the children. PMID- 10537659 TI - Improving patient satisfaction through unit-based team case management. PMID- 10537660 TI - Housing for infectious TB patients who are homeless: an alternative housing program. PMID- 10537661 TI - Dartmouth Atlas examines lack of prevention in US health care. PMID- 10537662 TI - Modeling of medical and surgical treatment costs of chronic pelvic pain: new paradigms for making clinical decisions. AB - Additional complexity has been added to the healthcare decision-making process by the socioeconomic constraints of the industry and a population that is increasingly educated about healthcare. As a result, decisions balanced on the basis of outcomes and economic realities are needed. This modeling of surgical versus medical treatment costs for chronic pelvic pain and endometriosis factors in the large number of women with chronic pelvic pain, direct and indirect costs of the condition, and clinical benefits, projected costs, and savings of the therapies. This process of calculation becomes an aid for decision making in the current healthcare system. PMID- 10537663 TI - Evaluation of Lovelace Health Systems chronic pelvic pain protocol. AB - Although laparoscopy has been considered the gold standard for the diagnosis of endometriosis, it often fails to detect the disease and provide lasting pain relief. Motivated by concerns for patient well-being, treatment efficacy, and cost containment, Lovelace Health Systems of Albuquerque, New Mexico, turned to the Lovelace Chronic Pelvic Pain Protocol, based on a chronic pelvic pain algorithm used to identify potential candidates for therapy with gonadotropin releasing hormone agonist (GnRH agonist). Since the protocol's introduction in January 1997, empiric therapy with GnRH agonist has proved beneficial to patients, physicians, and healthcare system budgets. PMID- 10537664 TI - The clinical and economic benefits of GnRH agonist in treating endometriosis. PMID- 10537665 TI - Focus on ED processes before making physical changes. PMID- 10537666 TI - Consider outsourcing IT projects when cutting-edge technology, specialized focus are needed. AB - Looking outside to meet information technology needs proves a smart way to avert extra staffing costs. Kaiser Permanente saves thousands each year by contracting out cutting-edge IT projects instead of hiring more full-time staff it doesn't need. Learn how the organization incorporates outsourcing and other temporary work methods into its IT staffing strategy. PMID- 10537667 TI - New study highlights ingredients for reengineering success. AB - Data benchmarks: A new study reveals specific elements to include in a reengineering program to boost its chances of success. The authors' review of the health care literature on reengineering reveals some startling discoveries, including the lack of consensus on defining reengineering and the fact that many hospitals are not finding the cost reductions they hoped for in the first years after completing their programs. PMID- 10537668 TI - System's internal grant program leads to clinical innovations. AB - Internal grant program inspires caregivers to dream up terrific clinical studies that improve care and post savings to the bottom line. BJC Health System in St. Louis has a long list of success stories from its novel quality improvement program. The one featured in this issue resulted in more than $100,000 in annual savings systemwide after a small change was made in the way suction catheters are used on ventilator machines in the ICU. PMID- 10537669 TI - Physicians take lead in the integration of medication and disease management. PMID- 10537670 TI - Putting evidence-based medicine to work: use of pocket guides in heart care. PMID- 10537671 TI - Technology and the future of medical equipment maintenance. AB - Maintenance of medical equipment has been changing rapidly in the past few years. It is changing more rapidly in developed countries, but changes are also occurring in developing countries. Some of the changes may permit improved maintenance on the higher technology equipment in developing countries, since they do not require onsite expertise. Technology has had an increasing impact on the development of medical equipment with the increased use of microprocessors and computers. With miniaturization from space technology and electronic chip design, powerful microprocessors and computers have been built into medical equipment. The improvement in manufacturing technology has increased the quality of parts and therefore the medical equipment. This has resulted in increased mean time between failures and reduced maintenance needs. This has made equipment more reliable in remote areas and developing countries. The built-in computers and advances in software design have brought about self-diagnostics in medical equipment. The technicians now have a strong tool to be used in maintenance. One problem in this area is getting access to the self-diagnostics. Some manufacturers will not readily provide this access to the owner of the equipment. Advances in telecommunications in conjunction with self-diagnostics make available remote diagnosis and repair. Since components can no longer be repaired, a remote repair technician can instruct an operator or an on-site repairman on board replacement. In case of software problems, the remote repair technician may perform the repairs over the telephone. It is possible for the equipment to be monitored remotely by modern without interfering with the operation of the equipment. These changes in technology require the training of biomedical engineering technicians (BMETs) to change. They must have training in computers and telecommunications. Some of this training can be done with telecommunications and computers. PMID- 10537672 TI - Tests show residential fire sprinklers will save lives. PMID- 10537673 TI - Hospital overcomes power problem. PMID- 10537674 TI - Hydrotherapy pool hygiene--the ultimate challenge. PMID- 10537675 TI - Act now in defusing the timebomb set to affect hospitals. PMID- 10537676 TI - Higher kitchen hygiene. PMID- 10537677 TI - Successful trial of telemedicine from a flying plane. PMID- 10537678 TI - The technical and financial impact of systematic maintenance and repair services within health systems of developing countries. PMID- 10537679 TI - A multiple-level analysis of hospital team effectiveness. AB - The purpose of this study was to identify factors at organizational, team, and individual levels that are most predictive of quality improvement team effectiveness. Numerous studies have been conducted on the use of teams in health care. The majority of these studies have focused on organizational-level issues. A few others have focused on team-level issues or individual-level issues. The authors believe that successful use of teams requires a more integrated approach. This study addresses this need by providing a multiple-level analysis of teams in three hospitals. Structured interviews were conducted with hospital administrators as well as quality improvement representatives and team leaders, and written questionnaires were administered to team members. Eight factors were related significantly to the effectiveness of team effort: frequency of team meetings, hours per week conducting team activities, willingness of members to serve on team, selection method of team members, communication of team plans, team member position/composition, team leader performance, and facilitator performance. PMID- 10537680 TI - A supervisory level self-directed work team in health care. AB - Nationwide changes in health care delivery precipitated shifts in management within health care facilities. Connecticut experienced growth of 24 percent in managed care contracting between 1992 and 1994, a result of private sector initiatives set in motion by the proposed Clinton Administration health care plan. The shift from fixed rate to capitated payment structures was expected to have a negative impact on health care facilities and hospitals in particular. Further, it was anticipated that managed care would push services out of hospitals and into price-competitive, freestanding facilities. The response of Hospital for Special Care included development of a supervisory self-directed work team. PMID- 10537681 TI - Organizational culture: the role of management and supervisors. AB - There is much more to organizational culture than the diversity issue. Establishing or changing a corporate culture is challenging for employers, management, and supervisors. In this article, the authors describe the importance of corporate culture, explore the roadblocks to improving it, and relate how successful managers and supervisors cope with this challenge. PMID- 10537682 TI - Managing the temperamental employee. AB - In today's fast-paced, competitive, and quality-centered health care environment, dealing effectively with a temperamental employee and the resultant impact on the organization presents particular challenges. Considerations include the requirement of timely action in order to alleviate the real problems of unprofessional behavior in the workplace. This article discusses the major issues associated with such situations, including why there often is a reluctance for management to take action, the consequences that are accentuated by inaction, prescriptions for preventing the problem, and steps to be taken in dealing effectively with the temperamental employee. PMID- 10537683 TI - Forms management for the health care supervisor. AB - The creation of necessary, efficient forms at the lowest possible cost is possible. However, it requires a willingness to diligently perform forms analysis, design, and control. Untrained forms designers may know what items they need on a form, but they do not necessarily know how to arrange items on a form or how to select the physical properties of the form. This article addresses the salient points of forms analysis, design, and control. PMID- 10537684 TI - Riding the waves of change in health system material management: team building in times of uncertainty. AB - Within the health care industry, reorganization brings new challenges and opportunities for improving work-related processes. This article examines one Midwestern health system's response to the systemic change that may occur after restructuring. The consolidation of diverse and separate material management departments within one integrated material management division prompted a concerted team development and training effort. In this model, management and non management staff worked collaboratively to achieve a unified work group that will continue to evolve and grow with the needs of the organization. PMID- 10537685 TI - Are hospitals facing a critical shortage of skilled workers? AB - A nationwide survey of health care human resource managers reveals that hospitals are experiencing a critical shortage of qualified job applicants in certain job categories. Computer systems personnel, registered nurses, pharmacists, and therapists are especially in short supply. Jobs that require only minimal training as well as those not directly involved in patient care are filled more easily. According to the survey respondents, shortages of qualified personnel negatively impact hospitals' operations in a variety of ways. To combat shortages of qualified job applicants, hospitals are employing a variety of tools to attract workers. Signing bonuses, above-market compensation, flexible work schedules, and myriad perks are being used to entice and retain qualified workers. PMID- 10537686 TI - The essentials of compassionate downsizing. AB - Downsizing remains a constant fixture of the organizational landscape. The process causes severe emotional stress on departing employees. It places functional and emotional strain on survivors. This article provides practical advice for executives and supervisors on how to minimize the traumatic effects of these staff reductions. It also discusses how staff reductions can be more efficient. Emphasis is on the importance of training following a downsizing. PMID- 10537687 TI - Key indicators of nursing care team performance: insights from the front line. AB - Self-directed nursing care teams (NCTs), comprised of a registered nurse (RN) team leader and two or more non-licensed caregivers, are a key feature of patient focused care reengineering. In well-functioning teams: (1) role overlap routinely occurs; (2) team members express satisfaction with interpersonal communication; and (3) team members express the belief that their shared purpose--and that of the health care organization--is the patient. Focus groups of team members in two case study hospitals expressed generalized dissatisfaction with performance on all three key imperatives of well-functioning teams, but also generally agreed that team delivery of bedside nursing care can work if properly managed. Based on study findings, recommendations for process improvement are made. PMID- 10537688 TI - A working manager's guide to effective and legal employee selection interviewing. AB - Employment interviewing as we know it today is an essential process but one fraught with potential traps and legal pitfalls. Overall, it is a far-from perfect means of selecting employees, but it the best such means available. Effective interviewing requires thorough preparation, including knowledge of how to seek out the most helpful kinds of information available, complete information about the position as it currently exists, and detailed knowledge of what kinds of questions can or cannot be asked legally. Effective interviewing also depends on the development of one's ability to seek out intangible and factual information and use all that is learned, recognizing that the well-cultivated "gut-feel" is fully as important as "facts" in evaluating an employment applicant. PMID- 10537689 TI - Federal district court explains, explores silent PPO. HCA Health Services of Georgia Inc. v. Employers Health Insurance Co. PMID- 10537690 TI - Dispute over Pap smear specimen slides resolved by Connecticut Supreme Court. Cornelio v. Stamford Hospital. PMID- 10537691 TI - "Web revolution" is changing healthcare. PMID- 10537692 TI - Catholic health ministry in transition. Church's unique vision remains stable in shifting healthcare landscape. PMID- 10537693 TI - Preserving our Catholic identity. If the health ministry is to remain faithful to its basic elements, it must first spell them out. PMID- 10537694 TI - Managed care: devils, angels, and the truth in between. PMID- 10537696 TI - The conversation underneath the truth in between. PMID- 10537695 TI - PSO helps restore trust in managed care. PMID- 10537697 TI - We need a realistic approach to healthcare financing. PMID- 10537699 TI - Catholic healthcare and the common good. An ancient concept can bring the transforming power of grace into the public healthcare debate. PMID- 10537698 TI - Building character for a new era. New temptations reveal the limitations of professional healthcare codes. PMID- 10537702 TI - Seeking leaders for the future. Interview by Ann Stockho. PMID- 10537700 TI - The common good and healthcare policy. Healthcare is a social construction for the good of all. PMID- 10537701 TI - Of what good is the "common good"? This abstract notion has the power to transform society. PMID- 10537703 TI - After the ink dries. Hospital encourages new partners to develop cultures compatible with Catholic health ministry. PMID- 10537704 TI - Toward a compensation philosophy. Leaders discuss ministry pay, benefits. PMID- 10537705 TI - Five common marketing mistakes. PMID- 10537706 TI - Community networks. Partnerships between Catholic Charities and Catholic healthcare organizations. Samaritan Place, Knoxville, TN. PMID- 10537707 TI - The thin line of life. PMID- 10537709 TI - How to spot the patient who's faking it. PMID- 10537708 TI - Telling your story: the importance of communications. PMID- 10537710 TI - What if a bankrupt insurer left you bare? PMID- 10537711 TI - Electronic medical records make sense--at last. PMID- 10537712 TI - Switching from paper to computerized charts. PMID- 10537713 TI - A shopper's guide to practice management software. PMID- 10537714 TI - We could treat the world with what we waste at home. PMID- 10537715 TI - Malpractice forecast. Clouds threaten, but sunshine breaks through. PMID- 10537716 TI - Will alternative-medicine referrals get you sued? PMID- 10537717 TI - Your newest competitors: alternative-medicine networks. PMID- 10537718 TI - Could you dig a financial hole this deep? PMID- 10537719 TI - Three specialists couldn't help her--but the patient sued me. PMID- 10537720 TI - She strengthens people's health--and their faith. PMID- 10537721 TI - Sloppy records--the kiss of death for a malpractice defense. PMID- 10537722 TI - Telling a prospective employer about malpractice claims. PMID- 10537723 TI - Why more doctors aren't testing for HIV. PMID- 10537724 TI - Alas, I've developed a bedside manner. PMID- 10537725 TI - Our senior partner isn't retired. He's "rehired". PMID- 10537726 TI - Would locum tenens practice suit you? PMID- 10537727 TI - Compassionate care--or murder? PMID- 10537728 TI - Wrestling with the managed care octopus, Part 4. Get plans to pay in full, on time. PMID- 10537729 TI - Repaying his town for a kindness long ago. PMID- 10537730 TI - Wrestling with the managed care octopus, Part 3. Get insurance insurance authorizations faster. PMID- 10537731 TI - A crisis made our practice a family. PMID- 10537732 TI - Doctor policing: too tough? Too easy? Medical boards: facing a disciplinary dilemma. PMID- 10537734 TI - Doctor policing: too tough? Too easy? Impaired physicians: giving rehab programs a new look. PMID- 10537733 TI - Doctor policing: too tough? Too easy? Peer review: breaking the code of silence. PMID- 10537735 TI - Doctor policing: too tough? Too easy? I'm a physician, and I'm a drug addict.... PMID- 10537736 TI - Don't put up with a partner's lousy penmanship. PMID- 10537737 TI - Don't send a patient running to a lawyer. PMID- 10537738 TI - Malpractice trials are more theater than law. PMID- 10537739 TI - Why I settled a malpractice suit I thought I could win. PMID- 10537740 TI - Your worst career move? A contract signed in haste. PMID- 10537741 TI - Will violence end patients' access to abortion? PMID- 10537742 TI - The best Web sites for doctors. PMID- 10537743 TI - Recommended resources on complexity theory. PMID- 10537744 TI - Culture in chaos: the need for leadership and followership in medicine. AB - The health care industry is changing at a dizzying pace and most of its players are struggling to maintain some form of the status quo. But resisting change will not prove fruitful--ultimately, it will rob physician executives of the opportunity to be architects in designing a new, more efficient health care system and their role in it. Because health care is a complex adaptive system (CAS)--change occurs rapidly and events are unpredictable--the old command and control style of leadership and a linear way of interpreting events is too rigid and, therefore, an ineffective model for guiding change. Complexity science offers insights about leading for change. In CASs, changes emerge in response to environmental demands for adaptability. Since the nature of these demands is unpredictable, the role of leadership is to manage the relationships and context out of which these changes emerge. A leadership style is called for that leads to purpose, makes positive changes by influencing context and relationships, and takes followers to a better place. PMID- 10537745 TI - The five "S" levels of enterprise health. AB - Whether pride, necessity, or inattention is at the root, some "slowly boiling" physicians find themselves working harder for fewer compensations of all sorts, and may not be fully cognizant of their circumstances. This article helps to diagnose and manage the health of physicians' practices and/or related enterprises. There are five levels of enterprise health, ranging from success (S 1) to shutdown (S-5), that serve as weather vanes about how the enterprise is adapting to changes in its environment. How should physicians respond to chaos and the threats of deteriorating enterprise health? A five-step approach is offered: (1) Discern what is important; (2) place and keep your program in alignment with those patient interests that will enhance your enterprise viability; (3) keep score with an internal balanced scorecard; (4) manage and shepherd your resources in a manner that demonstrably adds value to patient care; and (5) know the score and use it. PMID- 10537746 TI - Equity, equity, that's our cry. Interview by Richard L. Reece. AB - On February 11, 1999, Richard L. Reece, MD, interviewed J.D. Kleinke to talk about his new book entitled Bleeding Edge: The Business of Health Care in the New Century. A medical economist and author living in Denver, Kleinke advocates a true partnership between hospitals and physicians--a marriage with both parties contributing equally to the relationship. He believes that "physicians and people who are running the administrative infrastructures of hospitals and other facilities need to recognize that they are equal partners in a death struggle against the insurers for ultimate control of the premium and the consumer." Though physicians are sure to balk at the suggestion that they become "captive" to the hospital, Kleinke explains that, "captivity is a necessary condition before they can work functionally together and take on managed care and contract directly with consumers, employers, and the government." Kleinke discusses five trends that he explores in his book: risk assumption, consumerism, consolidation, integration, and industralization. PMID- 10537747 TI - Global theory and the nature of risk, Part I. How orthodox managed care stifles innovation. AB - The growing awareness that managed care is rapidly unraveling has not only produced a good deal of alarm, but also a call for prognostications regarding the future. Unfortunately, old habits die hard, and wedded ideologies die even harder. Instead of paving the way for innovation, most managed care pundits refuse to read the tea leaves properly and acknowledge that the orthodox regime is irretrievably comatose. Not understanding the fundamental flaws inherent in the old model, many persevere with rehashed predictions that only echo the very non-starters that got us in the present jam in the first place. Managed care has so far focused its energies on integrating the wrong objects, insurance and care, with all the predictably bad effects. Part 2 of this article will explore what this means and introduce the global theory of managed care as an alternative vision. Global theory lays a new foundation based on a more sound microeconomic model of risk, bifurcated markets, global fees for integrated episodes of care, and most important of all, patient/physician sovereignty. PMID- 10537748 TI - Health care trends, Part I. The promise and perils of evidence-based medicine ... panel discussion. AB - Numerous studies have demonstrated that there are wide variations in the way physicians manage similar patients. This suggests that an evidence-based approach could lead to better outcomes with less cost. But practicing evidence-based medicine requires new skills, such as using computerized databases and applying the rules of evidence to primary and integrative studies in the medical literature. The progress of evidence-based medicine will depend in large measure on how quickly these new skills can be developed and integrated into the practice environment. Here's how six experts see the promise and the perils of evidence based medicine, now and in the new millennium. Part 2 of the panel discussion will explore the new provider team, which includes nurses and, more recently, pharmacists, who are collaborating with physicians to provide disease management and drugs therapy management services. PMID- 10537749 TI - Newspapers foretell health care's future. AB - This article is based on a two-months snapshot (November 1998 to January 1999) of newspaper articles addressing various health care issues. Newspaper contents reflect the changing market share of competing societal concerns. Health care issues, particularly cost and choice, now preoccupy the American people. Health care trends percolate bottom-up through the pages of newspapers, not top-down from Washington, D.C, policymakers, or health care executives. By reviewing these articles, the author provides a big picture view of the prevailing and emerging health care trends. From the new thrust of consumerism and the public backlash against managed care organizations to the demise of HMOs and PPMCs, these observations signify not only the concerns that are bubbling to the surface but also the direction that health care is headed. Consumers are in the driver's seat and physician executives need to provide them with evidence of the value they desire--and understand what they perceive as value. PMID- 10537750 TI - Physician anger, Part 2. More on the dance of anger. AB - This article is a follow-up to an interview with Charles Dwyer, PhD, which appeared in the 1999 March/April issue of The Physician Executive. He described how physician executives can change the perceptions of today's beleaguered physicians and help them cope with change. We then asked him for some hands-on strategies to deal with physician anger, fear, and resentment. After much contemplation on providing a list of "fixes" that will restore each of us to a state of greater satisfaction, Dr. Dwyer concludes that there are no generalizable solutions because there are too many variables that come into play in each organization, individual, or group. Attending to the self can provide both individual rescue from these turbulent times and the best hope for changes in the system from which patients and health care providers can benefit. If physicians are to regain their power and maintain, or even improve, their quality of life, clearly changes are called for. And these are changes that require persistent effort and uncomfortable adjustments. PMID- 10537751 TI - The water is wide. AB - Trying to predict what cardiology will look like in 10, 25, or 50 years is an almost absurd act of imagination. Perhaps even more than most branches of medicine, it has been changing so fast that it is hard to recognize from year to year. When we contemplate the changes that cardiology and the rest of health care are facing, we know that none of these changes will ever be "just technical." They will all be deeply human, and each one will be very difficult for some people. As people and organizations, we all come to that point where we have to change. Where we are is not working, we can't go on like this, we have to do something different. After boldly setting off to go someplace new, after some basic work is done to escape the original presenting problem--we reach the Great River. This is a point at which small, incremental changes won't do and we truly need to see ourselves differently. There is no bridge, no easy way across. This is where we must gather our resources and courage, remind ourselves of why we came this far. Yet many of us don't, and settle for something comfortable rather than cross that wide, turbulent river. PMID- 10537752 TI - When to counter and when not. AB - It's a booming job market. Currently, job hunters are excited by the opportunities to get all they can, and headhunters and hiring organizations are pressured to find the best recruits. When the market is overheated, as it is, both candidates and recruiters are tempted to trust each other and not sweat the details. They are also tempted to believe that "a good candidate can fit in anywhere." It's time to share some cautionary tales that apply to job hunting physicians. The litany of mistakes extreme prosperity induces are described, including: (1) sloppy, superficial research, (2) rushing the process, (3) relying on things to "work out," and (4) soft peddling performance expectations. PMID- 10537753 TI - Characteristics of good mentors. AB - Physician executives who have advanced in their careers have had people who helped them. Six successful physician executives were interviewed about how mentors encouraged, taught, and helped them grow. Most agreed the term role model was more comfortable to them than the word mentor and that they only recognized these people as mentors when they looked back on their career paths--not at the time the interactions were happening. Sometimes they were role models and sometimes they were just the right person with the right information at the right time. Most were located in the person's city and organization and seen daily, but some were in another part of the country and seen on occasional visits but regularly talked to each other on the phone. Generally they were friends who created safe environments for learning, were protectors, gave specific feedback, viewed problems from a different angle, and stretched the thinking of those who sought their advice. PMID- 10537754 TI - Becoming a company person. AB - Physicians face a management challenge in adjusting to being part of a larger organization. By nature, physicians are not "company people." For physician executives, whose instincts can still lean toward being ferociously independent and individualistic--therein lies the struggle. Here are a few suggestions to help physician executives make the adjustments that are required of a "company person": (1) Be in the right place; (2) abandon critical/negative thinking; (3) focus on the issues; (4) maintain solidarity; and (5) make your boss look good. PMID- 10537755 TI - Cost-effectiveness and coverage policy. AB - Cost-effectiveness analyses have become a pervasive element of health care. But they have not had a major impact on medical coverage policy. The challenge of implementing cost-effectiveness as a medical coverage criterion is related to the following issues: (1) Contract language does not include cost-effectiveness as a coverage criterion; (2) cost-effectiveness analyses often take the societal, population-based perspective, while health care is delivered on an individual basis; (3) there is no standard methodology for cost-effective analysis; (4) there is no explicit cut-off between cost-effective and cost-ineffective; and (5) cost-effectiveness analyses are not time sensitive. PMID- 10537756 TI - Saving Medicare: premium supports. AB - Medicare, the universal health care program for the elderly and the disabled, is pending bankruptcy by the year 2008. Efforts to ensure its fiscal solvency are underway. The Bipartisan Commission on the Future of Medicare formed by the Balanced Budget Act of 1997 recently debated a bold new idea to transform this important program. There is some concern that this concept will not solve the problem without new funding. In addition, there are those who believe this is the time to correct one of Medicare's major flaws--the lack of a prescription benefit. Congressional action over the next several months will be critical to saving this program for future seniors. PMID- 10537757 TI - Nonlinear systems theory in medical care management. AB - Health care is provided to biological, not mechanical systems. The focus of attention is a human being, not an automobile or clock. Biological systems are highly complex and their behavior can be nonlinear. By understanding this difference, we can begin to answer the question of how can we deliver efficient and effective clinical care, while controlling spiraling costs. This article describes the behavior of complex systems and explores what has been learned about managing such systems in non health care fields. The author translates that learning into the health care setting and proposes a new model for the health care environment, based on the principles of complex adaptive systems. The approach advocated is to design models that can learn and adapt, and then to implement them with skill. Such models should include the elements of roadmaps, decision aids, and continuous improvement. PMID- 10537758 TI - Bibliography current world literature. Glaucoma. PMID- 10537759 TI - The role of medical therapy in the rank order of glaucoma treatment. AB - There is no consensus as to how treatment of open-angle glaucoma should be initiated. Medical therapy, laser trabeculoplasty, and filtration surgery each have their advocates as the best modality for initial treatment. The advantages and disadvantages of these three options are discussed here. It is pointed out that there may not be a single answer as to how to begin treatment. Patient factors such as age, general health, and stage of glaucoma as well as various socioeconomic and technologic factors may lead to different recommendations for different patients. At present, most patients with open-angle glaucoma are started on medical therapy. beta-Blockers are used most commonly as the initial drug of choice, but latanoprost and brimonidine are now being recommended by many as alternate first-line therapy. PMID- 10537760 TI - The risk profile of glaucoma filtration surgery. AB - Glaucoma surgery has evolved over the past 30 years from the full-thickness procedure to the guarded filtration procedure. However, many of the risks and complications attendant with the full-thickness procedure, including endophthalmitis, hypotony, cataract progression, and filtration failure, continue to plague the glaucoma surgeon performing the guarded filtration procedure, although at lower incidences. With proper modification of technique, such as with postoperative bleb titration and use of adjunctive antifibrotic therapy based on prognosticators for failure, the success rates of trabeculectomy reoperations can approach those of primary trabeculectomy. Such risk factors for failure include African-American race, higher preoperative intraocular pressures, previously failed filters, younger age, and uveitic and neovascular glaucomas. In this paper, we review a number of studies that analyze the risks, complications, and long-term results of glaucoma filtration surgery and discuss different surgical recommendations based on risk factors for failure as well as for performing concomitant cataract and glaucoma surgery. PMID- 10537761 TI - The contribution of vitreoretinal surgery to the management of refractory glaucomas. AB - Vitreoretinal techniques, although not new, are only gradually being recognized in the management of refractive glaucoma. Besides rare but established indications for vitrectomy, as in malignant glaucoma, vitreoretinal surgery may become even more valuable by offering an alternative outflow route to the choroid that remains open and functional. Vitreoretinal surgery in neovascular glaucoma is primarily meant to be antivasoproliferative surgery. Vitrectomy-lensectomy accesses the eye for panretinal laser coagulation, the only effective treatment of ocular neovascularization so far. Vitrectomy is a way of interfering with the pathogenesis of ocular new vessel formation, which otherwise inevitably leads to phthisis. PMID- 10537762 TI - The surgical management of leaking filtering blebs. AB - Leaking blebs may be encountered in the early postoperative period, or months to years after filtering surgery. Early postoperative bleb leaks are most often related to surgical trauma to the conjunctiva and can be avoided by careful surgical technique. Spontaneous late bleb leaks occur more frequently in glaucoma filtering surgery following adjunctive use of antimetabolites and full-thickness procedures. As we endeavor to achieve better long-term success with filtering surgery, antimetabolites have gained increasing popularity. With this change in clinical practice, a higher rate of bleb leaks is being recognized. These leaks may be uncomplicated or may be associated with sight-threatening complications such as endophthalmitis. The plethora of treatment options for bleb leaks described in the literature reflects the widespread nature of this problem. It also reflects the failure of any one particular approach not only to resolve bleb leaks but also to prevent their recurrence. This paper reviews the contemporary surgical management of leaking blebs and formulates a practical approach to their management. PMID- 10537764 TI - The diagnostic value of automated flicker threshold perimetry. AB - Automated perimetry techniques have advanced from the standard white-on-white threshold perimetry to a myriad of perimetric models. These models include motion detection, frequency-doubling contrast sensitivity, and spatial contrast sensitivity perimetry. The research findings for the more popular of the automated perimetry models, in particular those of blue-on-yellow and critical fusion frequency perimetry, are discussed and compared with findings for flicker threshold perimetry. Flicker threshold perimetry demonstrates resistance to such factors as test variability and retinal image blur and has great promise in its ability to detect early visual field loss in the presence of primary open-angle glaucoma. PMID- 10537763 TI - Glaucoma genetics: where are we? Where will we go? AB - The understanding of the genetic basis of the glaucomas has advanced rapidly. Mutations in the myocilin gene (previously known as TIGR) at the GLC1A locus on chromosome 1q21-q31 occur in a subset of patients with juvenile- and adult-onset primary open-angle glaucoma. Five other genetic localizations for primary open angle glaucoma have now been reported. In patients with primary congenital glaucoma, mutations have been found in the CYP1B1 gene on chromosome 2p21. At least one other locus for primary congenital glaucoma is mapped. In the developmental glaucomas, mutations in the PITX2 gene on chromosome 4q25 have been associated with Rieger syndrome, iris hypoplasia, and iridogoniodysgenesis. A second locus for Rieger syndrome resides on chromosome 13q14. Mutations in the FKHL7 gene on chromosome 6p25 have been described in patients with Axenfeld Rieger anomaly. A new ocular finding of glaucoma in pedigrees with the nailpatella syndrome has been described, and mutations in the LMX1B gene on chromosome 9q34 are now known to underlie nail-patella syndrome. Two loci for the pigment dispersion syndrome have been mapped. This paper provides an overview of recent literature, summarizes developments in glaucoma genetics, and addresses their potential relevance to the clinical management of glaucoma. PMID- 10537765 TI - The diagnostic significance of the multifocal pattern visual evoked potential in glaucoma. AB - The concept of objective perimetry is an exciting one because it strives to assess glaucoma damage without relying on psychophysical testing. The recent introduction of multifocal stimulus recording has enhanced our ability to examine the human visual field using electrophysiology. A multifocal pattern visual evoked potential can now be recorded, testing up to 60 sites within the central 25 degrees. The test requires only that the subject fixate on a target, while a cortically scaled dartboard pattern stimulus undergoes pseudorandom alternation within each of the test segments. In its present configuration the test requires at least 8 minutes recording time per eye. Modified bipolar electrode positions are required to ensure that adequate signals are detected from all parts of the visual field. In glaucoma patients, pattern visual evoked potential amplitudes have been shown to reflect visual field loss with reduction of signal amplitude in the affected areas. This technique represents the first major step toward objective detection of visual field defects in glaucoma. PMID- 10537766 TI - Glaucoma drainage devices: pros and cons. AB - Glaucoma drainage devices are an option in the management of complicated glaucomas that carry a high risk of failure from conventional filtering surgery. Examples include the glaucomas associated with aphakia or pseudophakia, neovascular glaucoma, and glaucomas associated with trauma, uveitis, epithelial downgrowth, iridocorneal endothelial syndrome, vitreoretinal disorders, and penetrating keratoplasty. Modifications in the various implant designs have been developed to limit the occurrence of postsurgical complications such as hypotony, serous and hemorrhagic choroidal detachment, tube and plate avulsion, tube exposure, and corneal endothelial damage. PMID- 10537767 TI - Glaucoma. PMID- 10537768 TI - The potential of neuroprotection in glaucoma treatment. AB - Visual field loss in glaucoma is due to death of retinal ganglion cells. Reducing or slowing down the loss of ganglion cells in glaucoma, a concept known as neuroprotection, would appear to be the only way forward. This does not imply that treatment of risk factors, such as elevated intraocular pressure, must not be continuously implemented. In this paper we point out that very little is known about the mechanisms of ganglion cell death in glaucoma and that data derived from studies on the "ideal animal model for glaucoma" must not be overemphasized. We also propose that the death processes of neurones in various diseases are fundamentally the same but vary in cause. Experimental data show that the death rate of neuronal populations is dependent on the impact of the insult and that neuroprotectants are more likely to benefit a patient in diseases in which the neurones die slowly, as in glaucoma, than in a disease in which the death of a set of neurones is rapid. We conclude that if a putative neuroprotectant can be administered in such a way that it reaches the retina in appropriate amounts and has insignificant side effects, it is likely to attenuate ganglion cell death and thus benefit the glaucoma patient. PMID- 10537769 TI - Glaucoma surgery: are there new perspectives in perioperative pharmacology? AB - The literature directed at perioperative pharmacologic advances in relation to glaucoma filtration surgery is reviewed. The successful use of subconjunctival anesthesia demonstrates a new alternative in preoperative glaucoma surgical anesthesia. The intraoperative use of the antimetabolites mitomycin C and 5 fluorouracil in both traditional filtration and glaucoma drainage implantation surgery has been expanded. The use of the antifibrinolytic agents urokinase and recombinant tissue plasminogen activator adds a new and controversial dimension to postoperative pharmacologic therapy. PMID- 10537770 TI - Perspectives in the medical treatment of glaucoma. AB - Many advances in medical therapy for chronic glaucoma have taken place in the past few years that have altered previous concepts of stepwise medical therapy for glaucoma. beta-adrenergic blockers are still the most common medicine prescribed as monotherapy. However, latanoprost and brimonidine are often given as monotherapy as well as early adjunctive therapy. In addition, newer treatments that are used as early adjunctive therapy are dorzolamide, brinzolamide, apraclonidine, and the combination products, consisting of dorzolamide/timolol maleate and pilocarpine/timolol maleate. Older adjunctive treatments are now often prescribed as late adjunctive therapy including pilocarpine, epinephrine compounds, and acetazolamide. The extent of maximum tolerated medical therapy and the decision to perform either laser or conventional filtration surgery depend on the physician's judgment and on the patient's needs and preferences. In the future, newer medical therapies that may protect the health of the optic nerve directly could be developed including blood flow-based, neuroprotective, and genetically based agents. PMID- 10537771 TI - Bibliography. Current world literature. Retina and vitreous disorders. PMID- 10537772 TI - Results of screening low-birth-weight infants for retinopathy of prematurity. AB - Retinopathy of prematurity (ROP) continues to be an important cause of potentially preventable blindness worldwide. The pattern of visual impairment from ROP in some middle-income countries--high rates affecting larger and more mature infants--resembles that seen in more developed countries two decades ago and has been called a "third epidemic" of the disease. Expert bodies in the United Kingdom and the United States have recently issued new guidelines for screening for ROP that utilize both birth weight and gestational age criteria. Studies in both countries suggest these criteria might be further revised to decrease time spent on screening without missing any significant disease. Population-based follow-up studies of extremely preterm infants suggest that although more preterm infants are surviving, with adequate screening and treatment, rates of blindness from ROP may be declining. Further information on the longer-term impact of ROP comes from a number of studies and particularly the Multicenter Trial of Cryotherapy for Retinopathy of Prematurity (CRYO-ROP) study. The risk of both myopia and strabismus is increased with any and each higher stage of ROP. Evidence is emerging that laser therapy for threshold disease may be associated with better visual outcome than cryotherapy, although complications following the former remain a concern. The fight against ROP may be enhanced by new information on the pathogenesis, including possible genetic predisposition and the role of vascular endothelial growth factor. PMID- 10537774 TI - Fundus lesions of adenomatous polyposis. AB - Congenital hypertrophy of the retinal pigment epithelium (CHRPE) is the most frequent extraintestinal manifestation of familial adenomatous polyposis. Present in 70% of families with familial adenomatous polyposis, CHRPE is a highly reliable and early marker of the disease. Studies over the past 5 years have addressed the histologic characteristics of the pigmented fundus lesions, the definition of universal positive fundus criteria, and mostly the genotype phenotype correlation. Indeed, the position of the mutation site of the APC (adenomatous polyposis coli) gene on chromosome 5 influences the retinal expressivity because CHRPE is present only if the mutation is located between exons 9 and 15. In CHRPE-positive families, fundus examination is simple, noninvasive, reproducible, inexpensive, and allows early detection of the mutant gene carriers. Knowing the CHRPE status of patients in a family with familial adenomatous polyposis helps to identify constitutional APC mutations. The combination of genetic analysis and fundus examination offers a 100% diagnostic predictability. PMID- 10537773 TI - Ultrasonography of metastases and melanomas of the choroid. AB - Choroidal metastases and melanomas are the most common intraocular neoplasms. Choroidal metastases may appear clinically similar to other amelanotic tumors, and the diagnosis may be difficult in the absence of a history of extraocular malignancy. Ultrasonography, using A- and B-mode criteria, offers an opportunity for high accuracy in the diagnosis of choroidal tumors. However, a considerable overlapping of ultrasonographic parameters, such as tumor solidity, vascularity, and choroidal excavation, has been demonstrated for choroidal melanomas and metastases. We have found that choroidal metastases are characterized by a significantly lower height-to-base ratio than melanomas, whereas reflectivity is significantly higher in metastases. Thus the combined use of height-to-base ratio and reflectivity provides a highly significant differentiation between choroidal melanomas and metastases. This observation has further improved the diagnostic value of ultrasonography in the differentiation of choroidal tumors. PMID- 10537775 TI - Retinal detachment associated with excimer laser. AB - Recent reports of retinal pathology associated with laser refractive surgery are documented. The overall risk of retinal disease following photorefractive keratectomy or laser-assisted in situ keratomileusis is discussed, and the potential causal associations between anterior segment laser and posterior segment pathology are assessed. PMID- 10537776 TI - Choroidal neovascularization in younger patients. AB - Choroidal neovascularization (CNV) is the most common cause of legal blindness in older adults in the United States. The most common cause for CNV in this age group is age-related macular degeneration, a condition manifesting with drusen (particularly soft drusen) and pigmentary alterations in the macular region. CNV can occur in younger people (< 50 years), who usually do not have conspicuous drusen or pigmentary abnormalities. In this age group CNV may occur as a secondary manifestation of many inherited and acquired conditions such as angioid streaks, high myopia, trauma, choroidal tumors, familial macular dystrophies, and inflammatory retinochoroidopathies. Occasionally, CNV in young people has no apparent antecedent cause, and these cases are termed "idiopathic CNV." This review examines the common reasons for CNV in young adults, with reference to some of the older literature as well as to recently published papers. PMID- 10537777 TI - Clinical applications of optical coherence tomography. AB - Optical coherence tomography is a new diagnostic tool for high-resolution imaging of ocular tissues. Ocular coherence tomography produces cross-sectional images of the retina with a longitudinal resolution of 10 microns. This provides the most highly resolved retinal images in vivo when compared with other available techniques. It has been used to study the anatomy and pathogenesis of various ocular disorders affecting the posterior segment. These include vitreomacular traction syndrome, macular hole, retinoschisis, macular edema, central serous chorioretinopathy, subretinal neovascularization, age-related maculopathy, optic nerve disorders, and nerve fiber layer evaluation in glaucoma. PMID- 10537778 TI - The present role of indocyanine green angiography in ophthalmology. AB - Recent developments in the clinical application of indocyanine green angiography have mainly concerned refining its role as an adjunct to fluorescein angiography in detecting and guiding the treatment of choroidal neovascularization. We now have a better understanding of the different patterns of abnormal hyperfluorescence seen with indocyanine green angiography in eyes with both wet and dry forms of macular degeneration. In exudative cases, the success rate of laser treatment guided by indocyanine green angiographic findings can vary considerably, and it is now known which angiographic presentations are not as likely to benefit. In dry macular degeneration, indocyanine green angiography appears to add clinically useful information, such as helping to identify plaques in fellow eyes with choroidal neovasculrization or watershed zones that may be predictive of future exudative transformation. In certain circumstances, indocyanine green angiography can be valuable in detecting choroidal neovascularization in other macular diseases or in helping to diagnose other choroidal conditions, especially when the clinical presentation is atypical, such as central serous chorioretinopathy in elderly patients. PMID- 10537779 TI - Transpupillary thermotherapy for choroidal melanoma. AB - The management of choroidal melanomas depends on many factors, most importantly, tumor size and location. Small choroidal melanoma in the posterior fundus is amenable to treatment options such as enucleation, radiotherapy, laser photocoagulation, and transpupillary thermotherapy or a combination of these methods. Transpupillary thermotherapy is a technique of tumor heating by infrared radiation delivered through the pupil into the tumor. This method causes dramatic tumor necrosis in choroidal melanomas up to 4 mm in thickness. With properly selected small choroidal melanomas, tumor control is approximately 94%. The heat induces cellular damage at the site of treatment with few remote side effects; therefore, complications are generally limited to the site of treatment and include retinal vascular obstruction (23%), retinal traction (20%), retinal neovascularization (6%), and retinal hole with detachment (< 1%). Tumors located temporal to the foveola demonstrate a statistically higher risk for retinal traction than those located in other quadrants. Tumors near the optic disk demonstrate a higher incidence of retinal neovascularization due to heat-induced obstruction of a major retinal vascular arcade. Overall, vision preservation is satisfactory after thermotherapy for choroidal melanoma, with more than 50% of patients maintaining the same or better vision after treatment, depending primarily on tumor location. In summary, small choroidal melanomas can be controlled with transpupillary thermotherapy, especially those near the optic disk and foveola in areas that are otherwise difficult to irradiate. Longer follow-up is necessary to assess for local recurrence and the impact of treatment on life prognosis. PMID- 10537780 TI - Laser treatment in eyes with drusen. AB - During the past several decades, many individuals have noted that drusen may resolve after macular laser photocoagulation. The lack of effective therapies against choroidal neovascularization (CNV) due to age-related macular degeneration (AMD) has prompted many invesigators to explore strategies designed to prevent CNV in eyes with high-risk drusen. Currently, several pilot studies in the United States, Spain, Sweden, Australia, and the United Kingdom are in progress; the Choroidal Neovascularization Prevention Trial (CNVPT) is the largest pilot study to date and is detailed herein. A definitive study, the Complications of Age-Related Macular Degeneration Prevention Trial, aims to evaluate the effects of laser treatment in patients with bilateral drusen and will commence this year. PMID- 10537782 TI - Photodynamic therapy: a new approach to the treatment of choroidal neovascularization secondary to age-related macular degeneration. AB - Visual loss as a result of choroidal neovascularization secondary to age-related macular degeneration continues to be a major challenge for all ophthalmologists. Photodynamic therapy represents an exciting and novel technique that uses light activated drugs and nonthermal light to achieve the selective destruction of choroidal neovascularization with minimal effects on the surrounding normal tissues. In Phase I-II clinical trials of photodynamic therapy with both benzoporphyrin derivative and tin ethyl etiopurpurin, closure of choroidal neovascularization was seen 24 hours after the treatment. However, recurrence of choroidal neovascularization can occur 2 to 3 months after treatment. Double blind, multicenter, randomized Phase III clinical trials with benzoporphyrin derivative and tin ethyl etiopurpurin are currently underway. PMID- 10537781 TI - Ocular manifestations of cat-scratch disease. AB - Bartonella henselae has only recently been isolated, characterized, and found to be the principal cause of cat-scratch disease (CSD). The availability of specific serologic investigations has allowed the recognition of a spectrum of ocular CSD syndromes that previously were ill defined and considered idiopathic. The primary inoculation complex causing regional lymphadenopathy is represented in the eye by Parinaud's oculoglandular syndrome; B. henselae is the most common cause. Leber's neuroretinitis has been identified for 80 years, and new data suggest that it is commonly a manifestation of CSD; the extent of the association remains to be determined. CSD optic neuritis is also described. The vitreoretinal manifestations include anterior uveitis, vitritis, pars planitis, focal retinal vasculitis, a characteristic retinal white spot syndrome, Bartonella retinitis, branch retinal arteriolar or venular occlusions, focal choroiditis, serous retinal detachments, and peripapillary angiomatous lesions. The pattern of ocular disease in AIDS-associated B. henselae infections is poorly delineated; unusual manifestations include conjunctival and retinal bacillary angiomatosis. The benefit of antimicrobial therapy for CSD in immunocompetent individuals has been difficult to establish, partly because most infections are self limited. Empirically, azithromycin, ciprofloxacin, rifampin, parenteral gentamicin, or trimethoprim-sulfamethoxazole provide the best therapeutic choices to minimize damage to the eye. PMID- 10537783 TI - Evidence-based medicine, utilities, and quality of life. AB - Evidence-based medicine provides the highest quality of information for medical practitioners. At the top of the pyramid of evidence-based medicine are the prospective, randomized clinical trials and meta-analysis. Evidence-based medicine can be incorporated with quality-of-life parameters; the latter can be quantified using utility theory. With utility theory, utility values range from 0.0 (death state) to 1.0 (perfect health state). The higher the utility value, the better a person's quality of life. Interventional treatment can change the utility level experienced by a patient. A change in utility value induced by an interventional treatment can be amalgamated with the duration of the treatment effect to provide the number of quality-adjusted life-years (QALYs) gained by a specific treatment (QALYs = [gain in utility value] x [duration of treatment effect]). Thus, this formula takes into account both the improvement in quality of life and the improvement in length of life gained by a treatment. The number of QALYs gained by a treatment can then be incorporated with medical costs (discounted for the time value of money) to arrive at a final common denominator of $/QALY (cost per QALY). The parameter $/QALY can be used to compare the cost effectiveness of interventional treatments across diverse specialties in medicine. In essence, this methodology allows a measure of the cost-effectiveness of a treatment that incorporates the highest quality of scientific information, clinical efficacy, patient quality-of-life preferences, and realistic costs. PMID- 10537784 TI - Cross-sectional era gives way to 3-D and 4-D imaging. PMID- 10537785 TI - Verdict on MQSA: quality up, but so is financial burden. PMID- 10537786 TI - Payment revisions could curtail use of contrast media. PMID- 10537787 TI - Multi-slice CT spirals past single-slice CT in diagnostic efficacy. PMID- 10537788 TI - Mobile imaging defies critics by continuing to roll along. PMID- 10537789 TI - MRI elucidates dementias and disorders of cerebral perfusion. PMID- 10537791 TI - Making wellness medicine work. PMID- 10537790 TI - A team approach to quality improvement. PMID- 10537792 TI - Medicare, 2000 and you. PMID- 10537793 TI - Improving patient communication in no time. PMID- 10537794 TI - Holding the gains in quality improvement. PMID- 10537795 TI - Becoming an effective physician leader. PMID- 10537796 TI - Physician-owned groups: the best strategy for success. PMID- 10537797 TI - A case study in developing a successful medical group. PMID- 10537798 TI - Developing 'groupthink' in a multispecialty group. PMID- 10537799 TI - Finding diamonds in the trenches with the nominal group process. PMID- 10537800 TI - GIS and public health policy: a new frontier for improving community health. PMID- 10537801 TI - Modern geographic information systems--promise and pitfalls. PMID- 10537802 TI - Geographic information systems (GIS) and community health: some statistical issues. PMID- 10537803 TI - Lead hot zones and childhood lead poisoning cases, Santa Clara County, California, 1995. PMID- 10537804 TI - Geographic distribution of mean blood lead levels by year in children residing in communities near the Bunker Hill lead smelter site, 1974-1983. PMID- 10537805 TI - Identifying predicted immunization "pockets of need," Hillsborough County, Florida, 1996-1997. PMID- 10537806 TI - Incidence rates of hepatitis A by ZIP code area, Salt Lake County, Utah, 1992 1996. PMID- 10537807 TI - Prevalence of benzoylecgonine (a cocaine metabolite) in newborn infants by ZIP code, Georgia, February 22 through April 23, 1994. PMID- 10537808 TI - Atlas of state and local geographic information systems (GIS) maps to improve community health. PMID- 10537809 TI - Predicting teen live birth rates using selected census-derived indicators, Lancaster County, South Carolina, 1990. PMID- 10537810 TI - Adequate prenatal care rates in North Dakota, 1991-1995. PMID- 10537811 TI - Automobile accidents to teenagers requiring emergency medical transport, Ventura County, California, 1996. PMID- 10537812 TI - Age-adjusted homicide rates by ZIP codes, Kansas City, Missouri, 1991-1995. PMID- 10537813 TI - Hospitalizations for all injuries, average annual rates per 1,000 adults ages 25 44 by ZIP code, Boston, Massachusetts, 1994-1996. PMID- 10537814 TI - Animal rabies cases in Central Palm Beach County, Florida, 1994-1998. PMID- 10537815 TI - Positive raccoon-strain rabies cases in Mahoning County, Ohio, 1997. PMID- 10537816 TI - Locations around the Hanford Nuclear Facility where average milk consumption by children in 1945 would have resulted in an estimated median iodine-131 dose to the thyroid of 10 rad or higher, Washington. PMID- 10537817 TI - Public notification to families with newborns at risk of methemoglobinemia from drinking water exposure, Clymer, New York, 1996-1998. PMID- 10537818 TI - Elevated nitrate levels in relation to bedrock depth, Linn County, Iowa, 1991 1996. PMID- 10537819 TI - Monitoring volatile organic compounds in private wells near a community landfill by tax parcel, Harford County, Maryland, 1986-1998. PMID- 10537820 TI - A computer simulation of groundwater withdrawal patterns for public water supply wells, Hutchinson, Kansas, 1998-2003. PMID- 10537821 TI - Location of East Carolina University School of Medicine telemedicine sites in relation to primary care health professional shortage areas for North Carolina's health service area VI, 1997. PMID- 10537822 TI - Client demographics (age and gender) at low-income clinics in Austin/Travis County, Texas, 1995-1996. PMID- 10537823 TI - Improving delivery of health and community services to welfare recipients, Columbia, South Carolina, 1997. PMID- 10537824 TI - Active patients attending the North Hill Child Health Clinic, Akron, Ohio, 1997. PMID- 10537825 TI - Percent of adults without health insurance in Ingham County, Michigan, 1994. PMID- 10537826 TI - Using marketing information to focus smoking cessation programs in specific census block groups along the Buford Highway corridor, DeKalb County, Georgia, 1996. PMID- 10537827 TI - Ranking priorities for a state family service integration initiative by census tract in northern New Castle County, Delaware, 1997. PMID- 10537828 TI - VISTA/PH software for community health assessment. PMID- 10537829 TI - Clackamas County Department of Human Services community health mapping engine (CHiME) geographic information systems project. PMID- 10537830 TI - An automated geographic information system for local health departments. PMID- 10537831 TI - Geographic information systems (GIS) for state and local public health practitioners, Part 1. PMID- 10537832 TI - Your first mapping project on your own: from A to Z. PMID- 10537833 TI - Geographic information system (GIS) hardware and software. PMID- 10537835 TI - State initiatives in geocoding vital statistics data. PMID- 10537834 TI - Development of childhood blood lead screening guidelines, Duval County, Florida, 1998. PMID- 10537836 TI - Methods to evaluate geographic access to health services. PMID- 10537837 TI - OSHA's latest latex legerdemain. PMID- 10537838 TI - Y2K: ready, set, go? AB - With the immovable deadline of the new millennium close at hand, many medical facilities will find they are scrambling to be prepared. Neither sophisticated integrated health systems or small medical offices will go unaffected or untouched. It is important for risk managers to make it their priority to ensure that all aspects are being addressed. PMID- 10537839 TI - Y2K, embedded chips, casualty hazards and due diligence in healthcare risk management. AB - Y2K raises challenges for healthcare risk managers that go beyond information technology issues. This article explains that (1) too little public attention is being paid to equipment which may well have Y2K faults and (2) few standards have been articulated for dealing with problems. Healthcare risk managers therefore must return to basic due diligence principles and develop their own standards and protocols. The article explains how to do due diligence and outlines suggested steps for dealing with the non-information technology side of compliance due diligence. PMID- 10537840 TI - Practical risk management principles for physicians. AB - Most medical schools and postgraduate residency programs do not focus adequate attention on risk management and quality management issues. This article will prepare physicians with an adequate working knowledge of risk management and quality management information, which will enable them to practice more effectively in today's litigious and regulatory climate. PMID- 10537841 TI - How to conduct a thorough sentinel event investigation. PMID- 10537842 TI - The dilemma over reprocessing single-use medical devices. AB - Making informed decisions about the reuse of single-use medical devices requires considerable analysis, study and management buy-in. This article addresses such issues as the prevalence of the practice, the changing standards and guidelines, institutional responsibilities, risk and insurance concerns and the ethical issues posed by the concept of reuse. PMID- 10537843 TI - When has the cause analysis reached the root? AB - Until a few years ago, the triennial JCAHO survey only peripherally involved the risk manager. With the advent of the sentinel event policy, this will certainly change regardless of whether the organization chooses to self-report. It is anticipated that the risk manager will be involved with the triennial survey particularly with regards to the PI standards. The mechanisms used by the organization to manage sentinel events will be scored within these standards and will be examined as part of the survey whether or not an event has occurred. The goal of ensuring that errors are managed in a manner that corrects the most fundamental contributing causes lies within the role of the risk manager. PMID- 10537844 TI - Physicians' cognitive errors and their liability consequences. AB - Diagnostic errors account for one-fifth of lawsuits against hospitals and physicians. Of greatest concern is the rising severity of lawsuits alleging diagnostic errors--over a 12-year period the average indemnity for these claims has risen by 258%. The hospital departments most affected are the emergency room, obstetrics, radiology and pathology. This article probes the structure of diagnostic errors and makes recommendations how to reduce the frequency and severity of these claims. PMID- 10537845 TI - Physician may be liable for failing to report child abuse. Stecker v. First Commercial Trust Co. PMID- 10537846 TI - Hospital may be liable for failing to act on disclosures to EAP counselors. Doe v. Garcia. PMID- 10537847 TI - Hospital's failure to prevent employee's abduction and murder not an intentional tort. Wake County Hospital System, Inc. v. Safety National Casualty Corp. PMID- 10537848 TI - On-call arrangement did not create joint venture. Rossi v. Oxley. PMID- 10537849 TI - Certain permanent injuries may cause an employee not to be "otherwise qualified". Webster v. Methodist Occupational Health Centers. PMID- 10537850 TI - Employee handbooks may be implied contracts with employees. Hansucker v. Josephine Sunset Home; Jewell v. North Big Horn Hospital District. PMID- 10537851 TI - The role of the workplace in the production and containment of health costs: the case of stress-related disorders. AB - Workplaces vary enormously in the amount of harmful stress they produce, even within specific economic sectors. Stress of certain kinds and at certain levels tend to produce health harms and costs that are borne not only by individual employees and employers but also by families and society at large. Variations in stress levels within economic sectors can be traced to variations in management practices that govern key conditions of work involving demand, effort, control and reward. The costs of stress-related disorders produced by adverse governance practices are transferred outside the workplace in varying degrees. The actual extent of this cost transfer depends on policies and programs within the workplace. We can characterize workplaces according to a typology in which the key dimensions are commitment to abate harm through participatory management practices and the effectiveness and efficiency of harm containment through programs such as employee assistance and health promotion. The most health promoting and cost-avoiding workplaces foster high control, high reward conditions and support employees with employee assistance and health promotion programs. The policy implications of this observation are drawn out. PMID- 10537852 TI - Creating health and health promoting hospitals: a worthy challenge for the twenty first century. AB - States that it seems self-evident that a hospital should be a healing environment, a healthy place to work, should not harm the health of the environment and should contribute to and be a source of health in the community, but argues that hospitals have not paid a great deal of attention to many of these issues until recently. Suggests that in recent years, a new and broader understanding of health promotion has led to a re-examination of the ways in which hospitals can be both healthy and health-promoting. Begins by exploring the broader concepts of health promotion that lay the foundation for the creation of healthy and health-promoting hospitals and provides some examples of how these approaches are being applied. PMID- 10537853 TI - Improving the quality of institutional care of urinary incontinence among the elderly: a challenge for governmental regulation. AB - This article describes actual UI prevalence and quality of care at Israeli LTC institutions for the elderly. The analysis is based on current regulatory data on 14,406 residents at 196 residential homes, and 8,278 patients at 159 hospitals for the chronically ill. It includes a calculation of summary indices of quality, the percentage of institutions with deficient items and of those showing change, and a description of functional status profiles. Multiple regression explains the deficiency rate variance through independent institutional variables. There is a higher prevalence of severe functional impairment and full incontinence at hospitals for the chronically ill than at residential homes. There were higher rates of deficiencies and lower rates of corrections for structural items than for process items at both. A major improvement occurred for process items (50-100 per cent). Regarding outcomes, 34 percent of the residents with UI during the first assessment were continent two years later. PMID- 10537854 TI - Self-assessment and business excellence. PMID- 10537855 TI - Using the business excellence model to develop a strategy for a healthcare organisation. AB - This article examines the appropriateness of the Business Excellence Model in developing a strategy for Bolton Hospitals NHS Trust to measure organisational performance. The need for a strategy to measure organisational performance and to improve organisational performance was highlighted with the production of the Government White Paper, The New NHS: Modern and Dependable. At the heart of recommendations there is emphasis on improving quality and driving efficiency. Greater emphasis will be placed on organisations measuring their performance. By utilising the conceptual framework, which consisted of The European Foundation for Quality Management (EFQM) Model, it became evident that, although tools were in existence within Bolton Hospitals to measure organisational performance, several critical areas needed addressing. By addressing these key areas, the organisation could begin to work towards its goal of business excellence. The conclusions drawn from this project demonstrated that there was scope for Bolton Hospitals to improve on organisational performance. It was highlighted that the Trust was functioning well in some areas of the EFQM Model, but not in others. For Bolton Hospitals NHS Trust to improve organisational performance, the EFQM Model should be adopted. PMID- 10537856 TI - Improving performance through self-assessment. AB - Wakefield and Pontefract Community Health NHS Trust uses the European Business Excellence Model self-assessment for continuous improvement. An outline of the key aspects of the model, an approach to TQM, is presented. This article sets out the context that led to the adoption of the model in the Trust and describes the approach that has been taken to completing self-assessments. Use of the model to secure continuous improvement is reviewed against Bhopal and Thomson's Audit Cycle and consideration is given to lessons learned. The article concludes with a discussion on applicability of the model to health care organisations. It is concluded that, after an initial learning curve, the model has facilitated integration of a range of quality initiatives, and progress with continuous improvement. Critical to this was the linking of self-assessment to business planning and performance management systems. PMID- 10537857 TI - Self-assessment of all the health centres of a public health service through the European Model of total quality management. AB - The Basque Country Public Health Service has moved in the last years from considering quality as an attribute of patient care to thinking that all management can be subject to improvement. Consequently, its general management team has promoted and supported a self-assessment experience of all their centres by means of the European Quality Model. This strategy has been facilitated by the Basque Country Government, which has strongly encouraged total quality management in companies, and has created the Basque Foundation for Quality Promotion, a key institution in this whole process. A total of 26 centres of the Public Health Service concluded a self-assessment process. As the main result of this, different improvement areas were detected, and various necessary actions were implemented in the centres assessed. Advantages, troubles and future work lines to extend and improve the use of the EFQM model in the health sector are discussed. PMID- 10537858 TI - Achieving a culture of continuous improvement by adopting the principles of self assessment and business excellence. AB - Following a brief description of the inception of self-assessment and the European Foundation for Quality business excellence model, this article describes how one clinical directorate in an NHS Trust used the principles of both to secure a culture of continuous improvement. The journey from a mainly hierarchical, bureaucratic, individualist culture to one where the norms, values and beliefs reflected teamwork, involvement and empowerment is described. The highs, lows and learning points are all included, in an attempt to enlighten other healthcare organisations considering the benefits and pitfalls of using the business excellence model to improve the quality of their healthcare delivery. PMID- 10537859 TI - EFQM approach and the Dutch Quality Award. AB - Different approaches to improve quality are used in organizations delivering health care. Donabedian introduced structure, process and outcome, from which other approaches like self-assessment, accreditation, visitation, International Standards Organisation (ISO) and European Foundation for Quality Management (EFQM) can be aligned. The EFQM model is one such approach that has been adopted and adapted by the Dutch Institute for Quality Management. This article describes the background and progress relating to the use of the EFQM business excellence model within Dutch health care organizations. In addition the process for applying for the European Quality Award and the Dutch Quality Award are described in detail. Finally, the reader is enlightened regarding the work of the European ExPeRT research group who are promoting the use of quality models within health care. PMID- 10537860 TI - The forgotten stakeholders: seniors' values concerning their health care. AB - Within the context of health care reform and its evaluation, a major gap exists in relation to our understanding of the values which seniors hold regarding their health care. This paper reports on a modified participatory, ethnographic study of such values, using transcribed interviews with ten seniors from across Canada. Members of the National Advisory Council of Canada, most of whom are themselves seniors, participated in designing the study, carrying out the interviews and interpreting the results. Clusters of values were identified concerning health care services, service providers and the overall health care system. While the numbers involved in this study preclude generalizing to the population, a number of recommendations emerged from the study which could impact on future research and begin to influence health policy at local and national levels. PMID- 10537861 TI - Quality of diagnostic services for cancer: a comparison of open access and conventional outpatient clinics. AB - OBJECTIVE AND STUDY DESIGN: To assess quality of a quick and early diagnosis route (QED) by determining effectiveness and cost-effectiveness of five clinics compared with three conventional outpatient clinics. Prospective economic evaluation. Six-month cohort of all referrals (November 1996-April 1997). SUBJECTS: All referrals for suspected cancers of: upper gastro-intestinal tract; urinary tract, prostate and testis; skin. EFFECTIVENESS: Median days saved between GP referral and date of: diagnostic appointment; consultant decision; intervention. RESULTS: GP referral to diagnostic appointment: QED was effective (median days) for all clinics. Diagnostic appointment to consultant decision: QED was effective for testicular and haematuria clinics. Consultant decision to intervention: QED was effective for haematuria, testicular and melanoma clinics. COST-EFFECTIVENESS: Extra (incremental) NHS cost per patient diagnosed. RESULTS: Less than 5 Pounds per day saved between GP referral and diagnostic appointment for: endoscopy; haematuria; prostate; testicular; melanoma. Less than 3 Pounds per day saved between GP referral and consultant decision for: testicular; haematuria. Less than 3 Pounds per day saved between GP referral and intervention for: endoscopy; haematuria; testicular; melanoma. CONCLUSION: A "quick and early" diagnostic route provides a higher quality service through improved effectiveness and cost-effectiveness compared to conventional outpatients. PMID- 10537862 TI - A comparison of service predispositions between NHS nurses and hospitality workers. AB - The following study sought to develop an instrument to elicit the service predispositions of nurses and hospitality foodservice workers. Results of a pilot study suggested that the service predisposition instrument (SPI) was valid and therefore appropriate to investigate the service attitudes of these workers. Service predispositions of nurses from two NHS Trusts were compared with those of hospitality foodservice workers in two large hotels. Overall, both nurses and foodservice workers were found to have similar positive service predispositions. However, significant differences were present between groups for certain service dimensions. PMID- 10537863 TI - Clinical governance: culture, leadership and power--the key to changing attitudes and behaviours in trusts. AB - Clinical governance places for legal duty for quality on Trust chief executives. The article deals with how chief executives can overcome the cultural and behavioural obstacles to change which impede the journey to developing effective structures for clinical governance. To establish effective structures for clinical governance, chief executives will need to address these features of organisational life--culture, power and leadership. The article seeks to provide practical ways in which they can do this. PMID- 10537864 TI - Relationship between primary payer and use of proactive immunization practices: a national survey. AB - OBJECTIVE: To quantify the national use and determinants of proactive immunization practices by examining the relationship to the primary practice payer. STUDY DESIGN: A standardized survey was conducted in 1995 by trained personnel using computer-assisted telephone interviewing. PATIENTS AND METHODS: A stratified random sample of family physicians, pediatricians, and general practitioners across the United States was selected from the American Medical Association master file of physicians list, which included nonmembers. The main outcome measures were use of reminder systems and assessment of immunization rates. RESULTS: Of the 1769 physicians who were contacted, 1236 participated. Use of reminder systems varied by the practice's primary payer: 31% of health maintenance organization (HMO), 41% of Medicaid, 27% of fee-for-service (FFS), and 28% of no predominant payment source physicians reported using a reminder system (P < 0.01). Use of computerized reminders also varied according to practice primary payer (HMO, 68%; Medicaid, 34%; FFS, 51%; and no predominant payment source, 42%; P < 0.01) as did assessment of immunization rates in the practice (HMO, 57%; Medicaid, 40%; FFS, 28%; and no predominant payment source, 30%; P < 0.01). A majority of Medicaid physicians (84%) required a physical examination before immunization, compared to 49% of HMO, 56% of FFS, and 63% of no predominant source physicians (P < 0.01). CONCLUSIONS: The primary payment source of a practice appears to influence use of proactive immunization practices. PMID- 10537865 TI - Polypharmacy management in Medicare managed care: changes in prescribing by primary care physicians resulting from a program promoting medication reviews. AB - OBJECTIVE: To examine the effects of medication reviews by primary care physicians on prescriptions written for elderly members of a Medicare managed care organization who were at risk for polypharmacy. STUDY DESIGN: Prospective study with follow-up survey. PATIENTS AND METHODS: We conducted a study in 1995 to demonstrate the prevalence of polypharmacy (defined as receiving 5 or more prescription medications during the 3-month study period) among elderly members of our managed care organization. Two years later, elderly members identified as being at risk for polypharmacy were sent a letter encouraging them to schedule a medication review with their primary care physician. Each primary care physician was provided with clinical practice guidelines on polypharmacy and patient specific medication management reports. Patients and physicians were subsequently mailed a survey to assess the impact of the medication review program on prescribing practices. RESULTS: Of 37,372 elderly members screened, 5737 (15%) were at risk for polypharmacy. Of these, 2615 (46%) responded to the follow-up survey. Of the survey respondents, 1087 (42%) had gone to their primary care physician for a medication review. During the review, 96% of patients discussed their prescription medications and 72% discussed nonprescription medications they were taking. Twenty percent reported that their physician discontinued medications, 29% reported that the physician changed the dose of a medication, and 17% informed their physician about a new prescription or nonprescription medication they were taking. Of the 275 primary care physicians surveyed, 56 (20%) returned the questionnaire. Of these, 61% reported that the medication review program was "very" or "somewhat useful." Thirty-five percent reported discontinuing unnecessary medications, and 23% reported decreasing the frequency of dosing. Overall, 45% of physicians reported making at least one change in their prescribing to a member at risk for polypharmacy. CONCLUSIONS: Our program promoting medication reviews between primary care physicians and their elderly patients resulted in significant changes in prescribing by physicians. This type of program is likely to decrease the risk of polypharmacy among older members of a Medicare managed care organization. PMID- 10537866 TI - Course and cost of treatment for depression with fluoxetine, paroxetine, and sertraline. AB - OBJECTIVE: To compare depression-related treatment costs and total healthcare costs for patients diagnosed with depression and treated with either sertraline, paroxetine, or fluoxetine. PATIENTS AND METHODS: Claims records from a national database of patients diagnosed with depression who began treatment with an SSRI in 1995, following an antidepressant medication-free period of at least 6 months, were included. Treatment course and associated depression-related treatment and total healthcare costs during the subsequent 12-month treatment period were examined using univariate and multivariate methods. RESULTS: Nine-hundred five (905) patients taking sertraline, 492 on paroxetine, and 945 on fluoxetine met inclusion criteria. The groups were similar and representative with respect to gender and age. Mean dose over the 12-month treatment period increased 24%, indicating significant titration in all cohorts. Patients treated with paroxetine had shorter treatment duration (157.0 days) than did patients treated with fluoxetine (192.6 days) or sertraline (166.9 days, P < 0.001). Patients receiving index treatment with paroxetine were most likely to switch to another SSRI (21.3%); those taking sertraline were second most likely to switch (16.1%); and those on fluoxetine were least likely (12.4%, P = 0.001). Mean costs for depression-related outpatient visits and hospitalizations were similar. Mean antidepressant prescription costs differed, being $586, $419, and $446 for fluoxetine, paroxetine and sertraline cohorts, respectively (P < 0.001). In this sample, the fluoxetine cohort did not have lower nonpharmaceutical healthcare costs to offset higher pharmaceutical acquisition costs. Conclusions from median and multivariate analyses were robust to these findings. CONCLUSIONS: During this study period when fluoxetine, paroxetine, and sertraline were all well established agents, similar depression-related treatment courses and cost characteristics among all 3 drugs were observed. PMID- 10537867 TI - Project LEAP of New Jersey: lower extremity amputation prevention in persons with type 2 diabetes. AB - OBJECTIVE: To reduce type 2 diabetes-related lower extremity amputations (LEAs) in New Jersey through a statewide training program for primary care providers at healthcare agencies in high-risk areas. STUDY DESIGN: Project LEAP provided 27 1 day training workshops to 560 healthcare professionals representing 85 organizations. The effect of training was evaluated based on a multiple-choice knowledge test, self-reported practice behaviors, and a medical records audit of practice behaviors, and pre- and postprogram LEA rates. PATIENTS AND METHODS: We evaluated statistically significant differences in pre- and postprogram knowledge scores using Student's t-tests. We also evaluated providers' intentions to change clinical foot-care practices and compared them with actual practices documented in medical records. We used analysis of variance to determine any statistically significant differences in pre- and postprogram LEA rates at various types of institutions. In addition, we assisted facilities in the development of self education programs containing specific foot-care modules. RESULTS: Participating providers were: 70.6% nurses, 7.8% physicians, 4.5% podiatrists, 4.2% dietitians, and 12.9% all others. Pre- and postprogram knowledge scores increased by 12% (T = 13.29; P < 0.0001) and were maintained for 9 months (T = 7.58; P < 0.05). Provider intentions to change clinical practice behaviors correlated with self reported practice changes 9 months postprogram (r = .51; P < 0.001). Medical record audits 1 year before and 9 months after training demonstrated marked improvement in foot-care practices in the following areas: (1) foot-care education given to patients by primary care providers; 2) documentation of peripheral vascular disease; 3) documentation of patient preventive care practices; and 4) referrals to diabetes educators, orthopedists, podiatrists, and diabetologists. Education programs with specific foot-care components increased 10%. The overall incidence of pre- and posttraining LEAs did not change significantly but differed depending on institution type. Hospitals and community healthcare centers were more likely to show postprogram reductions in LEAs than nursing homes and rehabilitation centers. CONCLUSION: Institutionalization of a LEAP program resulted in improved provider knowledge and certain clinical practice behaviors. There was a trend toward an overall reduction in the number of LEAs at participating institutions. PMID- 10537868 TI - Quality of care for chronic illness in primary care: opportunity for improvement in process and outcome measures. AB - OBJECTIVE: To describe adherence to a number of quality indicators and clinical outcomes for asthma, diabetes mellitus, hypertension, coronary heart disease, atrial fibrillation, and cerebrovascular disease in the primary care practices of the Practice Partner Research Network (PPRNet). STUDY DESIGN: Cross-sectional epidemiologic design. PATIENTS AND METHODS: PPRNet is a national research network of ambulatory, mostly primary care practices that use the Practice Partner Patient Records electronic medical records. Participating practices send anonymous clinical data on patients to the PPRNet data center monthly. Standard database management and statistical software are used to compile practice reports. These reports include measures of adherence to process and outcome measures for chronic illnesses, the subject of this report. RESULTS: Forty-eight PPRNet practices provided data for the first quarter of 1998. A total of 336,401 patients were active in these practices during this quarter. At least 2000 active patients had each of the conditions studied. Wide variation in guideline adherence among PPRNet practices was present for each of the performance measures. Better performance was present for physical examination measures and laboratory monitoring than for treatment interventions. Overall performance was excellent for blood pressure monitoring, poor for lipid monitoring in patients with CHD, and intermediate for glycosylated hemoglobin monitoring in patients with diabetes mellitus. CONCLUSION: The findings of this study are comparable to others in documenting that most clinical practice guidelines for chronic illness are not followed for a majority of patients and that large majorities do not reach desired clinical outcomes. PMID- 10537869 TI - Assessment of patient satisfaction with a formulary switch from omeprazole to lansoprazole in gastroesophageal reflux disease maintenance therapy. AB - OBJECTIVE: To determine if patients perceived a difference in the efficacy, side effects, and value of omeprazole versus lansoprazole for gastroesophageal reflux disease (GERD) maintenance therapy after a formulary conversion, and to evaluate the costs of the conversion. STUDY DESIGN: An unblinded questionnaire was mailed to patients who were currently receiving GERD maintenance therapy with lansoprazole from the Veterans Affairs San Diego Healthcare System. PATIENTS AND METHODS: Three hundred patients who had been on omeprazole for a minimum of 2 months prior to the formulary conversion and on lansoprazole for a minimum of 2 months after the formulary conversion were surveyed. Patients were asked to rate the severity and frequency of their symptoms (pain, heartburn, and regurgitation) on a scale from 0 to 9 for each medication. Questions regarding side effects, medication preference, and satisfaction with the formulary conversion process were also addressed. RESULTS: Fifty-two percent of the surveys were returned. There was no statistically significant difference between median total symptom scores for omeprazole and lansoprazole (1.33 vs. 1.34, respectively). More patients reported side effects to lansoprazole (P < 0.001) than to omeprazole. Sixty-four percent of patients preferred omeprazole (P < 0.005). The formulary conversion was estimated to save $29,000 per year. CONCLUSIONS: Omeprazole was the medication preferred by patients for GERD maintenance therapy. Patients were willing to pay an additional fee for their preferred agent. Fewer adverse events were reported with omeprazole. The potential cost savings of the formulary conversion may have been at the expense of patient satisfaction. PMID- 10537870 TI - Shift away from inpatient care spurs dramatic turnaround. PMID- 10537871 TI - Risk stabilization accounts keep hospitals afloat under Medicare risk. PMID- 10537872 TI - West Coast hospitals send mixed signals on their capitation experience. PMID- 10537873 TI - Profile network performance to manage contact capitation. AB - Often, the missing link in managing contact cap is a profiling system that gives specialists the data they need to make educated decisions. This system focuses on reducing intra-network variation. PMID- 10537874 TI - Clinical data hold key to profits under capitation. AB - One of the most pressing questions in health care information management is what data should be contained in the patient medical record to manage a budget under capitation. Answering this question is fundamental to the success of risk-bearing provider groups. PMID- 10537875 TI - Three strategies to improve market share and gain competitive advantage. PMID- 10537876 TI - Quantifying the sales process in healthcare. PMID- 10537877 TI - MemorialCare bets future is in the cards. PMID- 10537878 TI - Understand the overlap between FMLA and COBRA. AB - An employer's obligation to continue an eligible employee's medical insurance under the Family and Medical Leave Act overlaps with the employee's right to continued medical insurance coverage under the provisions of the Consolidated Omnibus Budget Reconciliation Act of 1985 (COBRA). To avoid liability traps resulting from the overlap, employers, plan sponsors and plan administrators must have a good understanding of the two laws and keep accurate records. PMID- 10537879 TI - Community health councils. Sold a pup. AB - CHCs have changed little since 1974. There are huge variations in their effectiveness. National policy is needed to set out their remit and lines of accountability. PMID- 10537880 TI - Practice management. Behind the lines. AB - The introduction of nurse telephone triage for patients requesting same-day attention in a GP practice has reduced the number of appointments and home visits for these patients. Patients welcome the opportunity to speak to a health professional when they telephone. The practice has found the scheme a useful method of demand management and hopes to expand it. PMID- 10537881 TI - Fair game. PMID- 10537882 TI - Devolution. Man of the people. PMID- 10537883 TI - Innovation. Trouble vision. PMID- 10537884 TI - Civil justice reforms. Woolf at the door. PMID- 10537885 TI - Workplace harassment. Defence mechanisms. PMID- 10537886 TI - Whistle blowing. In on the act? PMID- 10537887 TI - Human rights. Indefinite articles. PMID- 10537888 TI - Confederate flagging. PMID- 10537889 TI - Primary care groups. Lending a hand. AB - High-quality, dedicated management support was crucial to the GP commissioning pilots. A fifth of the pilots reported difficult relationships with their health authority. There are concerns about bringing prescribing 'outliers' into line. Involving the public will be a considerable challenge for PCGs. PMID- 10537890 TI - Our time has come. PMID- 10537891 TI - Clinical audit. Count the cost. AB - A system for calculating the cost of clinical audits in terms of money and staff time has been in operation at Brighton Health Care trust since 1995. The information has proved useful in cost-benefit analysis. At th e end of each audit an action plan is agreed with the lead clinician over required changes. Making the cost of audits explicit increases the likelihood of recommendations being implemented. PMID- 10537892 TI - General managers. Developing whirl. Interview by Lynn Eaton. PMID- 10537893 TI - The cycle of violence: important considerations for EMS providers. PMID- 10537894 TI - EMS & the domestic violence patient: a report card of existing policy, protocol & training. PMID- 10537895 TI - Seattle: bus off the Aurora Highway Bridge. PMID- 10537896 TI - Equity in the finance of health care: some further international comparisons. AB - This paper presents further international comparisons of progressivity of health care financing systems. The paper builds on the work of Wagstaff et al. [Wagstaff, A., van Doorslaer E., et al., 1992. Equity in the finance of health care: some international comparisons, Journal of Health Economics 11, pp. 361 387] but extends it in a number of directions: we modify the methodology used there and achieve a higher degree of cross-country comparability in variable definitions; we update and extend the cross-section of countries; and we present evidence on trends in financing mixes and progressivity. PMID- 10537897 TI - The redistributive effect of health care finance in twelve OECD countries. AB - The OECD countries finance their health care through a mixture of taxes, social insurance contributions, private insurance premiums and out-of-pocket payments. The various payment sources have very different implications for both vertical and horizontal equity and on redistributive effect which is a function of both. This paper presents results on the income redistribution consequences of the health care financing mixes adopted in twelve OECD countries by decomposing the overall income redistributive effect into a progressivity, horizontal inequity and reranking component. The general finding of this study is that the vertical effect is much more important than horizontal inequity and reranking in determining the overall redistributive effect but that their relative importance varies by source of payment. Public finance sources tend to have small positive redistributive effects and less differential treatment while private financing sources generally have (larger) negative redistributive effects which are to a substantial degree caused by differential treatment. PMID- 10537898 TI - Capitation contracts: access and quality. AB - The implications of competition amongst providers in both private and public systems for the quality of service and the number of care providers are analysed. Strong conditions must be imposed on preferences and cost conditions for quality to be efficiently supplied. In a median voter model of a public system the capitation fee and quality are lower than under competitive market equilibria and the number of practices inefficiently small. Entry control by a union which maximises gross provider income reduces the number of practices until the market is only just covered. PMID- 10537899 TI - The irrelevance of inference: a decision-making approach to the stochastic evaluation of health care technologies. AB - The literature which considers the statistical properties of cost-effectiveness analysis has focused on estimating the sampling distribution of either an incremental cost-effectiveness ratio or incremental net benefit for classical inference. However, it is argued here that rules of inference are arbitrary and entirely irrelevant to the decisions which clinical and economic evaluations claim to inform. Decisions should be based only on the mean net benefits irrespective of whether differences are statistically significant or fall outside a Bayesian range of equivalence. Failure to make decisions in this way by accepting the arbitrary rules of inference will impose costs which can be measured in terms of resources or health benefits forgone. The distribution of net benefit is only relevant to deciding whether more information is required. A framework for decision making and establishing the value of additional information is presented which is consistent with the decision rules in CEA. This framework can distinguish the simultaneous but conceptually separate steps of deciding which alternatives should be chosen, given existing information, from the question of whether more information should be acquired. It also ensures that the type of information acquired is driven by the objectives of the health care system, is consistent with the budget constraint on service provision and that research is designed efficiently. PMID- 10537900 TI - On the use of survival analysis techniques to estimate medical care costs. AB - Measurement of treatment costs is important in the evaluation of medical interventions. Accurate cost estimation is problematic, when cost records are incomplete. Methods from the survival analysis literature have been proposed for estimating costs using available data. In this article, we clarify assumptions necessary for validity of these techniques. We demonstrate how assumptions needed for valid survival analysis may be violated when these methods are applied to cost estimation. Our observations are confirmed through simulations and empirical data analysis. We conclude that survival analysis approaches are not generally appropriate for the analysis of medical costs and review several valid alternatives. PMID- 10537901 TI - On aggregating QALYs: a comment on Dolan. AB - Dolan [Dolan, P., 1998, The measurement of individual utility and social welfare. Journal of Health Economics, Vol. 17, pp. 39-52] in a recent paper suggested an empirical method for estimating the shape of the social welfare function. Using a simple theoretical model it is shown that Dolan's proposed empirical method has no theoretical foundation. The main problem with the approach is that it measures only the altruistic values that individuals attach to other peoples' health status and ignores the utility that individuals attach to their own health status. PMID- 10537902 TI - Drawing a veil over the measurement of social welfare--a reply to Johannesson. PMID- 10537903 TI - Medicaid fraud charged. Justice Department files lawsuit against Kansas City hospital. PMID- 10537905 TI - Radiologists top dogs. PMID- 10537904 TI - High court: FTC can regulate associations. PMID- 10537906 TI - Betting on rural markets. Head of start-up company aims to succeed by buying hospitals he sold a decade ago. PMID- 10537907 TI - Insurer, hospital partner. HealthPartners funds Wis. hospital project for 3 board seats. PMID- 10537908 TI - Kansas hospital dodges decertification. PMID- 10537909 TI - HHS: no quick fix for hospital woes. PMID- 10537910 TI - Practicing 'zero tolerance'. Regulatory agencies should require pre-employment and random drug tests at hospitals. PMID- 10537911 TI - At the center of cancer care. For-profit outpatient centers playing bigger role in treatment, clinical trials. AB - New treatments and technological advances mean the vasty majority of cancer patients are receiving their care on an outpatient basis. Increasingly, for profit cancer centers are providing that care. Although some hospitals have embraced the centers as partners, others see them as interlopers, drawing off lucrative business and threatening patients' continuity of care. PMID- 10537912 TI - Health lobbies go to bat for the uninsured. PMID- 10537913 TI - Integrating systems for chronic care. Study suggests partnering with others helps systems deliver better patient care. PMID- 10537914 TI - VHA to launch Internet package for docs. Alliance will compete with for-profits in offering on-line business and clinical support services. PMID- 10537915 TI - Investors aren't sold on healthcare bonds. PMID- 10537916 TI - Traditional Medicare. Providers campaign for 'a secure alternative'. PMID- 10537917 TI - Congress to consider budget-law relief bills. PMID- 10537919 TI - Alaska nurses vote for contract. Providence Alaska Medical Center gives nurses a voice in patient care, staffing ratios. PMID- 10537918 TI - Another HMO unplugged. Not-for-profit Sentara, Bon Secours scrapping Va. Medicare plan. PMID- 10537920 TI - Defense's turn nears in Columbia trial. PMID- 10537921 TI - Chasm grows between rich and poor. Top hospital performers get stronger as weaker competitors stumble, new research indicates. AB - Although recent studies show hospital operating margins dropped 45% in the fourth quarter of 1998, compared with the year-ago period, a new analysis of industrywide financial ratios reveals that the "average" hospital may be less representative of the whole. Instead, a chasm is widening around the median, with strong hospitals getting stronger and weak hospitals showing further decline. PMID- 10537922 TI - Tests may speed heart attack diagnosis. PMID- 10537923 TI - PPOs riding a wave of popularity. PMID- 10537924 TI - Spotlight on CFOs. As feds have grown savvier, bean counters increasingly have been targets in fraud probes. PMID- 10537925 TI - Life insurance: mortality trends and profitable business. PMID- 10537926 TI - Comparative mortality of persons with autism in California, 1980-1996. AB - The authors studied mortality rates of persons with autism, using the extensive California developmental disabilities registry. There was an overall mortality ratio (MR) of 213%. The MR for females (490%) was strikingly higher than for males (167%). The excess mortality rate (EDR) increased with age, while the mortality ratio (MR) decreased with age. Persons with autism are subject to increased mortality risk, as summarized in the provided tables. PMID- 10537927 TI - Underwriting implications of carcinoma in situ of the cervix. PMID- 10537928 TI - Homocysteinemia: new information about an old risk factor for vascular disease. AB - OBJECTIVE: To determine the importance of homocysteinemia as a risk factor for atherosclerotic vascular disease. DESIGN: Literature review of published studies homocysteine as risk factor for atherosclerotic vascular disease. METHODS: MEDLINE search from 1969 to 1998 using homocysteine and vascular disease as search terms, from which 13 articles were selected for review. RESULTS: Homocysteine is a sulfur containing amino acid derivative formed during methionine metabolism. Inherited deficiencies of cystathionine B synthase or MTHF reductase result in markedly elevated plasma homocysteine levels and homocystinuria. Although rare, hereditary homocystinuria results in a variety of life threatening vascular complications occurring at a young age. Lesser degrees of homocysteinemia may result from vitamin B12, folate and pyridoxine deficiencies as well as a recently described mutation of the MTHF reductase gene. Homocysteinemia from these causes has been shown to increase the risk of coronary artery disease, peripheral artery disease, stroke, and venous thrombosis. Postulated mechanisms for this association are discussed. CONCLUSION: Homocysteinemia is a risk factor for premature vascular disease. The strength of this association is similar to that due to hyperlipidemia and tobacco use. Although vitamin supplementation with folic acid, B12, and B6 is able to reduce homocysteine levels in many persons, proof of the effectiveness of vitamin treatment in preventing or halting the progression of vascular disease is not yet available. PMID- 10537929 TI - 5-year survival after bone marrow transplantation for aplastic anemia. AB - The International Bone Marrow Transplant Registry reported experience from 179 transplant teams worldwide on patients treated with bone marrow transplantation for severe aplastic anemia. The cohort consisted of 470 patients (283 male, 187 female), mean age 20 years, who received an HLA-identical sibling bone marrow transplant for aplastic anemia between 1988 and 1992. The etiology of aplastic anemia was usually idiopathic, with a small number of cases caused by hepatitis or toxin exposure. Congenital aplastic anemia was not considered. Expected mortality was based on the United States 1992 Total Population Mortality Table. Additional data were obtained from the author. PMID- 10537930 TI - Comparative mortality in cerebral palsy patients in California, 1980-1996. AB - BACKGROUND: The large database of the California Department of Developmental Services provides a data source for mortality rates in persons with mental retardation by age, sex, severity, cause and associated conditions. This study involves patients with a diagnosis of cerebral palsy. RESULTS: After a table of demographic data, four tables are used to show detailed age-related observed and expected mortality rates for Cerebral Palsy patients by sex and a severity factor that divides the patients into two groups of approximately equal size. The factor used was quadriplegia (all four limbs involved in motor dysfunction). Spasticity was the predominant feature of the motor dysfunction. CONCLUSION: Excess mortality was moderate in the less severe Cerebral Palsy patients, but was higher in those with quadriplegia (overall EDR--Excess Death Rate--about 6 per 1000 and 16 per 1000, respectively). In less severe cases EDR was higher at ages 1-4 years, the almost constant to age 49, then rose with advancing age. In case with quadriplegia EDR decreased in childhood and young adults to a relatively stable minimum at ages 25-49, then increased at older ages. There was little sex difference in EDR. PMID- 10537931 TI - Life table analysis. AB - Life table analysis is an effective way to present and evaluate survival data in a number of circumstances. The summary tables and survival curves that are commonly seen in the medical literature are frequently derived through life table analysis. A basic understanding of life table construction is of benefit to the medical director who wishes to abstract comparative mortality data from study reports. This article reviews the basic methodology of life table analysis with a particular focus on handling right-censored study participants as withdrawals. PMID- 10537932 TI - Medical underwriting on the eve of the millennium. AB - This article discusses the effects of the profusion of medical data on daily underwriting and then attempts to derive conclusions as to future developments in this field. Whereas the volume of medical information is constantly increasing, access to this information is in many cases being rendered difficult as a result of a highly critical attitude to the insurance industry on the part of applicants and medical practitioners. The second part of this article looks more closely at the handling of medical information and makes reference in this regard to the experience of insurance medical officers and directors of underwriting departments in the most important international markets. PMID- 10537933 TI - Genetic testing legislation: a review of the past two years. AB - The American Council of Life Insurance is a national trade association that represents the interests of legal reserve life insurance companies in legislative, regulatory and judicial matters at the federal, state and municipal levels of government and at the National Association of Insurance Commissioners. Its member companies hold more than 90 percent of the life insurance in force in the United States. PMID- 10537934 TI - Psoriatic arthritis. AB - Arthritis of various types can occur as a manifestation of psoriasis, and is classified as one of the seronegative spondyloarthropathies. Progression of psoriatic arthritis can be predicted by both clinical and biochemical markers. In some cases, mortality may be adversely effected. PMID- 10537935 TI - Alternative medicine use in the United States. PMID- 10537936 TI - Three-year survival and recurrence after stroke. PMID- 10537937 TI - Benefit of carotid endarterectomy in patients with symptomatic moderate or severe stenosis. PMID- 10537938 TI - Decrease in all-cause and CHD mortality with pravastatin over a broad range of initial cholesterol levels. PMID- 10537939 TI - Medical students and AIDS: knowledge, attitudes and implications for education. AB - Second year medical students at a large midwestern university were surveyed about their attitudes regarding AIDS. Results indicated: (1) students with homosexual and/or HIV-positive friends were significantly more tolerant toward AIDS patients, (2) over half the students believed that treating AIDS patients may be hazardous and that their education had not prepared them to treat these patients safely, (3) one-third believed they had the right to refuse to treat AIDS patients, and (4) AIDS-phobia was significantly associated with homophobia. These data suggest that medical educators may need to help students overcome AIDS phobia before some students will be able to incorporate instruction about AIDS since AIDS-phobia may inhibit this learning. Didactic instruction must be coupled with modeling by educators of non-prejudicial attitudes and strict adherence to medical professionalism. PMID- 10537940 TI - A review of the effectiveness of Smokebusters: community-based smoking prevention for young people. AB - Smokebusters is a community-based smoking prevention initiative for young children which aims to prevent them from starting to smoke. Despite the increase of Smokebusters clubs throughout the UK and Europe there is little published evidence of the effectiveness of this health promotion intervention. The aim of this study was to conduct a literature review of the effectiveness of established UK and Irish Smokebusters clubs. Over 60 clubs and agencies were contacted with a total of 36 reports received. Of those reviewed, most clubs have conducted process and impact evaluation to assess the popularity and quality of the programme. Attempts have been made to measure children's knowledge, attitudes and behaviour in relation to smoking and the Smokebusters intervention. Only three clubs have conducted long-term outcome evaluations which have measured changes in knowledge, attitudes and smoking behaviour. There is some evidence that changes occur in knowledge and attitudes after the establishment of clubs. To date, there are no reports of sustained change in smoking behaviour following the establishment of Smokebusters clubs. PMID- 10537941 TI - Health promotion research and the diffusion and institutionalization of interventions. AB - To examine the extent to which health promotion research is providing an empirical basis for the diffusion and institutionalization of effective interventions, we conducted a systematic audit of all articles in 12 public health and health promotion journals for the 1994 calendar year. We identified empirical/non-empirical and health promotion/non-health promotion articles. For each study, the health behaviours or outcomes studied, the target group, gender and setting were categorized. Each study was also categorized as belonging to one of four stages: basic research and development, innovation development, diffusion research, and research into institutionalization or policy implementation. Of all articles coded (n = 1210), 33.9% were identified as non-research, 39.5% were health promotion research and 26.6% were non-health promotion research. The vast majority of studies fell within the basic research and development stage (89.6%), with less than 1% categorized as diffusion research and only 5% as institutionalization or policy implementation research. The published studies reviewed provide a limited empirical basis for diffusion and institutionalization of health promotion programs. These findings suggest a need to more systematically monitor research input (funding) and research output (publications), and to develop a more explicit focus on the relevance of the stages of research innovation and development, the issues and/or behaviours addressed, the target population, and the research setting. PMID- 10537942 TI - Newspaper and wire service coverage of tobacco farmers. AB - This study examined print media coverage of tobacco farmers from the perspective of agenda setting, or the extent to which information is available to the public and perceived as important. A content analysis of 743 articles published between January 1, 1995 and June 30, 1997 was completed. The number of articles increased from 1995 to 1997. Of the topics analyzed, articles on tobacco settlement (7.1% of total) and diversification (15.6% of total) were the least prevalent. Because the settlement discussions did not occur until 1997 (when it comprised 26.4% of the total in the first 6 months), diversification was consistently the least covered topic. The two most frequent topics covered were tobacco companies (36.2%) and the tobacco price support program (32.3%). Except for one 6 month interval, there were substantially more articles in local/regional publications than in national publications. Public health professionals have called for tobacco farmers to diversify to non-tobacco enterprises. Yet, there is little discussion of diversification in print media. Without more attention to diversification, the public and policy makers will be ill-informed about opportunities and obstacles in this regard. PMID- 10537943 TI - Assessing decisional balance for smoking cessation among Southeast Asian males in the US. AB - This study examines the relationship of positive and negative perceptions of smoking to self-reported readiness to quit smoking among Southeast (SE) Asian males of Cambodian, Laotian or Vietnamese descent. In order to investigate this relationship, measures of decisional balance constructs (i.e. the pros and cons of smoking) appropriate for these ethnic groups were developed. Decisional balance was calculated by subtracting the cons from the pros. Following the criteria established by Prochaska and DiClemente, subjects were categorized into four levels of readiness to quit smoking (precontemplation, contemplation, preparation/action and maintenance). The expected pattern of relationship between decisional balance and stages of change included: (1) the cons of smoking being of less importance than the pros of smoking for those smokers in the precontemplation stage, (2) the pros and cons intersecting at the contemplation stage, and (3) the cons being of greater importance than the pros in the later stages of change. The SE Asian men in this study did not exhibit these decisional balance patterns, although mean decisional balance scores for precontemplators and contemplators were significantly more positive than mean scores for those in the preparation/action and maintenance stages. Decisional balance patterns differed across the three ethnic groups included in the sample. PMID- 10537944 TI - Counseled women's perspectives on their interactions with lay health advisors: a feasibility study. AB - Although the use of lay health advisors (LHAs) has become a popular intervention in public health promotion projects, few programs have conducted evaluations demonstrating program impact by interviewing people actually counseled by LHAs. This study used semistructured, in-person interviews with 29 older, black women to elicit their perceptions of their interactions with the North Carolina Breast Cancer Screening Program's LHAs, and the ways in which these interactions affected their mammography attitudes and behavior. Interview data indicate that a majority of the respondents felt that LHAs had helped them in some way; most said that talking to advisors made them think more positively about mammograms and/or consider getting one. LHAs influenced the women they counseled because the women knew the advisors well, felt comfortable talking to advisors about private issues, considered advisors to be credible sources of information about mammography and because advisors offered women support with respect to their mammography behavior. These results elucidate some of the mechanisms through which LHAs affect the attitudes and behavior of individuals in their social networks. PMID- 10537945 TI - Validity of scales measuring the psychosocial determinants of HIV/STD-related risk behavior in adolescents. AB - We examined the content, construct and concurrent validity of scales to assess beliefs and self-efficacy related to adolescents' sexual risk behavior. We addressed content validity in the scale development process by drawing on literature and theory, and by pre-testing items with focus groups. We used confirmatory factor analysis of two models, an intercourse involvement model and a condom use model, to assess construct validity. The final intercourse involvement model included three scales: norms about sexual intercourse, attitudes about sexual intercourse and self-efficacy in refusing sex. The final condom use model included five scales: norms about condoms, attitudes about condom use, self-efficacy in communicating about condoms, self-efficacy in buying/using condoms and barriers to condom use. After two alterations to the models, the chi 2 and other indices indicated that the data fit the models well. Supporting the concurrent validity of the scales, high school students who had never had sexual intercourse had more negative attitudes toward sexual intercourse among teenagers, perceived norms toward sexual intercourse among teenagers to be more negative and expressed greater self-efficacy in refusing sex than did those who had experienced sexual intercourse. Consistent condom users had more positive attitudes and norms about condoms, had higher self-efficacy in communicating about and buying/using condoms, and perceived fewer barriers to condom purchase and use than did inconsistent condom users. PMID- 10537946 TI - A story/dialogue method for health promotion knowledge development and evaluation. AB - Arguments have been made in favour of a constructivist or postpositivist approach to health promotion knowledge development and program evaluation, but little has been articulated about what such an approach would look like. This article describes a 'story/dialogue method' that was created with and for practitioners in response to their concerns that much of their practice did not lend itself to a positivist, or conventional, methodology. Derived from constructivist, feminist and critical pedagogical theory, and with roots in qualitative methods, the method structures group dialogue around case stories addressing particular generative practice themes. While intended for practitioner training, organizational development and evaluation, the method to date has been used primarily for training purposes. This article describes the method, provides an example of its application, and discusses its strengths, weaknesses and relevance to health promotion. PMID- 10537947 TI - The opportunities and effectiveness of the health promoting primary school in improving child health--a review of the claims and evidence. AB - School health programs have been part of schooling for most of this century. The health promoting school is a recently developed concept which seeks to provide a multifaceted approach to school health. Will it provide a better frame-work to assist schools address the health issues of their students? This paper examines the development of the health promoting school and identifies its structural components. It reviews the claims and evidence which have emerged from the school health research literature which focus on primary schools. Findings indicate health gains for primary school students are difficult to assess, and will most likely occur if a well-designed program is implemented which links the curriculum with other health promoting school actions, contains substantial professional development for teachers and is underpinned by a theoretical model. The paper concludes by discussing how improvements can be made in more accurately assessing the effectiveness of the health promoting primary school in improving school health. PMID- 10537948 TI - Adolescent alcohol use and the community health agenda: a study of leaders' perceptions in 28 small towns. AB - The study assessed leaders' perceptions of adolescent alcohol use as a public health issue in 28 small communities in northern Minnesota, as part of formative evaluation for a community-based intervention to reduce adolescent alcohol access and consumption. One hundred and eighteen leaders from five key community sectors were interviewed about their perceptions of social, health and alcohol-related problems in their communities. Analyses indicated that school representatives and police chiefs perceived adolescent alcohol use and related problems to be serious; newspaper editors mentioned other social problems more often; and mayors and business representatives did not perceive adolescent alcohol problems to be as serious. In relation to efforts to affect local policy, the study suggested government and business sectors in these communities may need to be educated about the problem to build its importance on the community agenda of health issues. Thus community leaders in some sectors may comprise a key target audience for intervention. PMID- 10537949 TI - Primary schoolchildrens' perceptions of smoking: implications for health education. AB - This paper suggests that there is a need, as early as Reception, to implement smoking intervention programmes in the local school curriculum. Findings from a cross-sectional study have shown that primary schoolchildren (4-8 years old) possess negative attitudes and beliefs about smoking, have as yet to establish regular patterns of smoking behaviour, and have a broad understanding of the nature of smoking. Health educators need to capitalize on this negative disposition toward smoking via early intervention; however, to date, there are no smoking-specific health education measures for this age group. The implementation of proactive programmes, before the habit manifests itself, has many supporters but little research has been conducted. This study was devised to fill this significant gap in the literature on smoking. Data was collected on a representative sample of primary schoolchildren in the city of Liverpool. A triangular methodology was adopted consisting of questionnaires (N = 1701), the Draw and Write investigative technique (N = 976), and semi-structured interviews (N = 50). The results highlight the need to implement smoking intervention programmes from Reception onward, the importance of developing a model that is more than just knowledge based and the necessity of involving the family in any school-based health education strategies. PMID- 10537950 TI - Drug education practice: results of an observational study. AB - Understanding normative practice in drug education is a key to identifying means of improving preventive intervention outcomes. In this paper, we report findings of an observational study in which drug education in multiple periods of 146 middle school classes was categorized minute-by-minute according to the type of instruction provided to students. Results indicate that nearly half of all drug education focused on providing students with knowledge. Alternative methods, particularly those that have shown programmatic effectiveness, and those that address risk and protective factors known to be highly predictive of drug use onset, were relatively ignored. Further, teachers showed relatively low consistency in understanding concepts other than knowledge based on comparisons of their ratings of intended instructions with those of trained observers. Nonetheless, there is evidence that some teachers systematically attempted to address drug prevention from either a social influence or an affective education perspective. These findings suggest that if improvements in the effectiveness of drug education are to be seen in the future, a relatively radical transformation of approaches to teaching will be needed. PMID- 10537951 TI - Student-school bonding and adolescent problem behavior. AB - Adolescent problem behavior, including substance use, school misconduct and delinquency, is a national concern. Implicit in the concept of middle school is the recognition that students who develop positive social bonds with their school are more likely to perform well academically, and refrain from misconduct and other antisocial behavior. However, little scientific attention has been given to the complex interactions between middle school students and the school environment. Prior to implementing a middle school problem behavior prevention program we conducted a survey in the seven middle schools in one US school district. Out of 4668 grade 6-8 students enrolled, 4263 (91.3%) completed the survey. Student-school bonding was positively correlated with school adjustment (r = 0.49) and perceived school climate (r = 0.77), but inversely correlated with problem behavior (r = -0.39 to -0.43). Problem behavior was significantly higher (P < 0.001) among males than females and among students in higher grades. Conversely, school bonding, climate and adjustment were significantly higher (P < 0.001) among females than males, but declined significantly from one grade to the next. The data support the conclusion that school bonding is associated with problem behavior. We describe the development of a multiple-component intervention in middle schools designed to increase student-school bonding and prevent problem behavior. PMID- 10537953 TI - Methodological hurdles in conducting pharmacoeconomic analyses. AB - As total healthcare spending increases throughout the world, greater emphasis is being placed on research which demonstrates value for medical interventions, including new and existing pharmaceuticals. Pharmacoeconomic evaluations can assist manufacturers, insurers, clinicians, governmental agencies, policy-makers and consumers to make informed, appropriate decisions about adoption and application of new medications. Because of the far-reaching implications of this research, it is important that researchers adequately address methodological challenges. In this article, we describe the uses of results of pharmacoeconomic trials, identify and discuss various study designs and methods for gathering nonclinical outcome data which may differ significantly from clinical outcome data, and consider the importance and difficulty of incorporating the patients' experience into such trials. Researchers in this area must give specific consideration to sample size estimation for economic outcomes, and carefully handle time issues including duration of observation for complications and discounting of future health and financial consequences. Costs from different perspectives associated with resource use should be assembled in a standard fashion. Use of charges which may not be standardised across geographical or organisational boundaries are discouraged. Inclusion of appropriate health related quality-of-life (HR-QOL) and utility instruments is increasingly important, but controversy over the best methods still exists. While there is little question of the importance of pharmacoeconomic evaluations, they are expensive. Well designed and executed pharmacoeconomic trials can justify this expense by helping decision-makers understand which treatments have value. PMID- 10537955 TI - Utility scores for chronic conditions in a community-dwelling population. AB - OBJECTIVE: The objective of this study was to determine utility scores for various chronic conditions. DESIGN AND SETTING: This study is a descriptive analysis. Health Utilities Index (HUI) scores for 20 chronic conditions were examined from the National Population Health Survey (NPHS) from 1994 to 1995. PATIENTS AND PARTICIPANTS: 17,626 individuals were surveyed (54.3% women). Chronic conditions included: acne (requiring medication), Alzheimer's disease, arthritis/rheumatism, asthma, back problems excluding arthritis, chronic bronchitis or emphysema, cancer, cataracts, diabetes, epilepsy, food allergies, glaucoma, heart disease, high blood pressure, migraine headaches, other allergies, sinusitis, stroke, stomach/intestinal ulcers and urinary incontinence. INTERVENTIONS: Health Utilities Index-Mark III (HUI-Mark III) scores for patients with and without a NPHS-defined chronic condition were collected. Utility scores were examined according to age, gender and comorbidity. MAIN OUTCOME MEASURES AND RESULTS: 42.6% of individuals reported having no NPHS-defined chronic condition. The most commonly reported health conditions were allergies other than food (17.6%) and rheumatism/arthritis (16.5%). The mean HUI-Mark III scores for patients without a health state was 0.933 +/- 0.079. Individuals with Alzheimer's disease (0.580 +/- 0.263), stroke (0.676 +/- 0.230) and urinary incontinence (0.698 +/- 0.230) had the lowest overall HUI-Mark III scores. Utility scores decreased as age and as the number of comorbid conditions increased. CONCLUSIONS: This study provides health economists, researchers and policy-makers with a reference for health utilities of various chronic conditions, different age groups, gender and comorbidities. PMID- 10537952 TI - The clinical and economic potential of cyclosporin drug interactions. AB - The introduction of cyclosporin significantly improved solid organ transplantation outcomes. However, the costs associated with immunosuppressive therapy increased from approximately $US1000 to $US2000 per patient per year with azathioprine (AZA) and prednisone to $US5000 to $US8000 per patient per year with the addition of cyclosporin (1997 values). Because of the financial demands placed on medical care in the current era, research has been directed towards developing drug combinations which potentiate the therapeutic effect of cyclosporin whereby reducing the amount of drug administered and consequently the costs of long term immunosuppressive therapy. To date, many drugs that interact with cyclosporin have been recognised. Included in this list are the azole antifungal drugs, ketoconazole, fluconazole and itraconazole; the calcium channel blockers, diltiazem, verapamil and nicardipine; and the macrolide antibacterials, erythromycin and related compounds. Although all of these drugs increase cyclosporin drug concentrations when used concomitantly, ketoconazole and diltiazem appear to be the best candidates on the basis of reducing financial pressures of chronic immunosuppressive therapy without sacrificing patients' well being. Studies of various regimens involving the combined use of ketoconazole and cyclosporin have shown that cyclosporin dosages can be reduced by approximately 70 to 85% while maintaining therapeutic blood concentrations in renal, cardiac and liver transplant recipients. The calcium channel blocker, diltiazem, allows a decrease in cyclosporin dosage by approximately 30 to 50% in this same group of organ transplant patients. These reductions in cyclosporin dosage have been achieved with no reported severe adverse effects that would discourage the use of these agents concurrently in practice. The combined use of cyclosporin and ketoconazole or diltiazem could reduce medication costs by approximately $US915 to $US3000 per year per patient. If all patients treated with cyclosporin are considered, these combinations could reduce medication costs by hundreds of millions of dollars per year in the US alone. While these are promising approaches, further characterisation of these drug interactions is necessary before this practice is adopted as standard protocol worldwide. The objective of this paper is to review the clinical and economic potential of cyclosporin sparing agents such as the azole antifungal drugs and calcium channel blockers in an attempt to decrease the costs associated with this expensive immunosuppressive agent. PMID- 10537954 TI - Recent advances in the methods of cost-benefit analysis in healthcare. Matching the art to the science. AB - This paper outlines recent advances in the methods of cost-benefit analysis (CBA). Economic evaluations in healthcare can be criticised for, amongst other things, the inappropriate use of incremental cost-effectiveness ratios and the reporting of benefits in terms of cost savings, such as treatment costs averted. Many such economic evaluations are, according to the 'scientific' definition, CBAs. The 'balance-sheet' (or opportunity cost) approach is a form of CBA which can be used to identify who bears the costs and who reaps the benefits from any change. Whilst the next stage in a CBA, as defined in health economics, would require that all costs and benefits be valued in monetary terms, the balance sheet approach, however, advocates that available monetary values can be augmented by other measures of cost and benefit. As such, this approach, which has a theoretical basis, is proposed as a practical prescription for CBA and highlights the notion that unquantified benefits are important and can be included within CBAs even when monetarisation is not possible. Recent methodological developments in monetary valuation for use in CBA are the development of the technique of willingness to pay, the use of conjoint analysis (CA) to elicit willingness-to-pay (WTP) values and advances in the debate on the inclusion of production gains in CBAs. Whilst acknowledging that there have been developments in each of these areas, it is claimed there has also been progress in using CBA as a framework for evaluation, as reflected by the balance-sheet approach. The paper concludes by stating that almost all types of economic evaluation have an element of the 'cost-benefit' approach in them. The important issue is to focus on the policy question to be addressed and to outline the relevant costs and benefits in a manner which assists the evaluation of welfare changes resulting from changes in healthcare delivery. The focus should not be on moulding a question to fit a hybrid definition of an analytical technique. PMID- 10537956 TI - Treatment of locally advanced pancreatic carcinoma in Sweden. A health economic comparison of palliative treatment with best supportive care versus palliative treatment with gemcitabine in combination with best supportive care. AB - OBJECTIVE: Patients with pancreatic cancer have only short survival after diagnosis, irrespective of treatment. The aim of this study was to perform a health economic evaluation of present standard treatment (in most cases, palliative treatment in combination with best supportive care) versus palliative treatment with gemcitabine in combination with best supportive care in patients with locally advanced pancreatic carcinoma. DESIGN: The use of resources and associated costs according to present treatment practice were estimated and calculated retrospectively. Costs were calculated from diagnosis until death. Actual costs and treatment effects for the patient population were compared with expected treatment costs for the same population if they additionally received gemcitabine. SETTING: This economic analysis is based on a hypothetical comparison and was performed from a societal point of view. PATIENTS AND PARTICIPANTS: The study population consisted of all patients diagnosed with pancreatic cancer during the year April 1994 to March 1995 and resident in Stockholm County, Sweden. After exclusions, 184 patients were included in the economic analysis. INTERVENTIONS: The effects of gemcitabine treatment on survival and disease-related symptoms were extrapolated from the results of a recent randomised clinical trial in North America. MAIN OUTCOME MEASURES AND RESULTS: The estimated additional costs for chemotherapy, treatment of adverse effects and in- and outpatient care associated with gemcitabine treatment were approximately 132,000 Swedish kronor (SEK) per life-year gained. This result is comparable with costs per life-year gained for other accepted treatments, for example those of home dialysis and kidney transplants for chronic renal failure. CONCLUSIONS: Treatment with gemcitabine in patients with pancreatic cancer may be a cost-effective alternative, but the results need to be confirmed in future randomised trials. PMID- 10537957 TI - Applying epidemiology-based outcomes research to clinical decision-making. A hypothetical model of biotechnology therapy in gram-negative sepsis. AB - OBJECTIVE: Sepsis occurs in a heterogeneous population. A prospective nationwide surveillance study found that populations stratified by infection type had significant differences in the incidence of sepsis syndrome, rate of complications and mortality. The objective of this study was to explore whether successful identification of population-specific risk factors for disease associated morbidity and mortality may allow for more accurate assessment of the cost effectiveness of treatment strategies. DESIGN: A decision analytic model was developed using outcomes data on incidence and resolution of major complications in sepsis syndrome. Healthcare resource utilisation data were based on length of hospital stay, intensive care unit stay versus hospital ward stay, and cost of treating sepsis-related complications. SETTING: This modelling study, conducted from the perspective of the healthcare institution, used actual outcomes data on 2 infection-specific patient populations. PATIENTS AND PARTICIPANTS: The 2 populations studied were patients with nosocomial respiratory tract infection or community-acquired urinary tract infection who subsequently developed sepsis syndrome. INTERVENTIONS: Treatment options modelled were standard therapy plus biotechnology therapy versus standard therapy alone in the treatment of gram negative sepsis complications. MAIN OUTCOME MEASURES AND RESULTS: The incremental cost-effectiveness ratios differed between the 2 study populations, due to differences in the incidence and rate of resolution of major sepsis-associated complications. The use of biotechnology therapy is always more cost effective in the respiratory tract infection population. CONCLUSIONS: Cost-effectiveness results for a therapy may change when the epidemiology of the disease state is known and incorporated into the decision analytic model. An infection-specific approach is important in the treatment of sepsis. PMID- 10537958 TI - Cost analysis of the treatment of severe spinal spasticity with a continuous intrathecal baclofen infusion system. AB - OBJECTIVE: The purpose of our study was to analyse and evaluate the costs of continuous intrathecal baclofen administration as a modality in the treatment of severe spasticity in the Netherlands. DESIGN: A cost analysis was conducted as part of a prospective, multicentre, multidisciplinary, randomised and placebo controlled clinical trial. The study covered the period from December 1991 to September 1995. The data on medical consumption and costs were collected over a 3 year period from different sources: administrative databases of health insurance companies, hospital registries and a patient survey. These data were structured by means of a flowchart analysis of the medical decision-making by specialists and general practitioners (GPs). They included data on in- and outpatient care, home care and care in nursing homes. The cost analysis was conducted using data from 18 patients included in the trial and from 15 so-called 'match' patients. The latter group are patients with comparable diseases leading to spasticity and living in comparable circumstances. Next to absolute costs (direct and indirect) of care and treatment for the 2 groups of patients, cost differences between the 2 groups were considered (differential cost analysis). SETTING: Per patient cost data, collected prospectively for 2 years during the phase of clinical evaluation, and retrospectively 1 year before implantation. The data were collected on patients from in- and outpatient care, home care and care in nursing home settings. PATIENTS AND PARTICIPANTS: The trial patients (8 men) had a mean age of 46 years; 11 patients had multiple sclerosis and 7 patients had spinal cord injuries. The match patients (7 men) had a mean age of 48 years; 9 patients had multiple sclerosis and 6 patients had spinal cord injuries. INTERVENTIONS: Trial patients were treated with a subcutaneously implanted programmable continuous infusion pump (SynchroMed, Medtronic), filled with baclofen (a muscle relaxant) to treat patients with chronic disabling spasticity who did not respond to a maximum dose of oral baclofen, dantrolene and tizanidine. MAIN OUTCOME MEASURES AND RESULTS: An analysis of hospital stay between both groups showed a significant difference during the implantation year. The average number of hospital days per patient in the year in the treated group was 31.5 days and in the match group was 18.7 days. Significant cost differences between both groups in the year that started with pump implantation and the following year can be attributed mostly to the costs of implantation of the pump and related hospitalisation days. The total costs of patient selection, testing, implanting the pump and follow-up amounted to $US28,473 for the first year. Savings must be taken into consideration as well. The savings of direct costs were due to withdrawal of oral medication (estimated annual total of between $US1950 and $US2800 per patient). Indirect savings on employment and nursing home costs, amounted annually to $US1047 and $US5814, respectively. Scenarios make it possible to consider policy consequences. The case of 'extending' the indications for this treatment to a larger population has been calculated and visualised. CONCLUSIONS: The costs of the therapy (continuous intrathecal infusion of baclofen) can be attributed mostly to implantation of the pump and related hospitalisation days. Savings originated from withdrawal of oral medication, job preservation and avoidance or delay of admission to a nursing home. PMID- 10537959 TI - Vinorelbine in advanced non-small cell lung cancer. A pharmacoeconomic review. AB - Vinorelbine is a semisynthetic vinca alkaloid that is effective against advanced non-small cell lung cancer (NSCLC). Myelosuppression is the primary dose-limiting toxicity; vinorelbine is otherwise relatively well tolerated. Two studies assessed the cost effectiveness of vinorelbine with or without cisplatin based primarily on data from a phase III comparison with vindesine plus cisplatin. Survival and cost data from this study were supplemented with those from other sources. One model simulated total management costs for the 4986 patients diagnosed with stage IV NSCLC in Canada in 1992. The other applied US cost data to the outcomes from the phase III trial. Using vinorelbine monotherapy or vinorelbine plus cisplatin produced a survival benefit and net cost savings compared with best supportive care according to the Canadian model (and preliminary data from a third analysis, conducted in the US). In the Canadian analysis, incremental cost effectiveness for inpatient or outpatient vinorelbine plus cisplatin ranged from 7450 Canadian dollars ($Can) to $Can30,770 (1993 values) per year of life saved (YLS) compared with outpatient cisplatin plus either etoposide or vinblastine. Cost-effectiveness ratios for vinorelbine plus cisplatin in the US analysis (1994 values) were $US18,000 (vs cisplatin plus etoposide) and $US15,500 (vs cisplatin plus vindesine) per YLS [all inpatient administration]. Detailed pharmacoeconomic comparisons with other current standard regimens (e.g. paclitaxel plus either cisplatin or carboplatin) are not available. Sensitivity analyses suggest that the cost effectiveness of vinorelbine-based therapy is robust to changes in assumptions regarding efficacy and the cost of managing toxicity. Limitations of the available pharmacoeconomic data include the retrospective nature of the analyses, inclusion of data from sources other than the main phase III trial (e.g. those for best supportive care and some chemotherapy regimens), and exclusion of some costs for hospitalisation and/or management of toxicity. CONCLUSIONS: Although some limitations apply, the available data suggest that vinorelbine alone or in combination with cisplatin is cost saving compared with best supportive care for NSCLC, and that vinorelbine plus cisplatin is cost effective compared with some other combination regimens. The pharmacoeconomic placing of vinorelbine in relation to a number of other currently recommended first-line treatments for NSCLC has yet to be resolved, and data from ongoing multicentre phase III trials are awaited with interest. In the meantime, vinorelbine-based chemotherapy appears to be a suitable choice for first-line treatment of advanced NSCLC from both clinical and pharmacoeconomic perspectives. PMID- 10537960 TI - Pharmacoeconomic evaluation in the real world. Effectiveness versus efficacy studies. AB - Pharmacoeconomic data may be obtained within the context of randomised clinical trials (RCTs) and from effectiveness studies in the 'real world'. The differences between the 2 types of study design have implications for the types of data that can be obtained and the interpretation of the resulting findings. Because RCTs are designed to assess the safety and efficacy of pharmaceuticals, and because the study design of RCTs emphasises internal validity over generalisability, the pharmacoeconomic data collected from them are limited. The data may not be applicable to the more heterogeneous patients encountered in actual clinical practice, and cost estimates may be inaccurate because of protocol requirements. Effectiveness studies, in which treatments are studied under real-world conditions, remedy some of these limitations. Generalisability to actual users is generally enhanced in effectiveness designs, but data may be biased in other ways. This brief review compares the 2 study designs as they relate to pharmacoeconomic evaluations in terms of the research questions they address, design differences and their implications for study bias, data collection and data analysis and the generalisability of their results. PMID- 10537961 TI - Prescribing at the interface between primary and secondary care in the UK. Towards joint formularies? AB - The current divisions in managing prescribing between primary and secondary care in the UK arise from separate budgetary arrangements. These divisions are neither sensible, organisationally efficient nor cost effective. Transition of patients across the interface of primary and secondary care has always been problematic, hindered by poor communication and coordination. Joint formularies would improve overall care and raise awareness of the need to consider overall costs within a unified National Health Service (NHS). There are, however, few examples of successful working of a joint formulary in the UK. It is likely that harmonisation of drug use in hospitals and in primary care will come about because of contracting and commissioning, and that it will largely be led by primary care, through the developing primary care groups (PCGs). Local decisions around availability and use of drug therapies will increasingly be superseded by the national decisions emanating from the newly formed National Institute for Clinical Effectiveness. PMID- 10537962 TI - Counting the costs of drug-related adverse events. AB - Adverse drug events occur frequently and lead to a significant number of fatalities each year. It has been estimated that fatalities directly attributable to adverse drug reactions are the fourth to sixth leading cause of death in US hospitals, exceeding deaths caused by pneumonia and diabetes. The economic burden resulting from drug-related morbidity and mortality is equally significant and has been conservatively estimated at $US30 billion dollars annually, and could exceed $US130 billion in a worst-case scenario. Since many adverse drug events are considered preventable, increased efforts should be made to avoid classes of drugs that are problem-prone and to initiate diligent monitoring of drugs with predictable toxicities. Programmes should also be implemented that improve medication use practices within institutions. Although nearly all drugs are capable of producing an injury, certain drugs are more likely to do so. Prevention of drug-related morbidity and mortality has become an increasingly important requirement for reducing healthcare expenditures. This article will review studies that examine the economic implications of drug-related adverse events. PMID- 10537963 TI - The revised Canadian Guidelines for the Economic Evaluation of Pharmaceuticals. AB - The first edition of the Guidelines for Economic Evaluation of Pharmaceuticals: Canada was published in November 1994. At that time, the Canadian Coordinating Office for Health Technology Assessment (CCOHTA) was assigned the task of maintaining and regularly updating the Canadian Guidelines. Since their introduction, a great deal of experience has been gained with the practical application of the guidelines. Their role has also evolved over time, from being a framework for pharmacoeconomic research to the point where a wide variety of decision-makers use economic evaluations based on the principles set out in the guidelines as a means of facilitating their formulary decisions. In addition, methodologies in certain areas (and the body of related research literature in general) have developed considerably over time. Given these changes in the science and the experience gained, CCOHTA convened a multi-disciplinary committee to address the need for revisions to the guidelines. The underlying principles of the review process were to keep the guidance nature of the document, to focus on the needs of 'doers' (so as to meet the information needs of 'users') and to provide information and advice in areas of controversy, with sound direction in areas of general agreement. The purpose of this review is three-fold: (i) to outline the process which lead to the revision of the Canadian Guidelines; (ii) to describe the major changes made to the second edition of this document; and (iii) to consider the 'next steps' as they relate to the impact of such guidelines and the measurement of outcomes related to economic assessments of pharmaceuticals in general. PMID- 10537964 TI - Clinical and economic outcomes of olanzapine compared with haloperidol for schizophrenia. Results from a randomised clinical trial. AB - OBJECTIVE: The purpose of this study was to compare, from the payor perspective, the clinical and economic outcomes of olanzapine to those of haloperidol for the treatment of schizophrenia. DESIGN AND SETTING: Clinical, quality-of-life and resource utilisation data were prospectively collected for US-residing patients with schizophrenia who were participating in a multicentre, randomised, double blind clinical trial comparing olanzapine and haloperidol. Direct medical costs were estimated by assigning standardised prices (1995 values) to the resource utilisation data. PATIENTS AND PARTICIPANTS: 817 patients with schizophrenia who had a baseline Brief Psychiatric Rating Scale score (BPRS) > or = 18 (items scored 0 to 6) and/or were no longer tolerating current antipsychotic therapy. INTERVENTIONS: Olanzapine 5 to 20 mg/day (n = 551) or haloperidol 5 to 20 mg/day (n = 266) for 6 weeks. Patients showing a predefined level of clinical response entered a 46-week maintenance phase. MAIN OUTCOME MEASURES AND RESULTS: After acute treatment, BPRS-based clinical improvements were seen in 38 and 27% of olanzapine and haloperidol patients, respectively (p = 0.002). Clinically important improvements on the Quality of Life Scale were achieved during acute treatment in 33% of olanzapine recipients and 25% of haloperidol recipients (p = 0.094). Olanzapine treatment in the acute phase led to significantly lower inpatient ($US5125 vs $US5795, p = 0.038) and outpatient ($US663 vs $US692, p = 0.001) costs, resulting in a significant overall reduction in mean total medical costs of $US388 (p = 0.033). This significant reduction in total costs was found despite olanzapine mean medication costs being significantly greater than haloperidol medication costs ($US326 vs $US15, p < 0.001). No significant differences in clinical improvement were observed in the maintenance phase. Maintenance phase olanzapine mean total medical costs were $US636 lower than haloperidol total costs (p = 0.128). Although olanzapine medication costs were significantly higher than haloperidol medication costs ($US3461 vs $US95, p < 0.001), this difference was offset by significantly lower inpatient ($US8322 vs $US10,662, p = 0.044) and outpatient ($US3810 vs $US5473, p = 0.038) costs. CONCLUSIONS: In this study, olanzapine treatment was more effective than haloperidol in producing clinical response in the acute phase. In addition, olanzapine treatment led to reductions in inpatient and outpatient costs that more than offset olanzapine's higher medication costs relative to haloperidol. PMID- 10537965 TI - Reconciling decision models with the real world. An application to anaemia of renal failure. AB - OBJECTIVE: The choice of evidence used in decision modelling of healthcare interventions divides analysts into 2 groups: (i) those who favour randomised clinical trial (RCT) data; and (ii) those who prefer 'real world' data. This preference may have serious consequences if the end result is to inform healthcare policy. This paper uses Medicare coverage of epoetin-alpha [erythropoietin (EPO)] as a case study to illustrate a technique which can be used to overcome some of the bias inherent in RCT data while avoiding some of the common pitfalls associated with the use of observational data. DESIGN AND SETTING: Cost analysis of 2 treatments for anaemia of renal failure primarily in an outpatient setting is modelled in a decision tree. This method can be used to analyse healthcare interventions or policies in any setting. PATIENTS AND PARTICIPANTS: Patients with nontransplanted end-stage renal disease (ESRD) who received either EPO or blood transfusion for treatment of anaemia at any time during the 1-year study period (July 1989 to June 1990) were included in the sample. METHODS: Outcome effects in the natural setting are decomposed into 2 parts: a treatment effect and a population effect. This is then extended to the special case of policy analysis. Logistic and multiple regression are used to estimate branch probabilities and payoffs, respectively, for 2 treatment options. MAIN OUTCOME MEASURES AND RESULTS: Under standard methods of decision analysis, an increase of $US7032 per patient following EPO coverage is observed. With the decomposition technique, the policy effect is estimated to be less, $US6172, the difference coming from the population effect. CONCLUSIONS: Failure to remove population effects from observed outcome effects may lead to biased decision making. Although not directly observable, the population effect can be imputed from secondary data. The decomposition and imputting technique allows for a more meaningful interpretation of the results for the purpose of policy analysis. PMID- 10537966 TI - Medical resource use and cost of venlafaxine or tricyclic antidepressant therapy. Following selective serotonin reuptake inhibitor therapy for depression. AB - OBJECTIVE: An analysis of administrative and claims data was performed to compare the resource use and costs to a managed-care organisation of venlafaxine, a serotonin and norepinephrine reuptake inhibitor (SNRI), versus tricyclic antidepressant (TCA) therapy, after switching from a selective serotonin reuptake inhibitor (SSRI). DESIGN: One-year costs and frequencies of all medical services, and of services coded for depression, were compared between patients who received venlafaxine and TCA therapy as second-line therapy using bivariate and multivariate statistical analyses. SETTING: Data were obtained from 9 individual health plans with more than 1.1 million covered lives affiliated with a national managed-care organisation. PATIENTS AND PARTICIPANTS: Health plan members were included if they had a diagnosis of depression between July 1993 and February 1997. They also had to have at least 2 months of prescriptions for SSRI therapy followed by at least 2 months of venlafaxine or TCA therapy, and continuous enrollment in the plan from at least 6 months prior to 12 months following initiation of venlafaxine or TCA therapy. 188 patients who received venlafaxine and 172 patients who received TCAs met the inclusion criteria. MAIN OUTCOME MEASURES AND RESULTS: Patients who received TCAs were slightly but significantly older (43 vs 40 years) than venlafaxine recipients and, during 6 months prior to initiating therapy, had significantly higher mean costs coded for depression ($US451 vs $US311) and costs not coded for depression ($US4500 vs $US2090). Psychiatrists prescribed a significantly higher proportion of venlafaxine than TCA prescriptions (46.3 vs 25.0%). Prior to adjusting for confounding characteristics, during 12 months following initiation of therapy, mean depression-coded costs were significantly higher for venlafaxine than TCA recipients ($US1948 vs $US1396) and mean costs not coded for depression were significantly lower ($US4595 vs $US6677). Overall costs were not significantly different ($US6543 for venlafaxine vs $US8073 for TCA). Significant cost differences were observed with primary care physicians as initial prescribers of second-line therapy but not with psychiatrists. However, costs between the 2 groups were similar after adjusting for confounding variables, including prior 6 month costs and initial prescriber of second-line therapy. CONCLUSIONS: Payer costs are similar among patients receiving venlafaxine and TCA therapy following SSRI therapy. Higher costs of venlafaxine pharmacotherapy relative to TCA therapy may be offset by lower costs of other medical services. Differences in prescribing patterns and costs between primary care physicians and psychiatrists warrant further investigation. PMID- 10537968 TI - Connectivity at core of POC growing pains. PMID- 10537969 TI - Combine sets up unrivaled leadership. PMID- 10537967 TI - Direct costs of hip fractures in patients over 60 years of age in Belgium. AB - OBJECTIVE: Osteoporosis-related costs are now considered a major burden for health authorities in most developed countries. An accurate and exhaustive evaluation of these costs would be a major contribution to health economic studies evaluating the efficiency of screening and prevention strategies. Osteoporosis is the most frequent underlying cause of femoral neck fractures in the elderly; these fractures weigh heavily on healthcare budgets. However, in Belgium, very few data on the financial burden of hip fractures are available and no updated estimates have been made. The goal of this paper is to estimate the direct medical expenditures associated with hip fractures in Belgium in 1996. DESIGN AND SETTING: This 1-year population-based cross-sectional study is conducted from the social security perspective. The target population in this study are men and women aged 60 years and over. PATIENTS AND PARTICIPANTS: We selected patients who had been hospitalised for a hip fracture during the year 1996 who were also affiliated with a registered social security organisation (covering 25% of the Belgian population). The sample constituted 2374 patients. INTERVENTIONS: For each of these patients, we collected an exhaustive and detailed list of healthcare resource use as well as nursing home admissions following the hip fracture event. Cost items investigated in the analysis were inpatient hospital costs and outpatient costs. Mean annual costs per case recorded in the sample were then extrapolated to the whole country on the basis of an exhaustive list of diagnoses having lead to all countrywide hospitalisations (1,700,000 hospital stays/year). MAIN OUTCOME MEASURES AND RESULTS: The mean hospital inpatient costs for hip fracture were evaluated at 332,148 Belgian francs (BeF) [$US8977] per case and BeF4,367,746,200 ($US118,047,194) for the whole country (10 million inhabitants). Patients with a hip fracture experienced an annual BeF27,825 ($US752) extra outpatient cost during the year following this fracture event, after correcting for costs related to additional comorbidity already present before the hip fracture. Finally, after a proximal femoral neck fracture, the rate of nursing home admission was higher, both for men and women at any age compared with age- and gender-matched population. CONCLUSIONS: With a total cost (acute hospital and outpatient costs) of BeF4,667,894,950 ($US126,159,323) per year in Belgium, proximal femoral neck fracture should be considered a major health economic problem and appropriate measures to prevent this disease should be rapidly undertaken. PMID- 10537970 TI - In search of risk-related genes for heart disease. PMID- 10537971 TI - Putting paper in its place. PMID- 10537972 TI - Automated immunoassay analyzers. PMID- 10537973 TI - Pushing proficiency testing results to their limits. AB - PT is a valuable component of laboratory quality control and management practice. PT results are useful to determine over time that a method is performing in a laboratory according to the manufacturer's design parameters, as exhibited by the performance characteristic of a large group of peer laboratories. PT peer group results give a good measure of the interlaboratory imprecision of a method, and summary reports document the state-of-the-art in user preference. PT results typically cannot be used to validate the accuracy of a method or its clinical utility or to compare the accuracy of various methods. These limitations are caused by the common and unpredictable matrix interferences with PT materials and most routine laboratory methods. In situations where a laboratory adjusts the calibration of one method to make patient results agree with those of another method, PT specimens results must be reported without calibration adjustments to allow those results to be compared to an appropriate peer group mean target value. PMID- 10537974 TI - Automated billing--new generation of systems provides tools to improve compliance. PMID- 10537975 TI - Keeping pace with managed behavioral health care accreditation trends. PMID- 10537976 TI - Poor public relations and liability. PMID- 10537977 TI - The state of the art of PPO quality and performance measurement. PMID- 10537978 TI - Handbook of telemedicine. PMID- 10537979 TI - Improving public financing for long-term care: the political challenge. PMID- 10537980 TI - Enabling informed consumer choice in the long-term care insurance market. AB - Provisions in the Health Insurance Portability and Accountability Act of 1996 (HIPAA) may increase private long-term care insurance sales without imposing substantially more stringent consumer-protection features. The ability of consumers to make informed choices when purchasing this complex product is examined in light of these changes. Data were collected through detailed examinations of policies and interviews with industry experts, insurance companies, agents, consumer groups, and regulators. Because of the complexity of this product, the goals of expanding, consumer choice and ensuring that consumers are able to make informed decisions often work against each other. Mechanisms are discussed through which the government can facilitate informed choice and improve consumer protection. The authors contend that, because the government is providing tax incentives that encourage consumers to purchase the product, it has the responsibility to ensure that consumers understand the long-term care insurance they purchase. PMID- 10537981 TI - Impacts of welfare reform on California grandparents raising grandchildren: reflections from the field. AB - Debate over the potential impacts of welfare reform largely has ignored the implications of these changes for the growing number of grandparents who are raising their grandchildren. Results of a qualitative study involving 36 key informants who were intimately involved in the crafting and/or implementation of California's welfare reform plan are presented. Particular attention is focused on time limits on aid, work requirements, and sanctions regarding teenage parenthood as these may impact on grandparent caregivers and their families. Cross-cutting themes also are presented. A case is made for greatly stepping up data collection and evaluative research that may help in determining the actual impacts of the legislation on intergenerational households headed by grandparents. PMID- 10537982 TI - Increasing older persons' employment in Finland: in search of a new strategy. AB - Using Finland as a case study, it is argued that early retirement will probably no longer be used on a large scale to reduce older-worker labor-force participation and unemployment among older workers. Instead, new strategies are needed to enhance the ability of older workers to remain productive and in the labor force, and to facilitate the reintegration of unemployed older persons back into working life. Both tasks require massive pioneering efforts. Reducing unemployment rates among older workers, particularly, requires completely new kinds of labor-market measures. PMID- 10537983 TI - Resident-centered care in assisted living. AB - Residents (n = 396) at 20 assisted living (AL) settings were interviewed as were program staff and administrators to understand how resident choice, getting needed care, and a sense of community were promoted or hindered. Residents reported relatively independent and autonomous lives, yet many experienced unmet health and long-term care needs and limited participation in meaningful activities or community life. Strong support was found for the hypothesis that AL program and site features influence resident experiences, particularly in regard to supporting independent lifestyles, minimizing avoidable care problems, and increasing community involvement. PMID- 10537984 TI - Gradual retirement in Germany. AB - In the summer of 1996, the German parliament enacted legislation (the "Gesetz zur Forderung cines gleitenden Ubergangs in den Ruhestand") that would allow a gradual transition from work to retirement by permitting workers 55 years and older to change from full-time to part-time work, and also by making it more difficult to take early retirement. The intent was to stop the trend towards early retirement. This article discusses the provisions of the new legislation, as well as the experiences with previous models that influenced its development. These experiences were mainly negative because older workers were unwilling to change to part-time work and employers did not provide attractive part-time work opportunities. The new legislation's chances of success are discussed and suggestions are offered for making the concept of gradual transition from work to retirement more attractive and viable. PMID- 10537985 TI - Patient focused care--better for patients and hospital staff. PMID- 10537986 TI - The best of health 3. Are we doing the right things, and are we doing those things right? PMID- 10537987 TI - A maternity cooperative that works. PMID- 10537988 TI - Integrated care. PMID- 10537989 TI - Dealing with CHE deficits is a high priority for THA (Transitional Health Authority). PMID- 10537990 TI - OSHA's working draft of an ergonomics standard. PMID- 10537991 TI - DuraPrep solution fire hazard. PMID- 10537992 TI - Developing human resources through discipline and morale. PMID- 10537993 TI - Role of computers in care of disabled. PMID- 10537994 TI - Absenteeism--a financial bottleneck and a resource drain. PMID- 10537995 TI - Laboratory medicine and hospital administration, interfaces at conceptual level. PMID- 10537996 TI - The cutting edge of HRD in health care organisations. PMID- 10537997 TI - Hospital waste disposal by incineration. AB - Hospital waste is becoming increasingly complex due to changing technologies and increase in the services that the hospitals perform for the community. Out of the available technology for the final disposal of solid wastes, incineration is best suited for hospital waste as it renders the waste nontoxic, non hazardous, non putrescible and reduces the volume of material for ultimate disposal. Present study was carried out in a service hospital to analyze the requirement of incinerator considering the state of art available in this country. Multi chambered oil fired incinerator installation as an on site facility for hospital solid waste disposal has been recommended as more environment. friendly option. PMID- 10537998 TI - Proper donor screening--a good prevention of transfusion transmitted AIDS and hepatitis. PMID- 10537999 TI - Socio-demographic pattern of drug and alcohol dependents. AB - The widespread abuse of drugs and Alcohol has become a human tragedy. Each year the abuse of Alcohol and Illicit drugs exact an enormous toll in deaths, decline in productivity, more crime and accidents and also increased expenditure in rehabilitation. The situation is likely to worsen and even may get out of hand if adequate measure are not taken to clearly identify the vulnerable group so as to provide them proper and maximal help. The present paper proposes to examine the sociodemographic correlates viz. Age, Sex group. Educational Status, Income Group, Marital Status and also possible causes of drug abuse of these patients. For this purpose patients of Dr. Vidya Sagar Hospital's drug-de-addition cum rehabilitation unit were statistically analysed. The findings have been discussed. PMID- 10538000 TI - Planning and designing of blood bank for a district hospital (200-300 beds). PMID- 10538001 TI - Laboratory services in India. PMID- 10538002 TI - Is the obstetric guideline of 30 minutes from decision to incision for Cesarean delivery clinically significant? AB - A Cesarean delivery may be critical to the health and wellbeing of a newborn. The time required to extract an infant from a hostile in utero environment is a frequent issue in medical negligence cases. The American College of Obstetricians and Gynecologists and the American Academy of Pediatrics suggest a time guideline of 30 minutes from decision for Cesarean delivery to the beginning (incision) of the procedure. This time frame is based on survey data from hospitals throughout the United States and is not based on clinical outcomes or the pathophysiology of obstetric events. This review focuses on specific Cesarean indications as noted by the specialty groups and analyzes them from an outcome point of view. The authors conclude that specific high-risk factors do indeed warrant delivery in as expedient a fashion as possible; however, compliance with the 30-minute guideline does not necessarily lead to a difference in outcome as far as the neonate is concerned. PMID- 10538003 TI - The early diagnosis project: a collaborative approach to risk management. AB - Diagnostic errors account for one-fifth of lawsuits against hospitals and physicians. Virtually all medical specialties are at risk for these lawsuits. To this point, risk managers have done little to assist physicians to reduce their risk of failure-to-diagnose (FTD) lawsuits. This article describes a collaborative effort to reduce FTD lawsuits in the primary care setting. PMID- 10538004 TI - Risk investigations involving genetic identification. AB - Risk managers should consider misidentifications as causes of otherwise unexplained diagnostic and process errors. Genetic tests are powerful tools that can resolve problem cases and indicate ways to improve patient-sample identification. Genetic typing, especially for DNA markers, has provided evidence of patient or sample identity in 21 of 22 hospital and laboratory cases of accidental or intentional patient misidentification, specimen mislabeling, and sample contamination. Identity is established with very high probability if infrequent genetic markers are observed in both unknown and reference specimens. The odds of a match of markers express both the infrequency of finding the match by chance (in specified populations) and the adequacy of testing. Genetic tests establish nonidentity with virtual certainty. PMID- 10538006 TI - Supreme Court issues decisions on sexual harassment. Burlington Industries v. Ellerth. PMID- 10538005 TI - Not to worry: directors and officers claims can be managed. AB - Directors and officers claims have become more prevalent due to the employment litigation surge in this country. This article provides examples of real claims, offers suggestions in managing claims, discusses trends in litigation and provides recommendations for loss prevention. PMID- 10538007 TI - Reports to and from NPDB may be defamatory. Anbar v. Leahan; Swafford v. Memphis Individual Practice Association; Le Baud v. Frische. PMID- 10538008 TI - Testing for HIV without consent may be invasion of privacy. Doe v. High-Tech Institute, Inc. PMID- 10538010 TI - Physician does not have duty to third parties to physician/patient relationship. Van Horn v. Chambers. PMID- 10538009 TI - Claims for negligent credentialing not covered by general liability policies. United States Fidelity and Guaranty Company v. St. Elizabeth Medical Center. PMID- 10538011 TI - Washington Statute of Repose held to be unconstitutional. DeYoung v. Providence Medical Center. PMID- 10538012 TI - Whistleblowers and nuclear medicine. AB - Healthcare facilities that practice nuclear medicine are subject to federal "whistleblower" protection laws when an employee reports a potentially unsafe radiological condition. This article addresses enforcement of the applicable sections of the Atomic Energy Act and the Nuclear Regulatory Commission's regulations in order to help such facilities avoid running afoul of those laws, which can result in fines, generate civil lawsuits by the claimant, and significantly disrupt the operation of a healthcare facility. PMID- 10538013 TI - Dispelling urban myths in healthcare risk management. AB - Applied research to explore and challenge myths in healthcare risk management is pivotal to the growth of the profession. The authors demonstrate this process through exploring patient safety and malpractice issues on weekdays compared with on weekends and holidays. Analysis suggests that claim volume is driven by service volume. PMID- 10538014 TI - High reliability perinatal units: an approach to the prevention of patient injury and medical malpractice claims. AB - Perinatal units differ in their ability to prevent patient injury and medical malpractice litigation. Obstetrical units with favorable performance are distinguished by common organizational and clinical features. Organizationally, they resemble what behavioral scientists define as "high-reliability organizations" (i.e., the ability to operate technologically complex systems essentially without error over long periods). Clinically, practices are based on nationally recognized guidelines and/or an operational philosophy of "safety first." These organizational and clinical features are described so that physicians, nurses, and administrators might view their own clinical environments in the context of this perspective. PMID- 10538015 TI - A case of Munchausen syndrome by proxy presenting as a medical negligence action. AB - The parents of a child who was transported lifeless to a hospital initiated a medical negligence action. Only after discovery did it become clear that the mother, suffering from a disorder known as Munchausen syndrome by proxy, caused the death of the child. The case presented a number of complex evidentiary and litigation issues likely to be encountered by other medical-negligence defense attorneys. PMID- 10538016 TI - Denial of privileges of disabled physician may violate ADA. Menkowitz v. Pottstown Memorial Medical Center. PMID- 10538017 TI - Hospital does not have nondelegable duty to ensure quality of emergency care. Baptist Memorial Hospital System v. Sampson. PMID- 10538018 TI - Kentucky statute does not protect peer review information. Sisters of Charity Health Systems, Inc. v. Raikes. PMID- 10538019 TI - Chiropractor does not have duty to refer to physician or obtain informed consent. Murphy v. Nordhagen. PMID- 10538021 TI - Medical leave of absence could be required as reasonable accommodation. Cehrs v. Northeast Ohio Alzheimer's Research Center. PMID- 10538020 TI - Disclosure of substance abuse problem may be required to obtain patient's consent. Cleveland v. Albany Urology Clinic, P.C. PMID- 10538023 TI - The joy of soy. PMID- 10538022 TI - Mental health reform: what it would really take. PMID- 10538024 TI - Diets for life. PMID- 10538025 TI - Turbocharge your taste. PMID- 10538026 TI - Never too old. Sexually active seniors are one of the fastest-growing HIV infected populations in the U.S. PMID- 10538027 TI - Kids and surgery. PMID- 10538028 TI - Contracting for high-tech health care for patients at home: a survey of purchaser responses. AB - Points out that the Department of Health's Executive Letter: EL(95)5 moved the finance of high technology treatment provided at home for chronically ill patients from the NHS prescribing budget onto a defined and consistent framework. The aim was to obtain better value for money by encouraging competition between potential homecare providers. Reports on a survey of prescribing advisers of purchasing health authorities, which focused on their response to these developments, and discusses the issues identified by purchasers in their implementation of EL(95)5. Notes that, although most purchasers chose to contract directly with a single commercial homecare organization in 1995-1996, there was no consensus about where contracts should be placed in the future, and that the purchasers identified inefficiencies in contracting for such care. Discusses methods of improving the purchasers' response to contracting. PMID- 10538030 TI - Barriers to complaints: a survey of mental health service users. AB - Reports on a survey of 222 mental health service users in two health service trusts, which provides evidence to support and elaborate Wood's analysis of barriers to effective complaints procedures. Identifies key confounding factors such as: lack of awareness of the existence of procedures, the fears of service users about making a complaint, and the lack of awareness of rights and expectations of services. Notes key implications for managers including: the provision of accurate, comprehensive information to service users about complaints procedures, the need to recognize the many factors which inhibit service users from using procedures and the need to inform service users about their rights and services. PMID- 10538029 TI - Health-care professionals and management development. AB - Discusses the impact of a self-governing hospital trust's accredited management development programme designed for health-care professionals responsible for managing natural clinical groups. The programme was a dual qualification: a level 5 national vocational qualification in management, and a diploma in management. Identifies key issues resulting from this type of programme. Discusses participants' evaluation of the two different formats for management development. Highlights the reservations of health-care professionals in respect of competence based management development, particularly regarding assessment of their work performance. Recognizes that when a group of senior health-care professionals are involved in a long-term in-house management development programme, they may be perceived as a threat by senior management. Concludes that health-care professionals will only engage proactively with management development activities which they perceive to have value for them. PMID- 10538031 TI - Decisions, decisions. AB - Describes changes in the organization of the NHS towards a trust-based system and explains that these changes have devolved decision making downwards. Points to the importance of strategic planning for the survival of hospital trusts and notes that strategy should be developed by doctors and managers working together. Outlines management concepts and theoretical constructs underpinning strategic decision making, applying them to the NHS. Concludes that the professionalization of NHS management has shifted power from doctors towards generalist managers but that there is evidence suggesting the value of clinicians as decision makers, given the right skills and an understanding of factors affecting decision processes. PMID- 10538032 TI - Incorporating psychometric measures in selecting and developing surgeons. AB - Provides a review of the recent literature on the selection and development of surgical trainees and surgeons, and discusses findings on the validity of psychometric measures of ability and personality in such processes. Drawing on this body of research, outlines the pilot project recently completed by the authors at St George's Hospital in London, which sought to test the appropriateness of incorporating psychometrics in supporting the career development of junior doctors applying for surgical training. The results of this pilot study tend to confirm those provided by similar research on surgeons, as well as other clinical groups--that psychometric measures have a significant role to play in aiding the assessment and career development of doctors. PMID- 10538033 TI - Management of queues in out-patient departments: the use of computer simulation. AB - Notes that patients attending public outpatient departments in Hong Kong spend a long time waiting for a short consultation, that clinics are congested and that both staff and patients are dissatisfied. Points out that experimentation of management changes in a busy clinical environment can be both expensive and difficult. Demonstrates computerized simulation modelling as a potential tool for clarifying processes occurring within such systems, improving clinic operation by suggesting possible answers to problems identified and evaluating the solutions, without interfering with the clinic routine. Adds that solutions can be implemented after they had proved to be successful on the model. Demonstrates some ways in which managers in health care facilities can benefit from the use of computerized simulation modelling. Specifically, shows the effect of changing the duration of consultation and the effect of the application of an appointment system on patients' waiting time. PMID- 10538034 TI - Health for All and British health policy: a comment on the quest for "healthy public policy". AB - Explains that Health for All is an international extra-governmental movement that seeks to pursue equity in access to health-related resources by broadening the scope of health policy. Notes that its major principles include social participation in state decision making, inter-sectoral collaboration in policy formulation and the improvement of conditions for the disadvantaged. Points out that its local initiatives often encompass health-service professionals and practitioners as well as the voluntary sector, social services and other local authority departments, and that the effect of this local activity on political understandings of health at a national level gives some indication of the extent to which this local time and effort have been justified. In this respect, notes two limits to the impact of the Health for All movement on the political debates about health in Britain. Suggests that these centre on a largely indifferent but powerful national government and an emphasis within the movement initiatives at the level of a politically marginalized local state. PMID- 10538035 TI - Be creative in your approach to healthcare IT staffing. PMID- 10538036 TI - Kids under the weather: a rainbow of care for sick children. PMID- 10538037 TI - Resource management and scheduling: managing basic costs. PMID- 10538038 TI - A new look at EDI. Single source solution leads to 60 percent decrease in cost per claim. PMID- 10538039 TI - Broken promises ... or wasted efficiencies? PMID- 10538040 TI - Data mining, distributed networks, and the laboratory. PMID- 10538041 TI - Track trends in staffing enterprise-wide. PMID- 10538042 TI - Streamline the registration process with EMPI. PMID- 10538043 TI - Managed care. Beam me up, Scotty. PMID- 10538044 TI - What works. OCR scanning system saves time and money, insures good first impression. PMID- 10538045 TI - What works. Automated ER triage process saves money, speeds patient care. PMID- 10538046 TI - Voice, data, video network offered with 1-step shopping. PMID- 10538047 TI - Hotlist: financial applications and services. PMID- 10538048 TI - Software components: which ones solve the implementation problem? PMID- 10538049 TI - The information system professionals behind the 100 top hospitals. PMID- 10538050 TI - Physicians, hospitals would like HMOs to be more forthcoming when sharing data. PMID- 10538051 TI - Compensation monitor. Health care executives most highly compensated. PMID- 10538052 TI - Single payer: can Uncle Sam make it right? PMID- 10538053 TI - Why physicians will embrace technology. Interview by Patrick Mullen. PMID- 10538054 TI - 'Will prudent layperson please report to the ER'. PMID- 10538055 TI - A better office visit for doctor and patient. PMID- 10538056 TI - Communication. Strengthening interviewing skills. PMID- 10538057 TI - Know what to do when agents search your office for evidence. PMID- 10538058 TI - How much should patients know about what it costs to treat them? PMID- 10538059 TI - Managed care outlook. PBM industry stabilizing amid mergers and spinoffs. PMID- 10538060 TI - Health care reform: past experiences and current status. PMID- 10538061 TI - Dealing with the conflict of interests in health care reform. PMID- 10538062 TI - Health care policy evaluation: a conceptual model using medical ethics. PMID- 10538063 TI - Supplying primary care practitioners: a transformation in state politics and policy. PMID- 10538064 TI - Health care reform: implications for human resources management. PMID- 10538065 TI - National health care reform failure: the political economy perspective. PMID- 10538066 TI - In a different voice: the attitudes of licensed practical nurses towards the future. AB - In the fall of 1997, the Idaho Commission of Nursing and Nursing Education undertook a survey of Idaho nurses to collect data on future workforce and educational needs. The results of the survey demonstrated substantial attitudinal differences between licensed practical nurses and registered nurses in the areas of educational motivation, role differentiation, skills by type of nurse, trends in nursing supply, and educational preparation. Given that the Idaho workforce is comparable to the national nursing workforce in numbers, educational background, and age, these findings have implications for health policy planners and nursing educators throughout the country. This article presents the findings and then uses the data as a basis for suggesting their implications for nursing education and workforce planning. PMID- 10538067 TI - An update on JCAHO: what you need to know to prepare for your next survey. PMID- 10538068 TI - Remembering universal precautions in the ED. PMID- 10538069 TI - Should you switch to point-of-service billing? PMID- 10538070 TI - Use simulation to learn results in advance. PMID- 10538071 TI - HCFA, Sen. Grassley preparing for nursing facility bankruptcies; letter instructs states to have contingency plans ready. PMID- 10538072 TI - Industry to use study showing inadequacy of PPS rates to press case for relief from HCFA, Congress. PMID- 10538073 TI - HCFA to use quality indicators as survey tool under new protocol. PMID- 10538074 TI - PPS rates should be adequate for most SNFs, says former HCFA project officer; industry groups dispute findings. PMID- 10538075 TI - HCFA to target transfers of patients from hospitals-within-hospitals. PMID- 10538076 TI - How to have a successful relationship with your fiscal intermediary. PMID- 10538077 TI - Avoiding the developing storm. PMID- 10538078 TI - By the numbers. Managed care offers quality cardiovascular care. PMID- 10538079 TI - Ombudsman programs. PMID- 10538080 TI - Consumer Advisory Committees. PMID- 10538081 TI - A commitment to clinical research. PMID- 10538082 TI - Disease management technology at the point of care. PMID- 10538084 TI - Rethinking medical decision making. Interview by Mark Hagland. PMID- 10538083 TI - STDs: the silent epidemic. PMID- 10538085 TI - Best practices in women's health: identifying exemplary care. Hormone replacement and mid-life issues. AB - Because health plans are organized systems of care providing comprehensive benefits with coordination and accountability, they can deliver care in a way that the old system could not. Health plans can track patients and patient groups and develop programs to increase the likelihood that patients receive preventive care, that chronic conditions are managed more effectively, and that various treatment approaches are monitored to track outcomes. None of these services were possible when individuals went to doctors and simply sent a bill to their insurance company for reimbursement. All over America, health plans are working to realize the potential of organized systems of care. In recent years, however, these efforts to improve care have been largely eclipsed by debates about the shortcomings--both real and perceived--of an evolving system of managed care. In 1996, The Commonwealth Fund provided a grant to the American Association of Health Plans (AAHP) to identify models of delivering care to women through health plans. The Fund views highlighting exemplary models as important to improving health care for women throughout the managed care industry. This article is part of a series based on that work, and is a follow-up to an article on hormone replacement therapy that appeared in the March/April 1999 issue of Healthplan (p. 65). Gleaned from more than 1,000 health plans across the country, this series showcases exemplary practices and programs in four women's health topics: breast cancer, domestic violence, obstetrics and prenatal care, and hormone replacement therapy and other mid-life issues. PMID- 10538086 TI - Health 2000: the next millennium ... panel discussion. PMID- 10538087 TI - Campaign 2000: will any presidential candidate dare to make health care the central issue? PMID- 10538088 TI - The drug formulary decision-making process. PMID- 10538089 TI - Making a name in managed care. PMID- 10538090 TI - Dietitians in management roles. Meeting the challenges of the future. PMID- 10538091 TI - Healing pressure ulcers. Determining the cost of medical nutrition therapy in long-term care. PMID- 10538092 TI - The laptop: an indispensable tool for consultant dietitians. PMID- 10538093 TI - Building bridges. How to establish a dialogue with difficult employees. PMID- 10538094 TI - Extra! Extra! Free data available on the web. AB - San Francisco-based Market Insights may shake up the industry with its new offering of free benchmarking data on the Web. Increasing competition and greater access to technology are making data much cheaper and easier to get. Data companies are shifting their focus from plain numbers to a more complex analysis. PMID- 10538095 TI - Southern hospitals keep the heat on. AB - Southern hospitals continue four-year reign at top of HCIA/Mercer 100 Top Hospitals list. Fierce managed care competition in some Southern states results in top-notch care. In the West, study found that lengths of stay are actually rising as levels of managed care increase. PMID- 10538096 TI - Top 100 hospitals form clinical research group. PMID- 10538098 TI - Disease management firm's enrollment booms: web-based system empowers patients. AB - Greensboro, NC-based Accordant Health Services, which specializes in managing 14 complex, chronic diseases, saw a 726% enrollment gain in 1998. At the end of 1998, Accordant reported a 53% reduction in hospital utilization among 754 patients with complex, chronic diseases. Company leaders say one key to their success has been the innovative use of technology that lets patients be at the center of their own treatment plan. PMID- 10538097 TI - Is your benchmarking missing the mark? PMID- 10538099 TI - Prepare for scrutiny of your role in Medicare appeals. PMID- 10538100 TI - Here's how to find social support for your frail Medicare members. AB - How to tap community resources for seniors. The HMO Workgroup on Care Management has issued a report on how plans and providers can access and work with community resources to meet the social needs of frail seniors. Find out what the panel suggests, and learn from the experiences of an Oregon medical group and a Georgia HMO. PMID- 10538101 TI - Seniors relying on Medicare as sole health care insurance may turn to Medicare+Choice. AB - Data File: Seniors most likely to join Medicare+Choice plans are those with no other source of medical insurance who want the most benefits they can get at the lowest out-of-pocket cost. A study by the Employee Benefit Research Institute finds the numbers of seniors in this category are increasing, especially among the poor, women, minorities, and those over age 85. PMID- 10538102 TI - Include smoking cessation programs in prenatal care. AB - Plan turns to providers to get pregnant Medicaid smokers to quit. After finding it tough to get pregnant Medicaid smokers to kick the habit on a plan level, a Wisconsin health cooperative relies on providers to persuade their pregnant Medicaid patients to stop smoking. See why some plans hope providers can make a difference. PMID- 10538103 TI - Marketplace. How health plans are tapping into the potential of minority populations. PMID- 10538104 TI - Perspectives. Drug benefits: a balancing act for Medicare as well as private HMOs. PMID- 10538105 TI - Marketplace. Wall Street: health plans will prosper, squeezing employers and providers. PMID- 10538106 TI - Perspectives. Salaried doctors turn to unions to stem losses of income, autonomy. PMID- 10538107 TI - Vulgarians at the gate. How ego, greed, and envy turned MedPartners from a hot stock into a Wall Street fiasco. PMID- 10538108 TI - Why CEOs fail. PMID- 10538109 TI - Live a lot longer. PMID- 10538110 TI - The HMO you love to hate is winning over Wall Street. PMID- 10538111 TI - Seven approaches to branding: take your pick. PMID- 10538113 TI - Hillestad: retailing offers opportunities to hospitals and health care delivery systems. PMID- 10538112 TI - Health care's top 10 trends that should ride into next decade. PMID- 10538114 TI - Retail enterprises bring fast, low-risk cash to health systems seeking new revenues. PMID- 10538115 TI - Some on 'Fastest Fifty' list of fast-growing hospitals say they don't belong there. PMID- 10538116 TI - Louisville's Jewish Hospital comfortable as leader on list of fast-growing hospitals. PMID- 10538117 TI - The 21st century health care leader. PMID- 10538118 TI - Should systems at risk turn to disease management? PMID- 10538119 TI - Patient service center concept promises enhanced experiences for patients. PMID- 10538120 TI - Reinventing health care: marketing consultant offers cures for what ails today's system. PMID- 10538121 TI - Jefferson Home Care sees OASIS not as a burden, but a tool to improve performance. PMID- 10538122 TI - Rumors of the death of integrated systems are greatly exaggerated. PMID- 10538123 TI - Bad news abounds for hospitals' bottom lines. PMID- 10538124 TI - Possible solutions to the bottom line blues. PMID- 10538125 TI - Price wars are over; go for higher MCO rates. PMID- 10538126 TI - Medicare claims processing instructions for pancreas transplantation issued. PMID- 10538127 TI - Office design can contribute to productivity. PMID- 10538128 TI - Baby boomers unprepared for long-term care costs. PMID- 10538129 TI - Quality indicators shift survey focus. PMID- 10538131 TI - Tough year for the top 50. PMID- 10538130 TI - The personal touch. Interview by Lynn Wagner and Kathleen Vickery. PMID- 10538132 TI - Cross training not just for athletes. PMID- 10538133 TI - Device failure poses Y2K threat. PMID- 10538134 TI - Looking good, feeling good. Quality cosmetology services can contribute to residents' care. PMID- 10538135 TI - Providers face the computer revolution. PMID- 10538136 TI - Finding their way to health. Health systems provide school-based comfort zones for at-risk teens. PMID- 10538137 TI - An incentive for community health. Linking CEO compensation to community goals. PMID- 10538138 TI - Trustees: commit to quality--now. PMID- 10538139 TI - Dick Carroll: redefining risk management. Interview by Laurie Larson. PMID- 10538140 TI - An eye to the future. PMID- 10538141 TI - Measuring up. Benchmarking tools can enhance executive performance. PMID- 10538142 TI - The perils of motherhood. Even in America, having babies remains dangerous for many. PMID- 10538143 TI - Making history with the humble fruit fly. Bug genes reveal roots of human behavior. PMID- 10538144 TI - Georgia Public Health Central Laboratory Decatur, Georgia. Facilities for training and for the sophisticated handling of biological specimens drive the design of this public health lab. PMID- 10538145 TI - Health care, Chicago style. PMID- 10538146 TI - An automated system for nonroutine staining. PMID- 10538147 TI - An anaerobic microbiology system for the clinical laboratory. PMID- 10538148 TI - Clearer communication in acid-base disorders: [H+] not pH. PMID- 10538149 TI - The mechanism and clinical applications of the NMP22 tumor marker immunoassay: a review. PMID- 10538150 TI - Noninvasive diagnostics: predicting flap viability with near-IR spectroscopy and imaging. PMID- 10538151 TI - Considerations in microscope design to avoid cumulative trauma disorder in clinical laboratory applications. PMID- 10538152 TI - Tracing the textures of an ethical relation to the dead: trauma, witnessing, and the "Montreal massacre". AB - This essay offers a consideration of English-language feminist memorial discourse as this has been sedimenting in Canada since the 1989 murder of 14 women at Ecole Polytechnique. The author suggests that remembrance now, almost a decade after the murders, exceeds the terms offered by a politic in which the living and the dead are connected through feminist alignment ["it could have been me"]. In its place, the author argues that there is a binding relation to the dead that is forged through understanding the murders as an event of historical trauma and rupture. She then contemplates and explores the implications of this rethinking of an ethics of relation through a situated analysis of annual memorial vigils. PMID- 10538153 TI - Practical discourse as policy making: an application of Habermas's discourse ethics within a community mental health setting. AB - This paper describes the application of Habermas's communication ethics in resolving an ethical dilemma that arose in the daily operation of a women's hostel. By generating the conditions of practical discourse, several obstacles to an open, non-hierarchical discussion among staff were removed. The process aided the clarification of differing views within an atmosphere of mutual respect. The discourse resulted in a modification of usual practice that might not have been achieved if an unexamined interpretation of standard policy had been applied. Some difficulties and limitations of this adaptation are also described. PMID- 10538154 TI - [Ethics and mental health: Are the most vulnerable always sacrificed? "The more things change, the more they remain the same!"]. PMID- 10538155 TI - Cultural interpretation services within a multicultural context: an exploration of the problematic and ethical issues facing social service institutions. AB - This paper explores the challenges for social service agencies which offer cultural interpretation services in their bid to meet the needs of service seekers and recipients from linguistic minorities. The author argues that cultural interpretation is provided by institutions that have done little more than add a service for clients from cultural minorities, while leaving intact their service structures--structures that have historically viewed language and "cultural differences" as problems. This orientation will need to change if these services are to be accessible and equitable for Canadians from linguistic and ethnic minorities. While cultural interpreters remain critical to service delivery, they need to work within institutions where service providers and administrators understand language as a cultural, social, and political instrument through which individuals articulate their identities, realities, and understandings of their cultural contexts and service needs. This paper concludes by identifying some of the ethical dilemmas and questions that attend the needed institutional changes. PMID- 10538156 TI - Communities, collectivities, and the ethics of research. AB - Ethical concerns have always placed limits on research, but the spirit of the times has led to an expansion of the territory covered by ethics. This new approach is found in the code of ethics presently in final revision by the three major granting agencies. This code will give unprecedented power to "collectivities" in the research process. Some see this as a long overdue corrective to hit-and-run research, while others see it as a threat to unfettered inquiry. This paper argues a different point: Involvement of collectivities is essential for ethical research relationships, but it ought not to limit the sorts of questions we study or publication of the answers we find. The difference will be illustrated by two examples in which aboriginal communities asserted their collective rights against researchers. The difference between the examples will lead to an examination of the debate between those who believe community work is "all politics," and those who try to underpin it with ethical principles. Finally, I argue that ethical practice requires a knowledge base created by valid research. We should support an improved relationship with host communities, but not let the political agendas of contending community groups constrain the questions we can ask about social problems or our assessment of measures designed to solve them. PMID- 10538157 TI - [Epidemiological research on violence against children: Ethical issues]. AB - This article reports on a seminar held among a group of researchers, practitioners, lawyers, service managers, and members of an ethics committee. This seminar examined various important ethical questions raised by an epidemiological (population) research project. The project's objective was to establish the incidence of violence against children in the province of Quebec. The ethics committee of the organization responsible for health and well-being surveys in Quebec (Sante-Quebec) had rejected the project on the grounds that several ethical constraints jeopardized the methodological requirements of the survey. The main issues discussed by the members of the seminar concerned: (a) balancing individual and collective rights, (b) identifying the status and the role of the researchers, (c) gauging the protection of individual children against the protection of research participants, (d) identifying the risks for children who could be linked to systematic reports of child maltreatment-like situations, and (e) dealing with the ambiguous notion of "reasonable doubt" as it presents itself in a survey context. Participants identified various paths for possible solutions to the problems which tackled legal, methodological, and pedagogical areas. PMID- 10538158 TI - Indoor environment quality--a question-and-answer session. PMID- 10538159 TI - Continuous quality improvement in health care organisations. AB - Quality Management has become a major concern in the delivery of health care. The demand of community, economics of medical practice especially due to technological advances, increasing legal action in malpractice cases and concern to protect the interest of clientele have focused the attention on quality management programmes for all hospital administrators, since the quality of health care has infinite number of elements. It is difficult to assess them all, yet we can demonstrate improvements in performed and elements of care, by putting each of these elements through measurable criteria. Quality assessment measures must cover the efficacy, efficiency, cost and outcome of health care technology. An elaborate valuation system should consist of detailed study of structure, process and outcome. This will establish a balanced equation between the performance and the resources. It is necessary to establish sound dependable methods like total quality management/continuous Quality Improvement in healthcare institutions. It is particularly important that quality care be ingrained as philosophy in Hospitals. PMID- 10538160 TI - Organisation of emergency services. PMID- 10538161 TI - Planning and designing of hospital laboratory services for a district hospital (200-300 beds). AB - The hospital Laboratory is a valuable investment and an economic asset to the health services. The average length of hospital stay is reduced by a rapid return of Laboratory results which facilitates early diagnosis, permits a more rapid patient turnover and accurately controls treatment. PMID- 10538162 TI - Correlation of CT SCAN with clinical provisional diagnosis--a NIMS study. PMID- 10538163 TI - A study of visitors in inpatient area of a teaching hospital beyond visiting hours. PMID- 10538164 TI - Absenteeism among group-D employers. AB - A study of absenteeism and man days lost in one hundred employees belonging to group "D" in a hospital at Jammu was conducted to explore the magnitude of problem of absenteeism and its remedy was sought. The study included the study of staffing pattern of the hospital, total group "D" employees, their jobs and work load and their attitudes, commencing from Sept. 1992 to Aug. 1995. Information regarding man days lost and man days schedule to work as records available in the hospital was used to calculate the trend, variations and fluctuations in absenteeism. For the purpose of calculations of absenteeism, nature of absence was divided into various headings like casual leave, sick leave, earned leave etc. A questionnaire was prepared and given to each employee and a detailed insight was sought with respect to his personality, social factors like accommodation, religious ceremonies, individual habits like drinking, gambling, job dissatisfaction and finally from the whole data absence rate, frequency rate and severity rate were calculated. From the observations it was concluded that absenteeism was more in females, in the age group of 33-45 years, married, in the months of Sept. to Nov. on Mondays and Saturdays and in Hindu backward classes. Though it seemed unreliable but all the employee opined that they were in harmony with their bosses and seemed satisfied with their jobs. PMID- 10538165 TI - Patient satisfaction survey of accident and emergency services of a tertiary care teaching institute of a metropolitan city. PMID- 10538167 TI - Proceedings of National Symposium on Medical Waste Management held at J.L. Auditorium, AIIMS, New Delhi on 27th & 28th Sept. 1996. PMID- 10538166 TI - Hospital management training in the North Eastern Region. AB - The article highlights the lack of training facilities for hospital management in the entire North Eastern Region, explores the quantum of need of the training in the region by identifying hospitals, other health institutions and manpower in the region which require, for their qualitative and quantitative improvement, the expertise in hospital management, and recommends suitable courses of hospital management to start with. PMID- 10538168 TI - Medical waste management--an overview. PMID- 10538169 TI - The medical care services at cross-roads. AB - Our hospitals of all types and sizes present a dismal picture and have miserably failed to meet the rising demands of its consumers, in terms of technology, quality, quantity and cost of care. The situation prevailing has been documented in various press reports and cases registered to the consumer forums in the country. The author while presenting the maladies involving the hospitals has made an attempt to analyse the cause and effect relationship of the maladies and suggested remedial measures. More emphasis has been laid on economic use of resources, cost containment, training and motivation of all categories of staff, and introduction of quality management techniques to improve functional efficiency for better output, in terms of quality, quantity, cost of care and consumer satisfaction. PMID- 10538170 TI - Medical waste management--issues and perspectives. PMID- 10538171 TI - Issues involved in hospital waste management--an experience from a large teaching institution. AB - Experience from a large teaching hospital regarding hospital waste management is presented. The quantum of hospital waste generated is 0.775 kg. per patient per day. Out of this biomedical waste constitute only 6.27 per cent. Though all resources have been provided, a large implementation gap for waste management is seen because of attitudinal problem. A particular difficulty is experienced for disposal of green coconut shells. Incineration has been advocated as a viable method of disposal. Cost of incineration is Rs. 2.71 per patient per day. PMID- 10538172 TI - Organisation of admission clinics in teaching hospitals--a continued study and comparative analysis. PMID- 10538173 TI - Equipment planning for DNA fingerprinting laboratory. PMID- 10538174 TI - Appraisal of equipment used in physical medicine and rehabilitation. AB - There is a wide variety of equipment, used in the practice of Physical Medicine and Rehabilitation (PMR). These equipment may be classified as Diagnostic and Therapeutic. In this paper, the basic essential equipment to run a PMR department in a community/district level hospital are presented. The basic equipment are discussed vis-a-vis benefits, obtained from the modern high-tech equipment by way of cost-benefit studies. It was found that some equipment are desirable at a teaching/referral hospital level, but most of the equipment simply add to the glamour rather than serving any useful purposes when compared to the exorbitant cost. Most of the equipment e.g. LASER, IFT, some diathermies. Combnation units do not show any appreciable therapeutic benefits when compared to placebos. The question arises, whether these equipment are essential, when accepting such equipment, do we have any authentic studies showing their significant benefits or we are simply playing puppets in the hands of manufacturers and vendors. In conclusion, barring a few basic essentials and some precise evaluation equipment, most of the equipment these days are marketed in the name of therapeutic PMR equipment have a questionable cost to benefit profile as compared to therapy given manually by a therapist with (in some cases 0 a little help of the so called out dated conventional equipment e.g. exercise table, weights & pulleys, hot packs etc. The Dictum: "A tablet of aspirin to kill pain is always beneficial to any electrotherapy or thermo therapy," should always be kept in mind while prescribing a therapeutic modality to the patient or ordering an equipment for the PMR department. PMID- 10538176 TI - Changing scenario in management of laboratory services. PMID- 10538175 TI - Costing of a cardiac catheterisation procedure. AB - An attempt has been made to identify the main factors involved in the costing of a Cardiac Catheterisation Procedure (CCP). The technique of historical costing was used. The cost of a CCP worked out to Rs. 4265.21. The direct costs contributed 62%, the indirect costs 2.6% and expenses contributed 35% to the total cost. All these factors were found to be sensitive to the volume and duration of CCP, hence administrators must assist the staff of cardiac catheterisation laboratory (CCL) in controlling these two factors for the cost effectiveness and efficient utilisation of the CCL. PMID- 10538177 TI - Modern concepts of architecture and functional design in operation theatres. PMID- 10538178 TI - Drug utilization in indoor ANC patients of Govt Medical College Hospital, Nagpur. AB - Drug utilization in the indoor patients of ANC ward of Govt. Medical College Hospital, Nagpur was studied in 42 patients. The prescriptions of these patients were audited to find number of drugs per prescription; prescribing trends and category-wise drug consumption. In most of the prescriptions drugs were prescribed by generic names (68.53%), Dosage form was mentioned, Frequency given, but duration was not mentioned. Dose in recommended units was not mentioned in 69.93% of prescriptions. Even though prescription of drugs was found to be rational, prescription writing was far from desired. The enquiry reveals these findings. PMID- 10538179 TI - Role of good communication in patient satisfaction. PMID- 10538180 TI - Fire safety in health care facilities. AB - All healthcare facilities must have a plan for the protection of all persons on their premises and for their evacuation from the building in case of fire. Written copies of this plan must be available to all supervisory personnel. All employees must periodically trained and informed of their duties in implementing the plan. All beds must be easily movable should evacuation be necessary. Emphasis should be placed on moving patients who are in he room of fire origin and others who are directly exposed to the fire, and on maintaing in their rooms the patients who are not immediately threatened during the fire. Fire exist drills must include actual transmission of a fire alarm signal alongwith a simulation of a fire alarm signal alongwith a simulation of a fire alarm signal alongwith a simulation of a fire condition. Quarterly drills on all shifts are required. All personnel including administrative staff, maintenance personnel and internal must be trained, as well as the nurses on duty each shift. A minimum of 12 drills must behold each year. All employees must be instructed in life safety procedures and use of devices. PMID- 10538181 TI - Proceedings of National Workshop on Curriculum Development for MD degree in hospital administration, organised by the Department of Hospital Administration, Kasturba Medical College, Manipal, on 9th January to 11th January 1997. PMID- 10538182 TI - Curbing trade of human organs--status of current policy regarding registration of hospitals. PMID- 10538183 TI - Gender differences in the historical trauma response among the Lakota. AB - The historical trauma response is a constellation of characteristics associated with massive cumulative group trauma across generations, similar to those found among Jewish Holocaust survivors and descendants. Trauma response features include elevated mortality rates and health problems emanating from heart disease, hypertension, alcohol abuse, and suicidal behavior. This article explores gender differences in the historical trauma response among the Lakota (Teton Sioux) and the correlation with health and mental health statistics. The theory of a Lakota historical trauma response is first explained. Traditional gender roles are described in combination with modifications engendered by traumatic Lakota history. Then, data from a study on Lakota historical trauma are presented, including gender differences in response to an experimental intervention aimed at facilitating a trauma resolution process. The data revealed significant gender differences. The sample of women presented initially with a greater degree of conscious affective experience of historical trauma. In contrast, the men reported more lifespan trauma associated with boarding school attendance and appeared to be at an earlier stage of grief. However, at the end of the intervention, women's experience of survivor guilt--a significant trauma response feature-decreased while men's consciousness of historical trauma and unresolved grief increased. Degree of traditional presentation-of-self, including phenotype, appeared to interact with gender to place male participants at greater risk for being traumatized over the lifespan and perhaps subsequently utilizing more rigid defenses against the conscious experience of the trauma with the exception of survivor guilt. The article concludes with a discussion of health and mental health implications for prevention and treatment of the trauma response which could positively impact the health status of the Lakota. Recommendations for future research are suggested. PMID- 10538184 TI - At what cost? The social impact of American Indian gaming. AB - American Indian gaming has been called the "new buffalo." It has the potential to greatly influence cultural traditions on American Indian reservations. This study looks at the social impact that American Indian gaming is having on one reservation in northern Minnesota. Tribal members share strong feelings, both positive and negative, about the issue. Concerns about gaming include an increase in gambling abuse and addiction; a lack of appropriate child care; and concern that gaming is replacing traditional social activities. Some express concern that American Indian values are being replaced by materialism. Supporters of gaming point out that gaming provides tribal members with an opportunity to learn job skills and have gainful employment. Implications for social policy are given. PMID- 10538186 TI - Whose genes are they? The Human Genome Diversity Project. AB - The Human Genome Diversity Project (HGDP) has targeted several hundred indigenous peoples worldwide as their source of genetic material. Proponents for this project claim that information derived by analyzing these materials may be used for a variety of purposes ranging from finding a cure for diabetes to resolving debates about human origins. However, the HGDP plan raises many issues for indigenous people. This paper describes the project as well as the possible ethical and policy implications for Native communities. PMID- 10538185 TI - Protecting the future of indigenous children and nations: an examination of the Indian Child Welfare Act. AB - The Indian Child Welfare Act is a landmark piece of legislation, passed in response to a long history of Native American children being alienated from their families and communities. The Act has far reaching implications for social workers and human service professionals who have any involvement with Native American children or families. Still, many professionals are either unaware of the Act all together or do not know how to effectively implement its provisions in their practice. This lack of awareness and other factors such as inadequate funding have meant that the Act has never realized its full potential to reduce the number of children in out-of-home care. In order to increase awareness about the Act and to make its implementation in day to day social services more practical, this article provides background information on the factors leading to the Act, information on the law itself, and recommendations for practitioners, administrators, and students in the human services. PMID- 10538187 TI - Interactions between American Indian ethnicity and health care. AB - Interventions in health care must be sensitive to the part that culture plays in treatment, recovery and healing of the American Indian patient. Cultural factors play an important part in how the family participates and copes with the intervention program. Interpreting communication and behavior from the perspective of the family's culture contributes to positive family-professional interaction. This paper addresses the most important cultural factors impinging on positive health care for American Indian families and addresses a process for assessment of cultural conflicts which may prevent positive outcomes in the delivery of health care to this population. In addition, this paper offers strategies throughout that can be used by health care professionals to assure culturally sensitive service delivery to American Indians. PMID- 10538188 TI - The changing spots of Alabama's public health. PMID- 10538189 TI - Cross-subsidization in hospital care: some lessons from the law and economics of regulation. PMID- 10538190 TI - The maladaptation of Miranda to advance directives: a critique of the implementation of the Patient Self-Determination Act. PMID- 10538191 TI - Who is my mother?: why states should ban posthumous reproduction by women. PMID- 10538192 TI - The new proposed safe harbors for certain managed care plans and risk-sharing arrangements: a history, analysis, and comparison with existing safe harbors and federal regulations. PMID- 10538193 TI - Canaries in the coal mine: the chronically ill in managed care. PMID- 10538194 TI - Lehigh Valley Hospital's scorecard program. PMID- 10538195 TI - A sample job charter--patient services manager. PMID- 10538196 TI - Optimal staffing for hospitals: in search of solutions. AB - The best staffing models are those that give nurses and other caregivers control and flexibility in their work, say administrators who are fighting higher acuity, tighter budgets and shorter patient stays as well as the threat of staff burnout. Specialty float pools, well-trained students and carefully defined job charters all can ease the strain, staffing experts say. PMID- 10538197 TI - The confidentiality countdown. PMID- 10538199 TI - The new wave of Gen X workers. PMID- 10538198 TI - Where the smoking ain't easy. PMID- 10538200 TI - The new direction in disability management. Putting it all together. PMID- 10538201 TI - Substance abusers. Terminate or treat? PMID- 10538202 TI - Harnessing the power of diversity. PMID- 10538203 TI - A case for the corner drug store. PMID- 10538204 TI - DataWatch. Where hospitals are behind the times. PMID- 10538205 TI - A sharper image of bias: three major disability decisions put stricter limits on who can sue employers for discrimination in hiring. PMID- 10538207 TI - Irreconcilable differences: why the doctor-patient relationship is disintegrating at the hands of health maintenance organizations and Wall Street. PMID- 10538206 TI - What is SAMe? S-Adenosylmethionine. PMID- 10538208 TI - Will group practices meet the Y2K deadline? PMID- 10538209 TI - Who will pay for provider ID? PMID- 10538210 TI - Starting over. Faced with an obsolete computer system, a Michigan payer decides to shut down operations and install a new one. PMID- 10538211 TI - Is third time the charm? A Dayton network tries a new intranet approach as a way to win physician support. PMID- 10538212 TI - This CEO is a firm believer. Interview by MargaretAnn Cross. PMID- 10538213 TI - Reaching out to rural providers. A tertiary care hospital uses information technology to lure new partners. PMID- 10538214 TI - Ringing out the century with a whimper. How the top health care software vendors are trying to make the best of a market besieged by worries. PMID- 10538215 TI - Health care software stocks bugged by Y2K. PMID- 10538216 TI - The Internet offers entrepreneurial opportunities. PMID- 10538217 TI - New software contracts share the risks, rewards. PMID- 10538218 TI - This is only a test.... are the risks involved in being a software test site worth taking in light of the potential rewards? PMID- 10538219 TI - CFOs turn to software to get the job done. PMID- 10538220 TI - At Kelsey-Seybold Clinic, unity in diversity. AB - Kelsey-Seybold Clinic (KSC), the Houston metro market's dominant physician organization, finds itself at a crossroads these days. As a tightly knit multispecialty group with strong physician leadership and significant opportunities for expansion in a rapidly-expanding healthcare market, KSC faces multiple challenges related to health plan consolidation and the ins and outs of capitation. Theirs is a situation epigrammatic of the challenges facing highly evolved medical organizations nationwide. PMID- 10538221 TI - Time for physicians to reconfigure. AB - The days when medical professionals made unilateral patient-care decisions are gone. Accelerating trends are converging to create a climate for what we call "consumer-centric healthcare," and that raises new and unsettling questions for physicians. PMID- 10538222 TI - An alternative view on PCP practice acquisition. New Towers Perrin study contradicts conventional wisdom on practice acquisition losses. PMID- 10538223 TI - Southern California hospitals find health plans less than hospitable. PMID- 10538224 TI - Care manager to the rescue. The key to successful 'gatekeeper' relationships. PMID- 10538225 TI - Expanding the market. Spend-down for assisted living seems to be a growing trend. PMID- 10538226 TI - Security blanket. Long-term care insurance is catching on--and could shape the future of long-term care. PMID- 10538227 TI - People who make a difference. Interview by Wendy L. Bonifazi. AB - In last year's 20th anniversary issue, Contemporary started an annual tradition of honoring people whose innovative business practices, research, advocacy, and other efforts have shaped long term care. This year's four honorees have created groundbreaking tools for care, campaigned to promote the healthy growth of a fledgling form of care, and championed a vision for improving quality of life for residents. PMID- 10538228 TI - Build it and they will come? Making sure your Web site is on the Internet's radar. PMID- 10538229 TI - Acquisition anxiety. The 1998 market was strong, but turbulence lies ahead. PMID- 10538230 TI - Matters of the heart. Life-saving drugs for cardiac disease are still underused. PMID- 10538231 TI - Minimizing the risk of suits for improper care. PMID- 10538232 TI - The oldest of the old. Interview by Yvonne Parsons. PMID- 10538233 TI - Winning the war on waits, delays requires 'continuous flow' systems. AB - The health care system continually battles the problem of waits and delays--but there are success stories. Waits and delays have a negative impact on all involved in a health care system. "Continuous flow" systems emphasize "doing today's work today." To a large extent, delays are built into the way health care delivery systems operate. Reducing waits and delays takes a willingness to change, the ability to communicate, and the spirit of cooperation. PMID- 10538234 TI - Asthma care specialists improve outcomes. AB - The incidence of asthma in the United States is widespread and growing, consuming a great deal of health care resources. A number of studies have concluded that treatment of asthma patients by specialists achieves superior outcomes and saves costs. Better use of aggressive techniques by allergists is central to their success in treating asthma patients. PMID- 10538235 TI - Association creates new standards for anesthesia. AB - Several recent high-profile tragedies have heightened public and practitioner concerns over the safety and quality of office-based anesthesia procedures. Inadequate office-based anesthesia procedures and precautions expose what some estimate to be 8 million people a year to unnecessary risk. The Accreditation Association for Ambulatory Health Care has established new quality standards for office-based, or mobile or "itinerant," anesthesia organizations. PMID- 10538236 TI - 'No secrets' approach pays off in satisfaction. AB - Health care organizations can encourage the free flow of information by making internal communications a high-priority, strategic function. A "no secrets" policy is a prime indicator of organizations that are doing an exemplary job of internal communications. Baptist Hospital in Pensacola, FL, provides a demonstration of best practices in internal communications within a health care organization. PMID- 10538238 TI - Supply and demand for radiation therapists: a United States perspective for 1997 2002. AB - The purpose of this paper is to review the current and future employment plans of radiation therapists and administrators that employ them so that the supply and demand for these individuals is quantified. The survey was funded by the American Society of Radiologic Technologists and implemented by the Task Force on Human Resources. In April 1997 a questionnaire was mailed to a sample of radiation therapy administrators identified by the mailing list of the Society of Radiation Oncology Administrators (SROA). Another questionnaire was mailed to a sample of radiation therapists holding active certificates with the American Registry of Radiologic Technologists. Results of the survey indicated an increased demand for radiation therapists based on plans for facility expansion. Individuals anticipating retirement in the next five years ranged from 7% of the radiation therapists and 12% of the administrators surveyed. A model was developed which projected a shortfall of therapists ranging from 0% (-13 positions) in 1998 to 3% (-403 positions) projected for 2002. The paper concludes that the supply of radiation therapists did not meet the demand in the time period measured; 1998 2002. An increase in newly educated radiation therapists is needed to fill the openings created by those leaving the field. The survey should be repeated at successive intervals to accurately track and project both the supply of and the demand for radiation therapists. PMID- 10538237 TI - The development of a community breast center. AB - Maximum capacity for mammography has been reached at Kaweah Delta Health Care District Imaging Center. Increased need and support in the community for access to care was a driving force in development of a Breast Center. What follows is a case study of implementing a Breast Center in a semi-rural community in Central California, which involves a collaborative effort between Cancer Program and Imaging Services. PMID- 10538239 TI - The key to excellence: successful executives keep the flame alive at work and home! PMID- 10538240 TI - The brave new world of healthcare e-commerce. PMID- 10538241 TI - Want to improve your facility's business? Try making your staff happier. PMID- 10538242 TI - Community project is low on costs, high on results. PMID- 10538243 TI - There are still a few unready hospitals. AHA survey reveals potentially alarming situation. PMID- 10538244 TI - Senate Y2K panel finds health care lagging. PMID- 10538245 TI - Flu vaccine programs get a shot in the arm. PMID- 10538246 TI - Patient privacy legislation moving forward. Senate working toward compromise measure. PMID- 10538247 TI - Medicare delivers a nasty outpatient surprise. BBA issues dominate legislative and regulatory agendas. PMID- 10538248 TI - DeParle delivers HCFA update. PMID- 10538249 TI - Armey, Thomas, Daschle share views on health care legislation. BBA hit private hospitals hardest, Congressional leaders say. PMID- 10538250 TI - Workshops focus on EMTALA, qui tam and union organizing. PMID- 10538251 TI - Chances for Medicare reform remain slight. Senate Finance Committee set to kick off debate. PMID- 10538252 TI - Post-mortem on the Tulane-Columbia partnership. Industry bellwether or one-time bonanza? PMID- 10538253 TI - Integrating behavioral health and primary care pays off. Model promises significant savings. PMID- 10538254 TI - Newly recognized PACE (Programs of All-Inclusive Care for the Elderly) programs benefit patients, providers, payers. PMID- 10538255 TI - Pharmaceutical manufacturers likely to be challenged by regulatory agencies over assertions of deceptive advertising. PMID- 10538256 TI - Pennsylvania hospitals find joint venture is a better solution than a merger. Health systems gain access to markets and services. PMID- 10538257 TI - Screening admission CT scans in patients with AIDS--a randomized trial. AB - OBJECTIVE: To determine if the length of hospital stay could be reduced for patients with AIDS by performing screening head and abdominal-pelvic computed tomography (CT) scans within 24 hours of admission, regardless of presenting signs and symptoms. DESIGN: Randomized, prospective trial. SETTING: Tertiary, academic medical center. PATIENTS: On presentation to the emergency department, 42 patients with AIDS were identified as being eligible to participate in our study. Twenty-two patients consented to participate and were assigned to screening CT or control group. INTERVENTION: Patients assigned to the screening CT group had head and abdominal-pelvic CT scans within 24 hours of admission, regardless of presenting signs or symptoms. The findings of the screening CT scans were immediately communicated to the patient's referring physician. Patients assigned to the control group had CT studies done solely at the discretion of their physician. MAIN OUTCOME MEASURE: Length of stay for patients in the screening CT and control groups. RESULTS: The average length of stay for patients in the screening CT group was 1.3 days longer than the average length of stay for patients in the control group (95% CI, 1.4 days shorter to 4 days longer). The study was terminated after 22 patients were enrolled. CONCLUSION: Screening CT scans of the head and abdomen and pelvis at the time of hospital admission do not reduce the length of stay for patients with AIDS. PMID- 10538258 TI - Institutional volumes and coronary angioplasty outcomes before and after the introduction of stenting. AB - CONTEXT: An increasing number of patients undergoing percutaneous transluminal coronary angioplasty (PTCA) are receiving coronary stents. OBJECTIVES: To assess whether the introduction of coronary stenting has changed hospital mortality or same-admission coronary artery bypass grafting (CABG) and whether the hospital's procedure volume affects these outcomes. DESIGN: Observational study using hospital claims. SETTING: Nonfederal hospitals that performed PTCA in California in 1993 and 1996. PATIENTS: 35,350 patients who underwent PTCA in 1993 (before the introduction of stenting) and 43,040 patients who had PTCA in 1996 (43% of whom received stents). MEASUREMENTS: Hospital stenting volumes for 1996 were divided into terciles; total PTCA procedures per year were categorized as low (< or = 200), medium (201 to 400), or high (> 400). Outcome variables included hospital death and coronary artery bypass grafting (CABG) performed during the same admission. Patients with a principal diagnosis of acute myocardial infarction (AMI) were analyzed separately from those without such a diagnosis. RESULTS: From 1993 to 1996, the characteristics of patients undergoing PTCA did not change substantially. The use of same-admission CABG decreased by 13% (from 6.0% to 5.2%; P = 0.008) in the AMI group and by 30% (from 3.7% to 2.6%; P < 0.001) in the no-AMI group. Hospital mortality did not change significantly in either group. Procedure volume was not related to hospital mortality. However, rates of same-admission CABG were significantly lower at hospitals with high annual stenting volumes than at low-volume centers (1.3% vs. 2.3% among patients in the no-AMI group; P < 0.001). CONCLUSIONS: Hospital mortality rates after PTCA have not changed considerably since the introduction and diffusion of coronary stenting. However, rates of same-admission CABG have decreased in recent years and are lowest at hospitals with high procedure volumes. PMID- 10538259 TI - Direct admission to an extended-care facility from the emergency department. AB - BACKGROUND: Many patients are admitted to acute-care hospitals when their medical needs might be more appropriately met in an extended-care facility (ECF). OBJECTIVE: To describe a cohort of patients who were admitted from an emergency department to an ECF. DESIGN: Observational cohort study. PARTICIPANTS: 121 enrollees of Harvard Vanguard Medical Associates who were admitted directly from an emergency department to an ECF between October 1, 1994, and December 31, 1997. OUTCOME MEASURES: Mean length of stay, charges per patient, and discharge disposition (discharged to home, discharged to a long-term-care facility, died, or transferred to an acute-care hospital within 30 days of ECF admission). RESULTS: Patients admitted directly to an ECF were generally frail and elderly (median age, 75 years). Mean length of stay in the ECF was 11 days; the mean per patient charge was $3290. Three quarters of patients were discharged from the ECF to their homes. Six percent (seven patients) were transferred from the ECF to an acute-care hospital within 30 days of ECF admission. None of these transfers clearly suggested that the initial decision to directly admit a patient to the ECF was inappropriate. Most patients were satisfied with direct ECF admission: Of the surviving, cognitively intact patients admitted to an ECF in 1997, 71% stated that they would choose direct admission to an ECF over admission to an acute-care hospital if they were "in a similar situation in the future." CONCLUSIONS: For selected patients, direct admission to an ECF seems to be feasible, safe, and acceptable. A randomized, clinical trial is needed to fully assess the safety and cost implications of direct ECF admission. PMID- 10538260 TI - Screening mammography rates by specialty of the usual care physician. AB - CONTEXT: Although Medicare began paying for screening mammography in 1991, utilization among enrollees has been low. PRACTICE PATTERN EXAMINED: The relation between the specialty of the usual care physician and the proportion of women 65 years of age and older receiving mammography. DATA SOURCE: 100% Medicare Part B claims for 186,526 female enrollees residing in Maine, New Hampshire, and Vermont during 1993 and 1994. RESULTS: Among women of the target screening age (65 to 69 years), 55.4%, received mammography during the 2-year period. The highest rates of mammography were observed in women whose usual care physician was a gynecologist (77.9%; 95% CI, 75.8 to 79.9), followed by those treated by an internist (67.1%; CI, 66.5 to 67.7), family practitioner (58.1%; CI, 57.4 to 58.9), general practitioner (47.4%; CI, 45.4 to 49.5), and other specialists (41.3%; CI, 40.1 to 42.5). The lowest rates were observed in women who had no physician visits during the 2-year period (9.5%; CI, 8.7 to 10.4). Although screening rates were lower in women aged 70 years and older, a similar pattern was observed. CONCLUSIONS: The probability of a Medicare enrollee's receiving screening mammography is strongly influenced by the specialty of her usual care physician. Covering a preventive service does not guarantee its use. PMID- 10538261 TI - Hospitalist staffing requirements. AB - CONTEXT: The use of hospitalists--physicians who spend a substantial portion of their time providing in-hospital care to the patients of primary care physicians- has been proposed as a way to decrease costs and increase the quality of inpatient care. COUNT: Number of full-time hospitalists. CALCULATIONS: Average daily census = annual admissions x length of stay divided by 365. Number of hospitalists = (average daily census divided by patients per hospitalist) + 1 extra hospitalist for night coverage. DATA SOURCES: The average number of patients per hospitalist was obtained from a National Association of Inpatient Physicians membership survey. A low estimate of 10 patients per hospitalist was used to account for the extra manpower needed for coverage during vacations and other time off. RESULTS: A hospital with 3000 admissions per year and an average length of stay of 5 days would have an average daily census of 41 patients and would need 5 full-time hospitalists. Hospitals with a lower patient volume would need fewer hospitalists and would probably need to find persons other than hospitalists to cover some nights and weekends. CONCLUSIONS: Simple calculations based on hospital admissions and length of stay can estimate the number of hospitalists required for adequate staffing. Requirements will vary with the hospitalists' workload; the patient case complexity; and the duties other than inpatient care that are required of hospitalists, such as consultations, skilled nursing facility coverage, quality improvement work, teaching, and research. PMID- 10538262 TI - Preparing manuscripts for submission to medical journals: the paper trail. AB - CONTEXT: Preparing a manuscript for publication in a medical journal is hard work. OBJECTIVE: To make it easier to prepare a readable manuscript. APPROACH: Start early--A substantial portion of the manuscript can be written before the project is completed. Even though you will revise it later, starting early will help document the methods and guide the analysis. Focus on high-visibility components--Pay attention to what readers are most likely to look at: the title, abstract, tables, and figures. Strive to develop a set of tables and figures that convey not only the major results but also the basic methods. Develop a systematic approach to the body of the paper--A standard framework can make it easier to write the introduction, methods, results, and discussion. An obvious organization with frequent subheadings and consistent labels makes the paper easier to read. Finish strong--Improve the paper by sharing it with others and by learning how to elicit and receive their feedback. Take the time to incorporate useful feedback by revising frequently. PMID- 10538263 TI - What is a hospital? PMID- 10538264 TI - A shared statement of ethical principles for those who shape and give health care: a working draft. The Tavistock Group. PMID- 10538265 TI - Efficient prenatal care: fewer visits, fewer sonograms. PMID- 10538266 TI - Specialty practices consolidate: new litigation emerges. Boman v. Southeast Medical Services Group. AB - This case is instructive in that it highlights a trend to use the Federal health care statutes in furtherance of a broad array of lawsuits. While the statutes were intended principally to protect the Medicare and Medicaid programs, the statutes are increasingly used to remedy a plethora of conduct. Here, the case is also helpful in that it does not dismiss or downplay the use of the Anti-Kickback Statute for such purpose. Rather, the court simply stated that the practice did not violate the Anti-Kickback Statute. PMID- 10538267 TI - Why the capping of AIDS benefits by self-funded employer welfare benefit plans should be actionable under Section 510 of ERISA. PMID- 10538268 TI - OIG Advisory Opinion 99-6 waiving co-payments. PMID- 10538269 TI - Gainsharing: a concept whose time has come? PMID- 10538270 TI - Summary of the new proposed regulations for ERISA claims procedures. PMID- 10538271 TI - Lawyers beware of criminal health care fraud: what attorneys can learn from the Kansas City health care attorney indictments. PMID- 10538272 TI - State action, due process, and regulated health care: the downstream effects of American Manufacturers Mutual Insurance Company v. Sullivan. PMID- 10538273 TI - When "other nonprofits" convert. PMID- 10538274 TI - On responsibility--Part II: The board. PMID- 10538275 TI - Promotional products effective in fund-raising campaigns. PMID- 10538276 TI - Big dogs make a difference. PMID- 10538277 TI - Are you destined to burn out? PMID- 10538278 TI - Toward 2000 and beyond: charitable and social change giving in the new millennium, Part I. AB - Commissioned by Craver, Mathews, Smith & Company (CMS), this study takes an in depth look at donors to charitable and progressive social change organizations at the close of the 20th century. As a follow-up to the benchmark donor study sponsored by CMS in 1990, the current research allows for some basic comparisons between donors at the beginning and end of the decade. PMID- 10538279 TI - The discipline of innovation. PMID- 10538280 TI - Appealing to the donors too much? AB - Many fund raisers believe that almost all donors feel they are receiving too many appeals. However, that is not always the case. Donors' perceptions of appeals vary widely from one organization to the next, and are also influenced by outside factors, such as age and overall non-profit mail received. PMID- 10538281 TI - The spotlight is on you--are you ready? PMID- 10538282 TI - Special report: new paradigms in credentialing. Here's how you should straddle the compliance-credentialing crosswalk. PMID- 10538283 TI - Special report: new paradigms in credentialing. Walking the new walk. PMID- 10538284 TI - Conduct a cost-effective RCA (root-cause analysis) by brainstorming. PMID- 10538285 TI - Clear up roles, expert says, don't 'mush them' together. PMID- 10538286 TI - Software 'scoreboard' used for competitor comparison. PMID- 10538287 TI - Involve governing board in patient safety effort. PMID- 10538288 TI - Pharmaceutical expenditure in Sweden. AB - Recently, the responsibility for prescribed pharmaceuticals in Sweden was transferred from national level to the regional health authorities (county councils). The purpose was that a closer integration and balance between pharmaceuticals and other factors of production in health care should produce better opportunities for a cost-effective use of the total health care resources. The purpose of this paper is to present a deeper analysis of pharmaceuticals as a production factor in Sweden, mainly during the 1990s, and to discuss the future development and future policy decisions in Sweden. Pharmaceuticals have increased their share of total health care expenditure in Sweden, from about 9% in 1990 to about 14% in 1995. The Swedish pharmaceutical market can be divided into sub markets, where the prescription sub-market accounts for the greater part of pharmaceutical expenditure. Further, a few disease categories account for a larger fraction of the cost of prescribed pharmaceuticals. The importance of pharmaceuticals as a production factor also differs between different age groups. Several factors are expected to contribute to a future increase in Swedish pharmaceutical expenditure, for instance an ageing population and the rapid introduction of expensive new pharmaceuticals. PMID- 10538289 TI - National health interview surveys in Europe: an overview. AB - In order to study the value of national health interview surveys for national and international research and policy activities, this paper examines the existence and content of recent and future health interview surveys in the 15 member states of the European Union (EU), Norway, Iceland and Switzerland. National health interview surveys are performed in most countries, but not in Greece (only regional surveys), Luxembourg, Ireland and Iceland (only multi-purpose surveys). The health interview surveys in the other 14 countries provide regular data on the main health topics. Of the 14 health topics that are examined in this inventory seven are measured in all countries. Questions on health status (e.g. self-assessed health, long-term physical disability, and height and weight) and medical consumption (e.g. consultations with the general practitioner, GP) are often included. Lifestyle topics are less often included, except smoking habits, information about which is sought in all countries. Topics like diet and drugs/narcotics are more often included in special surveys than in general health interview surveys. Despite differences in the content, frequency and methodology of national health interview surveys in different countries, these surveys are a valuable source of information on the health of Europeans. PMID- 10538290 TI - Regional disparities in health care supply in eleven European countries: does politics matter? AB - There are large differences both among and within European countries in the supply of health care facilities and personnel. In 1979 Smith posed the hypothesis that spatial disparities in health care supply will be smaller in countries with socialist (or social-democratic) governments. The aim of this paper is to examine this hypothesis by analysing whether or not regional disparities in health care supply within countries are smaller in countries that have been governed predominantly by socialist governments. We have collected regional data on the number of hospital beds and the number of physicians for 211 regions in 11 European countries for 1970 and 1990. Countries were classified according to the political composition of governments in the post-war era. It is concluded that: (1) the amount of regional variation is greater for hospital beds than for doctors; (2) for both aspects of supply, regional disparities decreased over time; (3) the decrease in regional disparities between 1970 and 1990, both for beds and for doctors in hospitals, was stronger for countries that had more years of socialist government in that period and (4) there is no relation between the number of years of socialist government between 1945 and 1990 and regional variation in health care supply in 1970, nor for government participation between 1970 and 1990 and variation in supply in 1990. PMID- 10538291 TI - Economics of screening programs for Tay-Sachs disease. PMID- 10538292 TI - The contingent valuation method in health care. AB - The contingent valuation method (CVM) is a survey-based, hypothetical and direct method to determine monetary valuations of effects of health technologies. This comprehensive review of CVM in the health care literature points at methodological as well as conceptual issues of CVM and on willingness to pay as a measure of benefits compared with other measures used in medical technology assessment. Studies published before 1998 were found by searching computerised databases and former review literature. Studies were included, when performing CVM using original data and meeting qualitative criteria. Theoretical validity of CVM was sufficiently shown and there were several indications of convergent validity. No results on criterion validity and only a few on reliability were found. There was widespread use of different elicitation formats, which make comparisons of studies problematic. Direct questions were seen problematic. First bids used in bidding games influenced the monetary valuation significantly (starting point bias). There were indications that the range of bids of payment cards also affected the valuation (range bias). However, no strategic bias was found. The influence of different states of valuation (ex-ante, ex-post) and of payment methods, as well as the possible aggregation of the results of decomposed scenarios rather than more complex holistic scenarios, were rarely investigated. Further methodological analysis and testing seems to be necessary before CVM may be used in health care decision making. Important research topics are the connection of assessment of different elicitation methods and criterion validity as well as tests on reliability according to methodological issues. Concerning conceptual issues, the analysis of the influence of different states of evaluation and of the status of the respondents as diseased or non-diseased, as well as the aggregation of results of decomposed scenarios, proved to be topics of further research. PMID- 10538293 TI - Mission possible: Texas facility upsizes. PMID- 10538294 TI - Smart and smarter. Intelligent buildings graduate to a new level. PMID- 10538295 TI - Power trip. Electricity deregulation sparks shopping sprees. PMID- 10538296 TI - Lighten up! High energy costs? Upgrade. PMID- 10538297 TI - Pipe line. NFPA '99 weighs piping system risks. PMID- 10538298 TI - Wash daze. High laundry costs spinning out of control? Try these tips. PMID- 10538299 TI - Y2K doesn't scare most U.S. hospitals, new survey shows. PMID- 10538300 TI - Price survey. Short-term deals reflect glove volatility. PMID- 10538301 TI - Group hunts for weak links in members' supply chains. PMID- 10538302 TI - Preventing Pyxis wars: without nurses, inventory control impossible. PMID- 10538303 TI - VHA back in limelight after spinning off purchasing. PMID- 10538304 TI - Was this sound medical judgment--or outright bias? PMID- 10538305 TI - The search for Medicare fraud. Don't let billing mistakes make you a target. PMID- 10538306 TI - The search for Medicare fraud. Will your patients turn into bounty hunters? PMID- 10538307 TI - The search for Medicare fraud. What to do--now--to protect yourself. PMID- 10538308 TI - Great advice for young doctors--from colleagues who learned the hard way. PMID- 10538309 TI - Charity medicine. Is managed care ripping a hole in this safety net? PMID- 10538310 TI - Latest twists in the Medicare melodrama. PMID- 10538311 TI - Was this doctor "unconventional" enough to lose his license? PMID- 10538313 TI - Teaching hospitals facing serious financial losses. PMID- 10538312 TI - Why you need real business pros on your board. PMID- 10538314 TI - OSHA issues controversial advisory on risks of latex. PMID- 10538315 TI - Synthetic gloves: an alternative. PMID- 10538316 TI - Latex allergy assessment raises nagging questions. PMID- 10538317 TI - OR roundtable. Managers' advice on OR staffing. PMID- 10538318 TI - Report card helpful planning resource. PMID- 10538319 TI - Adhesion barriers worth the cost? PMID- 10538320 TI - How to avoid being hijacked by anger. PMID- 10538321 TI - Sterilization & infection control. Which process is best to use? PMID- 10538322 TI - OR benchmarks. Rhinoplasty best performers. PMID- 10538323 TI - New glove standards are emerging. PMID- 10538324 TI - Social anxiety. For millions of Americans, every day is a struggle with debilitating shyness. PMID- 10538325 TI - Drug bazaar: getting medicine off the Web is easy, but dangerous. PMID- 10538326 TI - Outstanding regulatory aspects in the European pharmaceutical market. AB - This paper identifies and analyses a number of outstanding regulatory aspects in the completion of the European pharmaceutical single market. It discusses pricing and competition in pharmaceuticals in the aftermath of the 3 Frankfurt Roundtables and their results. It analyses the environment for generic competition in the European Union (EU) and the extent to which this environment needs to be amended in order for such competition to be promoted. It links the issues of parallel trade, standardisation, single trademark, the European databank and the definition of innovation with the current situation in the functioning of the single market, particularly the sovereignty of the member states in determining pricing and reimbursement levels. It argues that the above problems need to be tackled in conjunction with pricing and reimbursement. The paper further points at new developments, in particular biotechnology patenting and orphan drug regulation, where the EU has introduced or is about to introduce new legislation that has been needed for a long time and examines how this legislation can be beneficial. Finally, the paper analyses the implications for healthcare provision in the member states of 2 legal cases heard before the European Court of Justice in relation to the free movement of goods and healthcare provided across borders. The paper concludes that there is still a long list of regulatory aspects that remain unresolved despite the fact that significant progress has been made to date, and observes that economic analysis in pharmaceutical regulation is very much intertwined with political expediency. In addition, the definition of political expediency varies as one considers developments at the EU or national level, since it does not necessarily follow that individual actors coincide in opinion. PMID- 10538328 TI - Standard cost lists for healthcare in Canada. Issues in validity and inter provincial consolidation. AB - A standard cost list is a listing of recommended costs for a selected group of services. Standard costs are used in economic evaluation studies to eliminate that proportion of cost differences between interventions that are due to cost differences between providers. In this article we provide a summary of cost lists for pharmaceutical economic evaluation purposes which have been developed in 2 provinces in Canada-Alberta and Manitoba. We then assess these 2 lists from 2 different viewpoints. First, we developed criteria for the internal and external validity of costs and, in light of these validity criteria, we assessed how the 2 standard cost lists compared with the 'ideal' measure of long run marginal costs. Second, we identified the criteria for the inter-provincial consolidation of standard cost measures (in order to develop a single, consolidated cost list); in light of these criteria, we assessed whether the degree to which the 2 separate lists could be consolidated. The lists achieved a considerable degree of external validity, but fared less well in terms of internal validity. However, these results depend on the 'ideal' measure of cost which is used. The lists, in the forms which were developed, are not easily consolidated into a single list. Further refined cost data would be needed in order to achieve consolidation. PMID- 10538327 TI - Angiotensin converting enzyme (ACE) inhibitors and heart failure. The consequences of underprescribing. AB - Heart failure (HF) is a common and expensive cardiovascular disease, in economic terms as well as in lives lost. Angiotensin converting enzyme (ACE) inhibitors have been shown to significantly reduce mortality and hospitalisation in HF. However, recent surveys show that the prescription rate of ACE inhibitors for HF is far below what is considered to be optimal. Furthermore, prescribed dosages are usually lower than those recommended based on evidence from clinical trials. This article estimates the consequences, both economic and human, of underprescribing ACE inhibitors in patients with HF. The indication for prescribing an ACE inhibitor varies, and clinical trials have included different categories of patients; it is inappropriate to assess costs in all eligible patients without taking these factors into account. Therefore, we analysed the data with respect to 4 different groups: (i) asymptomatic left ventricular systolic dysfunction (LVSD)--an early stage leading to chronic HF; (ii) chronic HF; and post-myocardial infarction (MI) LVSD differentiated into (iii) post-MI asymptomatic LVSD and (iv) post-MI chronic HF. We also estimated the cost effectiveness of adding an ACE inhibitor to the treatment of patients with HF for whom an ACE inhibitor is not currently prescribed. If only patient populations in which large trials have shown a significant effect of ACE inhibition on mortality are included in the analysis (i.e. excluding asymptomatic patients with LVSD), increasing the number of Swedish patients receiving an ACE inhibitor could save in excess of 3700 lives each year, in addition to reducing the annual number of hospitalisations by 8400. The additional cost would be 101.5 million Swedish kronor (SEK), a cost per life saved of SEK27 200. Chronic HF is the most cost effective patient population to treat, generating cost savings under certain assumptions. A further 6700 hospitalisations can be avoided should the use of ACE inhibitors be extended to asymptomatic patients with LVSD. Increasing dosages to those used in the large clinical trials may generate additional savings in lives and hospitalisations. In conclusion, the use of ACE inhibitors in HF and LVSD has clearly been proven to be cost effective, and compares favourably with the cost effectiveness of treating hypertension or hypercholesterolaemia. At present, however, ACE inhibitors are not optimally utilised. Given the increasingly constrained resources for healthcare, every effort should be made to increase the use of cost-effective treatments, such as ACE inhibitors in chronic HF and post MI LVSD. PMID- 10538329 TI - Hospital selection for unit cost estimates in multicentre economic evaluations. Does the choice of hospitals make a difference? AB - OBJECTIVE: The objectives of this study were (i) to develop a conceptual framework for selecting hospitals for unit cost estimates in national and international multicentre trials and (ii) to test the impact of alternative hospital selection on the cost results. DESIGN AND SETTING: Within the conceptual framework, the following considerations which can be used when selecting a sample of hospitals for unit cost estimates in multicentre trials were identified: the number of hospitals; the sampling method; and the desired level of geographical subanalysis. Results from a recently completed international multicentre trial were used to explore changes in cost results obtained by using alternative methods of selecting and stratifying hospitals for unit cost estimates. PATIENTS AND PARTICIPANTS: The study included 5041 women from 72 hospitals in 6 countries with prelabour rupture of the membranes at term. INTERVENTIONS: The women were randomly assigned to induction of labour with intravenous oxytocin, induction of labour with prostaglandin E2 gel, or expectant management for up to 4 days with labour induced if complications developed. MAIN OUTCOME MEASURES AND RESULTS: Across each of the 4 management strategies of the study, the method of selecting and stratifying hospitals resulted in a 30 to 55% difference between the lowest and highest median unit cost estimates. In some cases, the relative ranking of the least to most expensive strategy varied across methods of hospital selection. The statistical comparisons across strategies found that the method used had a substantial impact on the conclusions of the economic evaluation. CONCLUSIONS: Unit cost information should be collected from as many hospitals as possible. Multivariate hospital cost studies are needed to identify important cost drivers that will assist with hospital selection in the future. PMID- 10538330 TI - A model to compute the medical cost of patients in intensive care. AB - OBJECTIVE: Our objective was to identify, among the information routinely collected on patients in intensive care units (ICUs), data that determine the total cost for a given patient. DESIGN: We developed a model that could help physicians in medical ICUs to estimate the cost of care for their patients when no cost data were available at the individual patient level. SETTING: A Medical ICU. PATIENTS AND PARTICIPANTS: The model was developed using a random sample of 73 patients admitted to the medical ICU in 1996 and 1997, validated by another random sample of 29 patients admitted during the same period. INTERVENTIONS: The actual medical variable cost per patient was computed from data on the total resources used (excluding personnel and fixed costs), collected from the patients' records plus pharmacy, laboratory and blood bank logs. The explanatory variables tested were: length of stay, nursing workload, severity of condition, and procedures recorded by a score [omega (omega)] including 3 components related to the frequency of procedure use. The model was constructed in a stepwise fashion, assuming a linear relation. Equations were tested on the basis of the residual mean square; criteria for inclusion and elimination of variables were the level of its partial regression coefficient and medical criteria. The model was validated by analysis of variance of the regression on a second population of 29 patients using the F-test. MAIN OUTCOME MEASURES AND RESULTS: The median length of stay was 7 days (range: 3 to 22 days). Mortality rate was 25%. Median medical variable cost was 805 Pounds (mean medical variable cost was 1738 Pounds, total cost was 6279 Pounds). The variables selected in the multiple regression model as relevant predictors of medical costs were: procedures recorded only once during the ICU stay irrespective of their reiteration (omega 1), procedures recorded every time they are performed (omega 2), procedures recorded daily in the ICU (omega 3) and the presence or absence of an invasive procedure (Kc). The final equation, calibrated with r2 of 0.826 and p > 0.0001, was: medical cost (Pounds) = 23 omega 1 + 53 omega 2 + 8 omega 3 + 2352Kc + 96. The validation with the other sample of 29 patients compared actual to predicted costs. Analysis of variance of the regression from the model was r2 = 0.596 (p > 0.05). CONCLUSIONS: Our standardised cost model is a possible approach to allow comparison of medical costs within and between ICUs. PMID- 10538331 TI - Costs of diabetes. A methodological analysis of the literature. AB - OBJECTIVE: To review studies on the costs of diabetes and its complications through a scheme designed specifically for assessing the quality of cost-of illness (COI) studies. DESIGN AND SETTING: The methodology of COI studies in diabetes was analysed in order to assess the significance of quantitative results. The scheme adopted 7 items identified as the main points for discussing the methodological choices governing the results. We also used a checklist based on questions related to the 7 items. MAIN OUTCOME MEASURES AND RESULTS: The answers showed that many studies appear not to give technical details, so it is hard to understand the method. Methodological choices varied widely between the studies. This is probably due to the lack of consensus on the methodology of COI studies. Based on the findings of this review, we suggest also some specific points that could help produce more reliable results on the costs of diabetes. CONCLUSIONS: Clearly, a general consensus on COI studies is still remote, making the value of any comparison of results questionable. PMID- 10538332 TI - Cost of schizophrenia to UK Society. An incidence-based cost-of-illness model for the first 5 years following diagnosis. AB - OBJECTIVE: This study estimated the cost to UK society of an annual cohort of newly diagnosed patients with schizophrenia over the first 5 years following diagnosis, using an incidence-based cost-of-illness framework. DESIGN AND SETTING: A discrete event model of the course of schizophrenia was constructed, based on a literature review and interviews among a panel of healthcare professionals (n = 7). Seven discrete disease states were defined within the model. Patients' movements between these disease states enabled 10 disease courses to be identified. In each disease state, the model estimated resource use and corresponding costs borne by the National Health Service (NHS), Local Authorities, the Home Office and society as a result of lost productivity. PATIENTS AND PARTICIPANTS: The model simulated patients' movements between disease states over the first 5 years following diagnosis. Since there are 7500 new cases of schizophrenia per year in the UK, the model was run for 7500 patient simulations. MAIN OUTCOME MEASURES AND RESULTS: The total discounted cost to society attributable to an annual cohort of newly-diagnosed patients with schizophrenia over the first 5 years following diagnosis was estimated at 862 million Pounds (range: 788 million Pounds to 926 million Pounds in sensitivity analysis). The discounted mean 5-year cost was estimated to be approximately 115,000 Pounds (range: 105,000 Pounds to 124,000 Pounds) per patient or approximately 23,000 Pounds (range: 21,000 Pounds to 25,000 Pounds) per patient per year. The NHS accounted for 38% of the total cost, Local Authorities for 12% and the Home Office for 1%. Indirect costs due to lost productivity accounted for 49%. Of the NHS costs, hospital admissions accounted for 69% and hospital visits (outpatient, day ward and day centre attendances) for a further 26%. Drugs (antipsychotics and adjunctive medications) accounted for 2%. CONCLUSIONS: NHS expenditure and lost productivity costs predominated, irrespective of disease course. This indicates that treatments that reduce hospitalisation and potentially enable patients to return to active employment could significantly reduce the societal burden of schizophrenia. PMID- 10538334 TI - A review of quality of life in Alzheimer's disease. Parts 1 and 2: Issues in assessing disease impact and drug effects. PMID- 10538335 TI - Telepathology in neurosurgery. AB - In most tumor cases of neurosurgery, we need to have a rapid section diagnostic of the tumor during the course of surgery. When we have received the results that diagnose the exact dignity of the tumor, we devise the operation strategy for the continuing course of surgery. The tissue sample is sent by taxi to the pathological institute in Heidelberg. It takes approximately 45-60 min until we receive the results by telephone. Many centers are far away from a pathological institute and the patient may need to be re-operated on there. In stereotaxy it is still more important. We need approximately 15-25 specimens during a stereotaxy, so we can be sure we have a sample with some of the tumor tissue present. With direct contact to a pathological institute we could reduce this dramatically. We prepared a methylen blue slide and used a histological microscope with a video camera attached to it and a digitizer interface for digitized pictures. By use of a modem we send the pictures by telephone line to the pathological institute, where they are assessed. We currently have direct contact with a pathologist so they can tell us from which part of the tumor we should send the other histological pictures. The procedure takes about twenty minutes. By using this procedure the distance to the pathological institute is irrelevant. The system costs approximately $10,000, covering the cost mostly of the microscope and the camera (but we already had the microscope) and can be built by anybody with minimal requirements. PMID- 10538336 TI - Efficient linear elastic models of soft tissues for real-time surgery simulation. AB - In this paper, we describe the basic components of a surgery simulator prototype developed at INRIA. We present two physical models which are well suited for surgery simulation. These models are based on linear elasticity theory and finite elements modeling. The former model can deforme large tetrahedral meshes in real time but does not allow any topological changes. On the contrary, the latter biomechanical model can simulate the cutting and tearing of soft tissue but must have a limited number of vertices to run in real-time. We propose a method for combining these two approaches into a hybrid model which may allow real time deformations and cuttings of large enough anatomical structures. PMID- 10538333 TI - Olanzapine. A pharmacoeconomic review of its use in schizophrenia. AB - Olanzapine is an atypical antipsychotic agent which is at least as effective as the conventional agent haloperidol and the atypical agent risperidone. Olanzapine may be superior to haloperidol in some respects, including treatment of negative symptoms. A major advantage of olanzapine over haloperidol is its lower risk of extrapyramidal symptoms. Olanzapine improves quality of life and other aspects of functioning to a greater extent than haloperidol, and improves quality of life to at least the same extent as risperidone. However, olanzapine has a high acquisition cost compared with conventional antipsychotics. Despite this, most pharmacoeconomic analyses indicate that treatment with olanzapine does not significantly increase, and may even decrease, the overall direct treatment costs of schizophrenia, compared with haloperidol. Total direct medical costs calculated from prospective resource utilisation data were lower with olanzapine than with haloperidol by $US388 (1995 values) per patient over 6 weeks and by $US55 per patient per month during 46 weeks extended treatment. In a mixed effects linear model of the same data, total costs over 1 year were $US10,301 (1996 values) per patient lower with olanzapine than haloperidol, and olanzapine was associated with 18.3 more symptom-free days per patient. Compared with risperidone, mean total direct medical costs over 28 weeks were $US493 (1995 values) per patient lower with olanzapine. In a Markov model of 5 years' treatment, olanzapine was associated with more time in a disability-free state than haloperidol at a total cost per patient that was lower by $US1539 (1995 values), 816 Pounds (1995/1996 values), 977 Dutch guilders (NLG; 1995 values) and 2296 Deutschmarks in US, UK, Dutch and German analyses, respectively. In a similar Spanish analysis, the overall total cost was higher with olanzapine, giving an incremental cost effectiveness for olanzapine of 32,516 pesetas (1995 values) per month of disability-free time gained. When risperidone was a comparator, the total cost per patient was $US1875 and NLG202 lower with olanzapine in US and Dutch analyses, respectively. CONCLUSIONS: The high acquisition cost of olanzapine is offset by reductions in other treatment costs in patients with schizophrenia. Compared with haloperidol, the drug improved patient outcome and quality of life, while overall direct treatment costs were generally not increased, or even decreased. Olanzapine has also been reported to decrease overall treatment costs compared with risperidone, but confirmation is required. Olanzapine is a cost-effective alternative to conventional agents for the treatment of moderately to severely ill patients with longstanding schizophrenia. PMID- 10538338 TI - A new hybrid renderer for virtual bronchoscopy. AB - PURPOSE: To improve the diagnosis of pathologic modified airways, a new hybrid visualization system has been developed and tested based on digital image analysis and synthesis of spiral CT as well as the visual simulation of bronchoscopy. METHOD/MATERIALS: 20 patients with pathologic modifications of the airways (tumors, lung transplantation) were examined with Spiral-CT. The shape of the airways and the lung tissue is defined by an automatic volume growing method and a following geometric reconstruction by the computation of geometric primitives. This is the basis of a multidimensional display system which visualizes volumes, surfaces and computation results simultaneously. The enable the intuitive and immersive inspection of the airways a virtual reality system, consisting of two graphic engines, a head mounted display system, data gloves and specialized software was integrated. RESULTS: In 20 cases the extension of the pathologic modification of the airways could be visualized with the virtual bronchoscopy. The user interacts with and manipulates the 3D model of the airways in an intuitive and immersive way. In contrast to previously proposed virtual bronchoscopy systems the described method permits truly interactive navigation, detailed quantitation of anatomic structures and a "see through" the bronchial wall. The system enables a user oriented and fast inspection of the volumetric image data. CONCLUSIONS: To support radiological diagnosis with additional information a virtual bronchoscopy was developed. It enables the immersive and intuitive interaction with 3D Spiral CTs by truly 3D navigation in the airways. The system was tested with 20 Spiral-CTs of bronchial tumors and obstructions and is well suited for the inspection of structures beyond the bronchialtree. PMID- 10538337 TI - Stereo augmented reality in the surgical microscope. AB - We present an augmented reality system that allows surgeons to view features from preoperative radiological images accurately overlaid in stereo in the optical path of a surgical microscope. The purpose of the system is to show the surgeon structures beneath the viewed surface in the correct 3-D position. The technical challenges are registration, tracking, calibration and visualisation. For patient registration, or alignment to preoperative images, we use bone-implanted markers and a dental splint is used for patient tracking. Both microscope and patient are tracked by an optical localiser. Calibration uses an accurately manufactured object with high contrast circular markers which are identified automatically. All ten camera parameters are modelled as a bivariate polynomial function of zoom and focus. The overall system has a theoretical overlay accuracy of better than 1 mm. Implementations of the system have been tested on seven patients. Recent measurements in the operating room conformed to our accuracy predictions. For visualisation the system has been implemented on a graphics workstation to enable high frame rates with a variety of rendering schemes. Several issues of 3-D depth perception remain unsolved, but early results suggest that perception of structures in the correct 3-D position beneath the viewed surface is possible. PMID- 10538339 TI - Visual clues in minimally invasive surgery: use of 2-D versus 6-D enhanced performance of complex minimally invasive complex skills. AB - In the recent past, we used two 2-D videoscopes to obtain both a close detailed view and simultaneously a panoramic view to improve the efficient and safe access for instruments into the microscopic working field by way of the benefits of the panoramic view. This bi-modal visual set of clues allows for (1) insertion of suture, (2) cutting of suture with scissors (3) retraction of tissue, and (4) removal of suture and needle. During these experiences, we observed the benefits accrued to the surgeon by allowing the focusing of his/her attention on the work (technical skills) without diffusing energy to other activities. Similarly, when training surgeons to perform micro-anastomoses, and while working to improve performance in micro-anastomoses, we hypothesize that two or more videoscopic views of the 3-dimensional working space would provided added visual information to the surgeon during the microscopic work. To examine this hypothesis, we have used a non-animate model, in the performance of complex skills in videoscopic surgery. METHODS: Inanimate videoscopic models for suturing and tying (24 studies) were used in this study. The technical skill studied was the sophisticated skill of suturing. The speed and accuracy of Free-Handed suturing and tying was determined in these studies. They were compared using a single 2-D system verses three videoscopic views reconstructing a 3-D effect. RESULTS: In each of these models, the delineation of multiple views allowed greater detailed 3-dimensional information for the surgeon. The sutures were placed faster, more accurately, and with fewer false motions. These data allow us to conclude the use of multiple high-resolution 2-D views will improve accuracy and efficiency in the performance of delicate and precise skills in videoscopic surgery. PMID- 10538340 TI - Evaluation of skill acquisition using a force feedback, virtual reality based surgical trainer. PMID- 10538341 TI - An intelligent, interactive platform for ophthalmic teaching, telemedicine, and telecollaboration: design considerations and prototype construction. AB - The development of technologies permitting processing, compression, and transmission of digital images and image sequences enables powerful methodologies for local and remote medical teleconsultation. We are developing a slit-lamp based ophthalmic augmented reality (image overlay) environment incorporating features to permit real-time, interactive teaching, telemedicine, and telecollaboration. A binocular slit-lamp biomicroscope interfaced to a CCD camera, framegrabber board, and PC permits acquisition and rendering of anterior segment and retinal images. Computer-vision algorithms facilitate robust tracking, registration, and near-video-rate image overlay of previously stored retinal photographic and angiographic images onto the real-time fundus image. Our algorithms facilitate shared control of pointing, drawing, and measuring functions registered with the retinal image video stream and direct audio communication between an examiner (student, generalist) and remote observer (instructor, specialist). Bandwidth and video compression considerations limit the frame rate and latency for video stream transmission. Excellent and acceptable performance are demonstrated in model eyes over a local area network and through a modem connection, respectively. These studies represent the first investigations towards the design and implementation of an intelligent platform for ophthalmic telemedicine and telecollaboration. PMID- 10538342 TI - Interactive navigation and bronchial tube tracking in virtual bronchoscopy. AB - An interactive virtual environment for simulation of bronchoscopy is developed. Medical doctor can safely plan their surgical bronchoscopy using the virtual environment without any invasive diagnosis which may risk the patient's health. The 3D pen input device of the system allows the doctor to navigate and visualize the bronchial tree of the patient naturally and interactively. To navigate the patient's bronchial tree, a vessel tracking process is required. While manual tracking is tedious and labor-intensive, fully automatic tracking may not be reliable. We propose a semi-automatic tracking technique called Intelligent Path Tracker which provides automation and enough user control during the vessel tracking. To support an interactive frame rate, we also introduce a new volume rendering acceleration technique, named as IsoRegion Leaping. The volume rendering is further accelerated by distributed rendering on a TCP/IP-based network of low-cost PCs. With these approaches, a 256 x 256 x 256 volume data of human lung, can be navigated and visualized at a frame rate of over 10 Hz in our virtual bronchoscopy system. PMID- 10538343 TI - Anatomic VisualizeR: realizing the vision of a VR-based learning environment. AB - The University of California, San Diego's Anatomic VisualizeR project has reached another milestone. As the period of DARPA-funded research and development comes to a close, UCSD's VR-based learning environment has matured to the point where curricular implementation is now underway. In this presentation, we will reflect on the process by which lessons in this virtual environment are realized, highlight the results of ongoing 3-D perception studies, and describe examples of how Anatomic VisualizeR is being used in medical school anatomy and high school biology classes. To conclude, we will outline the future of this project which will include full scale curricular implementation, learning outcomes assessment, and dissemination through industrial and academic partnerships. PMID- 10538344 TI - Some virtual reality and telemedicine applications useful for long duration spaceflight from a systems engineering perspective. AB - This paper presents a review of virtual reality, telemedicine, and systems engineering design concepts that relate to the human performance of mission tasks during long duration spaceflight. A Mini-Case Study of the MIR/Progress collision is discussed and serves to highlight some of the key issues. We review some physiologic changes which occur as a result of long duration spaceflight, discuss issues associated with performing required mission tasks, and integrate concepts from telemedicine and virtual reality as possible methods and technologies to alleviate some of the deleterious effects of extended space missions. PMID- 10538345 TI - Presence as an emotional experience. AB - Presence or the sense of "being there" has been discussed in the literature as an essential, defining aspect of Virtual Reality (VR). The VR literature includes definitions rooted in behavioral response, signal detection theory, and philosophy, but has generally ignored the emotional aspects of experience. The purpose of this paper is to reexamine the concept of presence in terms of people's emotional engagement with reality and their environment. Emotions are an essential part of how people experience the world. Any theory of presence must take emotional factors into account. This thesis has implications about how research should be conducted to further our understanding of presence. Validated psychological techniques for assessing emotions by subjective report, behavioral observations, and facial analysis can all be applied to increase our understanding of virtual presence. Further understanding of the interaction between presence and emotional state will improve our understanding of the construct of presence as well as better inform us about how virtual environments can be applied in creating emotional effects or treating emotional disorders. PMID- 10538346 TI - The use of an information system architecture modeling tool in the development of disease management systems. AB - A Disease Management System (DMS) refers to an integrated healthcare delivery system that provides patient centered care throughout the course of the disease independent of delivery site. A fundamental barrier for the development, implementation and monitoring of a DMS is lack of an appreciation by care providers of the complexity of these systems, and what is required for their maintenance. Foremost in the development of these systems is the presence of information systems that attempt to deal with the temporal, spatial and information needs of the DMS. PURPOSE: The Zachman Framework for Information Systems Architecture is used in many industries in the development of information systems. Its choice is based on the recognition of a need for a methodology in the conceptualization and modeling of complex information systems. This paper provides a brief overview of the Zachman Framework and its potential application in DMS development. In particular it will be the focus on the need for "perspective" clarification as the first step in the development of such complex systems. RESULTS: This paper reviews DMS and their potential information needs. The clarification of "perspectives" provides a method toward team building and unification of purpose by decreasing conflict and recognizing the unique contributions that each perspective holder makes. PMID- 10538347 TI - Gaze patterns in laparoscopic surgery. AB - By understanding surgeons' patterns of gaze, and what visual information is being obtained during a procedure, one can improve the operation via new techniques or instrumentation. Part of a larger project on Remote Manipulation in Endoscopic Surgery, we analyzed eye patterns of surgeons from videotape annotation. Three categories of eye patterns were defined: 1) eyes on (gaze on monitor); 2) eyes down (gaze on external operative space); 3) eyes off (gaze away from monitor/hands). In the context of hierarchical decomposition of procedures we compared eye patterns and sequential dependencies (gaze as a function of previous gaze) by procedure, surgical steps and tasks. Timelines showed transitions in eye patterns during the procedure. We determined what visual information is available and what visual information is needed by the surgeons. By comparing these, we suggest technology that can provide these needs. PMID- 10538348 TI - Virtual endoscopy using surface rendering and perspective volume rendering. AB - Noninvasive virtual endoscopy is a new method of diagnosis using computer processing of 3-D image data sets (such as CT or MRI scans). Conventionally, two methods have been used in virtual endoscopy. One is 3-D surface rendering method. 3-D surface of human organs can be explored in real time by using this method, but surface rendering algorithm has disadvantages such as the low quality of visualized image and the loss of the volume data. The other is perspective volume rendering method. The power of perspective volume rendering is that the intrinsic 3-D richness of the volume data is preserved. However, there are the difficulty in planning flight-path and the disability of real time flight for a computationally intense procedure. This paper presents virtual endoscopy using both surface and perspective volume rendering. By combining each merits of two methods, a user can not only process virtual endoscopy interactively in real time but also view lossless and high-resolution image using the flight path defined by surface rendering. As a result, the use of common fly-paths removes the burden to define a flight-path in conventional perspective rendering method. Also 3-D object is explored in real time and viewed as lossless and detail cine sequences. PMID- 10538349 TI - Teleimmersion for the doctor's office. AB - The purpose of our research is the development of a new technology, teleimmersion, which will make possible 3D visual communication between people distributed over the globe. We are putting together a low-end, PC-based, affordable hardware system, and are developing the software for a teleimmersion node (T-node). The software is based on accurate polynocular stereo. We will describe a testbed for the system, show results, and discuss challenges for future research and implementation. PMID- 10538350 TI - Telepresence surgery system enhances medical student surgery training. AB - The telepresence surgery system (TeSS) permits the surgeon to operate on a patient across distances. This is achieved through real-time 3D video vision, stereo audio, and remote instrument control with haptic feedback. Telepresence surgery has been proposed to be useful in providing specialist operative consultation to remote areas. Remotely mentoring medical students with no surgical experience through complex procedures provides an even greater challenge. Third-year medical students with no prior operative experience were mentored exclusively through use of TeSS during a standard surgical skills lab. This two-day laboratory includes abdominal procedures and thoracic procedures. The medical students were alone in the operating room and the teaching surgeon was in an entirely separate room. Anatomy, surgical principles, and adjunct techniques were taught to the students. The students felt the experience was better than standard because of the enhanced learning secondary to the required verbal accuracy in describing the procedures. In addition, they felt they had better visibility since the instructor was not standing in the way. The telepresence surgery system can be successfully used to remotely mentor and enhance introductory surgical training for inexperienced medical students. PMID- 10538352 TI - Virtual reality on the web: the potentials of different methodologies and visualization techniques for scientific research and medical education. AB - Academic and medical imaging are increasingly using computer based 3D reconstruction and/or visualization. Three-dimensional interactive models play a major role in areas such as preclinical medical education, clinical visualization and medical research. While 3D is comparably easy to do on a high end workstations, distribution and use of interactive 3D graphics necessitate the use of personal computers and the web. Several new techniques have been demonstrated providing interactive 3D via a web browser thereby allowing a limited version of VR to be experienced by a larger majority of students, medical practitioners and researchers. These techniques include QuickTimeVR2 (QTVR), VRML2, QuickDraw3D, OpenGL and Java3D. In order to test the usability of the different techniques, Mednet have initiated a number of projects designed to evaluate the potentials of 3D techniques for scientific reporting, clinical visualization and medical education. These include datasets created by manual tracing followed by triangulation, smoothing and 3D visualization, MRI or high-resolution laserscanning. Preliminary results indicate that both VRML and QTVR fulfills most of the requirements of web based, interactive 3D visualization, whereas QuickDraw3D is too limited. Presently, the JAVA 3D has not yet reached a level where in depth testing is possible. The use of high-resolution laserscanning is an important addition to 3D digitization. PMID- 10538351 TI - New software applications for interchangeable instrumentation in spinal stereotaxis. AB - Computer image-guided surgery has been widely accepted because it allows the surgeon to track an instrument through unvisualized critical structures of a patient in real-time, thus minimizing the risk of injury. Current spinal and cranial image-guided surgery is, however, limited by the lack of surgical instruments and software applications that would allow rapid interchange of useful instruments to perform the procedures. Most image-guided systems utilize a single standard probe or a few pre-defined instruments that are not necessarily useful for performing the actual surgical procedure. Present image-guided technology for screw placement in spinal surgery utilizes the standard probe only to confirm the entry point location and view the planned trajectory of the screw. The surgeon then resumes the procedure using standard surgical instruments to drill, tap and place screws without the benefit of image guidance. Our clinical laboratory experience with spinal image-guided surgery indicates that there is potential for error between each of these procedural steps of screw placement. Despite accurately locating an entry point, any deviation in the trajectory during drilling of a pilot hole, tapping or screw placement may result in significant errors in screw placement and potential neurovascular injury. We have developed custom software applications and universal hardware adaptation devices for spinal image-guided surgery that allow the use of standard instruments for intraoperative guidance. Utilizing universal dynamic registration hardware and software, standard surgical instruments are adapted for real-time image guided surgery. An array of light emitting diodes can be attached to essentially any rigid instrument with a definable tip and then calibrated to the system for intraoperative use. Laboratory tests using a cadaveric model indicate a difference in accuracy of less than 1.0 mm between the standard probe and a dynamically registered custom instrument and an absolute mean error of less than 2.0 mm for the image-guided system which is clinically insignificant in most cases. This technology is a significant step forward as it allows the surgeon to use a full array of instruments with image guidance and will ultimately make spinal and intracranial surgery safer and more accurate. PMID- 10538353 TI - Planning of skull base surgery in the virtual workbench: clinical experiences. AB - Based on the KRDL Virtual Workbench, we present a neurosurgical planning system called VIVIAN (Virtual Intracranial Visualization And Navigation). This VR environment allows for fast and intuitive interaction with three-dimensional multimodal (MRI, MRA, MRV, CT) patient specific data-sets. The user reaches behind a mirror into a 3D workspace where the 3D data is surrounded by interactive virtual tools-racks. Tumors, blood vessels, cranial nerves and surgically relevant parts of the brain are segmented by interactive control of density transfer-functions or by manual outlining and tracing tools. A neurosurgical procedure is planned by using various visualization and measurement tools and the system allows for the simulation of bone drilling and tissue removal. We have planned 16 cases which required tumor surgery at the cranial base. VIVIAN provided an efficient and comprehensive way to understand pre operatively the complexity of anatomical and pathological relationships. The ideal craniotomy and the extent of the required skull base exposure could be specified accurately. PMID- 10538354 TI - Knowledge optimization theory and application to point-of-care testing. AB - Point-of-care testing is defined as testing at or near the site of patient care wherever that medical care is needed. The number and types of tests available at the point of care are increasing dramatically. Point-of-care testing provides test results to the clinical team immediately, usually within a therapeutic turnaround time of five minutes. Immediacy of data is a primary benefit, but we must evaluate when, why, and where point-of-care testing should be used because in most cases this new modality is more expensive than conventional testing in clinical laboratories. Knowledge optimization integrates key components of testing, patient focusing, performance, and test clusters, in concert with synthesis of temporal and diagnostic-therapeutic process information that the physician can assimilate quickly and act on to improve patient outcomes. We identify through literature searches, meta-analysis, and documented outcomes results, the optimal applications of point-of-care testing in critical care and other medical settings. The basis for optimality is improved medical and/or economic outcomes. Results show that point-of-care testing is strongly justified for emergency resuscitations under any circumstances, in critical care, such as the operating room and intensive care unit, and in disease management. Long-range implications are: (1) the point-of-care testing trend will accelerate; (2) viable optimization tools include integrative strategies, clinical algorithms, care paths, and performance maps; (3) attention should be focused on rapid communication and understanding of critical results; and (4) enhanced human interfaces are crucial for efficient medical decision making and efficacious patient therapy. PMID- 10538355 TI - "KnowWare: virtual reality maps for blind people". AB - Much has been done to give blind people access to text-based information through computers. Unfortunately, the recent development of Windows operating systems and GUIs jeopardize these gains. Spatial information which is becoming more important to the sighted has never been widely available to blind people. Through a combination of gesture input with audio output, the KnowWare system offers blind users a new way to access spatial information. An invisible virtual map is defined on the desktop. When the user touches a feature on the map with the tip of his index finger, the computer tells him what he has touched by means of computer-generated speech or synthesized sound codes. Touching is detected by means of an overhead video camera that looks down at the user's hands as they rest upon the desktop. Specialized processors analyze the image of the user's hands and locate the fingertips. Because KnowWare is an electronic medium, it is possible to instantly zoom in on a particular state or country to examine it in more detail. Similarly, panning and overlays are possible. Twenty blind subjects have tested KnowWare and have been able to use it as well as they do traditional raised-line maps. PMID- 10538356 TI - Dynamic volume texture mapping and model deformation for visually realistic surgical simulation. AB - For computer assisted surgical simulation to be effective, objects in the simulated environment should respond to the user's actions dynamically with correct visual information. This includes dragging and cutting that cause changes in geometry, topology and appearance. Geometric object representation can be manipulated intuitively in real-time but does not preserve interior information. Volumetric data representation, on the other hand, preserves volume content but direct manipulation is compute-intensive. 3-D texture mapping provides an alternative in representing volumetric information. We present a surgical simulation system based on geometric models that allows interactive deformation and incision of objects while displaying correct volumetric information corresponding to these changes. This is accomplished by dynamic 3-D texture mapping. This method can be applied to anatomical data and patient CT and MR images to facilitate data/patient specific surgical simulations. PMID- 10538357 TI - Universal interfacing system for interactive technologies in telemedicine, disabilities, rehabilitation, and education. AB - A modular hardware and software system for human-computer interaction is described that allows for flexible, affordable interfacing of people, computers, and instruments. The approach is illustrated with an application in the disabilities area. Other application areas are outlined. PMID- 10538358 TI - Deformation simulation algorithms of elastic tissues in "real-time" based in elasticity theory. AB - In this paper, a new deformable model based in Boundary Elements Method (BEM) is presented. This model is characterised by its robustness and high deformation calculation speed. Experiments developed show that this model could calculate elastic deformations of elastic objects composed by 150 nodes with a 15 Hz. refresh rate (minim refresh rate acceptable in real time interactive systems). PMID- 10538359 TI - Virtual hand laboratory meets endoscopic surgery. AB - We describe two recent research projects: the Virtual Hand Laboratory, and Remote Manipulation in Endoscopic Surgery. The Virtual Hand Laboratory (VHL) is a prototype experimental tool for investigating human visuomotor coordination for object manipulation in augmented and virtual environments. The Remote Manipulation in Endoscopic Surgery (RMES) project examined surgeon's use of viewing and manipulating technologies in laparoscopic surgery, both in clinical and experimental settings. Current research brings together these two parallel research projects (VHL and RMES), for applications in planning and real-time execution of surgical procedures as well as for surgical training. We outline our research directions and detail current activities on superposition of display space on the workspace for the surgeon's hands. PMID- 10538360 TI - Interactive visualization of 3D fields and images on inexpensive workstations using VRML. AB - The visualization of volumetric medical images and static or time-dependent vector fields is performed on personal computers over the Web using an interactive 3D interface based on VRML and Java. The VRML client obtains field information and the surface models of examined objects from a server accessible over the Internet. Various virtual tools enable radiologists and referring physicians to visualize and manipulate complex data sets using a simple interface on low-cost computers. PMID- 10538361 TI - Virtual arthroscopy training: do the "virtual skills" developed match the real skills required? AB - The purpose of the study is to validate the training efficacy of the PC-based Sheffield Knee Arthroscopy Training System (SKATS, as described in MMVR6). Based on a task analysis of real arthroscopy, an evaluation module has been designed to test the core psycho-motor skills used in arthroscopy. The evaluation simulates a joint inspection and triangulation task, which is used to assess the research hypothesis that experienced arthroscopists will perform significantly better on the virtual arthroscopy simulator, than a trainee arthroscopist group and a control group. A group of experienced arthroscopic knee surgeons, a trainee surgeon group and a control group were tested on the simulator. The preliminary results indicate that experienced surgeons performed best with fewer instrument collisions and faster task-completion times. The results indicate that the core skills of arthroscopy used on the SKATS simulator are similar to those used in real arthroscopy. Further validation work is required to assess the training transfer effects. Once the simulator has been validated fully, it may prove beneficial in minimising patient risk. PMID- 10538362 TI - BRAVO/TeleTrend: a comprehensive WWW-based neuromonitoring system for the neurosurgery ICU. AB - This paper describes BRAVO/TeleTrend--a comprehensive client/server-based system for remote access, review and analyses of continuously acquired multiparametric physiological data from Intensive Care unit (ICU) patients. The system is designed as a distributed three tier model and implemented in Java (Sun Microsystems). TeleTrend is a data review package, which interfaces to existing physiological bedside monitors such as the BRAVO suite of products (Nicolet Biomedical, Madison, WI) and the vital signs monitors compatible with the Unity Network (Marquette Electronics, Milwaukee, WI). It does not transfer over the web the entire patient record, which can be hundreds of megabytes. Instead, it provides tools to view a compressed representation of the raw data in a trend display and to zoom into the raw data if needed. Thus, it eliminates the need for a high-bandwidth Internet connection and makes possible the use of a slower modem access to the vast amount of physiological data acquired per patient. In addition, TeleTrend features a rule-based module capable of generating clinical alerts, which is a potentially useful tool for neurointensivists and other critical care personnel. Finally, TeleTrend is intended as a multi-user, semi real-time telemedical application, which features built-in white-board and chat components. These components allow several physicians at different locations around the world to simultaneously view and brainstorm over critical chunks of continuously recorded raw and trend data. By allowing the end-user user to switch on-the-fly from monitoring patients in one ICU to those in another, and by integrating an HL7 interface TeleTrend steps over the boundaries of a single ICU. Thus, it can be provide a medical enterprise-wide solution to the remote access of an important component of the electronic patient medical record. Currently in house validation, verification and alpha testing of the system are underway. PMID- 10538363 TI - Anatomical and physiological models for surgical simulation. AB - A considerable amount of effort has been aimed towards developing real-time deformable objects for surgical simulation, but very little work has been aimed towards including physiology within the soft tissue models. A simulator that links the structural and functional aspects of the human body would allow the user to develop a better understanding of the intrinsic link between anatomy and physiology. This positional paper discusses the challenges facing the creation of and the development of an integrated physiological and anatomical soft tissue model for use in surgical simulators. It explores the artificial dichotomy between anatomy and physiology and the issues it raises, by considering a suturing simulator capable of modelling ischaemia. PMID- 10538364 TI - Virtual 3D cutting for bone segment extraction in maxillofacial surgery planning. AB - An important step toward our main goal of a completely computer-based maxillofacial surgical planning system is the availability of tools for the surgeon to define bone segments from skull and jaw bones. We have developed an easy-to-handle user interface that employs visual and force-feedback devices to define subvolumes of a patient's volume dataset. This interface is a main component of our maxillofacial surgical planning tool MeVisTo-Jaw [1]. The defined subvolumes together with their spatial arrangements lead to an operation plan. PMID- 10538365 TI - Immersive surgical robotic interfaces. PMID- 10538366 TI - PET supports the hypothesized existence of a male sexual brain algorithm which may respond to treatment combining psychotherapy with virtual reality. PMID- 10538367 TI - The Wearable Motherboard: a flexible information infrastructure or sensate liner for medical applications. AB - Research on the design and development of a Sensate Liner for Combat Casualty Care has led to the realization of the world's first Wearable Motherboard or an "intelligent" garment for the 21st Century. This Georgia Tech Wearable Motherboard (GTWM) provides an extremely versatile framework for the incorporation of sensing, monitoring and information processing devices. The principal advantage of GTWM is that it provides, for the first time, a very systematic way of monitoring the vital signs of humans in an unobtrusive manner. Appropriate sensors have been "plugged" into this motherboard using the developed Interconnection Technology and attached to any part of the individual being monitored, thereby creating a flexible wearable monitoring device. The flexible data bus integrated into the structure transmits the information to monitoring devices such as an EKG Machine, a temperature recorder, a voice recorder, etc. The bus also serves to transmit information to the sensors (and hence, the wearer) from external sources, thus making GTWM a valuable information infrastructure. GTWM is lightweight and can be worn easily by anyone--from infants to senior citizens. GTWM has enormous potential for applications in fields such as telemedicine, monitoring of patients in post-operative recovery, the prevention of SIDS (sudden infant death syndrome), and monitoring of astronauts, athletes, law enforcement personnel and combat soldiers. PMID- 10538368 TI - Semi-automated analysis for MRI of breast tumors. AB - Magnetic resonance imaging (MRI) techniques have the potential to greatly improve breast cancer detection, diagnosis, and treatment. Currently, a major problem associated with breast MRI is the overwhelming amount of data acquired during an exam, and the time-intensive analysis required to evaluate the images. We have developed a software platform for semi-automated analysis to assess both the tumor extent and overall grade or severity based on our diagnostic criteria. In a test subset of over 50 patients, the automated program produced results more accurate overall than those measurements taken manually, with a reduction in time for analysis from approximately 45 minutes down to 5 minutes per patient study. PMID- 10538369 TI - PC-based telerehabilitation system with force feedback. AB - A PC-based orthopedic rehabilitation system was developed for use at home, while allowing for remote monitoring from the clinic. The home rehabilitation station has a Pentium II PC with graphics accelerator, Polhemus tracker, and a novel Multipurpose Haptic Control Interface with its own Pentium board. This interface is used to sample patient's hand positions and to provide resistive forces using the Rutgers Master II (RMII) glove. A library of virtual rehabilitation routines was developed using WorldToolKit software. At the present time, it consists of two physical therapy exercises (DigiKey and Ball) and two functional rehabilitation exercises (Peg Board test and Ball game). All VR exercises allow automatic and transparent patient data collection into an Oracle database. A remote Pentium II PC is connected with the home-based PC over the Internet and an additional video-conferencing connection. The remote computer running Oracle server is used to maintain the patient database, monitor progress and change exercise level of difficulty. This allows for timely patient progress monitoring and repeat evaluations over time from the Clinic. The system will soon start clinical trails at Stanford Medical School, with progress being monitored remotely from Rutgers University. Other rehabilitation haptic interfaces under development include devices for elbow, and knee rehabilitation connected to the Multipurpose Haptic Control Interface. PMID- 10538370 TI - Digital image recording: an integral aspect of video endoscopy. AB - In today's medical environment, efficiency and accuracy are the keys to successful outcomes. This concept is especially applicable during video endoscopic imaging, where image clarity is of utmost importance for medical applications. One of the most exciting advances in video endoscopy is the development of digital image storage devices. We report on a fast and efficient way to grab and store images during endoscopic procedures. Images can be stored on standard 3.5" floppy disks using an innovative new digital image recorder. The images are stored as high resolution JPEG files (640 x 480 x 24) for excellent clarity and detail. Once loaded onto the computer or network, the images can be placed into electronic medical records, imported into internet web pages, incorporated into slide presentations, and most importantly, stored in well organized archives which take up a minimum of space and are easily accessed by multiple users. This digital image recorder provides a complete system for acquiring images on a standard format 3.5" floppy disk--the world's most ubiquitous storage format. PMID- 10538371 TI - A telementored trans-rectal ultrasound guided prostate biopsy. AB - The purpose of this project was to demonstrate the efficacy and feasibility of a real-time telementored transrectal ultrasound-guided prostate biopsy. The physician (P) was located at a remote site while the physician's extender (PE) was with the patient at another location. The (P) directed and guided the (PE) through the biopsy procedures utilizing video and audio contact, and viewing electronically transmitted ultrasound images. This procedure occurred at Walter Reed Army Medical Center in Washington, DC where the patient and (PE) were located in a surgical suite, and the (P) was located in another part of the hospital. Audio, video and ultrasound images were transmitted via a simulated T1 line. The success of this procedure demonstrates both feasibility and efficacy of invasive procedures telementored by a (P) from a remote location while the (PE) is with the patient and receiving remote instruction. The achievement of a real time telementored biopsy has significance in increasing access to care by those populations living in geographically remote areas or urban socioeconomically isolated areas. It also has significance for physicians whose primary clinical responsibilities are in urban or university environments who also desire to provide care to these underserved, difficult to serve or expensive to serve populations. This telementored procedure was the first step in development of a large program with the Medical University of South Carolina. The intent of the program is to conduct prostate screening and biopsy where the physician and patient can maintain visual and audio contact for consultation while the physician is simultaneously viewing ultrasound images and electronic patient records. The overall effectiveness of the program will be evaluated in terms of improvement in clinical process of care, improved health outcomes, improved access to care, and patient/physician satisfaction with the experience. PMID- 10538372 TI - The ergonomics of virtual reality: human factors in developing clinical-oriented virtual environments. AB - Virtual Reality (VR) is usually described as a collection of technological hardware: a computer capable of 3D real-time animation, a head-mounted display, data gloves equipped with one or more position trackers. However, this focus on technology is disappointing for clinicians interested in developing virtual environments to be used in assessment and therapy. To overcome this limitation this chapter describes VR as an advanced communication tool: a communication medium in the case of multi-user VR and a communication interface in single-user VR. Two are the core characteristics of VR as communication tool: the perceptual illusion of nonmediation and the sense of community. The first characteristic of a satisfying virtual environment is the disappearance of mediation, a level of experience where both the VR system and the physical environment disappear from the user's phenomenal awareness. The second characteristic is the sense of community developed by interaction. Through interaction made possible by multi user VR, individuals find or form groups that share interests. So, information exchange becomes the carrier for expressing self-concept and eliciting emotional support. Within this view, experiencing presence and telepresence does not depend so much on the faithfulness of the reproduction of 'physical' aspects of 'external reality'--which is also a social production, and not a primitive or 'natural' fact--as on the capacity of simulation to produce a context in which social actors may communicate and cooperate. The consequences of this approach for the design and the development of clinical oriented VR systems are presented, together with the methodological and technical implications for the study of advanced human-computer interaction. PMID- 10538373 TI - 3-D position measurement for microsurgical evaluation. AB - A compact system for high-resolution three-dimensional measurement of microsurgical instrument tip position in the laboratory is under development. The system is useful for quantification of tremor, wander, and drift, as well as for other studies characterizing surgeons' motion in microsurgery, and for evaluation of engineered devices for enhancement of microsurgical positioning accuracy. Preliminary results are presented. PMID- 10538374 TI - Surgeon-tool force/torque signatures--evaluation of surgical skills in minimally invasive surgery. AB - The best method of training for laparoscopic surgical skills is controversial. Some advocate observation in the operating room, while others promote animal and simulated models or a combination of surgical related tasks. The mode of proficiency evaluation common to all of these methods has been subjective evaluation by a skilled surgeon. In order to define an objective means of evaluating performance, an instrumented laparoscopic grasper was developed measuring the force/torque at the surgeon hand/tool interface. The measured database demonstrated substantial differences between experienced and novice surgeon groups. Analyzing forces and torques combined with the state transition during surgical procedures allows an objective measurement of skill in MIS. Teaching the novice surgeon to limit excessive loads and improve movement efficiency during surgical procedures can potentially result in less injury to soft tissues and less wasted time during laparoscopic surgery. Moreover the force/torque database measured in this study may be used for developing realistic virtual reality simulators and optimization of medical robots performance. PMID- 10538375 TI - New approaches to virtual environment surgery. AB - This research focused on two main problems: 1) low cost, high fidelity stereoscopic imaging of complex tissues and organs; and 2) virtual cutting of tissue. A further objective was to develop these images and virtual tissue cutting methods for use in a telemedicine project that would connect remote sites using the Next Generation Internet. For goal one we used a CT scan of a human heart, a desktop PC with an OpenGL graphics accelerator card, and LCD stereoscopic glasses. Use of multiresolution meshes ranging from approximately 1,000,000 to 20,000 polygons speeded interactive rendering rates enormously while retaining general topography of the dataset. For goal two, we used a CT scan of an infant skull with premature closure of the right coronal suture, a Silicon Graphics Onyx workstation, a Fakespace Immersive WorkBench and CrystalEyes LCD glasses. The high fidelity mesh of the skull was reduced from one million to 50,000 polygons. The cut path was automatically calculated as the shortest distance along the mesh between a small number of hand selected vertices. The region outlined by the cut path was then separated from the skull and translated/rotated to assume a new position. The results indicate that widespread high fidelity imaging in virtual environment is possible using ordinary PC capabilities if appropriate mesh reduction methods are employed. The software cutting tool is applicable to heart and other organs for surgery planning, for training surgeons in a virtual environment, and for telemedicine purposes. PMID- 10538376 TI - Automatic modeling of knee joint motion for the virtual reality dynamic anatomy (VRDA) tool. AB - A system for automatic modeling of anatomical joint motion for use in the Virtual Reality Dynamic Anatomic (VRDA) tool is described. The modeling method described in this article relies on collision detection. An original incremental algorithm use this information to achieve stable positions and orientations of the tibia on the femur for each angle considered between these two components on the range of motion. The stable states then become the basis for a look-up table employed in the animation of the motion of the joint. The strength of the method lies in its robustness to animate any "normal" anatomical joint, given a set of kinematic constraints for the joint type as well as an accurate 3D geometric model of the joint. The demonstration could be patient specific (based on a person's real anatomical data from an imaging procedure such as CT scanning) or scaled from a generic joint based on external patient measurements. The modeling method has been implemented on a generic knee model for use in the VRDA tool. PMID- 10538377 TI - Tests on reliability of a prostate biopsy telerobotic system. AB - This paper deals with the development of a robotic (mechanical-electronic) system that operates in the field of surgery, with new sensors and equipment ("Biopsy Program"). The system permits a rapid analysis of the mechanical needle + needle holder + mechanism system for penetration and aspiration for surgery, for prostate biopsies. These operations benefit from the use of a SR 8438 Sankyo Scara robot; they are also remote-controlled, i.e. via telerobotics. They require accurate positioning in known points of three-dimensional space with a high degree of precision. The safety of the surgical operations is guaranteed by the most complete observance of regulations under European Union Directives and Decrees 626 and 242 of Italian laws. The system proposed represents a big step for the application of industrial solutions, precisely for industrial companies in the medical and surgical field. PMID- 10538378 TI - Limitations of distributed segmentation for three-dimensional radiological modeling. AB - While the use of three-dimensional models has been shown to be useful clinically, the specialized computational equipment and expertise necessary for their construction and use keeps these tools out of reach of most physicians. This paper explores the construction of a Web-based Java application that allows medical radiological models to be built on a remote server and navigated locally on the physician's desktop PC. This paper will also address issues that arose from a public, unrestricted testing of usability over the Internet, such as model size management, easy navigation, processor loading and security. Based on observations and data collected, we suggest what steps are necessary to make a telemedicine application useable in a true clinical setting. PMID- 10538379 TI - Laser 3-D scanning for surface rendering in biomedical research and education. AB - A technique based on laser scanning is applied to body parts and organs. Laser light is projected onto the surface of objects and recorded by CCD sensors. This is a fast and flexible method for accurately scanning surface geometry. It also allows conversion to NURBS patches. We have, so far, made 'point-cloud' surface renderings from a head model, a plastic brain model, a human brain, moulds of bites and a cranium. The limits, quality, efforts and costs of employing the laser 3-D scanning technique are evaluated. The experiments are currently in progress and results give interesting 3D renderings and attempts to triangulated solid-models. PMID- 10538380 TI - Interactive volume visualization using "intelligent movies". AB - High quality visualization of medical volume models as performed by the VOXEL-MAN and similar systems is still too time consuming and the interaction complicated when sophisticated tools like dissection are used. We hence developed a new paradigm allowing to create simpler derivatives of the model, called "intelligent movies". These are in QuickTime or QuickTime VR format which allow interactive exploration with two degrees of freedom. As a decisive novelty, we extended it by a pixelwise link to the knowledge base which may be queried in the image context. Thus scenes emphasizing a selected aspect of the volume model may be created as intelligent movies, which a user (referring physician, student) can explore largely with the functionality of VOXEL-MAN, but in real time--on any standard PC -and also via a JAVA applet within web browsers. This is shown with the example of 3D interactive anatomical atlases and clinical cases. PMID- 10538381 TI - An interface for precise and comfortable 3D work with volumetric medical datasets. AB - We have developed a 3D/2D paradigm of interaction that combines manipulation of precise 3D volumetric data with unambiguous widget interaction. Precise 3D interaction is ensured by a combination of resting the lower arms on an armrest and pivoting the hands around the wrist. Unambiguous 2D interaction is achieved by providing passive haptic feedback by means of a virtual control panel whose position coincides with the physical surfaces encasing the system. We have tested this interface with a neurosurgical planning application that has been clinically used for 17 skull-base cases at two local hospitals. PMID- 10538382 TI - A portable virtual environment knee arthroscopy training system with objective scoring. AB - We have developed a Virtual Environment Knee Arthroscopy Training System (VE KATS). Formative analysis indicated several features which surgical trainees indicated were necessary for a system to be useful. We have addressed this need by incorporating both rigid and deformable objects, simultaneous use of camera and surgical instrument, improved optical modeling, portability and low cost. A major advance has been the incorporation of automated objective scoring of performance. VE-KATS has now progressed to a viable training system which will be of practical benefit to trainee orthopaedic surgeons. PMID- 10538383 TI - Early experience and validation work with Procedicus VA--the Prosolvia virtual reality shoulder arthroscopy trainer. AB - Shoulder arthroscopy is a difficult procedure, commonly used for both diagnostic and therapeutic purposes. Until now, the majority of training has been done in theatre, assisting and practising under supervision. Few good simulations exist. Procedicus VA, from Prosolvia Clarus, is a virtual reality simulation of shoulder arthroscopy, with interactive graphics and haptic feedback. The simulator has various modes including anatomy manipulation pathology subacromial decompression. We describe our experience with the simulator, attempting to validate some of the scoring mechanisms, and highlighting some of the pitfalls discovered as the simulator is first trialled by surgeons. This early experience has highlighted both successful aspects of the simulator, and some of the initial pitfalls. Our initial experience confirms the need for close collaboration between virtual programmers and surgical trainers. We are revising the assessment criteria over the coming months. PMID- 10538384 TI - Interactive volume visualizations for synchronous and asynchronous remote collaboration. AB - The value of information acquisition is not simply to acquire data at increased precision, but to make these data both available and usable to as many specialists as possible. We have developed a working prototype for intuitive realtime interaction over the Internet. Current scenarios include use by expert head and neck surgeons, for use in preoperative assessment of head and neck cancers, and remote synchronous collaboration scenarios such as expert-expert and expert-referring physician. This prototype demonstrates a shared virtual environment that allows multiple users to interactively manipulate large volumetric data sets across long distances. The system is easily extensible to many other applications that utilize discrete data sampling and thus is readily extensible to a wide range of scientific investigations. PMID- 10538385 TI - Virtual reality based surgery simulation for endoscopic gynaecology. AB - Virtual reality (VR) based surgical simulator systems offer very elegant possibilities to both enrich and enhance traditional education in endoscopic surgery. However, while a wide range of VR simulator systems have been proposed and realized in the past few years, most of these systems are far from able to provide a reasonably realistic surgical environment. We explore the basic approaches to the current limits of realism and ultimately seek to extend these based on our description and analysis of the most important components of a VR based endoscopic simulator. The feasibility of the proposed techniques is demonstrated on a first modular prototype system implementing the basic algorithms for VR-training in gynaecologic laparoscopy. PMID- 10538386 TI - Development of a robotic navigation system for neurosurgery. AB - This paper presents a robotic navigation system for image-guided neurosurgery, which can be applied to the treatment of Parkinson's disease and biopsy of brain tumor. The system integrates a computer for real-time display of brain anatomy, a magnetic tracking device for measuring the positions and orientations of surgical instruments, and a robot manipulator for guiding surgical instruments to the preplanned positions and orientations. The computer display of brain anatomy offers a convenient tool for surgeons to diagnose brain disease and to plan safe surgical paths; while the tracking device assists the robot manipulator to automatically guide surgical instruments to the preplanned direction. The registrations among the tracking device, the image system, and the robot are completed on the base of coordination mappings of external markers. An experiment of using a skull model for simulating a robotic biopsy of brain tumor has been done to verify the performance of the navigation system. The result shows that the system can accomplish a positioning accuracy around 2 mm. PMID- 10538387 TI - CathSim. PMID- 10538388 TI - CathSim: an intravascular catheterization simulator on a PC. AB - The development of a medical simulator that incorporates substantial training value and realism into an affordable product has been a huge challenge for the simulation community. A large hurdle to making an inexpensive simulator has been the high cost of the computers needed for adequate realism. We have met this challenge by developing CathSim, a low-cost medical simulator that integrates force feedback, multimedia, and 3D graphics simulation technology on an industry standard PC. This product is commercially available and is currently being used by numerous training institutions and hospitals. The CathSim system includes software and a force feedback interface device. The platform and device can be used to train health care providers to perform needle-stick medical procedures. Our first module teaches users the techniques of peripheral intravenous (i.v.) catheterization. Other training modules that will be added to the CathSim platform include central venous catheter (CVC) insertion and peripherally inserted central catheter (PICC) placement. This paper discusses the challenges of this project and the trade-offs and solutions that we developed to overcome them. We describe our process of analyzing and prioritizing the medical tasks necessary to correctly perform peripheral intravenous catheterization. This analysis and prioritization was used to decide which tasks would be included in the simulator and how the included tasks would be replicated. We discuss the method by which we obtained the needed realism in the 3D graphics rendering and in the tactile feedback of the input device. We illustrate how we blended together simulation and multimedia technology to ensure adequate immersion and training efficacy, while keeping the system cost to a minimum. PMID- 10538389 TI - Virtual reality and women's health: a breast biopsy system. AB - Minimally invasive procedures are becoming much more common in surgical practice because of the many advantages for patient comfort and convenience, and improved surgical access. However some of the major problems leading to occasional surgical errors with this minimal access method are restricted vision, limited sense of touch, difficulties in identification in 3D space of the position of the instrument tips, and their handling during delicate, short-distance movements toward the surgical target area. These factors emphasize the need for computer simulated training in surgical manipulations and procedures in preparation for conducting them in patients. The key new feature of our proof-of-concept training simulator is a preventive mechanism that serves at least two functions. As the surgical target (or a critical structure) is approached, a haptically generated preventive force forewarns the surgeon, making it possible to abort those maneuvers that may lead to adverse results. By announcing a potential collision of a virtual instrument tip with a surgical target, the time used for searching for the target is shortened, and the haptic signal minimizes the potential of tissue damage. This real-time, interactive, virtual reality based, haptic breast biopsy-training simulation is a PC/NT based multitasking, multithreading system. It is based upon an advanced force feedback device. The system monitors and indirectly guides the surgeon's movements, while providing high fidelity visual and force feedback cues as the area of surgical interest is approached. Our first application is with human breast. PMID- 10538390 TI - CAREN--Computer Assisted Rehabilitation Environment. AB - The CAREN project concerns the development of a virtual reality environment in which the agility of healthy subjects and patients can be tested in a variety of reproducible conditions. CAREN is made by customizing hardware and developing software to enable measurements of motion of a subject in detail as a response to a perturbation from the computer driven motion platform. After feeding the data in a human body model simulation, joint moments of force and muscle activation can be calculated. From the time patterns of these responses, inferences can be made concerning the motor programs the subjects launch. Any primary problem in a motor program, resulting in functional failure or inadequacy, can be identified down to the joint and muscle group. Secondary adaptations of patients to a limitation in the periphery (such as lack of muscle force) can be separated from the primary ones. Inadequacy of complete motor programs in children with movement disorders can be classified, recorded and related to progress that may occur. Especially the understanding of compensation strategies in patients may lead to a better therapeutic attitude. CAREN offers not only a test environment with means of almost unlimited exploratory behaviors for patients, but constitutes also a strong tool for motor control research. PMID- 10538391 TI - A technique for very high accuracy image intensifier calibration. AB - Accurate characterisation of the image distortion within a fluoroscopic image intensifier is critical if it forms the vision component of an image guided surgical system. By considering non-linear dynamic distortion it is possible to greatly increase the accuracy of the image intensifier, although at the cost of some image quality. PMID- 10538392 TI - Simulation of tissue cutting and bleeding for laparoscopic surgery using auxiliary surfaces. AB - Realistic simulation of tissue cutting and bleeding is important components of a surgical simulator that are addressed in this study. Surgeons use a number of instruments to perform incision and dissection of tissues during minimally invasive surgery. For example, a coagulating hook is used to tear and spread the tissue that surrounds organs and scissors are used to dissect the cystic duct during laparoscopic cholecystectomy. During the execution of these procedures, bleeding may occur and blood flows over the tissue surfaces. We have developed computationally fast algorithms to display (1) tissue cutting and (2) bleeding in virtual environments with applications to laparoscopic surgery. Cutting through soft tissue generates an infinitesimally thin slit until the sides of the surface are separated from each other. Simulation of an incision through tissue surface is modeled in three steps: first, the collisions between the instrument and the tissue surface are detected as the simulated cutting tool passes through. Then, the vertices along the cutting path are duplicated. Finally, a simple elastic tissue model is used to separate the vertices from each other to reveal the cut. Accurate simulation of bleeding is a challenging problem because of the complexities of the circulatory system and the physics of viscous fluid flow. There are several fluid flow models described in the literature, but most of them are computationally slow and do not specifically address the problem of blood flowing over soft tissues. We have reviewed the existing models, and have adapted them to our specific task. The key characteristics of our blood flow model are a visually realistic display and real-time computational performance. To display bleeding in virtual environments, we developed a surface flow algorithm. This method is based on a simplified form of the Navier-Stokes equations governing viscous fluid flow. The simplification of these partial differential equations results in a wave equation that can be solved efficiently, in real-time, with finite difference techniques. The solution describes the flow of blood over the polyhedral surfaces representing the anatomical structures and is displayed as a continuous polyhedral surface drawn over the anatomy. PMID- 10538393 TI - Virtual cutting of anatomical structures. AB - In the context of medicine cutting is one of the most important operations. Many surgical tasks start with an incision, allowing the surgeon to access the region of interest, using either conventional or minimal invasive surgery techniques. The work presented herein should be seen in the context of surgical training and preoperative planning using computer assisted medical simulation systems. The main result of this work is the proposed cutting algorithm based on predefined templates. To integrate this approach in the existing surgical training simulator and to handle additional tasks like soft tissue deformation, needed by the medical environment, a generic simulation framework was developed. This framework is able to utilize given multiprocessor- and network-environments to achieve the desired realtime interactivity. PMID- 10538394 TI - A voice-controlled robotic assistant for neuroendoscopy. AB - This paper describes experiments with a voice-controlled robot system to be used in endoscopic neurosurgery. The robot was a modified version of the robot described in previous publications of the group at Fraunhofer IPA and HSK. To control the robot a voice-controlled user interface was developed. The experiments were conducted on cadavers for three standard approaches in neuroendoscopy. The goal was to gain experience with a voice-controlled user interface and also with the set-up and use of the robotic system under clinical conditions. The results indicate that modifications to the robot and user interface are necessary. However the overall feasibility of the application was demonstrated. PMID- 10538395 TI - The correction of MR images distortion with phantom studies. AB - In the field of Image Guided surgery, Geometrical accuracy of images acquired from CT and MR is an essential prerequisite for good registration process. In this paper, we present the experiment with MR phantom. We visualize the distortion of MR images in 3-D shape and modify it by surface fitting algorithm using the control Point. PMID- 10538396 TI - Prostate biopsy schemes: 3-D visualization-based evaluation. AB - We have developed a prostate needle biopsy visualization system for the evaluation and optimization of biopsy schemes. Three-dimensional (3-D) prostate surface models have been reconstructed from the digitized whole-mount radical prostetactomy specimens with localized cancers. We have conducted evaluation of five major biopsy schemes with a total of 201 3-D prostate models. These are sextant, 10-pattern, 12-pattern, 14-pattern, and the 5-region schemes. The 10- and 12-pattern biopsy schemes had a 99.0% detection rate, while the rate of traditional sextant biopsy scheme was only 72.6%. The 5-region biopsy scheme had a 90.5% detection rate and the 14-pattern, which includes all the biopsies used in the above schemes, added only one additional positive case (99.5%). Our results suggest that biopsy schemes that use laterally placed biopsies based on the five region anatomical model are superior to the routinely used sextant biopsy scheme. Significant correlation is found between the tumor volume and the positive needle core volume for each of these five schemes. The 10-pattern scheme is the best in cancer detection among these five biopsy schemes. PMID- 10538397 TI - An improved stereotactic technique for cyst cannulation. AB - Stereotactic techniques for cannulation of cystic structures, within the brain, are well known. Superimposed structures (vessels, ventricles, etc.) may make this problematic as does the need to approach the cystic structure perpendicular to its tangent plane (rather than "glancing") as with a craniopharyngioma cyst. To facilitate a three-dimensional visualization of the trajectory, we have employed digital holography. Transparent holographic images of cystic structures, ventricles, and sulci are rendered from T2-weighted MR data. Holographic images of vascular structures are rendered from CT or MR angiographic data. Vascular holograms are superimposed over the brain holograms, demonstrating the spatial relationships of these structures with regard to each other. Holographic images of the skull are rendered from CT slices. A Laitinen stereotactic frame (Sandstrom) is placed on the patient prior to obtaining the CT. The skull, pre existing shunt catheters, and the stereotactic frame are all readily visible. The brain and vascular holograms are superimposed on these. The resulting image clearly demonstrates cystic structures, ventricles, vessels, pre-existing catheters, all within the skull and stereotactic frame. Using this holographic image as a "phantom", the actual Laitinen stereotactic frame is placed within its holographic image. The optical trajectory is then chosen, and the articulated arm of the stereotactic device is so adjusted. Subsequently, the frame is used to effect stereotactic placement of the cannula, in the usual manner. The major advantages of this technique are twofold. The first advantage lies with the fact that the surgeon can readily visualize the entire trajectory of the needle, and easily appreciate all structures which may be encountered by the needle on its passage from the skull to the target. Presumably, the surgeon's knowledge of anatomy would unable such knowledge to be apparent, but in complex cases the "safe" corridor may be rather small, and its limits may not be intuitively obvious. This is all the more the case, when obstacles along the pathway are pathologically distorted, or when they are not of tissue origin (shunt catheters, etc.). Employing this technique, we have successfully cannulated cystic structures in six patients, three of which presented with complex trajectory problems. PMID- 10538398 TI - A virtual instrument ergonomics workstation to measure surgeons' physical stress. AB - Videoendoscopic (VES) instruments have poor force transmission properties and often require surgeons to employ awkward hand and arm positions. In order to compare the physical workload of laparoscopic surgery to open surgery, we collected long-duration EMG records from the thumb (thenar compartment) of six surgeons performing suturing and knot tying in a training box using both open and VES techniques. EMG signals were acquired using a LabVIEW Virtual Instrument and analyzed using a Modified Exposure Variation Analysis (MEVA) algorithm. Standard EMG indices and the MEVA analysis demonstrated significantly greater amplitude and duration of EMG signals using VES technique compared to open technique. Our results suggest that the use VES techniques requires a greater intensity of physical effort than open surgery techniques. PMID- 10538399 TI - Fast finite element modeling for surgical simulation. AB - Given the geometric complexity of anatomical structures, realistic real-time deformation of graphical reconstructions is prohibitively computationally intensive. Instead, real-time deformation of virtual anatomy is roughly approximated through simpler methodologies. Since the graphical interpolations and simple spring models commonly used in these simulations are not based on the biomechanical properties of tissue structures, these "quick and dirty" methods typically do not accurately represent the complex deformations and force-feedback interactions that can take place during surgery. Finite element (FE) analysis is widely regarded as the most appropriate alternative to these methods. Extensive research has been directed toward applying the method to modeling a wide range of biological structures, and a few simple FE models have been incorporated into surgical simulations. However, because of the highly computational nature of the FE method, its direct application to real-time force-feedback and visualization of tissue deformation has not been practical for most simulations. This limitation is primarily due to the overabundance of information provided by the standard FE approaches. If the mathematics is optimized to yield only the information essential for the surgical task, computation time can be drastically reduced. Parallel computation and preprocessing of the model before the simulation begins can also reduce the size of the problem and greatly increase computation speed. Such methodologies are being developed in a combined effort between the Human Interface Technology Laboratory (HIT Lab) and the Mechanical Engineering Department of the University of Washington. We have created computer demonstrations which support real-time interaction with simple finite element soft tissue models. In collaboration with the Division of Dermatology, a real time skin surgery simulator is being developed using these fast FE methods. PMID- 10538400 TI - Defining the role of haptic feedback in minimally invasive surgery. AB - INTRODUCTION: The applications of Minimally Invasive Surgery (MIS) and Laparoscopy are rapidly expanding. Despite this expansion, the technology related to our understanding of the importance of haptic feedback related to laparoscopic surgery remains in its infancy. While many surgeons feel that the use of minimally invasive techniques eliminates force feedback and tactile sensation, the importance of haptics in MIS has not been fully evaluated. Moreover, there is considerable interest in the development of haptic simulators for MIS even though the importance of force feedback remains poorly understood. This study was designed to determine the ability of novice surgeons to interpret haptic feedback with respect to texture, shape and consistency of an object. METHOD: Subjects were presented objects in a random order and participants were blinded as to their identity. Inspection by direct palpation, palpation with conventional instruments, and palpation with laparoscopic instruments was performed on all objects. Statistical analysis of the data was performed using a Fischer exact probability test. RESULTS: Direct palpation provided the greatest degree of haptic feedback and was associated with the highest accuracy for texture discrimination, shape discrimination, and consistency discrimination. A significant decrease in the ability to identify shapes was noted with both CI and LI. A significant decrease in the ability to differentiate consistency was noted for LI only. When comparing palpation with conventional instruments to palpation with laparoscopic instruments, there was no significant difference in shape or texture discrimination. There was, however, a significant decrease in consistency discrimination. CONCLUSION: This data indicates that laparoscopic instruments do in fact provide the surgeon with haptic feedback. While the instruments change the information available to the surgeon, interpretation of the texture, shape and consistency of objects can be performed. Our ongoing work is directed at further defining force interactions. Through the use of force feedback impulse devices in VR simulators, one should be able to create a more realistic theatre in which the novice surgeon can learn operative skills that will readily translate into the operating room. PMID- 10538401 TI - Haptic rendering of isosurfaces directly from medical images. AB - Virtual environments for surgical training, planning and rehearsal have the potential to significantly enhance patient treatment and diagnosis. Haptic feedback devices provide forces to the physician through a manipulator, simulating palpation, scalpel cuts, or retraction of tissue. While haptics have been studied in other fields, medical applications of haptics remain in their infancy. We propose a method of haptically rendering isosurfaces (representing hard structures) directly from anatomical datasets rather than through traditional intermediate graphical representations such as polygons. Our algorithm determines an implicit surface representation of a volumetric isosurface on the fly, and renders the structure using standard haptic algorithms to calculate the forces felt by the user. This approach has the advantage of providing easy access to the rich volume dataset containing the actual anatomy. By relating Hounsfield units to density, haptic rendering has the ability to provide different resistances based on the tissue type being rendered. We developed and tested our algorithm using a quadric implicit surface, a common primitive in computer graphics, and have applied it to a variety of anatomical image datasets. Our paper describes the algorithm in sufficient detail to facilitate reproduction by others. PMID- 10538402 TI - Realtime simulation of tissue deformation for the nasal endoscopy simulator (NES). PMID- 10538403 TI - PreOp endoscopic simulator: a PC-based immersive training system for bronchoscopy. AB - The high cost of simulators that offer adequate realism for training has been a major challenge for the simulation community. The cost of the computers alone has been too high for most training institutions to afford. We have met this challenge by developing the PreOp Endoscopic Simulator, our second generation of low-cost medical simulators. The PreOp system integrates multimedia, 3D graphics simulation, and force feedback technology on a PC. This paper discusses the challenges of this project and the trade-offs and solutions that we developed to overcome them. We discuss our process of analyzing and prioritizing the medical tasks necessary to correctly perform flexible bronchoscopy. In addition, we illustrate how we blended together simulation and multimedia technology to ensure adequate immersion and training efficacy, while keeping the system cost to a minimum. PMID- 10538404 TI - A system for the simulation and planning of orthodontic treatment using a low cost 3D laser scanner for dental anatomy capturing. AB - The detection and correction of malocclusions and other dental abnormalities is a significant area of work in orthodontic diagnosis. To assess the quality of occlusion between the teeth the orthodontist has to estimate distances between specific points located on the teeth of both arches. Distance measuring is based on the observation, by the orthodontist, of a plaster model of the mouth. Gathering of information required to make the diagnosis is a time consuming and costly operation. On the other hand, obtaining and manipulation of plaster casts constitute a huge problem in clinics, due to both the large space needed and high costs associated with plaster casts manufacturing. For this problem we present a new system for three-dimensional orthodontic treatment planning and movement of teeth. We describe a computer vision technique for the acquisition and processing of three-dimensional images of the profile of hydrocolloids dental imprints taken by mean of a own developed 3D laser scanner. Profile measurement is based on the triangulation method which detects deformation of the projection of a laser line on the dental imprints. The system is computer-controlled and designed to achieve depth and lateral resolutions of 0.1 mm and 0.2 mm, respectively, within a depth range of 40 mm. The developed diagnosis software system (named MAGALLANES) and the 3D laser scanner (named 3DENT) are both commercially available and have been designed to replace manual measurement methods, which use costly plaster models, with computer measurements methods and teeth movement simulation using cheap hydrocolloid dental wafers. This procedure will reduce the cost and acquisition time of orthodontic data and facilitate the conduct of epidemiological studies. PMID- 10538405 TI - Hierarchical decomposition of laparoscopic procedures. AB - The purpose of this report is to outline the hierarchical decomposition of surgical procedures, from surgical steps through tasks and subtasks to tool motions, and highlight implications for surgical training systems. Three common laparoscopic procedures were analysed: cholecystectomy, inguinal hernia repair, and Nissen fundoplication. In laparoscopic training workshops and operating rooms, our observational research included split screen videotaping of both the endoscopic view and our video camera's view of the primary surgeon. Videotapes were extensively annotated and analysed to yield timelines of each procedure, with component surgical steps, substeps, tasks, and subtasks duration as a function of procedure. The hierarchical decomposition of surgical procedures provides a framework for structuring a systematic approach to training, in the real and simulated environment. An example comparing variations in the fundoplication procedure is presented. Our results also have important implications for the design and assessment of new technology and intelligent tools in endoscopic surgery. PMID- 10538406 TI - Virtual reality: a wholistic approach to rehabilitation. AB - Virtual Reality (VR) is proving to be an effective treatment tool in rehabilitation. Currently we are providing VR therapy to patients in an out patient clinic and an adult day center. The main focus to this paper is to show how VR is used in occupational therapy to improve balance and dynamic standing tolerance with geriatric patients. We will discuss our research findings of specific treatment approaches that we are using in therapy. The benefits of VR in rehab, as well as other rehab applications for this modality will also be discussed. We will illustrate the importance of addressing the visual and motor systems simultaneously for maximum efficiency in achieving our rehabilitation goals. The ultimate goal in occupational therapy is to increase a patient's level of independence in activities of daily living and therefore improving their quality of life. PMID- 10538407 TI - Thin walled models for haptic and graphical rendering of soft tissues in surgical simulations. PMID- 10538408 TI - Geographic Information Systems (GIS) in public health practice in the new millennium. PMID- 10538409 TI - Maps and health: the challenges of interpretation. PMID- 10538410 TI - Geographic Information Systems (GIS) for state and local public health practitioners, Part 2. PMID- 10538411 TI - Getting started with Geographic Information Systems (GIS): a local health department perspective. PMID- 10538412 TI - Local health departments and GIS: the perspective of the National Association of County and City Health Officials. PMID- 10538414 TI - Revitalizing communities with Geographic Information Systems (GIS): HUD's Community 2020 software. PMID- 10538413 TI - GIS and decision making for public health agencies: childhood lead poisoning and welfare reform. PMID- 10538415 TI - Epi Info and Epi Map: current status and plans for Epi Info 2000. PMID- 10538416 TI - Spatial analysis and mapping on the Internet. PMID- 10538417 TI - Toward a GIS sampling frame for surveys of local health departments and local boards of health. PMID- 10538418 TI - Sources of spatial data for community health planning. PMID- 10538419 TI - The future of GIS in public health management and practice. PMID- 10538420 TI - Hitting their mark. Two outpatient organizations partner with vendors to win award. PMID- 10538421 TI - Model citizens. Outsourcing helps start-up Medicare HMO. AB - Health Plans of Pennsylvania (HPP), the managed care arm of Crozer-Keystone Health System, in Media, Pa. PROBLEM: Selecting the information systems and building the infrastructure to support the start-up of a new Medicare HMO product. SOLUTION: HPP chose to outsource the information systems needed to integrate all the components of managed care administration into a cost-effective and cohesive program. RESULTS: Because of its aggressive programming and start-up of the MedCarePlus product offering, HPP became the first plan in the country to submit Medicare claims data electronically for encounter reporting to the Health Care Financing Administration (HCFA). KEYS TO SUCCESS: "Through an integrated team approach, an organization truly can benefit from the economies of scale gained through outsourcing." PMID- 10538422 TI - Great Britain: the Wellcome Trust goes online. PMID- 10538423 TI - Africa: telecom creates a small world. PMID- 10538424 TI - Steal the wheel. Companies like Ford, Hyatt and your local PD are steering healthcare executives to IT solutions. PMID- 10538425 TI - Picture perfect. Cost-effective solutions help advanced medical imaging make business sense. PMID- 10538426 TI - In for the long-term. The long-term care industry is in trouble. Careful IT planning may be its salvation. PMID- 10538427 TI - The regulatory process, the Food and Drug Administration, and the silicone breast implant controversy. PMID- 10538428 TI - Managed care: a critical review. AB - Widely perceived and accepted as the simplest way to control escalating health care costs, managed care is the way health care is now, and will continue to be, delivered in the foreseeable future. Growing numbers of Medicaid beneficiaries are required to participate in managed health care and Medicare beneficiaries are strongly encouraged to do so. In essence, America's poor and elderly are serving as the vanguard for health care reform. This article describes the evolution of managed care in the United States, then examines the implications of the transfer of financial risk to health care providers; the impact of managed care on existing inequalities in access to health care and services; the quality of managed care compared with fee-for-service arrangements; and the delivery of mental health services under managed care. We suggest that the role of the social work profession in managed care should be to mitigate the costs borne by clients, and offer specific suggestions for advocates in this arena. PMID- 10538429 TI - Medicare physician payment reform and the utilization of cardiovascular procedures. AB - Effective 1992, the US Congress implemented the Resource-Based Relative Value Scale (RBRVS) for pricing and reimbursing Medicare physician services. This study evaluates the post-1991 impacts of RBRVS on the utilization volumes of two leading cardiovascular procedures--Percutaneous Transluminal Coronary Artery (PTCA) and Coronary Artery Bypass Grafts (CABG). The regression model results based on HCFA-supplied data suggest that the new reimbursement policy reduced (increased) utilization volumes of the more (less) expensive CABG (PTCA) procedures. Physician adjustments to tightened HCFA reimbursements are partly aided by the increased percentage of Medicare patients enrolled in Medigap. The RBRVS fee schedule appears to be meeting its intended cost containment policy goals for the leading cardiovascular procedures. PMID- 10538430 TI - Community postpartum care needs assessment and systems development for low income families. AB - Decreased lengths of stay for U.S. childbirth hospitalization, infant morbidities, repeat adolescent pregnancies, and high no-show rates for postpartum visits among disadvantaged populations suggest barriers to continuity of maternity care. Findings of a survey of maternity health professionals (N = 78) providing postpartum case management with an urban Healthy Start project indicated less tracking and follow-up for postpartum care as compared to prenatal care as well as maternal postpartum health education, social support, and environmental needs. Recommendations included: (a) earlier timing of postpartum visit, (b) community care sites and home visiting, (c) coordinated postpartum maternal and infant care, and (d) increased postpartum psychosocial and environmental services. PMID- 10538431 TI - Prenatal, intrapartum, and newborn care provided by health maintenance organizations: Medicaid versus commercial enrollees. AB - A large majority of states have adopted a policy of enrolling Medicaid and other low-income populations in managed care organizations. Analysts have raised several questions relating to whether these populations will achieve appropriate access and utilization of services, including prenatal, intrapartum, and newborn care. Data from a national survey of health maintenance organizations are used to compare access to these kinds of care for Medicaid and commercial enrollees. Overall, utilization of services by the two populations is comparable. PMID- 10538432 TI - Public evaluation of an announcement of public interests communicated by an AIDS patient. AB - Announcements of public interests (APIs) on television (TV) have been an important means of AIDS prevention. Recently in Hong Kong, the first API on TV which was communicated by a male AIDS patient, named J.J., was displayed on the screen from April to December, 1995. The objective of this study is to examine the effectiveness of this API, as compared with other APIs previously shown in Hong Kong. Collecting data from a random sample of 1,275 residential respondents in Hong Kong through a telephone survey, this study demonstrated that the public gave very favorable evaluation on J.J.'s API, when compared with other APIs. Remarkable impacts of J.J.'s API on the public included increasing concern for AIDS, condom use, and testing blood for HIV infection, and reducing the number of sex partners. The impacts tended to be more salient on male than female viewers. Some of these impacts also appeared to vary by different levels of education and family income. PMID- 10538433 TI - Sexual harassment: an ominous liability for healthcare professionals. AB - Sexual harassment can arise in virtually any employment setting. Healthcare facilities must be careful to avoid sexual harassment of their employees both by other employees and by the myriad other persons who may come into contact with their employees in the workplace. Much of what makes sexual harassment an actionable offense are the perceptions of the victim. Employers should take a proactive approach to preventing, detecting and correcting instances of harassment. PMID- 10538434 TI - Genetic testing: do healthcare professionals have a duty to tell a patient's family members that they may be at risk? AB - The author describes the healthcare professional's dilemma of keeping a patient's genetic information confidential. Provided are case law, commentary and policy developments addressing confidentiality of patient information as weighed against third parties' needs to know genetic information regarding their family members. PMID- 10538435 TI - Two sides of a coin: an interview with two attorneys (reprinted from 1986). Interview by Steven MacLauchlan. PMID- 10538436 TI - Now that we own it, what do I do with it? Identifying risk management issues in acquired physician practices. AB - Healthcare risk managers now have responsibility for identifying and managing the unique exposures of the physician's office practice. This article focuses on exposures related to general communication systems and functions including telephone routing systems, voice mail, faxing, e-mail, and more, which can be found in the clinic environment. PMID- 10538437 TI - Risk management and pollution prevention: thinking outside the box. AB - The Medical University of South Carolina's risk management department received a 12-month grant from South Carolina's General Assembly "Hazardous Waste Management Research Fund" for the purpose of identifying pollution prevention opportunities within South Carolina healthcare facilities. One hundred and one healthcare facilities were invited to participate in this study. Facilities ranged in size from zero-licensed beds (clinics) to more than 300 beds. Seventy-six South Carolina facilities (75%) participated in this project. PMID- 10538438 TI - False memory cases yield mixed results. PMID- 10538439 TI - Damages under EMTALA limited by MICRA (California Medical Injury Compensation Reform Act) cap in California. Barris v. County of Los Angeles. PMID- 10538440 TI - Records of similar occurrences may be discoverable in Alabama. McCullough v. Dalraida Health Center. PMID- 10538441 TI - "Presumptive" coding results in false claims. U.S. ex rel. Trim v. McKean. PMID- 10538442 TI - Production of medical records allowed without patients' consent. U.S. ex rel. Ghaprial v. Quorum Health Resources. PMID- 10538443 TI - Who will swallow Medicare's bitter pills? PMID- 10538444 TI - Unionizing the E.R. PMID- 10538446 TI - My summer scare. One doctor, reading my breast X ray, ordered a surgical biopsy. I panicked--and I'm glad I did. PMID- 10538445 TI - Margarine misgivings. PMID- 10538447 TI - Senators acknowledge SNF cuts too much, likely action to restore funding in July. PMID- 10538448 TI - Group seeks additional clarification of medical review guidelines. PMID- 10538449 TI - Impact of PPS on publicly traded nursing facility chains called disastrous. PMID- 10538450 TI - Quarterly financial results of post acute/subacute companies. PMID- 10538451 TI - Variations in and correlates of length of stay in academic hospitals among patients with heart failure resulting from systolic dysfunction. AB - OBJECTIVE: Given the high cost of caring for patients with congestive heart failure, there are strong incentives to decrease hospital costs by shortening length of hospital stay. We sought to identify factors associated with length of stay among patients admitted for the treatment of heart failure resulting from systolic dysfunction. STUDY DESIGN: Retrospective cohort study. METHODS: We examined data from patients with a principal discharge diagnosis of congestive heart failure who had been admitted to 1 of the 49 academic hospitals across the United States that participated in the CHF Benchmark Project, a large collaborative quality improvement project coordinated by the University HealthSystem Consortium. Patients were discharged between January 1 and June 30, 1996. We obtained patient characteristics and hospitalization data by retrospectively reviewing medical records. We used linear regression models to identify major determinants of length of stay. RESULTS: Among the 1046 patients eligible for the study, 59% were women, 55% were white, and 58% were aged 65 years or older. Adjusting for patient demographic and admission clinical characteristics, the mean length of stay was 4.9 +/- 0.9 days. Length of stay varied significantly among hospitals, even after adjusting for differences in patient characteristics. In multivariate regression models, factors that were independently associated with a significantly longer length of stay were prior renal failure, peripheral edema, atrial fibrillation, hyponatremia, urinary catheter on admission, initiation of an antiarrhythmic or warfarin, and major complications. Patient characteristics and hospital events combined explained 16% of the variation in the length of stay. Adjusting for the individual hospitals explained an additional 10% of the variation in the length of stay. CONCLUSIONS: Although a number of patient and hospitalization factors were associated with length of stay in patients with congestive heart failure resulting from systolic dysfunction, much unexplained variation remained. Clinical factors alone explained about 50% more variation than did factors specific to the individual hospitals. PMID- 10538452 TI - Continuity of care: is it cost effective? AB - OBJECTIVE: To examine the association between the degree of healthcare provider continuity and healthcare utilization and costs. STUDY DESIGN: A longitudinal, prospective, observational study. PATIENTS AND METHODS: Data on patients with arthritis, asthma, epigastric pain/peptic ulcer disease, hypertension, and otitis media were collected at each of 6 health maintenance organizations (HMOs). Outcome variables included the number of prescriptions for the target disease and the cost, total number of prescriptions and the cost, the number of outpatient visits, and the number of hospital admissions. Disease-specific severity of illness, type of visit, and provider information were obtained at each encounter. HMO profit status, visit copay, gatekeeper strictness, formulary limitations, use of multisource (generic) drugs, gender, number of months in the study, age, and severity of illness were controlled in the analyses. RESULTS: There were 12,997 patients followed for more than 99,000 outpatient visits, 1000 hospitalizations, and more than 240,000 prescriptions. Increasing the number of primary or specialty care providers a patient encountered during the study generally was associated with increased utilization and costs when HMO and patient characteristics were controlled. The number of specialty care providers also increased as the number of primary care providers increased. The incremental increase in pharmacy costs per patient per year with each additional provider ranged between $19 in subjects with otitis media to $58 in subjects with hypertension. CONCLUSIONS: Continuity of care was associated with a reduction in resource utilization and costs. As healthcare delivery systems are designed, care continuity should be promoted. PMID- 10538453 TI - Nursing facilities and managed care. AB - OBJECTIVE: To examine the extent to which Illinois nursing facilities have developed relationships with other healthcare providers, particularly managed care organizations (MCOs). STUDY DESIGN: A cross-sectional survey of nursing facilities designed to determine: 1) relationship objectives; 2) obstacles to developing relationships; 3) currently available services; 4) staffing for these services and; 5) nursing facility approaches to networking. The survey was sent to a census sample of 867 nursing facilities serving the elderly in Illinois. Descriptive and multivariate logistic regression analyses of relationships determined to be formal/risk-sharing were performed. STUDY POPULATION: The sample included 523 Illinois nursing facilities. A total response rate of 60% was achieved (523/867). RESULTS: Higher strategic goals, urban location, nonprofit ownership status, higher percentages of private pay and/or Medicare clients (vs Medicaid), and provision of home care and subacute services were all significant predictors of formal or risk-sharing relationships with MCOs. CONCLUSIONS: Facilities with more relationships and higher goals have more formal/risk-sharing relationships with MCOs. Facilities in urban areas have more relationships, likely due to the fact that rural facilities have fewer options and operate in different markets. In addition, nursing facilities rely on Medicare referrals from hospitals, and these Medicare patients, especially those in urban areas, are increasingly controlled by MCOs. PMID- 10538454 TI - The role of respiratory care practitioners in a changing healthcare system: emerging areas of clinical practice. AB - OBJECTIVE: To evaluate shifts in respiratory care practice in the context of changing healthcare system and market dynamics. STUDY DESIGN: Telephone survey, structured interview, and case studies. METHODS: We conducted a telephone survey of 471 respiratory care practitioners (RCPs), drawn from the membership database of the American Association for Respiratory Care. We also interviewed 10 employers of RCPs and conducted 2 in-depth case studies to supplement our survey results. We used several statistical techniques to analyze our data, including calculation of population-weighted descriptive statistics and multivariate regression models. RESULTS: Changes in the healthcare system have prompted RCPs to broaden their practice settings, skills, and responsibilities. Respiratory care practitioners are taking part in managed care-related activities, such as cost control and disease management. We found that the need for certain skills and responsibilities varies by practice setting. In our interviews, employers considered RCPs cost effective providers for certain services. CONCLUSIONS: The practice of respiratory care is evolving to meet the changing needs of the healthcare system. A key challenge is to ensure appropriate growth and development of the respiratory care profession, as well as the delivery of appropriate services under new care management settings and processes. PMID- 10538455 TI - The impact of TennCare on patient satisfaction with care. AB - OBJECTIVE: To measure the level of satisfaction with care by Medicaid-eligible patients before and after implementation of a mandatory managed care plan known as TennCare. STUDY DESIGN: We used multivariate logit analysis of survey data to calculate the effects of TennCare on patient satisfaction for TennCare patients compared to those on traditional Medicaid, using North Carolina as a control state. PATIENTS AND METHODS: Patients were respondents to a survey conducted in late 1996 and early 1997 who had been admitted to hospitals in 1993 and 1995 for labor/delivery (n = 986), acute myocardial infarction (n = 457), and head trauma (n = 248). Dependent variables were yes/no responses to satisfaction questions for labor/delivery and 5-category ordered responses for adults. RESULTS: We found no statistically significant differences in satisfaction between TennCare and traditional Medicaid for either pediatric or adult hospital patients. Generally, TennCare recipients had satisfaction levels as good or better than traditional Medicaid recipients. For pediatric care, TennCare odds ratios ranged from 1.00 to 2.17, the latter for satisfaction with care received (P = 0.107). For adult care, odds ratios ranged from 0.77 to 1.23, the latter for satisfaction with cost of care (P = 0.547). For many dimensions of care, lower rates of satisfaction were reported for respondents who were uninsured, less educated, and in poor health. For adult care, blacks or Hispanics tended to be less satisfied with some aspects of care. CONCLUSION: TennCare did not reduce patient satisfaction with care among those who were hospitalized. PMID- 10538456 TI - Continuation of postmenopausal hormone replacement therapy in a large health maintenance organization: transdermal matrix patch versus oral estrogen therapy. AB - OBJECTIVE: To determine possible differences in continuation of postmenopausal estrogen replacement therapy among women initiating treatment with transdermal estradiol versus those initiating treatment with oral estrogen. STUDY DESIGN: A retrospective database search. PATIENTS AND METHODS: We analyzed estrogen use among 45- to 74-year-old women who filled index prescriptions for estrogen during 1996 for either once-a-week transdermal estradiol or daily oral estrogen. Prescription use was analyzed separately for each of 2 groups: 276 hysterectomized women who filled prescriptions for estrogen alone (ERT) and 4182 women who filled prescriptions for medroxyprogesterone acetate (MPA) with estrogen (HRT) on the same day. RESULTS: Risk of discontinuing therapy after 12 months ranged from 59% to 76% among the 4 subgroups: ERT with unopposed transdermal estradiol; ERT with unopposed oral estrogen; HRT with MPA-opposed transdermal estradiol; and HRT with MPA-opposed oral estrogen. The relative risk (RR) of discontinuation was significantly greater among women starting HRT with transdermal estradiol than among women starting oral estrogen (RR = 1.5; 95% confidence interval [CI] = 1.3 to 1.8). RR of discontinuation among women starting ERT with transdermal estradiol compared with women starting oral estrogen therapy was 1.3 (95% CI = 1.0 to 1.8). CONCLUSIONS: Approximately 2 of 3 women who start either ERT or HRT discontinue therapy within a year, regardless of hysterectomy status. Furthermore, women who start ERT or HRT with a transdermal estradiol system are more likely to discontinue therapy. PMID- 10538457 TI - Variation in length of stay in patients hospitalized with congestive heart failure. PMID- 10538459 TI - Stats & facts. Disenrollment rates: issues and concerns. PMID- 10538458 TI - Gauging individual patient needs versus those of the general population. PMID- 10538460 TI - Rheumatoid arthritis. PMID- 10538461 TI - A conversation with William T. McGivney, PhD. PMID- 10538462 TI - Internet pharmacies: opportunities and challenges. PMID- 10538463 TI - Faith or evidence: what is the purpose of disease state management? PMID- 10538464 TI - Outcomes of an educational component of a disease management program for hypertension. AB - Little data exist regarding educational efforts in disease state management in mixed-model health plans. The following article presents results and a discussion on the difficulties involved in measuring the effectiveness of an educational initiative for patients with moderate-to-severe hypertension. PMID- 10538465 TI - Provider-sponsored organizations: the state of the market. AB - The Balanced Budget Act of 1997 introduced the Medicare+Choice program, which expanded the types of organizations that could assume Medicare prepaid capitated risk. Among the many new options, the provider-sponsored organization (PSO) received the most attention, although there have been few PSO applications filed. Organizations considering the PSO opportunity can gauge their readiness by evaluating the ease with which they can adopt key aspects of successful HMOs. PMID- 10538466 TI - Physician organizing: a convergence of interests. AB - Economic forces and professional ideals are colliding to create further unrest in the health care industry in the form of organizing physicians into unions. Whereas self-employed or independent contractor physicians are barred from joining together to bargain collectively with third-party payers and HMOs, employed physicians may be able to avoid antitrust problems because they are eligible for protection under the National Labor Relations Act. Consequently, as the number of employed physicians has grown and the frustrations they encounter increase, unions (generally suffering a decrease in membership and political influence) are finding a receptive audience for their message of solidarity. However, will organizing into unions solve or aggravate the physicians' problems? PMID- 10538467 TI - Call centers held liable for 'medical services' provided. AB - A new insurance plan for disease management services is now available for health plans and vendors. Find out why it's important protection to have, what it covers, and how much it will cost. PMID- 10538468 TI - Slash rate of premature birth with early recognition and management of preterm labor. AB - Following clinical guidelines and incorporating these early warning signs of preterm labor into your prenatal treatment and education programs can significantly cut the rate of prematurity among your members. PMID- 10538469 TI - Engaging Medicaid populations in DM initiatives requires patience, labor and resources. AB - Slow to catch on, but great players when they finally do. That's how DM providers who work with Medicaid patients view their members. See how these programs have garnered better compliance and improved outcomes. PMID- 10538470 TI - Get to root cause of allergy symptoms and reduce the high cost of prescriptions. AB - Nothing to sneeze at: High pharmacy costs for antihistamines may be just the wrong prescription for patients and your health plan budget. Find out how new guidelines can help you manage allergies cost effectively, reduce costly referrals, and slash allergy-related drug costs. PMID- 10538471 TI - Get aggressive with early diagnosis, treatment of arthritis. PMID- 10538472 TI - Quality of telephone triage nursing comes under fire. AB - The quality and consistency of telephone triage services is increasingly coming under fire, fueled by a new study revealing inconsistent use of protocols. Here's what you can do to ensure your triage services are top notch. PMID- 10538473 TI - Hospital's case management program pays off with dramatic drop in utilization. AB - The administrators at this Ohio hospital are trying to keep people out of their facility, and they are not ashamed to say so. In fact, in a communitywide effort acclaimed by the American Hospital Association, the facility found a way to decrease its soaring readmission rates, and the community could not be happier. PMID- 10538474 TI - Lyme disease vaccine available for patients at high risk. PMID- 10538475 TI - Patients listen better when MDs give nutritional counseling. PMID- 10538476 TI - Diffusion of laparoscopic cholecystectomy in the Veterans Affairs health care system, 1991-1995. AB - CONTEXT: Laparoscopic cholecystectomy has become the most widely used treatment for gallbladder disease. In HMO, Medicare, and fee-for-service settings, cholecystectomy rates increased 28% to 59% after introduction of laparoscopic cholecystectomy. OBJECTIVE: To investigate the impact of the introduction of laparoscopic cholecystectomy on cholecystectomy rates and the operative mortality rate in Veterans Affairs (VA) hospitals. DESIGN: Sequential cross-sectional study. PATIENTS: All patients who underwent cholecystectomy from 1991 (before introduction of laparoscopic cholecystectomy) to 1995. SETTING: 133 VA hospitals. OUTCOME MEASURES: Cholecystectomy rates, use of laparoscopic or open cholecystectomy, and operative mortality rate. RESULTS: The annual number of cholecystectomies in the VA system increased by 10% from 1991 to 1995; the laparoscopic procedure accounted for 25% of the caseload in 1992 and 52% in 1995. Compared with patients having laparoscopic cholecystectomy, those having open cholecystectomy were more likely to be older, be male, and have acute cholecystitis or comorbid illnesses (P < 0.001). The operative mortality rate of open cholecystectomy increased by 46% during this 4-year period (from 2.4% to 3.4%) and was constant for laparoscopic cholecystectomy (about 0.5%). Given the increasing use of the laparoscopic procedure, however, the overall mortality rate of cholecystectomy during surgery decreased by 22% (from 2.4% to 1.8%). Despite increased use of the surgery, the absolute number of deaths decreased by 9%. CONCLUSIONS: The introduction of laparoscopic cholecystectomy in the VA system was not accompanied by a large increase in cholecystectomy rates, as it was in fee-for-service, Medicare, and HMO systems. Because the rate of operations has changed only slightly, the total number of cholecystectomy-related deaths has decreased. PMID- 10538477 TI - How well does a single question about health predict the financial health of Medicare managed care plans? AB - CONTEXT: Responses to simple questions that predict subsequent health care utilization are of interest to both capitated health plans and the payer. OBJECTIVE: To determine how responses to a single question about general health status predict subsequent health care expenditures. DESIGN: Participants in the 1992 Medicare Current Beneficiary Survey were asked the following question: "In general, compared to other people your age, would you say your health is: excellent, very good, good, fair or poor?" To obtain each participant's total Medicare expenditures and number of hospitalizations in the ensuing year, we linked the responses to this question with data from the 1993 Medicare Continuous History Survey. SAMPLE: Nationally representative sample of 8775 noninstitutionalized Medicare beneficiaries 65 years of age and older. MAIN OUTCOME MEASURES: Annual age- and sex-adjusted Medicare expenditures and hospitalization rates. RESULTS: Eighteen percent of the beneficiaries rated their health as excellent, 56% rated it as very good or good, 17% rated it as fair, and 7% rated it as poor. Medicare expenditures had a marked inverse relation to self assessed health ratings. In the year after assessment, age- and sex-adjusted annual expenditures varied fivefold, from $8743 for beneficiaries rating their health as poor to $1656 for beneficiaries rating their health as excellent. Hospitalization rates followed the same pattern: Respondents who rated their health as poor had 675 hospitalizations per 1000 beneficiaries per year compared with 136 per 1000 for those rating their health as excellent. CONCLUSIONS: The response to a single question about general health status strongly predicts subsequent health care utilization. Self-reports of fair or poor health identify a group of high-risk patients who may benefit from targeted interventions. Because the current Medicare capitation formula does not account for health status, health plans can maximize profits by disproportionately enrolling beneficiaries who judge their health to be good. However, they are at a competitive disadvantage if they enroll beneficiaries who view themselves as sick. PMID- 10538478 TI - Thriving in a busy practice: physician-patient communication training. AB - BACKGROUND: Despite growing concern about the potential impact of managed care on the physician-patient relationship, efforts to enhance the quality of communication between practicing clinicians and their patients have been limited. OBJECTIVE: To determine the effectiveness of a 1-day educational workshop. DESIGN: Clinician self-assessment of interviewing skills measured immediately before and 3 months after the workshop. SETTING: The Kaiser Permanente Medical Care Program. PARTICIPANTS: Practicing clinicians (n = 1384) in 22 workshops during a 5-year period. Nine hundred eleven participants (66% response rate) completed self-assessment questionnaires 3 months after the workshop. RESULTS: Self-assessed interviewing skills improved in all items 3 months after the workshop (P < 0.05). Clinicians also reported a decline in the proportion of visits that they characterized as frustrating. CONCLUSION: A 1-day educational intervention for large groups of practicing clinicians can improve confidence in medical interviewing skills and the ability to handle difficult encounters. PMID- 10538479 TI - Variation in the use of echocardiography. AB - CONTEXT: Geographic variation in population-based rates of invasive cardiac procedures has been described. However, little is known about variation in rates of noninvasive testing for cardiovascular disease. Echocardiography is the second most common cardiac diagnostic procedure. PRACTICE PATTERN EXAMINED: Population based rates of echocardiography, adjusted for age, sex, and race, in the United States. DATA SOURCE: 5% sample of Medicare Part B. RESULTS: 1 in 10 Medicare beneficiaries underwent echocardiography in 1995. Rates of echocardiography varied by state from 5% in Oregon to 15% in Michigan. Rates tended to be lowest in the Northern Great Plains, the Pacific Northwest, and the Rocky Mountains states. Among the 25 largest metropolitan areas, substantial variation was also apparent. For example, one fourth of Medicare beneficiaries in Miami, Florida, received echocardiography, and this proportion was more than four times greater than that seen in Seattle, Washington. CONCLUSION: The likelihood of Medicare beneficiaries having echocardiography is influenced by where they live. PMID- 10538480 TI - Changing disease definitions: implications for disease prevalence. Analysis of the Third National Health and Nutrition Examination Survey, 1988-1994. AB - CONTEXT: In the hope of extending treatment benefits to patients with early disease, various professional societies have recommended changing several common disease definitions by lowering the threshold value for diagnosis. COUNT: Number of Americans labeled "diseased" under new definitions for diabetes, hypertension, hypercholesterolemia, and being overweight. CALCULATION: [symbol: see text] DATA SOURCE: Adult participants (age > 17 years) in the Third National Health and Nutrition Examination Survey (1988-1994). RESULTS: Adopting the new definitions would dramatically inflate disease prevalence. Changing the threshold for diabetes from a fasting glucose level of > or = 140 mg/dL to > or = 126 mg/dL would result in 1.7 million new cases. Redefining hypertension as systolic blood pressure > or = 140 mm Hg instead of > or = 160 mm Hg or diastolic blood pressure > or = 90 mm Hg instead of > or = 100 mm Hg would create 13 million new hypertensive patients. For hypercholesterolemia (a cholesterol level of > or = 200 mg/dL instead of > or = 240 mg/dL) and being overweight (body mass index > or = 25 kg/m2 instead of > or = 27 kg/m2), the number of new cases would be 42 million and 29 million, respectively. The new definitions ultimately label 75% of the adult U.S. population as diseased. CONCLUSIONS: If these modest changes in disease definition were adopted, great numbers of people would be considered diseased. The extent to which new "patients" would ultimately benefit from early detection and treatment of these conditions is unknown. Whether they would experience important physical or psychological harm is an open question. PMID- 10538481 TI - Should this patient be screened for cancer? AB - CONTEXT: Advances in imaging technology have provided numerous opportunities for cancer screening but have also raised numerous questions. GENERAL QUESTION: Who should be screened and how exactly should screening be performed? SPECIFIC RESEARCH CHALLENGE: If spiral computed tomography (spiral CT) were being considered for lung cancer screening, for example, important questions would need to be answered: Should nonsmokers be screened? How often should screening take place? What should the diagnostic work-up be after abnormal findings were seen on spiral CT? STANDARD APPROACH: Randomized, controlled trials (RCTs) are the most valid method for determining which medical interventions are most effective. These trials are particularly useful in the evaluation of screening because they eliminate the early detection biases that may result in groosly misleading survival statistics. POTENTIAL DIFFICULTIES: Randomized, controlled trials of screening are subject to other biases, and their results may be difficult to generalize. In addition, because they require an enormous number of participants and many years of follow-up, RCTs can be applied only to a small proportion of the questions about cancer screening. ALTERNATE APPROACH: Quantitative decision analysis can be applied to the remaining questions and help inform decision making about cancer screening. PMID- 10538482 TI - Finding and redefining disease. PMID- 10538483 TI - Using the Internet as an effective customer channel. Creating sustainable market advantage through Web-based strategies. PMID- 10538484 TI - Five biggest challenges in marketing the integrated system: Part I. PMID- 10538485 TI - Hitting bad times, HMO strategists work on new entrees to the market. PMID- 10538486 TI - Pharmacies collaborate to track ADRs, medication use. PMID- 10538487 TI - Medical technology index helps hospitals spend money wisely. AB - New medical technology index isn't a crystal ball, but it's almost as good. If you're wondering what your health system needs--a strategic alliance, capital improvements, or a new surgical suite--this new index may be just what the doctor ordered. Find out how it can help you evaluate whether your hospital has the competitive edge. PMID- 10538488 TI - Rush to release physician report cards raises stakes in California market. AB - Provider report cards: Are they worth the paper they're printed on? Health plans and medical groups in California have begun releasing outcomes and service "report cards" in order to gain a competitive advantage in a crowded marketplace. Some observers charge that vagaries in reporting methodology are more likely to lead to confusion than enlightenment among patients and payers. However, a new initiative will develop standardized data analysis and reporting methods. PMID- 10538489 TI - 'Down and dirty' medical information system identifies high-risk patients. AB - Home-grown information system works for this PHO. Puyallup Valley Healthcare in Puyallup, WA, needed a way to identify patients who could benefit from case management, but the PHO didn't have a lot of money to spend on expensive information systems. See how the staff designed their own information system, combining data downloaded off the internet and the PHO's own software. PMID- 10538490 TI - Health care payments: how low can they go? AB - Data Library: A new national pricing database features Medicare fee benchmarking data to help payers and providers who are trying to set market-based fees for a variety of services. Find out how Salt Lake City-based Medicode devised this tool to help define the market value of health care services. PMID- 10538491 TI - Consumers rank HMOs high for quality of health care. AB - Health care consumers, physicians concur on HMO quality of care. In separate surveys HMO enrollees and primary care providers generally agreed that although they get a bad rap, HMOs really do provide good quality of care. PMID- 10538492 TI - Studying the effects of nurse prenatal and early infancy home visitations. PMID- 10538493 TI - Infection control in home care. A report by the Florida Hospital Association and Hospital Home Care Association of Florida's Infection Control Task Force. PMID- 10538494 TI - Influenza/pneumococcal vaccine study. A report by the Florida Hospital Association and Hospital Home Care Association of Florida's Infection Control Task Force, December 1998. PMID- 10538495 TI - Using readily available data to assist with asthma management. PMID- 10538496 TI - Case management of asthma in a primary care setting. PMID- 10538497 TI - Achieving success for freestanding ambulatory care centers. Start-up and initial operation. AB - This is the last in a series of three articles focusing on achieving success for freestanding ambulatory care centers. This article discusses start-up and initial operation of freestanding centers. The first article, published in the fall 1998 issue of Ambulatory Outreach, provided a framework for making the decision to invest in a freestanding ambulatory care center. The second article, published in the winter 1998 issue, discussed detailed planning for a center. The combined articles offer proven approaches for strategic decision-making, detailed planning and implementation of ambulatory care centers. PMID- 10538498 TI - Cost effective management of chronic illness. PMID- 10538499 TI - Operation clean break--a community health center response to gang related violence. PMID- 10538500 TI - Using OBQI (Outcome-Based Quality Improvement) to establish priorities for change. AB - OASIS and OBQI will touch every home care agency. Outcomes data, when properly used, may be of critical value to today's home care executives. Thus, it is important for agency executives to focus on positive ways to use the data to run their agencies more effectively. PMID- 10538501 TI - Six steps to smooth OASIS (Outcome Assessment and Information Set) implementation. AB - The Medicare Quality Improvement Demonstration Project has examined agencies' ability to integrate OASIS into standard admission, recertification, and discharge processes. One pilot agency succeeded in implementing OASIS smoothly and with very little increase in patient-admission time using a six-step strategic planning cycle and effective communication tools. PMID- 10538502 TI - Prat falls, missteps, & solutions in implementing change. AB - Agencies across the country are preparing for OASIS, PPS, and other changes. The Visiting Nurse Association of Northern California shares its story of preparing for changes while simultaneously shifting gears to achieve compliance with regulations amid competing priorities and financial pressures. PMID- 10538503 TI - Beyond OASIS (Outcome Assessment and Information Set). AB - Agencies that adopt the Outcome Based Quality Improvement (OBQI) methodology stand to make quantifable improvements in patient outcome. St. Mary's Home Care in Grand Junction, Colorado, successfully improved patient outcomes during its participation with the OBQI demonstration project. PMID- 10538504 TI - The accreditation program for home care aide services: an historical perspective. PMID- 10538505 TI - Volunteer program capitalizes on "neighborly" visits. AB - Turbulence and uncertainty mark today's health care environment. Health care providers must create new partnerships to deliver optimum care to their patients. Last year the staff of Forsyth Home Care and members of the Auxiliary of Forsyth Medical Center created one such partnership called the Neighbors Volunteer Program. PMID- 10538506 TI - OASIS (Outcome and Assessment Information Set). Will HCFA know more about your agency than you do? AB - OASIS data will provide a host of information to the Health Care Financing Administration. Agencies should look beyond HCFA's reasons for collecting the data and capitalize on the opportunity to use OASIS to improve their own services and better understand their patients' needs. PMID- 10538507 TI - Getting to the future first: a conversation with Gary Hamel. Interview by Joe Flower. PMID- 10538508 TI - Leadership and white space: the struggle for strategy innovation in health care. PMID- 10538509 TI - Knowledge management: moving the care model from a "snapshot" to a "story". PMID- 10538510 TI - The case for managing structural capital. PMID- 10538511 TI - Sharing knowledge and best practices: the hows and whys of tapping your organization's hidden reservoirs of knowledge. PMID- 10538512 TI - Web sites of tomorrow: how the Internet will transform healthcare. PMID- 10538513 TI - The health care profit pool: who stands to gain and lose in the digital economy? PMID- 10538514 TI - Informed shared decision making: is this the future of health care? PMID- 10538515 TI - The virtual team: strategies to optimize performance. PMID- 10538516 TI - A really balanced scorecard. PMID- 10538517 TI - The obsolescence of independent practice. Part 2: Inviting physicians back home- the hospital as a point of confluence. PMID- 10538518 TI - An innovation Rx. PMID- 10538520 TI - Not "I," but "We". PMID- 10538519 TI - Creature comforts. PMID- 10538521 TI - Junior partner woes. PMID- 10538522 TI - Fire-based EMS: the trend of the future? PMID- 10538524 TI - Surviving the interview process. PMID- 10538523 TI - Lifting the spirit of St. Louis. The St. Louis EMS Bureau is reborn through fire department merger. PMID- 10538525 TI - EMS incident management: traits and characteristics of the incident safety officer. PMID- 10538526 TI - An EMS approach to psychiatric emergencies. AB - Psychiatric patients present challenges not only to EMS, but in the ED as well. As we have tried to indicate in the case report, the presentation is not always clear-cut. There may not be a definite solution, and each case can be different. As an EMT or paramedic, how should you approach these issues? First, always make sure the scene is safe for you and your team. If you have doubts, contact law enforcement. With regard to patient care, err on the side of safety and what is in the best interest of the patient (which may include restraints). Follow established procedures and guidelines, and always document well. If you have any questions on scene, contact medical control, and document doing so. If there is a question, let the physician decide on competency issues. PMID- 10538527 TI - Critical care transport in Summit County. PMID- 10538528 TI - Is the IDS glass half empty or half full? AB - When the concept of integrated delivery began gaining popularity throughout the healthare field, some healthcare executies saw it as the solution for organizations struggling to contain rapidly rising costs. Others hoped that joining integrated delivery systems would help preserve smaller organizations that could not otherwise compete with larger providers in the same area. And many in healthcare thought that it had the potential to dramatically enhance quality of care and promote wellness. As integration becomes increasingly common, the practice is coming under heavy criticism from those who say it has not lived up to expectatations. At the same time, however, systems that are seeing success with integrated delivery are singing its praises. "The jury is still out on the effectiveness of integrated delivery systems," says Austin Ross, FACHE, a professor at the University of Washington's School of Public Health, Seattle. "So far, they have a spotty record. There have been some serious failures, but some positive things are taking place, too." Some experts say that integrated delivery systems are doomed; others believe there are good reasons to be optimistic. So is the glass half empty or half full? PMID- 10538529 TI - Outcomes and performance measurement: redefining how healthcare is strategized and delivered. AB - The prevalence of managed care, integrated delivery systems, and new financing structures is nudging healthcare into a value-driven rather than a cost controlling market, with a focus on community and improvement. Fueled by increasing demands from healthcare purchasers, consumers, accrediting bodies, and government agencies, providers are escalating their efforts to demonstrate that they offer the highest technical and service quality. PMID- 10538530 TI - High hospital CEO turnover trend continues. PMID- 10538531 TI - Assessing organizational viability. PMID- 10538532 TI - Linking with physician practices. PMID- 10538534 TI - Millennium stockpiling. PMID- 10538533 TI - Conducting internal service audits. PMID- 10538535 TI - Strategically evaluating your web site. PMID- 10538536 TI - Looking ahead to FY 2000. PMID- 10538537 TI - Integrated healthcare delivery. How are we shaping up? AB - For the past 10 years, healthcare organizations have been reshaping themselves- merging, acquiring, aligning, and partnering--positioning themselves to provide services across the continuum of care. But as the dust settles around these newly structured entities, some providers are wondering when they--and the communities they serve--will realize the full potential of integrated healthcare delivery. Healthcare experts point out that, although asset aquisition lays the foundation for integration, many challenges lie ahead. True clinical integration will be possible only when solid governance, management, and systems infrastructures are in place. PMID- 10538538 TI - Forecasting patient services: a 21st century vision of an academic health centre. AB - This article provides an overview of the development and implementation of the McGill University Health Centre model for forecasting patient services in the year 2004, and advice on how to apply the model. Critical success factors and case examples are highlighted. The insights provided will be of value to hospitals and other institutions that recognize the necessity of engaging in long range planning and forecasting. PMID- 10538539 TI - CONTINUUM-Activity Index: managing admission inappropriate stays in a community hospital. AB - The CONTINUUM-Activity Index was used as a concurrent tool to measure intensity of services delivered in an Acute Care Community Hospital. Applying these specific daily measures identified patients who did not meet admission appropriateness criteria on the first day of care or did not meet those criteria on two days subsequent to admission. These patients had a high probability that their entire stay would be inappropriate. Action was then taken to move the care process forward, resulting in a significant reduction in inappropriate hospital days. PMID- 10538540 TI - Team-based planning: new tools for new times. AB - The collaborative process of team-based planning draws upon the strengths of a team and develops the skills that healthcare providers need to build consensus during this time of significant change. This article describes three examples of team-based planning and shows how applying an integrated set of tools can help formulate plans and create new options to meet the challenges facing today's healthcare organizations. PMID- 10538541 TI - The creation of a profession leader role in an academic health sciences centre. AB - The introduction and administration of any new role in a dynamic changing environment requires strong leadership and specific, purposeful administrative strategies. This article describes the development of a professional leader role at one campus of a recently merged academic health sciences centre. Understanding the operational challenges may help other healthcare organizations that wish to introduce and refine professional leadership roles. PMID- 10538542 TI - Implementation of the resident assessment instrument: a Canadian experience. AB - As the population ages, the increasing number and complexity of needs of individuals requiring institutionalization will increase the demands for chronic care services. In this article, the authors describe the implementation process used to introduce interdisciplinary staff to the use of the Minimum Data Set (MDS) for assessment of both nursing home and chronic care residents. This screening tool assesses resident characteristics over a wide spectrum of dimensions. The assessment findings are then integrated into the clinical plan of care. PMID- 10538543 TI - Quality control (Part 2). Communication: implementation of the Resident Assessment Instrument (RAI): the benefits of a structured communication plan. AB - Following a government mandate to use the assessment portion of the Resident Assessment Instrument, S&WCHSC began to integrate the RAI into the care of its Aging Program residents. S&WCHSC have used this as an opportunity to provide a structure for assessment and care planning within the existing philosophy of Patient Focused Care. Implementation of the RAI required the development and implementation of a Communication Plan. An adaptation of a framework, "Effective Strategic Planning for Communications" provided guidelines for assessment of key stakeholders' communication needs. PMID- 10538544 TI - Leadership: the Winnipeg Community and Long-Term Care Authority. AB - The Winnipeg Community and Long Term Care Authority (WCA) was established in 1998 under the Regional Health Authorities Act of the Province of Manitoba. The WCA's role is to provide for the successful integration of Winnipeg's community-based healthcare delivery services through its three main portfolios: Community Care and Public Health, Home Care and Mental Health, and Long Term Care and Specialized Services. The WCA is dedicated to building a quality health future for Winnipeg. Various initiatives undertaken in the pursuit of quality are described. PMID- 10538546 TI - A review of the evaluation literature on health professions training and enrichment programs for minority/disadvantaged students. AB - This paper includes the authors' review of the outcome evaluation articles of federal and non-federal training programs to prepare minority/disadvantaged students for entry into health programs as managers and practitioners. It provides information from the senior author's 1987-1993 Health Careers Opportunity Program grant for comparison purposes. The paper makes an argument for the standardization of outcome variables so that cross-program comparisons may be made. Until now, the variables reported in the literature have been very disparate. This paper suggests which kind of training program may have the most chance for success. Finally, it provides information rarely found in the evaluation literature: success rates for different ethnic groups and costs for running such programs. These data are provided for Health Care Administration faculty who could use planning information for similar grant applications. PMID- 10538547 TI - Nurse executives: new roles, new opportunities. AB - As women have been nursing since the earliest days of recorded civilization, so nurses have been associated with health care since the earliest days of recorded medical history. Gender and function have been inextricably woven in ways that created a struggle for success within a male-dominated industry. Nurses, as women, have been undervalued as, until recently, their role in health care has been similarly undervalued. Changing realities in the health care environment have created an opportunity for women's unique skills and talents to be revalued in a way that offers new opportunities for nurses. Teamwork, global thinking, multitasking, creativity, and flexibility are characteristics that have assumed new importance in the marketplace. Nursing leaders possess these attributes, along with a strong clinical foundation that is integrated with knowledge of sound business principles. This combination now positions nurse executives to reach the highest levels of heath care administration. Critical to this achievement is the professional credibility obtained through education at the master's degree level in health care and nursing administration programs that provide the essential tools for professional success. New opportunities for nurse executives afford educators in health care and nursing administration similar opportunities to develop and market programs to this large group of health care professionals who are seeking graduate education in increasing numbers. PMID- 10538545 TI - Something strictly Nisga'a: British Columbia First Nation runs its own health system. Interview by Matthew D. Pavelich. PMID- 10538548 TI - A window on practice: developing "rounds" for health administration graduate students. PMID- 10538549 TI - Enhancing health administration competencies for physicians: an evaluation of medical school curricula and education for physicians. PMID- 10538550 TI - Complementary/alternative medicine for health care managers: a course design. PMID- 10538551 TI - Reduce the risk. Identify your service's risk for TB exposure and protect providers by utilizing personal protective equipment and other technologies. PMID- 10538552 TI - The mask. TB-preventive masks compared. PMID- 10538553 TI - Rules to live by. Official guidelines on preventing the spread of TB. U.S. Department of Labor/OSHA. PMID- 10538554 TI - TB by the numbers. The state of tuberculosis in the United States. PMID- 10538555 TI - Exposures in EMS. An EMS medical director's 3-step plan to protect providers from TB exposure. PMID- 10538556 TI - Triage toppers. PMID- 10538557 TI - Team EMS. Clarifying EMS roles at mass casualty incidents. PMID- 10538558 TI - Close encounters. Medical considerations for confined space operations. PMID- 10538559 TI - Kansas confined space rescue. A small department utilizes mutual aid to rescue a child trapped 16 feet below ground. PMID- 10538560 TI - Y2 worry. It's not too late to prepare for year 2000 computer problems. PMID- 10538561 TI - Multijurisdictional scenes: who's in charge? PMID- 10538562 TI - Medicare advisory panel takes shape; labs need clarification on billing trip fees. PMID- 10538563 TI - Answering your questions on dealing with a disgruntled employee and using call waiting in a POL. PMID- 10538564 TI - Easing the switch to medical necessity documentation. AB - The responsibility for educating clinicians on Medicare's new regulations for test coding and documentation has fallen to the nation's laboratories. See how this reference laboratory created its own tools to help them cope. PMID- 10538565 TI - Curbing overutilization: the silver lining to HCFA compliance. PMID- 10538567 TI - Performance appraisals: more than just going through the motions. PMID- 10538566 TI - Stopping the spread of vancomycin-resistant enterococci. PMID- 10538569 TI - Former Healthtrust exec to start firm. Essent Healthcare to target small community hospitals. PMID- 10538568 TI - The new technology for cervical cancer screening: costs versus reimbursement. PMID- 10538570 TI - Tenet to sell at least 18 hospitals. PMID- 10538571 TI - Flexibility at a price. State-run medical university gets some relief. PMID- 10538572 TI - AMA loses millions. 1% membership decline and Y2K expenses blamed. PMID- 10538573 TI - Peer-review records can be subpoenaed. PMID- 10538574 TI - Market deflates for provider-owned HMOs. Survey shows many plans are losing ground to growing national and regional players. AB - The market for provider-owned HMOs continues to deflate as the health plans lose ground to their rising national and regional counterparts. That's among the findings of Modern Healthcare's annual HMO/PPO survey. But analysts and executives say provider-owned plans can make a go of it if they find the right niche. PMID- 10538575 TI - Reformer, transformer. L.A. County's Finucane dodges politics as he leads system serving nation's most populous county. PMID- 10538576 TI - Rivals join forces on vascular centers. PMID- 10538577 TI - Infection control pays off. Illness-prevention plan at BJC Health System saves $5.8 million in patient treatment costs. PMID- 10538578 TI - Investors go with HealthStream's flow. On-line education company in Nashville attracts some of the biggest names in healthcare. PMID- 10538579 TI - Let workers make the insurance choices. PMID- 10538580 TI - Falling behind in the payment game. Providers are feeling squeezed by HMOs that don't pay on time and keep changing the rules. AB - When hospitals provide services, they naturally expect to be paid. But providers are waiting longer and longer for payment from HMOs, sometimes 90 to 120 days, putting the squeeze on cash flow. The problem has led states to enact prompt payment laws. Health plans defend their record, saying hospitals' billing mistakes and antiquated computer systems are often to blame for the tardy payments. PMID- 10538581 TI - Running at capacity. Some hospitals have more patients than they can handle, but that high occupancy can be costly. PMID- 10538582 TI - Healthcare is looking good in Salt Lake City. Hospitals, docs are efficient, residents are healthy--for now. PMID- 10538583 TI - Alta builds on past to revive L.A. hospitals. PMID- 10538584 TI - Welfare recipients go to work in healthcare. PMID- 10538585 TI - Report: assisted-living prices are at premium. PMID- 10538586 TI - Catalyst for change. Premier alliance to invest millions in database to improve quality of care. PMID- 10538587 TI - FPA asset sales leave creditors in lurch. PMID- 10538588 TI - Data security regulations on the way. PMID- 10538589 TI - How it's seniors vying with kids for dollars. PMID- 10538590 TI - Medicare pain: who pays? Latest lobbying threatens cuts in care for women and kids. PMID- 10538591 TI - Back to the old model. Buyer of MedPartners' Calif. operations seen as returning medical groups to docs' control. PMID- 10538592 TI - The big gets bigger. Houston's Texas Medical Center announces a $1.5 billion development plan. PMID- 10538593 TI - Feds pave way for AHERF deal. Proposed purchase of four remaining AHERF hospitals still faces other regulatory hurdles. PMID- 10538594 TI - HealthSouth spinoff unleashes growth. PMID- 10538595 TI - Fighting for control. Hospitals and their outside authorities sue each other over who is in charge of what. PMID- 10538596 TI - Software made easy. Firms poised to use Internet to bring basic computing up to speed at nonhospital locations. PMID- 10538597 TI - Tug of war over prescription drug benefit. PMID- 10538598 TI - Medicaid HMOs exit markets. Shrinking pool of managed-care plans could mean lower reimbursement to provider. PMID- 10538599 TI - Sit up, take notice. Justice Department official chides healthcare industry for lax attention span for fraud, abuse. PMID- 10538600 TI - Suit intensifies syndication scrutiny. PMID- 10538601 TI - Feds probe hospitals with association heads. PMID- 10538602 TI - Is this the mob or healthcare? K.C. fraud has executives bemoaning prosecutors' rough-and-tumble tactics. PMID- 10538603 TI - One hospital treated kindly by fed probers. PMID- 10538604 TI - Not on the same page. Consumers Union, Calif. attorney general split on methodology in charity-care report. PMID- 10538605 TI - JCAHO may revamp basic survey process. PMID- 10538606 TI - 'Bad luck' plagues Vista system [ews]. PMID- 10538607 TI - Group Health to drop coverage for 21,000. PMID- 10538608 TI - Docs push for HMO bargaining rights. PMID- 10538609 TI - Texas gives limited antitrust relief to docs. Critics say new collective bargaining law goes too far; some docs say it doesn't go far enough. PMID- 10538610 TI - Clock ticks for Aetna to sell Texas plans. PMID- 10538611 TI - AMA to wear union label. House of Delegates' decision pits docs against hospitals. PMID- 10538612 TI - Pitts' surprise. Mariner CEO's hasty exit seen as sign that Medicare isn't the only issue in post-acute care. PMID- 10538613 TI - Executive shake-up. McKesson HBOC fires four top officers in IT unit. PMID- 10538614 TI - Learning how to get along. In fits and starts, physician groups beginning to cooperate with fraud prosecutors. PMID- 10538615 TI - ANA to form collective-bargaining unit. PMID- 10538616 TI - Hospital groups right doc antitrust relief. PMID- 10538617 TI - On the ropes. Beefed-up anti-kickback laws, growing cohort of whistleblowers pound away at healthcare fraud. AB - The federal government, armed with more fraud-fighting personnel and bigger budgets, is combining beefed-up kickback laws with the Civil War-era False Claims Act to hit providers with a one-two legal punch. Increasingly, providers are finding their only option is to pay big settlements. PMID- 10538618 TI - Healthcare price clubs get popular. Prepaid referral programs bring significant discounts to uninsured, but lack preventive care. PMID- 10538619 TI - Plugging the holes in outpatient accounts. Hospitals work to expedite payer reimbursement by getting authorization, information upfront. PMID- 10538620 TI - Rural-relief bills win partial AHA blocking. PMID- 10538622 TI - Gain-sharing illegal. HHS: docs shouldn't be paid to limit care. PMID- 10538621 TI - Exodus, not boycott. Simultaneous Medicare pullouts are just a coincidence, not collusion, HMOs say. PMID- 10538624 TI - Bon Secours plan to move Va. hospital. PMID- 10538625 TI - Will healthcare do time for convictions? Hospital groups seek Medicare relief as industry anticipates impact of latest fraud verdicts. AB - Less than a week after two hospital executives were found guilty of Medicare fraud, hospital groups last week launched an advertising blitz aimed at securing more Medicare money. But that could be a hard sell. Its success will hinge on how much the industry's image has been damaged by the recent convictions in Florida, which came only a few months after convictions in a Kansas City kickback case. PMID- 10538623 TI - Allina changes tune in new-hospital bid. PMID- 10538626 TI - HIV treatment data soon to be on Web. PMID- 10538627 TI - Going, going, gone. New American to sell last Iowa for-profit hospital; Davenport location not rural enough. PMID- 10538628 TI - Providers fear paying for drug benefit. PMID- 10538629 TI - Holding the line (for you down there). Salaries flatlining for many, while CEOs see 7.4% increase in median total compensation. PMID- 10538630 TI - Battle is brewing over HMO premium dollars. PMID- 10538631 TI - Preventive medicine. Computerized system analyzes whether dosage, timing and type of drug are right for the patient. PMID- 10538632 TI - Bond issuance cools after scorcher of '98. PMID- 10538633 TI - The other shoe drops. While industry fights budget curbs, even some profitable hospitals are raising prices. PMID- 10538635 TI - Physicians unite! Doctors aspire to become trade unionists. PMID- 10538634 TI - Relief gestures fail to charm providers. PMID- 10538636 TI - Rx: plenty of bed rest and a gold microchip. Engineers tackle new ways to take medicine. PMID- 10538637 TI - Kid wrists at risk. Computer injuries can start at home or in the classroom. PMID- 10538638 TI - The effectiveness of a point-of-decision prompt in deterring sedentary behavior. AB - Individuals using a university library were observed using stairs or elevators to travel up one to three floors. After a baseline assessment, a sign was posted by the elevator to discourage elevator use over a five-week period. A hierarchical log linear analysis indicated that stair use increased significantly after the intervention moderated by day of the week, sex, and age. The study suggests that point-of-decision prompts to deter sedentary activity may be effective. PMID- 10538639 TI - Carrots and sticks: impact of an incentive/disincentive employee flexible credit benefit plan on health status and medical costs. AB - PURPOSE: Employee wellness programs aim to assist in controlling employer costs by improving the health status and fitness of employees, potentially increasing productivity, decreasing absenteeism, and reducing medical claims. Most such programs offer no disincentive for nonparticipation. We evaluated an incentive/disincentive program initiated by a large teaching hospital in western Michigan. METHODS: The HealthPlus Health Quotient program is an incentive/disincentive approach to health promotion. The employer's contribution to the cafeteria plan benefit package is adjusted based on results of an annual appraisal of serum cholesterol, blood pressure, tobacco use, body fat, physical fitness, motor vehicle safety, nutrition, and alcohol consumption. The adjustment (health quotient [HQ]) can range from -$25 to +$25 per pay period. We examined whether appraised health improved between 1993 and 1996 and whether the HQ predicted medical claims. RESULTS: Mean HQ increased slightly (+$0.47 per pay period in 1993 to +$0.89 per pay period in 1996). Individuals with HQs of less than -$10 per pay period incurred approximately twice the medical claims of the other groups (test for linear trend, p = .003). After adjustment, medical claims of employees in the worst category (HQ < -$10 per pay period) were $1078 (95% confidence interval $429-$1728) greater than those for the neutral (HQ between $2 and +$2 per pay period) category. A decrease in HQ of at least $6 per pay period from 1993 to 1995 was associated with $956 (95% confidence interval $264 $1647) greater costs in 1996 than was a stable HQ. CONCLUSIONS: The HealthPlus Health Quotient program is starting to yield benefits. Most employees are impacted minimally, but savings are accruing to the employer from reductions in medical claims paid and in days lost to illness and disability. PMID- 10538640 TI - Factors associated with attendance in a voluntary nutrition education program. AB - PURPOSE: This paper examines factors associated with attendance in a National Cancer Institute-funded randomized trial of nutrition education to increase fruit and vegetable consumption among women served by the Special Supplemental Nutrition Program for Women, Infants, and Children (WIC). SETTING: The study took place at 16 WIC sites in Maryland. SUBJECTS: The participants were 1528 women who were enrolled in WIC or who had children enrolled in WIC, were > or = 18 years of age, and planned to continue enrollment at that WIC site for at least 6 months (68% of eligible women approached agreed to participate). INTERVENTION: Women received personal invitations, letters, and telephone reminders from peer educators encouraging their attendance at three bimonthly nutrition sessions. MEASURES: Demographic data were collected in a baseline survey. Attendance data and telephone and address changes were also collected. The postintervention survey included a question regarding reasons for nonattendance. Focus groups were also held to ascertain reasons for attendance or nonattendance. Chi-square tests of trend and multiple logistic regression, adjusted for within-site correlation, were used in statistical analyses. RESULTS: Fifty-four percent of enrollees attended at least one session. Multiple logistic regression analysis showed increased odds of attending with higher age, breast-feeding, and/or knowledge of the recommendation to eat five or more servings of fruits and vegetables daily. There were decreased odds of attending for pregnant women who already had children. There were nonsignificant trends toward decreased attendance among unmarried women compared with married women and among blacks compared with nonblacks. Reasons given for nonattendance included withdrawal from WIC, moving, conflicting activities, negative feelings about nutrition education, and lack of transportation or child care. CONCLUSIONS: The results suggest that numerous barriers hinder participation in nutrition programs aimed at low-income women. These barriers should be considered by health care professionals when planning intervention programs. Overcoming these barriers presents a major challenge. PMID- 10538641 TI - Participation, retention, and adherence: implications for health promotion research and practice. PMID- 10538643 TI - Evaluation of a skin cancer prevention module for nurses: change in knowledge, self-efficacy, and attitudes. AB - PURPOSE: The purpose of this study was to evaluate the effectiveness of a 1-week didactic and clinical skin cancer prevention training module. The evaluation assessed both the immediate and the 3-month effects of the module on nurse participants. In addition, this study assessed whether the module had any secondary effects on skin cancer practices, including perceived support from colleagues, resources, time, and perceived responsibility to conduct skin cancer screening activities and education. METHODS: A quasi-experimental design with 32 intervention and 87 comparison subjects was employed. Instruments developed and validated specifically for this study were used to assess knowledge, self efficacy, priority of skin cancer, and organizational level constructs. RESULTS: The findings indicate that the module significantly increased general and prevention knowledge as well as screening ability; the increase was stable over time. The module was also found to improve self-efficacy to screen and to educate. There was no effect on the organizational-level constructs. CONCLUSIONS: Baseline knowledge assessments validated other studies indicating that nurses need more education about skin cancer. Despite promising results from program participants, system-level barriers could impose substantial barriers to implementation in health care practice. Knowledgeable nurses must educate their colleagues, their supervisors, and the public about the priority of skin cancer screening and develop strategies for creating organizational change to increase the likelihood that screening and patient education will occur for people at risk for skin cancer. PMID- 10538642 TI - The impact of California's smoking ordinances on worksite smoking policy and exposure to environmental tobacco smoke. AB - A subsample of 5776 respondents to the California Tobacco Survey who do not smoke and work indoors outside of their home was analyzed regarding worksite smoking policy and worksite exposure to environmental tobacco smoke. To obtain study results, survey responses were linked to tobacco ordinance data. Nonsmokers who worked in localities with moderate or strong laws were more likely to report worksite smoking policies than nonsmokers in localities without laws. Even in localities with strong laws, 23.5% of respondents reported no worksite policy, and 26.4% reported recent exposure to environmental tobacco smoke at the worksite. Comprehensive laws with minimal exemptions may be necessary to ensure adequate compliance and protection from environmental tobacco smoke. PMID- 10538644 TI - Self-efficacy and consumption of fruit and vegetables: validation of a summated scale. AB - PURPOSE: To develop and validate a scale to assess self-efficacy for increasing fruit and vegetable consumption, and to assess the ability of the scale to discriminate individuals at different stages of readiness to change. METHODS: Data were collected using a combination of self-administered mail questionnaires and phone interviews from a sample of 1200 Chinese Singaporeans randomly selected from residential phone listings. Principal-components analysis was conducted with half the sample, and model fit was measured using structural modeling procedures on the other half. Analyses of variance were used to determine whether self efficacy differed across the stages of change. SETTING: Data were collected as part of a larger study investigating factors influencing consumption of fruit, vegetables, and cereal foods. MEASURES: Fruit and vegetable intake was measured using a validated seven-item food frequency questionnaire. Subjects were assigned to stages using a phone-administered staging algorithm. Self-efficacy items were scored on a five-point Likert scale from very confident to not at all confident. RESULTS: Principal-components analysis revealed a two-factor structure that was highly stable across two split-half samples and gender, and accounted for 57% of the variance in self-efficacy. The two factors demonstrated good internal consistency (Cronbach's alpha = 0.77 and 0.89), with loadings ranging from 0.59 to 0.86 (mean = 0.70). Confirmatory factor analysis demonstrated good model fit (goodness-of-fit index = 0.99), with all parameters significant. Scores on the scale were significantly higher among subjects assigned to maintenance than among those assigned to precontemplation, contemplation, and preparation. CONCLUSIONS: Results of this study provide preliminary evidence for the utility of the scale to guide development and monitoring of community programs and therapeutic interventions. PMID- 10538646 TI - Ethics and AIDS drugs. PMID- 10538647 TI - Change of heart. A mitral-valve problem isn't as common--or as deadly--as your doctor might have told you. PMID- 10538645 TI - Acceptability of computer assessments among ethnically diverse, low-income smokers. AB - PURPOSE: To examine the acceptability of computer-based assessments among an ethnically diverse, low-income population of primary care patients. Although computers have been used to provide assessments and interventions in health care settings, members of ethnic minority and low-income households have less access to computers than other groups, and therefore the acceptability of computers as a health care assessment and delivery tool needs to be examined. DESIGN: We examined the acceptability of computers for providing assessments of smoking history, nicotine dependence, and other related variables among an ethnically diverse, low-income primary care population. No intervention was used in this study. SETTING: Three inner-city primary care clinics located in hospitals were used as sites for this study. These hospitals were located in areas of the city where low-income and ethnic minority households are overrepresented relative to the total population. SUBJECTS: Adult male and female smokers (n = 522) were recruited while awaiting appointments in each primary care clinic. MEASURES: A questionnaire assessing smoking rate, patterns, history, motivation to quit smoking, and other smoking-related variables was administered using either a paper-and-pencil format or a laptop computer. RESULTS: Frequency counts, analysis of variance, and chi 2 tests were used where appropriate. Most subjects (78.5%) used the computer to complete the baseline survey. Almost all subjects (92%) rated the computer "very easy" or "easy" to use. Subjects who were Spanish speaking, were born outside the United States, or were Hispanic tended to rate the program as slightly less easy to use than other subjects. CONCLUSIONS: Computer-based assessments appear highly acceptable to individuals in low-income populations. PMID- 10538648 TI - No money, no meds. PMID- 10538650 TI - Pharmacy computer systems--best-of-breed or one-vendor solutions? PMID- 10538649 TI - Wireless hospitals: new wave in healthcare technology. PMID- 10538652 TI - Successful steps to enterprisewide integration. PMID- 10538651 TI - Easy steps to point-of-care computing. PMID- 10538653 TI - How to approach IT outsourcing. PMID- 10538654 TI - Internet outsourced application provides IT cost reduction. PMID- 10538655 TI - Managing healthcare administration in the information age. PMID- 10538656 TI - Who owns your ideas? PMID- 10538657 TI - Where saving money meets saving lives. PMID- 10538658 TI - What works. Automated scheduling increases appointments, allows FTE reduction. PMID- 10538659 TI - What works. Rural medical center prepares for next century with end-to-end ATM. PMID- 10538660 TI - Enterprise integration: is there life after consolidation? PMID- 10538661 TI - Hotlist: wireless computers and networks. PMID- 10538662 TI - The value of integrated outcomes information for performance improvement initiatives. PMID- 10538663 TI - Candid comments on wireless. PMID- 10538666 TI - Records destruction--when? PMID- 10538665 TI - The Caldicott Report. PMID- 10538664 TI - Living wills. PMID- 10538667 TI - Cornish hospital records rescued. PMID- 10538668 TI - Market model fails: linkages between health policy and performance outcomes in western industrial countries. AB - In recent years, political discourse has sung the merits of the market model as a panacea for escalating health expenditures. While selected market mechanisms may be useful to control spending, there are reasons to question full-scale use of this model to shape health policy. Using the experiences of seven western industrialized countries, the argument developed in this article is that the market model's own assumptions cannot be upheld because ethical, sociological, and political factors underlying health policy make its application impossible. Specifically, a competitive equilibrium does not exist; sale of health care goods and services do not conform to the rules of marketability; and non-increasing returns are not demonstrable. A discussion of health outcome performance and spending data points out the need to attend to equity distributional concerns, state commitment to the health care access, and the extent and breadth of input into health care decision-making as other factors that should be considered. PMID- 10538669 TI - Fiscal policy dilemmas and health spending in South Africa. AB - There is broad agreement that government has an important role to play in the development of human capital, especially in health and education. Multilateral organizations, such as the World Bank, commonly call for public sector investment in human resources but the use of health spending to combat unemployment remains controversial. This article examines public sector expenditures in health and focuses on three arguments: 1) public goods; 2) investment in human capital; and 3) Keynesian spending in periods of high unemployment. PMID- 10538670 TI - Public and private perspectives on hospital conversions: proposed MUSC Columbia/HCA "partnership". AB - Conversion has been defined as "any type of transaction that results in the shift of all or a substantial portion of the assets of nonprofit health care organizations to for-profit use" (Claxton and Colleagues, 1997:10-11). Not surprisingly, efforts at such conversion create political conflict and opposition within communities with some groups supporting and others opposing the intended conversion. This study looks at a particular effort at conversion: the proposed "partnership" between three hospitals of the Medical University of South Carolina (MUSC) and Columbia/HCA, the largest for-profit health services network in the U.S. Beginning in 1995, the three-year effort (now at an impasse) has drawn local and state policy-markers into increasing controversy and acrimony. Beyond the details, the episode also raises fundamental issues about the nature of public health care and the extent to which we can convert public health facilities to private control while preserving their essential public missions. PMID- 10538671 TI - Health care as a social good: trauma care and the "kindness of strangers". AB - Urban trauma centers have been shown in the medical literature to be effective resources for dealing with traumatic injury in a manner which results in demonstrated increases in survival rates. Given that much debate exists over the relative efficacy of various technological medical interventions, the acceptance and diffusion of a "proven" technology, such as trauma centers, should be assured. Yet, the significant investment of resources required to staff, equip, and maintain a trauma center, coupled with a perceived fiscal deterioration of the provision of these services, has resulted in a retreat from the concept through closure of the services. PMID- 10538672 TI - Diversification strategy and performance: implications for health services research. AB - Health care represents a promising area of research due to its uniqueness. In recent years, considerable progress has been made in diversification strategy and performance research but not the study of health services strategy research. This article reviews diversification strategy and performance in health services domains. Adopting Datta, Rajagopalan, and Rasheed's (1991) framework, the authors evaluate the theoretical and empirical contributions of this research. The limitations and theoretical implications of these efforts are also explored. PMID- 10538673 TI - Burnout among leaders of Department of Veterans Affairs medical centers: contributing factors as determined by a longitudinal study. AB - Significant increases in the intensity of psychological burnout among leaders of local Department of Veterans Affairs (VA) medical centers occurred from 1989 to 1997. This longitudinal study was designed to analyze which demographic and behavioral variables were associated with the intensity of burnout. Responses of 83 Medical Center Directors, Associate Directors, and Chiefs of Staff to questionnaires sent in 1989, 1992, and 1997 were analyzed using path analysis. Burnout was assessed by the Maslach Burnout Inventory and the phase model for analysis. Questions also assessed antecedents (e.g., role characteristics and job support) and consequences (e.g., job satisfaction) of burnout. The presence of high levels of burnout rose from 25.3% in 1989 to 38.1% in 1997. Burnout phase in 1997 was directly related to burnout phase in 1992 and inversely related to the respondent's age. Phase in 1992 was inversely related to both resource availability and role clarity in 1989 if only accepted antecedents of burnout were included in the model. If consequences and antecedents were included in the model, burnout phase in 1997 was inversely related to job satisfaction and resource availability and directly related to intent to stay in the VA in 1989. Findings demonstrated that specific demographic and job site characteristics are associated with high levels of burnout in the VA. These could form the basis for interventional efforts. PMID- 10538674 TI - Revamp your perioperative services to save millions. PMID- 10538675 TI - Wisconsin hospital finds creative ways to reduce ADEs; avoids the "quick fix" approach. AB - Small changes can reduce adverse drug events. Hospitals looking for a quick fix to reduce adverse drug events usually find out it's not that easy. But implementing several minor changes can make a difference. That's what Luther/Midelfort in Eau Claire, WI, did when it revamped more than 10 processes over three years, resulting in fewer medication errors. PMID- 10538676 TI - Innovative maintenance contract for phone switches saves system more than $100,000 a year. PMID- 10538677 TI - Average cholecystectomy charges vary greatly depending on location of procedure. AB - DATA BENCHMARKS: Study finds wide variation in average charges for cholecystectomies. An in-depth analysis of claims admissions data from The Metropolitan Life Insurance Company compares average charges for in-hospital and outpatient laparoscopic and traditional open cholecystectomies by state and by region. The analysis also reveals significant differences by state in the percent that hospital charges represent of the total bill for those procedures. PMID- 10538678 TI - Adopt geriatric model to succeed in managed Medicare. PMID- 10538679 TI - Boost your bargaining clout in contract negotiations with Medicare managed care plans. AB - Boost your clout in negotiations with Medicare plans. Find out what Desert Physicians Association in Mesa, AZ, has learned in its dispute over reimbursement rates with CIGNA, and get some advice from a Medicare managed care consultant on how to negotiate contracts with Medicare plans. PMID- 10538680 TI - Providers develop health plan to manage Medicaid patients without woes of HMOs. AB - Managing Medicaid lives without HMOs. Providers in Pitt County, NC, have developed their own kind of managed care plan that brings together traditional Medicaid providers with private physicians seeking to improve care quality and patient access. Providers in the plan hope it can keep commercial HMOs out of the local Medicaid business. PMID- 10538682 TI - Study of five Medicare plans finds 10 budget-busting drugs. PMID- 10538683 TI - Cut inpatient utilization in half with proper use of home health services. PMID- 10538681 TI - Medicaid plans collaborate to improve birth outcomes, ease data reporting problems. AB - Collaboration in Medicaid solves providers' data reporting troubles. If you're looking for effective ways to give plans the information they want on pregnant Medicaid members and you want good programs for your high-risk patients, see how plans and providers in Philadelphia collaborated to improve birth outcomes. PMID- 10538685 TI - Kids with asthma lose weight, build strength and self-assurance. PMID- 10538684 TI - RTs, doctors team up to lead successful asthma program. PMID- 10538686 TI - HCFA projects may signal first step toward Medicare DM. AB - HCFA is finally clarifying the timing and other details behind two separate DM demonstration projects. The initiative could signal the first step toward integrating disease management into the Medicare payment system. PMID- 10538687 TI - Doctors at the heart of this cardiac DM program's success. AB - Don't let patients wait for a crisis event to jolt them into your coronary heart disease program. A new population-based effort developed by a trendsetting cardiac DM provider is winning praise as well as producing savings. PMID- 10538688 TI - Triage program provides direct access to ER care, but without the high cost. AB - A new program called ER Direct combines telephone triage with ER-based triage. The program is slashing the rate of retrospective ER claim denials and enhancing member access while still reining in emergency room costs. PMID- 10538689 TI - Federal and state privacy legislation moving ahead. AB - Federal and state legislators continue moving swiftly to enact legislation regarding the privacy of medical records. In this quick update, find out the latest and how it may affect your disease management efforts. PMID- 10538690 TI - Try these 10 tips to close loopholes at contract renewal. PMID- 10538691 TI - Risk-sharing 'pods' help PHO to manage global capitation. AB - A Connecticut PHO improved its management of capitation by reorganizing physicians into more effective decision-making groups, implementing a comprehensive medical management program, and developing a reporting system to support physician leaders. PMID- 10538692 TI - Are you benchmarking the right data to succeed under risk? PMID- 10538693 TI - Try this actuarial calculation for percent-of-premium contracts. PMID- 10538695 TI - Marketplace. Corporate wellness program now aim to improve a worker's entire wellbeing. PMID- 10538694 TI - Outside accounting help may be saving grace for capitated provider groups. PMID- 10538696 TI - Perspectives. Clinton plan offers drug benefit, new outlook on managed care. PMID- 10538697 TI - Perspectives. Women's health: pursuing quality and equality in a changing health system. PMID- 10538698 TI - A creative program that identifies and tracks the "unhealthy well". PMID- 10538699 TI - The role of a cancer risk assessment clinic. PMID- 10538700 TI - An amalgamation of four specialty hospitals. Lessons learned. PMID- 10538701 TI - Communication as a priority for success: lessons learned through change at St. Michael's Hospital. PMID- 10538702 TI - Crisis: the ultimate test for you and your hospital. PMID- 10538703 TI - Community relations: two perspectives. PMID- 10538704 TI - Why community relations is a strategic imperative. PMID- 10538705 TI - Consumers and hospital report cards: first efforts at definition. PMID- 10538706 TI - Cleveland's five-year experience with public reporting of hospital quality performance measurements. PMID- 10538707 TI - A progress report on Canadian Blood Services and its healthcare liaison projects. PMID- 10538708 TI - Building on strength: improving governance and accountability in Canada's voluntary sector. PMID- 10538710 TI - Legal aspects of supply chain continuity and contingency planning. PMID- 10538709 TI - Y2K supply chain continuity: the health sector's lifeline. PMID- 10538711 TI - Healthcare associations. Best practices in Internet communication. PMID- 10538712 TI - Ten predictions of the future of electronic medical record systems. PMID- 10538713 TI - What do Canadians think of the healthcare system? PMID- 10538714 TI - How do researchers influence decision-makers? Case studies of Mexican policies. AB - Though the problems translating or applying research in policy-making are legion, solutions are rare. As developing countries increase their capacities to develop effective local solutions to their health problems, they confront the research/policy dilemma. Yet few descriptive studies of research-policy links can be found from developing countries, and the relevance of European and North American models and data is questionable. We report the results of a descriptive study from Mexico of the relationship between health research and policy in four vertical programmes (AIDS, cholera, family planning, immunization). We interviewed 67 researchers and policy-makers from different institutions and levels of responsibility. We analyzed interviewee responses looking for factors that promoted or impeded exchanges between researchers and policy-makers. These were, in turn, divided into emphases on content, actors, process, and context. Many of the promoting factors resembled findings from studies in industrialized countries. Some important differences across the four programmes, which also distinguish them from industrialized country programmes, included extent of reliance on formal communication channels, role of the mass media in building social consensus or creating discord, levels of social consensus, role of foreign donors, and extent of support for biomedical versus social research. We recommend various ways to increase the impact of research on health policy-making in Mexico. Some of the largest challenges include the fact that researchers are but one of many interest groups, and research but one input among many equally legitimate elements to be considered by policy-makers. Another important challenge in Mexico is the relatively small role played by the public in policy making. Further democratic changes in Mexico may be the most important incentive to increase the use of research in policy-making. PMID- 10538715 TI - Political analysis of health reform in the Dominican Republic. AB - This article examines the major political challenges associated with the adoption of health reform proposals, through the experience of one country, the Dominican Republic. The article briefly presents the problems of the health sector in the Dominican Republic, and the health reform efforts that were initiated in 1995. The PolicyMaker method of applied political analysis is described, and the results of its application in the Dominican Republic are presented, including analysis of the policy content of the health reform, and assessment of five key groups of players (public sector, private sector, unions, political parties, and other non-governmental organizations). The PolicyMaker exercise was conducted in collaboration with the national Office of Technical Coordination (OCT) for health reform, and produced a set of 11 political strategies to promote the health reform effort in the Dominican Republic. These strategies were partially implemented by the OCT, but were insufficient to overcome political obstacles to the reform by late 1997. The conclusion presents six factors that affect the pace and political feasibility of health reform proposals, with examples from the case of the Dominican Republic. PMID- 10538716 TI - Using problem structuring methods in strategic planning. AB - In this paper we present approaches to problem structuring that have been employed to derive planning guidelines as part of a comprehensive strategic planning process. The approaches were developed for use in the context of a developing country, where quantitative data is particularly scarce. They rely heavily upon the informed judgement of technical planning officers. We discuss ways of ensuring that the approach remains flexible and participative. PMID- 10538717 TI - Health-care facility choice and the phenomenon of bypassing. AB - Health policy-makers in developing countries are often disturbed and to a degree surprised by the phenomenon of the ill travelling past a free or subsidized local public clinic (or other public facility) to get to an alternative source of care at which they often pay a considerable amount for health care. That a person bypasses a facility is almost certainly indicative either of significant problems with the quality of care at the bypassed facility or of significantly better care at the alternative source of care chosen. When it is a poor person choosing to bypass a free public facility and pay for care further away, such action is especially bothersome to public policy-makers. This paper uses a unique data set, with a health facility survey in which all health facilities are identified, surveyed, and located geographically; and a household survey in which a sample of households from the same health district is also both surveyed and located geographically. The data are analyzed to examine patterns of health care choice related to the characteristics and locations of both the facilities and actual and potential clients. Rather than using the distance travelled or some other general choice of type of care variable as the dependent variable, we are able actually to analyze which specific facilities are bypassed and which chosen. The findings are instructive. That bypassing behaviour is not very different across income groups is certainly noteworthy, as is the fact that the more severely ill tend to bypass and to travel further for care than do the less severely ill. In multivariate analysis almost all characteristics of both providers and facilities are found to have the a priori expected relationships to facility choice. Prices tend to deter use, and improved quality of services to increase the likelihood of a facility being chosen. The answer to the bypassing dilemma seems to be for providers to provide as good quality care relative to the money charged (if any), as other, often further away, providers. PMID- 10538718 TI - Unofficial fees in Bangladesh: price, equity and institutional issues. AB - The widespread collection of unofficial fees at health facilities is a common form of rent-seeking behaviour in Bangladesh. Typically, unofficial fees come in the form of cash payments for the performance of required services, for direct purchase of drugs and medical-surgical requisites, and for service access. Using observational and interview methods, this study explores linkages between official and unofficial fees at three Bangladesh health facility levels; primary care Thana Health Complexes, secondary or district hospitals, and medical college hospitals. The study estimates payment levels for different income classes and different payor types at these facilities, thereby highlighting potential equity, price and institutional questions associated with unofficial fees. Not only does the practice have clear income and equity effects, there also appear to be direct effects upon patient satisfaction, perception of quality, and the ability to pay for health services. The article concludes with a discussion of 'rent capture' processes at Bangladesh facilities and the effect of unofficial fees in six areas of health sector reform: displaced official policies, reduced merit goods production, upward income redistribution, distorted human resource development, growth of facility inefficiency, and obstruction of market reforms. PMID- 10538719 TI - The Spanish health care system: lessons for newly industrialized countries. AB - This article summarizes the organization, financing, and delivery of health care services in Spain, and discusses the elements that made it possible to maintain high levels of health among the population, while spending comparatively fewer resources on the health care system than most industrialized countries. The case of Spain is of particular interest for newly industrialized countries, because of the fast evolution that it has undergone in recent years. Considered, by United Nations' economic standards, a developing country until 1964, Spain became in a few years the fastest growing economy in the world after Japan. By the early 1970s the infant mortality rate was already lower than in Britain or the United States. PMID- 10538720 TI - Private health care in Nigeria: walking the tightrope. AB - The persistently low quality and inadequacy of health services provided in public facilities has made the private sector an unavoidable choice for consumers of health care in Nigeria. Ineffective state regulation, however, has meant little control over the clinical activities of private sector providers while the price of medical services has, in recent years, grown faster than the average rate of inflation. Reforms that are targeted at reorganizing the private sector, with a view to enhancing efficiency in the supply of services, are urgently required if costs are to be contained and consumers assured of good value for money. PMID- 10538721 TI - Identifying disability: comparing house-to-house survey and rapid rural appraisal. AB - This study compared house-to-house survey and rapid rural appraisal as methods used to identify people with disabilities in a sample rural population in South India. The research showed that by using these methods, two distinctly different populations were identified. The factors that influenced the identification processes were: local perceptions and definitions of disability; social dynamics, particularly those of gender and age; relationships within the rapid rural appraisal groups and between the health auxiliary and the respondents in the house-to-house survey; and the type of disability and the associated social implications and stigma of that disability. While a few more people were identified through the house-to-house survey, the rapid rural appraisal was a better approach for identifying disability in the community because of the greater community participation. The researchers believe that this community participation provided a greater understanding of the complex contextual dynamics influencing the identification of disability, thereby increasing the validity of the study findings. Another advantage of the rapid rural appraisal was the methodological and analytical simplicity. Both methods, however, failed to identify some individuals with disabilities who were later identified on the follow-up verification visits. Taking into account the factors discussed above, the researchers conclude that no single method could be used to comprehensively identify people with disability in a community. They suggest that a judicious combination of methods which takes into account local perceptions and priorities, includes more specific screening techniques, and facilitates informed voluntary referrals, would be the most effective approach. PMID- 10538722 TI - Does use of a government service depend on distance from the health facility? AB - To reduce mortality from common childhood illnesses such as diarrhoea and upper respiratory infections, it is important that health services are available and used appropriately. Physical accessibility to a health facility may influence its use, particularly in rural areas. We assessed whether use of government services for treatment of the three most common acute childhood illnesses (fever, diarrhoea and upper respiratory infections) was influenced by the physical accessibility of the government primary health care centres. We analyzed data from a household survey which was collected between November 1992 and January 1993, from 139 randomly selected villages located around 14 government facilities in Thatta, a rural district of Pakistan. There were 691 children under 5 years of age who suffered from the three acute illnesses; 85% of these children used either a government or a private service. Children living at less than 4 km from a government facility made 22% less use of that facility than those living 4 km or more away. After controlling for the effects of distance from a private facility and treatment cost in a multiple logistic regression model, children living less than 4 km from a government facility were no more likely to use the facility than those living 4 km or more away (Adjusted Odds Ratio: 1.01, 95% Confidence Interval: 0.68-1.50). These results suggest that factors other than distance are the primary determinants of use of government services for treating children in the Thatta district. To increase the use of government health services, policymakers should assess carefully the factors determining the use of existing facilities, before they plan the building of more health facilities. Further studies are needed to examine the management of health facilities and the clients' perception of health-care providers. PMID- 10538723 TI - Health impact assessment. AB - There is growing concern about the environmental, social and health consequences of development projects. Environmental impact assessment (EIA), which aims to address this concern, is often conducted with little input from the health sector. Quantifying the health benefits and risks of a project or policy requires an innovative synthesis of socio-demographic, environmental health, epidemiological and health systems data. This article provides a simple framework for health impact assessment (HIA), a method for describing and measuring the impact of a project or policy on health and wellbeing, and designing appropriate interventions. The key components of HIA are: review of available data; research and identification of priority health issues through the use of rapid assessment methods; design of a health action plan with stakeholder consultation; implementation of interventions and the monitoring of long-term health impacts. HIA can assist in ensuring that development and policies are 'health promoting' and that the health sector plays a meaningful role in EIA. PMID- 10538725 TI - Hospital settles patient/patient rape lawsuit after $2.5 million award. PMID- 10538724 TI - Intervention research in rational use of drugs: a review. AB - Many studies have been done to document drug use patterns, and indicate that overprescribing, multi-drug prescribing, misuse of drugs, use of unnecessary expensive drugs and overuse of antibiotics and injections are the most common problems of irrational drug use by prescribers as well as consumers. Improving drug use would have important financial and public health benefits. Many efforts have been undertaken to improve drug use, but few evaluations have been done in this field. This article provides an overview of 50 intervention studies to improve drug use in developing countries. It highlights what type of interventions exist and what is known about their impact. It reveals that commonly used interventions, such as an essential drug list and standard treatment guidelines, have rarely been systematically evaluated so far. The majority of intervention studies are focused on prescribers in a public health setting, while irrational use of drugs is also widespread in the private sector. Furthermore, the magnitude of inappropriate drug use at community level is often overlooked and few interventions address drug use from a consumer's perspective. More research on different types of intervention strategies in various health care settings is needed to draw conclusions on the effectiveness of a specific intervention strategy. Also more research is needed on socio-cultural factors influencing the impact of drug use interventions, particularly from a user perspective. To enhance evaluative research, more technical support will be needed for researchers in developing countries. The design of available studies from developing countries is generally weak, only six of the 50 studies included in this overview were randomized controlled studies. In order to provide technical support and coordination of future intervention research the establishment of an international resource centre for drug use intervention research is recommended. PMID- 10538726 TI - Answering your questions about safety, feasibility of satellite parking. PMID- 10538727 TI - Bill Kizorek on surveillance: what you can and cannot do today. PMID- 10538728 TI - 'Help' phones in hospital parking facilities not just for emergencies. PMID- 10538729 TI - Kankakee, IL: two hospitals; a middle-of-the-night train crash; 114 injured--how their disaster plans played out. AB - The emergency disaster plans of two Illinois hospitals were recently put to the test after an Amtrak train crash that occurred on the night of March 15 when a passenger train traveling from Chicago to New Orleans collided with a semi trailer truck, causing the train to derail about 50 miles south of Chicago in the city of Bourbonnais, IL. The injured were taken to Riverside Medical Center and Provena St. Mary's Hospital, Kankakee, IL--a city of 30,000. Out of the 216 passengers on Amtrak's City of New Orleans, 11 people, all located in the sleeper car, were killed and 114 were injured. The accident was the largest that the two area hospitals had ever dealt with. Here's how they handled it. PMID- 10538730 TI - Securing medical hotels for hospital patients and their families. PMID- 10538731 TI - A gap analysis of HMO service quality. PMID- 10538732 TI - Localization of relevant consequences in anti-drinking and driving PSAs (public service announcements): a new approach to targeting underage college students? AB - Drinking and driving is prevalent among college students, particularly among underage college students. Although most of these students know that drunk driving is a potentially harmful behavior, many of them still make the decision to do so. The purpose of this study was to determine why underage students drink and drive and what consequences, if any, these students fear when they engage in this behavior. Focus groups and in-depth interviews were conducted, using the Health Belief Model as a guide for discussion topics. Participants revealed that PSAs focusing on relevant, localized consequences would have more meaning to underage college students than the more general campaigns that they reported seeing. In addition, they revealed that the consequence that they fear most is being charged with a DUI, yet current anti-drinking and driving PSAs never portray this as a possible negative consequence. PMID- 10538733 TI - Asian American health care attitudes. AB - This paper describes the results of a survey of health care attitudes of a sample of respondents primarily of Asian American background. The importance of bilingualism, Asian background, age, and other attributes of a physician are discussed with relation to subgroups in the sample. The relative importance of the influence of doctors, family, and friends on the choice of physician and health care facility are also presented. The findings may help with the development of effective market segmentation and improved health care service to the Asian American community. PMID- 10538734 TI - Physician recruitment: understanding what physicians want. AB - One of the more dramatic changes in the healthcare industry has been the movement of physicians, particularly younger professionals, from private practice to some type of healthcare organization. In this study we examine the importance attached to specific incentives by physicians in making an affiliation decision and healthcare administrators. Our results suggest significant differences between the importance placed on certain recruiting incentives by physicians and healthcare administrators. Further, they suggest distinct differences in importance ratings by different types of physicians. Implications of this study argue for developing different compensation packages to appeal to different segments of physicians. PMID- 10538735 TI - Levels of service in independent community pharmacies: a strategic group analysis. AB - A sample of independent community pharmacies was analyzed for the existence of strategic groups utilizing service variables exclusively. Results indicate that strategic groups corresponding to high, moderate, and low levels of clinically related services do exist in this sample of independent community pharmacies. In addition the performance implications of strategic group membership was tested. Results indicate there was no statistical difference in performance across strategic groups. Implications for theory and practice are discussed. PMID- 10538736 TI - The applicability of SERVQUAL in different health care environments. AB - This paper reports on a study that investigates the applicability of a modified SERVQUAL instrument as a means of measuring service quality in two types of health service environments; medical care and health care (incorporating medical, social, cognitive and emotional elements). The research confirms a four factor structure that is stable for both environments, and similar to the service quality dimensions recognised in the literature. However, the relative importance of the dimensions of quality is inconsistent for the two types of health services. These results confirm the suggestion that importance values should be part of the measurement tool. Finally, the extra diagnostic advantage achieved by the use of gap scores to measure service quality, when compared to perception only scores is demonstrated. PMID- 10538737 TI - "The home infusion patient": patient profiles for the home infusion therapy market. AB - The authors review the relevant literature regarding home health care patient profiles. An empirical analysis is provided from archival data for a home infusion company servicing patients in urban and rural areas. The results are provided as a 2 x 2 matrix for patients in urban and rural areas seeing either a specialist or primary care physicians. A series of moderated regressions indicate that type of treating physician, patient's gender, geographic residence and level of acuity are cogent in predicting the complexity of prescribed infusion therapies. Managerial implications are provided for the home care marketer in segmenting patient markets for infusion services. PMID- 10538738 TI - Influence of pain-free dentistry and convenience of dental office on the choice of a dental practitioner: an experimental investigation. AB - Fear of dental pain or dental anxiety is not an uncommon phenomenon. Evidence supports dental patients' concern for dental pain. This study examines the influence of dental pain and convenience of dentist's office on attitude towards the dentist as well as the intention to switch to the dentist. The results reveal a strong effect of dental pain on patients' attitudes. The intention to switch to a pain-free dental facility, however, depends on satisfaction with the current dentist. PMID- 10538739 TI - Strategic planning in community mental health centers. PMID- 10538740 TI - Responsibility and risk: understanding how PSA messages can encourage condom use. AB - This article qualitatively examines how female college students interpret their responsibility for maintaining healthy sexual behaviors. A series of nine in depth interviews was conducted to assess their perceptions on a broad range of sexual health topics including personal fears, relationships, monogamy, and sexual relationships. In addition, the paper addresses how these women negotiate condom use, responsibility issues regarding this negotiation, and perceived risk for contracting AIDS during unprotected sex. This study has implications for health educators, as well as for designers of public service advertisements, as it provides an in-depth look at why college women do not use protection even when it is available to them. PMID- 10538741 TI - AIDS funding. The national lottery. AB - The government's funding of AIDS and HIV services in England is inequitable. Spending per patient in North Thames is four times that in Manchester. Deaths from AIDS in North Thames are significantly lower than in other parts of England. PMID- 10538742 TI - Hospital discharge. Smooth passage. AB - The introduction of care co-ordinators into a medical directorate two years ago has reduced bed blocking and cut length of stay from 8.5 to 5.9 days. All those appointed were already working in the hospital. There have been some tensions over boundaries with nursing and social work. PMID- 10538743 TI - Nursing recruitment. Starters' orders. AB - An 18-month support programme for newly qualified nurses at one trust has improved recruitment and retention. The programme includes training in clinical activities as well as ongoing supervision and support and a management course. Of 85 nurses who completed the programme only six have left the trust. PMID- 10538745 TI - Primary care groups. Chaos theory. PMID- 10538744 TI - US healthcare. A fistful of dollars. PMID- 10538746 TI - Read codes. Swallowed whole. PMID- 10538747 TI - Software. No soft options. PMID- 10538748 TI - Electronic results. Distant relations. PMID- 10538749 TI - Linking hospitals. Two-way traffic. PMID- 10538750 TI - Pharmaceuticals. Resistance fighters. PMID- 10538751 TI - Mental health. Open and shut case. AB - Acute mental care consumes about two-thirds of all mental health spending. But many beds are blocked by patients who no longer need intensive support. They stay in hospital because of a lack of alternative services. If each patient left hospital when they were deemed ready, inpatient costs would be almost halved. More than a third of these patients are re-admissions. A range of alternative crisis services, including 24-hour nursing care and staffed hostels, are needed. PMID- 10538752 TI - Right jab ... child immunisation targets. PMID- 10538753 TI - Health improvement programmes. Avon calling. AB - Organising a health improvement programme is a huge task. It is a challenge to meet participants' expectations, and many will be disappointed that much day-to day work of the NHS and social services is not included. The process is an opportunity to bring together people from different organisations and form new partnerships. It takes longer than expected. PMID- 10538754 TI - Lights out. PMID- 10538755 TI - Long-term care. Wait 'til your dad gets home. AB - Patients and carers were unaware of eligibility criteria for long-term care. Many patients interviewed in a qualitative study were not aware they had been assessed for services. Delays in providing rehabilitation services and mobility restrict independence. PMID- 10538756 TI - Long-term care. Strike up the bands. AB - Assessing elderly people for long-term care according to a banding system, with the assessors meeting weekly, can speed up placements. The system is believed to cut down on inappropriate placements. A rise in the number of elderly people in nursing home places funded by the local authority reflects increasing levels of dependence in the community. PMID- 10538757 TI - Overseas health care. Grandmothers' footsteps. AB - The Maori population of New Zealand has poorer health than the general population and finds access to traditional health services difficult. A programme using non professionals for health promotion has produced significant results. Take-up of immunisation has improved dramatically. PMID- 10538758 TI - On the evidence. Gynaecological cancers. PMID- 10538759 TI - Data briefing. Patients' views. PMID- 10538760 TI - Bristol inquiry. Iron fist, kid gloves. PMID- 10538761 TI - Obesity. About the size of it. AB - Severe obesity is on the increase in Britain and accounts for considerable excess hospital costs. The benefits of new treatments need to be assessed in terms of psychological, as well as physical, benefits to patients. Programmes aimed at changing lifestyles will be effective only when obesity is considered in terms of all its effects on the individual. PMID- 10538762 TI - Primary care groups. Sound backing for little voice. PMID- 10538763 TI - Contradiction in terms. PMID- 10538764 TI - Engaging the public in quality oversight. PMID- 10538766 TI - Special report. Standards revisions for 2000. PMID- 10538765 TI - Hot line increases awareness and access. PMID- 10538767 TI - O'Leary testifies on restraint. PMID- 10538768 TI - ORYX requirements simplified. PMID- 10538769 TI - Meeting the most challenging hospital standards. PMID- 10538770 TI - By the numbers. Miscellaneous. PMID- 10538771 TI - By the numbers. Healthcare economics. PMID- 10538772 TI - By the numbers. Medicare and Medicaid. PMID- 10538773 TI - By the numbers. Vital health statistics. PMID- 10538774 TI - By the numbers. Hospitals and health systems. PMID- 10538775 TI - By the numbers. Physicians. PMID- 10538776 TI - By the numbers. Insurers and managed care. PMID- 10538777 TI - By the numbers. Post-acute care. PMID- 10538778 TI - By the numbers. Public opinion. PMID- 10538780 TI - A crackdown on bad eggs. Egg-carton warnings sought to fight Salmonella outbreaks. PMID- 10538779 TI - Why Dr. Kildare stuck to medicine. PMID- 10538781 TI - Cancer's Trojan horse--tricking malignant cells to self-destruct. PMID- 10538782 TI - The low-fat life. You don't have to meditate and eat like a rabbit. PMID- 10538783 TI - The American Stop Smoking Intervention Study. Conceptual framework and evaluation design. AB - Reducing tobacco use, especially cigarette smoking, is a public health priority. The American Stop Smoking Intervention Study (ASSIST) was initiated in 1991 to prevent and reduce tobacco use primarily through policy-based approaches to alter the social-political environment. This article describes the conceptual design, research framework, evaluation components, and analytic strategies that are guiding the evaluation of this demonstration research endeavor. The ASSIST evaluation is a unique analysis of the complex relationships between the social context, public health activity at the state level, tobacco use, and individual behavior. The measures of tobacco control activity developed for this evaluation may be useful in ongoing national cancer control surveillance efforts, and the lessons learned will enhance the development of tobacco control programs. PMID- 10538784 TI - Outcome evaluation of an advocacy program to promote early childhood education for Israeli Arabs. AB - This article reports the results of an evaluation of the SHATIL early childhood education (ECE) project, a coalition-based advocacy initiative that aimed to achieve three main outcomes in Arab towns throughout Israel: growth in preschool enrollment rates, growth in number of preschool classes administered by local councils, and growth in number of certified preschool teachers working in the Arab sector. The study employed a quasi-experimental, repeated measures design. Outcomes were assessed via collection of archival data pertaining to each of the past 10 years in 45 Arab towns. This allowed for comparison of trends over time in the 5 years before project commencement with those in the 5 years since the project was launched. Findings indicated that the project was successful in recruiting its target population (the most needy towns) and that the desired changes in fact did occur, particularly in the target towns. Moreover, the authors were able to conclude, with a relatively high degree of confidence, that the project played a causal role in achieving these outcomes. PMID- 10538785 TI - Implications of the 65-mph speed limit for traffic safety. AB - This study evaluates the impact of the 65-mph speed limit on traffic safety. Using data for the years 1981 to 1995 for all 50 states, a pooled time series analysis is conducted. Separate models are estimated for state fatality rates on four categories of roads: rural interstate highways, rural noninterstate roads, all roads except for rural interstate highways, and all roads. It is reported that the 65-mph speed limit increased fatality rates on rural interstate highways but was correlated with a reduction in state fatality rates on the three other categories of roads. PMID- 10538786 TI - Differential attrition rates and active parental consent. AB - Active parental consent in survey research poses ethical and practical concerns. One common argument against the requirement of active consent procedures is its effect on participation rates. There is additional concern that higher risk groups may be underrepresented in the final sample. Empirical support of differential attrition, however, is lacking. In the current multisite longitudinal study, passive consent procedures were approved for the collection of pretest data. For subsequent years of data collection, active parental consent procedures were required. In this article, we use the pretest data to examine demographic, attitudinal, and behavioral differences between those students for whom active consent was provided and those for whom active consent was either denied or for whom no response was received. The results indicate that active consent procedures produce deleterious effects on participation rates and lead to an underrepresentation of at-risk youth in the sample. PMID- 10538787 TI - The results framework--an innovative tool for program planning and evaluation. AB - This article constitutes a case study of the development and implementation of the "results framework," an innovative planning and evaluation tool that is rapidly becoming a standard requirement for United States Agency for International Development (USAID) projects. The framework is used in a USAID funded regional initiative for HIV/AIDS prevention in Central America. This new program evaluation and monitoring tool provides many advantages over traditional evaluation approaches that use outside consultants to provide midterm and end-of project evaluations. The results-framework process, which spans the life of the project, provides an opportunity for program staff, donors, partners, and evaluators to work as a team to collect and use rich, longitudinal data for project planning, implementation, and evaluation purposes. PMID- 10538788 TI - Fast tracking trauma patients through the ICU. PMID- 10538790 TI - Evolution of Joint Commission ORYX initiative. PMID- 10538789 TI - Prevention of wrong-site surgery. PMID- 10538791 TI - Skill mix in primary care--creating a bibliography. AB - Skill mix in primary care should be governed by research-based evidence of how skills may best be distributed among health professionals in order to optimize the cost-effectiveness of health service delivery. There is a dearth of research in this area, yet many changes in skill mix within primary care have still to be adequately researched. Existing evidence of the nature and cost-effectiveness of skill mix change is scattered across the specialist literature of many different disciplinary groups, making it difficult to form a coherent overview of service provision. This paper reports on a project that sought to create a comprehensive database bringing together specialist literature relating to skill mix in primary care, as resource for health services researchers, providers and purchasers in the UK. PMID- 10538793 TI - Primary Care Sharing the Evidence (PRISE) project Website: bringing high-quality information to primary health care teams. PMID- 10538792 TI - Efficient literature searching in diffuse topics: lessons from a systematic review of research on communicating risk to patients in primary care. AB - Using the example of communication about risk in a primary care setting, this paper puts forward a method of developing and evaluating a detailed search strategy for locating the literature for a systematic review of a 'diffuse' subject. The aim of this paper is to show how to develop a search strategy that maximizes both recall and precision while keeping search outputs manageable. Six different databases were used, namely Medline, Embase, PsychLIT, CancerLIT, Cinahl and Social Science Citation Index (SSCI). The searches were augmented by hand-searching, contacting authors, citation searching and reference lists from included papers. Other databases were searched but yielded no extra references for this subject matter. Of the 99 papers included, 80 were indexed on Medline. The Medline search strategy identified 54 of them and the remaining 26 were located on other databases. The 19 further unique references were found using the other databases and methods of retrieval. A combination of several databases must be used to maximize recall and to increase the precision of searches on individual databases, thus improving the overall efficiency of the search. PMID- 10538794 TI - A checklist for clarifying issues in working with primary care. PMID- 10538795 TI - Designed to care: the Scottish perspective on primary care in the new NHS. PMID- 10538797 TI - Providing library support for the development of clinical guidelines. PMID- 10538796 TI - Do PCGs need information officers? PMID- 10538798 TI - Research into practice? PMID- 10538799 TI - The Wisdom Project: virtual education in primary care. AB - This article examines the development of the Wisdom Project, a pilot for the teaching of informatics to primary health care professionals, using an original educational model based on e-mail and Web pages. The article begins by placing the development of the Wisdom Project in the context of changes in medical education and training. The aims and objectives of Wisdom are outlined, and the methodologies for setting up and evaluating the project are described. The article then presents the results of the evaluation, including the identification of significant improvements in knowledge of CD ROMs (P = 0.01), e-mail (P = 0.03), Medline (P = 0.02), operating systems (P = 0.02), Web browsers (P = 0.003) and word processing (P = 0.03). Improvements in evidence-based practice (EBP) did not reach significance. Finally, a number of conclusions are presented, considering the lessons learnt for the future development of such projects. PMID- 10538801 TI - Community access to health information in Ireland. AB - This paper is based on a research project conducted on consumer health information (CHI) in the Republic of Ireland, the results of which were published in a report entitled Well Read: Developing Consumer Health Information in Ireland. The paper describes the research methodology and the Irish experience in relation to CHI followed by a discussion of access problems, illustrated with examples from the special needs and primary care sectors. The role of information providers in relation to primary healthcare and libraries is examined briefly, and finally the main research conclusions and recommendations are highlighted. PMID- 10538800 TI - Evidence into practice: an information service for primary care professionals. AB - This paper describes the development of the Primary Care Information Service (PCIS) in South Humber Health Authority--a practical example of an information service which aims to make it easier for primary care practitioners to have access to information services and resources, both by electronic and more traditional means. Also, issues to consider when establishing a service for primary care professionals, including barriers to be overcome, are outlined, and an examination is made of how the service has been developed around principles of evidence-based medicine. Finally, the achievements to-date are considered. PMID- 10538802 TI - Older people's health information needs. AB - This paper, originally given at the Health Libraries Group conference 'First aid for the front line' in September 1998, argues that older people's health information needs are in many respects little different to anyone else's. It looks at different formats of information that may be helpful for older people, and at the needs of two specific groups, carers and elders from ethnic groups. Direct user involvement in the development of information is seen as important. It is emphasized that there is a particular need for primary care workers to know how other services, such as those within social care and the voluntary sector, operate. The paper ends by looking at the information professional's role in providing resources and training to such workers, and suggests a broader remit for the profession as a whole, to help to ensure social inclusion for older people. PMID- 10538803 TI - Practice brief: the care and maintenance of charge masters. American Health Information Management Association. PMID- 10538804 TI - The vocabulary of experience. Interview by Jane E. Blumenthal. PMID- 10538805 TI - Healthcare legislation forges ahead. PMID- 10538807 TI - Coding compliance tips for hospital outpatient observation services. PMID- 10538806 TI - Roundtable takes on compliance challenges. PMID- 10538808 TI - An HL7 (Health Level Seven) overview. AB - HL7 is an organization with a short history and more activity now than ever before. Opportunities abound within HL& to assist in the development of the automation of healthcare and the deployment of information systems that create and support the electronic medical record. Finally, the continual advances in information systems technology enable HL7 to do more to meet the healthcare industry's needs. PMID- 10538810 TI - Three perspectives on coding. AB - It isn't just about assigning codes any more. These days, coders are breaking new ground in their profession. Three authors offer different perspectives on the role of the coder in today's healthcare environment--from their evolution in the world of HIM to broader connections with physicians and issues of compliance. PMID- 10538809 TI - Automated coding: the next step? PMID- 10538811 TI - Mining the CPR (and striking research gold). PMID- 10538812 TI - A new role for HIM in the knowledge economy. PMID- 10538813 TI - Cultivate a desire to learn. PMID- 10538814 TI - Best practices: what works? PMID- 10538815 TI - In search of best practices. PMID- 10538816 TI - AMAP (American Medical Accreditation Program) plots course for physician assessment. PMID- 10538817 TI - Job hunting in the electronic age. PMID- 10538820 TI - Tyrannosaurus, tortoises, and tunes: making quality learning fun. PMID- 10538818 TI - Group builds consensus on laboratory test policies. PMID- 10538821 TI - Nursing shortages: ten strategies to becoming a "magnet hospital" for RN recruitment and retention. PMID- 10538819 TI - Moving patients through: three top-performing emergency departments demonstrate how it's done. PMID- 10538822 TI - Long-term outlook: today's nurses may be tomorrow's nurses too. PMID- 10538824 TI - Regulation of health: case studies of Sweden and Switzerland. PMID- 10538823 TI - Magnet hospitals: ten strategies to becoming a model nursing employer. PMID- 10538825 TI - Regulation of health: case studies of Sweden and Switzerland. Introduction. PMID- 10538826 TI - Licensing of firms and institutions. PMID- 10538827 TI - Reimbursement of hospital services and hospital financing. PMID- 10538828 TI - Incentives for diffusion of new health care technology. PMID- 10538829 TI - The market for pharmaceuticals. PMID- 10538830 TI - Programs for the aged in Sweden and in Switzerland. PMID- 10538831 TI - Compensation for health-related loss of income. PMID- 10538832 TI - Taxes, premiums, user charges: financing from the point of view of consumers. PMID- 10538833 TI - Differences in taxation and regulation of health-affecting goods--alcohol and tobacco. PMID- 10538834 TI - Licensing of physicians. PMID- 10538835 TI - Overview of the two systems. PMID- 10538836 TI - Price setting for doctors. PMID- 10538837 TI - Is your ED overcrowded? Reduce risks with these aggressive tactics. PMID- 10538838 TI - Reduce risks of patients who leave the ED. PMID- 10538839 TI - Onsite visits identify ED-specific issues. PMID- 10538840 TI - Educate on-call consultants about EMTALA. PMID- 10538842 TI - Legislation delaying OSHA from issuing ergonomics regulation progresses in Congress. PMID- 10538841 TI - Court issues first ruling on prudent buyer therapy disallowances. PMID- 10538843 TI - Second year of PPS transition to bring new cost control challenges for SNFs. PMID- 10538844 TI - Risk-sharing, more ancillary service management critical for continued survival under PPS. PMID- 10538845 TI - Medical and economic impact of breast cancer treatment and prevention. PMID- 10538846 TI - Benefits foregone: too much of the wrong and too little of the right. Based on a presentation by Bernard S. Bloom, PhD. AB - Physicians, patients, and payers use some interventions that do not work, while they ignore others that have a scientific basis for efficacy. As a society, Americans are unwilling to address collectively healthcare issues, as witnessed by legislative inaction on reform. They have chosen instead to leave such matters up to the impersonal economic market. In doing so, the basic laws of economics must be taken into account: Healthcare resources are limited and conscious societal trade-offs must be made about who gets which services, how much, and who pays. Healthcare resources tend to be rationed according to income, even with Medicaid and other programs available to those with low incomes. The economic market lacks perfect or almost perfect information to enable rational choices to be made among alternatives. Even when the value of an intervention is known, physicians may over- or underuse it. Much of the literature focuses on overuse, but underuse is finally being recognized as a severe problem. For example, effective prevention and treatment measures for breast cancer exist and should be promoted for appropriate candidates, despite the low relative risk of adverse effects like other neoplasms. PMID- 10538847 TI - Breast cancer: overview of current management considerations of economic impact. Based on a presentation by Sandra E. Brooks, MD, FACS, FACOG. AB - Most women who are diagnosed with breast cancer have either no risk factors or only one. Recommendations for breast cancer screening for women between the ages of 40 and 49 are controversial, because the National Institutes of Health (NIH) Consensus Conference made no specific recommendation regarding the frequency of screening. In so doing, a possible reduction in mortality or morbidity is balanced against known false-positive or false-negative rates and the detection of ductal carcinoma in situ. Although genetic testing is available, many questions surround its use, including its likely emotional and financial impact on the patient and her family and the unresolved issue of how to interpret the results. PMID- 10538848 TI - An overview of cancer economics. Based on a presentation by C. Daniel Mullins, PhD. AB - The National Cancer Institute (NCI) has estimated that the aggregate outlay for cancer, including research and direct medical costs, was about $104 billion in 1996. Treatment expenditures for the 3 leading types of cancer--breast, lung, and prostate--total $16 billion a year, with breast cancer alone accounting for $6 billion. Early detection is key to reducing the cost of breast cancer because the costs per patient are higher for those who die than for those who survive and the cost effectiveness of detection and treatment are inseparable. Likewise, screening can be cost effective but only if options exist for treatment. Lack of access to care for financial and other reasons continues to be a major impediment to early detection. PMID- 10538849 TI - Breast cancer stage cost analysis in a managed care population. Based on a presentation by Kenneth L. McDonough, MD, MS. AB - A sample managed care plan provides the framework for examining data on breast cancer, employing a naturalistic setting as opposed to the randomized control trial model. Breast cancer costs are analyzed by disease stage, which is broken down further by treatment phase. This approach compares the costs of those with treated disease to a control group, essentially performing a case mix adjustment for comorbidities. Such an approach encompasses the claims database characteristics, defines the breast cancer stages and treatment phases, and describes the criteria used for the control group, as well as the methodology utilized in the case mix adjustment. PMID- 10538850 TI - The economics and challenges of breast cancer in a managed care environment. Based on a presentation by Alan H. Heaton, PharmD. AB - Breast cancer and its population effect are inseparable. One of the challenges managed care organizations (MCOs) face is instilling the idea that patients are part of a population, and in turn, that population is composed of patients. Therefore, there is a need to treat both patients individually and populations as a whole. Because breast cancer, like other major illnesses, involves large-scale expenditures for drugs, pharmaceutical benefit management companies are working with MCOs to look not only at drug costs but at global healthcare expenditures. Whereas treatment of breast cancer has direct costs to a healthcare plan, it is associated with a great deal of comorbidity as well. In dealing with such potential financial exposure, the challenge to health plans is to find individuals at risk, enable them to access the healthcare system, and see that they get proper care. A proactive communications effort involving such media as patient newsletters and a website can educate healthplan members, thereby facilitating the self-assessment of risk factors. PMID- 10538852 TI - Clinical benefits of combination therapy in managing hypertension: a paradigm for managed care. PMID- 10538851 TI - The cost effectiveness of tamoxifen in the prevention of breast cancer. AB - In the National Surgical Adjuvant Breast and Bowel Project P-1 Breast Cancer Prevention Trial (BCPT), women considered to be at high risk for developing breast cancer who received tamoxifen experienced 49% and 50% reductions in the risk of developing invasive and noninvasive breast cancer, respectively, compared with women receiving placebo. Although the BCPT addressed the clinical benefits of tamoxifen, this study sought to assess its cost effectiveness in the prevention of breast cancer in women at increased risk for developing the disease. Women were considered to be at an increased risk if they were: 1) 60 years of age or older, 2) age 35 to 59 years with a history of lobular carcinoma in situ, or 3) age 35 to 59 years with additional risk factors that made their 5 year predicted breast cancer risk at least as great as that of women 60 years of age. A decision-analysis model was used to estimate the incremental cost effectiveness of using tamoxifen compared with no intervention as preventive therapy in age-group defined cohorts of women who were at high risk for developing breast cancer. The analysis used data on the benefits and risks of tamoxifen as observed in the BCPT. In a subgroup analysis, tamoxifen's cost effectiveness was also evaluated in women who had had a hysterectomy, because of evidence that suggested an increased risk of endometrial cancer in women receiving tamoxifen. Under conservative assumptions from a base-case analysis, the incremental cost effectiveness of tamoxifen is $41,372 per life-year gained for women age 35 to 49 years, whereas for women age 50 to 59 years and 60 to 69 years, these values are $68,349 and $74,981, respectively. For women with a previous hysterectomy, tamoxifen's cost effectiveness is $46,060 per life-year gained. A strategy of using tamoxifen in high-risk women to prevent breast cancer in high-risk women may be cost effective, particularly in the 35-to-49 year-old age group and in those of any age who have had a hysterectomy. PMID- 10538853 TI - Cost effectiveness of combination therapy. Based on a presentation by Daniel Hilleman, PharmD. AB - The ultimate economic goal of hypertension management is to balance costs and benefits, but defining these entities may be difficult. The overall cost of treating high blood pressure includes direct costs, such as drug acquisition, physician fees, laboratory and diagnostic tests, and management of side effects, as well as indirect costs, such as inadequate blood pressure control, noncompliance with therapy, and loss to follow up. Determining actual costs can be complicated. For example, medical charges are rarely paid as billed to third party payers, and actual payments received for services are reimbursed at rates that vary from patient to patient and provider to provider. As difficult as determining treatment costs may be, quantifying the benefits and outcomes of treatment is probably even more difficult, especially because outcome can be classified as long term (with few available outcomes data on fixed-dose combinations), intermediate-term, and short-term. If blood pressure is considered a surrogate marker for mortality, it could be used in comparing the economic value of some antihypertensive agents. Cost-effectiveness studies evaluating hypertension treatment typically compare 2 or more alternatives, with the cost defined by 1 or more of 4 units of effectiveness. These units include: the money that needs to be spent to achieve the following: reach a specific mm-Hg reduction in blood pressure, reach a specific percentage reduction in blood pressure, treat a patient successfully to target blood pressure level, and treat a patient per quality-adjusted life-year gained. In studies evaluating fixed-dose combination therapy versus monotherapy in terms of response rates, costs per patient per year, and costs per successfully treated patient per year, combination therapy was found to be more effective in lowering blood pressure, but more expensive. However, the higher response rates seen with combination therapy either offset the added costs of managing patients with inadequately controlled hypertension or provided considerably better blood pressure control for only a few additional dollars per patient per year. Because prescription drug costs represent a significant percentage of the total costs of treating hypertension over time, several cost-containment interventions have been devised. These include formulary restriction, generic substitution, therapeutic interchange, prior authorization, and drug utilization. PMID- 10538855 TI - The role of combination therapy in achieving blood pressure control. PMID- 10538854 TI - Impact of combination therapy on managed care. Based on a presentation by Lisa Latts, MD, MSPH. AB - The major objectives of managed care are particularly applicable to hypertension, a very common disease that affects about 30% of the adult population. Although managed care organizations know that adequate blood pressure control has a considerable impact on reducing hypertension-related morbidity and mortality, managed care is, at present, doing a generally poor job of monitoring antihypertensive therapy. One of the major reasons for this is the difficulty in gathering the necessary data from administrative sources; only chart review, which is very costly and very time consuming, can provide the data needed. The value of disease management lies in its capacity to optimize clinical and economic outcomes as well as to improve service and quality of life, using inputs such as cost, time, clinical resources, and patient satisfaction. Disease management programs are instituted because they can lead to improved quality of life and reduced costs. While the focus in managed care has traditionally been on promoting risk modification behavior, providing patient education, and encouraging medication compliance, the treatment of hypertension itself has not been a major focus in most disease management programs. Measuring the effectiveness of the hypertension treatment is, however, one of the proposed measures for the Health Plan Employer Data and Information Set (HEDIS 2000) to profile health plan quality. From a managed care perspective, the potential benefits of combination therapy for hypertension include improved blood pressure control and improved patient compliance as a result of needing fewer pills and experiencing fewer side effects with low-dose therapy. Among the obstacles to combination therapy in the managed care setting are the addition of the combinations onto a formulary and lack of acceptance among physicians. Fixed-dose combination therapy with a calcium channel blocker and an angiotensin-converting enzyme (ACE) inhibitor can potentially reduce pharmacy costs if given to patients who are taking both drugs independently and to patients who are taking a calcium channel blocker but should probably also be taking an ACE inhibitor (e.g., patients with diabetes or congestive heart failure). A mini-pharmacoeconomic analysis based on claims data over a 6-month period from HMO Colorado, a health maintenance organization, shows that switching patients on dual therapy with a calcium channel blocker and an ACE inhibitor to 1-pill combination therapy with both agents can result in substantial cost savings. PMID- 10538856 TI - Pharmacoeconomics of emerging therapies for rheumatoid arthritis. Based on a presentation by Arthur Kavanaugh, MD. AB - Rheumatoid arthritis is a costly disease. Patients with this condition not only utilize substantial medical resources, but also incur high indirect costs in the form of work disability. These indirect costs are generally much greater than the direct costs. Therefore, a new therapy that is able to control this disease more effectively may be cost effective, even if the direct costs of the therapy itself are high. From a cost-analysis point of view, patients with refractory disease may be good candidates for a new therapy. All costs, direct and indirect, should be considered, and a global, long-term perspective on patient care should be taken. One way to estimate the worth of a therapy based on its ability to improve how patients feel would involve a utility or quality-of-life analysis. PMID- 10538857 TI - Assessing new therapies for RA: defining new expectations for treatment. New therapies in clinical practice: issues of cost and access. PMID- 10538858 TI - The state of behavioral health in managed care. PMID- 10538859 TI - A framework for assessing the effectiveness, efficiency, and equity of behavioral healthcare. AB - OBJECTIVE: To evaluate the effectiveness, efficiency, and equity of behavioral healthcare and to guide an assessment of the current state of the art of behavioral health-oriented health services research. STUDY DESIGN: The framework is grounded in previous conceptual work by the authors in defining a prevention- and outcomes-oriented continuum of healthcare and in identifying and integrating the concepts and methods of health services research and policy analysis for assessing healthcare system performance. PATIENTS AND METHODS: The defining assumptions are that (1) the denominator for behavioral healthcare services must encompass a look at the population, not just the patients, who manifest behavioral health risks; and (2) the delivery system to address these needs must extend beyond acute, treatment-oriented services to include both primary prevention and aftercare services for chronic relapsing conditions. RESULTS: Current policy and practice in behavioral healthcare reveal the absence of a comprehensive, coordinated continuum of care; substantial variation in policy and financial incentives to encourage such development; and poorly defined or articulated outcome goals and objectives. The current state of the art of research in this area reflects considerable imprecision in conceptualizing and measuring the effectiveness, efficiency, and equity criteria. Further, these 3 criteria have not been examined together in evaluating system performance. CONCLUSIONS: The first era of behavioral healthcare focused on cost savings in managed care alternatives; the second is focusing on quality and outcomes; a third must consider the issues of equity and access to behavioral healthcare, especially for the most seriously ill and vulnerable, in an increasingly managed care-dominated public and private policy environment. PMID- 10538860 TI - Implementing effectiveness research and improving care for schizophrenia in real world settings. AB - OBJECTIVE: To review recent advances in medication practices and standards of care in the treatment of schizophrenia and examine the disparity between the knowledge base and clinical practice. DATA SOURCES: Key literature on medication practices, novel pharmacotherapies, and the evolution of practice guidelines for schizophrenia were reviewed. DISCUSSION: Emerging data demonstrate a lack of consistent application of current knowledge and best practices, in part due to major structural inconsistencies in the public mental health system. Implementation of results from effectiveness research as well as the incorporation of practice guidelines may help bridge this gap. CONCLUSION: As standards of care for schizophrenia are developed, the following issues will need particular attention: coordination with the criminal justice system, comprehensive treatment of comorbid illnesses, outcomes based on symptoms in all domains, and continuous and integrated collection of data to produce rational cost justification. PMID- 10538862 TI - Managed public mental healthcare: issues, trends, and prospects. AB - OBJECTIVE: To describe the structure and status of public mental healthcare and the impact of managed behavioral healthcare on this system. STUDY DESIGN AND METHODS: The structure and financing of public mental health systems were reviewed. Because there are no controlled multisite studies of managed public sector behavioral healthcare, case examples were used to illustrate trends and issues. DISCUSSION: The methods, results, and impact of public managed behavioral healthcare are incomplete and uncertain. The complexity of the public sector system, the patients served in it, and the services provided are daunting. The variability of patient needs, the role of Medicaid versus state funding, and the variable governance structures of local systems in different states make managed care methods more complex than in private markets. CONCLUSIONS: The organization, structure, and financing of public mental health systems have developed rapidly in the past generation as care has been moved from hospital to community. Early efforts to apply managed behavioral healthcare methods used in the private, commercially paid sector have not been very successful, and most public sector managed care efforts have been limited to Medicaid-paid care. The trend in public mental health systems is to "unpack" managed care and use its tools selectively. PMID- 10538861 TI - New directions in alcohol and drug treatment under managed care. AB - OBJECTIVE: To examine the potential effects of the introduction and expansion of managed care on the financing and organization of public and private alcohol and drug abuse treatment systems by reviewing studies on managed care and substance abuse. STUDY DESIGN: Spending on treatment for alcohol and drug abuse, the organization of treatment, treatment workforce composition, provision of services, and their implications for access and treatment outcome were examined by review of the treatment literature. RESULTS: Managed care has had major effects on the organization of service delivery, the workforce, and the provision of services. Most of the changes have occurred without the benefit of clinical or policy research. Although managed care has the potential ability to address longstanding problems associated with alcohol and drug treatment, it also presents additional barriers to access and improving treatment outcome. CONCLUSIONS: The review suggests that organizational approaches, particularly the settings in which treatment is placed, will differ in their impact on ties between treatment agencies and the medical community, and ties with other health and social service agencies. Also of importance is a new emphasis on accountability of treatment through the mechanisms of outcomes monitoring and performance indicators. It remains to be seen whether these innovations will be meaningfully linked with outcomes research. It is incumbent on researchers and clinicians to explore these issues. PMID- 10538863 TI - Use of performance standards in behavioral health carve-out contracts among Fortune 500 firms. AB - OBJECTIVE: To determine the prevalence and nature of performance standards in specialty managed behavioral healthcare contracts among Fortune 500 companies. STUDY DESIGN: This was a cross-sectional survey of all companies listed on the Fortune 500 during 1994, 1995, or both. METHODS: From April 1997 to May 1998 we conducted a mailed survey with phone follow-up. Of the 68% of firms that responded, over one third reported carve-out contracts. The survey focused on whether companies had behavioral health carve-out contracts with specialty vendors and characteristics of these contracts, including the use of performance standards. RESULTS: More than three quarters of the Fortune 500 companies reporting specialty behavioral healthcare contracts used at least one performance standard. Most common were administrative standards (70.2%) and customer service standards (69.4%). About half of the companies used quality standards, whereas only a third used provider-related standards. Most (58.8%) companies using performance standards also specified financial consequences. Larger Fortune 500 firms were significantly more likely to use performance standards. Risk contracts and contracts that included all covered employees were also more likely to include some categories of standards. CONCLUSIONS: Administrative and customer service standards may be most common because companies find it easier to specify those standards, especially compared with clinical quality measures. To the extent that employers want to obtain the most value from their behavioral healthcare purchasing, we expect that more will begin to adopt quality standards in their contracts, especially as performance measures become more refined. Reliance on accreditation, however, is an alternative approach for employers. PMID- 10538864 TI - ACHCA recognizes national award recipients at convocation. PMID- 10538865 TI - Eden Alternative Principles hold promise for the future of long-term care. PMID- 10538866 TI - Pets bring therapeutic benefits to nursing homes. PMID- 10538867 TI - Medicaid reimbursement: are you really getting paid? PMID- 10538868 TI - Navigating new directions--finding your true north. PMID- 10538869 TI - Senate panel urges seniors to shop carefully for assisted living. PMID- 10538870 TI - Post-honeymoon, does patient-focused care deliver on its promises? PMID- 10538871 TI - Appointment time: know where your patient is? PMID- 10538873 TI - Survival skills! No discharge without them. PMID- 10538872 TI - Optimism may affect health as much as diet, exercise. PMID- 10538874 TI - Content of preplacement exam depends on goals. PMID- 10538875 TI - Death by questionnaire: quality of life measurement could seriously damage your health. PMID- 10538876 TI - Long-term care for older people: the unanswered questions. PMID- 10538877 TI - Can community leaders' preferences be used to proxy those of the community as a whole? AB - BACKGROUND: Community-based distribution of ivermectin and other drugs requires people in the endemic communities who are capable of distributing the drug. It is essential also to collect information on local people's views concerning different financing mechanisms and approaches to distributing ivermectin. However, studies at household level are resource-intensive. Eliciting the preferences of the community by interviewing a smaller number of community leaders offers an alternative strategy. METHODS: A comparison of information from community leaders and household heads on the financing and distribution of ivermectin through communities was conducted in three communities in Nigeria to determine whether rapidly collected information from key community leaders could represent broad community preferences. RESULTS: The preferences of community leaders and household heads were comparable in relation to the method of collecting payments, managing payments and making payments, who should set the level of payments and the drug distribution mechanisms. However, there were differences between community leaders' views and those of heads of households concerning how the scheme should be supervised. CONCLUSIONS: This study has shown that community leaders' views can only be used as a partial substitute for more laborious methods of data collection insofar as they have the attraction of being quicker and less costly to use. However, they should not be assumed to be identical with the views of the community as a whole. PMID- 10538878 TI - What determines geographical variation in rates of acceptance onto renal replacement therapy in England? AB - OBJECTIVE: To determine the independent effects of need and supply factors on the known geographical variation in acceptance rates onto renal replacement therapy (RRT) in England. METHODS: Data were obtained from all renal units in England on the characteristics of all cases aged 16 years and over, resident in England, who were accepted onto RRT in 1991 and 1992. Of these, 5715 (94.5%) had a valid postcode that could be matched to a census ward. Multilevel modelling using Poisson regression was used. The number of acceptances in each census ward within age bands 16-34, 35-64 and 65+ was the dependent variable. Independent effects modelled were: (1) individual factors (age, sex); (2) census ward need factors- ethnicity (expressed as the percentage of the ward population that was Asian or African-Caribbean), socio-economic deprivation--and supply factors--'access' to the nearest renal unit using crowfly and road travel time and distance, and services available to each ward expressed as number of haemodialysis stations per 100,000 catchment population of the nearest renal unit; (3) district health authority level effects. RESULTS: Age was a major determinant of acceptance, with a 7-fold higher rate in males aged over 64 years compared with younger men. Acceptance rates were lower in females, with a negative age-sex interaction in females aged over 64 years. The percentage of both Asian and African-Caribbean populations per ward was a highly significant positive determinant. Deprivation was also a significant determinant, best represented by a customised index. There was an inverse relation of acceptance with distance, especially road travel time. Other supply side variables had a significant effect though there was no independent district effect. There was some variation in the strength of these relationships by type of area (Greater London, urban and non-urban). CONCLUSIONS: Need and supply factors influence service use as expressed as acceptance onto RRT. Pressure to expand RRT services needs to be aimed at areas with large minority ethnic populations and those living far from existing units. PMID- 10538879 TI - Geographic variations in hospital discharge rates and discretionary hospital use. AB - OBJECTIVES: While past research has shown that there is greater geographic variability in hospital discharge rates for medical conditions where there is less agreement about proper treatment, there is little empirical evidence to support the corollary that high hospital use in a community is primarily the result of a greater volume of such discretionary hospitalizations. This study assesses the contribution of discretionary hospitalizations to higher overall rates of hospital use in communities. METHODS: Hospital discharge files and Medicare eligibility files were used to estimate adjusted rates of hospital discharge, days of care and adjusted mortality for a sample of 761 geographic communities in four states in the USA. Diagnostic information was used to classify hospitalizations into low, moderate and high discretion categories. Correlation and multiple regression analysis methods were used to test for systematic relationships between a community's overall rate of hospital use and discretion-level mix of hospitalizations. RESULTS: Although about half of the variance in overall rates of hospital use among communities was found to be related to the discretion-level mix of hospital discharges, only a small portion of the explained variance could be attributed specifically to community differences in the prevalence of high discretion hospitalizations. CONCLUSIONS: High overall rates of hospital use in communities were not found to be largely the result of high discretion hospital use. PMID- 10538880 TI - Shifting services from secondary to primary care: stakeholders' views of the barriers. AB - OBJECTIVES: To identify the barriers to shifting services from secondary to primary care perceived by the involved stakeholders. METHODS: Forty-five semi structured interviews with stakeholders from primary care, acute and community hospitals, purchasers (health authorities) and other agencies involved in two contrasting initiatives to shift services. RESULTS: Stakeholders perceived similar barriers in the two initiatives: disinvesting from existing providers; lack of information on activity and costs; uncertainty over the quality of the proposed alternative service; concern about an increasing workload in primary care; diversity of views within primary care; difficulties in communication between the many agencies involved; and lack of leadership by purchasers. CONCLUSIONS: Service shifts which involve disinvestment from existing providers and collaboration between agencies with different views and interests will inevitably face a range of barriers. Attempts to shift services by disinvesting from secondary care are likely to encounter the greatest difficulties. Attempts to shift without concomitant disinvestment may also be slow because of the difficulties of multi-agency collaboration. Frustration will be reduced if those involved have a realistic understanding of the difficulties rather than being surprised and overwhelmed by them. PMID- 10538881 TI - Improving the validity of economic evaluations alongside controlled trials. PMID- 10538882 TI - Commissioning health services research: an iterative method. AB - The standard linear method of commissioning research involves many stages, some lengthy. While assessment criteria are usually explicit, their weighting and interaction are not. Output is assessed on completion. This method is suitable where the research question is clear-cut. However, it has drawbacks when the research question and the form and scope of the research are not clear at the outset, as is often the case with research on the delivery and organisation of services. Also, it does not encourage potential users of the research to develop a sense of ownership. An alternative method is proposed by which the scope, form and content of research are not specified in advance but are developed iteratively. A programme director, advised by a group of potential users and research commissioners, has devolved authority to commit funding for the stages of the work as it unfolds, predicated on evolving need. There are foreseeable but avoidable risks of the group over-identifying with the researchers, of research management becoming cumbersome, and of unproductive friction between research groups when they are required to work together. The iterative method, being new and untried, is itself an organisational change requiring evaluation. However, from our local experience, it provides for productive dialogue between research commissioners, researchers and potential users. PMID- 10538883 TI - The third way in health care reform: does the emperor have any clothes? AB - The Labour government elected in the UK in May 1997 has described its approach to the National Health Service (NHS) as a third way in health care reform. This article seeks to analyse the defining features of the third way. It argues that the government has adopted an approach which combines central direction and local autonomy, sanctions and incentives, and planning and competition. The third way therefore entails a cocktail of different approaches and is both electic and pragmatic. In pursuing this policy, the government is seeking to deal with dilemmas that are as old as the NHS itself and is seeking a synthesis between approaches that in the past have been seen as exclusive. It can be argued that the search for a synthesis is unlikely to succeed and the government will have to decide whether it is really committed to a centralised health service in which the main emphasis is on planning and sanctions or whether it is willing genuinely to devolve power to a local level with reliance on incentives and elements of competition. The alternative interpretation is that by making use of a variety of instruments the government is increasing its chances of delivering its objectives. From this perspective, to insist on absolute consistency is to fail to recognise the complexity of the steering mechanisms required in modern public services. In the international context, the third way may find favour as a rhetorical device among politicians seeking to carve out a distinctive niche in the political market place, but its specific characteristics are likely to vary between systems. PMID- 10538884 TI - A review of the use of health status measures in economic evaluation. AB - OBJECTIVE: To review the use of measures of health status in the assessment of benefits in economic evaluation, whether or not the measures were designed for this purpose. METHODS: The review was based on a systematic search of the literature. It provides a comprehensive assessment of the evidence where it exists and a balanced overview of opinion otherwise. RESULTS: Over 3000 papers were identified, of which 632 were found to be relevant. The review provides a set of recommendations. These include: a checklist of questions for selecting a measure for use in economic evaluation; a list of circumstances in which non preference-based measures can be used; and recommendations surrounding the use of health state valuation techniques and multi-attribute utility scales. CONCLUSION: These recommendations should help to identify poor economic evaluations and hence guard against inefficient conclusions being drawn regarding the provision of health services. PMID- 10538885 TI - Evolutionary psychology and health: confronting an evolving paradigm. AB - In much the same way that developments in genetics have opened up new areas of activity in health services, the 'new genetics' has also stimulated a renewal in approaches that try to explain the nature of health behaviours within the context of human biological development. Evolutionary psychology, as an umbrella term for these views, stresses the importance of the brain as an intermediary between genes and individual behaviour. From such a perspective, social context is less important than an understanding of why certain behaviours are 'chosen' by the evolutionary process and how they are predicated on reproductive success. Health policy is a key area where these ideas are likely to become important given evolutionary psychology's focus on the interplay between physiological and psychological factors in determining health behaviours. Health research provides a fertile environment because it is already seeking the hidden biological pathways connecting social status with specific diseases. The challenge represented by evolutionary psychology needs to be taken seriously because of the way in which such ideas mesh with the individualistic basis of much health promotion and health policy. In particular, it poses a challenge when it purports to explain how inequalities in health are not necessarily the result of the unequal distribution of income in society but are natural phenomena. It is also important to engage with such ideas because they increasingly seem likely to occupy the empty ideological space created by the disappearance of politics in policy and as such may have a greater impact than would otherwise be the case. PMID- 10538886 TI - Documentation on trial: nine ways to protect your agency. AB - In today's environment, home care professionals are often overwhelmed with documentation requirements. Licensure and certification surveyors, Office of Inspector General staff, third-party payors, fraud and abuse inspectors, courtroom attorneys, and others seem to scrutinize home care documentation constantly. In short, home care documentation is always "on trial." These tips can help agencies protect themselves in this environment. PMID- 10538888 TI - DMEPOS (Durable Medical Equipment, Prosthetics, Orthotics and Supply) model compliance program: does the OIG have it right? AB - Companies of every size and structure can and should develop an ethical culture of compliance in which prevention, detection, and resolution of abusive practices are "core values." OIG has released a draft Compliance Guideline that includes a seven-element plan to which all agencies must comply. Yet, compliance programs simply are not a "one-size-fits-all" item. PMID- 10538887 TI - Bankruptcy protection and Medicare. AB - The changes in Medicare reimbursement for home health services have forced many agencies to close and still others to face difficult financial times. As a result, home care providers, like many other businesses, have turned to the federal bankruptcy courts in hopes of reorganizing their debts. PMID- 10538889 TI - Mediation: a welcome alternative to a formal PRRB appeal. PMID- 10538890 TI - Protect your agency from year 2000 failures and liability. PMID- 10538891 TI - Interim payment system and risk management. AB - Agencies need to be aware of the legal implications of the Medicare reimbursement system changes in the high-risk areas of patient care. Agencies must take deliberate steps to manage the risks that are sure to ensue under the new system of restricted payments and to minimize liability concerns. PMID- 10538892 TI - Beating the IPS blues. AB - The negative effects of the interim payment system (IPS) are just reaching their peak. The number of providers closing their doors because of IPS could leave elderly and frail people without access to needed services. In just one year, home care has gone from being the darling of Wall Street and the health care industry to the pariah. However, the aspect of IPS that has not been extensively reported is the human cost to the home care community. PMID- 10538893 TI - Home care and the ADA: do limitations on coverage violate the law? AB - To avoid liability, home care agencies must have a firm grasp of the basic requirements of the Americans with Disabilities Act (ADA) and Section 504 of the Rehabilitation Act of 1973 ("Section 504"). Agency leaders must understand the legal issues that arise whenever care is limited or denied to persons with disabilities. PMID- 10538894 TI - Building the Idea Factory. A conversation with John Kao. Interview by Joe Flower. PMID- 10538896 TI - Building ambidextrous organizations. Forming your own "skunk works". PMID- 10538895 TI - Positive turbulence. A climate for creativity. PMID- 10538897 TI - No special gift needed. Generating creative ideas for health care organizations. PMID- 10538898 TI - The process-centered organization: how one rural system made the switch. PMID- 10538899 TI - The obsolescence of independent practice. Part 1: An ocean of physician refugees search for a home. PMID- 10538900 TI - Complexity science: a route through hard times and uncertainty. PMID- 10538901 TI - The pressure is on: tying executive pay to community benefits. PMID- 10538902 TI - The hospital as airport: a new model for health care. PMID- 10538904 TI - Pulling it all together: a single knowledge repository reduces the overwhelming costs of complexity. PMID- 10538903 TI - Seven practices of successful organizations. Part 2: Invest in training, reduce status differences, don't keep secrets. AB - The economic benefits to those firms that really put their people first can be enormous--particularly in industries that are as dependent on people as health care. Extensive research shows that seven specific practices are related to such enhanced organizational performance. In part 1 of this article (in the January/February 1999 issue of this journal), I described four of these practices: employment security, selective hiring, self-managed teams and high compensation. Here, I discuss the remaining three: training, reduction of status differences and sharing information. Knowing what should be done and what it can accomplish should spur you and your colleagues to explore ways of actually implementing this knowledge. PMID- 10538905 TI - Therapeutic breakthroughs in the millennium: what to look for in the next two decades. Part 2: New therapeutics reverse old thinking. PMID- 10538906 TI - In an era of change ... we're changing! PMID- 10538907 TI - One kind word. PMID- 10538908 TI - The future of elder care. PMID- 10538909 TI - Insurance status and the decision to seek a legal opinion for a medical malpractice claim without merit. PMID- 10538910 TI - Health services technology: Part 2, Policy and managerial considerations. PMID- 10538911 TI - School-based clinics: creating children that thrive. PMID- 10538912 TI - Getting yourself motivated. PMID- 10538914 TI - Refugee care. Flight paths. PMID- 10538913 TI - Motivating the minimal performer. PMID- 10538915 TI - Special brew. PMID- 10538916 TI - Hospital information. Touch of class. PMID- 10538917 TI - Data briefing. Global inequalities. PMID- 10538918 TI - Impact of HIV/AIDS on the national economy of India. AB - BACKGROUND INFORMATION: Human immunodeficiency virus/acquired immune deficiency syndrome (HIV/AIDS) is a major public health problem in India. In general, it affects mainly young people who are at their most productive part of life. Despite initial fears that AIDS will be disastrous for the economy, recent experience and estimations have shown that there is a need for reappraisal of its economic impact on society. RESEARCH QUESTION: From the viewpoint of the society of India, what is the total cost and equivalent annual cost of HIV infections for the period 1986-1995 (10 years) in India? METHODS: Type of analysis: Cost descriptive based on predictive modelling cohort analysis using human capital approach. A discount rate of 5% was used. The cost of HIV infections include (i) loss of productivity among HIV patients due to sickness and death, (ii) productivity loss due to caregivers of AIDS patients, and (iii) cost of management of AIDS patients. To estimate the loss of productivity due to premature death attributable to AIDS, a life table approach using two cohorts, one with and one without HIV/AIDS infection at assumed rates was used. The demographic data of 1991 census were used. The difference in the person-years lived in the two scenarios gave the person-years lost due to HIV/AIDS. This was calculated separately for rural and urban areas. To convert this to monetary terms, national per capita income for 1992-93 of Rs. 5529 was used. The data on the days of inpatient care and the cost of management of AIDS patients were based on currently available data and 'expert opinion'. We analysed, using three different sets of assumptions for determination, the low, medium and high estimates of the impact of HIV/AIDS in India. Some of the costs were not included in the present analysis: (i) use of antiviral AZT, (ii) cost of retraining of new workforce, (iii) cost of strengthening of health care system, (iv) cost of research and development, (v) cost of communication activities, (vi) cost of prevention of vertical transmission, and (vii) the intangible cost of pain and suffering to the patients and their families. RESULTS: The total cumulative number of HIV-infected persons in India until 1995 was estimated to be 1.5 million (low estimate), 2.5 million (medium estimate) and 4.5 million (high estimate). The estimated total annual cost (in billion Rupees) of HIV/AIDS in India under low, medium and high assumptions was 6.73, 20.16 and 59.19, respectively. Cost of treatment of AIDS and loss in productivity were the two major components of the cost. CONCLUSIONS: The estimated annual cost of HIV/AIDS appears to be about 1% of the GDP of India if based on high assumptions. However, as mentioned earlier, all costs of HIV have not been taken into account. Its significance has to be assessed in the context of annual growth of GDP (3.5%) and cost of other major diseases in India. PMID- 10538919 TI - Public spending on health care: how are different criteria related? AB - At least nine different criteria are relevant for decisions about public spending for health care. These include economic efficiency criteria (public goods, externalities, catastrophic cost, and cost-effectiveness), ethical reasons (poverty, horizontal and vertical equity, and the rule of rescue), and political considerations (especially demands by the populace). Sometimes one criterion should be examined before another one is considered; that is, they are hierarchically related. Sometimes two criteria will not be compatible but will conflict, forcing difficult choices--particularly between efficiency and equity. Properly thought-out choices of which health care interventions to finance with public funds therefore depend not only on looking at all these criteria, but also on treating them in the appropriate sequence and taking account of their possible inconsistencies. Public funds should finance public and semi-public goods that are cost-effective and for which demand is inadequate; cost-effective interventions that preferentially benefit the poor; and catastrophically costly care, when contributory insurance will not work or there are good reasons to finance insurance publicly. PMID- 10538920 TI - The future of dental managed care in the US. AB - Managed care is slowly becoming more prominent within dental benefit programs, with nearly 30% of insured people participating in a dental managed care plan. This number is expected to increase steadily over time, but not with the same speed that it has with medicine. In combination with the changing demographics of the country and the diminishing supply of dental personnel, managed care will have less power over the dental market place than it has had over medicine. In the future, changes will be dependent on the demand for dental treatment and also the supply of dentists. With increased demand, decreased personnel and a minority of Americans having dental insurance, dentists are in a better position than their medical counterparts. If the influence of managed care is not diminished, the effects on the practice of dentistry will be dramatic. Managed care will eventually introduce outcomes assessment, utilization management, and reduced fees. This will transform the practice of dentistry into a two-tiered system, treating fee-for-service and managed care patients differently. PMID- 10538921 TI - Identification and priority setting for health technology assessment in The Netherlands: actors and activities. AB - This article describes the actual situation at the beginning of 1999 with regard to identification and priority setting for health technology assessment (HTA) on a national level in the Netherlands. For this purpose the literature on HTA published in 1980-1998, mainly national, was thoroughly reviewed. Many policy documents and other reports from the 'grey literature' of identification and priority setting for HTA in the Netherlands were also used. The results show that attempts to identify and set priorities for HTA is a new activity in the Netherlands. The three most important actors in the field are the Health Council, the Council for Health Research and the Health Insurance Council. Methodologies differ depending on the content and scope of each programme. In addition, the methods used are not always transparent and the activities are not co-ordinated. The lack of co-ordination is due to the fact that there is no single organisation that is authorized to identify and set priorities for HTA. Suggestions for improving co-ordination are proposed with the aim of developing a truly national effort in this field, which will enable a more balanced and efficient set of HTA activities. PMID- 10538922 TI - Assessment of biotechnology drugs: what are the issues? AB - Biotechnology is increasingly regarded as an important reservoir for the development of new and innovative, but generally expensive, pharmaceuticals. At the same time, concerns about cost containment have triggered a keen interest in evaluating and comparing the values of diverse health care interventions. In this paper we studied the process of assessment and diffusion of biotechnology drugs by studying three cases, i.e. nebacumab, colony stimulating factors and recombinant human growth hormone. These cases are evaluated in a standardised format, concerning safety, efficacy, cost-effectiveness and ethical, legal and social issues. Many factors that determine the fate of a biotechnology drug seemed to be similar to those of 'classical' drugs. The definition and measurement of clinically relevant outcomes has been identified as a key factor in the assessment process. Another important issue is the relatively small population for the primary indications of biotechnology drugs and the subsequent process of broadening of indications. Paradoxically, the current trend towards evidence-based medicine means that we will increasingly have to make decisions based on 'incomplete' knowledge'. PMID- 10538923 TI - Petitions, public opinion and hospital restructuring in Kitchener-Waterloo. AB - A survey was collected in 1996-97 in which, early in the petition-gathering process by the Save Our Hospital Campaign to prevent the closing of St. Mary's Hospital in Kitchener, Ontario, known petition signers and a general cross section of the public were sampled. The survey data demonstrated that signers of the "Save St. Mary's" petition did indeed have strong views about local health issues. The general public and the petition signers were differentiated not so much by personal health experiences or amount of contact with hospitals as by general concern among petitioners about the erosion of health care together with a conviction that the public should have a strong voice in health care decision making. PMID- 10538924 TI - Determinants of job stress and job satisfaction among supervisory and non supervisory employees in a large Canadian teaching hospital. AB - This article explores the extent to which hospital workers at a large teaching hospital at different managerial/supervisory levels (designated and non designated supervisors, and non-supervisory staff), experienced job stress and job satisfaction prior to the re-engineering of hospital services. For all groups, increased levels of job demands were associated with higher levels of stress. Lower levels of decision latitude were associated with increased job stress for designated supervisors. Increasing levels of decision latitude were associated with both job stress and satisfaction for the other two groups. Co worker support and teamwork contributed to increased job satisfaction for all groups. PMID- 10538925 TI - Regionalization and hospital utilization: Alberta 1991/2-1996/7. AB - BACKGROUND: In February 1994 Alberta Health announced a three-year business plan for the radical restructuring of the health care system in Alberta. The business plan outlined large reductions in funding for acute hospital care spending and the establishment of 17 Regional Health Authorities (RHAs). OBJECTIVES: The objectives of this study are to describe for the period 1991/2 to 1996/7: 1) Trends in overall acute hospital utilization by Alberta residents and residents of each of the 17 RHAs. 2) Trends in the provision of acute hospital services by each of the 17 RHAs and the Alberta Cancer Board. 3) Trends in the transfer of patients between RHAs. RESULTS: Between 1991/2 and 1996/7, the age-sex standardized separation rate, the age-sex standardized average length of stay, and age-sex standardized hospital days rate for Alberta residents fell by 25.6%, 18.7%, and 39.5% respectively. The age-standardized hospital days rate fell in all 17 RHAs. The total number of separations (Alberta residents and non residents) from Alberta acute care facilities fell by 19.6% while the average care intensity for all separations from Alberta acute care facilities rose by 8.7%. The ratio of the highest to lowest average RHA care intensity remained between 1.7 and 1.9 during the study period. RHA self-sufficiency indices increased dramatically in one RHA and remained largely unchanged in the remaining RHAs. RHA import indices decreased for most RHAs. CONCLUSIONS: Large reductions in the use of acute hospital services have occurred in Alberta during the period of major health care restructuring. Further research is needed to examine shifts in services to other sectors and to assess the impact of these reductions on patient outcomes. PMID- 10538927 TI - Manners. "Well, no one would mistake him for Richard the Lionhearted.". PMID- 10538926 TI - Where do we go from here? Preparing staff for hospital closure through education and support. PMID- 10538928 TI - Making sense of waiting lists. PMID- 10538929 TI - Open leadership key to regaining Canadians' trust in the blood system.. Interview by Matthew D. Pavelich. PMID- 10538930 TI - Price survey. Basic wound care prices level off. PMID- 10538931 TI - GPO hopes new name will end 'identity crisis'. PMID- 10538932 TI - Hospital finds many reasons to outsource forms. PMID- 10538933 TI - Strength in numbers: depts. unite to face vendors. PMID- 10538935 TI - Hospital autonomy: the experience of Kenyatta National Hospital. AB - An increasing number of countries are exploring the introduction or expansion of autonomous hospitals as one of the numerous health reforms they are introducing to their health system. Hospital autonomy is one of the forms of decentralization that is focused on a specific institution rather than on a political unit. It has gained much interest because it is an attempt to amalgamate the best elements of the public and private sectors in how a hospital is governed, managed and financed. This paper reviews the key elements of the concept of hospital autonomy, the reasons for its expanded use in many countries and a specific example of making a major teaching hospital autonomous in Kenya. A review of the successful experience of Kenyatta National Hospital and its process of introducing autonomy, with regard to governance, operations and management, and finances, lead to several conclusions on replicability. The legal framework is a critical element for successfully structuring the autonomous hospital. Additionally, success is highly dependent on the extent to which there is adequate funding during the process of attaining autonomy due to the length of the transition period needed. Autonomy must be granted within the context of the national health system and national health objectives and be consistent with those aims and their underlying societal values. Finally, as with decentralization, success is dependent upon the preparation done with the systems and management necessary for the proper governance and operation of autonomous hospitals. PMID- 10538934 TI - Public sector hospitals and organizational change: an agenda for policy analysis. AB - An important feature of health care systems in recent years is the change in the organizational position and relations of public sector hospitals. Health sector reforms have led to increasing heterogeneity in the organizational location and status of public sector hospitals and new organizational forms of public-private relations are being developed by and for hospitals. These changes can have important implications for health and health care. They raise issues around equity, control, accountability and performance of health care. Yet the policy process in practice may be failing to develop and implement appropriate forms of policy formulation on health sector reform. This paper focuses on the organizational position and relations of hospitals within public sector health services. It firstly outlines key elements of health sector reform and relates these to two dimensions of organizational change for hospitals: increasing heterogeneity and forms of public-private relations. The paper provides a descriptive format for classifying forms of hospital organizational change and proposes a framework of six questions for analysing these organizational forms. This may be used to assess the appropriateness of specific policies to particular country situations and to develop more open debate around hospital organizational forms. PMID- 10538936 TI - Rationalizing rural hospital services in Kazakstan. AB - The Soviet health care system placed great emphasis on specialist hospitalization. Primary care, in contrast, was viewed primarily as prophylactic and also identified patients for admission to hospital. This led to long lengths of stays, since patients were provided with outpatient type care in hospital, and unnecessary admissions. The reduction in funding for the health system has exacerbated the top heavy nature of the system and made restructuring of the sector essential. Rural areas in Kazakstan follow a similar structure to other parts of the former Soviet Union. In 1996 a project was undertaken to review the provision of hospital services in one rural rayon (district) just outside Almaty. The approach taken was to emphasize the relationship between activity and financial data. It did this by analysing the link between clinical decisions taken to reduce lengths of stay, management decisions to modify staffing and costs of care. It was shown that substantial savings could be made together with improvements in the quality of care, through a programme of planned restructuring. Some success in inducing change is reported but without a major change in approach to local level management. In order to achieve changes it is important that short and long term alternatives to hospitalization are developed. PMID- 10538937 TI - Indigence and access to health care in sub-Saharan Africa. AB - Access to health care services for the poor and indigent is hampered by current policies of health care financing in sub-Saharan Africa. This paper reviews the issue as it is discussed in the international literature. No real strategies seem to exist for covering the health care of the indigent. Frequently, definitions of poverty and indigence are imprecise, the assessment of indigence is difficult for conceptual and technical reasons, and, therefore, the actual extent of indigence in Africa is not well known. Explicit policies rarely exist, and systematic evaluation of experiences is scarce. Results in terms of adequately identifying the indigent, and of mechanisms to improve indigents' access to health care, are rather deceiving. Policies to reduce poverty, and improve indigents' access to health care, seem to pursue strategies of depoliticizing the issue of social injustice and inequities. The problem is treated in a 'technical' manner, identifying and implementing 'operational' measures of social assistance. This approach, however, cannot resolve the problem of social exclusion, and, consequently, the problem of excluding large parts of African populations from modern health care. Therefore, this approach has to be integrated into a more 'political' approach which is interested in the process of impoverishment, and which addresses the macro-economic and social causes of poverty and inequity. PMID- 10538938 TI - Survey measures nursing home quality. Not-for-profit homes received fewer citations for deficiencies than for-profits. PMID- 10538939 TI - AMA approves product-sale guidelines. Controversial rules direct docs who sell nonprescription health products in their offices. PMID- 10538940 TI - Mercy, Holy Cross form latest Catholic union. PMID- 10538941 TI - Clinton plan would cost providers. President focuses on Medicare reform policy, not politics; generates bipartisan support. PMID- 10538942 TI - E-commerce companies attract converts. Appointments of two purchasing industry veterans add credibility to fledgling companies. PMID- 10538943 TI - Exempla bailing out of HMO business. PMID- 10538944 TI - Inova also putting HMO up for sale. PMID- 10538945 TI - Alike, yet different. Columbia and its spinoffs share strategies, but the new companies work on separate identities. PMID- 10538946 TI - Nine health plans drop Medicare markets. AAHP President Karen Ignagni said HCFA has 'put this program into crisis'. PMID- 10538947 TI - Healthcare struggles with Stark reality. PMID- 10538948 TI - Rethinking home healthcare. A love of the profession doesn't abate thoughts that home care's future may lie in hospitals. PMID- 10538949 TI - Point of contention. Caregivers press for greater needlestick safety, but hospitals worry about the cost. PMID- 10538950 TI - Jury convicts two in Columbia fraud case. AB - Nearly three years after the federal government began a criminal investigation of Columbia/HCA Healthcare Corp. and its executives, a federal jury in Tampa, Fla., found Columbia financial executives Jay Jarrell and Robert Whiteside guilty of defrauding government health programs. A third executive, Michael Neeb, was acquitted in the case, and the jury couldn't reach a verdict on a fourth, Carl Lynn Dick. PMID- 10538951 TI - Technology supports outcomes studies. PMID- 10538952 TI - Round-the-clock education. Hospitals launch Health Channel to offer caregivers continuous access to continuing education. PMID- 10538954 TI - Associations join pro-union ranks. Doc, nurse organizations want to give their members a stronger voice, new services. PMID- 10538953 TI - Moody's gets real with ratings focus. Healthcare changes cause agency to shift its emphasis when evaluating hospital systems. PMID- 10538955 TI - McKesson HBOC slashes revenues. PMID- 10538956 TI - HMA (Health Management Associates, Inc.) shares plunge on earnings news. PMID- 10538957 TI - Hospitals push for special exceptions. PMID- 10538959 TI - Fla. PHO pulls plug, files bankruptcy. PMID- 10538958 TI - Blues leave 'em scrambling. As six plans bail out of the Medicare risk marketplace, critics question their mission. PMID- 10538960 TI - Senate softens patient protections. PMID- 10538961 TI - Driving change in Detroit. Charismatic and untraditional, DMC's new leader plans to turn around a troubled system. PMID- 10538962 TI - A growing concern. Employers' (and employees') frustrations with health plans lead to a new type of business. PMID- 10538963 TI - Private-letter ruling clarifies doc recruiting. PMID- 10538964 TI - Clinton: competition should drive Medicare. PMID- 10538965 TI - Hospitals using lenders of last resort. Some struggling providers say nontraditional financiers are abandoning them. AB - Hospitals and health systems are increasingly turning to nontraditional lenders to sell their receivables or back multimillion-dollar sales of troubled operations. But as some recent incidents show, the relationships with those financiers also can be troubled. Some critics contend that these lenders of last resort can be prone to panic. PMID- 10538966 TI - States grapple with risk and oversight. PMID- 10538967 TI - Conversion eases financial woes. Public facility is in the black after converting to private operation, escaping bureaucracy. PMID- 10538968 TI - House to vote on medication deduction. PMID- 10538969 TI - Striking out on their own. New "no-hospital" companies joining the game, but so far they're battling close to zero. PMID- 10538970 TI - High standards: ISO 9000 comes to health care. AB - ISO 9000, an international quality system, is catching on in health care. Providers see it as a much less expensive alternative or as a complement to the JCAHO. And because ISO has been used for years in manufacturing, it has one big advantage over JCAHO: employers know it and trust it. PMID- 10538972 TI - Net gain or net loss? Health care consumers become Internet savvy. AB - The Internet is overloaded with health care information, and consumers can't seem to get enough of it. As a result, patients are now more knowledgeable about their treatment options when they meet with doctors. That's the good news; the bad news is that the doctors themselves must sort through all the sometimes contradictory, sometimes inaccurate information. PMID- 10538971 TI - Defining healthy communities: the 1999 NOVA Awards. AB - "A healthy community isn't defined just by its rates of infection or inoculation," says Lawrence White Jr., CEO of St. Patrick Hospital, a winner of this year's NOVA Awards. Working with a wide array of community and health care organizations, the five winning hospitals have tackled knotty community problems- even ones not directly related to health care--in ways that reach beyond their traditional turf. PMID- 10538973 TI - Family caregivers: hospitals' most vulnerable partners. PMID- 10538974 TI - Partners. A common denominator. PMID- 10538976 TI - Community health. Taking it to the streets. PMID- 10538977 TI - Hospitals--designing inclusive places. PMID- 10538975 TI - Homeless patients: bridging the gap between hospital and health. PMID- 10538978 TI - Doernbecher Children's Hospital, Portland, Oregon. PMID- 10538979 TI - Celebration Health, Celebration, Florida. PMID- 10538980 TI - New York Psychiatric Institute, New York City. PMID- 10538981 TI - The CORE Center, Chicago, Illinois. PMID- 10538983 TI - How to minimize the legal risks of PACS and teleradiology. PMID- 10538982 TI - NCI-backed project aims for stronger radiology research. PMID- 10538984 TI - From RAM to RAID: digital storage options for PACS. PMID- 10538985 TI - The face of radiology's future. PMID- 10538986 TI - How to evaluate educational radiology sites on the Web. PMID- 10538987 TI - Latex debate: it's not just academic. PMID- 10538988 TI - Holy cow! What's so sacred about cover gowns? PMID- 10538989 TI - Performance art: benchmarking OR efficiency. PMID- 10538990 TI - Hoarding: the real Y2K bugaboo. PMID- 10538991 TI - Tools of the trade: from the stone age to the computer age. PMID- 10538993 TI - Stats. Lean times loom: BBA study. PMID- 10538992 TI - 7 ways to tap into a tight labor market. PMID- 10538994 TI - Compensation monitor. Study shatters big myth about primary care compensation. PMID- 10538995 TI - PPOs: a better brand of managed care? PMID- 10538996 TI - Antifraud push means plans need to focus on compliance. PMID- 10538997 TI - 'E' equals managed care squarely facing the future. PMID- 10538998 TI - The status quo must go!. Interview by Patrick Mullen. PMID- 10538999 TI - DM's motivation factor can skew study results. PMID- 10539000 TI - Is taking a class in ethics cruel, unusual punishment? PMID- 10539001 TI - Contract talks are like dating: ask those important questions. PMID- 10539002 TI - Managed care outlook. When medical necessity is disputed. PMID- 10539003 TI - As all-products clauses spread, physicians make tough choices. PMID- 10539005 TI - Responsible communication: a challenge to act with integrity. PMID- 10539004 TI - Civil rights claims and physician peer review. PMID- 10539006 TI - How bad faith lawsuits can be tailored to fit HMOs. AB - In January 1999, in the case of Goodrich v. Aetna, a California jury returned a record-breaking verdict of $120.5 million damages in favor of a widow as a result of Aetna's failure to act in good faith in the treatment of her terminally ill husband. The following article discusses the history and basis of this kind of lawsuit, major decisions specifically pertaining to HMOs, and the outlook for future liability in this area. PMID- 10539007 TI - Standardized scoring for group decision making. PMID- 10539008 TI - FPA, MedPartners, and claims-made malpractice insurance. PMID- 10539009 TI - Health care accounting: half-truths and sleight-of-hand. PMID- 10539010 TI - From paper charts to the spoken electronic medical record. PMID- 10539011 TI - Contact capitation for specialists. AB - Though still a relatively young concept, contact capitation has been successfully implemented in provider networks around the country. As a payment methodology, it holds the potential of offering broad physician panels, reduced medical management hassles, greater physician control of clinical decision-making and financial stability for practitioners. Contact capitation allows specialists to regain their voice in clinical decisions and offers them greater financial stability in risk sharing arrangements. PMID- 10539012 TI - What to do if you're behind on your payroll taxes. PMID- 10539013 TI - Kansas ruling threatens peer review. Adams v. St. Francis Medical Center. PMID- 10539014 TI - Magic bullets. PMID- 10539015 TI - The way it is, is not the way it will be. PMID- 10539016 TI - A brave new medicine. A conversation with William Haseltine.. Interview by Joe Flower. PMID- 10539017 TI - Age cannot wither: the golden age of the golden years. PMID- 10539018 TI - Nanomedicine. PMID- 10539019 TI - You gotta have heart: the future of cardiology. PMID- 10539020 TI - Paying for the biomedical future. PMID- 10539021 TI - Thinking in the future tense. PMID- 10539022 TI - The 1 percent problem. The organization of medicine has changed little in a century. PMID- 10539023 TI - No shots, no school. Minneapolis immunizes school-age children. PMID- 10539024 TI - Quantum leaps in healing. PMID- 10539026 TI - Diversity, compassion & exceptional leadership. Attributes of the 1999 emerging leaders. PMID- 10539025 TI - Quality care begins with patient-friendly environments. PMID- 10539027 TI - Making history. PMID- 10539028 TI - Contact capitation helps control specialists' costs. PMID- 10539029 TI - Integration the right strategy, despite health executives' increasing apprehensions. PMID- 10539030 TI - Physicians need to expect less from PPMCs, take more responsibility for practice success. PMID- 10539031 TI - Inspiring patient, employee satisfaction turns Florida hospital into top performer. PMID- 10539032 TI - Trends indicate 'wise' integrated systems should embrace complementary medicine. PMID- 10539033 TI - Hospital rating systems. Quality indicators and specious inferences. PMID- 10539034 TI - Hospital rating systems. HealthCareReportCards.com: valuable consumer information. PMID- 10539035 TI - What does a 'top 50' hospital look like? AB - A new guide rating U.S. hospitals, developed by The Center for HealthCare Industry Performance Studies (CHIPS), debunks some myths and offers some surprises. One surprise is the outstanding performance by a public hospital in Denver. Ed Egger analyzes the factors in the success of Denver Health Medical Center. PMID- 10539036 TI - Building physician/hospital partnerships: top 10 lessons. PMID- 10539037 TI - HCIA expands '100 Top Hospitals' program to include clinical research. AB - For six years, HCIA, Inc. has identified 100 hospitals that set benchmark standards of superior performance in management, patient care, and cost efficiency. Now this health care data management firm has expanded the program to include clinical research and identify best practices of top hospitals. PMID- 10539038 TI - Rural hospital turnaround comes from planning, winning support, collaborating. AB - Eleven years ago, rural Bladen County Hospital in North Carolina was on the verge of closing. Since then, its leaders have used planning, collaborating, and winning community support to implement many new initiatives and develop an award winning rural health network. PMID- 10539039 TI - Prioritists offer suggestions for hospital survival in current crisis environment. Interview by Ed Egger. PMID- 10539040 TI - Security now a burning issue as Internet becomes more than marketing tool. PMID- 10539041 TI - Effective physician integration strategy. PMID- 10539042 TI - Strategists face tough business decisions. PMID- 10539043 TI - What physicians want. PMID- 10539044 TI - Maximizing the role of your physician executive. PMID- 10539045 TI - Engaging physicians in true strategic partnership. PMID- 10539046 TI - Congress wrap-up ... ACHE's 1999 Congress on Healthcare Management. PMID- 10539047 TI - Evaluating executive compensation packages. PMID- 10539048 TI - ACHE's new strategic plan. PMID- 10539049 TI - Moving into management. PMID- 10539050 TI - Avoiding e-mail pitfalls. PMID- 10539051 TI - The ethics of managed care. PMID- 10539052 TI - Mystery shopping in your organization. PMID- 10539053 TI - The latest Medicare reform plan. PMID- 10539054 TI - Facts in court decision illustrate basis of OIG's and HCFA's EMTALA concern. Consumer Health Foundation v. Potomac Hospital. PMID- 10539055 TI - Court resolves federal-state law conflict. Seittelman v. Sabol. PMID- 10539056 TI - No ADA violation in case of HIV positive surgical technician. Mauro v. Borgess Medical Center. PMID- 10539057 TI - The future of Commendation. PMID- 10539058 TI - Top ten long-term care recommendations. PMID- 10539059 TI - Complying with challenging ambulatory care standards. PMID- 10539060 TI - Optional maintenance plan added to SOC (Statement of Conditions). PMID- 10539061 TI - Searching for effective alternatives to restraint use. PMID- 10539062 TI - Making the most of standards interpretations. PMID- 10539063 TI - PMCC (Performance Measurement Coordinating Council): integrating performance measurement efforts. PMID- 10539064 TI - Protecting personal health information. PMID- 10539065 TI - Outpatient payment impact could sock hospitals hard. PMID- 10539066 TI - CDC guidelines on surgical infection first since 1985. PMID- 10539067 TI - Highlights of CDC's recommendations. Centers for Disease Control and Prevention. PMID- 10539068 TI - Tips on orienting new OR staff. OR roundtable. PMID- 10539069 TI - Orientation stresses self-learning. PMID- 10539070 TI - Top performers in heart surgery. PMID- 10539071 TI - RNs are key in cardiac outcomes. PMID- 10539072 TI - Bringing diversity into the open. PMID- 10539073 TI - When will you be Y2K capable? PMID- 10539074 TI - OSHA plans action on sharps injuries. PMID- 10539075 TI - Reuse pow-wow resolves little. PMID- 10539076 TI - Unlocking the door to corporate compliance. PMID- 10539077 TI - Focusing admissions on resident needs. PMID- 10539078 TI - Mission creates winning approach to care. PMID- 10539079 TI - Rewriting the book on quality. PMID- 10539080 TI - Transforming data into quality. PMID- 10539081 TI - Setting best practices in motion. PMID- 10539082 TI - Earnings announcements. Skilled nursing facility chains; assisted living facility chains. PMID- 10539083 TI - Veterans' bill omits ban on bed glut. PMID- 10539084 TI - Jostling for elbow room. As competition intensifies, assisted living companies broaden their services and look for new markets. PMID- 10539085 TI - Top 30 assisted living chains. Mergers, expansion drive shift in assisted living rankings. PMID- 10539086 TI - Top 50 nursing facility chains. PMID- 10539087 TI - A few words with David Banks. Interview by Markian Hawryluk and Lynn Wagner. PMID- 10539088 TI - Keeping damages to a minimum. PMID- 10539089 TI - Beware of hospitals bearing gifts. PMID- 10539090 TI - No room for error. PMID- 10539091 TI - Avoiding unnecessary medical reviews. PMID- 10539092 TI - Time to abandon electroconvulsion as a treatment in modern psychiatry. AB - This review examines the evidence for the current use of electroconvulsive therapy (ECT) in psychiatry. The history of ECT is discussed because ECT emerged with no scientific evidence, and the absence of other suitable therapy for psychiatric illness was decisive in its adoption as a treatment. Evidence for the current recommendation of ECT in psychiatry is reconsidered. We suggest that ECT is an unscientific treatment and a symbol of authority of the old psychiatry. ECT is not necessary as a treatment modality in the modern practice of psychiatry. PMID- 10539093 TI - Efficacy and mode of action of an immunomodulator herbal preparation containing Echinacea, wild indigo, and white cedar. AB - Using the example of an allopathic herbal combined preparation containing Echinacea root, wild indigo root, and white cedar leaf tips (Echinaceae radix + Baptisiae tinctoriae radix + Thujae occidentalis herba = Esberitox N), the efficacy and mode of action of a phytoimmunomodulator, or immune system enhancer, is described. Efficacy of the immunomodulator has been demonstrated in studies of acute viral respiratory tract infections and infections requiring antibiotic therapy. In a recent study compliant to GCP, the therapeutic superiority of the herbal immunomodulator over placebo was confirmed as statistically significant and clinically relevant. The present overview describes a model of the antigen independent mode of action of phytoimmunomodulation ("immunobalancing"). PMID- 10539094 TI - Results of laboratory tests for improving risk adjustment for comparisons of hospitals' outcomes. PMID- 10539095 TI - Evaluating test performance criteria: concepts and practices. Part 3: Examining the dynamics of the normal distribution function applied to measurement error probability. PMID- 10539096 TI - Particle size measurement in suspensions: Part 1--A laboratory method for exploring food allergies and sensitivities in illness. PMID- 10539097 TI - Typical applications of molecular diagnostics in disease. PMID- 10539098 TI - A cost-effective system for paraffin-quality frozen sections. PMID- 10539099 TI - Determination of LD-Ig complex with counter immunoelectrophoresis using terylene cellulose acetate membrane as supporting media. AB - OBJECTIVE: In order to determine abnormal LD isoenzyme bands in serum, we established a method of counter immunoelectrophoresis using terylene cellulose acetate as supporting media. SETTING: Department of Clinical Laboratory, Southwestern Hospital, Chongqing 400038 China. PRACTICE DESCRIPTION: LD isoenzymes were electrophoretically fractionated with a Paragon electrophoretic system. LD anomaly was identified by counter immunoelectrophoresis using terylene cellulose acetate as supporting media. MAIN OUTCOME MEASUREMENTS: Identification of the LD-Ig complex using a simple and practical method. RESULTS: An abnormal LD 4 band and an extra band on the cathodic side of LD-5 were observed in the serum of a post-burn patient and proved to be an LD-IgG complex. CONCLUSION: We have established a useful and efficient protocol that uses both isoenzyme electrophoresis and counter immunoelectrophoresis to identify abnormal isoenzyme bands. The method of counter immunoelectrophoresis we describe, using terylene cellulose acetate as supporting media, is simple and suitable for routine clinical use. PMID- 10539101 TI - Ergonomics in the clinical laboratory. AB - Ergonomics can improve work quality, increase productivity, raise morale, reduce absenteeism, and reduce workers compensation. Laboratory managers are responsible for recognizing employees at risk of developing CTSs which represent 65 percent of all injuries reported. It is a wise investment to prevent injuries. It needs to be stressed to laboratorians that just because one is not suffering now, does not mean suffering will not occur in the future. All of these micro- (mini-) traumas add up over the years. Once there is pain, there are things that can be done to alleviate the discomfort or disability, but that part of the body is never the same again. CTDs are not inexpensive; the National Council on Compensation Insurance reports the average CTD victim compensation is $29,000. Policies and procedures are needed in every facility to address ergonomic safety issues. All employees should be educated about tasks or situations putting an individual at risk for CTDs. Employees should communicate with management and management should be open to information about potential or real risk situations. Safety is everyone's job. PMID- 10539100 TI - Drawing specimens for coagulation testing: is a second tube necessary? AB - Three recent studies discussed the possibility that the National Committee for Clinical Laboratory Standards (NCCLS) recommendations that the coagulation specimen should be the second or third tube collected are unnecessary. However, only one reagent/instrument was used in each study. Our protocol differed from the previous studies because we performed the assays on three different reagent/instrument systems on the same samples. Our study used photo-optic, mechanical, and nephelometric systems of clot detection. After obtaining informed consent, we obtained two blue-stoppered tubes of blood from 95 subjects: 15 normal patients and 80 patients currently on coumadin therapy. No discard tube was drawn for coagulation testing. A prothrombin time with an international normalized ratio and an activated partial thromboplastin time, were performed on each tube. Laboratory One used a MLA 1600C (Hemoliance) with Thromboplastin DS (Pacific-Hemostasis, ISI of 1.11) and APTT-LS (Pacific-Hemostasis). Laboratory Two used an STA (Diagnostica-Stago) with Neoplastine CI+ (Diagnostica-Stago, ISI of 1.14) and PTT-LT (Diagnostica-Stago). Laboratory Three used an ACL 300 with Plastinex (Biodata, ISI of 1.67) and Actin FSL (Dade Behring). No clinical or statistically significant differences were seen between the first or second tubes on any of the three reagent/instrument combinations in the PT in seconds, international normalized ratio reporting, or APTT results. Our results indicate that the NCCLS guidelines for obtaining a second tube when performing coagulation testing should be considered for elimination when new revisions are published. PMID- 10539102 TI - Microwaves in the laboratory: effective decontamination. AB - OBJECTIVE: We hypothesize that microwave irradiation of certain contaminated materials typically used in a clinical laboratory or a home healthcare setting could produce efficient and effective sterilization when compared to standard autoclave methods. DESIGN: A standard household carousel microwave oven unit used at the High setting was employed to expose certain materials that had been contaminated with either bacteria or yeast to microwaves for specific intervals of time. Following each time interval, materials were checked for effectiveness of decontamination using standard culture techniques and colony counting. Additionally, powdered media was prepared and microwave irradiated for specific times. The media was then poured into plates and checked for microbial contamination; another set of plates was examined to determine the ability of the irradiated media to support bacterial growth. SETTING: This study was carried out at Texas Tech University Health Sciences Center in Lubbock TX. MAIN OUTCOME MEASURE: Standard culture and colony counting techniques were used to determine the efficacy of microwave sterilization. RESULTS: The study indicated that microwave irradiation provided effective and efficient sterilization of all materials tested. Of the bacteria studied, only E. coli survived beyond 30 seconds of microwave exposure. Yeast did not survive beyond 15 seconds of microwave exposure. Swabs and gauze contaminated with bacteria or yeast were completely sterilized after 30 seconds. After three minutes in the microwave oven, powdered, prepared media was free of contamination while able to support growth when inoculated with S. aureus. CONCLUSION: We conclude that a household carousel microwave oven unit can provide fast, effective sterilization of certain contaminated materials typically used in a clinical laboratory, student laboratory, or home healthcare setting. PMID- 10539103 TI - Critical reading and writing across the curriculum in clinical laboratory science. PMID- 10539104 TI - Drug delivery: the strategic advantage of design controls. AB - An interesting dichotomy exists in the regulation of drug-delivery products. If they are regulated as devices, there is a strong likelihood that new products will be designed and developed under documented design control programmes. There is less chance of this if they are regulated as drugs. This article discusses the benefits and added value of designing and developing all drug delivery products under a documented design control programme. PMID- 10539105 TI - Consolidation and its consequences. AB - A small market place dominated by large companies is becoming the new business environment for the industry. But this will not result in stasis. Large companies will continually analyse and restructure their portfolios and niche companies will emerge and be acquired. This article explores current industry dynamics and their effects. PMID- 10539106 TI - Latest advances in nasal drug-delivery technology. AB - Nasal delivery devices are not only limited to local medical therapy. The nasal route is an alternative to invasive or oral drug administration. The penetration of bioactive molecules through the nasal mucosa has been shown to achieve good bioavailability, and nasal sprays offer patients greater convenience. This article reviews some of the latest devices for liquid and dry-powder formulations, including preservative-free systems, and the business benefits they offer. PMID- 10539107 TI - Patient pull: the changing influences on medical device design, Part I. AB - The medical device and pharmaceutical industries are in the process of a major paradigm shift, driven by increasing levels of regulatory control, cost containment and patient involvement in treatment programmes. The role of the patient is changing from passive participant to active partner, and this will have a significant impact on the design and development of medical devices. Part I of this two-part article discusses the key drivers of change, the implications for various stakeholders and how these will influence the design of medical devices. PMID- 10539108 TI - An insider's guide to knowledge transfer. From Academia to industry. AB - Higher education institutions (HEIs) can improve the quality of their research through working with industry. Industry can develop more profitable products as a result of working with HEIs. This article explores the process of forming a rewarding partnership and lists the dos and don'ts for academic and commercial people seeking to form a partnership with the "other side." PMID- 10539109 TI - What's going on in Germany? AB - The issues that are currently critical to business in Germany are highlighted in this round-up of news from BVMed, the federal association for the medical device industry in Germany. A huge overhaul of the health-care system has just been proposed, and this is being hotly debated by all sectors. The industry view is highlighted here. PMID- 10539110 TI - Drug-device products. AB - A drug-device product is generally regulated by the Medical Device Directive or by the Medicinal Product Directive. But for some types of product, cross references are made to specific requirements in both Directives. This article provides an overview of the regulation of these products and the conformity assessment procedures that are required. PMID- 10539111 TI - The parting of the ways? How biomaterials may differ from advanced materials in the next century. AB - A recent symposium on the future role of materials science in some critical industries suggests that the historical reliance of the medical device industry on the advances made in conventional materials development may not be sustained in the next few decades. PMID- 10539112 TI - Dry platelets with the Dideco Excel apparatus. AB - Dry platelets are required to prevent hemolytic and nonhemolytic febrile reactions after transfusion to ABO mismatched recipients, to reduce the risk of HLA immunization and to prevent allergic or anaphylactic reactions. Previously we have shown that the collection of single donor platelets almost free of plasma is possible with different cell separators. With the systems we described for the CS 3000+/Amicus and AS 104-204 apparatuses collection of dry platelets implies their resuspension in non plasma solutions after opening the circuit. With the new system developed by Dideco for the Excel apparatus this moreover is no longer required since the platelet bag can be connected to the resuspending medium through a dedicated line with an antibacterial filter. Platelets are collected cyclically and the resuspending solution is added when the procedure is over. In this study 53 collections were evaluated, 21 of which were erythrothrombocytapheresis. In 60-65 min 4.21 of anticoagulated blood (1/12) were processed with platelet collection automatically done after 6-700 ml cycles. The platelet yield averaged 4.67 +/- 0.7 x 10(11), with a platelet efficiency per minute of 7.18 +/- 0.9 x 10(9). The WBC contamination averaged 2.6 +/- 0.7 x 10(5) and contamination did not exceed 0.87 x 10(6). The quality of platelets was satisfactory as measured by aggregation, morphology score, and CD 62 membrane glycoprotein externalization. These results were comparable to those obtained with three other Excel apparatuses used in the conventional way to collect platelets resuspended in plasma or with the Amicus used to collect dry platelets using an open system. PMID- 10539113 TI - Quantitative PCR for counting residual white blood cells in blood products. AB - Leukocyte depleted blood components are frequently used to reduce alloimmunization and the risk of transfusion transmitted infection. Counting residual white blood cells in filtered blood products requires sensitive and reliable techniques. After separation of white blood cells from 500 microliters of 20 non-filtered and 54 filtered blood products we used polymerase chain reaction (PCR) and fluorimetric detection for the quantification of genomic DNA. The results were compared with results from Nageotte chamber counting. The accurate limit of detection of PCR was determined at 1 WBC/microliter (intra assay coefficient of variation: 16.3%). PCR correlated well with Nageotte chamber counts (r = 0.77, p < 0.001, n = 74). Concordant results were obtained in 51 filtered and 20 non-filtered blood products. Discrepant results were obtained in 3 filtered whole blood units: In these blood products > 12 WBC/microliters were counted in Nageotte chamber and PCR gave a negative result. After component preparation fresh-frozen plasma and red cell concentrates of these units contained < 1 WBC/microliter using both methods. In conclusion we describe a quantitative PCR method which had about the same sensitivity and specificity as Nageotte chamber testing. However, PCR is more laborious than the standard method. As well, as reliable PCR testing requires expensive instruments and staff experienced in molecular biology, the standard method is more cost effective. PMID- 10539114 TI - Annexin V and platelet antigen expression is not altered during storage of platelet concentrates obtained with the AMICUS cell separator. AB - During storage of platelet concentrate the so-called "storage lesion" occurs. During this time, platelets loose their morphological and functional capacities that are necessary for proper in vivo efficacy following transfusion. Annexin V represents a marker for apoptosis. In this study, Annexin V and additional antigens were analyzed by flow cytometry. Platelet concentrates were obtained with a new cell separator (AMICUS Separator, Fenwal). Following apheresis, platelet units were stored for an experimentally prolonged time of seven days. Daily aliquots of the platelet-rich plasma were obtained to measure Annexin V and platelet antigens CD62p, CD63, CD41a, CD42b, and the binding of fibrinogen. All analyses were performed using flow cytometry. During storage, no significant changes in mean channel fluorescence intensity (MCFI) of CD41a (P = 0.99) and CD42b (P = 0.29), percentage of CD62p+ and CD63+ platelets (P = 0.23 for CD62p; P = 0.52 for CD63), and the binding of fibrinogen to platelets occurred (P = 0.85). Also, the expression of Annexin V remained constant with no significant change (P = 0.36). This study shows that antigens of platelets, obtained with the AMICUS cell separator are well preserved during storage. Regarding Annexin V, no obvious signs of apoptosis can be detected by flow cytometry. These findings demonstrate the high degree of biocompatibility of the apheresis device and storage container. PMID- 10539116 TI - Flow cytometric analysis of platelet function in stored platelet concentrates. AB - Platelet activation occurs during the collection, processing and storage of platelet concentrates. The effect of the platelet activation on the functional state of stored platelets remains however undefined. We employed flow cytometric analysis to evaluate the extent of platelet activation and the physiological response to thrombin stimulation of platelets stored for up to five days under routine blood bank conditions. Platelet surface expression of the activation markers CD62 and CD63 was examined, along with modulation of platelet membrane glycoproteins (GP) Ib and IIbIIIa. Platelet dense granule content was determined using a mepacrine uptake assay and the extent of platelet microparticle generation was quantified. Thirteen random-donor platelet concentrates prepared under routine conditions by a platelet-rich-plasma protocol were examined. Platelets were found to be activated following preparation on day 1. Although a gradual increase was seen with increasing storage time, this was not statistically significant for CD62 or CD63 expression, GPIIbIIIa or GPIb modulation or dense granule release; the generation of platelet microparticles did, however, increase with increasing storage time. The characteristic increase in surface expression of CD62, CD63 and GPIIbIIIa and decrease in GPIb and dense granule content in response to thrombin stimulation was observed with all concentrates, but these measures of platelet functional reserve showed decreasing platelet function with increasing storage time. The results indicate that platelets are activated by day 1, likely as a consequence of manipulation during collection and processing, but are not further progressively activated with increasing storage time; they do, however, become relatively hypofunctional with increasing storage. PMID- 10539115 TI - Multiple red blood cell antibodies produced by donor B lymphocytes after ABO matched allogeneic bone marrow transplantation. AB - A 50-year-old man with AML[M2,t(8;21)] underwent BMT from his younger sister. At that time, he had no unexpected antibody and his blood type was O(+), CcDEe. The type of Kidd was not examined. The donor's blood type was O(+), CCDee, Jk(a+b-). One year after the BMT, the patient's blood type had changed to that of the donor's and anti-E antibody was detected. Despite the use of platelet concentrates (PCs) only, anti-c antibody was later identified. We conclude that there is a need to check red cell antibodies at regular intervals, even when using PCs only, for earlier detection of unexpected antibodies after BMT. PMID- 10539117 TI - What's happening? Cord blood derived stem cells: current views. PMID- 10539118 TI - Using healthcare claims data for outcomes research and pharmacoeconomic analyses. AB - Healthcare claims data are a practical complement to data from randomised controlled trials (RCTs) for evaluating health outcomes in non-experimental settings and for generalising results to a broader population. Claims data are a relatively inexpensive way to obtain useful information about patient demographics, as well as healthcare resources used for specific medical conditions and procedures from large numbers of patients over extended periods of time. With claims data, it is possible to identify patients who meet specific medical or sociodemographic criteria, estimate their costs, define episodes of medical care, and measure outcomes more globally than is possible with RCT data. Statistical methods exist to address some of the inherent issues with claims data due to their limited clinical detail. We also identify extensions of claims data to productivity issues, the use of centralised claims data such as in Canada, and the application of new statistical methods to outcomes research literature such as sample selection correction methods. PMID- 10539120 TI - Measuring sensitivity in pharmacoeconomic studies. Refining point sensitivity and range sensitivity by incorporating probability distributions. AB - OBJECTIVE: The aim of the present study is to describe a refinement of a previously presented method, based on the concept of point sensitivity, to deal with uncertainty in economic studies. DESIGN: The original method was refined by the incorporation of probability distributions which allow a more accurate assessment of the level of uncertainty in the model. In addition, a bootstrap method was used to create a probability distribution for a fixed input variable based on a limited number of data points. The original method was limited in that the sensitivity measurement was based on a uniform distribution of the variables and that the overall sensitivity measure was based on a subjectively chosen range which excludes the impact of values outside the range on the overall sensitivity. PATIENTS AND PARTICIPANTS: The concepts of the refined method were illustrated using a Markov model of depression. MAIN OUTCOME MEASURES AND RESULTS: The application of the refined method substantially changed the ranking of the most sensitive variables compared with the original method. The response rate became the most sensitive variable instead of the 'per diem' for hospitalisation. CONCLUSIONS: The refinement of the original method yields sensitivity outcomes, which greater reflect the real uncertainty in economic studies. PMID- 10539119 TI - Benefit valuation in economic evaluation of cancer therapies. A systematic review of the published literature. AB - Generic measures of benefit which employ individuals' preferences, such as the quality-adjusted life-year (QALY), are, in principle, the most appropriate outcome measure to use in the economic evaluation of cancer therapies. They can reflect the trade-offs between health-related quality of life (HR-QOL) and length of life, between different dimensions of HR-QOL and between the process (e.g. convenience) and outcome characteristics of treatments. This paper reviews the methods literature on preference-based measures of benefit in economic evaluation, with the aim of establishing good practice for applied studies using these methods. A systematic review of applied economic evaluations of cancer therapies which have used these types of benefit measure was performed with the aim of establishing whether studies in this area adhere to good practice. In total, 29 studies were reviewed, and the results showed that, in general, good methods are not being adopted. This may be due, in part, to authors not having the space in journals to detail their methods fully, but it is likely also to reflect the fact that good methods for the use of preference-based measures of benefit in economic evaluation have not been adequately disseminated. PMID- 10539121 TI - Economic analysis of carboplatin versus cisplatin in lung and ovarian cancer. AB - OBJECTIVE: To conduct an economic analysis on the use of carboplatin versus cisplatin over multiple courses in patients with lung [nonsmall cell lung cancer (NSCLC) and small cell lung cancer (SCLC)] or ovarian cancer. DESIGN: This 1-year study was a prospective, multicentre, cost-minimisation evaluation. Direct medical resource utilisation and costs associated with carboplatin and cisplatin administration over 3 to 6 courses of treatment were measured and compared. The perspective of this evaluation was that of the payer. SETTING: A convenience sample of 16 sites representing a mix of cancer centres, outpatient clinics, medical centres and managed-care sites in a general practice oncology setting participated. PATIENTS AND INTERVENTIONS: Patients were included in this study if they were newly diagnosed with NSCLC, SCLC or ovarian cancer, had not received prior chemotherapy, received either carboplatin or cisplatin as their treatment (additional chemotherapy agents were allowed), and received at least 3 courses of carboplatin or cisplatin therapy up to a maximum of 6 courses. Patients receiving more than 6 courses of therapy were included in this study, but data collection on those patients stopped after the sixth course. Individuals involved with data collection at all sites were trained via on-site and/or teleconference training. Site visits were made to assure reliability of at least 0.80. Data were collected and compiled via a fax transmission process that scans directly through optical mark and character recognition into a computer database. Outcome measures included costs of: medications, emergency room visits, physician/clinic/laboratory visits, home healthcare visits, transfusions, special procedures, consultations, hospitalisations and other/miscellaneous costs. MAIN OUTCOME MEASURES AND RESULTS: Of 220 patients, 164 met the study criteria (response rate = 74.2%) with 95 patients in the carboplatin group (NSCLC = 45, SCLC = 18, ovarian = 32) and 69 in the cisplatin group (NSCLC = 36, SCLC = 21, ovarian = 12). The average number of courses were: NSCLC = 4.3 and 4.2, SCLC = 4.3 and 4.8, and ovarian = 4.7 and 5.1, respectively, for carboplatin and cisplatin. The total costs (treatment and toxicity) associated with the use of carboplatin were higher in NSCLC, similar in SCLC but lower in ovarian cancer. CONCLUSIONS: These results indicate that overall treatment costs may vary depending on cancer type, even when the same drugs are used. The total costs (treatment plus toxicity costs) associated with the use of carboplatin were higher than those of cisplatin in patients with NSCLC, similar in SCLC, but lower in ovarian cancer. PMID- 10539122 TI - Cost of illness and disease severity in a cohort of French patients with Parkinson's disease. AB - OBJECTIVE: To assess the relationship between severity and progression of illness in Parkinson's disease and the use of healthcare resources. DESIGN AND SETTING: This was a prospective cost-of-illness study conducted in France based on clinical observation over a 6-month period of patients with Parkinson's disease treated in the hospital or community setting. Regression analyses were performed to construct the model that offered the best explanation for health expenditures using clinical and sociodemographic indicators. PATIENTS AND PARTICIPANTS: All patients included in the study had well-defined idiopathic Parkinson's disease, were aged > 35 years, were receiving treatment with levodopa or other antiparkinsonian agents, and were capable of completing questionnaires, alone or with the help of a household member. The final study population consisted of 294 patients, of whom 54 were enrolled by general practitioners and 240 by neurologists. INTERVENTIONS: Investigators completed a clinical questionnaire at the beginning and end of the 6-month observation period. Patients completed a questionnaire on their daily living conditions at the beginning and end of the study, and also completed monthly reports of healthcare use and loss of productivity. Patients with motor fluctuations also filled in fluctuation diaries on 4 consecutive days at the beginning and end of the 6-month period. Resource data collected included hospital stays, ancillary care, drug therapy, medical visits and transportation. Social costs were evaluated in nonmonetary terms, with the exception of costs of adapting the home environment. Transfer payments were analysed using reports from patients. MAIN OUTCOME MEASURES AND RESULTS: Hospital stays were the most expensive component of care (39% of costs), followed by ancillary care (30%) and drug therapy (22%). The mean medical cost was 308 euros (EUR) [$US357] for patients followed by a general practitioner and EUR2580 ($US2993) for patients followed by a neurologist. Costs also varied with age and motor fluctuations. Medical costs were strongly correlated with most clinical indicators and the cost generally progressed in line with the severity of the disease. The strongest correlation was between clinical indicators and ancillary care costs. CONCLUSIONS: These results confirm the importance of the social burden of Parkinson's disease. The regression results could be used to evaluate the benefit of novel treatments that reduce the intensity of motor fluctuations. PMID- 10539123 TI - Incremental cost-effectiveness analysis of intravenous ganciclovir versus oral ganciclovir in the maintenance treatment of newly diagnosed cytomegalovirus retinitis in patients with AIDS. AB - OBJECTIVE: To evaluate the cost effectiveness of a new product, oral ganciclovir, in comparison to a current therapy, intravenous (i.v.) ganciclovir, in the maintenance treatment of newly diagnosed cytomegalovirus (CMV) retinitis in patients with AIDS. DESIGN: This was a retrospective economic study of a prospective non-blinded randomised clinical trial. The model included i.v. ganciclovir induction, i.v. or oral ganciclovir maintenance and i.v. ganciclovir reinduction for patients whose CMV retinitis progressed. Safety and efficacy data were derived from the trial. A panel of Canadian infectious disease physicians and family physicians estimated the following in relation to i.v. ganciclovir treatment for CMV retinitis and related adverse events: healthcare resource utilisation, clinical practice patterns, patient out-of-pocket expenses and time loss from work. The incremental cost-effectiveness analysis is reported from a societal and a Ministry of Health perspective. SETTING: The trial was conducted in Canada (2 centres) and the US (13 centres) between March 1991 and November 1992. The model assumed that patients received either inpatient or outpatient care, or both. The model provided an analysis in a Canadian setting. PATIENTS AND PARTICIPANTS: Participants were patients with AIDS and newly diagnosed CMV retinitis. INTERVENTIONS: All patients received induction therapy with i.v. ganciclovir 5 mg/kg, twice daily for 14 days then once daily for 7 days. Patients whose CMV retinitis stabilised were randomised to maintenance therapy with either i.v. ganciclovir (5 mg/kg/day; n = 57) or oral ganciclovir (3000 mg/day; n = 60) and were followed for up to 140 days after the start of maintenance therapy. MAIN OUTCOME MEASURES AND RESULTS: The trial demonstrated that the mean time to progression of CMV retinitis was 57 days for oral ganciclovir compared with 62 days for i.v. ganciclovir maintenance therapy, as measured by masked fundus photography, and 96 days with i.v. ganciclovir compared with 68 days with oral ganciclovir according to the funduscopy results. There were more adverse events in the i.v. ganciclovir group compared with the oral ganciclovir group. The cost effectiveness results provide the dollar amount expended in order to continue to provide additional benefit using i.v. ganciclovir compared with oral ganciclovir. The incremental cost-effectiveness (C/E) ratio was 482 Canadian dollars ($Can: 1993 to 1995 values) per progression-free day gained with i.v. ganciclovir. Sensitivity analysis using funduscopy, rather than fundus photography, to document progression of CMV retinitis resulted in a C/E ratio of $Can42. CONCLUSIONS: This analysis found that i.v. ganciclovir provided additional days free of progression of CMV retinitis when compared with oral ganciclovir, but the costs were higher. PMID- 10539124 TI - Diclofenac/misoprostol. Pharmacoeconomic implications of therapy. AB - The combined formulation of diclofenac/misoprostol provides effective relief of pain and inflammation, with a 2- to 3-fold lower incidence of NSAID-associated gastroduodenal ulcers than diclofenac monotherapy. Both components of the combined formulation have been widely used and have well documented efficacy and tolerability profiles. Compared with other agents used as prophylaxis for NSAID induced gastropathies, misoprostol is generally considered to have the most extensive outcomes data establishing its efficacy in preventing both gastric and duodenal ulcers associated with long term NSAID use. Economic analyses conducted to date have shown that diclofenac/misoprostol is associated with similar or lower total direct medical treatment costs compared with other NSAIDs (with or without coprescribed misoprostol or an alternate prophylactic agent). As with pharmacoeconomic studies of coprescribed misoprostol with NSAIDs, the most favourable results with the combined formulation of diclofenac/misoprostol appear to be in patients at high risk of developing NSAID-associated gastroduodenal ulcers (e.g. the elderly). Although economic analyses with diclofenac/misoprostol were conducted in several different countries using a variety of methodologies and employing a wide range of clinical and economic assumptions, results have been generally favourable for the combined formulation. However, as is the case with pharmacoeconomic analyses in general, results of individual studies with diclofenac/misoprostol may not be generalisable between countries and are subject to change over time. Overall, clinical and economic data suggest that the optimal and most cost-effective use of the combined formulation of diclofenac/misoprostol is in patients requiring long term NSAID therapy who are at increased risk of developing NSAID-induced gastropathy, such as elderly patients with rheumatoid arthritis or osteoarthritis. PMID- 10539125 TI - The pharmacoeconomics of hormone replacement therapy. AB - Hormone replacement therapy (HRT) is a highly cost-effective treatment for symptoms of the menopause such as hot flushes (flashes). A number of economic evaluations have indicated that it may also be a cost-effective therapy for the prevention of cardiovascular disease and osteoporosis. However, these evaluations are based on the premise that HRT will reduce cardiovascular disease by 30 to 50%. Recent evidence casts doubt on its effectiveness at preventing cardiovascular disease, certainly as a secondary preventive therapy. Furthermore, HRT is likely to increase the incidence of breast cancer. If the effect of HRT on the cardiovascular system is slight or nonexistent, but its effect on breast cancer is modest or strong, then HRT is unlikely to be a cost-effective treatment for asymptomatic women at low risk of osteoporosis. However, the unwanted effects of HRT on the breast may be significantly reduced by targeting therapy to those women with low bone mass and who have other risk factors for fracture. Such a strategy is likely to be more cost effective than a strategy which allows asymptomatic women with low fracture risk to take HRT in the long term. As selective estrogen receptor modulators (SERMs) aggravate menopausal symptoms they are not likely to be an alternative for most perimenopausal women. Therefore, SERMs are more likely to be a competitor to existing and forthcoming bisphosphonates rather than HRT. PMID- 10539127 TI - With LOINC there's hope for mining lab data. PMID- 10539126 TI - Donepezil. Pharmacoeconomic implications of therapy. AB - Donepezil is a specific acetylcholinesterase inhibitor that can improve symptoms in patients with mild-to-moderate Alzheimer's disease; cognitive function is maintained above baseline levels for up to 1 year and normal decline of cognitive function is slowed. The ability of the patient to perform daily activities and neuropsychiatric symptoms may also be improved by donepezil, but data are limited. Donepezil is not expected to alter the underlying neurodegenerative process, and the response to the drug varies between individuals. In the absence of validated instruments to measure quality of life, it is not clear how donepezil affects this parameter. In a US survey of caregivers of patients with Alzheimer's disease who were being cared for at home at the start of the 6-month study period, treatment with donepezil did not increase overall direct medical costs. The acquisition cost of the drug was balanced by reduced institutionalisation costs. Economic analyses using Markov models from the US, UK and Canada suggest that donepezil initiated in the early stages of disease may be effectively cost neutral as a result of patients remaining in a nonsevere state of disease for a longer time. PMID- 10539128 TI - LMW heparins: from transition to fruition. PMID- 10539129 TI - What OR physicians want you to know about transfusion. PMID- 10539130 TI - Practical solutions: designing for improved human performance. PMID- 10539131 TI - Opening the gateway to change: creating a human-centered medical center- strategies for competing in the healthcare marketplace. PMID- 10539132 TI - Opening the gateway to change: the legal imperative for life-enhancing design. PMID- 10539133 TI - Opening the gateway to change: using the patient satisfaction survey to improve environmental quality. PMID- 10539134 TI - Long-term care design: life-enhancing design strategies at the Louis Feinstein Alzheimer Day Care Center. PMID- 10539135 TI - Long-term care design: what you need to know about designing life-enhancing environments. PMID- 10539136 TI - Long-term care design: National Alzheimer's Design Assistance Project--promoting innovation through exemplary settings. PMID- 10539137 TI - Children's health design: designing for family-centered care. PMID- 10539138 TI - The relationship between environmental design & patient medical outcomes. PMID- 10539139 TI - Children's health design: improving child health outcomes with Rehab 1, 2, 3. PMID- 10539140 TI - Design technology: what you need to know about circadian rhythms in healthcare design. PMID- 10539142 TI - Design technology: life-enhancing lighting design. PMID- 10539141 TI - Design technology: life-enhancing sound design. PMID- 10539143 TI - Clinical research data illuminating the relationship between the physical environment & patient medical outcomes. AB - The past 8 to 10 years have produced a large amount of information on the sensory development of the preterm infant. As a result of this information, it is evident that many of the environmental factors and care practices in the NICU do have a significant impact on infant sensory development. The factors that relate to light and noise clearly can be technologically modified to adapt better to infants' needs. Lights can be individually controlled. Focused lighting with limited scatter can be used, and a variety of techniques are available to produce barriers between light and the infant. Sound reduction and noise control in the NICUs are a constant problem. Much of the noise is generated by personnel. Much of it, however, is not essential and can be controlled. While the sound levels of the nursery do not produce any form of hearing loss, as measured by traditional hearing tests, it is now clear that they have the capacity to interfere with frequency discrimination and pattern recognition. These effects have been demonstrated in a wide range of animals and are currently under study in humans. The problems of sleep deprivation are very much tied to care practices and NICU organization. Again, the evidence supports the need for extended periods of undisturbed sleep in infants at all levels of maturity. This requires active involvement of the staff and the development of care plans and practices that allow infants clear rest periods undisturbed. PMID- 10539144 TI - Healthcare design competition. PMID- 10539145 TI - Healthcare environment awards. PMID- 10539146 TI - Nightingale product design competition. PMID- 10539147 TI - Exemplary healthcare facilities. PMID- 10539148 TI - Competing by design--what you need to know about tomorrow's business in healthcare. PMID- 10539149 TI - Through the patient's eyes. PMID- 10539150 TI - A theory of supportive design for healthcare facilities. PMID- 10539151 TI - Creating business success from user-driven design. PMID- 10539152 TI - Launching & sustaining the healthcare design revolution. PMID- 10539153 TI - Primary care design: case study of physician practices at Massachusetts General Hospital. PMID- 10539154 TI - Ambulatory care design: Brigham & Women's Hospital. PMID- 10539155 TI - Acute care design: Planetree addition to Griffin Hospital. PMID- 10539156 TI - Long-term care design: Hearthstone Alzheimer Care Center. PMID- 10539157 TI - Integrated care design: the Arizona Center for Health & Medicine--the integration of standard & alternative treatment practices under managed care. PMID- 10539158 TI - Life-enhancing design: a workshop for facility decision makers on the value of life-enhancing healthcare design. PMID- 10539159 TI - Current issues: designing the universal patient care room. PMID- 10539160 TI - Current issues: the secret lives of patients. PMID- 10539161 TI - Current issues: hospital design qualities to facilitate healing. PMID- 10539162 TI - A users' guide to healthcare design research. PMID- 10539163 TI - Practical solutions: high-performance healthcare textiles. PMID- 10539164 TI - Practical solutions: life-enhancing design--fast & on a tight budget. PMID- 10539166 TI - Investment in people. Making the case to top management. PMID- 10539165 TI - Helping seniors take their meds. PMID- 10539167 TI - Investment in people. Getting your money's worth from training. PMID- 10539168 TI - Coalitions keep quality alive. PMID- 10539169 TI - Assessing health plans from an IS perspective. PMID- 10539170 TI - The UR controversy. PMID- 10539171 TI - Kentucky takes on Aetna's doc policy. PMID- 10539173 TI - Do-it-yourself online records. PMID- 10539172 TI - Data watch. How employers are tackling Rx costs. PMID- 10539174 TI - A Phoenix rises from the ashes. Interview by MargaretAnn Cross. PMID- 10539175 TI - Private data on a public network. PMID- 10539176 TI - Inside health care's innovative Websites. PMID- 10539177 TI - A worldwide network of supplies. PMID- 10539178 TI - As Y2K work ends, CIOs set sights on electronic records. PMID- 10539179 TI - Obstacles and lessons on the road to a bug-free Y2K. PMID- 10539180 TI - HIMSS/RSNA collaboration achieves first goal. PMID- 10539181 TI - Associations name Wei-Tih Cheng CIO of the year. Interview by Tyler L. Chin. PMID- 10539182 TI - You've got mail? So do your patients. PMID- 10539183 TI - Diving into the Internet. PMID- 10539184 TI - Working the web. Massachusetts providers lobby for better online survey. PMID- 10539185 TI - OIG vey! Consolidated billing creates new liability risks. PMID- 10539186 TI - Tailor made: the time is right to customize your community. PMID- 10539187 TI - Rethinking rehab: make sure your rehab services suit the patients you serve. PMID- 10539188 TI - EEOC raises the bar. New guidance expands employer responsibility under the ADA. PMID- 10539189 TI - Pay check: Contemporary's 1999 nursing and administrative salary survey. PMID- 10539190 TI - Proceed with caution. Eight roadblocks to success in assisted living. PMID- 10539191 TI - Linking job and care quality. Interview by Yvonne Parsons. PMID- 10539192 TI - Perspectives. BBA just one of many forces shaping hospitals' financial fate. PMID- 10539193 TI - Marketplace. Festering dispute headed for court: do nonprofit HMOs have to shift risk? PMID- 10539194 TI - Perspectives. Senate cuts some slack to beleaguered HMO industry. PMID- 10539195 TI - Interview with James Rogers, president and CEO, Volunteers of America National Services. Interview by James A. Johnson. PMID- 10539196 TI - Influential leadership and change environment: the role leaders play in the growth and development of the people they lead. PMID- 10539197 TI - Rural managed care. PMID- 10539198 TI - Painful medicine: managed care and the fate of America's major teaching hospitals. AB - Healthcare spending in the United States has risen steadily throughout the post World War II period as the American healthcare system has been transformed from cottage industry to big business. The increasing rate of social investment in healthcare also transformed America's major teaching hospitals. As a case in point, the University of Iowa Hospitals and Clinics saw annual operating revenues rise from $1 million in 1945 to more than $350 million in 1995, which was accompanied by an extraordinary expansion in its physical facilities and in its multifaceted operations. In the 1970s and even more so in the 1980s, however, the unceasing climb in healthcare spending fueled concern among policy experts, politicians, employers, and insurers alike. In turn, the search for effective cost controls led to the current managed care revolution. While the end of that revolution is not yet in sight, managed care has, it appears, effected significant cost savings, but at no small cost to America's major teaching hospitals and their social missions of teaching, research, and patient care. Whether those missions can survive--and, if so, in what form--in a healthcare system dominated by the managed care ethos is an increasingly important concern. PMID- 10539199 TI - Communication patterns and group composition: implications for patient-centered care team effectiveness. AB - To assess how diversity affects team communication and to identify strategies to improve communication and patient care, focus groups of care production team members were held in two case study hospitals that have implemented the patient centered care model. Results indicate that care production team members generally support patient-centered care as a model that can work effectively in practice, even in an urban environment in which diversity concerns can affect team cohesiveness and communication. Successful implementation of the model, however, requires that hospitals consistently employ management strategies and reward structures that reinforce the value of teamwork and emphasize training and staff development. Key steps that healthcare executives can undertake to improve the performance of care production teams are detailed in this article and center around the following themes: team involvement in process improvement; a heightened emphasis on training (i.e., team and diversity training for all team members, task focused training for nonlicensed care givers; and leadership training for RNs); and the implementation of team-based reward and incentive structures. PMID- 10539200 TI - Mentoring in health administration: the critical link in executive development. AB - Career development in the next century will be predicated on the ability to forge hierarchical as well as lateral relationships within diverse organizational environments. Mentoring relationships may provide the critical link for future health executives to remain involved in their organization and their profession. This national study of 540 healthcare managers examined the prevalence of mentoring relationships according to demographic, organizational, and professional variables. Mentors appeared to influence access to promotional opportunities for managers in the healthcare industry and provide a mechanism for executive development. PMID- 10539202 TI - Economic credentialing: the propriety of managing physician costs through privileging. AB - Hospital executives face the unique task of managing the costs of an institution in which they have no direct managerial authority over the primary cost drivers, namely, the physicians who practice in the hospital. Perhaps the most controversial method of controlling physician costs consists of the application of economic factors to the credentialing process. Using the credentialing process as a technique to exert fiscal control over physicians affords hospital executives and their governing boards a tremendous cost-management opportunity. The legal propriety of economic credentialing remains unsettled. Many commentators, relying on limited case law, conclude that hospitals can engage in economic credentialing. Nevertheless, hospitals should exercise care when employing an economic rationale to restrict privileges lest they stir up legal challenges. Moreover, if hospitals use economic credentialing to limit medicaid patients' access to hospitals by excluding these patients' physicians from the hospital, the federal government may have the last word on the propriety of the practice. PMID- 10539201 TI - Cooperative ventures in a competitive environment: the influence of regulation on management decisions. AB - Two generalized game theory models are developed to explain observed management decisions between two large hospitals in Shelby County Tennessee within the regulatory context of competition for an advanced radiological technology pursued through the certificate-of-need process. The first model rationalizes each hospital's decision to submit competing certificate-of-need applications. The second model rationalizes the eventual joint venture agreement between the two hospitals and offers an explanation as to why the technology has yet to be acquired. The models are tested through interviews with the hospital administrators responsible for negotiating the joint venture agreement. The interviews confirm a preemptive motive behind each hospital's decision to compete and that the certificate-of-need requirement contributed to the eventual joint venture agreement. PMID- 10539204 TI - Hiring without firing. AB - Hiring executives has always been a daunting task--and today's economy makes it tougher than ever. The global scope and breakneck pace of business, the shrinking supply of job candidates, and the constant shift of organizational structures have increased the stakes exponentially; one wrong hire can quickly derail a company. Yet recent studies indicate that between 30% and 50% of executive-level hires end in firings or resignations. What makes hiring go wrong so often? And how can executives substantially improve the outcome of the process? This article provides some surprising answers to those questions. Fernandez-Araoz presents ten common hiring traps and many real-world examples of how those traps have scuttled business plans in a variety of industries worldwide. A large consumer goods company, for instance, slipped into the delegation gaffe trap when it handed over the screening and interviewing process to a mismatched team of managers that had an agenda different from the CEO's. And the ignoring emotional intelligence trap tripped up a U.S. telecommunications company that hired a CEO with a great track record--only to fire him less than a year later when his lack of cross-cultural social skills was discovered. Hiring well is a strategy-perhaps an organization's most important one, the author says. To sidestep the hiring traps, he suggests ways to systematically assess the company's needs and to determine how those needs mesh with the open job description--before candidates walk through the door. Fernandez-Araoz's search strategy incites managers with hiring responsibilities to be creative, determined, and courageous when embarking on a candidate search. PMID- 10539203 TI - Hospital customer service in a changing healthcare world: does it matter? AB - The healthcare industry is undergoing a rapid transformation to meet the ever increasing needs and demands of the patient population. Employers and health plans such as HMOs are demanding better service and higher quality care, and hospitals are trying to tackle reimbursement cutbacks, streamline services, and serve a diverse population. Hospitals have begun to realize that to overcome these obstacles and meet the needs of the health care plans and consumers, they must focus on the demands of the customer. Customer service initiatives increase patient satisfaction and loyalty and overall hospital quality, and many hospitals have found that consumer demands can be met through initiating and maintaining a customer service program. This article describes how the administrator can create, implement, and manage customer service initiatives within the hospital. PMID- 10539205 TI - Are you paying too much for that acquisition? AB - Despite 30 years of evidence demonstrating that most acquisitions don't create value for the acquiring company, executives continue to make more deals, and bigger deals, every year. There are plenty of reasons why value isn't created, but many times it's simply because the acquiring company paid too much. It's not, however, that acquirers pay too high a price in an absolute sense. Rather, they pay more than the acquisition is worth to them. What is that optimum price? The authors present a systematic way to arrive at it, involving several distinct concepts of value. In today's market, the purchase price of an acquisition will nearly always be higher than the intrinsic value of the company--the price of its stock before any acquisition intentions are announced. The key is to determine how much of that difference is "synergy value"--the value that will result from improvements made when the companies are combined. This value will accrue to the acquirer's shareholders rather than to the target's shareholders. The more synergy value a particular acquisition can generate, the higher the maximum price an acquirer is justified in paying. Just as important as correctly calculating the synergy value is having the discipline to walk away from a deal when the numbers don't add up. If returns to shareholders from acquisitions are no better in the next ten years than they've been in the past 30, the authors warn, it will be because companies have failed to create systematic corporate governance processes that put their simple lessons into practice. PMID- 10539206 TI - Bringing the environment down to earth. AB - The debate on business and the environment has typically been framed in simple yes-or-no terms: "Does it pay to be green?" But the environment, like other business issues, requires a more complex approach--one that demands more than such all-or-nothing thinking. Managers need to ask instead, "Under what circumstances do particular kinds of environmental investments deliver returns to shareholders?" This article presents five approaches that managers can take to identify those circumstances and integrate the environment into their business thinking. These approaches will enable companies with the right industry structure, competitive position, and managerial skills to reconcile their responsibility to shareholders with the pressure to be faithful stewards of the earth's resources. Some companies can distance themselves from competitors by differentiating their products and commanding higher prices for them. Others may be able to "manage" their competitors by imposing a set of private regulations or by helping to shape the rules written by government officials. Still others may be able to cut costs and help the environment simultaneously. Almost all can learn to improve their management of risk and thus reduce the outlays associated with accidents, lawsuits, and boycotts. And some companies may even be able to make systemic changes that will redefine competition in their markets. All five approaches can help managers bring the environment down to earth. And that means bringing the environment back into the fold of business problems and determining when it really pays to be green. PMID- 10539207 TI - The case of the religious network group. AB - GenCorp, a Connecticut-based paper-goods manufacturer, has long supported employee-organized network groups. Its social support group for African Americans, in fact, has been a particular success, having provided black employees with opportunities to further enhance their careers and helped the company retain top talent, meet its EEO goals, and gain favorable publicity. So when Alice Lawrence, a top accountant at GenCorp, called general manager Bill Thompson about the Christian network group being organized in one of the company's southern plants, Bill hardly flinched. After all, the Christian group was being organized by Russell Kramer, one of the company's most effective plant managers. What could be the problem there? But a couple of years ago, Alice noted, Russell had sent around a companywide letter that talked about the sinful nature of homosexuality. And that letter has made her and other gay and lesbian employees terribly uneasy. To complicate matters, the issue of "Christian rights" in the workplace was being widely discussed on radio talk shows, and several books on the topic had recently been published. An employee had even called the new region's head of human resources to get clarification on the topic. Up until now, GenCorp hadn't placed a lot of restrictions on network groups. But the emergence of a religious group was raising new questions for GenCorp's managers. Should the company accept religious groups or try to stop them? What policy, if any, should GenCorp adopt toward these network groups? Five experts comment on this fictional case study. PMID- 10539208 TI - Why good companies go bad. AB - One of the most common business phenomena is also one of the most perplexing: when successful companies face big changes, they often fail to respond effectively. Many assume that the problem is paralysis, but the real problem, according to Donald Sull, is active inertia--an organization's tendency to persist in established patterns of behavior. Most leading businesses owe their prosperity to a fresh competitive formula--a distinctive combination of strategies, relationships, processes, and values that sets them apart from the crowd. But when changes occur in a company's markets, the formula that brought success instead brings failure. Stuck in the modes of thinking and working that have been successful in the past, market leaders simply accelerate all their tried-and-true activities. In attempting to dig themselves out of a hole, they just deepen it. In particular, four things happen: strategic frames become blinders; processes harden into routines; relationships become shackles; and values turn into dogmas. To illustrate his point, the author draws on examples of pairs of industry leaders, like Goodyear and Firestone, whose fates diverged when they were forced to respond to dramatic changes in the tire industry. In addition to diagnosing the problem, Sull offers practical advice for avoiding active inertia. Rather than rushing to ask, "What should we do?" managers should pause to ask, "What hinders us?" That question focuses attention on the proper things: the strategic frames, processes, relationships, and values that can subvert action by channeling it in the wrong direction. PMID- 10539209 TI - Collaborating with congregations: opportunities for financial services in the inner city. AB - In all economies, financial systems perform a basic set of functions, which include the need to pool resources, to save and borrow, to make payments, and to collect information. And yet, in rich and poor communities, the ways in which those needs are met differ greatly. In part, this is because traditional financial service firms have found it too expensive to serve poor neighborhoods. But it is possible to work with less traditional institutions to meet those needs. According to the authors, inner cities face two core impediments to financial services: lack of economies of scale and lack of good information. An inner-city investor with $500, for instance, does not warrant much attention from financial service firms. But 20,000 parishioners investing $500 apiece can collectively wield $10 million. By partnering with strong social organizations such as churches, financial institutions can benefit both themselves and investors. A lack of good information concerning credit histories--a common problem in poor communities--also makes low-income customers much less appealing to financial institutions. Churches can help close such information gaps by vouching for parishioners' reliability. There are certainly problems that come with mixing business and religion, as the authors concede. Ethical issues, trust issues, and issues of experience all come to mind. But understanding that functions need to dictate the structure of the financial service sector may be the first step toward achieving inner-city prosperity. PMID- 10539210 TI - Turning goals into results: the power of catalytic mechanisms. AB - Most executives have a big, hairy, audacious goal. They write vision statements, formalize procedures, and develop complicated incentive programs--all in pursuit of that goal. In other words, with the best of intentions, they install layers of stultifying bureaucracy. But it doesn't have to be that way. In this article, Jim Collins introduces the catalytic mechanism, a simple yet powerful managerial tool that helps translate lofty aspirations into concrete reality. Catalytic mechanisms are the crucial link between objectives and performance; they are a galvanizing, nonbureaucratic means to turn one into the other. What's the difference between catalytic mechanisms and most traditional managerial controls? Catalytic mechanisms share five characteristics. First, they produce desired results in unpredictable ways. Second, they distribute power for the benefit of the overall system, often to the discomfort of those who traditionally hold power. Third, catalytic mechanisms have teeth. Fourth, they eject "viruses"- those people who don't share the company's core values. Finally, they produce an ongoing effect. Catalytic mechanisms are just as effective for reaching individual goals as they are for corporate ones. To illustrate how catalytic mechanisms work, the author draws on examples of individuals and organizations that have relied on such mechanisms to achieve their goals. The same catalytic mechanism that works in one organization, however, will not necessarily work in another. Catalytic mechanisms must be tailored to specific goals and situations. To help readers get started, the author offers some general principles that support the process of building catalytic mechanisms effectively. PMID- 10539211 TI - The toxic handler: organizational hero--and casualty. AB - You've watched them comfort colleagues, defuse tense situations, and take the heat from tough bosses. You've seen them step in to ease the pain during layoffs and change programs. Who are they? The authors call them toxic handlers--managers who voluntarily shoulder the sandness, frustration, bitterness, and anger of others so that high-quality work continues to get done. Toxic handlers are not new. They are probably as old as organizations themselves. But there has never been a systematic study of the role they play in business. In this article, the authors introduce the role of toxic handlers, explaining what they do and why. Managing the pain of others is hard work. Toxic handlers save organizations from self-destructing, but they often pay a high price--emotionally, professionally, and sometimes physically. Some toxic handlers experience burnout; others suffer far worse consequences, such as ulcers and heart attacks. The authors contend that these unsung corporate heroes have strategic importance in today's business environment. Effective pain management can--and does--contribute to the bottom line. No company can afford to let talented employees burn out. Nor can it afford to have a reputation as an unhappy place to work. The authors offer practical advice for managers and organizations about how to support toxic handlers--before a crisis strikes. The role of toxic handler needs to be given the attention it deserves for everyone's benefit, because the health of employees is a key element in the long-term competitiveness of companies and of society. PMID- 10539212 TI - Integrated health care delivery system conducts ad agency search as part of its brand-launching effort. AB - PennState Geisinger Health System, Hershey, Pa., conducted an extensive ad agency search after its inception in 1997. The integrated health care delivery system needed to introduce its brand to an audience that was confused by the wide array of available health care options. BVK/McDonald, Milwaukee, the agency selected, has created a branding campaign that revolves around the tag-line "The power of health." PennState Geisinger will tabulate the results of BVK/McDonald's multi million dollar campaign in 2000; at that time it will know whether its selection committee chose wisely. PMID- 10539213 TI - Health Midwest's pragmatic marketing avoids glitz in cluttered ad environment. PMID- 10539214 TI - Missouri Health Science Center finds the Web an effective way to communicate with employees. PMID- 10539216 TI - Provider sponsored organization hits the ground running with extensive marketing campaign. St. Joseph Medicare Plus, Albuquerque, NM. PMID- 10539215 TI - Newsletter key to overwhelming success of Seattle pediatric hospital's 1999 telethon.Children's Hospital & Regional Medical Center, Seattle. PMID- 10539217 TI - Baystate Health System launches successful cosmetic surgery campaign. PMID- 10539218 TI - Hate crime brings global attention to a northern Colorado hospital. Poudre Valley Health System, Fort Collins, CO. PMID- 10539219 TI - Radio talk can be valuable public relations tool. PMID- 10539220 TI - Direct mail magazine anchors hospital's marketing effort for its community wellness programs. Lehigh Valley Hospital and Health Network, Allentown, Pa. PMID- 10539221 TI - Computerized material management, inventory control, and purchase orders in the clinical laboratory. AB - The management of supplies, purchase orders, and equipment is of critical importance to the operation of the clinical laboratory. The InvMan software program has reduced hands-on time for performing the counting of inventory and the time required to generate purchase orders. These changes save the laboratory about $11,170 per year in personnel costs. While the use of a structured system does impose some constraints on the user, the program has helped the laboratory organize all of its supply, vendor, location, PO, and equipment data. It has allowed the laboratory to respond more rapidly and accurately to inquiries related to inventory. The varied functions of InvMan provide flexibility to the laboratory and permit it to define the inventory as best fits a particular situation. The application is well suited to its target audience. The program has performed well, allowing the laboratory to make significant improvements to its material management system. PMID- 10539223 TI - Psychological and perceived situational predictors of physical activity: a cross sectional analysis. AB - The purpose of this study was to identify psychological and self-reported situational factors that are associated with degree of involvement in moderate intensity physical activity at various stages of adult life. The study is grounded in Personal Investment Theory which proposes that personal incentives, sense of self and perceived options determine behavior. Participants aged 18 and above, selected by random-digit dialling, were invited to participate in a study on physical activity habits. Of 251 who agreed to participate, 41.4% were male (N = 104) and 58.6% were female (N = 147). These participants were asked the number of days per week that they engaged in physical activity which accumulated a total of 30 min or more. The 140 participants who indicated one or more days of activity answered questions concerning personal incentives for physical activity, sense of self and perceived barriers. Stepwise multiple regression analyses and discriminant function analysis indicated that Personal Investment Theory is able to predict up to 29% of the variance associated with voluntary participation in moderate-intensity physical activity. Discussion focuses on implications for physical activity programs for citizens at different stages of their adult life. PMID- 10539222 TI - Health technology assessment. PMID- 10539224 TI - Lay beliefs about the preventability of major health conditions. AB - Beliefs about the extent to which health problems can be prevented reflect an understanding that preventive measures can reduce adverse health events and the level of control individuals perceive that they hold over the factors that affect their health. A population survey of 1659 people conducted in 1995 in south western Sydney, Australia, found that only child drownings, tooth decay, skin cancer, and burns and scalds were considered all or mostly preventable by more than 50% of the sample. The majority of respondents did not believe that heart attacks, cervical cancer, high blood pressure, serious road injury, lung cancer and asthma deaths were all or mostly preventable. Logistic regression analysis showed that people born in an English speaking country, those with more than 10 years of education and men were significantly more likely to recognize a number of key conditions as highly preventable. The findings suggest that, in spite of the range of prevention efforts in Australia to date, these are not matched by strong beliefs within the community that prevention is possible. Communication of the opportunities and methods for prevention needs to be improved, particularly among certain population groups. The findings also indicate a need to examine social and environmental factors which are potentially reducing confidence, and subsequently and adoption of preventive behaviours. PMID- 10539225 TI - Understanding the intention to permanently follow a high folate diet among a sample of low-income pregnant women according to the Health Belief Model. AB - Despite folate fortification of the US food supply beginning January 1, 1998, evidence indicates that a substantial proportion of women of child-bearing age will continue to have folate intakes inadequate for the prevention of neural tube defects (NTDs). Therefore, health education remains an essential component of this public health campaign. The purpose of this study was to determine the applicability of the Health Belief Model (HBM) to understanding the intention to permanently follow a high folate diet among low-income pregnant women. A convenience sample of 251 low-income pregnant women participated in individual 15 min interviews assessing their folate attitudes and beliefs according to the model. Correlations consistent with the HBM were found between the perceived susceptibility, perceived severity, perceived benefits, perceived barriers, self efficacy and cues to action constructs, and participants' intention to permanently follow a high folate diet (folate intention). In regression analyses, the perceived benefits construct was consistently the most predictive of folate intention. Participants were generally unfamiliar with and had many misperceptions concerning both folate and NTDs. The HBM may offer an effective foundation for development of tailored educational interventions promoting permanent consumption of a high folate diet among low-income women. PMID- 10539226 TI - Health-related lifestyle in adolescence--origin of social class differences in health? AB - Survey data collected by mail, representing Finnish 16 year olds (N = 2977; response rate 83%), were used to identify which particular aspects of lifestyle are typical of adolescents who select various educational tracks and, thus, have different probabilities of ending up in low or high social positions. The dependent variable, educational track, was formed by classifying the respondents into five successive categories predicting their social position in adulthood. Lifestyle is measured by health behaviours, leisure-time activities and social relations. The probability of belonging to educational tracks with good social prospects in adulthood was high among adolescents who placed much emphasis on health-enhancing behaviours (not smoking, physical exercise, low milk-fat diet, dental hygiene, use of seatbelts, etc.), who did not spend much time watching TV or listening to music and who attended church or other religious meetings weekly. Health-related lifestyle, at the age of 16, is oriented towards the social group the individual is likely to belong to as an adult. The study provides evidence for a strong association between health-related lifestyle and educational track in adolescence. PMID- 10539227 TI - Changes in HIV/AIDS education, knowledge and attitudes among Scottish 15-16 year olds, 1990-1994: findings from the WHO: Health Behaviour in School-aged Children Study (HBSC). AB - There is concern about the high prevalence of adolescent sexual health problems, such as sexually transmitted diseases (STDs) and unwanted pregnancies, that currently exist in the UK. If young people are to reduce their risk from HIV/AIDS and other STDs it is imperative, in the first instance, they know what the risks are and how they can avoid them. However, effective school-based sex education can only be delivered if there are accurate data on young people's current levels of knowledge and existing sex education needs. This paper details findings from the WHO: Health Behaviours of School-aged Children Study on the changes that have occurred between 1990 and 1994 in Scottish school-children's knowledge, attitudes and perceived educational needs in relation to HIV/AIDS. There have been significant changes in knowledge and attitudes that may affect their sexual behaviour, e.g. in their attitudes to condom use, risk of HIV/AIDS and other STDs, and also other sexual health problems, such as the risk of unwanted pregnancies and abortions. Finally, areas that require future research and recommendations for future sexual health education interventions are highlighted. PMID- 10539228 TI - Measuring participatory strategies: instrument development for worksite populations. AB - A participatory strategies approach which involves employees in the planning and delivery of worksite health promotion programs was utilized in the 55 experimental worksites included in the national, NCI-funded Working Well Trial. According to study protocol, Employee Advisory Boards (EABs) were organized in each experimental worksite. This paper describes two substudies designed to develop and measure participatory strategies associated with the EABs in the Working Well Trial. Study 1 determined characteristics of the EABs, developed subscales and assessed the internal consistency of the scales. Study 2 used a confirmatory factor analysis to examine the structure of the developed questionnaire. The four subscales include: Autonomy/Independence, Management Involvement, Institutionalization/Commitment and Others Involvement. Results from Study 1 indicate that the four subscales of the 24-item instrument demonstrated strong internal consistency and three were sensitive enough to register differences by Study Center at the baseline. Study 2 results found that the EAB subscales again demonstrated good internal consistency, structural stability and acceptable sensitivity. An initial validity analysis was performed and yielded results which supported some but not all of the hypothesized associations. Implications for further refinement and application of this new instrument in worksite settings are explored. PMID- 10539229 TI - A critical appraisal of the draw and write technique. AB - The draw and write technique is increasingly popular in health education research with children. It is generally employed in the setting of the school classroom and is promoted as a 'bottom-up' approach which enhances participation by children. In this paper we critically appraise the use of this method. Against the background of a consideration of carrying out qualitative health promotion research with children we examine the origins and use of children's drawings in a number of disciplines and practice environments. We argue that, although the draw and write technique has made an important contribution to health education research, it fails to reflect the processes involved in the construction and collection of such data. A range of methodological, analytical and ethical issues are raised. We conclude that health education research with children must involve taking children seriously as social actors and query the assumption that drawing enables children to communicate their thought any more than does conversational language. We suggest that the development of research should be premised upon an appreciation of the social context and the world of the child. PMID- 10539230 TI - A brief motivational intervention to improve dietary adherence in adolescents. The Dietary Intervention Study in Children (DISC) Research Group. AB - Motivational interviewing offers health care professionals a potentially effective strategy for increasing a patient's readiness to change health behaviors. Recently, elements of motivational interviewing and the stages of change model have been simplified and adapted for use with patients in brief clinical encounters. This paper describes in detail a brief motivational intervention model to improve and renew dietary adherence with adolescents in the Dietary Intervention Study in Children (DISC). DISC is a randomized, multi-center clinical trial assessing the efficacy and safety of lowering dietary fat to decrease low-density lipoprotein cholesterol in high-risk children. In the first 3 years of follow-up covering ages 8-13, intervention participants (n = 334) were exposed to a family-based group intervention approach to change dietary choices. To address adherence and retention obstacles as participants moved into adolescence (age 13-17), an individual-level motivational intervention was implemented. The DISC motivational intervention integrates several intervention models: stages of change, motivational interviewing, brief negotiation and behavioral self-management. A preliminary test of the intervention model suggests that it was acceptable to the participants, popular with interventionists and appeared to be an age-appropriate shift from a family-based intervention model. PMID- 10539231 TI - Increased sexual abstinence among in-school adolescents as a result of school health education in Soroti district, Uganda. AB - A school health education programme in primary schools aimed at AIDS prevention in Soroti district of Uganda emphasized improved access to information, improved peer interaction and improved quality of performance of the existing school health education system. A cross-sectional sample of students, average age 14 years, in their final year of primary school was surveyed before and after 2 years of interventions. The percentage of students who stated they had been sexually active fell from 42.9% (123 of 287) to 11.1% (31 of 280) in the intervention group, while no significant change was recorded in a control group. The changes remained significant when segregated by gender or rural and urban location. Students in the intervention group tended to speak to peers and teachers more often about sexual matters. Increases in reasons given by students for abstaining from sex over the study period occurred in those reasons associated with a rational decision-making model rather than a punishment model. A primary school health education programme which emphasizes social interaction methods can be effective in increasing sexual abstinence among school-going adolescents in Uganda. The programme does not have to be expensive and can be implemented with staff present in most districts in the region. PMID- 10539232 TI - Padres Trabajando por la Paz: a randomized trial of a parent education intervention to prevent violence among middle school children. AB - This paper reports the results of a randomized trial to test the effectiveness of a theoretically derived intervention designed to increase parental monitoring among Hispanic parents of middle school students. Role model story newsletters developed through the process of Intervention Mapping were mailed to half of a subsample of parents whose children participated in Students for Peace, a comprehensive violence prevention program. The results indicated that parents in the experimental condition (N = 38) who had lower social norms for monitoring at baseline reported higher norms after the intervention than the parents in the control condition (N = 39) (P = 0.009). Children of parents in the experimental group reported slightly higher levels of monitoring at follow-up across baseline values, whereas control children who reported moderate to high levels of monitoring at pre-test reported lower levels at follow-up (P = 0.04). These newsletters are a population-based strategy for intervention with parents that show some promise for comprehensive school-based interventions for youth. PMID- 10539233 TI - Older adults in health education research: some recommendations. AB - A review of articles published in two health education journals is provided to examine the extent to which older adults were included in published research. The review suggests that older adults were included in about 15% of the research articles published in Health Education and Behavior and Health Education Research. Of the articles that include older adults, age differences in study processes and outcomes are rarely examined, and very few studies advance specific hypotheses based on a theoretical or conceptual model of aging or older adulthood. Several recommendations for health education research are suggested. PMID- 10539234 TI - A methodological inquiry into the evaluation of smoking cessation programmes. AB - This paper examines a methodological controversy and aims to show the advantages of introducing an alternative methodological approach, i.e. the 'scientific realist approach', into evaluation studies on health education programmes for smoking cessation. The methodological difficulties of the existing standard evaluation model, i.e. the quasi-experimental approach, are examined. This model fails to investigate how the programme setting influences outcomes (context problem) and draws attention away from understanding why programmes work because it adopts a 'successionist' logic (causation problem). An alternative methodology, the scientific realist approach, is proposed in order to cope with such problems. This approach adopts the 'generative' logic which looks at the matter of causation internally. This logic posits that the working of underlying mechanisms within a more basic level of reality causes relationships between visible events. According to the scientific realist approach, the actual outcomes of smoking cessation programmes follow from the workings of potential mechanisms whose functioning is afforded by contexts conducive to their operation. PMID- 10539235 TI - Becoming a physician executive: where to look before making the leap. PMID- 10539236 TI - The anatomy of a healing garden. PMID- 10539237 TI - Status report--an investigation to determine whether the built environment affects patients' medical outcomes. PMID- 10539238 TI - Healthcare Design Competition 1997. PMID- 10539239 TI - Healthcare Environment Awards 1997. PMID- 10539241 TI - Exemplary healthcare facilities. PMID- 10539240 TI - Nightingale Product Design Competition 1997. PMID- 10539242 TI - Loving leadership. PMID- 10539243 TI - The CQI imperative for supportive healthcare design. PMID- 10539245 TI - Creating organizations that inspire the soul. PMID- 10539244 TI - Sound health--designing a therapeutic auditory environment. PMID- 10539246 TI - Jerusalem house--a caring environment for homeless women living with AIDS & their children. PMID- 10539247 TI - The environmental risks of a construction project. PMID- 10539248 TI - Surviving & thriving in turbulent times--design as an essential and strategic investment. PMID- 10539249 TI - Using 3D animation for project success. PMID- 10539250 TI - Strategies for designing better Alzheimer's care environments. PMID- 10539251 TI - The Healthcare Design Research Alliance. PMID- 10539252 TI - Building homeness into existing long-term care facilities. PMID- 10539253 TI - Cultural expectations & locale--creating the eldergarden. PMID- 10539254 TI - The HealthEast 'caring touch' design project. PMID- 10539255 TI - Consumer perceptions of the healthcare environment--an investigation to determine what matters. PMID- 10539256 TI - Discovering the labyrinth as a tool for health & healing. PMID- 10539257 TI - The anatomy of a healthcare design research project. PMID- 10539258 TI - Seven communications habits that bring health to organizations & their staff. PMID- 10539259 TI - Connecting through words. PMID- 10539260 TI - Future development of healthcare institutions. PMID- 10539261 TI - Successful strategies for marketing healthcare design services. PMID- 10539262 TI - Michigan hospitals--building healthy communities. AB - The MHA Community Benefits Measurement Project is an ongoing, annual collection effort undertaken by the MHA and its hospital and health system members. This project quantifies the contributions made by nonprofit hospitals to their communities; from improving access to care for the underserved to bringing care right into the community through community-based programs. Over and above quantifying what is provided to the community in benefits, this survey helps to illustrate how Michigan hospitals are working to improve the overall health and welfare of the communities they serve. PMID- 10539263 TI - The customer's always right ... even when their attitude isn't. PMID- 10539264 TI - Integration: what's next in the new millennium? PMID- 10539265 TI - How to change the world. PMID- 10539267 TI - Employee handbooks: thin is not in. PMID- 10539266 TI - Our mission is the same yesterday, today and tomorrow. PMID- 10539269 TI - The system with the most physicians wins. PMID- 10539268 TI - Partnering for the community's well-being. AB - Health care organizations throughout Michigan continue to adjust to tightening budgets. Even so, there is a growing need to understand and act upon the more global and vexing health issues facing local communities. Managed care has greatly impacted community hospitals as well as community health agencies. The need to do more with less, and to do it more effectively whenever possible, has become a challenge for all involved in the health delivery system. PMID- 10539270 TI - Domestic violence is our business. PMID- 10539271 TI - The coming age. What's ahead for hospitals and health systems. AB - The health care delivery system is a reactive instrument: it conforms to the demands of the most powerful drivers. How the system responds to these drivers in terms of design of coverage and delivery of care represents in large part the major challenge of the coming decade. This article presents an overview of some of the major drivers of health care today--cost, customers and technology--and how hospitals and health systems are responding. PMID- 10539272 TI - Where does a community benefits program fit? PMID- 10539273 TI - Member satisfaction: a patient's perspective. PMID- 10539275 TI - Getting an edge on tomorrow. PMID- 10539274 TI - The 1999 Ludwig Award winners. Impacting Michigan's health--one community at a time. PMID- 10539277 TI - Integrative health care and demand management. PMID- 10539276 TI - The threshold of genetic research. PMID- 10539278 TI - Contraindications of nonprofit hospitals. PMID- 10539280 TI - Clearing the smoke from Medicare and Medicaid. PMID- 10539279 TI - Bringing people together for improvement. AB - The MHA North Central Hospital Council's first involvement in community health issues began in 1995 and focused initially on conducting a comprehensive assessment of health status in the North Central Council (NCC) region. More recently, the council's board identified a desire to collaborate across northern Michigan in making measurable improvement in youth physical activity. What follows is a description of the process that we have followed in our efforts to move beyond mere assessment to bringing people together for programs directed at improving health. PMID- 10539281 TI - Where is the money for education and research? PMID- 10539282 TI - How academic medicine can manage for the future. PMID- 10539283 TI - Health care becomes big business. PMID- 10539284 TI - Turning up the heat on managed care. PMID- 10539285 TI - The case of the vanishing inpatient. PMID- 10539286 TI - Is unity producing strength? PMID- 10539287 TI - Another reason to remember Pearl Harbor. PMID- 10539288 TI - Which care is worth the money? PMID- 10539290 TI - Quality health care--let's get out of the data dark ages. PMID- 10539289 TI - The rise of the health care consumer. PMID- 10539291 TI - Will wellness ever really catch on? PMID- 10539292 TI - Tough times ahead. PMID- 10539293 TI - Follow the money. PMID- 10539294 TI - Health care and community obligations. PMID- 10539295 TI - Threshold barriers to Title I and Title III of the Americans with Disabilities Act: discrimination against mental illness in long-term disability benefits. PMID- 10539296 TI - The role of new federalism and public health law. PMID- 10539297 TI - How reliable is medical malpractice law? A review of "Medical Malpractice and the American Jury: confronting the myths about jury incompetence, deep pockets, and outrageous damage awards" by Neil Vidmar. PMID- 10539298 TI - A proposal to recognize a legal obligation on physicians to provide adequate medication to alleviate pain. PMID- 10539300 TI - Not a run-of-the-mill profiling project. Initiative seeks to drive competition, shape industry. PMID- 10539299 TI - Battered women syndrome as a tort cause of action. PMID- 10539301 TI - Competition is increasing for international patients. PMID- 10539302 TI - Streamlined preadmission process slashes time for patient preparation. PMID- 10539303 TI - Are premium increases only a short-term antidote? PMID- 10539304 TI - Developmentally supportive care coming soon to your NICU. PMID- 10539305 TI - Legally speaking. PMID- 10539306 TI - Understanding research studies. PMID- 10539307 TI - AVs on a budget or "How to teach on the cheap". PMID- 10539308 TI - The Silver Book. Update to 1999 Buyer's Guide, new companies, EMS statistics. PMID- 10539309 TI - Creating an effective patient history. PMID- 10539310 TI - The road warriors of fund raising. PMID- 10539311 TI - iapps brings a new generation of Web sites to the non-profit sector. Organizations capitalize on Web sites built with Orgitecture. PMID- 10539312 TI - Toward 2000 and beyond: charitable and social change giving in the new millennium. Part 2. PMID- 10539314 TI - Your donors want to hear more about you...! PMID- 10539313 TI - Plan it right: create your architectural program and effectively use it. PMID- 10539315 TI - On accountability. PMID- 10539316 TI - Does your fund-raising program need a tune-up? PMID- 10539317 TI - Hell no, we won't go! PMID- 10539318 TI - Drug test. Online drugstores were ready to rock. There was just one little problem. PMID- 10539319 TI - When boilers go bad ... exploring safety. PMID- 10539320 TI - Chill factor: chiller safety is still a hot issue. PMID- 10539321 TI - The fright stuff: take the scare out of EtO. Learn to handle it safely. PMID- 10539322 TI - Energy buzz. ASHE Web site measures energy use. PMID- 10539323 TI - It cost $5,000 to get into this group--and a fortune to get out. PMID- 10539324 TI - Don't be suckered by drug seekers. PMID- 10539325 TI - Save time and please patients with e-mail. PMID- 10539326 TI - Will you be liable for Y2K computer glitches? PMID- 10539327 TI - Primary care feels the job squeeze. PMID- 10539328 TI - Patient education: helping the media get the facts straight. PMID- 10539329 TI - Medicare miscalculates, and doctors pay. PMID- 10539330 TI - Even small groups may need a fill-in doctor. PMID- 10539331 TI - A practice sold, a practice ruined. PMID- 10539332 TI - Undoing the damage: how to reclaim the practice you sold. PMID- 10539333 TI - Disease management to grow, but limited. PMID- 10539334 TI - Physicians' use of electronic medical records--identifying and crossing the barriers. PMID- 10539335 TI - Complementary and alternative medicine--a business opportunity? AB - This desire for health and well-being is driving the rapid growth of the complementary and alternative medicine (CAM) industry and points to a new role for health care professionals, including business opportunities for medical groups. CAM represents the opportunity to grow practice revenues, expand a group's tool kit for assisting patients with health care issues, and retain or increase market share by proactively responding to consumers. With respect to CAM, physician practices can lead their market, follow it or ignore it. PMID- 10539336 TI - Strategic planning: the basics and benefits. AB - Strategic planning can help a medical practice take an honest look at itself in light of the changes taking place in its environment and within the practice itself. The objective is for the group to design a plan, or road map, to its envisioned future. For medical practices, strategic planning is a four part process: 1) gaining buy-in for the process itself from the leadership and physicians; 2) gathering pertinent data about the group's environment through external resources, and about the group itself through interviews and surveys of physicians; 3) conducting a facilitated off-site retreat of key physicians and leaders in order to review data, discuss issues and develop a one to two year action plan; and, 4) carrying out the action plan developed at the retreat and measuring its outcomes. A follow-up mini-retreat about six months after the first retreat is highly recommended, as is instituting a process of sharing of the results and outcomes of the plan with all members of the organization. PMID- 10539337 TI - For patients and providers, it's all about access. AB - Increasingly, immediate patient service--whether scheduling an appointment or seeing a physician--is a realistic expectation. Smoothing patient access and hitting service targets takes sophisticated planning and a true scientific approach. The department of medicine at Boston's Children's Hospital has taken a new approach to visit management, using a model that could be applied to any practice in any setting. Staff have set up categories of care and guidelines for scheduling, realigning tasks in order to make better use of provider resources and expertise and restructuring associated supporting operations. PMID- 10539338 TI - How physician/administrator teams work in small groups. Six steps to make it happen. AB - The physician/administrator team is frequently supported as the preferred model for physician group governance. Perhaps an obvious model for large groups, it remains true that the largest percentage of physicians are practicing in groups of 10 or fewer. This article explores the applicability of the physician/administrator team concept for small group practices. The article covers the significance of the physician/administrator team in managed care settings, difference in governance structures between large and small groups, the need for physicians to be willing to share leadership in organizations they own, understanding empowerment in small groups, the manager's need to assume more responsibility and how to form the team. PMID- 10539339 TI - Is a PSO right for you? AB - The Balanced Budget Act of 1997 established the new Medicare+Choice program which provides a variety of alternatives to traditional Medicare Part A and Part B, including the provider sponsored organization (PSO). Over the next several years, a significant number of organizations will consider becoming a PSO. The decision requires a thorough and detailed review of critical success factors. This articles outlines those factors and defines some components of a successful PSO. PMID- 10539340 TI - What should be the primary focus of a group practice's marketing program? PMID- 10539341 TI - Paracelsus grooms for possible suitor. PMID- 10539343 TI - So what is a monopoly? Appeals court decision in Missouri case deals yet another blow to antitrust regulators. PMID- 10539342 TI - Report: JCAHO survey process fails public. PMID- 10539344 TI - Catholic system, Texas hospital partner. PMID- 10539345 TI - CHS kills deal to buy Calif. hospital. PMID- 10539346 TI - Judge OKs MedPartners reorganization. PMID- 10539347 TI - Panel seeks wider sharing or organs. PMID- 10539348 TI - Refocusing on fraud. Congress, HHS take another look at surety bonds as protection against home-care abuses. PMID- 10539349 TI - HCFA tones down spending projections. PMID- 10539350 TI - Monstrous problems. Medicare's not the only bogeyman haunting the long-term-care industry. AB - The problems in the long-term-care industry are positively monstrous. While many executives are pointing and screaming at Medicare cuts, lurking in the shadows are some self-imposed problems, such as the scary pile of debt many publicly traded companies are sitting on. PMID- 10539351 TI - Stent prices starting to retreat. PMID- 10539352 TI - Confessions of a Baptist Hospital exec. Financial manager tells HFMA attendees about mistakes that nearly caused system collapse. PMID- 10539353 TI - Tax-cut measure plays Medicare pawn. PMID- 10539354 TI - Tax court rules on joint ventures. PMID- 10539355 TI - What gain-sharing? Specialty hospitals in joint ventures with docs are cautious but undeterred by IRS warning. PMID- 10539356 TI - Sexual harassment policies can protect employers, too. PMID- 10539357 TI - Practice transition options for radiologists. PMID- 10539359 TI - Worried about Y2K? PMID- 10539358 TI - Pay-for-performance radiology: a new concept. PMID- 10539360 TI - Delivering radiology supplies just-in-time. AB - The radiology department at Dartmouth Hitchcock Medical Center (DHMC) adopted a just-in-time (JIT) inventory management system in 1992, reducing the volume of its in-house inventory of radiology supplies from a value of $400,000 to $16,000, just enough for four to five days of activity. An asset manager, the only person authorized to order supplies, was given responsibility for maintaining the department's supply of fixed and consumable assets. The first step in implementing the new system was to identify the supplies needed, standardize them and determine how often deliveries would be made. The JIT implementation team developed a request for proposal (RFP) that incorporated the standardized list of supplies. Three radiology supply vendors were invited to respond to the RFP. The team later determined that only one vendor was capable of implementing the JIT program. A three-year contract was awarded to that vendor. As that three-year contract reached completion, DHMC offered the JIT program to its eight affiliate hospitals and four outpatient clinics. The team decided to re-bid the contract for the entire network, which collectively performed 700,000 radiology exams annually. The new RFP encompassed 90 percent of the network's consumable supplies and offered customized delivery for each facility. The team identified eight criteria necessary for the evaluation of each vendor response to the RFP, rather than use price as the only consideration. The company that won the three-year contract furnished 90 percent of the radiology supplies for the DHMC network, allowing even further savings by the network, particularly for the smaller facilities and clinics. The program is continually monitored, adjusted and enhanced in order to incorporate changing departmental needs. PMID- 10539361 TI - Making the most of working with a consultant. AB - Key to working with a consultant is knowing when to use one, how to select the right one and how to work effectively with one. Most of the ordinary business of an organization can be carried out using its own internal resources. When this is not the case, the external viewpoint of a consultant can be helpful. Determining if a consultant can add value is considered a first step in the consulting process. Ideally, the imaging manager is involved in this step. Finding the right consultant, much like other major decisions, can be done by talking with friends or colleagues, even using the internet and AHRA Listserv. Depending on the size and nature of the engagement, a request for proposal (RFP) may be appropriate to identify the amount of expertise needed for a particular situation. When a list of potential consultants is created, the next step is to call the references of each to find the closet fit. The next step is to develop a contract or letter of understanding with the selected consultant. When agreement is reached, the consultant may then request preliminary information to begin work on the project. A further decision is to determine whether the consultant will only make recommendations or will actually help with implementation. The project typically concludes with delivery of the consultant's written report. A facility should expect some major and important new thoughts to come from the consultant's work. An effective partnership with a consultant can go far in attaining a goal of increased levels of service and decreased costs. PMID- 10539363 TI - Using instrumental leadership to manage change. AB - Effective leadership is the key factor in directing and influencing change in an organization. In today's healthcare environment, leaders have less time to make decisions that will ensure their organization's survival, a fact that has forced them to recreate the way they do business and be cognizant of ways to remain competitive. In the environment of change, a new type of leader is needed--the instrumental leader--one who combines the charismatic traits of envisioning, energizing and enabling with the three key facilitating behaviors of controlling, structuring and rewarding. To these attributes the instrumental leader adds critical thinking, problem solving and a feedback mechanism. Leaders who focus on employees as individuals and on the organization as a whole will achieve greater success. Today's leader must share a picture of the intended reality with the organization's staff, constituents and the community. Those most affected by oncoming change must be included in future planning. A shared vision is, however, an ongoing process, with communication crucial for its success. At some point in the change process, leaders must spend time building teams, clarifying behaviors and administering rewards and punishments. Part of this process involves energizing and aligning employees who will find ways to implement the shared new vision. The instrumental leader understands the need for teams that will form the backbone of the integrated system. Creating objective measures for performance, based on the new shared vision, demonstrates the organization's values better than communication or vision alone. Additional training or dialogue may better clarify the organization's expectations of employees. PMID- 10539362 TI - The development of a community breast center. AB - Maximum capacity for mammography services had been reached at the Kaweah Delta Health Care District, a 504-bed, multicampus hospital district in Visalia, Calif., so the community supported the idea of better and easier access to cancer care. Kaweah Delta Foundation, the hospital's development arm, helped raise funds for a new community breast center after hearing from local women that they disliked traveling to Los Angeles or San Francisco for state-of-the-art technology in diagnosis. They also requested better education and quicker exam results. The new Center was the result of a collaborative effort between imaging services and the cancer care program at Kaweah Delta. A nearby hospital, with more space for parking and room to offer an education program, became the site of the new Center. New equipment that met MQSA guidelines was purchased. An architectural firm designed a layout for patient comfort and privacy and efficient throughput for high volume work. The purchase of a second mammography unit allowed the Center to offer same-day and next-day appointments, which increased both physician and patient satisfaction. Consultation services with a radiologist are now offered. An education program that includes group support meetings and referrals to an oncology clinical nurse specialist are also offered. A new mobile mammography unit, housed at a newly acquired hospital 13 miles away, serves the needs of women in the two-county rural area who have no transportation. With careful planning and collaboration, the volume of mammography services has doubled in a year. Customer service ratings have soared. PMID- 10539364 TI - The check's in the mail, but death comes with it. PMID- 10539365 TI - The Arkansas healer. One gifted surgeon proves you need not go to medical meccas for top-notch care. PMID- 10539366 TI - The new Chicago hope. Can Northwestern pamper patients--and cut costs? PMID- 10539367 TI - Finding the right hospital for you. A step-by-step guide to getting the best care when you have to have it. PMID- 10539368 TI - America's best hospitals. Making the diagnosis: how we select the best. Keeping patients alive is a critical measure. PMID- 10539370 TI - Fundraising checklist. PMID- 10539369 TI - Offering new parents volunteer video support. PMID- 10539371 TI - Teetering on the edge of chaos. PMID- 10539372 TI - One foot in each world: volunteers forge links between hospitals and communities. PMID- 10539373 TI - Pen pets: a gift of love for children. PMID- 10539374 TI - Volunteer a mile in my shoes: hospital administrators see for themselves. PMID- 10539375 TI - Reading about volunteering on the Web. PMID- 10539376 TI - Virtual volunteering: helping others from your home or office. PMID- 10539377 TI - Connecting the disabled: a resource compendium. PMID- 10539378 TI - Efficacy and safety of a fixed-combination homeopathic therapy for sinusitis. AB - The efficacy and safety of a fixed-combination homeopathic medication (Sinusitis PMD) consisting of Lobaria pulmonaria, Luffa operculata, and potassium dichromate were investigated in an open-label practice-based study of 119 male and female patients, 12 to 57 years of age, with clinical signs of acute sinusitis not previously treated. At the first visit, after a mean of 4.1 days of treatment, secretolysis had increased significantly and typical sinusitis symptoms, such as headache, pressure pain at nerve exit points, and irritating cough, were reduced. Ninety-nine patients received only the test medication. Twenty patients were able to discontinue concomitant medication at the first visit. Only one patient needed an antibiotic. The average treatment duration was 2 weeks. At the end of treatment, 81.5% of patients described themselves as symptom free or significantly improved. Adverse drug effects were not reported. PMID- 10539379 TI - Transdermal fentanyl in patients with chronic, nonmalignant pain: a case study series. AB - Four cases of patients with chronic, nonmalignant pain who received transdermal fentanyl are presented. These indicate that some patients may benefit from such treatment but also serve to emphasize the importance of careful patient selection and assessment. PMID- 10539380 TI - Switching from depot antipsychotics to risperidone: results of a study of chronic schizophrenia. The Schizophrenia Treatment & Assessment Group. AB - Designed to provide information about patients with schizophrenia who switch from depot neuroleptics to the oral, atypical antipsychotic risperidone, this multicenter observational study enrolled patients who wished to stop the depot, had an unsatisfactory response, or experienced unacceptable side effects. Individuals remained on depot medication for 4 weeks and then received risperidone monotherapy for 3 months. Of the 143 patients who entered the study, 130 received risperidone, 109 completed the initial 16-week study, and 88 entered an optional 12-week follow-up. Symptoms and side effects did not change significantly during the depot phase (mean Positive and Negative Syndrome Scale [PANSS] score 72.2 at baseline, 71.6 at visit 2), but PANSS scores, global assessment of functioning, parkinsonism, and dyskinesia improved significantly during the risperidone phase (mean PANSS score decreased from 71.6 to 55.5 after 3 months). The number of contacts with healthcare professionals fell significantly during the risperidone phase; in addition, symptomatic improvements were maintained during follow-up, and movement disorders continued to decrease significantly. The investigators considered that 81% of patients had switched successfully. Patient acceptance of risperidone was significantly higher than for depot medication (83% vs 23%; P < .001), and 65% considered risperidone better than their previous treatment. Indications for depot medication should be reviewed, and patients may benefit from a switch to risperidone. PMID- 10539381 TI - Long-term results of radiotherapy alone for carcinoma of the vulva. AB - From 1953 to 1978, definitive radiotherapy for carcinoma of the vulva without surgery was used as a standard therapy at a university hospital in Germany. We retrospectively reviewed the records of 170 patients treated mainly with electron beam radiation to the vulva and Co-60 to the inguinal lymph nodes. Total doses to the vulva ranged from 6 to 90 Gy. A 30-year retrospective follow-up was accomplished. Tumor and node classification, tumor location, and parity were significant independent factors affecting survival. Actuarial survival was 52% for women with T1 disease, 25% for T2 disease, 10% for T3 tumors, and 0% for T4 tumors; survival with N0 tumors was 39% versus 13% with N2 lesions. Tumors of the labia minora were associated with a better survival rate (40.8% at 5 years) than were tumors at other locations (15%-21% at 5 years). Patients with zero to three children had significantly better results (32% at 5 years) than patients with four to seven children (11%). A dose of 60 Gy did not produce better long-term survival rates compared with lower doses, perhaps because low electron energy missed microscopic disease in deep vulvar tissues. On the basis of this review, patients with vulvar carcinoma should have at least a complete tumor resection. The therapeutic window between tumor control probability and the normal tissue complication rate in nonresected patients is too narrow: local recurrences are quite frequent with lower dose schedules, and survival rates are very poor, whereas complication rates are high with radical dose schedules that nevertheless provide acceptable cure rates. PMID- 10539382 TI - Risk factors for elder abuse. AB - The results of the first national incidence study of noninstitutionalized elder abuse and neglect in the United States are reviewed, as well as underlying causes of abusive relationships involving the elderly. It is estimated that approximately 1% to 2% of elders living in their own homes became abused in the United States during 1996, physically, emotionally, sexually, and/or financially. The abusers were predominantly adult children, spouses, and other relatives. More than 5 times as many new incidents of abuse and neglect were unreported than those reported to authorities responsible for addressing elder abuse. An individual who abuses an elder is often financially dependent on the elder, violent in other contexts, abuses alcohol and/or drugs, and has psychological problems. Although current rules and practices constrain the underwriting professional's use of this information in risk selection, public demand for financial institutions' reporting of elder abuse may provide an opportunity for open discussion about responsible handling of such cases. PMID- 10539383 TI - The journal of insurance medicine. PMID- 10539384 TI - Premalignant disease: the breast. AB - Premalignant disease of the breast is a controversial and evolving area of medical research. Breast cancer remains the number one cause of death in women in the US between the ages of 40 and 55 and ultimately causes about 4% of all deaths in women. Efforts to identify women at increased risk are of tremendous importance both clinically and from an insurance perspective. To make appropriate underwriting decisions, we need to evaluate proposed markers of increased risk with respect to what is known about how they affect short-term and/or long-term mortality. PMID- 10539386 TI - Myocardial bridging. AB - Human myocardial bridging is a normal anatomic variation in which a coronary artery is bridged by a short segment of myocardium. It can cause variable degrees of systolic obstruction. The majority of patients are asymptomatic. A wide variety of syndromes can occur, including myocardial infarction and sudden death. All patients with myocardial bridges have systolic artery compression, but it is postulated that ischemia develops only in those who have a concomitant decrease in diastolic coronary artery blood flow. Surgical removal of the myocardial bridge can be curative, and various other treatments can alleviate symptoms. The overall prognosis is good. PMID- 10539385 TI - Business quality control in issuing life insurance. AB - Fraud investigation and fraud control are entirely different processes. Similarly the auditing of a life insurance company and the issuing of life insurance policies and business quality control are not the same. Business quality control and fraud control have much in common. In this article, these similarities are explored and a case is made that companies issuing life insurance policies should consider the business quality control approach rather than the more traditional investigatory methods. PMID- 10539388 TI - Symptoms in peripheral artery disease. PMID- 10539387 TI - Submaximal heart rate response to exercise testing: independent predictor of mortality or inadequate test? PMID- 10539389 TI - Improved detection of coronary artery disease by exercise electrocardiography with the use of right precordial leads. PMID- 10539390 TI - Mortality risk in patients with coronary artery disease and depression. AB - Investigators at Duke University Medical Center studied the impact of depression on long-term mortality risk in patients with significant coronary artery disease (CAD), defined as > or = 75% narrowing of > or = 1 coronary artery. Participants were classified after testing into nondepressed, mildly depressed and moderately to severely depressed groups. From 5-10 years after the first cardiac catheterization, the mortality ratios were progressively high for the more depressed cohort. PMID- 10539391 TI - Efficacy of bilateral prophylactic mastectomy in women with a family history of breast cancer. PMID- 10539392 TI - Mortality of atrial fibrillation in a population selected to be free of major cardiovascular impairments. AB - The magnitude of additional mortality produced by the development of atrial fibrillation not associated with major cardiovascular risk factors is demonstrated. In a community-based population followed for 10 years, men aged 55 74 years had a mortality ratio of 260% and an excess death rate of 57. Women in the same age group had a mortality ratio of 335% and an excess death rate of 59. Were one to use an industry expected life table instead of the author's selected community population, the mortality ratios and excess death rates would be higher. Charging an extra premium for individuals with atrial fibrillation is supported by this increased mortality risk. PMID- 10539393 TI - Challenges to the economic evaluation of new biotechnological interventions in healthcare. AB - The next century is likely to bring unforeseen genetic and biotechnological discoveries, with new benefits, risks and costs. This paper explores some of the problem areas that healthcare biotechnologies are likely to encounter. Biotechnological interventions may present particular difficulties when they promise a major breakthrough in therapy, since economic evaluation early in the development of such technologies is inherently uncertain. Given the structure of the biotechnology industry, the perception of the high cost of new products and unknown uncertainties, governments may need to be proactive to ensure that meaningful clinical and economic data are generated in the product research phase and to manage the introduction and diffusion stages with further evaluation and surveillance. Progress usually comes at a price: both governments and the industry should prepare for this rather than offering bland reassurances about safety. PMID- 10539394 TI - A review of quality-of-life evaluations in prostate cancer. AB - Prostate cancer is a highly prevalent malignancy in older men. Because the disease and its treatments have the potential to cause substantial morbidity in affected individuals, prostate cancer has been the subject of great interest for quality-of-life (QOL) researchers. In this article, we review published QOL studies that have focused on individuals with prostate cancer. Generic survey instruments have generally been found to be insensitive to changes in health related quality of life (HR-QOL) related to prostate cancer and its treatments. Domain-specific survey instruments (such as those focusing on sexual function) have been more sensitive, but fail to capture all relevant impacts. At least 9 disease-specific instruments have been developed to measure the HR-QOL impact of prostate cancer. These instruments generally focus on specific symptoms related to the disease and its treatment--urinary function, bowel function, sexual function, physical function, psychological function and pain--however, the domains covered are not consistent from instrument to instrument, and the domains of emphasis within each instrument are rarely the same. In addition, no single instrument has been applied to all major therapies for prostate cancer across men at different ages and stages of disease. Finally, HR-QOL evaluations in some patient groups, such as those with advanced disease, have received relatively little attention to date. As a result of the proliferation of prostate cancer specific survey instruments and inconsistencies in their design and application, decision-makers face great difficulties evaluating HR-QOL across disease stages and comparing the HR-QOL impacts of alternative therapies, including conservative management ('watchful waiting'). In order for these tools to be useful for patient management and policy-making, coordination of instrument development efforts with the goal of consolidating the number of measures used is urgently needed. PMID- 10539395 TI - Review of quality-of-life evaluations in patients with angina pectoris. AB - Patients with angina pectoris have a reduced quality of life because of their symptoms, impaired activity and anxiety. However, there is no consensus on the best method of measuring quality of life. A systematic literature search of randomised controlled trials (RCTs) in angina showed that the most common generic questionnaire was the Nottingham Health Profile (NHP) Part 1, and the most common angina-specific measure was the Quality-of-Life after Acute Myocardial Infarction Questionnaire. A comparison of NHP scores with those of the healthy population revealed that patients with angina particularly seem to experience a lack of energy, poor sleep and decreased physical mobility. In the RCTs evaluated, antianginal drugs did not show a significant benefit over placebo in terms of quality of life. As a result of a lack of valid data from RCTs, a strong conclusion regarding the impact of revascularisation procedures on quality of life could not be derived. PMID- 10539396 TI - Cost effectiveness of riluzole in amyotrophic lateral sclerosis. Italian Cooperative Group for the Study of Meta-Analysis and the Osservatorio SIFO sui Farmaci. AB - OBJECTIVE: In patients with amyotrophic lateral sclerosis, long term treatment with riluzole has been reported to improve survival or tracheostomy-free survival in comparison with placebo. We conducted a pharmacoeconomic analysis for estimating the cost per life-year gained using this drug. DESIGN: This study was an incremental cost-effectiveness lifetime analysis. SETTING: The clinical material was derived from 2 placebo-controlled randomised controlled trials comparing riluzole versus usual care without riluzole, which were identified through a literature search based on the IOWA and the Medline systems. PATIENTS AND INTERVENTIONS: The study included 633 patients with amyotrophic lateral sclerosis. Patient-level information was retrieved from 313 patients treated with riluzole and 320 patients assigned to placebo. Survival after randomisation was compared between the 2 groups using standard statistics (log-rank test and Cox analysis), whereas the lifetime survival gain was estimated using Gompertz extrapolation. Cost data relative to the expenditure for healthcare resources were obtained from published information (using the US average wholesale price for the acquisition cost of riluzole). Sensitivity testing assessed the impact of different cost-of-illness assumptions for treated and untreated patients. MAIN OUTCOME MEASURES AND RESULTS: Our primary analysis showed that treatment with riluzole significantly prolonged survival [death risk = 0.77; 95% confidence interval (CI): 0.62 to 0.96; p = 0.022]. The lifetime survival gain (including 3% annual discounting) was, on average, 2.3 months per patient, while the incremental cost was around $US12,000 per patient. Hence, the cost-effectiveness ratio of riluzole versus usual care without riluzole was $US62,609 per life-year gained (discounted dollars per discounted years; 95% CI: $US13,458 to $US205,714). The sensitivity analysis, considering different values of national cost for riluzole, suggested an interval for this parameter ranging from $US45,048 to $US62,609. CONCLUSIONS: Our study indicates that in patients with amyotrophic lateral sclerosis, riluzole has an unfavourable cost-effectiveness ratio or, at best, a borderline pharmacoeconomic profile. PMID- 10539397 TI - Estimating long-term cost savings from treatment of Alzheimer's disease. A modelling approach. AB - OBJECTIVE: This paper puts forward a proposal for a modelling approach to the estimation of long term cost savings from the treatment of Alzheimer's disease (AD). DESIGN: In the proposed modelling approach, disease progression is defined in terms of intervals in the Mini-Mental State Exam (MMSE) scale. Clinical trial data are then used to determine the time at which a particular patient moved into a more severe stage of the disease. By comparing these durations across treatment groups, survival analysis is used to measure the impact of treatment in delaying the onset of a more costly stage of the disease. SETTING: Patients with varying severity of AD. PATIENTS AND PARTICIPANTS: The model uses clinical trial data on 1333 patients recruited internationally in 2 studies from 67 centres. INTERVENTIONS: The aim of these clinical studies was to evaluate the safety and efficacy of 2 non-overlapping dose ranges of rivastigmine relative to placebo over a 26-week treatment period in patients with probable AD. MAIN OUTCOME MEASURES AND RESULTS: The results indicate that the average cost savings with high-dose rivastigmine at the end of the trial period are quite low (approximately 29 Pounds per patient; 1997 values), but by extrapolating to a projected lifetime of 3 years, they rise to approximately 1100 Pounds per patient. The largest long term cost savings from treatment are obtained from treating those in the mild category (i.e. MMSE > 20). However, if the time horizon over which savings are estimated is short (i.e. if life expectancy is below 2 years), more costs are saved by prioritising patients with moderate AD (i.e. MMSE between 20 and 11). CONCLUSIONS: The model is a possible approach for estimating cost savings with treatment of AD, given the lack of long term data on resource use and drug efficacy. Caution should be used when extrapolating the results beyond the original study parameters. PMID- 10539398 TI - The influence of hospital-based prescribers on prescribing in general practice. AB - OBJECTIVE: To document the influence of hospital prescribers on prescribing in general practice. DESIGN AND PARTICIPANTS: Five percent of members of the Irish College of General Practitioners (n = 92) prospectively recorded 40 consecutive prescriptions. INTERVENTIONS: The name, dose and amount of medicine prescribed as well as the indication for therapy, details of their practice, distribution of private/GMS patients, and the number of years since qualification were recorded. The cost of individual prescriptions was calculated based on the ingredient cost and the number of days of treatment. This was subsequently correlated with the origin of the prescription. Each prescription was classified as either new or repeat. MAIN OUTCOME MEASURES AND RESULTS: Of 3286 prescriptions, 69% were for the state-supported General Medical Services (GMS) patients and 31% for private patients. Repeat prescriptions constituted 51%; 49% were new prescriptions. While hospital doctors initiated only 8% of private prescriptions, they initiated 38% of GMS prescriptions, particularly repeat prescriptions and those for cardiovascular, hormonal and centrally-acting agents. Prescriptions for anti infectives, oral contraceptives, dermatological preparations and musculoskeletal drugs were mostly initiated in general practice. The median cost for hospital initiated GMS prescriptions (5.93 Pounds) was greater than the cost of general practitioner (GP)-initiated prescriptions (3.49 Pounds; p < 0.01). Prescriptions from GPs who were qualified for less than 10 years and those with a mixed urban and rural practice were less costly (p < 0.05) than those issued by doctors qualified for over 10 years or working predominantly in an urban or rural area. These findings may also reflect differences in patient population, morbidity and demography. CONCLUSIONS: Our study indicates that hospital-initiated prescriptions are responsible for a significant proportion, both in volume and cost of GP prescribing. PMID- 10539399 TI - Cost comparison of antibacterial therapies for serious infections. A New Zealand 3-hospital study. AB - OBJECTIVE: The first aim was to identify and determine the economic costs of the regimens currently used in 3 New Zealand hospitals in the treatment of bacterial infections in haematology patients with febrile neutropenia and in intensive care patients with severe infections. The second was to develop a spreadsheet-based decision analytic model for use by hospital decision-makers as an aid in evaluating the comparative cost of drug regimens. DESIGN AND SETTING: The research utilised time and motion and microcosting techniques. The analytical perspective adopted for the study was that of a hospital administrator or clinical manager. PATIENTS AND INTERVENTIONS: Patients were eligible for inclusion in the study if either they were treated with the imipenem/cilastatin monotherapy, or could have been treated with this regimen. The final analysis considered 360 patient-treatment days and 8 antibacterials. MAIN OUTCOME MEASURES AND RESULTS: Drug acquisition cost ranged from 4.52 New Zealand dollars ($NZ; 1997 values) per patient-treatment day for gentamicin to $NZ104.81 for imipenem. The cost per patient-treatment day (when other cost components such as fluid additives, giving sets and needles were added) ranged from $NZ8.75 for gentamicin to $NZ129.12 for tazobactam. Drug acquisition cost, as a percentage of total drug preparation and administration cost, ranged from 52% for gentamicin to 93% for piperacillin. Giving sets and intravenous (i.v.) fluids were found to be important cost items when they were required specifically for the treatment regimen. There was a mean monitoring rate of 0.40 at a cost of $NZ6.41 per patient-treatment day for gentamicin. It was estimated that nephrotoxicity could add between $NZ23 and $NZ43 per day to the cost of aminoglycoside treatment. CONCLUSIONS: Although the small sample sizes of the study mean that results should be regarded as indicative rather than conclusive, there were sufficient information to construct a working model and show how the total cost of an antibacterial regimen could be evaluated in practical terms. The important cost drivers were found to be drug cost, the use of fluids and giving sets, and monitoring. PMID- 10539400 TI - Choice of cost-effectiveness measure in the economic evaluation of cholesterol modifying pharmacotherapy. An illustrative example focusing on the primary prevention of coronary heart disease in Canada. AB - OBJECTIVE: To evaluate the effect of using different cost-effectiveness measures in the economic evaluation of cholesterol-modifying pharmacotherapy. DESIGN AND SETTING: An economic model was used to examine the extent to which the relative cost effectiveness of cholesterol-modifying agents varies depending upon the cost effectiveness measure used. The perspective taken was that of the Canadian public healthcare system. PATIENTS: Individuals without coronary heart disease (CHD) with low-density lipoprotein cholesterol (LDL-C) levels in excess of 190 mg/dl. INTERVENTIONS: Cholesterol-modifying pharmacotherapies available in Canada. MAIN OUTCOME MEASURES AND RESULTS: Cost per 1% reduction in LDL-C level; incremental cost per life-year gained; least-cost agent achieving the LDL-C reduction required to meet the target level of 160 mg/dl; incremental cost per life-year gained of agents reaching the target LDL-C level of 160 mg/dl relative to no therapy; incremental cost per life-year gained of agents achieving the target LDL C level of 160 mg/dl relative to the least-cost agent reaching the target. Each cost-effectiveness measure had a different informational content to decision makers, both in terms of the usefulness of the information they provided, and in terms of the extent to which they showed one agent to be more cost effective than another. The most cost-effective treatment regimens were fluvastatin 20 mg per day, fluvastatin 40 mg per day, atorvastatin 10 mg per day and atorvastatin 20 mg per day, depending on the pretreatment LDL-C level and the cost-effectiveness measure used. CONCLUSIONS: We recommend that the cost effectiveness of cholesterol-modifying pharmacotherapy be measured using incremental cost per life year gained in reaching a predefined target LDL-C level. PMID- 10539401 TI - Using collectively-derived standards to evaluate individual performance: a cautionary note on clinical practice guidelines. AB - The purpose of this work is to demonstrate the problem of evaluating an individual physician's performance relative to practice guidelines which have typically been derived from group consensus or some measure of central tendency. It is argued that when evaluated against a set of criteria derived at the macro level, an individual physician's performance may justifiably vary due to the patient characteristics or the evolving process of care. It is also argued that it is not necessarily true that costs are reduced when practice variation is reduced. The results indicate that there are cost reduction in areas not targeted by the guidelines, suggesting a possible 'spillover effect' due to the increased vigilance in monitoring provider performance. The results also provide some evidence of increased costs following a reduction in variation. Caution should be exercised when evaluating individual physician performance relative to guidelines established at the aggregate level. Acceptable individual physician performance should be judged within the upper and lower boundaries of the implicit distribution of physicians' performances from which the established guidelines generated. PMID- 10539403 TI - Student utilization of health services at a university hospital in Turkey. AB - This paper gives the results of the first pilot study of health care utilization by Turkish university students who receive much of their student health-related services from an affiliated university hospital. The survey was distributed to 393 students and a response rate of 59.8% was obtained. In general students did not report satisfaction with the health services they received. This finding is significant because the hospital concerned is considered 'state of the art' in Turkey. Significant levels of dissatisfaction were noted across several treatment and provider variables, and comparisons with American health services are made. Suggestions for improving the student health services in the country are generated and future research recommendations are highlighted. PMID- 10539402 TI - Primary care: past and future. AB - There is anecdotal evidence of a crisis in the recruitment and retention of general practitioners nationwide and particularly in inner cities. The relationship between quality of service and single-handed or group practice is uncertain and confounded by other aspects of practice structure and populations. The review examines the factors that influence doctors to enter and remain in general practice. It explores whether initiatives designed to address problems of recruitment and retention have been evaluated in the past and suggests how could they inform current initiatives. PMID- 10539404 TI - The use of 'expectation enactment' as a conceptual framework for assessing managerial effectiveness: an NHS hospital trust example. PMID- 10539405 TI - The costs of quality: an interactive model of appraisal, prevention and failures. AB - The control of spending on health care while improving or maintaining quality is one of the most difficult problems confronting policy analysts. In this paper hypothetical data and an Excel spreadsheet are used to develop a model that estimates the costs of quality management. The focus is on the interaction between appraisal costs, prevention costs and the costs of failures. This approach enables the health service organization to estimate the costs associated with each of the three components and to assess the influence of appraisal and prevention on failure costs. The article concludes with a discussion of methods that might be used to assemble the required data and the benefits that might be derived by adopting the model. PMID- 10539406 TI - Conducting qualitative research in the health sector: researcher issues and dilemmas. AB - This article contributes to bridging the gap between research activity and the practical implementation of management decision making in the health sector by reflecting upon some of the issues and dilemmas for researchers, whether academics or managers, in conducting qualitative research in this sector. The article presents the methodological issues addressed by a team of researchers engaged on a project about manager learning and development in an NHS Trust, and highlights concerns about ethical issues that emerged from the research process. The study had involved a series of interviews with senior managers and clinical staff, doctors, nurses and therapists, and it addressed issues surrounding change within the organization, the impact on individuals' jobs, and the resultant learning and development required and undertaken. The article emphasizes that engaging in reflection on the research process is valuable and suggests that it should become a mainstream part of such research. It concludes that there is an important role of qualitative management research in the health sector and that for it to be acceptable and valued, it must be operationally sensitive, ethically robust and methodologically rigorous. PMID- 10539407 TI - Adoption of a managerial innovation: a study of physician impact analysis. AB - This article examines the adoption of physician impact analysis (PIA) among active treatment hospitals in Ontario, Canada. The influence of variables from three different levels of analysis (individual, organizational and contextual) were included as well as measures of key stakeholders' (Chief Executive Officer (CEO) and Medical Chief of Staff) assessments of the attributes of the innovation. A number of conclusions were drawn. First, by adding information about the perceived attributes of the innovation the model was able to account for a larger percentage of explained variance than has been seen in related work. Secondly, the adoption of PIA within a context of written guidelines agreed to by senior management, specifying process and structure concerns, is most likely in organizations which are large and where the CEO positively evaluates the innovation. PMID- 10539408 TI - Experiences of a heart failure clinic in preventing readmissions. PMID- 10539409 TI - Evolving practice and a culture of safety. PMID- 10539410 TI - Mother-baby transition program--one-year evaluation. PMID- 10539411 TI - See the light: Project Spectrum solves some of healthcare's most challenging IT problems. AB - BJC Health System, St. Louis. PROBLEM: Changes in healthcare, including the growth of managed care forced administrators at BJC to find new ways to address patients' needs across the entire continuum of care. To meet this challenge, disparate legacy information systems had to begin communicating. SOLUTION: BJC officials turned to IBM, along with Kodak, Southwestern Bell and Motorola, for the creation of Project Spectrum--a system that reaches beyond the boundaries of this acute care facility. RESULTS: Project Spectrum accomplishes three goals: allows access to clinical data from anywhere in the health system; is user friendly and fast; and provides the tools for better and efficient care. KEYS TO SUCCESS: "There is no quick fix. True integration means a long-term, institutional commitment to information system objectives." PMID- 10539412 TI - Standards matchmaking. PMID- 10539413 TI - Bug shots. PMID- 10539414 TI - Washington, we have a problem. PMID- 10539415 TI - "O" pioneers! Trail-blazing CIOs are leading healthcare across the CPR frontier. Interview by Charlene Marietti. PMID- 10539416 TI - Spotlight on CPR systems. Toward a CPR solution: survey of CPR vendors reflects industry growth. PMID- 10539417 TI - To err is human: systems analysis. PMID- 10539418 TI - When air sample results look bad. PMID- 10539419 TI - Strategic uses of information technology in health care: a state-of-the-art survey. AB - The general perception that the use of information technology (IT) in health care is ten to fifteen years behind IT in other industrial sectors such as banking, manufacturing, and airline is rapidly changing. Health care providers, faced with an unprecedented era of competition and managed care, are now exploring the opportunities for using IT to improve the quality while simultaneously reducing the cost of health care. A revolution is taking place in the health care industry, with IT playing an increasingly important role in its delivery. In recent years, for example, the industry spent approximately $12 billion to $14 billion a year on IT. Further exponential growth is expected as the health care industry implements electronic medical records, upgrades hospital information systems, sets up intranets for sharing information among key stakeholders, and uses public networks, such as the Internet, for distributing health-related information and for providing remote diagnostics. Along with these drastic changes and the new approach to health care, the field of health/medical informatics and telematics has also experienced significant growth in the last few years. This article identifies and surveys the critical information technologies that are being adopted to provide strategic benefits to the various health care constituencies including hospitals and health maintenance organizations (HMOs). PMID- 10539420 TI - Decision support systems in health economics. AB - This article describes a system addressed to different health care professionals for building, using, and sharing decision support systems for resource allocation. The system deals with selected areas, namely the choice of diagnostic tests, the therapy planning, and the instrumentation purchase. Decision support is based on decision-analytic models, incorporating an explicit knowledge representation of both the medical domain knowledge and the economic evaluation theory. Application models are built on top of meta-models, that are used as guidelines for making explicit both the cost and effectiveness components. This approach improves the transparency and soundness of the collaborative decision making process and facilitates the result interpretation. PMID- 10539421 TI - A decision technology system for health care electronic commerce. AB - Mounting costs have escalated the pressure on health care providers and payers to improve decision making and control expenses. Transactions to form the needed decision data will routinely flow, often electronically, between the affected parties. Conventional health care information systems facilitate flow, process transactions, and generate useful decision information. Typically, such support is offered through a series of stand-alone systems that lose much useful decision knowledge and wisdom during health care electronic commerce (e-commerce). Integrating the stand-alone functions can enhance the quality and efficiency of the segmented support, create synergistic effects, and augment decision-making performance and value for both providers and payers. This article presents an information system that can provide complete and integrated support for e commerce-based health care decision making. The article describes health care e commerce, presents the system, examines the system's potential use and benefits, and draws implications for health care management and practice. PMID- 10539422 TI - A knowledge-based patient image prefetching system: design, evaluation and management. AB - One fundamental clinical role of radiologists is to provide attending physicians with interpretations of an individual patient's radiological images essential to a treatment plan or overall patient management. Interpreting images from a newly taken radiological examination often requires reference to prior images of the same patient to establish a baseline from which to confirm a suspected pathological process or injury or to evaluate the progression of one that has been identified. Such image references are crucial to the radiologist's examination reading and when inappropriately supported can result in prolonged reading time, decreased report quality, and frustration. To address the problem of inadequate image prefetching methods used by many health care organizations, we took a knowledge-based approach and developed Image Retrieval Expert System (IRES), which incorporates relevant medical/radiological knowledge and contains image retrieval heuristics commonly shared by radiologists. This article describes the design of IRES, highlights its preliminary evaluation results, and discusses issues important for managing this and similar technologies in a health care organization. PMID- 10539423 TI - Nursing information systems: a survey of current practices. AB - This article reports the results of a recent survey on usefulness of computer based nursing information systems (NISs). To assess their usefulness, three research questions were asked: Are computer-based NISs useful to most nurses? What accounts for the nurses' assessment of NISs? What factors influence the usefulness of NISs? This framework can serve to guide empirical investigations into various aspects of information systems in hospitals. The findings lead to implications for nursing care management, as well as suggestions for nursing quality research opportunities. PMID- 10539424 TI - When health information systems fail. AB - The failure of health care information systems is a topic of critical importance for information management professionals. Such failure is also important for the consumers of health services who rely on the informed activity of health care workers for their well-being. This article presents case studies of two information systems projects within the British National Health Service that are generally viewed as having failed. The article provides an analysis of these failures, and examines whether the British National Health Service is particularly prone to the phenomenon of information systems failure. PMID- 10539425 TI - Integrating cost information with health management support system: an enhanced methodology to assess health care quality drivers. AB - Changes in health care delivery, reimbursement schemes, and organizational structure have required health organizations to manage the costs of providing patient care while maintaining high levels of clinical and patient satisfaction outcomes. Today, cost information, clinical outcomes, and patient satisfaction results must become more fully integrated if strategic competitiveness and benefits are to be realized in health management decision making, especially in multi-entity organizational settings. Unfortunately, traditional administrative and financial systems are not well equipped to cater to such information needs. This article presents a framework for the acquisition, generation, analysis, and reporting of cost information with clinical outcomes and patient satisfaction in the context of evolving health management and decision-support system technology. More specifically, the article focuses on an enhanced costing methodology for determining and producing improved, integrated cost-outcomes information. Implementation issues and areas for future research in cost-information management and decision-support domains are also discussed. PMID- 10539427 TI - Sharing in the gains: employees design their own incentive compensation program. PMID- 10539426 TI - The last phase of life: innovative approaches improve end-of-life care inexpensively. PMID- 10539428 TI - Materials pro makes waves in naval hospital. PMID- 10539430 TI - Tech assessment: just what the doctor ordered. PMID- 10539431 TI - Hard savings on soft goods. PMID- 10539429 TI - Buying on the Web: a site-seers guide. PMID- 10539432 TI - I'm from the Joint Commission and I'm here to help you ... roundtable discussion. PMID- 10539433 TI - Star wares: storage cabinets combat tight spaces. PMID- 10539434 TI - Eat your heart out. PMID- 10539435 TI - Hope meets hype. They talk about a breakthrough in Alzheimer's research, but what does that really mean? PMID- 10539436 TI - Political malpractice. PMID- 10539437 TI - Beyond Lyme: there's a new tick-borne disease to worry about. Here's what you need to know. PMID- 10539438 TI - Simplifying referrals communication. AB - Physicians at Washington University, St. Louis, use a standardized form to communicate with anticoagulation services at BJC Health System. The tool puts patient history, test scores and treatment goals in on a single page and tell staff when the physician should be notified about changes in the patient's status. PMID- 10539439 TI - PMDD diagnosable, treatable, distinct from depression. PMID- 10539440 TI - Coordinating care in an integrated delivery system. AB - To keep patients from falling through the cracks, each point of service in a healthcare system must understand its role in the patient's overall care and have access to the right information at the right time. Communication and a system's organization play significant roles in making coordinated care work: Some health systems prefer to keep patients with common diagnoses in service lines that take them from preadmission to home care; others set up separate organizational entities to coordinate care for all patients with chronic illness. Regular meetings on patient status, standardized forms and clear job descriptions all help minimize confusion. PMID- 10539441 TI - Outsmarting Alzheimer's. PMID- 10539442 TI - Is your HMO too stingy? PMID- 10539444 TI - So, how's your health? PMID- 10539443 TI - Nowhere to go for help. Isolated in Antarctica, a woman faces a cancer scare. PMID- 10539445 TI - Prescription drugs: issues of cost, coverage, and quality. AB - This Issue Brief closely examines expenditures on prescription drugs, and discusses their potential to substitute for other types of health care services. In addition, it describes employer coverage of prescription drugs, direct-to consumer advertising of prescription drugs, and potential legislation affecting the prescription drug market. Prescription drug expenditures grew at double-digit rates during almost every year since 1980, accelerating to 14.1 percent in 1997. In contrast, total national health expenditures, hospital service expenditures, and physician service expenditures growth rates decreased from approximately 13 percent in 1980 to less than 5 percent in 1997. Private insurance payments for prescription drugs increased 17.7 percent in 1997, after growing 22.1 percent in 1995 and 18.3 percent in 1996. This growth in prescription drug payments compares with 4 percent or less overall annual growth in private insurance payments for each of those three years. From 1993 to 1997, the overwhelming majority of the increases in expenditures on prescription drugs were attributable to increased volume, mix, and availability of pharmaceutical products. In 1997, these factors accounted for more than 80 percent of the growth in prescription drug expenditures. A leading explanation for the sharp growth in drug expenditures is that prescription drugs are a substitute for other forms of health care. While it is difficult to determine the extent to which this substitution occurs, various studies have associated cost savings with the use of pharmaceutical products in treating specific diseases. Evidence suggests that more appropriate utilization of prescription drugs has the potential to lower total expenditures and improve the quality of care. Also, some studies indicate the U.S. health care system needs to improve the way patients use and physicians prescribe current medications. Prescription drug plans offered by employers are likely to undergo changes to ensure that only the most efficacious drugs are covered. Anecdotal evidence suggests that copayments for prescriptions are going to increase. Some health plans are including prescription drug costs in their capitated payments to physicians. Furthermore, prescription drug plans are expected to use formularies more aggressively. In 1996, an average 5.47 outpatient prescriptions were written for those ages 55-64, compared with more than eight for those age 65 and older. Inevitably, this translated to significantly more spending for prescription drugs by the elderly. In 1994-1995, the average elderly individual (age 65 or older) spent $558 on prescription drugs, while the average 55-64-year-old spent $355. While prescription drugs are showing sharp price increases, they are also becoming more important in the treatment of many diseases. Consequently, both employers and policymakers must carefully balance the design and cost of a drug benefit so that continual innovation is preserved and the benefit can remain affordable and effective. PMID- 10539446 TI - Social Security, retirement incentives, and retirement behavior: an international perspective. AB - Escalating rates of early retirement are imposing fiscal pressure on retirement systems around the world. In some developed countries, the labor-force participation rates of men ages 60-64 have fallen by 75 percent over the last three decades. One explanation for this striking decline is social security program provisions which create disincentives to continued labor-force participation by older workers. There are substantial differences among developed nations in the labor-force participation of older workers. While two-thirds of 60 year-old American males are working, only one-quarter of men that age are working in Belgium. Over the entire 55-65 age range, 63 percent of American males are working, compared with only 40 percent of French males and 33 percent of Belgians males. There is strong evidence that the early retirement provisions of social security systems in developed countries determine the modal age of retirement. There is a strong relationship between early retirement ages and labor-force withdrawal rates; for example, in France, 60 percent of those working at the early entitlement age of 60 leave the labor force at that age. The core of this analysis is the construction of "implicit tax/subsidy rates" on additional work at older ages through each nation's social security system. These rates measure the change in a worker's retirement wealth entitlement from delaying retirement for one year, relative to the amount that would have been earned over that year. The U.S. Social Security system has an actuarial adjustment for delayed benefits claiming and other features that avoid financial incentives to leave the labor force at age 62 for a married worker, there is a slight disincentive to work for single workers and high wage earners. However, at ages 65 and older there is a stronger incentive to leave the labor force, with implicit tax rates on work of 19 percent for married workers and 33 percent for single workers. By comparison, other nations do not have actuarially fair adjustments, and as a result impose substantial taxes on additional work at older ages. In several countries, implicit tax rates on work at older ages approach or exceed 100 percent. This is because by delaying retirement, workers forgo benefits which often replace close to their full wage, in addition to having to pay the high payroll taxes required to finance generous social security benefits. There is a striking correlation across nations between high implicit tax rates on additional work and low labor force participation rates among older workers. This suggests that social security program incentives are an important determinant of retirement. These findings have important policy implications for reforming social security programs in the United States and abroad. Policymakers must consider how program reforms will affect incentives for continued work at older ages. PMID- 10539447 TI - Social Security reform: evaluating current proposals. Latest results of the EBRI SSASIM2 policy simulation model. AB - The present Social Security program has been shown to be financially unsustainable in the future without modification to the current program. The purpose of this Issue Brief, EBRI's fourth in a series on Social Security reform, is threefold: to illustrate new features of the EBRI-SSASIM2 policy simulation model not available in earlier EBRI publications, to expand quantitative analysis to specific proposals, and to evaluate the uncertainty involved in proposals that rely on equity investment. This analysis compares the Gregg/Breaux-Kolbe/Stenholm (GB-KS) and Moynihan/Kerrey proposals with three generic or "traditional" reforms: increasing taxes, reducing benefits, and/or increasing the retirement age. Both proposals would create individual accounts by "carving out" funds from current Social Security payroll taxes. This analysis also examines other proposed changes that would "add on" to existing Social Security funds through the use of general revenue transfers and/or investment in the equities market. President Clinton has proposed a general revenue transfer and the collective investment of some of the OASDI trust fund assets in equities. Reps. Archer and Shaw have proposed a general revenue tax credit to establish individual accounts that would be invested partially in the equities markets. When comparing Social Security reform proposals that would specifically alter benefit levels, the Moynihan/Kerrey bill compares quite favorably with the other proposals in both benefit levels and payback ratios, when individuals elect to use the individual account option. In contrast, the GB-KS bills do not compare quite as favorably for their benefit levels, but do compare favorably in terms of payback ratios. An important comparison in these bills is the administrative costs of managing the individual accounts, since benefits can be lowered by up to 23 percent when going from the assumed low to high administrative costs. Moreover, allowing individuals to decide whether to save the 2 percent of their OASDI taxable income or to receive higher takehome pay, as would be allowed in Moynihan/Kerrey, could lead to substantial differences in ultimate retirement income. Allowing for individual investment choices and using actual 401(k) participant allocation data, as opposed to an assumed average allocation for everyone, results in substantial differences in account balances. The Archer/Shaw approach mandates a 60 percent/40 percent equity/bond split specifically to avoid the variations in returns that arise from individual investment allocation decisions. Although there are greater chances for higher returns for equity investment in the president's proposal, there are also greater chances for worse outcomes. This is also true for other reforms that would invest Social Security assets in equities. PMID- 10539448 TI - Employment-based health insurance: a look at tax issues and public opinion. AB - This Issue Brief provides background information on the employment-based health insurance system and its alternatives. The report discusses the advantages and disadvantages of the current employment-based health insurance system, the current tax treatment of health insurance, and the strength and weaknesses of recent proposals to introduce tax credits. It presents findings from the public opinion survey conducted by the Employee Benefit Research Institute on public attitudes toward health insurance and summarizes recent research on the effects of tax changes on employment-based health benefits and the uninsured. Employment based health plans are the most common source of health insurance among nonelderly individuals in the United States, providing coverage to nearly two thirds of this population in 1997. Health insurance is probably the benefit most used and valued by workers and their families. Sixty-four percent of respondents to a recent survey rated employment-based health insurance benefits as the most important benefit. Despite essentially five years of very low health care cost increases and the recent increase in the percentage of Americans with employment based health insurance coverage, the uninsured population has continued to rise. This has resulted in a new interest among policymakers in finding ways to reverse this trend. One question that continues to be asked is whether the employment based health insurance system is the appropriate mechanism for expanding health insurance to the uninsured. Employment-based health plans are popular because they offer many advantages over other forms of health insurance and types of delivery systems. However, there are also potential drawbacks to the employment based system. The advantages include reduced risk of adverse selection, group purchasing efficiencies, employers acting as a workers' advocate, delivery innovation, and health care quality. The disadvantages include an unfair tax treatment, lack of portability and job lock, little choice of health plans, and lack of universal coverage. The tax credit proposals for health insurance, which come in all shapes and sizes, would either enhance the current employment-based health insurance system or put it at risk. This has potentially enormous public policy implications, since the vast majority of Americans get their health insurance coverage through employers. Such a change may also have political implications, as public opinion currently may not support such a fundamental change in the U.S. health insurance system. A recent public opinion survey conducted by the Employee Benefit Research Institute found that 68 percent of Americans with employment-based health insurance were satisfied with the current mix of benefits and wages. The EBRI survey found that under a changing tax code scenario, there is still strong support for the employment-based system. Strong support for the employment-based system may be the result of respondents' lack of confidence in their ability to choose the best health plan if their employer stopped offering health insurance. PMID- 10539449 TI - Lebensunwertes leben and the obligation to die. Does the obligation to die rest on a misunderstanding of community? AB - In this paper the authors address the recent argument that we have an obligation to seek or actively bring about our own death when we burden others too greatly. Some of the problems with this argument and some of the practical consequences of adopting such a point of view are discussed in this paper. We argue that the argument rests on an individualistic approach which sees the family being burdened as standing alone instead of seeing it as embedded in a burden-sharing community. PMID- 10539450 TI - Primary care groups and NHS rationing: implications of the Child B Case. AB - Implementing The new NHS and the 1997 NHS (Primary Care) Act will gradually extend cash-limiting into primary health care, especially general practice. UK policy-makers have avoided providing clear, unambivalent direction about how to 'ration' NHS resources. The 'Child B' case became an epitome of public debate about NHS rationing. Among many other decision-making processes which occurred, Cambridge and Huntingdon Health Authority applied an ethical code to this rationing decision. Using new data this paper analyses the rationing criteria NHS managers and clinicians used at local level in the Child B case; and the organisational structures which confronted them with such decisions. Primary Care Groups are likely to confront similar rationing decisions in respect of 'gate kept' NHS services. However, such rationing processes are not so easily transposed to open-access services such as general practice. NHS rationing decisions, especially in PCGs, will require a much more specific ethical code than hitherto used. PMID- 10539451 TI - A drunk driver, a sober pedestrian and the allocation of tragically scarce and indivisible emergency hospital treatment. AB - Le Grand describes a situation where a drunk driver, who has medical insurance, is the cause of an accident in which he and a sober pedestrian, who has no medical insurance, are both equally and seriously injured. At the private hospital to which they are both taken, there is available emergency treatment for one of them only. Who should receive it? The issues raised by Le Grand's example are shown to be more interesting, more complex and less clearcut than Le Grand suggests and implies. In particular, it is not the case that, unequivocally, the drunkenness of the driver establishes that the pedestrian rather than he should be treated nor that, unequivocally, the driver's possession of health insurance is morally irrelevant. PMID- 10539452 TI - A systemic and value-based approach to strategic reform of the mental health system. AB - Most writers now recognize that mental health policy and the mental health system are extremely resistant to real changes that reflect genuine biopsychosocial paradigms of mental disorder. Writers bemoaning the intransigence of the mental health system tend to focus on a small analytical level, only to find themselves mired in the rationalities of the existing system. Problems are acknowledged to be system-wide, yet few writers have used a method of analysis appropriate for systemic problems. Drawing upon the General System Theory (GST) analytical perspective, this article advances a systematic approach to understand the mental health system and to facilitate the development of reform strategies that recognize the system's complexity and changing nature. The article first discusses the failure of major reform efforts in the mental health system and the limitations of mainstream analysis of mental health politics and policies with respect to the objectives of analysis and reform. This article describes how systems thinking has thus far influenced the study of the mental health policy and politics system, and argues that a systemic perspective is profitable for reconceiving the mental health system, enabling a fresh basis for the development of reform strategies. The mental health system should be seen as a social system influenced by larger political and economic dimensions, not just as a 'delivery system' scientifically constructed by neutral experts. Furthermore, the policy planning process should be viewed as part and parcel of a mental health system modeled as complex and dynamic. The systemic perspective outlined here should help both to clarify the value-based objectives that we hold for the system and, consequently, to plan for the strategic reforms that have so far eluded us. PMID- 10539453 TI - Health care in the 21st century: what could be the shape of things to come? PMID- 10539454 TI - Is competition good for medicine? PMID- 10539455 TI - A new perspective on economic analysis in health care? A critical review of 'The Economics of Health Reconsidered' by Tom Rice. AB - A recently published book, 'The Economics of Health Reconsidered' by Tom Rice, provides a strong critique of the role of markets in health care. Many of the issues of 'market failure' raised by Rice, however, have been, to varying extents, recognised previously in the health economics literature (at least outside the U.S.). What perhaps sets Rice's book apart from previous attempts to document such issues is its elegance and the methodical manner in which this critique is delivered. Significantly the critique is based solely on conventional economic arguments. There has, however, been an emerging strand of the health economics literature not acknowledged in Rice's book which has approached some of these issues of market failure from a different perspective. Notably this research has involved, in part, borrowing from the ideas and methodological traditions of other disciplines. The emphasis in this work has been to expand the scope and the concerns of economic analysis in health care. PMID- 10539456 TI - Partnerships in healthcare: a Commonwealth perspective. AB - This article briefly outlines the nature and characteristics of partnership, and points to the need to build partnerships through deliberate efforts, since they do not occur by simply bringing people and/or resources together. It identifies some of the reasons why partnerships in health are invaluable in meeting the priority health needs of populations, and the importance which the Commonwealth places on partnerships given the unique characteristics which are common to its member countries. The article then uses practical examples to illustrate the value of partnerships in health at the national, regional and international level. For the benefit of front-line healthcare providers, some suggestions are given of how partnerships could be used to enhance the services which they give to the communities they serve. PMID- 10539457 TI - Future challenges for hospitals and health care: an international perspective on the NHS 50th anniversary. AB - This essay reviews research and poses questions for thought or discussion about the changing character of health care services and the impact of ageing, welfare, rising demand and related trends. It then asks what it would take to make the present medical service into a real health service and to empower patients and citizens. A third section worries about recruitment, retention and the morale of health care staff, and a final section considers the options for funding health care in the next 25 years. PMID- 10539458 TI - Building and funding hospitals the non-traditional way. PMID- 10539459 TI - Farewell speech of the chairman of the Dutch Healthcare Federation. PMID- 10539460 TI - Health care E-commerce and the Internet: ten strategies for health care providers and health plans doing business on the Web. PMID- 10539461 TI - Reinventing medicine on the Internet. PMID- 10539462 TI - The Internet: changing the way consumers receive health care. PMID- 10539463 TI - New HCFA rules may cut reimbursement up to 15% for outpatient services. PMID- 10539464 TI - 5 ways the new Medicare rules will change your job. PMID- 10539465 TI - What should EDs be worried about? PMID- 10539466 TI - Here are ACEP's main concerns about HCFA rules. PMID- 10539467 TI - Should you switch to template documentation? PMID- 10539468 TI - Legislation targets patient rights. PMID- 10539469 TI - Reaching the next level: governance in a time of turmoil. AB - Once considered an almost-arcane aspect of physician organization operations, governance issues are taking center stage as never before. Since medical groups, IPAs, and physician-based integrated systems face the toughest financial, operational, and strategic challenges ever in competitive healthcare markets across the country, there has never been a better--or riskier--time to test pet theories about the best governance and leadership strategies. PMID- 10539470 TI - Specialist ventures: new business partnerships with hospitals. AB - Hospital-physician partnering, a proposition perpetually loaded with opportunities and dangers, remains one of the key strategies for hospital-based organizations pursuing network- and integrated system-oriented strategies. Developing and pursuing well-thought-out models for partnerships with specialists is key to a successful outcome. PMID- 10539471 TI - President Clinton proposes $7.5 billion fund for providers; SNFs hopeful. PMID- 10539472 TI - HCFA to step up enforcement of nursing home initiatives in second year; facilities could face tougher scrutiny. PMID- 10539473 TI - Medicare cuts to SNFs projected to reach $2 billion per year; average payment, length of stay down significantly. PMID- 10539474 TI - To admit or not to admit: high-acuity residents are high reimbursement risks. PMID- 10539475 TI - Fiscal exercise: even if you don't plan to sell, you need an exit strategy. PMID- 10539476 TI - Odd couple: providers and ombudsmen find common ground. PMID- 10539477 TI - Continence by design. PMID- 10539478 TI - Playing computer catch-up: a wake-up call to the technologically timid. PMID- 10539479 TI - Prepare to win Y2K lawsuits: documentation is the key to filing or defending a claim. PMID- 10539480 TI - Finding better ways to pay for long-term care. PMID- 10539481 TI - Cost shifting: how will Medicare pay for prescription drug coverage? PMID- 10539482 TI - Overcoming denial about age. Interview by Yvonne Parsons. PMID- 10539483 TI - Dietitians in clinical roles. PMID- 10539484 TI - Violence in the workplace. Part I: Where does the problem come from? PMID- 10539485 TI - "Self-esteem" theory is waning. PMID- 10539486 TI - Computerized nutrition screening. PMID- 10539487 TI - Sizing up your nutrition care plans: 10 commandments for successful intervention. PMID- 10539488 TI - Violence in the workplace. Part II: Spotting and avoiding potential problems. PMID- 10539489 TI - Hunger and satiety: research focuses on the role of protein. PMID- 10539490 TI - Essential components of a contract: guidelines for food service and nutrition consultants. PMID- 10539491 TI - Conflict management: a fundamental part of leadership. PMID- 10539492 TI - Temperature measuring devices. PMID- 10539493 TI - The impact of schizophrenic patient functionality on service utilization and cost. Based on a presentation by Sandra L. Tunis, PhD. AB - With the advent of atypical agents in the treatment of schizophrenia, physicians and policy makers must consider the costs that may accompany greater clinical efficacy. Analyses reveal that olanzapine shows a greater clinical cost effectiveness, as well as a greater functional cost effectiveness, than haloperidol, and that functional outcomes, in particular, show promise as important measures of effectiveness. Functional outcomes can help differentiate medications and can be used to help demonstrate the cost effectiveness of atypical agents. Mental health and physical health functioning, as well as work status, are all measures of functioning that have been used to evaluate treatment strategies. When comparing olanzapine with haloperidol, cost savings are seen throughout the treatment period (1 year), with physical functioning most highly affected over time. Functional outcomes can therefore serve 2 purposes: to enhance compliance by improving health-related quality of life and to assist in making both treatment and formulary decisions. PMID- 10539494 TI - Evaluation of outcomes for atypical antipsychotic therapy and psychosocial rehabilitation in a community mental health center setting. Based on a presentation by Douglas Noordsy, MD. AB - Efficacy studies provide information on drug safety and its effect on symptoms, but their designs limit the general application of results to other settings. Functional outcomes, although difficult to measure, offer the most complete view of a medication's effect on the patient. A community mental health center (CMHC) is a common forum for treating schizophrenic patients, which presents an opportunity to study a drug's effect on patients in a natural setting. This study setting is useful because in the community patients face daily life situations, interact with family members and caregivers, and may suffer from comorbid illnesses or conditions that can affect outcomes. Douglas Noordsy, MD, Medical Director of the Mental Health Center of Greater Manchester, New Hampshire, has begun a study to examine the effectiveness of olanzapine compared with the effectiveness of typical antipsychotic medications in the CMHC setting. The initial data in Dr. Noordsy's study confirm the benefits of olanzapine for clinical symptoms and suggest positive results for functional outcomes in the future. PMID- 10539495 TI - Modeling of annual treatment costs and health outcomes of antipsychotic agents for schizophrenic populations. Based on a presentation by Josephine Mauskopf, PhD. AB - Schizophrenia is associated with the extensive use of inpatient services, so the costs of treating it are substantial. As a result, a treatment that can reduce the symptoms leading to inpatient care may decrease the overall costs and increase the quality of life for patients, their families, and society. Today, healthcare planners are faced with the delicate balancing act of providing the best care possible within limited budgets. A risk in budget assessment is looking at line items individually rather than at the total healthcare costs and clinical outcomes. For example, a drug may increase pharmacy budgets but lower inpatient services use or increase employment. A population model is described that can be used to predict the impact of new agents, such as atypical antipsychotics, and help decision makers plan budgets and revise current treatment strategies accordingly. Population models enable planners to plug in the cost and incidence numbers for their patient population and examine the total cost and patient outcomes. Such models provide a bigger picture of disease state management within a given setting. PMID- 10539496 TI - Assessment of quality-of-life outcomes in schizophrenic patients. PMID- 10539497 TI - The emerging role of psychiatric pharmacists. AB - The concept of pharmaceutical care has greatly expanded the role of the pharmacist, from that of strictly a drug dispenser to a more integrated member of a patient's healthcare team. In order for pharmaceutical care practice to succeed, the pharmacist must assume a more proactive role, using his or her knowledge of drug therapy and behavioral medicine to assume more responsibility in achieving improvement in patient health outcomes. The pharmacist must also develop open, professional relationships with patients, their families/caregivers, and other members of the healthcare team. Pharmaceutical care comprises 4 components: education, medical-legal issues, drug therapy knowledge, and communication. Through these efforts, and because pharmacists offer greater access to patients and a broader view of patient outcomes, pharmaceutical care affords the opportunity for these professionals to become patient advocates and prevent line-item decision making. Special considerations exist for psychiatric pharmacists practicing pharmaceutical care, especially in documentation and formulary decisions. Psychiatric pharmacists can ensure that patients have access to the safest, most efficacious (and cost-effective) drugs by considering more than just acquisition costs. PMID- 10539498 TI - Stats and facts. Breast cancer: progress and challenges. PMID- 10539499 TI - Women on the Internet: empowered, on-line, and making decisions. PMID- 10539500 TI - The era of accountability in behavioral health care. PMID- 10539501 TI - Raising the bar in the formulary decision process. PMID- 10539502 TI - Confidentiality: a threat for disease state management? PMID- 10539504 TI - Supreme Court approves application of racketeering statute to health insurer. AB - The Racketeer Influenced and Corrupt Organizations Act (RICO) was originally enacted to fight organized crime. Recently, however, the United States Supreme Court ruled that health insurers could be sued for fraud under the Act. This article examines the broad reach of RICO, and its potential application to MCOs in light of the Supreme Court's actions and several new lawsuits. PMID- 10539503 TI - Managed care and physician disability. AB - The number of disability claims by physicians has skyrocketed during the last decade. One of the primary reasons for this escalation is decreased job satisfaction brought about by managed care. Certain physician groups are more vulnerable to the stress of advanced managed care: solo practitioners, specialists and subspecialists, certain generalists, doctors with independent personalities, middle-aged or near-retirement physicians, impaired physicians, and those whose practices are almost solely contract driven. Based on analysis of physician disability claims, certain protective measures are recommended to relieve stress and promote survival in today's health care market. PMID- 10539505 TI - Not resting on its laurels, Sentara taps data analysis tool to fine-tune pathways. AB - Artificial intelligence data analysis leads health care system to fine-tune and improve new clinical pathways. Sentara Health Care in Norfolk, VA, developed a host of new clinical protocols in recent years, but instead of sitting back and enjoying initial improved outcomes and cost savings, it went the extra mile to enhance those pathways using a sophisticated data analysis tool. PMID- 10539506 TI - Use wireless phones to reduce unproductive labor, increase time with patients. AB - Wireless telephone systems help wipe out nurses' unproductive time. The Ohio State University Medical Center in Columbus installed a portable phone system throughout its hospital to increase efficiency in nursing shifts. The system proved so successful that more than 450 hospital personnel now use the phones. PMID- 10539507 TI - Total hip replacement surgery: comparing costs, performance against hospitals nationwide. AB - DATA BENCHMARKS: This study compares costs and performance of total hip replacement surgery against hospitals nationwide and by region. And it examines how managed care penetration in hospital markets impacts cost and length of stay. PMID- 10539508 TI - Steam sterilization takes spotlight from incineration. PMID- 10539509 TI - Innovative providers thrive under Medicaid managed care. AB - A publicly funded health care system embraces Medicaid managed care. To some safety net providers, providing quality care, serving the indigent, and providing medical training seem to conflict with managed care. See how the University Health System in San Antonio, TX, created new programs for its Medicaid and uninsured patients to meet its mission, compete with commercial Medicaid plans, and still survive. PMID- 10539510 TI - Medicaid plan, health centers reveal secrets to boosting HEDIS scores, quality of care. AB - How to do well on HEDIS measurement and boost quality of care for your Medicaid members. Neighborhood Health Plan in Boston, MA, attributes its top performance on Medicaid HEDIS measures to providers' care models, a commitment to quality, and the quest for performance data. PMID- 10539511 TI - Use these benchmarks to assess the strength of your Medicare HMO. AB - Data File: For Medicare HMOs, it's all about money. Benchmarks and trends data from InterStudy show how managed care penetration, capitation rates, and medical loss ratios can give providers clues on plan viability and performance. PMID- 10539512 TI - HCFA enrollment data show Medicaid managed care is alive, growing. AB - Also, Medicaid managed care enrollment now exceeds fee-for-service. Data recently compiled by HCFA reveal that although Medicaid managed care takes many forms, it now covers 54% of the country's total Medicaid population. Find out the latest trends in managed Medicaid enrollment. PMID- 10539513 TI - Manage your high-risk seniors to avoid financial ruin. PMID- 10539514 TI - Use physician education to cut prescription drug costs. AB - Can a physician education program effectively reduce drug costs in capitated systems? A study conducted by a Minneapolis pharmacy solutions firm suggests that physician education works. PMID- 10539515 TI - A common compensation plan can align network incentives. AB - When building a provider network through acquisition, how do you deal with the wide disparity in provider compensations and incentives? Here's one organization's solution. PMID- 10539516 TI - Use risk-reduction strategies to improve specialty cap contracts. AB - Organizations at risk for specialty care face a slew of challenges in limiting financial exposure. These risk protection strategies can improve contract performance. PMID- 10539517 TI - California IPA rings up profit with these cap rates. AB - Data Insight: Is capitation losing its luster in the Golden State? Not according to the results posted by an IPA in response to National Health Information's exclusive 1999 Capitation Survey. PMID- 10539518 TI - Competitive market analysis can reveal your core costs. AB - Peeling away layers of structure and exposing core costs allows provider groups to stay nimble in the marketplace. That's the value of a market performance analysis developed by a national consultant. PMID- 10539519 TI - Should you adopt ethics guidelines under capitation? PMID- 10539520 TI - The incidence of co-morbidities in the treatment of lymphedema. PMID- 10539521 TI - Warning: Y2K will delay payments. PMID- 10539522 TI - Perspectives. Road to Medicare FFS reform leads through middle ground occupied by private contractors. PMID- 10539523 TI - Marketplace. Buyers' market in malpractice insurance won't last as insurers stabilize prices. PMID- 10539524 TI - Perspectives. White House-CBO debate on cost of Medicare plan hinges on Medicaid, FFS reform issues. PMID- 10539525 TI - Want to bring the family into patient care? Look to pediatrics for a model. PMID- 10539526 TI - Attention, patient families: hospitals need your input. PMID- 10539527 TI - FDA: check Y2K problem list of high-risk equipment. PMID- 10539528 TI - T-graft technique promises longer-lasting bypasses: new configuration may avoid or delay re-CABGs. PMID- 10539529 TI - Electrophysiology test may predict SCD (sudden cardiac death) risk: investigators induce VT in CAD patients. PMID- 10539530 TI - Negative Medicare spending trends impact patient care, Scully tells Senate panel. PMID- 10539531 TI - Seventy-seven senators speak out on outpatient PPS. PMID- 10539532 TI - Administration announces sweeping Medicare reform proposal: plan includes billions in future hospital cuts. PMID- 10539533 TI - Prompt pay: three months is simply too long. PMID- 10539534 TI - A conversation with ... Bill Thomas, Republican Representative from California. PMID- 10539535 TI - Community Health Systems shares recipe for success. PMID- 10539536 TI - Hospital-based program improves outpatient asthma care. PMID- 10539537 TI - Will the Internet become the ultimate care management tool? PMID- 10539538 TI - Surgical treatment of early breast cancer: what would surgeons choose for themselves? AB - CONTEXT: Although breast-conserving surgery (BCS) is less invasive than mastectomy and results in similar survival, many women eligible for BCS continue to undergo mastectomy. Whether the persistent use of mastectomy means that women do not understand their options or reflects an informed preference is unknown. OBJECTIVE: To learn which treatment surgeons would choose when asked to imagine that they themselves had early-stage breast cancer. DESIGN: Cross-sectional survey. SAMPLE: Convenience sample of 40 staff and resident surgeons attending surgical grand rounds at Dartmouth-Hitchcock Medical Center in 1998. MAIN OUTCOME MEASURE: Choice of BCS or mastectomy for the treatment of stage I breast cancer. RESULTS: Twenty-six male and 14 female surgeons participated in the survey. Half chose BCS and half chose mastectomy for treatment of their hypothetical early stage breast cancer. Results did not differ by the sex of the surgeon. CONCLUSION: Even after being reminded of the equivalent 10-year survival statistics, half of the surgeons surveyed said that they would choose mastectomy over BCS for themselves. The assumption that BCS is the "right" choice for early stage breast cancer may be unwarranted because many patients may have an informed preference for mastectomy. PMID- 10539539 TI - Informed consent for PSA screening: does it happen? AB - CONTEXT: Screening for prostate cancer with serum prostate-specific antigen (PSA) is controversial. Ideally, patients should be aware of the potential benefits and risks related to testing. PURPOSE: To assess whether patients remembered having PSA screening and to determine whether they recalled having a discussion with their primary care provider about the pros and cons of such testing. METHODS: A questionnaire was sent to patients who had PSA screening ordered by a primary care practitioner during a 2-month period at a university-affiliated Veterans Affairs medical center. Approximately 3 months after the PSA test was done, patients were asked about their baseline health as well as their knowledge of and attitudes toward screening with PSA and treatment for prostate cancer. RESULTS: The overall response rate was 197 out of 421 (46%) patients. Among 173 eligible respondents without prostate cancer, 53 (31%) were unaware that their physician had ordered a PSA test. Among the 120 patients who were aware of receiving the test, only 56 (47%) recalled having a discussion with their primary care provider about the risks and benefits of screening. Support for the test was more common among patients who recalled having PSA screening than those who did not recall having the test (91% vs. 70%, respectively; P = 0.003). CONCLUSIONS: Patients who have PSA screening often are unable to recall relevant facts about the test and may have no knowledge of its associated risks and benefits. The role and effectiveness of obtaining verbal informed consent for PSA screening should be re evaluated. PMID- 10539540 TI - Women's knowledge and attitudes about genetic testing for breast cancer susceptibility. AB - OBJECTIVE: To assess female primary care patients' knowledge about breast cancer genetics and attitudes toward genetic testing. DESIGN: Self-administered survey. PARTICIPANTS: A convenience sample of 91 female patients awaiting appointments at a large primary care clinic of Group Health Cooperative in Seattle, Washington. RESULTS: Forty-seven percent of women had read or heard almost nothing about genetic susceptibility testing, and most did not know the answers to questions that assessed knowledge about breast cancer genetics. Eighty-one percent "somewhat" or "strongly" agreed that testing should be offered to everyone; women who had heard or read about genetic testing for breast cancer were more likely to agree that genetic testing should be offered only to people who have a reason to think that they have an altered gene. When asked whether they planned to have genetic testing for breast cancer, many women said "probably or definitely yes" (71% would do so if insurance covered the cost; 44% would do so even if they had to pay out-of-pocket). CONCLUSIONS: Although most women knew little about genetic testing, many expressed interest in being tested and believed that it should be offered to everyone. Primary care providers may be asked to educate women about cancer genetics and appropriate use of susceptibility testing. PMID- 10539541 TI - The Ottawa patient decision aids. AB - CONTEXT: Shared decision-making programs, or patient decision aids, have been developed for difficult decisions in which patients need to consider benefits versus risks. PRACTICE PATTERN EXAMINED: Decision aids currently used in practice in Ottawa, Ontario, Canada. DATA SOURCES: Published studies of patients faced with decisions about hormone therapy, prenatal testing, lung cancer treatments, and anticoagulation for atrial fibrillation; administrative data on distribution of decision aids; and a survey mailed to pulmonologists and surgeons. RESULTS: Although most patients considering health care options arrive for counseling with strong predispositions toward a particular option, some are uncertain about their choice and express the need for information, clarification of values, and advice about their options. Decision aids prepare patients for decision making by increasing their knowledge about expected outcomes and personal values. The aids are used in our local centers, and more than 6000 kits have been distributed in Canada, the United States, Europe, and Australia. They primarily affect the decisions of patients who are undecided at baseline and sometimes reduce the proportion of patients who choose more intensive options. CONCLUSION: The Ottawa patient decision aids assist patient decision making, particularly among those who are undecided. PMID- 10539542 TI - Valuing Viagra: what is restoring potency worth? AB - CONTEXT: The use of Viagra (sildenafil) (Pfizer, New York, New York) for treating impotence has increased dramatically. However, the cost of the drug and philosophical questions about what defines a medical condition have sparked controversy over whether insurance policies should cover impotence treatment. COUNT: The utility of life with impotence at which Viagra meets the conventional criterion for cost-effectiveness (i.e., < $50,000 per quality-adjusted life-year [QALY]). CALCULATION: We solved the following equation for utility of life with impotence: [formula: see text] RESULTS: Assuming that Viagra is used twice a week and that it costs $10 per pill, the utility of life with impotence would have to be less than 0.98 (compared with quality of life without impotence) for Viagra to meet the conventional criterion for cost-effectiveness. For patients using Viagra once or three times per week, the corresponding threshold utilities for impotence were 0.99 and 0.97, respectively. CONCLUSIONS: For men whose quality of life is sufficiently diminished by impotence, Viagra would be considered cost-effective relative to other commonly used health interventions. PMID- 10539543 TI - How can we help people make sense of medical data? AB - CONTEXT: Information is a basic prerequisite to informed medical decision making. GENERAL QUESTION: How can we help people interpret the quantitative data they need to make informed decisions? SPECIFIC RESEARCH CHALLENGE: To develop and evaluate interventions that will help people make sense of the quantitative data relevant to their health care decisions. STANDARD APPROACH: Traditional patient education interventions focus on providing disease-specific information (e.g., educational brochures about a single disease). POTENTIAL DIFFICULTIES: Interventions that focus on content--the provision of facts--may not be sufficient help for people facing medical decisions. Training that prepares people to make sense of the facts that they are given may be necessary. ALTERNATE APPROACH: We propose developing a generic (i.e., not disease-specific) tutorial to prepare people to better understand and more critically evaluate data on disease risk and the benefits and harms of treatment. This tutorial aims to improve critical reading skills by teaching people about risk (e.g., probability and rates) and showing them what to look for in statements about risk (e.g., time frame), how to put disease risk and treatment benefit in context (e.g., evaluating competing risks), how to interpret changes in risk, and whether to believe the statements about changes in risk. PMID- 10539544 TI - Patient decision making: in search of good decisions. PMID- 10539545 TI - What is a good decision? PMID- 10539546 TI - What do patients want? Help in making effective choices. PMID- 10539547 TI - Private Letter Ruling 99-10-060--joint venture diabetes clinic will not impact tax-exempt status or cause UBIT. PMID- 10539548 TI - Private Letter Ruling 99-13-035--IRS rules the formation of a limited liability company to jointly operate a hospital will not endanger 501(c)(3) exemption status. PMID- 10539549 TI - Is HMO exempt status becoming a "hot" issue?--recent status revocation and IRS business plan signal future attention. PMID- 10539550 TI - State supreme court overturns district court decision--permits property tax exemption. PMID- 10539551 TI - Supreme court decisions affecting home care. Interview by Val J. Halamandaris. PMID- 10539553 TI - Enhancing consumer independence through delegation. AB - Many forces are influencing the development of delegation options, and states are approaching the issue differently. In 1997 Oregon, Texas, New York, and Washington participated in a national symposium on delegation convened by The National Institute of Consumer-Directed Long-Term Services. These states do not necessarily represent four different approaches; however, each illustrates some of the issues involved and the ways that competing forces continue to work together to improve the policies and regulations governing delegation. PMID- 10539552 TI - Home services or euthanasia: at the heart of the debate. AB - Home care providers should be concerned about the mounting pressure to control health care expenditures and the reportedly growing public acceptance of assisted suicide and euthanasia. As the aging and disability communities organize politically for the expansion of flexible, consumer-driven, in-home support services, the euthanasia movement has begun to rally its forces behind accelerating death. PMID- 10539554 TI - Supreme Court to rule on rights of people with disabilities. AB - Since Congress enacted the Americans with Disabilities Act (ADA) in 1990, people with disabilities have demanded nationwide, "Enforce our civil rights," and "Our homes, not nursing homes." Now, in the case Olmstead v. L.C., the United States Supreme Court will decide whether the ADA will be a "civil rights statute" that enforces the rights of the disabled community and a means to achieve homebased services. PMID- 10539555 TI - Caregiving: insights into the minds and hearts of family caregivers. PMID- 10539556 TI - CHCE: professional credentialing for home care leaders. PMID- 10539557 TI - Meeting a client's spiritual needs. AB - Nurses and caregivers may feel unsure or uncomfortable discussing spiritual concerns with their patients. However, with the renewed emphasis on holistic care, the need to discuss spiritual issues arises frequently. Caregivers may gain confidence from guidelines on how to address the situation. PMID- 10539558 TI - Reaching out ... through communication. PMID- 10539559 TI - Home care and caring for the disabled. PMID- 10539560 TI - Fair play on the housing front. AB - People with disabilities are treated unfairly in many community-based housing programs. Forcing a person to participate in a program simply because he or she is a tenant is discriminatory and many advocacy groups are questioning the legality of the practice. People with disabilities must be able to choose where they wish to live and the services they need. PMID- 10539561 TI - Are you trigger-happy? PMID- 10539562 TI - Words. PMID- 10539563 TI - Taking it to the streets: telemedicine update. PMID- 10539565 TI - NAEMD (National Academy of Emergency Medical Dispatch) strives for universal certification. PMID- 10539564 TI - Enhancing a 9-1-1 service. Vermont transforms basic emergency response system. PMID- 10539567 TI - Train vs. truck: rescuers respond to Amtrak crash in Illinois. PMID- 10539566 TI - EMS hazardous materials operations. PMID- 10539568 TI - Railroad crossings: the many grades of danger. PMID- 10539569 TI - Fund drives vs. membership drives. PMID- 10539570 TI - Disease management. Health care to help you and your insurer. PMID- 10539571 TI - Changes in visitor control, security follow infant assaults at hospital. PMID- 10539572 TI - Publicizing information about baby births to media or on hospital web pages. PMID- 10539574 TI - Med center develops new electronic system to prevent baby switching. PMID- 10539573 TI - New security procedures installed after child is released by mistake. PMID- 10539575 TI - Dealing with mental hospital violence and abuse. AB - Heavy investments in security systems designed to stem violence at mental institutions in California and New Jersey have resulted from investigations by state officials. In one case, the challenge of a changing mix of mental patients, with a much greater percentage being referred by criminal courts, has spurred changes in security. In another, an extensive camera surveillance system is being installed to reduce incidents of abuse of patients by staff members. PMID- 10539576 TI - 'Code pink' infant abduction drills: what happened at two hospitals. PMID- 10539577 TI - Telemedicine for older adults. AB - Although telemedicine is not a new concept, health care assessment and treatment approaches which incorporate technology have expanded greatly over the past decade. Telemedicine has been used successfully with all age groups across the lifespan. However, telemedicine programs can serve an important function in home health care to support older adults in their own homes and communities. This article provides a summary of the types and outcomes of community-based telemedicine programs for elderly patients. In addition, practice and policy challenges in telemedicine are discussed. PMID- 10539578 TI - Factors of caregiver isolation in a rural midwest area. AB - The authors, working with a Veterans Affairs Home Based Primary Care Team in rural areas of Illinois and Indiana, noted the relative social isolation of many family caregivers of patients. They explored several factors that could contribute to this isolation: values held by the caregiver, transportation restraints, limited caregiver resources and caregiver health. Caregiver values, such as obligation and responsibility, stood out, contributing to generally excellent care for the elderly veteran patients, but also to the observed isolation. A solution would be increased funding for in-home respite, to help the family caregivers get needed rest and outings, thus responding to an expressed need, and enhancing their ability to provide "low-tech" in-home care to their loved ones. PMID- 10539579 TI - Physicians' capability in home health practice: home health nurses' perceptions. AB - OBJECTIVES: To examine home health nurses' attitudes towards physician capabilities in home health care, and whether nurses' attitudes are associated with their experience, practice setting, degree of physician interaction, or use of home health guidelines. DESIGN: A multiple regression analysis of a 90 item survey on agency characteristics, degree of interaction with physicians, and ratings of physicians capabilities across multiple dimensions of home health practice. SETTING/PARTICIPANTS: 86 registered visiting nurses from seven Chicago area home health agencies, who averaged 25 home visits and over one hour of direct contact with physicians weekly. MEASUREMENTS: Nurses' ratings of physician capability in home health practice were scaled from 18 survey items with high internal consistency reliability and correlated with nurses' practice characteristics. RESULTS: While most nurses (72%) felt that physicians responded adequately in emergencies and respected them as colleagues (70%), over 70% of respondents did not agree that physicians were adequately trained in home health. A majority of respondents rated physicians negatively on patient education, cross coverage and availability, discharge planning, support and medical supply services, and insurance issues. Respondents' years of home health experience correlated negatively (p = .004) and degree of contact with physicians correlated positively (p = .05) with ratings of physician capabilities. CONCLUSION: Nurses' attitudes about physicians' performance can provide important insights for improving the effectiveness of specialized disease and outcomes management programs which rely on care in the home setting. PMID- 10539580 TI - A national profile of primary and secondary household caregivers: estimates from the 1992 and 1993 surveys on income and program participation. AB - According to the 1992 and 1993 Surveys of Income and Program Participation (SIPP), an estimated 4,538,000 adults in the U.S. serve as primary assistants for one or more household members with disabilities. Another 530,000 serve as secondary assistants. Spouses are the main source of primary ADL and IADL assistance, while adult children and other family members are the main source of secondary assistance. Primary household assistants tend to be older and female, while secondary assistants tend to be younger and male. Primary assistants are more likely than the general adult population to rate their health as fair or poor, describe themselves as work disabled, and be limited in or need assistance with ADLs. Secondary assistants report slightly higher rates of work disability than the general population, but similar rates of ADL limitation and health status. Both assistance groups are more likely than the general adult population to have annual family incomes which fall below the federal poverty line, and working-age (aged 18-64) assistants are much less likely to work full time. PMID- 10539581 TI - Job satisfaction of home care assistants related to managerial practices. AB - This article addresses the question. "How do specific managerial practices support home care assistants' job satisfaction?" Staff from three home care agencies were surveyed regarding their perceptions of specific managerial practices and intrinsic job satisfaction. Results of a hierarchical regression model indicate that supportive leadership practices, client-centered in-service training style, and mission implementation together explained 52% of the variance in intrinsic job satisfaction. Supportive leadership was described as the extent to which a supervisor communicates effectively, shows personal concern or caring, and maintains high professional standards. Mission implementation was defined as how strongly the staff felt the mission influenced the hiring process, orientation, in-services, and everyday management. Effective in-services included discussions of types of clients and how to effectively handle common challenges. PMID- 10539582 TI - What has contributed to the change in life expectancy in Italy between 1980 and 1992? AB - Life expectancy at birth in southern Europe is known to be greater than expected in comparison with levels of economic development. This has been attributed to the 'Mediterranean diet'. There are, however, concerns that this comparative advantage is being lost. This paper examines the factors underlying changing life expectancy in Italy since 1980. The subjects of this analysis are obtained from data on all deaths in Italy between 1980 and 1992. Change in age specific death rates is calculated from selected causes and, using the method developed by Pollard, the contribution of deaths from different causes and at different ages to changing life expectancy at birth is estimated. Between 1980 and 1992, life expectancy at birth increased by 2.70 years for men and 2.75 years for women. Death rates have fallen among children and those over 40. In contrast, death rates have increased among men aged between 20 and 39 and have increased very slightly among women aged 25-29. Falling death rates from ischaemic heart disease are continuing to contribute to increasing life expectancy. Death rates from lung and breast cancer are rising among women but are compensated for by falling death rates from other cancers. Among men, falling death rates from cancer at younger ages are being offset by increases at older ages. The rising death rate among younger men is almost entirely due to AIDS, with accidents also making a small contribution. Life expectancy in Italy has improved throughout the 1980s, largely driven by falling death rates from cardiovascular diseases. Here are, however, some worrying trends, most notably the rising death rate among young men, due almost entirely to AIDS. The changing pattern of mortality has some similarities with Spain, another Mediterranean country, but there are also important differences. PMID- 10539583 TI - Productivity losses without absence: measurement validation and empirical evidence. AB - Productivity losses without absence are scarcely discussed in the literature. In this paper, the construct validity of three different measurement instruments for productivity losses without absence is investigated. The data were collected under employees of a Dutch trade firm, not in specific patient groups. On an average day, over 7% of the respondents were working with health problems, indicating that productivity losses without absence is quite a common problem. The amount of production losses related to these health problems are relatively small. However, for specific patient groups, the costs related to these productivity losses may be substantial. PMID- 10539584 TI - Association between health insurance and antibiotics prescribing in four counties in rural China. AB - A cross-sectional study was carried out at county, township and village health care facilities in four counties in rural China in order to describe and compare the effects of health financing systems on antibiotic prescribing in outpatient care. A total of 1232 outpatients at the health care facilities was selected by multi-stage random sampling and were interviewed over 2 weeks. The results showed that health financing systems appeared to influence antibiotic prescribing in outpatient care, both in terms of frequency and of the types prescribed. The insured group had lower prescribing of antibiotics at township and village health care facilities, and for respiratory tract infections, but had higher prescribing of newer antibiotics at county and village health care facilities, for respiratory tract and g-i infections. Because there was a high patient compliance rate (94.3%) in this study the prescribing of antibiotics (supply side behavior) reflected the use of antibiotics (demand side behavior) to a great extent. Thus the results imply that antibiotics prescribing and using might be biased by the patient's health financing systems and antibiotic prescribing was the result of the interaction between physicians and patients. PMID- 10539585 TI - Towards a reinforced agency role of health insurers in Belgium and The Netherlands. AB - This article describes some recent developments in health insurance in Belgium and the Netherlands. Both countries are moving towards greater financial responsibility of health insurers by means of risk-adjusted capitation payment systems. Although for the unwary observer it would appear as if both countries were following similar paths towards a common model, the authors make clear that rather different underlying rationales are driving these trends. In the Netherlands, the grand design 'Dekker proposal' for regulated competition has been replaced by a more gradual implementation of reforms with more limited scope. The ultimate goal remains a system of managed competition, albeit only for part of the health care services. In Belgium, prospective risk-adjusted capitation payment has always been at the heart of the original system in principle since its inception, but non-enforcement led to retrospective and inequitable financing in practice. Although the rhetoric of managed competition has never been used explicitly in any Belgian official government policy document, it seems unlikely that putting the insurers at financial risk without simultaneously also reinforcing their agency role by providing instruments for care management-like, for example, selective contracting--is viable in the longer run without jeopardizing the solvency of the insurers. The authors conclude that although the logic of the managed competition model is appealing, the lack of conclusive empirical evidence of success elsewhere makes governments reluctant to surrender their traditional cost containment tools. But making insurers financially accountable without simultaneously providing them with tools to take on the accountability seems useless and illogical. PMID- 10539586 TI - Is disease management relevant in Europe: some evidence from the United Kingdom. AB - Actions or approaches by the pharmaceutical industry, going under the general label 'disease management', have become very popular in the USA. However, there appears to be uncertainty about what exactly 'disease management' is and about the extent to which it can be applied in Europe. A postal questionnaire on disease management was sent out to senior personnel in the UK NHS and pharmaceutical industry. The survey aimed to explore the meaning of the term 'disease management' and its relevance to the NHS, assessing how perspectives differed between the two groups of respondents. Views on the barriers to the increase of disease management within the NHS were also sought. Finally, respondents were asked to indicate any involvement in joint disease management ventures. Most respondents agreed that disease management included estimating the total cost of managing a disease (92%) and the devising of clinical guidelines (97%). When asked about the particular role a pharmaceutical company might play, the level of agreement dropped in both groups of respondents, but by a greater degree in the NHS group. In defining disease management for themselves, just 4% of respondents referred to a 'partnership' between the NHS and the pharmaceutical industry. It would seem that, for the majority of respondents, 'joint ventures' are a possible, but not a necessary, means of undertaking disease management. Almost 30% of NHS respondents and 55% of industry respondents indicated that their Authority or company had experience of a joint venture in disease management. The major perceived barrier to an increase in disease management was NHS suspicion of pharmaceutical companies (86% of all respondents), with the difficulty in drawing up contracts coming a close second (79%). PMID- 10539587 TI - Price survey. Some hip implant prices still dropping. PMID- 10539588 TI - A school for savings: UPenn project nets $$. PMID- 10539589 TI - Policies curb, but don't eliminate, needles, latex. PMID- 10539590 TI - CHA pursues a national consensus on healthcare reform. PMID- 10539591 TI - Coming soon: second data bank on fraud and abuse. PMID- 10539592 TI - "Intranets" bring the revolution home. PMID- 10539593 TI - Talk ministry to me. PMID- 10539594 TI - Ministry hails U.S. decision on immigrants. Catholic providers hope the move will ease enrollment of Hispanic children in Medicaid and CHIP. PMID- 10539595 TI - Good news and bad. As a source of inspiration for healthcare reform, the Catholic tradition has both strengths and weaknesses. PMID- 10539596 TI - Let justice flourish. 84th Catholic Health Assembly, June 6-9, 1999. PMID- 10539597 TI - Keeping the hospitalized elderly healthy. Iowa hospital's program helps prevent function decline in older patients. PMID- 10539598 TI - Network nursing shared governance. As a healthcare network expands, nurses play key role in developing practice management. PMID- 10539600 TI - The "nameless children of Romania?" Ministry-sponsored fund helped disabled orphans in former iron curtain country. PMID- 10539601 TI - Why invest in other nation's health? Reasons Seton Institute provides aid to Third-World countries. PMID- 10539599 TI - Healthier communities through parish nursing. A South Dakota system finds multiple ways to support parish nursing programs. PMID- 10539602 TI - Helping babies grow normally. Project teaches Guatemalan mothers about the importance of infant nutrition. PMID- 10539603 TI - The spiritual side of illness. Spirit care process implements a systematic approach to spiritual healthcare. PMID- 10539604 TI - Community networks. Partnerships between Catholic charities and Catholic healthcare organizations. PMID- 10539605 TI - Videotapes promote long-term giving. PMID- 10539606 TI - Together we can do more. PMID- 10539607 TI - A profile of women CEOs/administrators in community and migrant health centers. AB - Most of the current research on women executives has focused on models in which few women achieve the highest position (e.g. hospital CEOs). This article looks at the nation's Community and Migrant Health Centers where substantial numbers of women hold the highest executive position. A national profile of women Community and Migrant Health Centers (C/MHCs) Chief Executive Officers/Administrators is provided in terms of their personal and work characteristics, as well as their values and beliefs regarding successful C/MHC attributes and important managerial practices. The study compares C/MHC Chief Executive Officers/Administrators based on gender. The study found that 41 percent of the CEO/Administrators were women and that they shared similar values and beliefs about functions/critical managerial factors and managerial characteristics of C/MHCs with their male colleagues. However, the study did find a comparable salary differential of over $11,000 in favor of male Chief Executive Officers/Administrators. The article reviews the literature of female executives in health care and concludes with recommendations for further study using the C/MHCs CEO/Administrators as a model study population. PMID- 10539609 TI - Productive working relationships in the midst of change in health care. AB - Traditional lines of authority that once provided identity and meaning in health care are blurring as delivery organizations adapt to the challenges of controlling cost and providing quality service. Teams and committees tackle work that was once the realm of individual managers. Men and women at different levels of authority in the organization now work together to make decisions. Previous lines of authority in the health administration education setting are also blurring as colleges and universities, under pressure, respond to customer needs. Academia is increasingly drawing upon adjunct and non-tenure-eligible professionals to streamline departments and save money. Within the structure of a department, workers in these temporary positions have less authority than tenured and tenure-track faculty. They have less authority in the classroom as well. In both the health care industry and academia, women endure these changes more than their male counterparts, since women often assume the variety of flexible roles required by these strategies. Changes in traditional academic authority produce anxiety and stress for everyone involved. However, faculty can teach students a flexible paradigm to navigate and find meaning in these situations to ensure successful and productive working relationships between men and women in the changing workplace. This paper identifies the pertinent components of this paradigm and its application in the health administration classroom. PMID- 10539608 TI - Career characteristics among graduates of a Midwestern M.H.S.A. program: variation by gender and length of time since graduation. AB - This study examined whether differences occur in the careers of men and women graduating from a Midwestern Master's degree program in hospital/health services administration (M.H.S.A.) during the calendar years of 1986-1997. These alumni were divided into two cohorts: 1986-90 and 1991-97, so that temporal differences could be examined. Men and women M.H.S.A. graduates of the 1986-90 cohort are currently working in similar settings. The most common setting is in acute care organizations. Men and women M.H.S.A. graduates in the 1991-97 cohort tend to be working more outside of acute care organizations than the 1986-90 graduates. The 1986-90 group of women graduates are currently in significantly different positions than their male counterparts. Compared to their male colleagues, these women are more likely to be in middle management positions, or out of the work force. A higher percentage of men from the 1986-90 cohort hold senior management positions (such as CEO or vice-president). The 1991-97 cohort of women graduates do not currently hold significantly different types of positions than their male colleagues, though about twice as many women were not working. PMID- 10539610 TI - The glass ceiling in academe: health administration is no exception. AB - This paper reviews gender issues in academe and presents findings of a limited survey of ACEHSA-accredited health administration graduate programs. The survey shows gender ratios adverse to women at the full, associate, and assistant professor levels. Men to women ratio among faculty was 1.98, among full-time faculty it was 2.24, and among tenured/tenure-track faculty it was 2.69, despite an excess of female students over male students in graduate programs, and despite equal proportions of women and men faculty holding doctoral degrees. Distribution by rank showed 48.5 percent full professors, 27.8 percent associate professors, and, 20.1 percent assistant professors among men, vs. 27.4 percent, 41.1 percent, and 31.5 percent respectively among women. In other academic fields similar gender ratios prevail, and many researchers have documented evidence of continuing gender inequities in tenure, promotion and salary, given comparable performance, despite the enactment of Title IX in 1972. Gender disparities are rooted in a complex web of gender-specific constraints interwoven with secular human capital and structural variables, and confounded by sexist discriminatory factors. In light of these issues, recommendations are made toward creating an equitable academic climate without compromising the ideal of meritocracy, through gender-sensitive initiatives and vigilance mechanisms to bring policies to fruition. PMID- 10539612 TI - Raising the "glass ceiling" for ethnic minority women in health care management. AB - Ethnic minority women are well represented in the work force and in the health care system in general, but do not have a similar level of representation in the management sector. This paper explores three strategies for schools of health administration to consider to lessen the effect of a "glass ceiling" that may be encountered by ethnic minority women aspiring to positions of leadership in health services agencies. These strategies are advancing affirmative action, valuing ethnic women in health administration education, and investigating diversity management. Inherent in each of the three strategies is the need for acknowledgment and more open discussion of the "glass ceiling." Problem-solving in relation to the potential for systemic discrimination adversely affecting ethnic minority women in senior health care management positions, and greater study of the three strategies using both qualitative and quantitative methodologies is also needed. PMID- 10539611 TI - Promoting career mobility of women faculty in health administration programs. AB - Faculty women have encountered the same glass ceiling in their pursuit of better paying, higher level positions that other women have experienced in the work world. Numerous factors have contributed to the continued inequality of salary and rank within the academic community. In order to overcome these barriers, faculty women and women executives in health administration have identified and implemented successful strategies. This article describes these barriers and discusses their impact on the career paths of faculty women and women graduate students in health administration programs. The strategies of role modeling, mentoring, and networking were examined in terms of their benefits, problems, and related gender factors. Finally, recommendations that will complement these strategies were determined and discussed. PMID- 10539613 TI - Addiction and discounting. AB - In 1988, Becker and Murphy [Becker, G.S., Murphy, K.M., 1988. A theory of rational addiction. Journal of Political Economy, 96, 675-700.] launched a theory in which they proposed that the perspective of rational decision-making could be applied also to cases of addictive behaviour. This paper discusses the theory's assumptions of interpersonal variation and stability in time preferences on the basis of estimates derived from three groups of people with different consumption levels of illegal intoxicants. We find that active injectors of heroin and amphetamine have a higher discount rate than a group reporting that they have never used the substances. Of greater interest, though not in accordance with Becker and Murphy's assumption of stability, we also find that the discount rate among active and former users differs significantly. These findings raise the question of whether a high time-preference rate leads to addiction or whether the onset of an addiction itself alters people's inter-temporal equilibrium. PMID- 10539614 TI - Firm behavior in a market with addiction: the case of cigarettes. AB - This paper investigates firm behavior when demand is linked over time. Among other things, the theoretical section shows that if firms are forward-looking, anticipated future events can affect current consumption of an 'addictive' good even when consumers are completely myopic. The empirical part of the paper reports a simulation of the 1983 federal cigarette excise tax increase. Both myopic and rational models of consumer demand give roughly the same predictions for per capita consumption, but neither model does very well predicting actual consumption. The problem appears to lie in the prediction of price, suggesting that supply considerations are important. PMID- 10539615 TI - An econometric analysis of smoking prevalence among lone mothers. AB - This paper uses panel data to examine the determinants of smoking among lone mothers over the period 1991-1996. Consideration is given to the initial conditions problem encountered when modelling dynamic panel probit models, and a recently suggested approach is applied to address this problem. PMID- 10539616 TI - Would you like suspenders to go with that belt? An analysis of optimal combinations of cost sharing and managed care. AB - When and why would it be efficient for a managed care insurance plan using managerial limits to add patient cost sharing? This paper uses a diagrammatic model to indicate that the use of patient point-of-service cost sharing can cause the managerial limits or guidelines to be less restrictive in limiting high value care for cases of severe illness. The model shows that cost-sharing is more likely to improve efficiency the greater the variation in illness severity and the smaller the degree of moral hazard. The model is extended to the case in which provider cost sharing is also used. PMID- 10539617 TI - Cost-sharing and pharmaceutical utilisation and expenditure in Russia. AB - This paper presents estimates of the impact of exemption status, and other socio economic variables, on pharmaceutical use in Russia. Estimates are derived from a newly collected household survey covering around four thousand households. Separate results for a zero-inflated negbin model of utilisation of prescriptions and for a two-part model of the overall level of household expenditure on pharmaceuticals are presented. Full exemption from prescription charges is shown to increase the utilisation of prescription items and reduce the probability of the households incurring drug expenditure. PMID- 10539618 TI - Physician fees and procedure intensity: the case of cesarean delivery. AB - While there is a large literature investigating the response of treatment intensity to Medicare reimbursement differentials, there is much less work on this question for the Medicaid program. The answers for Medicare may not apply in the Medicaid context, since a smaller share of a physician's patients will be Medicaid insured, so that income effects from fee changes may be dominated by substitution effects. We investigate the effect of Medicaid fee differentials on the use of cesarean delivery over the period 1988-1992. We find, in contrast to the backward-bending supply curve implied by the Medicare literature, that larger fee differentials between cesarean and normal childbirth for the Medicaid program leads to higher cesarean delivery rates. In particular, we find that the lower fee differentials between cesarean and normal childbirth under the Medicaid program than under private insurance can explain between one half and three quarters of the difference between Medicaid and private cesarean delivery rates. Our results suggest that Medicaid reimbursement reductions can cause real reductions in the intensity with which Medicaid patients are treated. PMID- 10539619 TI - The impact of malpractice fears on cesarean section rates. AB - A longstanding issue in the health care industry is whether physicians' malpractice fears lead to defensive medicine. We use national birth certificate data from 1990 through 1992 to conduct a county fixed-effects analysis of the impact of malpractice claims risk on cesarean-section rates and infant health. Malpractice claims risk is measured by obstetricians' malpractice premiums. The study provides evidence that physicians practice defensive medicine in obstetrics but that the impact of increased cesarean sections that results from malpractice fears on total obstetric care costs is small. The study also finds that physicians' defensive response varies with the socioeconomic status of the mother. PMID- 10539620 TI - Pastoral counseling as spiritual healing: a credo. AB - Notes that pastoral counseling is an ancient ministry which brings rich resources to the contemporary efforts to develop wholistic healing. Provides a historical background summary, a sketch of Judeo-Christian tradition, and clinical examples to illustrate ways in which modern explorations in mind/body wellness can be enhanced by including the pastoral counseling project. PMID- 10539621 TI - An interdisciplinary approach to integrative professional education. AB - Reports on a professional formation course in integrative clinical learning for senior nursing students. Outlines the objectives and methods utilized in the course. Concludes that, given the characteristics and results of the course, vocational formation for ministry needs to include similar dimensions in designing educational curricula. PMID- 10539622 TI - Designer genes: three photographs of the future and a call for debate. PMID- 10539623 TI - Dealing with cultural diversity: a hospital chaplain reflects on Gypsies and other such diversity. PMID- 10539624 TI - Describing what chaplains do in hospitals. AB - Reports on a research project that describes the unique activities in which chaplains engage as they provide pastoral care and counseling and spiritual direction to patients in a large, acute tertiary referral medical center in Melbourne, Australia. Analyzes 57,704 entries which described what hospital chaplains do when interacting with patients over a period of five and one-half years. Identifies as the core activities "promoting spiritual transcending" (57.6% of all entries), "promoting spiritual intactness" (36.1% of all entries) and "enacting ministry" (6.2% of all entries). PMID- 10539626 TI - Malpractice wars: brace yourself for a surge in premiums. PMID- 10539625 TI - Personal reflection--heart transplant. PMID- 10539627 TI - Malpractice survey snapshot: how's your insurance coverage? PMID- 10539628 TI - Malpractice wars. The newest threat: pressure to go beyond your expertise. PMID- 10539629 TI - Malpractice survey snapshot. Managed care's impact: does managed care increase malpractice risk? PMID- 10539630 TI - Malpractice wars. Tort reform battles: sometimes the force is with you. PMID- 10539631 TI - Malpractice survey snapshot. One guess who doctors blame--and what remedies they favor. PMID- 10539632 TI - HMO disputes. When patients and HMOs disagree. PMID- 10539633 TI - The Feds' good cop, bad cop routine. PMID- 10539634 TI - Malpractice wars. Our national survey confirms: even the best doctors are targets. PMID- 10539635 TI - Malpractice survey snapshot. Who's been sued--and who won. PMID- 10539636 TI - Malpractice wars: the sneaky games lawyers play. PMID- 10539637 TI - Malpractice survey snapshot: once burned, twice defensive. PMID- 10539638 TI - Malpractice wars: how doctors sabotage their own defenses. PMID- 10539639 TI - Is the managed care honeymoon over? PMID- 10539641 TI - Handhelds capture charges, speed billing cycle. PMID- 10539640 TI - Current group practice dynamics. PMID- 10539642 TI - Using compensation to improve primary care. AB - Progressive primary care networks are now placing significant portions of physician salaries at risk by linking compensation to quantifiable measures such as net medical revenue (collections), reduced practice expenses, cost and utilization, quality of care and patient satisfaction. For most networks, a combination of productivity increases and expense reductions are critical to ensure financial survival. This case study illustrates how one network's unique incentive compensation program targeted higher productivity levels by incentivizing desirable behaviors. PMID- 10539643 TI - The 'fixed cost effect' on practice management. AB - To obtain a better understanding of the behavior of "non-professional" costs in a medical practice, the authors analyzed the expenses of a 19-doctor practice. The analysis revealed that 80 percent of these expenses were fixed costs. Fixed costs, as opposed to variable costs, remain static in total but vary on a per unit basis as volume changes. Organizations with high fixed cost must maximize capacity to achieve profitability. Thus, the relationship among volume, capacity, cost and profit must be understood by medical practices negotiating rates for service units. PMID- 10539644 TI - How to design a hospitalist service. An academic health system case study. AB - Medical centers throughout the country are employing hospitalists--dedicated inpatient specialists--to care for patients during hospitalization in place of their primary care provider. There has been expansive growth of hospitalist programs as the number of practicing hospitalists has doubled in two years from approximately 1,500 to over 3,000. In July 1997, the department of medicine at University Hospitals of Cleveland, the primary teaching affiliate of Case Western Reserve University, created a hospitalist service. This paper will discuss the design of a hospitalist service at a major academic medical center, the factors that were considered during the design stage and financial feasibility. PMID- 10539645 TI - How to select and motivate physicians in managed care. AB - Recruiting a physician can be an extremely beneficial or an extremely costly move for any health care organization. The emotional matching of the person is always important but the ability of the new health care provider to operate efficiently and effectively is paramount to their success. This selection process begins even before the recruitment process and includes monthly meetings with physicians to provide feedback and discuss performance while they are practicing. This article addresses the needs of the several different managed care environments and offers insights to setting up effective utilization management. PMID- 10539646 TI - Changing dynamics in employer-sponsored health insurance. One market's perspective. AB - Using their sponsored health benefits as a mechanism for change, employers have been able to exert significant influence over the nation's health care system. By examining how employers design, purchase and manage these programs, much insight can be gained. Twenty-five mid- to large-sized companies in a Middle-Atlantic metropolitan area were interviewed during May-July 1998. The study was modeled after a similar effort conducted in 1991, the results of which were published in this journal in 1993. The study found that many of the dynamics in employer sponsored health insurance are changing. The findings suggest that a more distanced relationship between employers and employees is the major factor underlying the evolution, an intentional change to force a shift in medical care decision-making and responsibility to more of a shared process between the employee and the employer. These trends have important implications for local markets relevance and the national situation. PMID- 10539647 TI - Question: what is a "better-performing practice? PMID- 10539648 TI - Table of reference intervals. PMID- 10539650 TI - Critical values for drug levels. PMID- 10539649 TI - Reference laboratories for rare tests. PMID- 10539652 TI - Directory of diagnostic marketers. PMID- 10539651 TI - Table of critical limits. PMID- 10539653 TI - Cut-off and toxicity levels for drugs-of-abuse testing. PMID- 10539654 TI - Directory of professional organizations. PMID- 10539655 TI - MLO's national salary survey: MTs and MLTs respond. PMID- 10539656 TI - Index of tests, equipment, and services. PMID- 10539657 TI - Continuing education resources for laboratorians. PMID- 10539658 TI - Planning and implementing total laboratory automation at the North Shore-Long Island Jewish Health System Laboratories. AB - Lab automation and consolidation can be a daunting, risky, major reengineering project. Done right, it can mean decreased labor costs and space requirements, increased test volume, and more efficient use of personnel. See how this health system got the job done using a carefully defined, seven-step plan. PMID- 10539659 TI - How to defuse compliance time bombs. Part 2: Six hypothetical allegations. Panel discussion. AB - In the second and final part of this series, a panel of legal experts discusses more hypothetical allegations against a fictional laboratory, including billing for additional indices, using expired reagents, and falsifying the results of employee competency tests. Learn how to handle these dilemmas before they explode into full-blown legal violations. PMID- 10539660 TI - Reading the future: the increased relevance of laboratory medicine in the next century. AB - Through intelligent process control and data management, the laboratory may become the most frequently used--and the most important--source of diagnostic information in medicine. The central laboratory of the future is destined to become an esoteric testing center, whereas routine testing--administered at the patient bedside or at home--will become more economical. Point-of-care testing will soon become the most profitable way to provide laboratory services. Novel phlebotomy techniques and noninvasive tests may allow some diagnostic testing to be done through automated robotic companions that serve homebound patients or the elderly. PMID- 10539661 TI - A brief history of medical diagnosis and the birth of the clinical laboratory. Part 1--Ancient times through the 19th century. AB - From tasting urine to microscopy to molecular testing, the sophistication of diagnostic techniques has come a long way and continues to develop at break-neck speed. The history of the laboratory is the story of medicine's evolution from empirical to experimental techniques and proves that the clinical lab is the true source of medical authority. Part 1 in a 2-part series. PMID- 10539662 TI - Beverly may pay $225 million settlement. PMID- 10539663 TI - Picking up the pieces. After a long delay, Mo. hospitals try to resume merger plans. PMID- 10539664 TI - Healthcare mergers, acquisitions plunge. PMID- 10539665 TI - Mich. group achieves direct contracting. PMID- 10539666 TI - Tenet disappoints with earnings report. PMID- 10539667 TI - Beltway choice: big deal or train wreck. PMID- 10539668 TI - Physician compensation growth slows. PMID- 10539669 TI - Alexian system focuses on Chicago. PMID- 10539670 TI - Onus is on providers to handle Medicare Y2K. PMID- 10539671 TI - Investor jitters lead to grim six months. PMID- 10539672 TI - AHA to spend more fighting budget law. PMID- 10539673 TI - Hanging together. Flurry of legislative activity spurs some furious coalition building in healthcare industry. PMID- 10539674 TI - Has lap general surgery hit a plateau in growth? PMID- 10539675 TI - Bills would blunt impact of outpatient pay plan. PMID- 10539676 TI - A code of conduct for the OR. PMID- 10539677 TI - Surgery beyond the laparoscope. PMID- 10539678 TI - Benchmarks. Saving on supplies for AV shunt. PMID- 10539679 TI - Good analysis key to outsourcing. PMID- 10539680 TI - Getting ready for surgery online. PMID- 10539681 TI - Using data to measure improvement efforts. PMID- 10539682 TI - A checklist on exposure safety. PMID- 10539683 TI - What's best for monitoring steam? PMID- 10539684 TI - New standard on patient rights. PMID- 10539685 TI - Don't put off preparing for APC payment system. PMID- 10539686 TI - How can you improve end-of-life decision making? PMID- 10539687 TI - Passing the buck: accountability is everyone's responsibility. AB - In the rush to make a deal, corporate accountability often gets lost in the equation. Health care trustees need to ask tough questions, demand answers, and hold management's feet to the fire. BOTTOM LINE: if you don't do this, state attorneys general are itching to take control. PMID- 10539688 TI - The hot breath of the IRS: how trustees can avoid intermediate sanctions. AB - New regulations from the IRS, called "intermediate sanctions," impose stiff taxes and penalties on members of not-for-profit organizations responsible for "unreasonable" compensation. That means administrators, physicians, trustees, and others must be careful to comply with the letter of the law or risk having the IRS breathing heavily down your neck. PMID- 10539689 TI - There's no more room on the sidelines. PMID- 10539690 TI - Focusing on quality through employee satisfaction. PMID- 10539691 TI - Put yourself in the picture: planning by strategy development. PMID- 10539693 TI - The curse of Frankenfood. Genetically modified crops stir up controversy at home and abroad. PMID- 10539692 TI - When you hear that whistle blowing. How trustees can reduce the threat of "qui tam" lawsuits. AB - The number of qui tam--whistle-blower--lawsuits against hospitals has increased 1,500 percent in the last decade--in large part because of the high priority the government attaches to Medicare fraud and abuse. Even suits that never make it to court are financially draining and a blow to an organization's reputation. Here are some tips for protecting your hospital. PMID- 10539694 TI - Losing your mind? Memory lapses are usually normal, but some may signal deeper problems. PMID- 10539695 TI - Lots of lifts are for dumbbells: you can pump iron all day to get big muscles. Or try 30 minutes instead. PMID- 10539696 TI - Waiting for a chance to live: lack of organ donors undercuts allocation-reform efforts. PMID- 10539697 TI - In stroke, the immune system turns traitor. PMID- 10539698 TI - New hope from a 40-year-old drug: deaths from heart failure cut by 30 percent. PMID- 10539699 TI - Functional MRI explores mysteries of acupuncture. PMID- 10539700 TI - Technologists competent to perform basic GI exams. PMID- 10539701 TI - Government regulations may hinder fellowship training. PMID- 10539702 TI - Anxiety of MR patients extends beyond claustrophobia. PMID- 10539703 TI - Radiology networks pursue contracts while groups stay independent. PMID- 10539704 TI - Technologies that are changing the practice of radiology. PMID- 10539705 TI - Plan ahead to optimize clinical practice of fetal ultrasound. PMID- 10539706 TI - The conventional ultrasonic nebulizer proved inefficient in nebulizing a suspension. AB - A study was undertaken to compare the amount of nebulized budesonide suspension and nebulized terbutaline sulphate solution inhaled by healthy adult subjects when conventional jet and ultrasonic nebulizers were used. Ten healthy subjects (5 male; age range, 16-52 years) used two conventional nebulizers: the Spira Elektro 4 jet nebulizer (Respiratory Care Center, Hameenlinna, Finland) and the Spira Ultra ultrasonic nebulizer (Respiratory Care Center) in a breath synchronized mode with each drug. The amount of drug inhaled, the inhaled mass, was defined as the amount of drug deposited on a filter between the inspiratory port of the nebulizer and the mouthpiece. The amount of budesonide and terbutaline sulphate was determined by reversed-phase high-performance liquid chromatography. Single-dose respules were used (0.5 mg of budesonide and 5.0 mg of terbutaline sulphate), and nebulization time up to the defined gravimetric output was recorded. The inhaled mass of budesonide varied depending on the nebulizer used, whereas the inhaled mass of terbutaline was unaffected by the choice of nebulizer. The median inhaled mass of budesonide was 31.4% of the nominal dose (i.e., dose of drug in the respule per label claim) with the Spira Elektro 4 and 9.9% with the Spira Ultra, whereas the median inhaled mass of terbutaline was 50% with the Spira Elektro 4 and 52% with the Spira Ultra. It appears that a suspension is generally more difficult to nebulize than a solution and that the budesonide suspension should not be used in conventional ultrasonic nebulizers. PMID- 10539707 TI - Inspiratory flow rates through a novel dry powder inhaler (Clickhaler) in pediatric patients with asthma. AB - Inspiratory flow profiles through a novel dry powder inhaler (DPI) (Clickhaler; Innovata Biomed Ltd, St. Albans, UK) were recorded in 17 pediatric patients (aged 7-16 years) with stable mild to moderate asthma. Most patients (n = 14) could generate high peak inspiratory flows (PIFs) (> 60 L/min) and high flows early in the flow profile. These flows have been shown to be adequate to deaggregate and deliver micronized drug with this delivery system. PMID- 10539708 TI - Bench testing of nebulizers: a comparison of three methods. AB - Although nebulizers can vary widely in performance, there is no uniformly accepted method for bench testing these devices. In the present study, we compared three bench methods of measuring the performance of three commercial jet nebulizers (Whisper Jet [WJ; Marquest Medical, Englewood, CO], Sidestream [SS; Marquest Medical], and Vixone [VO; Westmed, Tucson, AZ] to assess the impact of the method of testing on reported nebulizer performance. Each nebulizer was charged with 3 mL of albuterol mixed with a radiotracer (technetium [99mTc]), and the radioactivity captured on a paper filter was expressed as a percentage of the nebulizer charge (% delivered). The nebulizers were tested with and without duplication of spontaneous respiration by a piston pump (spontaneous respiration and standing cloud methods, respectively). The nebulizers were also tested using a model of mechanical ventilation (mechanical ventilation method). For all three devices, the addition of the standardized breathing pattern significantly reduced the % delivered with all three nebulizers compared with the standing cloud method. For the standing cloud method, the presence of the T-piece/mouth-piece significantly reduced the % delivered with the WJ but not with the other two devices. The mechanical ventilation method had the lowest % delivered for all three devices. The magnitude of the differences between nebulizers varied with duration of treatment. The findings of this study emphasize the importance of bench testing that duplicates intended clinical usage, because significant differences in nebulizer performance may be manifested under certain clinical conditions but not under others. PMID- 10539709 TI - Effects of equipment dead space and pediatric breathing patterns on inhaled mass of nebulized budesonide. AB - Inhaled mass, the quantity of aerosolized drug actually inhaled by a patient, can be estimated in vitro by using a piston pump and an inhaled mass filter in a manner that simulates in vivo aerosol delivery. For pediatric patients, measurement with an inhaled mass filter with a large equipment dead space (VDEQ) in relation to a small tidal volume (VT) may underestimate the inhaled mass. The present study investigated the impact of VDEQ on the accuracy of in vitro measured inhaled mass of budesonide suspension for nebulization using Spira Module 1 jet nebulizers (Respiratory Care Center, Hameenlinna, Finland), inhaled mass filters, VDEQs of different sizes, and pediatric breathing patterns. The VDEQ varied between 14 and 108 mL, and the breathing patterns corresponded to a VT between 50 and 500 mL, to breathing frequencies of 40 to 12 per min-1, to a duty cycle of 0.5 for all breathing patterns, and to a nebulization time of 2 minutes. The results showed that the inhaled mass was a function of the VDEQ for each breathing pattern as defined by the inspiratory minute volume (VI). For a large VT, a small VDEQ affected the inhaled mass of budesonide only marginally, but as the VDEQ increased, the measured inhaled mass decreased to the point that for a VDEQ larger than the VT, the inhaled mass was zero. When the inhaled mass was expressed as a function of an effective volume (VEFF) (i.e., VI corrected for VDEQ), the results showed a linear correlation (R2 = 0.921) between the volume of aerosol inhaled through the nebulizer and the inhaled mass of budesonide. The results of the study indicate that VDEQ has a critical effect on the measurement of inhaled mass in vitro for conventional jet nebulizers using pediatric breathing patterns. This means that the in vitro measured inhaled mass of drug can seriously underestimate the in vivo value. When pediatric breathing patterns are used in vitro, a correction of the VT by the VDEQ should be made in order to more accurately reflect the in vivo inhaled mass of drug. PMID- 10539710 TI - A critical comparison of the dose delivery characteristics of four alternative inhalation devices delivering salbutamol: pressurized metered dose inhaler, Diskus inhaler, Diskhaler inhaler, and Turbuhaler inhaler. AB - Salbutamol is a short-acting beta 2 agonist which is effective as a rescue therapy in the treatment of asthma. This study uses in vitro test methods to compare the capability of four alternative devices to deliver an accurate and precise dose of salbutamol. It is demonstrated that the conventional metered dose inhaler (MDI) achieves excellent accuracy and precision in dose delivery. Additionally, it is the most efficient inhaler in terms of generating in-vitro a fine particle fraction from the dose. A spacer device has been shown to further enhance the dosing characteristics. When tested over a wide range of inspiratory air flow rates, the Diskus (GlaxoWellcome, Hertfordshire, UK) has comparable accuracy and precision to the MDI tested at 60 L/min, and it offers an advantage over two alternative dry powder inhalers (DPIs), delivering a more consistent dose across the range of flow rates tested and being more efficient at generating a fine particle fraction than either Turbuhaler (Astra, Lund, Sweden) or Diskhaler (GlaxoWellcome) at both 28 and 60 L/min inspiratory flow rates. Diskus, Diskhaler, Ventolin, Volumatic, and Rotadisk are trademarks of the GlaxoWellcome Group of companies. The Accuhaler is the alternative to the Diskus in those countries where the Diskus trademark is not available. Inspiryl and Turbuhaler are trademarks of the Astra Group of companies. PMID- 10539711 TI - An in vitro evaluation of a self-contained cardioplegia delivery device. AB - Focus on improved myocardial protection has prompted the development of delivery systems which can accommodate the demands of increasingly refined cardioprotective strategies. The purpose of this study was to evaluate the Sorin Blood Cardioplegia Console (BCC) for accuracy and precision in the delivery of cardioplegia solutions. An in vitro model was designed to evaluate the following performance characteristics of the BCC: delivery volume (blood and crystalloid) at 50, 250, 375, and 500 ml/min flow rates; potassium end-delivery concentration at blood:crystalloid ratios of 4:1, 8:1, and 16:1; intermittent cardioplegia delivery; and heat transfer of the internal heater/cooler. Differences in blood and cardioplegia volumes between measured and calculated values across all flow rates tested were found to be statistically significant and ranged from 1.4 to 21.5 ml; however, the differences were within the accepted variance of the instrument (+/- 5%). Across all tested ratios, the measured end-potassium concentrations were all within 1 mEq of the expected values, except for the 16:1 ratio at 50 ml/min, which had a 2.52 mEq variance. All significant differences were within the accepted variance of the instrument (+/- 5%). In conclusion, the BCC accurately and delivered cardioplegia volumes and potassium concentrations across all tested conditions with reproducible performance. PMID- 10539712 TI - Post cardiopulmonary bypass bleeding: an introductory review. AB - Cardiac surgery requiring cardiopulmonary bypass (CPB) has increased in both number and safety over the past four decades. Postoperative bleeding continues to be a major cause of perioperative morbidity. The reported incidence varies from 4 32%. This paper reviews the preoperative evaluation and specific hemostasis defects associated with cardiac surgery and extracorporeal circulation. Monitoring of anticoagulation and coagulation, methods to decrease the alterations of the coagulation system, as well as the specific therapy for and risks of post-CPB bleeding are also discussed. PMID- 10539713 TI - Systemic inflammatory response syndrome (SIRS) following emergency cardiopulmonary bypass: a case report and literature review. AB - A complication of emergency resuscitation is the development of the Systemic Inflammatory Response Syndrome (SIRS). In the past, this has been identified as multiple organ failure, with symptoms similar to sepsis. The hallmark of this syndrome is peripheral vasodilation, which is associated with a breakdown of capillary membranes and the accumulation of excess interstitial fluid. This case report discusses the development of SIRS in a patient following emergency cardiopulmonary bypass (CPB). The patient was a 53 year old male with significant left main coronary artery disease who developed sudden bradycardia and hypotension in the operating room and was emergently placed on cardiopulmonary bypass. During CPB, the patient was peripherally vasodilated, and required continuous alpha-adrenergic support to maintain normal systemic vascular resistance. In addition, metabolic acidosis was present despite high flow rates, high hematocrit, addition of colloids, and hemoconcentration. Despite excellent neurological and myocardial recovery following surgery, the patient died one week later in renal and hepatic failure. Several mechanisms for the development of this syndrome have been hypothesized. One of these theories is that the ischemic injury in the gastrointestinal tract disturbs the gut barrier function and allows enteric bacterial endotoxins to pass into the circulation producing sepsis-like symptoms. Other theories relate to the release patterns of cytokines associated with CPB. These mechanisms and the treatment of SIRS with new pharmacological agents and perfusion techniques are reviewed. PMID- 10539714 TI - Redo cardiac surgery in a patient with severe peripheral vascular disease and pericardial adhesions using subclavian arterial cannulation and port-access technology. AB - Patients viewed as conventionally inoperative candidates are now given alternative surgical choices. The ability to provide new technology such as the port-access minimally invasive approach, kinetic venous assist, and specialized cannulae have made this possible. This case report discusses the ability to apply and modify this new technology to provide a successful surgical outcome in a patient with severe peripheral vascular disease and dense mediastinal adhesions. PMID- 10539715 TI - Veno-arterial modified ultrafiltration in children after cardiopulmonary bypass. AB - A method of performing veno-arterial modified ultrafiltration is described that utilizes conventional blood flow through the aortic and venous cannulae. A dual pump blood cardioplegia console is adapted to aspirate blood from the cardiopulmonary bypass venous line. The blood is ultrafiltered, sent through the cardioplegia heat exchanger, and returned to the aorta via the cardioplegia needle. Veno-arterial modified ultrafiltration has produced no visual evidence of air entrainment in the cardiopulmonary arterial line. This method allows the immediate resumption of cardiopulmonary bypass without the need for the surgeon to recannulate or alter tubing. Thirty-five children underwent veno-arterial modified ultrafiltration; the results show significant increases in postoperative hematocrit, early extubation, and improved rheology. PMID- 10539717 TI - Will hospital report cards make the grade? AB - Hospital report cards that document patients' medical outcomes are attracting increasing attention for their role in guiding health care decisions by employers, consumers and providers. Significant questions remain, however, regarding the validity and utility of this information. This Issue Brief is based on a seminar held by the Center for Studying Health System Change at which two expert panels discussed whether report cards make the grade. The first panel approached this subject through a Socratic dialogue that focused on the release of a hypothetical community hospital report card. The second panel weighed in on two research presentations related to report cards. The panelists agreed that efforts to collect and report clinical outcomes data are flawed. Even so, release of the data can help improve clinical quality and foster an environment in which health care quality information ultimately has an impact on health care decision making. PMID- 10539716 TI - Reduction of vasoreactivity and thrombogenicity with laser-thermal angioplasty: comparison with balloon angioplasty. AB - The vasoreactivity and thrombogenicity of laser-thermal angioplasty were examined and compared with those of balloon angioplasty in an atherosclerotic rabbit iliac artery. Eight rabbits underwent laser-thermal angioplasty with a 1.7-mm hot-tip probe activated at 7 W with a probe temperature of 126 +/- 19 degrees C in one iliac artery. The other iliac artery was treated with balloon angioplasty using a 2.0-mm balloon. Angiographic luminal diameter increased from 0.19 +/- 0.15 to 1.54 +/- 0.35 mm by laser and from 0.29 +/- 0.22 to 1.84 +/- 0.20 mm by balloon (P less than 0.0001, respectively). However, it decreased to 1.34 +/- 0.42 for laser and 0.45 +/- 0.39 for balloon 60 minutes later (P less than 0.0001 vs immediately post). Both iliac arteries were visualized using angioscopy, which revealed thrombotic obstruction of 91% stenosis in the ballooned artery and 8% stenosis in the lased artery. Vasoreactivity of treated vessels was also investigated. Segments 3-mm long were obtained from either treated artery or control artery and examined for noradrenaline (10 -7 M) contraction. The segments were then mounted isometrically with 1 g tension in Krebs-bicarbonate buffer. Developed tension was 0.13 +/- 0.21 g for laser thermal and 2.33 +/- 0.4 g for its control (P less than 0.0001), and 0.15 +/- 0.16 g for balloon dilatation and 2.12 +/- 0.43 g for its control (P less than 0.0001). Neither acetylcholine at 10 -6 M or papaverine at 10 -4 M induced relaxation of treated segments. Histology showed slight thermal injury at thermally-treated sites without thrombus, and intimal and medial dissection with thrombus formation at balloon dilated site. IN CONCLUSION: (1) neither a laser-thermal recanalized or a balloon dilated obstructed artery is vasoreactive to constrictive or relaxant agents; and (2) laser-thermal angioplasty results in less thrombogenicity than balloon angioplasty under moderate probe temperature. PMID- 10539718 TI - Health care costs: will they start rising rapidly again? AB - Recent news reports have speculated that health care costs, which have been increasing at dramatically lower rates over the past few years, are about to take off again. But a panel convened by the Center for Studying Health System Change to discuss health care cost trends said the forces that have kept large cost increases at bay are still in effect and premium increases will be moderate. This Issue Brief reports on what the panel participants predicted is likely to happen to health care costs during the next several years and what underlying forces will shape these trends. PMID- 10539719 TI - Patients, profits and health system change: a Wall Street perspective. AB - Two years ago, the Center for Studying Health System Change brought together a group of highly respected Wall Street securities analysts who track health care companies. Their purpose: to discuss significant developments in the health care industry and possible future trends. The analysts met again this spring to discuss how the industry has changed since then. This Issue Brief reports on that roundtable discussion, during which the analysts addressed four major topics: managed care, health plans and providers, consolidation and public policy. PMID- 10539720 TI - Access to health care: bridging the gap between policy and research. AB - Access to care has long been studied by researchers, but emphasis has shifted recently from measuring whether access is worse for the poor and uninsured to monitoring how access is changing over time. Only a limited number of measures have been used consistently to monitor changes longitudinally, and these suggest that access by the uninsured is declining. There are potentially more valuable measures to track access, but data that are consistent over time have not been available. This Issue Brief, which is based on a seminar held by the Center for Studying Health System Change, discusses current research efforts to measure access to care. PMID- 10539721 TI - An inadequate supply of qualified people will slow the pace of health system change. AB - Executive recruiters bring a unique perspective on changes in the health system. By knowing the hiring needs of health care organizations, recruiters understand how the organizations are reacting to changes that have already occurred and how their strategies for dealing with the changes set the direction for future trends in the health system. This Issue Brief reports on a roundtable discussion during which a group of executive recruiters considered how health system changes are reflected in recruitment efforts in health care, and what the shifts in recruitment priorities indicate for the future direction and pace of health system change. The roundtable was sponsored by the Center for Studying Health System Change. PMID- 10539722 TI - Tracking health care costs: a slowing down of the rate of increase. AB - Throughout the 1990s there has been a dramatic slowing of health care cost increases. Virtually all studies show a substantial decline even after adjusting for general inflation, which dropped from 4.3 percent in 1990 to 2.5 percent in 1995. The pressures to contain health care costs are so intense that the low rates of increase could continue for some time. Consumers may not perceive this change, however, perhaps because of cost-shifting to them. This Issue Brief tracks changes in health care costs over time. The Center for Studying Health System Change plans to track these changes periodically. PMID- 10539723 TI - A primer on understanding health care cost trends: the story behind the numbers. AB - Virtually every study shows that rates of increase in health care costs have declined throughout the 1990s, but often the bottom-line numbers differ. There are numerous reasons for this disparity, including which data are used and how researchers analyze them. Unfortunately, there is no ideal source of data; the best sources vary widely with regard to accuracy, timeliness, and, most important, whether the information lines up with the questions that are being asked. This Issue Brief provides information about how to understand what the numbers really represent. PMID- 10539724 TI - Policy implications of risk selection in Medicare HMOs: is the federal payment rate too high? AB - For more than a decade, Medicare beneficiaries have had the option to enroll in risk-contract health maintenance organizations (HMOs) in which the federal payment is set at 95 percent of the estimated fee-for-service cost. Two questions have been raised by health policy researchers ever since: Are Medicare HMO enrollees healthier than the elderly who receive fee-for-service care? If so, does the government payment rate for Medicare HMOs accurately reflect the costs that would have been incurred by a healthier population? This Issue Brief discusses three recent studies of the extent to which risk-contract HMOs experience biased selection and the cost of this to Medicare. PMID- 10539726 TI - Tracking changes in the public health system: what researchers need to know to monitor and evaluate these changes. AB - Just as the medical care system is changing in communities across the country, so too is the public health system. Reduced resources, fragmentation of traditional public health functions, the spread of managed care, and developing new partnerships are key among these changes. Two dozen public health officials and health policy researchers met in April at the Center for Studying Health System Change to discuss the changes in the financing and delivery of public health services and the research needed to monitor and evaluate the impact of these changes. PMID- 10539725 TI - Medicaid eligibility policy and the crowding-out effect: did women and children drop private health insurance to enroll in Medicaid. AB - In the late 1980s and early 1990s, the federal government expanded Medicaid eligibility for children and pregnant women. By 1992, nearly a third of all children in the United States were eligible for Medicaid, and between 40 and 50 percent of women of childbearing age were eligible for Medicaid coverage for pregnancy-related services. During this period, the number of persons with employment-based insurance coverage declined, leading researchers to investigate whether Medicaid expansions have contributed to this decline--a so-called crowding-out effect. This Issue Brief discusses research findings and the health policy implications of the crowding-out effect. PMID- 10539727 TI - Managed care woes: industry trends and conflicts. AB - Although managed care plans have had success in controlling costs, they now face challenges on many fronts, including tighter profit margins, pressure for broader provider networks, increasing clout of hospitals and physicians and more demand for consumer protection regulation. Underlying these trends is a fundamental conflict between health plans and consumers, who are demanding--and, in many cases, getting--greater control over their health care delivery and services. This Issue Brief reports on a roundtable convened by the Center for Studying Health System Change to discuss these trends and conflicts and how they may play out over the next number of years. (1) Managed care challenges, (2) Redefining care delivery, (3) Investing in information systems, (4) Center survey findings about managed care, (5) Who's in control, (6) HCFA as a value purchaser, (7) Backlash fueled by many groups, (8) What about the future. PMID- 10539728 TI - Next steps in incremental health insurance expansions: who is most deserving? AB - President Clinton's proposal to allow near-elderly individuals--those between the ages of 55 and 64--to buy-in to Medicare is the latest initiative to expand health insurance coverage incrementally by targeting particular age groups. Health insurance expansions for children were passed last year, and assistance to young adults--those age 19 to 24--has also been discussed. The rationale for targeting individuals in these age groups is that they are presumed to be vulnerable. However, vulnerability to uninsurance includes not only the risk of being uninsured, but also the potential health and financial consequences that result from not having coverage, each of which varies substantially across age groups. This Issue Brief examines the vulnerability of the near-elderly and young adults with respect to uninsurance. PMID- 10539729 TI - CHIPing away at the problem of uninsured children. AB - The State Children's Health Insurance Program (CHIP), enacted one year ago this August, is the largest expansion of health insurance in more than three decades. One of the measures of its success will be whether state officials are able to enroll children who are eligible. Research conducted by Health System Change (HSC) shows that uninsured children are a diverse group, and that for CHIP to be successful, policy makers will need to target programs to specific groups and local market conditions. This Issue Brief discusses why children lack health insurance and the implications for implementing CHIP. PMID- 10539730 TI - The uninsured getting care: where you live matters. AB - A substantial number of Americans--41 million people--do not have health insurance; this represents a 16 percent increase in the uninsured since 1990. Further, many studies show that the uninsured have significantly more difficulty than the insured in getting needed care. This Issue Brief discusses Health System Change (HSC) findings from its Community Tracking Study showing that the ability of those without coverage to get needed care varies considerably across communities. In addition, HSC's study shows that the personal characteristics of the uninsured explain very little of this regional variation. These findings are the first step in helping decision makers understand how the dynamics of communities and the safety nets within them affect the medically indigents' ability to obtain needed care. PMID- 10539731 TI - Public health departments adapt to Medicaid managed care. AB - Millions of Medicaid beneficiaries have recently moved into private managed care plans across the country. Public health departments--which have acted as providers of primary care and other services for some Medicaid patients--are often not explicitly included in contracts between these designated Medicaid plans and the states. As a result, many of the 3,000 city and county public health agencies nationwide have lost both patients and significant revenue to plans. This Issue Brief describes how public health departments are adapting to this shift in state policy. According to our research conducted in 1997, many are de-emphasizing the delivery of direct health care services in favor of core public health functions, such as investigating community health problems and health promotion. Some are initiating new partnerships with Medicaid managed care plans. PMID- 10539732 TI - Wall Street comes to Washington: analysts' perspectives on health system change. AB - Security analysts took a hard look at the latest trends in the health care industry at the third annual Health System Change Wall Street roundtable held earlier this year. The roundtable focused on costs and premiums, evolving financial relationships among industry leaders, industry consolidation and pharmaceutical and technology development. The analysts' conclusions: some market strategies--including vertical integration, provider risk-sharing and equity arrangements and the use of formularies to rein in drug costs--haven't worked out as expected. Health plans and provider organizations have yet to find a magic bullet for controlling costs while responding to purchasers' and consumers' demands for broader choice. This Issue Brief reports on the trends discussed at the roundtable. PMID- 10539733 TI - The community tracking study: a focus on change in the health care system. AB - Recognizing that health care delivery is predominantly local, the Center for Studying Health System Change is investigating what is happening in health care financing and delivery at the community level. The Community Tracking Study focuses on changes in the health care system in 60 sites that are representative of the nation. Twelve of these communities are being studied intensively. The 48 additional communities studied less intensively will permit generalization to the nation as a whole and analysis of the relationship between health system characteristics and the effects of change on people. Data collection and analysis for the Community Tracking Study are planned in two-year cycles. The first cycle, which began in spring of 1996, will establish a baseline. PMID- 10539734 TI - The changing face of AIDS: translators needed. PMID- 10539735 TI - Metabolic markers of vitamin nutritional status. PMID- 10539736 TI - Ethanol and lipid metabolism. PMID- 10539737 TI - High doses of vitamin E in the treatment of disorders of the central nervous system in the aged. AB - Oxidative stress is a putative factor in the pathogenesis of many human disorders of the central nervous system. Therefore, antioxidants such as vitamin E have become attractive as therapeutic agents in the treatment of several diseases. In addition, vitamin E seems to play a specific role in the nervous system. As a result, vitamin E has been used in pharmacologic doses in the treatment of disorders such as Parkinson disease, Alzheimer disease, and tardive dyskinesia. One investigation showed that the use of 2000 IU all-rac-alpha-tocopheryl acetate is beneficial in the treatment of Alzheimer disease. Similar doses of vitamin E, however, were not beneficial for delaying the progression of Parkinson disease. In other studies, dosages >/=400 IU vitamin E/d were found to be beneficial in the treatment of tardive dyskinesia, although this finding was not confirmed in a larger cooperative study conducted by the Veterans Administration. Even though the efficacy of vitamin E in the management of cardiovascular disease has been shown, the potential role of vitamin E in the treatment of cerebrovascular disease remains essentially unknown. The experience from 2 large clinical trials involving the oral intake of 2000 IU vitamin E/d suggests that vitamin E is relatively safe at this dosage for periods <2 y. However, the safety and efficacy of supplemental vitamin E over periods of many years in the prevention of neurologic diseases has not been adequately explored. PMID- 10539738 TI - Do we facilitate the scientific process and the development of dietary guidance when findings from single studies are publicized? An American Society for Nutritional Sciences controversy session report. AB - This American Society for Nutritional Sciences Controversy Session presented at the 1997 Experimental Biology meeting considered whether publicity of findings from single studies facilitates or hampers the scientific process and the development of scientifically sound dietary guidance. In a 1995 survey, 78% of primary household shoppers believed it "very likely" or "somewhat likely" that in the next 5 y experts would have a completely different idea about which foods were healthy and which were not. This skepticism is fueled by the media's emphasis on reporting new and often controversial findings about food and nutrition. Media efforts are reinforced by the fact that some scientific journals regularly publicize newly published research findings. As a consequence, journalists frequently mediate scientific debate in a public forum-debate that previously was conducted among knowledgeable peers. Tight deadlines often make it difficult for reporters to thoroughly investigate findings publicized in press releases. Headlines can make results from single studies appear important, even when results are inconclusive. Finally, scientists and public policymakers have limited opportunity for making timely comments in response to an issue reported in the media. Nevertheless, the public has a right to be informed about health related research findings to help them make decisions about their diets. The media are a valuable resource for educating the public and maintaining public interest in the importance of diet in overall health status. Nutrition scientists should be more involved in helping the media accurately convey diet and health messages. PMID- 10539739 TI - Regions of the human brain affected during a liquid-meal taste perception in the fasting state: a positron emission tomography study. AB - BACKGROUND: The sensation of taste provides reinforcement for eating and is of possible relevance to the clinical problem of obesity. OBJECTIVE: Positron emission tomography (PET) was used to explore regions of the brain that were preferentially affected during the taste perception of a liquid meal by 11 right handed, lean men in the fasting state. DESIGN: After subjects had fasted for 36 h, 2 measurements of regional cerebral blood flow (rCBF) obtained immediately after subjects retained and swallowed 2 mL of a flavored liquid meal (the taste condition) were compared with 2 measurements of rCBF obtained immediately after subjects retained and swallowed 2 mL of water (the baseline condition). RESULTS: Compared with the baseline condition, taste was associated with increased rCBF (P < 0.005) in the left dorsolateral prefrontal cortex and superior temporal gyrus; the right ventrolateral prefrontal cortex, supramarginal gyrus, and anterior thalamus; and bilaterally in the hippocampal formation, posterior cingulate, midbrain, occipital cortex, and cerebellum. Taste was also associated with decreased rCBF (P < 0.005) in the right dorsolateral prefrontal cortex, superior temporal gyrus, and supplementary motor area, and bilaterally in the medial prefrontal cortex and inferior parietal lobule. CONCLUSIONS: This exploratory study provides additional evidence that the temporal cortex, thalamus, cingulate cortex, caudate, and hippocampal formation are preferentially affected by taste stimulation. The asymmetric pattern of activity in the dorsolateral prefrontal cortex and superior temporal gyrus may contribute to the taste perception of a liquid meal perceived as pleasant. Additional studies are required to determine how these regions are affected in patients with obesity or anorexia. PMID- 10539740 TI - Obesity at the age of 50 y in men and women exposed to famine prenatally. AB - BACKGROUND: It was shown that men who were conceived during the Dutch famine of 1944-1945 had higher rates of obesity at age 19 y than those conceived before or after it. OBJECTIVE: Our objective was to study the effects of prenatal exposure to the Dutch famine on obesity in women and men at age 50 y. DESIGN: We measured the body size of 741 people born at term between November 1943 and February 1947 in Amsterdam. We compared people exposed to famine in late, mid, or early gestation (exposed participants) with those born before or conceived after the famine period (nonexposed participants). RESULTS: The body mass index (BMI; in kg/m(2)) of 50-y-old women exposed to famine in early gestation was significantly higher by 7. 4% (95% CI: 0.7%, 14.5%) than that of nonexposed women. BMI did not differ significantly in women exposed in mid gestation (-2.1%; -7.0%, 3.1%) or in late gestation (-1.3%; -6.3%, 3.9%). In 50-y-old men, BMI was not significantly affected by exposure to famine during any stage of gestation: BMI differed by 0.4% (-3.5%, 4.5%) in men exposed to famine in late gestation, by -1.2% (-5.5%, 3.3%) in those exposed in mid gestation, and by 0.5% (-4.6%, 6.0%) in those exposed in early gestation compared with nonexposed men. CONCLUSIONS: Maternal malnutrition during early gestation was associated with higher BMI and waist circumference in 50-y-old women but not in men. These findings suggest that pertubations of central endocrine regulatory systems established in early gestation may contribute to the development of abdominal obesity in later life. PMID- 10539741 TI - Dietary fish as a major component of a weight-loss diet: effect on serum lipids, glucose, and insulin metabolism in overweight hypertensive subjects. AB - BACKGROUND: Obesity in hypertensive patients is associated with dyslipidemia and insulin resistance, both of which are improved by weight control. n-3 Fatty acids have diverse effects on mechanisms underlying atherosclerosis, including a decrease in serum triacylglycerols and an increase in HDL(2) cholesterol. OBJECTIVE: The objective was to examine whether dietary fish enhances the effects of weight loss on serum lipids, glucose, and insulin in 69 overweight, treated hypertensive patients. DESIGN: Overweight patients being treated for hypertension were randomly assigned to either a daily fish meal (3.65 g n-3 fatty acids), a weight-loss regimen, the 2 regimens combined, or a control group for 16 wk. RESULTS: Sixty-three subjects completed the study. Weight decreased by a mean (+/ SEM) of 5.6 +/- 0.8 kg with energy restriction. Weight loss decreased fasting insulin (P = 0.003) and the area under the curve for insulin (P = 0.003) and glucose (P = 0.047) during an oral-glucose-tolerance test. The greatest decrease occurred in the fish + weight-loss group. There was no independent effect of fish on glucose or insulin. Fish increased HDL(2) cholesterol (P = 0.004) and decreased HDL(3) cholesterol (P = 0.026) without altering total, LDL, or HDL cholesterol. Weight loss had no effect on these variables. Fasting triacylglycerols fell significantly with fish consumption (29%) and weight loss (26%). The fish + weight-loss group showed the greatest improvement in lipids: triacylglycerols decreased by 38% (P < 0.001) and HDL(2) cholesterol increased by 24% (P = 0.04) compared with the control group. CONCLUSIONS: Incorporating a daily fish meal into a weight-loss regimen was more effective than either measure alone at improving glucose-insulin metabolism and dyslipidemia. Cardiovascular risk is likely to be substantially reduced in overweight hypertensive patients with a weight-loss program incorporating fish meals rich in n-3 fatty acids. PMID- 10539742 TI - Sitostanol administered in lecithin micelles potently reduces cholesterol absorption in humans. AB - BACKGROUND: Phytosterol feeding in human clinical trials has had generally small and inconsistent effects on serum cholesterol concentrations, raising doubts about the importance of phytosterols in natural diets and supplements. OBJECTIVE: The hypothesis tested was that the low intestinal bioavailability of purified phytosterols can be increased by formulation with lecithin. DESIGN: The ability of sitostanol to reduce cholesterol absorption was measured directly by including hexadeuterated cholesterol tracer in a standard test breakfast and measuring plasma tracer concentration 4 and 5 d later by gas chromatography-negative ion mass spectrometry. The tracer amount after a test meal containing sitostanol was compared with that after an identical meal containing placebo. Each subject served as his or her own control and the order of testing was random. Sitostanol was formulated either as a powder or as a sonicated micellar solution with lecithin. A total of 38 single-meal tests were performed in 6 healthy subjects. RESULTS: Sitostanol powder (1 g) reduced cholesterol absorption by only 11.3 +/- 7.4% (P = 0.2), confirming in vitro data showing poor solubility of sitostanol powder in artificial bile. In contrast, sitostanol in lecithin micelles reduced cholesterol absorption by 36.7 +/- 4.2% (P = 0.003) at a dose of 700 mg and by 34.4 +/- 5.8% (P = 0.01) at a dose of 300 mg. CONCLUSIONS: Sitostanol reduced cholesterol absorption at doses lower than reported previously, but only if presented in lecithin micelles. Properly formulated sitostanol as well as naturally occurring complexes of phytosterol and phospholipid might be therapeutically useful for cholesterol lowering. PMID- 10539743 TI - Effect of trans fatty acids on calcium influx into human arterial endothelial cells. AB - BACKGROUND: A recent task force of The American Society for Clinical Nutrition and American Society for Nutritional Sciences recommended in a position paper on trans fatty acids that models be developed to assess the effects of changes in fat intake on disease risk. OBJECTIVE: The objective was to investigate, using human arterial endothelial cells as a model, the influence of trans fatty acids and magnesium on cell membrane composition and on calcium influx into arterial cells, a hallmark of atherosclerosis. DESIGN: Endothelial cells were cultured for 3 d in media with high (adequate) or low (inadequate) amounts of magnesium plus various concentrations of trans,trans linoelaidic; cis,cis linoleic; trans elaidic; oleic; or stearic acids. The cells were then harvested and the fatty acid composition and the amount of (45)Ca(2+) incorporated into the cell was determined. RESULTS: The percentage of fatty acids incorporated into the endothelial cells was proportional to the amount added to the culture medium. Adequate magnesium was crucial in preventing calcium influx into endothelial cells. Without an adequate amount of magnesium in the culture medium, linoelaidic and elaidic acids, even at low concentrations, increased the incorporation of (45)Ca(2+) into the cells, whereas stearic acid and oleic acid did not (P < 0.05). CONCLUSION: Our model indicated that a diet inadequate in magnesium combined with trans fat may increase the risk of calcification of endothelial cells. PMID- 10539744 TI - Comparative lipid and lipoprotein responses to solid-food diets and defined liquid-formula diets. AB - BACKGROUND: Liquid-formula diets (LFDs) are useful in metabolic studies of the cholesterolemic effects of dietary lipids because they can be formulated with accuracy, facilitating precise delivery of fatty acids of interest. However, because of differences in composition and nutrient delivery between LFDs and solid-food diets (SFDs), there is a need to determine differences in their effects. OBJECTIVE: Our objective was to compare lipid and lipoprotein responses to changes in total fat, saturated fatty acids (SFAs), and cholesterol in subjects consuming an SFD or LFD. DESIGN: Twenty-one healthy subjects consumed controlled diets representative of an average American diet [AAD; 37% of energy from fat (15% from SFAs), and <50 mg cholesterol/MJ] or a National Cholesterol Education Program (NCEP) Step II diet [26% fat (5% from SFAs) and <25 mg cholesterol/MJ]. Other nutrients were similar between diets. Diets were consumed for 23 d in a randomized, crossover design. RESULTS: For the AAD and NCEP Step II diet, there were no significant differences in lipids and apolipoproteins when the LFD or SFD versions were consumed. In contrast, consumption of the SFD was associated with significantly lower total cholesterol and triacylglycerols than was consumption of the corresponding AAD or Step II LFD (P < 0.05). Subjective ratings of satiety, hunger, and quality of life between diet forms did not differ significantly. CONCLUSIONS: Both LFDs and SFDs yield quantitatively similar cholesterolemic responses to changes in dietary fat, SFAs, and cholesterol. LFDs may offer advantages because they provide easily administered, complete, balanced nutrition without affecting satiety. PMID- 10539745 TI - Measurement of body water by multifrequency bioelectrical impedance spectroscopy in a multiethnic pediatric population. AB - BACKGROUND: Bioelectrical impedance spectroscopy (BIS) may provide a noninvasive, rapid method for the assessment of total body water (TBW), extracellular water (ECW), and intracellular water (ICW). Few studies, however, have examined the accuracy of BIS in pediatric populations. OBJECTIVE: Our objective was to evaluate the accuracy of BIS for the measurement of TBW, ECW, and ICW in healthy children. DESIGN: Dual-energy X-ray absorptiometry (DXA), total body potassium (TBK), and BIS measurements were performed in 347 children (202 males and 145 females aged 4-18 y). The reference values for TBW, ECW, and ICW were defined by using a DXA+TBK model. BIS values were evaluated by using the method of Bland and Altman. A randomly selected calibration group (n = 231) was used to derive new BIS constants that were tested in the remaining group (n = 116). RESULTS: BIS values were highly correlated with the reference values (r(2) = 0.94-0.97, P < 0.0001), but differences between the BIS and DXA+TBK models for individuals were significant (P < 0.001). Use of new BIS constants reduced the mean differences between the BIS and DXA+TBK models; the SDs of the mean differences were improved (1.8 L for TBW, 1.4 L for ICW, and 1.0 L for ECW) for the total population. CONCLUSIONS: On a population basis, BIS can be calibrated to replace the DXA+TBK model for the assessment of TBW, ECW, and ICW in healthy children. The accuracy of the BIS measurement in individual children may be refined further by using age and sex-specific adjustments for the BIS calibration constants. PMID- 10539746 TI - Metabolic and behavioral consequences of a snack consumed in a satiety state. AB - BACKGROUND: In view of the influence of dietary habits on obesity, human eating patterns merit study. OBJECTIVE: We investigated the behavioral and biological consequences of consumption of a 1-MJ snack by subjects in a satiety state. DESIGN: Eleven lean young men were deprived of time cues and subjected to continuous blood withdrawal over each of 4 sessions scheduled 2 wk apart. The first session was a basal session designed to determine the following in each subject: 1) the amount eaten in an ad libitum lunch; 2) the temporal patterns of plasma concentrations of glucose, insulin, fatty acids, and triacylglycerols between lunch and the spontaneous dinner request; and 3) the latency of the dinner request. In the 3 other sessions, each subject ingested the same lunch as in the basal session and a nutritionally well-balanced snack either 5 min before his individual peak of hyperglycemia observed in the first session, 40 min after this peak, or 120 min before the time he had requested his dinner in the first session. RESULTS: There was no significant difference in latency of the dinner request or the energy intake at dinner between sessions. Insulin secretion increased but glucose profiles did not change significantly regardless of the time of snack intake. CONCLUSION: A snack consumed in a satiety state fails to prolong the intermeal interval and would thus tend to favor storage. PMID- 10539747 TI - Bioelectrical impedance analysis in HIV-infected patients treated with triple antiretroviral treatment. AB - BACKGROUND: Triple antiretroviral treatment including protease inhibitors (PIs) delays the clinical progression of HIV infection and may thus reduce the risk of malnutrition. However, fat redistribution (lipodystrophy) was recognized recently as a metabolic side effect of PIs. OBJECTIVE: The study aimed to assess the effect of triple antiretroviral treatment on body composition and on the prevalence of malnutrition. DESIGN: Two cross-sectional studies, 1 in 1996 (t96; n = 247) and 1 in 1997 (t97; n = 266), were conducted in HIV-infected outpatients. Among patients who participated in both studies, 111 patients started a new antiretroviral treatment including a PI between t96 and t97 and were studied longitudinally. Total body water (TBW), intracellular water (ICW), extracellular water (ECW), and fat mass were estimated by monofrequency bioelectrical impedance analysis (BIA). RESULTS: Prevalence of malnutrition was reduced by 30-50% from t96 to t97, depending on the definition used. In the longitudinal study, TBW and the ratio between ICW and ECW increased and fat mass decreased (P < 0.001). BIA indicated a greater increase in ICW in 23 (21%) patients with clinically apparent fat redistribution than in patients without this syndrome, but estimates of fat mass changes were not significantly different. CONCLUSIONS: Triple antiretroviral treatment may protect HIV-infected patients against the development of malnutrition. Whole-body BIA data suggest an increase in appendicular body cell mass associated with improved antiretroviral treatment. However, the method is unreliable in detecting fat redistribution, and current prediction equations will need to be recalibrated for HIV-infected patients receiving highly active antiretroviral treatment. PMID- 10539748 TI - Use of the deuterated-retinol-dilution technique to assess total-body vitamin A stores of adult volunteers consuming different amounts of vitamin A. AB - BACKGROUND: The deuterated-retinol-dilution (DRD) technique provides a quantitative estimate of total body stores of vitamin A. However, it is not known whether the technique can detect changes in vitamin A pool size in response to different intakes of vitamin A. OBJECTIVE: Our objective was to determine the responsiveness of the DRD technique to 3 different daily supplemental vitamin A intakes during a period of 2.5-4 mo. DESIGN: Two oral doses of [(2)H(4)]retinyl acetate [52.4 micromol retinol equivalent (RE)] were administered on study days 1 and 91 to 26 men (18-32 y of age) who were consuming controlled, low-vitamin A diets, and receiving daily either 0, 5.2, or 10.5 micromol RE of unlabeled supplemental retinyl palmitate during a 75- or 129-d period. Plasma isotopic ratios of [(2)H(4)]retinol to retinol on day 115 were used to estimate final vitamin A body stores per Furr et al (Am J Clin Nutr 1989;49:713-6). RESULTS: Final ( +/- SD) estimated vitamin A pool sizes were 0.048 +/- 0.031, 0.252 +/- 0.045, and 0.489 +/- 0.066 mmol in the treatment groups receiving 0, 5.2, and 10.5 micromol RE/d, respectively (P < 0.001). Estimated mean changes in vitamin A pool sizes were similar to those expected for the vitamin A-supplemented groups [estimated:expected (95% CI of change in pool size): 1.08 (0.8, 1.2) and 1.17 (1.0, 1.3)]. CONCLUSIONS: The DRD technique can detect changes in total body stores of vitamin A in response to different daily vitamin A supplements. However, abrupt changes in dietary vitamin A intake can affect estimates of total body vitamin A stores. PMID- 10539749 TI - Increased dietary micronutrients decrease serum homocysteine concentrations in patients at high risk of cardiovascular disease. AB - BACKGROUND: Elevated blood homocysteine is a risk factor for cardiovascular disease. A 5-micromol/L increase is associated with an approximately 70% increase in relative risk of cardiovascular disease in adults. For patients with established risk factors, this risk is likely even greater. OBJECTIVE: Effects of increased dietary folate and recommended intakes of vitamins B-12 and B-6 on serum total homocysteine (tHcy) were assessed in individuals at high risk of cardiovascular disease. DESIGN: This trial was conducted at 10 medical research centers in the United States and Canada and included 491 adults with hypertension, dyslipidemia, type 2 diabetes, or a combination thereof. Participants were randomly assigned to follow a prepared meal plan (PMP; n = 244) or a self-selected diet (SSD; n = 247) for 10 wk, which were matched for macronutrient content. The PMP was fortified to provide >/=100% of the recommended dietary allowances for 23 micronutrients, including folate. RESULTS: Mean folate intakes at 10 wk were 601 +/- 143 microgram/d with the PMP and 270 +/ 107 microgram/d with the SSD. With the PMP, serum tHcy concentrations fell from 10.8 +/- 5.8 to 9.3 +/- 4.9 micromol/L (P < 0.0001) between weeks 0 and 10 and the change was associated with increased intakes of folate, vitamin B-12, and vitamin B-6 and with increased serum and red blood cell folate and serum vitamin B-12 concentrations. tHcy concentrations did not change significantly with the SSD. CONCLUSIONS: The PMP resulted in increased intakes and serum concentrations of folate and vitamin B-12. These changes were associated with reduced serum tHcy concentrations in persons at high risk of cardiovascular disease. PMID- 10539750 TI - Urinary iodine concentrations and thyroid function in adult Zimbabweans during a period of transition in iodine status. AB - BACKGROUND: In 1993 the compulsory iodization of salt was introduced in Zimbabwe, a country that was previously an area of severe iodine deficiency. OBJECTIVE: The objective of this study was to document urinary iodine excretion and biochemical thyroid function in seemingly healthy, community-dwelling adults after the introduction of iodization. DESIGN: A multistage, random sampling method was used in rural and urban settings to identify households from which the senior household member (aged >35 y) was recruited (alternating male and female recruits). Demographic data were collected for each subject and urinary and venous blood samples were taken. Urinary iodine excretion and serum thyroid hormone status (thyrotropin and total thyroxin) were evaluated according to age, sex, and area of residence. RESULTS: A total of 736 adults were recruited (253 men; mean age: 64 y). Urinary iodine concentrations were high [median (first and third quartiles): 4.41 (2.84, 6.78) micromol/L, or 560 (360, 860) microgram/L] and were significantly higher in rural areas than in urban areas [4.73 (3.07, 7.14) micromol/L, or 600 (390, 906) microgram/L, compared with 3.47 (2.05, 4.73) micromol/L, or 440 (260, 600) microgram/L; P < 0.001]. Urinary iodine excretion declined significantly with increasing age (r = -0.29, P < 0.001). Serum thyroid status suggested that the prevalence of biochemical hyperthyroidism in the study was 3%, with 13 of 415 cases in rural and 3 of 149 cases in urban subjects. CONCLUSION: This study reaffirms the need to continuously monitor iodine replacement programs to ensure efficacy. PMID- 10539751 TI - Decreased serum ubiquinone-10 concentrations in phenylketonuria. AB - BACKGROUND: Ubiquinone-10 is a lipid with important metabolic functions that may be decreased in phenylketonuria (PKU) because patients with PKU consume diets restricted in natural proteins. OBJECTIVE: We studied serum ubiquinone-10 concentrations in PKU patients. DESIGN: This was a retrospective, transversal study in which we compared serum ubiquinone-10, plasma cholesterol, plasma tyrosine, and plasma phenylalanine concentrations in 43 PKU patients with concentrations in a reference population (n = 102). Serum ubiquinone-10 concentrations were analyzed by HPLC with ultraviolet detection. Plasma tyrosine and phenylalanine were measured by ion-exchange chromatography. RESULTS: Serum ubiquinone-10 concentrations in PKU patients were significantly lower than in the reference population (P < 0.01 for patients aged 1 mo to <8 y and P < 0.00005 for patients aged 8-33 y). Moreover, 5 of 18 PKU patients (28%) in the younger age group and 10 of 23 (43%) in the older age group had serum ubiquinone-10 concentrations below the reference interval. CONCLUSIONS: Serum ubiquinone-10 deficiency appears to be related to the restricted diet of PKU patients. Because serum ubiquinone-10 plays a major antioxidant role in the protection of circulating lipoproteins, the correction of ubiquinone-10 concentrations should be considered in PKU patients. Further investigation seems advisable to elucidate whether the deficiency in serum ubiquinone-10 status is clinically significant. PMID- 10539752 TI - An estimation of selenium requirements for New Zealanders. AB - BACKGROUND: Current US dietary recommendations for selenium are based on maximization of plasma glutathione peroxidase (GSHPx) activity according to data from one study of Chinese men. OBJECTIVE: The effect of various amounts of supplemental selenium on GSHPx activities in blood of New Zealand adults was investigated to calculate a selenium requirement for New Zealanders. The effect on plasma selenoprotein P and thyroid hormones was also investigated. DESIGN: Fifty-two adults with low blood selenium concentrations ingested a placebo or 10, 20, 30, or 40 microgram Se as L-selenomethionine daily for 20 wk. RESULTS: Plasma and whole-blood GSHPx activities increased in all supplemented groups but reached a plateau only in the group receiving 40 microgram Se, as determined by statistical analysis. Increases in selenoprotein P were greater than those for selenium and GSHPx at all supplement intakes. Thyroxine concentrations decreased in supplemented groups but the decrease was significantly different from that in the control group only for the 10-microgram group and for all supplemented groups combined. CONCLUSIONS: An upper estimated requirement of 90 microgram Se/d was calculated as the intake necessary for maximization of plasma GSHPx activity, as used in the derivation of the US recommended daily allowance. Our lower estimated requirement of 39 microgram Se/d was the intake necessary to reach two-thirds of maximal GSHPx activity, as was used in calculating the World Health Organization normative requirement. The lower estimate is a realistic goal for New Zealand but the upper estimate could be achieved only with regular inclusion of high-selenium foods. PMID- 10539753 TI - Serum cobalamin, homocysteine, and methylmalonic acid concentrations in a multiethnic elderly population: ethnic and sex differences in cobalamin and metabolite abnormalities. AB - BACKGROUND: Low cobalamin concentrations and mild hyperhomocysteinemia are common in the elderly but ethnic differences have not been defined. OBJECTIVE: Our objective was to determine the demographic characteristics of cobalamin deficiency in the elderly and its role in their hyperhomocysteinemia. DESIGN: We measured serum cobalamin, total homocysteine (Hcys), and methylmalonic acid (MMA) concentrations in 725 subjects >60 y old, and folate concentrations in 520 subjects. RESULTS: After exclusion of subjects taking cobalamin supplements or with renal insufficiency, high prevalences of low cobalamin (11.8%), high MMA (16.6%), and high Hcys (26.1%) concentrations were seen. Most cobalamin concentrations <140 pmol/L appeared to reflect deficiency because 78. 3% of them were accompanied by abnormal metabolites. Subjects with cobalamin concentrations of 140-258 pmol/L had significantly fewer metabolic abnormalities. A low cobalamin concentration and renal insufficiency were the strongest predictors of abnormal Hcys concentrations. Elderly men had higher Hcys concentrations than did women (P = 0.0001). Whites and Latin Americans had lower cobalamin concentrations than did blacks and Asian Americans (P < 0.005). Whites also had higher Hcys concentrations than all the other groups (P < 0.05). When included in the analysis, renal insufficiency in subjects was associated with 23.8% of all high Hcys and 25.5% of all high MMA concentrations; most with renal insufficiency were Asian American and black men. CONCLUSIONS: Mild cobalamin deficiency is most common in elderly white men and least common in black and Asian American women. Hyperhomocysteinemia, which is most strongly associated with low cobalamin concentrations, is also most common in elderly whites, whereas that associated with renal insufficiency is more common in blacks and Asian Americans. Ethnic differences in cobalamin deficiency and the Hcys patterns associated with it or with renal insufficiency warrant consideration in supplementation strategies. Extending suspicion of deficiency to persons with cobalamin concentrations of 140 258 pmol/L appears to provide more disadvantages than advantages. PMID- 10539754 TI - Racial differences in prevalence of cobalamin and folate deficiencies in disabled elderly women. AB - BACKGROUND: Many previous investigations of cobalamin and folate status were performed in white populations. OBJECTIVE: Our objective was to determine whether there are racial differences in the prevalence of cobalamin and folate deficiency. DESIGN: The study was a cross-sectional comparison of baseline serum cobalamin, folate, methylmalonic acid (MMA), total homocysteine (tHcy), and creatinine concentrations, complete blood count, and vitamin supplementation in 550 white and 212 African American subjects from a cohort of physically disabled older women. RESULTS: The mean (+/-SD) serum MMA concentration was significantly higher in whites than in African Americans: 284 +/- 229 compared with 218 +/- 158 nmol/L (P = 0.0001). tHcy concentration was higher in African Americans than in whites: 12.4 +/- 7.0 compared with 10.9 +/- 4.6 micromol/L (P = 0.001). Serum cobalamin was lower in whites (P = 0.0002). Cobalamin deficiency (serum cobalamin <258 pmol/L and MMA >271 nmol/L) was more frequent in the white women (19% compared with 8%; P < 0.0003). Folate deficiency (serum folate <11.4 nmol/L, tHcy >13.9 micromol/L, and MMA <271 nmol/L) was more prevalent in African Americans than in whites (5% compared with 2%; P = 0.01). Multivitamin use was associated with lower tHcy but not with MMA concentrations. Regression models showed that age >85 y, African American race, serum creatinine >90 micromol/L, and high MMA concentration were all significantly correlated with higher tHcy. Creatinine > 90 micromol/L, white race, and folate concentration were positively associated with MMA concentration. CONCLUSIONS: Cobalamin deficiency with elevated serum MMA concentration is more prevalent in elderly white than in African American women and elevated serum tHcy and folate deficiency are more prevalent in elderly African American than in white women. PMID- 10539755 TI - Double-blind, randomized trial of a synthetic triacylglycerol in formula-fed term infants: effects on stool biochemistry, stool characteristics, and bone mineralization. AB - BACKGROUND: The low sn-2 palmitate content of infant formulas results in formation of fatty acid calcium soaps in the stools and reduced calcium absorption. OBJECTIVE: Our objective was to test the hypotheses that increasing the proportion of sn-2 palmitate in formula for term infants would result in greater skeletal mineral deposition and reduced stool hardness. DESIGN: Healthy term neonates were randomly assigned to receive standard formula (n = 103) or formula containing 50% sn-2 palmitate (high-sn-2 formula; n = 100) for 12 wk. One hundred twenty breast-fed infants were also studied. The main outcome measures were 1) radial (single-photon absorptiometry) and whole-body (dual-energy X-ray absorptiometry) bone mineral content (WBBMC) at 12 wk and 2) stool frequency, volume, and consistency at 6 and 12 wk. Secondary outcome measures included stool fatty acid content. RESULTS: Infants receiving high-sn-2 formula had higher WBBMC (128.1 +/- 9.7 compared with 122.7 +/- 10.1 g, adjusted for size and sex), softer stools at 6 and 12 wk, and a lower proportion of stool soap fatty acids than did infants receiving the control formula. Breast-fed infants had adjusted WBBMC values (128.3 +/- 9.1 g) similar to those of infants fed high-sn-2 formula and significantly higher than those of infants fed the control formula. CONCLUSIONS: Changing the stereoisomeric structure of palmitate in infant formula resulted in higher WBBMC, reduced stool soap fatty acids, and softer stools more like those of breast-fed infants. The greater bone mass measured could be important if it persists beyond the trial period; this merits further investigation. PMID- 10539756 TI - De novo lipogenesis, lipid kinetics, and whole-body lipid balances in humans after acute alcohol consumption. AB - BACKGROUND: Acute alcohol intake is associated with changes in plasma lipid concentrations and whole-body lipid balances in humans. The quantitative roles of hepatic de novo lipogenesis (DNL) and plasma acetate production in these changes have not been established, however. OBJECTIVE: We used stable-isotope mass spectrometric methods with indirect calorimetry to establish the metabolic basis of changes in whole-body lipid balances in healthy men after consumption of 24 g alcohol. DESIGN: Eight healthy subjects were studied and DNL (by mass-isotopomer distribution analysis), lipolysis (by dilution of [1,2,3,4-(13)C(4)]palmitate and [(2)H(5)]glycerol), conversion of alcohol to plasma acetate (by incorporation from [1-(13)C(1)]ethanol), and plasma acetate flux (by dilution of [1 (13)C(1)]acetate) were measured. RESULTS: The fractional contribution from DNL to VLDL-triacylglycerol palmitate rose after alcohol consumption from 2 +/- 1% to 30 +/- 8%; nevertheless, the absolute rate of DNL (0.8 g/6 h) represented <5% of the ingested alcohol dose; 77 +/- 13% of the alcohol cleared from plasma was converted directly to acetate entering plasma. Acetate flux increased 2.5-fold after alcohol consumption. Adipose release of nonesterified fatty acids into plasma decreased by 53% and whole-body lipid oxidation decreased by 73%. CONCLUSIONS: We conclude that the consumption of 24 g alcohol activates the hepatic DNL pathway modestly, but acetate produced in the liver and released into plasma inhibits lipolysis, alters tissue fuel selection, and represents the major quantitative fate of ingested ethanol. PMID- 10539759 TI - The alcohol paradox. PMID- 10539761 TI - Meta-analysis of the cholesterol-lowering effects of dietary fiber. PMID- 10539762 TI - Fat malabsorption in cystic fibrosis patients. PMID- 10539765 TI - Ubiquitin-dependent signaling: the role of ubiquitination in the response of cells to their environment. AB - The response of a cell to its external environment requires rapid and significant alteration of protein amount, localization and/or function. This regulation involves a complex combination of processes that control synthesis, localization and degradation. All of these processes must be properly regulated and are often interrelated. Intracellular proteolysis is largely accomplished by the ubiquitin dependent system and has been shown to be required for growth control, cell cycle regulation, receptor function, development and the stress response. Substrates subject to regulated degradation by this system include cyclins and cyclin dependent kinase inhibitors, tumor suppressors, transcription factors and cell surface receptors. In addition, proteins that are damaged by oxidation or that are improperly folded or localized are substrates whose degradation by this system often leads to antigen presentation on the surface of the cell in the context of Class I major histocompatibility complex molecules. A very large body of work in the last fifteen years has shown that degradation by this system requires the covalent attachment of a small protein called ubiquitin and that this modification serves to direct target proteins for degradation by a 26S proteolytic particle, the proteasome. Thus, the attachment of the ubiquitin domain is of vital importance in regulating normal growth and differentiation, as well as in defending against cellular damage caused by xenobiotics, environmental insults, infection and mutation. This review focuses on the role of ubiquitination in the cellular signaling pathways that deal with these external influences. PMID- 10539766 TI - Metabolic differences between genetically lean and fat chickens are partly attributed to the alteration of insulin signaling in liver. AB - Insulin signaling [tyrosine phosphorylation of insulin receptor (IR), insulin receptor substrate-1 (IRS-1), Src homology and collagen protein (Shc) and phosphatidyl inositol 3'-kinase activity (PI 3'-kinase)] was studied in the liver and thigh muscles of fat (FL) and lean (LL) chickens. These lines result from a divergent selection on abdominal fat pad size. The divergence is of metabolic origin. Extreme nutritional states were studied (fed, 48-h starved and 30-min refed). Such conditions significantly altered insulin signaling in chicken liver, but surprisingly not in the muscle (except the phosphorylation of Shc in the refed state). No major differences that could account for this divergence were found in muscle. Liver IR number and Shc protein did not differ between genotypes. Liver IRS-1 (protein and messenger) was lower in the fed state and higher in the starved state in FL compared to that in LL chickens. In the fed state, tyrosine phosphorylation of liver IR, IRS-1 and Shc action was higher in FL than in LL chickens that in the absence of insulin resistance rely on higher plasma insulin levels. In the starved state, phosphorylation of liver IR was lower, but the phosphorylation of IR and IRS-1 were higher in LL than in FL chickens, most likely in response to higher plasma glucose and insulin in the lean genotype. In the refed state, the phosphorylation of liver IR and IRS-1 did not differ between genotypes despite significantly lower plasma insulin in FL chickens. Finally, PI 3'-kinase was not affected by the genotype. A significant activation of early steps of insulin signaling in liver of fed FL chickens may at least partly account for their increased liver lipogenesis and ultimately their fattening. PMID- 10539767 TI - Chronic alcohol consumption induces genomic but not p53-specific DNA hypomethylation in rat colon. AB - Alcohol consumption has been implicated as an etiologic agent in colorectal carcinogenesis, but the mechanism by which alcohol enhances the development of colorectal cancer is not yet known. Recent reports indicate that alcohol consumption can diminish cellular S-adenosylmethionine levels, thus possibly altering normal patterns of DNA methylation, a phenomenon that is mediated by S adenosylmethionine and whose abnormalities are observed in colonic neoplasia. This study investigated the effect of chronic alcohol consumption on genomic DNA methylation of rat colonic epithelium and methylation of the p53 tumor suppressor gene, abnormalities of which have been implicated in colonic carcinogenesis. Two groups of rats (n = 10/group) were pair-fed either an alcohol-containing or an isocaloric control Lieber-DeCarli diet for 4 wk. The extent of genomic DNA methylation was assessed by incubating the extracted DNA with [(3)H]S adenosylmethionine and Sss1 methyltransferase. Gene-specific methylation was assessed by using semiquantitative polymerase chain reaction (PCR). Tritiated methyl uptake by colonic DNA (which is inversely correlated with genomic methylation) from alcohol-fed rats was 57% less than that in control DNA (P < 0.05). However, gene-specific DNA methylation, both in the p53 gene (exons 5-8) and in the beta-actin gene, a control gene, did not differ between the two groups. In conclusion, this study indicates that chronic alcohol consumption produces genomic DNA hypomethylation in the colonic mucosa. This may constitute a means by which carcinogenesis is enhanced, although further studies are required to establish causality. PMID- 10539768 TI - High levels of dietary vitamin E do not replace cellular glutathione peroxidase in protecting mice from acute oxidative stress. AB - Our objective was to determine whether high levels of dietary vitamin E replaced the protection of the Se-dependent cellular glutathione peroxidase (GPX1) against paraquat- or diquat-induced acute oxidative stress in mice. Two experiments were conducted using GPX1 knockout [GPX1(-/-)] mice and wild-type (WT) mice (n = 78/group). In Experiment 1, mice were fed torula yeast-based, Se-adequate (0.4 mg/kg as sodium selenite) diets + 0, 75, 750 or 7,500 mg all-rac-alpha-tocopheryl acetate for 5 wk before an intraperitoneal injection of 50 mg paraquat/kg body weight. In Experiment 2, mice were fed the diet + 0 or 750 mg all-rac-alpha tocopheryl acetate for 5 wk and were killed 1 or 3 h after an injection of diquat at 12, 24 or 48 mg/kg. In Experiment 1, all mice died of the injection and there were 8- to 15-fold differences (P < 0.001) in survival times between the GPX1(-/ ) and the WT mice. Although increasing tocopheryl acetate from 0 to 750 mg/kg extended the survival time of the GPX1(-/-) mice for 2 h (P = 0.06), the highest tocopheryl acetate level resulted in a decrease (P < 0.05) in survival time in the WT mice. The vitamin E-deficient GPX1(-/-) mice had the highest concentration of hepatic thiobarbituric acid reacting substances. In Experiment 2, the diquat induced formation of hepatic F(2)-isoprostanes was accelerated (P < 0.05) by vitamin E deficiency and was also affected by the GPX1 knockout. Diquat produced much greater (P < 0.01) dose-dependent increases in plasma alanine transaminase (ALT) activities in the GPX1(-/-) than in the WT mice. Hepatic phospholipid hydroperoxide GPX activities were decreased (P < 0.05) by the diquat injection only in the vitamin E-deficient GPX1(-/-) mice. Despite a potent inhibition of hepatic lipid peroxidation, high levels of dietary vitamin E do not replace the protection of GPX1 against the paraquat-induced lethality or the diquat-induced plasma ALT activity increase in mice. PMID- 10539769 TI - Altered expression and glucocorticoid-inducibility of hepatic CYP3A and CYP2B enzymes in male rats fed diets containing soy protein isolate. AB - Hepatic CYP3A and CYP2B enzymes were studied in male Sprague-Dawley rats derived from 5-7 litters fed diets in which the protein source was either casein or soy protein isolate. At age 65 d, rats were gavaged with corn oil (vehicle) or 50 mg/kg dexamethasone. Hepatic expression of CYP3A and CYP2B1 mRNA, apoprotein and associated monooxygenase activities were measured. Consumption of soy diets significantly increased monooxygenase activity toward the following: the CYP3A substrates erythromycin and ethylmorphine N-demethylase; corticosterone and testosterone 6beta-hydroxylase; and apoprotein and mRNA expression of CYP3A2 (P < 0.05). Dexamethasone significantly induced turnover of erythromycin and testosterone, expression of CYP3A apoprotein, and expression of CYP3A1 and CYP3A2 mRNA (P < 0.05). In addition, significant diet-inducer interactions were observed in the expression of CYP3A apoprotein and activities toward ethylmorphine, corticosterone and testosterone (P < 0.05). Significant diet-inducer interactions were also observed on CYP2B1-dependent pentoxyresorufin O-depentylase activity (P < 0.05). However, although dexamethasone significantly induced CYP2B1 expression at the apoprotein and mRNA level (P < 0.05), no significant diet effects were observed. These data suggest potential effects of soy consumption on the metabolism of a wide variety of CYP3A and CYP2B1 substrates, especially in situations involving coexposure to CYP inducers. PMID- 10539770 TI - Dietary calcium is a major factor in 1,25-dihydroxycholecalciferol suppression of experimental autoimmune encephalomyelitis in mice. AB - The active form of vitamin D (1,25-dihydroxycholecalciferol) is a potent immune system regulator. Treating mice with 1, 25-dihydroxycholecalciferol and feeding them diets high in calcium can completely suppress the induction of experimental autoimmune diseases such as experimental autoimmune encephalomyelitis (EAE). Experiments described here were carried out on mice in which development of EAE was induced. Mice were fed diets containing various amounts of calcium and 1,25 dihydroxychole-calciferol. Variables measured were as follows: 1) incidence and severity of EAE; 2) serum calcium concentrations; 3) body weight; 4) total number of cells in the lymph nodes; and 5) interleukin-4 (IL-4) and transforming growth factor-beta1 (TGF-beta1) mRNA levels. When calcium was removed from the diet, the incidence of EAE was reduced 20% in both males and females. Further, the lower the dietary level of calcium, the higher was the dose of 1,25 dihydroxycholecalciferol required to prevent the symptoms. Thus, 1, 25 dihydroxycholecalciferol was found most effective in mice fed a diet adequate or high in calcium. 1,25-Dihydroxycholecalciferol treatment of mice fed high dietary calcium resulted in a decreased number of lymphocytes in the lymph nodes and increased IL-4 and TGF-beta1 mRNA levels. When calcium was omitted from the diet, 1, 25-dihydroxycholecalciferol supplementation increased TGF-beta1 mRNA. Increased IL-4 mRNA and decreased lymphocytes in the lymph nodes in response to 1,25-dihydroxycholecalciferol occurred only when dietary calcium was adequate or high. Our results suggest that dietary calcium and 1,25-dihydroxycholecalciferol are both involved in the prevention of symptomatic EAE. PMID- 10539771 TI - Pinus pinaster oil affects lipoprotein metabolism in apolipoprotein E-deficient mice. AB - The aim of the present study was to assess the antiatherogenic properties of Pinus pinaster (maritime pine) seed oil. To this end, the effects of P. pinaster oil supplementation on lipoprotein levels and atherosclerotic lesions were compared to those of lard or sunflower oil in apolipoprotein E-deficient mice. Plasma total cholesterol (P < 0.0001) and VLDL + intermediary density lipoprotein (IDL)-cholesterol (P < 0.0001) levels were lower in mice fed P. pinaster and sunflower oil than in those fed the lard diet. In contrast, triglycerides (P < 0.0001) and VLDL + IDL-triglycerides (P < 0.0001) levels were higher in mice fed P. pinaster oil than sunflower oil or lard. The VLDL + IDL lipid composition of apolipoprotein E-deficient mice fed P. pinaster oil was intermediate between that of lard-fed transgenic mice and that of wild-type mice fed nonpurified diet. Using the Triton WR1339 method, the fractional catabolic rate of plasma triglycerides was found to be lower in mice fed P. pinaster oil (P < 0.0001) than sunflower oil or lard diet, suggesting a defective clearance of triglycerides in the P. pinaster group. Finally, the susceptibility of triglyceride-rich lipoproteins to in vitro lipoprotein lipase-mediated lipolysis was lower in the P. pinaster oil-fed group than in the lard-fed group. Despite the differences in VLDL + IDL level and lipid composition, the surface areas of aortic atherosclerotic lesions were not significantly different among mice fed P. pinaster, sunflower or lard diets. In conclusion, the results of the present study indicated that feeding P. pinaster oil had no better preventive effect on aortic atherosclerotic lesion extension in apolipoprotein E-deficient mice than other saturated or polyunsaturated fats. PMID- 10539772 TI - Histidine-imbalanced diets stimulate hepatic histidase gene expression in rats. AB - A high protein concentration in the diet induces the gene expression of several amino acid degrading enzymes such as histidase (Hal) in rats. It is important to understand whether the amino acid pattern of the dietary protein affects the gene expression of these enzymes. The purpose of the present work was to study the effect of a histidine-imbalanced diet on the activity and mRNA concentration of rat hepatic histidase. Seven groups of six rats were fed one of the following diets: 1) 6% casein (basal), 2) 20% casein, 3) 35% casein, 4) an imbalance diet containing 6% casein plus a mixture of indispensable amino acids (IAA) equivalent to a 20% casein diet without histidine (I-20), 5) 6% casein plus a mixture of IAA equivalent to a 35% casein diet without histidine (I-35), 6) a corrected diet containing 6% casein plus IAA including histidine equivalent to a 20% casein diet, 7) a corrected diet containing 6% casein plus IAA including histidine equivalent to a 35% casein diet. Serum histidine concentration was inversely proportional to the protein content of the diet, and it was significantly higher in rats fed the corrected diets compared to their respective imbalanced diet groups. Hal activity increased as the protein content of the diet increased. Greater histidine imbalance resulted in lower food intake and higher Hal activity. Rats fed histidine-corrected diets had lower activity than their respective imbalanced groups. Differences in Hal activity were associated with differences in the concentration of Hal mRNA. These results indicate that rats fed a histidine-imbalanced diet exhibit reduced food intake and weight gain and increased Hal gene expression as a consequence of an increased amino acid catabolism. PMID- 10539773 TI - Secretion of phospholipid transfer protein by human hepatoma cell line, Hep G2, is enhanced by sodium butyrate. AB - Hep G2 cells were used to study the synthesis and secretion of phospholipid transfer protein (PLTP). Upon incubation of the cells at confluence with serum free Dulbecco's modified Eagle's medium (DMEM), phosphatidylcholine (PC) transfer activity was found to accumulate in the culture media. The PC transfer activity in the media was effectively inhibited by rabbit anti-human PLTP immunoglobulin (Ig)G, thus indicating that the PC transfer activity was due to secreted PLTP. The molecular weight of Hep G2 PLTP was approximately 78 kDa by Western blot analysis, in agreement with the molecular weight obtained for purified human plasma PLTP. The PLTP secreted by Hep G2 also possessed an HDL conversion activity similar to that of human plasma PLTP. The addition of butyrate to the cell culture media resulted in a marked increase in the secretion of PLTP. After 24 h incubation with 4 mmol/L sodium butyrate, a more than twofold increase (P < 0.01) of PC transfer activity in the cell-conditioned media was obtained. The dose-dependent increase in the PC transfer activity in the media upon butyrate treatment was well correlated (r = 0.80, P < 0.01) with that of PLTP mass as determined by immuno-slot blot analysis of cell-conditioned media. The increased secretion of PLTP by Hep G2 treated with sodium butyrate was accompanied by a greater increase in the level of PLTP mRNA in the cells as determined by ribonuclease protection assay. In the presence of 4 mmol/L sodium butyrate, a fourfold increase (P < 0. 01) in mRNA level was obtained at 24 h. No stabilizing effect of butyrate on PLTP mRNA was apparent upon treatment of the cultured cells with the RNA synthesis inhibitor, actinomycin D. Thus, the up-regulatory effect of butyrate on PLTP gene expression seemed to have occurred at the transcriptional level. PMID- 10539774 TI - Nutritional value of [15N]-soy protein isolate assessed from ileal digestibility and postprandial protein utilization in humans. AB - The purpose of this work was to assess the true oro-ileal digestibility, and to concurrently quantify the deamination of absorbed dietary nitrogen to examine the postprandial nutritional value of a soy protein isolate (SPI) in humans. To assess bioavailability and bioutilization of SPI, 10 healthy volunteers ingested 30 g of SPI, intrinsically and uniformly [15N]-labeled, added with 100 g of sucrose and water up to a final volume of 500 mL. True ileal digestibility was assessed by the [15N]-dilution method for 8 h by means of a naso-intestinal intubation technique. To describe and quantify exogenous nitrogen deamination for the same time period, urine and plasma samples were collected. True oro-ileal digestibility of SPI nitrogen was 91%. The amount of absorbed SPI amino acids used for nonoxidative disposal, i.e., postprandial biological value, was 86% 8 h after meal ingestion. Hence, net postprandial protein utilization of SPI was 78%. Compared to previous data that were assessed under the same condition in humans, the nutritional value of SPI is 92% of that in milk protein concentrate. PMID- 10539775 TI - Plasma L-5-oxoproline carbon and nitrogen kinetics in healthy young adults. AB - L-5-oxoproline (OP), an intermediate of the gamma-glutamyl cycle of glutathione synthesis and degradation, may serve as a probe for the state of glutathione kinetics. We explored the whole-body carbon and nitrogen kinetics of OP in five male healthy subjects (75.2 kg; 181 cm; 26 y) after a 5-d adaptation to an adequate L-amino acid-based diet (160 mg N x kg(-1) x d(-1); 188 kJ x kg(-1) x d( 1)), using a crossover design. On day 6 of the diet period, we carried out an 8-h tracer protocol (3 h fast; 5 h fed; 2/3 of daily nitrogen intake) with intravenous infusion of L-[1-(13)C]oxoproline and L-[3, 3-(2)H]cysteine or, in randomized order, on the second occasion, L-[(15)N]oxoproline and L-[3,3 (2)H]cysteine. Plasma OP was isolated by cation exchange and after addition of internal standards (DL-[(2)H(3)]-5-oxoproline; L-[(15)N, U-(13)C(5)]-5 oxoproline; DL-[(2)H(3)]-glutamic acid) derivatized to form TBDMS esters and measured by gas chromatography/mass spectrometry. Plasma OP concentration did not differ between fed and fasted state (fast: 59. 4 +/- 8.3; fed 59.2 +/- 8.9 nmol/mL). (13)C- and (15)N OP flux during the fasted and fed state were 19 +/- 3.6, 21.2 +/- 3.2, and 22.6 +/- 3.9, 25.8 +/- 4.3 micromol x kg(-1) x 30 min(-1), respectively. OP oxidation was 15.6 +/- 3.6 and 17.9 +/- 3.5 micromol x kg(-1) x 30 min(-1), in fasting and feeding, respectively, (P < 0.05). More than 80% of the plasma flux was oxidized. These findings are compared with the published literature on GSH turnover in plasma of human subjects and underscore the need to define more completely the dynamic aspects of glutathione metabolism and of the intermediates of the gamma-glutamyl cycle. PMID- 10539776 TI - Fasting increases serum total cholesterol, LDL cholesterol and apolipoprotein B in healthy, nonobese humans. AB - Voluntary fasting is practiced by many humans in an attempt to lose body weight. Conflicting results have been published on the effects of food deprivation on serum lipids. To study the effect of acute starvation on serum lipids, 10 nonobese (93-124% of ideal body weight), healthy adults (6 men, 4 women, 21-38 y old) fasted (no energy) for 7 d. Fasting increased total serum cholesterol from 4.90 +/- 0.23 to 6.73 +/- 0.41 mmol/L (37.3 +/- 5.0%; P < 0.0001) and LDL cholesterol from 2.95 +/- 0.21 to 4.90 +/- 0.36 mmol/L (66.1 +/- 6. 6%; P < 0.0001). Serum apolipoprotein B (apo B) increased from 0.84 +/- 0.06 to 1.37 +/- 0.11 g/L (65.0 +/- 9.2%; P < 0.0001). The increases in serum cholesterol, LDL and apo B were associated with weight loss. Fasting did not affect serum concentrations of triacylglycerol and HDL cholesterol. Serum concentrations of insulin-like growth factor-I (IGF-I) decreased from 246 +/- 29 (prefast) to 87 +/ 10 microg/L after 1 wk of fasting (P < 0.0001). We conclude that, in nonobese subjects, fasting is accompanied by increases in serum cholesterol, LDL and apo B concentrations, whereas IGF-I levels are decreased. PMID- 10539777 TI - Oxidative stress in humans during work at moderate altitude. AB - Increased oxidative stress has been associated with work at high altitude; however, it is not known whether oxidative stress is a significant problem at moderate altitudes. The oxidative stress indicators, breath pentane (BP), 8 hydroxydeoxyguanosine (8-OHdG), oxygen radical absorption capacity (ORAC), 4 hydroxynonenal (4-HNE), malondialdehyde (MDA), and lipid peroxides (LPO) were measured in breath, blood and urine samples of U.S. Marines engaged in moderate altitude ( approximately 3000 m) cold weather field training. The test subjects were divided into a placebo and four antioxidant supplement groups (n = 15/group) and received the following supplements for 28 d: 1) vitamin E, 440 alpha tocopherol equivalents (alpha-TE); 2) vitamin A, 2000 retinol equivalents (RE) of beta-carotene; 3) vitamin C, 500 mg ascorbic acid; 4) a mixture of 440 alpha-TE, 2000 RE of beta-carotene, 500 mg ascorbic acid, 100 microg selenium and 30 mg zinc daily. Strenuous work ( approximately 23 MJ/d) in cold weather at moderate altitude was accompanied by increases in several indicators of oxidative stress that were not effectively controlled by conventional antioxidant supplements. The group receiving the antioxidant mixture exhibited lower BP (P < 0. 05) compared with those receiving single antioxidant supplements; however, not all markers of oxidative stress responded like BP. Because these markers did not respond in the same manner, it is important to include markers from more than one source to assess the effect of supplemental dietary antioxidants. PMID- 10539778 TI - Long-term weekly iron supplementation improves and sustains nonpregnant women's iron status as well or better than currently recommended short-term daily supplementation. AB - This 7-mo double-blind study compared the efficacy of two iron supplementation schemes in improving iron nutrition among 116 healthy fertile-age women. They were randomly distributed in three groups, receiving: Group 1, iron + folate (60 mg and 250 microg, respectively) daily for 3 mo (currently recommended scheme), and folate (250 microg) weekly the subsequent 4 mo. Group 2, folate daily, and 60 mg iron only once weekly for 3 mo, and then weekly iron + folate for 4 mo. Group 3, folate daily for 3 mo and then weekly for 4 mo. At baseline, 16% had depleted stores (plasma ferritin <15 microg/L) and 16% had hemoglobin levels <125 g/L. Eight percent had hemoglobin levels <120 g/L. In Group 1 hemoglobin and ferritin increased at 3 mo but returned to near basal conditions after 4 mo of weekly folate. In Group 2, hemoglobin and ferritin increased progressively throughout the 7 mo but mostly after 3 mo. Group 3 did not change. Side effects were highest with daily iron. Weekly iron supplementation over 7 mo (30 doses) improved and sustained iron nutrition at least as effectively and was better tolerated than 90 daily iron supplements consumed during 3 mo. PMID- 10539779 TI - Dietary vitamin A intakes of preschool-age children in South India. AB - The vitamin A intake of children aged 1-3 y (n = 683) was assessed using a quantitative food-frequency questionnaire in a vitamin A intervention study in South India. Trained field workers interviewed mothers about their children's usual consumption of common sources of vitamin A and collected information on portion sizes using standard cups. Mothers were asked to state the number of months in a year during which specific seasonal foods were available. Information about current breast-feeding was also obtained. Vitamin A intakes from nonbreast milk sources were extremely low at all ages. The median intake of total vitamin A, beta-carotene and retinol was 121, 100 and 21 retinol equivalents (RE), respectively. Maternal education and socioeconomic status (SES) were positively associated with total vitamin A and retinol intakes. Girls had significantly lower intakes than boys even after adjusting for differences in age, maternal education, SES and breast-feeding status. Breast-feeding was common, but declined to 60% by 24 mo and to 15% by 36 mo. Vitamin A intakes from nonbreast milk sources increased with age only for currently breast-fed children, who tended to be of lower SES. After taking into account the potential contribution of breast milk by using published estimates, nonbreast-fed children met only 60% of the Indian recommended dietary allowance (RDA; 250 RE/d), whereas breast-fed children met approximately 90% of the RDA during y 2 of life. PMID- 10539780 TI - Functional biochemical and nutrient indices in frail elderly people are partly affected by dietary supplements but not by exercise. AB - A decline in dietary intake due to inactivity and, consequently, development of a suboptimal nutritional status is a major problem in frail elderly people. However, benefits of micronutrient supplementation, all-round physical exercise or a combination of both on functional biochemical and hematologic indicators of nutritional and health status in frail elderly subjects have not been tested thoroughly. A 17-wk randomized controlled trial was performed in 145 free-living frail elderly people (43 men, 102 women, mean age, 78 +/- 5.7 y). Based on a 2 x 2 factorial design, subjects were assigned to one of the following: 1) nutrient dense foods, 2) exercise, 3) both (1) and (2) or 4) a control group. Foods were enriched with micronutrients, frequently characterized as deficient [25-100% of the recommended daily allowance (RDA)] in elderly people. Exercises focused on skill training, including strength, endurance, coordination and flexibility. Dietary intake, blood vitamin levels and nutritional and health indicators, including (pre)albumin, ferritin, transferrin, C-reactive protein, hemoglobin and lymphocytes were measured. At baseline, 28% of the total population had an energy intake below 6.3 MJ, up to a maximum of 93% having vitamin intakes below two thirds of the Dutch RDA. Individual deficiencies in blood at baseline ranged from 3% for erythrocyte glutathione reductase-alpha to 39% for 25-hydroxy vitamin D and 42% for vitamin B-12. These were corrected after 17 wk in the two groups receiving the nutrient-dense foods, whereas no significant changes were observed in the control or exercise group. Biochemical and hematologic indicators at baseline were within the reference ranges (mean albumin, 46 g/L; prealbumin, 0.25 g/L; hemoglobin, 8.6 mmol/L) and were not affected by any of the interventions. The long-term protective effects of nutrient supplementation and exercise, by maintaining optimal nutrient levels and thereby reducing the initial chance of developing critical biochemical values, require further investigation. Other indicative functional variables for suboptimal nutritional status, in addition to those currently selected, should also be explored. PMID- 10539781 TI - Dietary conjugated linoleic acids increase lean tissue and decrease fat deposition in growing pigs. AB - Conjugated linoleic acids (CLA) decrease the body fat content of rodents; the aim of this study was to determine whether dietary CLA altered carcass composition of pigs. Female Large White x Landrace pigs (n = 66) were used in this study. To obtain initial body composition, six pigs were slaughtered at 57 kg live weight, whereas the remaining pigs were allocated to one of six dietary treatments (0, 1.25, 2.5, 5.0, 7.5 and 10.0 g/kg CLA, containing 55% of CLA isomers). The diets, containing 14.3 MJ digestible energy (DE) and 9. 3 g available lysine per kg, were fed ad libitum for 8 wk. Dietary CLA had no significant effect on average daily gain (861 vs. 911 g/d for pigs fed diets with and without CLA, P = 0.15) or feed intake (2. 83 vs. 2.80 kg/d, P = 0.74). The gain to feed ratio was increased by dietary CLA by 6.3% (0.328 vs. 0.348, P = 0.009). Fat deposition decreased linearly (-8.2 +/- 2.09 g/d for each gram per kilogram increase in CLA concentration; P < 0.001) with increasing inclusion of CLA. At the highest level of CLA inclusion, fat deposition was decreased by 88 g/d (-31%). Similarly, the ratio of fat to lean tissue deposition decreased linearly (-0.093 +/- 0.0216 for each gram per kilogram increase in CLA concentration; P < 0.001) with increasing dietary CLA. The carcass lean tissue deposition response to dietary CLA was quadratic in nature and was maximized (+25%) at 5. 0 g/kg dietary CLA. Overall, dietary CLA increased the gain to feed ratio and lean tissue deposition and decreased fat deposition in finisher pigs. PMID- 10539782 TI - Dietary potassium bicarbonate and potassium citrate have a greater inhibitory effect than does potassium chloride on magnesium absorption in wethers. AB - We addressed the question whether the type of anion in potassium salts affects magnesium absorption and the transmural potential difference by using wethers (n = 8) fed a control diet and diets supplemented with equimolar amounts of KHCO(3), KCl or K-citrate according to a Latin-square design. The control diet contained 10.9 g K/kg dry matter and the high K diets contained 41.3 g K/kg dry matter. Compared with the control diet, KHCO(3) and K-citrate significantly reduced apparent Mg absorption by 9.5 and 6.5%, respectively. Supplemental KCl tended to reduce (P = 0.070) group mean magnesium absorption by 5.5%. Consumption of supplemental KHCO(3) and K-citrate produced a significant increase in the transmural potential difference (serosal side = positive) by 17.1 and 20.7 mV, respectively, whereas the addition of KCl to the diet did not. The individual values for the four diets tended to show a negative correlation (r = -0.336, n = 32, P = 0.060) between the transmural potential difference and apparent magnesium absorption. We conclude that different potassium salts have different effects on magnesium absorption in ruminants as caused by different effects on the transmural potential difference. PMID- 10539783 TI - Dietary marine algae (Schizochytrium sp.) increases concentrations of conjugated linoleic, docosahexaenoic and transvaccenic acids in milk of dairy cows. AB - Modification of milk fat to contain long-chain (n-3) fatty acids and increased concentrations of conjugated linoleic acid has potential for improving health of consumers. Natural modification of milk through nutritional manipulation of diets for dairy cows is preferable to post-harvest modification. The objectives of this study were to increase the concentrations of beneficial fatty acids in milk fat by feeding a diet rich in (n-3) fatty acids from algae to dairy cows. Cows were fed a control diet, a diet containing algae (Schizochytrium sp.) protected against ruminal biohydrogenation, or a diet containing unprotected algae for 6 wk. Feed intake and milk production were recorded daily. Milk samples were obtained weekly for analysis of milk composition and profile of fatty acids. Percentage of fat in milk of cows fed algae was lower (P < 0.01) than in milk from cows fed the control diet; however, energy-corrected milk production did not differ (P > 0.05). Inclusion of algae in diets decreased (P < 0.01) feed intake. Milk fat from cows fed algae contained greater (P < 0.01) concentrations of conjugated linoleic acid, (n-3) fatty acids (particularly docosahexaenoic acid), and transvaccenic acid. Concentrations of docosahexaenoic acid were greater (P < 0.01) in milk fat from cows fed protected algae compared to milk fat from cows fed unprotected algae. Milk fat from cows fed algae contained lower (P < 0.05) concentrations of total saturated fatty acids compared to cows fed the control diet. In conclusion, milk fat can be modified through nutritional management of dairy cows to provide more favorable fatty acids for consumers. PMID- 10539784 TI - Chronic feeding of a low boron diet adversely affects reproduction and development in Xenopus laevis. AB - The aims of this work were as follows: 1) to determine whether a purified diet currently used for studies with rats was acceptable for reproductive studies in frogs; and 2) to determine whether frogs are sensitive to a deficit of boron (B) in the diet. Adult Xenopus laevis were fed a nonpurified beef liver and lung (BLL) diet (310 microg B/kg), a purified diet supplemented with boron (+B; 1850 microg B/kg), or a purified diet low in boron (-B; 45 microg B/kg) for 120 d. Frogs fed the BLL and +B diets produced 11.3 and 12.2% necrotic eggs, respectively. Abnormal gastrulation occurred in <4% of the fertilized eggs in both groups, and 96-h larval survival exceeded 75% in both groups. In contrast, frogs fed the -B diet for 120 d produced a high proportion of necrotic eggs (54%). Fertilized embryos from the -B diet-fed frogs showed a high frequency of abnormal gastrulation (26.8%), and >80% of the embryos died before 96 h of development. Mean embryo cell counts at X. laevis developmental stage 7.5 (mid blastula) were significantly lower in the -B embryos than in the BLL or +B embryos. BLL and -B embryos grown in low boron culture media had a high frequency of malformations compared with embryos grown in boron-supplemented media. These studies show that a purified diet that has been used in rodent studies was acceptable for reproduction studies in X. laevis. This work also demonstrates that a diet low in boron markedly impairs normal reproductive function in adult X. laevis, and that administration of the low boron diet results in an increase in both incidence and severity of adverse effects. In addition, these studies demonstrate the usefulness of the X. laevis model in nutrition studies. PMID- 10539785 TI - Protein malnutrition affects the growth trajectories of the craniofacial skeleton in rats. AB - To investigate the effects of protein malnutrition on a normal growth trajectory, we radiographed Rattus norvegicus from 22 d (weaning) and continuing past adult size. We took measurements from longitudinal radiographs of rats fed a control diet and littermates fed an isocaloric low protein experimental diet. A Gompertz model was fit to each individual rat for body weight and 22 measurements of the craniofacial skeleton, producing parameters that described the rate and timing of growth. We tested for differences in these parameters due to diet, sex and litter with a mixed-model three-way ANOVA. Allometric analysis examined the scaling relationships between and within various regions of the skull. For most measurements, final sizes predicted by the model were not significantly different between rats fed the two diets, although the differences in final measurements showed small, but significant differences in growth between rats in the two diet groups. The instantaneous initial rate of growth, maximum rate of growth and deceleration of growth were significantly higher in the control rats for every measurement. Rats fed the low protein diet grew for a significantly longer period of time. The shape of the neurocranium was relatively conserved between diet groups; however, rats fed the low protein diet had shorter and relatively wider skulls than the controls. These results suggest that functional demands of the viscerocranium were greater after birth, and that growth in this area was faster. The viscerocranium reached functional adult proportions earlier and was therefore more susceptible to epigenetic perturbations such as dietary protein level. Protein malnutrition did not affect many aspects of adult size, but strongly altered the growth trajectory to achieve that size. PMID- 10539786 TI - Folic acid supplementation of pregnant mice suppresses heat-induced neural tube defects in the offspring. AB - Neural tube defects (NTD) are a group of malformations that result from the failure of the neural tube to close early in embryonic development and among the most common congenital malformations in humans. It has been reported that a substantial proportion of NTD in humans can be prevented by folic acid (FA) supplementation prior to conception and during the first months of pregnancy, and myo-inositol (MI) was shown to reduce the incidence of NTD in curly tail mice which are not prevented by FA. Brief maternal hyperthermia (HT) early in pregnancy has been implicated in NTD both in humans and laboratory animals, and anterior NTD including exencephaly and anencephaly are induced frequently when pregnant mice are exposed to HT. We examined the effect of FA or MI supplementation of pregnant mice on the occurrence of heat-induced NTD in the offspring. When pregnant mice were treated with FA (3 mg/kg) daily from gestational day (GD) 0.5 through GD 9.5 and heated at GD 8.5, the prevalence of NTD in the fetuses (26.6%) was significantly lower than the corresponding figure in the HT alone group (38.6%; P < 0.05). However we failed to detect the preventive effect of MI (500 mg/kg). The results of this study suggest that prenatal FA supplementation decreases HT-induced NTD in mice and sufficient FA intake during early pregnancy may be recommended to avoid the birth of malformed children. PMID- 10539787 TI - Dietary docosahexaenoic acid-enriched phospholipids normalize urinary melatonin excretion in adult (n-3) polyunsaturated fatty acid-deficient rats. AB - Melatonin (MEL) plays an essential role in physiologic functions associated with darkness. We examined the effects of docosahexaenoic acid (DHA)-enriched phospholipids from pig brains (BPL) or hen eggs (EPL), as sources of DHA, on lipid FA composition of pineal membranes and daytime and nighttime concentrations of 6-sulfatoxymelatonin (aMT6) in adult male control and (n-3)-deficient rats fed BPL and EPL diets for 5 wk. In two experiments, at 3 wk of age, rats were divided into subgroups and fed semipurified diets containing either peanut oil [(n-3) deficient group] or peanut plus rapeseed oil (control group) and two dietary formulas containing either 3.5 g/100 g diet of BPL (Experiment 1) or 5.0 g/100 g diet of EPL (Experiment 2). BPL and EPL diets provided approximately 200 mg of DHA/100 g diet. During the daytime, aMT6 concentrations were not significantly different among groups. Conversely, the (n-3)-deficient rats had significantly lower nighttime aMT6 concentrations than the control rats. BPL and EPL did not affect urinary nighttime aMT6 concentration in the control group, whereas (n-3) deficient + BPL or EPL groups exhibited significantly higher nighttime aMT6 concentrations than the (n-3)-deficient group (76 and 110%, respectively). The level of DHA was significantly higher in the pineal glands of control rats than in (n-3)-deficient rats. In rats fed EPL and BPL, the level of DHA reached a plateau, between 10 and 11 mg/100 mg total fatty acids in control + BPL or EPL and (n-3)-deficient + BPL or EPL groups. These findings suggest that new DHA enriched formulas may be used as an efficient alternative source of (n-3) polyunsaturated fatty acids to normalize MEL secretion. PMID- 10539788 TI - Psyllium shifts the fermentation site of high-amylose cornstarch toward the distal colon and increases fecal butyrate concentration in rats. AB - We examined the combination effects of psyllium (PS) and resistant starch on large bowel short-chain fatty acids (SCFA). Rats were fed one of the following four diets: low amylose (LAS) or high amylose cornstarch diets (HAS, 50 g/kg diet) with or without 15 g PS/kg diet (LAS/PS and HAS/PS diets). HAS and/or PS were substituted for the same amounts of LAS in diets. Cecal butyrate concentrations were significantly higher in rats fed the HAS and HAS/PS diets than in those fed the LAS and LAS/PS diets. However, butyrate and total SCFA concentrations in rats fed the HAS diet decreased along the length of the colon and fecal butyrate concentration was reduced to one-third of that in the cecum. In contrast, the HAS/PS diet maintained higher butyrate concentrations throughout the large bowel. Fecal butyrate concentration in the HAS/PS diet-fed group significantly exceeded the sum of the concentrations in rats fed the LAS/PS and HAS diets. PS supplementation to the HAS diet significantly increased fecal starch excretion by 10 fold compared with that of rats fed the HAS diet. There was a positive correlation between fecal butyrate concentration and fecal starch excretion (r = 0.709, P < 0.0001). In a further experiment, ileorectostomized rats were fed the HAS and HAS/PS diets. From the difference in fecal starch excretion between normal and ileorectostomized rats, starch degradation by large bowel microflora in rats fed the HAS and HAS/PS diets was deduced to be 96% and 63%, respectively. These findings support the hypothesis that PS may delay the fermentation rate of HAS in the cecum and shift the fermentation site of HAS toward the distal colon, leading to the higher butyrate concentration in the distal colon and feces. PMID- 10539789 TI - Docosahexaenoic and arachidonic acid prevent a decrease in dopaminergic and serotoninergic neurotransmitters in frontal cortex caused by a linoleic and alpha linolenic acid deficient diet in formula-fed piglets. AB - This study examined the effects of diets deficient (D) in linoleic [18:2(n-6)] and linolenic acid [18:3(n-3)] at 0.8 and 0.05% energy, respectively, or adequate (C) in 18:2(n-6) and 18:3(n-3) at 8.3 and 0.8% energy, respectively, without (-) or with (+) 0.2% energy arachidonic [20:4(n-6)] and 0.16% energy docosahexaenoic [22:6(n-3)] acid in piglets fed from birth to 18 d. Frontal cortex dopaminergic and serotoninergic neurotransmitters and phospholipid fatty acids were measured. Piglets fed the D- diet had significantly lower frontal cortex dopamine, 3,4 dihydroxyphenylacetic (DOPAC), homovanillic acid (HVA), serotonin and 5 hydroxyindoleacetic acid (5-HIAA) concentrations than did piglets fed the C- diets. Frontal cortex dopamine, norepinephrine, DOPAC, HVA, serotonin and 5-HIAA were higher in piglets fed the D+ compared to those fed the D- diet (P < 0.05) and not different between piglets fed the D+ and those fed the C- diets or the C- and C+ diets. Piglets fed the D- diet had lower frontal cortex phosphatidylcholine (PC) and phosphatidylinositol (PI) 20:4(n-6) and PC and phosphatidylethanolamine (PE) 22:6(n-3) than did piglets fed the C- diet (P < 0.05). Piglets fed the D+ diet had higher frontal cortex PC and PI 20:4(n-6) and PC, PE, PS and PI 22:6(n-3) than did piglets fed the D- diet. These studies show that dietary essential fatty acid deficiency fed for 18 d from birth affects frontal cortex neurotransmitters in rapidly growing piglets and that these changes are specifically due to 20:4(n-6) and/or 22:6(n-3). PMID- 10539791 TI - Response to drs. moya-camarena and belury PMID- 10539790 TI - The role of dietary fat in child nutrition and development: summary of an ASNS workshop. American Society for Nutritional Sciences. PMID- 10539792 TI - Diabetes and cardiovascular disease: current opinions and future directions. PMID- 10539793 TI - Type 2 diabetes and macrovascular disease: epidemiology and etiology. PMID- 10539794 TI - Treatment of diabetes mellitus: implications of the use of oral agents. PMID- 10539795 TI - FDA approach to the regulation of drugs for diabetes. PMID- 10539796 TI - Therapy of type 2 diabetes, cardiovascular death, and the UGDP. PMID- 10539797 TI - Hyperglycemia and hyperinsulinemia at diagnosis of diabetes and their association with subsequent cardiovascular disease in the United Kingdom prospective diabetes study (UKPDS 47). PMID- 10539799 TI - Diabetes and ischemic heart disease. PMID- 10539798 TI - Intensive insulin therapy in patients with type 2 diabetes: implications of the Veterans affairs (VA CSDM) feasibility trial. PMID- 10539800 TI - Relation between sulfonylurea therapy, complications, and outcome for elderly patients with acute myocardial infarction. PMID- 10539802 TI - Patients with diabetes did better with coronary bypass graft surgery than with percutaneous transluminal coronary angioplasty: was this BARI finding real? PMID- 10539801 TI - Diabetes and acute myocardial infarction: the role of insulin therapy. PMID- 10539804 TI - Hypertension and diabetes: current issues. PMID- 10539803 TI - Comparison of outcome after coronary angioplasty and coronary surgery for multivessel coronary artery disease in persons with diabetes. PMID- 10539805 TI - Atherosclerotic disease in non-insulin-dependent diabetes mellitus: role of abnormal lipids and the place for lipid-altering therapies. PMID- 10539806 TI - Insulin resistance syndrome and cardiovascular disease: genetics and connections to skeletal muscle function. PMID- 10539808 TI - First steps toward understanding the pathophysiologic link between air pollution and cardiac mortality. PMID- 10539807 TI - Concern for azotemia with angiotensin-converting enzyme inhibitors: public health implications and clinical relevance. PMID- 10539809 TI - ASDOS closures: congratulations and technical observations. PMID- 10539810 TI - Relation between the renin-angiotensin-aldosterone system and left ventricular structure and function in young normotensive and mildly hypertensive subjects. AB - BACKGROUND: High angiotensin II levels in relation to the corresponding urinary sodium excretion have been found to modulate left ventricular (LV) structure in middle-aged hypertensive patients. To analyze whether such a relation between the renin-angiotensin-aldosterone system and left ventricular structure is already present in young individuals, we examined the changes of angiotensin II and aldosterone in response to increased salt intake and their relations to LV structure and function. METHODS: In 119 young (aged 26 +/- 3 years) patients with normal or mildly elevated blood pressure, we determined LV structure and function (2-dimensional guided M-mode echocardiography and pulse wave Doppler sonography) and 24-hour ambulatory blood pressure (SpaceLabs 90207). Dietary sodium intake as estimated by 24-hour urinary sodium excretion, plasma renin activity, angiotensin II, and aldosterone concentrations were measured first on a normal diet and second at high salt intake to determine the extent of the resulting suppression of the renin-angiotensin-aldosterone system. RESULTS: Body mass index (r = 0.43, P <.001) and both systolic (r = 0.24, P <. 01) and diastolic (r = 0.19, P <.05) 24-hour ambulatory blood pressure correlated with LV mass. No straightforward relation was found between LV structure and baseline angiotensin II or aldosterone concentration. The increase of sodium excretion at high salt intake was related to a physiologically expected decrease of angiotensin II and aldosterone levels in normotensive (r = -0.36, P <.01 and r = -0.32; P =.016, respectively) but not in hypertensive patients. Changes in angiotensin II or aldosterone concentration were not related to LV structure in either hypertensive or normotensive young individuals. However, changes in aldosterone secretion correlated with diastolic filling parameters in hypertensive patients (velocity time integrals of the A over E wave: r = 0.32, P =.03; atrial contribution of LV filling: r = 0.33, P =. 025) but not in normotensive individuals. CONCLUSION: In contrast to middle-aged hypertensive patients, neither angiotensin II, aldosterone, nor their suppression in response to high salt intake were related to LV structure in young hypertensive patients. However, inadequate suppression of aldosterone after salt intake was associated with diastolic filling abnormalities in our young hypertensive patients, which may represent early changes in hypertensive heart disease and precede potential structural alterations. PMID- 10539811 TI - Angiotensin-converting enzyme inhibitor compliance and dosing among patients with heart failure. AB - BACKGROUND: The efficacy of angiotensin-converting enzyme (ACE) inhibitors in treating heart failure is well established, but there is concern that these agents are underutilized. Proper treatment is contingent both on appropriate medication dosing by the physician and on patient compliance with therapy. This study examined dosing and compliance with ACE inhibitors in routine clinical practice. METHODS AND RESULTS: Data were integrated medical and pharmacy claims from 869 patients with heart failure. Compliance and dosing of ACE inhibitors was examined for each patient over a 10- to 17-month period. Patients had ACE inhibitors available on 71% of the days assessed. At 180 days after their index prescription, 86% of patients continued to take an ACE inhibitor. The mean percentage of an adequate daily dose of ACE inhibitors dispensed per prescription was 79%, but only 34% of patients were dispensed >/=100% of an adequate daily dose. A number of variables were found to independently predict compliance and dosing levels in the multivariate analyses. CONCLUSIONS: Both physician-dependent and patient-dependent factors contributed significantly to ACE inhibitor underutilization. Each of these factors must be addressed to improve compliance and dosing of ACE inhibitors in routine clinical care. PMID- 10539812 TI - Variations in family physicians' and cardiologists' care for patients with heart failure. AB - BACKGROUND: Improved understanding of the reasons for underuse of diagnostic tests and treatments for congestive heart failure (CHF) may be helpful for designing future interventions to improve quality of care. METHODS: To determine differences between family physicians' and cardiologists' practice styles for diagnosis and treatment of CHF, a random sample of family physicians and cardiologists were surveyed with standardized case scenarios. RESULTS: Survey respondents were 182 family physicians and 163 cardiologists. Family physicians were less likely than cardiologists to rate measurement of left ventricular ejection fraction as "very important" for patients with new CHF, less likely to order an echocardiogram or test for ischemia, and much less likely to identify diastolic dysfunction as a cause of CHF. Family physicians were more likely to prescribe digoxin when it was not indicated (diastolic dysfunction) and less likely to prescribe digoxin and an angiotensin-converting enzyme (ACE) inhibitor when they were indicated (moderately to severely reduced left ventricular ejection fraction). Family physicians expressed more concern over the risks of ACE inhibitors in patients with blood pressure of 100/70 mm Hg or serum creatinine of 2.0 mg/dL and were less likely to prescribe an ACE inhibitor in these settings. Family physicians overestimated the risks of warfarin use for atrial fibrillation and were therefore less likely to prescribe warfarin. CONCLUSIONS: Family physicians appear to have less understanding of CHF pathophysiology (ie, systolic versus diastolic dysfunction) and how treatment differs according to the underlying disease process. Overestimation of the risk of ACE inhibitor and warfarin use may result in underprescribing these medications. PMID- 10539813 TI - Sex differences in the clinical care and outcomes of congestive heart failure in the elderly. AB - BACKGROUND: There is evidence for sex differences in treatment and outcome of ischemic heart disease. However, little and conflicting data exist about sex differences in the care and outcome of elderly patients with heart failure. METHODS: We compared mortality rate, readmission, and use of selected treatments and procedures between women and men in a database of 2445 patients (1426 women) aged >/=65 admitted for heart failure to 18 Connecticut hospitals in 1994 and 1995. Demographic and clinical data were abstracted from the medical records. RESULTS: Women were older and more likely to have a history of hypertension whereas men more often had previous coronary heart disease. Women had more preserved left ventricular systolic function and higher systolic blood pressure on presentation than men. Treatments on day 1 (aspirin, angiotensin-converting enzyme [ACE] inhibitors, and diuretics), procedures during admission (assessment of left ventricular function, coronary angiography, and revascularization), and use of ACE inhibitors among ideal candidates at discharge were similar in men and women. Six-month rehospitalization rates were also similar. Although 30-day mortality rate did not differ between men and women, 6-month and 1-year mortality rates were lower in women after age adjustment (relative risk for 6-month death 0.81, 95% confidence interval, 0.68-0.95). In multivariable analysis, sex differences in mortality rate were reduced (relative risk 0.90, 95% confidence intervals, 0.75-1.08). History of hypertension, systolic blood pressure on admission, and left ventricular function mostly explained the observed sex differences in mortality rate. CONCLUSIONS: Female and male patients hospitalized for heart failure have a similar hospital course, treatment pattern, and readmission rates, but women live longer than men. When baseline differences are accounted for, the mortality risk of women and men becomes very similar. PMID- 10539814 TI - ARCTIC: assessment of haemodynamic response in patients with congestive heart failure to telmisartan: a multicentre dose-ranging study in Canada. AB - BACKGROUND: The aim of this study was to examine the acute hemodynamic and neurohormonal effects of the angiotensin II antagonist telmisartan relative to placebo in patients with chronic symptomatic (New York Heart Association class II to III) congestive heart failure and to explore the dose-response relation for these effects. METHODS AND RESULTS: After baseline hemodynamic and neurohormonal measurements made with the use of a pulmonary artery and radial arterial catheter, 82 patients were randomly assigned to placebo or 10, 20, 40, or 80 mg of telmisartan in a double-blind fashion. Hemodynamic and neurohormonal measurements were carried out over 24 hours. Telmisartan caused significant decreases in systemic arterial, pulmonary arterial, and pulmonary capillary wedge pressures with evidence of a dose-response relation for each of these parameters. The drug had no significant effects on heart rate, cardiac index, or systemic vascular resistance. Telmisartan did not have consistent effects on either plasma norepinephrine or plasma atrial natriuretic peptide levels, although it did cause significant increases in both plasma renin activity and angiotensin II levels at higher doses. CONCLUSIONS: The acute administration of the angiotensin II antagonist telmisartan was associated with significant dose-dependent reductions in systemic arterial blood pressure and pulmonary pressures. Long-term follow-up studies are required to translate changes in hemodynamic parameters into a clinical benefit. PMID- 10539815 TI - Predictors of decreased renal function in patients with heart failure during angiotensin-converting enzyme inhibitor therapy: results from the studies of left ventricular dysfunction (SOLVD) AB - BACKGROUND: Although angiotensin-converting enzyme inhibitor therapy reduces mortality rates in patients with congestive heart failure (CHF), it may also cause decreased renal function. Little information is available to predict which patients are at highest risk for this complication. OBJECTIVE: To quantify specific clinical predictors of reduction in renal function in patients with CHF who are prescribed angiotensin-converting enzyme inhibitor therapy. METHOD: We analyzed data from the Studies of Left Ventricular Dysfunction (SOLVD), a randomized, double-blind, placebo-controlled trial of enalapril for the treatment of CHF. There were 3379 patients randomly assigned to enalapril with a median follow-up of 974 days and 3379 patients randomly assigned to placebo with a mean follow-up of 967 days. Decreased renal function was defined as a rise in serum creatinine >/=0.5 mg/dL (44 micromol/L) from baseline. We used time-to-event analysis to identify potential predictors of decrease in renal function including age, baseline ejection fraction, baseline creatinine, low systolic blood pressure (<100 mm Hg), history of hypertension, diabetes, and use of antiplatelet, diuretic, and beta-blocker therapy. RESULTS: Patients randomly assigned to enalapril had a 33% greater likelihood of decreased renal function than controls (P =.003). By multivariate analysis, in both the placebo and enalapril groups older age, diuretic therapy, and diabetes were associated with decreased renal function, whereas beta-blocker therapy and higher ejection fraction were renoprotective. Older age was associated with a greater risk of developing decreased renal function in both groups, but significantly more so in the enalapril group (enalapril: risk ratio [RR] 1.42 per 10 years, 95% confidence interval [CI] 1.32-1.52 with enalapril; placebo: RR 1.18, 95% CI 1.12-1.25). Diuretic therapy was likewise associated with a greater risk of decreased renal function in the enalapril group (RR 1.89, 95% CI 1.70-2.08) than in the placebo group (RR 1.35, 95% CI 1.09-1.66). Conversely, enalapril had a relative renoprotective effect (RR 1.33, 95% CI 1.13-1.53) compared with placebo (RR 1.96, 95% CI 1.57-2.44) in patients with diabetes. A lower risk of renal impairment was seen in both groups with beta-blocker therapy (RR 0.70, 95% CI 0.57-0.85) and higher baseline ejection fraction (RR 0.93 per 5% increment, 95% CI 0.91-0. 96). CONCLUSIONS: Enalapril use caused a 33% increase in the risk of decreased renal function in patients with CHF. Diuretic use and advanced age increased this risk. Diabetes was associated with an increased risk of renal impairment in all patients with CHF, but this risk was reduced in the enalapril group compared with the placebo group. beta-Blocker therapy and higher ejection fraction were renoprotective in all patients regardless of therapy. PMID- 10539816 TI - Static and pulsatile blood pressure correlates of left ventricular structure and function in black and white young adults: the CARDIA study. AB - OBJECTIVE: To determine the unbiased relative strength of the association of static (systolic and diastolic) and pulsatile (pulse pressure) blood pressure components with left ventricular mass and function. BACKGROUND: Blood pressure is correlated with left ventricular mass; however, the unbiased relative strength of static and pulsatile blood pressure components with left ventricular mass and function is unknown in young adults. METHODS: Cross-sectional analyses of 3918 young adult participants at 4 community-based CARDIA clinical centers during 1990 and 1991. RESULTS: Left ventricular mass positively correlated (P <.01) with systolic, diastolic, and pulse pressure in all ethnicity-sex groups except for diastolic blood pressure in white men (P =.19). The association rank ordered as systolic blood pressure > pulse pressure > diastolic blood pressure except in white men, in whom pulse pressure and systolic blood pressure reversed positions in this hierarchy. Systolic blood pressure was the third and fourth strongest independent correlate of left ventricular mass in men and women, respectively. Body mass index, followed by height, was the strongest correlate of left ventricular mass in both sexes. Left ventricular wall thickness/chamber radius ratio positively correlated with diastolic and systolic blood pressure (women only) (P <.05) but not with pulse pressure. In all groups, stroke volume positively correlated (P <.05) with pulse pressure but was unrelated to static blood pressure measures, except for systolic blood pressure in black men. Left ventricular mass and the ventricular wall thickness/chamber radius ratio were greater in blacks compared with whites. CONCLUSIONS: Although systolic blood pressure was consistently the strongest unbiased blood pressure correlate of left ventricular mass, this relation varied by ethnicity and sex. Pulse pressure correlated with favorable left ventricular function and geometry, suggesting an opposite meaning to the ominous prognosis of wide pulse pressure in hypertensive, older adults. PMID- 10539817 TI - Axial movement of the intravascular ultrasound probe during the cardiac cycle: implications for three-dimensional reconstruction and measurements of coronary dimensions. AB - BACKGROUND: Motion of the intravascular ultrasound (IVUS) probe within the coronary artery from cardiac contraction may result in artifacts during 3 dimensional ultrasound image reconstruction and inaccurate measurements of coronary compliance. The purpose of this study was to establish whether longitudinal movement of the IVUS transducer in the coronary artery occurs and to quantify such motion. METHODS: In 31 patients we positioned IVUS transducers at 59 coronary branch points: 41 in the left anterior descending coronary artery, 11 in the left circumflex coronary artery, and 7 in the right coronary artery. In each image sequence the branching vessel oscillated in and out of the imaging plane during the cardiac cycle, confirming longitudinal transducer movement. The extent of movement was estimated by IVUS from the dimension of the branch vessel traversed. In addition, angiographic visualization and measurement of IVUS probe motion was performed at 17 branch points in 12 patients. RESULTS: Average longitudinal transducer movement as measured by IVUS was 1.50 +/- 0.80 mm (n = 46, range 0.5 to 5.5 mm). Because IVUS could not account for probe motion that exceeded the vessel branch diameter, the values obtained represent minimum movement. Average probe motion as assessed by cineangiography in a subset of 12 patients was 2.43 +/- 1.42 mm (range 0.57 to 6.56 mm). CONCLUSIONS: This study establishes that longitudinal movement of IVUS transducers within coronary vessels occurs during the cardiac cycle. Because documented extent of motion may be sufficient to influence analysis, IVUS images are best obtained with electrocardiographic gating. PMID- 10539818 TI - Relation between exercise and dobutamine stress-induced wall motion abnormalities and severity and location of stenosis in single-vessel coronary artery disease. AB - BACKGROUND: Quantitative coronary angiography has been shown to allow accurate assessment of coronary stenosis. Exercise and dobutamine stress echocardiography both are established methods for assessing the functional importance of coronary stenosis. The relation, however, between exercise and dobutamine stress-induced wall motion abnormalities and the severity and location of stenosis remains controversial. METHODS AND RESULTS: Thirty patients with single-vessel coronary artery disease with >/=50% minimal luminal reduction and stable angina participated in the study. Severity of coronary artery stenosis was assessed by means of computed angiography. During peak exercise echocardiography 23 patients had wall motion abnormalities and 7 did not. A positive test result was associated with severity of stenosis >/=80% for 65% of stenoses (P <.05 versus severity of stenosis <80%) and with a proximal location of 94% of stenoses (P <.01 versus middle and distal stenoses). A significant correlation was found between area of stenosis and difference in wall motion score between rest and peak exercise (r = 0.53, P <.01). The proportion of positive exercise stress was greater among stenoses with severity <80% (62% versus 46% dobutamine stress, P <.05). During dobutamine stress echocardiography 18 patients had wall motion abnormalities and 12 patients did not. A positive test result was associated with severity of stenosis >/=80% in 72% of stenoses (P <.05 versus severity of stenosis <80%) and with a proximal location in 81% of stenoses (P <.01 versus middle and distal stenoses). A weak correlation was found between area of stenosis and difference in wall motion score between rest and peak dobutamine stress (r = 0.37, P <.05). CONCLUSIONS: Among patients with single-vessel coronary artery disease, positive stress echocardiographic test results usually are associated with proximal >/=80% stenosis. Patients with <80% stenoses are more likely to have a positive exercise stress test result than a positive dobutamine stress test result. PMID- 10539819 TI - Role of transesophageal echocardiography in assessing diastolic dysfunction in a large clinical practice: a 9-year experience. AB - BACKGROUND: Two-dimensional transthoracic echocardiography with respiratory monitoring has been used to characterize diseases that impair diastolic function. Transesophageal echocardiography (TEE) has emerged as a complementary technique to evaluate patients with these diseases. The purpose of this study was to evaluate in a large clinical practice the utility of TEE with respiratory monitoring for classification of patients with diastolic dysfunction. METHODS: Over a 9-year period TEE was used to examine 192 patients referred to an echocardiography laboratory for additional evaluation of abnormal diastolic function. We performed pulsed-wave Doppler TEE of the left ventricular inflow and pulmonary veins and respiratory monitoring to categorize patients as showing restrictive physiologic features, constriction with or without effusion, mixed constriction and restriction, abnormal relaxation, pseudonormalization, large pericardial effusion or tamponade, or normal diastolic function. RESULTS: Patients with diastolic dysfunction underwent 3% of the total number of transesophageal studies conducted during the study period. Among the 192 patients referred for TEE, abnormal diastolic function was found in 181 (94%); 11 (6%) had normal diastolic function. Seventy-one (39%) of the 181 patients had restrictive physiologic features. Constrictive pericarditis was found in 54 (30%) of the patients and was confirmed for all 31 patients who underwent pericardiectomy. Mixed constriction and restriction was present in 21 (12%) of the patients. The other 35 patients (19%) had abnormal relaxation, pseudonormalization, or large pericardial effusion or tamponade. The cause of diastolic dysfunction was idiopathic for 32% of the patients, previous cardiac operation for 26%, cardiac amyloidosis for 23%, radiation therapy for 11%, and hypertension or advanced ischemic heart disease for 8%. CONCLUSION: Two-dimensional and Doppler TEE with respiratory monitoring is useful in categorizing patients with impaired diastolic function, primarily into those with restrictive physiologic features or constrictive pericarditis. PMID- 10539820 TI - Heart rate variability associated with particulate air pollution. AB - BACKGROUND: Epidemiologic studies have linked fine particulate air pollution with cardiopulmonary mortality, yet underlying biologic mechanisms remain unknown. Changes in heart rate variability (HRV) may reflect changes in cardiac autonomic function and risk of sudden cardiac death. This study evaluated changes in mean heart rate and HRV in human beings associated with changes in exposure to particulate air pollution. METHODS: Repeated ambulatory electrocardiographic monitoring was conducted on 7 subjects for a total of 29 person-days before, during, and after episodes of elevated pollution. Mean HR, the standard deviation of normal-to-normal (NN) intervals (SDNN), the standard deviation of the averages of NN intervals in all 5-minute segments of the recording (SDANN), and the square root of the mean of squared differences between adjacent NN intervals (r-MSSD) were calculated for 24-hour and 6-hour time segments. Associations of HRV with particulate pollution levels were evaluated with fixed-effects regression models. RESULTS: After controlling for differences across patients, elevated particulate levels were associated with (1) increased mean HR, (2) decreased SDNN, a measure of overall HRV, (3) decreased SDANN, a measure that corresponds to ultralow frequency variability, and (4) increased r-MSSD, a measure that corresponds to high-frequency variability. The associations between HRV and particulates were small but persisted even after controlling for mean HR. CONCLUSIONS: This study suggests that changes in cardiac autonomic function reflected by changes in mean HR and HRV may be part of the pathophysiologic mechanisms or pathways linking cardiovascular mortality and particulate air pollution. PMID- 10539821 TI - Comparison of past versus recent physical activity in the prevention of premature death and coronary artery disease. AB - BACKGROUND: People who are physically active live longer, but it is unclear whether this is because of physical activity in the distant or more recent past. METHODS: We assessed activity levels in 5209 men and women in the Framingham Heart Study from 1956 to 1958 and again from 1969 to 1973. We included individuals who were alive and without cardiovascular disease in the period 1969 to 1973. The primary outcome was death from all causes during the 16 years after the 1969 to 1973 assessment. Secondary outcomes were incidence and mortality rate of cardiovascular disease. We used Cox proportional hazards regression to calculate the relative risk of being sedentary, both unadjusted and controlling for smoking, weight, systolic blood pressure, cholesterol, glucose intolerance, left ventricular hypertrophy, chronic obstructive pulmonary disease, and cancer. RESULTS: The overall 16-year mortality rate was 37% for men and 27% for women. When both distant and recent activity levels were included along with major cardiovascular disease risk factors, for recent activity the most active tertile had lower overall mortality rate than the least active tertile for men (risk ratio 0.58, 95% confidence interval, 0.43-0.79) and women (risk ratio 0.61, 95% confidence interval, 0.45-0.82). For distant activity there was no difference in overall mortality rate between the most and least active tertiles either for men or for women. Adjusting for major cardiovascular disease risk factors had little effect on the results. CONCLUSIONS: The reduction in overall mortality rates is more associated with recent activity than distant activity. These results suggest that for sedentary patients, it may never be too late to begin exercising. PMID- 10539822 TI - Decreased soluble interleukin-6 receptor in patients with acute myocardial infarction. AB - BACKGROUND: The plasma level of interleukin-6 (IL-6), a proinflammatory cytokine, has been reported to be elevated in patients with acute myocardial infarction (AMI). In addition to a specific cell-surface IL-6 receptor, a soluble IL-6 receptor (sIL-6R) exists in plasma as an extracellular domain of glycoprotein 80. The pathophysiologic roles of IL-6 and sIL-6R in AMI are still unknown. METHODS AND RESULTS: We measured plasma levels of IL-6 and sIL-6R in 17 patients with AMI to evaluate changes over time at 11 points in the acute phase and compared them with parameters of inflammation, myocardial injury, and atherosclerosis. IL-6 showed a triphasic increase with peaks at 3 hours (276.2 +/- 50.0 pg/mL), 2 days (153.6 +/- 35.7 pg/mL), and 5 days (180.7 +/- 52.3 pg/mL) after the onset of AMI. sIL-6R had biphasic dips at 12 hours (31.1 +/- 4.1 ng/mL) and 3 days (29.9 +/- 1.5 ng/mL) after the onset on AMI. The time-dependent changes in IL-6 paralleled those in sIL-6R from onset to 5 days. Thereafter, the changes in IL-6 and sIL-6R varied; that is, IL-6 gradually decreased from 5 days to 4 weeks, whereas sIL-6R gradually increased from 5 days to 4 weeks. Significant positive correlations were observed between the absolute increase in IL-6 and the decrease in sIL-6R and the changes in white blood cell count, erythrocyte sedimentation rate, C reactive protein, creatine kinase, and lactic dehydrogenase. Neither IL-6 nor sIL 6R strongly correlated with parameters of coronary atherosclerosis. CONCLUSIONS: These results demonstrate that IL-6 and sIL-6R are associated with the processes of inflammation and myocardial injury during the acute phase of AMI. PMID- 10539823 TI - Dynamic exercise normalizes resting blood pressure in mildly hypertensive premenopausal women. AB - BACKGROUND: Dynamic exercise acutely and transiently lowers resting blood pressure in hypertensive men and is termed postexercise hypotension (PEH). We examined 18 premenopausal women (7 hypertensive and 11 normotensive) to determine if PEH occurs in women and to elucidate possible hemodynamic and hormonal mechanisms. METHODS AND RESULTS: Patients wore an ambulatory blood pressure monitor throughout the day after 40 minutes of a rest sham session and 40 minutes of cycle exercise, of which 30 minutes was performed at 60% of maximal oxygen consumption. Cardiac output and total systemic vascular resistance were determined by Doppler echocardiography before and 15 minutes after sham and exercise. Catecholamines, plasma renin activity, and beta-endorphin were measured over this same period. PEH occurred only in the hypertensive women. Systolic, diastolic, and mean arterial blood pressure decreased in the hypertensive women by a mean of 9.5 +/- 2. 8 mm Hg (P <.01), 6.7 +/- 2.4 mm Hg (P <.05), and 7.7 +/- 2.4 mm Hg (P <.05), respectively, for up to 7 hours after versus before exercise, whereas blood pressure was similar in the normotensive women (P >.05). After exercise, total systemic vascular resistance was lower (P <.01), and cardiac output, catecholamines, and plasma renin activity were greater (P <.01) than before exercise in both groups of women. CONCLUSIONS: PEH was observed for up to 7 hours after exercise in mildly hypertensive women and was not explained by the hemodynamic and hormonal adjustments that occurred after exercise. The magnitude and duration of PEH may be sufficient to normalize the blood pressure of certain hypertensive women throughout most of the day. PMID- 10539824 TI - Elevated plasma immunoreactive leptin levels preexist in healthy offspring of patients with essential hypertension. AB - BACKGROUND: Plasma leptin levels and plasma insulin levels have been found to be elevated in patients with essential hypertension (EH) and have been suggested to be components of the metabolic syndrome. Increased heart rate (HR) may predict the development of EH in normal or borderline-hypertensive individuals. The aim of our study was to test the hypothesis that elevated plasma leptin and insulin levels as well as systolic blood pressure (SBP) and diastolic blood pressure (DBP) and increased resting HR preexist in the healthy offspring of patients with EH. METHODS AND RESULTS: Twenty-six (12 male, 14 female) healthy offspring of hypertensive patients, mean age 16 +/- 2.5 years and body mass index (BMI) of 21.5 +/- 2.8 kg/m(2) (group A), and 30 (14 male, 16 female) healthy offspring of normotensive patients, mean age 17 +/- 2.3 years and BMI of 21.9 +/- 2.4 kg/m(2) (group B), were studied. (The two groups were matched for sex, age, and BMI). Mean SBP, DBP, resting HR, plasma leptin, and plasma insulin levels (radioimmunoassay method) were determined in the whole study population. Mean SBP, DBP, and resting HR were significantly higher in group A than in group B (120 +/- 12 vs 112 +/- 9.5 mm Hg, 77 +/- 9 vs 72 +/- 7 mm Hg, 79 +/- 8 vs 75 +/- 5 beats/min, P <.01, P <.05, and P <.05, respectively). Plasma leptin and insulin levels were significantly higher in group A than in group B (9 +/- 5.06 vs 5.6 +/ 2.5 ng/mL and 20.11 +/- 11.3 vs 14.8 +/- 5.2 microIU/mL, P <.01 and P <.05, respectively). CONCLUSIONS: Our findings support the hypothesis that hyperleptinemia, hyperinsulinemia, and elevated blood pressure and resting HR preexist in the healthy offspring of patients with EH. PMID- 10539825 TI - Lack of effect of recent alcohol consumption on the course of acute myocardial infarction. AB - BACKGROUND AND OBJECTIVE: Alcohol has marked effects on hemodynamic and hemostatic variables that might alter the presentation of acute myocardial infarction that follows its use. We sought to determine whether recent alcohol consumption alters the course or complications of acute myocardial infarction. METHODS: In the Determinants of Myocardial Infarction Onset Study, we performed chart reviews and face-to-face interviews with 2161 patients who did not receive thrombolytic therapy. We assessed alcohol use before infarction, peak creatine kinase levels (1043 patients), electrocardiographic interpretations (1408 patients), and the presence of ventricular arrhythmias or congestive heart failure (all patients). RESULTS: Among the 2161 patients, 399 (18.5%) drank alcohol within 24 hours before myocardial infarction. We found no significant difference in mean peak creatine kinase level between those who had recently used alcohol and those who had not in an adjusted comparison (-6.1% difference; 95% confidence interval [CI] -20.3%-10.7%; P =.46). We also found no differences in adjusted risk for Q-wave infarction, congestive heart failure, or ventricular arrhythmias (odds ratios 1.03 [95% CI, 0.73-1.45; P =.88], 1.01 [95% CI, 0.67 1.54; P =.95], and 1.04 [95% CI, 0.66-1.65; P =.86]). Categorization of the duration since last alcohol use into 6-hour intervals revealed no trends between time since last use of alcohol and any of these outcomes. CONCLUSIONS: Recent alcohol use is not associated with changes in infarct size or risk for Q-wave infarction, congestive heart failure, or ventricular arrhythmia among this population. PMID- 10539826 TI - Improved detection of posterior myocardial wall ischemia with the 15-lead electrocardiogram. AB - BACKGROUND: A routine 12-lead electrocardiogram is commonly obtained to evaluate for possible acute myocardial infarction during the initial screening of patients with chest discomfort. Posterior myocardial infarction is commonly missed because it is not usually visible in the standard leads. In this study, we compared the sensitivity and specificity of posterior chest leads (V(7), V(8), and V(9)) and 12-lead electrocardiography in detecting posterior injury pattern during single vessel percutaneous transluminal coronary angioplasty. METHODS AND RESULTS: Three posterior chest leads in addition to the routine 12-lead electrocardiogram were monitored simultaneously during single-vessel percutaneous transluminal coronary angioplasty of the right, circumflex, and left anterior descending coronary arteries in a total of 223 patients. Posterior injury patterns (95%) were detected mostly during circumflex coronary occlusion. Posterior leads were able to detect injury pattern in 49% (36 of 74) of patients, whereas the 12-lead electrocardiogram was able to detect only 32% (P <.04). When all 15 leads were used to detect all ST elevations, sensitivity increased to 57%, with a specificity of 98% for the circumflex coronary artery. If maximal ST depressions in leads V(2) to V(3) are considered to be from posterior myocardial injury, then the overall sensitivity is increased to 69%. CONCLUSIONS: Posterior leads significantly increased the detection of posterior injury pattern compared with the standard 12-lead electrocardiogram. Using all 15 leads significantly further improved the detection of circumflex coronary-related injury pattern over the standard 12-lead electrocardiogram. PMID- 10539827 TI - Transcatheter closure of atrial-septal defects and patent foramen ovale in adults: optimal anatomic adaptation of occlusion device. AB - BACKGROUND: For transcatheter closure of atrial-septal defects, different occlusion systems are available. The purpose of this study was to examine the clinical feasibility of the ASD Occlusion System (ASDOS, Dr Osypka GmbH, Grenzach Wyhlen, Germany) and to evaluate the short- and long-term results. METHODS AND RESULTS: The study was composed of 20 consecutive patients with atrial-septal secundum defect (n = 13) or patent foramen ovale (n = 7). The device implantation was successful in all patients. For optimal closure of the defect, left atrial and right atrial umbrellas of different sizes were required in 10 of 20 patients. No major short- or long-term complications occurred. During the mean follow-up period of 13.9 +/- 5 months, 5 strut fractures without dislocation were observed, and in 8 (40%) of 20 patients transesophageal echocardiography revealed a small residual shunt. CONCLUSION: The ASDOS double umbrella system is suitable for transcatheter closure of interatrial defects in selected patients. This system showed a high procedural safety and has the unique advantage of individual adaptation of the occluding device on the defect anatomy that results in high closure effectiveness. PMID- 10539828 TI - Long-term invasive and noninvasive results of percutaneous balloon pulmonary valvuloplasty in children, adolescents, and adults. AB - BACKGROUND: Short-term and mid-term results of percutaneous balloon pulmonary valvuloplasty (BPV) are well known. However, data documenting long-term effectiveness of BPV are scarce. METHODS AND RESULTS: The long-term results of 62 patients were assessed by catheterization and Doppler echocardiography 1 to 10 years (mean 6.4 +/- 3.4) after BPV. Mean age of the patients was 13.5 +/- 10.5 years (range 9 months to 44 years). Twenty patients were 16 years of age or older. Right ventricular peak systolic pressure was systemic or suprasystemic in 72% of patients. A double-balloon technique was used in 29 patients. The balloon to-pulmonary valve diameter ratio was 1.4 +/- 0.38 (range 1 to 1.8). Total systolic transpulmonary pressure gradient in excess of 50 mm Hg in all patients before BPV decreased from 98 +/- 40 to 32 +/- 23 immediately after BPV and to 19 +/- 9 mm Hg at follow-up (P <.001). Infundibular gradient increased from 8 +/- 10 to 14 +/- 24 mm Hg after BPV and fell to 1 +/- 4 mm Hg at follow-up (P <.01). In 16 patients it was >/=20 mm Hg and virtually disappeared spontaneously in all at follow-up. The valvar gradient fell from 93 +/- 39 to 19 +/- 11 (P <.001) and was 18 +/- 9 mm Hg at follow-up. It remained unchanged in 3 patients (range 36 to 45 mm Hg). In 3 (4.8%) other patients, a new gradient >35 mm Hg developed that was >/=50 mm Hg in all 3. Among 5 patients having dysplastic valves, 3 had a gradient >35 mm Hg. There were no predictors of a gradient >35 mm Hg at long-term follow up by univariate or multivariate Cox proportional hazards analysis. Mild to moderate pulmonary regurgitation was present in 39% of patients. On electrocardiography, right ventricular hypertrophy decreased significantly in 90% of patients. CONCLUSIONS: BPV as a treatment of typical pulmonic valve stenosis produces excellent long-term results. Restenosis is rare (4.8%) and occurs more frequently in patients with dysplastic valves. There is a constant spontaneous regression of associated infundibular obstruction. PMID- 10539829 TI - Hemodynamic effects of a single oral dose of enalapril among children with asymptomatic chronic mitral regurgitation. AB - BACKGROUND: Angiotensin-converting enzyme inhibitors have been shown to have beneficial effects in the short- and long-term treatment of adult patients with chronic mitral regurgitation. The safety and efficacy of such treatment have not been established for children. The objective of this study was to assess the effect of the angiotensin-converting enzyme inhibitor enalapril on the severity of valvar mitral regurgitation and the systolic performance of overloaded left ventricle of children. METHODS: Ten patients 3 to 16 years of age (mean age 9.6 +/- 3.8 years) with moderate to severe chronic mitral insufficiency were examined by means of Doppler echocardiography before and 2 hours after receiving a single oral dose of enalapril (0.40 mg/kg). Effective regurgitant orifice area, regurgitant volume and fraction, left ventricular end-diastolic and end-systolic volumes indexed for body surface area, left ventricular pump function (total ejection fraction), left ventricular contractility (stress-adjusted velocity of shortening) and afterload (peak systolic and end-systolic circumferential wall stress), and systemic vascular resistance were calculated before and after treatment. RESULTS: The following values decreased significantly compared with baseline values: effective regurgitant orifice area (36.2 +/- 17.4 versus 25.9 +/ 16.5 mm(2), P =.00008), regurgitant volume (53.6 +/- 27.4 versus 36.1 +/- 24.5 mL, P =.0002), regurgitant fraction (56.7 +/- 14.5% versus 39.9 +/- 17.0%, P =. 0009), left ventricular end-diastolic volume indexed for body surface area (81.3 +/- 17.4 versus 76.1 +/- 16.1 mL/m(2), P =.005), left ventricular end-systolic volume indexed for body surface area (26.7 +/- 9.1 versus 22.6 +/- 8.9 mL/m(2), P =.02), afterload (peak systolic circumferential wall stress 135.8 +/- 15.3 versus 123.5 +/- 19.7 g/cm(2), P =.005; end-systolic circumferential wall stress 57.8 +/ 12.4 versus 48.3 +/- 12.8 g/cm(2), P =.005), and systemic vascular resistance (2012.2 +/- 536.1 versus 1622.7 +/- 389 dyne. sec. cm(-5), P =.005). Left ventricular pump function increased (total ejection fraction 67.6 +/- 5.7% versus 71.7 +/- 6.5%, P =. 005) without significant changes in left ventricular contractility (stress-adjusted velocity of shortening -0.35 +/- 0.8 versus -0.21 +/- 1.3 SD, P not significant). CONCLUSIONS: The data showed that for pediatric patients single-dose treatment with oral enalapril reduces the severity of mitral regurgitation and improves left ventricular loading conditions and systolic performance without impairment of myocardial contractility. Persistence of these unloading effects in long-term therapy might slow the evolution of left ventricular dysfunction caused by overload-induced myocardial damage and possibly delay the time at which surgical repair or replacement of the mitral valve becomes necessary. PMID- 10539830 TI - Intracoronary surface changes after Palmaz-Schatz stent implantation: serial observations with coronary angioscopy. AB - BACKGROUND: The objective of this study was to evaluate the appearance of the intraluminal surface after Palmaz-Schatz stent implantation by using coronary angioscopy. METHODS AND RESULTS: Coronary angioscopy was performed immediately after stenting and at 1, 3, and 6 months later in 43 patients with 45 lesions. The presence or absence of red thrombus and/or dissection and the extent of neointimal coverage of the stent struts were analyzed. Immediately after stenting, red thrombus and dissection were observed in 9 (41%) and 12 (56%) of 22 lesions, respectively, and these rates decreased with time. Complete coverage of the stent struts by smooth white neointima was observed in 55% of 11 lesions at 1 month and in 80% of 21 lesions at 3 months. However, incomplete neointimal coverage was seen in 3 lesions at both 3 and 6 months. CONCLUSIONS: In human coronary arteries, neointimal coverage of an implanted Palmaz-Schatz stent may take as long as 6 months or more. PMID- 10539831 TI - Impact of tranilast on restenosis after coronary angioplasty: tranilast restenosis following angioplasty trial (TREAT). AB - BACKGROUND: Tranilast is an antiallergic drug that suppresses the release of cytokines such as platelet-derived growth factor, transforming growth factor beta1, and interleukin-1beta and prevents keloid formation after skin injury. Treatment with this drug reduced the restenosis rate after percutaneous transluminal coronary angioplasty in a preliminary study. METHODS AND RESULTS: We conducted a multicenter, randomized, double-blind, placebo-controlled trial. A total of 255 patients with 289 lesions were randomly assigned to treatment with the oral administration of 600 mg/d tranilast, 300 mg/d tranilast, or a placebo for 3 months after successful angioplasty. Angiographic follow-up was done at 3 months, and a clinical follow-up examination was performed at 12 months. Two hundred ten (72.7%) lesions of 188 (73.7%) of the patients met the criteria and were eligible for the assessment of restenosis. The restenosis rates defined as >/=50% loss of the initial gain were 14.7% in the 600 mg/d tranilast group, 35.2% in the 300 mg/d tranilast group, and 46.5% in the placebo group (P <. 0001 for 600 mg/d tranilast vs placebo). The restenosis rates defined as percent diameter stenosis of >/=50% at follow-up were 17. 6% in the 600 mg/d tranilast group, 38.6% in the 300 mg/d tranilast group, and 39.4% in the placebo group (P =.005 for 600 mg/d tranilast vs placebo). CONCLUSIONS: The oral administration of 600 mg/d of tranilast for 3 months markedly reduced the restenosis rate after percutaneous transluminal coronary angioplasty. PMID- 10539832 TI - Effect of institutional volume and academic status on outcomes of coronary interventions: the IMPACT-II experience. AB - BACKGROUND: Rates of morbidity and mortality after interventional procedures are reported to be inversely associated with institutional volume. METHODS: This study assessed both procedural volume and academic status at the 82 US centers that participated in the IMPACT-II trial. Interventional volumes at the sites ranged from 90 to 3300 cases per year. Patients were randomly assigned to a platelet glycoprotein IIb/IIIa inhibitor (eptifibatide) or placebo during procedures done by experienced operators. The primary end point was the composite of death, myocardial infarction, nonelective repeat coronary intervention, or nonelective coronary artery bypass surgery at 30 days, or placement of an intracoronary stent for abrupt closure during the initial procedure. RESULTS: Baseline patient characteristics and median length of stay were similar between the academic and nonacademic centers. In univariable analysis, volume as a continuous variable had a nonlinear relation with the incidence of the composite end point, with better outcomes noted at the highest volume institutions. Academic status did not predict outcome. When added to a predictive model that contained the variables unstable angina, weight, prior coronary artery bypass grafting, heart rate, and platelet count, procedural volume continued to be associated with the composite outcome (P =.04). CONCLUSIONS: We conclude that among hospitals participating in this trial, there is a nonlinear relation between annual interventional volume and outcomes. This relation is complex, involving variations in periprocedural infarction rates and additional, undefined institutional differences (other than academic status) that result in differences in procedural outcome. PMID- 10539833 TI - Multivessel coronary artery bypass surgery without cardiopulmonary bypass. AB - BACKGROUND: There is limited experience with complete myocardial revascularization on a beating heart. Using a mechanical stabilization system, we sought to determine if complete coronary revascularization is feasible without cardiopulmonary bypass and what the short-term clinical outcome would be. METHODS: From February through September 1998, 26 patients underwent complete myocardial revascularization with Medtronics Octopus Tissue Stabilization System. Mean age for the group was 62 +/- 7 years (range 48 to 78 years); 11% had prior interventions. Mean preoperative ejection fraction was 49% +/- 14% (range 20% to 66%); 38% of operations were performed urgently. The mean number of vessels grafted was 3.0 +/- 0.9 (range 1 to 5 grafts/patient). In 96% of patients, at least one arterial graft was used. Fifteen percent of patients had 2 or more arterial grafts. In 58% of patients, a branch of circumflex coronary artery was bypassed. RESULTS: The median time to extubation was 2 hours (range 0 to 37 hours). None of the patients had perioperative myocardial infarction, cerebrovascular accident, or renal failure requiring dialysis. The 30-day survival rate was 100%. Angiographic follow-up was not available. At a mean follow-up period of 3.8 +/- 2.9 months, all patients remained free of angina and none has required cardiac reintervention. CONCLUSIONS: Complete myocardial revascularization on a beating heart can be achieved with the currently available stabilization systems and is associated with low perioperative complications and satisfactory short-term clinical outcomes. The long-term outcomes and graft patency remain to be determined. PMID- 10539834 TI - Persistent T-wave changes after radiofrequency catheter ablation of an accessory connection (Wolff-parkinson-white syndrome) are caused by "cardiac memory". AB - BACKGROUND: The purpose of this study was to determine the incidence and origin of T-wave changes after ablation of an accessory atrioventricular connection (AC), which could either be a sign of damage to the coronary circulation or a result of persistent abnormal repolarization secondary to previously abnormal ventricular activation ("cardiac memory"). METHODS AND RESULTS: Ninety of 107 consecutive patients (33 women and 57 men, mean age 36 +/- 5 years) undergoing successful catheter ablation of an AC were studied. Patients with bundle branch block or more than 1 AC were excluded. Sixty-four patients had manifest preexcitation (group 1) and 26 had a concealed AC (group 2). Immediately after loss of preexcitation, 38 (59%) patients with a manifest AC showed T-wave abnormalities. In contrast, none of the patients with a concealed AC had T-wave abnormalities after ablation (P <.05). The T-wave changes (1) did not correlate with the number or duration of energy applications or with markers of tissue injury; (2) correlated with the location of the AC and the degree of preexcitation, respectively; and (3) completely resolved over a period of weeks to months. None of the patients had recurrence of preexcitation or tachycardia during a mean follow-up of 16 +/- 7 months. CONCLUSIONS: T-wave changes after ablation are most likely caused by "cardiac memory" and are not a sign of myocardial or coronary injury. PMID- 10539836 TI - Reduced risk for thromboembolism in atrial fibrillation and mitral regurgitation. PMID- 10539835 TI - Effect of dofetilide on survival in patients with supraventricular arrhythmias. AB - OBJECTIVES: This study compared survival between patients taking dofetilide and patients taking placebo in a pooled analysis of randomized clinical trials of patients with supraventricular arrhythmias. BACKGROUND: Clinical trials of antiarrhythmic drugs used to treat supraventricular arrhythmias rarely include enough patients to assess whether the drug being tested has an effect on survival. Pooling data from many trials provides useful information on safety. METHODS: Data from randomized clinical trials of antiarrhythmic drug therapy of supraventricular arrhythmias were pooled to assess the effect on survival of dofetilide (n = 1346) compared with placebo (n = 677) in this patient population. RESULTS: The unadjusted hazard ratio for risk of death (dofetilide/placebo) was 1.4 with 95% confidence interval 0.4-5.1. After adjusting for effects of arrhythmia diagnosis, age, sex, and structural heart disease, the hazard ratio was 1.1 (confidence interval 0.3-4.3). CONCLUSIONS: The pooled survival analysis provided reassurance regarding the safety of dofetilide in patients with supraventricular arrhythmias. PMID- 10539837 TI - C-reactive protein and coronary disease. PMID- 10539839 TI - Atherosclerosis is an inflammatory disease. PMID- 10539840 TI - Stable versus unstable atherosclerosis: clinical aspects. PMID- 10539841 TI - Diet and atherosclerosis. PMID- 10539842 TI - Role of infections in atherosclerosis. AB - A growing amount of epidemiologic, experimental, and clinical evidence has linked infection as a risk factor to variousatherosclerotic diseases including acute myocardial infarction and cerebral infarction. Bacteremic infections with and without endocarditis carry a high risk for both stroke and acute myocardial infarction. During the last decade, chronic bacterial infections such as Chlamydia pneumoniae and dental infections have been associated as risk factors for various atherosclerotic diseases. These chronic bacterial infections are risk factors for acute cardiovascular events, but they may also have some role in the etiopathogenesis of atherosclerotic process itself. There are many known mechanisms that might explain the observed association of infection and atherosclerotic diseases, but it is probable that these mechanisms are complex and multifactorial and probably differ from infection to infection and from patient to patient. Infection theory is by no means against classic risk factor theory in the etiopathogenesis of atherosclerosis. Infection may also act as a synergistic risk factor together with classic risk factors in the development of various atherosclerotic diseases. PMID- 10539843 TI - Coronary heart disease, Helicobacter pylori, dental disease, Chlamydia pneumoniae, and cytomegalovirus: meta-analyses of prospective studies. PMID- 10539844 TI - Functionality of specific immunity in atherosclerosis. PMID- 10539845 TI - Autoimmunity and atherosclerosis. PMID- 10539846 TI - Molecular mediators of arterial inflammation: a role for microbial products? PMID- 10539847 TI - Viruses and atherosclerosis: a critical review of the epidemiologic evidence. PMID- 10539848 TI - Viral activation of the coagulation cascade. PMID- 10539849 TI - Atherosclerosis induced by infection with Marek's disease herpesvirus in chickens. AB - BACKGROUND: This research was suggested after crystals that we observed in herpesvirus-infected cell cultures were identified as cholesterol. Other reports and the development of defined reagents led us to select the use of Marek's disease herpesvirus (MDV) infection of chickens to demonstrate a potential role of herpesviruses in the pathogenesis of atherosclerosis. Available for our use were a clone-purified strain of MDV of known virulence, genetically selected, specific pathogen-free chickens, and appropriate isolation facilities to design controlled experiments to fulfill Koch's postulates. METHODS AND RESULTS: Experiments were performed to test the roles of both MDV and dietary cholesterol in atherosclerosis. The birds were examined 7 months after MDV infection with and without cholesterol feeding for gross and microscopic arterial lesions. Atherosclerotic lesions were found only in infected normocholesterolemic or hypercholesterolemic birds. The character and distribution of these lesions closely resembled those found in the chronic human arterial disease. Atherosclerotic lesions were not found in uninfected birds even if the birds were hypercholesterolemic. CONCLUSIONS: Evidence was obtained from other experiments that after MDV infection, cholesterol and cholesteryl esters accumulated in cell cultures and in atherosclerotic lesions. These changes were associated with altered enzymatic activity of the cholesterol synthesis cycle. Immunization with turkey herpesvirus vaccine or SB-1 vaccine prevented atherosclerotic lesions. PMID- 10539850 TI - Coronary artery disease after heart transplantation and its relation to cytomegalovirus. AB - The most common cause of death and retransplantation after heart transplantation is a rapidly progressive, obliterative vascular disease involving the coronary arteries, termed cardiac allograft vasculopathy (CAV). Most believe that this is a form of chronic rejection. Several clinical series have suggested an association between cytomegalovirus and CAV. Rat cytomegalovirus enhances the development of CAV in rat heterotopic heart or aortic transplantation models. The mechanism(s) by which cytomegalovirus might have an impact on the severity of chronic rejection include the augmentation of vascular growth factors, the alteration in the alloimmune response directly or the alteration of cytokines and cell adhesion molecules, enhancing cellular and humoral interactions. We previously reported that the infection of smooth muscle cells by cytomegalovirus resulted in the alteration of major histocompatibility complex class I molecules on the smooth muscle cell surface. In a subsequent report we demonstrated that a sublethal inoculum of cytomegalovirus produced no cytopathology in smooth muscle cells yet had the same viral burden as fibroblasts, which demonstrated cytopathology. The identical effects on major histocompatibility complex class I were observed in smooth muscle cells, and cytokine gene transcription was altered, favoring a proinflammatory milieu. These and most in vitro studies are carried out with the use of traditional laboratory strains of cytomegalovirus. We have subsequently demonstrated major genotypic differences between laboratory and clinical strains of cytomegalovirus that are associated in differences in biological activity in vitro. These include differences in tropism for vascular cells, differences in cell surface antigen expression, and differences in mesenchymal growth factor gene expression. All of these may have important implications with regard to associating cytomegalovirus with CAV. PMID- 10539851 TI - Cytomegalovirus is involved in vascular pathology. PMID- 10539852 TI - Potential role of cytomegalovirus in the pathogenesis of restenosis and atherosclerosis. PMID- 10539853 TI - Enteroviruses and myocardial infarction. AB - BACKGROUND: Several lines of evidence suggest that some microbial infections are involved in the pathogenesis of atherosclerosis. The aim of the studies referred to here was to evaluate the possible role of enteroviral infections as potential risk factors for cardiac events. METHODS AND RESULTS: The association of enterovirus-specific antibodies and cardiac events was analyzed in 3 large, prospective population studies with the nested case-control design. Stored sera collected at the study baseline were tested for enterovirus group-specific immunoglobulin G antibodies. The study samples from the North Karelia Project, the Helsinki Heart Study, and the Mobile Clinic Health Service were composed of 183, 241, and 276 men with myocardial infarction, respectively, and their matched control patients. In 2 of the 3 studies, male cases without evidence of heart disease at the baseline had significantly higher levels of antibodies to enteroviruses than their matched control patients. High enterovirus antibody level was found to be a definite independent risk factor for future cardiac events. In the North Karelia study the risk was high in men aged 25 to 49 years, whereas in the Mobile Clinic Health Service study the risk was particularly strong in those with low levels of serum cholesterol. No association with high levels of enterovirus antibodies and cardiac events was seen in the Helsinki Heart Study, which consisted of hypercholesteremic men. CONCLUSIONS: If a high level of enterovirus group-specific antibodies can be considered as a sign of frequent enterovirus infections in the past, these studies suggest that enterovirus infections increase the risk of myocardial infarction at least in normocholesteremic men. PMID- 10539854 TI - Cytomegalovirus vaccine. AB - Congenital cytomegalovirus disease is an unsolved public health problem, unlikely to be solved by means other than immune prophylaxis. Development of a vaccine has been hampered by low awareness of the problem, which is caused by the often delayed detection of abnormalities after birth. Nevertheless, cytomegalovirus vaccine development is active. An attenuated, live vaccine has been studied extensively, and an improved strain may result from genetic manipulation. An immunogenic viral glycoprotein (gB) vaccine is currently in clinical trial to determine if antibodies alone will be protective. The idea of a combined vaccine has been proposed, in which a canarypox recombinant containing several cytomegalovirus genes is used both to generate cellular immunity and to prime for augmented antibody responses to the viral glycoprotein. Finally, DNA plasmids containing cytomegalovirus genes are being investigated for their utility as vaccines. PMID- 10539855 TI - Chlamydia pneumoniae and atherosclerosis: links to the disease process. AB - Chlamydia pneumoniae is an obligate intracellular prokaryotic human pathogen responsible for a significant portion of atypical pneumonia and associated with a variety of chronic sequelae, the most significant of which is atherosclerosis. The organism is endowed with several attributes that may contribute to the development of atherosclerotic lesions or promote tissue damage at the site of an existing lesion. Two key events that are directly involved in the atherogenic process include the development of foam cells from macrophages and the oxidation of lipoproteins at the site of lesion development. The former process allows for deposition of cholesterol-containing low-density lipoprotein (LDL) and the latter can contribute directly to tissue damage locally. We have hypothesized that C pneumoniae may interact with mononuclear phagocytes in ways that are consistent with the view that this organism contributes to atherosclerotic lesion development. We have demonstrated that the presence of C pneumoniae causes macrophage foam cell formation and lipid oxidation with murine and human cells cocultured in the presence of LDL. In addition, we have provided evidence that implicates 2 putative chlamydial virulence factors in the development of these pathologic processes. Chlamydial lipopolysaccharide has been shown to cause macrophages to develop into foam cells in the presence of LDL, and the 60-kDa chlamydial heat shock protein (cHsp60), a known pathogenesis-inducing protein, has been found to contribute to oxidation of LDL in the presence of macrophages. Work is currently underway to define mechanisms involved in these processes and to further refine the putative role of C pneumoniae in atherogenesis and atherosclerotic lesion development. PMID- 10539856 TI - Molecular biology of Chlamydia pneumoniae surface proteins and their role in immunopathogenicity. AB - BACKGROUND: The association of Chlamydia pneumoniae with the development of atherosclerosis is based on serology and on detection of C pneumoniae-specific DNA by polymerase chain reaction in the atheromas. METHODS AND RESULTS: Because the humoral immune response frequently recognizes epitopes present on the surface of the bacteria, we analyzed what components are present on the C pneumoniae surface. We identified a family of proteins, the GGAI or Omp4-15 proteins, of which at least 3 are present on the surface of C pneumoniae. We immunized rabbits with recombinant GGAI proteins and used these antibodies in immunofluorescence microscopy of experimentally infected mice. In lung sections, a massive infiltration with polymorph nuclear neutrophil cells was observed. In the bronchial epithelial cells, C pneumoniae inclusions were seen. Evidence was found of differential expression of the GGAI proteins. CONCLUSIONS: On the basis of surface localization, differential expression, and the fact that the proteins are recognized by the human humoral immune response, we speculate whether these proteins, in addition to the lipopolysaccharides, are of importance for the immunopathogenesis of C pneumoniae. PMID- 10539857 TI - Pathologic manifestation of Chlamydial infection. PMID- 10539858 TI - Epidemiology of Chlamydia pneumoniae in atherosclerosis. AB - Chlamydia pneumoniae is a common, ubiquitous respiratory tract agent causing, apart from upper respiratory tract infections, approximately 10% of all pneumonias worldwide. Antibody prevalence starts to rise early in life in the developing countries, but in industrialized countries they only begin to rise when the children start school. In early adulthood, antibody prevalence reaches approximately 50%, with men having greater prevalence than women. This prevalence rises toward old age. The pitfalls in seroepidemiologic studies associating C pneumoniae infection with various syndromes include the difficulties in testing for C pneumoniae antibodies, the necessity for careful choice of control patients, the frequently old age of the matched control patients, and the possible prevailing epidemic situation reflecting antibody saturation in control patients. However, more than 20 studies that used different serologic methods in different laboratories all over the world have established the association of C pneumoniae with atherosclerosis. Circulating immune complexes and immunoglobulin A antibodies appear to be the best markers of chronic infection in C pneumoniae seroepidemiologic studies. PMID- 10539859 TI - Interaction of Chlamydia pneumoniae infection with other risk factors of atherosclerosis. AB - BACKGROUND: Seroepidemiologic studies have provided information on the interaction of Chlamydia pneumoniae with other known risk factors of coronary heart disease. C pneumoniae infection appears to be more common in smokers than in nonsmokers, suggesting that smoking predisposes to the development of chronic C pneumoniae infection. METHODS AND RESULTS: In identical twins, C pneumoniae specific immunoglobulin A levels were found to be higher and cell-mediated immunity, measured as a lymphoproliferation response, lower in smoking twins than in their nonsmoking counterparts, suggesting that chronic C pneumoniae infections are common in smokers whose cell-mediated protective immunity appears to be lowered. Infections also may have an effect on lipid metabolism. Thus even in acute pneumonia caused by C pneumoniae, high-density lipoprotein (HDL) values are lower and triglyceride values higher than in pneumonia caused by viruses and other bacteria. Furthermore, chronic C pneumoniae infection has been associated with elevated triglyceride and lowered HDL levels in otherwise healthy Finnish men. We also have explored the combined effect of chronic C pneumoniae infection and markers of the metabolic syndrome (elevated body mass index, blood glucose and systolic blood pressure, and lowered HDL cholesterol) on the risk of cardiac events. The results suggest that chronic C pneumoniae infection enhances the effect of the metabolic syndrome on the risk of coronary heart disease. CONCLUSIONS: The known risk factors of coronary heart disease may be explained partly by their interaction with chronic C pneumoniae infection. Further studies are needed to elucidate the pathogenetic mechanisms underlying these interactions. PMID- 10539860 TI - In vitro infection and pathogenesis of Chlamydia pneumoniae in endovascular cells. AB - The strength of the epidemiologic and clinical associations of Chlamydia pneumoniae with atherosclerosis can be increased by the demonstration that C pneumoniae can initiate and sustain growth in human vascular cells as well as in animal models. To investigate the biological basis for the dissemination and proliferation of this organism in vascular cells, the in vitro growth of C pneumoniae was studied in 2 macrophage cell lines, peripheral blood monocyte (PBMC)-derived macrophages, human bronchoalveolar lavage (BAL) macrophages, several endothelial cell lines, and aortic artery smooth muscle cells. Five of 5 strains of C pneumoniae were capable of 3 passages in human U-937 macrophages and in murine RAW 246.7 macrophages. Titers were suppressed in both macrophage types with each passage as compared with growth in HEp-2 cells. Both human BAL macrophages and PBMC-derived macrophages were able to inhibit C pneumonia eafter 96 hours' growth. Eleven C pneumoniae strains were capable of replicating in normal human aortic artery-derived endothelial cells, umbilical vein-derived endothelial cells, and pulmonary artery endothelial cells. Infection in human aortic artery smooth muscle cells was also established for 13 strains of C pneumoniae. C pneumoniae was also capable of growing in endothelial cells derived from human cadaver coronary artery endothelial cells (CAEC). U-937 human macrophages that were infected with C pneumoniae were capable of transmitting the infection to CAEC when they were brought into contact with the endothelial cells by centrifugation, rocking overnight, and direct layering overnight, with and without using artificial laboratory tissue culture enhancements, such as centrifugation of the inoculum and cycloheximide in the growth media. The in vitro ability of C pneumoniae to maintain infections in macrophages, endothelial cells, and aortic smooth muscle cells may provide support for the hypothesis that C pneumoniae can infect such cells, which when followed by an immune response may contribute to atheroma formation in vivo. Stimulation of cytokine responses by infection with C pneumoniae has indicated that this organism is capable of interacting with the immune system. In vitro infection by C pneumoniae of U-937 macrophages stimulated the production of IL-1beta, IFN-gamma, and TNF-alpha in tissue culture. Human CAEC that are infected with C pneumoniae produce more IL-8 compared with those inoculated with killed C pneumoniae or negative control cells, indicating a chemokine response to infection that may play a role in recruitment of inflammatory cells to sites of infection in vascular cells. When IFN-gamma was used to up regulate HEp-2 and U-937 cells before infection by C pneumoniae, inhibition of a lytic growth cycle occurred in a dose related response. However, removal of the IFN-gamma after 24 to 48 hours' exposure allowed subsequent productive growth in the cells, perhaps indicating the prior induction of a persistent infection. More studies are needed to study the complex relationship between lytic infection and persistence, the ability of C pneumoniae to affect the immune response of vascular cells, and the potential for C pneumoniae to influence the initiation of or progression of atheromatous lesions. PMID- 10539861 TI - Value of animal models for Chlamydia pneumoniae-related atherosclerosis. AB - Chlamydia pneumoniae is strongly implicated in the pathogenesis of atherosclerosis in human beings. Animal models are important to help establish causality, to understand the mechanism of infection induced atherogenesis, to examine interaction of other factors or variables, to explore treatment regimens and their efficacy, and to help develop a vaccine for prevention. To date, the rabbit model is the only animal model shown to develop de novo atherosclerotic changes with C pneumoniae infection. However, the mouse model may be useful to show enhancement with other factors such as hypercholesterolemia and to explore pathogenic mechanisms. In our studies, we have shown that C pneumoniae respiratory infection in the rabbit results in early atherosclerotic changes in 26% with single inoculation and in 35% after triple inoculation, but sham infection or infection with Mycoplasma pneumoniae does not result in similar changes. Early treatment (5 days after inoculation) with 30 mg/kg per day azithromycin once every 6 days was 87% effective in preventing atherosclerotic changes, but delayed treatment (6 weeks after inoculation) was ineffective. Further studies are needed with longer or more aggressive regimens or possible combination of agents to determine whether it is possible to reverse preformed lesions. An effective vaccine for prevention of C pneumoniae -induced pneumonia and possibly atherosclerotic lesions in human beings would have tremendous application and would circumvent the shortcomings of antibiotic therapy. PMID- 10539862 TI - Animal models of chlamydia and atherosclerosis. PMID- 10539863 TI - Mouse models of Chlamydia pneumoniae infection and atherosclerosis. PMID- 10539864 TI - Transgene as vaccine for chlamydia. AB - BACKGROUND: We evaluated the potential utility of DNA immunization with the major outer membrane protein (MOMP) gene of Chlamydia trachomatis mouse pneumonitis (MoPn) strain for induction of protective immunity to chlamydial infection in mice. METHODS AND RESULTS: Groups of Balb/c mice were immunized with naked DNA intramuscularly or intranasally or with MOMP DNA-transfected Salmonella typhimurium delivery orally. Mice were challenged with MoPn through the pulmonary route to assay for protective immunity. All 3 routes of DNA immunization elicited protective immunity. Mucosal delivery appeared more efficacious than intramuscular delivery. CONCLUSIONS: DNA immunization with the chlamydia MOMP gene may be suitable for vaccine development. PMID- 10539865 TI - Helicobacter and atherosclerosis. PMID- 10539866 TI - Dental infections and atherosclerosis. AB - In most countries, coronary heart disease is one of the leading causes of morbidity and death. This report reviews the current evidence indicating that oral conditions (specifically periodontitis) may be a risk factor for atherosclerosis and its clinical manifestations and provides new preliminary data. This review is done in the context of the research indicating that inflammation plays a central role in atherogenesis and that there is a substantial systemic microbial and inflammatory burden associated with periodontal disease. Our review concentrates on 5 longitudinal studies that show oral conditions being associated with the onset of coronary heart disease while controlling for a variety of established coronary heart disease risk factors. In addition to published evidence, preliminary findings from our Dental Atherosclerosis Risk in Communities study also indicate that periodontal disease is associated with carotid intimal-medial wall thickness, a measure of subclinical atherosclerosis, adjusting for factors known to be associated with both conditions. PMID- 10539868 TI - Emerging role of antibiotics in atherosclerosis. AB - It has been shown that plaque composition changes significantly in the setting of acute events, macrophages and T cells being the predominant pattern at the immediate site of fissure or erosion. There appears to be a relation between physical blood stream factors, plaque morphology, and the distribution of inflammatory cells. Furthermore, there is cumulative evidence for the presence of intracellular pathogens in the arterial wall, namely Chlamydia pneumoniae and cytomegalovirus, which affect endothelial cells, monocytes, and macrophages. The ROXIS trial has shown some encouraging evidences for the potential role of intracellular pathogens in acute coronary syndromes. The ongoing WIZARD trial evaluates in a large population whether the addition of an antibiotic provides better outcome for coronary patients. PMID- 10539867 TI - Multiple infections in carotid atherosclerotic plaques. AB - BACKGROUND: Chlamydia pneumoniae, cytomegalovirus, herpes simplex virus, and recently, periodontal disease, have been associated with human atherosclerosis. Porphyromonas gingivalis and Streptococcus sanguis are major pathogens associated with periodontitis, a common chronic inflammatory condition in adults. Investigators have found that these infectious agents may influence vascular cell functions by inducing thrombus formation, vascular cell proliferation, apoptosis, and cell death. METHODS AND RESULTS: The main purpose of our study was to investigate the relation between the presence of multiple infectious agents in human carotid endarterectomy specimens and pathoanatomic features of the corresponding carotid plaques. Histologically, plaque rupture of the fibrous cap and communication of the luminal thrombus with the central necrotic lipid core was seen in or at proximity to the macrophage-rich shoulder (unstable plaque region). Thrombus within the lipid core without plaque rupture was occasionally found near the internal elastic lamina, associated with increased vascularity and lymphocytic infiltrate. Apoptosis, as detected by both the immunohistochemical staining of apoptosis-related proteins and in situ labeling of internucleosomally degraded DNA, was common in atherosclerotic plaques. Immunostainings for C pneumoniae, cytomegalovirus, herpes simplex virus-1, P gingivalis, and S sanguis were positive in the carotid plaques. From 1 to 4 organisms were found in the same specimen. The micro-organisms were immunolocalized in plaque shoulders and lymphohistiocytic infiltrate, associated with ulcer and thrombus formation, and adjacent to areas of strong labeling for apoptotic bodies. CONCLUSIONS: Our data provide evidence that multiple infectious agents may be found in atherosclerotic plaques, and sometimes in the same specimen. The current study is the first to report the detection of 2 major odontopathogens, P gingivalis and S sanguis, in atherosclerotic plaques. The immunolocalization of these micro-organisms within unstable plaque regions and their association with plaque ulceration, thrombosis, and apoptosis in vascular cells are intriguing. Multiple infectious agents may alter vascular cell function and provide a "trigger" for acute ischemic stroke events. Further evidence from human studies and animal models will be needed. PMID- 10539869 TI - Chlamydia pneumoniae, monocyte activation, and azithromycin in coronary heart disease. PMID- 10539870 TI - WIZARD and the design of trials for secondary prevention of atherosclerosis with antibiotics. AB - Clinical trials to assess the merit of antibiotic intervention in the treatment of ischemic cardiovascular disease are now underway, spurred on by an association between Chlamydia pneumoniae and atherogenesis noted in epidemiologic investigations, histopathologic studies, and results from various animal models. The design of such clinical trials must take into account a number of issues: the primary event as strictly defined by objective criteria, the event rate in the chosen population, the potential treatment effect, the availability of patients, the underlying cause of their atherosclerotic disease, the determination of the C pneumoniae-infected population to study, the dose and duration of the antibiotic, and the length of follow-up. In the design of the WIZARD study (Weekly Intervention with Zithromax for Atherosclerosis and its Related Disorders), an attempt was made to take these issues under consideration. Patients were randomly assigned either to 600 mg/d zithromax for 3 days then 600 mg/wk for 11 additional weeks or to placebo. Patients in the study had a myocardial infarction at least 6 weeks previously, had no recent coronary artery bypass graft or percutaneous transluminal coronary angioplasty, and did not required long-term administration of antibiotics. Patients were required to have an immunoglobulin G titer to C pneumoniae of >/=1:16. The primary end point was the time to a composite of all cause death, myocardial infarction, a revascularization procedure, or hospitalization for angina. The study enrolled 3500 patients, sufficient to detect a 25% reduction in the presumed 8% placebo event rate with 90% power. Follow-up will continue through the prespecified number of end points. PMID- 10539871 TI - Secondary prevention trials for coronary artery disease with antibiotic treatment for Chlamydia pneumoniae: design issues. PMID- 10539872 TI - Fulfillment of Koch's postulates and the causes of atherosclerosis. PMID- 10539873 TI - How to design studies to confirm a link between bacterial infection and atherosclerosis. PMID- 10539874 TI - Designing studies to confirm a link between viral infection and atherosclerosis. PMID- 10539875 TI - Design of future intervention studies for Chlamydia pneumoniae in atherosclerosis. PMID- 10539876 TI - How to design vaccination trials to prevent atherosclerosis. PMID- 10539877 TI - Conclusions. PMID- 10539878 TI - The perioperative complication rate of orthopedic surgery in sickle cell disease: report of the National Sickle Cell Surgery Study Group. AB - Orthopedic disease affects the majority of sickle cell anemia patients of which aseptic necrosis of the hip is the most common, occurring in up to 50% of patients. We conducted a multicentered study to determine the perioperative complications among sickle cell patients assigned to different transfusion regimens prior to orthopedic procedures: 118 patients underwent 138 surgeries. The overall serious complication rate was 67%. The most common of these were excessive intraoperative blood loss, defined as in excess of 10% of blood volume. The next most common complication was sickle cell-related events (acute chest syndrome or vaso-occlusive crisis), which occurred in 17% of cases. While preoperative transfusion group assignment did not predict overall complication rates, higher risk procedures were associated with significantly higher rates of overall complications. Transfusion complications were experienced by 12% of the patients. Two patients died following surgery. Both deaths were associated with an acute pulmonary event. The 52 patients undergoing hip replacements experienced the highest rate of complications with excessive intraoperative blood loss occurring in the majority of patients. Sickle cell-related events occurred in 19% of patients, and surgical complications occurred after 15% of hip replacements and included postoperative hemorrhage, dislocated prosthesis, wound abscess, and rupture of the femoral prosthesis. There were twenty-two hip coring procedures. Acute chest syndrome occurred in 14% of the patients. Overall, decompression coring was a safer, shorter operation. A randomized prospective trial to determine the perioperative and long-term efficacy of core decompression for avascular necrosis of the hip in sickle cell disease is needed. In conclusion, this study demonstrates a high rate of perioperative complications despite compliance with sickle cell perioperative care guidelines. Pulmonary complications and transfusion reactions were common. This study supports the results previously published by the National Preoperative Transfusion in Sickle Cell Disease Group. These results stated that a conservative preoperative transfusion regimen to bring hemoglobin concentration to between 9 and 11 g/dl was as effective as an aggressive transfusion regimen in which the hemoglobin S level was lowered to 30%. PMID- 10539879 TI - All trans-retinoic acid decreases early mortality in patients with promyelocytic leukemia and can be given entirely on an outpatient basis. AB - The results of the treatment of 43 patients with acute promyelocytic leukemia (PML) are reported: 27 were treated initially with all-trans-retinoic acid (ATRA), whereas 16 were treated with conventional chemotherapy. All patients received myelosuppressive chemotherapy after the initial treatment. Respectively, the complete remission rate was 92% and 37% (P < 0.01), the 5-day mortality rate was 0% and 44% (P < 0.001), and the 28-day mortality rate was 4% and 44% (P < 0.001). The median disease-free survival was 12 and 1 months (P < 0.01), whereas the 12-month disease-free survival was 50% and 13% (P < 0.01) and the 36-month disease-free survival was 41% and 9% (P < 0.01). Thirteen of the patients treated with ATRA were given the treatment fully as outpatients. ATRA given as initial therapy decreased significantly early mortality in promyelocytic leukemia patients; because some promyelocytic leukemia patients given ATRA as initial therapy can be treated as outpatients, the costs of this treatment modality may be diminished. PMID- 10539880 TI - Familial and metachronous malignant lymphoma: absence of constitutional p53 mutations. AB - Familial and metachronous aggregations of malignant lymphoma are well-documented, but the molecular basis of a predisposition for development of lymphoma is as yet unclear. Malignant lymphomas have been described as part of the spectrum of neoplasias in Li-Fraumeni syndrome (LFS), which is associated with constitutional mutations of p53. However, p53 germline mutations have also, albeit less frequently, been described in patients not fitting the clinical definition of LFS. To clarify whether a genetic predisposition for lymphoma is associated with constitutional p53 mutations, DNA from normal blood lymphocytes of 12 lymphoma patients with a family history of lymphoma and/or with metachronous lymphoma (median age 37 years) was examined for mutations of p53 exons 4-8. One patient had four first-degree relatives with Hodgkin's disease, acute leukemia, and carcinomas, but the family history did not fulfill criteria of LFS. Four patients with Hodgkin's disease were diagnosed with metachronous non-Hodgkin's lymphoma as a second malignant neoplasm. No constitutional p53 mutations were detected in any of these patients, implying that outside the clinical spectrum of LFS, constitutional p53 mutations are rare in patients with lymphomas. PMID- 10539881 TI - mRNA expression of variant Fas molecules in acute leukemia cells. AB - Fas (Apo-1/CD95) is a cell membrane receptor involved in apoptotic cell death. Soluble variant forms (sFas) lacking the transmembrane domain due to alternative splicing have been identified. Up-regulation of sFas expression is reportedly implicated in prereceptorial blockage of Fas-induced apoptosis in a dose dependent manner. We examined mRNA expression of Fas and sFas in fresh leukemia cells. All leukemia cells expressed both mRNAs of full-length Fas (FasFull) and sFas with deletion of exon6 (FasDel6). The ratio of FasFull/FasDel6 mRNA expression was not always correlated with Fas-mediated growth inhibition. Interestingly, in a 6-year-old boy with acute myelogenous leukemia, blast cells obtained at onset and at the time of bone marrow relapses expressed distinct amounts of FasDel6 mRNA. Furthermore, the level of FasDel6 expression appeared to be correlated with Fas-resistance in leukemia blasts. In addition, sFas protein levels were elevated in patients' sera at onset with subsequent return to normal levels after complete remission. These results indicated that sFas could be synthesized and released by leukemia blasts and suggested that up-regulation of Fas variant transcript might render leukemia blasts resistant to Fas-mediated growth inhibition in certain cases. PMID- 10539882 TI - Altered expression of CD45 isoforms in differentiation of acute myeloid leukemia. AB - Specific expression of different CD45 isoforms can be seen in various stages of differentiation of normal nucleated hematopoietic cells. Association of membrane expression of CD45 isoforms and differential levels of leukemia cells was studied in 91 cases with de novo acute myeloid leukemia (AML). Membrane expression of CD45RA and CD45RO was analyzed by flow cytometry and their expression patterns were compared with AML subtypes classified according to the French-American British (FAB) classification. CD45RA was essentially expressed in all of the FAB myelocytic subtypes (M0-M3). Its expression in percentage was lower in the most differentiated subtype of AML (M3) when compared with other myelocytic subtypes. CD45RO expression was rarely observed in cases with myelocytic subtypes (1/56 cases of M0, M1, M2, and M3) except for the minimally differentiated myelocytic subtype (M0) or those with potential for differentiation to T-cell lineage where three of 12 cases showed CD45RO expression. When leukemia cells of an M3 case were differentiated to mature granulocytes by treatment of all-trans-retinoic acid, they showed increasing expression of CD45RO. In subtypes with a monocytic component (M4 and M5), both of CD45RA and CD45RO expression were observed and mutually exclusive. When 10 cases of M5 were subdivided by the differential level into undifferentiated (M5a) and differentiated monocytic leukemia (M5b), expression of CD45RA and CD45RO was strictly restricted to cases with M5a and M5b, respectively. These results suggest that CD45 isoform expression in AML characterizes differential levels both in myelocytic and monocytic lineages and specifically disturbed in each subtype. The assessment of CD45 isoform expression appears to provide an insight on biological characteristics and a useful supplementary test for differential diagnosis of AML subtypes. PMID- 10539883 TI - Use of a novel platelet function analyzer (PFA-100) with high sensitivity to disturbances in von Willebrand factor to screen for von Willebrand's disease and other disorders. AB - The PFA-100 is a new platelet function analyzer which uses whole blood and high shear stress blood flow to simulate primary hemostasis and assess platelet function. A small volume of blood is introduced into a disposable cartridge, and forced through a capillary tube. Platelet adhesion and aggregation is then initiated following exposure to either collagen/ADP [C/ADP] or collagen/epinephrine [C/Epi] coated membranes. Movement of blood through the capillary, and its subsequent occlusion is monitored and yields the measured endpoint (closure time [CT] in seconds). Using two approaches, we assessed the sensitivity of this system to disturbances in the function of von Willebrand Factor (VWF). Firstly, we assessed the ability of the PFA-100 to detect the presence of von Willebrands Disease (VWD). Using normal individuals (N = 18), CTs (in seconds; mean [range = mean +/- 2SD]) were (i) C/ADP, 95 [66-124], (ii) C/Epi, 128 [98-158]. A panel of 47 patients undergoing evaluation for clinical hemostatic defects inclusive of VWD were also evaluated. All samples from patients confirmed to have VWD following specific VWF studies [N = 9; 3 x Type 1, 1 x Type 3, 1 x Type 2A, 4 x Type 2B] gave prolonged CTs (>/= 200 s) for both C/ADP and C/Epi membranes; in contrast, all patients yielding normal CT values were found to yield normal VWF results (i.e., were found not to suffer from VWD). Patients with hemophilia (1 x hemophilia A, 1 x hemophilia C) gave normal PFA-100 CT, while those with clinical thrombocytopaenia (N = 3) gave prolonged PFA-100 CT. A number of other patient samples also gave abnormal CT values which in some cases could be linked to recent aspirin consumption. In the second evaluation process, and using normal blood, we have assessed the ability of various antibodies to influence the CT. Of the monoclonal antibody panel tested [N = 20], only a proportion of those against VWF [6/10] or gp1b/IX [CD42; 2/5] were found to be inhibitory (i.e., prolonged the CT). Data using polyclonal antibodies (against platelets, VWF, fibrinogen and fibronectin) is more complex but largely confirms the sensitivity of the system to VWF. On the basis of these results, we conclude that the PFA-100 is highly sensitive to disturbances in VWF and to the presence of VWD and may thus provide a valuable screening test for VWD in certain specific circumstances (i.e., acute need conditions or remote testing sites; normal CT result generally effective as negative predictor, i.e. not severe VWD). However, since abnormal CT values were obtained in clinical situations other than evident VWD, the PFA-100 cannot be used as a specific diagnostic tool to establish the presence of VWD. Thus, any abnormal PFA-100 CT result should be thoroughly evaluated by follow-up specific testing to establish the true clinical disorder affecting the individual under investigation, inclusive of appropriate VWF assays if VWD is clinically suspected. PMID- 10539884 TI - Paroxysmal nocturnal hemoglobinuria: An acquired genetic disease. AB - Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired clonal hematopoietic stem cell disorder characterized by an intravascular hemolytic anemia. Abnormal blood cells lack a series of glycosylphosphatidylinositol (GPI)-anchored proteins. The lack of GPI-anchored complement regulatory proteins, such as decay accelerating factor (DAF) and CD59, results in complement-mediated hemolysis and hemoglobinuria. In the affected hematopoietic cells from patients with PNH, the first step in biosynthesis of the GPI anchor is defective. At least four genes are involved in this reaction step, and one of them, an X-linked gene termed PIG A, is mutated in affected cells. The PIG-A gene is mutated in all patients with PNH reported to date. Here, we review recent advances in the understanding of the molecular pathogenesis of PNH. PMID- 10539885 TI - Long-term treatment of refractory thrombocytopenia in a patient with Wiskott Aldrich syndrome with vincristine, immunoglobulin, and methylprednisolone. AB - We report a child with Wiskott-Aldrich syndrome with severe, refractory, symptomatic thrombocytopenia who achieved an excellent response to combination therapy with vincristine 1.5 mg/m(2) x 1 day, intravenous immunoglobulin 1 g/kg x 3 days, and methylprednisolone 25 mg/kg x 3 days (VIM) for 7 years after failing multiple treatments. He did not have a histocompatible donor for bone marrow transplantation. When the patient ceased to respond to this regimen, he was rescued with pulse dexamethasone. Vincristine, immunoglobulin, and methylprednisolone might serve as a novel treatment option for the patient with refractory thrombocytopenia. Our patient had a sustained remission of symptomatic thrombocytopenia without toxicity. Furthermore, pulse dexamethasone might be an alternative treatment option to which patients with Wiskott-Aldrich syndrome may respond. PMID- 10539886 TI - Association of unstable hemoglobin variants and heterozygous beta-thalassemia: example of a new variant Hb Acharnes or [beta53(D4) Ala --> Thr]. AB - We report here the functional and structural characterization of Hb Acharnes [beta53(D4) Ala --> Thr], an unstable and electrophoretically silent variant, that was found associated in trans with a beta(0)-thalassemic mutation (IVSI-1 G -> A), in a patient with thalassemia intermedia syndrome. This case is discussed in comparison with other sporadic cases that we have previously investigated, resulting from the co-inheritance of a beta(0)-thalassemic mutation (CD39 C --> T) with two other types of unstable hemoglobins, Hb Koln [beta98(FG5) Val --> Met], and Hb Arta [beta45(CD4) Phe --> Cys]. It may be concluded that, in these associated forms, both the degree of instability of the variant and the altered oxygen binding properties (affecting the degree of tissue hypoxia) are major determinants of their clinical expression. PMID- 10539887 TI - Clonality of acquired primary pure red cell aplasia. AB - Acquired primary pure red cell aplasia (PRCA) has frequently been shown to be associated with T cells that inhibit marrow erythropoiesis. A 41-year-old female presented with anemia in December 1985. Bone marrow examination revealed 1.8% erythroid cells. A diagnosis of PRCA was made. She received prednisolone, and her hemoglobin level recovered to 12 g/dl. In February 1995, her hemoglobin level decreased to 5.8 g/dl, and acquired primary PRCA recurred. Surface markers of peripheral blood mononuclear cells demonstrated CD4/8 ratio inversion. The T-cell receptor (TCR)-beta chain gene showed germ line configuration by Southern blot analysis of the mononuclear cells in peripheral blood. However, stepdown polymerase chain reaction analysis revealed that the TCR-beta gene of peripheral blood mononuclear cells was rearranged. With highly sensitive polymerase chain reaction analysis, clonality of T cells was confirmed. We propose that some acquired primary PRCA patients have a T-cell clonal disorder, similar to some PRCA patients with large granular lymphocytes leukemia or thymoma. PMID- 10539888 TI - Factitious methemoglobinemia. AB - We report the case of a 26 year-old female who was treated on numerous occasions for methemoglobinemia believed secondary to surreptitious abuse of dapsone as part of a factitious disorder. PMID- 10539889 TI - Feasibility of restriction enzyme protocols for the molecular diagnosis of abnormal hemoglobins in Turkish population. PMID- 10539890 TI - Mesenteric vein thrombosis secondary to combined protein C deficiency and double heterozygosity for factor V Leiden and prothrombin G20210A. PMID- 10539891 TI - Salvage therapy and long-term remission with danazol and cyclosporine in refractory Evan's syndrome. PMID- 10539892 TI - Is it time to revisit the classification system for cervicovaginal cytology? PMID- 10539893 TI - A portrait in history. Sir James Paget: surgeon to Queen Victoria. PMID- 10539894 TI - Patient outcomes and pathology practice: An introduction to the College of American Pathologists Conference XXXIV on Molecular Pathology: Role in Improving Patient Outcome. AB - This article provides an abbreviated conceptual framework for viewing general issues of outcomes research and management and consideration of the emerging interest in patient-referenced outcomes in medicine and pathology. Specific issues addressed are the reasons for increased interest in outcomes research within the past decade; a review of the current language of outcomes management and research; a critique of the advantages and weaknesses of contemporary outcomes analysis and research methods; a summary of the responses of government, regulatory and accreditation organizations, medical societies, and the College of American Pathologists to the outcomes movement; and, in conclusion, a discussion of opportunities provided through outcomes management for enhanced involvement by pathologists in the care of patients. PMID- 10539895 TI - Molecular pathology: role in improving patient outcome: Overview. PMID- 10539896 TI - Molecular methods in the detection and identification of mycobacterial infections. AB - Nucleic acid amplification (NAA) tests for direct detection of Mycobacterium tuberculosis complex in respiratory specimens have the potential to provide a more rapid diagnosis of pulmonary tuberculosis (TB) than is currently possible by conventional stain, culture, and identification tests. Currently, 2 NAA tests enhanced Amplified Mycobacterium Tuberculosis Direct (MTD) Test (Gen-Probe, Inc) and Amplicor Mycobacterium tuberculosis Test (Roche Molecular Systems, Inc)-have been approved by the Food and Drug Administration for testing respiratory specimens that are smear positive for acid-fast bacilli (AFB). This restriction to AFB smear-positive specimens was based on data from the initial clinical trials conducted to evaluate these products that showed low sensitivity (ie, 48% 53%) and less-than-optimal specificity (ie, 96%-99%) in AFB smear-negative specimens. Data from the clinical trial for the enhanced MTD test and from 2 subsequent studies, however, suggest that this version of the MTD test is a reliable tool for rapid diagnosis of pulmonary TB, regardless of the AFB smear result. Both NAA tests have been evaluated for diagnosis of extrapulmonary TB, and results were comparable to the results of tests performed with respiratory specimens. The NAA tests also appear to be reliable for rapid identification of M tuberculosis complex in positive broth cultures of all specimen types except blood. The impact of the NAA tests on patient outcome varies based on the AFB smear result. With smear-positive results, public health and hospital infection control resources are predominantly affected. With smear-negative results, however, the potential for affecting patient outcome is much greater. In patients with smear-negative results, the NAA test can result in earlier diagnosis of TB and subsequent initiation of therapy. Use of these tests also may eliminate the need for invasive diagnostic procedures, which are costly and pose an added risk to the patient, and they may allow earlier discharge of hospitalized patients. PMID- 10539897 TI - Molecular epidemiology in the care of patients. AB - Several different epidemiologic typing methods have been applied in studies of microbial pathogens. These methods include the more traditional nonmolecular approaches as well as the more sophisticated molecular typing methods. Application of traditional epidemiologic typing methods, such as antibiogram, serotyping, biotyping, and phage typing, have occasionally been useful in describing the epidemiology of infectious diseases. However, these methods have generally been considered to be too variable, labor intensive, and slow to be of practical value in epidemiologic investigations. In response to these limitations, several techniques have been adopted from the molecular biology field for use as epidemiologic typing methods and have been applied in studies of bacteria, fungi, viruses, and protozoa. The most widely used molecular typing methods are the DNA-based methods, such as plasmid profiling, restriction endonuclease analysis of plasmid and genomic DNA, Southern hybridization analysis using specific DNA probes, and chromosomal DNA profiling using either pulsed field gel electrophoresis or polymerase chain reaction-based methods. The various molecular typing methods may be applied to the investigation of outbreaks of infections or may be used in the context of epidemiologic surveillance. For outbreak investigation, typing methods are used to compare isolates from a suspected outbreak to delineate clonally related and unrelated strains with the goal of short-term control of transmission. In the context of epidemiologic surveillance, molecular typing methods may be used to monitor geographic spread and prevalence shifts of epidemic and endemic clones with the goal of long-term evaluation of preventive strategies or for the detection and monitoring of emerging and reemerging infections. The specific typing method selected may vary with the task at hand; however, the typing studies must always be used to supplement, rather than replace, careful epidemiologic investigation. PMID- 10539898 TI - Impact of viral load testing on patient care. AB - Quantitative human immunodeficiency virus (HIV) type 1 RNA tests have been essential tools in increasing our understanding of HIV pathogenesis and antiretroviral therapy. The plasma HIV RNA level is among the most powerful predictive tests in modern medicine for disease progression and has rapidly become the standard of practice for guiding clinicians in initiating, monitoring, and changing antiretroviral therapy. In this article the scientific rationale and clinical indications for viral load testing in HIV infection are reviewed. PMID- 10539899 TI - Infection, immunity, and cancer. AB - A significant percentage of human cancers worldwide are associated with infections due to known viruses, including human papillomaviruses (cervical cancer and other skin cancers), human T-lymphotropic viruses (adult T-cell leukemias and lymphomas in endemic areas), hepatitis B virus (liver cancer), and Epstein-Barr virus (Burkitt lymphoma and nasopharyngeal carcinoma). The fraction of human cancers attributable to infection may now need to be revised in light of the fact that new viral associations have been discovered and other nonviral associations have been identified. This article addresses the increasingly recognized role of infectious agents as precipitants of human neoplasia and the possibility that novel diagnostic, therapeutic, and chemopreventive strategies may emanate directly from research directed at identifying and understanding these agents. PMID- 10539900 TI - Laboratory determination of hereditary susceptibility to breast and ovarian cancer. AB - Inherited mutations in the genes BRCA1 and BRCA2 are associated with a significantly increased risk of breast cancer, particularly before the age of 50 years, as well as an increased risk of ovarian cancer. Patients with early-onset breast cancer or ovarian cancer at any age with a family history of either disease are at higher risk of carrying a mutation in BRCA1 or BRCA2. Laboratory analysis of these genes can determine whether a patient has inherited an increased risk of breast and ovarian cancer. In the absence of a mutation that has been previously identified in a family member, most tests for hereditary breast-ovarian cancer risk analyze the entire coding sequences of BRCA1 and BRCA2. The gene sequencing process itself can be automated, but the data must be interpreted by an individual with training in molecular diagnostics. Management options generally available to individuals with hereditary susceptibility to breast and ovarian cancer include heightened surveillance, prophylactic surgery, and chemoprevention. The use of genetic techniques to identify women with increased risk of cancer demonstrates the application of recent advances in the understanding of the genetic basis of malignancy to laboratory medicine and clinical care. PMID- 10539901 TI - Colon cancer testing and screening. AB - Laboratory testing has the potential to favorably affect the outcome of patients with neoplasia by applications in screening, risk identification, surveillance, diagnosis, prognosis, prediction of tumor response or resistance to therapies, and prediction of toxic effects from therapies. Molecular testing directed at colorectal neoplasia is currently in use for diagnosis and characterization of rare, highly penetrant inherited syndromes due to germline mutation of the APC suppressor gene or DNA nucleotide mismatch repair genes. Routine application of molecular assays of colorectal tumors as prognostic and predictive markers is likely to occur in the near future, but major advances in technology are needed to begin to move molecular testing into screening of the general population and surveillance of subjects at increased risk. It seems certain that molecular methods will become increasingly important in improving the outcome of patients with colorectal neoplasia and in contributing to continued decline of the death rate from this common cancer. PMID- 10539902 TI - Minimal residual disease in hematologic disorders. AB - In almost no other area of medical oncology has the introduction of new drugs, combinations of chemotherapeutic agents, and novel biologic treatments caused such dramatic responses as it has in the treatment of malignant hematologic disorders. However, despite some therapeutic success, many patients relapse and die from recurrence of their disease. The implications of minimal residual disease (MRD), a term referring to disease that is undetectable by conventional morphologic methods, have therefore attracted increasing attention in recent years. New and powerful laboratory tools such as polymerase chain reaction assays have extraordinary sensitivity and provide exciting new insights into the detection, nature, quantification, and kinetics of MRD. This article summarizes methods used in the identification of MRD and its importance as exemplified in the case of acute leukemias and chronic myelogenous leukemia. PMID- 10539903 TI - Molecular diagnosis and prognosis in gynecologic oncology. AB - Currently, molecular pathology plays a limited role in improving patient outcome in gynecologic oncology. However, molecular investigation is providing important insights into the epidemiology, pathogenesis, and progression of female genital cancers. Future roles should include prediction of poor outcome in low-risk cases, more accurate staging of multifocal tumors, identification of new precursor lesions, and prediction of response to specific therapeutic regimens. Gene therapy of some malignant tumors may become important in the near future. In the immediate future, however, the most significant role of molecular pathology may be in the screening and triage of putative cervical cancer precursors and in the possible prophylaxis of these lesions by means of a vaccine or vaccines against human papillomaviruses. PMID- 10539904 TI - Cystic fibrosis: molecular diagnosis, population screening, and public policy. AB - OBJECTIVE: To review the current status of scientific knowledge and opinion regarding molecular genetic testing of mutations in the CFTR gene for purposes of diagnosis and population carrier screening of cystic fibrosis (CF). DATA SOURCES: Published research findings on the nature of the CFTR gene, pilot population screening studies in the United States and Europe, and ongoing deliberations of professional and governmental agencies considering implementation of widespread testing. STUDY SELECTION: Findings relevant to the molecular heterogeneity of CFTR mutations and its implications for population carrier screening were considered. DATA EXTRACTION: Information was extracted from studies published by us and others, as made available to recent consensus panels and professional committees. DATA SYNTHESIS: These data were reevaluated in light of recent movements in professional and public policy regarding acceptability and desirability of widespread CF mutation testing. Effects to date of such testing on patient outcomes is reported. CONCLUSIONS: The ability to test for CFTR mutations at the molecular level has already improved the diagnosis of symptomatic patients and expanded the reproductive options of family members of CF patients. The same technology also holds promise of identifying asymptomatic carriers and at-risk couples without family history in the general population so that they too might be offered prenatal diagnosis or other options. However, a number of key questions remain to be worked out before a widespread national screening program can be put into practice. These include the target population to be offered testing (the entire population vs high-risk ethnic groups), the size and nature of the mutation test panel (universal vs ethnic specific), the inclusion or exclusion of CFTR variants that do not cause classical CF, the optimal testing technology, appropriate standards for laboratory quality assurance, and the development of sufficient educational materials and genetic counseling resources for test delivery, reporting, and interpretation. The answers to these questions will be relevant not only to CF testing but also to many other large-scale molecular genetic screening programs being considered in the future. PMID- 10539905 TI - Utility of RET mutation analysis in multiple endocrine neoplasia type 2. AB - OBJECTIVE: To review the role of RET mutation analysis in the diagnosis of multiple endocrine neoplasia type 2 (MEN 2) and in presymptomatic screening for this disorder. DATA SOURCES: Review of the medical literature and current clinical practice. CONCLUSIONS: RET mutation analysis is a sensitive and specific test for MEN 2. It plays a pivotal role in the diagnosis and management of patients and families with MEN 2 and in the individual who presents with an apparently sporadic medullary thyroid carcinoma or pheochromocytoma. These disorders may first come to the attention of either the anatomic or clinical pathologist, who has the opportunity to see that appropriate testing is done. As with any familial disease, professional genetic counseling is an important part of the care of these patients. PMID- 10539906 TI - Muscular dystrophy: identification and use of genes for diagnostics and therapeutics. AB - The application of cloned genes and their protein products to molecular diagnostics has been an increasingly important area of pathology. The first gene to be identified by positional cloning was the Duchenne muscular dystrophy gene, mutations of which cause one of the most common and most devastating human inherited conditions. The identification of the responsible gene and the encoded dystrophin protein has resulted in a large series of studies concerning the other components of the membrane cytoskeleton of myofibers and their involvement in different forms of muscular dystrophy. Through the study of patients deficient in specific components of the muscle fiber, much is being learned about normal myofiber structure and function and dysfunction in disease states. A new frontier is the application of the normal genes and proteins toward patient therapeutics (gene therapy). Although highly experimental, delivery of therapeutic genes promises to become an important medical practice. PMID- 10539907 TI - Hereditary hemochromatosis: impact of molecular and iron-based testing on the diagnosis, treatment, and prevention of a common, chronic disease. AB - OBJECTIVE: To review the current state-of-the-art regarding the role of iron- and DNA-based testing on the detection, treatment, and prevention of hereditary hemochromatosis (HH), the most common single-gene disorder in white people. SOURCES: Review of the medical literature, with particular emphasis on recent reports of the impact of DNA-based testing on the detection of symptomatic and presymptomatic patients with HH. CONCLUSIONS: Hereditary hemochromatosis, a common autosomal recessive iron overload disorder (with a population prevalence of 0.3%-0.8%), is a common cause of preventable liver, heart, joint, and endocrine disease. Since the associated clinical signs and symptoms are nonspecific, an accurate HH diagnosis demands both a high index of suspicion and the direct laboratory demonstration of elevated iron parameters. The substantial public health burden of HH as a common, deadly, detectable, and treatable chronic disease has led the College of American Pathologists to recommend that "systematic screening for hemochromatosis is warranted for all persons over the age of 20 years." The recent discovery that most HH cases are the result of a single well-conserved homozygous missense mutation (C282Y) within a novel transferrin-receptor binding protein (HFE) has given rise to diagnostic clinical tests for the DNA-based detection of this pathologic mutation. This direct HFE mutation test can now be used not only to confirm the diagnosis of HH in those with symptomatic disease, but also, perhaps more importantly, to detect those with presymptomatic iron overload in whom future disease manifestations may be prevented (with phlebotomy therapy). PMID- 10539908 TI - Impact of molecular (DNA) testing on determination of parentage. AB - OBJECTIVE: To examine changes in parentage testing practices since the introduction of DNA polymorphisms. METHODS: Comparison of data from American Association of Blood Banks (AABB) annual questionnaires and the responses of participants to AABB-College of American Pathologists proficiency test panels. RESULTS: DNA polymorphisms have led to a complete change in the technical methods used by parentage testing laboratories. CONCLUSIONS: The widespread use of DNA methods has increased the power of the information routinely provided to the courts in cases of disputed paternity, to agencies needing information about relatedness, and to the individuals who are tested. PMID- 10539909 TI - Impact of DNA typing on standards and practice in the forensic community. AB - This article reviews the history of DNA-based human identification from its inception in 1985. Since the development of the technology, experts called for setting of standards and use of proficiency tests for quality assurance measures. The response of the National Institute of Standards and Technology to DNA forensic standards needs was catalyzed by the Technical Working Group on DNA Analysis Methods, sponsored by the Federal Bureau of Investigation with funding provided by the National Institute of Justice. Standard reference materials were developed for the original technologies used in DNA identification and for the newer polymerase chain reaction-based technologies. Adoption of recommended standards developed through the Federal Bureau of Investigation-commissioned DNA Advisory Board show the acceptance of National Institute of Standards and Technology standards for calibration of laboratory protocols. New technologies will require a process of validation and continued testing through the use of proficiency tests, such as those provided through the College of American Pathologists. Robotics and parallel processing of samples will lead to increased efficiency in DNA testing. The use of DNA data banks of convicted felons will increase dramatically with the the Federal Bureau of Investigation's national implementation of a computerized identification system known as the Combined DNA Index System. This system that will make major use of short, tandem, repeat genetic systems and will be the major driver of technology for the next 5 to 10 years. Finally, sample collection and training are of major concern for those who look at the long-term impact of DNA testing in forensic laboratories. PMID- 10539911 TI - Financial determinants of outcomes in molecular testing. AB - For the benefits of molecular pathology to outweigh its inherent costs, testing procedures must be integrated into total disease assessment to realize the true financial impact. Major financial benefits are achievable from molecular testing because the tests reduce the use of less sensitive and less specific tests, unnecessary diagnostic procedures, and ineffective therapies. In this review, the financial determinants of outcomes for molecular-based testing for disease predisposition, screening, early detection, and directed therapy are presented. PMID- 10539910 TI - Efforts to regulate the collection and use of genetic information. AB - Public fascination with and support for genetic medicine is complicated by a deeply held fear that genetic information will be used by third parties (eg, insurers, employers, school systems) in ways that will harm the individuals from whom it was derived. Since the mid-1990s there has been much state and some federal legislative activity to address 2 closely related issues: the maintenance of genetic privacy and the prevention of genetic discrimination. These laws have had to confront several challenging questions such as what constitutes a genetic test, is genetic information qualitatively different from other medical information, and is there a means to distinguish between the two. In general the state laws are not well crafted. I will argue that a far more preferable policy is to draft a global, comprehensive medical records privacy law and to develop a model statute that defines the role of predictive genetic information in insurance underwriting. Concerns over misuse of genetic information also pose major issues for the conduct of genomic research. Among those I discuss are ownership of the DNA sample, significant changes in the scope of consent that must precede the decision to volunteer as a subject in genomic research, the reuse of long-archived samples, the challenges to intellectual property rights that flow from research, and the rise of the doctrine of community consent. PMID- 10539912 TI - Ethical issues in molecular pathology: paradigms in flux. AB - Recent advances in molecular pathology and molecular genetics have created new concerns about the use of human biologic materials in research. Since researchers now have the ability to extract and amplify DNA from minuscule archived samples, virtually any human tissue sample can potentially become the template for a test that provides information that may relate to the inherited genes of an individual. Researchers using human biologic materials should follow the 3 basic principles that have been defined for all ethical human subjects research: respect for persons, beneficence, and justice. Institutional Review Boards are responsible for providing review of the risks and benefits of research proposals to safeguard the rights and welfare of human subjects. Currently, there is considerable debate concerning the role of informed consent procedures and the Institutional Review Board oversight process in situations when researchers use human biologic materials that have been anonymized or coded. In 1999, the National Bioethics Advisory Commission is expected to make recommendations to President Clinton and the National Science and Technology Council that are expected to clarify the balance between respect for personal autonomy and the societal need to pursue biomedical research to improve the health and welfare of all individuals. PMID- 10539913 TI - Interobserver variability in subclassification of squamous intraepithelial lesions: Results of the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology. AB - OBJECTIVE: To determine whether, on a national cytology proficiency test, a competent cytologist can consistently distinguish grades of squamous intraepithelial lesions. DESIGN: Results for low- and high-grade squamous intraepithelial lesion referenced slides from the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology for 1996 and 1997 were analyzed including educational, nongraded vs graded validated slides. RESULTS: The discrepant rate between low- and high- grade lesions ranged from 9.8% to 15% for cytotechnologist, pathologist, laboratory, and all responses. There was a statistically significant difference in performance on graded, validated slides vs educational slides with better performance on validated slides. CONCLUSION: This significant interobserver variability in subclassification of squamous lesions should be considered in management guidelines for abnormal Papanicolaou test results and implementation of national cytology proficiency testing. PMID- 10539914 TI - Prostate-specific antigen expression and lipochrome pigment granules in the differential diagnosis of prostatic adenocarcinoma versus seminal vesicle ejaculatory duct epithelium. AB - BACKGROUND: Lipochrome pigment granules (LPGs) and prostate-specific antigen (PSA) localization have been cited as helpful adjuncts in differentiating atypical histologic patterns of seminal vesicle-ejaculatory duct (SVED) from prostatic adenocarcinoma. However, LPGs have been described in both benign and neoplastic prostatic acini, and PSA expression within the intraprostatic SVED has not been fully explored. DESIGN: Fifty radical prostatectomy specimens were studied for LPGs and 9 cases for PSA expression. RESULTS: Two morphologic types of LPGs (type 1 and type 2) were observed. The reproducibility in classifying LPGs was evaluated by kappa statistics, which demonstrated a strong agreement between 4 observers. Type 1 was restricted to SVED in all 50 specimens. Type 2 was subclassified into 2A and 2B. Type 2 LPGs were observed in prostatic acini of different zones, high-grade prostatic intraepithelial neoplasia, prostatic adenocarcinoma, and occasionally with type 1 LPG in SVED. Focal reactivity for PSA in the distal portion of SVED near urethra was noted in 1 of 9 cases. CONCLUSION: Awareness about morphologic differences between the 2 types of LPGs could help to avoid a potential diagnostic pitfall of misinterpreting SVED epithelium for adenocarcinoma. Caution is recommended in interpreting PSA expression, since rare focal PSA reactivity was observed in the distal SVED. PMID- 10539915 TI - Lack of Epstein-Barr virus infection in cervical carcinomas. AB - CONTEXT: The Epstein-Barr virus (EBV) is a ubiquitous microorganism strongly associated with lymphoproliferative disorders and a large number of human neoplasms, mainly undifferentiated nasopharyngeal carcinoma and Burkitt lymphoma. The viral DNA has been detected in other tumors, such as carcinomas from tonsil, salivary glands, and thymus, and malignancies of the female genital tract. Some authors have proposed that EBV could play a role in the carcinogenesis of cervical tumors; however, other studies do not support this hypothesis. OBJECTIVE: To assess whether EBV is associated with female genital tract neoplasms. DESIGN: Sixty-five biopsy specimens (5 in situ carcinomas, 24 invasive squamous cell carcinomas, 6 lymphoepithelioma-like carcinomas, and 30 endocervical adenocarcinomas) were used to perform EBV detection through RNA in situ hybridization. RESULTS: None of the cervical carcinoma cases studied was positive for EBV infection. CONCLUSIONS: The results suggest that it is still premature to incriminate EBV in the carcinogenesis of cervical carcinoma. PMID- 10539916 TI - Evaluation of the enhanced amplified Mycobacterium tuberculosis direct test for direct detection of Mycobacterium tuberculosis complex in respiratory specimens. AB - OBJECTIVE: To evaluate the performance of the enhanced Mycobacterium Tuberculosis Direct Test (E-MTD), for the direct detection of M tuberculosis complex (MTBC) in respiratory specimens. DESIGN: Two hundred seventy-four respiratory specimens from 151 patients in respiratory isolation were tested with the E-MTD, and the results were compared with the results of mycobacterial smear, culture, and the earlier form of the test, MTD-1. RESULTS: Forty-one specimens were culture positive for mycobacteria (20 MTBC and 21 nontuberculous mycobacteria), 23 of which were smear positive (16 MTBC, 7 nontuberculous mycobacteria). Twenty-four specimens were positive by E-MTD, and 21 were positive by MTD-1. Of the 20 MTBC culture-positive specimens, 19 were positive by the E-MTD and 19 were positive by the MTD-1. The remaining specimens were MTBC negative by all methods. After resolution of discrepancies, the sensitivity, specificity, and positive and negative predictive values were 95.2%, 100%, 100%, 99.6% for the MTD-1 and 95.2%, 98.8%, 87.0%, and 99.6%, for the E-MTD. For the E-MTD smear-positive and smear negative specimens, these same values were 93.8%, 100%, 100%, and 87.5% and 100%, 98.8%, 62.5%, and 100%, respectively. CONCLUSION: The results suggest that the E MTD is a reliable method for the direct detection of MTBC in smear-positive respiratory specimens. PMID- 10539917 TI - Metastatic Leydig cell tumor with sarcomatoid differentiation. AB - Leydig cell tumors of the testis are uncommon. Only about 10% of cases have a malignant course. It has been stated that the only definite criterion for malignancy is presence of metastasis. We present a 47-year-old patient with metastatic Leydig cell tumor 17 years after initial diagnosis, to our knowledge the longest reported interval between diagnosis and the development of metastasis. The primary tumor did not exhibit convincing features of malignancy. The initial metastasis in the right perirenal fat tissue showed a biphasic tumor with sarcomatoid differentiation not described previously in a metastatic Leydig cell tumor. PMID- 10539918 TI - Invasive cystic hypersecretory ductal carcinoma of breast: a case report and review of the literature. AB - Few individual cases of invasive cystic hypersecretory ductal carcinoma of the breast have been described. Review of 33 cases of cystic hypersecretory carcinoma, including the current case, indicate that only 6 cases presented with invasive disease. Two of these cases had positive nodes and 2 had distal metastases. The case presented here is unique in an additional aspect: the contralateral breast harbored lobular breast carcinoma 10 years after mastectomy of the first malignancy. Bilateral breast disease resulting in bilateral mastectomies over long-term follow-up, as in the case presented here, was reported in 3 of 33 cases. PMID- 10539919 TI - Spurious elevation of automated platelet counts in secondary acute monocytic leukemia associated with tumor lysis syndrome. AB - The intent of this article is to describe the effect of tumor lysis on automated platelet counts in therapy-related, secondary acute monocytic leukemia. The first patient was a 69-year-old man with large cell carcinoma of the lung who developed acute monocytic leukemia 1(1/2) years after initiation of radiation and chemotherapy for his carcinoma. The second patient was a 72-year-old female with peripheral T-cell lymphoma who developed acute monocytic leukemia 1 year after initiation of chemotherapy for her lymphoma. Platelet counts were determined by the automated Coulter (STKS) counter. Both patients had clinical and laboratory evidences of tumor lysis syndrome and disseminated intravascular coagulation. The peripheral blood smears revealed numerous fragments of leukemic cells and apoptotic cells with pyknotic nuclei. The Coulter machine enumerated these cellular fragments as platelets, resulting in falsely elevated platelet counts. Awareness of this laboratory artifact in secondary acute monocytic leukemia with tumor lysis syndrome is important so that potential life-threatening thrombocytopenia is not overlooked. PMID- 10539920 TI - Squamous cell carcinoma arising in a ciliated hepatic foregut cyst. AB - Ciliated hepatic foregut cysts are rare congenital lesions derived from the embryologic foregut. They are considered benign, and a review of 64 published cases revealed no instances of malignant transformation. We report a case of squamous cell carcinoma arising in a ciliated hepatic foregut cyst in a 51-year old man. The tumor was found during a routine cholecystectomy and involved the adjacent mesentery and duodenal wall. There was histologic evidence of perineural and perivascular involvement. Despite an en bloc resection of the tumor and contiguous areas of gross involvement, the patient died 2 months later. Although aspiration of cyst contents is an accepted treatment for asymptomatic lesions, this case suggests that most ciliated hepatic foregut cysts should be excised, especially when radiologic studies yield equivocal results. PMID- 10539921 TI - Familial occurrence of acinic cell carcinoma of the parotid gland. AB - We report the familial occurrence of acinic cell carcinoma involving the parotid gland, the first such report of which we are aware. The familial occurrence of any salivary gland neoplasm is rare. Several reports are present in the literature, including pleomorphic adenoma, Warthin tumor, carcinoma of the submandibular gland, and malignant lymphoepithelial lesion. We report the case of a 35-year-old man who underwent excision of a left parotid gland acinic cell carcinoma. Eight years later, his daughter presented at the age of 16 years with a nontender parotid gland mass that was excised and found also to be acinic cell carcinoma. The histologic features of both neoplasms were typical of acinic cell carcinoma. While this may represent a coincidental event, the possibility that this familial occurrence is a manifestation of common genetic or environmental risk cannot be excluded. PMID- 10539922 TI - Pathologic quiz case: an unusual infection in a human immunodeficiency virus positive man. Pathologic diagnosis: rhinosporidiosis. PMID- 10539923 TI - Barrett esophagus-associated small cell carcinoma. PMID- 10539925 TI - Clinical hematology atlas PMID- 10539926 TI - Serum folate and homocysteine concentrations in large population samples of US ethnic and racial groups. PMID- 10539928 TI - Why cholesterol-lowering diets should still be encouraged in the face of effective pharmaceutical interventions. PMID- 10539931 TI - Fatal pulmonary fibrosis after a low cumulated dose of bleomycin: role of alpha1 antitrypsin deficiency? PMID- 10539932 TI - Practice guidelines for autopsy pathology: autopsy reporting. Autopsy Committee of the College of American Pathologists. AB - The Autopsy Committee of the College of American Pathologists has prepared this revised guideline to reflect changes that have occurred in the reporting of autopsies since the original guideline was published in February 1995. It is intended to be an instrument to assist pathologists in the reporting of autopsies. The guideline is to be regarded as being primarily an educational tool. Application of these recommendations on autopsy reporting is to be made on the basis of the judgment of the pathologist engaged in a specific case. PMID- 10539941 TI - The Raw and the Stolen. Cooking and the Ecology of Human Origins. AB - Cooking is a human universal that must have had widespread effects on the nutrition, ecology, and social relationships of the species that invented it. The location and timing of its origins are unknown, but it should have left strong signals in the fossil record. We suggest that such signals are detectable at ca. 1.9 million years ago in the reduced digestive effort (e.g., smaller teeth) and increased supply of food energy (e.g., larger female body mass) of early Homo erectus. The adoption of cooking required delay of the consumption of food while it was accumulated and/or brought to a processing area, and accumulations of food were valuable and stealable. Dominant (e.g., larger) individuals (typically male) were therefore able to scrounge from subordinate (e.g., smaller) individuals (typically female) instead of relying on their own foraging efforts. Because female fitness is limited by access to resources (particularly energetic resources), this dynamic would have favored females able to minimize losses to theft. To do so, we suggest, females formed protective relationships with male co defenders. Males would have varied in their ability or willingness to engage effectively in this relationship, so females would have competed for the best food guards, partly by extending their period of sexual attractiveness. This would have increased the numbers of matings per pregnancy, reducing the intensity of male intrasexual competition. Consequently, there was reduced selection for males to be relatively large. This scenario is supported by the fossil record, which indicates that the relative body size of males fell only once in hominid evolution, around the time when H. erectus evolved. Therefore we suggest that cooking was responsible for the evolution of the unusual human social system in which pair bonds are embedded within multifemale, multimale communities and supported by strong mutual and frequently conflicting sexual interest. PMID- 10539942 TI - Socioeconomic Growth, Culture Scale, and Household Well-Being. A Test of the Power-Elite Hypothesis. AB - Socioeconomic growth is an elite-directed process that concentrates social power in direct proportion to increases in culture scale. Power elites have controlled social power to their own advantage in at least three different ways: domestically, by means of kinship, politically, by means of rulers, and commercially, by means of the market. Each method produces its own growth trajectory and scale of culture and a distinctive distribution of elite power and household living standards. Ethnographic data on urban property ownership in 27 municipalities in the Palouse region of eastern Washington suggest that when power is commercially organized and villages become towns and cities, there is a dramatic increase not only in the number of prosperous households but even more in the number of poor and maintenance-level households. Elite property owners, who most benefit from growth, assume a larger role in municipal government, where they can encourage further growth through municipal annexations and zoning changes. Thus, as elite power becomes increasingly concentrated, the growth process itself tends to become self-perpetuating. In the Palouse example, small, no-growth municipalities appear to be politically more democratic than larger scale, growing municipalities and household well-being in them more equitably distributed. PMID- 10539943 TI - Are East African Pastoralists Truly Conservationists? AB - Controversy exists among anthropologists, conservation biologists, and development workers as to whether the concept of the "ecologically noble savage" is a myth. Central to this debate are the problem of how to identify conservationist behavior and the issue of whether sound management of common property is likely to evolve. While social scientists have documented instances of restraint in the use of resources, those who adopt an evolutionary perspective are challenged to identify the selective mechanisms whereby such altruistic conservation acts might be maintained in a population. Here a game-theoretical approach is used to analyze the case of pastoralist grazing reserves. We demonstrate that under some conditions conservation can be the result of narrow self-interest and there is no collective-action problem. However, the range of these conditions is much broader for wealthy individuals, and thus the wealthy may also find it advantageous to coerce others into conserving. In conclusion, we propose an extension of the definition of conservation that is of greater generality for use in nonforaging populations and incorporates the essential political element of how conflicts over resource use are resolved. PMID- 10539945 TI - Rhinoceros 2. PMID- 10539944 TI - Archaeological Narratives and Other Ways of Telling. AB - With a few exceptions, archaeologists have been far less concerned with the form of their texts or problems of authorship than have ethnographers. Typically, archaeologies are presented in the form of narratives understood as sequential stories. Approaches to narrative analysis drawn from literary theory, philosophy, and sociology and definitions of characters, events, and plots are examined, together with particular problems these may pose for the discipline of archaeology. It is suggested that neither literary analysis nor the tendency to write and evaluate archaeological and historical narratives in terms of explanatory value takes sufficient account of the often hybrid nature and aims of these texts and the contexts in which they were produced. This argument is illustrated with particular reference to stories of the Mesolithic-Neolithic transition in Europe. It is argued that reconsidering archaeology's positioning across the 19th-century science-humanities divide suggests a broader approach to the idea of what constitutes a narrative which can offer fresh opportunities for useful reflexivity and experimentation in presentation. Further roles and possibilities of narrative and non-narrative ways of writing archaeologies are also considered. PMID- 10539946 TI - On Diet and Gut Size in Non-human Primates and Humans: Is There a Relationship to Brain Size? PMID- 10539947 TI - On Regional Geochemistry and Statistical Method. PMID- 10539948 TI - An Interview with Sydel Silverman. PMID- 10539949 TI - Aurignacian Settlement Patterns in the Vezere Valley. PMID- 10539950 TI - Population History and the Islamization of the Iberian Peninsula: Skeletal Evidence from the Lower Alentejo of Portugal. PMID- 10539951 TI - Meat Eating and Hominid Evolution. PMID- 10539952 TI - When Nomads Settle: The Effects of Commoditization, Nutritional Change, and Formal Education on Ariaal and Rendille Pastoralists. PMID- 10539954 TI - The biomechanics of fast-starts during ontogeny in the common carp cyprinus carpio AB - Common carp Cyprinus carpio L. were reared a constant temperature of 20 degrees C from the larval (7 mm total length) to the juvenile (80 mm) stage. Body morphology and white muscle mass distribution were measured. Fast-start escape responses were recorded using high-speed cinematography from which the velocities, accelerations and hydrodynamic power requirements were estimated. All three measures of fast-start performance increased during development. White muscle contraction regimes were calculated from changes in body shape during the fast-starts and used to predict the muscle force and power production for all longitudinal positions along the body. Scaling arguments predicted that increases in body length would constrain the fish to bend less rapidly because the cross sectional muscle area, and hence force production, does not increase at the same rate as the inertial mass that resists bending. As predicted, the increases in body length resulted in decreases in muscle shortening velocity, and this coincided with increases in both the force and power produced by the muscles. The hydrodynamic efficiency, which relates the mechanical power produced by the muscles to the inertial power requirements in the direction of travel, showed no significant change during ontogeny. The increasing hydrodynamic power requirements were thus met by increases in the power available from the muscles. The majority of the increases in fast-start swimming performance during ontogeny can be explained by size-dependent increases in muscle power output. For all sizes, there was a decrease in muscle-mass-specific power output and an increase in muscle stress in a posterior direction along the body due to systematic variations in fibre strain. These changing strain regimes result in the central muscle bulk producing the majority of the power requirements during the fast start, and this power is transmitted to the tail region of the fish and ultimately to the water via muscle in the caudal myotomes. PMID- 10539953 TI - Primitive organization of cytosolic Ca(2+) signals in hepatocytes from the little skate Raja erinacea. AB - Cytosolic Ca(2+) (Ca(i)(2+)) signals begin as polarized, inositol 1, 4,5 trisphosphate (InsP3)-mediated Ca(i)(2+) waves in mammalian epithelia, and this signaling pattern directs secretion together with other cell functions. To investigate whether Ca(i)(2+) signaling is similarly organized in elasmobranch epithelia, we examined Ca(i)(2+) signaling patterns and InsP3 receptor (InsP3R) expression in hepatocytes isolated from the little skate, Raja erinacea. Ca(i)(2+) signaling was examined by confocal microscopy, InsP3R expression by immunoblot, and the subcellular distribution of InsP3Rs by immunochemistry. ATP induced a rapid increase in Ca(i)(2+) in skate hepatocytes, as it does in mammalian hepatocytes. Unlike in mammalian hepatocytes, however, the Ca(i)(2+) increase in skate hepatocytes began randomly throughout the cell rather than in the apical region. In cells loaded with heparin ATP-induced Ca(i)(2+) signals were inhibited, but de-N-sulfated heparin was not inhibitory, suggesting that the increases in Ca(i)(2+) were mediated by InsP3. Immunoblot analysis showed that the type I but not the types II or III InsP3R was expressed in skate liver. Confocal immunofluorescence revealed that the InsP3R was distributed throughout the hepatocyte, rather than concentrated apically as in mammalian epithelia. These findings demonstrate that ATP-induced Ca(i)(2+) signals are mediated by InsP3 in skate hepatocytes, as they are in mammalian hepatocytes. However, in skate hepatocytes Ca(i)(2+) signals begin at loci throughout the cell rather than as an organized apical-to-basal Ca(i)(2+) wave, which is probably because the InsP3R is distributed throughout these cells. This primitive organization of Ca(i)(2+) signaling may in part explain the observation that Ca(2+)-mediated events such as secretion occur much less efficiently in elasmobranchs than in mammals. PMID- 10539955 TI - The relationship between leg stepping pattern and yaw torque oscillations in curve walking of two crayfish species AB - Curve walking in two species of crayfish, Procambarus clarkii and Astacus leptodactylus, was investigated to test whether the mechanism underlying curve walking is the synchronous action of a centrally pre-programmed leg tripod or whether it is the action of one principal leg that produces the main body yaw torque. Curve walking was induced by an optomotor visual stimulus, and the yaw torque produced by the tethered animals was measured in open-loop conditions. Our main results suggest that the yaw torque oscillations in both P. clarkii and A. leptodactylus are related to the movement of outer leg 4 (i.e. leg 4 on the outside of the turn). That is, the peaks in the yaw torque occur, on average, in synchrony with the power stroke of outer leg 4. When comparing the results of this open-loop experiment on P. clarkii with results previously obtained for curve walking in untethered individuals of the same species, we found a much higher variability in leg coordination in the open-loop situation. Similarly, here we did not find the same level of synchrony in the tripod (formed by outer leg 4 and inner legs 2 and 5) observed during untethered free walking. Therefore, we suggest that tethered conditions may diminish the need for stability and thus allow outer leg 4 to produce a body rotation regardless of the leg stepping configuration. The characteristics of leg 4 are in line with its major role in turning. According to previous studies, legs 4 provide the largest force and the largest step amplitude during walking, and their force includes both a pulling and a pushing component which can facilitate the control of turning. Although it is apparent that outer leg 4 is not the only leg that can produce an inward yaw torque, its major role in modulating the yaw torque suggests that there may be a specific, centrally generated control of outer leg 4 during curve walking in crayfish. PMID- 10539957 TI - The sites of respiratory gas exchange in the planktonic crustacean daphnia magna: an in vivo study employing blood haemoglobin as an internal oxygen probe AB - Recent studies on Daphnia magna have revealed that the feeding current is important for uptake of oxygen from the ambient medium. Respiratory gas exchange should therefore mainly occur within the filtering chamber, whose boundaries are formed by the trunk and the extended carapace shell valves. The precise site of gas exchange in the genus Daphnia is, however, a matter of conjecture. We have developed a method of imaging the haemoglobin oxygen-saturation in the circulatory system of transparent animals, which provides an opportunity to localize oxygen uptake from the environment and oxygen release to the tissues. Experiments were carried out at 20 degrees C on 2.8-3.0 mm long parthenogenetic females maintained in hypoxic culturing conditions, which had resulted in an increased haemoglobin content in the haemolymph. In lateral views of D. magna, the highest values of haemoglobin oxygen-saturation occurred near the posterior margin of the carapace and, surprisingly, in the rostral part of the head. The ambient oxygen partial pressures at which haemoglobin was half-oxygenated were 15 mmHg (2.0 kPa) for the posterior carapace region and 6 mmHg (0.8 kPa) for the rostrum. Although not all parts of the circulatory system could be analyzed using this technique, the data obtained from the accessible regions suggest that the inner wall of the carapace is a major site of respiratory gas exchange. Taking the circulatory pattern and the flow pattern of the medium in the filtering chamber into consideration, it becomes clear that the haemolymph, after passing from the limbs to the carapace lacuna, becomes oxygenated while flowing through the ventral part of the double-walled carapace in a posterior direction. The laterally flattened rostral region, where sensory and central nervous system structures are located, seems to have direct diffusive access to ambient oxygen, which could be especially advantageous during severe hypoxia when the convective transport systems fail to supply enough oxygen to that region. PMID- 10539956 TI - The effects of intensity on the energetics of brief locomotor activity. AB - The energetic costs associated with locomotion are often estimated only from the energy expended during activity and do not include the costs incurred during recovery. For some types of locomotion, this method overlooks important aspects of the metabolic costs incurred as a result of the activity. These estimates for energetic cost have also been predicted from long-duration, low-intensity activities that do not necessarily reflect all the behavior patterns utilized by animals in nature. We have investigated the effects of different activity intensities on the metabolic expenditure (per unit distance traveled) associated with brief exercise, and offer a more inclusive analysis of how the energetics of short-duration activities might be analyzed to estimate the costs to the animal. Mice ran on a treadmill for 15 or 60 s at 25 %, 50 % or 100 % of maximum aerobic speed (MAS) while enclosed in an open-flow respirometry system. Following the run, each mouse was allowed to recover while remaining enclosed in the respirometry chamber. Excess exercise oxygen consumption (EEOC), the excess volume of oxygen consumed during the exercise period, increased with the duration and increased linearly with the intensity of exercise. In contrast, the volume of oxygen consumed during the recovery period, or excess post-exercise oxygen consumption (EPOC), was independent of exercise intensity and duration and accounted for more than 90 % of the total metabolic cost. The net cost of activity (C(act)), calculated by summing EEOC and EPOC and then dividing by the distance run, increased as both activity duration and intensity decreased. The values for C(act) ranged from 553 ml O(2 )g(-)(1 )km(-)(1) for a 15 s run at 25 % MAS to 43 ml O(2 )g(-)(1 )km(-)(1) for a 60 s run at 100 % MAS. Combining these data with data from a companion paper, we conclude (1) that EPOC is independent of both the duration and intensity of activity when exercise duration is brief in mice, (2) that EPOC accounts for a majority of the oxygen consumed as a result of the activity when exercise durations are short, and (3) that animals can minimize their energy expenditure per unit distance by running faster for a longer period. PMID- 10539958 TI - Morphological basis of kinematic diversity in feeding sunfishes AB - The effects of differences among species in the scaling of lower jaw levers on the scaling of prey-capture kinematics are explored in three species of centrarchid fishes. We consider the jaw opening and closing lever systems and calculate the consequences of differences in the scaling of the in-levers for the scaling of the time taken to open the mouth (T(o)) and the time taken to close the mouth (T(c)) during prey capture. Predictions of T(o) and T(c), based on differences in the scaling of jaw in-levers, are compared with the observed scaling of T(o) and T(c) in three centrarchid fishes. Video recordings (200 and 400 images s(-)(1)) were made of prey capture in largemouth bass Micropterus salmoides (33-206 mm standard length, SL), spotted sunfish Lepomis punctatus (24 145 mm SL) and bluegill sunfish Lepomis macrochirus (24-220 mm SL), and the fastest values of T(o) and T(c) were taken from the fastest recorded feeding event for each fish. The scaling exponents of T(o) and T(c) regressed on fish SL for largemouth bass were 0.592 and 0.572, respectively. Exponents observed for sunfishes were not significantly different from predicted values, based on scaling exponents in largemouth bass and interspecific differences in jaw lever proportions. Two conclusions are emphasized. First, between 25 and 220 mm SL, the time taken to open and close the mouth during the strike increases with body size in all three species, suggesting a general pattern for this family. Second, evolutionary changes in jaw lever mechanics are a major determinant of the diversity of prey-capture kinematics in this sample of centrarchid fishes. PMID- 10539959 TI - Shear stress experienced by echinoderm eggs in the oviduct during spawning: potential role in the evolution of egg properties AB - Shear stresses experienced by eggs in the oviduct of the echinoid Arbacia punctulata during spawning were calculated using engineering equations that describe laminar flow through pipes. Shear stresses in the oviduct ranged from 0 to 58.7 Pa. Two properties of eggs were identified that have the potential either to minimize the shear stress in the oviduct or to reduce the damage experienced by eggs exposed to high shear stress. These properties are the viscosity of the eggs and the presence of extracellular layers on eggs of A. punctulata. The viscosity of eggs decreases with increasing shear rates, which reduces the magnitude of shear stress experienced in the oviduct, while the extracellular layers mitigate the effect of shear stress on the eggs. Eggs with intact extracellular layers were damaged less frequently than were those with the extracellular layers removed. The results of this research indicate that physical stresses may be important selective factors in the evolution of gamete properties. PMID- 10539960 TI - Precise monitoring of porpoising behaviour of Adelie penguins determined using acceleration data loggers. AB - A new method using acceleration data loggers enabled us to measure the porpoising behaviour of Adelie penguins (Pygoscelis adeliae), defined as a continuous rapid swimming with rhythmic serial leaps. Previous hydrodynamic models suggested that leaping would be energetically cheaper when an animal swims continuously at depths of less than three maximum body diameters below the water surface. In the present study, free-ranging Adelie penguins leapt at a mean speed of 2.8 m s( )(1) above the predicted threshold speed (0.18-1. 88 m s(-)(1)). Wild penguins reduced drag by swimming deeper (0.91 m) and did not swim continuously within the high-drag layer while submerged. This indicates that previous calculations may be incomplete. Moreover, leaps represented an average of only 3.8 % of the total distance travelled during the porpoising cycle, which would make energy savings marginal. Among the six penguins used in our study, two did not porpoise and three porpoised for less than 7 min, also indicating that this behaviour was not important during travel to and from foraging sites, as has been previously suggested. Birds mainly porpoised at the start and end of a trip. One explanation of porpoising might be an escape behaviour from predators. PMID- 10539961 TI - Accommodation in the cuttlefish (Sepia officinalis). AB - We have studied natural accommodation in the eye of six specimens of cuttlefish (Sepia officinalis) as they were fed with fish and shrimp. Using infrared photoretinoscopy, we observed (1) that the resting refractive state of the cuttlefish was emmetropic or slightly hyperopic, (2) that accommodation took place only a fraction of a second before a strike and (3) that accommodation focused selectively only in the frontal visual field while no change in refraction could be measured in the lateral field of view. Accommodation was bilateral and amounted to approximately 5 diopters (the reciprocal of the focal length expressed in meters). Simultaneously, the eyes converged. It appears that, as in most teleost fishes, accommodation in the cuttlefish involves a movement of the crystalline lens perpendicular to the axis of the eye. In histological sections, we observed the position and arrangement of the ciliary muscles, confirming earlier anatomical descriptions, and developed a model of how accommodation could be achieved. PMID- 10539962 TI - Reactive oxygen intermediate production by oyster hemocytes exposed to hypoxia AB - Oysters are frequently exposed to severely hypoxic conditions, especially during summer months. During the summer, there are also large numbers of disease-related oyster mortalities. This research was conducted to determine whether exposure to environmental hypoxia reduces the ability of oyster hemocytes to produce reactive oxygen intermediates (ROIs), an important part of their defense system. Oysters of the species Crassostrea virginica were held in normoxic (P(O)(2)=20.0-20.7 kPa, pH 7.8-8.0) and hypoxic conditions (P(O)(2)=4.0-6.7 kPa, pH 7.1-7.4). In vivo hemolymph variables (P(O)(2), P(CO)(2) and pH) were measured after both 1 hour and 2 days in each treatment to determine the appropriate environment for subsequent hemocyte experiments. Production of reactive oxygen intermediates by hemocytes was measured using luminol-enhanced chemiluminescence (CL). During CL tests, hemocytes were held under the following conditions: air (P(O)(2)=20.7, P(CO)(2)<0.07, pH 7.6), in vivo hemolymph conditions of normoxic oysters (P(O)(2)=5.2, P(CO)(2)=0.27, pH 7.6), and in vivo hemolymph conditions of hypoxic oysters (P(O)(2)=1.47, P(CO)(2)=0.53, pH 7.1). Production of ROIs under hypoxic conditions was 33 % of that under normoxia. This decrease was the result of specific and independent effects of lower oxygen levels and decreased pH. It was not due to any direct effect of CO(2). PMID- 10539964 TI - Ferromagnetic material in the eastern red-spotted newt notophthalmus viridescens AB - Behavioral results obtained from the eastern red-spotted newt (Notophthalmus viridescens) led to the suggestion of a hybrid homing system involving inputs from both a light-dependent and a non-light-dependent mechanism. To evaluate the possible role of a receptor based on biogenic magnetite in this animal, we performed magnetometry experiments on a set of newts previously used in behavioral assays. The natural remanent magnetization (NRM) carried by these newts was strong enough to be measured easily using a direct-current-biased superconducting quantum interference device functioning as a moment magnetometer. Isothermal remanent magnetizations were two orders of magnitude higher than the NRM, suggesting that ferromagnetic material consistent with magnetite is present in the body of the newt. The NRM has no preferential orientation among the animals when analyzed relative to their body axis, and the demagnetization data show that, overall, the magnetic material grains are not aligned parallel to each other within each newt. Although the precise localization of the particles was not possible, the data indicate that magnetite is not clustered in a limited area. A quantity of single-domain magnetic material is present which would be adequate for use in either a magnetic intensity or direction receptor. Our data, when combined with the functional properties of homing, suggest a link between this behavioral response and the presence of ferromagnetic material, raising the possibility that magnetite is involved at least in the map component of homing of the eastern red-spotted newt. PMID- 10539963 TI - Flight-muscle adenylate pool responses to flight demands and thermal constraints in individual Colias eurytheme (Lepidoptera, pieridae). AB - We study here the connections among body temperature variation, flight performance and flight 'fuel' metabolism in Colias eurytheme butterflies, to begin re-examining the metabolic reasons for animal thermoregulation. Methods are presented for (a) stable extraction of adenylates (and other metabolites) from the flight muscles of individual Colias eurytheme, (b) automated separation and quantitative analysis of individual adenylate samples by high-pressure liquid chromatography and (c) reliable, low-variance assay of inorganic phosphate levels in the same extracts. Correlations among metabolite concentrations and two indices of muscle cytosol ATP maintenance occur as expected on general metabolic principles. [ATP] and [ATP]/[ADP] decline from resting levels to reach a plateau in the first minute of free, interrupted flight, while [AMP] increases at the same time; these concentrations do not vary further for up to 6 min total flight time. In an initial test of the alternative metabolic bases of the thermoregulation of Colias eurytheme, we find that [ATP]/[ADP] rises between a body temperature, T(b), of 31 and 35 degrees C, at the base of the behavioral thermal optimum for flight, but then decreases again at T(b)=39 degrees C, at the top of the behavioral thermal optimum and well short of damaging temperatures. This is not consistent with the view that metabolic effectiveness increases monotonically up to the lower limits of thermal damage to enzymes, but supports an alternative hypothesis that the narrowness of thermoregulation results from a system-based constraint on the breadth of temperature over which maximal energy processing is possible. PMID- 10539965 TI - Temperature-dependence of neuronal performance in the motion pathway of the blowfly calliphora erythrocephala AB - Raising the head temperature within a behaviourally relevant range has strong effects on the performance of an identified neuron, the H1 neuron, in the visual motion pathway of blowflies. The effect is seen as an increase in the mean amplitude of the responses to motion under both transient and steady-state conditions, a considerable decrease in the response latency and an improvement in the reliability of the responses to motion. These temperature-dependent effects are independent of whether the animal is exposed to transient temperature changes or is maintained continuously at the same temperature for its entire life. The changes in the neuronal response properties with temperature may be of immediate functional significance for the animal under its normal operating conditions. In particular, the decrease in latency and the improvement in the reliability with increasing temperature may be relevant for the fly when executing its extremely virtuosic flight manoeuvres. PMID- 10539966 TI - Phenotypic flexibility of the avian gizzard: rapid, reversible and repeated changes of organ size in response to changes in dietary fibre content AB - Evolutionary biology presumes that organ capacities match their natural loads. Therefore, in fluctuating conditions, organ systems are expected to show a reversible, repeatable and rapid phenotypic response that is directional and scaled. In this study, phenotypic responses of the gizzard of adult Japanese quail (Coturnix japonica) to experimental mismatches of load and capacity were tested by a series of diet-switching experiments, involving an increased content of non-digestable fibre (NDF) in the diet. The results of all experiments were in accordance with the predictions of the hypothesis that there is matching between loads and capacities. (1) The observed phenotypic responses are directional and scaled to the demands, i.e. increasing NDF elicits an increase in gizzard size. When the proportion of NDF in the diet was raised from 1 % to 45 %, the gizzard was more than twice as large as in the control group. (2) Size responses were reversible, and reduced NDF was followed by a decrease of gizzard size. (3) Phenotypic responses could be elicited repeatedly in three successive trials. (4) Excess capacities were downregulated and insufficient capacities were upregulated. (5) The responses followed changes of loads with almost no time lag, with size changes measurable within 24 h. PMID- 10539967 TI - The scaling of acceleratory aquatic locomotion: body size and tail-flip performance of the california spiny lobster panulirus interruptus AB - Tail-flipping is a crucial escape locomotion of crustaceans which has been predicted to be limited by increased body mass (M(b)). Given isometric growth, one may predict that with growth event duration will decrease as M(b)(-)(1/3), translational distances will increase as M(b)(1/3), translational velocity will be independent of M(b), translational acceleration will decrease as M(b)(-)(1/3), angular displacement will be independent of M(b) and angular velocity and angular acceleration will decrease as M(b)(-)(1/3). We tested these hypotheses by examining the scaling of 12 morphological variables, five kinematic variables and six performance variables of tail-flipping by the California spiny lobster Panulirus interruptus. Growth approximated isometry, which validated the use of the proposed scaling hypotheses. For animals from 1 to 1000 g M(b), the predicted scaling relationships for tail-flip duration and translational distance and velocity variables were supported; however, translational acceleration performance was much better than predicted. Predictions for rotation and rotational velocity variables were not supported, while the rotational acceleration data closely matched the predicted relationship. The increase in tail-flip duration as predicted suggests that muscle shortening velocity decreases with growth; the sustained acceleration performance (similar to findings for shrimp and fish fast-starts) suggests that muscle force output may increase at a greater rate than predicted by isometry. The scaling of rotational acceleration indicates that the torque produced during the tail-flip scales with a mass exponent greater than 1. Comparison of the tail-flip performance of Panulirus interruptus with those of other crustacean species reveals a wide range in performance by animals of similar body size, which suggests that the abdominal muscle may show interesting differences in contractile properties among different species. PMID- 10539968 TI - Target organ specificity of major neuropeptide stimulants in locust excretory systems AB - The major stimulant of ileal fluid reabsorption in Locusta migratoria and Schistocerca gregaria corpora cardiaca, ion-transport peptide (ITP), had no stimulatory action on fluid secretion by isolated Malpighian tubules of S. gregaria, nor did it have a synergistic or antagonistic effect in combination with locustakinin (Lom-K) or Locusta-diuretic hormone (Locusta-DH). Stimulants of locust Malpighian tubules (Lom-K and Locusta-DH) had no action on either active transport of Cl(-) (measured as short-circuit current, I(sc)) or the rate of fluid reabsorption across S. gregaria ilea and recta in vitro. Thus, hormonal control of these major organs of the excretory system appears to be clearly separated. Lom-K and Locusta-DH acted synergistically to stimulate secretion by S. gregaria Malpighian tubules, and the diuretic response was more rapid than the response of the ileum and rectum to hindgut stimulants. Taken together, these data suggest that, in the initial phase of post-prandial diuresis, urine flow will exceed fluid uptake in the hindgut, thereby allowing excess water to be eliminated. PMID- 10539969 TI - Cardiorespiratory response to progressive hypoxia and hypercapnia in the turtle trachemys scripta AB - The ventilatory responses of chelonian reptiles to hypoxic and hypercapnic stress have been fairly well described. As turtles are capable of large cardiac shunts, changes in pulmonary perfusion may be an equally viable and potent response to these stressors. To test this hypothesis, conscious unrestrained turtles were unidirectionally ventilated while blood flow in the left pulmonary artery ( q_dot (LPA)) and left aortic arch ( q_dot (LAo)) was monitored. Turtles were exposed to step changes (2.5 h step(-)(1)) in O(2) tension (30, 15, 5, 2.5 or 0 % O(2); CO(2) inflow maintained constant) on day 1 followed by step changes in CO(2) tension (0, 2, 4, 8 % CO(2); O(2) inflow maintained constant) on day 2. Steady state cardiorespiratory variables were recorded for the last 30 min of each step change in gas tension.Progressive hypoxia resulted in progressive increases in ventilation, q_dot (LPA) and q_dot (LAo) and a small, but non-significant, increase in heart rate. Progressive hypercapnia resulted in a progressive increase in ventilation, while q_dot (LPA) and q_dot (LAo) did not change at any level of CO(2). These results suggest that information from the O(2)-sensitive chemoreceptors appears to be stimulatory to both the cardiovascular and ventilatory control systems, while CO(2) chemoreception appears to affect primarily the ventilatory control system. These results also suggest that, in animals capable of intracardiac shunting, increasing pulmonary perfusion may be an integral component of the reflex response to hypoxia. PMID- 10539970 TI - Contractile properties of muscles used in sound production and locomotion in two species of gray tree frog. AB - The sound-producing muscles of frogs and toads are interesting because they have been selected to produce high-power outputs at high frequencies. The two North American species of gray tree frog, Hyla chrysoscelis and Hyla versicolor, are a diploid-tetraploid species pair. They are morphologically identical, but differ in the structure of their advertisement calls. H. chrysoscelis produces very loud pulsed calls by contracting its calling muscles at approximately 40 Hz at 20 degrees C, whereas, H. versicolor operates the homologous muscles at approximately 20 Hz at this temperature. This study examined the matching of the intrinsic contractile properties of the calling muscles to their frequency of use. I measured the isotonic and isometric contractile properties of two calling muscles, the laryngeal dilator, which presumably has a role in modulating call structure, and the external oblique, which is one of the muscles that provides the mechanical power for calling. I also examined the properties of the sartorius as a representative locomotor muscle. The calling muscles differ greatly in twitch kinetics between the two species. The calling muscles of H. chrysoscelis reach peak tension in a twitch after approximately 15 ms, compared with 25 ms for the same muscles in H. versicolor. The muscles also differ significantly in isotonic properties in the direction predicted from their calling frequencies. However, the maximum shortening velocities of the calling muscles of H. versicolor are only slightly lower than those of the comparable muscles of H. chrysoscelis. The calling muscles have similar maximum shortening velocities to the sartorius, but have much flatter force-velocity curves, which may be an adaptation to their role in cyclical power output. I conclude that twitch properties have been modified more by selection than have intrinsic shortening velocities. This difference corresponds to the differing roles of shortening velocity and twitch kinetics in determining power output at differing frequencies. PMID- 10539971 TI - Power output of sound-producing muscles in the tree frogs Hyla versicolor and Hyla chrysoscelis. AB - Sound-producing muscles provide the opportunity of studying the limits of power production at high contractile frequencies. We used the work loop technique to determine the power available from the external oblique muscles in two related species of North American gray tree frog, Hyla chrysoscelis and Hyla versicolor. These trunk muscles contract cyclically, powering high-intensity sound production in anuran amphibians. The external oblique muscles in H. chrysoscelis have an in vivo operating frequency of 40-55 Hz at 20-25 degrees C, whereas in H. versicolor these muscles contract with a frequency of 20-25 Hz at these temperatures. In vivo investigations have shown that these muscles use an asymmetrical sawtooth length trajectory (with a longer shortening phase compared with the lengthening phase) during natural cycles. To study the influence of this particular length trajectory on power output, we subjected the muscles to both sinusoidal and sawtooth length trajectories. In both species, the sawtooth trajectory yielded a significantly higher power output than the sinusoidal length pattern. The maximum power output during sawtooth cycles was similar in both species (54 W kg(-)(1) in H. chrysoscelis and 58 W kg(-)(1) in H. versicolor). These values are impressive, particularly at the operating frequencies and temperatures of the muscle. The sinusoidal length trajectory yielded only 60 % of the total power output compared with the sawtooth trajectory (34 W kg(-)(1) for H. chrysoscelis and 36 W kg(-)(1) for H. versicolor). The optimum cycle frequencies maximizing the power output using a sawtooth length pattern were approximately 44 Hz for H. chrysoscelis and 21 Hz for H. versicolor. These frequencies are close to those used by the two species during calling. Operating at higher frequencies, H. chrysoscelis maximized power at a strain amplitude of only 8 % compared with a value of 12 % in H. versicolor. These strains match those used in vivo during calling. The stimulus timing observed in vivo during calling was also similar to that yielding maximum power at optimal frequency in both species (6 ms and 8 ms before the start of shortening in H. chrysoscelis and H. versicolor, respectively). As expected, twitch duration in H. chrysoscelis is much shorter than that in H. versicolor (23 ms and 37 ms, respectively). There was a less remarkable difference between their maximum shortening velocities (V(max)) of 13.6 L(0 )s( )(1) in H. chrysoscelis and 11.1 L(0 )s(-)(1) in H. versicolor, where L(0) is muscle length. The force-velocity curves are very flat, which increases power output. At the myofibrillar level, the flat force-velocity curves more than compensate for the lower peak isometric force found in these muscles. The data presented here emphasize the importance of incorporating in vivo variables in designing in vitro studies. PMID- 10539972 TI - Loading conditions and cortical bone construction of an artiodactyl calcaneus. AB - Customary nonuniform distributions of physiological bone strains are thought to evoke heterogeneous material adaptation in diaphyseal cortices of some limb bones. Recent studies of artiodactyl calcanei have suggested that the regional prevalence of specific mechanical strain features such as mode and magnitude correlate with specific variations in cortical bone ultrastructure, microstructure and mineralization. These data are also consistent with predictions of current algorithms of mechanically induced bone adaptation. However, detailed characterization of the customary functional strain environment of these bones is needed to understand better the mechanisms of these adaptations. An in vitro loading method and rosette strain gauges were used to record principal strains, maximum shear strains and principal strain angles at multiple locations on ten calcanei of adult male mule deer (Odocoileus hemionus hemionus). Each hind limb was fixed in an apparatus to mimic the mid-support phase of the gait and loaded via the Achilles tendon over a broad range of functional loads (0 to 2943 N). Strains were recorded on the craniolateral, craniomedial, caudal, medial and lateral cortices at mid-diaphysis. Loading variations included the progressive elimination of the ligament and tendon along the caudal calcaneus. The results showed that the cranial cortex experiences longitudinal compressive strains that are nearly equal to the principal minimum strains and that the caudal cortex receives longitudinal tensile strains that are nearly equal to the principal maximum strains. With a 981 N load, the mean principal compressive strain on the cranial cortex was -636+/-344 micro(&egr;) (mean +/- s.d., N=9) and the mean principal tensile strain on the caudal cortex was 1112+/-68 micro;(&egr;)x (N=9). In contrast to the cranial and caudal cortices, principal strains in the medial and lateral cortices displayed relatively large deviations from the longitudinal axis (medial, 24 degrees cranial; lateral, 27 degrees caudal). Although shear strains predominated at all gauge sites, variations in maximum shear strains showed no apparent regional pattern or consistent regional predominance. The plantar ligament and tendon of the superficial digital flexor muscle were shown to have important load-sharing functions. These results demonstrate that the functionally loaded artiodactyl calcaneus generally behaves like a cantilevered beam with longitudinal compression and tension strains predominating in opposing cranial and caudal cortices, respectively. Differences in osteon remodeling rates, osteon morphology and mineral content reported previously between the cranial and caudal cortices correlate, in part, with the magnitudes of the principal compressive and tensile strains, respectively. However, material differences that distinguish the medial and lateral cortices from the cranial and caudal cortices could not be primarily attributed to locally increased shear strains as previously suggested. Variations in osteon and/or collagen fiber orientation may correlate more strongly with principal strain direction. PMID- 10539973 TI - Effects of lipid phase transitions on cuticular permeability: model membrane and in situ studies AB - The role of lipid physical properties in cuticular water loss was examined in model membranes and intact insects. In model experiments, pure hydrocarbons of known melting point (T(m)) were applied to a membrane, and the effects of temperature on permeability were quantified. Arrhenius plots of permeability were biphasic, and transition temperatures for water loss (T(c)) were similar to T(m). In grasshoppers Melanoplus sanguinipes, changes in cuticular water loss were measured using flow-through respirometry. Transition temperatures were determined and compared with T(m) values of cuticular lipids, determined using Fourier transform infrared spectroscopy, for the same individuals. Individual variation in T(m) was highly correlated with T(c), although T(m) values were slightly higher than T(c) values. Our results show that, in both intact insects and model membranes, lipid melting results in greatly increased cuticular permeability. PMID- 10539974 TI - Ionic regulatory properties of brain and kidney splice variants of the NCX1 Na(+) Ca(2+) exchanger. AB - Ion transport and regulation of Na(+)-Ca(2+) exchange were examined for two alternatively spliced isoforms of the canine cardiac Na(+)-Ca(2+) exchanger, NCX1.1, to assess the role(s) of the mutually exclusive A and B exons. The exchangers examined, NCX1.3 and NCX1.4, are commonly referred to as the kidney and brain splice variants and differ only in the expression of the BD or AD exons, respectively. Outward Na(+)-Ca(2+) exchange activity was assessed in giant, excised membrane patches from Xenopus laevis oocytes expressing the cloned exchangers, and the characteristics of Na(+)(i)- (i.e., I(1)) and Ca(2+)(i)- (i.e., I(2)) dependent regulation of exchange currents were examined using a variety of experimental protocols. No remarkable differences were observed in the current-voltage relationships of NCX1.3 and NCX1.4, whereas these isoforms differed appreciably in terms of their I(1) and I(2) regulatory properties. Sodium-dependent inactivation of NCX1.3 was considerably more pronounced than that of NCX1.4 and resulted in nearly complete inhibition of steady state currents. This novel feature could be abolished by proteolysis with alpha chymotrypsin. It appears that expression of the B exon in NCX1.3 imparts a substantially more stable I(1) inactive state of the exchanger than does the A exon of NCX1.4. With respect to I(2) regulation, significant differences were also found between NCX1.3 and NCX1.4. While both exchangers were stimulated by low concentrations of regulatory Ca(2+)(i), NCX1.3 showed a prominent decrease at higher concentrations (>1 microM). This does not appear to be due solely to competition between Ca(2+)(i) and Na(+)(i) at the transport site, as the Ca(2+)(i) affinities of inward currents were nearly identical between the two exchangers. Furthermore, regulatory Ca(2+)(i) had only modest effects on Na(+)(i) dependent inactivation of NCX1.3, whereas I(1) inactivation of NCX1.4 could be completely eliminated by Ca(2+)(i). Our results establish an important role for the mutually exclusive A and B exons of NCX1 in modulating the characteristics of ionic regulation and provide insight into how alternative splicing tailors the regulatory properties of Na(+)-Ca(2+) exchange to fulfill tissue-specific requirements of Ca(2+) homeostasis. PMID- 10539977 TI - The future JMG: www.jmedgenet.com. PMID- 10539978 TI - Call for a moratorium on the use of the Rorschach Inkblot Test in clinical and forensic settings. AB - A call is issued for a moratorium on the use of the Rorschach Inkblot Test in clinical and forensic (but not research) settings. The moratorium should last until we have determined which Rorschach scores are valid and which ones are invalid. Unfortunately, for most Rorschach scores, results from meta-analyses have been uninformative. Also, incremental validity has not been studied for most Rorschach scores. Furthermore, positive findings for Rorschach scores have rarely been independently replicated. Finally, selective reporting of results has been a problem: Some investigators report significant results but not nonsignificant results. The magnitude of this problem has not been determined. Unless a moratorium is adopted, clinicians will continue to interpret invalid scores along with valid scores. PMID- 10539975 TI - High-level expression, functional reconstitution, and quaternary structure of a prokaryotic ClC-type chloride channel. AB - ClC-type anion-selective channels are widespread throughout eukaryotic organisms. BLAST homology searches reveal that many microbial genomes also contain members of the ClC family. An Escherichia coli-derived ClC Cl(-) channel homologue, "EriC," the product of the yadQ gene, was overexpressed in E. coli and purified in milligram quantities in a single-step procedure. Reconstitution of purified EriC into liposomes confers on these membranes permeability to anions with selectivity similar to that observed electrophysiologically in mammalian ClC channels. Cross-linking studies argue that EriC is a homodimer in both detergent micelles and reconstituted liposomes, a conclusion corroborated by gel filtration and analytical sedimentation experiments. PMID- 10539976 TI - Voltage sensitivity and gating charge in Shaker and Shab family potassium channels. AB - The members of the voltage-dependent potassium channel family subserve a variety of functions and are expected to have voltage sensors with different sensitivities. The Shaker channel of Drosophila, which underlies a transient potassium current, has a high voltage sensitivity that is conferred by a large gating charge movement, approximately 13 elementary charges. A Shaker subunit's primary voltage-sensing (S4) region has seven positively charged residues. The Shab channel and its homologue Kv2.1 both carry a delayed-rectifier current, and their subunits have only five positively charged residues in S4; they would be expected to have smaller gating-charge movements and voltage sensitivities. We have characterized the gating currents and single-channel behavior of Shab channels and have estimated the charge movement in Shaker, Shab, and their rat homologues Kv1.1 and Kv2.1 by measuring the voltage dependence of open probability at very negative voltages and comparing this with the charge-voltage relationships. We find that Shab has a relatively small gating charge, approximately 7.5 e(o). Surprisingly, the corresponding mammalian delayed rectifier Kv2.1, which has the same complement of charged residues in the S2, S3, and S4 segments, has a gating charge of 12.5 e(o), essentially equal to that of Shaker and Kv1.1. Evidence for very strong coupling between charge movement and channel opening is seen in two channel types, with the probability of voltage independent channel openings measured to be below 10(-9) in Shaker and below 4 x 10(-8) in Kv2.1. PMID- 10539979 TI - Behavioral science foundations of the Rorschach test: research and clinical applications. AB - Never without its critics, the Rorschach Test continues to be widely used in clinical settings. The test continues to be criticized vigorously. Rorschach critics appear to fall into two broad groups: those leveling valid methodological concerns about the test s behavioral science foundations and method critics who appear to deny the validity of the test on strictly a priori or theoretical considerations. Many critics do not appear to be acquainted with the extensive Rorschach research literature. The current paper provides an overview of several domains of applied and laboratory Rorschach behavioral science, including statistical power analysis, interobserver agreement and interrater reliability, Rorschach assessment of thought disorder, and emerging research linking Rorschach variables with diagnostic criteria from the DSM-IV, as a means of educating both adherents and detractors alike concerning the test s scientific track record and applicability to clinical assessment. PMID- 10539980 TI - What the Rorschach can do for you: incremental validity in clinical applications. AB - Psychological assessment instruments vary in how much structure they provide and the extent to which their meaning and purpose are apparent. The Rorschach Inkblot Method (RIM) is a relatively unstructured instrument whereas the MMPI-2 is a relatively structured instrument: People respond to these two instruments at different levels of conscious awareness concerning the possible significance of their responses. Because of its relatively unstructured nature, the RIM is less susceptible than the MMPI-2 to impression management. This complementarity makes it possible for Rorschach findings to enrich clinical assessments, especially when efforts to fake good result in MMPI-2 protocols that provide little reliable information. There is solid conceptual basis in psychology for employing multi method assessment, and clinical applications in which Rorschach data contribute to fuller or more accurate formulations than would otherwise be possible attest the incremental validity that can derive from including relatively unstructured measures in a test battery. PMID- 10539981 TI - The Rorschach Inkblot Test: a case of overstatement? AB - In the 1940s, inflated claims were often made regarding the Rorschach Inkblot Test. Over half a century later, overstatements regarding the test are still common. The present article identifies problems with the Rorschach regarding norms, cultural sensitivity, interrater reliability, test-retest reliability, validity, factor structure, and accessibility of supporting studies. Contrary to overstated claims made on behalf of the Rorschach, the test continues to be a highly problematic instrument from a psychometric standpoint. PMID- 10539983 TI - Performance of the Personality Inventory for Youth validity scales. AB - Response sets as well as cognitive and academic deficits compromise the validity of child and adolescent self-report of emotional adjustment. Three studies using clinical and asymptomatic samples of 4th to 12th grade students detail applications of the four validity scales of the Personality Inventory for Youth (PIY), namely, (a) Validity (VAL) a scale of six highly improbable statements, (b) Inconsistency (INC) consisting of pairs of highly correlated statements, (c) Dissimulation (FB) constructed of statements that were infrequent and characteristic of intentional distortion, and (d) Defensiveness (DEF) an extension of the Lie scale of the parent-report Personality Inventory for Children. The effects of minimizing, malingering, and random response sets on the PIY validity scales are reported. The importance of such validity scales derived from child and adolescent response is discussed. PMID- 10539982 TI - Heritability of MMPI Harris-Lingoes and Subtle-Obvious subscales in twins reared apart. AB - A large sample of identical and fraternal twins who had been reared apart was used to examine the genetic and environmental architecture of the MMPI Subtle Obvious and Harris-Lingoes subscales. Univariate genetic analyses indicated significant heritability for all 28 of the Harris-Lingoes subscales (estimates ranged from.23 to.61), all five Obvious subscales (estimates ranged from.37 to.56) and four of the five Subtle subscales (estimates ranged from.27 to.35). Two randomly constructed scales were analyzed as controls; neither of these scales showed significant heritability. Exploratory correlational findings suggested that three of the Wiener-Harmon Subtle subscales may tap aspects of psychological health, naivete, or repression. Ma-S may come closest to Wiener and Harmon s intent. Although they apparently diverge from their original purpose, it may be too early to abandon the low face valid items of the Subtle subscales. PMID- 10539984 TI - Internship training in psychological assessment: has managed care had an impact? AB - Training directors of 84 APA-approved internship programs in psychology responded to a survey on the current status and recent changes in internship training in psychological assessment. There appears to be a slight decline in emphasis on assessment in general, although it is still considered to be an important component of training. There was a surprising endorsement of traditional assessment instruments as important for clinical practice, in spite of a trend toward decreased emphasis on projective methods. The majority of programs have maintained the same degree of emphasis on objective measures, intelligence tests, behavioral methods, and the interview, while increasing emphasis on neuropsychological assessment. Less than half (43%) of the training directors reported that their programs have been significantly affected by managed care, suggesting that the majority of internship sites are somewhat insulated from present trends in marketplace pressures regarding reimbursement for psychological testing. The implications of potential future trends are discussed. PMID- 10539985 TI - Measuring perceived barriers to condom use: psychometric evaluation of the Condom Barriers Scale. AB - A programmatic series of three studies developed and evaluated the Condom Barriers Scale (CBS), an instrument measuring women s perceived barriers to condom use for prevention of HIV and other sexually transmitted diseases. Following item generation and selection, Study 1 evaluated the CBS in a sample of minority women (N = 178), reduced the number of items, assessed the factor structure, evaluated the internal consistency, and explored the convergent validity of the CBS. In Study 2, the CBS was administered to a cross-validation sample (N = 278). Confirmatory factor analysis and internal consistency were compared against the original sample and construct, criterion, and discriminant validity were assessed. In Study 3 (N = 30), temporal stability of the CBS was evaluated. The resulting instrument appears to have sound psychometric properties and can be used to measure a key construct in the leading theoretical models of health behavior for which a measure with known psychometric properties previously has not been available. PMID- 10539986 TI - Efficient computational algorithms for docking and for generating and matching a library of functional epitopes I. Rigid and flexible hinge-bending docking algorithms. AB - In this, and the next review article (1), we present highly efficient, computer vision and robotics based algorithms for docking and for the generation and matching of epitopes on molecular surfaces. We start with descriptions of molecular surfaces, and proceed to utilize these in both rigid-body and flexible matching routines. These algorithms originate in the computer vision and robotics disciplines. Frequently used approaches, both in searches for molecular similarity and for docking, i.e., molecular complementarity, strive to obtain highly accurate correspondence of respective molecular surfaces. However, owing to molecular surface variability in solution, to mutational events, and to the need to use modeled structures in addition to high resolution ones, utilization of epitopes might prove to be a judicious approach to follow. Furthermore, through the deployment of libraries of epitopes which represent recurring features, or motifs in a given family of receptors or of enzymes, in principle we a priori focus on the more critical groups of atoms, or amino acids, essential for the binding of the two molecules. Utilization of recurring motifs may prove more robust than single molecule matchings. In addition, via utilization of epitopes one can make use of information derived from evolutionary related molecules. All of the above combine to represent an approach which may be highly advantageous. Combinatorial approaches have proven their immense utility in the wet laboratory. The combination of efficient computational approaches and the utilization of such libraries may well be particularly profitable. Our highly efficient techniques are amenable to such a task. In this review we focus on rigid and flexible docking algorithms. In the second review (1) we address the generation of epitopes in families of molecules. These may be used by the docking algorithms to identify the more likely bound interfaces. PMID- 10539987 TI - Efficient computational algorithms for docking and for generating and matching a library of functional epitopes II. Computer vision-based techniques for the generation and utilization of functional epitopes. AB - This is the second review in a two-part series. In the first review (1) we described the computational complexity involved in the docking of a ligand onto a receptor surface. In particular, we focused on efficient algorithms designed to handle this computational task. Such a procedure results in a large number of potential, geometrically feasible solutions. The difficulty is to pinpoint which of these is the more likely candidate. While there exists a number of approaches to rank these solutions according to different criteria, such as the size of the interface or some approximation of their binding energetics, none of the existing methods has been shown to be consistently successful in this endeavor. If the binding site is unknown a priori, the magnitude of the task is awesome. Here we propose one way of addressing this problem, i.e., via derivation and utilization of binding epitopes. If a library of such epitopes is available, particularly for a large number of protein families, it may be used to predict more likely binding sites for a given ligand. We describe an efficient, computer-vision based method to construct binding epitopes focusing on two ways through which such a library can be generated, (i) molecular surface-based, or (ii) residue-based. Alternatively, the two can be combined. We further describe how such a library may be used efficiently in the matching/docking procedure. PMID- 10539988 TI - A high throughput platform for systematic evolution of ligands by exponential enrichment (SELEX). AB - The systematic evolution of ligands by exponential enrichment (SELEX) process is a combinatorial chemistry method for the isolation of nucleic acid ligands (aptamers) that bind to a desired target molecule with high affinity. In order to increase throughput via automation, we have adapted the SELEX process for protein targets to a robotics-compatible microtiter plate format. A remarkable feature of the platform is that targets are immobilized by hydrophobic adsorption onto the plate surface. Hydrophobic immobilization procedures are simple and require no specialized modification of the protein target. This format was tested by manually performing four independent SELEX experiments. All were concluded within 8 rounds of selection and yielded aptamers that bind in solution to their respective protein target, calf intestinal alkaline phosphatase, human alpha thrombin or human platelet derived growth factor, with equilibrium dissociation constants below 3 x 10-10 M. PMID- 10539989 TI - Application of homogeneous time-resolved fluorescence (HTRFTM) to monitor poly ubiquitination of wild-type p53. AB - Rapid degradation of wild-type p53 in the human uterine cervix is induced by the infection of high-risk human papilloma virus (HPV) types 16 and 18. HPV-E6 protein plays a critical role in the poly-ubiquitination of wild-type p53 by mediating the association of p53 with E6-associated protein (E6AP). As a result, the poly-ubiquitinated p53 is rapidly and selectively degraded by the 26S proteasome. We have established a high throughput assay system to monitor poly ubiquitination of wild-type p53 using a new fluorescence homogeneous technology known as Homogeneous Time-Resolved Fluorescence (HTRFTM). The Europium Cryptate [Eu(K)]-labeled ubiquitins are incorporated into poly-ubiquitin chains conjugated with the biotinylated p53. In the HTRF assay, Europium cryptate-labeled ubiquitin and streptavidin-labeled allophycocyanin (XL665) are used as the fluorescence donor and acceptor, respectively. The biotinylated p53 is ubiquitinated by ubiquitination enzymes, then by the addition of streptavidin-labeled XL665, the donor and acceptor molecules are brought in close proximity, thereby generating fluorescent signals. This time-resolved fluorescence assay system shows a sufficient signal for its application in synthetic compound screening and having almost the same level of sensitivity as that monitored by the scintillation proximity assay (SPA) using 125I-labeled ubiquitin. The detection of poly ubiquitination of wild-type p53 by using the HTRFTM or SPA systems described here is much easier and quicker than by using conventional methods. Therefore, these new systems would be appropriate for high throughput screening of compounds for the discovery of new inhibitors of poly-ubiquitination of wild-type p53. PMID- 10539990 TI - Identification of synthetic by-products in combinatorial libraries using high performance liquid chromatography-electrospray ionization mass spectrometry. AB - High performance liquid chromatography (HPLC), electrospray ionization mass spectrometry (ESI) and high performance liquid chromatography coupled to mass spectrometry (LC-MS) were used to analyze randomly chosen samples from parallel syntheses carried out on derivatized polypropylene crowns compatible with a Multipin solid support system. Side-reactions and by-products were clearly identified, and the yields of the expected molecules were unexpectedly low for most samples. LC-MS was superior to HPLC with absorbance detection or electrospray mass spectrometry alone for determining the identity and purity of each desired combinatorial compounds. PMID- 10539991 TI - beta-Lactamases of increasing clinical importance. AB - Resistance to b-lactam-containing antimicrobial agents continues to increase, frequently due to the presence of b-lactamases in Gram-negative bacteria. Over the past twenty-five years broad-spectrum enzymes such as TEM- and SHV-variants and the metallo-b-lactamases have become more prolific. As a result of the ability of plasmids to continue to acquire additional resistance determinants, many of the b-lactamase-producing Gram-negative pathogens have become multi-drug resistant. In combination with decreased permeability, the organisms can become virtually untreatable with current therapies. The major groups of b-lactamases that pose the most serious therapeutic problems include the extended-spectrum b lactamases, the plasmid-mediated cephalosporinases, the inhibitor-resistant TEM- or SHV-derived b-lactamases and the carbapenem-hydrolyzing b-lactamases. Those enzymes that can be transferred on mobile elements are the most serious of the newer b-lactamases, and include enzymes in each of the four groups outlined above. PMID- 10539992 TI - SHV-type beta-lactamases. AB - The group of plasmid-mediated SHV b-lactamases includes SHV-1 and at least twenty three variants, most of which possess extended-spectrum (ES) activity against the newer broad-spectrum cephalosporins. Their likely ancestor is a chromosomal penicillinase of Klebsiella pneumoniae. SHV enzymes belong to the molecular class A of serine b-lactamases and share extensive functional and structural similarity with TEM b-lactamases. The three-dimensional structure of the SHV-1 b-lactamase possesses an active site wider than that of TEM-1 b-lactamase by 0.7 to 1.2 A. This results in subtle, yet important, differences in the positioning of critical active-site residues. SHV-1 b-lactamase behaves as a typical penicillinase hydrolyzing penicillins and early generation cephalosporins. SHV-1 b-lactamase has spread, via plasmids, to virtually all enterobacterial species but is encountered mostly in K. pneumoniae. ES SHV b-lactamases are found with increasing frequency in K. pneumoniae and other enterobacterial isolates and are now considered the most prevalent ES b-lactamases. These ES SHV b-lactamases confer a wide spectrum of resistance to b-lactams, including the new generation cephalosporins and monobactams, and are usually encoded by self-transmissible multi-resistant plasmids that are highly mobile. Extension of the hydrolytic spectrum of ES SHV enzymes to include oximino-b-lactams is seen as a result of substitutions of critical amino acid residues that alter the properties of the active site. These mutational changes, however, result in diminished hydrolytic activity against penicillins and an increased susceptibility to mechanism-based inhibitors. Understanding the substrate evolution, properties and modes of spread of these clinically important b-lactamases can help in formulating effective antibiotic policies and developing new antimicrobial agents. PMID- 10539993 TI - OXA-type beta-lactamases. AB - The OXA-type (oxacillin-hydrolysing) enzymes are widespread and have been mostly described in Enterobacteriaceae and in P. aeruginosa. They usually confer resistance to amino- and ureidopenicillin and possess high-level hydrolytic activity against cloxacillin, oxacillin, and methicillin. Their activities are weakly inhibited by clavulanic acid but sodium chloride (NaCl) possesses a strong inhibition activity. Oxacillin-hydrolysing b-lactamases belong to Ambler class D and thus possess an active serine site as classes A and C b-lactamases. Overall amino-acid identities between class D and class A or class C b-lactamases is about 16%. Until now, 24 Ambler class D enzymes, named OXA-1 to OXA-22, AmpS and LCR-1, have been characterised, either by sequence and/or by biochemical analyses, but for none of them a three dimensional structure is yet available. While some oxacillinases present a significant degree of amino-acid identity (for example, OXA-1 and OXA-4; OXA-10 (PSE-2) derivatives; OXA-2 and OXA-3), most of them are only weakly related (20% to 30% amino-acid identity). Oxacillinases usually display a restricted-spectrum phenotype. However extension of their spectrum towards oxyimino cephalosporins and/or imipenem has recently been observed mostly as a consequence of point mutations in OXA-2 or OXA-10 derivatives. Their frequent plasmid- and/or integron-location provide them a mean for a wide diffusion. PMID- 10539994 TI - Regulation of inducible AmpC beta-lactamase expression among Enterobacteriaceae. AB - AmpC ss-lactamases are active-site serine enzymes that are primarily cephalosporinases. In many gram negative organisms, including Enterobacter spp.,Citrobacter freundii, Serratia marcescens, Morganella morganii and Pseudomonas aeruginosa, the expression of chromosomal ampC genes is low but inducible in response to ss-lactams and other stimuli. The current working model for AmpC induction requires exposure of bacterial cells to ss-lactam drugs or other stimuli and is linked to the cell wall recycling pathway. Induction of ampC appears to involve several gene products associated with this pathway. These gene products include AmpR, AmpD, and AmpG. In addition, anhydro forms of cell wall precursor muropeptides are believed to act as cofactors for AmpC induction. These cofactors bind to the DNA binding protein, AmpR, and define the role of AmpR as activator. Recent debate has ensued in the literature as to the identification of the precursor muropeptide involved in the activation process. Two candidate muropeptides include 1,6-anhydro-N-acetylmuramic acid L-Ala-D-Glu-meso diaminopimelic acid (anhydro-MurNAc-tripeptide) and anhydro-MurNAc-L-Ala-D-Glu meso-diaminopimelic acid- D-Ala-D-Ala (pentapeptide). The intent of this review is to address the general mechanism involved in AmpC induction. In doing so, the genes and gene products required for the process of AmpC induction are described. In addition, we review the data addressing cell wall recycling as it relates to AmpC induction. PMID- 10539995 TI - The reactivity of beta-lactams, the mechanism of catalysis and the inhibition of beta-lactamases. AB - Four membered b-lactam rings do not show unusual reactivity compared with their acyclic amide analogues and there is no evidence of concerted mechanisms for nucleophilic substitution reactions at the carbonyl centre. The identity of the general base/acid catalyst in the serine b-lactamases, which catalyse the hydrolysis of b-lactams, is unknown. There are no ideal transition state analogue inhibitors for these enzymes which involve several intermediates and transition states. The class C serine b-lactamase enhances the rate of phosphonylation of its active site serine residue by a similar magnitude to the enzyme rate enhancement factor for the hydrolysis of b-lactams. Comparisons are made between the stereochemical consequences of tetrahedral and trigonal bipyramidal intermediates for hydrolysis and phosphonylation respectively. Class B zinc b lactamases are inhibited by thiol dipeptides with a D configuration at the cysteine centre analogous to the L configuration at C6 in penicillins. The mechanism of hydrolysis catalysed by the metallo-b-lactamases probably involves a di-anionic tetrahedral intermediate stabilised by zinc(II). PMID- 10539997 TI - Structural and mechanistic aspects of evolution of beta-lactamases and penicillin binding proteins. AB - Penicillin-binding proteins (PBPs) and b-lactamases are related enzymes, the former are the targets for b-lactam antibiotics and the latter are resistance enzyme to these antibiotics. The two families of enzymes share structural topologies and certain mechanistic features. However, these classes of enzymes have diversified substantially and have broadened the reaction repertoire for their catalytic properties. This report addresses the issues of the evolution of function of these proteins. PMID- 10539996 TI - Class B beta-lactamases: the importance of being metallic. AB - The structural and functional features of class B b-lactamases, which are metal dependent, are reviewed in this article. Enzymes from different bacterial strains exhibit a common fold and sequence similarity in their active sites. However, the protein scaffold fine tunes the metal binding affinity and substrate selectivity. In this way, some metallo-b-lactamases seem to be functional with only one Zn(II) equivalent per enzyme, whereas others require a binuclear active site. The sequence similarity leads to a subdivision of these enzymes into three subclasses. The substrate specificities are rather broad, except for enzymes belonging to subclass B2. Some inhibitors have been designed and tested, but none of them is able to exhibit a broad spectrum against these enzymes. PMID- 10539998 TI - Recent advances in the chemistry of beta-lactam compounds as selected active-site serine beta-lactamase inhibitors. AB - The b-lactamases catalyze the hydrolysis of the b-lactam bond of a variety of b lactam antibiotics destroying their antibacterial activity. During the last decades, there has been an inexorable spread of b-lactamase genes into species that previously were not known to possess them. One approach to combat the action of the b-lactamase is to inhibit the enzyme. However, inhibition of b-lactamase alone is not sufficient. The ability to penetrate the external membrane of Gram negative bacteria, chemical stability, pharmacokinetics and other factors are also important in determining whether an inhibitor is suitable or not for therapeutic use. This review takes recent examples of synthetic b-lactam compounds developed as active-site serine b-lactamase inhibitors, emphasizing information on their structures and their activity against Ambler classes A, C and D b-lactamases. In addition, examples based on rational design by computerized molecular modeling of crystal structure of the native enzyme and mechanism of the enzyme action are highlighted. PMID- 10539999 TI - Solid-phase and combinatorial synthesis in beta-lactam chemistry. AB - Combinatorial chemistry has became a core technology for the rapid development of novel lead compounds in the pharmaceutical industry and for the optimization of therapeutic efficacy. The effort to prepare libraries of compounds by combinatorial chemistry has led to an unprecedented growth in solid phase organic synthesis (SPOS), particularly for the preparation of non-oligomeric small molecules. In this context, the clinically valuable b-lactam compounds are very attractive targets for research using these new techniques. In recent years, b lactam compounds have been recognized not only as unique antibacterial agents but also as potent enzyme inhibitors, drug delivery agents, and versatile synthetic intermediates. This review gives a comprehensive up-date for the application of solid-phase and combinatorial synthesis to b-lactam compounds. PMID- 10540000 TI - Preface PMID- 10540002 TI - The ALSFRS-R: a revised ALS functional rating scale that incorporates assessments of respiratory function. BDNF ALS Study Group (Phase III). AB - The ALS Functional Rating Scale (ALSFRS) is a validated rating instrument for monitoring the progression of disability in patients with amyotrophic lateral sclerosis (ALS). One weakness of the ALSFRS as originally designed was that it granted disproportionate weighting to limb and bulbar, as compared to respiratory, dysfunction. We have now validated a revised version of the ALSFRS, which incorporates additional assessments of dyspnea, orthopnea, and the need for ventilatory support. The Revised ALSFRS (ALSFRS-R) retains the properties of the original scale and shows strong internal consistency and construct validity. ALSFRS-R scores correlate significantly with quality of life as measured by the Sickness Impact Profile, indicating that the quality of function is a strong determinant of quality of life in ALS. PMID- 10540001 TI - Consensus guidelines for the design and implementation of clinical trials in ALS. World Federation of Neurology committee on Research. AB - BACKGROUND: In 1994 consensus guidelines were developed for conducting clinical trials in ALS. With growing experience in clinical trials, it has become clear that a number of further guidelines were needed. METHODS: Under the auspices of the World Federation of Neurology Committee on Research, a multinational group of neurologists, statisticians, patient advocates, representatives from the pharmaceutical industry as well as regulatory agencies developed consensus about a number of revisions to the existing guidelines during a 2 day conference in April 1998. RESULTS: Expanded areas of focus include greater protection of patient rights, more detailed guidelines for outcome measures statistical analyses, disclosure of study results and improved interaction between investigators and the corporate sector. COMMENT: Substantial progress has been made in standardizing and improving the quality of clinical trials in ALS through these consensus guidelines. PMID- 10540003 TI - The emotional lability questionnaire: a new measure of emotional lability in amyotrophic lateral sclerosis. AB - In an ALS Clinic, use of the Pathological Laughter and Crying Scale of Robinson et al. [Robinson RG, Parikh RM, Lipsey JR, Starkstein SE, Price TR. Pathological laughter and crying following stroke: validation of a measurement scale and double-blind treatment study. American Journal of Psychiatry 1993;150(2):286-293] revealed several problems: reliance on self-rating, insufficient period of assessment, inadequate exploration of 'appropriateness of emotion', lack of an item for abnormal smiling, amusement rather than happiness being the cause of laughter in ALS, and a frequency-based rating system. The necessary modifications produced a new Emotional Lability Questionnaire (ELQ) that was tested in 43 ALS patients and 43 healthy controls. The self-rated version of the ELQ was administered as a structured interview to each participant, and the independent relationship between subscales of the ELQ for both versions, thus confirming its internal validity. Greater emotional lability appeared associated with pseudobulbar symptoms. The question why 14 patients rated themselves as severely labile in the crying domain alone and four in the laughter domain alone, required further study. PMID- 10540004 TI - Quality of life assessment in MND: development of a social withdrawal scale. AB - A scale to measure 'social withdrawal,' was developed specifically for use with MND patients. Scale design was based upon patient-focused interviews which were used to explore patients' concerns. The impact the condition had upon their social interactions with others was salient for all patients and affected their overall quality of life. Using issues raised by patients, a 23-item scale was generated to specifically and quantitatively measure social withdrawal and this scale was psychometrically evaluated. The scale was administered to a sample of 23 patients at varied stages of progression of MND, and to a control group of patients with arthritis with similar levels of physical disability. For MND patients withdrawal from the community was found to be closely related to severity of physical symptoms and depression. Specific patients were identified who are particularly susceptible to withdrawal and depression. Comparison of the pattern of withdrawal in the MND and arthritis patient groups suggested that there may be differences between the factors governing social withdrawal in different patient populations. PMID- 10540005 TI - Acoustic analysis of dysarthria profile in ALS patients. AB - Dysarthria is a leading disability in ALS patients with motor neurone degeneration in the bulbar region. Although different approaches have been tried in the past, currently, no test is available to detect and follow the progression of dysarthria. We studied 53 patients with definite (n=27) or probable (n=26) ALS (the bulbar onset group n=15, the limb onset group n=38, mean age 53. 66/29-76 years/) according to El Escorial criteria. Each patient was seen by a neurologist every 10-12 weeks and clinical performance was assessed using the Norris scale. To evaluate dysarthria we developed a computer-based acoustic method. All patients had computer-analysed speech sound tests done three times. The most significantly affected vowels were selected for further studies. A method based on the Euclidian principle was used and the results were compared with 30 age, sex-matched, healthy control subjects. Our results demonstrated the existence of a specific dysarthria profile in ALS patients with most significantly affected vowels: 'B', 'O', 'I', 'W', 'T' in the bulbar group, and: 'B', 'I', 'T', 'W', 'O' in the limb group. This study suggests that it is possible to detect and monitor the progression of the disease based on the acoustic analysis of only several sounds. Abnormalities detected in the dysarthria profile may appear prior to any clinical symptoms of the disease. PMID- 10540006 TI - Pathological laughing and crying in amyotrophic lateral sclerosis: an association with prefrontal cognitive dysfunction. AB - Pathological laughing and crying (PLC) frequently occurs in amyotrophic lateral sclerosis (ALS). The etiology of the syndrome is unclear, but frontal-subcortical circuits are implicated, given their known association with mood and affect regulation. Ten ALS patients with PLC, eight patients without, and ten healthy controls were compared on a number of psychometric measures. Three indices of prefrontal cortical function were given: the Wisconsin Card Sort Test (WCST), the novel 'Gambling task' and a measure of olfactory discrimination. Global cognitive ability, psychiatric symptoms, and illness variables were also examined. No significant between-groups differences emerged with respect to global cognitive ability, mood, olfaction, and performance on the Gambling task. On the WCST, however, patients with PLC made significantly more total errors than the other two groups, and showed a strong trend in a similar direction for perseverative errors. A discriminant function analysis revealed that the WCST variable 'total errors' correctly predicted the presence or absence of pathological affect in 75% of cases. Thus, PLC appears to be associated with impairment in the functional integrity of the prefrontal cortex. Although this was not found for all prefrontal measures, further investigation of this area appears warranted. PMID- 10540008 TI - Clinical characteristics of SOD1 gene mutations in UK families with ALS. AB - Five to ten percent of patients with ALS have a family history of the disease, inheritance is usually autosomal dominant. Mutations of the SOD1 gene were first identified in a proportion of families with ALS by Rosen et al. The SOD1 gene encodes the enzyme copper zinc superoxide dismutase. Patients were studied from throughout the UK, where more than one individual in the family had ALS. Clinical history and examination of the individual and family were obtained, and DNA extracted from leukocytes of whole blood samples. Mutations were identified by standard sequencing methods. To date, 12 different mutations of SOD1 have been identified in 17 different families, representing around 20% of all ALS families studied. The mutations were mainly single base substitutions - H48Q, G72S, G93R, G93V, E100G, D101N, D101G, G108V, I113T, D125H, I149T - and also an insertion mutation - 132insTT - leading to a premature stop codon. The mutations were present in exons 2-5. We did not identify mutations in exon 1, although these have been identified by others in different patient samples. We have identified SOD1 mutations in around 20% of UK families with ALS studied. This is similar to that reported in other populations. Mutations have now been identified in all exons of SOD1. The individual mutations do not precisely predict disease severity, and generally it is difficult to give a specific prognosis based on the individuals' SOD1 mutations. We continue to investigate the possible pathogenic mechanisms of the SOD1 mutations. We have studied the neuropathology in patients with SOD1 mutations. We are also performing linkage studies to identify the genes involved in the 80% of families where an SOD1 mutation has not been identified. PMID- 10540007 TI - Motor system abnormalities in heterozygous relatives of a D90A homozygous CuZn SOD ALS patient of finnish extraction. AB - Presently, 64 mutations in the gene encoding the enzyme CuZn-superoxide dismutase have been found in a small fraction of amyotrophic lateral sclerosis patients worldwide. All but one of these mutations show autosomal dominant inheritance. In Scandinavia, the D90A mutation is inherited as an autosomal recessive trait and patients have an easily recognizable characteristic phenotype with little variation among patients, even amongst different families. Importantly, all D90A heterozygous relatives of Scandinavian D90A homozygous patients have been reported as clinically unaffected. We have investigated a Canadian family of Finnish extraction in which the D90A homozygous proband developed ALS with the characteristic phenotype. Remarkably, two D90A heterozygous relatives show slight symptoms and signs of motor system involvement, suggesting that the final phenotype of an individual with a CuZn-superoxide dismutase mutation is shaped by the combination of the particular CuZn-SOD mutation, other polymorphic modifying genes elsewhere in the genome, stochastics and possible environmental factors. PMID- 10540009 TI - Respiratory disorders in ALS: sleep and exercise studies. AB - Sleep disruption in ALS/MND is related to hypoventilation and nocturnal O(2) saturation. Maximal inspiratory pressure (PI(max)) proved sensitive in predicting nocturnal O(2) saturation. However, PI(max) is highly dependent on patient collaboration; on the other hand Mouth Occlusion Pressure (MOP) is a reliable, non-volitional parameter index of central respiratory drive. Since exercise testing (ET) is also part of the assessment of ventilatory regulation the authors aimed to determine whether MOP and ET are sensitive and reliable parameters predictive of nocturnal O(2) saturation and clinical evolution. We conducted a Polysomnographic (PSG) study in two groups of 14 patients, selected according to their MOP level. Patients performed at admission an ET, Respiratory Function tests (RFT) and clinical evaluation with Norris spinal and bulbar scores (SNS and BNS). All patients in Group I (Low MOP) had decreased O(2) saturation during ET (P<0.001). Correlation study showed correlation between ET and MOP (R=0.6); PI(max) slope and PE(max) slope correlated with ET (R=-0.4; -0.6), respectively. ET also correlated with nocturnal O(2) saturation and SNS slope (R=0.8; -0.5), respectively. SNS and BNS slopes correlated with nocturnal O(2) saturation (R= 0.4; -0.7), respectively. The best correlations found were between MOP slope and BNS slope and SNS slope (R=0.8; 0.7), respectively. The high predictive values of MOP and ET at admission to nocturnal O(2) saturation (predicted value=80%) suggested the need of nocturnal pulse oximetry as a standard procedure. MOP and ET should also be used in evaluation protocols of ALS/MND. PMID- 10540010 TI - Can amyotrophic lateral sclerosis patients with respiratory insufficiency exercise? AB - The authors have shown in a recent paper that survival with amyotrophic lateral sclerosis (ALS) can be increased by the use of non-invasive methods of assisted ventilation (Bipap). However, the progression of muscle weakness was not affected and the quality of life was not positively enhanced. In ALS, reduced physical activity may partially be secondary to alveolar hypoventilation syndrome. This leads to deconditioning of ALS/motor neuron disease (ALS/MND) patients. The authors decided to investigate the possibility of reducing motor decline by exercising these patients to the anaerobic threshold, but simultaneously compensating the respiratory insufficiency with the Bipap STD. We conducted a controlled single blind study, exercising eight consecutive ALS/MND patients and used a control group of 12 ALS/MND patients. The patients were all evaluated during a 1 year period. Respiratory function tests (RFT) were performed at entry and then at 6 month intervals. Barthel, Functional Independent Mobility scale (FIM) and Spinal and Bulbar Norris scores were recorded every 3 months. There was a significant difference between the two groups with respect to FIM scores (P<0.03), but not Barthel scores (P<0.8). A slower clinical course (Spinal Norris score P<0.02) and a significant difference in the slope of the RFT (P<0.008) were observed in the treated group, suggesting that exercise may be beneficial in ALS patients once Bipap is used to control peripheral and muscle oxygenation. PMID- 10540011 TI - Multifocal motor neuropathy mimicking motor neuron disease: nine cases. AB - Multifocal motor neuropathy with persistent conduction block (MMN) is a rare clinical entity, mimicking motor neuron disease (MND). In order to research which are the most frequent nerves and segments where conduction block (CB) can be identified, we reviewed the clinical and neurophysiological data of nine patients with MMN who were studied and followed by the authors. Weakness and muscle atrophy of the dominant hand was the most frequent presentation. Lower limbs were involved later in the disease evolution. The ulnar and median nerves were the most affected nerves. They had conduction blocks mostly at the forearm and at Erb's point-elbow (or above elbow) segments. Both common peroneal and tibial nerves were frequently affected at their distal segments, but proximal segments were also probably involved. The presence of anti-GM1 antibodies was variable, and their determination was not essential for the diagnosis of MMN. Eight patients given IV immunoglobulin therapy had no disease progression. One patient was responsive to corticosteroids. The CB identification in our patients allowed us to clearly distinguish MMN from MND. The good prognosis and need for management with IV immunoglobulin, support the crucial role of a careful neurophysiological study to diagnose this clinical entity. PMID- 10540012 TI - Reflex sympathetic dystrophy associated with amyotrophic lateral sclerosis. AB - Reflex sympathetic dystrophy (RSD) is a syndrome characterised by severe distal pain and vasomotor changes. It is believed to be caused by sympathetic nervous system overactivity. Trauma is the most frequent precipitant event. An association with amyotrophic lateral sclerosis (ALS) has been reported only once. We report three patients with ALS in whom the occurrence of RSD, in one of them at a very early clinical stage, seemed to have precipitated a more rapid clinical evolution. New sprouting re-innervating fibres have abnormal ion channels which might increase the risk of RSD. On the other hand, motor changes have been described in RSD, as well as motor strength improvement after RSD treatment. The complex relation of ALS with RSD is discussed. In all ALS patients pain followed by further loss of function should prompt a search for RSD. PMID- 10540013 TI - 1H-magnetic resonance spectroscopy in amyotrophic lateral sclerosis. AB - 1H-magnetic resonance spectroscopy (MRS) is potentially a powerful tool for the investigation of the chemicals of the brain in vivo in health and disease. Levels of N-acetyl-aspartate (NAA) in the motor cortex and brainstem of patients with amyotrophic lateral sclerosis (ALS) have been reported to be reduced by up to 68%, and in one report the level of glutamate in the brainstem was increased by 58%. We studied levels of metabolites in the cerebral cortex and brainstem of 20 ALS patients and 14 age-matched controls with a 1.5 Tesla Picker magnet using MRS. We used the same spectra for determining both the area of the metabolite peaks expressed as a ratio of the area of the creatine (Cr) peak, and the absolute concentrations using the Provencher LC model. These produced different results. With the LC model, the NAA content of the motor cortex of ALS patients was reduced by 7.7% (P=0.015), and that of the brainstem was reduced by 21.5% (P=0.035), compared with controls. The degree of reduction of NAA was related to the severity of upper motor neuron abnormalities. No effect of treatment with anti-glutamate agents on NAA concentration could be detected. Concentrations of other metabolites were not affected in ALS. It appears that MRS is a technique that is still in development, and that further refinement is required before it can be used to understand disease mechanisms and investigate treatment in ALS. PMID- 10540014 TI - A receptor for advanced glycosylation endproducts (AGEs) is colocalized with neurofilament-bound AGEs and SOD1 in motoneurons of ALS: immunohistochemical study. AB - Neurofilament (NF)-bound AGEs colocalize immunochemically with SOD1 in the motoneurons of patients with ALS. Among three types of AGE receptors reported in the human brain, AGE-R1 (oligosaccharyltransferase family) and AGE-R2 (substrate of protein kinase C) have been found in neurons, while AGE-R3 is restricted to glia. The present study investigates which of these receptors may be responsible for binding AGEs in the NF conglomerates of motoneurons. Immunostaining of paraffin sections from eight ALS patients (five sporadic and three familial) and three control cases was performed with antibodies directed against R1 and R2, in parallel with those against AGEs and SOD1. The sites of AGE-R1 immunoreactivity (IR) in motoneurons were in conformity to those of NF-associated AGE and SOD1 IRs. By contrast, the IR of R2 was negative in NF conglomerates. Negative R2 IR for NF conglomerates was outlined by surrounding coarse R2 immunopositive granules in the perikaryon. No IR for R1 or R2 was found in hyaline or Bunina inclusions. There was no extraneuronal expression of IR for AGE-R1 or AGEs in microglia or astroglia around the NF accumulation. The colocalization of AGE, AGE R1, and SOD1 at NF conglomerates in motoneurons supports the notion that AGE mediated oxidative stress and protein aggregation may be implicated in NF conglomeration and ALS pathogenesis. PMID- 10540015 TI - Cortical silent period in patients with amyotrophic lateral sclerosis. AB - Transcranial magnetic stimulation (TMS) during a muscle contraction induces a motor-evoked potential (MEP) in the skeletal muscle followed by a cessation of EMG activity, the cortical silent period (C-SP). The C-SP is a useful parameter to indicate the activation of the motor system. Accurate determination of the C SP can be important in amyotrophic lateral sclerosis (ALS), a progressive disorder of unknown etiology characterised by degeneration of upper and lower motor neurons. The aim of this study was to evaluate the sensitivity of C-SP as an index of motor system involvement, in ten patients affected by ALS, with a mean duration of the disease: 5. 5+/-3.4 months, by means of an objective computer-aided method to measure C-SP and its relationship to stimulation intensity. C-SP duration was significantly reduced in ALS patients compared to controls at low stimulation intensity corresponding to an MEP threshold increased by 15%. While in less severely affected patients C-SP duration approached control values at higher stimulation intensities (25 and 50% upper MEP threshold), in more severe ALS subjects it showed a further reduction, allowing them to be discriminated. PMID- 10540016 TI - The effect of riluzole in amyotrophic lateral sclerosis: a study with cortical stimulation. AB - A population of 31 patients with sporadic amyotrophic lateral sclerosis (ALS) was selected for a prospective open study based on treatment with riluzole. A neurophysiological evaluation was performed by means of single and paired transcranial magnetic stimulation (TMS). The examined parameters, excitability threshold, motor evoked potential (MEP) duration, silent period (SP) duration and time course of intracortical inhibition to paired TMS after 6 months treatment, were matched against those recorded from the patients themselves before the beginning of treatment and from 20 (single TMS) or 10 (paired TMS) age-matched control subjects. Normal behaviour of the SP in response to increasing TMS was found in the treated patients; they showed a significant linear correlation between these two parameters (r=0.96) comparable to that calculated for controls (r=0.98), and significantly different with respect to drug-free patients (r=0.8, P=0.014). A significant reduced size of the 'conditioned' MEPs to paired stimulation was documented in the treated patients compared with the untreated patients (P=0.002). Our neurophysiological contribution to the assessment of the effect of riluzole on the motor cortical inhibitory property in ALS may be considered a setting for controlled trials in extended patient series, even in a pre-clinical phase. PMID- 10540017 TI - Prospective study of palliative care in ALS: choice, timing, outcomes. AB - To understand palliative care choices among people with ALS, it is important to follow a group of recently diagnosed patients and record when patients reach well defined palliative care milestones. We began following such a cohort prospectively in 1996, when 121 ALS patients were enrolled from a tertiary care clinic. These patients are assessed every 4 months to determine their experience with ameliorative and palliative care. Domains include adjuvant therapies (e.g. speech therapy), adaptive aids (e.g. wheelchair use, augmentative communication), home health care, PEG placement, pulmonary support, health care directives, psychosocial care (e.g. participation in support group, pastoral counseling), and hospitalization. The median follow-up time for the cohort, to date, was 12 months. In the group that entered the cohort within 1 year of their diagnosis (n=93), 53.8% have died, 22.6% have had PEG, 19.4% have used Bi-PaP, and 4.3% have had a tracheostomy. Many patients did not take advantage of palliative care options before death; for example, 36.6% used hospice, 48% had signed a power of attorney form, and 18% had 'do not resuscitate' orders in their medical charts. Examining time to such endpoints captures important features of patient and family experience with the disease. PMID- 10540018 TI - Pulmonary evaluation and prevalence of non-invasive ventilation in patients with amyotrophic lateral sclerosis: a multicenter survey and proposal of a pulmonary protocol. AB - In order to evaluate the current standard of care for the management of respiratory failure in patients with amyotrophic lateral sclerosis (ALS), a questionaire was mailed to the Medical Directors of 48 multidisciplinary ALS centers in the United States. Twenty centers reported information on 2357 patients, mean of 124 patients per center. Pulmonary function tests were performed at each visit in 17/20 institutions. Arterial blood gases, maximal expiratory pressures and maximal inspiratory pressures were followed in three centers and serum chloride was monitored in only four centers. The use of non invasive ventilation (NIV) was extremely variable (range 0-50%) and included 360 patients (15%). The majority of centers used symptoms/signs of hypoventilation and worsening forced vital capacity (FVC) to initiate NIV with no established protocol. A FVC between 20 and 40% was used by most centers to initiate NIV. Due to great variability in the approach to monitoring pulmonary function among ALS centers and the modest effects of current medications to slow disease progression, we propose the use of a structured protocol which can prospectively study the role of NIV in prolonging survival and improving quality of life. PMID- 10540020 TI - Estrogen replacement therapy in women with amyotrophic lateral sclerosis. AB - Amyotrophic Lateral Sclerosis (ALS) occurs more commonly in men than in women, and women get the disease later in life compared to men. This epidemiologic aspect of the disease raises the question as to whether estrogen may be neuroprotective in delaying or preventing ALS. Postmenopausal women with ALS were separated into two groups dependent upon whether or not they took estrogen replacement therapy. Women who used estrogen had onset of their disease at an earlier age compared to those not taking hormonal replacement. There was no difference in survival in those patients taking estrogen compared to those not on the medication. Women with ALS were more likely to take estrogen compared to a control group of patients with neurological diseases other than motor neuron disease. Therefore, no evidence for a neuroprotective role of estrogen in postmenopausal women with ALS was found. PMID- 10540019 TI - A retrospective study of percutaneous endoscopic gastrostomy in ALS patients during the BDNF and CNTF trials. AB - Percutaneous endoscopic gastrostomy (PEG) provides a reliable route for nutrition and hydration in ALS patients with dysphagia. We performed a retrospective analysis of the CNTF and BDNF databases to determine the clinical status of ALS patients within 30 days preceding PEG insertion. This analysis revealed an approximately 50% reduction of function across multiple measures of ALS disease status. A trend to earlier intervention with PEG was apparent upon review of published studies and the CNTF and BDNF studies. By comparing the rate of decline pre- and post-PEG, nutritional supplementation via PEG stabilized the weight loss experienced by patients. Death within 30 days post-PEG was associated with a marked reduction in forced vital capacity (FVC) and identified a group of ALS patients in whom PEG should be cautiously performed. These data emphasize the importance of sequential measurement of FVC in managing ALS patients to guide the timing of nutritional intervention with PEG. PMID- 10540021 TI - Percutaneous gastrojejunostomy in amyotrophic lateral sclerosis. AB - We have performed a retrospective review of the use of a percutaneous gastrojejunostomy in patients with amyotrophic lateral sclerosis (ALS). Forty-one patients with initial bulbar manifestations of ALS and 32 patients with initial limb manifestations underwent a percutaneous gastrojejunostomy under fluoroscopic control using the Rankin gastrojejunostomy tube. Survival characteristics were compared with 86 bulbar onsetting and 207 limb onsetting ALS patients who did not require nutritional support. The 30-day mortality rate was 9.6% (respiratory death in three bulbar onsetting patients and four limb onsetting patients) and the 30 day morbidity rate was 4.1% (one operative site infection and intraperitoneal leakage in two patients). The most frequent long-term complication was the requirement for tube changing (blockage in six; dislodgment in two). Gastric reflux was not described amongst the treated patients. Overall survivorship (symptom onset to death) was less in the bulbar onsetting patients receiving a gastrojejunostomy tube than in the control population (median survival 22.0 vs. 33.7 months, respectively, P=0.005). As a group, the median survivorship for limb onsetting patients was not different for those receiving a gastrojejunostomy than for those who did not. However, a significant reduction in survival was observed in limb onsetting patients receiving a gastrojejunostomy early in the course of their disease (P=0.001) compared to those with a longer duration prior to the procedure. This was not observed in the bulbar onsetting patients. In both patient populations, no relationship was observed between survival post-gastrojejunostomy and the severity of pulmonary involvement at the time of the intervention, serum chloride, or age at onset. These studies demonstrate that a percutaneous gastrojejunostomy is a well-tolerated and safe alternative technique for enteral nutritional support in ALS patients. It also offers the advantage of not requiring either a general anaesthetic at the time of the procedure or instrumentation through the oropharynx. We have also observed that limb onsetting patients requiring a gastrojejunostomy early in the course of their illness are in a distinctive, less favorable, prognostic group. PMID- 10540022 TI - Visualization of defective mitochondrial function in skeletal muscle fibers of patients with sporadic amyotrophic lateral sclerosis. AB - The mitochondrial function in skeletal muscle was investigated in skeletal muscle biopsies of 26 patients with sporadic amyotrophic lateral sclerosis (ALS) and compared with investigations of 28 age-matched control muscle samples and biopsies of 6 patients with spinal muscular atrophy (SMA) and two patients with Tay-Sachs disease. In comparison to the control, SMA and Tay-Sachs biopsies, we observed in the ALS samples a significant about two-fold lower activity of complex I of mitochondrial respiratory chain. To visualise the distribution of the mitochondrial defect in skeletal muscle fibers we applied confocal laser scanning microscopy and video fluorescence microscopy of NAD(P)H and fluorescent flavoproteins. The redox change of mitochondrial NAD(P)H and flavoproteins on addition of mitochondrial substrates, ADP, or cyanide were determined by measurement of fluorescence intensities with dual-photon UV-excitation and single photon blue excitation. In skeletal muscle fibers of ALS patients with abnormalities of mitochondrial DNA (multiple deletions, n=1, or lower mtDNA levels, n=14) we observed a heterogeneous distribution of the mitochondrial defects among individual fibers and even within single fibers. In some patients (n=3) a mitochondrial defect was also detectable in cultivated skin fibroblasts. These findings support the viewpoint that the observed impairment of mitochondrial function in muscle of certain ALS patients is caused by an intrinsic mitochondrial defect which may be of pathophysiological significance in the etiology of this neurodegenerative disease. PMID- 10540024 TI - Prevention by insulin-like growth factor-I and riluzole in motor neuron death after neonatal axotomy. AB - Transection of the sciatic nerve in neonatal rats results discernable loss of motor neurons in the spinal cord. This neuronal death could be due to lack of retrogradely transported target derived neurotrophic factors, since some of these factors have been shown to be effective in injury induced motor neuron death. Another hypothesis suggests that glutamate and its receptors has been implicated as possible mechanism for motor neuron death, because inhibitor of glutamate release and antagonists of glutamate receptors are effective in preventing axotomized motor neuron death. To investigate the effect of insulin-like growth factor-I (IGF-I) and riluzole, a drug that inhibits glutamate release, on axotomy induced motor neuron death. Newborn rats were anesthetized with hypothermia. Sciatic nerve was cut near the obturator tendon in the left thigh. Animals were then treated daily with different doses of IGF-I and riluzole for 14 days with intraperitoneal injections. Control rats received PBS in the same fashion. After the treatment, the number of surviving motor neurons and the motor neuron diameter in the L(4) was assessed. Both IGF-I (1.0 mg/kg) and riluzole (5.0 mg/kg) rescued motor neuron death in a similar way. Co-administration of IGF-I (1.0 mg/kg) and riluzole (5.0 mg/kg) was more effective than either agent alone and there was a statistically significant difference between co-administration and IGF-I alone. However there was no significant difference between simultaneous treatment and riluzole alone. As for diameter of motor neurons, riluzole (5.0 mg/kg) preserved the motor neuron diameter in the lesion side. Nonetheless, no further increase in motor neuron diameter was seen when riluzole (5 mg/kg) and IGF-I (1.0 mg/kg) were applied in combination. Both agents did not affect diameter of motor neurons in the non-axotomy side. Riluzole is available in amyotrophic lateral sclerosis (ALS) and the positive results of clinical trials with IGF-I suggests that combination treatment of IGF-I and riluzole in ALS remains to be determined. PMID- 10540023 TI - Comparative analysis of motoneuron loss and functional deficits in PMN mice: implications for human motoneuron disease. AB - We have investigated the correlation between functional and morphological deficits in PMN mice, an animal model of human motoneuron disease. Electrophysiologic investigations showed first abnormalities, i.e. reduction of M response amplitudes, already at postnatal d 13 when the disease was not yet phenotypically apparent, and when motoneuron and motor axon numbers were still normal. After d 27, a loss of more than 30% of motoneuron axons and cell bodies was detectable in the phrenic nerve and facial nucleus, respectively. At that stage, PMN mice showed severe functional and electrophysiological deficits. At later stages of the disease when still more than 50% of motor axons and at least 60% of motoneuron cell bodies were present, the distal compound muscle action potential amplitude decreased by more than 95% in small foot muscles after sciatic nerve stimulation. We conclude that functional deficits precede structural deficits in this animal model of human motoneuron disease. Our findings are in agreement with the concept of the 'sick motoneuron' in this animal model of motoneuron disease rather than the idea of progressive loss of motoneurons resulting in disease only after a significant number of motoneurons has degenerated. PMID- 10540025 TI - Effect of sympathectomy on the expression of NMDA receptors in the spinal cord. AB - The expression of NMDA receptors in the intermediolateral (IML) region of the upper thoracic spinal cord, was studied in 3 week old rats. The effect of section of the cervical sympathetic nerve on neuronal cell number and receptor expression was examined up to two weeks after the operation. Age-matched sham-operated and unoperated animals were used as controls. It was shown using quantitative autoradiography with the NMDA receptor antagonist [(3)H]MK-801 (dizocilpine maleate), that there was a marked downregulation of receptors in all groups of animals, beginning at approximately 4 weeks of age. However after sympathectomy, which resulted in the death of 44% of neurones in the IML by 7 days, there was a significant increase in receptor density per neurone compared to sham-operated controls. In the control animals there was a significant increase in the Kd value of the binding between 21 and 24 days after birth indicating an increased expression of a low affinity receptor, but no such increase was seen after axotomy. The results are consistent with two populations of NMDA receptors being transiently expressed in the IML in developing animals, and the higher affinity receptor being down-regulated between 4 and 5 weeks of age. The presence of the high affinity receptor subtype may predispose neurones to die after axotomy. PMID- 10540026 TI - Motor neurone metabolism. AB - The cell and molecular mechanisms which determine the motor neurone (MN) phenotype are unclear. Tissue culture models offer a unique system for the study of a wide variety of MN features. For instance, since the neurone-astrocyte metabolic interactions play a critical role in the selective MN loss observed in amyotrophic lateral sclerosis (ALS), the glutamatergic MN toxicity could be reanalyzed in vitro, after a careful evaluation of the role of astrocytes. Ca(2+) appears to be important in inducing MN loss from in vitro studies. It was shown primarily in culture that apoptotic or necrotic death of neurones after injury depends upon the cell energetic status. Also, SOD-1 mutations were successfully expressed in cultured MNs, providing a critical assay to sequence the molecular processes responsible for MN degeneration due to an identified genetic defect. Purified human developing MNs and astrocytes were recently obtained from the spinal cord anterior horn. The effects of molecules affecting MN survival, neurite extension, and metabolism can easily be tested in long-term cultures. Interactions at the single cell level can be studied today using a series of RNA amplification techniques. Understanding the properties of human MNs in vitro may represent a critical tool in defining regional metabolic changes that could constitute the first pathogenic event of cell degeneration in ALS. PMID- 10540027 TI - Neurofilament metabolism in sporadic amyotrophic lateral sclerosis. AB - Although the role of intraneuronal neurofilamentous aggregates in the pathogenesis of ALS is unknown, their presence forms a key neuropathological hallmark of the disease process. Conversely, the experimental induction of neurofilamentous aggregates in either neurotoxic or transgenic mice gives rise to motor system degeneration. To determine whether alterations in the physiochemical properties of NF are present in sporadic ALS, we purified NF subunit proteins from cervical spinal cord of ALS and age-matched control patients. The cytoskeleton-enriched, Triton X-100 insoluble fraction was further separated into individual NF subunits using hydroxyapatite HPLC. We observed no differences between control and ALS in the characteristics of NFH, including migration patterns on 2D-IEF, sensitivity to E. coli, alkaline phosphatase mediated dephosphorylation, peptide mapping, or proteolysis (calpain, calpain/calmodulin mediated, phosphorylated or dephosphorylated NFH). NFL showed no differences in 2D-IEF migration patterns, peptide mapping, or the extent of NFL nitrotyrosine immunoreactivity in either the Triton soluble or insoluble fractions. The latter observation demonstrated that NFL nitration is a ubiquitous occurrence in neurons and suggests that NFL might function as a sink for free reactive nitrating species. In contrast to the lack of differences in the post-translational processing of NF in ALS, we did observe a selective suppression of NFL steady state mRNA levels in the limb innervating lateral motor neuron column of ALS. This occurred in the absence of modifications in NFH, NFM or neuronal nitric oxide synthase (Type I NOS; nNOS) steady state mRNA levels. Coupled with previous observations of nNOS immunoreactivity co-localizing with NF aggregates in ALS motor neurons, this suggests activation of the nNOS enzyme complex in ALS, which would be predicted to contribute directly to the generation of reactive nitrating species. Given this, the isolated suppression of NFL steady state mRNA levels in ALS may indicate that ALS motor neurons are at an intrinsic deficit in the ability to buffer free reactive nitrating species. PMID- 10540028 TI - Primary lateral sclerosis: disease, syndrome, both or neither? PMID- 10540030 TI - Axonal degeneration in the pathogenesis of multiple sclerosis. AB - Axonal degeneration plays an important role in the accumulation of disability in patients with multiple sclerosis (MS). Pathological studies have demonstrated axonal damage, particularly in areas of acute inflammation and demyelination, and in chronic lesions. Axonal loss and its progression, which is associated with neurological disability, has also been demonstrated by magnetic resonance imaging (MRI) studies. The mechanisms of axonal loss are uncertain, but may involve axonal degeneration secondary to demyelination, or damage to the axonal cytoskeleton. Inflammatory mediators, including cytokines and proteolytic enzymes may contribute to axonal damage, as may nitric oxide. Axonal destruction may also be due to immune attack directed at axonal components. The realisation that axonal degeneration is a fundamental component of MS that may occur early in the disease course should alter the approach to management and open avenues to a more targeted immunotherapy aimed at reducing the progression of disability. PMID- 10540029 TI - What is primary lateral sclerosis? AB - Primary lateral sclerosis (PLS) is a rare degenerative disorder of the upper motor neuron. Its nosological status and relationship to other motor neuron syndromes, especially amyotrophic lateral sclerosis (ALS), is uncertain. Diagnostic criteria have been proposed. We discuss the history of this rare clinical disorder, its relationship to the motor neuron disease syndrome, and reports of overlapping clinico-pathological conditions. Two patients with the clinical syndrome of PLS are described to illustrate current understanding of the clinical, laboratory, and neurophysiological features. PMID- 10540031 TI - Bell's palsy and herpes viruses: to (acyclo)vir or not to (acyclo)vir? AB - The majority of peripheral seventh cranial nerve palsy cases remain without an identified etiology and will eventually be diagnosed as idiopathic or Bell's palsy. Some features of this condition may be characteristic of a viral infection. Indeed, several herpes viruses have been implicated as potential causative pathogens. Besides varicella-zoster virus, shown to cause Bell's palsy under the Ramsay-Hunt syndrome, recent years have seen an increased interest and focus on the possible herpes simplex virus type 1 (HSV-1) etiology in idiopathic facial paralysis. We review the clinical, biological and virological basis for the potential herpetic cause of Bell's palsy and the rational for antiviral therapy in this condition. PMID- 10540032 TI - Cellular and molecular studies in muscle and cultures from patients with multiple mitochondrial DNA deletions. AB - In the last decade, several mitochondrial encephalomyopathies have been pathogenically associated with large-scale mitochondrial DNA deletions that are sporadic, or with point mutations that are maternally inherited. The mutations were also demonstrated in cultures of muscle satellite cells obtained from the patients. Subsequently, multiple deletions in mitochondrial DNA were found in several families. The affected members had progressive external ophthalmoplegia, cataracts and limb weakness, inherited as an autosomal dominant trait, or progressive external ophthalmoplegia with neurogastrointestinal encephalomyopathy or with cardiomyopathy, inherited as an autosomal recessive trait. To better understand the developmental pathobiology and localization of the multiple deletions, we performed comparative molecular genetic studies in muscle and cultures from patients. Whereas multiple deletions were found in muscle fragments from which muscle satellite cells were removed by enzymatic digestion, no deletions were found in the satellite cells or their cultured progeny. Our results suggest that multiple mitochondrial DNA deletions arise as somatic mutations during later stages of muscle development, or in terminally differentiated myofibers. PMID- 10540033 TI - Elevated S100 blood level as an early indicator of intraventricular hemorrhage in preterm infants. Correlation with cerebral Doppler velocimetry. AB - The aim of this study was to assess the use of S100 protein in blood as a means of identifying preterm infants at risk of intraventricular hemorrhage. In 25 preterm newborns, S100 blood concentrations were measured by an immunoradiometric assay during the first 48 h. Cerebral Doppler velocimetry waveform patterns were also tested at the time the blood sample was taken, when clinical and cerebral ultrasound scanning were still normal. Of the 25 newborns studied, 14 were controls and 11 developed intraventricular hemorrhage as revealed by ultrasound scanning more than 72 h after birth, and clinically confirmed by neurological examination on the seventh day of follow-up. S100 blood concentrations were significantly higher (P<0.002) in infants with intraventricular hemorrhage than in control infants and also correlated significantly (r=0.81, P<0.003) with the grade of hemorrhage. A significant correlation (r=0.70, P<0.05) between the S100 blood concentration and the middle cerebral artery pulsatility index was also observed. The present data show that S100 blood concentrations offer a measurable parameter of brain lesion in preterm infants before a radiological assessment of hemorrhage can be performed, when clinical symptoms may be silent and preventive/therapeutic action could be especially useful. PMID- 10540034 TI - MRI of the cauda equina in CIDP: clinical correlations. AB - Isolated reports have documented enhancement and/or enlargement of spinal nerve roots on magnetic resonance imaging (MRI) in patients with chronic inflammatory demyelinating polyradiculoneuropathy (CIDP). This work examines those findings in a consecutive series of 16 patients with CIDP, with blinded comparison to MRI in 13 disease controls, including five patients with Charcot-Marie-Tooth disease type 1A. MRI sequences consisted of T1 weighted sagittal and axial views, before and after administration of gadolinium. Blinded MRI interpretation was performed independently by two neuroradiologists. MRI results were correlated with data collected from chart review. Enhancement of the cauda equina was seen in 11 of 16 CIDP patients (69%), and in none of 13 control subjects. Nerve roots were enlarged, most significantly in the extraforaminal region, in three CIDP patients, and in one patient with Charcot-Marie-Tooth type 1A. MRI findings did not correlate with disease activity and severity, nor with any clinical or laboratory features in patients with CIDP. PMID- 10540035 TI - The RNA of the glutamate transporter EAAT2 is variably spliced in amyotrophic lateral sclerosis and normal individuals. AB - Impaired re-uptake of synaptic glutamate, and a reduced expression of the glutamate transporter EAAT2 have been found in the motor cortex of patients with amyotrophic lateral sclerosis (ALS). Two splice forms of the EAAT2 RNA resulting from retention of intronic sequences (EAAT2/Int) and deletion of one protein coding exon (EAAT2/C1) have been reported to account for the EAAT2 protein loss in ALS. In this study we investigated the presence of two known (EAAT2/C1; EAAT2/Int) and three novel (EAAT2/C2-4) EAAT2 RNA in motor cortex of 17 ALS cases and 11 controls. Reverse transcription and PCR were carried out to amplify the complementary DNA of the complete and variably spliced EAAT2 transcripts. Nested PCR was followed to generate amplicons specific for EAAT2/C1-4 and EAAT2/Int. EAAT2/Int was detected in 59% of ALS specimens as compared to 36% of controls showing a trend but no statistical significance of a more frequent expression in ALS (Type I error 24.6%). EAAT2/C1-4 were found to be equally expressed in ALS patients and controls. Our results indicate that the involvement of EAAT2 transcripts in ALS is unlikely to be primary, and more complex than previously recognized. Alterations of quantitative expression of distinct EAAT2 splice forms in ALS cannot be excluded from this study and remain to be investigated. PMID- 10540036 TI - Transcranial magnetic stimulation compared with upper motor neuron signs in patients with amyotrophic lateral sclerosis. AB - If patients with amyotrophic lateral sclerosis (ALS) present without upper motor neuron signs (UMNS) they do not meet current ALS research criteria. To compare how sensitively degeneration of upper motor neurons is detected clinically and by transcranial magnetic stimulation, 35 patients with ALS were studied. Nineteen patients had definite UMNS, nine patients had probable UMNS, and seven patients had no UMNS. Cortex, cervical nerve roots, and lumbar plexus were stimulated with a magnetic stimulator. Compound muscle action potentials from abductor digiti minimi and from anterior tibial muscles were recorded with surface electrodes. Responses to transcranial magnetic stimulation were considered abnormal if central motor conduction time was above the 99% upper limits or if there was no response to cortical but to peripheral stimulation. In all patients with definite UMNS central motor conduction was abnormal. In patients with probable UMNS it was abnormal in 67%, and in patients without UMNS it was abnormal in 71%. Abnormality of central motor conduction was neither correlated with the duration nor with the severity of the disease. The high rate of abnormalities of central motor conduction found in patients with ALS but without definite UMNS suggests that, in these patients, the diagnosis of ALS can be made more reliably if transcranial magnetic stimulation studies are performed. PMID- 10540037 TI - Focal cognitive impairment in mitochondrial encephalomyopathies: a neuropsychological and neuroimaging study. AB - Mitochondrial encephalomyopathies (ME) are a multisystemic group of diseases characterized by a wide range of biochemical and genetic mitochondrial defects with a variable mode of inheritance. We studied the neuropsychological profile, magnetic resonance imaging (MRI) and single photon emission computed tomography (SPECT) data in a group of ME patients in order to look for common or specific cognitive defects and a possible correlation with related brain areas. Three main cognitive areas were assessed: general intelligence, memory functions and visuo perceptual skills. Our sample included 16 ME patients (nine males, seven females) aged 25-68 years (mean age 45.2, SD 13.0). No sign of mental deterioration was found in the group of elderly subjects. Despite subjects showing no global cognitive impairment they scored lower in nonverbal versus verbal tasks. Visuo spatial skills and short-term memory were selectively impaired. There was no correlation between neuropsychological results and age, illness duration, age of onset, clinical phenotypes, genetic mitochondrial alterations and pharmacological therapy. The most frequent SPECT pattern observed was the hypoperfusion of temporal lobes, with a direct localization in the temporal cortex and with prevalent mesial involvement. The neuropsychological profile and SPECT imaging revealed similarities with focal defects. PMID- 10540038 TI - S-1 radiculopathy as a possible predisposing factor in focal myositis with unilateral hypertrophy of the calf muscles. AB - Associated with chronic S-1 radiculopathy, a 44-year-old man developed unilateral hypertrophy of the calf muscles. Electromyography revealed neurogenic alterations in the corresponding limb compatible with S-1 radiculopathy. In addition, MR tomographic and bioptic findings were consistent with a focal inflammatory myopathy of the enlarged right gastrocnemius muscle. Predisposing factors for the localisation of a focal myositis are unknown. This case report highlights the diagnostic difficulties in distinguishing focal myositis and denervation hypertrophy following S-1 radiculopathy or secondary inflammation related to denervation. We consider the possibility that in our case the inflammatory process might have been triggered by electromyographically proven chronic denervation related to radiculopathy. PMID- 10540039 TI - Recurrent steroid-responsive trismus and painful ophthalmoplegia. AB - A 63-year-old woman experienced two episodes of trismus and painful ophthalmoplegia at an interval of six years. She suffered left visual loss, and enhanced CT scan and MR imaging revealed heterogeneous enlargement of the left extraocular muscles extending to the orbital apex. In addition, the left pterygopalatine fossa was filled with a mass isointense with muscle without evidence of surrounding tissue invasion; 67Ga scintigraphy showed high uptake in this lesion. Steroid administration dramatically resolved the trismus, and the mass in the orbit and extraorbit vanished completely. Orbital pseudotumor is characterized by self-limited, relapsing, steroid-responsive painful ophthalmoplegia, and this case could be a variant of this entity with inflammation extending into the extraorbital area. PMID- 10540040 TI - Review article: liver support systems in acute hepatic failure. AB - The treatment of acute hepatic failure has developed rapidly over the last 40 years, reducing morbidity and mortality from this syndrome. Whilst this has been partly attributed to significant improvements in the specialist medical management of these patients, advances in surgical techniques and pharmaceutical developments have led to the establishment of successful liver transplantation programmes, which have improved mortality significantly. This review will examine the clinical impact of alternative methods that have been used to provide extra corporeal hepatic support. Non-biological, bio- logical and hybrid hepatic extra corporeal support will be explored, offering a comprehensive historical overview and an appraisal of present and future advances. PMID- 10540041 TI - Review article: nonsteroidal anti-inflammatory drug-associated gastrointestinal complications--guidelines for prevention and treatment. AB - Chronic ingestion of NSAIDs increases the risk for gastrointestinal complications, which range from dyspepsia to gastrointestinal bleeding, obstruction, and perforation. Among patients using NSAIDs, 0.1 to 2.0% per year suffer serious gastrointestinal complications. Patients who require analgesic therapy should be carefully assessed for the lowest possible dosage and shortest duration of NSAID use and for the potential of treatment with a non-NSAID pain reliever. These patients should also be assessed for factors that increase their risk of gastrointestinal complications, including increased age, concomitant anticoagulant or corticosteroid use, and past history of NSAID-associated gastrointestinal complications. The exact association between Helicobacter pylori infection and NSAID-related ulcer disease is unclear, and the routine testing and treatment of all NSAID using patients for H. pylori infection is not recommended at this time. NSAID-using patients who suffer from dyspepsia should have NSAIDs discontinued, the dosage changed, or be changed to a different class of NSAID. If NSAIDs cannot be discontinued, then an antisecretory agent should be initiated. Misoprostol prevents NSAID-associated gastrointestinal complications. Proton pump inhibitors are the most effective at healing NSAID-associated ulcers among patients who cannot discontinue NSAID therapy. PMID- 10540042 TI - The effect of dosing with omeprazole on the accuracy of the 13C-urea breath test in Helicobacter pylori-infected subjects. AB - BACKGROUND: The 13C-urea breath test (13C-UBT) is an accurate means of Helicobacter pylori diagnosis. However, proton pump inhibitors may suppress H. pylori and cause false negative results. AIM: To study the kinetics of H. pylori suppression by omeprazole during and after short-term use. METHODS: Volunteers underwent a baseline 13C-UBT (13C-urea 100 mg). H. pylori-positive subjects took omeprazole 20 mg daily for 14 days. Those who remained 13C-UBT positive (delta13CO2 >/= 5) continued omeprazole for a further 14 days. 13C-UBTs were performed weekly on omeprazole and then every second day after it was stopped. False negatives occurred when delta13CO2 fell to < 5. RESULTS: In 25 H. pylori positive subjects (mean age 43.9 +/- 2.4 years; 21 females, 4 males) the mean baseline delta13CO2 was 28.1 +/- 3.4. False negative breath tests occurred in three subjects after 7 days of omeprazole and in a further four subjects after 14 days. A further six subjects developed negative tests between Days 14 and 28. Following cessation of omeprazole, the 13C-UBT became positive again in 12/13 subjects within 4 days and in all within 6 days, with a mean recovery to 99.9 +/- 18.6% of baseline delta13CO2. CONCLUSIONS: False negative 13C-UBTs are common during treatment with omeprazole and occur after as little as 7 days. Return to positive test results is rapid after cessation of omeprazole. These findings are relevant to the timing of testing in clinical practice. PMID- 10540043 TI - Seroprevalence of cytotoxin-associated gene A positive Helicobacter pylori strains in Changle, an area with very high prevalence of gastric cancer in south China. AB - BACKGROUND: Helicobacter pylori, especially the CagA-positive strains, are closely associated with peptic ulcers and gastric cancers. We performed a large scale gastric cancer screening project and examined the prevalence of H. pylori and CagA-positive strains in Changle, China, an area with one of the World's highest gastric cancer mortality. We also compared the prevalence with that in Hong Kong which has one-tenth of the gastric cancer mortality of that in Changle. METHODS: A total of 2424 subjects in Changle and 523 subjects in Hong Kong had endoscopic examination and venesection. Sera were tested for anti-H. pylori antibody and anti-CagA antibody and correlated with endoscopic findings. RESULTS: In Changle, 80. 9% of the subjects were H. pylori carriers. Out of 551 carriers, 408 (74%) were positive for anti-CagA antibody. A total of 76% and 87% of the asymptomatic and gastric cancer patients were positive for anti-CagA antibody, respectively (P > 0.05). Compared to Hong Kong, there was a significantly (P < 0.0001) higher prevalence of CagA-positive strains in asymptomatic subjects in Changle (76%) than in Hong Kong (28%), but not in peptic ulcers or gastric cancers. CONCLUSIONS: Subjects in Changle had a high prevalence of H. pylori infection and a high prevalence of the CagA-positive strains. The contrast in the prevalence of CagA-positive strains, in asymptomatic subjects in two areas with differing gastric cancer mortality, supports the pathogenic role of CagA-positive strains in gastric carcinogenesis. PMID- 10540044 TI - The effect of Helicobacter pylori eradication therapy on gastric antral myoelectrical activity and gastric emptying in patients with non-ulcer dyspepsia. AB - BACKGROUND: Dysmotility of the gastroduodenal region and delayed gastric emptying have been considered to play roles in non-ulcer dyspepsia (NUD). Helicobacter pylori-induced inflammation of the gastric mucosa may affect gastric motility. AIM: To evaluate the effects of H. pylori eradication therapy on gastrointestinal motility and symptoms in NUD patients. METHODS: : Forty-six NUD patients were examined for gastric emptying, antral myoelectrical activity, H. pylori infection, and symptom scores. In H. pylori-positive NUD patients, gastric emptying, antral myoelectrical activity, and symptom scores were also analysed 2 months after cure of H. pylori infection. RESULTS: Sixty-seven per cent of NUD patients were H. pylori-positive. Both abnormal gastric emptying and antral myoelectrical activity were observed in NUD patients. H. pylori-positive NUD patients were divided into three groups according to their gastric emptying: the delayed group, the normal group, and the rapid group. In the delayed and rapid gastric emptying groups, the emptying and symptom scores were improved significantly by eradication. There was no improvement in symptom scores in the normal gastric emptying NUD group by the eradication therapy. CONCLUSIONS: Disturbed gastric emptying and antral myoelectrical activity play roles in NUD. H. pylori-induced disturbed gastric emptying may cause some NUD symptoms. Gastric emptying and symptom scores are improved by H. pylori eradication therapy in NUD patients with disturbed gastric emptying; H. pylori eradication therapy is effective in H. pylori-positive NUD patients with disturbed gastric emptying. PMID- 10540045 TI - Seven-day 'rescue' therapy after Helicobacter pylori treatment failure: omeprazole, bismuth, tetracycline and metronidazole vs. ranitidine bismuth citrate, tetracycline and metronidazole. AB - BACKGROUND: Eradication therapy with omeprazole (O), amoxycillin (A) and clarithromycin (C) is used extensively, although it often fails. A 'rescue' therapy with a quadruple combination of O, bismuth (B), tetracycline (T) and metronidazole (M) has been recommended. AIM: : To assess ranitidine bismuth citrate (Rbc) instead of O and B for treatment failure. METHODS: Sixty consecutive patients (13 duodenal ulcer, 47 non-ulcer dyspepsia) in whom a previous eradication trial with O, A and C had failed were randomized to receive one of two regimens for 7 days: O (20 mg b.d.), B (120 mg q. d.s.), T (500 mg q.d.s.) and M (250 mg q.d.s.) (group OBTM, n=30); or Rbc (400 mg b.d.), T (500 mg q.d.s.) and M (250 mg q.d.s.) (group RbcTM, n=30). Eradication was defined as a negative 13C-urea breath test 1 month after completing therapy. RESULTS: Mean age +/- s.d. was 45 +/- 12 years, 47% were males. Distribution of studied variables (age, sex, smoking, duodenal ulcer/non-ulcer dyspepsia) was similar in both therapeutic groups. Per protocol eradication was achieved in 17 out of 29 patients (59%) in group OBTM and in 25 out of 29 patients (86%) in group RbcTM (P < 0.05). Intention-to-treat eradication was achieved, respectively, in 17 out of 30 (57%) and in 25 out of 30 (83%) (P < 0.05). In the multivariate analysis the variables which influenced on H. pylori eradication were the type of therapy (odds ratio, OR=3.9; 95%CI: 1.02-15; P < 0.05) and diagnosis (duodenal ulcer/non ulcer dyspepsia) (OR=0.1; CI: 0.02-0.4). Adverse effects were infrequent and mild with both regimens. CONCLUSION: Therapy with RbcTM is a promising option after H. pylori eradication failure with OCA, achieving a higher efficacy than quadruple therapy with OBTM. PMID- 10540046 TI - Helicobacter pylori treatment instead of maintenance therapy for peptic ulcer disease: the effectiveness of case-finding in general practice. AB - BACKGROUND: Maintenance therapy with acid-inhibiting medication is common in general practice. Since the eradication of Helicobacter pylori has become the treatment of choice for peptic ulcer disease, H. pylori treatment could replace maintenance therapy in patients with an ulcer history. AIM: To determine the effectiveness of a full peptic ulcer disease history case-finding strategy, together with subsequent H. pylori testing and treatment, in discontinuing maintenance therapy. METHOD: Patients were included from seven general practices, who had been using acid-inhibiting medication for more than 3 months in the period May 1996-May 1997. Patients with a history of proven peptic ulcer disease were tested, and treated with proton pump inhibitor-triple therapy if positive. Maintenance therapy was discontinued and restart within 12 months was monitored. RESULTS: Long-term acid suppression was used by 2.8% of the practice-populations. A peptic ulcer disease history was found in 18% of the patients, 73% of whom were offered the 'H. pylori test and treat' alternative. The majority responded: 92% of the H. pylori-infected patients were treated, 78% of whom successfully discontinued long-term medication. CONCLUSION: Implementing an 'H. pylori test and treat' strategy enabled one-third of the patients with a peptic ulcer disease history to stop maintenance therapy successfully. The strategy contributes to reduction of long-term drug use, but compliance needs improvement. PMID- 10540047 TI - Low molecular weight heparin as adjuvant therapy in active ulcerative colitis. AB - BACKGROUND: Heparin given intravenously has shown beneficial effects in the treatment of refractory ulcerative colitis in open trials. Low molecular weight heparin (LMWH) offers advantages in the method of administration but have not been evaluated in inflammatory bowel disease conditions. AIM: To assess the tolerability and safety of subcutaneous self-administered LMWH in outpatients with refractory ulcerative colitis and to evaluate any potential adjuvant therapeutic effect. PATIENTS AND METHODS: Twelve patients with mild to moderately active ulcerative colitis were included in the trial. The patients had either responded poorly to treatment with conventional therapy, including oral and/or rectal glucocorticosteroids, or had experienced a rapid relapse during or shortly after GCS therapy. Dalteparin sodium 5000 units s.c. injection was administered twice daily for 12 weeks. Patients were monitored for possible adverse events and changes in clinical symptoms, and endoscopic and histological scores were analysed. Leucocyte scanning was performed at inclusion and at the end of the study. RESULTS: Tolerability and compliance were excellent and no serious adverse events occurred. Eleven patients improved symptomatically and six (50%) attained complete remission after 12 weeks of treatment. Endoscopic, scintigraphic and histological scores were found to be significantly improved. CONCLUSION: Self administered LMWH given s.c. may be a safe adjuvant therapy for patients with active, glucocorticosteroids-refractory ulcerative colitis. A controlled trial should be undertaken to confirm the positive effects found in this study. PMID- 10540048 TI - Interferon alfa-2b alone or in combination with ketoprofen as treatment for interferon-naive chronic hepatitis C patients. AB - BACKGROUND: Non-steroidal anti-inflammatory drugs may amplify the anti-viral effect of alpha-interferon in vitro but in vivo data are still controversial. AIM: : To test the hypothesis that ketoprofen may increase the rate of response to alpha-interferon of chronic hepatitis C patients. METHODS: Fifty patients with chronic hepatitis C who had never received alpha-interferon were randomly assigned to receive 3-8 MU of alpha2b-interferon, three times weekly for 6 months, alone or in association with ketoprofen at a dose of 200 mg/day five times weekly. The virological response to treatment (undetectable HCV RNA in serum) was evaluated after 3 months and at the end of treatment, and 6 and 12 months after therapy withdrawal. RESULTS: One patient under combination therapy stopped the ketoprofen for persisting epigastric pain. Complete response under treatment was observed in 15 out of 24 (62.5%) patients receiving alpha2b interferon alone and in 14 out of 26 (53.8%) patients under combination therapy (P=N.S.). One year after the end of treatment, a sustained response was seen in 4 out of 24 (16.2%) patients treated with alpha2b-interferon and in 5 out of 26 (19.2%) patients having received the combination (P=N.S.). CONCLUSION: Administration of ketoprofen does not increase either the primary or the sustained response to alpha2b-interferon therapy of interferon-naive chronic hepatitis C patients. PMID- 10540049 TI - Plasma hydroxy-metronidazole/metronidazole ratio can detect early changes in hepatic function in ethanol-induced liver injury. AB - AIMS: To evaluate the usefulness of plasma hydroxy-metronidazole/metronidazole (OH-MET/MET) ratios as a dynamic liver function test in ethanol abusers with or without liver cirrhosis. METHODS: Metronidazole was administered intravenously for 20 min to healthy volunteers, and to patients with alcohol-induced, non cirrhotic hepatopathy and liver cirrhosis. Plasma concentrations of metronidazole and hydroxy-metronidazole were measured by high performance liquid chromatography in samples collected 5, 10, 20 and 30 min after the metronidazole infusion. RESULTS: Patients with non-cirrhotic alcoholic hepatopathy had significantly elevated aminotransferase levels compared to healthy volunteers and Child A patients. Child-Pugh C patients had significantly prolonged prothrombin times when compared to healthy volunteers and patients with non-cirrhotic hepatopathy. Metronidazole metabolism, as measured by the OH-MET/MET ratio following the intravenous administration of 500 mg of the drug, was significantly impaired in all ethanol-abusing individuals, including patients with non-cirrhotic alcoholic hepatopathy. CONCLUSIONS: Metronidazole metabolism was impaired in ethanol abusers, even in the absence of liver cirrhosis, indicating that ethanol was capable of affecting liver function in the early stages of alcohol-induced liver disease. PMID- 10540050 TI - Atrophic gastritis during long-term omeprazole therapy affects serum vitamin B12 levels. AB - BACKGROUND: Omeprazole maintenance therapy for gastro-oesophageal reflux disease (GERD) has been associated with an increased incidence of atrophic gastritis in H. pylori-infected patients and with a decreased absorption of protein-bound, but not of unbound cobalamin. AIM: : To test the hypothesis that the combination of decreased cobalamin absorption and atrophic gastritis decreases serum cobalamin levels during omeprazole therapy. METHODS: Forty-nine H. pylori-positive GERD patients were treated with omeprazole for a mean (+/- s.d.) period of 61 (25) months. At the start of omeprazole treatment (T0) and at the latest follow-up visit (T1), serum was obtained for measurement of cobalamin. Corpus biopsy specimens were obtained at entry and follow-up for histopathological scoring according to the updated Sydney classification. RESULTS: At inclusion, none of the 49 patients had signs of atrophic gastritis. During follow-up, 15 patients (33%) developed atrophic gastritis, nine of whom had moderate to severe atrophy. These 15 patients did not differ from the other 34 patients with respect to age, serum cobalamin at T0 or the duration of follow-up. During follow-up, no change was observed in the median serum cobalamin level in the 34 patients without atrophy; (T0) 312 (136-716) vs. (T1) 341 (136-839) pmol/L (P=0.1). In the 15 patients who developed atrophy, a decrease in cobalamin was seen from 340 (171 to 787) at baseline to 285 (156-716) at latest follow-up (P < 0.01). CONCLUSIONS: The development of atrophic gastritis during omeprazole treatment in H. pylori positive GERD patients is associated with a decrease of serum vitamin B12 levels. PMID- 10540051 TI - Comparison of two different formulations of botulinum toxin A for the treatment of oesophageal achalasia. The Gismad Achalasia Study Group. AB - BACKGROUND: Intrasphincteric injection of botulinum toxin has been reported as a safe and effective alternative treatment in oesophageal achalasia, especially in high-risk and elderly patients. AIM: : To compare two formulations of botulinum toxin in the management of achalasia. PATIENTS AND METHODS: We randomly compared the efficacy and safety of 100 U of Botox (Allergan, Irvine, USA) and 250 U of Dysport (Ipsen, Milan, Italy), injected through a sclerotherapy needle at the level of the lower oesophageal sphincter, in 78 consecutive patients with achalasia. Symptom score, oesophageal manometry and 24 h pH-metry were recorded (before and 1 month after therapy). Symptom score was also obtained 6 months after treatment. RESULTS: One month after treatment, the effects of the toxin on symptoms and oesophageal tests were similar for both formulations. Lower oesophageal sphincter pressure decreased from 31 +/- 12 to 18 +/- 5 mmHg after Botox, and from 35 +/- 9 to 18 +/- 10 after Dysport. At the end of the follow-up period (6 months), symptom score decreased from 5 +/- 1.2 to 1.2 +/- 0.8 after Botox and from 5.2 +/- 1.5 to 1.5 +/- 1 after Dysport. Moreover, the percentages of patients who failed to respond to treatment (10% and 17.5%) and who relapsed during follow-up (12% and 24%) did not differ significantly. No patient complained of reflux symptoms after treatment, although abnormal acid exposure was documented in two subjects. CONCLUSIONS: Both formulations of botulinum toxin have comparable efficacy in the treatment of oesophageal achalasia, for up to 6 months of follow-up. PMID- 10540052 TI - The ulcer healing effect of protamine sulphate in rat stomach. AB - BACKGROUND: Protamine sulphate has been reported to stimulate nitric oxide production from blood vessels, which is a pivotal factor for gastric ulcer healing. Our preliminary study also showed that protamine sulphate potentiated the ulcer healing effect of heparin. METHODS: Male SD rats with acetic acid induced gastric ulcers were given protamine sulphate (40-80 mg/kg, s.c.) twice daily for 4 or 7 days. L-NG-nitroarginine methyl ester (L-NAME, 5 mg/kg), an inhibitor of nitric oxide synthase (NOS), was given s.c. prior to protamine sulphate (80 mg/kg) treatment. Ulcer healing, angiogenesis, mucosal histological changes, NOS activity and growth factors were determined. RESULTS: Protamine sulphate dose-dependently accelerated gastric ulcer healing, which was accompanied by a significant increase in angiogenesis, mucosal regeneration and constitutive NOS activity. Inhibition of gastric secretion was observed. Epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), tumour necrosis factor-alpha (TNF-alpha) or inducible NOS activity was also affected. L NAME completely blocked the beneficial effects of protamine sulphate. CONCLUSIONS: Protamine sulphate accelerates gastric ulcer healing through a mucosal nitric oxide-dependent and possibly also the EGF-and bFGF-associated pathways, which are followed by an increase of angiogenesis and mucosal regeneration. Acid inhibition contributes in part to the ulcer healing action of protamine sulphate. PMID- 10540053 TI - Injurious effect of Helicobacter pylori culture fluid to gastroduodenal mucosa, and its detoxification by sucralfate in the rat. AB - BACKGROUND: Helicobacter pylori plays an important role in the pathogenesis of peptic ulcer. Although several cytotoxins related to H. pylori have been reported, their effects on gastroduodenal mucosa have not been well evaluated in vivo. AIM: To investigate the effects of the combination of acid and toxic substances derived from H. pylori on gastroduodenal mucosa, and to observe the effect of sucralfate on such factors in the rat. METHODS: Male Sprague-Dawley rats were fasted overnight and anaesthetized. The pylorus was ligated, and a double-lumen cannula was inserted into the forestomach for gastric luminal perfusion. In other animals, a cannula was inserted to perfuse the proximal duodenum. 51Cr-EDTA was administered intravenously and mucosal integrity was monitored by measuring the blood-to-lumen 51Cr-EDTA clearance. After 72 h of culture of H. pylori (NCTC11637 and Sydney strain 1), Brucella broth containing 3% FBS was filtered to remove the bacteria (supernate of H. pylori culture fluid; HPsup). HPsup was acidified (pH=2.0) with HCl, and tested for its injurious action on gastric or duodenal mucosa by luminal perfusion. HPsup was incubated with sucralfate for 30 min. The supernate was collected by centrifugation and the pH was readjusted to 2.0. This sucralfate-treated HPsup was used to test the effect of sucralfate against H. pylori-related mucosal injurious factors. RESULTS: Non-acidified and acidified HPsup did not cause any detectable injury to the gastric mucosa. Non-acidified HPsup did not cause injury in the duodenal mucosa. However, acidified HPsup induced a significantly greater increase in 51Cr EDTA clearance and greater histological damage than in controls. Sucralfate completely reversed this. CONCLUSION: These results suggest that an H. pylori related toxic substance may aggravate duodenal acid injury by acting on luminal surfaces, and that the detoxification of this substance by sucralfate may contribute to its anti-ulcer action. PMID- 10540054 TI - The Do-Not-Attempt-Resuscitation ('DNAR') order: a lever to improve outcome and deliver preventative care. PMID- 10540055 TI - Personality traits of anaesthetists and physicians: an evaluation using the Cloninger Temperament and Character Inventory (TCI-125). AB - The personality profiles of Specialist Anaesthetists, Trainee Anaesthetists and Specialist Physicians were examined using Cloninger's Temperament and Character Inventory. These were compared with validated Community Sample 'average values' and a historical Norwegian Physician sample. Completed forms were returned from 364 doctors (Specialist Anaesthetists 222, Trainee Anaesthetists 75, Physicians 67), an overall response rate of 71%. Specialist Anaesthetists were more Cooperative, Harm Avoidant and Self-Directed than the Community Sample but less Reward Dependent, Novelty Seeking and Persistent than the Community Sample. Physicians were more Cooperative than their Specialist Anaesthetist colleagues, but both more so than were the general population. Trainee anaesthetists appear to be more Novelty Seeking and Reward Dependent than the Specialist Anaesthetists, this factor being predominately age related. Extreme/Mild personality traits were identified in 33% of Specialists, 41% of Trainees and 33% of Physicians, whilst personality disorders were found at the expected rates (Specialist Anaesthetists 9%, Trainee Anaesthetists 10%, Physicians 2%). Personality assessment has implications for recruitment, crisis management and professional development within anaesthesia. PMID- 10540056 TI - Ventilatory characteristics in mechanically ventilated patients during manual hyperventilation for chest physiotherapy. AB - We measured the tidal volumes and peak inflation pressures generated during manual hyperventilation for chest physiotherapy in 25 adult ventilated patients. The average tidal volume ranged from 711 to 1511 ml, with a mean (SD) of 1120 (274) ml. There was a negative correlation (p < 0.05) between the average tidal volume and the lung injury, as measured by the Murray lung injury score. The average peak inflation pressure ranged from 37 to 74 cmH2O with a mean (SD) of 51.5 (7.6) cmH2O. There was a positive correlation (p < 0.05) between average peak inflation pressure and the lung injury score. Mean (SD) PaO2 improved by 18.3 (14.3) kPa from baseline after physiotherapy (p < 0.05). Mean (SD) PaCO2 decreased by 0.1 (0.4) kPa. As the lung score increases higher inflation pressures and smaller tidal volumes are used suggesting an increased potential for barotrauma or volutrauma in susceptible lungs. PMID- 10540057 TI - An environmental survey of compliance with Occupational Exposure Standards (OES) for anaesthetic gases. AB - Environmental monitoring of nitrous oxide and volatile agents was carried out between August 1996 and October 1997 within operating theatre areas in eight hospitals within the Bro Taf Health Authority. Static monitoring and personal sampling were undertaken to assess compliance with the Occupational Exposure Standards introduced in January 1996 by the Health and Safety Executive for anaesthetic agents. The monitoring concentrated on nitrous oxide with the results showing that compliance was being achieved. Limited monitoring was carried out of the volatile agents, which again were well below the Occupational Exposure Limits. Monitoring was also carried out in nontheatre areas in which anaesthetic agents were used. The results show that many of these locations, such as delivery suites and radiology units, have inadequate ventilation and no anaesthetic gas scavenging, both of which combined to produce levels that exceeded the standards. PMID- 10540058 TI - The effect of rectal diclofenac on pruritus in patients receiving intrathecal morphine. AB - In this prospective randomised study, pruritus and pain were evaluated in patients undergoing abdominal surgery in which intrathecal morphine was administered. Each patient received intrathecal morphine 0.3 mg prior to induction, followed by a standard anaesthetic. The patients were randomly allocated to one of two groups. One group received 100 mg of rectal diclofenac immediately post-induction. Patients receiving diclofenac had significantly lower pruritus scores at 30 min (p = 0.0076), 2, 4, 8 and 24 h postoperatively, as well as significantly reduced pain scores at each time point (p < 0.0001 at each study interval). Morphine consumption in the first 24 h was also significantly lower in this group. In conclusion, rectal administration of diclofenac significantly reduces the incidence and severity of postoperative pruritus. It also significantly reduces pain and further analgesic requirements postoperatively. PMID- 10540059 TI - Intensive care: preparations for the Millennium. A survey of hospitals in the north of England. AB - In order to assess preparedness for the Millennium, we carried out a telephone survey of all Intensive Care Units in the former Northern Region. The survey disclosed wide variation in the current and planned provision of back-up power and central services, proposed staffing levels and expected demand. PMID- 10540060 TI - Definitions in obstetric anaesthesia: how should we measure anaesthetic workload and what is 'epidural rate'? AB - Crude delivery rate is used to calculate requirements for consultant anaesthetic sessions in the UK, but this calculation is arbitrary and ignores differences in case-mix between units. The term 'epidural rate' is commonly used to indicate regional anaesthetic activity but has never been defined. We challenge both these concepts and illustrate our argument by applying different definitions of obstetric anaesthetic activity to prospectively collected maternity data from 31 211 deliveries over 5 years in two hospitals. Number of anaesthetic interventions is a more accurate reflection of obstetric anaesthetic activity than number of deliveries, with Northwick Park Hospital having about 200-600 more deliveries per year than Chelsea & Westminster Hospital but about 300-400 fewer anaesthetic interventions per year. 'Epidural rate' varied by up to 30% according to the definition used. We conclude that number of anaesthetic interventions should replace crude number of deliveries as a measure of obstetric anaesthetic activity, and that the term 'regional anaesthesia rate' should replace 'epidural rate'. PMID- 10540061 TI - Syringe labels in anaesthetic induction rooms. AB - A survey of 35 hospitals in the United Kingdom has uncovered a wide variety of syringe drug labels. Use of different systems in different hospitals may result in wrong drug administrations, particularly when trainees move from one hospital to another. There is an urgent need to standardise the colour coding of syringe labels in the United Kingdom. Such standards are already in place in Australia, New Zealand and in the United States of America. This survey of syringe drug labels highlights the existing risks and recommendations for change are made. PMID- 10540062 TI - Testing of adult and paediatric ventilators for use in a magnetic resonance imaging unit. AB - We have assessed the performance of a series of ventilators (modified versions of the ventiPAC, paraPAC and babyPAC ventilators; SIMS pneuPAC Ltd, Luton, UK) in a magnetic resonance imaging (MRI) scanning environment, with MR safety and compatibility issues being addressed. Following initial modifications to remove ferromagnetic components and replace them with MR-safe materials, all three ventilators performed well in a series of tests in static magnetic fields up to 2 T. Ventilator performance was unaffected by static fields, switching gradients or radio frequency fields within the MR suite. Furthermore, the devices produced no degradation of image quality when used during MR scanning. We discuss management strategies for the care of critically ill ventilated patients during MR procedures. PMID- 10540063 TI - A simple method to control tracheal cuff pressure in anaesthesia and in air evacuation. AB - The pressure within latex balloons remains constant despite the balloons being inflated to more than 40 times their initial volume. We used this property to enable improved tracheal cuff pressure control. A latex balloon with an initial volume of 5 ml was connected via a vinyl duct attached with a roller clamp and three-way stopcock to a standard tracheal tube cuff. The 5 ml latex balloon was then inflated with 250 ml of air. The pressure within the tracheal tube cuff was monitored throughout anaesthesia with the inflated latex balloon acting as a pressure controller. Throat symptoms were recorded on the first four postoperative days. The controller kept the tracheal tube cuff pressures constant, and reduced the incidence of postoperative throat symptoms. Variations in cuff pressures with and without the controller were investigated in an altitude chamber to simulate flight. In the altitude chamber, cuff pressure reached over 200 cmH2O at 10 000 feet without the controller, whereas such variations were practically eliminated when the controller was used. PMID- 10540064 TI - A cuff pressure controller for tracheal tubes and laryngeal mask airways. AB - A simple device (U.K. Patent application no. 9907876.8) for fine adjustment of the cuff pressure of tracheal tubes and the laryngeal mask airway is described. In vitro tests confirm its efficiency. It is also a simple tool for training anaesthetic assistants in the operating theatre and nurses in the intensive care unit. PMID- 10540065 TI - Delayed pneumothorax and hydrothorax with central venous catheter migration. AB - We report a case of delayed pneumothorax, central venous catheter migration and iatrogenic hydrothorax in a 22-year-old female. The left subclavian central venous catheter initially transfixed the lung apex; pneumothorax occurred 24 h later following initiation of positive pressure ventilation. Lung collapse as a result of the pneumothorax caused catheter migration and hydrothorax. Catheter removal and chest drainage led to an uneventful recovery. PMID- 10540066 TI - Spinal cord injury following an attempted thoracic epidural. AB - Unsuccessful attempts were made to insert a thoracic epidural in an anaesthetised patient. Signs of spinal cord damage were observed the following day. Magnetic resonance imaging demonstrated a haematoma anterior to the spinal cord. Surgical exploration revealed an intradural haematoma and a needle puncture of the cord. The patient suffered a permanent paraparesis. PMID- 10540067 TI - Anaesthesia for caesarean section in a patient with recent subarachnoid haemorrhage and severe pre-eclampsia. AB - Subarachnoid haemorrhage is a leading 'indirect' cause of maternal death in the UK. We describe the case of a 43-year-old woman who presented with headache, photophobia and neck stiffness of sudden onset at 32 weeks' gestation. Cerebral computed tomography demonstrated subarachnoid blood in the cisterns around the midbrain, and oral nimodipine was started to prevent vasospasm. Preparations were made for endovascular coil embolisation in the event of identification of a posterior circulation aneurysm. However, angiography under general anaesthesia failed to reveal any vascular abnormality. On emergence from anaesthesia, headache persisted, and over the next 24 h severe pre-eclampsia developed. Magnesium sulphate was started, and urgent Caesarean section performed under general anaesthesia without incident. The rationale for the neuroradiological, obstetric and anaesthetic management is discussed. PMID- 10540068 TI - The effect of ionised magnesium on coagulation using thromboelastography. AB - Magnesium is an ionised mineral with therapeutic uses. There is laboratory evidence that it may have an anticoagulant activity although recent research has been to the contrary. The clinical implications of the effect of a therapeutic dose of magnesium on coagulation have yet to be resolved conclusively. In our study, 10 healthy volunteers were given 4 g of magnesium sulphate intravenously. Thromboelastographs were recorded and blood analysed for haematological indices, before and after the infusion. All variables associated with coagulation remained unchanged except the alpha angle on the thromboelastograph which increased significantly. We conclude that in our in vivo study, the effect of magnesium sulphate on coagulation is not clinically significant. PMID- 10540069 TI - Comparison of sevoflurane and total intravenous anaesthesia for daycase urological surgery. AB - Target-controlled total intravenous anaesthesia using propofol and remifentanil was compared with inhalational anaesthesia using sevoflurane and alfentanil in patients undergoing daycase urological surgery. Seventy-one patients were randomly allocated to receive either a target-controlled infusion of an admixture of propofol and remifentanil (125 microg of remifentanil added to 500 mg of propofol), or inhalational anaesthesia with sevoflurane and intra-operative alfentanil. There was no difference in time to fitness for discharge, time to fitness for transfer from primary to secondary recovery, time to first oral intake or adverse anaesthetic induction occurrences. Patient satisfaction assessed at 24 h post-discharge was high in both groups with no significant difference between groups. The incidence of nausea and vomiting was low in both groups. We conclude that, for ultra-short stay surgery, both the techniques we describe offer satisfactory anaesthesia with very early resumption of street fitness. PMID- 10540070 TI - Vaporisers. PMID- 10540071 TI - Ventricular fibrillation during spinal surgery PMID- 10540073 TI - MRSA contamination of a laryngoscope blade: a potential vector for cross infection. PMID- 10540074 TI - An unusual complication of tracheostomy. PMID- 10540075 TI - Cervical spine movements during laryngoscopy. PMID- 10540076 TI - No evidence for seeker needles. PMID- 10540078 TI - Damage to a central venous catheter. PMID- 10540079 TI - The spread and side-effects of intrathecally administered bupivacaine. PMID- 10540082 TI - Cephalad spread of epidural blockade in a 15-degree head-down position. PMID- 10540083 TI - Spinal haematoma following epidural anaesthesia in a patient with eclampsia. PMID- 10540084 TI - Nausea and vomiting during Caesarean section. PMID- 10540085 TI - A long 17-G vygon epidural needle PMID- 10540086 TI - Time to recheck the checklist? PMID- 10540087 TI - The left-handed laryngoscope. PMID- 10540088 TI - A long and dangerous journey: maternal mortality in Africa. PMID- 10540089 TI - Target-controlled remifentanil in combination with propofol for spontaneously breathing day-case patients. AB - Remifentanil is a new potent opioid with a very short duration of action irrespective of duration of infusion. It may have a role in day-case anaesthesia as part of a balanced total intravenous anaesthetic technique with propofol. We examined the respiratory depressant effects of remifentanil in 20 patients undergoing day-case anaesthesia. The target plasma concentration of remifentanil was varied while maintaining a constant target-controlled infusion of 4.5 microg x ml-1 propofol. In only 12 patients was satisfactory spontaneous respiration maintained. In these patients the median remifentanil target concentration was 1.6 ng x ml-1 and was achieved with a median infusion rate of 0.05 microg x kg-1 x min-1. The range of target concentrations associated with satisfactory spontaneous respiration was wide and varied over a 4.7-fold range from 0.6 to 2.8 ng x ml-1. PMID- 10540090 TI - Comparison of remifentanil with alfentanil or suxamethonium following propofol anaesthesia for tracheal intubation. AB - Sixty ASA physical status I and II, premedicated patients were administered propofol 2 mg x kg-1 and remifentanil 2 microg x kg-1 (group R), alfentanil 50 microg x kg-1 (group A) or suxamethonium 1 mg x kg-1 (group S) as a rapid bolus. One minute after study drug administration, tracheal intubation was performed. Intubation conditions were then scored. Excellent or good conditions were observed in only 35% in group R compared with groups S and A (100% and 85%, respectively; p < 0.001). The haemodynamic response to tracheal intubation was blunted in groups R and A compared with group S (p < 0.001). The mean heart rate in groups R and A was significantly lower than group S (p < 0.001). We conclude that remifentanil 2 microg x kg-1 given as a rapid bolus will not produce intubating conditions as good as those obtained with alfentanil 50 microg x kg-1 or suxamethonium 1 mg x kg-1 if administered after propofol 2 mg x kg-1. PMID- 10540091 TI - Dosing study of remifentanil and propofol for tracheal intubation without the use of muscle relaxants. AB - Sixty ASA I and II patients, premedicated with midazolam, were administered propofol 2 mg x kg-1 and remifentanil 3 microg x kg-1 (group R3), remifentanil 4 microg x kg-1 (group R4) and remifentanil 5 microg x kg-1 (group R5). Laryngoscopy and intubation were performed 1 min after the administration of the study drugs and the intubating conditions were assessed. Good to excellent conditions were observed in 12 patients in group R3 compared with 19 patients each in groups R4 and R5 (p = 0.004). Significant reductions in mean arterial pressure (MAP) and heart rate (HR) after administration of the study drug were observed in each group, p < 0.01. There was, however, no difference in mean MAP and HR between the three groups at all time points. We conclude that remifentanil 4-5 microg x kg-1 may reliably provide good to excellent conditions for tracheal intubation when administered after propofol 2 mg x kg-1. PMID- 10540092 TI - Respiratory mechanics during and after anaesthesia for major vascular surgery. AB - To evaluate the effects of major vascular surgery on respiratory mechanics, 11 patients undergoing general anaesthesia for abdominal aortic surgery were studied. Before aortic cross-clamping, chest wall elastance and resistance both increased (by 126% and 58%, respectively) when surgical retractors were placed. After aortic cross-clamping, lung elastance increased by 29%, accompanied by a decrease in cardiac index (22%) and an increase in pulmonary (17%) and systemic (15%) vascular resistance. After aortic unclamping, lung elastance decreased, although it remained higher than baseline values (by 12%). All cardiovascular variables returned to the values obtained before aortic cross-clamping. PMID- 10540093 TI - A comparison of ICU mortality prediction using the APACHE II scoring system and artificial neural networks. AB - The aim of this study was to compare the ability of artificial neural networks and the Acute Physiology and Chronic Health Evaluation II score to predict mortality in adult intensive care units. The same physiological variables were used in both predictive models to predict hospital mortality from a data set of 8796 patients collected from 26 adult intensive care units in the United Kingdom and Ireland as part of the Intensive Care Society study. The results from the two models were compared with the actual outcome. The overall prediction accuracy and the overall goodness-of-fit of all the models were assessed. Both predictive models showed similar goodness-of-fit and prediction discrimination. The overall predictive and classification performance of the artificial neural network developed matched and in some aspects was better than that of Acute Physiology and Chronic Health Evaluation II. PMID- 10540094 TI - Blood transfusion for Caesarean section in Malawi. A study of requirements, amount given and effect on mortality. AB - A prospective study of mothers needing Caesarean section was undertaken to examine the need for blood transfusion, the actual number of units given and the effectiveness of blood transfusion in preventing maternal mortality. Of 3665 mothers in 22 hospitals in Malawi, 11.1% were assessed as needing a transfusion and 7.2% were transfused. There were significant differences between district and central hospitals in transfusion rates. Of those mothers who were considered to need blood, there was no significant difference in mortality between those who received a transfusion and those who did not. PMID- 10540095 TI - Double-blind comparative study of droperidol, granisetron and granisetron plus dexamethasone as prophylactic anti-emetic therapy in patients undergoing abdominal, gynaecological, breast or otolaryngological surgery. AB - In this double-blind study the clinical efficacy of a single pre-operative intravenous dose of droperidol 1.25 mg (137 patients), granisetron 1 mg (130 patients) and granisetron 1 mg plus dexamethasone 5 mg (130 patients) was investigated for the prevention of postoperative nausea and vomiting after gynaecological surgery, breast surgery, abdominal surgery and ear, nose and throat surgery. The incidence of nausea in the first 24 h postoperatively was 52% in the droperidol group, 48% in the granisetron group and 34% with the combination, respectively. Both granisetron and granisetron/dexamethasone performed better than droperidol in their effects on vomiting or combined nausea and vomiting (incidence in the first 24 h 22%, 18% and 42%, respectively). The number of emetic episodes during the 5-day study period was significantly higher in the droperidol group (198) than in the granisetron (73) or combination group (78). PMID- 10540096 TI - Structured sedation programme for magnetic resonance imaging examination in children. AB - One thousand, eight hundred and fifty-seven patients underwent magnetic resonance imaging following the establishment of a structured sedation programme. Forty eight of these patients came from the intensive care unit with a secure airway and were therefore excluded from any further analysis. Oral sedation was to be given to children aged 5 years and below. For children >/= 6 years old, oral sedation could be given only if their level of co-operation was judged to be inadequate by the referring physician. Oral sedation consisted of chloral hydrate 90 mg x kg-1 (maximum 2.0 g) orally with or without rectal paraldehyde 0.3 ml x kg-1. All magnetic resonance imaging requests for children who failed oral sedation as well as those referred for general anaesthesia from the outset were reviewed by a consultant anaesthetist who then allocated patients to undergo the procedure with either general anaesthesia or intravenous sedation. Scans requiring intravenous sedation or general anaesthesia were performed in the presence of a consultant anaesthetist. Intravenous sedation consisted of either a propofol 0.5 mg x kg-1 bolus followed by an infusion (maximum 3 mg x kg-1 x h-1) or midazolam 0.2-0.5 mg x kg-1 boluses. General anaesthesia was given using spontaneous ventilation with a mixture of 66% nitrous oxide in oxygen and isoflurane following either inhalation (sevoflurane) or intravenous (propofol) induction. One thousand and thirty-nine (57.4%) of the scans were done without sedation whereas 93 scans were performed during the consultant anaesthetist supervised sessions. Oral sedation failed in 50 out of 727 patients (6.9%). Eighty-seven per cent of children aged 5 years and below needed sedation compared with 4.5% of those aged over 10 years. Two patients who had only received chloral hydrate developed significant respiratory depression. This structured sedation programme has provided a safe, effective and efficient use of limited resources. PMID- 10540097 TI - Organophosphorus poisoning and anaesthesia. AB - Organophosphorus compounds, used as insecticides and agents of chemical warfare, are a major global cause of health problems. These irreversible inhibitors of cholinesterase produce three well-recognised clinical entities: the initial cholinergic phase, which is a medical emergency often requiring management in an intensive care unit; the intermediate syndrome, during which prolonged ventilatory care is necessary; and delayed polyneuropathy. In addition, disturbances of body temperature and endocrine function, electrolyte imbalances, immunological dysfunction and disorders of reproduction have been reported in animals and man. Vocal cord paralysis, pancreatitis, cardiac arrhythmias and a wide range of neuropsychiatric disorders are known to follow acute and chronic exposure to organophosphorus compounds. As a result of the inhibition of plasma cholinesterase, there can be increased sensitivity to drugs hydrolysed by this enzyme, e.g. suxamethonium and mivacurium. The inhibition of acetylcholinesterase causes dysfunction at the neuromuscular junction which can produce altered responses to nondepolarizing neuromuscular blockers. Anaesthetists may encounter patients exposed to organophosphorus compounds either following acute poisoning, trauma (warfare) or as patients with a wide range of nonspecific disorders presenting for surgery. The traditional use of oximes and atropine in treatment has failed to reduce the morbidity and mortality associated with poisoning. The roles of agents that have reduced the toxicity of organophosphorus compounds in animal experiments are discussed as potential therapeutic agents. There is an urgent need for accurate information on the problems associated with exposure to organophosphorus compounds. This would best be achieved by collaborative research between technologically advanced countries and developing countries, where organophosphorus compounds are a leading cause of ill health. PMID- 10540098 TI - Use of the cuffed oropharyngeal airway for manual ventilation by nonanaesthetists. AB - We studied the use of the cuffed oropharyngeal airway in 100 ASA I and II anaesthetised patients. In the first 50 patients (group A), an experienced anaesthetist inserted the airway. The optimum sizes and cuff volumes for manual ventilation in adult males and females were found to be sizes 11 and 10 with up to 60 ml and 50 ml in each cuff, respectively. Manual ventilation was clinically successful in 49/50 (98%) of these patients. Using these recommendations and following a brief tutorial, a group of 50 nonanaesthetic, basic life-support providers attempted to insert a cuffed oropharyngeal airway and manually ventilate the lungs of a subsequent 50 patients (group NA). Clinically adequate tidal volumes were achieved within 45 s in 47/50 (94%) patients in this group. A persistent leak was present in 21/49 (43%) and 24/47 (51%) of the successful insertions in each group, but this did not affect the ability to ventilate the lungs adequately. The cuffed oropharyngeal airway may offer an effective method of providing adequate ventilation during resuscitation by nonanaesthetic hospital staff. PMID- 10540099 TI - 'Near-miss' hyperkalaemic cardiac arrest associated with rapid blood transfusion. AB - A case is presented in which a relatively modest blood transfusion resulted in acute hyperkalaemia with a 'near-miss' cardiac arrest. While transfusion-related hyperkalaemia usually occurs in association with massive transfusions, several factors may have increased the risk of such an acute reaction. A high index of suspicion is required, especially in patients with risk factors. Anaesthetists should not be lulled into a false sense of security simply because modest volumes of blood are being transfused. PMID- 10540100 TI - Aspiration in severe trauma: a prospective study. AB - The incidence and origin of contamination of the vocal cords in 53 trauma patients was studied when tracheal intubation was performed before hospital admission. Eighteen patients (34%) had gross contamination which was blood in 15 patients and gastric contents in three patients. This has implications for prehospital airway management and particularly for use of the laryngeal mask airway. PMID- 10540101 TI - Relationship between stimulating current and accelographic train-of-four response at the great toe. AB - We investigated the accelographic train-of-four response evaluated at the great toe at varying stimulating currents. Fifteen adult patients undergoing elective general anaesthesia were studied. The mean current at which a supramaximal T1 value could be elicited was > 49 (9) mA [mean (SD)]. Ratios of accelographic T1 values at 50, 40, 30, 20 and 10 mA to accelographic T1 value at 60 mA were 0.90 (0. 18), 0.58 (0.37), 0.38 (0.37), 0.19 (0.26) and 0.00 (0.00), respectively [mean (SD), p < 0.05 for 50 mA vs. 30, 20 and 10 mA, p < 0.05 for 40 mA vs. 20 and 10 mA, and p < 0.05 for 30 vs. 10 mA]. Threshold currents for train-of-four (the lowest currents at which any train-of-four response could be elicited) before and after induction of anaesthesia were 30 (10) and 31 (10) mA, respectively. The train-of-four ratio (T4/T1) measured at varying currents did not differ significantly. However, in the patients in whom threshold currents for train-of-four were 40, 30 and 20 mA, the train-of-four ratio recorded at the threshold current was significantly less than at 50 mA. We conclude that at the great toe, the mean current at which a supramaximal response to train-of-four can be yielded is as high as > 49 mA. The mean threshold currents for TOF before and after induction of anaesthesia were 30 and 31 mA, respectively. Train-of-four ratio measured at the threshold current is less than that at 50 mA. PMID- 10540102 TI - An epidural scoring scale for arm movements (ESSAM) in patients receiving high thoracic epidural analgesia for coronary artery bypass grafting. AB - Thoracic epidural analgesia appears to improve the outcome of patients undergoing coronary artery bypass graft surgery. Cranial extension of nerve blockade involving the third, fourth and fifth cervical nerve roots can cause apnoea. However, progressive paraesthesia and weakness due to cephalad spread of thoracic epidural analgesia will affect the arms before the diaphragm. A scale was designed to test three active movements of the arms bilaterally: hand grip (T1/C8), wrist flexion (C8/7) and elbow flexion (C6/5). This epidural scoring scale for arm movements (ESSAM) consists of four grades (0-3) based on the number of absent movements, and suggests appropriate action. The reliability of this scale was tested in 40 patients undergoing coronary artery bypass surgery. Twelve of the 40 patients had their epidural infusion reduced on the basis of the scale. Of these 12 patients, eight had a worst ESSAM score of 1, three had a worst score of 2 and one had a worst score of 3. In each patient, motor power returned following the reduction in infusion rate, taking between 30 min and 3 h. This scale appears to be a simple and reliable method for the early detection of the cephalad spread of thoracic epidural analgesia. PMID- 10540103 TI - Dental anaesthesia. PMID- 10540104 TI - Anaesthetic machines. PMID- 10540105 TI - The use of mini-dose suxamethonium to facilitate the insertion of a laryngeal mask airway. PMID- 10540106 TI - The obstructed airway. PMID- 10540107 TI - Documentation of grade of laryngoscopy and intubation difficulty. PMID- 10540108 TI - The last resort--follow the instructions! PMID- 10540109 TI - The Triennial Report--was it a 'direct death'? PMID- 10540110 TI - Ambulatory labour analgesia. PMID- 10540111 TI - Local anaesthetic loss during spinal injection. PMID- 10540112 TI - Faulty Tuohy needle. PMID- 10540113 TI - Electrical hazards from surgical equipment, identified by the diathermy machine. PMID- 10540114 TI - Inadequate training? PMID- 10540115 TI - Meningioma revealed after general anaesthesia. PMID- 10540116 TI - Peri-operative asystole in a patient with diabetic autonomic neuropathy. PMID- 10540117 TI - Methoxamine in the management of severe anaphylaxis. PMID- 10540118 TI - All under the umbrella of anaesthesia. PMID- 10540119 TI - A new use for Magill's forceps. PMID- 10540120 TI - Hypoxia caused by body piercing. PMID- 10540121 TI - Adjuvant radiotherapy for rectal cancer. PMID- 10540122 TI - Clinical dilemma. Retrosternal goitre. PMID- 10540123 TI - Evidence that metastasis is less common in cirrhotic than normal liver: a systematic review of post-mortem case-control studies. AB - BACKGROUND: It has been hypothesized that the cirrhotic liver is afforded protection against metastasis. The evidence has been examined and the plausibility of such a phenomenon is reviewed. METHODS: A systematic literature review was conducted with analysis of combined data from post-mortem case-control studies. RESULTS: Overall, the crude rate of metastasis to normal liver was 37.3 per cent, while the rate to cirrhotic liver was 23.7 per cent. The Mantel Haenszel (MH) fixed-effects estimate of the odds ratio was 0. 47 (95 per cent confidence interval (c.i.) 0.41-0.53; chi2 = 136, 11 d.f., P < 0.001). The DerSimonian-Laird (DL) random-effects estimation of the odds ratio was 0.42 (95 per cent c.i. 0.31-0.58; chi2 = 28, 1 d.f., P < 0.001). For tumours arising within the distribution of the portal vein, the crude rate of metastasis to normal liver was 47.6 per cent, whereas the rate to cirrhotic liver was 29.8 per cent. The MH estimate of the odds ratio was 0.45 (95 per cent c.i. 0.37-0.54; chi2 = 68.2, 5 d.f., P < 0.001). The DL pooled odds ratio was 0.44 (95 per cent c.i. 0.28-0.70; chi2 = 12.3, 1 d.f., P < 0.001). The MH and DL pooled estimates of the odds ratio were similar for groups of patients from the East (Japan) and the West (Europe and the USA). CONCLUSION: The post-mortem evidence reviewed suggests that the likelihood of metastasis to the cirrhotic liver is lower than that to normal liver. The degree of protection for tumours arising from within the distribution of the portal vein is neither greater nor less than it is overall. Eastern and Western populations appear to have a similar degree of risk reduction. The differences noted were significant on testing in the meta analysis, but confounding bias accounting for these differences has not been excluded. PMID- 10540124 TI - Illustrated review of new imaging techniques in the diagnosis of abdominal wall hernias. AB - BACKGROUND: The assessment of abdominal wall hernias has long been a clinical skill that only occasionally required the supplementary radiological assistance of herniography. However, with the advent of cross-sectional imaging, a new range of diagnostic tools is now available to help the clinician in difficult cases. METHODS: This review explores the ability of computed tomography and magnetic resonance imaging to demonstrate many of the hernias encountered in the anterior abdominal wall. Also discussed is the role of imaging techniques in the management of a variety of hernias. RESULTS AND CONCLUSION: Cross-sectional imaging techniques are being employed with increasing frequency for the assessment of hernias. Although the anatomical detail can usually be delineated clearly, the accuracy of the various methods and their place in the clinical management of hernias has yet to be fully determined. PMID- 10540126 TI - Digest PMID- 10540125 TI - Digest PMID- 10540127 TI - Partial ileal resection for hypercholesterolaemia in patients undergoing surgery for obesity. PMID- 10540128 TI - Clinical study of adjuvant photodynamic therapy to reduce restenosis following femoral angioplasty. AB - BACKGROUND: Photodynamic therapy (PDT) reduces neointimal hyperplasia and negative remodelling following balloon injury in small and large animal models. This clinical study investigated the role of adjuvant PDT following femoral percutaneous transluminal angioplasty (PTA). METHODS: Eight PTAs in seven patients (two women) with a median age of 70 (range 59-86) years were performed with adjuvant PDT. All patients had previously undergone conventional angioplasty at the same site which resulted in symptomatic restenosis or occlusion between 2 and 6 months. Each was sensitized with oral 5-aminolaevulinic acid 60 mg/kg, 5-7 h before the procedure. Following a second femoral angioplasty, up to 50 J/cm2 red light (635 nm) was delivered to the angioplasty site via a laser fibre within the angioplasty balloon. Patients were kept in subdued light overnight and discharged the following day. Outcome was assessed by duplex imaging at 24 h, 1, 3 and 6 months and by intravenous digital subtraction angiography at 6 months. A peak systolic velocity ratio (PSVR) of more than 2.0 at the angioplasty site was taken to represent restenosis. RESULTS: All patients tolerated the procedure well without adverse complications or death. All were rendered asymptomatic which was sustained throughout the study interval. All vessels remained patent and no lesion attained the duplex definition of restenosis. Median (interquartile range) PSVR across stenotic segments was 4.7 (3.7-5.7) before angioplasty, 1.1 (0.9-1.3) at 24 h and 1.4 (1.0-1.8) at 6 months after intervention (P = 0.04 compared with preoperative value). CONCLUSION: This pilot study suggests that endovascular PDT is safe and may reduce restenosis follow- ing angioplasty. The data justify a randomized controlled trial. PMID- 10540129 TI - Comparison of the expression of fibrosis-associated genes in glomeruli after renal transplantation between conventional cadaveric and non-heart-beating donors. AB - BACKGROUND: The main difference between cadaveric heart-beating donors and non heart-beating donors (NHBDs) is the degree of warm ischaemia to which the kidney is subjected. This study was designed to see if this affected the expression of fibrosis-associated genes in the early period after transplantation. METHODS: A series of 29 cadaveric and 19 NHBD renal transplants was studied. Patients underwent protocol needle-core renal transplant biopsies at 1 week, 3 months and 6 months after transplantation. At least two individual glomeruli were isolated from each biopsy. Messenger RNA was extracted and genes of interest were amplified by reverse transcriptase-polymerase chain reaction, then quantified in an enzyme-linked immunosorbent assay system. RESULTS: Delayed graft function was common in NHBD (17 of 19) compared with cadaveric transplants (six of 29) (P < 0.0001). Acute rejection rates were similar. The level of tissue inhibitor of metalloproteinase 1, an inhibitor of extracellular matrix degradation, was higher in kidneys from NHBDs at 1 week (P = 0.02). There were no other statistically significant differences in the expression of fibrosis-associated genes between the two groups. CONCLUSION: Although the increased ischaemic injury in kidneys retrieved from NHBDs leads to a higher rate of delayed graft function, this does not translate into increased expression of fibrosis-associated genes after the first week. PMID- 10540130 TI - Oral nifedipine reduces resting anal pressure and heals chronic anal fissure. AB - BACKGROUND: Topical preparations have been used in the treatment of anal fissure. However, they are not universally successful and there is confusion over the site and dose of application. This study assessed the effectiveness of oral nifedipine in reducing resting anal pressure and on fissure healing. METHODS: Anal manometry was performed on eight healthy volunteers and 15 patients with chronic anal fissure before and after oral administration of nifedipine 20 mg. Nifedipine was taken twice daily. Fissure healing was assessed over an 8-week period and pain scores were monitored. RESULTS: Oral nifedipine produced an initial reduction in maximum resting anal pressure (MRP) of 35 per cent (P < 0.001) and of 28 per cent after 5 days (P < 0.001) in healthy volunteers. A reduction in MRP of 36 per cent (P < 0.001) was observed in patients with fissure. Pain scores were significantly reduced during the treatment period. Healing was complete in nine patients after 8 weeks and a further three were asymptomatic. Ten patients experience flushing and four had mild headaches. There were no episodes of postural hypotension or incontinence. CONCLUSION: Oral nifedipine reduces resting anal pressure. It is well tolerated and offers an alternative treatment for chronic anal fissure. PMID- 10540131 TI - Treatment and outcome of intracystic papillary carcinoma of the breast. PMID- 10540132 TI - Improving cytological diagnosis and surgical management of parotid adenolymphoma. AB - BACKGROUND: The role of fine-needle aspiration cytology (FNAC) in the diagnosis and management of discrete parotid swellings remains controversial. Controlled enucleation can be appropriate with accurate preoperative diagnosis. This study (1985-1995) reviewed the role of FNAC in the diagnosis and surgical management of adenolymphoma. METHODS: Review of cytological smears by two observers concentrated on the features of infarction and squamoid metaplasia. Sensitivity, interobserver and intraobserver variation were evaluated statistically in a two run 'blinded' analysis of 80 cytological slides from a variety of lesions. RESULTS: Of 222 epithelial neoplasms of the parotid, 33 were adenolymphomas. FNAC was performed before operation in 32, producing 34 slides, and a correct cytological diagnosis was made in 21 patients. Retrospective review of the 34 slides, to examine specific features of squamoid metaplasia and infarction, improved diagnostic accuracy. The reliability and reproducibility of cytodiagnosis was confirmed by analysis of interobserver and intraobserver agreement. The sensitivity was high (0.76-0.88). Controlled enucleation was performed in 12 patients and superficial parotidectomy in 11. There were no tumour recurrences. CONCLUSION: Attention to the features of squamoid metaplasia and infarction improves cytological diagnosis and directs appropriate surgical management. PMID- 10540133 TI - Emerging trends in the management of the impalpable testis. AB - BACKGROUND: The management of the impalpable undescended testis is controversial. The study examines emerging trends in the management of this problem. METHODS: Two groups of boys were treated consecutively and recorded prospectively from 1974 to 1984 and from 1990 to 1998 inclusive. A consistent policy of using the preperitoneal approach for impalpable testis was adopted during both time intervals but during the second study period examination under anaesthesia and diagnostic laparoscopy were introduced to ascertain testicular presence and location. RESULTS: Some 919 boys were treated for cryptorchidism during the study period. Ninety boys in the first group (23 per cent) underwent preperitoneal explorations for impalpable testes. Anorchia was present in 18 and orchidectomy was performed in two boys. Thirty boys in the later group (5 per cent) were diagnosed as having impalpable testes. Fifteen boys underwent successful preperitoneal orchidopexy, anorchia was present in 11 and four underwent orchidectomy, carried out for high intra-abdominal testes. CONCLUSION: Examination under anaesthesia and subsequent laparoscopic assessment for all impalpable testes has reduced the need for preperitoneal exploration for the impalpable undescended testis. In this large series, division of the testicular vessels in order to secure scrotal placement of the testis was required in one instance only. PMID- 10540134 TI - Experience with the preperitoneal 'plug and patch' inguinal hernia repair. PMID- 10540135 TI - Survival of patients with colorectal cancer diagnosed in a randomized controlled trial of faecal occult blood screening. AB - BACKGROUND: Analysis of survival of subjects with colorectal cancer diagnosed by different modalities can provide insight into the mechanism by which screening has an effect. It can also give an indication of the feasibility of using prognostic indicators as surrogate outcome measures to predict mortality in future studies. METHODS: This paper examines the survival of individuals with colorectal cancer diagnosed in the Nottingham trial and explores the role of selected prognostic factors as possible surrogate outcome measures. RESULTS: Survival was significantly better in subjects with screen-detected cancers than in controls, even after adjusting for tumour stage and accounting for lead-time bias. Survival was inversely related to stage of tumour, with patients with stage A tumours having the best survival. Subjects with well or moderately differentiated tumours had a significantly better survival than those with poorly differentiated tumours. CONCLUSION: Screening for colorectal cancer by means of faecal occult blood testing improved survival among subjects with screen-detected cancers. Differences in prognostic factors largely explain the differences in survival between both non-responders and subjects with interval cancers and those in the control group, but not the improved prognosis for patients with screen detected cancers. The use of such factors as surrogate outcome measures may therefore be inappropriate. PMID- 10540136 TI - Randomized controlled trial to examine the influence of thoracic epidural analgesia on postoperative ileus after laparoscopic sigmoid resection. AB - BACKGROUND: The aim of the study was to evaluate whether perioperative epidural analgesia had any effect on the duration of postoperative ileus after laparoscopic sigmoid resection. METHODS: Twenty patients were randomized to surgery either with (group 1; n = 10) or without (group 2; n = 10) thoracic epidural analgesia. The major endpoint of the study was the time to the first postoperative bowel movement. Secondary endpoints were the interval until oral feeding was tolerated, incidence of postoperative vomiting, postoperative analgesic consumption use of patient-controlled analgesia (PCA) until the fourth day after operation, subjective pain perception and the incidence of epidural related side-effects. RESULTS: Age, sex and American Society of Anesthesiologists classification were similar in the two groups. The first bowel movement was documented after a median of 54 (95 per cent confidence interval 32-127) h in group 1 and 77 (31-99) h in group 2 (P = 0.8). Oral feeding without additional parenteral therapy was tolerated after 48 (40-64) h in group 1 and after 56 (48 64) h in group 2 (P = 0.6). Postoperative vomiting occurred in two patients from each group. During epidural therapy the use of PCA was lower in group 1 (0.30 (0.19-0.96) mg morphine per kg) than in group 2 (0.56 (0.37-0. 80) mg/kg) (P < 0.05). Postoperative pain perception during rest and while coughing was similar in both groups. Three patients experienced reversible side-effects of epidural therapy (motor deficit, two patients; bladder dysfunction, one). CONCLUSION: Perioperative thoracic epidural analgesia did not have a clinically relevant effect on the duration of postoperative ileus after laparoscopic sigmoid resection. PMID- 10540138 TI - High early mortality rate from acute pancreatitis in Scotland, 1984-1995. AB - BACKGROUND: Death from acute pancreatitis within the first week after admission is usually a consequence of multiple organ dysfunction. Reports from specialist centres suggest that, with improvements in resuscitation and supportive care, such deaths are becoming uncommon but it is unclear if this is reflected in a decrease in early mortality rate from acute pancreatitis in the general population. METHODS: Data concerning patients discharged with a diagnosis of acute pancreatitis (International Classification of Disease-9 code 577.0) between 1984 and 1995 were obtained from the Information and Statistics Division, National Health Service in Scotland, and analysed on a computer database. RESULTS: The incidence of acute pancreatitis in Scotland continues to increase in both sexes. The in-hospital mortality rate (death from all causes) was 7.5 per cent and showed a slight but significant downward trend over the period of study. Death within 7 days of hospital admission accounted for 53.7 per cent of all deaths and the proportion of early deaths did not decline over the study interval. CONCLUSION: These results suggest that scope remains for considerable improvement in the early management of acute pancreatitis. There is an urgent need to improve the early recognition of severe pancreatitis coupled to a willingness on behalf of clinicians to transfer these patients at an early stage to a centre with high-dependency and intensive care facilities supervised by a multidisciplinary team with expertise in the endoscopic, radiological and surgical management of these patients. PMID- 10540137 TI - Early ascorbic acid depletion is related to the severity of acute pancreatitis. AB - BACKGROUND: Ascorbic acid (AA) is an important endogenous antioxidant in plasma and has been shown to be decreased at the time of hospital admission in patients with acute pancreatitis. The aim of this study was to determine whether plasma AA concentration continues to decrease after admission and whether the extent of decrease is related to the severity of pancreatitis. METHODS: Consecutive patients with mild (n = 62) and severe (n = 23) acute pancreatitis had plasma AA concentration measured on the day of recruitment and on days 2 and 5 by high performance liquid chromatography. RESULTS: The plasma AA concentration in patients with acute pancreatitis was significantly less than that in normal volunteers on days 0, 2 and 5 (P < 0.0001) and this was more marked in those with severe disease. There was a decrease in plasma AA concentration from day 0 to day 2 in patients with mild (P < 0.0001) and severe (P = 0.0005) pancreatitis, and from day 2 to day 5 in patients with severe pancreatitis (P = 0.023). CONCLUSION: Endogenous plasma AA continues to decrease over the first 5 days in hospital and the extent is related to the severity of acute pancreatitis. Presented to a meeting of the Australasian Surgical Research Society, Auckland, New Zealand, August 1995 and published in abstract form as Aust N Z J Surg 1996; 66: 243 PMID- 10540139 TI - Evaluation of pylorus-preserving pancreatoduodenectomy with the Imanaga reconstruction by hepatobiliary and gastrointestinal dual scintigraphy. AB - BACKGROUND: Following pylorus-preserving pancreatoduodenectomy (PPPD), most surgeons use gastrointestinal reconstruction with an end-to-side duodenojejunostomy placed distally to the pancreatojejunostomy and choledochojejunostomy. In contrast, the authors have consistently used PPPD with the Imanaga reconstruction (PPPD-Imanaga) which consists of end-to-end duodeno- jejunostomy, end-to-side pancreatojejunostomy and choledochojejunostomy, performed in that order. In this study, the movement of bile and food after PPPD Imanaga was evaluated to document the functional advantages of this method. METHODS: Twenty-four patients who had undergone PPPD-Imanaga were subjected to hepatobiliary and gastrointestinal dual scintigraphy. The interval between operation and scintigraphy ranged from 28 days to 67 months. Six of the 24 patients underwent repeated dual scintigraphy for the observation of temporal changes in gastrointestinal function. RESULTS: The incidence of biliogastric reflux and bile stasis in the jejunal loop was markedly decreased at times later than 2 months after operation. Delay of gastric emptying and bile evacuation, sometimes accompanied by stasis in the jejunal loop, affected the mixing status of bile and food at 1 h after the beginning of imaging. A majority of the patients, however, had a satisfactory mixing status at 2 h. CONCLUSION: The Imanaga reconstruction appears to be a recommendable procedure following PPPD, in light of the bile and food movement achieved in the gastrointestinal tract. PMID- 10540141 TI - A 14-year experience with 6 cm as a criterion for surgical treatment of abdominal aortic aneurysm. AB - BACKGROUND: It remains unclear when to recommend operation for an asymptomatic abdominal aortic aneurysm (AAA). This study examined a prospective series of patients for whom standard criteria were applied. METHODS: Some 584 consecutive patients with an AAA of diameter 3 cm or greater detected by ultrasonographic screening have been observed for up to 14 years. Repeat ultrasonographic examinations have been performed at intervals. Surgery was not considered unless the aneurysm measured 6 cm in diameter, expanded at a rate equivalent to at least 1 cm per year, caused the patient symptoms, or an iliac aneurysm was present that required treatment. RESULTS: Operation was performed on 127 patients; the majority (80; 63 per cent) had an aneurysm that reached 6 cm in diameter. Use of the above criteria prevented rupture in all but 24 (4 per cent) of the 584 patients over the 14-year interval. Of these 24 patients, 11 were unfit for planned surgery and eight declined operation or follow-up. Rupture in the five remaining patients (1 per cent) who were available for treatment compared favourably with the reported 30-day mortality rate for elective surgical treatment of 1.4-12 per cent. CONCLUSION: Repeated observation is preferable to surgical intervention until an aortic aneurysm measures 6 cm in diameter, expands by 1 cm per annum or causes symptoms. Presented as a poster to the 52nd Annual Meeting of the Society for Vascular Surgery, San Diego, California, USA, June 1998 PMID- 10540140 TI - Endoscopic totally extraperitoneal repair of bilateral inguinal hernias. AB - BACKGROUND: Recurrence rates associated with bilateral inguinal hernia repair with a giant prosthesis (Stoppa procedure) are low. Endoscopic totally extraperitoneal bilateral inguinal hernia repair with a giant prosthesis combines the low recurrence rate of the Stoppa repair and the advantages of minimally invasive surgery. The aim of this retrospective study was to investigate whether extraperitoneal bilateral inguinal hernia repair could be performed by the minimally invasive, totally extraperitoneal approach. METHODS: From February 1993 to January 1998, 98 patients with bilateral inguinal hernias underwent surgery. A polypropylene 30 x 10 cm rectangular mesh or a 30 x 10/15 cm 'slipmesh' was used. Follow-up, including a physical examination, of 96 per cent of patients was performed. RESULTS: Median operative time was 60 min. Mostly minor intraoperative complications occurred. Conversion was required for two patients. Apart from one patient with a necrotic fasciitis who died from respiratory failure, only minor postoperative complications (10 per cent) occurred. Median hospital stay was 1 (range 1-21) days. Median recuperation time was 5 (range 1-22) days. Median follow-up (96 per cent) was 32 (range 7-57) months; there were six recurrences among 34 hernias in the group of 17 patients treated with 10 x 30 cm mesh and two (1 per cent) in the group that received 30 x 10/15 cm mesh (162 hernias in 81 patients). CONCLUSION: The endoscopic approach for the Stoppa procedure for bilateral inguinal hernia repair is a reliable method with minor complications. It ensures a short recuperation time and the recurrence rate is low owing to adequate overlap of the hernial defect when a 'slipmesh' is used. PMID- 10540142 TI - External anal sphincter atrophy on endoanal magnetic resonance imaging adversely affects continence after sphincteroplasty. AB - BACKGROUND: There is still considerable debate about the value of preoperative anorectal physiological parameters in predicting the clinical outcome after sphincteroplasty. Recently it has been reported that atrophy of the external anal sphincter can be clearly shown with endoanal magnetic resonance imaging (MRI). The aims of this study were to investigate the prevalence of external anal sphincter atrophy in women with anterior sphincter defects due to obstetric injury and to determine the impact of external anal sphincter atrophy on the outcome of sphincteroplasty. METHODS: In this prospective study, 20 consecutive women (median age 50 (range 28-75) years) with faecal incontinence due to obstetric trauma were assessed before operation with endoanal ultrasonography and endoanal MRI. The external anal sphincter was examined and evaluated for the presence of atrophy. The clinical outcome of sphincteroplasty was interpreted without knowledge of the magnetic resonance and ultrasonographic images. RESULTS: In all patients anterior sphincter defects could be demonstrated with ultrasonography and MRI. External anal sphincter atrophy could only be demonstrated on MRI. Eight of 20 patients had external anal sphincter atrophy. Continence was restored in 13 patients. Outcome was significantly better in those without external anal sphincter atrophy (11 of 12 patients versus two of eight; P = 0.004). CONCLUSION: External anal sphincter atrophy can only be visualized on endoanal MRI and affects continence after sphincteroplasty. Endoanal MRI is valuable in the preoperative assessment of patients with faecal incontinence. Presented to the American Society of Colon and Rectal Surgeons in Philadelphia, Pennsylvania, USA, June 1997 PMID- 10540143 TI - Frequency of early colorectal cancer in patients undergoing colonoscopy. AB - BACKGROUND: Early colorectal cancer is defined as carcinoma limited to the mucosa or submucosa. Up to 20 per cent of all colorectal cancers treated in some Japanese institutions are early cancers. These cancers are sometimes flat or depressed, and may be less than 1 cm in diameter. The aim of this study was to identify the frequency and morphology of early colonic cancers detected at colonoscopy by a surgeon aware of and looking for such lesions. METHODS: A review was made of all colonoscopies performed by or under the supervision of a single endoscopist between 1990 and 1998. Follow-up and outcome of all patients with early colorectal cancer was undertaken. RESULTS: Ninety-five invasive colorectal cancers were identified from 2198 colonoscopies. Eighteen were early colorectal cancers (T1). Macroscopically these were flat (nine tumours), villous (four) and pedunculated (five). Two patients had lymph node metastasis. The median size of flat cancers was 20 (range 9-30) mm. Median follow-up was 3 years. One patient had local recurrence, and another, whose early cancer was metachronous, died from metastatic cancer. CONCLUSION: This study identified early colonic cancer with similar frequency and morphology to that reported by the Japanese. Colonoscopy should be considered as the investigation of choice for patients with large bowel symptoms. PMID- 10540144 TI - Outcome in patients with colorectal cancer managed by surgical trainees. AB - BACKGROUND: The surgeon is an important variable that influences outcome following colorectal cancer surgery. Operative training of suitable quality and quantity is essential if intersurgeon variation is to be reduced. The aim of this study was to examine the outcome of colorectal cancer surgery when a high proportion of the operations were performed by trainee surgeons. METHODS: A prospective 7-year (1989-1996) audit of 306 consecutive colorectal cancers referred to a single general surgeon with a colorectal interest was carried out. The outcome (anastomotic leakage, 30-day mortality rate, local recurrence and cancer-related survival) of operations performed by the consultant was compared with that of his trainees. RESULTS: Some 245 (92.5 per cent) of 265 patients undergoing laparotomy had a resection. Seventy (28.6 per cent) and 67 (27.3 per cent) of operations were performed by supervised and independent trainees respectively. There was no difference between the consultant, supervised and independent trainees for 30-day mortality rate (6.5, 6 and 4 per cent respectively), clinical anastomotic leakage rate (9, 2 and 5 per cent) and local recurrence rate (2, 3 and 7 per cent). There was no difference between the three groups for adjusted 5-year disease-related survival rates. CONCLUSION: Properly supervised trainees can resect a high proportion of colorectal cancers without compromising immediate outcome or long-term survival. Presented in part to the annual meeting of the Association of Surgeons of Great Britain and Ireland, Bournemouth, UK, April 1997, and published in abstract form as Br J Surg 1997; 84(Suppl): 56 PMID- 10540145 TI - Treatment of persistent pruritus ani in a combined colorectal and dermatological clinic. AB - BACKGROUND: Pruritus ani is a common and socially embarrassing condition which is often poorly managed. It is often classified as idiopathic where the symptoms are usually transitory or secondary when a more persistent itch is experienced. The aim of this study was to establish the cause of pruritus ani in a group of patients referred to a combined colorectal and dermatological clinic, and to determine the most appropriate treatment. METHODS: Forty consecutive patients with pruritus ani were referred over a 6-month period from either the general practitioner or another hospital consultant to a combined colorectal and dermatological clinic. They were assessed by history, completion of a general health questionnaire, full examination of the skin, digital rectal examination, proctoscopy, sigmoidoscopy and patch testing. Patients were treated according to clinical findings at assessment. RESULTS: Thirty-four patients had a recognizable dermatosis, three had superficial perianal fissuring and three had a normal perineum; two required surgical intervention. Eighteen patients had a positive reaction when patch tested. All patients have shown an improvement or complete resolution of symptoms with treatment. CONCLUSION: This series has shown that the majority of patients presenting with pruritus ani have a dermatosis as the underlying cause of their symptoms and that many of them have developed contact sensitivities to the various topical medications used. These findings suggest that referral to a dermatologist in the first instance may be more appropriate. PMID- 10540147 TI - Improved perineal healing after internal sphincter-preserving proctectomy in ulcerative colitis. PMID- 10540146 TI - Prospective study of primary anastomosis without colonic lavage for patients with an obstructed left colon. AB - BACKGROUND: Traditionally, left-sided colon obstruction is managed by a multistaged defunctioning colostomy and resection. However, there is growing acceptance of one-stage primary resection and anastomosis with on-table antegrade irrigation. This paper presents a series of patients managed prospectively by primary anastomosis without intraoperative colonic lavage. METHODS: Emergency resection of acutely obstructed left-sided colonic carcinomas was performed. This was followed by primary anastomosis without on-table lavage after bowel decompression using a new technique. RESULTS: Fifty-eight consecutive, unselected patients underwent bowel decompression, resection and primary colocolic anastomosis. Only one patient developed a leak at the anastomotic site, requiring pelvic abscess drainage and transverse loop colostomy. One death occurred 12 h following surgery. Autopsy confirmed that this was due to myocardial infarction. Mean hospital stay was 9.8 days. CONCLUSION: Emergency surgery on the obstructed left colon can be carried out safely after decompression alone, without intraoperative colonic lavage. PMID- 10540148 TI - Touch imprint cytological analysis of sentinel lymph nodes for detecting axillary metastases in patients with breast cancer. AB - BACKGROUND: Sentinel lymph node biopsy is a procedure that examines the first tumour-draining lymph node. Touch imprint cytology may provide a quick method for intraoperative screening of sentinel lymph nodes for the presence of metastases. METHODS: Touch imprint cytological analysis of sentinel lymph nodes was compared prospectively with the findings obtained on routine paraffin sections. Touch imprint slides from 55 patients with breast cancer were prepared during operation from multiple sections of sentinel lymph nodes, stained with haematoxylin and eosin. A cytopathologist blinded to the histological results interpreted the smears. RESULTS: The concordance between touch imprint and paraffin sections of sentinel lymph nodes was 98 per cent (54 of 55). When touch imprint analysis of sentinel lymph nodes was compared with paraffin sectioning of all lymph nodes from the axillary node dissection, the concordance was 95 per cent (52 of 55). The sensitivity and specificity of sentinel lymph node touch imprints in detecting metastases were 82 and 100 per cent respectively. The positive and negative predictive values were 100 and 93 per cent respectively. CONCLUSION: Touch imprint cytology is potentially useful for the intraoperative evaluation of sentinel lymph nodes in patients with breast cancer. PMID- 10540149 TI - Plasma concentrations of glutathione S-transferase isoenzyme are raised in patients with intestinal ischaemia. AB - BACKGROUND: The mortality rate associated with acute mesenteric ischaemia (AMI) remains high. Diagnosis is frequently confounded by the non-specific history and physical signs, in conjunction with the absence of a reliable biological assay. Glutathione S-transferase (GST) is an enzyme with a crucial role in cellular homoeostasis, the alpha isoenzyme of which is highly specific to small bowel and liver. This study assessed alphaGST as a marker for AMI. METHODS: Twenty-six patients with acute abdominal pain were enrolled. Each patient manifested a diagnostic dilemma, with a potential diagnosis of AMI. Plasma was reserved for alphaGST assay during routine blood testing and stored at -20 degrees C for analysis. A final diagnosis was made by autopsy, laparotomy, a definitive other investigation or a return to full health. RESULTS: Twelve patients had AMI. Plasma alphaGST was significantly increased in patients with AMI (P < 0.0001). Although pH differed and other biochemical changes occurred, only alphaGST accurately predicted AMI. CONCLUSION: A threshold of 4 ng/ml for alphaGST was 100 per cent sensitive and 86 per cent specific for AMI. If these observations can be confirmed, evaluation of alphaGST may reliably predict the presence or absence of AMI. PMID- 10540150 TI - Treatment of gallbladder cancer by radical resection. PMID- 10540152 TI - The modulation of B7.2 and B7.1 on B cells by immunosuppressive agents. AB - Several recent studies demonstrate that B7.2, but not B7.1, play an important role in allergic inflammation and IgE production. Agents that down-regulate B7.2 may therefore be of benefit for the treatment of Th2-driven allergic diseases. Our current study was carried out to investigate the effect of immunosuppressive agents, cyclosporin A (CsA) and dexamethasone, on B7.2 and B7.1 expression on B cells stimulated with the superantigen, toxic shock syndrome toxin-1 (TSST-1). The analysis of B7.2 and B7.1 on the same cells by flow cytometry demonstrated that TSST-1 up-regulated B7.2+B7.1- but not B7.1+B7.2- on B cells in a dose dependent fashion. CsA and dexamethasone significantly down-regulated B7.2+B7.1- but up-regulated B7.2-B7.1+ B cells in the presence or absence of TSST-1 (100 ng/ml). Interestingly, the combination of CsA and dexamethasone was much more potent in the inhibition of B7.2 expression than either of these agents alone. As CD40 is known to up-regulate B7.2 expression on B cells, the mechanism of B7.2 down-regulation by CsA and dexamethasone was further studied by investigating the effect of these agents on CD40 expression on B cells. TSST-1 significantly increased CD40 expression on B cells. However, the addition of CsA or dexamethasone significantly down-regulated CD40 expression. Anti-CD40 MoAb significantly reversed the effects of CsA or dexamethasone on B7.2 and B7.1 expression, suggesting that T cell engagement of CD40 plays a role in the mechanisms by which CsA and dexamethasone acts on B cells. These data demonstrate the modulatory effect of CsA and dexamethasone on B7.2 and B7.1 expression on B cells and the potential role of CD40 in mediating this effect. PMID- 10540153 TI - Effect of different sensitizing doses of antigen in a murine model of atopic asthma. AB - The dose of antigen is assumed to be one of the important factors in the polarized development of helper T cell subsets, i.e. Th1 or Th2 cells. We investigated the effect of the sensitizing antigen dose in a murine model of atopic asthma, which involved sensitization with ovalbumin (OVA) followed by repeated exposure to OVA aerosols. BALB/c mice were primed with varying doses of OVA (0, 10, 100 and 1000 microg) plus Al(OH)3 on days 0, 7 and 14, and were challenged with OVA aerosols (50 mg/ml for 20 min) on days 15-20. There were striking antigen dose-related differences in OVA-specific antibodies: high IgE and low IgG2a titres were found in mice sensitized at 10 microg, while low IgE and high IgG2a titres were seen at 1000 microg. The sensitizing dose was inversely correlated with the total cell count and the eosinophil count in bronchoalveolar lavage fluid (BALF), as well as with the extent of histological changes such as goblet cell hyperplasia of the bronchial epithelium and cellular infiltration into bronchovascular bundles. Antigen-induced bronchial hyper responsiveness (BHR) to methacholine was observed with sensitization at 10 microg but not at 1000 microg. Splenic mononuclear cells (SMNC) obtained from mice sensitized at either dose showed proliferation in response to OVA. Production of IL-4 and IL-5 by OVA-stimulated SMNC was inversely correlated with the dose of sensitizing antigen. High-dose sensitization resulted in general suppression of cytokine production by SMNC, including interferon-gamma (IFN-gamma). The BALF levels of IL-4 and IL-5 were increased by low-dose sensitization, whereas IFN gamma and IL-12 levels were increased by high-dose sensitization. These results suggest that the dose of sensitizing antigen defines the phenotypic changes in the present murine asthma model, presumably by influencing the pattern of cytokine production. PMID- 10540154 TI - Sodium butyrate blocks interferon-gamma (IFN-gamma)-induced biosynthesis of MHC class III gene products (complement C4 and factor B) in human fetal intestinal epithelial cells. AB - Human intestinal epithelial cells have been established as local sites for complement biosynthesis. In this study, we investigated the effects of IFN-gamma and sodium butyrate on biosynthesis of MHC class III gene products (complement C4 and factor B) in the human fetal intestinal epithelial cell line INT-407. IFN gamma induced a dose- and time-dependent increase in C4 and factor B secretion. However, sodium butyrate dose-dependently inhibited IFN-gamma-induced C4 and factor B secretion. These effects were also observed at the mRNA level. Immunoblotting indicated that IFN-gamma induced a rapid activation of Stat1alpha, and fluorescence immunohistochemistry detected a translocation of Stat1alpha into the nucleus within 1 h. However, the translocation of Stat1alpha was not affected by the addition of sodium butyrate. Nuclear run-on assay indicated that IFN-gamma induced a weak increase in the transcription rate of factor B gene, and sodium butyrate did not affect this response. IFN-gamma and sodium butyrate induced a counter-regulatory effect on C4 and factor B secretion: IFN-gamma acted as a potent inducer, but sodium butyrate potently abrogated these responses. These are mainly regulated through the post-transcriptional mechanism. PMID- 10540155 TI - Counter-regulatory effect of sodium butyrate on tumour necrosis factor-alpha (TNF alpha)-induced complement C3 and factor B biosynthesis in human intestinal epithelial cells. AB - The various biological activities of butyrate have been well documented. In this study, we tested the effects of butyrate on TNF-alpha-induced complement C3 and factor B biosynthesis in human intestinal epithelial cells. The biosynthesis of C3, factor B and IL-8 was evaluated at the protein and mRNA levels. To evaluate transcriptional activation, the nuclear run-on assay was performed. The transcription factor-DNA binding activity was assessed by an electrophoretic gel mobility shift assay (EMSA). In the intestinal epithelial cell lines HT-29, T84 and Caco-2, sodium butyrate enhanced TNF-alpha-induced C3 secretion, but suppressed TNF-alpha-induced factor B and IL-8 secretion. Nuclear run-on assay revealed that transcriptional regulatory mechanisms are involved in the effects of sodium butyrate. The EMSAs indicated that sodium butyrate suppressed TNF-alpha induced nuclear factor (NF)-kappaB- and activation protein (AP)-1-DNA binding activity, but enhanced TNF-alpha-induced activation of CCAAT/enhancer-binding protein (C/EBP)beta (NF-IL-6)-DNA binding activity. Sodium butyrate induced a counter-regulatory effect on TNF-alpha-induced C3 and factor B biosynthesis in human intestinal epithelial cells. Butyrate action has been discussed with its activity to induce histone hyperacetylation, but its counter-regulatory effect on complement biosynthesis may be closely associated with the modulation of transcription factor activation. PMID- 10540157 TI - Analysis of the secretion pattern of monocyte chemotactic protein-1 (MCP-1) and transforming growth factor-beta 2 (TGF-beta2) by human retinal pigment epithelial cells. AB - Retinal pigment epithelial (RPE) cells, situated between the neurosensory retina and the vascularized choroid, form part of the blood-eye barrier and are important for homeostasis of the outer retina. These cells are able to produce a variety of cytokines which may play a role in the maintenance of the immunosuppressive milieu inside the eye and in intraocular inflammatory responses. In the present study, we investigated whether RPE cells secreted the anti-inflammatory cytokine TGF-beta2 and the proinflammatory cytokine MCP-1 in a polarized manner. Monolayers of human donor RPE cells were cultured on transwell filters. Secretion of TGF-beta2 and MCP-1 at either the apical or basal side of the RPE cell monolayers, that were not treated or stimulated with IL-1beta (200 U/ml), was analysed by ELISA. All three cell lines examined had a different TGF beta2 secretion pattern. In two of the three donor RPE cell lines tested, TGF beta2 secretion was polarized, but not in the same direction. TGF-beta2 secretion was not up-regulated by stimulation with IL-1beta. In contrast, IL-1beta strongly induced MCP-1 secretion preferentially into the basal compartment of all RPE monolayers tested. These data indicate that human RPE cells are able to secrete TGF-beta2 and MCP-1 in a polarized fashion. Our results suggest that MCP-1 can be secreted by RPE cells in the direction of choroidal vessels during inflammatory responses in the posterior part of the eye, which may limit damage to the neurosensory retina. PMID- 10540156 TI - Qualitative and quantitative studies of autoantibodies to phospholipids in diabetes mellitus. AB - Diabetes mellitus is associated with vascular and neurological complications. We have investigated the presence of antibodies to phospholipids and to phospholipid binding plasma proteins in blood samples collected from 68 clinically and biochemically characterized type I and type II diabetic patients and from 252 healthy blood donor controls. Each sample was analysed for antibodies to three phospholipids (cardiolipin, phosphatidylserine and phosphatidylethanolamine), the antibody isotypes (IgA, IgG and IgM), and whether antibody activity was plasma protein-dependent. Patients were considered to have anti-phospholipid antibodies when one or more of these 18 tests was found above predetermined control values. The results of these experiments revealed an increased incidence of anti phospholipid antibodies in diabetic patients compared with control subjects. The incidence of IgA isotype to phosphatidylethanolamine was higher than the incidence of other isotypes to other phospholipids, and their reactivities were independent of phospholipid-associated proteins. In addition, these antibody findings were studied for associations with prothrombin degradation products, activated factor VII and activated protein C, and with the incidence of diabetic complications. The anti-phosphatidylethanolamine antibody association with proliferative retinopathy was significant. PMID- 10540158 TI - Suppressive effect of Chinese medicinal herb, Acanthopanax gracilistylus, extract on human lymphocytes in vitro. AB - We studied the effect of a Chinese medicinal herb, Acanthopanax gracilistylus, extract (AGE), on human lymphocytes in vitro. AGE markedly suppressed the proliferative responses of human peripheral blood lymphocytes stimulated with mitogens concanavalin A (Con A) and Staphylococcus aureus Cowan I (SAC). Both T cell and B cell activities-production of interferon-gamma and immunoglobulin-were suppressed by AGE. The mechanism of AGE-induced suppression of lymphocytes is to arrest the cell cycle at the G0/G1 stage without a direct cytotoxic effect. AGE also suppressed the alloantigen-specific cytotoxic T lymphocyte response. However, natural killer cell activity was less sensitive to the suppressive activity of AGE. In contrast, AGE markedly enhanced monocyte function to produce cytokines. These activities of AGE were associated with a 60-kD protein which was sensitive to treatment with pronase E, but not with NaIO4. These results suggest that AGE has an immunomodulating activity on human lymphocytes and its properties could be clinically applied in the treatment of several diseases such as autoimmune and allergic diseases. PMID- 10540159 TI - Turkeys are protected from infection with Chlamydia psittaci by plasmid DNA vaccination against the major outer membrane protein. AB - Plasmid DNA expressing the major outer membrane protein (MOMP) of an avian Chlamydia psittaci serovar A strain has been tested for its ability to raise an immune response and induce protection against challenge with the same serovar. A combined parenteral (intramuscular injection) and mucosal route (DNA drops administered to the nares) of DNA inoculation was compared with gene gun-based immunization. The gene gun delivery of pcDNA1/MOMP as well as the intramuscular intranasal DNA delivery primed both T-helper and B cell memory, although rMOMP expressing cells did not induce high antibody responses. Evidence for the priming of the memory was provided by the fact that the pcDNA1/MOMP inoculations raised antibodies belonging to the IgG and not IgM isotype. However, in response to challenge only five out of 15 vaccinated turkeys showed four-fold increases in serum IgG after challenge. By contrast, evidence for the priming of T cell memory in response to challenge was found in all vaccinated turkeys, as shown by the significantly heightened proliferative responses of peripheral blood lymphocytes following vaccination. Both immunization methods produced similar serological and lymphocyte proliferative responses. Notwithstanding the immunization method, a significant level of protection was observed in all pcDNA1/MOMP-immunized turkeys. The efficacy of MOMP-based DNA vaccination as a means of preventing severe clinical signs, lesions and chlamydia excretion in a turkey model of C. psittaci infection was demonstrated. PMID- 10540160 TI - HLA-DRB1 leprogenic motifs in nigerian population groups. AB - Amino acid residues involved in the peptide binding groove of HLA-DRB1 alleles were examined in three Nigerian ethnic groups with leprosy (n = 287) and 170 controls to determine the role of DRB1 alleles in disease outcome with Mycobacterium leprae. Nine positively charged motifs and two others with neutral charge to the binding groove were detected. These motifs occurred more frequently in leprosy (leprogenic) than was expected by chance (P < 0.0001). In contrast, five motifs with net negative or 'modified' neutral charges to the pocket were negatively associated with leprosy. We conclude that clinical outcome of infection with M. leprae is largely determined by a shared epitope in DRB1 alleles marked by several motifs. These motifs occur in otherwise normal DRB1 alleles, characterized by net positive or neutral charges in the binding groove. We hypothesize that these polarities cause poor binding of DRB1 to M. leprae. On presentation, the signal via the T cell receptor results in muted cell-mediated immunity. The resulting response translates to various forms of leprosy depending on degree of charge consonance between M. leprae and host DRB1 allele. Other factors within or without the HLA complex, such as the T cell receptor repertoire, may also influence the resulting disease. PMID- 10540161 TI - Perforin and Fas/Fas ligand-mediated cytotoxicity in acute and chronic woodchuck viral hepatitis. AB - The Fas ligand (FasL)/Fas and the perforin-granzyme cytotoxic pathways presumably play a central role in the development of hepatocellular injury in viral hepatitis. To recognize the potential contribution of FasL and perforin-based cell killing in hepadnaviral infection, we adopted a cytotoxic assay using murine Fas+ P815 and human Fas- K562 cells as targets. Freshly isolated peripheral blood mononuclear cells (PBMC) from woodchucks with newly acquired woodchuck hepatitis virus (WHV) infection (n = 6), with chronic WHV hepatitis (n = 9), and from healthy animals (n = 11) were used as effector cells. We have found that woodchuck lymphoid cells kill cell targets via both the FasL/Fas and the perforin death pathways. The contribution of Fas-dependent cytolysis was ascertained in blocking experiments with anti-Fas antibody and by incubation of PBMC with cyclohexamide to prevent de novo synthesis of FasL. The involvement of the perforin pathway was confirmed by treatment of K562 cells with colchicine to inhibit the microtubule-dependent perforin release. Comparative analysis showed that peripheral lymphoid cells from acute WHV hepatitis, but not those from chronic WHV infection, are more cytotoxic and that this increase seems to be entirely due to activation of perforin-mediated killing. The data indicate that acute infection in woodchucks is associated with the augmented capacity of lymphoid cells to elicit perforin-dependent killing, but in chronic infection, independent of the severity of liver disease and duration of chronicity, these cells have the same or lower cytotoxic potential as PBMC from healthy controls. These findings suggest a role for non-specific cellular immunity, presumably natural killer (NK) cells, in the control of early WHV infection and in the progression of chronic hepatitis. PMID- 10540162 TI - Interferon-alpha (IFN-alpha) enhances cytotoxicity in healthy volunteers and chronic hepatitis C infection mainly by the perforin pathway. AB - Cell-mediated cytotoxicity is exerted via perforin and Fas ligand (FasL). We have recently shown that IFN-alpha up-regulates FasL expression in T cells isolated from healthy volunteers and augments activation-induced T cell death. Since the Fas/FasL system is implicated in the pathogenesis of hepatic failure and both molecules have been shown to be up-regulated in hepatitis C virus (HCV) infection, we studied whether cytotoxicity via the FasL system is enhanced by IFN alpha and therefore could contribute to hepatic injury. We investigated FasL and perforin expression in peripheral blood mononuclear cells (PBMC) derived from HCV+ donors by Northern analysis and soluble FasL synthesis by ELISA. Natural killer (NK) cell and cytotoxic T lymphocyte (CTL) cytotoxicity was studied by 51Cr-release assays. IFN-alpha up-regulates FasL mRNA and protein synthesis in mitogen-activated PBMC of HCV+ individuals, as previously found in healthy subjects. Stimulation with IFN-alpha increases perforin mRNA levels in PBMC. In NK cytotoxicity assays, the enhancement of cytotoxicity by IFN-alpha is mainly due to the perforin pathway, while the FasL pathway plays only a minor role. In CTL cytotoxicity experiments neither the FasL nor the perforin pathway is further enhanced by IFN-alpha. Our data suggest that up-regulation of perforin by IFN alpha results in elevated cytotoxicity, suggesting that IFN-alpha might support elimination of virally infected cells via this pathway. In contrast, the major effect of IFN-alpha on the Fas/FasL system might be the enhanced elimination of activated T cells as a means of finally limiting a T cell response. PMID- 10540163 TI - Induction of HIV-1-specific T cell responses by administration of cytokines in late-stage patients receiving highly active anti-retroviral therapy. AB - Highly active anti-retroviral therapy (HAART) is associated with reduction in the morbidity and mortality of patients with advanced HIV-1 disease. The ability of such treatment to improve immune responses against HIV-1 and opportunistic pathogens is variable and limited. Addition of cytokine immunotherapy to this treatment may improve immune responses. IL-2 with or without granulocyte macrophage colony-stimulating factor (GM-CSF) was administered to HIV-1+ individuals receiving HAART with undetectable viral loads, and CD4 counts < 100 cells/microl. In one patient presenting with Mycobacterium avium complex (MAC) infection, we evaluated the effect of cytokine immunotherapy on lymphocyte phenotype; plasma viral load; proliferative responses to mitogens, recall and HIV 1 antigens; cytokine production and message in response to non-specific and specific stimuli; and natural killer (NK) cell activity. Proliferation assays were performed in two similar patients. Before cytokine immunotherapy the predominant CD8+ population was mainly CD28-. No proliferation or IL-2 production was seen in response to mitogens, recall or HIV-1 antigens; and no HIV-1 peptide specific interferon-gamma (IFN-gamma)-secreting cells were present. Low levels of IL-4 were detected in response to antigens to which patients had been exposed, associated with up-regulated expression of costimulatory molecules influenced by IL-4. Following IL-2 administration, loss of IL-4 was associated with increased NK cell activity and HIV-1 peptide-specific and non-specific IFN-gamma-producing cells. Proliferative responses associated with IL-2 production and responsiveness were only seen after subsequent concomitant administration of GM-CSF with IL-2. These changes mirrored clinical improvement. An imbalance of lymphocyte subsets may account for immune unresponsiveness when receiving HAART. Restoration of responses following immunotherapy suggests a shift towards a lymphocyte profile with anti-pathogen activity. PMID- 10540164 TI - CCR5 and CXCR4 chemokine receptor expression and beta-chemokine production during early T cell repopulation induced by highly active anti-retroviral therapy. AB - Expression of chemokine receptors and beta-chemokine production by peripheral blood mononuclear cells (PBMC) were determined in HIV-1-infected individuals before and after highly active anti-retroviral therapy (HAART) and their relationship to viral load, T cell phenotype and the expression of immunological activation markers was examined. We found that the expression of CCR5 is up regulated in HIV-1-infected individuals while CXCR4 appears down-regulated on both CD4 and CD8 T cells compared with normal controls. These alterations are associated with the high levels of viral load. In addition, a relationship was observed between the degree of immune activation and chemokine receptor expression on T cells. However, after 3 months of combined anti-retroviral regimen, expression of CXCR4 significantly increased while CCR5 decreased when compared with pretherapy determinations. This was seen in strict association with a dramatic decrease of viral load and an increase of both CD45RA+/CD62L+ (naive) and CD45RA-/CD62L+ or CD45RA+/CD62L- (memory) T cells accompanied by a significant decrease of the expression of immune activation markers such as HLA DR and CD38. At enrolment, both spontaneous and lectin-induced RANTES, macrophage inflammatory protein-1alpha (MIP-1alpha) and MIP-1beta production by PBMC were higher in HIV-1-infected individuals compared with normal controls, although differences for MIP-1beta were not statistically significant. However, RANTES and MIP-1alpha production decreased during HAART at levels closer to that determined with normal controls, while MIP-1beta production was less consistently modified. These data indicate that the expression of chemokine receptors CCR5 and CXCR4 and the production of beta-chemokines are altered in HIV-infected individuals, and suggest that their early modifications during HAART reflect both the peripheral redistribution of naive/memory T cell compartments and the decrease in levels of T cell activation. Such modifications in the expression of host determinants of viral tropism and the production of anti-viral molecules may play a role in the emergence of virus variants when a failure of HAART occurs. PMID- 10540165 TI - Cytokine gene expression in peripheral blood mononuclear cells (PBMC) from patients with pharmacologically treated cystic echinococcosis. AB - The influence of pharmacological treatment on the immune response of patients with Echinococcus granulosus infection was evaluated by reverse transcriptase polymerase chain reaction (RT-PCR) to determine mRNA expression for IL-12 p35, IL 12 p40, interferon-gamma (IFN-gamma), tumour necrosis factor-alpha (TNF-alpha) and IL-4 in PBMC from 12 patients before and after chemotherapy and from seven uninfected controls. Most patients' PBMC showed measurable amounts of IL-12 p35, IL-4, IFN-gamma and TNF-alpha mRNA in parasite antigen-stimulated and unstimulated cultures. Conversely, IL-12 p40 mRNA was detected almost exclusively in successfully treated patients (86%) after therapy. In these patients semiquantitative analysis of RT-PCR products showed a significant difference between IL-12 p40 mRNA mean levels before and after therapy (P = 0.03 in parasite antigen-stimulated cultures; P = 0.001 in unstimulated cultures). IL-4 mRNA was weakly expressed before therapy and more highly so after treatment in both groups of patients and under both culture conditions; IL-4 mRNA reached its highest level in post-therapy PBMC from patients in whom therapy failed (stimulated cultures). IFN-gamma and TNF-alpha mRNA expression increased in patients who responded to therapy and decreased in patients who did not. In contrast to IL-12 p35, IFN-gamma and TNF-alpha mRNAs, IL-12 p40 and IL-4 mRNAs were detected exclusively in patients, suggesting a close relationship between these two cytokines and cystic echinococcosis. Our findings indicate that chemotherapy influences the immune response, thus determining changes in Th1/Th2 cytokine mRNA patterns, predominantly in IL-12 p40 and IL-4 mRNA expression. PMID- 10540166 TI - Clinical and immunological evaluation of patients with mild IgG1 deficiency. AB - Serum IgG subclass concentrations were determined in patients visiting, the pulmonology out-patient clinic with chronic respiratory tract problems. A total of 24 patients with a serum IgG1 concentration < 4.9 g/l (i.e. below the reference range) and normal values for IgG2, IgM and IgA were included. Patients with a selective IgG1 deficiency were vaccinated with a 23-valent pneumococcal polysaccharide vaccine. There were nine patients with a poor antibody response to pneumococcal capsular polysaccharide antigens. Responsiveness to protein antigens was intact in all patients. Patients with pneumonia showed a significantly lower anti-polysaccharide response in the IgG2 subclass than patients without pneumonia. Patients with recurrent sinusitis showed a significantly lower response in the IgA isotype after vaccination with pneumococcal polysaccharide vaccine compared with non-sinusitis patients. It can be concluded that patients with recurrent sinopulmonary infections and a mild IgG1 subclass deficiency have an impaired IgG1 anti-polysaccharide response, which can extend to decreased IgG2 and IgA anti-polysaccharide responses. PMID- 10540167 TI - Cytolytic mechanisms involved in non-MHC-restricted cytotoxicity in Chediak Higashi syndrome. AB - To determine the mechanisms responsible for the impaired lymphocyte-mediated cytotoxicity in Chediak-Higashi syndrome (CHS), we investigated the killing ability of peripheral blood lymphocytes (PBL) from three patients with CHS using several kinds of target cells that were sensitive to perforin, Fas ligand (FasL), and/or tumour necrosis factor-alpha (TNF-alpha). Freshly isolated CHS PBL did not kill K562 target cells, killing of which by normal PBL was perforin-dependent, as demonstrated by complete inhibition by concanamycin A (CMA), an inhibitor of perforin-based cytotoxicity. In contrast, the CHS PBL exhibited substantial cytotoxicity against Jurkat cells, which was only partially inhibited by CMA treatment but not by the addition of neutralizing anti-FasL or anti-TNF-alpha antibodies. IL-2-activated CHS PBL exhibited substantial levels of cytotoxicity against K562 and Jurkat cells, the levels being 74% and 83% of the respective normal control values, respectively. CMA treatment showed that while the cytotoxicity of IL-2-activated CHS PBL against K562 was largely dependent on perforin, that against Jurkat was largely not. IL-2-activated CHS PBL expressed FasL mRNA, and killed Fas transfectants. These findings indicate that CHS PBL have an ability to kill some target cells via a perforin-mediated pathway, especially when they are activated by IL-2. It was also demonstrated that CHS PBL can exert cytotoxicity against certain target cells by utilizing FasL and an undefined effector molecule other than perforin, FasL, or TNF-alpha. PMID- 10540168 TI - Dendritic cell-derived nitric oxide is involved in IL-4-induced suppression of experimental allergic encephalomyelitis (EAE) in Lewis rats. AB - Cytokines play a crucial role in initiating and perpetuating EAE, an animal model of multiple sclerosis (MS). A low dose of IL-4, administered by the nasal route over 5 days (100 ng/rat per day) prior to immunization, improved clinical scores of EAE induced in Lewis rats with myelin basic protein (MBP) peptide 68-86 (MBP 68-86). We examined whether dendritic cells (DC) may have contributed to the amelioration of the disease process. These professional antigen-presenting cells (APC) not only activate T cells, but also tolerize T cells to antigens, thereby minimizing autoimmune reactions. We found that IL-4 administration enhanced proliferation of DC. In comparison with DC of PBS-treated rats, DC from IL-4 treated rats secreted high levels of interferon-gamma (IFN-gamma) and IL-10. Nitric oxide (NO) production by DC was also strongly augmented in IL-4-treated rats. In vitro studies showed that IL-4 stimulated DC expansion and that IFN gamma enhanced NO production by DC. DC-derived NO promoted apoptosis of autoreactive T cells. These results indicate that nasal administration of IL-4 promotes activation of DC and induces production of IFN-gamma and IL-10 by DC. IL 10 suppresses antigen presentation by DC, while IFN-gamma induces NO production by DC which leads to apoptosis in autoreactive T cells. Such a DC-derived negative feedback loop might contribute to the clinical improvement observed in EAE. PMID- 10540169 TI - IgM anti-myeloperoxidase antibody-secreting lymphocytes are present in the peripheral repertoire of lupus mice but rarely differentiate into IgG-producing cells. AB - Two IgM, kappa anti-myeloperoxidase (MPO) monoclonal antibodies, 6D6 and 9B5, bound to MPO in a solid-phase enzyme-linked immunosorbent assay were derived from the splenocytes of (NZB x NZW) F1 and MRL/lpr-lpr mice, respectively. 6D6 gave a characteristic perinuclear immunofluorescence staining pattern on ethanol-fixed human neutrophils, bound to the native form of MPO by immunoblotting and had a high constant affinity for MPO as demonstrated by real-time specific interaction. 9B5 produced a cytoplasmic immunofluorescence staining pattern, reacted with the heavy chain of MPO and had a low constant affinity for MPO. The heavy-and light chain variable region genes of monoclonal antibodies (mAb) 6D6 and 9B5 were sequenced and found to be highly homologous to germline genes and to contain negatively charged amino acids in the complementarity determining regions. IgM MPO-binding activity was observed in most BW and MRL/lpr-lpr mouse sera, which may correspond to polyclonal activation of B cells, whereas IgG anti-MPO antibodies could be rarely detected. Thus, this study indicates that (i) BW and MRL/lpr-lpr mice do not delete IgM anti-MPO secreting B cells, do not maintain these B cells in a state of anergy, but most individuals are not able to spontaneously induce the class-switching of this autoantibody population; (ii) IgM anti-MPO antibodies can recognize different epitopes on MPO and produce different immunofluorescence staining pattern on ethanol-fixed human neutrophils, as demonstrated by the immunochemical properties of the two lupus-mouse derived mAb. PMID- 10540170 TI - Natural killer (NK) T cells are significantly decreased in the peripheral blood of patients with rheumatoid arthritis (RA). AB - The number of NK T cells was measured in relation to the Th1/Th2 imbalance observed in RA. Peripheral blood samples of patients with RA (n = 60) and healthy controls (n = 36) were stained with anti-NK receptor 1A (anti-NKR-P1A), anti CD56, and anti-CD3 MoAbs, and examined by three-colour flow cytometry. NK T (NKR P1A+CD3+) cells in the peripheral blood were decreased in RA compared with the controls: 25 +/- 20/microl versus 143 +/- 53/microl (P < 0.0001). CD56+CD3+ cells were also decreased in RA: 60 +/- 46/microl versus 116 +/- 54/microl (P < 0.0001). The decrease was significant when adjusted to the number of total lymphocytes (P < 0.0001) or NK (CD56+CD3-) cells (P < 0.0001), and showed no correlation with age, sex, disease duration, disease activity, functional class, x-ray stage, drug treatment, joint score, grip strength, C-reactive protein, rheumatoid factor or erythrocyte sedimentation rate of the patients. The results show that the levels of NK T cells are depressed in the peripheral blood of patients with RA, suggesting that the measurement of NK T cells in peripheral blood may have clinical importance for a Th1-type autoimmune disease like RA. PMID- 10540171 TI - Tumour necrosis factor (TNF) production by T cell receptor-primed T lymphocytes is a target for low dose methotrexate in rheumatoid arthritis. AB - Methotrexate (MTX) is an effective immunosuppressive agent in various chronic inflammatory diseases such as rheumatoid arthritis (RA). However, its mechanisms of action are only partially understood. In this study, we assessed the effects of MTX on the differentiation of peripheral blood (PB) CD4+CD45RA 'naive' and CD4+CD45RO 'memory' T cells from healthy controls and patients with RA. Accordingly, purified T cells were primed and restimulated in vitro via the T cell receptor (TCR) in the presence of IL-2 to generate effector T cells secreting large amounts of Th1 and Th2 cytokines. We observed that low doses of MTX strongly suppress TNF and to a lesser extent interferon-gamma (IFN-gamma) production by T cells from both healthy donors and RA patients when present during T cell priming via the TCR. Similar data were obtained for TCR-primed synovial fluid mononuclear cells in RA. In contrast, production of IL-4 by TCR primed CD45RA T cells was significantly increased upon MTX treatment. Interestingly, MTX did not enhance IL-4 production when present during restimulation of effector CD45RO T cells, although it still suppressed TNF production. The results indicate that MTX effects depend on the stage of T cell activation and identify TNF production by TCR-primed T lymphocytes as a target for low-dose MTX treatment in RA. These findings could explain the delayed clinical effects of MTX and may contribute to its potent anti-inflammatory and immunoregulatory properties. PMID- 10540172 TI - Antibody production in autoimmune BXSB mice. I. CD40L-expressing B cells need fewer signals for polyclonal antibody synthesis. AB - Male, but not female, BXSB mice develop severe lupus associated with multiple immune system defects. It was recently shown that one immunological abnormality found in male BXSB mice encompasses B cell expression of CD40 ligand (CD40L) by an expanded population of large B cells. The present study was undertaken to determine how the CD40L-expressing large B cells in male BXSB mice compared with size-matched B cells from female mice in terms of their ability to secrete antibody. It was shown that the large B cells from female mice, similar to the small B cells from either male or female mice, required CD40 signalling, immunoglobulin cross-linking and cytokines for optimal antibody synthesis. In contrast, large B cells from male BXSB mice produced high levels of antibody when stimulated with only two of the three signals, and made significantly more total IgM and IgG, and anti-ssDNA antibody than size-matched B cells from female mice when stimulated with IL-4/IL-5 alone, IL-4/IL-5 plus low levels of anti-IgD dextran, or IL-4/IL-5 plus anti-CD40 MoAb. The ability of the large B cells from male mice to produce antibody under suboptimal stimulatory conditions correlated with their expression of CD40L, and was inhibited by CD40-immunoglobulin. Taken together, these findings suggested that large CD40L-expressing B cells from male BXSB mice may be able to bypass a need for CD40 signalling from T cells, thus contributing to autoimmune disease by promoting antibody production in the absence of cognate T cell help. PMID- 10540173 TI - CD4 cytotoxic and dendritic cells in the immunopathologic lesion of Sjogren's syndrome. AB - The existence of CD4+ T lymphocytes with cytotoxic activity in minor salivary gland (MSG) biopsies from Sjogren's syndrome (SS) patients was investigated using in situ double immunohistochemistry technique. The presence of dendritic cells (DC) in SS lesions was examined by using single and double immunohistochemistry methods and a panel of different MoAbs to specific cell surface markers (i.e. CD3, CD11c, DRC). Furthermore, the ultrastructural morphology of DC was characterized by electron microscopy (EM). Immunogold labelling technique using the DRC surface marker was also applied. Finally, we investigated the existence of germinal centres (GC) in the salivary gland lesions of SS patients. Seven patients with primary SS and five patients with non-specific sialadenitis were the subjects of this study. Our results indicate the existence of a CD4+ cytotoxic cell population that utilizes perforin-mediated cell destructions as they expressed perforin mRNA. Quantitative analysis of these cells revealed that they comprised approximately 20% of the existing T lymphocytes. We also identified a population of CD4+ T cells that expressed the CD11c activation marker. Furthermore, we observed a distinct cell subtype which expressed the DRC cell surface marker. These cells had the characteristic ultrastructural morphology of DC and were DRC+ when examined by immunoelectron microscopy. Finally, the formation of GC structures in the histopathologic lesions of the salivary glands was observed. The above findings indicate that both CD4+ cytotoxic T lymphocytes (CTL) and DC may be involved in the initiation and perpetuation of SS pathogenesis. Moreover, the formation of GC in the lesions reveals a possible mechanism for in situ differentiation and proliferation of activated B lymphocytes. PMID- 10540174 TI - Keratinocytes from patients with lupus erythematosus show enhanced cytotoxicity to ultraviolet radiation and to antibody-mediated cytotoxicity. AB - Keratinocyte cytotoxicity is an important component of the immunopathology of photosensitive lupus erythematosus, and antibody-dependent cell-mediated cytotoxicity (ADCC) has been shown to be an important mechanism by which autoantibodies, especially those specific for SS-A/Ro, can induce keratinocyte damage in models of photosensitive lupus. We provide further evidence that keratinocytes from patients with photosensitive lupus show significantly greater ultraviolet radiation (UVR)-induced cytotoxicity, and that ADCC of these targets is especially enhanced by autologous patient's serum or by anti-SS-A/Ro+ sera. Keratinocytes from normal uninvolved skin of 29 patients with cutaneous lupus erythematosus (LE) were grown in cell culture and tested as targets in cytotoxicity experiments in vitro. Cultured keratinocytes from patients with systemic lupus erythematosus (SLE) and subacute cutaneous lupus erythematosus (SCLE) showed significantly greater cytotoxicity following UVR treatment than did keratinocytes from normal adult controls or from neonatal foreskins (P < 0.01). The same cultures also showed greater UVR-induced binding of IgG from fractionated anti-SS-A/Ro+ preparations. ADCC experiments were also performed using keratinocytes cultured from patients with SLE, SCLE, discoid lupus erythematosus (DLE), and normal controls. When keratinocytes were incubated in autologous serum plus a standard mononuclear cell effector population, the percentage of ADCC observed was significantly greater in cultures containing keratinocytes and sera from the SLE and SCLE patients (P < 0.001). When cultured keratinocytes were added to different IgG antibody probes, plus standard mononuclear effector populations, greater ADCC was seen using the anti-SS-A/Ro probe and keratinocytes from patients with SLE or SCLE. With normal human neonatal keratinocyte targets, the anti-SS-A/Ro probe induced greater ADCC than that seen with anti-ssDNA or normal human serum. We have shown that keratinocytes from patients with some forms of lupus erythematosus (SLE and SCLE) show greater cytotoxicity in vitro when irradiated with UVR, and greater susceptibility to ADCC whether the antibody source is their own serum or an anti-SS-A/Ro probe. PMID- 10540175 TI - The activation of the neutrophil respiratory burst by anti-neutrophil cytoplasm autoantibody (ANCA) from patients with systemic vasculitis requires tyrosine kinases and protein kinase C activation. AB - The ability of antineutrophil cytoplasm autoantibodies (ANCA) from patients with systemic vasculitis to stimulate protein kinase C (PKC) and tyrosine kinases was examined in human neutrophils. Using the superoxide dismutase-inhibitable reduction of ferricytochrome C, the kinetics of ANCA-induced superoxide (O2-) production were characterized and subsequently manipulated by specific inhibitors of PKC and tyrosine kinases. With this approach, ANCA IgG, but not normal IgG or ANCA F(ab')2 fragments caused a time and dose dependent release of O2- from TNF alpha primed neutrophils. The kinetics of ANCA-induced O2- production showed an initial 10-15 min lag phase compared to the N-formyl-L-methionyl-L-leucyl-L phenylalanine response, suggesting differences in the signalling pathways recruited by these two stimuli. Inhibitor studies revealed that ANCA-activation involved members of both the Ca2+-dependent and -independent PKC isoforms and also tyrosine kinases. ANCA IgG resulted in the translocation of the betaII isoform of PKC at a time corresponding to the end of the lag phase of O2- production, suggesting that PKC activity may be instrumental in processes regulating the activity of the NADPH oxidase in response to ANCA. Tyrosine phosphorylation of numerous proteins also peaked 10-15 min after stimulation with ANCA but not normal IgG. These data suggest that PKC and tyrosine kinases regulate O2- production from neutrophils stimulated with autoantibodies from patients with systemic vasculitis. PMID- 10540176 TI - The fas and fas ligand pathways in liver allograft tolerance. AB - The Fas and Fas ligand (Fas/FasL) pathways may play a central role in cytotoxicity or immunoregulation in liver transplantation. Here, in an attempt to examine the role of Fas/FasL on drug-free tolerance, we measured mRNA levels of Fas/FasL in livers by reverse transcriptase-polymerase chain reaction (RT-PCR), and also protein levels of Fas/FasL in livers by immunohistochemistry and in serum by dot blot assay. PVG recipients bearing DA livers showed serious rejection between post-operative (POD) days 7 and 14, but this rejection was naturally overcome without any immunosuppression. Fas gene and protein products were expressed on almost every cell in livers taken from naive rats, and at any time point in both syngeneic and allogeneic orthotopic liver transplantation (OLT) rats. In contrast, FasL mRNA in DA livers was detectable at POD 2, peaked at POD 14, and declined at POD 63 in allogeneic OLT (DA-PVG). Although the FasL gene was detectable in isografts at POD 14, its expression was much lower than in allografts. The time course and localization of FasL expression indicated that the expression of FasL gradually switched from infiltrating cells to hepatocytes when the rejection was naturally overcome and tolerance was induced in this OLT model. Soluble Fas could constitutively be detected at any time point in the serum of the tolerogenic OLT (DA-PVG) rats and was not diminished during the rejection phase. Soluble FasL peaked at POD 14 in allogeneic OLT, while sFasL was significantly lower in the serum of normal and syngeneic OLT rats. These findings suggest that the Fas and FasL pathways, including soluble forms, may contribute to the control of the immune response in this drug-free tolerance OLT model. PMID- 10540177 TI - Properdin deficiency and meningococcal disease--identifying those most at risk. PMID- 10540178 TI - Differences in cytokine production by peripheral blood mononuclear cells (PBMC) between patients with atopic dermatitis and bronchial asthma. AB - It is widely accepted that type 2 helper T (Th2) lymphocytes play a crucial role in the pathogenesis of atopic dermatitis (AD) as well as bronchial asthma (BA). We measured the amounts of IL-5 and interferon-gamma (IFN-gamma) produced by PBMC upon stimulation with house dust mite (HDM) or Candida albicans (CA) in 17 children (3-15 years) with AD, and compared these values with those of 16 children with BA. Although IL-5 production by PBMC upon stimulation with HDM in patients with AD was significantly higher than that in 13 non-atopic controls (geometric mean = 23.4 pg/ml versus 5.9 pg/ml, P < 0.05), it was significantly lower than that in patients with BA (177.8 pg/ml, P < 0.001). The amount of IL-5 produced by PBMC upon stimulation with CA was also significantly lower in patients with AD than in those with BA (7.2 pg/ml versus 100.0 pg/ml, P < 0.001). The production of IFN-gamma by PBMC stimulated with HDM or CA was also significantly lower in patients with AD than in those with BA (HDM 4. 3 pg/ml versus 12.6 pg/ml, P < 0.05; CA 6.5 pg/ml versus 60.3 pg/ml, P < 0.001). Consequently the ratio of IL-5 to IFN-gamma production was high not only in patients with BA but also in those with AD. These findings suggest that there are some differences in the regulation of in vivo cytokine production between patients with AD and those with BA, although a Th2-dominant profile is common to both. PMID- 10540179 TI - A lack of evidence for down-modulation of CD3 zeta expression in colorectal carcinoma and pregnancy using multiple detection methods. AB - Loss of the T cell receptor-associated CD3 zeta chain has been proposed as a possible mechanism of the acquired immunosuppression in both tumour-bearing hosts, and in symptomatic patients with HIV infection. However, other reports suggest that the zeta-chain loss may in part be caused by protease activity of contaminating phagocytes ex vivo. Using flow cytometry and Western blot analysis on highly purified T cells, and ensuring adequate addition of protease inhibitors, we have studied the expression of CD3zeta on peripheral blood T cells from patients with colorectal carcinoma, and compared these with normal controls, and pregnant donors, as a further example of an immunocompromised state. Immunohistochemistry was performed on tumour sections from patients with colorectal carcinoma to measure CD3zeta expression in tumour infiltrating T cells, and compared with normal mucosa and tonsil. Using these three approaches, our data provide no evidence for downregulation of CD3zeta chain expression either in colorectal carcinoma or pregnancy and suggest that this explanation is unlikely to fully account for the reduced T cell function associated with these conditions in all patients. PMID- 10540180 TI - Limitations of the semisynthetic library approach for obtaining human monoclonal autoantibodies to the thyrotropin receptor of Graves' disease. AB - Graves' disease (GD) is characterized by the presence of autoantibodies against the TSH-receptor (TSH-R) which are pathogenic and, upon binding to the receptor, trigger intracellular signal transduction. The autoantibodies are oligoclonal and as they are responsible for disease activity, their characterization would lead to a better understanding of the development of GD. Attempts to isolate anti-TSH R antibodies from patients have proved to be difficult due to the exceedingly low serum levels due to rarity of these B cells, together with difficulties in obtaining purified TSH-R capable of interacting with patients autoantibodies. We employed phage antibody display technology and performed selection with a previously characterized semisynthetic antibody library on the purified extracellular ectodomain of the TSH-R. We report the isolation of six different anti-TSH-R monoclonal phage antibodies (moPhabs) from this library. All the moPhabs recognized TSH-R and its recombinant fragments by Western blotting, but failed to recognize the native TSH-R by flow cytometry. Consequently, the moPhabs did not lead to TSH-R activation. As these were the first moPhabs to TSH-R, they were analysed in terms of nucleotide and amino acid sequence and epitope specificity on the receptor. The moPhabs used immunoglobulin VH1 and VH3 germ line genes, all associated with Vlambda3 genes. Interestingly, the CDR3 regions of all moPhabs were remarkably similar, though not identical. In light of the common CDR3 usage, the epitopes recognized on TSH-R appeared to be restricted to amino acids residues 405-411 and 357-364. In summary, our results show that semisynthetic libraries may be limited in isolating human monoclonal antibodies that resemble pathogenic antithyrotropin receptor autoantibodies present in patients with GD. It is likely that until preparations of purified TSH-R that can be recognized by patients autoantibodies become available, similar to the recently described glycosylphosphatidylinositol (GPI) anchored TSH-R ectodomain, monoclonal antibodies from phage antibody display to TSH-R will be limited for isolating the rare, pathogenic antibodies of GD. PMID- 10540181 TI - Platelet expression of tumour necrosis factor-alpha (TNF-alpha), TNF receptors and intercellular adhesion molecule-1 (ICAM-1) in patients with proliferative diabetic retinopathy. AB - Microvascular complications of insulin-dependent diabetes mellitus (IDDM) have been strongly associated with platelet abnormalities, whilst TNF-alpha has been implicated in the pathogenesis of this condition. However, at present it is not clear whether human circulating platelets express TNF-alpha or TNF receptors (TNF R) or whether impaired expression of these molecules and of the TNF-reactive adhesion molecule ICAM-1 may be associated with platelet abnormalities in patients with IDDM. On this basis we investigated the platelet expression of these molecules in patients with IDDM complicated or uncomplicated by proliferative diabetic retinopathy (PDR) and in healthy subjects. We observed that the proportion of platelets staining for TNF-alpha was significantly higher in IDDM patients with active PDR than in patients without microvascular complications (P = 0.0078), quiescent PDR (P = 0.003) or healthy subjects (P = 0.0013). Patients with active PDR also showed a higher proportion of platelets expressing TNF-RI (P = 0. 0052) and TNF-RII (P = 0.015) than healthy controls or patients with quiescent PDR (P = 0.009 and 0.0006, respectively). In addition, the percentage of ICAM-1+ platelets was significantly higher in patients with active PDR than in patients with quiescent PDR (P = 0.0065) or normal subjects (P = 0.013). There was a direct correlation between platelet expression of TNF-alpha and that of TNF-R in PDR patients, indicating that platelet staining for TNF alpha may be due to binding of this cytokine to its receptors. The results suggest that increased platelet expression of TNF-alpha, TNF-R and ICAM-1 in IDDM patients may constitute important markers of thrombocyte abnormalities during the development of microvascular complications of diabetes mellitus. PMID- 10540182 TI - Externalization of tropomyosin isoform 5 in colon epithelial cells. AB - Ulcerative colitis (UC) is associated with autoantibody response to a cytoskeletal protein, human tropomyosin (hTM) isoform-5 (hTM5). Because hTM5 is an intracellular protein, it may remain inaccessible to the autoantibodies. Therefore, we have investigated the possibility of externalization of hTM5 in colon epithelial cells. Freshly isolated colonic and small intestinal epithelial cells and LS-180 colon cancer cell line were examined for surface expression of hTM5 by flow cytometric analysis using hTM isoform-specific MoAbs. The extracellular release of hTM5 was determined by Western blot and radioimmunoprecipitation analyses. Physical association of hTM5 with a membrane associated colon epithelial protein (CEP) was examined by co-immunoprecipitation of hTM5 with anti-CEP MoAb, and CEP with anti-hTM5 MoAb. Cell surface expression of hTM5 was observed in colonic epithelial and LS-180 cells but not in small intestinal epithelial cells. LS-180 cells spontaneously released hTM5 as well as CEP into the culture medium that was significantly stimulated by a calcium ionophore, A23187, but inhibited by phorbol-12-myristate-13-acetate, monensin and methylamine. Co-immunoprecipitation experiments revealed that hTM5 forms a complex with CEP. We conclude that hTM5 is externalized in colon but not in small intestinal epithelial cells. The physical association of hTM5 with CEP suggests a possible chaperone function of CEP in the transport of hTM5, a putative target autoantigen in UC. PMID- 10540183 TI - In vitro activated CD4+ T cells from interferon-gamma (IFN-gamma)-deficient mice induce intestinal inflammation in immunodeficient hosts. AB - To investigate the role of IFN-gamma in the immunopathogenesis of inflammatory bowel disease (IBD), severe combined immunodeficient (SCID) mice were transplanted with in vitro activated CD4+ T cells from either wild-type (WT) or IFN-gamma-deficient (IFN-gammaKO) BALB/c mice. In vitro, the two types of T cells displayed comparable proliferation rates and production of tumour necrosis factor alpha (TNF-alpha), IL-2, IL-4 and IL-10 after concanavalin A (Con A) stimulation. When transplanted into SCID mice, WT CD4+ blasts induced a lethal IBD, whereas IFN-gammaKO blasts induced a less severe intestinal inflammation with moderate weight loss. Intracellular cytokine staining of lamina propria lymphocytes (LPL) revealed comparable fractions of CD4+ T cells positive for TNF-alpha, IL-2 and IL 10 in the two groups of transplanted SCID mice, whereas a two-to-three-fold increase in the fraction of IL-4-positive cells was found in IFN-gammaKO transplanted SCID mice. Flow cytometric analyses showed strong up-regulation of MHC class II expression of colonic epithelial cells of WT-CD4+ T cell transplanted compared with IFN-gammaKO-transplanted SCID mice. A significantly higher fraction of CD4+ LPL were found to enter the cell cycle, i.e. to incorporate bromo-deoxy-uridine, and to undergo apoptosis in vivo in WT transplanted compared with IFN-gammaKO-transplanted SCID mice. These data point towards an important role for IFN-gamma in the development of IBD in SCID mice. The inflammation might be initiated and subsequently enhanced by the ability of IFN-gamma to induce de novo MHC class II expression in the colonic epithelium, a change which could lead to increased antigen processing and production of local proinflammatory cytokines, CD4+ T cell turnover and thereby to exaggeration of disease. PMID- 10540184 TI - Impaired production of proinflammatory cytokines in response to lipopolysaccharide (LPS) stimulation in elderly humans. AB - Ageing is associated with decreased resistance to bacterial infections and concomitant increased circulating levels of inflammatory cytokines. The purpose of the present study was to research age-related changes in levels of early mediators of the acute-phase response in whole blood supernatants following LPS stimulation, representing an ex vivo model of sepsis. Levels of tumour necrosis factor-alpha (TNF-alpha), IL-1beta and IL-6 in whole blood supernatants were measured after in vitro LPS stimulation for 24 h in 168 elderly humans aged 81 years from the 1914 cohort in Glostrup, Denmark and in 91 young controls aged 19 31 years. Levels of TNF-alpha and IL-1beta were significantly lower in elderly humans compared with young controls, whereas no difference was detected with regard to IL-6. Elderly humans with low body mass index had the lowest levels of IL-1beta. Young women had lower levels of proinflammatory cytokines compared with young men, but this difference was blurred by ageing. No relation was found between circulating plasma levels of TNF-alpha and levels after in vitro LPS stimulation. In conclusion, decreased production of TNF-alpha and IL-1beta after exposure to LPS may reflect impaired host defence against infections in the elderly and be of importance in elderly humans with underlying health disorders. However, the clinical relevance is questionable in healthy elderly people because decreased levels were found compared with young men but not compared with young women. PMID- 10540185 TI - Perioperative serum levels of tumour-necrosis-factor alpha (TNF-alpha), IL-1 beta, IL-6, IL-10 and soluble IL-2 receptor in patients undergoing cardiac surgery with cardiopulmonary bypass without and with correction for haemodilution. AB - Cardiac surgery with cardiopulmonary bypass (CPB) leads to a systemic inflammatory response with secretion of cytokines. Alterations in the serum concentrations of cytokines have important prognostic significance. Reports on cytokine release during cardiac surgery with CPB have yielded conflicting results. Haemodilution occurs with the onset of CPB resulting in large fluid shifts during the perioperative course of cardiac procedures. In the present study we compare the perioperative course of serum concentrations of TNF-alpha, IL-1beta, IL-6, IL-10 and sIL-2R with and without correction for haemodilution in patients undergoing coronary artery bypass grafting (CABG) surgery. Twenty male patients undergoing elective CABG surgery with CPB and general anaesthesia using a balanced technique with sufentanil, isoflurane and midazolam were enrolled into the study. Serum levels of TNF-alpha, IL-1beta, IL-6, IL-10 and sIL-2R were measured using commercially available ELISA kits. Simultaneous haematocrit values were obtained at all sample times. Statistical analysis was performed by non parametric analysis of variance and t-tests for data corrected for haemodilution and data that were not corrected for haemodilution. Adjusted significance level was P < 0.01. Intra-operatively, up to the second post-operative day PCV values were significantly decreased compared with preoperative values. Cytokine measurements not corrected for haemodilution were significantly lower than the corrected values. The perioperative haemodilution and decrease in PCV may lead to an underestimation of the cytokine secretion in post-operative patients. We conclude that cytokine measurements were significantly influenced by the perioperative haemodilution and the subsequent decrease in PCV and may in part account for the varying results reported in the literature regarding cytokine release in patients undergoing CABG surgery. PMID- 10540186 TI - Peripheral blood antigen-presenting cells from African-Americans exhibit increased CD80 and CD86 expression. AB - Despite the increased incidence and severity of many autoimmune diseases and transplant rejection in African-Americans (AA) compared with Caucasians (CS), very few studies have addressed issues of racial variation during antigen presentation. This investigation was performed as a preliminary exploration of differences in peripheral blood cell costimulatory functions between healthy AA (n = 20) and CS (n = 20) subjects. The expression of surface costimulatory molecules on peripheral blood cells, mononuclear cells enriched by Ficoll density centrifugation, and plastic adherent antigen-presenting cells (APC) was determined by flow cytometry using fluorescent-labelled MoAbs. The expression of both B7 costimulatory molecules was significantly higher on the cells from AA subjects compared with cells from CS subjects (CD80, P < 0.05; CD86, P < 0.05). Also, following 18 h of culture with rhIL-1beta, there was a significant increase in the percentage of APC from AA expressing high levels of the costimulatory molecule CD80 (P < 0.05). Costimulatory function during mitogen and antigen presentation was determined by 3H-thymidine incorporation during T cell proliferation. Purified T cells from AA subjects demonstrated significantly increased proliferation to phytohaemagglutinin (PHA). The differences reported here suggest that racial variations in peripheral blood APC characteristics may exist. Given the importance of costimulation in maintaining long-term immune responses, these data suggest a further direction for the investigation of racial disparity in autoimmune disease pathology and transplant rejection rates. PMID- 10540187 TI - Activation of human neutrophils by mycobacterial phenolic glycolipids. AB - The interaction between mycobacterial phenolic glycolipids (PGLs) and phagocytes was studied. Human neutrophils were allowed to interact with each of four purified mycobacterial PGLs and the neutrophil production of reactive oxygen metabolites was followed kinetically by luminol-/isoluminol-amplified chemiluminescence. The PGLs from Mycobacterium tuberculosis and Mycobacterium kansasii, respectively, were shown to stimulate the production of oxygen metabolites, while PGLs from Mycobacterium marinum and Mycobacterium bovis BCG, respectively, were unable to induce an oxidative response. Periodate treatment of the M. tuberculosis PGL decreased the production of oxygen radicals, showing the importance of the PGL carbohydrate moiety for the interaction. The activation, however, could not be inhibited by rhamnose or fucose, indicating a complex interaction which probably involves more than one saccharide unit. This is in line with the fact that the activating PGLs from M. tuberculosis and M. kansasii contain tri- and tetrasaccharides, respectively, while the nonactivating PGLs from M. marinum and M. bovis BCG each contain a monosaccharide. The complement receptor 3 (CR3) has earlier been shown to be of importance for the phagocyte binding of mycobacteria, but did not appear to be involved in the activation of neutrophils by PGLs. The subcellular localization of the reactive oxygen metabolites formed was related to the way in which the glycolipids were presented to the cells. PMID- 10540188 TI - Human cytokine responses induced by gram-positive cell walls of normal intestinal microbiota. AB - The normal microbiota plays an important role in the health of the host, but little is known of how the human immune system recognizes and responds to Gram positive indigenous bacteria. We have investigated cytokine responses of peripheral blood mononuclear cells (PBMC) to Gram-positive cell walls (CW) derived from four common intestinal indigenous bacteria, Eubacterium aerofaciens (Eu.a. ), Eubacterium limosum (Eu.l.), Lactobacillus casei (L.c.), and Lactobacillus fermentum (L.f.). Our results indicate that Gram-positive CW of the normal intestinal microbiota can induce cytokine responses of the human PBMC. The profile, level and kinetics of these responses are similar to those induced by lipopolysaccharide (LPS) or CW derived from a pathogen, Streptococcus pyogenes (S.p.). Bacterial CW are capable of inducing production of a proinflammatory cytokine, tumour necrosis factor-alpha (TNF-alpha), and an anti-inflammatory cytokine, IL-10, but not that of IL-4 or interferon-gamma (IFN-gamma). Monocytes are the main cell population in PBMC to produce TNF-alpha and IL-10. Induction of cytokine secretion is serum-dependent; both CD14-dependent and -independent pathways are involved. These findings suggest that the human cytokine responses induced by Gram-positive CW of the normal intestinal microbiota are similar to those induced by LPS or Gram-positive CW of the pathogens. PMID- 10540189 TI - The effect of site-specific monoclonal antibodies directed to toxic shock syndrome toxin-1 in experimental Staphylococcus aureus arthritis. AB - Staphylococcus aureus produces a large number of potential virulence factors, among these the superantigen toxic shock syndrome toxin-1 (TSST-1). We have recently demonstrated that TSST-1 is involved in the pathogenesis of septic arthritis. Recent data show that the TSST-1 molecule is composed of two distinct domains, one proposed to interact with T cell receptor (TCR) and one with the MHC class II. The aim of this study was to assess if interaction between TSST-1 specific MoAbs directed to sites on the MHC and/or TCR Vbeta affects the development of experimental S. aureus-induced arthritis. For that purpose we used a panel of seven MoAbs, which were injected intraperitoneally before and after inoculation with a TSST-1-producing S. aureus strain. Administration of antibodies did not affect the development of arthritis, suggesting inefficacy of such a procedure in neutralization of exotoxin-mediated disease manifestations. PMID- 10540190 TI - Benznidazole, a drug employed in the treatment of Chagas' disease, down-regulates the synthesis of nitrite and cytokines by murine stimulated macrophages. AB - Benznidazole (BZL) is a nitroheterocyclic drug employed in the chemotherapy of Chagas' disease, a protozoan disease caused by Trypanosoma cruzi. Because this parasite mostly replicates in macrophages, we investigated whether BZL was likely to modify the synthesis of macrophage mediators such as nitrite, tumour necrosis factor-alpha (TNF-alpha), IL-1beta, IL-6 and IL-10. Control and stimulated murine macrophages (lipopolysaccharide (LPS) and/or interferon-gamma (IFN-gamma)) were treated with BZL and measurements were carried out in culture supernatants collected 24 h later. Synthesis of nitrite, IL-6 and IL-10 was maximal upon combined stimulation with LPS + IFN-gamma, whereas lower amounts of the three mediators were detected when both stimuli were given alone. BZL treatment significantly reduced nitrite, IL-6 and IL-10 production, to undetectable levels in some cases, particularly IL-6 and IL-10. LPS was the most potent stimulus of IL-1beta and TNF-alpha production, followed by LPS + IFN-gamma and IFN-gamma in decreasing order. BZL partly inhibited TNF-alpha synthesis, but this effect was smaller than that observed for nitrite, IL-6 and IL-10. LPS-induced production of IL-1beta was also affected by BZL. Semiquantification of gene expression for inducible nitric oxide synthase (iNOS) showed that BZL completely inhibited iNOS gene induction by IFN-gamma, and resulted in respective inhibitions of 30% and 50% with LPS- and LPS + IFN-gamma-stimulated cells. BZL was not cytotoxic on macrophage cultures, as shown by the lactate dehydrogenase activity. Besides its trypanocidal activity, BZL may also alter the balance between pro- and anti inflammatory mediators with important consequences for the course of T. cruzi infection. PMID- 10540191 TI - Properdin deficiency in a large Swiss family: identification of a stop codon in the properdin gene, and association of meningococcal disease with lack of the IgG2 allotype marker G2m(n). AB - Properdin deficiency was demonstrated in three generations of a large Swiss family. The concentration of circulating properdin in affected males was < 0.1 mg/l, indicating properdin deficiency type I. Two of the nine properdin-deficient males in the family had survived meningitis caused by Neisseria meningitidis serogroup B without sequel. Two point mutations were identified when the properdin gene in one of the properdin-deficient individuals was investigated by direct solid-phase sequencing of overlapping polymerase chain reaction (PCR) products. The critical mutation was found at base 2061 in exon 4, where the change of cytosine to thymine had generated the stop codon TGA. The other mutation was positioned at base 827 in intron 3. The stop codon in exon 4 was also demonstrated by standard dideoxy sequencing in three additional family members. The question was asked if genetic factors such as partial C4 deficiency and IgG allotypes could have influenced susceptibility to meningococcal disease in the family. No relationship was found between C4 phenotypes and infection. Interestingly, the two properdin-deficient males with meningitis differed from the other properdin-deficient persons in that they lacked the G2m(n) allotype, a marker known to be associated with poor antibody responses to T-independent antigens. This implies that the consequences of properdin deficiency might partly be determined by independent factors influencing the immune response. PMID- 10540192 TI - Expression of the BLM gene in human haematopoietic cells. AB - Bloom's syndrome (BS) is a rare autosomal recessive disorder characterized by stunted growth, sun-sensitive erythema and immunodeficiency. Chromosomal abnormalities are often observed. Patients with BS are highly predisposed to cancers. The causative gene for BS has been identified as BLM. The former encodes a protein, which is a homologue of the RecQ DNA helicase family, a family which includes helicases such as Esherichia coli RecQ, yeast Sgs1, and human WRN. WRN is encoded by the gene that when mutated causes Werner's syndrome. The function of BLM in DNA replication and repair has not yet been determined, however. To understand the function of BLM in haematopoietic cells and the cause of immunodeficiency in BS, expression of the BLM gene in various human tissues and haematopoietic cell lines was analysed and the involvement of BLM in immunoglobulin rearrangement examined. In contrast to WRN, BLM was expressed strongly in the testis and thymus. B, T, myelomonocytic and megakaryocytic cell lines also expressed BLM. All of the examined sequences at the junction of the variable (V), diversity (D) and joining (J) regions of the immunoglobulin heavy chain genes were in-frame, and N-region insertions were also present. The frequency of abnormal rearrangements of the T cell receptor was slightly elevated in the peripheral T cells of patients with BS compared with healthy individuals, whereas a higher frequency of abnormal rearrangements was observed in the cells of patients with ataxia-telangiectasia (A-T). In DND39 cell lines, the induction of sterile transcription, which is required for class switching of immunoglobulin heavy-chain constant genes, was correlated with the induction of the BLM gene. Taking into consideration all these results, BLM may not be directly involved in VDJ recombination, but is apparently involved in the maintenance of the stability of DNA. PMID- 10540194 TI - Release of CXC-chemokines by human lung microvascular endothelial cells (LMVEC) compared with macrovascular umbilical vein endothelial cells. AB - In the present study, the sensitivity of LMVEC and human umbilical vein endothelial cells (HUVEC) to lipopolysaccharide (LPS) and the proinflammatory cytokines IL-1, tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) was compared. To this end, the production of the CC- (MCP-1), CXC- (IL-8, ENA-78, Groalpha, NAP-2, GCP-2) and CX3C (fractalkine) chemokines was studied. A low basal production of these chemokines was observed in both cell types. TNF-alpha, IL-1 and LPS up-regulated all chemokines tested. IFN-gamma however was only able to up-regulate MCP-1 production. LMVEC were more sensitive to IL-1 and LPS compared with HUVEC, since LMVEC produced significantly more MCP 1, ENA-78 and Groalpha (P < 0. 01) under these conditions. Maximal production of MCP-1 in LMVEC was achieved with TNF-alpha (28.4 ng/ml, P < 0.01), whereas IL-1 was the most potent stimulator of ENA-78 (2.78 ng/ml, P < 0.001) and Groalpha (29.2 ng/ml, P < 0.001). IL-8 production in LMVEC cells was maximal after LPS stimulation (28.4 ng/ml), but lower than on HUVEC (P < 0.01). LPS, TNF-alpha and IL-1 stimulation strongly up-regulated all chemokine mRNA. No quantitative differences in mRNA expression between LMVEC and HUVEC were detected for MCP-1 and Groalpha after LPS stimulation. mRNA expression of ENA-78, GCP-2, CX3C and NAP-2 was however higher in LMVEC under LPS stimulation. In contrast, IL-8 mRNA was slightly more expressed in HUVEC under these conditions. These results further support the hypothesis that the microvascular lung endothelium plays an active role in the induction and perpetuation of acute lung injury. PMID- 10540193 TI - Overlapping but distinct specificities of anti-liver-kidney microsome antibodies in autoimmune hepatitis type II and hepatitis C revealed by recombinant native CYP2D6 and novel peptide epitopes. AB - Anti-liver-kidney microsome antibodies (anti-LKM) occur in autoimmune hepatitis (AIH) type II and in a subset of patients with hepatitis C. Anti-LKM1 in AIH are directed against cytochrome P4502D6 (CYP2D6), but conflicting data exist concerning the specificity of anti-LKM in hepatitis C. The aim of this study was to evaluate binding specificities of anti-LKM antibodies in both diseases using novel test antigens as well as their inhibitory capacity on CYP2D6 enzyme activity. Sera from 22 patients with AIH type II and 17 patients with hepatitis C being anti-LKM-positive in the immunofluorescence test were investigated for binding to native recombinant CYP2D6 and liver microsomes by ELISA and immunoblotting, and to synthetic peptides covering the region 254-339 (254-273, 257-269, 270-294, 291-310, 307-324, 321-339, 373-389) as well as the novel peptide 196-218 by ELISA. Furthermore, all sera were tested for inhibition of CYP2D6-dependent bufuralol 1'-hydroxylase activity. Twenty of the 22 AIH type II sera (91%) and nine of the 17 hepatitis C sera (53%) were positive for CYP2D6 by ELISA and/or immunoblotting. The previously described major peptide epitope comprising CYP2D6 amino acids 257-269 was recognized by 16 of the 22 AIH sera but by only one hepatitis C serum. A further epitope, 196-218, could be defined for the first time as another immunodominant epitope for AIH because it was recognized by 15 of the 22 AIH (68%) but only three of the 17 hepatitis C sera (18%). With the exception of the peptide 254-273, the other peptides showed no significant reactivity. Analysing the inhibitory properties of anti-LKM antibodies it emerged that 95% of AIH sera and 88% of hepatitis C sera inhibited enzyme function. These data indicate that anti-LKM antibodies in AIH and hepatitis C react with CYP2D6, as shown by their inhibitory activity, and that besides the known epitope 257-269 a further immunodominant epitope exists on CYP2D6 which is recognized by sera from patients with AIH II but hardly by sera from patients with hepatitis C. PMID- 10540195 TI - Increased levels of circulating acetylcholine receptor (AChR)-reactive IL-10 secreting cells are characteristic for myasthenia gravis (MG). AB - Antibodies to the nicotinic AChR are pivotal in the immunopathogenesis of MG. Cytokines produced by T-helper cells are important regulators of humoral immune responses. IL-10 is considered an anti-inflammatory cytokine, but it promotes B cell activation and worsens experimental autoimmune MG in Lewis rats, an experimental model of MG. To study IL-10 and, as a control, interferon-gamma (IFN gamma) in MG, we used an enzyme-linked immunospot (ELISPOT) assay, thereby assessing numbers of blood mononuclear cells (MNC) secreting IL-10 and IFN-gamma spontaneously and after stimulation with AChR. Low numbers of IL-10-secreting cells were regularly found in peripheral blood from patients with MG as well as in controls with other neurological diseases and healthy subjects. However, only MG patients had elevated blood levels of AChR-reactive IL-10- and IFN-gamma secreting cells. The MG patients showed no responses to the control autoantigen myelin basic protein, underlining the specificity of the IL-10 and IFN-gamma responses. Immunosuppressive treatment reduced numbers of AChR-reactive IFN-gamma secreting cells but increased the numbers of IL-10-secreting cells. The numbers of IL-10-secreting cells tended to be higher in patients with generalized versus ocular MG, further suggesting that the augmented IL-10 responses may be important in the pathogenesis and perpetuation of MG. PMID- 10540196 TI - Induced nitric oxide (NO) synthesis in heterologous nephrotoxic nephritis; effects of selective inhibition in neutrophil-dependent glomerulonephritis. AB - Increased NO synthesis, due to inducible NO synthase (iNOS) activity, is found in macrophage-associated glomerulonephritis. Little is known about NO in neutrophil dependent immune complex inflammation, and its role remains controversial. We therefore studied early phase heterologous nephrotoxic nephritis (HNTN) induced in rats by nephrotoxic globulin and the effects of selective iNOS inhibition of this model. At 2 h of the model iNOS mRNA was induced and nitrite (NO-2) was generated in glomeruli incubated ex vivo (5.2 +/- 1.0 nmol/2000 glomeruli per 24 h). There were 14.7 +/- 2.2 polymorphonuclear neutrophils (PMN)/glomerulus (normal controls 0.1 +/- 0.1). At 8 h urinary protein was 69 +/- 15.3 (normal controls 0. 6 +/- 0.2 mg/24 h). Peritoneal PMN expressed iNOS and produced significant NO-2 (basal 11.2 +/- 0.3 nmol/106 cells per 24 h). Selective iNOS inhibition with L-N6-(1-iminoethyl)-lysine (L-NIL) in vitro inhibited nephritic glomerular and PMN NO-2 synthesis. In HNTN L-NIL in vivo significantly suppressed elevated plasma NO-2/NO-3 levels (representative experiment: 17 +/- 2 microM, untreated 40 +/- 4 microM, P = < 0.01, normal control 18 +/- 2 microM). This inhibition did not affect leucocyte infiltration into glomeruli or induce thrombosis. There was no consistent effect on proteinuria. This is the first demonstration of glomerular iNOS induction and high output NO production in the acute phase of PMN-dependent acute immune complex glomerulonephritis. Selective iNOS inhibition does not affect the primary mechanism of injury (leucocyte infiltration) in this model. PMID- 10540197 TI - Thalidomide analogue CC1069 inhibits development of rat adjuvant arthritis. AB - The cytokine tumour necrosis factor-alpha (TNF-alpha) has been implicated in the aetiology of rheumatoid arthritis in humans as well as of experimental arthritis in rodents. Thalidomide, and to a greater extent the new thalidomide analogue CC1069, inhibit monocyte TNF-alpha production both in vitro and in vivo. The aim of the present study is to establish whether these drugs block production of TNF alpha as well as IL-2 by rat leucocytes and whether this inhibition affects the development of rat adjuvant arthritis (AA). Cultured splenocytes were stimulated with either lipopolysaccharide (LPS) or concanavalin A (Con A) in the presence of thalidomide, CC1069, or solvent, and the production of TNF-alpha and IL-2 were compared. Next, adjuvant was injected into the base of the tail of rats without or with daily intraperitoneal injections with 100-200 mg/kg per day thalidomide or 50-200 mg/kg per day CC1069. Disease activity, including ankle swelling, hind limb radiographic and histological changes, weight gain, and ankle joint cytokine mRNA levels, were monitored. CC1069, but not the parent drug thalidomide, inhibited in vitro production of TNF-alpha and IL-2 by stimulated splenocytes in a dose-dependent manner. In vivo, a dose-dependent suppression of AA disease activity occurred in the CC1069-treated animals. In contrast, thalidomide-treated rats experienced comparable arthritis severity to placebo-treated animals. There was also a reduction in TNF-alpha and IL-2 mRNA levels in the ankle joints of CC1069-treated rats compared with thalidomide- and placebo-treated arthritic rats. Early initiation of CC1069 treatment suppressed AA inflammation more efficiently than delayed treatment. We conclude that thalidomide, which did not suppress TNF-alpha or IL-2 production in vitro by Lewis rat cells, did not suppress development of rat AA. However, the development of rat AA can be blocked by the thalidomide analogue CC1069, which is an efficient inhibitor of TNF-alpha production and IL-2 in vitro. PMID- 10540198 TI - Antibodies from systemic lupus erythematosus (SLE) sera define differential release of autoantigens from cell lines undergoing apoptosis. AB - SLE is an autoimmune disease characterized by a wide range of anti-cellular and anti-nuclear autoantibodies. Many of these antigens are exposed or altered during apoptosis when the nucleus is dismantled in a controlled manner by caspases. We used Western blotting techniques to demonstrate that autoantibodies in SLE sera recognize antigens released during apoptosis. Reproducible bands, not seen in the untreated cells, with the characteristics of histones were seen when staining apoptotic cell lysates with SLE sera. Normal sera recognized some of these bands but much less strongly. Different triggers of apoptosis did not produce marked differences in the antigens recognized. We also compared different cell lines (Jurkat and U937) and found that the staining differed for one autoantigen in particular. The differential release of autoantigens by apoptotic cells may have relevance to the variety of autoantibodies seen in SLE. PMID- 10540200 TI - Primary immunodeficiency diseases. Report of an IUIS Scientific Committee. International Union of Immunological Societies. PMID- 10540199 TI - Up-regulation of macrophage migration inhibitory factor in acute renal allograft rejection in the rat. AB - Recent studies have identified a key role for macrophage migration inhibitory factor (MIF) in a number of immune cell-mediated diseases. The current study investigated the potential role of MIF in acute allograft rejection. Lewis rats underwent bilateral nephrectomy and then received an orthotopic DA renal allograft or an orthotopic Lewis renal isograft. Groups of six animals were killed at day 1 or 5 after transplantation. No immunosuppression was used. Animals receiving a renal allograft exhibited severe rejection on day 5, as shown by high levels of serum creatinine, very low rates of creatinine clearance, and severe tubulitis with a dense macrophage and T cell infiltrate. In contrast, isografts had normal renal function on day 5 with no histological evidence of rejection. Northern blotting showed that renal MIF mRNA expression was unchanged at day 1, but was increased 3.5-fold on day 5. In situ hybridization showed a marked increase in MIF mRNA expression by tubular cells and MIF mRNA expression by many infiltrating mononuclear cells in day 5 allografts. Immunostaining confirmed an increase in tubular MIF protein expression, particularly in areas of severe tubular damage with prominent leucocytic infiltration. Double staining showed that many infiltrating macrophages and T cells expressed the MIF protein in day 5 allografts. There was only a minor increase in MIF expression in day 5 isografts, demonstrating that neither surgical injury nor stress cause significant up-regulation of MIF expression in allograft rejection. In conclusion, this study has demonstrated that local MIF production is specifically increased in acute renal allograft rejection. These results suggest that MIF may play an important role in the cellular immune response mediating acute allograft rejection. PMID- 10540201 TI - Safety and availability of immunoglobulin replacement therapy in relation to potentially transmissable agents. IUIS Committee on Primary Immunodeficiency Disease. International Union of Immunological Societies. PMID- 10540202 TI - Cholera toxin-induced alteration of the phenotype and behaviour of an ovarian carcinoma cell line, SR8. AB - Cholera toxin (CT) has been reported to cause a variety of effects on several different cell types. Recently, CT has been shown to increase the susceptibility of ovarian carcinoma cells to cytotoxicity mediated by a variety of effector cells (natural killer, lymphokine-activated killer cells and tumour-associated lymphocytes derived from ascites of ovarian cancer patients) of both autologous and allogenic background. In the present study, CT demonstrated several effects on a newly established ovarian carcinoma line (SR8)1 when added to the culture medium at a concentration of 12.5 ng/mL for 2 days. Cholera toxin altered SR8 morphology to a uniform polygonal cellular shape, with less cell dispersion than the non-CT treated cells. Cholera toxin prolonged the population doubling time by approximately 10 h. The CT-treated SR8 cells exhibited reduced epidermal growth factor receptor expression (39 versus 50%), and increased carbohydrate antigen 125 expression (45 versus 2%) in both immunocytochemical and quantitative flow cytometric analyses. These changes in morphology and tumour marker expression were reversible when CT was removed from the culture. The CT-treated SR8 cells showed reduced capacity to generate tumours in female nude mice in comparison with non-CT treated cells, which produce both subcutaneous and intraperitoneal xenografts with local invasion in an animal model. Cytogenetic analysis of the cell line SR8 before and during treatment with CT showed no new clonal rearrangements. The possible mechanisms involved and the influence of CT on the biological behaviour of ovarian tumour cells are discussed. PMID- 10540203 TI - Preproenkephalin is a Th2 cytokine but is not required for Th2 differentiation in vitro. AB - Preproenkephalin (PPNK) mRNA expression has been detected in many cells of the immune system, including monocytes and lymphocytes. In the present paper, the expression of PPNK mRNA in purified CD4+ Th1 and Th2 lymphocyte subpopulations is investigated and correlated with the presence of the opioid neuropeptides leu- and met-enkephalin. We found that PPNK mRNA and met-enkephalin were present at higher levels in the Th2 cultures compared with the Th1 cultures. Lymphocytes from PPNK-deficient mice were then used to look at the role of PPNK in Th2 lymphocyte differentiation. Lymphocytes from these mice could be driven into a Th2 phenotype, suggesting that cultures containing IL-4 do not require PPNK for Th2 differentiation. PMID- 10540204 TI - Random length assortment of human and mouse T cell receptor for antigen alpha and beta chain CDR3. AB - In view of the recently determined three-dimensional structures of complexes formed by the T cell receptor for antigen (TCR), the processed peptide and the MHC class I molecule, it is expected that the combined configuration formed by the third complementarity determining regions (CDR3) of TCR alpha and beta chains will be very restricted in size and shape due to the limited length variations of the processed peptides. Thus, the combined TCR alpha and beta chain CDR3 lengths should have a fairly narrow distribution. This feature can be due to the selective association of long alpha chain CDR3 with short beta chain CDR3 and vice versa or due to random assortment of alpha and beta chain CDR3 of even narrower length distribution. Based on existing translated amino acid sequence data, it has been found that the latter mechanism is responsible. PMID- 10540205 TI - The effect of molecular weight and beta-1,6-linkages on priming of macrophage function in mice by (1,3)-beta-D-glucan. AB - 1,3-beta-D-glucans (glucans) are structural elements in the cell walls of yeast and fungi with immunomodulatory properties, mediated through their ability to activate macrophages. This study assessed the activation of cells of the peritoneal cavity between 3 and 90 days after i.p. injection of particulate yeast glucan differing in molecular weight (MW) and degree of (1,6)-linkages. Female QS mice, 7-9 weeks of age, were injected, i.p., with varying doses of low (< 5 x 10(5)), medium (1-2 x 10(6)) or high (> 3 x 10(6)) MW glucans, all with low (< 5%) beta-(1,6)-linkages, or high MW (> 3 x 10(6)) glucan with high 1,6-linkages (> 20%). All glucans induced a transient increase in the proportion of neutrophils and eosinophils and a reduction in mast cell numbers in the peritoneal cavity. Peritoneal macrophages showed an altered morphology, increased intracellular acid phosphatase, increased LPS-stimulated NO production and increased PMA-stimulated superoxide production. There were no significant changes in serum lysozyme levels. Most macrophage activities returned to control levels by 28 days post injection of 1, 3-beta-D-glucan. There was a trend for higher MW or (1,6)-linked, (1, 3)-beta-D-glucans to be more stimulatory. It was concluded that particulate yeast (1,3)-beta-D-glucan is an effective stimulator of immune function, the efficiency of which may be influenced by the MW and degree of (1,6) linkages. PMID- 10540207 TI - Dendritic cell origins: puzzles and paradoxes. AB - In the present review, a series of studies on the origins of dendritic cells of mice and humans are summarized. Several subsets of mature dendritic cells found in vivo are described and these may correspond to distinct lineages. There is evidence that some dendritic cells are myeloid-derived and that others are lymphoid-derived. The different ways of generating dendritic cells are examined and an attempt to reconcile the differences seen using mouse and human culture models is made. The particular case of Langerhans cells is discussed and an historical overview of the biology of the plasmacytoid T cells, which may represent a distinct 'lymphoid-related' dendritic cell lineage, is given. It is concluded that three or four different pathways lead to the development of different subtypes of dendritic cells. PMID- 10540206 TI - Dendritic cells at the end of the millennium. AB - We have recently proposed a dual role for dendritic cells (DC) in the amplification of innate immune responses and in the activation of adaptive immune responses. The DC are localized along the major routes of entry of micro organisms, where they perform a sentinel function for incoming pathogens. Soon after interaction with appropriate stimuli, DC undergo a coordinated process of maturation and respond to danger signals by re- programming their functions. The DC first regulate leucocyte recruitment at the site of inflammation, through the production of chemokines, inflammatory cytokines and interferons, and then they acquire migratory properties and undergo a rapid switch in chemokine receptor expression. This allows them to leave the inflamed tissue and to reach the lymph node T cell area. During this migration, DC complete their maturation process and acquire the ability to prime T cell responses. Thus, DC bridge innate and adaptive immunity. PMID- 10540208 TI - Dendritic cells: the driving force behind autoimmunity in rheumatoid arthritis? AB - Dendritic cells (DC) are likely to play a significant role in immune-mediated diseases such as autoimmunity and allergy. To date there are few treatments capable of inducing permanent remission in rheumatoid arthritis (RA) and elucidation of the role of DC may provide specific strategies for disease intervention. Dendritic cells have proven to be powerful tools for immunotherapy and investigations are under way to determine their clinical efficacy in transplantation and viral and tumour immunotherapy. The present review will focus on the current view of DC and their role in autoimmunity, in particular RA. Two possible roles for DC in the pathogenesis of RA will be proposed, based on recent advances in the field. PMID- 10540209 TI - Regulation of T helper cell differentiation by respiratory tract dendritic cells. AB - Studies on dendritic cells (DC) of the respiratory and gastric mucosae have identified an extensive network of cells that represent the predominant antigen presenting cell type at these sites. Under steady-state conditions, respiratory tract DC (RTDC) are specialized for antigen uptake and spontaneously migrate to local lymph nodes, although in vivo transfer studies have shown that the T-cell priming activity of these cells is restricted to low-level, Th2-skewed responses. Following exposure to inflammatory stimuli, the migration of RTDC to lymph nodes is accelerated and is associated with a rapid and dramatic increase in the ability of these cells to induce both Th1- and Th2-dependent immunity. Under normal circumstances, however, responsiveness of epithelial RTDC to maturation stimuli is regulated by locally produced micro-environmental factors, including pro-inflammatory cytokines, reactive oxygen species and prostanoids. These studies have led to a greater understanding of airway DC function and their role in T helper cell differentiation and provide the basis for future studies to determine the role of the cells in the aetiology and pathogenesis of respiratory immunoinflammatory disorders. PMID- 10540210 TI - Long-term stroma-dependent cultures are a consistent source of immunostimulatory dendritic cells. AB - A long-term culture (LTC) system has been established that supports the continuous production of dendritic cells (DC) from haemopoietic cells present in the culture. The production of cells depends on the presence of an intact stromal cell layer containing a mixture of fibroblasts and endothelial cells. Cells are shed from foci of dividing cells in contact with the stromal cell matrix. They resemble DC in terms of morphology and cell surface marker expression. The LTC can be derived from different lymphoid tissues, but most success has been achieved with murine spleen. Different LTC vary in capacity to produce immunostimulatory DC. Some LTC produce DC that are very effective APC and can stimulate both mixed lymphocyte and antigen-specific T cell responses. The DC produced in others are weak APC. Different LTC appear to produce DC reflecting different stages of maturation or development, reflected by different phenotypic and functional characteristics. The production of cells within LTC occurs independently of added cytokines and is dependent on maintenance of the stromal cell layer and the presence of a subset of smaller progenitor cells. Long-term cultures remain a valuable source of cells for study of DC development and function. PMID- 10540211 TI - Monocyte-derived dendritic cells as a model for the study of HIV-1 infection: productive infection and phenotypic changes during culture in human serum. AB - Dendritic cells (DC) have been implicated in the initial selection for macrophage tropic HIV-1 during transmission and in the generation of high-level virus replication during interactions with CD4 T cells. The role of DC as viral reservoirs and the extent of productive infection is unclear, but the ability to generate large numbers of DC from blood monocytes has produced a tractable model for study of DC-HIV-1 interactions. When cultured in granulocyte-macrophage colony stimulating factor and IL-4, sorted CD14+ monocytes rapidly lost phagocytic function for both 93 nm and 977 nm latex particles and developed the surface markers and function of DC. After 7 days, when returned to medium containing human serum without cytokines, some monocyte-derived dendritic cells (MDDC) became adherent, but retained the costimulatory markers CD80 and CD86 and continued to express CD83 and CD40. The MDDC stimulated allogeneic CD4 T cells, did not express new macrophage markers and remained non-phagocytic. With or without TNF-alpha, MDDC generated in cytokines were infected by macrophage and T cell-tropic virus and produced higher reverse transcriptase levels than did the autologous monocyte-derived macrophages (MDM). When added to T cells, the infected MDDC were able to infect T cells with a wider range of viral isolates than were MDM. PMID- 10540212 TI - Dendritic cell immunotherapy for cancer: application to low-grade lymphoma and multiple myeloma. AB - The confirmation that most cancers express one or more molecular changes, which may act as tumour-associated antigens (TAA), combined with the knowledge that T lymphocytes recognize even single amino acid differences in MHC presented peptides has stimulated renewed clinical interest in immunotherapeutic strategies. Dendritic cells (DC) are now recognized as specialist antigen presenting cells, which initiate, direct and regulate immune responses. Recent data suggest that DC are not recruited into, or activated by, cancers and that other abnormalities in DC function are associated with malignancy, including multiple myeloma. This provides a rationale for designing immunotherapeutic strategies, which exploit DC as nature's adjuvant either in vivo or in vitro. Low grade lymphoma and multiple myeloma are slowly progressive malignancies, which generally express a unique immunoglobulin idiotype as a potential TAA. Data from animal models and clinical studies suggest that DC-based immunotherapy strategies, applied when the patient has minimal residual disease, may improve the long-term prognosis in these diseases. PMID- 10540213 TI - Langerhans' cell histiocytosis is caused by dysregulation of the E-cadherin-beta catenin cascade: a hypothesis. AB - Langerhans' cell histiocytosis (LCH) is a proliferative disease of cells of the dendritic cell lineage, closely resembling activated Langerhans' cells. The clinical picture of LCH is greatly variable, suggesting a scale of aberrancies at the cellular level. Despite progress in clinical treatment, the aetiology and pathogenesis of this disease remain unknown. In the present paper, we present the hypothesis that dysregulation of the E-cadherin-beta-catenin-Wnt cascade, which has both adhesive and transcriptional functions, may be fundamental to the development of LCH. This hypothesis is founded upon two notions: (i) careful regulation of this cascade is essential in normal Langerhans cell activation; and (ii) abnormalities in the E-cadherin-beta-catenin cascade are a major cause of epithelial neoplastic proliferation. On the basis of this hypothesis, we present three alternative scenarios that may describe the initial steps in the pathogenesis of LCH. PMID- 10540215 TI - Induction of dendritic cell costimulator molecule expression is suppressed by T cells in the absence of antigen-specific signalling: role of cluster formation, CD40 and HLA-class II for dendritic cell activation. AB - Full activation of T lymphocytes by dendritic cells (DC) during antigen presentation is known to require the interaction of several inducible receptor ligand pairs. We have postulated that the reciprocal activation of DC by T lymphocytes is also important. Potential signalling molecules that might increase the stimulatory capacity of DC during antigen presentation to T lymphocytes were tested using an in vitro model. Fresh human blood DC were cocultured with CD4+ and CD8+ allogeneic or with autologous T lymphocytes plus Staphylococcus superantigen A (SEA). Surprisingly, costimulator expression on DC cocultured with T lymphocytes was reduced in comparison to DC cultured alone. However, the minority (10-30%) of DC clustering with T lymphocytes showed antigen-specific up regulation of the CD40, CD80 and CD86 costimulator molecules, whereas the non clustered DC (70-90%) had less up-regulation than control DC cultured alone and did not respond to antigen-specific triggering. Monoclonal antibodies (mAb) to CD40 ligand (CD40L) and human leucocyte antigen (HLA)-DR, but not lymphocyte function-associated antigen-1 (LFA-1), LFA-3 or HLA-class I, significantly inhibited the T-lymphocyte induction of DC costimulator expression. Since HLA class II, but not HLA-class I mAb, inhibited allogeneic T-lymphocyte-mediated activation of DC, CD4 T lymphocytes appear to be the main subset activating DC in the mixed lymphocyte reaction. Cross-linking of CD40, but not HLA-class II, up regulated DC or B-cell costimulator expression. Although direct class II signalling does not appear to play a role in DC activation, antigen-specific T cell recognition contributes via other mechanisms to regulate DC activation. PMID- 10540214 TI - CD40-deficient dendritic cells producing interleukin-10, but not interleukin-12, induce T-cell hyporesponsiveness in vitro and prevent acute allograft rejection. AB - The induction of an immune response or tolerance is mediated by corresponding subsets of dendritic cells (DC). However, the property of tolerogenic DC is not clear. Recently, we have characterized a population of CD11c+ splenic DC derived from long-term mixed leucocyte culture (LT-MLC), which are able to proliferate upon stimulation and have a strong primary mixed leucocyte reaction (MLR) stimulating activity in conventional MLR. In this study, we show that, in contrast to the irradiated ones, non-irradiated LT-MLC-derived DC induce polyclonal antigen-specific T-cell hyporesponsiveness when cocultured with allogeneic splenocytes for 3-11 days. The degree of the hyporesponsiveness increased with the length of coculture. Although these DC expressed major histocompatibility complex class II and B7 costimulatory molecules, which are down-regulated during coculture, they expressed very low or undetectable CD40 before and after coculture, respectively. The CD40-deficient DC spontaneously produce interleukin-10 (IL-10), but not IL-12. The skewed balance between IL-10 and IL-12 is associated with their capability to induce T-cell hyporesponsiveness, because a neutralizing antibody to IL-10, exogenous recombinant IL-12 or lipopolysaccharide (LPS) significantly blocked the hyporesponsiveness. Accordingly, infusion of a small number of non-irradiated LT MLC-derived DC (5x105) significantly prolonged the survival of a vascularized heterotopic murine heart transplant, whereas irradiated DC accelerated graft rejection. These data suggest that CD40-deficient DC producing IL-10, but not IL 12 can induce T-cell hyporesponsiveness in vitro and in vivo. PMID- 10540216 TI - Analysis of mouse dendritic cell migration in vivo upon subcutaneous and intravenous injection. AB - Dendritic cells (DC) have an increasingly important role in vaccination therapy; therefore, this study sought to determine the migratory capacity and immunogenic function of murine bone-marrow (BM)-derived DC following subcutaneous (s.c.) and intravenous (i.v.) injection in vivo. DC were enriched from BM cultures using metrizamide. Following centrifugation, the low-buoyant density cells, referred to throughout as DC, were CD11c(high), Iab(high), B7-1(high) and B7-2(high) and potently activated alloreactive T cells in mixed lymphocyte reactions (MLR). In contrast, the high-density cells expressed low levels of the above markers, comprised mostly of granulocytes based on GR1 expression, and were poor stimulators in MLR. Following s.c. injection of fluorescently labelled cells into syngeneic recipient mice, DC but not granulocytes migrated to the T-dependent areas of draining lymph nodes (LN). DC numbers in LN were quantified by flow cytometric analysis, on 1, 2, 3, 5 and 7 days following DC transfer. Peak numbers of around 90 DC per draining LN were found at 2 days. There was very little migration of DC to non-draining LN, thymus or spleen at any of the time-points studied. In contrast, following i.v. injection, DC accumulated mainly in the spleen, liver and lungs of recipient mice but were largely absent from peripheral LN and thymus. The ability of DC to induce T-cell-mediated immune responses was examined using trinitrobenzenesulphate (TNBS)-derivatized DC (TNBS-DC) to sensitize for contact hypersensitivity responses (CHS) in naive syngeneic recipients. Following s.c. injection, as few as 105 TNBS-DC, but not TNBS granulocytes, sensitized for CHS responses. However, the same number of TNBS-DC failed to induce CHS following i.v. injection. In summary, this study provides new and quantitative data on the organ specific migration of murine BM-derived DC following s.c. and i.v. injection. The demonstration that the route of DC administration determines the potency of CHS induction, strongly suggests that the route of immunization should be considered in the design of vaccine protocols using DC. PMID- 10540217 TI - Expression of the RelB transcription factor correlates with the activation of human dendritic cells. AB - The RelB gene product is a member of the nuclear factor (NF)-kappaB family of transcription factors. It has been identified recently within mouse antigen presenting cells and human monocyte-derived dendritic cells (DC). Disruption of the mouse RelB gene is accompanied, amongst other phenotypes, by abnormalities in the antigen-presenting cell lineages. In order to define RelB expression during human DC differentiation, we have analysed RelB mRNA by reverse transcriptase polymerase chain reaction and RelB protein by intracellular staining in CD34+ precursors and different types of DC preparations. RelB mRNA was not detected in CD34+ precursor populations. Fresh blood DC (lineage-human leucocyte antigen-DR+ (lin-HLA-DR+)) lacked RelB mRNA and cytoplasmic RelB protein but a period of in vitro culture induced RelB expression in blood DC. Purified Langerhans' cells (LC) (CD1a+ HLA-DR+) failed to express RelB mRNA. Immunocytochemical staining identified RelB protein in human skin epithelium. RelB protein was expressed in a very few CD1a+, CD83+ or CMRF-44+ dermal DC but was not present in CD1a+ LC. Tonsil DC (lin-HLA-DR+ CMRF-44+) were positive for RelB mRNA and RelB protein. Intestinal DC (HLA-DR+) also lacked immunoreactive RelB protein. The majority of interdigitating CD83+, CMRF-44+, CMRF-56+ or p55+ DC located in paracortical T lymphocyte areas of lymph node and tonsil contained RelB protein. The expression of RelB mRNA and RelB protein correlates with the activated phase of blood DC and the postmigration cell (activated) stage of tissue DC development. PMID- 10540218 TI - Activation of complement receptor 3 on human monocytes by cross-linking of very late antigen-5 is mediated via protein tyrosine kinases. AB - Bordetella pertussis interacts with very-late antigen-5 (VLA-5) receptors on the human monocyte resulting in cross-linking of these receptors followed by activation of complement receptor 3 (CR3) and firm adhesion of B. pertussis to these monocytes. In the present study we investigated whether protein tyrosine kinases are involved in the activation of CR3 on monocytes, which was assessed by the binding of C3bi-coated erythrocytes (EC3bi). Pre-incubation of monocytes with tyrphostin-A47, a specific protein tyrosine kinase inhibitor, before adherence of the cells to an anti-VLA-5 monoclonal antibody-coated surface, or addition of tyrphostin-A47 within 10 min of the adherence to such surface, reduced the binding of EC3bi to monocytes significantly. Pre-incubation of monocytes with tyrphostin-A47 reduced the binding of B. pertussis to such monocytes as well. Inhibitors of protein kinase A and/or C had no effect on EC3bi binding to monocytes. Cross-linking of VLA-5 on monocytes resulted in tyrosine phosphorylation of several proteins. Together, these results indicate that protein tyrosine kinases are involved in the VLA-5-induced activation of CR3 on human monocytes. PMID- 10540219 TI - Intermediate stages in monocyte-macrophage differentiation modulate phenotype and susceptibility to virus infection. AB - The kinetics of monocyte-macrophage differentiation was analysed using two Swine Workshop Cluster (SWC) CD molecules: SWC1 and SWC9. Myeloid cells were selected by labelling for the common myeloid antigen, SWC3. Confirmation of macrophage identification used acid phosphatase and phagocytosis activities. During differentiation, SWC1 was gradually lost. SWC9 was absent on monocytes but up regulated early. Consequently, monocytes were SWC1+ SWC9- and macrophages were SWC1- SWC9+. An additional, intermediate, cell population was identified as SWC1+ SWC9+. Size and granularity characteristics mirrored the monocyte, macrophage and intermediate-cell phenotypes. Overall, SWC9 up-regulation was central in macrophage differentiation and dependent on plasma factors. The concomitant loss of SWC1 was independent of these factors, but always associated with mature macrophages. Upon up-regulation of SWC9, the SWC1+ SWC9+ intermediate monocytic cells became susceptible to African swine fever virus infection. These results demonstrate the heterogeneity of monocytic cell differentiation and the importance of these characteristics for interaction with monocytotropic viruses. PMID- 10540220 TI - ATP-induced Ca2+ response mediated by P2U and P2Y purinoceptors in human macrophages: signalling from dying cells to macrophages. AB - The activation of macrophages by various stimuli leading to chemotactic migration and phagocytosis is known to be mediated by an increase in intracellular Ca2+ concentration ([Ca2+]i). We measured changes in [Ca2+]i using a Ca2+ imaging method in individual human macrophages differentiated from freshly prepared peripheral blood monocytes during culture of 1-2 days. A transient rise in [Ca2+]i (duration 3-4 min) occurred in 10-15 macrophages in the vicinity of a single tumour cell that was attacked and permeabilized by a natural killer cell in a dish. Similar Ca2+ transients were produced in 90% of macrophages by application of supernatant obtained after inducing the lysis of tumour cells with hypo-osmotic treatment. Ca2+ transients were also evoked by ATP in a dose dependent manner between 0.1 and 100 microm. The ATP-induced [Ca2+]i rise was reduced to less than one-quarter in Ca2+-free medium, indicating that it is mainly due to Ca2+ entry and partly due to intracellular Ca2+ release. UTP (P2U purinoceptor agonist) was more potent than ATP or 2-chloro-ATP (P2Y agonist). Oxidized ATP (P2Z antagonist) had no inhibitory effect. Both cell lysate- and ATP induced Ca2+ responses were inhibited by Reactive Blue 2 (P2Y and P2U antagonist) to the same extent, but were not affected by PPADS (P2X antagonist). Sequential stimuli by cell lysate and ATP underwent long-lasting desensitization in the Ca2+ response to the second stimulation. The present study supports the view that macrophages respond to signal messengers discharged from damaged or dying cells to be ingested, and ATP is at least one of the messengers and causes a [Ca2+]i rise via P2U and P2Y receptors. PMID- 10540221 TI - Differential modes of regulation of interleukin-1beta expression by extracellular matrices. AB - Extracellular matrices (ECMs) can stimulate human monocytic cells to express interleukin-1beta (IL-1beta), a proinflammatory cytokine implicated in the regulation of tissue inflammation. In this study, we explored the intracellular mechanisms responsible for ECM induction of IL-1beta using human promonocytic U937 cells transfected with the full-length human IL-1beta gene promoter connected to a reporter gene. Using this system, we demonstrated that the ECM molecules fibronectin (FN), type I collagen (Coll), fibrin (Fb) and laminin (Lm) induced transcription of the IL-1beta gene (which was associated with a modest increase in IL-1beta protein secretion) in suspended cells, when used in their soluble monomeric form. This effect was mimicked or blocked by anti-integrin monoclonal antibodies (mAbs) and was dependent on the activation of protein kinase C (PKC). Three of the ECMs tested (FN, Coll and Fb) induced the activation of mitogen-activated protein kinases (MAPKs), whereas Lm had no effect. FN, Coll and Fb (but not Lm) also induced DNA binding of the transcription factor activator protein-1 (AP-1), but not that of nuclear factor-kappaB. Co transfection of U937 cells with a competing AP-1 oligomer blocked the IL-1beta response induced by FN, but not that induced by the other ECMs. By inhibiting AP 1 translocation, glucocorticoids also blocked the FN-induced response, but not that of the other ECMs. These studies suggest that the signalling pathways which mediate ECM induction of IL-1beta expression in human monocytic cells converge at the level of PKC activation. However, they diverge in other aspects, as demonstrated by the differential activation of MAPKs and the need for diverse transcription factors. PMID- 10540222 TI - Selective correlation of interferon-gamma, tumour necrosis factor-alpha and granulocyte-macrophage colony-stimulating factor with immunoglobulin G1 and immunoglobulin G3 subclass antibody in leprosy. AB - Dysregulation of both B- and T-cell responses is observed in leprosy. Immunoglobulin G1 (IgG1) and IgG3 antibody subclasses are selectively elevated towards the lepromatous or disseminated form of the disease accompanied by a depression of T-cell responses. T-cell and macrophage cytokines influence antibody class switching, differentiation and proliferation of B cells. To understand the dynamic nature of the immune response in leprosy, we examined the relationship between circulating Mycobacterium leprae-specific antibodies and secreted cytokines [interferon-gamma (IFN-gamma), interleukin-2 (IL-2), IL-5, IL 10, IL-6, tumour necrosis factor-alpha (TNF-alpha) and granulocyte-macrophage colony-stimulating factor (GM-CSF)] in leprosy patients (19 lepromatous patients; 25 tuberculoid patients) and their exposed household contacts (HC=14) in response to M. leprae antigens. Paired comparison revealed a highly significant negative correlation between IFN-gamma and IgG (P=0.016), IgG1 (P<0.001) and IgG3 (P=0. 007) antibodies. No significant relationship was observed with other T-cell cytokines (IL-2, IL-5 and IL-10). These results strongly suggest that IFN-gamma may play a role in down-regulating antigen-specific IgG1 and IgG3 antibodies. Among the macrophage cytokines, TNF-alpha and GM-CSF which have not been shown to play a role in B-cell activation were positively associated with IgG1 (TNF-alpha, P=0.0005; GM-CSF, P=0.001) and IgG3 (TNF-alpha, P=0.001; GM-CSF, P=0.021) antibodies. Since macrophages have high-affinity Fc receptors for IgG1 and IgG3, it is possible that antigen uptake via these receptors may influence cytokine expression of TNF-alpha, a key modulator of disease pathogenesis in mycobacterial diseases. We are currently investigating the role of Fc receptors on activated macrophages, in expression of pro-inflammatory cytokines in mycobacterial diseases. PMID- 10540223 TI - Expression and translocation of Rac2 in eosinophils during superoxide generation. AB - Eosinophils induce tissue injury by releasing granule-associated cytotoxic proteins, lipid mediators and superoxide anions in response to appropriate stimuli. Superoxide generation associated with respiratory burst is largely dependent on the assembly of the NADPH oxidase complex in the membrane, consisting of membrane-bound cytochrome b558 and translocated p47phox and p67phox. The activation of this complex is critically dependent on the translocation of GTP-bound Rac1, or its homologue Rac2, from the cytosol to the membrane in neutrophils. Rac expression has not yet been fully characterized in eosinophils. We proposed that eosinophils translate and express Rac2 and its GDP dissociation inhibitor, RhoGDI. Furthermore, we hypothesized that Rac2 translocates along with p47phox and p67phox proteins from the cytosol to the plasma membrane during respiratory burst. By reverse transcription-polymerase chain reaction analysis and sequencing of the amplified product, guinea-pig eosinophils were found to express Rac2 mRNA, exhibiting 93% homology with the human Rac2 sequence. Rac1 mRNA was also detected in eosinophils but not its translated product. In contrast, Rac2 protein expression was detected using a specific antibody. In subcellular fractions, Rac2 was found to translocate, along with p47phox and p67phox, from cytosol to plasma membrane-associated fractions following phorbol myristate acetate stimulation, while RhoGDI remained within cytosolic fractions. These findings suggest that Rac2 is preferentially expressed and activated in eosinophils, and is likely to be a crucial regulator of the release of reactive oxygen species from these cells during inflammatory reactions. PMID- 10540224 TI - Adhesion of human mast cells to extracellular matrix provides a co-stimulatory signal for cytokine production. AB - Engagement of integrin receptors during cell adhesion leads to changes in the morphology and the state of activation of cells. We therefore examined whether mast cell adhesion to extracellular matrix proteins affects the synthesis and release of various proinflammatory cytokines. Cells of the human mast cell line HMC-1 were added to fibronectin (FN)-, vitronectin (VN)- or, as a control, bovine serum albumin (BSA)-coated wells and were stimulated with phorbol 12-myristate 13 acetate (PMA) and/or calcium ionophore A23187 (ionophore). Cytokine production was evaluated using semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis of cell extracts and enzyme-linked immunosorbent assay (ELISA) analysis of cell supernatants. After a 4-hr incubation, mRNA expression of interleukin (IL)-8 (and weakly of IL-6) was up-regulated in matrix-adherent cells, with further increase in the presence of PMA and/or ionophore, compared with unstimulated cells. High-level de novo expression of IL-3 and of granulocyte macrophage colony-stimulating factor (GM-CSF) was observed mainly in matrix adherent cells. These changes were paralleled by the secretory pattern of HMC-1 cells after a 24-hr stimulation. Unstimulated cells adherent to FN or VN had already released small amounts of IL-8, and both VN- and FN-adherent cells produced, almost invariably, a higher level of cytokines than BSA-exposed cells after additional stimulation. These results show that mast cell adhesion to matrix proteins by itself has only selected and minor effects, but additional activation of mast cells by secretory stimuli causes significantly enhanced cytokine gene expression and secretion, suggesting that mast cells are far more active in their natural tissue environment than hitherto suggested from data in suspension cultures. PMID- 10540225 TI - Regulation of VH gene repertoire and somatic mutation in germinal centre B cells by passively administered antibody. AB - Immunization with T-dependent antigens induces a rapid differentiation of B cells to plasmacytes that produce the primary immunoglobulin M (IgM) and IgG antibodies with low affinities for the immunogen. It is proposed that the IgG antibody forms immune complexes with the residual antigen which provide an important stimulus for the formation of germinal centres (GC) and the activation of somatic mutation. This hypothesis was tested by passive administration of hapten-specific antibody into mice shortly after the immunization with nitrophenyl (NP) coupled to chicken gamma globulin (NP-CGG) in an environment of limited T-cell help. Athymic mice that received normal T helper cells at 72 hr after the administration of antigen produced low levels of anti-NP antibody and the splenic GC formation was delayed until day 12 after the antigen administration. The analysis of VDJ segments from NP-reactive GC B cells showed very few mutations in the VH genes. Passive injection of anti-NP IgG1 monoclonal antibody - but, not IgM - stimulated the GC formation up to normal levels and the somatic mutation activity in the GC B cells was fully restored. In addition, GC B cells in the recipients of IgG1 antibody demonstrated a change in the usage of germline encoded VH genes which was not apparent among the primary antibody-forming cells. These results suggest the existence of a specific feedback mechanism whereby the IgG antibody regulates the GC formation, clonotypic repertoire and somatic mutation in GC B cells. PMID- 10540226 TI - The preventive effects of incomplete Freund's adjuvant and other vehicles on the development of adjuvant-induced arthritis in Lewis rats. AB - The present study showed a novel finding that the development of adjuvant-induced arthritis (AA) in Lewis rats was completely prevented by incomplete Freund's adjuvant (IFA) injected 21 or 28 days before complete Freund's adjuvant (CFA) challenge. Hexadecane also completely prevented AA and squalane, methyl oleate and pristane moderately prevented AA, though pristane by itself induced mild arthritis in two out of five rats. Concanavalin A-stimulated lymph node cells (LNCs) isolated from AA rats were able to adoptively transfer the severe polyarthritis to all the naive recipients or even to the IFA pretreated recipients with earlier onset and more rapid progression than those of AA. The LNCs from the donors who had been pretreated with IFA and subsequently challenged with CFA could induce mild arthritis in only two out of eight naive recipients, whereas all the recipients who were challenged with CFA immediately after intravenous injection of these LNCs developed significantly less severe arthritis. However, the LNCs from IFA-pretreated donors failed to prevent AA. According to the T helper type 1 (Th1)/Th2 paradigm, it was suggested that the adjuvant-active vehicles such as IFA, hexadecane, squalane, methyl oleate and pristane, can affect and deviate the Th1/Th2 balance of immune responses in host. CFA could promote the propagation of Th2 cells rather than Th1 cells in these vehicle-pretreated rats through as yet undetermined mechanisms, eventually resulting in the prevention of AA. Finally, we discussed a regulatory role of adjuvant vehicles for induction and suppression of AA. PMID- 10540227 TI - T-cell recognition of lipid peroxidation products breaks tolerance to self proteins. AB - Peroxidation of polyunsaturated fatty acids in lipoproteins and cell membrane phospholipids occurs in many situations in the body, both under normal and pathological conditions. Low-density lipoprotein is particularly prone to oxidation and is believed to be a pathogenetic component in atherogenesis. Both antibody responses and T-cell responses to oxidatively modified lipoproteins have been demonstrated in humans as well as in animal models. However, little is known about how these responses arise or how T cells recognize these antigens. In the present study, mice were immunized with homologous albumin covalently modified with a series of defined aldehydes which are known to be generated during lipid peroxidation. T-cell hybridomas from immunized animals demonstrated major histocompatibility complex-restricted and protein sequence-dependent responses to modified albumin, but not to native albumin. In addition to the response to modified epitopes, some aldehyde modifications resulted in strong antibody responses also to the non-modified protein. This T-cell-dependent break of tolerance constitutes a novel pathway for induction of autoimmunity by lipid peroxidation. The findings have implications in many situations where lipid peroxidation products are generated, including atherosclerosis and inflammatory and infectious diseases. PMID- 10540228 TI - Inhibitory effects of endogenous and exogenous interferon-gamma on bronchial hyperresponsiveness, allergic inflammation and T-helper 2 cytokines in Brown Norway rats. AB - Interferon-gamma (IFN-gamma) is an important cytokine involved in the regulation of allergen-induced immune responses. We examined the role of IFN-gamma in a Brown-Norway rat model of bronchial hyperresponsiveness (BHR) and airway eosinophilia, and its effects on the mRNA expression of T helper type 1 (Th1)/Th2 cytokine. Ovalbumin (OA)-sensitized animals were given either exogenous IFN-gamma (105 U/rat over 3 days, intraperitoneally) or anti-IFN-gamma blocking antibody (DB-1 0.3 mg/rat, intravenously) prior to exposure to OA aerosol and were studied 18-24 hr later. In sensitized animals, OA induced significant BHR, accumulation of eosinophils, T lymphocytes and neutrophils in bronchoalveolar lavage (BAL) fluid, and also increased eosinophils and CD8+ T cells in the airways. Exogenous IFN-gamma attenuated allergen-induced BHR (P<0.02, compared with sham-treated animals) together with a significant reduction in eosinophil and neutrophil numbers in BAL fluid (P<0. 005), and eosinophils and CD8+ T cells in airways (P<0.05). By contrast, anti-IFN-gamma antibody increased airway CD4+ T cells and BHR. Using reverse transcriptase-polymerase chain reaction, significant increases in Th2 [interleukin-4 (IL-4), IL-5 and IL-10], and IFN-gamma cytokine mRNA were found in the lungs of sensitized and OA-exposed animals, while exogenous IFN gamma significantly suppressed IL-4, IL-5 and IL-10 mRNA expression, and anti-IFN gamma antibody increased IL-4 and IL-5 mRNA expression. These results indicate that Th1 effects, such as those mediated by IFN-gamma, play a down-regulatory role to suppress the Th2 responses associated with allergen-induced BHR and eosinophilic inflammation. PMID- 10540229 TI - Lipopolysaccharide-dependent down-regulation of CD27 expression on T cells activated with superantigen. AB - To investigate the mechanisms underlying T-cell responses during superantigen (SAg) stimulation, we analysed the effects of SAg on CD27 expression with or without lipopolysaccharide (LPS) as a novel regulator of T-cell function. CD27 is expressed on the majority of resting peripheral blood T cells (CD27low). Activation of T cells by SAg induces high levels of CD27 surface expression (CD27high) accompanied with simultaneous CD30 receptor expression. After prolonged activation in vitro, the level of CD27 expression became intermediate. The effects of LPS on down-regulation of CD27high expression on CD30+ T cells were dose-dependent. Separating LPS-stimulated monocytes from T cells by mechanical dispersion abolished its inhibitory effect, indicating the requirement for direct interactions between monocytes and T cells. We also found that SAg up regulated CD80 expression on CD14+ monocytes and LPS inhibited SAg-induced CD80 expression after 24 hr of stimulation. Up-regulation of CD152 (CTLA-4) was selective, since it was found to be preferentially expressed on the CD30+ population. Competitive experiments using soluble blocking peptides showed that addition of CD28 or CD80 peptide recovered LPS-induced down-regulation of CD27high expression on CD30+ T cells. These observations suggested that the presence of low levels of CD80 on monocytes may partially inhibit CD27 expression due to inefficient delivery of positive signals via CD28/CD80 interaction, and that the increased levels of CD80 enhance the inhibition through interactions with CD152 which is expressed at the highest levels after 48 hr of activation. PMID- 10540230 TI - Cell surface expression and metabolism of major histocompatibility complex class II invariant chain (CD74) by diverse cell lines. AB - We previously described the processing of antibodies to CD74 (the major histocompatibility complex class II-associated invariant chain, Ii), by B-cell lymphoma cell lines. These cells expressed relatively low levels of Ii on the surface, but the molecules were rapidly internalized and replaced by new molecules, so that approximately 8 x 10(6) antibody molecules per cell were taken up per day. We herein report the results of similar studies with other cell types, namely a melanoma, a colon carcinoma, a T-cell lymphoma and B lymphoblastoid cell lines. The melanoma and the carcinoma were treated with interferon-gamma to induce high levels of the antigen. The T-cell lymphoma, HUT 78, was selected specifically because it was previously reported to lack cell surface Ii, while expressing the molecule intracellularly. However, HUT 78 displayed Ii on the cell surface, as did the other cell lines tested, and catabolism of the antibody was very fast on all of the cell lines. The capacity of four of the cell lines for cumulative antibody uptake was evaluated, using 'residualizing' radiolabels, which are trapped within the cell after catabolism of the antibody to which they were conjugated. A high level of uptake was observed in all cases, although there was significant variation between the cell lines. With melanoma SK-MEL-37, the total LL1 uptake in 24 hr was nearly 10(7) molecules per cell and the average turnover time for Ii on the cell surface was 4 min; with carcinoma HT-29, the total LL1 uptake in 24 hr was approximately 10(6) molecules per cell, and the average turnover time for Ii on the cell surface was 27 min. Based on the cell content of mature class II antigens (alphabeta), these data suggest that a large fraction, or all, of immature class II molecules (alphabetaIi) reach the cell surface before entering the peptide-loading compartment, independent of the particular cell type. PMID- 10540231 TI - Structural characterization of mouse CD97 and study of its specific interaction with the murine decay-accelerating factor (DAF, CD55). AB - CD97 is a newly identified, activation-associated human leucocyte antigen with seven putative transmembrane domains. It has an extended extracellular segment containing several adhesion molecule structure motifs, and has been shown to interact with the human complement regulator, decay-accelerating factor (DAF, CD55). To understand further the interaction between CD97 and DAF, as well as the structure and function of CD97 in general, we have cloned the mouse CD97 cDNA and studied the encoded protein for its membrane association property and ability to interact specifically with the murine decay-accelerating factor. The full-length mouse CD97 cDNA that we have cloned and characterized encodes a protein that is 60% identical to the three epidermal growth factor (EGF) domain-containing form of human CD97 but does not contain the Arg-Gly-Asp (RGD) motif which is present in human CD97. Two other alternatively spliced forms of mouse CD97 were also identified. These forms differ by the number of EGF-like sequence repeats present in the N-terminal region. Northern blot analysis revealed that CD97 is expressed widely in mouse tissues and in resting as well as activated cultured mouse splenocytes. Transient transfection of human embryonic kidney (HEK) 293 cells with the mouse CD97 cDNA in a green-fluorescence protein vector (pEGFP-N1) showed plasma membrane targeting of the expressed protein. Western blot analysis confirmed its membrane association and identified the existence of a processed C terminal fragment, supporting the notion that CD97 on the cell membrane is composed of post-translationally generated subunits. Adhesion studies demonstrated that normal, but not DAF knockout mouse erythrocytes and splenocytes adhered to mouse CD97-transfected HEK cells. The interaction of CD97 and DAF was found to be species-restrictive in that human erythrocytes were unable to bind to mouse CD97-transfected HEK cells. These results indicate that the general structure, membrane association property and DAF-binding ability of CD97 are conserved and that the adhesive interaction between CD97 and DAF is independent of the RGD motif. The finding that CD97 is distributed widely among various mouse tissues suggests that CD97 may have other roles beyond lymphocyte activation. PMID- 10540232 TI - Comparative responses of Pseudomonas stutzeri and Pseudomonas aeruginosa to antibacterial agents. AB - The sensitivity of six strains of Pseudomonas stutzeri (NCIMB 568, 10783, 11358, 11359, JM 302, JM 375) to cationic antiseptics, mercury compounds, the parabens, phenolics, EDTA and various antibiotics was compared with Pseudomonas aeruginosa NCIMB 8626. All Ps. stutzeri strains were highly sensitive to chlorhexidine diacetate, organomercurials and triclosan, but rather less so to quarternary ammonium compounds (QACs). They were also sensitive to other biocidal agents and more sensitive to many antibiotics than the strain of Ps. aeruginosa. There was little correlation between uptake of chlorhexidine diacetate or cetylpyridinium chloride by dense suspensions of organisms, leakage of intracellular constituents and loss of cell viability. PMID- 10540233 TI - Phospholipid molecular species distribution of some medically important Candida species analysed by fast atom bombardment mass spectroscopy. AB - The aim of this study was to obtain detailed information on phospholipids (PL) of the medically important Candida species and to determine their possible chemotaxonomic significance. Lipids were extracted from 22 strains representing 8 Candida species and their PL molecular species distributions were determined by Fast Atom Bombardment Mass Spectroscopy (FAB MS) in negative ion mode. Fifteen major lower mass peaks (m/z 221 to 289) were attributable to the expected presence of carboxylate anions and 24 major higher mass peaks (m/z 557 to 837) were attributable to phospholipid anions. Major carboxylate peaks were of the following m/z and identities : 253, C16:1; 255, C16:0; 277, C18:3; 279, C18:2; 281, C18:1; and 283, C18:0. The most abundant peaks consistent with the presence of phospholipid molecular species anions include those of m/z 673, 743, 833, 834 and 836 tentatively identified as phosphatidic acid (PA) (34:1), phosphatidylglycerol (PG) (34:3), phosphtidylinositol (PI) (34:2) and two unknown molecular species. This profile is diagnostic for the genus Candida. Quantitative differences were observed between different Candida species. Thus, polar lipid molecular species distribution in Candida spp. has chemotaxonomic significance, especially so in the case of carboxylate anions. PMID- 10540235 TI - Bile salt deconjugation by lactobacillus plantarum 80 and its implication for bacterial toxicity AB - The effects of bile salts on the survival of lactobacilli were investigated using glycocholic acid, cholic acid and deoxycholic acid as model compounds and the bile salt hydrolase active Lactobacillus plantarum 80 (BSH+) and its BSH negative mutant. The detrimental effects of cholic acid, i.e. growth inhibition and cytotoxicity at a concentration of 1 and 5 mmol l-1, respectively, were considered to be due to the hydrophobic protonated form of the molecule, which brings about membrane damage. The conversion of glycocholic acid to cholic acid by the BSH active L. plantarum 80 caused a growth inhibition which was comparable with the inhibition observed in the broth supplemented with 1 mmol l-1 cholic acid. Deoxycholic acid caused toxicity through membrane damage when the compound was in solution. Its toxicity disappeared in the culture broth as the molecule precipitated. In case of cholic acid, the toxicity could be removed by buffering the solution at pH 7.0. It was calculated that at this pH most of the cholic acid molecules were ionized. The results led to the formulation of an extended hypothesis about the ecological significance of bile salt transformations. Primary deconjugation is carried out to counteract intracellular acidification. Yet, the deconjugated molecule can be harmful at moderately acidic pH-values. In this case, the BSH+ strains could effectively profit from their activity in case they are associated with 7alpha-dehydroxylating bacteria which dehydroxylate the deconjugated bile salts. The dehydroxylated molecule has a low solubility and precipitates at moderately acidic pH. PMID- 10540234 TI - Combined use of acetic acid treatment and modified atmosphere packaging for extending the shelf-life of chilled chicken breast portions. AB - Samples of chicken breasts with skin were treated with a 1% acetic acid solution or untreated and packaged in a 70% CO2/30% N2 modified atmosphere. Two different types of films were studied to establish their usefulness within the above pre determined conditions. After 3, 7, 14 and 21 d of storage at 4 degrees C, the samples were evaluated for spoilage microbial growth, odour and slime, as well as the gas composition in the headspace volume in the package. As a result of this, it was found that both films were adequate for using them as barriers. Samples treated with the acetic acid solution smelt slightly acidic and pleasant, while the untreated ones had 'off' odours at the end of the storage periods. However, all samples showed acceptable overall aspect by that time. Acetic acid treatment produced decreases in counts in all genera studied. Results of this study indicate that using modified atmosphere packaging (MAP) on chicken breasts previously decontaminated with acetic acid is a worthwhile technology to extend samples shelf-life. PMID- 10540236 TI - Effects of cultural conditions on denitrification by pseudomonas stutzeri measured by membrane inlet mass spectrometry AB - Denitrification is a globally important process leading to loss of fertiliser efficiency and the production of the greenhouse gas nitrous oxide and nitric oxide, an ozone depleter. Membrane inlet mass spectrometry (MIMS) was employed to study the effect of different variables on the process of denitrification by Pseudomonas stutzeri in a defined salts medium. MIMS was used for concomitant measurements of nitrous oxide, nitrogen and oxygen and showed that denitrification occurred in the presence of dissolved oxygen. A nitrate concentration of 15 mmol l-1 and a nitrite concentration of 5 mmol l-1 were found to be optimum for complete denitrification of nitrate or nitrite to nitrogen and varying these concentrations had a marked effect on the ratio of gaseous products released. Denitrification products were also dependant on pH with neutral or alkaline conditions being best for production of gaseous end products. Our results suggest that under nutrient rich conditions the most important factor in the regulation of denitrification by Ps. stutzeri is the amount of nitrite generated at the first enzymatic stage of the process. This appears to cause inhibition of the denitrification pathway above 5 mmol l-1 and at high enough concentrations (15 mmol l-1) restricts growth. PMID- 10540238 TI - Hemicellulase activity of antarctic microfungi AB - The mannanase (endo-beta-1,4-mannanase; E.C. 3.2.1.78) and xylanase (endo-beta 1,4-xylanase; E.C. 3.2.1.8) activity of five microfungal isolates from Antarctica were characterized at different temperatures and pH. In general, the hemicellulase activity of the antarctic strains occurred at least 10 degrees C and as much as 30 degrees C lower than that of a mesophilic reference strain. At 0 degrees C, two strains, a Phoma and a Penicillium, produced in excess of 40% of their measured maximum activity of mannanase. All strains had maximum hemicellulase activity in the range pH 4-5, with Penicillium, Phoma and Alternaria strains exhibiting high (in excess of 80% of maximum) mannanase activity at pH 10. Three of the antarctic isolates exhibited high levels of xylanase activity over a pH range of 3-11. PMID- 10540237 TI - The influence of hay-packing techniques on the presence of saccharopolyspora rectivirgula. AB - Environmental factors influencing the growth, distribution and viability of thermophilic actinomycetes, especially Saccharopolyspora rectivirgula as an agent of extrinsic allergic alveolitis in farms workers, were studied. Total microbial count, eumycetes and thermophilic actinomycetes were determined on 96 hay samples, randomly collected, from small prismatic and large cylindrical bales, 30 air samples before and after animal feeding, and various surfaces in two farms located in the province of Reggio Emilia, Italy. The number of thermophilic actinomycetes (potentially responsible for hypersensitivity pneumonitis) was higher in hay samples from large cylindrical bales than in those from small prismatic bales. The structural characteristics of the buildings (barns with stalls, poor ventilation) and the feeding practices (manual handling of hay, constant presence of hay in feedings corridors) contributed to the dispersion of high levels of thermophilic actinomycetes spores (potentially responsible for extrinsic allergic alveolitis). The ventilation system proved to be inadequate in reducing the number of microorganisms present. PMID- 10540239 TI - Water activity and temperature effects on germination and growth of Eurotium amstelodami, E. chevalieri and E. herbariorum isolates from bakery products. AB - This study investigated the effect of temperature (5-30 degrees C), water activity (0.775-0.90 aw) and their interactions on the temporal rates of germination and mycelial growth of three species of Eurotium on flour wheat sucrose medium. Germination was quite rapid at aw >0.85, with an almost linear increase with time for all isolates. However, under more extreme water stress, germination was slower. The aw minima for germination were usually lower than those for growth and varied with temperature. The effect of aw x temperature interactions on the lag phases (h) prior to germination and on the germination rates (h-1) were predicted using the Gompertz model modified by Zwietering. Eurotium spp. had shown short lag times at 0.90 aw over a wide range of temperatures. At marginal temperatures, lag phases were significantly longer, especially at >15 degrees C. The temperature x aw profiles for mycelial growth varied between species in terms of rates (mm d(-1)). Predictions of the effect of important environmental factors, such as temperature, aw and their interactions on lag times to germination, germination rates and mycelial growth, are important in the development of hurdle technology approaches to predict fungal spoilage in food products. PMID- 10540240 TI - Characterization of chlorobenzoate degraders isolated from polychlorinated biphenyl-contaminated soil and sediment in the Czech Republic. AB - Two polychlorinated biphenyl-contaminated sites in the Czech Republic, a soil at Zamberk and a sediment sludge at Milevsko, were screened for the presence of chlorobenzoate degraders. Sixteen different chlorobenzoate degraders were isolated from the soil compared with only three strains isolated from the sediment. From these strains, only four soil degraders and one strain isolated from the sediment, respectively, were shown to possess a complete chlorobenzoate (CB) pathway. Bacteria isolated from the soil have expressed more flexibility for CB degradation, namely in the case of ortho-chlorinated benzoates. They all possessed large plasmids, the restriction patterns of which were compared. Plasmids in Pseudomonas sp. A7, A8, A18 and A19, respectively, were cured and found to encode at least part of the metabolic pathway involved in the growth on ortho-chlorinated benzoates. PMID- 10540241 TI - Effect of process temperature, pH and suspended solids content upon pasteurization of a model agricultural waste during thermophilic aerobic digestion. AB - Thermophilic aerobic digestion(TAD), or liquid composting, is a versatile new process for the treatment and stabilization of high strength wastes of liquid or, perhaps more importantly, slurry consistency. The pattern of inactivation of various pathogenic and indicator organisms was studied using batch digestions under conditions that may be expected to be found in full-scale TAD processes. Rapid inactivation of test populations occurred within the first 10 min from the start of digestion. The inactivation rate was slightly lower when digestions were conducted below 60 degrees C. In some instances, a 'tail' was apparent, possibly indicating the survival of relatively resistant sub-populations particularly in the case of Serratia marcescens and Enterococcus faecalis, or of clumping or attachment of cells to particulate materials. The effect of pH on the inactivation of the test populations depended on the temperature of digestion, but varied with the test population. At 55 degrees C Escherichia coli was more sensitive to temperature effects at pH 7 than at pH 8, but was more sensitive at pH 8, 60 degrees C. The reverse was the case at 60 degrees C for Ent. faecalis. An increase in the solid content of the digesting waste caused a progressive increase in the protection of test organisms from thermal inactivation. Challenging a TAD process with test strains allows (via estimation of D-values) a quantification of the cidal effects of such processes, with a view to manipulating process variables to enhance such effects. PMID- 10540242 TI - Heat transfer analysis of staphylococcus aureus on stainless steel with microwave radiation. AB - Staphylococcus aureus (NCTC 6571; Oxford strain) on stainless steel discs was exposed to microwave radiation at 2450 MHz and up to 800 W. Cell viability was reduced as the exposure time increased, with complete bacterial inactivation at 110 s, attaining a temperature of 61.4 degrees C. The low rate of temperature rise, RT, of the bacterial suspension as compared with sterile distilled water or nutrient broth suggests a significant influence of the microwave sterilization efficacy on the thermal properties of the micro-organisms. The heat transfer kinetics of thermal microwave irradiation suggest that the micro-organism has a power density at least 51-fold more than its surrounding liquid suspension. When the inoculum on the stainless steel disc was subjected to microwave radiation, heat conduction from the stainless steel to the inoculum was the cause of bacteriostasis with power absorbed at 23.8 W for stainless steel and 0.16 W for the bacteria-liquid medium. This report shows that the microwave killing pattern of Staph. aureus on stainless steel was mainly due to heat transfer from the stainless steel substrate and very little direct energy was absorbed from the microwaves. PMID- 10540243 TI - Study of the potential relationship between the morphology of infectious somatic coliphages and their persistence in the environment. AB - The proportions of different morphological types of infectious somatic coliphages were determined in faecally polluted freshwaters. Myoviridae, followed by Siphoviridae, were the most frequently isolated morphological types in raw sewage, treated sewage and river water collected a few metres downstream from a sewage outfall. However, in river water collected further downstream from the pollution point, in river water after 'in situ' inactivation experiments and in chlorinated raw and treated sewage significant changes in the proportions of the different somatic coliphage morphological types occurred. In all cases, Siphoviridae, especially those with flexible and curled tails, became more abundant to the detriment of Myoviridae. PMID- 10540244 TI - Reduced toxicity of expression, in Escherichia coli, of antipollutant antibody fragments and their use as sensitive diagnostic molecules. AB - Single-chain antibody fragments (scAb), specific for the chlorophenoxy acid herbicide mecoprop, have been expressed and purified from the bacterium Escherichia coli. Co-expression with the colE1-compatible, arabinose-inducible, skp expression vector pHELP1 prevented bacterial lysis and significantly increased both total and functional expression yield. The periplasmic protein, SKP, may have a role as a generic detoxification protein. Surface plasmon resonance (BIAcore 2000) analysis confirmed that the purified scAb retained similar binding kinetics to the monoclonal antibody (Mab) from which it was cloned. In competition ELISA, the bacterial scAb showed the same specificity for mecoprop and a related herbicide, MCPA, as the Mab but an increase in sensitivity for free antigen in all ELISA formats. Bacterially expressed antibody fragments provide a simple, sensitive and cost-effective alternative to the traditional production of diagnostic Mabs via tissue culture. PMID- 10540245 TI - Detection of low numbers of Salmonella in environmental water, sewage and food samples by a nested polymerase chain reaction assay. AB - A polymerase chain reaction (PCR) assay with two nested pairs of primers selected from conserved sequences within a 2.3 kb randomly cloned DNA fragment from the Salmonella typhimurium chromosome was developed. The nested PCR assay correctly identified 128 of a total of 129 Salmonella strains belonging to subspecies I, II, IIIb and IV. One strain of Salm. arizona (ssp. IIIa) tested negative. No PCR products were obtained from any of the 31 non-Salmonella strains examined. The sensitivity of the assay was 2 cfu, as determined by analysis of proteinase K treated boiled lysates of Salm. typhimurium. The performance of the assay was evaluated for environmental water, sewage and food samples spiked with Salm. typhimurium. Water and sewage samples were filtered and filters were enriched overnight in a non-selective medium. Prior to PCR, the broth cultures were subjected to a rapid and simple preparation procedure consisting of centrifugation, proteinase K treatment and boiling. This assay enabled detection of 10 cfu 100 ml(-1) water with background levels of up to 8700 heterotrophic organisms ml(-1) and 10000 cfu of coliform organisms 100 ml(-1) water. Spiked food samples were analysed with and without overnight enrichment in a non selective medium using the same assay as above. Nested PCR performed on enriched broths enabled detection of <10 cfu g(-1) food. Variable results were obtained for food samples examined without prior enrichment and most results were negative. This rapid and simple assay provides a sensitive and specific means of screening drinking water or environmental water samples, as well as food samples, for the presence of Salmonella spp. PMID- 10540246 TI - Effect of protective solutes on leakage from and survival of immobilized lactobacillus subjected to drying, storage and rehydration AB - When lactic acid bacteria are used industrially as fermentation starters it is important to obtain stable and highly viable bacterial cultures. Six strains of Lactobacillus encapsulated in Ca-alginate gel beads were investigated to determine whether dehydration, storage and rehydration may inflict injury. A negative relationship between leakage of lactate dehydrogenase and survival rates was found. Mesophilic lactobacilli showed only negligible leakage compared with thermophilic strains when dehydrated at 30 degrees C to a level of 0.11 g H20 (g dry wt)-1. The choice of an appropriate suspending medium to be introduced before drying was therefore very important for thermophilic lactobacilli in order to increase the survival rates during dehydration, storage and rehydration. The osmoregulatory solutes tested were adonitol, betaine, glycerol and reconstituted non-fat milk solids (NFMS). Less injury was inflected during dehydration for Lactobacillus helveticus with adonitol, glycerol and NFMS. Survival rates for the strains subjected to immobilization, dehydration, storage and rehydration varied with the strain and the protective solute when fluidized-bed drying was used at 5 degrees C to a level as high as 0.34 g H20 (g dry wt)-1. Non-fat milk solids gave the best protection for thermophilic lactobacilli, while adonitol and NFMS were best for mesophilic lactobacilli. PMID- 10540247 TI - Nisin, temperature and pH effects on growth and viability of pectinatus frisingensis, a gram-negative, strictly anaerobic beer-spoilage bacterium AB - Pectinatus frisingensis, a Gram-negative and strictly anaerobic beer spoilage bacterium is sensitive to nisin. An increase in nisin concentration (0 to 1100 IU ml-1) added to the culture medium prolonged the lag phase, and decreased the growth rate of the bacterium. In addition, late exponential cells of P. frisingensis exposed to low concentrations of nisin lost immediately a part of their intracellular K+. Presence of Mg2+ up to 15 mmol l-1 did not protect P. frisingensis from nisin-induced loss of viability and K+ efflux. Potassium leaks were also measured in P. frisingensis late exponential phase cells exposed to combined effects of nisin addition (100-500 IU ml-1), 10 min mild heat-treatment (50 degrees C) or rapid cooling (2 degrees C), and pH (4.0 and 6.2). Net K+ efflux from both starving and glucose-metabolizing cells, was more important at pH 6.2, whatever the temperature treatment and nisin addition. Reincubation at 30 degrees C of P. frisingensis glucose-metabolizing cells exposed to a preliminary combination of nisin addition and mild heat or cooling down treatment, showed that cells exposed to rapid cooling reaccumulated more K+ than heat-treated cells, whatever the pH conditions. A combination of nisin and mild heat-treatment could thus be of interest to prevent P. frisingensis growth in beers. PMID- 10540249 TI - Ecological basis for biocontrol of damping-off disease by pseudomonas fluorescens 54/96 AB - Pseudomonas fluorescens 54/96, originally isolated from the rhizosphere of sugar beet, has been shown to be commercially effective in field trials for the suppression of a number of fungal diseases of seedlings. In vitro and microcosm based assays revealed that both the timing and method of application of bacteria were important for effective control of Pythium ultimum, the causative agent of damping-off disease. Following transposon mutagenesis (Tn5lac), mutants deficient for the suppression of Pythium ultimum infections of peas were isolated. Three major classes of insertional mutants of Ps. fluorescens 54/96 were identified which either inhibited sporulation, reduced mycelial growth or affected the regulation of bacterial metabolic activity. Evaluation of the metabolic capability of pathogen and antagonist revealed evidence for direct competition, as both the fungus and bacterium had similar sole carbon source nutrient utilization profiles. Further comparisons of the activity of the transposon mutants indicated that although the mechanisms of disease control were multifactorial, the most significant factor was the prevention of rapid spore germination in the presence of pea seeds. PMID- 10540248 TI - Recovery and assay of African swine fever and swine vesicular disease viruses from pig slurry. AB - Assaying samples for infectious virus is more difficult when the sample is toxic to cells used in the assay, e.g. with samples of infected pig slurry. Various techniques were compared for the recovery of African swine fever virus (ASFV) and swine vesicular disease virus (SVDV) in pig slurry. Extraction with Freon led to 80-100% recovery of SVDV added to pig slurry. The assay sensitivity enabled undiluted, centrifuged sample to be put directly onto monolayers of IB-RS2 cells, allowing a minimum detection level of 100.7 pfu ml-1. ASFV was difficult to recover intact, and the best technique allowed a recovery of 60% with a minimum detectable level of 101.8 HAD50 ml-1, due to toxicity to the cells at low sample dilutions. Extraction with the addition of an equal volume of ox serum to inoculated slurry was best at recovering ASFV. Poor recoveries with the other techniques may have been due to the inactivation of the virus while in the slurry rather than as a result of the inability of the method to extract ASFV. PMID- 10540250 TI - AFM tips: how sharp are they? AB - From both simple estimates and a 'blind' reconstruction based on cryo-AFM images of filamentous actin, we find that the radius of curvature at the apex of Si3N4 tips can be as small as 1 nm with a conical angle in the range 30 approximately 40 degrees, revealing a relatively high aspect ratio that is much greater than previously anticipated. Our results show that commercially available cantilevers are often sharp enough for routine high resolution imaging of biological materials, and suggest that factors other than an inherent blunt tip are probably responsible for frequent occurrences of poor resolution. PMID- 10540251 TI - Possible roles of extracellular matrix and cytoskeleton in leech body wall muscles. AB - Round circomyarian fibres of leeches are peculiar helical muscles. The fibres are characterized by a lack of junctions, being separated by a thick extracellular matrix, and by scarce end-plates. Even so, the fibres grouped in units show the same degree of contraction. Biochemical, immunocytochemical and ultrastructural studies were performed in order: (a) to demonstrate the presence in the extracellular matrix of fibronectin, collagen type IV and laminin and in the cytoskeleton of desmin and alpha-actinin; (b) to show the possible link of extracellular matrix with the scaffold of intermediate filaments; (c) to evaluate how the extracellular matrix can play a role in the transduction of a signal during contraction-relaxation-superelongation phases. PMID- 10540253 TI - Signal components in the environmental scanning electron microscope. AB - We demonstrate that the gas-amplified secondary electron signal obtained in the environmental scanning electron microscope has both desired and spurious components. In order to isolate the contributions of backscattered and secondary electrons, two sets of samples were examined. One sample consisted of a pair of materials having similar secondary emission coefficients but different backscatter coefficients, while the other sample had a pair with similar backscatter but different secondary emission coefficients. Our results show how the contribution of the two electron signals varies according to the pressure of the amplifying gas. Backscatter contributions, as well as background due to gas ionization from the primary beam, become significant at higher pressure. Furthermore, we demonstrate that the relative amplification efficiencies of various electron signals are dependent upon the chemistry of the gas. PMID- 10540252 TI - A procedure to prepare cultured cells in suspension for electron probe X-ray microanalysis: application to scanning and transmission electron microscopy. AB - We describe a simple procedure to prepare cultured cells in suspension to analyse elemental content at the cellular level by electron probe X-ray microanalysis. Cells cultured in suspension were deposited onto polycarbonate tissue, culture plate well inserts, centrifuged at low g, washed to remove the extracellular medium, cryofixed and freeze-dried, and analysed in the scanning mode of a scanning electron microscope. We tested the effect of different washing solutions (150 mM ammonium acetate, 300 mM sucrose, and distilled water) on the elemental content of cultured cells in suspension. The results demonstrated that distilled water was the best washing solution to prepare cultured cells. In addition, the low Na content, high K content and high K/Na ratio of the cells indicated that this procedure, based on the centrifugation at low g followed by cryopreparation, constitutes a satisfactory method to prepare cultured cells in suspension. We also investigated the effects of different accelerating voltages on X-ray signal collection. The results showed that moderate accelerating voltages, i.e. 10-11 kV, should be used to analyse whole cells in the scanning mode of the scanning electron microscope. We show that this method of preparation makes it possible to prepare cryosections of the cultured cells, thus permitting analysis of the elemental content at the subcellular level, i.e. nucleus, cytoplasm and mitochondria, using a scanning transmission electron microscope. PMID- 10540254 TI - Cryo-techniques applied to stratum corneum with description of a new sample holder for cryo-scanning electron microscopy of freeze-fractured samples. AB - Stratum corneum structure greatly differs from that of the living epidermis and specific sample cryo-preparation techniques have to be used. Practical aspects of these cryo-techniques applied to stratum corneum are discussed. Emphasis is placed on scanning electron microscopy of cryo-fixed samples. A new sample holder designed for cryo-scanning electron microscopy of freeze-fractured stratum corneum is described. PMID- 10540255 TI - Unexpected property of trehalose as observed by cryo-electron microscopy. AB - Trehalose is an agent useful in maintaining the integrity of many biological systems submitted to various stresses. It is also presumed to improve specimen preparation for electron microscopy and to reduce beam damage. Here we study the effect of trehalose on the preparation and observation by cryo-electron microscopy of thin vitrified films of biological suspensions. We observe that trehalose, as compared to sucrose, can indeed reduce electron beam damage to biological particles, as determined from the dose necessary for the onset of bubbling. Surprisingly, we also find that the contrast of biological particles is higher in a vitrified solution of trehalose than in one of sucrose. This effect can be explained if the water evaporation during the specimen preparation is less in the presence of trehalose than with sucrose, but we do not yet understand the underlying reasons since the evaporation properties of both sugars are similar at a macroscopic level. We conclude that trehalose is truly a remarkable substance and that more investigation is needed in order to fully understand its properties, and that the addition of ca. 3-5% trehalose to biological suspensions is a simple and useful method to reduce commonly arising drying artefacts and water evaporation in the thin film vitrification method. PMID- 10540256 TI - Electron diffraction from micro- and nanoparticles of hydroxyapatite. AB - Hydroxyapatite (HAP) obtained from aqueous solutions under different conditions has been examined by high-resolution transmission electron microscopy (HRTEM) and electron diffraction, including selected-area electron diffraction (SAED) and microdiffraction. A Philips CM300 field-emission gun electron microscope with a Schottky W/ZrO field-emission tip and a spherical aberration constant of 0.65 mm was used at 300 kV. The HAP crystals had different sizes, ranging from a few nanometres to a few micrometres. Single-crystal diffraction patterns have been obtained from the largest microcrystals using the conventional SAED technique. Assemblies of nanoparticles gave only broad diffuse rings. Nevertheless, microdiffraction with electron microprobes 3.5-10 nm in diameter clearly indicated the crystalline character of the nanoparticles in these assemblies. Experimental HRTEM images, Fourier transforms and calculated images exhibited the fine structure of the HAP crystals. PMID- 10540257 TI - Proximal electromagnetic shear forces. AB - We perform a simple model calculation to estimate the electromagnetically induced shear force caused by a current dissipation when a charged tip is moved parallel to a conducting material. For parameters typical in shear force imaging, the force is many orders of magnitude below reported values. Thus, proximal electromagnetic tip-sample forces can be neglected in discussions of shear force imaging. PMID- 10540258 TI - Image cytometric method for quantifying the relative amount of DNA in bacterial nucleoids using Escherichia coli. AB - An image cytometric method for quantifying integrated fluorescence was developed to measure the relative DNA contents of bacterial nucleoids. Image analysis was performed with newly developed macros in combination with the program Object Image, all downloadable from http://simon.bio.uva.nl/object-image.html. Four aspects of the method were investigated. (i) Good linearity was found over a ten fold range of fluorescence intensity in a test with a calibration kit of fluorescent latex spheres. (ii) The accuracy of the method was tested with a narrowly distributed Escherichia coli population, which was obtained by growing cells into stationary phase. The width of the image cytometric distribution was approximately 6%, in good agreement with results obtained by flow cytometry. (iii) The error contribution of manual focusing could be kept below 2%, although a strong dependency between integrated fluorescence and focus position was observed. (iv) The results were verified with a flow cytometer, which gave similar distributions for the DNA contents per cell expressed in chromosome equivalents (4.8 fg of DNA). We used the presented method to evaluate whether the DNA conformation had any effect on the total fluorescence of bacterial nucleoids. Experiments using nucleoids with the same amount of DNA in either a dispersed or a compact conformation showed no significant difference in integrated fluorescence, indicating that it is possible to determine the DNA content per nucleoid independently of the actual organization of the DNA. PMID- 10540259 TI - Pronounced loss of cell nuclei and anisotropic deformation of thick sections. AB - The local deformation and variations in section thickness are studied in 100 microm thick vibratome sections of well-fixed human brain tissue. During processing, including drying on glass slides, the section thickness is reduced to less than half, but close to the edges there is less shrinkage of the section thickness. Close to both surfaces there is a pronounced reduction in the number of neuronal nucleoli. At the scale of the original section, the upper 15 microm and the lower 10 microm are depleted. The loss is most pronounced at the upper surface, which is unprotected during processing. In the central 70% of the section height, where one would ordinarily use an optical disector for sampling, there is no indication of non-uniform shrinkage. The simplest explanation for the observed loss of nucleoli is that all cells opened by the knife may lose their nuclei across an unprotected section surface. The observations do not generalize to other tissues and other preparation techniques, but illustrate the magnitude of some of the problems for uniform sampling and unbiased estimation in very thick sections. The uniform optical disector sampling of nucleoli in thick sections, as opposed to that of cell nuclei, raises a special problem, which is discussed briefly. PMID- 10540261 TI - Introduction PMID- 10540260 TI - An exploration of the accuracy of an invasive method for measuring the volume of specimens of Amoeba proteus. AB - A method is described for measuring the volume of individual specimens of Amoeba proteus which utilizes an easily constructed compressor to flatten the specimen to a known thickness. The microscopic image of the flattened specimen is captured on tape, digitized and analysed with the NIH Image software. The results from one specimen are given to illustrate the sources and magnitude of errors affecting these volume measurements. PMID- 10540262 TI - Collected studies on interfaces and interphases as related to the behaviour of fibre-reinforced aluminium alloy composites AB - This paper is an essentially practical treatment of interphases and interfaces and of their influence on the properties of a number of metal matrix composites (MMCs). The illustrations are drawn from the authors' experiences and have been chosen to underline the importance of detailed microstructural analysis for elucidating the fabrication behaviour and the mechanical performance of this group of materials. The work involves a series of MMCs based upon different combinations of aluminium alloy and ceramic/carbon fibre (both continuous and short) and made using the method of low-pressure liquid metal infiltration (LMI). Detailed analyses of the composite microstructures are given, with particular attention being paid to the interface regions. The data are used to categorize an interface according to the type of bond, that is a mechanical bond resulting from thermal mismatch between the fibre and metal matrix, or a chemical bond, with or without second phase, caused by chemical reaction. The information is then employed to account for aspects of composite fabrication, such as the cast microstructure produced by the LMI method and the effect of heat treatment, and to elucidate composite properties such as stiffness, yield stress and failure strength. PMID- 10540263 TI - Aluminium-aluminium nitride composites fabricated by melt infiltration under pressure AB - Aluminium-matrix composites containing approximately 55 vol.% AlN particles were fabricated by melt infiltration of aluminium into an AlN preform under a pressure of up to 130 MPa. Two different AlN powders (H.C. Starck, Goslar, Germany, and ESK, Elektroschmelzwerk, Kempten, Germany) and four types of aluminium alloy (2024, 1070, 6060 and 5754) were used. The initial AlN powders were characterized by scanning electron microscopy. The composites were studied by light microscopy, scanning and transmission electron microscopies and energy-dispersive X-ray spectroscopy. Particle-matrix interfaces were observed using high-resolution electron microscopy. As a result of the melt infiltration process, the composites are very dense and the microstructure shows a homogeneous distribution of the reinforcement. The interfaces are clean with very little porosity. Some Al2Cu precipitates were observed in the 2024 matrix. PMID- 10540264 TI - Effect of SiC reinforcement on the deformation and fracture micromechanisms of Al Li alloys AB - The effect of SiC reinforcement on the microstructure of a naturally aged 8090 Al alloy as well as on the deformation and fracture micromechanisms was investigated. To this end, the microstructural characteristics (grain and reinforcement morphology, precipitate structure) were determined in the unreinforced alloy and in the composite reinforced with 15 vol.% SiC particles. The materials were tested under monotonic tension and fully reversed cyclic deformation and then carefully analysed through scanning and transmission electron microscopy to find the dominant deformation and failure processes for each material and loading condition. It was found that the dispersion of the SiC particles restrained the formation of elongated grains during extrusion and inhibited the precipitation of Al3Li. As a result, the plastic deformation in the composite was homogeneous, while strain localization in slip bands was observed in the unreinforced alloy specimens tested in tension and in fatigue. The unreinforced alloy failed by transgranular shear along the slip bands during monotonic deformation, whereas fracture was initiated by grain boundary delamination, promoted by the stress concentrations induced by the slip bands, during cyclic deformation. The fracture of the composite was precipitated by the progressive fracture of the SiC reinforcements during monotonic and cyclic deformation. PMID- 10540265 TI - Scanning and transmission electron microscopy study of the microstructural changes occurring in aluminium matrix composites reinforced with SiC particles during casting and welding: interface reactions AB - Processing of aluminium matrix composites (AMCs), especially those constituted by a reactive system such as Al-SiC, presents great difficulties which limit their potential applications. The interface reactivity between SiC and molten Al generates an aluminium carbide which degrades the composite properties. Scanning and transmission electron microscopes equipped with energy-dispersive X-ray spectroscopes are essential tools for determining the structure and chemistry of the Al-SiC interfaces in AMCs and changes occurring during casting and arc welding. In the present work, an aluminium-copper alloy (AA2014) reinforced with three different percentages of SiC particles was subjected to controlled remelting tests, at temperatures in the range 750-900 degrees C for 10 and 30 min. Arc welding tests using a tungsten intert gas with power inputs in the range 850-2000 W were also carried out. The results of these studies showed that during remelting there is preferential SiC particle consumption with formation of Al4C3 by interface reaction between the solid SiC particle and the molten aluminium matrix. The formation of Al4C3 by the same mechanism has also been detected in molten pools of arc welded composites. However, in this case there was formation of an almost continuous layer of Al4C3, which protects the particle against further consumption, and formation of aciculate aluminium carbide on the top weld. Both are formed by fusion and dissolution of the SiC in molten aluminium followed by reaction and precipitation of the Al4C3 during cooling. PMID- 10540266 TI - Microstructural analysis of Al alloys dispersed with TiB2 particulate for MMC applications AB - A dispersion of TiB2 particulates in an Al alloy matrix was formed via the in situ reaction between mixtures of K2TiF6 (K2ZrF6), KBF4 and molten aluminium. The dispersion of the ceramic phase in the aluminium matrix was also achieved in some experiments by adding exogenous TiB2 particles to the fluoride melt in contact with molten aluminium. In this work, we have examined the microstructure of the as-cast metal matrix composites using analytical electron microscopy and X-ray diffraction techniques. The phases formed as a result of the reaction between the molten fluoride flux and liquid aluminium have been identified. These were (Ti, Zr, Al)B2, Al3Ti and possibly AlB12 in the Al-matrix, and KAlF4 and KMgF3 in the solidified flux. The mechanism of formation of TiB2 and Al3Ti is explained. The role of alloying elements is also explained in the context of interfacial chemistry and dispersion. PMID- 10540267 TI - Imaging damage evolution in a small particle metal matrix composite AB - It is difficult to study effectively microstructural damage in metal matrix composites (MMCs) due to artefacts arising from traditional metallographic sample preparation techniques. The sectioning and imaging capabilities of the focused ion beam (FIB) microscope provide an excellent method for studying damage accumulation in MMCs. The capabilities of the FIB system have been used to carry out a study of damage evolution in a powder-processed/hot-extruded Al2080/SiCp MMC. Microvoid damage is found to be preserved accurately during FIB sectioning, allowing measurements of the fraction of decohered particles and the void area fraction. These microscopic damage measurements are correlated with the macroscopic damage parameter, D, as determined by density measurements. Using transmission electron microscopy, the evolution of dislocation structures at the SiC-matrix interfaces has been examined. A previously unreported decohesion mechanism has been observed. PMID- 10540268 TI - Local variations of the chemistry in as-cast gamma-TiAl (TiBx) alloys and its consequence for thermomechanical treatments AB - The first part of the paper presents an accompanying investigation to a thermomechanical processing route of gamma-TiAl turbine blades. By means of scanning electron microscopy and energy dispersive X-ray (EDX) analysis, the chemical homogeneity and the microstructures of gamma-TiAl as-cast ingots, work pieces and the final turbine blades are determined. We find that the local Al content in as-cast ingots can vary by more than 1.5 at.%. Large chemical inhomogeneities present in as-cast ingots can only be eliminated to a certain degree by subsequent thermomechanical processing. An additional aim of the study is to assess the influence of a thermomechanical processing on the morphology of titanium boride precipitates and the alpha2-Ti3Al-phase. The second part of the paper contains a detailed analytical study devoted to homogenization of a range of gamma-TiAl cast alloys. Different microstructures are generated in a laboratory-scale argon-arc furnace by both rapid and slow solidification rates and an additional homogenization treatment, respectively. Quantitative EDX analysis shows that only rapid solidification of ingots with a subsequent homogenization treatment leads to a nearly chemically homogeneous microstructure. PMID- 10540269 TI - The response of SiC fibres to vacuum plasma spraying and vacuum hot pressing during the fabrication of titanium matrix composites AB - Vacuum plasma spraying (VPS) and vacuum hot pressing (VHP) have been used to fabricate Ti-6Al-4V matrix composite material reinforced longitudinally with DERA Sigma C coated SiC 1140+ fibres. VPS of Ti-6Al-4V onto Sigma 1140+ SiC fibres caused no fibre/matrix interfacial reaction. During VHP a fibre/matrix reaction occurred, producing a mixture of fine (< 50 nm) TiCx (x /= 31 year-old group. This is consistent with findings in other countries. This is the first report of the prevalence of HEV infection in Bolivia. PMID- 10540301 TI - In vitro antiplasmodial activity of Central American medicinal plants. AB - The in vitro antiplasmodial activities of 14 plant species traditionally used in Central America for the treatment of malaria or fever were evaluated. Lipophilic extracts of Piper hispidum, Siparuna andina, S. pauciflora, S. tonduziana, and Xylopia cf. frutescens, proved to be active against both a chloroquine-sensitive and a resistant strain of Plasmodium falciparum. IC50 values ranged between 3.0 microg/ml and 21.9 microg/ml; however, moderate cytotoxicity of active extracts was observed. Bioactivity-guided fractionation of Piper hispidum yielded 2',4, 6' trihydroxy-4'-methoxydihydrochalcone (asebogenin) as an active compound. PMID- 10540300 TI - Comparative clinical trial of four regimens of dihydroartemisinin-mefloquine in multidrug-resistant falciparum malaria. AB - We conducted a randomized, comparative trial at the Bangkok Hospital for Tropical Diseases during 1996-98 to evaluate the clinical efficacy and tolerability of four combination regimens of dihydroartemisinin-mefloquine. 207 male patients aged 18-25 years, weighing 49.3-55.1 kg were randomized to receive a single oral dose of 300 mg dihydroartemisinin plus one or two doses of mefloquine as follows: regimen I (n = 26): 750 mg mefloquine concurrently, or regimen II (n = 22): 750 mg mefloquine 24 h later, or regimen III (n = 78): 750 and 500 mg mefloquine at 24 and 30 h, or regimen IV (n = 81): 750 and 500 mg mefloquine (at 0 and 24 h). All patients improved clinically within 24 h of initiation of treatment. The initial therapeutic response was rapid and identical in all treatment groups (median PCT vs. FCT: 36 vs. 24, 36 vs. 28, 36 vs. 26, and 34 vs. 26 h, for regimen I, II, III and IV, respectively). All combination regimens generally showed acceptable tolerability profiles. Compliance with follow-up (42 days) was achieved by 86.5% (179 cases). Recrudescent parasitaemia was significantly higher in patients treated with low-dose mefloquine combinations (regimens I, II:8/23, 9/16) than in those who received high-dose mefloquine (regimens III, IV: 2/70, 3/70). No RII or RIII type of response was observed. There were no significant differences in susceptibility to mefloquine between primary and recrudescent isolates. Dose-adjusted whole blood mefloquine concentrations were significantly higher in high-dose mefloquine regimens (III and IV). Patients who vomited within the first hour of mefloquine administration had markedly lower whole blood mefloquine concentrations than those who did not vomit. PMID- 10540302 TI - An enzyme-linked immunosorbent assay for detection of IgG1 antibodies specific to human cystic echinococcosis in Egypt. AB - Human cystic hydatidosis (cystic echinococcosis) is a chronic zoonotic disease that results from infection with the dog tapeworm Echinococcus granulosus. In Egypt, cystic echinococcosis (CE) is recognized in slaughtered livestock by veterinarians, however, there is little information about human CE infection rates. We describe an immunological assay useful for the diagnosis of human cystic hydatidosis. Sera were collected from surgically confirmed hydatid cases (34), nonendemic subjects free from parasitic infection (20) and from subjects (109) infected with other helminths (Hymenolepis nana, Schistosoma mansoni, Fasciola hepatica and Ancylostoma duodenale). Hydatid cyst fluid (HCF) of camel origin was used as antigen in an ELISA format to measure total E. granulosus specific IgG antibodies and IgG subclasses. Sensitivity measurements of total IgG, and IgG1-4 were 100, 100, 79.4, 61.8 and 55.9%, respectively, whereas respective specificity reached 65.1, 97.7, 98.4, 96.1 and 83. 7%. The diagnostic value of measuring IgG1 (97.7%), as assessed by a rating index (J) for combined sensitivity and specificity, was superior to total IgG (65.1%) and IgG2-4 (77.8, 57.9 and 39.6%, respectively). These findings set the stage for field evaluation of the IgG1 assay in areas endemic with human cystic hydatidosis. PMID- 10540303 TI - Lung function, blood gases, pH and serum electrolytes of small-scale miners exposed to chrome ore dust on the Great Dyke in Zimbabwe. AB - We measured and compared lung function indices and some blood parameters (gases, electrolytes, glucose, pH, red cell indices) of 54 male small-scale miners (SSM) chronically exposed to chrome ore dust to those of 50 nonmining control subjects (NMC) and 46 large-scale chrome miners (LSM) who had taken internationally recommended precautionary measures (secondary control). The respirable dust level in the SSM environment (6.0 +/- 0.5 mg/m3) was significantly higher (P < 0.001) than in the NMC and LSM environments (0.3 +/- 0.1 mg/m3 and 0.5 +/- 0.3 mg/m3, respectively). There were no significant differences in neither dust levels nor lung function status between NMC and LSM environments. The values of FVC, FEV1, PEFR and FEV1% of the SSM were 3.5 +/- 0.09 l, 2.61 +/- 0. 09 l, 6.07 +/- 0.36 l/second and 76.19 +/- 2.36%, respectively. These values were significantly lower than those of NMC (P < 0.01, respectively). However, the blood parameters of the SSM and NMC were not significantly different. The results are indicative of both restrictive and obstructive ventilatory defects in the SSM which may be attributed exposure to chrome ore dust in the environment. Associated risk factors appear to be advancing age, prolonged exposure to chrome ore dust and acid base disturbance. PMID- 10540304 TI - Contact dermatitis due to tea tree oil. PMID- 10540306 TI - Dogmas and misunderstandings in East Coast fever. AB - East Coast fever (ECF) is the most important tick-borne disease in eastern, central and southern Africa and caused an estimated loss of US $186 million in 1989 in the 11 countries where it occurs. It was brought to southern Africa with cattle from Tanzania in 1901 and, over the next 3 years, devastated the cattle that had survived the rinderpest pandemic of the 1890s. Chemical control of ticks using arsenical compounds was introduced in the early 1900s and became the main control measure for both ticks and the diseases they transmit. This method of control has become less reliable over the last 30 years for many reasons, including reduced government spending on livestock and extension, the cost of acaricides, acaricide resistance, poor management of dips and spray races, and poor application of cattle movement control and quarantine. Significant advances in immunization and treatment have been made in the last 30 years, and more robust integrated strategies combining immunization, reduced frequency of chemical control and treatment are being adopted or considered. Throughout its history, ECF has been a source of great anxiety and cost to farmers, and of intense interest to research workers. Many dogmas and misconceptions have become established, some of which still flourish while others took years to demolish. This paper briefly reviews these as well as the history of the disease and explores recent epidemiological findings and their relevance to applying effective control. PMID- 10540307 TI - Immunization against diseases caused by Theileria parva: a review. AB - Theileria parva is the causative agent of three epidemiologically different diseases, East Coast fever (ECF), Corridor disease and January disease, caused by 3 types of T. parva, T. p. parva, T. p. lawrencei and T. p. bovis, respectively. The history of immunization against these diseases has been marked by salient discoveries such as the immune status in recovered animals, the activity of tetracyclines during the incubation period, the possibility for cryopreserving supernatant of prefed ticks and the development of useful serological tests. The possibility of simultaneous administration of stabilate and long-acting tetracycline have greatly contributed to making the infection and treatment method operational. The importance of antigenic diversity in T. parva has been reflected in the difficulties related to the selection of the immunizing stock or combinations of stocks: a 'cocktail' of East African isolates may give broad protection against field challenge by ECF (T. parva parva), but Corridor disease is more problematic. On the other hand, certain single isolates may give equally good protection against ECF field challenge. Studies on the immunology of T. parva infection and the application of molecular tools have led to the discovery that sera of recovered animals neutralize sporozoites of various isolates, and to the p67 molecular vaccine; yet so far the only available method of immunizing against T. parva infections is the infection and treatment method or, in the case of T. parva bovis, the use of sublethal stabilate doses. Infection and treatment is applied on a fairly large scale in Zambia, and on a more limited scale in a few other countries. Immunity by this rather crude method is long-lasting and solid, but cross-immunity problems against some field strains remain. Furthermore, as immunized animals remain carriers, immunization may contribute to attaining and improving endemic stability in endemic areas in indigenous breeds with an adequate level of genetic tolerance to ECF. On the other hand, carrier animals may constitute a risk for spreading the disease into ECF-free areas where the vector is present. Other disadvantages of the method are that immunization of cattle during the incubation of naturally contracted East Coast fever will not prevent the disease and jeopardize its reputation. Furthermore, stabilates have to be cryopreserved, often a technical drawback, and contamination with undesirable pathogens may occur in tick-derived material. Therefore the need remains for the development of effective molecular vaccines and it must be remembered that immunization must be cost-effective and sustainable and it is only one aspect of integrated control of theileriosis and other tick-borne diseases. There is no universally valid strategy. Several factors have to be considered: value and susceptibility of cattle to theileriosis and to other tick borne and tick-associated diseases, infestation by various ticks present in the area, the type of theileriosis (ECF, Corridor disease or January disease) and the epidemiological situation where immunization is taking place. The optimal age for immunization of the calves in endemic areas needs to be determined: when calf mortality by naturally occurring theileriosis is a problem, the sooner calves are immunized the better, but a proportion will have contracted natural infection before they can be reached, and immunization of very young calves might not be fully effective. PMID- 10540309 TI - The current epidemiological status of bovine theileriosis in eastern Zambia. AB - Results of a longitudinal study conducted in the eastern province of Zambia from 1994 to 1997 indicate that it is doubtful whether a state of endemic stability of East Coast fever (ECF) can be reached in the near future. Even in endemic areas, the mortality of Theileria parva infections is still estimated above 50%. The main factors limiting progress towards endemic stability are high innate susceptibility of the Zebu cattle, the virulence of the parasite and the climate. The unimodal rainfall pattern results in restricted activity of Rhipicephalus appendiculatus instars and year-to-year variation in rainfall causes fluctuations in tick phenology and T. parva transmission. Adult tick activity invariably peaks during the rains and is associated with the highest ECF incidence. Nymphal transmission of T. parva to cattle appears to be less important. Second periods of activity of both adult and nymphal instars are pronounced only when the climate is suitable. These second waves of tick activity ensure a more continuous and efficient transmission of T. parva and also play a key role in the dynamics of prolonged outbreaks in epidemic areas. ECF control methods may have an important influence on ECF epidemiology. Immunizations as well as chemotherapy of clinical cases create a reservoir of virulent parasites in susceptible cattle, resulting in artificial endemic stability. PMID- 10540308 TI - Molecular epidemiology of Theileria parva in the field. AB - Molecular tools based on seminested RFLP-PCR techniques to characterize field parasites in bloodspots dried on filter paper permitted investigation of the extent and the dynamics of diversity of Theileria parva populations in the field. Parallel molecular studies explored the long-term genome stability of various isolates by probing Southern blots of EcoRI digested total genomic DNA with four different reference nucleic acid probes. Three polymorphic single copy loci encoding for antigen genes were developed for seminested PCR detection in order to apply them for a multilocus approach in population genetic studies. Seven alleles were identified for the polymorphic immunodominant molecule (PIM) locus by using restriction enzymes, and 4 alleles each for the p150 and p104 loci. A simple DNA extraction method gave good results in amplifying these loci from carrier animals using samples of blood dried on filter papers. Results from probing Southern blots of cultures taken at sequential timepoints indicate relative genome stability in T. parva in comparison to other parasitic protozoa such as Plasmodium. Comparatively homogeneous profiles in sympatric isolates from Zambia were identified using all four probes and PCR amplified products which contrasted with the variety found amongst Kenyan stocks. Preliminary characterization of T. parva field samples from the Southern Province of Zambia strongly suggest clonal expansion of one of the components of a non-Zambian trivalent vaccine used on a limited scale in the Province from 1985 until 1992. PMID- 10540310 TI - Epidemiological uses of a population model for the tick Rhipicephalus appendiculatus. AB - The spatial and temporal risk of tick-borne disease depends fundamentally on the distribution, abundance and seasonal dynamics of the vector ticks. The latter factor exerts a major quantitative influence on the transmission dynamics of tick borne parasites. The population model for Rhipicephalus appendiculatus applies throughout the range of this tick in eastern Africa, and predicts all three fundamental risk factors on the basis of the local temperature and rainfall conditions. Satellite imagery can provide more detailed, real-time measures of environmental conditions over extensive areas than climatic data. There is preliminary evidence to suggest that the population model could be driven by satellite-derived surrogates of its climatic predictors, thus providing wide scale predictive risk maps of theileriosis. PMID- 10540311 TI - Spatial and temporal variation in Rhipicephalus appendiculatus size in eastern Zambia. AB - The size of Rhipicephalus appendiculatus collected at different altitudes in the Eastern Province of Zambia between February 1985 and May 1986 and between October 1994 and December 1996 showed distinct variation dependent on altitude and season. The ticks were smallest during the dry season and at the start of the rains, and specimens were larger as the rainy season progressed. Second generation adults where on average smaller than first-generation ticks. At higher altitudes, where a one-generation-per-annum phenology dominates, ticks were larger than at intermediate altitudes, where two generations per year are common. Larger size, associated with increased survival, is also favoured in low-lying, drier areas. Selective mortality of smaller adult ticks in years with a delayed rainy season appears to play an important role in the variation in size between years. PMID- 10540312 TI - Economics of theileriosis control in Zambia. AB - For an economic analysis of theileriosis control, we adopted the total economic cost (TEC) method, which calculates the sum of output losses from tick damage, theileriosis mortality and morbidity, and expenditures for treatment or prevention of the disease. At farm level, the TEC can be minimized by a specific combination of vector control and/or immunization and an acceptable level of losses. Expenditures for vector control include acaricides, construction of dipping or spraying facilities and their maintenance, and variable costs such as those for water and labour. Economics of vector control depend on the herd size and the method of application of the acaricide. Morbidity, mortality and tick damage losses are effectively reduced by correct and intensive vector control programmes. Expenditures for vector control are estimated at US$ 8. 43, 13.62 and 21.09 per animal per year for plunge dipping, hand spraying and pour-on, respectively. Immunization costs comprise production of parasite stabilates, storage and application, delivery and treatment. At US$ 9.5 per animal, immunization limits losses caused by Theileria parva, but ticks still may reduce the productivity of the animals. Expenditures for treatment after natural infection involve drugs, transport, veterinary fees and farm labour costs. Treatment has a moderate success rate, hence both morbidity and mortality remain important factors. Equally, it does not affect the vector, which may continue to reduce overall productivity of cattle. Expenditures for treatment range between US$ 9.04 and US$ 27.31 per animal. To compare different TECs in relation to different control strategies, assumptions have to be made on disease occurrence, case fatality, value and productivity of the cattle, reductions in productivity due to morbidity and number of animals under a specific control regime. Calculations based on data from Southern Province, Zambia show that large-scale immunization reduces the TEC by 90% compared to no intervention. Treatment, which is the second-best option, reduces the TEC by 60%. Appendix 1 Summary of factors influencing total economic cost PMID- 10540313 TI - Theileriosis control modelling (experiences from Southern Province, Zambia). AB - Effects of different tick-borne disease control strategies on cattle productivity are simulated based on a 30-year herd projection, calculated by a modified Markov Chain model. Input data can be grouped in technical, economic and epidemiological parameters. The output is a set of economic parameters such as benefit/cost ratio (BCR), net present value (NPV) of the profit, internal rate of return (IRR), total economic cost (TEC) as well as graphs showing animal production over time. Shadow prices are obtained for input and output in kind. Throughout the calculations a distinction is made between transactions in cash and transactions in kind. A case study was run for Southern Province, Zambia, to illustrate the model. Either vector control or treatment, or a combination of these, controls theileriosis at farm level after natural infection. Preventive immunization against the parasite is also possible. Although the calculations are based on a mixture of data obtained from literature, field experience, expert opinion and assumptions, the importance of theileriosis control is clearly indicated. Immunization gives better economic results than chemotherapy. Vector control can only be used as a last resort. PMID- 10540314 TI - Evaluation of recombinant sporozoite antigen SPAG-1 as a vaccine candidate against Theileria annulata by the use of different delivery systems. AB - The major sporozoite surface antigen of Theileria annulata (SPAG-1) is a candidate for inclusion in a subunit vaccine. In this paper we summarize the results of 4 vaccination experiments using recombinant SPAG-1 expressed in different systems and presented in different adjuvants. The antigen has been presented as either a C terminal 108 amino acid peptide (called SR1) expressed as both beta-galactosidase and hepatitis B core antigen fusions or as a full-length form expressed as a GST fusion with an N terminal His6 tag. We used different adjuvants, namely Freund's, saponin, ISCOMs and a proprietary adjuvant supplied by SmithKline Beecham, which we call SKBA. The data point to the conclusion that SPAG-1 can elicit partial protection and is therefore suitable for inclusion in an eventual multicomponent subunit vaccine. PMID- 10540315 TI - Mechanism(s) of attenuation of Theileria annulata vaccine cell lines. AB - Attenuated vaccines are an important means of controlling Theileria annulata infection of cattle. Production is by prolonged cultivation of macroschizont infected cells. The mechanism of attenuation remains unclear. There are three general nonmutually exclusive possibilities: Selection of avirulent subpopulations, genome rearrangements and alterations in gene expression. Several groups, including ours, have provided evidence that the population structure usually tends to simplify during attenuation. Our data on the T. annulata (Ta) Ankara cell line show that attenuation is not necessarily accompanied by the population becoming clonal. We have been unable to detect large DNA rearrangements. Evidence for alterations in host and parasite gene expression during attenuation is available. With respect to the host we have shown that attenuation is accompanied by loss of expression of parasite induced matrix metalloproteinases (MMPs). However, in different lines different protease activities are involved. In the T. annulata Ode line we have shown that 8 activities (including MMP9) are downregulated and that this correlates with a loss of metastatic behaviour. This has previously been shown in vitro using reconstituted basement membrane (Matrigel) and is demonstrated in vivo using scid mice in this study. Thus part of the pathology, namely the ability to disseminate, mediated by host MMPs, is lost upon attenuation. Re-isolation experiments have shown that the reduction/loss of MMP is a stable transferable trait. A logical extension is that loss of MMP activity (and virulence in general) must be at the most fundamental level a genetic trait of the parasite. Evidence for loss of parasite gene expression is implied by the loss of the ability to differentiate into merozoites on attenuation. Specific evidence for loss of parasite gene expression has been obtained using differential RNA display. We view virulence as a multifactorial phenomenon involving interacting subpopulations of cells and attenuation is a threshold effect whereby the number of virulence factors is reduced below a critical level. On this basis there will be many different ways to achieve attenuation. PMID- 10540317 TI - Microglia induce myelin basic protein-specific T cell anergy or T cell activation, according to their state of activation. AB - Microglial cells are non-professional antigen-presenting cells (APC) the function of which is still controversial. Here, we studied the function of microglia derived from H-2(u) mice. We show that these microglia express a low level of B7.2 and CD40 and, interestingly, lack surface expression of B7.1. Resting and IFN-gamma-activated microglia were unable to activate naive and primed myelin basic protein (MBP)-specific CD4(+) T cells in the presence of MBP and encephalomyelitic MBP Ac1-11 peptide. Furthermore, in the presence of Ac1-11 peptide, CD4(+) TCR-transgenic T cells became anergized. Microglia became professional APC only after a multistep activation process involving both stimulation through cytokines [granulocyte-macrophage colony-stimulating factor (GM-CSF) and IFN-gamma] and cognate signaling (B7-CD28 and CD40-CD40 ligand interactions). As such they were able to present MBP to both unprimed and primed T cells. Co-culture of microglia with GM-CSF up-regulated co-stimulatory molecules, in particular B7.1. Additional activation with IFN-gamma induced MHC class II and CD40 up-regulation. CD40-CD40 ligand interaction significantly enhanced microglial ability to prime TCR-transgenic T cells and was essential for presentation of MBP to in vivo primed non-transgenic T cells. We propose that microglia may serve different functions under different inflammatory conditions, depending on the cytokine milieu and the type of cognate interaction they are involved in. PMID- 10540316 TI - Involvement of the N-methyl-D-aspartate receptor in neuronal cell death induced by cytotoxic T cell-derived secretory granules. AB - The mechanisms underlying neurotoxicity mediated by cytotoxic T lymphocytes (CTL) and their secretory granule proteins perforin and granzymes remain unclear. We evaluated the possible role of the neurotransmitter glutamate in cell death observed in differentiated neurons exposed to CTL-derived granules. Excitotoxicity induced by excessive concentrations of extracellular glutamate is associated with a rise in intracellular calcium and can lead to generation of NO through the activation of glutamatergic N-methyl-D-aspartate (NMDA) receptors. Consistent with an involvement of glutamate, we found that cell death in mature cerebral granule cells was inhibited by 65-80% by two NMDA receptor blockers (MK 801 and D-2-amino-5-phosphonovaleric acid) or a NO synthase blocker (N(G)-nitro-L arginine methylester). Furthermore, neurons treated with secretory granules responded with a biphasic rise in the intracellular calcium concentration ([Ca2+]i). Whereas MK-801 did not interfere with the immediate rise of [Ca2+]i, the second wave of calcium accumulation starting at 40 min was delayed by 20 min and reduced in amplitude in the presence of MK-801. In immature, NMDA receptor negative neurons, MK-801 prevented neither the cytotoxicity nor the calcium influx observed 5 min after addition of cytotoxic granules. The demonstration that NMDA receptors and NO are involved in granule-mediated killing of mature neurons opens new avenues in the treatment of neuronal cell death in CTL-mediated diseases such as viral encephalitis. PMID- 10540318 TI - A1 expression is stimulated by CD40 in B cells and rescues WEHI 231 cells from anti-IgM-induced cell death. AB - Engagement of the antigen receptor on murine immature B cells leads to growth arrest followed by apoptosis. Concomitant signaling through CD40 sustains proliferation and rescues the cells from apoptosis. We show here that cross linking CD40 stimulates the expression of A1, a member of the anti-apoptotic Bcl 2 family, in primary murine B lymphocytes. CD40-dependent stimulation of A1 was confirmed in WEHI 231 cells, an immature murine B cell lymphoma line. We transduced WEHI 231 cells with a bicistronic recombinant retroviral vector coding for A1 and a chimeric selection marker comprising the enhanced yellow fluorescent protein and the zeocin resistance protein. A1-transduced WEHI 231 cells showed a significant higher survival rate after engagement of the antigen receptor. In contrast, constitutive expression of A1 did not abrogate anti-IgM-induced c-myc down-regulation. Consistent with this, A1 did not release anti-IgM-induced cell cycle arrest. Our data indicate that CD40-stimulated A1 expression permits WEHI 231 cells to survive in the presence of anti-IgM antibodies and suggests a protective role for A1 in antigen receptor-mediated apoptosis in B cells. PMID- 10540319 TI - Reduced antilisterial activity of TNF-deficient bone marrow-derived macrophages is due to impaired superoxide production. AB - Mice deficient for TNF ligand and receptor type 1 have demonstrated the importance of TNF in the host defense against Listeria monocytogenes. To investigate the particular deficiency of macrophages derived from TNF/lymphotoxin (LT)-alpha(-/-) mice in antilisterial growth control, bone marrow-derived macrophages (BMDM) were used for in vitro infection experiments. After the combined treatment with IFN-gamma and lipopolysaccharide (LPS), production of NO by wild-type (wt) and TNF/LT-alpha(-/-) BMDM was induced to comparable levels, but only wt BMDM controlled L. monocytogenes growth efficiently. Nevertheless, inhibition of NO production led to a remarkable loss of antilisterial activity. This suggests that presence of NO is necessary but not sufficient for L. monocytogenes killing and that elimination of L. monocytogenes requires additional effector molecules. The LPS-inducible superoxide production of TNF/LT alpha(-/-) BMDM was impaired. Accordingly both scavenging of superoxide and peroxynitrite led to reduced L. monocytogenes killing by wt BMDM. In addition, peroxynitrite was able to kill L. monocytogenes in vitro. Together these findings suggest that the defective host defense of TNF/LT-alpha-deficient mice against L. monocytogenes partially stems from reduced superoxide production of macrophages due to the absence of TNF and imply a function for peroxynitrite, the reaction product of NO and superoxide, in the intracellular killing of L. monocytogenes. PMID- 10540320 TI - Cyclic adenosine monophosphate-responsive elements are involved in the transcriptional activation of the human IL-10 gene in monocytic cells. AB - IL-10 plays an important role in the regulation of immune responses. We and others have demonstrated recently that cyclic adenosine monophosphate (cAMP) elevating substances up-regulate monocytic IL-10 expression in vitro and in vivo. Computer analysis of the IL-10 promoter/enhancer region localized four putative cAMP-responsive elements (CRE1- 4) with homology to the CRE consensus motif. In electrophoretic mobility shift assays CRE1 and CRE4 bound protein complexes consisting of transcription factors CREB-1 and ATF-1, while CRE3 bound only marginal amounts of CREB-1/ATF-1 in combination with unknown protein(s). CRE2 showed no protein binding activity. In vitro mutation of CRE1 and CRE4 reduced the level of cAMP-stimulated transactivation in reporter gene assays in comparison to the wild-type promoter by 20 % and 50 %, respectively, while mutation of CRE3 had no effect. The main action of CRE4 on cAMP-dependent stimulation is probably based on its adjacent localization to the TATA box and its sequence comprising a perfect half site. Experiments with double and triple mutants and with deleted promoter fragments indicated the participation of additional elements beside the CRE motifs in the cAMP-dependent stimulation. Our data suggest that intracellular cAMP may directly affect expression of the immunoregulatory cytokine IL-10 in monocytic cells via activation of the eukaryotic transcription factors CREB-1 and ATF-1 and their binding to CRE1 and CRE4 in the upstream enhancer of the IL-10 promoter. PMID- 10540321 TI - The lectin-like domain of tumor necrosis factor-alpha increases membrane conductance in microvascular endothelial cells and peritoneal macrophages. AB - Herein, we show that TNF exerts a pH-dependent increase in membrane conductance in primary lung microvascular endothelial cells and peritoneal macrophages. This effect was TNF receptor-independent, since it also occurred in cells isolated from mice deficient in both types of TNF receptors. A TNF mutant in which the three amino acids critical for the lectin-like activity were replaced by an alanine did not show any significant effect on membrane conductance. Moreover, a synthetic 17-amino acid peptide of TNF, which was previously shown to exert lectin-like activity, also increased the ion permeability in these cells. The amiloride sensitivity of the observed activity suggests a binding of TNF to an endogenous ion channel rather than channel formation by TNF itself. This may have important implications in mechanisms of TNF-mediated vascular pathology. PMID- 10540323 TI - Targeting and subsequent selection of somatic hypermutations in the human V kappa repertoire. AB - The number and distribution of nucleotide substitutions in human VkappaJkappa genes were examined using a PCR technique that analyzed nonproductive and productive rearrangements amplified from genomic DNA of individual B cells. The results indicate that the mutational mechanism introduces replacement (R) mutations comparably throughout the length of the VkappaJkappa rearrangement, but tends to target specific triplets. Moreover, hotspots of mutational activity were identified in complementarity determining regions (CDR). A marked increase in the frequency of R mutations in CDR was noted when productive were compared to nonproductive rearrangements, indicating that these were selected into the expressed repertoire. Of note, amino acids encoded by codons adjacent to hotspots of mutation were also positively selected implying that similar regions were targeted for hypermutation and subsequent selection. In contrast to the distribution of CDR mutations, R mutations in the framework (FR) regions tended to be eliminated from productive VkappaJkappa rearrangements, implying that the somatic hypermutational machinery frequently introduced amino acid changes that were deleterious to the structural integrity of the kappa chain protein. The difference in the ratio of R to silent mutations in CDR and FR in the expressed repertoire, therefore, reflects the summation of positive selection of R mutations in the CDR and the elimination of R mutations in the FR. The data indicate that the balance between targeted mutation of VkappaJkappa rearrangements and subsequent selection and elimination governs the pattern of mutations manifest within the expressed kappa repertoire. PMID- 10540322 TI - H-2 class I knockout, HLA-A2.1-transgenic mice: a versatile animal model for preclinical evaluation of antitumor immunotherapeutic strategies. AB - H-2 class I-negative, HLA-A2.1-transgenic HHD mice were used for a comparative evaluation of the immunogenicity of HLA-A2.1-restricted human tumor-associated cytotoxic T lymphocyte (CTL) epitopes. A hierarchy was established among these peptides injected into mice in incomplete Freund's adjuvant which correlates globally with their capacity to bind and stabilize HLA-A2.1 molecules. Co injection of a helper peptide enhanced most CTL responses. In contrast, classical HLA class I-transgenic mice which still express their own class I molecules did not, in most cases, develop HLA-A2.1-restricted CTL responses under the same experimental conditions. Different monoepitope immunization strategies of acceptable clinical usage were compared in HHD mice. Recombinant Ty-virus-like particles, or DNA encoding epitopes fused to the hepatitis B virus middle envelope protein gave the best results. Using this latter approach and a melanoma based polyepitope construct, CTL responses against five distinct epitopes could be elicited simultaneously in a single animal. Thus, HHD mice provide a versatile animal model for preclinical evaluation of peptide-based cancer immunotherapy. PMID- 10540324 TI - Glutenin is involved in the gluten-driven mucosal T cell response. AB - Gluten ingestion causes coeliac disease in susceptible individuals. Gluten is a heterogeneous mixture of glutenin and gliadin, the latter of which is considered responsible for disease induction. By combining high-performance liquid chromatography purification steps of gluten with a T cell bioassay and mass spectral analyses, we have identified a glutenin peptide (glt04 707-742) that activates T cells from the small intestine of a coeliac disease patient and results in the secretion of large amounts of IFN-gamma. The minimal T cell stimulatory core of the peptide (residues 724-734) is repetitively present in glutenin molecules. Moreover, it was observed that a large number of naturally occurring variants of this peptide are recognized by the T cells. These data suggest that the large heterogeneity of glutenin proteins dramatically increases the number of available T cell epitopes. Together, the results provide new insight into the nature of the gluten antigens that lead to coeliac disease and suggest that glutenin, next to gliadin-derived antigens, may be involved in the disease process. PMID- 10540325 TI - T cells are the main cell type expressing B7-1 and B7-2 in the central nervous system during acute, relapsing and chronic experimental autoimmune encephalomyelitis. AB - T cell co-stimulation through the CD28 receptor on T cells is critical to the induction of experimental autoimmune encephalomyelitis (EAE). In this study, expression of the co-stimulatory ligands B7-1 (CD80) and B7-2 (CD86), as well as the receptors CD28 and CTLA-4, were quantitated in central nervous system (CNS) tissues from mice at various stages of EAE. Immunohistochemistry and flow cytometry of CNS-infiltrating cells revealed a high percentage of infiltrating T cells expressing B7-1 and B7-2 during acute, chronic and relapsing EAE. Of the infiltrating cells 10-20% were CTLA-4(+), most of which were CD4(+) T cells. B7-1 and B7-2 expression within the CNS during active EAE might increase the potential for local activation of autoimmune T cells; however, the high level of expression of B7 molecules may also provide a mechanism for the autoregulation of activated CTLA-4(+) T cells. PMID- 10540326 TI - Molecular and functional characterization of IRp60, a member of the immunoglobulin superfamily that functions as an inhibitory receptor in human NK cells. AB - In this study we describe the functional and molecular characterization of IRp60 (inhibitory receptor protein 60), an inhibitory receptor expressed on all human NK cells. The IRp60 molecule has been identified by the generation of three novel monoclonal antibodies (mAb). Cross-linking of IRp60 by specific mAb strongly inhibits the spontaneous cytotoxicity of NK cells as well as the NK-mediated cytolytic activity induced via different non-HLA-specific or HLA-specific activating receptors. IRp60 is a 60-kDa glycoprotein that, upon sodium pervanadate treatment, becomes tyrosine phosphorylated and associates with the SH2-containing phosphatases SHP-1 and SHP-2. The IRp60 gene is located on human chromosome 17 and encodes a molecule belonging to the immunoglobulin (Ig) superfamily characterized by a single V-type Ig-like domain in the extracellular portion. The cytoplasmic tail contains three classical immunoreceptor tyrosine based inhibitory motifs. Southern blot analysis revealed cross-hybridization with monkey and mouse genomic DNA, thus suggesting that IRp60 may be conserved among different species. Moreover, based on the use of different anti-IRp60 mAb, we could identify two IRp60 allelic variants. Since IRp60 is also expressed by other cell types, including T cell subsets, monocytes and granulocytes, it may play a more general role in the negative regulation of different leukocyte populations. PMID- 10540327 TI - Leukocyte-associated immunoglobulin-like receptor-1 (LAIR-1) is differentially expressed during human B cell differentiation and inhibits B cell receptor mediated signaling. AB - Leukocyte-associated Ig-like receptor-1 (LAIR-1) belongs to the growing family of immunoreceptor tyrosine-based inhibitory motif-bearing receptors and is expressed on the majority of peripheral mononuclear cells, including NK cells, T cells, B cells, monocytes, and dendritic cells. In this study, we investigated the distribution and the capacity of LAIR-1 to function as an inhibitory receptor on human B cells. LAIR-1 is expressed from early on during B cell differentiation, but is absent on approximately half of the memory B cells, and all germinal center B cells, plasmablasts, and terminally differentiated plasma cells. In vitro stimulation of naive B cells via the B cell receptor (BCR) or CD40, triggering proliferation and differentiation into Ig-producing plasma cells, is accompanied by loss of LAIR-1 expression. We previously reported that LAIR-1 can function as an inhibitory receptor on NK cells and T cells. Here, we demonstrate that it can also function as a negative regulator of BCR-mediated signaling, since simultaneous cross-linking of LAIR-1 and the BCR reduces the increase of intracellular Ca(2+) evoked by BCR ligation. Taken together, this suggests that the inhibitory mechanism of LAIR-1 is functional in multiple components of the hematopoietic system. PMID- 10540329 TI - Tyrosine kinase-dependent ubiquitination of CD16 zeta subunit in human NK cells following receptor engagement. AB - We investigated whether aggregation of the low-affinity immunoglobulin G receptor (CD16) on human NK cells results in receptor ubiquitination. We found that the CD16 zeta subunit becomes ubiquitinated in response to receptor engagement. We then investigated whether protein tyrosine kinase (PTK) activation is required for CD16-mediated receptor ubiquitination. Pretreatment with the PTK inhibitor genistein substantially decreased ligand-induced zeta ubiquitination, suggesting a requirement for PTK activation in receptor ubiquitination. We further analyzed PTK involvement in controlling receptor ubiquitination by using the vaccinia virus expression system. Overexpression of wild-type active lck, but not a kinase deficient mutant, enhanced both ligand-induced tyrosine phosphorylation and ubiquitination of the CD16 zeta subunit. Taken together, our data demonstrate that CD16 engagement induces zeta chain ubiquitination and strongly suggest a role for lck in regulating this modification. PMID- 10540328 TI - Bcl-2 leads to expression of anti-DNA B cells but no nephritis: a model for a clinical subset. AB - Transgenic mice expressing anti-DNA antibodies have been extensively studied as a model for understanding B cell regulation in systemic lupus erythematosus (SLE). BALB/c mice transgenic for the R4A-gamma2b heavy chain of an anti-double-stranded DNA (dsDNA) antibody produce two populations of high-affinity anti-dsDNA B cells, one deleted, and the other anergized. We generated double-transgenic BALB / c mice expressing both the R4A-gamma2b heavy chain and the anti-apoptotic bcl-2 gene in the B cell compartment to study whether bcl-2 overexpression differentially affected anergic and deleted B cells. The double-transgenic mice (R4A/bcl-2) express elevated serum titers of both high- and low-affinity anti dsDNA antibodies and display rescue of autoreactive B cells that are normally either deleted or anergized. Despite the presence of anti-dsDNA antibodies in their serum, R4A/bcl-2-transgenic mice do not develop nephritis, demonstrating that overexpression of bcl-2 is not by itself sufficient to allow disease progression. This phenotype resembles that of some SLE patients who have high titers of anti-DNA antibodies without nephritis. PMID- 10540330 TI - Role of autologous CD4+ T cell clones in human B non-Hodgkin's lymphoma: aborted activation and G1 blockade induced by cell-cell contact. AB - This article describes the study of the functional relationship between auto tumor-reactive CD4(+) T cell clones (TCC) and autologous malignant B cells. Four auto-tumor-reactive CD4(+) TCC were derived from tumor-infiltrating T lymphocytes (TIL-T) from a freshly isolated human follicular lymphoma by the following technique: total CD4(+) TIL-T were negatively purified by an immunomagnetic procedure, then CD4(+) TCC were obtained by limiting dilution in the presence of IL-2 and autologous non-irradiated follicular lymphoma cells as feeders. After expansion, these CD4(+) TCC were co-cultured with non-irradiated autologous malignant B cells. All four TCC were activated by B lymphoma cells and proliferated, as assessed by CD25 expression and cell cycle analysis. Activation and proliferation of B lymphoma cells were studied in response to activated CD4(+) T cells. Although all four TCC were able to induce B lymphoma cell activation (Ki-67 antigen induction and CD40 up-regulation), cells were subsequently blocked in G1 phase. Activation of B-NHL cells was mediated by TCR HLA class II interaction, as shown by a blocking experiment using an anti-CD4 monoclonal antibody (mAb). Since anti-CD40 mAb with or without IL-4 did not induce proliferation of B lymphoma cells in contrast to normal B cells, we suggest that the blockade in G1 phase is due to the presence of abnormalities in B lymphoma cells. This is the first evidence that autologous reactive CD4(+) TCC can engage follicular lymphoma B cells to enter the cell cycle and induce an aborted activation stage. PMID- 10540331 TI - Urokinase-type plasminogen activator inhibits alpha 4 beta 1 integrin-mediated T lymphocyte adhesion to fibronectin independently of its catalytic activity. AB - The urokinase-type plasminogen activator (u-PA)/plasmin system plays an important role in promoting cell migration and invasion, an effect which is largely ascribed to the proteolytic activity of these enzymes. We investigated whether u PA modulates integrin-dependent T lymphocyte migration and adhesion on fibronectin independently of its plasminogen activator function. Here we report that u-PA reduced the spontaneous and phorbol 12-myristate 13-acetate-induced migration of peripheral blood T lymphocytes on fibronectin by 20-50%, decreased the T lymphocyte and alpha4beta1(+)/alpha5beta1(+) K562 cell adhesion on fibronectin by 30-40%, and completely suppressed integrin alpha4beta1-dependent T lymphocyte and alpha4beta1(+)/alpha5beta1(+) K562 cell adhesion to the LDV containing 40-kDa fibronectin fragment. The u-PA receptor was not essential for this effect. In contrast, adhesion of alpha4beta1(-)/alpha5beta1(+) K562 cells to an RGD-containing fibronectin fragment was unaffected. A recombinant protein comprising the N-terminal fragment of u-PA, but lacking its proteolytic domain, had the same inhibitory effect. Decreased adhesion was neither associated with a diminished cell surface expression of alpha4beta1 nor with a suppression of alpha4beta1 ligand-binding function. Our results demonstrate that u-PA inhibits alpha4beta1- but not alpha5beta1-mediated lymphocyte/leukocyte adhesion to fibronectin independently of its proteolytic activity. This finding provides additional evidence that matrix proteinases may participate in cell adhesion and migration control independently of their matrix-degrading activity. PMID- 10540332 TI - The assignment of chemokine-chemokine receptor pairs: TARC and MIP-1 beta are not ligands for human CC-chemokine receptor 8. AB - Identification of chemokine receptors and their associated ligands is crucial to the understanding of most immune reactions. Three human chemokines [I-309, thymus and activation-regulated chemokine (TARC) and macrophage inflammatory protein 1beta (MIP-1beta)] have been reported to be ligands for CC-chemokine receptor 8 (CCR8). In this report, we present evidence that TARC and MIP-1beta did not bind to or induce chemotaxis through CCR8 on a stable transfected cell line (1D-21) and did not bind to CCR8 on in vitro differentiated human CD4(+) Th(2) cell cultures. Also, I-309-dependent calcium mobilization in 1D-21 cells and in Th(2) cells was desensitized by I-309 but not by MIP-1beta or TARC. These results provide strong evidence that, at physiologically relevant concentrations, I-309 is the only known human ligand for CCR8. These data also provide a framework for suggesting minimum requirements for the assignment of chemokine receptor-ligand pairs. PMID- 10540333 TI - Anti-CD40 antibody enhances responses to polysaccharide without mimicking T cell help. AB - Protection against infection with encapsulated bacteria is mediated by IgG antibodies against the capsular polysaccharides. Production of such antibodies is impaired during infancy, when susceptibility to bacterial meningitis is greatest. Recent studies have proposed the use of anti-CD40 antibody to increase responsivenesses to polysaccharide antigens. We show here that the IgG response to a model polysaccharide antigen is greatly increased, but retains thymus independent characteristics--switching continues to be mainly to IgG3 and neither germinal centers nor memory B cells are formed. Furthermore, anti-CD40 causes striking splenomegaly in mice, which is accompanied by dramatic cellular redistribution and proliferation of dendritic cells, macrophages, T cells and endothelium, as well as B cells. These findings raise the possibility that the anti-CD40 effect is due mainly to increased activity of accessory cells that affect plasmablast growth and differentiation rather than mimicry of T cell help. PMID- 10540334 TI - Expression and release of chemokines associated with apoptotic cell death in human promonocytic U937 cells and peripheral blood mononuclear cells. AB - To characterize mechanisms which may determine the fate of apoptotic cells, we investigated chemokine expression in apoptotic promonocytic U937 cells or peripheral blood mononuclear cells (PBMC). Exposure of U937 cells to etoposide (VP-16) or the nitric oxide (NO) donor DETA-NO, both inducers of apoptosis in these cells, was associated with increased expression of the chemokines IL-8 and macrophage inflammatory protein-1 alpha. Up-regulation of IL-8 mRNA expression by VP-16 or DETA-NO was observed as early as 4 h or 6 h, respectively, after onset of treatment and was still detectable after 19 h of exposure. A serine protease inhibitor prevented both VP-16-induced apoptosis and release of IL-8, whereas inhibition of p38 MAP kinases reduced IL-8 secretion only. Moreover, we observed that incubation with 2-chlorodeoxyadenosine (CdA) up-regulated release of IL-8 from adherent PBMC in parallel to induction of apoptosis. In these cells a modest but significant induction of TNF-alpha release by CdA was also detected. In addition, CdA augmented release of IL-8 from whole blood cultures. By facilitating adequate recruitment of phagocytes to sites of cell death, stress induced up-regulation of chemokines associated with apoptosis may contribute to mechanisms aiming at efficient removal of apoptotic cells. PMID- 10540335 TI - The extrafollicular-to-follicular transition of human B lymphocytes: induction of functional globotriaosylceramide (CD77) on high threshold occupancy of CD40. AB - Amongst lymphocytes, expression of CD77 (globotriaosylceramide, Gb3) is exclusive to B cells of the germinal center (GC). Its acquisition by extrafollicular B cells may thus herald their commitment to a follicular response. Here we show that high threshold occupancy of CD40 by its cognate ligand (CD40L) promotes rapid induction of CD77 expression in non-GC (CD38(lo)) B cells. The kinetics of CD77 acquisition mirrored those of GC-related markers CD95 and CD86 but contrasted with the more delayed increase in CD38 expression. Induction of CD77 was not a simple consequence of cell cycle entry: other conditions of stimulation equally capable of driving proliferation failed to promote CD77 expression. CD77 was functional in that cells were now sensitive to Verotoxin-1, an Escherichia coli-derived ligand of Gb3. These data indicate that acquisition by extrafollicular B cells of CD77 results from high threshold occupancy of CD40, a situation that should be reached physiologically only once a critical level of T cell priming has been achieved. PMID- 10540336 TI - Inhibition of dendritic cell maturation by herpes simplex virus. AB - Maturation of dendritic cells (DC), leading to migration and increased T cell stimulatory capacity, is essential for the initiation of immune responses. This process is triggered by a variety of stimuli, such as inflammatory cytokines, bacterial and viral products. Using a recombinant disabled infectious single cycle herpes simplex virus 1 (HSV-1) encoding green fluorescent protein, we show that the infected DC are defective in up-regulating co-stimulatory molecules, do not produce cytokines, and do not acquire responsiveness to chemokines required for migration to secondary lymphoid organs. These results reveal yet another strategy used by HSV-1 to evade the immune response, namely the inhibition of signaling pathways involved in DC maturation. PMID- 10540337 TI - Inhibition of indoleamine 2,3-dioxygenase in human macrophages inhibits interferon-gamma-induced bacteriostasis but does not abrogate toxoplasmastasis. AB - Induction of indoleamine 2,3-dioxygenase (IDO) by IFN-gamma results in growth inhibition of Toxoplasma and Chlamydia spp. as well as tumor cells. This is caused by the degradation, and therefore depletion, of L-tryptophan necessary for cell protein synthesis. Human macrophages stimulated with IFN-gamma express IDO and inhibit the growth of intracellular toxoplasma and chlamydia as well as that of extracellular bacteria such as group B streptococci. Here we describe experiments in which the L-tryptophan analog, 6-chloro-DL-tryptophan (CDLT) caused a dose-dependent inhibition in the IFN-gamma-induced IDO-mediated L tryptophan degradation in monocyte-derived macrophages and glioblastoma cells. An inhibition of IDO activity of up to 80 % was observed at concentrations of CDLT of 750 microM. Expression of IDO at this concentration, as shown by Northern blot analysis, was unimpaired. This inhibition of IDO was coupled in glioblastoma cells by a complete abrogation of the IFN-gamma-induced toxoplasmastasis in these cells. IDO inhibition by CDLT in human macrophages resulted in a complete abrogation of the IFN-gamma-induced growth inhibition of streptococci and staphylococci. In contrast to this, IFN-gamma-induced toxoplasmastasis was not inhibited in human macrophages by CDLT-mediated IDO inhibition. PMID- 10540338 TI - Homeostatic regulation of CD8+ T cells by perforin. AB - To prevent uncontrolled expansion, the massive proliferation of T cells during an acute immune response has to be followed by controlled deletion. Here we show that similar to Fas, perforin is not only an important effector molecule of cytotoxic T lymphocytes (CTL) but also involved in down-regulating peripheral T cells. Mice deficient for both the CTL effector molecule perforin and the apoptosis-inducing Fas ligand spontaneously develop infiltration of highly activated CD8(+) T cells in kidney and liver and die between 5 and 12 weeks of age. Injection of staphylococcal enterotoxin B (SEB) into perforin-deficient mice results in dramatically increased selective expansion and prolonged persistence of CD8(+), but not CD4(+), SEB-reactive T cells. Also, secondary immunization of TCR transgenic perforin-deficient mice with the lymphocytic choriomeningitis virus glycoprotein-derived epitope peptide leads to an increased proliferation of transgenic CD8(+) T cells, that is not explained by failure to deplete professional antigen-presenting cells. These results point to a novel mechanism of T cell homeostasis in which the acquisition of perforin-dependent cytotoxic activity regulates the expansion and persistence of CD8(+) effector T cells in vivo. PMID- 10540339 TI - Memory CD4 cells do not migrate into peripheral lymphnodes in the absence of antigen. AB - Memory T cells are thought to protect against previously encountered pathogens in part by preferentially recirculating through the lymphoid tissues where they were primed and where challenge with antigen (Ag) is likely to occur. In this study, we examined the distribution of memory CD4 cells after priming, and analyzed their capacity to localize in lymph nodes after transfer to normal and Ag-primed recipients. Immunization induced a high frequency of Ag-specific CD4 cells in the primary response in draining lymph nodes and spleen. Thereafter, the numbers in lymph nodes declined dramatically whereas frequencies in the spleen were unchanged, suggesting that memory CD4 cells primarily reside in or recirculate through the spleen. Indeed, memory CD4 cells, unlike naive CD4 cells, failed to home to lymph nodes after adoptive transfer to normal recipients and were detected predominantly in the spleen for extended periods, suggesting that recirculation through lymph nodes was limited. Memory cells also did not home to lymph nodes recipients in response to specific Ag, but subsequently, recruitment that could be blocked with monoclonal antibodies to CD44 and LFA-1 and was independent of naive cells did occur. The data indicate that memory and naive CD4 cells can be distinguished on the basis of their patterns of circulation. PMID- 10540341 TI - MHC class I-restricted epitope spreading in the context of tumor rejection following vaccination with a single immunodominant CTL epitope. AB - Epitope spreading is a process whereby epitopes distinct from and non-cross reactive with an inducing epitope become targets of an evolving immune response. This phenomenon has been associated most notably with the progression of naturally occurring or experimentally induced chronic autoimmune diseases. We have investigated the potential occurrence of epitope spreading in the context of antitumor cytotoxic T cell (CTL) responses using chicken ovalbumin (OVA) as a model antigen. Our results indicate that following rejection of OVA-expressing EG.7 tumor cells effectuated by a CTL response which is induced against the MHC class I-restricted immunodominant epitope OVA257-264, there occurs intramolecular diversification of the CTL response to two additional OVA-derived epitopes, OVA176-183 and OVA55-62, as well as intermolecular spreading to other endogenous tumor-derived determinants. It seems that CTL-mediated tumor cell destruction in vivo favors cross-presentation of additional epitopes with the consequent activation of additional tumor-reactive lymphocytes. The process of epitope spreading in that context has obvious important implications for the design of antigen-specific antitumor immunotherapies. PMID- 10540340 TI - CD1high B cells: a population of mixed origin. AB - A specialized subpopulation of T lymphocytes is reactive to the MHC class I-like molecule CD1d. It is not clear which cells are the major antigen-presenting cells in vivo in the activation of CD1-restricted immune responses. We have characterized a subset of B lymphocytes expressing six- to eightfold higher levels of CD1 than the bulk of B cells. The cells have a surface phenotype (CD21(high), CD23(low), IgM(high), IgD(low)) found previously to characterize B cells residing in the splenic marginal zones. CD1(high) B cells localize preferentially to the spleen, and appear late in ontogeny, at 3 - 4 weeks of age. The CD1(high) B cells were present in mice lacking conventional helper T cells, ruling out an exclusive origin from T cell-dependent immune responses. Still, some CD1(high) B cells had been involved in T cell-dependent immune responses as suggested by mutations in their rearranged immunoglobulin gene regions. The population could still be found in mice with severely reduced B cell reactivity to bacterial lipopplysaccharides (C3H / HeJ mice) and in mice unable to respond to thymus-independent type 2 antigens (NFR.Xid mice), as well as in germ-free mice, indicating that bacterial antigens are not major stimuli for the induction of CD1(high) B cells. In contrast, the CD1(high) B cell population was severely reduced in CD19-deficient mice. Taken together, the results imply that the CD1(high) population is heterogenous and of mixed origin, dependent for its development or maintenance on signaling through the CD19 molecule. PMID- 10540343 TI - Thymic dependence of invariant V alpha 14+ natural killer-T cell development. AB - Both thymic and extrathymic bone marrow (BM)-derived pathways for the development of CD1 reactive, Valpha14-Jalpha281(+) NK1.1(+) T cells have been suggested. In this report, we sought evidence for extrathymic NK-T cell development using two approaches. First, BM cells from gammac-deficient mice were examined for the presence of Valpha14-Jalpha281 transcripts. Since intrathymic NK-T cell selection is gammac independent, we predicted that gammac(-) BM cells should also harbor these specific TCRalpha chains. Second, Valpha14-Jalpha281 transcripts were analyzed in BM cells from lethally irradiated, thymectomized mice reconstituted with fetal liver hematopoietic precursors. All donor-derived T cell development in these chimeras is by definition extrathymic. In both cases, we failed to detect invariant Valpha14(+) TCRalpha chain transcripts. These experiments call into question the significance of an extrathymic pathway of development for Valpha14(+) NK1.1(+) CD1-reactive T cells. PMID- 10540342 TI - Role of Syk in Fc gamma receptor-coupled tyrosine phosphorylation of Cbl in a manner susceptible to inhibition by protein kinase C. AB - Fcgamma receptors (FcgammaR) of guinea pig neutrophils were ligated and anti-Cbl immunoprecipitates prepared therefrom were assayed for the associated protein tyrosine kinase activity, which increased upon ligation of FcgammaR. The increases were overcome upon activation of cellular protein kinase C by simultaneous addition of phorbol 12-myristate 13-acetate (PMA) to the ligated cells. Syk proved to be the most important tyrosine kinase bound to Cbl that served as the major substrate; essentially no tyrosine phosphorylation occurred in the anti-Cbl immunoprecipitates prepared from the cell lysate that had been depleted of Syk by prior immunoprecipitation with anti-Syk antibodies. Exposure of the (32)P-labeled cells to PMA resulted in phosphorylation of cellular Cbl on serine residues. Thus, protein kinase C-induced serine phosphorylation of Cbl suppressed its tyrosine phosphorylation by Syk as a result of tyrosine kinase inhibition by unknown mechanisms, leading to inhibition of Cbl-mediated signaling such as phosphatidylinositol 3-kinase activation. PMID- 10540344 TI - Negative regulation of antigen receptor-mediated signaling by constitutive association of CD5 with the SHP-1 protein tyrosine phosphatase in B-1 B cells. AB - CD5, a membrane-associated glycoprotein, has been shown to negatively regulate antigen receptor-mediated growth responses in peritoneal B lymphocytes, thymocytes and mature T cells. The CD5-expressing peritoneal B cells (B-1) that are normally unresponsive to B cell receptor (BCR)-mediated growth signals mount a proliferative response to BCR cross-linking if the CD5 gene is deleted or if the CD5 molecule is sequestered away from the BCR. SHP-1, a cytosolic protein tyrosine phosphatase, has also been implicated in the negative regulation of antigen receptor-mediated signaling. The present study shows that SHP-1 is constitutively associated with the BCR in B-1 cells. This association is mediated in part by CD5, as it is reduced substantially after antigen receptor ligation in CD5(-/-) B-1 cells, and upon sequestration of CD5 from the antigen receptor complexes in wild-type B-1 cells. Prior cross-linking of CD5 also restores a normal calcium mobilization response as well as NF-kappaB activation in B-1 cells. These data support a model whereby CD5 negatively regulates antigen receptor-mediated growth signals by recruiting SHP-1 into the BCR complex in B-1 cells. PMID- 10540345 TI - A MAGE-A4 peptide presented by HLA-A2 is recognized by cytolytic T lymphocytes. AB - The MAGE-encoded antigens that are recognized by cytolytic T lymphocytes (CTL) are shared by many tumors and are strictly tumor specific. Clinical trials involving therapeutic vaccination of cancer patients with MAGE antigenic peptides or proteins are in progress. To increase the range of patients eligible for therapy with peptides, it is important to identify additional MAGE epitopes. We have used a method to identify CTL epitopes, which selects naturally processed peptides. CD8(+) T cells, obtained from individuals without cancer, were stimulated with autologous dendritic cells infected with a recombinant adenovirus containing the MAGE-A4 coding sequence. Responder cell microcultures that specifically lysed autologous EBV-transformed B cells infected with vaccinia-MAGE A4 were cloned using autologous stimulator cells infected with a Yersinia enterocolitica carrying the MAGE-A4 sequence. An anti-MAGE-A4 CTL clone was obtained and the epitope was found to be decapeptide GVYDGREHTV (amino acids 230 239) presented by HLA-A2 molecules. The CTL clone lysed HLA-A2 tumor cells expressing MAGE-A4. This is the first reported antigenic peptide encoded by MAGE A4. It may be valuable for cancer immunotherapy because MAGE-A4 is expressed in 51% of lung carcinomas and 63% of esophageal carcinomas, whereas about 50% of Caucasians and Asians express HLA-A2. PMID- 10540347 TI - Autoimmune diseases developed in athymic nude mice grafted with embryonic thymus of xenogeneic origin. AB - Reconstitution of T cell functions in athymic BALB / c nude mice was investigated after transplantation of embryonic thymus grafts under the renal capsule (TG nude mice). Thymi were obtained from mice, rats, rabbits, hamsters, guinea pigs, swine and cows. All of the grafted thymi grew and formed proper thymic structures with host CD90(+) cells. The TCRalpha beta expression pattern of the lymphocytes in the grafted thymi was quite similar to that of normal mouse thymocytes. Sufficient CD4(+) populations were observed in the peripheral lymphoid organs of all groups of TG nude mice. Plaque-forming cell assay revealed that the TG nude mice had acquired considerable helper T cell functions. Histological and serological studies, however, showed multiple organ-localized inflammatory diseases in TG nude mice of all xenogeneic thymus groups, but not with syngeneic or allogeneic thymi. The organs affected were the thyroid, the lacrimal, submandibular and sublingual glands, the eyes, stomach and ovaries. The incidence of lesions was variable depending on the species of origin of the grafted thymus. Lesions from TG nude mice were successfully transferred into naive nude mice by host CD4(+) cells. These results together suggested that the microenvironment of grafted thymi, even if xenogeneic, is able to educate host T cell precursors. However, this reconstitution of functions does not always induce tolerance to certain autoantigens, resulting in development of multiple autoimmune lesions. PMID- 10540346 TI - Expansion of circulating V gamma 9/V delta 1 T cells in a patient with a syndrome of recurrent fever: evidence for an unusual antigen-driven process leading to selection of recurrent motifs within TCR junctional loops of diverse lengths. AB - Polyclonal expansions of human Vdelta1 T cells have been described in diverse physiopathological situations without strong TCR structural data for an antigen driven selection. Here, we have analyzed the phenotype and TCR repertoire of gamma delta T cells obtained from the peripheral blood of a 19-year-old patient with a syndrome of recurrent fever, which accounted for up to 40% of CD3(+) T cells and expressed predominantly Vgamma9 and Vdelta1 TCR regions and a memory phenotype. Sequence analysis of Vdelta1-Jdelta1 transcripts derived from peripheral blood lymphocytes (PBL) indicated that, while Vdelta1-Jdelta1 junctional sequences were diverse in length, all but one contained several recurrent motifs at conserved positions from both the 5'- and 3'-ends of the complementarity-determining region (CDR)3 loop. Analysis of gamma delta T cell clones derived from patient PBL demonstrated that Vgamma9(+) but not Vgamma9(-) T cell clones frequently expressed Vdelta1 chains with these characteristics and unveiled a hierarchy between the constraints imposed on the 5'- vs. the 3' motifs of the Vdelta1 CDR3 loops. These results constitute the first strong evidence for a nominal antigen-driven selection of Vdelta1 T cells in vivo and also suggest that the hierarchy of the constraints imposed by antigens respectively on the length and amino acid composition of TCR CDR3 loops differs between alpha beta and gamma delta T cells. PMID- 10540348 TI - Selection of conserved TCR VDJ rearrangements in chronic psoriatic plaques indicates a common antigen in psoriasis vulgaris. AB - Psoriasis vulgaris is a common HLA-associated inflammatory skin disease. Although its etiology is still unknown, it is thought to involve T cell-mediated inflammatory mechanisms. In examining the lesional psoriatic TCR beta chain (TCRB) usage in a pair of identical twins concordant for psoriasis, we observed repetitive TCR VDJ rearrangements which indicated antigen-specific oligoclonal T cell expansion. Several of these TCRB rearrangements were identical or highly homologous in the amino acid composition of the complementarity determining region 3 (CDR3), suggesting that T cells with these TCR might be important for disease manifestation. This conclusion was strengthened by TCR analysis of other psoriasis patients. Several repetitive lesional TCRB rearrangements were found that were similar to the conserved CDR3 seen in the twins. Since TCR antigen specificity is largely determined by the beta chain CDR3, selection of T cells with conserved TCRB CDR3 motifs could indicate the presence of a common antigen as a major target of the lesional psoriatic immune response. PMID- 10540349 TI - The interplay between T helper subset cytokines and IL-12 in directing human B lymphocyte differentiation. AB - This study asks how T helper (TH) subset cytokines impact upon IL-12-directed change in B cells engaged in signaling via the B cell receptor and CD40, essential components in the initiation of T-dependent B cell responses. For B cells stimulated in this way, IL-12 promoted a distinct phenotype highlighted by the hyper-expression of CD38: the Th1 cytokine IFN-gamma reproduced the IL-12 effects while neutralizing antibody to IFN-gamma reversed IL-12-dependent change. The divergent pathway of differentiation promoted by the Th2 cytokine IL-4 (characterized by hyper-induction of CD23) was left unchecked by IL-12. IL-10 was found to dampen IL-12 actions by suppressing IL-12-dependent IFN-gamma production but failed to perturb the effects of exogenous IFN-gamma. Thus, IL-12--by invoking autocrine IFN-gamma production--promotes phenotypic deviation in B cells engaging T-dependent signals. The reversal of such Th1 driving of B cells by IL 10 only when the source of IFN-gamma is endogenous and the inability of IL-12 to impact upon IL-4-directed differentiation suggest a progressive and hierarchical commitment of B cells to polarization during a developing T-dependent response dominated at the level of the Th cell rather than that of the dendritic cell. PMID- 10540351 TI - Combining DNA and protein vaccines for early life immunization against respiratory syncytial virus in mice. AB - Early life responses to respiratory syncytial virus (RSV)-F DNA and RSV-F protein immunization were studied in murine models of neonatal immunization. RSV-F DNA induced similar antibody (Ab) responses, antigen-specific IFN-gamma production and cytotoxic T lymphocyte (CTL) responses in 1-week-old and adult BALB / c mice. In contrast, RSV-F protein induced much higher IL-5 responses in early life. Both vaccines elicited Ab and CTL responses in spite of maternal Ab, but with distinctive kinetics. Sequential RSV-F DNA priming / protein boosting primed 1 week-old mice for RSV-F-specific CTL responses, reduced IL-5 production and enhanced Ab responses. In contrast, IL-5 exceeded IFN-gamma responses when young mice were primed with protein and boosted with DNA. Last, when protein and DNA immunization were combined, a single vaccine dose induced early Ab responses, preferential IL-5 responses but strong CTL responses. Sequential or combined DNA / protein immunization thus represent interesting strategies for early life immunization. PMID- 10540350 TI - Modulation of E2F activity in primary mouse B cells following stimulation via surface IgM and CD40 receptors. AB - Since signals via CD40 and the B cell receptor are known to synergize to induce B cell activation, we have analyzed the pocket protein/E2F complexes in mouse B lymphocytes following stimulation by anti-IgM, anti-CD40, alone or together. We find that E2F4 and DP1 form the predominant E2F heterodimers in the G0 and G1 phases of the cell cycle, complexed with hypophosphorylated p130. During late G1 and S phase this complex is replaced by at least three different E2F complexes, one of which is an E2F complex containing p107 or pRB as well as two "free" E2F complexes consisting of E2F4/DP1 and E2F1-3/DP1. These effects were mirrored by the levels and phosphorylation status of the three pocket proteins. We also observed an increase in electrophoretic mobility of DP1 and E2F4 as B cells progressed from G0 into early G1, resulting from their dephosphorylation. This is known to correlate with a decrease in DNA binding capacity of these proteins and could also be important for derepression of genes negatively regulated through E2F sites in their promoters. These results therefore indicate that the pRB/E2F pathway integrates proliferative signals emanating from the sIgM and CD40 receptors. PMID- 10540352 TI - Domain deletions in the human polymeric Ig receptor disclose differences between its dimeric IgA and pentameric IgM interaction. AB - The human polymeric Ig receptor (pIgR), or transmembrane secretory component, is basolaterally expressed on secretory epithelial cells; its function is to transport externally J chain-containing dimeric IgA and pentameric IgM. The ligand-binding extracellular part of this receptor contains five disulfide stabilized domains which show considerable homology with the variable domains of Ig chains. The N-terminal domain 1 (D1) mediates the initial noncovalent ligand interaction. In this study we made deletions of the human pIgR D2 and D3 (pIgRDelta2,3), or D4 and D5 (pIgRDelta4,5), to investigate the influence of these domains in receptor binding and transport of dimeric IgA and pentameric IgM across MDCK cells transfected with the truncated receptors. Both mutants were found to bind pentameric IgM, but only pIgRDelta4,5 bound dimeric IgA. These results showed that the two ligands interact differently with human pIgR; binding of pentameric IgM apparently depends fully on strong interactions with D1, while binding of dimeric IgA in addition depends on elements within D2 and / or D3 to support the initial noncovalent binding to D1. Moreover, our studies imply that dimeric human IgA binds differently to pIgR from various species. This observation cautions against interpretation of functional studies performed with non-homologous receptor-ligand pairs. PMID- 10540353 TI - Autoimmune insulitis and diabetes in the absence of antigen-specific contact between T cells and islet beta-cells. AB - Autoimmune diabetes develops following recognition of organ-specific antigens by T cells. The disease begins with peri-islet infiltration by mononuclear cells, proceeds with insulitis and becomes manifest with destruction of insulin producing islet beta-cells. T cells are necessary to induce insulitis and diabetes, but it is not clear by what mechanisms they can do so, i. e. whether the T cells need to make antigen-specific contact with the beta-cell or whether other interactions are sufficient to induce beta-cell death. In the present study we have constructed chimeric mice in which the bone marrow-derived antigen presenting cells, but not the islet beta-cells, are capable of presenting antigen to monospecific T cells. We show that both insulitis as well as beta-cell destruction can proceed in the absence of islet beta-cell surface antigen recognition by T cells. Our results support the notion that diabetes can be caused by distinct effector mechanisms. PMID- 10540354 TI - L-type calcium channels in the photoreceptor ribbon synapse: localization and role in plasticity. AB - Calcium (Ca(2+)) influx through voltage-gated Ca(2+)channels stimulates a variety of neural activities, including process outgrowth, neurotransmission, and synaptic plasticity. In general, L-type channels control Ca(2+) influx into the soma and dendrites, whereas other Ca(2+) channel types control presynaptic activities. Neurons that make ribbon synapses, however, are among a select group of nerve cells whose presynaptic Ca(2+)-dependent secretion is linked to L-type channels. Recently, photoreceptor ribbon synapses have been shown to be capable of dramatic structural remodeling and neuritic outgrowth. Here, we have examined 1) the distribution of dihydropyridine (DHP)-sensitive (L-type) Ca(2+) channels in photoreceptor presynaptic structures and 2) the role of these channels in axonal plasticity and process outgrowth in culture. Using anti-alpha(1C) and the fluorescent dihydropyridine, (-)-DM-BODIPY DHP, L-type channels were localized in the outer plexiform layer of retinal sections and in presynaptic terminals of freshly isolated photoreceptors. In the rod terminal, dense patches of label were present; their distribution and number matched that of synaptic ribbons. After 1 7 days in vitro, punctate alpha(1C) staining occurred along newly formed neurites and presynaptic varicosities. Functional channels were present throughout the culture period, as determined by fura-2 imaging. Channel blockage by nicardipine, a DHP antagonist, inhibited axonal remodeling. Specifically, it prevented axon retraction and lamellipodium formation, reduced neurite growth, and produced long, thin processes on some, primarily cone, photoreceptors. L-type Ca(2+) channel activity, therefore, not only stimulates neurotransmission but contributes to presynaptic structural plasticity at the ribbon synapse. PMID- 10540356 TI - Cellular heterogeneity in cerebral cortex: a study of the morphology of pyramidal neurones in visual areas of the marmoset monkey. AB - The morphological characteristics of the basal dendritic fields of layer III pyramidal neurones were determined in visual areas in the occipital, parietal, and temporal lobes of adult marmoset monkeys by means of intracellular iontophoretic injection of Lucifer yellow. Neurones in the primary visual area (V1) had the least extensive and least complex (as determined by Sholl analysis) dendritic trees, followed by those in the second visual area (V2). There was a progressive increase in size and complexity of dendritic trees with rostral progression from V1 and V2, through the "ventral stream," including the dorsolateral area (DL) and the caudal and rostral subdivisions of inferotemporal cortex (ITc and ITr, respectively). Neurones in areas of the dorsal stream, including the dorsomedial (DM), dorsoanterior (DA), middle temporal (MT), and posterior parietal (PP) areas, were similar in size and complexity but were larger and more complex than those in V1 and V2. Neurones in V1 had the lowest spine density, whereas neurones in V2, DM, DA, and PP had similar spine densities. Neurones in MT and inferotemporal cortex had relatively high spine densities, with those in ITr having the highest spine density of all neurones studied. Calculations based on the size, number of branches, and spine densities revealed that layer III pyramidal neurones in ITr have 7.4 times more spines on their basal dendritic fields than those in V1. The differences in the extent of, and the number of spines in, the basal dendritic fields of layer III pyramidal neurones in the different visual areas suggest differences in the ability of neurones to integrate excitatory and inhibitory inputs. The differences in neuronal morphology between visual areas, and the consistency in these differences across New World and Old World monkey species, suggest that they reflect fundamental organisational principles in primate visual cortical structure. PMID- 10540355 TI - Zonal organization of the ventral lateral geniculate nucleus in the cat: cholera toxin mapping. AB - A zonal organization of the ventral lateral geniculate nucleus (LGNv) in the cat was studied with the bidirectional tracing method by using the subunit b of cholera toxin (CTb) as the marker. The prime objectives of the present study were to examine precise distribution of terminals of the retinal afferents to the LGNv, and to correlate it with the origins and terminations of the other central connections of the nucleus. The results obtained are summarized as follows: (1) the LGNv of the cat is divided into 3 zones according to the terminations of retinal afferents: lateral, intermediate, and medial; (2) the lateral zone receives afferents from the retina in which ipsi- and contralateral fibers terminate in a complementary fashion. According to the density of labeling of retinal terminals, the lateral zone is further divided into several areas. It also receives fibers from the visual related cortex; (3) the intermediate zone, which does not receive fibers from the retina nor the visual cortex, is reciprocally connected with the midbrain, primarily with the superficial layers of the superior colliculus (SC), and gives rise to thalamic projections to the nucleus centralis lateralis bilaterally; and (4) the medial zone, from which commissural fibers arise, receives afferents bilaterally from the retina, and sends fibers to the SC bilaterally. These results suggest that the LGNv of the cat has three different zones. Functional participation of each zone has been discussed. PMID- 10540358 TI - Efferent neurons and specialization of abdominal segments in grasshoppers. AB - A specialized behavior, oviposition, is produced by the eighth and ninth abdominal segments of female grasshoppers. To begin to understand how these segments produce the behavior, which is not displayed by males or pregenital regions of the abdomen in females, the structure and function of efferent neurons in abdominal ganglia of both sexes were examined. In females, the eighth and ninth segments are specialized differently for oviposition: 20 ovipositor motor neurons were found in the eighth segment, and 26 were found in the ninth segment. Males had fewer motor neurons in their eighth segment, but the same number in the ninth segment, which is the only genital segment in males. However, the axons of several of the ninth segmental male motor neurons traveled to the periphery in the genital nerve, which is only found in males. In both sexes, pregenital ganglia had the most motor neurons, but these neurons, for the most part, had morphologies that strongly resembled those of genital segments. Efferent modulatory neuron numbers were not sexually dimorphic in the segments examined, except that males had a greater number in their ninth segment. Experimental methods that activate oviposition were found to also activate a rhythmical motor pattern in pregenital abdominal segments of both sexes. In females, the pattern was phase-coupled to oviposition, but persisted after the connections with the terminal abdominal ganglion were severed. The preponderance of similarities among efferent neurons and elicited motor activity suggests a common pattern of neural circuitry in the behaviorally diverse abdominal segments of grasshoppers. PMID- 10540357 TI - Glutamate carboxypeptidase II is expressed by astrocytes in the adult rat nervous system. AB - The enzyme glutamate carboxypeptidase II (GCP II) has been cloned from rat brain and human prostate. This enzyme, which catabolizes the neuropeptide N acetylaspartylglutamate, has also been known as N-acetylated alpha-linked acidic dipeptidase (NAALADase), and is identical to the prostate-specific membrane antigen and to the jejunal folylpoly-gamma-glutamate carboxypeptidase. The goals of the present study were to elucidate the cell specificity and regional pattern of GCP II expression in the rat nervous system by using Northern blots and enzymatic assays of brain and subfractionated primary neuronal and glial cultures together with in situ hybridization histochemistry (ISHH) in sections of adult rat tissue. GCP II activity was assayed in astrocyte cultures (4.4 pmol/mg protein per minute), neuronal-glial cocultures (2.5 pmol/mg protein per minute) and neuron-enriched cultures (0.38 pmol/mg protein per minute), with the activity in each preparation correlating to its astrocytic content (r = 0.99). No activity was detected in cultured oligodendrocytes or microglia. Northern blots probed with a GCP II cDNA detected mRNAs exclusively in activity-positive cell preparations. ISHH results show that GCP II is expressed by virtually all astrocytes, by Bergmann glial cells in cerebellum, by Muller cells in retina and by the satellite cells in dorsal root ganglia. Astrocytes in select groups of nuclei (e.g., habenula, supraoptic nucleus, pontine nucleus) contained pronounced levels of GCP II message. The data of the present study suggest that GCP II is expressed in the adult rat nervous system exclusively in astrocytic glial cells. PMID- 10540359 TI - Immunocytochemical localization of cannabinoid CB1 receptor and fatty acid amide hydrolase in rat retina. AB - Cannabinoids have major effects on central nervous system function. Recent studies indicate that cannabinoid effects on the visual system have a retinal component. Immunocytochemical methods were used to localize cannabinoid CB1 receptor immunoreactivity (CB1R-IR) and an endocannabinoid (anandamide and 2 arachidonylglycerol) degradative enzyme, fatty acid amide hydrolase (FAAH)-IR, in the rat retina. Double labeling with neuron-specific markers permitted identification of cells that were labeled with CB1R-IR and FAAH-IR. CB1R-IR was observed in all cells that were protein kinase C-immunoreactive (rod bipolar cells and a subtype of GABA-amacrine cell) as well as horizontal cells (identified by calbindin-IR). There was also punctate CB1R-IR in the distal one third of the inner plexiform layer (IPL) that could not be assigned to a cell type. FAAH-IR was most prominent in large ganglion cells, whose dendrites projected to a narrow band in the proximal IPL. Weaker FAAH-IR was observed in the soma of horizontal cells (identified by calbindin-IR); the soma of large, but not small, dopamine amacrine cells (identified by tyrosine hydroxylase-IR); and dendrites of orthotopic- and displaced-starburst amacrine cells (identified by choline acetyltransferase-IR) but in less than 50% of the starburst amacrine cell somata. The extensive distribution of CB1R-IR on horizontal cells and rod bipolar cells indicates a role of endocannabinoids in scotopic vision, whereas the more widespread distribution of FAAH-IR indicates a complex control of endocannabinoid release and degradation in the retina. PMID- 10540360 TI - Trigeminal projections to hypoglossal and facial motor nuclei in the rat. AB - This study was undertaken to identify the trigeminal nuclear regions connected to the hypoglossal (XII) and facial (VII) motor nuclei in rats. Anterogradely transported tracers (biotinylated dextran amine, biocytin) were injected into the various subdivisions of the sensory trigeminal complex, and labeled fibers and terminals were searched for in the XII and VII. In a second series of experiments, injections of retrogradely transported tracers (biotinylated dextran amine, gold-horseradish peroxidase complex, fluoro-red, fluoro-green) were made into the XII and the VII, and labeled cells were searched for in the principal sensory trigeminal nucleus, and in the pars oralis, interpolaris, and caudalis of the spinal trigeminal nucleus. Trigeminohypoglossal projections were distributed throughout the ventral and dorsal region of the XII. Neurons projecting to the XII were found in all subdivisions of the sensory trigeminal complex with the greatest concentration in the dorsal part of each spinal subnucleus and exclusively in the dorsal part of the principal nucleus. Trigeminofacial projections reached all subdivisions of the VII, with a gradual decreasing density from lateral to medial cell groups. They mainly originated from the ventral part of the principal nucleus. In the spinal nucleus, most of the neurons projecting to the VII were in the dorsal part of the nucleus, but some were also found in its central and ventral parts. By using retrograde double labeling after injections of different tracers in the XII and VII on the same side, we examined whether neurons in the trigeminal complex project to both motor nuclei. These experiments demonstrate that in the spinal trigeminal nucleus, neurons located in the pars caudalis and pars interpolaris project by axon collaterals to XII and VII. PMID- 10540361 TI - Region-specific mRNA expression of phosphatidylinositol 3-kinase regulatory isoforms in the central nervous system of C57BL/6J mice. AB - Activation of phosphatidylinositol 3-kinase (PI 3-kinase) by receptor tyrosine kinases for growth factors is crucial for neuronal cell survival and proliferation. This class of kinases is comprised of heterodimers, each consisting of one regulatory and one catalytic subunit. Multiple isoforms of regulatory subunits exist, including p85alpha and its alternative splice products p50alpha and AS53/p55alpha, and p85beta and p55(PIK), which are derived from different genes. The regional distribution of these PI 3-kinase regulatory isoforms was mapped in the adult murine brain by in situ hybridization histochemistry. All isoforms were demonstrated in abundance in choroid plexus and anterior pituitary. In neuronal compartments, however, PI 3-kinase isoforms were distributed in a regionally specific manner. In general, the mRNAs for p85alpha, p50alpha, AS53, and p85beta were widespread, with the highest level in the olfactory system, in neuronal groups of the forebrain and hypothalamus, in the hippocampus, cortex, inferior and superior colliculus, pituitary, and cerebellum. However, each isoform had specific variations. Lower expression levels of these isoforms were found in the thalamus, diencephalon, mesencephalon, and brainstem. In contrast, abundant mRNA expression of p55(PIK) was limited to cerebellum and anterior pituitary, with moderate levels of p55(PIK) in the olfactory bulb and hippocampus and low levels elsewhere. The distribution pattern of PI 3-kinase isoforms in the brain indicates pluripotent signaling properties for PI 3-kinase isoforms p85alpha, p50alpha, AS53/p55alpha, and p85beta for a variety of receptor tyrosine kinases, whereas the restricted expression of p55(PIK) implies a regionally specific role for this isoform in neuronal signaling. The unique integrated expression profiles of PI 3-kinase isoforms in distinct neuronal compartments denote complex intracellular signaling pathways for each neuronal region to ensure specificity of receptor tyrosine kinase signal transduction. PMID- 10540363 TI - Erratum: rapid extension of axons into the CA3 region by adult-generated granule cells. J comp neurol 413:146-154 AB - Hastings NB, Gould E. 1999. Rapid extension of axons into the CA3 region by adult generated granule cells. J Comp Neurol 413:146-154. In the Acknowledgments section of the above article, Patima Tanapat's name was incorrectly printed as Patrick Tanapat. The acknowledgment should read as follows: We gratefully acknowledge Patima Tanapat for helpful comments in the manuscript and Joseph Goodhouse for assistance with confocal imaging. The Publisher regrets the error. PMID- 10540362 TI - Stimulant-induced exocytosis from neuronal somata, dendrites, and newly formed synaptic nerve terminals in chronically decentralized sympathetic ganglia of the rat. AB - Loss of preganglionic neurones underlies the autonomic failure of human multiple system atrophy. In rat sympathetic ganglia decentralization leads to new synapse formation. We explored whether these synapses are functional, and whether chronically decentralized neurones respond normally to activation, in terms of exocytosis. Potassium depolarization and cholinergic agonists were applied to freshly excised rat superior cervical sympathetic ganglia, preganglionically denervated with prevented reinnervation 5 months earlier. Ganglia were incubated and stimulated in the presence of tannic acid, which stabilizes released vesicle cores for subsequent electron microscopy. In denervated ganglia exocytosis was observed from newly formed synaptic nerve terminals, and from nonsynaptic surfaces of neurone somata and dendrites. The results demonstrated that the new intraganglionic synapses, which are mostly catecholaminergic, can function and that chronically decentralized sympathetic neurones remain capable of stimulant induced exocytosis from somata and dendrites. The maximal release upon potassium depolarization did not differ significantly between denervated and contralateral ganglia. Relative to this, the exocytotic responses of decentralized somata and dendrites to nicotine resembled those of contralateral ganglia. Responses to muscarine were significantly less in denervated than in contralateral ganglia, indicating inhibition in dendrites. Responses to carbachol suggested interactions between nicotinic and excitatory muscarinic effects. Nerve terminals in denervated ganglia showed high basal release. Their responses to muscarine and carbachol resembled those of the decentralized neurones, from which most may originate. Their response to nicotine evidenced inhibition. Their actions, coupled with nonsynaptic effects of soma-dendritic exocytosis, might modulate responses of the decentralized neurone population to other surviving inputs. This modulation could be influential in disease-induced decentralization in man. PMID- 10540364 TI - Treatment of Guillain-Barre syndrome: a cost-effectiveness analysis. AB - Acute Guillain-Barre syndrome is the most common cause of neuromuscular paralysis. Plasma exchange and intravenous immune globulin (IV IgG) are both effective treatments for this condition and the purpose of this report was to compare the cost-effectiveness of these two modalities. A MEDLINE search was performed to identify randomized studies that compared the use of IV IgG and plasma exchange for treatment of acute Guillain-Barre syndrome to determine if one modality was more effective and/or safer for the management of this condition. A decision analysis was structured around the alternatives facing neurologists who must choose a treatment regimen for patients diagnosed with acute Guillain-Barre syndrome who require active therapy. Cost information was obtained directly from product manufacturers and hospital sources. Two head-to head trials comparing the effectiveness of plasma exchange and IV IgG for treatment of acute Guillain-Barre syndrome determined that there was insufficient evidence to suggest one therapy was more effective than the other; therefore, a cost minimization analysis was performed. The costs per patient of plasma exchange and IV IgG for the treatment of acute Guillain-Barre syndrome were $6,204 and $10,165, respectively. A sensitivity analysis determined the model was sensitive to the cost of IV IgG. The cost savings per patient treatment for the use of plasma exchange varied from $304 to $6,625 depending on the IV IgG product selected. Plasma exchange and IV IgG are both effective treatments for Guillain Barre syndrome. However, our analysis determined plasma exchange on average was almost $4,000 less costly per patient than IV IgG. Further research is required to determine the impact of patient and physician preferences on the treatment of this disorder. PMID- 10540365 TI - Pentastarch as partial replacement fluid for therapeutic plasma exchange: effect on plasma proteins, adverse events during treatment, and serum ionized calcium. AB - We compared the safety, effect on plasma proteins, and contribution to hypocalcemic toxicity of 10% pentastarch vs. 5% albumin in plasma exchange. Thirty-two neurology patients underwent 161 plasma exchange procedures. Subjects were randomized to receive either 10% pentastarch (n = 17) or 5% albumin (n = 15) as the first (1/2) of colloid replacement. The second (1/2) of colloid replacement was 5% albumin in all cases. NaCl (plus small amounts of ACD-A) accounted for (1/4) of the total return fluid. Mean total exchange volume was 3,842.6 +/- 450 ml. Hemoglobin, platelet count, coagulation parameters, and plasma fibrinogen were similar between the two groups at the outset of plasma exchange and at the time of the last (4th or 5th) procedure of the series. Immunoglobulin levels were equivalent at the outset and were reduced by plasma exchange to the same extent in the two groups. Serum albumin concentration fell significantly faster in the group receiving pentastarch. On the other hand, serum calcium, corrected for the albumin concentration, fell significantly further in the control group than in the group receiving pentastarch. Mean serum ionized calcium fell approximately 25 % in procedures using only albumin but only approximately 16% when pentastarch was used (P < 0.0001). This was reflected in the occurrence of hypocalcemic toxicity in 33.3% of procedures performed using only albumin but in only 8.1% of those using pentastarch (P = 0.0002). Pentastarch may be suitable for partial colloid replacement in plasma exchange and would have saved our hospital $127,050 to $434,280 in 1998 if used in all procedures. PMID- 10540366 TI - Erythrocytapheresis for chronically transfused children with sickle cell disease: an effective method for maintaining a low hemoglobin S level and reducing iron overload. AB - Cerebrovascular accident (CVA) is a major complication of sickle cell disease during childhood. Long-term transfusion reduces the hemoglobin S level and generally prevents recurrent stroke, but it also results in progressive iron overload that requires regular chelation therapy. Erythrocytapheresis offers an alternative approach aimed at reducing the iron accumulation. We reviewed the results of erythrocytapheresis in eight sickle cell patients (mean age of 12.1 years) at high risk for a first or recurrent stroke. They were maintained at the standard pre-transfusion hemoglobin S (Hb S) level of 30%. Over an average of 9 months of erythrocytapheresis, none of the patients developed complications related to the procedure or to the increased blood use. Ferritin levels decreased by a mean of 26.5% in all patients. When evaluating the ferritin level in five patients, who remained on chelation therapy with deferoxamine (DFO), the level dropped by a mean of 32%. The levels remained stable in the three patients who were not on DFO. The procedure is safe and effective in reducing iron overload and can obviate the need for chelation therapy, even when the target Hb S is maintained at the standard 30% range. PMID- 10540367 TI - Experience of double filtration plasmapheresis in the treatment of Guillain-Barre syndrome. AB - Therapeutic plasma exchange (TPE) is a standard treatment in Guillain-Barre syndrome. TPE may require exogenous fluid for replacement of plasma and, depending on the equipment used, varying extracorporeal volumes. Potential adverse effects include allergic reaction, infection, and hypotension. From September 1993 to December 1997, we treated 16 patients with Guillain-Barre syndrome by a newly developed method of automated double filtration plasmapheresis (DFPP). Patients (ten males and six females, age ranged from 16 to 73) suffering from acute ascending motor weakness and fulfilling the diagnostic criteria for GBS were chosen for DFPP. Each patient received at least five sessions of apheresis in 7 to 10 days and approximately 2.5 to 3.0 L of plasma was treated in each session. Patients were evaluated by disability grade according to a Hughes scale. The mean grade of disability was 3.62 at treatment and improved to 2.37 four weeks after the start of DFPP. The median time to grade 2 (walk without support) was 19 days. There were five patients (41.6%) in need of respirator support. The median time to weaning off the respirator was 9 days. Only two patients (12.5%) could not reach grade 2 at the end of 6 months. Our results were comparable to previously published results of TPE. We conclude that DFPP may be as effective as TPE in the treatment of GBS. PMID- 10540369 TI - Risk of transmission of Creutzfeldt-Jakob disease by transfusion of blood, plasma, and plasma derivatives. AB - Studies in experimental animals and case-reports of transmission of Creutzfeldt Jakob Disease (CJD) by blood transfusion or by albumin products have raised the possibility that CJD may be transmitted by transfusion. The risk of transmission of CJD by transfusion remains theoretical, since no confirmed case of CJD has ever been causally attributed to the receipt of a blood transfusion, no confirmed case of CJD has developed in recipients of clotting factor concentrates, and no cluster of CJD cases has been reported following the administration of a pooled plasma derivative to which a donor who subsequently developed CJD had contributed. However, based on a review of the hitherto available data, it is impossible to conclude at this time that CJD is not transmitted by blood or plasma transfusion or by the administration of pooled plasma derivatives. This review discusses the findings of the animal experiments and the human studies that investigated the potential for transmission of CJD among humans by transfusion, and explains the statistical difficulties associated with proving the negative hypothesis that CJD is not transmitted by transfusion. PMID- 10540368 TI - Intravascular stents do not cause microangiopathic hemolysis or thrombotic microangiopathy. AB - An increased incidence of TTP has been noted among patients receiving intravascular stents to improve patency in diseased coronary, renal, and peripheral arteries. Placement of transjugular intrahepatic porto-systemic shunt stents is often associated with subsequent development of severe hemolysis. We have prospectively studied the development of microangiopathic hemolysis or TTP in patients undergoing intravascular stent placement for peripheral vascular or renal artery disease. Hemolysis was evaluated both before and after stent placement by measuring complete blood count, total bilirubin, lactate dehydrogenase (LDH), haptoglobin and reticulocyte count, and examining peripheral blood films of all patients. Coagulation parameters, blood urea nitrogen and creatinine were measured to exclude disseminated intravascular coagulation or thrombotic thrombocytopenic purpura as a potential cause of hemolysis. Seventeen patients (median age 69 years) were evaluated. One patient was on ticlopidine. Mean hematocrit fell from 41.8% pre-stenting to 35.5% post-stenting (P = 0.003) but without significant change in reticulocyte count (1.7 vs. 1.6%, P = 0.605), LDH (546 vs. 560 IU/l; P = 0.836), bilirubin (0.62 vs. 0.63 mg/dl; P = 1.0), or haptoglobin (183 vs. 158 mg/dl; P = 0.083). Thus, this drop in hematocrit could not be attributed to hemolysis. Peripheral blood films revealed fewer than 1% schistocytes before and after stent placement in all cases. Absence of significant changes in mean platelet count (240 vs. 210 x 10(9)/L; P = 0.088), fibrinogen (385 vs. 378 mg/dl; P = 0.789), BUN (24.5 vs. 16.8; P = 0.079), and creatinine (1.38 vs. 1.24; P = 0.757) argue against development of TTP or DIC resulting from stent placement. No patient developed new renal impairment, a neurological syndrome, or unexplained fever after stent placement. At a mean of 6 weeks follow-up after stent placement, patients have not developed signs of hemolytic anemia or worsening renal function. Our findings argue against a primary risk of microangiopathic hemolytic anemia or TTP due to intravascular stents in patients not receiving ticlopidine. PMID- 10540370 TI - Therapeutic plasma exchange for acute inflammatory demyelinating syndromes of the central nervous system. AB - Idiopathic inflammatory demyelinating diseases (IIDDs) of the central nervous system, of which multiple sclerosis is the prototype, represent a family of monophasic, recurrent or progressive diseases with overlapping clinical and pathological manifestations. While most patients recover spontaneously or following a brief course of high-dose corticosteroids, occasional patients, particularly those with fulminant severe IIDDs, such as the Marburg variant, do not respond to corticosteroids and have severe, residual neurological deficits. While it is widely believed that IIDDs are mediated by T lymphocytes, as is experimental allergic encephelomyelitis, additional, possibly humoral, factors may be essential to generate the extensive demyelination seen in these conditions. Anecdotal reports over the past two decades have suggested that patients with acute, severe neurological deficits resulting from IIDDs, who fail to improve after high-dose intravenous corticosteroids, may benefit from plasma exchange. A randomized, sham-controlled, crossover study has recently been completed at the Mayo Clinic, which addresses these observations. PMID- 10540371 TI - Clinical features, evaluation, and treatment of patients with polyneuropathy associated with monoclonal gammopathy of undetermined significance (MGUS). AB - A number of common disorders of the peripheral nervous system are closely linked to a monoclonal gammopathy. In a minority of patients, the neuropathy represents the sentinel feature of a malignant plasma cell dyscrasia, such as multiple myeloma or its osteosclerotic variant, Waldenstrom's disease, amyloidosis, cryoglobulinemia or lymphoma; the vast majority have so-called "monoclonal gammopathy of undetermined significance" (MGUS). Sensory symptoms predominate with paresthesias, numbness, imbalance, and gait ataxia. Electrodiagnostic studies show mixed demyelinating and axonal features and often may be indistinguishable from findings in chronic inflammatory demyelinating polyneuropathy. Some have a pure axonal polyneuropathy, and in these patients the relationship to the paraprotein is less certain. With limited success, correlations have been made between the immunoglobulin type (IgM, IgG, or IgA) and the clinical and electromyographic characteristics of the neuropathy. The treatment of MGUS neuropathies poses a considerable challenge. Patients with IgG/IgA-MGUS have improved with corticosteroids or intravenous immune globulin. Only the benefit of plasma exchange has been substantiated in a controlled trial. The IgM neuropathies tend to be more refractory but often improve with similar regimens, particularly if cytotoxic agents are added in doses sufficient to reduce the amount of the M-protein. In addition to plasma exchange, chlorambucil, and cyclophosphamide, interferon-alpha is a novel therapy that holds promise for patients with IgM neuropathies associated with anti-myelin associated antibodies. PMID- 10540372 TI - Transfer of EBV-specific CTL to prevent EBV lymphoma post bone marrow transplant. AB - EBV-associated lymphoproliferative disorders (EBV-LPD) are a significant problem after hemopoietic stem cell transplantation from unrelated donors or mismatched family members. Risk factors include T-cell depletion, MHC mismatch, and intensity of immunosuppression. New therapeutic strategies involve cellular immunotherapy approaches and both donor T-cells and EBV-specific cytotoxic T lymphocytes (CTLs) have proven to be effective therapies. EBV-specific CTL has also proved to have a major impact on the incidence of this complication when used prophylactically. PMID- 10540373 TI - Coordination Chemistry of Aluminum, Gallium, and Indium at Transition Metals. AB - The current upswing in the interest in organoelement chemistry of Group 13 metals is attributed not least to the establishment of the coordination chemistry of R(a)E fragments (E=Al, Ga, In; a=1, 2) at d-block metals (M). Recently the availability of low-valent organoelement compounds as building blocks for synthesis has substantially enriched the structure chemistry of this class of compounds. The M-E bonding conditions and the question of the significance of M(d(pi))-E(p(pi)) backbonding as well as potential applications in materials science, for example, as single-source precursors for the deposition of thin intermetallic films by chemical vapor deposition, are discussed. PMID- 10540374 TI - Sleep and Its Modulation by Drugs That Affect GABA(A) Receptor Function. AB - Most sleeping pills are made up of chemical compounds that are ligands for allosteric modulatory binding sites on the GABA(A) receptor. Polysomnographic studies demonstrate that these hypnotics effectively increase the ability to fall and to stay asleep, but disrupt the physiological sleep profile (typical electroencephalograms (EEG) for the awake state and the different states of sleep are shown). Hence, there is an urgent need for sleep-promoting substances with a different mechanism of action. GABA analogues are one class of promising molecules. PMID- 10540375 TI - DNA Microarrays. AB - The complete human genes (ca. 100 000) as well as the whole spectrum of biological diversity should soon be able to be analyzed simultaneously by means of DNA microarrays using the fast technical advances that are occurring in this area. The particular strength of array analysis, typically based on the hybridization of nucleic acid probes attached to microchips with labeled RNA or DNA samples, results from the highly redundant measurement of many parallel hybridization events (see picture), which leads to an extraordinary level of assay validation. PMID- 10540376 TI - Self-Assembled, Sub-Micrometer Diameter Semipermeable Capsules. AB - Semipermeable nanoscopic hollow spheres or cages with diameters in the sub micrometer to the 100 nm range are accessible by the self-assembly of polymers onto sub-micrometer sized particles (Mohwald et al.) or by the formation of shell cross-linked polymeric micelles (Wooley et al.) followed by chemical degradation of the underlying particle or micelle core (see scheme). PMID- 10540377 TI - Asymmetric Catalytic Aminoalkylations: New Powerful Methods for the Enantioselective Synthesis of Amino Acid Derivatives, Mannich Bases, and Homoallylic Amines. AB - Modern methodologies such as the design of chiral catalysts using combinatorial techniques play a key role in the development of asymmetric aminoalkylations. Thus, powerful enantioselective syntheses for many attractive target compounds using chiral Lewis acid or base catalysts like 1 or 2 were created. PMID- 10540378 TI - Planar-Tetracoordinate Carbon in a Neutral Saturated Hydrocarbon: Theoretical Design and Characterization. AB - Exact planarity at the central carbon atom is achieved, according to molecular orbital calculations, in the strained polycyclic cage hydrocarbon dimethanospiro[2.2]octaplane (see structure). There are no glaringly long C-C bonds, which might have reflected inherent instability in this molecule that is yet to be synthesized. PMID- 10540379 TI - meso Myths: What Drives Assembly of Helical versus meso- AB - Both a triple helix as well as a meso complex are formed by the Ga(III) and Al(III) complexes with a bis-bidentate bis-hydroxypyridinone ligand H(2)L. The two forms are in equilibrium in solution, though formation of the helical structure in the presence of water, which as guest molecule finds sufficient space in the cavity of the helix, is favored (the structure of the helical H(2)O subset[Al(2)L(3)] complex is shown). PMID- 10540380 TI - Self-Assembly of a Three-Dimensional AB - A near trigonal antiprism with metal-metal distances in the nanometer regime is formed by the six metal ions in the crystalline, homochiral [Ga(6)(L(2))(6)] (see structure). This metal-ligand "cylinder" is based on a threefold symmetric, beta diketone ligand, and represents a new geometry for metal-ligand clusters. PMID- 10540381 TI - "Inverted" Solvent Effect on Charge Transfer in the Excited State. AB - Faster in cyclohexane than in acetonitrile is the fluorescence quenching of the azoalkane 2,3-diazabicyclo[2.2.2]oct-2-ene (DBO) by amines and sulfides. Although this photoreaction is induced by charge transfer (CT; see picture) and exciplexes are formed, the increase in the dipole moment of the exciplex is not large enough to offset the solvent stabilization of the excited reactants, and an "inverted" solvent effect results. PMID- 10540382 TI - An Inorganic Tire-Tread Lattice: Hydrothermal Synthesis of the Layered Vanadate AB - Two distinct V(9)O(23) building blocks having different vanadium coordination environments are intergrown in the unusual layered vanadate [N(CH(3))(4)](5)V(18)O(46) (see picture). Lattice strain is relieved by alternation of the vanadate strips formed from the blocks in both directions (akin to a three-dimensional tire tread). The formation of this lattice has implications for the assembly mechanism of this compound and related materials in hydrothermal synthesis. PMID- 10540383 TI - Combinatorial Methods for the Synthesis of Aluminophosphate Molecular Sieves. AB - Automatic dispensing of reagents into autoclave blocks followed by synthesis, isolation, and automated structure analysis with X-ray diffractometry represents an efficient methodology for the combinatorial synthesis of microporous materials. The figure shows a typical diffractogram of as-synthesized AFI-type molecular sieves taken with a CCD camera. PMID- 10540384 TI - "Scaffold-Hopping" by Topological Pharmacophore Search: A Contribution to Virtual Screening. AB - A chemically advanced template search (CATS) based on topological pharmacophore models has been developed as a technique for virtual screening. This technique has successfully identified novel potent Ca(2+) antagonists (such as 2) that have a similar activity to 1 (a known T-channel blocking agent) in a library of several hundred thousand compounds on the basis of a correlation vector representation. PMID- 10540385 TI - Cascade Reactions in Quantitative Solid-State Syntheses. AB - 100 % yield in the absence of the liquid phase: A one-pot synthesis of highly substituted pyrroles, which proceeds in solution with moderate yields, functions quantitatively in the solid-solid variant, and that at much lower temperatures, although at least four reaction steps are required. The reaction execution in the absence of liquid phases avoids product workup because of the 100 % yield and is thus resource-saving and environmentally friendly. PMID- 10540386 TI - Novel Fluorescent Probes for Singlet Oxygen. AB - The first fluorescent chemical traps for (1)O(2) have been developed. DPAXs react specifically with (1)O(2) to yield the corresponding endoperoxides, DPAX-EPs (see scheme; X = H, Cl, F). DPAXs scarcely fluoresce, while DPAX-EPs are strongly fluorescent. Since the fluorescence of these probes is unaffected by H(2)O(2), superoxide, and nitric oxide, they are useful for the selective detection of (1)O(2) in biological systems. PMID- 10540387 TI - Natural Product Synthesis on Polymeric Supports-Synthesis and Biological Evaluation of an Indolactam Library. AB - Potent activators of protein kinase C in fibroblasts: This property was determined for several indolactam V analogues (1) with a new cell-based assay system. This tumor-promoting indole alkaloid and analogues thereof can be synthesized efficiently on the solid phase. The key steps of the combinatorial approach are a regioselective amination of the indole ring and an enantioselective enzymatic reaction. PMID- 10540388 TI - Guest-Controlled Formation of a Hydrogen-Bonded Molecular Capsule. AB - The dimerization of the self-complementary resorcarene tetraesters is triggered by the entrapment of a tropylium cation in the pi-basic cavity. Eight intermolecular OH small middle dot small middle dot small middle dotOC hydrogen bonds together with host-guest interactions such as charge transfer and C-H small middle dot small middle dot small middle dotpi bonding are responsible for the high stability of this assembly (see picture). Hereby neither the host-guest interaction nor the interaction of two resorcarene molecules through hydrogen bonds is sufficient by itself to form the complex. PMID- 10540389 TI - A Catalytic Enantioselective Electron Transfer Reaction: Titanocene-Catalyzed Enantioselective Formation of Radicals from meso-Epoxides. AB - A rationally designed titanium(III) catalyst allows the opening of epoxides with high enantioselectivity. This reaction [Eq. (1)] constitutes the first example of an enantioselective transition metal catalyzed radical reaction that proceeds by electron transfer. PMID- 10540390 TI - Noncatalytic Organic Synthesis Using Supercritical Water: The Peculiarity Near the Critical Point. AB - Pinacol and Beckmann rearrangements can be carried out effectively by using supercritical water (scH(2)O) both as an acid catalyst as well as a medium. In the near-critical region not only are the rates significantly accelerated (see picture), but the nature of supercritical water can be adjusted to give weak acidity, which opens a new reaction pathway to a Diels-Alder adduct between dehydrated products. PMID- 10540391 TI - A 10(10) Rate Enhancement of Phosphodiester Hydrolysis by a Dinuclear Aminopeptidase-Transition-State Analogues as Substrates? AB - Streptomyces dizinc aminopeptidase (sAP) shows a specific activity of 33.7 nmol min(-1) mg(-1) toward the hydrolysis of the transition-state analogue bis-p nitrophenylphosphate with a rate constant of k(cat)/K(m)=100 M(-1) s(-1) and a first-order rate enhancement of about 10(10), which is much superior to several Zn chemical models and comparable to some phosphodiesterases. This study suggests that sAP can serve as a novel dinuclear model system to provide further insight into dinuclear hydrolysis. PMID- 10540392 TI - [n]-Polyurethanes: Synthesis and Characterization. AB - More than 50 years after Otto Bayer's detailed description of [m,n] polyurethanes, the first general synthesis of [n]-polyurethanes 1 is described. This series of aliphatic [n]-polyurethanes is synthesized by the in situ polymerization of the corresponding alpha,omega-isocyanato alcohol monomers, which in turn are made out of linear alpha,omega-amino alcohols and di-tert butyltricarbonate. Polymers of high molecular weight possessing a uniform microstructure are obtained, while their melting points show a strong odd-even effect. PMID- 10540393 TI - Isoporphycene: The Fourth Constitutional Isomer of Porphyrin with an N(4) Core Occurrence of E/Z Isomerism. AB - Liberation of the ligand from the nickel complex 1 obtained by template synthesis yielded isoporphycene (as the octaethyl derivative 2), the first constitutional isomer of porphyrin with an N(4) core for which E/Z isomerism is involved: Compound 2 is present as the E isomer, which is in rapid, presumably acid catalyzed equilibrium with a small amount (2 %) of the Z isomer. The remaining unknown constitutional isomers of porphyrin are considerably higher in energy than the already rather labile isoporphycene, so that the latter should mark the border of existence for this type of structural variant of porphyrin. PMID- 10540394 TI - The C-F small middle dot small middle dot small middle dotH-C "Anti-Hydrogen Bond" in the Gas Phase: Microwave Structure of the Difluoromethane Dimer. AB - A shortening of the C-H bond lengths and a blue shift of the C-H stretching frequencies for the C-F small middle dot small middle dot small middle dotH-C groups indicates that anti-hydrogen bonds are present the difluoromethane dimer. The most stable conformer has three such interactions (shown schematically). PMID- 10540395 TI - [Al] AB - A "naked" aluminum atom links two aluminum tetrahedra in the [Al(7){N(SiMe(3))(2)}(6)](-) ion (see picture), which results from the reaction of a metastable AlCl solution with LiN(SiMe(3))(2) and crystallizes with [Li(OEt(2))(3)](+) as cation. This unique structure among molecular metal atom clusters represents a small but characteristic section of cubic close-packed aluminum. PMID- 10540396 TI - Hyperbranched Polyether Polyols with Liquid Crystalline Properties. AB - The attachment of mesogens as end groups to hyperbranched polyglycerol (degree of polymerization 22-45; see schematic representation, the rigid mesogens are shown as rods and the flexible alkyl chains as lines) leads to liquid crystalline polymers with narrow polydispersity, whose liquid crystalline behavior is induced by the mesogenic end groups only. PMID- 10540397 TI - Ultrastable Mesoporous Aluminosilicates by Grafting Routes. AB - A combination of postsynthesis grafting and hydrothermal treatment offers an excellent route for the synthesis of ultrastable mesoporous aluminosilicates with enhanced acidity and catalytic activity. The stability observed (>150 h in boiling water; 4 h at 1000 degrees C) is, for mesoporous silicates, remarkable. Unusually the hydrothermal treatment is beneficial with respect to the use of the stable aluminosilicates as solid acid catalysts. PMID- 10540398 TI - Total Synthesis of (+/-)-Gelsemine. AB - A complex molecular reorganization (1-->2), a sequential anionic aza-Cope rearrangement and Mannich cyclization, and an unprecedented intramolecular Heck reaction of the tetrasubstituted double bond of a vinylogous carbamate are key steps in a new total synthesis of (+/-)-gelsemine (3). MOM=methoxymethyl, DBU=1,8 diazabicyclo[5.4.0]undec-7-ene. PMID- 10540399 TI - A Five-Membered Ring with Three Negative Charges and Solvent-Free Lithium Counterions. AB - Remarkably short distances to the ring plane are shown by the eta(5)-bound lithium ions in the first compound with a triply negatively charged five-membered ring, 1, which was obtained by reduction of 2 with lithium. R=CH(SiMe(3))(2), Dur=2,3,5,6-tetramethylphenyl. PMID- 10540400 TI - Ramberg-Backlund Approaches to the Synthesis of C-Linked Disaccharides. AB - Readily available S-glycoside dioxides were utilized in a Ramberg-Backlund rearrangement for the construction of C-linked disaccharides. This approach is ideally suited to analogue synthesis simply by variation of the alkylating agent, and is illustrated here by the synthesis of beta,beta-C-trehalose (see reaction scheme), a higher homologue of C-trehalose, and methyl C-gentiobioside. Bn=benzyl. PMID- 10540401 TI - Charge Transfer and Environment Effects Responsible for Characteristics of DNA Base Pairing. AB - A hitherto unresolved discrepancy between theory and experiment is unraveled. Charge transfer and the influence of the environment in the crystal are vital for understanding the nature and for reproducing the structure of hydrogen bonds in DNA base pairs. The introduction of water molecules and a sodium counterion into the theoretical model (see picture) deforms the geometry of AT and GC in such a way that excellent agreement with the experimental structures is obtained. PMID- 10540402 TI - Stereoselective Synthesis of Coordination Compounds: Self-Assembly of a Polymeric Double Helix with Controlled Chirality. AB - Its information content is infinitely smaller than that of DNA, but its structure (see picture) ressembles the double-stranded helix produced by nature. This self assembled, configurationally predetermined coordination polymer is built up from enantiopure chiral bipyridine-type ligands and silver ions. PMID- 10540403 TI - A Novel Dinuclear Diaminoplatinum(II) Glutathione Macrochelate. AB - Both oxidized and reduced glutathione (gamma-L-Glu-L-Cys-Gly) react with the anticancer complex [Pt(en)Cl(2)] to form the bicyclic complex illustrated (en=ethylenediamine). This unprecedented structure, which was determined from extensive NMR experiments, contains a ten-membered macrochelate ring. PMID- 10540404 TI - Carbohydrate Recognition through Noncovalent Interactions: A Challenge for Biomimetic and Supramolecular Chemistry. AB - Carbohydrates are not always as "sticky" as one might expect. Even in organic solvents they are difficult targets for the supramolecular chemist, due to their complex, three-dimensional structures. In their natural environment (water) they are especially elusive, presenting challenges which will occupy synthetic and theoretical chemists for some time to come. The complex of an octaamide supramolecular receptor with beta-D-glucopyranose, which binds through apolar and polar contacts, is shown. PMID- 10540406 TI - Milestones in the Biochemistry of Silicon: From Basic Research to Biotechnological Applications. AB - 6.7 Gigatonnes of silicon are processed each year by marine organisms. Since it was known that silicon is an essential element for many biological systems, significant advances in the biochemistry of this element have been achieved from the classical viewpoint of silicon being a purely inorganic element. This article describes the proteins, genes, and molecular mechanisms of silicon metabolism in diatoms and sponges. These studies may help to reveal the role of silicon for optimal development and growth in many plants and animals as well as initiate the development of new technological methods for the shape-controlled production of new patterned silicone-based materials. PMID- 10540405 TI - Chemistry in Supercritical Water. AB - Water is a special liquid-and not only under "normal" conditions. Water in the supercritical state (shaded area in the phase diagram) is distinguished by special properties which are wholly at variance with those of normal water. So far these have usually been exploited for the purification of waste water, for example for the complete oxidation of poorly degradable substances. But what is the potential of supercritical water for synthesis? Answers are provided here on the basis of thermodynamic and kinetic studies as well as with reference to measurements of corrosion. PMID- 10540407 TI - Towards Efficient and Wide-Scope Metal-Catalyzed Alkyl-Alkyl Cross-Coupling Reactions. AB - The simplest organic fragment, C(sp(3))-C(sp(3)), is unfortunately the most difficult to prepare by metal-catalyzed cross-coupling reactions (shown schematically) in the presence of functional groups. Recent advances show, however, that alkane moieties can be built within functionalized molecules by a careful choice of catalysts and conditions. PMID- 10540408 TI - Heterogeneous Catalysis with Cross-Linked Lyotropic Liquid Crystal Assemblies: Organic Analogues to Zeolites and Mesoporous Sieves. AB - Organic, nanoporous heterogeneous catalysts based on a carboxylate-containing, amphiphilic mesogen catalyze the Knoevenagel condensation (see schematic representation). These networks maintain their order in solution and can be recycled. Enhanced basicity, excellent site accessibility, and substrate size exclusion are features of these nanostructured systems. PMID- 10540409 TI - Atom-Efficient Oxidation of Alkenes with Molecular Oxygen: Synthesis of Diols. AB - Dihydroxylations of simple alkenes were carried out for the first time in excellent yields and selectivities with molecular oxygen as oxidant [(Eq. (a)]. Both oxygen atoms are used productively and are incorporated into the product in this transition metal catalyzed alkene oxidation. PMID- 10540410 TI - Valence Isomerization of a 1,3-Diphosphacyclobutane-2,4-diyl: Photochemical Ring Closure to 2,4-diphosphabicyclo AB - The bond stretch isomer 1,3-diphosphacyclobutane-2,4-diyl 1 was transformed photochemically to give the previously unknown 2,4-diphosphabicyclo[1.1.0]butane 2, which itself can be converted thermally into gauche-1,4-diphosphabutadiene 3. The crystal structures of these three energy-rich valence isomers of 1,2 diphosphete have been determined. R=SiMe(3); Mes*=2,4,6-tBu(3)C(6)H(2). PMID- 10540411 TI - 1,3-Diphosphacyclobutane-2,4-diyl-2-ylidenide: A Unique Carbene and Its Trimethylalane Complex. AB - A unique bonding situation is displayed by the lithium 1,3-diphosphacyclobutane 2,4-diyl-2-ylidenide 2 small middle dot[Li(thf)(n)](+) (Ar=2,4,6-tBu(3)C(6)H(2)) obtained by deprotonation of 1. According to ab initio calculations, the anion 2 can viewed as a cyclic bis(phosphanyl)carbene. Reaction with trimethylaluminum gives the complex 3 small middle dot[Li(thf)(4)](+), whose crystal structure is presented. PMID- 10540412 TI - Charting No-Man's Land in d(0) Transition Metal Six-Coordination: Structure Predictions for the Complexes AB - Unusual structures between octahedral and trigonal-prismatic are found by density functional calculations for the "simple" six-coordinate d(0) title complexes. While a distorted octahedron is still slightly lower in energy than a prismatic structure for [WCl(5)CH(3)], intermediate structures start to appear for [WCl(4)(CH(3))(2)] (see picture). In the case of [WCl(3)(CH(3))(3)], prismatic arrangements are already more favorable. These and many related compounds should be accessible experimentally. PMID- 10540413 TI - Trapping of an Organic Radical by an O=Cr(VI) Function. AB - Not only capable of generating organic radicals by H abstraction, chromyl chloride can also trap them again (see scheme). This is confirmed by the crystal structure of a Cr(V) oxo alkoxide, which was formed in three successive steps when bisadamantylidene oxide was allowed to react with CrO(2)Cl(2). PMID- 10540414 TI - Highly Ordered Columnar Structures from Hexa-peri-hexabenzocoronenes-Synthesis, X ray Diffraction, and Solid-State Heteronuclear Multiple-Quantum NMR Investigations. AB - Improved long-range ordering in the columnar mesophase of hexa(para-n dodecylphenyl)hexabenzocoronene 1 has been achieved by inserting phenyl rings between the extended aromatic core of hexabenzocoronene and the alkyl side chains, which are needed to form liquid crystalline phases. The long-range hexagonal order of the columns is demonstrated by X-ray scattering, while the improved packing of the aromatic cores within the columns and the molecular mobility is probed by a newly developed heteronuclear multiple-quantum MAS NMR technique. PMID- 10540415 TI - Rhodium-Promoted Linear Tetramerization and Cyclization of 3,3-Dimethylbut-l-yne. AB - With tetrahydrofuran as the solvent [Rh(thf)(2)(cod)](+) promotes the selective coupling of tBuC(2)H to form triene-yne 1; at high alkyne concentrations, however, an unusual C-H activation process intervenes, leading to the formation of complex 2, which contains two linked five-membered rings. cod=1,5 cyclooctadiene. PMID- 10540416 TI - An Atmospherically Driven Optical Switch. AB - The atmospheric regulation of the photophysical properties of 1 gives rise to a new optical switch. This optical information can be translated with the utmost spatial and molecular economy using confocal single molecule spectroscopy. R=OH. PMID- 10540417 TI - Novel Catalytic Hydrogenolysis of Trimethylsilyl Enol Ethers by the Use of an Acidic Ruthenium Dihydrogen Complex. AB - The heterolytic cleavage of H(2) is the key to the novel catalytic hydrogenolysis of trimethylsilyl enol ethers catalyzed by [RuCl(eta(2)-H(2))(dppe)(2)]OTf (dppe = 1,2-bis(diphenylphosphanyl)ethane, OTf = trifluoromethanesulfonate), which results in the formation of a ketone and Me(3)SiH (see scheme). In addition, the stoichiometric, ruthenium-assisted protonation of a prochiral lithium enolate with H(2) gave a chiral ketone with high enantioselectivity (up to 75 % ee). PMID- 10540418 TI - Direct Observation by Electrospray Ionization Mass Spectrometry of AB - The use of methanol as solvent is essential for the formation of the double bookshelf-type oxide cluster [(Cp*Rh)(2)Mo(6)O(20)(OMe)(2)](2-) from [{Cp*Rh(u Cl)Cl}(2)] and four equivalents of [Mo(2)O(7)](2-). The reaction proceeds via [Cp*RhMo(3)O(8)(OMe)(5)](-). The proposed structure for this key intermediate (shown schematically) is supported by electrospray ionization mass spectrometry and labeling experiments with CD(3)OD as solvent. Cp*=eta(5)-C(5)Me(5). PMID- 10540419 TI - The First Bismuth Phosphide Complex: AB - Thermally unstable crystals of the title compound-the first bismuth phosphide complex to be structurally characterized (see picture)-are obtained by the reaction of [Bi(NMe(2))(3)] with [tBuPHLi] (1:3) in THF/hexane. Berry pseudorotation of the pseudo-trigonal-bipyramidal [{(tBuP)(3)}(2)Bi](-) ion is prevented for steric reasons. PMID- 10540420 TI - alpha-Alkyl-alpha-Amino-beta-Lactam Peptides: Design, Synthesis, and Conformational Features. AB - Remarkable asymmetric induction is achieved in the alkylation of the lithium enolate of the beta-lactam 1. This allows the first time access to a new family of peptidomimetics 2 with predictable conformational constraints. PMID- 10540421 TI - Double Insertion of Coordinated Phosphanylalkyne Ligands into a Pt-C(6)F(5) Bond. AB - Enhanced reactivity is shown by uncoordinated C identical withC bonds in the proximity of a metal in phosphanylacetylene complexes. cis [Pt(C(6)F(5))(2)(thf)(2)] reacts with [M(C(6)F(5))(2)(PPh(2)C identical withCPh)(2)] (M=Pt, Pd) to form binuclear complexes containing the novel 2,3 bis(diphenylphosphanyl)-1,3-butadien-1-yl bridging ligand. Substitution of the solvent ligands with, for example, PPh(2)H (see picture) provides species that could be characterized by X-ray crystallography. PMID- 10540422 TI - Exploiting the Self-Assembly Strategy for the Design of Selective Cu(II) Ion Chemosensors. AB - Simply by mixing in water, a liphophilic dipeptide (L), a surfactant (S), and a fluorophore (F) self-assemble to give a sensor able to detect Cu(II) ions (see scheme). Despite the ease of construction, the sensor displays high selectivity and a low detection limit for the target ion. This new modular approach to sensing devices allows easy variations of the components, making optimization of the system very simple and fast. PMID- 10540423 TI - N,N-Diethanolaminomethyl Polystyrene: An Efficient Solid Support to Immobilize Boronic Acids. AB - The key to the efficiency of N,N-diethanolaminomethyl polystyrene (DEAM-PS), the first solid support capable of coupling to boronic acids, is the formation of a stable, resin-bound boronic ester ate adduct (see scheme). With this resin it is now possible to efficiently immobilize a wide variety of boronic acids including functionalized ones that can be derivatized by solid-phase combinatorial synthesis. PMID- 10540424 TI - Electrochemical Synthesis of Ba(2)Ag(8)S(7), a Quasi-One-Dimensional Barium Silver(I) Sulfide Containing Mixed S(2-)/S(2)(2-) Ligands. AB - A stingray pattern is adopted by Ba(2)Ag(8)S(7). Crystals of this compound were grown in a two-electrode chemical cell from a BaS(3)/ethylenediamine solution. In the pseudo-one-dimensional structure columns of (1)(infinity)[Ag(8)S(7)] (shown in the picture) are stacked head-to-tail to create voids where the barium atoms reside. The coexistence of S(2-) and S(2)(2-) indicates a reductive decomposition [Eq. (1)]. PMID- 10540425 TI - NMR Spectroscopic Structural Determination of Organozinc Reagents: Evidence for "Highly Coordinated" Zincates. AB - The structures of (13)C-labeled methylzinc reagents have been studied by NMR spectroscopy. Analysis of the spectra gives information on the number of chemically equivalent methyl groups that are attached to the metal centers. Shown is the structure of trimethylzincate with the spin couplings that form the basis of the spin system used in the calculation. PMID- 10540426 TI - An Exceedingly Stable Diiron(II,III) Complex Ion AB - A record equilibrium constant of K(c)=10(19) was found for the comproportionation of the diiron(II,III) complex 1 in acetonitrile. In water the K(c) value is strongly diminished (10(7.9)), but still exceeds that of the Creutz-Taube ion. Intervalence charge transfer bands occur at 2520 nm (in CH(3)CN) and 2250 nm (in D(2)O), and all evidence points to a strongly coupled system with delocalized valences. PMID- 10540427 TI - Conversion of Molecular Oxygen into a Hydroperoxo Species by Ring-Opening Protonation of a Cyclic eta(2)-Peroxo Intermediate: Characterization of the eta(2)-Peroxo and Hydroperoxo Complexes. AB - A transition metal-hydroperoxo species is formed by the oxygenation of a low valent rhodium precursor followed by a protonation of the resulting eta(2)-O(2) ligand; the latter process is assisted by an intramolecular hydrogen-bonding interaction (see scheme). This process is the first structural evidence for an effective method for the activation of molecular oxygen as postulated for the cytochrome P-450 system. PMID- 10540428 TI - Macroscopic, Hierarchical, Two-Dimensional Self-Assembly. AB - By tailoring capillary interactions at a fluid-fluid interface, a hierarchical two-dimensional self-assembly of hexagonal millimeter-sized poly(dimethylsiloxane) plates has been demonstrated (see picture). The strength and direction of capillary forces between plates was controlled by patterning of the surfaces of the plates to be hydophobic or hydrophilic. The thick lines indicate hydrophobic faces whose mutual attraction forms the basis of capillarity. PMID- 10540429 TI - Stereocontrolled Synthesis of (-)-5,11-Dideoxytetrodotoxin. AB - New derivatives of an intriguing marine natural product are now accessible. The first asymmetric synthesis of the simple tetrodotoxin analogue, 5,11 dideoxytetrodotoxin (3), was achieved. Hydroxylation at position C8 of the key intermediate 1 relied on the neighboring trichloroacetamide group, and stereoselective elaboration of the vinyl group gave alpha-hydroxylactone 2, which was transformed into the title compound through a new guanidylation method. PMID- 10540431 TI - High-Yielding Enantioselective Synthesis of the Macrolactam Aglycon of Sch 38516 from Two Units of (2R)-2-Ethyl-4-penten-1-ol. AB - The same precursor-namely, (2R)-2-ethyl-4-penten-1-ol-was used to obtain fragments C9-C13 and C1-C8 of 1, the aglycon of Sch 38516 (which is active against Candida sp.) and fluvirucin B(1) (which is active against influenza A virus). The key steps of the synthesis were the aldol-like reaction between the two fragments and the macrolactamization of a 13-azidotridecanoic acid derivative (see scheme). MOM=methoxymethyl, Py=2-pyridyl. PMID- 10540430 TI - Regio- and Diastereoselective Aldol Products through Three-Component Coupling Reactions of Difluoroboroxy Fischer Carbene Molybdenum Complexes. AB - The reaction of difluoroboroxy Fischer carbene complexes, vinyl ketones, and aldehydes leads to the regioselective formation of aldol-like products (see reaction scheme). The reaction also shows a high degree of diastereofacial selectivity when chiral aldehydes are used. These results show that this is a very attractive process since the regioselective formation of aldols from nearly symmetrical ketones still remains an unsolved problem. PMID- 10540432 TI - Enantioselective Total Synthesis of Avarol and Avarone. AB - Formation of the C11-C1' bond through application of Barton's radical decarboxylation and quinone addition is the cornerstone of a new convergent and concise synthesis of the marine metabolites avarol (1) and avarone (2; see scheme), for which antimitotic, antileukemic, and antiviral effects have been reported. PMID- 10540496 TI - The effect of tubing length, gas flow, and number of heaters on maximum gas temperature for aerosol circuits used for cold water near-drowning or hypothermia. AB - BACKGROUND: Clinicians who treat patients suffering from cold water near-drowning or hypothermia routinely warm inspire gases greater than body temperature in accordance with care guidelines promulgated by the various organizations. However, humidifiers are designed to prevent heating gases beyond 41 C (assuming the use of a standard six foot aerosol circuit) in order to meet International Standards Organization regulations (ISO). Clinicians must modify equipment in order to deliver care. There are several factors, which can effect the highest temperature that a particular circuit will achieve. Among the factors that are considered most important for maximum circuit are tubing length, gas flow, and the number of heaters (heat source). METHODS: The maximum temperature that a circuit could achieve was measured after varying tubing lengths (1.5 feet, 3 feet, and 6 feet), gas flow (opening or closing a venturi receiving a fixed flow rate of 10 L/min), and the size of the heat source (one or two heated humidifiers in aerosol circuit). A total of ten runs were made in each of the possible twelve combinations. RESULTS: Univariate statistics showed significant differences for Venturi open/close (p = .0001) and the number of heaters (p = .0001) but not the tubing length (p = 0.19). However, the multivariate analysis revealed significance for tubing length, number of heaters, and venturi open/closed (p = .01). CONCLUSION: All factors (tubing length, number of heaters, and tubing length) were important determinants of maximum gas temperature. The effect of tubing length can be overwhelmed by higher gas flows. PMID- 10540497 TI - Expired gas analysis of field simulated CPR. AB - The concentration of expired oxygen and carbon dioxide that a rescuer may give during cardiopulmonary resuscitation was originally determined without adjusting for conditions that can exist in real life. This study was devised to investigate how simulated field conditions might effect the concentrations of these gases that a rescuer would give a victim. The expired gases were measured from volunteers who performed one person cardiopulmonary resuscitation on manikins for two minutes. They then stopped the resuscitation, walked or ran 200 meters, and performed another two minutes of cardiopulmonary resuscitation while listening to loud ambulance sirens. For the thirty-eight volunteers that were tested, it was found that the percentage of carbon dioxide increased by an average of 11% and the oxygen concentration only changed by a negligible amount. This amounted to a statistically significant increase in the carbon dioxide concentration (p << 0.01), without a significant change in the oxygen concentration (p > 0.05). In conclusion, exertion before performing CPR will cause an increase in the percent of CO2 expired, while not affecting the concentration of O2. PMID- 10540498 TI - The challenge of ongoing Haemophilus influenzae type B carriage and transmission in Alaska. AB - Cases of invasive Haemophilus influenzae type b disease in Alaskan children quickly dropped 10-fold after widespread vaccination with a conjugate vaccine (PRP-OMP) began in 1991. However, reemergence of invasive disease in 1996-97 soon followed a change to a combination diphtheria-tetanus toxoid-pertussis/H. influenzae type b vaccine which incorporates a different conjugate vaccine (HbOC). Previously unrecognized persistence of H. influenzae type b carriage in rural Alaska, coupled with characteristics of the immune response to HbOC, are the likely explanations for disease reemergence. The current vaccine recommendation--PRP-OMP for the first dose, followed by HbOC to complete the vaccination series--appears to protect Alaskan infants even in the face of continuing carriage and transmission. Successful control of invasive H. influenzae type b disease in Alaskan children will require not only appropriate immunization, but also continuing surveillance for both disease and carriage, identification of factors associated with carriage, and investigation into the feasibility of using vaccination plus antimicrobial drugs to eliminate this pathogen. PMID- 10540499 TI - Decrease birth defects by increasing the number of women who take folic acid before they are pregnant. PMID- 10540500 TI - [Weight and fertility: nutritional regulation of reproductive function]. PMID- 10540501 TI - [Theory and practice in daily prescriptions in an obstetric office. Antibiotics]. PMID- 10540502 TI - [Markers for trisomy 21]. PMID- 10540503 TI - [Clomiphene citrate: for or against?]. PMID- 10540504 TI - [Ovulation]. AB - This review summarizes our knowledge on ovulation process. After gonadotropin surge or LH injection, 13 factors are involved in the follicular rupture. All of them plays a role since the inhibition of their synthesis or activity prevents the rupture of preovulatory follicles or reduces the number of ovulations. Most of them are involved in all inflammatory reactions. At the apex of the follicle, blood vessels constriction, free TNF alpha and probably factors from the ovarian epithelium cells are involved in the full dissociation of follicle layers and cell death, allowing the localized rupture of the follicle wall. Gonadotrophins and angiotensine II are the only factors able to induce both follicle rupture and meiotic resumption. PMID- 10540505 TI - [Intra-uterine insemination]. AB - Artificial insemination has been proposed for a number of years in the treatment of unexplained or male factors related to infertility with very low results. In recent years, the association of intra-uterine insemination with gonadotropin ovulation induction has demonstrated its effectiveness and it is now the first treatment to propose in these cases before in vitro fertilization. PMID- 10540506 TI - [Anti-estrogens, selective estrogen receptor modulators (SERM), tibolone: modes of action]. AB - The regulation of estrogen-induced cellular effects is a multistep molecular process. The diversity of estrogen and anti-estrogen effects on cellular functions is also modulate by tissue and gene specificity. This diversity may be explained by different levels of molecular regulation, including the presence of two distinct estrogen receptor isoforms (ER alpha and ER beta), their binding to activator or corepressor transcriptional proteins, and their affinity to different DNA binding domains of target genes (estrogen responsive element or API). These mechanisms may account for the specific responses to estrogens or anti-estrogens according to tissue, cell or gene level. The anti-estrogen tamoxifen, in vitro, inhibits the estrogen-induced proliferation of breast cancer cells and, in vivo, enhances long-term prognosis of patients having ER positive breast cancer when it is used as an adjuvant treatment. The partial agonist effect of anti-estrogens such as raloxifene, is observed only on bones and vessels but not in endometrium. Tibolone is another class of ER modulators which acts as a prodrug. It is metabolized in compounds activating nuclear receptors (androgen and progesterone receptors) which modify the ER level and estrogen metabolism. The improvement of current knowledge on the cellular mechanisms of estrogen and anti-estrogens action should allow the elaboration new therapeutic approaches on specific functions involved in estrogen-dependent pathologies. PMID- 10540507 TI - [From the prescription of contraceptives to their application. A study of the on going use of contraceptive methods in immigrant circles during the post-partum period]. AB - PURPOSE: The research presented here aimed at establishing a possible link between the level of integration of an immigrant population and the rate of continuity following a prescription for contraception given to women on leaving the maternity clinic. MATERIALS AND METHODS: From June 1995 to November 1996, a study was conducted in Valence among 71 women who came from Turkey, sub-Saharan Africa and the Maghreb. Out of 87 women interviewed at the maternity, 71 were seen a second time. As well as taking into consideration socio-demographical factors, the study compared the degree of integration with the use of the prescribed contraceptive method six months after leaving the maternity clinic. RESULTS: The study shows that there is no significant link between the level of integration and the continued use of a contraceptive method six months after birth. The only salient fact is that women who had lived less than 10 years in France systematically used a contraceptive method (P = 0.004, relative risk: 1.44, 95% confidence interval: 1.12-1.84). DISCUSSION: Our study would tend towards the following observation: the more recently the immigrant arrived, the more receptive they were towards proposals made by the medical personnel. We consider that rather than it being the case of either total confidence in, or distrust of, medical contraception, the cultural factor remains important and must be taken into account in any approach among this kind of population. It is more a question of community health education than of medical prescription. PMID- 10540508 TI - [Profound endometriosis and infertility. Fertility results after laparoscopic treatment of profound endometriosis infiltrating the uterosacral ligaments]. PMID- 10540509 TI - [Teaching biopsychosocial approach in the carrying out of clinical interviews before teaching to recognize emotional disturbances]. PMID- 10540510 TI - Training general practitioners in mental health skills. PMID- 10540511 TI - Comprehensive, research-based interviewing guidelines in general practice settings. PMID- 10540512 TI - Detection and management of mental distress and psychiatric disorders in primary care settings. AB - OBJECTIVE: Epidemiological and clinical studies indicate that 10-50% of primary care patients suffering from clinically relevant psychiatric distress are not diagnosed by their physician and only a minority of them receive an appropriate treatment. The improvement of physicians' ability to detect mental distress and psychiatric disorder, in their routine clinical activity, represents a crucial point to reduce the social impact of mental illnesses, prevent their worsening and chronicity and, eventually, relieve mental health services of an excessive burden of care and costs. The aim of this article is to examine a number of factors which intervene in the process of detection of mental distress by the physician. Then, we will examine factors related to the management of psychiatric disorders most commonly co-occurring with physical illness in general health care sector. METHOD: The method used for this review was essentially a recension of the literature concerning detection and treatment of psychiatric disorders in primary care settings, having in view to see the factors connected with these processes. RESULTS: Among factors intervening in the process of identification of mental distress in primary care settings, both the characteristics of the physician and the characteristics of the patient should be taken into account. Primary care physicians and psychiatrists are being asked to work together more frequently in this era of community care. The principal aim of such invoked collaboration is the amelioration of quality of care and reduction of costs for mentally ill patients. An important issue within this collaboration is the referral by primary care physicians to specialist services. PMID- 10540513 TI - [The Italian law on psychiatric hospitals. Toward the model of deinstitutionalization]. AB - An evaluation is currently underway concerning the changes which have taken place in Italy in the twenty years following the reform law. The qualitative and quantitative changes are being analyzed based on a possible shared definition of the processes of deinstitutionalization. This theme is generally the object of misunderstandings and cliches. The need for change in clinical and institutional psychiatry is the indispensable premise for the development of community psychiatry and the growth of a culture of public psychiatry in general. In this framework, an attempt is being made to define the meaning of change through the growth of the active participation of person affected with mental disorders and their families in treatment, the participation of ordinary citizens, the spread of services in the community and the quantitative increase of the number of personnel involved in public community services. Emblematic of this change is the increase in the number of psychiatrists working in the public sector, from 700 to 7,000, over this twenty year period. The changes which must take place in psychiatric practice must also be emphasized: the heirarchies, the relationships, the search for non-health resources and enhancing the value of operators as subjects outside of their institutional role. The various forms of resistance which have retarded, and continue to retard, the process of change are also considered: the persistence of clinical cultural models, administrative inertia, the defense of acquired privileges by medical and nursing lobbies, the interests of the private, commercial and religious sectors and political manipulation. In any case, the beginning of a process of change which contains all the potential of a real project for prevention is judged positively. PMID- 10540514 TI - [The closure of The St. Lazarus Psychiatric Hospital in Reggio Emilia]. AB - This is not a scientific paper. It is the report of the actions carried out during 1996 and 1997 to close the Psychiatric Hospital S. Lazzaro in Reggio Emilia. PMID- 10540515 TI - [Problems of patients with schizophrenic disorders and of their families]. AB - OBJECTIVE: To evaluate psychopathological symptoms, disabilities and family burden in schizophrenic patients and to analyse predictors of family burden and relatives' satisfaction. DESIGN: Descriptive study of 203 patients with an ICD 10 -F2 diagnosis (schizophrenia and related disorders) in contact with the Desio Department of Mental Health on 31st December 1994. SETTING: The Desio Department of Mental Health. MAIN OUTCOME MEASURES: The patients have been evaluated in three areas: disability (by ADC-DAS), psychiatric symptoms (by 24 items BPRS) and family burden (by Family Problems questionnaire). The outpatient, hospital and residential care contacts of the patients have been collected for six months by our service information system. For each area (DAS, BPRS and FP) a principal component analysis and a rotation of the significant components have been performed. Eleven factors, derived from three scales, have been retained as explanatory variables. Finally, a multiple regression analysis has been performed to assess the influence of explanatory variables on the set of response variables regarding family burden and relatives' satisfaction. RESULTS: One third of patients suffer of moderate-severe positive symptoms, while negative symptoms are less frequent. Manic symptoms are rare while depressive ones more frequent. Disability, related to work and sexual problems, is frequent; social withdrawn, underactivity, lack of participation in household duties and lack of self care are less frequent. Family burden is severe in one third of relatives, mainly in social relationships. Disability is the main predictor of family burden; manic and positive symptoms, time spent by the carer with the patient and carer's social support are less important. Satisfaction with services is predicted by family burden. CONCLUSIONS: To be more responsive to the needs of patients and relatives we should increase activities in rehabilitation and family support areas. Further analysis of severity of psychosocial and psychiatric problems, based on an epidemiological based sample, could give interesting results on the case-mix of different services. PMID- 10540516 TI - [Camberwell assessment of need (CAN). Preliminary introduction]. PMID- 10540517 TI - The utilization of nurse-midwives as providers of obstetric triage services. Results of a national survey. AB - Obstetric triage services are rapidly advancing, and the concept is becoming a popular practice pattern. As more pregnant women are evaluated in ambulatory settings, especially in high volume obstetric tertiary centers, it is now realized more and more that labor and other complaints cannot be addressed solely in labor and delivery units, nor are most of these complaints solely labor related. This article presents the results of a national survey designed to discover what contributions nurse-midwives are making to obstetric triage services. In addition, the results provide initial benchmark data on obstetric triage components against which other midwifery services can address practice issues. PMID- 10540518 TI - Midwifery triage and management of trauma and second/third trimester bleeding. AB - Trauma affects approximately 8% of all pregnancies, and bleeding affects nearly 5% of gestations. These two conditions are potentially life-threatening and require immediate management by the midwife. Trauma in pregnancy is commonly caused by motor vehicle accidents, falls, and assault. Although abruption resulting from trauma is a rare occurrence, injury caused by domestic violence, is associated with the greatest risk of obstetric complications. Bleeding in pregnancy has a number of etiologies. Midwives are well-prepared to safely and competently make a differential diagnosis of bleeding in the second and third trimesters. A sequence for midwifery triage of clients who present to the emergency room/triage area for trauma and bleeding is presented. Considerations for stabilization, history, physical examinations, diagnostic testing, initial management, and follow-up are described. Practical considerations for midwifery services incorporating provisions for triage into their caseloads are also provided. PMID- 10540519 TI - Preterm labor in the triage setting. AB - Preterm birth is one of the most devastating problems facing obstetrics today. Despite all of the available sophisticated research and therapeutic technology, the preterm birth rate has remained the same for the last 40 years. One birth in 10 occurs prematurely. Preterm labor manifests itself in a variety of ways. It is essential to promptly differentiate true preterm labor from preterm contractions or other conditions that present with similar symptoms. True preterm labor requires prompt clinical intervention in the obstetric triage setting. PMID- 10540520 TI - Triage issues in an out-of-hospital birth center. AB - Effective triage in an out-of-hospital birth center helps low-risk women avoid high-risk care. Background issues include the contributions of evidence-based practice, informed consent, patient education, problem-focused documentation, after-hours access to client data, and the value of intuition. Telephone triage, immediate referral, birth center management, and follow-up with counseling are outlined for common out-of-hospital triage problems: first trimester bleeding, nausea and vomiting, second and third trimester bleeding, urinary tract symptoms, decreased fetal movement, contractions < 37 weeks, rupture of membranes, contractions > or = 37 weeks, and "emergency" delivery. PMID- 10540521 TI - Telephone triage. A challenge for practicing midwives. AB - Telephone triage is the process by which a health care provider communicates with a client via the telephone and, thereby, assesses the presenting concerns, develops a working diagnosis, and determines a suitable plan of management. Determination of the seriousness of the situation will dictate whether a client can be cared for at a distance or whether a more comprehensive in-person evaluation is in order. The process of telephone triage is fraught with potential problems, including difficulty in establishing a reliable database, environmental distractions, cost concerns, liability issues, and, frequently, inadequate documentation. This article will describe an approach to these concerns by discussing the use of appropriate communication techniques, the development of a working diagnosis, the establishment of a plan of intervention, and the appropriate documentation of care. Such steps will go far toward diminishing the growing legal threats that arise to midwives who utilize this technology to render care to their patients. PMID- 10540522 TI - Ultrasound in obstetric triage. AB - The use of limited ultrasound as a tool in obstetric triage has become increasingly popular. While most midwives only perform ultrasound in the third trimester, some institutions do include earlier sonographic testing as part of the midwifery management plan. This article addresses the professional issues related to limited ultrasound in obstetric triage protocol, the education and training needed to acquire the skill, the process for integrating the skill into clinical practice, and a discussion of obstetric triage procedures. PMID- 10540523 TI - Obstetric triage in 10 U.S. midwifery practices. PMID- 10540524 TI - Efficacy of selective venous sampling to localize a small ovarian androgen producing tumor. AB - Two cases of androgen-producing tumors, including a Sertoli-Leydig cell tumor in a woman of reproductive age and a Leydig cell tumor in a postmenopausal woman, are reported herein. In both cases, only selective venous sampling was able to detect the presence of the androgen-producing ovarian tumors. PMID- 10540525 TI - Morphological aspects of ectopia cordis: four case reports and a review of the literature. AB - Ectopia cordis is a rare congenital anomaly. We present 4 cases of ectopia cordis, 1 of which is the first report of an affected fetus in a triplet pregnancy. The morphological relationship between the types of ectopia cordis and their outcomes were investigated in all 4 cases. In addition, the literature on ectopia cordis in Japan was reviewed and discussed. PMID- 10540526 TI - A clinicopathological study of postoperative pulmonary metastasis of uterine cervical carcinomas. AB - OBJECTIVE: To investigate the clinicopathological backgrounds and prognostic factors of uterine cervical carcinomas metastatic to the lung. METHODS: A total of 519 patients with invasive cervical carcinoma (Stage pTIb-IIb) treated by abdominal radical hysterectomy at the Saitama Cancer Center from January 1, 1976 to December 31, 1989 were analyzed clinicopathologically. RESULTS: The frequencies of pulmonary metastasis were 6.4% (24/377) and 11.3% (16/142) in patients with negative and positive pelvic lymph nodes, respectively. Among 24 negative lymph node patients, 15 had pulmonary metastasis only. The overall 5 year survival rate of these 15 patients was 36% after relapse. Of the 15, the prognosis of 12 patients with 1-3 pulmonary metastases only was better, that is, 46% after surgical resection (mean size of resectable tumor = 2.8 cm) and/or chemotherapy. But the other patients died within 3.3 years after relapse. CONCLUSIONS: The occurrence of pulmonary metastasis only, its number (1-3) and size (mean size = 2.8 cm), and no lymph node metastasis are important prognostic factors. For these patients, active surgical resection of the pulmonary lesion(s) and further chemotherapy are recommended in order to improve their prognosis. PMID- 10540527 TI - Effect of vaginal delivery on the Q-Tc interval in a patient with the long Q-T (Romano-Ward) syndrome. AB - Twelve ECG leads were monitored continuously during peripartum in a 23-year-old Japanese woman diagnosed as having the long Q-T (Romano-Ward) syndrome. Corrected Q-T (Q-Tc) intervals determined by 2 investigators blinded from the clinical informations disclosed that the Q-Tc interval increased during labor, suggesting that physical and/or emotional stress during labor might cause prolonged Q-Tc intervals in women with the long QT syndrome. PMID- 10540528 TI - Comparison of the anterior colporrhaphy procedure and the Marshall-Marchetti Krantz operation in the treatment of stress urinary incontinence among women. AB - OBJECTIVE: To compare retrospectively the efficacy of the anterior colporrhaphy procedure (AC) and the Marshall-Marchetti-Krantz operation (MMK) in the treatment of stress urinary incontinence (SUI). METHODS: A retrospective analysis through a review of the medical records of Japanese women with stress urinary incontinence who were surgically treated at Kyushu University Hospital from 1980 through 1996. A questionnaire regarding the current status of urinary incontinence was sent to all patients. RESULTS: A total of 103 patients could be evaluated, 77 of whom had undergone an AC, and 26 of whom had undergone an MMK. Postoperative complications were more common in the AC group (p < 0.05). There were no significant differences between the 2 groups in terms of the duration of hospital stay or postoperative catheterization. The long-term subjective cure rates of the ACs and the MMKs were 55% and 58%, respectively. CONCLUSIONS: The AC and the MMK were equally effective in treating stress urinary incontinence, and both showed decreased long-term subjective cure rates. The recurrence rate did not differ between the AC and the MMK. The AC had more postoperative complications and shorter recurrence intervals. PMID- 10540529 TI - A case report: pregnancy complicated by blue rubber-bleb nevus syndrome. AB - Blue rubber-bleb nevus (BRBN) syndrome, first reported in 1958 by Bean, manifests with multiple hemangiomas located in the skin and gastarointestinal tract. Characteristic laboratory data include chronic anemia with iron deficiency and consumption coagulopathy. We describe herein a pregnancy complicated by BRBN syndrome resulting in the delivery of a male infant by cesarean section. PMID- 10540530 TI - An analysis of breastfeeding patterns and menses returning in Chengdu, China. AB - OBJECTIVE: To study the relationship between infant feeding practices and the duration of lactational amenorrhea, WHO conducted a multi-centered prospective study in 7 countries from 1989. This paper reports the preliminary results based on the data collected in Chengdu center in China. METHODS: A detailed follow-up survey was conducted among 541 pairs of mothers and infants from delivery to the returning of menses of mothers at the interval of every 2 weeks. RESULTS: The results showed that the mean number of breastfeeding episodes and mean duration of breastfeeds had little correlation with the time since delivery. Within 6 months since delivery, the percentage of infants' feeds consisting of breastmilk was over 90%, and this percentage dropped to 70% at 1 year's age of infants. The time to the start of regular supplementation was 153 days postpartum. The proportion of women in exclusive breastfeeding at 2 weeks postpartum was 73.4%, and these figures at 3 and 6 months postpartum were about 60% and 40%, respectively. Until 18 months postpartum, nearly 80% mothers were partial breastfeeding their infants. The cumulative probabilities of menses returning were 0.0150, 0.0395, 0.2345 and 0.6820 at 61, 89, 187 and 369 days postpartum respectively. The median duration of lactational amenorrhea was 282 days. CONCLUSION: These results indicated that the duration of lactational amenorrhea would be prolonged if the time of exclusive/predominant breastfeed was extended and the supplementary food was introduced later. The paper concluded that the first menses returning was the signal for initiating contraceptive methods for women. PMID- 10540531 TI - Remission of idiopathic hypoparathyroidism during lactation: a case report. AB - A 35-year-old woman with idiopathic hypoparathyroidism achieved a spontaneous remission during lactation even though 1-alpha-hydroxyvitamin D3 therapy was discontinued after delivery. Urinary excretion of cyclic adenosine 3',5' monophosphate and phosphate was significantly increased during lactation, probably in response to the increased levels of circulating parathyroid hormone related protein derived from the breast tissue. PMID- 10540532 TI - A case of huge ovarian cyst of 21-year-old young woman. AB - Huge ovarian tumors are rarely seen in modern surgical practice. As health care education and access to hospitals have improved over the past 30 years, the number of these reports have become almost negligible. However, these huge ovarian tumors still present many challenge, even life-threatening risks due to severe cardiovascular, pulmonary, and circulatory problems, including technical difficulties of surgery, massive hemorrhage, and postoperative complications. A knowledge of the deranged physiology and its management may avert these complications. We present the case of a 21-year-old woman with huge ovarian tumor. The total weight of the tumor was 136 pounds (62 kg). She was treated surgically with good results. PMID- 10540533 TI - High-risk types of human papillomavirus associated with the progression of cervical dysplasia to carcinoma. AB - OBJECTIVES: In the past several years, much evidence has accumulated that strongly implicates human papillomavirus (HPV) as an etiological agent of cervical cancer. However, the natural history of HPV infection is not yet completely understood, and at present there is no good marker to predict progression to invasive cancer in individual patients with dysplasia. METHODS: Tissue specimens from 45 patients with cervical dysplasia were classified as showing progression to carcinoma (progression group; 26 women), spontaneous regression (regression group; 16 women), and persistence (persistence group; 3 women). The presence of HPV 16, 18, 33, and 52 DNA was examined in 90 formalin fixed, paraffin-embedded surgical and biopsy specimens of dysplasia using the PCR method, and the relationship between the presence of HPV and cervical carcinogenesis was analyzed. The mean follow-up period was 25.8 months in the progressed group and 34.0 months in both the persistent and regressed groups. The mean ages were 46.4 in the progressed group and 46.8 in the persistent and regressed groups. RESULTS: HPV 16, 18, 33, and 52 DNA were detected in 19 of the 26 patients (73.1%) in the progressed group. All patients with HPV 16, 18, or 52 DNA showed progression to carcinoma. The HPV 33 DNA was detected in 5 of the 16 patients (31.3%) in the regressed group and in 2 of the 3 patients (66.7%) in the persistent group, while HPV 16, 18, and 52 DNA were not detected in the 19 patients that comprised the regressed and persistent groups. CONCLUSION: This retrospective study suggests that cervical dysplasia in patients with high-risk types of HPV possessed the potential to progress to carcinoma in situ and eventually to invasive carcinoma as well. PMID- 10540534 TI - The effect of medroxyprogesterone acetate and heparin in the prevention of postsurgical adhesion formation in the rat uterine model. AB - OBJECTIVE: To evaluate the effects of depot medroxyprogesterone acetate and heparin in preventing postsurgical adhesion formation in the rat model. METHODS: A hundred and five female Wistar rats were divided into 7 groups. Groups 1 and 2 were injected with intramuscular 15 mg medroxyprogesterone acetate 3 weeks before surgery and at the end of laparotomy. Groups 3 and 4 were given 15 mg medroxyprogesterone acetate by intramuscular injection, 3 weeks before surgery. An equal volume of intramuscular sterile saline was injected to control groups, 3 weeks before and at the end of surgery. Before abdominal closure, 2 ml of Ringer's lactate was instilled into the peritoneal cavity of all rats, except group 7. Groups 1, 4, and 5 were given 2 ml of intraperitoneal Ringer's lactate containing 500 U heparin/ml. A standardized surgical injury was performed in all rats. Two weeks after surgery, the adhesions were scored on a scale of 0 to 3 according to their thickness-tenacity and vascularity. Kruskall-Wallis and Mann Whitney U statistical test were used. RESULTS: The preoperative and postoperative administration of medroxyprogesterone acetate resulted in the least number of and the least severe adhesions, when compared with single dose medroxyprogesterone acetate treated rats and controls (p < 0.05). However, the combination of medroxyprogesterone acetate and intraperitoneal heparin did not enhance the adhesion reducing capacity of medroxyprogesterone acetate. CONCLUSIONS: Concurrent preoperative and postoperative administration of medroxyprogesterone acetate results in the most significant reduction of postsurgical adhesions. The combination treatment of medroxyprogesterone acetate and heparin does not show any additional effect in the reduction of adhesion formation, when compared with medroxyprogesterone acetate treatment alone. PMID- 10540535 TI - Burn care, one of the most neglected health care issues in Pakistan. PMID- 10540536 TI - Immunohistochemical evaluation of small round cell tumors of childhood. AB - OBJECTIVE: This study was done to evaluate the pediatric undifferentiated small round cell tumors with immunohistochemical staining. SETTING: The present study included consecutive cases of small round cell tumors which were diagnosed in children (< 15 years) in the section of Histopathology at the Aga Khan University Hospital, Karachi during the period of two years. METHODS: The group of undifferentiated small round cell tumors were evaluated immunohistochemically by using a panel of antibodies on sections from routinely processed, formalin fixed, paraffin embedded tissue blocks. RESULTS: The category of undifferentiated small round cell tumors included rhabdomyosarcoma (23.2%), primitive neuroectodermal tumor (17.9%), non-Hodgkin's lymphoma (16.1%), neuroblastoma (14.2%), Ewing's sarcoma (10.7%) in order of frequency. Osteosarcoma (Small cell variant), retinoblastoma and medulloblastoma comprised 1.8% each. In seven cases (12.5%), the immunohistochemical analysis was inconclusive. CONCLUSION: Immunohistochemistry is a very valuable diagnostic tool which helps in distinguishing the undifferentiated tumors especially small round cell tumors. The immunohistochemical staining needs to be performed routinely for undifferentiated tumors in diagnostic histopathology. PMID- 10540537 TI - The effects of glibenclamide on serum lipids and lipoproteins in type II non insulin dependent diabetes mellitus. AB - OBJECTIVE: To examine the effects of glibenclamide treatment on plasma lipids and lipoprotein levels. SETTINGS: Out patients of Type II diabetics from department of Baqai Diabetes and Endocrine Centre and two other diabetic clinics of Karachi. METHODS: The effects of glibenclamide on blood glucose and various aspects of lipoproteins has been studied in 26 (14 male, 12 female) Type II Diapetes patients before and after 12 weeks of glibenclamide therapy. Treatment was initiated with 5 mg oral glibenclamide with diet control. The initial dosage of glibenclamide was 5 mg/day taken half an hour before meal; this was increased to 5 mg per week and was adjusted according to the patient's tolerance to the drug and their glycemic control. RESULTS: The results demonstrated that fasting blood glucose declined from 221.53 + 7.84 to 165.02 + 5.12 mg/dl, (P < 0.001). There was a statistically significant increase in the plasma high-density lipoprotein cholesterol from 33.60 + 1.00 to 37.07 + 1.05 mg/dl, (P << 0.05). Total cholesterol, triglycerides, low-density lipoprotein cholesterol and very-low density lipoprotein cholesterol did not change significantly. CONCLUSION: Improved glycaemic control in patients treated with glibenclamide with Type II Diabetes was achieved which lead to changes in lipoprotein metabolism. There was no evidence of changes in lipoproteins in directions associated with an increased risk for atherosclerosis. PMID- 10540538 TI - Perceived gynecological morbidity among young ever-married women living in squatter settlements of Karachi, Pakistan. AB - BACKGROUND: Community-based information on obstetric and gynecological morbidity in developing countries is meager and nearly non-existent in Pakistan. OBJECTIVES: To estimate the prevalence of specific gynecological morbidities and investigate the predictors of pelvic inflammatory disease. METHODS: Users [404] and non-users [313] of modern contraceptives were identified from eight squatter settlements of Karachi, Pakistan and detailed information on basic demographics, contraceptive use, female mobility, decision-making and gynecological morbidities were elicited. RESULTS: The perceived prevalence of menstrual disorders were 45.3%, uterine prolapse 19.1%, pelvic inflammatory disease 12.8% and urinary tract infection 5.4%. The magnitude of gynecological morbidity was high with about 55% of women reporting at least one gynecological morbidity though fewer [20%] reported at least two gynecological morbidities. Significant predictors of pelvic inflammatory disease were intrauterine contraceptive device users (OR = 3.1; 95% CI 1.7-5.6), age < or = 20 years (OR = 2.3; 95% CI 1.1-4.8) and urban life style (OR = 2.1; 95% CI 1.0-4.6). CONCLUSION: There is an immense burden of reproductive ill-health and a significant association between ever users of intrauterine contraceptive device and pelvic inflammatory disease. We therefore suggest improvement in the quality of reproductive health services generally, but specifically for family planning services. PMID- 10540539 TI - Carotid Doppler ultrasonography in young stroke patients. AB - BACKGROUND: The present study focuses on the role of carotid doppler ultrasonography (CDUS) in the diagnosis and management of carotid stenosis in young stroke patients. METHODS: The findings of carotid doppler in 45 ischemic stroke patients between 15-45 years of age were reviewed retrospectively. The variables of interest for this study included risk factors for atherosclerotic disease, primary abnormality detected on carotid doppler ultrasonography (ulceration vs. stenosis), degree of stenosis and the type of plaque (soft vs. calcified). RESULTS: The prevalence of hypertension and diabetes was 50% and 35% respectively. The rate of carotid stenosis in the study population was found to be 31%. The degree of stenosis was mild in 35% and moderate in 21%. High-grade stenosis was found in 21% of patients. The plaque was soft in the majority of cases (43%). CONCLUSION: The proportion of carotid stenosis in young stroke patients was relatively high compared with previous studies. This may be due to an increase in the risk factors for atherosclerotic disease in developing countries. PMID- 10540541 TI - Neuralgic amyotrophy (Parsonage-Turner syndrome): an often misdiagnosed diagnosis. PMID- 10540540 TI - Autoimmune hemolytic anemia in visceral leishmaniasis. PMID- 10540542 TI - Evaluation and management of malaria in general practice. PMID- 10540543 TI - Aspiration pneumonia in pediatric age group: etiology, predisposing factors and clinical outcome. AB - INTRODUCTION: Aspiration pneumonia in children is an important disease in terms of the morbidity and mortality associated with it. The objective of this study is to characterize the cases of aspiration pneumonia on the basis of the predisposing factors, types of aspiration syndromes, materials aspirated and their clinical outcome. METHODS: A total of 107 patients diagnosed as having aspiration pneumonia, were included in this study. Cases were between 0-15 years of age, admitted to the Aga Khan University Hospital (AKUH) over five years. RESULTS: The most common form of aspiration syndrome seen was chemical pneumonitis (52.1%). The three most common factors predisposing to pulmonary aspiration were accidental ingestion (37.4%), altered consciousness (34.6%) and neurologic disorders (29%). Children who aspirated oropharyngeal flora were at higher odds to require mechanical ventilation than those aspirating inert fluids and particulate matter (OR = 6.4, 95% CI: 1.5-29.2, p = 0.003). Milk (31.8%), kerosene (21.5%) and oral secretions (19.6%) were the most common materials aspirated. Betel nuts were the most commonly aspirated foreign body. Patients aspirating oral secretions and milk were seen to have a relatively worse clinical outcome than those aspirating kerosene oil. CONCLUSION: Aspiration pneumonia is a relatively uncommon clinical entity at AKUH in children. However, it does cause significant morbidity and mortality. PMID- 10540544 TI - [Surgical sepsis-definition of the notion. Problems of terminology]. AB - The problems of terminology are discussed, as well as classification and definition of sepsis. Comparative analysis of traditional for our country and up to-date foreign classifications is carried out. The criteria for diagnosis of sepsis are discussed. On the basis of 20-year experience in examination and treatment of patients with sepsis, the authors set forth their own stand, suggesting two varieties of sepsis for consideration: common complication of surgical infection, a rare disease. PMID- 10540545 TI - [On the problems of sepsis classification]. AB - The article is devoted to classification of sepsis. Two classifications are compared: the Russian one, developed in A.V. Vishnevsky Institute of Surgery (phase I--the initial one or toxemia, phase 2--septicemia; 3--septicopyemia), and the American one, accepted in 1991 in Chicago which singles out three forms of sepsis (sepsis, severe sepsis or sepsis-syndrome, and septic shock). Both classifications are thoroughly analysed and it is stated that there is no principal disagreement in the definition of phases of the course and description of clinical stages of sepsis. The necessity for universal classification of sepsis is emphasized. PMID- 10540546 TI - [Debatable questions of surgical sepsis treatment and diagnosis]. AB - On the basis of critical analysis of the results of combined treatment of 73 patients, the authors suggested that sepsis could be diagnosed only in the presence of systemic inflammatory response, multiorganic insufficiency and obligatory isolation of hemoculture. The microorganisms' specimen from the infected area and the blood are not identical both in primary and repeated inoculation. Impossibility to verify the pathogenic microorganism in the blood prevents from carrying out adequate and purposeful treatment. The absence of correlation between the severity of clinical course of sepsis, microflora species and laboratory immunography data, confirms the opinion that the organism responds to the inflammatory process by complex general physiologic reaction of defence, which includes, besides immunologic system, fermentative, endocrine, vegetative and other systems of homeostasis. PMID- 10540547 TI - [Intensive care in surgical sepsis]. AB - The experience in treatment of 740 patients with surgical sepsis is presented. The problems of the diagnosis, pathogenesis of sepsis, clinical and laboratory features of its course, and disturbances of homeostasis are discussed. Score systems for evaluation of the severity of patients' conditions are analysed. The problems of intensive care of surgical sepsis are considered, absolute and relative components are singled out. The principles for application of antibacterial and immune therapy, restoration of protein and energy losses, infusional and transfusional therapy are discussed. The algorithm for intensive care measures is suggested. PMID- 10540548 TI - [Some aspects of intensive care for severe forms of anaerobic non-sporeforming infections of soft tissues]. AB - The results of treatment (in Hospital N.N. Burdenko) of 167 patients with anaerobic nonsporeforming ("nonclostridial") infection of soft tissues (ANIST) of various location are presented. Principal errors of intensive care in this category of patients, substantial difference in the course of infectious process in patients with ANIST depending the diseased area are shown. Programs of treatment for the patients with limited (up to 1600 cm2) and extended (over 1600 cm2) forms of ANIST are proposed. The effectiveness of such components of intensive care as ozonotherapy, correction of metabolic disturbances, HBO has been studied. Practical application of these programs allowed active influence on intoxication syndrome in ANIST which resulted in a decrease of lethality up to 10.2%. PMID- 10540549 TI - [Biliary sepsis: some peculiarities of pathogenesis]. AB - Results of clinical study of 87 biliary sepsis patients and experimental study on 54 rats with obstructive jaundice and cholangitis are presented. Own and literary data are compared. Specific immune and portal haemodynamic changes, provoced by obstructive jaundice are main pathogenic factors defining specific course of biliary sepsis. These changes are: 1) gut bacterial and endotoxin translocation, portal endotoxaemia; 2) reduction of RES and Kupfer cell function and endotoxin break into the systemic circulation; 3) liver parenchyma ischemia and milliary abscess formation; 4) portal blood flow shunting into the general circulation additionally increasing systemic endotoxaemia. These factors determine rapid, even fulminate development of milliary abscesses of the liver and multiorganic failure. The authors suggest that etiologic and pathogenic factors, causing peculiarities of the clinical course should be indicated in the diagnosis of septic patient. PMID- 10540550 TI - [Antimicrobial chemotherapy in patients with pyo-septic diseases in intensive care units]. AB - The analysis of the treatment of 900 patients with large festered wounds of various genesis and location for the period from 1973 to 1998 years in the intensive care department has shown, that infection of respiratory ways is encountered in 30% of cases (in patients with nonsporeforming anaerobic bacteria- in 11-12%), bacteriuria--in 70-80%, bacteriamia--in 75% of patients with sepsis. In acute pyogenous diseases of soft tissues microbes from the wounds in monoculture were isolated out in 83.3% of cases, associations of gram-positive and gram-negative bacteria--in 16.7%, in chronic pyogenous diseases of soft tissues--in 60 and 40% of cases, respectively. In sepsis associations of gram positive and gram-negative microbes were isolated in 55.6% of cases. Most often (91%) pathogenic staphylococcus was found in hemocultures. Uring in 62% of cases contained association of gram-positive and gram-negative microorganisms, in sputum gram-positive microflora in monoculture (69%) prevailed. In the group of patients with peritonitis, phlegmon of the anterior abdominal wall, diabetic phlegmon and gangrene, crush syndrome the association of anaerobic and aerobic microflora (from 57.1 to 98.8%) prevailed in the wounds. Application of up-to date antimicrobial means in the intensive care unit resulted in a decrease of mortality rate in sepsis and complicated course of wound infection up to 23%, and in anaerobic nonsporeforming infection--up to 15%. PMID- 10540551 TI - [Modern aspects of wound sepsis in war surgical trauma]. AB - The results of the treatment of 1020 wounded with symptoms of wound infection have been studied. The wounded were admitted with developed complications from the hospitals in Afghanistan. The syndrome of systemic inflammatory reaction detected in 239 (23.4%) wounded was the indication for application of the whole complex of antiseptic treatment. Sepsis in patients with complicated gunshot wounds was revealed only in 54 wounded, i.e. in 5.4%. Early diagnosis and multicomponent therapy resulted in a decrease of mortality rate in wound sepsis up to 9.3%. PMID- 10540552 TI - [Peculiarities of diagnosis and treatment for diabetic feet]. AB - Multifactorial pathogenesis of the "diabetic foot" syndrome suggests advisability to single out some clinical pathogenetic forms, depending on basic causes of the lesions: due to neuropathy, osteoarthropathy, neuro-ischemic factors. The authors has developed the algorithm which enables to diagnose various forms of the syndrome and differentiate the treatment. The most important treatment modalities include unloading and podiatric measures as well as surgery, systemic antibacterial and glucolytic therapy. Conservative or surgical antiischemic measures are indicated only in demonstration of the degree of the ischemic damage in the lower extremities by Doppler ultrasonography and measurement of local oxygenation of the skin. The combined differentiated treatment in 142 patients with diabetic lesions of the feet has resulted 97.2% favourable outcomes in neuropathy and 86.5% successful outcomes in neuroischemic damages. Local oxygenation of dorsal skin of the foot (< 20 mm Hg) indicates that amputation above the ankle is highly probable. PMID- 10540553 TI - [Substantiation and varieties of complex surgical treatment tactics for pyo necrotic forms of diabetic foot]. AB - For the period from 1996 to 1998 in the Division of wounds and wound infection of A.V. Vishnevsky Institute of Surgery 92 patients with pyonecrotic forms of "diabetic foot" underwent thorough examination and treatment. The patients were divided into groups by the form of "diabetic foot": with pyonecrotic forms of "diabetic foot" without critical ischemia (group 1) and with it (group 2). In 18 patients of both groups the data of electron autoradiography were used to reveal peculiarities of the wound process. Group 1 patients had at admittance a large number of neutrophiles in various stages of destruction in biopsies of the wound. In patients of group 2 a great majority of the vessels in biopsies of the wound were in different stages of destruction with lost connections between their separate cells or some of their part absent. Separate cells (endotheliocytes and pericytes) which make up the walls of destroying vessels, were synthesizing RNA and were functionally active. In both groups, the studied parts of the wound before plastic repair of its defect usually represented as well developed granulation tissues with a number of microvessels and cells. Intensive synthesis of PNA in the cells of microvascular wall evidenced of their high functional activity, and the synthesis of DNA in them showed their ability for proliferation, i.g.--for growth. Thus, microangiopathy was reversible, and the solution of the problem of critical ischemia should be considered in the light of macroangiopathy. Thus, in patients of group 1 the cause of pyonecrotic damage consists in infection process, while in patients of group 2--in combination of infection with ischemia of the extremity. In both groups pyonecrotic disease of the extremity ruses at the background of severe disturbances of cellular immunity. PMID- 10540554 TI - [Prevention of extremities amputation in patients with diabetic foot complications]. AB - In order to decrease the number of amputations for "diabetic foot", these patients should undergo elective or delayed operations. It is obligatory before the operation to correct carbohydrate metabolism and hemodynamics. Sodium succinate in combination with conventional angioprotective treatment is used for this purpose. Microcirculation is evaluated using oxymonitor ISM-2 and tetrapolar rheography. Blood flow disturbances are evaluated according to ultrasound dopplerography data. Operations are performed with the use of a primary or delayed suture at the definite level of the extremity where oxygen tension of the skin is not lower than 33 mm Hg, and the index of minute blood flow--not lower than 1.8 ml/min per 100 cm3 of the tissue. When operating on the foot it is obligatory to leave loose excessive brims of tissues to facilitate the placement of broad-grip sutures without tension. It is advisable to use through flowing- aspiration drainage and to perform surgical treatment of the deep phlegmon of the foot through club-shaped approach. PMID- 10540555 TI - [Treatment of diabetic foot complications]. AB - Of 3721 patients with diabetic lesions of the foot treated 2416 (65%) patients have been operated. The rise of urgent hospitalization rate (over 90%) unfavourably influenced the results of the treatment and contributed to increased frequency of high amputations of the extremity. The proposed system for evaluation of the condition and scope of the combined treatment resulted in decrease of mortality among operated patients from 14.7% to 9.8% and among not operated ones--from 20.5% to 5.5%. The suggested classifications of complicated forms of the "diabetic foot" syndrome provided proper evaluation of the prognosis and individual approach to the choice of treatment measures. PMID- 10540556 TI - [Anesthesia in surgical treatment of pyo-necrotic forms of diabetic foot]. AB - Examinations of 48 patients with the syndrome of "diabetic foot" (IV-V degree according to Wagner classification), have shown advantages of prolonged epidural anesthesia (EA) carried out during pre-, intra-, and postoperative (6-7 days) period. This method has relieved pain during postoperative period with minimal impact on carbohydrate metabolism and central hemodynamics. Moreover prolonged EA is a basic prophylactic method against the development of phantom painful syndrome in patients with amputated extremity. The prolonged postoperative EA is a reliable method for modification of surgical stress-response, being especially important in patient, afflicted with a severe form of diabetes mellitus who represent a high surgical and anesthesiological risk group. PMID- 10540558 TI - [Plastic repair of wound defects in purulent surgery of wrist]. AB - Data on the application of plastic methods for filling wound defects after surgical management of 48 patients with purulent disease of the upper extremity are presented. 6 various modes were used depending on the size and location of the wounds. To achieve complete rehabilitation of the operated extremity and favorable functional and cosmetic results the earliest filling of wounds in purulent surgery is obligatory. PMID- 10540557 TI - [Clinical features, diagnosis and treatment for trophic, late radiation ulcers and ulcers undergone malignant transformation]. AB - Clinical and histological examinations of 1562 patients with trophic and advanced radiation ulcers were carried out during 30 years. Malignant transformation of the ulcers has been revealed in 14 of them: sarcoma--in 1, and cancer--in 13. From all the patients with advanced radiation ulcers, malignant transformation was detected in 7.07%, and in cases of ulcers of the other genesis--in 0.81% of cases. Malignant transformation of ulcers of non-radiation genesis occurs on the average after 19 years, and of radiation one--after 3.8 years. Early diagnosis of skin cancer, at the site of the ulceris rather difficult and depends thoroughly on oncological alert and timely morphological examination of various skin areas. A new, rather perspective and relatively simple method of treatment for ulcerative forms of cancer of the skin is photodynamic therapy (PhDT). This method showed to be effective even in those patients who failed conventional methods of treatment (relapses), or had contraindications. PhDT has broaden the armory of the oncologists and oncodermatologists for treatment of complicated forms of cancer of the skin. PMID- 10540559 TI - [Choice of anesthesia in patients with suppurative surgical infections and diabetes mellitus]. AB - 500 anesthesias were performed in patients with surgical diseases in the presence of diabetes millitus (90% of patients with II type diabetes). The methods of intravenous anesthesia are proposed for short-term procedures with preservation of spontaneous ventilation and stable hemodynamics. The method of epidural anesthesia with introduction of major dose of anesthetic in two stages was developed for the operation on the lower extermities, it provides for stable hemodynamics and adequate anesthesia. Factors, influencing the choice of anesthesia were determined. Basic principles of intensive care in patients with surgical infection in diabetes mellitus were established. PMID- 10540560 TI - [Multicomponent dressing media in treatment of suppurated wounds]. AB - The multicomponent dressing medium (MDM) has been developed for treatment of festered wounds. The first layer consists of hydrofibrosed gauze with chlorhexidine being introduced into its structure in quantity of 0.2-0.5 mass %. The second layer of MDM is chlorhexidinum salt of monocarboxylcellulose with carboxyl group content of 12-22 mass % and chlorhexidine content being 1-2 mass %. The third layer is polyamide pellicle with the pores less than 1 micron in diameter. The created MDM possess significant capillar-transport, absorption, anti-adhesive and prolongated antimicrobial properties, prevents development of secondary infection and "hotbed" effect in the wound. PMID- 10540561 TI - [Clinico-cytological features of local treatment for slow-granulating soft tissue wounds with 0,2% curiosine solution in phase II of healing process (data from registration studies)]. PMID- 10540562 TI - [Laser autofluorescent spectroscopy--new method for express-diagnosis in surgery]. PMID- 10540563 TI - [Asthma phenotype during the first six years of life]. AB - Asthma is one of the main wheezing causes during the first years of life. In our country it is a common respiratory chronic illness but insufficient studies still exist on the asthma phenotypes during the first years of life. OBJECTIVE: To know the phenotype of asthma in a group of children younger than 6 years old. MATERIAL AND METHODS: 185 children of both sexes were studied with antecedent of having presented wheezing (tree episodes or more) and it registered data about the antecedents of family and personal allergy, dietary habits during the first year of life, infections, data on the beginning and the evolution of the condition, and they were practiced determinations of peripheral eosinophilia, total serum IgE and gastric gammagram to discard illness for gastroesophageal reflux. All were carried out skin tests for foods and aeroallergens. RESULTS: In the group of 185 patients of both sexes, they had data that supported the allergic process, in 137. It was correlated the atopy antecedents significantly, positive skin tests, eosinophilia (more than 300), with elevated IgE for the age (p < 0.05). The gastric gamagrama was carried out in 144 patients, of which were positive results for gastroesophageal reflux in 64 (44%) and in 79 (54%) it was reported doubtful or negative. It was related the gastroesophageal reflux presence and the positive skin tests significantly (p < 0.05). CONCLUSIONS: The more common phenotype of asthma in our patients corresponds to a wheezing pattern that persist after the 3 years old, in relation to an allergic component. Furthermore in most of those children a positive gastroesophageal reflux was an important finding. PMID- 10540564 TI - [Dry powdered formoterol, twice a day versus aerosolized salbutamol, four times a day, in patients with stable asthma]. AB - MATERIAL AND METHODS: The objective of this multicenter, open randomized study was to compare the effect of inhaled formoterol (dry powder with ISF system) 12 mcg twice daily versus salbutamol (200 mg qid) in patients with bronchial asthma. A total of 160 patients were evaluated with such diagnosis in four specialized centers; the main variable was the maximal respiratory flow (MEF) assessed prior to drug administration (morning and afternoon). In addition to this, vital capacity (VC), forced respiratory volume over one second (FEV-1), and other safety variables were also determined. RESULTS: With regard to MEF the administration of formoterol showed better results (P < 0.05) ever since the first month of treatment. The frequency of adverse events was similar between treatment groups; the formoterol group had fewer nightly wake up periods. CONCLUSIONS: That formoterol is a safe, efficacious and long-acting Beta 2 agonist which can be administered twice daily. PMID- 10540566 TI - [Leukocyte adhesion deficiency syndrome: case report]. AB - INTRODUCTION: Leukocyte adhesion deficiency syndrome (LAD) is an altered phagocytic disorder characterised by the deficiency of one or several integrins which are included within the adhesion molecules group and cell surface receptors superfamily. OBJECTIVE: To describe the clinical features of a rare primary immunodeficiency case. CLINICAL CASE: A nineteen days-old male newborn was referred to the pediatrics infectology service because a 15 days clinical course characterised by delayed cord detachment; fever and skin lesions in several arcas that evolved to, cellulitis and dermal necrosis: Then he was admitted with the diagnosis of septicemia secondary to omphalitis. There were a partial response to antimicrobial treatment. Thereafter he had recurrent respiratory and gastrointestinal fungal and bacterial infections. Then he suffered psychomotor impairment and severe malnutrition. The patient died because septicemia at 5 months-old. WBC counts showed persistent leukocytosis between 42,000 and 133,000 cells/mm3, mostly neutrophils (64%-88%). We also found defective neutrophil quimiotaxis. By flow cytometer it was detected CDII/18 adhesins deficiency. Otherwise immunological, bone marrow biopsy and viral tests were, normal. CONCLUSIONS: Although its prevalence is rare, leukocyte adhesion defects must be considered in those patients with delayed cord detachment, recurrent severe infections and both persistent and elevated neutrophilia and with other primary and secondary immunodeficiencies previously discarded. PMID- 10540565 TI - [Air flow measurement in children with mite allergies before and after skin prick test]. AB - OBJECTIVE: To determine the obstruction of the air flow by flujimetry subsequent to the application of tests cutaneous specific (Dermatophagoides pteronisinnus) in sensitive children. MATERIAL AND METHODS: They were studied 44 patient of one and other sex with diagnostic of asthma and antecedent of sensibility to the mite Dermatophagoides pteronisinnus through tests cutaneous for prick, with an average age of 6 to 16 years, captured of the external consult of the allergy service of the Hospital Infantil de Mexico Federico Gomez. It is a longitudinal study, prospective, blind, cross, in the one which previously was made a challenge test with antigen standardized of Dermatophagoides pteronisinnus or glycerine, previous reading of flujimetry. RESULTS: Of the patients challenged with antigen in 23 there was decrease of the respiratory maximum flow, with a p < 0.05, but without clinical meaning. When they were challenged with placebo only it reduced in patient seven the respiratory maximum flow, also significative statistically, but without clinic relevancy. CONCLUSION: The cutaneous tests are a useful tool in allergy and sure, since almost they do not produce serious systemic reactions. PMID- 10540567 TI - [Allergic fungal sinusitis: recent developments]. AB - Allergic fungal sinusitis is a recently reported disease of the nose and paranasal sinuses. Since the first reports by Lamb et al and Katzenstein there has been controversy about its diagnosis and treatment. Recently diagnostic criteria have been suggested. To our judgement they have a high degree of specificity. Allergy to fungi elements is essential. Currently surgical treatment consist in an adequate ventilation of the nose and paranasal sinuses followed by the use of oral and topical steroids. Immunotherapy is controversial and more prospective studies are needed to evaluate its possible use. PMID- 10540568 TI - [Alcohol--risk factor for overweight]. AB - Overweight is accepted as risk factor for various diseases such as hypertension and diabetes mellitus, dyslipidemias and cardiovascular diseases. In this study the correlation of alcohol consumption and overweight was investigated among members of the Swiss association of inn keepers and hotel managers (Gastrosuisse). A questionnaire was mailed to all members of the Gastrosuisse in the cantons of Zurich, Graubunden, and the German speaking part of Freiburg. The following parameters were assessed: sex, birth date, weight, height, and cardiovascular risk factors. The frequency of alcohol consumption per week, the type and amount of alcoholic beverage (wine, beer, liquor) consumed for each drinking occasion were asked for. The daily alcohol intake (g/d) was computed. The health status, physical activity levels at work and for recreational activities, tendency for abdominal fat accumulation, self-judged stress, fat content of the diet were assessed with visual analogue scales. The present analysis was limited to non-smoking men (n = 573). A j-shaped relationship was found between alcohol consumption and the body weight: the lowest body weight was found in subjects with a mean daily consumption between 13 and 36 g/d (26.3 +/- 0.3 and 26.2 +/- 0.4 kg/m2, mean +/- sem). Non-consumers and subjects with an intake > 36 g/d had a higher body weight. The relationship was independent of age and physical activity levels as well as the beverage type. Limiting the analysis to daily alcohol consumers only a linear relationship was found. Our data suggest that alcohol consumption, especially daily consumption, should be regarded as a risk factor for an increased body weight and obesity. PMID- 10540569 TI - [Pericardial effusion in the hospital--diagnosis and therapy]. AB - A pericardial effusion is a relatively common disease confronting the clinician. The most frequent causes are neoplasias (lung, breast and ovarial carcinoma, leukemia and lymphoma) uremia or idiopathic. Infections (frequently virus and seldom bacteria), myocardial infarction and rheumatic disease are also common. We present the clinical picture, the differential diagnosis and the various investigations of the pericardial effusion. PMID- 10540570 TI - [Unexpected development during rehabilitation for suspected rheumatic disorder]. AB - The 57 year old woman presented with diffuse muscle spasms and delirium. Prior to presentation, she complained of progressive muscle pain, weakness and a weight loss of 10 kg over several months. Laboratory investigation showed hypopituitarism and a syndrome of inappropriate antidiuretic hormone secretion. Magnetic resonance imaging revealed an empty sella. The primary and secondary syndromes of empty sella are discussed. PMID- 10540571 TI - [Acquired angioedema during ACE-inhibitor therapy]. PMID- 10540573 TI - Tuberculous pancreatic abscess in HIV-positive patients. A report of 3 cases and a review of the literature. AB - Three cases of tuberculous pancreatic abscess (TPA) in HIV-positive patients are reported. Pancreatic tuberculosis (PTB) is a rare pathological entity with nonspecific symptomatology that presents a diagnostic challenge. Ultrasound or computed tomography-guided fine-needle aspiration biopsy (FNAB) is recommended, as this may be diagnostic and negate the need for operative intervention. PMID- 10540572 TI - J.H. Louw Memorial Lecture. From puppy dogs to molecules: small-bowel atresia and short-gut syndrome. PMID- 10540574 TI - Slow continuous dilatation of oesophageal strictures using the Didcott dilator, with reference to its wider use. AB - Acute dilatation or bouginage of strictures gives only temporary relief, and slow continuous dilatation was therefore tried and found to give superior results in treating benign and malignant strictures, particularly of the oesophagus. Slow stretch methods are discussed and compared with other methods. Methods are described that were evolved for dilating both by the 'acute' and slow-continuous methods, including use of the Didcott dilator (DD), invented in 1956. For oesophageal cancer this, combined with brachytherapy, has resulted in increased longevity and quality of life. Mortality from the dilatation and introduction of a DD for a week, followed by its removal without anaesthesia, is less than 2%. Relief of dysphagia lasts 2-10 months. Thereafter the procedure can be repeated and finally, when the patient is obviously near-terminal, a permanent indwelling stent can be used. This can be a modified DD stent or a Livingstone or Celestin tube. These are also used in tracheo-oesophageal fistulas. Complete cure is often possible in benign strictures, especially if short. PMID- 10540575 TI - Pyogenic spondylitis. AB - Twenty-nine patients with pyogenic vertebral osteomyelitis were reviewed retrospectively after an average follow-up of 3.7 years. We identified 17 patients with predisposing factors (10 diabetes, 4 urinary tract infection, 2 HIV positive, 1 rheumatoid arthritis). No patient presented with a febrile illness. The lumbar spine was involved in 15 patients. Eighteen patients had neurological impairment at presentation. Eleven patients who were neurologically intact had needle biopsies and the remaining 18 patients who were neurologically compromised had an open decompression. Staphylococcus aureus was cultured in 14 patients. Although spinal tuberculosis is relatively common in our environment it is important to obtain a tissue diagnosis in order to exclude pyogenic vertebral osteitis. PMID- 10540576 TI - Guidelines on safety standards for instruments used in minimal access surgery (MAS). Recommendations of the South African Society of Endoscopic Surgeons (SASES). PMID- 10540577 TI - [Anticardiolipin antibodies in patients with Buerger's disease]. AB - The aim of this study was to estimate the incidence of occurrence of anticardiolipin autoantibodies in patients with thromboangitis obliterans (TAO). Patients with Buerger's disease had statisticaverified significant higher frequency of anticardiolipin IgM antibodies than control group. This antibody may play a role in pathogenesis of TAO, although this results should be verified because of the small number of patients and diagnostic criteria. PMID- 10540578 TI - [Surgical intraduodenal drainage of bile duct: an introductory report]. AB - Reconstructing operations, performed on extrahepatic bile ducts, usually need protection of anastomosis by means of drainage. Classic Kehr's drainage is loaded with the risk of complications. Since 1993 the authors have been applying the alter way of drainage: an intraduodenal drainage by endoprothesis Y (DEY). Endoscopical removal of the drain was performed in 3-4 months after the operation. This way of drainage was used between III 1993--V 1997 in 23 patients. We did not observed any complications or pancreas reactions. Clinical control of patients was made after operation with ECW. In 4 cases the results of treatment were unsuccessful. In other 16 patients results of performed operations were favourable. CONCLUSION: Proposed way of protection of bile duct's reconstruction's site using DEY decreases the number of complications connected with classic Kehr's drainage. PMID- 10540579 TI - [Neonatal polycythemia: risk of microcirculation disorders]. AB - The clinical picture and results of treatment of 13 polycythaemic neonates were presented. The possibility of permanent organs failure and even death caused by polycythaemia was considered. PMID- 10540580 TI - [Extended small bowel resection with right hemicolectomy after massive superior mesenteric artery embolism, management in early postoperative period]. AB - Intensive care management in recent postoperative period in nine patients with superior mesenteric artery embolism was provided. In all patients during surgical treatment an extensive resection of small bowel and right colectomy were performed. On the basis of physiopathological mechanisms of occlusional bowel ischemia and septic shock development the appropriate therapeutic procedure during pre and postoperative period was submitted. The authors suggest usefulness of the antibiotic prophylaxis in patients with high risk of measenteric embolism in order to decrease the dynamics of septic complications in the cases with bowel necrosis. PMID- 10540581 TI - [Comparative analysis of occurrence and susceptibility to gram-negative bacilli isolated from intubation tubes and tracheostomy tubes in patients at intensive care units]. AB - The purpose of this work was to determine the frequency of occurrence and susceptibility of Gram-negative bacilli isolated from intubation tubes and tracheotomy tubes in neonates and adults in intensive care units. Nonfermenting rods were the most often isolated from the tubes especially Pseudomonas aeruginosa from 39.2% to 45.2% of Gram-negative bacilli and Acinetobacter spp. from about 10.9% of Gram-negative bacilli. The rods of Enterobacteriaceae family were isolated too. It was found that the most frequently isolated bacteria was Escherichia coli--from 17.8% to 21.4% of Gram-negative bacilli. The other rods were isolated occasionally. PMID- 10540582 TI - [The role of methotrexate in the management of severe steroid-dependent asthma]. AB - Low-dose methotrexate therapy has been recently proposed as an alternative therapy for patients with severe steroid-dependent asthma. This study examined the role of low-dose methotrexate in the management of asthma. Fourteen subjects with stable, severe asthma were enrolled. Patients received orally methotrexate 15 mg/week. Only 2 of 14 patients responded to methotrexate 2 to 3 years after the study's termination. The doses of steroids could be reduced. Side effects were observed in nearly 50% of patients; 7 subjects required the discontinuation of treatment. Low-dose methotrexate therapy may have probably a role in small selected group of steroid-dependent asthmatics. PMID- 10540583 TI - [Comparative evaluation of postoperative complication in the reconstructive surgery of the esophagus]. AB - The paper presents 78 patients with stenosis of the middle thoracic segment of the esophagus. The substitute, being pedunculated intestinal graft, was brought to the neck through the retrosternal space. Early post-operative complications occurred in 8.9% of patients. Post-operative mortality rate was 7.6%. The most dangerous complication after esophageal plastic surgery is blood supply insufficiency. It was observed in 2 cases. Other complications included anastomotic leaks and respiratory distress syndrome. The authors emphasize that frequency of post-operative complications and mortality are related with the patient general condition and extent of the surgery. PMID- 10540584 TI - [The management in vascular trauma of extremities]. AB - Between 1990-1998 in the Department of Vascular Surgery, Medical Academy in Wroclaw, 104 patients were operated due to artery trauma: 28 women, 76 men. Mean age was 37 years. In 58 cases acute ischaemia occurred, in 46--haemorrhage. Preoperative procedure consisted of preparation and diagnostics, in some cases angiography. Predominantly autogenous material was used for artery reconstruction. In 25 cases prosthetic grafts were implanted. In 53 patients (79.1%) good results were obtained, complications occurred in 14 cases (20.9%). PMID- 10540585 TI - [Clinical picture in sleep apnea syndrome]. AB - The aim of the study was to assess the clinical picture of patients with sleep apnea syndrome (SAS). The study group consisted of 54 patients (51 men, 3 women) mean age 49.7 +/- 8.7 years, mean body mass index (BMI) 33.1 +/- 5.8. In all cases polisomnography confirmed the diagnosis of SAS. Mean apnea and hypopnea index (AHI) was 66.6 +/- 30.7 and mean minimum arterial blood oxygen saturation was 67.57 +/- 11.58%. It allowed us to qualify 69.4% of patients to the group with a severe SAS. Snoring (93%), apneas (83%), excessive daytime sleepiness (80%), morning weakness (81%), nycturia (66%) were the most common symptoms. The most frequently accompanying diseases in patients with SAS were overweight (89%), depression (67%), arterial hypertension (51%), impaired glucose tolerance (41%). PMID- 10540586 TI - [Diffuse alopecia in women: classification and views of etiopathogenesis]. AB - Contemporary views of feminine diffuse alopecia's etiopathogenesis have been presented on the grounds of literature data. In the considerable part of cases feminine diffuse alopecia is a symptom of many systemic diseases and hypovitaminosis. Sideropenia is often diagnosed. However it seems that neuroses and psychic traumas provoke alopecia in some cases. PMID- 10540587 TI - [Intravascular ultrasound]. AB - An intravascular ultrasound (IVUS) is a new cardiological diagnostic technique which provides detailed cross-sectional images of the vessel wall, allows to investigate lumen, as well as structure of the coronary artery. This imaging technique has the unique potential to provide an image of atherosclerotic plaque, characterizes its composition, and severity of stenosis. "Soft" plaque, dense fibrous "hard" plaque, calcification and thrombosis have all been identified using IVUS. Intravascular ultrasound imaging has become a method for supporting interventional decision making by selecting patients who are to undergo Percutaneous Transluminal Coronary Angioplasty, Stent placement, Directional Coronary Atherectomy or Laser Angioplasty. Quality of measurements should allow more widespread use to guide interventional techniques. A series of IVUS imagings could assess the influence of plaque composition on mechanism of immediate lumen enlargement after revascularization. With utilization of IVUS guidance and high pressure balloon inflation--stent implantation could now be performed "optimally" and safely without anticoagulation. This original diagnostic tool is becoming a clinically useful adjunct to angiography in the assessment coronary arteries of heart transplant recipient and patients with higher risk of restenosis. PMID- 10540588 TI - [The role of vasoactive intestinal peptide in human body]. AB - The quick progress of medical sciences changed somewhat our point of view on the role of Vasoactive Intestinal Peptide in human body. In the following article we have tried to gather all the latest information concerning VIP and its regulatory function. The role in menstrual cycle regulation, in prolactine and in hypothalamo-pituitary-adrenal axis secretion as well as in pancreas' hormones secretion has been described. We also emphasized the role in maintaining the blood pressure. The influence on the digestion, hematopoiesis, cell growth and differentiation was also discussed. PMID- 10540589 TI - [Sigmoid-vesical fistula in the course of long-term conservative treatment for recurrent sigmoid diverticulitis]. AB - The authors describe a case of sigmoidovesical fistula developed in a man with sigmoid diverticular disease. Appropriate surgical procedure has been done thanks to proper diagnosis. In authors' opinion early operation performing is the best therapy of colon diverticulitis even if a patient suffer from other serious disorders. PMID- 10540590 TI - [Low obturation ileus caused by gallstone incarceration in sigmoid colon]. AB - Authors presents two cases of low obturation ileus caused by gallstone which had obstructed sigmoid colon. They pay attention to diagnostic problems and usefulness of sonography and X-ray techniques also in case of biliary ileus. Authors shows difficulties in surgical procedures, prefering enterotomy and evacuation of concrement, without simultaneous internal biliary fistula liquidation, especially in cases of increased risk and in old age patients. PMID- 10540591 TI - [Vaginal evisceration with secondary torsion of small intestine]. AB - The case of vaginal evisceration in 87 year old woman is described. The evisceration occurred on a day preceding the admission of the patient to the hospital. On admission the intestinal ischemia due to torsion of the bowel loop was noticed and was immediately reversed in emergency room. On laparotomy the viability of the intestine was confirmed and repair of vaginal tear was performed. The brief review of the literature is also presented. PMID- 10540592 TI - [A palatine tonsil as an origin site of non-Hodgkin's lymphoma: a report of two cases]. AB - The authors describe the clinical picture of non-Hodgkin's lymphoma originated from palatine tonsil in two children: 7-year old boy and 15-year old girl. After intensive polychemotherapy the complete remission was achieved. The period after the end of treatment is now 5 months and 24 months long, respectively. PMID- 10540593 TI - Racism as a stressor for African Americans. A biopsychosocial model. AB - Various authors have noted that interethnic group and intraethnic group racism are significant stressors for many African Americans. As such, intergroup and intragroup racism may play a role in the high rates of morbidity and mortality in this population. Yet, although scientific examinations of the effects of stress have proliferated, few researchers have explored the psychological, social, and physiological effects of perceived racism among African Americans. The purpose of this article was to outline a biopsychosocial model for perceived racism as a guide for future research. The first section of this article provides a brief overview of how racism has been conceptualized in the scientific literature. The second section reviews research exploring the existence of intergroup and intragroup racism. A contextual model for systematic studies of the biopsychosocial effects of perceived racism is then presented, along with recommendations for future research. PMID- 10540594 TI - Does cigarette smoking cause stress? AB - Smokers often report that cigarettes help relieve feelings of stress. However, the stress levels of adult smokers are slightly higher than those of nonsmokers, adolescent smokers report increasing levels of stress as they develop regular patterns of smoking, and smoking cessation leads to reduced stress. Far from acting as an aid for mood control, nicotine dependency seems to exacerbate stress. This is confirmed in the daily mood patterns described by smokers, with normal moods during smoking and worsening moods between cigarettes. Thus, the apparent relaxant effect of smoking only reflects the reversal of the tension and irritability that develop during nicotine depletion. Dependent smokers need nicotine to remain feeling normal. The message that tobacco use does not alleviate stress but actually increases it needs to be far more widely known. It could help those adult smokers who wish to quit and might prevent some schoolchildren from starting. PMID- 10540595 TI - Drug use harm. PMID- 10540596 TI - Reducing the harm of a failed drug control policy. PMID- 10540597 TI - The inferior petrosal sinus: assessment by transcranial Doppler ultrasound using the suboccipital approach. AB - Recently, intracranial veins and sinuses have been successfully insonated using the transtemporal and transoccipital approaches by transcranial Doppler ultrasound. The purpose of this study was to prove the capacity of the Doppler method to evaluate the inferior petrosal sinus via the suboccipital approach. Venous transcranial ultrasound was performed with a range-gated 2-MHz transducer in 80 healthy volunteers and patients without central nervous system disorders ranging in age from 15-84 years (mean +/- standard deviation [SD], 37.6 +/- 15.2 years). A venous signal with a flow directed toward the probe was considered to originate from the inferior petrosal sinus because of its proximity to the basilar artery. The inferior petrosal sinus was insonated in 96.3% of the cases at least on one side. It was found bilaterally in 48 (60%), on the right side in 74 (92.5%), and on the left side in 51 (63.8%) subjects, respectively. Mean blood flow velocity ranged from 8-53 cm/s (mean +/- SD, 19.6 +/- 8.7 cm/s). A significant age dependency of venous velocities was found. Weak but significant side-to-side differences were observed, reflecting the known right-sided predominance of venous outflow in humans. Using the suboccipital approach, the inferior petrosal sinus can be insonated in a high percentage of subjects without major difficulties and is defined by its vicinity to the basilar artery. PMID- 10540598 TI - Blood flow velocities in the vertebral veins of healthy subjects: a duplex sonographic study. AB - Along with the jugular veins, the vertebral veins serve as an important pathway for venous blood returning from the brain. In this study, the authors report duplex sonographic findings in 138 healthy subjects without central nervous disease. Successful insonation was possible in 70.7% of all examined vessels. Bilateral insonation was achieved in 86 subjects (62.3%). Only 1 vertebral vein was detected in 23 persons (16.7%), whereas no vein was found in 29 persons (21%). The authors observed a marked variation of peak flow velocities ranged (5 81 cm/s, mean +/- standard deviation, 23.9 +/- 12.3 cm/s). No significant gender related or side-to-side differences or age influences on flow velocities were detected. The authors' findings may be of relevance when discussing flow velocities in the vertebral veins in cases of cerebrovenous disorders (e.g., dural sinus thrombosis) or in patients after neck dissection. PMID- 10540599 TI - Does an increase in sulcal or ventricular fluid predict where brain tissue is lost? AB - Quantitative volumes of cerebrospinal fluid (CSF) and brain tissue were measured on magnetic resonance images (MRIs) of 287 individuals from 5 diagnostic groups: Alzheimer's disease (AD), chronic alcoholics (ALC), individuals positive for human immunodeficiency virus (HIV), schizophrenia subjects (SZ), and normal comparison subjects (NC) older than 50 years of age. Within each group, mean volumes were calculated for ventricular CSF, cortical (sulcal) CSF, cortical gray matter, total white matter, basal ganglia gray matter, and thalamic gray matter. Correlations of CSF measures with brain tissue measures were determined, and multiple regression analyses were performed to try and predict volume of gray matter or white matter region from volume of CSF compartment. Results indicated the following: 1. Enlarged cortical fluid volume significantly predicts cortical gray matter deficits for subjects with AD and those who are ALC and SZ but not for subjects with HIV or NC. 2. Enlarged cortical fluid volume also significantly predicts white matter deficits in all five groups. 3. Enlarged ventricular fluid volume significantly predicts basal ganglia deficits in AD, HIV, and NC but not in SZ or ALC. 4. Enlarged ventricular volume has no predictive value for thalamic volume for any of the groups. This study supports the clinical practice of predicting brain tissue volume loss from CSF enlargement but not for all brain regions in all diagnoses. PMID- 10540600 TI - The role of quantitative ictal SPECT analysis in the evaluation of nonepileptic seizures. AB - Nonepileptic seizures may represent difficult diagnostic problems. Identifying their presence and frequency is critical for determining appropriate treatment. The authors investigated the value of quantitative perfusion changes as measured by ictal single-photon emission tomography (SPECT) difference images in differentiating nonepileptic from epileptic seizures. Eleven patients with a clinical suspicion of nonepileptic events had ictal and interictal technetium-99m hexamethylpropylene amine SPECT scans during continuous audiovisual surface electroencephalogram (EEG) monitoring. The authors analyzed perfusion difference images based on registration, normalization, and subtraction of ictal and interictal SPECT images. The difference images were registered to each patient's magnetic resonance imaging scan to anatomically localize ictal perfusion changes. Three of 11 patients also carried the diagnosis of epilepsy and were taking antiepileptic medication. Five patients were taking antiepileptic drugs, but the diagnosis of epilepsy was not confirmed. In all patients, continuous video EEG monitoring revealed no ictal EEG findings. In nine of these patients, visual interpretation of ictal SPECT was suggestive of localized increased (n = 6) or decreased perfusion (n = 3). In all patients, however, no blood flow changes were noted on quantitative SPECT analysis with injections performed during the seizure like event, suggesting the diagnosis of pseudoseizures. The authors' results suggest that quantitative ictal SPECT analysis is a useful tool in the diagnosis of nonepileptic seizures. PMID- 10540601 TI - Brain uptake and plasma metabolism of [11C]vinpocetine: a preliminary PET study in a cynomolgus monkey. AB - Vinpocetine, a vinca alkaloid, is a widely used therapeutic agent in patients with acute and chronic stroke. To reveal the mechanisms of vinpocetine action in the brain, vinpocetine was labeled with 11C. Positron emission tomography (PET) was used to determine the uptake and distribution of [11C]vinpocetine in brain regions and the trunk of a cynomolgous monkey in two independent measurements. The concentration of vinpocetine and its labeled metabolites was determined in blood and plasma using high-performance liquid chromatography (HPLC). Almost identical measurements were obtained in the two independent studies. After intravenous administration, following an initial peak, the total concentration of radioactivity in blood was relatively stable with time, whereas the concentration of the unchanged compound decreased with time in an exponential manner. The uptake of [11C]vinpocetine in brain was rapid, and 5% of the radioactivity totally injected was present in the brain 2 minutes after drug administration, indicating that the compound entered the brain readily. The radioactivity uptake was heterogeneously distributed among brain regions and was highest in the thalamus, the basal ganglia, and certain neocortical regions. The high brain uptake and the heterogeneous regional distribution indicate that direct central nervous system (CNS) effects of vinpocetine must be considered as explanation for the therapeutic effects. The detailed exploration of this suggestion requires further studies. PMID- 10540602 TI - Single-photon emission computed tomography of the dopamine transporter in parkinsonism. AB - The known dopaminergic abnormalities in Parkinson's disease have facilitated the development of radiolabeled biomarkers for diagnostic and research applications in humans. Presynaptic, intrasynaptic, and postsynaptic imaging now is possible using single-photon emission computed tomography. In particular, the development of new radiotracers that target the dopamine transporter located on degenerating dopamine neurons in Parkinson's disease and related disorders is directly relevant to improved clinical diagnosis, disease monitoring, and assessment of putative neuroprotective strategies in patients. In addition, the ability to characterize in vivo neuronal degeneration in these disorders provides a powerful research tool to better understand the natural course of these disorders and could provide clues to etiology. PMID- 10540604 TI - Hyperacute cerebral enhancement: the earliest predictor of hemorrhage by MR imaging? AB - A test to detect very early hemorrhage in acute cerebral infarct could offer a substantial increase in the safety and success of advanced stroke therapies, particularly when the use of thrombolytic therapies is contemplated. Currently, computed tomography is the standard test for the detection of cerebral hemorrhage but is not a valid predictor of potential areas of hemorrhagic transformation. A technique to evaluate the risk of hemorrhagic transformation in infarcted cerebral tissue has been conducted with contrast-enhanced magnetic resonance imaging in various animal stroke models. Knight demonstrated Gadolinium-DTPA enhancement in the territory of occluded vessels immediately in rats after reperfusion. Gadolinium enhancement was thought to predict areas of hemorrhagic transformation. Yenari and associates demonstrated in rabbit models that contrast enhanced T1-weighted scans can reveal regions of blood-brain barrier disruption, characterized as hemorrhagic transformation in ischemic tissue. The authors report a clinical example in which hyperacute contrast-enhanced magnetic resonance imaging was the first indication of hemorrhagic transformation within 24 hours of onset of an acute cerebral infarct. PMID- 10540603 TI - Development and applications of 4-D ultrasound (dynamic 3-D) in neurosonology. AB - The development and application of color-coded data in three-dimensional (3-D) reconstruction or four-dimensional (4-D) imaging (equal to dynamic 3-D) are demonstrated. In 4-D imaging, electrocardiography-triggered data acquisition of consecutive phases during the heart cycle are stored to form a multiphase 3-D data set. The option of color-coded data gives a new insight into such hemodynamic information. In the past, 3-D reconstructions were simple unicolor images, as in power mode, and the color-coded hemodynamic information was lost. These new options are presented here, along with color-coded data in examples of angiographically controlled pathologic results in extracranial and intracranial vessels. PMID- 10540605 TI - Transcranial sonography in a patient with a persistent trigeminal artery. AB - Transcranial ultrasonography was performed in a patient with a persistent trigeminal artery, a remnant of fetal cerebral circulation that bridges the carotid and basilar territories. The study revealed low flow and reversal of flow in the vertebral and basilar arteries, respectively. The interpretation and significance of these findings are discussed. PMID- 10540606 TI - Monitoring of the extracranial and intracranial course of single emboli of cardiac origin: a preliminary report. AB - Simultaneous monitoring of emboli in extracranial and intracranial arteries recorded with identical probes, in a patient with an artificial cardiac valve, allowed the identification and characterization of pairs of signals, which most likely represent single emboli flowing through the common carotid artery into the middle cerebral artery. This technique offers new insight into emboligenesis with obvious therapeutic implications. PMID- 10540608 TI - Radial microbrain form of micrencephaly: possible association with carbamazepine. AB - A premature infant exposed to carbamazepine in utero had a markedly undersized brain on cranial ultrasonogram. Postmortem examination of the brain revealed no evidence of hypoxic-ischemic injury, hemorrhage, infarction, congenital infection, or calcification. The normal cortical gyral pattern, normal residual germinal matrix, and normal cortical lamination suggested the diagnosis of a radial microbrain form of micrencephaly. PMID- 10540607 TI - Basilar artery dissection in a young woman: a case report. AB - The case of a young woman with basilar artery dissection, possibly precipitated by trauma, is presented, and the literature is reviewed. PMID- 10540609 TI - Hemispheric language dominance testing by means of fMRI. PMID- 10540610 TI - Nurturing our own. PMID- 10540611 TI - Managing postoperative CABG pain. PMID- 10540612 TI - Teaching pursed-lip breathing. PMID- 10540613 TI - Living with irritable bowel syndrome. PMID- 10540614 TI - When to use hydrogel dressings. PMID- 10540615 TI - Myths & facts ... about systemic lupus erythematosus. PMID- 10540616 TI - Action stat. Retinal detachment. PMID- 10540617 TI - Riding out a diabetic emergency. AB - Acute complications of diabetes are like a runaway roller coaster. Diabetes or its treatment can rocket your patient's blood glucose level to dizzying heights or plunge it to life-threatening lows. Hypoglycemia, the most common endocrine emergency, typically occurs in a known diabetic patient whose therapy with insulin or oral diabetes agents goes awry. At the opposite extreme, soaring blood glucose levels mark the acute conditions diabetic ketoacidosis (DKA) and hyperosmolar hyperglycemic nonketotic state (HHNS). These complications may send the patient to the emergency department (ED) before he even knows he has diabetes. In this article, I'll explain how these problems develop and spell out nursing measures to get your patient back on track. PMID- 10540618 TI - I.v. infiltration. Not just a peripheral problem. PMID- 10540619 TI - How to build your "hope skills". PMID- 10540620 TI - Recognizing elder abuse. PMID- 10540621 TI - Pretty baby girl. PMID- 10540622 TI - Getting a slant on syncope. PMID- 10540623 TI - It's a wonderful life. PMID- 10540624 TI - Taking steps to calm restless legs syndrome. PMID- 10540625 TI - Fexofenadine and prolonged QT intervals. PMID- 10540626 TI - Clamping down on circumcision. PMID- 10540627 TI - Using SOAP, SOAPIE, and SOAPIER formats. PMID- 10540628 TI - Growing up without Rebekah. PMID- 10540629 TI - 7 strategies to protect your heart. PMID- 10540630 TI - Exploding myths about women and heart disease. PMID- 10540632 TI - Nutrition matters. PMID- 10540631 TI - How to take (and keep) the weight off. PMID- 10540633 TI - The future of medicine and radiology: Part II. PMID- 10540634 TI - New digital mammography systems may require different X-ray spectra and, therefore, more general normalized glandular dose values. PMID- 10540635 TI - Probably benign breast lesions: when should follow-up be recommended and what is the optimal follow-up protocol? PMID- 10540636 TI - Six-month follow-up: an alternative view. PMID- 10540637 TI - Glandular breast dose for monoenergetic and high-energy X-ray beams: Monte Carlo assessment. AB - PURPOSE: To extend the utility of normalized glandular dose (DgN) calculations to higher x-ray energies (up to 120 keV) and to provide the tools for investigators to calculate DgN values for arbitrary mammographic and x-ray spectra. MATERIALS AND METHODS: Validated Monte Carlo methods were used to assess DgN values. One million x-ray photons (1-120 keV, in 1-keV increments) were input to a semicircular breast geometry of thicknesses from 2 to 12 cm and breast compositions from 0% to 100% glandular. DgN values for monoenergetic (1-120 keV) x-ray beams, polyenergetic (40-120 kV, tungsten anode) x-ray spectra, and polyenergetic mammographic spectra were computed. Skin thicknesses of 4-5 mm were used. RESULTS: The calculated DgN values were in agreement within approximately 1%-6% with previously published data, depending on breast composition. DgN tables were constructed for a variety of x-ray tube anode-filter combinations, including molybdenum anode-molybdenum filter, molybdenum anode-rhodium filter, rhodium anode-rhodium filter, tungsten anode-rhodium filter, tungsten anode-palladium filter, and tungsten anode-silver filter. DgN values also were graphed for monoenergetic beams to 120 keV and for general diagnostic x-ray beams to 120 kV. CONCLUSION: The tables and graphs may be useful for optimizing mammographic procedures. The higher energy data may be useful for investigations of the potential of dual-energy mammography or for calculation of dose in general diagnostic or computed tomographic procedures. PMID- 10540638 TI - Cost-effectiveness of MR imaging and core-needle biopsy in the preoperative work up of suspicious breast lesions. AB - PURPOSE: To assess the clinical and economic consequences of the use of preoperative breast magnetic resonance (MR) imaging and core-needle biopsy (CNB) to avert excisional biopsy (EXB). MATERIALS AND METHODS: A decision-analytic Markov model was constructed to compare MR imaging, CNB, and EXB without preoperative testing in a woman with a suspicious breast lesion. Stage-specific cancer prevalence, tumor recurrence, progression rates, and MR imaging and CNB sensitivity and specificity were obtained from the literature. Cost estimates were obtained from the literature and from the Medicare fee schedule. RESULTS: EXB without preoperative testing was associated with the greatest quality adjusted life expectancy, followed by MR imaging and CNB; life expectancies were 17.409, 17.405, and 17.398 years, respectively. EXB resulted in the greatest lifetime treatment cost ($31,438), followed by MR imaging ($29,072) and CNB ($28,573). Results were robust over a wide range of cancer prevalence, stage distribution, tumor progression rates, and procedure and treatment costs. Incremental cost-effectiveness ratios showed that preoperative testing was cost effective, but the choice between MR imaging and CNB was highly dependent on the accuracy of each test and to patient preferences. CONCLUSION: Preoperative testing of most suspicious breast lesions was cost-effective. More precise estimates of MR imaging and CNB test performance characteristics are needed. Until those are available, patient preferences should inform individual decisions regarding preoperative testing. PMID- 10540639 TI - Ventilation-perfusion lung scintigraphy as a guide for pulmonary angiography in the localization of pulmonary emboli. AB - PURPOSE: To assess the appropriateness of ventilation-perfusion (V-P) scintigraphic abnormalities as a guide to pulmonary angiography for the diagnosis of pulmonary embolism (PE). MATERIALS AND METHODS: V-P scintigrams and pulmonary angiograms of 104 patients with angiographically proved PE were reviewed by two nuclear medicine physicians and two interventional radiologists. For V-P scintigrams, the lung with the larger amount of perfusion abnormality was determined followed by identification of specific lobes. Pulmonary angiograms were similarly evaluated for lateralization and lobar distribution of PE. Conclusions were initially reached independently and subsequently by consensus. RESULTS: Interobserver agreement for lateralization was 88% (kappa = 0.75) for V P scintigraphy and 98% (kappa = 0.96) for pulmonary angiography. In 72 patients, V-P scintigrams predicted unilateral embolus; 64 patients underwent pulmonary angiography of the suspected side. Eight patients underwent contralateral angiography only. Of the 64 patients, 61 (95%) had PE on the predicted side at angiography. V-P scintigrams predicted lobar distribution in 55 patients. Of these, PE was found in the predicted lobe in 42 (76%). CONCLUSION: Localization of perfusion abnormalities at V-P scintigraphy provides useful information for the interventional radiologist and serves as an accurate guide for determining the initial approach for pulmonary angiography. PMID- 10540640 TI - The fissure sign. PMID- 10540641 TI - Cosmetic outcome in patients receiving an interstitial implant as part of breast conservation therapy. AB - PURPOSE: To study factors related to breast cosmetic outcome in patients treated with an interstitial implant as part of breast-conservation therapy. MATERIALS AND METHODS: One hundred fifty-six patients with stage I or II breast carcinoma who received 50 Gy of external-beam irradiation followed by a 20-Gy interstitial boost were examined. The dose homogeneity index (DHI) was calculated for each evaluable implant and was examined in light of other patient-, treatment-, and tumor-related variables previously demonstrated to affect cosmesis. RESULTS: Of the variables examined, both the DHI (P = .021) and the total excision volume (P = .019) were significantly related to cosmetic outcome (excellent vs less than excellent) in a univariate model. In the multivariate analysis, only the total excision volume remained significant (P = .032). The mean total excision volume +/- SD in patients with excellent cosmetic outcome (81.8 cm3 +/- 84.0) was significantly less than that in patients with less than excellent cosmetic outcome (120 cm3 +/- 84). The probability of excellent cosmetic outcome linearly increased with an increase in DHI. The mean DHI was 0.74 +/- 0.12 for the cases with excellent cosmetic outcome and 0.68 +/- 0.10 for those with less than excellent cosmetic outcome. CONCLUSION: To achieve optimal cosmesis, DHI should be maximized. The volume of tissue removed, however, remains the most significant determinant. PMID- 10540642 TI - Resectable esophageal carcinoma: local control with neoadjuvant chemotherapy and radiation therapy. AB - PURPOSE: To evaluate the usefulness of neoadjuvant chemotherapy and radiation therapy before esophagectomy for invasive cancer of the esophagus or gastroesophageal junction (GEJ). MATERIALS AND METHODS: The authors conducted a retrospective analysis of 154 patients who underwent esophagectomy for invasive cancer between September 1, 1991, and December 31, 1995. The end points evaluated were overall, disease-free, local-regional relapse-free, and systemic relapse free survival. RESULTS: Seventy of the 154 patients received neoadjuvant combined modality therapy (CMT) consisting of concurrent cisplatin and fluorouracil administration and accelerated, hyperfractionated radiation therapy. The remaining 84 patients underwent immediate esophagectomy. With a median follow-up of 34.7 months, the 3-year overall, disease-free, and distant metastatic relapse free survival rates were 38.0%, 41.9%, and 56.0%, respectively. Although neoadjuvant therapy did not appear to prevent distant metastases, there was a dramatic effect on local control. After CMT, the 5-year local control rate was 90% compared to 64% after surgery (P < .001). Tumors in the GEJ recurred more frequently (P = .01); however, multivariate analysis showed CMT was the only independent predictor of local control. Postoperative mortality was 15.7% after CMT versus 5.9% without CMT (P = .05). CONCLUSION: Local control of esophageal cancer is excellent following neoadjuvant chemotherapy and radiation therapy. However, the effects of CMT on overall and disease-free survival are less clear due to significant differences between the treatment groups. PMID- 10540643 TI - Posttransplantation lymphoproliferative disorder of the abdomen: CT evaluation in 51 patients. AB - PURPOSE: To study the appearance and distribution of posttransplantation lymphoproliferative disorder (PTLD) at abdominal computed tomography (CT). MATERIALS AND METHODS: The authors retrospectively analyzed pretreatment abdominal CT scans in 51 patients (mean age, 36 years) with PTLD after solid organ transplantation. All diagnoses were proved at either abdominal (n = 26) or extraabdominal (n = 25) pathologic examination. Presence or absence of abdominal involvement, appearance and distribution of disease, and association with abdominal symptoms were all analyzed. RESULTS: CT scans were abnormal in 36 of the 51 patients (71%). Fifteen patients (29%) had no CT, clinical, or pathologic evidence of abdominal involvement. Of the 36 patients with abdominal PTLD at CT, 22% had lymph node enlargement, 28% splenic involvement, and 81% extranodal or extrasplenic involvement. Extranodal abdominal sites included liver (53%), small bowel (25%), kidney (17%), mesentery (14%), adrenal gland (8%), abdominal wall (8%), colon (6%), stomach (3%), and gallbladder (3%). Frequency of abdominal involvement was greater for heart and liver transplant recipients (94%) than for lung and kidney transplant recipients (58%) (P < .01). Seventeen of 36 patients (47%) with abdominal PTLD had no evidence of extraabdominal disease. CONCLUSION: Extranodal involvement is more common than splenic or nodal involvement in patients with abdominal PTLD. The presence of such findings in a patient after transplantation strongly suggests the diagnosis of PTLD and warrants aggressive evaluation. PMID- 10540644 TI - Mucinous versus nonmucinous rectal carcinomas: differentiation with MR imaging. AB - PURPOSE: To determine if quantitative and qualitative magnetic resonance (MR) imaging measures can help differentiation of mucinous from nonmucinous rectal tumors. MATERIALS AND METHODS: In 26 patients with pathologically proved mucinous (n = 9) and nonmucinous (n = 17) rectal tumors, MR imaging was performed with T1 weighted spin-echo (SE) and T2-weighted fast SE sequences in all patients and with a gadolinium-enhanced T1-weighted sequence in 18. With use of the signal intensity (SI) measurements in the tumors and reference tissues, tumor-to-muscle, tumor-to-fat, and tumor-to-urine SI ratios were calculated. In addition, the SI and contrast-enhancement patterns in the tumors were assessed qualitatively by three blinded readers. RESULTS: Mucinous tumors had a much higher SI on the T2 weighted fast SE images. Tumor-to-muscle, tumor-to-fat, and tumor-to-urine SI ratios were significantly higher in the mucinous compared with the nonmucinous tumors (P = .0004, P = .0008, and P = .00002, respectively). Qualitative evaluation of the SI correlated well between readers 1 and 2 (r = 0.93), readers 1 and 3 (r = 0.94), and readers 2 and 3 (r = 0.91). Agreement for the contrast enhancement patterns was 67%, 72%, and 67%, respectively, with most mucinous tumors having predominantly high SI and a peripheral contrast-enhancement pattern. CONCLUSION: Mucinous and nonmucinous rectal tumors can be differentiated with MR imaging because mucinous tumors show high SI on T2-weighted fast SE images. PMID- 10540645 TI - Liver metastases: comparison of current MR techniques and spiral CT during arterial portography for detection in 20 surgically staged cases. AB - PURPOSE: To compare spiral computed tomography during arterial portography (CTAP) with current magnetic resonance (MR) imaging, including hepatic arterial-dominant phase, gadolinium-enhanced, spoiled gradient-echo imaging, for the prospective detection of liver metastases in 20 patients who subsequently underwent surgery to confirm findings. MATERIALS AND METHODS: Twenty patients underwent spiral CTAP and MR imaging within 1 week. Spiral CTAP and MR images were interpreted separately in blinded fashion. All patients subsequently had intraoperative confirmation. Sensitivity, specificity, and positive and negative predictive values were determined for lesion detection and segmental distribution. RESULTS: CTAP and MR images demonstrated, respectively, 54 and 60 true-positive lesions, six and one false-positive lesions, 15 and 22 true-negative (i.e., benign) lesions, and eight and two false-negative lesions. CTAP and MR images demonstrated, respectively, 57 and 62 true-positive segmental involvements, six and one false-positive segmental involvements, 89 and 95 true-negative segmental involvements, and eight and two false-negative segmental involvements. No significant difference in lesion detection was observed. CONCLUSION: Spiral CTAP and MR imaging were approximately equivalent for lesion detection in patients who were evaluated preoperatively for resection of liver metastases. The lower cost and fewer problems with artifacts may suggest that MR imaging is the preferred modality for preoperative assessment of patients for surgical treatment of liver metastases. PMID- 10540646 TI - Liver metastases from melanoma: detection with multiphasic contrast-enhanced CT. AB - PURPOSE: To assess the clinical utility of multiphasic computed tomography (CT) of the liver in patients with metastatic melanoma. MATERIALS AND METHODS: Nonenhanced and biphasic hepatic CT examinations were performed in 28 patients with metastatic melanoma, and liver lesion conspicuity was graded. CT studies in 20 patients met the eligibility criteria, and 13 patients had liver lesions. RESULTS: A total of 57 liver lesions were seen on CT studies: 48 on hepatic arterial phase images, 49 on portal venous phase phase images, and 30 on delayed phase images. Of eight lesions overlooked on portal venous phase images, six were seen on nonenhanced images, and six were seen on arterial phase images. Twenty eight lesions were graded as more conspicuous on portal venous phase images; 10 were graded as more conspicuous on arterial phase images. CONCLUSION: CT images obtained only during the portal venous phase would have resulted in eight (14%) overlooked lesions, which implies that more than one phase is needed for hepatic CT in patients with malignant melanoma. The combination of nonenhanced and portal venous phase CT was as effective as the combination of arterial and portal venous phase CT in these patients. Delayed phase CT did not improve lesion detection either alone or in combination with CT at other phases. PMID- 10540647 TI - Primary melanoma of the esophagus: radiologic and clinical findings in six patients. AB - PURPOSE: To evaluate the clinical and radiologic findings of primary melanoma of the esophagus. MATERIALS AND METHODS: A computer search of pathology, radiology, and cancer registry records from 1973 to 1998 revealed six patients with primary malignant melanoma of the esophagus whose radiographs were available for review. Six esophagograms, three contrast material-enhanced chest computed tomographic (CT) scans, and four chest radiographs were reviewed. Medical records were reviewed for presenting symptoms and clinical course. RESULTS: Six patients (age range, 63-78 years; mean age, 70 years) had histopathologically proved primary malignant melanoma of the esophagus. All patients presented with dysphagia or odynophagia of 6 weeks duration or less. Esophagography and chest CT showed polypoid, nonobstructing esophageal masses, which were mucosal (n = 5) or submucosal (n = 1) and which were located in the middle (n = 3), distal (n = 2), or proximal (n = 1) third of the esophagus. Five patients underwent esophagogastrectomy: Three died a mean of 5 months afterward, two were lost to follow-up, and one was alive 7 months later. CONCLUSION: Primary melanoma of the esophagus is rare. It is usually polypoid, intraluminal, and nonobstructive. As with other esophageal malignancies, the prognosis is dismal despite resection. PMID- 10540648 TI - Diagnosis please. Case 15: congenitally corrected transposition of the great arteries. PMID- 10540649 TI - Abdominal US for diagnosis of pancreatic tumor: prospective cohort analysis. AB - PURPOSE: To elucidate the accuracy of abdominal ultrasonography (US) in the diagnosis of pancreatic tumors. MATERIALS AND METHODS: In all patients referred for pancreatic US during 1988-1990, data on malignant disease and survival were analyzed by using the Swedish Death and Cancer Registries. Nine hundred nineteen patients were entered into the analysis. In 140 of them, a clinical diagnosis of tumor in the pancreatic area was confirmed within 1 year after US. These tumors were primary pancreatic tumors (n = 102), common bile duct and duodenal cancers (n = 17), and metastases in the pancreatic area (n = 21). RESULTS: The sensitivity of US in the detection of all tumors in the pancreatic area was 88.6% (124 of 140 patients), which was similar to that for the detection of exocrine pancreatic cancer, 90% (79 of 88 patients). There were nine false-positive US examinations, for a specificity of 98.8% (770 of 779 patients). Systematic sampling of 94 investigations confirmed an association between US accuracy and presence of clinical symptoms of pancreatic cancer. Significant differences in the sensitivity (P < .05) and accuracy (P < .01) of diagnosis were observed between three experienced investigators. CONCLUSION: Study results support the use of US as a first-line diagnostic examination in patients suspected of having pancreatic tumor. Dependency on the investigator's experience with US mandates continuous evaluation of its performance. PMID- 10540650 TI - Gastric retention of zinc-based pennies: radiographic appearance and hazards. AB - PURPOSE: To determine the radiographic appearance and features of corrosion in U.S. coins exposed to gastric acid. MATERIALS AND METHODS: Six U.S. copper-based pre-1982 pennies, 12 zinc-based post-1982 pennies, a quarter, a nickel, and a dime were exposed to postprandial concentrations of gastric acid (0.15N HCl) for 7 days, and radiographs were obtained daily. Half the zinc-based coins were scraped to disrupt their copper coating. Coins were weighed at the start and completion of the study. RESULTS: Post-1982 zinc-based pennies developed radiolucent corrosive changes within 24 hours. Erosions on the coins became more apparent over time. Frank holes were present on day 2. The weights of these coins decreased 5%-8% during the study. Pre-1982 copper pennies and "silver-colored" coins showed no change on radiographs over 7 days. CONCLUSION: Unexpected radiolucent corrosions may develop in post-1982 zinc alloy pennies when retained in the stomach. Coins have long been considered innocuous foreign bodies in the gastrointestinal tracts of children. However, because of the potential for ulceration and zinc-related morbidity, closer clinical and radiographic observation is warranted. Coins with scalloped edges or holes should be endoscopically removed, as they have likely been retained longer than 1 or 2 days. PMID- 10540651 TI - Cyclic cystography: diagnostic yield in selected pediatric populations. AB - PURPOSE: To test the hypothesis that the diagnostic yield of cyclic cystography is related to the prevalence of vesicoureteral reflux (VUR) in the population being evaluated. MATERIALS AND METHODS: Two groups of children were examined prospectively: 124 with severe urinary tract infection, defined as patient hospitalization or a maximum temperature greater than 39.5 degrees C, and 135 with previously diagnosed VUR. Nuclear cystography was performed in 249 patients, and fluoroscopic cystography was performed in 10. If VUR was not seen during the first cycle of bladder filling and voiding, a second cycle was performed. RESULTS: VUR was present during cycle 1 in 40 (32%) of 124 patients with severe urinary tract infection and 90 (67%) of 135 children in the VUR follow-up group (P < .001). VUR was demonstrated during cycle 2 in seven (9%) of 76 of the severe urinary tract infection group and eight (24%) of 34 of the VUR follow-up group (P = .045). Of 15 patients with VUR during cycle 2, two had grade III VUR and 13 had grade I or II VUR. CONCLUSION: The second cycle of cyclic cystography has a higher diagnostic yield in patients undergoing VUR follow-up than in patients with severe urinary tract infection. The decision to perform a second cycle of bladder filling and voiding should take into account the pretest probability of VUR in the child being examined. PMID- 10540652 TI - Defining and categorizing leukoencephalopathies of unknown origin: MR imaging approach. AB - PURPOSE: To categorize leukoencephalopathies of unknown origin into a few major groups by using magnetic resonance (MR) imaging criteria to facilitate further studies, and to assess the possibility of defining "new" (i.e., until now unknown) disease entities within these major groups. MATERIALS AND METHODS: MR images of 92 patients (55 male, 37 female; mean age, 9.3 years) with a leukoencephalopathy were examined by using a scoring list of 68 items. Seven major categories were defined according to the predominant location of the white matter abnormalities. Statistical analysis was used to assess the validity of these seven categories. RESULTS: Statistical analysis results showed that the seven categories could be well distinguished by either using the defining variables initially accepted as inclusion criteria or selecting a few other variables found to have discriminating value. The additional variables confirmed that the categories are essentially distinct and vary systematically with regard to items other than the inclusion criteria. The existence of two recently defined leukoencephalopathies was confirmed, but no consistent evidence of other new disease entities could be provided. CONCLUSION: Establishing these seven categories helps in the interpretation of individual studies by demonstrating features that the patient has in common with other patients, and it may facilitate further research on homogeneous subgroups of patients and allow pooling of data across multiple centers. PMID- 10540653 TI - Brain tumors: complications of cerebral angiography accompanied by intraarterial chemotherapy. AB - PURPOSE: To evaluate the rate of complications associated with diagnostic cerebral angiography accompanied by intraarterial chemotherapy for the treatment of primary and metastatic brain tumors. MATERIALS AND METHODS: Three hundred ninety-two consecutive transfemoral cerebral angiographic procedures accompanied by intraarterial chemotherapy were performed in 48 patients (28 men, 20 women), and complications were evaluated. RESULTS: The most common local complications were groin hematomas, which occurred in 10 (2.6%) of the 392 procedures and none of which required therapy. Two carotid arterial dissections (0.5%) were reported in two patients who were asymptomatic and did not require further treatment. Both improved at follow-up examinations. Only one patient required surgery for a delayed popliteal embolus. Systemic transient complications occurred five times (1.3%). There were seven (1.8%) transient neurologic events, which were paresis and visual disturbances. Six (1.5%) transient seizure events were recorded. There were no permanent neurologic complications. CONCLUSION: Intraarterial chemotherapy for brain tumors is a safe procedure with a low complication rate. PMID- 10540654 TI - CT assessment of cerebral perfusion: experimental validation and initial clinical experience. AB - PURPOSE: To validate a dynamic single-section computed tomographic (CT) method to measure cerebral blood volume (CBV) and cerebral blood flow (CBF) by using a noncarotid artery as the input and to demonstrate the feasibility of this method in a pilot series of patients. MATERIALS AND METHODS: Twelve dynamic contrast material-enhanced CT studies were performed in beagles. CBV, CBF, and mean transit time (MTT) values were calculated by using an internal carotid artery (ICA) and a noncarotid artery as the input artery to the brain. Patient studies with use of the radial artery as the input were performed (a) repetitively in two patients after subarachnoid hemorrhage, (b) in a patient with a symptomatic ICA occlusion before and after the intravenous injection of 1 g of acetazolamide, and (c) in a patient with a malignant brain tumor. RESULTS: Linear regression analyses revealed highly significant correlations (P < .001) between CBV (r, 0.98; slope, 0.96), CBF (r, 0.89; slope, 0.87), and MTT (r, 0.80; slope, 0.76) values calculated with the ICA and the noncarotid inputs. The CT-derived patient data correlated well with ancillary clinical and neuroradiologic findings. CONCLUSION: Dynamic single-section CT scanning to measure CBV and CBF on the basis of a noncarotid input is a highly accessible and cost-effective blood flow measurement technique. PMID- 10540655 TI - Acute stroke: improved nonenhanced CT detection--benefits of soft-copy interpretation by using variable window width and center level settings. AB - PURPOSE: To assess the use of nonstandard, variable window width and level review settings in computed tomography (CT) without contrast material administration in the detection of acute stroke. MATERIALS AND METHODS: Nonenhanced CT was performed in 21 patients with acute (< 6 hours) middle cerebral arterial stroke and nine control patients. Two blinded neuroradiologists rated all scans for presence of parenchymal hypoattenuation. Images were reviewed at a picture archiving and communication system (PACS) workstation, with standard, locally determined center level and window width settings of 20 and 80 HU and with variable soft-copy settings initially centered at a level of 32 HU with a width of 8 HU. Reviewers altered settings to accentuate gray and white matter contrast. RESULTS: With standard viewing parameters, sensitivity and specificity for stroke detection were 57% and 100%. Sensitivity increased to 71% with variable window width and center level settings, without loss of specificity. Receiver operating characteristic analysis revealed a significant improvement in accuracy with nonstandard, soft-copy review settings (P = .03, one-tailed z test). CONCLUSION: In nonehanced CT of the head, detection of ischemic brain parenchyma is facilitated by soft-copy review with variable window width and center level settings to accentuate the contrast between normal and edematous tissue. PMID- 10540656 TI - Comprehensive MR imaging protocol for stroke management: tissue sodium concentration as a measure of tissue viability in nonhuman primate studies and in clinical studies. AB - PURPOSE: To investigate sodium magnetic resonance (MR) imaging for monitoring tissue viability in stroke. MATERIALS AND METHODS: A comprehensive MR imaging protocol used to measure apparent diffusion coefficient and perfusion parameters was extended to include sodium imaging. Tissue sodium concentration was estimated by using a two-compartment model. This protocol lasted less than 45 minutes. These parameters were followed over the first 6 hours in a nonhuman primate model (n = 2) of acute embolic stroke without or with thrombolytic therapy. This protocol was used in patients in whom acute (< 24 hours, n = 11) or nonacute (> or = 24 hours, n = 31) stroke was ultimately confirmed. RESULTS: The animal model showed abnormal diffusion and perfusion parameters in the lesion immediately after embolization, and these remained abnormal for over 6 hours. Tissue sodium concentration increased with time (5.7 mmol/L/h) unless halted with thrombolytic therapy. Regions with sodium concentrations over 70 mmol/L were histochemically verified as being infarcted. In patients in whom stroke older than 6 hours was clinically confirmed, sodium concentrations over 70 mmol/L were found in the appropriate brain regions. CONCLUSION: Tissue sodium concentration provides a sensitive measure of tissue viability that is complementary to the diagnostic role of diffusion and perfusion imaging for ischemic insult. PMID- 10540657 TI - Asymptomatic carotid arterial disease in young patients following neck radiation therapy for Hodgkin lymphoma. AB - PURPOSE: To determine the prevalence and severity of asymptomatic carotid arterial disease in young patients following neck radiation therapy for Hodgkin lymphoma and to compare the prevalence of carotid arterial disease following radiation therapy alone with that following radiation therapy and chemotherapy. MATERIALS AND METHODS: Forty-two survivors of childhood or early adult Hodgkin lymphoma aged 18-37 years who had undergone radiation therapy more than 5 years earlier underwent carotid arterial ultrasonography. Common carotid intima-media thickness was measured; carotid vessels were assessed for intima-media abnormalities. Results were compared with those from 33 control subjects. RESULTS: Patients had a significantly greater number of abnormal scans than did control subjects (11 [26%] vs one [3%]; P < .01). Ten patients (24%) had intima media abnormalities that did not cause significant stenosis; one patient had diffuse bilateral intima-media thickening (mean, 1.99 mm) with greater than 70% stenosis of both common carotid arteries. Intima-media thickness was significantly greater in patients (0.51 mm) than in control subjects (0.43 mm; P < .005). The number of abnormalities in patients with radiation therapy plus chemotherapy (six [19%] of 31 patients) did not differ significantly from the number in patients with only radiation therapy (five [45%] of 11 patients; P = .12); there was no significant difference between median intima-media thicknesses (0.50 mm vs 0.51 mm, P > .2). CONCLUSION: Asymptomatic carotid arterial disease occurs frequently in young patients following neck radiation therapy for Hodgkin lymphoma. No difference in prevalence was shown between only radiation therapy and radiation therapy plus chemotherapy. PMID- 10540658 TI - T1 and T2 lip cancer: a superselective method of facial arterial infusion therapy -preliminary experience. AB - PURPOSE: To formulate and evaluate a facial arterial infusion chemotherapy for squamous cell lip carcinoma. MATERIALS AND METHODS: The study included six patients (age range, 46-84 years) with squamous cell carcinoma of the lower lip. There were two T1 tumors, three T2 tumors, and one T1-compatible postoperative recurrent tumor. A 4-F, double-lumen balloon catheter was inserted into the external carotid artery through the superficial temporal artery and placed for selective infusion into the tumor-feeding facial artery. Patients received a combination of mitomycin C (4.4 mg/m2 per body surface area) on day 1 and 3.2 mg/m2 of peplomycin sulfate on days 1-7 (22.4 mg/m2 per week), or, when peplomycin sulfate was contraindicated, 16 mg/m2 of cisplatin only on days 1-5 (80 mg/m2 per week). Two to three cycles of chemotherapy were given until tumor disappearance was histologically confirmed. RESULTS: Complete tumor disappearance was achieved in all cases. One patient had a self-limiting asthma attack during peplomycin sulfate treatment, and another had transient partial hair loss. No disfigurement, recurrence, or late complications were observed at a mean follow up of 5.0 years (range, 2.3-11.2 years). CONCLUSION: The described facial arterial infusion chemotherapy appears to be a safe and curative treatment for T1 and T2 squamous cell lip carcinomas. PMID- 10540659 TI - Synthetic dialysis shunts: thrombolysis with the Cragg thrombolytic brush catheter. AB - PURPOSE: To evaluate the effectiveness of the Cragg thrombolytic brush catheter for declotting of synthetic arteriovenous dialysis shunts. MATERIALS AND METHODS: In this randomized controlled trial, 77 patients with synthetic forearm loop shunts that were thrombosed were randomly assigned to undergo pharmacomechanical thrombolysis with a pulsed spray (n = 34) or a thrombolytic brush catheter (n = 43). The following findings were evaluated: declotting time, urokinase dose, procedure time, complications, and shunt patency at the first dialysis session and at 3 months. All data were collected prospectively in an unblinded manner. RESULTS: The total amount of urokinase used, including secondary interventions, was 243,657 IU with the catheter versus 476,563 IU with the pulsed spray (P = .001). At 15 minutes, clot lysis was successful in 66% of the patients with the catheter versus in 19% with the pulsed spray (P = .001). At 30 minutes, clot lysis was successful in 98% with the catheter versus 47% with the pulsed spray (P = .001). Procedure complication rates and patency at 3 months were similar for the catheter and the pulsed-spray groups. CONCLUSION: Use of the Cragg catheter with urokinase offered faster and more complete clot lysis than did use of the pulsed spray with urokinase. The amount of urokinase used with the catheter was half that used with the pulsed spray. Shunt patency at 3 months was similar for the two treatment methods. PMID- 10540660 TI - Thoracic aorta: rapid black-blood MR imaging with half-Fourier rapid acquisition with relaxation enhancement with or without electrocardiographic triggering. AB - PURPOSE: To evaluate and compare findings for thoracic aortic disease with three black-blood magnetic resonance (MR) pulse sequences: half-Fourier rapid acquisition with relaxation enhancement (RARE), with and without electrocardiographic (ECG) triggering, and ECG-triggered turbo spin echo (SE). MATERIALS AND METHODS: Axial black-blood MR images of the chest acquired at 1.5 T with a phased-array coil were obtained in 38 consecutive patients referred for evaluation of thoracic aortic disease. ECG-triggered and nontriggered half Fourier RARE images were compared with T1-weighted ECG-triggered turbo SE images. Two readers independently scored images for each of the following parameters: ghosting artifacts; clarity of the mediastinum, cardiac chambers, and aortic wall; conspicuity of abnormality; intraluminal signal void uniformity; and overall image quality. RESULTS: Both half-Fourier RARE sequences outperformed the turbo SE sequence for all measured parameters. Scores for the ECG-triggered half Fourier RARE sequence were significantly (P < .05) higher than those for the nontriggered version for clarity of the mediastinum and aortic wall, conspicuity of any abnormality other than aortic dissection, and overall image quality. Mean acquisition times for the ECG-triggered (48 seconds) and nontriggered (30 seconds) sequences were significantly shorter than that for the turbo SE sequence (2 minutes 20 seconds). CONCLUSION: Rapid black-blood half-Fourier RARE sequences, with or without ECG triggering, can replace ECG-triggered turbo SE sequences for evaluation of thoracic aortic disease. PMID- 10540661 TI - Intraaortic growth of hydatid cysts causing occlusion of the aorta and of both iliac arteries: case report. AB - A woman who had been operated on previously for a paraspinal hydatid cyst presented with claudication of the lower limbs. Computed tomographic and magnetic resonance images showed multiple cysts in the soft tissues of the back, retroperitoneum, and lumen of the aorta and iliac arteries. Occlusion of the aorta and iliac arteries by recurrent hydatid cysts after previous surgery was confirmed with angiography and subsequent surgical exploration. The authors present the imaging findings of this unusual manifestation of cystic echinococcosis. PMID- 10540662 TI - Can chest CT be used to exclude aortic injury? AB - PURPOSE: To determine whether chest computed tomography (CT) can be used to exclude aortic injury. MATERIALS AND METHODS: Patients in whom there was very high suspicion of traumatic aortic injury were examined with aortography only. Other patients were examined with contrast material-enhanced CT. Follow-up aortography was performed in all patients with moderate to high suspicion of traumatic aortic injury and in all patients with CT scans that were positive for traumatic aortic injury. CT scans were regarded as positive when they showed mediastinal hematoma or direct findings of aortic injury. During a 4 1/2-year period, 1,009 patients (263 female, 746 male; age range, 3-90 years) were evaluated for possible traumatic aortic injury. RESULTS: Of the 207 patients who underwent aortography directly without CT, 10 had traumatic aortic injury. Of the 802 patients who were examined with CT, 382 underwent follow-up aortography. In this group, there were 10 true-positive and no false-negative CT scans. CT had 100% sensitivity and a 100% negative predictive value for the detection of traumatic aortic injury. PMID- 10540663 TI - Acute cervical spine injuries: prospective MR imaging assessment at a level 1 trauma center. AB - PURPOSE: To determine the weighted average sensitivity of magnetic resonance (MR) imaging in the prospective detection of acute neck injury and to compare these findings with those of a comprehensive conventional radiographic assessment. MATERIALS AND METHODS: Conventional radiography and MR imaging were performed in 199 patients presenting to a level 1 trauma center with suspected cervical spine injury. Weighted sensitivities and specificities were calculated, and a weighted average across eight vertebral levels from C1 to T1 was formed. Fourteen parameters indicative of acute injury were tabulated. RESULTS: Fifty-eight patients had 172 acute cervical injuries. MR imaging depicted 136 (79%) acute abnormalities and conventional radiography depicted 39 (23%). For assessment of acute fractures, MR images (weighted average sensitivity, 43%; CI: 21%, 66%) were comparable to conventional radiographs (weighted average sensitivity, 48%; CI: 30%, 65%). MR imaging was superior to conventional radiography in the evaluation of pre- or paravertebral hemorrhage or edema, anterior or posterior longitudinal ligament injury, traumatic disk herniation, cord edema, and cord compression. Cord injuries were associated with cervical spine spondylosis (P < .05), acute fracture (P < .001), and canal stenosis (P < .001). CONCLUSION: MR imaging is more accurate than radiography in the detection of a wide spectrum of neck injuries, and further study is warranted of its potential effect on medical decision making, clinical outcome, and cost-effectiveness. PMID- 10540664 TI - Oblique meniscomeniscal ligament: another potential pitfall for a meniscal tear- anatomic description and appearance at MR imaging in three cases. AB - Three patients with an arthroscopically proved normal variant, the oblique meniscomeniscal ligament, underwent prospective magnetic resonance (MR) imaging of the knee. In the first case, the ligament was misinterpreted as a displaced flap tear of the posterior horn of the lateral meniscus. In the two subsequent cases, the ligament was identified correctly at MR imaging as the oblique meniscomeniscal ligament. PMID- 10540665 TI - Histologic evaluation of platinum coil embolization in an aneurysm model in rabbits. AB - PURPOSE: To characterize the histologic response to platinum coil embolization by using a rabbit aneurysm model. MATERIALS AND METHODS: Saccular aneurysms were created in New Zealand White rabbits by using vessel ligation with intraluminal elastase incubation. Aneurysms were subsequently embolized by using platinum coils. Subjects were sacrificed at various intervals up to 12 weeks following coil embolization. The aneurysm cavities and adjacent vessels were embedded in methylmethacrylate, were sectioned, and were stained for histologic examination. RESULTS: Two weeks following coil implantation, aneurysms were filled predominantly with unorganized thrombus. Six weeks following coil implantation, histologic features included complete filling of the aneurysm lumen with either prominent laminated but unorganized thrombus or areas of unorganized thrombus interspersed among areas of cellular infiltration. At 12 weeks following coil implantation, aneurysms were filled with the loosely packed, disordered cells contained within the extracellular matrix. Fibrosis or smooth muscle cell infiltration was not present in any of the 6- or 12-week samples. CONCLUSION: Platinum coils placed into experimental saccular aneurysms in New Zealand White rabbits failed to elicit a fibrotic response. This model can be used for the testing of biologic modifications of platinum coils aimed at increasing intra aneurysmal fibrosis. PMID- 10540666 TI - Endovascular creation of an in vivo bifurcation aneurysm model in rabbits. AB - PURPOSE: To develop a rabbit model of an intracranial bifurcation aneurysm to test new endovascular therapies. MATERIALS AND METHODS: An experimental aneurysm model was created in rabbits by means of endovascular balloon occlusion of the left common carotid artery, which created an aneurysm at the bifurcation formed by the aortic arch and the brachiocephalic trunk. A total of 18 aneurysms were created. In eight rabbits, the aneurysms were incubated with intraluminal elastase to induce degeneration of the elastic laminae. The animals were followed up with angiography for as long as 3 months. The animals were sacrificed at various times, and histologic evaluation of the aneurysm was performed. RESULTS: Ten aneurysms created without elastase infusion were all very small or completely closed at 1-3 months. Six aneurysms created with elastase infusion had long-term patency (two were patent at 1 month and four, at 3 months). The elastase aneurysms had a mean width of 3 mm (range, 2-3.5 mm) and a mean length of 5 mm (range, 3-7 mm). Histologic evaluation revealed destruction of the normal elastin layers, which allowed the artery to become aneurysmal. CONCLUSION: This aneurysm model re-created the hemodynamic forces and size of human cerebral bifurcation aneurysms and maintained the integrity of the endothelium. The creation of the aneurysms was rapid, reliable, and reproducible. PMID- 10540667 TI - Hemodynamics and wall mechanics after stent placement in swine iliac arteries: comparative results from six stent designs. AB - PURPOSE: To compare the hemodynamics and wall mechanics of swine iliac arteries after placement of six types of stent. MATERIALS AND METHODS: Stents were placed in the iliac artery of 18 pigs (three pigs each underwent placement with one of six types of stent); 16 untreated pigs served as control animals. Iliac arterial hemodynamics and wall mechanics were measured 4 days after placement. RESULTS: Four stents (Palmaz-Schatz, Cordis, Warren, NJ; and Strecker, Cragg, and Symphony, Boston Scientific/Vascular, Natick, Mass) caused decreased pulsatile flow rate in the treated and contralateral iliac arteries; one (Memotherm; Bard, Covington, Ga) caused increased flow pulsatility; and one (Wallstent; Schneider, Plymouth, Minn) had no effect. No compliance mismatching was noted for the Cragg, Symphony, and Memotherm stents, whereas a decrease in compliance was noted for the Palmaz-Schatz, Strecker, and Wallstent designs. The Palmaz-Schatz and Strecker stents caused increased arterial wall rigidity, the Symphony and Wallstent designs had no effect, and the Memotherm and Cragg stents caused decreased wall rigidity. Stents made of stiff metal yielded different early results than did stents made of the less rigid nitinol. CONCLUSION: Soon after implantation, the six stent designs elicited varying changes in blood flow, arterial compliance, and arterial wall mechanics. Contralateral arterial flow also was affected. PMID- 10540668 TI - Reperfused myocardial infarction as seen with use of necrosis-specific versus standard extracellular MR contrast media in rats. AB - PURPOSE: To measure the difference in size of reperfused myocardial infarction with necrosis-specific (bis-gadolinium-mesoporphyrin [hereafter, mesoporphyrin]) and standard extracellular (gadopentetate dimeglumine) magnetic resonance (MR) contrast media. MATERIALS AND METHODS: Echo-planar (for T1 measurement) and spin echo (for infarction size) MR imaging were conducted in 32 rats subjected to reperfused reversible (n = 16) and irreversible (n = 16) myocardial injuries. All animals received gadopentetate dimeglumine 1 hour after reperfusion and underwent imaging. Sixteen rats received mesoporphyrin at 2 hours, the other 16 rats received gadopentetate dimeglumine at 24 hours, and all animals underwent imaging at 24 hours. RESULTS: Mesoporphyrin produced prolonged (22 hours) reduction in T1 in irreversibly, but not in reversibly, injured myocardium. The size of the mesoporphyrin-enhanced region (37% +/- 4 [SEM] of left ventricular surface area) closely correlated with the true infarction size as measured by means of histomorphometry (36% +/- 3, r = 0.90). The size of the gadolinium-enhanced region overestimated (48% +/- 2 and 43% +/- 1 at 1 and 24 hours of reperfusion, respectively) the size of true infarction (36% +/- 3, P < .05, r = 0.02), but it was close to the size of the area at risk (r = 0.93). CONCLUSION: The sizes of hyperenhanced regions displayed by using mesoporphyrin and gadopentetate dimeglumine differed from each other. The difference in size of the hyperenhanced region demarcated by mesoporphyrin and gadopentetate dimeglumine may provide an estimation of potentially salvageable myocardium. PMID- 10540669 TI - Correlation between renal vascular resistance, pulse pressure, and the resistive index in isolated perfused rabbit kidneys. AB - PURPOSE: To assess the effect of acute changes in renal vascular resistance (RVR) and pulse pressure on the resistive index (RI) measured by using Doppler ultrasonography (US). MATERIALS AND METHODS: Rabbit kidneys were perfused by using a pulsatile perfusion system in which RVR, systolic and diastolic pulse pressures, and pulse kinetics were controlled and monitored while simultaneously measuring the RI. RESULTS: When RVR was increased fivefold with phenylephrine hydrochloride, the RI increased only slightly (from 0.45 at baseline up to 0.50). There was a virtually linear relationship between the RI and the pulse pressure index ([systolic pressure-diastolic pressure]/systolic pressure) in the range of 0.30-0.80. The RI was not affected by the pulse rate or fraction of time that systolic pressure was applied during the pulse cycle. CONCLUSION: Contrary to conventional teaching, which is based on theoretic considerations, the RI is not readily affected by acute changes in RVR. This indicates a need to reconsider the conventional explanations used to explain increases in RI that are frequently found in patients with renal disease or ureteral obstruction. PMID- 10540671 TI - Autosomal dominant polycystic kidney disease types 1 and 2: assessment of US sensitivity for diagnosis. AB - PURPOSE: To estimate the sensitivity and specificity of ultrasonography (US) in the diagnosis of autosomal dominant polycystic kidney disease (ADPKD) types 1 and 2, as compared with those of genetic linkage analysis. MATERIALS AND METHODS: A renal US and DNA analysis for ADPKD was performed in 319 patients who were at risk, 161 of whom were younger than 30 years, from 54 families with ADPKD. The sensitivity of US for diagnosis was estimated by comparing the US results with genotypes inferred from linkage studies. RESULTS: The sensitivity of US in individuals younger than 30 years who were at risk was 95% for ADPKD type 1 but only 67% for ADPKD type 2. The sensitivity of US for either ADPKD type 1 or ADPKD type 2 in individuals aged 30 years or older who were at risk was 100%. The overall sensitivity in individuals younger than 30 years was 93%. For both ADPKD types 1 and 2 in all patients, US demonstrated a sensitivity of 97%, a specificity of 100%, and an accuracy of 98%. CONCLUSION: US is the first-line imaging technique that should be used in the diagnosis of ADPKD. The sensitivity in individuals aged 30 years or older is 100%, but if there is a clinical suspicion of ADPKD type 2 in individuals younger than 30 years, linkage analysis should also be considered. PMID- 10540670 TI - Prostate cancer tumor grade differentiation with dynamic contrast-enhanced MR imaging in the rat: comparison of macromolecular and small-molecular contrast media--preliminary experience. AB - PURPOSE: To differentiate prostate cancers of different histopathologic grades with dynamic gadolinium-enhanced magnetic resonance (MR) imaging. Results with a conventional small-molecular contrast medium (CM) were compared to those with a prototypic macromolecular CM. MATERIALS AND METHODS: High- and low-grade tumors, sublines of the Dunning R3327 rat prostate cancer line, were subcutaneously implanted into the flanks of 12 male Copenhagen rats. Dynamic contrast material enhanced MR imaging was performed with small-molecular CM and macromolecular CM at an interval of 1 day. Microvascular permeability, as estimated with the endothelial transfer coefficient, and fractional plasma volume were calculated for each tumor and each CM by means of a two-compartmental, bidirectional kinetic model. RESULTS: Mean endothelial transfer coefficient values for both macromolecular CM and small-molecular CM were significantly different between the two tumor sublines (P = .0004 and P = .01, respectively). For the high- and low grade tumors, no overlap of values was seen with macromolecular CM, but a broad overlap was seen with small-molecular CM despite a significant difference in mean values. CONCLUSION: Dynamic contrast-enhanced MR imaging permits differentiation of histopathologic prostatic tumor types. Quantitative microvascular permeability characteristics estimated from macromolecular CM-enhanced data were significantly superior to those derived from small-molecular CM-enhanced data. PMID- 10540672 TI - Pulmonary nodules resected at video-assisted thoracoscopic surgery: etiology in 426 patients. AB - PURPOSE: To determine the etiology of pulmonary nodules resected at video assisted thoracoscopic surgery (VATS) and establish the probabilities that single or multiple nodules resected at VATS represent malignancy in patients with or patients without known cancer. MATERIALS AND METHODS: Pathology reports from VATS performed between January 1995 and July 1997 were searched for data on gross specimens revealing pulmonary nodules 3 cm or smaller. Findings were correlated with clinical and histologic data. RESULTS: In 254 patients with one nodule resected at VATS, the nodules were malignant in 108 patients with and in 32 patients without known cancer (P < .03). Among 172 patients with multiple nodules resected, at least one nodule was malignant in 85 patients with and in 20 patients without known cancer (P > .05). Nodules larger than 1 cm were more likely to be malignant than were smaller nodules (P < .002). In patients with known malignancy, nodules smaller than 0.5 cm were more likely to be benign, whereas nodules larger than 0.5 cm but smaller than 1 cm were more likely to be malignant (P < .001). CONCLUSION: A single pulmonary nodule resected at VATS was more likely to be malignant in patients with known cancer. Nodules larger than 1 cm but smaller than 3 cm resected at VATS were more likely to be malignant. Nodules smaller than 0.5 cm were more likely to be benign. PMID- 10540673 TI - Cystic fibrosis: usefulness of thoracic CT in the examination of patients before lung transplantation. AB - PURPOSE: To evaluate the usefulness of thoracic computed tomography (CT) in the pre-lung transplantation examination of patients with cystic fibrosis (CF). MATERIALS AND METHODS: Fifty-six patients (age range, 12-42 years) with CF were evaluated for possible lung transplantation from 1991 to 1997. Twenty-six of these patients underwent bilateral lung transplantation, 19 were awaiting transplantation at the time of the study, seven died before transplantation, and four were excluded for psychosocial concerns. Preoperative chest radiographic and CT findings were reviewed and correlated with clinical, operative, and pathology records. RESULTS: In seven patients, discrete, 1-2-cm pulmonary nodules were detected at CT. Five of these patients underwent transplantation; the nodules were found to be mucous impactions. No malignancy was found in any of the patients who underwent transplantation. Pretransplantation sputum cultures grew Aspergillus fumigatus in seven patients, none of whom had radiologic findings suggestive of Aspergillus infection. Radiographic or CT findings were suggestive of mycetoma in five cases, but no such tumors were found at transplantation. The accuracies of chest radiography and CT for the detection of pleural disease in 48 hemithoraces were 81% (n = 39) and 69% (n = 33), respectively. The radiologic findings of pleural thickening did not influence the surgical approach in any patient. CONCLUSION: Thoracic CT has little utility in the routine pre-lung transplantation examination of patients with CF. PMID- 10540675 TI - US probes: risk of cross infection and ways to reduce it--comparison of cleaning methods. AB - After their use at ultrasonography (US) in the intensive therapy unit, probes were used to directly inoculate blood agar plates before and after various cleaning procedures. The uncleaned probes transmitted large numbers of clinically important microbes. Simple cleaning methods were effective in reducing transmission among certain patients: fit patients, double paper wipe; patients at risk of contracting infection, single paper wipe followed by alcohol wipe; patients with a potential source of infection, single paper wipe followed by alcohol wipe. PMID- 10540674 TI - Pulmonary nodules: experimental and clinical studies at low-dose CT. AB - PURPOSE: To compare the number of pulmonary nodules detected at helical low- and standard-dose computed tomography (CT) and to investigate the diagnostic value of low-dose CT with a radiation exposure equivalent to that used at chest radiography. MATERIALS AND METHODS: Two radiologists recorded pulmonary nodules at standard-dose (250 or 100 mA, pitch of 1; 200 mA, pitch of 2) or low-dose CT (50 or 25 mA, pitch of 1 or 2) in five postmortem specimens and 75 patients. Nodules were assessed by size (5 mm or smaller, 6-10 mm, or larger than 10 mm) and by diagnostic confidence ("definite nodule," "definite lesion, not classic nodule," or "questionable lesion, possibly representing a vessel") with the Wilcoxon signed rank test. Artifacts depicted at low-dose CT were recorded. RESULTS: There were no statistically significant differences in the number of nodules detected at standard- or low-dose CT except in nodules 5 mm or smaller that were assessed as definite nodules at standard- or low-dose CT (25 mA, pitch of 2) (472 vs 397, P < .05). Artifacts that possibly interfered with nodule detection were observed exclusively at CT with 25 mA and a pitch of 2. CONCLUSION: Pulmonary nodules were detected reliably at CT with 50 mA and pitch of 2 or with 25 mA and a pitch of 1. However, further reduction of the dose to that used at chest radiography was associated with a significant decrease in the number of nodules 5 mm or smaller that were detected, possibly due to artifacts. PMID- 10540676 TI - Hemodialysis graft: use as access for upper and lower extremity arteriography and interventional procedures--initial experience. AB - Functioning hemodialysis grafts were used as access sites for peripheral vascular arteriography and interventional procedures. In 11 patients with end-stage renal disease and ischemia, upper extremity (n = 8) or lower extremity (n = 3) arteriography was performed successfully. Angioplasty and other interventional procedures were performed via the same route in two of the patients. No bleeding complications occurred, and all patients were ambulatory immediately after the procedure. PMID- 10540677 TI - Tunneled jugular small-bore central catheters as an alternative to peripherally inserted central catheters for intermediate-term venous access in patients with hemodialysis and chronic renal insufficiency. AB - In 34 patients with chronic renal insufficiency or failure, 43 small-bore central catheters were placed via the internal or external jugular veins: right internal jugular vein, 28; left internal jugular vein, 14; right external jugular vein, one. Central venous access was achieved in all patients (mean catheter dwell time, 28 days; range, 3-99 days), with two minor complications (arterial puncture and catheter damage during suturing). Tunneled jugular small-bore central catheters are a vein-preserving alternative to peripherally inserted central catheters in this population. PMID- 10540678 TI - Perils of PACS. PMID- 10540679 TI - Perils of PACS. PMID- 10540680 TI - Pneumococcal vaccine. PMID- 10540681 TI - Launch of a rural advanced maternity care curriculum. PMID- 10540682 TI - Getting the message across. PMID- 10540683 TI - Why was the study not included? PMID- 10540684 TI - Publish critical reviews only, please. PMID- 10540685 TI - John's story. PMID- 10540686 TI - Patients' wishes are very important. PMID- 10540687 TI - Martyrdom by toads' tongues. Early Canadian doctors and their victims. PMID- 10540688 TI - Dextromethorphan. Extrapolation of findings from reproductive studies in animals to humans. AB - QUESTION: One of my patients, who is now 8 weeks pregnant, just read in the newspaper that dextromethorphan (DM), an antitussive found in a variety of cough medicines, caused birth defects in chicken embryos. The author of the study stated that even one dose could be dangerous and that he would never allow his wife to use this drug if she were pregnant. My patient was understandably very concerned because last week she was suffering from a nasty cough and had been advised by her pharmacist to use a cough mixture containing DM, which she subsequently took for several days. ANSWER: You may reassure your patient that she did not put her baby at risk by using this substance. Dextromethorphan has been on the market for many years and has never been implicated as a human teratogen. Furthermore, chick embryos are not a good model for predicting teratogenic potential in humans and, consequently, were abandoned as such more than 30 years ago. PMID- 10540689 TI - Dermacase. Sebaceous adenomas (angiofibromas). PMID- 10540690 TI - Radiology rounds. Osteopoikilosis. PMID- 10540691 TI - Domestic violence is the leading cause of injury to women aged 15 to 44. PMID- 10540692 TI - Functional (non-ulcer) dyspepsia and Helicobacter pylori infection. To treat or not to treat? PMID- 10540694 TI - Treatment of obesity. PMID- 10540693 TI - Orlistat. No hurry.... AB - Treatments for obesity are disappointing. None has yet shown an effect on morbidity and mortality. Nondrug treatments are poorly assessed. Stable long-term weight loss necessitates long-term management. Orlistat (Xenical, Hoffman-La Roche), a gastrointestinal lipase inhibitor, is indicated, in combination with a low-calorie diet, for management of obesity. The assessment file is bulky and methodologically sound, at least in terms of the weight loss end point. During medium-term trials (12 to 24 months), orlistat administered at a dose of 120 mg three times daily and combined with dietary intervention had a moderate positive effect on body weight (-3.5 kg on average). No longer-term trials have been done. It is unknown whether this drug affects morbidity and mortality linked to obesity. In clinical trials, patients treated with orlistat had an increased frequency of breast cancer. This potential risk is currently being assessed in a specific trial. Gastrointestinal adverse effects are frequent. Treatment is costly. PMID- 10540695 TI - Conservative management of spontaneous abortions. Women's experiences. AB - OBJECTIVE: To describe women's experiences with expectant management of spontaneous abortions. DESIGN: Descriptive survey using questionnaires with fixed choice and open-ended questions. The latter were analyzed for themes, using qualitative methods. SETTING: Urban and suburban private primary care family practices. PARTICIPANTS: A convenience sample of family practice patients (59 of 80 eligible) pregnant for less than 12 weeks who had spontaneous abortions without surgery. Response rate was 84.7%; 50 questionnaires were received from the 59 women. METHOD: Women were asked about their physical experiences, including amount of pain and bleeding; emotional effects; their satisfaction with medical care; and their suggestions for improving care. MAIN FINDINGS: The mean worst pain experienced during a spontaneous abortion on an 11-point scale was 5.9. Bleeding varied, but was often very heavy. Satisfaction rate was 92.9% with family physician care and 84.6% with hospital care. Women described the emotional effect of "natural" spontaneous abortions and made recommendations for improving care. CONCLUSIONS: A better understanding of the physical and emotional experiences of the women in this study might help physicians better prepare and support patients coping with expectant management of spontaneous abortions. PMID- 10540696 TI - Reducing surgery in management of spontaneous abortions. Family physicians can make a difference. AB - OBJECTIVE: To test the effectiveness of physician and patient education in reducing the rate of surgery in management of spontaneous abortions in family practice. DESIGN: A before-after intervention trial. SETTING: Urban and suburban family doctors' practices in greater Vancouver, BC. PARTICIPANTS: Family practice patients (56 physicians contributed 417 patients) who had spontaneous abortions between June 1997 and August 1998. INTERVENTIONS: Seminars for doctors and educational pamphlets for patients. MAIN OUTCOME MEASURES: Rate of surgeries, and rates of referrals and complications. RESULTS: In the 2 years before the intervention, the rate of surgery was 45.8% (n = 299); after, it was 32.2% (n = 118). No transfusions were required. Before the intervention, 17% of women had hemorrhages; after, 13%. Rates of infection were 3.7% and 0.8%, respectively. Rates of referral to gynecologists were 54.0% and 40.2%, respectively. CONCLUSIONS: Patients of family doctors who attended seminars and agreed to join the study had significantly reduced rates of surgery after spontaneous abortions. Rates of referral for these patients were also lower, and there was no increase in complications. PMID- 10540697 TI - How accurately do we estimate patients' weight in emergency departments? AB - OBJECTIVE: To assess the accuracy of estimates of patients' weight made by physicians, nurses, and patients themselves in emergency departments. DESIGN: Observational prospective study. SETTING: Tertiary referral centre in Vancouver, BC. PARTICIPANTS: Eleven attending physicians, 26 nurses, and a convenience sample of 117 patients. INTERVENTIONS: Patients themselves, attending physicians, and nurses independently estimated the weight of 117 patients. An investigator weighed each patient. MAIN OUTCOME MEASURES: Mean error was determined by subtracting actual weight from estimated weight and dividing by actual weight; 95% confidence intervals (CI) were calculated. RESULTS: Mean error in estimates was 3.1% (95% CI 2.7 to 3.5) for patients, 8.4% for nurses (CI 7.6 to 9.2), and 8.1% (CI 7.1 to 9.1) for physicians. Weight was estimated within 5% of actual weight by 32% of nurses, 39% of physicians, and 82% of patients. Weight was estimated within 10% of actual weight by 66% of nurses, 66% of physicians, and 97% of patients. Estimates out by more than 15% were made by 11% of nurses, 16% of physicians, and 1% of patients. CONCLUSIONS: Patient estimates were most accurate. Physician and nurse estimates were unreliable. PMID- 10540699 TI - Diagnosing and treating asymptomatic tuberculosis infection. AB - OBJECTIVE: To summarize relevant parts of the guidelines recommended by the Canadian and American Thoracic Societies for diagnosis and management of asymptomatic tuberculosis (TB) infection. QUALITY OF EVIDENCE: The latest guidelines published by the Canadian and American Thoracic Societies were reviewed. Unfortunately, neither of these guidelines state explicitly how recommendations were derived. The references accompanying each set of guidelines, however, suggest that they were developed by extensive literature review of the subject and consensus among expert panels. MAIN MESSAGE: Only higher-risk patients should receive a TB screening test (Mantoux test) to minimize the possibility of false-positive test results. The cutoff points for positive tests vary to reflect the pretest likelihood of TB infection. An induration 5 mm or greater is considered positive in patients at highest risk of TB infection, that is, HIV-infected patients, close contacts of active TB cases, and patients with chest x-ray abnormalities suggestive of previous untreated TB. All other patients are considered positive if they have induration greater than 10 mm according to the Canadian guideline. A 15-mm cutoff point, however, is used for patients without risk factors in the American guideline. All patients with positive Mantoux test results should be considered infected with TB. Infected patients should be offered 6 to 12 months of isoniazid prophylaxis if they have HIV infection, if they have medical conditions that increase the risk of TB activation, or if they are younger than 35 years. CONCLUSIONS: Prophylactic treatment of infected individuals effectively prevents the spread of TB infection. Family physicians, who most often see patients in the asymptomatic stage of TB infection, are uniquely situated to prevent secondary cases of TB by offering appropriate patients prophylactic treatment. Patients should be counseled about the risk and benefit of prophylactic treatment so they give informed consent for it. PMID- 10540698 TI - Clinical effectiveness of pneumococcal vaccine. Meta-analysis. AB - OBJECTIVE: To determine the clinical effectiveness of pneumococcal vaccine. DATA SOURCES: Computerized searches of MEDLINE, EMBASE, and SCISEARCH databases were performed, reference lists of retrieved articles were reviewed, and first authors of published studies were contacted. STUDY SELECTION: Studies of use of pneumococcal vaccines in adults were included if the study design was a randomized or quasi-randomized controlled trial and at least one of the following clinical outcomes was reported: vaccine-type systemic pneumococcal infection, systemic pneumococcal infection, vaccine-type pneumococcal pneumonia, pneumococcal pneumonia, non-vaccine-type pneumococcal pneumonia. SYNTHESIS: Study quality was assessed and descriptive information concerning the study populations, interventions, and outcome measurements was extracted for 13 trials involving more than 65,000 patients. Estimates of vaccine efficacy, based on a meta-analysis of randomized and quasi-randomized trials, were determined for clinical outcomes. CONCLUSIONS: Vaccination with pneumococcal polysaccharide vaccine can be expected to reduce the risk of systemic infection due to pneumococcal types included in the vaccine by 83% and systemic infection due to all pneumococci by 73%. We found no evidence that the vaccine was less efficacious for the elderly, institutionalized people, or those with chronic disease. PMID- 10540700 TI - How advance directives affect hospital resource use. Systematic review of the literature. AB - OBJECTIVE: To assess whether advance directives influence resource use by hospitalized patients. DATA SOURCES: A systematic search of computerized medical databases, reference lists from relevant articles, and personal files was conducted to identify studies examining the association between advance directives and resource use. STUDY SELECTION: Primary studies assessing the effect of advance directives on hospital resource use were selected if they had a clear quantitative measure of hospital resource use, hospitalized patients as a study population, a control group for comparison, and a description of the advance directive being studied. Data on the following topics were abstracted from studies meeting inclusion criteria: study methods and design, resource use, source of financial data, description of advance directive, population size and composition, length of assessment. SYNTHESIS: Six studies met inclusion criteria. Three retrospective studies showed significant reductions in resource use associated with documentation of advance directives while three prospective studies (two randomized, one not randomized) showed no association between advance directives and reduced resource use. Studies were limited to narrowly defined patient populations in US tertiary care hospitals. CONCLUSIONS: Little evidence supports the hypothesis that advance directives reduce resource use by hospitalized patients. Some retrospective studies have shown savings, but their conclusions are weakened by shortcomings in study design. Prospective trials, which have better experimental methods, have demonstrated no evidence of cost savings with the use of advance directives. PMID- 10540701 TI - Joint position paper on training for rural family practitioners in advanced maternity skills and cesarean section. College of Family Physicians of Canada, Society of Rural Physicians of Canada, Society of Obstetricians and Gynaecologists of Canada. PMID- 10540702 TI - Building or renovating your clinic. PMID- 10540703 TI - [Electroconductive lithotripsy: clinical results of the Sonolith sigma]. AB - The electroconductive lithotripter (ECL) is a new concept for shockwave generation in which a highly conductive solution channels the discharge between the anode and cathode. Out of 152 patients treated, complete follow up data were available on 151 patients. The average number of shocks per treatment was 2,138. At 3 months the overall stone-free rate was 73.5%. Success rate, defined as stone free or asymptomatic residual fragments measuring 4 mm or less, was 84.9% for renal and 94.9% for ureteral calculi. The overall success rate for all calculi was 89%. PMID- 10540705 TI - [Clinical study of transurethral electrovaporization of the prostate for benign prostatic hyperplasia]. AB - We report our clinical experience of transurethral electrovaporization of the prostate (TUVP) for benign prostatic hyperplasia (BPH) using VaporTrode developed by Circon ACMI. From April to November 1995, we treated 22 patients with symptomatic BPH with TUVP. The mean I-PSS decreased significantly, from 21.8 at baseline to 8.1, 4.5, 4.3 and 5.4 at 1, 3, 6 and 12 months after operation, respectively. The mean QOL index also decreased significantly, from 5.2 to 1.5, 1.0, 1.3 and 1.3. The mean peak flow rate increased significantly, from 9.0 preoperatively to 17.4, 17.7, 20.8 and 16.5 ml/sec at 1, 3, 6 and 12 months after TUVP, respectively. The mean prostate volume decreased significantly, from 41.5 to 22.9, 18.6, 18.8 and 19.9 ml. The mean residual urine decreased significantly, from 90.0 to 17.6, 20.6, 24.1 and 9.4 ml. As for overall efficacy, the rate of excellent and good cases at 1, 3, 6 and 12 months was 77.3, 95.5, 95.0 and 84.2%, respectively. No serious complications were observed. Our clinical results suggest that TUVP using VaporTrode has several potential advantages including good efficacy, minimal morbidity and lower cost compared with other less invasive procedures, and may become the useful way of surgical treatment for BPH. PMID- 10540704 TI - [Intra-arterial infusion chemotherapy for superficial bladder cancer]. AB - Intra-arterial infusion chemotherapy (AI) is widely used in the treatment of invasive bladder cancer, but few studies have been reported on its efficacy for superficial bladder cancer. We retrospectively examined the anti-tumor effect and prophylactic effect of AI in 18 cases which were either a case with multiple or extensive tumors which could not be controlled completely by transurethral resection (TUR), a case with grade 3 tumors or a recurrent case after TUR and/or intravesical chemotherapy. Fifty mg of adriamycin and 100 mg of cisplatin were administered into bilateral internal iliac arterys. This treatment was repeated 1 3 times every three weeks. Concerning the anti-tumor effect, 8 showed a complete response, 5 showed partial response and 5 showed no change; the overall response rate (CR + PR) was 72%. Concerning the prophylactic effect, 1-, 2- and 3-year recurrence-free rates were 58.8%, 41.1% and 32.9%, respectively. This study demonstrated the efficacy of AI in the anti-tumor treatment, but not in the prophylactic treatment of superficial bladder cancer. PMID- 10540706 TI - [A case of extragonadal germ cell tumor with inferior vena caval tumor thrombus]. AB - We report a case of retroperitoneal extragonadal germ cell tumor with tumor thrombus in the inferior vena cava. The patient referred to our hospital with lumbago. Computed tomography (CT) showed a bulky mass in the retroperitoneum. The levels of alpha-fetoprotein (AFP) and beta-human chorionic gonadotropin (beta HCG) in the serum were elevated. Histological examinations indicated embryonal cell carcinoma. Bilateral testicles did not contain any palpable mass upon careful palpation. No tumor mass was detected in the bilateral testicles on ultrasonography. Clinically, the diagnosis was a retroperitoneal extragonadal germ cell tumor associated with para-aortic lymph-node involvement. After the combination chemotherapy (BEP 1 course and EP 3 courses), the tumor mass was reduced in size and the tumor marker was normalized. Retroperitoneal lymph node dissection (RPLND) was performed and tumor thrombus in the inferior vena cava was resected. There was no involvement of the viable cells in the resected tumor. The patient has been in good condition with no evidence of disease. PMID- 10540707 TI - [Complete remission of lung and hepatic metastases from renal cell carcinoma by interferon alpha-2b therapy: a case report]. AB - A right renal tumor was found in a 74-year-old man with multiple metastases to the lungs and liver. Tumor thrombus extending into the inferior vena cava and a right spermatic varicocele were also noted at the first visit. Interferon alpha 2b and interferon gamma were administered for treatment. Partial remission of lung metastases, complete remission of hepatic metastases, and disappearance of the varicocele occurred after 4, 6 and 8 weeks, respectively. Then the primary right renal tumor was resected. Although only interferon alpha-2b was continued twice weekly by self-injection, complete remission of the lung metastases was obtained 13 weeks after the initiation of therapy. No evidence of recurrence or new metastasis has been found after 18 months. These results indicate that even advanced renal cell carcinoma may show a rapid response to interferon alpha. Interferon alpha is worth trying for metastatic renal cell carcinoma and should be continued for at least a few months. PMID- 10540708 TI - [A case of pulmonary metastasis from renal cell carcinoma with complete response to interferon-alpha and tegafur/uracil (UFT) but possibly UFT-induced liver dysfunction and leukoencephalopathy-like symptoms]. AB - A 61-year-old man presented with gross hematuria. He underwent left radical nephrectomy under a diagnosis of left renal cell carcinoma without distant metastasis, but bilateral multiple pulmonary metastases appeared 2.5 months after the operation. Though the metastases responded well to combination therapy of interferon-alpha and a 1:4 mixture of tegafur and uracil (UFT), the side effects of liver dysfunction and leukoencephalopathy-like symptoms due to UFT appeared 7 months after the beginning of the chemotherapy. These side effects were improved after the cessation of UFT administration. PMID- 10540709 TI - [A case of emphysematous pyelonephritis with emphysematous cystitis]. AB - A 74-year-old woman with diabetes mellitus had a high fever, and was treated with antibiotics and insulin in another hospital. She was referred to our department, because CT scan showed the right hydronephrosis and the abnormal gas shadow in the right renal calyces. Ureteral catheterization was performed on the right side and cloudy urine was drained. Urine culture yielded E. coli. Since submucosal emphysematous changes were demonstrated in the bladder mucosa by cystoscopy, she was diagnosed with emphysematous pyelonephritis with emphysematous cystitis associated with diabetes mellitus. Administration of antibiotics and insulin and the ureteral catheter drainage improved her condition immediately. Abnormal gas shadow on CT scan and submucosal emphysema on cystoscopy disappeared. We reviewed 110 cases of emphysematous pyelonephritis and 23 cases of emphysematous cystitis including our case in Japan, and report their clinical characteristics. PMID- 10540711 TI - [Acute renal failure due to over-infusion in a child with bilateral obstruction of the pyeloureteral junction]. AB - A 2-year-old boy was hospitalized with the chief complaint of oliguria and dyspnea. Bilateral hydronephrosis and obstruction of the pyeloureteral junction were detected by ultrasonography. Pulmonary edema was also found on chest radiographs. The clinical diagnosis was acute post renal failure due to bilateral pyeloureteral obstruction and pulmonary edema due to overtransfusion. After we performed bilateral percutaneous nephrostomy, the patient recovered from renal failure and pulmonary edema. Both nephrostomies were removed after we confirmed a non-obstructing pattern using the Whitaker test. PMID- 10540710 TI - [A case of pyonephrosis caused by ureteral stones with elevated serum levels of CA19-9]. AB - A case of pyonephrosis with high levels of serum CA19-9 antigen is reported. A 71 year-old woman was admitted with right flank pain. Computed tomography and ultrasonography showed severe hydronephrosis and hydroureter due to a right ureter stone. Laboratory data revealed a high level of serum CA19-9. However, no tumor was found in the pancreas, gallbladder, liver, gastrointenstinal tract or genitourinary tract. Drip infusion pyelography showed a non-functioning pattern of right kidney. Therefore, right nephroureterectomy was performed for right pyonephrosis. Histological examination revealed chronic inflammation. Malignant cells were not seen in the resected specimen. The serum CA19-9 levels before and after operation were 102.9 U/ml and 24 U/ml, respectively, being normal after the operation. Immunohistochemical examination revealed the presence of CA19-9 antigen in the urethelium, indicating its expression in the specimen. To our knowledge this might be the first case of pyonephrosis associated with high levels of serum CA19-9 antigen. PMID- 10540712 TI - A case of adenocarcinoma with clear cell carcinoma of the bladder. AB - A case of adenocarcinoma with clear cell carcinoma of the bladder in a 65-year old male is reported. Our patient had a walnut-sized nodular tumor located on the anterior wall of the bladder. The patient underwent radical cystoprostatectomy with urethral hemi-Koch pouch. Histopathological examination revealed a lesion composed of poorly-differentiated adenocarcinoma and clear cell carcinoma with diffuse sheet patterns of cells with abundant, clear cytoplasm. The patient died of general metastasis 18 months after operation. To our knowledge this is the first case of adenocarcinoma with clear cell carcinoma arising from the anterior wall of the bladder in a male. PMID- 10540713 TI - [Small cell neuroendocrine carcinoma of the urinary bladder: a case report]. AB - A 60-year-old male was referred to our hospital with a complaint of asymptomatic gross hematuria. Cystoscopic examination revealed a non-papillary broad-based tumor on the posterior wall of the urinary bladder. Computed tomography revealed no evidence of metastases. Transurethral resection of bladder tumor (TUR-BT) was performed and muscle invasion was detected by histological examination of the specimen. Total cystectomy and ileal conduit formation were performed at the preoperative diagnosis of T2-3N0M0. Hematoxylin-eosin staining of the specimen revealed small cancer cells with hyperchromatic nucleus and scanty cytoplasm growing in the muscle layer of the urinary bladder and in the left obturator lymph nodes. Immunohistochemistry for neurospecific enolase showed diffuse staining in the cytoplasm of cancer cells, and ultrastructural study showed dense core granules. From these findings, the patient was diagnosed with small cell neuroendocrine carcinoma of the urinary bladder at the stage of pT3bpN1M0. Three courses of adjuvant chemotherapies with cis-platinum (CDDP) and etoposide were administered. The patient is still alive with no evidence of any recurrence at 22 months after the operation. This case suggests that treatment with combined total cystectomy and adjuvant CDDP and etoposide chemotherapies is effective against neuroendocrine carcinoma of the urinary bladder with regional lymph node metastases. PMID- 10540714 TI - [A case of malignant lymphoma of the testis: characterization with ultrasonography and magnetic resonance imaging]. AB - We report a case of stage IE non-Hodgkin's lymphoma of the testis including the sonographic and magnetic resonance imaging (MRI) findings. A 68-year-old male noticed a painless swelling of his right scrotal contents. The serum alpha fetoprotein and beta-HCG levels were within the normal ranges. Scrotal ultra sonography revealed a hypoechoic lesion in the right testis. This tumor showed isointensity on T1-weighted imaging and also showed low intensity on T2-weighted imaging. A high orchiectomy was performed and the histological diagnosis was non Hodgkin's lymphoma. Adjuvant chemotherapy (cyclophosphamide, doxorubicin, vincristine and predonisolone) was administered. At 6 months postoperatively, this patient shows no signs of recurrence. PMID- 10540715 TI - [Metastatic tumor of the epididymis from pancreatic carcinoma: a case report]. AB - A 58-year-old male who complained of painful left scrotal swelling consulted a local clinic in August 1998. Because his symptoms did not improve after antibiotic therapy, he was transferred and admitted to Jyouhoku City Hospital on September 14, 1998. Pelvic computed tomography (CT) was performed, and revealed left epididymitis. However, antibiotic treatment did not improve the condition. Then, because carcinoma of the epididymis was suspected, left inguinal orchiectomy was performed. We found a tumor in the spermatic cord and another tumor in the epididymis. The pathological diagnosis was adenocarcinoma, and metastatic carcinoma from the digestive tract was suspected. Therefore, examinations were performed to detect the primary cancer. CT and magnetic resonance imaging (MRI) demonstrated an invasive irregular tumor from the pancreas to the left kidney. Irregular mucosa was observed by gastrointestinal fiberscopy. A biopsy was performed and the pathological diagnosis was adenocarcinoma. Based on these findings, the patient was diagnosed as having a metastatic tumor of the epididymis and spermatic cord caused by pancreatic carcinoma. This is the 3rd case of adenocarcinoma of the pancreas that presented as an epididymal nodule, and this is the 12th case of adenocarcinoma of the pancreas that presented as a spermatic cord nodule. PMID- 10540716 TI - [Prenatal diagnosis: an attempt at evaluation of effectiveness and risk based on the experience of one center]. AB - We present the results of prenatal diagnosis in 2241 women carried out in one centre in the period 1985-1994. Indications were cytogenetic in 84% of the cases, of those in 77% it was maternal age 35 years and over. The second most frequent indication was open neural tube defect in a previously born child (7.5%). Abnormal results of prenatal tests in whole material were obtained in 60 cases (2.4%); in 47 cases this was chromosomal aberration. Abnormal result of prenatal test did not necessarily mean selective termination of pregnancy. In 17.5% of chromosomal fetal aberrations pregnancy was continued (it concerned mostly aberrations involving sex chromosomes). The risk of prenatal diagnosis (miscarriage due to the procedure) according to our estimation was between 0.3 and 0.6% of the tested pregnancies. Sociological analysis of the tested group showed clearly that women with better education (secondary and higher level) in Poland have much better access to prenatal diagnosis. Most of the tested woman (72%) considered a prenatal test a sine qua non condition of their procreation. PMID- 10540717 TI - [The effect of astrocytoma cells on secretion of cytokines by autologous lymphocytes]. AB - Immunotherapy is one of the experimental methods used in the treatment of brain tumours. The development of astrocytomas is accompanied by decreased activity of lymphocytes, namely inhibition of mitogen-induced proliferation and decreased cytokine secretion, which is an adverse event in the course of disease. In the present study, astrocytoma cells obtained from human tumours and cocultured with autologous lymphocytes inhibited cytokine secretion by those lymphocytes. PMID- 10540719 TI - [The difference among patients with acute ischemia stroke and with reversed flow in the ophthalmic artery and the ones with a correct direction of the flow]. AB - METHOD: Fifty-four patients with acute ischaemic stroke (AIS) and with reversed flow direction in the ophthalmic artery (OA) were compared prospectively with patients with normal flow in that artery. Age, sex, presence of atrial fibrillation (AF) and ischaemic heart disease, the course in the acute period within 30 days after stroke onset were analysed. On the first day the following examinations were performed in all patients: routine laboratory investigations, X ray, ECG and Doppler and USG examinations of the extracranial arteries. In some cases CT of the head was obtained. The condition of the patients was assessed by Rankin scale on the first and 30-th days after stroke onset. Improvement was accepted if on the 30-th day the score was 1 point better than on the first assessment. RESULTS: The mean age in the group with reversed blood flow in OA was 60.7 years, and in the group with normal flow it was 69 years (p < 0.0001). The male/female ratio was statistically significantly different in these groups (p = 0.01): in the younger group with reversed flow males accounted for 76% and in the older group with normal flow--52.5%. AF in the younger group was present in a low proportion of cases (3.7%) but in the older group it was 21.3%, the difference was statistically significant (p = 0.005). No significant difference was found in the presence of ischaemic heart disease. In the patients scoring 4 and 5 in Rankin scale in the first assessment no worsening was noted in the group with reversed blood flow, while in the other group 24% of the patients were worse or died (statistically significant difference, p = 0.004). The course of mild and medium severe stroke within 30 days was similar in both groups. CONCLUSIONS: Patients with AJS and reversed flow in OA-differed from those with normal flow in OA, they were younger, more often were males. AF was present in isolated cases and the course of major stroke was milder. PMID- 10540718 TI - [Fibrinogen and lipids: associated risk factors for ischemic cerebral stroke]. AB - The study included 55 patients (18 females, 37 males); aged 32-75 yr. who divided into three groups according to the severity of clinical picture: 12 people with reversible ischaemic stroke (RIS), 20 with progressive ischaemic stroke (PIS), 23 with complete stroke (CS). Levels of total cholesterol, high density lipoproteins (HDL), apolipoproteins A1 and B (ApoA1 and ApoB), fibrinogen (Fb) and Lp (a) were measured. Lipid factor of atherosclerosis (ATHi) was quantified. Qualitative evaluation of lipids contents in cerebrospinal fluid (CSR) was performed. Distribution of cholesterol--containing lipids among the fractions, despite low values, had clearly atherogenic profile. 12% patients with irreversible ischaemic stroke, 16% with progressive ischaemic stroke and 85% with complete stroke had Fb level above 4 g/l. Lp (a) levels in all cases were significantly higher in the cells isolated from CSF. The severity of the stroke correlated with increasing levels of lipids in the cells isolated from SF. There was correlation between LDL cholesterol and content of lipids in the cells from CSF. PMID- 10540720 TI - [Epidemiology of multiple sclerosis in the region of Szczecin: prevalence and incidence 1993-1995]. AB - In the Province of Szczecin in the period 1993-1995 the incidence was 2.16 and the prevalence 55.32/100,000 on December 31 1995. Incidence and prevalence were higher in women (the difference was higher in Szczecin City) what was not connected with a significantly greater number of women in the population of the province. The study confirmed the presence in the southern part of the province of a focus with prevalence 110.54/100,000. The mean age at disease onset was 37.05 +/- 8.91. In women it was significantly higher than in men and it was nearly 10 years longer in relation to the mean age at onset in the years 1960 1984. PMID- 10540721 TI - [Surgical treatment of GnRH secretory hypothalamic hamartomas causing precocious puberty]. AB - Hypothalamic hamartomas (HHs) are benign lesions often associated with central precocious puberty. Resection of HHs has been recommended as a treatment option for selected cases, however recent reports stressed the role of effective medical management with a long-acting GnRH agonist. This paper describes incomplete response to the initial GnRH therapy and results of total resection of HHs. Five children two boys and three girls with physical signs of puberty at a mean age of 20.2 months (range 5-36 months) have been treated at our institution. All children had a pedunculated mass below the tuber cinereum. Two children were initially treated with GnRH agonist and had received follow-up care for 11 and 12 months respectively. These patients had incomplete regression of endocrinological disturbances to prepubertal level. Both patients were subsequently operated on. All five children underwent total surgical removal of HHs. The hamartomas were excised through pterional approach. Postoperatively two children showed transient third nerve palsy and one diabetes incipidus. There was no permanent disability due to surgical intervention. The clinical and biochemical symptoms and signs of precocious puberty completely regressed postoperatively including two cases treated initially with GnRH analog. The children have been followed for 1 to 6 years and no recurrence of puberty was observed. Surgical excision of pedunculated HHs is still a valuable option in the treatment of precocious puberty in small children. This alternative should be considered if the initial GnRH therapy failed to suppress++ puberty and reduce bone age advancement. PMID- 10540722 TI - [Cases of cavernous hemangioma discovered in patients with post traumatic intracerebral hematoma]. AB - Cavernous haemangioma has various forms. The cases with spontaneous intracerebral haemorrhage are most common, but post-traumatic intracerebral haematoma was not reported. The aim of this report is to present cases of cavernous haemangioma with unusual clinical course. In seven patients with post-traumatic intracerebral haematoma, fragments of histologically confirmed cavernous haemangioma tissue situated in the place of haematoma were found. In one case, delayed intracerebral haematoma twenty four hours after trauma and initial CT-scan was observed. On the initial CT-scan in this patient only traumatic changes in the brain without haematoma or tumour were present. In our cases, cavernous haemangioma was situated most frequently in frontal and temporal region, shown as haematoma usually 24 hours after trauma, mainly in men in age range 30-44 years with disorders of consciousness (GCS 11). As the result of operation, the majority of patients (6 out of 7) were discharged as self-independent. The authors reviewed the literature for cavernous haemangioma. The tumour may be present in any region of the brain including infratentorial region. The lesion is disclosed most frequently in children as spontaneous brain haemorrhage and rarely as seizures or intracranial hypertension. Final diagnosis is based on cerebral angiography or MRI. CT-scans are not typical and before the appearance of haematoma do not suggest the presence of tumour. Surgical removal of haematoma with tumour fragments is the main method of therapy, but radiation is possible in order to diminish the mass. The authors conclude that in each patient with post-traumatic intracerebral haematoma, the presence of cavernous haemangioma is possible. PMID- 10540723 TI - [Sensory neuronopathy]. AB - The authors present the question of sensory neuronopathy which are disorders affecting intervertebral ganglia. The neuropathological background and clinical symptoms of sensory neuronopathy are emphasised, as well as diagnostic difficulties resulting from a variety of ethiological conditions: toxic, inflammatory, and autoimmunological ones, and from lack of unequivocal clinical criteria enabling a difference diagnosis along with neuropathy and radiculopathy, which in turn requires a broad spectrum of diagnostic tests and prolonged observation of patients. The authors discuss also the clinical outcome, prognosis, and current therapeutic possibilities focusing on intensive immunosuppressive management. PMID- 10540724 TI - [Autosomal dominant spinocerebellar ataxia]. AB - The modern classification is presented based on genetic criteria of the group of degenerative nervous system diseases inherited as autosomal dominant trait and called collectively spinocerebellar ataxia (SCA). They belong mostly to the class of diseases of similar mutation mechanism in which amplification is present of the trinucleotide sequence (CAG)n. Clinical picture and neuropathological changes in various SCA types are compared. PMID- 10540725 TI - [Prospects of measles and subacute sclerosing panencephalitis (SSPE) elimination in Poland]. AB - Following the introduction of vaccinations against measles in 1975 supplemented in recent years with booster vaccinations at the age of 6 years the epidemiological situation with respect to measles and SSPE has been gradually improving, particularly recently. In the paper discussion is presented on the question in what degree the present epidemiological situation of measles, the epidemiological supervision and vaccinations against measles in Poland meet the operative aim of measles and, consequently, SSPE elimination, as recommended by the WHO Regional Bureau. Attention is called to incomplete reliability of measles diagnosis based on clinical manifestations, economic difficulties in conducting serological investigations (detection of IgM antibodies to measles) necessary for measles confirmation, and shortcomings in vaccination organization. PMID- 10540726 TI - [How does the immune system communicate with the brain?]. AB - It is now evident, that the immune system and the central nervous system (CNS) are functionally connected and interact with one another. The entrance of pathogenic micro-organisms or their products into the body evokes an array of systemic responses, i.e. acute-phase proteinemia, neutrophilic leucocytosis, changes in the circulating levels of various hormones, and fever with specific changes of behaviour. It is now generally recognised that many of these responses are modulated by the brain. Until 1995 it was thought that the transport of immune signals to the brain is mediated by a humoral mechanism, mainly by certain cytokines, produced by activated monocytes and macrophages. The mechanism for the transfer of cytokines through the blood-brain barrier was however not completely understood until recently. It was also not clear, how the fast febrile phase can be signalled within 15 min, if the sequence: external pyrogen-->monocytes- >cytokines-->prostaglandins-->stimulation of the hypothalamus-->fever, requires 40-60 minutes to be activated. In addition to the "classical" humoral mode of transfer, it has recently been proposed that the immune signals are transported to the brain by certain peripheral nerves, predominantly by the vagus. The macrophages of the liver (Kupfer cells) start producing small amounts of cytokines in response to their activation by bacterial lipopolisaccharides, coupled into a complex with the already pre-activated anaphylatoxic component of the complement. These cytokines can immediately activate adjacent sensory paraganglia of the hepatic vagus nerve, which carries these stimuli to the nucleus tractus solitarius, which in turn sends them out to the hypothalamus via the ascending noradrenergic pathways. This fast avenue of communication between the immune system and the brain is activated not only in pathological situations, corresponding to fever, but is an important component of systemic homeostasis, e.g. regulation of body temperature. PMID- 10540727 TI - [Subcortical infarcts (caudate nucleus) in a case of bilateral anterior cerebral artery occlusion]. AB - The authors describe a patient with bilateral anterior cerebral artery (ACA) occlusion. CT and MRI revealed bilateral encephalomalacia in the regions supplied by Heubner arteries and/or by perforating branches of ACA. The patient presented mainly with frontal symptomatology resulting from caudate nuclei lesion. Frontal symptomatology due to caudate impairment is discussed in the sense of frontal subcortical circuits: lateral orbitofrontal and anterior cingulate ones. We emphasise a similarity of behavioural and cognitive disorders in early Huntington's disease and in frontal lobe lesion. PMID- 10540728 TI - [A sporadic case of Creutzfeldt-Jakob disease with peculiar neuropathological lesions]. AB - The authors report a case of Creutzfeldt-Jakob disease in a man aged 60 years from a family without a history of similar disease. The disease extended over 11 months. In the clinical picture initially equilibrium disturbances and dementia with psychotic symptoms predominated, EEG pattern was not typical of CJD. Neuropathological examination revealed extensive spongiform lesions in the cortex of all cerebral lobes, in striatum and substantia nigra, moreover a considerable number of kuru plaques was found in cerebellar cortex. The authors consider that the case meets the criteria accepted for the sporadic form of CJD but believe that the final differentiation from the Gerstmann-Streussler-Scheinker syndrome should be based on genetic studies. PMID- 10540729 TI - [Successful treatment with bacteriophage in purulent cerebrospinal meningitis in a newborn]. AB - The subject of this report is the case of purulent meningitis in new-born caused by Klebsiella pneumoniae. As the intensive antibiotic therapy turned out to be ineffective phage therapy was applied. Oral administration of specific phage preparate for the period of 5 weeks resulted in complete sterilization of cerebrospinal fluid and unquestionable improvement of child's health. However, after several ventriculopunctures some complications appeared (haemorrhage into central nervous system, extra infection). They were treated in standard way. Because of increasing internal hydrocephalus and necessity of operation, the child was sent to suitable hospital for further treatment. PMID- 10540731 TI - [Good and bad habits of presentations of papers. Advice and comments of the chairman]. PMID- 10540730 TI - [A rare case of intracranial hematoma complicating bacterial endocarditis]. AB - Endocarditis is an infectious and inflammatory disease of cardiac valves with other coexisting symptoms affecting heart and other organs and systems. Patients with cardiac valves diseases are in the group of high risk. The authors present a case of successfully treated endocarditis affecting mitral valve prosthesis in the course of staphylococcus septicaemia complicated by intracranial haematoma. PMID- 10540732 TI - [The effect of an excessive calcium intake during growth and embryonal development of calcium metabolism and skeletal development in the dog]. PMID- 10540733 TI - [Nutrition disorders in a dairy cow operation]. PMID- 10540734 TI - [Adelbert van Miert: what is the future of pharmacology?. Interview by Marianne Sloet van Oldruitenborgh-Oosterbaan]. PMID- 10540735 TI - [Nondominant aggression in golden retrievers: an inherited problem?]. PMID- 10540736 TI - Digestibility of the hydrogenated derivative of an isomaltooligosaccharide mixture by rats. AB - The digestibility of the hydrogenated derivative of an isomaltooligosaccharide mixture (IMO-H) was investigated. In an in vitro experiment, the digestibility of IMO-H was examined by models of the digestive system. IMO-H was resistant to two types of alpha-amylase and to artificial gastric juice. Enzymes in the rat small intestinal mucosa hydrolyzed tri-, tetra- and higher saccharide alcohols to disaccharide alcohol, removing successive glucose units from the non-reducing ends of the chains. The hydrolysis ratio for IMO-H was intermediate between the values for maltose and maltitol. In an in vivo study, growing rats were fed on an experimental diet containing IMO-H, maltitol, or hydrogenated palatinose in the range from 5% to 20%. The growth parameters of the rats fed on the test sugar show that the availability of IMO-H was about 1.2 to 1.25 times that of maltitol or hydrogenated palatinose. PMID- 10540737 TI - Responses of Mentha suspension-cultured cells to 2,4-dichlorophenoxyacetic acid and accumulation of esterified phenolic acids in their cell walls. AB - Two distinct types of cell growth of suspension-cultured Mentha were formed when the cells maintained in the medium containing 1000 micrograms l-1 2,4-D were subcultured into different 2,4-D concentrations. Few cell elongation of Mentha (average cell length: 34-40 microns) was observed after division in the medium containing 1-200 micrograms l-1 2,4-D; and significant cell elongation (average cell length: 95-130 microns) was observed after cell division in the medium containing 500-2000 micrograms l-1 2,4-D. A close correlation between culture medium and water content in the cells indicated that 2,4-D promoted cell elongation by water uptake. Amounts of phenolic acid in cell walls were much higher in unelongated cell walls than in elongated ones during the cultivation, and there was a close correlation between the amounts and the level of PAL activity in elongated and unelongated cells. However, there was no significant difference in cell wall components and its neutral sugar composition between elongated and unelongated cells. PMID- 10540738 TI - Characterization of Bacillus subtilis ExoA protein: a multifunctional DNA-repair enzyme similar to the Escherichia coli exonuclease III. AB - To discover the physiological role of the Bacillus subtilis ExoA protein, which is similar in amino acid sequence to Escherichia coli exonuclease III, an exoA::Cm disruption was constructed in the chromosomal DNA of B. subtilis. There was no clear difference in tolerance to hydrogen peroxide and alkylating agents between the disruptant and the wild type strain. An expression plasmid of the ExoA in E. coli was constructed by inserting the exoA gene into the expression vector pKP1500. The purified ExoA was used to clarify enzymatic characterizations using synthetic DNA oligomers as substrates. A DNA oligomer containing a 1', 2' dideoxyribose residue as an AP site, a DNA-RNA chimera oligomer, and a 3' end 32P labeled oligomer were synthesized. It has been shown that the ExoA has AP endonuclease, 3'-5' exonuclease, ribonuclease H, and 3'-phosphomonoesterase activities. Thus, it has been confirmed that ExoA is a multifunctional DNA-repair enzyme in B. subtilis that is very similar to E. coli exonuclease III except that ExoA has lower 3'-5' exonuclease activity than that of E. coli exonuclease III. PMID- 10540739 TI - Thermostabilization by proline substitution in an alkaline, liquefying alpha amylase from Bacillus sp. strain KSM-1378. AB - alpha-Amylase (LAMY) from alkaliphilic Bacillus sp. strain KSM-1378 is a novel semi-alkaline enzyme which has 5-fold higher specific activity than that of a Bacillus licheniformis enzyme. The Arg124 in LAMY was replaced with proline by site-directed mutagenesis to increase thermostability of the enzyme. The wild type and engineered LAMYs were very similar with respect to specific activity, kinetic values, pH-activity curve, and degree of inhibition by chelating reagents. Thermostability and structure stiffness of LAMYs as measured by fluorescence were increased by the proline substitution. The change of Arg124 to proline is assumed to stabilize the loop region involving amino acid residues from 122 to 134. This is the first report that thermostability of an alpha amylase is improved by proline substitution. PMID- 10540740 TI - Involvement of cytochrome a in iron oxidation of a moderately thermophilic iron oxidizing bacterium, strain TI-1. AB - The iron-oxidizing activity of a moderately thermophilic iron-oxidizing bacterium, strain TI-1, was located in the plasma membrane. When the strain was grown in Fe2+ (60 mM)-salts medium containing yeast extract (0.03%), the plasma membrane had iron-oxidizing activity of 0.129 mumol O2 uptake/mg/min. Iron oxidase was solubilized from the plasma membrane with 1.0% n-octyl-beta-D glucopyranoside (OGL) containing 25% (v/v) glycerol (pH 3.0) and purified 37-fold by a SP Sepharose FF column chromatography. Iron oxidase solubilized from the plasma membrane was stable at pH 3.0, but quite unstable in the buffer with the pH above 6.0 or below 1.0. The optimum pH and temperature for iron oxidation were 3.0 and 55 degrees C, respectively. Solubilized enzyme from the membrane showed absorption peaks characteristic of cytochromes a and b. Cyanide and azide, inhibitors of cytochrome c oxidase, completely inhibited iron-oxidizing activity at 100 microM, but antimycin A, 2-n-heptyl-4-hydroxyquinoline-N-oxide (HOQNO) and myxothiazol, inhibitors of electron transport systems involved with cytochrome b, did not inhibit enzyme activity at 10 microM. The absorption spectrum of the most active enzyme fraction from SP Sepharose FF column chromatography (4.76 mumol O2 uptake/mg/min) compared with lower active fractions from the chromatography (0.009 and 2.10 mumol O2 uptake/mg/min) showed a large alpha-peak of cytochrome a at 602 nm and a smaller alpha-peak of cytochrome b at 560 nm. The absorption spectrum of pyridine ferrohemochrome prepared from the most highly purified enzyme showed an alpha-peak characteristic of heme a at 587 nm, but not the alpha peak characteristic of heme c at 550 nm. The cytochrome a, but not cytochrome b, in the most highly purified enzyme fraction was reduced by the addition of ferrous iron at pH 3.0, indicating that electrons from Fe2+ were transported to cytochrome a, but not cytochrome b. These results strongly suggest that cytochrome a, but not cytochromes b and c, is involved in iron oxidation of strain TI-1. PMID- 10540741 TI - Steady-state inhibitory kinetic studies on the ligand binding modes of Aspergillus niger glucoamylase. AB - Inhibitory activities of 1-deoxynojirimycin and gluconolactone on Aspergillus niger glucoamylase were studied in relation to the subsite structure of the enzyme. Although both of these inhibitors are considered to bind at subsite 1 of the enzyme active site, 1-deoxynojirimycin showed competitive type inhibition but gluconolactone was a mixed type (or noncompetitive type) inhibitor for the hydrolysis of p-nitrophenyl alpha-D-glucoside. The former type of inhibition suggested that the main binding mode of the substrate was productive, but the latter, nonproductive. A possible way of explaining these apparent inconsistent results is to assume that the main binding mode of the substrate is productive and gluconolactone forms a nonproductive ternary complex with the enzyme and the substrate. PMID- 10540742 TI - Role of olfaction in food preference as evaluated in an animal model. AB - Food preference in individual animals is regulated by brain activity. Two murine model systems for investigating food preference were developed by focusing on fruit juices. In a home-cage, two-bottle test, the volume of apple juice consumed was found to be much larger than that of orange juice. In a two-nozzle "Drinkometer" test, by which each mouse was kept in a 38 cm (W) x 32 cm (D) cage and each drinking event was recorded by an electronic "Drinkometer" device, it was again found that the mice preferred drinking apple juice to orange juice. To elucidate the role of olfaction in this food preference, mice were subjected to an olfactory bulbectomy to remove the olfaction capability. In the home-cage two bottle test, the preference for apple juice over orange juice was apparent even after the olfactory bulbectomy, indicating that olfaction was not essential for the formation of food preference behavior. In contrast, in the two-nozzle "Drinkometer" test, the preference for apple juice over orange juice was found to be abrogated by this surgery, implying the involvement of olfaction-based memory on food preference behavior. PMID- 10540743 TI - Inhibition of Candida rugosa lipase by berberine and structurally related alkaloids, evaluated by high-performance liquid chromatography. AB - It is known that certain microorganisms produce extracellular lipase to better colonize the skin and mucosal surfaces. Since different extracts from medicinal plants have anti-lipase activity (Shimura et al., Biosci. Biotechnol. Biochem., 56: 1478-1479, 1992), we examined the effects of selected natural substances on Candida rugosa lipase. In the presence of the compounds under examination, the enzyme was incubated with beta-naphthyl laurate, and beta-naphthol, produced by the enzymatic reaction, was extracted with ethyl acetate and analyzed by reversed phase HPLC, using a C-18 column. Thus, the inhibitory activity was calculated by a proper formula based on the variations of the area under the chromatographic peak of beta-naphthol. The method was validated by analyzing substances with known anti-lipase activity such as saturated fatty acids (C10-16) and tetracycline. Berberine and a number of structurally related alkaloids such as chelidonine, chelerythrine, and sanguinarine appeared active. This property of berberine and sanguinarine is of interest because they are used in pathological conditions in which microbial lipases could play a pathogenic role. PMID- 10540744 TI - Selective and continuous degradation of carbazole contained in petroleum oil by resting cells of Sphingomonas sp. CDH-7. AB - Microbial degradation of carbazole (CA), a model of hard-removal heterocyclic nitrogen compounds contained in petroleum oil, was examined using Sphingomonas sp. CDH-7 isolated from a soil sample by screening for CA-assimilating microorganisms. CDH-7 used CA as a sole source of carbon and nitrogen, and metabolized CA to ammonia via anthranilic acid as an intermediate product. When CDH-7 was cultivated in the medium containing CA at the concentration of 500 mg/l (2.99 mM), CA was completely degraded within 50 h. By the reaction with the resting cells of CDH-7, 500 mg/l of CA was completely degraded within 4 h, with 1.64 mM of ammonia accumulated in the reaction mixture. When CA was added at the concentration of 100 mg/l (0.599 mM) periodically to the reaction mixture ten times, 925 mg/l (5.54 mM) of CA was degraded within 48 h by the resting cells, and 4.50 mM of ammonia was accumulated in the reaction mixture with a 75.1% molar conversion yield based on total CA added. The resting cells could almost completely degrade CA in a two-liquid-phase system which consists of water and organic solvent, even in the presence of 20% (v/v) isooctane, n-hexane, cyclohexane, and kerosene as a model petroleum oil. In the presence of an organic solvent system such as 20% (v/v) pxylene, toluene, and heptanol, however, CA degradation yields decreased. PMID- 10540745 TI - Effects of a probiotic on the lipid metabolism of cocks fed on a cholesterol enriched diet. AB - The effects of a probiotic (a mixture of Bacillus, Lactobacillus, Streptococcus, Clostridium, Saccharomyces and Candida) on the lipid metabolism, and caecal flora and metabolites of cocks were studied. The cholesterol level of the liver and serum was significantly decreased in the cocks fed on the cholesterol-enriched diet containing the probiotic. The distribution and frequency of occurrence of flora, and the chemical characteristics of the metabolites in the caecal content of the cocks were also affected by the inclusion of the probiotic in the basal and cholesterol-enriched diets. The Enterobacteriaceae species were significantly decreased in number, while the Bacillus, Streptococcus, Bifidobacterium and Lactobacillus species were significantly increased. The presence of yeast was observed, and the ammonia level was significantly reduced. The pH value, however, was not affected. The concentration of short-chain fatty acids in the caecal content of the cocks fed on the cholesterol-enriched diet supplemented with the probiotic was increased. It is, therefore, suggested that the incorporation of a probiotic in the diet would improve the balance of the intestinal flora and metabolites. Furthermore, it would also suppress the serum and liver cholesterol levels of cocks fed on the cholesterol-enriched diet. PMID- 10540746 TI - Expression of a recombinant molt-inhibiting hormone of the kuruma prawn Penaeus japonicus in Escherichia coli. AB - The crustacean molt-inhibiting hormone (MIH) suppresses ecdysteroid synthesis by the Y-organ. The MIH of the kuruma prawn Penaeus japonicus has recently been isolated and its cDNA cloned. In this study, we expressed the MIH in Escherichia coli to obtain a large quantity of this hormone with biological activity. The MIH cDNA was processed and ligated into an expression plasmid. E. coli was transformed with this plasmid, and then the recombinant MIH (r-MIH) was expressed. The r-MIH was put through the refolding reaction and was purified by reverse-phase HPLC. N-terminal amino acid sequence and time-of-flight mass spectral analyses supported the idea that the r-MIH had the entire sequence. By in vitro bioassay using the Y-organ of the crayfish, the r-MIH was found to be comparable to natural MIH in inhibiting ecdysteroid synthesis. PMID- 10540747 TI - Molecular cloning of isomaltotrio-dextranase gene from Brevibacterium fuscum var. dextranlyticum strain 0407 and its expression in Escherichia coli. AB - The gene encoding an extracellular isomaltotrio-dextranase (IMTD), designed dexT, was cloned from the chromosomal DNA of Brevibacterium fuscum var. dextranlyticum strain 0407, and expressed in Escherichia coli. A single open reading frame consisting of 1923 base pairs that encoded a polypeptide composed of a signal peptide of 37 amino acids and a mature protein of 604 amino acids (M(r), 68,300) was found. The primary structure had no significant similarity with the structure of two other reported exo-type dextranases (glucodextranase and isomalto dextranase), but had high similarity with that of an endo-dextranase isolated from Arthrobacter sp. Transformed E. coli cells carrying the gene encoding mature protein of IMTD overproduced IMTD under the control of the T7 phage promoter induced by IPTG. The purified recombinant enzyme showed the same optimum pH, lower specific activity, and similar hydrolytic pattern, as to those of native IMTD. PMID- 10540748 TI - Cloning, sequencing, and expression of the gene encoding the Clostridium stercorarium xylanase C in Escherichia coli. AB - The nucleotide sequence of the Clostridium stercorarium F-9 xynC gene, encoding a xylanase XynC, consists of 3,093 bp and encodes a 1,031-amino acids with a molecular weight of 115,322. XynC is a multidomain enzyme composed of an N terminal signal peptide and six domains in the following order: two thermostabilizing domains, a family 10 xylanase domain, a family IX cellulose binding domain, and two S-layer homologous domains. Immunological analysis indicated the presence of XynC in the culture supernatant of C. stercorarium F-9 and in the cells, most likely on the cell surface. XynC purified from a recombinant E. coli was highly active toward xylan and slightly active toward p nitrophenyl-beta-D-xylopyranoside, p-nitrophenyl-beta-D-cellobioside, p nitrophenyl-beta-D-glucopyranoside, and carboxymethylcellulose. XynC hydrolyzed xylan and xylooligosaccharides larger than xylotriose to produce xylose and xylobiose. This enzyme was optimally active at 85 degrees C and was stable up to 75 degrees C at pH 5.0 and over the pH range of 4 to 7 at 25 degrees C. PMID- 10540749 TI - Induction of isoflavonoid and retrochalcone branches of the flavonoid pathway in cultured Glycyrrhiza echinata cells treated with yeast extract. AB - Yeast extract-treated suspension cultures of a new cell line, AK-1, of Glycyrrhiza echinata were induced to produce an isoflavonoid phytoalexin (medicarpin) and metabolites of retrochalcone/flavone pathway (echinatin, licodione, and 7,4'-dihydroxyflavone). From these cells, putative full-length cDNAs encoding cytochrome P450s, (2S)-flavanone 2-hydroxylase and isoflavone 2' hydroxylase, were cloned. PMID- 10540750 TI - Efficient production of recombinant human erythropoietin by replenishment of microcarriers in the hollow fiber culture cassette. AB - Human erythropoietin (EPO)-producing recombinant BHK cells were cultured in culture medium containing microcarriers, and then microcarriers attached with cells were replenished in the hollow fiber culture cassette. By culture for 14 days, it was possible to produce 450 micrograms of the recombinant EPO, which corresponded to over two-fold of the recombinant EPO production by control hollow fiber culture without microcarriers. PMID- 10540751 TI - (Z)-3-hexenyl and trans-linalool 3,7-oxide beta-primeverosides isolated as aroma precursors from leaves of a green tea cultivar. AB - 6-O-beta-D-Xylopyranosyl-beta-D-glucopyranosides (beta-primeverosides) of (Z)-3 hexenol and trans-linalool 3,7-oxide were newly isolated from fresh leaves of a tea cultivar (Camellia sinensis var. sinensis cv. Yabukita). In addition, the already identified beta-primeverosides of benzyl alcohol, methyl salicylate, and trans-linalool 3,6-oxide from an oolong tea cultivar were isolated from the Yabukita cultivar. It was confirmed that all aglycones of the linalool oxide glycosides isolated here were of the optically active S-form by chiral GC after enzymatic hydrolysis with glycosidase. PMID- 10540752 TI - Hypertriglyceridemia in rats induced by consumption of a food-derived carcinogen, 2-amino-1-methyl-phenylimidazo[4,5b]pyridine (PhIP). AB - 2-Amino-1-methyl-phenylimidazo[4,5b]pyridine (PhIP), the most abundant mutagenic heterocyclic amine produced in cooked meat and fish, is known to be a carcinogen for rats and mice. This study provides the first evidence for hypertriglyceridemia in rats exposed to PhIP, suggesting its potential risk to induce not only carcinogenesis, but also atherosclerosis, and highlighting the potential importance of PhIP for humans. PMID- 10540753 TI - New potent antioxidative o-dihydroxyisoflavones in fermented Japanese soybean products. AB - A potent antioxidative 6-hydroxydaidzein (6-OHD) was newly isolated from soybean koji fermented with Aspergillus oryzae. 6-OHD, in addition to 8-hydroxydaidzein and 8-hydroxygenistein, were found to be present in various fermented soybean products, including their koji. Considering that these o-dihydroxyisoflavones had strong antioxidative activities, they may contribute to protecting from oxidative deterioration during the processing of fermented soybean products. PMID- 10540754 TI - 3-Geranyl-4-hydroxy-5-(3'-methyl-2'-butenyl)benzoic acid as an anti-inflammatory compound from Myrsine seguinii. AB - Bioassay-guided isolation of anti-inflammatory compounds from the methanol extract of Myrsine seguinii yielded an anti-inflammatory compound (1). The structure of compound 1 was elucidated to be 3-geranyl-4-hydroxy-5-(3'-methyl-2' butenyl)benzoic acid on the basis of its spectroscopic data. Compound 1 strongly suppressed 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced inflammation on mouse ears at a dose of 500 micrograms (inhibitory effect (IE): 65%). The acetate and the methyl ether of 1 showed moderate activity at a 500-microgram application, with IE 38% and 27%, respectively. However, the methyl ester and the dimethyl derivative of 1 did not show activity at the same dose. The related compounds of 1, o-, m- and p-hydroxybenzoic acids also did not exhibit notable activity. These results indicate that the carboxylic acid and lipophilic terpene moieties of 1 were significant structural features for anti-inflammatory activity. PMID- 10540755 TI - 1,2-Hydride Shift in the Biosynthesis of Pinguisane-type Sesquiterpenes. AB - A (2)H-NMR analysis of 6alpha-hydroxy-3-oxo-pinguis-5(10)-ene-11,6-olide produced by axenic culture of liverwort Aneura pinguis in the presence of [4-(2)H2] labeled mevalonate clarified the presence of a 1,2-hydride shift and retention of deuterium at the C-4 position in the biosynthesis of pinguisane-type sesquiterpenes from FPP. PMID- 10540756 TI - Ego functions and ego development: defense mechanisms and intelligence as predictors of ego level. AB - This study considers the contribution of two ego functions--intelligence and defense mechanisms--to ego developmental level. Two independent assessments of ego level were related to IQ and defense mechanism use in a sample of 89 young adults. Whereas IQ and defense were themselves found to be unrelated, both variables predicted ego level: The relation with IQ was linear, whereas the relation with defense was curvilinear. In addition, the relation between defense and ego level varied as a function of IQ level. At low levels of IQ, stronger use of Denial and Projection was associated with higher ego levels. At high IQ levels, strong use of Denial was associated with lower ego levels, whereas moderate use of Projection was associated with higher ego levels. PMID- 10540757 TI - Susceptibility to affect: a comparison of three personality taxonomies. AB - This study had three major goals: to clarify the relationships between Eysenck's, Gray's, and Cloninger's personality taxonomies, to show that traits from these taxonomies predict differential sensitivities to emotional states, and to explore the relationship between sensitivity to an emotional state and how much that state is actually experienced. A factor analysis of traits from Eysenck's, Gray's, and Cloninger's personality taxonomies resulted in three factors that were named reward sensitivity, impulsivity-thrill seeking, and punishment sensitivity. These factors predicted a global measure of affect, emotional reactions to a laboratory mood induction, and self-reported affect in daily life. Generally, reward sensitivity predicted positive, but not negative emotions, whereas punishment sensitivity predicted negative, but not positive emotions. Impulsivity-thrill seeking predicted few emotions in either context. Coherence among the relationships found across methodological contexts suggests that the traits that predict emotion susceptibilities in the laboratory similarly predict emotional experience in ongoing daily life. PMID- 10540758 TI - Body image and day-to-day social interaction. AB - Participants maintained a social interaction diary and completed a measure of body image. Body image was found to have three factors, body attractiveness, social attractiveness (how attractive people believed others found them to be), and general attractiveness. For both men and women, self-perceptions of body attractiveness and of social attractiveness were positively related to the intimacy they found in interaction. Self-perceptions of social attractiveness were positively related to women's confidence in social interaction and their perceived influence over interaction, whereas for men, confidence and influence were unrelated to social attractiveness. For both men and women, body image was unrelated to how enjoyable people found interactions to be and was weakly related to how responsive they felt others were to them. For both men and women, body image was also unrelated to how socially active people were and to the relative distribution of same- and opposite-sex interactions. PMID- 10540759 TI - Self and identity in advanced old age: validation of theory through longitudinal case analysis. AB - Case studies drawn from a 20-year longitudinal study of aging were examined for the support they provide to two theoretical viewpoints on the self in later life: one focusing on management of self-esteem, the other on development of identity as story. The five cases selected for scrutiny represented diverse trajectories of self-esteem. They furnished ample illustrations of certain key aspects of both theories, including assimilative processes of coping, depression related to absence of accommodation, maintenance of life story themes, and life review processes. They did not, however, give strong support to the dichotomy, drawn within both theoretical models, between younger and older old age. Examples of accommodation, disengagement, and self-transcendence, hypothesized to typify advanced old age, were relatively few in number and emerged only toward the very end of life. It is argued that examination of prototypical cases provides a useful approach to validating and developing theory. A conclusion drawn from this study is that more analysis should be carried out on the lives of persons who exemplify the theoretically ideal characteristics of advanced old age. PMID- 10540760 TI - [The last fling syndrome]. PMID- 10540761 TI - [The historical setting of cystine lithiasis]. AB - W. H. Wollaston, who first described cystine stones, as well as the most outstanding contemporary figures and their contribution to the understanding of this uncommon type of lithiasis are described. PMID- 10540762 TI - [Fournier's gangrene. Our experience over 10 years. A review of the literature]. AB - OBJECTIVE: To report our experience over the last 10 years with Fournier's gangrene, an extensive fulminant infection of the perineoscrotal region, and to review the literature. METHODS: The medical records of 9 patients with Fournier's gangrene that had been diagnosed from January 1988 to December 1997 were reviewed. Patient age, etiology and predisposing factors, microbiological findings, duration of hospital stay, treatment and outcome were analyzed. RESULTS: The mean age of the patients was 53.8 years (range 43-71). The source of the gangrene was perirectal (22.22%), urinary (66.66%) and cutaneous (11.11%). Predisposing factors included diabetes mellitus, alcoholism, malnutrition and low socio-economic status. All patients were treated with surgical debridement and broad-spectrum antimicrobial therapy. Two patients underwent delayed reconstructive surgery. Cystostomy was performed in 100% of the cases. Two patients died from severe sepsis. CONCLUSIONS: Necrotizing fasciitis of the perineum and genitalia is a severe condition with a high morbidity and mortality. Good management is based on aggressive debridement, broad-spectrum antibiotics and intensive supportive care. PMID- 10540763 TI - [Inflammatory changes in the obstructed prostate: the correlation between the bacteriological and histological findings]. AB - OBJECTIVES: To analyze the incidence and nature of the prostatic inflammatory changes, to determine the prevalence of prostatic colonization or infection in patients undergoing surgery for benign hyperplasia of the prostate (BPH) and to correlate the inflammatory lesions with the bacteriological findings. METHODS: A prospective study was conducted on 175 patients undergoing surgery for BPH. All patients were entered into a study protocol that included quantitative bacterial cultures of the prostatic tissue and histological analyses of the surgical specimens. Data of previously defined variables were entered into a data base for subsequent analysis comprised of a descriptive and an analytical study. RESULTS: 44 of the 175 patients (25.1%) had a positive bacterial culture of prostatic tissue. Histological lesions indicating prostatitis associated with BPH were found in 68 of the 175 patients (38.9%), regardless of the presence or absence of bacteria. Of these 68 patients with a histologically demonstrated inflammation of the prostate, only 19 (27.9%) had positive prostatic tissue cultures. The incidence of granulomatous prostatitis was 1.1%. CONCLUSIONS: Histological lesions indicating prostatitis associated with BPH were found in 68 of the 175 patients (38.9%). The presence of bacteria was demonstrated in the prostates of a significant number of patients (25.1%) who underwent adenomectomy for BPH. No differences were found between the patients with a positive or negative bacterial culture and histological evidence of prostatitis. PMID- 10540764 TI - [The clinical and flowmetric results of the treatment of benign prostatic hyperplasia with doxazosin]. AB - OBJECTIVE: To evaluate the clinical efficacy of doxazosin for 6 months in the treatment of benign prostatic hyperplasia (BPH). METHODS: A prospective clinical and uroflowmetric study was conducted on 65 males with BPH treated with doxazosin 4 mg daily for 6 months. Patient mean age was 66.7 years (range 45-79). Clinical evaluation (IPSS and andrologic data) and blood analyses were performed before and after treatment. IPSS data were obtained according to the WHO validation and Spanish translation. RESULTS: A significant improvement was found between the mean pre- and post-treatment IPSS scores (19.8 +/- 4.8 vs 11.9 +/- 4.6; p < 0.001). Maximum flow rate before treatment was 9.13 ml/sec, which increased to 16.23 ml/sec after treatment (p < 0.001). Postvoid residual urine dropped from 21.7% to 12.5% (p < 0.01). All the patients were normotensive before (135.9 mean systolic and 78.9 mean diastolic blood pressure) and after treatment (135.4 mean systolic and 77.8 mean diastolic blood pressure). Mean heart rate was similar before and after treatment (71.9 +/- 5.8 vs 71.8 +/- 5.9). A relationship between low IPSS score before treatment and urinary symptoms improvement was demonstrated (coeff. -0.45939). No relationship was found between prostate volume (digital rectal examination or transabdominal ultrasound) and improvement in the IPSS score. No statistical relationship was found between the IPSS and postvoid residual urine or peak flow. No modifications of sexual activity was demonstrated with doxazosin treatment. Pre- and post-treatment blood analytical data fell within the normal ranges. Transient side effects were observed in 12 patients (20%): headache in 6 patients (10%), fatigue in 6 (10%), dizziness in 3 (5%) and somnolence in (5%). CONCLUSIONS: Doxazosin, at a daily dose of 4 mg daily for 6 months, is a safe and effective treatment in patients with BPH. PMID- 10540765 TI - [BCG and radiotherapy: the compatibility of 2 conservative treatments for cancer of the urinary bladder]. AB - OBJECTIVE: To review the efficacy of radiotherapy and BCG in the treatment of transitional cell carcinoma of the urinary bladder in its different forms of presentation, with special reference to patients with infiltrating bladder tumors receiving radiotherapy and those in whom the lesion recurs as a high grade superficial bladder tumor. METHODS/RESULTS: 10 patients who previously received radiotherapy for T2-4 infiltrating bladder tumor that recurred as a high grade superficial tumor were treated with BCG. Four patients are alive and disease-free with a preserved bladder at 2-8 years follow-up. Four other patients who required cystectomy for persistence or progression of the tumor to the bladder wall, are alive and disease free at 3-7 years follow-up. The remaining two patients who were not amenable to major surgery died from the disease more than two years after treatment with BCG. BCG was well-tolerated by 70% of the patients and the rest showed minor complications. CONCLUSIONS: 28.3% of recurrences after radiotherapy are superficial tumors and 7% are carcinoma in situ. The appearance of carcinoma in situ or T1 G3 lesions following radiotherapy of the bladder questions its efficacy against these superficial forms for which cystectomy is reserved. BCG has been found to be effective in high grade superficial bladder tumors that have not been previously irradiated, therefore it would be acceptable to extend its application to those patients in whom radiotherapy has achieved control of the infiltrating tumor but not the high grade superficial tumor. The 40-70% of patients who are alive with a preserved bladder appears to be sufficient to recommend BCG salvage for high grade superficial bladder tumors post-radiotherapy. BCG therapy does not entail major complications or compromise patient survival, including those cases that will require cystectomy. PMID- 10540766 TI - [The prevention of stage-T1 superficial bladder tumors with 27 mg. of BCG weekly over 6 weeks]. AB - OBJECTIVE: To evaluate the utility of prophylactic treatment of stage T1 superficial tumors of the bladder with 27 mg BCG weekly for 6 weeks and to compare the results reported in the literature. METHODS: BCG instillations were offered to 235 patients and was accepted by 111 (group A) and refused by 124 (group B). Three weeks thereafter, intravesical instillation of 27 mg BCG was administered for 6 weeks. The patients were controlled regularly according to the standard control procedures utilized in our setting. RESULTS: 39% of the patients in group A showed recurrence versus 71.7% of those in group B (p < 0.001). No differences in progression of tumor stage was observed; 6.3% for group A and 10.9% for group B. By grade, significant differences were found in the number of recurrence in those with G1 (28.5% vs 69%; p < 0.001) and G2 (47% vs 72%; p < 0.01) tumors, but not for G3 (53% vs 77%; p = n.s.). No differences were found in the number of progressions. For those with G2 and G3, the results were not as good as those reported in the literature. The incidence of toxicity was 33%. CONCLUSIONS: The results achieved in patients with G2 and especially G3 tumors were not as good as those reported in the literature, therefore we do not recommend this approach. For those with G1 tumors and assuming a toxicity rate of 33%, the results are similar to those reported elsewhere using higher doses, and therefore this approach could be utilized. PMID- 10540767 TI - [Renal artery stenosis in the transplant patient]. AB - OBJECTIVE: To present our experience in the diagnosis and treatment of renal artery stenosis in kidney transplants. METHODS: A review of 601 renal transplants performed in adults showed 32 cases of renal artery stenosis. The diagnostic techniques utilized were arteriography in 18 patients, DIVAS in 15, echo-Doppler in 11 patients and MAG with captopril test on two occasions. RESULTS: Arterial hypertension was the most common symptom (92.8%), alone (53.1%) or in association with impaired renal function (43.7%). 46.8% of the cases could be managed by drug therapy. Percutaneous transluminal angioplasty was performed in 14 patients. Surgery was required on two occasions. CONCLUSIONS: The incidence of renal artery stenosis in our series of renal transplants in adults up to 1997 was 5.3%. Arterial hypertension with or without impairment of renal function was the most common symptom. Currently, echo-doppler and MAG with captopril test are the most widely utilized diagnostic techniques. Percutaneous transluminal angioplasty is the treatment of choice in renal artery stenosis when arterial hypertension is refractory to drug therapy. Good results are achieved in 57%, although it is not free from complications. In case of failure, revascularization surgery is the alternative approach. PMID- 10540768 TI - [The usefulness of perineal ultrasound in urinary incontinence in women]. AB - OBJECTIVE: To analyze the utility of perineal ultrasound in the evaluation of female urinary incontinence. METHODS: The present study was conducted on 50 women with urinary incontinence. The distance between the bladder neck and symphysis pubis with and without stress was measured by perineal ultrasound. A displacement of the bladder greater than 10 mm was considered cystocele, and if the bladder neck and urethra formed a funnel greater than 13 mm, it was considered urinary incontinence. RESULTS: The distance between the bladder neck and the symphysis pubis in all 50 patients ranged from 10 mm to 18 mm (mean 12.8 mm) without stress and from 15 mm to 42 mm (mean 32.6 mm) with stress (Valsalva maneuver and/or cough). Perineal ultrasound was useful in all cases since one could evaluate the descent of the angle of the bladder neck in the case of cystocele and a lesser or greater funneling of the urethra, which is an essential sign in urinary incontinence. CONCLUSIONS: Perineal ultrasound can determine which patient has a cystocele, urinary incontinence or both. It is useful in the evaluation and follow-up of patients undergoing surgery for urinary incontinence. PMID- 10540769 TI - [An aggressive inguinal (parafunicular) angiomyxoma in a male patient]. AB - OBJECTIVE: To report a case of aggressive inguinal angiomyxoma in a male patient. METHODS: An 82-year-old male patient presented with a well-defined, 6 cm. parafunicular mass in the right groin. The mass was located adjacent to the spermatic cord and had been noted 8 years earlier. Patient evaluation included CT, ultrasound and immunohistochemical studies. RESULTS: The CT and US findings suggested lymph node enlargement. Microscopic analysis showed a myxoid tumor with partially infiltrating margins, vascular channels of small-sized vessels with thick walls occasionally with hyalinization and spindle-shaped or stellate mesenchymal cells with ill-defined margins without atypia or mitosis that were positive for vimentin and negative for actin, desmin, keratins, CD34 and protein S-100. No tumor recurrence or metastasis has been observed at 26-months' follow up. CONCLUSIONS: To our knowledge, this is one of the few cases of inguinal angiomyxoma in male patients; 16 have been reported to date. This neoplasm appears to originate from pelvic soft tissue fibroblasts. PMID- 10540770 TI - [A renocolic fistula diagnosed by retrograde pyelography]. AB - OBJECTIVE: To describe a case of renocolic fistula diagnosed by retrograde pyelography. METHODS: A case of renocolic fistula associated with pyonephrosis and left pleural empyema in a 48-year-old female with a history of recurrent pyelonephritis is presented. RESULTS: The ultrasound findings were suggestive of pyonephrosis and an excretory urogram showed a non-functioning left kidney. Retrograde pyelography showed passage of contrast medium into the descending colon. Surgical treatment consisted of nephrectomy, exteriorization of the compromised colon and pleurotomy. The patient died from septic shock. CONCLUSIONS: The utility of retrograde pyelography in the diagnosis of renocolic fistula is demonstrated in the case described herein. PMID- 10540771 TI - [A bifid ureter with a blind branch]. AB - OBJECTIVE: To report on a case of bifid ureter with a blind-ending branch. METHODS: A case of bifid right ureter with a blind-ending branch and complete duplication of the contralateral ureter in a 38-year-old female patient is described. The patient had consulted for pain referred to the right iliac fossa and recurrent urinary infection. RESULTS: An excretory urogram disclosed the ureteric anomaly, which was confirmed by retrograde pyelography. Uretero-ureteric reflux to the blind-ending branch was found during surgery and excision of the blind-ending branch was performed. CONCLUSIONS: This ureteric anomaly is basically diagnosed radiologically. Treatment is initially conservative, although complications or severe symptoms may warrant surgical excision of the blind ending ureter. PMID- 10540772 TI - [Squamous carcinoma of the male urethra, its presentation as a scrotal abscess]. AB - OBJECTIVE: To report a case of urethral carcinoma presenting as scrotal abscess. METHODS: A 53-year-old man presented with swelling, redness and pain in the scrotum. He had a history of urethral stricture and multiple scrotal abscesses. Incision and drainage were performed and the patient was treated with antibiotics without improvement. Incision and drainage were repeated and a biopsy of the tissue edges of the abscess were performed. RESULTS: Histopathological analysis disclosed a squamous cell carcinoma arising from the urethra. CONCLUSION: Patients with a history of urethral stricture should be followed closely. A scrotal biopsy should be performed in patients who do not improve with antibiotic therapy. PMID- 10540774 TI - [Intravesical hemangiopericytoma]. AB - OBJECTIVE: To report on a case of hemangiopericytoma of the urinary bladder mimicking a urothelial tumor. METHODS: A case of hemangiopericytoma arising from the urinary bladder in a 78-year-old man is described. The initial symptoms included gross hematuria, and it was confused with a urothelial tumor. The difficulty in making the diagnosis in regard to the site of origin and histological findings are discussed. The literature is briefly reviewed. RESULTS/CONCLUSIONS: Ultrasound and urographic evaluation showed an intravesical mass which was suspected to be a urothelial tumor. The patient was submitted to transurethral surgery. Histological analysis and immunohistochemical techniques demonstrated a hemangiopericytoma. PMID- 10540773 TI - [A simple renal cyst, the origin of Wunderlich's syndrome]. AB - OBJECTIVE: An additional case of Wunderlich syndrome arising from a ruptured simple renal cyst is presented. METHODS/RESULTS: A 50-year-old female presented with pain, a mass in the left renal fossa and hypovolemia that could not be managed conservatively. The imaging techniques disclosed a large collection of blood in the left perirenal area. Surgical drainage of the hematoma and nephrectomy were performed. A simple cyst containing blood was found in the lower pole of the kidney. Its wall had been ruptured and had caused the hemorrhage. CONCLUSIONS: If surgical management of hemorrhage is decided in a patient with Wunderlich's syndrome, a conservative procedure should be performed and nephrectomy avoided if the underlying condition is benign. PMID- 10540776 TI - [Recurrent orchiepididymitis, 6 months after miliary tuberculosis]. PMID- 10540775 TI - [Bladder leiomyoma with extramural growth. An infrequent cause of pelvic pain]. AB - OBJECTIVE: A case of extravesical leiomyoma that had been incidentally discovered in a patient presenting with unspecific pelvic pain is described. METHODS/RESULTS: A 51-year-old male patient consulted for unspecific pelvic pain. Patient evaluation disclosed a well-defined mass. A partial cystectomy was performed. Pathological analysis confirmed the diagnosis of leiomyoma of the bladder. CONCLUSIONS: Mesenchymal tumors of the urinary bladder are rare and account for 1%-5% of all bladder tumors. Leiomyoma is the most common benign non epithelial tumor. The symptoms caused by this tumor type is varied, unspecific and in most of the cases depends on its location (intravesical, intramural or extramural). Treatment is by surgery, which achieves excellent results. PMID- 10540777 TI - Outcome of colposuspension in patients with stress urinary incontinence and abnormal cystometry. AB - OBJECTIVE: To analyze the prognostic value of preoperative cystometric alterations in the outcome of women undergoing colposuspension for stress incontinence. PATIENTS AND METHODS: Over a 5 year period, 220 women were operated on for stress urinary incontinence using the Burch or Marshall colposuspension techniques. An abnormal cystometry was found pre-operatively in 44 (20%), which was associated with urge incontinence in 11 (25%). Cystometric abnormalities comprised 3 subgroups: detrusor instability (DI), low bladder compliance (LBC) and small detrusor contractions (SDC). Women with an abnormal cystometry had responded partially to anticholinergic therapy. Detailed postoperative questioning was undertaken to differentiate stress from urge incontinence, as well as storage symptoms. Results of patients with cystometric abnormalities were compared to an age-matched group of 44 patients with a stable bladder on the preoperative study. RESULTS: Bladder compliance was statistically lower in the preoperative CMG of patients with abnormal cystometry (p < 0.005). Groups were followed for a mean of 39 (abnormal CMG) and 36 months (stable bladder), respectively. The presence of the aforementioned cystometric alterations was not associated with lower cure rates of the stress incontinence. However, the group with DI referred more postoperative storage symptoms. "De novo" DI was found in 20% of patients with a previously stable bladder who referred storage symptoms postoperatively. CONCLUSIONS: Small detrusor contractions are not a contraindication for colposuspension. Patients with DI and low bladder compliance who also have stress incontinence showed more storage symptoms on postoperative evaluation. PMID- 10540778 TI - Laparoscopic cholecystectomy in routine practice: duct injury as an index event. PMID- 10540779 TI - Outcome of diabetic pregnancy with spontaneous labour after 38 weeks. AB - One hundred and forty-eight patients with well controlled insulin dependent diabetes that were allowed to labour spontaneously from 1981 to 1994 were reviewed. There were 2 perinatal deaths, giving a perinatal mortality rate of 13.5/1000. One hundred and twenty-four patients (84 per cent) had a normal vaginal delivery, 13 (9 per cent) forceps delivery and 11 (7 per cent) caesarean section. Twenty-one infants (14 per cent) required admission to a Special Care Baby Unit. One third of infants weighed 4 Kg or more, however there was only 1 case of shoulder dystocia. We compared these results with those of the general hospital population of 1987. The 2 main differences are; 1) the Caesarean section rate in labour was higher for this diabetic group than for the general hospital population, 7 per cent versus 3.4 per cent, 2) the birth weight was heavier, 33 per cent of infants of the diabetic group weighed 4 Kg or more versus 18 per cent of the general hospital population. The other parameters were comparable. We conclude that conservative management of pregnancy in well controlled diabetic women is advantageous, resulting in a high vaginal delivery rate without an increase in shoulder dystocia, and a low perinatal morbidity and mortality rate. PMID- 10540780 TI - Idiopathic mesenteric thrombosis following caesarean section. AB - Mesenteric venous thrombosis, "the great mimicker", is a very rare disorder in pregnancy and the puerperium, particularly when not associated with any pre existing thrombophilia or autoimmune states. We describe a patient requiring a resection of 150 cm of gangrenous small bowel after uncomplicated elective Caesarean section. The only risk factor for thrombosis was recovery from an elective Caesarean section, a condition classified by the Royal College of Obstetricians and Gynaecologists as "low risk". Death from thromboembolism is the leading cause of maternal mortality and should always be considered with unusual post partum symptoms. Early diagnosis of mesenteric vascular occlusion is difficult and recent evidence suggests that elevated GST isoenzyme may be helpful. In all cases of MVT anti-coagulation is the basis of treatment. Patients who are not anti-coagulated after surgery have a recurrence rate of 25 per cent compared with 13 per cent of heparinised post-operative patients. As no other pre existing cause for MVT was found, management was with warfarin for 6 months, the oral contraceptive pill was contraindicated and heparin prophylaxis was recommended for future pregnancies. PMID- 10540781 TI - Application of red cell distribution width to screening for coeliac disease in insulin-dependent diabetes mellitus. AB - Coeliac disease has been reported to occur in 2-5 per cent of insulin-dependent diabetic patients (IDDM). Suitable non-invasive screening tests would allow identification of these patients. The aim of this study was to determine the value of the red cell distribution width (RDW) in detecting unrecognised coeliac disease in insulin-dependent diabetic patients (IDDM). All patients (n = 208) attending the Diabetic out-patient clinics at 2 adjacent centres who had a full blood picture and RDW carried out in the past 18 months were included. IDDM patients with an elevated RDW were identified and their charts were reviewed to determine if they had symptoms or laboratory abnormalities compatible with coeliac disease. They were invited to attend for serological screening. Ninety five of 208 patients had an elevated RDW of whom 66 had non-insulin dependent diabetes mellitus and 29 had IDDM. Two of the 29 IDDM patients had died in the interim period. Six of the remaining 27 IDDM patients had previously been tested for serological markers associated with coeliac disease, of whom 1 had a positive antigliadin antibody titre (IgA-AGA 199 EU) and normal duodenal biopsy. Eighteen of the remaining 21 patients with IDDM consented to serological testing of whom only 1 had a positive titre of antiendomysial antibody (IgA-EMA) and villous atrophy. Although the RDW is known from previous studies to be a sensitive predictor for coeliac disease, this study has demonstrated its poor specificity in predicting IDDM patients who may have coeliac disease. The RDW is not recommended as a screening test for coeliac disease in patients with IDDM. PMID- 10540782 TI - Pain syndromes post cardiac transplantation. PMID- 10540783 TI - National school teachers' knowledge of asthma and its management. AB - Asthma is the most common chronic medical condition that school teachers may encounter among their pupils. However management of asthma in schools and the role school teachers adopt in this condition has only recently been explored. The aim of this study was to determine teachers' knowledge of asthma and its management. A postal questionnaire was circulated to 199 school teachers from 46 schools in Dublin City. A 74 per cent response rate was obtained. The number of children with asthma as identified by teachers was 7.8 per cent which suggests that asthma may be unrecognised in a number of pupils. Knowledge on signs and symptoms of asthma, provoking factors of asthma and the nature of the disease was generally satisfactory. However, knowledge on asthma medications, the purpose of inhalers and teachers' understanding of the treatment and management of asthma was considered poor. Knowledge on exercise-induced asthma was limited. There is a need to provide school teachers with education on asthma and its management. School policies on asthma also need to be developed with particular reference to action necessary in the event of an acute severe attack of asthma. PMID- 10540785 TI - Alcoholic ketoacidosis presenting as diabetic ketoacidosis. PMID- 10540784 TI - The identification and assessment of undernutrition in patients admitted to the age related health care unit of an acute Dublin general hospital. AB - OBJECTIVES: To profile those over 65 yr admitted to an acute geriatric medical service. To identify and assess their undernutrition risk and quantify the nutritional intervention they received. METHODS: Forty-nine consecutive admissions were recruited, 23 various parameters were assessed. A diagnosis of undernutrition was made according to a specifically designed flow chart. RESULTS: Eighty-four per cent of recruits were at risk of undernutrition on admission and 80 per cent were moderately to severely at risk. There was deterioration from baseline nutritional status in 29 per cent of previously well nourished patients with hospitalization. In the undernourished group, an improvement and/or stasis from baseline was achieved in 75 per cent, with intervention. CONCLUSIONS: The risk of undernutrition on admission to hospital and during treatment is an indicator of the need for nutrition services and nutritional screening for all acute medical services for older people. PMID- 10540786 TI - The detection and quantitation of 7-aminoflunitrazepam, the major urinary metabolite of flunitrazepam, by gas chromatography/mass spectrometry. AB - A sensitive gas chromatography/mass spectrometry method for the detection and quantitation of 7-aminoflunitrazepam, the major urinary metabolite of flunitrazepam, is described. The method is based upon solvent extraction followed by derivatisation with methyl-bis-trifluoroacetamide (MBTFA), to give a trifluoroacetyl derivative. A profile of 7-aminoflunitrazepam urinary concentrations following ingestion of 0.5 to 4 mg oral doses is reported. The method has been found to be reproducible and can be used for the confirmation of flunitrazepam administration. PMID- 10540787 TI - Effect of Doppler flow and lipid studies in a geriatric population--increased flow left internal carotid artery only. AB - Twenty geriatric subjects were treated with recombinant human growth hormone and results compared with a control group. The maximum dose of 3 units per day for 4 months was achieved in all patients. All patients had established atherosclerosis. Doppler Flow Studies were carried out at zero, 6 and 16 weeks. No alteration in plaque size or intima media thickness was observed. An increase in peak velocity in the treated group with a decrease in the control group in the left internal carotid artery only was observed. The study demonstrated a reduction in antero-posterior diameter of the left internal carotid in the treated group but the lesion grading remained unaltered. The expected reduction in total cholesterol LDL was however accompanied by a significant transient increase in triglyceride and P.A.I. 1. Fibrinogen levels were unaltered. PMID- 10540788 TI - Prevalence of rheumatoid arthritis in Dublin, Ireland: a population based survey. AB - The prevalence of Rheumatoid Arthritis (RA) in Ireland has never been established. Studies from different countries show varying rates, being almost 100 per cent greater in the highlands of Scotland (10/1,000) than in rural Lesotho (6/1,000). A recent study also suggests a fall in the prevalence of RA among women in the London urban area. Given these variations the validity of extrapolating prevalence rates established for other countries to Ireland is questionable. This study aimed to establish a prevalence rate for RA in a defined Dublin population. A self-administered questionnaire was sent to 2,500 people chosen at random from the electoral register. The questionnaire was designed to select out both undiagnosed patients and those with definite arthritis. Respondents whose replies indicated an arthritic process, but in whom no diagnosis had been made, were asked to attend for further assessment and investigations as appropriate. Those who responded that they had been diagnosed with arthritis were asked for consent to inspect their hospital or general practitioner records. A diagnosis of RA was based on American Rheumatism Association (ARA) criteria. Valid responses were received from 1,227 people surveyed (response rate = 49 per cent). Six cases of RA were identified including 2 previously undiagnosed cases. A prevalence rate of 5/1,000 has been estimated based on these findings. PMID- 10540789 TI - Tonic and phasic activity in smooth muscle. AB - We have studied the electrical and mechanical behaviour of two very different smooth muscle preparations, mesenteric lymphatic ducts and proximal urethra. These tissues generate different patterns of spontaneous contraction adapted to fulfil their contrasting functions. While lymphatics undergo regular phasic contractions and relaxations, suited to their role in propelling lymph, the urethra remains in a state of contracture to maintain urinary continence. The challenge is to understand how both of these achieve their respective roles. Interestingly, electrical activity of lymphatics resembles that in the heart in having a one to one relationship between the action potential and phasic contraction. Patch clamp studies have shown that lymphatic cells express 3 ionic currents that are not present in urethral cells, but are shared with cardiac muscle. These are, i) fast Na+ current, ii) T-type Ca2+ current and iii) a hyperpolarization-activated cation current, Ir. The fast Na+ current is ideally suited to the propagation of the action potential over large distances, as required by a vessel capable of generating a rapid well co-ordinated contraction along its length. The T-current and Ir, on the other hand, appear to be involved in electrical pacemaking as they are in the heart. The urethra does not usually undergo regular phasic contractions and it lacks these currents. Instead, urethral tone may depend on an interaction between L-type Ca2+ current and a large Ca(2+)-activated Cl- current. Activation of Cl- channels (perhaps by spontaneous release of Ca2+ from intracellular stores) would depolarize the membrane potential to within the 'window current' range for L-type Ca2+ channels and result in Ca2+ influx and contraction. This process may be maintained for a time by positive feedback whereby the influx of Ca2+ continues to activate the Cl channels. PMID- 10540790 TI - Why we need poets in palliative care. PMID- 10540791 TI - Cancer pain management in home hospice settings: a comparison of primary care and oncologic physicians. AB - We compare the effectiveness of terminal cancer pain management delivered by primary care physicians with that of specialists in home-based hospice settings. Each visit record for 223 outpatients in three hospice programs was abstracted for physician type, patient age, medication usage, level of pain reported, cancer type, and metastatic status. Thirteen percent of patients reported no pain at any visit and 19% reported pain at all visits; half of the patients reported pain at two thirds of their visits. No difference was found in the presence or absence of pain between primary care and oncologic patients. When available, the level of pain reported (0-10 scale) was statistically (p < 0.01) but not clinically different between physician groups; average pain rating for primary care patients was 3.7 while the mean pain rating for oncologic patients was 3.1. The reported pain level varied significantly among facilities, as did physician mix. Multivariate analysis revealed that program and an interaction term between program and physician type, but not physician type independently, explained a significant amount of variation in pain level. Overall, reported pain remained higher than optimal. Research elsewhere has shown that application of the World Health Organization (WHO) cancer pain management guidelines can control 70%-90% of cancer pain. Strategies for implementing pain guidelines that emphasize a systems approach may be effective. PMID- 10540792 TI - Temporal distribution of deaths in cancer patients admitted to a palliative care unit. AB - The timing of death has received much attention, particularly in the area of sudden cardiac death. Many studies have demonstrated that sudden cardiac death and other sudden deaths follow a circadian pattern. Deaths have also been reported to vary around dates that are especially meaningful to patients and families. To test these reported observations in a cancer palliative care population, we reviewed the date and time of death of 626 consecutive patients admitted to the palliative care unit of a western Canadian hospital. All patients were adults with advanced metastatic or locally recurrent cancer. A circadian distribution in the time of deaths was observed; 225 deaths occurred between 20:00 and 06:00 (261 deaths expected) versus 401 deaths between 06:00 and 20:00 (365 deaths expected) (p = 0.0037). The distribution of deaths did not change significantly according to day of the week or month of the year. Based on patient birthday, 41 deaths occurred during the three weeks before a birthday (33.5 deaths expected) versus 26 deaths during the three weeks after a birthday (33.5 deaths expected) (p = 0.067). There appear to be fewer deaths during the evening and night; there does not appear to be a decline in deaths before the patient's birthday; and any temporal distribution of deaths in this population appears to be minimal. PMID- 10540794 TI - Do hospice clinicians sedate patients intending to hasten death? PMID- 10540793 TI - The Edmonton Symptom Assessment Scale (ESAS) as an audit tool. AB - To ensure quality of care, palliative care programs need to document the effectiveness of their relief of physical and psychological distress. The Edmonton Symptom Assessment Scale (ESAS) is a validated, reliable instrument developed to measure 9 different symptoms in palliative care patients. To see if symptom management could be compared across institutions, we first reviewed the charts of 188 successive admissions to the palliative care unit at St. Boniface General Hospital, Winnipeg, Manitoba. Our study showed that the ESAS is a useful audit tool for assessing patterns of palliative symptom control that allows for institutional comparisons. Procedures that ensure completeness of data collection remain to be developed. PMID- 10540795 TI - Palliative care for patients with hematological malignancies--if not, why not? PMID- 10540796 TI - Family surrogacy and cancer disclosure: physician-family negotiation of an ethical dilemma in Japan. PMID- 10540797 TI - Thirty years' experience with cancer and non-cancer patients in palliative home care. PMID- 10540798 TI - The potential dangers of complementary therapy use in a patient with cancer. PMID- 10540799 TI - Case report: is this patient palliative? Taube AW, Bruera E. J Palliat Care 1999: 15(1): 53-55. PMID- 10540800 TI - [Inaccurate or insignificant?]. PMID- 10540802 TI - [Knee endoprosthesis. Indications--principles of surgery--follow-up treatment]. PMID- 10540801 TI - [Topical pharmacology. COX-2 inhibitors]. PMID- 10540803 TI - [Heart transplantation. Current state and future alternatives]. PMID- 10540804 TI - [Placebo surgery for clinical studies?]. PMID- 10540805 TI - The role of interference in memory span. AB - In two experiments, we investigated the possibility that susceptibility to proactive interference (PI) affects performance on memory span measures. We tested both younger and older adults (older adults were tested because of the suggestion that they are differentially susceptible to PI). We used two different span measures and manipulated testing procedures to reduce PI for these tasks. For older adults, span estimates increased with each PI-reducing manipulation; for younger adults, scores increased when multiple PI manipulations were combined or when PI-reducing manipulations were used in paradigms in which within-task PI was especially high. The findings suggest that PI critically influences span performance. We consider the possibility that interference-proneness may influence cognitive behaviors previously thought to be governed by capacity. PMID- 10540806 TI - Sentence interference in the Stroop task. AB - In two experiments on Stroop interference, we examined whether sentences can be processed without the intention of the reader. Participants named the ink colors in which words in sentences were printed, and the ink colors in which the same words, randomly arranged, were printed. In Experiment 1, sentences yielded longer response times (RTs) and more errors than did nonsentences, but only when they included words that were highly relevant to the color-naming task (i.e., color and color-related words). In Experiment 2, sentences yielded more errors than did nonsentences, and sentences in which the color words matched the set of ink colors yielded longer RTs than did nonsentences. The results indicate that sentence processing can be obligatory when the component words are highly relevant to the task. PMID- 10540807 TI - Working memory and intrusions of irrelevant information in a group of specific poor problem solvers. AB - An important body of evidence has shown that reading comprehension ability is related to working memory and, in particular, to the success in Daneman and Carpenter's (1980) reading and listening span test. This research tested a similar hypothesis for arithmetic word problems, since, in order to maintain and process the information, they require working memory processes. A group of children possessing average vocabulary but poor arithmetic problem-solving skills was compared with a group of good problem solvers, matched for vocabulary, age, and socioeconomic status. Poor problem solvers presented lower recall and a greater number of intrusion errors in a series of tasks testing working memory and memory for problems. The results obtained over a series of six experimental phases, conducted during a 2-school-year period, offer evidence in favor of the hypotheses that groups of poor problem solvers may have poor performance in a working memory test requiring inhibition of irrelevant information (Hypothesis 1), but not in other short-term memory tests (Hypothesis 2), that this difficulty is related to poor recall of critical information and greater recall of to-be inhibited information (Hypothesis 3), that poor problem solvers also have difficulty in remembering only relevant information included in arithmetic word problems (Hypothesis 4) despite the fact that they are able to identify relevant information (Hypothesis 5). The results show that problem-solving ability is related to the ability of reducing the memory accessibility of nontarget and irrelevant information. PMID- 10540808 TI - Division by multiplication. AB - In two experiments, item-specific transfer was examined in simple multiplication and division with prime and probe problems separated by four to six trials. As was predicted by Rickard and Bourne's (1996) identical-elements model, response time (RT) savings were larger with identical (e.g., prime 63 divided by 7, probe 63 divided by 7) than with inverted (63 divided by 9 and 63 divided by 7) division problems, whereas identical (7 x 9 and 7 x 9) and inverted (9 x 7 and 7 x 9) multiplication problems produced equivalent transfer. Nonetheless, there was statistically significant transfer between inverted division problems. Furthermore, RT savings in the multiplication-to-division transfer conditions (e.g., prime 7 x 9, probe 63 divided by 7) indicated that multiplication mediated large-number division problems. These latter effects are not predicted by the identical-elements model but may be reconciled with the model by distinguishing associative transfer (facilitation owing to strengthening of a common problem node in memory) from mediated transfer (facilitation owing to mediation by a strengthened, related problem). Skilled adults can exploit the conceptual correspondences between multiplication and division facts in a highly efficient way to facilitate performance. PMID- 10540809 TI - More on the relation between division and multiplication in simple arithmetic: evidence for mediation of division solutions via multiplication. AB - Adults (N = 32) solved simple multiplication (e.g., 8 x 7) and corresponding division problems (e.g., 56/8). Self-reports of solution processes were given by half of the participants. Latency patterns and error rates were closely related across operations and were similar in self-report and no-report conditions. Solution of division problems, however, facilitated solution of multiplication problems more than the reverse. On large division problems, participants reported that they "recast" problems as multiplication (e.g., 56/8 as 8 x = 56). These results support the hypothesis that multiplication and division are stored in separate mental representations but that solution of difficult division problems sometimes involves access to multiplication. PMID- 10540810 TI - The organization of verbs of knowing: evidence for cultural commonality and variation in theory of mind. AB - Cross-cultural commonality and variation in folk theories of knowing were studied by examining the organization of verbs of knowing in German and Japanese adults. German and Japanese adults performed one of two tasks: a similarity judgment task and an attribute rating task. Organizational structure was assessed for the similarity judgment task using multidimensional scaling and additive similarity tree analyses. The attribute rating task was used to describe the characteristics that organized the dimensions and clusters emerging from the scaling solutions. The folk theory of mind displayed was an information processing model with constructive components, although the constructive aspects were more salient for the Germans than for the Japanese. PMID- 10540811 TI - Another word on parsing relative clauses: eyetracking evidence from Spanish and English. AB - Ambiguity as to what the relative clause modifies in phrases such as Someone shot the maid of the actress who was divorced/Alguien disparo contra la criada de la actriz que estaba divorciada tends to be resolved differently in different languages (and in different forms of complex noun phrases). In English, there is a weak but seldom significant tendency for the relative clause to be taken as modifying the second noun phrase, the actress, but in Spanish, several researchers have found a significant preference for the relative clause's modifying the first noun phrase, la criada. The present experiments compare Spanish and English readers' eye movements while reading exactly comparable sentences in their native languages and find a significant reading time advantage in Spanish when it is forced to modify the first noun phrase, but in English when the relative clause is forced to modify the second noun phrase. Theoretical implications of the findings for previous explanations of the phenomenon are discussed. PMID- 10540812 TI - On-line predictive inferences in reading: processing time during versus after the priming context. AB - Prior research suggests that predictive inferences take time to construct on line. The present study examines the relative contribution of time available during and after reading an inducing context. In six experiments, we manipulated the presentation rate of the context and the delay between the onset of the last word in the context and a target word. A predicting, or a control, sentence context was followed by a target word, which represented the predicted event or an unlikely event. The results indicated that increasing the time available during reading of the context improved comprehension of explicit information, but it did not affect construction of inferences. In contrast, increasing the delay at the end of the context did not affect explicit comprehension, but it enhanced the probability of inferences, as revealed by shorter latencies in naming the predictable target word after the inducing context, relative to the control context. These findings show that readers defer making predictive inferences until 1 sec after the sentence context has been read, regardless of the time available when they are processing the context. PMID- 10540813 TI - Justification effects on the judgment of analogy. AB - Many of us share a strong intuition that justification forces us to better understand the situations we face. And there is substantial evidence indicating that this is often the case. However, there is a growing body of research showing that, under certain circumstances, explanation and justification can impair performance on a variety of cognitive tasks. In the present research, the effects of justification on judgment of the soundness of analogies were examined. Subjects judged the quality of the match between pairs of stories with varying degrees of superficial and analogical similarity. Experimental subjects either provided reasons for their judgments or wrote recollections of the target stimuli. These subjects rated the match between stimulus pairs as more sound than did control subjects. Also, providing reasons led to poorer discrimination between superficially similar aspects of the stimuli and analogous aspects. Explanations of these findings are proposed, and implications for problem solving and confidence judgment are discussed. PMID- 10540814 TI - What is learned in knowledge-related categories? Evidence from typicality and feature frequency judgments. AB - When a category's features are tied together by integrative knowledge, subjects learn the category faster than when the features are not directly related. What do subjects learn about the category in such circumstances? Some research has suggested that the subjects can use the knowledge itself in performing the category learning task and, thus, do not learn the details of the category's features. Two experiments investigated this hypothesis by collecting feature frequency estimates after category learning. The results showed that integrative knowledge about a category did not decrease subjects' sensitivity to feature frequency--if anything, knowledge improved it. A third experiment found that integrative knowledge did reduce sensitivity to feature frequency in typicality ratings. The results suggest that knowledge does not inhibit the learning of detailed category information, though it may replace its use in some tasks. PMID- 10540815 TI - Learning categories by touch: on the development of holistic and analytic processing. AB - The development of holistic and analytic processing often studied in the visual domain was investigated in haptics. Children 3 to 9 years of age and adults had to categorize haptic exemplars that varied systematically in four attributes (size, shape, surface texture, and weight). The subjects could learn the categories either analytically--that is, by focusing on a single attribute--or holistically--that is, in terms of overall similarity. The data show that even the youngest children learned the haptic categories far more often in an analytic mode than in a holistic mode. Nevertheless, an age trend was observed, referring to the attributes that the analytic learners used for their categorization. The children preferred substance-related attributes, especially surface texture, whereas the adults preferred structure-related attributes, especially shape. Thus, it appears that analytic and/or holistic processing in category learning develops in a similar manner in the visual and haptic domains. PMID- 10540816 TI - Orientation-specific effects in picture matching and naming. AB - In two experiments, subjects made timed decisions about the second of two sequentially presented rotated drawings of objects. When the two objects were physically identical, response times to decide whether the two drawings depicted the same object varied as a function of the shortest distance between the orientation of the second drawing and either the orientation of the previous drawing or the upright. This was found for both short (250-msec) and long (2-sec) interstimulus-intervals. The result was also obtained when subjects named the second drawing after deciding whether the first drawing faced left or right. Following repeated experience with the drawings in the left/right task over four blocks of trials, time to name the second drawing in the same-object sequences was independent of orientation. These results suggest that, initially, object- and orientation-specific representations can be formed following a single presentation of a rotated object and subsequently used to identify drawings of the same object at either the same or different orientations. Alignment of the second drawing with either the canonical representation or the new representation at the previous orientation is achieved by normalization through the shortest path. Following experience with the objects, orientation-invariant representations are formed. PMID- 10540817 TI - An examination of the effects of adult age on explicit and implicit learning of figural sequences. AB - Memory for previously learned figural sequences and item-to-item covariations within figural sequences was examined under explicit and implicit instructional conditions in three age groups: young adults (17-23 years); middle-aged adults (35-45 years); and older adults (55-65 years). In Phase 1 of the experiment, the acquisition phase, half the subjects in each age group learned sequences of three to eight items in which the item-to-item changes conformed to an artificial grammar, and the other half of the subjects in each age group learned strings in which the item-to-item changes were nongrammatical. In Phase 2, the implicit/explicit test phase, subjects made forced-choice judgments about parts of the strings that they learned in Phase 1, under either explicit or implicit instructions. Analyses of Phase 2 data revealed that subjects in both instructional conditions used item-to-item covariations in making decisions about grammatical strings. However, use of previously learned covariations as well as the number of correct judgments about previously learned strings was greater in the explicit condition than in the implicit condition. An age-related deficit was found for explicit recognition of grammar-following sequences. PMID- 10540818 TI - Long-term memory for temporal structure: evidence form the identification of well known and novel songs. AB - In three experiments, long-term memory for temporal structure was examined by having participants identify both well-known (e.g., "I've Been Working on the Railroad") and novel songs. The target songs were subjected to a number of rhythmic alterations, to assess the importance of four critical features of identification performance. The four critical features were meter, phrasing, rhythmic contour (ordinal scaling of note durations), and the ratio of successive durations. In contrast with previous work, the unaltered version of each song was identified significantly better than any altered version. This indicates that rhythm is stored in long-term memory. Furthermore, the results demonstrated that all four critical features play a role in the identification of songs. These results held for both well-known and novel tunes. PMID- 10540819 TI - Levels-of-processing effects in subject-performed tasks. AB - In memory for subject-performed tasks (SPTs), subjects encode a list of simple action phrases (e.g., thumb through a book, knock at the door) by performing these actions during learning. In three experiments, we investigated the size of the levels-of-processing effects in SPTs as compared with those in standard verbal learning tasks (VTs). Subjects under SPT and VT conditions learned lists of action phrases in a surface or a conceptual orienting task. Under both encoding conditions, the subjects recalled fewer items with surface orienting tasks than with conceptual orienting tasks, but the levels-of-processing effects were strongly reduced in the SPT condition. In the SPT condition, items that were encoded in a surface orienting task were still substantially recalled. The items were recalled almost as well as the conceptually encoded items in the VT condition. The distinct reduction of the levels-of-processing effect is caused by the fact that, in SPT encoding even with a verbal surface orienting task, subjects process conceptual information in order to perform the denoted action. We attribute the small conceptual advantage, which remains with SPT despite the conceptual processing for performing, to the fact that items are not as well integrated into memory as they are when conceptual processing is focused on the action component, rather than on the semantic contexts. This lower integration reduces the accessibility of items in the verbal surface task, even with SPT encoding. PMID- 10540820 TI - Positional information in short-term memory: relative or absolute? AB - Evidence suggests that short-term memory for serial order includes information about the positions of items in a sequence. This information is necessary to explain why substitution errors between sequences tend to maintain their position within a sequence. Previous demonstrations of such errors, however, have always used sequences of equal length. With sequences of different length, both transpositions between groups (Experiment 1) and intrusions between trials (Experiment 2) are shown to respect position relative to the end as well as to the start of a sequence. These results support models in which position is coded by start and end markers, but not models in which position is coded in temporal or absolute terms. Possible interpretations of an end marker are discussed. PMID- 10540821 TI - Hypermnesia: the role of multiple retrieval cues. AB - We demonstrate that encoding multiple cues enhances hypermnesia. College students were presented with 36 (Experiment 1) or 60 (Experiments 2 and 3) sets of words and were asked to encode the sets under single- or multiple-cue conditions. In the single-cue conditions, each set consisted of a cue and a target. In the multiple-cue conditions, each set consisted of three cues and a target. Following the presentation of the word sets, the participants received either three cued recall tests (Experiments 1 and 2) or three free recall tests (Experiment 3). With this manipulation, we observed greater hypermnesia in the multiple-cue conditions than in the single-cue conditions. Furthermore, the greater hypermnesic recall resulted from increased reminiscence rather than reduced intertest forgetting. The present findings support the hypothesis that the availability of multiple retrieval cues plays an important role in hypermnesia. PMID- 10540822 TI - [Infections of the mouth mucosa (I). HIV infection--an epidemiological, clinical and therapeutic update]. AB - Infections of the oral mucosa have become important with respect to acquired immunodeficiency syndrome (AIDS), particularly as opportunistic infections. In the first part of this overview the epidemiologic, clinical and therapeutical aspects of the HIV infection are addressed. By the end of 1998, WHO had registered 2 million cases of AIDS globally and 33.4 million HIV infections were estimated. Every day 16,000 new infections occur worldwide. Sub-Saharan Africa is the region which has been affected most. By the end of 1998, 18,000 cases of AIDS were registered in Germany, with an estimated 50,000-60,000 HIV infections. The majority of new infections was registered among homosexual men; however, heterosexual transmissions are increasing. Antiretroviral combination therapies had an impact on the clinical course of HIV infection, affecting both mortality and morbidity. The effectiveness of antiretroviral therapy is monitored by determination of the viral load in plasma and CD4(+)-cell counts. Triple therapy resulted in longer survival and a reduction in opportunistic infections. Oral opportunistic infections are also markedly reduced after the introduction of combination therapies. Combination therapies require stringent compliance and are often accompanied by serious side effects. PMID- 10540823 TI - [Clinical studies on the pathophysiology of odontogenic abscesses]. AB - In 26 patients with abscesses in the maxillofacial area, the electrolyte concentrations, pH and osmotic and hydrostatic pressures of the pus fluid were measured and calculated. The main cations identified were sodium (134 +/- 38 mmol/l) and potassium (37 +/- 16 mmol/l) and as anions chloride (183 +/- 46 mmol/l) and bicarbonate (10 +/- 4 mmol/l). The pH value of the pus liquid was 6.164 +/- 0.233. The calculated mean osmotic pressure of the pus liquid was 7910 +/- 1455 mm Hg, whereas the measured physical pressure inside the abscess was 49 +/- 13 mm Hg. Both pressure types show time-dependent pressure curves. With time, the real pressure inside the abscess cavity increases, whereas the osmotic pressure decreases. There was no relationship between the two pressure types and the different species of microorganisms responsible for the inflammation. The results of the study reveal that abscesses can be regarded as osmotically active systems, and the mechanism by which the abscess is formed might be as follows. After penetration of virulent microorganisms into the tissue space, the area of acute inflammation is walled off by the collection of inflammatory cells. Destruction of tissue by products of the polymorphonuclear leucozytes takes place and results in liquefactive necrosis and a hypertonic abscess cavity. The inwards directed flow of tissue fluids into the cavity via the abscess membrane causes volume expansion and generates pressure, two facts that can explain the swelling dynamics and typical symptoms of abscesses in the maxillofacial area. PMID- 10540824 TI - [Diagnostic and therapeutic concepts of canine fossa abscess. Evaluation of a multicenter study of 55 German-speaking departments of oromaxillofacial surgery]. AB - A possible complication of fossa canina infections is reactive thrombosis of the vena angularis, which can lead to cavernous sinus phlebothrombosis. According to the literature there are different opinions about the treatment protocol of fossa canina abscesses. Therefore a multicentre study was undertaken in which 55 departments of oral surgery were interviewed using a standardized questionnaire about their procedures. The basis of diagnosis consists of clinical examination (100%), antibiogram (77.1%) and radiography (97.1%). A total of 28.6% use additional imaging procedures. Sonography is used by 20.0% and is found to be helpful in the diagnosis of vena angularis thrombosis. The painful palpation of the medial angle of the eye is of major diagnostic importance (100%) although anatomic variations of the vena angularis have to be considered. Extraoral incisions are not used in most cases (25.7% in exceptional cases) as well as ligations of the vena angularis in case of thrombosis (8.6% in exceptional cases). There is an agreement in the use of antibiotics (elective administration in 14.3%, obligatory administration in 85.7%); 77.1% don't use heparin to treat thrombosis of the vena angularis. In heparin therapy risks such as bleeding complications and thrombocytopenia have to be considered. PMID- 10540825 TI - [Cowden syndrome (multiple hamartoma syndrome). Role of the dentist in early detection]. AB - First lesions of Cowden syndrome appear in the oral cavity and on the skin. Malignant transformation is a late, common event in thyroid and breast. The early diagnosis of Cowden disease prior to the development of internal malignancy, particularly of the breast and the thyroid gland, is very important. We emphasize that the dentist may be the first health care professional who recognizes the syndrome, and this is a crucial step in the prevention and cure of the predictable malignancy. This article presents a typical case of Cowden disease. PMID- 10540827 TI - [Cytokeratin and p53 expression of odontogenic cysts]. AB - The histopathologic diagnosis of odontogenic cysts is based mainly on the morphological nature of the epithelial lining of cysts and their origin. We used the international histologic classification set up by the World Health Organisation in 1992. The aim of this study was to investigate the differentiation of various types of cyst using an immunohistochemical technique rather than by conventional morphological assessment. A standard immunocytochemical method (APAAP method), applying anticytokeratin monoclonal antibodies and a p53 antibody, was used for the diagnosis of odontogenic cysts. A total of 57 jaw cysts were diagnosed according to clinical, radiological and pathological criteria as radicular cysts (20), dentigerous cysts (20) and keratocysts (17). The results proved that cyst type can be distinguished by the pattern of staining using the monoclonal antibodies CK7, CK19, CK20 for cytokeratins and the clone DO-7 for the p53 protein. Staining with the monoclonal antibodies CK7 and CK20 did not distinguish type. CK19 was not detected in keratocysts and p53 was only expressed in keratocysts. This may prove to be diagnostically useful for the more precise distinction between different cyst types. PMID- 10540826 TI - [Experimental chemotherapy of xenotransplanted oral squamous epithelial carcinoma: effectiveness of docetaxel (taxotere) in the nude mouse model]. AB - Originating from plants, the taxoids, in particular docetaxel (Taxotere), represent progress in antitumoral chemotherapy. In addition to their use as palliative treatment they have also proved to be increasingly important for adjuvant and neoadjuvant treatment of oral squamous cell carcinoma (SCC). In the present nude mouse model the efficacy of chemotherapeutic agents against exemplary oral SCC were examined using cell line HNSCC 001. Typical biological properties such as expression of serum tumor markers and treatment-related alteration were reviewed. Intraperitoneal administration of 30 mg docetaxel per kilogram body weight at a time resulted in significant growth inhibition (P < 0.001), however without complete remission. Mean values of relative tumor volume ranged from 95% to 131% in the treated groups as compared to 311% in animals without treatment. Application more frequently than weekly did not result in a significant increase in antitumor activity. From the present experimental study no final conclusion can be drawn regarding weekly docetaxel administration mostly used in clinical phase II trials. Except for SCC, for which values correlated well with tumor volume (r = 0.85 without treatment and r = 0.87 with treatment), on the one hand, and a distinct treatment-related decrease, on the other, the tested tumor markers TPA and TPS proved to be less valuable for screening of treatment and follow-up in this murine model. PMID- 10540828 TI - [The Stromeyer hook. Life and work of the man behind the eponym]. AB - Georg Friedrich Stromeyer (1804-1876) is generally known as one of the founders of orthopedics and orthopedic surgery and also made many contributions to modern military medicine. Furthermore, every oral and maxillofacial surgeon in Germany knows him because of the "Stromeyer hook", which is used for elevation of zygomatic arch fractures. This special aspect as well as Stromeyer's biography is presented in this article from the history of medicine. PMID- 10540829 TI - [Biodegradable miniplates (lactosorb) in cranio-osteoplasty--experimental results with the rapidly maturing, juvenile minipig]. AB - As a passive intracranial transmission (PIT- effect) has been described for metallic osteosyntheses materials in the infant growing skull. Thereby the use of resorbable plates and screws might be an alernative fixation device in infant craniofacial surgery. For evaluating the biological behaviour, craniotomies were performed in the frontoorbital region of four infant minipigs, six weeks of age and 6.1 kg of weight. After turning and orthotopical repositioning the full thickness bone graft were fixed with resorbable plates and screws made of LactoSorb on the left side after epiperiosteal, on the right side after subperiosteal preparation. The animals were sacrificed after 3, 6, 9 and 18 months. Histologically, a PIT- effect was detected similar to metallic microplates and screws being significant diminished after epiperiosteal preparation and plate positioning. The biodegradation was not affected by intraosseous translocation. Even in case of intrasinuidal transmission no inflammatory reactions werde observed. No contraindications for the clinical use of this specific PLLA-PGA copolymer could be found when implanted in the rapidly growing craniofacial bone surfaces. PMID- 10540830 TI - [Tracheobronchial perforations--a complication after mouth, maxillary and facial surgery]. AB - After difficult endotracheal intubation in oral and maxillofacial surgery, esophageal or tracheal injuries can cause mediastinal, pericardial or cervicofacial soft tissue emphysema. If a patient has of thoracic pain after general anesthesia, mediastinal emphysema should be taken into consideration because of the possibility of subsequent life-threatening complications. Diagnosis can be established with computed tomography and fiber-endoscopy. We present the diagnostic and therapeutic management of two patients. PMID- 10540831 TI - [Time saving and effective method for temporary intraoperative repositioning of mandibular fractures]. PMID- 10540832 TI - [Interleukin-8 and airway inflammation]. AB - Airway inflammation is a prominent feature of chronic obstructive diseases of the airways, including asthma, bronchiectasis, chronic bronchitis, and diffuse panbronchiolitis. Neutrophils are implicated in the pathogenesis of these diseases. The present review discusses the role of interleukin-8 (IL-8), a neutrophil chemo-attractant, in neutrophil accumulation in the airways, and the mechanisms of inducing IL-8 expression. IL-8 presents in the sputum of patients with inflammatory airway diseases, and accounts in large part for the chemo attractant activity present. Focusing on Pseudomonas aeruginosa as the stimulus, it was discovered that when a supernatant of bacterial culture is introduced into the airways in vivo, bacterial products induce IL-8 expression in surface airway epithelial cells and the recruitment of neutrophils into the airways. The neutrophil chemotactic activity of the airway fluid was inhibited by an IL-8 antibody. The luminal IL-8 concentration increased in response to instillation of bacteria, and an inhibitor of neutrophil recruitment markedly reduced the IL-8 levels. From these results, it was speculated that bacteria-induced neutrophil accumulation in the airways involves a cascade of events, and that early neutrophil recruitment in response to bacteria is due to epithelium-derived IL-8, while the amplification of the response is due, at least in part, to IL-8 induction in the neutrophils themselves. PMID- 10540833 TI - [A study of the long-term effectiveness of an outpatient pulmonary rehabilitation program]. AB - Pulmonary rehabilitation is one of the most important components of comprehensive care for patients with significant disability due to chronic respiratory failure. Because pulmonary rehabilitation has not been popular in Japan, the long-term effectiveness of pulmonary rehabilitation has rarely been reported. We therefore examined the long-term effectiveness of an outpatient pulmonary rehabilitation program for patients with chronic respiratory failure. Our program was composed of a once-a-week introduction program for 2 months and a support program that was continued every 4 weeks as long as possible. Thirty stable patients with chronic respiratory failure were enrolled in the program; 21 patients (COPD: 15, lung complications of tuberculosis: 6) completed the 9-week introduction program and the ensuing 6-month support program. Good compliance with the home training regimen was maintained during the period. The introduction program significantly alleviated dyspnea (Fletcher's grade: 3.3 to 3.0, p < 0.01) and improved the data for activity (Spector's score: 5.3 to 5.8, p < 0.01) and 6-minute walking distance (319 to 384 m, p < 0.01). These benefits were sustained during the 6 month support program. We concluded that outpatient pulmonary rehabilitation can alleviate dyspnea and improve the activity and exercise tolerance of patients with chronic respiratory failure, and that the effectiveness of training can be well maintained with a minimal support program. PMID- 10540834 TI - [Continuous oximetry monitoring in patients undergoing home oxygen therapy for chronic respiratory failure]. AB - We investigated the clinical usefulness of continuous nocturnal oxygen saturation monitoring in patients undergoing home oxygen therapy (HOT). The subjects were 11 patients with chronic respiratory disease in the process of healing from acute exacerbation. None were mechanically ventilated. Each subject underwent full overnight oximetry. One patient was excluded from further investigation because of periodic desaturation suggestive of sleep apnea. The remaining 10 subjects included 5 patients with sequelae of pulmonary tuberculosis, 2 with diffuse panbronchiolitis, 1 with chronic pulmonary emphysema, 1 with chronic bronchitis, and 1 with kyphoscoliosis. All underwent full overnight and 30 min daytime oximetry monitoring for 23.7 +/- 7.4 (mean +/- SD) consecutive days. Daytime oximetry was performed when subjects were awake and resting in supine position. Mean nocturnal oxygen saturation (NmSpO2) and mean daytime oxygen saturation (DmSpO2) were calculated from data obtained from 0:00 through 5:00 hrs and from data obtained during a stable 10 min daytime period, respectively. The difference between NmSpO2 and DmSpO2 (delta SpO2), the percentage of total sleep time with SpO2 < or = 90% (DST 90) and nocturnal lowest oxygen saturation (NLSpO2) were calculated once daily for each subject. There were significant (p < 0.05) correlations between NmSpO2 and NLSpO2, between NmSpO2 and DST 90, and between NLSpO2 and DST 90 in all subjects. However, significant (p < 0.05) correlations between NmSpO2 and DmSpO2 were observed in only 6. During acute exacerbation, NmSpO2 was lower than DmSpO2, and delta SpO2 increased. Conversely, with the amelioration of acute symptoms, delta SpO2 decreased and NmSpO2 was higher than DmSpO2. There was a significant (p < 0.05) reverse correlation between NmSpO2 and delta SpO2 in 9 subjects. We concluded that monitoring nocturnal oxygen saturation is clinically useful to assessments of oxygenation status in patients undergoing HOT, and that it may assist the early diagnosis of acute exacerbation of respiratory failure. PMID- 10540835 TI - [Relationship between diabetes mellitus-associated obstructive sleep apnea syndrome and hyperinsulinemia]. AB - The purpose of this study was to examine the prevalence of sleep disordered breathing and background factors, especially hyperinsulinemia, in diabetic patients. The subjects were 70 patients randomly selected from 143 noninsulin dependent diabetic patients hospitalized for educational purposes. Obstructive sleep apnea syndrome (OSAS) was diagnosed in 13 subjects, equivalent to a prevalence of 18.6%. The mean +/- S.D. immunoreactive insulin (IRI) values for 11 of the 13 OSAS patients (excluding insulin-treated patients) and for 49 non-OSAS patients were 10.7 +/- 6.8 microU/ml and 5.8 +/- 3.5 microU/ml, respectively. The value for the OSAS group was higher than that for the non-OSAS group (p = 0.04). However, a positive correlation between body mass index (BMI) and IRI was observed in both the OSAS (Y = 1.148 X - 20.006, r = 0.834, p = 0.001) and non OSAS (Y = 0.466 X - 5.820, r = 0.524, p = 0.0001) groups. Multivariate analysis demonstrated that the presence of OSAS had a statistically significant influence on IRI (p = 0.001), but not on BMI (p = 0.391). These findings suggest that hyperinsulinemia may be exacerbated in diabetic patients with OSAS regardless of BMI. PMID- 10540836 TI - [A case of common variable immunodeficiency that responded to long-term erythromycin chemotherapy]. AB - A 32-year-old woman with common variable immunodeficiency (CVID) accompanied by sinopulmonary infection was evaluated for purulent sputum, cough, and nasal obstruction that did not respond to regular intravenous immunoglobin (IVIG) infusion. Chest X-ray films revealed bronchiectasis affecting both lung bases, and a bacteriological examination of sputum was positive for Pseudomonas aeruginosa. Long-term chemotherapy with erythromycin (EM) was started, and the patient's respiratory symptoms gradually subsided. Sinopulmonary infection is the dominant clinical complication in patients with CVID. This case suggested that long-term EM chemotherapy is useful for the treatment of IVIG-refractory sinopulmonary infection associated with CVID. PMID- 10540837 TI - [Interstitial pneumonia induced by the inhalation of hard metal]. AB - A 51-year-old man visited Okayama Rousai Hospital with the chief complaints of dyspnea and emaciation. His occupational history included 23 years as a hard metal polisher for a shipyard. Physical examination disclosed digital clubbing and fine crackles audible in the inferior posterior lung fields. Laboratory examination revealed hypoxemia and a remarkably reduced vital capacity of the lungs. Chest x-ray films and computed tomograms disclosed interstitial pneumonia predominantly in the upper lung lobes. Lung fibrosis progressed rapidly, and the patient died of exacerbation of chronic respiratory failure 2 years after his first visit to our hospital. The histopathologic findings from tissue specimens obtained by open lung biopsy and necropsy revealed mixed patterns of atypical and usual interstitial pneumonia, but no giant cell interstitial pneumonia. X-ray analysis detected tungsten in the lung tissue and mediastinal lymph nodes, but no cobalt was found. The interstitial pneumonia observed in this patient was thought to be induced by the occupational inhalation of hard metal. PMID- 10540838 TI - [Paragonimiasis Miyazakii with variable X-ray shadows]. AB - A 77-year-old woman was admitted to our hospital with hemoptysis and weight loss. She had eaten 15 raw freshwater crabs about 5 months before the onset of her clinical symptoms. Chest X-ray films obtained on the first admission showed left pleural effusion. After 1 week of chemotherapy with SBTPC, the pleural effusion disappeared. Two months later, the patient was re-admitted with recurrent hemoptysis. Chest X-ray films showed a solitary nodular lesion in the right lung. Eosinophilia and increased serum IgE levels were detected. The solitary nodular lesion moved from the middle to upper field of the right lung during the patient's 3-week stay in the hospital. Serologic tests yielded a conclusive diagnosis of Paragonimiasis miyazakii infection. Praziquantel administration relieved the patient's symptoms. PMID- 10540839 TI - [Constriction of liver tissue herniated through a diaphragmatic defect, simulating a pulmonary tumor]. AB - Defects in the dome of the right diaphragm are a rarity among diaphragmatic abnormalities. Here we report the case of an elderly man who presented with an intrathoracic nodular opacity adjacent to the right diaphragm on chest radiographs. A radionuclide liver scan and maximum intensity projection (MIP) images of a computed tomographic (CT) scan together yielded a diagnosis of diaphragmatic defect with herniation of liver tissue. The herniated liver was so constricted that the angle made by the nodule and diaphragm became acute, resulting in close resemblance to a pulmonary tumor. This unusual configuration of herniated liver tissue was reviewed and the cause of constriction and diagnostic procedure were discussed. To avoid unnecessary examinations, herniation of liver tissue should be included in the differential diagnosis of intrathoracic nodular opacities, particularly when they are in contact with the diaphragm. PMID- 10540840 TI - [Thymic cyst with elevated SLX levels in serum and cystic fluid]. AB - A 24-year-old man was admitted to our hospital in June 1996 with complaints of anterior chest discomfort. Chest X-ray films on admission showed an abnormal mediastinal shadow with well-defined margin. Chest X-ray examinations about 6 weeks earlier had not detected any abnormalities. Laboratory tests on admission showed a high serum concentration of Siaryl Lewis X-i antigen (SLX). A computed tomographic scan of the chest showed a large (6 x 6 x 12 cm) homogeneous mass in the right anterior mediastinum. The mass was removed completely and histologically diagnosed as a thymic cyst. Biochemical analysis of fluid from the cyst revealed remarkably high levels of SLX, CA 19-9, and CEA. In immunohistochemical studies, epithelial cells from the cystic walls stained positive for SLX, CA 19-9, and CEA. After the operation, the level of serum SLX returned almost to normal. PMID- 10540841 TI - [An autopsy case of lung cancer accompanied by pneumoperitoneum]. AB - We report a case of pneumoperitoneum at the end stage of lung cancer. The patient was an 81-year-old man who experienced only mild abdominal pain and fullness. The pneumoperitoneum spontaneously disappeared. Ruptured viscera and peritonitis were not observed at autopsy. We speculated that air had moved from the lung cancer lesion to the mediastinum, and then through the retroperitoneal space into the peritoneal space. When pneumoperitoneum is observed, lung cancer and other intrathoracic diseases should be suspected as causes. Also, it may be necessary to carefully examine the patient for fever, general appearance, abdominal findings, and laboratory findings, including white blood cell count and C reactive protein level, and on that basis differentiate from ruptured viscus, which requires an emergency laparotomy. PMID- 10540842 TI - [Pulmonary thromboembolism accompanied by abnormal plasminogen]. AB - A 38-year-old man was admitted to our hospital because of sudden chest pain and bloody sputum. Lung perfusion scintigraphy disclosed segmental defects in both lungs. An enhanced thin-section computed tomographic scan of the chest showed a low-density area in the right main pulmonary artery. These findings yielded a diagnosis of pulmonary thromboembolism. Serum plasminogen activity was low, not only in the patient but in his elder brother and daughter. Gene analysis revealed a point mutation at exon 15 of the plasminogen gene, suggesting abnormal plasminogen. Abnormal plasminogen is more prevalent in Japan than in the USA or Europe, and is usually asymptomatic. Thromboembolism in patients with abnormal plasminogen is very rare. Further studies are needed to elucidate the relationship between plasminogen abnormalities and pulmonary thromboembolism. PMID- 10540843 TI - [Alpha 1-antitrypsin deficiency (Siiyama) with pulmonary emphysema]. AB - A 44-year-old man was admitted to the hospital with dyspnea on exertion. Chest radiographs and pulmonary function tests showed evidence of pulmonary emphysema. Serum alpha 1-antitrypsin (alpha 1-AT) could not be detected by nephelometry, immuno-electrophoresis, or iso-electric focusing. However, allele-specific PCR revealed a genotype homozygotic for an alpha 1-AT deficient variant of the Siiyama allele. An elder sister of the proband was also a homozygous carrier of the Siiyama allele. The amino acid sequence for normal alpha 1-AT variants had been substituted by Arg101-Val213-Glu376 in the proband, demonstrating that the alpha 1-antitrypsin-deficient Siiyama variant in this pedigree was derived from M 1 (Val213). PMID- 10540844 TI - [Primary ciliary dyskinesia treated with living-donor lobar lung transplantation]. AB - We report a case of primary ciliary dyskinesia in which a living-donor lobar lung transplant was performed. A 24-year-old woman with a diagnosis of primary ciliary dyskinesia and bronchiectasis was admitted to Shinshu University Hospital because of persistent dyspnea and pyrexia over a period of 4 months. Although she was given various antibiotics, neutrophilia, elevated plasma C-reactive protein (CRP) levels, and respiratory failure persisted. Chest roentgenograms and computed tomography disclosed severe bronchiectasis and diffuse infiltrative shadows in both lung fields. Pseudomonas aeruginosa was detected in a sputum culture. Although a variety of conventional therapies were administered, the patient's oxygenation progressively deteriorated. She was intubated and assisted by mechanical ventilation. The patient and her family proposed lung transplantation, and we concluded that a living-donor lobar lung transplant would be a suitable treatment for her disease. We transported the patient to Okayama University Hospital by helicopter 10 days after intubation. A living-donor lobar lung transplant was successfully performed with lung tissues donated by the patient's mother and sister for each transplant site. PMID- 10540845 TI - [Thymic enlargement exhibiting remarkable respiration-induced changes in form]. AB - The patient was a 30-year-old woman. During an examination for a painful bruise on her back, an anterior mediastinal mass lesion was detected by computed tomographic (CT) scan. A second CT scan 6 days later showed pronounced expansion of the anterior mediastinal mass shadow. A third CT scan performed 18 days later, however, disclosed that the mass had contracted back to the size observed with the initial CT scan. Respiration-induced changes in size were suspected, and breathing dynamic cine magnetic resonance imaging (MRI) was performed. The MRI findings clearly demonstrated that the anterior mediastinal mass shadow contracted on inspiration and expanded on expiration. The patient was admitted for suspected thymoma, and hyperthyroidism was diagnosed. After her thyroid function normalized, a subtotal thyroidectomy and thymectomy were simultaneously performed. The pathologic diagnosis was thymus enlargement and hyperthyroidism, respectively. Breathing dynamic cine MRI provided extremely valuable films that demonstrated remarkable respiration-induced changes in the shape of the enlarged thymus. PMID- 10540846 TI - [A model for the Japanese health center to meet the needs of health crisis management in the 21st century]. PMID- 10540847 TI - [A review of methodology on community health nursing diagnosis]. AB - PURPOSE: To develop a systematic method and model for community health nursing diagnosis to be used in teaching and in community practices. METHOD: From searching the databases of Medline (from Jan. 1966 to May 1997) and the Japana Centra Revuo Medicina (from Jan. 1987 to Jan. 1997), literature on community diagnosis, community health nursing, diagnosis, assessment and analysis were classified into keywords, purposes, subjects, health problems and methods. RESULTS: 1. As an explanation of the process of nursing diagnosis the community as-partner model (Anderson and McFarlane; 1995) is useful for understanding the target community and the use of the community health nursing diagnosis process. 2. The methodology of the community health nursing diagnosis is based on three strategies of public health diagnosis. The method of interview surveys was strengthened by incorporating the ethnographic method. 3. Several case studies in the partnership between communities and universities in USA were introduced. CONCLUSION: Changes in community health policy require that public health nurses develop specialized and comprehensive practices in their communities. The authors presented the model of the community health nursing diagnosis process and proposed a partnership between communities and universities. The construction of community health nursing diagnosis process in this paper is based on the public health diagnosis framework consisting of three strategies, to which analysis of existing data, a social survey utilized in epidemiological community diagnosis, and free interviewing from ethnographic methods are incorporated. Developing this systematic diagnosis process of facilitates the search for potential or actual community health problems or concerns, the practice of applying data from surveys and the discussion of concrete strategies toward problem solving. It is useful for educational and research processes and in practice in the community. PMID- 10540848 TI - [Multiple chemical sensitivities: case definition, etiology and relations to allergy, poisoning, psychogenic illness etc]. AB - Multiple Chemical Sensitivities (MCS) have been defined as an acquired disorder characterized by recurrent symptoms, referable to multiple organ systems, occurring in response to demonstrable exposure to many chemically unrelated compounds at doses far below those established in the general population to cause harmful effects; no single widely accepted test of physiologic function can be shown to correlate with symptoms (Cullen MR, 1987). The etiology of MCS is hypothesized as a toxicant-induced loss of tolerance to multiple chemicals with subsequent manifestation of multiple-organ symptoms triggered by low-level exposure to such chemicals. The involvement of multiple organs might be attributed to a neurogenic switching mechanism. The final diagnosis of MCS is to rely on provocation of symptoms in a exposure chamber by a double-blind method. Relations of MCS to allergy, poisoning, psychogenic illness, chemical sensitivity, idiopathic environmental intolerances etc. are discussed in terms of case definition and etiology of these disorders. PMID- 10540849 TI - [Development of professional competence in public health nurses]. AB - PURPOSE: Significant changes that are occurring in the community health care system, require that public health nurses who work for local governments to not only provide direct care but also to coordinate health care teams and participate in policy making. The purpose of this study is to investigate the current system of developing professional competence in public health nurses, and to consider ways to improve it. METHOD: The subjects, randomly chosen, were 100 chief public health nurses and 298 staff nurses in Hokkaido. Sixty-four chief nurses, 44 beginner nurses, 87 proficient nurses and 88 expert nurses responded. The data were collected with a self-administered questionnaire which necessitated the subjects to make a self evaluation of their practical competence and circumstances of its development. RESULT: Self evaluated competence, in making accurate assessments of individual needs and initiating direct care, developed with their experiences in the job, and was generally high. However, the questionnaire showed that self evaluation of their work and ability to do theoretical analysis and research was low and did not progress in conjunction with the length of work experience. Policy making experience was limited and policy making competence was evaluated as low, but there was an expectation that this competence would develop in time. Almost all of the respondents expected their professional competence to progress to higher levels. In particular, beginner nurses wanted to gain practical care competence; proficient and expert nurses wanted to develop their powers of theoretical analysis and do more research, while chief nurses were keen to gain competence in the area of policy making. The respondents reported that they sometimes attended academic conferences, but hardly did any research. CONCLUSION: Low self-evaluation of public health nurses reflect a basic immaturity as a profession. They need to establish their profession and to increase their self-evaluated competencies as their careers develop. These findings showed the importance of establishing a system of continuing education that will cultivate competence in various aspects of their job and also motivate self study. These findings also reveal the importance of collaboration between the university as a vehicle for theoretical work and research and the work place as the embodiment of practical application. PMID- 10540850 TI - [Validity of urinary glucose test for diabetes screening in workplace regular medical checkups]. AB - It has been acknowledged that urinary glucose level varies depending on the time after meal which may hence affect validity of urinary glucose test as a screening test for diabetes. However, sample collection time has not been standardized for urinary glucose test in workplace regular medical checkups. In this study, we investigated the effect of timing of urine sampling on results of urinary glucose screening tests, and using data obtained from the 75 g oral glucose tolerance tests (OGTT) evaluated the validity of urinary glucose test in workplace medical checkups. Also, we made a survey among industrial physicians about the timing of sample collection. Between 1991 and 1996, 455 males and 116 females who participated in a 2-day health examination including the OGTT in a hospital in Chiba were used as study subjects. These did not include an additional 22 subjects who had already been diagnosed as having diabetes but received the OGTT. The examinees observed a strict fast after supper the previous evening and urine and blood samples were collected before and 2 hours after glucose intake for the OGTT. Diagnosis of diabetes was made following the criteria of WHO (1985) and screening test results were defined positive when urinary glucose level was equal to or more than 40 mg/dl. Sensitivity and specificity were calculated for diabetes and impaired glucose tolerance separately for male and female. Industrial physicians who participated in a lecture course for qualification were surveyed about the timing of urine sample collection for urinary glucose test in workplace regular medical checkups. For the urine samples collected 2 hours after glucose intake, sensitivity was very high (male 84%, female 100%), in addition to a relatively high specificity (male 71%, female 92%). However the samples collected before glucose intake showed extremely poor sensitivity (male 11%, female 0%). Similar results were obtained when screening was made for impaired glucose tolerance and both diabetes and impaired glucose tolerance combined. The survey among the industrial physician revealed that in the majority (58%) of workplace medical checkups, the urine sample had been collected when examinees were fasting. The largest reason for this was that the urinary glucose test was performed together with upper gastro-intestinal examination or measurement of serum triglyceride both of which require fasting of examinees. The results of the present study showed that urinary glucose tests have been frequently performed at a timing that produces low validity. The procedures of workplace regular medical checkups should be evaluated with the concept of Evidence Based Medicine. The standard of the timing of urine sample collection for urinary glucose test should be carefully examined. PMID- 10540851 TI - [Cross-sectional study of factors related to Achilles bone mineral density measured by an ultrasound system]. AB - A cross-sectional study was conducted to examine the relationship between bone mineral density (BMD) and life-style related factors including exercise and dietary habits in 1016 pre-menopausal women and 856 post-menopausal women in Ishikawa Prefecture, Japan. The achilles BMD in 1,872 women ages between 19 and 85 years were measured from 1995 to 1996 by an ultrasound system. The stiffness index calculated by the Lunar Achilles ultrasound machine was used as the BMD in this analysis. Self-administered questionnaires were used to obtain the following information: medical history, pregnancy, delivery and menstrual history, height at 20-years of age, present number of teeth, fracture history, sports exercise history, food intake frequency, smoking and drinking history, and daily physical activity. Analysis of covariance and multiple regression analysis were performed to evaluate the contribution of life-style related factors to BMD after adjustment for age and BMI (Body mass index) in pre- and post-menopausal women, respectively. Results were as follow: 1) BMD was inversely associated with increasing age in pre- and post-menopausal women. The BMD level of post menopausal women were lower than that of pre-menopausal women in each 5-year age group. The pearson's correlatin coefficient between age and BMD was significant at -0.25 and -0.44 in pre- and post-menopausal women, respectively. 2) Body mass index (BMI) and BMD were positively correlated in pre- and post-menopausal women. 3) In pre-menopausal women, lower BMD was associated with the following factors: age, lower BMI, no history of joining a sports club in junior high school, absence of current regular sports, being inactive in daily life, having joint pains, lower number of remaining teeth and lower dairy product intake. 4) In post menopausal women, lower BMD was associated with the following factors: age, lower BMI, no history of joining a sports club in junior high school, past history of fracture and longer post-menopausal years. Factors associated with lower BMD in this study were regarded as risk factors for future osteoporotic fractures in the elderly, or signs of lower BMD. Therefore, the information of these factors should be employed in health education for the prevention of osteoporosis. Especially, participating in a sports club while in junior high school may be a recommendation for acquiring higher BMD even in the post-menopausal period. PMID- 10540852 TI - [Relationship of dietary habits pattern and body build of parents to child obesity]. AB - Patterns of eating habits were analyzed to elucidate its relationship to the temporal change of body build from childhood through school age in subjects of the Toyama study. Survey questionnaires at the time of entrance to elementary school were used. Subjects were 6,452 (males 3,293, and females 3,159). Subjects were classified into 6 clusters among the males, 8 clusters among females based on the results of cluster analysis of eating habits. The cluster in males that preferred egg, milk, dairy products, fats, fish and shellfish, soybeans, fruits, green yellow vegetables indicated more frequent subjects whose BMI were less that 14. The cluster in girls that preferred fats indicated more frequent subjects whose BMI were more or equal to 18. The ANOVA showed significant relation of parental body build on their children. Even after grouping by parental body build, the cluster based on patterns of eating habits showed different frequencies of obese children. Preference for intake of milk indicated less frequent obese children among the similar parental body build for boys, while preference for intake of fats indicated more frequent obese children among a similar parental body build for girls. In conclusion, the obesity of a school child has a close relationship to parent's body build. However, the temporal changes of obesity were seen among eating habits clusters even if body builds of their parents are the same. It was shown that patterns of eating habit are important in school children's obesity development. PMID- 10540853 TI - [Development of a smoking cessation program during health checkups. Preliminary study to evaluate the usefulness of this program]. AB - Development of a simple and effective smoking cessation program is needed to provide cessation counseling during health checkups. A new cessation program, which consists of brief individual counseling and 4 follow-up telephone calls, was developed based on the stage model for life-style change. This program was performed during health checkups in the town of Nose to evaluate its usefulness. Smoking status questionnaires were completed to assess the smoking habits of subjects and to evaluate their smoking stage before the counseling session. Then, stage-matched cessation counseling was provided using a self-help guide. During the counseling, carbon monoxide measurement of expired air and Health Risk Appraisal feedback were performed to enhance self-perception of smoking. Follow up calls were provided for only those clients who set a quit date during the individual counseling. It was easy to implement this program, and it required between 15 and 20 minutes to conduct. The cessation rate was 19% at 8 months after the health checkups. This result was more effective than data for other programs reported previously. Therefore, this program was effective and could be used at health checkups. This trial had no control group, so further studies are needed to clarify the efficacy and effectiveness of this program. In addition, training courses for health professionals must be developed to disseminate this program into general use. PMID- 10540854 TI - [Intra- and inter-individual variations in diets of the middle-aged and the elderly]. AB - This study was conducted to examine intra- and inter-individual variations in diets of the middle-aged and the elderly (40 years or older, 46 men and 42 women). The coefficients of variations for intakes of nutrients and food groups were computed from four 4-day weighed dietary records performed at 3-month intervals from June 1996. The results were as follows: a) The highest intra individual variation (%) for nutrient intake was observed in retinol (men 293.5, women 283.8) and the lowest in carbohydrate (men 17.7, women 22.1). b) The highest inter-individual variation (%) was found in retinol (58.2) in men, and in carotene (56.7) in women. The lowest inter-individual variation (%) was observed in magnesium (17.0) in men, and in carbohydrate (14.4) in women. c) Nuts and seeds showed the highest intra-individual variation (%) for food group intake (men 291.5, women 391.8), while rice presented the lowest (men 30.5, women 38.9). d) The highest inter-individual variation for food group intake (%) was seen in milk and dairy products (111.7) in men and in alcoholic beverages (162.3) in women. The lowest inter-individual variation was observed in potatoes and starches (20.7) in men and in pulses (26.0) in women. e) The number of days necessary to estimate true average nutrient intake was much longer for such vitamins as retinol and carotene (over 50 days) than for macronutrients (3-5 days) except for fat. More than one year was required to estimate intake of nuts and seeds in both sexes and alcoholic beverages or seaweeds in women, whereas only 9-15 days for intake of rice. In conclusion, energy, protein and carbohydrate can be estimated by short-period dietary recalls or records, since their intra-individual variations were relatively small. On the other hand, many days, were found to be required to estimate usual dietary intake of such vitamins as retinol or carotene and that of each food group except for rice. It would therefore be very difficult to estimate usual intake of these nutrients and food groups by short-period dietary recalls or records. PMID- 10540855 TI - [Lifestyle and health status of homeless people around Shibuya Station, Tokyo]. AB - The number of homeless people in Tokyo is estimated to be 3,200-3,300. While studies on the health status of homeless people, including illness, injury and deaths have been previously reported, most of these reports concern the homeless who resided in housing facilities for the homeless or who admitted to hospitals. We undertook a comparison of lifestyle and health status between homeless people and people who live in houses (as a control group). Health status was also analyzed for differences among homeless people. Subjects were asked by questionnaire regarding their age, the length of being homeless, former and present employment, sleeping condition, food, whether they have friends or not, the amount of smoking per day, and Short-Form-36 Health Survey (SF36). As objective findings, measurement of blood pressure and blood testing were also performed. Fifty-three homeless people, 49 male, 4 female, average age 52, from the areas around Shibuya station and Yoyogi park, were enrolled. While 98% of the homeless people had previous employment, 73% were not working when the study was performed. Compared with control group, the homeless had fewer meals per day, fewer friends, excessive smoking, greater history of gastro-duodenal ulcer and injury, greater limitation due to physical problems, and higher general mental health as measured by SF36. The diastolic blood pressure of the homeless was higher than that of the control. The blood testing showed higher white blood cell counts and platelet counts. It was suggested that changes in the social structures were largely influential in causing life, and that access to health care was limited because of financial and social barriers. Further studies with more samples, survey of social volunteers involved in care of homeless and qualitative data would be necessary to find and develop better support system for the homeless. PMID- 10540856 TI - [A personal computer system providing various graphs of mortality rates]. AB - PURPOSE: We devised observational methods for showing relations of mortality rates to age and period in various ways. And we contrived a personal computer system to realize the methods. The purpose of this report is to evaluate the methods and the system. METHODS AND MATERIALS: We contrived the system which provides some graphs useful for observing age-period-specific mortality rates. The program was written in Visual Basic on a Windows personal computer. The graphs provided by the system are bird's-eye view, contour map and mortality curves. The system is also made to present any graph desired for any specific required time. A smoothing method is also provided with the system to diminish random error, therefore the characteristics of mortality can be grasped easily. The system was applied to the mortality rates of malignant tumor for the last 45 years (1950-1994) in Japan. RESULTS AND CONCLUSION: Examples which show that one could easily observe the trend and structure of mortality rates by using the method were presented. Because the system can present any of the graphs instantly, so we can observe mortality step-by-step; we can seen the mortality graphs as a whole from a bird's eye view, then observe it in detail by a contour map graph, and furthermore look into the point of interest by mortality curves. Thus, the system will be useful to observe mortality rates. PMID- 10540857 TI - [Clinical vitaminology]. AB - This paper deals with overview on clinical vitaminology including as follows: 1. Historical trends of prevalence of vitamin deficiency in Japan. 2. Recent status of number of patient with vitamin deficiency referred to "patients survey" by Ministry of Health and Welfare. 3. Notion and characteristic of vitamin dependency. 4. Types of vitamin deficiency. i.e. a simple type and a tape caused by disturbance of utilization. 5. Vitamin metabolism of hospitalized elderly patients. 6. Pharmacological role of vitamins. PMID- 10540858 TI - [New aspects of vitamin and coenzyme research]. AB - Vitamins are defined as essential organic micronutrients that are not synthesized by mammals. Coenzymes are defined as organic compounds with low molecular weight that are required to show enzyme activities by reversibly binding with their apoenzymes. Most of vitamins and coenzymes show various biofunctions besides their functions as vitamins and coenzymes. Accordingly, it is more appropriate to understand both as effective biofactors. Various retinoid derivatives synthesized bind to retinoid binding proteins to regulate gene expressions and show other biofunctions. Pyridoxal phosphate serves as an inhibitor of catepsins, and regulates the gene expression. Several novel built-in coenzymes such as topaquinone and lysyltyrosylquinone have been demonstrated in mammalian and microbial enzymes. PMID- 10540859 TI - [Basic aspects of vitamin A--chemical structure, physiological functions and metabolism]. AB - Vitamin A research has been facing the third wave which was initiated both by a cloning of nuclear retinoid receptors and therapeutic application of retinoic acid for acute promyelocytic leukemia. The third wave also gives rise to confusion between vitamin A, retinoids, and retinoate analogues. Physiological functions of vitamin A still remain almost unresolved at a molecular level except a visual cycle. Future study may unravel a molecular involvement of vitamin A in sensation such as smelling, hearing and tasting, and a specific role of vitamin A in dopaminergic neuron will be presented in the near future. A refined control mechanism for intracellular level of retinoic acid is also discussed with retinal dehydrogenase II and cytochrome P450 RAI (CYP26). PMID- 10540860 TI - [Vitamin B1]. AB - Vitamin B1 (thiamin), taken-up into cells, is converted to thiamin diphosphate (TDP), and TDP acts as a cofactor for several enzymes involving in carbohydrate metabolism. CoA-dependent oxidative decarboxylation of pyruvate is catalyzed by pyruvate dehydrogenase multienzyme complex (PDC) with NAD+ as an electron acceptor in most organisms involving mammals and higher plants. PDC consists of three component enzymes, one of which is pyruvate dehydrogenase (lipoamide) which contains TDP as a prosthetic group. Similar multienzyme complex for 2 oxoglutarate or branched chain 2-oxoacids is also found in mammals. In anaerobic bacteria, archaebacteria and anaerobic protozoa, pyruvate:ferredoxin oxidoreductase (PFOR) functions for the oxidative decarboxylation of pyruvate with ferredoxin in place of NAD+. PFOR contains TDP as a cofactor; however its structure is quite different from PDC and 1-3[4Fe-4S] clusters are involved as redox centers. Pyruvate:NADP+ oxidoreductase (PNOR), which catalyzes the oxidative decarboxylation of pyruvate with NADP+ as an electron acceptor, occurs in mitochondria of Euglena gracilis, a protist containing chloroplasts. PNOR consists of two functional domains, one of which contains TDP and 3[4Fe-4S] clusters and resembles PFOR. Another domain involves FMN and FAD as redox centers and its structure is similar to NADPH-cytochrom P450 reductase. PMID- 10540861 TI - [Chemical and functional properties of flavin coenzymes]. AB - The yellow-colored compounds with the basic structural frame work of 7,8-dimethyl 10-alkylisoalloxazine are generally termed as flavins. The 10-ribityl derivative, riboflavin, is the most abundant flavin found in nature and is known as vitamin B2. Riboflavin is a precursor of the flavocoenzymes, FMN (flavin mononucleotide) and FAD (flavin adenine dinucleotide) which function as prosthetic groups of flavocoenzymes. While flavocoenzymes are usually bound noncovalently to apoproteins of flavoenzymes, covalently-bound flavocoenzymes also occur in nature, though much less often. Flavin molecules can exist in three different redox states, i.e., oxidized, one-electron reduced and two-electron reduced states, and therefore can participate in redox reactions as either one- or two electron mediator, making the flavoenzymes extremely versatile in terms of substrate and type of reactions catalyzed. We classified flavoenzymes according to the electron-transfer process in their reductive and oxidative half-reactions and the mechanism of each class of flavoenzymes is discussed in detail. PMID- 10540862 TI - [Vitamin B6]. AB - 1934, vitamin B6 was discovered by P. Gyorgy. Vitamin B6 consists of 3 closely related pyrimidine derivatives: pyridoxine, pyridoxal and pyridoxamine and their phosphate esters. The tumor-specific compound has been recently found to be as a new B6 derivatives. The active form of vitamin B6 is pyridoxal phosphate (PLP) which is produced by multi-metabolizing enzymes including pyridoxine kinase and PNP/PMP oxidase. PLP functions as the coenzymes of many enzymes which catalyze biochemical reactions such as transamination and decarboxylation. A novel physiological function of PLP which is the negative modulator of the steroid dependent gene expression and the albumin gene expression has been also revealed. PMID- 10540863 TI - [Vitamin B12]. AB - Vitamin B12 is unique among all the vitamins in that it contains not only a complex organic molecule but also an essential trace element, cobalt. Vitamin B12 is synthesized in some bacteria but not in animals and plants. Intestinal absorption and subsequent plasma transport of vitamin B12 are mediated by specific vitamin B12-binding proteins and their receptors in mammals. Vitamin B12 taken up by the cells is enzymatically converted to coenzyme forms of vitamin B12, methyl- and adenosyl-vitamin B12, which function as coenzymes of methionine synthase (EC 2.1.13), involved in methionine biosynthesis, and methylmalonyl-CoA mutase (EC 5.4.99.2), involved in oxidation of odd-numbered fatty acids and amino acids (valine, isoleucine, and threonine), respectively. Chemical properties, physiological function, and intracellular metabolism of vitamin B12 are summarized in this section. PMID- 10540864 TI - [Nicotinic acid and nicotinamide]. AB - Nicotinic acid and nicotinamide are called niacin. They are the antipellagra vitamin essential to many animals for growth and health. In human being, niacin is believed necessary together with other vitamins for the prevention and cure of pellagra. Niacin is widely distributed in nature; appreciable amounts are found in liver, fish, yeast and cereal grains. Nicotinamide is a precursor of the coenzyme NAD and NADP. Some of the most understood metabolic processes that involve niacin are glycolysis, fatty acid synthesis and respiration. Niacin is also related to the following diseases: Hartnup disease; blue diaper syndrome; tryptophanuria; hydroxykynureninuria; xanthurenic aciduria; Huntington's disease. PMID- 10540865 TI - [Pantothenic acid]. AB - Pantothenic acid is the antipellagra vitamin essential to many animals for growth and health. It is widely distributed in nature; appreciable amounts are found in liver and some microorganisms. Bound forms of pantothenic acid, such as coenzyme A and 4'-phosphopantetheine, play important roles in various metabolic processes, especially, in fatty acid synthesis and degradation. PMID- 10540866 TI - [Vitamin C: structure-activity correlation and cytoprotective actions through free radical scavenging and extracellular matrix construction]. AB - Conventional studies on vitamin C have been conducted by single-dosage administration with ascorbic acid itself being so labile as to undergo irreversible degradation. In contrast, enrichment of intracellular ascorbate is accomplished by pro-vitamin C in which its 2,3-enediol moiety is protected with phosphate ester, being thereafter enzymatically esterolyzed. Ascorbic acid-2-O phosphate (Asc2P) gradually releases ascorbate, which is cumulatively taken up into diverse human or mammalian cells, and prevents cell injuries such as post ischemic reperfusional injury in the liver or heart, age-dependent telomeric DNA shortening in endothelial or epithelial cells, UV-B irradiational injury in the skin and lipid peroxide-induced injury in the vascular endothelium, and tumor invasion and metastasis. PMID- 10540867 TI - [Vitamin D: its chemistry, metabolism and biological functions]. AB - Vitamin D is metabolized to 25-hydroxyvitamin D(25(OH)D) in the liver and subsequently to 1 alpha,25-dihydroxyvitamin D(1 alpha,25(OH)2D) or 24R,25 dihydroxyvitamin D(24R,25(OH)2D) in the kidney. 1 alpha,25(OH)2D, the active form of vitamin D, stimulates intestinal calcium absorption and bone resorption, resulting in the elevation of plasma calcium. Recent studies have revealed that 1 alpha,25(OH)2D exerts a wide variety of biological actions such as cellular differentiation and proliferation in addition to its role in calcium homeostasis. Most cellular actions of 1 alpha,25(OH)2D are mediated by alterations in the transcription of vitamin D-dependent genes. In this review article, the mode of action and the biological function of 1 alpha,25(OH)2D are summarized. PMID- 10540868 TI - [Trend of vitamin E]. AB - The term vitamin E covers several related tocopherols and tocotrienols (alpha-, beta-, gamma-, delta-tocopherols and tocotrienols), which have been isolated from natural sources, but the most active of these is the alpha-form, Fig. 1). Vitamin E is an essential nutrient for higher animals including man. A daily recommended level of this vitamin has been established in 1989, and been revised in 1999. In man, a limited number of deficiency symptoms (Cerebella ataxia, Muscular weakness et. al.) have been noted in adults but only after prolonged impairment of fat absorption and genetic defects. The beneficial effects of this vitamin intake are reported by many studies. PMID- 10540869 TI - [Essential unsaturated fatty acids]. AB - Linoleic and alpha-linolenic acids are essential unsaturated fatty acids which were referred to as vitamin F much earlier. Their biological significance is to be converted to eicosapolyenoic acids including arachidonic acid which are the precursors of bioactive eicosanoids such as prostaglandins, thromboxanes and leukotrienes. Arachidonic acid is released from phospholipids by the catalysis of phospholipase A2, and then subjected to oxygenation by 5-lipoxygenase for leukotriene production and that by cyclooxygenase for prostaglandin and thromboxane formation. There are two isozymes of cyclooxygenase (COX-1 and COX 2). COX-2 is a product of an immediate early gene and an inducible enzyme, which is considered to play an important role in inflammation. COX-1 is a house-keeping enzyme found in platelets, kidney, stomach and many other tissues. Nonsteroidal anti inflammatory drugs currently used inhibit both COX-1 and COX-2 more or less, and causes gastric erosion and ulceration by inhibiting local production of prostaglandin E2. In view of such important roles of COX-2 in inflammation, inhibitors specific for COX-2 have been developed as anti-inflammatory drugs without stomach injury. PMID- 10540870 TI - [Vitamin K]. AB - It is well-known that vitamin K has a strong blood coagulation activity by acting a cofactor for gamma-carboxylase which catalizes the conversion of specific glutamic acid residue to gamma-carboxyglutamic acid residue (Gla). Some of the Gla-containing proteins, such as osteocalcin and matrix Gla protein have been found in calcifying tissues. These proteins are considered to play an important role in Ca-deposition. Recent studies have clearly demonstrated the substantial role of vitamin K in bone metabolism that has been developed for clinical use. Furthermore, diverse physiological activities have been found subsequently as follows: regulation of glucose metabolism, anti-arteriosclerosis, and induction of cell differentiation. Here I introduce the mainly physiological activities of vitamin K2, making a comparison with vitamin K1. PMID- 10540871 TI - [Folic acid]. AB - One-carbon derivatives of tetrahydrofolate, the coenzyme form of the vitamin folic acid, play a key role in DNA synthesis and cell replication, through their involvement in the biosynthesis of purine nucleotides and the amino acids. Although the most conspicuous symptom of folic acid deficiency is pernicius anemia, it has been observed that the lack of folate intake during the pregnancy induce the incidence of neural tube defects such as apina bifida. Recently, it has been reported that clinical progression of coronary heart disease and cerebral peripheral vascular disease occurred at a high rate in hyperhomocysteinemia with low folate content in plasma. Consequently, the nutritional importance of folate has been increasingly recognized. PMID- 10540872 TI - [Enhancement of glucose-induced insulin secretion and modification of glucose metabolism by biotin]. AB - Biotin causes improvements in disordered glucose metabolism by stimulating glucose-induced insulin secretion in pancreatic beta-cells and by accelerating glycolysis in liver and pancreas. Biotin is known to regulate hepatic and pancreatic glucokinase expression at both transcriptional and translational levels, and to regulate hepatic phosphoenolpyruvate carboxykinase expression at the transcriptional level. The effects of biotin on glucose-induced insulin secretion were investigated using the method of isolated pancreas perfusion. The pancreas of the biotin-deficient rat has an impaired insulin response to both glucose and arginine. In control rats as well as biotin-deficient rats, the insulin response to glucose stimulation was enhanced by the addition of 1 mM biotin to the perfusate. Biotin-induced enhancement of glucose-induced insulin release was evident within the first few minutes of perfusion. Since any effects on the glucokinase synthesis pathway would not be seen for at least 30 minutes, these results indicate that biotin may have the ability to act directly on the insulin secreting function of pancreatic beta-cells. Biotin perfusion was not found to cause enhancement of the arginine-induced insulin response, suggesting that biotin has no significant effects on the distal portion of the signaling pathway involved in insulin secretion. These results indicate that the administration of a high concentrations of biotin may improve the metabolism and/or utilization of glucose in patients with non-insulin-dependent diabetes mellitus. PMID- 10540873 TI - [Carnitine as a vitamin-like biofactor]. AB - Carnitine is a well-known cofactor for the beta-oxidation of long-chain fatty acid. It also plays a role in transport of acetyl moity for fatty acid and cholesterol synthesis, excretion of organic acid and xenobiotic acid as carnitine ester, and control of ratio of acetylCoA to CoA. Therapeutic effect of acetylcarnitine on Alzheimer disease and HIV-infection, and aberrant incorporation acetylcarnitine into brain under chronic fatigue syndrome have been reported. Carnitine deficiency causes hyperammonemia through suppression of gene expression of urea cycle enzymes. On the other hand, a large amount of carnitine has a therapeutic effect on hyperammonemia by still unclear mechanism. These suggest carnitine as a multifunctional biofactor. PMID- 10540874 TI - [Molecular mechanism of vitamin E transport in the body]. AB - Vitamin E occurs in nature in eight different forms, but animal body is enriched in alpha-tocopherol compared with other forms. alpha-Tocopherol transfer protein (alpha-TTP) is a liver cytosolic protein, which specifically binds alpha tocopherol, and plays an important role in the discrimination of various tocopherols in the body. Furthermore, alpha-TTP is a product of the causative gene for familial isolated vitamin E deficiency. Using cell culture system, it was shown that alpha-TTP functions to enhance secretion of alpha-tocopherol from liver cells and that the reaction utilizes a novel non-Golgi mediated pathway which may be linked to cellular cholesterol metabolism and/or transport. PMID- 10540875 TI - [Gene structure and transcriptional regulation of vitamin A, D binding proteins and nuclear receptors]. AB - Vitamin A and D play an important role in many biological processes including cell differentiation, proliferation and bone metabolism. These effects are believed to be mediated by specific nuclear receptors such as retinoic acid receptors (RARs) or vitamin D receptor (VDR), that regulate the transcription of a particular set of target genes. Hetero-dimers of RAR or VDR to retinoid X receptors (RXRs) bind target enhancer elements in their gene promoters. The target enhancer elements are referred as RA response elements (RAREs) or vitamin D response elements (VDREs), and composed of two hexamer core motife. DNA-bound RAR/VDR control transcription in a ligand-binding dependent way in co-operation with a multiprotein complex containing RNA polymerase II and a series of auxiliary factors, TFIIA, B, D, E, F and H. During process of ligand-induced transactivation by nuclear receptors, nuclear coactivators interacting with the AF-2 including ligand binding domain (LBD) seems to be involved. Several transcriptional coactivators and corepressors have been recently identified, and their function is currently under investigation. PMID- 10540876 TI - [Regulation of gene expression and vitamin]. AB - Action of 1,25-dihydroxyvitamin D, the most active metabolite of vitamin D, is exerted by the nuclear vitamin D receptor (VDR) mediated gene expression. Toward the expression of vitamin D function, several steps including 1,25 dihydroxyvitamin D production, tissue specific expression of VDR and transcription of target gene by VDR are involved. One of the important progress in vitamin D metabolism is the cloning of 25-hydroxyvitamin D-24-hydroxylase gene and 1 alpha-hydroxylase gene. Structural organization of the human VDR chromosomal gene and its promoter was also important to understand the amount of VDR expressed in tissues. A part of mechanism of tissue specific expression of VDR have recently been reported. Furthermore, VDR recruit several coactivators to achieve vitamin D-induced transactivation. Selective coactivator interaction with VDR may specify the array of biological actions of vitamin D. PMID- 10540877 TI - [Molecular basis of vitamin-responsive inborn errors of metabolism]. AB - In inborn errors where the defective enzyme had a cofactor requirement it was found that the administration of large amounts of the vitamin resulted in a clinical and biochemical improvement. A mutation that imposes an exaggerated requirement for a vitamin affects the apoenzyme directly, the conversion of a vitamin into coenzyme, or the attachment of vitamin to the apoenzyme. Direct proof has so far been provided that molecular defects in the said gene underlie both the vitamin-responsive and vitamin-nonresponsive forms. HCS deficiency is an exception, all HCS-deficient patients being responsive to biotin administration. We examined the relationship between the kinetic characteristics of HCS mutants and the clinical and biochemical features of the HCS deficient patients. PMID- 10540878 TI - [Tissue distribution of vitamin B12 and its dynamics in nervous tissues]. AB - Tissue distributions and intracellular localization of vitamin B12 (abbr B12) were described in spinal cords, roots and peripheral nerves of humans. The followings are results. 1) B12 contents of spinal cords are found to increase with centripetal slope. 2) Higher contents of B12 were found in ventral horn than lateral and posterior columns. 3) Motor nerves contained higher B12 contents than sensory nerves. 4) Considerable amounts of cyano-B12 was main component among B12 derivatives in nerve tissues. 5) Intracellular B12 was rich in mitochondrial fraction. 6) By ligation of sciatic nerves, B12 bound to nerves was released from nerve tissues and cyanide ions were later released, though its mechanisms remains unknown. PMID- 10540879 TI - [Cellular immunity and vitamins]. AB - The purpose of this review is to introduce the informations on cellular immunity and vitamins. Until now, many literatures have addressed the evidences showing the close relationship between cellular immunity and vitamins. In water-soluble vitamins, it is well-known that their deficiences induce the marked decrease of cellular immunity, although their supplementations have little effect. In contrast, lipid-soluble vitamins such as vitamin A and E markedly affect cellular immunity in both deficient and excess state. Vitamin A supplementation induces the increase of cellular immunity such as phagocytic and tumoricidal activities in human monocytes or mouse peritoneal macrophages. High intake of vitamin E has an ability to improve the decreased cellular immunity in the aged, which appears to be associated with the decreased production of prostaglandin E2 (PGE2). In summary, since vitamins are important nutrients to maintain and promote cellular immunity, the beneficial use of vitamins for the health of human should be considered. PMID- 10540880 TI - [Vitamins and apoptosis--induction of apoptosis in human cancer cells by nicotinic acid-related compounds]. AB - It was found that picolinic acid, dipicolinic acid, and isonicotinamide strongly induce apoptosis in human acute myelomonocytic leukemia cells, HL-60. Cinchomeronic acid, quinolinic acid, N1-methylnicotinamide, 6-aminonicotinamide, and picolinamide were weak inducers of the apoptosis. After treatments with picolinic acid, dipicolinic acid, and isonicotinamide, apoptosis started within 4 hr and it was induced in about 50% of the cells within 8 hr. These compounds also induced apoptosis in human chronic myelogenous leukemia cells, K562 and human cervical carcinoma cells, HeLa. However, apoptosis was not induced by these three compounds in quiescent normal human lymphocytes. A wide spectrum caspase inhibitor perfectly prevented DNA fragmentation induced by these compounds. But, caspase-1 inhibitor and caspase-3 inhibitor did not block DNA fragmentation. PMID- 10540881 TI - [Oxidative stress and vitamins]. AB - The importance of specific determination of key molecules involved in radical reaction is discussed in the evaluation of oxidative stress in animal tissues. As an effective index of oxidative stress, the concentration of lipid hydroperoxides, which is determined by a specific and sensitive method developed by the authors, is discussed firstly. The efficiency of the indicator is demonstrated by several results that it increases in tissues of aged, vitamin C deficient, vitamin E-deficient, iron-overloaded, or streptozotocin-induced diabetic rats. The other indicator, tissue vitamin C, which is also determined by a specific and sensitive method developed by the authors, is discussed. Based on the method, the profile of vitamin C decrease in tissues of ODS rats, which cannot synthesize the vitamin inherently, was determined and the in vivo interaction between vitamin C and vitamin E was demonstrated for the first time. The roles of oxidative stress in diabetes mellitus, atherosclerosis, and cell death (necrosis and apoptosis) were also discussed. PMID- 10540882 TI - [The current state on development of novel vitamin derivatives]. AB - In this review, we outline the current state on developments of novel vitamin A, C and D derivatives. We focused on the topics obtained with these novel vitamin A derivatives in relation to cell differentiation, apoptosis and gene regulation. As for vitamin D derivatives, the results obtained on cell differentiation and calcium metabolism were summarized. Lastly, we described some stable ascorbates, and further introduced stable (AA-2G) and lipophilic (6-Acyl-AA-2G) derivatives which have been recently developed in our laboratory. Both of them were demonstrated by us to express vitamin C activity in vivo. PMID- 10540883 TI - [Vitamin deficiencies and hypervitaminosis]. AB - There have recently been very few deficiencies with respect to fat soluble and water soluble vitamins in Japan All-trans-retinoic acid as induction or maintenance treatment improves disease free and overall survival against acute promyelocytic leukemia. In the isolated vitamin E deficiencies gene mutation has been cleared for alpha-tocopherol transferprotein. Recently, a relation of nutritional vitamin K intake and senile osteoporosis in women was epidemiologically demonstrated on a prospective study. Thiamin was yet noticed as development of deficiency in alcoholism, while the importance of supplemental folic acid during pregnancy has become especially clear in light of studies showing that folic acid supplements reduce the risk of neural tube defects in the fetus. With respect to hypervitaminosis, the Council for Responsible Nutrition (CRN), USA, has established safe intakes by identifying the NOAEL (No Observed Adverse Effect Level) and LOAEL (Lowest Observed Adverse Effect Level). Summaries of NOAEL and LOAEL for individual vitamins were shown. PMID- 10540885 TI - [Clinical roles of vitamins in hematopoietic disorders]. AB - Vitamins are essential organisms which promote various metabolisms and physiological systems. Several vitamins play important roles in hematopoietic system. Vitamin B12, C and folic acid are associated with DNA synthesis of erythroid nucleus, the deficiency of which causes the megaloblastic anemia. Some megaloblatic anemia and sideroblastic anemia might response to vitamin B1 and B6, respectively. Vitamin K participates in some coagulation factors in coagulation fibrinogenolysis system. It has been reported that vitamins A, D and K potentially differentiate leukemic cells and then induce the apoptosis, suggesting that they would be new therapeutic agents in acute leukemia. PMID- 10540884 TI - [Atherosclerosis]. AB - Atherosclerotic lesions can be characterized as accumulation of cholesterol esters and pathologic reactions by various cell groups. The pathogenesis of atherosclerosis has been discussed primarily on the basis of these two phenomena. A well-known concept to explain the etiology of atherosclerosis is the theory of response to injury. According to this theory, physiologically active substances such as platelet-derived growth factor (PDGF) and macrophage colony-stimulating factor (M-CSF) are released in response to injury of the vascular wall, and these substances induce pathologic reactions by the cells constituting the vascular wall. In the presence of excessive amounts of low-density lipoprotein (LDL), denatured LDL modified by oxidation or other reactions on the vascular wall is taken up by macrophages via scavenger receptors, resulting in the formation of foam cells and accumulation of cholesterol esters. Vitamins including Vitamin C and Vitamin E may play an important role in form cell formation by preventing the oxidation. PMID- 10540886 TI - [Therapeutic use of vitamin D and its analogues for rickets and osteomalacia]. AB - Recently, It has become clear that the mutant gene in X-linked hypophosphatemic rickets, vitamin D dependent rickets type I, and vitamin D dependent rickets type II were identified and they were caused by the disorder in the activation of vitamin D and the intracellular defect in vitamin D receptor. In this paper, the pathophysiology in various type of rickets or osteomalacia and the treatment by vitamin D agents were reported. So far, 1, alpha-(OH) D3 and 1, 25-(OH)2D3 have been effectively used as the treatment of these disease. However, vitamin D poisoning caused by these agents is still a serious issue for the treatment. In the near future, more effective treatment using genetic engineering is expected to treat the disorder of phosphate metabolism in X-linked hypophosphatemic rickets. PMID- 10540887 TI - [Diabetes and vitamin levels]. AB - The plasma vitamin levels are discussed in association with human diabetic condition. 1) Plasma vitamin B1 level of diabetic patients is revealed in the state of marginal deficiency. 2) Vitamin B6, as the coenzyme pyridoxal phosphate, plays an important role in the metabolism of carbohydrates, therefore B6 has been associated with impairments in gluconeogenesis and abnormal glucose intolerance. 3) Vascular complications of diabetes mellitus, such as atherosclerosis and retinopathy are considered to be related with glycation of low density lipoprotein which induces oxidative injuries to vascular endothelium. Administration of vitamins to diabetic patients reduces insulin requirement and attracts much attention for improvement of vascular complications. Vitamins play as not only nutritional supplements for deficiency, but pharmacological agents for treatment. PMID- 10540888 TI - [Vitamin E disturbances in chronic renal failure]. AB - Plasma levels of alpha-tocopherol (vitamin E) in chronic renal insufficiency (CRI) patients may be decreased, normal or elevated. However, an abnormal distribution of vitamin E in each lipoprotein has been reported. In comparison to control subject low-density lipoprotein (LDL), patient LDL contained less vitamin E. On the contrary, malondialdehyde (MDA) in patient LDL was enhanced. According to the evaluation of the susceptibility of LDL to in vitro oxidation and the rate of lipid peroxidation by fluorescence development during copper exposure, the susceptibility of patient LDL was enhanced, suggesting a possible relationship between excessive LDL peroxidation and accelerated atherosclerosis. In a clinical small uncontrolled trial, the increments of an aortic calcification index estimated by CT scan in patients treated with vitamin E were suppressed compared to those treated without vitamin E, suggesting that vitamin E might prevent the progress of atherosclerosis in CRI patients. PMID- 10540889 TI - [Hepatobiliary and pancreatic disorders as risk factors for fat-soluble vitamin deficiencies]. AB - The aim of this article is to describe the fat-soluble vitamin status in patients with hepatobiliary and pancreatic diseases, and the contribution of these vitamin deficiency or excess to hepatic injury. A considerable number of patients with advanced liver disease and cholestasis might actually be fat-soluble vitamin deficient, although clinical signs of deficiency are uncommonly seen in patients with vitamin A and E deficiency. Increased bone resorption may be the predominant cause of hepatic osteodystrophy. On the other hand, the possible causes of vitamin K deficiency seen in patients with hepatobiliary disease are the decrease of vitamin K absorption from intestine, the disturbance of vitamin K cycle and the decrease of pool area for vitamin K storage. The intake of vitamin A may be associated with the risk of liver cirrhosis in lifetime teetotalers, although the retinyl palmitate reduces hepatic fibrosis in rats. PMID- 10540890 TI - [Recent advances in geriatric vitaminology]. AB - The recent report concludes that elderly people have satisfying the recommended daily allowance in nutritional consumption except for calcium. However, those who are institutionalized, of low income, alcoholic and living alone are at high risk of vitamin deficiency. The high incidence in vitamin deficiency is shown when vitamin levels were measured with biological activity. Physiological range of vitamin seems to be enough to normalize this deficiency and nutritional supplementation would be useful. Those who are with the lack of calcium consumption or of time spent under the sunshine are at higher risk of having fracture. This is because of secondary hyperparathyroidism triggered by the decreased concentrations in serum calcium and 25-hydroxy-vitamin D3 level, and it is suspected to be the cause of osteoporosis among the elderly. To prevent the progression of osteoporosis in the elderly, serum PTH level needs to be kept in the normal range, and 100 IU/day, the recommended daily allowance of vitamin D in Japan, is far too small. PMID- 10540892 TI - [Vitamin and dermatology]. AB - Vitamin A, B1, B2, B6, B12, biotin, nicotinic acid, panthotenic acid, vitamin C, E and K have been used for various skin disorders. The use is mostly based on the similarity of the skin manifestations seen in their deficiencies, except for the rare cases of clear deficiency like pellagra. Recent introduction of vitamin A and D analogues for psoriasis and keratinization disorders resulted in significant progress in clinical dermatology. Application of vitamin C, E and beta-carotene++ for UV-induced skin damages are being studied, and the vitamins will be more important in dermatology in the future. PMID- 10540891 TI - [Vitamins in pregnancy]. AB - Vitamins are essential for the growth and normal function of human body. There are very few reports on vitamin A deficiency in pregnant women and newborn. Vitamin B complex includes various fractions essential to proper nutrition. The absence of vitamin C results Barlow disease in newborn. Vitamin D, the antirachitic Vitamin, is of great importance in safeguarding the mother and fetus from its relation to calcium and phosphorus metabolism. Vitamin E is also useful for the treatment of toxemia of pregnancy, intra-uterine growth retardation (IUGR), and neonatal jaundice. Vitamin K is well known for the protection of neonatal hemorrhage. The 6th recommended dietary allowances for Japanese in 1999 is shown in the table. PMID- 10540893 TI - [Disease prevention by vitamins based on epidemiological investigations in humans]. AB - Epidemiological studies in humans clearly indicate that higher consumption of vegetables and fruits, or a plant based diet, reduce the risk of cancer and coronary heart disease. Antioxidant vitamins or micro-nutrients contained in vegetables and fruits seem to be active components in the prevention of cancer and coronary heart disease. Intervention studies, however, failed to demonstrate protective effects. Further intervention studies with different study design are needed to clarify the role of supplemental vitamins or micro-nutrients in the prevention of cancer and coronary heart disease. The results of a few cohort and intervention studies have provided a good evidence that vitamin E supplement is protective against coronary heart disease. PMID- 10540894 TI - [Pathogenesis and conservative treatment of glaucoma]. AB - Glaucoma is a chronic disease which, if not treated, can lead to blindness. The reason for deterioration of function is neuropathia n. optici developed during the disease. Earlier increased ocular tension was considered to be the cause of neuropathia. By now we have realised that increased ocular tension (that above 21 Hgmm) in only one of the risk factors. The decay of optic nerve fibres is caused by circulatory failure on the one hand, and by the necrosis of ganglion cells on the other hand. In the conservative treatment of glaucoma pilocarpin was used earlier but nowdays the first place has been taken over by the group of betareceptor blockers, which are applied twice a day in the form of dropping. If this proves unsatisfactory, the treatment is complemented with carbonanhydrase inhibitor-drops 2-3 times daily. In certain cases this is followed by prostaglandin F2alfa analog drops once a day, dripped in the evening hours. Carboanhydrase inhibitor can be administered per oral as well: 1-2 times weekly. This latter cannot be given continuously: it is only a temporal solution for a few months in addition to other conservative therapy. Cholinerg drops can join in at any time of conservative treatment. The future method of conservative therapy is the combination of drops with varions effect, which decreases the frequency of daily drippings and enhances the efficacy of treatment. PMID- 10540895 TI - [Surgical management of villous and tubulovillous adenomas of the rectum]. AB - One hundred four cases of middle and low rectal villous and tubulovillous adenomas have been operated on with transanal polypectomy (8), transanal endoscopic microsurgery--TEM (80), anterior rectum resection with double stapled straight sigmoideorectosomy (7), and deep rectum resection, bi-directional mucosectomy and hand sutured straight sigmoideoanostomy (9). The option of the authors to remove the tumours in 5 cm to the dentate line are the transanal polypectomy or transanal mucosectomy corresponding to their size. The transanal endoscopic microsurgical technic is recommended to manage the polyps smaller than 4 to 5 cm in the middle rectum. The best radicallity in removal of the circular, extended villous adenomas could be achieved with deep rectum resection, bi directional mucosectomy and transanal straight, hand sewn sigmoideoanostomy. PMID- 10540896 TI - [The combined effect of psychotherapy and fluoxetine on obesity]. AB - Obesity as psychosomatic disease is a mass phenomenon. The number of obese males (BMI > 30) became doubled in the last ten years. In the etiology of obesity play an important role the reactive obesity. In the background of "yo-yo syndrome" often could be found depression, or other psychotic disorder. The low self esteem, body dissatisfaction, tension, anxiety disorders is well-known in a slimming diet. Obese subjects (n = 29) who were admitted on their request with a view to losing weight were examined (Hamilton Depressive Scala, Hamilton Anxietas Scala, Eating Attitude Test) Physical Conditioning and internal Medicine Department of National Sports Medicine Institute, Budapest. Among obesities with mild and severe depression as treatment of somatic complications was used fluoxetine, in severe cases and depression with severe anxiety was associated with supportive or cognitive-behavioral treatment. The prevalence of binge eating disorders were at 57% and bulimia nervosa was at 3% in using population (n = 29). Decreasing of anxiety and grade of depression significantly correlated with body mass index (p < 0.023, F = 1.997, p < 0.034, F = 3.131). The treatment of fluoxetine significantly correlated with body mass index (T1: p < 0.023, T2: p < 0.03, T3: p < 0.004). The patients indicated their well being as fluoxetine reduced eating, satiety and lower binges. PMID- 10540897 TI - [The third venous system of the lower extremity and its clinical significance]. AB - In deep venous thrombotic and aplasia cases superficial veins become dilated. With the resulting incompetence of the valves, venous blood stream is not directed to the heart, but to the ankle as well. In these cases the superficial system does not support the venous drainage of the limb, but gives a further load. The question is if one can ameliorate the venous drainage with removal of insufficient varicose veins, or does it make the outcome worse? A new examination was developed to determine if collateral veins in the subfascial space are enough to drain the venous blood of the limb. A tensiometer cuff was placed on the limb and inflated. The patient was asked to walk, if it was performed without complaints then the results was negative and the superficial veins were removed. In these cases no venous disturbances were detected during the operation or following that, in spite of the absence of deep venous circulation and radical removal of varicose venous bed. It means, that in the absence of deep venous circulation develops a collateral circulation not only in the subcutaneous, but in the subfascial space as well and it can alone maintain the venous drainage of the limb, this is the third venous system of the leg. PMID- 10540898 TI - [Confessions of Joannes Baptista Van Helmont]. PMID- 10540899 TI - [Herbs--trees--flowers]. PMID- 10540901 TI - Managed care law provisions require clarification. PMID- 10540900 TI - [Basedow disease]. PMID- 10540902 TI - Push continues for lawsuit abuse reform. PMID- 10540903 TI - Merging medicine, managed care, and public health. Interview by Linda Walburn. AB - In an effort to enhance communications and cooperation among Pennsylvania leaders in medicine, managed care, and public health, The Foundation of the Pennsylvania Medical Society, and its coalition, Keystones of Public Health, sponsored a one day symposium on September 8, 1999, at State Society headquarters. More than 70 key leaders gathered to identify and plan a project that will positively impact the health status of Pennsylvania. Nancy W. Dickey, MD, immediate past president, American Medical Association, delivered the keynote address. In this interview, Dr. Dickey shares her insights on the importance and impact of this statewide initiative. PMID- 10540904 TI - The paperless office: here for the asking. PMID- 10540905 TI - Understanding drug/drug interactions: cisapride. PMID- 10540906 TI - 'Growth of bacterial cultures' 50 years on: towards an uncertainty principle instead of constants in bacterial growth kinetics. AB - Ever since Monod's efforts to study bacterial cultures in quantitative terms, the growth of Escherichia coli on sugars like glucose has appeared an attractive subject for the mathematical description of nutrient conversion into biomass. But instead of simplicity, it is becoming evident that bacterial adaptations affect 'constants' such as K(s) (growth affinity constant) and are, in turn, a complex function of nutrient concentration. Instead of a single affinity, bacteria exhibit a continuum of nutrient scavenging abilities peaking at micromolar sugar levels; there is lower affinity with millimolar or submicromolar glucose in the medium. Similar problems arise in defining parameters such as Y (growth yield constant), because nutrient-limited growth at low exponential growth rates induces a continuum of hunger and starvation responses. Autocatalytic changes to the environment caused by growth (as well as external factors) ensure that bacteria present an ever-adapting interface to the outside world. The regulatory interaction between the organism and environment means that no universal kinetic constants describe bacterial growth. PMID- 10540908 TI - Genetic studies of a thermoregulated gene in the psychrotrophic bacterium Pseudomonas fluorescens. AB - In the psychrotrophic bacterium Pseudomonas fluorescens, some genes are thermoregulated: they are maximally expressed at a particular temperature within the broad range of temperatures that allow growth of this bacterium. To study this regulation, random transcriptional insertion fusions were obtained by means of mini-Tn5lacZ1 or mini-Tn5luxAB transposition. One fusion was studied in which beta-galactosidase production was maximal at a low-growth temperature. The mutated gene (that we call xsf) was highly homologous to xseA from Escherichia coli (and from other bacteria) which encodes the large subunit of exonuclease VII. Genetic tools were constructed in order to analyse and manipulate this fusion: a plasmid derived from R68.45 was used for chromosome transfer and a replacement vector was constructed to allow in situ marker exchange of the mini Tn5lacZ1 by an Hg(r) interposon. This vector was used to make double mutants and hence to study the effect of the insertion in xsf on the expression of other fusions. Six genes were thereby identified with a decreased expression in an xsf- background and with different characteristics of thermoregulation. PMID- 10540907 TI - Evolution of the linear chromosomal DNA in Streptomyces: is genomic variability developmentally modulated? AB - Genome rearrangements are responsible for the variability observed at the ends of the chromosome among Streptomyces species. The characterization of mutators, which are stimulated for genome plasticity, and of mutants produced at different stages of development support the idea that genome instability is developmentally modulated. PMID- 10540909 TI - Temperature-dependent flagellar antigen phase variation in Escherichia coli. AB - A new kind of flagellar phase (H antigen) variation which is dependent on the temperature of growth is described for Escherichia coli strains, all but one of which belong to serogroup O148, isolated in different geographical regions. At 37 degrees C the strains simultaneously displayed two different H antigen specificities, H40 and H53, while in cultures grown at 30 degrees C only a single flagellar antigen, H53, was detected. It was shown that the bacteria possess two separate flagellin genes, fliC40 and flkA53. An element controlling the temperature-dependent expression of fliC was localized in the region of flkA53. Relevant problems in H antigen serotyping in E. coli are discussed. PMID- 10540910 TI - Induction of lactate production associated with a decrease in NADH cell content enables growth resumption of Clostridium cellulolyticum in batch cultures on cellobiose. AB - When grown in batch cultures in fermentors with 23.4 mM cellobiose, Clostridium cellulolyticum displayed biphasic growth kinetics not associated with sequential substrate consumption and which led to a twofold higher production of biomass than previously reported. In the first growth phase, acetate was the major product of cellobiose metabolism, since lactate and ethanol productions remained low. Furthermore, an accumulation of intracellular NADH was observed. The transition towards the second growth phase was accompanied by an induction of lactate production, in such a way that lactate became the major product of C. cellulolyticum metabolism. In addition, a decrease in NADH concentration was measured, concomitant with this induction of lactate production and with the growth resumption. During both growth phases, the NADH-ferredoxin reductase hydrogenase system played a major function in NADH regeneration, since H2 production was 1.4- to 1.5-fold higher than that of CO2. Thus, we found that lactate production serves as an additional catabolic pathway enabling C. cellulolyticum to cope with excesses of carbon and NADH produced. Growth experiments on C. cellulolyticum under an atmosphere of carbon monoxide mimicked this phenomenon and confirmed that a high intracellular level of NADH can provide a barrier to bacterial growth. PMID- 10540911 TI - Comparison of iron uptake in different Helicobacter species. AB - Comparison of iron uptake of four Helicobacter species (Helicobacter pylori, Helicobacter felis, Helicobacter acinonyx, and Helicobacter mustelae), associated with various degrees of gastritis in their respective host, with five other species which colonize the intestinal tract of various animals (Helicobacter fennelliae, Helicobacter cinaedi, Helicobacter muridarum, Helicobacter bilis, and Helicobacter hepaticus), demonstrated that the iron acquisition system differed according to the ecological niche of the organism. Gastric Helicobacter, except for H. pylori, which used iron from human lactoferrin, were nonsiderophore producing organisms and were only able to obtain iron from heme and hemoglobin. Nongastric Helicobacter produced siderophores and were able to use for growth a wide range of iron sources (bovine and human lactoferrin and transferrin, heme, hemoglobin). PMID- 10540912 TI - Single DNA sequence common to all chlamydial species employed for PCR detection of these organisms. AB - Chlamydial infection is responsible for a wide spectrum of diseases of the eye, genitourinary tract, and lung. This group of organisms is also implicated in the pathogenesis of coronary artery disease as well as arthritis. Since cross-species infection is widely reported (though probably underestimated), it is an advantage to have a rapid and reliable method to detect all forms of chlamydiae in patient samples. We have identified a 160/163-bp DNA fragment in Chlamydia which is highly conserved in all chlamydial species. A polymerase chain reaction method based on this sequence has been developed to detect, in clinical samples, chlamydiae which have been shown to be positive by fluorescent-staining immunoassay; this method can be utilized in combination with restriction endonuclease cleavage to identify individual chlamydial species. Thus we have developed a sensitive and rapid detection method and have used it on samples from patients with respiratory and genital infections. PMID- 10540914 TI - Functional remodelling and left ventricular dysfunction after repeated ischaemic episodes. A chronic experimental porcine model. AB - This experimental study was set up to investigate left ventricular function and remodelling after repeated ischaemic episodes using magnetic resonance imaging (MRI). A significant reduction in mortality due to coronary heart disease (CHD) has been explained by both a decline in the incidence of acute myocardial infarction (AMI) and an improved post-AMI survival rate, suggesting a change in the natural history of CHD. Experimental intracoronary microembolization can induce different ischaemic patterns and the functional impact of repeated ischaemic episodes different from occlusion of central epicardial arteries can be studied. In this study repeated intracoronary microembolizations were performed in 20 domestic pigs. After 129 d, MRI was performed for assessment of left ventricular volume, mass and wall stress. Six pigs underwent serial MRI at baseline, immediately after embolization and at the end of the observation period. Microembolizations induced acute myocardial infarct expansion and increased left ventricular wall stress preceding chronic remodelling. End systolic and end diastolic volumes increased from 15.1 +/- 2.7 cm3 to 41.3 +/- 11.5 cm 3 (p < 0.002), and from 52.0 +/- 6.7 cm3 to 81.1 +/- 9.2 cm3 (p < 0.0007), respectively. End systolic wall stress increased from and 17.5 +/- 2.7 to 29.7 +/- 6.2 N/m2 (p < 0.001). Left ventricular filling pressures and cardiac index were unchanged. Histological examination revealed a diffuse pattern of perivascular fibrosis covering 12 +/- 3% of the left ventricular wall. This study demonstrates that repeated ischaemic episodes different from confined regional myocardial infarctions induce acute infarct expansion and chronic left ventricular remodelling in pigs. Serial assessment of absolute left ventricular volumes and mass is important during acute/chronic remodelling. PMID- 10540913 TI - The extended Biocor stentless aortic bioprosthesis. Early clinical experience. AB - In the "Extended" Biocor stentless aortic bioprosthesis, supra- and subvalvular extensions to a bovine pericardium ring carry three porcine leaflets. The extensions cover the "non-coronary" sinus of the prosthesis and allow optional enlargement of the aortic root down towards the mitral valve as well as upwards into the aortotomy. Seventy-one patients with this stentless valve (62 with predominantly aortic stenosis, 28 with concomitant CABG) are being prospectively studied. This paper reports follow-up one year after insertion. The upper and lower pericardial extensions were used in 61 and 11 patients, respectively. The average prosthetic valve size was 23.2 +/- 1.6 mm. Early mortality was 7% (5/71); late mortality (4/66, 5%/patient year) was not valve-related. Symptoms of thromboembolism (new neurological defects) occurred in four patients. There was no valve failure or late endocarditis. One year postoperatively the transvalvular mean pressure difference for all valves was 7.9 (3.1-18.4) mmHg. None of the patients had haemodynamically significant aortic regurgitation at follow-up; nine had trivial regurgitation. The "Extended" Biocor stentless bioprosthesis thus has a favourable haemodynamic profile and can be advantageous in elderly patients with narrow aortic roots, and often with thin and/or calcified aortic walls. PMID- 10540915 TI - The role of pulmonary and systemic circulation in the tracheal blood supply in rats. AB - The different roles of bronchial and pulmonary circulation in the tracheal blood supply were investigated in 26 female rats: a control group (CG, n = 7), a group with pulmonary hilar ligation (PL, n = 5), another with tracheal transsection (TL, n = 9) and a group with both these procedures (TL&PL, n = 5). Technetium 99 m was injected into the left ventricle postoperatively, and the radioactivity of tracheal samples was calculated as a percentage of injected activity/g tissue (%ID/g). The tracheal uptake averaged 1.9 in group CG, and 1.7, 1.3 and 1.5% ID/g in groups PL, TL and TL&PL, respectively. Tracheal transsection (TL) thus reduced the tracheal blood supply by 29.7% compared with the control group (p < 0.05), whereas the reduction of tracheal blood supply following pulmonary hilar ligation (PL) was only 10.9% (n.s.). Tracheal transsection combined with hilar ligation (TL&PL) effected a reduction of 19.9% (n.s.). We conclude that only 10.9% of the tracheal blood supply comes from the pulmonary circulation. PMID- 10540916 TI - Health-related quality of life in elderly patients with heart failure. AB - OBJECTIVE: To assess health-related quality of life (HRQL) in elderly patients with congestive heart failure (CHF) and correlate these to clinical and demographic variables. PATIENTS AND METHODS: HRQL was evaluated in 191 patients with CHF, aged 65-84 years, using a self-administered questionnaire including the Nottingham Health Profile (NHP), Quality of Life Questionnaire in Heart Failure and Patients' Global Self-Assessment. RESULTS: HRQL was more impaired in women than to men (p < 0.05), New York Heart Association functional class correlated to HRQL (p < 0.01) and HRQL, as assessed by NHP, was impaired in CHF patients compared to a previously evaluated, age and sex matched, normal reference population. CONCLUSION: Measurement of HRQL in heart failure patients provides important information in addition to a clinical evaluation, and inclusion of HRQL assessments in clinical practice is feasible and warranted. Specific intervention should be aimed at improving HRQL in those most severely affected. PMID- 10540917 TI - Mediastinal lymph node infiltration in non-small cell lung cancer and its role in curative surgery. AB - Despite the importance of lymph node infiltration for the classification and prognosis of non-small cell lung cancer (NSCLC), there are no accepted standards for quality of mediastinal lymphadenectomy. In 270 consecutive patients undergoing potentially curative surgery for NSCLC, including complete ipsilateral lymph node dissection, we investigated the possibility of a correlation between tumour location and lymph node infiltration. The tumours were classified as UICC stage I (n = 115), II (n = 42) or IIIa (n = 113). Patients with N2-disease (n = 68) showed up to 81% skip metastasis. Because of the observed dissemination of lymph node metastasis, tumour location could not predict nodal infiltration. The variability of nodal involvement and the frequent occurrence of skip metastasis thus make complete ipsilateral lymphadenectomy mandatory for curative management of NSCLC. PMID- 10540918 TI - Predictors of chest pain after coronary artery bypass grafting. AB - To identify preoperative biopsychosocial factors characterizing patients who will experience chest pain (self-reported) one year after coronary artery bypass grafting (CABG), 111 patients under 61 years of age were evaluated by questionnaire before CABG and 12 months postoperatively. A "Coronary Health Profile" was evolved to study quality-of-life indicators, e.g. "Sense of Coherence" (SOC), emotional state (loneliness, depressed mood, stress, anxiety) and social support as well as experience of chest pain, and the results were correlated to biomedical data. Chest pain was experienced in the first postoperative year by 34% of the patients. These patients, who were younger than those without chest pain, generally had a body mass index >25, as well as lower preoperative values for SOC, poorer emotional state and social support. Independent predictors in a multivariate stepwise logistic regression analysis were moderate/weak SOC, ejection fraction <50%, and moderate/severe mood depression. We conclude that biomedical as well as psychosocial factors have a significant impact as predictors of chest pain (of any origin) after CABG, and must be considered in preoperative evaluation. The findings indicate the need for biopsychosocial support/intervention before as well as after CABG. PMID- 10540919 TI - Acute ischemic chest pain is not associated with increased calcitonin gene related peptide (CGRP) levels in peripheral plasma nor in the coronary circulation. AB - Calcitonin gene-related peptide (CGRP) and substance P co-exist in capsaicin sensitive primary sensory neurons and are released from the myocardium after activation of sensory nerve fibres as well as by ischemia in animals. This study was undertaken to try to clarify the potential involvement of immunoreactive (ir) CGRP in anginal pain and myocardial ischemia in humans. One clinical group (n = 87) and one experimental group (n = 14) were studied. The clinical group was admitted to a coronary care unit with suspected or definite acute myocardial infarction (AMI). The experimental group consisted of patients with severe angina pectoris (NYHA III-IV). This group was subjected to atrial pacing up to the appearance of angina pectoris. Mean irCGRP levels at admission for the clinical group with and without AMI showed no significant difference. Neither were any significant differences found in irCGRP concentrations between patients with pain as compared to those without pain or in the group who had had chest pain >30 min before hospital admission as compared to those with chest pain <30 min. Extraction ratios for lactate and irCGRP was calculated in the experimental group. No statistically significant covariance was found between irCGRP extraction ratio and lactate extraction ratio (r(xy) = -0.006) at the time of appearance of angina during atrial pacing. Despite the facts that CGRP may be liberated by a variety of physiological stimuli and may act as a potent vasodilator in the human vasculature, no evidence has been found in this study that CGRP release is increased as a consequence of ischemia or ischemic pain. PMID- 10540920 TI - Acute effects of intravenous magnesium on ventricular refractoriness and monophasic action potential duration in humans. AB - The objective of this study was to examine the antiarrhythmic mechanisms of magnesium (Mg). The 12 patients who were selected for the study were given 12 mmol Mg sulphate followed by an infusion of 8 mmol/h. Ventricular effective refractory period (VERP). monophasic action potential duration at 50% and 90% repolarization during spontaneous rhythm (MAPD50, MAPD90) and atrial pacing at a cycle length of 600 ms (MAPD(50)600, MAPD(90)600) were measured before and after the intervention. Plasma magnesium concentration rose from 0.83 +/- 0.05 to 1.85 +/- 0.24 mmol/l (p < 0.001). Shortening of MAPD(50) (250 +/- 45 vs 240 +/- 46 ms; p < 0.05), MAPD(90) (294 +/- 42 vs 283 +/- 41 ms; p < 0.05) and shortening of sinus cycle length (SCL) (783 +/- 153 vs 742 +/- 160 ms; p < 0.01) were detected. During controlled cycle length, magnesium decreased MAPD(50)600 (230 +/- 28 vs 221 +/- 28 ms; p < 0.01), MAPD(90)600 (274 +/- 25 vs 261 +/- 29 ms; p < 0.01) and VERP (247 +/- 25 vs 241 +/- 21 ms: p < 0.05). QRS duration, QT interval and blood pressure remained unchanged. The change in SCL correlated with the change in MAPD(50) (r = 0.58; p < 0.05) and MAPD(90) (r = 0.55; p = 0.06 ) but not with MAPD(50)600, MAPD(90)600 or VERP. Magnesium acutely shortens ventricular monophasic action potential duration and refractoriness. This effect is partly mediated by mechanisms that increase heart rate. The reductions might bring salutary effects in arrhythmias evoked by prolonged repolarization. PMID- 10540921 TI - Recurrent massive hyperplasia of the thymus. AB - Differentiation of massive thymic hyperplasia from malignant lesions requires early resection. We report a case in which thoracoscopic thymectomy was performed for massive hyperplasia recurring 16 years after steroid therapy. This case provides additional information on the natural history, surgical management and histology of the disease. PMID- 10540922 TI - Unusual sites of uncommon endobronchial foreign bodies. Reports of four cases. AB - Foreign body aspiration occurs most commonly in children and can have serious consequences. In adults, it is associated with surgery, trauma and accidents. We report four unusual cases of foreign body inhalation. In one case a spike of wild barley entered the trachea through a tracheostomy cannula and migrated from the chest wall. In the second case a piece of coarse cloth which was introduced through a tracheostomy stoma aided by a wood sliver was retained in the trachea. In another patient an inhaled sewing needle migrated to the pericardium, and in the fourth case the head of a metal stud penetrated the trachea percutaneously through the neck and lodged in the right main bronchus. The incidence, causes, complications and management of such cases are discussed and the literature is briefly reviewed. PMID- 10540923 TI - Recurrence of uterine intravenous leiomyomatosis with intracardiac extension. Diagnostic considerations and surgical removal. AB - A 28-year-old woman (gravida 2, para 2) was admitted 20 months after a hysterectomy because of fibromyoma. The hysterectomy specimen had shown intravenous leiomyomatosis. The patient presented with unspecific abdominal symptoms, serologic signs of hepatic and renal failure and clinical right-sided heart failure. Progression despite treatment with a gonadotropin-releasing hormone analogue promoted transferral to the present centre. Abdominal ultrasonography, phlebography and transoesophageal echocardiography showed a left pelvic mass and a seemingly free-floating tumour extending from the left main iliac vein via the inferior caval vein to the right ventricle. During a combined cardiac and distal caval approach using extracorporeal circulation, a 45 cm massive leiomyoma was removed successfully. Seven weeks later the left pelvic tumour was removed radically together with left oophorectomy. At control 12 months later the patient was well and without any remaining symptoms. PMID- 10540924 TI - Intravascular ultrasound in equivocal coronary angiography. AB - Intravascular ultrasound (IVUS) is a well-established diagnostic tool that supplements coronary angiography in the evaluation of angiographical intermediate lesions as well as guiding Percutaneous transluminal coronary angioplasty. In this case report we describe the benefit of IVUS in diagnosing pseudostenosis as opposed to angiographically suspected guidewire induced dissection, and suggest the use of IVUS in all cases where angiography is equivocal. We also report a case of preoperative IVUS where the IVUS finding resulted in further coronary artery bypass grafting and suggest IVUS as a feasible alternative to probing of coronary arteries suspected of stenosis during coronary artery bypass grafting. PMID- 10540925 TI - Biological monitoring of sterilizers and sterilization failures in Norwegian dental offices in 1985 and 1996. AB - It is essential that dental office sterilizers be regularly challenged with biological indicators (BIs) in order to prove that the test spores are being killed during sterilization. The aims of the study were to biologically monitor Norwegian dental office sterilizers and to identify factors contributing to sterilization failure. In 1985, participants received a packet containing: (i) 4 BI units; (ii) a set of instructions; (iii) a questionnaire concerning operation (including biological monitoring) of the office sterilizer(s), and (iv) a return address envelope. In 1996, offices were sent (i) a survey which included demographic questions and inquiries concerning instrument sterilization processes; (ii) 2 sets of 3 BI units with instructions for their use on 2 different days; (iii) 1 control BI unit that was not to be processed, and (iv) a return-address envelope. Both private and public offices participated. Response rate to the 1996 study was 60%, which was 9.1% of all dental offices in Norway. Testing results indicated a 6.3% overall sterilization failure rate. Three out of 163 steam autoclaves (SAs) (1.8% of total) and 14 out of 109 dry heat (DH) ovens (12.8% of total) failed. DH ovens were over 7 times more likely to fail BI testing than were SAs (chi2, P < 0.01). Demographic or hygiene procedural factors could not be correlated to sterilization performance (chi2, P > 0.05). The failure rate for SAs (n = 216) in 1985 was almost 5 times greater than in 1996 (8.8% vs 1.8%). Improvement in sterilizer performance during the decade may be related to issuance in 1986 of Norway's 1st infection control guidelines for dentistry and greater awareness of infection control practices and/or to increases over the previous 10 years in the number of postgraduate courses offered in infection control. The current Norwegian guidelines on infection control practices in public health services, including dentistry, recommend regular biological monitoring of sterilizers without specifying how often. There is a lack of information among Norwegian dentists as to how frequently dental office sterilizers should be regularly monitored by BI. PMID- 10540926 TI - Simplified sampling methods for estimating levels of lactobacilli in saliva in dental clinical practice. AB - The aim of the present study was to evaluate whether estimation of lactobacilli was possible with simplified saliva sampling methods. Dentocult LB (Orion Diagnostica AB, Trosa, Sweden) was used to estimate the number of lactobacilli in saliva sampled by 3 different methods from 96 individuals: (i) Collecting and pouring stimulated saliva over a Dentocult dip-slide; (ii) direct licking of the Dentocult LB dip-slide; (iii) contaminating a wooden spatula with saliva and pressing against the Dentocult dip-slide. The first method was in accordance with the manufacturer's instructions and selected as the 'gold standard'; the other 2 methods were compared with this result. The 2 simplified methods for estimating levels of lactobacilli in saliva showed good reliability and specificity. Sensitivity, defined as the ability to detect individuals with a high number of lactabacilli in saliva, was sufficient for the licking method (85%), but significantly reduced for the wooden spatula method (52%). PMID- 10540927 TI - Effect of a chemo-mechanical caries removal system (Carisolv) on dentin topography of non-carious dentin. AB - The purpose of the present study was to examine the morphology of healthy dentin surfaces after treatment with Carisolv followed by conditioning with phosphoric acid and EDTA, since surface morphology may be of interest for dentin bonding. Another purpose was to evaluate the effect of treatment with Carisolv on healthy non-carious dentin surfaces with exposed collagen fibers. Scanning electron microscopy was utilized to carry out a detailed morphological examination of the dentin surfaces with regard to presence or absence of both smear layer and collagen fibers. Twelve premolars extracted for orthodontic reasons from young adults were used. The two etchants appeared to have produced two distinctly different surfaces. Etching with phosphoric acid following Carisolv treatment resulted in a porous dentin surface, while EDTA etching without prior Carisolv treatment appeared to have uncovered an intact collagen network. In contrast, the surfaces treated with Carisolv prior to EDTA etching displayed smooth intertubular surfaces with only occasional fibers. Apparently, the ability of EDTA to expose collagen in the dentin surface is counteracted or inhibited by the Carisolv treatment. Furthermore, it cannot be excluded that the Carisolv treatment in itself may have dissolved collagen fibers. Since most bonding systems claim bonding to the collagenous component of dentin, the question arises which of the etched surfaces is preferable and to what degree the collagenous component contributes to bonding strength. Further studies are thus needed to evaluate the micromechanical retention of a restoration to the different surfaces described in the present study. PMID- 10540928 TI - Rectal sedation with diazepam or midazolam during extractions of traumatized primary incisors: a prospective, randomized, double-blind trial in Swedish children aged 1.5-3.5 years. AB - The aim of this study was to compare rectal sedation with diazepam and rectal sedation with midazolam with regard to sedative effect, treatment acceptance, and amnesia. Ninety children, 1.5-3.5 years of age, consecutively referred for extractions of traumatized primary incisors were randomly sedated with diazepam (0.7 mg/kg body weight) or midazolam (0.3 mg/kg body weight). The study design was randomized and double-blind. The level of sedation (state of mind) was assessed prior to and 10 and 60 min after administration of the drug by use of a behavioral scale (Wilton). The children's acceptance of procedures was assessed using another behavioral scale (Holst) during administration of the sedative, application of topical anesthesia, injection of a local anesthesia, and extraction. Amnesia was evaluated by the parents on the following day, with the child being asked standardized questions. Parental ratings of the child's and their own distress during and after treatment were made on a visual analog scale (VAS). No differences were found between the sedatives concerning level of sedation during treatment, acceptance of procedures, or amnesia. At discharge, 60 min after administration of the sedative, the children receiving diazepam were significantly more agitated (P=0.006). Parental rating on a VAS of the child's discomfort after treatment was significantly higher in the diazepam group (P=0.006). There was a tendency for children with poor acceptance of the rectal administration to display a more negative acceptance of the dental treatment. In conclusion, the present results, in combination with known pharmacological advantages, indicate that midazolam is preferable in outpatients when sedation is needed and amnesia is desirable. PMID- 10540929 TI - Tunnel restorations in general practice. Influence of some clinical variables on the success rate. AB - Using bitewing radiographs and clinical inspection, the success rate for tunnel restorations was assessed in a population with low caries activity. The material consisted of 242 tunnel restorations in permanent premolars and molars in 142 individuals (mean age = 18.8 years). The median DFSappr value (decayed and filled approximal surfaces) at the time of restoration was 4.0. The mean follow-up time was 25 months. Bivariate associations between the outcome variable (success/failure of the tunnel restoration) and conceivable explanatory variables were investigated. In a multivariate logistic regression analysis, the independent variables tooth type (premolars vs molars), surface site (mesial vs distal), radiographic stage of approximal carious progression and age of patient at the time of restoration (9-15 years vs > 15 years) were used to estimate the effect on the dependent variable success/failure. Using the life table method, the estimated cumulative proportion of successful restorations was 81% after 2 years and 64% after 3.5 years. The success rate was not related to caries activity and did not differ between the two types of tunnel preparation techniques nor between different follow-up periods. In the multivariate regression analysis, tooth type (molars vs premolars) was the only factor significantly associated with failure. Thus, a failure occurred about 5 times as often in molars as in premolars. Of the failures, half were due to caries; either radiographically observed adjacent to the restoration or progressing enamel caries on the outer proximal surface. Marginal ridge fractures constituted 26% of the failures. From the present results it can be concluded that in a population with low caries activity, the tunnel restoration technique can be recommended for premolars. PMID- 10540930 TI - Acceptance and side effects of nitrous oxide oxygen sedation for oral surgical procedures. AB - Two hundred and forty-one treatment sessions with nitrous oxide oxygen sedation were performed in 194 patients undergoing ambulatory oral surgery procedures. Removal of mesiodentes and tooth transplants were the most frequent procedures in age groups under 13 years, while removal of impacted teeth was predominant in older age groups. Local anesthesia was used in addition to inhalation sedation in 238 sessions. Median gas volume rate was 10 l/min, median concentration 50% and median duration of procedures 31 min. In 10 sessions (4.1%) sedation was not accepted, while in 25 (10.4%) sessions the procedure could be completed with some difficulty. No potentially dangerous complications were noted. Side effects occurred in 18 sessions in 16 patients. All side effects were minor and easily handled. Logistic regression analysis revealed that failure, defined as poor acceptance and/or presence of side effects, was associated with ASA class 2 and general apprehension, especially based on previous negative experience with medical or dental treatment. Nitrous oxide oxygen sedation is a reliable, efficient and safe adjunct to local anesthesia in both healthy children and adults undergoing ambulatory oral surgery procedures. PMID- 10540931 TI - Machine learning methods applied on dental fear and behavior management problems in children. AB - The etiologies of dental fear and dental behavior management problems in children were investigated in a database of information on 2,257 Swedish children 4-6 and 9-11 years old. The analyses were performed using computerized inductive techniques within the field of artificial intelligence. The database held information regarding dental fear levels and behavior management problems, which were defined as outcomes, i.e. dependent variables. The attributes, i.e. independent variables, included data on dental health and dental treatments, information about parental dental fear, general anxiety, socioeconomic variables, etc. The data contained both numerical and discrete variables. The analyses were performed using an inductive analysis program (XpertRule Analyser, Attar Software Ltd, Lancashire, UK) that presents the results in a hierarchic diagram called a knowledge tree. The importance of the different attributes is represented by their position in this diagram. The results show that inductive methods are well suited for analyzing multifactorial and complex relationships in large data sets, and are thus a useful complement to multivariate statistical techniques. The knowledge trees for the two outcomes, dental fear and behavior management problems, were very different from each other, suggesting that the two phenomena are not equivalent. Dental fear was found to be more related to non-dental variables, whereas dental behavior management problems seemed connected to dental variables. PMID- 10540932 TI - Clinical performance of indirect composite resin inlays/onlays in a dental school: observations up to 34 months. AB - The aim of this retrospective clinical study was to evaluate the clinical performance of indirect composite resin inlays and onlays. Patients among the dental school clientele in need of posterior approximal filings and preferring esthetic restorations were included. Clinical teachers or trained students under supervision carried out the preparations, made impressions and prepared stone casts. Inlays made from either Tetric, Z100 or Maxxim were light-cured and placed in a light oven for secondary curing, before being luted with a dual cure cement. At recall, the inlays were evaluated using slightly modified US Public Health Service (USPHS) criteria. Twenty-two patients with 50 fillings presented for the assessment. The right censored observation periods ranged from 12 to 34 months, with a mean of 20. With the only exception of an early fracture of one onlay, all restorations were classified as successful. This was based on 15 "A" (optimal) and 34 "B" (acceptable) ratings, each of which representing the lowest rating for the individual restoration. The major reason for the "B" ratings was imperfect gingival marginal adaptation due to a small surplus of bonding material and/or luting cement. PMID- 10540933 TI - Root resorption and signs of repair in Papillon-Lefevre syndrome. A case study. AB - The aim of this investigation was to describe some tooth-related histological features of prepubertal periodontitis. Teeth extracted during treatment of two Papillon-Lefevre syndrome patients were processed by means of the sawing and grinding technique. Light microscopy examination revealed little or no cementum in the coronal parts of the roots. Resorptions of various depths (0.02 to 1.5 mm) and to various extents (affecting up to 1/3 of the root surface) were observed in the 5 investigated teeth. Some resorptive defects on 1 of the examined incisors showed signs of spontaneous repair. Extrinsic fibers were inserted into the new cellular intrinsic fiber cementum which had formed directly on the bottom of the defect. Intact acellular extrinsic fiber cementum was found where fibers were still attached. Here, the characteristic of pristine cementum, a hyaline layer of peripheral dentin, could be identified. If resorption was not present, the cementum did not show any signs of hypoplasia. Thus, histological features of prepubertal periodontitis in the current material were (i) areas of extensive resorption, (ii) signs of spontaneous repair, and (iii) healthy cementum. PMID- 10540934 TI - Dentists' selection of measures for assessment of oral health risk factors for Finnish young adults. AB - Dentists' selection of measures for assessing oral health risk factors for young adults, in relation to their oral health status and to those dentists' characteristics, was studied in one administrative unit of the Finnish public oral health service. A random selection (n = 239) was made of all young adults born in the period 1966-71 and clinically examined during 1994. On the original oral health records of those selected, all notes were scrutinized concerning the most recent clinical examination and treatment course; in total 208 (87%) records. We found that assessment of risk factors to oral health was rare. The patient's diet had been recorded as assessed in 7% of all cases, use of fluoride in 8%, and oral hygiene habits in 14%. No salivary tests were performed; nor was patients' use of tobacco assessed. No correlation was detected between measures used by these dentists and their patients' oral health status (DMFT and DT scores, number of approximal incipient lesions, and number of healthy sextants by CPITN). The oral health status impelled only slightly assessments by bite-wing radiographs. Fewer than half (44%) of the dentists performed and recorded any kind of assessment measure; 4% assessed diet, hygiene, and use of fluoride for all their patients in our sample. A dentist's gender showed no correlation with number of measures used; younger dentists tended to perform and record assessments slightly more often than did older dentists, but in all age groups there were those who had not done this. The practice of risk-factor assessment should be more widespread and standardized, contributing to needs-based treatment and allocation of resources. PMID- 10540935 TI - Gender differences in knowledge, attitude, behavior and perceived oral health among adolescents. AB - A cross-sectional dental questionnaire census survey was conducted in classrooms of 17,280 students aged 13-18 years in Skaraborg County, Sweden. The overall response rate, based on school attendance on the test day, was 91% with no gender differences at the senior level, and 86% (boys 87%, girls 85%) at the upper secondary level. The aim was to examine gender differences in knowledge, attitude, behavior and perceived oral health. A retest study showed good agreement. Thirty-one percent of the girls and 21% of the boys flossed regularly. Eleven percent reported daily candy consumption, with no significant gender difference. Girls, however, more often than boys considered their own consumption to be too high. This gender difference in attitude was most pronounced among older daily consumers (odds ratio (OR) = 5.8 [3.7-9.2]). Oral health was regarded as important by a majority of the students (95%). Girls considered sound teeth to be more important than did boys, both among the younger (OR = 1.7 [1.4-2.1]) and the older (OR = 2.4 [1.9-3.1]) adolescents. It is concluded that most adolescents had a positive dental attitude and perceived their own oral health to be good. Poorer knowledge and behaviors concerning oral health were demonstrated. Gender differences existed in most issues. Girls scored more favorably on behavioral measures, showed more interest in oral health, and perceived their own oral health to be good to a higher degree than did boys. PMID- 10540936 TI - Extracorporeal photochemotherapy for cutaneous T-cell lymphoma: a 9.7-year experience. AB - Cutaneous T-cell lymphoma (CTCL) is an indolent lymphoma usually of CD4+ T lymphocytes in which the aggressive treatment for the advanced stages does not increase survival. Photopheresis has been established as an alternative modality for the therapy of erythrodermic CTCL and reportedly improves survival in patients with advanced stages of the disease. The objective of this study is to review the experience of treating patients with erythrodermic CTCL with extracorporeal photochemotherapy (ECP) at the New York Veteran Affairs Medical Center/NYU Medical Center between September 1987 and April 1997. Forty-one patients with erythrodermic CTCL (stages III and IV) received photopheresis; 25 of them fulfilled the inclusion criterion, i.e., the completion of greater than or equal to 6 cycles of photopheresis. Skin score was defined as a product of severity and percentage of involved surface area. Complete clinical response was defined as disappearance of measurable disease for at least one month, and partial response was defined as greater than or equal to 50% clearance of measurable disease for at least one month. The profile of the patients was: 20 men, 5 women; average age: 64.2 years; 17 patients had stage III disease, and 8 had stage IV disease. Five of the 25 patients (20%) achieved complete clinical response, another 15 (60%) had partial response, and 5 (20%) had no response. The mean time (+/- SD) to achieve complete clinical clearance was 12.6 +/- 10 months (range: 4-30 months) and the mean time (+/- SD) to obtain partial clinical response, including complete response, was 9.7 +/- 5.3 months (range: 4-17 months). Remission duration ranged from 9 to 67 months. The median survival time from the time of initiation of photopheresis is estimated at 70 months. The complete responder group had a lower median CD4/CD8 ratio compared to the non responders at baseline (3.8 vs 7.2, respectively), although the difference was not statistically significant (P = 0.40). At the time of maximal response, the CD4/CD8 ratio of the complete responder group decreased towards normal values (median = 1.2), whereas this ratio increased among the non-responders (median = 11.0; P = 0.04). Side effects were minimal. Extracorporeal photochemotherapy is an effective and safe treatment for erythrodermic CTCL. In some of these patients, it can induce a long-term and complete clinical remission. PMID- 10540937 TI - The photoprotective effect of ascorbic acid, acetylsalicylic acid, and indomethacin evaluated by the photo hen's egg test. AB - The aim of the present study was to evaluate the supposed photoprotective effects of ascorbic acid, acetylsalicylic acid, and indomethacin by the photo hen's egg test, a recently developed new model for phototoxicity. Therefore, in three independent experimental settings the blood vessel system of the embryo's yolk sac of 24 incubated hens' eggs (2 test groups) were exposed to 60 mJ/cm2 ultraviolet B (UVB) to induce severe phototoxic damage. Before UVB irradiation, one of these test groups was exposed additionally to one of the test substances and the other one to 0.9% sodium solution alone. To exclude plain toxic reactions, two additional test groups were exposed only to 0.9% sodium chloride solution or to one of the test substances alone. Over a test observation period of 24 h, the embryo lethality as well as the morphological changes of the yolk sac blood vessel system were observed. Ascorbic acid led to a significant and remarkable reduction of the UVB-induced damage. Acetylsalicylic acid also showed a significant but lower photoprotective capacity. In contrast, indomethacin showed no photoprotective effects in the photo hen's egg test. PMID- 10540939 TI - Immediate pigment darkening thresholds of human skin to monochromatic (362 nm) ultraviolet A radiation are fluence rate dependent. AB - When human skin is irradiated with ultraviolet radiation (340-400 nm) there is an immediate pigment response, termed Immediate Pigment Darkening (IPD). This reaction is thought to be due to photooxidation of preexisting melanin, precursors and/or melanin metabolites because it appears during exposure. It has been demonstrated that UVA-induced skin reactions, including erythema and pigmentation, are oxygen dependent. Therefore, these reactions should also be irradiance (fluence rate) dependent. The purpose of this investigation was to determine the dependence of the IPD threshold on fluence rate. We exposed the forearm of 12 volunteers (skin type II-V) to monochromatic UVA radiation (362 nm) at a range of fluence rates of 6-115 mW/cm2 and determined the fluence at which pigment was perceptible. The threshold fluence for the IPD reaction increased by a factor of 2.7 as the fluence rate increased by a factor of 18. Therefore, we conclude that the IPD reaction following exposure to 362 nm radiation is dependent on fluence rate, and independent of skin type. PMID- 10540938 TI - Effect of stretch on the ultraviolet spectral transmission of one type of commonly used clothing. AB - The spectral ultraviolet (UV) transmission through stockings was measured in field and laboratory based trials using a spectroradiometer. From these spectral UV measurements, the ultraviolet protection factor (UPF) was calculated. The UPF of stockings measured in the field was generally higher than that measured in the laboratory when using a quartz tungsten halogen light as the UV source. The UPF of 50 denier stockings decreased 868% when stretched 30% from their original size. Doctors recommending and patients using high denier stockings for patient photoprotection should be aware of the dramatic decrease in UPF when the stocking is in a stretched position, such as over a human leg. PMID- 10540941 TI - The clinical features and management of actinic prurigo: a retrospective study. AB - Actinic prurigo (AP) is a rare photodermatosis, mostly affecting young American Indian girls. A retrospective descriptive study was done in the National Skin Centre, Singapore. Our patients have different characteristics compared to the previous reports. Of the 11 cases found between 1990 and 1998, 10 were male. All of the patients had the onset in adulthood. The condition was recognised by the presence of papules and nodules on the sun-exposed areas, predominantly on forearms and back of hands. Phototests revealed lowered minimal erythemal dose (MED) to ultraviolet A (UVA) alone in 2 patients and lowered MED to both UVA and UVB in another 4 patients. Patch, photopatch and histological examination did not show any significant finding. Sun protection, emollients and topical steroid were the baseline treatment for all patients. Intralesional steroid, systemic steroid, psoralen plus ultraviolet A (PUVA), azathioprine and thalidomide were used in some patients, with variable results. PMID- 10540940 TI - Temperatures reached inside stand-up ultraviolet treatment boxes. AB - Heat generated within ultraviolet treatment units can exacerbate eczema. To document the actual temperature changes within the treatment units, we measured the air temperatures in standard stand-up psoralen plus ultraviolet A (PUVA), narrowband ultraviolet B (UVB), broadband UVB, and combination UVA/UVB cabinets using a thermocouple thermometer. For the latter unit, we also measured the air temperatures with and without ventilation systems, and actual skin temperatures on individuals undergoing light treatment. The air temperatures rose significantly in all the treatment units, more so with PUVA and narrowband UVB boxes, and were highest with the ventilation systems shut off. Skin temperatures also rose significantly, but less dramatically. Ventilation is essential in maintaining comfortable temperatures within ultraviolet treatment units. PMID- 10540942 TI - Hand and foot PUVA soaks: an audit of the Massachusetts General Hospital's experience from 1994 to 1998. AB - Palmoplantar psoriasis and eczema can be chronic recalcitrant dermatoses. Oral psoralen plus ultraviolet A (PUVA) has proven effective for these, but has a number of unwanted side effects. Previous studies have shown that regional PUVA therapy using the 8-methoxypsoralen (8-MOP) soak method followed by immediate UVA irradiation is also beneficial and well tolerated. The purpose of this audit was to determine the efficacy of hand and foot PUVA soaks by reviewing the experience of the Massachusetts General Hospital's Phototherapy Unit with this treatment modality. Phototherapy records of all patients who underwent hand and foot PUVA soaks from 1994 to 1998 were reviewed. Details regarding the treatment procedure, patient compliance, patient response and incidence of side effects were noted and summarized. Of the 56 patients who underwent at least 20 treatments, 29% had excellent response, 42% had minimal to moderate response, and 29% had poor response. The average number of treatments to induce clearing was 41. The average maximum treatment dose at clearing was 5.85 J/cm2, while the average cumulative dose to achieve clearing was 267.17 J/cm2. 8-MOP PUVA soak therapy is quite useful for chronic hand and foot dermatoses. Patient compliance must be emphasized to obtain favorable results. PMID- 10540943 TI - Ultraviolet radiation-blocking characteristics of contact lenses: relevance to eye protection for psoralen-sensitised patients. AB - Nine brands of contact lens marketed as "UV protective" were tested for ultraviolet (UV) transmission in order to assess potential suitability for psoralen-sensitised patients. UV-transmission characteristics of hydrated lenses was tested with a Bentham monochromator spectro-radiometer system. All lenses showed minimal transmission loss in the visible band. The performance of the nine lenses was uniform for ultraviolet B radiation with negligible transmission, but showed variation in transmission for ultraviolet A radiation. None of the lenses complied with UV-transmission criteria used previously to assess UV-blocking spectacles. Only two lenses had UV-blocking characteristics which came close to the arbitrary criteria used. The performance of ordinary soft and hard lenses was very similar, with negligible blocking of UV radiation. None of the nine contact lenses marketed as "UV protective" excluded sufficient UVA to comply with criteria in current use to assess UV protection in spectacles for psoralen sensitised patients. However, the improved UV-blocking characteristics of contact lenses identified in this paper compared to previous studies suggests that such a contact lens will soon become available. Meanwhile, contact lens-wearing systemically sensitised PUVA patients should continue to wear approved spectacles for eye protection whilst photosensitised with psoralen. PMID- 10540944 TI - Steady-state mRNA levels of interleukin-1, integrins, cJun, and cFos in hairless mouse skin during short-term chronic UV exposure and the effect of topical tretinoin. AB - We have proposed that UV activation of cytokine and integrin signaling pathways may initiate the photoaging process and that one of the effects of tretinoin treatment may be to alter the cytokine and integrin patterns. In previous results, steady-state mRNA levels of interleukin-1alpha, tumor necrosis factor alpha, transforming growth factor beta, collagenase, stromelysin, collagen, and integrins (alpha1 and alpha2) were increased in the skin of hairless mice that were either UV treated or concurrently treated with UV followed by topical tretinoin for 5 weeks. The aim of this study was to focus on the expression of alpha1, alpha2 and alpha5 integrins, IL-1alpha, IL-1beta, cJun, and cFos at an earlier time point (3 weeks). Animals were UV irradiated thrice weekly for 3 weeks and were treated topically with either 0.05% tretinoin or the vehicle immediately after each exposure. Total RNA was prepared and used in RT-PCR with radiolabeled dCTP and specific primers. UV slightly increased steady-state mRNA levels for alpha1, alpha2 and alpha5 integrins whereas UV + tretinoin increased their expression (3-, 2- and 7-fold respectively). Steady-state mRNA levels for IL-1alpha, IL-1beta and cJun were increased with UV (3-, 12- and 6-fold respectively) and with UV + tretinoin (6-, 7- and 9-fold respectively). In contrast, cFos expression was unchanged. In situ staining for IL-1alpha mRNA was slightly more abundant in mice treated for 3 weeks with UV and UV + tretinoin than in controls whereas 5 weeks of UV + tretinoin treatment gave strongly positive staining. Results are consistent with cytokines and integrins mediating the effects of UV on the skin, with modulation of these effects by tretinoin. PMID- 10540945 TI - Simple changes to PUVA phototherapy may minimize the photocarcinogenic risks. PMID- 10540947 TI - Characterization of Chabertia ovina by isoenzyme gel electrophoresis: comparative study with Oesophagostomum venulosum. AB - The glucose-6-phosphate dehydrogenase (G6PD, EC.1.1.1.49), glucose phosphate isomerase (GPI, EC.5.3.1.9), and malate dehydrogenase (MDH, EC.1.1.1.37) isoenzymatic patterns of Chabertia ovina were determined by starch-gel electrophoresis. The G6PD and GPI isoenzymatic patterns were characterized by the existence of three phenotypes: (1) a single and slow anodic band, (2) a single and fast anodic band, and (3) a large spot matching its migration with bands 1 and 2. These three phenotypes may be explained as the existence of only one gene locus for the G6PD and GPI in C. ovina. Allelic frequencies and the Hardy Weinberg test were determined. This test indicated that the population was not in Hardy-Weinberg equilibrium. The MDH isoenzymatic pattern of C. ovina was characterized by the presence of two bands with anodic and cathodic migration. Furthermore, comparative isoenzyme studies were carried out between Oesophagostomum venulosum and C. ovina. The different G6PD, GPI, and MDH isoenzymatic patterns observed for the two species allowed us to distinguish them and, therefore, to use isoenzymatic patterns as a diagnostic tool to discriminate these species. PMID- 10540946 TI - A study of the systematics of Theileria spp. based upon small-subunit ribosomal RNA gene sequences. AB - The systematics of benign and moderately pathogenic Theileria isolates from cattle and deer originating from different geographic regions was undertaken by small-subunit ribosomal RNA (SSU rRNA) gene nucleotide-sequence analysis. A maximum-likelihood phylogenetic tree constructed from these sequences resulted in two major divisions, each with a common ancestor. One major division branches into four relatively divergent groups, including (1) bovine Theileria sp. Type D (USA and Korea), (2) T. mutans Intona and Theileria sp. MSD (Africa), (3) T. cervi (USA), and (4) well-characterized pathogenic Theileria spp. (Africa). The other major division branches into two groups: (1) T. buffeli Warwick and T. buffeli Marula and (2) a second branch of closely related isolates with SSU rRNA gene Types B, B1, C, E, and H. Putative geographically associated diversity was noted only in the Korean bovine Theileria spp. with SSU rRNA gene types C and H and in African T. mutans Intona and Theileria sp. MSD. The current results show that the United States bovine Theileria isolates are not T. mutans because they have T. buffeli Marula (Type A) and/or Type D (species undesignated) SSU rRNA gene sequences. The taxonomic separation of T. buffeli Warwick from African T. mutans is confirmed in this study. PMID- 10540948 TI - Taxonomy and phylogeny of some Eimeria (Apicomplexa:Eimeriidae) species of rodents as determined by polymerase chain reaction/restriction-fragment-length polymorphism analysis of 18S rDNA. AB - The 18S rDNA genes of 10 Eimeria species from rodents (E. albigulae, E. arizonensis, E. falciformis, E. langebarteli, E. nieschulzi, E. onychomysis, E. papillata, E. reedi, E. separata, E. sevilletensis) were polymerase-chain reaction (PCR)-amplified, digested with 12 restriction endonucleases, and electophoresed in agarose gels. The resulting fragment patterns (riboprints) distinguished all species except E. sevilletensis from E. falciformis, and E. arizonensis from E. albigulae; the sporulated oocysts of the latter two species and of E. onychomysis are often indistinguishable morphologically. When the restriction fragment data were analyzed using distance and parsimony phylogenetic methods a clade was found consistently, which contained E. arizonensis, E. albigulae, E. onychomysis, E. reedi, and E. papillata. This finding and other results of the phylogenetic analyses agreed and supplemented previous phylogenetic work on the Eimeria of rodents. Riboprinting appears to provide useful data for taxonomic and phylogenetic studies on the genus Eimeria and may be especially practical when samples do not contain enough oocysts for other molecular-based methods. PMID- 10540949 TI - Genus-specific PCR for the differentiation of eggs or larvae from gastrointestinal nematodes of ruminants. AB - Using sequences of the ribosomal second internal transcribed spacer (ITS2), PCR primers were designed for the differentiation of the gastrointestinal nematode genera Ostertagia, Cooperia, Nematodirus, Haemonchus and Trichostrongylus. Single eggs or larvae from faeces could be differentiated without previous DNA extraction. Quantification of the PCR result proved to be difficult because the DNA content between eggs from fresh or 24-h-old faeces varied considerably. PMID- 10540950 TI - Cryptosporidium is more closely related to the gregarines than to coccidia as shown by phylogenetic analysis of apicomplexan parasites inferred using small subunit ribosomal RNA gene sequences. AB - The phylogenetic placement of gregarine parasites (Apicomplexa: Gregarinasina) within the Apicomplexa was derived by comparison of small-subunit ribosomal RNA gene sequences. Gregarine sequences were obtained from Gregarina niphandrodes Clopton, Percival, and Janovy, 1991, and Monocystis agilis Stein, 1848 (Eugregarinorida Leger 1900), as well as from Ophriocystis elektroscirrha McLaughlin and Myers, 1970 (Neogregarinorida Grasse 1953). The sequences were aligned with several other gregarine and apicomplexan sequences from GenBank and the resulting data matrix analyzed by parsimony and maximum-likelihood methods. The gregarines form a monophyletic clade that is a sister group to Cryptosporidium spp. The gregarine/ Cryptosporidium clade is separate from the other major apicomplexan clade containing the coccidia, adeleids, piroplasms, and haemosporinids. The trees indicate that the genus Cryptosporidium has a closer phylogenetic affinity with the gregarines than with the coccidia. These results do not support the present classification of the Cryptosporidiidae in the suborder Eimerioirina Leger, 1911. PMID- 10540951 TI - Schistosomal granuloma modulation. III. Schistosma haematobium worms accelerate S. mansoni soluble egg antigen-induced hepatic granuloma formation in vivo. AB - Recurrent experimental evidence indicates that schistosomal egg granuloma formation at least in the murine model results from a host response generated against both egg- and worm-derived antigens. Further experiments aimed at identifying the existence in vivo of cross-sensitization between Schistosoma haematobium worms and S. mansoni-derived egg antigens were performed with respect to S. mansoni egg antigen-induced granuloma formation and fibrogenesis in the liver. Male OF1 mice bisexually infected with S. haematobium or S. mansoni were hepatically challenged (cecal vein injection) with S. mansoni SEA (soluble egg antigen)-coupled Sepharose beads at the end of prepatent infection (8-10 days prior to the start of egg deposition). The mean granuloma volume (MGV) of in-vivo generated synchronized hepatic granulomas (8 days old) and the fibrotic response were estimated. Just like S. mansoni-infected rodents, mice carrying an S. haematobium infection generated an accelerated hepatic granulomogenesis [respective MGVs 4.72 +/- 0.56 and 5.41 +/- 0.75 x 10(6) microm3; P < 0.0001 versus unsensitized (MGV 3.00 +/- 0.40 x 10(6) microm3) mice] and an enhanced fibrotic response against S. mansoni SEA. They also had significantly enlarged spleens (P < 0.0001) and moderately enlarged livers (P = 0.02) as compared with S. haematobium-infected mice that were not challenged with SEA. From these observations we infer that in vivo, S. haematobium worms can positively modulate S. mansoni egg antigen-induced granuloma formation and hepatic fibrogenesis, resulting in more severe liver pathology. PMID- 10540952 TI - Ultrastructure of Myxidium trachinorum sp. nov. from the gallbladder of the lesser weever fish Echiichthys vipera. AB - Myxidium trachinorum sp. nov. is described from the gallbladder of the lesser weever fish Echiichthys vipera. Pseudoplasmodia attach themselves to the gallbladder epithelium by filose processes, which are inserted between host cells. Pseudoplasmodia undergo endogenous cell formation at the secondary and tertiary levels. In the proliferative cycle, primary and endogenous cells are packed with digestive vacuoles formed by phagocytosis. In the sporogonic cycle the pseudoplasmodium becomes a pericyte enclosing two secondary cells (lacking digestive vacuoles) in a vacuole. These give rise to five cells each two valvogenic, two capsulogenic and a binucleate sporoplasm, which mature into spores. Comparison of the disporic M. trachinorum with polysporic species of Myxidium revealed significant differences in plasmodial ultrastructure, especially their attachments to host cells, surface characteristics and mode of nutrition, and in formation of generative cells. These suggest that the genus Myxidium may require revision. PMID- 10540953 TI - Enhanced absorption of pour-on ivermectin formulation in rats by co administration of the multidrug-resistant-reversing agent verapamil. AB - The effect of verapamil, a multidrug-resistance (Mdr)-reversing agent on the absorption of a pour-on formulation of ivermectin was evaluated in rats. Absorption of ivermectin was effectively enhanced (40%) by the presence of verapamil, suggesting that absorption of ivermectin involves Mdr-P-glycoprotein and that verapamil should act as a competitive inhibitor for the transport and extrusion of ivermectin by P-glycoprotein. This hypothesis is consistent with other studies describing verapamil as a blocking agent of P-glycoprotein involved in the efflux of ivermectin in a resistant strain of Haemonchus contortus. PMID- 10540954 TI - Characterization of a genomic region encoding the 32-kDa dense granule antigen of Sarcocystis muris (Apicomplexa). AB - Two subgenomic libraries constructed from Sarcocystis muris total DNA were screened with a cDNA probe, specific for a 32-kDa protein associated with the dense granules. Two clones reacted positively and were isolated, gDG 32/1 and gDG 32/2. Genomic clone gDG32/1 and part of clone gDG 32/2 have been sequenced. The composite nucleotide sequence of these genomic clones comprises 4.34 kb. It contains a 5' region of 2.14 kb, a first coding region (222 bp, exon I), a noncoding region (608 bp, intron), a second coding region (660 bp, exon II), and a 3' region of 693 bp. The upstream region shows a eukaryotic promoter structure and a consensus sequence for the 5' and 3' splicing sites. Thus the open reading frame (ORF DG32) coding for the 32-kDa protein of the dense granules of S. muris cyst merozoites is interrupted by an intron. To our knowledge, dg32 is the first sarcosporidian mosaic gene to be characterized. PMID- 10540955 TI - Prevalence of intestinal helminths of dogs and foxes from Jordan. AB - Necropsy of 340 stray and semi-stray dogs (Canis familiaris) and nine red foxes (Vulpes vulpes) from Jordan revealed that 239 dogs (70.3%) and all foxes were infected with at least one intestinal helminth species. No trematodes were found in the intestine of these hosts. The overall infection rates with cestodes, nematodes and acanthocephalans in dogs were 66.8%, 4.4% and 2.9%, respectively. The following cestodes were identified: Echinococcus granulosus (9.4%), Taenia pisiformis (11.8%), T. hydatigena (7.4%), T. ovis (4.4%), T. multiceps (3.8%), T. taeniaeformis (2.9%), Dipylidium caninum (19.4%), Joyeuxiella (3.2%), Diplopylidium (2.4%), and Mesocestoides (0.9%). Other intestinal worms in dogs were Toxascaris (2.6%), Toxocara canis (1.2%), and Protospirura (0.6%) nematodes, and gigantorhynchiid acanthocephalans (2.9%). Intestinal helminths found in foxes included cestodes (D. caninum, Joyeuxiella, Diplopylidium, Mesocestoides), nematodes (Protospirura, Uncinaria stenocephala and Oxynema) and an acanthocephalan (Macracanthorhynchus). In both hosts, most helminths were recovered from the second intestinal segment of four equally divided segments. PMID- 10540957 TI - Detection of early developmental stages of Myxobolus cerebralis in fish and tubificid oligochaete hosts by in situ hybridization. AB - The myxosporean and actinosporean spores of Myxobolus cerebralis develop through many stages in their respective hosts, salmonid fishes and a tubificid oligochaete. Using a modified, non-radioactive in situ hybridization protocol, the parasite, which exhibits radically different structural forms during its development in each host, could be specifically detected in paraffin-embedded tissues of both fish and oligochaetes. Our study aims to demonstrate the application of the technique for detection of early stages of M. cerebralis in both hosts. PMID- 10540956 TI - Differential sensitivity of erythrocytic stages of the rodent malaria parasite Plasmodium chabaudi chabaudi to dioncophylline B, a highly active naphthylisoquinoline alkaloid. AB - Four-week-old OF1 mice, infected with synchronized Plasmodium chabaudi chabaudi blood forms, were intraperitoneally injected with the naphthylisoquinoline alkaloid dioncophylline B (10 mg kg(-1) day(-1)) at three consecutive days. The respective groups were treated when rings, trophozoites, and schizonts were predominant. Microscopical observations of thin blood smears were made every two hours after the start of the experiment. A clear dependency of the effectiveness of dioncophylline B treatments on the timing of drug administration was demonstrated. Based upon the evolution of total parasitaemia and the survival rates, it was concluded that ring stages are insensitive to dioncophylline B, while the drug is highly effective when given at the trophozoite stage and partially effective when given at the schizont stage. Dioncophylline B seems to act by inhibiting the haemozoin degradation, as indicated by pigment clumping, and by impairing the segmentation of schizonts. PMID- 10540958 TI - The possibility of testing antiparasitical substances using the cockroach as a model. AB - Products used against parasitic arthropods must be tested for their efficacy. Because the supply of parasitic organisms for such tests may be a limiting factor, it is best to conduct these tests with a model organism. We show that the cockroach is a suitable model organism because it is easy to keep and to breed. Moreover, all stages of these animals are more resistant than, or at least as resistant as, the corresponding parasites, so the results with cockroaches can also be taken as valid for ectoparasites. PMID- 10540959 TI - Phenotypic analysis of peripheral lymphocyte subpopulations in hydatid patients. AB - Peripheral T-lymphocytes were analyzed in three groups of people: (1) individuals with current liver hydatid disease (hydatid patients, n = 20), (2) persons who had undergone surgical cyst removal at least 2 years previously (recovered patients, n = 9), and (3) a control group of healthy volunteers (uninfected controls, n = 13). Group 1 was subdivided according to cyst status, relapse of disease, and the presence or absence of symptoms. Percentages of lymphocytes expressing CD3, CD4, CD8, CD56, CD25, CD45RA, CD45RO, and HLA-DR were determined. Symptomatic patients had proportionally fewer CD3+ CD8 + lymphocytes than the control group (P=0.038). Hydatid patients with active cysts had proportionally more natural killer cells (CD56 + CD8-) than the control group (P = 0.028). PMID- 10540960 TI - Influence of olanzapine on cognitive functions and catalepsy in rats after single and chronic administration. AB - The aim of our investigations was to supplement the scarce information about behavioral effects of olanzapine and especially to find out if the effects change during prolonged treatment. It was established that at a dose of 0.5 mg/kg, which did not evoke sedation, olanzapine had distinct anxiolytic effects, both after acute and chronic treatment. Olanzapine improved memory in the maze test (food finding time), but only in chronic experiments after a lag of 14 days. Olanzapine at the dose of 0.5 mg/kg did not cause catalepsy but a higher dose of 1 mg/kg elicited strong sedating effects, which would interfere with the catalepsy test. The results of Porsolt's test (immobility time in swimming rats) were interesting because a shortening of immobility time occurred only after single drug injection. Our results are in agreement with the statement of other authors that olanzapine has greater potency in antagonizing responses mediated rather by 5-HT than D2 receptors. PMID- 10540961 TI - Modification of avoidance responding by amphetamine and dopamine receptor antagonists. AB - This study was designed to investigate the effects of preferential D1 (SCH 23390) and D2 (haloperidol) dopamine (DA) receptor antagonists and their interaction with low-dose (1 mg kg) D-amphetamine during the acquisition of two-way shuttle avoidance in rats. In the course of training, the dissociation of drug effects on response latencies was observed. Haloperidol (0.05 mg/kg) caused significant reduction in the frequency of short-latency avoidance responses that was only partially compensated by concomitant amphetamine administration. In contrast, SCH 2339 (0.025 mg/kg) did not affect the frequency of short-latency responses but lowered probability of avoidances emitted toward the end of 5 s interval between the onsets of conditoned and unconditioned stimuli (CS-UCS interval). Amphetamine compensated for this impairment by increasing frequency of short-latency avoidances well above the control level. These results argue for different nature of short- and long-latency avoidance responses, and suggest involvement of DA D2 receptors in the process of response initiation facilitated by amphetamine. Interestingly, a profound behavioral breakdown was observed under higher dose of SCH 23390 (0.05 mg/kg), but only when applied together with amphetamine. The latter result seems to confirm the notion that behavioral output of dopaminergic transmission may depend more on the balance between D1 and D2 receptors than on the independent modulation of particular receptor system. PMID- 10540962 TI - Effect of desipramine during infancy and preweanling on dorsal striatal dopamine D1 and D2 receptor binding in adolescence in the rat. AB - Desipramine, a monoamine uptake inhibiting antidepressant, was given once a day sc from the 6th to the 22nd postnatal days, which is during infancy and preweanling in ontogenesis, to rats and its effects on dorsal striatal dopamine D1 and D2 receptor binding in adolescence were examined. The rats were decapitated, their dorsal striata were dissected on approximately the 34th postnatal day, and the maximal densities (Bmax) and affinities (Kd) of dopamine D1 and D2 receptors were assayed using [3H] SCH 23390 and [3H] spiperone as ligands. Desipramine did not affect the densities or affinities of dopamine D1 or D2 receptor binding, and tended to increase the D1/D2 density ratio. The Bmax of dopamine D2 receptors was higher in male than in female rats (p<0.01), the D1/D2 Bmax ratio tended to be lower in male than in female rats (p<0.07), and the Kd for D1 receptors tended to be lower in male than in female rats (p<0.05). There was no interaction between treatment and gender. We conclude that desipramine given at infancy and preweanling increases the susceptibility of adolescent rats to behavioral effects of dopamine agonists with an optimal, and relatively high SCH 23390/spiperone binding site density ratio. Furthermore, gender differences in dorsal striatal spiperone binding site densities, SCH 23390/spiperone density ratio, and SCH 23390 affinities may render male rats more vulnerable than female rats to expression of excessive stereotyped behavior. PMID- 10540963 TI - Pharmacological profile of milnacipran, a new antidepressant, given acutely. AB - Pharmacological effects of acute treatment with milnacipran (MIL), a clinically active antidepressant (a noradrenaline [NA] and 5-hydroxytryptamine [5-HT] reuptake inhibitor without any affinity for neurotransmitter receptors) were studied in mice and rats. MIL inhibited the reserpine- or apomorphine-induced hypothermia in mice and enhanced the L-5-hydroxytryptophan-induced head twitches in rats. It reduced the immobility time in Porsolt's test in mice and rats, but either did not change the locomotor activity (mice) or decreased it (rats). MIL changed neither the clonidine-induced aggressiveness in mice nor the behavioral syndrome induced by oxotremorine in rats. The obtained results indicate that MIL, given acutely, shows a pharmacological profile similar to that of tricyclic NA and 5-HT reuptake inhibitors. In contrast to the antidepressants mentioned above, MIL does not exhibit an alpha1-adrenolytic or cholinolytic activity (in vivo tests). PMID- 10540964 TI - Effects of the NMDA/glycine receptor antagonist, L-701,324, on morphine- and cocaine-induced place preference. AB - Effects of the novel NMDA/glycine receptor antagonist, L-701,324, on morphine- and cocaine-induced conditioned place preference (CPP) were examined in male Wistar rats. After determination of initial preference, animals were conditioned with morphine (5 mg/kg, i.p.) or cocaine (5 mg/kg, i.p.) for 3 conditioning trials, alone or in combination of these drugs with L-701,324 (2.5 mg/kg and 5 mg/kg, p.o.). L-701,324 prevented acquisition of the place preference produced by morphine and cocaine. Administration of L-701,324 on the test day attenuated the expression of morphine-induced CPP, whereas it had no effect on cocaine CPP. When L-701,324 was given alone it did not affect dependent variables (i.e. time spent in non-preferred compartment) suggesting that L-701,324 did not display any reinforcing properties by itself. Our current data suggest that glycine site on the NMDA receptor complex may be involved in the mediation of the rewarding effects of drugs of abuse. PMID- 10540965 TI - Submandibular gland peptide-T (SGP-T) modulates ventricular function in response to intravenous endotoxin. AB - A novel peptide hormone isolated from salivary glands, submandibular gland peptide-T (SGP-T), protects against the enhanced hypotensive response to intravenously administered lipopolysaccharide (LPS; 3.5 mg/kg; Salmonella typhosa) in rats with their submandibular glands removed (sialadenectomy). In this study, we examined the effects of SGP-T on LPS-provoked perturbations of heart function. Sialadenectomy did not alter basal heart function, although the sialadenectomized rats exhibited more pronounced reductions in ventricular peak systolic pressure (VPSP), ventricular pressure at max dP/dt (Pmax dP/dt), and maximum ventricular negative dP/dt (-dP/dt) relative to unoperated controls following LPS challenge. These changes were primarily due to changes in afterload (blood pressure). However, in sialadenectomized rats LPS-induced changes in Pmax dP/dt (ventricular pressure at maximum -dP/dt) did not correlate with changes in VPSP, which suggests that sialadenectomy may alter the systolic component of the heart beat. The peptide SGP-T, at doses of 1 and 3.5 microg/kg, corrected Pmax dP/dt and all other endotoxin-induced changes in heart function. Since Pmax dP/dt is a measure of relaxation during diastole, the submandibular glands appear to play a role in protecting the heart against the adverse effects of acute endotoxin administration on ventricular relaxation. These effects of the submandibular glands may be mediated by the release of the peptide SGP-T. PMID- 10540966 TI - Effect of oxidative stress and erythropoietin on cytoskeletal protein and lipid organization in human erythrocytes. AB - Phenylhydrazine (PHX)-mediated damage in human red blood cells has been assessed by monitoring the release of tyrosine from cell proteins as well as using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). PHX-treated red blood cells exhibited concentration- and time-dependent tyrosine release. ATP has no effect on the release of tyrosine. This observation is supported by SDS-PAGE pattern of RBC membrane proteins, which shows a correlation between tyrosine release and cytoskeletal protein degradation. PHX requires the presence of erythrocyte cytosolic fraction for the degradation, possibly due to the presence of a proteolytic enzyme in the cytosol. PHX treatment renders the membrane proteins susceptible to the proteolytic attack. Treatment of PHX-exposed erythrocyte with bee venom phospholipase A2 induces the translocation of phosphatidylserine (PS) and phosphatidylethanolamine (PE) to the outer surface of the cell membrane. At the same time, phosphatidylcholine (PC) was translocated towards the inner surface, altering the membrane phospholipid asymmetry. Interestingly, increased tyrosine production followed by translocation of phospholipids across the red blood cell membrane by PHX treatment is completely inhibited by 0.2 units of erythropoietin (EP). Our findings suggest that exposure of red blood cells to an oxidant like PHX causes degradation of cytoskeletal protein by an ATP-independent proteolytic pathway and this in turn allows the transbilayer movement of phospholipids across the cell membrane. EP, by scavenging the hydroxyl radicals produced during interaction of PHX with red blood cells, protects the erythrocytes from oxidative attack. PMID- 10540967 TI - 9-substituted 1,2,3,4-tetrahydro-beta-carbolin-1-ones, new 5-HT1A and 5-HT2A receptor ligands. AB - Three series of new 9-substituted 1,2,3,4-tetrahydro-beta-carbolin-1-ones with 2 , 3- and 4-membered alkyl chain (1, 2 and 3, respectively) were synthesized, and the effect of some structural modifications on their 5-HT1A and 5-HT2A receptor affinities and functional in vivo properties was discussed. Radioligand binding measurements showed that the majority of compounds had a distinct affinity for 5 HT1A (1b, 2a, 2b, 2c, 3b; Ki = 0.3-64 nM) and 5-HT2A receptors (1b, 2b, 2c, 3b; Ki = 0.9-80 nM). The most potent 5-HT1A (1b, 2a, 2b, 3b) and 5-HT2A (1b, 2b, 3b) ligands were evaluated in in vivo tests. The obtained results indicate that 1,2,3,4-tetrahydro-beta-carbolin-1-ones containing 1-(o-methoxyphenyl)piperazine (1-3b) show pharmacological profile of 5-HT1A postsynaptic antagonists (with very weak agonistic component) and 5-HT2A antagonists, compound with 1,2,3,4 tetrahydroisoquinoline (2a) is a pure 5-HT1A postsynaptic antagonist. Summing up, the connection of 1,2,3,4-tetrahydro-beta-carbolin-1-one moiety through the 2-4 membered alkyl spacer with 1-(o-methoxyphenyl)-piperazine, which is present in a variety of 5-HT1A ligands, allowed us to obtain the compounds with high and equal affinity for 5-HT1A/5-HT2A receptors and the expected functional properties, i.e. distinct antagonistic and weak agonistic activity at 5-HT1A postsynaptic receptors and antagonistic at 5-HT2A ones. PMID- 10540969 TI - Comparison of effectiveness of two schedules of rapid transcranial magnetic stimulation on enhancement of responsiveness to apomorphine. AB - Locomotor response to apomorphine (0.5 mg/kg s.c.) was tested in rats pre-treated with multiple rapid transcranial magnetic stimulation (rTMS) or electroconvulsive shock (ECS). rTMS (B = 1.6T, f = 20 Hz, t = 300 s) was delivered 9 times during 18 days (group rTMS1, total number of stimulations 54,000) or 18 times during 18 days (group rTMS2, total number of stimulations 108,000); ECS (I = 150 mA, f = 50 Hz, t = 500 ms) was delivered 9 times at 48 h intervals. Apomorphine given 24 h after the last treatment induced hyperactivity that was significantly more pronounced in rats receiving ECS and rTMS. The effect was most pronounced for electroconvulsive treatment, the least in the rTMS1 group. The results indicate that an increase in the total number of impulses during the rTMS may increase the responsiveness of rats to dopaminergic stimulation to the level comparable with that induced by ECS. PMID- 10540968 TI - Valproate- and aminophylline-induced 'wet dog shakes'--a function of dose and time. AB - The time course of appearance of 'wet dot shakes' (WDS) was examined following valproic acid (VPA, 100, 200, 300, 400 mg/kg i.p.) and aminophylline (AMP, 50, 100, 150 mg/kg i.p.) injections. VPA and AMP at various doses showed a qualitative difference in their ability to induce WDS with no difference in intensity, confirming 'all or none' nature of the phenomenon. There was a significant (p<0.001), dose-dependent increase in the number of whole body shakes following first three doses of VPA but not after the administration of its highest dose (400 mg/kg). In contrast, the numbers of WDS produced by AMP were inversely proportional to its increasing doses. The maximum numbers of WDS were observed at 300 mg/kg of VPA and 50 mg/kg of AMP, within 10 min and 20-30 min during 1 h and 1 h 30 min observation period, respectively. The present stereotyped behavior induced by acute, single dose administration of VPA and AMP in non-toxic doses, being a reproducible phenomenon, lasting for a brief period may be anticipated to serve as a tool to explore mechanisms underlying WDS. PMID- 10540970 TI - Participation of opioid mechanism in the antinociceptive effects induced by oxaprotiline enantiomers in mice. AB - The purpose of the present study was to assess the activity of (+)-oxaprotiline [(+)-OXA] (a noradrenaline uptake inhibitor) and (-)-oxaprotiline [(-)-OXA] (with unknown mechanism of action) in two experimental models of pain in mice, a hot plate test and a writhing syndrome induced by phenylbenzoquinone (PHBQ), and to determine whether the opioidergic system may be engaged in their antinociceptive effects. Morphine was used as a reference drug. Administration of (+)-OXA (0.31-5 mg/kg) and (-)-OXA (20 mg/kg) produced a statistically significant elevation of the nociceptive threshold, measured by the increased latencies in the hot plate test. Moreover, (+)-OXA (0.62-5 mg/kg) and (-)-enantiomer (5-20 mg/kg) decreased the number of writhing episodes induced by PHBQ in mice, (+)-enantiomer being more effective than (-)-OXA in either test. In the hot plate test, the analgesic effect induced by (+)-OXA (0.31 mg/kg) or (-)-OXA (20 mg/kg) was abolished by naloxone (2 mg/kg), an opioid receptor antagonist. In the writhing test, naloxone (2 mg/kg) partially, but not significantly, reduced the antinociceptive responses induced by (+)-OXA (0.62 mg/kg) or (-)-OXA (5 mg/kg). The obtained results show that both OXA enantiomers produce antinociception in mice which can be, at least partially, connected with opioid system. PMID- 10540971 TI - Cohort changes in illegal drug use among arrestees in Manhattan: from the Heroin Injection Generation to the Blunts Generation. AB - This paper identifies three inner-city cohorts differing by birth year and preferred drugs that routinely passed through Manhattan's criminal justice system from 1987 through 1997: The Heroin Injection Generation born 1945-54, the Cocaine/Crack Generation born 1955-69, and the Blunts (marijuana plus tobacco) Generation born since 1970. The future prospects for the Blunts Generation may be modestly enhanced by their continued avoidance of cocaine, crack, and heroin- despite the fact that many of them are being reared in severely distressed households and are developing few skills for legal jobs. PMID- 10540972 TI - Drug use, AIDS knowledge, and HIV risk behaviors of Cuban-, Mexican-, and Puerto Rican-born drug injectors who are recent entrants into the United States. AB - To date, relatively little research attention has been devoted to the HIV-risky behaviors of persons who are newly arrived in the United States and who use drugs. Data gathered from street-recruited injection drug users (IDUs) recruited in 10 United States cities who were born in Mexico, Cuba, and Puerto Rico and who are recent entrants into the United States suggest that, in comparison to US-born IDUs, Mexican-born subjects are at elevated risk for acquiring and transmitting HIV as a result of sharing needles with friends and running partners; sharing drug injection implements such as cookers, cotton, and rinse water; frequent injection in HIV-risky settings; use of unsterilized needles; and relatively frequent trading of sex for drugs or money. Puerto-Rican-born IDUs were found to inject drugs relatively frequently, and to do so relatively often in high-risk settings in which sterile injecting equipment and cleaning materials often are scarce. These data also show generally lower levels of AIDS knowledge among the in-migrant IDUs than among US-born IDUs. Respondents from each nationality group most often cited television as the source of their most useful and reliable AIDS information, but also tended to regard community outreach workers as a significant source of reliable AIDS and needle cleaning information. The high levels of involvement in HIV-risky behaviors, deficits in knowledge concerning the means of HIV transmission, and relative ease of mobility of the at-risk (for HIV) individuals examined here indicate a need for a comprehensive public health prevention initiative to limit the future spread of HIV. At a minimum, such an undertaking would do well to incorporate group-specific, culturally appropriate behavioral interventions as well as an information campaign. PMID- 10540973 TI - Relapse patterns among adolescents treated for chemical dependency. AB - This study examined levels, timing, and patterns of posttreatment alcohol and drug use in adolescents (n = 113) during the 12 months following completion of primary treatment. Data were collected from clinical records, random urine screens, and interviews with the adolescents and their parents. In all, 61.1% relapsed to pretreatment levels of use during the 12 months after treatment, 79.6% of the males and 59.3% of the females. Of those with alcohol as the drug of dependence, 45.9% relapsed to pretreatment levels of use in 12 months. Likewise, 75.0% marijuana users, 70.6% combined alcohol and marijuana users, and 50.2% other drug users relapsed to pretreatment levels of use in the 12 months. Relapse curves are presented to demonstrate the specific timing and patterns of relapse. Implications for primary treatment and aftercare are discussed. PMID- 10540974 TI - Identifying offspring of problem-drinking parents: comparison of five self-report measures. AB - Although there is general consensus that self-report measures are reliable in offspring identification of parental problem drinking, studies in which these measures are used differ in two important ways: 1) different self-report measures are used across investigations, and 2) when identical measures are used, idiosyncratic cutoff criteria are employed. The purpose of this study was to compare five self-report measures commonly used in college-age populations to identify problem-drinking parents. When the most conservative criterion was employed, each of the five measures identified similar percentages of offspring as having problem-drinking parents (10% for fathers and 4% for mothers). Interrelationships among the five measures were examined, and each method appeared to contribute both to the common and unique variance of the construct "parental problem drinking." Therefore no one measure can capture all aspects of a parent's drinking problem as reported by their offspring. PMID- 10540975 TI - Community-based intervention to reduce demand for drugs in Northern Thai tribal villages. AB - This is an evaluation study of a community-based intervention model used in a project designed to reduce the demand for and use of opium, heroin, and other drugs among 85 tribal villages located in Northern Thailand. The Integrated Drug Abuse Prevention (IDAP) Project was conducted from 1995 to 1997 and used a community-based approach which included innovative methods such as multimedia awareness raising campaigns, networking between villages and local government agencies, and village-based drug detoxification and treatment to assist villages in solving their drug problems. The intervention model was successfully implemented in most villages and demonstrated very good results in improving awareness, decreasing the number of active drug users living in the villages, and preventing new cases of addiction. However, a follow-up study at 6 months after project termination indicated problems with sustainability of demand reduction activities and outcomes. These problems were attributed in part to a lack of empowerment among village leaders to continue activities without assistance from project staff. Also, village leaders expressed problems in resisting drug dealers who returned to the area, which suggested that support from law enforcement is critical to the viability of drug demand reduction programs. PMID- 10540976 TI - Use of AUDIT for alcohol screening among emergency room patients in Thailand. Alcohol Use Disorders Identification Test. AB - This study evaluated the Alcohol Use Disorders Identification Test (AUDIT) against blood alcohol levels and medical diagnoses. The population under study included 695 current drinkers admitted to emergency rooms of four regional Thailand hospitals. The AUDIT positivity rate was 61% among 343 patients who drank prior to admission and 32% among 352 patients who did not drink alcohol before admission. Breath alcohol levels were positively associated with AUDIT scores. The sensitivity against a previous or current alcohol-related medical diagnosis was 89%. We concluded that the AUDIT is a satisfactory instrument for alcohol screening in this population. PMID- 10540977 TI - Does Alcoholics Anonymous work? The results from a meta-analysis of controlled experiments. AB - This article reviews the outcome (usually abstinence at 12 months) of 21 controlled studies of AA, with emphasis on methodological quality. Severe selection biases compromised all quasi-experiments. Randomized studies yielded worse results for AA than nonrandomized studies, but were biased by selection of coerced subjects. Attending conventional AA meetings was worse than no treatment or alternative treatment; residential AA-modeled treatments performed no better or worse than alternatives; and several components of AA seemed supported (recovering alcoholics as therapists, peer-led self-help therapy groups, teaching the Twelve-Step process, and doing an honest inventory). PMID- 10540978 TI - Syringe disposal options for injection drug users: a community-based perspective. AB - This study was a qualitative exploration of syringe disposal interventions for injection drug users (IDUs). Data were collected through in-depth interviews with 26 community members who injected drugs and 32 noninjecting community members in Atlanta, Georgia. Both groups supported syringe exchange programs as syringe disposal interventions, while noninjecting community members favored a one-way drop box. IDUs identified fear of arrest for possession of syringes as the most salient barrier to safe syringe disposal, revealing the negative consequences of drug paraphernalia laws. PMID- 10540979 TI - Proximate composition and mineral content of two edible species of Cnidoscolus (tree spinach). AB - Proximate composition and mineral content of raw and cooked leaves of two edible tree spinach species (Cnidoscolus chayamansa and C. aconitifolius), known locally as 'chaya', were determined and compared with that of a traditional green vegetable, spinach (Spinicia oleraceae). Results of the study indicated that the edible leafy parts of the two chaya species contained significantly (p<0.05) greater amounts of crude protein, crude fiber, Ca, K, Fe, ascorbic acid and beta carotene than the spinach leaf. However, no significant (p>0.05) differences were found in nutritional composition and mineral content between the chaya species, except minor differences in the relative composition of fatty acids, protein and amino acids. Cooking of chaya leaves slightly reduced nutritional composition of both chaya species. Cooking is essential prior to consumption to inactivate the toxic hydrocyanic glycosides present in chaya leaves. Based on the results of this study, the edible chaya leaves may be good dietary sources of minerals (Ca, K and Fe) and vitamins (ascorbic acid and beta-carotene). PMID- 10540980 TI - Effect of blanching time and salt concentration on pectolytic enzymes, texture and acceptability of fermented green beans. AB - Effect of blanching time, salt concentration and fermentation on texture of green beans (var. Tuf) was determined. Beans were blanched at 90 degrees C for 1, 10 or 20 minutes and fermented with brine containing 3.08 and 4.6% salt (corresponding to 1.2 and 1.8% salt in the finished product). The beans were assessed for texture mechanically using Universal Testing Machine (Model 1011, Instron). Sensory evaluation was carried out to assess the taste, texture and overall acceptability of the products. Pectolytic enzyme (pectinestarase and exopolygalacturonase) activities were determined in fresh and fermented beans. Increase in blanching time improved the texture significantly. Significant (p<0.05) decreases in shearing energy of the beans were observed in the first two days of fermentation and, thereafter, there were no significant (p>0.05) changes except for samples blanched for one minute. Salt concentration showed a small but significant effect on texture only in samples blanched for one minute and this was probably due to activation by salt of residual pectolytic enzymes in the beans. Most acceptable beans were those blanched for 20 minutes and fermented with 1.2% salt brine. PMID- 10540981 TI - Protein evaluation of four oat (Avena sativa L.) cultivars adapted for cultivation in the south of Brazil. AB - Four oat cultivars adapted for soil and climate conditions in the southern region of Brazil were evaluated for protein nutritive value. Evaluations were done both in vitro and in vivo. In vitro evaluation was done by essential amino acid profile, available lysine, amino acid scoring, and protein digestibility corrected amino acid-scoring (PDCAAS). Nitrogen balance indices and PER were determined in vivo with rats. In all four cultivars (UFP-15, UFP-16, CTC-03, UFRGS-14), lysine was the most limiting amino acid. Available lysine, amino acid score and PDCAAS were highest for cultivar UFRGS-14 and lowest for CTC-03. When compared to casein, only nitrogen retention for UFRGS-14 did not differ statistically (p>0.05); all other indices of protein quality were inferior to casein for the oat cultivars. The oat cultivars tended to be identical among themselves, except for apparent protein digestibility which was significantly higher in the UFRGS-14 and CTC-03 cultivars. On average, the PER values of the oat cultivars were 82% of casein; the net protein utilization was 88% of casein as determined in vivo and 49% by the estimation in vitro (PDCAAS). PMID- 10540982 TI - Environmental variables affect the hard-to-cook phenomenon of cowpea (Vigna unguiculata) seed. AB - Changes in moisture content and cooking rate of cowpea (Vigna unguiculata) seeds which were sun-dried for 5 hours on cement, wood, or corrugated iron sheet surfaces and packaged for 6 months in jute or polythene bags were studied. Relationships and effects of interacting variables studied were examined using the contrast analysis technique. From day zero to about 2 months of storage, the sun-dried samples had significantly (p<0.01) lower moisture content and longer cooking times than the corresponding control samples. However, moisture-gain and cooking time increased progressively throughout the storage period for all samples studied. The relationship between these two variables, tested at p = 0.01 using contrast analysis technique, was dependent on the choice of packaging material. PMID- 10540983 TI - Production and organoleptic assessment of akara from bambara groundnut (Voandzeia subterranea (L.) Thouars). AB - Bambara groundnut (BGN) seeds were pretreated by soaking, dry-milling or autoclaving and used to produce pastes which were then used to prepare akara. Proximate analysis and organoleptic tests were conducted. Method of pretreatment affected the proximate composition of the akara. The moisture, protein and fiber contents of akara prepared from BGN seeds, which were cracked and soaked or autoclaved, were significantly (p< or =0.05) different than those from soaked whole grains and flour. The cooked batch weights of the akara ranged from 258-272 g, akara from soaked whole and autoclaved seeds having the lowest and highest values, respectively. Akara from autoclaved BGN seeds was more highly preferred by panelists compared to akara from other pretreatments. Except for appearance and color, no significant differences were found between BGN akara and the cowpea akara used as reference. Heat treatment of BGN seeds prior to dehulling appeared to influence the level of acceptability of the akara. PMID- 10540984 TI - Effects of infestation on the nutrient content and physiocochemical properties of two cowpea (Vigna unguiculata) varieties. AB - Two cowpea varieties (Ife-brown and Kano-white varieties) were used for the study. The effects of insect infestation on the chemical composition and physicochemical properties of these cowpea seeds were studied. The proximate composition, mineral content, total starch, total soluble sugars, bulk density, fat and water absorption capacities, viscosity, gelation capacity and emulsion properties of infested cowpea varieties were compared with those of uninfested cowpeas. Effects of infestation on nitrogen solubility and on the protein fractions were also determined. Infestation depleted the protein, starch and soluble sugar contents of cowpeas. Oil and water absorption capacities were increased while emulsification, foam and viscosity properties were reduced. The nitrogen solubility pattern was altered. Uninfested Kano-white cowpeas (UKW) possessed better foam properties than uninfested Ife-brown cowpeas (UFB). On the other hand, UFB had better emulsification properties than UKW. PMID- 10540985 TI - Changes in nitrogenous and other chemical constituents, protein fractions and in vitro protein digestibility of germinating fluted pumpkin (Telfairia occidentalis Hook) seed. AB - The effect of 7 days of germination on levels of nitrogenous and other nutrition related parameters, protein fractions and in vitro protein digestibility of fluted pumpkin (Telfairia occidentalis) seed was studied. The non-protein nitrogen gradually increased and the protein nitrogen content decreased during germination. Albumin and globulin fractions were found to be the major seed proteins of fluted pumpkin seeds, constituting about 58.6% of the total protein of the ungerminated (raw) seeds. The protein fractions, albumin and glutelin, were observed to increase by 61.5% and 57.0%, respectively, while a 54.6% decrease was noted in the prolamine fraction. The globulin fraction increased at the beginning of germination but decreased at the end. Germination significantly (p< or =0.05) increased the crude protein, nitrogen solubility and in vitro protein digestibility but decreased the fat, phytic acid and polyphenol contents of the seeds. PMID- 10540986 TI - Effect of plant fruits--Indian gall nut, bedda nut and gooseberry--on hypercholesterolemic rats. AB - The effect of supplementation of three fruits, Indian gall nut, bedda nut and gooseberry, on serum lipid levels and excretion of bile acids was investigated. Rats made hypercholesterolemic by feeding hypercholesterolemia inducing diet (HID) for a period of 30 days were used as the test model. Feeding of a dried powder of these fruits along with the HID resulted in significant (p<0.01) reduction in total cholesterol, LDL cholesterol and triglycerides. HDL cholesterol remained unchanged in groups fed gall nut and bedda nut. However, the levels were significantly (p<0.01) higher in groups fed mixed and gooseberry diets in comparison to the control diet. Excretion of bile acids was found to be significantly (p<0.01) higher in animals receiving the three fruits in combination in comparison to those receiving the individual fruits. PMID- 10540987 TI - Comparative effects of scopoletin and cyanide on rat brain, 1: histopathology. AB - Four week old male Wistar rats were used to study the effects of scopoletin and cyanide on the histopathology of rat brain. The rats were divided into a control and three experimental groups (2-4) and fed rations containing 0.07 microg scopoletin/100 g, 0.07 microg scopoletin + 1.8 mg cyanide/100 g and 1.8 mg cyanide/100 g, respectively. These levels of scopoletin and cyanide corresponded to levels found in a processed cassava diet. The first group was fed the same ration as the others but without scopoletin and cyanide. The rats were fed these rations for twelve months. Rats from each group were sacrificed at the third, sixth, ninth and twelfth months; the relative brain weight of the rats (% of body weight) and histology of their brains were studied. The lipid peroxide levels of the rat brains were also studied at the twelfth month. The results showed that the relative brain weights of the rats fed scopoletin + cyanide were significantly (p<0.05) less than that of the control from the third month. There were no significant changes in the lipid peroxide levels of the rat brains in the various groups. Histological examination of the brains of the rats suggested that scopoletin is involved in the pathogenesis of the neuropathy seen in cassava consuming populations. PMID- 10540988 TI - Hypocholesterolemic effect of germinated fenugreek seeds in human subjects. AB - The effect of consumption of germinated fenugreek seed powder at two different levels, i.e., 12.5 g and 18.0 g on the blood lipid profiles of twenty hypocholesterolemic adults of both sexes in the age range of 50-65 years was studied. The subjects were divided into two groups, i.e., Group I and Group II who were asked to incorporate the powder into any dish of their choice at the rates of one packet per day containing 12.5 g and 18.0 g of the germinated powder, respectively, for a period of one month. Fasting blood was drawn intravenously one day before and at the end of 30 days feeding trials. The findings revealed that germination had brought distinct changes in soluble fiber content of the seeds. Consumption of the seed at both the levels resulted in a hypocholesterolemic effect. Between the two levels, higher levels of consumption, i.e., 18.0 g of the germinated seed resulted in a significant reduction in total cholesterol and LDL levels. No significant changes were found in HDL, VLDL and triglyceride levels in all the subjects. PMID- 10540990 TI - Biotechnological production of oil: fatty acid composition of microbial oil. AB - The fatty acid profile and the fatty acid composition of microbial lipids obtained from molds revealed that oil from Aspergillus sydowii, Fusarium oxysporum and F. equisetti had a high percentage of unsaturated fatty acids, particularly oleic acid, and had a similarity to the edible oils, groundnut and palm oil. This study sheds light on the possibilities of exploring the use of these oils as supplement to other edible fats and for other non-edible industrial purposes. PMID- 10540991 TI - International issue. PMID- 10540989 TI - The effect of processing on total organic acids content and mineral availability of simulated cassava-vegetable diets. AB - Changes in pH, titratable acidity and mineral content (Ca, Fe, Mg, Zn) were estimated in processed cassava products while the mineral content of raw and blanched amaranthus vegetable was determined. pH of fresh cassava (6.5) decreased as total organic acid (0.07%) increased with fermentation period. Fufu and lafun had the lowest pH and the highest total organic acids contents. Fermentation of cassava increased the total calcium and iron contents, reduced magnesium level while zinc remained fairly constant in grated cassava but was reduced when soaked in water (for the preparation of fufu and lafun). Fermentation also increased the availability of these selected minerals in both cassva products and simulated cassava-vegetable diets. Blanching reduced the mineral content of amaranthus vegetable but increased mineral availability. Fermentation of cassava and blanching of vegetables play an important role in making minerals more available and these processing methods should be encouraged to potentially ameliorate the disease states associated with mineral deficiency. PMID- 10540992 TI - Lengthening of the lateral column and reconstruction of the medial soft tissue for treatment of acquired flatfoot deformity associated with insufficiency of the posterior tibial tendon. AB - We analyzed our results of surgery for acquired flatfoot deformity after dysfunction of the posterior tibial tendon. This included lengthening the proximal lateral column by calcaneal osteotomy and reconstructing the medial soft tissue. Nineteen patients (9 women and 10 men; average age, 52.9 years [range, 24 72 years]) were treated for stage II and stage II-III insufficiency of the posterior tibial tendon. The medial soft tissue surgery included 18 reconstructions of the tendon, 11 transfers of the flexor digitorum longus tendon, 13 repairs of the deltoid ligament, and 3 repairs of the spring ligament. At follow-up (mean, 23.4 months), all patients had satisfactory restoration of their medial longitudinal arch, reduction of abduction in the forefoot, and restored height in the arch. All patients were able to bear weight fully on the foot that underwent surgery, and all but one were satisfied with the result achieved. The clinical result was rated as excellent in 6, good in 11, and fair in 2 cases. In all but one case, no loss of achieved correction in the foot was found. In one case, the calcaneocuboid joint had to undergo arthrodesis after 5 months because of painful degenerative joint disease. In the pes planovalgus and abductus deformities occurring in stage II disease, calcaneal osteotomy and reconstruction of the medial tendon and ligament seem to play a significant role in operative management. This was the case only when degenerative joint disease and significant subluxation of the subtalar or talonavicular joint or both had not already occurred. They seem to function by restoring more normal biomechanics, which allows reconstructed or transferred tendon to function successfully. PMID- 10540993 TI - Metatarsal neck osteotomy with rigid internal fixation for the treatment of lesser toe metatarsophalangeal joint pathology. AB - Metatarsalgia associated with metatarsophalangeal (MTP) joint instability and/or plantar callosity formation is a difficult problem to treat. During a 15-month period, we performed 50 osteotomies of the metatarsal neck with rigid internal fixation in 47 feet of 42 patients. Three patients were excluded from the study, leaving 47 osteotomies in 44 feet of 39 patients for review. There were 6 men and 33 women, with a mean age of 57 years. In addition to lesser MTP joint pain with or without instability, the majority of patients had first ray pathologic condition, which was also addressed at the time of surgery. All but one of the osteotomies were united radiologically at 6 weeks. The mean shortening was 4.1 mm (range, 2-12 mm), and the mean follow-up was 9 months. There were no cases of malunion, nonunion, or avascular necrosis. At follow-up, 33 patients were asymptomatic. Eight patients (nine feet) had a degree of persisting pain at follow-up (seven mild and two moderate), but the source of this pain was only the metatarsal or MTP joint that was operated on in three cases. In this article, we describe the indications, the technique, and the results of the osteotomy. PMID- 10540994 TI - Clinical and functional outcome after anatomic and nonanatomic ankle ligament reconstruction: Evans tenodesis versus periosteal flap. AB - The present study investigated the effects of two different surgical procedures for the treatment of chronic ankle instability. Ten patients treated with an anatomic reconstruction using a periosteal flap were compared with a second group that received an Evans tenodesis. All patients were evaluated before and after surgery with clinical and radiographic examinations as well as dynamic pedobarography. Patient satisfaction and radiographic and functional results were comparable in both groups and revealed a good restoration of joint stability and gait symmetry. Our results indicate that both methods of ankle ligament reconstruction achieve a comparable clinical and functional outcome within 1 year after surgery. PMID- 10540995 TI - Hindfoot alignment of hallux valgus evaluated by a weightbearing subtalar x-ray view. AB - A new radiographic view was proposed to evaluate alignment of the hindfoot under weightbearing condition. The ankle joint and the middle and posterior facets of the subtalar joint were clearly visualized in all radiographs. A comparative study was made of 104 feet with hallux valgus in 58 female patients and 67 normal feet in 57 normal female subjects (control group). The mean value of the angle between the axis of the tibia and a line on the surface of the ankle joint on the talus was significantly larger in the group with hallux valgus than in the control group. Likewise, the mean value of the angle between the axis of the tibia and a line on the surface of the posterior facet of the subtalar joint on the calcaneus in the group with hallux valgus was 95.3 degrees, significantly larger than the 87.9 degrees in the control group. These findings showed that the ankle joint and the posterior facet of the subtalar joint in hallux valgus have valgus deviation. The hindfoot in a foot with hallux valgus has a tendency toward pronation. No previous study has measured the inclination of the posterior facet of the subtalar joint directly in weightbearing. PMID- 10540996 TI - Comparison of the reliability of plantar pressure measurements using the two-step and midgait methods of data collection. AB - The reliability of plantar pressure measurements obtained using the traditional midgait method compared with the two-step method was investigated. Using an EMED SF system, the parameters of contact area, contact time, maximum force, and peak pressure at seven sites of the foot were assessed for reliability of measurement in 10 normal subjects. The results of the study indicate that the two-step method is as reliable as the midgait method and may be preferred for use in both research and clinical settings. PMID- 10540997 TI - Mobility of the first tarsometatarsal joint in relation to hallux valgus deformity: anatomical and biomechanical aspects. AB - Hypermobility of the first tarsometatarsal (TMT 1) joint is suggested to be an important factor in the cause and progression of hallux valgus deformity. Hypermobility of the TMT 1 joint is tested clinically in the sagittal plane, but an important deformation also exists in the transversal plane: metatarsus primus varus. This in vitro study was undertaken to investigate the relation between the mobility of the TMT 1 joint in these two planes and to investigate the correlation of the mobility with morphological variables. A second aim was to study the possible stabilizing effect of the tibialis anterior muscle, flexor hallucis longus muscle, and peroneus longus muscle on the TMT 1 joint. Nine embalmed human specimens were tested under standardized conditions. A 30-N force was applied to the head of the first metatarsal (MT 1) to pull in either the dorsal or medial direction. To simulate muscle force, 21 N was applied to the three tendons: all seven possible combinations of muscle action were tested in each plane of motion. Angular displacements were measured using 2-dimensional LED video registration. TMT 1 mobility is a relevant factor in MT 1 mobility in the sagittal and transversal planes, the peroneus longus has a stabilizing effect on this joint, and the effect of the flexor hallucis longus on this joint is different in both planes. When considering a Lapidus procedure for surgically correcting a hallux valgus, the mobility of MT 1 in the transversal plane should also be assessed, but so far no objective clinical test in this plane has been described. PMID- 10540998 TI - Arthrodesis of the distal tibiofibular joint for a large osteochondroma in an adult. AB - A case of a large osteochondroma of the distal tibia with distortion of the distal tibiofibular joint is presented. This could not be managed by traditional means, as excision would have resulted in ankle and tibiofibular joint instability. The problem was overcome by performing an arthrodesis. Only enough bone from both the tibia and the fibula was excised to provide a host bed for bone graft. We believe that symptomatic osteochondromata should usually be excised. However, if this would result in damage, then the method described offers an alternative management strategy. PMID- 10540999 TI - Peroneus quartus muscle: a case report and review of the literature. AB - The peroneus quartus is a supernumerary muscle of the lateral compartment of the lower leg. There are several variations of this accessory muscle, and its terminology is very confusing. The peroneus quartus is found with a frequency varying from 10 to 21.7% of observed individuals. It is rarely involved in pathologic processes of the foot and ankle. Only a few reports exist in the literature involving peroneus quartus in a "retromalleolar conflict" (lateral ankle stenosis), sometimes in association with longitudinal attrition and tears of the peroneus brevis. We present a novel case, together with a review of the clinical, anatomic, and radiologic references about this muscle, and we try to make order in classification and terminology of its different forms. PMID- 10541000 TI - The mechanical properties of the heel pad in unilateral plantar heel pain syndrome. AB - Plantar heel pain syndrome has been attributed to entrapment neuropathy, plantar fasciitis, calcaneal spurs, and stress fractures of the calcaneus. Although deteriorated mechanical properties of the heel pads may play an important role in the pathogenesis of heel pain syndrome, this has received little notice. In this study, a specially designed compression relaxation device with a push-pull scale and a 10-MHz linear array transducer was used to determine thickness of the heel pad under different loading conditions. Twenty consecutive patients aged 29 to 77 years with unilateral plantar heel pain syndrome were enrolled. Thickness of heel pad bilaterally was measured when the heel pad was compressed by serial increments of 0.5 kg to a maximum of 3 kg and then relaxed sequentially. The load displacement curve during a loading-unloading cycle was plotted, and the compressibility index and energy dissipation ratio of the heel pad were calculated accordingly. Phase I displacement of the heel pad (from 0 to 1 kg load) on the painless side was greater than that on the painful side (P < 0.01), but there was no statistically significant difference between painless and painful sides in thickness of unloaded heel pads, compressibility index, or energy dissipation ratio (P > 0.05). In conclusion, the affected heel pad in plantar heel pain syndrome was stiffer under light pressure than the heel pad on the painless side. The changed nature of chambered adipose tissue in a painful heel pad may be responsible for its increased stiffness under light pressure. PMID- 10541001 TI - Is the course of brain development in schizophrenia delayed? Evidence from onsets in adolescence. AB - A degree of ventricular enlargement, together with a reduction of total cortical mass and loss of asymmetry is reported in schizophrenia, but the meaning is obscure. These changes may reflect an anomaly of brain development. Brain structure was assessed on a 1.5-Tesla MRI scan in a series of 29 adolescents at the time of a first episode of schizophrenia and compared with 15 adolescents with other serious psychiatric disturbance (mostly psychotic) and 20 normal adolescent controls. The age at scan ranged between 13 and 20 years. In the adolescents with a diagnosis of schizophrenia, total brain volume increased with age in a way that differed significantly (p=0.007) from that seen in patients with other psychiatric disturbance and normal controls. Thus, brain growth, as assessed by this index, had reached a plateau in the control group by the age of 13 years, but this was not true of patients with schizophrenia. The measure that most clearly distinguished the groups (p<0.001 after co-varying for height and sex) was the volume of the left lateral ventricle the ventricle was significantly larger in patients with schizophrenic illness, and ventricular size increased with age to a greater extent in the patient group, although not significantly so, than in normal controls. Thus, aspects of brain growth are delayed in patients with early onset schizophrenia, and the greatest severity of illness is reflected in a component of growth that is lateralized to the dominant hemisphere. Individuals who develop serious psychiatric illness, including schizophrenia, represent a fraction of the population in whom a component of the relative development of the cerebral hemispheres occurs late. PMID- 10541002 TI - Ventriculomegaly and reduced hippocampal volume following intrauterine growth restriction: implications for the aetiology of schizophrenia. AB - Structural alterations in the brains of some schizophrenic patients suggest an impairment of brain development, possibly as a result of intrauterine compromise. In this study we have tested the hypothesis that placental insufficiency during the second half of pregnancy in the guinea pig results in structural alterations similar to those seen in some schizophrenic patients. Placental insufficiency was induced in pregnant guinea pigs via uterine artery ligation at midgestation. At 60 days gestation (term: 68 days gestation) the fetal brains were prepared for quantitative histological and immunohistochemical analysis and compared with controls. Placental insufficiency resulted in growth-restricted animals with significantly larger cerebral ventricles, reduced cross-sectional area of the cerebral cortex and the striatum and reduced hippocampal volume compared with controls. There were fewer neuronal nitric oxide synthase (nNOS)-positive cells in layers 5-6 of the cingulate cortex, and in layer 1 of the frontal and temporal cortices. In contrast, there were no significant alterations in the optical density of tyrosine hydroxylase (TH), a rate-limiting enzyme in the biosynthesis of catecholamines and the dopamine transporter (DAT) in the striatum in growth restricted animals compared with controls. These findings indicate that developmental disturbances can produce anatomical changes that resemble those found in some individuals with schizophrenia. PMID- 10541003 TI - The synaptic-vesicle-specific proteins rab3a and synaptophysin are reduced in thalamus and related cortical brain regions in schizophrenic brains. AB - Two synaptic-vesicle proteins, rab3a and synaptophysin, have been studied on post mortem brain tissues of schizophrenics and healthy controls. We found significantly reduced levels of rab3a in thalamus (p<0.001); for both proteins in gyrus cinguli and hippocampus (p<0.0001); for rab3a in frontal and parietal cortex (p<0.05); and no differences in temporal cortex or cerebellum in schizophrenics compared with controls. Reduced synaptic density may be a prominent feature of the molecular neuropathology of schizophrenia. PMID- 10541004 TI - Association between a promoter polymorphism in the dopamine D2 receptor gene and schizophrenia. AB - Genetic factors and dopamine receptor dysfunction have been implicated in the pathophysiology of schizophrenia. Recently, an association between a putative functional promoter polymorphism (-141C Ins/Del) in the dopamine D2 receptor gene and schizophrenia was reported. We investigated unrelated Swedish schizophrenic patients (n = 129) and control subjects (n = 179) for the same polymorphism. Similarly to a previous Japanese report, the - 141C Del allele frequency was significantly lower in patients than controls (chi2=4.4, 1 df, p<0.05; odds ratio 0.49, 95% confidence interval 0.26-0.91). The present and previous results may indicate that the -141C Ins/Del dopamine D2 receptor gene polymorphism affects susceptibility to schizophrenia. PMID- 10541005 TI - Tyrosine transport is regulated differently in patients with schizophrenia. AB - Previous PET studies of tyrosine transport have suggested that the transport of tyrosine from blood to brain compartment is not dependent on its plasma concentration in patients with schizophrenia. In order to examine this relationship, the transport constant (K1) of tyrosine was determined in five patients with schizophrenia and five normals. L-[1-11C]Tyrosine was injected i.v. and arterial blood samples were taken during PET scanning. The tyrosine transport was assessed during baseline conditions and after oral administration of L tyrosine at a dose (175 mg/kg) that significantly elevated the plasma levels. K1 was determined from tracer kinetic modelling. The transport rate dropped in the normals after tyrosine loading, which is consistent with the prevailing notion that the brain transport system for neutral amino acids works close to saturation, whereas it was virtually unchanged in the schizophrenics. The results demonstrated that tyrosine transport was not saturated in the patients with schizophrenia and thus could lead to elevated brain concentrations of tyrosine. PMID- 10541006 TI - Cytogenetic findings in 250 schizophrenics: evidence confirming an excess of the X chromosome aneuploidies and pericentric inversion of chromosome 9. AB - Chromosomal abnormalities may be of help in identifying disease genes. To search for susceptibility loci for schizophrenia, we have performed chromosomal examinations by using the GTG banding technique for 250 schizophrenics. We found five cases with an aneuploidy of the X chromosome and ten cases with pericentric inversion of chromosome 9 [inv (9)]. These results confirmed an excess of the X chromosome aneuploidies in schizophrenia, indicating a possible involvement of the X chromosome in the pathogenesis of the illness. The observed incidence (4.0%) of inv (9) in our schizophrenic sample was significantly higher (p=0.013) than that reported in the general population in Japan (1.7%). Although inv (9) has been considered to be a normal variant, our observation implies a possible association between inv (9) and schizophrenia, suggesting that a susceptibility locus for the disease may be located at a breakpoint of the inversion on chromosome 9. PMID- 10541007 TI - Co-distribution of sensory gating and impaired niacin flush response in the parents of schizophrenics. AB - Complex illnesses may result from the interaction or addition of multiple factors. We examined the familial co-distribution of two abnormalities common in schizophrenia: impaired auditory sensory gating and impaired flush response to niacin. In ten families, the obligate carrier parent had sensory-gating deficits, while eight of the ten parents without a family history of schizophrenia had impaired flush response. No parents had both deficits. The data are consistent with a theory suggesting the interaction of these two factors in some cases of schizophrenia. PMID- 10541008 TI - Age at onset anticipation in familial schizophrenia. Does the phenomenon even exist? AB - The discovery of dynamic mutations, like the expansion of unstable CG-rich trinucleotide repeat sequence mutations, has revived the interest in investigating the phenomenon of anticipation of age at onset of the illness (AAO) in familial schizophrenia. In those studies of parent-offspring pairs analyzed for AAO anticipation published to date, however, several ascertainment biases were not adequately controlled for. The present study focuses mainly on the age at investigation (AAI) bias, neglected so far, by investigating 96 schizophrenic parent offspring pairs and 26 aunt/uncle-niece/nephew pairs. When not controlling for a potential AAI bias, AAO differences in the parent-offspring sample in favor for anticipation were found in the same magnitude as reported by other authors (12.5 years, p < 0.0001). However, when controlling for AAI, these positive anticipation findings were compensated for (1.32 years, p=0.129). Additional selection procedures such as the exclusion of late-onset schizophrenia, the analysis of pairs where both members were through the age of risk, or the selection of aunt/uncle-niece/nephew pairs could not circumvent the AAI effect. These results suggest that the AAI effect is an essential bias in investigating anticipation, leading to false-positive AAO anticipation results if not taken into account. PMID- 10541009 TI - A population-based register study of the association between schizophrenia and rheumatoid arthritis. AB - The authors investigated the association between schizophrenia and rheumatoid arthritis. The design is a population-based case-control and follow-up study. The cases were 20495 patients admitted for schizophrenia and registered in the Danish Psychiatric Case Register. A total of 204912 persons matched on age and gender and chosen from the general population served as controls. Admissions for rheumatoid arthritis and other non-autoimmune, musculoskeletal disorders were checked in the Danish National Patient Register. Odds ratios and relative risks were estimated by the Mantel-Haenszel estimator and Poisson regression. The same analyses were carried out for 10242 patients with bipolar affective disorder and 102420 controls for comparison. Individuals with schizophrenia had a reduced risk for being admitted with rheumatoid arthritis [odds ratio 0.44 (CI 0.24-0.81)] in the case-control study. A similar result was found in the follow-up study, but the incidence of the degenerative disorders in the musculoskeletal system was equally significantly lower in both studies. The incidence of rheumatoid arthritis among the bipolar patients was the same as in the control population. The negative association between schizophrenia and rheumatoid arthritis may thus be the result of ascertainment bias and selection due to under reporting and treatment of the medical illness. Clinicians are reminded of the difficulties in detecting medical illness among individuals with schizophrenia. PMID- 10541010 TI - Inhibitory processing in visuospatial attention in healthy adults and schizophrenic patients. AB - This study assessed visuospatial attention in healthy adults and medicated schizophrenic patients. Participants performed a visual orientation task in which a peripheral cue was followed. at different intervals, by a target presented either at valid or invalid locations. When the long stimulus onset asynchrony (SOA) was used, participants were presented with either a single peripheral cue (single-cue condition) or two cues, the peripheral cue followed by a central cue (the double-cue condition). Healthy adults showed marginal facilitation effects with the short SOA and similar inhibition of return effects with the long SOA in both single-cue and double-cue conditions. Schizophrenic individuals showed a big facilitation effect with the short SOA and normal inhibition of return with the long SOA in both cue conditions. Results with the short SOA replicated previous findings (Huey, E.D., Wexler, B.E., 1994. Schizophrenia Research 14, 57-63) but, in contrast, we did not observe blunted inhibition of return with the long SOA. An inspection of the differences in the procedures used in both studies may help both to account for the discrepancies and to reveal what processes involved in visuospatial attention are affected in schizophrenia. PMID- 10541011 TI - Brain morphology in adolescents at genetic risk for schizophrenia assessed by qualitative and quantitative magnetic resonance imaging. PMID- 10541012 TI - The administration of neuroleptic drugs decreased the arousal level, as reflected in a reduction of early CNV amplitude. PMID- 10541013 TI - A baculovirus anti-apoptosis gene homolog of the Trichoplusia ni granulovirus. AB - An inhibitor of apoptosis (iap) gene homolog (Tn-iap) of the Trichoplusia ni granulovirus (TnGV) was cloned, sequenced and mapped on the genome of TnGV. Tn iap encoded a protein (Tn-IAP) of 301 amino acids with a predicted molecular mass of 35 kDa. The Tn-IAP contained the two sequence motifs, BIRs and RING finger, characteristic of IAP proteins, and shared identities of 21-27% and similarities of 28-53% with IAP proteins of Cydia pomonella GV (Cp-IAP), Orgyia pseudotsugata multinucleocapsid nucleopolyhedrovirus (MNPV) (Op-IAP1, 3), Autographa californica MNPV (Ac-IAP1), Bombyx mori NPV (Bm-IAP1), Lymantria dispar MNPV (Ld IAP3) and Buzura suppressaria single nucleocapsid NPV (Bs-IAP1). However, Tn-IAP shared no significant homology with baculovirus IAP2 proteins. Using an antisense Tn-iap probe, two major transcripts of approximately 800 nt and 1600 nt were detected by Northern blot analysis of RNA extracted from the fat body of T. ni larvae infected with the TnGV. Unlike Cp-IAP and Op-IAP3, however, Tn-IAP did not rescue virion occlusion in SF21 cells infected with a p35-deficient AcMNPV mutant. Tn-IAP's synthesis in vivo but failure to rescue p35-deficient AcMNPV in SF21 cells suggests it is a functional IAP that is only effective in certain cell types. PMID- 10541014 TI - Sequence variations of Epstein-Barr virus LMP2A gene in gastric carcinoma in Japan. AB - The latent EBV gene products expressed in Epstein-Barr virus (EBV)-associated gastric carcinoma (EBVaGC), are only LMP2A, EBNA-1, BARF-0 and EBERs. To examine the correlation between LMP2A sequence variation in EBVaGC and transformation of the cells, the complete sequence of the LMP2A gene was determined in three cases of Japanese EBVaGC and compared with the prototype B95-8 strain. In addition, the sequences of exons 2,6 and 7 of LMP2A were determined in four to six EBVaGC cases. The results of sequence analysis indicated that LMP2A of EBVaGC was structurally very similar to B95-8, but contained a significant nucleotide variation. Ten nucleotide substitutions were identified in almost all cases tested, and three of these caused amino acid changes. Of these three, two amino acid substitutions were not expected to change any known functions of LMP2A. The other amino acid substitution from serine to threonine was located at codon 348 within one of the target epitopes of EBV-specific cytotoxic T-lymphocytes. The LMP2A of EBV in peripheral blood lymphocytes from six healthy individuals showed serine (4/6 cases) or threonine (2/6 cases) substitution at codon 348, while LMP2A with the threonine substitution was the major form (5/6 cases) observed in EBVaGC, indicating that EBV with the threonine substitution may confer an advantage for viral persistence in tumor cells. However, our sequencing results suggested that the LMP2A protein in EBVaGC is functionally similar to that of the B95-8 strain and is not unique to gastric carcinoma, indicating the importance of LMP2A for EBV latency. PMID- 10541015 TI - Sequence analysis of the NSP4 gene from human rotavirus strains isolated in the United States. AB - Two major and one minor genotype of the rotavirus NSP4 gene have been described. The sequences of 29 NSP4 genes from rotavirus isolates obtained in the United States during the 1996-1997 rotavirus season (types P[8]G1, P[8]G9, P[4]G2 and P[6]G9) and 10 strains isolated during previous rotavirus seasons (types P[8]G1 and P[4]G2) were determined. All NSP4 genes from strains with short E types (6 P[4]G2, 4 P[6]G9) belonged to genotype NSP4A, whereas all 19 strains with long E types (16 P[8]G1, 3 P[8]G9) had NSP4 genes of genotype NSP4B. Genetic variation within genotypes was low ( < or = 2.3% for both NSP4A and NSP4B), confirming that the NSP4 genes are highly conserved. Nonetheless, at least two distinct sub lineages could be detected within each genotype: strains isolated in the same year, regardless of geographic location, were more closely related or even identical at the deduced amino acid level; strains isolated in different years were more distinct. Thus, geographic distance did not affect genetic distance. Northern hybridization analysis with NSP4A and NSP4B total gene probes failed to detect any unusual combinations of the VP6 and NSP4 genes in 31 additional isolates from the 1996-1997 rotavirus season. PMID- 10541017 TI - Molecular characterization of an isolate of citrus tristeza virus that causes severe symptoms in sweet orange. AB - The complete sequence (19,249 nucleotides) of the genome of citrus tristeza virus (CTV) isolate SY568 was determined. The genome organization is identical to that of the previously determined CTV-T36 and CTV-VT isolates. Sequence comparisons revealed that CTV-SY568, a severe stem-pitting isolate from California, has more than 87% overall sequence identity with CTV-VT, a seedling yellows isolate from Israel. Although SY568 has an overall sequence identity of 81% with CTV-T36, a quick decline isolate from Florida, the sequence identity in the 3' half of the genome is over 90% while the sequence identity in the 5' half of the genome is as low as 56%. Based on the sequence alignments of these three isolates, sequences in the 3' half of the genome are generally well conserved, while the sequences in the 5' half are relatively divergent. Sequence data of independent overlapping clones from the CTV-SY568 genome revealed two regions with highly divergent sequences. In open reading frame 1b (RNA dependent RNA polymerase), there were 118 nucleotide differences that lead to 16 amino acid changes. In the open reading frame of the divergent coat protein gene, 5 amino acid changes result from 48 nucleotide differences. Most differences occurred in the third position of the codons, and resulted in silent amino acid substitutions. RNase protection assays demonstrated that most of the clones obtained are representative of the major RNA species of this isolate. Northern analysis indicated that CTV-SY568 accumulated more viral RNA including genomic and certain subgenomic RNAs than isolates VT or T36 in sweet orange. PMID- 10541016 TI - Active human hepatitis B viral polymerase expressed in rabbit reticulocyte lysate system. AB - Human HBV polymerase has been expressed in reticulocyte lysate system. The expressed protein shows the DNA-dependent DNA polymerase activity. In vitro transcription and translation produces a major protein product with an apparent molecular weight of approximately 100 kD. The HBV DNA polymerase has been characterized biochemically in the condition that the contaminating cellular DNA polymerases were fairly suppressed by aphidicolin and NEM. The polymerization reaction is optimal at pH 7.5 and 37 degrees C and the polymerase requires either MnCl2 or MgCl2, with a preference for MnCl2. The protein represented an optimal activity in the presence of either 75 mM NaCl or 100 mM KCl, with a higher activity at 75 mM NaCl than 100 mM KCl. Study of the polymerizing activity of the deleted versions of the polymerase protein suggests that the terminal protein is essential for full polymerase function and the spacer region may decrease the stability of the P protein. PMID- 10541018 TI - Infectious bronchitis virus S2 gene sequence variability may affect S1 subunit specific antibody binding. AB - The S2 gene of several strains of infectious bronchitis virus (IBV) belonging to the Arkansas, Connecticut, and Florida serotypes was sequenced. Phylogenetic analysis of the S2 gene nucleotide and deduced amino acid sequence data resulted in groups of strains that were the same as groupings observed when S1 sequence data was used. Thus, it appears that S2 subunits are conserved within a serotype but not between serotypes. Although the sequence differences were small, we found that only a few amino acid differences were responsible for different secondary structure predictions for the S2 subunit. It is likely that these changes create different interactions between the S1 and S2 subunits, which could affect the conformation of the S1 subunit where serotype specific epitopes are located. Based on this sequence data, we hypothesize that the S2 subunit can affect specific antibody binding to the S1 subunit of the IBV spike glycoprotein. PMID- 10541021 TI - Changes found in nursing home admission practices since advent of PPS. Prospective-pricing-system. PMID- 10541020 TI - Movement to temperature monitoring loses steam. PMID- 10541019 TI - Specific interaction of a 25-kilodalton cellular protein, a 40S ribosomal subunit protein, with the internal ribosome entry site of hepatitis C virus genome. AB - Translation initiation of hepatitis C virus (HCV) RNA is controlled by an internal ribosome entry site (IRES) contained in 5' noncoding region (NCR) and in several nucleotides of the coding region. The ability of a 25-kilodalton cellular protein (p25) to bind the HCV 5' NCR is correlated with the efficiency of translation initiation of HCV RNA, indicating that this protein plays a critical role in HCV translation (S. Fukushi, C. Kurihara, N. Ishiyama, F. B. Hoshino, A. Oya, and K. Katayama, J Virol 71, 1662-1666, 1997). We have extended the study for identification of the IRES region required for p25 binding. For this purpose, we have performed UV cross-linking competition analyses using 5'- or 3'- deleted mutants of the HCV 5' NCR as competitor RNAs for binding of p25 to wild-type HCV 5' NCR. Competitor RNAs lacking nucleotides (nt) 47-74 or nt 279-331 did not inhibit p25 binding to the HCV IRES, indicating that these regions are necessary for interaction of the p25 and HCV IRES. Since p25 binding was not observed in the IRES elements of encephalomyocarditis virus and poliovirus in UV cross linking competition analyses, the p25 binding may be specific for the HCV IRES. p25 bound to the HCV IRES was detected when a purified 40S ribosomal subunit was used for UV cross-linking experiment, indicating that p25 is one of 40S ribosomal subunit proteins. These results reveal an unique interaction between the 40S ribosomal subunit and HCV IRES to contribute to translation initiation of the HCV genome. PMID- 10541022 TI - Patients need more information about HMO formularies, says report on Medicare plans. PMID- 10541023 TI - Most U.S. overseas drug donations are useful; attention needed to donor requests and product age and relevance. PMID- 10541024 TI - North Carolina, Louisiana legislate collaborative drug therapy management. PMID- 10541026 TI - Coalition continues work on medication-use performance measures. PMID- 10541025 TI - North Carolina vaccine program linked to improved childhood immunization rates. PMID- 10541027 TI - Nonmalignant pain goes untreated in one fourth of nursing home residents, study shows. PMID- 10541028 TI - Getting to the bottom of a sentinel event. PMID- 10541029 TI - Perindopril erbumine. PMID- 10541030 TI - Phosphatidylserine. PMID- 10541031 TI - Urokinase activity after freezing: implications for thrombolysis in intraventricular hemorrhage. AB - The retention of urokinase activity after frozen storage was studied. Urokinase powder was reconstituted aseptically in sterile water for injection or preservative-free 0.9% sodium chloride injection to a final concentration of 5000 IU/mL. Samples were stored in 5-mL plastic syringes at -20 or -70 degrees C for up to six months. Samples containing urokinase 25,000 IU/mL were similarly prepared by using sodium chloride injection as the diluent and were stored frozen at the same temperatures for up to 93 days. Urokinase activity was measured with a chromogenic assay at each test interval. Samples were also cultured after thawing to evaluate their potential to support microbial growth. The activity of urokinase at either concentration did not change appreciably during the study period. The method of thawing-at room temperature or in a refrigerator-had no effect on urokinase activity. No microbial growth was observed. Urokinase 5000 IU/mL did not show any changes in activity when reconstituted with sterile water for injection or 0.9% sodium chloride injection and frozen for up to six months. Urokinase 25,000 IU/mL in sodium chloride injection was also stable after 93 days of frozen storage. PMID- 10541032 TI - Monitoring temperature-sensitive vaccines and immunologic drugs, including anthrax vaccine. AB - The experience of the U.S. Army Medical Materiel Center, Europe (USAMMCE), in monitoring temperature-sensitive vaccines and immunologic drugs, including anthrax vaccine, during storage and shipment is discussed. USAMMCE uses an electronic monitoring device to monitor and archive the time-temperature history of shipments of various vaccines, immunoglobulins, and other drugs requiring refrigeration. Using these monitors, USAMMCE can track its carriers' performance, reduce product loss, and validate quality. USAMMCE trains people to pack refrigerated items and to activate and place the monitoring device inside the packing container. Over 1200 temperature-monitor readings from 44 U.S. military logistical depots, hospitals, and clinics located outside the United States are evaluated annually by the USAMMCE pharmacist; each reading represents one shipment or packed box. When deactivated during unpacking, the device flashes green for a successful shipment (all temperature readings within the ideal range) or red for a potentially problematic shipment. From January through October 1998, the device was used in 750 temperature-sensitive shipments; 72% of the devices were returned to USAMMCE in green condition and the remainder in red. Of the red flashing monitors, 15% were determined to signal that the drugs were received in unacceptable condition. USAMMCE successfully shipped more than 26,000 vials of anthrax vaccine from February through October 1998 within the manufacturer's guidelines for storage temperature. Temperature monitoring is essential for proper storage and transport of vaccines and immunologic drugs. PMID- 10541033 TI - Bioactivity of cryopreserved alteplase solutions. PMID- 10541034 TI - Antiparkinsonian drug prescribing in elderly inpatients receiving risperidone therapy. PMID- 10541035 TI - Rectal administration of warfarin. PMID- 10541037 TI - Current literature. PMID- 10541036 TI - Plasticizers in perspective. PMID- 10541038 TI - Molecular mechanisms of pigment transport in melanophores. AB - We present an overview of the research on intracellular transport in pigment cells, with emphasis on the most recent discoveries. Pigment cells of lower vertebrates have been traditionally used as a model for studies of intracellular transport mechanisms, because these cells transport pigment organelles to the center or to the periphery of the cell in a highly co-ordinated fashion. It is now well established that both aggregation and dispersion of pigment in melanophores require two elements of the cytoskeleton: microtubules and actin filaments. Melanosomes are moved along these cytoskeletal tracks by motor proteins. Recent studies have identified the motors responsible for pigment dispersion and aggregation in melanophores. We propose a model for the possible roles of the two cytoskeletal transport systems and how they might interact. We also discuss the putative mechanisms of regulation of pigment transport, especially phosphorylation. Last, we suggest areas of research that will receive attention in the future in order to elucidate the mechanisms of organelle transport. PMID- 10541039 TI - Identification of microtubule-organizing centers in interphase melanophores of Xenopus laevis larvae in vivo. AB - The morphological characteristics of microtubule-organizing centers (MTOCs) in dermal interphase melanophores of Xenopus laevis larvae in vivo at 51-53 stages of development has been studied using immunostained semi-thick sections by fluorescent microscopy combined with computer image analysis. Computer image analysis of melanophores with aggregated and dispersed pigment granules, stained with the antibodies against the centrosome-specific component (CTR210) and tubulin, has revealed the presence of one main focus of microtubule convergence in the cell body, which coincides with the localization of the centrosome specific antigen. An electron microscopy of those melanophores has shown that aggregation or dispersion of melanosomes is accompanied by changes in the morphological arrangement of the MTOC/centrosome. The centrosome in melanophores with dispersed pigment exhibits a conventional organization, and their melanosomes are situated in an immediate vicinity of the centrioles. In melanophores with aggregated pigment, MTOC is characterized by a three-zonal organization: the centrosome with centrioles, the centrosphere, and an outlying radial arrangement of microtubules and their associated inclusions. The centrosome in interphase melanophores is presumed to contain a pair of centrioles or numerous centrioles. Because of an inability of detecting additional MTOCs, it has been considered that an active MTOC in interphase melanophores of X. laevis is the centrosome. We assume that remaining intact microtubules in the cytoplasmic processes of mitotic melanophores (Rubina et al., 1999) derive either from the aster or the centrosome active at the interphase. PMID- 10541040 TI - Ultimate fate of rod outer segments in the retinal pigment epithelium. AB - To investigate the degradation pathway of rod outer segments (ROS) in vivo, we injected gold-labeled ROS into the subretinal space of rabbits using a pars plana approach. Histology and electron microscopy performed on the specimens 72 hr after ROS injection revealed that the retina over the injection site was reattached, the retinal pigment epithelial (RPE) cells were intact, and gold granules were localized inside melanin granules and melanosomes. These results indicate that, in RPE, in vivo degradation of ROS is associated with melanosomes. PMID- 10541041 TI - Ephelides are more related to pigmentary constitutional host factors than solar lentigines. AB - Ephelides and solar lentigines are benign pigmented spots, which are currently associated with an increased risk of skin cancer. These two pigmented spots are known to be discriminated by their clinical, histological, and electron microscopic characteristics, even though occasional misclassification can occur because of their similarity. It has also been questioned whether these spots are not one and the same. In this study, we have attempted to differentiate between these two pigmented spots with the use of a standardized protocol for clinical examinations on 272 healthy volunteers, paying particular consideration to their pigmentary and constitutional host factors. We found that solar lentigines 1) are more prevalent than ephelides, 2) increase in prevalence and number with higher age, and 3) are most prevalent on the trunk and occur more frequently in males than in females. A trend is also observed whereby ephelides 1) loose their prevalence with age, 2) become equally distributed on the face, arms, and trunk, and 3) occur more frequently in females. An intimate association of ephelides, but not solar lentigines, has been found with hair color and skin type. All of these findings are in agreement with most of those reported in the literature, supporting the view that ephelides and solar lentigines are different types of pigmented lesions. PMID- 10541042 TI - Hypophysial and meningeal melanocytes in the Zucker rat. AB - Melanocytes have not been described in the pituitary of mammals or in the meninges of the rat. In this paper, we report the presence of the cluster of melanocytes in the intermediate lobe of the pituitary and around the median eminence of the hypothalamus forming an 'infundibulo-hypophysial circle', and also describe the characteristics of meningeal melanocytes in Zucker rats. In the leptomeninges, numerous melanocytes were found on the ventrolateral surface of cerebral hemisphere in the area of the middle cerebral artery. Pigment granules were also observed in the surrounding tissue outside the melanocytes as well as incorporated in the cytoplasm of neural and epithelial cells. Electron microscopy revealed that melanosomes in hypophysial and meningeal melanocytes were in different (II-IV) stages of maturity. In the leptomeninges of Zucker rats, HMB-45 immunoreactivity was found in round non-melanosome-containing cells, while no HMB 45 reaction was found in the leptomeninges of the albino rat. We conclude that both obese and lean Zucker rats possess functionally active melanocytes in the meninges and the pituitary and transfer pigment granules to neighboring cells. The distributions of melanocytes in proximity to blood vessels in the leptomeninges and in the 'infundibulo-hypophysial circle' suggest an endocrine secretory function. PMID- 10541043 TI - Location and catalytic characteristics of a multipotent bacterial polyphenol oxidase. AB - The melanogenic marine bacterium Marinomonas mediterranea contains a multipotent polyphenol oxidase (PPO) able to oxidize substrates characteristic for tyrosinase and laccase. Thus, this enzyme shows tyrosine hydroxylase activity and it catalyzes the oxidation of a wide variety of o-diphenol as well as o-methoxy activated phenols. The study of its sensitivity to different inhibitors also revealed intermediate features between laccase and tyrosinase. It is similar to tyrosinases in its sensitivity to tropolone, but it resembles laccases in its resistance to cinnamic acid and phenylthiourea, and in its sensitivity to fluoride anion. This enzyme is mostly membrane-bound and can be solubilized either by non-ionic detergent or lipase treatments of the membrane. The expression of this enzymatic activity is growth-phase regulated, reaching a maximum in the stationary phase of bacterial growth, but L-tyrosine, Cu(II) ions, or 2,5-xylidine do not induce it. This enzyme can be separated from a second PPO form by gel permeation chromatography. The second PPO is located in the soluble fraction and shows a sodium dodecyl sulfate (SDS)-activated action on the characteristic substrates for tyrosinase, L-tyrosine, and L-dopa, but it does not show activity towards laccase-specific substrates. The involvement of the multipotent PPO in melanogenesis and its relationship with the SDS-activated form and with the alternative functions proposed for multicopper oxidases in other microorganisms are discussed. PMID- 10541044 TI - Role of CXCR4 in HIV-1-induced apoptosis of cells with a CD4+, CXCR4+ phenotype. PMID- 10541045 TI - Apoptosis and HIV infection: T-cells fiddle while the immune system burns. AB - Enhancement of programmed cell death (apoptosis) of CD4 T-cells by human immunodeficiency virus (HIV) is thought to be an important factor in the pathogenesis of HIV disease. Recent studies have cast doubt on this concept, however, portraying apoptosis as a potent antiviral strategy to eliminate infected cells. These studies have shed new light on the role of apoptosis in HIV infection. While cellular and immunologic mechanisms of apoptosis purge the HIV infected lymphoid cell population, HIV thwarts apoptosis in myeloid cells, particularly monocyte/macrophages. Although HIV protease inhibitor therapy partially reverses the lymphoid cell process, this therapeutic approach fails to counter the persistence of HIV infection in myeloid cells. Thus apoptosis of T cells may be a futile host attempt to control the spread of HIV while the infection smoulders in monocyte/macrophages. In other words, the antiviral defense system fiddles while the immune system burns. PMID- 10541046 TI - Monocyte chemoattractant protein-1 and macrophage inflammatory protein-2 production is inhibited by taurine chloramine in rat C6 glioma cells. AB - Taurine monochloramine (Tau-Cl) is formed through the actions of a halide dependent myeloperoxidase system associated with polymorphonuclear leukocytes (PMN). Tau-Cl inhibits production of inflammatory mediators by activated macrophages, and PMN. Recently, Tau-Cl was shown to inhibit production of nitric oxide and prostaglandin E2 by activated C6 glioma cells. Since chemokines, secreted by activated glial cells, play a prominent role in eliciting inflammatory responses in the central nervous system, the effects of Tau-Cl on production of monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-2 (MIP-2) were determined in activated C6 glioma cells. Tau Cl inhibited production of MCP-1 and MIP-2 in a concentration-dependent manner, and was most potent against MCP-1. Tau-Cl exerted a transient inhibition of the temporal expression of MCP-1 and MIP-2 mRNAs during the first 4 h of activation. Although both chemokine mRNA levels were similar to those of control cells after 8-24 h of activation, production of the chemokine proteins, especially MCP-1, remained markedly low. These results suggest that Tau-Cl inhibits production of MCP-1 and MIP-2 in activated C6 cells primarily through post-transcriptional mechanisms. PMID- 10541047 TI - Immunological parameters and respiratory functions in patients suffering from atopic bronchial asthma after intravenous treatment with salmon calcitonin. AB - This study reports the effect of salmon calcitonin on airway function and peripheral blood parameters in asthmatic subjects. The premise for the study is that calcitonin is given to asthmatics that require systemic corticosteroids as a way to counter problems with calcium balance and osteoporosis, and that it has an immunosuppressive effect. Salmon calcitonin (100 IU) was administered to 18 patients with atopic bronchial asthma, and the following spirometric parameters were evaluated: forced vital capacity (FVC), forced expiratory volume in the first second (FEV1), peak respiratory flow rate (PEFR) and forced expiratory flow rates at 25%, 50% and 75% of the forced vital capacity (FEF25%, FEF50% and FEF75%). Calcitonin significantly decreased the levels of FVC and FEV1 by 20 min after starting the infusion. The effect of 500 mg aminophylline, used as a reference drug in this study, was much more profound, with a significant increase in all investigated parameters. Also, the effect of salmon calcitonin on some immune parameters (white blood cell count, number of eosinophils, serum levels of immunoglobulins IgG, IgM and IgA, and serum levels of lymphocytes subpopulations CD3, CD4, CD8 and CD19) was determined in another group of 30 patients suffering from atopic bronchial asthma. Calcitonin at a dose of 100 IU/day subcutaneously for 3 days did not alter the immune parameters studied, thus rendering it safe for such and similar treatment schedules in a variety of medical conditions. PMID- 10541048 TI - Immunogenicity of H-2Kb-low affinity, high affinity, and covalently-bound peptides in anti-tumor vaccination. AB - CTL induction by immunization with synthetic peptide epitopes has been shown to inhibit tumor growth and its metastatic spread. Ex vivo pulsing of peptides on MHC class I-bearing cells such as RMA-S cells or professional APCs elicits an effective CTL response. Since the stability of the MHC-peptide complex is strongly correlated with the overall immunogenecity, we compared the effect of immunization with low affinity, high affinity, and irreversibly bound MHC peptides in the context of immunotherapy of metastasis. MUT1, a tumor-associated antigen peptide that was isolated from 3LL Lewis lung carcinoma, is a low H-2Kb binder. MUT1 was modified into a high binder by changing positions 3, 5, and 8 to the favorable anchor residues. In addition, we introduced a photo-active chemical moiety, which can bind irreversibly to MHC upon illumination. These peptides, loaded onto RMA-S, were used to immunize mice against the 3LL tumor. Vaccination via the covalent conjugation of the low binder peptide was found to increase the CTL response measured against MUT1 loaded cells and against H-2Kb transfected D122 cells relative to the native MUT1 peptide. However, the photo cross-linking of the high affinity peptide to the MHC did not significantly improve the induction of specific CTL. The level of CTL activity was elevated to the same extent by either cross-linking the peptide to the MHC or by modifying it into a high-binder peptide. The protective capacity of all the peptide-based vaccines against D122 metastatic spread to the lungs was found to be comparable. These results indicate that augmentation of the affinity of a TAA peptide to the RMA-S surface MHC molecules, by conversion to a high-affinity mimotope or by photo conjugation, can significantly enhance the immune response. There seems to be, however, a ceiling beyond which increase in the peptide-binding affinity does not lead to a corresponding enhancement of the overall immunogenicity of the peptide. PMID- 10541049 TI - The influence of dietary protein antigen on Th1/Th2 balance and cellular immunity. AB - Dietary administration of ovalbumin (OVA) antigen (Ag) into OVA-specific T cell receptor alphabeta-transgenic (TCR-Tg) mice resulted in the induction of activated CD4+ Th cells expressing CD69 early activation Ag. However, the number of CD4+ Th cells rather decreased by dietary administration of OVA antigen. The production of Th1-cytokines such as IFN-gamma and IL-2 markedly reduced in spleen of OVA-fed mice compared to mice fed with normal diet. In sharp contrast, the production of Th2-cytokine, IL-4 greatly increased in spleen of OVA-fed mice though the number of CD4+ T cells decreased to less than 10% of control mouse spleen. The decrease of IFN-gamma production and the increase of IL-4 production by CD4+ T cells was demonstrated at a single cell level by intracellular cytokine staining analysis. Moreover, such a polarized cytokine production pattern was also demonstrated using highly purified CD4+ T cells obtained from mice fed with OVA. In addition to the decrease of Th1-cytokine production, TCR-Tg mice fed with OVA-containing diet showed greatly reduced in vivo generation of NK cells, LAK cells and CTL. These results suggested that dietary protein antigen caused the polarization of Th1/Th2 balance into Th2-dominant immunity and inhibited cellular immunity. PMID- 10541050 TI - In vivo acquired mechanisms of tumor cells local defense against the host innate immunity effectors: implication in specific antitumor immunity. AB - As shown earlier, the cells transformed in vitro by several different oncogenes, or spontaneously, during in vivo growth in normal hosts would be gradually replaced by the highly-tumorigenic descendants co-expressing high H2O2 catabolizing and PGE2-releasing activities. Acquisition of (H2O2(CA) + PGE(S)) phenotype provides the cells with local defense mechanisms against the host innate immunity effectors. However, it remained unknown, whether the expression of (H2O2(CA) + PGE(S)) phenotype is implicated in susceptibility of tumor cells expressing tumor-specific transplantation antigens to rejection in immune animals. Here, with the use of SV40 in vitro transformed parental cells, negative in expression (H2O2(CA) + PGE(S)) phenotype, and their in vivo selected descendant tumor cell lines expressing this phenotype, we show that: (1) the rates of in vivo selection of the parental SV40 tumor cells expressing (H2O2(CA) + PGE(S)) phenotype are the same in normal and SV40-immune animals; (2) in vivo selected SV40 tumor cells expressing (H2O2(CA) + PGE(S)) phenotype, although they retain specific immunosensitivity, are 100 times less effectively rejected in SV40-immunized animals, as compared with their in vitro SV40-transformed parental cells. Thus, in vivo acquired immunologically non-specific local mechanisms of tumor cells defense against the host innate immunity effectors, significantly decreases the effectiveness of their specific immunorejection. PMID- 10541051 TI - Mechanisms of protein A superantigen-induced signal transduction for proliferation of mouse B cell. AB - Apart from many of the biological properties of protein A (PA) of Staphylococcus aureus, it has been recognized recently as a B-cell superantigen. Therefore, we investigated the molecular mechanisms of PA superantigen-induced mice splenic B cell proliferation. Treatment of resting B cells with PA-evoked cell proliferation. Binding of PA to B cells led to a cascade of signal transduction mechanisms involving tyrosine kinase that activated phospholipase C, which in turn activated protein kinase C (PKC), and translocated it from cytosol to membrane. Mitogen-activated protein (MAP) kinase has been found to be activated down-stream of PKC in this signal pathway, which ultimately caused an activation of serum-responsive factor (SRF). Inhibition at any step of this signaling cascade could block B-cell proliferation. PA could also stimulate the Bcl-2 gene expression at protein level thereby supporting the pro-proliferative effect of PA. Thus, the molecular mechanisms related to PA-induced B cell proliferation has been delineated in this report as tyrosine kinase > PLC > PKC > MAP kinase > SRF > Bcl-2. Knowledge gathered from these observations might be of immense help to study the immune cell proliferation as a part of immunoactivation process. Also, the development of suitable inhibitors of the signaling pathway outlined here might provide clues as to how to abrogate pathologic antibody production in many disease processes. PMID- 10541052 TI - Occurrence of adrenergic nerve fibers and of noradrenaline in thymus gland of juvenile and aged rats. AB - Adrenergic nerve fibers (ANF) were studied in juvenile, adult and old rats by biochemical and morphological methods and by quantitative analysis of images (QAI). After chemical sympathectomy with neurotoxin 6-OH dopamine, the greater part of ANF disappeared. The whole thymus was drawn in juvenile normal or sympathectomized rats, in adult normal or sympathectomized rats and in old normal or sympathectomized rats. Thymuses from the above-mentioned animals were weighed, measured and dissected. Thymic slices were stained with eosin orange for the detection of the microanatomical details and with Bodian's method for the recognition of the whole nerve fibers. Histofluorescence microscopy was used for staining of ANF while immunofluorescence microscopy was employed for staining of neuropeptide Y (NPY)-like immunoreactivity. Biochemical dosage of proteins and of noradrenaline amount was performed. Finally, all morphological results were subjected to QAI. Our results suggest that: (1) total innervation of the thymus increases with age; (2) ANF do not change with age; (3) the content of noradrenaline in the thymus increases with age; and (4) NPY-like immunoreactive structures in the thymus decrease with age. Biochemical results are in accordance with the morphological ones and both are confirmed by means of QAI. The probable function of sympathetic innervation of rat thymus is also discussed. PMID- 10541054 TI - CD11a expression and soluble ICAM-1 levels in peripheral blood in high-risk and overt type 1 diabetes subjects. AB - A pair of correspondent adhesion molecules: LFA-1 (CD11aCD18) and ICAM-1 (CD54) was shown to be involved in autoimmune insulitis in animal models. Anti-LFA-1 or anti-ICAM-1 monoclonal antibodies administered in vivo had a very strong preventive effect on the development of spontaneous diabetes with a marked reduction of insulitis. On the other hand elevated levels of the soluble form of ICAM-1 (sICAM-1) were documented in subjects at risk for type 1 diabetes. Recently sICAM-1 was shown to play an immunoregulatory role as an inhibitor of islet insulitis. The aim of the present study was to evaluate CD11a + mononuclear cells (lymphocytes and monocytes) and soluble sICAM-1 levels in the peripheral blood of subjects with preclinical and overt type 1 diabetes to assess their role in the development of the autoimmune process and their possible associations with the humoral autoimmune markers. The study was carried out in three groups of subjects: 26 first degree relatives of type 1 diabetes patients (prediabetics) with the combinations of autoantibodies against pancreatic B cells (ICA, GADA, IA 2A, IAA), 22 patients with a recent onset of type 1 diabetes and age and sex matched 24 healthy volunteers (control group). We observed an increased fluorescence intensity of CD11a on mononuclear cells in overt diabetes subjects and a positive correlation between CD11a fluorescence intensity on monocytes and ICA titre. The highest sICAM-1 levels we obtained in the peripheral blood in the prediabetics in comparison to patients with clinical diabetes and the healthy controls. We found a positive correlation between slCAM-1 and values of ICA, GADA and a total number of antibodies present. In conclusion our study suggests that LFA-1 and sCAM-1 play an important role in the pathogenesis of type 1 diabetes. The assessment of the CD11a bearing monocytes and sICAM-1 levels are potential markers of the preclinical stage of the autoimmune diabetes, but further prospective studies in high risk diabetes type 1 subjects are needed. PMID- 10541053 TI - Macrophages that kill glioma cells expressing the membrane form of macrophage colony stimulating factor are resistant to prostaglandin E2 and interleukin-10. AB - Malignant rat T9 glioma cells retrovirally transduced with the membrane form of macrophage colony stimulating factor (mM-CSF) were killed by bone marrow derived macrophages in 24 h cytotoxicity assays. Prostaglandin E2 (PGE) and interleukin 10 (IL10) were tested for their ability to block this tumoricidal reaction. Only at very high nonphysiological concentrations of PGE (10(-5) and 10(-6) M) was this cytotoxicity inhibited. Use of high doses of theophylline, a phosphodiesterase inhibitor, also prevented macrophages from killing the mM-CSF transduced target cells. IL10 did not alter the killing potential of the mM-CSF tumoricidal macrophages, even though IL10 reduced the production of nitric oxide by macrophages in response to tumor necrosis factor and lipopolysaccharide. IL10 enhanced the growth of bone marrow macrophages suggesting that IL10 has a complex role in the regulation of tumoricidal macrophages. Thus, the mM-CSF may be an ideal agent to treat tumors that utilize either of these two immunosuppressive defense mechanisms that may block other forms of treatment. PMID- 10541055 TI - Central inhibition of sexual response in the male: a theoretical perspective. AB - A theoretical model for central inhibition of sexual response is proposed, postulating individual variability in the propensity for such inhibition. Whereas such inhibition is typically adaptive, individuals with high propensity may be vulnerable to sexual dysfunction, and those with low propensity to high risk sexual behavior. Evidence of the existence and localization of such inhibitory mechanisms from both the animal and human literature is reviewed. Evidence of central neurotransmitters with sexual inhibitory effects is substantial, though in most cases the inhibition is not specific to sexual response or behavior. Recent studies have identified centers in the brain stem and lateral hypothalamus which appear to have specific inhibitory effects on sexual response. A variety of adaptive mechanisms involving inhibition of sexual response are considered, some involving perception of threat, others occurring more directly as consequences of previous sexual activity. These different adaptive functions may well involve different inhibitory mechanisms. This theoretical model opens a new agenda for experimental research into adaptive sexual behavior, both human and animal. PMID- 10541056 TI - How do cell assemblies encode information in the brain? AB - The present review discusses why cell-assembly coding, i.e. ensemble coding by functionally connected neurons, is a tenable view of the brain's neuronal code and how it operates in the working brain. The cell-assembly coding has two major properties, i.e., partial overlapping of neurons among assemblies and connection dynamics within and among the assemblies. The former is the ability of one neuron to participate in different types of information processing. The latter is the capability for functional synaptic connections, detected by activity correlations of the neurons, to change among different types of information processing. An example of a series of experiments which detected these two major properties is then given. Several relevant points concerning the detection of the actual dynamics of cell-assembly coding are also enumerated. They include the dependence of the type of cell-assembly coding on types of information-processing in different structures of the brain, sparse coding by distributed overlapped assemblies, and coincidence detection as a role of individual neurons to bind distributed neurons into cell assemblies. PMID- 10541057 TI - A behavioral stages model of classical (Pavlovian) conditioning: application to cognitive aging. AB - In the present article, it is argued that a five-stage sequential model of the behavioral and neurophysiological events that occur when organisms are exposed to signals predicting significant events suggests that classical conditioning produces multiple memory traces involving both excitatory and inhibitory processes. Further, these multiple brain structures and associated neurophysiological mechanisms are beginning to be understood; thus, using Pavlovian conditioning techniques to study aging and cognitive functions may provide insights into which brain structures or mechanisms are responsible for more general age-related declines in associative learning and memory. The evidence for this model is briefly reviewed and studies suggesting age-related effects on classical conditioning of various response systems are described within the context of the brain structures implicated by the model. PMID- 10541058 TI - Role of local anesthetics on both cholinergic and serotonergic ionotropic receptors. AB - A great body of experimental evidence indicates that the main target for the pharmacological action of local anesthetics (LAs) is the voltage-gated Na+ channel. However, the epidural and spinal anesthesia as well as the behavioral effects of LAs cannot be explained exclusively by its inhibitory effect on the voltage-gated Na+ channel. Thus, the involvement of other ion channel receptors has been suggested. Particularly, two members of the neurotransmitter-gated ion channel receptor superfamily, the nicotinic acetylcholine receptor (AChR) and the 5-hydroxytryptamine receptor (5-HT3R type). In this regard, the aim of this review is to explain and delineate the mechanism by which LAs inhibit both ionotropic receptors from peripheral and central nervous systems. Local anesthetics inhibit the ion channel activity of both muscle- and neuronal-type AChRs in a noncompetitive fashion. Additionally, LAs inhibit the 5-HT3R by competing with the serotonergic agonist binding sites. The noncompetitive inhibitory action of LAs on the AChR is ascribed to two possible blocking mechanisms. An open-channel-blocking mechanism where the drug binds to the open channel and/or an allosteric mechanism where LAs bind to closed channels. The open-channel-blocking mechanism is in accord with the existence of high-affinity LA binding sites located in the ion channel. The allosteric mechanism seems to be physiologically more relevant than the open-channel-blocking mechanism. The inhibitory property of LAs is also elicited by binding to several low-affinity sites positioned at the lipid-AChR interface. However, there is no clearcut evidence indicating whether these sites are located at either the annular or the nonannular lipid domain. Both tertiary (protonated) and quaternary LAs gain the interior of the channel through the hydrophilic pathway formed by the extracellular ion channel's mouth with the concomitant ion flux blockade. Nevertheless, an alternative mode of action is proposed for both deprotonated tertiary and permanently-uncharged LAs: they may pass from the lipid membrane core to the lumen of the ion channel through a hydrophobic pathway. Perhaps this hydrophobic pathway is structurally related to the nonannular lipid domain. Regarding the LA binding site location on the 5-HT3R, at least two amino acids have been involved. Glutamic acid at position 106 which is located in a residue sequence homologous to loop A from the principal component of the binding site for cholinergic agonists and competitive antagonists, and Trp67 which is positioned in a stretch of amino acids homologous to loop F from the complementary component of the cholinergic ligand binding site. PMID- 10541059 TI - Is there sexual dimorphism in obsessive-compulsive disorder? AB - The present review addresses the question of sexual dimorphism in obsessive compulsive disorder. It enumerates examples that could be interpreted to suggest the existence of such dimorphism from the fields of epidemiology, phenomenology, pharmacology, neuropsychology, neuroimaging and genetics. We conclude that data, at this point, are too scarce to warrant a firm conclusion. On the contrary it seems that there are enough indications in the literature that hint at the possibility of sexual dimorphism to stimulate further research in the field. PMID- 10541060 TI - The conditioned avoidance response test re-evaluated: is it a sensitive test for the detection of potentially atypical antipsychotics? AB - The present review discusses the history and paradigm of the conditioned avoidance response (CAR) in rats for the detection of potential antipsychotic activity of drugs. In addition, the role of dopamine (DA) D2, serotonin (5 HT)2A/2C, alpha1, 5-HT1A, DA D4, muscarinic and glutamate receptors in the suppression of CAR induced by various classes of drugs is evaluated. Finally, data investigating brain sites of action for the mediation of CAR behavior is discussed. It is concluded that the CAR test, originally found to be sensitive for the detection of antipsychotic drugs with high affinity as antagonists for brain dopamine receptors, is also sensitive for the detection of potentially antipsychotic compounds acting primarily via neurotransmitter receptors other than the DA D2 receptor. Furthermore, the review confirms the importance of the nucleus accumbens(shell) in the mediation of effects on CAR produced by traditional, as well as atypical antipsychotic drugs. PMID- 10541061 TI - Neurochemical mechanisms of the defensive behavior in the dorsal midbrain. AB - Some regions in the mesencephalon, such as dorsal periaqueductal gray, inferior colliculus and deep layers of superior colliculus have been grouped together as a continuous strip of midbrain structures involved in the integration of the different components of aversive states in the brain. In fact, escape behavior and defensive, or fear-like behavior often result when these sites are electrically or chemically stimulated. Moreover, the behavioral responses induced by stimulation of these structures are, in general, accompanied by increases in mean arterial blood pressure, heart rate and respiration, and by analgesia. Both the behavioral and autonomic consequences of electrical stimulation of the mesencephalic tectum was shown to be attenuated by minor tranquilizers, probably through enhancement of GABAergic neurotransmission. Besides GABAergic interneurons which exert a tonic inhibitory control on neural circuits responsible for the behavioral correlates of the aversion in the above-mentioned structures, several other mechanisms such as opioid, neuropeptides, serotonergic and excitatory amino acids have also been implicated in the regulation of these processes. As to the analgesia that accompanies these aversive states it is mediated by non-opioid mechanisms, particularly by serotonergic ones through 5 HT2 receptors. Now, efforts have been made to characterize the mode of action of these neurotransmitters on their multiple receptors and how they interact with each other to produce or regulate the neural substrates of aversion in the midbrain. PMID- 10541062 TI - Theoretical review: altered pain regulatory systems in chronic pain. AB - This review synthesizes the existing literature regarding the relationship between resting blood pressure and pain sensitivity, and the literature indicating possible endogenous opioid dysfunction in chronic pain. Adaptive interactions between the cardiovascular and pain regulatory systems occur in healthy individuals, with greater blood pressure associated with decreased acute pain sensitivity. Endogenous opioids appear necessary for full expression of this relationship. There is ample evidence indicating diminished endogenous opioid CSF/plasma levels in chronic pain patients, yet little is known about the functional effects of these opioid changes. A theoretical model is proposed based upon the literature reviewed suggesting progressive dysfunction in endogenous opioid systems with increasing chronic pain duration. This dysfunction is hypothesized to result in dysregulation of normally adaptive relationships between the cardiovascular and pain regulatory systems, resulting in increased chronic pain intensity and increased acute pain sensitivity among chronic pain patients. Preliminary data are consistent with the hypothesis of progressive opioid changes resulting in dysfunctional alterations in the adaptive blood pressure-pain relationship. Clinical implications of this theory are discussed. PMID- 10541063 TI - NACOB presentation Keynote lecture. Cancellous bone biomechanics. North American Congress on Biomechanics. AB - Cancellous bone is both a biological and a mechanical structure. The interaction between these two aspects of cancellous bone is sufficiently strong that understanding the mechanical properties of the tissue is not possible without consideration of the biology. This manuscript is a mathematical expansion of a portion of the first author's Keynote lecture at the 1998 NACOB presentation. The cellular activity of cancellous bone proceeds in part by the transport of metabolites between trabecular hard tissue and marrow. The anatomical observation is that human trabeculae are seldom internally served by a blood supply, suggesting that the transport mechanisms for trabecular survival are diffusion and a collection of mechanisms for active transport of metabolites independent of blood flow. It will be demonstrated that metabolite transport by diffusion can explain two notable empirical relationships for bone: (a) the close relationship between the bone surface and the bone volume, and (b) the exponential decline in the bone volume fraction during periods of mechanical disuse. A mathematical model is also developed showing how mechanical loading can effect bone volume fraction by increasing metabolite transport between the tissue compartments. PMID- 10541064 TI - NACOB presentation CSB New Investigator Award. Balance recovery from medio lateral perturbations of the upper body during standing. North American Congress on Biomechanics. AB - Postural control strategies have in the past been predominantly characterized by kinematics, surface forces, and EMG responses (e.g. Horak and Nashner, 1986, Journal of Neurophysiology 55(6), 1369-1381). The goal of this study was to provide unique and novel insights into the underlying motor mechanisms used in postural control by determining the joint moments during balance recovery from medio-lateral (M/L) perturbations. Ten adult males received medio-lateral (M/L) pushes to the trunk or pelvis. The inverted pendulum model of balance control (Winter et al., 1998, Journal of Neurophysiology 80, 1211-1221) was validated even though the body did not behave as a single pendulum, indicating that the centre of pressure (COP) is the variable used to control the centre of mass (COM). The perturbation magnitude was random, and the central nervous system (CNS) responded with an estimate of the largest anticipated perturbation. The observed joint moments served to move the COP in the appropriate direction and to control the lateral collapse of the trunk. The individual joints involved in controlling the COP contributed differing amounts to the total recovery response: the hip and spinal moments provided the majority of the recovery (approximately 85%), while the ankles contributed a small, but significant amount (15%). The differing contributions are based on the anatomical constraints and the functional requirements of the balance task. The onset of the joint moment was synchronous with the joint angle change, and occurred too early (56-116 ms) to be result of active muscle contraction. Therefore, the first line of defense was provided by muscle stiffness, not reflex-activated muscle activity. PMID- 10541065 TI - NACOB presentation to ASB Young Scientist Award: Postdoctoral. The impact of boundary conditions and mesh size on the accuracy of cancellous bone tissue modulus determination using large-scale finite-element modeling. North American Congress on Biomechanics. AB - The apparent properties of cancellous bone are determined by a combination of both hard tissue properties and microstructural organization. A method is desired to extract the underlying hard tissue properties from simple mechanical tests, free from the complications of microstructure. It has been suggested that microCT voxel-based large-scale finite element models could be employed to accomplish this goal (van Rietbergen et al., 1995, Journal of Biomechanics, 28, 69-81). This approach has recently been implemented and it is becoming increasingly popular as finite element models increase in size and sophistication (Fyhrie et al., 1997, Proceedings of the 43rd Annual Meeting of the Orthopaedic Research Society, San Francisco, CA, p. 815; van Rietbergen et al., 1997, Proceedings of the 43rd Annual Meeting of the Orthopaedic Research Society, San Francisco, CA, p. 62). However, no direct quantitative measurements of the accuracy of this method applied to porous structures such as cancellous bone have been made. This project demonstrates the feasibility of this approach by quantifying its best-case accuracy in determining the trabecular hard tissue modulus of analogues fabricated of a material with known material properties determined independently by direct testing. In addition we were able to assess the impact of mesh size and boundary conditions on accuracy. We found that the assumption of a frictionless boundary condition in the parallel plate compression loading configuration was a significant source of error that could be overcome with the use of rigid end-caps similar to those used by Keaveny et al. (1997 Journal of Orthopaedic Research, 15(1), 101-110). In conclusion, we found that this approach is an effective method for determining the average trabecular hard tissue properties of human cancellous bone with an expected practical accuracy level better than 5%. PMID- 10541066 TI - Partial prevention of monocyte and granulocyte activation in experimental vein grafts by using a biomechanical engineering approach. AB - Leukocytes interact with endothelial cells and contribute to the development of vascular diseases such as thrombosis and atherosclerosis. These processes are possibly influenced by mechanical factors. This study focused on the role of mechanical stretch in the activation of monocytes and granulocytes in experimental vein grafts. Two models were created by using rats: a nonengineered vein graft with increased tensile stress, which was created by grafting a jugular vein into the abdominal aorta, and an engineered vein graft with reduced tensile stress, which was created by restricting the vein graft into a cylindrical sheath constructed by using fixative-treated intestinal tissue. The density of activated monocytes and granulocytes, which attached to the endothelium, and the distribution of the intercellular adhesion molecule (ICAM)-1 in endothelial cells were examined using immunohistological assays. It was found that, in nonengineered vein grafts, the density of activated monocytes and granulocytes increased significantly compared to that in normal jugular veins at day 1, 5, 10 and 20. At each observation time, the cell density in the proximal region of the nonengineered vein grafts was significantly higher than that in the middle and distal regions, and the cell density in the distal region was significantly higher than that in the middle region. These changes were associated with ICAM-1 clustering at day 1 and 5 and focal ICAM-1 un-regulation at day 10 and 20. In engineered vein grafts, the density of activated monocytes and granulocytes decreased significantly compared to that in nonengineered vein grafts at all observation times, although it was significantly higher than that in normal jugular veins. At each observation time, the cell density in the proximal and distal regions was significantly higher than that in the middle region, but no significant difference was found between the proximal and distal regions. ICAM-1 clustering along endothelial cell borders was found at day 1 and 5, but no apparent focal ICAM-1 up-regulation was found at day 10 and 20. These results suggested that mechanical stretch due to exposure to increased tensile stress contributed to the activation of monocytes and granulocytes in experimental vein grafts, and this event could be partially prevented by reducing tensile stress using a biomechanical engineering approach. PMID- 10541067 TI - Streaming potential of human lumbar anulus fibrosus is anisotropic and affected by disc degeneration. AB - The streaming potential responses of non-degenerate and degenerate human anulus fibrosus were measured in a one-dimensional permeation configuration under static and dynamic loading conditions. The goal of this study was to investigate the influence of the changes in tissue structure and composition on the electrokinetic behavior of intervertebral disc tissues. It was found that the static streaming potential of the anulus fibrosus depended on the degenerative grade of the discs (p = 0.0001) and on the specimen orientation in which the fluid flows (p = 0.0001). For a statically applied pressure of 0.07 MPa, the ratio of streaming potential to applied pressure ranged from 5.3 to 6.9 mV/MPa and was largest for Grade I tissue with axial orientation and lowest for Grade III tissue with circumferential orientation. The dynamic streaming potential responses of anulus fibrosus were sensitive to the degeneration of the disc: the total harmonic distortion factor increased by 108%, from 3.92 +/- 0.66% (mean +/- SD) for Grade I specimens to 8.15 +/- 3.05% for Grades II and III specimens. The alteration of streaming potential reflects the changes in tissue composition and structure with degeneration. To our knowledge, this is the first reported data for the streaming potential of human intervertebral disc tissues. Knowledge of the streaming potential response of the intervertebral disc provides an understanding of potentially important signal transduction mechanisms in the disc and of the etiology of intervertebral disc degeneration. PMID- 10541068 TI - The influence of friction and interference on the seating of a hemispherical press-fit cup: a finite element investigation. AB - The formation of gaps in the polar region of acetabular cups is seen as a drawback of press-fit fixation of non-cemented acetabular cups. Recent findings indicate a link between long-term polar gaps and the gaps present directly after implantation. In this study the process of press-fitting is simulated with a linear-elastic two-dimensional axisymmetric finite-element model. The aim of this paper is to investigate the possible importance of friction and interference on the formation of these gaps. A range of cup-bone friction coefficients (mu = 0.1 0.5) is assigned to the cup-bone interface in order to represent the unknown amount of friction occurring during press-fitting. The cup is modeled with a radius of 27 mm, whereas the radius of the cavity is varied between 26.50 and 26.75 mm, thus, creating 0.50 and 0.25 mm radial interference fits. The difference in cavity radius represents the discrepancy between the radius of the last-reamer-used and radius of the cavity it creates. The subchondral plate is considered as being completely removed during reaming. The effects of impact blows via the surgeon's mallet during surgery are modeled as a series of four load pulses, in which peak force is gradually increased from 0.5 to 4.0 kN. The effects of load removal as well as those of load application are investigated. On load application, the cup penetrates into the cavity, and on load removal, the cup rebounds. Depending on the friction, interference and load applied, the position of the cup after the load pulse is somewhere between its position at peak force and its position at the beginning of the pulse. Although the simplifications and conditions involved in the creation of the model necessitate caution when interpreting the results for all clinical cases, it is found that the seating of hemispherical cups in trabecular bone could be more satisfactory for intermediate values of friction (mu = 0.2-0.3) and smaller interference fits (0.25 mm). PMID- 10541069 TI - Measuring muscle and joint geometry parameters of a shoulder for modeling purposes. AB - An extensive set of muscle and joint geometry parameters was measured of the right shoulder of an embalmed male. For all muscles the optimal muscle fiber length was determined by laser diffraction measurements of sarcomere length. In addition, tendon length and physiological cross-sectional area were determined. The parameter set was needed to enhance the reliability of a computer model of the shoulder (Van der Helm, 1994a,b Journal of Biomechanics 27, 527-550, 551 569). With the model, an abduction of the arm was simulated in seven positions, at 30 degrees intervals. In each of the simulated arm positions, actual sarcomere lengths were calculated from the lengths of 104 muscle elements, distributed over 16 shoulder muscles. For most muscle elements, the simulated abduction appeared to take place within the sarcomere length range in which the muscle elements can exert force. The muscle elements can then act on the ascending limb as well as on the plateau and on the descending limb of the relative force-length curves of sarcomeres. The produced data set is not only important for the refinement of shoulder modeling, but also for functional analyses of shoulder movements in general. PMID- 10541070 TI - Compression data on bovine bone confirms that a "stressed volume" principle explains the variability of fatigue strength results. AB - The literature contains many measurements of the fatigue properties of compact bone, but these experimental results have been difficult to interpret and use due to a large amount of apparent scatter: variation in the number of cycles to failure for a given cyclic stress or strain range. Recently Taylor (1998a, Journal of Orthopaedic Research, 16, 163-169) showed that much of this scatter could be explained using a statistical model which took into account specimen size, or more specifically stressed volume. The present paper describes an attempt to test this model by using it to predict some new data, for bovine bone tested in compressive loading at room temperature at physiological loading rates. Twenty specimens were tested at the same applied load range (100 MPa). The theory was able to predict the mean behaviour of the specimens very well, with an accuracy (expressed in terms of stress) of 2%. It was also able to predict the degree of scatter (i.e. the variation of Nf), which was shown to be similar to that measured by other workers. PMID- 10541071 TI - A telescopic inverted-pendulum model of the musculo-skeletal system and its use for the analysis of the sit-to-stand motor task. AB - For field applicability of biomechanical methodologies aiming at assessing motor ability in disabled, or at risk of disablement (e.g. elderly), subjects, measurements must be carried out using a least perceivable to the subject and essential experimental apparatus. Since data thus obtained do not necessarily lend themselves to straightforward interpretation, they should be fed to a model of the portion of the musculo-skeletal system involved that already embodies the invariant aspects of both the modelled system and the motor task. Through such a minimum measured-input model, richer, physiology-related, and thus easier to interpret, information may be expected. In this framework, the present study investigated the sit-to-stand motor task using information obtained only from a force plate located under seat and subject's feet, a seat uniaxial load-cell and basic anthropometric parameters. Data were collected in a sample of 12 able bodied subjects while executing the motor task at different speeds. The musculo skeletal system was modelled as a telescopic inverted pendulum (TIP) that could vary its length (shortening or elongation) by effect of a force actuator and its orientation in space by effect of two couple actuators that were looked upon as muscle equivalent effectors. The TIP model output consisted in the kinematics and dynamics of these actuators. It allowed the identification of four functional phases in which the seat-to-stand motor task could be divided, and a detailed description of the relevant mechanics in terms of balance control and centre of mass elevation. Motor strategy modifications associated with speed variation could also be identified. For a global evaluation of the motor act it showed to be no less informative than more demanding multi-segment models. Although it is true that specific musculo-articular functions can only be inferred, the more compact information yielded by the TIP model is expected to facilitate subject and/or disability classification. PMID- 10541072 TI - Total shoulder and relative muscle strength in the scapular plane. AB - Strength profiles of the shoulder joint are measured experimentally for two arm positions in "the scapular plane" in order to present quantitative data on the shoulder strength. Apart from yielding the actual force a subject can exert in various directions, these measurements also exhibit e.g. the strongest and weakest directions, in fact the relative strength in all directions. The inter individual variation of the direction of maximal force was at most 14 degrees (sd). The experimental profiles are compared with the corresponding theoretical profiles, obtained by using a shoulder model. The calculations were made both with default muscle parameters and individually adapted parameters. The results show that the employed shoulder model, which is based on data from an elderly population, may be adapted to other populations and that the necessary changes in relative muscle strength are those expected on biomechanical grounds. Without model changes the difference between measured (in the mean) and predicted maximal force directions was at most 50 degrees. Muscle parameter adjustment reduced this difference to 23 degrees. The strength profiles clearly indicate in what direction a person can produce larger forces and which muscles that contribute. PMID- 10541073 TI - A new method for estimating the axis of rotation and the center of rotation. AB - A new method is presented for estimating the parameters of two different models of a joint. The two models are: (1) A rotational joint with a fixed axis of rotation, also referred to as a hinge joint and (2) a ball and socket model, corresponding to a spherical joint. Given the motion of a set of markers, it is shown how the parameters can be estimated, utilizing the whole data set. The parameters are estimated from motion data by minimizing two objective functions. The method does not assume a rigid body motion, but only that each marker rotates around the same fixed axis of rotation or center of rotation. Simulation results indicate that in situations where the rigid body assumption is valid and when measurement noise is present, the proposed method is inferior to methods that utilize the rigid body assumption. However, when there are large skin movement artefacts, simulation results show the proposed method to be more robust. PMID- 10541074 TI - Mathematical modelling of stress in the hip during gait. AB - A mathematical model is developed for calculating the contact stress distribution in the hip for a known resultant hip force and characteristic geometrical parameters. Using a relatively simple single nonlinear algebraic equation, the model can be readily applied in clinical practice to estimate the stress distribution in the most frequent body positions of everyday activities. This is demonstrated by analyzing the data on the resultant hip force obtained from laboratory observations where a stance period of gait is considered. PMID- 10541075 TI - Parameters influencing the accuracy of the point of force application determined with piezoelectric force plates. AB - Errors up to +/- 30 mm in determining the point of force application with piezoelectric force plates have been reported in the literature (Kistler, 1984. Multicomponent Measuring Force Plate for Biomechanics and Industry. Kistler, Switzerland; Bobbert and Schamhardt, 1990. Journal of Biomechanics 23, 705-710; Sommer et al., 1997. Proceedings of the XVI th I.S.B. Congress). To explain the main factors influencing the systematic errors the force plate system is modeled as a two-dimensional beam structure. By this model it is strongly indicated that the cause for the errors in determining the point of force application are bending moments in the measurement posts. The main parameters influencing the shape and magnitude of the error function are the ratios between the bending stiffness of the plate and the bending and compressive stiffnesses of the measurement posts. In the current design it is therefore not possible to eliminate the cause for the errors by changing the constructive parameters. By comparing the error functions derived with the beam model to the correction formulas given in the literature an improved algorithm is proposed. This paper shall help biomechanists in understanding the basic concepts of determining the point of force application with force plates and in constructing custom-made force plates for specific applications. PMID- 10541077 TI - Mechanical strength of the cement-bone interface is greater in shear than in tension. AB - The objective of this study was to determine the relative mechanical properties of the cement-bone interface due to tensile or shear loading. Mechanical tests were performed on cement-bone specimens in tensile (n = 51) or shear (n = 55) test jigs under the displacement control at 1 mm/min until complete failure. Before testing, the quantity of bone interdigitated with the cement was determined and served as a covariate in the study. The apparent strength of the cement-bone interface was significantly higher (p < 0.0001) for the interface when loaded in shear (2.25 MPa) when compared to tensile loading (1.35 MPa). Significantly higher energies to failure (p < 0.0001) and displacement before failure (p < 0.01) were also determined for the shear specimens. The post-yield softening response was not different for the two test directions. The data obtained herein suggests that cement-bone interfaces with equal amounts of tensile and shear stress would be more likely to fail under tensile loading. PMID- 10541076 TI - Development of a fluorescent light technique for evaluating microdamage in bone subjected to fatigue loading. AB - A new method using fluorescent light microscopy has been developed to visualize and evaluate bone microdamage. We report the findings of two different experiments with a common aim of comparing the fluorescent light technique to the brightfield method for quantifying microdamage in bone. In Experiment 1, 36 canine femurs were tested in four-point cyclic bending until they had lost between 5 and 43% of their stiffness. The loaded portion of the bone was stained en bloc with basic fuchsin for the presence of damage. Standard point counting techniques were used to calculate fractional damaged area (Dm.Ar = Cr.Ar/B.Ar, mm2/mm2) under brightfield and fluorescent microscopy. In Experiment 2, bone microdamage adjacent to endosseous implants, subjected to fatigue loading (150,000 cycles, 2 Hz and 37 degrees C) ex vivo was examined. The bone around the implant was either allowed to heal (adapted specimen) for 12 weeks after placement in dog mid-femoral diaphyses prior to testing or was loaded immediately to simulate non-healed bone surrounding endosseous implants (non-adapted). Crack numerical density (Cr.Dn = Cr.N/B.Ar, #/mm2), crack surface density (Cr.S.Dn = Tt.Cr.Le/B.Ar, mm/mm2) and fractional damaged area were calculated separately by both techniques in the adapted and non-adapted specimens. In both Experiments 1 and 2, significantly more microdamage was detected by the fluorescent technique than by the brightfield method. Also, there was a trend towards higher intraobserver repeatability when using the fluorescent method. These results suggest that the brightfield technique underestimates microdamage accumulation and that the fluorescent technique better represents the actual amounts of microdamage present. The results demonstrate that the fluorescent method provides an accurate and precise approach for bone microdamage evaluation, and that it improves the prediction of stiffness loss from damage accumulation. PMID- 10541078 TI - Linear elastic and poroelastic models of cartilage can produce comparable stress results: a comment on Tanck et al. (J Biomech 32:153-161, 1999) PMID- 10541079 TI - PACS: new age radiology. Picture archiving and communication systems. PMID- 10541080 TI - Doctor and the doll. PMID- 10541081 TI - The missed breast cancer: perceptions and realities. PMID- 10541082 TI - Impact of filmless radiology on frequency of clinician consultations with radiologists. AB - OBJECTIVE: The purpose of the study was to determine the impact of filmless operation on the relative frequency of in-person consultations in the radiology department between radiologists and clinicians. CONCLUSION: The transition to filmless operation at the Baltimore Veterans Affairs Medical Center was associated with an 82% reduction in the in-person consultation rate for general radiography and a 44% reduction for cross-sectional imaging despite an increase in the volume of studies. The major reason for this decrease was the convenient access to current and prior images provided by the PACS (picture archiving and communication system). Radiology departments contemplating a transition to filmless operation should prepare for communication with clinicians to shift from being mostly in person to being conducted more and more through electronic forms of communication. PMID- 10541083 TI - PACS and radiology practice: enjoy the benefits but acknowledge the threats. Picture archiving and communications systems. PMID- 10541084 TI - PACS in sonography: accuracy of interpretation using film compared with monitor display. Picture archiving and communication systems. AB - OBJECTIVE: The goal of this study was to determine the relative accuracy of interpretation of sonography when viewed on a monitor or on film. MATERIALS AND METHODS: Four radiologists twice interpreted a series of 440 sonograms using the following sequences of display formats for initial and second interpretations: film-film, film-monitor, monitor-film, and monitor-monitor. Reporting discrepancies between the initial and subsequent interpretation were reviewed by an arbitration panel unaware of the display mode. Results were analyzed for differences in error rate attributable to film versus monitor display format, chronology of interpretation, individual observer, and observer seniority. RESULTS: We found no statistically significant difference in the error rate for film (10.3%) versus monitor display format (14.6%) (p = .09). Likewise, we found no significant differences in the error rates attributable to chronology of interpretation (p = .13), individual observer (p = .54), or observer seniority (p = .87). CONCLUSION: Interpretative accuracy is similar whether sonograms are interpreted on a monitor or on film. PMID- 10541085 TI - Radiographic anatomy: multimedia interactive instructional software on CD-ROM. AB - OBJECTIVE: We wanted to create a filmless radiographic anatomy curriculum, a didactic software with a digital image database on CD-ROM for first-year medical students. CONCLUSION: We created a CD-ROM that includes an introduction, radiographic anatomy tutorial, and interactive questions. Additional features include Boolean text searching, links to related images, and Internet accessibility. The software can be updated. PMID- 10541086 TI - Spectrum of imaging findings of the liver in end-stage cirrhosis: Part II, focal abnormalities. AB - Cirrhosis, through the process of necrosis, fibrosis, regeneration, and malignant transformation, creates multiple focal benign and malignant hepatic masses. The detection and appearance of these masses varies with the severity of the cirrhotic process and the imaging technique used. Although some overlap exists in the imaging appearance of the benign and malignant masses, in most instances recognition of a few characteristic features will yield the correct diagnosis. PMID- 10541087 TI - Cirrhotic liver enhancement on dual-phase helical CT: comparison with noncirrhotic livers in 146 patients. AB - OBJECTIVE: The purpose of this study was to prospectively determine any differences in vascular and liver enhancement between patients with cirrhosis and patients without cirrhosis during both the arterial and portal venous phases on dual-phase helical CT. SUBJECTS AND METHODS: Fifty-eight patients with histologically proven cirrhosis (group 1) and 88 without cirrhosis (group 2 = normal findings on CT, group 3 = metastases, group 4 = other liver diseases) underwent dual-phase helical CT of the liver. Attenuation values of liver and vessels were measured on unenhanced scans and on scans obtained during the arterial and portal venous phases. The mean enhancement values per time interval (5 sec) were determined. Results were analyzed taking into account various intrinsic patient parameters. RESULTS: We found no statistically significant difference in terms of mean vascular enhancement and mean liver enhancement during the arterial imaging phase for each time interval among all the groups. The mean peak enhancement and mean liver enhancement during the portal venous phase were significantly lower in group 1 than in other groups. Time to peak enhancement was significantly delayed in group 1. CONCLUSION: In spite of the hepatic arterial buffer response, mean liver enhancement during the arterial phase was not significantly different in patients with cirrhosis compared with patients without cirrhosis. Although portal vein enhancement did not differ significantly, enhancement of cirrhotic liver was significantly lower during the portal venous phase and delayed, presumably because of decreased peripheral portal perfusion. The contrast injection protocol may be tailored to optimize conspicuity of hypovascular tumor. PMID- 10541088 TI - Percutaneous liver biopsy: a cost-benefit analysis comparing sonographic and CT guidance. AB - OBJECTIVE: We compared the relative cost of a liver biopsy performed with sonographic guidance with that of one performed with CT guidance in a cost benefit analysis model. MATERIALS AND METHODS: Variables were estimated from a search of the literature and from clinical experience with 437 hepatic biopsies at our institution. Probability variables included the probability of obtaining an adequate sample and the probability of a major complication. Cost variables included the direct and indirect costs, the cost of a major complication, and the opportunity costs of foregone revenue from preempted diagnostic studies. One-way and two-way sensitivity analyses were performed. RESULTS: Using baseline values, CT guidance was 1.89 times more expensive than sonographic guidance. Sensitivity analyses indicate that CT and sonographic guidance costs would be equivalent if the success rate with sonographic guidance was 39.8%, the opportunity costs of CT guidance were 3.13 times less than best estimates, and the opportunity costs of sonography were 3.15 times greater than best estimates. CONCLUSION: Sonographic guidance for hepatic biopsies is substantially more economical than CT guidance across a wide range of estimated costs. PMID- 10541089 TI - Comparison of biopsy devices: sonographically guided percutaneous liver biopsies in rabbits. PMID- 10541090 TI - Detection of hepatocellular carcinoma: comparison of low- and high-spatial resolution dynamic MR images. AB - OBJECTIVE: The purpose of our study was to compare the diagnostic performance of low- and high-spatial-resolution gadolinium chelate-enhanced triphasic dynamic gradient-recalled echo (GRE) MR images in the detection of hepatocellular carcinoma. MATERIALS AND METHODS: Triphasic dynamic MR images obtained with low (256 x 128) and high (512 x 224) image matrices in 28 patients with 65 hepatocellular carcinomas (HCCs) were retrospectively analyzed. Image review was conducted on a segment-by-segment basis; a total of 215 liver segments, including 56 segments with tumor burden, were reviewed for the presence of HCC by three independent radiologists. Detectability was evaluated with relative sensitivity, specificity, and receiver operating characteristic (ROC) analysis. Image quality was evaluated with rank order analysis. RESULTS: Relative sensitivity was statistically significantly better with high-spatial-resolution images than with low-spatial-resolution images (p < .005). Relative specificity was statistically significantly better with low-spatial-resolution images than with high-spatial resolution images (p < .001). Diagnostic accuracy determined by ROC curve analysis was marginally higher with high-spatial-resolution (area under ROC curve [Az] = .97) than with low-spatial-resolution (Az = .94, p < .09) images. Image quality was statistically significantly better with high-spatial-resolution images (p < .005). CONCLUSION: High-spatial-resolution dynamic GRE images were superior to low-spatial-resolution images in sensitivity of detecting HCC and in image quality. Triphasic dynamic GRE imaging in the screening and follow-up programs of patients with suspected HCC should be performed using high image matrices. PMID- 10541091 TI - Vascular changes in hepatocellular carcinoma: correlation of radiologic and pathologic findings. AB - OBJECTIVE: Our objective was to analyze the hemodynamic properties and vascular supply changes in the carcinogenesis of hepatocellular carcinoma. MATERIALS AND METHODS: Ten nodules (nine patients) (one early, three early-advanced, and six advanced cases of hepatocellular carcinoma) less than 3 cm in diameter were selected from 45 patients (50 nodules) who underwent CT arteriography and CT during arterial portography. These images were correlated with histopathologic findings. Ratios of all microscopically counted (normal hepatic and abnormal) arteries, normal hepatic arteries, and portal veins in each nodule to those in the surrounding liver were calculated. RESULTS: Early hepatocellular carcinoma (one early case and early areas in three early-advanced cases) had low attenuation on CT arteriography and isoattenuation on CT during arterial portography. Advanced hepatocellular carcinoma (six advanced cases and advanced areas in three early-advanced cases) had high attenuation on CT arteriography and low attenuation on CT during arterial portography. In early hepatocellular carcinoma, the ratios of all arteries, normal hepatic arteries, and portal veins were 1.21 +/- 0.07, 0.60 +/- 0.07, and 0.73 +/- 0.06, respectively. In advanced hepatocellular carcinoma, the ratios were 2.66 +/- 0.26, 0.08 +/- 0.04, and 0.07 +/- 0.03, respectively. CONCLUSION: In early hepatocellular carcinoma, the combination of normal hepatic artery degeneration and preserved portal veins results in low attenuation on CT arteriography and isoattenuation on CT during arterial portography. In advanced hepatocellular carcinoma, the combination of neoplastic (abnormal) arterial development by angiogenesis and obliteration of portal veins results in high attenuation on CT arteriography and low attenuation on CT during arterial portography. These findings are a characteristic difference between early and advanced hepatocellular carcinoma. PMID- 10541092 TI - CT fluoroscopy-assisted needle puncture and ethanol injection for hepatocellular carcinoma: a preliminary study. AB - OBJECTIVE: We assessed the usefulness of real-time CT fluoroscopy for needle guidance and evaluated the clinical usefulness of a unified CT fluoroscopy and angiography system in the treatment of hepatocellular carcinoma. SUBJECTS AND METHODS: A single-session percutaneous ethanol injection was performed with CT fluoroscopy guidance and monitoring for 15 hepatocellular carcinomas with an average size of 2.5 cm (range, 0.7-4.7 cm) in 10 consecutive patients. Of these, seven lesions were not seen on sonography. To mark the lesion for puncture, we performed CT arteriography or arterial injection of iodized oil. A puncture guide was applied to 12 lesions. RESULTS: The average depth from the skin's surface to the lesion was 9.3 cm (range, 4.5-11.5 cm), and the puncture route was transthoracic in five lesions and transabdominal in 13. The overall success rate in puncturing the lesions was 94.4% (17/18 sessions). The average number of punctures was 3.3, and it significantly decreased after introduction of a puncture guide compared with freehand puncture (p < .01). The average amount of injected ethanol was 12.7 ml (range, 4-27 ml). The ratio of injected ethanol dose to calculated ethanol dose was 0.6. Local recurrence occurred in four (26.7%) of 15 lesions after an average of 5 months. CONCLUSION: Using CT fluoroscopy for guidance of the needle and for monitoring ethanol infusion in the target lesion, we have found single-session percutaneous ethanol injection to be possible for hepatocellular carcinomas smaller than 5 cm or not revealed by sonography. The puncture guidance equipment was helpful for accurate insertion of the needle into the lesion, allowing a minimum number of punctures and minimal radiation exposure. PMID- 10541093 TI - Small hepatocellular carcinoma treated with percutaneous RF ablation: MR imaging follow-up. AB - OBJECTIVE: The purpose of our study was to determine the usefulness of MR imaging in the follow-up evaluation of small hepatocellular carcinoma lesions treated with RF ablation. SUBJECTS AND METHODS: The study group included 37 patients with a single hepatocellular carcinoma lesion less than 3 cm in diameter. A strict protocol required follow-up MR imaging every 6 months after RF treatment. At each follow-up visit, the findings on unenhanced and dynamic gadolinium-enhanced MR images were correlated with those on contrast-enhanced CT images and with results of fine-needle aspiration biopsy. In five patients who underwent surgical resection after the 6-month follow-up examination, comparison with histologic findings of surgical specimens was also performed. RESULTS: Correct diagnosis of complete or partial tumor necrosis was made in 32 (86%) of the 37 patients with the use of unenhanced and dynamic gadolinium-enhanced MR images. Hypointensity on T2-weighted images and loss of enhancement on dynamic MR images corresponded to completely necrotic lesions in all patients. Conversely, intratumoral regions of hyperintensity on T2-weighted images and enhancement on dynamic MR images did not always correlate to residual viable tumor. MR imaging and CT findings agreed in the evaluation of therapeutic response in all patients. CONCLUSION: Our experience confirms that MR imaging is useful for evaluating the effectiveness of RF therapy in achieving tumor regression. PMID- 10541094 TI - Percutaneous microwave coagulation therapy for hepatocellular carcinoma located on the surface of the liver. AB - OBJECTIVE: Percutaneous microwave coagulation therapy was recently introduced as a new treatment for hepatocellular carcinoma in our country. We performed this study to evaluate the efficacy and safety of this therapy for treatment of hepatocellular carcinoma, especially for tumors located on the surface of the liver. CONCLUSION: Percutaneous microwave coagulation therapy can be performed safely even in patients with cirrhosis and can achieve complete remission of small hepatocellular carcinomas (< or = 2.0 cm) located on the surface of the liver. PMID- 10541095 TI - Hepatic changes in benign obstruction of the hepatic inferior vena cava: CT findings. AB - OBJECTIVE: The objective of this study was to describe CT findings of changes in the liver associated with benign obstruction of the hepatic inferior vena cava (IVC). MATERIALS AND METHODS: For a 10-year period, 35 patients with benign obstruction of the hepatic IVC underwent contrast-enhanced CT of the abdomen. These patients were included in this retrospective study. CT scans were analyzed for morphologic changes and abnormal enhancement of the liver, changes in intrahepatic vessels, and additional findings that might be related to obstruction of the IVC. RESULTS: Morphologic changes of the liver included hypertrophy of the caudate lobe (91%) and the left lobe (57%), atrophy of the right lobe (49%), and a nodular surface (74%). The most common pattern of attenuation change was areas of linear, irregular, or wedge-shaped hypoattenuation predominantly located in the peripheral portion of the liver (63%). Diffuse hypoattenuation was seen in six patients (19%) and was frequently found in areas in which hepatic veins filled with hypoattenuated thrombosis (67%). On CT, segmental IVC obstruction (80%) was seen as an obliterated segment of the hepatic IVC. However, membranous IVC obstruction (20%) was not seen on CT. The IVC below the level of obstruction was often revealed as rounded (89%) and occasionally contained thrombus (37%) or calcification (26%). CONCLUSION: CT shows a broad spectrum of morphologic and attenuation changes of the liver and of the hepatic vessels in benign obstruction of the hepatic IVC. PMID- 10541096 TI - Small peripheral cholangiocarcinoma with undisturbed transiting portal vein: radiologic-pathologic correlation. PMID- 10541097 TI - Blood flow in healthy gallbladder walls on color and power Doppler sonography: effect of wall thickness and gallbladder volume. AB - OBJECTIVE: The purpose of this study was to determine the effect of gallbladder contraction on the conspicuity of flow in the normal gallbladder wall. SUBJECTS AND METHODS: Ten healthy adult volunteers without clinical evidence of gallbladder disease participated in the study. After patients fasted overnight, the gallbladder was scanned using gray-scale, color Doppler, and power Doppler sonography. The subjects were then given a standard meal consisting of 478 ml of a carbohydrate-rich dietary supplement, and the imaging sequence was repeated 20 and 45 min thereafter. Mural flow was graded using a four-step grading scheme. Gallbladder volume, wall thickness, and visibility of mural flow at all three time points were statistically compared. RESULTS: Enhanced mural flow was seen after meal consumption in all but one volunteer. Overall, mural flow was significantly greater 45 min after eating than at baseline or 20 min on color Doppler sonography (p = .004) and power Doppler sonography (p = .008). CONCLUSION: Flow in the gallbladder wall is a normal finding that is seen more easily when the gallbladder is contracted. This fact must be kept in mind when sonography is used with patients in whom acute cholecystitis is suspected, particularly if they do not fast before sonography. PMID- 10541098 TI - Anomalies and anatomic variants of the biliary tree revealed by MR cholangiopancreatography. PMID- 10541099 TI - Intramural varices of the bile duct: an unusual pattern of cavernous transformation of the portal vein. PMID- 10541100 TI - A specific sign of pneumoperitoneum on sonography: enhancement of the peritoneal stripe. AB - OBJECTIVE: Failure to reveal pneumoperitoneum is a recognized weakness of abdominal sonography. Our objective is to describe a reliable and reproducible sign of pneumoperitoneum that was first identified in an animal model and then confirmed in patients who had undergone laparoscopy. SUBJECTS AND METHODS: We injected 300 ml of degassed water into the peritoneal cavity of a 15-kg anesthetized pig. Sonographic images were obtained of the anterior peritoneal area after intraperitoneal injection of a single bubble, a series of bubbles, and, subsequently, a 10-ml bolus of air. Later, abdominal sonography was performed in nine patients who had undergone laparoscopy. Close attention was paid to the anterior peritoneal area and signs of free air observed in the animal model. Ten healthy volunteers functioned as a control group. RESULTS: In the pig, minute amounts of intraperitoneal air showed on sonography as enhancement of the peritoneal stripe. Larger volumes of intraperitoneal air showed as enhancement of the peritoneal stripe associated with dirty shadowing or distal multiple reflection artifacts. The stripe enhanced each time it appeared in the reflection artifact. Intraluminal gas was associated with a normal thin peritoneal stripe, superficial and distinct from the underlying gas artifact. The patients who had undergone laparoscopy showed findings suggestive of small and large pockets of free air, as we saw in the pig model. The control group showed findings consistent with intraluminal gas only. CONCLUSION: On sonography, enhancement of the peritoneal stripe alone or with reflection artifacts involving the peritoneal stripe is an accurate sign of pneumoperitoneum. PMID- 10541101 TI - Focal fatty infiltration of the pancreas: MR characterization with chemical shift imaging. PMID- 10541102 TI - Upper gastrointestinal series and CT findings of primary gastric plasmacytoma: report of two cases. PMID- 10541103 TI - CT cystography versus conventional cystography in evaluation of bladder injury. AB - OBJECTIVE: The objective of this study was to evaluate prospectively the use of CT cystography, using retrograde filling of the bladder with diluted iodinated contrast material, versus conventional cystography to identify bladder injury in patients with hematuria after blunt abdominal trauma. SUBJECTS AND METHODS: Inclusion criteria consisted of the adult hemodynamically stable abdominal trauma patient with hematuria referred for abdominopelvic CT and also being considered for cystography. An initial abdominopelvic CT scan using IV iodinated contrast material was obtained, as would have been done routinely in the trauma victim. A second CT scan through the pelvis was obtained after retrograde distention of the bladder with dilute iodinated contrast material. CT cystography revealing bladder injury was followed with appropriate therapy. CT cystograms not revealing injury were followed by conventional cystography. Results of patient outcome were evaluated. RESULTS: Over a 21-month period from January 1995 through September 1996, CT cystography was performed on 55 patients who presented with hematuria after blunt abdominal trauma. Five of the 55 patients had bladder injury on CT cystography. The injury in each of these five patients was confirmed intraoperatively. In the remaining 50 patients, both CT and conventional cystography did not reveal bladder injury. CONCLUSION: CT cystography is an accurate method for evaluating bladder injury in the blunt abdominal trauma victim with hematuria. CT cystography, performed in conjunction with routine CT of the abdomen and pelvis for evaluating traumatic hematuria, would therefore preclude conventional cystograms in these patients. PMID- 10541104 TI - Triphasic helical CT of the kidneys: contribution of vascular phase scanning in patients before urologic surgery. AB - OBJECTIVE: The purpose of this study was to evaluate the potential benefits of performing vascular phase scanning of the kidneys in addition to unenhanced and parenchymal phase contrast-enhanced CT in patients being examined for urologic surgery. MATERIALS AND METHODS: Parenchymal and vascular phase images from triphasic renal helical CT of 50 patients were sequentially evaluated in a randomized, retrospective fashion by two independent observers. The number of renal arteries and veins and the presence of vein or collecting system anomalies were recorded for each scan phase along with a subjective 10-point-scale score of the visibility of the vasculature and collecting system. Correlation of these findings was made with surgical or angiographic findings in 67 of the 87 kidneys and was made by consensus review in the remaining 20 kidneys. RESULTS: Accessory renal arteries were seen significantly more often (p < .05, chi-square test) on the vascular phase scans. The subjective scores for the visibility of the renal arteries and renal veins were significantly higher on the vascular phase scans (p < .0001, Wilcoxon's rank sum test). The subjective scores for the visibility of the filling of the collecting system and renal pelvis were significantly higher for the parenchymal phase scans, despite the use of a small contrast bolus before each scan (p < .0001, Wilcoxon's rank sum test). CONCLUSION; Triphasic renal CT better reveals the artery and vein anatomy of the kidney than does parenchymal phase imaging only. Triphasic helical CT is indicated in patients undergoing planning for urologic surgery when vascular anatomy is clinically important. PMID- 10541105 TI - Evaluation of dynamic gadolinium-enhanced breath-hold MR angiography in the diagnosis of renal artery stenosis. AB - OBJECTIVE: The aim of our study was to evaluate a three-dimensional gadolinium enhanced breath-hold MR angiography sequence using standard MR gradients in detecting renal artery stenosis. SUBJECTS AND METHODS: Forty-two patients referred for angiography for suspected renal artery stenosis underwent both conventional digital subtraction angiography (DSA) and MR angiography. MR angiography was performed on a 1.5-T scanner with standard gradients. A fast multiplanar spoiled gradient-echo sequence was used with the following parameters: TR/TE, 10.3/1.9; flip angle, 45 degrees; field of view, 36 x 32 cm; matrix size, 256 x 128; one excitation; volume thickness, 70 mm; and partitions, 28. Gadolinium was administered IV as a dynamic bolus of 30-40 ml. Conventional and MR angiographic images were interpreted by two radiologists in consensus. RESULTS: DSA revealed 87 renal arteries, of which 79 were in 35 patients with native kidneys and eight arteries were in seven patients with transplanted kidneys. Gadolinium-enhanced MR angiography showed 85 (98%) of 87 renal arteries. Seventeen patients had 20 significant (>50% stenosis) renal artery stenoses and five patients had five occluded renal arteries revealed by DSA. MR angiography revealed 85 renal arteries (98%), 20 stenoses (100%), and five occlusions (100%). Gadolinium-enhanced MR angiography led to one false-positive interpretation for renal artery stenosis and no false-negative interpretations. Thus, the sensitivity, specificity, and accuracy of MR angiography for renal artery stenosis were 100%, 98%, and 99%, respectively. CONCLUSION: The MR angiography pulse sequence we used was an effective and reliable technique for the diagnosis of renal artery stenosis. The sequence can be performed on widely available MR equipment that does not require fast gradient hardware. PMID- 10541106 TI - Interobserver variability in the interpretation of renal digital subtraction angiography. AB - OBJECTIVE: Our purpose was to analyze interobserver variability in the interpretation of renal digital subtraction angiography and to describe the main factors associated with observer discrepancies. MATERIALS AND METHODS: Forty-nine cases of unilateral atheromatous renal artery stenosis of more than 60% were quantified first by local investigators in a multicenter study and then by five other radiologists. Differences between radiologists for the minimum diameter (Dmin), the reference diameter (Dref), and the percentage of stenosis of the renal arteries were analyzed. Interpretations by the local investigators were then compared with the gold standard, defined as the mean for the five radiologists. RESULTS: The average SD for estimation of all renal artery stenoses by all radiologists was 7% for stenosis percentage, 0.5 mm for Dmin, and 0.7 mm for Dref. Main discrepancies occurred more frequently in cases of weakly opacified renal artery stenosis and poststenotic dilatation. The observations of local investigators disagreed by more than two SDs (14%) with the gold standard for 11 of 49 cases (22%). CONCLUSION: The accuracy of digital subtraction angiography in renal artery interpretations is poor because of variations in evaluating both Dmin and Dref. Precise and reproducible methods for quantification of renal artery stenosis are required. PMID- 10541107 TI - Sonographic detection of acute parenchymal injury in an experimental porcine model of renal hemorrhage: gray-scale imaging using a sonographic contrast agent. AB - OBJECTIVE: The purpose of this study was to determine the usefulness of contrast enhanced sonography in the detection of acute parenchymal injury. SUBJECTS AND METHODS: In a model of acute renal injury in pigs, four separate renal parenchymal bleeds were created by puncturing an interlobar artery of the upper and lower poles of the kidneys. B-mode gray-scale scans of the kidneys before and after injury, and after the administration of i.v. and intraarterial (i.a.) contrast agents were recorded on videotape for 5 min for each condition (baseline, after injury, after i.v. contrast administration, and after i.a. contrast administration). For each condition and injury, selected frames were analyzed with regions of interest of the normal renal parenchyma, the area of injury, and the perinephric space. Randomized videotape clips from each of the experimental conditions were rated by three sonologists as to the presence or absence of increased intrarenal parenchymal echogenicity, perinephric echogenicity, and confidence as to whether renal injury was present. RESULTS: Areas of renal injury were isoechoic with normal parenchyma on unenhanced scans. After both i.v. and i.a. contrast material injection, areas of injury were visible as areas of increased echogenicity. Contrast increased from 0.2 on unenhanced images to 4.0 and 4.5, respectively, after i.v. and i.a. administration of the new contrast agent. The three observers' ability to diagnose renal injury increased from 0.61, 0.64, and 0.54 to 0.71, 0.70, and 0.74 after i.v. injection and to 0.93, 0.92, and 0.97 after i.a. injection as indicated by the area under the curve in the receiver operating characteristic analysis. CONCLUSION: Transabdominal contrast-enhanced gray-scale sonography can reveal the area of acute renal hemorrhage. This procedure may be applicable in patients when sonographic contrast agents, imaging procedures, and modes of contrast administration are optimized for clinical use in trauma. PMID- 10541108 TI - Epidermoid cysts of the testicle: sonographic and MR imaging features. AB - OBJECTIVE: The purpose of this study is to analyze the appearance of testicular epidermoid cysts on high-resolution sonography and MR imaging and correlate imaging features with histopathologic findings. CONCLUSION: Intratesticular epidermoid cysts may show imaging features that correlate with their histopathologic findings. Concentric rings of alternating hypo- and hyperechogenicity on sonography and alternating high and low signal intensity on MR imaging ("onion ring" appearance) correspond to the pathologic finding of multiple layers of keratin debris. Absence of flow on color Doppler sonography and absence of contrast enhancement on MR imaging is also consistent with the avascular nature of these lesions. The ability of preoperative imaging studies to suggest the diagnosis of epidermoid cyst may prompt a testis-sparing surgery instead of an orchiectomy. PMID- 10541109 TI - Adnexal ring sign and hemoperitoneum caused by hemorrhagic ovarian cyst: pitfall in the sonographic diagnosis of ectopic pregnancy. PMID- 10541110 TI - Are malignant cells displaced by large-gauge needle core biopsy of the breast? AB - OBJECTIVE: The purpose of this paper is to determine the rate of tumor displacement resulting from large-gauge needle core biopsy in patients with breast carcinoma. MATERIALS AND METHODS: Three hundred fifty-two cancer excisions in patients who had undergone large-gauge needle core biopsy were evaluated for evidence of tumor displacement. Three needle procedures were compared: vacuum assisted, automated gun, and core biopsy guided by palpation. Needle track visualization, presence and amount of tumor displacement, tumor morphology, and interval between core biopsy and surgical excision were recorded for each case. RESULTS: Seventy-six cases showed tumor displacement of one or two cell clusters, and 38 cases-showed displacement of multiple tumor fragments. Tumor displacement was identified in 37% of automated gun specimens, 38% of specimens obtained with palpable guidance, and 23% of specimens obtained with a vacuum-assisted needle. Tumor displacement was seen in 42% of patients with an interval between biopsy and excision of less than 15 days, in 31% of patients with an interval of 15-28 days, and in 15% of tumors excised more than 28 days after core biopsy (p < .005). CONCLUSION: Tumor cell displacement was observed in 32% of patients who had undergone large-gauge needle core biopsy. The incidence and amount of tumor displacement was inversely related to the interval between core biopsy and excision. This relation suggests that tumor cells do not survive displacement. PMID- 10541112 TI - Clinical usefulness of MR imaging of the breast in the evaluation of the problematic mammogram. AB - OBJECTIVE: This study was undertaken to determine the usefulness of MR imaging of the breast as an adjunct to mammography in problematic cases in which the significance, presence, or location of an abnormality could not be determined. MATERIALS AND METHODS: From January 1993 through February 1998, 86 lesions for which histologic or mammographic follow-up was available were evaluated by breast MR imaging because of equivocal findings on mammography. MR studies were performed with a dedicated breast multicoil on a 1.5-T scanner. Early studies were done using a T1-weighted two-dimensional spin-echo sequence before and after the administration of contrast material. Later studies were performed using a three-dimensional fast spoiled gradient sequence with fat suppression. Studies were considered to be positive for an abnormality if a focal area of enhancement was seen after contrast administration. RESULTS: MR imaging had positive findings in 38 sites. Twenty-six of these sites corresponded in location to the mammographic abnormality that had prompted the recommendation for MR imaging. The remaining 12 sites occurred in areas not suspected mammographically. At biopsy, 10 (26%) of the 38 positive sites were malignant. MR imaging had negative findings at 60 other sites that had been suspected mammographically. Of these 60 sites, six were treated with excision, all with benign results; the remaining 54 sites showed mammographic stability on follow-up that ranged from 5 to 66 months (mean, 19 months). CONCLUSION: MR imaging of the breast can be a valuable adjunct to mammography for selected problematic cases. PMID- 10541111 TI - Complete percutaneous excision of infiltrating carcinoma at stereotactic breast biopsy: how can tumor size be assessed? AB - OBJECTIVE: The purpose of this study was to determine the frequency of complete excision of infiltrating carcinoma at stereotactic 11-gauge directional vacuum assisted breast biopsy and to evaluate the feasibility of measuring tumor size in stereotactic biopsy specimens in infiltrating carcinomas that were percutaneously excised. MATERIALS AND METHODS: We performed retrospective review of 51 infiltrating carcinomas diagnosed using stereotactic 11-gauge directional vacuum assisted biopsy that underwent subsequent surgery. For lesions yielding no residual infiltrating carcinoma at surgery, the maximal dimension of the tumor was measured in stereotactic biopsy specimens using ocular micrometry. RESULTS: In 10 (20%) (95% confidence intervals, 9.8-33.1%) of 51 infiltrating carcinomas diagnosed at stereotactic biopsy, surgery revealed no residual infiltrating carcinoma. Complete excision of infiltrating carcinoma was more frequent if 14 or more specimens were obtained (32% versus 0%, p < .004), if the mammographic lesion was removed (35% versus 7%, p < .03), and if the mammographic lesion size measured 0.7 cm or less (50% versus 16%, p = .08). Tumor size in stereotactic biopsy specimens was within 3 mm of mammographic lesion size in six (60%) of 10 lesions, including five (71%) of seven masses and one (33%) of three calcification lesions, but was smaller than the mammographic lesion size in eight (80%) of 10 lesions. CONCLUSION: Surgery revealed no residual infiltrating carcinoma in 10 (20%) of 51 infiltrating carcinomas diagnosed at stereotactic 11 gauge biopsy. Although tumor size can be assessed in stereotactic biopsy specimens in these lesions, such measurements may underestimate the maximal dimension of the tumor. Further study is needed to evaluate the usefulness of these measurements in guiding treatment decisions. PMID- 10541113 TI - Management of complex breast cysts. AB - OBJECTIVE: This study was undertaken to evaluate the various strategies currently in use to manage complex cysts and specifically address the need for intervention. MATERIALS AND METHODS: A review of 4562 breast sonograms obtained during an 18-month period revealed 308 complex cysts in 252 women. Data collected from review of patient records included the patient's age and risk factors for breast cancer, aspiration or biopsy results (or both), follow-up imaging studies, and management recommendations. RESULTS: Management recommendations for complex cysts were 1-year follow-up in 13 patients, 6-month follow-up in 148, sonographically guided aspiration in 82, aspiration with possible core biopsy in 62, and excisional biopsy in three. No malignancies were diagnosed in the group treated with follow-up imaging, sonographically guided aspiration, or excisional biopsy. One malignancy, a papilloma with a 3-mm focus of ductal carcinoma in situ, was diagnosed in one of the patients who underwent core biopsy. CONCLUSION: Of the lesions classified as complex cysts, the malignancy rate was 0.3% (1/308). This malignancy rate is lower than that for lesions classified as probably benign using mammographic criteria (i.e., for lesions classified as category 3 lesions using the Breast Imaging Reporting and Data System). Because the accepted standard practice for management of probably benign lesions is follow-up studies, the low yield of malignancy in this series suggests that complex cysts can be managed with follow-up imaging studies instead of intervention. PMID- 10541114 TI - Effect of patients' being prone during FDG PET for the diagnosis of breast cancer. PMID- 10541115 TI - Thymic hyperplasia after high-dose chemotherapy and autologous stem cell transplantation: incidence and significance in patients with breast cancer. AB - OBJECTIVE: The purpose of this study was to determine the incidence and clinical significance of thymic hyperplasia after high-dose chemotherapy and autologous stem cell transplantation for treatment of metastatic or high-risk primary (with at least four positive lymph nodes) breast cancer. MATERIALS AND METHODS: We retrospectively reviewed clinical records and CT scans of 102 breast cancer patients treated with high-dose chemotherapy and autologous stem cell transplantation. Patients were 26-63 years old (mean, 46 years). The length and width of the thymus gland were measured on serial CT scans obtained before and after treatment. Moderate thymic hyperplasia was recorded if a focal or diffuse increase was seen in the oblong, triangular soft-tissue opacity conforming to the configuration of the normal gland within the anterior mediastinum after therapy. Minimal hyperplasia was recorded when a minimal increase was seen in soft-tissue attenuation conforming to the configuration of the normal bilobed thymus gland within the anterior mediastinum, but no discrete mass was visible. RESULTS: CT showed no thymic hyperplasia in 91 (89%) of the 102 patients. CT showed thymic hyperplasia in the other 11 patients (11%). Three patients (3%) had moderate hyperplasia, and eight patients (8%) had minimal hyperplasia. When comparing patients with and without hyperplasia, we found no difference in mean age or survival. CONCLUSION: Thymic hyperplasia is rare after high-dose chemotherapy and autologous stem cell transplantation in adult patients with metastatic or high risk primary breast cancer. In this population, thymic hyperplasia does not appear to correlate with survival. PMID- 10541116 TI - CT-guided catheter drainage of loculated thoracic air collections in mechanically ventilated patients with acute respiratory distress syndrome. AB - OBJECTIVE: We report our experience with CT-guided percutaneous catheter drainage of loculated thoracic air collections in mechanically ventilated patients with acute lung injury or acute respiratory distress syndrome. MATERIALS AND METHODS: Nine critically ill patients had 17 air collections (13 pneumothoraces, three pneumatoceles, one tension pneumomediastinum) that either developed despite the presence of standard surgical chest tubes or were in loculated sites that were difficult to access. All nine patients were ventilated mechanically for a clinical diagnosis of acute respiratory distress syndrome. Catheter size ranged from 7- to 28-French. Response was measured by imaging follow-up, ventilatory parameters, and clinical outcome. RESULTS: On follow-up imaging studies, all 17 air collections were shown to have been evacuated successfully. Catheters remained in place for a mean of 11 days (range, 4-28 days). No major complications occurred. Sixteen air collections were treated successfully with CT guided catheter placement alone; the remaining air collection, a pneumothorax, was treated with subsequent placement of a chest tube by the surgeon at the patient's bedside. No surgery was undertaken for the air collections. Improvement in gas exchange was documented by increase in the hypoxemia ratio (arterial oxygen pressure divided by the inspired fraction of oxygen) in seven of 12 drainages; the other drainages were accompanied either by no improvement or by deterioration. Eight (89%) of the nine patients eventually were extubated and discharged from the hospital. The ninth patient died. CONCLUSION: CT-guided percutaneous catheter drainage provided effective treatment for loculated thoracic air collections and obviated surgical intervention in these critically ill, high-surgical-risk patients. PMID- 10541117 TI - Portable CT: assessing thoracic disease in the intensive care unit. AB - OBJECTIVE: The objective of this study was to assess the usefulness of a portable CT scanner to evaluate and treat thoracic disease in patients in the intensive care unit. MATERIALS AND METHODS: Fourteen patients who were being treated in the intensive care unit underwent 20 portable CT scans. Twice a CT scan was obtained to guide an interventional chest procedure. The remaining 18 scans were assessed for findings not evident on portable chest radiography and for findings that altered treatment. Image quality was judged in comparison with fixed CT scans. RESULTS: Unsuspected abnormalities, most relating to the pleura or chest wall, were found in 13 of the 17 available portable CT scans. Treatment was affected in four (25%) of the 16 cases in which medical records were available for review. Two interventional procedures were performed successfully using portable CT guidance. Scan quality was judged to be comparable with that of fixed CT for mediastinal windows and somewhat inferior for lung windows. CONCLUSION: Portable CT gives images of diagnostic quality and allows confident guidance during interventional procedures in critically ill patients who therefore need not leave the intensive care unit environment. PMID- 10541118 TI - Predicting proper endotracheal tube placement in underexposed radiographs: tangent line of the aortic arch. AB - OBJECTIVE: We investigated an indirect radiographic means of predicting proper endotracheal tube placement on underexposed portable chest radiographs. CONCLUSION: When an endotracheal tube is 3.4-5.0 cm above the tangent line of the aortic arch, it is in proper position 95% of the time. PMID- 10541119 TI - Iatrogenic bronchopleural cutaneous fistula. PMID- 10541120 TI - CT diagnosis of laceration of the main pulmonary artery after blunt trauma. PMID- 10541121 TI - Sonography of the normal fetal heart: a practical approach. PMID- 10541122 TI - Prenatal sonography of congenital renal malformations. PMID- 10541123 TI - Intrahepatic pregnancy: sonography and CT findings. PMID- 10541124 TI - MR imaging findings of lateral ulnar collateral ligament abnormalities in patients with lateral epicondylitis. AB - OBJECTIVE: The purpose of this paper was to use MR imaging to determine whether a relationship exists between lateral epicondylitis and abnormalities of the lateral ulnar collateral ligament. SUBJECTS AND METHODS: The study group comprised 35 consecutive patients who were referred for MR imaging to rule out lateral epicondylitis. On MR imaging, "lateral epicondylitis" was defined as increased signal intensity of the extensor tendons close to their insertion on the lateral epicondyle. The severity of the lateral epicondylitis was graded as mild, moderate, or severe. The origin of the lateral collateral ligamentous complex was characterized, and the lateral ulnar collateral ligament was graded as normal, thickened, partially torn, or torn. Eleven patients underwent elbow surgery after the initial MR examination. RESULTS: In 15 patients, MR imaging revealed characteristics of mild lateral epicondylitis. In 13 of these patients, the lateral ulnar collateral ligament was normal; one patient showed a thickened ligament; and one patient had a thinned ligament. In 11 patients, MR imaging showed features of moderate lateral epicondylitis. In eight of these patients, the lateral ulnar collateral ligament was thickened, and in the remaining three patients the ligament was normal. All nine patients with severe lateral epicondylitis showed abnormalities of the lateral ulnar collateral ligament on MR imaging. In one of these patients the lateral ulnar collateral ligament was thickened, in three patients we saw a partial tear, and in the remaining five patients we saw a complete tear of the ligament. CONCLUSION: In our study, MR imaging features of lateral epicondylitis were often associated with thickening and tears of the lateral ulnar collateral ligament. PMID- 10541125 TI - MR imaging of subchondral osteonecrosis of the vertebral body after percutaneous laser diskectomy. AB - OBJECTIVE: We describe four cases of subchondral osteonecrosis of the vertebral body that occurred after percutaneous laser diskectomy. Follow-up MR imaging after laser intervention showed abnormal findings in the vertebral body immediately adjacent to the site of diskectomy that are consistent with subchondral osteonecrosis. CONCLUSION: MR imaging features of this complication include a wedge-shaped low signal intensity on T1-weighted images, high and low signal intensities on T2-weighted images, and a contrast-enhanced area corresponding to high signal intensity on T2-weighted images. Possible causative mechanisms include thermal injury and photoacoustic shock. PMID- 10541126 TI - MR imaging of penetrating spinal trauma. PMID- 10541128 TI - Using CO2-enhanced arteriography to investigate acute gastrointestinal hemorrhage. AB - OBJECTIVE: The objective of the study was to compare the efficacy of CO2-enhanced arteriography with that of arteriography enhanced with standard iodinated contrast material in the detection of gastrointestinal hemorrhage. CONCLUSION: For acute gastrointestinal hemorrhage, we did not find an advantage to using CO2 to show active bleeding. In fact, CO2 was inferior to iodinated contrast material in revealing nonbleeding vascular anomalies. PMID- 10541127 TI - Prospective randomized comparison of valved versus nonvalved peripherally inserted central vein catheters. AB - OBJECTIVE: The purpose of this study was to evaluate whether a valved peripherally inserted central catheter (PICC) design would result in a lower incidence of occlusion, infection, and malfunction than a clamped catheter. SUBJECTS AND METHODS: Three hundred sixty-two study patients (233 men, 129 women; mean age, 44 years) were randomized to receive a clamped (n = 182) or valved (n = 180) 5-French single-lumen PICC. Catheters were placed under fluoroscopic (n = 331) or sonographic guidance (n = 31). The valved PICC was flushed with saline solution, and the clamped PICC was flushed with a heparin-saline solution. All patients were prospectively followed up at least weekly for catheter status and complications. RESULTS: Percutaneous placement with the catheter tip in the central veins was successful in 99% of patients. Mean dwell time was 34 days. Twenty-six occlusive or infectious complications occurred in the clamped catheter group and 12 in the valved catheter group (p = .02). The clamped and valved catheter groups had 13 and five occlusions, respectively (p = .06), and 12 and five catheter-related blood stream infections, respectively (p = .09). Most occlusions (68%) were treated successfully with urokinase, and site infection or sepsis was treated by catheter removal. CONCLUSION: We found a statistically significant difference in the complication rate for the valved PICC compared with the clamped PICC. With the valved PICC, occlusion and infection were reduced, and patients having these catheters did not require heparin flushes. PMID- 10541129 TI - Gadolinium-based contrast agents in angiography and interventional radiology. PMID- 10541130 TI - Attenuation difference between tumor and surrounding normal pancreas. PMID- 10541131 TI - Wherefore ABBI? Advanced Breast Biopsy Instrumentation. PMID- 10541132 TI - ACR standard for luminance and illuminance. American College of Radiology. PMID- 10541133 TI - Bilateral adnexal hydatidosis in primary infertility. PMID- 10541134 TI - Myoepithelioma of the nasal piriform aperture: CT findings. PMID- 10541135 TI - Subarachnoid gadolinium enhancement mimicking subarachnoid hemorrhage on FLAIR MR images. Fluid-attenuated inversion recovery. PMID- 10541136 TI - Detection of primary pericardial liposarcoma by CT. PMID- 10541137 TI - "Hilar eyes," or the Gatling gun revisited. PMID- 10541138 TI - Unexpected hip arthrogram during cystography in a patient with extraperitoneal bladder rupture and acetabular fracture. PMID- 10541140 TI - Wish list for the new millennium PMID- 10541139 TI - Leaders of the pack. PMID- 10541141 TI - Madman in the OR. PMID- 10541142 TI - Pathomechanics and clinical relevance of disc degeneration and annular tear: a point-of-view review. AB - Annular tear is a major cause of intervertebral disc degeneration that results in disabling back pain. Many of the stresses resulting in this type of lesion are common in the workplace: compression, torsion, compression combined with flexion, and vibration. Age-related disc degeneration begins early in adulthood, and progresses thereafter, altering disc morphology and mechanical properties in ways that predispose to disc herniation, and should not be misconstrued as "old age." Acute trauma may produce disc herniation whether or not there are predisposing factors, such as age-related degeneration, but disc herniation in the absence of acute injury requires the presence of preexisting degenerative changes. PMID- 10541143 TI - Geriatric intertrochanteric hip fractures: an economic analysis. AB - Hip fractures have a high economic and social cost to society. Rising healthcare costs, coupled with managed healthcare systems, have forced a close inspection of healthcare expenditures. To evaluate the impact of geriatric intertrochanteric hip fractures upon a hospital system, an economic cost-analysis was undertaken. An analysis was made of financial and hospital data of elderly patients who sustained an intertrochanteric hip fracture and subsequently underwent open reduction and internal fixation. Increasing patient age did not correlate with length of stay, overall hospital cost, or net hospital income. When costs were subdivided, there were no statistically significant differences relative to patient age for costs of pharmacy, radiology, respiratory therapy, or operating room supply. There were, however, statistically significant correlations between patient age and costs of the operating room, blood, and anesthesia. During the 36 month study period, a case manager was added to the orthopedic surgical team. There was a trend toward decreased length of stay after the addition of the case manager, but the overall cost of patient care was not affected. PMID- 10541144 TI - Evaluation of the upper sacrum by three-dimensional computed tomography. AB - Axial and sagittal computed tomographic (CT) scans of 40 sacrum specimens were obtained. The measurements of the upper sacral canal and S1-2 and S2-3 anterior sacral foramina were performed on axial scans, and the evaluation of the upper sacral pedicle was based on sagittal scans. The results showed that there were statistically significant differences between male and female specimens in 3 of 29 measurements. In general, the measurements of male specimens were slightly larger than those of the female specimens, and the linear dimensions of the sacral canal, anterior foramina, and pedicle decreased from the S1 to S3. This study indicated that the critical area of the sacral pedicle for screw insertion lies in the junction between the pedicle and vertebral body. CT scans provide more accurate information about the essential sacral anatomy. PMID- 10541145 TI - Traumatic osteochondral defect of the first metatarsal head: a case report. AB - Osteochondral injury of the first metatarsophalangeal joint is described in most literature as "osteochondritis dissecans" and an early stage of hallux rigidus. Traumatic osteochondral lesions of the knee and ankle are relatively common and well described. A case of a traumatic osteochondral defect of the first metatarsal head is presented. PMID- 10541146 TI - Acute compartment syndrome complicating a distal tibial physeal fracture in a neonate. AB - This case report of a neonate who developed an acute compartment syndrome secondary to a minimally displaced distal tibial physeal injury represents the youngest patient to be reported with such a condition. After undergoing emergency four-compartment decompression fasciotomies, the 4-week-old child had a return of normal neuromuscular function and anatomic remodeling of the fracture. It is difficult to diagnose compartment syndrome in a neonate. The patient can neither give a history, nor follow commands to cooperate with the exam. The physician must rely primarily on the physical examination; however, the quantitative measurement of intracompartmental pressure can corroborate the diagnosis of compartment syndrome. We have found using a monometer to measure intracompartmental pressure to be helpful in conjunction with a physical exam when evaluating a neonate suspected of having a compartment syndrome. PMID- 10541147 TI - Infection in total knee arthroplasty: Part II. Treatment. PMID- 10541148 TI - Posterior fracture through the sternoclavicular physis associated with a clavicle fracture: a case report and literature review. AB - Three reported cases of fractures in teenagers of the medial clavicle associated with a posterior disruption at the sternoclavicular joint have similar findings. The medial fragment is rotated 90 degrees to the coronal plane. The medial fragment is usually stripped of its periosteum. Treatment requires open reduction and internal fixation of the clavicle fracture. We are reporting similar findings in a fourth case. PMID- 10541150 TI - Visual complaints from healthy children. AB - It is common for healthy children with specific visual complaints to be seen for eye examinations. After a complete eye examination has ruled out pathologic conditions as the cause of these complaints, it is appropriate for the clinician to explore the possibility that normal entoptic or physiologic visual phenomena might have provoked the child's report of vision problems. Some of these normal visual experiences are frequent causes of children's complaints of vision problems, such as physiologic diplopia, relaxation of the near synkinesis during reading, and vitreous body floaters. Some complaints are common, even though the underlying entoptic or physiologic phenomenon may be speculative or obscure, such as the report that objects look bigger or smaller than they actually are. When the clinician encounters such situations, the parents and the child will be much more satisfied by an explanation of the normal system anatomy and physiology than by the simple reassurance that everything is all right. PMID- 10541149 TI - Iatrogenic choroidal neovascularization. AB - Iatrogenic choroidal neovascularization is an uncommon complication of laser photocoagulation and other ocular surgical procedures. It appears to be the result of a number of conditions, including damaged Bruch's membrane and/or retinal pigment epithelium, whose reparative processes trigger the release of angiogenic factors. Inflammatory cells and choroidal ischemia may also play a role. The prognosis varies depending on the underlying disease and the type of choroidal neovascularization (subretinal, chorioretinal, or choriovitreal). Minimizing the amount of laser energy used during laser procedures, avoiding repeat laser treatment to the same retinal area, and minimizing direct mechanical trauma to the retinal pigment epithelium and choroid decrease the chance of inducing iatrogenic choroidal neovascularization. PMID- 10541151 TI - Phacoemulsification and modern cataract surgery. AB - The techniques and results of cataract surgery have changed dramatically during the past three decades. In the USA, we have moved from intracapsular cataract extraction as the preferred technique to almost exclusively extracapsular techniques. Smaller incisions have become the standard, with phacoemulsification now being the method of choice for most surgeons. Along with these advances have come improved intraocular lens materials and designs, especially well suited for use with smaller incisions. Phacoemulsification as a method to remove the cataractous lens was first proposed more than 20 years ago. Advances in techniques and equipment have led to a dramatic increase in the popularity of phacoemulsification with increased safety and efficiency. Viscoelastic agents have been developed synchronously with modern phacoemulsification techniques, playing an integral role in the success of this new technology. Improved surgical techniques for removing the anterior lens capsule have decreased the incidence of both intraoperative and postoperative capsular complications. Nucleus removal, formerly performed primarily in the anterior chamber, is now performed in the posterior chamber, decreasing damage to the corneal endothelium. Improved wound construction allows many wounds to be left unsutured, and smaller wounds allow shorter recovery time and greater intraoperative control and safety. Intraocular lenses can have smaller optic sizes and still maintain accurate centration. Foldable intraocular lenses can take advantage of the smaller incision, even further shortening the time to visual recovery. Continual evolution of this technology promises to further improve patient outcomes after cataract surgery. PMID- 10541152 TI - A bad eye and a sore lip. AB - A 48-year-old woman developed painful visual loss in the left eye, meningismus, and painful oral ulcers. Magnetic resonance imaging of the brain with gadolinium demonstrated enhancement of the left optic nerve. Lumbar puncture showed a lymphocytic pleocytosis, and a biopsy specimen of one of the oral ulcerations was consistent with Behcet's disease. Epidemiologic factors and diagnostic criteria for Behcet's disease are discussed. PMID- 10541153 TI - Sildenafil (Viagra) and ophthalmology. AB - Sildenafil citrate (Viagra) is a new oral medication that inhibits phosphodiesterase-5 (PDE5) in the corpus cavernosum to facilitate penile erection for the treatment of male impotence. The drug also has a mild inhibitory effect on PDE6, which controls the level of cyclic guanosine monophosphate in the retina, and it may cause a perception of bluish haze or increased light sensitivity in some patients. Long-term retinal damage has not been reported, but long-term electroretinographic studies have not been performed. Sildenafil causes a mild lowering of blood pressure and is absolutely contraindicated in patients taking any form of nitrate medication. A number of cardiovascular deaths and retinal vascular events in patients taking sildenafil have been reported, but so far the rate of these complications does not exceed expectation for an elderly population. Ophthalmologists should alert patients to the ocular side effects and potential risks of this new drug until further clinical experience has been obtained. PMID- 10541154 TI - What ailed Goya? AB - At age 46, Francisco de Goya (1746-1828) suffered from a severe illness that lasted several months. It caused loss of vision and hearing, tinnitus, disorientation, weakness, abdominal distress, and general malaise. After a few months he recuperated but was left deaf forever. In addition to the physical effects, his emotional health and artwork were affected. The precise cause of this illness has long been debated. One early, but unlikely, hypothesis was that he had syphilis. Later conjectures have included Vogt-Koyanagi-Harada disease and lead toxicity. Cogan's syndrome and vasculitis are additional possibilities, although neither is likely to have been Goya's diagnosis. An infectious disease such as meningitis, encephalitis, or malaria is far more likely. Quinine toxicity (cinchonism) may have complicated the illness. PMID- 10541155 TI - Aligning the Goldmann tonometer tip by means of the "precontact whitish rings". AB - A set of whitish rings observed before contact is made during Goldmann-type applanation tonometry can help simplify and speed the examination. The orientation and size of the rings are useful both in aligning the tonometer and in estimating the distance remaining before contact is made. Relying on these whitish rings can also avoid some of the shear-related damage exerted on the corneal epithelium from alignment movements of the tonometer tip after contact with the cornea has been made. PMID- 10541156 TI - Congenital cystic eye: report of two cases and review of the literature. AB - A 13-month-old boy and a 2-week-old girl, who were considered to be anophthalmic and who later each developed a cystic lesion in the left orbit with protrusion of the lower eyelid, were studied. The fellow eye in case 1 was subsequently found to be microphthalmic with cyst and was normal in case 2. Histopathologic study of each case revealed a cyst lined externally by dense fibrous connective tissue to which skeletal muscle and adipose tissue were attached. The inner aspect of the cyst was lined by neuroglial tissue, possible immature retinal tissue, and cuboidal epithelium. No fully developed ocular structures or microphthalmos were identified. Fourteen cases of congenital cystic eye, including our cases, have been published in the English-language literature since 1964. We discuss and illustrate the findings in our cases and 10 others in which histopathologic findings were reported. Congenital cystic eye, microphthalmos with cyst, and microphthalmos with cystic teratoma should be suspected in patients with a small or unrecognizable eye and an orbital cystic mass that is detected by palpation or visualization. PMID- 10541157 TI - Strabismus: accommodative component treated by LASIK. PMID- 10541158 TI - A plasticity for blood vessel remodeling is defined by pericyte coverage of the preformed endothelial network and is regulated by PDGF-B and VEGF. PMID- 10541159 TI - Molecular diagnosis of bilateral coronal synostosis. AB - The authors performed a prospective study evaluating molecular diagnosis in patients with bilateral coronal synostosis. The patients were divided into two groups: (1) those clinically classified as having Apert, Crouzon, or Pfeiffer syndrome and (2) those clinically unclassified and labeled as having brachycephaly. Blood samples were drawn for genomic DNA analysis from 57 patients from 1995 to 1997. Polymerase chain reactions were performed using primers flanking exons in FGFR 1, 2, and 3. Each exon was screened for mutations using single-strand confirmation polymorphism, and mutations were identified by DNA sequencing. Mutations in FGFR2 or FGFR3 were found in all patients (n = 38) assigned a phenotypic (eponymous) diagnosis. All Apert syndrome patients (n = 13) carried one of the two known point mutations in exon 7 of FGFR2 (Ser252Trp and Pro253Arg). Twenty-five patients were diagnosed as having either Crouzon or Pfeiffer syndrome. Five patients with Crouzon syndrome of variable severity had mutations in exon 7 of FGFR2. Fifteen patients (12 with Crouzon, 3 with Pfeiffer) had a mutation in exon 9 of FGFR2, many of which involved loss or gain of a cysteine residue. A wide phenotypic range was observed in patients with identical mutations, including those involving cysteine. Two patients labeled as having Crouzon syndrome had the Pro250Arg mutation in exon 7 of FGFR3. All three patients with the crouzonoid phenotype and acanthosis nigricans had the same mutation in exon 10 of FGFR3 (Ala391Glu). This is a distinct disorder, characterized by jugular foraminal stenosis, Chiari I anomaly, and intracranial venous hypertension. Mutations were found in 14 of 19 clinically unclassifiable patients. Three mutations were in exon 9, and one was in the donor splice site of intron 9 on FGFR2. The most common mutation discovered in this group was Pro250Arg in exon 7 of FGFR3. These patients (n = 10) had either bilateral or unilateral coronal synostosis, minimal midfacial hypoplasia with class I or class II occlusion, and minor brachysyndactyly. No mutations in FGFR 1, 2, or 3 were detected in five patients with nonspecific brachycephaly. In conclusion, a molecular diagnosis was possible in all patients (n = 38) given a phenotypic (eponymous) diagnosis. Different phenotypes observed with identical mutations probably resulted from modulation by their genetic background. A molecular diagnosis was made in 74 percent of the 19 unclassified patients in this series; all mutations were in FGFR2 or FGFR3. Our data and those of other investigators suggest that we should begin integrating molecular diagnosis with phenotypic diagnosis of craniosynostoses in studies of natural history and dysmorphology and in analyses of surgical results. PMID- 10541160 TI - Complications of systemic corticosteroid therapy for problematic hemangioma. AB - Systemic corticosteroid therapy has been used to treat hemangiomas for 30 years; yet, there are no studies of possible complications. We reviewed the database of the Vascular Anomalies Center at the Boston Children's Hospital and gathered information on short- and long-term side effects in children who were given systemic corticosteroids for problematic hemangiomas. In addition, a questionnaire regarding early and late consequences was sent to the families of children who were treated with corticosteroids from 1983 to 1997. Of 300 patients with hemangiomas, 80 children were identified as having received a full course of systemic corticosteroids for problematic tumors. Complete data were collected on 62 of these children. The response rate to the questionnaire was 78 percent (n = 62 of 80). The initial dose of corticosteroid varied from 2 to 3 mg/kg/ day. Duration of therapy ranged from 2 to 21 months (mean, 7.9 months; median, 6.5 months). The follow-up interval from the cessation of therapy ranged from 6 months to 15 years (mean, 4 years; median, 3 years). Short-term complications included cushingoid facies (n = 44; 71 percent), personality changes (n = 18; 29 percent), gastric irritation (n = 13; 21 percent), fungal (oral or perineal) infection (n = 4; 6 percent), and diminished gain of height (n = 22; 35 percent) and weight (n = 26; 42 percent). A total of 91 percent of children who had diminished gain of height (n = 20) returned to their pretreatment growth curve for height by 24 months of age. One child, who was treated at another institution with a dose of 20 mg/kg/day for 6.5 months that was slowly tapered over 18 months, was petite 6 years after ending therapy. Another child treated with an initial dose of 2 mg/kg/day for 5 months was smaller than predicted at the age of 6 years, but she was born prematurely and was on ventilatory support for respiratory distress. Three children treated with the standard dose and duration were at a low percentile for weight 4, 5, and 10 years after the cessation of therapy. Statistical analysis showed a correlation between diminished gain of height with duration of therapy and age at initiation of treatment. One child had corticosteroid myopathy that resolved with cessation of therapy. We found no evidence for immunologic suppression, i.e., there was no increase in the number of bacterial infections during corticosteroid administration. In conclusion, systemic corticosteroids can be safely given to treat endangering hemangiomas in infants at doses of 2 to 3 mg/kg/day, which are slowly tapered and stopped before the age of 1 year. Short-term side effects were minor and transient, and no serious long-term complications occurred. PMID- 10541161 TI - Long-term comparison of a newly designed gold implant with the conventional implant in facial nerve paralysis. AB - Patients with complete facial nerve palsy are at risk for eye complications resulting from exposure of the cornea and loss of the blinking reflex. Failure of protection predisposes the patient to exposure keratitis, corneal abrasion and, in rare cases, blindness. The mainstays of non-surgical therapy are cumbersome, obscure vision, and are mostly helpful in patients with acute facial paralysis in whom recovery of orbicularis oculi function is expected. Methods of lid-loading using metal implants and gold eyelid weights have been reported in the literature. Between October of 1988 and March of 1995, 32 patients with lagophthalmos due to facial nerve palsy underwent a total of 34 procedures for the insertion of a gold eyelid weight. Each patient had a gold weight inserted into a small pocket between the orbicularis oculi and the tarsal plate of the upper eyelid. The gold implant is curved to fit the curvature of the eye and contains holes for fixation to the tarsus with sutures. Ingrowth of fibrous tissue through the holes may also help fix the weight in position. Between 1988 and 1991, 10 patients received 10 commercially available rectangular gold implants with 2 holes; these implants resulted in adverse effects, such as infection and exposure in up to 30 percent of the cases. Because of the high complication rate with the rectangular gold implant, the authors began using a new, elliptical gold implant with 3 holes, which is longer, thinner, wider in the center, and narrower in the peripheral portion. This new elliptical implant was used on 22 patients (24 implants) from December of 1991 through March of 1995. The mean follow-up time for the 32 patients in the study was 41.3 months (range, 6 to 63 months), 49.8 months for patients with rectangular implants and 32.8 months for patients with elliptical implants. The elliptical gold implant resulted in dynamic closure of the eyelid and in excellent protection and cosmesis. Lagophthalmos and exposure keratitis resolved, visual acuity significantly improved without complications, and most patients could dispense with eyedrops and salves. A lower eyelid supporting procedure (conchal cartilage graft) should be performed simultaneously in patients with lagophthalmos of a moderate or severe degree to achieve complete closure of the eyelid. Use of a tall pillow decreased the incidence of eyelid opening during sleep. Double eyelid fold operations'were performed on the contralateral eyelid after 6 months, resulting in a symmetrical and beautiful eyelid. PMID- 10541162 TI - Progress in larynx-sparing surgery for glottic cancer through tracheal transplantation. AB - The current surgical treatment for unilateral, advanced glottic cancer is a total laryngectomy. Usually, the noninvolved hemilarynx needs resection because the resulting laryngeal defect cannot be reconstructed after adequate tumor resection. Experimental findings suggest that segments of autologous trachea may restore extended laryngeal defects. The authors used tracheal transplantation to save laryngeal function after the removal of advanced glottic cancer. In this case series review, 10 patients were treated during a 1.5-year period, with an average follow-up of 8 months. Evaluated factors included survival of the tracheal transplant and functional outcome with regard to the onset and quality of the airway, speech, and deglutition. The authors showed that segments of cervical trachea may restore extended laryngeal defects after initial revascularization by a radial forearm fascial flap. The fascial flap served as a vascular carrier for the transplanted trachea. Follow-up showed the stability of the reconstruction. Compared with a total laryngectomy, a striking improvement in patient comfort and function was noticed. Transplantation of the trachea is a technique that may save laryngeal function after the treatment of advanced-stage glottic cancer. These findings may improve laryngeal preservation strategies in treating laryngeal cancer. PMID- 10541163 TI - Somatosensory status after pedicled or free TRAM flap surgery: a retrospective study. AB - The transverse rectus abdominis musculocutaneous (TRAM) flap is widely used in autologous breast reconstructions. In transferring tissue as a pedicled or free flap, alterations in sensibility are unavoidable. This study evaluated somatosensory function in the reconstructed breast at least 2 years after pedicled or free TRAM flap surgery. Thirteen patients who had a pedicled TRAM flap and 13 patients who had a free TRAM flap participated in the study. The patients completed a questionnaire regarding subjective sensibility and their general opinion of the reconstructed breast. Somatosensory examinations to study the sensations of touch, warmth, cold, and pain were performed using nonquantitative and quantitative techniques. An age-matched control group of eight women who had never had breast surgery was also examined because the majority of the women who had breast reconstruction also had a mammaplasty performed on the contralateral breast, which disqualified it as a control. The majority of the patients reported that the reconstructed breast felt like a real breast. However, sensibility to touch, warmth, cold, and pain was decreased in the study groups compared with the control group. No clinically significant differences existed in sensibility between the pedicled and free TRAM flap groups. PMID- 10541164 TI - Thoracodorsal vessels as recipient vessels for the free TRAM flap in delayed breast reconstruction. AB - The internal mammary vessels have been recommended as the first choice recipient vessels for delayed breast reconstruction with the free TRAM flap. This approach has avoided surgery in the previously operated axilla, has required a shorter pedicle length, and has allowed for more medial placement of the TRAM tissue. Frequency of nonusable axillary vessels has been reported at 11 percent, with a 6 percent incidence of flap loss in the delayed reconstructive setting. We reviewed our experience with the thoracodorsal vessels as recipient vessels in delayed free TRAM breast reconstruction to assess more accurately the adequacy of these potential recipient vessels. All patients undergoing delayed TRAM reconstruction were reviewed. Forty-seven of 300 consecutive TRAM procedures were for planned delayed free reconstruction. In seven of the patients (15 percent), the thoracodorsal vessels were found to be inadequate for free reconstruction. A supercharged pedicled TRAM was used for reconstruction in each of these seven patients. Average operating room time was 7 hours. Mean follow-up time was 38 months. Nineteen percent of all patients developed at least one complication. Twelve percent of free TRAM patients developed a complication, whereas 57 percent of supercharged patients developed a postoperative complication. The difference in complication rates was statistically significant. The thoracodorsal vessels have provided an adequate recipient vessel in 85 percent of delayed free TRAM reconstructions, comparable to previous reports. Pedicling and supercharging the flap, in those situations in which the thoracodorsal vessels were inadequate, were associated with an increased incidence of postoperative complications. This suggests that in the delayed reconstructive setting, higher-risk patients benefit from free reconstruction over supercharged reconstructions. A second recipient vessel should be used when the thoracodorsal vessels are inadequate for planned free TRAM reconstruction. In these circumstances, we would recommend the use of the internal mammary vessels followed by the thoracoacromial vessels as reliable alternative recipient sites for delayed free TRAM reconstruction. PMID- 10541165 TI - Internal mammary vessels as a model for power Doppler imaging of recipient vessels in microsurgery. AB - The aim of this interdisciplinary study was to evaluate power Doppler imaging as a method of collecting reliable preoperative data concerning the diameters and topography of exemplary internal mammary vessels as recipient vessels in reconstructive microsurgery. Thirteen female patients (range, 37 to 58 years; mean, 45.6 years) were examined preoperatively with power Doppler imaging from the first to the fifth intercostal space parasternally and bilaterally. These data are compared with measurements obtained intraoperatively in each individual. Mean velocity in the artery in the second intercostal space on the right side is 47.11 cm/sec (range, 15 to 90 cm/sec) and on the left side is 42.25 cm/sec (range, 18 to 95 cm/sec). Mean velocity in the vein in the second intercostal space on the right side is 17.80 cm/sec (range, 10 to 30 cm/sec) and on the left side is 13.06 cm/sec (range, 5.3 to 32 cm/sec). The topographic results are in close agreement with intraoperative measurements and previous anatomical studies. Sonographic preoperative data of arteries (mean, 1.88 mm) show slightly smaller diameters than intraoperative measurements (mean, 2.08 mm), whereas veins show slightly larger diameters in sonography (mean, 2.33 mm) than intraoperatively (mean, 2.12 mm). Mean sonographic diameter of artery ranges from 2.14 mm (second intercostal space) to 1.46 mm (fifth intercostal space), of the vein from 2.76 (second intercostal space) to 1.25 mm (fifth intercostal space). In one case, a vein was not detectable. This noninvasive method leads to confirmation of the preoperative choice of the optimal recipient vessels for free tissue transfer and does not harm the patient. PMID- 10541166 TI - The premature removal of tissue expanders in breast reconstruction. AB - The role of tissue expanders in breast reconstruction is well established. Little information exists, however, regarding the incidence and etiology of premature removal of the tissue expander before planned exchange to a permanent breast implant. The purpose of this study was to review our 10-year experience with tissue expander breast reconstruction and identify factors relating to the premature removal of the tissue expander. This study is a retrospective review of 770 consecutive patients who underwent breast reconstruction with tissue expanders over the past 10 years. Breast reconstruction was immediate in 90 percent of patients. Patients were expanded weekly, and adjuvant chemotherapy was begun during the expansion process when required. Factors potentially affecting premature expander removal (chemotherapy, diabetes, obesity, radiation therapy, and smoking) were evaluated. Fourteen patients (1.8 percent) with a mean age of 47 years (range, 38 to 62 years) required premature removal of their tissue expander. Expanders were removed a mean of 3.2 months (0.1 to 8 months) after insertion. Causes for premature removal of the tissue expander included infection (7 patients), exposure (2), skin necrosis (2), patient dissatisfaction (2), and persistent breast cancer (1). Positive wound cultures were obtained in four of the seven infected patients (57 percent), requiring expander removal for infection. Tissue expanders were removed in 11 patients for complications directly related to the expander. Among these, six (55 percent) were receiving adjuvant chemotherapy, and one was a smoker. Diabetes, obesity, other concomitant medical illnesses, and prior mantle irradiation were not associated with expander removal. Premature removal of the tissue expander was required in only 1.8 percent of the patients in this series. Infection was the most common complication necessitating an unplanned surgical procedure to remove the expander. This study demonstrates that the use of tissue expanders in breast reconstruction is reliable, with the vast majority of patients completing the expansion process. PMID- 10541167 TI - Angiogenesis and vascular growth factor receptor expression in malignant melanoma. AB - The growth and metastases of many solid tumors are dependent on the recruitment of new blood vessels. Tumor angiogenesis is most likely initiated by paracrine release of growth factors that bind to their corresponding endothelial cell surface receptors. To determine whether angiogenesis and growth factor receptor expression are consistent findings in malignant melanoma, primary human melanomas were examined for mRNA expression of receptors for fibroblast growth factors (FGFR-1, FGFR-2), vascular endothelial growth factor (VEGFR-1, VEGFR-2), and the receptors Tiel and Tie2. Charts were reviewed and archival formalin-fixed, paraffin-embedded primary tumors were obtained from patients with thin (<1 mm; n = 10), intermediate (1 to 4 mm; n = 10), or thick malignant melanoma (>4 mm; n = 8). Also examined was whether melanoma cell lines could induce endothelial growth factor receptor synthesis by metabolic labeling. It was found that tumor vascularity did not correlate with clinical stage, melanoma thickness, or clinical outcome. It was also found that melanoma cell lines were not capable of directly regulating endothelial cell synthesis of growth factor receptors. However, expression of Tiel and VEGFR-2 mRNA by the tumor vasculature in select stage IA-IIB patients, and FGFR-1 mRNA expression by the tumor cells in the same clinical stages was found. The expression of these growth factor receptors did not correlate with clinical outcome. These data suggest that angiogenesis is not a prominent characteristic of primary malignant melanoma lesions and that the endothelial cell expression of Tiel and VEGFR-2 in vivo is probably not directly induced by the tumor. PMID- 10541168 TI - Geometric limit of multiple local Limberg flaps: a flap design. AB - The Limberg rhombic flap is a reliable and widely used technique in head and neck surgery. Since Limberg introduced his original design in 1946, several modifications of the technique have been described. Although a single Limberg flap is frequently used at the face to close small to medium defects, multi Limberg flap techniques can help the surgeon to cover moderate to large defects of the extremities, trunk, and back. In this study, a design of four neighboring local Limberg flaps to cover a moderate to large defect without using a skin graft is introduced. It is believed that this design is the geometric limit of multiple Limberg flaps that can entirely cover a single large rhombic defect, because one Limberg flap unit can only be adjoined by three others, one from the tip and two from the sides. This flap design of four local Limberg flaps is also the only geometrically possible design that can keep all the bases of these four flaps free of incisions if one attempts to prepare four small Limberg flaps around a large rhombic defect. PMID- 10541169 TI - Limb-sparing surgery with reinnervated free-muscle transfer following radical excision of soft-tissue sarcoma in the extremity. AB - Limb-sparing surgery is the preferred approach in the management of patients with high-grade soft-tissue sarcomas when local disease can be completely resected. However, conventional treatment focuses only on restoration of basic functions to the remnant limb. Lost functions are not restored to normal, leaving the patient with variable degrees of functional disabilities. This in turn may necessitate further massive reconstructive procedures. Transferred reinnervated free muscles were used to reconstruct functions lost after radical resection of malignant soft tissue sarcoma of the extremities in 17 patients. The long-term functional outcome included survival of transplanted muscle, speed of neural recovery, and muscle strength and disabilities. All muscles survived. Postoperative follow-up ranged from 27 to 106 months. All muscles except those in a 75-year-old patient were successfully reinnervated. Powerful strength and almost normal limb functions were obtained. Functional scoring of the patients according to the rating system of the Musculoskeletal Tumor Society was 87 percent for the lower extremity and 93 percent for the upper extremity. All patients are presently disease-free. Use of the reinnervated free-muscle transfer in limb-sparing surgery after resection of soft-tissue sarcoma in the extremity may be indicated in the young adult when radical excision of the tumor will result in severe motor functional loss, provided adequate clearance can be obtained and that there is no presence of distant metastasis. PMID- 10541170 TI - Operative management and outcome of complex wounds following total knee arthroplasty. AB - This study describes the treatment protocol for and the outcome of the management of complex wounds around total knee replacements. An analysis of 28 patients (29 knees) with complex defects who had surgery between January 1, 1986, and July 30, 1996, was performed. A specific management protocol was applied to each knee on the basis of the size and depth of the wound, the presence of infection, and the quality of soft tissue. Primary treatment included local wound care, debridement, and skin grafting or coverage with a fasciocutaneous flap, pedicled muscle flap, or free muscle transfer. Postoperatively, knees were evaluated using the Knee Society objective score. Successful salvage of the lower extremity was obtained in 28 knees (97 percent) and of the knee prosthesis in 24 of 29 knees (83 percent). Secondary plastic surgery procedures were necessary in five knees (17 percent), and secondary orthopedic procedures were necessary in four knees (14 percent). Successful salvage of total knee arthroplasty in the presence of a complex wound requires early identification of infection, aggressive irrigation and debridement, and early appropriate soft-tissue coverage. The use of our proposed algorithm will facilitate management of these complex wounds. PMID- 10541172 TI - Reconstruction of fingertip amputations with full-thickness perionychial grafts from the retained part and local flaps. AB - The treatment of fingertip amputations distal to the distal interphalangeal joint when the amputated part is saved is difficult and controversial. Both reattachment of the amputated portion as a composite graft and microvascular anastomosis are prone to failure in this distal location. The authors have evolved a reconstructive plan that uses the nail matrix, perionychium, and hyponychium of the amputated fingertip as a full-thickness graft when the amputation is between the midportion of the nail bed andjust proximal to the eponychial fold. Various flaps are used to lengthen and augment the finger pulp, and skeletal pinning is carried out as necessary. The charts of 15 patients who underwent this procedure over a 38 month period were evaluated retrospectively. Seven returned to the office for examination at least 1 year after the fingertip reconstruction described above; four others were interviewed by telephone. Nail deformity, fingertip sensation, and joint range of motion were evaluated, and the reconstructed fingertips were photographed in standardized views. In six of the seven patients seen in the office, aesthetic and functional results were judged as good by both patient and physician; one of the six had minimal nail curvature. The seventh patient had no nail growth, although finger length was retained and there was no functional disability. The four patients interviewed by phone reported normal fingertip use with no dysesthesias or cold intolerance; all had nail growth, although three patients described slight nail curvature that required care in trimming. The authors favor salvage of all perionychial parts when a distal fingertip amputation occurs. Reconstruction of the fingertip with grafting of the hyponychium, perionychium, and nail matrix from the amputated part combined with local flaps can provide a very satisfactory functional and aesthetic result. PMID- 10541171 TI - Reconstruction of bilateral metacarpal hands with multiple-toe transplantations. AB - Bilateral metacarpal hands, if not treated properly, leave a patient without prehensile ability in both hands. Since 1990, six patients with bilateral metacarpal hands caused by accidents have undergone reconstruction with multiple toe transplantations. Four or five toes were used for each patient, with a total of 27 toes transplanted to the hands. There was no toe loss. One nonunion in a middle-finger reconstruction was treated successfully with bone grafting. Secondary operations for functional improvement included one joint fusion and one flexor tendon tenolysis. Only one patient required excision of a plantar callus 42 months postoperatively, whereas the other five patients reported no major donor-site problems in an average 57 months of follow-up time. The six patients continue all their daily activities independently. Although their jobs were changed, all adult male patients were able to return to regular work. Principles of reconstruction to achieve satisfying prehensile function combined with minor donor-site morbidity in bilateral metacarpal hands include an adequate soft tissue coverage before toe transplantations, selection of digits to be reconstructed based on functional and individual requirements, selection of toes and number of toes to be harvested based on consideration of usefulness for the hands and of foot morbidity, and consideration of thenar function in planning the sequence of transplantations. In conclusion, given thorough planning, multiple toe-to-hand transplantations can provide adequate prehensile function in reconstructed bilateral metacarpal hands with acceptable donor-site morbidity. PMID- 10541173 TI - Mersilene suture as a vehicle for delivery of growth factors in tendon repair. AB - Extensive clinical and laboratory studies have demonstrated that growth factors accelerate and modulate the wound-healing process. The purpose of this experiment was to apply the principles of growth factor-enhanced wound healing to an in vitro rat tendon model. A method was developed for covalently binding a biologically active peptide to nonabsorbable braided polyester suture (Mersilene). Sutures were treated with various growth factors, which included epidermal growth factor, platelet-derived growth factor, and keratinocyte growth factor, and bovine serum albumin was the control. Spectrophotometric assessment was used to verify the peptide's activity. The suture was subsequently placed through individual harvested rat flexor tendons, which were arranged in standard tissue culture conditions. Markedly increased cellular proliferation along the suture was appreciated on the tendons treated with epidermal growth factor-bound suture. Platelet-derived growth factor was shown to have a lesser effect, whereas keratinocyte growth factor had no visible effect on cellular proliferation. This preliminary study describes a new technique of binding growth factors to suture. It also demonstrates that the presence of growth factors may help facilitate flexor tendon healing and allow early postoperative rehabilitation to decrease adhesion formation. PMID- 10541174 TI - Effect of early mobilization on healing of nerve repair: histologic observations in a canine model. AB - The effect of early mobilization on the healing of nerve repair was studied in a canine model. Median and ulnar nerves in the left wrist of 16 adult mongrel dogs were transected and immediately repaired. No motion of the repaired forelimb was allowed in the immobilized group (n = 10), while controlled passive motion between 30 and 90 degrees of wrist flexion was begun on the first postoperative day for 10 minutes twice daily in the mobilized group (n = 6). The pattern of revascularization and collagen formation at neurorrhaphy was examined by transillumination of India ink-injected specimen and by conventional histologic sections. Revascularization of nerve repair was found to occur by ingrowth of capillaries from proximal and distal nerve ends, which typically crossed the neurorrhaphy by 3 weeks in the immobilized group. Following early mobilization, there was a persistent "hypovascular zone" at the nerve repair site for up to 6 weeks. In addition, more scar tissue was generated by early motion according to gross observation and quantitative collagen analysis. Early mobilization, therefore, seems to impede nerve regeneration by delaying revascularization and enhancing scar formation. PMID- 10541175 TI - Sutureless microvascular anastomoses by a biodegradable laser-activated solid protein solder. AB - A new sutureless technique to successfully anastomose the abdominal aorta of rats (1.3 mm in diameter) by using a fully biodegradable, laser-activated protein solder is presented. A total of 90 rats were divided into two groups randomly. In group one, the anastomoses were performed by using conventional microsuturing technique, whereas in group two, the anastomoses were performed by using a new laser welding technique. In addition, each of the two groups were divided into five subgroups and evaluated at different follow-up periods (10 minutes, 1 hour, 1 day, 1 week, and 6 weeks). At these intervals, the anastomoses were evaluated for patency and tensile strength. Three anastomoses in each subgroup were processed for light and electron microscopy. All anastomoses were found to be patent. The mean clamp time of the anastomoses performed with conventional suturing was 20.6 minutes compared with 7.2 minutes for the laser-activated welded anastomoses (p < 0.001). The strain measurements showed a stronger mechanical bond of the sutured anastomoses in the initial phase. However, at 6 weeks the tensile strength of the laser-welded anastomoses was higher compared with the conventional suture technique. Histologic evaluations revealed a near complete resorption of the solder after 6 weeks. The junction site of the vessel ends cannot be determined on the luminal side of the artery. In conclusion, a resorbable protein used as a solder, activated by a diode laser, can provide a reliable, safe, and rapid arterial anastomosis, which could be performed by any microsurgeon faster than conventional suturing after a short learning curve. PMID- 10541176 TI - Development of a human adipocyte synthetic polymer scaffold. AB - Because of the need for methods for successful transplantation of autologous fat, the differentiation of human preadipocytes on surgical mesh coated with various extracellular matrix components was investigated. Biopsy specimens of human adipose tissue were collected from seven different patients and were subjected to collagenase digestion and selective filtration, resulting in primary cultures of human preadipocytes. Fluortex monofilament-expanded polytetrafluoroethylene (52 /xm pore size) was as a template for coating with either human collagen, albumin, or fibronectin, followed by sodding with established cultures of human preadipocytes. Sodding efficiency on the different matrices was determined by trypsinization of attached cells at different time periods. Preadipocytes did not attach to uncoated polytetrafluoroethylene, but did attach to protein-coated mesh, and in a variable manner. Fibronectin-coating resulted in the highest efficiency of sodding, with differentiation of preadipocytes to adipocytes as assessed by scanning electron -microscopy and conventional Oil Red O staining. Similar results were achieved by using rat (n = 6) perirenal adipose tissue. This new method of adipocyte scaffolding may be used for improving soft-tissue augmentation and serving as a delivery system for growth factors important in wound healing. PMID- 10541177 TI - Independent function in a tendon transfer of the split flexor carpi ulnaris. AB - This case demonstrates the efficacy of the split flexor carpi ulnaris transfer to restore thumb and finger extension. The humeral and ulnar compartments retained their viability and their function. In addition, they were able to work independently, and the patient has secured two separate functions from one muscle. The versatility and simplicity of this technique give it advantages over more complex reconstructive procedures. PMID- 10541178 TI - Straightening of aberrant carotid arteries with age in velocardiofacial syndrome. AB - Two cases of velocardiofacial syndrome demonstrated spontaneous straightening over a 4-year period of previously deviated carotid arteries on contrast enhanced computed tomography. Based on this finding, uncomplicated traditional pharyngeal flaps were performed on both patients. It is unknown whether straightening of aberrant vessels with age is the rule or the exception in this patient population. PMID- 10541179 TI - Sensitive areolar reconstruction in using a neurocutaneous island flap based on the medial antebrachial cutaneous nerve. AB - Sensory reconstruction has recently been stressed in breast reconstruction. However, there are no reports concerning the reconstruction of a sensitive areola. The bilateral reconstruction of a sensitive areola using a neurocutaneous flap based on the medial antebrachial cutaneous nerve is reported. The flap was harvested from the distal third of the forearm as an island flap and tunneled to reach the apex of the new breast, which was previously reconstructed using a 135 cc, gel-filled, silicone prosthesis covered by a latissimus dorsi myocutaneous flap. Six months later, fine sensibility in the reconstructed areola was demonstrated. The patient could perceive light touch, pain, and 14 mm two-point discrimination. At 2 months after surgery, 50 percent of cutaneous faulty stimulus location was observed. However, at 4 and 6 months after surgery, faulty location disappeared. Six months after harvesting the medial antebrachial cutaneous nerve, the sensory deficit was minimal; it included a hypoesthesic zone of 4 to 7 cm and an anesthesic zone of 2.5 to 5 cm on the middle third of the forearm. Fifteen months after the procedure, no hypoesthesic zone was observed; only a 2 to 3 cm anesthesic zone on the proximal medial side of the forearm existed. This sensory deficit passed unnoticed by the patient. The technique developed here is a refinement in breast reconstruction, and we think it should be used in selected patients. PMID- 10541180 TI - Hypotelorism: an extracranial correction technique. PMID- 10541181 TI - A missing portrait of Gaspare Tagliacozzi found 400 years after his death. PMID- 10541182 TI - Alloplastic implantation. AB - LEARNING OBJECTIVES: After studying this article, the participant should be able to: 1. Define an alloplastic material and know the differences between an alloplast and other types of implants available for surgical use. 2. Determine the biologic response to alloplastic implantation and the material and host characteristics that contribute to long-term reconstruction success with their use. 3. Review the criteria for choosing a specific alloplastic material for a reconstruction site and the principles of surgical technique for its proper placement. 4. Evaluate the various alloplastic material types that are currently available for surgical use and be able to discuss several physical properties of each as they relate to handling and clinical implantation. 5. Discuss the complication of alloplastic infection, its pathogenesis, preoperative and intraoperative measures for its avoidance, and the postoperative management of its occurrence. PMID- 10541183 TI - Safety of silicone breast implants: scientific validation/vindication at last. PMID- 10541184 TI - Relationship between muscle activity of the frontalis and the associated brow displacement. AB - Although the relationship between regional facial muscle activity and resultant displacement of the face is important to the refinement of many of the treatments of facial paralysis and palsy, this relationship has not been adequately explored. We analyzed the relationship between frontalis muscle activity as measured with surface electromyography and the associated eyebrow displacement as measured with the maximal static response assay in 16 human volunteers. We hypothesized that graded, sustained muscle activity would correlate with graded, sustained elevation of the eyebrow. We found that (1) the relationship between the muscle activity and the corresponding displacements was best described by activity versus displacement squared (r2 = 0.993); (2) there was a wide variation between individual brows for the relationship between muscle activity and displacement; (3) there was a normal amount of asymmetry of the relationship between muscle activity and displacement between the two brows of a subject; (4) left and right frontalis generated the same amount of muscle activity, but left brows elevated significantly higher; (5) the method of simultaneous measurement of muscle activity and displacement used in this study yielded results similar to those previously reported; and (6) the methods used in this study are useful to further investigate the relationship between muscle activity and displacement. PMID- 10541185 TI - A review of 685 rhytidectomies: a new method of analysis based on digitally processed photographs with computer-processed data. AB - It is the aim of this work to present a simple method to assess the surgery used for facial aging (face-neck lift), based on a digital computerized analysis of images, and to include an evaluation of the results obtained from its use on 685 cases of face lift, which our team resolved from January of 1990 to December of 1997. To this end, we reviewed 4110 preoperative and postoperative slides of these patients and processed them using a computer, obtaining the results presented herein. It is worthy of note that in this new method of analysis that we have used, data, measurements, or aspects of facial aging have revealed the most homogeneity and reliability when comparing the results obtained with the reality observed by both the surgeon and the patient. Likewise, we rejected other analyzed data, as they gave different or unreal results when compared with the reality. PMID- 10541186 TI - Suction-assisted lipectomy for lipodystrophy syndromes attributed to HIV-protease inhibitor use. AB - The addition of HIV-protease inhibitors to the arsenal of therapies for the treatment of HIV infection has resulted in significant suppression of viral load such that HIV-positive individuals experience reduced morbidity and extended life expectancy. Recently, a number of syndromes have been described involving abnormal fat distribution that may be associated with prolonged HIV-protease inhibitor therapy. These syndromes include hypertrophy of the cervicodorsal fat pad ("buffalo hump"); a tendency toward increased central adiposity ("protease paunch"); adiposity in the submental, mandibular, and lateral cheek regions of the face; and hypertrophy of adipose tissue in the breast in women. A peripheral lipodystrophy, or fat-wasting, in the extremities and face (particularly the malar and nasolabial fold regions) has also been observed. As these patients live longer and healthier lives, many are beginning to seek surgical correction of the disfigurements. In this regard, we present a review of the literature regarding these recently described syndromes to familiarize plastic and reconstructive surgeons with the unique deformities encountered in this ever-increasing patient population. We also present our results with suction-assisted lipectomy for treatment of these deformities. Physical findings, pathogenesis, and surgical management are discussed. PMID- 10541187 TI - Ketamine-assisted intravenous sedation with midazolam: benefits and potential problems. AB - A review of 134 cases of ketamine-induced intravenous sedation was undertaken. It was concluded that (1) whereas properly titrated midazolam with low-dose ketamine (0.5 mg/kg) can provide almost complete absence of behavioral problems and complete analgesia, transient oxygen desaturation may be seen, and (2) the induction phase of ketamine is an opportunity for the surgeon to rehearse mask ventilation. PMID- 10541188 TI - Elicitation of the oculocardiac reflex during endoscopic forehead lift surgery. AB - Elicitation of the oculocardiac reflex is a well-documented phenomenon encountered during ophthalmologic surgical procedures. Familiarity with and prompt recognition of this entity has significantly reduced the morbidity associated with it; however, potentially lethal arrhythmias and cardiac arrest still occur. We report elicitation of the reflex during manipulation of the supraorbital nerve during endoscopic forehead lift surgery. To our knowledge this is the first case of elicitation of the oculocardiac reflex reported during endoscopic forehead lift surgery. PMID- 10541189 TI - Face lift and hair transplantation as a single procedure. AB - Frequently, when a face lift procedure is performed, several pieces of healthy scalp are discarded as waste unless a prehairline incision is used. In selected cases, such as patients with hair loss, these pieces of scalp may be used to create micrografts (grafts with 1 to 2 hairs) and minigrafts (grafts with 3 to 4 hairs) and transplanted to the areas of need in the same session. I have found particularly rewarding the combination of face lift and hair transplantation, because patients who need both procedures benefit immensely by doing them together. This way, the pieces of healthy scalp that normally would have gone to waste are recycled. In a preliminary fashion, a strip of retroauricular and occipital scalp that normally would be discarded is harvested from one side and handed to my assistants. Under magnification, they dissect it into micrografts and minigrafts as I do the face lift on that side. When I go to the second side of the face lift, I give them the other strip of scalp; again, as they dissect it into grafts, I continue with the face lift. Usually, we generate about 1000 micrografts and minigrafts from those strips that would have normally been discarded. If I want more grafts, I would (in a preliminary fashion) harvest the donor strip of the size required. As the face lift with or without eyelids is completed, we usually have the grafts ready for insertion. Today, we are able to transplant approximately 1000 grafts in about 1 hour. Therefore, combining the two procedures adds only about an hour to our surgical and anesthesia time. PMID- 10541190 TI - Cerclage suture method for closed-tip rhinoplasty. AB - This article presents a method for passing a cerclage suture around the nasal tip complex to narrow it. The method does not require elevation of the skin from the lower lateral cartilages. The method requires a double-pointed straight needle with the suture swaged on in the center. PMID- 10541191 TI - Chin surgery: I. Augmentation--the allures and the alerts. AB - The correction of sagittal deformities of the chin presents a seemingly simple surgical challenge. However, several authors have reported negative sequelae from such chin surgery, During the past 11 years, the senior author (B.M.Z.) has evaluated more than 100 such cases of adverse results after chin augmentation. Many surgeons, it seems, use chin implants unnecessarily and, thus, get into trouble. Because alloplastic chin augmentation is deceptively easy, it tends to be overused in certain situations. Either the surgeon's evaluation is too narrowly focused or his/her abilities to perform other types of surgery (e.g., osseous genioplasty) are limited. Herein, the authors present a diagnostic evaluation protocol, QUAC (Quick Analysis of the Chin), to assist in avoiding simple mistakes in alloplastic chin augmentation. This protocol will alert the surgeon to situations that, if unrecognized, will cause problems and create an unhappy patient. This article will specifically focus on (1) lower lip analysis; (2) the effect of the labiomental fold; (3) chin pad evaluation, both static and dynamic; (4) the anatomy of the cleft chin; (5) special situations; and (6) how to troubleshoot three common problems. The accompanying article, Chin Surgery II, will present a new operation that treats a chin problem that was previously difficult to correct. PMID- 10541192 TI - Chin surgery: II. Submental ostectomy and soft-tissue excision. AB - At the present time, surgical reduction of an isolated large chin is not a simple procedure. Essentially, two surgical procedures exist for chin reduction: osteotomy with setback or prominence reduction by burring. Both of these procedures have potential negative aesthetic sequelae, including mental nerve injuries, bony contour irregularities, increasing submental soft-tissue fullness, and chin pad ptosis. In this report, the authors present a new approach to chin reduction: submental ostectomy with soft-tissue excision. This technique reduces the prominent chin and avoids ptosis by soft-tissue adjustment. PMID- 10541193 TI - The superficial subciliary cheek lift, a technique for rejuvenating the infraorbital region and nasojugal groove: a clinical series of 71 patients. AB - The superficial subciliary cheek lift is an aesthetic operation to rejuvenate the infraorbital region and midface. It smoothes and rejuvenates the nasojugal groove, the nasolabial fold, and cheekpad with a direct pull upward. This technique is a technically straightforward procedure that does not involve dissection at a subperiosteal level, endoscopic equipment, a formal canthotomy, or drill fixation. It does not produce superior displacement of the lateral canthus. The entire operation is performed through an extended subciliary incision. Fixation is accomplished to the intermediate temporal fasciajust lateral to the lateral orbital rim. No reliance is placed on the lower eyelid for tightening the midface region. The superficial subciliary cheek lift can be performed as a separate operation or in conjunction with a face lift. It is an aesthetic procedure that is an adjunct to a subciliary lower blepharoplasty when rejuvenation of the upper midface and nasojugal area is also required. A series of 71 patients is presented, with excellent 6-month to 3-year results and an acceptably low complication rate. PMID- 10541194 TI - Dual-plane lipoplasty for the superficial and deep layers. AB - Two embryologically and histologically distinct layers of the subcutaneous adipose tissue were treated individually by different modalities. The authors performed ultrasound-assisted lipoplasty at the superficial layer and traditional suction-assisted lipectomy in the deep subcutaneous tissue. The ultrasound procedure allowed tissue-specific destruction of the superficial layer supported by dense fibrous networks without disrupting them. Skin retractions caused by this procedure could diminish the necessity of surgical dermolipectomy such as an abdominoplasty in moderately deformed cases. On the other hand, the traditional liposuction was performed to remove the excessive deep fat contained in the loose fibrous network in a limited area. Although the advancement of ultrasound devices and regimens has now reduced their operating times significantly, the ultrasonic lipoplasty was slower than traditional liposuction at the beginning of our procedure. In the abdominal wall, the procedure was performed only in a confined area, because the skin retractions made over the locally managed areas created a circumferential tightness around the trunk, using the nontreated area as a bridge. Confinement on the managed area prevented unnecessary blood loss and tissue destruction. It was less a matter of fat volume to remove in the superficial layer; rather, a precise application to the exact target area was required to get a more natural result. Dual-plane lipoplasty was performed for 2 years in 35 patients, mostly for the abdomen. No serious complication such as seromas or skin loss was observed in our series of patients. PMID- 10541195 TI - Large-volume circumferential liposuction with tumescent technique: a sure and viable procedure. AB - During a 4-year period, 152 female and 9 male patients underwent large-volume liposuction, with ages ranging from 19 to 65 years (mean of 36 years), with a weight previous to surgery between 57 and 126 kg (mean of 72 kg). Tumescent liposuction was done simultaneously by two surgeons in several corporal areas according to the necessities of each case. In 28 patients (17 percent), 500 ml of whole blood was required previous to the surgery by self donation. By means of liposuction, volumes between 5 and 22.3 liters (mean of 8.7 liters) were obtained with an average relation of 860 cc of fat for 140 cc of liquid. The reduction of hemoglobin and hematocrit at 1 week after surgery was of 3.8 g and 12 percent, respectively. The weight after surgery during the patient's follow-up varied from 54 to 111 kg, with an average of 66 kg. Major complications were not presented. Minor complications consisted of two superficial cutaneous necroses (1.2 percent) and 18 seromas (11.2 percent), which were drained without leaving sequelae; 24 patients (14.9 percent) presented postsurgical palpable irregularities, visible in only 8 patients (5 percent); 148 patients (92 percent) expressed important satisfaction with the results of the surgery, with the remaining 13 (8 percent) expressing some disagreement due to persistent irregularities. These complications had a direct relationship to some factors of the surgical technique and some characteristics of the patients. The amount of fat liposuctioned, the ideal height-weight relationship of the patient, the diameter of the cannulas used, and the experience acquired during the time were the most important factors that were associated with the complications. Based on these results, we concluded that large-volume circumferential liposuction with tumescent technique is a viable and sure alternative to achieve improvement of the body contour and weight loss. PMID- 10541196 TI - Lidocaine is not necessary in liposuction. AB - Lidocaine is an integral part of most wetting solutions used in liposuction. Although the Physician's Desk Reference states that the permissible dose of lidocaine is 7 mg/kg, doses as high as 75 mg/kg have been used in liposuction. Lidocaine is used in the wetting solution even when the procedure is performed under epidural or general anesthesia. The justification for this is a reduction in postoperative pain. This study compared the pain between paired, mirrored sides of 10 patients when lidocaine was used on only one side. There was no statistically significant difference between the postoperative pain at 5, 30, 60, and 120 minutes and on the first postoperative day. Because there was no difference in pain whether or not lidocaine was used, and because lidocaine is potentially toxic and lethal, this study concludes that lidocaine is not necessary in liposuction. PMID- 10541197 TI - Factor XI deficiency: implications for management of patients undergoing aesthetic surgery. AB - We report our experience in patients with an abnormal partial thromboplastin time elevation due to factor XI deficiency (Rosenthal syndrome) who presented for aesthetic surgery consideration. Preoperative evaluation included a thorough history, physical examination, coagulation profile, and hematological consultation. Nine of 10 patients underwent 12 elective aesthetic procedures without undue intraoperative or postoperative bleeding. Based on these findings, we stratified patients as low risk or high risk. Low-risk patients were those with greater than 15 percent factor XI levels, or those with 5 to 14 percent factor XI levels but a history of multiple major surgical procedures without bleeding complications. High-risk patients were those with factor XI levels less than 15 percent, history of bleeding either spontaneously or with surgery, and a family history of bleeding diathesis from factor XI deficiency. Low-risk patients had fresh frozen plasma available for the procedure, whereas high-risk patients received fresh frozen plasma 2 hours before surgery. We conclude that (1) in these patients with abnormally high partial thromboplastin time values and no prior known bleeding disorder, we have identified factor XI deficiency as the prevalent coagulopathy; (2) partial thromboplastin time does not necessarily correlate with factor XI levels; (3) patients can be classified as high or low risk for elective surgery based on factor XI levels and prior surgical or family history; (4) recommendations for perioperative management can be made based on this risk profile; and (5) aesthetic surgery can be performed successfully and safely on patients with factor XI deficiency on a case-by-case basis when appropriate guidelines are enforced. PMID- 10541198 TI - Reduction mammaplasty with a circular folded pedicle technique. PMID- 10541199 TI - Deep vein thrombosis prophylaxis. American Society of Plastic and Reconstructive Surgeons. PMID- 10541200 TI - A modification of the Kernahan "Y" classification in cleft lip and palate deformities. PMID- 10541201 TI - Correction of the crooked nose. PMID- 10541202 TI - Repair of nasal and septal perforations. PMID- 10541203 TI - Acquired clefting as a consequence of palate infection. PMID- 10541204 TI - Is the tumescent solution bacteriocidal? PMID- 10541205 TI - Antibacterial effects of tumescent lidocaine. PMID- 10541207 TI - Solving the problem of color mismatch in nipple-areola reconstruction. PMID- 10541206 TI - Pectoralis major myocutaneous flap. PMID- 10541208 TI - Endoscopic versus open carpal tunnel release: is it a toss-up? PMID- 10541209 TI - Treatment for plantar ulcers in Hansen's disease. PMID- 10541210 TI - Unusual complications of circumcision. PMID- 10541211 TI - A simple apparatus for fluid evacuation: syringe suction drain. PMID- 10541212 TI - Do the oil-based breast implants bleed a lot? PMID- 10541213 TI - Pricing strategy for aesthetic surgery: economic analysis of a resident clinic's change in fees. PMID- 10541214 TI - Continuous renal replacement therapy compared with intermittent hemodialysis in intensive care: which is better? PMID- 10541215 TI - Connective tissue growth factor: a potential stimulus for glomerulosclerosis and tubulointerstitial fibrosis in progressive renal disease. PMID- 10541216 TI - Endothelin antagonists for hypertension and renal disease. AB - The endothelin system has been implicated in the pathogenesis of arterial hypertension and renal disorders. Endothelin-1, the predominant isoform of the endothelin peptide family, regulates vasoconstriction and cell proliferation in tissues both within and outside the cardiovascular system through activation of Gi-protein-coupled ET(A) and ET(B) receptors. Endothelin synthesis is regulated through autocrine mechanisms by endothelin converting enzymes, chymases, and non endothelin converting enzyme metalloproteases. In-vitro experiments have demonstrated that endothelin-1 stimulates growth in vascular smooth muscle and in the kidney. Recent studies indicate that endothelin mRNA and protein are also increased in vivo in the kidney and vasculature in hypertension and renal disease. Studies using molecular or pharmacological inhibition of the endothelin system demonstrate that endothelin-1 contributes to the functional and structural changes associated with arterial hypertension and glomerulosclerosis, and that these effects are only in part dependent on blood pressure. These experimental studies and first clinical trials suggest that endothelin antagonists may offer therapeutic potential to reduce end-organ damage in diseases associated with vascular remodeling and renal injury. PMID- 10541217 TI - Targeting the complement system. AB - Interest has blossomed in the development of complement inhibitors, in parallel with a growth in our understanding of the biology of the complement cascade. The first generation of designed inhibitors was based on naturally occurring complement receptors and regulatory molecules. These agents provided useful tools for exploring the role of complement in experimental models of disease, but may have limited therapeutic application in humans because of their short half-lives, limited bioavailability and possible antigenicity. More recently, humanized antibodies and synthetic molecules that block the activation of complement have been developed, which look as though they may overcome some of these difficulties. The possibility for precision inhibition of a limited part of the complement cascade, or for inhibition confined to a single organ, may offer effective therapeutic results, while avoiding the disadvantages of nonselective complement blockade. This review examines the recent evidence that complement inhibition will reduce tissue damage resulting from organ transplantation, ischaemia-reperfusion injury, cancer, glomerulonephritis and the use of extracorporeal circuits. PMID- 10541218 TI - Non-transplant uses of mycophenolate mofetil. AB - The immunosuppressive drug mycophenolate mofetil is a potential new treatment for autoimmune renal disease. In addition to its effects in modulating the autoimmune response, mycophenolate mofetil reduces adhesion molecule expression and delays the progression of renal failure. Data from experimental models of glomerulonephritis, especially of lupus nephritis, have demonstrated that mycophenolate mofetil reverses both inflammatory and autoimmune aspects of disease and influences outcome. As yet, clinical experience with mycophenolate mofetil remains anecdotal but provides a strong platform for the scientific evaluation of mycophenolate mofetil as a new therapeutic agent for these conditions in the future. PMID- 10541219 TI - Pharmacologic prevention of vascular access stenosis. AB - Vascular access dysfunction continues to result in substantial morbidity for chronic hemodialysis patients. Pharmacologic and molecular biologic approaches to prevention of vascular access dysfunction, if clinically successful, will be cost effective and improve quality of life for chronic dialysis patients. This review summarizes currently available information and future prospects in pharmacologic and molecular biologic approaches to preventing vascular access stenosis. PMID- 10541220 TI - Erythropoietin therapy in chronic uremia: the impact of normalization of hematocrit. AB - The target hematocrit to be achieved when treating anemia in hemodialysis patients with erythropoietin is controversial. Current evidence-based recommendations suggest a target hematocrit range of 33% to 36%. Small studies suggest that normalization of hematocrit with erythropoietin may benefit hemodialysis patients in terms of brain function, physical performance, quality of life, and prevention of progressive left ventricular dilatation. However a recent study of the effects of erythropoietin-induced normalization of hematocrit in hemodialysis patients with symptomatic heart disease has shown an increase in both mortality and the rate of vascular access thrombosis. Currently, normalization of hematocrit in patients with symptomatic heart disease is not recommended, nor is it possible to conclude that possible benefits of normalization of hematocrit will outweigh risks in hemodialysis patients without symptomatic heart disease. PMID- 10541221 TI - Immortalized kidney epithelial cells as tools for hormonally regulated ion transport studies. AB - The development of transgenic mice carrying the simian virus-40 large T antigen gene or the temperature-sensitive simian virus-40 large T antigen gene, either alone or placed under the control of the 5'-regulatory regions of tissue-specific or ubiquitous genes, has permitted the production of differentiated, polarized kidney epithelial cells. This review covers the immortalized cell lines issued from the various parts of the renal tubule and, in particular, the recently established collecting duct cell lines that have been used as ex-vivo cell models to analyze the regulation of ion transport processes by hormones. PMID- 10541222 TI - Calcium transport in the kidney. AB - Recent discoveries of calcium-regulating and calcium-transporting proteins have paved the way for a heightened understanding of the mechanisms and control of renal calcium transport. In this review, new findings regarding the multifunctional megalin receptor, chloride channels, a putative calcium entry channel, and the calcium-sensing receptor are discussed. PMID- 10541223 TI - H+,K+-ATPase. AB - The H+,K+-ATPases comprise a group of integral membrane proteins that belong to the X+,K+-ATPase subfamily of P-type cation-transporting ATPases. Although these H+,K+-ATPase isoforms share approximately 60-70% amino acid identity, they exhibit discrete kinetic and pharmacological properties when expressed in heterologous systems. HK alpha2 has been categorized by its insensitivity to Sch 28080, an inhibitor of the gastric H+,K+-ATPase, and partial sensitivity to ouabain, an inhibitor of the Na+,K+-ATPase. This functional profile contrasts with the pharmacological sensitivities ascribed to HK alpha2 in transport studies in rat isolated medullary collecting ducts perfused in vitro and in mouse medullary collecting duct cell lines. HK alpha2 mRNA and protein abundance appears to be both tissue and site-specifically upregulated in response to chronic hypokalemia. This regulatory response has been localized to the outer and inner medulla. To reconcile these expressed sensitivities to those reported in vitro in isolated tubules and cells in culture, it would be necessary to invoke modification of the pharmacologic insensitivity of the colonic H+,K+-ATPase to Sch-28080. Although a 'unique' beta-subunit has been reported recently, this beta subunit (beta(c)) is identical at the amino acid level to the recently cloned beta3-Na+,K+-ATPase. Moreover, while HK alpha2 can assemble indiscriminately with any X+,K+-ATPase beta-subunit, HK alpha2 has been reported to assemble stably with beta1-Na+,K+-ATPase in the renal medulla and in the distal colon. It remains conceivable that subunit assembly could be tissue specific and might respond to different physiological and pathophysiological stimuli. Furthermore, recent studies have suggested that the H+,K+-ATPase is both Na+-dependent and localized to the apical membrane in the distal colon. Therefore, future studies will need to resolve these discrepancies by determining if a unique, yet undiscovered H+,K+ ATPase isoform exists in kidney, or if post-translational modifications of the alpha- and/or beta-subunits could account for these functional diversities. PMID- 10541225 TI - Bibliography. Current world literature. Pharmacology and therapeutics. PMID- 10541224 TI - Assembly of signaling complexes by the sodium-hydrogen exchanger regulatory factor family of PDZ-containing proteins. AB - The sodium-hydrogen exchanger regulatory factor (NHERF) was first identified as an essential cofactor for cyclic AMP-mediated inhibition of the epithelial isoform of rabbit kidney sodium-hydrogen exchanger (NHE3). More recent work shows that NHERF constitutes a family of PSD-95/DIg/ZO-1 (PDZ) domain-containing adapter proteins, only some of which associate with the NHE3 antiporter. Other targets of the NHERF proteins include members of the ezrin-radixin-moesin family of cytoskeletal proteins. In the current model for NHE3 regulation, NHERF links NHE3 to the protein kinase A-anchoring protein, ezrin, and thereby facilitates its phosphorylation and inhibition by protein kinase A. Recent studies have also established the interaction of NHERF and its homologs with the beta2-adrenergic receptor and the platelet-derived growth factor receptor tyrosine kinase that facilitates signal transduction by these receptors. Association with NHERF may also regulate the cystic fibrosis transmembrane conductance regulator and the sodium-bicarbonate transporter. With the rapid increase in the intracellular targets identified for NHERF, the emerging data point to a broad role for these PDZ-containing proteins in the organization of signaling complexes and control of cell physiology. PMID- 10541226 TI - Bibliography. Current world literature. Molecular cell biology and physiology of solute transport. PMID- 10541227 TI - Changes in ankle spasticity and strength following selective dorsal rhizotomy and physical therapy for spastic cerebral palsy. AB - OBJECT: In this investigation the authors quantified changes in ankle plantarflexor spasticity and strength following selective dorsal rhizotomy (SDR) and intensive physical therapy in patients with cerebral palsy (CP). METHODS: Twenty-five patients with cerebral palsy (CP group) and 12 able-bodied volunteers (AB controls) were tested with a dynamometer. For the spasticity measure, the dynamometer was used to measure the resistive torque of the plantarflexors during passive ankle dorsiflexion at five different speeds. Data were processed to yield a single value that simultaneously encompassed the three key elements associated with spasticity: velocity, resistance, and stretch. For the strength test, the dynamometer rotated the ankle from full dorsiflexion to full plantarflexion while a maximum concentric contraction of the plantarflexors was performed. Torque angle data were processed to include the work done by the patient or volunteer on the machine. Plantarflexor spasticity values for the CP group were significantly greater than similar values for the AB control group prior to surgery but not significantly different after surgery. Plantarflexor strength values of the CP group were significantly less than those of the AB control group pre- and postsurgery. Postsurgery strength values did not change relative to presurgery values. CONCLUSIONS: The spasticity results of the present investigation agreed with those of previous studies indicating a reduction in spasticity for the CP group. The strength results did not agree with the findings of most previous related literature, which indicated that a decrease in strength should have occurred. The strength results agreed with a previous investigation in which knee flexor strength was objectively examined, indicating that strength did not decrease as a consequence of an SDR. The methods of this investigation could be used to improve SDR patient selection. PMID- 10541228 TI - Treatment of cerebral origin spasticity with continuous intrathecal baclofen delivered via an implantable pump: long-term follow-up review of 18 patients. AB - OBJECT: The goal of this study was to assess the long-term benefits of managing severe spasticity by using continuous infusion of intrathecal baclofen delivered via an implantable pump. METHODS: Eighteen patients with severe spasticity of cerebral origin, who failed to respond adequately to more conservative treatments, have-been treated with continuous infusion of intrathecal baclofen delivered via an implanted pump. Follow-up review of these patients has lasted between 12 months and 9 years. The patients have been assessed using a variety of tools. Seventeen have had a significant reduction in tone and all have benefited by a reduced need for nursing care or increased function or both. CONCLUSION: Long-term continuous infusion of intrathecal baclofen delivered via an implantable pump offers an effective method for dealing with otherwise intractable spasticity. PMID- 10541229 TI - Failure of the competitive N-methyl-D-aspartate antagonist Selfotel (CGS 19755) in the treatment of severe head injury: results of two phase III clinical trials. The Selfotel Investigators. AB - OBJECT: Excessive activity of excitatory amino acids released after head trauma has been demonstrated to contribute to progressive injury in animal models and human studies. Several pharmacological agents that act as antagonists to the glutamate receptor have shown promise in limiting this progression. The efficacy of the N-methyl-D-aspartate receptor antagonist Selfotel (CGS 19755) was evaluated in two parallel studies of severely head injured patients, defined as patients with post resuscitation Glasgow Coma Scale scores of 4 to 8. METHODS: A total of 693 patients were prospectively enrolled in two multicenter double-blind studies. Comparison between the treatment groups showed no significant difference with regard to demographic data, previous incidence of hypotension, and severity of injury. As the study progressed, the Safety and Monitoring Committee became concerned about possible increased deaths and serious brain-related adverse events in the treatment arm of the two head injury trials, as well as deaths in the two stroke trials being monitored concurrently. The Selfotel trials were stopped prematurely because of this concern and because an interim efficacy analysis indicated that the likelihood of demonstrating success with the agent if the studies had been completed was almost nil. CONCLUSIONS: Subsequently, more complete data analysis revealed no statistically significant difference in mortality rates in all cases between the two treatment groups in the head injury trials. In this report the authors examine the studies in detail and discuss the potential application of the data to future trial designs. PMID- 10541230 TI - The impact of raised intracranial pressure on cerebral venous hemodynamics: a prospective venous transcranial Doppler ultrasonography study. AB - OBJECT: The effect of increased intracranial pressure (ICP) on cerebral venous blood flow has been the subject of very few clinical and experimental studies. The authors assessed the usefulness of venous transcranial Doppler (TCD) ultrasonography as a noninvasive monitoring tool for predicting raised ICP. METHODS: Serial venous TCD studies of the basal vein of Rosenthal and the straight sinus (SS) were prospectively performed in 30 control volunteers and 25 patients with raised ICP. Correlations with ICP data were calculated using a multivariate regression model. Venous blood flow velocities (BFVs) in the basal vein of Rosenthal showed, within a certain range, a linear relationship between mean ICP and maximal venous BFV (r = 0.645; p<0.002). Moreover, a linear relationship was found for maximal venous BFVs in the SS and mean ICP (r = 0.928; p<0.0003). CONCLUSIONS: Venous TCD studies may provide an additional noninvasive monitoring tool for raised ICP and give further insights into the cerebral venous hemodynamics present during raised ICP. PMID- 10541232 TI - Computerized tomography angiography in patients with subarachnoid hemorrhage: from aneurysm detection to treatment without conventional angiography. AB - OBJECT: The purpose of this study was to determine prospectively whether and to what extent computerized tomography (CT) angiography can serve as the sole imaging method for a preoperative workup in patients with ruptured intracranial aneurysms. METHODS: During a 1-year period, all patients who presented to the authors' hospital with subarachnoid hemorrhage demonstrated by unenhanced CT scanning or lumbar puncture underwent CT angiography. Two radiologists evaluated the CT angiography source images and maximum intensity projection slabs and arrived at a consensus. They categorized the quality of the CT angiography as adequate or inadequate and classified aneurysms that were detected as definitely or possibly present. The parent artery of anterior communicating artery aneurysms was identified by asymmetrical anterior cerebral artery size and asymmetrical aneurysm location. The parent artery was indicated by the larger A1 segment in cases of asymmetrical A1 size. Only CT angiograms of adequate quality that revealed aneurysms classified as definitely present and with an unequivocal parent artery were presented to the neurosurgeons, who decided whether preoperative digital subtraction (DS) angiography should still be performed. Forty-nine of the 100 studied patients did not undergo surgery because of poor clinical condition, nonaneurysmal cause of the hemorrhage, or endovascular treatment of the ruptured aneurysm. Of the 51 patients who underwent surgery, radiologists required DS angiography in 17 patients; the imaging technique provided greater certainty in 13 instances. The neurosurgeons required DS angiography 11 times; this provided additional information in two instances. Twenty-three (45%) of the 51 patients were surgically treated successfully on the basis of CT angiography findings alone. CONCLUSIONS: Computerized tomography angiography can replace DS angiography as the preoperative neuroimaging technique in a substantial proportion of patients with ruptured intracranial aneurysms. PMID- 10541231 TI - Increased incidence and impact of nonconvulsive and convulsive seizures after traumatic brain injury as detected by continuous electroencephalographic monitoring. AB - OBJECT: The early pathophysiological features of traumatic brain injury observed in the intensive care unit (ICU) have been described in terms of altered cerebral blood flow, altered brain metabolism, and neurochemical excitotoxicity. Seizures occur in animal models of brain injury and in human brain injury. Previous studies of posttraumatic seizures in humans have been based principally on clinical observations without a systematic approach to electroencephalographic (EEG) recording of seizures. The purpose of this study was to determine prospectively the incidence of convulsive and nonconvulsive seizures by using continuous EEG monitoring in patients in the ICU during the initial 14 days post injury. METHODS: Ninety-four patients with moderate-to-severe brain injuries underwent continuous EEG monitoring begin-ning at admission to the ICU (mean delay 9.6+/-5.4 hours) and extending up to 14 days postinjury. Convulsive and nonconvulsive seizures occurred in 21 (22%) of the 94 patients, with six of them displaying status epilepticus. In more than half of the patients (52%) the seizures were nonconvulsive and were diagnosed on the basis of EEG studies alone. All six patients with status epilepticus died, compared with a mortality rate of 24% (18 of 73) in the nonseizure group (p<0.001). The patients with status epilepticus had a shorter mean length of stay (9.14+/-5.9 days compared with 14+/ 9 days [t-test, p<0.031). Seizures occurred despite initiation of prophylactic phenytoin on admission to the emergency room, with maintenance at mean levels of 16.6+/-2.8 mg/dl. No differences in key prognostic factors (such as the Glasgow Coma Scale score, early hypoxemia, early hypotension, or 1-month Glasgow Outcome Scale score) were found between the patients with seizures and those without. CONCLUSIONS: Seizures occur in more than one in five patients during the 1st week after moderate-to-severe brain injury and may play a role in the pathobiological conditions associated with brain injury. PMID- 10541233 TI - Pericollicular approaches to the rhomboid fossa. Part II. Neurophysiological basis. AB - OBJECT: The authors describe their technique of electrophysiological mapping to assist pericollicular approaches into the rhomboid fossa. METHODS: Surgical approaches to the rhomboid fossa can be optimized by direct electrical stimulation of superficially located nuclei and fibers. Electrophysiological mapping allows identification of facial nerve fibers, nuclei of the abducent and hypoglossal nerves, motor nucleus of the trigeminal nerve, and the ambiguous nucleus. Stimulation at the surface of the rhomboid fossa performed using the threshold technique allows localization above the area that is located closest to the surface. Simultaneous bilateral electromyographic (EMG) recordings from cranial motor nerves obtained during stimulation document the selectivity of evoked EMG responses. With respect to stimulation parameters and based on morphometric measurements, the site of stimulation can be assumed to be the postsynaptic fibers at the axonal cone. Strict limitation to 10 Hz with a maximum stimulation intensity not exceeding 2 mA can be considered safe. Direct side effects of electrical stimulation were not observed. CONCLUSIONS: Electrical stimulation based on morphometric data obtained on superficial brainstem anatomy defines two safe paramedian supra- and infracollicular approaches to the rhomboid fossa and is particularly helpful in treating intrinsic brainstem lesions that displace normal anatomical structures. PMID- 10541234 TI - Endoscopy of the posterior fossa and dissection of acoustic neuroma. AB - OBJECT: The authors evaluated the importance of endoscopes in eliminating the disadvantages of the posterior fossa approach, such as the lack of adequate visualization of the lateral aspect of the internal acoustic canal (IAC). METHODS: Between 1989 and 1998, 32 patients underwent removal of acoustic neuroma (AN) via a combined retro-sigmoid-retrolabyrinthine approach. Endoscopes were used at different stages of the operation, and their use was evaluated with regard to elimination of the disadvantages of the posterior fossa approach. All patients in whom AN had been diagnosed underwent surgery in which a standard retrosigmoid-retrolabyrinthine approach was used. Standard sinus endoscopes of 0 degree, 30 degrees, and 70 degrees were introduced into the cerebellopontine angle before debulking the tumor, and the IAC was inspected at the end of the operation. Neurovascular integrity as well as the relationship between the AN and surrounding structures were evaluated. The IAC was inspected for residual tumor, and if any was found, endoscopically guided tumor dissection was performed. CONCLUSIONS: Endoscopes have facilitated an understanding of the anatomy between an AN and neighboring neurovascular structures. For surgery in which the posterior fossa approach is used, endoscopes can make operations safer by eliminating the disadvantages of the approach. In addition to allowing inspection of the fundus, it is possible to perform endoscopically guided tumor dissection within the IAC. PMID- 10541235 TI - Expression of angiogenic growth factors in dural arteriovenous fistula. AB - OBJECT: Although various mechanisms of the development of dural arteriovenous fistula (AVF) have been described, the exact course of its pathogenesis, including molecular processes mediating its genesis, is still unknown. Recently, the importance of sinus thrombosis and venous hypertension has been reported in experimental and clinical studies. Additionally, a role of angiogenic growth factors in the pathogenesis of vascular malformations of the central nervous system has been reported. In this study, the authors investigated the existence of sinus thrombosis in dural AVF and the expression of angiogenic growth factors (basic fibroblast growth factor [bFGF] and vascular endothelial growth factor [VEGF]) in nine patients with dural AVFs that were surgically resected. METHODS: The authors examined histological features of dural AVFs that involved the transverse/sigmoid sinus in seven patients and the superior sagittal sinus in two. Sinus thrombosis was verified angiographically in seven cases and histologically in all cases. In surgically resected specimens the angiogenic growth factors bFGF and VEGF were examined immunohistochemically in nine patients with dural AVFs, with five dural sinuses from cadavers with unrelated central nervous system diseases serving as a normal control group. The media and perivascular connective tissues of the arteries in the wall of the normal dural sinuses stained faintly for bFGF; on the other hand, the expression of VEGF was not detected. In all patients with dural AVFs, the thick wall of the dural sinus stained strongly for bFGF, mainly in the subendothelial layer and media of the strongly proliferative vessels in the sinus wall, in addition to the perivascular connective tissues. In all nine cases VEGF was expressed in the endothelium of the sinus and perivascular connective tissues. In two cases, VEGF was expressed in many capillaries proliferating in the granulation-like tissues in sinuses that were obliterated by organized thrombi. CONCLUSIONS: It is concluded that the pathogenesis of dural AVF is still unknown, but that angiogenic growth factors, which might be produced by the healing process due to sinus thrombosis, may participate in the genesis of dural AVF. Understanding the mechanism of molecular pathogenesis in the development of dural AVF might aid in the establishment of a new therapeutic strategy for this dynamic vascular disease. PMID- 10541236 TI - Localization of language-specific cortex by using magnetic source imaging and electrical stimulation mapping. AB - OBJECT: In this paper the authors demonstrate the concordance between magnetic source (MS) imaging and direct cortical stimulation for mapping receptive language cortex. METHODS: In 13 consecutive surgical patients, cortex specialized for receptive language functions was identified noninvasively by obtaining activation maps aided by MS imaging in the context of visual and auditory word recognition tasks. Surgery was then performed for treatment of medically intractable seizure disorder (eight patients), and for resection of tumor (four), or angioma (one). Mapping of language areas with cortical stimulation was performed intraoperatively in 10 patients and extraoperatively in three. Cortical stimulation mapping verified the accuracy of the MS imaging-based localization in all cases. CONCLUSIONS: Information provided by MS imaging can be especially helpful in cases of atypical language representation, including bihemispheric representation, and location of language in areas other than those expected within the dominant hemisphere, such as the anterior portion of the superior temporal gyrus, the posteroinferior portion of the middle temporal gyrus, the basal temporal cortex, and the lateral temporooccipital cortex. PMID- 10541237 TI - Functional brain mapping using positron emission tomography scanning in preoperative neurosurgical planning for pediatric brain tumors. AB - OBJECT: The purpose of this report is to demonstrate the value of functional brain mapping using the positron emission tomography (PET) method for preoperative neurosurgical planning in children with brain tumors. Brain maps were used to characterize the relationship between potentially resectable tumors and functionally eloquent brain areas. METHODS: Five children, ranging in age from 3 to 13 years, with hemispheric brain tumors adjacent to eloquent cortex were studied. Magnetic resonance (MR) imaging was used to identify the brain tumors; PET imaging after injection of [18F] fluorodeoxyglucose (FDG), [11C]L methionine (CMET), or a combination of the two was performed to grade the tumors; and a [15O] H2O uptake study was used to characterize the anatomical relationships of the tumors to functional cortex. The cortical activation maps were obtained during control periods and during behavioral tasks and were used to document motor, visual, and speech and language organizational areas. Wada tests were performed in two patients. Language and speech activation was concordant with the results of Wada testing. CONCLUSIONS: Functional brain mapping using PET scans and coregistered MR images provided the neurosurgeon with precise definitions of structural and functional cortical areas; this altered surgical management in some cases and/or was used to predict outcome. The combination of PET imaging with FDG and/or CMET and measurements of [15O] water uptake was useful in characterizing and grading tumors and instrumental in achieving effective neurosurgical planning. Postoperative results in the five cases suggest that preoperative functional brain mapping has the potential to improve outcome by defining a surgical plan to maximize resection and minimize the risk of neurological sequelae. PMID- 10541238 TI - A mobile high-field magnetic resonance system for neurosurgery. AB - OBJECT: The authors' goal was to place a mobile, 1.5-tesla magnetic resonance (MR) imaging system into a neurosurgical operating room without adversely affecting established neurosurgical management. The system would help to plan accurate surgical corridors, confirm the accomplishment of operative objectives, and detect acute complications such as hemorrhage or ischemia. METHODS: The authors used an actively shielded 1.5-tesla magnet, together with 15 mtesla/m gradients, MR console computers, gradient amplifiers, a titanium, hydraulic controlled operating table, and a radiofrequency coil that can be disassembled. The magnet is moved to and from the surgical field by using overhead crane technology. To date, the system has provided unfettered access in 46 neurosurgical patients. In all patients, high-definition T1- and/or T2-weighted images were rapidly and reproducibly acquired at various stages of the surgical procedures. Eleven patients underwent craniotomy that was optimized after preincision imaging. In four patients who harbored subtotally resected tumor, intraoperative MR imaging aided the surgeon in removing the remaining tumor. Interestingly, the intraoperative administration of gadolinium demonstrated a dynamic expansion of enhancement beyond the preoperative contrast contour in patients with malignant glioma. These zones of new enhancement proved, on examination of biopsy samples, to be tumor. CONCLUSIONS: The authors have demonstrated that high-quality MR images can be obtained in the operating room within reasonable time constraints. Procedures can be conducted without compromising or altering traditional neurosurgical, nursing, or anesthetic techniques. It is feasible that within the next decade intraoperative MR imaging may become the standard of care in neurosurgery. PMID- 10541239 TI - Gliomas in rodent whisker barrel cortex: a new tumor model. AB - OBJECT: Surgical treatment of gliomas is difficult because they are invasive. Invasion of essential cortex often limits or precludes surgical resection. A tumor model was developed in which the rodent whisker barrel cortex was used to examine how gliomas affect cortical function and structure. METHODS: Both DBT (mouse) and C6 (rat) glioma cell lines were grown in culture and labeled with the fluorescent marker Dil in vitro. Labeled tumor cells were then injected into the whisker barrel cortex of adult mice and rats. Neurological assessments were made daily and magnetic resonance (MR) images were obtained. Animals were killed by perfusion 6 to 14 days after injection, and histological sections were prepared and studied. Tumors were found in all 20 rats and 10 mice that had been injected with the C6 and DBT cell lines, respectively. The animal cells had been labeled with Dil in vitro, and all in vivo tumors proved to be Dil positive. The MR images revealed the tumor locations and serial MR images demonstrated tumor growth. Histological evaluation confirmed the location of the tumor and the disruption of barrel cortex architecture. CONCLUSIONS: Both DBT and C6 glioma cell lines can be used to generate malignant glial tumors reproducibly in the whisker barrel cortex. Fluorescent labeling and cytochrome oxidase staining permit visualization of tumor growth patterns, which disrupt the barrel cortex by microscopic invasion and by gross tissue deformation. Magnetic resonance imaging demonstrates the anatomical extension of these tumors in live rodents. Using this model for further studies on the effects of malignant glioma growth on functional cerebral cortex should advance our understanding of the neurological issues and management of patients with these tumors. PMID- 10541241 TI - Expression of vascular permeability factor in craniopharyngioma. AB - OBJECT: The expression of vascular endothelial growth/permeability factor (VEG/PF) has recently been correlated with the presence of tumor-associated cysts in some intracranial tumors. The purpose of this study was to evaluate a possible relationship between the presence of VEG/PF and the formation of cysts in craniopharyngiomas. METHODS: The expression of VEG/PF was studied in histological specimens from a series of 12 craniopharyngiomas. In this series, the tumors were classified as presenting a mainly solid pattern with small macroscopic cysts (four patients) or a mainly cystic pattern (eight patients). The mainly solid tumors containing small macroscopic cysts showed little or no VEG/PF positivity, which was mainly present in tumor cells surrounding cysts. Nevertheless, mainly cystic craniopharyngiomas showed a moderate or high degree of VEG/PF positivity in tumor cells. CONCLUSIONS: These findings indicate that the predominance of a cystic or solid macroscopic appearance of craniopharyngiomas may be influenced by the degree of VEG/PF expression within the tumor cells. PMID- 10541240 TI - Cell cycle arrest and astrocytic differentiation resulting from PTEN expression in glioma cells. AB - OBJECT: Genetic alterations of the PTEN gene (also known as MMAC1 or TEP1) have frequently been identified in high-grade gliomas, indicating that inactivation of PTEN plays a crucial role in human glioma progression. The aim of this study was to assess the biological significance of PTEN inactivation in the development of glioma. METHODS: The authors introduced wild-type PTEN complementary DNA into four human glioma cell lines (T98G, U-251MG, U-87MG, and A172) containing endogenous aberrant PTEN alleles. The number of colonies transfected with the wild-type PTEN was reduced to 15 to 32% of those found after transfection of a control vector, suggesting growth suppression by the exogenous PTEN. To analyze phenotypic alterations produced by PTEN expression, T98G-derived clones with inducible PTEN expression were further established using a tetracycline-regulated inducible gene expression system. Induction of PTEN expression suppressed the in vitro growth of T98G cells with accumulation of G1 phase cells. Furthermore, when cells were cultured in the presence of the extracellular matrix (ECM), PTEN expression caused distinct morphological changes, with multiple and elongated cytoplasmic processes similar to those of normal astrocytes. The level of glial fibrillary acidic protein, an intermediate protein specifically expressed in differentiated astrocytes, was upregulated concomitantly. CONCLUSIONS: These findings strongly indicate that exogenous PTEN expression inhibits the proliferation of glioma cells by inducing G1 arrest and elicits astrocytic differentiation in the presence of the ECM. Inactivation of PTEN would play an important role in the enhancement of unregulated growth of undifferentiated glioma cells. PMID- 10541242 TI - Neuronal damage in gerbils caused by intermittent forebrain ischemia. AB - OBJECT: The purpose of this study was to investigate the possibility of preventing cumulative neuronal damage after repetitive severe ischemia. METHODS: The authors monitored ischemic depolarization in the gerbil hippocampus, which has recently been shown to be a good experimental model of the effects of brief ischemia on the brain, and evaluated neuronal damage in the CA1 subregion 7 days after the ischemic insult. In a single-ischemia paradigm, the results indicate that induction of ischemia-induced neuronal damage depended on the duration of ischemic depolarization. Neuronal damage can be detected in the CA1 subregion after a period of depolarization lasting 210 seconds. Using a double-ischemia paradigm in which the animals were subjected to two periods of ischemia, there was apparently no accumulation of neuronal damage from the first ischemic episode to the second, provided the duration of the first period of ischemic depolarization did not exceed 90 seconds. Neuronal damage accumulated when the duration of the first ischemia episode exceeded 90 seconds, regardless of the duration of the reperfusion interval between the two ischemic insults. Finally, when the ischemic insult was spread over four separate episodes, each lasting 90 seconds (with a reperfusion interval of 5 minutes), neuronal damage was not found when the total depolarization period was less than 420 seconds. CONCLUSIONS: The authors conclude that cumulative neuronal damage may be avoided by adopting an intermittent ischemia approach. The implications of these results for human surgery requiring temporary occlusion of the cerebral arteries are discussed. PMID- 10541243 TI - Induction of heat shock protein 70 in the rat brain following intracisternal infusion of autologous blood: evaluation of acute neuronal damage. AB - OBJECT: Investigation into a potential treatment for the acute period following onset of spontaneous subarachnoid hemorrhage (SAH) is hampered by the lack of a standardized experimental model. For that purpose the authors elaborated on a small-animal model in which computer-controlled intracisternal blood infusion is used and investigated whether this model can reliably reproduce acute neuronal injury after SAH. METHODS: Whole autologous blood (blood-infused group) or isotonic saline (control group) was infused into the cisterna magna or olfactory cistern of rats. The infusions decreased exponentially during a 5-minute period. Throughout the infusion period, intracranial pressure (ICP) was monitored. Neuronal injury was quantified by observing tissue immunoreactivity to a 70-kD heat shock protein (HSP70) and comparing this with the tissue's reaction to hematoxylin and eosin staining. On Days 1, 3, and 5, the CA1, CA3, and dentate gyrus regions of the hippocampus were analyzed, respectively. During saline infusion ICP increased within seconds beyond 80 mm Hg and afterward decreased in accordance with the infusion rate. During the infusion of blood, the same initial pressure peak was found, but the ICP remained increased beyond this pressure level throughout the 5-minute infusion period. The HSP70 immunoreactivity in the saline-infused group was found only on Day 1 in the CA1 region and the dentate gyrus, but not in the CA3. After injection of whole blood, there was HSP70 positive staining in the CA1, CA3, and dentate gyrus regions throughout the observation period. CONCLUSIONS: The controlled cisternal infusion of blood caused neuronal injury that resembled that of previous experimental models that produce SAH by rupture of intracranial vessels with endovascular techniques. Unlike those experiments, the intracisternal infusion technique presented by the authors provides more standardized bleeding with regard to ICP, the volume of subarachnoid blood, and the extent of acute cellular injury. PMID- 10541244 TI - Skull bone regeneration in primates in response to basic fibroblast growth factor. AB - OBJECT: The feasibility of using a biodegradable hydrogel incorporating basic fibroblast growth factor (bFGF) to induce bone regeneration at the site of a skull defect in monkeys was investigated. METHODS: Basic fibroblast growth factor was incorporated into a bioabsorbable hydrogel, which was prepared through glutaraldehyde crosslinking of gelatin. Following treatment of monkey skull defects measuring 6 mm in diameter (six defects/experimental group) with gelatin hydrogel incorporating bFGF, skull bone regeneration was evaluated using soft x ray studies, dual x-ray absorptometry, and histological examinations. The water content of the hydrogels varied according to the glutaraldehyde concentration in the hydrogel preparation. Gelatin hydrogels incorporating 100 microg of bFGF significantly promoted bone regeneration and the skull defect was completely closed 21 weeks after implantation. This is in marked contrast with the effect of the same dose of bFGF in solution form. Bone mineral density (BMD) measured at the sites of skull defect was enhanced by the bFGF-incorporating hydrogels. The BMD enhancement was more prominent at lower water contents of hydrogel. Empty gelatin hydrogels neither induced nor interfered with skull bone regeneration. CONCLUSIONS: The findings of this study indicate that bFGF coupled with bioabsorbable hydrogel is a very promising tool to assist in the regrowth of bone at the site of a skull defect, which clinically has been recognized as almost impossible. PMID- 10541245 TI - Recovery of cutaneous pain sensitivity after end-to-side nerve repair in the rat. AB - OBJECT: The hypothesis that collaterally sprouting axons from an uninjured donor nerve may provide recovery of pain sensitivity in the skin after end-to-side nerve repair was investigated in rats. In addition, the effect of this technique on the donor nerve was examined. METHODS: The distal stump of the transected peroneal nerve was sutured end to side to the intact sural nerve. No epineurial window or perineurial slit was made in the sural nerve at the site of coaptation. Other nerves in the leg were transected and ligated. Eighteen weeks later, the sural nerve was transected at a site distal from the coaptation site. The residual pain sensitivity in the peroneal innervation field in the instep was documented using the skin pinch test in three of 11 animals. The area of sensitivity encompassed 19 to 40% of the maximum nociceptive innervation area of the normal peroneal nerve. The nerve pinch test revealed functional sensory axons in all communicating peroneal nerves, in which 277+/-119 myelinated axons (mean +/- standard deviation) were found by histological investigation. CONCLUSIONS: The authors conclude that at least partial recovery of sensory function due to collateral sprouting of axons after end-to-side nerve repair is possible in principle. However, the presence of functional sensory axons in the peroneal nerve stumps did not guarantee the recovery of skin sensitivity to pain in all animals. No functional or morphological evidence of an untoward effect of collateral sprouting into the end-to-side communicating nerve was detected in the axons of the donor nerve itself. PMID- 10541246 TI - Involution of enhancing intrinsic tectal tumors after endoscopic third ventriculostomy. Report of two cases. AB - For benign intrinsic tectal tumors causing triventricular obstructive hydrocephalus, cerebrospinal fluid diversion followed by neuroimaging is a widely accepted treatment plan. In this report, the authors describe two children with focal enhancing tectal lesions that caused acute, symptomatic hydrocephalus. One child had neurofibromatosis Type 1 (NF1). In both children the hydrocephalus was effectively treated by endoscopic third ventriculostomy. Following this procedure, serial imaging studies revealed not only that the ventriculomegaly had resolved, but also that the enhancing tectal tumors had regressed and disappeared over time. The time to complete involution of these tumors was 18 months for the child with NF1 and 12 months for the other child. To the authors' knowledge, this is the first report of the involution of enhancing tectal tumors after endoscopic third ventriculostomy. The possible mechanisms for this unexpected result are discussed. PMID- 10541247 TI - Sagittal sinus occlusion by intraluminal dural cysts. Report of two cases. AB - Lesions involving the sagittal sinus typically present as masses compressing the sinus externally. The authors describe two cases of lesions entirely within the lumen of the sagittal sinus. In one of the cases, syncope was the presenting symptom and surgical resection of the cyst was performed. An entirely intraluminal cyst, consistent with a dural cyst, was resected, followed by reconstruction of the sinus and resolution of symptoms. Entirely intraluminal lesions of the sagittal sinus have rarely been reported as incidental findings. This represents the first report of symptomatic occlusion of a venous sinus by an intraluminal cyst. PMID- 10541248 TI - Ruptured anterior communicating artery aneurysm encased in a tuberculum sellae meningioma. Case report. AB - This 70-year-old woman suffered a subarachnoid hemorrhage (SAH) from a ruptured anterior communicating artery aneurysm encased in a meningioma in the tuberculum sellae. Although preoperative magnetic resonance imaging disclosed that the aneurysmal complex was completely enclosed in the tumor, angiographic studies did not reveal arterial narrowing. The embedded aneurysm caused diffuse SAH rather than intratumoral hemorrhage. These factors indicated very little adhesion between the tumor and the encased arteries. Surgery was performed on the 20th day post-SAH. Intraoperative findings revealed that the tumor did not adhere to the enclosed vasculature except at the point of rupture of the aneurysm. The authors were able to clip the aneurysm safely after piecemeal removal of the tumor, which was finally extirpated without fear of aneurysm rupture. Careful stepwise procedures were essential to treat the aneurysm and the tumor simultaneously. PMID- 10541249 TI - Fatal secondary increase in serum S-100B protein after severe head injury. Report of three cases. AB - The S-100B protein is a small cytosolic protein that is found in astroglial or Schwann cells. It is highly specific for brain tissue and is increasingly being investigated as a diagnostic tool to assess the neurological damage after head injury, stroke, subarachnoid hemorrhage, and cardiopulmonary bypass. The authors report on three patients with severe head injury with otherwise normal cerebral perfusion pressure, SaO2, PaCO2, and controlled intracranial pressure (ICP), in whom a secondary excessive increase in serum S-100B was observed. In all cases, the S-100B increase was followed by an increase in ICP. All three patients died within 72 hours after the excessive increase in S-100B. These findings indicate that major secondary brain damage may occur at a cellular level without being identified by current neuromonitoring techniques. PMID- 10541250 TI - Iatrogenic pneumocephalus secondary to intravenous catheterization. Case report. AB - The presence of pneumocephalus in a patient without a history of undergoing intracranial or intrathecal procedures is a significant radiographic finding that portends a violation of the dural barrier or the presence of infection. The authors report a case of iatrogenic pneumocephalus that confounded the evaluation of a patient with unrelated neurological disorders, resulting in unnecessary transfer of the patient and utilization of medical resources. A review of 100 sequential computerized tomography scans obtained in patients for any indication in the emergency department revealed a 6% incidence of iatrogenic intravenous pneumocephalus. Computerized tomography scans revealing pneumocephalus had been obtained for altered mental status, focal motor deficit, seizure, and trauma. More careful intravenous catheterization and recognition of the condition on imaging may avoid similar problems. PMID- 10541251 TI - Stereotactic radiofrequency ablation for the treatment of gelastic seizures associated with hypothalamic hamartoma. Case report. AB - The author presents the case of a patient with gelastic seizures associated with a hypothalamic hamartoma, in whom partial resection of the hamartoma followed by temporal lobectomy and orbitofrontal corticectomy failed to reduce the seizures. Subsequent stereotactic radiofrequency ablation of the hamartoma resulted in progressive improvement in the seizure disorder during a 28-month follow-up period. There is support in the literature for the concept that gelastic seizures originate directly from the hamartoma; however, direct surgical approaches to these lesions pose significant risks. It is proposed that the technique of radiofrequency ablation provides a minimally invasive, low-risk approach for the treatment of hypothalamic hamartomas. PMID- 10541252 TI - Posteroventral pallidotomy for midbrain tremor after a pontine hemorrhage. Case report. AB - This 49-year-old man gradually developed a disabling action tremor in the proximal right upper extremity 8 months after suffering a pontine tegmental hemorrhage. The intraoperative microrecording in the nucleus ventralis intermedius (VIM) of the left thalamus revealed tremor-synchronous grouped discharges with a vigorous (2.7 Hz) action tremor predominantly in the shoulder and upper arm. High frequency electrical stimulation in the VIM did not affect the tremor. A posteroventral pallidotomy (PVP) was performed and resulted in the successful alleviation of all tremor activity. Posteroventral pallidotomy is known to alleviate parkinsonian tremors, especially those occurring in the contralateral lower extremity, trunk, and proximal segment of the contralateral upper extremity. The authors consider the pallidoreticular pathway to be an important tremor-mediating pathway for the proximal segment of the upper extremities and believe it can be controlled more effectively by PVP than by VIM thalamotomy, as demonstrated by the PVP-induced resolution of the midbrain tremor observed in this case. PMID- 10541253 TI - Partial myotomy/myectomy of the trapezius muscle with an asleep-awake-asleep anesthetic technique for treatment of cervical dystonia. Technical note. AB - The authors describe a technique for performing partial sectioning and myectomy of the trapezius muscle in patients with severe cervical dystonia that is unresponsive to conservative treatment. Asleep-awake-asleep anesthesia allows intraoperative control of the sectioning procedure to avoid causing postoperative weakness of arm elevation above the horizontal plane. The procedure has been performed successfully in three patients. In all cases the dystonic posture of the shoulder and local pain were improved postoperatively. There were no new deficits. This technique can be used as an adjunct to other peripheral surgical procedures in patients with marked laterocollis and dystonic elevation and ante version of the shoulder. PMID- 10541254 TI - Transciliary subfrontal craniotomy for anterior skull base lesions. Technical note. AB - Microsurgical approaches are the procedures of choice for high-risk patients with lesions requiring surgical treatment. The use of a microscope reduces the extent of the surgical invasion, thus minimizing the handling of healthy tissues. The authors present a surgical approach described for the first time in 1981, which has been used for the past 17 years in more than 260 patients with different tumors and vascular lesions of the anterior cranial fossa. The modification set forth in this article makes better exposure possible, allows more space for instrument handling, and improves cosmetic results. This particular report was based on the treatment of 41 patients who were observed for longer than 3 months. All of the patients were satisfied with the cosmetic result. PMID- 10541255 TI - Proliferation index and prophylactic radiosurgery. PMID- 10541256 TI - Regulatory aspects of oxalate secretion in enteric oxalate elimination. AB - Enteric excretion of oxalate was studied in rats with chronic renal failure (CRF) by measuring the magnitude and direction of oxalate fluxes in vitro across short circuited preparations of distal colon in Ussing chambers. The net absorptive flux of oxalate that is characteristic of colonic tissues removed from control rats was significantly changed to a net secretory flux in CRF rats. Injecting CRF rats with a specific angiotensin II (AngII) receptor (AT1) antagonist, losartan, results in a reversal of the CRF-induced net secretory flux (-13.87+/-0.08 pmol x cm(-2) x h(-1)) to an absorptive flux (+7.32+/-3.68 pmol x cm(-2) x h(-1)) by normalizing the unidirectional fluxes of oxalate. Similarly, oxalate fluxes were normalized across CRF colonic tissues by acute, in vitro application of losartan to the serosal bathing solution. It was also possible to simulate the CRF-induced secretory flux of oxalate (Jsm) in vitro across colonic tissues removed from control rats. Serosal application of AngII at 10(-6), 10(-5), and 10(-4) M resulted in significant increases in the s-->m flux of oxalate (increasing deltaJsm = 4.06+/-1.2, 8.41+/-0.94, and 13.8+/-3.8 pmol x cm(-2) x h(-1), respectively). Taken together, these results suggest that CRF-induced oxalate secretion is at least partly mediated by AngII, which is consistent with previous findings of a twofold upregulation of AT1 receptors in CRF colonic mucosa. PMID- 10541257 TI - Intestinal hyperabsorption of oxalate in calcium oxalate stone formers: application of a new test with [13C2]oxalate. AB - In up to one-third of patients with calcium oxalate stones, a hyperoxaluria can be detected. Hyperoxaluria can result from increased endogenous production, from excessive oxalate content of the food, or from intestinal hyperabsorption. For a causal therapy, it is important to discriminate between metabolic and hyperabsorptive hyperoxaluria. Our new 13C-oxalate test allows this differentiation. Under standardized conditions, 50 mg of disodium salt of [13C2]oxalic acid was applied. From the amount of labeled oxalate excreted in urine as measured by a gas chromatographic-mass spectrometric assay, the intestinal absorption was calculated. Seventy patients with recurrent calcium oxalate urolithiasis who had no signs of inflammatory bowel disease were tested. Their mean intestinal oxalate absorption was 9.2+/-5.1%. This was significantly higher than the mean absorption of 50 healthy volunteers (6.7+/-3.9%). There was no difference in oxalate absorption between male (n = 25) and female volunteers. Oxalate absorption correlated with the oxalate excretion in the 24-h urine (volunteers: r = 0.46, P < 0.01; patients: r = 0.62, P < 0.001). Oxalate hyperabsorption was defined as an absorption exceeding 10%. According to this definition, 34% of the patients had oxalate hyperabsorption; 20% of the volunteers showed a hyperabsorption, too. The 13C-oxalate absorption test allows reliable determination of intestinal oxalate absorption. Because of the use of a stable isotope, this test may be repeated as often as required. It will allow the control of therapeutic regimens and also help to unravel genetic influences in stone formation. PMID- 10541258 TI - Direct correlation between hyperoxaluria/oxalate stone disease and the absence of the gastrointestinal tract-dwelling bacterium Oxalobacter formigenes: possible prevention by gut recolonization or enzyme replacement therapy. AB - Oxalobacter formigenes is a specific oxalate-degrading, anaerobic bacterium inhabiting the gastrointestinal tracts of vertebrates, including humans. This bacterium maintains an important symbiotic relationship with its host by regulating oxalate homeostasis, primarily by preventing enteric absorption. Increased absorption of oxalate can lead to multiple complications associated with hyperoxaluria, especially recurrent calcium oxalate urolithiasis. Detection of O. formigenes in the gastrointestinal tract has attracted attention because the absence of this bacterium appears to be a risk factor for development of hyperoxaluria and/or recurrent calcium oxalate kidney stone disease. In the present study, epidemiologic studies with patients at high risk for calcium oxalate urolithiasis showed a direct correlation between the number of recurrent kidney stone episodes and the lack of O. formigenes colonization. As expected, the lack of O. formigenes revealed a clear association with prophylactic antibiotic therapy. To confirm the importance of O. formigenes in regulating hyperoxaluria, laboratory rats known to be noncolonized were colonized with live bacteria or treated with a preparation of oxalate-degrading enzymes derived from O. formigenes to determine any subsequent increased resistance to high oxalate challenge. Rats receiving either bacteria or enzyme replacement therapy excreted far lower levels of oxalate, did not develop the crystalluria observed with control rats, and resisted the formation of calcium oxalate crystals in their nephrons. These observations, taken together, support the concept that O. formigenes is important in maintaining oxalate homeostasis, that its absence from the gut increases the risk for hyperoxaluria and recurrent kidney stone disease, and that replacement therapy is an efficient procedure to prevent hyperoxaluria and its complications. PMID- 10541259 TI - Importance of oxalate precursors for oxalate metabolism in rats. AB - Three metabolic precursors of oxalate were compared after intravenous administration to rats by measuring the urinary excretion of oxalate and related substances using capillary electrophoresis. Urine specimens were collected hourly from eight male Wistar rats (approximately 200 g) in each group. Glyoxylate (2 mg), glycolate (10 mg), and hydroxypyruvate (100 mg) were almost equally oxalogenic based on urinary oxalate excretion, with 22.0, 6.1, and 0.4% of the respective doses being converted into oxalate, 3, 8.9, and 0.2% into glycolate, and 1, 0.1, and 0.003% into glyoxylate. The mean urinary excretion of oxalate peaked between 1 and 2 h, while that of glycolate peaked at 1 h. The baseline urinary excretion of glycolate and glyoxylate was 0.11 to 0.24 micromol/h and 0.0 to 8.3 nmol/h, respectively, and all three agents caused a significant increase of urinary glycolate excretion for 2 to 3 h. Only glyoxylate administration increased urinary glyoxylate excretion at 1 h. Hydroxypyruvate administration significantly increased urinary hydroxypyruvate, glycerate, and citrate excretion at 1 to 2 h. The increase of urinary citrate excretion remains to be explained. PMID- 10541260 TI - Glycolate metabolism by Hep G2 cells. AB - The pathways of oxalate synthesis in humans are not well defined despite their clinical significance in primary hyperoxaluria and idiopathic calcium oxalate nephrolithiasis. Furthermore, the functional roles, if any, of this synthesis have not been elucidated. This study examines pathways of oxalate synthesis from glycolate in Hep G2 cells, a human hepatoma cell line. Incubation of these cells with glycolate has revealed that a pathway may function to synthesize oxalate from glycolate that does not depend on the oxidation of glycolate to glyoxylate by glycolate oxidase. Labeling cells with 14C-glycolate and chromatographic analyses indicated that detectable amounts of 14C-glyoxylate were not formed. A radioactive peak that coeluted with oxalate on ion exclusion chromatography was the only peak yet identified. A detailed examination of glycolate metabolism in these cells should help clarify the terminal steps associated with oxalate synthesis and aid in our understanding of two-carbon metabolism. PMID- 10541261 TI - Recent developments in our understanding of primary hyperoxaluria type 2. AB - Hydroxypyruvate reductase (HPR) has been partially purified from human liver and can be separated into at least two forms by chromatofocusing; these forms therefore differ in their pI values. Both forms, one with a pI of >7.2 (peak A) and the other with a pI between pH 6.5 and 5.5 (peak B), use NADPH as a cofactor. However, only peak B was able to reduce hydroxypyruvate and glyoxylate, with a Km of 2.3 mM for the latter substrate. Peak A coeluted with lactate dehydrogenase and could represent lactate dehydrogenase (which is known to reduce hydroxypyruvate) alone or a mixture of proteins with HPR activity. The Km for hydroxypyruvate of the enzyme(s) in peak A (8 mM) was 80 times greater than that of peak B (0.1 mM), suggesting that the HPR enzyme contained in peak B may be more important physiologically, where the hydroxypyruvate concentrations are in the micromolar range. The data presented provide a biochemical explanation for the previously observed differences in the tissue distribution of HPR and glyoxylate reductase activities in human subjects and support the claim that diagnoses of primary hyperoxaluria type 2 should be made by measurement of glyoxylate reductase activity in the liver. PMID- 10541262 TI - Aspects of calcium oxalate crystallization: theory, in vitro studies, and in vivo implementation. AB - There are three main approaches to urolithiasis research: theory, basic science, and clinical implementation. Although each approach has yielded meaningful results, there does not appear to be complete synergy between them. This article examines these approaches as they pertain to urinary calcium oxalate crystallization processes. Theoretical calculations were performed to examine the role of oxalate concentration on calcium oxalate supersaturation. The effects of magnesium, citrate, and combinations thereof on calcium oxalate crystallization kinetics were examined in a mixed suspension, mixed product removal crystallizer. Finally, male volunteers were given supplements of calcium alone and binary combinations of calcium, magnesium, and citrate to investigate their effects on the urinary supersaturation of calcium oxalate. Calculations showed that oxalate is 23 times more potent than calcium in its effect on the supersaturation of calcium oxalate. In the in vitro experiments, magnesium and citrate reduced the growth and nucleation kinetics as well as the supersaturation. In combination, these two components were more effective than the individual components in reducing the growth rate and the supersaturation. All of the supplements favorably altered the kinetic and thermodynamic risk factors. Calcium was the most effective in reducing the urinary excretion of oxalate. Articulation of these three approaches is essential for the meaningful investigation and understanding of urolithiasis. PMID- 10541263 TI - Nucleation at surfaces: the importance of interfacial energy. AB - The nucleation and growth of stone-forming minerals on the surfaces of other crystalline phases, cellular material, and immobilized macromolecules must be important in the formation of stones in the urinary tract. The nucleation and growth of calcium oxalate monohydrate (COM) crystals were studied using the constant composition kinetics technique, in solution supersaturated with respect to COM (sigmaCOM = 1.44). The solid phases during the reaction were examined by x ray diffraction, scanning electron microscopy, and diffuse reflectance Fourier transform infrared spectroscopy. Human serum albumin was found to nucleate COM crystals when immobilized on hydroxyapatite (HAP) surfaces. The induction period for nucleation of COM on HAP surfaces preadsorbed with albumin significantly decreased to about 65 min from about 230 min for pure HAP particles. The initial growth rate of COM on pure HAP particles, Rm approximately/= 0.56 X 10(-7) mol/min per m2, was slower than that for HAP surfaces preadsorbed with albumin, 2.14 x 10(-7) mol/min per m2. The surface properties were characterized using contact angle measurements by sessile drop and thin layer wicking. The thermodynamic results suggested that surfaces with high Lewis base parameter values (gamma-) and low interfacial tension with water (gammaSL) are more effective in the nucleation and growth of crystal phases. PMID- 10541264 TI - Precipitation of calcium oxalate monohydrate at phospholipid monolayers. AB - Studies of calcium oxalate monohydrate (COM) precipitation at Langmuir monolayers of the phospholipids dipalmitoylphosphatidylglycerol (DPPG), dipalmitoylphosphatidylcholine (DPPC), and dipalmitoylphosphatidylserine (DPPS) are reported. The precipitation is heterogeneous and selective, with a majority of crystals orienting with the COM (101) face toward the monolayer interface for each phospholipid. The number density of COM crystals depends on the identity of the phospholipid monolayer, decreasing in the order DPPG > DPPS > DPPC. It was also found that COM precipitation increases as the surface pressure of the monolayer is lowered and with the addition of low levels (<0.1%) of cholesterol. The possibility that changes in monolayer fluidity influence COM attachment is discussed. PMID- 10541265 TI - Risk factors for crystallization in the nephron: the role of renal development. AB - Several studies indicate that crystallization, the essential first step for stone formation, starts in the nephron. First a calcium phosphate mineral precipitates in the loop of Henle and this may induce formation of calcium oxalate in the late nephron segments. This study investigated the factors that determine the risk of the first calcium phosphate crystallization step in the loop of Henle. Data from a theoretical model that describes the fluid composition in the different nephron segments are combined with data from nucleation experiments. From this, an assessment was made regarding how changes in plasma and urine composition, tubular functions, and renal anatomy effect the chance of initial crystallization of calcium phosphate in the loop of Henle. The results show that parameters like hypercalciuria and hyperoxaluria do not completely reflect the risk for the initial nucleation step. A combination with data on plasma composition and on tubular function is needed to assess this risk. Renal growth from birth to adulthood and the concomitant increase in renal concentrating capacity are shown to increase the risk for crystallization in the loop of Henle. This coincides with the increasing incidence of calcium oxalate urolithiasis. Treating crystallization and stone formation as a nephron event opens new ways for investigating and understanding the process of urinary stone formation. PMID- 10541266 TI - Some aspects of the intratubular precipitation of calcium salts. PMID- 10541267 TI - Role of sex hormones in experimental calcium oxalate nephrolithiasis. AB - It is unclear why men have a higher incidence of calcium oxalate nephrolithiasis than women. This study examined the role of sex hormones on urinary oxalate excretion and kidney stone formation in an experimental model of urolithiasis. Adult male and female Sprague Dawley rats with different sex hormone modulations were given 0.75% ethylene glycol for 2 wk to induce hyperoxaluria and kidney calcium oxalate crystal deposition. The study groups were: intact male and female rats; castrated male and female rats; intact male or female rats with opposite sex hormone implants; and castrated male and female rats with either testosterone or estradiol implants. Overall, a significant negative correlation between urinary oxalate and plasma estradiol/testosterone ratio was found. None of the estradiol-implanted rats, whether male or female, intact or castrated, developed kidney crystal deposits. The three groups of testosterone-implanted rats had a 43 to 88% rate of kidney calcium oxalate crystal deposition. These results indicate that androgens increase and estrogens decrease urinary oxalate excretion, plasma oxalate concentration, and kidney calcium oxalate crystal deposition. These findings may partly explain why nephrolithiasis is a predominantly male disease. PMID- 10541268 TI - Specific modulatory effect of arachidonic acid on human red blood cell oxalate transport: clinical implications in calcium oxalate nephrolithiasis. AB - Greater arachidonic acid (AA) contents, which were correlated with erythrocyte transmembrane oxalate (Ox) transport, were observed in plasma and erythrocyte membrane phospholipids of patients with idiopathic calcium renal stones, suggesting a link between membrane phospholipid fatty acid composition and cellular Ox transport. To confirm this hypothesis, the effects of exogenous red blood cell incorporation of three different fatty acids (i.e., oleic acid, AA, and eicosapentaenoic acid) on Ox transport and the phosphorylation status of band 3 protein, which has been shown to mediate red blood cell Ox flux, were investigated. Preincubation of erythrocytes with AA induced a dose-dependent increase in the phosphorylation level of band 3 protein and an increase in transmembrane Ox self-exchange. In contrast, inhibitory effects on both parameters were observed after the incorporation of oleic and eicosapentaenoic acids. These data, together with previous observations of dietary effects on erythrocyte Ox transport and urinary Ox excretion, indicate that genetic and/or nutritional changes in membrane phospholipid fatty acid composition play a crucial role in modulating cellular Ox transport in idiopathic calcium Ox nephrolithiasis. PMID- 10541269 TI - Role of urinary bikunin in the inhibition of calcium oxalate crystallization. AB - Several urinary macromolecules are known to modulate calcium oxalate (CaOx) crystallization. One of these is urinary bikunin, the light chain of inter-alpha inhibitor. Bikunin has been demonstrated to be an efficient inhibitor of CaOx crystal growth; however, its inhibitory activity against other events in CaOx crystallization has not been fully investigated. To assess the potential of urinary bikunin as an effective inhibitor, its effects on CaOx crystal nucleation and aggregation were evaluated. Nucleation and aggregation of CaOx crystals were studied by measuring turbidity at 620 nm. In the nucleation assay, crystallization was induced by mixing calcium chloride and sodium oxalate, at final concentrations of 3 and 0.5 mM, respectively. Both solutions were buffered with 0.05 M Tris, 0.15 M NaCl, pH 6.5. Nucleation measurements were performed at 37 degrees C, with stirring at 800 rpm. Inhibition of nucleation was estimated by comparing the induction time in the presence of the inhibitor with control values. In the aggregation assay, the optical density of the solution containing CaOx monohydrate crystals was monitored. Inhibition of aggregation was evaluated by comparing the turbidity slope in the presence of the inhibitor with control values. The data showed that urinary bikunin, at concentrations of 2.5 to 20 microg/ml, retarded crystal nucleation by 67 to 58% and inhibited crystal aggregation by 59 to 80%. According to these results, it seems that urinary bikunin is an efficient inhibitor of crystal nucleation and aggregation. Its presence in the kidneys and urine may protect subjects against CaOx crystallization and kidney stone formation. PMID- 10541271 TI - Analysis of urinary concentrations of calcium phosphate crystal-associated proteins: alpha2-HS-glycoprotein, prothrombin F1, and osteopontin. AB - It has been reported that prothrombin F1 and osteopontin (OPN) have strong inhibitory effects on calcium oxalate crystallization and are produced in stone forming kidneys in animal models. It is important to evaluate urinary concentrations of these proteins for patients with renal stones and healthy control subjects. Urinary macromolecules were collected from nine healthy individuals, nine stone-formers, and five patients with primary hyperparathyroidism (HPT). Each 50-mg aliquot of urinary macromolecules was mixed with calcium phosphate solution, and calcium phosphate crystal-precipitated proteins (alpha2-HS-glycoprotein, prothrombin F1, and OPN) were obtained. The proteins were analyzed by anion-exchange chromatography. Furthermore, OPN levels in whole urine from 18 healthy individuals, 31 stone-formers, and two patients with HPT were measured using a new enzyme immunoassay system. The elution peaks for prothrombin and OPN were significantly smaller for the stone-formers and patients with HPT, compared with the healthy control subjects. Urinary concentrations of OPN assessed using the enzyme-linked immunosorbant assay were significantly lower for stone-formers. Lower urinary excretion of prothrombin F1 and OPN by stone-formers might be one of the reasons for stone formation. PMID- 10541270 TI - Effects of microgravity on urinary osteopontin. AB - Increased risk of renal stone formation during space flight has been linked primarily to increased calcium excretion from bone demineralization induced by space flight. Other factors contributing to increased risk include increased urinary calcium oxalate supersaturation, while urinary citrate, magnesium and volume are all decreased. The aim of this study was to increase the predictive value of stone risk profiles for crew members during space flight by evaluating the excretion of urinary protein inhibitors of calcium crystallization so that more comprehensive stone risk profiles could relate mineral saturation to the concentrations of inhibitor proteins. Levels of urinary osteopontin (uropontin) are reported in a series of 14 astronauts studied before, during, and after space flights. During space flight, a compensatory increase in uropontin excretion was not observed. However, the uropontin excretion of a majority of astronauts was increased during the period after space flight and was maximal at 2 wk after landing. The downward shift in the molecular size of uropontin observed in samples obtained during space flight was shown to result from storage at ambient temperature during flight, rather than an effect of microgravity on uropontin synthesis. PMID- 10541272 TI - Hyaluronans: crystallization-promoting activity and HPLC analysis of urinary excretion. AB - This work follows from our earlier report of a crystallization-promoting population in the leading electrophoretic fractions of pooled urinary glycosaminoglycan extracts of renal stone formers (GAG(SF)), but not in those of healthy individuals (GAG(H)). Preliminary HPLC analysis of disaccharide products of the fractions indicated the presence of chondroitin sulfates (CS) and hyaluronan (HA). Because both commercial CS and urinary CS of healthy individuals showed a basal crystallization-promoting property, we hypothesized that the observed property was due to HA excreted by SF and not by healthy individuals. Defined quantities of a commercial preparation of reference HA (pig skin, 40 to 60 kD) were first tested in urine ultrafiltrate (prepared at pH 5.3, 1250 mosmol/kg and nominal cutoff of 10 kD) by a freezing procedure to assess the crystallization-promoting property. The reference HA moderately promoted crystallization, yielding fewer crystals (5 to 10 times those in neat urine ultrafiltrate) than similar hexuronate concentrations of the HA-containing GAG(SF) population (25 to 30 times those in neat urine ultrafiltrate) reported earlier. Urinary GAG were then recovered individually from random urine samples of SF and healthy individuals. Products of sequential digestion by Streptomyces hyaluronidase and then chondroitinase ABC were analyzed by HPLC. Both GAG(SF) and GAG(H) indicated the presence of HA. Taken together, these results suggest that the crystallization-promoting property of HA(SF) does not generally apply to all HA preparations and that although healthy individuals also excrete HA, HA(H) does not share the property of HA(SF). PMID- 10541273 TI - Prothrombin gene expression in rat kidneys provides an opportunity to examine its role in urinary stone pathogenesis. AB - Urinary form of prothrombin (PT) fragment 1 is the most abundant protein in calcium oxalate crystals generated in human urine. The protein has also been detected in human calcium-containing stones. In its purified form, the protein inhibits calcium oxalate crystal growth and, more importantly, aggregation in aqueous inorganic solutions and undiluted human urine. Recently, PT gene expression has been reported in human kidneys. However, access to human renal tissue for studies is limited, and it is not possible to easily manipulate PT biosynthesis in human subjects. The aim of this investigation, therefore, was to determine whether PT gene expression is present in rat kidneys. Samples of total RNA were isolated from the kidneys and livers (positive controls) of 12 male hooded Wistar rats. Using reverse transcription-PCR, mRNA corresponding to the thrombin and F1+2 regions of PT was analyzed by agarose gel electrophoresis. The expression of the "housekeeping" gene glyceraldehyde-3-phosphate dehydrogenase was also examined, to determine the availability of amplifiable substrate in each specimen. The amplified products were also sequenced, to determine their identities. All rat kidneys displayed evidence of expression of the thrombin and F1+2 domains of the PT gene. This similarity between human and rat kidneys allows the possibility of using established rat models of stone disease to evaluate therapeutic strategies to reduce stone formation. PMID- 10541274 TI - Gene expression of prothrombin in human and rat kidneys: basic and clinical approach. AB - Prothrombin has remarkable affinity for calcium oxalate crystals. It is produced in renal tubular cells and is detected as a urinary form of prothrombin F1. The aim of this basic study was (1) to isolate prothrombin mRNA from normal human and rat kidneys; (2) to confirm expression level changes in stone-forming rat kidneys; and (3) to analyze the DNA sequence of renal prothrombin. The aim of the clinical investigation was to measure the serum levels of renal prothrombin in clinical cases of various urologic diseases. The expression of prothrombin mRNA in human kidneys and male Wistar rat kidneys was investigated using reverse transcription-PCR, with prothrombin (F1, F2, and thrombin) primers. Renal prothrombin levels were measured in the sera of patients with renal cell carcinoma, renal transplant donors, patients with chronic renal failure, and renal transplant recipients, using an enzyme-linked immunosorbent assay. Expression of cyclophilin as well as prothrombin mRNA could be detected. Prothrombin mRNA expression levels seemed to be increased in stone-forming rats. The DNA sequence of renal prothrombin differed from that of liver prothrombin at three points. Repeated measurements of renal prothrombin showed that values were high during the acute tubular necrosis period and tended to decrease with the recovery of renal function. Prothrombin mRNA expression could be confirmed in human and rat kidneys, as well as in stone-forming rat kidneys. Serum concentration measurements can be considered useful for assessment of recovery from acute tubular necrosis after renal transplantation and for diagnosis of acute rejection. PMID- 10541276 TI - Oxalate: from crystal formation to crystal retention. AB - Idiopathic calcium oxalate stone formation is a multifactorial disease. It is therefore unlikely that a single underlying condition will be responsible for entire spectrum of the disease; however, it appears that one important factor in the pathogenesis is an abnormality in oxalate metabolism. Whatever the cause, two critical parameters for stone formation are crystal formation and crystal retention in the renal tubules. Although crystal formination and role of oxalate in crystal formation have been evaluated extensively, it is only recently that crystal retention has been addressed. Previous studies from our laboratories demonstrated that oxalate exposure to renal epithelial cells in culture resulted in initiation of a program of events including DNA synthesis and cell death. The present studies evaluated effects of oxalate on cell proliferation and damage to distal tubular (Madin-Darby canine kidney cells) and proximal (LLC-PK1 cells) cells. Effects of oxalate exposure on calcium oxalate monohydrate (COM) crystal adherence to these cells were also evaluated. Results presented herein demonstrate that proximal tubular cells are more sensitive to oxalate than distal tubular cells. Furthermore, oxalate exposure to proximal tubular cells resulted in reinitiation of DNA synthesis, whereas no such effect was observed in distal tubular cells. Higher levels of oxalate (> 1 mM) resulted in cell loss of both proximal and distal tubular cells, as observed by crystal violet staining. Despite these differences, oxalate exposure to both proximal and distal tubular cells resulted in increased COM crystal adherence. Thus, oxalate exposure may promote crystal adherence to renal epithelial cells either secondarily to cell death and proliferation or by a yet unidentified mechanism. These studies provide the first direct evidence for the role of oxalate in promoting COM crystal retention by the urothelium. PMID- 10541277 TI - Nucleation, adhesion, and internalization of calcium-containing urinary crystals by renal cells. AB - Renal tubular fluid in the distal nephron is supersaturated favoring nucleation of the most common crystals in renal stones, which are composed of calcium oxalate and calcium phosphate. The mechanisms whereby these newly formed crystals can be retained in the nephron and develop into calculi are not known. Calcium oxalate monohydrate and hydroxyapatite (calcium phosphate) crystals rapidly adhere to anionic sites on the surface of cultured renal epithelial cells, but this process is inhibited by specific urinary anions such as citrate, glycosaminoglycans, uropontin, or nephrocalcin, each of which can coat the crystals. Therefore, competition for the crystal surface between soluble anions in tubular fluid and anions anchored on the apical cell surface could determine whether a crystal binds to a tubular cell. Crystals of calcium oxalate dihydrate can also nucleate directly on the surface of cultured BSC-1 cells in a face specific manner, suggesting another potential pathway for crystal deposition in the nephron. Once present on the cell surface, calcium oxalate monohydrate, calcium oxalate dihydrate, and hydroxyapatite crystals are quickly internalized by renal cells; alterations in gene expression and initiation of proliferation may then ensue. Calcium oxalate crystals can also dissolve after renal cells internalize them, but this process may require up to several weeks. Increased knowledge about cell-crystal interactions, including identification of molecules in tubular fluid and on the cell surface that modulate the process, appear critical for understanding the pathogenesis of nephrolithiasis. PMID- 10541275 TI - Molecular detection of heparan sulfate proteoglycan mRNA in rat kidney during calcium oxalate nephrolithiasis. AB - The present study used reverse transcription (RT)-PCR to examine heparan sulfate proteoglycan (HSPG) mRNA expression in rat kidneys. Total mRNA in kidney was isolated and converted to cDNA. To confirm the exact expression level of HSPG mRNA, quantitative competitive (QC)-PCR was performed for each sample. PCR products were resolved by electrophoresis on 1.5% agarose gel and visualized with ethidium bromide. Fragment intensity and area were measured using an image analyzer. In QC-PCR, target DNA and competitive DNA were expressed, using gene specific primer for HSPG mRNA, as 506- and 345-bp bands, respectively. The level of HSPG mRNA expression apparently increased in the nephrolithic rat kidney. Immunohistochemical study revealed that increased production of heparan sulfate was detected in both distal and proximal tubules during nephrolithiasis. These findings suggest that increased expression of HSPG may play a significant role during calcium oxalate stone formation. PMID- 10541278 TI - Attachment sites for particles in the urinary tract. AB - The adherence of crystals to the surface of renal tubule epithelial cells is one of the initial events in the development of nephrolithiasis. The accumulation of crystalline material in the kidney will sooner or later result in the formation of a stone. Calcium crystals occasionally are present in the urine of even healthy individuals, and mechanisms responsible for the selective attachment of crystals to the tubular epithelium of stone-forming individuals must exist. Although several types of cell surface molecules, including phosphatidylserine (PS) and sialic acid, have been proposed as receptors for crystals in the tubular system, the exact nature of these crystal-binding sites has not yet been revealed. Previously, it was demonstrated that calcium oxalate monohydrate crystals adhere to subconfluent, but not to confluent, Madin-Darby canine kidney I cultures. This model was used here to investigate whether the surface of cells with affinity for crystals is enriched with one of the proposed crystal-binding molecules. Annexin V was used for the detection of PS at the cell surface, and Sambucus nigra lectin was used to reveal terminal sialic acid in a (alpha2,6) linkage to galactose units. FITC-annexin V binding studies showed that PS was not exposed at the surface of proliferating or growth-inhibited cells, unless they were pretreated with an apoptosis-inducing cytotoxic agent. Sambucus nigra lectin binding, of which the specificity was confirmed by blocking with N acetylneuraminyl-lactose, demonstrated the abundant presence of (alpha2,6)-linked sialic acid residues at the cell surface of both subconfluent and confluent cultures. While these results seem to rule out a role for PS in the adherence of calcium oxalate monohydrate crystals to the surface of maturating Madin-Darby canine kidney-I cells, they question the role for cell surface-associated sialylated glycoconjugates in this process. PMID- 10541279 TI - Bikunin prevents adhesion of calcium oxalate crystal to renal tubular cells in human urine. AB - Crystal-renal tubular cell interactions are important factors in crystal retention and development of kidney stones. It has been reported that human urine, especially its macromolecular fraction, distinctively prevented calcium oxalate monohydrate (COM) crystal adhesion to tubular cells. This study was designed to find and isolate a specific substance in human urine with a strong inhibitory effect against crystal adhesion. A protein from the urine was purified by two anion exchange chromatography columns and one gel filtration column. The inhibition activity for COM crystal adhesion to Madin-Darby canine kidney cells was determined quantitatively. Amino acid sequence of the protein was analyzed and then subjected to homology search in the GenBank protein database. A specific human urine protein that inhibited the COM crystal adhesion to the cells was isolated and identified. Molecular mass of the protein was approximately 35 kD. The first 20-amino acid sequence from the N-terminal of the purified protein was structurally homologous with the light chain of inter-alpha-trypsin inhibitor, also called bikunin. The isolated bikunin inhibited crystal adhesion at a minimum concentration of 10 ng/ml, and blocked completely at 200 ng/ml. It is concluded that bikunin may contribute to the regulation of crystal adhesion and retention within tubules during kidney stone formation. PMID- 10541280 TI - Oxalate-induced exposure of phosphatidylserine on the surface of renal epithelial cells in culture. AB - A molecular mechanism of crystal attachment to renal cells after injury has been proposed in which the exposure of phosphatidylserine (PS) on the cell membrane surface following injury provides attachment sites for calcium-containing crystals. Annexin V was used to determine whether injury to kidney cells by oxalate in culture resulted in PS exposure on the cell surface. When continuous cultures of intermedullary collecting duct cells were exposed to various levels of oxalate, a dose-dependent increase in PS exposure was observed on the cell surfaces. Initially, only scattered cells expressed PS on the surface. However, as the level of oxalate increased, groups of cells began to express PS, suggesting that the injured cells may have an influence on neighboring cells. Exposure of PS on the cell membrane surface correlated with a corresponding increase in calcium oxalate monohydrate crystal attachment to the cells. This indicates that damage to kidney epithelial cells by elevated concentrations of urinary components, in this case oxalate, could result in exposure of PS on cells, which could provide a point of fixation or nucleation for calcium containing crystals. PMID- 10541281 TI - Oxalate-induced changes in the viability and growth of human renal epithelial cells. AB - Previous studies on the porcine renal epithelial LLC-PK1 cell line demonstrated that oxalate exposure produces concentration-dependent effects on renal cell growth and viability via process(es) involving free radicals. The present studies were conducted to determine whether these findings could be extended to a renal proximal tubular epithelial cell line derived from the human kidney. These studies examined oxalate-induced changes in membrane integrity after short-term exposure (4 h) and changes in cell survival after longer-term exposure (24 to 72 h). Oxalate-induced changes were also assessed in the expression of two genes: egr-1, a zinc-finger transcription factor, and osteopontin, a protein associated with tissue remodeling. The present studies also determined whether oxalate induced changes in either cell viability or gene expression depended on free radicals. Oxalate at a concentration > or = 175 microM (free) produced membrane damage within 4 h. This effect was inhibited by Mn(III) tetrakis (1-methyl-4 pyridyl) porphyrin (MnTMPyP), a superoxide dismutase mimetic, but not by N-acetyl cysteine, a glutathione precursor, or by deferoxamine, an iron chelator. Acute oxalate-induced injury was followed by cell loss within 24 h, an effect maintained at 48 and 72 h at high concentrations of oxalate. Oxalate also promoted DNA synthesis. This mitogenic effect offset cell loss at lower oxalate concentrations (88 microM) leading to a small but significant increase in cell number at 72 h. Treatment with oxalate also increased expression of egr-1 mRNA within 1 h, a response that was attenuated by MnTMPyP; oxalate treatment for 8 h also increased abundance of osteopontin mRNA. These studies suggest that oxalate exposure produces changes in human renal cell growth and viability via a process(es) dependent on reactive oxygen intermediates. Such changes may play a role in the development and/or progression of renal disease via generation of reactive oxygen intermediates. PMID- 10541282 TI - Cells of proximal and distal tubular origin respond differently to challenges of oxalate and calcium oxalate crystals. AB - LLC-PK1 and Madin-Darby canine kidney (MDCK) cells were used to study the role of free radicals in renal epithelial injury during exposure to oxalate ions (Ox) and calcium oxalate monohydrate (COM) crystals. The cell cultures were exposed for 120 or 240 min to 1.0 mmol Ox or 1.0 mmol Ox plus 500 microg/ml of COM crystals averaging 1.0 microm in size. Exposure of both LLC-PK1 and MDCK cells to Ox alone increased the leakage of lactate dehydrogenase, which was further enhanced when cells were exposed to Ox + COM crystals. The release of lactate dehydrogenase from the LLC-PK1 cell line, however, was significantly higher than that from MDCK cells. LLC-PK1 cells also showed a significant increase in malondialdehyde (MDA) content on Ox challenge. MDA content was even higher when LLC-PK1 cells were challenged with Ox + COM crystals. However, in MDCK cells, the elevated MDA content was similar in both treatment groups, suggesting that these cells may be more resistant to the calcium oxalate crystals. Glutathione peroxidase activity was decreased in both LLC-PK1 and MDCK cells. Challenging cells with Ox + COM resulted in decreased catalase activity in LLC-PK1, but increased catalase activity in MDCK cells. Superoxide dismutase activity and reduced glutathione content were not significantly different in either cell type when challenged with Ox or Ox + COM. Previous in vivo animal studies yielded indirect evidence for the increased lipid peroxidation during hyperoxaluria-induced nephrolithiasis. However, in an animal model, it is difficult to separate the effect of Ox from Ox in combination with COM crystals. This study suggests that the injury to renal tubular epithelial cells is accompanied by lipid peroxidation when exposed to Ox. The injury is augmented when COM crystals are included. LLC-PK1 cells are more susceptible to Ox-associated injury than MDCK cells. PMID- 10541283 TI - Crystal-cell interaction and apoptosis in oxalate-associated injury of renal epithelial cells. AB - Two renal epithelial cell lines, LLC-PK1 and Madin-Darby canine kidney (MDCK), were grown in monolayers and exposed to oxalate (Ox) and/or calcium oxalate (CaOx) crystals to investigate cellular responses to these challenges. In addition, LLC-PK1 cells were exposed to high concentrations of Ox for various time periods to investigate the role of apoptosis in Ox-associated cell injury. Both cell types showed signs of damage when exposed to Ox. However, LLC-PK1 cells appeared more sensitive than MDCK cells. There was a significant increase in release of lactate dehydrogenase into the medium and decrease in trypan blue exclusion by cells in the monolayer. Most noticeable was the detachment of cells from the substrate. Exposure of cells to CaOx crystals resulted in their attachment to cell surfaces followed by internalization. Using flow cytometry for quantification of apoptotic cells, transmission electron microscopy for morphology, and electrophoresis for DNA laddering detection, we observed significant apoptotic changes including condensation and margination of nuclear chromatin, DNA fragmentation, and migration of phosphatidylserine of the plasma membrane from inside to the cell surface. However, these cells also showed some necrotic changes such as loss of plasma membrane integrity and release of lactate dehydrogenase, indicating that the apoptotic process was interrupted. PMID- 10541284 TI - NS-398 upregulates constitutive cyclooxygenase-2 expression in the M-1 cortical collecting duct cell line. AB - The cortical collecting duct (CCD) is a major site of intrarenal prostaglandin E2 (PGE2) synthesis. This study examines the expression and regulation of the prostaglandin synthesizing enzymes cyclooxygenase-1 (COX-1) and -2 in the CCD. By indirect immunofluorescence using isoform-specific antibodies, COX-1 and -2 immunoreactivity was localized to all cell types of the murine M-1 CCD cell line. By immunohistochemistry, both COX-1 and COX-2 were localized to intercalated cells of the CCD on paraffin-embedded mouse kidney sections. When COX enzyme activity was measured in the M-1 cells, both indomethacin (COX-1 and -2 inhibitor) and the specific COX-2 inhibitor NS-398 effectively blocked PGE2 synthesis. These results demonstrate that COX-2 is the major contributor to the pool of PGE2 synthesized by the CCD. By Western blot analysis, COX-2 expression was significantly upregulated by incubation with either indomethacin or NS-398. These drugs did not affect COX-1 protein expression. Evaluation of COX-2 mRNA expression by Northern blot analysis after NS-398 treatment demonstrated that the COX-2 protein upregulation occurred independently of any change in COX-2 mRNA expression. These studies have for the first time localized COX-2 to the CCD and provided evidence that the intercalated cells of the CCD express both COX-1 and COX-2. The results also demonstrate that constitutively expressed COX-2 is the major COX isoform contributing to PGE2 synthesis by the M-1 CCD cell line. Inhibition of COX-2 activity in the M-1 cell line results in an upregulation of COX-2 protein expression. PMID- 10541286 TI - Increased nitric oxide synthase-3 expression in kidneys of deoxycorticosterone acetate-salt hypertensive rats. AB - In addition to its hemodynamic effects, nitric oxide (NO) may play a role in the renal tubular handling of sodium. Experiments were conducted to determine possible changes in renal nitric oxide synthase-3 (NOS3) expression in rats treated with deoxycorticosterone acetate (DOCA) and high salt. All rats were uninephrectomized, and either a placebo or DOCA pellet was implanted subcutaneously. Placebo-treated rats were then given tap water to drink ad libitum, and DOCA-treated rats received a 0.9% NaCl solution to drink. Once a week, rats were placed in metabolic cages so that a 24-h urine sample could be collected. After 3 wk, the animals were sacrificed and the kidneys removed and prepared for subsequent immunohistochemical or Western blot analysis. Urinary excretion of nitrate and nitrite (NOx) was measured to provide an indication of the intrarenal production of NO. DOCA-salt hypertensive rats exhibited increased urinary NOx excretion (2.43 +/- 0.48 micromol NOx/mg creatinine) compared with the placebo control animals (1.17 +/- 0.06 micromol NOx/mg creatinine). Western blot analysis revealed that NOS3 protein levels in both the cortex and medulla were greater in DOCA-salt rats compared with placebo-treated animals. Immunohistochemical analysis of kidneys revealed that NOS3 expression in placebo rats was localized in vascular endothelial cells with slight, but detectable, immunoreactivity in medullary collecting ducts. In DOCA-salt rats, a very large increase in the intensity of immunostaining was detected in tubular epithelia of the proximal tubule, thick ascending limb of Henle's loop, and cortical and medullary collecting duct; immunoreactivity in endothelial cells appeared unchanged. These data suggest that increased tubular expression of NOS3 is responsible, at least in part, for the increased renal production of NO in DOCA salt hypertension, and are consistent with a role for NO in the renal tubular response to salt loading. PMID- 10541285 TI - Zonal heterogeneity in action of angiotensin-converting enzyme inhibitor on renal microcirculation: role of intrarenal bradykinin. AB - The present study examined the role of intrarenal bradykinin in angiotensin converting enzyme inhibitor (ACEI)-induced dilation of renal afferent (AFF) and efferent arterioles (EFF) in vivo, and further evaluated whether ACEI-stimulated bradykinin activity differed in superficial (SP) and juxtamedullary nephrons (JM). Arterioles of canine kidneys were visualized with an intravital charge coupled device camera microscope. E4177 (an angiotensin receptor antagonist, 30 microg/kg) dilated AFF and EFF in SP (15 +/- 3% and 19 +/- 5%) and JM (15 +/- 3% and 18 +/- 4%). Subsequently, cilazaprilat (30 microg/kg) caused further dilation of both AFF (29 +/- 4%) and EFF (36 +/- 4%) in JM, whereas in SP it dilated only EFF (29 +/-3%). Similarly, in the presence of E4177, cilazaprilat caused further increases in sodium excretion. This cilazaprilat-induced vasodilation and natriuresis was abolished by a bradykinin antagonist (N(alpha)-adamantaneacetyl-D Arg-[Hyp3,Thi5,8,D-Phe7]b radykinin). In parallel with these results, cilazaprilat increased renal bradykinin content, more greatly in the medulla than in the cortex (5.7 +/- 0.4 versus 4.6 +/- 0.1 ng/g). Similarly, cilazaprilat elicited greater bradykinin-dependent increases of nitrite/nitrate in the medulla. In conclusion, zonal heterogeneity in renal bradykinin/nitric oxide levels and segmental differences in reactivity to bradykinin contribute to the diverse responsiveness of renal AFF and EFF to ACEI. ACEI-enhanced kinin action would participate in the amelioration of glomerular hemodynamics and renal sodium excretion by ACEI. PMID- 10541287 TI - Bioflavonoid quercetin inhibits interleukin-1-induced transcriptional expression of monocyte chemoattractant protein-1 in glomerular cells via suppression of nuclear factor-kappaB. AB - Flavonoids are semiessential food components that possess anti-inflammatory properties. This report describes a novel potential of bioflavonoid quercetin as an inhibitor of monocyte chemoattractant protein-1 (MCP-1) in glomerular cells. Cultured mesangial cells as well as isolated glomeruli expressed MCP-1 mRNA in response to interleukin-1beta (IL-1beta). Quercetin dramatically inhibited the cytokine-triggered MCP-1 expression. To explore the mechanisms involved, effects of quercetin on the putative transcriptional activators of MCP-1, nuclear factor kappaB (NF-kappaB) and activator protein-1 (AP-1), were examined. Exposure of the cells to IL-1beta caused activation of NF-kappaB without significant upregulation of AP-1 activity. NF-kappaB inhibitor MG132 diminished the IL-1-induced expression of MCP-1 in mesangial cells and isolated glomeruli, whereas c-Jun/AP-1 inhibitor curcumin did not affect this process. Consistently, NF-kappaB-inactive mesangial cells expressing a super-repressor mutant of IkappaBalpha showed blunted expression of MCP-1 by IL-1beta. In contrast, AP-1-inactive mesangial cells expressing a dominant-negative mutant of c-Jun exhibited the same level of MCP-1 mRNA as that in control cells. These results suggest that: (1) quercetin has the ability to attenuate activation of NF-kappaB; and (2) it inhibits IL-1 triggered MCP-1 expression via suppression of NF-kappaB, but not AP-1, in glomerular cells. PMID- 10541288 TI - ATP depletion increases tyrosine phosphorylation of beta-catenin and plakoglobin in renal tubular cells. AB - This study examines the hypothesis that the loss of integrity of the junctional complex induced by ATP depletion is related to alterations in tyrosine phosphorylation of the adherens junction proteins beta-catenin and plakoglobin. ATP depletion of cultured mouse proximal tubular (MPT) cells induces a marked increase in tyrosine phosphorylation of both beta-catenin and plakoglobin. The tyrosine phosphatase inhibitor vanadate has the same effect in ATP-replete (control) monolayers, whereas genistein, a tyrosine kinase inhibitor, reduces phosphorylation of both proteins in ATP-replete monolayers and prevents the hyperphosphorylation of these proteins with ATP depletion. This study also demonstrates that the fall in the transepithelial resistance of MPT monolayers induced by ATP depletion can be reproduced by treatment of ATP-replete monolayers with vanadate, whereas genistein substantially ameliorates the fall in transepithelial resistance induced by ATP depletion. Also, using immunofluorescence microscopy it was demonstrated that ATP depletion results in a marked diminution of E-cadherin staining in the basolateral membrane of MPT cells. Vanadate mimics this effect of ATP depletion, whereas genistein ameliorates the reduction in the intensity of E-cadherin staining induced by ATP depletion. Because it is has been well established that hyperphosphorylation of the catenins leads to dissociation of the adherens junction and to dysfunction of the junctional complex, it is proposed that the increase in tyrosine phosphorylation of catenins observed in MPT cells during ATP depletion contributes to the loss of function of the junctional complex associated with sublethal injury. PMID- 10541289 TI - Secretion of platelet-activating factor is mediated by MDR1 P-glycoprotein in cultured human mesangial cells. AB - MDR1 P-glycoprotein (Pgp), the product of the MDR1 gene involved in multidrug resistance in cancer cells, is also expressed in normal tissues. In the human kidney, it is localized in the mesangium, the proximal tubule, the thick ascending limb of Henle's loop, and the collecting duct. Pgp actively transports lipophilic xenobiotics, peptides, steroids, and lipids, and perhaps endogenous substrates. It has been shown previously that human mesangial cells in culture express active Pgp and that the expression of Pgp can be down-regulated by exposure to antisense oligonucleotides. Mesangial cells do not express multidrug resistance-related protein (MRP). Experiments were performed to determine whether 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine (generically platelet-activating factor, PAF) is a substrate of Pgp in human mesangial cells in culture. This study found: (1) PAF C-16 and analogs inhibited Pgp-mediated efflux of rhodamine 123 by 59 to 88% in multidrug-resistant KBV-1 cells and by 85 to 97% in cultured human mesangial cells. (2) In mesangial cells stimulated with A23187, the secretion of endogenously produced PAF was inhibited by >80% by the Pgp blockers verapamil, cyclosporin A, PSC-833, vinblastine, and adriamycin. (3) Preincubation with MDR1 antisense oligonucleotides also blocked PAF secretion by human mesangial cells. PAF analogs do not modify the transport of MRP substrates in MCF 7/VP cells expressing MRP but not Pgp. These results indicate that PAF is an endogenous substrate of Pgp in human mesangial cells. Inhibition of Pgp transport may be useful in reducing glomerular damage occurring in pathologic conditions where PAF secretion is elevated. PMID- 10541290 TI - Chemokine and chemokine receptor expression in a novel human mesangial cell line. AB - Chemokines are thought to play a pivotal role in mediating the selective migration of leukocytes into sites of tissue injury. The local production of chemokines by mesangial cells (MC) has been linked to inflammatory processes within the glomerulus. To study the chemokine biology of human MC, an immortalized human MC line was generated and then chemokine and chemokine receptor expression was examined in response to various proinflammatory stimuli. The results show that human MC have a specific and limited repertoire of chemokine expression. The stimulus-specific regulation of the chemokines monocyte chemoattractant protein- (MCP- 1), regulated upon activation, normal T cell expressed and secreted (RANTES), interleukin-8 (IL-8), and IP-10 was demonstrated using RNase protection assays. Transcripts for the chemokines MIP-1alpha, MIP 1beta, I-309, or lymphotactin could not be detected. The expression of CC chemokine receptors was investigated by reverse transcription-PCR and RNase protection assays. MC stimulated with interferon-gamma (IFN-gamma) expressed mRNA for the chemokine receptor CCR1. The expression could be further increased by activating the cells with a combination of tumor necrosis factor-a (TNF-alpha), IL-1beta, and IFN-gamma. Under these conditions, no mRNA for CCR2, CCR3, CCR4, CCR5, or CCR8 was detected. A comparison of the immortalized human mesangial cells with primary cells showed identical expression patterns of chemokine receptors. To demonstrate functional activity of chemokine receptors expressed by human MC, chemotaxis assays were performed. MC stimulated with a combination of TNF-alpha, IL-1beta, and IFN-gamma, but not unstimulated MC, migrated toward a RANTES gradient. Eotaxin did not enhance the migratory activity of human MC. In summary, a novel human mesangial cell line was established and the pattern of chemokine expression was examined. For the first time, the inducible expression of functionally active CCR1 by human MC was shown. PMID- 10541291 TI - Complement membrane attack complex (C5b-9) mediates interstitial disease in experimental nephrotic syndrome. AB - Accumulating evidence suggests that the generation of complement activation products from filtered complement components in urine with nonselective proteinuria leads to tubulointerstitial disease, resulting in progressive loss of renal function. To elucidate the role of C5b-9 in complement-mediated effects on renal tubular cells exposed to proteinuric urine, equivalent levels of proteinuria were induced (using the aminonucleoside of puromycin) in normocomplementemic and genetically C6-deficient piebald viral glaxo (PVG) rats. Semiquantitative histologic analysis revealed that complement-sufficient animals developed more severe tubulointerstitial disease than did C6-deficient rats. Amelioration of tubulointerstitial damage in C6-deficient animals was confirmed by studies with three independent markers of tubular damage, i.e., vimentin, osteopontin, and proliferating cell nuclear antigen. More tubular epithelial cells expressed osteopontin (an early marker of tubular injury) in normocomplementemic rats, compared with C6-deficient rats, at both days 7 and 12. Staining of vimentin in the tubules, near areas of tubular damage, was increased in normocomplementemic rats at day 12, and more proliferating cell nuclear antigen-positive tubular cells were observed at day 12 in complement-sufficient animals. The tubulointerstitial damage in complement-sufficient rats was also associated with greater accumulation of extracellular matrix (fibronectin) at day 12. These studies document for the first time an important role for C6, and therefore C5b-9, in the pathogenesis of nonimmunologic tubulointerstitial injury induced by proteinuria. These findings suggest that C5b-9 formation resulting from proteinuria contributes to the loss of nephron function by damaging the tubulointerstitium and that prevention of C5b-9 formation in tubules could slow the deterioration of renal function. PMID- 10541292 TI - Identification of a unique glomerular factor X activator in murine lupus nephritis. AB - The role of glomerular procoagulant activity (PCA) was studied in mice (MRL/lpr, NZBxWF,, and BXSB) that are known to develop lupus nephritis. In young mice (6 to 8 wk) without renal disease, there was no increase in spontaneous glomerular PCA. In contrast, older (5 to 8 mo) autoimmune mice had significant augmentation in glomerular PCA, coinciding with the histologic appearance of severe glomerulonephritis and renal fibrin deposition. The PCA was characterized as a serine protease that directly activated factor X. This factor X activator is not tissue factor because (1) expression of PCA was not dependent on factor VII; (2) a monoclonal antibody against the factor X activator inhibited glomerular PCA, but not tissue factor; (3) the molecular weight (66 kD) of the activator was different from that of tissue factor; and (4) concanavalin A inhibited tissue factor but not glomerular PCA. Immunohistochemical studies localized the factor X activator to the glomerular mesangium and capillary wall of 4- to 6-mo-old diseased MRL/lpr mice. Immunogold-labeled antibody bound to the dense deposits, macrophages, and endothelial cells of diseased glomeruli. These studies define the role of a unique glomerular factor X activator in murine lupus nephritis. PMID- 10541293 TI - Aberrant splicing in the PKD2 gene as a cause of polycystic kidney disease. AB - It is estimated that approximately 15% of families with autosomal dominant polycystic kidney disease (ADPKD) have mutations in PKD2. Identification of these mutations is central to identifying functionally important regions of gene and to understanding the mechanisms underlying the pathogenesis of the disorder. The current study describes mutations in six type 2 ADPKD families. Two single base substitution mutations discovered in the ORF in exon 14 constitute the most COOH terminal pathogenic variants described to date. One of these mutations is a nonsense change and the other encodes an apparent missense variant. Reverse transcription-PCR from patient lymphoblast RNA showed that, in addition, both mutations resulted in out-of-frame splice variants by activating cryptic splice sites via different mechanisms. The apparent missense variant produced such a strong splicing signal that the processed transcript from the mutant chromosome did not contain any of the normally spliced, missense product. A third mutation, a nonconservative missense change effecting a negatively charged residue in the third transmembrane span, is likely pathogenic and defines a highly conserved residue consistent with a potential channel subunit function for polycystin-2. The remaining three mutations included two frame shifts resulting from deletion of one or two bases in exons 6 and 10, respectively, and a nonsense mutation due to a single base substitution in exon 4. The study also defined a novel intragenic polymorphism in exon 1 that will be useful in analyzing "second hits" in PKD2. Finally, the study demonstrates that there are reduced levels of normal polycystin-2 protein in lymphoblast lines from PKD2-affected individuals and that truncated mutant polycystin-2 cannot be detected in patient lymphoblasts, suggesting that the latter may be unstable in at least some tissues. The mutations described will serve as critical reagents for future functional studies in PKD2. PMID- 10541294 TI - Primary hyperoxaluria type I: a model for multiple mutations in a monogenic disease within a distinct ethnic group. AB - Primary hyperoxaluria type 1 is an autosomal recessive inherited metabolic disease in which excessive oxalates are formed by the liver and excreted by the kidneys, causing a wide spectrum of phenotypes ranging from renal failure in infancy to mere renal stones in late adulthood. Mutations in the AGXT gene, encoding the liver-specific enzyme alanine:glyoxylate aminotransferase, are responsible for the disease. Seven mutations were detected in eight families in Israel. Four of these mutations are novel and three occur in children living in single-clan villages. The mutations are scattered along various exons (1, 4, 5, 7, 9, 10), and on different alleles comprising at least five different haplotypes. All but one of the mutations are in a homozygous pattern, reflecting the high rate of consanguinity in our patient population. Two affected brothers are homozygous for two different mutations expressed on the same allele. The patients comprise a distinct ethnic group (Israeli Arabs) residing in a confined geographic area. These results, which are supported by previous data, suggest for the first time that the phenomenon of multiple mutations in a relatively closed isolate is common and almost exclusive to the Israeli-Arab population. Potential mechanisms including selective advantage to heterozygotes, digenic inheritance, and the recent emergence of multiple mutations are discussed. PMID- 10541295 TI - Systemic factors are involved in the pathogenesis of proteinuria-induced glomerulosclerosis in adriamycin nephrotic rats. AB - This study aims to dissociate the respective roles of systemic nephrosis and of the intrarenal effects of proteinuria in the pathogenesis of focal segmental glomerulosclerosis (FGS) in adriamycin nephrosis. To this purpose, this study examined proteinuria and FGS in bilateral (BAP) and unilateral proteinuria (UAP) in two different rat strains. UAP was obtained by protecting one kidney from exposure to adriamycin by temporary clipping of one renal artery during adriamycin injection. At sacrifice (week 12), FGS was present in BAP and in exposed kidneys in UAP, but not in unexposed kidneys. FGS correlated significantly with proteinuria per kidney in BAP and UAP. Remarkably, for a given proteinuria per kidney, the sclerosis score was higher in BAP than in UAP, reflected by a higher ratio of FGS score per mg proteinuria per kidney (Wistar: 0.09 +/- 0.01 in BAP versus 0.05 +/- 0.01%/mg protein per d in UAP, P < 0.05; Lewis: 0.12 +/- 0.01 in BAP versus 0.07 +/- 0.01%/mg protein per d in UAP, P < 0.05), indicating that the local damaging effects of proteinuria are modified by other factors. Cholesterol correlated with total proteinuria in BAP and UAP. FGS score was positively correlated with cholesterol. The latter correlation was similar in BAP and UAP, indicating that cholesterol was a more uniform predictor for FGS than proteinuria per kidney. This was independent of strain-specific factors. On multilinear regression analysis, cholesterol turned out to be the most consistent predictor of FGS in proteinuric kidneys, with a stronger predictive value than proteinuria per kidney. It is concluded that although systemic sequelae of nephrosis do not induce renal damage in nonproteinuric kidneys, they modify the severity of proteinuria-induced FGS in proteinuric kidneys. PMID- 10541297 TI - Growth hormone receptor antagonism prevents early renal changes in nonobese diabetic mice. AB - The growth hormone (GH)/insulin-like growth factor (IGF) axis is involved in diabetic renal disease. The role of a specific GH receptor (GHR) antagonist in the development of early renal changes in nonobese diabetic (NOD) mice was investigated. Female diabetic (nonketotic) NOD mice treated with a polyethylene glycol-treated GHR antagonist (2 mg/kg, every other day) (DA group) or saline (D group) and their nonhyperglycemic age-matched littermates (control animals) were euthanized 3 wk after the onset of diabetes. Body weights at euthanasia were similar among the groups. Serum GH levels were markedly elevated, and serum IGF-I levels were significantly decreased in D and DA animals, compared with controls. The increases in kidney weights and glomerular volumes observed for the D group were absent in the DA group. Albuminuria was increased in the D group but was normalized in the DA group. Extractable renal IGF-I protein levels were increased in the D group but were partially normalized in the DA group. Renal IGF-binding protein 1 mRNA levels were increased in the D group but returned to almost normal levels in the DA animals. Kidney IGF-I and GHR mRNA levels were decreased in both the D and DA groups. Renal GH-binding protein mRNA levels remained unchanged in both diabetic groups. GHR antagonism had a blunting effect on renal/glomerular hypertrophy and albuminuria in diabetic NOD mice. These salutary effects were associated with concomitant inhibition of increased renal IGF-I protein levels and were obtained without affecting either somatic growth or circulating GH and IGF-I levels. Therefore, modulation of GH effects may have beneficial therapeutic implications in diabetic nephropathy. PMID- 10541296 TI - Nutrition and chronic renal failure in rats: what is an optimal dietary protein? AB - In chronic uremia (CRF), malnutrition is an important determinant of morbidity in adults and impaired growth in children. Causes of malnutrition include anorexia and abnormal protein and amino acid metabolism. To determine how different levels of dietary protein and CRF interact to influence growth and nutritional status, CRF and sham-operated, pair-fed control rats were fed isocaloric diets containing 8, 17, or 30% protein for 21 d to mimic dietary regimens recommended for CRF patients: the minimum daily requirement; the recommended daily allowance; or an excess of dietary protein. Serum creatinine did not differ between groups of CRF rats but blood urea nitrogen was lowest in CRF rats fed 8% protein (P < 0.001). CRF rats eating 30% protein gained less weight and length compared to their controls or CRF rats fed 8 or 17% protein (P < 0.05); they also had acidemia. CRF rats fed 8% protein had the highest efficiency of utilization of protein for growth, while 17% protein promoted the highest efficiency of utilization of food and calories for growth. Notably, CRF rats eating 30% protein had the lowest protein efficiency; their calorie intake was also the lowest because of anorexia. Plasma branched-chain amino acids were progressively higher in control rats eating 8, 17, or 30% protein. CRF rats fed 8 or 17% protein had lower branched chain amino acid concentrations compared with CRF rats fed 30% protein. In CRF, it is concluded that excessive dietary protein impairs growth but a low-protein diet does not impair nutritional responses and permits utilization of protein for growth if calories are sufficient. PMID- 10541298 TI - The paradox of the low-renin state in diabetic nephropathy. AB - Although diabetic nephropathy is often a low renin state, the renin system appears to be implicated in its pathogenesis. In this study, it was hypothesized that the low plasma renin activity (PRA) is misleading, masking and perhaps reflecting an activated intrarenal renin system. PRA and renal vascular responses (inulin and para-aminohippurate clearance) to graded doses of an angiotensin II (AngII) antagonist, irbesartan, were assessed in eight healthy volunteers and 12 patients with type 2 diabetes mellitus and nephropathy on a 10 mmol Na intake, to activate the renin system. Basal PRA was suppressed in type 2 diabetes mellitus compared with the healthy subjects (0.58 +/- 0.14 versus 1.58 +/- 0.28 ng/L per s, mean +/- SEM; P < 0.01). Despite the low PRA, renal perfusion rose more in response to irbesartan in type 2 diabetes mellitus (714 +/- 83 to 931 +/- 116 ml/min; P = 0.002) than normal (624 +/- 29 to 772 +/- 49 ml/min; P = 0.008). The youngest patients were hyperfiltrating and showed the largest rise in renal plasma flow in response to irbesartan, whereas renal plasma flow rose less and GFR fell in patients with low basal GFR. PRA rose in response to irbesartan more gradually in the patients with type 2 diabetes mellitus, but ultimately matched the normal response. To account for the apparent paradox of a heightened renal hemodynamic response to an AngII antagonist in the face of a low PRA in type 2 diabetes mellitus, and the rise in PRA following the AngII antagonist, it is proposed that there is increased intrarenal AngII production in type 2 diabetes mellitus. This increase could account for suppressed circulating renin, the exaggerated renal vasodilator response to irbesartan, and the therapeutic effectiveness of interrupting the renin system in diabetic nephropathy. PMID- 10541299 TI - Pharmacokinetics of novel erythropoiesis stimulating protein compared with epoetin alfa in dialysis patients. AB - Novel erythropoiesis stimulating protein (NESP) is a hyperglycosylated analogue of recombinant human erythropoietin (Epoetin) which has an increased terminal half-life in animal models. The aim of this study was to extend these observations to humans. Using a double-blind, randomized, cross-over design, the single-dose pharmacokinetics of Epoetin alfa (100 U/kg) and an equivalent peptide mass of NESP were compared following intravenous bolus in 11 stable peritoneal dialysis patients. This was followed by an open-label study to determine the single-dose pharmacokinetics of an equivalent peptide mass of NESP by subcutaneous injection in six of these patients. The mean terminal half-life for intravenous NESP was threefold longer than for intravenous Epoetin (25.3 versus 8.5 h), a difference of 16.8 h (95% confidence interval, 9.4 to 24.2 h, P = 0.0008). The area under the serum concentration-time curve was significantly greater for NESP (291.0 +/- 7.6 ng x h per ml versus 131.9 +/- 8.3 ng x h per ml; mean +/- SEM; P < 0.0005), and clearance was significantly lower (1.6 +/- 0.3 ml/h per kg versus 4.0 +/- 0.3 ml/h per kg; mean +/- SEM; P < 0.0005). The volume of distribution was similar for NESP and Epoetin (52.4 +/- 2.0 ml/kg versus 48.7 +/-2.1 ml/kg; mean +/-SEM). The mean terminal half-life for subcutaneous NESP was 48.8 h. The peak concentration of subcutaneous NESP was approximately 10% of that following intravenous administration, and bioavailability was approximately 37% by the subcutaneous route. The longer half-life of NESP is likely to confer a clinical advantage over Epoetin by allowing less frequent dosing in patients treated for anemia. PMID- 10541300 TI - Acute renal failure after cardiopulmonary bypass in related to decreased serum ferritin levels. AB - Acute renal failure (ARF) requiring dialysis occurs in up to 4% of patients after cardiopulmonary bypass (CPB). CPB leads to the generation of intravascular free hemoglobin, resulting in increased endothelial and renal tubular cell free iron, which is associated with renal injury. Conversely, renoprotection is conferred by processes that upregulate heme and iron sequestration pathways, such as ferritin. This study evaluates the influence of free hemoglobin generation during CPB and the capacity to sequester free iron on the occurrence of post-CPB renal insufficiency. Thirty consecutive patients undergoing CPB were enrolled in the study. Serum creatinine, free hemoglobin, and ferritin were measured preoperatively, at the end of bypass, and 24 and 48 h after surgery. Renal injury, as determined by an increase in the serum creatinine of > or =25% (ARF) by 48 h after surgery, occurred in 40% (12 of 30) of patients, and dialysis was necessary in 6.6% (2 of 30). Free hemoglobin levels increased in all patients but did not correlate with postoperative ARF. However, patients with preoperative serum ferritin levels < or =130 microg/L, the median value for the group, had a sixfold greater likelihood of developing ARF compared to patients with levels above this value (P = 0.03). Lower serum ferritin levels appear to be associated with the development of ARF. Serum ferritin levels may signify intravascular as well as endothelial and renal epithelial cell ability to bind free iron generated during CPB-induced hemolysis, and thus may help provide information regarding the risk for ARF. PMID- 10541301 TI - Serum from hemodialysis patients inhibits basal and cytokine-stimulated tissue factor expression in vitro. AB - Hemorrhagic complications are common among hemodialysis (HD) patients. The mechanisms by which HD perturbs the coagulation cascade are still being defined. This study evaluated the influence of HD serum on cellular expression of tissue factor (TF), a procoagulant membrane-associated protein that is a pivotal regulator of blood coagulation. Serum was collected immediately before dialysis and 15, 30, and 180 min into HD using polysulfone membranes. Serum was then assessed for its ability to influence basal and cytokine-stimulated TF activity in human umbilical vein endothelial cells and ECV304 cells. Predialysis serum did not influence basal levels of TF activity. HD was associated with the appearance of a serum factor that suppressed basal TF activity (TF units/microg protein: predialysis serum 8.2 +/- 0.9; 180-min dialysis serum 4.9 +/- 0.6; P < 0.05) and TF activity induced by the cytokine tumor necrosis factor-alpha (TNFalpha) (TF units/microg protein: TNFalpha alone 15.9 +/- 0.7; TNFalpha + 180-min dialysis serum 5.9 +/- 0.9; P < 0.01). This response was not mimicked by heparin, suggesting production of an endogenous inhibitor of TF activity during HD. Dialysis was associated with a striking increase in circulating levels of tissue factor pathway inhibitor (TFPI), a physiologic inhibitor of the TF/VIIa complex. The lack of temporal correlation between TFPI levels and suppression of TF activity, however, suggested the presence of additional TFPI independent pathway(s) for modulation of TF activity. Dialysis-related suppression of TF expression may contribute to hemorrhagic complications in HD patients. PMID- 10541302 TI - Relationship between erythropoietin and chronic heart failure in patients on chronic hemodialysis. AB - In the present study, the relationship between the blood erythropoietin level and cardiac function was investigated in 15 patients on chronic hemodialysis who developed chronic heart failure. Another 45 patients without cardiac dysfunction were selected as a control group that was matched for gender, age, and the duration of dialysis. The erythropoietin level was 256.3 +/- 481.8 mU/ml in the heart failure group, which was significantly higher than that in the control group (17.0 +/- 10.0 mU/ml, P < 0.01). Eight of the 15 patients in the heart failure group maintained a hematocrit of more than 30% without receiving recombinant human erythropoietin therapy, whereas 29 of the 45 patients in the control group required erythropoietin. In the heart failure group, the erythropoietin level was significantly correlated with the levels of atrial natriuretic peptide and brain natriuretic peptide (P < 0.01). These results suggest that heart failure can increase the erythropoietin level in proportion to the severity of cardiac dysfunction, even in patients on long-term dialysis. PMID- 10541303 TI - Acute regulation of proximal tubule apical membrane Na/H exchanger NHE-3: role of phosphorylation, protein trafficking, and regulatory factors. PMID- 10541304 TI - Dietary protein restriction and the progression of chronic renal disease: what have all of the results of the MDRD study shown? Modification of Diet in Renal Disease Study group. AB - The Modification of Diet in Renal Disease (MDRD) Study was the largest randomized clinical trial to test the hypothesis that protein restriction slows the progression of chronic renal disease. However, the primary results published in 1994 were not conclusive with regard to the efficacy of this intervention. Many physicians interpreted the failure of the MDRD Study to demonstrate a beneficial effect of protein restriction over a 2- to 3-yr period as proving that this therapy does not slow disease progression. The authors believe that this viewpoint is incorrect, and is the result of misinterpretation of inconclusive evidence as evidence in favor of the null hypothesis. Since then, numerous secondary analyses of the MDRD Study have been undertaken to clarify the effect of protein restriction on the rate of decline in GFR, urine protein excretion, and onset of end-stage renal disease. This review describes some of the principles of secondary analyses of randomized clinical trials, presents the results of these analyses from the MDRD Study, and compares them with results from other randomized clinical trials. Although these secondary results cannot be regarded as definitive, the authors conclude that the balance of evidence is more consistent with the hypothesis of a beneficial effect of protein restriction than with the contrary hypothesis of no beneficial effect. Until additional data become available, physicians must continue to make recommendations in the absence of conclusive results. The authors suggest that physicians incorporate the results of these secondary analyses into their interpretation of the findings of the MDRD Study. PMID- 10541305 TI - Unraveling the mechanisms of glomerular ultrafiltration: nephrin, a key component of the slit diaphragm. PMID- 10541306 TI - Anti-glomerular basement membrane disease. PMID- 10541308 TI - Prediction of male adult stature using anthropometric data at birth: a nationwide population-based study. AB - Short stature and excess weight in adulthood are both associated with an increased risk of health problems. In a population-based investigation, data on birth length, birth weight, and gestational age for males born in Sweden in 1976 were used to predict the risk of being short or overweight in adulthood. The Swedish Birth Register was used to identify singleton males, born to Nordic mothers, who were without malformations and alive at 18 y of age. After individual record linkage between the Birth Register and the Swedish Conscript Register, information about height and weight at 18-21 y was obtained for 90% (n = 39901) of the birth cohort. Logistic regression analyses were used to estimate the risk of being short or overweight at conscription. The odds ratio (OR) was used to estimate relative risk. At conscription, mean height (+/-SD) was 179.5+/ 6.6 cm, mean weight 72.1+/-11.2 kg, and mean body mass index 22.3+/-3.1 kg/m2. The risk of short adult stature (<166.3 cm) was associated with being short for gestational age (OR = 5.9), having a low birth weight for gestational age (OR = 1.7), and being born at a gestational age below 32 wk (OR = 2.6). The risk of being overweight (body mass index > +2 SD) was primarily associated with a high ponderal index (> +2 SD; OR = 1.8). In conclusion, anthropometric birth data are better predictors of short stature than of being overweight in adulthood. Among anthropometric data at birth, birth length is the most important predictor of adult height. PMID- 10541307 TI - Transfer of specific unresponsiveness to organ allografts by thymocytes. PMID- 10541309 TI - Inverse relationship between serum inhibin B and FSH levels in prepubertal boys with cryptorchidism. AB - To investigate the gonadal control of FSH secretion in prepuberty, we studied the relationship between circulating inhibin B and FSH levels in 16 prepubertal boys with cryptorchidism (age range, 1-8 y). The effect of Leydig cell stimulation on the secretion of inhibin B, sex steroids, and FSH was investigated in nine boys who were given human chorionic gonadotropin (hCG) treatment. In these boys, serum inhibin B, testosterone, estradiol, and gonadotropin levels were measured before and on the fourth day of the last (third) hCG injection, given at 1-wk intervals. Except for one boy with both high inhibin B and FSH concentrations, basal serum levels of these hormones correlated negatively (r(s) = -0.79, n = 15, p < 0.005). This inverse relationship remained significant in the subgroup of boys younger than 2 y of age (r(s) = -0.84, n = 11, p = 0.008) who also had greater variance of serum FSH concentrations than 14 control boys of similar age with normally located testes (p < 0.01). hCG stimulation increased serum testosterone and suppressed serum FSH concentrations in each boy (n = 9, p < 0.005). In the four oldest subjects, the serum inhibin B level increased from the mean of 91 to 135 pg/mL (p < 0.05). These findings suggest that inhibin B regulates FSH secretion in early childhood. Moreover, the hCG-induced suppression of FSH secretion was probably mediated by sex steroids rather than by inhibin B. Finally, the increase in serum inhibin B concentration during the hCG treatment was likely to be indirect via Leydig cell-Sertoli cell or Sertoli cell-germ cell interaction(s). PMID- 10541310 TI - Alpha-mannosidosis in the guinea pig: a new animal model for lysosomal storage disorders. AB - Alpha-mannosidosis is a lysosomal storage disorder resulting from deficient activity of lysosomal alpha-mannosidase. It has been described previously in humans, cattle, and cats, and is characterized in all of these species principally by neuronal storage leading to progressive mental deterioration. Two guinea pigs with stunted growth, progressive mental dullness, behavioral abnormalities, and abnormal posture and gait, showed a deficiency of acidic alpha mannosidase activity in leukocytes, plasma, fibroblasts, and whole liver extracts. Fractionation of liver demonstrated a deficiency of lysosomal (acidic) alpha-mannosidase activity. Thin layer chromatography of urine and tissue extracts confirmed the diagnosis by demonstrating a pattern of excreted and stored oligosaccharides almost identical to that of urine from a human alpha mannosidosis patient. Widespread neuronal vacuolation was observed throughout the CNS, including the cerebral cortex, hippocampus, thalamus, cerebellum, midbrain, pons, medulla, and the dorsal and ventral horns of the spinal cord. Lysosomal vacuolation also occurred in many other visceral tissues and was particularly severe in pancreas, thyroid, epididymis, and peripheral ganglion. Axonal spheroids were observed in some brain regions, but gliosis and demyelination were not observed. Ultrastructurally, most vacuoles in both the CNS and visceral tissues were lucent or contained fine fibrillar or flocculent material. Rare large neurons in the cerebral cortex contained fine membranous structures. Skeletal abnormalities were very mild. Alpha-mannosidosis in the guinea pig closely resembles the human disease and will provide a convenient model for investigation of new therapeutic strategies for neuronal storage diseases, such as enzyme replacement and gene replacement therapies. PMID- 10541311 TI - Intrauterine growth retardation associated with maternal uniparental disomy for chromosome 6 unmasked by congenital adrenal hyperplasia. AB - We report the first case of maternal uniparental disomy for chromosome 6 (UPD6mat) ascertained through congenital adrenal hyperplasia (CAH), which arose because of reduction to homozygosity of an autosomal recessive mutation. This case suggests that UPD6mat is associated with intrauterine growth retardation (IUGR). A case of paternal UPD (involving only the short arm of chromosome 6) ascertained as CAH has previously been reported, but was not stated to have IUGR. Our patient was born with IUGR followed by extraordinarily good catch-up growth. She had a history of a marked lag in motor development. She presented at 2.65 y of age with pubarche of 3 mo duration, clitoral enlargement, and an advanced bone age. Simple virilizing CAH was diagnosed by elevations of plasma 17 hydroxyprogesterone and testosterone. Mutation analysis showed that the CAH was due to homozygosity for the 1172N exon 4 mutation. When parental DNA was examined, the mother was found to be heterozygous for the uncommon exon 4 mutation, while the father had no detectable mutations. DNA microsatellite analysis was subsequently performed on the patient and parents using polymorphic markers spanning the entire chromosome 6. Seven markers were informative for inheritance of a single maternal allele and absence of paternal alleles in the proband. Analysis of microsatellite markers from other chromosomes confirmed biparental inheritance at these loci. This combination of findings is diagnostic of UPD6mat. The only other reported case of UPD6mat was discovered serendipitously when genotyped for renal transplantation; this patient had a history of IUGR. Since both cases of UPD6mat had IUGR, the phenotype appears to include IUGR as well as the potential to unmask an autosomal recessive trait. PMID- 10541312 TI - Pulmonary hemodynamics in newborn piglets during hypoxemia and reoxygenation: blocking of the endothelin-1 receptors. AB - The effects of blocking endothelin (ET) receptors in pulmonary circulation during hypoxemia and reoxygenation were studied in five groups of piglets. Ten minutes before hypoxemia, the Hyp group (n = 10) was given saline and the 1-mg (n = 9) and 5-mg group (n = 9), respectively, were given 1 and 5 mg/kg i.v. SB 217242 (an ET receptor antagonist). Two groups served as normoxic controls. The piglets were ventilated with 8% O2 until base excess was <-20 mmol/L or mean arterial blood pressure was <20 mm Hg. Reoxygenation was performed with air. The increase of mean pulmonary artery pressure was significantly attenuated during hypoxemia and reoxygenation in the 1-mg group (p = 0.006). The pulmonary vascular resistance index increased significantly at the end of hypoxemia in the Hyp and 5-mg groups but was comparable to baseline in the 1-mg group. During the study period, the changes in pulmonary vascular resistance index were significantly attenuated in the 1-mg group compared with the 5-mg group. Stroke volume index was significantly attenuated compared with baseline in the 5-mg group during both hypoxemia and reoxygenation, whereas, in the Hyp and 1-mg group, stroke volume index was attenuated only at the end of hypoxemia. During hypoxemia, plasma ET-1 decreased from 1.9+/-0.2 to 1.3+/-0.3 ng/L (p = 0.008) in the Hyp group, remained unchanged in the 1-mg group, and increased from 1.6+/-0.2 to 6.6+/-1.6 ng/L (p = 0.008) in the 5-mg group. We conclude that blocking ET receptors attenuates pulmonary vasoconstriction during hypoxemia and reoxygenation in piglets. PMID- 10541313 TI - KGF and FGF-10 stimulate liquid secretion in human fetal lung. AB - During fetal life, the pulmonary epithelium secretes liquid that distends the airways and is important for normal lung growth and development. The factors regulating human fetal lung liquid secretion are poorly understood; however, recent studies in murine models show that keratinocyte growth factor (KGF, FGF-7) and fibroblast growth factor 10 (FGF-10) stimulate liquid secretion. We asked whether KGF and FGF-10 stimulate liquid secretion in human fetal lung. First trimester fetal lung explants developed dose-dependent increases in intraluminal volume in response to KGF and FGF-10. Although there were no acute changes in explant transepithelial potential difference in response to KGF (0.1-1000 ng/mL), exposure to 5-50 ng/mL KGF over 60 h depolarized transepithelial potential difference compared with controls. We used ribonuclease protection assays to quantitate the ontogeny and regulation of mRNA expression for KGF and its receptor. Both mRNA were expressed in fetal and postnatal lung. Because the promoter region of the human KGF gene contains cAMP and IL-6 response elements, we asked whether cAMP or IL-6 stimulated expression of KGF or its receptor. We have previously shown that cAMP stimulates liquid secretion in this model. Both cAMP and IL-6 significantly increased expression of KGF but not KGF receptor during a 48-h experiment. Thus, stimulation of liquid secretion in explant models by cAMP may be mediated in part by induction of KGF expression. KGF and FGF-10 may be important paracrine factors regulating liquid secretion in human fetal lung. PMID- 10541314 TI - Altered surfactant protein B levels in transgenic mice do not affect clearance of bacteria from the lungs. AB - To determine the role of surfactant protein B (SP-B) in bacterial clearance from the airways, three groups of mice expressing different levels of SP-B were studied: wild-type mice, hemizygous SP-B mice, and SP-B overexpressing transgenic mice. SP-B levels in overexpressing mice were increased 5-fold relative to hemizygous mice and 2- to 3-fold over wild-type littermates. Mice from each group were infected intratracheally with the common airway pathogens, group B streptococci or Pseudomonas aeruginosa. There was no significant difference in the number of recoverable viable bacteria at 6 h (group B streptococci and P. aeruginosa) and at 24 h (P. aeruginosa) among the three groups. Similarly, systemic dissemination of bacteria was not different among the three groups for both pathogens and at both time points. We conclude that SP-B levels in vivo do not influence clearance of bacteria from the lungs. PMID- 10541315 TI - Ventilatory responses to hypercapnia and hypoxia in Mash-1 heterozygous newborn and adult mice. AB - Normal control of breathing is characterized by maintenance of CO2 and O2 arterial pressures at constant levels by appropriate ventilatory responses to changes in CO2 production and O2 consumption. Abnormal development of this regulatory system during embryogenesis may produce early impairments in chemosensitivity, as in congenital central hypoventilation syndrome. The present study addresses the role of the mammalian achaetescute homologous gene (Mash-1) in the development of respiratory control. We analyzed ventilatory responses to hypercapnia (8% CO2, 21% O2, 71% N2) and hypoxia (10% O2, 3% CO2, 87% N2) in newborn and adult Mash-1 heterozygous mice (Mash-1+/-) and their wild-type littermates (Mash-1+/+). Ventilation, breath duration, and tidal volume were measured using whole-body plethysmography. Ventilatory responses to hypercapnia were significantly weaker in newborn male Mash-1+/- compared with Mash-1+/+ mice as a result of a weaker breath-duration response. No differences were observed between adult Mash-1+/- and Mash-1+/+ mice. Our data suggest that Mash-1 may be involved in respiratory control development via mechanisms linked to the X chromosome. PMID- 10541316 TI - Erythropoietin in the cerebrospinal fluid of neonates who sustained CNS injury. AB - We previously reported that erythropoietin (Epo) is present in human cerebrospinal fluid (CSF). It is not known whether CSF Epo concentrations change under conditions of CNS injury or, if so, whether this change reflects loss of blood-brain barrier integrity or increased CNS Epo synthesis. We hypothesized that CSF Epo increases in conditions of neural injury including hypoxia, meningitis, and intraventricular hemorrhage (IVH) and that CSF Epo concentrations are independent of plasma Epo concentrations. To test these hypotheses, Epo concentrations were measured in 122 paired CSF and blood samples obtained from neonates and children categorized as follows: 16, asphyxia; 31, meningitis; 11, IVH; 41, controls. Twelve infants treated with recombinant Epo (rEpo) and 11 additional samples from children with miscellaneous neurologic problems were also evaluated. CSF and plasma Epo concentrations were significantly higher in asphyxiated infants than in controls (225.0+/-155.0 versus 4.5+/-0.5 mU/mL; mean +/- SEM, p < 0.05, respectively, in CSF; 1806.7+/-1254 versus 5.2+/-0.5, p < 0.05 in plasma). Neonates with IVH had higher CSF Epo concentrations than controls (p < 0.01) but did not have higher plasma Epo concentrations than controls. Patients with meningitis did not have elevated CSF or plasma Epo concentrations. There was no correlation between CSF and plasma Epo concentrations in infants treated with rEpo. We conclude that Epo is selectively increased in the CSF by hypoxia, less so by IVH, and not at all by meningitis. rEpo treatment does not elevate CSF Epo. These findings suggest that rEpo does not cross the blood-brain barrier and that hypoxia induces increased CNS synthesis of Epo. PMID- 10541317 TI - Familial dominant thrombocytopenia: clinical, biologic, and molecular studies. AB - Inherited thrombocytopenias are a heterogenous group of disorders. Different criteria have been suggested to classify the forms, such as the inheritance mechanism and the platelet volume as well as the number and morphology of megakaryocytes. However, the classification is often descriptive, and the precise mechanism of thrombocytopenia still remains unknown. We describe the clinical, biologic, and molecular findings of an autosomal dominant thrombocytopenia in a large family. The 17 patients had normocellular bone marrow and normal platelet volume. Platelets also showed a normal aggregation test and normal response to ADP and thrombopoietin (TPO). In the affected subjects, the mean +/- SD levels of platelet count and plasma TPO were 62+/-25 and 258+/-151, respectively. Comparative analysis showed that the patients with platelet count <70000 had higher plasma TPO concentration. The data are consistent with a mild clinical form of the disease associated with only a few episodes of bleeding. To exclude the possible role of TPO and its receptor c-mpl in the etiology of this condition, linkage analysis was performed using microsatellite markers close to the TPO and c-mpl genes on chromosomes 3q26.3-q27 and 1p34, respectively. The absence of cosegregation within the affected family indicated that these genes, as well as two other candidate loci on chromosomes 11 and 21, are not responsible for this hereditary dominant form of thrombocytopenia. A genome-wide search and subsequent identification of the gene will provide new insight into the pathogenesis of this disorder. PMID- 10541318 TI - Identification of new prognosis factors from the clinical and epidemiologic analysis of a registry of 229 Diamond-Blackfan anemia patients. DBA group of Societe d'Hematologie et d'Immunologie Pediatrique (SHIP), Gesellshaft fur Padiatrische Onkologie und Hamatologie (GPOH), and the European Society for Pediatric Hematology and Immunology (ESPHI). AB - Diamond-Blackfan anemia (DBA) is a constitutional disease characterized by a specific maturation defect in cells of erythroid lineage. We have assembled a registry of 229 DBA patients, which includes 151 patients from France, 70 from Germany, and eight from other countries. Presence of malformations was significantly and independently associated with familial history of DBA, short stature at presentation (before any steroid therapy), and absence of hypotrophy at birth. Two hundred twenty-two patients were available for long-term follow-up analysis (median, 111.5 mo). Of these individuals, 62.6% initially responded to steroid therapy. Initial steroid responsiveness was found significantly and independently associated with older age at presentation, familial history of DBA, and a normal platelet count at the time of diagnosis. Severe evolution of the disease (transfusion dependence or death) was significantly and independently associated with a younger age at presentation and with a history of premature birth. In contrast, patients with a familial history of the disease experienced a better outcome. Outcome analysis revealed the benefit of reassessing steroid responsiveness during the course of the disease for initially nonresponsive patients. Bone marrow transplantation was successful in 11/13 cases; HLA typing of probands and siblings should be performed early if patients are transfusion dependent, and cord blood should be preserved. Incidence of DBA (assessed for France over a 13-y period) is 7.3 cases per million live births without effect of seasonality on incidence of the disease or on malformative status. Similarly, no parental imprinting effect or anticipation phenomenon could be documented in families with dominant inheritance. PMID- 10541319 TI - Treatment of the Kasabach-Merritt syndrome with pegylated recombinant human megakaryocyte growth and development factor in mice: elevated platelet counts, prolonged survival, and tumor growth inhibition. AB - Kasabach-Merritt Syndrome (KMS) is seen in children with large vascular tumors. KMS is characterized by very low platelet counts and a consumption of coagulation factors causing life-threatening complications. It has been proposed that thrombopenia in these patients is caused by intratumoral trapping of platelets. The truncated form of the cMpl-receptor ligand thrombopoietin, pegylated human megakaryocyte growth and development factor (Peg-rHuMGDF), is an agent that stimulates platelet production. We hypothesized that stimulation of the platelet production would prevent the life-threatening complications of patients with KMS owing to low platelet counts. In a mouse model of KMS, with tumors derived from a hemangioendothelioma cell line, we studied the effect of Peg-rHuMGDF. Treatment with Peg-rHuMGDF (10 microg/kg/day intraperitoneally) increased platelet counts by 7-8-fold compared with control tumor-bearing mice after 11 d of treatment (p < 0.001, n = 8). Survival was significantly increased, with 50% of treated animals alive at 1 mo versus 0% in untreated controls. Interestingly, we also observed an inhibition of tumor growth by 75% (p < 0.001, n = 8). Hematoxylin and eosin staining showed fresh fibrin clots in the treated tumors, suggesting that higher platelet counts caused intravascular thrombosis of tumor vessels. We conclude that increased platelet production in this model of KMS resulted in an antivascular tumor effect via platelet trapping. Further, we propose that thrombopoietin may be of critical value in preventing life-threatening complications from KMS. PMID- 10541321 TI - Transforming growth factor-beta1: a possible signal molecule for posthemorrhagic hydrocephalus? AB - Posthemorrhagic hydrocephalus remains a complication of preterm birth for which we lack a clear understanding and a curative therapy. Transforming growth factor beta (TGF-beta) is a cytokine that upregulates the production by fibroblasts of extracellular matrix proteins. We hypothesized that TGF-beta might be released into cerebrospinal fluid (CSF) after intraventricular hemorrhage and play a role in posthemorrhagic hydrocephalus. Total TGF-beta1 and TGF-beta2 were measured by immunoassay in CSF samples from 12 normal preterm infants, nine preterm infants with transient posthemorrhagic ventricular dilation, and 10 infants who subsequently developed permanent hydrocephalus. Five infants received intraventricular tissue plasminogen activator, and two infants were treated by drainage irrigation and fibrinolytic therapy. Median TGF-beta1 in normal CSF was 0.495 ng/mL. In infants with transient posthemorrhagic ventricular dilation, median initial CSF TGF-beta1 was 2.1 ng/mL. Infants who subsequently had permanent hydrocephalus had median initial CSF TGF-beta1, 9.7 ng/mL (differences between groups p < 0.01). Intraventricular recombinant tissue plasminogen activator was followed by a rise in CSF TGF-beta1 (p = 0.0007). Drainage irrigation and fibrinolytic therapy was followed by a fall in CSF TGF-beta1. TGF beta2 was detected in CSF and showed similar trends, but the CSF concentration of TGF-beta1 was more than 20 times higher. These findings support the hypothesis that TGF-beta1 is released into CSF after intraventricular hemorrhage and may play an important part in hydrocephalus. The results help to explain the failure of intraventricular fibrinolytic therapy. PMID- 10541320 TI - Maternal infection, fetal inflammatory response, and brain damage in very low birth weight infants. Developmental Epidemiology Network Investigators. AB - Echolucent images (EL) of cerebral white matter, seen on cranial ultrasonographic scans of very low birth weight newborns, predict motor and cognitive limitations. We tested the hypothesis that markers of maternal and feto-placental infection were associated with risks of both early (diagnosed at a median age of 7 d) and late (median age = 21 d) EL in a multi-center cohort of 1078 infants <1500 x g. Maternal infection was indicated by fever, leukocytosis, and receipt of antibiotic; fetoplacental inflammation was indicated by the presence of fetal vasculitis (i.e. of the placental chorionic plate or the umbilical cord). The effect of membrane inflammation was also assessed. All analyses were performed separately in infants born within 1 h of membrane rupture (n = 537), or after a longer interval (n = 541), to determine whether infection markers have different effects in infants who are unlikely to have experienced ascending amniotic sac infection as a consequence of membrane rupture. Placental membrane inflammation by itself was not associated with risk of EL at any time. The risks of both early and late EL were substantially increased in infants with fetal vasculitis, but the association with early EL was found only in infants born > or =1 after membrane rupture and who had membrane inflammation (adjusted OR not calculable), whereas the association of fetal vasculitis with late EL was seen only in infants born <1 h after membrane rupture (OR = 10.8; p = 0.05). Maternal receipt of antibiotic in the 24 h just before delivery was associated with late EL only if delivery occurred <1 h after membrane rupture (OR = 6.9; p = 0.01). Indicators of maternal infection and of a fetal inflammatory response are strongly and independently associated with EL, particularly late EL. PMID- 10541322 TI - Bone and collagen markers in preterm infants: relationship with growth and bone mineral content over the first 10 weeks of life. AB - In a longitudinal study of 25 preterm infants, we have examined the relationship of bone-specific alkaline phosphatase (ALP), C-terminal propeptide of type I collagen (PICP), N-terminal propeptide of type III procollagen (P3NP), C-terminal telopeptide of type I collagen, urinary pyridinoline (Pyd) and deoxypyridinoline (Dpd), with rates of gain in weight, length, and lower leg length and with bone mineral content (BMC), all measured at weekly intervals over the first 10 wk of life. Concentrations of all collagen markers were 10-fold higher than in older children. Each marker showed a distinctive pattern of postnatal change, with early increases in PICP and P3NP and decreases in ICTP reflecting postnatal growth. Once markers had reached a plateau during weeks 4-10, P3NP was positively correlated, whereas Pyd and Dpd were negatively correlated with rate of weight gain (r = +0.44, -0.46, and -0.40, respectively, p < 0.05). P3NP was also positively correlated with overall linear growth (r = +0.44, p < 0.05). PICP was strongly correlated with mean BMC (r = +0.63,p < 0.01) and with total BMC attained by the end of the study period (r = +0.81, p < 0.001). Bone ALP was positively correlated with the rate of bone mineral accretion (r = +0.55, p = 0.01). We conclude that the marker of soft-tissue collagen formation, P3NP, is a good marker for overall ponderal and linear growth in preterm infants, whereas the markers of collagen breakdown, Pyd and Dpd, have inverse relationships with weight gain. The osteoblast markers, PICP and bone ALP, seem to be good surrogate markers for bone mineralization in preterm infants. Markers may provide information on whole-body turnover of bone and collagen that is complementary to traditional physical measures of growth and bone mineralization. PMID- 10541323 TI - Correction of ureagenesis after gene transfer in an animal model and after liver transplantation in humans with ornithine transcarbamylase deficiency. AB - We report effects of gene transfer and liver transplantation on urea synthesis in ornithine transcarbamylase deficiency (OTCD). We measured the formation of [15N] urea after oral administration of 15NH4Cl in two girls with partial OTCD before and after liver transplantation. Ureagenesis was less than 20% of that observed in controls before transplantation, and was normalized afterward. Studies performed on the OTCD sparse fur (spf/Y) mouse showed discordance between OTC enzyme activity and ureagenesis with modest increases in OTC enzyme activity after gene transfer resulting in significant improvement in ureagenesis. This study suggests that both liver transplantation and gene therapy may be effective in improving ureagenesis in OTCD. PMID- 10541324 TI - Molecular correlations in phenylketonuria: mutation patterns and corresponding biochemical and clinical phenotypes in a heterogeneous California population. AB - We studied 133 California phenylketonuria (PKU) patients and one obligate heterozygote to delineate the molecular basis of PKU in a population with greater ethnic diversity than in previous studies, and to determine whether a correlation exists between genotype and clinical phenotype, with the latter defined by both the diagnostic pretreatment blood phenylalanine (PHE) level and cognitive (IQ) test scores. To determine PAH genotypes, we used PCR-mediated amplification, denaturing gradient gel electrophoresis, and direct sequencing on dried whole blood samples. Where possible, mutation severity was defined according to predicted in vitro PAH enzyme activity estimated by using Cos cell expression analysis for a given mutation. We then asked whether mutation severity, as defined by such expression analysis, correlated with pretreatment PHE levels or with IQ test results. A mutation was identified in 236 (88%) of 267 mutant alleles. Seventeen new mutant alleles were found; A47E, T81P, I102T, E182G, T328D, Y343P, K371R, Y387H, A389E, E422K, IVS9nt5, IVS11nt20, delS70, del364 368/del198-220, delF299, delT323, and -1C/T. In striking contrast to a number of studies in other populations, in this study, based on predicted PAH activity, we observed no correlation between mutation severity and pretreatment PHE levels. There was also no correlation between genotype and IQ. We conclude that in samples collected from an ethnically heterogeneous population, there is no correlation of mutation severity with either pretreatment PHE levels or IQ measurement in treated patients. We caution that genetic counseling in PKU should incorporate the notion that prognosis may not be predicted with precision based on mutation analysis in a given patient. PMID- 10541325 TI - Age-dependent increase of IgE-binding and FcepsilonRI expression on circulating basophils in children. AB - Peripheral blood basophils are sparse in the circulation, but they express high affinity receptors for IgE (FCepsilonRI) and bind IgE efficiently. The present study was performed to elucidate the role of IgE bound on the basophil surface in the development of allergic responses during infancy and early childhood. IgE binding and FcepsilonRI expression on basophils were evaluated by two-color flow cytometry. Basophil-bound IgE increased rapidly and reached adult levels during infancy in atopic patients, while it gradually increased with advancing age in parallel with serum IgE in normal controls. IgE-binding and FcepsilonRI expression in atopic children were higher than in normal controls among various age groups. They correlated with serum IgE levels but reached a plateau when serum IgE exceeded 300 ng/mL. A low, but significant level of FcepsilonRI expression was observed on cord blood basophils, although IgE-binding was usually undetectable. Incubation of cord blood with IgE rapidly saturated the preexisting IgE receptors and basophil-bound IgE levels increased. When neonatal basophils were cultured for 48 h with IgE, FcepsilonRI expression was upregulated dose dependently and IgE-binding increased further. The up-regulation of FcepsilonRI was completely inhibited by cycloheximide, indicating that it was dependent on de novo protein synthesis. These results suggest that IgE-binding on basophils serves as a sensitive indicator of allergic sensitization, and that IgE functions as a positive regulator of FcepsilonRI expression in vivo. PMID- 10541326 TI - Local complement genes expression in the mammary gland: effect of gestation and inflammation. AB - Complement components in breast milk may enhance the local immune response in the gut of infants. In this study, we investigated the expression of complement genes in the mammary gland and attempted to determine possible regulatory mechanisms. We have studied the expression of C3, C4, factor B, and HLA-DRalpha mRNA by in situ hybridization in gestational mammary gland specimens and compared these findings to those in breast tissue affected with an inflammatory process, lactating adenoma or idiopathic gynecomastia. In normal resting breast, only C4 mRNA was noted in some ductal epithelium. In gestational mammary gland, there was a diffuse expression of C4, C3, and factor B mRNA in the epithelial cells of the acini. A similar pattern of complement gene expression was found in localized areas of an infectious inflammatory process. In addition, in the inflammatory specimens, there was also expression of C3 mRNA in infiltrating macrophages (CD 68 positive cells). In gynecomastia, C4 mRNA was noted in ductal epithelium, and there was a marked increased expression of C3 mRNA in the proliferating epithelium of the lactating adenoma. HLA-DRalpha was observed only in macrophages involved in the inflammatory response. Our findings, which reflect the hormonal and inflammatory events in vivo, provide new insights as to in situ complement gene expression. PMID- 10541328 TI - Development of a system to record cardiac output continuously in the newborn. AB - Intermittent recordings of Doppler flow velocity and cardiac output are of value during intensive care of the sick newborn infant but result in repeated disturbance of the child. We describe a new device for making continuous precordial recordings of Doppler flow velocity from the pulmonary artery in healthy resting newborn infants. Optimal probe siting was evaluated in six babies, and signals were found to be best when the pulmonary artery was insonated from the mid left parasternum. Continuous recordings were made in 13 other babies. Pulmonary artery velocities and, by calculation, cardiac output were measured continuously over periods ranging from 24 to 60 min. Median right ventricular output ranged widely from 148 to 246 mL x kg(-1) x min(-1). In contrast, for individual babies, the values were remarkably stable: the interquartile ranges varied from 13.2 to 29.9 mL x kg(-1) x min(-1). The simultaneous display of signal power allowed independent assessment of artifactual changes in cardiac output. This technique is feasible in healthy term infants and now requires evaluation in the intensive care setting where it may provide useful information concerning trends and short-term variability in right ventricular output. PMID- 10541327 TI - Granulocyte-macrophage colony-stimulating factor increases surfactant phospholipid in premature rabbits. AB - Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a hematopoietic cytokine that is low in airway specimens from immature lungs at birth. In adult mice, an absence of GM-CSF causes excessive accumulation of alveolar surfactant due to a lack of catabolism. Our aim was to investigate whether recombinant human GM-CSF (rhGM-CSF) affects the pool sizes or the turnover of disaturated phosphatidylcholine (DPC) in preterm (gestation 29 d) rabbits at birth and in term rabbits, age 3 d. 3H-labeled dipalmitoyl phosphatidylcholine, 14C-acetate, and either rhGM-CSF (125 or 25 microg/kg body weight) or placebo were given intratracheally. Thereafter, the intra- and extracellular surfactant fractions were isolated and quantified for DPC and radioactivity. In preterm animals, GM CSF increased dose-dependently within 24 h both the pool sizes of surfactant DPC and the 3H,14C-labeling of surfactant DPC (p < 0.05). The expression of surfactant protein B mRNA was unaffected, whereas surfactant protein B in bronchoalveolar lavage increased. The number of cells in the whole lung, the type II alveolar epithelial cells, and the lavageable alveolar macrophages were unaffected. At term, rhGM-CSF increased the turnover but did not affect the pool sizes of surfactant DPC. Intraperitoneal rhGM-CSF increased blood eosinophils but had no effect on surfactant DPC. Depending on the degree of lung maturity, GM-CSF in the alveolar space may either up-regulate the pool size or increase the turnover of surfactant phospholipid after birth. PMID- 10541329 TI - Human meconium has high phospholipase A2 activity and induces cellular injury and apoptosis in piglet lungs. AB - Aspiration of meconium produces an inflammatory reaction resulting in necrotic changes in lung tissue. To further investigate the mechanisms of the meconium induced early pulmonary injury, twenty 10-12-d-old piglets were studied for lung tissue ultrastructural and apoptotic changes and phospholipase A2 activity. Twelve piglets received an intratracheal bolus (3 mL/kg) of a 20-mg/mL (thin, n = 6) or 65-mg/mL (thick, n = 6) mixture of human meconium, and control piglets (n = 5) received the same amount of intratracheal saline. Three ventilated piglets with no aspiration were also studied. Pulmonary hemodynamics and systemic oxygenation were followed for 6 h after meconium or saline insufflation. In the control groups, the pulmonary tissue showed open alveolar spaces and intact vascular walls, whereas meconium administration resulted in severe pneumonitis, with alveolar spaces filled with inflammatory exudate. Meconium instillation additionally resulted in edematous changes in the vascular walls and alveolar epithelium, whereas type II pneumocytes were intact. The amount of apoptotic cells was increased, especially in the respiratory epithelium, and the catalytic activity of phospholipase A2 in lung tissue samples was significantly elevated after thick meconium instillation. This activity rise proved to be mainly because of human group I phospholipase A2, introduced by meconium. Our data thus show that aspiration of meconium leads to severe lung tissue inflammation with early ultrastructural changes in the pulmonary alveolar walls and is associated with apoptotic cell death in the epithelium, already during the first hours after the insult. These results further suggest that high phospholipase A2 activity, mainly introduced into the lungs within the meconium, may have an important role in the initiation of these alterations in neonatal lungs. PMID- 10541330 TI - Antenatal dexamethasone suppresses tumor necrosis factor-alpha expression in hypoplastic lung in nitrofen-induced diaphragmatic hernia in rats. AB - The hypoplastic lung in congenital diaphragmatic hernia (CDH) has both a quantitative and qualitative reduction in surfactant. Tumor necrosis factor-alpha (TNF-alpha) drastically decreases surfactant phospholipids synthesis by isolated human type II pneumocytes. Recently, it was shown that TNF-alpha mRNA expression is increased in human hypoplastic CDH lung. Antenatal glucocorticoid therapy demonstrates improved surfactant biochemical immaturity in an animal CDH model. The aim of this study was to investigate the effect of antenatal dexamethasone (Dex) on TNF-alpha protein and gene expression in nitrofen-induced CDH hypoplastic lung in rats. A CDH model was induced in pregnant rats after the administration of nitrofen on d 9.5 of gestation. Dex was given intraperitoneally on d 18.5 and 19.5. Cesarean section was performed on d 21. In situ hybridization was performed with a rat TNF-alpha-specific and digoxigenin-labeled oligonucleotide probe. TNF-alpha level was measured in solubilized lung tissue extracts by ELISA. In control lung, TNF-alpha mRNA expression was weak or absent. In contrast, strong TNF-alpha mRNA expression was demonstrated in type II pneumocytes and bronchiolar epithelium in CDH lung. In Dex-treated CDH lung, TNF alpha mRNA expression was weak in both type II pneumocytes and the bronchiolar epithelium. The level of TNF-alpha was elevated significantly in CDH lung compared with levels in control lung extracts (p < 0.01). In Dex-treated CDH lung, TNF-alpha protein was significantly decreased compared with CDH lung (p < 0.05). Our findings suggest that the reduction in the local production of TNF alpha may be one contributing mechanism by which antenatal glucocorticoid therapy improves pulmonary parenchymal immaturity, including surfactant. PMID- 10541331 TI - Introduction to the Seventh Conference on Radioimmunodetection and Radioimmunodetection and Radioimmunotherapy of Cancer. PMID- 10541332 TI - Tribute to David A. Goodwin and Claude F. Meares, recipients of the Immunomedics Science Award. PMID- 10541333 TI - Biodistribution study of 188Re-labeled trisuccin-HuCC49 and trisuccin HuCC49deltaCh2 conjugates in athymic nude mice bearing intraperitoneal colon cancer xenografts. AB - The trihydroxamate bifunctional chelating agent (BCA), trisuccin, has been shown to be a potential ligand for radiolabeling of monoclonal antibodies (MAbs) with rhenium radioisotopes, through an indirect postconjugation approach. The use of this trihydroxamate BCA made it possible to prepare stable BCA-MAb conjugates in pure form that could be radiolabeled with carrier-free 188Re. The anti-TAG-72 murine MAb, CC49, and its humanized derivatives are promising agents in the treatment of a number of malignancies with the CH2 domain-deleted MAb (HuCC49deltaCH2), which is of particular interest due to its rapid blood clearance. The biodistribution of 188Re-labeled conjugates of trisuccin with both humanized CC49 (HuCC49) and HuCC49deltaCH2 in athymic nude mice implanted i.p. with LS174T human colon carcinoma was studied. Trisuccin-MAb conjugates were synthesized at different BCA:MAb ratios by the 6-oxoheptanoic acid method using trisuccin hydrazide. The conjugates were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy for the number of incorporated trisuccin molecules. The conjugates were radiolabeled with carrier free, generator-produced 188Re and purified by gel filtration on Sephadex G-25. Labeling yields and homogeneity of the labeled conjugates were analyzed by high pressure liquid chromatography and instant TLC. Athymic nude mice were injected i.p. with LS174T human colon carcinoma cells, 7 days prior to injection of the labeled antibodies. 188Re-labeled MAbs were injected i.p., and the mice were sacrificed 24 h postinjection. Matrix-assisted laser desorption/ionization time of-flight analyses showed stable incorporation of trisuccin into each MAb, with the measured ligand:MAb values positively correlating with the theoretical ratios. Labeling of the conjugates with 188Re proceeded with high yields, producing homogeneous 188Re-MAbs with good stabilities as shown by instant TLC and biodistribution analyses. Biodistribution of the radiolabeled MAbs at 24 h after injection showed median tumor uptake values of 23.5%ID/g and 17.6%ID/g for the 188Re-HuCC49deltaCH2 and 188Re-HuCC49, respectively. The blood clearance of the domain-deleted MAb was faster than that of the intact antibody. The blood values at 24 h after injection were 0.7%ID/g for 188Re-HuCC49deltaCH2 and 3.2%ID/g for 188Re-HuCC49. The results indicate that trisuccin is a promising agent for postconjugation labeling of antibodies with 188Re. Additionally, these results illustrate the potential of 188Re-HuCC49deltaCH2 in radioimmunodiagnosis and radioimmunotherapy of cancer. PMID- 10541334 TI - Radiolabeling of an anti-carcinoembryonic antigen antibody Fab' fragment (CEA Scan) with the positron-emitting radionuclide Tc-94m. AB - The goal of this work was to test whether an antibody-based agent approved for use as a single-photon-emitting imaging agent when radiolabeled with technetium 99m could be labeled comparably with a positron-emitting nuclide, technetium-94m. "Instant kits" containing lyophilized NP-4 antibody Fab' fragment of an anticarcinoembryonic antigen IgG (CEA-Scan) from the same manufactured lot were reconstituted with either Tc-99m or Tc-94m, as solutions of sodium pertechnetate in isotonic saline solution. Radioanalyses of the labeled Fab' fragments by size exclusion high-performance chromatography and TLC were carried out. Equivalent results were obtained for radioimmunoconjugates when each was analyzed with both methods. Facile incorporation of Tc-94m into tumor-targeting Fab' antibody fragments will enable investigation of such agents for tumor-specific imaging using positron emission tomography. PMID- 10541335 TI - I-131 anti-B1 therapy/tracer uptake ratio using a new procedure for fusion of tracer images to computed tomography images. AB - In patients with non-Hodgkin's lymphoma being treated by I-131-radiolabeled anti B1 monoclonal antibody, we test the hypothesis that the activity taken up in tumors during therapy is the same as that observed during tracer evaluation, except for scaling by the ratio of administered activities. Chemotherapy-relapsed patients are imaged only with planar conjugate views, whereas previously untreated patients are imaged with planar conjugate views and with single-photon emission computed tomography (SPECT). The SPECT tracer activity quantification requires computed tomography (CT) to SPECT image fusion, for which we devised a new procedure: first, the tracer SPECT images are fused to the therapy SPECT images. Then, that transformation is combined with the therapy SPECT-to-CT transformation. We also use (a) the same volumes of interest defined on CT for both tracer and therapy image sets, and (b) a SPECT counts-to-activity conversion factor that adapts to background and rotation radius. We define R as the ratio of therapy activity percentage of infused dose over tracer activity percentage of infused dose at 2-3 days after monoclonal antibody infusion. For 31 chemotherapy relapsed patients, the R ratio for 60 solitary or composite tumors averages 0.931 +/- 0.031. The hypothesis of R being 1 is rejected with greater than 95% confidence. However, the difference from 1 is only 7.4%. The range of R is 0.43 1.55. For seven previously untreated patients, R averages 1.050 +/- 0.050 for 24 solitary tumors evaluated by SPECT. For six of these patients, R averages 0.946 +/- 0.098 for one of these solitary tumors and for five composite tumors, evaluated by conjugate views. Both results agree with the hypothesis that R is 1. The range of R for the SPECT tumors is 0.71 +/- 0.03 to 1.82 +/- 0.53, and for the conjugate view tumors, it is 0.70-1.35. Plots of R versus tumor volume yield small correlation coefficients. That from SPECT approaches a statistically significant difference from zero correlation (P = 0.06). In summary, on average, the tumor percentage of infused dose following tracer administration is predictive of therapeutic percentage of infused dose within 8%. For greater accuracy with individual tumors, however, an intratherapy evaluation is probably necessary because the range of R is large. PMID- 10541336 TI - Effect of 67Cu-2IT-BAT-Lym-1 therapy on BCL-2 gene and protein expression in a lymphoma mouse model. AB - Radioimmunotherapy using monoclonal antibodies against tumor-associated antigens has been particularly promising in the treatment of radiosensitive malignancies such as lymphoma. 67Cu has excellent physical and biochemical properties for radioimmunotherapy. 67Cu-2IT-BAT-Lym-1 has been used in preclinical and clinical trials, where an exceptionally long residence time of 67Cu on tumor was observed. BCL-2, a proto-oncogene that promotes cell survival by blocking apoptotic cell death, is overexpressed in most B-cell lymphomas including Raji human Burkitt's lymphoma cells. In this study, therapeutic efficacy and BCL-2 gene and protein expression levels were examined in Raji xenografts in mice after 67Cu-2IT-BAT-Lym 1 radioimmunotherapy. 67Cu-2IT-BAT-Lym-1 therapy induced a response rate (complete and partial responses) of approximately 50%. BCL-2 gene expression was decreased 3 h after radioimmunotherapy, followed by a decrease in Bcl-2 protein by 24 h. Decreases in BCL-2 gene and protein expression preceding observations of 67Cu-2IT-BAT-Lym-1 therapeutic effect suggest that down-regulation of BCL-2 leaves cells more likely to be killed by low dose-rate radiation from radioimmunotherapy. PMID- 10541337 TI - A method for patient-specific absorbed dose estimation for internal beta emitters. AB - The purpose of the study was to determine a technique for estimating patient specific absorbed radiation doses in radioimmunotherapy and other internal emitter therapies. Beta Radiation sources were considered, with 90Y being the radionuclide of primary interest. Organ uptake of activity was determined using a merged set of computed tomography and planar nuclear images. Estimation of local absorbed dose was accomplished using a voxel source kernel. Voxel size was 0.2 x 0.2 x 0.5 cm; dimensions were from the digital resolution of the nuclear and computed tomography data sets. Dose-volume histograms were also obtained due to the voxel nature of the estimations. Organ dose estimates were made for two patients receiving the chimeric anticarcinoembryonic antigen antibody cT84.66. Considerable variation was observed when comparing the voxel kernel results with medical internal radiation dosimetry values obtained via the MIRDOSE3 program. Primary uncertainty in the organ dose estimates was determined to be due to the variation in organ mass. By correcting the S values in that program by the organ mass ratio, we found generally good agreement between our method and MIRDOSE3. We conclude that patient-specific absorbed doses can be estimated for 90Y-labeled antibodies. PMID- 10541338 TI - Validation of an analytical expression for the absorbed dose from a spherical beta source geometry and its application to micrometastatic radionuclide therapy. AB - The purpose of this study was to validate an analytical expression for the absorbed-dose calculation from the spherical source of beta-emitting radionuclides and to apply it to micrometastases treated with radiolabeled monoclonal antibodies. The self-absorbed fractions from I-131 and P-32 uniform spherical sources were calculated using the analytical expression introduced by P. K. Leichner (J. Nucl. Med., 35: 1721-1729, 1994). The calculated absorbed fractions were compared with previously reported values and were found to be in reasonable agreement, with a maximum difference of 15% for smaller masses and a long-range beta emitter. The expression was subsequently applied to estimate the absorbed dose within spheroid models with nonuniform penetration of radiolabeled antibody. The corresponding absorbed dose for I-131 was compared with reported micro-thermoluminescence dosimeter measurements and found to be in good agreement. This work has independently substantiated the methodology outlined by Leichner and may be reliably incorporated into new software developments for radionuclide dosimetry treatment planning. PMID- 10541339 TI - Can current models explain the lack of liver complications in Y-90 microsphere therapy? AB - Normal liver complications have not been observed in Y-90 microsphere therapy of hepatic tumors [selective internal radiation (SIR)], despite clinical studies reporting estimated absorbed doses to normal liver between 100 and 150 Gy. The purpose of the study was to see whether predictions of normal tissue complication probability (NTCP) models for liver based on clinical data from external beam therapy are consistent with clinical results of SIR. Liver NTCP was calculated using a parallel architecture model and normal liver dose-volume histograms that have been proposed for SIR. A parallel model including internal functional subunit structure is also proposed. Dose rate effects are incorporated. A criterion for comparing model calculations with clinical data is presented. For the parallel architecture model, the predicted NTCP is sensitive to the dose distribution in normal liver and to the model parameters, particularly the repair time. With reasonable assumptions about the microsphere distribution, the parallel model with parameters deduced from external beam therapy outcome analysis is consistent with the observed lack of liver complications. Inclusion of FSU structure widens the range of assumptions under which consistency is found. The parallel model can be consistent with the clinically observed lack of liver complications in SIR. More information about the activity distribution and the radiobiology of normal liver under conditions typical of microsphere therapy should be sought. PMID- 10541340 TI - Studies on the red marrow dosimetry in radioimmunotherapy: an experimental investigation of factors influencing the radiation-induced myelotoxicity in therapy with beta-, Auger/conversion electron-, or alpha-emitters. AB - Usually, the red marrow (RM) is the first dose-limiting organ in radioimmunotherapy. However, several studies have obtained only poor correlations between the marrow doses and the resulting toxicities. Furthermore, RM doses are mostly not determined directly but are derived from blood doses by assuming a ratio that is, over time for the respective conjugates, more or less constant between blood and marrow activities. The aim of this study was to determine, in a mouse model, this RM:blood activity ratio for various immunoconjugates, to investigate whether there may be differences between complete IgG and its fragments with various labels ((125/131)I versus (111)In, (88/90)Y, or 213Bi), and to analyze, in more detail, factors other than just total dose, such as dose rate or relative biological effectiveness factors, that may influence the resulting myelotoxicity. The maximum tolerated activities (MTAs) and doses (MTDs) of several murine, chimeric, and humanized immunoconjugates as complete IgG or fragments (F(ab)2 and Fab), labeled with beta(-)-emitters (such as 131I or 90Y), Auger electron-emitters (such as 125I or (111)In), or alpha-emitters (such as 213Bi) were determined in nude mice. Blood counts were monitored at weekly intervals; bone marrow transplantation was performed to support the assumption of the RM as dose-limiting. The radiation dosimetry was derived from biodistribution data of the various conjugates, accounting for cross-organ radiation; besides the major organs, the activities in the blood and bone marrow (and bone) were determined over time. Whereas no significant differences were found for the RM:blood ratios between various IgG subtypes, different radiolabels or various time points, differences were found between IgG and bi- or monovalent fragments: typically, the RM:blood ratios were approximately 0.4 for IgG, 0.8 for F(ab')2, and 1.0 for Fab'. Nevertheless, at the respective MTAs, the RM doses differed significantly between the three conjugates: e.g., with 131I-labeled conjugates, the maximum tolerated activities were 260 microCi for IgG, 1200 microCi for F(ab)2, and 3 mCi for Fab, corresponding to blood doses of 17, 9, and 4 Gy, respectively. However, initial dose rates were 10 times higher with Fab as compared to IgG, and still 3 times higher as compared to F(ab)2; interestingly, all three deliver approximately 4 Gy within the first 24 h. The MTDs of all three conjugates were increased by BMT by approximately 30%. Similar observations were made for 90Y-conjugates. Higher RM doses were tolerated with Auger-emitters than with conventional beta(-)-emitters, whereas the MTDs were similar between alpha- and beta(-)-emitters. In accordance to dose rates never exceeding those occurring at the single injection MTA, two subsequent injections of two doses of 80% of the single shot MTA of 131I- or 90Y-labeled Fab' and two doses of 100% of the single shot MTA of 213Bi-labeled Fab' were tolerated without increased lethality, if administered 24-48 h apart. In contrast, reinjection of bivalent conjugates was not possible within 6 weeks. These data suggest that the RM:blood activity ratios differ between IgG and fragments, although there is no anatomical or physiological explanation for this phenomenon at this point. In contrast to the current opinion, indication for a strong influence of the dose rate (or dose per unit time), not only total dose, on the resulting toxicity is provided, whereas the influence of high-linear energy transfer (alpha and Auger/conversion electrons) versus low-linear energy transfer (beta and gamma) type radiation seems to be much lower than expected from previous in vitro data. The lower myelotoxicity of Auger-emitters is probably due to the short path length of their low-energy electrons, which cannot reach the nuclear DNA if the antibody is not internalized into the stem cells of the RM. PMID- 10541341 TI - A strategy to reduce red marrow dose for intraperitoneal radioimmunotherapy. AB - The aim of this study was to determine whether shorter-lived radionuclides can reduce red marrow (RM) toxicity for i.p. radioimmunotherapy (RIT). The potential radionuclides, which included Lu-177, I-131, Y-90, Re-186, Re-188, and Ho-166, were attached to antibody CC49. Each radiopharmaceutical was assumed to have identical in vivo pharmacokinetics. Blood and whole body retention data acquired from 26 patients who received i.p. RIT with Lu-177 CC49 were used as input. The average biological half-time of Lu-177 CC49 in the whole body was 280 h, and the average Lu-177 concentration in plasma increased to a maximum at 2 days postinfusion, followed by steady clearance. The residence time and RM doses were calculated for each radionuclide. In the current model, Re-188 was found to deliver the lowest RM dose, primarily because it had the shortest half-life, whereas Y-90 delivers the highest dose. Re-188 delivers 60% of the RM dose as compared with Lu-177 and can increase the dose to metastatic sites in the i.p. space by a similar factor. Based on limiting the RM dose to 200 cGy, the maximum administered activity of each radionuclide is as follows: (a) 106 mCi, Lu-177; (b) 58 mCi, I-131; (c) 34 mCi, Y-90; (d) 70 mCi, Re-186; (e) 169 mCi, Re-188; and (f) 110 mCi, Ho-166. Because of the delayed steady leakage of radiopharmaceuticals from the i.p. cavity to the plasma, short-lived radionuclides may offer special advantages for i.p. RIT. PMID- 10541343 TI - Ion exchange tumor targeting: a new approach. AB - Connective tissues are distinguished by the types, concentrations, and organizations of material in the extracellular matrix. Many physiological functions are determined largely by the nature and organization of the extracellular components. The components are characterized by their content and distribution of charged, mostly anionic groups. The distinct roles played by the charges are sometimes modeled by analogy to the transport theory of ion exchange resins. The intent of this study was to investigate whether the properties of the tumor matrix could be used for selective, charge-dependent accumulation of charge modified dextran. Ten patients with diagnosed superficial urinary bladder carcinoma were included in the study. They received intravesical instillations of technetium-99m-labeled charge-modified dextran derivatives (approximately 0.1-1 mg; approximately 50 MBq in saline; 30-min incubation). After treatment and resection, samples were taken from normal and diseased tissue. The result clearly demonstrated a charge-dependent difference in the quotient of radioactive uptake in tumor tissue: normal tissue. Instillations of cationic dextran yielded a high quotient, up to 3000. Normal tissue had background activity. Anionic dextran yielded a low quotient, 1.8-2, with increased background (i.e. uptake in normal tissue). Neutral dextran gave a quotient of up to 90. No radioactivity could be detected in blood. The tumors in this study apparently displayed cation exchanging properties. We will continue this investigation and determine whether this is a general property of bladder carcinomas and whether other carcinomas display ion exchange properties. If this is the case, the finding could have important implications for the local treatment of several cancers. PMID- 10541342 TI - Targeting strategies for cancer radiotherapy. AB - Novel strategies to increase the therapeutic ratio in clinical radioimmunotherapy studies are needed. Limitations to radioimmunotherapy include bone marrow suppression due to the long circulating half-life of radiolabeled monoclonal antibodies (mAbs) and heterogeneous tumor penetration of the high-molecular weight mAb. An approach to overcome these problems is the use of genetically engineered mAbs. The engineered mAb discussed in this paper contains a deletion in the constant region of the mAb that increases its tumor penetration and blood clearance compared with the intact mAb. Radiolabeling of this mAb should lead to a similar radiation-absorbed dose to tumor compared with the intact mAb, but reduce the radiation absorbed dose to bone marrow. In addition, low or variable expression of tumor-associated target antigens or receptors may lead to low or heterogeneous tumor uptake of radiolabeled mAbs. This report also discusses a novel approach toward systemic radiotherapy that combines gene transfer techniques (to increase tumor receptor expression) with radiolabeled peptides that target the induced receptor. The radiolabeled peptides achieve good tumor uptake, rapid tumor penetration, and rapid blood clearance. A humanized construct of the CC49 (HuCC49) high-affinity anti-TAG-72 mAb, as well as a construct with the CH2 region deleted (HuCC49deltaCH2), were labeled with 131I and 177Lu. Biodistribution of the radiolabeled constructs was evaluated 24 h after regional i.p. injection in athymic nude mice bearing i.p. LS174T human colon cancer xenografts. The 131I-HuCC49deltaCH2 showed a median tumor uptake of 5.5% ID/g which was similar to that of 131I-HuCC49 at 5.2% ID/g. However, the median blood concentration of 131I-HuCC49deltaCH2 was 0.2% ID/g which was significantly lower than 0.8% ID/g for 1311-HuCC49. The uptake of the constructs in other normal tissues were similar. The 177Lu-HuCC49deltaCH2 showed a median tumor uptake of 9.4% ID/g, which was slightly higher than that of 177Lu-HuCC49 at 7.9% ID/g. The median blood concentration of 177Lu-HuCC49deltaCH2 was 0.2% ID/g, which was significantly lower than 0.4% ID/g for 177Lu-HuCC49. The uptake of the antibody constructs in other normal tissues were similar except for the kidney. The tumor:blood ratios of 177Lu-HuCC49 and 177Lu-HuCC49deltaCH2 were 19.4 and 60.2, respectively, at 24 h after injection. The purpose of the second aspect of the study was to determine the biodistribution of 64Cu-1,4,8,11 tetraazacyclotetradecane-1,4,8,11-tetraacetic acid (TETA)-octreotide in a human ovarian cancer model induced to express human somatostatin receptor subtype 2 (SSTr2) using gene transfer techniques as a prelude to future therapy studies. Mice bearing i.p. SKOV3.ip1 tumors transduced with an adenoviral vector encoding the cDNA for SSTr2 (AdSSTr2) and injected i.p. with 64Cu-TETA-octreotide showed a median uptake of 24.3% ID/g in tumor at 4 h postinjection compared with 4.9% ID/g at 18 h after injection. Also, tumor uptake of 64Cu-TETA-octreotide at 4 h was not significantly different when administered either 2 or 4 days after injection of AdSSTr2 (P = 0.076). 64Cu-TETA-octreotide should be useful for targeted radiotherapy against tumors that are genetically induced to express high levels of SSTr. These two novel targeting strategies show promise for improved cancer radioimmunotherapy. PMID- 10541344 TI - Comparison of 125I- and (111)In-labeled monoclonal antibody BR96 for tumor targeting in combination with extracorporeal immunoadsorption. AB - Extracorporeal whole blood immunoadsorption (ECIA) accelerates the clearance of radiolabeled monoclonal antibodies (mAbs) without significantly affecting tumor uptake by removing the excess of these mAbs from the blood, thus increasing tumor:normal tissue (T:N) ratios. The present study is focused on comparing the capacity of ECIA in tumor targeting with the same mAb (chiBR96-biotin) labeled with either (111)In or 125I. Forty-five Brown Norwegian rats with syngeneic rat colon carcinoma isografted both in liver and intramuscularly were used. chiBR96 is a highly tumor-specific mAb directed against the Lewis-type antigen. ECIA of whole blood was started 15 h after the injection of 125I- or (111)In-labeled BR96 biotin. The procedure lasted for 2 h and was repeated for (111)In-labeled BR96 biotin in a few rats after 3 or 24 h. ECIA reduced the whole body activity by the same magnitude (between 39% and 52%), irrespective of whether (111)In- or 125I labeled chiBR96 was used. A similar observation was also made for the reduction in blood radioactivity after ECIA (79-94%). Time-activity curves during ECIA showed that the major reduction (approximately 85%) in blood radioactivity occurred during the first 45-60 min. Repeating the ECIA with (111)In-BR96 caused only an additional minimal reduction of blood activity, whereas a further reduction of whole body activity of 14-20% was achieved. The T:N uptake ratios were significantly enhanced immediately after ECIA with (111)In- or 125I-labeled chiBR96. Due to greater accumulation of (111)In-BR96 in tumors, a long-term improvement in T:N ratios was obtained after ECIA compared with 125I-labeled BR96. Our results therefore indicate that (111)In/(90Y)-labeled BR96-biotin could be more advantageous than 125I/131I for radioimmunotargeting/radioimmunotherapy in combination with ECIA due to better activity retention by the tumor. PMID- 10541345 TI - Epithelial mucin-1 (MUC1) expression and MA5 anti-MUC1 monoclonal antibody targeting in multiple myeloma. AB - Multiple myeloma (MM) is the second most common hematological cancer in the United States. It is typically incurable, even with myeloablative chemotherapy and stem-cell transplantation. The epithelial mucin-1 (MUC1) glycoprotein is expressed by normal and malignant epithelial cells but has also been shown to be expressed by MM cells. MUC1 is a useful antigenic target in solid tumors for clinical diagnostic and therapeutic monoclonal antibody (mAb)-based approaches. The MA5 mAb, as well as other anti-MUC1 mAbs reactive with the MUC1 variable number tandem repeat domain, exhibited moderate to strong reactivity with both MM cell lines and clinical samples. To explore the biochemical nature and potential of MUC1 as an antigenic target in MM, studies were performed to: (a) compare the mRNA and the MUC1 glycoprotein species between epithelial cancer and MM cell lines; and (b) develop and use a human MM tumor xenograft model system to study the biodistribution of the MA5 mAb. MA5 mAb was strongly reactive with six of eight human MM cell lines by flow cytometry. In seven of eight MM patient samples (bone marrow and/or peripheral blood) reactivity was found in 10-90% of the cells, whereas normal control (n = 5) and leukemia and lymphoma (n = 5) cells showed only 0-6% reactivity. 125I-labeled MA5 whole-cell binding studies showed quantitatively similar amounts of binding between strongly positive MM lines and high-MUC1-expressing breast carcinoma lines. mRNA expression was assessed by Northern blotting and reverse transcription-PCR. MM cell lines were positive by both methods, with strong similarity in the sizes of the mRNAs and cDNAs that were obtained. Finally, biodistribution experiments were carried out with 131I labeled MA5 versus a nonbinding control 125I-labeled mAb in a s.c. MM xenograft model. Selective MM tumor uptake of the MA5 mAb was demonstrated, with a potential for delivering a tumor radiation absorbed dose of 8540 cGy/mCi of injected dose compared with 3099 cGy/mCi of tumor-absorbed dose delivered by nonspecific antibody. PMID- 10541346 TI - Idiotypic-anti-idiotypic antibody interactions in experimental radioimmunotargeting. AB - Idiotypic-anti-idiotypic antibody interactions can be used in vivo to regulate the serum levels of specific radiolabeled antibodies. Anti-idiotypic antibodies can also be used as clearing agents for radiolabeled antibodies in radioimmunolocalization and radioimmunotherapy. The present study describes the immunochemical interactions between the monoclonal idiotype (H7) and three generated monoclonal anti-idiotypic antibodies (alphaH7:1, alphaH7:35, and alphaH7:38). An unexpected variability in complex formation could be demonstrated in vitro, revealing three different stable complex patterns, i.e., low molecular weight 1:1 complexes, ladder formation with oligomeric, consecutively added constituents, and large linear polymeric complexes of high molecular weight. Within 24 h, the anti-idiotypes were able to cause a significant decrease in total body radioactivity, and the antibody generating a ladder formation (alphaH7:38) was found to be the most efficient at removing radiolabeled idiotypes from the circulation. It is concluded that monoclonal anti-idiotypic antibodies may be valuable tools in improving radioimmunolocalization and radioimmunotargeting. PMID- 10541347 TI - Targeting human cancer xenografts with monoclonal antibodies labeled using radioiodinated, diethylenetriaminepentaacetic acid-appended peptides. AB - A new nonmetabolizable peptide approach to the production of residualizing radioiodine was evaluated in nude mice bearing xenografts of human lung adenocarcinoma (Calu-3) and B-cell lymphoma (Ramos). Monoclonal antibodies (MAbs) RS7 (anti-epithelial glycoprotein-1) and LL2 (anti-CD22) were radioiodinated using the thiol-reactive diethylenetriaminepentaacetic acid-D-peptide adducts IMP R1 and IMP-R2. 125I-IMP-R1- and 125I-IMP-R2-labeled MAbs were compared to the MAbs iodinated by the conventional chloramine-T approach, (111)In, and 131I dilactitoltyramine (DLT). In vivo biodistribution studies demonstrated a significant improvement in the tumor accretion of radiolabel using the 125I-IMP R1 labeled MAbs compared with the conventionally iodinated antibodies. For example, at day 7, the percentage of injected dose per gram of tissue in Calu-3 was 7.9 +/- 4.1% and 18.1 +/- 7.9% (P < 0.05) for the conventional 131I- and 125I IMP-R1-RS7, respectively, and tumor:nontumor ratios were 2.6-4.5-fold higher with the 125I-IMP-R1-RS7. It is estimated that 131I-IMP-R1-RS7 would deliver a dose to tumor (at the estimated maximum tolerated dose) 3.9 times greater than conventional 131I-labeled RS7, 1.4 times greater than 90Y-labeled RS7, and 0.7 times that of 131I-DLT-labeled RS7. Tumor accretion of 125I-IMP-R2-RS7 was also improved compared with conventionally iodinated antibody. However, this label also caused a large increase in kidney accretion. Similar improvements in tumor accretion and tumor:nontumor ratios were observed when 125I-IMP-R1-LL2 was used in the Ramos model. IMP-R1 offers a practical and useful residualizing radioiodine label because labeling efficiency is at least 10 times greater than that of the residualizing label DLT, without MAb aggregation. Structural modifications can be envisioned for further improvements in radioiodine incorporation, specific activity, and tumor dosimetry, and efforts along these lines are under way. PMID- 10541348 TI - Development of a hyperimmune anti-MUC-1 single chain antibody fragments phage display library for targeting breast cancer. AB - Radioimmunotherapy (RIT) has demonstrated potential for improving clinical cancer therapy. Optimizing the approach has proven difficult thus far. Antibody phage display libraries provide unique molecules that could improve RIT. A phage display library of single chain antibody fragments (scFv) against the MUC-1 mucin molecule, which is expressed on 90% of human breast cancers, was produced from the spleen cells of MUC-1 hyperimmunized BALB/c mice. Increased serum IgG levels, 15 times baseline, were detected following the third immunization. RNA from the spleen cells was isolated, cDNA was made, and variable heavy and variable light immunoglobulin chain gene regions were amplified using PCR technology. The variable heavy and variable light chain gene regions were combined with a flexible linker, ligated into the pCANTAB 5E phagemid vector, and electroporated into TG1 Escherichia coli cells. A library of 10(7) initial colonies was compiled. Forty-six of 288 colonies screened for reactivity demonstrated binding to MUC-1-expressing MCF-7 breast cancer cell membrane fragments. Anti-MUC-1 library diversity evaluated by BstNI digest demonstrated that 52% of the anti-MUC 1 scFv binding MCF-7 possessed individual banding patterns representative of approximately 5 x 10(5) colonies likely able to recognize distinct epitopes present on MUC-1 positive human breast cancers. In summary, the anti-MUC-1 scFv antibody phage library contains diverse scFv molecules, which should provide unique characteristics and epitope recognition. These molecules will be used in the development of pretargeting RIT strategies designed to improve the clinical outcome of patients with breast cancer. PMID- 10541349 TI - Humanization of Immu31, an alpha-fetoprotein-specific antibody. AB - Immu31 is a murine monoclonal antibody (Ab) specific for alpha-fetoprotein (AFP), a tumor-associated marker. The excellent tumor targeting ability of Immu31 has led to the development of a Immu31-based radioimmunodiagnostic agent, AFP-Scan, for hepatocellular carcinoma and other AFP-producing tumors. To enhance the capability of Immu31-based immunoconjugates being used in diagnostic and therapeutic procedures in humans, a humanized version of Immu31 (hImmu31) was constructed by grafting the complementarity determining regions (CDRs) of murine variable domains for the heavy (VH) and kappa (Vkappa) chain to the respective human VH and Vkappa framework regions (FRs). The cDNA encoding the VH and Vkappa of Immu31 was cloned by reverse transcription-PCR from hybridoma cells, and a chimeric Immu31 (cImmu31) composed of murine V and human C domains was constructed. Competitive ELISA assays showed identical AFP binding activity between the chimeric and murine Abs, confirming the authenticity of the cloned V genes. Based on sequence homology, the EU FR1, FR2, and FR3 and the NEWM FR4 were selected as the scaffold for grafting VH CDRs and REI FRs for Vkappa CDRs of Immu31. The amino acid residues in murine FRs that are considered to be in contact with the CDRs of the Ab were maintained in the humanized version. hImmu31, thus constructed and expressed, showed comparable immunoreactivity in a competitive binding ELISA assay to that of murine Immu31 and cImmu31. High-level production was achieved by expressing hImmu31 in a dhfr-based amplifiable system, and the productivity has exceeded 100 mg/liter in terminal cultures. PMID- 10541350 TI - Generation and monitoring of cell lines producing humanized antibodies. AB - Antibody humanization has eliminated or reduced the human antimouse antibody response associated with the administration of murine antibodies. We have successfully humanized three different antibodies: (a) hMN-3 (granulocyte targeting); (b) hMu-9 (colorectal cancer targeting); and (c) hWI2 (anti-idiotype to the anti-carcinoembryonic antigen antibody MN-14). All humanized antibodies demonstrated immunoreactivities comparable to their parent counterparts. Previously, we reported the generation of high productivity cell lines for hMN-14 and hLL2 using the amplifiable vector pdHL2. Through amplification, selection, and cloning procedures, cell lines capable of large scale production were established, and further enhancement of production was achieved by a fed perfusion bioreactor process. Using a similar and improved approach, we have enhanced the production of the above-mentioned humanized antibodies by gene amplification induced by a stepwise increase in the concentration of methotrexate in the culture media. A reliable IgG determination method is essential to monitor amplification, especially at the final cloning stage, for the selection of the subclones with the highest productivity. We found that measurement of humanized IgG concentration in culture media supplemented with more than 1 microM methotrexate by a standard ELISA assay could be unreliable and misleading. Whereas the determination of antibody by adsorption/elution on protein A from a 100-ml culture is accurate and reproducible, the method is time-consuming, tedious, and labor intensive. We have recently developed a Western blot assay that enables us to monitor the productivity of the cultures. The assay is simple and sensitive, and it makes simultaneous determinations of relative antibody production from individual clones at the 96-well stage feasible. With this method, amplification, cloning, and adaptation to serum-free conditions of multiple cell lines can be monitored in an efficient manner. PMID- 10541351 TI - The effects of domain deletion, glycosylation, and long IgG3 hinge on the biodistribution and serum stability properties of a humanized IgG1 immunoglobulin, hLL2, and its fragments. AB - Antibody (Ab) fragments are preferred agents for imaging applications because of their rapid clearance from the blood, thereby providing high tumor:blood ratios within a few hours. Several preclinical studies have also suggested that Ab fragments might be preferred for therapeutic applications over an intact IgG. The purpose of this project was to develop engineered Ab fragments using a humanized anti-carcinoembryonic antigen and anti-CD22 Ab as the parent. Three types of variants were prepared: a deltaCH2 (deletion mutant missing the CH2), a gamma3 F(ab')2 containing the human IgG3 hinge, and three glycosylated variants. The gamma3 F(ab')2 and glycosylated variants were developed because of the potential for site-specific linkage to the Ab in its divalent or monovalent fragment. The gamma3 F(ab')2 variant contains 10 cysteine residues that could be used for direct coupling using thiol chemistry, whereas the glycosylated variants have N linked glycosylation sites engineered in the CH1 domain (two variants) as well as the VK domain (one variant). All of these variants were successfully prepared and shown to react with the target antigen. All Abs could be purified to a single peak by size-exclusion HPLC, but the deltaCH2 variant showed two distinct peaks, which were believed to be both the divalent and monovalent forms of this fragment. The two CH1 glycosylated variants showed differences in the extent of glycosylation. Modeling studies suggest that one variant would be better suited for site-specific coupling than the other because the carbohydrate chain is extended further away from the antigen-binding site. The Abs were radioiodinated to determine their pharmacokinetic behavior in mice. All of the humanized Ab divalent fragments cleared nearly 20 times faster from the blood than the murine parent F(ab')2 over a 24-h period. The glycosylated fragments showed some added stability compared to the other fragments over 4 h, but by 24 h, they had cleared to the same extent. Size-exclusion high-performance liquid chromatography of blood samples indicated that the humanized Ab fragments were quickly degraded in the blood. Thus, there is an inherent instability of the divalent fragments from these humanized IgG1 constructs that may affect their utility in imaging or therapy applications. PMID- 10541352 TI - Generation and characterization of a single gene-encoded single-chain-tetravalent antitumor antibody. AB - Monoclonal antibody (mAb) CC49, a murine IgG1, reacts with the tumor-associated glycoprotein-72 expressed on a variety of carcinomas. In clinical trials, radiolabeled CC49 has shown excellent tumor localization to a variety of carcinomas. To minimize the immunogenicity of CC49 mAb in patients, a humanized CC49 (HuCC49) was generated by complementarity-determining region (CDR) grafting. The relative affinity of HuCC49 was 2-3-fold less than that of the murine mAb. With the aim of improving tumor targeting, attempts have been made to enhance the avidity of the HuCC49 mAb. Previous research has yielded a single gene-encoded immunoglobulin, SCIgcCC49deltaCH1, which is a dimer of a single chain consisting of CC49 single-chain Fv linked to the NH2 terminus of the human gamma1 Fc through the hinge region. This molecule is comparable to the mouse-human chimeric CC49 in terms of in vitro antigen binding properties, cytolytic activity, and rate of plasma clearance in athymic mice bearing human tumor xenografts. Recently, a single gene encoding a single-chain immunoglobulin consisting of a HuCC49 diabody attached to human gamma1 Fc via the hinge region was constructed. The diabody, a bivalent antigen-binding structure, is made up of variable heavy (V(H))/variable light (V(L)) domains and V(L)/V(H) domains. In each of the variable domain pairs, the V(H) and V(L) domains are linked through a short linker peptide. Meanwhile, the two pairs are linked via a 30-residue Gly-Ser linker peptide to yield two antigen-binding sites by lateral and noncovalent association of the V(L) of one pair with the V(H) of the other. Transfectomas expressing the single-gene immunoglobulin secrete a homodimer of about Mr 160,000 that reacts to tumor associated glycoprotein-72. This tetravalent humanized antitumor immunoglobulin molecule may potentially be an efficacious therapeutic and diagnostic reagent against a wide range of human carcinomas. PMID- 10541354 TI - The effect of various therapeutic solutions including colloidal chromic 32P via an intratumoral injection on the tumor physiological parameters of AsPC-1 human pancreatic tumor xenografts in nude mice. AB - To overcome the physiological barrier in solid tumors (i.e., tumor hypertension), a large volume of material is required via an intratumoral injection. Alternatively, a method of reduction in tumor hypertension is also feasible. In this study, we focused on the physiological response after an intratumoral infusion of various therapeutic agents. Tumor interstitial fluid pressure (TIFP) was intermittently monitored for up to 7 days after treatment using AsPC-1 human pancreatic tumors in nude mice. Macroaggregated albumin (MAA), colloidal chromic 32P (32P-CP), albumin, dexamethasone, 5-fluoro-2'deoxyuridine, dextrose, saline, and trypan blue increased TIFP within approximately 5 min, and TIFP returned to the original level within 1 h, except in the case of MAA and 32P-CP. We also found that the maximal uptake for AsPC-1 tumors in both the exponential and plateau growth phases occurred at approximately 100 min postincubation; the maximum value in the exponential growth phase was approximately 2 times less than that of plateau growth phase (P < 0.01). Therefore, this study supports intralesional 32P-CP brachytherapy for nonresectional pancreatic cancer patients. This may offer a promising treatment modality for delivering high doses of tumor selective radiation, mainly due to two physiological mechanisms: (a) the high adherence of 32P-CP to the infused regions; and (b) reduction in either tumor blood flow or TIFP by this therapeutic colloid. PMID- 10541353 TI - Cholecystokinin-B/gastrin receptor binding peptides: preclinical development and evaluation of their diagnostic and therapeutic potential. AB - The high sensitivity of pentagastrin stimulation in detecting primary or metastatic medullary thyroid cancer (MTC) suggests widespread expression of the corresponding receptor type on human MTC. Indeed, autoradiographic studies demonstrated cholecystokinin (CCK)-B/gastrin receptors not only in >90% of MTCs but in a high percentage of small cell lung cancers and potentially a variety of gastrointestinal adenocarcinomas. In a pilot study, we have demonstrated the feasibility of radiolabeled gastrin-I to target CCK-B receptor-expressing tissues in vivo in animals and patients (T. M. Behr et al., Eur. J. Nucl. Med., 25: 424 430, 1998). The aim of the present study was to systematically optimize, in a preclinical model, suitable radioligands for targeting CCK-B receptors in vivo. For this purpose, a variety of CCK/gastrin-related peptides, all having in common the COOH-terminal CCK-receptor binding tetrapeptide sequence Trp-Met-Asp-PheNH2 or derivatives thereof, were studied. They were radioiodinated by the Iodogen or Bolton-Hunter procedures. The peptides tested were members of the gastrin- or cholecystokinin families or possessed characteristics of both, which differ by the intramolecular position of a tyrosyl moiety (occurring in native or sulfated form). Their stability and affinity were studied in vitro and in vivo; their biodistribution and therapeutic efficacy were tested in nude mice bearing s.c. human MTC xenografts. Diethylene-triamine-pentaacetate derivatives of suitable peptides were synthesized, evaluated, and labeled with (111)In. All members of the CCK or gastrin family were stable in serum (with t(1/2)s of several hours at 37 degrees C); nevertheless, the stability of those peptides was highest that bore the NH2-terminal pGlu residues (e.g., big gastrin, gastrin-I, caerulein, and others) or D-amino acids. In accordance to their comparably low affinity, nonsulfated members of the CCK family showed fairly low uptake in the tumor and other CCK-B receptor-expressing tissues (e.g., the stomach). Sulfated CCK derivatives performed significantly better but additionally displayed a high uptake in normal, CCK-A receptor-expressing tissues (such as the liver/gallbladder, pancreas, and bowel). Best tumor uptake and tumor:nontumor ratios were obtained with members of the gastrin family, probably because of their selectivity and affinity for the CCK-B receptor subtype. Pilot therapy experiments in MTC bearing animals showed significant antitumor efficacy as compared with untreated controls. (111)In-Labeled diethylene-triamine pentaacetate derivatives of minigastrin showed excellent targeting of CCK-B receptor-expressing tissues in animals and a normal human volunteer. These data suggest that CCK/gastrin analogues may be a useful new class of receptor binding peptides for diagnosis and therapy of CCK-B receptor-expressing tumors, such as MTC or small cell lung cancer. Nonsulfated gastrin derivatives may be preferable because of their CCK-B receptor selectivity, and hence, lower accretion in normal CCK-A receptor-expressing organs. Further preclinical as well as clinical studies are ongoing. PMID- 10541355 TI - Retargeting interleukin 13 for radioimmunodetection and radioimmunotherapy of human high-grade gliomas. AB - A vast majority of patients with glioblastoma multiforme (GBM), a high-grade glioma, overexpress abundant amounts of a receptor for interleukin (IL)-13 in situ. This receptor is more restrictive because it is IL-4-independent and therefore differs from the IL-13/4 signaling receptor of normal tissue that is shared with IL-4. We previously identified one of the sites on the human IL (hIL) 13 molecule that is important for its interaction with the IL-13/4 receptor, a residue of glutamic acid at position 13. In this study, we mutated the cytokine and produced hIL-13.E13Y, in which the glutamic acid was substituted by tyrosine. This additional tyrosine residue was therefore strategically located within the region of IL-13 interaction with the signaling physiological receptor. hIL 13.E13Y did not transduce signals through the IL-13/4 receptor, whereas its interaction with the more restrictive, GBM-associated receptor remained intact. The mutated hIL-13 could be readily radiolabeled. Radiolabeled hIL-13.E13Y produced specific autoradiographic images of human GBM specimens. We demonstrate an effective way to redirect hIL-13 to its more restrictive receptor found in high-grade gliomas by mutagenizing the cytokine, and, concomitantly, we equipped hIL-13 with an additional tyrosine residue for higher specific activity radiolabeling. PMID- 10541356 TI - Internal radionuclide therapy of primary osteosarcoma in dogs, using 153Sm ethylene-diamino-tetramethylene-phosphonate (EDTMP). AB - Fifteen dogs were referred because of a spontaneous bone tumor, lameness, and local pain. The osteosarcoma diagnosis was established by clinical examination, X ray, bone scintigraphy, and histological examination of biopsy material. The tumors were located in the extremities (n = 12), scapula (n = 1), maxilla (n = 1), and the frontal bone (n = 1). The dogs were given one to four i.v. injections of 153Sm-labeled ethylene-diamino-tetramethylene-phosphonate (153Sm-EDTMP; 36-57 MBq/kg body weight). Three dogs had surgery in addition to the radionuclide treatment. Platelet and WBC counts showed a moderate and transient decrease. No other toxicity was observed. Average tumor doses after a single injection were approximately 20 Gy, considerably higher in some areas because of inhomogeneous uptake. Macroscopically distant metastases were detected in seven dogs at autopsy. One dog died from an intercurrent disease, free of cancer, 5 months after the radionuclide treatment. None of the dogs was cured. The median and mean survival times from the first treatment to death or euthanasia were 150 and 252 days, respectively. Nine of the dogs had obvious pain relief, and five of them seemed pain-free: one for 20 months and one for 48 months. It is concluded that high tumor doses may be deposited in dog osteosarcomas by 153Sm-EDTMP, and the ratio between tumor dose and the dose to surrounding tissues is favorable. The treatment gives pain relief and in some cases tumor growth delay. In combination with surgery, 153Sm-EDTMP may prolong life significantly and possibly cure the disease because the development of metastases are seemingly postponed. No serious side effects were observed. PMID- 10541357 TI - Comparison of multiple bolus and continuous injections of 131I-labeled CC49 for therapy in a colon cancer xenograft model. AB - One of the problems in achieving cures with radioimmunotherapy is that hematological toxicity limits the quantity of radiolabeled monoclonal antibody (MAb) that can be administered. The MAb CC49 binds with high affinity to the TAG 72 antigen expressed in many human adenocarcinomas. We investigated tumor growth inhibition, survival, and tumor and bone marrow dosimetry after multiple bolus injections or continuous infusion of 131I-labeled CC49 MAb in a human colon cancer xenograft model to determine which method of administration results in the highest therapeutic ratio. Groups of athymic nude mice bearing established s.c. LS174T human colon cancer xenografts received three i.p. bolus injections (3X) of 131I-labeled CC49 (3X, days 0, 3, and 7) or were implanted i.p. with mini-osmotic pumps delivering 131I-labeled CC49 over 7 days. The total radionuclide doses administered were broken down into low-dose (< or = 450 microCi), medium-dose (450-800 microCi), and high-dose (> 800 microCi) groups. At the medium-dose level, the bolus-therapy animals did not have a significantly longer survival time but did have a significantly longer time-to-tumor doubling than the pump therapy animals. The median survival for medium-dose bolus and pump therapy was 157 and 105 days, respectively, and the median time-to-tumor doubling was at least 114 and 77 days, respectively. At the low-dose level, the bolus-therapy animals had a significantly longer survival time but not a significantly longer time-to-tumor doubling than the pump-therapy animals. The median survival for low dose bolus and pump therapy was 95.5 and 59 days, respectively, and the median time-to-tumor doubling was 73 and 38 days, respectively. The high-bolus dose was toxic. A comparison of the overall survival rate of pump therapy versus bolus therapy, excluding high-dose, resulted in the bolus-therapy animals having a longer survival time and a longer time-to-tumor doubling than the pump-therapy animals. Serial section autoradiography was used to reconstruct tumor activity density distributions over time. Average dose values calculated from total uptake data for 900 microCi administered activity yielded 158 Gy (3X) and 141 Gy (pump). Average three-dimensional doses using the radial histograms to calculate the absorbed fractions were 139 Gy and 123 Gy, respectively. This calculation includes energy loss external to the tumor. With cell proliferation parameters set to single fraction 60Co recurrence results, the effective dose (D(eff)) for local control was 11 Gy and 9 Gy, respectively. Three bolus injections resulted in a more uniform dose rate over a longer period, resulting in a calculated 19% improvement in D(eff) compared with pump administration. Dose to bone marrow was calculated assuming an activity concentration in bone marrow of 0.24 times the concentration in blood and an absorbed fraction of 0.63. For the 900-microCi 131I labeled CC49 injected activity, pump administration resulted in an 80% higher calculated D(eff) to bone marrow compared with 3X bolus injection. These results demonstrate that 3X bolus injections were clearly superior to pump administration in terms of survival, tumor growth inhibition, tumor absorbed dose, and bone marrow dose. PMID- 10541358 TI - Radioimmunotherapy using vascular targeted 213Bi: the role of tumor necrosis factor alpha in the development of pulmonary fibrosis. AB - A monoclonal antibody (201B) specific to murine thrombomodulin, covalently linked to cyclohexyl diethylenetriaminepentaacetic acid, successfully delivers chelated 213Bi, an alpha-particle emitter, (213Bi-201B) rapidly to lung vascular endothelium. When injected at doses of 1 MBq/mouse, 213Bi-201B destroyed most of the 100 colonies of EMT-6 mammary carcinomas growing as lung tumors of up to 2000 cells/colony. Some mice were cured of lung tumors, and others had extended life spans compared to untreated control animals but eventually succumbed to tumor recurrence. At injected doses of 4-6 MBq/mouse, 100% of lung tumor colonies were eliminated; however, 3-4 months later, these mice developed pulmonary fibrosis and died. The mechanisms leading to the fibrotic response in other pulmonary irradiation models strongly implicate tumor necrosis factor alpha (TNF-alpha), released from damaged tissues, as the pivotal inflammatory cytokine in a cascade of events that culminate in fibrosis. Attempts to prevent the development of pulmonary fibrosis, by using antibodies or soluble receptor (rhuTNFR:Fc) as inhibitors of TNF-alpha, were unsuccessful. Additionally, mice genetically deficient for TNF-alpha production developed pulmonary fibrosis following 213Bi 201B treatment. Interestingly, non-tumor-bearing BALB/c mice receiving rhuTNFR:Fc or mice genetically deficient in TNF-alpha production and treated with 213Bi 201B, had significantly reduced life spans compared to mice receiving no treatment or 213Bi-201B alone. We speculate that in normal mice, although TNF alpha may induce an inflammatory response following alpha-particle radiation mediated tumor clearance and pulmonary damage, its effects in the post-tumor clearance time period may actually retard the development of fibrosis. PMID- 10541359 TI - Validation of 213Bi-alpha radioimmunotherapy for multiple myeloma. AB - The efficacy of radioimmunotherapy (RIT) with beta emitters has been clinically demonstrated in the treatment of refractory forms of lymphoma. Alpha-emitting radionuclides with a short half-life are also good potential candidates for RIT directed at tumor targets easily accessible to radioimmunoconjugate molecules and small enough to benefit from the short range of alpha particles (<100 microm). The purpose of this study was to demonstrate the feasibility of ex vivo purging of multiple myeloma-invaded bone marrow. Tumor cells were targeted by a specific monoclonal antibody (B-B4) coupled to 213Bi by a chelating agent (pentaacetic triamine diethylene p-aminobenzyl acid). The efficacy of alpha-RIT was assessed in vitro by analysis of thymidine incorporation, cell mortality, apoptosis of myeloma cells, and the study of nonspecific irradiation of hematopoietic cell lines not recognized by B-B4-pentaacetic triamine diethylene p-aminobenzyl acid immunoconjugate. High dose-dependent cell mortality of myeloma cells was found with radiolabeled B-B4, and this mortality was total at 30 kBq/10(5) cells. Cells were found in apoptotic state at rates of up to 40% for a dose of 7.4 kBq/10(5) cells. Nonspecific mortality was low compared with specific mortality (up to 1%). PMID- 10541360 TI - Growth and characterization of LNCaP prostate cancer cell spheroids. AB - Cells from the prostate tumor cell line LNCaP have been grown as spheroids. The growth kinetics of the spheroids have been characterized by fitting a Gompertz equation to spheroid growth curves. The proliferation state of cells within spheroids of different diameters was assessed by bromodeoxyuridine staining. Scanning and electron transmission microscopy were performed to determine the ultrastructure of the spheroids. Prostate-specific antigen (PSA) secretion was monitored throughout spheroid growth. Consistent with Gompertzian kinetics, the volume of LNCaP spheroids initially increased exponentially and then reached a plateau. The doubling time during the exponential phase was 29 +/- 4 h. A core of nonproliferating cells was seen in spheroids with a diameter of 400 microm; at a diameter of 600 microm, a necrotic core had formed. In smaller, 200-microm diameter spheroids, a core of nonproliferating cells was not seen, but proliferating cells were concentrated at the spheroid periphery. Electron microscopy showed that the spheroids were enveloped by an extracellular matrix and that cell adhesion within the spheroids was due in part to desmosomes. PSA secretion by the spheroids could be modeled as originating from a spherical shell whose thickness was independent of overall spheroid diameter. The shell thickness obtained by fitting an appropriate equation to the data was consistent with that determined from the bromodeoxyuridine studies. LNCaP cells exhibit several important features of prostate cancer cells; in vivo, they are androgen responsive, and they express prostatic acid phosphatase, PSA, and prostate specific membrane antigen. LNCaP spheroids provide a simple but relevant model for the study of drug delivery and response in prostate cancer. PMID- 10541361 TI - Immunohistology of carcinoembryonic antigen (CEA)-expressing tumors grafted in nude mice after radioimmunotherapy with 131I-labeled bivalent hapten and anti-CEA x antihapten bispecific antibody. AB - We have developed a pretargeting strategy, called the Affinity Enhancement System (AES), which uses bispecific antibodies (BsF(ab')2) to target radiolabeled bivalent haptens to tumor cells. We performed several radioimmunotherapy (RIT) experiments in nude mice grafted with LS174T colon carcinoma or TT medullary thyroid cancer. Mice were treated with 131I-labeled di-DTPA-indium-tyrosyl-lysine bivalent hapten (75-112 MBq) administered 15-48 h after anti-CEA x anti-DTPA indium BsF(ab')2. Immunohistological studies were performed on tumors at their minimal relative volume (TT), on stabilized tumor nodules (LS174T), and on regrowing tumors (TT and LS174T). Untreated tumors were used as controls. On microscopic examination, regrowing tumors (2 months posttherapy) were similar to untreated tumors with cells showing their respective typical morphology (large cells with a high nucleocytoplasmic ratio for TT, small and very undifferentiated cells for LS174T). However, regrowing tumors showed larger necrotic areas and a higher mitotic index correlated with Ki-67 antigen staining. Immunostaining for CEA was as strong as for controls. By contrast, the immunohistology of TT tumors at their minimal relative volume (1 month posttherapy) or of LS174T residual nodules (8 months posttherapy) showed decreased mitotic indices correlated with poor Ki-67 antigen staining. Some clusters of LS174T presented with features of glandular lumen, which suggested a more differentiated and less aggressive status. In TT tumors, CEA expression remained unchanged (80-100% membrane and cytoplasmic staining), whereas only 70% of the LS174T tumors were stained, with 58% loss of the membrane expression. Repeated treatment early after the tumor has reached its minimal relative volume should thus be efficient and improve the overall efficacy of AES RIT. PMID- 10541362 TI - Toxicity and efficacy of radioimmunotherapy in carcinoembryonic antigen-producing medullary thyroid cancer xenograft: comparison of iodine 131-labeled F(ab')2 and pretargeted bivalent hapten and evaluation of repeated injections. AB - This study compared the toxicity and efficacy of 131I-labeled bivalent hapten pretargeted by anti-carcinoembryonic antigen (CEA)/anti-N alpha (diethylenetriamine-N,N,N',N''-tetraacetic acid-indium(F6-734) bispecific antibody [affinity enhancement system (AES) reagents] with 131I-labeled anti-CEA F(ab')2 (131I-F6) in mice grafted with a human medullary thyroid carcinoma. Repeated injections of AES reagents were also evaluated. Mice bearing TT tumor xenografts were treated with 37, 74, or 92.5 MBq of AES reagents, two injections of 74 MBq of AES reagents 45 days apart, or 37 or 92.5 MBq of 131I-F6. Control groups were treated with nonspecific 131I-labeled F(ab')2, nonspecific AES reagents, nonradiolabeled F6, F6-734 bispecific antibody, and nonradiolabeled bivalent hapten or received no injection. For AES treatments, bispecific antibody was injected 48 h before the hapten. Animal weight, hematological toxicity, tumor volume, and serum thyrocalcitonin were monitored during 5 months. At 92.5 MBq, weight loss was significantly lower after AES than F6 treatment (P = 0.004). The percentages of leukocyte count changes were significantly lower after AES than F6 at 37 and 92.5 MBq (P = 0.01 and 0.04, respectively). The percentage of platelet count changes was significantly lower with AES at the 92.5-MBq dose level (P = 0.04). In the group injected twice with AES reagents, toxicity was not significantly increased after the second treatment. Tumor response was observed in all cases but was significantly longer with repeated treatments of 74 MBq AES reagents than with a single treatment (P = 0.004). Two complete responses were observed with repeated treatments. Changes in thyrocacitonin level paralleled those in tumor volume. These results indicate that pretargeted radioimmunotherapy was at least as efficient as one-step radioimmunotherapy and markedly less toxic. Repeated treatments with AES reagents increased efficacy without increasing toxicity. PMID- 10541363 TI - Radioimmunotherapy in medullary thyroid cancer using bispecific antibody and iodine 131-labeled bivalent hapten: preliminary results of a phase I/II clinical trial. AB - The toxicity and therapeutic efficacy of escalating doses of anti carcinoembryonic antigen x anti-N alpha-(diethylenetriamine-N,N,N',N'' tetraacetic acid)-In bispecific monoclonal antibody (F6-734) and iodine 131 labeled bivalent hapten were determined in a Phase I/II trial. A total of 26 patients with recurrences of medullary thyroid cancer documented by imaging and a rise in serum thyrocalcitonin were enrolled. Twenty to 50 mg of F6-734 and 40-100 mCi of 131I-hapten were injected 4 days apart. Quantitative scintigraphy was performed after the second injection for dosimetry estimations in eight cases. Clinical, biological, and morphological follow-up was carried out for 1 year after treatment. The mean percentage of injected activity per gram of tumor at the time of maximum uptake was 0.08% (range, 0.003-0.26%). The tumor biological half-life ranged from 3 to 95 days, and tumor doses ranged from 2.91 to 184 cGy/mCi. The estimated tumor-to-nontumor dose ratios were 43.8 x 53.4, 29.6 x 35.3, 10.9 x 13.6, and 8.4 x 10.0 for total body, red marrow, liver, and kidney, respectively. Grade III/IV hematological toxicity was observed in seven patients, most of them with bone metastases. Among the 17 evaluable patients, 4 pain reliefs, 5 minor tumor responses, and 4 biological responses with decrease of thyrocalcitonin were observed. Nine patients developed human anti-mouse antibody. Dose-limiting toxicity was hematological, and maximum tolerated activity was 48 mCi/m2 in this group of patients, most of whom had suspected bone marrow involvement. The therapeutic responses observed in patients mainly with a small tumor burden are encouraging for the performance of a Phase II trial with minimal residual disease. PMID- 10541364 TI - Assessment of combined radioimmunotherapy and chemotherapy for treatment of medullary thyroid cancer. AB - We have shown previously significant antitumor effects using 90Y-MN-14 anti-CEA monoclonal antibody (MAb) for radioimmunotherapy (RAIT) of human medullary thyroid cancer (MTC) xenografts using the TT cell line. The purpose of this investigation was to determine the effect of combining chemotherapy and RAIT with 90Y-MN-14 in MTC. In particular, the toxicity and efficacy of various dose schedules of RAIT and doxorubicin were examined and compared with that at the maximum tolerated dose (MTD) of each single modality treatment. The MTD of RAIT of 105 microCi of 90Y-MN-14 was given alone and combined with 100 and 75% of the MTD of doxorubicin (60 mg/m2); and the MTD of doxorubicin was given alone and combined with 100 and 75% of the MTD of RAIT. In addition, 75% of each agent was also administered in combination. The MTD of RAIT was also evaluated in combination with 58 and 78% of the MTD of Taxol. Whereas 90Y-MN-14 (105 microCi) led to significant antitumor effects (P < 0.0001), doxorubicin at 60 mg/m2 or Taxol at 225 mg/m2 yielded only a slight tumor growth delay. The combinations of 100% of the MTD of RAIT and 75% of the MTD of doxorubicin and 100% of the MTD of doxorubicin and 75% of the MTD of RAIT were equitoxic to the MTD of RAIT alone and appear to result in improved efficacy compared with either RAIT or doxorubicin alone. For the 100% RAIT and 75% doxorubicin combination, the therapeutic efficacy was similar when doxorubicin was administered on the same day or 1 day after RAIT, but the treatment was less effective when doxorubicin was administered 2 days after RAIT (P < 0.03). Prolonged retardation of tumor progression was also observed in animals treated with the MTD of RAIT combined with 175 mg/m2 of Taxol, without increases in toxicity above that observed with RAIT alone. In conclusion, the combination of RAIT and chemotherapy appears to augment the antitumor effects of either treatment alone without a significant increase in toxicity. In addition, the timing of drug administration relative to RAIT in the combined therapy appears to be important. PMID- 10541365 TI - Maximum tolerated dose and large tumor radioimmunotherapy studies of 64Cu-labeled monoclonal antibody 1A3 in a colon cancer model. AB - The purpose of this study was 2-fold: to determine the maximum tolerated dose (MTD) of 64Cu-bromoacetamidobenzyl- 1,4,8,11-tetraazacyclotetradecane N,N',N'',N'''-tetraacetic acid (BAT)-2-iminothiolane (2IT)-monoclonal antibody (MAb) 1A3 in hamsters, and second, to determine the therapeutic efficacy of 64Cu BAT-2IT-MAb 1A3 at various dose levels in hamsters with large (600 mg), 7-day-old GW39 human colorectal carcinoma tumors. In the MTD studies, non-tumor-bearing hamsters were injected with varying amounts of Cu-64-BAT-2IT MAb 1A3 (>10 mCi) normalized to mCi injected/kg of hamster body weight. Results indicated that the MTD was 150 mCi of Cu-64/kg of body weight. Hamsters receiving higher doses (170 190 mCi/kg) lost greater than 20% of their body weight, and all died between 8 and 13 days (n = 3). All hamsters receiving doses < or = 150 mCi/kg (120-150 mCi; n = 13) survived to the experimental end point (6 weeks) with an overall gain in weight. WBC and platelet counts were depressed in all animals 7 days after treatment but returned to normal values in the survivors by 2 weeks. For larger tumor therapy studies, 40% (8 of 20) of hamsters receiving a single dose of 7.0 mCi and 62.5% (5 of 8) of hamsters receiving 15 mCi of Cu-64-BAT-2IT-MAb 1A3 remained tumor free 4 months after treatment. In dose fractionation studies, hamsters received two 3.5 mCi doses separated by 24 or 48 h with 44% (4 of 9) and 25% (2 of 8) survival, respectively. In every large tumor experimental group, 100% of animals experienced tumor growth inhibition compared to saline control animals. Together, the MTD and the large tumor therapy studies confirm that 64Cu labeled agents are excellent candidates for radioimmunotherapy trials. PMID- 10541366 TI - Antibody phage libraries for the next generation of tumor targeting radioimmunotherapeutics. AB - Pretargeting techniques have shown promise for enhancement of the therapeutic index of radioimmunotherapy for cancer. However, methods to vary and compare antibody configurations and select optimal combinations have proved rather formidable. New options for the construction of pretargeting molecules are provided by sophisticated use of the diversity and malleability of antibody genes. Diverse arrays of single-chain antibody fragments (scFvs) can now be obtained reactive with virtually any target antigen by selection from human naive phage antibody libraries. ScFvs can also be cloned directly from hybridoma for construction of phage libraries that facilitate subsequent manipulation: e.g., affinity maturation and modification of specificity. ScFvs affinity selected from these sources to their specific antigen targets have demonstrated a wide spectrum of binding characteristics. ScFvs selected from a large human naive phage antibody library by binding Cu-1,4,8,11-tetra-azacyclotetradecane-N,N',N'',N''' tetraacetic acid (TETA) or Y-1,4,7,10-tetra-azacyclododecane-N,N',N'',N''' tetraacetic acid (DOTA) have shown diversity by DNA fingerprints. DNA sequence information confirmed that the anti-TETA scFv represented diverse scFv gene families. ScFvs for Y-DOTA and those for lymphoma-associated HLA DR10 (Lym-1) were selected in a similar manner from mouse antibody gene libraries derived from hybridoma. ScFv clones for each of these antigens were chosen for further study based on the results of ELISA assays involving the respective cell membrane or metal chelate antigens. A PCR primer system built to pCANTAB 5E expression vector sequence was designed to facilitate cloning of antibody heavy (V(H)) and light (V(L)) genes from selected scFvs as cassettes into diabody modules. Thus, chosen scFvs could be expressed in the same diabody format for comparative study. Selected mouse anti-DOTA scFv and Lym-1 scFv genes were linked as V(HA) anti-DOTA link-V(LB) Lym-1; V(HB) anti-DOTA-link-V(LA) Lym-1 and ligated into the pCANTAB 5E vector. Corresponding diabodies were expressed in Escherichia coli and purified by affinity chromatography. Here we provide a perspective on the power of antibody phage libraries and the possibilities of creating simple molecular formats that can be used en route to the development of new tumor targeting and pretargeting molecules. PMID- 10541367 TI - Strategies for developing effective radioimmunotherapy for solid tumors. AB - Single-agent radioimmunotherapy (RIT) has proven efficacy as a treatment for hematological malignancies, particularly non-Hodgkin's lymphoma. Although promising, RIT has been less effective for solid tumors, in part because they are less radiosensitive. Bone marrow transplantation permits the administration of larger radiopharmaceutical doses, but the results of bone marrow transplantation supported RIT for solid tumors have been marginal. The purpose of this publication is to provide an overview of promising RIT strategies for solid tumors. It is apparent that combination therapy is required, but optimization of the radiopharmaceutical should be the first step. Metallic radionuclides provide higher tumor radiation doses but not necessarily an improved therapeutic index, that is, the ratio of tumor:normal tissue radiation doses. Biodegradable peptide linkers between the chelated metal and the antibody improve the therapeutic index. Further improvements depend on identification of synergistic therapies which recognize that: (a) continuous, low-dose radionuclide therapy acts through apoptosis; and (b) apoptosis is often blocked because most tumors have ineffective p53 and increased Bcl-2. Taxanes are particularly attractive as synergistic agents for RIT because they induce cell cycle arrest in the radiosensitive G2-M phase and p53-independent apoptosis. Optimal sequence and timing for combined modality RIT are critical to achieve synergy. Data from preclinical and clinical studies will be reviewed to illustrate the potential of these strategies. PMID- 10541368 TI - Initial clinical experience evaluating Yttrium-90-chimeric T84.66 anticarcinoembryonic antigen antibody and autologous hematopoietic stem cell support in patients with carcinoembryonic antigen-producing metastatic breast cancer. AB - cT84.66 is a human/murine IgG1 with high affinity and specificity for carcinoembryonic antigen (CEA). An earlier Phase I trial defined the maximum tolerated dose for 90Y-diethylenetriaminepentaacetic acid (DTPA)-cT84.66 at 22 mCi/m2. Dose-limiting toxicities were reversible leukopenia and thrombocytopenia. The purpose of this Phase I trial was to evaluate the feasibility and toxicities of administering higher activities of 90Y-DTPA-cT84.66 with stem cell support in patients with CEA-producing breast cancer. Patients with CEA-producing breast cancer refractory to standard therapies underwent peripheral stem cell collection followed by infusion of 111indium-DTPA-cT84.66. Those patients demonstrating tumor targeting received a single therapy dose of 90Y-DTPA-cT84.66, followed by Ca-DTPA infusion for 72 h posttherapy. Stem cells were reinfused following a divided schedule. To date, seven patients have been accrued to this trial. Each patient received an imaging dose of (111)In-cT84.66. Six patients demonstrated tumor imaging and received a single cycle of 90Y-cT84.66 at 15 mCi/m2 (three patients) and 22.5 mCi/m2 (three patients). One patient did not demonstrate tumor imaging and was not treated. At these administered activities, 90Y-cT84.66 was well tolerated. No dose-limiting toxicities have been observed. All patients demonstrated hematopoietic recovery after stem cell infusion. One patient demonstrated stable disease for 4 months; one patient had stable disease and reduction of bone pain for 3 months; and a third patient experienced >50% reduction of an ovarian metastasis, resolution of malignant pleural effusion, stable pleural metastases, and stable bone scan for 14 months. Preliminary results from this ongoing Phase I trial are promising and demonstrate the feasibility and potential for antitumor effects of stem cell supported 90Y cT84.66 therapy in patients with CEA-producing breast cancers. PMID- 10541370 TI - Dosimetry-based therapy in metastatic breast cancer patients using 90Y monoclonal antibody 170H.82 with autologous stem cell support and cyclosporin A. AB - Radioimmunoconjugates of 170H.82 (m170), a panadenocarcinoma monoclonal antibody, are effective for imaging primary and metastatic breast cancer. To evaluate m170 as a targeting agent for therapy, we developed (111)In- and 90Y-2-iminothiolane-2 [p-(bromoacetamido)benzyl]-1,4,7,10 tetraazacyclododecane-N,N',N'',N''' tetraacetic acid-m170 immunoconjugates with 99% purity by molecular sieving and immunoreactivity comparable to unmodified antibody. (111)In-m170 pharmacokinetic studies were performed prior to each therapy to determine the maximum dose of 90Y m170 that could be administered without exceeding a limit of 800 rad to the liver, lungs, or kidneys or 250 rad to the whole body or bone marrow for each of three cycles of treatment. Peripheral blood stem cells (PBSCs) were harvested and cyclosporin A (5 mg/kg twice daily) was administered as strategies to ameliorate myelosuppression and prevent the development of HAMA, respectively. An (111)In imaging/pharmacokinetic study was performed, and the 90Y dose was calculated and administered. The liver was the 90Y dose-limiting organ. The mean and range of calculated doses (in rad/mCi) for the five patients evaluated were as follows: whole body, 2.3 (2.1-2.4); liver, 17.8 (12.7-22.2); lung, 6.4 (4.8-7.2); kidney, 6.9 (6.3-11.5); marrow, 3.6 (1.9-4.4); and tumors (n = 25), 71.5 (14.1-141.5). Of the three patients treated, with doses of 37, 54, and 57 mCi of 90Y, one had a partial response, one had measurable tumor reduction but less than a partial response, and one had stable disease for more than 1 month. PBSCs prevented prolonged myelosuppression. The therapeutic responses, coupled with an absence, thus far, of significant adverse sequelae, suggest that this dosimetry-based approach combined with PBSCs may lead to effective therapy when higher 90Y doses are reached. PMID- 10541371 TI - A phase II study of intraperitoneal radioimmunotherapy with iodine-131-labeled monoclonal antibody OC-125 in patients with residual ovarian carcinoma. AB - Standard treatment of advanced ovarian cancer is a combination of surgery and chemotherapy. Additional therapies using the i.p. route are considered as a potential means of improving the locoregional control rate. This Phase II study evaluated the efficacy of i.p. radioimmunotherapy (RIT) in patients with minimal residual ovarian adenocarcinoma after primary treatment with surgery and chemotherapy. Between February 1995 and March 1996, six patients with residual macroscopic (<5 mm) or microscopic disease as demonstrated by laparotomy and multiple biopsies received i.p. RIT. All had initial stage III epithelial carcinoma and were treated with debulking surgery and one line (four patients) or two lines (two patients) of chemotherapy. RIT was performed with 60 mg of OC 125 F(ab')2 monoclonal antibody labeled with 4.44 GBq (120 mCi) of 131I injected 5-10 days after the surgical procedure. Systematic laparoscopy or laparotomy with multiple biopsies performed 3 months after RIT in five patients (clinical progression was seen in one patient) showed no change in three patients and progression in two patients. Toxicity was mainly hematological, with grade III neutropenia and thrombocytopenia in two patients. Human antimouse antibody production was demonstrated in all six patients. This study showed little therapeutic benefit from i.p. RIT in patients with residual ovarian carcinoma. PMID- 10541369 TI - Radioimmunotherapy of small volume disease of colorectal cancer metastatic to the liver: preclinical evaluation in comparison to standard chemotherapy and initial results of a phase I clinical study. AB - At the time of surgery, occult metastases (micrometastases) are present in more than 50% of colorectal cancer patients, and the liver is the most frequent site of apparent metastatic disease. Frequently, adjuvant chemotherapy is unable to prevent tumor recurrence. Thus, novel therapeutic strategies are warranted. The aim of this study was to establish a model of human colon cancer metastatic to the liver of nude mice, to assess, in this setting, the therapeutic efficacy of radioimmunotherapy (RAIT) compared to standard chemotherapy and to evaluate, in a Phase I/II trial, the toxicity and therapeutic efficacy of RAIT in colorectal cancer patients with small volume disease metastatic to the liver. Multiple liver metastases of the human colon cancer cell line GW-39 were induced by intrasplenic injection of a 10% tumor cell suspension. Whereas controls were left untreated, therapy was initiated on day 10 or 20 after tumor inoculation with the 131I labeled, low affinity anticarcinoembryonic antigen (anti-CEA) monoclonal antibody (MAb), F023C5 (Ka = 10(7) liters/mol), or the high-affinity anti-CEA MAb, MN-14 (Ka = 10(9) liters/mol), or chemotherapy (5-fluorouracil/leucovorin (folinic acid) versus irinotecan) at their respective maximum tolerated doses (MTDs). Twelve colorectal cancer patients with small volume disease metastatic to the liver (all lesions < or = 2.5 cm) were entered into a mCi/m2-based Phase I dose escalation study with 131I-labeled humanized version of MN-14, hMN-14. The patients were given single injections, starting at 50 mCi/m2 and escalating in 10 mCi/m2 increments. The MTD was defined as the dose level at which < or = 1 of 6 patients develop grade 4 myelotoxicity. In the mice, untreated controls died from rapidly progressing hepatic metastases at 6-8 weeks after tumor inoculation. The life span of mice treated with 5-fluorouracil/leucovorin was prolonged for only 1 3 weeks, whereas irinotecan led to a 5-8-week prolongation. In contrast, at their respective MTDs, the 131I-labeled low-affinity anti-CEA MAb, F023C5, led to a 20% permanent cure rate, and the high affinity MAb, MN-14, led to an 80% permanent cure rate, when therapy was initiated at 10 days after tumor inoculation. In the 20-day-old tumor stage, although it prolonged life, 131I-F023C5 was unable to achieve cures, whereas 131I-MN-14 was still successful in 20%. Histologically, no remaining viable tumor cells could be demonstrated in these animals surviving > 6 months. In patients, the MTD was reached at 60 mCi/m2 of hMN-14 (at 70 mCi/m2, two of three grade 4 myelotoxicities). Of 11 assessable patients, 2 had partial remissions (corresponding to an objective response rate of 18%), and 5 (45%) had minor/mixed responses or experienced stabilization of previously rapidly progressing disease. These data suggest that in small volume disease, RAIT may be superior to conventional chemotherapy. Antibodies of higher affinity seem to be clearly superior. The clinical response rates in patients with small volume disease are encouraging, being comparable to the response rates of conventional chemotherapeutic regimens but with fewer side effects. Ongoing studies will show whether treatment at the MTD will further improve therapeutic results. PMID- 10541373 TI - Radioimmunotherapy of small cell lung carcinoma with the two-step method using a bispecific anti-carcinoembryonic antigen/anti-diethylenetriaminepentaacetic acid (DTPA) antibody and iodine-131 Di-DTPA hapten: results of a phase I/II trial. AB - As small cell lung carcinoma (SCLC) is frequently a widespread disease at diagnosis, highly radiosensitive and often only partially responsive to chemotherapy, radioimmunotherapy (RIT) would appear to be a promising technique for treatment. We report the preliminary results of a Phase I/II trial of RIT in SCLC using a two-step method and a myeloablative protocol with circulating stem cells transplantation. Fourteen patients with proved SCLC relapse after chemotherapy were treated with RIT. They were first injected i.v. with a bispecific (anti-carcinoembryonic antigen/anti-diethylenetriaminepentaacetic acid) monoclonal antibody (20-80 mg in 100 ml of saline solution) and then 4 days later with di-(In-diethylenetriaminepentaacetic acid)-tyrosyl-lysine hapten labeled with 1.48-6.66 GBq (40-180 mCi) of I-131 and diluted in 100 ml of saline solution. In patients receiving 150 mCi or more, circulating stem cells were harvested before treatment and reinfused 10-15 days later. Treatment response was evaluated by CT and biochemical data during the month before and 1, 3, 6, and 12 months after treatment. All patients received the scheduled dose without immediate adverse reactions to bispecific antibody or 1-131 hapten. Toxicity was mainly hematological, with two cases of grade 2 leukopenia and three cases of grade 3 or 4 thrombopenia. Body scanning 8 days after injection of the radiolabeled hapten generally showed good uptake at the tumor sites. Estimated tumor dose was 2.6-32.2 cGy/mCi. Among the 12 patients evaluated to date, we have observed 9 progressions, 2 partial responses (one almost complete for 3 months), and 1 stabilization of more than 24 months. Efficiency and toxicity were dose related. The maximal tolerable dose without hematological rescue was 150 mCi. These preliminary results are encouraging, and dose escalation is currently continuing to reach 300 mCi. RIT should prove to be an interesting therapeutic method for SCLC, although repeated injections and hematological rescue will probably be required, as well as combination with other treatment modalities. PMID- 10541372 TI - Phase II study of interferon-enhanced 131I-labeled high affinity CC49 monoclonal antibody therapy in patients with metastatic prostate cancer. AB - Adjuvant Interferon (IFN) was given to increase tumor antigen expression and enhance localization with 131I-labeled CC49 radioimmunotherapy in a Phase II trial for hormone resistant metastatic prostate cancer. Patients received four doses of alpha-IFN (3 x 10(6) IU) s.c. on alternate days, from day -5 to day +1 of 75 mCi/m2 131I-CC49 treatment. Toxicity was well tolerated, with the majority of patients experiencing transient grade 3 or 4 neutropenia and/or thrombocytopenia (maximal at 4-6 weeks). The absorbed dose was >25 Gy in four of eight tumors visualized, which represents an increase of >20 fold over whole body radiation dose. Two patients had radiographic minor responses by 6 weeks post therapy, whereas five of six patients experiencing pain had symptom relief without radiographic changes. The protocol provided modest antitumor effects (pain relief in five of six patients and two minor radiographic responses). This study suggests that the addition of IFN enhanced tumor uptake and antitumor effects as compared to a prior Phase II trial of 131I-CC49 alone. PMID- 10541374 TI - Phase I radioimmunotherapy of metastatic renal cell carcinoma with 131I-labeled chimeric monoclonal antibody G250. AB - Clinical tumor targeting studies with chimeric monoclonal antibody G250 (cG250) in renal cell carcinoma (RCC) patients indicated the potential use of this antibody for radioimmunotherapy. Here we report on a phase I activity dose escalation study to determine the safety, the maximum tolerable dose (MTD), and the possible therapeutic potential of 131I-labeled cG250 in patients with progressive metastatic RCC. All patients (n = 12) received a diagnostic i.v. infusion of 5 mg of cG250 labeled with 222 MBq of 131I. If accumulation of the antibody in metastatic lesions was observed, patients were hospitalized and a second, therapeutic, i.v. infusion of 5 mg of cG250 labeled with a high dose of 131I was administered (n = 8). Three patients per dose level were entered, starting at 1665 MBq/m2. If no dose-limiting toxicity occurred, the study continued at the next dose level (555 MBq/m2 increase). Most patients experienced mild nausea without vomiting. No other complaints were reported during hospitalization. In two of two patients who received a dose of 2775 MBq/m2, grade IV hematological toxicity was observed, which was defined as dose limiting. Thus, the MTD was set at 2220 MBq/m2. In one patient (2220 MBq/m2), stable disease (lasting 3-6 months) was achieved, whereas another patient (2220 MBq/m2) showed a partial response that is ongoing (>9 months). The minor responses observed in this phase I trial in patients with an advanced stage of RCC are encouraging and warrant further study in a phase II setting at the MTD to determine the efficacy of radioimmunotherapy for metastatic RCC. PMID- 10541375 TI - Loco-regional radioimmunotherapy of high-grade malignant gliomas using specific monoclonal antibodies labeled with 90Y: a phase I study. AB - A Phase I radioimmunotherapy trial was conducted in which radioconjugated monoclonal antibody (MAb) was directly infused into the tumor or postoperative tumoral bed in patients with high-grade malignant glioma. BC-4, a murine MAb that recognizes tenascin, was used in these studies. The MAb was labeled with 90Y, a pure beta emitter with maximum energy of 2.284 MeV, which can penetrate into tissue up to 0.5-0.7 cm. Stable 90Y-labeled MAb conjugates were prepared using the chelator p-isothiocyanatobenzyl derivative of diethylenetriaminepentaacetic acid (ITC-Bz-DTPA), obtaining >95% labeling efficiency and conserving the antibodies' immunoreactivity (>85%). Twenty patients, 2 with anaplastic astrocytoma and 18 with glioblastoma, were included in the study. All of the patients had been treated previously with conventional therapies (surgery, external radiotherapy, and chemotherapy) and presented with progressive disease not amenable to further treatment. A dose-escalation study was performed using doses ranging from 5-30 mCi (185-1110 MBq) of 90Y-labeled MAb BC-4. The protein dose of MAb was always 1 mg. Three patients were treated at the 5, 10, 15, and 20 mCi levels, and the 25- and 30-mCi doses were each administered to 4 patients. Systemic toxicity was completely absent in all of the patients. The maximum tolerated dose to the brain was 25 mCi (925 MBq). The average dose to the tumor was 3200 cGy/mCi. Doses to the liver, bone marrow, and kidneys were below 10 cGy/mCi in all of the cases. Biodistribution studies demonstrated that the 90Y labeled MAb accreted exclusively in the neoplastic area without any diffusion into the normal brain or other normal organs. No clinical responses were recorded because of the very advanced stage of disease at the time of radioimmunotherapy. PMID- 10541376 TI - Radioimmunotherapy of relapsed non-Hodgkin's lymphoma with zevalin, a 90Y-labeled anti-CD20 monoclonal antibody. AB - Approximately 55,400 new cases of non-Hodgkin's lymphoma (NHL) are diagnosed each year, with the overall prevalence of the disease now estimated to be 243,000. Until recently, treatment alternatives for advanced disease included chemotherapy with or without external beam radiation. Based on the results of several clinical trials, the chimeric monoclonal antibody Rituximab has now been approved by the United States Food and Drug Administration as a treatment for patients with relapsed or refractory, low-grade or follicular, B-cell NHL. Several other monoclonal antibodies in conjugated and unconjugated forms have been evaluated in the treatment of NHL. Ibritumomab, the murine counterpart to Rituximab, radiolabeled with 90Y (Zevalin), is presently being evaluated in clinical trials. The success of radioimmunotherapy is dependent upon the appropriate choice of antibody, isotope, and chelator-linker. The Ibritumomab antibody targets the CD20 antigen. The antibody is covalently bound to the chelator-linker tiuxetan (MX DTPA), which tightly chelates the isotope 90Y. To date, two Phase I/II Zevalin clinical trials have been completed in patients with low-grade, intermediate grade, and mantle cell NHL. The overall response rate was 64% in the first trial and 67% in the later trial. Phase II and III trials are ongoing. PMID- 10541377 TI - Radioimmunotherapy using 131I-labeled anti-CD22 monoclonal antibody (LL2) in patients with previously treated B-cell lymphomas. AB - Experience in using rapidly internalizing antibodies, such as the anti-CD22 antibody, for radioimmunotherapy of B-cell lymphomas is still limited. The present study was conducted to assess the efficacy and toxicity of a 131I-labeled anti-CD22 monoclonal antibody (mAb), LL2, in patients with B-cell lymphomas failing first- or second-line chemotherapy. Eligible patients were required to have measurable disease, less than 25% B cells in unseparated bone marrow, and an uptake of 99mTc-labeled LL2Fab' in at least one lymphoma lesion on immunoscintigram. Eight of nine patients examined with immunoscintigraphy were unequivocally found to have an uptake, and therapy with 131I-labeled anti-CD22 [1330 MBq/m2 (36 mCi/m2)] preceded by 20 mg of naked anti-CD22 mAb was administered. Three patients achieved partial remission (duration, 12, 3, and 2 months), and one patient with progressive lymphoma showed stable disease for 17 months. Four patients exhibited progressive disease. The toxicity was hematological. Patients with subnormal counts of neutrophils or platelets before therapy seemed to be more at risk for hematological side effects. Radioimmunotherapy in patients with B-cell lymphomas using 131I-labeled mouse anti-CD22 can induce objective remission in patients with aggressive as well as indolent lymphomas who have failed prior chemotherapy. PMID- 10541378 TI - Pharmacokinetics, dosimetry, and initial therapeutic results with 131I- and (111)In-/90Y-labeled humanized LL2 anti-CD22 monoclonal antibody in patients with relapsed, refractory non-Hodgkin's lymphoma. AB - The pharmacokinetics, dosimetry, and immunogenicity of 131I- and (111)In-/90Y humanized LL2 (hLL2) anti-CD22 monoclonal antibodies were determined in patients with recurrent non-Hodgkin's lymphoma. Fourteen patients received tracer doses of 131I-hLL2 followed 1 week later by therapeutic doses intended to deliver 50-100 cGy to the bone marrow. Another eight patients received (111)In-hLL2 followed by therapy with 90Y-hLL2 also delivering 50 or 100 cGy to the bone marrow. The blood T(1/2) (hours) for the tracer infusions of 131I-hLL2 was 44.2 +/- 10.9 (mean +/- SD) compared with 54.2 +/- 25.0 for the therapy infusions, whereas the values were 70.7 +/- 17.6 for (111)In-hLL2 and 65.8 +/- 15.0 for 90Y-hLL2. The estimated average radiation dose from 131I-hLL2 in tumors >3 cm was 2.4 +/- 1.9 cGy/mCi and was only 0.9-, 1.0-, 1.1-, and 1.0-fold that of the bone marrow, lung, liver, and kidney, respectively. In contrast, the estimated average radiation dose from 90Y hLL2 in tumors >3 cm was 21.5 +/- 10.0 cGy/mCi and was 3.7-, 2.5-, 1.8-, and 2.5 fold that of the bone marrow, lung, liver, and kidney, respectively. No evidence of significant anti-hLL2 antibodies was seen in any of the patients. Myelosuppression was the only dose-limiting toxicity and was greater in patients who had prior high-dose chemotherapy. Objective tumor responses were seen in 2 of 13 and 2 of 7 patients given 131I-hLL2 or 90Y-hLL2, respectively. In conclusion, 90Y-hLL2 results in a more favorable tumor dosimetry compared with 131I-hLL2. This finding, combined with the initial anti-tumor effects observed, encourage further studies of this agent in therapeutic trials. PMID- 10541379 TI - Low- versus high-dose radioimmunotherapy with humanized anti-CD22 or chimeric anti-CD20 antibodies in a broad spectrum of B cell-associated malignancies. AB - Both CD22 and CD20 have been used successfully as target molecules for radioimmunotherapy (RAIT) of low-grade B cell non-Hodgkin's lymphoma. Because both CD20 and CD22 are highly expressed relatively early in the course of B cell maturation, and because its expression is maintained up to relatively mature stages, we studied the potential of the humanized anti-CD22 antibody, hLL2, as well as of the chimeric anti-CD20 (chCD20) antibody, rituximab (IDEC-C2B8), for low- or high-dose (myeloablative) RAIT of a broad range of B cell-associated hematological malignancies. A total of 10 patients with chemorefractory malignant neoplasms of B cell origin were studied with diagnostic (n = 5) and/or potentially therapeutic doses (n = 9) of hLL2 (n = 4; 0.5 mg/kg, 8-315 mCi of 131I) or chCD20 (n = 5; 2.5 mg/kg, 15-495 mCi of 131I). The diagnostic doses were given to establish the patients' eligibility for RAIT and to estimate the individual radiation dosimetry. One patient suffered of Waldenstrom's macroglobulinemia, eight patients had low- (n = 4), intermediate- (n = 2) or high (n = 2) grade non-Hodgkin's lymphoma, and one patient had a chemorefractory acute lymphatic leukemia, after having failed five heterologous bone marrow or stem cell transplantations. Three of these 10 patients were scheduled for treatment with conventional (30-63 mCi, cumulated doses of up to 90 mCi of 131I) and 7 with potentially myeloablative (225-495 mCi of 131I) activities of 131I labeled hLL2 or chCD20 (0.5 and 2.5 mg/kg, respectively); homologous (n = 6) or heterologous (n = 1) stem cell support was provided in these cases. Good tumor targeting was observed in all diagnostic as well as posttherapeutic scans of all patients. In myeloablative therapies, the therapeutic activities were calculated based on the diagnostic radiation dosimetry, aiming at lung and kidney doses < or = 20 Gy. Stem cells were reinfused when the whole-body activity retention fell below 20 mCi. In eight assessable patients, five had complete remissions, two experienced partial remissions (corresponding to an overall response rate of 87%), and one (low-dose) patient had progressive disease despite therapy. In the five assessable, actually stem-cell grafted patients, the complete response rate was 100%. Both CD20 and CD22 seem to be suitable target molecules for high-dose RAIT in a broad spectrum of hematological malignancies of B cell origin with a broad range of maturation stages (from acute lymphatic leukemia to Waldenstrom's macroglobulinemia). The better therapeutic outcome of patients undergoing high dose, myeloablative RAIT favors this treatment concept over conventional, low dose regimens. PMID- 10541380 TI - Pharmacokinetics of radiolabeled polyclonal antiferritin in patients with Hodgkin's disease. AB - The objective was to identify pharmacokinetic parameters predictive for tumor response and normal tissue side effects after i.v. administered radiolabeled rabbit antihuman ferritin IgG. Twenty-eight patients with recurrent Hodgkin's disease received 2 mg of rabbit antihuman ferritin i.v., labeled with 4-7 mCi of In-111 followed by two doses of 0.25, one dose of 0.3, or one dose of 0.4 mCi of Y-90-labeled antiferritin per kg of body weight 1 week later. Radioactivity and HPLC measurements of blood and urine samples and liver and tumor volumes identified on sequential whole-body scans provided the data for a pharmacokinetic analysis covering the first 6 days after the administration of the radioimmunoconjugate. Side effects and tumor response were recorded. Temporary hematological toxicity was noted in all patients. Sixteen patients showed a tumor response. The Y-90 blood level at 1 h after administration correlated with the severity of subsequent hematological toxicity. The rapid blood elimination half life of radioactivity was 4.4 h. Less than 5% of the administered radioactivity was eliminated in the first 24 h urine. The slow blood elimination half-life was 44 and 37 h for In-111 and Y-90, respectively. One of 12 retreated patients produced anti-rabbit IgG antibodies. The volume of distribution was larger for Y 90 than for In-111-labeled antiferritin (160 versus 110% of estimated blood volume). Accidentally extravasated rabbit IgG was rapidly catabolized in perivascular tissues with an effective half-life of less than 35 h. Slower catabolism was noted for rabbit IgG in blood (t(1/2) = 40 h), liver (t(1/2) = 62 h) or tumor (t(1/2) = 40-80 h). Twelve of 13 patients with an effective tumor half-life > 57 h showed a tumor response. I.v. administered polyclonal rabbit antihuman ferritin, labeled with In-111 or Y-90 is stable in vivo and targets Hodgkin's disease. Intravascular Y-90 causes a vascular leak and a larger volume of distribution for antiferritin. Elevated Y-90 blood levels at 1 h and a tumor half-life of >57 h predict for hematological toxicity and tumor response, respectively. PMID- 10541381 TI - Fractionated radiolabeled antiferritin therapy for patients with recurrent Hodgkin's disease. AB - The objective of this study was to determine the therapeutic ratio of fractionated radiolabeled immunoglobulin therapy (RIT) for patients with recurrent Hodgkin's disease. Ninety patients with recurrent Hodgkin's disease received 2 mg of yttrium-90-labeled polyclonal rabbit antihuman ferritin IgG i.v. Fifty-seven patients received a single (unfractionated) administration per treatment cycle; 11 of them were treated with 0.3 mCi/kg body weight, 39 were treated with 0.4 mCi/kg body weight, and 7 received 0.5 mCi/kg body weight per treatment cycle. Thirty-three patients had their radiolabeled immunoglobulin administration separated (fractionated) in 2 x 0.25 mCi/kg body weight (total activity, 0.5 mCi/kg). The interval between fractions was 1 week. Radioimmunoconjugates did not cause serious acute side effects. In vivo radioimmunoconjugates were stable. Human antirabbit IgG antibodies were found in 2 of 50 retreated patients (<5%). Hematological toxicity was the only side effect noted in all patients, and it was usually temporary. Response rates (RRs) were 20%, 61%, and 86% after 0.3, 0.4, or 0.5 mCi/kg unfractionated yttrium-90-labeled antiferritin. The RR for patients treated with fractionated RIT was 42%. In the fractionated RIT group, complete responses were decreased, and progressive disease increased (P < 0.05). Complete responses had a medium duration of 6 months. Median survival times were 390 days for 1 x 0.4 mCi/kg and 300 days for the 2 x 0.25 mCi/kg patient group. Fractionation did not provide the expected decrease in hematological toxicity or the expected increase in tumor RRs. PMID- 10541383 TI - Importance of timing of radioimmunotherapy after granulocyte colony-stimulating factor administration for peripheral blood stem cell harvest. AB - Nine radioimmunotherapy (RAIT)-naive patients with medullary thyroid cancer received high doses of 131I-MN-14 F(ab)2 anti-carcinoembryonic antigen monoclonal antibody (232-486 mCi), five in combination with bone marrow harvest, without prior granulocyte colony stimulating factor (G-CSF) injections (group 1) and the other four using peripheral blood stem cell harvest (PBSCH) preceded by G-CSF administration of 10 microg/kg per day for 5 days for stem cell mobilization, 6-8 days before RAIT (group 2). The amounts of radioactivity (mCi) given in both groups were similar (312 +/- 93 versus 424 +/- 65; P = NS). The percent platelet loss at nadir, duration of grade 4 thrombocytopenia, and time to complete recovery (TTCR, measured from the day of treatment), were 83 +/- 17%, 2.5 +/- 0.7 days, and 45 +/- 8 days in group 1, respectively, compared with 88 +/- 6%, 3.0 +/ 2.6 days, and 50 +/- 24 days in group 2 (P = NS), respectively. In contrast, the percent WBC loss at nadir, duration of grade 4 leukopenia, and TTCR of WBCs were 72 +/- 12%, 0.0 +/- 0.0 day, and 42 +/- 12 days in group 1, respectively, compared with 93 +/- 3%, 8.0 +/- 3.6 days, and 263 +/- 136 days in group 2, respectively (P < 0.02, 0.03, and 0.05 for differences of percent loss, duration of nadir, and TTCR, respectively). The difference in WBC toxicity after RAIT with bone marrow harvest and PBSCH is thought to be due to the administration of G-CSF for stem cell mobilization within 1 week before RAIT, which may sensitize the "endogenous" granulocyte precursors to subsequent RAIT. Preclinical data of RAIT in mice showed that the time of G-CSF administration before RAIT is critical: increased WBC toxicity was seen in mice given RAIT 3 or 7 days after a 5-day course of G-CSF (81 and 57% WBC loss, respectively) compared with those given no G-CSF or G-CSF 10 or 14 days before RAIT (45-50%) WBC loss). In conclusion, our data indicate that the timing of RAIT after the administration of G-CSF for PBSCH may influence WBC toxicity and recovery after this treatment and may have important implications on the design of high-dose RAIT trials combined with PBSCH. PMID- 10541382 TI - 67Copper-2-iminothiolane-6-[p-(bromoacetamido)benzyl-TETA-Lym-1 for radioimmunotherapy of non-Hodgkin's lymphoma. AB - Copper-67 (67Cu) has ideal properties for radioimmunotherapy. The 62-h half-life is similar to the residence time of antibodies in tumor, and the therapeutic beta emission of 67Cu is comparable to that of 131I. 67Cu, however, has gamma emissions similar to 99mtechnetium that are favorable for imaging. The macrocyclic chelating agent 1,4,7,11-tetraazacyclotetradecane-N,N',N'',N''' tetraacetic acid (TETA) binds 67Cu tightly and selectively, facilitating linkage to Lym-1, a mouse monoclonal antibody that preferentially targets malignant lymphocytes. The safety, efficacy, and practicality of 67Cu-2-iminothiolane (2IT) 6-[p-(bromoacetamido)benzyl]-TETA (BAT)-Lym-1 was assessed in this Phase I/II clinical trial for patients with non-Hodgkin's lymphoma (NHL) who had failed standard therapy. Up to four doses of 67Cu-2IT-BAT-Lym-1, 25 or 50-60 mCi/m2/dose (0.93 or 1.85-2.22 GBq/m2/dose, respectively) were administered; the lower dosage was used when NHL was detected in the bone marrow. 67Cu-2IT-BAT-Lym-1 provided good imaging of NHL, had favorable radiation dosimetry, and had a response rate of 58% (7 of 12). Hematological toxicity was dose-limiting, but no significant nonhematological toxicity was observed. The ability to image and treat NHL patients with a single radiopharmaceutical with useful physical properties makes 67Cu-labeled monoclonal antibody an option for future clinical trials, as this study showed that 67Cu-2IT-BAT-Lym-1 was safe, effective, and practical. PMID- 10541384 TI - The role of magnetic resonance imaging on spinal trauma. AB - Spinal MR has an increasingly important role in the assessment of spinal trauma. The ability to visualise clearly the spinal cord, nerve roots, ligaments, intervertebral discs and adjacent vascular structures allow a more accurate assessment of the extent of injury, and necessity for further management and provide a prognosis for recovery. PMID- 10541385 TI - The value of ultrasound and aspiration in differentiating vaso-occlusive crisis and osteomyelitis in sickle cell disease patients. AB - We have studied 50 patients with sickle cell disease who presented with musculoskeletal pain over a 2-year period to assess the use of ultrasound in differentiating infection from infarction. All the patients were evaluated by ultrasound. Five had soft tissue oedema and no fluid collection adjacent to the bone. Forty-five had a subperiosteal fluid collection. Twelve patients whose collections were not aspirated were diagnosed according to clinical evaluation. The remaining 33 patients underwent aspiration under ultrasound guidance to distinguish between an infection and infarction. Twenty-three of these were diagnosed as osteomyelitis and 10 as vaso-occlusive crises. In 21 out of the 23 infected cases, the fluid collection was greater than 10 mm at its thickest point perpendicular to the bone surface and all those with infarction had fluid less than 10 mm thickness. Aspiration under ultrasound guidance is a useful method to differentiate the two clinical entities. In patients suffering from osteomyelitis, identification of the organisms guides antibiotic administration. Needle decompression can help to relieve pain in osteomyelitis and vaso-occlusive crisis. PMID- 10541386 TI - Intrapulmonary lymph nodes: CT and pathological features. AB - Intrapulmonary lymph nodes are not uncommon and may be seen frequently in high quality computed tomography (CT) images and chest radiographs. We report four patients, older than 55 years, who had a long history of heavy smoking. Four intrapulmonary lymph nodes were located in the subpleural region (within 3 mm of the visceral pleural surface) of the right or left lower lobes. The lymph nodes were ovoid or round, homogeneous, well-defined and ranged from 9 to 10 mm in diameter. In one case, coexistent small nodules in the same or in other lobes on initial CT studies increased slightly in size over the following 3 years. All nodules contained lymphoid follicles and anthrocotic pigment, and in one case adjacent small aggregates of lymphocytes along interlobular septa were seen. Intrapulmonary lymph nodes have non-specific CT and clinical features. Follow-up CT may be useful in patients with suspected intrapulmonary lymph nodes. PMID- 10541387 TI - Mammographic features of ductal carcinoma in situ (DCIS) present on previous mammography. AB - AIM: To review previous mammograms of women found later to have DCIS and identify features which may have been missed or misinterpreted as benign. METHODS: The previous mammograms of 50 women who developed DCIS were analysed. The mammographic features at diagnosis and on the prior mammograms were compared. RESULTS: 11 (22%) of the previous mammograms were in retrospect abnormal; 5 (45%) of these had previously been assessed for the abnormality. All showed microcalcification. The following features were commoner at diagnosis than on previous films; rod shaped calcification (64 vs. 27%, P = 0.03) and a ductal distribution of calcification (76 vs. 45%, P = 0.05). Predominantly punctate calcification (64 vs. 12%, P = 0.001) and less than 10 calcifications in the cluster (54 vs. 24%, P = 0.05) were more common on the previous films. No difference was found in the frequency of granular calcification, branching calcification, irregularity in density, size or shape of calcification between the two groups. CONCLUSION: Features of DCIS missed on previous mammography include small cluster size, less than 10 calcifications in the cluster, the absence of rod shaped calcifications, the absence of a ductal distribution and the presence of predominantly punctate calcification. Features frequently seen both at diagnosis and on previous films which might have allowed earlier diagnosis were granular calcifications which vary in size, density and shape in an irregularly shaped cluster. Focal clustered calcification deserves aggressive investigation. PMID- 10541388 TI - The CT appearance of pleural and extrapleural disease in lymphoma. AB - OBJECTIVE: Pleural effusions in patients with lymphoma that are assumed to be related to malignancy are attributed to either lymphatic obstruction by tumour with resultant decreased clearance of pleural fluid, or direct tumour involvement of the pleura. The purpose of our study was to determine how often pleural or extrapleural disease was detected by computed tomography (CT) of patients with pleural effusions and primary or recurrent lymphoma. METHODS AND MATERIALS: We reviewed CT examinations showing evidence of pleural effusion in 61 patients with a diagnosis of primary or recurrent lymphoma and no history of other systemic disorders, including infection. The study population consisted of patients with non-Hodgkin's lymphoma (n = 44) or Hodgkin's disease (n = 17); both primary disease (n = 11) and recurrent disease (n = 50) were represented. Each CT examination was evaluated for the presence of disease involving the visceral and parietal pleura and extrapleural space, mediastinal adenopathy, and pulmonary parenchymal disease. RESULTS: Fourteen patients (23%) (nine with non-Hodgkin's lymphoma and five with Hodgkin's disease) had parietal pleural disease (thickening or nodules). Eighteen patients (30%) (14 with non-Hodgkin's lymphoma, four with Hodgkin's disease) had tumour or enlarged lymph nodes in the extrapleural space. Forty-three patients (70%) had mediastinal lymphadenopathy. Patients who received intravenous contrast did not have evidence of visceral pleural abnormalities or underlying pulmonary parenchymal disease. CONCLUSION: Forty-one percent of the patients with lymphoma and pleural effusions had CT evidence of pleural and/or extrapleural disease. The majority of the patients with extrapleural disease had adjacent posterior mediastinal disease. PMID- 10541389 TI - Breath-hold 3D gadolinium-enhanced subtraction MRA in the detection of transplant renal artery stenosis. AB - AIMS: The purpose of this study was to assess the value of breath-hold 3D gadolinium-enhanced subtraction magnetic resonance angiography (GD-MRA) in the detection of transplant renal artery stenosis (TRAS). PATIENTS AND METHODS: Seven patients with suspected post-transplant renal artery stenosis were studied. GD MRA was performed at 1.5T with a 3D fast spoiled gradient recalled echo (FSPGR) pulse sequence. Before injection of contrast medium, the 3D pulse sequence was performed to obtain a set of non-contrast images for subtraction purposes. Dynamic 3D imaging was performed simultaneously with the bolus injection of 40 ml of gadopentetate dimeglumine. Angiographic images were reconstructed using the Advantage Window workstation (version 2.0 GE Medical Systems) and subtraction was made with the pre-contrast image data. Any signal intensity cut-off or narrowing of more than 50% was regarded as significant stenosis. Ultrasound Doppler (USD) study was performed with both colour and spectral studies. Peak systolic velocity (PSV) of greater than 2.0 m/s and acceleration time (AT) greater than 120ms was regarded as positive for TRAS. These were then compared with the digital subtraction angiography (DSA) as the gold standard. RESULTS: A total of nine examinations performed in seven patients were included in the analysis. MRA correlated with the DSA findings in eight examinations, with one false negative. USG correlated with DSA in six examinations, with two false negative and one false positive case. CONCLUSION: In our opinion, GD-MRA is a promising and non invasive technique in the detection of TRAS. PMID- 10541390 TI - Carotid stenosis in the real world--can Doppler ultrasound replace angiography in a district general hospital setting? AB - Large trials have confirmed the benefit of carotid endarterectomy in the prevention of stroke in patients with transient ischaemic attacks and > or =70% stenosis of the ipsilateral internal carotid artery. Invasive confirmatory angiography carries some risk, but these patients can be identified by Doppler ultrasound. Non-invasive confirmatory testing with spiral computed tomographic angiography or magnetic resonance angiography is not easily available in many hospitals. In this study, criteria have been developed for use in this unit to identify significant carotid artery stenosis and enable selection for surgery after Doppler ultrasound alone, with known degrees of sensitivity, specificity and accuracy. Carotid arteriography is reserved for a minority of cases. PMID- 10541391 TI - The diagnostic performance of a PC-based teleradiology link. AB - AIM: Evaluation of the diagnostic performance of a personal computer based teleradiology link. MATERIALS AND METHODS: Two experienced radiologists assessed 100 cases, all based on chest and skeletal films using teleradiology for 50. These assessments were compared with the consensus of a panel of three independent radiologists. RESULTS: Diagnostic performance of teleradiology and conventional film was similar (sensitivity 88 vs. 90%; specificity 96 vs. 90%; accuracy 91 vs. 90%; not significant). However, the quality of teleradiology images was rated poorer, and the confidence in diagnosis was lower with teleradiology. ROC curve analysis, taking into account diagnostic confidence, showed significantly poorer performance for teleradiology at all thresholds when chest X-rays only were considered. There was no significant difference for skeletal images, although the two smooth curves crossed, suggesting teleradiology might be better when the specificity is high. CONCLUSION: These findings suggest that when this type of teleradiology system is used, the value of rapid reporting must be balanced against poorer image quality, particularly for chest X-rays. PMID- 10541392 TI - Radiologic findings of bronchogenic carcinoma with pulmonary metastases at presentation. AB - PURPOSE: To describe the radiologic findings in patients with primary bronchogenic carcinoma and pulmonary metastases at presentation. MATERIALS AND METHODS: A retrospective review of patients with bronchogenic carcinoma who at presentation had pulmonary metastases. RESULTS: Fourteen (52%) men and 13 (48%) women with a mean age of 60 years were identified. Adenocarcinoma was the most common histology (70%). The number of nodules varied, although 78% of patients had greater than 50 nodules. Nodules size ranged from 2 to 30 mm, but 82% of patients had nodules less than 10 mm in diameter. Mediastinal lymphadenopathy was seen in 41% of patients, and pleural disease in 44% of patients. Only 37% had radiologic evidence of extrathoracic disease, with bone metastases (30%) being the most common. CONCLUSION: Multiple pulmonary nodules may be the presenting thoracic manifestation of primary bronchogenic carcinoma, with patterns of metastases and survival rates similar to other stage IV patients. PMID- 10541393 TI - Curvilinear areas in the perinephric fat seen on MR images. AB - AIM: On magnetic resonance (MR) images, strands correspond to curvilinear areas running in the perinephric fat, and haloes to those lying on the renal surface. Our aim was to examine the diagnostic significance and histopathological basis of these areas. PATIENTS AND METHODS: MR images obtained in 46 patients without renal disease and 96 patients with renal disease were assessed for the signal intensity and extent of strands and haloes, and their degree of right-left asymmetry. RESULTS: Strands usually revealed low signal intensity on T1-weighted MR images and high signal intensity on fat-suppressed T2-weighted images and contrast-enhanced fat-suppressed T1-weighted images. Haloes revealed high signal intensity on fat-suppressed T2-weighted images, but most of haloes were not clearly depicted on T1-weighted images or contrast-enhanced fat-suppressed T1 weighted images. Strands and haloes were common and usually symmetrical or only mildly asymmetrical in both patient groups. However, in 11 of the 96 patients with renal disease, prominent strands and/or haloes appeared with remarkable asymmetry and likely represented definite changes in the perinephric fat. At histopathology, vascular loose fibrous tissue was found at the sites of strands and haloes. CONCLUSION: Strands and haloes usually represent normal anatomical variations. However, the presence of prominent strands or haloes with remarkable right-left asymmetry implies abnormality and may provide additional information in the evaluation of renal disease. PMID- 10541394 TI - How reliable is modern breast imaging in differentiating benign from malignant breast lesions in the symptomatic population? AB - AIM: To assess the ability of mammography and ultrasound individually and in combination to predict whether a breast abnormality is benign or malignant in patients with symptomatic breast disease. MATERIALS AND METHODS: Patients included were those in whom histological confirmation of the abnormality following surgical excision was available. Mammographic and ultrasound appearances were prospectively classified using a four-point scale (1 = no significant lesion, 2 = benign lesion, 3 = possibly malignant, 4 = probably malignant). RESULTS: Histological confirmation following surgical excision was available in 559 patients, of which 303 were benign and 256 were malignant. The imaging classification was correlated with histology in these 559 lesions. In predicting final histology, the sensitivity and specificity of mammography alone were 78.9 and 82.7%, respectively, of ultrasound alone were 88.9 and 77.9%, respectively, and of mammography and ultrasound in combination were 94.2 and 67.9%, respectively. Only one patient had both a mammogram and ultrasound reported as normal (category 1 for both tests) in whom subsequent histology revealed a carcinoma (0.4% of all carcinomas). CONCLUSION: We found that the extensive use of ultrasound increases the cancer detection rate in this selected population by 14%. PMID- 10541395 TI - Ultrasound guided compression of femoral artery pseudoaneurysms: modified digital technique shortens repair time. AB - Ultrasound guided graded manual compression therapy has become the standard treatment for femoral artery pseudoaneurysms. It entails blunt ultrasound probe compression on the neck of the aneurysm for considerable periods of time. We describe a technique using digital compression after ultrasound localization of the neck enabling easier and more effective targeting of the manual pressure. Preliminary experience in six consecutive cases has shown it to be effective, safe and to have relatively short compression times. Ultrasound guided digital compression has the potential to deliver more effective pressure making this often onerous procedure less demanding on both patient and operator. PMID- 10541396 TI - CT appearance of midgut volvulus with malrotation in a young infant. AB - Computed tomography (CT) appearances of small bowel malrotation and midgut volvulus (MGV) have rarely been described in paediatric patients. We present spiral CT images of a surgically proven case in a young infant. The literature on imaging techniques to diagnose these conditions is reviewed. Radiation doses of upper gastrointestinal series (UGI) and spiral CT are estimated and compared. PMID- 10541397 TI - Intra-abdominal bleeding caused by spontaneous rupture of an accessory spleen: the CT findings. AB - Accessory spleens are common. Their clinical importance lies in the need to include their removal when performing a splenectomy for primary haematological disorders, or as the source of 'preservable' splenic tissue in cases of ruptured primary spleen. Rupture of a normal spleen almost always occurs because of trauma, spontaneous rupture is rare. In pathological spleens, however, 'spontaneous' rupture is more widely reported, although it is argued that minor trauma is often still responsible in these cases. We report a case of spontaneous isolated rupture of a histologically normal accessory spleen and show the CT findings. PMID- 10541398 TI - Purulent pericarditis presenting as acute abdomen in children: abdominal imaging findings. AB - Purulent pericarditis is rapidly fatal if untreated [1,2]. With increased development of bacterial resistance to antibiotics, severe bacterial infections in children are becoming more frequent [3,4]. We report two children with purulent pericarditis who presented in a 1-month period for evaluation of acute abdominal distention and signs of sepsis. In both, one evaluated with computed tomography (CT) and one with ultrasound, abdominal findings included periportal edema, gallbladder wall thickening, and ascites secondary to right heart failure from cardiac tamponade. Radiologists should be aware that children with purulent pericarditis may have a normal heart size on radiographs, present with acute abdominal symptoms, and demonstrate findings of right sided heart failure on abdominal imaging. PMID- 10541399 TI - Alveolar rhabdomyosarcoma presenting with symmetrical bony and renal metastases in infancy. AB - Alveolar Rhabdomyosarcoma is a small round cell tumour in which may be radiologically and histologically difficult to differentiate from other small round cell tumours such as lymphoma, neuroblastoma and Ewing's tumours. We report a case in infancy of disseminated alveolar rhabdomyosarcoma with symmetrical renal and bony metastases. PMID- 10541400 TI - Endorectal ultrasound in a small cell carcinoma of the rectum; staging and assessment of response to chemotherapy. PMID- 10541401 TI - Low cost technique for vessel sizing. PMID- 10541402 TI - Hyperechoic renal papillae in neonates. PMID- 10541403 TI - Genetic analysis of the D1S80 locus in five North Indian populations. AB - The distributions of the D1S80 alleles and genotypes in five endogamous populations (Lobanas, Jat Sikhs, Brahmins, Khatris and Scheduled castes) was determined by amplified fragment length polymorphism (AMP-FLP) technique. The distributions of the observed genotypes for the five populations conformed to Hardy-Weinberg expectations. Two alleles D1S80*18 and D1S80*24 were observed in all populations at frequencies similar to many Caucasian populations, but they showed significant inter-population variability within the region. Allele *24 varied from 33% (Scheduled caste) to 49% (Lobanas), while the allele *18 frequency was lowest in Lobanas (15%) and highest in Jat Sikhs (25%). There was significant overall heterogeneity among the populations studied for this locus. The heterozygosity, probability of exclusion, match probability and discrimination probability estimates demonstrate the usefulness of this locus for paternity and forensic purposes in Indian populations. PMID- 10541404 TI - Evidence of environmental suppression of familial resemblance: height among US Civil War brothers. AB - This study examines, with historical data, whether within family correlations in height varied across environments and whether variability in height was greater in worse environments. To investigate these hypotheses, brothers were identified who were mustered into the Union Army of the US Civil War, using linked records from the 1850 and 1860 censuses and military and medical records. Heights were available for 3898 men aged 18 and older, of whom 595 were further identified as belonging to 288 family sets of two, three or four brothers. Generalized estimating equations were used to concurrently model the mean height, the variance and the correlation between brothers as a function of county population. Heights decreased as county population size increased (p<0.001). The correlation between brothers' heights decreased significantly (p = 0.032) with increasing county population, and the variance increased (p = 0.026). The correlation ranged approximately from 0.63 in the least populous to 0.24 in the most populous counties. The degree of familial resemblance was lower in environments where mean height was lower, and the variability in height was greater, suggesting that the environmental contribution to the variability in height is of greater relative importance in populations reared, on average, in worse environments. PMID- 10541406 TI - Early nutritional history and motor performance of Senegalese children, 4-6 years of age. AB - The effects of undernutrition on motor coordination and performance of 139 4.0 6.5 year-old Senegalese children were studied. The sample was partitioned into three nutritional history groups: 54 children exposed chronically to a mild-to moderate form of undernutrition (group A), 52 children hospitalized for severe undernutrition during infancy and nutritionally rehabilitated but who had been subsequently exposed to moderate undernutrition (group B); and 33 children from well-off urban households (group C). Tests included six items from the McCarthy (arm coordination) and the Charlop-Atwell (gross motor coordination) scales, and five motor fitness items (endurance run, shuttle run, distance throw, standing long jump, grip strength). Performances improved with age, and boys performed better than girls in all motor fitness tests except the jump, but not in motor coordination items. In general, group C performed better than group A and B in most of the tests. Body dimensions explained a significant part of variance of motor performance, and stature was the main predictor. After removing the effect of age and body size, differences between nutritional groups disappeared in motor performance, but persisted in certain motor coordination items. It is concluded that chronic undernutrition reflected by reduced body size and perhaps muscle mass is an important determinant of the motor performance of preschool Senegalese children. PMID- 10541405 TI - A review of recent dietary intervention trials in the United Kingdom to reduce blood cholesterol levels. AB - A review of 14 UK studies conducted between 1980 and January 1997 showed that blood cholesterol levels can be reduced through screening followed by dietary and behavioural intervention in both general population and high risk individuals (hyperlipidaemic and angina patients). In most studies cholesterol levels were lowered moderately while changes in other risk factors were also in a positive direction. However, it is unclear whether the cholesterol reductions are sufficiently large to have a significant impact in lowering the risk of heart disease in the whole population. PMID- 10541407 TI - Height and weight percentile curves of Beijing children and adolescents 0-18 years, 1995. AB - Nationwide growth survey has been performed once every 10 years in Mainland China since 1975. However, no percentile growth curves for children from birth to 18 years have ever been constructed for clinical use. The third survey was performed in 1995 and from this survey, data of the Beijing population were retrieved to construct the height for age and weight for age percentile curves. A total of 12218 healthy children were examined for height and weight, using the standardized methods. Height, but not weight, was normally distributed. Smoothing was performed separately for each sex, using JPA2 model for mean height for age > 2 years. Otherwise, smoothing was performed for the following curves by polynomial equations up to the order of the 5th degree for height and 12th degree for weight, mean height for age < 2 years, SD curve of height for age from birth to 18 years, median weight for age < 2 years, median weight for age > 2 years, differences between the 3rd, 10th, 25th, 75th, 90th, 97th centiles and the median for age from birth to 18 years. The predicted adult height was 173.43cm for boys and 160.86 cm for girls, with age of peak velocity at 13 years and 11.5 years for boys and girls respectively. Compared to those in Hong Kong, they were 2.3-2.5 cm taller signifying a geographical difference in growth even within one country. Compared to the Beijing data collected in 1975 and 1985 the secular changes in growth were more obvious in the first decade, suggesting that the Beijing population was approaching the optimal adult height. PMID- 10541408 TI - Endogamy and inbreeding since the 17th century in past malarial communities in the Province of Cosenza (Calabria, Southern, Italy). AB - Many authors stressed the importance of considering mating patterns, migration and consanguinity when analysing micro-geographic differences in the distribution of the frequency of genetic traits (thalassaemia and glucose 6 phosphate dehydrogenase deficiency (G6PD) in populations living in areas of past malarial endemy. Therefore, the present work was aimed at estimating the reproductive isolation of Calabria, an Italian region that experienced endemic malaria until very recently. The research was carried out on 15311 records of marriage from Parish Books of four villages located in the past malarial area, and four situated in the non-malarial region. Endogamy rates were high in every village and decreased only in the present century as a consequence of the breakdown of isolation. In the earlier periods, the rates ranged between 93-84% in non malarial villages, and between 96-66% in the past malarial area. The rate of consanguineous marriages was low in all villages: in the malarial area it was 2.15% on average, whereas in the non-malarial villages it ranged between 2-16%. Its trend increased with time almost everywhere. Concerning values, differences between past malarial and non-malarial villages in earlier periods are not consistent as they ranged from 0.1 x 10(-3) to 1 x 10(-3). In the present century, however, a was higher in the non-malarial villages. Observed changes of the coefficient a since the 19th century are due to the increased frequency of first cousin marriages. Isonymy rates were lower than 2% in all past malarial towns in all periods, whereas in nonmalarial villages they ranged between 1.2 9.5% and increased with time. Inbreeding coefficients F are always higher than alpha values, but show the same trend with time. They were between 0.0006-0.0045 in past malarial towns, and between 0.0017-0.024 in non-malarial villages. In non malarial villages Fn displayed noticeable negative values in two situations in the earlier periods. In conclusion, given the above mating patterns and the observed distribution of frequencies of G6PD deficient hemizygous and thalassemic heterozygous in the investigated villages, there is clear evidence in this area for the absence of any specific role of reproductive isolation and consanguinity on the distribution of genetic traits related to past malaria presence. PMID- 10541409 TI - Consanguineous marriages in Denizli, Turkey. AB - For the study 1000 families were interviewed during 1996 in the city of Denizli, which is situated in Western Anatolia and has a population of 79211 families. The overall rate of consanguinity was 11.7%, with a mean inbreeding coefficient of 0.00873. The principal type of consanguineous marriage recorded was between first cousins, which accounted for 49.6% of all unions. For both sexes, a significant negative association was observed between consanguinity and mean age at marriage and level of education. PMID- 10541410 TI - Wrinkles induced by the use of smoothing procedures applied to serial growth data. PMID- 10541411 TI - Attribution of intent and role-taking: cognitive factors as mediators of aggression with people who have mental retardation. AB - An assessment task was devised to explore the attributional style and role-taking ability of 22 aggressive and 22 nonaggressive adults with moderate to mild mental retardation. Aggressive participants showed a hostile bias in their interpretation of others' intentions towards themselves within ambiguous interpersonal situations, although no difference was found across groups within clearly provocative situations. Furthermore, the role-taking ability of the aggressive participants was superior to the nonaggressive group, undermining the focus on global problems of interpersonal understanding as a causal factor of aggression for people with mental retardation. PMID- 10541412 TI - Craniofacial maturity and perceived personality in children with Down syndrome. AB - In this pair of studies, we examined whether the common perception of a positive Down syndrome personality is associated with a youthful craniofacial appearance, similar to Zebrowitz's (1997) "babyface." In Study 1, 43 observers rated photographs of age-matched children with Down syndrome, another mental retardation syndrome (5p- syndrome), and typically developing children. Those with Down syndrome were perceived as being more physically babyfaced and more likely to behave in an immature manner. We controlled for the effect of familiarity with Down syndrome in Study 2 by employing a within-etiology design in which 128 observers rated 12 pictures of 10-year-old children with Down syndrome. Results showed that more physically babyfaced children with Down syndrome are more subject to the overgeneralization. PMID- 10541413 TI - Mothers and fathers of children with Down syndrome: parental stress and involvement in childcare. AB - Parental stress was examined in socioeconomically matched samples of mothers and fathers of children with Down syndrome and typically developing children. Parents of children with Down syndrome perceived more caregiving difficulties, child related stress (distractibility, demandingness, unacceptability), and parent related stress (incompetence, depression, health problems, role-restriction) than did parents of typically developing children. For the combined groups of parents, mothers' stress was associated with children's caregiving difficulties; fathers' stress, with children's group status (Down syndrome, typically developing). Mothers who reported more responsibility for childcare perceived more difficulties with health, role restriction, and spousal support. Fathers who reported more responsibility for childcare perceived fewer difficulties with attachment and parental competence. Partner stress was associated both with mothers' and with fathers' stress. PMID- 10541414 TI - Needs and supports reported by Latino families of young children with developmental disabilities. AB - We interviewed 200 Latino parents (50 Mexican couples, 50 Puerto Rican couples) living in the United States to determine needs and supports related to raising a child with a disability and to identify variables related to reported needs and supports. The pattern of needs expressed was similar to that found in previous studies, but the number was substantially higher. More support was reported from family and formal sources than from friends or informal sources. Using repeated measures of analysis of covariance involving six family variables and three child variables, we found that English language proficiency was the only variable to account for significant variance in needs and supports. PMID- 10541415 TI - Leaving or launching? Continuing family involvement with children and adolescents in placement. AB - Postplacement involvement and well-being of 53 families who had placed their child into a residential facility were studied. Parents were interviewed an average of 1, 2, and 3.5 years following placement. Visitation remained moderately high, with some type of visit reported at least monthly at Time 3 by 83% of families. Expressed attachment was high and stable, and most parents reported experiencing less guilt than at the time of placement. Parents were still thinking and talking about the child frequently. They were more likely, over time, to view the placement as permanent, and for about two thirds, the ideal placement was a residential facility. Virtually all parents reported family life to be better following placement, especially in recreation, social life, and relationships with their other children. PMID- 10541416 TI - Puerto Rican families caring for an adult with mental retardation: role of familism. AB - The role of familism (a cultural value including interdependence among nuclear and extended family members for support, loyalty, and solidarity) in caregiving was explored for Puerto Rican mothers with children with mental retardation living at home. Familism--defined here as direct caregiving provided by family members to the person with mental retardation, mothers' social support networks, and mothers' obligations to other family members--was hypothesized to account for variation in maternal well-being. Better maternal well-being was predicted by larger social support networks, greater satisfaction with social support, and more minor children living in the household. A troubling but not unexpected finding is that these mothers faced many socioeconomic challenges and were in poor health in addition to the challenges of parenting a child with mental retardation. PMID- 10541417 TI - Impact of managed care on quality of care, research, and teaching. PMID- 10541418 TI - Comparative clinical effectiveness of long-term controller therapy for asthma. PMID- 10541419 TI - Chlamydia pneumoniae, asthma, and COPD: what is the evidence? AB - LEARNING OBJECTIVES: Reading this article will familiarize the reader with (1) the unique chlamydial intracellular life cycle and the propensity for human chlamydial infections to become persistent and to result in immunopathologic (inflammatory) damage in target organs and (2) current evidence linking Chlamydia pneumoniae (Cpn) infection to obstructive lung diseases (asthma and chronic obstructive pulmonary disease, COPD). Potential therapeutic implications of the Cpn-asthma association are also discussed. DATA SOURCES: All Medline articles (January 1985 to March 1999) that cross-referenced the exploded MESH headings "lung diseases, obstructive" and "Chlamydia pneumoniae" (N = 76). Additional referenced articles, published abstracts, book chapters, and conference proceedings were also utilized. STUDY SELECTION: (1) Case reports and case series that identified Cpn infection in asthma and/or COPD and (2) epidemiologic studies of markers for Cpn infection in asthma and/or COPD that included one or more control groups. RESULTS: Of 18 controlled epidemiologic studies (over 4000 cases/controls), 15 found significant associations between Cpn infection and asthma using organism detection (polymerase chain reaction (PCR) testing (n = 2 studies) or fluorescent antigen testing (n = 1)), Cpn-specific secretory IgA (sIgA) antibody testing (n = 1), and/or specific serum IgE (n = 2), IgA (n = 4), IgG (n = 3) or other antibody criteria (n = 7). Eight case reports and 13 case series of Cpn infection in asthma (over 100 patients) also include descriptions of improvement or complete disappearance of asthma symptoms after prolonged antibiotic therapy directed against Cpn. Significant associations with COPD (over 1000 cases/controls) were reported in 5 of 6 studies. Results of treating chronic chlamydial infections in COPD patients have not been reported. CONCLUSIONS: Although the full clinical significance of these Cpn-obstructive lung disease associations remains to be established, reports of asthma improvement after treatment of Cpn infection deserve further investigation. Clinicians who manage asthma should be aware of this information since it may help to manage difficult cases. The hypothesis that Cpn infection in COPD can amplify smoking-associated inflammation and worsen fixed obstruction also deserves further study. PMID- 10541421 TI - Comparison of the efficacy of inhaled fluticasone propionate, 880 microg/day, with flunisolide, 1500 microg/day, in moderate-to-severe persistent asthma. AB - BACKGROUND: Inhaled corticosteroids have become the mainstay of asthma therapy. Few studies however, have compared inhaled steroids in children. We compared the efficacy of inhaled fluticasone propionate (FP), 880 microg/day (2 puffs of 220 microg/puff, BID) with inhaled flunisolide (FLU), 1500 microg/day (3 puffs of 250 microg/puff, BID). METHODS: Thirty children with moderate to severe asthma, mean age 12.7 years (range 10 to 17 years), mean duration of asthma 8.4 years, initially received flunisolide 1500 microg/day for 1 year, and then were switched to fluticasone propionate 880 microg/day and followed for an additional year. Pulmonary function tests (PFTs) were monitored and analyzed before and after the switch for the duration of study. Mean percent predicted for age values for FVC, FEV1, FEF25-75%, and FEFR were compared at 1 month, 2 to 6-month intervals, and 7 to 12-month intervals and during the same season of the year. Pulmonary function tests within 3 weeks of an exacerbation were not included in the study. The number of asthma exacerbations, emergency room visits, hospital admissions, and number of school days lost were also compared. RESULTS: There was significant improvement in mean asthma exacerbations/patient/year (1.7 +/- 1.66 SD) versus (4 +/- 2.6) (P < .0002); mean emergency room visits/patient/year (0.23 +/- 0.62) versus (1.2 +/- 1.74) (P = .004); mean hospital admissions for asthma/patient/year (0.2 +/- 0.61) versus (1.13 +/- 1.45) (P < .0002); and number of school days lost/patient/year (1.4 +/- 2.38) versus (7.93 +/- 6.7) (P < .0002) while patients were receiving fluticasone propionate as compared with flunisolide. Also, the mean percent values predicted for age in all time-periods (at 1 month, 2 to 6 months, and 7 to 12 months) revealed significant improvement in FEV1 and FEF25-75% (P < .05 for both parameters). As PFT can be affected by seasonal changes, PFT parameters were compared during the same season of the year and significant improvement in FVC and FEV1 was observed in all seasons while patients were receiving fluticasone propionate (FP) compared with flunisolide (FLU) (P < .05 for all parameters). Significant improvement in PEFR and FEF25-75% was observed only in spring and summer season. CONCLUSION: Fluticasone propionate 880 microg/day improved lung function and quality of life in adolescents with moderate-to-severe asthma when compared with flunisolide 1500 microg/day. PMID- 10541420 TI - A six-month-old boy with neonatal lupus and cardiac enlargement. PMID- 10541422 TI - Sensitivity to the house dust mite and airway hyperresponsiveness in a young adult population. AB - BACKGROUND: The pathogenic mechanisms of airway hyperresponsiveness (AHR) in asthma are unknown and only a few studies have examined the importance of sensitivity to antigens in AHR in young adults. OBJECTIVE: We investigated the correlation between AHR and sensitivity to specific antigens, atopy, history of childhood asthma and spirometry in a young adult population. METHODS: Based on the results of interviews with 447 students at our university, 308 non-smoker students were classified into six groups. Group 1 comprised subjects with intermittent mild bronchial asthma; group 2, subjects with history of childhood asthma; group 3, subjects with atopic disease, and a RAST score for Dermatophagoides farinae (Def) of > or = 2; group 4, normal subjects with a RAST score for Def of > or = 2; group 5, subjects with cedar pollinosis; and group 6, normal subjects. We measured AHR to methacholine (MCh), spirometry, immunoglobulin E-radioimmunosorbent test (IgE-RIST), IgE-radioallergosorbent test to six common antigens, eosinophil cationic protein (ECP), and eosinophil count in peripheral blood in each subject. RESULTS: Airway hyperresponsiveness to MCh did not correlate with IgE-RIST, eosinophil count, or ECP. The highest AHR to MCh was present in groups 1 and 2 and lowest in groups 5 and 6. Multiple regression analysis showed that sensitivity to Def was the only factor that significantly influenced AHR to MCh. Airway hyperresponsiveness to MCh of groups with a RAST score for Def of 0/1 was lower than groups with a RAST score of 2 to 6. Airway hyperresponsiveness to MCh did not correlate with the degree of positivity to Def antigen among positive sensitized groups (RAST score 2 to 6). CONCLUSIONS: Sensitivity to mite antigen may be important in the pathogenesis of AHR and Def is a major contributing antigen in young adults in Japan. Once asthma occurs, AHR remains positive for a long time even after the disappearance of asthma-related symptoms. PMID- 10541423 TI - Once-daily fexofenadine HCl improves quality of life and reduces work and activity impairment in patients with seasonal allergic rhinitis. AB - BACKGROUND: Fexofenadine HCl (Allegra, Telfast) is approved in the US for twice daily dosing for treatment of seasonal allergic rhinitis. OBJECTIVE: To determine the effect of once-daily fexofenadine HCl on patient-reported quality of life and impairment at work, in the classroom, and in daily activities due to seasonal allergic rhinitis symptoms. METHODS: This placebo-controlled, double-blind, randomized study included patients aged 12 to 65 years with moderate-to-severe seasonal allergic rhinitis symptoms. Outcomes were assessed using self administered questionnaires at baseline, week 1, and week 2. Outcome measures included change from baseline in: overall Rhinoconjunctivitis Quality of Life Questionnaire (RQLQ) score; individual RQLQ domain scores; work, classroom, and daily activity impairment measured using the Work Productivity and Activity Impairment (WPAI) instrument; and ratings in 3 generic health domains from the SF 36 Health Survey. RESULTS: Intent to treat efficacy analyses included 845 patients from 40 sites. Patients receiving either 120 or 180 mg QD fexofenadine HCl reported significantly greater improvement (P < or = .006) in overall RQLQ score than patients receiving placebo. Similarly, both fexofenadine treatment groups reported significantly greater reductions in overall work impairment and daily activity impairment compared with the placebo group (P < or = .004). There was a trend for improvement in classroom impairment with fexofenadine treatment, although differences from placebo were not statistically significant. Generic health measures demonstrated fexofenadine HCl treatment had a positive effect on general health. CONCLUSION: Once-daily fexofenadine HCl, 120 or 180 mg, significantly improved patient-reported quality of life and reduced performance impairment in work and daily activities due to seasonal allergic rhinitis symptoms compared with placebo. PMID- 10541424 TI - Comparison of the efficacy, safety, and onset of action of mizolastine, cetirizine, and placebo in the management of seasonal allergic rhinoconjunctivitis. MIZOCET Study Group. AB - BACKGROUND: Mizolastine is a potent and selective H1-receptor antagonist with additional anti-allergic properties. OBJECTIVE: The aim of this European multicenter, randomized, double-blind study was to compare the efficacy of mizolastine 10 mg (n = 122), cetirizine 10 mg (n = 125), and placebo (n = 128) once daily for 28 days in patients with seasonal allergic rhinoconjunctivitis (SAR), with focus on the onset of action. METHODS: Symptoms were evaluated by the investigator using a total symptom score (TS) and by the patient (first week). Responders (R) were patients with a TS decrease of at least 50%. Safety was assessed according to the spontaneous reporting of adverse events. RESULTS: Both mizolastine and cetirizine were effective in relieving the symptoms of SAR. After 7 days of treatment, the improvement in TS and responder's rate were significantly (P < .05) greater in patients treated with mizolastine (TS change versus baseline, mean +/- SD: -6.40 +/- 5.71; R: 55%) and cetirizine (TS change versus baseline: -6.24 +/- 5.24; R: 53%) than with placebo (TS change versus baseline: -4.11 +/- 5.91; R: 40%). Both drugs acted rapidly, within 2 hours of the first intake. During the first 3 days, mizolastine relieved symptoms more effectively than cetirizine, the difference being significant on the second (P = .027) and third (P = .050) day. Both mizolastine and cetirizine were well tolerated. CONCLUSION: Mizolastine 10 mg once daily is at least as effective as cetirizine in relieving symptoms of SAR, onset of action is rapid with clinical effect evident within 2 hours. PMID- 10541425 TI - Placebo-controlled, comparative study of the efficacy and safety of triamcinolone acetonide inhalation aerosol with the non-CFC propellant HFA-134a in patients with asthma. Azmacort HFA Clinical Study Group. AB - BACKGROUND: Triamcinolone acetonide (TAA) inhalation aerosol (Azmacort Inhalation Aerosol), a well-established corticosteroid treatment for bronchial asthma, utilizes the chlorofluorocarbon (CFC) propellant P-12, which will be phased out because of environmental concerns. Two TAA aerosol formulations have been developed using a non-chlorofluorocarbon propellant, HFA-134a (Azmacort HFA Inhalation Aerosol delivering TAA 75 microg/puff or 225 microg/puff). OBJECTIVE: This study compared the efficacy and safety of the new 225 microg/puff formulation (TAA-HFA 225) to the marketed TAA inhalation aerosol (TAA-CFC) and to placebo in adult patients with moderate-to-severe persistent asthma. METHODS: After a 5-day to 21-day baseline period during which all patients received TAA CFC 150 microg/day, 538 patients were randomized to one of the following treatment schedules: TAA-HFA 450, 900, or 1800 microg/day; TAA-CFC 450 or 900 microg/day; or placebo for 12 weeks. RESULTS: All active treatment groups showed statistically significant improvement compared with placebo in pulmonary function (FEV1, FEF25-75%, morning and evening PEF), use of rescue albuterol, and asthma symptom scores. Improvements in all variables occurred within 1 week of treatment. CONCLUSIONS: The TAA-HFA 225 exhibited similar safety and efficacy profiles to the two equivalent doses of TAA-CFC studied. Our findings indicate that TAA-HFA is a safe and effective replacement for the currently marketed CFC containing product. The higher strength 225 microg/puff formulation provides effective control of asthma with fewer inhalations. PMID- 10541427 TI - Glucocorticosteroid treatment for cerebrospinal fluid eosinophilia in a patient with ventriculoperitonial shunt. AB - BACKGROUND: Cerebrospinal fluid (CSF) eosinophilia commonly occurs in patients with ventriculoperitoneal (VP) shunts and is associated with shunt complications such as obstruction or infection. Glucocorticosteroids (GCS) are effective in reducing eosinophilia and eosinophils in skin, nasal mucosa, and airway epithelium. Effects of GCS on CSF eosinophils has not been reported. OBJECTIVE: To demonstrate glucocorticosteroid effects on the CSF eosinophil levels and to propose that GCS may be used as a therapeutic agent for CSF eosinophilia. RESULT: A case report of a patient with congenital hydrocephalus and a VP shunt developed CSF eosinophilia associated with latex allergy and shunt malfunction. Daily treatment with 2 mg/kg of methylprednisolone was associated with reduced peripheral eosinophilia and slightly reduced CSF eosinophil counts. Pulse methylprednisolone, 15 mg/kg, was associated with complete reduction of CSF eosinophils and prolonged VP shunt survival. CONCLUSION: Systemic glucocorticosteroids effectively reduce CSF eosinophils. Glucocorticosteroids may be beneficial for treatment of CSF eosinophilia associated with VP shunt malfunction. PMID- 10541426 TI - Age-dependent tendency to become sensitized to other classes of aeroallergens in atopic asthmatic children. AB - BACKGROUND: Several longitudinal studies report that allergic sensitization increases with age from childhood to adulthood. OBJECTIVE: To evaluate whether an age-dependent tendency to become sensitized to new classes of allergens is present in atopic children, we studied retrospectively the changes in allergic sensitization in 165 asthmatic patients, monosensitized (ie, sensitized to only one class of allergens) in the first survey. METHODS: All the children (18 months to 8 years at enrollment), attended our outpatient clinics twice, at time intervals ranging from 2 to 10 years. On each visit, sensitization to house dust mites, pollens, animal danders, and molds was determined by skin prick test. RESULTS: We found that 43.6% (n = 72) of the patients became polysensitized on the second survey. According to age on first survey, the patients were further divided into two age groups: (1) group 1 = 18 months to < 5 years old (n = 98) and (2) group 2 = 5 to 8 years (n = 67). The transition from monosensitization to polysensitization observed in the entire population was present in both groups: 47 (47.9%) of the 98 children in group 1 and 25 (37.3%) of the 67 children in group 2 showed to be sensitized to more classes of allergens, as compared with first survey. Both in the whole population and in the two age subgroups, the changes in the frequency of monosensitization between the two evaluations were time-dependent (P < .05, each Chi(2)). Finally, to investigate whether monosensitization to a specific class of allergens could favor the development of polysensitization, we evaluated the frequency of polysensitization in the second survey in patients originally monosensitized to house dust mites or to pollens. We found that of the 130 patients originally monosensitized to house dust mites, 59 became polysensitized (45.4%), while of the 28 patients originally monosensitized to pollens, 9 became polysensitized (32.1%) (P > . 1). Similar results were obtained when patients were divided into age groups. CONCLUSION: These data demonstrate that (1) monosensitized children are likely to become polysensitized and (2) house dust mite sensitization and, at a lower degree, pollen sensitization, apparently seem to play a "triggering" role in the development of polysensitization, since a high proportion of children originally monosensitized to house dust mites or to pollens became polysensitized. PMID- 10541428 TI - Relationships between atopy and lung function: results from a sample of one hundred medical students in Japan. AB - BACKGROUND: The prevalence of allergic diseases has been increasing dramatically and several studies have shown that atopy is related to asthmatic symptoms and bronchial hyperresponsiveness. OBJECTIVE: To observe the relationships between atopic status and asthmatic predisposition (obstructive change in lung function) in apparently healthy young adults in Japan. METHODS: A sample of 100 healthy Japanese medical students were subjected to a skin prick test for 11 common aeroallergens and food allergens, and their spirometric lung function was measured. RESULTS: Surprisingly, 90% of them showed a positive prick test result for at least one of the 11 allergens tested, and 59% of them showed allergic responses to more than three allergens. The positive rate for Dermatophagoidesfarinae (Der) was the highest (71.0%), followed by house dust (57.0%), Dactylois gloinerata (42.0%), Cryptomeria gromerata (Cry) (40.0%), and cat fur (39.0%). Furthermore, there was no statistical difference in the positive rates for Der and Cry between groups with and without either the present illness or past history of any of the three major allergic diseases: bronchial asthma (BA), atopic dermatitis (AD), or allergic rhinitis (AR). Compared with the positive rates for these aeroallergens, those for food allergens were much lower (4% to 9%). Several lung function parameters, including the levels of FEV1% and %V50 which reflect obstructive pulmonary changes, showed significant negative correlation to the number of skin prick test-positive allergens. The same correlation was observed for groups without either the present illness or past history of BA. CONCLUSION: These data suggest that those who are multi-allergic tend to feature subclinical asthma-like changes in their lung functions. Further studies are needed to determine whether this multi-allergic status can lead to future onset of asthma or other allergic diseases. PMID- 10541429 TI - Leukotriene modifiers in chronic urticaria. PMID- 10541430 TI - An automated technique for most-probable-number (MPN) analysis of densities of phagotrophic protists with lux AB labelled bacteria as growth medium. AB - An automated modification of the most-probable-number (MPN) technique has been developed for enumeration of phagotrophic protozoa. The method is based on detection of prey depletion in micro titre plates rather than on presence of protozoa. A transconjugant Pseudomonas fluorescens DR54 labelled with a luxAB gene cassette was constructed, and used as growth medium for the protozoa in the micro titre plates. The transconjugant produced high amounts of luciferase which was stable and allowed detection for at least 8 weeks. Dilution series of protozoan cultures and soil suspensions were inoculated into micro titre plates amended with a suspension of the transconjugant. After 45 days measurement of light emission allowed detection of individual wells in the titre plates, where protozoan grazing had removed the inoculated bacteria. PMID- 10541431 TI - Measurement of charge transfer during bacterial adhesion to an indium tin oxide surface in a parallel plate flow chamber. AB - An experimental method is described for the measurement of charge transfer during bacterial adhesion in situ to a transparent, semiconducting indium tin oxide (ITO) coated glass plate in a parallel plate flow chamber. Bacterial adhesion is measured simultaneously with either the electric potential or the capacitance of the surface. Initial bacterial adhesion was accompanied by a change in electric potential of the surface with no measurable change in capacitance. Consequently, it can be assumed that the change in electric potential of the surface is due to charge transfer between bacteria and the surface, and it can be calculated that, on average, a charge of about 10(-14) C per bacterium is exchanged during initial adhesion, which corresponds to only several percent of the total surface charge of a bacterium. Charge transfer could either be to or from the bacterial cell surface, dependent on the bacterial strain involved and the ionic strength used. PMID- 10541432 TI - Sustained mitogen-activated protein kinase activation is induced by transforming erbB receptor complexes. AB - We used a genetic approach to characterize features of mitogen-activated protein kinase (MAPK) activation occurring as a consequence of expression of distinct erbB receptor combinations in transformed human cells. Kinase-deficient erbB proteins reduced epidermal growth factor (EGF)-induced tyrosine phosphorylation of endogenous Shc proteins and also reduced immediate and sustained EGF-induced ERK MAPK activities in human glioblastoma cells, although basal ERK MAPK activities were unaffected. Basal and EGF-induced JNK and p38 MAPK kinase activities were equivalent in parental cancer cells and EGFR-inhibited subclones. When ectopically overexpressed in murine fibroblasts and human glioblastoma cells, a constitutively activated human EGF receptor oncoprotein (deltaEGFR) induced EGF-independent elevation of basal ERK MAPK activity. Basal JNK MAPK kinase activity was also specifically induced by deltaEGFR, which correlated with increased phosphorylation of a 54-kDa JNK2 protein observed in deltaEGFR containing cells. The JNK activities in response to DNA damage were comparably increased in cells containing wildtype EGFR or deltaEGFR. Consistent with the notion that transforming erbB complexes induce sustained and unregulated MAPK activities, coexpression of p185(neu) and EGFR proteins to levels sufficient to transform murine fibroblasts also resulted in prolonged EGF-induced ERK in vitro kinase activation. Transforming erbB complexes, including EGFR homodimers, deltaEGFR homodimers, and p185(neu)/EGFR heterodimers, appear to induce sustained, unattenuated activation of MAPK activities that may contribute to increased transformation and resistance to apoptosis in primary human glioblastoma cells. PMID- 10541433 TI - Targeted gene transfer system using a streptavidin-transforming growth factor alpha chimeric protein. AB - The previously reported streptavidin-TGFalpha chimeric protein-based delivery system (Ohno and Meruelo, DNA Cell Biol. 15:401-406, 1996) could efficiently transfer protein molecules into A431 cells via the epidermal growth factor (EGF) receptor. We have modified this delivery system for the transfer of DNA. For this purpose, we have linked the chimeric protein ST-TGFalpha to DNA through biotinylated polylysine molecules. We show with this system, in the presence of the endosome-destabilizing reagent chloroquine, an average of 50-fold increase in reporter gene expression in comparison with polylysine DNA complexes alone. This gene expression is specific for EGF receptor-expressing cells and is blocked by EGF-binding molecules. These results suggest that the ST-TGFalpha biotinylated polylysine system could be used to deliver DNA to targeted cells. PMID- 10541434 TI - NF-kappaB inhibits expression of the alpha1(I) collagen gene. AB - Fibrosis results from an increase in the synthesis and deposition of type I collagen. Fibrosis is frequently associated with inflammation, which is accompanied by increased levels of tumor necrosis factor-alpha (TNFalpha) and activation of the transcription factor NF-kappaB. However, several agents known to activate NF-kappaB, such as phorbol 12-myristate 13-acetate (PMA) and TNFalpha, result in decreased expression of type I collagen. Therefore, we directly examined the effects of NF-kappaB on alpha1(I) collagen gene expression in two collagen-producing cells, NIH 3T3 fibroblasts and hepatic stellate cells (HSCs). Transient transfections of NIH 3T3 cells or HSCs using NF-kappaB p50, p65, and c-Rel expression plasmids with collagen reporter gene plasmids demonstrated a strong inhibitory effect on transcription of the collagen gene promoter. Dose-response curves showed that p65 was a stronger inhibitor of collagen gene expression than was NF-kappaB p50 or c-Rel (maximum inhibition 90%). Transient transfections with reporter gene plasmids containing one or two Spl binding sites demonstrated similar inhibitory effects of NF-kappaB p65 on the activity of these reporter genes, suggesting that the inhibitory effects of NF kappaB p65 are mediated through the critical Spl binding sites in the alpha1(I) collagen gene promoter. Cotransfection experiments using either a super-repressor I[ke]B or Spl partially blocked the inhibitory effects of p65 on collagen reporter gene activity. Coimmunoprecipitation experiments demonstrated that NF kappaB and Spl do interact in vivo. Nuclear run-on assays showed that NF-kappaB p65 inhibited transcription of the endogenous alpha1(I) collagen gene. Together, these results demonstrate that NF-kappaB decreases transcription of the alpha1(I) collagen gene. PMID- 10541435 TI - A G1 cell cycle arrest induced by ligands of the reovirus type 3 receptor is secondary to inactivation of p21ras and mitogen-activated protein kinase. AB - The reovirus type 3 S1 gene product (type 3 hemagglutinin; HA3) is the viral protein responsible for binding to a mammalian cell-surface receptor. It has been shown that HA3 binding to its receptor inhibits cell growth, even in the continuous presence of serum mitogens. Here, receptor-mediated signal transduction leading to growth arrest was studied after binding with synthetic or recombinant ligands in the absence of viral infection. Receptor ligation caused rapid inactivation of p21(ras), a decrease in Raf phosphorylation and in mitogen activated protein kinase (MAPK) enzymatic activity, and G1 cell cycle arrest. Transfection and expression of constitutively active v-Has-ras prevented the G1 arrest, indicating that inactivation of p21(ras) is causative. Interestingly, v Has-ras expression also decreased the efficiency of reoviridae replication, suggesting that inactivation of p21(ras) signals is required at some step of the viral cycle. This study may define new mechanisms regulating cell growth and support the approach of using viral proteins to identify and study cellular receptors. Synthetic receptor ligands with antiproliferative properties may be useful in drug development with the aim of blocking mitosis. PMID- 10541436 TI - DNA priming-protein boosting enhances both antigen-specific antibody and Th1-type cellular immune responses in a murine herpes simplex virus-2 gD vaccine model. AB - It has previously been reported that herpes simplex virus (HSV)-2 gD DNA vaccine preferentially induces T-helper (Th) 1-type cellular immune responses, whereas the literature supports the view that subunit vaccines tend to induce potent antibody responses, supporting a Th2 bias. Here, using an HSV gD vaccine model, we investigated whether priming and boosting with a DNA or protein vaccine could induce both potent antibody and Th1-type cellular immune responses. When animals were primed with DNA and boosted with protein, both antibody and Th-cell proliferative responses were significantly enhanced. Furthermore, production of Th1-type cytokines (interleukin-2, interferon-gamma) was enhanced by DNA priming protein boosting. In contrast, protein priming-DNA boosting produced antibody levels similar to those following protein-protein vaccination but failed to further enhance Th-cell proliferative responses or cytokine production. DNA priming-protein boosting resulted in an increased IgG2a isotype (a Th1 indicator) profile, similar to that induced by DNA-DNA vaccination, whereas protein priming DNA boosting caused an increased IgG1 isotype (a Th2 indicator) profile similar to that seen after protein-protein vaccination. This result indicates that preferential induction of IgG1 or IgG2a isotype is determined by the type of priming vaccine used. Thus, this study suggests that HSV DNA priming-protein boosting could elicit both potent Th1-type cellular immune responses and antibody responses, both of which likely are important for protection against HSV infection. PMID- 10541437 TI - Secretion of metalloendopeptidase 24.15 (EC 3.4.24.15). AB - The metalloendopeptidase EP24.15 (EC3.4.24.15) is a neuropeptide-metabolizing enzyme present in neural and endocrine tissues, presumably functioning extracellularly. Because the majority of the EP24.15 activity is identified in the soluble fraction of cellular homogenates, suggesting that the enzyme is primarily an intracellular protein, we addressed the issue of how EP24.15 arrives in the extracellular environment. We utilized a model system of neuroendocrine secretion, the AtT20 cell. According to both enzymatic activity and immunologic assays, EP24.15 was synthesized in and released from AtT20 cells. Under basal conditions and after stimulation by corticotropin-releasing hormone or the calcium ionophore A23187, EP24.15 activity accumulated in the culture medium. This secretion was not attributable to cell damage, as judged by the absence of release of cytosolic enzyme markers and the ability to exclude trypan blue dye. Pulse-chase analysis and subcellular fractionation of AtT20 cell extracts suggested that the mechanism of EP24.15 secretion is not solely via classical secretory pathways. Additionally, drugs which disrupt the classical secretory pathway, such as Brefeldin A and nocodazole, blocked A23187-stimulated EP24.15 release yet had no effect on basal EP24.15 release, suggesting differences in the basal and stimulated pathways of secretion for EP24.15. In summary, EP24.15 appears to be secreted from AtT20 pituitary cells into the extracellular milieu, where the enzyme can participate in the physiologic metabolism of neuropeptides. PMID- 10541438 TI - Determinants of the DNA-binding specificity of the Avian homeodomain protein, AKR. AB - AKR (Avian Knotted-Related) was the first example of a vertebrate homeodomain protein with a highly divergent Ile residue at position 50 of the DNA-recognition helix. The protein was cloned from a liver cDNA expression library of a day-9 chick embryo by virtue of its ability to bind to the F' site in the proximal promoter of the avian apoVLDLII gene. Expression of the apoVLDLII gene is completely estrogen dependent, and mutation or deletion of the F' site decreases estrogen inducibility 5- to 10-fold. Subsequent data indicated that AKR is capable of repressing the hormone responsiveness of the apoVLDLII promoter, specifically through binding to F'. Involvement of the F' site in the hormone dependent activation of apoVLDLII gene expression, as well as AKR-mediated repression, strongly suggests that both positive and negative regulatory factors interact with this site. Although several mammalian proteins have now been isolated whose homeodomains share many of the structural features of AKR, including the Ile at position 50, little is known of their functions in vivo or the identities of the genes they regulate. Consequently, the elements through which they exert their effects and the structural determinants of their binding specificities remain largely uncharacterized. In this study, we defined the sequence specificity of binding by AKR using polymerase chain reaction-assisted optimal site selection and determined the affinity with which the protein binds to both the optimized site and the F' site. Additionally, we generated a three dimensional model of the AKR homeodomain binding to its optimized site and probed the validity of the model by examining the consequences of mutating amino acid residues in recognition helix 3 and the N-terminal arm on the binding specificity of the homeodomain. Finally, we present evidence that the F' site itself may act as an estrogen response element (ERE) when in the vicinity of imperfect or canonical EREs and that AKR can repress hormone inducibility mediated via this site. PMID- 10541439 TI - Sports medicine in the new millennium: a vision for 2020. AB - Globalisation, empowerment and technological change will determine the emerging directions in sports medicine in the new millennium. Networks and alliances of scientist and clinician services, as well as electronic profiling of athletes' learning styles, genetic predisposition and other variables, will enhance the spectrum of sports medicine services. Visionary direction will require changes in the organisational paradigms employed, the communication of information to athletes and coaches and the methodologies of assessment. An emphasis on prevention science and clinical and educational interventions will require a clearer focus. The sports medicine scientist and clinician of today must utilise the endowments suggested by Covey and the multiple intelligence models advanced by Gardner in capturing the clarity of focus for sports medicine in the new millennium. PMID- 10541440 TI - Nutritional aspects of immunosuppression in athletes. AB - The literature suggests that a heavy schedule of training and competition leads to immunosuppression in athletes, placing them at a greater risk of opportunistic infection. There are many factors which influence exercise-induced immunosuppression, and nutrition undoubtedly plays a critical role. Misinterpretation of published data and misleading media reports have lead many athletes to adopt an unbalanced dietary regimen in the belief that it holds the key to improved performance. Some sports have strict weight categories, whilst in others low body fat levels are considered to be necessary for optimal performance or seen as an aesthetic advantage. This leads some athletes to consume a diet extremely low in carbohydrate content which, whilst causing rapid weight loss, may have undesirable results which include placing the athlete at risk from several nutrient deficiencies. Complete avoidance of foods high in animal fat reduces the intake of protein and several fat-soluble vitamins. On the other hand, diets with a very high carbohydrate content are usually achieved at the expense of protein. In addition, anecdotal and media reports have often promoted the supposed performance benefits of certain vitamins and minerals, yet most athletes do not realise that micronutrient supplementation is only beneficial when correcting a deficiency, and to date there is little scientific evidence to substantiate claims that micronutrients act as an ergogenic aid. Moreover, excessive intakes of micronutrients can be toxic. Deficiencies or excesses of various dietary components can have a substantial impact on immune function and may further exacerbate the immunosuppression associated with heavy training loads. This review examines the role of nutrition in exercise-induced immunosuppression and the effect of both excessive and insufficient nutrient intake on immunocompetence. As much of the present literature concerning nutrition and immune function is based on studies with sedentary participants, the need for future research which directly investigates the relationship between exercise, training, immunity and nutrition is highlighted. PMID- 10541441 TI - Effects of exercise and training on natural killer cell counts and cytolytic activity: a meta-analysis. AB - Meta-analysis techniques have been used to accumulate data from 94 studies describing the natural killer (NK) cell response of some 900 volunteers to acute and chronic exercise. NK cell numbers have been indicated in terms of CD3 CD16+CD56+, CD16+ or CD56+ phenotypes, and cytolytic activity has been expressed per 10,000 peripheral blood mononuclear cells or in terms of lytic units. Acute exercise has been categorised as sustained moderate (50 to 65% of aerobic power), sustained vigorous (>75% of aerobic power), brief maximal or 'supramaximal', prolonged, eccentric or resistance, and repeated exercise. In general, there was a marked increase in NK cell count at the end of exercise, probably attributable to a catecholamine-mediated demargination of cells. Following exercise, cell counts dropped to less than half of normal levels for a couple of hours but, except in unusual circumstances (e.g. prolonged, intense and stressful exercise), normal resting values are restored within 24 hours. If activity is both prolonged and vigorous, the decrease in NK cell counts and cytolytic activity may begin during the exercise session. Although the usual depression of NK cell count seems too brief to have major practical importance for health, there could be a cumulative adverse effect on immunosurveillance and health experience in athletes who induce such changes several times per week. There is a weak suggestion of an offsetting increase in resting NK cell counts and cytolytic action in trained individuals, and this merits further exploration in studies where effects of recent training sessions are carefully controlled. PMID- 10541444 TI - Symposium on antioxidants and health. Bordeaux, France 18-20 March 1998. Proceedings. PMID- 10541442 TI - The influence of air travel on athletic performance. AB - Rapid transmeridian flight is a common reality for modern athletes and it has often been assumed that air travel has detrimental effects on athletic performance. The plausibility of this assumption is supported by established deteriorations in sleep and mood following transmeridian flight. However, the scientific evidence supporting the assumption is neither consistent nor compelling. Studies that have assessed athletic performance following transmeridian flight have produced mixed results and are characterised by major methodological flaws. Recent retrospective assessments of athletic team performance based on distance travelled have generally failed to indicate performance impairments following transmeridian flight. The plausibility of transmeridian air travel impairing athletic performance would be indicated by demonstration of an internally-driven circadian rhythm of athletic performance, or of deleterious performance consequences following sleep deprivation or desynchronisation between the circadian system and the environment. More rigorous research is needed to establish whether athletic performance is influenced by air travel. PMID- 10541443 TI - Anterior shoulder instability in sport: current management recommendations. AB - In the young athlete, anterior shoulder dislocations are common injuries that usually result in recurrent instability, and often require surgical treatment. Non-operative treatment remains the initial recommended course for most conditions. Operative treatment has advanced to more anatomical repairs, both open and arthroscopic. The purpose of this paper is to review the evaluation and treatment of anterior shoulder instability, to include acute dislocations, acute subluxations and recurrent instability. PMID- 10541445 TI - Type of alcohol and mortality from cardiovascular disease. AB - Many epidemiological studies have described a U-shaped relation between alcohol intake and all-cause mortality (Boffetta and Garfinkel, 1990; Fuchs et al., 1995; Gronbaek et al., 1994; Marmot et al., 1981). Most researchers attribute the 'U' to a combination of beneficial and harmful effects of ethanol itself. It has, on the other hand, been explained as an artefact due to misclassification or confounding (Shaper et al., 1998). Most of the studies of the effect of total alcohol intake have found that the descending leg of the curve mainly is attributable to death from cardiovascular disease (Rimm et al., 1991; Stampfer et al., 1988). Until recently, most studies addressed the effect of the three beverages taken together as ethanol. Studies of the correlation between wine intake per capita in different countries and incidence of ischaemic heart disease gave rise to the hypothesis that there is a a more beneficial effect of wine than of beer and spirits. Leger et al., Renaud and de Lorgeril and later Criqui and Rigel found an inverse relation between incidence rates of ischemic heart disease and wine consumption in different countries, but no such relation for the other types of beverages (Criqui and Rigel, 1994; Leger et al., 1979; Renaud and de Logeril, 1992). PMID- 10541446 TI - "The SU.VI.MAX Study": a primary prevention trial using nutritional doses of antioxidant vitamins and minerals in cardiovascular diseases and cancers. SUpplementation on VItamines et Mineraux AntioXydants. AB - The "SUpplementation en VItamines et Mineraux AntioXydants" (SU.VI.MAX) Study is a randomized double-blind, placebo-controlled, primary-prevention trial which started in 1994 in France. This epidemiologic study is designed to test the efficacy of a daily supplementation with antioxidant vitamins (vitamin C, 120 mg, vitamin E, 30 mg, and beta-carotene, 6 mg) and minerals (selenium, 100 microg, and zinc, 20 mg) at nutritional doses, in reducing the main causes of premature death (cancers and cardiovascular diseases); 12,735 eligible subjects (women aged 35 to 60 years, and men aged 45 to 60 years) were included in 1994 and and will be followed up for 8 years. Participants undergo a yearly visit consisting, every other year, of either biological sampling or clinical examination. They also regularly provide information on health events and dietary intake by filling out computerized questionnaires using the Minitel Telematic Network. After 2 years of supplementation, biochemical indicators of vitamin and trace element status reach reasonable level without reaching concentrations as high as those observed in intervention studies, which tested relatively high doses of antioxidants, and ended up with higher risk of pathology. PMID- 10541447 TI - Hydroquinone: genotoxicity and prevention of genotoxicity following ingestion. AB - Plant-derived polyphenolics and other chemicals with antioxidant properties have been reported to inhibit the expression of genotoxic activity by pro-oxidant chemicals (Sai et al., 1992, 1994; Teel and Castonguay, 1992). In vitro and in vivo studies with ionizing radiation suggest that hydroquinone (HQ) may have similar protective effects (Babaev et al., 1994). The present study was conducted to determine whether HQ is capable of inhibiting the induction of micronuclei in the bone marrow of mice following exposure to an oxidant, potassium bromate or KBrO3 (Nakajima et al., 1989; Sai et al., 1992, 1994). To be able to interpret the results of this work, it was also necessary to determine whether HQ is itself genotoxic when fed in the diet. HQ diets (0.8%) fed to mice for 6 days reduced the background incidence of micronuclei compared with the basal diet. KBrO3 dosed ip (12.5-100 mg/kg) produced a dose-dependent increase in micronuclei as reported by others. Mice fed 0.8% HQ diets 6 days, and then dosed intraperitoneally with KBrO3, showed a 36% reduction in micronuclei across the range of KBrO3 dose levels. This effect was associated with a reduction in the background micronucleus response as well as a reduction in response to KBrO3. Statistical significance (P < or = 0.05), observed at a dose of 25 mg/kg KBrO3 in the mice fed the control diet, was abolished in the group fed 0.8% HQ. When mice were given 50 mg HQ/kg by oral gavage and then given 50 mg KBrO3/kg ip 20 min later, the micronucleus response induced by KBrO3, was lower in animals given HQ. The results of this study demonstrate that large doses of HQ may be given orally without induction of micronuclei or bone marrow depression, that HQ reduces the background micronucleus response in animals fed a basal diet, and that the HQ reduces the micronucleus response to KBrO3 as well as background incidence of micronuclei in KBrO3-dosed animals. The protective effect of HQ may be due to enzyme induction or a direct antioxidant effect of HQ against oxidants commonly present in the diet. PMID- 10541448 TI - Dietary flavonoids: intake, health effects and bioavailability. AB - Flavonoids are polyphenolic compounds that occur ubiquitously in foods of plant origin. Over 4000 different flavonoids have been described. They may have beneficial health effects because of their antioxidant properties and their inhibitory role in various stages of tumour development in animal studies. An estimation of the total flavonoid intake is difficult, because only limited data on food contents are available. It is estimated that humans ingest a few hundreds of milligram per day. The average intake of the subclasses of flavonols and flavones in The Netherlands was 23 mg/day. The intake of flavonols and flavones was inversely associated with subsequent coronary heart disease in most but not all prospective epidemiological studies. A protective effect of flavonols on cancer was found in only one prospective study. Flavonoids present in foods were considered non-absorbable because they are bound to sugars as beta-glycosides. However, we found that human absorption of the quercetin glycosides from onions (52%) is far better than that of the pure aglycone (24%). Flavonol glycosides might contribute to the antioxidant defences of blood. Dietary flavonols and flavones probably do not explain the cancer-protective effect of vegetables and fruits; a protective effect against cardiovascular disease is not conclusive. PMID- 10541449 TI - Mechanisms of action of antioxidants as exemplified in vegetables, tomatoes and tea. AB - Most chronic diseases, including coronary heart disease and many types of cancer depend on the in vivo conversion of cellular macromolecules or of carcinogens to specific reactive, oxidized forms. For that reason, health promoting nutrition involves the daily intake of five to 10 vegetables and fruits, fruit juices, red wine and tea that are rich sources of micronutrients with antioxidant properties, including the antioxidant vitamins C, E and beta-carotene. Tomatoes contain lycopene, a stable, active antioxidant. Many vegetables contain quercetin and related polyphenolic compounds. Tea is a source of epigallocatechin gallate, in green tea, and theaflavin and the associated thearubigins, in black tea. Red wine contains resveratrol. The diverse antioxidants in foods, red wine and tea provide the necessary antioxidant resources for the body to control oxidation reactions in the body with possible adverse consequences. For example, the oxidation of low density lipoprotein (LDL) cholesterol yields a product that damages the vascular system. Thus, a lower intake of saturated fats to decrease the levels of LDL cholesterol, together with an adequate intake of antioxidants, is the optimal approach to lower heart disease risk. Cancer of the stomach involves the consumption of salted, pickled foods yielding direct-acting carcinogens, and their formation is inhibited by vitamins C and E. Cancer in the colon, breast, prostate and pancreas may be caused by a new class of carcinogens, the heterocyclic amines, formed during the broiling or frying of creatinine containing foods, including fish and meats. Their formation and action can be inhibited by antioxidants such as those in soy, tea, vitamin C and also by the synthetic antioxidants BHA or BHT. The growth, cell proliferation and development of abnormal preneoplastic and neoplastic cells also involves oxidation reactions, including the formation of active oxygen or peroxy compounds. Such reactions can be inhibited by antioxidants, such as those in tea, tomatoes or vegetables. Even ageing and longevity in good health would be favoured by the availability of adequate amounts of varied antioxidants. Prevention of the formation and of action of reactive products by antioxidants as present in fruits, vegetables, tomatoes, red wine and tea is of great public health importance in decreasing the risk of major diseases. Prevention is the optimal approach to disease control, and also as an effective route to lower costs of medical care. PMID- 10541450 TI - Intracellular antioxidants: from chemical to biochemical mechanisms. AB - Intracellular antioxidants include low molecular weight scavengers of oxidizing species, and enzymes which degrade superoxide and hydroperoxides. Such antioxidants systems prevent the uncontrolled formation of free radicals and activated oxygen species, or inhibit their reactions with biological structures. Hydrophilic scavengers are found in cytosolic, mitochondrial and nuclear compartments. Ascorbate and glutathione scavenge oxidizing free radicals in water by means of one-electron or hydrogen atom transfer. Similarly, ergothioneine scavenges hydroxyl radicals at very high rates, but it acts more specifically as a chemical scavenger of hypervalent ferryl complexes, halogenated oxidants and peroxynitrite-derived nitrating species, and as a physical quencher of singlet oxygen. Hydrophobic scavengers are found in cell membranes where they inhibit or interrupt chain reactions of lipid peroxidation. In animal cells, they include alpha-tocopherol (vitamin E) which is a primary scavenger of lipid peroxyl radicals, and carotenoids which are secondary scavengers of free radicals as well as physical quenchers of singlet oxygen. The main antioxidant enzymes include dismutases such as superoxide dismutases (SOD) and catalases, which do not consume cofactors, and peroxidases such as selenium-dependent glutathione peroxidases (GPx) in animals or ascorbate peroxidases (APx) in plants. The reducing coenzymes of peroxidases, and as a rule all reducing components of the antioxidant network, are regenerated at the expense of NAD(P)H produced in specific metabolic pathways. Synergistic and co-operative interactions of antioxidants rely on the sequential degradation of peroxides and free radicals as well as on mutual protections of enzymes. This antioxidant network can induce metabolic deviations and plays an important role in the regulation of protein expression and/or activity at the transcriptional or post-translational levels. Its biological significance is discussed in terms of environmental adaptations and functional regulations of aerobic cells. PMID- 10541451 TI - Gluthathione: in defence of the lung. AB - Oxidative stress is implicated in the pathology of numerous diseases of the lung. These include cystic fibrosis, chronic obstructive airway disease and asthma. All these conditions are characterised by an imbalance between the amounts of reactive oxygen species (ROS) and available antioxidant defences. In the lung, ROS arise from endogenous sources, such as the influx of inflammatory cells or exogenous sources, such as from air pollution and cigarette smoke. When ROS production increases the redox balance of the airways alters, and this can lead to bronchial hyperactivity and further inflammation. The lung, like many other tissues, has a range of antioxidant defences which help to maintain a balanced redox status. These antioxidants are present in the intracellular, the vascular and extracellular respiratory tract lining fluid (RTLF) compartments. The reduced glutathione (GSH) content of RTLF is particularly high and new findings are beginning to reveal the role that the RTLF GSH pool plays in defending the lung. PMID- 10541452 TI - Molecular mechanism of cellular uptake and intracellular translocation of alpha tocopherol: role of tocopherol-binding proteins. AB - Vitamin E (alpha-tocopherol) is a lipid-soluble antioxidant which is present in cellular membranes where it plays an important role in the suppression of free radical-induced lipid peroxidation. There are eight naturally occurring homologues of vitamin E which differ in their structure and in biological activity in vivo and in vitro. Various studies have suggested that the tocopherol distribution system favours the accumulation of alpha-tocopherol both in the plasma and different tissues. Mechanisms involved in the preferential accumulation of alpha-tocopherol are not yet well established; however, recent data indicate that both intracellular and membrane alpha-tocopherol-binding proteins may be involved in these processes. A 30 kDa alpha-tocopherol-binding protein (TBP) in the liver cytoplasm is now known to regulate plasma vitamin E concentrations by preferentially incorporating alpha-tocopherol into nascent very low density (VLDL) whereas the 15 kDa TBP may be responsible for intracellular distribution of alpha-tocopherol. The 30 kDa TBP is unique to the hepatocyte whereas the 15 kDa TBP is present in all major tissues. The 15 kDa TBP specifically binds alpha-tocopherol in preference to the delta- and gamma tocopherol and may exclusively transport alpha-tocopherol to these intracellular sites. In addition, the presence of a membrane TBP (TBPpm) in tissues may regulate their alpha-tocopherol levels. Activity of erythrocyte TBPpm appears to be reduced in smokers, which may lead to reduced levels of alpha-tocopherol in these cells despite smokers have similar plasma levels of vitamin E as in non smokers. The current status of the evidence for this directed flow of alpha tocopherol through interactions with these proteins (TBP and TBPpm) is discussed. PMID- 10541453 TI - Antioxidant functions of sulforaphane: a potent inducer of Phase II detoxication enzymes. PMID- 10541454 TI - Modulation of pancreatic carcinogenesis by antioxidants. AB - Previously performed short-term (4-month) studies demonstrated that vitamins C and E, beta-carotene and selenium modulate growth of early putative preneoplastic acinar lesions induced in rat pancreas by azaserine. The present paper summarizes the results of long-term studies performed with azaserine-treated rats maintained on diets high in either beta-carotene, vitamins C and E or selenium. It appeared that rats given a diet high in beta-carotene, vitamin C or selenium, but not vitamin E, developed fewer pancreatic tumours than controls. The chemopreventive effects of these micronutrients were most pronounced when beta-carotene and/or selenium were given during the promotion phase of the carcinogenic process. Surprisingly, cell proliferation in azaserine-induced preneoplastic acinar lesions was higher in rats given beta-carotene and/or selenium via the diet in comparison to controls. It is considered unlikely that any antioxidant alone can be associated with protection against cancer. It is concluded that dietary supplementation of combinations of antioxidants may have practical application in chemoprevention of cancer. PMID- 10541455 TI - Phenolics: blocking agents for heterocyclic amine-induced carcinogenesis. AB - Chemopreventive effects of synthetic and naturally occurring antioxidants on heterocyclic amine (HCA)-induced rat carcinogenesis and mechanisms of inhibition were assessed. In a medium-term liver bioassay, combined treatment with 0.03% 2 amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and synthetic antioxidants such as 1-O-hexyl-2,3,5-trimethylhydroquinone (HTHQ), butylated hydroxyanisole (BHA), butylated hydroxutoluene (BHT), tert-butylhydroquinone (TBHQ) or propyl gallate, each at a dose of 0.25%, inhibited development of preneoplastic glutathione S-transferase placental form (GST-P) positive foci as compared with MeIQx alone, after initiation with diethylnitrosamine (DEN). Of these antioxidants, HTHQ showed the greatest activity. 8-Hydroxydeoxyguanosine (8 OHdG), a marker for DNA damage induced by active oxygen species, and malonedialdehyde and 4-hydroxynonenal levels were not largely influenced by the treatment with MeIQx or antioxidants, either alone or in combination. In the same medium-term liver bioassay, effects of some naturally occurring antioxidants, such as green tea catechins (GTC), hesperidin, chlorogenic acid, quercetin, rutin, curcumin, daidzin, ferulic acid and genistein were also examined. Of these antioxidants, only GTC tended to inhibit GST-P positive foci development, while quercetin, rutin, curcumin, daidzin, ferulic acid and genistein all exerted significant enhancing effects. Examination of HTHQ influence in a medium term liver bioassay with HCA Glu-P-1, in which the experimental period was extended for up to 26 weeks, also demonstrated a significant decrease in the incidence of liver tumours to 40% in the group treated with 0.5% HTHQ and 0.03% 2-amino-6 methyldipyrido[1,2-a:3',2'-d]imidazole (Glu-P-1) as compared with the Glu-P-1 alone value of 89%. Effects of HTHQ on colon carcinogenesis induced by 2-amino-1 methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) were evaluated in a two-stage colon carcinogenesis model using 1,2-dimethylhydrazine (DMH) as an initiator. At week 36, the multiplicity of colon tumours induced by 0.02% PhIP after DMH initiation (9.1+/-6.2/rat) was dose-dependently decreased by the combined treatment with 0.5% HTHQ (3.6+/-1.8, P < 0.001) and 0.125% HTHQ (6.2+/-3.2, not significant). Similarly, the incidence of mammary carcinomas in female F344 rats induced by oral administration of 0.02% PhIP (40%) for 52 weeks was significantly decreased by simultaneous treatment with 0.5% HTHQ (5%). Alpha-tocopherol and chlorophyllin only reduced the multiplicity of carcinomas. Analysis of the influence of HTHQ on metabolic activation of Glu-P-1 or PhIP after incubation with rat S9 mixture and NADPH by HPLC, revealed that each major metabolite was strongly reduced by the addition of HTHQ. Immunohistochemically detected PhIP-DNA adduct positive nuclei in the colon induced by continuous oral treatment with 0.02% PhIP for 2 weeks decreased by the combined treatment with 0.5 or 0.125% HTHQ. These results indicate that synthetic antioxidant HTHQ is a very strong chemopreventor of heterocyclic amine (HCA)-induced carcinogenesis and that depressed metabolic activation rather than antioxidant activity is responsible for the observed effect. PMID- 10541456 TI - In vivo studies on butylated hydroxyanisole. PMID- 10541457 TI - Diet and antioxidant status. PMID- 10541458 TI - Repair of oxidative DNA damage in vitro: a tool for screening antioxidative compounds. AB - Reactive oxygen species (ROS) provoke the formation of base DNA alterations that are processed by an excision step of the lesion followed by a repair synthesis and ligation step to restore the strand continuity. We have reported previously the detection of DNA adducts by an in vitro chemiluminescence DNA repair synthesis assay (Salles et al., 1995) which allows the measurement of repair synthesis by cell-free extracts in damaged plasmid DNA adsorbed on sensitized microplate wells. The 3D (DNA damage detection) assay was performed in the presence of biotin-dUTP which was incorporated during the repair synthesis step. The extent of repair synthesis was measured in an ELISA reaction with ExtrAvidin horse radish peroxidase and chemiluminescence detection. The 3D assay allows detection of any type of base alterations including base oxidation. Interestingly, under controlled production of ROS a screening procedure of antioxidants might be carried out with the 3D assay. By taking advantage of plasmid DNA adsorption, oxidative base damage can be recognized by the Escherichia coli Fpg protein which was detected in an ELISA reaction with specific antibody and chemiluminescence measurement (4D assay). With the sceening procedure of antioxidative compounds in mind, the development of such assays and their drawbacks are discussed. PMID- 10541460 TI - Safety assessment of butylated hydroxyanisole and butylated hydroxytoluene as antioxidant food additives. AB - Butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT) are widely used antioxidant food additives. They have been extensively studied for potential toxicities. This review details experimental studies of genotoxicity and carcinogenicity which bear on cancer hazard assessment of exposure to humans. We conclude that BHA and BHT pose no cancer hazard and, to the contrary, may be anticarcinogenic at current levels of food additive use. PMID- 10541459 TI - Comparative in vitro and in vivo effects of antioxidants. AB - We have investigated the effects of reactive oxygen species (ROS) in a wide variety of systems, both in vitro and in vivo, with particular attention to their genetic effects. Some of the results of these studies, as they relate to the protective effects of antioxidants, are discussed. PMID- 10541461 TI - What is beneficial for health? The concept of functional food. AB - 'Functional Food' is now a very popular term. The conceptual approach developed in the EU-founded FUFOSE (Functional Food Science in Europe) project is rather restrictive, making functional food a food product to be part of the usual dietary pattern. Functional food science that supports the development of functional foods is and must remain part of the science of nutrition. Finally, all that exercise, that extended over the last 3 years, was function-driven because the functions and their modulation are universal, as opposed to a food or food component-driven approach,which is likely to be very much influenced by local, traditional or cultural characteristics. PMID- 10541462 TI - Toward a molecular explanation for long-term potentiation. PMID- 10541463 TI - On beyond LTP. Long-term potentiation. PMID- 10541464 TI - The role of dendritic filtering in associative long-term synaptic plasticity. AB - Several forms of synaptic plasticity in the neocortex and hippocampus depend on the temporal coincidence of presynaptic activity and postsynaptic trains of action potentials (APs). This requirement is consistent with the Hebbian, or correlational, type of cellular learning rule used in many studies of associative synaptic plasticity. Recent experimental evidence suggests that APs initiated in the axosomatic area are actively back-propagated to the dendritic arborization of neocortical and pyramidal cells. High-frequency trains of postsynaptic APs that are used as conditioning stimuli for the induction of Hebbian-like plasticity in both neocortical and hippocampal pyramidal cells display attenuation of the dendritic AP amplitude during the train. This attenuation has been shown to be modulated by neurotransmitters and by electrical activity. We suggest here that both spike train attenuation in the dendrite and its modulation by neurotransmitters and electrical activity may have important functional consequences on the magnitude and/or the sign of the synaptic plasticity induced by a Hebbian pairing procedure. PMID- 10541465 TI - Impaired memory retention and decreased long-term potentiation in integrin associated protein-deficient mice. AB - Previously, we have demonstrated that integrin-associated protein (IAP) mRNA level is approximately fourfold higher in rats showing good retention performance (600 sec) than rats showing poor retention performance (< 80 sec) in an inhibitory avoidance learning paradigm. In the present study, we have used the gene-targeted IAP-deficient mice to further investigate the role of IAP involved in memory formation and hippocampal long-term potentiation (LTP) in vivo. Results revealed that there was a significant impairment in memory retention and a significant reduction in the magnitude of LTP in IAP-deficient mice when compared with the wild-type and heterozygote mice, whereas the wild-type and heterozygote animals did not show marked differences on these measures. Furthermore, the impairment in retention performance of IAP-deficient mice was not due to different sensitivities of these animals to the electric shock. When we examined locomotor activity and rotarod treadmill performance, no differences were observed among these three groups of animals either. Western blot analysis confirmed the lack of IAP protein in IAP-deficient mice, whereas IAP expression was similar in both the wild-type and heterozygote controls. These results together demonstrate that IAP plays an important role in the process of memory formation and synaptic plasticity in mice. PMID- 10541466 TI - Inhibition of the endothelial isoform of nitric oxide synthase impairs long-term memory formation in the chick. AB - Previous studies with general inhibitors of nitric oxide synthase have yielded variable and contradictory results with respect to their effects on memory. This may have been due to differential effects of blocking the various isoforms of this enzyme. We show that day-old chicks trained on a single-trial passive avoidance task suffered significant memory loss from approximately 40 min post training following post-training intracranial administration of a potent inhibitor of eNOS. Administration of a specific nNOS or iNOS inhibitor at the same time had no effect on retention, although a role for either of these isoforms when administered at a different time after learning has yet to be fully investigated. The onset of memory loss following eNOS inhibition is the same as observed following general NOS inhibition, which suggests that amnestic effects observed in previous studies using nonspecific inhibitors may be attributable to blocking the function of eNOS. The findings indicate that eNOS may play a role in memory formation for this task, which is at least distinct from any role that may be played by nNOS. PMID- 10541467 TI - Orphanin FQ suppresses NMDA receptor-dependent long-term depression and depotentiation in hippocampal dentate gyrus. AB - We reported previously that orphanin FQ (OFQ) inhibited NMDA receptor-mediated synaptic currents and consequently suppressed induction of long-term potentiation (LTP) in the hippocampal dentate gyrus. This study examines the effect of OFQ on several other forms of long-term synaptic plasticity in the lateral perforant path of mouse hippocampal dentate gyrus. (1) Long-term depression (LTD): a low frequency stimulation (1 Hz, 15 min) applied to the lateral perforant path induced a long-lasting reduction in the dentate field potentials in slices from 22- to 30-day-old mice. This LTD was sensitive to the NMDA receptor blocker D AP5, and could be significantly attenuated by bath application of OFQ (1 microM, 25 min). (2) Primed LTD: induction of LTD in slices from 50- to 65-day-old mice required a priming procedure consisting of multiple high frequency stimulus trains delivered in the presence of D-AP5 before the low-frequency stimulation. OFQ applied during the low-frequency stimulation, but not during the priming trains, blocked induction of primed LTD. (3) Depotentiation: high-frequency train induced dentate LTP could be reversed by a subsequent low-frequency stimulation. This depotentiation was also attenuated by either OFQ or D-AP5 applied during low frequency stimulation. These results, together with our previous findings, suggest that OFQ inhibits bidirectional changes in synaptic strength in the dentate; and its multiple actions on NMDA receptor-dependent, long-term synaptic plasticity might work in tandem to regulate hippocampus-dependent learning and memory. PMID- 10541468 TI - A necessity for MAP kinase activation in mammalian spatial learning. AB - Although the biochemical mechanisms underlying learning and memory have not yet been fully elucidated, mounting evidence suggests that activation of protein kinases and phosphorylation of their downstream effectors plays a major role. Recent findings in our laboratory have shown a requirement for the mitogen activated protein kinase (MAPK) cascade in hippocampal synaptic plasticity. Therefore, we used an inhibitor of MAPK activation, SL327, to test the role of the MAPK cascade in hippocampus-dependent learning in mice. SL327, which crosses the blood-brain barrier, was administered intraperitoneally at several concentrations to animals prior to cue and contextual fear conditioning. Administration of SL327 completely blocked contextual fear conditioning and significantly attenuated cue learning when measured 24 hr after training. To determine whether MAPK activation is required for spatial learning, we administered SL327 to mice prior to training in the Morris water maze. Animals treated with SL327 exhibited significant attenuation of water maze learning; they took significantly longer to find a hidden platform compared with vehicle-treated controls and also failed to use a selective search strategy during subsequent probe trials in which the platform was removed. These impairments cannot be attributed to nonspecific effects of the drug during the training phase; no deficit was seen in the visible platform task, and injection of SL327 following training produced no effect on the performance of these mice in the hidden platform task. These findings indicate that the MAPK cascade is required for spatial and contextual learning in mice. PMID- 10541469 TI - Lesions of periaqueductal gray dissociate-conditioned freezing from conditioned suppression behavior in rats. AB - It is commonly assumed that suppression of an ongoing behavior is an indirect measure of freezing behavior. We tested whether conditioned suppression and freezing are the same or distinct conditioned responses. Rats were trained to press a bar for food and then given fear-conditioning sessions in which a tone was paired with a foot shock (two pairings a day for 2 days). They then received either sham or electrolytic lesions of the periaqueductal gray (PAG). Post training PAG lesions blocked freezing to the conditioned stimulus (CS), but had no effect on the suppression of operant behavior to the same CS. Thus, conditioned suppression and freezing, which both cause a cessation in activity, appear to be mediated by separate processes. PMID- 10541471 TI - Serotonin depletion does not prevent intrinsic sensitization in the leech. AB - Intrinsic sensitization is a form of behavioral facilitation that is distinct from the extrinsic sensitization normally studied. To examine whether intrinsic and extrinsic sensitization are mediated by different physiological processes, the effects of 5,7-dihydroxytryptamine-induced serotonin (5-HT) depletion on intrinsic sensitization of the leech whole-body shortening response were observed. Previous experiments have shown that 5-HT depletion disrupts dishabituation and extrinsic sensitization of this behavior in the leech. Intrinsic sensitization was observed in preparations from both control and 5-HT depleted animals, indicating that this form of behavioral facilitation was not affected by 5-HT depletion. The differences in the effects of 5-HT depletion on intrinsic versus extrinsic sensitization suggest that there are distinct neurophysiological processes mediating these two forms of behavioral facilitation. In addition, 5-HT depletion appeared to disrupt a putative extrinsic form of habituation of the shortening reflex. These data support the hypothesis that both intrinsic and extrinsic processes of neuromodulation mediate habituation and sensitization. PMID- 10541472 TI - Impaired learning and motor behavior in heterozygous Pafah1b1 (Lis1) mutant mice. AB - Heterozygous mutation or deletion of Pafab1b1 (LIS1) in humans is associated with syndromes with type 1 lissencephaly, a severe brain developmental disorder resulting from abnormal neuronal migration. We have created Lis1 heterozygous mutant mice by gene targeting. Heterozygous mutant mice are viable and fertile, but display global organizational brain defects as a result of impaired neuronal migration. To assess the functional impact of the mutation, Lis1 heterozygous mice and their wild-type littermates were evaluated on a wide variety of behavioral tests. Lis1 mutant mice displayed abnormal hindpaw clutching responses and were impaired on a rotarod test. Lis1 heterozygous mice were also impaired in the spatial learning version of the Morris water task. Impaired motor behavior and spatial learning and memory in Lis1 mutant mice indicates that impaired neuronal migration can have functional effects on complex behavioral responses. The behavioral findings also support the use of the Lis1 mutant mice as a model from human type 1 lissencephaly. PMID- 10541470 TI - Brain gene expression during REM sleep depends on prior waking experience. AB - In most mammalian species studied, two distinct and successive phases of sleep, slow wave (SW), and rapid eye movement (REM), can be recognized on the basis of their EEG profiles and associated behaviors. Both phases have been implicated in the offline sensorimotor processing of daytime events, but the molecular mechanisms remain elusive. We studied brain expression of the plasticity associated immediate-early gene (IEG) zif-268 during SW and REM sleep in rats exposed to rich sensorimotor experience in the preceding waking period. Whereas nonexposed controls show generalized zif-268 down-regulation during SW and REM sleep, zif-268 is upregulated during REM sleep in the cerebral cortex and the hippocampus of exposed animals. We suggest that this phenomenon represents a window of increased neuronal plasticity during REM sleep that follows enriched waking experience. PMID- 10541473 TI - Sarcoglycan complex: a muscular supporter of dystroglycan-dystrophin interplay? AB - In striated muscle, the cytoskeletal protein dystrophin, the protein product of the Duchenne muscular dystrophy gene, is associated with a number of sarcolemmal glycoproteins to form a large oligomeric complex, the dystrophin-glycoprotein complex (DGC). Over the last 10 years, four of these sarcolemmal glycoproteins, alpha-, beta-, gamma- and delta-sarcoglycans, have been shown to form a distinct subcomplex, the sarcoglycan complex, in the DGC. Furthermore, the genetic defects of alpha-, beta-, gamma- and delta-sarcoglycans have been identified as the causes of four distinct forms of muscular dystrophies, which are now collectively called sarcoglycanopathy. Current studies are beginning to focus on the biological functions of the sarcoglycan complex and the molecular mechanism by which its dysfunction leads to muscle cell degeneration. PMID- 10541474 TI - Semaphorins and their receptors in olfactory axon guidance. AB - The mammalian olfactory system is capable of discriminating among a large variety of odor molecules and is therefore essential for the identification of food, enemies and mating partners. The assembly and maintenance of olfactory connectivity have been shown to depend on the combinatorial actions of a variety of molecular signals, including extracellular matrix, cell adhesion and odorant receptor molecules. Recent studies have identified semaphorins and their receptors as putative molecular cues involved in olfactory pathfinding, plasticity and regeneration. The semaphorins comprise a large family of secreted and transmembrane axon guidance proteins, being either repulsive or attractive in nature. Neuropilins were shown to serve as receptors for secreted class 3 semaphorins, whereas members of the plexin family are receptors for class 1 and V (viral) semaphorins. The present review will discuss a role for semaphorins and their receptors in the establishment and maintenance of olfactory connectivity. PMID- 10541475 TI - An unexpected effect of an ouabain-sensitive ATPase activity on the amount of antigen-antibody complexes formed in situ. AB - A classical method of indirect immunofluorescence was applied on various kinds of lightly fixed and permeabilized cells to analyze the formation of the complexes between a nuclear antigen and its antibody (AAC). The amount of AAC decreased dramatically when the incubation with the first antibody was realized in the presence of ATP in a sodium-rich medium with 0.5 mM KCl. Addition of sodium vanadate, a general inhibitor of ATPases, ouabain or tetrabutylammonium ion, specific inhibitors of the Na+,K+-ATPase, prevented this effect. The established role of this enzyme is to increase free-K+ concentration and decrease free Na+ concentration in the cell. It is not surprising to find an ATPase still active since light fixation and permeabilization do not destroy phosphatases. But it is rather surprising to find something looking like Na+,K+-ATPase activity in permeabilized cells. The importance of potassium in this puzzling result is suggested by the fact that appearance of ACC was equally suppressed if the incubation was made in the absence of ATP in a potassium-rich medium without sodium. Results are discussed, taking into account the properties of cell associated water and recently found interrelation between Na+,K+-ATPase and tubulin. In any case, the results seem interesting in the field of immunocytochemistry. PMID- 10541476 TI - Long-term microstructural analyses of hydroxyapatite implanted in rats using laser-Raman spectrometry and scanning electron microscopy. AB - To investigate the long-term surface microstructure of a synthetic auditory ossicle (Apaceram) composed of dense hydroxyapatite (HA), thin HA disks were implanted subcutaneously into the interscapular regions of 12 rats. After 6, 14 and 20 months, implanted HA surfaces were observed using stereoscopic microscopy, scanning electron microscopy (SEM) and laser-Raman spectrometry. Visual observation by SEM at 6 months and by stereoscopic microscopy at 14 months indicated a progressive degradation of the HA disk surfaces implanted in the subcutaneous tissue. Visual observation by SEM at 14 and 20 months and by stereoscopic microscopy at 20 months indicated a progressive redeposition on the surfaces of the implants. Raman spectra compared half-peak breadths of v1 signal (PO4(3-), 960 cm(-1)) on the gray and white surface areas of implanted HA disks observed by stereoscopic microscopy. Analysis demonstrates that demineralization at 14 months and remineralization at 20 months occur on the gray areas; demineralization at 6 months and remineralization at 14 months occur on the white areas. PMID- 10541477 TI - Rapid aldosterone effects on tyrosine phosphorylation in vascular smooth muscle cells. AB - Non-genomic aldosterone effects are characterized by their rapid onset, their specificity for mineralocorticoids and their insensitivity both to the mineralocorticoid type 1 receptor antagonist spironolactone and to the inhibitors of transcription and translation, cycloheximide and actinomycin D. The aim of the present study was to further characterize the second messenger system involved in the non-genomic pathway of aldosterone with particular emphasis on protein phosphorylation. The rapid increase of free intracellular calcium by aldosterone in VSMC is sensitive to genistein, so that tyrosine kinase activity appears likely to be involved in the signaling pathway. Here, the effect of 100 nmol/l aldosterone (10 min.) on tyrosine protein-phosphorylation was determined in VSMC. Our findings show that aldosterone (100 nmol/l) in combination with shear stress as additional stimulus induces a rapid (within 10 min.) small but consistent increase in tyrosine-phosphorylation compared with aldosterone or shear stress alone. Immunoprecipitation of the MAPK-isoforms ERK 1 and ERK 2 showed an increased phosphorylation after 3 and 5 min. PMID- 10541478 TI - Liver and kidney peroxisomes in lactating rats and their pups after treatment with ciprofibrate. Biochemical and morphometric analysis. AB - We administered the hypolipidemic drug ciprofibrate to lactating rats and examined the enzymatic content and ultrastructural features of liver and kidney peroxisomes, both in treated animals and in their pups. The peroxisomal morphometric parameters, in particular, were measured in specimens submitted to the cytochemical reaction for the marker enzyme catalase. In liver of treated rats, the activities of peroxisomal enzymes involved in the fatty acid catabolism were significantly increased, while D-amino acid oxidase activity was lower than in controls; increments were also found in relative volume and pleiomorphism degree of the peroxisomal compartment, where a catalase dilution was supposed to occur. In the kidney, the treatment induced generalized increases of all examined enzymes; values significantly higher than controls were found in peroxisomal relative volume and numerical density, while the peroxisomal mean diameter practically did not change. The two organs, moreover, were affected by the drug in an age-dependent way, the pups being more responsive than the adults. The organ- and age-specific responses to the drug are interpreted as possibly related to the tissue-specific distribution of the peroxisomal proliferator activated receptor isotypes. PMID- 10541479 TI - Detection by immunohistochemistry of c-erbB2 oncoprotein in breast carcinomas and benign mammary lesions. AB - The presence of the c-erbB2 oncoprotein was demonstrated by immunohistochemistry in a study involving 173 mammary lesions. The lesions included infiltrating cancers, non-invasive neoplasia, as well as atypical and benign lesions. Our aim was to investigate the correlation between the c-erbB2 oncoprotein overexpression and the morphological features of the different mammary tissues analyzed to obtain a better characterization for the growth potential of certain lesions, with emphasis placed on the non-invasive neoplasia and the atypical lesions. Nearly 30% of infiltrating ductal carcinomas (27/89 cases) and 2 out of 24 infiltrating lobular carcinomas were positive. The comedocarcinomas were mostly stained (83%). In contrast, the intraductal carcinomas of cribriform or papillar patterns were consistently negative. No staining was observed in the atypical epithelial hyperplasia located in the vicinity of positive cancers for anti oncoprotein c-erbB2 antibody. Furthermore, the only 5 positive cases for c-erbB2 out of 32 cancer-free cases were three fibroadenomas and two fibrocystic diseases with atypical ductal hyperplasia. A close correlation was thus observed between c erbB2 oncoprotein overexpression and cancerous cell morphology, characterized by a marked nuclear hypertrophy often associated with cellular pleomorphism. However, predictive abnormalities of malignant transformation in non-neoplastic epithelial proliferation was difficult to identify, considering only the c-erbB2 expression. A group of tumors with little nuclear abnormalities were found positive for c-erbB2 immunostaining. These probably corresponded to a particular cellular phenotype. Further studies involving other oncogenes should lead to a better characterization of the different tumor phenotypes and help to clarify breast carcinogenesis. PMID- 10541480 TI - First searchable database for DNA profiles of human cell lines: sequential use of fingerprint techniques for authentication. AB - The authenticity and freedom from cross-contaminants of a cell line are important prerequisites for any research, development or production programs involving cell lines. Mini- and microsatellites in the human genome harboring variable-numbers of tandem repeat (VNTR) DNA markers allow individualization at the DNA level and are of practical value for genetic linkage mapping, forensic legal medicine, paternity testing, monitoring of bone marrow transplants, and individualization of established cell lines. We have validated fingerprint techniques of different single- and multiple-locus VNTRs enabling the establishment of a searchable database of DNA profiles. As a result, multiplexed polymerase chain reaction amplification fragment length polymorphism (AmpFLP) of four prominent and highly polymorphic minisatellite VNTR loci was proven as the best tool for screening the uniqueness of DNA profiles in a fingerprint database. In order to avoid false positivity, identical or similar DNA profiles based on AmpFLP VNTR were tested further using a multi-locus fingerprint system. Our data demonstrate that misidentification remains a chronic problem among human continuous cell lines (detailed information at URL http://www.dsmz.de). The combination of rapidly generated DNA profiles based on single-locus VNTR loci, their authentication by screening the fingerprint database, and confirmation of duplicate banding patterns using multilocus fingerprints constitute a highly reliable and robust method, which enables high fidelity and quality of maintenance independent from the quantity of individual cell lines. PMID- 10541481 TI - Supplementation with Lactobacillus reuteri or L. acidophilus reduced intestinal shedding of cryptosporidium parvum oocysts in immunodeficient C57BL/6 mice. AB - The effect of L. acidophilus supplementation to reduce fecal shedding of Cryptosporidium parvum oocysts was compared to L. reuteri using C57BL/6 female mice immunosuppressed by murine leukemia virus (strain LP-BM5) inoculation. After 12 weeks post LP-BM5 inoculation, 15 immunosuppressed mice each were randomly assinged to one of the following treatment groups: historical control (group A), LP-BM5 control (group B), C. parvum (group C), L. reuteri plus C. parvum (group D) or L. acidophilus plus C. parvum (group E). Mice were pre-fed the L. reuteri or L. acidophilus bacteria strains daily for 13 days, challenged with C. parvum oocysts and thereafter fed the specified Lactobacillus regimens daily during the experimental period. Animals supplemented with L. reuteri shed fewer (p<0.05) oocysts on day-7 post C. parvum challenge compared to controls. Mice supplemented with L. acidophilus also shed fewer (p<0.05) oocysts on days 7 and 14 post challenge compared to controls. Overall, Lactobacillus supplementation reduced C. parvum shedding in the feces but failed to suppress the production of T-helper type 2 cytokines [interleukin-4 (IL-4), IL-8)] which are associated with immunosuppression. Additionally, Lactobacillus supplementation did not restore T helper type 1 cytokines (interleukin-2 (IL-2) and gamma interferon (IFN-gamma), which are required for recovery from parasitic infections. Altered T-helper types 1 and 2 cytokine production as a consequence of immunodysfunction permitted the development of persistent cryptosporidiosis while mice with intact immune system were refractory to infection with C. parvum. Reduction in shedding of oocysts observed in the Lactobacillus supplemented mice during deminished IL-2 and IFN gamma production may be mediated by factors released into the intestinal lumen by the Lactobacillus and possibly other host cellular mechanisms. These observations suggest that L. reuteri or L. acidophilus can reduce C. parvum parasite burdens in the intestinal epithelium during cryptosporidiosis and may serve potential benefits as probiotics for host resistance to intestinal parasitic infections. L. acidophilus was more efficacious in reducing fecal shedding than L. reuteri and therefore may also have implication in the therapy of cryptosporidiosis during immunosuppressive states including human AIDS. PMID- 10541482 TI - Identification of altered DNA-protein interactions at the lamin A proximal promoter in quiescent hepatocytes. AB - The nuclear lamina of differentiated cells is composed of A-type and B-type lamins. The expression of the A-type lamins has been shown to be primarily under transcriptional control in proliferating cells. In order to gain insights into the factors involved in downregulation of the promoter in quiescent cells, we have compared the DNA-protein interactions at the rat lamin A proximal promoter in fetal (dividing) hepatocytes and adult (quiescent) hepatocytes by DNase I footprinting analysis, electrophoretic mobility shift assays and UV crosslinking experiments. We have identified proteins from adult hepatocyte nuclear extracts that are bound to sequences within 28 bp downstream of the transcription initiation site (62 and 48 kDa proteins) and within 20 bp upstream of the initiation site (54 kDa protein), which may function as transcriptional repressors of the lamin A proximal promoter. PMID- 10541483 TI - Effects of subclinical bovine paratuberculosis on in-vitro polymorphonuclear neutrophil migration. AB - Migration of polymorphonuclear neutrophils (PMNs) [unstimulated or stimulated with zymosan-activated serum (ZAS)] from 18 cows was measured in a microwell filter assay. Of these animals, 10 were subclinically infected with Mycobacterium paratuberculosis and shown by culture to be excreting the organism in the faeces; the remaining eight were clinically normal and negative for M. paratuberculosis on faecal culture. PMN "net migration" (stimulated minus unstimulated cells) of the infected cows was significantly lower than that of the uninfected cows. Migration of unstimulated cells in the infected cows did not differ from that in the uninfected cows. It would therefore appear that the infection influenced only the migratory response of the ZAS-stimulated cells. 1999 Harcourt Publishers Ltd. PMID- 10541484 TI - Revalidation is the answer. PMID- 10541485 TI - Dangerous people with severe personality disorder. British proposals for managing them are glaringly wrong-and unethical. PMID- 10541487 TI - Screening for osteoporosis. No point until we have resolved issues about long term treatment. PMID- 10541488 TI - How much to do at the accident scene? PMID- 10541486 TI - Meningococcal disease and healthcare workers. PMID- 10541490 TI - Scientists identify enzymes implicated in Alzheimer's PMID- 10541489 TI - Government orders inquiry as price of generic drugs soars. PMID- 10541491 TI - Surgeons were "fall guys" for a wider problem, inquiry told. PMID- 10541492 TI - Audit commission calls for improvements to critical care PMID- 10541493 TI - In brief PMID- 10541494 TI - Tobacco chiefs sell shares in their own company. PMID- 10541495 TI - Medicines control agency takes over GP research database. PMID- 10541496 TI - Nerve gas antidote a possible cause of gulf war illness. PMID- 10541497 TI - Euthanasia campaigner to stand in byelection. PMID- 10541498 TI - COX 2 inhibitors might be useful in cancer prevention PMID- 10541499 TI - Furby not guilty as "charged" PMID- 10541500 TI - Europe should give Alzheimer's disease higher priority. PMID- 10541501 TI - Radiologists accused in "scan scam". PMID- 10541502 TI - US research scientist found guilty of fraud PMID- 10541504 TI - Economic transition and changing relation between income inequality and mortality in Taiwan: regression analysis. AB - OBJECTIVE: To examine the changing relation between income inequality and mortality through different stages of economic development in Taiwan. DESIGN: Regression analysis of mortality on income inequality for three index years: 1976, 1985, and 1995. SETTING: 21 counties and cities in Taiwan. MAIN OUTCOME MEASURES: All age mortality and age specific mortality in children under age 5. RESULTS: When median household disposable income was controlled for, the association between income inequality and mortality became stronger in 1995 than in 1976. Especially, the association between income inequality and mortality in children aged under 5, with adjustment for differences in median household disposable income, changed from non-significant in 1976 to highly significant in 1995. In 1995, the level of household income after adjustment for income distribution no longer had a bearing on mortality in children under 5. CONCLUSION: The health of the population is affected more by relative income than by absolute income after a country has changed from a developing to a developed economy. PMID- 10541503 TI - Helicobacter pylori infection and early onset myocardial infarction: case-control and sibling pairs study. AB - OBJECTIVES: To examine the association between coronary heart disease and chronic Helicobacter pylori infection. DESIGN: Case-control study of myocardial infarction at young ages and study of sibling pairs with one member affected and the other not. SETTING: United Kingdom. PARTICIPANTS: 1122 survivors of suspected acute myocardial infarction at ages 30-49 (mean age 44 years) and 1122 age and sex matched controls with no history of coronary heart disease; 510 age and sex matched pairs of siblings (mean age 59 years) in which one sibling had survived myocardial infarction and one had no history of coronary heart disease. MAIN OUTCOME MEASURES: Serological evidence of chronic infection with H pylori. RESULTS: 472 (42%) of the 1122 cases with early onset myocardial infarction were seropositive for H pylori antibodies compared with 272 (24%) of the 1122 age and sex matched controls, giving an odds ratio of 2.28 (99% confidence interval 1.80 to 2.90). This odds ratio fell to 1.87 (1.42 to 2.47; P<0.0001) after smoking and indicators of socioeconomic status were adjusted for and to 1.75 (1.29 to 2.36) after additional adjustment for blood lipid concentrations and obesity. Only 158 of the 510 pairs of siblings were discordant for H pylori status; among these, 91 cases and 67 controls were seropositive (odds ratio 1.33 (0.86 to 2.05)). No strong correlations were observed between H pylori seropositivity and measurements of other risk factors for coronary heart disease (plasma lipids, fibrinogen, C reactive protein, albumin, etc). CONCLUSION: In the context of results from other relevant studies, these two studies suggest a moderate association between coronary heart disease and H pylori seropositivity that cannot be fully accounted for by other risk factors. But even if this association is causal and largely reversible by eradication of chronic infection, very large randomised trials would be needed to show this. PMID- 10541505 TI - Visual field defect associated with vigabatrin: observational cohort study. PMID- 10541507 TI - Email submissions from outside the united kingdom PMID- 10541506 TI - Association of variant alleles of mannose binding lectin with severity of pulmonary disease in cystic fibrosis: cohort study. PMID- 10541508 TI - Influence on general practitioners of teaching undergraduates: qualitative study of London general practitioner teachers. AB - OBJECTIVE: To examine the perceived effect of teaching clinical skills and associated teacher training programmes on general practitioners' morale and clinical practice. DESIGN: Qualitative semistructured interview study. SETTING: General practices throughout north London. SUBJECTS: 30 general practitioners who taught clinical skills were asked about the effect of teaching and teacher training on their morale, confidence in clinical and teaching skills, and clinical practice. RESULTS: The main theme was a positive effect on morale. Within teacher training this was attributed to developing peer and professional support; improved teaching skills; and revision of clinical knowledge and skills. Within teaching this was attributed to a broadening of horizons; contact with enthusiastic students; increased time with patients; improved clinical practice; improved teaching skills; and an improved image of the practice. Problems with teaching were due to external factors such as lack of time and space and anxieties about adequacy of clinical cover while teaching. CONCLUSION: Teaching clinical skills can have a positive effect on the morale of general practitioner teachers as a result of contact with students and peers, as long as logistic and funding issues are adequately dealt with. PMID- 10541509 TI - Reliability of patients measuring blood pressure at home: prospective observational study. PMID- 10541511 TI - ABC of complementary medicine. The manipulative therapies: osteopathy and chiropractic. PMID- 10541510 TI - Urinary tract infection in children. PMID- 10541512 TI - Take a lodger PMID- 10541513 TI - Revalidation in the United Kingdom: general principles based on experience in general practice. PMID- 10541514 TI - Recertification in the United States. PMID- 10541516 TI - Revalidation of doctors in Canada. PMID- 10541515 TI - Revalidation in Australia and New Zealand: approach of Royal Australasian College of Physicians. PMID- 10541517 TI - Reregistration of medical specialists in the Netherlands. PMID- 10541519 TI - What is health? PMID- 10541520 TI - Treatment of shock PMID- 10541518 TI - Staffing of hospitals: future needs, future provision. PMID- 10541521 TI - People who condemn eugenics may be in minority now. PMID- 10541522 TI - Treatment of toenail onychomycosis. Will Paper's key message soon appear in promotional material for drug? PMID- 10541523 TI - CT scanning can differentiate between ischaemic attack and haemorrhage. PMID- 10541524 TI - Getting letters published in journals is good aim for medical students. PMID- 10541525 TI - In general practice Scottish NHS and English NHS differ considerably. PMID- 10541526 TI - Study confirms tendency towards lower risk of myocardial infarction with second generation oral contraceptives in UK. PMID- 10541527 TI - Numbers needed to treat derived from meta-analysis. Are an absurdity. PMID- 10541528 TI - Cholesterol lowering margarine is effective. PMID- 10541529 TI - Trends in emergency admissions. Shop floor experience suggests a rise. PMID- 10541530 TI - Effect of supplementation with vitamin A or beta carotene on mortality related to pregnancy. Pooling of groups may not be appropriate. PMID- 10541532 TI - When I use a word. "Liar" can indeed be found in dictionary PMID- 10541531 TI - Human rights are based on consensus. PMID- 10541534 TI - Sixty one PCTs could be in place in 2000 PMID- 10541533 TI - Desmond bardon PMID- 10541535 TI - Infections and human cancer: cancer surveys, volume 33; microbes and malignancy: infection as a cause of human cancers PMID- 10541536 TI - Poems PMID- 10541537 TI - Selling drugs to consumers PMID- 10541538 TI - Drug advertising PMID- 10541539 TI - The blues and pellagra: a public health detective story PMID- 10541541 TI - Revalidating and rethinking PMID- 10541542 TI - H pylori is weakly associated with heart disease PMID- 10541540 TI - Have you heard who's on ward 3? PMID- 10541543 TI - Association between income distribution and health changes with level of economic development PMID- 10541544 TI - Teaching undergraduates affects general practitioners' morale positively PMID- 10541545 TI - Patients probably measure their own blood pressure accurately PMID- 10541546 TI - Making a difference to patient care is the challenge for all methods of recertification PMID- 10541547 TI - ORC binding, gene amplification, and the nature of metazoan replication origins. PMID- 10541548 TI - X-inactivation by chromosomal pairing events. PMID- 10541549 TI - XRCC3 promotes homology-directed repair of DNA damage in mammalian cells. AB - Homology-directed repair of DNA damage has recently emerged as a major mechanism for the maintenance of genomic integrity in mammalian cells. The highly conserved strand transferase, Rad51, is expected to be critical for this process. XRCC3 possesses a limited sequence similarity to Rad51 and interacts with it. Using a novel fluorescence-based assay, we demonstrate here that error-free homology directed repair of DNA double-strand breaks is decreased 25-fold in an XRCC3 deficient hamster cell line and can be restored to wild-type levels through XRCC3 expression. These results establish that XRCC3-mediated homologous recombination can reverse DNA damage that would otherwise be mutagenic or lethal. PMID- 10541550 TI - Drosophila ORC specifically binds to ACE3, an origin of DNA replication control element. AB - In the yeast Saccharomyces cerevisiae, sequence-specific DNA binding by the origin recognition complex (ORC) is responsible for selecting origins of DNA replication. In metazoans, origin selection is poorly understood and it is unknown whether specific DNA binding by metazoan ORC controls replication. To address this problem, we used in vivo and in vitro approaches to demonstrate that Drosophila ORC (DmORC) binds to replication elements that direct repeated initiation of replication to amplify the Drosophila chorion gene loci in the follicle cells of egg chambers. Using immunolocalization, we observe that ACE3, a 440-bp chorion element that contains information sufficient to drive amplification, directs DmORC localization in follicle cells. Similarly, in vivo cross-linking and chromatin immunoprecipitation assays demonstrate association of DmORC with both ACE3 and two other amplification control elements, AER-d and ACE1. To demonstrate that the in vivo localization of DmORC is related to its DNA binding properties, we find that purified DmORC binds to ACE3 and AER-d in vitro, and like its S. cerevisiae counterpart, this binding is dependent on ATP. Our findings suggest that sequence-specific DNA binding by ORC regulates initiation of metazoan DNA replication. Furthermore, adaptation of this experimental approach will allow for the identification of additional metazoan ORC DNA-binding sites and potentially origins of replication. PMID- 10541551 TI - Crystal structure of an OCA-B peptide bound to an Oct-1 POU domain/octamer DNA complex: specific recognition of a protein-DNA interface. AB - We have determined the crystal structure, at 3.2 A, of a ternary complex containing an OCA-B peptide, the Oct-1 POU domain, and an octamer DNA site. The OCA-B peptide binds in the major groove near the center of the octamer site, and its polypeptide backbone forms a pair of hydrogen bonds with the adenine base at position 5 of the octamer DNA. Numerous protein-protein contacts between the OCA B peptide and the POU domain are also involved in the ternary complex. In particular, the hydrophobic surface from a short alpha-helix of OCA-B helps to stabilize the complex by binding to a hydrophobic pocket on the POU-specific domain. The structure of this ternary complex is consistent with previous biochemical studies and shows how peptide-DNA and peptide-protein contacts from OCA-B provide structural and functional specificity in the regulation of immunoglobulin transcription. PMID- 10541553 TI - INK4a/ARF mutations accelerate lymphomagenesis and promote chemoresistance by disabling p53. AB - The INK4a/ARF locus encodes upstream regulators of the retinoblastoma and p53 tumor suppressor gene products. To compare the impact of these loci on tumor development and treatment response, the Emu-myc transgenic lymphoma model was used to generate genetically defined tumors with mutations in the INK4a/ARF, Rb, or p53 genes. Like p53 null lymphomas, INK4a/ARF null lymphomas formed rapidly, were highly invasive, displayed apoptotic defects, and were markedly resistant to chemotherapy in vitro and in vivo. Furthermore, INK4a/ARF(-/-) lymphomas displayed reduced p53 activity despite the presence of wild-type p53 genes. Consequently, INK4a/ARF and p53 mutations lead to aggressive tumors by disrupting overlapping tumor suppressor functions. These data have important implications for understanding the clinical behavior of human tumors. PMID- 10541552 TI - Disruption of the ARF-Mdm2-p53 tumor suppressor pathway in Myc-induced lymphomagenesis. AB - Transgenic mice expressing the c-Myc oncogene driven by the immunoglobulin heavy chain enhancer (Emu) develop B-cell lymphoma and exhibit a mean survival time of approximately 6 months. The protracted latent period before the onset of frank disease likely reflects the ability of c-Myc to induce a p53-dependent apoptotic program that initially protects animals against tumor formation but is disabled when overtly malignant cells emerge. In cultured primary mouse embryo fibroblasts, c-Myc activates the p19(ARF)-Mdm2-p53 tumor suppressor pathway, enhancing p53-dependent apoptosis but ultimately selecting for surviving immortalized cells that have sustained either p53 mutation or biallelic ARF deletion. Here we report that p53 and ARF also potentiate Myc-induced apoptosis in primary pre-B-cell cultures, and that spontaneous inactivation of the ARF-Mdm2 p53 pathway occurs frequently in tumors arising in Emu-myc transgenic mice. Many Emu-myc lymphomas sustained either p53 (28%) or ARF (24%) loss of function, whereas Mdm2 levels were elevated in others. Its overexpression in some tumors lacking p53 function raises the possibility that Mdm2 can contribute to lymphomagenesis by interacting with other targets. Emu-myc transgenic mice hemizygous for ARF displayed accelerated disease (11-week mean survival), and 80% of these tumors lost the wild-type ARF allele. All ARF-null Emu-myc mice died of lymphoma within a few weeks of birth. About half of the tumors arising in ARF hemizygous or ARF nullizygous Emu-myc transgenic mice also overexpressed Mdm2. Therefore, Myc activation strongly selects for spontaneous inactivation of the ARF-Mdm2-p53 pathway in vivo, cancelling its protective checkpoint function and accelerating progression to malignancy. PMID- 10541554 TI - Bmi-1 collaborates with c-Myc in tumorigenesis by inhibiting c-Myc-induced apoptosis via INK4a/ARF. AB - The bmi-1 and myc oncogenes collaborate strongly in murine lymphomagenesis, but the basis for this collaboration was not understood. We recently identified the ink4a-ARF tumor suppressor locus as a critical downstream target of the Polycomb group transcriptional repressor Bmi-1. Others have shown that part of Myc's ability to induce apoptosis depends on induction of p19arf. Here we demonstrate that down-regulation of ink4a-ARF by Bmi-1 underlies its ability to cooperate with Myc in tumorigenesis. Heterozygosity for bmi-1 inhibits lymphomagenesis in Emu-myc mice by enhancing c-Myc-induced apoptosis. We observe increased apoptosis in bmi-1(-/-) lymphoid organs, which can be rescued by deletion of ink4a-ARF or overexpression of bcl2. Furthermore, Bmi-1 collaborates with Myc in enhancing proliferation and transformation of primary embryo fibroblasts (MEFs) in an ink4a ARF dependent manner, by prohibiting Myc-mediated induction of p19arf and apoptosis. We observe strong collaboration between the Emu-myc transgene and heterozygosity for ink4a-ARF, which is accompanied by loss of the wild-type ink4a ARF allele and formation of highly aggressive B-cell lymphomas. Together, these results reinforce the critical role of Bmi-1 as a dose-dependent regulator of ink4a-ARF, which on its turn acts to prevent tumorigenesis on activation of oncogenes such as c-myc. PMID- 10541556 TI - Recruitment of Nanos to hunchback mRNA by Pumilio. AB - Translational regulation of hunchback (hb) mRNA is essential for posterior patterning of the Drosophila embryo. This regulation is mediated by sequences in the 3'-untranslated region of hb mRNA (the Nanos response elements or NREs), as well as two trans-acting factors-Nanos and Pumilio. Pumilio recognizes the NREs via a conserved binding motif. The mechanism of Nanos action has not been clear. In this report we use protein-protein and protein-RNA interaction assays in yeast and in vitro to show that Nanos forms a ternary complex with the RNA-binding domain of Pumilio and the NRE. Mutant forms of the NRE, Nos, and Pum that do not regulate hb mRNA normally in embryos do not assemble normally into a ternary complex. In particular, recruitment of Nos is dependent on bases in the center of the NRE, on the carboxy-terminal Cys/His domain of Nos, and on residues in the eighth repeat of the Pum RNA-binding domain. These residues differ in a closely related human protein that also binds to the NRE but cannot recruit Drosophila Nos. Taken together, these findings suggest models for how Nos and Pum collaboratively target hb mRNA. More generally, they suggest that Pum-like proteins from other species may also act by recruiting cofactors to regulate translation. PMID- 10541555 TI - Functional antagonism of the Polycomb-Group genes eed and Bmi1 in hemopoietic cell proliferation. AB - The murine Polycomb-Group (PcG) proteins Eed and Bmi1 govern axial patterning during embryonic development by segment-specific repression of Hox gene expression. The two proteins engage in distinct multimeric complexes that are thought to use a common molecular mechanism to render the regulatory regions of Hox and other downstream target genes inaccessible to transcriptional activators. Beyond axial patterning, Bmi1 is also involved in hemopoiesis because a loss-of function allele causes a profound decrease in bone marrow progenitor cells. Here, evidence is presented that is consistent with an antagonistic function of eed and Bmi1 in hemopoietic cell proliferation. Heterozygosity for an eed null allele causes marked myelo- and lymphoproliferative defects, indicating that eed is involved in the negative regulation of the pool size of lymphoid and myeloid progenitor cells. This antiproliferative function of eed does not appear to be mediated by Hox genes or the tumor suppressor locus p16(INK4a)/p19(ARF) because expression of these genes was not altered in eed mutants. Intercross experiments between eed and Bmi1 mutant mice revealed that Bmi1 is epistatic to eed in the control of primitive bone marrow cell proliferation. However, the genetic interaction between the two genes is cell-type specific as the presence of one or two mutant alleles of eed trans-complements the Bmi1-deficiency in pre-B bone marrow cells. These studies thus suggest that hemopoietic cell proliferation is regulated by the relative contribution of repressive (Eed-containing) and enhancing (Bmi1-containing) PcG gene complexes. PMID- 10541557 TI - A large-scale insertional mutagenesis screen in zebrafish. AB - It is estimated that approximately 2500 genes are essential for the normal development of a zebrafish embryo. A mutation in any one of these genes can result in a visible developmental defect, usually followed by the death of the embryo or larva by days 5-7 of age. We are performing a large-scale insertional mutagenesis screen in the zebrafish with the goal of isolating approximately 1000 embryonic mutations. We plan to clone a significant fraction of the mutated genes, as these are the genes important for normal embryogenesis of a vertebrate. To achieve this goal, we prepared approximately 36, 000 founder fish by injecting blastula-stage embryos with one of two pseudotyped retroviruses. We estimate that together these fish harbor between 500,000-1,000,000 proviral insertions in their germ lines. The protocol we have devised and the size of our facility allow us to breed approximately 80,000-150,000 of these insertions to homozygosity within 2 years. Because a pilot screen conducted earlier in our laboratory revealed that the frequency of mutations obtained with this type of insertional mutagen is 1 embryonic lethal mutation per 70-100 proviral insertions, screening 100,000 insertions should yield at least 1000 mutants. Here we describe the protocol for the screen and initial results with the first of the two retroviral vectors used, a virus designated F(5). We screened an estimated 760 insertions among F(3) progeny from 92 F(2) families and obtained 9 recessive embryonic lethal mutations. Thus, the efficiency of mutagenesis with this viral vector is approximately one-ninth that observed with the chemical mutagen ENU in zebrafish. We have also obtained two dominant mutations, one of which is described here. As expected, mutated genes can be readily identified. So far, genes mutated in four of the nine recessive mutants and one of the two dominant mutants have been cloned. Further improvements to this technology could make large-scale insertional mutagenesis screening and rapid gene cloning accessible to relatively small zebrafish laboratories. PMID- 10541558 TI - Organization and dynamics of the Mu transpososome: recombination by communication between two active sites. AB - Movement of transposable genetic elements requires the cleavage of each end of the element genome and the subsequent joining of these cleaved ends to a new target DNA site. During Mu transposition, these reactions are catalyzed by a tetramer of four identical transposase subunits bound to the paired Mu DNA ends. To elucidate the organization of active sites within this tetramer, the subunit providing the essential active site DDE residues for each cleavage and joining reaction was determined. We demonstrate that recombination of the two Mu DNA ends is catalyzed by two active sites, where one active site promotes both cleavage and joining of one Mu DNA end. This active site uses all three DDE residues from the subunit bound to the transposase binding site proximal to the cleavage site on the other Mu DNA end (catalysis in trans). In addition, we uncover evidence that the catalytic activity of these two active sites is coupled such that the coordinated joining of both Mu DNA ends is favored during recombination. On the basis of these results, we propose that the DNA joining stage requires a cooperative transition within the transposase-DNA complex. The cooperative utilization of active sites supplied in trans by Mu transposase provides an example of how mobile elements can ensure concomitant recombination of distant DNA sites. PMID- 10541560 TI - Regulation of sulphotransferase expression in the endometrium during the menstrual cycle, by oral contraceptives and during early pregnancy. AB - The endometrium plays a key role in reproduction, and this function is tightly regulated by endogenous and xenobiotic steroids. Sulphation, catalysed by members of the sulphotransferase (SULT) enzyme family, is a major deactivating mechanism for steroid hormones and we have investigated the expression and regulation in vivo of SULT in the human endometrium. In the normal cycling endometrium, expression of the phenol sulphotransferases SULT1A1 and SULT1A3 and the oestrogen sulphotransferase SULT1E1 were observed, with SULT1A1 and SULT1E1 expression being higher in the luteal phase than in the follicular phase. No expression of the hydroxysteroid sulphotransferase SULT2A1 was detected at any time in the endometrium. In endometrium from women taking the combined oral contraceptive pill (OCP), SULT1E1 expression was virtually absent, and SULT1A1 expression was substantially reduced. Similarly, in early pregnancy (i.e. first trimester) endometrium, SULT1E1 expression was absent, although SULT1A1 and SULT1A3 expression were unaffected. Our results with normal endometrium support in-vitro data showing that SULT1E1 expression is regulated by progesterone. However, the data obtained from OCP and early pregnancy endometrium suggest that factors other than the concentration of circulating progesterone are involved in the regulation of the expression of this important enzyme in the endometrium. PMID- 10541561 TI - Insulin and insulin-like growth factor-I and -II modulate human granulosa-lutein cell steroidogenesis: enhancement of steroidogenic acute regulatory protein (StAR) expression. AB - Insulin and insulin-like growth factors (IGF)-I and -II stimulate granulosa cell steroidogenesis. Since steroidogenic acute regulatory protein (StAR) regulates the rate-limiting step in steroid hormone biosynthesis, the ability of insulin and IGF to modulate StAR protein and mRNA expression was examined in two human granulosa cell culture systems: (i) proliferating granulosa-lutein cells and (ii) luteinized-granulosa cells derived during in-vitro fertilization (IVF). In proliferating granulosa-lutein cells, IGF-I and IGF-II increased StAR protein approximately 4-5-fold, while insulin and 8-bromoadenosine 3',5'-cAMP (8-Br-cAMP) increased amounts of StAR protein 2.5- and 6-fold respectively. A combination of IGFs/insulin and 8-Br-cAMP increased StAR 7-9-fold. Luteinized granulosa cells also had increased StAR expression after treatment with IGF-I (2. 8-fold), IGF-II (3-fold), insulin (2.5-fold) and 8-Br-cAMP (4. 5-fold). Progesterone production generally followed a pattern similar to StAR protein in both cell systems. In proliferating granulosa-lutein cells, both IGF-I and insulin increased StAR mRNA (3-fold) and 8-Br-cAMP increased StAR mRNA 4-fold, whereas a combination of IGF-I and 8-Br-cAMP had an additive effect on StAR mRNA expression (7-fold). Transient transfection of proliferating granulosa-lutein cells with StAR promoter luciferase reporter constructs demonstrated that IGF-I, IGF-II, and insulin had no effect on the StAR promoter activity, while 8-Br-cAMP stimulated StAR promoter function. The results indicate that: (i) IGFs and insulin stimulate StAR mRNA and protein expression in human granulosa-lutein cells; (ii) IGF-I and 8-Br-cAMP have an additive effect on StAR gene expression; and (iii) IGF-I increases StAR mRNA and protein by a mechanism that does not involve activation of the proximal StAR gene promoter. PMID- 10541559 TI - Tagging chromatin with retrotransposons: target specificity of the Saccharomyces Ty5 retrotransposon changes with the chromosomal localization of Sir3p and Sir4p. AB - Retrotransposon and retroviral insertions are not randomly distributed on chromosomes, suggesting that retroelements actively select integration sites. This is the case for the yeast Ty5 retrotransposons, which preferentially integrate into domains of silent chromatin at the HM loci and telomeres. Here we demonstrate that loss of Sir3p or Sir4p-components of silent chromatin-causes a greater than ninefold decrease in Ty5 targeting to the HM loci and largely randomizes chromosomal integration patterns. Strains with a deletion of SIR4 also display an approximately 10-fold increase in cDNA recombination, which is due both to the expression a- and alpha-mating-type information and the loss of Sir4p. It is known that in old yeast cells or in strains carrying the sir4-42 allele, the Sir complex relocalizes to the rDNA. About 26% of Ty5 insertions occur within the rDNA in sir4-42 strains compared with 3% in wild type. Ty5, therefore, is sensitive to changes in chromatin, indicating that retrotransposons may be useful for dissecting chromatin dynamics that occur during developmental programs such as aging. PMID- 10541562 TI - The involvement of nitric oxide in corpus luteum regression in the rat: feedback mechanism between prostaglandin F(2alpha) and nitric oxide. AB - In the corpus luteum (CL), prostaglandin F(2alpha) (PGF(2alpha)) is a physiological agent with luteolytic actions. Nitric oxide (NO) is a messenger molecule capable of modulating diverse pathophysiological processes. The aim of the present study was to investigate the role of ovarian NO in PGE (a luteotrophic prostanoid) and PGF(2alpha) (a luteolytic prostanoid) production and in progesterone synthesis during CL regression in the rat. To obtain a longer functional CL, we used a pseudopregnant (PSP) rat model. By means of intrabursa ovarian sac treatment of two competitive nitric oxide synthase (NOS) inhibitors, N(G)-monomethyl-L-arginine (L-NMMA, 1 mg/kg) and N(W)-nitro-L-arginine methyl ester (L-NAME; 3 mg/kg), and sodium nitroprusside (SNP, 0.05 mg/kg) as a NO generator, we found that NO, produced by the ovarian tissue during the last 2 days of CL development (days 8 and 9), increased PGF(2alpha) production in the ovary and diminished serum progesterone concentrations leading to CL involution. We also proposed a positive feedback mechanism between PGF(2alpha) and NO, to ensure luteal regression. Thus, we injected intraperitoneally a luteolytic dose (3 microg/kg) of a synthetic PGF(2alpha) during the mid and late phase of CL development. Ovarian NOS activity was evaluated. The results confirmed our hypothesis; we did not see any effect in the mid-stage of CL development, but increased ovarian NOS activity was found in PGF(2alpha)-injected late pseudopregnant rats. PMID- 10541563 TI - Regulation of human sperm capacitation by a cholesterol efflux-stimulated signal transduction pathway leading to protein kinase A-mediated up-regulation of protein tyrosine phosphorylation. AB - Protein tyrosine phosphorylation is an important intracellular event accompanying the in-vitro capacitation of mouse, bovine and human spermatozoa. Here, we demonstrate that bovine serum albumin (BSA) and NaHCO(3) are required for protein tyrosine phosphorylation in ejaculated human spermatozoa. The absence of protein tyrosine phosphorylation in media minus these two constituents could be recovered by addition to the media of cAMP analogues and/or phosphodiesterase inhibitors. Since BSA is postulated to modulate capacitation by removal of cholesterol from the sperm plasma membrane, we determined whether cholesterol release leads to changes in protein tyrosine phosphorylation. Incubation of spermatozoa in media containing BSA resulted in the release of significant amounts of cholesterol when compared with media devoid of BSA. Preloading BSA with cholesterol-SO(4) inhibited protein tyrosine phosphorylation, as well as capacitation, and this inhibitory effect was overcome by the addition of dibutyryl cAMP plus isobutylmethylxanthine (IBMX). The functional significance of BSA-mediated cholesterol release, protein tyrosine phosphorylation and capacitation was confirmed by examining the effects of the cholesterol-binding heptasaccharides, methyl-beta-cyclodextrin or OH-propyl-beta-cyclodextrin. Both cyclodextrins caused cholesterol efflux from the spermatozoa, increased protein tyrosine phosphorylation, and stimulated capacitation. Therefore, cholesterol release is associated with the activation of a signal transduction pathway involving protein kinase A and tyrosine kinase second messenger systems, and resulting in protein tyrosine phosphorylation and capacitation. PMID- 10541565 TI - Analysis of gene expression in single oocytes and embryos by real-time rapid cycle fluorescence monitored RT-PCR. AB - Rapid cycle DNA amplification is a refinement of the polymerase chain reaction (PCR) method that permits increased product specificity while reducing amplification time by an order of magnitude. Combined with the use of micro volume capillaries, minute samples can be examined by this technique. Thus, this approach is ideally suited to the analysis of gene expression in individual cells. As the current understanding of early developmental processes is still rudimentary, further characterization of transcription in single oocytes and embryos may provide additional insight into the molecular mechanisms directing these events. In this study, we examined the suitability of fluorescence monitored reverse transcription (RT)-PCR for the study of gene expression during oogenesis and embryogenesis using transcripts of the housekeeping gene, beta actin, as an experimental model. Product accumulation was monitored by either the double-stranded DNA dye SYBR Green I or sequence-dependent hybridization of reporter molecules called molecular beacons. Dyes bind generically and are economical to use. However, both specific and non-specific products are labelled. Hybridization probes permit very specific and sensitive target recognition but they can be costly to manufacture. Once molecular markers indicative of optimal development are identified, this technology could be used in a clinical in-vitro fertilization laboratory as a diagnostic tool. PMID- 10541564 TI - PI-PLC releases a 25-40 kDa protein cluster from the hamster oolemma and affects the sperm penetration assay. AB - The effects of phosphatidylinositol-specific phospholipase C (PI-PLC) on human sperm-hamster oocyte interaction were investigated to determine whether PI-PLC cleavable glycosylphosphatidyinositol (GPI)-anchored proteins are involved in sperm-egg binding and fusion. Two-dimensional electrophoresis was then utilized to visualize proteins released from hamster oocytes following PI-PLC treatment. For the binding and fusion assay, either spermatozoa or eggs were treated with 1 IU/ml PI-PLC for 30 min and washed prior to gamete co-incubation. Treatment of human spermatozoa with PI-PLC significantly (P /=140 and/or diastolic blood pressure >/=90 mmHg). After MIBG examination, blood pressure was measured regularly in all NIDDM patients. There were significant differences between 65 normotensive and 17 hypertensive NIDDM patients with respect to age (55+/-11 vs 63+/-12 years, respectively, P<0.05), prevalence of diabetic retinopathy (12% vs 35%, respectively, P<0.05) and systolic blood pressure (120+/-12 vs 145+/-16 mmHg, respectively, P<0.001). The H/M ratio in hypertensive NIDDM patients was significantly lower than in the control group (1. 81+/-0.29 vs 2.27+/-0.20, respectively, P<0.01). During the follow-up period (18+/- 12 months), 17 NIDDM patients newly developed hypertension after MIBG examination. There were no significant differences in their clinical characteristics compared with persistently normotensive or hypertensive NIDDM patients. %WR in patients with new onset hypertension was significantly higher than in the control group (30.88%+/-16.87% vs 12.89%+/-11.94%, respectively, P<0.05). Moreover, in these patients %WR correlated with duration from the date of MIBG scintigraphy to the onset of hypertension (r=-0.512, P<0.05). Five NIDDM patients died during the follow-up period (four newly hypertensive patients and one normotensive patient). There were significant statistical differences between the control group and non survivors in terms of age (54+/-11 vs 73+/-11 years, respectively, P<0.01), H/M ratio (2. 27+/- 0.20 vs 1.64+/-0.36, respectively, P<0.01) and %WR (12. 89%+/ 11.94% vs 42.52%+/-22.39%, respectively, P<0.01). In conclusion, cardiac sympathetic denervation using MIBG scintigraphy observed in hypertensive NIDDM patients, and was more profound in non-survivors. MIBG scintigraphy proved useful for the evaluation of NIDDM patients with new onset hypertension, and it was found that NIDDM patients with abnormalities on MIBG scintigraphy needed to be observe carefully. PMID- 10541831 TI - Contribution of cholescintigraphy to the early diagnosis of acute acalculous cholecystitis in intensive-care-unit patients. AB - Thirty-two intensive care unit patients (78% on long-term total parenteral nutrition) suspected of having acute acalculous cholecystitis (AAC) were studied prospectively. All of these patients underwent abdominal ultrasonography and cholescintigraphy with technetium-99m mebrofenin. Morphine sulphate (0.04 mg/kg) was administered only if the gallbladder was not visualised after 1 h (16 patients). The final diagnosis was reached after clinical improvement, or upon the discovery of another aetiology for the symptoms presented, or on the basis of histopathology following cholecystectomy (when this was performed). We analysed the contribution of individual cholescintigraphic findings (I: non-visualisation of the gallbladder during the first 60 min of the examination; II: persistent non visualisation of the gallbladder 30 min following morphine administration; III: non-visualisation of the small bowel for at least 90 min) and their various combinations. We obtained a sensitivity of 79% and a specificity rate 100% using the interpretative criteria "I and II or III". Excluding obstructive syndrome ("I and II"), the sensitivity and specificity figures were 70% and 100% respectively (28 patients). We had no false-positive results in our patient population. Cholescintigraphy was found to complement ultrasonography, which had either good sensitivity (93%) and poor specificity (17%), when at least two of the three major signs were present (sludge, thickened wall, gallbladder distension), or poor sensitivity (36%) and good specificity (89%) when all three signs were present. We conclude that cholescintigraphy is a useful tool for early diagnosis of AAC in critically ill patients, in whom ultrasonography alone does not provide enough information to permit a sufficiently early decision regarding the use of surgery. PMID- 10541832 TI - Efficacy of iodine-131 6beta-methyl-iodo-19-norcholesterol scintigraphy and computed tomography in patients with primary aldosteronism. AB - In order to define the role of scintigraphy in determining the aetiology of primary aldosteronism, 41 patients were examined by computed tomography (CT) scan and adrenal scintigraphy using iodine-131 6beta-methyl-iodo-19-norcholesterol with the dexamethasone suppression test. Hormonal and scintigraphic examinations were conducted while avoiding interference by medical treatment. The aetiological diagnosis was established by taking account of the clinical context, the endocrine profile, and CT scan and scintigraphic data, as well as possible hormone assays after catheterization of the adrenal veins (12 cases) and postoperative pathology data (14 cases). The aetiological diagnoses established were Conn's adenoma (insensitive to angiotensin II) in 12 cases, idiopathic hyperplasia in 11 and macronodular hyperplasia (with functional autonomy of the nodules) in 18. Unilateral and bilateral lesions were correctly distinguished by scintigraphy in 92% of cases as compared with only 58% using CT scan alone; this was because the CT scan appearance was normal in 3/12 cases of adenoma and because a single nodule was visible in 2/11 cases of idiopathic hyperplasia and in 12/18 cases of macronodular hyperplasia. It is concluded that scintigraphy using noriodocholesterol with the dexamethasone suppression test should be performed systematically in conjunction with hormonal tests and adrenal CT scan in all cases of primary aldosteronism, as part of a strategy aimed not only at detecting adenoma but also at determining whether the hyperfunctional lesions are bilateral. PMID- 10541833 TI - An alternative approach to estimation of the brain perfusion index for measurement of cerebral blood flow using technetium-99m compounds. AB - Cerebral blood flow (CBF) has been quantified non-invasively using the brain perfusion index (BPI) determined from radionuclide angiographic data generated by technetium-99m hexamethylpropylene amine oxime( )((99m)Tc-HMPAO) or technetium 99m ethyl cysteinate dimer( )((99m)Tc-ECD). The BPI is generally calculated using graphical analysis (GA). In the present study, BPI was measured using spectral analysis (SA), and its usefulness evaluated in comparison with GA. The BPI was calculated from the sum of spectral data obtained by SA. We applied this method to radionuclide angiographic data collected from the bilateral brain hemispheres of 20 patients with various brain diseases using (99m)Tc-HMPAO and from those of 20 patients using (99m)Tc-ECD. We also measured BPI using GA. The BPI values obtained by SA (BPI(S)) (x) and by GA (BPI(G)) (y) correlated closely (y=0.708x+0.038, r=0.945 for (99m)Tc-HMPAO and y=0.559x+0.093, r=0.931 for (99m)Tc-ECD). However, the BPI(G) values were underestimated by 22.9%+/-6.6% (mean+/-SD) for (99m)Tc-HMPAO and by 27.9%+/-7.5% for (99m)Tc-ECD as compared with the BPI(S) values. The extent of underestimation tended to increase with increasing BPI(S) values. These findings were considered to be a result of the BPI(G) values being affected by the first-pass extraction fraction of the tracer. We also compared the BPI(S) and BPI(G) values with those of CBF measured using N isopropyl-p-[(123)I]iodoamphetamine (CBF(IMP)) in 16 patients (six for (99m)Tc HMPAO and ten for (99m)Tc-ECD). Although both BPI(S) and BPI(G) values correlated significantly with the CBF(IMP) values, the correlation coefficient in BPI(S) was always better than that in BPI(G) (r=0.869 for (99m)Tc-HMPAO and r=0.929 for (99m)Tc-ECD in BPI(S), r=0.629 for (99m)Tc-HMPAO and r=0.856 for (99m)Tc-ECD in BPI(G)). These results suggest that SA can provide a more reliable BPI for quantifying CBF using (99m)Tc-HMPAO or (99m)Tc-ECD than the conventional method using GA. Our method will be useful especially when using a tracer with a low first-pass extraction fraction and/or when performing activation studies using pharmacological intervention. PMID- 10541834 TI - Glucose utilisation in the lungs of septic rats. AB - Sequestration and degranulation of leucocytes in the pulmonary microcirculation is considered to be a key event in the development of acute respiratory distress syndrome in patients with sepsis. Glucose serves as the main source of energy in activated leucocytes. The aim of this study was to assess whether glucose utilisation in the lungs can be used as an indicator of pulmonary leucocyte accumulation in an experimental model of sepsis of intra-abdominal origin. Sepsis was induced in rats by abdominal implantation of a gelatine capsule containing bacteria and rat colonic contents. Empty gelatine capsules were implanted in control animals. Animals were studied 6 and 12 h after sepsis induction. Glucose utilisation was measured as the tissue uptake of fluorine-18-fluorodeoxyglucose ((18)FDG) 1 h after intravenous injection of the tracer. Micro-autoradiography was also performed after injection of tritiated deoxyglucose. We found increased uptake of (18)FDG in the lungs of septic animals. The uptake also increased with time after sepsis induction. (18)FDG uptake in circulating leucocytes was increased in septic animals compared with controls, and micro-autoradiography showed intense accumulation of deoxyglucose in leucocytes in the lungs of septic animals. We conclude that glucose utilisation is increased in the lungs of septic rats. Measurements of pulmonary glucose utilisation as an index of leucocyte metabolic activity may open new possibilities for studies of the pathophysiology of sepsis and for evaluation of therapeutic interventions. PMID- 10541835 TI - Dual time point fluorine-18 fluorodeoxyglucose positron emission tomography: a potential method to differentiate malignancy from inflammation and normal tissue in the head and neck. AB - Fluorine-18 fluorodeoxyglucose (FDG) positron emission tomography (PET) studies imaging FDG PET imaging is used to detect and stage head and neck cancers. However, the variable physiologic uptake of FDG in different normal structures as well as at inflammatory sites may either obscure a tumor focus or be falsely interpreted to represent tumor activity. Twenty-one patients (9 men, 12 women, median age 59) were scanned serially at two time points, one at 70 min (range 47 112) and the second at 98 min (77-142) after the intravenous injection of 4.3 MBq/kg of FDG. The mean interval between emission scans was 28 min (13-49). Transmission scans were performed and regions of interest (ROIs) were overlayed on the fully corrected images. Standardiued uptake values (SUVs) were generated for the cerebellum, tongue, larynx, every lesion, and a matched contralateral site. Follow-up and pathologic studies revealed 18 squamous cell carcinomas and nine inflammatory or infectious lesions. Tumor SUVs were 4.0+/-1.6 (mean +/- SD) for the first scan and 4. 5+/-2.2 for the second scan. Contralateral SUVs were 1.2+/-0.5 and 1. 1+/-0.5 for the two scans. Tumor SUVs increased by 12%+/-12% as compared with a 5%+/-17% decrease for contralateral sites (P<0.05). SUVs for inflammatory sites (2.0+/-0.7 and 2.0+/-0.9), cerebellum (4. 2+/-1.3 and 4.3+/ 1.4), tongue (1.8+/-0.4 and 1.9+/-0.5) and larynx (1.5+/-0.6 and 1.5+/-0.6) remained constant over time (+0.6%, +2.8%, +1.4%, and -2.4%; P<0.05 when compared with tumor SUV changes). The ratio tumor/contralateral SUV increased by 23%+/-29% over time while this ratio for inflamed sites increased by only 5%+/-15% (P=0.07). The time interval between scans correlated with increase in SUV for tumors (r=0.55, P<0.05) but not for any of the other ROIs. Separation was superior when studies were performed more than 30 min apart (P<0.05). These preliminary data suggest that dual time point imaging compatible with a clinical study protocol is helpful in differentiating malignant lesions from inflammation and normal tissues, especially when separated by a sufficient time interval. PMID- 10541836 TI - Reduction of contamination risks during clinical studies with technegas. AB - During patient studies with the Technegas equipment in our department, we regularly detected small to considerable contaminations of the operator and in the area surrounding the apparatus. These contaminations were found to be of different origin: residual activity in the tubing from the apparatus to the patient which diffuses after deconnection, residual activity and small particles of the destroyed carbon crucible in the apparatus which are dispersed during opening of the door of the gas preparation chamber, leakage at the patient site during studies in uncooperative patients and a dysfunctioning valve inside the apparatus. To reduce the contamination risks, we made some adaptations to the apparatus. In the first place, the dysfunctioning valve was replaced. In addition, a powerful air exhaust pump with an efficient filter was installed. It was connected with (1) a newly installed transparent box in front of the door of the gas preparation chamber, (2) a dome on a flexible arm which can be positioned above the patient's face during the examination and (3) a nipple on which the mouthpiece can be placed after the study. After these adaptations, a study showed the absence of measurable contamination on the clothing of the personnel handling the apparatus. Occasionally, considerable contamination was still measured on the gloves worn during filling of the carbon boat with generator eluate, but only small contaminations (up to 9.25 kBq) were measured on the mouthmask worn by the operator during administration of the Technegas. This results in a maximum effective dose equivalent from activity deposited in the lungs of 0.008 microSv per study. The total body dose of the operator from external radiation for one Technegas examination was determined to be 2 microSv. The highest dose rate was measured during filling of the crucible (0.2 mSv/h). PMID- 10541837 TI - Differential uptake of FDG and DG during post-ischaemic reperfusion in the isolated, perfused rat heart. AB - Fluorine-18 2-fluoro-2-deoxyglucose (FDG) and 2-deoxyglucose (DG) are widely used as tracers of glucose uptake in the myocardium. Although there is agreement that the two analogues behave similarly to glucose under control conditions, there is growing evidence that some interventions (e.g. insulin stimulation or ischaemia/reperfusion) cause differential changes in their behaviour. The addition of a two-surface coil nuclear magnetic resonance (NMR) probe and a dual perfusion cannula to our recently developed PET and NMR dual-acquisition (PANDA) system allows us to collect PET (FDG) images and phosphorus-31 NMR (2 deoxyglucose-6-phosphate) spectra simultaneously from each independently perfused coronary bed of the heart. We have used this technique to study the effect of regional ischaemia/reperfusion on FDG and DG uptake in the isolated, perfused rat heart. During control perfusion, FDG uptake was almost identical in both coronary beds. When one coronary bed was made ischaemic, FDG uptake ceased on that side but continued on the control side. Reperfusion failed to restore FDG uptake. In contrast, NMR spectra showed that, during reperfusion, the uptake and phosphorylation of DG did not differ between the two coronary beds. The results thus demonstrate that regional myocardial ischaemia/reperfusion has different effects on the uptake of FDG and DG in the isolated, perfused rat heart. PMID- 10541838 TI - Single-photon emission tomography imaging of serotonin transporters in the nonhuman primate brain with [(123)I]ODAM. AB - We have described previously a selective serotonin transporter (SERT) radioligand, [(123)I]IDAM. We now report a similarly potent, but more stable IDAM derivative, 5-iodo-2-[2-[(dimethylamino)methyl]phenoxy]benzyl alcohol ([(123)I]ODAM). The imaging characteristics of this radioligand were studied and compared against [(123)I]IDAM. Dynamic sequences of single-photon emission tomography (SPET) scans were obtained on three female baboons after injection of 375 MBq of [(123)I]ODAM. Displacing doses (1 mg/kg) of the selective SERT ligand (+)McN5652 were administered 120 min after injection of [(123)I]ODAM. Total integrated brain uptake of [(123)I]ODAM was about 30% higher than [(123)I]IDAM. After 60-120 min, the regional distribution of tracer within the brain reflected the characteristic distribution of SERT. Peak specific binding in the midbrain occurred 120 min after injection, with an equilibrium midbrain to cerebellar ratio of 1. 50+/-0.08, which was slightly lower than the value for [(123)I]IDAM (1.80+/- 0.13). Both the binding kinetics and the metabolism of [(123)I]ODAM were slower than those of [(123)I]IDAM. Following injection of a competing SERT ligand, (+)McN5652, the tracer exhibited washout from areas with high concentrations of SERT, with a dissociation kinetic rate constant k(off)=0.0085+/ 0.0028 min(-1) in the midbrain. Similar studies using nisoxetine and methylphenidate showed no displacement, consistent with its low binding affinity to norepinephrine and dopamine transporters, respectively. These results suggest that [(123)I]ODAM is suitable for selective SPET imaging of SERT in the primate brain, with higher uptake and slower kinetics and metabolism than [(123)I]IDAM, but also a slightly lower selectivity for SERT. PMID- 10541839 TI - Normal variants, artefacts and interpretative pitfalls in PET imaging with 18 fluoro-2-deoxyglucose and carbon-11 methionine. AB - Interpretation of studies from all imaging modalities requires a knowledge of the possible pitfalls that may occur due to normal variation, artefacts and processes which may mimic pathology. The applications and use of not only 18-fluoro-2 deoxyglucose but also l-[methyl-(11)C] methionine positron emission tomography (PET) are widening and it is timely that the currently recognised interpretative pitfalls are reviewed as the number of dedicated PET scanners and coincidence gamma cameras increases. PMID- 10541841 TI - Analysis of methods for reporting severe and mortal lipoplasty complications. AB - During the past 2 years, media attention has focused on catastrophic outcomes associated with liposuction. A critical review of the lipoplasty literature was undertaken to determine the incidence of severe and mortal complications. Reported lipoplasty complications and patient outcome studies published in the English literature through January 1, 1999, were reviewed. From these and from ASPRS questionnaire surveys of experienced, board-certified plastic surgeons, it is apparent that (1) plastic surgeons do not often voluntarily report severe and mortal complications (either as case reports or in self-reported series), and (2) while survey studies provide the most accurate estimate of complications due to lipoplasty, they are subject to an underreporting bias because they exclude complications occurring in the hands of residents and junior attendings. The mortality from lipoplasty procedures is higher than the 0.003 to 0.02% reported in the literature and may be as high as 0.1%. PMID- 10541842 TI - Ultrasound-assisted lipoplasty: is It really safe? AB - During the last decade liposuction has become the most common aesthetic procedure. It has also become the most common aesthetic procedure performed by physicians not trained in plastic surgery. New developments such as the tumescent technique, finer cannulas, and, finally, the technique of ultrasound-assisted lipoplasty (UAL) have been paralleled by reports of larger amounts of extracted fat. At the same time we see an increasing number of fatal complications. I have undertaken an investigation of 28 patients during 16 months, undergoing liposuction with UAL, to find out how the procedure affected them in terms of blood loss. I found that, while undergoing UAL, our patients lost up to 53% of their blood volume on postoperative day 1, and the average patient still had a loss of 20% of the blood volume 1 week postoperatively. PMID- 10541843 TI - Endermologie: humoral repercussions and estrogen interaction. AB - Endermologie is a motorized rhythmic folding-unfolding and suction technique of the panniculus adiposus. Our study shows that one 40-min Endermologie session produces no noticeable changes in biological parameters, except for plasma estradiol levels, which vary significantly, first by decreasing during the session, then by increasing afterward. Such an Endermologie/estrogen interaction can be compared to the clinical effects observed in some patients undergoing regular Endermologie treatment: return of menses in amenorrheal patients and a trophic effect on skin and subcutaneous connective tissue comparable to that observed during postmenopausal hormone replacement therapy. Understanding such an interaction with the estrogen metabolism requires additional studies and opens many paths for research on therapeutic applications before and after menopause beyond refinement of the body contour and improvement of the appearance of the panniculus adiposus. PMID- 10541844 TI - Creation of an acellular dermal matrix from frozen skin. AB - At present, one of the treatments of choice for closure of full-thickness skin loss is to use a cultured epidermal autograft when skin loss is extensive. In this study, we investigated a simple method of processing frozen surplus skin to produce an acellular, structurally intact, dermal matrix. First, the acellular dermal matrix prepared from normal human skin (ADM) we processed was observed using a transmission electron microscope and a scanning electron microscope. The matrix maintained the basement membrane complex and the extracellular matrix structure of the dermis despite frozen skin being used. Next, using an animal model, we transplanted the ADM and Pelnac, which is used as a contrast in full thickness wounds onto nude rats. The dermal matrix supported fibroblast infiltration and neovascularization. These results suggest that skin processed by our simple method has the potential to be used as a dermal template together with the cultured epidermis in the closure of full-thickness wounds. PMID- 10541845 TI - Breast augmentation by an umbilical approach. AB - Augmentation mammaplasty through an umbilical approach is a technique that, since it was first described by Johnson and Christ in 1993, has gathered both supporters and detractors. The possibility of introducing the prosthesis through the umbilicus without visible scars on the breast area has attracted a considerable number of surgeons and patients. The purpose of this paper is to share our experience with 145 patients operated on between 1994 and 1998 in which this technique was used to insert inflatable saline implants. The indications, aesthetic outcome, and complications of this unique technique are discussed. Furthermore, we have modified the technique by utilizing a dismountable prosthesis introducing clamp (PIC) that was designed by us. PMID- 10541846 TI - Periareolar reduction mammaplasty utilizing the inferior dermal pedicle. AB - The critical points which should not be overlooked when performing reduction mammaplasty are to minimize scar on the breast and to ensure a sufficient blood supply for the viability of the nipple-areolar complex. Periareolar reduction mammaplasty has been widely used because it left only one scar around the areola. However, with the typical periareolar reduction mammaplasty technique, it is difficult to remove a large amount of breast tissue and mobilize the remaining breast tissue. It may result in necrosis of the nipple-areolar complex in some cases. To overcome these limitations we combined the periareolar incision with the inferior dermal pedicle, which has a relatively good blood supply. This new technique was employed in 22 consecutive women (44 breasts) with hypertrophy and a varying degree of ptosis. Infiltration of a tumescent solution and liposuction were performed in all cases. After periareolar incision, dissection of the skin was performed, and the breast was elevated from the fascia of the pectoralis major muscle, leaving the inferior dermal pyramidal pedicle. An adequate amount of tissue was resected in the superior, medial, and lateral areas. After mastopexy, closure was done with a purse-string suture. The amount of tissue resected ranged from 180 to 1510 g per breast, and the mean was 466.1 g. The mean length of elevation of the nipple was 10.6 cm along the meridian of the breast. There were a few complications which needed revision operation: hematoma collection in one breast (2.3%), wound dehiscence in one breast (2.3%), and fat necrosis in one breast (2.3%). There was no necrosis of the nipple-areolar complex. With this new technique of periareolar reduction mammaplasty utilizing the inferior dermal pedicle, we were able to minimize the scar, preserve the nipple-areolar complex, and improve the motility of the breast tissue. But we also observed a flat or square appearance in the case of a large amount of resection in the patients with poor skin elasticity. This technique is safe and versatile and produces aesthetically acceptable results in selected patients. PMID- 10541847 TI - Changes in ocular globe-to-orbital rim position with age: implications for aesthetic blepharoplasty of the lower eyelids. AB - Changes in the relationship of the anterior globe to the orbital rim, orbital fat, and cheek mass are examined in the present study. Two groups of individuals (N = 28) were studied, young versus old, using three-dimensional computer tomography. A computer-derived soft tissue reformat of the data allowed the anterior-posterior changes to be evaluated at the midpupillary plane. Analysis of the data brings to light two important changes which occur with aging. First, the orbital rim moves posteriorly relative to the anterior cornea with age (p = 0.0007). This is important because overresection of orbital fat during lower blepharoplasty accentuates the proptotic appearance of the eye which occurs naturally with age due to orbital remodeling. A second finding is that there is a tendency for the cheek mass to move posteriorly with age relative to the anterior cornea (p = 0.0038). The negative vector, a warning sign for lower blepharoplasty, becomes more common with advancing age. It is suggested that the presence of a negative vector is a sign of generalized maxillary hypoplasia. Certain individuals with a negative vector can be further identified preoperatively by the clinical triad of scleral show, prominent medial fat, and a prominent nasojugal crease. These individuals likewise exhibit maxillary hypoplasia and may be more prone to complications after blepharoplasty. Lastly, a summated model of skeletal remodeling is presented. The significant points are as follows: (1) contrary to previous work, the craniofacial skeleton remodels throughout adulthood, (2) changes in the skeletal architecture impart their effects on the overlying soft tissues, and (3) facial aging is a summation of both hard and soft tissue changes which occur throughout life. PMID- 10541848 TI - Laser double-eyelid operation. AB - The author applied the laser technique in double-eyelid operations. A Sharplan CO(2) laser was used as a cutting tool and also as a hemostatic tool in the defocused mode during all procedures. Eighty cases of laser double-eyelid operations were compared with the conventional method. The results were analyzed during the operation, immediately postoperation, and 3, 7, and 10 days and 1 and 3 months after the operation. The laser double-eyelid technique showed advantages such as shortening of the operation time and minimal bleeding. The operation was safe and efficient, and the healing process was as fast as with the conventional scalpel method. Complications occurred in several cases, including a case of inadvertent tearing of the conjunctival mucosa, a case of hematoma, and two cases of minor wound disruptions immediately after stitch removal. PMID- 10541849 TI - Sex differences in the zygomatic angle in japanese patients analyzed by MRI with reference to Moire fringe patterns. AB - To quantitate the degree of zygomatic protrusion, a new item, "zygomatic angle," was measured on MRI cross sections in the orbitomeatal plane. The sex differences in the zygomatic angle were highly significant on both sides, being larger in males than in females. The shape of the zygomatic protrusion was classified into angulate, intermediate, and massive types according to the value of the zygomatic angle. The incidence of the angulated type was significantly more frequent in females, and that of the massive type was significantly more frequent in males. Another analysis was made of horizontal cross sections in the orbitomeatal plane in moire photographs. In these cross sections, it was possible to judge the above mentioned three types of protrusions. The number of Moire fringes in the infraorbital region was measured in relation to the amount of zygomatic protrusion. The results on Moire photographs coincided with those obtained by MRI analysis. PMID- 10541850 TI - True intraoral reduction malarplasty with a minimally invasive technique. AB - In the Orient, faces are usually wide and short. This physical characteristic is often undesirable in many Asian cultures. Consequently, reduction malarplasty is one of the most common aesthetic procedures performed in the Orient. Previously described techniques for malar reduction such as intraoral chiseling or the burring-down of the zygomatic body and arch often result in minimal invasiveness, with no external scarring. However, they are less effective techniques. Other techniques, such as osteotomy and repositioning of a prominent malar complex by means of a coronal approach also proved fallible, by often resulting in a prominent, visible scar. Furthermore, a combined approach, using both the intraoral and the external (preauricular or sideburn) routes, often results not only in an external scar, but also in possible facial nerve (frontotemporal branch) damage. Thus, we performed a true intraoral osteotomy and reduction malarplasty through upper buccal sulcus incisions, resulting in minimal tissue damage without external scarring. Aesthetic facial contouring surgery has been performed in increasing numbers over the past decade, especially for malar and mandibular angle protrusion. The prominence of the malar area varies according to differences in the inherited bony structure. Essentially, Orientals are mesocephalic, whereas Caucasians are dolichocephalic; the face is wider and shorter in the former than in the latter. An even greater prominence of the zygoma among Orientals, which causes the face to be wider and shorter, is widely regarded as an unattractive feature in the Oriental culture [1,2]. In contrast, a flat cheek bone in Caucasians makes the face appear masculine, in addition to making the nasal and chin prominences more noticeable. Of the previously described techniques for malar reduction, shaving and contouring by the intraoral approach, without the external approach [1,4] is often less effective due to the limitations of shaving. Likewise, osteotomy and repositioning of a prominent malar complex by the coronal approach can result in an extensive visible scar [2,5,6,12]. Additionally, the combined approach by means of the intraoral and external (preauricular or temporal, sideburn) routes [4,7] can result in an external scar and the distinct possibility of facial nerve damage [8]. Therefore, our team created a simple and effective way of conducting a true intraoral osteotomy and reduction malarplasty without external scarring and the instigation of facial nerve damage. PMID- 10541851 TI - Combined frontal bone reshaping and forehead lift for frontal sinus hypertrophy. AB - A technique to correct frontal sinus hypertrophy is presented. The outer table of the frontal sinus is removed and divided into three pieces with an oscillation saw. The pieces are trimmed at the margin, fixed with two microplates, and regrafted. Bilateral lateral thick portions of the supraorbital rim are shaved with a surgical burr. At the same time, a forehead lift is performed. A satisfactory result with a flat and wide forehead can be obtained by employing this technique. PMID- 10541852 TI - Umbilical restoration in abdominal dermolipectomy: A simple double-Y, double-M technique. AB - The authors present a simple technique for restoration of the umbilicus in abdominal dermolipectomy. This procedure is represented by a double-Y cutaneous incision on the abdominal skin and by a double-M cutaneous incision on the umbilical skin. This technique restores a neoumbilicus with multiple small skin flaps and conceals the periumbilical scar at the bottom of the umbilicus. Different designs and figures are proposed to explain the procedure. PMID- 10541853 TI - Treatment of depressed scars with a dissecting cannula and an autologous fat graft. AB - Contemporary options for the improvement of depressed scars include scar revision with an elliptical excision, z-plasty, w-plasty, and geometric broken-line closure. Dermabrasion and laser treatment has been used to obtain a uniform skin surface. When scars are hypertrophic, intralesional steroids and silicone pressure therapy may be useful. Occasionally, scars may be adherent to the underlying fascia. The resulting depression along the length of the scar worsens the aesthetic deformity. Fat injection is an established method for treating depressions and contour deformities. We report encouraging results with the use of this fat injection technique into a pocket made with a sharp cannula in treating 30 patients with postsurgical scars that were depressed and adherent to the underlying fascia. This technique is a useful addition to the surgeon's resources when treating scars. PMID- 10541854 TI - A new surgical technique for the correction of the inverted nipple. AB - Many techniques have been proposed over the years to correct the inverted nipple, a condition which causes both aesthetic and functional problems. In more severe cases, other than causing infections and inflammations, breastfeeding is impossible because of the lack of nipple erection. Reconstructive surgical techniques today are oriented toward methods that allow adequate filling to maintain the nipple permanently everted. In the technique we propose, the nipple is pulled out and extruded by way of a periareolar incision after sectioning the galactophorous ducts and fibrous tissue. To guarantee a permanent eversion, a single trilobed dermoglandular flap is created, overturned, and fixed to fill the "dead space" below the nipple after the lobes have been sutured together. Finally, two transfixed U-shaped sutures are employed to keep the flap in place. From an analysis of the various techniques and results obtained, this method appears to be effective above all in resolving the aesthetic problem in a stable manner and is simpler than the techniques that employ multiple flaps. PMID- 10541855 TI - Mechanisms of mitochondrial DNA escape to the nucleus in the yeast Saccharomyces cerevisiae. AB - The transfer of organelle nucleic acid to the nucleus has been observed in both plants and animals. Using a unique assay to monitor mitochondrial DNA escape to the nucleus in the yeast Saccharomyces cerevisiae, we previously showed that mutations in several nuclear genes, collectively called yme mutants, cause a high rate of mitochondrial DNA escape to the nucleus. Here we demonstrate that mtDNA escape occurs via an intracellular mechanism that is dependent on the composition of the growth medium and the genetic state of the mitochondrial genome, and is independent of an RNA intermediate. Isolation of several unique second-site suppressors of the high rate of mitochondrial DNA-escape phenotype of yme mutants suggests that there are multiple independent pathways by which this nucleic acid transfer occurs. We also demonstrate that the presence of centromeric plasmids in the nucleus can reduce the perceived rate of DNA escape from the mitochondria. We propose that mitochondrial DNA-escape events are manifested as unstable nuclear plasmids that can interact with centromeric plasmids resulting in a decrease in the number of observed events. PMID- 10541856 TI - A 'natural' mutation in Saccharomyces cerevisiae strains derived from S288c affects the complex regulatory gene HAP1 (CYP1). AB - The HAP1 gene encodes a complex transcriptional regulator of many genes involved in electron-transfer reactions and is essential in anaerobic or heme-depleted conditions. We show here that strains derived from S288c carry a defective Ty1 element inserted in the 3' region of the HAP1 ORF. This mutant allele acts as a HAP1 null allele in terms of cytochrome c expression and CYC1 UAS1-dependent transcription, but is able to sustain limited growth in heme-depleted conditions. PMID- 10541857 TI - Mutational analysis of yeast mitochondrial translational activator Cbs2p and of YHR063Cp, a protein with similarity to Cbs2p. AB - Translation of mitochondrial cytochrome b in Saccharomyces cerevisiae requires the nuclearly encoded proteins Cbs1p, Cbs2p and Cbp6p. So far no homologs have been identified, except for the product of the S. cerevisiae orf YHR063C, which has some similarity to Cbs2p. Here we analyze the effect of a null mutation of YHR063C and show that it is not required for mitochondrial respiration. In addition, we report on the importance of the carboxyl-terminus of Cbs2p for its function. We show that truncations and some directed mutations in the carboxyl terminal region of Cbs2p render the protein non-functional. The importance of the COOH-terminus is further underscored by the finding that mutational alteration of the cbs2-1 allele results in the substitution of Ile(372) by Lys. PMID- 10541858 TI - Rpc19 and Rpc40, two alpha-like subunits shared by nuclear RNA polymerases I and III, are interchangeable between the fission and budding yeasts. AB - The cDNAs and genes encoding the common subunits Rpc19 and Rpc40 of nuclear RNA polymerases I and III of Schizosaccharomyces pombe were isolated from cDNA and genomic libraries of the fission yeast and tested for their ability to substitute for the homologous genes in Saccharomyces cerevisiae by heterospecific complementation of corresponding null alleles and temperature-sensitive mutations. The results obtained indicate that both Sz. pombe genes (rpc19(+) and rpc40(+)) are able to replace their S. cerevisiae counterparts in vivo. The primary structure and general organization of both genes were established: rpc40(+) is an intronless gene, while rpc19(+) contains three introns (73, 48 and 77 bp long); rpc19(+) is situated on the long arm of chromosome I and rpc40(+) on the long arm of chromosome II. PMID- 10541859 TI - Identification of an ARS element and development of a high efficiency transformation system for Pichia guilliermondii. AB - An Autonomously Replicating Sequence element adjacent to the RIB1 gene encoding GTP cyclohydrolase II of the yeast Pichia guilliermondii was identified by transformation experiments. Detailed sequence analysis unveiled two potential ARS elements located 5' and 3' of the RIB1 open reading frame. The chromosomal fragment containing the ARS-like sequence 3' to the RIB1 structural gene, called PgARS, conferred high transformation frequencies of 10(4)-10(5) transformants/microg of DNA to a pUC19-derived plasmid in P. guilliermondii. The PgARS element also conferred autonomous replication to hybrid plasmids in this host. Based on this element a series of Escherichia coli shuttle vectors for efficient transformation of the flavinogenic yeast P. guilliermondii was developed. PMID- 10541860 TI - Phylogenetic relationships between mating-type sequences from homothallic and heterothallic ascomycetes. AB - To gain a deeper insight into the evolution of mating-type genes from filamentous ascomycetes, a comprehensive sequence analysis of PCR-amplified sequences corresponding to A- and a-specific mating-type sequences was undertaken. The study included nine homothallic (compatible) and eight heterothallic (incompatible) members of the genera Neurospora and Sordaria. Distance and parsimony trees based on gene fragments from the mat a-1 and mat A-1 genes were compared with trees derived from partial DNA sequences of the gpd glyceraldehyde 3-phosphate dehydrogenase gene. In contrast to the sequences from the gpd gene, mating-type genes show striking sequence differences, suggesting that these genes evolve very rapidly. Strong inter-relationships were found among homothallic, as well as among heterothallic, members of both genera, indicating that in each genus a change from one reproductive strategy to another might result from one single event. PMID- 10541863 TI - ? PMID- 10541861 TI - A pair of invertedly repeated genes in Chlamydomonas reinhardtii encodes a zygote specific protein whose expression is UV-sensitive. AB - Uniparental inheritance of the chloroplast genome has been observed in a wide variety of green plants. In Chlamydomonas this phenomenon, which can be selectively inhibited by UV irradiation of mt(+) gametes, has been shown cytologically to be due to the preferential degradation of mt(-)-derived chloroplast nucleoids in young zygotes. The zygote-specific pair of zys1 genes, zys1A and zys1B, is expressed earliest among five genes isolated from a "10-min" zygote library. We report here that the ZYS1 protein, which is encoded by the invertedly duplicated zys1 gene, accumulates in zygotes and is localized in nuclei. In addition, when mt(+) gametes (but not mt(-) gametes) are UV-irradiated before mating, only very limited accumulation of ZYS1 protein can be detected in the resulting zygotes. PMID- 10541862 TI - A color-selectable and site-specific integrative transformation system for gene expression studies in the dematiaceous fungus Wangiella (Exophiala) dermatitidis. AB - To explore potential virulence factors in the dematiaceous (melanized) fungus Wangiella dermatitidis, we established a gene expression system with properties of homologous transformation and color identification. Using a polyketide synthase gene (WdPKS1) fragment for targeting, we found that 52% of transformants became albinos easily distinguishable from nonspecific transformants. Southern analysis confirmed that the integrations were at the WdPKS1 locus, which however did not affect transformant growth. With a heterologous promoter, P-glaA, enhanced expression of lacZ was found at 37 degrees C. Our results indicated that this system allows the efficient production of isogenic strains for gene function analysis in W. dermatitidis. PMID- 10541864 TI - Validation of a western immunoblotting procedure for bovine PrP(Sc) detection and its use as a rapid surveillance method for the diagnosis of bovine spongiform encephalopathy (BSE). AB - In this report we document the results of several independent studies testing the sensitivity, specificity and reliability of the Prionics Western blotting (PWB) procedure to detect bovine and ovine disease-specific, protease-resistant prion protein (PrP(Sc)). Validation of the technique was obtained by blind analysis of samples from cattle affected with bovine spongiform encephalopathy (BSE), clinically normal animals or cattle with neurological diseases unrelated to BSE. Overall, very high sensitivity, specificity and reliability was observed. It became clear that sampling of the correct brain region and the method used for protein extraction are important factors for correct diagnosis. Furthermore, we tested the usefulness of the PWB technique as an instrument for surveillance purposes. We analyzed animals from a culling scheme as well as older animals from abattoirs to determine the number of subclinical BSE cases detectable by histopathological examination, immunohistochemistry for PrP(Sc) and PWB. In both studies, BSE-affected animals with no overt clinical symptoms were detected. These results demonstrate the usefulness of the PWB procedure in surveillance systems serving as a rapid diagnostic tool to identify animals subclinically infected with BSE. PMID- 10541865 TI - Functional evidence for a role of combined CDKN2A (p16-p14(ARF))/CDKN2B (p15) gene inactivation in malignant gliomas. AB - Homozygous chromosome 9p deletions in gliomas commonly include the CDKN2A and CDKN2B genes, which code for the structurally highly homologous cdk inhibitors/tumor suppressors p16 and p15, respectively. Alternative splicing of the CDKN2A gene results in the expression of p14(ARF). Interestingly, not only p16 and p15, but also the structurally unrelated p14(ARF) appear to function as negative cell cycle regulators. Concerted inactivation of p16, p15 and p14(ARF) could be demonstrated in seven of nine glioblastoma cell lines. Strong suppression of tumorigenicity after transfection with p16 and p15 alone or in combination was seen in cell lines containing neither endogenous p16 nor p15 but functional pRB. Significantly weaker growth suppression was observed in tumors either retaining expression of both p16 and p15 or p15 only. p14(ARF) proved to be a potent tumor suppressor in the presence of wild-type p53, while mutant p53 substantially reduced growth inhibition by p14(ARF). No differences between p16 and p15 effects could be observed, suggesting a largely overlapping function of p16 and p15. To facilitate further research into p16/p15 effects, three cell lines with conditional, tetracycline-controlled p16 expression were established. Reversible growth suppression mediated by p16 was observed in these models. Combined inactivation of CDKN2A and CDKN2B, i.e., loss of both p16 and p15 as well as p14(ARF), results in disruption of two major growth control pathways involving pRB and p53 in malignant gliomas. Therefore, homozygous co-deletions of CDKN2A and CDKN2B rather than mutations targeting individual transcripts are frequently selected for in these tumors. PMID- 10541867 TI - Disease associated prion protein may deposit in the peripheral nervous system in human transmissible spongiform encephalopathies. AB - There is increasing evidence indicating involvement of the peripheral nervous system (PNS) in the pathogenesis of transmissible spongiform encephalopathies (TSEs). Immunocytochemically detectable deposits of TSE-specific abnormal prion protein (PrP(sc)) are considered as a surrogate marker for infectivity. We used anti-PrP immunocytochemistry to trace PrP(sc) deposition in spinal and enteric ganglia, and peripheral nerve in Creutzfeldt-Jakob disease (CJD), Gerstmann Straussler-Scheinker disease (GSS), and fatal familial insomnia. Discrete PrP(sc) deposits were detectable only in a few posterior root nerve fibers in an adaxonal location in one of nine CJD and the one GSS patients examined. Follicular dendritic cells of the gut and enteric nervous system were not labeled. Thus, PrP(sc) may spread to the PNS in different forms of human prion disease. In contrast to our observations in experimental scrapie (Groschup et al., Acta Neuropathol, this issue), the deposits were scant. Possible explanations for this discrepancy comprise strain difference, or centripetal (experimental scrapie) versus centrifugal (sporadic and genetic human prion diseases) spread of PrP(sc), resulting in different patterns and amounts of PrP(sc) accumulation in the PNS. PMID- 10541866 TI - Deposition of disease-associated prion protein involves the peripheral nervous system in experimental scrapie. AB - There is some evidence that the peripheral nervous system (PNS) is involved in the pathogenesis of transmissible spongiform encephalopathies (TSEs). The TSE specific abnormal prion protein (PrP(sc)) is considered as surrogate marker for infectivity. We traced the deposition of PrP(sc) by immunocytochemistry in sheep and hamsters inoculated intraperitoneally with scrapie. The trigeminal, dorsal root, celiac, thoracic, and nodose ganglia contained ganglion cells and fewer satellite cells with prominent granular PrP(sc) deposition. As a novel deposition pattern, punctate deposits in adaxonal location were seen along nerve fibers of peripheral nerve adjacent to ganglia. Such prominent involvement of the PNS in two different experimental scrapie models emphasizes the need to consider the PNS in natural scrapie and other TSEs including bovine spongiform encephalopathy as potential source of infectivity. PMID- 10541868 TI - Alpha-synuclein immunoreactive Lewy bodies and Lewy neurites in Parkinson's disease are detectable by an advanced silver-staining technique. AB - Immunostaining with anti-alpha-synuclein is used to detect Lewy bodies and Lewy neurites in cases of Parkinson's disease and related disorders. To prove that the result of a modern silver method is equivalent to that achieved with immunoreactions for alpha-synuclein, individual sections were successively processed using both methods. The silver-stained sections showed all of the immunoreactive Lewy bodies, and thin Lewy neurites were detected equally well by both techniques. The present study, therefore, points to the capabilities of a modern silver-staining method which is less time consuming and less expensive than immunocytochemical techniques. PMID- 10541869 TI - Disturbance of Notch-1 and Wnt signalling proteins in neuroglial balloon cells and abnormal large neurons in focal cortical dysplasia in human cortex. AB - Determination of neuroglial cell fate and neural tube development during embryogenesis is influenced by the Notch/Wnt signalling pathway. We have studied the localisation of several proteins in the Wnt pathway in focal cortical dysplasia (FCD). This disorder, thought to be due to abnormalities of neuronal migration and differentiation, contains regions of morphologically normal neocortex interrupted by areas of neuronal laminar disorganisation, heterotopic white matter neurons, abnormal large neurons and balloon cells of uncertain histogenesis. We found that the subcellular distribution of several proteins involved in the Wnt pathway differed in regions of FCD from normal cortex, and that within the areas of FCD, the pattern varied with cellular phenotype. Thus, in balloon cells, elevated cytoplasmic levels of dishevelled (Dvl-1) protein were accompanied by an absence of Notch-1, increased adenomatous poliposis coli (APC), altered cytoplasmic beta-catenin, and reduced nuclear expression of beta-catenin. A contrasting pattern of expression was found in abnormal large neurons, which demonstrated elevated levels of Notch-1, and glycogen synthase kinase-3beta (GSK 3beta), and normal levels of APC. Our results are consistent with the notion that Wnt/Notch signalling influences neuroglial cell fate, and suggest that a perturbation of Wnt/Notch signalling may contribute to the neuropathology of FCD. PMID- 10541870 TI - Apoptosis of oligodendrocytes occurs for long distances away from the primary injury after compression trauma to rat spinal cord. AB - We evaluated by in situ nick end labeling the presence of apoptotic glial cells in the spinal cord of rats which have sustained a moderate and severe compression injury at the level of T8-9, resulting in a severe but reversible paraparesis and irreversible paraplegia, respectively. In a previous investigation we found apoptotic glial cells (oligodendrocytes) in the immediate vicinity of the primary lesion (T7 and T10). The present study was designed to evaluate the extent of such cells in the spinal cord even at long distances away from the primary injury. Rats sustaining a moderate and severe compression injury and surviving 4 and 9 days showed a significant increase in the number of apoptotic glial cells at the T1, T5, T7, T12 and L2 levels. At the T10 level the elevation was significant only after day 9. There was no significant increase in the number of these cells at 4 h and 1 day after moderate and severe compression. In general, the apoptotic cells were most often seen in segments adjacent to the compression. They were randomly located in the ventral, lateral and dorsal tracts but were rarely present in the gray matter of the cord. In conclusion, compression trauma to rat spinal cord induces signs of apoptosis in glial cells, presumably oligodendrocytes of the long tracts. This newly discovered type of secondary injury is widely distributed in the damaged spinal cord and occurs even at long distances remote from the initial compression injury. Apoptotic cell death of oligodendrocytes will induce myelin degeneration and cause additional disturbances of axonal function. This cell damage may be a target for future therapy since it occurs after a delay and chemical compounds are now available by which apoptotic cell death can be modified. PMID- 10541871 TI - Detection and localisation of HIV-1 DNA and RNA in fixed adult AIDS brain by polymerase chain reaction/in situ hybridisation technique. AB - In the brain of patients with AIDS, HIV-1 is localised in a productive form in mononuclear cells. One issue that still needs clarification is whether HIV is localised in cells other than those of mononuclear lineage. Gene amplification by polymerase chain reaction/in situ hybridisation (PCR-IS) could shed light on it. In this study, formalin-fixed, paraffin-embedded brain tissue from ten adult AIDS sufferers was used. Five of them showed evidence of HIV encephalitis (HIVE), five did not show any abnormality. Nested PCR revealed HIV-1 DNA in all HIVE cases and in three of the group without HIVE. HIV-1 DNA and RNA were also detected in situ in seven cases (all seven were also HIV-1 DNA positive in tube). A higher signal was located in the white than in the grey matter. HIV-1 DNA was found in microglia, macrophages, perivascular cells, multinucleated gaint cells (MGC) and in CD68-negative cells. Some of them were identified as endothelial cells, astrocytes and oligodendrocytes. Reverse transcriptase-PCR-IS was positive in macrophages, MGC, endothelial and glial cells. These results confirm infection of endothelial cells and other glial cells and give clues about the route of entry of virus into the central nervous system and the pathogenesis of the disease. This study did not give any convincing evidence supporting an infection of neurons by HIV-1. PMID- 10541872 TI - Presenilin-1 expression in Pick's disease. AB - Recent studies have reported that neuronal populations expressing low levels of presenilin-1 (PS-1) display increased vulnerability in late-onset sporadic Alzheimer's disease (AD). To examine whether this phenomenon also occurs in other neurodegenerative diseases, we performed a quantitative immunocytochemical study of PS-1 distribution in the cerebral cortex of Pick's disease (PiD) cases and non demented individuals. In PiD cases, the percentage of PS-1-containing, Pick body (PB)-free neurons was significantly elevated only in cortical areas showing neuronal loss. In these areas, PS-1 levels, measured by immunoblotting, were often higher in PiD compared to non-demented cases. Moreover, PS-1 immunoreactivity was significantly reduced in PB-containing neurons. These data suggest that as previously shown in AD, low cellular expression of PS-1 may be associated with increased neuronal loss and cellular degeneration. PMID- 10541873 TI - Ultrastructural localization of phosphorylated eIF2alpha [eIF2alpha(P)] in rat dorsal hippocampus during reperfusion. AB - During post-ischemic brain reperfusion there is a substantial reduction of protein synthesis in selectively vulnerable neurons. Normal protein synthesis requires a functional translation initiation complex, a key element of which is eukaryotic initiation factor 2 (eIF2), which in a complex with GTP introduces the met-tRNA(i). Phosphorylation of Ser(51) on the alpha subunit of eIF2 [eIF2alpha(P)] generates a competitive inhibitor of eIF2B, thereby preventing the replenishment of GTP onto eIF2, thus blocking translation initiation. It has been shown that the conditional expression of an eIF2alpha mutant (Asp substituted for Ser(51)) imitating the negative charge of Ser(51) (P) induces apoptosis. During the first 10 min of post-ischemic reperfusion, there is an approximately 20-fold increase in eIF2alpha(P) seen in the cytoplasm of CA1 hippocampal neurons, and, by 1 h, there is also accumulation of eIF2alpha(P) in the nucleus. We utilized post-embedding electron microscopical immunogold methods to examine the localization of eIF2alpha(P) during reperfusion. Immunogold particles (10 nm) were concentrated chiefly along the rough endoplasmic reticulum and in association with the membranes of the nuclear envelope in CA1 neurons. Aggregations of gold particles in the nucleus were concentrated: (1) within and around the nucleolus, (2) associated to strands of heterochromatin, and (3) along putative nuclear filaments. The presence of eIF2alpha(P) in the nucleolus probably reflects its association with nascent ribosomal subunits. The beta subunit of eIF2 has a zinc finger and polylysine blocks analogous to those on other proteins that affect transcription. The association of eIF2alpha(P) with chromatin may have important implications for transcription. PMID- 10541874 TI - Variant Gerstmann-Straussler syndrome with the P105L prion gene mutation: an unusual case with nigral degeneration and widespread neurofibrillary tangles. AB - We present here a case of variant Gerstmann-Straussler syndrome (GSS) with a codon 105 mutation of the prion protein gene. A 57-year-old woman developed dementia and gait disturbance dissimilar to the spastic paraparesis that is observed in most cases with codon 105 mutation. The clinical course of the disease in this case was 12 years. The brain weighed 900 g, and the frontal lobe, pallidum and thalamus were markedly atrophic. Severe neuronal loss was observed in the deep layer of the frontal and temporal cortices, and fibrillary gliosis and a marked loss of neurons was observed in the globus pallidus, thalamus and substantia nigra. Many amyloid plaques and some ballooned neurons were present in the frontal, temporal and parietal cortices. However, no spongiform changes were seen. The cerebellum was relatively well preserved. Numerous neurofibrillary tangles (NFTs) were recognized in the cerebral cortices, and scattered NFTs were observed in the basal nucleus of Meynert, thalamus, substantia nigra, periaqueductal gray matter, raphe nuclei and locus ceruleus. The case presented here indicates the presence of variations in the pathological findings of cases with codon 105 mutation, and that the formation of cortical and brain stem NFTs might have something to do with the duration of illness and/or the degree of brain tissue destruction that had occurred. PMID- 10541875 TI - Sporadic amyotrophic lateral sclerosis with multiple system degeneration: a report of an autopsy case without respirator administration. AB - This report concerns an autopsy case of amyotrophic lateral sclerosis (ALS) with unusual clinical and neuropathological findings. The patient was a Japanese man without hereditary burden who was 49 years old at the time of death. His clinical manifestation included dysarthria at age 48, followed by dysphagia, atrophy and fasciculation of the tongue, muscle weakness in the four extremities, tremor, rigidity, increased deep tendon reflexes in the upper and lower extremities, and incoordination of the four extremities. He died of respiratory failure 12 months after the disease onset. No respirator administration was performed throughout the clinical course. The neuropathological examination revealed not only degeneration of upper and lower motor neuron systems, including the presence of Bunina bodies and ubiquitin-immunoreactive neuronal inclusions in the lower motor neurons, but also prominent degeneration of the substantia nigra and dentate nucleus with slight neuronal loss in the locus ceruleus and pontine nucleus. To our knowledge, this is the first reported case of sporadic ALS without dementia and respirator support, showing degeneration of the substantia nigra and dentate nucleus. This report may contribute to the resolution of the question concerning the neuropathological heterogeneity of sporadic ALS with respiratory support. PMID- 10541876 TI - Neuropathology at the Turn of the Millenium 44th Annual Meeting paragraph signDeutsche Gesellschaft fur Neuropathologie und Neuroanatomie e.V.Bonn, October 6-9, 1999 paragraph signPresident: Otmar D. Wiestler, Bonn paragraph sign Assistants: Barbel Prushoff and Benedict Wiestler. PMID- 10541877 TI - Tumor markers in peripheral blood of patients with malignant melanoma: multimarker RT-PCR versus a luminoimmunometric assay for S-100. AB - The identification of circulating tumor cells in the peripheral blood of patients with malignant melanoma by detection of melanoma associated protein transcripts using the reverse transcriptase polymerase chain reaction (RT-PCR) technique has been introduced as a noninvasive and sensitive technique for early detection of tumor progression and metastatic disease. An alternative approach is the analysis of S-100 protein in the serum of melanoma patients by a luminoimmunometric assay (LIA). In this study, the sensitivities of RT-PCR and LIA were compared. Seventy seven blood samples of 59 melanoma patients were analyzed for tyrosinase, Melan A/MART-1, MAGE-3, gp100, and p97 expression by multimarker RT-PCR; 540 serum samples of 352 melanoma patients were analyzed for S-100 protein concentration by LIA. In stage III 23.8% and in stage IV 37.5% of the samples were positive for at least one marker in multimarker RT-PCR, versus 8.1% and 48.1% of elevated S-100 levels analyzed by LIA, respectively. In a direct comparison, 31 identical samples were analyzed by multimarker RT-PCR and by S-100 LIA. In stage III 18.2% and in stage IV 45% of the samples were positive by multimarker RT-PCR versus 45.5% and 80% by S-100 LIA, respectively. S-100 LIA was more sensitive in detection of metastatic disease in melanoma patients than multimarker RT-PCR and should be evaluated in further studies. RT-PCR might be more useful in the analysis of micrometastases in anatomic compartments other than peripheral blood. PMID- 10541878 TI - T lymphocytes and altered keratinocytes express interferon-gamma and interleukin 6 in lichen planus. AB - Lichen planus is assumed to represent a delayed hypersensitivity reaction, in the course of which cytokines control the proliferation and differentiation of cytotoxic T lymphocytes which attack the epidermis and cause apoptosis of undifferentiated keratinocytes. Since interferon-gamma and interleukin 6 are known to be markedly generated in lichen planus, we investigated the cellular localization of these cytokines in affected skin/oral mucosa biopsy specimens using in situ hybridization for interferon-gamma and in situ reverse transcription-polymerase chain reaction for interleukin 6 mRNA. In the upper subepithelial connective tissue interferon-gamma mRNA was noted within proliferating CD3+ T lymphocytes. In this tissue compartment interleukin 6 mRNA was detected in infiltrating CD4+ and CD8+ T lymphocytes. In the epithelium, expression of interferon-gamma mRNA and interleukin 6 mRNA was observed in the basal and suprabasal keratinocytes of altered skin/oral mucosa. In contrast, normal skin did not reveal any interferon-gamma or interleukin 6 expression, although a few CD4+ and CD8+ T lymphocytes were noted in the dermis as well as the epidermis. These findings indicate that in lichen planus the proinflammatory cytokines interferon-gamma and interleukin 6 are produced not only by activated T lymphocytes but also by altered keratinocytes, and suggest that stimulated keratinocytes may amplify the course of lichen planus. PMID- 10541880 TI - The action of a novel vitamin D3 analogue, OCT, on immunomodulatory function of keratinocytes and lymphocytes. AB - Topical vitamin D3 has relatively recently been introduced for the treatment of psoriasis. Synthetic vitamin D3 analogues with a high potential for inducing differentiation of cells, but with a low hypercalcemic effect have recently been developed. One such synthetic analogue of 1,25-dihydroxyvitamin D3 (calcitriol), 22-oxacalcitriol (OCT), is a novel agent for the topical treatment of psoriasis. The activity of OCT in vitro was investigated and compared with that of a series of vitamin D3 analogues as to their ability to inhibit murine T lymphocyte proliferation stimulated by con-A, to suppress IL-6 and IL-8 production by keratinocytes stimulated with IL-1alpha and TNFalpha, and to inhibit AP-1- and NFkappaB-dependent reporter gene expression. OCT inhibited the proliferation of lymphocytes and suppressed IL-8 and IL-6 production by keratinocytes to the same extent as the other vitamin D3 analogues. It also inhibited AP-1- and NFkappaB controlled luciferase activity to the same extent as the other vitamin D3 analogues, which demonstrates its mechanism of action in the suppression of inflammatory processes. PMID- 10541879 TI - Measurement of 5-methoxypsoralen and 8-methoxypsoralen in saliva of PUVA patients as a noninvasive, clinically relevant alternative to monitoring in blood. AB - BACKGROUND: Monitoring of psoralen concentration and time-course in PUVA patients is vital for efficient PUVA therapy. Blood sampling is invasive and labour intensive and thus unsuited for routine use and repeat measurements over the course of therapy. OBJECTIVE: Psoralen pharmacokinetics in saliva were investigated and validated as a noninvasive, simple and biologically relevant alternative to measurements in blood. METHODS: The time-course of psoralen concentration was measured in saliva and serum of volunteers and patients receiving PUVA or extracorporeal photopheresis therapy. The samples were analysed by high-performance liquid chromatography. Three commonly used oral psoralen preparations were tested: Psoraderm5 (5-methoxypsoralen; 5-MOP), Meladinine and Oxsoralen (both 8-methoxypsoralen; 8-MOP). RESULTS: The pharmacokinetic parameter Cmax in saliva averaged 10% (range 6-20%) of the serum values for 8-MOP, and < or = 4% for 5-MOP. These concentrations correspond to the therapeutically relevant, non-albumin-bound fraction of psoralen in serum that is available to diffuse into the tissues. The parameter tmax in saliva and serum coincided, indicating that psoralens diffuse rapidly between the two compartments. CONCLUSION: Monitoring of psoralens in saliva is a valuable, noninvasive alternative to measurements in serum, suitable for routine use. A series of five or six saliva samples is sufficient to determine tmax in a patient beginning photochemotherapy. To determine Cmax, three independent saliva measurements at t = tmax are recommended. PMID- 10541881 TI - Expression of vitamin D3 25-hydroxylase (CYP27) mRNA after induction by vitamin D3 or UVB radiation in keratinocytes of human skin equivalents--a preliminary study. AB - The presence of both 1alpha-hydroxylase activity and 25-hydroxylase activity is a biochemical prerequisite for the enzymatic generation of 1alpha,25(OH)2D3 from vitamin D3 in keratinocytes. In contrast to 1alpha-hydroxylase, however, activity of a 25-hydroxylase in keratinocytes has not previously been demonstrated. In this study, using RT-PCR we detected vitamin D3 25-hydroxylase mRNA in human keratinocytes. The expression of vitamin D3 25-hydroxylase mRNA in keratinocytes was identical to that of CYP27 expressed in mitochondria of human hepatoma HepG2 cells. CYP27 mRNA in keratinocytes is not constitutively expressed but is inducible both by vitamin D3 (780 nM) and by UVB radiation (300 nm, 30 mJ/cm2). PMID- 10541882 TI - Determinants of melanocyte density in adult human skin. AB - The distribution of melanocytes in human skin has been observed to vary within and among individuals, yet little is known of the factors that determine the density of these pigment cells. These factors were explored in a molecular epidemiological study conducted among a population-based sample of 97 male subjects aged over 50 years in Queensland, Australia. Information relating to environmental and phenotypic factors was collected through face-to-face interviews and physical examination of all participants. In addition, 2-mm biopsies of representative skin were taken from the dorsum of the hand and another anatomical site. Melanocytes were identified by cytoplasmic staining with the B8G3 (anti-TRP1) monoclonal antibody using standard immunohistochemical techniques. Melanocyte counts were performed blind by two observers. On crude analysis, melanocyte density decreased with advancing age (P = 0.0002), and increased with increasing number of naevi (P = 0.01). Other pigmentary characteristics (such as hair and eye colour and depth of tan) were not associated with epidermal melanocyte density. Melanocyte density varied significantly by anatomical site (P = 0.02), with highest densities observed on the back/shoulders (n = 50, 17.1 +/- 8.8 cells/mm, mean +/- SD) followed by the upper limbs (n = 11, 12.6 +/- 8.8 cells/mm) and lower limbs (n = 14, 14.4 +/- 5.9 cells/mm). Lowest melanocyte densities were recorded on the anterior trunk (n = 3, 3.2 +/- 2.4 cells/mm). These findings confirm the results of earlier studies in which site-specific differences in melanocyte density have been found. We speculate that the unequal distribution of melanocytes may partially explain the site-specific incidence of melanoma, offering fresh perspectives on the aetiology of this cancer. PMID- 10541883 TI - Effect of barrier perturbation on cutaneous penetration of salicylic acid in hairless rats: in vivo pharmacokinetics using microdialysis and non-invasive quantification of barrier function. AB - The penetration of topically applied drugs is altered in diseased or barrier damaged skin. We used microdialysis in the dermis to measure salicylic acid (SA) penetration in hairless rats following application to normal (unmodified) skin (n = 11) or skin with perturbed barrier function from (1) tape-stripping (n = 5), (2) sodium lauryl sulphate (SLS) 2% for 24 h (n = 3) or (3) delipidization by acetone (n = 4). Prior to the experiment, transepidermal water loss (TEWL) and erythema were measured. Two microdialysis probes were inserted into the dermis on the side of the trunk and 5% SA in ethanol was applied in a chamber overlying the probes. Microdialysis sampling was continued for 4 h, followed by measurements of probe depth by ultrasound scanning. SA was detectable in all samples and rapidly increasing up to 130 min. Microdialysates collected between 80 and 200 min showed mean SA concentrations of 3 microg/ml in unmodified and acetone-treated skin, whereas mean SA concentrations were 280 microg/ml in SLS-pretreated skin and 530 microg/ml in tape-stripped skin (P < 0.001). The penetration of SA correlated with barrier perturbation measured by TEWL (P < 0.001) and erythema (P < 0.001). A correlation between dermal probe depth and SA concentration was found in unmodified skin (P = 0.04). Microdialysis sampling in anatomical regions remote from the dosed site excluded the possibility that SA levels measured were due to systemic absorption. Microdialysis sampling of cutaneous penetration was highly reproducible. Impaired barrier function, caused by irritant dermatitis or tape stripping, resulted in an 80- to 170-fold increase in the drug level in the dermis. This dramatic increase in drug penetration could be relevant to humans, in particular to topical treatment of skin diseases and to occupational toxicology. PMID- 10541884 TI - UVA1 and UVB radiation but not PGE2 stimulate IL-8 release in normal human keratinocytes. PMID- 10541885 TI - Production of reactive oxygen intermediates by human macrophages exposed to soot particles and asbestos fibers and increase in NF-kappa B p50/p105 mRNA. AB - Alveolar macrophages (AM) play a decisive role in the immunologic defense system of the lung and in inflammatory pulmonary pathomechanisms. AM and blood monocytes (BM) were exposed to chrysotile B, soot FR 101, and Printex 90 (P 90). We evaluated the reactive oxygen intermediate (ROI) release of AM and BM after particle exposure. ROI release was measured by chemiluminescence. Thirty-minute exposure caused a significant (up to 2.5-fold) increase in ROI release of AM (100 micrograms/10(6) cells) compared with control experiments (p < 0.01). Identical exposure conditions for BM resulted in a similar reaction pattern (maximum 2.2 fold increase in ROI release; p < 0.05). After a 90-min particle exposure at concentrations of 10 and 100 micrograms/10(6) cells, we investigated the steady state level of p50/p105 mRNA encoding for the precursor protein of the p50 subunit of nuclear factor kappa B (NF-kappa B) by semiquantitative reverse transcription-polymerase chain reaction. One hundred micrograms Chrysotile B, FR 101, or P 90 induced a significant maximum 4.0-fold up-regulation of NF-kappa B gene expression in AM and a 3.3-fold up-regulation in BM (p < 0.05). The addition of superoxide dismutase (200 U/ml) to particle- and fiber-exposed macrophages resulted in inhibition of ROI release and a decrease in NF-kappa B mRNA expression (70%). NF-kappa B is an important transcription factor involved in the regulation of numerous genes (e.g., for inflammatory cytokines, and cytokine receptors). These cytokines are supposed to be involved in inflammatory pathomechanisms in bronchial epithelial cells, which result, for example, in chronic obstructive pulmonary disease. Our results suggest that particle-induced ROI release is associated with an increase in NF-kappa B (p50/p105) mRNA steady state level. PMID- 10541886 TI - Variability of airway responses in mice. AB - We examined the effect of animal strain, type of spasmogen, and mode of spasmogen administration on the pattern of lung mechanical responses in intubated and mechanically ventilated mice. We determined the response in inspiratory respiratory system resistance (R(rs)) and inspiratory static respiratory system compliance (C(rs)) to increasing doses of inhaled or intravenous carbachol or serotonin in Balb/C and C57BL/6 mice. R(rs) responsiveness was quantitated by calculating, by interpolation, the inhaled spasmogen concentration (PC(150)) and intravenous spasmogen dose (PD(150)) causing an increase in R(rs) to 150% of baseline. C(rs) responsiveness was calculated similarly for a decrease in C(rs) to 85% of baseline (PC(85) for inhaled and PD(85) for intravenous spasmogen). Baseline R(rs) and C(rs) were similar in all groups. R(rs) responsiveness to inhaled and intravenous carbachol and serotonin tended to plateau and was not different in the two strains. In contrast, C(rs) responses were variable and had a greater mean PC(85) for inhaled serotonin than carbachol in both strains and a greater fall in mean C(rs) at PC(150) for carbachol in Balb/C mice; no interstrain and interdrug differences in PD(85) were noted for intravenous spasmogens. Intravenous atropine attenuated the R(rs) response to high-dose inhaled and intravenous serotonin, suggesting the involvement of a vagal reflex. In contrast, atropine attenuated C(rs) responses only for intravenous serotonin in Balb/C mice. These results suggest that animal strain, spasmogen, and mode of administration determine the extent to which induced airflow resistance is accompanied by increases in elastic recoil. PMID- 10541887 TI - Apoptotic activity is increased in the newly formed fibromyxoid connective tissue in bronchiolitis obliterans organizing pneumonia. AB - Bronchiolitis obliterans organizing pneumonia (BOOP) and usual interstitial pneumonia (UIP) are clinically and histologically distinguishable interstitial lung diseases both sharing the presence of the newly formed fibromyxoid connective tissue as a histologic feature. In BOOP the newly formed connective tissue is susceptible to even complete reversal, but in UIP it participates in the remodeling of the pulmonary tissue. In this study we have tested the hypothesis that the apoptotic activity in the fibromyxoid lesions is higher in BOOP than in UIP. Apoptotic activity in cells of the newly formed connective tissue in BOOP and UIP was visualized by TUNEL. Apoptosis-regulating proteins bcl 2, mcl-1, and bax were visualized by immunohistochemistry. The number of the apoptotic events was given as an apoptotic index giving the percentage of the apoptotic cells and bodies of the total cell number. Our results show that the apoptotic activity is clearly higher in the fibromyxoid lesions of BOOP (mean, 0.7; S.D., +/-0.51) compared with those of UIP (mean, 0. 13; S.D., +/-0.14, p < 0.003). The results thus suggest that apoptosis has an important role in the resolution process of the newly formed connective tissue in BOOP. The low level of apoptosis in UIP could, on the other hand, suggest that a decreased apoptosis in cells of the fibromyxoid stroma might pathogenetically relate to a decreased resolution of these lesions in UIP. No significant difference was found in the expression of bcl-2, mcl-1, and bax in BOOP and in UIP, suggesting that regulation of apoptosis might partly bypass the influence of these proteins. PMID- 10541888 TI - Th1 cells that adoptively transfer experimental hypersensitivity pneumonitis are activated memory cells. AB - Cultured murine CD4+ T cell lines from Saccharopolyspora rectivirgula-sensitized donors with cytokine secretion characteristics of Th1 cells can adoptively transfer murine experimental hypersensitivity pneumonitis (EHP), whereas Th2 CD4+ cell lines cannot (Cell Immunol 177:169-175, 1997). To assess the differences between these cell lines that may be related to the ability to transfer EHP, we determined cell surface markers that distinguish naive from activated/memory cells that indicate activation and that mediate endothelial adhesion. Both Th1 and Th2 T cell lines are CD4+, CD11a+, ICAM-1+, and L-selectin negative. Th1 cells are CD49d (alpha 4) and LPAM (alpha 4 beta 7) positive, with 32% and 42% of the apparent membrane site density quantitated as the mean molecules of equivalent soluble fluorochromes (MESF) values of unstimulated spleen cells, respectively. Th2 cells are weakly alpha 4 and alpha 4 beta 7 positive, with 15% and 11% of the MESF of unstimulated spleen cells. Th1 cell lines are CD45Rb negative and CD44+, whereas Th2 cell lines are CD45Rb intermediate and CD44-/low. Th1 cells are CD25 (IL-2 receptor) low and Th2 cells CD25 high. We conclude that Th1 cells capable of transfer are activated/memory T cells, and Th2 cells incapable of transfer lack some characteristics of memory/activated T cells (i.e., increase of CD44 and decrease of CD45Rb). Both Th1 and Th2 cell lines express alpha 4 beta 7 and alpha 4 (Th1 > Th2), suggesting that alpha(4) integrin may be important in conferring ability to cells to adoptively transfer EHP. PMID- 10541896 TI - ICG-guided laser photocoagulation of juxtafoveal and extrafoveal occult choroidal neovascularization. AB - BACKGROUND: The most common cause of legal blindness in the Western world in people over 50 years of age is age-related macular degeneration (AMD) complicated by choroidal neovascular membranes (CNV). We conducted a prospective pilot study to evaluate the functional and anatomical results of indocyanine green (ICG) guided laser photocoagulation of juxta- and extrafoveal occult CNV. METHODS: Seventeen consecutive patients (17 eyes) with occult CNV in AMD were included. All showed occult CNV appearing as late-phase fluorescein leakage of undetermined source on fluorescein angiography and as a well-defined hot spot or plaque hyperfluorescence on ICG angiography. Fibrovascular retinal pigment epithelial detachments (PED) and serohemorrhagic PED were excluded. Laser photocoagulation was performed using an argon green laser. The patients had serial follow-up examinations, including fluorescein and ICG angiography, at 15 days and 1, 2, 3, 6, 9 and 12 months after photocoagulation. RESULTS: At the end of the follow-up, the visual acuity (VA) was stable (within +/-3 lines of the initial VA) in 76% (13) of the eyes. A moderate decrease in VA was observed in 24% (4). A complete resolution of exudative signs was observed in 65% (11). CONCLUSION: ICG-guided laser photocoagulation of occult CNV, presenting at baseline examination as late phase fluorescein leakage of undetermined source and as a well-defined hyperfluorescent lesion on ICG angiography, may constitute a subgroup of occult CNV that benefits from ICG-guided laser photocoagulation. A multicentric randomized controlled clinical study is, however, mandatory to confirm this result. PMID- 10541889 TI - Previously unrecognized obstructive sleep apnea in Chinese subjects with essential hypertension. AB - Obstructive sleep apnea (OSA) is found to affect 2-4% of the middle-aged population in several Caucasian studies, whereas the prevalence among other ethnic groups have not been clearly documented. It has been reported that OSA and systemic hypertension are highly associated; we therefore conducted a study on Chinese subjects who were receiving treatment for essential hypertension to assess the prevalence of OSA among this group. Ninety-two consecutive patients being followed up at a hypertension clinic were recruited for a questionnaire survey. The entire study group was aged 54.7 +/- 11.7 years, with 40 men. One male subject had a diagnosis of obstructive sleep apnea on nasal continuous positive airway pressure (nCPAP) treatment and 46 subjects agreed to an overnight sleep study. Those who underwent sleep study showed selection bias with a higher body mass index and more symptoms associated with OSA. Of the 46 who underwent sleep study, 16 (34.8%) had an obstructive apnea-hypopnea (AHI) score of >/=5 and excessive daytime sleepiness, with a median score of 26.2 (range, 8.3-64.9). Patients in the group with obstructive sleep apnea syndrome (OSAS) thus defined compared with those without OSAS had more men (64.7 vs 17.20%, p = 0.001) and an excess of smokers (31.5 vs 3.3%. p = 0.01) and had significantly more symptoms of excessive daytime sleepiness (p = 0.001), daytime fatigue (p = 0.007), and witnessed apneas (p = 0.008). Seven patients accepted treatment with nCPAP and reported improvement in symptoms, but there was no detectable change in clinic blood pressure measurements after 3 months of nCPAP treatment. This study demonstrated a high prevalence of previously unidentified OSAS among Chinese patients with essential hypertension. Increased awareness of both doctors and patients toward this potentially treatable problem is warranted. PMID- 10541897 TI - A prospective randomized comparison of two techniques of combined cataract glaucoma surgery. AB - BACKGROUND: Small-incision cataract surgery combined with trabeculectomy offers new options for surgical treatment of patients with glaucoma and cataract. The purpose of this prospective randomized study was to compare the efficacy and safety of two different techniques of combined surgery: a one-site and a two-site approach. METHODS: Fifty eyes of 50 patients were included in this study. Twenty five patients were randomly assigned to the one-site procedure and 25 patients to the two-site procedure. The one-site approach consisted of a superior tunnel phacoemulsification under a scleral flap with subsequent trabeculectomy. The two site approach included a temporal corneal phacoemulsification combined with a separate-incision superior trabeculectomy. RESULTS: The preoperative mean intraocular pressure (IOP) of 29.8+/-4.9 mmHg dropped significantly to 15.9+/-3.2 mmHg. The mean follow-up time was 19+/-4. 3 months (range 4-25 months). The reduction of IOP was more pronounced in the two-site group (50.1%) than in the one-site group (43%), but the difference was not statistically significant. Patients needed 2.2+/-1.7 antiglaucomatous medications preoperatively vs 0.52+/ 1.39 postoperatively. Three patients (6%) required needling of encapsulated bleb, and two patients underwent a reoperation to control IOP. Mean visual acuity improved from 0. 14+/-0.36 to 0.38+/-0.30 postoperatively. The most common complications after combined surgery were fibrinous exudation (24%) and hyphema (12%). CONCLUSION: Both techniques of combined cataract and glaucoma surgery proved to be efficient and safe procedures to control IOP and to improve visual acuity. The reduction of IOP did not differ between the one-site approach and the two-site approach. PMID- 10541898 TI - Cystoid macular edema following immune recovery and treatment with cidofovir for cytomegalovirus retinitis. AB - BACKGROUND: Cytomegalovirus (CMV) retinitis is the most common opportunistic ocular infection in AIDS patients. Cidofovir has proved to be highly effective in treatment of CMV retinitis. Iritis and bulbar hypotony are known as the major complications after intravenous and intravitreal use of this antiviral drug. Cystoid macular edema (CME) after intravenous application of cidofovir has not been reported. METHODS: We analyzed retrospectively the incidence of CME after intravitreal or intravenous application of cidofovir and its correlation with CD4 cell counts of the patients. RESULTS: Two (22.2%) of 9 eyes in the intravenous and 3 (18.8%) of 16 eyes in the intravitreal injection group developed CME. It occurred between 3 and 48 weeks after cidofovir administration. In all eyes CMV retinitis was inactive. All patients received highly active antiretroviral treatment (HAART). CME was correlated with a rapid and sustained improvement in CD4 cell counts. CONCLUSION: We interpret the occurrence of CME as an immune recovery phenomenon for the following reasons. All CMEs were seen in eyes with inactive CMV retinitis and the unaffected contralateral side never developed CME. The time range of appearance between 3 and 48 weeks after cidofovir administration makes direct toxicity of cidofovir unlikely. All patients had a sustained improvement of CD4 cell counts due to HAART. No CME was reported during the use of cidofovir before the HAART era. PMID- 10541899 TI - Infrared imaging of cystoid macular edema. AB - BACKGROUND: Cystoid macular edema (CME), a cause of central visual loss, is described in various pathologies. Typically, the fluorescein angiogram confirms the diagnosis and provides qualitative information as to the extent of leakage. This study was performed to investigate the features of cysts and quantify the extent of CME using non-invasive infrared imaging. METHODS: Eighteen eyes of 16 successive patients with CME in association with a broad spectrum of diseases were included in the study. The diagnosis of CME was established clinically and confirmed by fluorescein angiography. Digital infrared imaging was performed with a research scanning laser ophthalmoscope with different apertures, providing direct confocal and indirect imaging modes, to discriminate superficial features from deeper ones and to emphasize sources of multiple laterally scattered light. RESULTS: CME was easily detected with infrared imaging in all eyes. Confocal mode visualized the cysts themselves, while indirect mode emphasized borders. Large central cysts were detected as distinct, non-confluent structures. In addition, folds detected with infrared imaging in the macula in 12 of the 18 eyes were not always observed clinically. CONCLUSION: Infrared imaging provides a quick and safe diagnostic tool for patients with CME. The cystoid structures are readily localized and quantified, useful for monitoring CME. Despite differences in the pathophysiology, cysts did not differ qualitatively in a variety of diseases with infrared imaging. PMID- 10541901 TI - Perimetric defects after a single acute angle-closure glaucoma attack. AB - BACKGROUND: This study was carried out to determine the effect of an acute attack of angle-closure glaucoma on the visual field. METHODS: A total of 53 eyes were examined 36-48 h after remission of an acute glaucoma attack by means of computerised perimetry (Humphrey 630 perimeter, 30-2 program). Perimetry was repeated after at least 1 month in 22 eyes. RESULTS: Perimetric defects, varying greatly in severity and primarily of the generalised type or concerning at least wide sectors of the field, were detected in 45 (85%) of 53 cases. The visual field was normal in the remaining 8 patients (15%). The upper nasal quadrants were the most frequently affected and the degree of eccentricity was most frequently involved within the 9 degrees -21 degrees area. In 7 of the 22 cases in which perimetry was repeated after 1 month, complete normalisation was noted in the visual field. CONCLUSIONS: An isolated attack of acute glaucoma produces in most cases a perimetric defect of generalised or mixed type. This may be reversible. The most affected zones were the upper half of the visual field and the 9 degrees -21 degrees area. PMID- 10541900 TI - Iris angiographic changes in multifocal chorioretinitis with panuveitis. AB - BACKGROUND: Multifocal chorioretinitis with panuveitis (MCP) is a chronic inflammatory disease of the peripheral retina and choroid with typical clinical appearance. Although obvious involvement of the anterior segment is often mild, severe chronic inflammatory reactions can occur after cataract surgery. Explantation of an intraocular lens (IOL) or primary aphakia may be necessary. In this pilot study we therefore examined the iris of patients with MCP by means of fluorescein angiography (IAG) to investigate iris vessel involvement. MATERIALS AND METHODS: Twenty-one eyes of 13 patients with MCP (12 women, 1 man) were examined by IAG. The average age of the patients was 72.5+/-6.2 years, and the average duration of the disease prior to examination was 13 months. In 9 of 21 eyes a pars plana vitrectomy (PPV) was performed because of marked vitreous opacification. IAG was performed before and after surgery. RESULTS: Although clinically unremarkable the iris of 14/21 eyes showed avascular zones in IAG; 13/21 had irregular vessels such as vascular collaterals, and 10/21 exhibited neovascularization. All eyes showed leakage of dye at the pupillary margin, and in 15/21 there was leakage out of peripheral iris vessels. In 2 of 9 eyes angiographic changes such as avascular zones regressed after PPV. CONCLUSION: Irides in patients with MCP that are unremarkable on slit-lamp examination may show marked angiographic changes. Thus IAG in those cases with planned cataract extraction and IOL, if necessary combined with PPV for vitreous opacification, may be warranted in order to better assess the prognosis after surgery. PMID- 10541902 TI - Corneal biopsy in keratitis performed with the microtrephine. AB - BACKGROUND: The aetiology of most cases of keratitis remains unclear because the causative agents respond to broad-spectrum antibiotics. Problems occur when they become resistant to local therapies. Further diagnostic measures such as corneal scrapings or biopsies are then necessary. In order to ensure early and gentle biopsy followed by effective diagnosis within 24 h, corneal biopsy specimens were obtained with a microtrephine. PATIENTS AND METHODS: Microbiopsies were obtained from 28 patients suffering from corneal infiltrates or ulcerative keratitis. Different stainings were used to identify the pathogens. Photographs of the clinical healing process were taken immediately after biopsy and during the follow-up. RESULTS: One hundred and ten microbiopsies were performed. One hundred and eighteen specimens could be obtained. No perforation occurred. In 5 of 10 cases in which herpetic keratitis was predicted, herpes DNA could be confirmed. The other five cases were found to be caused by other microbes. In 15 of 18 cases, the bacterial pathogen could be confirmed by Gram's stain diagnosis after microtrephination. Corneal smear was positive in only 7 of these cases. In 2 of 6 cases, predicted to be caused by fungi, lactophenol-blue staining of the microbiopsies showed positive results. Corneal smear was positive in only 1 of these 2 fungal cases. No intraoperative or postoperative complications occurred. No worsening of the disease as a result of treatment could be observed. CONCLUSIONS: The confirmation of microbial cause of keratitis is more effective using microbiopsy than with corneal smears. As a result of the effective treatment after biopsy diagnosis, the majority of cases of keratitis healed. Local therapy seems to have been optimised due to the unroofing of infection during biopsy as well. Therefore microbiopsy in combination with laboratory diagnosis may prove to be a very useful diagnostic and possibly therapeutic method in the clinical routine. PMID- 10541903 TI - Topical application of methotrexate for inhibition of corneal angiogenesis. AB - PURPOSE: Methotrexate (MTX) is a folic acid antagonist used in chemotherapy regimens. Additional therapeutic applications have been suggested based on effect as an immuno-modulating drug in systemic rheumatoid disease and associated uveitis. Since chronic inflammatory disease is often associated with a neovascular response, we investigated the use of MTX for treatment of corneal angiogenesis. METHODS: Neovascularizations were induced by fibroblast growth factor in a corneal pocket model. Vessels were examined biomicroscopically. MTX was applied topically to rabbit corneas in a concentration of 0.2 mg/day. MTX level was measured in aqueous humor and plasma. RESULTS: On day 9 the vascularized area was 12.0+/-6.9 mm(2) in control eyes and significantly smaller, 2. 2+/-1.86 mm(2), in treated eyes. Treated animals showed no local side effects such as epithelial defects. Although therapeutic levels were measured in the aqueous humor, MTX could not be detected in the serum of treated animals. CONCLUSION: The antiangiogenic mechanism of MTX might be due to inhibition of both macrophage invasion during early angiogenesis and endothelial cell proliferation. The high levels in the aqueous humor indicate a possible application of topical MTX for inflammations of the anterior segment of the eye. PMID- 10541904 TI - The role of costimulatory molecules B7-1 and B7-2 in mice with experimental autoimmune uveoretinitis. AB - BACKGROUND: Onset of experimental autoimmune uveoretinitis (EAU) is believed to involve a CD4-positive type 1 T helper cell (Th1) immune response, with inhibition involving a Th2 immune response. Development of Th1 and Th2 responses involves the participation of the costimulatory molecules B7-1 and B7-2, respectively. The purpose of this study was to investigate the role of B7-1 and B7-2 in the EAU model in mice. METHODS: B10.A mice were immunized with interphotoreceptor retinoid-binding protein (IRBP) and given daily intraperitoneal injections of either phosphate-buffered saline (control), mouse monoclonal antibody (mAb) to B7-1, mAb to B7-2, or mAb to both B7-1 and B7-2. Eyes were evaluated by histopathological criteria and cytokines were assayed in culture medium of IRBP-stimulated lymphocytes. Cellular immune responses were measured by cell proliferation assay under IRBP stimulation. RESULTS: Rates of EAU onset were 5/10 (50%) for control mice, 1/9 (11%) for mice treated with anti B7-1 mAb, 5/6 (83%) for mice treated with anti-B7-2 mAb, and 2/6 (33%) for mice treated with both anti-B7-1 and anti-B7-2 mAb. Mean histopathological severity scores were 2. 4+/-0.8, 1.0+/-0, 2.6+/-1.0, and 1.0+/-0, respectively. Production of IL-5 was significantly increased in mice treated with anti-B7-1 mAb, while IFN gamma was increased in mice treated with anti-B7-2 mAb. Spleen cell proliferation was significantly reduced in mice treated with anti-B7-1 mAb. CONCLUSIONS: These results suggest that the costimulatory molecules B7-1 and B7-2, via their influence on generating Th1 and Th2 immune responses, play an important role in the clinical outcome of EAU in mice immunized with IRBP. PMID- 10541905 TI - Human fetal retinal pigment epithelium suppresses the activation of CD4(+) and CD8(+) T-cells. AB - BACKGROUND: The suppressive effect of human fetal retinal pigment epithelium (HFRPE) on the activation of human CD4(+) and CD8(+) T-cells was evaluated in vitro. METHODS: Pure populations of CD4(+) and CD8(+) T-cells were isolated from human peripheral blood-derived buffy coats by negative immunomagnetic selection. The purity of the cells was examined by flow cytometry using anti-CD3-FITC, anti CD4-FITC, anti-CD8-PE, and anti-CD20-PE mAbs. HFRPE cells were isolated from fetal eyes and pure cultures were obtained. The effect of normal or IFN-gamma activated HFRPE cells at early (P3) or late (P6) passages on the activation of CD4(+) and CD8(+) T-cells was assessed in two different T-cell activation assays. In both activation models anti-CD3 mAb (OKT3) provided the antigen-specific signal. The secondary signal for the activation of CD4(+) and CD8(+) T-cells was provided with anti-CD18 mAb (TS1/18) and anti-CD28 mAb (9.3) in the first and the second assay respectively. Cross-linking of these soluble mAbs was performed with sheep-anti-mouse IgG-coated latex beads. The T-cell activation was determined by cell proliferation measured by [(3)H]thymidine incorporation. In each activation assay T-cells were incubated with HFRPE cells in a ratio of T-cells to HFRPE of 1:1 or 1:4. RESULTS: CD4(+) and CD8(+) T-cells were activated by cross-linking CD3 and CD18 in the first assay (CD3/CD18) and CD3 and CD28 in the second assay (CD3/CD28). In both assays HFRPE inhibited the activation of CD4(+) and CD8(+) T cells. IFN-gamma-activated HFRPE cells totally suppressed the T-cell activation at a 1:1 ratio. This suppressive effect was weaker at lower cell ratios. Some donor variation was observed in the inhibition at the lower cell ratios, especially for the inhibition of CD8(+) T-cell activation with anti-CD3/CD18. The passaging of HFRPE cells did not alter their suppressive effect on CD4(+) and CD8(+) T-cells. CONCLUSIONS: HFRPE cells suppressed the activation of both CD4(+) and CD8(+) T-cells in vitro. These findings suggest that RPE-induced immune suppression may play a significant role in maintaining immune privilege in the subretinal space. PMID- 10541906 TI - Human and porcine anterior lens capsule as support for growing and grafting retinal pigment epithelium and iris pigment epithelium. AB - PURPOSE: To establish a method for transplantation of cultured monolayers of RPE and IPE into the subretinal space, anterior lens capsule was evaluated for its suitability to serve as growth support and carrier for transplantation procedures. MATERIALS AND METHODS: Twenty-four anterior lens capsules were obtained from porcine eyes. The same number of human lens capsules was obtained during cataract surgery. Six lens capsules of each species were stored at -80 degrees C. Subsequently, the capsules were transferred onto type-I collagen. A second set of six lens capsules was treated identically except for the cryo treatment. A third set of six capsules was initially exposed to 0.05% trypsin for 30 min. Suspended porcine RPE and IPE cells (5 x 10(4) cells/well) were seeded on the top of each capsule. The remaining six lens capsules served as controls and were incubated in uncoated 12-well dishes without undergoing experimental treatment. The cultures were maintained in a water-saturated atmosphere at 37 degrees C with 5% CO(2). Six days later, viability, morphology, and cell density were determined. The capsules covered by a confluent monolayer of cells were transferred into uncoated wells and cultivated for another 10 days. At the end of the experiment, light and phase-contrast microscopy was performed on all capsules. RESULTS: Storage at -80 degrees C and exposure to trypsin resulted in significant reduction of cellular contamination. The highest cell density was found after 5 days when capsules which had undergone cryopreservation or trypsin exposure served as support for RPE and IPE. The pigment cell layer was firmly attached to the capsules and permitted a transfer to other culture flasks without significant cell loss. The IPE cell layer remained confluent after transfer to uncoated culture flasks, while the RPE cell layer ceased to proliferate 10 days after transfer. CONCLUSIONS: Lens capsules may be suitable for growing and supporting monolayers of pigment epithelial cells. Especially IPE cells formed stable monolayers on anterior lens capsules which could be transferred to secondary culture flasks without inflicting damage on the cells. PMID- 10541907 TI - Occupational medicine in Taiwan. PMID- 10541908 TI - Critical review of the epidemiology literature on the potential cancer risks of methylene chloride. AB - OBJECTIVE: To critically review and summarize the epidemiological evidence published to date on the carcinogenicity of methylene chloride to humans. METHODS: Papers for review were identified through Medline (National Library of Medicine) and were limited to epidemiology studies. Studies were classified using three categories. Primary studies focused on the association between methylene chloride and cancer among occupational cohorts primarily exposed to methylene chloride. Secondary studies identified methylene chloride a priori as a potential exposure of interest, and the investigators either characterized the methylene chloride exposure or described results for the methylene chloride-exposed workers separately. Tertiary studies evaluated cohorts either minimally exposed to methylene chloride or presumed exposed but for which no exposure estimation or separate classification was made. RESULTS: No strong or consistent finding for any site of cancer was apparent despite several studies of large occupational cohorts of workers potentially exposed to high concentrations of methylene chloride. Sporadic and weak associations were reported for cancers of the pancreas, liver and biliary passages, breast, and brain. Although these studies collectively cannot rule out the possibility of any cancer risk associated with methylene chloride exposure, they do support a conclusion of no substantive cancer risk. CONCLUSIONS: Continued follow-up of the established cohorts may elucidate the few and inconsistent relationships reported to date; however, it appears likely that risks associated with methylene chloride exposure, if any, are small and limited to rare cancers. The usefulness of additional cohort studies for the evaluation of cancer risks associated with methylene chloride exposure will depend largely on whether the relevant exposure period has passed and whether exposure characterization (e.g., peak or intermittent exposure or intensity) can be improved. PMID- 10541909 TI - Endotoxins and IgG antibodies as indicators of occupational exposure to the microbial contaminants of metal-working fluids. AB - OBJECTIVES: The aim of this study was to evaluate workers' exposure to microbes and bacterial endotoxins during the use of metal-working fluids (MWF). METHODS: Air and bulk sampling with biomonitoring of workers' serum IgG antibodies were used to estimate the exposure to biological agents at 18 workplaces. The types of emulsified MWF used were synthetic fluid, mineral oil or rape seed oil, in grinding, turning and drilling work. RESULTS: The endotoxin concentrations in the air ranged from 0.04 to 600 ng/m(3) when the endotoxin levels in MWF were 0.03 25,000 ng/ml. A high correlation was found between the endotoxin levels and the bacterial counts from MWF, as well as between the total culturable bacteria and the gram-negative bacteria concentrations in the air. Comamonas testosteroni and C. acidovorans were the most common strains in the samples but also colonies of Ochrobactrum anthropi, Pantoea agglomerans and Stenotrophomonas maltophilia were isolated from the workplaces. Fungi like Aspergillus, Cladosporium and Penicillium species were identified in the air but only rarely in the MWF. Positive IgG antibodies were found in the sera of 22 of the 25 MWF workers examined. Antibodies against S. maltophilia, P. agglomerans and C. acidovorans were the most common, appearing in 72%, 64% and 64%, respectively, of the cases. The MWF workers showed significantly higher IgG antibody responses to bacterial antigens than did the controls. CONCLUSIONS: The results clearly proved that in occupational hygiene measurements, endotoxins serve as excellent indicators of exposure to the microbial contaminants of MWF. IgG antibodies against antigens identified from workplace samples could be a practical tool for occupational health physicians. PMID- 10541911 TI - Assessment of autonomic nervous activity in hand-arm vibration syndrome patients using time- and frequency-domain analyses of heart rate variation. AB - OBJECTIVES: The aim of the present study was to non-invasively assess autonomic nervous activity, using time- and frequency-domain analyses of heart rate variation (HRV), and to investigate the relationship between indices of HRV and duration of exposure to vibration (DEV), time since retirement from work involving vibration (TR) and time undergoing treatment (TT) in a group of patients with hand-arm vibration syndrome (HAVS). SUBJECTS AND METHODS: Twenty one HAVS patients who were no longer exposed to vibration and were undergoing standard treatment for HAVS, and 10 healthy control subjects, similar to the patients in age, height, weight and number of current smokers and drinkers, volunteered for this study. Indices of HRV [time-domain indices (the mean of R-R intervals, standard deviation and coefficient of variation) and normalized units of frequency-domain indices [low frequency (LF) and high frequency (HF) components], indicating parasympathetic nervous activity, were calculated from 2 min electrocardiographic data recorded during spontaneous breathing by subjects in supine rest. RESULTS: The LF and HF components of the patients were significantly lower than those of the healthy controls (P < 0.05). When Pearson correlation analysis was applied for the patient group, using indices of HRV with age, weight, height, DEV, TR and TT, the LF components positively related to TR and TT (P < 0.01). The patients were thus divided into three groups as follows, according to TR: group A (1 to <5 years) and group C (>/=5 to 1 to <5 years) and group Z (>/=5 to CAG) has been detected in nucleotide position 1948 (codon 648) of the polymerase delta gene from Novikoff cells, resulting in an Arg to Gln change. Position 648 lies just proximal to the conserved region VI, which is part of the "fingers" subdomain of alpha-like polymerases. This subdomain is involved in dNTP binding. Upon comparison of biochemical characteristics of partially purified DNA polymerase delta from both Novikoff cells and rat liver, the following properties of the enzyme from Novikoff cells were found to be altered: (i) K(50) values for nucleotide analogs (e.g. butylphenyl-dGTP) were lower, (ii) sensitivity to various antineoplastic drugs (e.g. doxorubicin, topotecan and distamycin) was enhanced, (iii) copying fidelity was decreased when primer templates containing O(6)-methylguanine were used, and (iv) the activity of DNA polymerase delta from Novikoff tumor cells was less stimulated by lactate dehydrogenase than the enzyme from normal cells. The altered biochemical characteristics of DNA polymerase delta from Novikoff cells suggest mutator properties. We conclude that the point mutation detected in the cDNA might be causally related to the observed changes in inhibition characteristics and copying fidelity. PMID- 10541968 TI - A limited-sampling model for the pharmacokinetics of carboplatin administered in combination with paclitaxel. AB - PURPOSE: Carboplatin doses are often determined by using modified Calvert formulas. It has been observed that the area under the concentration versus time curve (AUC) for free carboplatin is lower than expected when modified formulas are used for carboplatin/paclitaxel chemotherapy combination regimens. By using limited-sampling models, the carboplatin AUC actually reached can easily be verified, and the dose adjusted accordingly. METHODS: In this report, we describe the development and validation of a limited-sampling model for carboplatin from 77 pharmacokinetic curves, when carboplatin is used in combination with paclitaxel. RESULTS: The following single-point model was selected as optimal: AUC carboplatin (min mg(-1) ml(-1)) = 418. c(2.5 h)(mg/ml) + 0.43 (min mg(-1) ml( 1)), where c(2.5 h) is the concentration (mg/ml) of carboplatin 2.5 h after the start of a 30-min infusion. This model proved to be unbiased (mean prediction error = 3.4 +/- 1.6%) and precise (root mean square error = 10.1 +/- 1.5%). CONCLUSIONS: The proposed model can be very useful for ongoing and future carboplatin/paclitaxel studies aimed to optimise and individualize treatment. PMID- 10541970 TI - Efficacy, toxicity and mode of interaction of combination radioimmunotherapy with 5-fluorouracil in colon cancer xenografts. AB - PURPOSE: The feasibility of radioimmunotherapy (RAIT) combined with 5 fluorouracil (5-FU) was examined in colon cancer xenografts. The mode of interaction of the two treatments was also investigated. METHODS: Mice bearing human colon cancer were treated with a combination of 4.63 MBq (L-RAIT) or 9.25 MBq (H-RAIT) (131)I-A7, an IgG1 against 45-kDa glycoprotein, and 5-FU at a dose of 30 mg kg(-1)day(-1) for 5 days. Myelotoxicity was monitored by blood cell counts and intestinal toxicity was assessed by the dosimetry. The results were compared with those of a single-modality therapy. RESULTS: The combination of 5 FU with H-RAIT enhanced the antitumor effect, improving the tumor quadrupling time from 25.3 +/- 9.59 days to 31.3 +/- 8.32 days (P < 0.05) and inducing tumor regression in 7 out of 10 mice, compared to 3 out of 9 mice treated with H-RAIT alone. The efficacy of L-RAIT was also improved by the combination. Analysis of the dose/response relationship showed an additive interaction of the two modalities. The combination of 5-FU with RAIT induced slightly more severe myelotoxicity than a single-modality treatment, but blood cell counts recovered similarly. Dose estimation suggested that RAIT does not increase the intestinal toxicity of 5-FU. CONCLUSION: The combination of two modalities would be feasible for the treatment of colon cancer, increasing antitumor effect with minor effect on toxicity. PMID- 10541969 TI - Melanoma-associated expression of vascular endothelial growth factor and its receptors FLT-1 and KDR. AB - The expression patterns of vascular endothelial growth factor (VEGF) and its two receptors, flt-1 and KDR, were assessed in normal human melanocytes, transformed melanocytes expressing the simian virus 40 Tgene (SV40T), and melanoma cells derived from primary and metastatic lesions. Constitutive expression of VEGF, flt 1, and KDR mRNA and proteins was observed in the majority of primary and metastatic melanoma cell lines, and in SV40T-transformed melanocytes. VEGF expression in melanoma cell lines was further enhanced by exogenous growth factors including insulin and fetal calf serum. By contrast, neonatal melanocytes did not express VEGF or VEGF receptors and VEGF expression could not be induced by exogenous growth factors. Exogenous VEGF had no significant effects on melanoma cell proliferation or on production of a transcriptional target for VEGF, urokinase-type plasminogen activator. Down-regulation of VEGF expression in the metastatic melanoma cell line WM164 through transfection of a VEGF antisense construct similarly did not affect proliferation of the transfected cells in the presence or absence of exogenous VEGF. In summary, coexpression of VEGF and its receptors is a tumor-associated phenomenon in melanoma development. However VEGF production does not support autocrine proliferation of the melanoma cell lines tested. PMID- 10541971 TI - Severe non-haematological toxicity after treatment with gemcitabine. AB - The authors report that 4 out of a series of 56 patients (7.1%) treated with gemcitabine developed an unexplained non-cardiogenic pulmonary distress syndrome most likely related to gemcitabine. One further patient developed ventricular arrhythmia immediately after gemcitabine exposure, leading to cardiac arrest. Between 1995 and 1998 56 patients with locally advanced or metastatic carcinoma were treated with gemcitabine. The patients suffered from breast cancer (n = 17), pancreatic cancer (n = 17), lung cancer (n = 12), cancer of unknown primary (n = 5), ovarian cancer (n = 2), oral cavity cancer (n = 2) and cancer of the bladder (n = 1). Their median age was 55 years, and there were 33 female and 23 male patients. Fifteen patients had been pretreated with radiation therapy: 2 had received radiation therapy involving the mediastinum as treatment for non-small cell lung cancer and cancer of unknown origin respectively, 11 patients had had prior neoadjuvant or adjuvant radiation therapy of the chest wall for breast cancer and 2 patients had received radiation therapy for head/neck cancer. All patients received gemcitabine on days 1, 8 and 15 and this was repeated on day 29 at a dose of 1000 mg/m(2) as a 30-min infusion in 250 ml isotonic NaCl. In 4 patients gemcitabine treatment was combined with cisplatinum, in 7 patients with a somatostatin analogue and in 1 patient with epirubicin. All other patients received gemcitabine as a single agent. We assume that the pulmonary or cardiac toxicity of 5 patients was related to gemcitabine. In 3 patients re-exposure resulted in repeated toxicity. One patient did not receive gemcitabine again because of the life-threatening nature of the primary response. Two patients had received prior radiation to the mediastinum with 62 Gy and 50 Gy respectively, 3 years and 1 year before gemcitabine application. In our experience pulmonary toxicity after gemcitabine treatment is more common than initially anticipated. Gemcitabine should be used with caution in patients who have received prior radiation to the mediastinum. PMID- 10541972 TI - E-cadherin expression in oesophageal carcinoma treated with high-dose radiotherapy; correlation with pretreatment parameters and treatment outcome. AB - BACKGROUND AND PURPOSE: E-cadherin plays an important role in the cell-cell contact of normal epithelium. Loss of E-cadherin expression may be related to tumour invasiveness and metastatic potential. In a group of patients treated for oesophageal carcinoma by radiotherapy only, we found that immunohistochemically detected p53 expression correlated with reduced survival, mainly because of the occurrence of distant metastases. We questioned whether, in this group of patients, E-cadherin expression was concomitantly altered and served as a predictive factor for the development of distant metastases. MATERIALS AND METHODS: Immunostaining for E-cadherin was performed on paraffin- embedded biopsy specimens from patients with adenocarcinoma and squamous cell carcinoma of the oesophagus. E-cadherin status and its correlation with regard to pretreatment parameters and treatment outcome were determined. RESULTS: An aberrant staining pattern of E-cadherin did not correlate with any of the pretreatment parameters. In a univariate analysis, a significantly reduced metastatic potential was found for tumours that had an aberrant cellular staining pattern for E-cadherin, which was strongest for squamous cell carcinomas. However, in a multivariate analysis only p53 status correlated significantly with the occurrence of distant metastases. CONCLUSION: Although, in univariate analysis, aberrant E-cadherin expression served as a better, rather than a worse prognostic factor, p53 status remained the only significant parameter in multivariate analysis, in this group of patients with oesophageal carcinoma. No relationship between p53 status and E cadherin expression was found. PMID- 10541973 TI - Expression of CD44 variants in osteosarcoma. AB - The standard form of CD44 (CD44H) is a transmembranous glycoprotein, widely distributed on a variety of human lymphoid cells, epithelial cells and tumours. CD44 has many variant forms, which are generated by alternative splicing. In recent years, CD44 has been reported to be related to the degree of tumour differentiation, tumour cell invasion, and metastasis. We investigated 44 tumour specimens in 39 patients with osteosarcoma immunochemically to analyse the expression of CD44 standard (CD44H) and variant exon-encoded gene products (CD44v3, v4, v5, v6, v7, v9, and v10). Furthermore, the relationship between CD44 expression and the clinical outcome of patients with osteosarcoma was analysed. Membrane accentuation and exclusive cytoplasmic reactivity were analysed as separate staining patterns. Tumour cells and some multinucleated giant cells were markedly stained. CD44H, v3, v4, v5, v6, v7, v9, and v10 were expressed in 85%, 49%, 54%, 59%, 46%, 5%, 28%, and 10% of the specimens respectively. The cumulative 5-year metastasis-free survival was 58% in CD44v6-negative cases and 24% in CD44v6-positive cases (P=0.046). However, the cumulative 5-year metastasis free survival was not significantly different between cases positive and negative for other variants of CD44. Multivariate analysis (Cox proportional-hazard model) with CD44v6 expression (positive or negative), chemotherapy (intensive or non intensive), tumour site (proximal or distal), and age (at least 30 years or less than 30 years) showed that expression of CD44v6 and chemotherapy were important prognostic factors in patients with osteosarcoma. Overexpression of CD44 isoforms containing variant v6 is correlated with poor prognosis in patients with osteosarcoma. PMID- 10541974 TI - Psychotropics in supportive care: first assess, then prescribe. PMID- 10541976 TI - Awareness and acceptance of dying in cancer patients and their relatives. PMID- 10541975 TI - Vascular access investigation comes of age. PMID- 10541977 TI - Caring for terminally ill patients. The Johannes Hospiz in Munich: a palliative care unit in Southern Bavaria, Germany. PMID- 10541978 TI - Psychopharmacology in supportive care in cancer: a review for the clinician. I. Benzodiazepines. AB - Benzodiazepines are widely utilized in the cancer setting. Most aspects relevant to their use in supportive care are reviewed to assist the clinical oncologist in prescribing such drugs. This review covers pharmacokinetics, indications, adverse effects and drug interactions, and compares the profiles of different benzodiazepines. Finally, controversial issues with regard to benzodiazepines are discussed. PMID- 10541979 TI - Intravascular catheters impregnated with antimicrobial agents: a milestone in the prevention of bloodstream infections. AB - Vascular catheters impregnated with antimicrobial agents have been shown to decrease the risk of catheter-related colonization and bloodstream infections. Various antimicrobials and antiseptics have been used. In a recent meta-analysis of 12 studies, catheters coated with chlorhexidine and silver sulfadiazine (CH/SS) were shown to be significantly less likely to be associated with catheter related bloodstream infections than uncoated catheters. However, these catheters were coated only on the external surface and they are associated with short antimicrobial durability (3-7 days). In addition, anaphylactic reactions to them were reported in Japan. Vascular catheters impregnated with minocycline and rifampin (M/R) were found to be highly efficacious in preventing catheter-related infections. In a recent prospective, randomized trial, the likelihood of catheter related bloodstream infections associated with the use of M/R catheters was one twelfth of that observed with catheters coated with CH/SS. The M/R catheters are coated on the external and internal surfaces and have an antimicrobial durability of 4 weeks. Although no resistance to either minocycline or rifampin has been seen in two trials, further studies are required to determine whether the risk of resistance outweighs the benefits derived from their use. In conclusion, antimicrobial catheters have been shown to be highly cost effective in decreasing the risk of catheter-related bloodstream infection. PMID- 10541980 TI - Strategies for prevention of catheter-related bloodstream infections. AB - Prevention of catheter-related bloodstream infections is critically dependent on an accurate knowledge of the two main routes by which intravascular devices become contaminated: the extraluminal (skin-related) and the intraluminal (hub related) routes. Extraluminal catheter seeding results from infection of the catheter entry site by microorganisms and leads to bacteremia most often during the week following catheter placement. The main ways of preventing it are appropriate skin disinfection and the adoption of maximal antiseptic barriers at the time of catheter insertion. Avoiding the internal jugular and the femoral veins, whenever possible, will reduce the likelihood of bacteremia. Intraluminal contamination is the consequence of improper handling of the catheter hub at the time of connection and disconnection of the administration set. It is the most common origin of catheter infections after the first week of catheter placement. Multiple-lumen catheters, side-ports and multipurpose catheters particularly increase the risk of endoluminal contamination. To prevent it, strict asepsis should be observed in hub handling and hubs should be protected against environmental soiling with an antiseptic impregnated gauze at all times. New technology is available for prevention of catheter infections: antibiotic and antiseptic-coated catheters, antiseptic hubs, disinfecting caps and flushing solutions are currently undergoing scientific assessment. PMID- 10541981 TI - Psychological impact of informed consent in hospitalized cancer patients. A sequential study of anxiety and depression using the hospital anxiety and depression scale. AB - Patients with cancer receive an explanation of their disease and the recommended treatment when they are asked to give informed consent (IC). In the course of this process patients suffer severe distress, including anxiety and depression, but physicians tend to underestimate it. The goal of this study was to reveal the magnitude of such stress and any changes to this during the IC process by means of the Hospital Anxiety and Depression (HAD) scale, a self-assessment scale. Of 171 in-patients newly diagnosed with lung cancer, 119 were assessable for serial HAD scale scores on admission, immediately after the IC process, and at 1 and again at 2 weeks after the IC. Both anxiety and depression scores increased significantly immediately after IC. Female patients had significantly higher anxiety and depression scores than males at 1 week after the IC. The patients with poor performance status demonstrated high anxiety scores on admission and immediately after the IC, and substantial depression persisted longer in these patients. The prevalence of high scores of more than 11 (judged as adjustment disorder or more severe state) immediately after the IC was 50% for anxiety and 31% for depression. The prevalence decreased significantly within 1 or 2 weeks, but 41% and 14% of the patients still showed high anxiety and depression scores, respectively. Physicians should be aware of these facts and pay special attention to their patients' psychological distress in routine clinical practice. PMID- 10541982 TI - The European Organization for Research and Treatment of Cancer Quality of Life Questionnaire (EORTC QLQ-C30): translation and validation study of the Iranian version. AB - The objective of this study was to test the reliability and validity of the Iranian version of the European Organization for Research and Treatment of Cancer (EORTC) Quality of Life Questionnaire (QLQ-C30). The English-language version of the questionnaire was translated into Persian (Iranian language), and its final form was approved by the EORTC Study Group on Quality of Life before it was used in this study. The questionnaire was administered at two time points to a consecutive sample of 168 newly diagnosed breast cancer patients, and almost all of them (99%) found the questions easy to understand and acceptable. Crohnbach's alpha coefficient for multi-item scales (to test reliability) ranged from 0.48 to 0.95 at baseline and from 0.52 to 0.98 at follow-up administration of the questionnaire. Validity was checked using two methods: inter-scale correlation and known-groups comparison. Almost all inter-scale correlations were statistically significant in the expected direction. Known-groups comparison analysis showed that all functioning and symptom scales discriminated between subgroups of patients differing in clinical status as defined by their performance status and disease stage. In general, the findings of this study indicate that the Iranian version of the EORTC QLQ-C30 is a reliable and valid measure of quality of life in cancer patients and can be used in clinical trials and studies of outcome research in oncology. PMID- 10541983 TI - Quantity and quality of information desired by Portuguese cancer patients. AB - Disclosure of the diagnosis of cancer to the patients affected has always been a controversial issue in the doctor-patient relationship. Undoubtedly this is so not only because of differences between countries and cultures, but also because there have been changes of opinion over the years. The aim of this study was to evaluate the quality and quantity of information desired by Portuguese cancer patients, and how and from whom they want to hear this information. Our sample comprised a total of 193 cancer patients, 87 men and 106 women. We found that 68.9% knew what their diagnosis was. In our sample, 74% wanted "as much information as possible, good or bad"; 85% said they wanted to know if their disease was cancer; 95% wanted to know the best or worst likely outcome of their disease; and 96.4% wanted to know the chances of getting cured. Most patients said they would prefer to be informed by physicians (92.7%) and have access to a telephone help-line, books and television. In conclusion, most patients wanted to know as much as possible about their illness and treatment, and the majority preferred to be involved in treatment decisions. PMID- 10541984 TI - Antiemetic therapy for high-dose chemotherapy with transplantation: report of a retrospective analysis of a 5-HT(3) regimen and literature review. AB - The goal of this work was to evaluate the efficacy of 5-HT(3) receptor antagonist regimens in the transplant setting through a retrospective analysis of clinic patients and a review of the literature. A retrospective study was performed to evaluate the efficacy of an antiemetic regimen in 24 patients receiving high-dose chemotherapy with bone marrow transplantation. The antiemetic agent used in 96% of the patients was single-agent ondansetron 0.15 mg/kg as a continuous infusion every 12 h starting on the first day of chemotherapy. Efficacy or failure was defined by the need for administration of rescue antiemetics, as documented by nurses' notes and pharmacy records. Overall, 23 (96%) patients experienced antiemetic failure within the first 3 days. Rescue antiemetics yielded improved control of nausea and vomiting in 16 of 23 (70%) patients. These data demonstrate poor antiemetic efficacy of single-agent, continuous-infusion ondansetron in a small group of patients receiving high-dose chemotherapy with bone marrow transplantation. These data are considered in the context of a thorough literature review that shows the limitations of previously reported antiemetic studies in this setting, potential pharmacokinetic interactions, and also highlights the utilization of combination therapy, administered orally or intravenously, to improve efficacy and tolerability of antiemetic therapy. A number of recent investigations suggest that appropriate doses of 5-HT(3) receptor antagonists with phenothiazine and corticosteroids can improve the safety and efficacy of antiemetic regimens in patients undergoing transplantation. PMID- 10541985 TI - Serologic evidence of heparin sensitization in cancer patients receiving heparin flushes of venous access devices. AB - Cancer patients with venous access devices (VADs) often receive daily flushes of heparin. Even this relatively small heparin exposure has been reported to induce immune-mediated thrombocytopenia. To estimate how frequently this occurs we tested for heparin-related antibodies in 49 patients receiving daily heparin flushes of their VADs. Although one-third of the patients showed evidence of heparin sensitization on at least one occasion during their surveillance, their antibody titers were generally low and typical of those found in other cohorts of patients who become sensitized to heparin but do not develop secondary thrombocytopenia. However, one patient developed a positive serotonin release assay indicative of a more significant sensitization, but without thrombocytopenia. Therefore, our observations suggest that heparin-induced thrombocytopenia (HIT) related to heparin flushes of VADs is uncommon but still an important diagnosis to entertain. PMID- 10541986 TI - Nosocomial candidaemias due to species other than Candida albicans in cancer patients. Aetiology, risk factors, and outcome of 45 episodes within 10 years in a single cancer institution. AB - Forty-five cases of fungaemia due non-albicans Candida spp. (NAC) in a single National Cancer Institution within 10 years were analysed for aetiology, risk factors and outcome. There had been 12 cases of fungaemia that were due to C. krusei, 14 due to C. parapsilosis, 7 due to C. (T.) glabrata, 6 to C. tropicalis, 2 to C. guillermondii, 2 to C. lusitaniae, 1 to C. stellatoidea, and 1 to C. rugosa. Comparison of 45 NAC fungaemia with 75 episodes of C. albicans fungaemia revealed differences only in two risk factors: previous empiric therapy with amphotericin B (16.0 vs 2.2%, P<0.01) appeared more frequently in cases of C. albicans fungaemia, and prior prophylaxis with fluconazole (8.9 vs 0%, P<0.02) was conversely more frequently observed with NAC. The incidence of other risk factors, such as underlying disease, chemotherapy, antibiotic prophylaxis or therapy, treatment with corticosteroids, catheter insertion, mucositis, cytotoxic chemotherapy, and neutropenia, was similar in both groups. There was no difference either in attributable or in overall mortality between NAC and C. albicans fungaemia in our cancer patients. PMID- 10541987 TI - Successful antidepressant treatment for five terminally ill cancer patients with major depression, suicidal ideation and a desire for death. AB - In the debate on euthanasia and physician-assisted suicide, we have to exclude terminally ill patients in whom the desire for death is caused by major depression. However, it is still not clear to what degree major depression can be treated by psychiatric intervention in this setting. We evaluated the effect of antidepressant treatment in terminally ill cancer patients. Six cancer patients with suicidal ideas thought to be due to major depression were treated with tricyclic antidepressants. Three had requested terminal sedation to relieve them from their suffering. The median survival of five of these patients was 4 weeks after diagnosis; one was lost to follow-up. The efficacy of the antidepressant treatment was assessed using the Hamilton Rating Scale for Depression (HRSD). One week after the start of treatment with antidepressants, five of the six patients showed a marked improvement in their mood and showed no further suicidal thoughts or requests for terminal sedation. The average reduction in the HRSD score was 23.4 points (14-38; SD = 9. 9). Antidepressant treatment can be effective in alleviating the desire for death due to major depression, even in terminally ill cancer patients. PMID- 10541988 TI - General practitioners caring for terminally ill patients resident in a hospice. AB - The goal of this study was to investigate why GPs have little or no involvement in the medical process relating to any of their patients when they are admitted to a palliative care unit (PCU) and what solutions they suggest. The study took the form of a descriptive pilot study based on a short questionnaire. It emerged that GPs felt their involvement was influenced by their job description, by practical factors (time investment, distance between practice and PCU, remuneration, referral) and personal issues (e.g. dealing with dying). It is concluded that GPs need education in palliative/supportive care and approved remuneration as well as knowledge about their task in the PCU. PMID- 10541989 TI - Forthcomming meetings PMID- 10542017 TI - Classical swine fever virus is genetically stable in vitro and in vivo. AB - Phylogenetic analyses of large numbers of classical swine fever strains have revealed a high degree of sequence conservation in the genomic regions examined, suggesting either a recent common ancestor or a low evolution rate. This low variability is in contrast to findings with other RNA viruses. To investigate the consequence of this apparent genetic stability on phylogenetic examinations, the Belgian field isolate Wingene'93 was passaged in pigs as well as in cell culture by various methods. Sequence analyses of viruses collected after various passages in three target regions proposed for phylogenetic studies (5' NTR, E2, and NS5B) revealed a complete sequence conservation. Only when the amount of passaged virus was lowered, mimicking a genetic bottleneck, a single point mutation was observed in the E2 gene. Additionally, only four nucleotide substitutions were observed when the genome of a virus obtained after 96 cell passages in persistently infected cells was compared with its parental virus, the recombinant virus derived from an infectious cDNA clone of CSFV strain Alfort/187. This low mutation frequency observed both in vitro and in vivo demonstrates that classical swine fever virus is genetically stable. Hence, even minor mutations can be considered significant in molecular epidemiological studies. PMID- 10542018 TI - Determination of 3'-terminal nucleotide sequence of pepper vein banding virus RNA and expression of its coat protein in Escherichia coli. AB - Pepper vein banding virus (PVBV) is an important virus infecting chilli pepper in south India. Earlier reports suggested it to be a distinct potyvirus. The nucleotide sequence of PVBV RNA from the 3'-end (3862 nt) was determined. Analysis of the nucleotide and deduced amino acid sequence revealed that it encompasses a partial open reading frame encoding the partial sequence of VPg, NIa-protease, NIb, coat protein (CP) and 3'-untranslated region (UTR). Comparison of the amino acid sequence of CP and the nucleotide sequence of 3'-UTR with those of other potyviruses confirmed an earlier observation that PVBV is a distinct member of the Potyvirus sub-group and it had significant similarity to a recently characterized virus infecting chilli pepper, chilli vein-banding mottle virus (CVbMV), from Thailand. The analysis showed that both PVBV and CVbMV might represent strains of the same virus. Further, the PVBV CP gene was overexpressed in E. coli, which assembled into potyvirus-like particles (PVLPs). The assembled particles were shown to encapsidate the CP mRNA. PMID- 10542019 TI - Amino acid substitutions at position 481 differently affect the ability of the measles virus hemagglutinin to induce cell fusion in monkey and marmoset cells co expressing the fusion protein. AB - The hemagglutinin (H) protein of the measles virus (MV) Edmonston strain induced cell fusion in Cos (monkey) and B95a (marmoset) cells, when co-expressed with the fusion (F) protein, whereas the H protein of the wild-type KA strain induced fusion in B95a cells, but not in Cos cells. Asparagine residue at position 481 of the KA H protein was replaced by various amino acids through site-directed mutagenesis. Substitution with tyrosine, which was found at position 481 of the Edmonston H protein, enabled the mutant KA H protein (N481Y) to induce cell fusion in Cos cells co-expressing the F protein, which could be completely blocked by anti-CD46 antibody. This mutant, however, did not cause CD46 downregulation, unlike the Edmonston H protein. The other H protein mutants (N481S, N481T, N481D, N481H, N481F) did not produce syncytia in Cos cells. On the other hand, all of the mutants retained the ability to induce cell fusion in B95a cells. Thus, while tyrosine at position 481 was indispensable for the MV H protein's interaction with CD46, the residue at this position does not appear to be critically involved in the interaction with the receptor for wild-type strains present on B95a cells. PMID- 10542020 TI - E. coli expressed proteins as diagnostic reagents for typing of foot-and-mouth disease virus. AB - Truncated proteins corresponding to the C-terminal half of VP1 of four vaccine strains and two field variants of foot-and-mouth disease virus (FMDV) were expressed in E. coli. The expressed proteins were affinity purified and their type specific reactivity was confirmed by immunoprecipitation with anti-virus antibodies. Antibodies were raised against the purified proteins in guinea pigs and the type specificity of the anti peptide antibodies was confirmed by antigen capture reverse transcription polymerase chain reaction (Ag-RT/PCR) where the sera against a particular type captured the homologous virus. Antibodies were purified by immuno-affinity chromatography and tested for specificity by various serological tests. Using the purified proteins and the antibodies raised against them, tests like ELISA, Ag-RT/PCR, and latex agglutination test (LAT) were standardized. Application of the reagents in various tests was studied by screening a few field samples and by nucleotide sequencing. Specific reactivity of antibodies raised against expressed protein was seen with both vaccine virus and field samples. Thus E. coli expressed proteins and antibodies to them may form an alternative and cheap source of diagnostic reagents. The studies showed that antibodies against peptides were mono-specific and therefore may be used in LAT for rapid typing of FMDV and Ag-RT/PCR for typing ELISA negative field samples. PMID- 10542021 TI - Human parvovirus B19 infection in acute fulminant liver failure. AB - We previously reported detection of human parvovirus B19 DNA in livers from patients requiring transplantation for acute fulminant liver failure. In this study, we used immune adherence PCR (IA-PCR) to bind B19 virions in recipient native liver onto solid phase with specific monoclonal antibodies followed by PCR amplification of virion DNA. IA-PCR had sensitivity and specificity similar to conventional PCR. We examined liver tissue from 16 patients with non-A, non-B, non-C, non-E (NA-E) acute fulminant liver failure (AFLF) (6 of unknown etiology associated with aplastic anemia (AA), 4 of unknown etiology without AA; and 6 patients with AFLF of known etiology). IA-PCR detected B19 virions in 5 of 6 (83%) of livers from patients with idiopathic NA-E AFLF associated with AA and in 2 of 3 (75%) without AA, compared to 1 of 6 (17%) of livers from patients with AFLF of known etiology and to 6 of 34 (18%) of 34 control patients with chronic or neoplastic liver disease. Viral mRNA encoding the structural protein was detected in the liver tissue from three B19 IA-PCR positive patients with AFLF. Detection of B19 virions and mRNA for capsid proteins provided strong evidence for B19 infection during the course of NA-E AFLF and argues for involvement of B19 virus in liver injury. PMID- 10542022 TI - Characterization and expression of the coat protein-coding region of banana bract mosaic potyvirus, development of diagnostic assays and detection of the virus in banana plants from five countries in southeast Asia. AB - We have sequenced the entire coat protein (CP)-coding region and 5' 162 nucleotides of the 3' untranslated region (UTR) of nine different isolates of banana bract mosaic virus (BBrMV) from five different countries. Further, we have sequenced the 3' 621 nucleotides of the NIb-coding region of a Philippines isolate. This is the first report of BBrMV in Thailand, Vietnam and Western Samoa. When the sequences of the CP-coding region and 3' UTR were compared to each other, variability of between 0.3% and 5.6%, and 0.3% and 4. 3%, was observed at the nucleotide and amino acid levels, respectively. Phylogenetic analysis of the BBrMV isolates did not reveal any relationship between the geographic location of the isolates. The BBrMV CP was expressed in Escherichia coli as a fusion protein and the purified recombinant protein was used to produce a high titre BBrMV-specific polyclonal antiserum. This antiserum was used to develop a F(ab')(2) indirect double antibody sandwich ELISA and compared with immuno-capture PCR (IC-PCR) and reverse transcription PCR (RT-PCR) assays for BBrMV detection. RT-PCR was shown to be the most sensitive test followed by ELISA and IC-PCR. http://link.springer. de/link/service/journals/00705/bibs/9144009/91441725.htmPfr conversion. The strongest effect of mutation on the chromoprotein assembly, leading to an almost complete loss of the chromophore binding capability, was found for the exchanges of His322 by Leu (H322L) and Pro318 by Lys (P318K), whereas a corresponding alanine mutant (P318A) showed wild-type behavior. The second histidine (H319) is also involved in chromophore fixation, as indicated by a slower assembly rate upon mutation (H319L). For the other mutants, an assembly process very similar to that of the wild-type protein was found. The light induced Pr-->Pfr conversion kinetics is altered in the mutations H319L and S320K and in the double mutant L323R/Q324D, all of which exhibited a significantly faster I700 decay and accelerated Pfr formation. P318 is also involved in the Pr- >Pfr conversion, the millisecond steps (formation of Pfr) being significantly slower for P318A. Lacking sufficient amounts of W366F, assembly kinetics could not be determined in this case, while the fully assembled mutant underwent the Pr ->Pfr conversion with kinetics similar to wild-type protein. PMID- 10542064 TI - Tissue variant effects of heme inhibitors on the mouse cytochrome c oxidase gene expression and catalytic activity of the enzyme complex. AB - The in vivo effects of heme biosynthesis inhibitors, succinylacetone and CoCl2 on the cytochrome c oxidase (COX) gene expression and enzyme activity in different mouse tissues were investigated. Succinylacetone and CoCl2 showed tissue-specific differences in their ability to modulate heme aa3 content. A single dose of succinylacetone treatment for 8 h reduced the heme aa3 content of kidney mitochondria with no effect on the liver. CoCl2 treatment for 8 h, however, selectively affected the heme aa3 level in the liver. Reduced mitochondrial heme aa3 with both treatments was accompanied by approximately 50% reduced, mitochondrial genome-encoded COX I and II mRNAs and nuclear genome-encoded COX Vb mRNAs, but no change in COX IV mRNA level. Use of isolated mouse liver and brain mitochondrial systems showed a 50-80% reduction in mitochondrial transcription and translation rates in heme-depleted tissues. Blue native gel electrophoresis followed by immunoblot analysis showed that the complex from heme-depleted tissues contained a 30-50% reduction in levels of subunits I, IV, Vb and near normal levels of subunit VIc, indicating altered subunit content. Treatment of submitochondrial particles with protein kinase A and ATP resulted in partial dissociation of COX, suggesting a mechanistic basis for the reduced subunit content of the complex from heme-depleted tissues. Surprisingly, the enzyme from heme-depleted tissues showed twofold to fourfold higher turnover rates for cytochrome c oxidation, suggesting alterations in the kinetic characteristics of the enzyme following heme reduction. This is probably the first evidence that the tissue heme level regulates not only the mammalian COX gene expression, but also the catalytic activity of the enzyme, probably by affecting its stability. PMID- 10542067 TI - Lack of oxygen sensing by mitochondria in platelets. AB - The range over which cells are sensitive to changes in oxygen concentration remains uncertain. Wilson and colleagues [Wilson, D.F. (1994) Med. Sci. Sports Exerc. 26, 37-43] have suggested that cytochrome oxidase is sensitive to oxygen concentrations below about 40 microM, but proposed that this sensitivity is obscured in intact cells because an increase in reduction state of cytochrome c acts to maintain oxygen consumption. We have tested this hypothesis in platelets, which are small cells (2-4 micrometer diameter, < 0.5 micrometer thick) that do not decrease their rate of oxygen consumption until oxygen concentrations fall below 2.5 microM. Contrary to the expectations of the hypothesis, the reduction state of cytochrome c, the concentration of NADH and the rate of glycolytic output are not changed as oxygen concentration declines from 40 microM down to 5 microM. Therefore, we conclude that at least some cell types contain mitochondria that are not capable of sensing oxygen above 5 microM by the mechanism proposed by Wilson and colleagues. PMID- 10542066 TI - Isolation and characterization of hydrophobic polypeptides in human bile. AB - Polypeptides were isolated from human bile by extraction with chloroform/methanol, followed by reversed-phase chromatography in methanol/ethylene chloride and gel filtration in chloroform/methanol. Peptides were characterized by SDS/PAGE, sequence analysis and matrix-assisted laser desorption ionization/time-of-flight mass spectrometry. This identified haemoglobin alpha chain, ATP synthase lipid-binding protein subunit 9, an N terminal fragment of mac25/insulin-like growth factor-binding protein 7 and an internal fragment of monocyte differentiation antigen CD14, all not described previously in bile. In addition, alpha1-antitrypsin, known in bile from previous work, was also identified. The hydrophobic character of haemoglobin alpha chain is not apparent from its amino acid sequence, but the other polypeptides all have major hydrophobic segments. These results show that several proteins are removed upon organic solvent extraction used for delipidation during the preparation of samples for proteome analysis. Several of the polypeptides found are unexpectedly present in bile, suggesting that specific excretion mechanisms may be involved. PMID- 10542068 TI - Determination of the sites required for the allosteric inhibition of serine acetyltransferase by L-cysteine in plants. AB - Serine acetyltransferase (SATase; EC 2.3.1.30) catalyzes the formation of O acetylserine from L-Ser and acetyl-CoA in plants and bacteria. In plants, two types of SATase have been described. One is allosterically inhibited by L-Cys, and the second is not sensitive to L-Cys inhibition. However, the allosteric site in SATase has not been identified. To understand better the mechanism of L-Cys inhibition of plant SATases, we constructed several chimeric SATase enzymes from watermelon SATase (WaSATase) (sensitive type) and Arabidopsis SAT-p (insensitive type). These enzymes were expressed in Escherichia coli, and inhibition of the mutated SATase activity by L-Cys was analyzed. Mutated WaSATase, in which Met280 was changed to Ile, was no longer inhibited by L-Cys. Analysis of the inhibition the chimeric enzymes indicated that the C-terminal region of WaSATase from Pro276 to Phe285, in which five amino acids are different from those of SAT-p, was responsible for the determination of the sensitivity to L-Cys. In particular, Gly277 in the C-terminal region of WaSATase was primarily responsible for the L Cys inhibition. The N-terminal half of the protein, which does not contain the catalytic domain, was also important for the sensitivity to L-Cys. These results indicate that the sensitivity of SATase to L-Cys is due to the N-terminal and C terminal regions rather than to the catalytic domain. PMID- 10542069 TI - GroEL is involved in activation of Escherichia coli RNA polymerase devoid of the omega subunit in vivo. AB - Highly purified Escherichia coli RNA polymerase contains a small subunit termed omega that has a molecular mass of 10 105 Da and is comprised of 91 amino acids. E. coli strains lacking omega (omega-less) are viable, but exhibit a slow-growth phenotype. Renaturation of RNA polymerase isolated from an omega-less mutant, in the presence of omega, resulted in maximum recovery of activity. The omega-less RNA polymerase from omega-less strains recruits the chaperonin, GroEL (unlike the wild-type enzyme), suggesting a structural deformity of the mutant enzyme. The GroEL-containing core RNA polymerase interacts efficiently with sigma70 to generate the fully functional holoenzyme. However, when GroEL was removed, the enzyme was irreversibly nonfunctional and was unable to bind to sigma70. The damaged enzyme regained activity after going through a cycle of denaturation and reconstitution in the presence of omega or GroEL. GroES was found to have an inhibitory effect on the core-sigma70 association unlike the omega subunit. The omega subunit may therefore be needed for stabilization of the structure of RNA polymerase. PMID- 10542070 TI - Transcriptional control is relevant in the modulation of mosquito ferritin synthesis by iron. AB - In yellow fever mosquito cells (Aag2 clone), iron treatment induces a threefold increase in ferritin message (fer mRNA) and protein (ferritin) by 16 h. These data contrast with work in mammalian hepatocytes and fibroblasts in which fer mRNA levels do not change with iron stimulation, but ferritin levels increase 50 fold. Pretreatment of the Aag2 cells with actinomycin D blocks induction of fer mRNA and reduces the ferritin subunit synthesis, suggesting that iron induction of ferritin subunit synthesis is subjected to transcriptional control. A putative iron-regulatory protein has also been identified in cytoplasmic extracts from Aag2 cells. PMID- 10542071 TI - Structural elucidation of the O-antigenic polysaccharides from Escherichia coli O21 and the enteroaggregative Escherichia coli strain 105. AB - The structure of the O-antigen polysaccharide of the lipopolysaccharide from an enteroaggregative Escherichia coli (strain 105) has been elucidated, using primarily one-dimensional and two-dimensional NMR experiments. The sequence of residues was deduced with heteronuclear multiple-bond correlation and NOESY experiments. The structure of the repeating unit of the polysaccharide from the enteroaggregative E. coli is as follows:[sequence: see text] The structure of the O-antigen from enteroaggregative E. coli strain 105 was shown to be identical with that of E. coli O21 by sugar and methylation analyses as well as by 1H-NMR and 13C-NMR spectroscopy. PMID- 10542072 TI - Structural studies utilizing 13C-enrichment of the O-antigen polysaccharide from the enterotoxigenic Escherichia coli O159 cross-reacting with Shigella dysenteriae type 4. AB - The structure of the O-antigen polysaccharide from Escherichia coli O159 has been determined using primarily NMR spectroscopy of the 13C-enriched polysaccharide. The sequence of the sugar residues could be determined by heteronuclear multiple bond connectivity NMR experiments. The polysaccharide is composed of a pentasaccharide repeating unit with the following structure: [sequence: see text] Matrix assisted laser desorption ionization mass spectrometry was performed on intact lipopolysaccharide and from the resulting molecular mass the O-antigen part was estimated to contain approximately 23 repeating units. Cross-reactivity of this O-antigen to that of Shigella dysenteriae type 4 was confirmed using enzyme-linked immunoabsorbant assay. PMID- 10542073 TI - Expression and compartmentation of the glucose repressor CRE1 from the phytopathogenic fungus Sclerotinia sclerotiorum. AB - The glucose repressor from the phytopathogenic fungus Sclerotinia sclerotiorum is encoded by the cre1 gene. Polyclonal antibodies were raised against a fusion protein (gluthathione S-transferase) GST-CRE1 in order to study cre1 expression. Western blot analyses revealed that CRE1 synthesis is regulated by the nature of the extracellular carbon source. High CRE1 levels are induced by glucose and remain stable after transfer into pectin medium, suggesting the existence of post translational mechanisms which inactivate CRE1 to allow transcription of glucose repressed genes. Subcellular fractionation demonstrated that CRE1 is localized in the nuclei of glucose grown hyphae and in the cytoplasm when glucose is removed from the culture medium. CRE1 fused to green fluorescent protein (GFP) was introduced into Aspergillus nidulans. Fluorescence microscopy showed the nuclear localization of the GFP-CRE1 fusion protein according to the presence of glucose in the culture medium, suggesting homologous post-translational regulations of glucose repressors in fungi. We propose that filamentous fungi regulate the activity of the glucose repressor by controlling its nuclear translocation. PMID- 10542074 TI - RNA hairpin loops repress protein synthesis more strongly than hammerhead ribozymes. AB - A general study has been carried out to determine how well hammerhead ribozymes might reduce levels of specific protein synthesis in living cells, compared with RNA hairpin loops as stable but noncleaving controls. Four different experiments are described. First, a wide variety of hammerhead ribozymes, as well as hairpin loops, was cloned into a gene-expression cassette for beta-galactosidase, upstream of the coding sequences for that reporter gene, and expressed from plasmids in several strains of Escherichia coli. The results show that ribozymes, when acting intramolecularly in E. coli, do not significantly reduce the amount of protein synthesized from any construct. As a control, long RNA hairpin loops do greatly reduce the amount of protein made. Secondly, we studied the transcription-translation of these same plasmids in a cell extract from E. coli. Once again, hammerhead ribozymes show no effect on levels of beta-galactosidase, whereas long RNA hairpin loops produce a strong reduction, by apparent attentuation at the level of translation. Thirdly, we added an SV40 promoter to each plasmid, in order to study the effects of these gene-regulators on protein synthesis in Chinese hamster ovary cells. Here active intramolecular ribozymes produce a slight reduction in beta-galactosidase, whereas long RNA hairpin loops produce an even stronger reduction than before. Those hairpin loops apparently induce degradation of their own mRNA in Chinese hamster ovary cells, by a mechanism not seen in E. coli. Finally, analyses of total RNA by S1-trimming show that hammerhead ribozymes will self-cleave a mRNA by a total of no more than 45 50% in E. coli, compared with 70-80% in vitro. Other analyses using Northern blotting were unable to detect any ribozyme cleavage in E. coli or Chinese hamster ovary cells. In summary, the ability of hammerhead ribozymes to reduce protein synthesis appears weak or nonexistent in all the cellular systems tested. By comparison, long RNA hairpin loops reduce protein synthesis strongly: by an apparent attentuation mechanism in E. coli or by a novel degradation of their own mRNA in Chinese hamster ovary cells. PMID- 10542075 TI - Functional analysis of the 5'-flanking region of FTA for expression of rat GDP-L fucose:beta-D-galactoside 2-alpha-L-fucosyltransferase. AB - The tissue-specific and species-specific expression of the ABH antigens is well known among vertebrate species and it is regulated by the alpha(1,2)fucosyltransferase that forms the H antigen, a precursor of the A and B antigens. To investigate the mechanisms governing the tissue-specific and species specific expression of this alpha(1,2)fucosyltransferase, we characterized the gene structure, including the promoter region, of FTA, a rat orthologous homolog of human FUT1 that encodes the H alpha(1, 2)fucosyltransferase and is responsible for the expression of the ABH antigens on human red blood cells. Northern blot and 5'-RACE analyses suggested that at least two forms of FTA mRNA (2.9 and 2.6 kb), which use alternative transcription start sites, are present in the cancer cell lines RCN-9 (rat colon cancer) and PC12 (rat pheochromocytoma), whereas only the 2.6 kb form was detected in normal colon, stomach and pancreas. Transcriptional activity of the 5'-flanking sequence, which contains three putative Sp1-binding sites, but lacks both TATA and CAAT boxes, was examined. Transient transfection experiments of promoter-reporter gene constructs showed high promoter activity in RCN-9, PC12 and human colon cancer (WiDr) cell lines, weak activity in human vascular endothelial (ECV304) cells and no activity in human erythroleukemia (HEL) cells. The results suggest that the 5'-flanking region of FTA contains a tissue-specific promoter. Deletional analysis of the 5' flanking sequence revealed regions containing cell-type-specific positive acting element(s) and negative regulatory element(s), which are related to the promoter activity. PMID- 10542076 TI - A Drosophila gene encoding multiple splice variants of Kazal-type serine protease inhibitor-like proteins with potential destinations of mitochondria, cytosol and the secretory pathway. AB - A Drosophila gene (KAZ1), mapped to cytological position 61A1-2 on chromosome 3, has been cloned and found to encode multiple splice variants of Kazal-type serine protease inhibitor-like proteins. KAZ1 consists of five exons and four alternatively retained introns to produce six transcripts of type AB, C1, C2, C3, D and E. The AB transcript contains two ORFs, of which the upstream one produces a polypeptide alpha, which has a mitochondrial sorting signal. Localization to mitochondria was confirmed by expression in COS1 cells. The downstream ORF is shared partially with type C1, C2, C3, D and E transcripts and produces polypeptides beta, gamma, delta and epsilon when expressed in Drosophila cells. Type C1, C2 and C3 transcripts differ only in the 5'-noncoding sequence and thus all produce type gamma. Polypeptides gamma and epsilon have a signal sequence at their N-termini and are secreted into the medium while beta and delta lack this sequence and remain in the cytoplasm. Isoforms beta and epsilon share a common C terminal sequence distinct from that shared by polypeptides gamma and delta. The N-terminal sequences of isoforms beta to epsilon contain a PEST region which could induce rapid intracellular degradation of isoforms beta and delta. Sequence analysis of the Kazal-type domain suggests a similar folding pattern as observed for rhodniin and SPARC/BM-40. Northern analysis and in situ hybridization showed that the type C3 transcript is predominant and the expression is highest in midgut at larval stage. PMID- 10542077 TI - The presence of the alternatively spliced A2 cassette in the vacuolar H+-ATPase subunit A prevents assembly of the V1 catalytic domain. AB - Vacuolar ATPases (V-ATPases) are multisubunit enzymes that couple the hydrolysis of ATP to the transport of H+ across membranes, and thus acidify several intracellular compartments and some extracellular spaces. Despite the high degree of genetic and pharmacological homogeneity of V-ATPases, cells differentially modulate the lumenal pH of organelles and, in some cells, V-ATPases are selectively targetted to the plasma membrane. Although the mechanisms underlying such differences are not known, the subunit isoform composition of V-ATPases could contribute to altered assembly, targeting or activity. We previously identified an alternatively spliced variant of the chicken A subunit in which a 30 amino acid cassette (A1) containing the Walker consensus sequence for ATP binding is replaced by a 24 amino acid cassette (A2) that lacks this feature. We have examined the ability of chimeric yeast/chicken A subunits containing either the A1 or the A2 cassette to restore the V-ATPase activity of yeast that lack the A subunit. The A1-containing chimeric subunit, but not the chimera that contains the A2 cassette, partially restores the ability of the mutated yeast to grow at neutral pH. Both chimeric proteins are expressed, although at lower levels than the similarly transfected yeast A subunit. The A2-containing subunit fails to associate with the vacuolar membrane or support the assembly of V-ATPase complexes. Thus, the substitution of the A1 sequence by A2 not only removes the Walker nucleotide binding sequence but also compromises the ability of the A subunit to assemble with other V-ATPase subunits. PMID- 10542078 TI - Two branches of the lupeol synthase gene in the molecular evolution of plant oxidosqualene cyclases. AB - Two new triterpene synthase cDNAs, named as OEW and TRW, were cloned from olive leaves (Olea europaea) and from dandelion roots (Taraxacum officinale), respectively, by the PCR method with primers designed from the conserved sequences found in the known oxidosqualene cyclases. Their ORFs consisted of 2274 bp nucleotides and coded for 758 amino acid long polypeptides. They shared high sequence identity (78%) to each other, while they showed only about 60% identities to the known triterpene synthases LUPI (lupeol synthase clone from Arabidopsis thaliana) and PNY (beta-amyrin synthase clone from Panax ginseng) at amino acid level. To determine the enzyme functions of the translates, they were expressed in an ERG7 deficient yeast mutant. Accumulation of lupeol in the cells of yeast transformants proved both of these clones code for lupeol synthase proteins. An EST (expression sequence tag) clone isolated from Medicago truncatula roots as a homologue of cycloartenol synthase gene, exhibits high sequence identity (75-77%) to these two lupeol synthase cDNAs, suggesting it to be another lupeol synthase clone. Comparatively low identity (approximately 57%) of LUP1 from Arabidopsis thaliana to either one of these clones leaves LUP1 as a distinct clone among lupeol synthases. From these sequence comparisons, now we propose that two branches of lupeol synthase gene have been generated in higher plants during the course of evolution. PMID- 10542079 TI - Deficient coacervation of two forms of human tropoelastin associated with supravalvular aortic stenosis. AB - Human tropoelastin associates by coacervation and is subsequently cross-linked to make elastin. In Williams syndrome, defective elastin deposition is associated with hemizygous deletion of the tropoelastin gene in supravalvular aortic stenosis (SVAS). Remarkably, point-mutation forms of SVAS correspond to incomplete forms of tropoelastin which include in-frame termination by nonsense mutations, yet the resulting phenotype of these disorders is not explained because expression variably occurs from both normal and mutant alleles. Proteins corresponding to two truncated tropoelastin mutants were expressed and purified to homogeneity. Coacervation of these proteins occurred as expected with increasing temperature, but substantially contrasted with that of the performance of a normal tropoelastin. Significantly, association by coacervation of the truncated SVAS tropoelastin molecules was negligible at 37 degrees C, which contrasted with the substantial coacervation seen for normal tropoelastin. Furthermore their midpoints of coacervation increased and correlated with the extent of deletion, in accord with the loss of hydrophobic regions required for tropoelastin association. Their secondary structures are similar, as evidenced by CD studies. We propose a model for point-mutation SVAS in which aberrant tropoelastin molecules are incompetent and are mainly excluded from participation in coacervation and consequently in elastogenesis. These forms of SVAS may consequently be considered functionally similar to a hemizygous deletion, and mark point-mutation SVAS as a disorder of defective coacervation. PMID- 10542080 TI - The best customer is an educated customer. PMID- 10542081 TI - Finding the right fold. AB - Using mutational analysis, three groups have compared the transition states for the folding of two pairs of homologous proteins. The results of these studies suggest that protein folding mechanisms are conserved and are defined primarily by the overall topology of the native structures, as opposed to specific details of the interactions stabilizing these structures. PMID- 10542082 TI - A chaperone with a hydrophilic surface. AB - The folding of native tubulin involves at least seven different chaperone proteins: prefoldin, the cytosolic chaperonin CCT and five tubulin-specific chaperone proteins named cofactors A-E. The structure of the yeast homolog of cofactor A, Rbl2p, shows it to be a dimer with largely hydrophilic surfaces, reflecting the fact that it interacts with quasi-native, not unfolded, beta tubulin. PMID- 10542083 TI - Training ribozymes to switch. AB - Ribozymes that are sensitive to cyclic nucleotides have been selected in vitro. Remarkably, the cGMP-dependent ribozymes are specifically activated by a factor of 5,000 - the largest allosteric ribozyme activation by a small molecule seen to date. PMID- 10542084 TI - Folding alphabets. AB - A new computational approach optimizes searches for reduced protein folding alphabets that use fewer than 20 types of amino acids. The predicted optimal five letter alphabet happens to be in agreement with the suggestive results of a recent experiment, but whether highly reduced alphabets are sufficient for truly protein-like properties remains an open experimental question. PMID- 10542087 TI - The ribosome revealed. AB - Several recently reported structures reveal the details of ribosome architecture and provide new insights into the mechanism of protein synthesis. PMID- 10542088 TI - Picture story. Keeping HIV out. PMID- 10542089 TI - Prebiotic chemistry. PMID- 10542090 TI - Mutational analysis of acylphosphatase suggests the importance of topology and contact order in protein folding. AB - Muscle acylphosphatase (AcP) is a small protein that folds very slowly with two state behavior. The conformational stability and the rates of folding and unfolding have been determined for a number of mutants of AcP in order to characterize the structure of the folding transition state. The results show that the transition state is an expanded version of the native protein, where most of the native interactions are partially established. The transition state of AcP turns out to be remarkably similar in structure to that of the activation domain of procarboxypeptidase A2 (ADA2h), a protein having the same overall topology but sharing only 13% sequence identity with AcP. This suggests that transition states are conserved between proteins with the same native fold. Comparison of the rates of folding of AcP and four other proteins with the same topology, including ADA2h, supports the concept that the average distance in sequence between interacting residues (that is, the contact order) is an important determinant of the rate of protein folding. PMID- 10542091 TI - The folding transition state between SH3 domains is conformationally restricted and evolutionarily conserved. AB - The protein engineering analysis of the alpha-spectrin SH3 domain at three different stability conditions (pH 7.0, 3.5 and 2.5) reveals a folding transition state structured around the distal loop beta-hairpin and the 310-helix. This region is impervious to overall changes in protein stability, suggesting a transition state ensemble with little conformational variability. Comparison with the Src SH3 domain (36% sequence homology) indicates that the transition state in this protein family may be conserved. Discrepancies at some positions can be rationalized in terms of the different interactions made by the different side chains in both domains. Bronsted plot analysis confirms the straight phi(doubledagger-U) results and shows two folding subdomains for this small protein. These results, together with previous data on circular permutants of the alpha-spectrin SH3 domain, indicate that polypeptide topology and chain connectivity play a major role in the folding reaction of this protein family. PMID- 10542092 TI - Experiment and theory highlight role of native state topology in SH3 folding. AB - We use a combination of experiments, computer simulations and simple model calculations to characterize, first, the folding transition state ensemble of the src SH3 domain, and second, the features of the protein that determine its folding mechanism. Kinetic analysis of mutations at 52 of the 57 residues in the src SH3 domain revealed that the transition state ensemble is even more polarized than suspected earlier: no single alanine substitution in the N-terminal 15 residues or the C-terminal 9 residues has more than a two-fold effect on the folding rate, while such substitutions at 15 sites in the central three-stranded beta-sheet cause significant decreases in the folding rate. Molecular dynamics (MD) unfolding simulations and ab initio folding simulations on the src SH3 domain exhibit a hierarchy of folding similar to that observed in the experiments. The similarity in folding mechanism of different SH3 domains and the similar hierarchy of structure formation observed in the experiments and the simulations can be largely accounted for by a simple native state topology-based model of protein folding energy landscapes. PMID- 10542093 TI - Single protein misfolding events captured by atomic force microscopy. AB - Using single protein atomic force microscopy (AFM) techniques we demonstrate that after repeated mechanical extension/relaxation cycles, tandem modular proteins can misfold into a structure formed by two neighboring modules. The misfolding is fully reversible and alters the mechanical topology of the modules while it is about as stable as the original fold. Our results show that modular proteins can assume a novel misfolded state and demonstrate that AFM is able to capture, in real time, rare misfolding events at the level of a single protein. PMID- 10542094 TI - Crystal structure of the post-chaperonin beta-tubulin binding cofactor Rbl2p. AB - The folding pathway of tubulins includes highly specific interactions with a series of cofactors (A, B, C, D and E) after they are released from the eukaryotic chaperonin CCT. The 2.2 A crystal structure of Rbl2p, the Saccharomyces cerevisiae homolog of beta-tubulin specific cofactor A, shows alpha helical monomers forming a flat, slightly convex dimer. The surface of the molecule is dominated by polar and charged residues and lacks hydrophobic patches typically observed for chaperones that bind unfolded or partially folded proteins. This post-chaperonin cofactor is therefore clearly distinct from typical chaperones where hydrophobicity is a hallmark of substrate recognition. PMID- 10542095 TI - A computational approach to simplifying the protein folding alphabet. AB - What is the minimal number of residue types required to form a structured protein? This question is important for understanding protein modeling and design. Recently, an experimental finding by Baker and coworkers suggested a five residue solution to this problem. We were motivated by their results and by the arguments of Wolynes to study reductions of protein representation based on the concept of mismatch between a reduced interaction matrix and the Miyazawa and Jernigan (MJ) matrix. We find several possible simplified schemes from the relationship of minimized mismatch versus the number of residue types (N = approximately 2-20). As a specific case, an optimal reduction with five types of residues has the same form as the simplified palette of Baker and coworkers. Statistical and kinetic features of a number of sequences are tested. Comparison of results from sequences with 20 residue types and their reduced representations indicates that the reduction by mismatch minimization is successful. For example, sequences with five types of residues have good folding ability and kinetic accessibility in model studies. PMID- 10542096 TI - Solution structure of the hRPABC14.4 subunit of human RNA polymerases. AB - The protein hRPABC14.4 is an essential subunit of human RNA polymerases I, II, and III and is required for the transcription of all human nuclear genes. The structure of hRPABC14.4 was determined by nuclear magnetic resonance spectroscopy. The protein fold comprises a highly conserved central domain forming two antiparallel alpha-helices flanked by the less conserved N- and C terminal regions forming a five-stranded beta-sandwich. Amino acids from the two helices participate in the generation of a hydrophobic surface area which is conserved in all eukaryotic and archaeal homologous subunits, and likely constitutes a critical macromolecular interaction interface. The hRPABC14.4 structure accounts for mutagenesis results in Saccharomyces cerevisiae and provides a structural working model for elucidating the role of this subunit in the molecular architecture and function of the human nuclear RNA polymerases. PMID- 10542097 TI - Arginase-boronic acid complex highlights a physiological role in erectile function. AB - The crystal structure of the complex between the binuclear manganese metalloenzyme arginase and the boronic acid analog of L-arginine, 2(S)-amino-6 boronohexanoic acid (ABH), has been determined at 1.7 A resolution from a crystal perfectly twinned by hemihedry. ABH binds as the tetrahedral boronate anion, with one hydroxyl oxygen symmetrically bridging the binuclear manganese cluster and a second hydroxyl oxygen coordinating to Mn2+A. This binding mode mimics the transition state of a metal-activated hydroxide mechanism. This transition state structure differs from that occurring in NO biosynthesis, thereby explaining why ABH does not inhibit NO synthase. We also show that arginase activity is present in the penis. Accordingly, the tight binding and specificity of ABH allows us to probe the physiological role of arginase in modulating the NO-dependent smooth muscle relaxation required for erection. Strikingly, ABH causes significant enhancement of nonadrenergic, noncholinergic nerve-mediated relaxation of penile corpus cavernosum smooth muscle, suggesting that arginase inhibition sustains L arginine concentrations for NO synthase activity. Therefore, human penile arginase is a potential target for therapeutic intervention in the treatment of erectile dysfunction. PMID- 10542098 TI - Structure of the TRAIL-DR5 complex reveals mechanisms conferring specificity in apoptotic initiation. AB - TRAIL, an apoptosis inducing ligand, has at least four cell surface receptors including the death receptor DR5. Here we report the crystal structure at 2.2 A resolution of a complex between TRAIL and the extracellular region of DR5. TRAIL forms a central homotrimer around which three DR5 molecules bind. Radical differences in the surface charge of the ligand, together with variation in the alignment of the two receptor domains confer specificity between members of these ligand and receptor families. The existence of a switch mechanism allowing variation in receptor domain alignment may mean that it is possible to engineer receptors with multiple specificities by exploiting contact positions unique to individual receptor-ligand pairs. PMID- 10542099 TI - Calculating the electrostatic properties of RNA provides new insights into molecular interactions and function. AB - Solutions to the nonlinear Poisson-Boltzmann equation were used to obtain the electrostatic potentials of RNA molecules that have known three-dimensional structures. The results are described in terms of isopotential contours and surface electrostatic potential maps. Both representations have unexpected features: 'cavities' within isopotential contours and areas of enhanced negative potential on molecular surfaces. Intriguingly, the sites of unusual electrostatic features correspond to functionally important regions, suggesting that electrostatic properties play a key role in RNA recognition and stabilization. These calculations reveal that the electrostatic potentials generated by RNA molecules have a variety of functionally important characteristics that cannot be discerned by simple visual inspection of the molecular structure. PMID- 10542100 TI - Allosteric selection of ribozymes that respond to the second messengers cGMP and cAMP. AB - RNA transcripts containing the hammerhead ribozyme have been engineered to self destruct in the presence of specific nucleoside 3',5'-cyclic monophosphate compounds. These RNA molecular switches were created by a new combinatorial strategy termed 'allosteric selection,' which favors the emergence of ribozymes that rapidly self-cleave only when incubated with their corresponding effector compounds. Representative RNAs exhibit 5,000-fold activation upon cGMP or cAMP addition, display precise molecular recognition characteristics, and operate with catalytic rates that match those exhibited by unaltered ribozymes. These findings demonstrate that a vast number of ligand-responsive ribozymes with dynamic structural characteristics can be generated in a massively parallel fashion. Moreover, optimized allosteric ribozymes could serve as highly selective sensors of chemical agents or as unique genetic control elements for the programmed destruction of cellular RNAs. PMID- 10542101 TI - The energetics of T4 lysozyme reveal a hierarchy of conformations. AB - We have used native state exchange to examine the energy landscape of the well characterized protein T4 lysozyme. Although the protein exhibits two-state behavior by traditional probes, the energy landscape determined here is much more complex. The average stability of the C-terminal subdomain is significantly higher than that for the N-terminus suggesting at least two regions of unfolding. At a more detailed level, there appears to be a broad continuum of stabilities throughout each region. The overall subdomain hierarchy of energies does not mirror data on the folding pathway for this protein, challenging the relationship between energy landscapes and folding trajectories. PMID- 10542102 TI - A method for extraction of high-quality and high-quantity genomic DNA generally applicable to pathogenic bacteria. AB - In this study, we report a modified procedure for extraction of high-quality genomic DNA that is rapid, simple, biologically nonhazardous, and generally applicable to pathogenic bacteria. Bacterial cells were pretreated with 70% ethanol prior to enzymatic digestion with lysozyme. Exposure of bacterial cells to 70% ethanol sterilized the cultures, making the process biologically safe and increased the susceptibility of the cells to lysozyme-induced lysis. Consistently high yields of genomic DNA (mean average yield, 0.5-2.5 mg/ml) were obtained from 465 isolates representing over 30 clinically important bacterial species. Genomic DNA obtained was determined to be suitable for further analysis, including bacterial fingerprinting techniques like restriction endonuclease analysis, Southern hybridization, and repetitive PCR. Availability of a generally applicable procedure for extraction of high-quality and high-quantity genomic DNA would be immensely beneficial for laboratories engaged in molecular surveillance of nosocomial and community-based outbreaks. PMID- 10542103 TI - Determination of kinetic isotope effects for nucleoside hydrolases using gas chromatography/mass spectrometry. AB - Kinetic isotope effects are widely used to determine the transition state of chemical and enzymatic reactions. Radioactive isotopes are used most often to determine these kinetic isotope effects. However, stable isotopes offer a number of advantages over the use of radioactive isotopes. These advantages include ease of handling and disposal along with increased safety in the laboratory. [1' (13)C]Inosine and [1'-(2)H]inosine kinetic isotope effects were determined using a gas chromatograph in conjunction with a mass selective detector for nucleoside hydrolase, a purine-metabolizing enzyme. Three ion pairs were used to determine kinetic isotope effects. These ion pairs were 158/159, 187/188, and 217/218. The average isotope effects for all ion pairs were 1.021 +/- 0.006 for [1' (13)C]inosine and 1.113 +/- 0.008 for [1'-(2)H]inosine. The transition state consistent with these isotope effects is also consistent with the transition state proposed by Schramm and Horenstein using radioactive substrates. PMID- 10542104 TI - Development of a capillary electrophoresis assay based on free sulfate determination for the direct monitoring of sulfoesterase activity. AB - A capillary electrophoresis assay of sulfoesterase activity was developed that overcomes the main drawbacks encountered with the usual methods for sulfate determination in complex biological medium. Conditions are described allowing direct measurement of inorganic sulfate that is enzymatically produced in the reaction mixture. The main features of this method are electrokinetic sample introduction, which allows selective extraction of sulfate from the matrix into the separation capillary, counter-electroosmotic flow migration mode, indirect absorbance detection and use of an internal standard for quantitative performances. Likewise, perfect linearity was obtained for concentrations of sulfate up to 40 ppm. The limits of detection and quantification were 0.2 and 0.6 ppm, respectively. The run-to-run and day-to-day precision are 1 and 4.5%, respectively, for sulfate concentrations varying from 35 ppm down to 1 ppm. The accuracy was established for the synthetic p-nitrocatechol sulfate substrate by comparison with the classical spectrophotometric assay. The method was applied to the kinetic monitoring of the activity of a sulfoesterase extracted from the marine mollusc Pecten maximus on fucoidan, a bioactive sulfated fucose-based polysaccharide derived from brown algae. For the first time, a sulfoesterase activity was shown to be effective on such sulfated polysaccharides. PMID- 10542105 TI - Mass spectrometry to characterize the binding of a peptide to a lipid surface. AB - The binding of an amphipathic alpha-helical peptide to small unilamellar lipid vesicles has been examined using chemical derivitization and mass spectrometry. The peptide is derived from the sequence of human apolipoprotein C-II (apoC-II), the protein activator of lipoprotein lipase (LpL). ApoC-II(19-39) forms approximately 60% alpha-helix upon binding to model egg yolk phosphatidylcholine small unilamellar vesicles. Measurement of the affinity of the peptide for lipid by spectrophotometric methods is complicated by the contribution of scattered light to optical signals. Instead, we characterize the binding event using the differential labeling of lysine residues by the lipid- and aqueous-phase cross linkers, disuccinimidyl suberate (DSS) and bis(sulfosuccinimidyl) suberate (BS(3)), respectively. In aqueous solution, the three lysine residues of the peptide are accessible to both cross-linkers. In the presence of lipid, the C terminal lysine residue becomes inaccessible to the lipid-phase cross-linker DSS, but remains accessible to the aqueous-phase cross-linker, BS(3). We use mass spectrometry to characterize this binding event and to derive a dissociation constant for the interaction (K(d) = 5 microM). We also provide evidence for the formation of dimeric cross-linked peptide when high densities of peptide are bound to the lipid surface. PMID- 10542106 TI - 32P-postlabeling assay for the quantification of the major platinum-DNA adducts. AB - To allow more sensitive, selective, and routine analyses of platinum(Pt)-GG and AG intrastrand cross-links we have significantly improved our quantitative (32)P postlabeling assay (M. J. P. Welters et al. Carcinogenesis 18, 1767-1774, 1997). Instead of off-line scintillation counting we introduced an on-line flow radioisotope detector into the HPLC system. Furthermore, the isolation protocol for the adducts was significantly modified and optimized to reduce interfering background peaks that prevented quantification of low levels of the cisplatin-DNA adducts in white blood cells obtained from patients. Reduction of background signals was obtained by boiling the samples, followed by phenol/chloroform/isoamylethanol extraction after the DNA digestion step. The labeling efficiency for the adducts was increased by 40% by using Na-formate instead of NH(4)-formate for elution of the adducts from the strong cation exchange columns. Finally, a calibration curve and quality controls were implemented. The labeling efficiencies were not different between the dinucleotides. The between- and within-run precision for the Pt-GG and Pt-AG adducts measured at the lower limit of quantification of 87 and 53 amol/microg DNA, respectively, was less than 20% CV. The adducts were stable in DNA stored for a 2-month time period at -80 degrees C. The assay is now routinely used for high-precision analyses of patient and cell line samples containing very low adduct levels. PMID- 10542107 TI - Mass accuracy and sequence requirements for protein database searching. AB - To elucidate the role of high mass accuracy in mass spectrometric peptide mapping and database searching, selected proteins were subjected to tryptic digestion and the resulting mixtures were analyzed by electrospray ionization on a 7 Tesla Fourier transform mass spectrometer with a mass accuracy of 1 ppm. Two extreme cases were examined in detail: equine apomyoglobin, which digested easily and gave very few spurious masses, and bovine alpha-lactalbumin, which under the conditions used, gave many spurious masses. The effectiveness of accurate mass measurements in minimizing false protein matches was examined by varying the mass error allowed in the search over a wide range (2-500 ppm). For the "clean" data obtained from apomyoglobin, very few masses were needed to return valid protein matches, and the mass error allowed in the search had little effect up to 500 ppm. However, in the case of alpha-lactalbumin more mass values were needed, and low mass errors increased the search specificity. Mass errors below 30 ppm were particularly useful in eliminating false protein matches when few mass values were used in the search. Collision-induced dissociation of an unassigned peak in the alpha-lactalbumin digest provided sufficient data to unambiguously identify the peak as a fragment from alpha-lactalbumin and eliminate a large number of spurious proteins found in the peptide mass search. The results show that even with a relatively high mass error (0.8 Da for mass differences between singly charged product ions), collision-induced dissociation can help identify proteins in cases where unfavorable digest conditions or modifications render digest peaks unidentifiable by a simple mass mapping search. PMID- 10542108 TI - The use of chromogenic reference substrates for the kinetic analysis of penicillin acylases. AB - Determination of kinetic parameters of penicillin acylases for phenylacetylated compounds is complicated due to the low K(m) values for these substrates, the lack of a spectroscopic signal, and the strong product inhibition by phenylacetic acid. To overcome these difficulties, a spectrophotometric method was developed, with which kinetic parameters could be determined by measuring the effects on the hydrolysis of the chromogenic reference substrate 2-nitro-5 [(phenylacetyl)amino]benzoic acid (NIPAB). To that end, spectrophotometric progress curves with NIPAB in the absence and presence of the phenylacetylated substrates and their products were measured and analyzed by numerical fitting to the appropriate equations for competing substrates with product inhibition. This analysis yielded kinetic constants for phenylacetylated substrates such as penicillin G, which are in close agreement with those obtained in independent initial velocity experiments. Using NIPAB analogs with lower k(cat)/K(m) values, kinetic parameters for the hydrolysis of cephalexin and penicillin V were determined. This method was suitable for determining the kinetic constants of penicillin acylases in periplasmic extracts from Escherichia coli, Alcaligenes faecalis, and Kluyvera citrophila. The use of chromogenic reference substrates thus appears to be a rapid and reliable method for determining kinetic constants with various substrates and enzymes. PMID- 10542110 TI - Simultaneous assay of Src SH3 and SH2 domain binding using different wavelength fluorescence polarization probes. AB - pp60(c-src) is a prototypical nonreceptor tyrosine kinase and may play a role in diseases as diverse as cancer and osteoporosis. In Src, the SH3 domain (Src homology 3) binds proteins at specific, proline-rich sequences, while the SH2 domain (Src homology 2) binds phosphotyrosine-containing sequences. Inhibition of Src SH3 and SH2 domain function is of potential therapeutic value because of their importance in signaling pathways involved in disease states. We have developed dual-wavelength fluorescent peptide probes for both the Src SH3 and the Src SH2 domains, which allow the simultaneous measurement of compounds binding to each domain in assays based on the technique of fluorescence polarization. We demonstrate the utility of these probes in a dual-binding assay (suitable for high-throughput screening) to study the interactions of various peptides with these domains, including a sequence from the rat protein p130(CAS) which has been reported to bind simultaneously to both Src SH3 and SH2 domains. Utilizing this dual-binding assay, we confirm that sequences from p130(CAS) can simultaneously bind Src via both its SH3 and its SH2 domains. We also use the dual-binding assay as an internal control to identify substances which inhibit SH3 and SH2 binding via nonspecific mechanisms. PMID- 10542109 TI - Measurement of responses from Gi-, Gs-, or Gq-coupled receptors by a multiple response element/cAMP response element-directed reporter assay. AB - We have established a rapid, sensitive, high-throughput assay that requires one assay condition to detect agonist effects from Gi-, Gs-, and Gq-coupled receptors. We utilized a vector containing a promoter with three multiple response elements, the vasoactive intestinal peptide promoter and a cAMP response element controlling the transcription of the luciferase gene. An adrenergic agonist, para-aminoclonidine, inhibited forskolin-stimulated luciferase expression when cells were cotransfected with the Gi-coupled alpha(2)-C adrenergic receptor and the MRE/CRE reporter vector. Further, we demonstrate that gastrin-releasing peptide, which activates a Gq-coupled GRP receptor, isoproterenol, which activates a Gs-coupled beta-adrenergic receptor, calcium ionophores, and phorbol 12-myristate 13-acetate, a stimulator of protein kinase C, can mediate increases in luciferase expression in the presence of forskolin but not in its absence. The effect at Gi-coupled receptor activation correlates with the phosphorylation of the CRE binding protein (CREB); however, the mechanisms mediating the responses to Gq- and Gs-coupled receptors are more complex. We demonstrate that this assay is useful for pharmacological analysis of both agonists and antagonists and has the potential to associate orphan G-protein coupled receptors with their corresponding ligands. PMID- 10542111 TI - Colorimetric determination of cell numbers by Janus green staining. AB - A colorimetric assay using the basic azo dye Janus green has been developed to assess cell numbers in anchorage-dependent cell cultures, with special regard to the enumeration of osteoblastic cells. Therefore, cells are fixed in ethanol and stained with a 0.2% solution of Janus green for 3 min, followed by a destaining step of 1 min in tap water. The addition of diluted hydrochloric acid easily and immediately leads to dye elution from stained cell layers into the acidic supernatant which consequently is transferred into 96-well plates and read on a microplate reader at 595 nm. Working under standardized conditions, Janus green uptake in several cell lines is shown to be linearly correlated with cell numbers over a broad range of cell densities, in MC3T3-E1 cells from about 3% up to more than 300% of confluency. Absolute sensitivity of the assay allows detection of less than 1000 cells/cm(2). In comparison to many other colorimetric assays, the Janus green technique is simple to perform, fast, precise, stable, cheap, and well suited for processing large quantities of samples. Moreover, it is applicable to any culture formate and size, from irregular formed carriers up to 96-multiwell plates. PMID- 10542112 TI - Rapid detection of CYP1A1, CYP2D6, and NAT variants by multiplex polymerase chain reaction and allele-specific oligonucleotide assay. AB - Drugs and carcinogens are excreted from the body after metabolic conversion involving enzymes mediating oxidative metabolism and conjugation. Many of the corresponding genes exhibit functional polymorphisms that contribute to individual cancer susceptibility. To increase the efficiency and to facilitate genotyping, we developed a combined approach (PCR-ASO) which includes multiplex PCR and allele-specific oligonucleotide (ASO) hybridization. PCR primer pairs were used to amplify the following alleles/variants: CYP1A1*1, *2A, *2B; CYP2D6*3, *4; NAT1*4, *3, *10, *11, *14, *15; and NAT2*4, *5A, *5B, *5C, *6A, *7B. The products were dot-blotted and polymorphisms were detected by hybridization with ASO probes for both wild-type and variant sites in parallel. This approach was validated by genotyping DNA samples from a French-Canadian population that was previously analyzed by PCR-RFLP. The variants frequencies were compared with the data on other populations available in the literature. The PCR-ASO assay appears to be simple, efficient, and cost-effective, particularly if a large number of samples are to be screened for several DNA variants. This approach has potential for automation with microplates and robotic workstations for high throughput. PMID- 10542113 TI - Identification of N-acetyltransferase 2 genotypes by continuous monitoring of fluorogenic hybridization probes. AB - Three polymorphic sites in the N-acetyltransferase 2 (NAT2) gene were detected using rapid cycle DNA amplification with allele-specific fluorescent probes and melting curve analysis. Two fluorogenic adjacent hybridization probes were designed to NAT2*5A (C(481)T), NAT2*6A (G(590)A), and NAT2*7A (G(857)A). During amplification, probe hybridization is observed as fluorescence resonance energy transfer. The fluorescence increases every cycle as the product accumulates during amplification. A single base mismatch resulted in a melting temperature shift (T(m)) of 5 to 6 degrees C, allowing for the easy distinction of a wild type allele from the mutant allele. The protocol is rapid, requiring 40 min for the completion of 45 cycles including the melting curves. It is also a simple and flexible method, since DNA templates prepared from different sources, including DNA from serum and paraffin-embedded tissue sections, could be used without adverse effects. Fluorescence genotyping of all three polymorphisms in a total of 155 DNA samples correlated perfectly with our previously validated genotyping by restriction enzyme digestion (PCR-RFLP). This new facile approach allows for the easy detection of NAT2 polymorphisms in hundreds of samples in only a day. PMID- 10542114 TI - Characterization of recombinant human monoclonal tissue necrosis factor-alpha antibody using cation-exchange HPLC and capillary isoelectric focusing. AB - Cation-exchange liquid chromatography (CIEX) and capillary isoelectric focusing (cIEF) methods have been developed for the routine analysis of a recombinant, human anti-tumor necrosis factor monoclonal antibody D2E7. Both of these methods can separate heavy-chain C-terminal variants of this antibody. Various enzymatic digestion methods have also been developed for the identification of the antibody C-termini lysine (Lys) variants. A comparison of conventional CIEX-HPLC and cIEF methods has been made for the analysis of antibodies. cIEF can also be used to determine the isoelectric points (pI) of antibody variants based on the use of internal pI standards. Different C-termini Lys variants have been separated and collected from the CIEX column and subsequently analyzed by cIEF and mass spectrometry. PMID- 10542116 TI - High-resolution electrophoretic DNA fragment analysis using Spreadex gels. PMID- 10542115 TI - Characterization of novel peptide agonists of the alpha mating factor of Saccharomyces cerevisiae. AB - Alpha-factor [WHWLQLKPGQPMY], a secreted tridecapeptide pheromone, is required for mating between the a- and alpha-haploid mating types of Saccharomyces cerevisiae (MATa, MATalpha). New analogues of alpha-factor were synthesized and evaluated by morphogenesis assays and receptor binding studies. The Y(0)Nle(12)F(13) analogue [YWHWLQLKPGQPNleF] (MFN5) caused growth arrest and morphological alteration in MATa cells in a fashion identical to that of the native pheromone. Binding of (125)I-labeled MFN5 was saturable, and reversible as shown by equipotent label displacement by MFN5 and native alpha-mating factor. Scatchard analysis of equilibrium binding data on plasma membranes and intact cells indicated the existence of a single high-affinity binding site (K(d) = 6.4 x 10(-8)). Specific binding of (125)I-labeled MFN5 was significantly reduced by guanosine nucleotides. Affinity cross-linking of (125)I-labeled MFN5 to MATa cell membranes identified a specifically labeled 49-kDa protein. The novel synthetic alpha-factor analogue MFN5 can be easily iodinated and used as a probe for the alpha-factor receptor. PMID- 10542117 TI - Quantitative nonisotopic immuno-dot-blot method for the assessment of cellular poly(ADP-ribosyl)ation capacity. PMID- 10542118 TI - Separation and analysis of plasmid denatured forms using hydrophobic interaction chromatography. PMID- 10542119 TI - Single-step method for estimating nanogram quantities of protein. PMID- 10542120 TI - Preparation of heteroduplex DNA containing a mismatch base pair with magnetic beads. PMID- 10542121 TI - Salivary duct carcinoma in five cats. AB - Carcinomas of salivary gland ducts are described in five cats. The typical histological pattern was the formation of large cell aggregates resembling dilated ducts, often with central necrosis and a looping pattern. All tumours were labelled with antibody to cytokeratins (CKs) 5, 6, 8, 14, 17 and 19. Labelling of tumour cells with CK14 suggested basal cell differentiation. All tumours stained with Jack bean (Canavalia ensiformis) agglutinin (Con A); this is a feature of normal salivary gland ducts but is seen in other salivary gland tumours. Staining of tumour cells at the luminal surface of ductal structures with wheat germ (Triticum vulgaris) agglutinin (WGA) in the cat tumours was similar to that seen in ducts of normal cat salivary glands but occurs in other cat tumours. Other immunohistochemical staining results were unremarkable. 1999 Harcourt Publishers Ltd. PMID- 10542123 TI - Exercise-induced haemorrhagic lesions in the dorsocaudal extremities of the caudal lobes of the lungs of young thoroughbred horses. AB - The dorsocaudal extremities of the caudal lobes of the lungs of racehorses are vulnerable to exercise-induced pulmonary haemorrhage (EIPH). The morphology of the lungs at these sites was studied in 13 Thoroughbred horses aged 18 to 22 months. These animals, which had been performing low-intensity exercise on a track at maximum running speeds of approximately 5-8.5 metres/second (m/s), were withdrawn from the racehorse training programme for reasons of unsuitability. Lung lesions observed in the dorsocaudal lung extremities in 10 of the 13 horses were not found in the craniodorsal or cranioventral portions of the lungs. The lesions, which resembled those previously found in Thoroughbred racehorses aged 5 to 11 years with a history of EIPH, were of two main types, namely, multifocal bronchiolar distortion and alveolar epithelialization. EIPH lesions were found only in horses that had been trained at maximum speeds greater than approximately 7.0 m/s. It would seem, therefore, that exercise intensity is an important factor in the pathogenesis of EIPH and that running speeds greater than approximately 7.0 m/s may be sufficient to generate the pulmonary vascular pressures necessary to cause EIPH lesions in young Thoroughbreds. PMID- 10542122 TI - Lesions in cattle exposed to Mycobacterium bovis-inoculated calves. AB - Nine calves were housed for periods ranging from 24 to 117 days in close contact with cattle inoculated intranasally with Mycobacterium bovis. These "in-contact" calves were examined immunologically and bacteriologically during the period of exposure, and pathologically and immunocytochemically post mortem. Three became infected by day 14, as indicated by the detection of M. bovis in nasal mucus. In vitro interferon-gamma production and lymphocyte proliferation were detected after stimulation of peripheral blood with M. bovis antigens in the majority of in-contact animals by day 28; this provided support for the role of immunological mechanisms in pathogenesis. Tuberculous lesions were found in the submandibular and bronchomediastinal lymph nodes and in the lungs of the in-contact calves; in distribution and appearance the lesions resembled those observed in naturally occurring disease. The distribution of M. bovis antigen and the numbers of mycobacteria within pulmonary lesions are reported. 1999 Harcourt Publishers Ltd. PMID- 10542124 TI - In-situ hybridization for the detection of inflammatory cytokines (IL-1, TNF alpha and IL-6) in pigs naturally infected with Actinobacillus pleuropneumoniae. AB - The detection and distribution of interleukin-1 (IL-1), tumour necrosis factor alpha (TNF-alpha) and IL-6 were studied, by in-situ hybridization with a non radioactive digoxigenin-labelled probe, in formalin-fixed paraffin wax- embedded lung tissue from 10 pigs naturally infected with Actinobacillus pleuropneumoniae. A strong hybridization signal for IL-1, TNF-alpha and IL-6 was detected in "streaming" degenerate alveolar leucocytes (the so-called "oat cells") bordering zones of coagulative necrosis, and a less intense signal was seen in the dense zone of degenerate cells in granulation tissue surrounding the necrotic areas. IL 1 expression was also prominent in scattered endothelial cells bordering zones of coagulative necrosis. Simultaneous expression of all three cytokines was always associated with pleuropneumonic lung lesions. Expression of inflammatory cytokines was minimal in non-lesional lung tissue of the infected pigs and in normal lung from control pigs. The results suggest that these cytokines play a crucial role in mediating and regulating inflammation through cells of several types in A. pleuropneumoniae infection. 1999 Harcourt Publishers Ltd. PMID- 10542125 TI - Nuclear DNA fragmentation and immune reactivity in bovine spongiform encephalopathy. AB - To investigate whether apoptosis contributes to neuronal degeneration in bovine spongiform encephalopathy (BSE), morphological changes consistent with apoptosis were sought and in-situ end labelling (ISEL) was applied, in a series of 20 BSE cases and 10 age-matched normal control cattle. Apoptotic changes were not found in neurons but were occasionally seen in glial cells. Relatively few ISEL positive neurons were found, but many labelled nuclei were seen in glial cells in certain areas. None of the labelled cells showed morphological features of apoptosis. ISEL(+)cells occurred in areas of spongiform change and other areas of grey matter lacking spongiform change. Some association was found between degree of cellular DNA fragmentation and accumulation of abnormal prion protein (PrP(Sc)). Interestingly, small or moderate numbers of T lymphocytes, not present in the normal central nervous system (CNS), were detected in the CNS parenchyma in most BSE cases. There was a pronounced astrogliosis, but markers of macrophage or microglial activation were only slightly increased. The results indicate that nuclear DNA vulnerability is enhanced in certain neuroanatomical areas in BSE, but evidence that apoptosis plays a role in neuronal loss in BSE was very limited. 1999 Harcourt Publishers Ltd. PMID- 10542126 TI - Experimental infection of specific pathogen-free New Zealand White rabbits with five strains of amyxomatous myxoma virus. AB - Myxomatosis is a specific disease of the European rabbit (Oryctolagus cuniculus) due to a virus belonging to the genus Leporipoxvirus. Forty-seven years after its deliberate introduction into Europe, the clinical aspects and the epizootiology of myxomatosis have changed. Two forms (nodular and amyxomatous) of the disease have been identified to date. A comparative study was made of the clinical signs, pathogenesis and gross lesions observed in male specific pathogen-free New Zealand White rabbits inoculated with five strains of amyxomatous myxoma virus. All five strains induced the characteristic amyxomatous myxomatosis clinical syndrome with clinical signs that differed only in intensity. The varying clinical intensity, together with the results of virological examination question the virulence of at least three of the five strains. Genomic analysis confirmed that the five strains came from the Lausanne strain introduced in 1952 in France and not from an unnoticed introduction of a Californian strain of myxoma virus. No link was found between the amyxomatous myxoma virus strains and the SG33 vaccine strain. 1999 Harcourt Publishers Ltd. PMID- 10542128 TI - Volume contents, volume 120 PMID- 10542127 TI - Immunohistochemical study of the inflammatory infiltrate associated with equine squamous cell carcinoma. AB - The distribution of T (CD3), B (CD79) lymphocytes, immunoglobulin (IgG, IgM and IgA)-producing plasma cells, macrophages (lysozyme, Mac387) and MHC Class II antigen was analysed in the inflammatory infiltrate associated with 19 equine squamous cell carcinomas (SCCs) and six cases of precancerous lesions (actinic keratosis). The SCCs came from the penis (11 cases), conjunctiva (four), skin (two), nasal cavity (one) and oral cavity (one). Seven cases were well differentiated and 12 moderately differentiated. Nine cases showed no invasion of peritumoral deep tissues (locally invasive), whereas the remaining 10 cases were highly invasive. An abundant inflammatory infiltrate was associated with the majority of the SCCs and with lesions of actinic keratosis. This infiltrate was composed mainly of CD3(+)T lymphocytes, CD79(+)B cells and numerous IgG(+)plasma cells; IgM- and IgA-producing plasma cells were scarce and variable, respectively. Macrophages were usually numerous. Macrophages, lymphocytes, intra epithelial dendritic cells and fibroblasts expressed MHC Class II antigen. No significant correlation was found between the nature of the inflammatory infiltrate and the SCC histological grade or degree of invasion, suggesting that the local anti-tumour immune response failed to prevent tumour invasion or metastasis. MHC Class II was expressed by a variable number of neoplastic epithelial cells in four SCCs, all of which were only locally invasive. In addition, in areas where SCC cells expressed Class II antigen, numerous CD3(+)T lymphocytes were present and some of them were associated with degenerate tumour cells. These findings suggest that the expression of MHC Class II by neoplastic cells induces an improved local anti-tumour immune response. PMID- 10542129 TI - The role of the E-cadherin/catenin adhesion complex in the development and progression of cancer. AB - The E-cadherin/catenin protein complex regulates the functional integrity of epithelia by mediating specific intercellular adhesion, Defects in the transmembrane E-cadherin protein play an important role in several human cancer types. E-cadherin-inactivating mutations were mainly found in sporadic lobular breast carcinoma and in both familial and sporadic diffuse gastric carcinoma. Armadillo proteins such as beta-catenin and p120ctn are complexed to the cytoplasmic tail of E-cadherin, whereas the vinculin-related alphaE-catenin protein forms a link to the actin cytoskeleton. The latter shows inactivating deletions in various tumor cell lines. Apparently, both E-cadherin and alphaE catenin serve as tumor suppressor and invasion suppressor molecules. On the other hand, protein-stabilizing oncogenic mutations of beta-catenin were found at high frequency in particular human tumor types. Mutated beta-catenin protein is imported into the nucleus, and its binding to LEF/TCF transcription factors modulates transcription of intriguing target genes. Also p120ctn was recently found to arrive in the nucleus and to interact with a transcription factor. Furthermore, a wide variety of mechanisms have been described to regulate in a reversible way E-cadherin/catenin-mediated cell adhesion and differentiation. These phenomena appear to be crucial in human cancer development and progression. PMID- 10542130 TI - Light regulation of nitrate reductase gene expression in maize involves a G protein. AB - This paper reports three lines of evidence to demonstrate the presence of heterotrimeric G-proteins in maize and their involvement in the regulation of nitrate reductase gene expression by light: (1) Southern blot analysis of maize genomic DNA using a human Ha-ras cDNA probe revealed specific bands indicating the presence of G-protein (alpha subunit) gene(s) in maize. Northern blot analysis of maize total RNA using the same probe revealed that the putative Galpha gene(s) is transcriptionally active. (2) Western blots containing purified plasma membrane proteins from maize leaves showed specific binding of gamma [35S] labeled GTP in a red light-dependent manner, indicating the involvement of G proteins in mediating the light signal. The size of the putative Galpha gene product (approximately 45 kDa) indicates that it may be a heterotrimeric G protein. (3) Cholera toxin mimicked the effect of red light to enhance the transcript levels of nitrate reductase (NR), indicating that G-proteins may mediate light regulation of NR gene expression. PMID- 10542131 TI - Cloning and expression of a murine histone deacetylase 3 (mHdac3) cDNA and mapping to a region of conserved synteny between murine chromosome 18 and human chromosome 5. AB - Histone deacetylases (HDACs) are enzymes that play a pivotal role in transcription, differentiation, and cell cycle progression. We previously cloned human HDAC3 cDNA and showed that its transfection into THP-1 cells results in G2/M cell cycle accumulation. Using bioinformatic screening and PCR, we have now cloned the murine Hdac3 cDNA, which encodes a 428-amino-acid protein with near complete identity to its human ortholog. To establish a link to a potential disease locus, we performed PCR-based chromosomal mapping for the mHdac3 gene and chromosomal fluorescence in situ hybridization (FISH) for the human gene. mHdac3 localizes to chromosome 18 and human HDAC3 gene localizes to a syntenic region in chromosome 5 at band q31.3-q32 telomeric to the cytokine gene cluster. Transfection of mHdac3 into HeLa cells led to accumulation in G2/M. Our results suggest a cell cycle function for murine Hdac3, reflecting the complex regulatory roles of this gene family. PMID- 10542132 TI - Heterologous upregulation of nerve growth factor-TrkA receptors in PC12 cells by pituitary adenylate cyclase-activating polypeptide (PACAP). AB - The capacity for the neurotrophic factor PACAP38 to regulate expression of nerve growth factor (NGF)-trkA receptors in PC12 cells has been examined. Treatment of PC12 cells with 5 nM PACAP38 for 48 h elicited a 2.5-fold increase in 125I-NGF binding sites. FACS and Western analysis of trkA receptor protein indicate an abundance of receptors. The PACAP38-selective antagonist PACAP 6-38 blocked trkA receptor upregulation elicited by PACAP38. The expression of epidermal growth factor receptors was not affected by PACAP38 suggesting that upregulation of trkA represents a selective effect of this neurotrophic peptide. Similarly, expression of the pan-neurotrophin binding receptor p75 was not altered by PACAP38 treatment. In addition to effects on trkA observed in wild-type PC12 cells, PACAP38 stimulated an increase in the level of expressed human trkA receptors stably transfected into PC12 cells. PACAP38 provoked an increase in basal and NGF stimulated phosphorylation of trkA. Enhanced phosphorylation of trkA was detected as early as 6 h following addition of PACAP38 and was maximal at 48 h. Increased incorporation of phosphate occurs on both serine and tyrosine residues of trkA. These results suggest that PACAP38 is able to promote upregulation of trkA receptors, an event associated with elevated serine/tyrosine phosphorylation of trkA. PMID- 10542133 TI - Molecular characterization and genomic mapping of human IPM 200, a second member of a novel family of proteoglycans. AB - We report herein the characterization of the cDNA for a novel human chondroitin sulfate proteoglycan, designated IPM 200, and the chromosomal location of its gene, designated IMPG2. IPM 200 was isolated from the retinal interphotoreceptor matrix, a unique extracellular matrix that occupies the subretinal space between the apices of the retinal pigment epithelium and the neural retina. The cDNA contains an open reading frame of 3,726 bp that codes for a core protein with a deduced molecular weight of 138.5 kDa. The deduced IPM 200 core protein contains a putative transmembrane domain, two EGF-like repeats, numerous N- and O-linked glycosylation consensus sequences and one consensus sequence for glycosaminoglycan attachment. IMPG2 maps to human chromosome 3q12.2-12.3. Based on homologies within their amino acid sequences we propose that IPM 200 and a previously described human proteoglycan, IPM 150, form a new family of extracellular matrix glycoconjugates. PMID- 10542134 TI - cbl-b inhibits EGF-receptor-induced apoptosis by enhancing ubiquitination and degradation of activated receptors. AB - Studies in C. elegans and Drosophila melanogaster suggest that cbl proteins are inhibitors of epidermal growth factor receptor (EGFR) function. Here we describe that overexpression of cbl-b, a homologue of the c-cbl protooncogene, inhibits EGFR-induced apoptosis in MDA-MB-468 breast cancer cells. Overexpression of cbl-b results in a shortened duration of EGFR activation upon EGF stimulation. This is demonstrated by decreased amounts of phosphorylated EGFR as well as by inhibition of multiple downstream signaling pathways. The inhibition of signaling by cbl-b results from increased ubiquitination and degradation of the activated EGFR. The inhibitory effects of cbl-b overexpression on apoptosis and on EGFR signaling are reversed by blocking proteosomal degradation of the EGFR. These data demonstrate that the mechanism by which cbl-b inhibits EGFR-induced apoptosis is by activation-dependent degradation of the EGFR. They imply that this mechanism may be a general one whereby cbl proteins regulate intracellular signaling. PMID- 10542135 TI - Nuclear localization of hypoxia-inducible factor-2alpha in bovine arterial endothelial cells. AB - Hypoxia-inducible factor (HIF)-2alpha is a recently identified hypoxia-inducible transcription factor abundantly expressed in vascular endothelial cells. As well as HIF-1alpha, HIF-2alpha forms a heterodimeric complex with the aryl hydrocarbon receptor nuclear translocator and upregulates hypoxia-inducible genes such as vascular endothelial growth factor. We found in this study that using green fluorescent protein (GFP) fusion constructs, the subcellular localization of HIF 2alpha was different from that of HIF-1alpha in bovine arterial endothelial cells (BAEC). HIF-1alpha was localized in the cytoplasm under normoxic cells and translocated from the cytoplasm into the nucleus in response to hypoxic induction. In contrast, HIF-2alpha was clearly localized in the nucleus of BAEC even under normoxic conditions. The regulation of HIF-2alpha might differ from that of HIF-1alpha in BAEC. We further showed that nuclear localization of HIF 2alpha was inhibited by either deletion or a single amino acid substitution within the C-terminal end of the protein. The amino acid sequence surrounding Lys737 and Arg738 functions as a nuclear localization signal of HIF-2alpha. PMID- 10542136 TI - Cholera toxin induces tumor necrosis factor alpha production in human monocytes. AB - Cholera toxin covalently ADP-ribosylates the a subunit of Gs proteins. The modified Gsalpha activates adenylate cyclase and leads to a dramatic increase in intracellular cAMP. The effect of cholera toxin on the production of tumor necrosis factor (TNF-alpha), a critical mediator of toxicity for a number of bacterial and viral infections, has not been examined. Here we show that cholera toxin stimulated human monocytes to secrete TNF-alpha. The subunit A of cholera toxin alone also induced TNF-alpha production, suggesting that TNF-alpha production is mediated through ADP-ribosylation activity of the toxin. Inhibitors of ADP-ribosylation such as 3-aminobenzamide and niacinamide blocked TNF-alpha induction. However, cyclic AMP analogs and adenylate cyclase activator forskolin did not induce TNF-alpha production in monocytes, suggesting that TNF-alpha induction is independent of cAMP. Furthermore, cholera toxin-induced TNF-alpha production was suppressed by protein kinase C inhibitors H7 and sphingosine and by phospholipase C inhibitors U73122 and ET-18-OCH3, suggesting that PLC and PKC mediate TNF-alpha induction. Cholera toxin-mediated induction of TNF-alpha occurs at the transcription level as demonstrated by the time-dependent expression of TNF-alpha mRNA. These results raise the possibility that TNF-alpha may play an important role in cholera toxin-mediated toxicity and demonstrate that cholera toxin activates TNF-alpha production through PLC-dependent and cAMP-independent pathways. The probable mechanisms of signal transduction from cholera toxin to PLC in monocytes will be discussed. PMID- 10542137 TI - Development of human and rabbit vaginal smooth muscle cell cultures: effects of vasoactive agents on intracellular levels of cyclic nucleotides. AB - In this study, we subcultured and characterized human and rabbit vaginal smooth muscle cells and investigated the synthesis of second messenger cyclic nucleotides in response to vasodilators and determined the activity and kinetics of phosphodiesterase (PDE) type 5 (EC 3.1.4.35 3',5'-cyclic GMP phosphodiesterase). Cultured vaginal cells exhibited growth characteristics typical of smooth muscle cells and immunostained with antibodies against alpha smooth muscle actin. The cells retained functional prostaglandin E and beta adrenergic receptors as demonstrated by increased intracellular cAMP synthesis in response to PGE1, or isoproterenol. The response to these vasoactive substances was augmented with forskolin, suggesting stabilization of G-protein-activated adenylyl cyclases. Treatment with the nitric oxide donor, sodium nitroprusside, in the presence of sildenafil, a PDE type 5 inhibitor, enhanced intracellular cGMP synthesis and accumulation. Incubation of rabbit vaginal tissue with sildenafil, sodium nitroprusside, and PGE1 or forskolin produced a marked increase in intracellular cGMP. These observations were similar to findings with cultured cells and suggest that subcultured cells retain functional characteristics exhibited in intact tissue. The cells retained phosphodiesterase type 5 expression as shown by specific cGMP hydrolytic activity. Sildenafil and zaprinast inhibited cGMP hydrolysis competitively and bound with high affinity (Ki = 7 and 250 nM, respectively). These observations suggest that cultured human and rabbit vaginal smooth muscle cells retained their metabolic functional integrity and this experimental system should prove useful in investigating the pathway of nitric oxide and PDE type 5 inhibitors in modulating vaginal smooth muscle tone. PMID- 10542139 TI - The presenilins in Alzheimer's disease--proteolysis holds the key. AB - Alzheimer's disease (AD) research has shown that patients with an inherited form of the disease carry mutations in the presenilin proteins or the amyloid precursor protein (APP). These disease-linked mutations result in increased production of the longer form of amyloid-beta (the primary component of the amyloid deposits found in AD brains). However, it is not clear how the presenilins contribute to this increase. New findings now show that the presenilins affect APP processing through their effects on gamma-secretase, an enzyme that cleaves APP. Also, it is known that the presenilins are involved in the cleavage of the Notch receptor, hinting that they either directly regulate gamma-secretase activity or themselves are protease enzymes. These findings suggest that the presenilins may prove to be valuable molecular targets for the development of drugs to combat AD. PMID- 10542140 TI - Imaging magma transport during the 1997 seismic swarm off the izu peninsula, japan AB - The spatio-temporal evolution of a propagating magma-filled crack was estimated from inversion of Global Positioning System (GPS) data, tiltmeters, and leveling. The dike opened at a maximum rate of 50 millimeters per day and had a peak magma flux of 2 x 10(6) cubic meters per day. Although the spatial resolution was limited, slow upward propagation was resolved during the 9-day-long intrusion. In contrast, the earthquakes migrated rapidly upward during the first 12 hours of the swarm, and nearly all of the seismic energy was released in the first 2 days. Comparison of inversion results with accurate hypocenter locations will lead to improved understanding of magma transport through the brittle crust and of the causes of volcanic seismicity. PMID- 10542138 TI - High-cell-density phorbol ester and retinoic acid upregulate involucrin and downregulate suprabasal keratin 10 in autocrine cultures of human epidermal keratinocytes. AB - Autocrine growth of human epidermal keratinocytes is initiated in subconfluent cell cultures in the absence of exogenous growth factors, at low calcium concentration of the medium and with sufficient cell density. Culture confluence inhibits keratinocyte proliferation and upregulates expression of early, keratin 10 (K10), and late, involucrin, markers of differentiation. In this report, the phenotype of autocrine keratinocytes was studied at high cell density (postconfluence), specifically after treatment with 12-O-tetradecanoylphorbol 13 acetate (TPA), or all-trans retinoic acid (RA). At postconfluence, K10 is decreased but not involucrin. TPA upregulates involucrin expression, but not K10 in subconfluent keratinocytes. Treatment of confluent keratinocytes with RA downregulates K10, but upregulates involucrin. This in vitro culture model, unlike others, simulates for the first time the in vivo effects of RA, a member of the retinoid family which potently modulates keratinocyte differentiation and the expression of selected gene products. It thus can be developed to further examine epidermal differentiation. PMID- 10542141 TI - Abrupt climate change at the end of the last glacial period inferred from trapped air in polar Ice AB - The last glacial period was terminated by an abrupt warming event in the North Atlantic approximately 15,000 years before the present, and warming events of similar age have been reported from low latitudes. Understanding the mechanism of this termination requires that the precise relative timing of abrupt climate warming in the tropics versus the North Atlantic be known. Nitrogen and argon isotopes in trapped air in Greenland ice show that the Greenland Summit warmed 9 +/- 3 degrees C over a period of several decades, beginning 14,672 years ago. Atmospheric methane concentrations rose abruptly over a approximately 50-year period and began their increase 20 to 30 years after the onset of the abrupt Greenland warming. These data suggest that tropical climate became warmer or wetter (or both) approximately 20 to 80 years after the onset of Greenland warming, supporting a North Atlantic rather than a tropical trigger for the climate event. PMID- 10542142 TI - 16 degrees C rapid temperature variation in central greenland 70,000 years Ago AB - Variations in the (29)N(2)/(28)N(2) ratio of air bubbles trapped in polar ice cores and their relation to variations of the (18)O/(16)O of the ice allow past surface temperature variations and ice age-gas age differences to be determined. High-resolution measurements of (29)N(2)/(28)N(2) in Dansgaard-Oeschger event 19 (around 70,000 years before the present) in ice from Central Greenland show that at the beginning of the event, the ice age-gas age difference was 1090 +/- 100 years. With the use of a combined firn densification, temperature, and gas diffusion model, the delta(18)O(ice)-temperature coefficient alpha was determined to be 0. 42 +/- 0.05 per mil per kelvin. This coefficient implies a mean surface temperature change of 16.0 kelvin (between 14.3 and 18.1 kelvin), which differs substantially from values derived from borehole temperatures and modern spatial delta(18)O(ice)-surface temperature correlations. PMID- 10542143 TI - Seismic consequences of warm versus cool subduction metamorphism: examples from southwest and northeast japan AB - Warm and cool subduction zones exhibit differences in seismicity, seismic structure, and arc magmatism, which reflect differences in metamorphic reactions occurring in subducting oceanic crust. In southwest Japan, arc volcanism is sparse and intraslab earthquakes extend to 65 kilometers depth; in northeast Japan, arc volcanism is more common and intraslab earthquakes reach 200 kilometers depth. Thermal-petrologic models predict that oceanic crust subducting beneath southwest Japan is 300 degrees to 500 degrees C warmer than beneath northeast Japan, resulting in shallower eclogite transformation and slab dehydration reactions, and possible slab melting. PMID- 10542144 TI - In situ observation of the formation of 10 A phase from talc + H(2)O at mantle pressures and temperatures AB - Development of an x-ray-transparent capsule that is suitable for water-saturated experiments has allowed the high-pressure and high-temperature synthesis of 10 angstrom phase from talc + H(2)O to be observed in situ in a multianvil apparatus with synchrotron radiation. The reaction talc + H(2)O = 10 angstrom phase was observed within 20 minutes at 6 gigapascals and 500 degrees C. The higher pressure vapor-absent reaction talc = 10 angstrom phase + enstatite + coesite was also observed. The 10 angstrom phase is therefore a good candidate for transporting water into Earth's mantle at subduction zones. The in situ technique may allow the hydration of other high-pressure mantle phases to be observed under dynamic conditions. PMID- 10542145 TI - Field-effect flow control for microfabricated fluidic networks AB - The magnitude and direction of the electro-osmotic flow (EOF) inside a microfabricated fluid channel can be controlled by a perpendicular electric field of 1.5 megavolts per centimeter generated by a voltage of only 50 volts. A microdevice called a "flowFET," with functionality comparable to that of a field effect transistor (FET) in microelectronics, has been realized. Two flowFETs integrated with a channel junction have been used to generate opposite flows inside a single EOF-pumped channel, thus illustrating the potential of the flowFET as a controlling and switching element in microfluidic networks. PMID- 10542146 TI - Organic-inorganic hybrid materials as semiconducting channels in thin-film field effect transistors AB - Organic-inorganic hybrid materials promise both the superior carrier mobility of inorganic semiconductors and the processability of organic materials. A thin-film field-effect transistor having an organic-inorganic hybrid material as the semiconducting channel was demonstrated. Hybrids based on the perovskite structure crystallize from solution to form oriented molecular-scale composites of alternating organic and inorganic sheets. Spin-coated thin films of the semiconducting perovskite (C(6)H(5)C(2)H(4)NH(3))(2)SnI(4) form the conducting channel, with field-effect mobilities of 0.6 square centimeters per volt-second and current modulation greater than 10(4). Molecular engineering of the organic and inorganic components of the hybrids is expected to further improve device performance for low-cost thin-film transistors. PMID- 10542147 TI - The root of angiosperm phylogeny inferred from duplicate phytochrome genes. AB - An analysis of duplicate phytochrome genes (PHYA and PHYC) is used to root the angiosperms, thereby avoiding the inclusion of highly diverged outgroup sequences. The results unambiguously place the root near Amborella (one species, New Caledonia) and resolve water lilies (Nymphaeales, approximately 70 species, cosmopolitan), followed by Austrobaileya (one species, Australia), as early branches. These findings bear directly on the interpretation of morphological evolution and diversification within angiosperms. PMID- 10542148 TI - A species of small antisense RNA in posttranscriptional gene silencing in plants. AB - Posttranscriptional gene silencing (PTGS) is a nucleotide sequence-specific defense mechanism that can target both cellular and viral mRNAs. Here, three types of transgene-induced PTGS and one example of virus-induced PTGS were analyzed in plants. In each case, antisense RNA complementary to the targeted mRNA was detected. These RNA molecules were of a uniform length, estimated at 25 nucleotides, and their accumulation required either transgene sense transcription or RNA virus replication. Thus, the 25-nucleotide antisense RNA is likely synthesized from an RNA template and may represent the specificity determinant of PTGS. PMID- 10542149 TI - TCR-Mediated internalization of peptide-MHC complexes acquired by T cells. AB - Peptide-major histocompatibility complex protein complexes (pMHCs) on antigen presenting cells (APCs) are central to T cell activation. Within minutes of peptide-specific T cells interacting with APCs, pMHCs on APCs formed clusters at the site of T cell contact. Thereafter, these clusters were acquired by T cells and internalized through T cell receptor-mediated endocytosis. During this process, T cells became sensitive to peptide-specific lysis by neighboring T cells (fratricide). This form of immunoregulation could explain the "exhaustion" of T cell responses that is induced by high viral loads and may serve to down regulate immune responses. PMID- 10542150 TI - Epigenetic inheritance of active chromatin after removal of the main transactivator. AB - The Drosophila Polycomb and trithorax group proteins act through chromosomal elements such as Fab-7 to maintain repressed or active gene expression, respectively. A Fab-7 element is switched from a silenced to a mitotically heritable active state by an embryonic pulse of transcription. Here, histone H4 hyperacetylation was found to be associated with Fab-7 after activation, suggesting that H4 hyperacetylation may be a heritable epigenetic tag of the activated element. Activated Fab-7 enables transcription of a gene even after withdrawal of the primary transcription factor. This feature may allow epigenetic maintenance of active states of developmental genes after decay of their early embryonic regulators. PMID- 10542151 TI - A direct estimate of the human alphabeta T cell receptor diversity. AB - Generation and maintenance of an effective repertoire of T cell antigen receptors are essential to the immune system, yet the number of distinct T cell receptors (TCRs) expressed by the estimated 10(12) T cells in the human body is not known. In this study, TCR gene amplification and sequencing showed that there are about 10(6) different beta chains in the blood, each pairing, on the average, with at least 25 different alpha chains. In the memory subset, the diversity decreased to 1 x 10(5) to 2 x 10(5) different beta chains, each pairing with only a single alpha chain. Thus, the naive repertoire is highly diverse, whereas the memory compartment, here one-third of the T cell population, contributes less than 1 percent of the total diversity. PMID- 10542152 TI - Arabidopsis NPH3: A NPH1 photoreceptor-interacting protein essential for phototropism. AB - Phototropism of Arabidopsis thaliana seedlings in response to a blue light source is initiated by nonphototropic hypocotyl 1 (NPH1), a light-activated serine threonine protein kinase. Mutations in three loci [NPH2, root phototropism 2 (RPT2), and NPH3] disrupt early signaling occurring downstream of the NPH1 photoreceptor. The NPH3 gene, now cloned, encodes a NPH1-interacting protein. NPH3 is a member of a large protein family, apparently specific to higher plants, and may function as an adapter or scaffold protein to bring together the enzymatic components of a NPH1-activated phosphorelay. PMID- 10542153 TI - Four evolutionary strata on the human X chromosome. AB - Human sex chromosomes evolved from autosomes. Nineteen ancestral autosomal genes persist as differentiated homologs on the X and Y chromosomes. The ages of individual X-Y gene pairs (measured by nucleotide divergence) and the locations of their X members on the X chromosome were found to be highly correlated. Age decreased in stepwise fashion from the distal long arm to the distal short arm in at least four "evolutionary strata." Human sex chromosome evolution was probably punctuated by at least four events, each suppressing X-Y recombination in one stratum, without disturbing gene order on the X chromosome. The first event, which marked the beginnings of X-Y differentiation, occurred about 240 to 320 million years ago, shortly after divergence of the mammalian and avian lineages. PMID- 10542154 TI - Opposite patterns of synchrony in sympatric disease metapopulations. AB - Measles epidemics in UK cities, which were regular and highly synchronous before vaccination, are known to have become irregular and spatially uncorrelated in the vaccine era. Whooping cough shows the reverse pattern, namely a shift from spatial incoherence and irregularity before vaccination to regular, synchronous epidemics afterward. Models show that these patterns can arise from disease specific responses to dynamical noise. This analysis has implications for vaccination strategies and illustrates the power of comparative dynamical studies of sympatric metapopulations. PMID- 10542155 TI - Small molecule inhibitor of mitotic spindle bipolarity identified in a phenotype based screen. AB - Small molecules that perturb specific protein functions are valuable tools for dissecting complex processes in mammalian cells. A combination of two phenotype based screens, one based on a specific posttranslational modification, the other visualizing microtubules and chromatin, was used to identify compounds that affect mitosis. One compound, here named monastrol, arrested mammalian cells in mitosis with monopolar spindles. In vitro, monastrol specifically inhibited the motility of the mitotic kinesin Eg5, a motor protein required for spindle bipolarity. All previously known small molecules that specifically affect the mitotic machinery target tubulin. Monastrol will therefore be a particularly useful tool for studying mitotic mechanisms. PMID- 10542156 TI - Prokaryotic nitrate reduction: molecular properties and functional distinction among bacterial nitrate reductases. PMID- 10542157 TI - Regulated antisense RNA eliminates alpha-toxin virulence in Staphylococcus aureus infection. AB - The ability to selectively disrupt gene function remains a critical element in elucidating information regarding gene essentiality for bacterial growth and/or pathogenesis. In this study, we adapted a tet regulatory expression system for use in Staphylococcus aureus, with the goal of downregulating gene expression via induction of antisense RNA. We demonstrate that this system exhibits a 50- to 100 fold dose-dependent level of induction in bacterial cells grown in culture (i.e., in vitro) and also functions in mice (i.e., in vivo) following oral administration of inducer. To determine whether induced antisense RNA could interfere with chromosomally derived gene expression, we cloned a fragment of the S. aureus alpha-toxin gene (hla) in antisense orientation downstream of the tet promoter system and introduced the construct into S. aureus. Induced antisense hla RNA downregulated chromosomally derived hla gene expression in vitro approximately 14-fold. Similarly, induction of hla antisense RNA in vivo dramatically reduced alpha-toxin expression in two different murine models of S. aureus infection. Most importantly, this reduction completely eliminated the lethality of the infection. These results indicate that the tet regulatory system functions efficiently in S. aureus and induced antisense RNA can effectively downregulate chromosomal gene expression both in vitro and in vivo. PMID- 10542158 TI - Functional interactions of a homolog of proliferating cell nuclear antigen with DNA polymerases in Archaea. AB - Proliferating cell nuclear antigen (PCNA) is an essential component of the DNA replication and repair machinery in the domain Eucarya. We cloned the gene encoding a PCNA homolog (PfuPCNA) from an euryarchaeote, Pyrococcus furiosus, expressed it in Escherichia coli, and characterized the biochemical properties of the gene product. The protein PfuPCNA stimulated the in vitro primer extension abilities of polymerase (Pol) I and Pol II, which are the two DNA polymerases identified in this organism to date. An immunological experiment showed that PfuPCNA interacts with both Pol I and Pol II. Pol I is a single polypeptide with a sequence similar to that of family B (alpha-like) DNA polymerases, while Pol II is a heterodimer. PfuPCNA interacted with DP2, the catalytic subunit of the heterodimeric complex. These results strongly support the idea that the PCNA homolog works as a sliding clamp of DNA polymerases in P. furiosus, and the basic mechanism for the processive DNA synthesis is conserved in the domains Bacteria, Eucarya, and Archaea. The stimulatory effect of PfuPCNA on the DNA synthesis was observed by using a circular DNA template without the clamp loader (replication factor C [RFC]) in both Pol I and Pol II reactions in contrast to the case of eukaryotic organisms, which are known to require the RFC to open the ring structure of PCNA prior to loading onto a circular DNA. Because RFC homologs have been found in the archaeal genomes, they may permit more efficient stimulation of DNA synthesis by archaeal DNA polymerases in the presence of PCNA. This is the first stage in elucidating the archaeal DNA replication mechanism. PMID- 10542159 TI - Contribution of the cell wall component teichuronopeptide to pH homeostasis and alkaliphily in the alkaliphile Bacillus lentus C-125. AB - A teichuronopeptide (TUP) is one of major structural components of the cell wall of the facultative alkaliphilic strain Bacillus lentus C-125. A mutant defective in TUP synthesis grows slowly at alkaline pH. An upper limit of pH for growth of the mutant was 10.4, while that of the parental strain C-125 was 10.8. Gene tupA, directing synthesis of TUP, was cloned from C-125 chromosomal DNA. The primary translation product of this gene is likely a cytoplasmic protein (57. 3 kDa) consisting of 489 amino acid residues. Introduction of the tupA gene into the TUP defective mutant complemented the mutation responsible for the pleiotropic phenotypes of the mutant, leading to simultaneous disappearance of the defect in TUP synthesis, the diminished ability for cytoplasmic pH homeostasis, and the low tolerance for alkaline conditions. These results demonstrate that the acidic polymer TUP in the cell wall plays a role in pH homeostasis in this alkaliphile. PMID- 10542161 TI - Characterization of a Hank's type serine/threonine kinase and serine/threonine phosphoprotein phosphatase in Pseudomonas aeruginosa. AB - Pseudomonas aeruginosa is an opportunistic pathogen that causes infections in eye, urinary tract, burn, and immunocompromised patients. We have cloned and characterized a serine/threonine (Ser/Thr) kinase and its cognate phosphoprotein phosphatase. By using oligonucleotides from the conserved regions of Ser/Thr kinases of mycobacteria, an 800-bp probe was used to screen P. aeruginosa PAO1 genomic library. A 20-kb cosmid clone was isolated, from which a 4.5-kb DNA with two open reading frames (ORFs) were subcloned. ORF1 was shown to encode Ser/Thr phosphatase (Stp1), which belongs to the PP2C family of phosphatases. Overlapping with the stp1 ORF, an ORF encoding Hank's type Ser/Thr kinase was identified. Both ORFs were cloned in pGEX-4T1 and expressed in Escherichia coli. The overexpressed proteins were purified by glutathione-Sepharose 4B affinity chromatography and were biochemically characterized. The Stk1 kinase is 39 kDa and undergoes autophosphorylation and can phosphorylate eukaryotic histone H1. A site-directed Stk1 (K86A) mutant was shown to be incapable of autophosphorylation. A two-dimensional phosphoamino acid analysis of Stk1 revealed strong phosphorylation at a threonine residue and weak phosphorylation at a serine residue. The Stp1 phosphatase is 27 kDa and is an Mn(2+)-, but not a Ca(2+)- or a Mg(2+)-, dependent Ser/Thr phosphatase. Its activity is inhibited by EDTA and NaF, but not by okadaic acid, and is similar to that of PP2C phosphatase. PMID- 10542160 TI - On the origin of branches in Escherichia coli. AB - Some Escherichia coli strains with impaired cell division form branched cells at high frequencies during certain growth conditions. Here, we show that neither FtsI nor FtsZ activity is required for the development of branches. Buds did not form at specific positions along the cell surface during high-branching conditions. Antibiotics affecting cell wall synthesis had a positive effect on branch formation in the case of ampicillin, cephalexin, and penicillin G, whereas mecillinam and D-cycloserine had no substantial effect. Altering the cell morphology by nutritional shifts showed that changes in morphology preceded branching, indicating that the cell's physiological state rather than specific medium components induced branching. Finally, there was no increased probability for bud formation in the daughters of a cell with a bud or branch, showing that bud formation is a random event. We suggest that branch formation is caused by abnormalities in cell wall elongation rather than by aberrant cell division events. PMID- 10542162 TI - Molecular evolution of the pathogenicity island of enterotoxigenic Bacteroides fragilis strains. AB - Enterotoxigenic Bacteroides fragilis (ETBF) strains, which produce a 20-kDa zinc metalloprotease toxin (BFT), have been associated with diarrheal disease in animals and young children. Studying a collection of ETBF and nontoxigenic B. fragilis (NTBF) strains, we found that bft and a second metalloprotease gene (mpII) are contained in an approximately 6-kb pathogenicity island (termed B. fragilis pathogenicity island or BfPAI) which is present exclusively in all 113 ETBF strains tested (pattern I). Of 191 NTBF strains, 100 (52%) lack both the BfPAI and at least a 12-kb region flanking BfPAI (pattern II), and 82 of 191 NTBF strains (43%) lack the BfPAI but contain the flanking region (pattern III). The nucleotide sequence flanking the left end of the BfPAI revealed a region with the same organization as the mobilization region of the 5-nitroimidazole resistance plasmid pIP417 and the clindamycin resistance plasmid pBFTM10, that is, two mobilization genes (bfmA and bfmB) organized in one operon and a putative origin of transfer (oriT) located in a small, compact region. The region flanking the right end of the BfPAI contains a gene (bfmC) whose predicted protein shares significant identity to the TraD mobilization proteins encoded by plasmids F and R100 from Escherichia coli. Nucleotide sequence analysis of one NTBF pattern III strain (strain I-1345) revealed that bfmB and bfmC are adjacent to each other and separated by a 16-bp GC-rich sequence. Comparison of this sequence with the appropriate sequence of ETBF strain 86-5443-2-2 showed that in this ETBF strain the 16-bp sequence is replaced by the BfPAI. This result defined the BfPAI as being 6,036 bp in length and its precise integration site as being between the bfmB and bfmC stop codons. The G+C content of the BfPAI (35%) and the flanking DNA (47 to 50%) differ greatly from that reported for the B. fragilis chromosome (42%), suggesting that the BfPAI and its flanking region are two distinct genetic elements originating from very different organisms. ETBF strains may have evolved by horizontal transfer of these two genetic elements into a pattern II NTBF strain. PMID- 10542163 TI - The Oenococcus oeni clpX homologue is a heat shock gene preferentially expressed in exponential growth phase. AB - Using degenerated primers from conserved regions of previously studied clpX gene products, we cloned the clpX gene of the malolactic bacterium Oenococcus oeni. The clpX gene was sequenced, and the deduced protein of 413 amino acids (predicted molecular mass of 45,650 Da) was highly similar to previously analyzed clpX gene products from other organisms. An open reading frame located upstream of the clpX gene was identified as the tig gene by similarity of its predicted product to other bacterial trigger factors. ClpX was purified by using a maltose binding protein fusion system and was shown to possess an ATPase activity. Northern analyses indicated the presence of two independent 1.6-kb monocistronic clpX and tig mRNAs and also showed an increase in clpX mRNA amount after a temperature shift from 30 to 42 degrees C. The clpX transcript is abundant in the early exponential growth phase and progressively declines to undetectable levels in the stationary phase. Thus, unlike hsp18, the gene encoding one of the major small heat shock proteins of Oenococcus oeni, clpX expression is related to the exponential growth phase and requires de novo protein synthesis. Primer extension analysis identified the 5' end of clpX mRNA which is located 408 nucleotides upstream of a putative AUA start codon. The putative transcription start site allowed identification of a predicted promoter sequence with a high similarity to the consensus sequence found in the housekeeping gene promoter of gram-positive bacteria as well as Escherichia coli. PMID- 10542164 TI - Purification, characterization, and gene analysis of a chitosanase (ChoA) from Matsuebacter chitosanotabidus 3001. AB - The extracellular chitosanase (34,000 M(r)) produced by a novel gram-negative bacterium Matsuebacter chitosanotabidus 3001 was purified. The optimal pH of this chitosanase was 4.0, and the optimal temperature was between 30 and 40 degrees C. The purified chitosanase was most active on 90% deacetylated colloidal chitosan and glycol chitosan, both of which were hydrolyzed in an endosplitting manner, but this did not hydrolyze chitin, cellulose, or their derivatives. Among potential inhibitors, the purified chitosanase was only inhibited by Ag(+). Internal amino acid sequences of the purified chitosanase were obtained. A PCR fragment corresponding to one of these amino acid sequences was then used to screen a genomic library for the entire choA gene encoding chitosanase. Sequencing of the choA gene revealed an open reading frame encoding a 391-amino acid protein. The N-terminal amino acid sequence had an excretion signal, but the sequence did not show any significant homology to other proteins, including known chitosanases. The 80-amino-acid excretion signal of ChoA fused to green fluorescent protein was functional in Escherichia coli. Taken together, these results suggest that we have identified a novel, previously unreported chitosanase. PMID- 10542165 TI - Activation of the cryptic aac(6')-Iy aminoglycoside resistance gene of Salmonella by a chromosomal deletion generating a transcriptional fusion. AB - Salmonella enterica subsp. enterica serotype Enteritidis BM4361 and BM4362 were isolated from the same patient. BM4361 was susceptible to aminoglycosides, whereas BM4362 was resistant to tobramycin owing to synthesis of a 6'-N acetyltransferase type I [AAC(6')-I]. Comparative analysis of nucleotide sequences, pulsed-field gel electrophoresis patterns, and Southern hybridizations indicated that the chromosomal aac(6')-Iy genes for the enzyme in both strains were identical and that BM4362 derived from BM4361 following a ca. 60-kb deletion that occurred 1.5 kb upstream from the resistance gene. Northern hybridizations showed that aac(6')-Iy was silent in BM4361 and highly expressed in BM4362 due to a transcriptional fusion. Primer extension mapping identified the transcriptional start site for aac(6')-Iy in BM4362: 5 bp downstream from the promoter of the nmpC gene. Study of the distribution of aac(6')-Iy by PCR and Southern hybridization with a specific probe indicated that the gene, although not found in S. enterica subsp. arizonae, was specific for Salmonella. In this bacterial genus, aac(6')-Iy was located downstream from a cluster of seven open reading frames analogous to an Escherichia coli locus that encodes enzymes putatively involved in carbohydrate transport or metabolism. This genomic environment suggests a role in the catabolism of a specific sugar for AAC(6')-Iy in Salmonella. PMID- 10542166 TI - Acetate metabolism in a pta mutant of Escherichia coli W3110: importance of maintaining acetyl coenzyme A flux for growth and survival. AB - In order to study the physiological role of acetate metabolism in Escherichia coli, the growth characteristics of an E. coli W3100 pta mutant defective in phosphotransacetylase, the first enzyme of the acetate pathway, were investigated. The pta mutant grown on glucose minimal medium excreted unusual by products such as pyruvate, D-lactate, and L-glutamate instead of acetate. In an analysis of the sequential consumption of amino acids by the pta mutant growing in tryptone broth (TB), a brief lag between the consumption of amino acids normally consumed was observed, but no such lag occurred for the wild-type strain. The pta mutant was found to grow slowly on glucose, TB, or pyruvate, but it grew normally on glycerol or succinate. The defective growth and starvation survival of the pta mutant were restored by the introduction of poly-beta hydroxybutyrate (PHB) synthesis genes (phbCAB) from Alcaligenes eutrophus, indicating that the growth defect of the pta mutant was due to a perturbation of acetyl coenzyme A (CoA) flux. By the stoichiometric analysis of the metabolic fluxes of the central metabolism, it was found that the amount of pyruvate generated from glucose transport by the phosphoenolpyruvate-dependent phosphotransferase system (PTS) exceeded the required amount of precursor metabolites downstream of pyruvate for biomass synthesis. These results suggest that E. coli excretes acetate due to the pyruvate flux from PTS and that any method which alleviates the oversupply of acetyl CoA would restore normal growth to the pta mutant. PMID- 10542167 TI - The hetC gene is a direct target of the NtcA transcriptional regulator in cyanobacterial heterocyst development. AB - The heterocyst is the site of nitrogen fixation in aerobically grown cultures of some filamentous cyanobacteria. Heterocyst development in Anabaena sp. strain PCC 7120 is dependent on the global nitrogen regulator NtcA and requires, among others, the products of the hetR and hetC genes. Expression of hetC, tested by RNA- DNA hybridization, was impaired in an ntcA mutant. A nitrogen-regulated, NtcA-dependent putative transcription start point was localized at nucleotide 571 with respect to the hetC translational start. Sequences upstream from this transcription start point exhibit the structure of the canonical cyanobacterial promoter activated by NtcA, and purified NtcA protein specifically bound to a DNA fragment containing this promoter. Activation of expression of hetC during heterocyst development appears thus to be directly operated by NtcA. NtcA mediated activation of hetR expression was not impaired in a hetC mutant, indicating that HetC is not an NtcA-dependent element required for hetR induction. PMID- 10542168 TI - Exponential-phase glycogen recycling is essential for growth of Mycobacterium smegmatis. AB - Bacterial glycogen is a polyglucose storage compound that is thought to prolong viability during stationary phase. However, a specific role for glycogen has not been determined. We have characterized SMEG53, a temperature-sensitive mutant of Mycobacterium smegmatis that contains a mutation in glgE, encoding a putative glucanase. This mutation causes exponentially growing SMEG53 cells to stop growing at 42 degrees C in response to high levels of glycogen accumulation. The mutation in glgE is also associated with an altered growth rate and colony morphology at permissive temperatures; the severity of these phenotypes correlates with the amount of glycogen accumulated by the mutant. Suppression of the temperature-sensitive phenotype, via a decrease in glycogen accumulation, is mediated by growth in certain media or multicopy expression of garA. The function of GarA is unknown, but the presence of a forkhead-associated domain suggests that this protein is a member of a serine-threonine kinase signal transduction pathway. Our results suggest that in M. smegmatis glycogen is continuously synthesized and then degraded by GlgE throughout exponential growth. In turn, this constant recycling of glycogen controls the downstream availability of carbon and energy. Thus, in addition to its conventional storage role, glycogen may also serve as a carbon capacitor for glycolysis during the exponential growth of M. smegmatis. PMID- 10542169 TI - Metabolic flux ratio analysis of genetic and environmental modulations of Escherichia coli central carbon metabolism. AB - The response of Escherichia coli central carbon metabolism to genetic and environmental manipulation has been studied by use of a recently developed methodology for metabolic flux ratio (METAFoR) analysis; this methodology can also directly reveal active metabolic pathways. Generation of fluxome data arrays by use of the METAFoR approach is based on two-dimensional (13)C-(1)H correlation nuclear magnetic resonance spectroscopy with fractionally labeled biomass and, in contrast to metabolic flux analysis, does not require measurements of extracellular substrate and metabolite concentrations. METAFoR analyses of E. coli strains that moderately overexpress phosphofructokinase, pyruvate kinase, pyruvate decarboxylase, or alcohol dehydrogenase revealed that only a few flux ratios change in concert with the overexpression of these enzymes. Disruption of both pyruvate kinase isoenzymes resulted in altered flux ratios for reactions connecting the phosphoenolpyruvate (PEP) and pyruvate pools but did not significantly alter central metabolism. These data indicate remarkable robustness and rigidity in central carbon metabolism in the presence of genetic variation. More significant physiological changes and flux ratio differences were seen in response to altered environmental conditions. For example, in ammonia-limited chemostat cultures, compared to glucose-limited chemostat cultures, a reduced fraction of PEP molecules was derived through at least one transketolase reaction, and there was a higher relative contribution of anaplerotic PEP carboxylation than of the tricarboxylic acid (TCA) cycle for oxaloacetate synthesis. These two parameters also showed significant variation between aerobic and anaerobic batch cultures. Finally, two reactions catalyzed by PEP carboxykinase and malic enzyme were identified by METAFoR analysis; these had previously been considered absent in E. coli cells grown in glucose-containing media. Backward flux from the TCA cycle to glycolysis, as indicated by significant activity of PEP carboxykinase, was found only in glucose-limited chemostat culture, demonstrating that control of this futile cycle activity is relaxed under severe glucose limitation. PMID- 10542170 TI - Stable packaging of phage PRD1 DNA requires adsorption protein P2, which binds to the IncP plasmid-encoded conjugative transfer complex. AB - The double-stranded DNA bacteriophage PRD1 uses an IncP plasmid-encoded conjugal transfer complex as a receptor. Plasmid functions in the PRD1 life cycle are restricted to phage adsorption and DNA entry. A single phage structural protein, P2, located at the fivefold capsid vertices, is responsible for PRD1 attachment to its host. The purified recombinant adsorption protein was judged to be monomeric by gel filtration, rate zonal centrifugation, analytical ultracentrifugation, and chemical cross-linking. It binds to its receptor with an apparent K(d) of 0.20 nM, and this binding prevents phage adsorption. P2 deficient particles are unstable and spontaneously release the DNA with concomitant formation of the tail-like structure originating from the phage membrane. We envisage the DNA to be packaged through one vertex, but the presence of P2 on the other vertices suggests a mechanism whereby the injection vertex is determined by P2 binding to the receptor. PMID- 10542172 TI - Purification and characterization of (per)chlorate reductase from the chlorate respiring strain GR-1. AB - Strain GR-1 is one of several recently isolated bacterial species that are able to respire by using chlorate or perchlorate as the terminal electron acceptor. The organism performs a complete reduction of chlorate or perchlorate to chloride and oxygen, with the intermediate formation of chlorite. This study describes the purification and characterization of the key enzyme of the reductive pathway, the chlorate and perchlorate reductase. A single enzyme was found to catalyze both the chlorate- and perchlorate-reducing activity. The oxygen-sensitive enzyme was located in the periplasm and had an apparent molecular mass of 420 kDa, with subunits of 95 and 40 kDa in an alpha(3)beta(3) composition. Metal analysis showed the presence of 11 mol of iron, 1 mol of molybdenum, and 1 mol of selenium per mol of heterodimer. In accordance, quantitative electron paramagnetic resonance spectroscopy showed the presence of one [3Fe-4S] cluster and two [4Fe 4S] clusters. Furthermore, two different signals were ascribed to Mo(V). The K(m) values for perchlorate and chlorate were 27 and <5 microM, respectively. Besides perchlorate and chlorate, nitrate, iodate, and bromate were also reduced at considerable rates. The resemblance of the enzyme to nitrate reductases, formate dehydrogenases, and selenate reductase is discussed. PMID- 10542171 TI - Transcriptional activation of the chlorocatechol degradative genes of Ralstonia eutropha NH9. AB - Ralstonia eutropha (formerly Alcaligenes eutrophus) NH9 degrades 3-chlorobenzoate via the modified ortho-cleavage pathway. A ca. 5.7-kb six-gene cluster is responsible for chlorocatechol degradation: the cbnABCD operon encoding the degradative enzymes (including orfX of unknown function) and the divergently transcribed cbnR gene encoding the LysR-type transcriptional regulator of the cbn operon. The cbnRAB orfXCD gene cluster is nearly identical to the chlorocatechol genes (tcbRCD orfXEF) of the 1,2, 4-trichlorobenzene-degrading bacterium Pseudomonas sp. strain P51. Transcriptional fusion studies demonstrated that cbnR regulates the expression of cbnABCD positively in the presence of either 3 chlorobenzoate or benzoate, which are catabolized via 3-chlorocatechol and catechol, respectively. In vitro transcription assays confirmed that 2-chloro cis,cis-muconate (2-CM) and cis, cis-muconate (CCM), intermediate products from 3 chlorocatechol and catechol, respectively, were inducers of this operon. This inducer-recognizing specificity is different from those of the homologous catechol (catBCA) and chlorocatechol (clcABD) operons of Pseudomonas putida, in which only the intermediates of the regulated pathway, CCM for catBCA and 2-CM for clcABD, act as significant inducers. Specific binding of CbnR protein to the cbnA promoter region was demonstrated by gel shift and DNase I footprinting analysis. In the absence of inducer, a region of ca. 60 bp from position -20 to position -80 upstream of the cbnA transcriptional start point was protected from DNase I cleavage by CbnR, with a region of hypersensitivity to DNase I cleavage clustered at position -50. Circular permutation gel shift assays demonstrated that CbnR bent the cbnA promoter region to an angle of 78 degrees and that this angle was relaxed to 54 degrees upon the addition of inducer. While a similar relaxation of bending angles upon the addition of inducer molecules observed with the catBCA and clcABD promoters may indicate a conserved transcriptional activation mechanism of ortho-cleavage pathway genes, CbnR is unique in having a different specificity of inducer recognition and the extended footprint as opposed to the restricted footprint of CatR without CCM. PMID- 10542173 TI - Cloning and sequencing of a novel meta-cleavage dioxygenase gene whose product is involved in degradation of gamma-hexachlorocyclohexane in Sphingomonas paucimobilis. AB - Sphingomonas (formerly Pseudomonas) paucimobilis UT26 utilizes gamma hexachlorocyclohexane (gamma-HCH), a halogenated organic insecticide, as a sole source of carbon and energy. In a previous study, we showed that gamma-HCH is degraded to chlorohydroquinone (CHQ) and then to hydroquinone (HQ), although the rate of reaction from CHQ to HQ was slow (K. Miyauchi, S. K. Suh, Y. Nagata, and M. Takagi, J. Bacteriol. 180:1354-1359, 1998). In this study, we cloned and characterized a gene, designated linE, which is located upstream of linD and is directly involved in the degradation of CHQ. The LinE protein consists of 321 amino acids, and all of the amino acids which are reported to be essential for the activity of meta-cleavage dioxygenases are conserved in LinE. Escherichia coli overproducing LinE could convert both CHQ and HQ, producing gamma hydroxymuconic semialdehyde and maleylacetate, respectively, with consumption of O(2) but could not convert catechol, which is one of the major substrates for meta-cleavage dioxygenases. LinE seems to be resistant to the acylchloride, which is the ring cleavage product of CHQ and which seems to react with water to be converted to maleylacetate. These results indicated that LinE is a novel type of meta-cleavage dioxygenase, designated (chloro)hydroquinone 1, 2-dioxygenase, which cleaves aromatic rings with two hydroxyl groups at para positions preferably. This study represents a direct demonstration of a new type of ring cleavage pathway for aromatic compounds, the hydroquinone pathway. PMID- 10542174 TI - A novel cellulosomal scaffoldin from Acetivibrio cellulolyticus that contains a family 9 glycosyl hydrolase. AB - A novel cellulosomal scaffoldin gene, termed cipV, was identified and sequenced from the mesophilic cellulolytic anaerobe Acetivibrio cellulolyticus. Initial identification of the protein was based on a combination of properties, including its high molecular weight, cellulose-binding activity, glycoprotein nature, and immuno-cross-reactivity with the cellulosomal scaffoldin of Clostridium thermocellum. The cipV gene is 5,748 bp in length and encodes a 1,915-residue polypeptide with a calculated molecular weight of 199,496. CipV contains an N terminal signal peptide, seven type I cohesin domains, an internal family III cellulose-binding domain (CBD), and an X2 module of unknown function in tandem with a type II dockerin domain at the C terminus. Surprisingly, CipV also possesses at its N terminus a catalytic module that belongs to the family 9 glycosyl hydrolases. Sequence analysis indicated the following. (i) The repeating cohesin domains are very similar to each other, ranging between 70 and 90% identity, and they also have about 30 to 40% homology with each of the other known type I scaffoldin cohesins. (ii) The internal CBD belongs to family III but differs from other known scaffoldin CBDs by the omission of a 9-residue stretch that constitutes a characteristic loop previously associated with the scaffoldins. (iii) The C-terminal type II dockerin domain is only the second such domain to have been discovered; its predicted "recognition codes" differ from those proposed for the other known dockerins. The putative calcium-binding loop includes an unusual insert, lacking in all the known type I and type II dockerins. (iv) The X2 module has about 60% sequence homology with that of C. thermocellum and appears at the same position in the scaffoldin. (v) Unlike the other known family 9 catalytic modules of bacterial origin, the CipV catalytic module is not accompanied by a flanking helper module, e.g., an adjacent family IIIc CBD or an immunoglobulin-like domain. Comparative sequence analysis of the CipV functional modules with those of the previously sequenced scaffoldins provides new insight into the structural arrangement and phylogeny of this intriguing family of microbial proteins. The modular organization of CipV is reminiscent of that of the CipA scaffoldin from C. thermocellum as opposed to the known scaffoldins from the mesophilic clostridia. The phylogenetic relationship of the different functional modules appears to indicate that the evolution of the scaffoldins reflects a collection of independent events and mechanisms whereby individual modules and other constituents are incorporated into the scaffoldin gene from different microbial sources. PMID- 10542175 TI - Loss of cytochrome c oxidase activity and acquisition of resistance to quinone analogs in a laccase-positive variant of Azospirillum lipoferum. AB - Laccase, a p-diphenol oxidase typical of plants and fungi, has been found recently in a proteobacterium, Azospirillum lipoferum. Laccase activity was detected in both a natural isolate and an in vitro-obtained phase variant that originated from the laccase-negative wild type. In this study, the electron transport systems of the laccase-positive variant and its parental laccase negative forms were compared. During exponential (but not stationary) growth under fully aerobic (but not under microaerobic) conditions, the laccase-positive variant lost a respiratory branch that is terminated in a cytochrome c oxidase of the aa(3) type; this was most likely due to a defect in the biosynthesis of a heme component essential for the oxidase. The laccase-positive variant was significantly less sensitive to the inhibitory action of quinone analogs and fully resistant to inhibitors of the bc(1) complex, apparently due to the rearrangements of its respiratory system. We propose that the loss of the cytochrome c oxidase-containing branch in the variant is an adaptive strategy to the presence of intracellular oxidized quinones, the products of laccase activity. PMID- 10542176 TI - Genetic and structural analyses of cytoplasmic filaments of wild-type Treponema phagedenis and a flagellar filament-deficient mutant. AB - Unique cytoplasmic filaments are found in the treponeme genus of spirochete bacteria. Their function is unknown, but their location underneath the periplasmic flagellar filaments (PFF) suggests a role in motility and/or cell structure. To better understand these unique structures, the gene coding for the cytoplasmic filaments, cfpA, was identified in various treponemal species. Treponema phagedenis cfpA was 2,037 nucleotides long, and the encoded polypeptide showed 78 to 100% amino acid sequence identity with the partial sequence of CfpA from T. denticola, T. vincentii, and T. pallidum subsp. pertenue. Wild-type T. phagedenis and a PFF-deficient isolate were analyzed by electron microscopy to assess the structural relationship of the cytoplasmic filaments and the PFF. The number of cytoplasmic filaments per cell of T. phagedenis (mean, 5.7) was compared with the number of PFF at each end of the cell (mean, 4.7); the results suggest that there is no direct one-to-one correlation at the cell end. Moreover, a structural link between these structures could not be demonstrated. The cytoplasmic filaments were also analyzed by electron microscopy at different stages of cell growth; this analysis revealed that they are cleaved before or during septum formation and before the nascent formation of PFF. A PFF-deficient mutant of T. phagedenis possessed cytoplasmic filaments similar to those of the wild type, suggesting that intact PFF are not required for their assembly and regulation. The extensive conservation of CfpA among pathogenic spirochetes suggests an important function, and structural analysis suggests that it is unlikely that the cytoplasmic filaments and the flagellar apparatus are physically linked. PMID- 10542177 TI - Bacterial phylogenetic clusters revealed by genome structure. AB - Current bacterial taxonomy is mostly based on phenotypic criteria, which may yield misleading interpretations in classification and identification. As a result, bacteria not closely related may be grouped together as a genus or species. For pathogenic bacteria, incorrect classification or misidentification could be disastrous. There is therefore an urgent need for appropriate methodologies to classify bacteria according to phylogeny and corresponding new approaches that permit their rapid and accurate identification. For this purpose, we have devised a strategy enabling us to resolve phylogenetic clusters of bacteria by comparing their genome structures. These structures were revealed by cleaving genomic DNA with the endonuclease I-CeuI, which cuts within the 23S ribosomal DNA (rDNA) sequences, and by mapping the resulting large DNA fragments with pulsed-field gel electrophoresis. We tested this experimental system on two representative bacterial genera: Salmonella and Pasteurella. Among Salmonella spp., I-CeuI mapping revealed virtually indistinguishable genome structures, demonstrating a high degree of structural conservation. Consistent with this, 16S rDNA sequences are also highly conserved among the Salmonella spp. In marked contrast, the Pasteurella strains have very different genome structures among and even within individual species. The divergence of Pasteurella was also reflected in 16S rDNA sequences and far exceeded that seen between Escherichia and Salmonella. Based on this diversity, the Pasteurella haemolytica strains we analyzed could be divided into 14 phylogenetic groups and the Pasteurella multocida strains could be divided into 9 groups. If criteria for defining bacterial species or genera similar to those used for Salmonella and Escherichia coli were applied, the striking phylogenetic diversity would allow bacteria in the currently recognized species of P. multocida and P. haemolytica to be divided into different species, genera, or even higher ranks. On the other hand, strains of Pasteurella ureae and Pasteurella pneumotropica are very similar to those of P. multocida in both genome structure and 16S rDNA sequence and should be regarded as strains within this species. We conclude that large-scale genome structure can be a sensitive indicator of phylogenetic relationships and that, therefore, I-CeuI-based genomic mapping is an efficient tool for probing the phylogenetic status of bacteria. PMID- 10542179 TI - Imbalanced base excision repair increases spontaneous mutation and alkylation sensitivity in Escherichia coli. AB - Inappropriate expression of 3-methyladenine (3MeA) DNA glycosylases has been shown to have harmful effects on microbial and mammalian cells. To understand the underlying reasons for this phenomenon, we have determined how DNA glycosylase activity and substrate specificity modulate glycosylase effects in Escherichia coli. We compared the effects of two 3MeA DNA glycosylases with very different substrate ranges, namely, the Saccharomyces cerevisiae Mag1 and the E. coli Tag glycosylases. Both glycosylases increased spontaneous mutation, decreased cell viability, and sensitized E. coli to killing by the alkylating agent methyl methanesulfonate. However, Tag had much less harmful effects than Mag1. The difference between the two enzymes' effects may be accounted for by the fact that Tag almost exclusively excises 3MeA lesions, whereas Mag1 excises a broad range of alkylated and other purines. We infer that the DNA lesions responsible for changes in spontaneous mutation, viability, and alkylation sensitivity are abasic sites and secondary lesions resulting from processing abasic sites via the base excision repair pathway. PMID- 10542178 TI - Influence of S-adenosylmethionine pool size on spontaneous mutation, dam methylation, and cell growth of Escherichia coli. AB - Escherichia coli strains that are deficient in the Ada and Ogt DNA repair methyltransferases display an elevated spontaneous G:C-to-A:T transition mutation rate, and this increase has been attributed to mutagenic O(6)-alkylguanine lesions being formed via the alkylation of DNA by endogenous metabolites. Here we test the frequently cited hypothesis that S-adenosylmethionine (SAM) can act as a weak alkylating agent in vivo and that it contributes to endogenous DNA alkylation. By regulating the expression of the rat liver SAM synthetase and the bacteriophage T3 SAM hydrolase proteins in E. coli, a 100-fold range of SAM levels could be achieved. However, neither increasing nor decreasing SAM levels significantly affected spontaneous mutation rates, leading us to conclude that SAM is not a major contributor to the endogenous formation of O(6)-methylguanine lesions in E. coli. PMID- 10542180 TI - The CpxRA signal transduction system of Escherichia coli: growth-related autoactivation and control of unanticipated target operons. AB - In Escherichia coli, the CpxRA two-component signal transduction system senses and responds to aggregated and misfolded proteins in the bacterial envelope. We show that CpxR-P (the phosphorylated form of the cognate response regulator) activates cpxRA expression in conjunction with RpoS, suggesting an involvement of the Cpx system in stationary-phase survival. Engagement of the CpxRA system in functions beyond protein management is indicated by several putative targets identified after a genomic screening for the CpxR-P recognition consensus sequence. Direct negative control of the newly identified targets motABcheAW (specifying motility and chemotaxis) and tsr (encoding the serine chemoreceptor) by CpxR-P was shown by electrophoretic mobility shift analysis and Northern hybridization. The results suggest that the CpxRA system plays a core role in an extensive stress response network in which the coordination of protein turnover and energy conservation may be the unifying element. PMID- 10542181 TI - The Vibrio cholerae O139 Calcutta bacteriophage CTXphi is infectious and encodes a novel repressor. AB - CTXphi is a lysogenic, filamentous bacteriophage. Its genome includes the genes encoding cholera toxin (ctxAB), one of the principal virulence factors of Vibrio cholerae; consequently, nonpathogenic strains of V. cholerae can be converted into toxigenic strains by CTXphi infection. O139 Calcutta strains of V. cholerae, which were linked to cholera outbreaks in Calcutta, India, in 1996, are novel pathogenic strains that carry two distinct CTX prophages integrated in tandem: CTX(ET), the prophage previously characterized within El Tor strains, and a new CTX Calcutta prophage (CTX(calc)). We found that the CTX(calc) prophage gives rise to infectious virions; thus, CTX(ET)phi is no longer the only known vector for transmission of ctxAB. The most functionally significant differences between the nucleotide sequences of CTX(calc)phi and CTX(ET)phi are located within the phages' repressor genes (rstR(calc) and rstR(ET), respectively) and their RstR operators. RstR(calc) is a novel, allele-specific repressor that regulates replication of CTX(calc)phi by inhibiting the activity of the rstA(calc) promoter. RstR(calc) has no inhibitory effect upon the classical and El Tor rstA promoters, which are instead regulated by their cognate RstRs. Consequently, production of RstR(calc) renders a CTX(calc) lysogen immune to superinfection by CTX(calc)phi but susceptible (heteroimmune) to infection by CTX(ET)phi. Analysis of the prophage arrays generated by sequentially integrated CTX phages revealed that pathogenic V. cholerae O139 Calcutta probably arose via infection of an O139 CTX(ET)phi lysogen by CTX(calc)phi. PMID- 10542182 TI - Structural characterization of the symbiotically important low-molecular-weight succinoglycan of Sinorhizobium meliloti. AB - The production of succinoglycan by Sinorhizobium meliloti Rm1021 is required for successful nodule invasion by the bacterium of its host plant, alfalfa. Rm1021 produces succinoglycan, an acidic exopolysaccharide composed of an octasaccharide repeating unit modified with acetyl, succinyl, and pyruvyl moieties, in both low- and high-molecular-weight forms. Low-molecular-weight (LMW) succinoglycan, previously thought to consist of monomers, trimers, and tetramers of the repeating unit, has been reported as being capable of promoting the formation of nitrogen-fixing nodules by succinoglycan-deficient derivatives of strain Rm1021. We have determined that the three size classes of LMW succinoglycan species are in fact monomers, dimers, and trimers of the repeating unit and that the trimer is the species active in promoting nodule invasion. A detailed structural analysis of the components of LMW succinoglycan by using various chromatographic techniques, along with nuclear magnetic resonance analyses, has revealed that there is considerable heterogeneity within the LMW succinoglycan oligomers in terms of noncarbohydrate substitutions, and we have determined the structural basis of this heterogeneity. PMID- 10542183 TI - Outer membrane lipoprotein e (P4) of Haemophilus influenzae is a novel phosphomonoesterase. AB - Haemophilus influenzae exists as a commensal of the upper respiratory tract of humans but also causes infections of contiguous structures. We describe the identification, localization, purification, and characterization of a novel, surface-localized phosphomonoesterase from a nontypeable H. influenzae strain, R2866. Sequences obtained from two CNBr-derived fragments of this protein matched lipoprotein e (P4) within the H. influenzae sequence database. Escherichia coli DH5alpha transformed with plasmids containing the H. influenzae hel gene, which encodes lipoprotein e (P4), produced high levels of a membrane-associated phosphomonoesterase. The isolated approximately 28-kDa enzyme was tartrate resistant and displayed narrow substrate specificity with the highest activity for arylphosphates, excluding 5-bromo-4-chloro-3-indolylphosphate. Optimum enzymatic activity was observed at pH 5.0 and only in the presence of divalent copper. The enzyme was inhibited by vanadate, molybdate, and EDTA but was resistant to inorganic phosphate. The association of phosphomonoesterase activity with a protein that has also been recognized as a heme transporter suggests a unique role for this unusual phosphohydrolase. PMID- 10542184 TI - Role of crotonyl coenzyme A reductase in determining the ratio of polyketides monensin A and monensin B produced by Streptomyces cinnamonensis. AB - The ccr gene, encoding crotonyl coenzyme A (CoA) reductase (CCR), was cloned from Streptomyces cinnamonensis C730.1 and shown to encode a protein with 90% amino acid sequence identity to the CCRs of Streptomyces collinus and Streptomyces coelicolor. A ccr-disrupted mutant, S. cinnamonensis L1, was constructed by inserting the hyg resistance gene into a unique BglII site within the ccr coding region. By use of the ermE* promoter, the S. collinus ccr gene was expressed from plasmids in S. cinnamonensis C730. 1/pHL18 and L1/pHL18. CCR activity in mutant L1 was shown to decrease by more than 90% in both yeast extract-malt extract (YEME) medium and a complex fermentation medium, compared to that in wild-type C730.1. Compared to C730.1, mutants C730.1/pHL18 and L1/pHL18 exhibited a huge increase in CCR activity (14- and 13-fold, respectively) in YEME medium and a moderate increase (3.7- and 2. 7-fold, respectively) in the complex fermentation medium. In the complex fermentation medium, S. cinnamonensis L1 produced monensins A and B in a ratio of 12:88, dramatically lower than the 50:50 ratio observed for both C730.1 and C730.1/pHL18. Plasmid (pHL18)-based expression of the S. collinus ccr gene in mutant L1 increased the monensin A/monensin B ratio to 42:58. Labeling experiments with [1, 2-(13)C(2)]acetate demonstrated the same levels of intact incorporation of this material into the butyrate-derived portion of monensin A in both C730.1 and mutant C730.1/pLH18 but a markedly decreased level of such incorporation in mutant L1. The addition of crotonic acid at 15 mM led to significant increases in the monensin A/monensin B ratio in C730.1 and C730.1/pHL18 but had no effect in S. cinnamonensis L1. These results demonstrate that CCR plays a significant role in providing butyryl-CoA for monensin A biosynthesis and is present in wild-type S. cinnamonensis C730.1 at a level sufficient that the availability of the appropriate substrate (crotonyl-CoA) is limiting. PMID- 10542186 TI - Evidence of horizontal transfer of the EcoO109I restriction-modification gene to Escherichia coli chromosomal DNA. AB - A DNA fragment carrying the genes coding for EcoO109I endonuclease and EcoO109I methylase, which recognize the nucleotide sequence 5'-(A/G)GGNCC(C/T)-3', was cloned from the chromosomal DNA of Escherichia coli H709c. The EcoO109I restriction-modification (R-M) system was found to be inserted between the int and psu genes from satellite bacteriophage P4, which were lysogenized in the chromosome at the P4 phage attachment site of the corresponding leuX gene observed in E. coli K-12 chromosomal DNA. The sid gene of the prophage was inactivated by insertion of one copy of IS21. These findings may shed light on the horizontal transfer and stable maintenance of the R-M system. PMID- 10542187 TI - Mutations in the regulatory gene hrpG of Xanthomonas campestris pv. vesicatoria result in constitutive expression of all hrp genes. AB - hrpG is a key regulatory gene for transcriptional activation of pathogenicity genes (hrp) of Xanthomonas campestris pv. vesicatoria. We identified three mutations in hrpG which render hrp gene expression constitutive in normally suppressing medium. The mutations in hrpG result in novel amino acid substitutions compared to mutations in related proteins, such as OmpR. In addition, mutated hrpG enhances the timing and intensity of plant reactions in infection assays. PMID- 10542185 TI - Cloning and expression of Mycobacterium tuberculosis and Mycobacterium leprae dihydropteroate synthase in Escherichia coli. AB - The genes for dihydropteroate synthase of Mycobacterium tuberculosis and Mycobacterium leprae were isolated by hybridization with probes amplified from the genomic DNA libraries. DNA sequencing revealed an open reading frame of 840 bp encoding a protein of 280 amino acids for M. tuberculosis dihydropteroate synthase and an open reading frame of 852 bp encoding a protein of 284 amino acids for M. leprae dihydropteroate synthase. The dihydropteroate synthases were expressed under control of the T5 promoter in a dihydropteroate synthase deficient strain of Escherichia coli. Using three chromatography steps, we purified both M. tuberculosis and M. leprae dihydropteroate synthases to >98% homogeneity. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed molecular masses of 29 kDa for M. tuberculosis dihydropteroate synthase and 30 kDa for M. leprae dihydropteroate synthase. Gel filtration of both enzymes showed a molecular mass of ca. 60 kDa, indicating that the native enzymes exist as dimers of two identical subunits. Steady-state kinetic parameters for dihydropteroate synthases from both M. tuberculosis and M. leprae were determined. Representative sulfonamides and dapsone were potent inhibitors of the mycobacterial dihydropteroate synthases, but the antimycobacterial agent p aminosalicylate, a putative dihydropteroate synthase inhibitor, was a poor inhibitor of the enzymes. PMID- 10542188 TI - The BldD protein from Streptomyces coelicolor is a DNA-binding protein. AB - Gel mobility shift assays with His-tagged BldD isolated from Escherichia coli have illustrated that BldD is capable of specifically recognizing its own promoter region. DNase I and hydroxyl radical footprinting assays have served to delimit the BldD binding site, revealing that BldD recognizes and binds to a site just upstream from, and overlapping with, the -10 region of the promoter. How BldD binds to its promoter and the effect this binding has on the expression of BldD are discussed. PMID- 10542189 TI - Expression of the Escherichia coli ada regulon in stationary phase: evidence for rpoS-dependent negative regulation of alkA transcription. AB - The Escherichia coli Ada protein activates sigma(70)-dependent transcription at three different promoters (ada, aidB, and alkA) in response to alkylation damage of DNA. During stationary phase, however, the methylated form of Ada shuts off expression of alkA; this repression is specific for sigma(S)-dependent transcription. Thus, at the alkA promoter, the Ada protein can act as both a positive and negative modulator of the adaptive response to alkylation damage, depending on the cell's physiological state. PMID- 10542190 TI - Identification and disruption of lisRK, a genetic locus encoding a two-component signal transduction system involved in stress tolerance and virulence in Listeria monocytogenes. AB - lisRK encodes a two-component regulatory system in the food pathogen Listeria monocytogenes LO28. Following identification of the operon in an acid-tolerant Tn917 mutant, a deletion in the histidine kinase component was shown to result in a growth phase variation in acid tolerance, an ability to grow in high ethanol concentrations, and a significant reduction in virulence. PMID- 10542191 TI - The IntI1 integron integrase preferentially binds single-stranded DNA of the attC site. AB - IntI1 integrase is a member of the prokaryotic DNA integrase superfamily. It is responsible for mobility of antibiotic resistance cassettes found in integrons. IntI1 protein, as well as IntI1-COOH, a truncated form containing its carboxy terminal domain, has been purified. Electrophoretic mobility shift assays were carried out to study the ability of IntI1 to bind the integrase primary target sites attI and aadA1 attC. When using double-stranded DNA as a substrate, we observed IntI1 binding to attI but not to attC. IntI1-COOH did not bind either attI or attC, indicating that the N-terminal domain of IntI1 was required for binding to double-stranded attI. On the other hand, when we used single-stranded (ss) DNA substrates, IntI1 bound strongly and specifically to ss attC DNA. Binding was strand specific, since only the bottom DNA strand was bound. Protein IntI1-COOH bound ss attC as well as did the complete integrase, indicating that the ability of the protein to bind ss aadA1 attC was contained in the region between amino acids 109 and 337 of IntI1. Binding to ss attI DNA by the integrase, but not by IntI1-COOH, was also observed and was specific for the attI bottom strand, indicating similar capabilities of IntI1 for binding attI DNA in either double-stranded or ss conformation. Footprinting analysis showed that IntI1 protected at least 40 bases of aadA1 attC against DNase I attack. The protected sequence contained two of the four previously proposed IntI1 DNA binding sites, including the crossover site. Preferential ssDNA binding can be a significant activity of IntI1 integrase, which suggests the utilization of extruded cruciforms in the reaction mechanisms leading to cassette excision and integration. PMID- 10542192 TI - The Agrobacterium tumefaciens chaperone-like protein, VirE1, interacts with VirE2 at domains required for single-stranded DNA binding and cooperative interaction. AB - Agrobacterium tumefaciens transfers single-stranded DNA (ssDNA) into plants. Efficient tumorigenesis requires VirE1-dependent export of ssDNA-binding (SSB) protein VirE2. VirE1 binds VirE2 domains involved in SSB and self-association, and VirE1 may facilitate VirE2 export by preventing VirE2 aggregation and the premature binding of VirE2 to ssDNA. PMID- 10542193 TI - The structure and assembly of secreted mucins. PMID- 10542194 TI - A detailed molecular belt model for apolipoprotein A-I in discoidal high density lipoprotein. AB - Apolipoprotein A-I (apoA-I) is the principal protein of high density lipoprotein particles (HDL). ApoA-I contains a globular N-terminal domain (residues 1-43) and a lipid-binding C-terminal domain (residues 44-243). Here we propose a detailed model for the smallest discoidal HDL, consisting of two apoA-I molecules wrapped beltwise around a small patch of bilayer containing 160 lipid molecules. The C terminal domain of each monomer is ringlike, a curved, planar amphipathic alpha helix with an average of 3.67 residues per turn, and with the hydrophobic surface curved toward the lipids. We have explored all possible geometries for forming the dimer of stacked rings, subject to the hypothesis that the optimal geometry will maximize intermolecular salt bridge interactions. The resulting model is an antiparallel arrangement with an alignment matching that of the (nonplanar) crystal structure of lipid-free apoA-I. PMID- 10542195 TI - Homology between egg white sulfhydryl oxidase and quiescin Q6 defines a new class of flavin-linked sulfhydryl oxidases. AB - The flavin-dependent sulfhydryl oxidase from chicken egg white catalyzes the oxidation of sulfhydryl groups to disulfides with the reduction of oxygen to hydrogen peroxide. Reduced proteins are the preferred thiol substrates of this secreted enzyme. The egg white oxidase shows an average 64% identity (from randomly distributed peptides comprising more than 30% of the protein sequence) to a human protein, Quiescin Q6, involved in growth regulation. Q6 is strongly expressed when fibroblasts enter reversible quiescence (Coppock, D. L., Cina Poppe, D., Gilleran, S. (1998) Genomics 54, 460-468). A peptide antibody against Q6 cross-reacts with both the egg white enzyme and a flavin-linked sulfhydryl oxidase isolated from bovine semen. Sequence analyses show that the egg white oxidase joins human Q6, bone-derived growth factor, GEC-3 from guinea pig, and homologs found in a range of multicellular organisms as a member of a new protein family. These proteins are formed from the fusion of thioredoxin and ERV motifs. In contrast, the flavin-linked sulfhydryl oxidase from Aspergillus niger is related to the pyridine nucleotide-dependent disulfide oxidoreductases, and shows no detectable sequence similarity to this newly recognized protein family. PMID- 10542196 TI - The mutagenesis protein UmuC is a DNA polymerase activated by UmuD', RecA, and SSB and is specialized for translesion replication. AB - Replication of DNA lesions leads to the formation of mutations. In Escherichia coli this process is regulated by the SOS stress response, and requires the mutagenesis proteins UmuC and UmuD'. Analysis of translesion replication using a recently reconstituted in vitro system (Reuven, N. B., Tomer, G., and Livneh, Z. (1998) Mol. Cell 2, 191-199) revealed that lesion bypass occurred with a UmuC fusion protein, UmuD', RecA, and SSB in the absence of added DNA polymerase. Further analysis revealed that UmuC was a DNA polymerase (E. coli DNA polymerase V), with a weak polymerizing activity. Upon addition of UmuD', RecA, and SSB, the UmuC DNA polymerase was greatly activated, and replicated a synthetic abasic site with great efficiency (45% bypass in 6 min), 10-100-fold higher than E. coli DNA polymerases I, II, or III holoenzyme. Analysis of bypass products revealed insertion of primarily dAMP (69%), and to a lesser degree dGMP (31%) opposite the abasic site. The UmuC104 mutant protein was defective both in lesion bypass and in DNA synthesis. These results indicate that UmuC is a UmuD'-, RecA-, and SSB activated DNA polymerase, which is specialized for lesion bypass. UmuC is a member of a new family of DNA polymerases which are specialized for lesion bypass, and include the yeast RAD30 and the human XP-V genes, encoding DNA polymerase eta. PMID- 10542197 TI - The biotin domain peptide from the biotin carboxyl carrier protein of Escherichia coli acetyl-CoA carboxylase causes a marked increase in the catalytic efficiency of biotin carboxylase and carboxyltransferase relative to free biotin. AB - Acetyl-CoA carboxylase catalyzes the first committed step in the biosynthesis of long-chain fatty acids. The Escherichia coli form of the enzyme consists of a biotin carboxylase activity, a biotin carboxyl carrier protein, and a carboxyltransferase activity. The C-terminal 87 amino acids of the biotin carboxyl carrier protein (BCCP87) form a domain that can be independently expressed, biotinylated, and purified (Chapman-Smith, A., Turner, D. L., Cronan, J. E., Morris, T. W., and Wallace, J. C. (1994) Biochem. J. 302, 881-887). The ability of the biotinylated form of this 87-residue protein (holoBCCP87) to act as a substrate for biotin carboxylase and carboxyltransferase was assessed and compared with the results with free biotin. In the case of biotin carboxylase holoBCCP87 was an excellent substrate with a K(m) of 0.16 +/- 0.05 mM and V(max) of 1000.8 +/- 182.0 min(-1). The V/K or catalytic efficiency of biotin carboxylase with holoBCCP87 as substrate was 8000-fold greater than with biotin as substrate. Stimulation of the ATP synthesis reaction of biotin carboxylase where carbamyl phosphate reacted with ADP by holoBCCP87 was 5-fold greater than with an equivalent amount of biotin. The interaction of holoBCCP87 with carboxyltransferase was characterized in the reverse direction where malonyl-CoA reacted with holoBCCP87 to form acetyl-CoA and carboxyholoBCCP87. The K(m) for holoBCCP87 was 0.45 +/- 0.07 mM while the V(max) was 2031.8 +/- 231.0 min(-1). The V/K or catalytic efficiency of carboxyltransferase with holoBCCP87 as substrate is 2000-fold greater than with biotin as substrate. PMID- 10542198 TI - Rapid STAT phosphorylation via the B cell receptor. Modulatory role of CD19. AB - Engagement of the B cell receptor (BCR) initiates multiple signaling cascades which mediate different biological responses, depending on the stage of B cell differentiation, antigen binding affinity, and duration of stimulation. Aggregation of co-receptors such as CD19 with the antigen receptor has been suggested to modulate the signals necessary for the development and functioning of the humoral immune system. In this study, we demonstrate that engagement of the antigen receptor on peripheral blood B cells, but not naive splenic B lymphocytes, leads to rapid phosphorylation of signal transducers and activators of transcription 1 (STAT1) on Tyr-701 and Ser-727. Interestingly, phosphorylation on tyrosine diminished with increased stimulation, whereas serine phosphorylation correlated directly with the level of BCR cross-linking. In contrast, phosphorylation of STAT3 occurs exclusively on serine and is sensitive to inhibitors of the PI3-kinase and the ERK1/2 pathways. Finally, we show that co ligation of CD19 with the BCR results in increased tyrosine phosphorylation of STAT1 relative to BCR cross-linking alone, establishing CD19 as a positive modulator of BCR-mediated STAT activation. PMID- 10542199 TI - Cdc2 and Cdk2 kinase activated by transforming growth factor-beta1 trigger apoptosis through the phosphorylation of retinoblastoma protein in FaO hepatoma cells. AB - The signaling pathway leading to TGF-beta1-induced apoptosis was investigated using a TGF-beta1-sensitive hepatoma cell line, FaO. Cell cycle analysis demonstrated that the accumulation of apoptotic cells was preceded by a progressive decrease of the cell population in the G(1) phase concomitant with a slight increase of the cell population in the G(2)/M phase in response to TGF beta1. TGF-beta1 induced a transient increase in the expression of Cdc2, cyclin A, cyclin B, and cyclin D1 at an early phase of apoptosis. During TGF-beta1 induced apoptosis, the transient increase in cyclin-dependent kinase (Cdk) activities coincides with a dramatic increase in the hyperphosphorylated forms of RB. Treatment with roscovitine or olomoucine, inhibitors of Cdc2 and Cdk2, blocked TGF-beta1-induced apoptosis by inhibiting RB phosphorylation. Overexpression of Bcl-2 or adenovirus E1B 19K suppressed TGF-beta1-induced apoptosis by blocking the induction of Cdc2 mRNA and the subsequent activation of Cdc2 kinase, whereas activation of Cdk2 was not affected, suggesting that Cdc2 plays a more critical role in TGF-beta1-induced apoptosis. In conclusion, we present the evidence that Cdc2 and Cdk2 kinase activity transiently induced by TGF-beta1 phosphorylates RB as a physiological target in FaO cells and that RB hyperphosphorylation may trigger abrupt cell cycle progression, leading to irreversible cell death. PMID- 10542200 TI - Purinoceptors evoke different electrophysiological responses in pancreatic ducts. P2Y inhibits K(+) conductance, and P2X stimulates cation conductance. AB - In epithelia, extracellular nucleotides are often associated with regulation of ion transporters, especially Cl(-) channels. In this study, we investigated which purinoceptors are present in native pancreatic ducts and how they regulate ion transport. We applied whole-cell patch-clamp recordings, intracellular Ca(2+) and pH measurements, and reverse transcription-polymerase chain reaction (RT-PCR) analysis. The data show two types of purinoceptors and cellular responses. UTP and ATP produced large Ca(2+) transients, a decrease in intracellular pH, 8-10-mV depolarization of the membrane voltage, and a decrease in the whole-cell conductance. The membrane effects were due to closure of K(+) channels, as confirmed by dependence on extracellular K(+). UTP/ATP effects could be associated with P2Y(2) purinoceptors, and RT-PCR revealed mRNAs for P2Y(2) and P2Y(4) receptors. On the other hand, 2', 3'-O-4-benzoylbenzoyl-ATP induced Ca(2+) influx and approximately 20-mV depolarization of the membrane voltage with a concomitant increase in the whole-cell conductance. These effects were dependent on extracellular Na(+), not Cl(-), indicating opening of cation channels associated with P2X(7) purinoceptors. RT-PCR showed mRNAs for P2X(7) and P2X(4) receptors. In microperfused ducts, luminal (but not basolateral) ATP caused large depolarizations of membrane voltages recorded with microelectrodes, consistent with luminal localization of P2X(7) receptors. Thus, P2Y(2) (and possibly P2Y(4)) purinoceptors inhibit K(+) channels and may not support secretion in native ducts. P2X(7) (and possibly P2X(4)) receptors are associated with cation channels and may contribute to regulation of secretion. PMID- 10542201 TI - Acid-labile ATP and/or ADP/P(i) binding to the tetraprotomeric form of Na/K ATPase accompanying catalytic phosphorylation-dephosphorylation cycle. AB - The Na/K-ATPase has been shown to bind 1 and 0.5 mol of (32)P/mol of alpha-chain in the presence [gamma-(32)P]ATP and [alpha-(32)P]ATP, respectively, accompanied by a maximum accumulation of 0.5 mol of ADP-sensitive phosphoenzyme (NaE1P) and potassium-sensitive phosphoenzyme (E2P). The former accumulation was followed by the slow constant liberation of P(i), but the latter was accompanied with a rapid approximately 0.25 mol of acid-labile P(i) burst. The rubidium (potassium congener)-occluded enzyme (approximately 1.7 mol of rubidium/mol of alpha-chain) completely lost rubidium on the addition of sodium + magnesium. Further addition of approximately 100 microM [gamma-(32)P]ATP and [alpha-(32)P]ATP, both induced 0.5 mol of (32)P-ATP binding to the enzyme and caused accumulation of approximately 1 mol of rubidium/mol of alpha-chain, accompanied by a rapid approximately 0.5 mol of P(i) burst with no detectable phosphoenzyme under steady state conditions. Electron microscopy of rotary-shadowed soluble and membrane bound Na/K-ATPases and an antibody-Na/K-ATPase complex, indicated the presence of tetraprotomeric structures (alphabeta)(4). These and other data suggest that Na/K ATP hydrolysis occurs via four parallel paths, the sequential appearance of (NaE1P:E.ATP)(2), (E2P:E.ATP:E2P:E. ADP/P(i)), and (KE2:E.ADP/P(i))(2), each of which has been previously referred to as NaE1P, E2P, and KE2, respectively. PMID- 10542202 TI - Modification of the ADP-ribosyltransferase and NAD glycohydrolase activities of a mammalian transferase (ADP-ribosyltransferase 5) by auto-ADP-ribosylation. AB - Mono-ADP-ribosylation, a post-translational modification in which the ADP-ribose moiety of NAD is transferred to an acceptor protein, is catalyzed by a family of amino acid-specific ADP-ribosyltransferases. ADP-ribosyltransferase 5 (ART5), a murine transferase originally isolated from Yac-1 lymphoma cells, differed in properties from previously identified eukaryotic transferases in that it exhibited significant NAD glycohydrolase (NADase) activity. To investigate the mechanism of regulation of transferase and NADase activities, ART5 was synthesized as a FLAG fusion protein in Escherichia coli. Agmatine was used as the ADP-ribose acceptor to quantify transferase activity. ART5 was found to be primarily an NADase at 10 microM NAD, whereas at higher NAD concentrations (1 mM), after some delay, transferase activity increased, whereas NADase activity fell. This change in catalytic activity was correlated with auto-ADP-ribosylation and occurred in a time- and NAD concentration-dependent manner. Based on the change in mobility of auto-ADP-ribosylated ART5 by SDS-polyacrylamide gel electrophoresis, the modification appeared to be stoichiometric and resulted in the addition of at least two ADP-ribose moieties. Auto-ADP-ribosylated ART5 isolated after incubation with NAD was primarily a transferase. These findings suggest that auto-ADP-ribosylation of ART5 was stoichiometric, resulted in at least two modifications and converted ART5 from an NADase to a transferase, and could be one mechanism for regulating enzyme activity. PMID- 10542203 TI - Structure of the vacuolar ATPase by electron microscopy. AB - The structure of the vacuolar ATPase from bovine brain clathrin-coated vesicles has been determined by electron microscopy of negatively stained, detergent solubilized enzyme molecules. Preparations of both lipid-containing and delipidated enzyme have been analyzed. The complex is organized in two major domains, a V(1) and V(0), with overall dimensions of 28 x 14 x 14 nm. The V(1) is a more or less spherical molecule with a central cavity. The V(0) has the shape of a flattened sphere or doughnut with a radius of about 100 A. The V(1) and V(0) are joined by a 60-A long and 40-A wide central stalk, consisting of several individual protein densities. Two kinds of smaller densities are visible at the top periphery of the V(1), and one of these seems to extend all the way down to the stalk domain in some averages. Images of both the lipid-containing and the delipidated complex show a 30-50-kDa protein density on the lumenal side of the complex, opposite the central stalk, centered in the ring of c subunits. A large trans-membrane mass, probably the C-terminal domain of the 100-kDa subunit a, is seen at the periphery of the c subunit ring in some projections. This large mass has both a lumenal and a cytosolic domain, and it is the cytosolic domain that interacts with the central stalk. Two to three additional protein densities can be seen in the V(1)-V(0) interface, all connected to the central stalk. Overall, the structure of the V-ATPase is similar to the structure of the related F(1)F(0) ATP synthase, confirming their common origin. PMID- 10542204 TI - Function of the factor I modules (FIMS) of human complement component C6. AB - In order to elucidate the function of complement component C6, truncated C6 molecules were expressed recombinantly. These were either deleted of the factor I modules (FIMs) (C6des-748-913) or both complement control protein (CCP) modules and FIMs (C6des-611-913). C6des-748-913 exhibited approximately 60-70% of the hemolytic activity of full-length C6 when assayed for Alternative Pathway activity, but when measured for the Classical Pathway, C6des-748-914 was only 4 6% as effective as C6. The activity difference between C6 and C6des-748-913 for the two complement pathways can be explained by a greater stability of newly formed metastable C5b* when produced by the Alternative Pathway compared with that made by the Classical Pathway. The half-lives of metastable C5b* and the decay of (125)I-C5b measured from cells used to activate the Alternative Pathway were found to be about 5-12-fold longer than those same parameters derived from cells that had activated the Classical Pathway. (125)I-C5 binds reversibly to C6 in an ionic strength-dependent fashion, but (125)I-C5 binds only weakly to C6des FIMs and not at all to C6des-CCP/FIMs. Therefore, although the FIMs are not required absolutely for C6 activity, these modules promote interaction of C6 with C5 enabling a more efficient bimolecular coupling ultimately leading to the formation of the C5b-6 complex. PMID- 10542205 TI - 1H NMR investigation of the distal hydrogen bonding network and ligand tilt in the cyanomet complex of oxygen-avid Ascaris suum hemoglobin. AB - The O(2)-avid hemoglobin from the parasitic nematode Ascaris suum exhibits one of the slowest known O(2) off rates. Solution (1)H NMR has been used to investigate the electronic and molecular structural properties of the active site for the cyano-met derivative of the recombinant first domain of this protein. Assignment of the heme, axial His, and majority of the residues in contact with the heme reveals a molecular structure that is the same as reported in the A. suum HbO(2) crystal structure (Yang, J., Kloek, A., Goldberg, D. E., and Mathews, F. S. (1995) Proc. Natl. Acad. Sci. U. S. A. 92, 4224-4228) with the exception that the heme in solution is rotated by 180 degrees about the alpha,gamma-meso axis relative to that in the crystal. The observed dipolar shifts, together with the crystal coordinates of HbO(2), provide the orientation of the magnetic axes in the molecular framework. The major magnetic axis, which correlates with the Fe-CN vector, is found oriented approximately 30 degrees away from the heme normal and indicates significant steric tilt because of interaction with Tyr(30)(B10). The three side chain labile protons for the distal residues Tyr(30)(B10) and Gln(64)(E7) were identified, and their relaxation, dipolar shifts, and nuclear Overhauser effects to adjacent residues used to place them in the distal pocket. It is shown that these two distal residues exhibit the same orientations ideal for H bonding to the ligand and to each other, as found in the A. suum HbO(2) crystal. It is concluded that the ligated cyanide participates in the same distal H bonding network as ligated O(2). The combination of the strong steric tilt of the bound cyanide and slow ring reorientation of the Tyr(30)(B10) side chain supports a crowded and constrained distal pocket. PMID- 10542206 TI - Biochemical characterization of various catalytic complexes of the brain platelet activating factor acetylhydrolase. AB - Brain intracellular platelet-activating factor acetylhydrolase (PAF-AH) isoform I is a member of a family of complex enzymes composed of mutually homologous alpha(1) and alpha(2) subunits, both of which account for catalytic activity, and the beta subunit. We previously demonstrated that the expression of one catalytic subunit, alpha(1), is developmentally regulated, resulting in a switching of the catalytic complex from alpha(1)/alpha(2) to alpha(2)/alpha(2) during brain development (Manya, H., Aoki, J., Watanabe, M., Adachi, T., Asou, H., Inoue, Y., Arai, H., and Inoue, K. (1998) J. Biol. Chem. 273, 18567-18572). In this study, we explored the biochemical differences in three possible catalytic dimers, alpha(1)/alpha(1), alpha(1)/alpha(2), and alpha(2)/alpha(2). The alpha(2)/alpha(2) homodimer exhibited different substrate specificity from the alpha(1)/alpha(1) homodimer and the alpha(1)/alpha(2) heterodimer, both of which showed similar substrate specificity. The alpha(2)/alpha(2) homodimer hydrolyzed PAF and 1-O-alkyl-2-acetyl-sn-glycero-3-phosphorylethanolamine (AAGPE) most efficiently among 1-O-alkyl-2-acetyl-phospholipids. In contrast, both alpha(1)/alpha(1) and alpha(1)/alpha(2) hydrolyzed 1-O-alkyl-2-acetyl-sn-glycero 3-phosphoric acid more efficiently than PAF. AAGPE was the poorest substrate for these enzymes. The beta subunit bound to all three catalytic dimers but modulated the enzyme activity in a catalytic dimer composition-dependent manner. The beta subunit strongly accelerated the enzyme activity of the alpha(2)/alpha(2) homodimer but rather suppressed the activity of the alpha(1)/alpha(1) homodimer and had little effect on that of the alpha(1)/alpha(2) heterodimer. The (His(149) to Arg) mutant beta, which has been recently identified in isolated lissencephaly sequence patients, lost the ability to either associate with the catalytic complexes or modulate their enzyme activity. The enzyme activity of PAF-AH isoform I may be regulated in multiple ways by switching the composition of the catalytic subunit and by manipulating the beta subunit. PMID- 10542207 TI - Glucokinase overexpression restores glucose utilization and storage in cultured hepatocytes from male Zucker diabetic fatty rats. AB - Zucker diabetic fatty rats develop type 2 diabetes concomitantly with peripheral insulin resistance. Hepatocytes from these rats and their control lean counterparts have been cultured, and a number of key parameters of glucose metabolism have been determined. Glucokinase activity was 4.5-fold lower in hepatocytes from diabetic rats than in hepatocytes from healthy ones. In contrast, hexokinase activity was about 2-fold higher in hepatocytes from diabetic animals than in healthy ones. Glucose-6-phosphatase activity was not significantly different. Despite the altered ratios of glucokinase to hexokinase activity, intracellular glucose 6-phosphate concentrations were similar in the two types of cells when they where incubated with 1-25 mM glucose. However, glycogen levels and glycogen synthase activity ratio were lower in hepatocytes from diabetic animals. Total pyruvate kinase activity and its activity ratio as well as fructose 2,6-bisphosphate concentration and lactate production were also lower in cells from diabetic animals. All of these data indicate that glucose metabolism is clearly impaired in hepatocytes from Zucker diabetic fatty rats. Glucokinase overexpression using adenovirus restored glucose metabolism in diabetic hepatocytes. In glucokinase-overexpressing cells, glucose 6-phosphate levels increased. Moreover, glycogen deposition was greatly enhanced due to the activation of glycogen synthase. Pyruvate kinase was also activated, and fructose 2,6-bisphosphate concentration and lactate production were increased in glucokinase-overexpressing diabetic hepatocytes. Overexpression of hexokinase I did not increase glycogen deposition. In conclusion, hepatocytes from Zucker diabetic fatty rats showed depressed glycogen and glycolytic metabolism, but glucokinase overexpression improved their glucose utilization and storage. PMID- 10542208 TI - The helicase from hepatitis C virus is active as an oligomer. AB - The helicase from hepatitis C virus (HCV NS3h) residing on the C-terminal domain of nonstructural protein 3 was considered to be monomeric by several researchers. Here we demonstrate, based on biochemical kinetic data, that the HCV helicase acts as an oligomer. The increase in the ATPase k(cat) of the NS3h protein with increasing protein concentration provided evidence for oligomerization. A sharp decrease in the unwinding rate was observed when the wild type NS3h was mixed with the ATPase deficient mutants of NS3h protein. This provided strong support for both mixed oligomer formation and subunit interactions for the HCV helicase. Chemical cross-linking of NS3h protein was an inefficient process, but yielded cross-linked protein oligomers of various sizes. The information currently available for HCV helicase is consistent with the hypothesis that oligomers of NS3h are not stable and the helicase subunits exchange during unwinding. Nevertheless, oligomerization of HCV helicase stimulates the ATPase activity, and it is required for the helicase activity. PMID- 10542209 TI - Nucleosome structural features and intrinsic properties of the TATAAACGCC repeat sequence. AB - Nucleosomes, the fundamental building blocks of chromatin, play an architectural role in ensuring the integrity of the genome and act as a regulator of transcription. Intrinsic properties of the underlying DNA sequence, such as flexibility and intrinsic bending, direct the formation of nucleosomes. We have earlier identified genomic nucleosome-positioning sequences with increased in vitro ability for nucleosome formation. One group of sequences bearing a 10-base pair consensus repeat sequence of TATAAACGCC had the highest reported nucleosome affinity from genomic material. Here, we report the intrinsic physical properties of this sequence and the structural details of the nucleosome it forms, as analyzed by footprinting techniques. The minor groove is buried toward the histone octamer at the AA steps and facing outwards at the CC steps. By cyclization kinetics, the overall helical repeat of the free DNA sequence was found to be 10.5 base pairs/turn. Our experiments also showed that this sequence is highly flexible, having a J-factor 25-fold higher than that of random sequence DNA. In addition, the data suggest that twist flexibility is an important determinant for translational nucleosome positioning, particularly over the dyad region. PMID- 10542210 TI - Expression of the gene for mitoribosomal protein S12 is controlled in human cells at the levels of transcription, RNA splicing, and translation. AB - The human gene RPMS12 encodes a protein similar to bacterial ribosomal protein S12 and is proposed to represent the human mitochondrial orthologue. RPMS12 reporter gene expression in cultured human cells supports the idea that the gene product is mitochondrial and is localized to the inner membrane. Human cells contain at least four structurally distinct RPMS12 mRNAs that differ in their 5' untranslated region (5'-UTR) as a result of alternate splicing and of 5' end heterogeneity. All of them encode the same polypeptide. The full 5'-UTR contains two types of sequence element implicated elsewhere in translational regulation as follows: a short upstream open reading frame and an oligopyrimidine tract similar to that found at the 5' end of mRNAs encoding other growth-regulated proteins, including those of cytosolic ribosomes. The fully spliced (short) mRNA is the predominant form in all cell types studied and is translationally down-regulated in cultured cells in response to serum starvation, even though it lacks both of the putative translational regulatory elements. By contrast, other splice variants containing one or both of these elements are not translationally regulated by growth status but are translated poorly in both growing and non growing cells. Reporter analysis identified a 26-nucleotide tract of the 5'-UTR of the short mRNA that is essential for translational down-regulation in growth inhibited cells. Such experiments also confirmed that the 5'-UTR of the longer mRNA variants contains negative regulatory elements for translation. Tissue representation of RPMS12 mRNA is highly variable, following a typical mitochondrial pattern, but the relative levels of the different splice variants are similar in different tissues. These findings indicate a complex, multilevel regulation of RPMS12 gene expression in response to signals mediating growth, tissue specialization, and probably metabolic needs. PMID- 10542211 TI - Kaposi's sarcoma-associated herpesvirus-encoded G protein-coupled receptor activation of c-jun amino-terminal kinase/stress-activated protein kinase and lyn kinase is mediated by related adhesion focal tyrosine kinase/proline-rich tyrosine kinase 2. AB - The Kaposi's sarcoma-associated herpesvirus (KSHV) (also known as human herpesvirus 8) has been implicated in the pathogenesis of Kaposi's sarcoma and B cell primary effusion lymphomas. KSHV encodes a G protein-coupled receptor (GPCR) that acts as an oncogene and constitutively activates two protein kinases, c-Jun amino-terminal kinase (JNK)/stress-activated protein kinase (SAPK) and p38 mitogen-activated protein kinase. It also induces the production of vascular endothelial growth factor. These processes are believed to be important in KSHV GPCR-related oncogenesis. We have characterized the signaling pathways mediated by KSHV-GPCR in a reconstituted 293T cell model in which the related adhesion focal tyrosine kinase (RAFTK) was ectopically expressed. RAFTK has been shown to play an important role in growth factor signaling in endothelium and in B cell antigen receptor signaling in B lymphocytes. KSHV-GPCR induced the tyrosine phosphorylation of RAFTK. Expression of wild-type RAFTK enhanced GPCR-mediated JNK/SAPK activation, whereas dominant-negative mutant constructs of RAFTK, such as K457A (which lacks kinase activity) and Y402F (a Src-binding mutant), inhibited KSHV-GPCR-mediated activation of JNK/SAPK. RAFTK also mediated the KSHV GPCR-induced activation of Lyn, a Src family kinase. However, RAFTK did not mediate the activation of p38 mitogen-activated protein kinase induced by KSHV GPCR. Human interferon gamma-inducible protein-10, which is known to inhibit KSHV GPCR activity, was found to reduce RAFTK phosphorylation and JNK/SAPK activation. These results suggest that in cells expressing RAFTK/proline-rich tyrosine kinase 2, such as endothelial and B cells, RAFTK can act to enhance KSHV-GPCR-mediated downstream signaling to transcriptional regulators such as JNK/SAPK. PMID- 10542212 TI - Interleukin-10 signaling blocks inhibitor of kappaB kinase activity and nuclear factor kappaB DNA binding. AB - The transcription factor nuclear factor kappaB (NF-kappaB) coordinates the activation of numerous genes in response to pathogens and proinflammatory cytokines and is, therefore, pivotal in the development of acute and chronic inflammatory diseases. In its inactive state, NF-kappaB is constitutively present in the cytoplasm as a p50-p65 heterodimer bound to its inhibitory protein IkappaB. Proinflammatory cytokines, such as tumor necrosis factor (TNF), activate NF-kappaB by stimulating the activity of the IkappaB kinases (IKKs) which phosphorylate IkappaBalpha on serine residues 32 and 36, targeting it for rapid degradation by the 26 S proteasome. This enables the release and nuclear translocation of the NF-kappaB complex and activation of gene transcription. Interleukin-10 (IL-10) is a pleiotropic cytokine that controls inflammatory processes by suppressing the production of proinflammatory cytokines which are known to be transcriptionally controlled by NF-kappaB. Conflicting data exists on the effects of IL-10 on TNF- and LPS-induced NF-kappaB activity in human monocytes and the molecular mechanisms involved have not been elucidated. In this study, we show that IL-10 functions to block NF-kappaB activity at two levels: 1) through the suppression of IKK activity and 2) through the inhibition of NF kappaB DNA binding activity. This is the first evidence of an anti-inflammatory protein inhibiting IKK activity and demonstrates that IKK is a logical target for blocking inflammatory diseases. PMID- 10542213 TI - Identification of the C-terminal part of Bordetella dermonecrotic toxin as a transglutaminase for rho GTPases. AB - Bordetella dermonecrotic toxin (DNT) causes the deamidation of glutamine 63 of Rho. Here we identified the region of DNT harboring the enzyme activity and compared the toxin with the cytotoxic necrotizing factor 1, which also deamidates Rho. The DNT fragment (DeltaDNT) covering amino acid residues 1136-1451 caused deamidation of RhoA at glutamine 63 as determined by mass spectrometric analysis and by the release of ammonia. In the presence of dansylcadaverine or ethylenediamine, DeltaDNT caused transglutamination of Rho. Deamidase and transglutaminase activities were blocked in the mutant proteins Cys(1292) --> Ala, His(1307) --> Ala, and Lys(1310) --> Ala of DeltaDNT. Deamidation and transglutamination induced by DeltaDNT blocked intrinsic and Rho- GTPase activating protein-stimulated GTPase activity of RhoA. DeltaDNT deamidated and transglutaminated Rac and Cdc42 in the absence and presence of ethylenediamine, respectively. Modification of Rho proteins by DeltaDNT was nucleotide-dependent and did not occur with GTPgammaS-loaded GTPases. In contrast to cytotoxic necrotizing factor, which caused the same kinetics of ammonia release in the absence and presence of ethylenediamine, ammonia release by DeltaDNT was largely increased in the presence of ethylenediamine, indicating that DeltaDNT acts primarily as a transglutaminase. PMID- 10542214 TI - Delineation of the structural basis for the activation properties of the dopamine D1 receptor subtypes. AB - To delineate the structural determinants involved in the constitutive activation of the D1 receptor subtypes, we have constructed chimeras between the D1A and D1B receptors. These chimeras harbored a cognate domain corresponding to transmembrane regions 6 and 7 as well as the third extracellular loop (EL3) and cytoplasmic tail, a domain referred herein to as the terminal receptor locus (TRL). A chimeric D1A receptor harboring the D1B-TRL (chimera 1) displays an increased affinity for dopamine that is indistinguishable from the wild-type D1B receptor. Likewise, a chimeric D1B receptor containing the D1A-TRL cassette (chimera 2) binds dopamine with a reduced affinity that is highly reminiscent of the dopamine affinity for the wild-type D1A receptor. Furthermore, we show that the agonist independent activity of chimera 1 is identical to the wild-type D1B receptor whereas the chimera 2 displays a low agonist independent activity that is indistinguishable from the wild-type D1A receptor. Dopamine potencies for the wild-type D1A and D1B receptor were recapitulated in cells expressing the chimera 2 or chimera 1, respectively. However, the differences observed in agonist mediated maximal activation of adenylyl cyclase elicited by the D1A and D1B receptors remain unchanged in cells expressing the chimeric receptors. To gain further mechanistic insights into the structural determinants of the TRL involved in the activation properties of the D1 receptor subtypes, we have engineered two additional chimeric D1 receptors that contain the EL3 region of their respective cognate wild-type counterparts (hD1A-EL3B and hD1B-EL3A). In marked contrast to chimera 1 and 2, dopamine affinity and constitutive activation were partially modulated by the exchange of the EL3. Meanwhile, hD1A-EL3B and hD1B-EL3A mutant receptors display a full switch in the agonist-mediated maximal activation, which is reminiscent of their cognate wild-type counterparts. Overall, our studies suggest a fundamental role for the TRL in shaping the intramolecular interactions implicated in the constitutive activation and coupling properties of the dopamine D1 receptor subtypes. PMID- 10542216 TI - The kinetic mechanism of EcoRI endonuclease. AB - Steady-state parameters governing cleavage of pBR322 DNA by EcoRI endonuclease are highly sensitive to ionic environment, with K(m) and k(cat) increasing 1,000 fold and 15-fold, respectively, when ionic strength is increased from 0.059 to 0.23 M. By contrast, pre-steady-state analysis has shown that recognition, as well as first and second strand cleavage events that occur once the enzyme has arrived at the EcoRI site, are essentially insensitive to ionic strength, and has demonstrated that the rate-limiting step for endonuclease turnover occurs after double-strand cleavage under all conditions tested. Furthermore, processive cleavage of a pBR322 variant bearing two closely spaced EcoRI sites is governed by the same turnover number as hydrolysis of parental pBR322, which contains only a single EcoRI sequence, ruling out slow release of the enzyme from the cleaved site or a slow conformational change subsequent to double-strand cleavage. We attribute the effects of ionic strength on steady-state parameters to nonspecific endonuclease.DNA interactions, reflecting facilitated diffusion processes, that occur prior to EcoRI sequence recognition and subsequent to DNA cleavage. PMID- 10542215 TI - Na(+)-dependent glutamate transporters (EAAT1, EAAT2, and EAAT3) of the blood brain barrier. A mechanism for glutamate removal. AB - Na(+)-dependent transporters for glutamate exist on astrocytes (EAAT1 and EAAT2) and neurons (EAAT3). These transporters presumably assist in keeping the glutamate concentration low in the extracellular fluid of brain. Recently, Na(+) dependent glutamate transport was described on the abluminal membrane of the blood-brain barrier. To determine whether the above-mentioned transporters participate in glutamate transport of the blood-brain barrier, total RNA was extracted from bovine cerebral capillaries. cDNA for EAAT1, EAAT2, and EAAT3 was observed, indicating that mRNA was present. Western blot analysis demonstrated all three transporters were expressed on abluminal membranes, but none was detectable on luminal membranes of the blood-brain barrier. Measurement of transport kinetics demonstrated voltage dependence, K(+)-dependence, and an apparent K(m) of 14 microM (aggregate of the three transporters) at a transmembrane potential of -61 mV. Inhibition of glutamate transport was observed using inhibitors specific for EAAT2 (kainic acid and dihydrokainic acid) and EAAT3 (cysteine). The relative activity of the three transporters was found to be approximately 1:3:6 for EAAT1, EAAT2, and EAAT3, respectively. These transporters may assist in maintaining low glutamate concentrations in the extracellular fluid. PMID- 10542217 TI - Isoforms of the Lutheran/basal cell adhesion molecule glycoprotein are differentially delivered in polarized epithelial cells. Mapping of the basolateral sorting signal to a cytoplasmic di-leucine motif. AB - Lu and Lu(v13) are two glycoprotein (gp) isoforms that belong to the immunoglobulin superfamily and carry both the Lutheran (Lu) blood group antigens and the basal cell adhesion molecule epithelial cancer antigen. Lu (85 kDa) and Lu(v13) (78 kDa) gps, which differ only in the length of their cytoplasmic domain, are adhesion molecules that bind laminin. In nonerythroid tissues, the Lu/basal cell adhesion molecule antigens are predominantly expressed in the endothelium of blood vessel walls and in the basement membrane region of normal epithelial cells, whereas they exhibit a nonpolarized expression in some epithelial cancers. Here, we analyzed the polarization of Lu and Lu(v13) gps in epithelial cells by confocal microscopy and domain-selective biotinylation assays. Differentiated human colon carcinoma Caco-2 cells exhibited a polarized expression of endogenous Lu antigens associated with a predominant expression of the Lu isoform at the basolateral domain of the plasma membrane and a very low expression of the Lu(v13) isoform at both the apical and basolateral domains. Analysis of transfected Madin-Darby canine kidney cells revealed a basolateral expression of Lu gp and a nonpolarized expression of Lu(v13) gp. Delivery of Lu(v13) to both apical and basolateral surfaces showed similar kinetics, indicating that this isoform is directly transported to each surface domain. A dileucine motif at position 608-609, specific to the Lu isoform, was characterized as a dominant basolateral sorting signal that prevents Lu gp from taking the apical delivery pathway. PMID- 10542218 TI - trans-Retinoic acid blocks platelet-derived growth factor-BB-induced expression of the murine monocyte chemoattractant-1 gene by blocking the assembly of a promoter proximal Sp1 binding site. AB - Proper regulation of the CC chemokine MCP-1 (monocyte chemoattractant protein-1) is important for normal inflammatory responses. MCP-1 is regulated by a wide variety of agents, including platelet-derived growth factor-BB (PDGF-BB) and tumor necrosis factor-alpha (TNF). Using both in vivo and in vitro assays, the elements required for expression between these two cytokines were compared. In vivo genomic footprinting showed that PDGF-BB induction occurred through the occupancy of the proximal regulatory region, and unlike TNF induction, no changes in the NF-kappaB binding, distal regulatory region occurred. Treatment of cells with trans-retinoic acid, an inhibitor of PDGF-BB activity, resulted in a 50% reduction in PDGF-BB-mediated induction and a concomitant block in the assembly of the proximal regulatory region. trans-Retinoic acid had minimal effect on TNF induction or promoter occupancy. An inhibitor of histone deacetylation was found to stimulate expression of MCP-1 in a manner that correlated with increased accessibility to the proximal regulatory region. These results show that the mechanisms of PDGF-BB and TNF activation of MCP-1 are distinct, although they both require the proximal regulatory region Sp1 binding site. The results also suggest that part of the mechanism used by both of these cytokines involves a process that regulates transcription factor access to the regulatory regions. PMID- 10542220 TI - Cloning of the human thiamine transporter, a member of the folate transporter family. AB - We have isolated a cDNA from human placenta, which, when expressed heterologously in mammalian cells, mediates the transport of the water-soluble vitamin thiamine. The cDNA codes for a protein of 497 amino acids containing 12 putative transmembrane domains. Northern blot analysis indicates that this transporter is widely expressed in human tissues. When expressed in HeLa cells, the cDNA induces the transport of thiamine (K(t) = 2.5 +/- 0.6 microM) in a Na(+)-independent manner. The cDNA-mediated transport of thiamine is stimulated by an outwardly directed H(+) gradient. Substrate specificity assays indicate that the transporter is specific to thiamine. Even though thiamine is an organic cation, the cDNA-induced thiamine transport is not inhibited by other organic cations. Similarly, thiamine is not a substrate for the known members of mammalian organic cation transporter family. The thiamine transporter gene, located on human chromosome 1q24, consists of 6 exons and is most likely the gene defective in the metabolic disorder, thiamine-responsive megaloblastic anemia. At the level of amino acid sequence, the thiamine transporter is most closely related to the reduced-folate transporter and thus represents the second member of the folate transporter family. PMID- 10542219 TI - Distinct roles for PP1 and PP2A in phosphorylation of the retinoblastoma protein. PP2a regulates the activities of G(1) cyclin-dependent kinases. AB - The function of the retinoblastoma protein (pRB) in controlling the G(1) to S transition is regulated by phosphorylation and dephosphorylation on serine and threonine residues. While the roles of cyclin-dependent kinases in phosphorylating and inactivating pRB have been characterized in detail, the roles of protein phosphatases in regulating the G(1)/S transition are not as well understood. We used cell-permeable inhibitors of protein phosphatases 1 and 2A to assess the contributions of these phosphatases in regulating cyclin-dependent kinase activity and pRB phosphorylation. Treating asynchronously growing Balb/c 3T3 cells with PP2A-selective concentrations of either okadaic acid or calyculin A caused a time- and dose-dependent decrease in pRB phosphorylation. Okadaic acid and calyculin A had no effect on pRB phosphatase activity even though PP2A was completely inhibited. The decrease in pRB phosphorylation correlated with inhibitor-induced suppression of G(1) cyclin-dependent kinases including CDK2, CDK4, and CDK6. The inhibitors also caused decreases in the levels of cyclin D2 and cyclin E, and induction of the cyclin-dependent kinase inhibitors p21(Cip1) and p27(Kip1). The decrease in cyclin-dependent kinase activities were not dependent on induction of cyclin-dependent kinase inhibitors since CDK inhibition still occurred in the presence of actinomycin D or cycloheximide. In contrast, selective inhibition of protein phosphatase 1 with tautomycin inhibited pRB phosphatase activity and maintained pRB in a highly phosphorylated state. The results show that protein phosphatase 1 and protein phosphatase 2A, or 2A-like phosphatases, play distinct roles in regulating pRB function. Protein phosphatase 1 is associated with the direct dephosphorylation of pRB while protein phosphatase 2A is involved in pathways regulating G(1) cyclin-dependent kinase activity. PMID- 10542221 TI - Vasoactive intestinal peptide and pituitary adenylate cyclase-activating polypeptide inhibit interleukin-12 transcription by regulating nuclear factor kappaB and Ets activation. AB - The vasoactive intestinal peptide (VIP) and the structurally related neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP) act as "macrophage deactivating factors". We showed previously that VIP and PACAP inhibit the production of macrophage-derived tumor necrosis factor-alpha, interleukin (IL)-6, nitric oxide, and IL-12. This study examines the molecular mechanisms involved in the VIP/PACAP inhibition of IL-12 production. VIP and PACAP inhibit IL-12 (p40) gene expression by affecting both NF-kappaB binding and the composition of the Ets-2 binding complex. Both neuropeptides prevent the activation-induced nuclear translocation of the NF-kappaB components p65 and c-Rel by inhibiting the reduction in cytoplasmic IkappaBalpha. Moreover, VIP and PACAP inhibit the synthesis of the interferon responsive factor-1. The decrease in nuclear interferon responsive factor-1 and c-Rel results in alterations of the Ets-2 binding complex. Two transduction pathways, a cAMP-dependent and a cAMP independent pathway, are involved in the inhibition of IL-12 gene expression and appear to differentially regulate the transcriptional factors involved. Because IL-12 participates in T cell activation and cytolytic T lymphocyte activity and promotes the differentiation of T helper cells into the Th1 subset, the understanding of the mechanisms that affect IL-12 production in normal and pathological conditions could contribute to immune response-based therapies or vaccine designs. PMID- 10542222 TI - A novel signaling intermediate, SHEP1, directly couples Eph receptors to R-Ras and Rap1A. AB - The Eph family of receptor tyrosine kinases has been implicated in many developmental patterning processes, including cell segregation, cell migration, and axon guidance. The cellular components involved in the signaling pathways of the Eph receptors, however, are incompletely characterized. Using a yeast two hybrid screen, we have identified a novel signaling intermediate, SHEP1 (SH2 domain-containing Eph receptor-binding protein 1), which is expressed in the embryonic and adult brain. SHEP1 contains an Src homology 2 domain that binds to a conserved tyrosine-phosphorylated motif in the juxtamembrane region of the EphB2 receptor and may itself be a target of EphB2 kinase activity, since it becomes heavily tyrosine-phosphorylated in cells expressing activated EphB2. SHEP1 also contains a domain similar to Ras guanine nucleotide exchange factor domains and binds to the GTPases R-Ras and Rap1A, but not Ha-Ras or RalA. Thus, SHEP1 directly links activated, tyrosine-phosphorylated Eph receptors to small Ras superfamily GTPases. PMID- 10542223 TI - Transcription initiation at the TATA-less spliced leader RNA gene promoter requires at least two DNA-binding proteins and a tripartite architecture that includes an initiator element. AB - Eukaryotic transcriptional regulatory signals, defined as core and activator promoter elements, have yet to be identified in the earliest diverging group of eukaryotes, the primitive protozoans, which include the Trypanosomatidae family of parasites. The divergence within this family is highlighted by the apparent absence of the "universal" transcription factor TATA-binding protein. To understand gene expression in these protists, we have investigated spliced leader RNA gene transcription. The RNA product of this gene provides an m(7)G cap and a 39-nucleotide leader sequence to all cellular mRNAs via a trans-splicing reaction. Regulation of spliced leader RNA synthesis is controlled by a tripartite promoter located exclusively upstream from the transcription start site. Proteins PBP-1 and PBP-2 bind to two of the three promoter elements in the trypanosomatid Leptomonas seymouri. They represent the first trypanosome transcription factors with typical double-stranded DNA binding site recognition. These proteins ensure efficient transcription. However, accurate initiation is determined an initiator element with a a loose consensus of CYAC/AYR (+1), which differs from that found in metazoan initiator elements as well as from that identified in one of the earliest diverging protozoans, Trichomonas vaginalis. Trypanosomes may utilize initiator element-protein interactions, and not TATA sequence-TATA-binding protein interactions, to direct proper transcription initiation by RNA polymerase II. PMID- 10542225 TI - Contractile activity modifies Fru-2,6-P(2) metabolism in rabbit fast twitch skeletal muscle. AB - Modification of muscular contractile patterns by denervation and chronic low frequency stimulation induces structural, physiological, and biochemical alterations in fast twitch skeletal muscles. Fructose 2,6-bisphosphate is a potent activator of 6-phosphofructo-1-kinase, a key regulatory enzyme of glycolysis in animal tissues. The concentration of Fru-2,6-P(2) depends on the activity of the bifunctional enzyme, 6-phosphofructo-2-kinase/fructose-2,6 bisphosphatase (PFK-2/FBPase-2), which catalyzes the synthesis and degradation of this metabolite. This enzyme has several isoforms, the relative abundance of which depends on the tissue metabolic properties. Skeletal muscle expresses two of these isoforms; it mainly contains the muscle isozyme (M-type) and a small amount of the liver isozyme (L-type), whose expression is under hormonal control. Moreover, contractile activity regulates expression of muscular proteins related with glucose metabolism. Fast twitch rabbit skeletal muscle denervation or chronic low frequency stimulation can provide information about the regulation of this enzyme. Our results show an increase in Fru-2,6-P(2) concentration after 2 days of denervation or stimulation. In denervated muscle, this increase is mediated by a rise in liver PFK-2/FBPase-2 isozyme, while in stimulated muscle it is mediated by a rise in muscle PFK-2/FBPase-2 isozyme. In conclusion, our results show that contractile activity could alter the expression of PFK-2/FBPase 2. PMID- 10542224 TI - The hydrophobic residues phenylalanine 184 and leucine 187 in the type-1 parathyroid hormone (PTH) receptor functionally interact with the amino-terminal portion of PTH-(1-34). AB - Recent mutagenesis and cross-linking studies suggest that three regions of the PTH-1 receptor play important roles in ligand interaction: (i) the extreme NH(2) terminal region, (ii) the juxtamembrane base of the amino-terminal extracellular domain, and (iii) the third extracellular loop. In this report, we analyzed the second of these segments in the rat PTH-1 receptor (residues 182-190) and its role in functional interaction with short PTH fragment analogs. Twenty-eight singly substituted PTH-1 receptors were transiently transfected into COS-7 cells and shown to be fully expressed by surface antibody binding analysis. Alanine scanning analysis identified Phe(184), Arg(186), Leu(187), and Ile(190) as important determinants of maximum binding of (125)I-labeled bovine PTH-(1-34) and (125)I-labeled bovine PTH-(3-34) and determinants of responsiveness to the NH(2) terminal analog, PTH-(1-14) in cAMP stimulation assays. Alanine mutations at these four sites augmented the ability of the COOH-terminal peptide [Glu(22), Trp(23)]PTHrP-(15-36) to inhibit the cAMP response induced by PTH-(1-34). At Phe(184) and Leu(187), hydrophobic substitutions (e.g. Ile, Met, or Leu) preserved PTH-(1-34)-mediated cAMP signaling potency, whereas hydrophilic substitutions (e.g. Asp, Glu, Lys, or Arg) weakened this response by 20-fold or more, as compared with the unsubstituted receptor's response. The results suggest that hydrophobicity at positions occupied by Phe(184) and Leu(187) in the PTH-1 receptor plays an important role in determining functional interaction with the 3 14 portion of PTH. PMID- 10542226 TI - Substrate diversity of herpes simplex virus thymidine kinase. Impact Of the kinematics of the enzyme. AB - Herpes simplex virus type 1 (HSV 1) thymidine kinase (TK) exhibits an extensive substrate diversity for nucleobases and sugar moieties, in contrast to other TKs. This substrate diversity is the crucial molecular basis of selective antiviral and suicide gene therapy. The mechanisms of substrate binding of HSV 1 TK were studied by means of site-directed mutagenesis combined with isothermal calorimetric measurements and guided by theoretical calculations and sequence comparison. The results show the link between the exceptionally broad substrate diversity of HSV 1 TK and the presence of structural features such as the residue triad His-58/Met-128/Tyr-172. The mutation of Met-128 into a Phe and the double mutant M128F/Y172F result in mutants that have lost their activity. However, by exchanging His to form the triple mutant H58L/M128F/Y172F, the enzyme regains activity. Strikingly, this triple mutant becomes resistant toward acyclovir. Furthermore, we give evidence for the importance of Glu-225 of the flexible LID region for the catalytic reaction. The data presented give new insights to understand mechanisms ruling substrate diversity and thus are crucial for both the development of new antiviral drugs and engineering of mutant TKs apt to accept novel substrate analogs for gene therapeutic approaches. PMID- 10542227 TI - 2-Keto-3-deoxy-D-glycero-D-galacto-nononic acid (KDN)- and N-acetylneuraminic acid-cleaving sialidase (KDN-sialidase) and KDN-cleaving hydrolase (KDNase) from the hepatopancreas of oyster, Crassostrea virginica. AB - KDN (2-keto-3-deoxy-D-glycero-D-galacto-nononic acid), a sialic acid analog, has been found to be widely distributed in nature. Despite the structural similarity between KDN and Neu5Ac, alpha-ketosides of KDN are refractory to conventional sialidases. We found that the hepatopancreas of the oyster, Crassostrea virginica, contains two KDN-cleaving sialidases but is devoid of conventional sialidase. The major sialidase, KDN-sialidase, effectively cleaves alpha ketosidically linked KDN and also slowly cleaves the alpha-ketosides of Neu5Ac. The minor sialidase, KDNase, is specific for alpha-ketosides of KDN. We were able to separate these two KDN-cleaving enzymes using hydrophobic interaction and cation-exchange chromatographies. The rate of hydrolysis of 4-methylumbelliferyl alpha-KDN (MU-KDN) by KDN-sialidase is 30 times faster than that of MU-Neu5Ac in the presence of 0.2 M NaCl, whereas in the absence of NaCl this ratio is only 8. KDNase hydrolyzes MU-KDN over 500 times faster than MU-Neu5Ac and is not affected by NaCl. KDN-sialidase purified to electrophoretically homogeneous form was found to have a molecular mass of 25 kDa and an isoelectric point of 8.4. One of the three tryptic peptides derived from KDN-sialidase contains the consensus motif, SXDXGXTW, that has been found in all conventional sialidases. Kinetic analysis of the inhibition of the hydrolysis of MU-KDN and MU-Neu5Ac by 2, 3-dehydro-2-deoxy KDN (KDN2-en) and 2,3-dehydro-2-deoxy-(Neu5Ac2-en) suggests that KDN-sialidase contains two separate active sites for the hydrolysis of KDN and Neu5Ac. Both KDN sialidase and KDNase effectively hydrolyze KDN-G(M3), KDNalpha2-->3Gal beta1- >4Glc, KDNalpha2-->6Galbeta1-->4Glc, KDNalpha2-->6-N-acetylgalactosaminitol, KDNalpha2-->6(KDNalpha2-->3)N-acetylgalactosaminitol, and KDNalpha2- >6(GlcNAcbeta1-->3)N-acetylgalactosaminitol. However, only KDN-sialidase also slowly hydrolyzes G(M3), Neu5Acalpha2-->3Galbeta1-->4Glc, and Neu5Acalpha2- >6Galbeta1-->4Glc. These two KDN-cleaving sialidases should be useful for studying the structure and function of KDN-containing glycoconjugates. PMID- 10542228 TI - The role of DOC-2/DAB2 protein phosphorylation in the inhibition of AP-1 activity. An underlying mechanism of its tumor-suppressive function in prostate cancer. AB - DOC-2/DAB2, a novel phosphoprotein with signal-transducing capability, inhibits human prostatic cancer cells (Tseng, C.-P., Ely, B. D., Li, Y., Pong, R.-C., and Hsieh, J.-T. (1998) Endocrinology 139, 3542-3553). However, its mechanism of action is not understood completely. This study delineates the functional significance of DOC-2/DAB2 protein phosphorylation and demonstrates that in vivo activation of protein kinase C (PKC) by 12-O-tetradecanoylphorbol-13-acetate (TPA) induces DOC-2/DAB2 phosphorylation, including a serine residue at position 24. Mutation of Ser(24) to Ala reduced DOC-2/DAB2 phosphorylation by PKC. Using a synthetic Ser(24) peptide (APS(24)KKEKKKGSEKTD) or recombinant DOC-2/DAB2 as substrates, PKCbetaII, PKCgamma, and PKCdelta (but not casein kinase II) directly phosphorylated Ser(24) in vitro. This indicates that DOC-2/DAB2 is a PKC-specific substrate. Since expression of wild-type DOC-2/DAB2, but not the S24A mutant, inhibited TPA-induced AP-1 activity in prostatic epithelial cells, phosphorylation of Ser(24) appears to play a critical role in modulating TPA induced AP-1 activity. Taken together, these data suggest that PKC-regulated phosphorylation of DOC-2/DAB2 protein may help its growth inhibitory function. PMID- 10542229 TI - Deprotonation of the 33-kDa, extrinsic, manganese-stabilizing subunit accompanies photooxidation of manganese in photosystem II. AB - Photosystem II catalyzes photosynthetic water oxidation. The oxidation of water to molecular oxygen requires four sequential oxidations; the sequentially oxidized forms of the catalytic site are called the S states. An extrinsic subunit, the manganese-stabilizing protein (MSP), promotes the efficient turnover of the S states. MSP can be removed and rebound to the reaction center; removal and reconstitution is associated with a decrease in and then a restoration of enzymatic activity. We have isotopically edited MSP by uniform (13)C labeling of the Escherichia coli-expressed protein and have obtained the Fourier transform infrared spectrum associated with the S(1) to S(2) transition in the presence either of reconstituted (12)C or (13)C MSP. (13)C labeling of MSP is shown to cause 30-60 cm(-1) shifts in a subset of vibrational lines. The derived, isotope edited vibrational spectrum is consistent with a deprotonation of glutamic/aspartic acid residues on MSP during the S(1) to S(2) transition; the base, which accepts this proton(s), is not located on MSP. This finding suggests that this subunit plays a role as a stabilizer of a charged transition state and, perhaps, as a general acid/base catalyst of oxygen evolution. These results provide a molecular explanation for known MSP effects on oxygen evolution. PMID- 10542230 TI - Helix 4 of the Bacillus thuringiensis Cry1Aa toxin lines the lumen of the ion channel. AB - The mode of action of Bacillus thuringiensis insecticidal proteins is not well understood. Based on analogies with other bacterial toxins and ion channels, we hypothesized that charged amino acids in helix 4 of the Cry1Aa toxin are critical for toxicity and ion channel function. Using Plutella xylostella as a model target, we analyzed responses to Cry1Aa and eight proteins with altered helix 4 residues. Toxicity was abolished in five charged residue mutants (E129K, R131Q, R131D, D136N, D136C), however, two charged (R127E and R127N) and one polar (N138C) residue mutant retained wild-type toxicity. Compared with Cry1Aa and toxic mutants, nontoxic mutants did not show greatly reduced binding to brush border membrane vesicles, but their ion channel conductance was greatly reduced in planar lipid bilayers. Substituted cysteine accessibility tests showed that in situ restoration of the negative charge of D136C restored conductance to wild type levels. The results imply that charged amino acids on the Asp-136 side of helix 4 are essential for toxicity and passage of ions through the channel. These results also support a refined version of the umbrella model of membrane integration in which the side of helix 4 containing Asp-136 faces the aqueous lumen of the ion channel. PMID- 10542231 TI - The SH3 domains of endophilin and amphiphysin bind to the proline-rich region of synaptojanin 1 at distinct sites that display an unconventional binding specificity. AB - The proline-rich domain of synaptojanin 1, a synaptic protein with phosphatidylinositol phosphatase activity, binds to amphiphysin and to a family of recently discovered proteins known as the SH3p4/8/13, the SH3-GL, or the endophilin family. These interactions are mediated by SH3 domains and are believed to play a regulatory role in synaptic vesicle recycling. We have precisely mapped the target peptides on human synaptojanin that are recognized by the SH3 domains of endophilins and amphiphysin and proven that they are distinct. By a combination of different approaches, selection of phage displayed peptide libraries, substitution analyses of peptides synthesized on cellulose membranes, and a peptide scan spanning a 252-residue long synaptojanin fragment, we have concluded that amphiphysin binds to two sites, PIRPSR and PTIPPR, whereas endophilin has a distinct preferred binding site, PKRPPPPR. The comparison of the results obtained by phage display and substitution analysis permitted the identification of proline and arginine at positions 4 and 6 in the PIRPSR and PTIPPR target sequence as the major determinants of the recognition specificity mediated by the SH3 domain of amphiphysin 1. More complex is the structural rationalization of the preferred endophilin ligands where SH3 binding cannot be easily interpreted in the framework of the "classical" type I or type II SH3 binding models. Our results suggest that the binding repertoire of SH3 domains may be more complex than originally predicted. PMID- 10542232 TI - Activation of transcription by estrogen receptor alpha and beta is cell type- and promoter-dependent. AB - Tamoxifen acts as a strong estrogen antagonist in human breast but as an estrogen agonist in the uterus. The action of tamoxifen is mediated through estrogen receptors (ERalpha and ERbeta), which bind to a variety of responsive elements, to activate transcription. To examine the role of these varied elements in the response to antiestrogens, we studied the activation of a panel of differing promoters, by these compounds, in human breast, bone, and endometrial derived cell lines. No agonistic activity was observed in breast cells, whereas all antiestrogens, particularly tamoxifen, exhibited agonistic effects in uterine cell lines. All antiestrogens studied were agonistic in co-transfections of a collagenase reporter gene and ERbeta, but tamoxifen alone was agonistic with ERalpha in (uterine) HEC-1-A cells. The ERalpha mediated, agonism of tamoxifen was not observed in primary cultures of human uterine stromal cells, whereas the ERbeta-mediated agonism of all selective estrogen receptor modulators was present. This suggests that the two receptors operate by distinct pathways and that the response of cells to antiestrogens is dependent on the ER subtypes expressed. PMID- 10542233 TI - Regulation of a cytosolic and nuclear O-GlcNAc transferase. Role of the tetratricopeptide repeats. AB - The O-GlcNAc transferase (OGT) is a unique nuclear and cytosolic glycosyltransferase that contains multiple tetratricopeptide repeats. We have begun to characterize the mechanisms regulating OGT using a combination of deletion analysis and kinetic studies. Here we show that the p110 subunit of the enzyme forms both homo- and heterotrimers that appear to have different binding affinities for UDP-GlcNAc. The multimerization domain of OGT lies within the tetratricopeptide repeat domain and is not necessary for activity. Kinetic analyses of the full-length trimer and the truncated monomer forms of OGT suggest that both forms function through a random bi-bi kinetic mechanism. Both the monomer and trimer have similar specific activities and similar K(m) values for peptide substrates. However, they differ in their binding affinities for UDP GlcNAc, indicating that subunit interactions affect enzyme activity. The findings that recombinant OGT has three distinct K(m) values for UDP-GlcNAc and that UDP GlcNAc concentrations modulates the affinity of OGT for peptides suggest that OGT is exquisitely regulated by the levels of UDP-GlcNAc within the nucleus and cytoplasm. PMID- 10542234 TI - Decreased capacity of recombinant 45/47-kDa molecules (Apa) of Mycobacterium tuberculosis to stimulate T lymphocyte responses related to changes in their mannosylation pattern. AB - The Apa molecules secreted by Mycobacterium tuberculosis, Mycobacterium bovis, or BCG have been identified as major immunodominant antigens. Mass spectrometry analysis indicated similar mannosylation, a complete pattern from 1 up to 9 hexose residues/mole of protein, of the native species from the 3 reference strains. The recombinant antigen expressed in M. smegmatis revealed a different mannosylation pattern: species containing 7 to 9 sugar residues/mole of protein were in the highest proportion, whereas species bearing a low number of sugar residues were almost absent. The 45/47-kDa recombinant antigen expressed in E. coli was devoid of sugar residues. The proteins purified from M. tuberculosis, M. bovis, or BCG have a high capacity to elicit in vivo potent delayed-type hypersensitivity (DTH) reactions and to stimulate in vitro sensitized T lymphocytes of guinea pigs immunized with living BCG. The recombinant Apa expressed in Mycobacterium smegmatis was 4-fold less potent in vivo in the DTH assay and 10-fold less active in vitro to stimulate sensitized T lymphocytes than the native proteins. The recombinant protein expressed in Escherichia coli was nearly unable to elicit DTH reactions in vivo or to stimulate T lymphocytes in vitro. Thus the observed biological effects were related to the extent of glycosylation of the antigen. PMID- 10542235 TI - Cloning and characterization of PBP 1C, a third member of the multimodular class A penicillin-binding proteins of Escherichia coli. AB - All proteins of Escherichia coli that covalently bind penicillin have been cloned except for the penicillin-binding protein (PBP) 1C. For a detailed understanding of the mode of action of beta-lactam antibiotics, cloning of the gene encoding PBP1C was of major importance. Therefore, the structural gene was identified in the E. coli genomic lambda library of Kohara and subcloned, and PBP1C was characterized biochemically. PBP1C is a close homologue to the bifunctional transpeptidases/transglycosylases PBP1A and PBP1B and likewise shows murein polymerizing activity, which can be blocked by the transglycosylase inhibitor moenomycin. Covalently linked to activated Sepharose, PBP1C specifically retained PBP1B and the transpeptidases PBP2 and -3 in addition to the murein hydrolase MltA. The specific interaction with these proteins suggests that PBP1C is assembled into a multienzyme complex consisting of both murein polymerases and hydrolases. Overexpression of PBP1C does not support growth of a PBP1A(ts)/PBP1B double mutant at the restrictive temperature, and PBP1C does not bind to the same variety of penicillin derivatives as PBPs 1A and 1B. Deletion of PBP1C resulted in an altered mode of murein synthesis. It is suggested that PBP1C functions in vivo as a transglycosylase only. PMID- 10542236 TI - Deoxyhypusine synthase from tobacco. cDNA isolation, characterization, and bacterial expression of an enzyme with extended substrate specificity. AB - Deoxyhypusine synthase catalyzes the formation of a deoxyhypusine residue in the translation eukaryotic initiation factor 5A (eIF5A) precursor protein by transferring an aminobutyl moiety from spermidine onto a conserved lysine residue within the eIF5A polypeptide chain. This reaction commences the activation of the initiation factor in fungi and vertebrates. A mechanistically identical reaction is known in the biosynthetic pathway leading to pyrrolizidine alkaloids in plants. Deoxyhypusine synthase from tobacco was cloned and expressed in active form in Escherichia coli. It catalyzes the formation of a deoxyhypusine residue in the tobacco eIF5A substrate as shown by gas chromatography coupled with a mass spectrometer. The enzyme also accepts free putrescine as the aminobutyl acceptor, instead of lysine bound in the eIF5A polypeptide chain, yielding homospermidine. Conversely, it accepts homospermidine instead of spermidine as the aminobutyl donor, whereby the reactions with putrescine and homospermidine proceed at the same rate as those involving the authentic substrates. The conversion of deoxyhypusine synthase-catalyzed eIF5A deoxyhypusinylation pinpoints a function for spermidine in plant metabolism. Furthermore, and quite unexpectedly, the substrate spectrum of deoxyhypusine synthase hints at a biochemical basis behind the sparse and skew occurrence of both homospermidine and its pyrrolizidine derivatives across distantly related plant taxa. PMID- 10542237 TI - Peroxisome proliferator-activated receptor alpha negatively regulates the vascular inflammatory gene response by negative cross-talk with transcription factors NF-kappaB and AP-1. AB - Interleukin-6 (IL-6) is a pleiotropic cytokine, whose plasma levels are elevated in inflammatory diseases such as atherosclerosis. We have previously reported that peroxisome proliferator-activated receptor alpha (PPARalpha) ligands (fibrates) lower elevated plasma concentrations of IL-6 in patients with atherosclerosis and inhibit IL-1-stimulated IL-6 secretion by human aortic smooth muscle cells (SMC). Here, we show that aortic explants isolated from PPARalpha null mice display an exacerbated response to inflammatory stimuli, such as lipopolysaccharide (LPS), as demonstrated by increased IL-6 secretion. Furthermore, fibrate treatment represses IL-6 mRNA levels in LPS-stimulated aortas of PPARalpha wild-type, but not of PPARalpha-null mice, demonstrating a role for PPARalpha in this fibrate action. In human aortic SMC, fibrates inhibit IL-1-induced IL-6 gene expression. Furthermore, activation of PPARalpha represses both c-Jun- and p65-induced transcription of the human IL-6 promoter. Transcriptional interference between PPARalpha and both c-Jun and p65 occurs reciprocally, since c-Jun and p65 also inhibit PPARalpha-mediated activation of a PPAR response element-driven promoter. This transcriptional interference occurs independent of the promoter context as demonstrated by cotransfection experiments using PPARalpha, p65, and c-Jun Gal4 chimeras. Overexpression of the transcriptional coactivator cAMP-responsive element-binding protein-binding protein (CBP) does not relieve PPARalpha-mediated transcriptional repression of p65 and c-Jun. Finally, glutathione S-transferase pull-down experiments demonstrate that PPARalpha physically interacts with c-Jun, p65, and CBP. Altogether these data indicate that fibrates inhibit the vascular inflammatory response via PPARalpha by interfering with the NF-kappaB and AP-1 transactivation capacity involving direct protein-protein interaction with p65 and c-Jun. PMID- 10542238 TI - Role of the first extracellular loop in the functional activation of CCR2. The first extracellular loop contains distinct domains necessary for both agonist binding and transmembrane signaling. AB - The physiological cellular responses to monocyte chemoattractant protein-1 (MCP 1), a potent chemotactic and activating factor for mononuclear leukocytes, are mediated by specific binding to CCR2. The aim of this investigation is to identify receptor microdomains that are involved in high affinity agonist binding and receptor activation. The results from our functional studies in which we utilized neutralizing antisera against CCR2 are consistent with a multidomain binding model, previously proposed by others. The first extracellular loop was of particular interest, because in addition to a ligand-binding domain it contained also information for receptor activation, crucial for transmembrane signaling. Replacement of the first extracellular loop of CCR2 with the corresponding region of CCR1 decreased the MCP-1 binding affinity about 10-fold and prevented transmembrane signaling. A more detailed analysis by site-directed mutagenesis revealed that this receptor segment contains two distinct microdomains. The amino acid residues Asn(104) and Glu(105) are essential for high affinity agonist binding but are not involved in receptor activation. In contrast, the charged amino acid residue His(100) does not contribute to ligand binding but is vital for receptor activation and initiation of transmembrane signaling. We hypothesize that the interaction of agonist with this residue initiates the conformational switch that allows the formation of the functional CCR2-G protein complex. PMID- 10542239 TI - Identification of inhibitory autophosphorylation sites in casein kinase I epsilon. AB - Casein kinase I epsilon (CKIepsilon) is a widely expressed protein kinase implicated in the regulation of diverse cellular processes including DNA replication and repair, nuclear trafficking, and circadian rhythm. CKIepsilon and the closely related CKIdelta are regulated in part through autophosphorylation of their carboxyl-terminal extensions, resulting in down-regulation of enzyme activity. Treatment of CKIepsilon with any of several serine/threonine phosphatases causes a marked increase in kinase activity that is self-limited. To identify the sites of inhibitory autophosphorylation, a series of carboxyl terminal deletion mutants was constructed by site-directed mutagenesis. Truncations that eliminated specific phosphopeptides present in the wild-type kinase were used to guide construction of specific serine/threonine to alanine mutants. Amino acids Ser-323, Thr-325, Thr-334, Thr-337, Ser-368, Ser-405, Thr 407, and Ser-408 in the carboxyl-terminal tail of CKIepsilon were identified as probable in vivo autophosphorylation sites. A recombinant CKIepsilon protein with serine and threonine to alanine mutations eliminating these autophosphorylation sites was 8-fold more active than wild-type CKIepsilon using IkappaBalpha as a substrate. The identified autophosphorylation sites do not conform to CKI substrate motifs identified in peptide substrates. PMID- 10542240 TI - Intramolecular backfolding of the carboxyl-terminal end of MxA protein is a prerequisite for its oligomerization. AB - Mx proteins are large GTPases, which play a pivotal role in the interferon type I mediated response against viral infections. The human MxA inhibits the replication of several RNA viruses and is organized in oligomeric structures. Using two different experimental approaches, the mammalian two-hybrid system and an interaction dependent nuclear translocation approach, three domains in the carboxyl-terminal moiety were identified that are involved in the oligomerization of MxA. The first consists of a carboxyl-terminal amphipathic helix (LZ1), which binds to a more proximal part of the same molecule. This intramolecular backfolding is a prerequisite for the formation of an intermolecular complex. This intermolecular interaction is mediated by two domains, a poorly defined region generated by the intramolecular interaction and a domain located between amino acids 363 and 415. Co-expression of wild-type MxA with various mutant fragments thereof revealed that the presence of the carboxyl-terminal region comprising the amphipathic helices LZ1 and LZ2 is necessary and sufficient to exert a dominant negative effect. This finding suggests that the functional interference of the carboxyl-terminal region is due to competition for binding of an as yet unidentified cellular or viral target molecules. PMID- 10542241 TI - 3-Formyl-1-butyl pyrophosphate A novel mycobacterial metabolite-activating human gammadelta T cells. AB - Most human blood gammadelta T cells react without major histocompatibility complex restriction to small phosphorylated nonpeptide antigens (phosphoantigens) that are abundantly produced by mycobacteria and several other microbial pathogens. Although isopentenyl pyrophosphate has been identified as a mycobacterial antigen for gammadelta T cells, the structure of several other stimulating compounds with bioactivities around 1000-fold higher than isopentenyl pyrophosphate remains to be elucidated. This paper describes the structural identification of 3-formyl-1-butyl-pyrophosphate as the core of several non prenyl mycobacterial phosphoantigens bioactive at the nM range. Recognition of this molecule by gammadelta T cells is very selective and relies on its aldehyde and pyrophosphate groups. This novel pyrophosphorylated aldehyde most probably corresponds to a metabolic intermediate of the non-mevalonate pathway of prenyl phosphate biosynthesis in eubacteria and algae. The reactivity to 3-formyl-1 butyl-pyrophosphate supports the view that human gammadelta T cells are physiologically devoted to antimicrobial surveillance. PMID- 10542242 TI - Inactivation of p42 mitogen-activated protein kinase is required for exit from M phase after cyclin destruction. AB - By using cycling Xenopus egg extracts, we have previously found that if mitogen activated protein kinase (p42 MAPK) is activated on entry into mitosis (M-phase), the extract is arrested with condensed chromosomes and spindle microtubules. Here we show that these arrested extracts have high levels of M-phase promoting factor (MPF, Cyclin B/Cdc2) activity, stabilized levels of Cyclin B, and sustained M phase-specific phosphorylations. We also examined the role of p42 MAPK in DNA damage checkpoint-arrested extracts that were induced to enter M-phase by the addition of Cdc25C protein. In these extracts, Cdc25C protein triggers the abrupt, premature activation of MPF and entry into M-phase. MPF activity then drops suddenly due to Cyclin B proteolysis, just as p42 MAPK is activated. Unexpectedly, however, M-phase is sustained, as judged by maintenance of M-phase specific phosphorylations and condensed chromosomes. To determine if this M-phase arrest depended on p42 MAPK activation, we added PD98059 (PD), an inhibitor of p42 MAPK activation, to egg extracts with exogenous Cdc25. Both untreated and PD treated extracts entered M-phase simultaneously, with a sharp peak of MPF activity. However, only PD-treated extracts subsequently exited from M-phase and entered interphase. In PD-treated extracts, p42 MAPK was not activated, and the transition to interphase was accompanied by the formation of decondensed nuclei and the disappearance of M-phase-specific phosphorylation of proteins. These results show that although entry into M-phase requires the activation of MPF, exit from M-phase even after cyclin destruction, is dependent on the inactivation of p42 MAPK. PMID- 10542243 TI - The nuclear factor-kappaB engages CBP/p300 and histone acetyltransferase activity for transcriptional activation of the interleukin-6 gene promoter. AB - Expression of the pleiotropic cytokine interleukin (IL)-6 can be stimulated by the proinflammatory cytokine tumor necrosis factor (TNF) and the microbial alkaloid staurosporine (STS). In this report, the transcriptional mechanisms were thoroughly investigated. Whereas transcription factors binding to the activator protein-1-, cAMP-responsive element-, and CAAT enhancer-binding protein responsive sequences are necessary for gene activation by STS, nuclear factor (NF)-kappaB alone is responsible and sufficient for inducibility by TNF, which reveals distinct signaling pathways for both compounds. At the cofactor level, cAMP-responsive element-binding protein-binding protein (CBP) or p300 potentiate basal and induced IL-6 promoter activation via multiple protein-protein interactions with all transcription factors bound to the promoter DNA. However, the strongest promoter activation relies on the p65 NF-kappaB subunit, which specifically engages CBP/p300 for maximal transcriptional stimulation by its histone acetyltransferase activity. Moreover, treatment of chromatin-integrated promoter constructions with the histone deacetylase inhibitor trichostatin A exclusively potentiates TNF-dependent (i.e. NF-kappaB-mediated) gene activation, while basal or STS-stimulated IL-6 promoter activity remains completely unchanged. Similar observations were recorded with other natural NF-kappaB-driven promoters, namely IL-8 and endothelial leukocyte adhesion molecule (ELAM). We conclude that, within an "enhanceosome-like" structure, NF-kappaB is the central mediator of TNF-induced IL-6 gene expression, involving CBP/p300 and requiring histone acetyltransferase activity. PMID- 10542245 TI - Interaction between collagen and the alpha(2) I-domain of integrin alpha(2)beta(1). Critical role of conserved residues in the metal ion-dependent adhesion site (MIDAS) region. AB - A docking model of the alpha(2) I-domain and collagen has been proposed based on their crystal structures (Emsley, J., King, S., Bergelson, J., and Liddington, R. C. (1997) J. Biol. Chem. 272, 28512-28517). In this model, several amino acid residues in the I-domain make direct contact with collagen (Asn-154, Asp-219, Leu 220, Glu-256, His-258, Tyr-285, Asn-289, Leu-291, Asn-295, and Lys-298), and the protruding C-helix of alpha(2) (residues 284-288) determines ligand specificity. Because most of the proposed critical residues are not conserved, different I domains are predicted to bind to collagen differently. We found that deleting the entire C-helix or mutating the predicted critical residues had no effect on collagen binding to whole alpha(2)beta(1), with the exception that mutating Asn 154, Asp-219, and His-258 had a moderate effect. We performed further studies and found that mutating the conserved surface-exposed residues in the metal ion dependent adhesion site (MIDAS) (Tyr-157 and Gln-215) significantly blocks collagen binding. We have revised the docking model based on the mutagenesis data. In the revised model, conserved Tyr-157 makes contact with collagen in addition to the previously proposed Asn-154, Asp-219, His-258, and Tyr-285 residues. These results suggest that the collagen-binding I-domains (e.g. alpha(1), alpha(2), and alpha(10)) bind to collagen in a similar fashion. PMID- 10542244 TI - Mechanisms of transcriptional activation of bcl-2 gene expression by 17beta estradiol in breast cancer cells. AB - bcl-2 gene expression is induced by 17beta-estradiol (E2) in T47D and MCF-7 human breast cancer cells, and the mechanism of E2 responsiveness was further investigated by analysis of the bcl-2 gene promoter. The -1602 to -1534 distal region (bcl-2j) of the promoter was E2-responsive; however, in gel mobility shift assays, the estrogen receptor alpha (ER(alpha)) did not bind [(32)P]bcl-2j, whereas Sp1 protein formed a retarded band complex. Further analysis demonstrated that the upstream region (-1603 to -1579) of the bcl-2 gene promoter contained two GC/GA-rich sites at -1601 (5'-GGGCTGG-3') and -1588 (3'-GGAGGG-5') that bound Sp1 protein. Subsequent studies confirmed that transactivation by E2 was dependent on ER(alpha)/Sp1 interactions with both GC-rich sites, and this was confirmed by in vitro footprinting. In contrast, a 21-base pair E2-responsive downstream region (-1578 to -1534) did not bind Sp1 or ER(alpha) protein; however, analysis of a complex binding pattern with nuclear extracts showed that ATF-1 and CREB-1 bound to this motif. These data coupled with results of transient transfection studies demonstrated that transcriptional activation by E2 of the -1578 to -1534 region of the bcl-2 gene promoter was dependent on induction of cAMP and subsequent activation through a cAMP response element. Thus, hormone regulation of bcl-2 gene expression in breast cancer cells involves multiple enhancer elements and E2-mediated transactivation does not require direct binding of the estrogen receptor with promoter DNA. PMID- 10542246 TI - Unique cellular events occurring during the initial interaction of macrophages with matrix-retained or methylated aggregated low density lipoprotein (LDL). Prolonged cell-surface contact during which ldl-cholesteryl ester hydrolysis exceeds ldl protein degradation. AB - A critical event in atherogenesis is the interaction of arterial wall macrophages with subendothelial lipoproteins. Although most studies have investigated this interaction by incubating cultured macrophages with monomeric lipoproteins dissolved in media, arterial wall macrophages encounter lipoproteins that are mostly bound to subendothelial extracellular matrix, and these lipoproteins are often aggregated or fused. Herein, we utilize a specialized cell-culture system to study the initial interaction of macrophages with aggregated low density lipoprotein (LDL) bound to extracellular matrix. The aggregated LDL remains extracellular for a relatively prolonged period of time and becomes lodged in invaginations in the surface of the macrophages. As expected, the degradation of the protein moiety of the LDL was very slow. Remarkably, however, hydrolysis of the cholesteryl ester (CE) moiety of the LDL was 3-7-fold higher than that of the protein moiety, in stark contrast to the situation with receptor-mediated endocytosis of acetyl-LDL. Similar results were obtained using another experimental system in which the degradation of aggregated LDL protein was delayed by LDL methylation rather than by retention on matrix. Additional experiments indicated the following properties of this interaction: (a) LDL-CE hydrolysis is catalyzed by lysosomal acid lipase; (b) neither scavenger receptors nor the LDL receptor appear necessary for the excess LDL-CE hydrolysis; and (c) LDL-CE hydrolysis in this system is resistant to cellular potassium depletion, which further distinguishes this process from receptor-mediated endocytosis. In summary, experimental systems specifically designed to mimic the in vivo interaction of arterial wall macrophages with subendothelial lipoproteins have demonstrated an initial period of prolonged cell-surface contact in which CE hydrolysis exceeds protein degradation. PMID- 10542247 TI - Selective loss of poly(ADP-ribose) and the 85-kDa fragment of poly(ADP-ribose) polymerase in nucleoli during alkylation-induced apoptosis of HeLa cells. AB - Alkylation treatment of HeLa cells results in the rapid induction of apoptosis as revealed by DNA laddering and cleavage of poly(ADP-ribose) polymerase (PARP) into the 29-and 85-kDa fragments (Kumari S. R., Mendoza-Alvarez, H. & Alvarez Gonzalez, R. (1998) Cancer Res. 58, 5075-5078). Here, we performed a time-course analysis of (i) poly(ADP-ribose) synthesis and degradation as well as (ii) the subnuclear localization of PARP and its fragments by using confocal laser scanning immunofluorescence microscopy. PARP was activated within 15 min post treatment, as revealed by nuclear immunostaining with antibody 10H (recognizing poly(ADP-ribose)). This was followed by a late, time-dependent, progressive decline of 10H signals that coincide with the time of PARP cleavage. Strikingly, nucleolar immunostaining with antibodies 10H and C-II-10 (recognizing the 85-kDa PARP fragment) was lost by 15 min post-treatment, whereas F-I-23 signals (recognizing the 29-kDa fragment) persisted. We hypothesize that the 85-kDa PARP fragment is translocated, along with covalently bound poly(ADP-ribose), from nucleoli to the nucleoplasm, whereas the 29-kDa fragment is retained, because it binds to DNA strand breaks. Our data (i) provide a link between the known time dependent bifunctional role of PARP in apoptosis and the subcellular localization of PARP fragments and also (ii) add to the evidence for early proteolytic changes in nucleoli during apoptosis. PMID- 10542248 TI - Biosynthesis of vascular endothelial growth factor-D involves proteolytic processing which generates non-covalent homodimers. AB - Vascular endothelial growth factor-D (VEGF-D) binds and activates the endothelial cell tyrosine kinase receptors VEGF receptor-2 (VEGFR-2) and VEGF receptor-3 (VEGFR-3), is mitogenic for endothelial cells, and shares structural homology and receptor specificity with VEGF-C. The primary translation product of VEGF-D has long N- and C-terminal polypeptide extensions in addition to a central VEGF homology domain (VHD). The VHD of VEGF-D is sufficient to bind and activate VEGFR 2 and VEGFR-3. Here we report that VEGF-D is proteolytically processed to release the VHD. Studies in 293EBNA cells demonstrated that VEGF-D undergoes N- and C terminal cleavage events to produce numerous secreted polypeptides including a fully processed form of M(r) approximately 21,000 consisting only of the VHD, which is predominantly a non-covalent dimer. Biosensor analysis demonstrated that the VHD has approximately 290- and approximately 40-fold greater affinity for VEGFR-2 and VEGFR-3, respectively, compared with unprocessed VEGF-D. In situ hybridization demonstrated that embryonic lung is a major site of expression of the VEGF-D gene. Processed forms of VEGF-D were detected in embryonic lung indicating that VEGF-D is proteolytically processed in vivo. PMID- 10542250 TI - SPARC regulates the expression of collagen type I and transforming growth factor beta1 in mesangial cells. AB - The matricellular protein SPARC is expressed at high levels in cells that participate in tissue remodeling and is thought to regulate mesangial cell proliferation and extracellular matrix production in the kidney glomerulus in a rat model of glomerulonephritis (Pichler, R. H., Bassuk, J. A., Hugo, C., Reed, M. J., Eng, E., Gordon, K. L., Pippin, J., Alpers, C. E., Couser, W. G., Sage, E. H., and Johnson, R. J. (1997) Am. J. Pathol. 148, 1153-1167). A potential mechanism by which SPARC controls both cell cycle and matrix production has been attributed to its regulation of a pleiotropic growth factor. In this study we used primary mesangial cell cultures from wild-type mice and from mice with a targeted disruption of the SPARC gene. SPARC-null cells displayed diminished expression of collagen type I mRNA and protein, relative to wild-type cells, by the criteria of immunocytochemistry, immunoblotting, and the reverse transcription-polymerase chain reaction. The SPARC-null cells also showed significantly decreased steady-state levels of transforming growth factor-beta1 (TGF-beta1) mRNA and secreted TGF-beta1 protein. Addition of recombinant SPARC to SPARC-null cells restored the expression of collagen type I mRNA to 70% and TGF beta1 mRNA to 100% of wild-type levels. We conclude that SPARC regulates the expression of collagen type I and TGF-beta1 in kidney mesangial cells. Since increased mitosis and matrix deposition by mesangial cells are characteristics of glomerulopathies, we propose that SPARC is one of the factors that maintains the balance between cell proliferation and matrix production in the glomerulus. PMID- 10542249 TI - p53 represses CAAT enhancer-binding protein (C/EBP)-dependent transcription of the albumin gene. A molecular mechanism involved in viral liver infection with implications for hepatocarcinogenesis. AB - p53 is a transcription factor that is activated by genotoxic stress and mediates cell cycle arrest and apoptosis. Here we demonstrate that infection of mouse liver with recombinant E1/E3-deleted adenovirus leads to p53 activation and simultaneously to the down-regulation of albumin gene expression. In vitro transcription assays indicate that transcriptional mechanisms mediated through the albumin promoter are responsible for reduced albumin mRNA levels during viral infection. Albumin expression is maintained in the liver by a combination of liver-enriched transcription factors such as CAAT enhancer-binding protein (C/EBP)alpha and C/EBPbeta. We show that p53 wild type and tumor-derived p53 mutations repress C/EBP-mediated transactivation of the albumin promoter. The binding of C/EBPalpha or -beta to its cognate sequence in the albumin promoter is not inhibited by p53 expression. Deletion analysis and domain swapping experiments show that repression of C/EBPbeta-mediated transactivation is dependent on the N-terminal domain of p53 and the transactivation domain, leucine zipper domain, and the inhibitory domain II (amino acids 163-191) of C/EBPbeta. Our results provide a molecular explanation for the p53-mediated down-regulation of liver-specific gene expression after viral infection. Additionally, as overexpression of p53 mutants is frequently found in undifferentiated hepatocellular carcinomas, the same mechanisms may contribute to the lack of liver-specific gene transcription in these tumors. PMID- 10542251 TI - A C-terminal-truncated PrP isoform is present in mature sperm. AB - PrP(C), the normal isoform of the prion component PrP(Sc), is a 33-35-kDa glycophosphatidylinositol-anchored glycoprotein expressed in the plasma membrane of many cells and especially in the brain. The specific role of PrP(C) is unknown, although lately it has been shown to bind copper specifically. We show here that PrP(C) is present even in mature sperm cells, a polarized cell that retains only the minimal components required for DNA delivery, movement, and energy production. As opposed to PrP(C) in other cells, PrP in ejaculated sperm cells was truncated in its C terminus in the vicinity of residue 200. Sperm PrP, although membrane-bound, was not released by phosphatidylinositol phospholipase C as well as not localized in cholesterol-rich microdomains (rafts). Although no infertility was reported for PrP-ablated mice in normal situations, our results suggest that sperm cells originating from PrP-ablated mice were significantly more susceptible to high copper concentrations than sperm from wild type mice, allocating a protective role for PrP in specific stress situations related to copper toxicity. Since the functions performed by proteins in sperm cells are limited, these cells may constitute an ideal system to elucidate the function of PrP(C). PMID- 10542253 TI - The human tap nuclear RNA export factor contains a novel transportin-dependent nuclear localization signal that lacks nuclear export signal function. AB - The human Tap protein mediates the sequence-specific nuclear export of RNAs containing the constitutive transport element and is likely also critical for general mRNA export. Here, we demonstrate that a previously defined arginine-rich nuclear localization signal (NLS) present in Tap acts exclusively via the transportin import factor. Previously, transportin has been shown to mediate the nuclear import of several heterogeneous nuclear ribonucleoproteins, including heterogeneous nuclear ribonucleoprotein (hnRNP) A1, by binding to a sequence element termed M9. Although the Tap NLS and the hnRNP A1 M9 element are shown to compete for transportin binding, they show no sequence homology, and the Tap NLS does not conform to the recently defined M9 consensus. The Tap NLS also differs from M9 in that only the latter is able to act as a nuclear export signal. The Tap NLS is therefore the first member of a novel class of transportin-specific NLSs that lack nuclear export signal function. PMID- 10542252 TI - G(s)alpha repression of adipogenesis via Syk. AB - G(s)alpha regulates the differentiation of 3T3-L1 mouse embryonic fibroblasts to adipocytes, a process termed adipogenesis. Inducers of adipogenesis lead to a loss of G(s)alpha and derepress differentiation to adipocytes. The broad spectrum tyrosine kinase inhibitor genistein is shown to block induction of adipogenesis, suggesting an early role of tyrosine phosphorylation in adipogenesis. Staining of phosphotyrosine identified prominent staining of a approximately 70-kDa protein, hypothesized to be the tyrosine kinase Syk. Reverse transcription and polymerase chain reaction amplification established the expression of Syk mRNA in these embryonic fibroblasts. Immunoprecipitations with Syk-specific antibodies demonstrated the presence of Syk in fibroblasts and a rapid increase in the amount of phospho-Syk, peaking at 24 h post induction. Clones constitutively expressing G(s)alpha, which can no longer be induced to differentiate, no longer display increased phospho-Syk levels in response to inducers. The linkage between G(s)alpha and Syk was probed by immunoprecipitations revealing association of Syk with G(s)alpha in the absence of induction. Upon induction of adipogenesis, G(s)alpha levels decline and phospho-Syk levels as well as Syk kinase activity increase. Expression of wild-type Syk both potentiates the ability of inducers to act as well as induces adipogenesis itself. Expression of the kinase-deficient Syk had no such effects on adipogenesis. These data provide a new insight into the control of adipogenesis, suggesting that G(s)alpha represses adipogenesis via Syk. Treatment with the inducers promotes a decline in G(s)alpha, increases in levels of phospho-Syk, and adipogenesis. PMID- 10542254 TI - Structure and regulation of the mouse ing1 gene. Three alternative transcripts encode two phd finger proteins that have opposite effects on p53 function. AB - The human ING1 gene encodes nuclear protein p33(ING1), previously shown to cooperate with p53 in cell growth control (Garkavtsev, I., Grigorian, I. A., Ossovskaya, V. S., Chernov, M. V., Chumakov, P. M., and Gudkov, A. V. (1998) Nature 391, 295-298). p33(ING1) belongs to a small family of proteins from human, mouse, and yeast of approximately the same size that show significant similarity to one another within the C-terminal PHD finger domain and also contain an additional N-terminal region with subtle but reliably detectable sequence conservation. Mouse ing1 is transcribed from three differently regulated promoters localized within a 4-kilobase pair region of genomic DNA. The resulting transcripts share a long common region encoded by a common exon and differ in their 5'-exon sequences. Two transcripts are translated into the same protein of 185 amino acids, the mouse equivalent of the human p33(ING1), while the third transcript encodes a longer protein that has 94 additional N-terminal amino acids. Overexpression of the longer protein interferes with the accumulation of p53 protein and activation of p53-responsive promoters after DNA damage. Between the two products of ing1, only the longer one forms a complex with p53 detectable by immunoprecipitation. These results indicate that a single gene, ing1, encodes both p53-suppressing and p53-activating proteins that are regulated by alternative promoters. PMID- 10542256 TI - Molecular cloning and biological activity of a novel Ha-Ras suppressor gene predominantly expressed in skeletal muscle, heart, brain, and bone marrow by differential display using clonal mouse EC cells, ATDC5. AB - We cloned a cDNA encoding a novel mouse protein, named A-C1, by differential display between two mouse cell lines: embryonic fibroblast C3H10T1/2 and chondrogenic ATDC5. The deduced amino acid sequence of A-C1 consists of 167 amino acids and shows 46% identity with that of a ras-responsive gene, rat Ha-rev107. Northern blot analysis showed a distinct hybridization band of 3.2 kilobases. Expression of A-C1 mRNA was detected in undifferentiated ATDC5 cells and myoblastic C2C12 cells, while none of C3H10T1/2 cells, NIH3T3 fibroblasts, Balb/c 3T3 fibroblasts, osteoblastic MC3T3-E1 cells, and ST2 bone marrow stromal cells expressed A-C1 mRNA in vitro. Moreover, A-C1 mRNA was expressed in skeletal muscle, heart, brain, and bone marrow in adult mice. By in situ hybridization, A C1 gene expression was localized in hippocampus as well as bone marrow cells. By immunocytochemistry, A-C1 protein was detected in the cytoplasm as well as perinuclear region of the cells. Transfection of A-C1 cDNA into Ha-ras transformed NIH3T3 cell line caused increase in the number of flat colonies and inhibition of cell growth. Our data indicate that A-C1 is expressed in some specific tissues in vivo and modulates Ha-ras-mediated signaling pathway. PMID- 10542255 TI - Determinants of ligand binding specificity of the alpha(1)beta(1) and alpha(2)beta(1) integrins. AB - The alpha(1)beta(1) and alpha(2)beta(1) integrins are cell surface collagen receptors. Cells expressing the alpha(1)beta(1) integrin preferentially adhere to collagen IV, whereas cells expressing the alpha(2)beta(1) integrin preferentially adhere to collagen I. Recombinant alpha(1) and alpha(2) integrin I domains exhibit the same collagen type preferences as the intact integrins. In addition, the alpha(2) integrin I domain binds echovirus 1; the alpha(1) I domain does not. To identify the structural components of the I domains responsible for the varying ligand specificities, we have engineered several alpha(1)/alpha(2) integrin I domain chimeras and evaluated their virus and collagen binding activities. Initially, large secondary structural components of the alpha(2) I domain were replaced with corresponding regions of the alpha(1) I domain. Following analysis in echovirus 1 and collagen binding assays, chimeras with successively smaller regions of alpha(1) I were constructed and analyzed. The chimeras were analyzed by ELISA with several different alpha(2) integrin monoclonal antibodies to assess their proper folding. Three different regions of the alpha(1) I domain, when present in the alpha(2) I domain, conferred enhanced collagen IV binding activity upon the alpha(2) I domain. These include the alpha3 and alpha5 helices and a portion of the alpha6 helix. Echovirus 1 binding was lost in a chimera containing the alphaC-alpha6 loop; higher resolution mapping identified Asn(289) as playing a critical role in echovirus 1 binding. Asn(289) had not been implicated in previous echovirus 1 binding studies. Taken together, these data reveal the existence of multiple determinants of ligand binding specificities within the alpha(1) and alpha(2) integrin I domains. PMID- 10542257 TI - The interferon-induced double-stranded RNA-activated protein kinase PKR will phosphorylate serine, threonine, or tyrosine at residue 51 in eukaryotic initiation factor 2alpha. AB - The family of eukaryotic initiation factor 2alpha (eIF2alpha) protein kinases plays an important role in regulating cellular protein synthesis under stress conditions. The mammalian kinases PKR and HRI and the yeast kinase GCN2 specifically phosphorylate Ser-51 on the alpha subunit of the translation initiation factor eIF2. By using an in vivo assay in yeast, the substrate specificity of these three eIF2alpha kinases was examined by substituting Ser-51 in eIF2alpha with Thr or Tyr. In yeast, phosphorylation of eIF2 inhibits general translation but derepresses translation of the GCN4 mRNA. All three kinases phosphorylated Thr in place of Ser-51 and were able to regulate general and GCN4 specific translation. In addition, both PKR and HRI were found to phosphorylate eIF2alpha-S51Y and stimulate GCN4 expression. Isoelectric focusing analysis of eIF2alpha followed by detection using anti-eIF2alpha and anti-phosphotyrosine specific antibodies demonstrated that PKR and HRI phosphorylated eIF2alpha-S51Y on Tyr in vivo. These results provide new insights into the substrate recognition properties of the eIF2alpha kinases, and they are intriguing considering the potential for alternate substrates for PKR in cellular signaling and growth control pathways. PMID- 10542258 TI - A novel NE-dlg/SAP102-associated protein, p51-nedasin, related to the amidohydrolase superfamily, interferes with the association between NE-dlg/SAP102 and N-methyl-D-aspartate receptor. AB - The membrane-associated guanylate kinase proteins have been known to interact various membrane receptors with their N-terminal segments designated the PDZ domains and to cluster these receptors at the target site of the cell membrane. NE-dlg/SAP102, a neuronal and endocrine tissue-specific MAGUK family protein, was found to be expressed in both dendrites and cell bodies in neuronal cells. Although NE-dlg/SAP102 localized at dendrites was shown to interact with N-methyl D-aspartate receptor 2B via the PDZ domains to compose postsynaptic density, the binding proteins existing in the cell body of the neuron are still unknown. Here we report the isolation of a novel NE-dlg/SAP102-associated protein, p51-nedasin. Nedasin has a significant homology with amidohydrolase superfamily proteins and shows identical sequences to a recently identified protein that has guanine aminohydrolase activity. Nedasin has four alternative splice variants (S, V1, V2, and V3) that exhibited different C-terminal structures. NE-dlg/SAP102 is shown to interact with only the S form of nedasin which is predominantly expressed in brain. The expression of nedasin in neuronal cells increases in parallel with the progress of synaptogenesis and is mainly detected in cell bodies where it co localizes with NE-dlg/SAP102. Furthermore, nedasin interferes with the association between NE-dlg/SAP102 and NMDA receptor 2B in vitro. These findings suggest that alternative splicing of nedasin may play a role in the formation and/or structural change in synapses during neuronal development by modifying clustering of neurotransmitter receptors at the synaptic sites. PMID- 10542259 TI - Cloning and uterus/oviduct-specific expression of a novel estrogen-regulated gene (ERG1) AB - The steroid hormone estrogen profoundly influences growth and differentiation programs in the reproductive tract of cycling and pregnant mamals. It is thought that estrogen exerts its cellular effects by regulating the expression of specific target genes. We utilized a messenger RNA differential display method to identify the genes whose expression is modulated by estrogen in the preimplantation rat uterus. Here we report the cloning of a novel gene (ERG1) that is tightly regulated by estrogen in two key reproductive tissues, the uterus and oviduct. Spatio-temporal analyses reveal that ERG1 mRNA is expressed in a highly stage-specific manner in the uterus and oviduct, and its expression is restricted to the surface epithelium of both of these tissues. Nucleotide sequence analysis of the full-length ERG1 cDNA indicates that it has an open reading frame of 1821 nuceotides encoding a putative protein of 607 amino acids with a single transmembrane domain and a short cytoplasmic tail. The extracellular part of the protein contains several distinct structural motifs. These include a zona pellucida binding domain, which is present in a number of proteins such as the zona pellucida sperm binding proteins, and uromodulin, In addition, there is a repeat of a motif called CUB domain, which exists in a number of genes involved in development and differentiation such as bone morphogenetic protein 1 (BMP1). Although the precise function of ERG1 eludes us presently, its unique pattern of expression in the uterus and oviduct and its regulation by estrogen, a principal reproductive hormone, lead us to speculate that this novel gene plays an important role in events during the reproductive cycle and early pregnancy. PMID- 10542260 TI - Coagulation factors VIIa and Xa induce cell signaling leading to up-regulation of the egr-1 gene. AB - Intracellular signaling induced by the coagulation factors (F) VIIa and Xa is poorly understood. We report here studies on these processes in a human keratinocyte line (HaCaT), which is a constitutive producer of tissue factor (TF) and responds to both FVIIa and FXa with elevation of cytosolic Ca(2+), phosphorylation of extracellular signal-regulated kinase (Erk) 1/2, p38(MAPK), and c-Jun N-terminal kinase, and up-regulation of transcription of the early growth response gene-1 (egr-1). Using egr-1 as end point, we observed with both agonists that phosphatidylinositol-specific phospholipase C and the mitogen activated protein kinase/Erk kinase/Erk pathway were mediators of the responses. The responses to FVIIa were TF-dependent and up-regulation of egr-1 mRNA did not require presence of the TF cytoplasmic domain. Antibodies to EPR-1 and factor V had no effect on the response to FXa. We have provided evidence that TF is not the sole component of the FVIIa receptor. The requirement for proteolytic activity of both FVIIa and FXa suggests that protease-activated receptors may be involved. We now report evidence suggesting that protease-activated receptor 2 or a close homologue may be a necessary but not sufficient component of this particular signal transduction pathway. The up-regulation of egr-1 describes one way by which the initiation of blood coagulation may influence gene transcription. The ability of these coagulation proteases to induce intracellular signals at concentrations at or below the plasma concentrations of their zymogen precursors suggests that these processes may occur also in vivo. PMID- 10542261 TI - Microfibril-associated protein 4 is present in lung washings and binds to the collagen region of lung surfactant protein D. AB - We have purified a glycoprotein from bovine lung washings using affinity chromatography on a maltose-affinity column. On SDS-polyacrylamide gel electrophoresis the protein showed a molecular mass of 36 kDa in the reduced state and 66 kDa in the unreduced state. On gel permeation chromatography the apparent molecular mass was 250 kDa. N-terminal sequencing showed homology to the human matrix protein microfibril-associated protein (hMFAP4), and the glycoprotein was designated bovine MFAP4 (bMFAP4). Lung surfactant protein D (SP D) was also purified from lung washings, and calcium-dependent binding was demonstrated between bMFAP4 and SP-D. hMFAP4 was cloned, and recombinant hMFAP4 showed the same binding pattern to SP-D as bMFAP4. No binding was seen to recombinant SP-D composed of the neck region and carbohydrate recognition domain of SP-D, indicating that the interaction between MFAP4 and SP-D is mediated via the collagen region of SP-D. MFAP4 also showed calcium-dependent binding to mannan, which was partially inhibited by maltose. Our findings indicate that MFAP4 has two binding specificities, one for collagen and one for carbohydrate, and we suggest that MFAP4 may fix the collectins in the extracellular compartment during inflammation. PMID- 10542262 TI - Transgenic overexpression of beta(2)-adrenergic receptors in airway smooth muscle alters myocyte function and ablates bronchial hyperreactivity. AB - beta(2)-Adrenergic receptors (beta(2)AR) act to relax airway smooth muscle and can serve to counteract hyperresponsiveness, although the effect may not be ablative even in the presence of exogenous agonist. Within this signaling cascade that ultimately transduces smooth muscle relaxation, a significant "spare receptor" pool has been hypothesized to be present in the airway. In order to modify the relationship between beta(2)AR and downstream effectors, transgenic mice (TG) were created overexpressing beta(2)AR approximately 75-fold in airway smooth muscle using a mouse smooth muscle alpha-actin promoter. While >90% of these receptors were expressed on the smooth muscle cell surface, the percentage of receptors able to form the agonist-promoted high affinity complex was less than that found with nontransgenic (NTG) cells (R(H) = 18 versus 36%). Nevertheless, beta(2)AR signaling was found to be enhanced. Intact airway smooth muscle cells from TG had basal cAMP levels that were greater than NTG cells. A marked increase in agonist-stimulated cAMP levels was found in the TG ( approximately 200% stimulation over basal) compared with NTG ( approximately 50% over basal) cells. Adenylyl cyclase studies gave similar results and also showed a 10-fold lower EC(50) for TG cells. Tracheal rings from TG mice that were precontracted with acetylcholine had an enhanced responsiveness (relaxation) to beta-agonist, with a 60-fold decrease in the ED(50), indicating that the enhanced signaling imposed by overexpression results in an increase in the coordinated function of the intact airway cells. In vivo studies showed a significantly blunted airway resistance response to the inhaled bronchoconstrictor methacholine in the TG mice. Indeed, with beta-agonist pretreatment, the TG mice displayed no response whatsoever to methacholine. These results are consistent with beta(2)AR being the limiting factor in the transduction system. Increases in the initial component of this transduction system (the beta(2)AR) are sufficient to markedly alter signaling and airway smooth muscle function to the extent that bronchial hyperresponsiveness is ablated, consistent with an anti-asthma phenotype. PMID- 10542263 TI - Association of beta-arrestin with G protein-coupled receptors during clathrin mediated endocytosis dictates the profile of receptor resensitization. AB - Resensitization of G protein-coupled receptors (GPCRs) following agonist-mediated desensitization is a necessary step for maintaining physiological responsiveness. However, the molecular mechanisms governing the nature of GPCR resensitization are poorly understood. Here, we examine the role of beta-arrestin in the resensitization of the beta(2) adrenergic receptor (beta(2)AR), known to recycle and resensitize rapidly, and the vasopressin V2 receptor (V2R), known to recycle and resensitize slowly. Upon agonist activation, both receptors recruit beta arrestin to the plasma membrane and internalize in a beta-arrestin- and clathrin dependent manner. However, whereas beta-arrestin dissociates from the beta(2)AR at the plasma membrane, it internalizes with the V2R into endosomes. The differential trafficking of beta-arrestin and the ability of these two receptors to dephosphorylate, recycle, and resensitize is completely reversed when the carboxyl-terminal tails of these two receptors are switched. Moreover, the ability of beta-arrestin to remain associated with desensitized GPCRs during clathrin-mediated endocytosis is mediated by a specific cluster of phosphorylated serine residues in the receptor carboxyl-terminal tail. These results demonstrate that the interaction of beta-arrestin with a specific motif in the GPCR carboxyl terminal tail dictates the rate of receptor dephosphorylation, recycling, and resensitization, and thus provide direct evidence for a novel mechanism by which beta-arrestins regulate the reestablishment of GPCR responsiveness. PMID- 10542265 TI - Identification of the RNA binding domain of T4 RegA protein by structure-based mutagenesis. AB - The T4 translational repressor RegA protein folds into two structural domains, as revealed by the crystal structure (Kang, C.-H. , Chan, R., Berger, I., Lockshin, C., Green, L., Gold, L., and Rich, A. (1995) Science 268, 1170-1173). Domain I of the RegA protein contains a four-stranded beta-sheet and two alpha-helices. Domain II contains a four-stranded beta-sheet and an unusual 3/10 helix. Since beta-sheet residues play a role in a number of protein-RNA interactions, one or both of the beta-sheet regions in RegA protein may be involved in RNA binding. To test this possibility, mutagenesis of residues on both beta-sheets was performed, and the effects on the RNA binding affinities of RegA protein were measured. Additional sites for mutagenesis were selected from molecular modeling of RegA protein. The RNA binding affinities of three purified mutant RegA proteins were evaluated by fluorescence quenching equilibrium binding assays. The activities of the remainder of the mutant proteins were evaluated by quantitative RNA gel mobility shift assays using lysed cell supernatants. The results of this mutagenesis study ruled out the participation of beta-sheet residues. Instead, the RNA binding site was found to be a surface pocket formed by residues on two loops and an alpha-helix. Thus, RegA protein appears to use a unique structural motif in binding RNA, which may be related to its unusual RNA recognition properties. PMID- 10542264 TI - Smad7 differentially regulates transforming growth factor beta-mediated signaling pathways. AB - Smad7 has been identified as a negative regulator of transforming growth factor beta (TGF-beta) signaling by interfering with the phosphorylation of other Smad proteins by TGF-beta receptor type I (TbetaRI). We established a mink lung epithelial (Mv1Lu) cell line where ectopic expression of Smad7 is tightly controlled by doxycycline using an improved Tet-on system. Once induced by doxycycline, the recombinant Smad7 was localized predominantly in the perinuclear region and in the cytoplasm. However, the type of culture surface alters the subcellular localization of Smad7: on plastic or on fibronectin-coated glass, Smad7 was localized in the cytoplasm; but when the cells were cultured on glass, nuclear localization was observed. TGF-beta stimulation did not alter substantially the cellular distribution of Smad7. Importantly, the expression of recombinant Smad7 differentially inhibited TGF-beta signaling pathways. Consistent with previous studies, Smad7 inhibited TGF-beta-stimulated induction of type 1 plasminogen activator inhibitor as measured by p3TP-Lux reporter. However, expression of Smad7 had little effect on TGF-beta-induced growth inhibition. PMID- 10542266 TI - Heregulin induces phosphorylation of BRCA1 through phosphatidylinositol 3 Kinase/AKT in breast cancer cells. AB - The breast cancer susceptibility gene BRCA1 encodes a nuclear phosphoprotein that acts as a tumor suppressor. Phosphorylation of BRCA1 has been implicated in altering its function, however, the pathway(s) that leads to the phosphorylation of BRCA1 has not been described. Here, a signaling pathway by which heregulin induces cell cycle-independent phosphorylation of BRCA1 was delineated. We showed that heregulin stimulation induced the phosphorylation of BRCA1 and concomitant activation of the serine/threonine kinase AKT in T47D human breast cancer cells. Heregulin-induced phosphorylation of BRCA1 was abrogated by phosphatidylinositol 3-kinase (PI3K) inhibitors and by a dominant-negative AKT. In the absence of heregulin, the ectopic expression of the constitutively active p110 subunit of PI3K was sufficient to induce BRCA1 phosphorylation. Furthermore, the purified glutathione S-transferase/AKT kinase phosphorylated BRCA1 in vitro. We have also shown that the phosphorylation of BRCA1 by AKT occurs on the residue Thr-509, which is located in the nuclear localization signal. These results reveal a novel signaling pathway that links extracellular signals to the phosphorylation of BRCA1 in breast cancer cells. PMID- 10542267 TI - A GCM motif protein is involved in placenta-specific expression of human aromatase gene. AB - A new cis-element, trophoblast-specific element 2 (TSE2) is located in the placenta-specific enhancer of the human aromatase gene that dictates its tissue specific expression. In the minimum enhancer region, an element similar to the trophoblast-specific element (TSE), originally described for the human chorionic gonadotropin alpha-subunit gene, also exists (Yamada, K., Harada, N., Honda, S., and Takagi, Y. (1995) J. Biol. Chem. 270, 25064-25069). The co-presence of TSE and TSE2 is required to direct trophoblast-specific expression driven by a heterologous thymidine kinase promoter. A 2562-base pair cDNA clone encoding a 436-amino acid protein that binds to TSE2 was isolated from a human placental cDNA library using a yeast one-hybrid system with the TSE2 as a reporter sequence. The protein was revealed to be identical to hGCMa, a mammalian homologue of the Drosophila GCM (glia cells missing) protein. Expression of hGCMa is restricted to the placenta. The protein also binds to PLE1 in the leptin promoter among other cis-elements reported to confer placenta-specific expression, suggesting that hGCMa is a placenta-specific transcription regulator, possibly involved in the expression of multiple placenta-specific genes. PMID- 10542268 TI - An atypical form of alphaB-crystallin is present in high concentration in some human cataractous lenses. Identification and characterization of aberrant N- and C-terminal processing. AB - Two unique polypeptides, 22.4 and 16.4 kDa, were prominent in some human cataracts. Both proteins were identified as modified forms of the small heat shock protein, alphaB-crystallin. The concentration of total alphaB-crystallin in most of these cataracts was significantly increased. The 22.4-kDa protein was subsequently designated as alphaB(g). Mass spectrometric analyses of tryptic and Asp-N digests showed alphaB(g) is alphaB-crystallin minus the C-terminal lysine. alphaB(g) constituted 10-90% of the total alphaB-crystallin in these cataracts and was preferentially phosphorylated over the typical form of alphaB-crystallin. Human alphaB(g) and alphaB-crystallin were cloned and expressed in Escherichia coli. The differences in electrophoretic mobility and the large difference in native pI values suggest some structural differences exist. The chaperone-like activity of recombinant human alphaB(g) was comparable to that of recombinant human alphaB-crystallin in preventing the aggregation of lactalbumin induced by dithiothreitol. The mechanism involved in generating alphaB(g) is not known, but a premature termination of the alphaB-crystallin gene was ruled out by sequencing the polymerase chain reaction products of the last exon for the alphaB-crystallin gene from lenses containing alphaB(g). The 16.4-kDa protein was an N-terminally truncated fragment of alphaB(g). The high concentration of alphaB-crystallin in these cataracts is the first observation of this kind in human lenses. PMID- 10542270 TI - Uptake, degradation, and release of fibrillar and soluble forms of Alzheimer's amyloid beta-peptide by microglial cells. AB - Microglia are phagocytic cells that are the main inflammatory response cells of the central nervous system. In Alzheimer's disease brain, activated microglia are concentrated in regions of compact amyloid deposits that contain the 39-43-amino acid Abeta peptide. We examined the uptake, degradation, and release of small aggregates of fibrillar Abeta (fAbeta) or soluble Abeta (sAbeta) by microglia. We found that although some degradation of fAbeta was observed over 3 days, no further degradation was observed over the next 9 days. Instead, there was a slow release of intact Abeta. The poor degradation was not due to inhibition of lysosomal function, since the rate of alpha2-macroglobulin degradation was not affected by the presence of fAbeta in the late endosomes/lysosomes. In contrast to fAbeta, internalization of sAbeta was not saturable. After internalization, sAbeta was released rapidly from microglia, and very little was degraded. These data show that fAbeta and sAbeta interact differently with microglia but that after internalization a large fraction of both are released without degradation. PMID- 10542269 TI - Expression of beta-amyloid precursor protein-CD3gamma chimeras to demonstrate the selective generation of amyloid beta(1-40) and amyloid beta(1-42) peptides within secretory and endocytic compartments. AB - Amyloid beta-protein (Abeta) is the main constituent of amyloid fibrils found in senile plaques and cerebral vessels in Alzheimer's disease (AD) and is derived by proteolysis from the beta-amyloid precursor protein (APP). We have analyzed the amyloidogenic processing of APP using chimeric proteins stably transfected in Chinese hamster ovary cells. The extracellular and transmembrane domains of APP were fused to the cytoplasmic region derived from the CD3 gamma chain of the T cell antigen receptor (CD3gamma). CD3gamma contains an endoplasmic reticulum (ER) retention motif (RKK), in the absence of which the protein is targeted to lysosomes without going through the cell surface (Letourneur, F., and Klausner, R.D. (1992) Cell 69, 1143-1157). We used the wild-type sequence of CD3gamma to create an APP chimera predicted to remain in the ER (gammaAPP(ER)). Deletion of the RKK motif at the C terminus directed the protein directly to the lysosomes (gammaAPP(LYS)). A third chimera was created by removing both lysosomal targeting signals in addition to RKK (gammaAPP(DeltaDelta)). This last construct does not contain known targeting signals and consequently accumulates at the cell surface. We show by immunofluorescence and by biochemical methods that all three APP chimeras localize to the predicted compartments within the cell, thus providing a useful model to study the processing of APP. We found that Abeta(1-40) is generated in the early secretory and endocytic pathways, whereas Abeta(1-42) is made mainly in the secretory pathway. More importantly, we provide evidence that, unlike in neuronal models, both ER/intermediate compartment- and endocytic derived Abeta forms can enter the secretable pool. Finally, we directly demonstrate that lysosomal processing is not involved in the generation or secretion of either Abeta(1-40) or Abeta(1-42). PMID- 10542271 TI - Sp1 and NF-Y synergistically mediate the effect of vitamin D(3) in the p27(Kip1) gene promoter that lacks vitamin D response elements. AB - Vitamin D(3) promotes myeloid leukemic cell lines to differentiate terminally into monocytes/macrophages. It has been reported that overexpression of the cdk inhibitor p27(Kip1) results in the differentiation of the myelomonocytic U937 cell line and that this gene is the target of vitamin D(3). To identify the sequences required for the positive regulation of p27(Kip1) transcription by vitamin D(3), a 3.6-kilobase 5'-flanking region of the human p27(Kip1) gene was examined by transiently transfecting luciferase reporter constructs into U937 cells. The transcriptional activity of this construct was activated by vitamin D(3). Deletion and mutational analysis revealed that both a GGGCGG sequence ( 545/-539) and a CCAAT sequence (-525/-520) were necessary to induce p27(Kip1) gene expression. Importantly, the region containing both of these elements conferred positive responsiveness to vitamin D(3) to a heterologous promoter. Gel shift assays showed that Sp1 binds to the GGGCGG sequence and that NF-Y binds to the CCAAT sequence. Consistent with the roles of these transcription factors, treatment with vitamin D(3) stimulated the DNA binding activities of these factors to each element and induced the change of one NF-Y subunit. We conclude that vitamin D(3) stimulates transcription of the p27(Kip1) gene by a novel mechanism involving Sp1 and NF-Y, but not the vitamin D receptor, during the early stages of U937 cell differentiation. PMID- 10542272 TI - Studies on the ADP-ribose pyrophosphatase subfamily of the nudix hydrolases and tentative identification of trgB, a gene associated with tellurite resistance. AB - Four Nudix hydrolase genes, ysa1 from Saccharomyces cerevisiae, orf209 from Escherichia coli, yqkg from Bacillus subtilis, and hi0398 from Hemophilus influenzae were amplified, cloned into an expression vector, and transformed into E. coli. The expressed proteins were purified and shown to belong to a subfamily of Nudix hydrolases active on ADP-ribose. Comparison with other members of the subfamily revealed a conserved proline 16 amino acid residues downstream of the Nudix box, common to all of the ADP-ribose pyrophosphatase subfamily. In this same region, a conserved tyrosine designates another subfamily, the diadenosine polyphosphate pyrophosphatases, while an array of eight conserved amino acids is indicative of the NADH pyrophosphatases. On the basis of these classifications, the trgB gene, a tellurite resistance factor from Rhodobacter sphaeroides, was predicted to designate an ADP-ribose pyrophosphatase. In support of this hypothesis, a highly specific ADP-ribose pyrophosphatase gene from the archaebacterium, Methanococcus jannaschii, introduced into E. coli, increased the transformant's tolerance to potassium tellurite. PMID- 10542273 TI - Site-specific heterodimerization by paired class homeodomain proteins mediates selective transcriptional responses. AB - Alx4 is a paired class homeodomain protein involved in defining anterior/posterior polarity in the developing limb bud. The paired class of homeodomain proteins cooperatively bind palindromic DNA elements as homodimers or as heterodimers with other paired homeodomain proteins. Previous characterization demonstrates that the strength of the cooperativity as well as the preference for targets is dictated largely by the identity of amino acid 50 of the homeodomain. Here we compare and contrast the DNA binding properties of a glutamine 50 paired homeodomain protein, Alx4, and a lysine 50 paired homeodomain protein, Goosecoid. We demonstrate that Alx4 homodimers, Gsc homodimers, and Alx4/Gsc heterodimers each have distinct DNA binding properties, and each can discriminate between highly related palindromic elements. Using reporter gene assays, we show that Alx4 activates transcription in a site-specific manner, and that Gsc is capable of antagonizing Alx4-mediated activation only from promoter elements that support heterodimer formation. These data demonstrate that paired homeodomain proteins with different DNA binding properties are able to form heterodimeric complexes with unique DNA binding and transcriptional activities. Thus, heterodimerization regulates the DNA binding specificity of these transcription factors and may partially explain how paired homeodomain proteins direct specific developmental functions. PMID- 10542274 TI - p42/44 MAP kinase-dependent and -independent signaling pathways regulate caveolin 1 gene expression. Activation of Ras-MAP kinase and protein kinase a signaling cascades transcriptionally down-regulates caveolin-1 promoter activity. AB - Caveolin-1 is a principal component of caveolae membranes in vivo. Caveolin-1 mRNA and protein expression are down-regulated in NIH 3T3 cells in response to transformation by activated oncogenes, such as H-Ras(G12V) and v-Abl. The mechanisms governing this down-regulation event remain unknown. Here, we show that caveolin-1 gene expression is directly regulated by activation of the Ras p42/44 MAP kinase cascade. Down regulation of caveolin-1 protein expression by Ras is independent of (i) the type of activating mutation (G12V versus Q61L) and (ii) the form of activated Ras transfected (H-Ras versus K-Ras versus N-Ras). Treatment of Ras or Raf-transformed NIH 3T3 cells with a well characterized MEK inhibitor (PD 98059) restores caveolin-1 protein expression. In contrast, treatment of v-Src and v-Abl transformed NIH 3T3 cells with PD 98059 does not restore caveolin-1 expression. Thus, there must be at least two pathways for down regulating caveolin-1 expression: one that is p42/44 MAP kinase-dependent and another that is p42/44 MAP kinase-independent. We focused our efforts on the p42/44 MAP kinase-dependent pathway. The activity of a panel of caveolin-1 promoter constructs was evaluated using transient expression in H-Ras(G12V) transformed NIH 3T3 cells. We show that caveolin-1 promoter activity is up regulated approximately 5-fold by inhibition of the p42/44 MAP kinase cascade. Using electrophoretic mobility shift assays we provide evidence that the caveolin 1 promoter (from -156 to -561) is differentially bound by transcription factors in normal and H-Ras(G12V)-transformed cells. We also show that activation of protein kinase A (PKA) signaling is sufficient to down-regulate caveolin-1 protein expression and promoter activity. Thus, we have identified two signaling pathways (Ras-p42/44 MAP kinase and PKA) that transcriptionally down-regulate caveolin-1 gene expression. PMID- 10542276 TI - The C terminus of a chloroplast precursor modulates its interaction with the translocation apparatus and PIRAC. AB - The import of proteins into chloroplasts involves a cleavable, N-terminal targeting sequence known as the transit peptide. Although the transit peptide is both necessary and sufficient to direct precursor import into chloroplasts, the mature domain of some precursors has been shown to modulate targeting and translocation efficiency. To test the influence of the mature domain of the small subunit of Rubisco during import in vitro, the precursor (prSSU), the mature domain (mSSU), the transit peptide (SS-tp), and three C-terminal deletion mutants (Delta52, Delta67, and Delta74) of prSSU were expressed and purified from Escherichia coli. Activity was then evaluated by competitive import of (35)S prSSU. Both IC(50) and K(i) values consistently suggest that removal of C terminal prSSU sequences inhibits its interaction with the translocation apparatus. Non-competitive import studies demonstrated that prSSU and Delta52 were properly processed and accumulated within the chloroplast, whereas Delta67 and Delta74 were rapidly degraded via a plastid-localized protease. The ability of prSSU-derived proteins to induce inactivation of the protein-import-related anion channel was also evaluated. Although the C-terminal deletion mutants were less effective at inducing channel closure upon import, they did not effect the mean duration of channel closure. Possible mechanisms by which C-terminal residues of prSSU modulate chloroplast targeting are discussed. PMID- 10542275 TI - A novel element and a TEF-2-like element activate the major histocompatibility complex class II transactivator in B-lymphocytes. AB - Major histocompatibility complex (MHC) class II molecules play a central role in immune responses, and transcription of this family of genes requires the MHC class II transactivator (CIITA). CIITA has four promoters, which are transcribed in a tissue-specific manner. CIITA promoter III is constitutively active in mature B-lymphocytes. This report now describes the minimal 319-base pair promoter region necessary for maximal transcriptional activity in B-lymphocytes. Ultraviolet light and dimethylsulfate in vivo genomic footprinting analyses reveal five occupied DNA sequence elements present in intact B-lymphocytes. Functional analysis of these elements using promoter deletions and site-specific mutations demonstrates that at least two of the sites occupied in vivo are critical for transcriptional activity. In vitro protein/DNA analysis suggests that one of the sites is a TEF-2-like element and the other is occupied by a novel transcription activator. In addition, nuclear factor-1 associates with the promoter both in vivo and in vitro. In myeloma cell lines, loss of CIITA transcription correlates with a completely unoccupied CIITA promoter III. These findings suggest that CIITA transcription in B-lymphocytes is activated through at least two strong promoter elements, while loss of expression in myeloma cells is mediated through changes in promoter assembly. PMID- 10542277 TI - The yeast nucleoporin Nup2p is involved in nuclear export of importin alpha/Srp1p. AB - The importin alpha.beta heterodimer mediates nuclear import of proteins containing classical nuclear localization signals. After carrying its cargo into the nucleus, the importin dimer dissociates, and Srp1p (the yeast importin alpha subunit) is recycled to the cytoplasm in a complex with Cse1p and RanGTP. Nup2p is a yeast FXFG nucleoporin that contains a Ran-binding domain. We find that export of Srp1p from the nucleus is impaired in Deltanup2 mutants. Also, Srp1p fusion proteins accumulate at the nuclear rim in wild-type cells but accumulate in the nuclear interior in Deltanup2 cells. A deletion of NUP2 shows genetic interactions with mutants in SRP1 and PRP20, which encodes the Ran nucleotide exchange factor. Srp1p binds directly to an N-terminal domain of Nup2p. This region of Nup2p is sufficient to allow accumulation of an Srp1p fusion protein at the nuclear rim, but the C-terminal Ran-binding domain of Nup2p is required for efficient Srp1p export. Formation of the Srp1p.Cse1p. RanGTP export complex releases Srp1p from its binding site in Nup2p. We propose that Nup2p may act as a scaffold that facilitates formation of the Srp1p export complex. PMID- 10542278 TI - Identification of hMutLbeta, a heterodimer of hMLH1 and hPMS1. AB - hMLH1 and hPMS2 function in postreplicative mismatch repair in the form of a heterodimer referred to as hMutLalpha. Tumors or cell lines lacking this factor display mutator phenotypes and microsatellite instability, and mutations in the hMLH1 and hPMS2 genes predispose to hereditary non-polyposis colon cancer. A third MutL homologue, hPMS1, has also been reported to be mutated in one cancer prone kindred, but the protein encoded by this locus has so far remained without function. We now show that hPMS1 is expressed in human cells and that it interacts with hMLH1 with high affinity to form the heterodimer hMutLbeta. Recombinant hMutLalpha and hMutLbeta, expressed in the baculovirus system, were tested for their activity in an in vitro mismatch repair assay. While hMutLalpha could fully complement extracts of mismatch repair-deficient cell lines lacking hMLH1 or hPMS2, hMutLbeta failed to do so with any of the different substrates tested in this assay. The involvement of the latter factor in postreplicative mismatch repair thus remains to be demonstrated. PMID- 10542279 TI - Analysis of the molecular mechanism for the antagonistic action of a novel 1alpha,25-dihydroxyvitamin D(3) analogue toward vitamin D receptor function. AB - We have recently reported that 23(S)-25-dehydro-1alpha-hydroxyvitamin D(3)-26,23 lactone (TEI-9647) efficiently blocks the differentiation of HL-60 cells induced by 1alpha,25-dihydroxyvitamin D(3) (1alpha,25(OH)(2)D(3)) (Miura, D., Manabe, K., Ozono, K., Saito, M., Gao, Q., Norman, A. W., and Ishizuka, S. (1999) J. Biol. Chem. 274, 16392-16399). To clarify the molecular mechanisms of this antagonism, we examined whether TEI-9647 antagonizes the genomic effects of 1alpha,25(OH)(2)D(3). 10(-7) to 10(-9) M TEI-9647 inhibited the transactivation effect of 10(-8) M 1alpha,25(OH)(2)D(3) in a dose-dependent manner, while TEI 9647 alone did not activate the reporter activity driven by SV40 promoter containing two vitamin D response elements in Saos-2 cells. The antagonistic effect of TEI-9647 was also observed using the rat 24-hydroxylase gene promoter, but the effect was weaker in HeLa and COS-7 cells than in Saos-2 cells. TEI-9647 also exhibited antagonism in an assay system where the VDR fused to the GAL4 DNA binding domain and the reporter plasmid containing the GAL4 binding site were used in Saos-2 cells, but did not in HeLa cells. TEI-9647 reduced the interaction between VDR and RXRalpha according to the results obtained from the mammalian two hybrid system in Saos-2 cells, but did not in HeLa cells. The two-hybrid system also revealed that the interaction between VDR and SRC-1 was reduced by TEI-9647 in Saos-2 cells. These results demonstrate that the novel 1alpha,25(OH)(2)D(3) analogue, TEI-9647, is the first synthetic ligand for the VDR that efficiently antagonizes the action of 1alpha, 25(OH)(2)D(3), although the extent of its antagonism depends on cell type. PMID- 10542280 TI - Hyperglycemia enhances angiotensin II-induced janus-activated kinase/STAT signaling in vascular smooth muscle cells. AB - We have shown previously that angiotensin II (Ang II) activates the janus activated kinase (JAK)/signal transducers and activators of transcription (STAT) pathway in vascular smooth muscle cells (VSMCs) and that activation of the JAK/STAT pathway is required for Ang II induction of VSMC proliferation. In the present study, we examined the effects of hyperglycemia (HG) on Ang II-induced JAK/STAT signaling events in cultured VSMCs. HG increases Ang II-induced JAK2 tyrosine phosphorylation and promotes a partial tyrosine phosphorylation of the enzyme under basal conditions. In addition, HG increases both basal and Ang II induced complex formation of JAK2 with the Ang II AT(1) receptor. The extent of STAT1 and STAT3 tyrosine and serine phosphorylation are also increased under HG conditions. Furthermore, the tyrosine phosphorylation and activities of the SHP-1 and SHP-2 tyrosine phosphatases, enzymes that regulate Ang II-induced JAK2 tyrosine phosphorylation, are altered by HG. SHP-1, which is responsible for JAK2 tyrosine dephosphorylation in VSMC, is completely deactivated in HG, resulting in a prolonged duration of JAK2 phosphorylation under HG conditions. HG also enhances Ang II induction of VSMC proliferation. Taken together, these data suggest that HG augments Ang II induction of VSMC proliferation by increasing signal transduction through the JAK/STAT pathway. PMID- 10542281 TI - Recombinant human aggrecan G1-G2 exhibits native binding properties and substrate specificity for matrix metalloproteinases and aggrecanase. AB - A recombinant human aggrecan G1-G2 fragment comprising amino acids Val(1) Arg(656) has been expressed in Sf21 cells using a baculovirus expression system. The recombinant G1-G2 (rG1-G2) was purified to homogeneity by hyaluronan Sepharose affinity chromatography followed by high performance liquid chromatography gel filtration, and gave a single band of M(r) 90,000-95,000 by silver stain or immunoblotting with monoclonal antibody 1-C-6. The expressed G1 G2 bound to both hyaluronan and link protein indicating that the immunoglobulin fold motif and proteoglycan tandem repeat loops of the G1 domain were correctly folded. Further analysis of secondary structure by rotary shadowing electron microscopy confirmed a double globe appearance, but revealed that the rG1-G2 was more compact than its native counterpart. The size of rG1-G2 by SDS polyacrylamide gel electorphoresis was unchanged following digestion with keratanase and keratanase II and reduced by only 2-5 kDa following digestion with either O-glycosidase or N-glycosidase F. Recombinant G1-G2 was digested with purified matrix metalloproteinases (MMP), isolated aggrecanase, purified atrolysin C, or proteinases present in conditioned medium from cartilage explant cultures, and the products analyzed on SDS gels by silver stain and immunoblotting. Neoepitope antibodies recognizing the N-terminal F(342)FGVG or C terminal DIPEN(341) sequences were used to confirm MMP cleavage at the Asn(341) downward arrow Phe bond, while neoepitope antibodies recognizing the N-terminal A(374)RGSV or C-terminal ITEGE(373) sequences were used to confirm aggrecanase cleavage at the Glu(373) downward arrow Ala bond. Cleavage at the authentic MMP and aggrecanase sites revealed that these proteinases have the same specificity for rG1-G2 as for native aggrecan. Incubation of rG1-G2 with conditioned medium from porcine cartilage cultures revealed that active soluble aggrecanase but no active MMPs, was released following stimulation with interleukin-1alpha or retinoic acid. Atrolysin C, which cleaves native bovine aggrecan at both the aggrecanase and MMP sites, efficiently cleaved rG1-G2 at the aggrecanase site but failed to cleave at the MMP site. In contrast, native glycosylated G1-G2 with or without keratanase treatment was cleaved by atrolysin C at both the aggrecanase and MMP sites. The results suggest that the presence or absence per se of keratan sulfate on native G1-G2 does not affect the activity of atrolysin C toward the two sites. PMID- 10542283 TI - Purification and biophysical characterization of a new [2Fe-2S] ferredoxin from Azotobacter vinelandii, a putative [Fe-S] cluster assembly/repair protein. AB - During the purification of site-directed mutant variants of Azotobacter vinelandii ferredoxin I (FdI), a pink protein, which was not observed in native FdI preparations, appeared to associate specifically with variants that had mutations in ligands to FdI [Fe-S] clusters. That protein, which we designate FdIV, has now been purified. NH(2)-terminal sequence analysis revealed that the protein is the product of a previously described gene, herein designated fdxD, that is in the A. vinelandii iscSUA operon that encodes proteins involved in iron sulfur cluster assembly or repair. An apoprotein molecular mass of 12,434.03 +/- 0.21 Da was determined by mass spectrometry consistent with the known gene sequence. The monomeric protein was shown to contain a single [2Fe-2S](2+/+) cluster by UV/visible, CD, and EPR spectroscopies with a reduction potential of 344 mV versus the standard hydrogen electrode. When overexpressed in Escherichia coli, recombinant FdIV holoprotein was successfully assembled. However, the polypeptide of the recombinant protein was modified in some way such that the apoprotein molecular mass increased by 52 Da. Antibodies raised against FdIV and EPR spectroscopy were used to examine the relative levels of FdIV and FdI in various A. vinelandii strains leading to the conclusion that FdIV levels appear to be specifically increased under conditions where another protein, NADPH:ferredoxin reductase is also up-regulated. In that case, the fpr gene is known to be activated in response to oxidative stress. This suggests that the fdxD gene and other genes in the iron-sulfur cluster assembly or repair operon might be similarly up-regulated in response to oxidative stress. PMID- 10542282 TI - Accumulation of pre-apocytochrome f in a Synechocystis sp. PCC 6803 mutant impaired in cytochrome c maturation. AB - Cytochrome c maturation involves heme transport and covalent attachment of heme to the apoprotein. The 5' end of the ccsB gene, which is involved in the maturation process and resembles the ccs1 gene from Chlamydomonas reinhardtii, was replaced by a chloramphenicol resistance cartridge in the cyanobacterium Synechocystis sp. PCC 6803. The resulting Delta(M1-A24) mutant lacking the first 24 ccsB codons grew only under anaerobic conditions. The mutant retained about 20% of the wild-type amount of processed cytochrome f with heme attached, apparently assembled in a functional cytochrome b(6)f complex. Moreover, the mutant accumulated unprocessed apocytochrome f in its membrane fraction. A pseudorevertant was isolated that regained the ability to grow under aerobic conditions. The locus of the second-site mutation was mapped to ccsB, and the mutation resulted in the formation of a new potential start codon in the intergenic region, between the chloramphenicol resistance marker and ccsB, in frame with the remaining part of ccsB. In this pseudorevertant the amount of holocyt f increased, whereas that of unprocessed apocytochrome f decreased. We suggest that the original deletion mutant Delta(M1-A24) expresses an N-terminally truncated version of the protein. The stable accumulation of unprocessed apocytochrome f in membranes of the Delta(M1-A24) mutant may be explained by its association with truncated and only partially functional CcsB protein resulting in protection from degradation. Our attempt to delete the first 244 codons of ccsB in Synechocystis sp. PCC 6803 was not successful, suggesting that this would lead to a lack of functional cytochrome b(6)f complex. The results suggest that the CcsB protein is an apocytochrome chaperone, which together with CcsA may constitute part of cytochrome c lyase. PMID- 10542284 TI - Identification and characterization of falcilysin, a metallopeptidase involved in hemoglobin catabolism within the malaria parasite Plasmodium falciparum. AB - The malaria parasite Plasmodium falciparum degrades hemoglobin in its acidic food vacuole for use as a major nutrient source. A novel metallopeptidase activity, falcilysin, was purified from food vacuoles and characterized. Falcilysin appears to function downstream of the aspartic proteases plasmepsins I and II and the cysteine protease falcipain in the hemoglobin proteolytic pathway. It is unable to cleave hemoglobin or denatured globin but readily destroys peptide fragments of hemoglobin. Falcilysin cleavage sites along the alpha and beta chains of hemoglobin are polar in character, with charged residues located in the P1 and/or P4' positions. In contrast, plasmepsins I and II and falcipain prefer hydrophobic residues around the scissile bond. The gene encoding falcilysin has been cloned. Its coding sequence exhibits features characteristic of clan ME family M16 metallopeptidases, including an "inverted" HXXEH active site motif. Falcilysin shares primary structural features with M16 family members such as insulysin, mitochondrial processing peptidase, nardilysin, and pitrilysin as well as with data base hypothetical proteins that are potential M16 family members. The characterization of falcilysin increases our understanding of hemoglobin catabolism in P. falciparum and the unusual M16 family of metallopeptidases. PMID- 10542285 TI - The COOH terminus of Rho-kinase negatively regulates rho-kinase activity. AB - Rho-kinase is implicated in the phosphorylation of myosin light chain downstream of Rho, which is thought to induce smooth muscle contraction and stress fiber formation in non-muscle cells. Here, we examined the mode of action of inhibitors of Rho-kinase. The chemical compounds such as HA1077 and Y-32885 inhibited not only the Rho-kinase activity but also the activity of protein kinase N, one of the targets of Rho, but had less of an effect on the activity of myotonic dystrophy kinase-related Cdc42-binding kinase beta (MRCKbeta). The COOH-terminal portion of Rho-kinase containing Rho-binding (RB) and pleckstrin homology (PH) domains (RB/PH (TT)), in which point mutations were introduced to abolish the Rho binding activity, interacted with Rho-kinase and thereby inhibited the Rho-kinase activity, whereas RB/PH (TT) had no effect on the activity of protein kinase N or MRCKbeta, suggesting that the COOH-terminal region of Rho-kinase is a possible negative regulatory region of Rho-kinase. The expression of RB/PH (TT) specifically blocked the stress fiber and focal adhesion formation induced by the active form of Rho or Rho-kinase in NIH 3T3 cells, but not that induced by the active form of MRCKbeta or myosin light chain. Thus, RB/PH (TT) appears to specifically inhibit Rho-kinase in vivo. PMID- 10542286 TI - Novel effector control through modulation of a preexisting binding site of the aromatic-responsive sigma(54)-dependent regulator DmpR. AB - The Pseudomonas derived sigma(54)-dependent DmpR activator regulates transcription of the (methyl)phenol catabolic dmp-operon. DmpR is constitutively expressed, but its transcriptional promoting activity is positively controlled in direct response to the presence of multiple aromatic effectors. Previous work has led to a model in which effector binding by the amino-terminal region of the protein relieves repression of an intrinsic ATPase activity essential for its transcriptional promoting property. Here, we address whether the observed differences in the potencies of the multiple effectors (i) reside at the level of different aromatic binding sites, or (ii) are mediated through differential binding affinities; furthermore, we address whether binding of distinct aromatic effectors has different functional consequences for DmpR activity. These questions were addressed by comparing wild type and an effector specificity mutant of DmpR with respect to effector binding characteristics and the ability of aromatics to elicit ATPase activity and transcription. The results demonstrate that six test aromatics all share a common binding site on DmpR and that binding affinities determine the concentration at which DmpR responds to the presence of the effector, but not the magnitude of the responses. Interestingly, this analysis reveals that the novel abilities of the effector specificity mutant are not primarily due to acquisition of new binding abilities, but rather, they reside in being able to productively couple ATPase activity to transcriptional activation. The mechanistic implications of these findings in terms of aromatic control of DmpR activity are discussed. PMID- 10542287 TI - Zinc inhibits protein synthesis in neurons. Potential role of phosphorylation of translation initiation factor-2alpha. AB - In the central nervous system, Zn(2+) is concentrated in the cerebral cortex and hippocampus and has been found to be toxic to neurons. In this study, we show that exposure of cultured cortical neurons from mouse to increasing concentrations of Zn(2+) (10-300 microM) induces a progressive decrease in global protein synthesis. The potency of Zn(2+) was increased by about 2 orders of magnitude in the presence of Na(+)-pyrithione, a Zn(2+) ionophore. The basal rate of protein synthesis was restored 3 h after Zn(2+) removal. Zn(2+) induced a sustained increase in phosphorylation of the alpha subunit of the translation eukaryotic initiation factor-2 (eIF-2alpha), whereas it triggered a transient increase in phosphorylation of eukaryotic elongation factor-2 (eEF-2). Protein synthesis was still depressed 60 min after the onset of Zn(2+) exposure while the state of eEF-2 phosphorylation had already returned to its basal level. Moreover, Zn(2+) was less effective than glutamate to increase eEF-2 phosphorylation, whereas it induced a more profound inhibition of protein synthesis. These results suggest that Zn(2+)-induced inhibition of protein synthesis mainly correlates with the increase in eIF-2alpha phosphorylation. Supporting further that Zn(2+) acts at the initiation step of protein synthesis, it strongly decreased the amount of polyribosomes. PMID- 10542288 TI - Self-catalyzed cleavage of the yeast nucleoporin Nup145p precursor. AB - Nup145p is a component of the nuclear pore complex of Saccharomyces cerevisiae and is essential for mRNA export. Nup145p and its apparent vertebrate homologue are the only known nucleoporins to be composed of two functionally independent peptide moieties resulting from the post-translational cleavage of a large precursor molecule. In this study, the proteolytic cleavage site of Nup145p has been mapped upstream of an evolutionary conserved serine residue. Cleavage occurs at the same site when a precursor is artificially expressed in Escherichia coli. A hydroxyl-containing residue is critical for the reaction, although a thiol containing residue offers an acceptable replacement. In vitro kinetics experiments using a purified precursor molecule demonstrate that the cleavage is self-catalyzed and that the catalytic domain lies within the N-terminal moiety. Taken altogether, our data are consistent with a proteolytic mechanism involving an N>O acyl rearrangement and a subsequent ester intermediate uncovered in other self-processing proteins. PMID- 10542289 TI - Characterization of the gene encoding serine acetyltransferase, a regulated enzyme of cysteine biosynthesis from the protist parasites Entamoeba histolytica and Entamoeba dispar. Regulation and possible function of the cysteine biosynthetic pathway in Entamoeba. AB - The enteric protist parasites Entamoeba histolytica and Entamoeba dispar possess a cysteine biosynthetic pathway, unlike their mammalian host, and are capable of de novo production of L-cysteine. We cloned and characterized cDNAs that encode the regulated enzyme serine acetyltransferase (SAT) in this pathway from these amoebae by genetic complementation of a cysteine-auxotrophic Escherichia coli strain with the amoebic cDNA libraries. The deduced amino acid sequences of the amoebic SATs exhibited, within the most conserved region, 36-52% identities with the bacterial and plant SATs. The amoebic SATs contain a unique insertion of eight amino acids, also found in the corresponding region of a plasmid-encoded SAT from Synechococcus sp., which showed the highest overall identities to the amoebic SATs. Phylogenetic reconstruction also revealed a close kinship of the amoebic SATs with cyanobacterial SATs. Biochemical characterization of the recombinant E. histolytica SAT revealed several enzymatic features that distinguished the amoebic enzyme from the bacterial and plant enzymes: 1) inhibition by L-cysteine in a competitive manner with L-serine; 2) inhibition by L-cystine; and 3) no association with cysteine synthase. Genetically engineered amoeba strains that overproduced cysteine synthase and SAT were created. The cysteine synthase-overproducing amoebae had a higher level of cysteine synthase activity and total thiol content and revealed increased resistance to hydrogen peroxide. These results indicate that the cysteine biosynthetic pathway plays an important role in antioxidative defense of these enteric parasites. PMID- 10542290 TI - Critical flanking sequences of PU.1 binding sites in myeloid-specific promoters. AB - The myeloid-specific transcription factor PU.1 is essential for expression of p47(phox), a component of the superoxide-forming phagocyte NADPH oxidase. The consensus PU.1 binding sequence (GAGGAA) is located on the non-coding strand from position -40 to -45 relative to the transcriptional start site of the p47phox promoter. A promoter construct extending to -46 was sufficient to drive tissue specific expression of the luciferase reporter gene, but extension of the promoter from -46 to -48 resulted in a significant increase in reporter expression. Mutations of the nucleotides G at -46 and/or T at -47 reduced both reporter expression and PU.1 binding, whereas mutations at -48 had no effect. The PU.1 binding avidity of these sequences correlated closely with their capacity to dictate reporter gene transcription. In parallel studies on the functional PU.1 site in the promoter of CD18, mutations of nucleotides G and T at positions -76 and -77 (corresponding to -46 and -47, respectively, of the p47phox promoter) reduced PU.1 binding and nearly abolished the contribution of this element to promoter activity. We conclude that the immediate flanking nucleotides of the PU.1 consensus motif have significant effects on PU.1 binding avidity and activity and that this region is the dominant cis element regulating p47phox expression. PMID- 10542291 TI - F1Aalpha, a death receptor-binding protein homologous to the Caenorhabditis elegans sex-determining protein, FEM-1, is a caspase substrate that mediates apoptosis. AB - Apoptosis is an evolutionarily conserved process that is critical for tissue homeostasis and development including sex determination in essentially all multicellular organisms. Here, we report the cloning of an ankyrin repeat containing protein, termed F1Aalpha, in a yeast two-hybrid screen using the cytoplasmic domain of Fas (CD95/APO-1) as bait. Amino acid sequence analysis indicates that F1Aalpha has extensive homology to the sex-determining protein FEM 1 of the Caenorhabditis elegans, which is required for the development of all aspects of the male phenotype. F1Aalpha associates with the cytoplasmic domains of Fas and tumor necrosis factor receptor 1, two prototype members of the "death receptor" family. The F1Aalpha protein also oligomerizes. Overexpression of F1Aalpha induces apoptosis in mammalian cells, and co-expression of Bcl-XL or the dominant negative mutants of either FADD or caspase-9 blocks this effect. Deletion analysis revealed the center region of F1Aalpha, including a cluster of five ankyrin repeats to be necessary and sufficient for maximum apoptotic activity, and the N-terminal region appears to regulate negatively this activity. Furthermore, F1Aalpha is cleaved by a caspase-3-like protease at Asp(342), and the cleavage-resistant mutant is unable to induce apoptosis upon overexpression. F1Aalpha is therefore a member of a growing family of death receptor-associated proteins that mediates apoptosis. PMID- 10542292 TI - Molecular identification and characterization of novel membrane-bound metalloprotease, the soluble secreted form of which hydrolyzes a variety of vasoactive peptides. AB - One class of zinc metalloproteases, represented by neutral endopeptidase 24.11 and endothelin-converting enzyme, has been shown to be involved in proteolytic activation or inactivation of many regulatory peptides. Here, we report molecular cloning and characterization of a novel member of this type II membrane-bound metalloprotease family, termed soluble secreted endopeptidase (SEP). Alternative splicing results in the generation of another transcript, SEP(Delta), which lacks a 69-base pair nucleotide segment following the transmembrane helix. Both SEP and SEP(Delta) mRNA are detected in all mouse tissues examined. Transfection of an SEP cDNA expression construct resulted in the expression of the membrane-bound form of SEP in the early secretory pathway as well as the soluble secreted form of the enzyme in the culture medium. In contrast, transfection of the SEP(Delta) cDNA only results in the expression of the membrane-bound form. In vitro enzymological analysis of the recombinant soluble form of SEP demonstrated that it hydrolyzes a variety of vasoactive peptides, including endothelin-1, atrial natriuretic peptide, and angiotensin I. This activity of SEP was inhibited by phosphoramidon and the neutral endopeptidase 24.11 specific inhibitor thiorphan, but it was only partially inhibited by the endothelin-converting enzyme specific inhibitor FR901533. These findings suggest that SEP is a novel metalloprotease that possesses a broad substrate specificity and that it may be involved in the metabolism of biologically active peptides intracellulary as well as extracellularly. PMID- 10542293 TI - Amino-terminally modified RANTES analogues demonstrate differential effects on RANTES receptors. AB - Modification of the amino terminus of regulated on activated normal T-cell expressed (RANTES) has been shown to have a significant effect on biological activity and produces proteins with antagonist properties. Two amino-terminally modified RANTES proteins, Met-RANTES and aminooxypentane-RANTES (AOP-RANTES), exhibit differential inhibitory properties on both monocyte and eosinophil chemotaxis. We have investigated their binding properties as well as their ability to activate the RANTES receptors CCR1, CCR3, and CCR5 in cell lines overexpressing these receptors. We show that Met-RANTES has weak activity in eliciting a calcium response in Chinese hamster ovary cells expressing CCR1, CCR3, and CCR5, whereas AOP-RANTES has full agonist activity on CCR5 but is less effective on CCR3 and CCR1. Their ability to induce chemotaxis of the murine pre B lymphoma cell line, L1.2, transfected with the same receptors, consolidates these results. Monocytes have detectable mRNA for CCR1, CCR2, CCR3, CCR4, and CCR5, and they respond to the ligands for these receptors in chemotaxis but not always in calcium mobilization. AOP-RANTES does not induce calcium mobilization in circulating monocytes but is able to do so as these cells acquire the macrophage phenotype, which coincides with a concomitant up-regulation of CCR5. We have also tested the ability of both modified proteins to induce chemotaxis of freshly isolated monocytes and eosinophils. Cells from most donors do not respond, but occasionally cells from a particular donor do respond, particularly to AOP-RANTES. We therefore hypothesize that the occasional activity of AOP RANTES to induce leukocyte chemotaxis is due to donor to donor variation of receptor expression. PMID- 10542294 TI - Integrin-mediated muscle cell spreading. The role of protein kinase c in outside in and inside-out signaling and evidence of integrin cross-talk. AB - Muscle cell survival depends upon the presence of various integrins with affinities for different extracellular matrix proteins. The absence of either alpha(5) or alpha(7) integrins leads to degenerative disorders of skeletal muscle, muscular dystrophies. To understand the cell survival signals that are mediated by integrin engagement with matrix proteins, we studied the early signaling events initiated by the attachment of muscle cells to fibronectin, an interaction that is mediated primarily by alpha(5) integrins. Cells that express alpha(5) integrin rapidly spread on fibronectin, and this process is associated with the phosphorylation of focal adhesion kinase (FAK). Cells deficient in alpha(5) integrin failed to spread or promote FAK phosphorylation when plated on fibronectin. For alpha(5)-expressing cells, both spreading and FAK phosphorylation could be blocked by inhibitors of protein kinase C (PKC), indicating that PKC is necessary for this "outside-in signaling" mediated by alpha(5) integrin. Surprisingly, activators of PKC could promote spreading and FAK phosphorylation in alpha(5)-deficient muscle cells plated on fibronectin. This PKC-induced cell spreading appeared to be due to activation of alpha(4) integrins ("inside-out signaling") since it could be blocked by peptides that specifically inhibit alpha(4) integrin binding to fibronectin. A model of integrin signaling in muscle cells is presented in which there is a positive feedback loop involving PKC in both outside-in and inside-out signaling, and the activation of this cycle is essential for cell spreading and downstream signaling to promote cell survival. In addition, the data indicate a cross-talk that occurs between integrins in which the outside-in signaling via one integrin can promote the activation of another integrin via inside-out signaling. PMID- 10542295 TI - Signal regulatory proteins negatively regulate immunoreceptor-dependent cell activation. AB - Signal regulatory proteins of the alpha subtype (SIRPalpha) are ubiquitous molecules of the immunoglobulin superfamily that negatively regulate protein tyrosine kinase receptor-dependent cell proliferation. Their intracytoplasmic domain contains four motifs that resemble immunoreceptor tyrosine-based inhibition motifs (ITIMs) and that, when tyrosyl-phosphorylated, recruit cytoplasmic SH2 domain-bearing protein tyrosine phosphatases (SHPs). ITIMs are borne by molecules that negatively regulate cell activation induced by receptors bearing immunoreceptor tyrosine-based activation motifs (ITAMs). Because SIRPalpha are coexpressed with ITAM-bearing receptors in hematopoietic cells, we investigated whether SIRPalpha could negatively regulate ITAM-dependent cell activation. We found SIRPalpha transcripts in human mast cells, and we show that a chimeric molecule having the transmembrane and intracytoplasmic domains of SIRPalpha could inhibit IgE-induced mediator secretion and cytokine synthesis by mast cells. Inhibition required that the SIRPalpha chimera was coaggregated with ITAM-bearing high affinity IgE receptors (FcepsilonRI). It was correlated with the tyrosyl phosphorylation of the SIRPalpha chimera and the recruitment of SHP-1 and SHP-2. The phosphorylation of FcepsilonRI ITAMs was decreased; the mobilization of intracellular Ca(2+) and the influx of extracellular Ca(2+) were reduced, and the activation of the mitogen-activated protein kinases Erk1 and Erk2 was abolished. SIRPalpha can therefore negatively regulate not only receptor tyrosine kinase-dependent cell proliferation but also ITAM-dependent cell activation. PMID- 10542296 TI - Inhibition of the FixL sensor kinase by the FixT protein in Sinorhizobium meliloti. AB - Nitrogen fixation in symbiotic rhizobia is subject to multiple levels of gene regulation. In Sinorhizobium meliloti, the alfalfa symbiont, the FixLJ two component regulatory system plays a major role in inducing nitrogen fixation and respiration gene expression in response to the low ambient O(2) concentration of the nodule. Here we report on the mode of action of the FixT protein, a recently identified repressor of nitrogen fixation gene expression in S. meliloti. First, we provide evidence that FixT prevents transcription of the intermediate key regulatory genes nifA and fixK by counteracting the activity of the FixLJ two component system under otherwise inducing microoxic conditions. Second, we demonstrate that FixT acts as an inhibitor of the sensor hemoprotein kinase FixL, preventing the production or the accumulation of its phosphorylated form. FixT is thus a new example of a regulatory protein that blocks signal transduction in two component systems at the level of the sensor kinase. PMID- 10542298 TI - Oxidized low density lipoprotein displaces endothelial nitric-oxide synthase (eNOS) from plasmalemmal caveolae and impairs eNOS activation. AB - Hypercholesterolemia-induced vascular disease and atherosclerosis are characterized by a decrease in the bioavailability of endothelium-derived nitric oxide. Endothelial nitric-oxide synthase (eNOS) associates with caveolae and is directly regulated by the caveola protein, caveolin. In the present study, we examined the effects of oxidized low density lipoprotein (oxLDL) on the subcellular location of eNOS, on eNOS activation, and on caveola cholesterol in endothelial cells. We found that treatment with 10 microgram/ml oxLDL for 60 min caused greater than 90% of eNOS and caveolin to leave caveolae. Treatment with oxLDL also inhibited acetylcholine-induced activation of eNOS but not prostacyclin production. oxLDL did not affect total cellular eNOS abundance. Oxidized LDL also did not affect the palmitoylation, myristoylation or phosphorylation of eNOS. Oxidized LDL, but not native LDL, or HDL depleted caveolae of cholesterol by serving as an acceptor for cholesterol. Cyclodextrin also depleted caveolae of cholesterol and caused eNOS and caveolin to translocate from caveolae. Furthermore, removal of oxLDL allowed eNOS and caveolin to return to caveolae. We conclude that oxLDL-induced depletion of caveola cholesterol causes eNOS to leave caveolae and inhibits acetylcholine-induced activation of the enzyme. This process may be an important mechanism in the early pathogenesis of atherosclerosis. PMID- 10542297 TI - Catalytically active TYK2 is essential for interferon-beta-mediated phosphorylation of STAT3 and interferon-alpha receptor-1 (IFNAR-1) but not for activation of phosphoinositol 3-kinase. AB - TYK2, a Janus kinase, plays both structural and catalytic roles in type I interferon (IFN) signaling. We recently reported (Rani, M. R. S., Gauzzi, C., Pellegrini, S., Fish, E., Wei, T., and Ransohoff, R. M. (1999) J. Biol. Chem. 274, 1891-1897) that catalytically active TYK2 was necessary for IFN-beta to induce the beta-R1 gene. We now report IFN-beta-mediated activation of STATs and other components in U1 (TYK2-null) cell lines that were complemented with kinase negative (U1.KR930) or wild-type TYK2 (U1.wt). We found that IFN-beta induced phosphorylation on tyrosine of STAT3 in U1.wt cells but not in U1.KR930 cells, whereas STAT1 and STAT2 were activated in both cell lines. Additionally, IFN-beta mediated phosphorylation of interferon-alpha receptor-1 (IFNAR-1) was defective in IFN-beta treated U1.KR930 cells, but evident in U1.wt cells. In U1A-derived cells, the p85/p110 phosphoinositol 3-kinase isoform was associated with IFNAR-1 but not STAT3, and the association was ligand-independent. Further, IFN-beta treatment stimulated IFNAR-1-associated phosphoinositol kinase activity equally in either U1.wt or U1.KR930 cells. Our results indicate that catalytically active TYK2 is required for IFN-beta-mediated tyrosine phosphorylation of STAT3 and IFNAR-1 in intact cells. PMID- 10542299 TI - Involvement of DNA-dependent protein kinase in UV-induced replication arrest. AB - Cells exposed to UV irradiation are predominantly arrested at S-phase as well as at the G(1)/S boundary while repair occurs. It is not known how UV irradiation induces S-phase arrest and yet permits DNA repair; however, UV-induced inhibition of replication is efficiently reversed by the addition of replication protein A (RPA), suggesting a role for RPA in this regulatory event. Here, we show evidence that DNA-dependent protein kinase (DNA-PK), plays a role in UV-induced replication arrest. DNA synthesis of M059K (DNA-PK catalytic subunit-positive (DNA-PKcs(+))), as measured by [(3)H]thymidine incorporation, was significantly arrested by 4 h following UV irradiation, whereas M059J (DNA-PKcs(-)) cells were much less affected. Similar results were obtained with the in vitro replication reactions where immediate replication arrest occurred in DNA-PKcs(+) cells following UV irradiation, and only a gradual decrease in replication activity was observed in DNA-PKcs(-) cells. Reversal of replication arrest was observed at 8 h following UV irradiation in DNA-PKcs(+) cells but not in DNA-PKcs(-) cells. Reversal of UV-induced replication arrest was also observed in vitro by the addition of a DNA-PK inhibitor, wortmannin, or by immunodepletion of DNA-PKcs, supporting a positive role for DNA-PK in damage-induced replication arrest. The RPA-containing fraction from UV-irradiated DNA-PKcs(+) cells poorly supported DNA replication, whereas the replication activity of the RPA-containing fraction from DNA-PKcs(-) cells was not affected by UV, suggesting that DNA-PKcs may be involved in UV-induced replication arrest through modulation of RPA activity. Together, our results strongly suggest a role for DNA-PK in S-phase (replication) arrest in response to UV irradiation. PMID- 10542300 TI - Echocardiography in stroke and thromboembolism: transoesophageal imaging for all? PMID- 10542301 TI - Tilt table testing in the diagnosis of unexplained syncope. PMID- 10542302 TI - Pregnancy does not adversely affect renal transplant function. AB - Women with functioning transplanted kidneys often become fertile again. Indeed, renal function, endocrine status and libido rapidly improve after renal transplantation, and 1:50 women of childbearing age become pregnant. However, there is concern regarding the haemodynamic changes of pregnancy, which could lead to a decline in graft function (temporary or permanent). We examined obstetric data and renal parameters in 29 patients and 33 pregnancies. Mean serum creatinine and creatinine clearance remained stable throughout pregnancy and 1 year postpartum. However, there was a significant increase in proteinuria from a mean of 0.45 g/24 h around the time of conception to 1.11 g/24 h at delivery (p<0.05). The proteinuria resolved to baseline levels at 3 months postpartum. We highlight certain parameters to be considered before conception to allow a good obstetric outcome and prolong stable renal function: serum creatinine <150 micromol/l, proteinuria <1 g/day, absence of histological evidence of chronic allograft rejection, controlled blood pressure (140/90) and stability of maintenance immunosuppression. PMID- 10542303 TI - Equity of renal replacement therapy utilization: a prospective population-based study. AB - This 1-year prospective survey assessed the incidence and characteristics of all patients starting renal replacement therapy (RRT) for end-stage renal disease in Scotland, and whether there is equity of utilization of RRT in terms of age, domicile and social circumstance. In the year studied, 104 patients per million population (533 patients) started RRT (390 per million population aged 65-75). In 23.5% the cause of ESRD could not be determined. Diabetes was the single most frequently identified cause (16%). The requirement for RRT rose with age, but over the country as a whole, patients aged over 75 years were under-represented. The majority of health boards provided RRT at a rate within 20% of the national rate. There was no difference in the median age at starting RRT between health boards. The spectrum of social deprivation of patients starting RRT was the same as that of the general population. There was no evidence that social deprivation influences acceptance on to the RRT program, although the relationship between ESRD and deprivation is complex. The utilization of RRT exceeded the minimum rate recommended by the Renal Association, although there was fluctuation between health board areas. The national requirement for resources to provide RRT is likely to rise further to care for an increasingly elderly population. PMID- 10542304 TI - Accuracy of CT scanning and adrenal vein sampling in the pre-operative localization of aldosterone-secreting adrenal adenomas. AB - In primary hyperaldosteronism, it is important to distinguish between unilateral and bilateral disease, as management strategies differ. In the period 1983-95, we identified 34 patients with primary hyperaldosteronism. Following further investigations, a diagnosis of aldosterone-secreting adenoma was made in 17 patients, and surgery was performed. Computed tomography clearly localized an apparent adenoma (discrete adenoma=1 cm diameter; normal contralateral gland) in only 10 of these patients (59%); two of these 'adenomas' were subsequently shown to be hyperplastic glands without adenomas. Histological examination showed adrenal adenomas in the remaining 15 patients. An 'adenoma' also appeared to be clearly localized in 3/17 patients later classified as having bilateral adrenal hyperplasia by adrenal vein sampling. CT scanning, therefore clearly localizes adenomas in only 50% of histologically proven cases, and can also produce misleading results. Adrenal vein sampling results altered our management approach in one third of cases. On the basis of our detailed results we would recommend surgery if there is clear evidence of unilateral aldosterone secretion along with CT findings which may not be strictly localizing but are in keeping with the dominant side on adrenal vein sampling. The decision to refer for surgery in primary hyperaldosteronism can be difficult, and we would caution against too heavy a reliance on CT results when recommending adrenalectomy, and suggest that adrenal vein sampling should remain a routine part of the investigation of patients with primary hyperaldosteronism. PMID- 10542305 TI - Titres of anti-neutrophil cytoplasmic antibodies in inflammatory bowel disease are not related to disease activity. AB - In the systemic vasculitides, serial measurement of titres of anti-neutrophil cytoplasmic autoantibodies (ANCA) is useful for follow-up of disease activity and prediction of relapses. ANCA have been detected in patients with inflammatory bowel disease, but their relation to disease activity in these diseases is unclear. We analysed the relation between disease activity and ANCA titres as determined by indirect immunofluorescence in paired samples obtained during active disease and at remission from individual patients with ulcerative colitis (n=60) and Crohn's disease (n=101). In addition, patients were followed prospectively, to study the fluctuations of ANCA with time in relation to disease activity. We did not detect a correlation between disease activity and ANCA titres, either in paired samples from active disease and remission, or in serial samples, either in ulcerative colitis or in Crohn's disease. In contrast to the ANCA-associated systemic vasculitides, serial measurement of ANCA titres is not useful in the monitoring of disease activity in patients with inflammatory bowel disease. PMID- 10542307 TI - Decision analysis and the implementation of evidence-based medicine. AB - The evidence-based medicine movement has received enthusiastic endorsement from editors of major medical journals. Hardly anyone can disagree with the aim of helping clinicians to make judicious use of the best scientific evidence for decisions in patient care. Evidence-based medicine, however, because of its dependence on randomized trials, cannot be applied to all individuals seen in daily practice. Specifically, patients may differ in age, severity of illness, presence of comorbidity and myriad of other clinical nuances. In response to these limitations, decision analysis, a technique which allows to consider multiple health outcomes, such as the patient's preferences for different states of health, and to measure the consequences of many strategies for which randomized trials are not feasible, provides a rational means of allowing health professionals to move from finding evidence to implementing it. Such formal approach may reconcile evidence-based medicine with 'real life' and patient's preference. It should therefore be considered complementary to evidence-based medicine. PMID- 10542306 TI - Out-patient parenteral antimicrobial therapy--a viable option for the management of cutaneous leishmaniasis. AB - Cutaneous infection with Leishmania braziliensis complex requires treatment with parenteral pentavalent antimonials to prevent development of mucocutaneous leishmaniasis. Patients with imported disease are usually managed in hospital because of concerns over drug toxicity. This study describes the clinical features and outcome of infection treated in the UK in an out-patient setting. Thirteen marines (aged 19-35 years) who acquired leishmaniasis in Belize were studied prospectively. Three had at least two lesions (0. 6-3 cm diameter), eight had regional lymphadenopathy and one had localized painless lymphatic thickening. Histology for amastigotes and PCR for Leishmania braziliensis complex was positive in all. Culture was positive in six. Patients received 1.5-2 g (mean 1.7 g) (20 mg/kg) sodium stibogluconate intravenously daily for 20 days. All developed transient musculoskeletal symptoms and asymptomatic hepatitis. Eleven developed biochemical pancreatitis, and one thrombocytopenia. Three developed transient ECG changes and one herpes zoster. There were four device-related infections, two requiring hospitalization (one required surgical drainage of an abscess). All lesions re-epithelialized. A total of 250 bed-days were saved over a 67-day period. These results indicate that in selected patients, out-patient therapy for cutaneous leishmaniasis with parenteral high-dose sodium stibogluconate may be appropriate, provided there is adequate monitoring of therapy. PMID- 10542309 TI - Lower cardiac mortality in smokers following thrombolysis. PMID- 10542308 TI - Beta-blockers for heart failure--time to think the unthinkable? PMID- 10542310 TI - Cloning and expression profile of human inorganic pyrophosphatase. AB - We have cloned a 1.23 kb cDNA from a human heart library which encodes a 32 kDa protein that is 94% identical to bovine inorganic pyrophosphatase. The protein contains an aspartate-rich signature sequence that was previously identified in yeast and prokaryotic pyrophosphatases. Our clone detects a single band on Northern blots and is expressed at modest levels in all tissues examined. The cDNA shows linkage to markers on the long arm of chromosome 10. PMID- 10542311 TI - DNA triple helix stabilisation by covalent attachment of a triplex-specific ligand. AB - We have prepared oligonucleotides with a naphthylquinoline triplex-binding ligand covalently tethered to the 5'-end and have used UV-melting and DNase I footprinting to examine the stability of intra- and inter-molecular triplexes containing this modification. We find that covalent attachment of the ligand increases the melting temperature of intramolecular 6-mer triplexes by about 14 K, and increases the binding of 9-mer oligonucleotides to their duplex target sites by about 60-fold. PMID- 10542312 TI - Metazoan cellulase genes from termites: intron/exon structures and sites of expression. AB - Endogenous endo-beta-1,4-glucanase (EGase, EC 3.2.1.4) cDNAs were cloned from representatives of the termite families Termitidae and Rhinotermitidae. These EGases are all composed of 448 amino acids and belong to glycosyl hydrolase family 9 (GHF9), sharing high levels of identity (40-52%) with selected bacterial, mycetozoan and plant EGases. Like most plant EGases, they consist of a single catalytic domain, lacking the ancillary domains found in most microbial cellulases. Using a PCR-based strategy, the entire sequence of the coding region of NtEG, a gene putatively encoding an EGase from Nasutitermes takasagoensis (Termitidae), was determined. NtEG consists of 10 exons interrupted by 9 introns and contains typical eukaryotic promoter elements. Genomic fragments of EGase genes from Reticulitermes speratus (Rhinotermitidae) were also sequenced. In situ hybridization of N. takasagoensis guts with an antisense NtEG RNA probe demonstrated that expression occurs in the midgut, which contrasts to EGase expression being detected only in the salivary glands of R. speratus. NtEG, when expressed in Escherichia coli, was shown to have in vitro activity against carboxymethylcellulose. PMID- 10542313 TI - Functional activity of hepatocyte nuclear factor-1 is specifically decreased in amino acid-limited hepatoma cells. AB - Limitation of cultured rat hepatoma cells for an essential amino acid results in a specific decrease in expression of several genes that are preferentially expressed in the liver, including the serum albumin and transthyretin genes. In the work presented here, we examined whether the coordinate repression of these genes is caused by decreased activity of one or more of the liver-enriched transcription factors, hepatocyte nuclear factor-1 (HNF-1), HNF-3, HNF-4 or C/EBP. To address this question, HepG2 human hepatoma cells were transiently transfected with luciferase reporter constructs containing multiple copies of individual transcription factor binding sites. Limitation for an essential amino acid resulted in specific repression of a construct in which luciferase expression was directed by HNF-1. A single HNF-1 binding site located adjacent to the TATA box plays a major role in transcription directed by the serum albumin promoter in transient transfection assays. Amino acid limitation of cells transfected with an albumin promoter/luciferase reporter construct resulted in specific repression of promoter activity. In addition, bacterial methylation or site-directed mutagenesis of the HNF-1 binding site in the albumin proximal promoter region eliminated the regulation of an albumin promoter-luciferase reporter construct under conditions of amino acid limitation. These results demonstrated that the HNF-1 binding site played a major role in regulation of the albumin promoter by amino acid availability. Deletion analysis of the albumin promoter confirmed regulation through the HNF-1 binding site and also identified a second amino acid regulatory element in the upstream region of the albumin promoter, which has been shown previously to contain a functional binding site for HNF-3. The repression of albumin promoter and HNF-1 reporter constructs in amino acid-limited cells occurred without a change in the DNA binding activity of HNF-1. Moreover, HNF-3 DNA binding activity was also not decreased in amino acid limited cells. These results suggest that the regulation of transcription by amino acids occurs at the level of transcriptional activation by HNF-1 and HNF-3, rather than by alteration of the DNA binding activity of either factor. PMID- 10542314 TI - The v-Myb oncoprotein activates C/EBPbeta expression by stimulating an autoregulatory loop at the C/EBPbeta promoter. AB - Previous studies have implicated the CCAAT box/enhancer binding protein beta (C/EBPbeta) in the regulation of cell-type specific gene expression in myelomonocytic cells and in the activation of target genes by the transcription factor v-Myb. To better understand the role of C/EBPbeta in myelomonocytic cells we have cloned the chicken C/EBPbeta gene and studied its regulation. The chicken C/EBPbeta promoter contains a number of C/EBP binding sites and is activated by C/EBPbeta, suggesting that the C/EBPbeta gene is autoregulated by its own protein product. Interestingly, the C/EBPbeta promoter is not activated by C/EBPalpha, another C/EBP family member highly expressed in myelomonocytic cells, indicating that the autoregulation is specific for C/EBPbeta. Comparison of different C/EBP inducible promoters shows that the relative transactivation potential of C/EBPalpha and beta is extremely dependent on the promoter context. By using the promoters of the mim-1 and C/EBPbeta genes and by exchanging the DNA-binding domains between C/EBPalpha and beta we show that the observed promoter preferences of C/EBPalpha and beta are not due to differential DNA-binding but instead depend on the transactivation domains of these proteins. The C/EBPbeta promoter also contains several Myb binding motifs, suggesting that the C/EBPbeta gene is also myb-inducible. We show that the C/EBPbeta promoter is activated synergistically by v-Myb and C/EBPbeta and that transcription of the endogenous C/EBPbeta gene is increased by v-Myb. Thus, our results identify the C/EBPbeta gene as a novel v-Myb target gene. Taken together, our data suggest a model for the regulation of C/EBPbeta expression in which v-Myb stimulates the synthesis of C/EBPbeta by enhancing an autoregulatory loop acting on the C/EBPbeta promoter. PMID- 10542315 TI - Molecular markers for UV-B stress in plants: alteration of the expression of four classes of genes in Pisum sativum and the formation of high molecular mass RNA adducts. AB - Sixteen ultraviolet-B radiation-regulated pea genes were identified. Functionally, the corresponding proteins were divided into four groups. (i) Chloroplast-localized proteins. Genes for these proteins were down-regulated, underlining the deleterious effects of UV-B on this organelle. A novel down regulated photosystem I light-harvesting chlorophyll a/b-binding protein gene (PsLhcA4), was cloned and sequenced. (ii) Protein turnover enzymes. Levels of mature mRNAs for the PU1 and PsUBC4 genes, encoding proteins of the ubiquitin protein degradation pathway, were up- and down-regulated, respectively, implying alteration of plant cell protein content by changes in both gene expression and protein degradation. (iii) Proteins involved in intracellular signalling. Expression of genes for small GTPases, rab and rho homologues, were altered. (iv) Phenylpropanoid or flavonoid biosynthesis. Expression of three genes encoding enzymes in these pathways were up-regulated and one of them, the novel PsC450R1, was cloned and sequenced. Moreover, unexpected high molecular mass psbA RNA adducts were found to appear after UV-B exposure. In addition, a large increase in corresponding high molecular mass adducts were also found for PsLhcA4, and PsUBC4 mRNA and 23S rRNA. These RNA species do not contain protein and probably appear due to cross-linking of two or more RNA molecules, or are the result of UV B-induced failure of transcription termination. PMID- 10542316 TI - cDNA cloning and expression analysis of mouse zf9, a Kruppel-like transcription factor gene that is induced by adipogenic hormonal stimulation in 3T3-L1 cells. AB - Using a differential hybridization method, we have cloned a zinc finger transcription factor gene whose expression was enhanced by adipogenic hormones in preadipocyte 3T3-L1 cells. Cloning of this gene revealed that it encodes a mouse homologue of rat Zf9 and human CPBP/GBF, previously identified as a wound-induced transcription factor and GC-rich binding protein, respectively. The mRNA for this clone consisted of 0.9 kb coding region and 3.2 kb long 3' untranslated region. Northern blot analysis revealed that it was ubiquitously expressed, among adult tissues, in which abundant expression was observed in lung, ovary and thymus. The transcript was transiently induced by different stimuli such as serum, cAMP and 12-O-tetradecanoylphorbol 13-acetate. Nuclear run-on and RNA synthesis inhibitor chase experiments indicated that the transient induction of the mRNA was regulated both at transcriptional and post-transcriptional levels. PMID- 10542318 TI - Polyethylenimine shows properties of interest for cystic fibrosis gene therapy. AB - Before being considered for a cystic fibrosis (CF) gene therapy trial, any gene delivery agent must be able to show that it produces low levels of toxicity as well as being able to protect the DNA from nuclease degradation. Here we show that complexes of linear polyethylenimine (L-PEI) and DNA can repeatedly be administered to animals (up to 21 consecutive days) without eliciting an immune response against PEI/DNA particles or inducing toxic side effects due to accumulation of PEI in the lungs. However, the host response to the exogenous protein resulted in some decrease of expression. PEI-mediated transfection was unaffected by treatment of the complexes with DNase (frequently used to reduce the viscosity of lung secretions in CF patients). Taken together, these properties make L-PEI a valuable vector for gene therapy of CF. PMID- 10542317 TI - Identification of an oxygen responsive enhancer element in the glyceraldehyde-3 phosphate dehydrogenase gene. AB - The glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is induced by hypoxia in endothelial cells (EC). Upregulation occurs primarily at the level of transcription and occurs to a much greater extent in EC than in other cell types. To characterize EC specific hypoxia response elements within the GAPDH gene, we performed transient transfection studies in EC, fibroblasts and smooth muscle cells using portions of the GAPDH promoter linked to a CAT reporter gene. These initial studies identified an EC specific hypoxia responsive region that was further characterized (using SV40-promoter-CAT reporter constructs) as a 19 nucleotide sequence (-130 to -112) containing both an hypoxia inducible factor-1 (HIF-1)-binding site and a novel flanking sequence. Electrophoretic mobility shift assays confirmed inducible EC protein binding to this fragment. Mutation of either the HIF-1-binding site or the flanking sequence resulted in complete loss of function and loss of inducible protein binding. Thus, a single HIF-1-binding site is necessary, but not sufficient, for hypoxic regulation of GAPDH in EC. Furthermore, the novel HIF-1 flanking sequence required for GAPDH upregulation and the protein(s) that bind to it may be EC specific. PMID- 10542319 TI - Promoter analysis of the membrane protein gp64 gene of the cellular slime mold Polysphondylium pallidum. AB - We cloned a genomic fragment of the membrane protein gp64 gene of the cellular slime mold Polysphondylium pallidum by inverse PCR. Primer extension analysis identified a major transcription start site 65 bp upstream of the translation start codon. The promoter region of the gp64 gene contains sequences homologous to a TATA box at position -47 to -37 and to an initiator (Inr, PyPyCAPyPyPyPy) at position -3 to +5 from the transcription start site. Successively truncated segments of the promoter were tested for their ability to drive expression of the beta-galactosidase reporter gene in transformed cells; also the difference in activity between growth conditions was compared. The results indicated that there are two positive vegetative regulatory elements extending between -187 and -62 bp from the transcription start site of the gp64 promoter; also their activity was two to three times higher in the cells grown with bacteria in shaken suspension than in the cells grown in an axenic medium. PMID- 10542321 TI - Characterization of the regulatory region of the human testis-specific form of the pyruvate dehydrogenase alpha-subunit (PDHA-2) gene. AB - The alpha-subunit of human pyruvate dehydrogenase (E(1)) is encoded by two separate genes. The gene located on chromosome X (PDHA-1) is expressed in somatic tissues, whereas the second gene (PDHA-2), located on chromosome 4, is expressed only in post-meiotic spermatogenic cells. A genomic fragment harboring the human gene encoding PDHA-2 has been isolated and approximately 800 nucleotides of the promoter region have been characterized. Functional studies of the promoter indicate the presence of both enhancer and repressor elements that are common to other genes that are only expressed in mature sperm. PMID- 10542320 TI - Positive and negative regulation of the human heme oxygenase-1 gene expression in cultured cells. AB - To elucidate the regulation of the human heme oxygenase-1 (hHO-1) gene expression, we assessed approximately 4 kb of the 5'-flanking region of the hHO-1 gene for basal promoter activity and sequenced approximately 2 kb of the 5' flanking region. A series of deletion mutants of the 5'-flanking region linked to the luciferase gene was constructed. Basal level expression of these constructs was tested in HepG2 human hepatoma cells and HeLa cervical cancer cells. By measuring luciferase activity, which was transiently expressed in the transfected cells, we found a positive regulatory region at position -1976 to -1655 bp. This region functions in HepG2 cells but not in HeLa cells. A negative regulatory region was also found at position -981 to -412 bp that functions in both HepG2 cells and HeLa cells. PMID- 10542322 TI - Molecular cloning and characterization of the promoter region of murine natural killer cell receptor 2B4. AB - Natural killer (NK) cells are bone marrow-derived lymphocytes that have the ability to kill certain tumor cells and virally infected cells. The activation of NK cells is mediated by a balance of negative and positive signals from cell-cell interactions and from responses to cytokines. However, the molecular basis of NK cell activation and recognition of target cells is poorly understood. We have previously identified, cloned and characterized a receptor, 2B4, expressed on murine NK cells. 2B4 is not only expressed on all NK cells, but also on a subset of T-cells which have NK-like killing properties. Structural analysis indicated that 2B4 belongs to the CD2 subset of immunoglobulin superfamily. We have also shown 2B4 to interact with CD48 with nine times more affinity than that of CD2 CD48 interaction. In order to understand the transcriptional regulation as well as the mechanisms controlling the restricted expression of the 2B4 gene, we obtained a genomic 2B4 clone including the sequence of the 5'-flanking region. To define the start site of transcription, we performed primer extension and 5'-RACE assays and found that the 2B4 gene may be initiated at multiple start sites and driven by a TATA-less promoter. Transient transfections of nested 5'-fragments of the 2B4 promoter to drive CAT expression revealed tissue specific expression in CTLL-2 cells, a mouse T-cell line. A promoter fragment of 348 bases upstream from the first base of the mouse 2B4 cDNA clone p2B4.8 produced maximal CAT activity in CTLL-2 cells. The presence of the region -653 to -540 on the other hand, drastically reduced transcription. Sequence analysis of this promoter region has identified potential recognition motifs for a number of lymphocyte-restricted in addition to ubiquitous transcription factors, which may play a role in the transcriptional regulation of the mouse 2B4 gene. PMID- 10542323 TI - cDNA cloning, genomic structure and chromosomal localization of the human BUP-1 gene encoding beta-ureidopropionase. AB - A full-length cDNA clone encoding human beta-ureidopropionase was isolated. A 1152-nucleotide open reading frame which corresponds to a protein of 384 amino acids with a calculated molecular weight of 43? omitted?158 Da, surrounded by a 5'-untranslated region of 61 nucleotides and a 3'-untranslated region of 277 nucleotides was identified. The protein showed 91% similarity with the translation product of the rat beta-ureidopropionase cDNA. Expression of the human cDNA in an Escherichia coli and eukaryotic COS-7 expression system revealed a very high beta-ureidopropionase enzymatic activity, thus confirming the identity of the cDNA. Since human EST libraries from brain, liver, kidney and heart contained partial beta-ureidopropionase cDNAs, the enzyme seems to be expressed in these tissues, in agreement with the expression profile of this enzyme in rat. Using the human cDNA as a probe a genomic P1 clone could be isolated containing the complete human beta-ureidopropionase gene. The gene consist of 11 exons spanning approximately 20 kB of genomic DNA. Fluorescence in situ hydridization localized the human beta-ureidopropionase gene to 22q11.2. PMID- 10542324 TI - Membrane-bound transferrin-like protein (MTf): structure, evolution and selective expression during chondrogenic differentiation of mouse embryonic cells. AB - Mouse membrane-bound transferrin-like protein (MTf) cDNA was cloned to examine its expression during chondrogenic differentiation in the mouse embryonic cell line ATDC5, and to analyze the phylogenetic relationships among the MTfs of four animal species and 23 other transferrin members. Phylogenetic analysis indicated that the MTf gene diverged from the common ancestor gene earlier than the genes of the other transferrins such as serum transferrin, lactoferrin and ovotransferrin, and that the divergence occurred after the divergence of vertebrates and invertebrates. MTf, as well as the other transferrins, consists of two repeated domains. The similarity between the N-terminal and the C-terminal domains of MTf is much higher than that of the other transferrins, although the five amino acid residues required for iron binding were not conserved in the C terminal domain of MTf in contrast to the conservation of these residues in both domains of the other transferrins. Among various adult mouse tissues, MTf mRNA was expressed at the highest level in cartilage and at a moderate level in the testis. MTf mRNA was expressed only at very low levels in the brain, spleen, thymus, muscle, lung, skin and intestine, and hardly detected in the heart, kidney, stomach and liver. In cultures of the mouse ATDC5 cell line, MTf is developmentally expressed in parallel with the expression of type II collagen and aggrecan, in the pattern commensurate with the onset of chondrogenesis to form cartilage nodules. The structural characteristics and the expression pattern suggest that during development and in adult tissues, MTf has some functions that are different from those of other transferrins. PMID- 10542325 TI - Cloning, tissue-specific expression, gene structure and chromosomal localization of human phosphatidylcholine transfer protein. AB - Phosphatidylcholine transfer protein (PC-TP) is a cytosolic protein that catalyzes intermembrane transfer of phosphatidylcholines in vitro. We have cloned a cDNA encoding the human ortholog of PC-TP and have determined its tissue specific expression as well as genomic organization. Radiation hybrid mapping localized the human gene, PCTP, to chromosome 17q21-22 and PCR-based single strand conformation polymorphism analysis of an interspecific backcross assigned mouse Pctp to the region of syntenic conservation on chromosome 11. PMID- 10542326 TI - Cloning, sequencing and differential expression of alphaB-crystallin in the zebrafish, Danio rerio. AB - Here we report the cloning and expression of alphaB-crystallin from the zebrafish. 5'- and 3'-RACE was used to isolate a 900-bp transcript that contained insertions and deletions that differentiate it from both alphaA-crystallin and HSP-27. The deduced amino acid sequence of zebrafish alphaB-crystallin revealed that it lacked four residues in the C-terminus implicated in protein-protein interactions in other vertebrate species. In addition, the sequence contained two substitutions at sites implicated in phosphorylation in other vertebrate species. Northern analysis and semi-quantitative RT-PCR indicate that zebrafish alphaB crystallin is expressed at extremely low levels outside of the lens. PMID- 10542327 TI - Isolation, cloning, and expression of a new murine zinc finger encoding gene. AB - With the aim of identifying genes involved in cartilage differentiation, we have used a subtractive hybridization strategy with cDNAs from a chondrocytic cell line (MC615) and mRNAs from a mesenchymal precursor cell line (10T1/2). We have isolated a cDNA clone representing a novel mouse gene. The predicted 368-amino acid protein, designated ZF-12, contains four C(2)H(2)-type zinc finger motifs and one region homologous to the LeR domain, a finger-associated structural domain. ZF-12 mRNAs are expressed during embryonic development and in different organs in adult, including rib cartilage. These data suggest that ZF-12 might play an important role not only in cartilage differentiation, but also in basic cellular processes. PMID- 10542328 TI - Characterization of mouse glycogenin-1 cDNA and promoter region. AB - Glycogenin-1 is an autocatalytic, self-glucosylating protein which acts as the primer for glycogen synthesis in mammalian skeletal muscle. In this study, we have cloned the glycogenin-1 cDNA from mouse skeletal muscle. Mouse glycogenin-1 has a predicted molecular mass of 37? omitted?399 Da, and the deduced amino acid sequence exhibited 87% homology with human glycogenin-1. Northern blot analysis specifically detected mouse glycogenin-1 transcript in skeletal muscle and heart, and to a lesser extent in kidney, lung and brain. 5'-RACE analysis revealed the major transcription start site to be localized 47 bp upstream of the initiation of translation codon. Sequence analysis of approximately 2 kb of the 5'-flanking region revealed potentially important regions of homology between the mouse and human glycogenin-1 promoters. Several conserved but putative elements, including a TATA box, Sp1 site, and a cyclic AMP responsive element, were observed proximal to the transcription start site. Significantly, Northern blot analysis revealed dibutyryl-cAMP treatment of cultured mouse C2C12 myotubes markedly reduced the levels of glycogenin-1 mRNA in a dose-dependent manner. PMID- 10542329 TI - PLP-I: a novel prolactin-like gene in rodents. AB - In this report, we describe molecular cloning and characterization of cDNAs encoding a novel rat prolactin-like protein. The rat cDNAs were isolated from the decidua and the gene was named PLP-I. cDNAs for the mouse equivalent were also cloned by the cross-hybridization technique. Pregnancy-specific expression of the rat PLP-I gene was observed in the rat placenta by Northern analysis. Location of signal peptide cleavage sites in rat and mouse pre-PLP-I proteins was predicted using a theoretical method. A molecular phylogenetic tree for the growth hormone prolactin superfamily including the novel member, PLP-I, constructed using the neighbor-joining method, places rat/mouse PLP-I closest to rat/mouse placental lactogen I and II. PMID- 10542330 TI - Analysis and regulation of the secY gene(1) from Streptomyces griseus N2-3-11 and interaction of the SecY protein with the SecA protein. AB - The chromosomal region encoding the secY gene of Streptomyces griseus N2-3-11 was cloned and analyzed. The secY gene encodes a polypeptide of 438 aa with a molecular mass of 47.5 kDa. The transcriptional start point of the secY gene was determined. Northern blot analysis revealed a growth phase-dependent secY expression supporting our previous findings for secA gene expression in S. griseus. Overproduction of the SecY protein was obtained when using Streptomyces lividans TK23 as host. The interaction of the SecY proteins of S. griseus, S. lividans, and Escherichia coli, respectively, with the purified SecA protein of S. griseus was demonstrated for the first time by using ligand affinity blot assays. PMID- 10542331 TI - Identification of the true human orthologue of the mouse Zp1 gene: evidence for greater complexity in the mammalian zona pellucida? AB - The mammalian zona pellucida is a mixture of glycoproteins, believed to be encoded by three distinct genes, ZP1/ZPB, ZP2/ZPA, and ZP3/ZPC. We have now determined that the true human orthologue of the mouse Zp1 gene is not ZPB, but that there is a distinct human ZP1 gene. Comparison of the human ZP1 and murine Zp1 genes indicates significant conservation of nucleotide and amino acid sequences, of intron-exon size and organisation, and of regulatory sequences. In addition, the mouse and human ZP1 genes are in a region of conserved synteny between human chromosome 11 and mouse chromosome 19. PMID- 10542332 TI - Genetic linkage and radiation hybrid mapping of the three human GABA(C) receptor rho subunit genes: GABRR1, GABRR2 and GABRR3. AB - GABA(C) receptors mediate rapid inhibitory neurotransmission in retina. We have mapped, in detail, the human genes which encode the three polypeptides that comprise this receptor: rho1 (GABRR1), rho2 (GABRR2) and rho3 (GABRR3). We show that GABRR1 and GABRR2 are located close together, in a region of chromosome 6q that contains loci for inherited disorders of the eye, but that GABRR3 maps to chromosome 3q11-q13.3. Our mapping data suggest that the rho polypeptide genes, which are thought to share a common ancestor with GABA(A) receptor subunit genes, diverged at an early stage in the evolution of this gene family. PMID- 10542333 TI - Alternative splicing of the Drosophila PTEN gene. AB - Mutations in the human PTEN gene have been identified in a number of different tumour types, and in the hamartomatous polyposis syndromes Cowden disease and Bannayan-Zonana syndrome. The PTEN gene encodes a phosphatase that antagonises phosphoinositide 3-kinase (PI3K) signalling by removing the 3' phosphate from phosphatidylinositol 3, 4,5-trisphosphate (PtdIns (3,4,5)P(3)). Here we show that the PTEN gene is conserved in the invertebrate Drosophila melanogaster and demonstrate that the gene undergoes alternative splicing. PMID- 10542335 TI - Structural analysis of phylogenetically conserved J domain protein gene. AB - Novel cDNAs encoding evolutionarily conserved J Domain Proteins (JDPs) were investigated from Drosophila and mouse. Each of the full coding sequences potentially encodes a conserved J domain, but lacks additional characteristic structures present in DnaJ family proteins. The expression was restricted to head in Drosophila. However, ubiquitous expression was observed in mice with the highest level in kidney. PMID- 10542334 TI - Molecular cloning of rabbit CARP cDNA and its regulated expression in adriamycin cardiomyopathy. AB - A full-length rabbit cDNA of cardiac adriamycin responsive protein (CARP) has been cloned. It shows high levels of identity at the amino acid sequence level (>86%) with the rat, mouse and human homologues. CARP mRNA levels are highly regulated in adriamycin-cardiomyopathy in rabbits. PMID- 10542336 TI - Sequence and expression of the monkey homologue of the ER-golgi intermediate compartment lectin, ERGIC-53. AB - We obtained the cDNA sequence of the monkey homologue of the intermediate compartment protein ERGIC-53 by both cDNA library screening and RT-PCR amplification. The final sequence of 2422 nts of the monkey ERGIC-53 cDNA is 96.2% identical to the human ERGIC-53 cDNA and 87% and 67% identical to the rat and amphibian cDNA, respectively. The translated CV1 ERGIC-53 protein is 96.47% identical to the human ERGIC-53, 87% identical to the rat p58 and 66. 98% to the Xenopus laevis protein. Southern blot analysis of multiple genomic DNAs shows the presence of sequences similar to ERGIC-53 in different species. ERGIC-53 is expressed as a major transcript of about 5.5 kb in either monkey CV1 or in human CaCo2. A shorter transcript of 2.3 kb was detected in both cell lines and in mRNAs derived from human pancreas and placenta. PMID- 10542337 TI - Human alpha-catulin, a novel alpha-catenin-like molecule with conserved genomic structure, but deviating alternative splicing. AB - A new human cDNA was cloned and termed alpha-catulin, based on sequence similarity with both alpha-CATenins and vincULIN. The mRNA is present ubiquitously, although low expression levels are found in neural tissues. The genomic organization of the alpha-catulin gene CTNNAL1 is closely related to that of the alphaE-catenin gene CTNNA1, but not at all to that of the vinculin gene. Alternative splicing of the last exon generates a frameshift, resulting in a truncated protein with a new carboxy-terminus. PMID- 10542338 TI - Isolation of a zeta class wheat glutathione S-transferase gene. AB - A new Zeta class glutathione S-transferases (GST) gene, pGST, has been cloned from wheat for the first time by the differential display PCR (DD-PCR) method. The genomic sequence of pGST, TA-GSTZ1, contains nine exons that encode a polypeptide of 213 amino acids and eight introns. The deduced amino acid sequence of TA-GSTZ1 as well as the exon:intron placement are more similar to the GSTs of the Zeta class than to the two wheat GSTs reported earlier. The pGST cDNA gene product expressed in Escherichia coli and purified by affinity chromatography showed typical Zeta class GST and glutathione peroxidase activities. Sequence polymorphism in the 3' untranslated region (UTR) of TA-GSTZ1 gene in wheat has been discovered. In this study, an 89 bp sequence is present in the 3' UTR of TA GSTZ1gene in 16 wheat cultivars but absent in the other five. Although the biological importance of this polymorphism is unknown, it can be useful as a genetic marker in wheat breeding. PMID- 10542339 TI - Identification and characterization of five cspA homologous genes from Myxococcus xanthus. AB - Escherichia coli contains a large CspA family consisting of nine homologues, in which four are cold-shock inducible and one is stationary-phase inducible. Here, we demonstrate that Myxococcus xanthus possesses at least five CspA homologues, CspA to CspE. Hydrophobic residues forming a hydrophobic core, and aromatic residues, which are included in functional motifs RNP-1 and RNP-2 involved in binding to RNA and ssDNA, are well conserved. These facts suggest that M. xanthus CspA homologues have a similar structure and function as E. coli CspA. However, in contrast to the E. coli CspA family, the expression of M. xanthus csp genes as judged by primer extension analysis is not significantly regulated by temperature changes, except for cspB of which expression was reduced to less than 10% upon heat shock at 42 degrees C. Such constitutive expression of the csp genes may be important for M. xanthus, a soil-dwelling bacterium, to survive under conditions of exposure to various environmental changes in nature. PMID- 10542340 TI - Unaliasing by fourier-encoding the overlaps using the temporal dimension (UNFOLD), applied to cardiac imaging and fMRI. AB - In several applications, MRI is used to monitor the time behavior of the signal in an organ of interest; e.g., signal evolution because of physiological motion, activation, or contrast-agent accumulation. Dynamic applications involve acquiring data in a k-t space, which contains both temporal and spatial information. It is shown here that in some dynamic applications, the t axis of k t space is not densely filled with information. A method is introduced that can transfer information from the k axes to the t axis, allowing a denser, smaller k t space to be acquired, and leading to significant reductions in the acquisition time of the temporal frames. Results are presented for cardiac-triggered imaging and functional MRI (fMRI), and are compared with data obtained in a conventional way. The temporal resolution was increased by nearly a factor of two in the cardiac-triggered study, and by as much as a factor of eight in the fMRI study. This increase allowed the acquisition of fMRI activation maps, even when the acquisition time for a single full time frame was actually longer than the paradigm cycle period itself. The new method can be used to significantly reduce the acquisition time of the individual temporal frames in certain dynamic studies. This can be used, for example, to increase the temporal or spatial resolution, increase the spatial coverage, decrease the total imaging time, or alter sequence parameters e.g., repetition time (TR) and echo time (TE) and thereby alter contrast. Magn Reson Med 42:813-828, 1999. PMID- 10542341 TI - Titration of the BOLD effect: separation and quantitation of blood volume and oxygenation changes in the human cerebral cortex during neuronal activation and ferumoxide infusion. AB - Most functional magnetic resonance imaging (fMRI) techniques are sensitive to susceptibility variations and rely on the change in blood oxygenation level in response to neuronal activation (BOLD). The BOLD effect is accompanied by a change in cerebral blood flow (rCBF) and cerebral blood volume (rCBV). Intravascular contrast agents, such as magnetite nanoparticles, can be used to measure changes in rCBV. A new measuring protocol has been developed that enables the separate quantification of changes in blood volume and oxygenation levels. A combination of alternating acoustic stimulation blocks and infusion of a superparamagnetic contrast agent offers the possibility to disentangle the competing influences of oxygenation and blood volume changes. Serial blood sampling during infusion was used to assess the actual contrast agent concentration during infusion in order to calculate absolute blood volume changes during neuronal resting and activation states. Magn Reson Med 42:829-836, 1999. PMID- 10542342 TI - Coronary venous oximetry using MRI. AB - Based on the Fick law, coronary venous blood oxygen measurements have value for assessing functional parameters such as the coronary flow reserve. At present, the application of this measure is restricted by its invasive nature. This report describes the design and testing of a noninvasive coronary venous blood oxygen measurement using MRI, with a preliminary focus on the coronary sinus. After design optimization including a four-coil phased array and an optimal set of data acquisition parameters, quality tests indicate measurement precision on the order of the gold standard optical measurement (3%O(2)). Comparative studies using catheter sampling suggest reasonable accuracy (3 subjects), with variability dominated by sampling location uncertainty ( approximately 7%O(2)). Intravenous dipyridamole (5 subjects) induces significant changes in sinus blood oxygenation (22 +/- 9% O(2)), corresponding to flow reserves of 1.8 +/- 0.4, suggesting the potential for clinical utility. Underestimation of flow reserve is dominated by right atrial mixing and the systemic effects of dipyridamole. Magn Reson Med 42:837-848, 1999. PMID- 10542343 TI - Investigation of BOLD signal dependence on cerebral blood flow and oxygen consumption: the deoxyhemoglobin dilution model. AB - The relationship between blood oxygenation level-dependent (BOLD) MRI signals, cerebral blood flow (CBF), and oxygen consumption (CMR(O2)) in the physiological steady state was investigated. A quantitative model, based on flow-dependent dilution of metabolically generated deoxyhemoglobin, was validated by measuring BOLD signals and relative CBF simultaneously in the primary visual cortex (V1) of human subjects (N = 12) during graded hypercapnia at different levels of visual stimulation. BOLD and CBF responses to specific conditions were averaged across subjects and plotted as points in the BOLD-CBF plane, tracing out lines of constant CMR(O2). The quantitative deoxyhemoglobin dilution model could be fit to these measured iso-CMR(O2) contours without significant (P TR) have found important applications in fMRI and MR thermometry since their introduction. The technique increases T *(2) weighting in BOLD imaging and temperature change sensitivity in phase-based temperature imaging compared to FLASH sequences. Yet, inconsistent observations have previously been reported when variants of this technique were used in various MRI experiments. Previous understanding of the ES sequences (the "FLASH-like" postulation) was not sufficient to explain these observations. This work provides an in-depth study on the various ES sequences. It was found that there are two types of ES sequences: ES-FLASH that spoils coherent transverse magnetization and ES-GRE, which is based on SSFP signals. A signal expression was derived for the clinically popular TR-periodic ES-GRE sequence with one echo shift. This signal expression reduced to the "FLASH-like" postulation under certain conditions. The new knowledge about the echo formation mechanism in ES sequences helps explain the experimental observations previously reported. Moreover, the resonance offset angle based analysis demonstrates an elegant methodology to analyze short TR imaging sequences. Magn Reson Med 42:864 875, 1999. PMID- 10542345 TI - Fluctuating equilibrium MRI. AB - A new fast, spectrally selective imaging method called fluctuating equilibrium magnetic resonance is presented. With all gradients refocused over a repetition interval, certain phase schedules of radiofrequency excitation pulses produce an equilibrium magnetization that fluctuates from excitation to excitation, thus permitting simultaneous acquisition of several images with different contrast features. For example, lipid and water images can be rapidly acquired. The effective echo time can be adjusted using the flip angle, thus providing control over the T(2) contribution to the contrast. Several applications of the technique are presented, including fast musculoskeletal, abdominal, breast, and brain imaging, in addition to MR angiography. A technique for combining lipid and water images generated with this sequence for angiography is described and other potential applications are suggested. Magn Reson Med 42:876-883, 1999. PMID- 10542346 TI - 1H double-quantum-filtered MR imaging as a new tool for assessment of healing of the ruptured Achilles tendon. AB - 1H double-quantum-filtered magnetic resonance imaging (DQF MRI) was applied to monitor the healing process of the Achilles tendons in rabbits after tenotomy. DQF MRI provides a new contrast, which is based on the non-zero average of the dipolar interaction caused by anisotropic motion of water molecules, determined mainly by their interaction with the ordered collagen fibers. Tissues are characterized by the dependence of their DQF signal on the DQ creation time, tau. With the use of DQF MRI, higher tissue contrast is obtained between tendon, bone, skin, and muscle. The tendons, which give weak signals in standard MRI techniques, are highlighted in the (1)H DQF image. The image changed dramatically during the healing process of the injured Achilles tendon. These changes matched the phases of the healing process. By using a tau-weighted contrast, the DQF images indicate the part of tendon that has not completely healed, even after the conventional MRI appeared normal. Magn Reson Med 42:884-889, 1999. PMID- 10542347 TI - Effect of transit times on quantification of cerebral blood flow by the FAIR T(1) difference approach. AB - The effect of finite transit times for the tagging bolus is known to be a significant error source for perfusion quantification using the flow-sensitive alternating inversion recovery (FAIR) technique. It is shown that, in the presence of transit times, both the slice-selective (SS) and nonselective (NS) inversion recovery experiments actually consist of an NS period followed by an SS period. This mixed process can be described using a newly defined time constant called the "switching time," which separates the two periods. Calculations predict that finite transit times always lead to decreased flow values in the signal-intensity-difference approach, but that the measured flows in the T(1) difference approach may be decreased or increased. This theory well explains our recent experimental flow results on cat brain as a function of predelay. The results show the signal-intensity-difference method is superior over the T(1) difference approach in terms of convenience and ease of quantification. Magn Reson Med 42:890-894, 1999. PMID- 10542348 TI - Effects of blood sugar level on rat transient focal brain ischemia consecutively observed by diffusion-weighted EPI and (1)H echo planar spectroscopic imaging. AB - The effects of blood sugar level on transient focal brain ischemia were examined by consecutive diffusion-weighted EPI and (1)H echo planar spectroscopic imaging. A remote-controlled rat intraluminal suture middle cerebral artery occlusion (MCAO) model was prepared. Animals were divided into three experimental groups: control, 1 g/kg, and 2 g/kg glucose groups (n = 6 for each). Saline or glucose was infused intraperitoneally 30 min prior to MCAO. The glucose-loaded groups showed increased lactate accumulation and marked decreases in average diffusion coefficient in the ischemic region during 40-min MCAO. These changes were correlated with blood sugar levels at the onset of MCAO. After reperfusion, all rats in the control and 1 g/kg groups recovered from the ischemic changes, but three rats with marked hyperglycemia in the 2 g/kg group showed irreversible changes. The adverse effects of hyperglycemia on transient focal brain ischemia were clearly demonstrated by sequential 2D images. Magn Reson Med 42:895-902, 1999. PMID- 10542349 TI - Methods and reproducibility of cardiac/respiratory double-triggered (1)H-MR spectroscopy of the human heart. AB - Localized (1)H-MR spectroscopy is sensitive to motion and has mostly been applied to the brain. For the human heart, cardiac and respiratory motion lead to displacements on the order of the localized voxel and lead to substantial variations of voxel content, lineshape, water suppression, and signal phase and amplitude. Combined respiratory and cardiac double triggering can avoid these complications to a large extent. Three methods of double triggering are evaluated, with reproducibility established in nine subjects for a method based on respiratory modulation of the ECG amplitude and a visual feedback mechanism. Quantitated with respect to water, within-subject reproducibilities for this setup were 9% for trimethylammonium compounds, 10% for creatine/phosphocreatine, and 13% for lipids. ANOVA showed significant differences between subjects which may relate to natural variability between subjects or exact location within the heart. Unresolved issues for this technique are its susceptibility to precise placement of ECG electrodes and the reasons for failure in 20% of examination. With this technique it is possible to investigate open questions in cardiac pathophysiology, such as the creatine content in chronic heart disease. Variants of this triggering method may also improve cardiovascular MRI methods relying on data acquired in several heartbeats. Magn Reson Med 42:903-910, 1999. PMID- 10542351 TI - Diffusion-weighted spin-echo fMRI at 9.4 T: microvascular/tissue contribution to BOLD signal changes. AB - The nature of vascular contribution to blood oxygenation level dependent (BOLD) contrast used in functional MRI (fMRI) is poorly understood. To investigate vascular contributions at an ultrahigh magnetic field of 9.4 T, diffusion weighted fMRI techniques were used in a rat forepaw stimulation model. Tissue and blood T(2) values were measured to optimize the echo time for fMRI. The T(2) of arterial blood was 40.8 +/- 3.4 msec (mean +/- SD; n = 5), similar to the tissue T(2) of 38.6 +/- 2.1 msec (n = 16). In comparison, the T(2) of venous blood at an oxygenation level of 79.6 +/- 6.1% was 9. 2 +/- 2.3 msec (n = 11). The optimal spin-echo time of 40 msec was confirmed from echo-time dependency fMRI studies. The intravascular contribution was examined using a graded diffusion-weighted spin-echo echo-planar imaging technique with diffusion weighting factor (b) values of up to 1200 sec/mm(2). Relative BOLD signal changes induced by forepaw stimulation showed no dependence on the strength or direction of the diffusion sensitizing gradients, suggesting that the large vessel contribution to the BOLD signal is negligible at 9.4 T. However, gradient-echo fMRI performed with bipolar diffusion sensitizing gradients, which suppress intravascular components from large vessels, showed higher percent signal changes in the surface of the brain. This effect was attributed to the extravascular contribution from large vessels. These findings demonstrate that caution should be exercised when interpreting that higher percent changes obtained with gradient-echo BOLD fMRI are related to stronger neural activation. Magn Reson Med 42:919-928, 1999. PMID- 10542352 TI - Texture analysis of spinal cord pathology in multiple sclerosis. AB - Texture analysis was applied to MR images of the spinal cord in an attempt to quantify pathological changes that occur in multiple sclerosis (MS). Texture features quantify macroscopic lesions and also the microscopic abnormalities that may be undetectable using conventional measures of lesion volume and number. Significant differences in texture between normal controls and MS patients were seen. Texture differences were detected between normal controls and relapsing remitting patients before detectable spinal cord atrophy. There was also significant correlation between texture and disability. The segmentation and texture analysis technique demonstrates intraobserver coefficients of variation ranging from 0. 6-8.2%. Texture analysis has potential as a tool for monitoring changes associated with the development of disability in patients with MS. Reproducibility and sensitivity must be improved to use the technique for serial monitoring in individuals. Magn Reson Med 42:929-935, 1999. PMID- 10542350 TI - Water diffusion, T(2), and compartmentation in frog sciatic nerve. AB - A potential relationship between structural compartments in neural tissue and NMR parameters may increase the specificity of MRI in diagnosing diseases. Nevertheless, our understanding of MR of nerves and white matter is limited, particularly the influence of various water compartments on the MR signal is not known. In this study, components of the (1)H transverse relaxation decay curve in frog peripheral nerve were correlated with the diffusion characteristics of the water in the nerve. Three T(2) values were identified with nerve. Water mobility was found to be unrestricted on the timescale of 100 msec in the component of the signal with the intermediate T(2) time, suggesting some contribution from the interstitial space to this T(2) component. Restricted diffusion was observed in the component with the longest T(2) time, supporting the assignment of at least part of the spins contributing to this component to an intracellular compartment. The observed nonexponential behavior of the diffusion attenuation curves was investigated and shown to be potentially caused by the wide range of axon sizes in the nerve. Magn Reson Med 42:911-918, 1999. PMID- 10542353 TI - Biochemical modulation of the catabolism and tissue uptake of the anticancer drug 5-fluorouracil by 5-bromovinyluracil: assessment with metabolic (19)F MR imaging. AB - Using chemical shift-selective (19)F magnetic resonance (MR) imaging, we investigated the biomodulating action of 5-bromovinyluracil (BVU) on the degradation of the anticancer drug 5-fluorouracil (5-FU) to its major catabolite alpha-fluoro-beta-alanine (FBAL) and the tissue uptake of 5-FU in ACI rats with transplanted Morris hepatoma. Rats in the control group (n = 7) received 200 mg/kg body weight of 5-FU intravenously, whereas the rats in the BVU group (n = 7) additionally received 30 mg/kg body weight of BVU intraperitoneally about 45 min before 5-FU injection. In each animal examination, three selective (19)F MR images were acquired sequentially after 5-FU administration with an acquisition time of 32 min each: an early 5-FU image (dominant Fourier line, 8 min p.i.) that characterized the early uptake of the drug into the various tissues, an FBAL image (dominant Fourier line, 56 min p.i.) that reflected the catabolism of the drug, and a late 5-FU image (dominant Fourier line, 78 min p.i.) that assessed the retention ("trapping") of unmetabolized 5-FU and its MR-visible anabolites. Pretreatment with BVU resulted in a highly statistical significant decrease (P < 0.001) of the FBAL signal in the liver. The marked effect of BVU on 5-FU degradation, however, improved neither the early uptake nor the retention of 5-FU in skeletal muscle and tumor tissue (P > 0.7). Moreover, our results indicate that 5-FU tumor uptake is not only dependent on the plasma concentration of unmetabolized 5-FU but is also determined by tumor-specific factors, these showing considerable variations between individual neoplasms. Magn Reson Med 42:936-943, 1999. PMID- 10542354 TI - MRI measurement of the temporal evolution of relative CMRO(2) during rat forepaw stimulation. AB - This study reports the first measurement of the relative cerebral metabolic rate of oxygen utilization (rCMRO(2)) during functional brain activation with sufficient temporal resolution to address the dynamics of blood oxygen level dependent (BOLD) MRI signal. During rat forepaw stimulation, rCMRO(2) was determined in somatosensory cortex at 3-sec intervals, using a model of BOLD signal and measurements of the change in BOLD transverse relaxation rate, the resting state BOLD transverse relaxation rate, relative cerebral blood flow (rCBF), and relative cerebral blood volume (rCBV). Average percentage changes from 10 to 30 sec after onset of forepaw stimulation for rCBF, rCBV, rCMRO(2), and BOLD relaxation rate were 62 +/- 16, 17 +/- 2, 19 +/- 17, and -26 +/- 12, respectively. A poststimulus undershoot in BOLD signal was quantitatively attributed to the temporal mismatch between changes in blood flow and volume, and not to the role of oxygen metabolism. Magn Reson Med 42:944-951, 1999. PMID- 10542355 TI - SENSE: sensitivity encoding for fast MRI. AB - New theoretical and practical concepts are presented for considerably enhancing the performance of magnetic resonance imaging (MRI) by means of arrays of multiple receiver coils. Sensitivity encoding (SENSE) is based on the fact that receiver sensitivity generally has an encoding effect complementary to Fourier preparation by linear field gradients. Thus, by using multiple receiver coils in parallel scan time in Fourier imaging can be considerably reduced. The problem of image reconstruction from sensitivity encoded data is formulated in a general fashion and solved for arbitrary coil configurations and k-space sampling patterns. Special attention is given to the currently most practical case, namely, sampling a common Cartesian grid with reduced density. For this case the feasibility of the proposed methods was verified both in vitro and in vivo. Scan time was reduced to one-half using a two-coil array in brain imaging. With an array of five coils double-oblique heart images were obtained in one-third of conventional scan time. Magn Reson Med 42:952-962, 1999. PMID- 10542356 TI - Motion correction with PROPELLER MRI: application to head motion and free breathing cardiac imaging. AB - A method for motion correction, involving both data collection and reconstruction, is presented. The PROPELLER MRI method collects data in concentric rectangular strips rotated about the k-space origin. The central region of k-space is sampled for every strip, which (a) allows one to correct spatial inconsistencies in position, rotation, and phase between strips, (b) allows one to reject data based on a correlation measure indicating through-plane motion, and (c) further decreases motion artifacts through an averaging effect for low spatial frequencies. Results are shown in which PROPELLER MRI is used to correct for bulk motion in head images and respiratory motion in nongated cardiac images. Magn Reson Med 42:963-969, 1999. PMID- 10542357 TI - Heart motion adapted cine phase-contrast flow measurements through the aortic valve. AB - A method for magnetic resonance cine velocity mapping through heart valves with adaptation of both slice offset and angulation according to the motion of the valvular plane of the heart is presented. By means of a subtractive labeling technique, basal myocardial markers are obtained and automatically extracted for quantification of heart motion at the valvular level. The captured excursion of the basal plane is used to calculate the slice offset and angulation of each required time frame for cine velocity mapping. Through-plane velocity offsets are corrected by subtracting velocities introduced by basal plane motion from the measured velocities. For evaluation of the method, flow measurements downstream from the aortic valve were performed both with and without slice adaptation in 11 healthy volunteers and in four patients with aortic regurgitation. Maximum through-plane motion at the aortic root level as calculated from the labeled markers averaged 8.9 mm in the volunteers and 6.5 mm in the patients. The left coronary root was visible in 2-4 (mean: 2.2) time frames during early diastole when imaging with a spatially fixed slice. Time frames obtained with slice adaptation did not contain the coronary roots. Motion correction increased the apparent regurgitant volume by 5.7 +/- 0.4 ml for patients with clinical aortic regurgitation, for an increase of approximately 50%. The proposed method provides flow measurements with correction for through-plane motion perpendicular to the aortic root between the valvular annulus and the coronary ostia throughout the cardiac cycle. Magn Reson Med 42:970-978, 1999. PMID- 10542359 TI - Migration and chemical modification of silicone in women with breast prostheses. PMID- 10542360 TI - Migration and chemical modification of silicone in women with breast prostheses. PMID- 10542361 TI - Migration and chemical modification of silicone in women with breast prostheses. PMID- 10542363 TI - Response PMID- 10542362 TI - A critical review of MR studies concerning silicone breast implants. PMID- 10542364 TI - Characterization of microtubule-associated proteins in teleosts. AB - Although microtubules are known to play an important role in many cellular processes, they have been virtually neglected in fish. In this report, microtubule-associated proteins (MAPs) in fish (teleost) were characterized using antibodies (Abs) directed against the mammalian MAPs tau, MAP1A and B, and MAP 2. Two different populations of tau-like proteins (TLPs) were found in fish brain using the anti-tau Abs Tau-1, Tau-2, tau5', and tau3'. The TLPs that were recognized by Tau-1, Tau-2, and tau5' were (1) heat-stable; (2) the same molecular weight as mammalian TLPs: 59-62 kDa; (3) not enriched in microtubules prepared from catfish brain; and (4) localized to the cell body of neurons in fish brains. While the TLPs recognized by tau3' Abs were (1) heat-stable; (2) lower molecular weight than mammalian TLPs: 32-55 vs. 50-65 kDa; (3) enriched in microtubule fractions prepared from catfish brain, and (4) localized to the axons of neurons. These results are consistent with two different populations of TLPs being present in fish brains. While MAP2 was found to be approximately the same molecular weight, 250 kDa, in zebrafish and goldfish as in mammals and to be distributed to dendrites in the fish brain, both MAP1A and MAP1B were found to be about 25% the mass of their mammalian homologs. These results suggest that MAPS in fish have some characteristics similar to their mammalian counterparts, but also possess some unique properties that require further study to elucidate their function. PMID- 10542365 TI - Attenuation of plasminogen activator inhibitor type-1 promoter activity in serum stimulated renal epithelial cells by a distal 5' flanking region. AB - Urokinase plasminogen activator (u-PA) and its fast acting type-1 inhibitor (PAI 1) localize to cellular focal adhesive structures and the adjoining proximal undersurface region, respectively (Kutz et al., J. Cell. Physiol. 176:8-18, 1997). PAI-1 may function in this locale to modulate pericellular proteolytic activity, cell-to-substrate adhesion, or matrix-dependent motility. While PAI-1 synthesis is regulated in an immediate-early response manner in growth "activated" renal cells coincident with cytoskeletal restructuring, adhesive influences both repress the amplitude and prolong the time course of serum induced PAI-1 transcription (Ryan et al., Biochem. J. 314:1041-1046, 1996). To identify potential adhesion-responsive elements within the PAI-1 gene that function in this complex mode of expression control, reporter constructs containing defined directionally deleted PAI-1 5' genomic fragments cloned upstream of a CAT gene were employed in transient transfection assays. A 483-bp distal PAI-1 flanking segment (corresponding to nucleotides -2395 to -1912) conferred significant adhesion-dependent attenuation on serum-induced PAI-1 transcription. This 483-bp distal PAI-1 segment functioned as a repressor of reporter (CAT) activity under both adhesive and suspension culture conditions, however, when ligated upstream of either an SV40 promoter/enhancer or a minimal PAI-1 promoter. These data suggest that repressor elements located between -2395 and -1912 bp interact with more proximal adhesion-dependent regulatory elements to affect PAI-1 expression attenuation during serum stimulation of adherent renal epithelial cells. PMID- 10542367 TI - Monastral bipolar spindles in meiosis II of male Trichosia pubescens (Sciaridae): early stages of spindle formation and chromosome orientation. AB - The metaphase spindle of male meiosis II in fungus gnats (Sciaridae) is one example of naturally occurring monastral bipolar spindles. To gain further insights into how the bipolar spindle is formed in the presence of only one polar center, prometaphase of male meiosis II was investigated in the sciarid Trichosia pubescens by means of anti-tubulin immunofluorescence, DAPI chromosome staining, and electron microscopy of ultrathin serial sections. The first step in spindle formation after interkinesis seems to be the organization of an astral half spindle, probably by MTOC activity of the astral region. With the exception of the non-disjunctional X chromosome, which always lies close to the aster, the chromosomes are found to occupy various positions with respect to the astral region, revealing different orientations of their chromatid kinetochores. It was observed that some of the mal-oriented kinetochores are associated with microtubules that, due to their orientation perpendicular to the spindle axis, are unlikely to originate from the astral region. Therefore, these mal-oriented microtubules are taken as an indication of a dispersed MTOC activity near the chromosomes or at kinetochores. According to recent models of chromosome-induced spindle self-organization [e.g., Merdes et al., 1997: J. Cell Biol. 138:953-956], they could be responsible for the formation of the other (anastral) half-spindle and for amphitelic (bipolar) orientation of the chromosomes. PMID- 10542366 TI - Flagellar arrest behavior predicted by the Geometric Clutch model is confirmed experimentally by micromanipulation experiments on reactivated bull sperm. AB - The central tenet of the Geometric Clutch hypothesis of flagellar beating is that the internal force transverse to the outer doublets (t-force) mediates the initiation and termination of episodes of dynein engagement. Therefore, if the development of an adequate t-force is prevented, then the dynein-switching necessary to complete a cycle of beating should fail. The dominant component of the t-force is the product of the longitudinal force on each outer doublet multiplied by the local curvature of the flagellum. In the present study, two separate strategies, blocking and clipping, were employed to limit the development of the t-force in Triton X-100 extracted bull sperm models. The blocking strategy used a bent glass microprobe to restrict the flagellum during a beat, preventing the development of curvature in the basal portion of the flagellum. The clipping strategy was designed to shorten the flagellum by clipping off distal segments of the flagellum with a glass microprobe. This limits the number of dyneins that can contribute to bending and consequently reduces the longitudinal force on the doublets. The blocking and clipping strategies both produced an arrest of the beat cycle consistent with predictions based on the Geometric Clutch hypothesis. Direct comparison of experimentally produced arrest behavior to the behavior of the Geometric Clutch computer model of a bull sperm yielded similar arrest patterns. The computer model duplicated the observed behavior using reasonable values for dynein force and flagellar stiffness. The experimental data derived from both blocking and clipping experiments are fully compatible with the Geometric Clutch hypothesis. PMID- 10542368 TI - Characterization of outer arm dynein in sea anemone, Anthopleura midori. AB - Outer arm dynein was purified from sperm flagella of a sea anemone, Anthopleura midori, and its biochemical and biophysical properties were characterized. The dynein, obtained at a 20S ATPase peak by sucrose density gradient centrifugation, consisted of two heavy chains, three intermediate chains, and seven light chains. The specific ATPase activity of dynein was 1.3 micromol Pi/mg/min. Four polypeptides (296, 296, 225, and 206 kDa) were formed by UV cleavage at 365 nm of dynein in the presence of vanadate and ATP. In addition, negatively stained images of dynein molecules and the hook-shaped image of the outer arm of the flagella indicated that sea anemone outer arm dynein is two-headed. In contrast to protist dyneins, which are three-headed, outer arm dyneins of flagella and cilia in multicellular animals are two-headed molecules corresponding to the two heavy chains. Phylogenetic considerations were made concerning the diversity of outer arm dyneins. PMID- 10542369 TI - Phosphorylation of MAP2c and MAP4 by MARK kinases leads to the destabilization of microtubules in cells. AB - Microtubules serve as transport tracks in molecular mechanisms governing cellular shape and polarity. Rapid transitions between stable and dynamic microtubules are regulated by several factors, including microtubule-associated proteins (MAPs). We have shown that MAP/microtubule affinity regulating kinases (MARK) can phosphorylate the microtubule-associated-proteins MAP4, MAP2c, and tau on their microtubule-binding domain in vitro. This leads to their detachment from microtubules (MT) and an increased dynamic instability of MT. Here we show that MARK protein kinases phosphorylate MAP2 and MAP4 on their microtubule-binding domain in transfected CHO cells. In CHO cells expressing MARK1 or MARK2 under control of an inducible promoter, MARK2 phosphorylates an endogenous MAP4-related protein. Prolonged expression of MARK2 results in microtubule-disruption, detachment of cells from the substratum, and cell death. Concomitant with microtubule disruption, we also observed a breakdown of the vimentin network, whereas actin fibers remained unaffected. Thus, MARK seems to play an important role in controlling cytoskeletal dynamics. PMID- 10542371 TI - Structure and expression of the rat relaxin-like factor (RLF) gene. AB - The relaxin-like factor (RLF) is a novel member of the insulin-IGF-relaxin family of growth factors and hormones, and its mRNA is expressed very specifically in the Leydig cells of the testis and in the theca and luteal cells of the ovary. Here we report the cloning of the RLF gene and cDNA from the rat. The 0.8kb mRNA is produced from a small gene comprising two exons situated less than 1 kb downstream of the gene for the signalling factor JAK3. Northern hybridization confirms high RLF mRNA expression in the adult rat testis, and low expression in the ovary, but in no other tissues examined. Northern analysis of fetal and neonatal gonadal tissues showed that RLF mRNA is highly upregulated in the testes of day 19 embryos, but not in later neonatal stages, nor in any ovarian tissue from this period. This would indicate that RLF is a marker for the mature fetal as well as the adult-type Leydig cell, but is not expressed in premature, precursor, or dedifferentiated Leydig cells of either cell type. Finally, RNA was analysed from the testes of rats which had been treated with ethylene dimethane sulfonate (EDS), an alkylating agent that specifically destroys rat Leydig cells. RLF mRNA was absent from the acutely treated testes, but became detectable between 15 and 20 days post-treatment, concomitant with the repopulation of the testes by new Leydig cells. Continuous testosterone substitution of EDS-treated rats suppressed the production of gonadotropins, and LH-dependent Leydig cell differentiation, with the result that RLF mRNA remained undetectable throughout the study period. In conclusion, RLF is a very specific marker for the mature Leydig cell phenotype in both the adult-type and fetal Leydig cell populations of the rat testis. PMID- 10542372 TI - Upregulation of CHOP-10 (gadd153) expression in the mouse blastocyst as a response to stress. AB - CHOP-10 (also known as gadd153 or Ddit3) is one of the genes overexpressed by mammalian cells exposed to cytotoxic agents or to nutrient stress. The response of this gene to stress was studied in the mouse blastocyst and in F9 embryonal carcinoma cells. When mouse blastocysts were exposed to the alkylating agent MMS, the metabolic inhibitor sodium arsenite or an inhibitor of protein glycosylation tunicamycin, levels of the CHOP-10 mRNA were increased by two- to threefold relative to the mRNA for beta-actin. There was no increase in gene expression when blastocysts were treated with the inhibitor of nucleotide synthesis PALA. These results show that the response of CHOP-10 is dependent on the type of stress applied to the embryo. When F9 embryonal carcinoma cells were treated with MMS or sodium arsenite, CHOP-10 expression was induced by fourfold within 4 hr of treatment. The induction following tunicamycin treatment was slower requiring at least 24 hr. The response to tunicamycin was greater in cells treated with retinoic acid to induce differentiation. The results suggest that there is a link between the extent of glycoprotein synthesis and the sensitivity of CHOP-10 to tunicamycin. The inhibitor PALA did not change CHOP-10 expression in the presence or absence of retinoic acid. In F9 cells an increase in the expression of CHOP-10 was followed by cell death due to apoptosis. The overexpression of CHOP-10 may be a marker for one of the pathways that lead to apoptosis in the blastocyst. These results suggest that there is more than one control system regulating growth arrest in the blastocyst and the fetal outcome may differ depending on the type of stress encountered in culture. PMID- 10542373 TI - Expression of protein gene product 9.5, a neuronal ubiquitin C-terminal hydrolase, and its developing change in sertoli cells of mouse testis. AB - Protein gene product 9.5 (PGP9.5), originally isolated as a neuron-specific protein, belongs to a family of ubiquitin carboxyl-terminal hydrolases that play important roles in the nonlysosomal proteolytic pathway. Antibodies against PGP9.5 have been used for immunohistochemical detection of neural elements, although some non-neuronal cells are also immunoreactive for PGP9.5. In the present study, developing testes of the mouse were immunostained after autoclave pretreatment of sections. In the testes of days 8 and 16, PGP9.5 was only localized on the spermatogonia, whereas on day 30 and in adults it appeared not only on spermatogonia, but also on Sertoli cells. In the testis of the male sterile W/W(v) mutant, very little, but strong, immunoreactivity was detected at some Sertoli cells, which were phagocytizing Sertoli cell aggregations that had fallen from the basal membrane. Additionally, it was confirmed that the nucleotide sequence of PGP9.5 in mice was highly conserved, like that in other mammals. These results suggest that PGP9.5 is a useful marker for activated Sertoli cells, playing an important role in degradation of abnormal proteins. PMID- 10542374 TI - Relationship between dead cells and DNA fragmentation in bovine embryos produced in vitro and stored at 4 degrees C. AB - DNA fragmentation and its relationship with dead cells were examined in bovine blastocysts produced in vitro and stored at 4 degrees C for 1-5 days. Survival and development to the hatching and hatched blastocyst stage decreased with increasing storage time. Both were significantly lower at 72 hr than at 48 hr. None of the embryos stored for 120 hr developed to the hatching or hatched blastocyst stage. The proportion of dead cells per embryo increased progressively as the time of storage increased, until 69% of embryonic cells were dead after 120 hr of storage. There was no significant difference between the proportions of DNA fragmentation per embryo stored for 0 and 24 hr (12% vs 16%). However, the proportion of DNA fragmentation in embryos stored for longer than 48 hr was significantly greater than that in embryos stored for less than 24 hr. There were no significant differences among those stored for longer than 48 hr (28-33%). These results suggest that the reduced developmental competence of bovine embryos stored at 4 degrees C is characterized by necrotic change rather than apoptotic change. PMID- 10542376 TI - Selenium-independent epididymis-restricted glutathione peroxidase 5 protein (GPX5) can back up failing Se-dependent GPXs in mice subjected to selenium deficiency. AB - We have previously characterized and cloned a secreted sperm-bound selenium independent glutathione peroxidase protein (GPX5), the expression of which was found to be restricted to the mouse caput epididymidis. Because of the lack of selenium (Se) in the active site of this enzyme, unlike the other animal GPXs characterized to date, it was suspected that GPX5 does not function in the epididymis as a true glutathione peroxidase in vivo. In the present report, following dietary selenium deprivation which is known to reduce antioxidant defenses and favor oxidative stress in relation with depressed Se-dependent GPX activities, we show that the epididymis is still efficiently protected against increasing peroxidative conditions. In this model, the caput epididymides of selenium-deficient animals showed a limited production of lipid peroxides, a total GPX activity which was not dramatically affected by the shortage in selenium availability and an increase in GPX5 mRNA and protein levels. Altogether, these data strongly suggest that the selenium-independent GPX5 could function as a back-up system for Se-dependent GPXs. PMID- 10542375 TI - Quantitation of IGF-I, IGF-II, and multiple insulin receptor family member messenger RNAs during embryonic development in rainbow trout. AB - The IGF system has been shown to be important for normal embryonic growth in mice. Characterization of the IGF system in lower vertebrates is still in progress. To gain a greater understanding of the IGF system during embryonic development in teleosts, a competitive reverse transcription-polymerase chain reaction (RT-PCR) assay was developed and used to quantitate the levels of IGF-I and IGF-II mRNAs from rainbow trout embryos isolated from a staged series. The absolute number of molecules of IGF-I mRNA/microg total RNA was significantly lower than the absolute number of molecules of IGF-II mRNA/microg total RNA both during early and late embryonic development. The recent identification of multiple IGF type I receptor (rtIGFR) and insulin receptor (rtIR) cDNAs in rainbow trout has provided us with a tool to investigate the expression of these mRNAs. A relative quantitative RT-PCR assay was used to determine the steady state levels of two forms of rtIGFR and three forms of rtIR mRNAs in rainbow trout embryos. The relative levels of rtIGFR mRNAs were greater in embryos compared to adult tissues while the relative levels of rtIR mRNAs were generally lower. In a RT-PCR based assay, a differential ability to detect rtIGFR and rtIR mRNAs was shown, suggesting developmental regulation of polyadenylation. Our results suggest that IGF-II mRNA is the predominant IGF expressed in rainbow trout embryos. Our characterization of IGF ligand and receptor mRNA levels in rainbow trout embryos suggests that a functional IGF system exists during embryonic development in teleosts. PMID- 10542377 TI - Intracellular regulation of estradiol and progesterone production by cultured bovine granulosa cells. AB - The objective of the present study was to investigate the implication of protein kinase A (PKA), protein kinase C (PKC), and receptor protein tyrosine kinase (R PTK) pathways in the regulation of estradiol (E2) and progesterone (P4) production by bovine granulosa cells. Cells were harvested from bovine follicles (8-15 mm diameter) and cultured without serum for an initial 3 days (37 degrees C; 5% CO(2) in air; D1-D3). On the fourth day of culture (D4), E2 and P4 production were stimulated with FSH (1-6 ng/ml) or forskolin (FSK) in the presence or absence of intracellular effectors of PKA, PKC, and R-PTK. Culture medium was collected and replaced each day. Stimulation of granulosa cell adenylate cyclase activity with FSK (0.06-3.75 microM) mimicked FSH, inducing a quadratic increase (P < 0.001) of E2 production and a continuous elevation of P4 (P < 0.01). Inhibition of R-PTK activity with genistein (25-50 microM) increased the sensitivity of cells to FSH as demonstrated by a leftward shift in the dose response curve (P < 0.001). Treatment with transforming growth factor-alpha (TGFalpha; 0. 1 ng/ml) abolished the FSH-induced E2 production (P < 0.001) and this effect was not reversed (P < 0.001) by FSK or by genistein. Furthermore, the inhibitory effect of TGFalpha on FSH-induced E2 production was reproduced by phorbol 12-myristate 13-acetate (PMA; 1. 25-2.5 microM), a PKC activator (P < 0.001). Interestingly, genistein inhibited P4 production (P < 0.05). From these results, we conclude that E2 production by bovine granulosa cells is mediated by intracellular factors and can be stimulated downstream from the FSH receptor. The results also suggest that stimulation of R-PTK and/or PKC activities, as probably occurs with TGFalpha, negatively affects the PKA pathway, thus decreasing E2 production. Furthermore, inhibition of R-PTK leads to an increase production of E2 and may limit luteinization of bovine granulosa cells. PMID- 10542378 TI - Porcine sperm surface beta1,4galactosyltransferase binds to the zona pellucida but is not necessary or sufficient to mediate sperm-zona pellucida binding. AB - The binding of sperm to the zona pellucida is an integral part of the mammalian fertilization process, investigated most extensively in the mouse. Several sperm receptors for the murine zona pellucida have been studied (Snell WJ, White JM. 1996. Cell 85:629-637; Wassarman PM. 1999. Cell 96:175-183), but the most compelling evidence exists for beta-1,4-galactosyltransferase (GalTase). Considering that GalTase is present on the surface of porcine sperm (Larson JL, Miller DJ. 1997. Biol Reprod 57:442-453), we investigated the role of GalTase in porcine sperm-zona binding. Sperm surface GalTase catalyzed the addition of uridine diphosphate-[(3)H]galactose to the 55 kDa group of the porcine zona pellucida proteins implicated in sperm binding, demonstrating that GalTase binds the porcine zona. The functional importance of GalTase-zona pellucida binding was tested. Addition of uridine diphosphate galactose, a substrate that completes the GalTase enzymatic reaction and disrupts GalTase mediated adhesion, had no effect on binding of sperm to porcine oocytes. Furthermore, removal of the GalTase zona ligand by incubation of oocytes with N-acetylglucosaminidase had no effect on binding of sperm to oocytes. These results suggest that GalTase is not necessary for sperm to bind to the zona pellucida. Digestion of isolated porcine zona proteins with N-acetylglucosaminidase did not affect the biological activity of soluble porcine zona proteins in competitive sperm-zona binding assays, suggesting that GalTase alone is not sufficient to mediate sperm-zona attachment. From these results, it appears that, although GalTase is able to bind porcine zona proteins, its function in porcine sperm-zona binding is not necessary or sufficient for sperm-zona binding. This supports the contention that porcine sperm-zona binding requires redundant gamete receptors. PMID- 10542379 TI - Effects of cooling germinal vesicle-stage bovine oocytes on meiotic spindle formation following in vitro maturation. AB - Attempts to cryopreserve bovine oocytes result in low survival because of their sensitivity to temperatures near 0 degrees C. This study evaluates the effects of chilling germinal vesicle-stage (GV) oocytes on their formation of microtubules and the meiotic spindle. In experiment 1, five groups of GV-stage oocytes, each consisting of approximately 90 oocytes, were held at 39 degrees C as controls, or at 31 degrees C, or cooled to 24, 4 or 0 degrees C for 10 min. After being treated, all oocytes were cultured at 39 degrees C for 24 hr. Compared to the controls, holding oocytes for 10 min at 31 or 24 degrees C did not significantly alter the formation of normal spindles, but chilling them to 4 or 0 degrees C did. After 24 hr of maturation, the respective percentages of oocytes containing normal meiotic spindles observed in the controls or those held at 31 or 24 degrees C were 69.8%, 71.9%, or 69.4% (P > 0.05). In contrast, the percentages of oocytes with normal spindles after they had been cooled to 4 or 0 degrees C were 44.0% or 29.1%, respectively. In experiment 2, approximately 90 oocytes/group were cooled to 4 degrees C for various times before being warmed and cultured. Regardless of the time of exposure, cooling oocytes to 4 degrees C reduced the formation of normal spindles. The percentages of oocytes cooled to 4 degrees C for 10, 20, 30, 45, or 60 min with normal spindles were 44.0%, 38.4%, 37.5%, 34.5% and 30.9%, respectively. In experiment 3, approximately 60 oocytes per group that had been held at 31 degrees C or cooled to 24, 4 or 0 degrees C for 10 min were allowed to mature for 24 hr before being subjected to in vitro fertilization. The cleavage rates of oocytes subjected to various chilling treatments exhibited the same pattern as that of oocytes with normal spindles. That is, there were no significant differences in cleavage rates among the control oocytes and those held at 31 or 24 degrees C (70.4%, 71.8%, and 72.4%; P > 0.05). However, only 37. 0% and 30.4% of oocytes chilled to 4 or 0 degrees C cleaved after fertilization. These results suggest that: (1) chilling bovine oocytes no lower than 24 degrees C does not reduce formation of normal meiotic spindles; (2) however, chilling oocytes to 4 degrees C or lower for as little as 10 min drastically reduces the formation of normal meiotic spindles and of fertilization; (3) the rates of fertilization and cleavage of resultant zygotes mimic that of formation of normal spindles. PMID- 10542380 TI - Regulation of intracellular pH in bovine oocytes and cleavage stage embryos. AB - This study investigated the mechanisms for the regulation of intracellular pH in bovine oocytes and embryos. Na(+)/H(+) antiporter activity for the regulation of intracellular pH in the acid to neutral range was detected in both in vitro matured bovine oocytes and in vitro produced embryos. However, the activity of the antiporter was significantly reduced in oocytes compared to 2-cell, 4-cell, and 8-cell embryos. HCO(3)(-)/Cl(-) exchanger activity could be detected in oocytes and embryos using the chloride removal method, however the ability of this transporter to regulate intracellular pH against an alkaline load was poor and intracellular pH could not be re-established. The inability of the HCO(3)( )/Cl(-) exchanger to adequately regulate intracellular pH was further highlighted by the arrest of embryos at the 8-16 cell stage when challenged with a small alkaline load. Therefore, bovine embryos are extremely sensitive to alterations in intracellular pH above the resting value of around 7.2. This sensitivity could account in part for impaired development and viability of bovine embryos produced in vitro. PMID- 10542381 TI - Cysteine-containing histone H1-like (PL-I) proteins of sperm. AB - We have determined the presence of cysteine in the protein PL-I from the sperm of the surf clam Spisula solidissima. The existence of cysteine in this histone H1 related protein is responsible for its previously described aggregation behavior. The location of this residue, within the trypsin-resistant domain of the protein, has been established. We have also shown that cysteine is ubiquitously present in the PL-I proteins from the sperm of other bivalve mollusks but is absent from other PL of smaller molecular mass (PL-II, PL-III, PL-IV). We have also found cysteine to be present in the PL-I from a tunicate (Chelysoma productum) but absent in a PL-I from a fish (Mullus barbatus). The possible significance of the unusual occurrence of cysteine in these histone-H1-related proteins is discussed. PMID- 10542382 TI - Phosphate induced developmental arrest of hamster two-cell embryos is associated with disrupted ionic homeostasis. AB - Culture of hamster embryos with 0.35 mM inorganic phosphate results in developmental arrest at the 2-cell stage. These arrested 2-cell embryos were found to have significantly elevated levels of both intracellular pH and intracellular free calcium. Culture of 2-cell embryos with both glucose and phosphate did not further alter intracellular ionic homeostasis. Developmental arrest of 2-cell embryos was dependent on the concentration of phosphate used. Culture with 1.25 microM phosphate did not alter development, while concentrations of 2.5 microM and 5.0 microM resulted in a percentage of embryos arresting development at the 2-cell stage. Analysis of intracellular levels of pH and calcium after culture with different phosphate concentrations revealed a significant negative correlation between intracellular calcium levels and development beyond the 2-cell stage. There was no correlation between the increase in intracellular pH and embryo development in the presence of phosphate. The increase in intracellular calcium levels after culture with phosphate appears to be derived from intracellular pools, as preventing the influx of extracellular calcium did not alter development beyond the 2-cell stage. Therefore, it is apparent that a disruption in ionic homeostasis is associated with developmental arrest of hamster embryos cultured with phosphate. PMID- 10542384 TI - The role of virtual organizations in the 21st century. PMID- 10542386 TI - Water insoluble drugs set for oral delivery. PMID- 10542383 TI - Determination of the intracellular dissociation constant, K(D), of the fluo-3. Ca(2+) complex in mouse sperm for use in estimating intracellular Ca(2+) concentrations. AB - In order to calculate the actual, rather than the relative, intracellular Ca(2+) concentration (Ca(2+))(i) in mammalian sperm cells, using fluorescent probes whose fluorescence emission differs between the probe. Ca(2+) complex and free probe, the value of the dissociation constant for the probe. Ca(2+) complex, K(D), is required. Interaction of the probe with cellular components may change the intracellular value of K(D) from that determined in buffered solution. We had previously shown that fluo-3, whose Ca(2+) complex is highly fluorescent whereas free fluo-3 is not, could be used to monitor changes of (Ca(2+))(i) in mouse sperm. In this report, we describe a method for determining K(D) for the fluo-3. Ca(2+) complex in mouse sperm suspended in medium MJB, a medium in which the sperm remain viable, but which contains high Ca(2+). The method involved treating the sperm with ionomycin to provide a plasma membrane Ca(2+) carrier, with nigericin to eliminate pH gradient, and with gramicidin D to eliminate membrane potential, such that (Ca(2+))(i) equilibrates with medium Ca(2+) concentration (Ca(2+))(e), then titrating (Ca(2+))(e) with EGTA in added aliquots to near nil concentration. At EGTA concentrations in excess of total medium Ca(2+), an approximation algorithm was used to calculate (Ca(2+))(e), based on the known K(D) for the EGTA. Ca(2+) complex. The fluorescence of the intracellular fluo-3. Ca(2+) complex, F, decreased with increasing additions of EGTA; (Ca(2+))(i) = (Ca(2+))(e) was plotted as a linear function of F/[F(max) - F]; the slope gives K(D). At 37 degrees C, intracellular K(D) was calculated to be 0.636 +/- 0.018 microM (+/-SEM, n = 8). At 37 degrees C and 20 degrees C, K(D) values in MJB were calculated to be 0.502 +/- 0.022 and 0.578 +/- 0.029 (+/-SEM, n =8 and n = 6), respectively. The higher intracellular K(D) value implies probe interaction with cytosol components, primarily those in the head, as this compartment is the major contributor to sperm fluorescence. Changes in (Ca(2+))(i), monitored with fluo-3 fluorescence, that occur on interaction of capacitated mouse sperm with the zona pellucida and may now be quantified, using 0.636 microM for K(D) of the intracellular fluo-3. Ca(2+) complex. PMID- 10542388 TI - Novel vaccination approaches on the horizon? PMID- 10542385 TI - Corrigendum. PMID- 10542387 TI - Haemophilia B gene therapy using adeno-associated virus. PMID- 10542389 TI - Strategies for particle design using supercritical fluid technologies. AB - Major advances in drug delivery and targeting over recent years have highlighted the limitations of conventional particle formation and pretreatment processes in fine-tuning the characteristics required. The alternative strategy of using supercritical fluid technologies for crystal and particle engineering of pharmaceutical materials and drug delivery systems shows great promise in this area. The design of particles for specific drug delivery needs - such as particle size control or polymorphic purity - is increasingly seen as a viable option. In describing recent progress in this field, this review provides a perspective of the current position of this platform technology and considers the possibilities and challenges for future applications and developments. PMID- 10542390 TI - Polymer conjugates for tumour targeting and intracytoplasmic delivery. The EPR effect as a common gateway? AB - Tumour capillaries are frequently hyperpermeable compared with normal vasculature, and thus they offer a much sought-after gateway for targeted delivery of cancer chemotherapy. Phase I clinical trials reported recently describe the first synthetic polymer-drug conjugate to be tested in man. N-(2 Hydroxypropyl)methacrylamide copolymer-doxorubicin (PK1, FCE 28068) displayed antitumour activity in chemotherapy-refractory patients, considerably reduced toxicity compared with doxorubicin, and evidence of tumour-selective targeting. With increasing understanding of the vector- and tumour-related factors that govern vascular permeability, non-viral vectors are being designed for tumour selective targeting and subsequent intracytoplasmic delivery of macromolecular medicines such as genes, antisense oligonucleotides, proteins and peptides. PMID- 10542391 TI - Delivery of peptide and non-peptide drugs through the respiratory tract. AB - The respiratory tract, and the nose in particular, offers opportunities for improved drug delivery. Many drugs are rapidly and efficiently absorbed from the nasal cavity and, as a result, the nasal route may be used in crisis treatments (for example, for pain and nausea). Polar drugs, such as peptides and proteins, are not well absorbed across the nasal mucosa, unless they are delivered with an absorption enhancing material. Agents, such as the polysaccharide chitosan, that are able to open tight junctions between cells can offer important opportunities. The nasal route can also be used for the delivery of vaccines. This review makes a comparison between nasal and pulmonary delivery. PMID- 10542392 TI - Metabolism-based drug design and drug targeting. AB - Even at the early stages of drug discovery and structure-based drug design, the pharmacokinetic, pharmacodynamic and toxicological consequences of drug metabolism cannot be ignored. Drug metabolism is also of interest to medicinal chemists in the design of drugs with controlled, predictable deactivation after achieving the therapeutic objective in prodrug design and in chemical-enzymatic drug targeting. In this review, the authors provide an overview of concepts that can be utilized from drug discovery to pharmaceutical development to overcome problems associated with drug metabolism, or that may be used to take advantage of 'designed-in' metabolic activation to achieve drug targeting. PMID- 10542393 TI - Monitor: progress and profiles. PMID- 10542394 TI - Antagonistic Analogs of Growth Hormone-releasing Hormone: New Potential Antitumor Agents. AB - Recently, new potent antagonistic analogs of growth hormone-releasing hormone (GH RH) have been synthesized. These GH-RH antagonists bind to pituitary receptors for GH-RH and inhibit the release of GH in vitro and in vivo. This suggests that they could be clinically useful in conditions such as acromegaly. The main applications of GH-RH antagonists would be in the field of insulin-like growth factor I (IGF-I)- and IGF-II-dependent cancers. GH-RH antagonists inhibit the growth of various human cancer cell lines xenografted into nude mice, including mammary cancers, androgen-independent prostate cancers, small-cell lung carcinomas, non-small-cell lung carcinomas, renal adenocarcinomas, pancreatic cancers, colorectal carcinomas and malignant gliomas. These effects could, in part, be exerted indirectly through inhibition of the secretion of GH and the resulting reduction in levels of hepatic IGF-I. However, the principal action of GH-RH antagonists in vivo appears to be the direct suppression of the autocrine and/or paracrine production and expression of the genes encoding IGF-I (IGF1) and IGF-II (IGF2) in tumors. In vitro, antagonists of GH-RH inhibit the proliferation of mammary, prostatic, pancreatic and colorectal cancer cell lines, reducing the expression of IGF2 mRNA in the cells and the secretion of IGF-II. The presence of the GH-RH ligand has been demonstrated in human ovarian, endometrial, mammary and lung cancers, suggesting that GH-RH could be a growth factor. Further development of GH-RH antagonists should lead to potential therapeutic agents for IGF dependent cancers. PMID- 10542395 TI - Signaling Mechanism of the AT2 Angiotensin II Receptor: Crosstalk between AT1 and AT2 Receptors in Cell Growth. AB - The peptide angiotensin (Ang) II exerts a range of actions in the cardiovascular, renal, reproductive and central nervous systems. At least two distinct Ang II receptor subtypes have been defined and designated as type 1 (AT1) and type 2 (AT2). The function and signaling mechanism of these receptor subtypes are quite different, and these receptors exert opposite effects on cell growth. PMID- 10542396 TI - Mechanisms of Glucocorticoid-receptor-mediated Repression of Gene Expression. AB - It is hoped that this review will give the reader a taste of some of the mechanisms used by the glucocorticoid receptor to repress gene function. These mechanisms include direct binding to DNA, antagonism of other transcription factor families and sequestration of necessary cofactors. Each of these mechanisms, and others, are discussed. PMID- 10542397 TI - Multimeric Coactivator Complexes for Steroid/Nuclear Receptors. AB - Nuclear receptors regulate transcription in direct response to their cognate hormonal ligands. Ligand binding leads to the dissociation of corepressors and the recruitment of coactivators. Many of these factors, acting in large complexes, have emerged as chromatin remodelers through intrinsic histone modifying activities or through other novel functions. In addition, other ligand recruited complexes appear to act more directly on the transcriptional apparatus, suggesting that transcriptional regulation by nuclear receptors might involve a process of both chromatin alterations and direct recruitment of key initiation components at regulated promoters. PMID- 10542398 TI - Spatial Compartmentalization in the Regulation of Glucose Metabolism by Insulin. AB - Despite intense investigation, major gaps remain in our understanding of the cellular mechanisms that underlie the actions of insulin, as well as the regulation of the enzymes and transport proteins crucial to the orderly control of glucose metabolism. In recent years, the compartmentalization of signaling molecules and metabolic enzymes has been suggested to play an important role in ensuring metabolic balance. We will discuss examples of recent findings, suggesting that spatial compartmentalization and protein translocation might be the keys to understanding the specificity of insulin in the regulation of glucose metabolism. PMID- 10542399 TI - Real-time Analysis of Glucose Metabolism by Microscopy. AB - Glucose metabolism has traditionally been assayed via biochemical means. Fluorescence monitoring of NAD(P)H levels has provided a non-invasive method to assay glucose metabolism in cells and tissues. However, these measurements have traditionally been of low resolution (no subcellular information) because of limitations imposed by optical and cellular photodamage problems. The recent advent of two-photon excitation microscopy as a dependable tool for biological research has opened the possibility of real-time, high-resolution analysis of glucose metabolism in living cells. Such measurements have the potential to provide subcellular information from intact tissue that cannot be obtained by other techniques. PMID- 10542400 TI - Vitamin D and Hip Fracture. AB - Hip fractures are recognized as a major public health problem worldwide. Demographic changes will lead to enormous increases in the number of hip fractures and projections indicate that the number of hip fractures occurring worldwide each year will rise from 1.26 million in 1990 to 4.5 million by 2050. However, preventive strategies are available. Supplementation with Ca2+ and vitamin D restores bone quality through suppression of secondary hyperparathyroidism and decreases the risk of falling through improvement of neuromuscular coordination and body sway. This leads to a reduction of hip fracture risk of 43% in the vitamin D-insufficient elderly. Treatment with the bisphosphonate alendronate increases bone strength and results in a 51% reduction of hip fracture risk. Hip protectors absorb energy during a fall and reduce hip fracture risk by 56%. Combining these three procedures could prevent a large proportion of hip fractures in the future. PMID- 10542401 TI - Divergence of RGS proteins: evidence for the existence of six mammalian RGS subfamilies. PMID- 10542402 TI - Discovering new ADP-ribose polymer cycles: protecting the genome and more. PMID- 10542403 TI - Multistability: a major means of differentiation and evolution in biological systems. AB - Very simple biochemical systems regulated at the level of gene expression or protein function are capable of complex dynamic behaviour. Among the various patterns of regulation associated with non-linear kinetics, multistability, which corresponds to a true switch between alternate steady states, allows a graded signal to be turned into a discontinuous evolution of the system along several possible distinct pathways, which can be either reversible or irreversible. Multistability plays a significant role in some of the basic processes of life. It might account for maintenance of phenotypic differences in the absence of genetic or environmental differences, as has been demonstrated experimentally for the regulation of the lactose operon in Escherichia coli. Cell differentiation might also be explained as multistability. PMID- 10542404 TI - Origin of eukaryotic cells: was metabolic symbiosis based on hydrogen the driving force? PMID- 10542406 TI - The SPFH domain: implicated in regulating targeted protein turnover in stomatins and other membrane-associated proteins. PMID- 10542408 TI - How membrane proteins travel across the mitochondrial intermembrane space. AB - A newly discovered family of small proteins in the yeast mitochondrial intermembrane space mediates import of hydrophobic proteins from the cytoplasm into the inner membrane. Loss of one of these chaperone-like proteins from human mitochondria results in a disease that causes deafness, muscle weakness and blindness. PMID- 10542409 TI - Progress in protrusion: the tell-tale scar. AB - The crawling movement of a cell involves protrusion of its leading edge, in coordination with the translocation of its cell body, and depends upon a cytoplasmic machinery able to respond to signals from the environment. Protrusion is now understood to be driven by actin polymerization, and signalling from membrane receptors to actin has been shown to be mediated by the Rho family of GTPases. However, a major gap in our understanding of regulated motility has been how to connect the signalling pathway to the motile machinery itself. Recent structural, biochemical and genetic studies have identified some of the missing links and provided a strong working model for the pathways and mechanisms by which the signals are interpreted and implemented. PMID- 10542412 TI - The PH superfold: a structural scaffold for multiple functions. AB - Pleckstrin homology (PH) domains form a structurally conserved family that is associated with many regulatory pathways within the cell. Domains with a nearly identical fold are found in other families that share no sequence similarity, suggesting the existence of a stable PH superfold. The PH domains generally function as regulated membrane-binding modules that bind to inositol lipids and respond to upstream signals by targeting the host proteins to the correct cellular sites. The other domains with a similar fold, such as the phosphotyrosine binding domains, recognize protein ligands. PMID- 10542411 TI - The economics of ribosome biosynthesis in yeast. AB - In a rapidly growing yeast cell, 60% of total transcription is devoted to ribosomal RNA, and 50% of RNA polymerase II transcription and 90% of mRNA splicing are devoted to ribosomal proteins (RPs). Coordinate regulation of the approximately 150 rRNA genes and 137 RP genes that make such prodigious use of resources is essential for the economy of the cell. This is entrusted to a number of signal transduction pathways that can abruptly induce or silence the ribosomal genes, leading to major implications for the expression of other genes as well. PMID- 10542413 TI - Chain-length determination mechanism of isoprenyl diphosphate synthases and implications for molecular evolution. AB - In the synthesis of isoprenoids, isoprenyl diphosphate synthases catalyze the consecutive condensation of isopentenyl diphosphate with allylic diphosphates to produce a variety of prenyl diphosphates with well-defined chain lengths. Site directed mutagenesis in conjunction with X-ray crystallographic studies have identified specific amino acid residues responsible for chain-length determination. Simple combinations of these residues within a characteristic motif are not only sufficient to confer product specificities to all isoprenyl diphosphate synthases but represent structural features that reflect the enzyme family's evolutionary course. PMID- 10542414 TI - Phosphorelay signal transduction: the emerging family of plant response regulators. AB - Homologs of bacterial two-component signal transduction elements are emerging as key players in eukaryotic signaling systems. The recent identification of a large gene family in Arabidopsis that is similar to two-component response regulators emphasizes the importance of this signaling mechanism in plants. The understanding of the function of these response regulator genes is only rudimentary but the transcriptional induction of a subset by cytokinin suggests a role for some of these regulators in the response to this important plant hormone. PMID- 10542415 TI - Rapid-flow techniques and their contributions to enzymology. PMID- 10542416 TI - DNA switches for thermal control of gene expression. PMID- 10542417 TI - Entamoeba histolytica: a derived, mitochondriate eukaryote? PMID- 10542418 TI - Regulation of virulence factor expression in group A streptococcus. PMID- 10542419 TI - The pathogenic potential of endogenous retroviruses: a sceptical view. PMID- 10542421 TI - Response from Lower PMID- 10542420 TI - Human endogenous retroviruses and pathogenicity: genomic considerations. PMID- 10542422 TI - The photosynthetic apparatus of Rhodobacter sphaeroides. AB - Functional and ultrastructural studies have indicated that the components of the photosynthetic apparatus of Rhodobacter sphaeroides are highly organized. This organization favors rapid electron transfer that is unimpeded by reactant diffusion. The light-harvesting complexes only partially surround the photochemical reaction center, which ensures an efficient shuttling of quinones between the photochemical reaction center and the bc1 complex. PMID- 10542424 TI - Pressure-regulated metabolism in microorganisms. AB - There has been a renewal of interest in the survival strategies employed by deep sea, high-pressure-adapted (piezophilic) microorganisms as well as in the effects of high pressure on mesophilic, 1-atmosphere-pressure-adapted microorganisms. This is partly the result of a greater appreciation of the adaptations of microorganisms to life in extreme environments and partly the result of the development of new techniques for examining physiological and molecular processes as a function of pressure. PMID- 10542423 TI - Hydrogenosomes: eukaryotic adaptations to anaerobic environments. AB - Like mitochondria, hydrogenosomes compartmentalize crucial steps of eukaryotic energy metabolism; however, this compartmentalization differs substantially between mitochondriate aerobes and hydrogenosome-containing anaerobes. Because hydrogenosomes have arisen independently in different lineages of eukaryotic microorganisms, comparative analysis of the various types of hydrogenosomes can provide insights into the functional and evolutionary aspects of compartmentalized energy metabolism in unicellular eukaryotes. PMID- 10542425 TI - The EGFR as a target for viral oncoproteins. AB - The epidermal growth factor receptor (EGFR) is a potent stimulator of the mitogen activated protein kinase (MAPK) signaling pathway. Chronic stimulation of the EGFR and of multiple steps in the MAPK signaling pathway is involved in the development of cancer. Several tumor viruses encode proteins that induce EGFR expression or stimulate EGFR-mediated signaling and are thus likely to play an important role in the transformation of virus-infected cells. PMID- 10542426 TI - Stress resistance in Staphylococcus aureus. AB - Staphylococcus aureus is a major human pathogen of increasing importance as a result of the spread of antibiotic resistance. It causes a wide range of diseases and survives outside the host by virtue of its adaptability and resistance to environmental stress. Several cellular components involved in Staphylococcus aureus stress resistance have begun to be characterized. PMID- 10542427 TI - Retinotectal maps: molecules, models and misplaced data. AB - The mechanisms underlying the formation of topographic maps in the retinotectal system have long been debated. Recently, members of the Eph and ephrin receptor ligand family have been found to provide a molecular substrate for one type of mechanism, that of chemospecific gradient matching, as proposed by Sperry. However, experiments over several decades have demonstrated that there is more to map formation than gradient matching. This article briefly reviews the old and new findings, argues that these two types of data must be properly integrated in order to understand map formation fully, and suggests some experimental and theoretical ways to begin this process. PMID- 10542428 TI - Clinical trials with neuroprotective drugs in acute ischaemic stroke: are we doing the right thing? AB - Ischaemic stroke is a leading cause of death and long-lasting disability. Several neuroprotective drugs have been developed that have the potential to limit ischaemic brain damage and improve outcome for patients. While promising results with these drugs have been achieved in animal stroke models, all Phase III trials conducted so far indicate that these drugs have failed to live up to their promise. Despite the limits of animal models, which cannot mimic the clinical situation, the disappointing results of neuroprotective trials might largely be due to methodological problems. Future trials with neuroprotective drugs should be performed in stroke (care) units, after sufficient information regarding therapeutic time window, dosage, duration of therapy and safety has been gathered from pilot studies, and a better selection of target patients has been made. Much of this information can now be obtained by techniques that visualize the penumbra, such as combined diffusion-weighted and perfusion MRI. Consideration should also be given to clinical trials with well-designed combinations of treatments. PMID- 10542430 TI - Consciousness in mind: a correlate for ACh? PMID- 10542429 TI - Cholinergic correlates of consciousness: from mind to molecules. PMID- 10542431 TI - Reply to Holscher PMID- 10542432 TI - Estimation of information in neuronal responses. PMID- 10542433 TI - Reply to victor and purpura PMID- 10542434 TI - Tracking with the mind's eye. AB - The two components of voluntary tracking eye-movements in primates, pursuit and saccades, are generally viewed as relatively independent oculomotor subsystems that move the eyes in different ways using independent visual information. Although saccades have long been known to be guided by visual processes related to perception and cognition, only recently have psychophysical and physiological studies provided compelling evidence that pursuit is also guided by such higher order visual processes, rather than by the raw retinal stimulus. Pursuit and saccades also do not appear to be entirely independent anatomical systems, but involve overlapping neural mechanisms that might be important for coordinating these two types of eye movement during the tracking of a selected visual object. Given that the recovery of objects from real-world images is inherently ambiguous, guiding both pursuit and saccades with perception could represent an explicit strategy for ensuring that these two motor actions are driven by a single visual interpretation. PMID- 10542435 TI - Sensing effectors make sense. AB - 'Housekeepers' of living organisms maintain salt and water balance, monitor blood sugar and schedule their work to the season and the time of day. In order to perform their chores, they rely on information about the status quo. The traditional concept of a sensor that communicates with a central comparator authorizing an effector, which was inspired by engineers, has become blurred in the search for morphological correlates of such regulatory cascades. In many cases, neurones, which are both sensory and neurosecretory, and endocrine cells equipped with smart detectors, reliably regulate autonomous functions by using local rather than central computing. Like the well-trained staff of a smoothly run household, such 'sensing effectors' translate information into action. PMID- 10542436 TI - Nicotinic receptors in the brain: correlating physiology with function. AB - Nicotinic ACh receptors (nAChRs) have been implicated in a variety of brain functions, including neuronal development, learning and memory formation, and reward. Although there are substantial data indicating that nAChR subunits are found in many brain regions, the precise cellular roles of these subunits in neuronal functions have remained elusive. Until recently, nAChRs were thought primarily to serve a modulatory role in the brain by regulating neurotransmitter release from nerve terminals. However, new evidence has revealed that nAChRs also function in a postsynaptic role by mediating fast ACh-mediated synaptic transmission in the hippocampus and in the sensory cortex, and are found at somatodendritic as well as nerve terminal sites in the reward system. It is possible that presynaptic and postsynaptic nAChRs mediate changes in the efficacy of synaptic transmission in these brain regions. These changes could underlie the proposed functions of nAChRs in cognitive functions of the hippocampus and cerebral cortex, in neuronal development in the sensory cortex, and in reward. PMID- 10542439 TI - Inhibitory monoclonal antibodies to human cytochrome P450 enzymes: a new avenue for drug discovery. PMID- 10542437 TI - Long-term potentiation in the amygdala: a mechanism for emotional learning and memory. AB - In the mammalian brain, LTP is an enduring form of synaptic plasticity that is posited to have a role in learning and memory. Compelling new evidence for this view derives from studies of LTP in the amygdala, a brain structure that is essential for simple forms of emotional learning and memory, such as Pavlovian fear conditioning in rats. More specifically, antagonists of the NMDA receptor block both amygdaloid LTP induction and fear conditioning, fear conditioning induces increases in amygdaloid synaptic transmission that resemble LTP, and genetic modifications that disrupt amygdaloid LTP eliminate fear conditioning. Collectively, these results provide the most-convincing evidence to date that LTP mediates learning and memory in mammals. PMID- 10542440 TI - Protective activity of adenosine receptor agonists in the treatment of organophosphate poisoning. PMID- 10542441 TI - The female sex hormone oestrogen as a neuroprotectant. AB - It is well recognized that oestrogen regulates sex differentiation and maturation of sex organs via binding to specific intracellular receptors. However, oestrogen receptors (ERs) are expressed in a variety of other tissues, including the nervous system, which suggests that oestrogen's effects are not limited to primary and secondary sex organs. Increasing evidence supports the role of oestrogen as a neuroprotective compound that can act dependently or independently of ER activation; oestrogen has recently been shown to exhibit intrinsic antioxidant activity that is ER independent. Thus, oestrogen might represent a potential 'chemical shield' for neurones. In this article, some recent advances in the elucidation of oestrogen's beneficial activities on nerve cell survival are discussed. PMID- 10542442 TI - A unified nomenclature for short-chain peptides isolated from scorpion venoms: alpha-KTx molecular subfamilies. AB - Peptidyl toxins are used extensively to determine the pharmacology of ion channels. Four families of peptides have been purified from scorpion venom. In this article, the classification of K+-channel-blocking peptides belonging to family 2 peptides and comprising 30-40 amino acids linked by three or four disulfide bridges, will be discussed. Evidence is provided for the existence of 12 molecular subfamilies, named alpha-KTx1-12, containing 49 different peptides. Because of the pharmacological divergence of these peptides, the principle of classification was based on a primary sequence alignment, combined with maximum parsimony and Neighbour-Joining analysis. PMID- 10542443 TI - Corrigendum PMID- 10542444 TI - Molecular pharmacology of the CFTR Cl- channel. AB - Dysfunction of the cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel is associated with a wide spectrum of disease. In the search for modulators of CFTR, pharmacological agents that interact directly with the CFTR Cl- channel have been identified. Some agents stimulate CFTR by interacting with the nucleotide-binding domains that control channel gating, whereas others inhibit CFTR by binding within the channel pore and preventing Cl- permeation. Knowledge of the molecular pharmacology of CFTR might lead to new treatments for diseases caused by the dysfunction of CFTR. PMID- 10542445 TI - Ecto-protein kinases: ecto-domain phosphorylation as a novel target for pharmacological manipulation? AB - An increasing number of studies document the presence of protein kinases facing outwards at the cell surface of a diverse array of cells. These ecto-protein kinases phosphorylate cell-surface proteins and soluble extracellular substrates, and thus could affect many physiological processes involving cell-cell contacts, cellular differentiation and proliferation, ion fluxes and cellular activation. To date, only limited attention has been paid to exploring ecto-protein kinases as possible pharmacological targets. Here, the identification and physiological role of ecto-protein kinases in different biological systems is described; it is suggested that ecto-protein kinases are attractive and novel candidates for pharmacological manipulation under various (patho)physiological conditions. PMID- 10542446 TI - Novel natural vanilloid receptor agonists: new therapeutic targets for drug development. AB - The discovery that compounds lacking a recognizable vanillyl-like motif might act as vanilloids has given new impetus to a search for novel vanilloid receptor agonists and antagonists in compound libraries. The availability of cell lines transfected with a cloned human vanilloid receptor will further expedite this search. In this article, the pharmacological properties of unsaturated dialdehydes and triprenyl phenols that represent two newly discovered chemical classes of vanilloids will be discussed. The existence of vanilloid receptors in several brain nuclei as well as in non-neuronal tissues predicts novel, innovative therapeutic indications for vanilloids. However, these findings also suggest that vanilloids might cause side-effects. An exploration of the uses of unsaturated dialdehydes in indigenous medicine might help identify new therapeutic targets for vanilloids and avoid unwanted actions. PMID- 10542448 TI - Ants, agriculture and antibiotics. PMID- 10542447 TI - Cyclooxygenase 2 inhibitors: discovery, selectivity and the future. AB - The recent marketing of two selective cyclooxygenase 2 (COX-2) inhibitors climaxes the first phase of an exciting and fast-paced effort to exploit a novel molecular target for nonsteroidal anti-inflammatory drugs (NSAIDs). Much has been written in the lay and scientific press about the potential of COX-2 inhibitors as anti-inflammatory and analgesic agents that lack the gastrointestinal side effects of traditional NSAIDs. Although research on COX-2 inhibitors has focussed mainly on inflammation and pain, experimental and epidemiological data suggest that COX-2 inhibitors could be used in the treatment or prevention of a broader range of diseases. In this review, some key points and unresolved issues related to the discovery of COX-2 inhibitors, the kinetic and structural basis for their selectivity, and possible complications in their development and use will be discussed. PMID- 10542449 TI - The paradox of the stickleback: different yet the same. PMID- 10542450 TI - The names of animals. PMID- 10542451 TI - Sticklers for sympatry. PMID- 10542452 TI - Elasticity analysis as an important tool in evolutionary and population ecology. AB - Elasticity analysis estimates the proportional change in the population growth rate for a proportional change in a vital rate (i.e. survival, growth or reproduction). It can be used to pinpoint those parts of an organism's life history that should be the focus of management effort, or those that contribute most to fitness. Recent theoretical work has emphasized some limitations of the technique, has overcome other problems, and has shown that it is robust to some violations of its underlying assumptions. Thus, although care is needed, elasticity analysis is a simple first step in answering important questions in evolutionary and population ecology. PMID- 10542453 TI - Life at the front: history, ecology and change on southern ocean islands. AB - Terrestrial ecosystems of southern ocean islands have enjoyed renewed attention recently owing to the discovery that their climates are changing dramatically. This has led to an enhanced understanding of the biogeography of this region, and an increased awareness that these ecosystems provide unrivalled opportunities for investigating the impacts of environmental change on interactions between invasive and indigenous species. Recent studies have revealed increases in the abundance of established alien species and in the strength of their negative impacts on local biota, especially through indirect interactions. Also, increases in island temperature and human visitor frequency are likely to result in increasing numbers of successful alien colonization events. PMID- 10542454 TI - Cooperation among unrelated individuals: the ant foundress case. AB - Ant foundress associations are an example of cooperation among non-kin. Across a dozen genera, queens able to found a colony alone often join unrelated queens, thereby enhancing worker production and colony survivorship. The benefits of joining other queens vary with group size and ecological conditions. However, after the first workers mature, the queens fight until only one survives. The presence of cofoundresses, and their relative fighting ability, also affects the extent of cooperative investment before worker emergence. This reveals previously overlooked early conflicts among queens, which reduce the mutualistic benefits of cooperation. PMID- 10542456 TI - Other ant invaders. PMID- 10542455 TI - Trophic cascades revealed in diverse ecosystems. AB - New studies are documenting trophic cascades in theoretically unlikely systems such as tropical forests and the open ocean. Together with increasing evidence of cascades, there is a deepening understanding of the conditions that promote and inhibit the transmission of predatory effects. These conditions include the relative productivity of ecosystems, presence of refuges and the potential for compensation. However, trophic cascades are also altered by humans. Analyses of the extirpation of large animals reveal loss of cascades, and the potential of conservation to restore not only predator populations but also the ecosystem level effects that ramify from their presence. PMID- 10542457 TI - Reply from D.A. Holway and A.V. Suarez. PMID- 10542458 TI - Relating populations to habitats. PMID- 10542459 TI - Reply from M.S. Boyce, L.L. McDonald and B.F.J. Manly. PMID- 10542460 TI - The behavior-conservation interface. PMID- 10542461 TI - The evolution of mating systems in tropical reef corals. AB - The life histories of tropical reef corals (Scleractinia) include two traits that can strongly bias mating systems towards inbreeding: (1) most species express both sexes simultaneously, creating the potential for self-fertilization; and (2) there is philopatric dispersal of planktonic or demersal larvae. Recent studies have confirmed that all hermaphrodite species with broad dispersal potential are either completely, or almost completely, self-incompatible. By contrast, species with limited dispersal potential have high, but variable, rates of self fertilization. This interspecific variation in coral mating systems is similar to that found in terrestrial plants. Understanding the selective forces that drive mating-system variation in marine environments will undoubtedly broaden our understanding of the evolution of inbreeding and outbreeding in sessile plants and animals. PMID- 10542463 TI - Persistence without intervention: assessing success in wildlife reintroductions. PMID- 10542462 TI - The modern synthesis, Ronald Fisher and creationism. AB - The 'modern evolutionary synthesis' convinced most biologists that natural selection was the only directive influence on adaptive evolution. Today, however, dissatisfaction with the synthesis is widespread, and creationists and antidarwinians are multiplying. The central problem with the synthesis is its failure to show (or to provide distinct signs) that natural selection of random mutations could account for observed levels of adaptation. PMID- 10542464 TI - [ [In Process Citation] PMID- 10542465 TI - A theoretical model of the evolution of virulence in sexually transmitted HIV/AIDS. AB - INTRODUCTION: The evolution of virulence in host-parasite relationships has been the subject of several publications. In the case of HIV virulence, some authors suggest that the evolution of HIV virulence correlates with the rate of acquisition of new sexual partners. In contrast some other authors argue that the level of HIV virulence is independent of the sexual activity of the host population. METHODS: Provide a mathematical model for the study of the potential influence of human sexual behaviour on the evolution of virulence of HIV is provided. RESULTS: The results indicated that, when the probability of acquisition of infection is a function both of the sexual activity and of the virulence level of HIV strains, the evolution of HIV virulence correlates positively with the rate of acquisition of new sexual partners. CONCLUSION: It is concluded that in the case of a host population with a low (high) rate of exchange of sexual partners the evolution of HIV virulence is such that the less (more) virulent strain prevails. PMID- 10542466 TI - [Tolerance and resistance: abortion from the point of view of traditional midwives in a rural area of Mexico]. AB - OBJECTIVE: An evaluation of the perception, resources and practices regarding abortion of traditional midwives in a rural area in the municipality of Yecapixtla, state of Morelos, located in the central region of Mexico. METHODS: A qualitative methodology consisting of a detailed interview, focal groups and participating observation, was used. The subjects investigated were socio cultural aspects, reproduction, sexuality and health related to abortion. Nine midwives were interviewed and a focal group was formed in which 16 midwives participated. RESULTS: The results demonstrated a profound rejection of abortion whether inducted or spontaneous. The former was considered a major sin and the latter a serious failure of a womans reproductive function. Women who abort are called "pigs", "hogs" or "bitches" and the midwives are reluctant to attend them. However, a common practice among the women in the community is to "regulate the menstruation", that is, to use substances that provoke menstruation when this is delayed. This specific practice is not considered abortive by these women. CONCLUSION: Local popular beliefs about abortion are indispensible for the construction of effective strategies, which when provided by the institutional health services, reinforce the bonds between these and the traditional midwives in such a way as to increase accessibility to the health services as well as the quality of care to women. PMID- 10542467 TI - [Factors associated to the trial of labor in primipara women with one previous cesarean section]. AB - OBJECTIVE: The purpose of this study was to identify medical and non-medical factors associated to the performance of a trial of labor during the second delivery of women with one previous cesarean section. METHODS: This was a nested case control study, with a secondary data analysis from a retrospective population based cohort study. It was primarily performed on a population of women who had had their first children in Campinas, SP, Brazil, during 1985. The study population was constituted of the 1,352 women of the cohort study who had had their first deliveries by cesarean section and also had their second deliveries no matter when. The group of cases (333 women, almost 25%) was constituted of those women who had a trial of labor during their second deliveries and the control group (1,019 women) of those who had not had it. For each possible associated factor evaluated, the Odds Ratio and its respective 95% Confidence Interval were calculated. For the ordered categorical variables, the chi2 for trend was also calculated. Finally, a non conditional multivariate regression analysis was performed, identifying the significant factors and then estimating their adjusted Odds Ratio. RESULTS: The main factors associated with the trial of labor in this situation were a low monthly family income, having public medical insurance by the national health system, a low maternal age, the occurrence of rupture of membranes during the second delivery, and having been in childbirth during the first delivery. CONCLUSION: It is concluded that the main determinants for a trial of labor among primipara women with one previous cesarean section were basically social and economic factors, rather than medical ones. PMID- 10542468 TI - Assessing morbidity in the paediatric community. AB - INTRODUCTION: Morbidity information is easily available from medical records but its scope is limited to the population attended by the health services. Information on the prevalence of diseases requires community surveys, which are not always feasible. These two sources of information represent two alternative assessments of disease occurrence, namely demand morbidity and perceived morbidity. The present study was conceived so as to elicit a potential relationship between them so that the former could be used in the absence of the latter. METHODS: A community of 13,365 families on the outskirts of S. Paulo, Brazil, was studied during the period from 15/Nov/1994 to 15/Jan/1995. Data regarding children less than 5 years old were collected from a household survey and from the 2 basic health units in the area. Prevalence of diseases was ascertained from perceived morbidity and compared to estimates computed from demand morbidity. RESULTS: Data analysis distinguished 2 age groups, infants less than 1 year old and children 1 to less than 5. The most important groups of diseases were respiratory diseases, diarrhoea, skin problems and infectious & parasitical diseases. Basic health units presented a better coverage for infants. Though disease frequencies were not different within or outside these units, a better coverage was found for diarrhoea and infectious & parasitical diseases in the infant group, and for diarrhoea in the older age group. Equivalence between the two types of morbidity was found to be limited to the infant group and concerned only the best covered diseases. The odds of a disease being seen at the health service should be of at least 4:10 to ensure this equivalence. CONCLUSION: It was concluded that, provided that health service coverage is good, demand morbidity can be taken as a reliable estimate of community morbidity. PMID- 10542469 TI - [Health and nutritional status of infants in a population of the Center-Western region of Brazil]. AB - OBJECTIVE: To evaluate the nutritional status of infants who reside in the city of Campo Grande, State of Mato Grosso do Sul, Brazil. METHODS: The method used was that of a cross sectional household study by means of an anthropometric social survey, sampling 652 children from 0 to 59 months of age. RESULTS: A low prevalence of nutritional deficit, excepting that of height-for-age, starting in the first year of life, was found. The nutritional status proved to be influenced by the socioeconomic conditions, especially concerning per capita family income. Nearly all children started breast-feeding but were weaned during the first month. Exclusive breast-feeding is of short duration and soon replaced by infant formula. The survey of mother-child assistance demonstrated an excellent coverage of prenatal care, but inadequacy in the follow-up of the child's health at all income levels. CONCLUSIONS: The need to carry out changes in the approach to preventive actions and in the monitoring of the nutritional situation of the children, concerning the problems identified in this study in order to allow for the development of differential actions in the nutritional field has been identified. PMID- 10542470 TI - [Mortality among children of less than one year of age in the Southwestern region of Brazil]. AB - OBJECTIVE: A description of infant mortality in children of less than one year of age on the basis of official systems of information. METHODS: The data were obtained from two official information systems of the Health Ministry: of mortality and live-births. Health service and home visits were carried out to complete the information on death certificates. RESULTS: Infant mortality was 49.7. It was higher among boys, low birthweight children, twin births, among those born by cesarean section, resident in the outskirts, among children of mothers of little schooling, greater number of children on above 35 years of age. The data on death certificates were poorly reported. Data on birth certificates were complete. CONCLUSION: The results suggested that a health information system should be implemented, with a view to reducing infant mortality. PMID- 10542471 TI - [Measles seroprevalence among pediatricians in a teaching hospital]. AB - OBJECTIVE: A measles outbreak occurred in S. Paulo state, during 1996 and 1997, resulting in 20,921 cases. Forty seven percent of the cases occurred in people between 20 and 29 years of age, and one of the control strategies of the Department of Health was the vaccination of health care workers. The prevalence of antibodies against measles among the hospital pediatricians was investigated. METHODS: One hundred and fifty samples were taken from volunteer pediatricians to test for measles antibodies using ELISA. A questionnaire about their having had measles and the vaccine was filled out. RESULTS: Of the 150 doctors, 122 (81.4%) were female and 28 (18.6%) male, of between 23 and 46 years of age (mean and median 27 years). The majority (98%-147/150) had protective levels of antibodies against measles (>100 UI/ml); 118 (80.3%-118/147) without and 29 (19.7%-29/147) with a history of measles. Only 3 pediatricians (2%-3/150), had negative serology, 2 without and 1 with a history of measles. Out of the 118 without history of measles, 79 (67%-39/118) in spite of the protective level of antibodies against measles, did not know if they, had been vaccinated. Out of the 79 vaccinated pediatricians, 64 (81%-64/79), had been vaccinated 25 years before, and still maintained protective levels of antibodies. Of the 3 doctors with negative serologies only one declared that he had been vaccinated. CONCLUSIONS: Measles seroprevalence among pediatricians of this hospital is high, especially due to preceding vaccination. On the other hand, the 2% of pediatricians with negative serology, in an epidemic situation could constitute a significant population for the acquisition and dissemination of the disease. PMID- 10542472 TI - [Monitoring of parasites in domestic sewage]. AB - OBJECTIVE: The evaluation of the presence of parasites in semisolid and liquid sewage in Argentinian Patagonia in view of the fact that this is a restriction for its use. METHODS: The samples taken at 4 Domestic Sewage Plants were analyzed in accordance with Standard Methods for the Examination of Water and Wastewater, Environmental Protection Agency, World Health Organization and some other classifications. RESULTS: Only 2 of 6 semisolid samples analyzed had non-viable Ascaris lumbricoides eggs. Of the 10 liquid samples analyzed, only 2 did not contain eggs whereas the remaining ones had pathogens of categories I (Giardia sp., Hymenolepis nana and Enterobius vermicularis) and III (Ascaris lumbricoides, Ancylostoma duodenale and Trichuris trichiura). CONCLUSIONS: All semisolid samples turned out to be satisfactory for use as fertilizer as no viable Ascaris lumbricoides eggs were found in any of them. But only 6 of the liquid samples were satisfactory for use as they had no eggs or their concentrations was equal to or less than 1 egg per litre. PMID- 10542473 TI - [Prevalence of dental caries and treatment needs in 6 to 12 year-old schoolchildren at public schools]. AB - OBJECTIVE: To assess dental caries prevalence and treatment needs of schoolchildren in the State of Goias, Brazil. METHODS: The study population consisted of 6-12-yr-old schoolchildren (n=1,419), male and female, attending 25 public schools located in the urban area of 9 provincial cities in the State of Goias. RESULTS: Percentage of caries-free schoolchildren was very low at all ages (4.4% at age 12). Mean DMF-T ranged from 0.41 at age 6 to 5.19 at age 12. Mean dmf-t in this age group ranged from 4.93 to 0.29. Treatment needs were higher than the proportion of treated teeth in both deciduous and permanent dentition. CONCLUSION: The high prevalence of dental caries found in schoolchildren in the provincial cities of Goias suggests the need for oral health education and preventive programs targeted at the underlying causes of the disease on a population level. PMID- 10542474 TI - [A cross-sectional study about mental health of farm-workers from Serra Gaucha (Brazil)]. AB - OBJECTIVE: In view of the shortage of population-based rural studies, this research project evaluated the associations between the characteristics of rural work and the occurrence of minor psychiatric disorders (MPD). METHODS: A cross sectional study was carried out on the 1,282 farm workers of 446 farms. Information about the farms (land extension, agricultural activities, technology and pesticide use) was collected. Demographic and socioeconomic data, characteristics of the work process and mental health indicators were obtained from the workers. RESULTS: MPD were found in 37.5% of the farm workers. The risk was higher on farms with a land extension of from 26 to 50 hectares, and lower where there was an increased level of job technology and schooling. The prevalence of MPD was higher among bean producers and lower among apple producers. Despite the impossibility of defining the direction of the causal link, pesticide poisoning was strongly associated with MPD. CONCLUSION: The results call attention to the dimension of the problem and to the importance of adopting new policies for the protection of farm workers' mental health. PMID- 10542475 TI - [Assessment of a health program for the non-insured population]. AB - OBJECTIVE: Present the results of the evaluation of a program for the non-insured population of the four poorest states of the country implemented by the Ministry of Health of Mexico between 1991 and 1995. METHODS: The effects of the program were evaluated in three areas: i) increase in health services coverage; ii) delivery of personal health services, and iii) changes in health conditions of the target population. The extension of coverage was measured by the increase in potential access due to the construction of health infrastructure projects and the use of additional health manpower, mainly primary health care workers. For the evaluation of the impact of the program on the delivery of services, three surveys were developed: one for service utilization, another one for accessibility, and a third for quality of care. The impact on health conditions was evaluated by changes in health indicators of children under five and women of reproductive age. RESULTS AND CONCLUSIONS: The Program had a positive impact on coverage, accessibility and quality of services. Its impact on health conditions was also positive. However, these last changes cannot be attributed only to the program, but to the sum of several concurrent activities. PMID- 10542476 TI - [Human exposure to trihalomethanes in drinking water]. AB - Halogenated hydrocarbon compounds, some of them recognized as carcinogenic to different animal species can be found in drinking water. Chloroform, bromodichloromethane, dibromochloromethane and bromoform are the most important trihalomethanes found in potable water. They are produced in natural waters during chlorinated desinfection by the halogenation of precursors, specially humic and fulvic compounds. The review, in the MEDLINE covers the period from 1974 to 1998, presents the general aspects of the formation of trihalomethanes, sources of human exposure and their toxicological meaning for exposed organisms: toxicokinetic disposition and spectrum of toxic effects (carcinogenic, mutagenic and teratogenic). PMID- 10542477 TI - [Mathematical location models applied in the spatial organization of health units]. AB - Mathematical location models have been increasingly applied in the health services at the international level. In Brazil, although incipient, there exists an enormous potential for the use of such models in the area of public health. In this paper several location models that can be applied to public health are presented initially, and the location of non-emergency services, of emergency services and of services hierarchically related are analysed. A hierarchical model is then applied to the location of maternal and perinatal assistance in the municipality of Rio de Janeiro. In this part, after presenting some related data for the municipality, a four-level hierarchical model (location of out-patient units, maternity hospitals, neonatal hospitals and general hospitals) is proposed and the impact that the adoption of this methodology would have as compared with that of the present system is analysed. PMID- 10542484 TI - The disgrace of poor care. PMID- 10542485 TI - Not because they are old. PMID- 10542487 TI - Ageing in the USA. PMID- 10542486 TI - Devolution of care. PMID- 10542488 TI - Volunteering: a healthy choice? PMID- 10542489 TI - Three steps to effective wound care. AB - Optimal care of leg ulcers and pressure sores does not depend simply on the choice of dressing. The cause of the wound must be investigated, and other systemic factors related to wound healing must be assessed, then the wound environment can be evaluated and the most suitable dressing applied. Georgina Casey looks at these three steps to good wound care. PMID- 10542490 TI - Staying out. PMID- 10542491 TI - Individual choice. PMID- 10542493 TI - Elderly care reviews initiatives and resources related to nutrition and older people. PMID- 10542492 TI - Lifting. PMID- 10542494 TI - Taking charge of shifts. PMID- 10542495 TI - Promoting the three Rs: rehabilitation, recruitment and retention. PMID- 10542496 TI - Degrees of competence. PMID- 10542497 TI - Inspector nurse. PMID- 10542498 TI - Nursing care of Parkinson's disease. PMID- 10542500 TI - Thirty years of service. PMID- 10542499 TI - Parkinson's disease and the role of nurse specialists. PMID- 10542501 TI - High Court rules on care home funding. PMID- 10542502 TI - Opening up to the Katz Index. AB - As assessment of functional ability becomes more and more important to nurses as well as other members of the interdisciplinary team, the Katz index is a useful instrument to have available. The Katz Index is a simple instrument. This is both a strength and a deficit. The IADLs are not evaluated with the index. It has some deficits that make it less useful in rehabilitation and of more value in nursing and interdisciplinary research. In any case, it is a useful tool with which nurses should be familiar when seeking a way to measure functional ability of older adults EC. PMID- 10542503 TI - The basics of quality care. PMID- 10542504 TI - Remembering yesterday, caring today. PMID- 10542505 TI - Dementia. PMID- 10542506 TI - The colour coded Moorside home for older people. PMID- 10542508 TI - Major issues relating to older people in 1999. PMID- 10542507 TI - The blind hunter of dementia. PMID- 10542509 TI - Investing in continence. PMID- 10542510 TI - Strong men do cry. PMID- 10542512 TI - Altered body image. PMID- 10542511 TI - Sexuality and Parkinson's disease. PMID- 10542513 TI - A personal tragedy. PMID- 10542514 TI - Parkinson's aware in primary care. PMID- 10542515 TI - A vision for elderly care. PMID- 10542516 TI - Attitudes of student nurses to older patients in Nigeria. PMID- 10542517 TI - Conquering loneliness. AB - It is important to identify signs of clinical depression such as pessimism, refusal to eat, diminished concern about personal appearance, and reluctance to make decisions. Medical referral for clinical depression is imperative because there are medications and other interventions that can effectively alleviate the feelings of sadness and despair that accompany clinical depression. One must determine if loneliness is a symptom of depression or an emotional state that can be changed by one's own efforts. Whether the lonely person is the health care giver or a health care receiver, the ability to use creative strategies for coping with loneliness can turn loneliness into a more comfortable experience. PMID- 10542518 TI - Compression bandaging. PMID- 10542519 TI - Nursing homes. Examining policy--using experience. PMID- 10542520 TI - Nursing homes. Preferred providers. PMID- 10542521 TI - The future is bright--the future is older. PMID- 10542522 TI - RCN tackles ageism in nursing. PMID- 10542523 TI - Parkinson's disease. Drug treatment in old age. PMID- 10542525 TI - The risks of poisoning in later life. PMID- 10542524 TI - Parkinson's disease. Neurosurgical procedures. PMID- 10542526 TI - Assessing nutrition in older patients. PMID- 10542527 TI - Dehydration and electrolyte disturbance. PMID- 10542528 TI - A duty to protect. PMID- 10542529 TI - Developing improved care for patients with dementia. PMID- 10542530 TI - A rehabilitation programme for people with Parkinson's disease. PMID- 10542531 TI - Whose need does A&E serve? PMID- 10542532 TI - Can nurses remove spinal boards and cervical collars safely? PMID- 10542533 TI - Painless loss of vision. PMID- 10542534 TI - Key issues in nurse requested X-rays. PMID- 10542535 TI - Why A&E nurses feel inadequate in managing patients who deliberately self harm? PMID- 10542537 TI - Refusing to be anyone's victim. PMID- 10542536 TI - ENP scheme: highlighting the barriers. PMID- 10542538 TI - Braveheart. Constitutional change in Scotland. PMID- 10542539 TI - Risk management in major incident and emergency planning. PMID- 10542540 TI - I'm talking to you! Managing bullying and harassment in the workplace. PMID- 10542541 TI - Pesticide poisoning: herbicides. PMID- 10542542 TI - Electrical injuries. AB - Never underestimate the severity of the injury. History taking must be meticulous and thorough. Accurately assess percent of skin loss--fluid resuscitate as necessary. Monitor for the presence of cardiac arrhythmias. Observe for the development of compartment syndrome. Observe volume and colour of passed urine. Seek expert opinion. PMID- 10542543 TI - [Case report and nursing report of a child with lymphangioma]. PMID- 10542544 TI - [Accidents in children--causes and tips for prevention]. PMID- 10542546 TI - [Feasible limits--medical ethics and modern biomedicine]. PMID- 10542545 TI - [Protection from infectious diseases]. PMID- 10542547 TI - [Mentoring first year nursing students. Trying out a school project]. PMID- 10542548 TI - [Quality management in neonatal intensive care]. PMID- 10542549 TI - [Reuse of medical one-way products may become problematic]. PMID- 10542550 TI - [Under what circumstances should antipyretics be used in children with febrile infections if there are no risk factors for febrile convulsions]. PMID- 10542551 TI - [Sepsis and multiple organ failure in surgery]. PMID- 10542553 TI - [Latex allergy: diagnosis, clinical aspects, therapy and prevention]. PMID- 10542552 TI - [Simple treatment of migraine pain]. PMID- 10542554 TI - [Areas of care in the operating room]. PMID- 10542555 TI - [Scientific study confirms experience]. PMID- 10542556 TI - [How effective is beta-sistosterin?]. PMID- 10542557 TI - [Effect of beta-sitosterin on the growth of the prostate]. PMID- 10542558 TI - [New social legislation in Denmark since 1996. Preventive nursing and geriatric care]. PMID- 10542559 TI - [Search for factors preventing wound healing. A motivating therapy concept in chronic decubitus ulcers]. PMID- 10542561 TI - [Skin protection for intensive care patients]. PMID- 10542560 TI - [How much fasting is necessary before and after anesthesia?]. PMID- 10542562 TI - [Safety in blood sugar measurements has to be guaranteed. Test strips must be the original thing]. PMID- 10542564 TI - [Disturbed wellbeing or illness: are men with hormone deficiencies taken seriously]. PMID- 10542563 TI - [Use of EMLA Creme in leg ulcers: usefulness and limits]. PMID- 10542565 TI - [Helicobacter pylori infections: current evaluation of their relevance]. PMID- 10542566 TI - [Report from a meeting--what many physicians do not know: insufficient treatment of Helicobacter pylori infections]. PMID- 10542567 TI - [Alzheimer's disease: a breakthrough in research?]. PMID- 10542569 TI - [New findings in vacuum sealing of coloplast]. PMID- 10542568 TI - [Modern insulin mixtures now also for type 2 diabetics]. PMID- 10542570 TI - [The realities of dementia therapy]. PMID- 10542571 TI - [A new test for the early diagnosis of dementias]. PMID- 10542572 TI - [Patients' experiences with insulin lispro in daily use. Yesterday's innovation- today's standard therapy]. PMID- 10542573 TI - Massage therapy: Part II. PMID- 10542574 TI - Population-focused care: a new rubric in the role definition for geriatric nurse practitioners in primary care practice? AB - Traditionally, geriatric nurse practitioners (GNPs) provide care to individual older adults and their families in a primary care practice. Although the goal is to provide high-quality, cost-effective care, GNPs may be providing ineffective care by narrowly focusing on individuals and their families. Given today's health care climate, it is essential that GNPs practice with a wider perspective. This is done by noting health issue trends among the specific older adult population that are targeted for care and planning that care with a population focus delineated by either health issues or characteristics of the older adult population. PMID- 10542575 TI - Culturally sensitive care for the elderly. AB - The opportunities for nurse practitioners to work with elderly minorities and immigrants are on the rise. Both populations are witnessing steady growth. The nurse practitioner is in a unique position to serve this population because of advanced clinical skills and traditional nursing values. When combined, these competencies make the practitioner more capable of interacting with individuals of other cultures and ages. However, skill and professional concepts still require a foundation. This article proposes that a key element in interacting with members of other cultures is understanding one's own history and sensitivities and then approaching the individual with respect. PMID- 10542576 TI - Care for life: a faculty practice with a Lifecare community. AB - A combined clinical faculty practice model is shown in the partnership that was developed between the University of Maryland, School of Nursing, Baltimore, MD, and Roland Park Place, Baltimore, MD, a continuing care retirement community. This model encourages reciprocal relationships among clinical work, research, and what is taught and learned by the students. PMID- 10542577 TI - Integration and collaboration: the advanced practice nursing role in gerontologic rehabilitation. AB - The Penn Nursing Network, the Academic Practices of the University of Pennsylvania School of Nursing, seeks to create practice settings that provide cost-effective, high-quality care and an environment for the development and testing of new models of nursing services. The Collaborative Assessment and Rehabilitation for Elders (CARE) Program is one such practice. As a Comprehensive Outpatient Rehabilitation Facility (CORF), the CARE Program offers opportunities for a nurse-managed model of care and four distinct advanced practice roles. PMID- 10542578 TI - An innovative approach to the medical management of the nursing home resident: the EverCare experience. AB - Providing quality medical care to the frail elderly who reside permanently in nursing homes is associated with a variety of problems and impediments. This article describes the solution devised by two geriatric nurse practitioners who created a program of care called EverCare. This program, now a Health Care Financing Administration (HCFA) demonstration project, incorporates the nurse practitioner in a collaborative relationship with the primary care physician to provide early detection and intervention for myriad health and medical problems found in the nursing home population and to enhance the care of the frail elderly overall. PMID- 10542579 TI - Aging in place: a proposal for rural community-based care for frail elders. AB - Rural health care is hampered by uneven development of services and multiple definitions of rurality. One of the most vulnerable populations in this setting is older residents. This article explores existing models of long-term care for the older population and proposes a model for rural community-based care. This model emphasizes the contributions of advanced practice nurses as coordinators of a collaborative system of care for the targeted population. Issues of cost and quality, as well as the strengths and limitations of the model, are addressed. PMID- 10542580 TI - Therapeutic touch--what is the controversy and why does controversy exist? PMID- 10542581 TI - Understanding and treating atopic dermatitis. AB - Atopic dermatitis is a chronic, relapsing inflammatory pruritic skin disease commonly associated with respiratory allergy. The keys to successful management of this disease continue to include accurate diagnosis, hydration of the skin, control of pruritus and infections, appropriate use of topical corticosteroids and emollients, and identification and elimination of exacerbating factors. Treatment can be complicated and educating the patient regarding disease and therapy is critical for success. PMID- 10542582 TI - Superficial fungal infections in children and adolescents. AB - Children with superficial fungal infections are commonly seen in clinical practice. Although tinea capitis and tinea corporis are the most common childhood mycoses, thrush and candida diaper dermatitis also occur frequently in infants. At times, diagnosis can be a challenge, but is made easier with the use of the potassium hydroxide microscopy and fungal cultures. Most childhood superficial fungal infections are adequately treated with topical antifungal medication. These medications are effective and the majority are safe for use in children. Oral antifungal drugs are required for children with tinea capitis, tinea unguium, and those who are immunosuppressed either from disease or therapy. Griseofulvin is the current systemic drug of choice to use in children. Several newer systemic antimycotics are currently being investigated for pediatric use. Terbinafine appears to have the best safety profile and the least risk of drug interactions. Itraconazole and fluconazole are also potential substitutes for griseofulvin in the future. The new agents, fluconazole, itraconazole, and terbinafine, have definitely improved the treatment of tinea unguium. Despite the availability of effective medications for treatment of superficial fungal infections, failure to take local and environmental measures to prevent transmission and reinfection will nullify the use of any treatment. PMID- 10542583 TI - Contact dermatitis for primary care providers. AB - Contact dermatitis is an eczematous dermatitis developing at a site where the skin has been in direct contact with the cutaneous irritant or allergen from the environment. Irritant contact dermatitis (ICD) is the most common form. Allergic contact dermatitis (ACD) is inflammation of the skin caused by an antigen that elicits a type IV hypersensitivity reaction. Clinical evaluation and judgment are a critical part of the process. Patch testing in an objective method to aid in differentiating ACD from ICD and of verifying responsible allergens. PMID- 10542584 TI - Wound healing: new understandings. AB - Understanding wound healing has moved from the cellular to the molecular level. In the clinical arena, however, care is often based on evaluation of the external manifestations of the wound, whereas treatments emanate primarily from tradition. Advances in knowledge make it necessary to understand healing at multiple levels to provide sound treatments. This article expands several molecular explanations for altered healing and suggests treatments aimed at restoring chronic wounds to a milieu for repair. PMID- 10542585 TI - Dermatologic complications in HIV. AB - The skin is unique in its accessibility for the diagnosis of diseases acquired because of decreased immunity in human immunodeficiency virus (HIV) infection. A majority of HIV-infected patients will have dermatologic problems at some time during their illness. Many of the cutaneous conditions identified will provide the means by which systemic opportunistic infections, neoplasms, and complications of therapy can be identified. The skin provides an opportunity for the recognition of new or foreign conditions as well as exotic manifestations of diseases that complicate HIV infection. A high degree of suspicion and a willingness to examine the entire skin thoroughly and frequently can enable practitioners to reduce the morbidity and mortality associated with HIV infection. PMID- 10542586 TI - Topical corticosteroids: a review of properties and principles in therapeutic use. AB - Topical corticosteroids are used to treat a variety of dermatologic conditions. Desired therapeutic endpoints can usually be achieved when an appropriate drug selection is made from the myriad topical corticosteroid agents available for use. Proper drug selection is based on the dermatosis being treated and its anticipated corticosteroid responsiveness: the anatomic site of the dermatosis, potency of the corticosteroid, appropriateness of the delivery vehicle, and the existence of particular patient factors that can effect response to therapy. Proper drug choice, coupled with good application technique, can maximize efficacy and minimize the potential of side effects experienced with topical corticosteroids. PMID- 10542588 TI - Lobbying for children's nursing issues. PMID- 10542587 TI - Common skin cancers in the United States: a practical guide for diagnosis and treatment. AB - Cutaneous malignancies are the most common cancers found in the primary care setting. It is imperative that all primary care providers become competent in evaluating skin lesions. Actinic keratoses are the most common premalignant lesions. These rough scaly plaques are the direct result of ultraviolet and other carcinogenic exposure. Actinic keratoses may be the first clinical sign to alert primary care practitioners of severe solar dermatitis and herald the development of skin cancer. Treatment is cryotherapy or topical chemotherapeutic agents such as 5-fluorouracil. Basal and squamous cell carcinomas are the most common nonmelanoma skin cancers. The primary cause is cumulative exposure to ultraviolet radiation from the sun, although other factors exist. Treatment is generally surgical excision performed by a practitioner skilled in this type of procedure contingent on tumor type, size, location, aggressiveness, and other factors. Other common treatments include electrodesiccation and curettage and cryotherapy. The incidence of malignant melanoma is the fastest rising cancer in the United States. Early detection and prevention are the mainstays of a good outcome. Depth of the lesion is the primary determinant in staging and prognosis, although other factors are also important. As the incidence of skin cancer increases, primary care practitioners play an integral role in the diagnosis, treatment, and prevention of skin cancer. The importance of early detection and appropriate referral by primary care providers will become even more crucial in the prognosis of afflicted patients. PMID- 10542590 TI - Who teaches nurses about child and adolescent mental health? PMID- 10542589 TI - Spare the rod--protect the child. PMID- 10542591 TI - Restraining children for painful procedures. PMID- 10542592 TI - Guided imagery: a review of effectiveness in the care of children. PMID- 10542593 TI - Hepatitis C: the silent killer. PMID- 10542594 TI - Food allergy in children. PMID- 10542595 TI - Nurses' perception of the value of clinical supervision. AB - Literature on clinical supervision includes few references to the experiences of those being supervised. In this qualitative study two groups of intensive care nurses were surveyed for their views on clinical supervision: 10 NICU staff who had experience of one-to-one supervision and 15 PICU staff without that experience. In addition, six NICU nurses were interviewed to elicit their understanding and perceptions of aspects of the process. A range of themes and categories emerged which identified issues for practice and management, including the value of supervision, its effect on care and requirements for its successful implementation. PMID- 10542596 TI - Promoting health: breastfeeding in PICU. PMID- 10542597 TI - Reflections on the 2nd International Congress on Child Care Nursing. PMID- 10542598 TI - Machines versus humans in clinical practice. PMID- 10542599 TI - A strategy for nursing, midwifery and health visiting: a commentary. PMID- 10542600 TI - Positive parenting: the role of the children's nurse. PMID- 10542601 TI - Oral premedication for children undergoing cardiac catheterisation. PMID- 10542602 TI - Specialist courses: education for the future. PMID- 10542603 TI - Interpreting family-centred care within neonatal nursing. PMID- 10542604 TI - How do nurses learn about family-centred care? AB - How do student nurses develop their understanding of the nebulous and ill-defined concept of family-centred care? Semi-structured interviews were used to explore how 10 child branch students described the concept and how they perceived the development of their understanding of it. According to the students, the concept incorporates interaction between the child, family and nurses to provide holistic care. Students learnt through reflection and by applying theory to practice. Although the practice they observed was varied, some of the students used more experienced nurses as role models. These findings emphasize the importance of clarifying the concept of family-centred care, of monitoring and updating the practice of supervisors in clinical areas and of encouraging reflection to help bridge the theory/practice gap. PMID- 10542605 TI - Collaborative care documentation. PMID- 10542606 TI - A re-examination of the history of children's community nursing. PMID- 10542608 TI - [Guidance is a craft]. PMID- 10542607 TI - Nutritional management of renal disease. AB - This article discusses the nutritional management of children with chronic renal failure and renal replacement therapy. It is important to remember that there is no set diet for the treatment of renal disease. Diets are prescribed for each individual child and must be reviewed regularly. PMID- 10542610 TI - [In Process Citation] PMID- 10542609 TI - [Focus on enteral nutrition. The journey to the moon was the hour of birth for enteral nutrition]. PMID- 10542611 TI - [Nurses are responsible for the instruction of family members]. PMID- 10542612 TI - [Care of a child with leukemia. Infections are particularly dreaded]. PMID- 10542613 TI - [Families of patients in intensive care: nurses help them throughout the crisis]. PMID- 10542614 TI - [Mobilization of an immobile patient: kinesthetics activates patient resources and spares the nurses]. PMID- 10542615 TI - [Use of electronic data processing in nursing: using the computer for one's own goals]. PMID- 10542616 TI - [Proof of execution: it is necessary within the framework of the nursing process]. PMID- 10542617 TI - [Foreign experience in nursing: "Leonardo da Vinci" is promoting professional education in Europe]. PMID- 10542618 TI - [Project management and the development of organization: change of topics in guidance seminars is necessary]. PMID- 10542619 TI - [Sexuality in nursing: to be able to talk about it shows professionality]. PMID- 10542620 TI - [Development of a concept of care on the unit]. PMID- 10542621 TI - [A culture of disobedience is needed]. PMID- 10542622 TI - [Model project, nursing assistant: the work satisfaction of nurses can be increased]. PMID- 10542623 TI - [Nursing conference at the Augsburg Caritas Association: "Uni on tour"--nursing science for real]. PMID- 10542624 TI - [Practical aspects of tracheotomy: care of the patient requires professional knowledge and attention to the patient]. PMID- 10542625 TI - [Aims of rehabilitation in nursing: patient activation means more than mobilization]. PMID- 10542626 TI - [Giving an intravenous injection: the Hochstetter method is the safest]. PMID- 10542628 TI - [Decision by the Federal Finance Court: home nursing service is a business]. PMID- 10542627 TI - [Teaching project in geriatric care: students are experiencing handicaps]. PMID- 10542629 TI - [Palliative care in nursing education: the promotion of personal competence stands in the foreground]. PMID- 10542631 TI - [Nursing models in practice: a common organizational image promotes the identity of the profession]. PMID- 10542630 TI - [The organizational model "primary nursing": nursing and the management of the nursing station are separated]. PMID- 10542632 TI - [Electronic data processing in nursing: a system successful in hospital practice]. PMID- 10542633 TI - [Measurements for evaluation in nursing. Thoughts on the possibilities and limits of evaluation of efficiency, using indexing and electronic data processing]. PMID- 10542634 TI - Speak freely on the Net, but think first. PMID- 10542636 TI - Critical care close-up. PMID- 10542635 TI - Ethics in action. Durable power of attorney. PMID- 10542637 TI - Another source of stem cells. AB - Blood in umbilical cords contains stem cells, which are used to help fight many diseases. Nurses are in a position to ensure that cord blood is not wasted by letting expectant parents know they can donate it or store it for future use by members of their immediate family. PMID- 10542638 TI - How to keep float nurses from sinking. AB - Floating. It's a word that makes nurses cringe. If you're a charge nurse with staffing responsibilities in acute care, you can use the strategies discussed here to help ensure that float nurses have a positive experience working on your unit--and a positive impact on patient care. PMID- 10542639 TI - Latex allergy. What you need to know. AB - Latex allergy may not be as prevalent as we think, but it's still a serious problem that affects patients and healthcare workers alike. You can minimize its effects by knowing who's at risk, how it's diagnosed, and how to reduce the amount of latex in your workplace. PMID- 10542640 TI - It matters to me. PMID- 10542641 TI - A quick way to identify compatible drugs. AB - This RN developed a way for staff nurses in her hospital to determine which commonly used drugs could be administered safely in combination with each other. Here's how she did it, and how you can create a similar brochure specific to your unit. PMID- 10542643 TI - Preventing med errors. PMID- 10542642 TI - Iatrogenic injuries. Pneumothorax. PMID- 10542644 TI - Local MSN programs make US News top rankings. PMID- 10542645 TI - Find your passion and get involved. PMID- 10542646 TI - Sticks and stones. PMID- 10542647 TI - Holistic care for children with hemophilia. PMID- 10542648 TI - Clinical research associate course available. PMID- 10542649 TI - Public health RN takes on Shaken Baby syndrome. PMID- 10542650 TI - Cervical cancer prevention. PMID- 10542651 TI - Erectile dysfunction. PMID- 10542652 TI - "Move on". PMID- 10542653 TI - Rattlesnake bites--treatment or mistreatment? PMID- 10542654 TI - Mini PICUs in the home. PMID- 10542655 TI - AACN doubts preceptor pay feasible. PMID- 10542656 TI - Preparing for the inevitable. PMID- 10542657 TI - Opening critical care to graduate nurses. PMID- 10542659 TI - Nursing students score scholarships and fellowships PMID- 10542658 TI - Sigma Theta Tau induction. PMID- 10542660 TI - The bed bath blues. PMID- 10542662 TI - So you have to give a speech. PMID- 10542661 TI - Networking for career advancement. PMID- 10542663 TI - Could you lose your license? PMID- 10542664 TI - Hospice founder inspires nurses around the world. Interview by Renee Pevour. PMID- 10542665 TI - Diving without water: hyperbaric nursing. PMID- 10542666 TI - RNs and the media: thin coverage. PMID- 10542667 TI - What's newsworthy, what's not. PMID- 10542668 TI - Six tips for taming Mike Wallace. PMID- 10542669 TI - When headlines wreak havoc on your patients. PMID- 10542671 TI - Hepatitis C uncovered. PMID- 10542670 TI - Agency nursing--too good to be true? PMID- 10542673 TI - Building a team. PMID- 10542672 TI - Therapeutic advances: new hope for patients with ALS. PMID- 10542674 TI - Nursing in the hot zone. PMID- 10542675 TI - Within reach: educational mobility for Hispanic RNs. PMID- 10542676 TI - The killer seeds. PMID- 10542677 TI - Christmas 1962: a year of firsts. PMID- 10542678 TI - Why I learned Spanish. PMID- 10542679 TI - A nurse for all seasons. Interview by Mary Jane Zusy. PMID- 10542681 TI - NEOP enables low-income students to become RNs. PMID- 10542680 TI - Interdisciplinary teams in action. PMID- 10542682 TI - Click here for research. PMID- 10542683 TI - Take the time to train. PMID- 10542684 TI - "Our house". PMID- 10542685 TI - Aviation nurses form corporation. PMID- 10542686 TI - African-American women and breast cancer. PMID- 10542687 TI - Latex allergy alert. PMID- 10542688 TI - "Gus" Hoya scores at Georgetown. PMID- 10542689 TI - Paying your preceptor? Food for thought. PMID- 10542690 TI - Shared governance and managed care. PMID- 10542691 TI - Letters of leadership: CEO, COO, and RN. PMID- 10542692 TI - Chi Eta Phi Sorority, Inc., RNs serving humanity. PMID- 10542693 TI - An eye on the federal budget. PMID- 10542694 TI - Earaches and antibiotics. PMID- 10542695 TI - The nurse/physician relationship: can it be saved? PMID- 10542696 TI - Managed care: nursing's friend or foe. PMID- 10542697 TI - Everlasting tribute to a nurse hero. PMID- 10542698 TI - Treating multiple personality patients. PMID- 10542699 TI - Emergency contraception. PMID- 10542700 TI - Nursing behind bars. PMID- 10542701 TI - Herbal treatments. PMID- 10542702 TI - What is energy work? PMID- 10542703 TI - Earning degrees by distance education. PMID- 10542705 TI - Feng Shui--a complementary therapy. PMID- 10542704 TI - Nurse, heal thyself. PMID- 10542706 TI - The business of breastfeeding. PMID- 10542707 TI - Energy fields--grounded in reality? PMID- 10542708 TI - Protocol for detecting child abuse. PMID- 10542709 TI - Cost-conscious consortium education. PMID- 10542710 TI - Vagal stimulator helps patients control seizures. PMID- 10542711 TI - Narcolepsy--a rude awakening. PMID- 10542712 TI - New JCAHO challenge: sentinel event reporting. PMID- 10542713 TI - Taking an exposure history. PMID- 10542714 TI - RSDS: difficult to Dx, difficult to Tx. PMID- 10542715 TI - Nurse substance use varies by specialty. PMID- 10542716 TI - It's time to wake up to cancer fatigue. PMID- 10542717 TI - Immune globulin therapy: new indications. PMID- 10542718 TI - The waiting game. PMID- 10542720 TI - Nurse of the year impacts the health of a nation PMID- 10542719 TI - Apheresis specialist. PMID- 10542721 TI - Five points of nursing excellence PMID- 10542723 TI - On to graduate school: step by step. PMID- 10542724 TI - The backbone of nursing--more nominees PMID- 10542722 TI - The mystery of lupus. PMID- 10542725 TI - Job outlook bright for trend-savvy student nurses. PMID- 10542726 TI - Pressure sores: the persistent problem. PMID- 10542727 TI - Are men a step higher on the ladder of success? PMID- 10542728 TI - A good nurse is never off duty. PMID- 10542730 TI - Transformational nurse executives. PMID- 10542729 TI - Rezulin pulled in the UK. PMID- 10542731 TI - Nursing with a disability. PMID- 10542732 TI - The RN of yesteryear ... the more things change PMID- 10542733 TI - Frederick school health nurses--a class act. PMID- 10542734 TI - School health nurses: sorely needed. PMID- 10542735 TI - Better transfer of records: the HARP for MED. PMID- 10542736 TI - Depression doesn't affect just adults. PMID- 10542737 TI - An elderly couple's example to live by. PMID- 10542738 TI - Drug interactions with medications and food. PMID- 10542739 TI - Germ warfare. PMID- 10542740 TI - Neonatal resource nurse--new in critical care. PMID- 10542741 TI - Breast cancer hopes. PMID- 10542742 TI - On the lookout for lice. PMID- 10542743 TI - Next generation lipids testing. PMID- 10542744 TI - Transitioning from staff to management. PMID- 10542745 TI - Sharing nursing knowledge down under. PMID- 10542746 TI - Genetics nursing: helping patients make tough choices. PMID- 10542747 TI - Anxiety disorders. PMID- 10542748 TI - Solving the puzzle of chronic pain. PMID- 10542749 TI - Making your course evaluations count. PMID- 10542750 TI - Slow codes: a time to die? PMID- 10542751 TI - Nursing art in Washington, DC. PMID- 10542752 TI - Transplant coordinator--racing the clock. PMID- 10542753 TI - Pharmacogenomics: tailoring the drug to the patient. PMID- 10542754 TI - A matter of style--communicating effectively. PMID- 10542755 TI - Cardiac invasive procedures: pre- and postprocedure care. PMID- 10542756 TI - Ecumenical institute clears path to health ministry. PMID- 10542757 TI - First days. PMID- 10542758 TI - Procedure room RN makes cardioversion easier. PMID- 10542759 TI - Advances in pediatric pain management. PMID- 10542760 TI - Suicide survivors. PMID- 10542761 TI - Fellowship opens OR suite to novices. PMID- 10542762 TI - Conscious sedation crisis: are you prepared? PMID- 10542763 TI - Seeking Athena. PMID- 10542765 TI - Breast cancer susceptibility genes. PMID- 10542764 TI - The power of culture. PMID- 10542766 TI - Build yourself up--don't beat yourself down. PMID- 10542767 TI - Nurses and politics: politically correct combination. PMID- 10542768 TI - When duty calls, military nurses answer. PMID- 10542769 TI - NIWI returns to DC in March. PMID- 10542770 TI - Testicular cancer: a young man's disease. PMID- 10542771 TI - Nursing in war--wounded healers. PMID- 10542772 TI - Clinical implications of genetic testing for inherited breast and ovarian cancer risk. PMID- 10542773 TI - Post traumatic stress disorder and Vietnam vets: an update. PMID- 10542774 TI - Turning ideas into products and profits. PMID- 10542775 TI - A parent's prayer. PMID- 10542776 TI - Chlamydia infections on the rise. PMID- 10542777 TI - International consulting in community health nursing. PMID- 10542778 TI - Dana Reeve: caregiver, activist, and superwoman. Interview by Tova Navarra. PMID- 10542779 TI - Calling all case managers. PMID- 10542780 TI - Caring for one of our own. PMID- 10542781 TI - More than a quota: the Health Education Partnerships Act of 1998. PMID- 10542782 TI - Pew Commission recommends CE and credentialing reform. PMID- 10542783 TI - Assessment and treatment of the pediatric traumatic brain injury patient. PMID- 10542785 TI - Understanding money: it's all in the process. PMID- 10542784 TI - Branding your RN-friendly organization. PMID- 10542786 TI - Moving the psychiatric patient closer to recovery. PMID- 10542787 TI - From trash to treasure. PMID- 10542788 TI - Operation Smile at CUA's School of Nursing. PMID- 10542790 TI - Becoming an RN chiropractor. PMID- 10542789 TI - Critical care enrichment serves students/staff. PMID- 10542791 TI - You are a valuable asset--here's why. PMID- 10542792 TI - Critical care nursing: past, present, and future. Interview by Catherine Campion. PMID- 10542793 TI - Pulmonary artery catheter concerns. PMID- 10542794 TI - Tooting the self-care horn. PMID- 10542795 TI - Promoting medication adherence in HIV treatment. PMID- 10542796 TI - Resolved--resources for critical care nurses. PMID- 10542797 TI - Critical care--emotion amid the equipment. PMID- 10542798 TI - Facial what? PMID- 10542799 TI - Is angioplasty safe at community hospitals without cardiac surgery? PMID- 10542800 TI - World's first live-born octuplets. PMID- 10542801 TI - Fewer RNs mean higher postop risk. PMID- 10542802 TI - Creating a mission statement for work and for life. PMID- 10542803 TI - Who says I'm depressed? PMID- 10542804 TI - ED team develops bereavement booklet. PMID- 10542805 TI - Limb lengthening attracts international attention. PMID- 10542806 TI - RN cuts cord as neighbors pop corks. PMID- 10542807 TI - Home massage for hospice patients. PMID- 10542808 TI - The midlife doctoral nurse's journey. PMID- 10542809 TI - Bring your own sutures--Peruvian mission. PMID- 10542810 TI - Forensic nursing and violent schoolboys. PMID- 10542811 TI - Worn out? You're not alone. PMID- 10542812 TI - Code Delta: water outage. PMID- 10542813 TI - Real-time documentation: hand-held computers. PMID- 10542814 TI - The Fox phenomenon: raising awareness of young Parkinson's. PMID- 10542815 TI - 1999: the countdown to Y2K compliance. PMID- 10542816 TI - Computerizing a nursing department. PMID- 10542818 TI - When drugs are in short supply. PMID- 10542817 TI - To the point: the contemporary body piercing and tattooing renaissance. PMID- 10542819 TI - St. John's Wort: latest results. PMID- 10542820 TI - Cognitive and sociodemographic risk factors for mortality in the Seattle Longitudinal Study. AB - The relationship between cognitive function and survivorship was examined in a community-dwelling sample. Survival analysis was used to examine how level and change in intellectual functioning, verbal memory, perceptual speed, and psychomotor speed were related to mortality in a sample of 601 individuals who subsequently died (decedents; n = 342 men; n = 259 women; M = 73.81 years of age) and a control group of 609 survivors (n = 296 men; n = 313 women; M = 71.96). The sample of survivors was selected to be of similar age and to have a similar level of education as the decedents. Individuals in the lowest 25th percentile of performance (crystallized abilities, visualization abilities, verbal memory, and perceptual and psychomotor speed) had a significant risk for subsequent mortality compared to individuals in the highest 25th percentile. However, after adjusting for demographic variables and psychomotor speed, only perceptual speed remained a significant risk factor for mortality. Significant 7-year declines (lowest 25th percentile) in measurements of Verbal Meaning, Spatial Ability, Reasoning Ability, and Psychomotor Speed were risk factors for subsequent mortality relative to those who had the least amount of decline. The relationship between mortality and cognitive function tended to be a specific rather than a pervasive phenomenon, even after adjusting for sociodemographic factors and psychomotor speed. Decrease in cognitive performance tended to be a better predictor of subsequent mortality than was the level of cognitive performance. PMID- 10542821 TI - Mode of physical activity and self-efficacy in older adults: a latent growth curve analysis. AB - A randomized controlled trial examined the effect of two physical activity modes on changes in self-efficacy over the course of a 12-month period in older, formerly sedentary adults (N = 174, M age = 65.5 years). Participants were randomized into either an aerobic activity group or a stretching and toning group. Structural equation modeling was employed to conduct multiple sample latent growth curve analyses of individual growth in exercise and physical self efficacy over time. Results revealed a curvilinear growth pattern for both types of efficacy with increases occurring over the first 6 months followed by declines at the 6-month follow-up. There was a significant treatment by mean level growth interaction for exercise efficacy with both groups increasing over time, but the aerobic group evidenced a twofold increase in growth over the stretching group. Structural analyses indicated that frequency of exercise participation was a significant predictor of overall growth in efficacy, and improvements in fitness were only related to exercise efficacy growth in the stretching group. Findings are discussed in terms of social cognitive theory and further application of latent growth curve modeling to studies of physical activity effects in older adults. PMID- 10542822 TI - Neuropsychological correlates of self-reported performance in instrumental activities of daily living and prediction of dementia. AB - This study examines the relationships between performance on neuropsychological tests and 4 instrumental activities of daily living (IADL) associated with an increased risk of dementia, in 1,792 nondemented elderly people included in the Personnes Agees Quid (PAQUID) study. There was a decline of neuropsychological performance on each test with increasing IADL dependency, in particular between Grades 1 and 2. Performances of the independent participants were very homogeneous. Each IADL had different specific associations with the neuropsychological tests. A principal component analysis showed three main factors explaining 49.6% of variance. The first factor was the common cognitive component of both instrumental tasks and neuropsychological tests. The second factor was specific to three IADL, whereas use of transportation had its main loading on the third factor. Only the first factor was predictive of the risk of incident dementia, suggesting that the predictive value of the 4 IADL was explained by their cognitive component. PMID- 10542823 TI - Psychosocial consequences of age-related visual impairment: comparison with mobility-impaired older adults and long-term outcome. AB - Indices of behavioral competence (activities of daily living [ADLs], instrumental activities of daily living [IADLs], use of outdoor resources, leisure activity level) as well as emotional adaptation (subjective well-being, future orientation) were used to investigate the psychosocial consequences of age related vision impairment in a threefold manner: (a) comparison of visually impaired and unimpaired elders, (b) comparison of visually impaired and mobility impaired elders, and (c) long-term adaptation across 5 years. The research design used (a) 42 severely visually impaired elders, (b) 42 blind elders, (c) 42 mobility-impaired elders, and (d) 42 unimpaired elders. Compared with the mobility impaired, the visually impaired demonstrated lower IADL competence but no difference in emotional adaptation. The long-term adjustment of the visually impaired remained relatively stable in the behavioral domain, although lower compared with the unimpaired elders. Emotional adaptation decreased over the 5 year longitudinal interval in the visually impaired and the unimpaired group, but the decrease was generally higher in the visually impaired group. Conceptual ideas from environmental gerontology as well as psychological resilience are used to interpret these results. PMID- 10542824 TI - One voice too many: adult age differences in language processing with different types of distracting sounds. AB - An experiment is reported that investigated factors that might contribute to age differences in the ability to process spoken language under conditions of competition from various types of background noise. Age differences in recall of spoken sentences were shown to depend on the type of background noise as well as its intensity. Increased intensity levels of just one competing speaker produced differentially greater impairment in older adults than in young adults. Analyses showed that listening performance was predicted not only by individual differences in hearing ability but also by speed of processing, which underscores the combined role of age-related auditory and cognitive changes in processing spoken language. PMID- 10542825 TI - Clock drawing is sensitive to executive control: a comparison of six methods. AB - We examined six clock-drawing task (CDT) scoring systems relative to the Executive Interview (EXIT25, a measure of Executive Control Function [ECF]) and the Mini-Mental State Exam (MMSE). Subjects included n = 33 National Institute of Neurological, Communicative Disorders, and Stroke "probable" Alzheimer's disease (AD) cases and n = 52 independent living controls. AD cases and controls differed on the EXIT25, MMSE, and all CDTs. All CDTs were significantly correlated with the EXIT25 (ranging from r = .56 to r = .78). These associations generally persisted after adjusting for Age, Education, and MMSE scores. In backwards stepwise linear multivariate regression models, only CLOX: An Executive Clock Drawing Task scores contribute significantly to EXIT25 scores (R2 = .68) and MMSE scores (R2 = .72). Clock drawing draws upon both executive and general cognitive resources. CLOX explains incrementally more variance in ECF than other CDTs. PMID- 10542826 TI - Variation in the impact of social network characteristics on physical functioning in elderly persons: MacArthur Studies of Successful Aging. AB - OBJECTIVES: Social support and social networks have been shown to exert significant effects on health and functioning among elderly persons. Although theorists have speculated that the strength of these effects may differ as a function of sociodemographic characteristics and prior health status, few studies have directly tested the moderating effects of these variables. METHODS: Longitudinal data from the MacArthur Study of Successful Aging were used to examine the effects of structural and functional social support on changes in physical functioning over a 7-year period, measured by the Nagi scale, in a sample of initially high-functioning men and women aged 70 to 79 years. Multiple regression analyses were used to test the main effects of social support and social network variables, as well as their interactions with gender, income, and baseline physical performance. RESULTS: After controlling for potential confounding effects, respondents with more social ties showed less functional decline. The beneficial effects of social ties were stronger for respondents who were male or had lower levels of baseline physical performance. DISCUSSION: The effects of social support and social networks may vary according to the individual's gender and baseline physical capabilities. Studies of functional decline among elderly persons should not ignore this population variation in the effects of social networks. PMID- 10542827 TI - Work transitions and health in later life. AB - OBJECTIVES: The goal of the analysis was to examine the association between health status and work behavior among men aged 55-69. We specifically examined the conditions under which health is most strongly associated with labor force exit and reentry. METHODS: The association between health and labor force transitions was examined using logistic regression analyses, based on data from the 1984 and 1985 panels of the Survey of Income and Program Participation. RESULTS: We found that for men aged 55-69 in the mid-1980s, poor health was positively associated with labor force exit, and negatively associated with returns to work. Although these main effects are very strong, we found that health was particularly important among individuals for whom retirement was least attractive. Health had its most substantial association with work transitions among men with working wives, as well as among men who were younger, or who had limited nonwork financial resources. Health also had a particularly strong association with work transitions among Black men, but only with reference to reentry decisions. DISCUSSION: Our results suggest that continued work may have limited appeal for men who are prepared for retirement, even when they are in excellent health. PMID- 10542828 TI - Association between executive attention and physical functional performance in community-dwelling older women. AB - OBJECTIVES: Executive functions supervise the contents of working memory, where information from long-term memory is integrated with information in the immediate present. This study examined whether executive attentional abilities were uniquely associated with the performance of complex, instrumental activities of daily living (IADLs) in cognitively intact and physically high-functioning older women. METHODS: Participants were 406 community-residing, older women aged 70-80 years in the Women's Health and Aging Study (WHAS) II, screened to be physically high functioning and cognitively intact using the Mini-Mental State Exam. Hierarchical regression models, adjusted for demographic and disease variables, were used to evaluate the association of cognitive domains, including executive attention, memory, psychomotor speed, and spatial ability with summary measures of IADL (e.g., looking up and dialing a telephone number) and mobility-based ADL (e.g., walking 4 meters) function. RESULTS: Tests of executive attention were associated with performance on IADLs (6.6%) and, to a lesser degree, mobility based ADLs (1%), adjusting for demographic and disease variables. In particular, the mental flexibility component of the Trail Making Test accounted for the majority of attentional variance in IADL performance. Older age, lower education, and African American race were also associated with poorer physical test performances. DISCUSSION: Executive difficulties in flexibly planning and initiating a course of action were selectively associated with slower performance of higher-order IADL tests, relative to other domains of cognition, in a high functioning, community-based older cohort. These results suggest that executive functions may be important in mediating the onset and progression of physical functional declines. PMID- 10542829 TI - Time use of old and very old Berliners: productive and consumptive activities as functions of resources. AB - OBJECTIVES: The aims of this study were to examine time use of elderly women and men and to explain age- and gender-related variance in activity levels in terms of differences in available resources. METHODS: Activities reported in an elderly sample stratified for age and gender (N = 485, Age: 70-103 years) were aggregated into three classes: regenerative, productive, and consumptive activities and regressed on income, presence of a partner in the household, education, walking mobility, and labor force participation. RESULTS: Levels in productive and consumptive activities were decreased in the old-old. Moreover, young-old women spent more time working in the household than any other group. Most of the age- and gender-related variance in activity levels could be explained by differences in available resources. For men, living with a partner was associated with less time spent for productive activities and enhanced leisure time, whereas for women, the reverse was observed. The expected reduction of unpaid work time as a function of income (income effect) was not observed. DISCUSSION: Elderly individuals do contribute to societal production. Allocation of time to productive and consumptive activities is a function of available resources, amounts and effects of which differ for young-old and old-old as well as for women and men. PMID- 10542830 TI - How older people in the United States and Germany fared in the growth years of the 1980s: a cross-sectional versus a longitudinal view. AB - OBJECTIVES: The goal of the study was to show that cross-sectional and longitudinal data yield dramatically different answers to a basic question: "How did older persons fare in the recovery years of the 1980s?" METHODS: The United States Panel Study of Income Dynamics and the German Socio-Economic Panel are used cross-sectionally to capture changes in the economic well-being of older persons in the trough and peak years of the 1980s business cycle, and longitudinally to trace how the economic well-being of a given cohort of older persons changed over those years. Kernel density estimation is then used to show how the distribution of economic well-being of these populations changed over these years. RESULTS: Cross-sectional comparisons confirm that persons aged 65 and over in the peak year were better off than persons aged 65 and over in the trough year in both countries. Longitudinal comparisons, however, show that persons aged 65 and over in the trough year who survived to the peak year received a substantially smaller share of the rewards of economic recovery than cross-sectional comparisons imply. Moreover, the entire income distribution of older persons in the United States shifted downwards. DISCUSSION: Compositional changes in the cross-sectional data, caused by the entry of high-income persons who are young in the peak year but old in the trough year, obscure the decline in the economic well-being of the cohort of older persons who survived the trough year, in cross-sectional comparisons of older populations in the United States in the 1980s. PMID- 10542831 TI - A comparison of correlates of cognitive functioning in older persons in Taiwan and the United States. AB - OBJECTIVES: This article compares patterns of association between cognitive functioning and a number of sociodemographic and health correlates among older persons in Taiwan and the United States. METHODS: The study uses data from the 1993 Survey of Health and Living Status of the Elderly in Taiwan and the 1993 Study of Asset and Health Dynamics Among the Oldest Old in the United States. Separate multivariate regression models are employed for each country to examine the effects of sociodemographic and health factors on cognitive functioning, and to examine the marginal impact of cognitive functioning on activities of daily living (ADL) and instrumental ADL (IADL) functioning. RESULTS: Results of the multivariate analyses show similar patterns of association across the two countries and replicate findings from previous studies. Increasing age, female gender, lower education, depression, and selected health conditions are associated with lower cognitive functioning. In addition, although a significant predictor of both ADL and IADL impairments, cognitive functioning is more powerful with respect to explaining IADL impairments. DISCUSSION: Study findings suggest that the cognitive measures are capturing similar dimensions in Taiwan and the United States, and that factors associated with cognitive functioning and its consequences with respect to physical functioning are similar in the two countries. PMID- 10542833 TI - After Littleton. PMID- 10542832 TI - Stability and change in older adults' social contact and social support: the Cardiovascular Health Study. AB - OBJECTIVES: The aim of this study was to examine the degree of individual change in structural indicators of social support (family network contact and close friend network contact) and functional indicators of social support (belonging, appraisal, and tangible support) during late life. METHODS: Using a large population-based sample of older adults, hierarchical linear modeling was applied to examine the extent of change in social contact and support as well as sociodemographic characteristics (age, race, gender, and education) that might explain individual variability in contact and support at baseline and over time. RESULTS: Consistent with predictions, small yet significant increases were observed in belonging support and tangible support. Contrary to predictions, no evidence was found for significant individual change in family network contact, close friend network contact, or appraisal support. Sociodemographic characteristics were more consistent predictors of variability in contact and support at baseline than variability over time. DISCUSSION: The findings of this study add to a growing literature suggesting that late life is not typically characterized by a decline in important social resources. PMID- 10542834 TI - Solving the microgram/kilogram puzzle. PMID- 10542835 TI - Preserving patient privacy. PMID- 10542836 TI - Aiding nurses with AIDS. PMID- 10542837 TI - Hypoglycemia treatment. PMID- 10542838 TI - Trouble with bubbles. PMID- 10542839 TI - Staking a claim. PMID- 10542841 TI - Talking with physicians about pain. PMID- 10542842 TI - APNs in home care. PMID- 10542843 TI - Tax debate. Key health care programs in danger. PMID- 10542844 TI - Restraint-free care: is it possible? PMID- 10542845 TI - Emergency! Pericarditis. PMID- 10542846 TI - Nurses as leaders. Then & now. PMID- 10542847 TI - What is homocysteine? PMID- 10542848 TI - Clinical snapshot. Fibromyalgia. PMID- 10542849 TI - Endothelial dysfunction and the promise of ACE inhibitors. PMID- 10542850 TI - Gamma knife radiosurgery. PMID- 10542851 TI - Continued competence: assuring quality health care. PMID- 10542852 TI - Opioid conversion guidelines for managing adult cancer pain. PMID- 10542853 TI - When all else fails, try harm reduction. PMID- 10542854 TI - Working with UAPs. PMID- 10542855 TI - How to practice 'activist geriatrics'. PMID- 10542856 TI - Geriatrics photo quiz. Chagas disease. PMID- 10542857 TI - Enlarged papules on the foot. PMID- 10542858 TI - Urinary incontinence: keys to diagnosis of the older woman.1. AB - Up to 38% of women age 65 and older experience urinary incontinence to some degree, although the prevalence in this population may be even greater. Aging, childbirth, and hormonal changes affect the muscle and tissue support of the urethra and bladder, decreasing their ability to hold urine. Neurologic injury related to disease, trauma, or surgery may impair the pathways between the brain and bladder, leading to inappropriate urgency and frequency. Older women often do not talk about their incontinence, because they are embarrassed or believe there is no cure. Therefore, it's important for clinicians to ask about the involuntary loss of urine. A careful history alone can often reveal 80 to 90% of the diagnosis. PMID- 10542859 TI - Alternative medicine: what the data say about common herbal therapies. AB - An increasing number of Americans are turning to complementary and alternative medicine to help manage or prevent the onset of chronic disease, improve cognitive function, boost overall general well-being, and increase longevity. Some of the more widely-used herbal preparations designed to help accomplish these objectives include St. John's wort, ginkgo biloba, echinacea, garlic, and ginger. In general, the clinical trial data on these preparations is in the embryonic stages, whereas the popularity of these compounds is fueled in part by anecdotal evidence. Given the embrace by Americans--especially older persons--of these alternative remedies, knowledge of their uses and potential side effects can help the primary care physician better collaborate on a course of care that makes effective use of the best treatments, both traditional and alternative. PMID- 10542860 TI - Parkinson's disease: therapeutic choices and timing decisions in patient management. Interview by Wayne Kuznar. AB - Parkinson's disease is a progressive neurodegenerative disorder characterized by striatal dopaminergic loss. Carbidopa/levodopa is the most effective drug treatment for disease management. It reduces bradykinesia and rigidity, but is less effective against tremor. Whether carbidopa/levodopa should be used at the time of initial diagnosis or delayed until symptoms become disabling is controversial. A clinical trial is in progress to help resolve this dilemma. As carbidopa/levodopa loses efficacy with continued use, adjunct therapies using catechol-O-methyl-transferase inhibitors or dopamine agonists may be considered. In younger patients exhibiting parkinsonian symptoms, dopamine agonists may be used as first-line therapy. A new, reversible surgical intervention known as deep brain stimulator placement is being used to control disabling tremor in patients not responding to optimal drug therapy. PMID- 10542861 TI - Viscosupplementation: treatment alternative for osteoarthritis of the knee. PMID- 10542862 TI - [State of the progeny of the 1st generation conceived at different times after low-dose irradiation of female rats]. AB - The relation between the antenatal and early postnatal ontogenesis of the progeny and the degree of maturity of oocytes at the time of the whole single gamma irradiation with doses of 0.25, 0.5 and 1 Gy was studied on 473 first pregnant Wistar rats and 1402 rats of the first generation. Ontogenesis disorders ot the progeny of these females after irradiation of the matured oocytes were more marked than after irradiation of the maturing oocytes. The distinction was noticeable after a dose of 0.25 Gy. PMID- 10542863 TI - The effect of essentiale on histones and nucleic acids in liver and blood-forming tissues of rats irradiated with gamma-rays. AB - In this paper, the influence of the hepatoprotective drug Essentiale on the target (normal and regenerating liver) and the non-target (spleen and bone marrow) rat tissues was studied after whole body irradiation with the dose of 5.7 Gy gamma-radiation. The application of the drug 24 h before irradiation alleviated all the radiation induced changes of the histones and nucleic acids in the normal and regenerating liver tissue. In the non-target tissues only mild radioprotective effect was observed. The application of the preparation 30 min after irradiation was less effective than the application before irradiation. The repeated application of Essentiale after irradiation did not increase the beneficial effect of the previous preparation application. PMID- 10542864 TI - [Functional interactions of components of the adenylate cyclase system in the rat liver after prenatal exposure to gamma irradiation]. AB - The effects of long-term prenatal gamma-radiation (0.5 Gy) on the glucagon signalling through adenylyl cyclase in adult rat liver have been investigated. The coupling of receptor/Gs-protein/adenylyl cyclase was tested using kinetic constants for stimulation of adenylyl cyclase by GTP, Gp(NH)pp, AlF4- and glucagon. The estimated data suggests that prenatal chronic gamma-radiation prompted (i) a decrease in rate of GTP hydrolysis on Gs-protein; (ii) a reduction of glucagon potency to accelerate the exchange GDP for GTP on Gs-protein. PMID- 10542865 TI - [Effects of different doses of x-ray irradiation on the contents of microsomal hemoproteins in the rat liver]. AB - The content of cytochromes P-450 and b5 in microsomal fraction of rat liver was studied at 1, 3, 6, 12 and 24 hours after X-irradiation with doses of 4, 8 and 12 Gy. It was found that post-irradiation changes in the cytochromes content were already observed in the first hours after X-irradiation independently of a dose of ionizing radiation. PMID- 10542866 TI - [Dependence of the cytogenetic adaptive response in the rat bone marrow cells on the dose of chronic gamma irradiation in vivo]. AB - The induction of chromosome aberrations in bone marrow cells of rats exposed to chronic gamma-irradiation and subsequently to acute gamma-irradiation was studied. Adult male rats were exposed ot 3-40 cGy (2.9 cGy/day) of chronic gamma irradiation (adaptive dose) and subsequently to 4 or 6 Gy of gamma-rays (47 cGy/min, challenge dose). The yield of chromosome aberrations in marrow cells induced by adaptive and challenge dose was lower than the sum of the yields separately induced by chronic and acute gamma-irradiation. The most effective dose for induction of the adaptive response was 0.4 Gy. PMID- 10542867 TI - [Separate and combined mutagenic effect of radiation and asbestos on the micronucleus test in experiments]. AB - We investigated the isolated and combined mutagenic effect (ME) of radiation (gamma-irradiation, 0.5 or 2 Gy) and asbestos (i.p. 10 mg/mice) in mice CBA. We studied also the antioxidant activity and malonic dialdehyde concentration in blood serum as possible mechanism of ME and its possible modification. For the ME the micro-nuclei incidence in polychromatic bone marrow erythrocytes was scored. The reciprocal modification (potentiation) of both radiation and asbestos ME was established for combination "radiation, 2 Gy + asbestos", the additivity--for combination "radiation, 0.5 Gy + asbestos". PMID- 10542869 TI - [Late radiation effects in the microcirculation system of the mouse brain after chronic low-dose irradiation]. AB - The radiation reaction of the mouse brain microvascular system was investigated 15 months after chronic (dose-rate was 3 cGy/day) and acute (dose-rate 3.2 Gy/min) gamma-irradiation. Late pathological changes in the microvascular system after the acute irradiation (3 Gy) were not found. In the same time, after chronic irradiation with equivalent dose (the accumulated dose was 3 Gy) the depletion of endothelial cell population, and multiple foci of minor pathological changes of endotheliocyte ultrastructure at as many as 50% of the population were revealed. It may be more likely to lead to reduction of tolerance of brain microvascular system to an acute radiation (10 Gy) in the late period after chronic exposure. The mechanism of impairment of brain microcirculation after low chronic irradiation will have to understand. PMID- 10542868 TI - [Effects of halogen derivative hydrocarbons on radiation damage of biological membranes]. AB - Sharp increase of radiation-induced damages of isolated erythrocyte membrane lipids and proteins upon irradiation in presence of some halogen derivative hydrocarbon are shown. On chloroform and CCl4 examples it is shown that marked effects displayed under relatively low (submillimoles) concentrations and radiation doses that induced only insignificant oxidative membrane damages in absence of modifiers. The data are received which evidenced in favour of decisive role in these processes of halogenated organic radicals forming under one electron reduction of halogen derivative hydrocarbons by hydrated electrons. PMID- 10542870 TI - [Assessment of genotoxicity and late effects of radiation and chemical exposures]. AB - The 3-month-old female rats received with drinking water PbCl2 or Hg2(NO3)2 in concentrations 100 or 1 mkg/l in account to metal, respectively, during 1 month before and 1 month after single gamma-irradiation with 25 or 50 cGy doses. The features of action of lead- and mercury-ions revealed in combination with low doses radiation. In case of the mercury ions introduction the relationships had been shown between early postradiation biochemical effects (changes of blood leucocyte DNA indexes) and blood granulocyte changes in 1 month after irradiation, and correlation of postradiation repair results with life span shortening as well. PMID- 10542871 TI - [Effects of chronic radiation exposure on erythrocyte metabolism and functioning]. AB - The structural and functional state of blood erythrocytes in people exposed to radiation during Chernobyl NPP accident was studied two or six years later. The changes in oxygen transport system of erythrocytes and in the ratio between some organic phosphate fractions were observed. The beta-carotene per os administration to rats during low-level irradiation increased hemolytic stability and activity of glycolytic enzymes, that bears witness to the increasing of organism adaptive reactions at provitamin administration during irradiation. PMID- 10542872 TI - [Radiation damage of the thyroid gland as a probable cause of increased incidence of non-Hodgkin lymphoma and other hematologic diseases]. AB - Th distribution of autoimmune thyroiditis in the patients with diseases of blood system was investigate. The attribute of autoimmune thyroiditis was revealed by the detection of antimicrosomal antibodies. It was established that the autoimmune thyroiditis are more often in patients with various hematological diseases than in control group. It is supposed that the increase in frequency of some hematological diseases in residents suffered from the Chernobyl accident can be defined not only by the influence of the radiation on blood system, but also can be connected with damage to thyroid glands. PMID- 10542873 TI - [Effects of emotional stress on postradiation regeneration of the hematopoietic system in rats under protective action of indralin]. AB - The rat experiments were designed to study the effect of the emotional stress, developed before or after gamma-irradiation with a dose of 6.0 Gy (LD40-50/30), on the radioprotective effectiveness of indralin. It was shown that the emotional stress, developed before irradiation, did not change the radioprotactive effect of indralin on hemopoietic system. In contrast with this, emotional stress, developed after irradiation, inhibited the radioprotactive effect of preparation on the medullary hemopoiesis. PMID- 10542874 TI - [Stress in the etiology of radiation injury. Role of regulatory mechanisms]. AB - Radiation damage as stress is regarded in the review. The place of lipid peroxidation in the starting of stress reaction and the possibility of the application of antioxidants and thymus preparations as factors which lower the hardness of radiation damage are considered. PMID- 10542875 TI - [Radiation safety of the population and agricultural production industry (standardization of the radionuclide content of agricultural products)]. AB - The topical problems of standardization of the radionuclide content in food products and radiation control in agricultural production have been considered. The fundamental principles for establishing the permissible radionuclide concentrations in foodstuffs and acceptable contamination levels of agricultural land and products were justified. PMID- 10542876 TI - [DNA-protein cross-links in leukocytes of the field mouse (Apodemus agrarius) and the bank vole (Clethrionomys glareolus) dwelling in the territory of a radioactive waste storage station]. AB - DNA-protein cross-links in white blood cells of Apodemus agrarius and Clethrionomys glareolus dwelling in the territory of the guarded zone of the radioactive waste storage in Sergievo Posad (area with gamma-background 50-400 microR/h) have been determined. As a control the animals from sanitary protected area of Sergievo Posadsky department (gamma-background on the soil surface is 8 10 microR/h) was used. It was found that the doses received by Apodemus agrarius and Clethrionomys glareolus, resulted in increasing of the DNA-protein cross-link level in the Apodemus agrarius white blood cells by a factor of 1.8, and in Clethrionomys glareolus by a factor of 1.4, in comparison with control. The increase of DNA-protein cross-link number reliably correlates with the beta emitting nuclide content in animal bodies. The changes in the number white of cells and in the white blood composition were not observed. PMID- 10542877 TI - [Distribution of Cs-137 in bodies of inhabitants of the Vitebsk Region]. AB - The content of 137Cs in the whole human body and certain internal organs has been determined using gamma-spectrometry method for inhabitants of Vitebsk Region. The average activity of 137Cs in the whole human body was obtained to be 0.038 mkCu. The unevenness of radiocesium accumulation in the investigated internal organs has been noted. The average annual doses from 137Cs for some human organs have been calculated. PMID- 10542878 TI - [Effects of natural zeolite-clinoptilollite on processes of removal of Cs-137 from the rat body]. AB - The selectivity of natural and synthetic zeolites to 137Cs in experiments in vitro has been investigated. The influence of natural zeolite-clinoptilolite on the dynamics of withdrawal of 137Cs from rats' organism was estimated. PMID- 10542880 TI - [Surgical treatment of malignant ovarian tumors]. PMID- 10542879 TI - Treatment of gynecologic cancer: the US experience. PMID- 10542881 TI - [Role of surgical staging in epithelial ovarian tumors of initial stages]. PMID- 10542882 TI - [First-line chemotherapy in advanced ovarian cancer]. PMID- 10542883 TI - [Promising new drugs in the treatment of ovarian cancer, and of gynecologic tumors, in general]. PMID- 10542884 TI - [Pharmacologic basis of the effectiveness of topotecan in the treatment of ovarian carcinoma]. PMID- 10542885 TI - [BRCA genes and ovarian neoplasms]. PMID- 10542886 TI - [Prognostic significance of folate binding protein in ovarian neoplasms]. PMID- 10542887 TI - [Ovarian carcinoma: pilot study of hyperthermic intraoperative peritoneal perfusion of antiblastic agents]. PMID- 10542888 TI - [Locoregional chemotherapy with hypoxic perfusion in residual ovarian cancer]. PMID- 10542889 TI - [Intraoperative chemotherapy with mitoxantrone and cisplatin after extensive debulking in the treatment of stage III and IV epithelial neoplasms of the ovaries]. PMID- 10542890 TI - [Statistical considerations in clinical trials with particular reference to phase III studies]. PMID- 10542891 TI - [Hormone replacement therapy in patients with ovarian cancer]. PMID- 10542892 TI - [Intensive care medicine and intensive care medicine research in Germany- difference between appearance and reality]. PMID- 10542893 TI - [Piracetam in anesthesia for prevention of postoperative delirium]. AB - Delirium is a serious postoperative complication after general anaesthesia. The incidence is estimated to be about 10-15%. In the case of additional risk factors, such as old age, previously existing cerebral-vasculous and psychic deficiencies, and anaesthesia of long duration the incidence is much higher. Delirium impairs postoperative mobilisation and convalescence of the patient, can lead to a longer hospital stay and is associated with higher mortality rates. In a series of clinical studies it could be shown that the perioperative administration of the nootropic piracetam led to a shorter recovery period after anaesthesia and had a favourable influence on the delirious symptoms. Especially patients with risk factors for postoperative delirium profited from prophylactic application of piracetam. The success of this medication can be explained by a protective influence of the substance on central neurons against hypoxia, ischemia and intoxication, all of which are discussed as possible causes for postoperative delirium. Next to the pathogenetic mechanisms of postoperative delirium, the mode of action of piracetam is shown in a review and a summary of references on the clinical use in anaesthesia is given. PMID- 10542894 TI - [Meta-analysis of controlled randomized studies on droperidol for prevention of postoperative phase vomiting and nausea]. AB - OBJECTIVE: Randomised, controlled trials using prophylactic droperidol to prevent postoperative nausea and vomiting (PONV) were included in a meta-analysis to estimate efficiency and dose-response of treatment. MATERIALS AND METHODS: Studies were systematically extracted from Medline, the manufacturer's database, and a supplemental search of references lists and current issues of locally available anaesthesia journals. Complete prevention of PONV defined as absence of nausea, retching, and vomiting within 6 hours (early PONV) and within 48 hours (late PONV) was chosen as the main end point. Additional information such as dose of droperidol, time and way of administration, and biometric data of the patients were extracted from each study. The pooled relative risk and the number-needed-to treat (NNT) were calculated. RESULTS: A total of 72 studies with 107 comparative subgroups were accepted for analysis according to the prospectively defined criteria. Of these sixty-nine trials reported a lower incidence of PONV with droperidol. The incidence of early and late PONV among the 5370 patients receiving droperidol was 23.4% and 38.2%, respectively. The corresponding incidence among the 3954 control patients was 40.7% and 53.9%. The relative risk for patients receiving prophylactic droperidol of suffering from early PONV was 0.58 and 0.71 for late PONV. The NNT for preventing one patient from PONV was 5.8 and 6.4 for early and late PONV, respectively. Treatment with droperidol was more effective when the baseline risk for PONV was higher than 25% for early PONV and 35% for late PONV. Under these circumstances the NNT was between 2.6 and 5.6. There was no dose response relationship for droperidol when the drug was applied in doses ranging from 0.5 to 300 micrograms.kg-1 body weight. It was not possible to derive reliable information about the incidence of side-effects of the droperidol administration. CONCLUSION: Droperidol is an effective antiemetic drug. The drug can be administered to patients with an increased risk of suffering from PONV without antiemetic prophylaxis. Since a positive dose response is lacking, droperidol should only be administered in doses of 1 mg or less. PMID- 10542895 TI - [The accumulation of different substituted hydroxyethyl starches (HES) following repeated infusions in healthy volunteers]. AB - AIM OF THE STUDY: Accumulation of hydroxyethyl starch (HES) after repeated applications of starches with different molar substitution and similar molecular weight was investigated. METHODS: Treatment with five consecutive infusions of hydroxyethyl starch was carried out using two medium molecular weight hydroxyethyl starches with a molar substitution of 0.5 and 0.62. Healthy volunteers received 500 ml 6% HES 200/0.62 (30 g) or 500 ml 10% HES 200/0.5 per day over a period of five consecutive days. Blood samples were taken in the morning before infusions were started (7.A.M.) and at each hour during the infusion period of 4 hours post infusionem until 4 hours after the infusions. During the first 10 days and on the 20th and on the 30th day after the last infusion blood samples were taken. RESULTS: Both HES solutions were subjectively and objectively well tolerated by healthy volunteers. No side effects were observed. However, pharmacokinetics of the investigated HES-formulations were significantly different. The model-independent calculated elimination half life time (T1/2) increased from day to day. During the five days T1/2 was prolonged for 20 h by high substituted HES (200/0.62) and for 2.5 h for medium substituted HES (200/0.5). The half life times related to the three compartment model calculation were with 0.6 h, 11.6 h and 211 h for HES 200/0.62 two fold higher than the times for HES 200/0.5 with 0.39 h, 6.98 h and 113 h. Plasma clearance for HES 200/0.5 (4.86 ml/min) was five fold higher than that from HES 200/0.62 with 0.98 ml/min. With the exception of the first day of infusion serum concentrations of HES 200/0.5 although only 30 g HES 200/0.6 versus 50 g HES 200/0.5 were infused. No difference of the hemodilution effects between the two HES-formulations were observed. The hemorheologic parameters were similar in both groups with the exception of plasma viscosity which was significantly higher after infusion of HES 200/0.62. CONCLUSION: High substituted HES accumulate in serum more than medium substituted HES. Especially when HES must be applied in multiple doses, high substituted HES should not be used or the infusion interval must be adapted to the elimination half life time of the used HES. PMID- 10542896 TI - [Intensive care research in Germany--an analysis of papers in important international journals]. AB - OBJECTIVE: The present study was designed to assess who is responsible for publications in important english-written intensive care journals. METHODS: 10 english-written journals that exclusively or mainly publish intensive care papers were analysed over the past 10 years (from 1988 to 1997). All German universities were included and publications were specified according to the speciality (e.g. anesthesiology, internal medicine, neurosurgery) and the university of origin of the publication. Letters, abstracts, editorials, meeting-reports, and news were not included in the analysis. RESULTS: A total of 804 publications was found in the analysed journals. 343 publications were originated from interne medicine departments (433 of all publications), 207 were from anesthesiology departments (26%). Papers from surgical departments were only rarely found (cardiac surgery: 14 papers; neurosurgery: 12 papers; general/trauma surgery: 34 papers). 150 publications originated from neurology departments, and only 44 pediatric intensive care papers were found. A wide range of contributions from the specific universities were seen (ranging from 0 to 43 papers). Universities from former East Germany published very rarely in international intensive care journals (3%). CONCLUSION: Only few German universities have published in important intensive care journals over the past 10 years. Some interne medicine and some anesthesia departments were markedly engaged in this speciality, whereas several other universities and most universities from former East Germany did not participate significantly in the international intensive care literature. PMID- 10542897 TI - [Anesthesia in patients with cardiovascular diseases. 1. General and specific aspects of coronary heart disease and arterial hypertension]. PMID- 10542898 TI - [Normalization of the vascular picture with sympathetic block in severe arterial ischemia from ergotism]. AB - This case report presents a 44-year-old woman with severe arterial ischemia leading to claudicatio and acute pain in rest caused by an ergotism. In the history was an abuse of suppositories containing caffeine and ergotamine induced by chronic headache. The initial angiography showed occlusions of the femoral arteries. After excluding other vascular diseases, intraarterial infusions of prostaglandin E were administered. Additionally, physiotherapeutic treatment followed. An progrediency of the symptoms made a epidural catheter for sympathicolysis and treatment of the acute pain necessary. As the results of this intervention were encouraging, a sympathetic blockade with injection of 96% ethanol at the level of L 2/3 and 3/4 was performed. After treatment, the clinical symptoms and the blood flow measured by Doppler ultrasonography normalised. A final angiography demonstrated a now normal arterial status. Ergotism, indication and methods of sympathetic blockades are discussed. PMID- 10542900 TI - Trauma anesthesia at the verge of the third millennium. Epidemiologic, socioeconomic and medical aspects. PMID- 10542899 TI - [Rett's syndrome: pathophysiology and anesthesiological implications]. AB - Rett's syndrome is a neurodevelopmental disorder which is caused by a mutation on the x-chromosome; thus, it only affects the female sex. After seemingly normal postnatal development affected girls lose already acquired mental, motoric and social skills. The last stage of the syndrome is characterized by microcephaly, severe mental retardation, spastic paraparesis, epilepsia, respiratory dysrhythmia, neurogenic scoliosis, abnormal joint alignment and muscle contractures. Rett's syndrome is probably the leading cause for progressive mental retardation in girls, but still it is relatively unknown. This paper describes Rett syndrome and its pathophysiology. The following case report discusses special anesthesiological implications due to the immature cardiorespiratory system and describes a coagulation disorder following treatment with valproic acid. PMID- 10542901 TI - [The polytrauma patient, triage and priorities in early treatment]. PMID- 10542902 TI - [Skull-brain injury in polytrauma--results of neurosurgery]. PMID- 10542903 TI - [Treatment priorities in preclinical management of polytraumatized patients]. PMID- 10542904 TI - [Acute treatment of the polytraumatized patient in the emergency room (diagnostic and therapeutic steps)]. PMID- 10542905 TI - [Vertebromedullary injuries in polytrauma]. PMID- 10542906 TI - [Combined skull-brain injury and thoracic trauma: treatment concepts]. PMID- 10542907 TI - [Special features in the diagnosis and treatment of polytraumatized children]. PMID- 10542908 TI - [Differential diagnosis and treatment control of blunt chest trauma with computed tomography]. PMID- 10542909 TI - [Cerebral pressure autoregulation and management of cerebral perfusion pressure (CPP) following head-skull trauma]. PMID- 10542910 TI - [Metabolic monitoring: can monitoring of brain tissue oxygen partial pressure replace measurement of jugular venous oxymetry?]. PMID- 10542911 TI - [Laboratory diagnosis in polytrauma]. PMID- 10542912 TI - [Intra-hospital transport of patients with increased intracranial pressure]. PMID- 10542913 TI - [Transport trauma: fact or fiction?]. PMID- 10542914 TI - [The importance of neurophysiologic monitoring in polytrauma]. PMID- 10542915 TI - [Increased intracranial pressure: therapeutic options]. PMID- 10542916 TI - Bacterial meningitis and deafness. PMID- 10542917 TI - The management of the solitary thyroid nodule: a review. PMID- 10542918 TI - Microbial colonization of silicone voice prostheses used in laryngectomized patients. AB - The aim of this study was to identify the microbial colonization of dysfunctioning voice prostheses in laryngectomized patients and determine the influence of patient radiation therapy on prosthesis life span. In a 40-month period, 257 outpatient voice prosthesis replacements were carried out in a laryngectomized group of 31 patients. The voice prostheses were all removed from the tracheo-oesophageal fistula after dysfunctioning of the prosthesis. Of the replaced prostheses 183 were cultured. The microbial cultures showed a predominant colonization with Candida albicans and commensal oral microflora. Radiation therapy induced xerostomia shortened the lifetime of the first inserted prosthesis in particular. PMID- 10542920 TI - The effect of cold water immersion on the nasal mucosa. AB - The response of the nasal mucosa to cold water immersion is not well known. We have attempted to document this response in normal individuals. Seventeen individuals with no history of nasal disease or allergy were studied. All subjects were asked to perform sustained cold water (15 degrees C) immersion of their hand and forearm on the side of the obstructed nostril for a period of 5 min. The nasal cross-sectional area was measured on both sides of the nose using an acoustic rhinometer. The individuals were then rested for at least 30 min and the test repeated with immersion of the opposite hand. There was a significant fall in nasal cross-sectional area on the side of immersion (median change = 0.32 cm2, P = 0.0003) with a significant rise in nasal cross-sectional area on the none test side (median change = 0.35 cm2, P = 0.0003). There were no significant differences between these results and those obtained by immersion on the opposite side. The results indicate that cold water immersion produces nasal obstruction and that both afferent and efferent arms of this reflex are side-specific. PMID- 10542919 TI - Distinguishing levels of tinnitus distress. AB - Degrees of tinnitus distress were explored in a sample of 216 patients who completed audiological measures and were assessed in a structured interview conducted by a clinical psychologist. The Klockhoff and Lindblom grading system was used and its inter-rater reliability assessed in a subsample showing a high degree of correspondence. Results from the interview are reported in terms of variability of tinnitus, characteristics of problematic situations, distress caused by tinnitus, possibilities to cope, and other influencing factors. Finally, a set of discriminant analyses were conducted on the data set resulting in a final model which included pitch, minimal masking level (MML), tolerance in relation to onset, and avoidance of situations because of tinnitus. This model correctly classified 73% of the subjects into the two levels of distress (grade II and III). There may be a potential role for MML as an outcome variable in tinnitus treatment research. PMID- 10542921 TI - The effect on nasal resistance of an external nasal splint during isometric and isotonic exercise. AB - The now commonplace wearing of external nasal splints by sportsmen and athletes has never been scientifically evaluated. The present study looks into the effect of both isometric and isotonic exercise on nasal resistance and examines if this is altered by the wearing of an external nasal splint. Twenty subjects who did not suffer from rhinitis were tested. Nasal resistance measurements were recorded using an anterior rhinomanometer before and after exercise, with and without an external nasal splint. Pulse and blood pressure were measured using a Criticare Inc. model 508 physiological monitor before and after exercise. Significant changes were observed in pulse (P < 0.001) and both systolic (P < 0.002) and diastolic (P < 0.001) blood pressure in response to isotonic exercise and pulse (P < 0.0001) and diastolic blood pressure (P < 0.0006) in isometric exercise. Significant differences were seen in nasal resistance when the splint was applied before (P < 0.001) and after exercise in both groups (P < 0.003). No significant difference was observed between the post-isotonic exercise groups with and without the splint (P = 0.167) but significant differences were seen in the isometric group (P < 0.0001). External nasal splints decrease nasal resistance at rest but are of little proven value when performing isotonic exercise however significantly reduce nasal resistance during isometric exercise. PMID- 10542922 TI - Chronic otitis media with effusion during infancy, have parent-reported symptoms prognostic value? A prospective longitudinal study from 0 to 2 years of age. AB - As a part of a prospective study (age, 0-2 years), the prognostic value of parent reported symptoms relative to chronic otitis media with effusion (COME) was examined in a group of 122 infants. The occurrence of hearing loss, ear infection, mouth breathing, snoring and common cold was inventoried using a standardised questionnaire filled in by parents at 3-monthly intervals. Tympanometric and otoscopic records were combined for assessment of middle ear status. Subjects were categorized into three groups: none (n = 13), mild (n = 78) and severe (n = 31) COME. Analysis revealed that all symptoms in the first year of life were significantly associated with severe COME. In the second year, only hearing loss was associated with a higher risk for severe COME. The risk for severe COME increased when symptoms were combined. In conclusion, a questionnaire based on only symptoms during the first year of life may assist in screening and managing severe COME. PMID- 10542923 TI - Insertion of ventilation tubes: does the site matter? AB - Several factors are known to affect the length of time a ventilation tube remains in the tympanic membrane. These include the design of ventilation tube, the insertion technique and the presence of intercurrent infection. In addition there are theoretical reasons to suggest that a ventilation tube placed superiorly should remain longer than one placed inferiorly. A randomised prospective study was undertaken on 54 children to test this theory. It showed that there is no significant difference in the extrusion rates for a particular type of ventilation tube (Shah grommet) when comparing the anterosuperior quadrant with the anteroinferior quadrant of the tympanic membrane. PMID- 10542924 TI - Serum vascular endothelial growth factor in patients with head and neck squamous cell carcinoma. AB - Vascular endothelial growth factor (VEGF) is a key pro-angiogenic cytokine expressed by most human tumours. Two isoforms, VEGF121 and VEGF165, are soluble and can be assayed in serum. Serum VEGF has been shown to be significantly raised in patients with solid tumours and shows some promise as a potentially useful tumour marker. Serum levels of VEGF were assayed in 52 patients with untreated head and neck squamous cell carcinoma (HNSCC) and 104 healthy controls. Serum VEGF is significantly raised in patients with HNSCC (P < 0.001), but there was no association with either tumour stage or specifically the presence of nodal metastases. Sixteen patients (31%) had a higher serum VEGF than 95th centile of controls, suggesting that serum VEGF measurement is of little practical use as an initial diagnostic tool. The finding that patients with HNSCC have significantly raised serum VEGF probably relates to enhanced platelet aggregation in these patients. PMID- 10542925 TI - Parameters of nasal airway anatomy on magnetic resonance imaging correlate poorly with subjective symptoms of nasal patency. AB - Forty-four patients undergoing magnetic resonance imaging (MRI) head scans for non-nasal disease were asked to complete a questionnaire immediately after the scan. Subjective patency was scored for each nasal airway, patients were also asked about other nasal symptoms, hay fever, upper respiratory tract infections, medication and any history of nasal surgery or trauma. The following measurements from MRI scans were made: the cross-sectional area of the nasal airway at the anterior end of the middle turbinate, the horizontal width of the inferior turbinate and maximum septal mucosal thickness. In addition the presence of any septal deviation and the thickness or the mucosa of the paranasal sinuses was assessed. Correlation between subjective airway patency and the anatomical parameters studied was generally very weak. However, patients with sinus mucosal thickening on MRI scanning had significantly lower subjective patency scores (left P = 0.003, right P = 0.029) for both nasal airways. Assessment of the nasal airway on MRI correlates poorly with symptoms of nasal obstruction. However, patients with sinus mucosal thickening (> 5 mm) had significantly more symptoms of nasal obstruction on both sides. PMID- 10542926 TI - Systemic absorption of gentamicin in the management of active mucosal chronic otitis media. AB - A prospective study was performed on patients with active mucosal chronic otitis media who were being treated with the gentamicin-containing preparation Gentisone HC. In 27 patients plasma gentamicin levels were measured. Detectable levels were found in 7/27 (26%). Pre- and post-treatment audiometry was performed on 16 patients. There was no statistically significant difference in the change of the mean bone conduction thresholds as a result of treatment, between the treatment and control ears (P = 0.07, Wilcoxon signed Ranks Test at 95% C.I.). We conclude that there is evidence of systemic absorption of gentamicin that would ultimately be absorbed into the perilymph. Gentamicin is known to be ototoxic affecting the vestibular system in lower doses and the cochlea in high doses, hence audiometric assessment is not an appropriate screen for ototoxicity when using topical gentamicin-containing drops. Alternative topical preparations should be further investigated. PMID- 10542927 TI - The radiographic incidence of bony defects in the lateral lamella of the cribriform plate. AB - The purpose of this study was to evaluate the incidence and radiographic characteristics of bony defects of the lateral lamella of the cribriform plate using a coronal computerised tomography (CT) scan. By retrospectively reviewing the coronal CT scans of 410 sinuses from 205 subjects, the overall incidence and the relationship to the subjects' age and depth of cribriform plate were analysed. Bony defects were identifiable in 59.5% of the sinuses. The incidence of a bony defect was not significantly different between the sinuses of the paediatric age group and the adult group. The sinuses having a deeper cribriform plate showed a higher incidence of bony defects in comparison with the sinuses with a less deep cribriform plate. The high incidence of radiologically detectable bony defects in this region may emphasize the particular risk of this area during endoscopic sinus surgery. PMID- 10542928 TI - Manipulation of the fractured nose: a comparison of local infiltration anaesthesia and topical local anaesthesia. AB - Reduction of simple nasal fractures under local anaesthetic is now an accepted practice. The anaesthetic is usually administered using an external percutaneous approach, coupled with topical intranasal cocaine. Topical local anaesthetic with intranasal cocaine is an alternative method. Fifty-four adult patients with simple displaced nasal fractures were randomised to three different groups prior to fracture reduction in the outpatient department. Group A received external local infiltration, Group B received topical EMLA cream whilst Group C received topical AMETOP gel. All patients received topical intranasal cocaine. The cosmetic result and airway patency were comparable in all groups. However, patients in group A perceived the procedure as significantly more painful than the patients of groups B and C. Despite the increased procedure time, we recommend topical anaesthesia as the method of choice to reduce simple nasal fractures in the outpatient department. It offers a similar outcome, whilst being significantly less painful than external infiltration. PMID- 10542929 TI - Paediatric epiglottitis: the influence of the Haemophilus influenzae b vaccine, a ten-year review in the Sheffield region. AB - Cases of acute epiglottitis in children have become very uncommon in recent years. This study set out to find whether the introduction of the Haemophilus influenzae b (Hib) vaccine in the UK in October 1992 has influenced the incidence of acute epiglottitis in the Sheffield region, and whether the pathogenesis has altered. A 10-year retrospective case note review was undertaken. A total of 30 children were treated for acute epiglottitis in Sheffield Children's Hospital over that time period. A sharp decline in the number of cases was found after the vaccine was introduced. Most children presenting with the disease after October 1992 had not been vaccinated. In addition, the pathogens isolated in those children who had received the vaccine were all Streptococci. This is the first study in the UK to examine the influence of the Hib vaccine on acute paediatric epiglottitis. PMID- 10542930 TI - A prospective case-controlled study of 197 men, 50-60 years old, selected at random from a population at risk from hyperlipidaemia to examine the relationship between hyperlipidaemia and sensorineural hearing loss. AB - Hyperlipidaemia has been reported as an aetiological factor in sensorineural hearing loss. Reports on this subject have been retrospective, lacked adequate controls, or have been based on a series of cases which may represent incidental findings as hyperlipidaemia is prevalent in the normal population. The published evidence that hyperlipidaemia causes hearing problems is contradictory and warrants further controlled trials: A prospective case-controlled study was conducted of 197 men between 50 and 60 years old selected at random from a population at risk from ischaemic heart disease or hyperlipidaemia. Hearing thresholds, relevant variables and--on two occasions--their fasting cholesterol levels, were recorded. The control subjects were recruited from patients undergoing nasal surgery for either internal or external structural abnormalities. All study and control groups were also compared with the National Study of Hearing (NSH) data. When hearing thresholds were grouped together as 0.5, 1, 2 and 4 kHz, 0.5, 1, 2 kHz or 4, 6 and 8 kHz there were no statistically significant differences except at 4, 6 and 8 kHz in the worse hearing ear where the at risk group had better hearing thresholds, P = 0.0475. However, the differences which existed became reduced across all frequencies when the subgroup with fasting lipids > 6.5 mmol/l was compared with the at risk group as a whole against the NSH data. PMID- 10542931 TI - Computed tomography and magnetic resonance imaging features of olfactory neuroblastoma: an analysis of 22 cases. AB - The computed tomography (CT) and magnetic resonance imaging (MRI) studies of 22 patients with a histologically proven olfactory neuroblastoma were retrospectively reviewed. The tumours displayed a variety of imaging characteristics and aggressiveness. The expansile tendency of olfactory neuroblastoma is characterised by bowing of the sinus walls. The destructive aspect is manifested as tumour replacing the turbinates, septum, and sinus walls with extension into contiguous areas. The density/signal and enhancement characteristics are non-specific. Olfactory neuroblastoma should be suspected in all ages following identification of a mass in the superior nasal cavity demonstrating both expansile and destructive growth patterns. The otorhinolaryngologist and the radiologist should be aware of this tumour entity, as early diagnosis appropriately guides therapy and predicts survival. PMID- 10542932 TI - Sudden sensorineural hearing loss with fracture of the stapes footplate following sneezing and parturition. AB - Three rare cases of sudden high frequency sensorineural hearing loss with longitudinal fracture of the stapes footplate are presented. In two patients it occurred after they suppressed a sneeze. In the third patient after the exertion of parturition. At exploratory tympanotomy all were found to have longitudinal fractures of the stapes footplate and two had a perilymph fistula at that site. The history and audiometric profiles in such patients should raise a high index of suspicion regarding the possibility of a stapes footplate fracture. PMID- 10542933 TI - Re: Surgical workshop: bipolar scissor tonsillectomy. PMID- 10542934 TI - Re: Bipolar scissor tonsillectomy. PMID- 10542935 TI - Tamoxifen and the endometrium. PMID- 10542936 TI - Expression of steroid receptors, Ki-67, and epidermal growth factor receptor in tamoxifen-treated endometrium. AB - Endometrial specimens of 34 (25 premenopausal and 9 postmenopausal) breast cancer patients receiving tamoxifen were immunohistochemically examined using estrogen receptor (ER), progesterone receptor (PR), Ki-67, and epidermal growth factor receptor (EGFR) antibodies. Proliferative (n = 6), secretory (n = 9), and postmenopausal (n = 6) endometria served as controls. The ER and PR expressions of the glandular cells in tamoxifen-treated patients did not differ from those of the glandular cells in the control women regardless of menopausal status. The Ki 67 index of glandular cells in tamoxifen-induced amenorrheic women was found to be lower than that of the proliferative glandular cells in the control women (p < 0.03), whereas the Ki-67 index of glandular cells in the tamoxifen-treated postmenopausal patients was higher than that of the glandular cells in the control women (p < 0.02). No EGFR overexpression was found in the glandular cells of the tamoxifen-treated premenopausal patients, but expression of EGFR was high in glandular cells of the tamoxifen-treated postmenopausal patients associated with a high Ki-67 index. In competition with ovarian estrogen secretion, tamoxifen may have an antiestrogenic effect on the endometrium, but tamoxifen probably has an estrogenic effect in the absence of ovarian estrogen secretion. This estrogenic effect of tamoxifen may be associated with an EGFR autocrine system. PMID- 10542937 TI - Endometrial cancers in postmenopausal breast cancer patients with tamoxifen treatment. AB - To evaluate possible predicting factors for endometrial cancer in postmenopausal breast cancer patients with tamoxifen treatment, we compared various clinical features between 12 postmenopausal breast cancer tamoxifen-treated patients with endometrial cancer and a control group of 261 otherwise similar patients without this endometrial pathology. These comparisons were based on a long-term prospective follow-up. Several clinical factors such as longer duration of breast disease, older patient age, lower frequency of chemotherapy administration, and higher frequency of postmenopausal bleeding were found among the tamoxifen treated patients with endometrial cancers, and were significantly different when compared to the control group. Only eight (66.7%) had postmenopausal bleeding, and a preoperative diagnosis of endometrial cancer was made in only six (50.0%). When considering postmenopausal bleeding as a marker for endometrial cancer in the study patients, sensitivity was 67% and specificity was 98%. PMID- 10542938 TI - Myxoid and fibrous endometrial stromal tumors of the uterus: a report of 10 cases. AB - Ten endometrial stromal tumors of the uterus with a prominent myxoid or fibrous appearance, or both, that led to problems in interpretation are reported. The patients were 32 to 52 (mean 39) years of age. Three presented with dysfunctional uterine bleeding and one with abdominal pain. An enlarged uterus or a pelvic mass was palpated in five patients; the tumor was an incidental postpartum finding in one patient. All patients underwent hysterectomy. The tumors ranged from 4 to 20 cm in greatest dimension. Six were soft, polypoid intracavitary masses and four were predominantly intramyometrial; two were gelatinous. On microscopic examination, nine tumors infiltrated the myometrium (stromal sarcomas) and one was well circumscribed (stromal nodule). Six tumors had a predominantly fibrous component with the neoplastic cells separated by variable amounts of collagen; extensive areas of hyalinization were present in three tumors. Two tumors were predominantly composed of hypocellular areas with an abundant myxoid matrix, and two had both components in roughly equal proportions. Alcian blue staining was positive, with the staining eliminated by hyaluronidase predigestion, in the myxoid areas. The typical morphologic features of endometrial stromal neoplasia were present focally in four tumors. All of them contained numerous small thin walled vessels. Vimentin and smooth muscle actin were positive in nine of nine and seven of nine tumors, respectively, whereas desmin was negative in six of nine tumors and only focally positive in the other three. One patient had omental nodules at the time of the initial diagnosis and another had a pelvic recurrence 2 years after hysterectomy. Follow-up information is unavailable or short in the other cases. These tumors should be considered of endometrial stromal origin in view of the typical location of most of them, their growth pattern, content of characteristic arterioles, presence of typical endometrial stromal neoplasia in the primary or recurrent tumor in some cases, and absence of evidence of origin from a cell type other than endometrial stroma. These tumors may be identical, in some instances at least, to tumors referred to in the older literature as "myxofibrosarcomas." PMID- 10542939 TI - The use of microsatellite instability in the distinction between synchronous endometrial and colonic adenocarcinomas. AB - The association of endometrial carcinoma with other gynecologic neoplasms, especially ovarian and fallopian tube carcinoma, has been well documented and is usually interpreted as a result of a field defect. Sporadic synchronous primary carcinomas occurring in the endometrium and colon are extremely rare, especially in the absence of the familial genetic abnormalities seen in hereditary nonpolyposis colorectal carcinoma (HNPCC) syndrome, and may present a diagnostic dilemma. Two cases of synchronous adenocarcinomas of the endometrium and colon were studied for genetic abnormalities and differences to test for the presence of two primary tumors. Primary tumors, metastases, and normal tissues were microdissected from formalin-fixed, paraffin-embedded tissues. PCR amplification was performed for microsatellite DNA markers on chromosome 17q and 11q13. The colonic tumors were moderately and poorly differentiated, invasive, nonmucinous adenocarcinomas, whereas one uterine tumor was endometrioid adenocarcinoma and the other was papillary serous carcinoma. Although microsatellite instability, as evidenced by changes in the lengths of the amplified PCR products, was detected at 17q and 11q13 loci in the uterine and colonic neoplasms, the patterns of instability differed between the two primary tumor sites. Moreover, the lymph node metastasis in one colonic tumor had genetic alterations that differed from that of the primary tumor. In both patients, the molecular studies suggested the presence of two synchronous primary tumors. Molecular techniques may assist in distinguishing two separate primaries by determining the contraction and expansion of microsatellite regions in DNA obtained by microdissection from the primary tumors and associated metastases. PMID- 10542940 TI - Post-hysteroscopic ablation reaction: a histopathologic study of the effects of electrosurgical ablation. AB - Ablation of the endometrium is now an established treatment modality for dysfunctional uterine bleeding (DUB). There have been few reports of the inflammatory and healing reaction within the endomyometrium subsequent to this therapy, and its identifying characteristics. The morphologic reaction after electrosurgical ablation in particular has received scant attention. In this retrospective study, 19 women were identified who had undergone both an endometrial ablation using electrosurgical rollerball ablation and a subsequent endometrial sampling or hysterectomy. The average age of the patients was 47 years (range, 28 to 60). Fifteen patients had been ablated for DUB, while four had been ablated immediately after a resection that later revealed atypical hyperplasia. Histopathologic specimens were examined from 1 to 48 months postablation. The six specimens examined at 3 months or less after ablation all exhibited necrotic myometrium, and in five of these six cases, a florid foreign body and granulomatous reaction to necrotic myometrium and spicules of thermally damaged myometrium. A variable degree of acute inflammation was evident in all six cases and was exclusively present in one case. The remaining 13 cases were examined at > 3 months posttreatment. Necrotic myometrium was no longer evident, but a persistent granulomatous, a foreign-body reaction, or both was detected in 5 of 12 cases up to 16 months postablation. In most cases (9 of 12), there was striking endometrial scarring. The morphologic response of the endometrium after electrosurgical endometrial ablation is similar to that reported previously for both resection and laser ablation. Post-hysteroscopic ablative reaction should be recognized and distinguished from other causes of granulomatous endometritis. PMID- 10542941 TI - The stromal component of large endometrial polyps. AB - Benign endometrial polyps belong in the differential diagnosis of adenofibroma and adenosarcoma. There is, however, little information about the range of stromal mitotic activity, stromal cellularity, and stromal atypia in benign endometrial polyps, rendering the differential diagnosis with the aforementioned tumors problematic. In this study, the stroma of 66 polyps 1 cm or more in greatest dimension from 56 patients was analyzed for stromal mitotic activity, cellularity, and atypia. Sixteen (24%) had an almost completely fibrotic stroma that had rare mitoses, little cellularity, and no atypia. However, 50 polyps (76%) had stroma that was predominantly endometrial or was a mixture of endometrial-type stroma and fibrous stroma. In these polyps stromal mitoses were relatively common, averaging 1.2/10 MFs/HPFs (range, 0-5.8 MFs/10 HPFs). Stromal cellularity was frequently equal to or mildly increased over adjacent nonpolypoid endometrial stroma and mild nuclear atypia (enlarged stromal nuclei) was also common. Twelve polyps (24%) from the group of 50 had two or more MFs/10 HPFs, a mitotic rate present in some adenosarcomas. None of these polyps had other features necessary for the diagnosis of adenofibroma or adenosarcoma and follow up in all patients was uneventful (average follow-up, 96 months). It is concluded that benign polyps that retain areas of endometrial-type stroma often have mitotic activity and that significant stromal mitotic activity (> or = 2 MFs/HPFs) is relatively common. These polyps do not have significant stromal atypia nor do they have a marked increase in stromal cellularity. Thus, in the absence of other supportive features, stromal mitotic activity alone should not be regarded as a worrisome finding. PMID- 10542942 TI - p53, c-erbB, and Ki-67 expression in ovaries removed prophylactically from women with a family history of ovarian cancer. AB - Prophylactically removed ovaries from 64 women were compared with those from 30 women with no known family history of ovarian cancer for expression of the p53 tumor suppressor protein, the c-erbB-2 oncoprotein, and for the proliferation antigen Ki-67. All analyses were performed without knowledge of the family history. Ki-67 was expressed in rare nuclei in both the surface and cyst epithelial cells, whereas p53 was expressed in rare nuclei only in cyst epithelial cells. Neither the proportion of positive cases nor the intensity of staining differed between groups. c-erbB2 was not expressed in surface or cyst epithelium in any case from either group. Nuclei of granulosa and granulosa lutein cells expressed both Ki-67 and p53. In conclusion, increased expression of p53, c-erbB2, and Ki-67 was not found in the epithelium of prophylactically removed ovaries, suggesting that increased expression occurs later in the development of carcinoma or invasive tumor evolves too quickly to identify expression of these proteins in preinvasive epithelium. PMID- 10542943 TI - Expression of metallothionein and nuclear size in discrimination of malignancy in mucinous ovarian tumors. AB - Metallothioneins (MTs) are low molecular weight proteins that control cell proliferation via their metalloregulatory function. Several studies in various tumors have shown their influence in determining response to chemotherapy and prognosis. Because there has been no such study pertaining to ovarian tumors, we investigated MT expression and nuclear size in mucinous ovarian neoplasms (12 benign, 6 borderline, and 8 malignant). The percentage of MT-positive stained cells was significantly higher in the borderline than in the benign tumors, but lower than in the malignant tumors. Single layers of cells in the borderline tumors showed mild immunostaining in 50% of the cells and moderate staining in the remaining 50%, while 83.3% of cells within multilayered epithelium showed moderate to strong immunostaining. In the carcinomas, 87.5% of tumors showed moderate to strong staining in single-layered epithelium and moderate to strong staining of all the cells in multilayered epithelium. Morphometry measurements showed that the mean nuclear area of cells in the carcinomas was significantly larger than in the borderline or benign tumors. The nuclear area of cells in the carcinomas with early recurrence or metastasis was also significantly larger than in carcinomas without recurrence or metastasis. It is concluded that MT protein expression and nuclear size are possible markers for the evaluation of the progression of malignancy in mucinous ovarian tumors. PMID- 10542944 TI - Prognostic factors in Paget's disease of the vulva: a study of 21 cases. AB - Twenty-one cases of vulvar Paget's disease were studied to assess possible prognostic indicators, including presence and depth of invasion, status of resection margins, tumor DNA cell content, and immunoreactivity for p53 and estrogen receptor proteins. Immunostaining for cytokeratin 7 (CK7), cytokeratin 20 (CK20), and gross cystic disease fluid protein-15 (GCDFP) were also performed. Patients were 45 to 82 years of age (mean, 66.9 years). Ten of 21 patients (47.6%) had invasive Paget's disease. Dermal invasion was < or = 1 mm in 7 of 10 cases and 2 mm, 3 mm, and 8 mm in the remaining three invasive tumors. Of the seven patients with minimally invasive Paget's disease (< or = 1 mm depth of invasion), five are alive with no evidence of disease, one died of an unrelated illness, and one is alive with biopsy-proven in situ Paget's disease, having refused operative treatment. Of the three patients with more than minimally invasive Paget's disease (> 1 mm), all had nodal metastases; one patient is alive with no evidence of disease, one died of undertermined causes, and one died of metastatic Paget's disease. The remaining 11 patients had Paget's disease confined to the epidermis and its adnexal structures. Seven of these patients were alive at last follow-up with no evidence of disease. Of the remaining four patients, one died of metastatic cervical cancer, one died of metastatic bladder cancer, one died of an unrelated illness, and one patient is alive with biopsy proven in situ Paget's disease and awaiting operative treatment. Twenty of the 21 cases represented primary vulvar Paget's disease while one represented possible local spread from a cervical adenocarcinoma. The immunoprofiles were GCDFP+/CK7+/CK20- in 14 cases, GCDFP+/CK7+/CK20+ in 4 cases, and GCDFP-/CK7+/CK20 in 2 cases. All tumors were estrogen receptor-negative. Immunostaining for p53 was positive in 16 tumors and negative in four tumors. Seven of 12 (58%) patients with positive margins experienced local recurrence of Paget's disease, while the disease recurred in 1 of 4 patients with negative margins. Recurrence was observed in 3 of 5 patients with diploid tumors and in 4 of 10 patients with aneuploid tumors. Neither of these differences is statistically significant. This study supports the recognition of a category of minimally invasive vulvar Paget's disease that has a low risk of distant metastasis and death caused by disease. Status of surgical resection margins, tumor cell DNA ploidy, estrogen receptor expression, and p53 immunoreactivity are not predictive of local recurrence. PMID- 10542946 TI - Dermatofibrosarcoma protuberans of the vulva: clinicopathologic and immunohistochemical analysis of four cases, one with fibrosarcomatous change, and review of the literature. AB - Only 18 cases of dermatofibrosarcoma protuberans (DFSP) of the vulva have been reported. This article describes the clinicopathologic and immunohistochemical findings of four additional vulvar DFSPs. The median patient age was 54 years (range, 36 to 69 years) and the most common clinical presentation was a slowly growing vulvar mass. The tumors occurred in the left paraclitoral area (1 case), right labium majus (1 case), left labium majus (1 case), and mons pubis (1 case). The tumor size, known in three cases, ranged from 1.2 to 5.0 cm in greatest dimension. Microscopically, the tumors were composed of bland spindle cells with oval or wavy nuclei arranged in a cartwheel pattern; in one case, fibrosarcomatous areas were apparent. CD34 immunostaining was positive in the three cases in which it was performed; positive staining was also seen in the area of fibrosarcoma. Estrogen and progesterone receptor immunostains were negative in three cases. Of the three patients who underwent surgery, one had tumor recurrences every 2 to 4 years for 18 to 20 years but has remained free of tumor in the 7 years since the last excision, one was free of tumor for 12 years before dying of an unrelated cause, and the other has been free of disease for 6 months. The fourth patient refused treatment and has persistent disease in the vulva. This study shows that the behavior of DFSP in the vulva is similar to its behavior in its more common locations. PMID- 10542945 TI - Localization of chromogranin, synaptophysin, serotonin, and CXCR2 in neuroendocrine cells of the minor vestibular glands: an immunohistochemical study. AB - Sections of vulvar tissue containing minor vestibular glands (15 sections from 14 women with vulvar vestibulitis syndrome and six control sections from five asymptomatic women) were immunohistochemically stained with antibodies against chromogranin, synaptophysin, serotonin, and CXCR2. The degree of inflammation in vestibular tissue from women with and without vulvar vestibulitis syndrome was not significantly different. All vulvar minor vestibular glands contained neuroendocrine cells that expressed chromogranin to some degree. Only one case failed to express synaptophysin. The number of cells expressing chromogranin and synaptophysin was the same regardless of the degree of inflammation. However, moderate to severe inflammation was associated with a statistically significant increase in the number of cells expressing serotonin (p < 0.001) and CXCR2 (p < 0.02). It is concluded that neuroendocrine cells are present within minor vestibular glands of the vulva. The number of cells expressing the inflammatory mediator serotonin and CXCR2, the shared interleukin-8 receptor, are upregulated with inflammation. Chronic inflammation is a normal finding in vestibular tissue and does not serve as a histologic marker for vulvar vestibulitis. PMID- 10542947 TI - Frequent allelic imbalance of tumor suppressor gene loci in cervical dysplasia. AB - In addition to human papillomavirus (HPV) infection, loss of heterozygosity (LOH) at tumor suppressor gene loci has been frequently observed in cervical cancer. Thus, it may be assumed that detection and characterization of specific LOH profiles in preneoplastic lesions, in addition to HPV typing, might facilitate assessment of progression risk of cervical dysplasia. In this study, the type and frequency of allelic imbalance (allelic loss or allelic reduction) were analyzed in 24 unrelated cervical lesions using 14 polymorphic microsatellite markers at different tumor suppressor gene loci. No allelic loss was observed in four condylomatous lesions, whereas 2 of 13 (15%) CIN I lesions displayed allelic loss at 3p25 and 5q11-13. In high-grade lesions, however, allelic loss occurred in four of six (66%) cases at multiple chromosomal regions (3p14-25, 5p15, 5q11, 5q21, 11p15, and 17q21). Allelic reduction was observed in 4 of 13 (30%) low grade lesions and 3 of 6 (50%) high-grade lesions. LOH was confined to lesions infected by high-risk HPV types. These data suggest that chromosomal instability is an early event in cervical carcinogenesis. The detection of LOH on multiple chromosome 3p loci in 50% of high-grade lesions suggests that a specific marker panel encompassing this region might enable better assessment of which lesions are likely to regress, persist, or progress. PMID- 10542948 TI - Frequent expression of beta-human chorionic gonadotropin (beta-hCG) in squamous cell carcinoma of the cervix. AB - Human chorionic gonadotropin (beta-hCG) has been detected within tissue homogenates, culture fluid, and sera of patients with squamous cell carcinoma of the cervix. Studies regarding in vivo localization of beta-hCG in squamous cell carcinoma of the cervix are scant and conflicting. Cervical samplings (biopsy and/or curettage specimens) of 63 cases of poorly differentiated invasive squamous cell carcinoma of the cervix were initially stained by the immunoperoxidase technique for the presence of beta-hCG and human placental lactogen (hPL). Based on beta-hCG reactivity, patients were divided into beta-hCG positive and beta-hCG-negative groups. Thirty-three of the 63 (52%) cases showed localization of beta-hCG in tumor cells. Subsequent specimens of patients, who underwent surgical treatment, were likewise examined for beta-hCG reactivity. These surgical specimens showed focal beta-hCG reactivity in the beta-hCG positive group only. The beta-hCG reactivity was seen in both high-grade SIL (CIN III), invasive squamous cell carcinoma, and its metastases. The focal beta-hCG reactivity was predominantly confined to the peripheral tumor cells at the stromal-epithelial interface in noninvasive and invasive lesions. Intensity of immunostaining was moderate to strong. The beta-hCG staining was observed in different cancer stages and in various age groups. No hPL reactivity was seen in any cases. Poorly differentiated squamous cell carcinoma of uterine cervix showing immunoreactivity for beta-hCG should be distinguished from choriocarcinoma and other trophoblastic tumors. PMID- 10542949 TI - Endometrial carcinoma in tamoxifen-treated breast cancer patient: clinicopathological, immunohistochemical, and genetic analysis. AB - Endometrial polyps and endometrial neoplasms are a recognized complication of chronic tamoxifen treatment. This study describes an endometrial carcinoma that developed in a woman receiving low-dose tamoxifen treatment for breast cancer. Little is known about steroid receptor status, somatic alterations in oncogenes and tumor suppressor genes, and inherited susceptibility in endometrial carcinomas associated with tamoxifen use. In the present case, the endometrial carcinoma was negative for estrogen receptors and weakly positive for progesterone receptors. In addition, analysis of K-ras, c-erbB2/neu, cyclin D1, and p53 status revealed a codon 12 point mutation in the K-ras oncogene. The patient was determined not to be a carrier of germ-line mutations in cytochrome P 450 1A1 (CYP1A1), an estrogen-metabolizing gene previously associated with enhanced endometrial cancer risk, but she was a carrier of a methylenetetrahydrofolate reductase gene variant related with putative alterations in DNA methylation. PMID- 10542950 TI - Endocrine cell micronests in an ovarian mucinous cystadenofibroma: a mimic of microinvasion. AB - An ovarian mucinous cystadenofibroma with peculiar neuroendocrine cell micronests is described in a 59-year-old Japanese woman. Aggregates of epithelial cells resembling microinvasive carcinoma cells were scattered throughout the adenofibromatous area. These micronests were composed of small uniform cells with argentaffin and argyrophil granules. Numerous small cells with neuroendocrine granules were also seen within mucinous glands. This is the first report of neuroendocrine micronests in an ovarian neoplasm, a finding that should be distinguished from microinvasion. PMID- 10542952 TI - Uterine Wilms' tumor. PMID- 10542951 TI - Collision of uterine rhabdoid tumor and endometrioid adenocarcinoma: a case report and review of the literature. AB - Extrarenal malignant rhabdoid tumors have been reported in a variety of anatomic sites but infrequently in the female genital tract. In the uterus, they have been described as a pure tumor, in association with endometrial stromal sarcomas, and as a component of a malignant mullerian mixed tumor. This study reports an unusual uterine neoplasm in a 49-year-old woman, in which a malignant rhabdoid tumor occurred as a collision tumor with a well-differentiated endometrioid adenocarcinoma. The tumor was a 14-cm polypoid mass that filled the endometrial cavity. The two neoplastic components were distinct on microscopic and immunohistochemical examination. Ultrastructural examination confirmed the rhabdoid phenotype of the sarcomatous component. The patient died of disease 4 months after diagnosis with progression of the malignant rhabdoid tumor. The highly aggressive behavior of the rhabdoid (i.e., nonepithelial) component in this collision tumor lends support for a distinction of this neoplasm from a malignant mullerian mixed tumor, with which it may be confused. PMID- 10542953 TI - [Regression of Hensen's node and axial growth of the embryo]. AB - Hensen's node is the gastrulation center in the avian embryo. It is the homologue of the amphibian dorsal blastopore lip and the zebrafish shield. It contains the progeny of all midline cells (floor plate of the neural tube, notochord and dorsal endoderm). However, microsurgical experiments on Hensen's node allow to think that organizer function is due to an extremely limited region situated in the caudal part of Hensen's node which corresponds to the boundary between prospective axial mesoderm rostrally and paraxial mesoderm caudally. This interface is essential for Hensen's node regression and organization of the caudal part of the body. PMID- 10542954 TI - [Segmentation in vertebrates: a molecular clock linked to periodic somite formation]. AB - In the vertebrate embryo, somites constitute the basis of the segmental body pattern. They give rise to the axial skeleton, the dermis of the back and all striated muscles of the body. In the chick embryo, a pair of somites buds off, in a highly coordinated fashion, every 90 minutes, from the cranial end of the presomitic mesoderm (PSM) while new mesenchymal cells enter the paraxial mesoderm as a consequence of gastrulation. The processes leading to the segmentation of the somite are not yet understood. We have identified and characterised c-hairy1, an avian homologue of the Drosophila segmentation gene, hairy. c-hairy1 is strongly expressed in the presomitic mesoderm where its mRNA exhibits a cyclic posterior-to-anterior wave of expression whose periodicity corresponds to the formation time of one somite (90 min). Fate mapping of the rostral half of the PSM using the quail-chick chimera technique supports a model of cryptic segmentation within the presomitic mesoderm, and indicates that c-hairy1 expression dynamics are not due to massive cell displacement. Analysis of in vitro cultures of isolated presomitic mesoderm demonstrates that rhythmic c hairy1 mRNA production and degradation is an autonomous property of the paraxial mesoderm. Rather than resulting from the caudal-to-rostral propagation of an activating signal, it arises from pulses of c-hairy1 expression that are coordinated in time and space. Blocking protein synthesis does not alter the propagation of c-hairy1 expression, indicating that negative autoregulation of c hairy1 expression is unlikely to control its periodic expression. Most of the segmentation models proposed for somite formation rely on the existence of an internal clock coordinating the cells to segment together to form a somite. These results provide the first molecular evidence of a developmental clock linked to segmentation and somitogenesis of the paraxial mesoderm, and support the possibility that segmentation mechanisms used by invertebrates and vertebrates have been conserved. PMID- 10542955 TI - [Somite patterning and segregation of different somite lineages]. AB - The somite is a transient embryonic mesodermal structure, found only in vertebrates. In amniotes, somites give rise to the dermis of the back, to the striated skeletal muscles of the trunk and limbs and to the vertebral column and ribs. Segregation of these different lineages is linked to the establishment of two somitic polarity axes, a dorso-ventral one and a medio-lateral one. While the establishment of the former relies essentially on extrinsic cues, that of the latter obeys to both intrinsic and extrinsic mechanisms. Concerning the environmental cues regulating the establishment of both axes, somitic regionalisation results from antagonistic or combinatorial influences mediated by diffusible factors, such as Sonic Hedgehog, Wnt and Bmp-4, that act in gradients. PMID- 10542956 TI - [Molecular aspects of vertebral chondrogenesis]. AB - In this article, we review the data showing that dorso-ventral polarization of the vertebral cartilage involves two distinct molecular pathways, acting sequentially during development. The first one is mediated by the notochord and the ventral part of the neural tube, by the action of the Sonic Hedghehog protein (SHH): it directs the formation of the cartilaginous pieces located in a deep position within the embryo. The second one is responsible for the differentiation of the dorsal parts of the vertebra, differentiating under the ectoderm; it is initiated by the roof plate of the neural tube, by the action of the secreted protein BMP4. Those two pathways cannot be exchanged, rather they are antagonists. These data allow us to draw a new model for vertebral chondrogenesis. PMID- 10542958 TI - [Corticosteroid hormones and the brain]. AB - The anatomical and functional links between the hormone stress axis and the cortico-limbic brain regions which integrate emotion and motivation are well documented. It is important, considering the consequences of stress on the brain, to take into account the regulatory buffer capacities of the personality cognitive processes. Another point of interest is evaluation of the long term effects of repeated life events on chronic environmental pressures which induce brain negative feedback defects and, subsequently, insidious cellular changes in regions such as the hippocampus that lead to memory or adaptive impairments. An example is provided by perinatal stress that induces, later in life, both hormonal and cognitive deleterious changes. PMID- 10542957 TI - [Steroid receptors and mechanism of action of sex steroids]. AB - Steroid hormone receptors define a large family of proteins. Recently, a new estradiol receptor has been identified. This discovery suggests the existence of a previously unrecognized pathway of estrogen signalling. Moreover, it implies important pharmacological consequences. Receptors activation induces the modulation of transcription of specific genes. Proteins involved in this effect have been identified: coactivators, corepressors and cointegrators. Their mechanism of action have been characterized. They modify histone acetylation of the corresponding promotor. Sex steroid receptors are located in the nucleus. This nuclear localization is in fact a dynamic situation, resulting from a continuous shuttling of the receptor between the cytoplasm and the nucleus. Non genomic effects of steroids have also been described. PMID- 10542959 TI - [Neurosteroids: trophic effects in the nervous system]. AB - Some steroids, named "neurosteroids", can be synthesized from cholesterol within both the central and peripheral nervous systems. Thus, pregnenolone and progesterone persist in the brain and in peripheral nerves long after removal of the steroidogenic endocrine glands by castration and adrenalectomy. The role of neurosteroids during the development of the nervous system is not well known, although they are synthesized by glial cells and some populations of neurons already during embryonic life. Cell culture experiments suggest that neurosteroids may influence the survival and differentiation of neurons and glial cells. In the adult nervous system, neurosteroids play an important role during regeneration. Progesterone is indeed synthesized by Schwann cells in peripheral nerves, where it plays an important role in the formation of new myelin sheaths after lesion. This is the first demonstration of a vital role for a neurosteroid in the nervous system. PMID- 10542960 TI - [Neurosteroids: behavioral aspects and physiological implications]. AB - The term "neurosteroids" applies to those steroids that are both formed in the nervous system from sterol precursors, and accumulate in the nervous system, at least in part, independently of peripheral steroidogenic glands secretion. Neurosteroids that are active on the central nervous system include, mainly, pregnenolone (PREG), dehydroepiandrosterone (DHEA) and their sulfate esters (PREG S and DHEA-S), as well as the reduced metabolite of progesterone, 3 alpha,5 alpha TH PROG also called allopregnanolone. These neuroactive neurosteroids alter neuronal excitability by modulating the activity of several neurotransmitter receptors and thus can influence behavior. PREG-S decreases the sleeping time in rats anesthetized with a barbiturate, which is consistent with its antagonist action on the GABAA receptor (GABAA-R). Allopregnanolone is anxiolytic in rats tested in a conflict paradigm, through an interaction at a site specific for the benzodiazepine (BZ) receptor inverse agonist RO15-4513 and/or at the picrotoxinin site on GABAA-R. The contribution of the amygdala, a key region involved in the control of anxiety, is also demonstrated for the anxiolytic action of allopregnanolone. An anti-agressive effect of DHEA can be observed in castrated male mice who become agressive in the presence of lactating females. This inhibition of agressiveness by DHEA is associated to a selective decrease in the brain of PREG-S, which may, in turn, trigger an increase of endogenous GABAergic tone. Finally, cognitive performances of aged rats tested in the Morris water maze and the Y-maze can be correlated with individual concentrations of PREG-S in the hippocampus, i.e. poor performance in both tasks with low levels of PREG-S. Remarkably, the memory deficits are significantly improved, albeit transiently, by an intra-hippocampal injection of PREG-S in impaired aged rats. Promnesiant PREG-S may then reinforce some neurotransmitter systems that can decline with age. This brief review provides evidence of the pharmacology and physiological correlates of neurosteroids involved in behavioral phenomena. However, neurobiological mechanisms of behavioral effects of neurosteroids await further investigation. PMID- 10542961 TI - [Models for the study of memory and neurosteroids]. AB - The steroids dehydroepiandrosterone sulfate (DHEA-S) and pregnenolone sulfate (Preg-S) are naturally synthetized in the brain. They improve short term and long term memory performances in a variety of learning tasks and models of amnesia in rodents. DHEA-S and Preg-S modulate GABAergic and glutamatergic synaptic transmission through direct interactions with GABA-A, NMDA and/or sigma 1 membrane receptors. In addition, these two neurosteroids facilitate the release of acetylcholine and modulate synaptic plasticity phenomena in cerebral structures, such as the hippocampus, known to play a role in learning and memory processes. The possible links between these actions and the promnestic effects of DHEA-S and Preg-S are discussed in the present review. PMID- 10542962 TI - [Perspectives and hopes in infectious pathology]. AB - Over the last few decades, changes in socio-economic conditions and social practices as well as aggressive therapy of many diseases have led to the emergence of new infectious pathologies. These new pathologies are either associated with newly identified microbial species or the emergence of known microbes which have encountered new environments in which they are able to cause disease. Recent progress has allowed us to understand the mechanisms by which these pathogens express their virulence and will certainly allow us to diagnose and treat these infections more efficiently in the future. PMID- 10542963 TI - [Prion diseases. The "prion" a remarkable infectious agents]. AB - Prion diseases are a set of brain degenerative syndromes developed by many mammals. The epidemiological characteristics are remarkable, the origin of the disease is either infectious, genetics or sporadic. A protein synthesised by the host, the so-called prion protein (PrP), seems to be both the etiologic agent and it is also responsible for the induced pathology. This protein is found under two very different conformations. The normal cellular form (PrPC) is alpha-helix rich while the pathological (PrPSc) conformation is mainly composed of beta-sheet structures and resist proteinase-K attack. The conversion of the PrPC isoform to a structure resisting to proteinase-K has been demonstrated in vitro. In order to understand these phenomena, physico-chemical models have been proposed. PMID- 10542964 TI - [Molecular dialogue between human retroviruses and host cells]. AB - Several exogenous retroviruses are etiologic agents of severe human diseases. Retrovirus replication depends on a plethora of highly specific interactions with the host cell resulting in the subversion of multiple cellular signal transduction pathways. Understanding the molecular cross-talk that regulates the relationship between the virus and the host cell is currently the objective of many researchers and should contribute to gain insight into puzzling features of the physiopathology of retrovirus infections. A large body of recent data indicates that retroviruses utilize a variety of unrelated cell surface receptors to initiate infection and modulate the homeostasis of the target cell following receptors binding. As an example, I discuss here the tremendous capacity of HIV-1 envelope glycoproteins to modulate kinases activation and nuclear translocation of transcription factors following binding to surface receptors CD4 and CXCR4. Viral envelope glycoproteins interaction with CD4 controls cell activation and proliferation required for virus production whereas interaction with CXCR4 favors apoptosis thereby decreasing the host immune response. PMID- 10542965 TI - [Cellular defence reactions and their depression in insects]. AB - In insects the main cellular defence reactions are phagocytosis and encapsulation of foreign bodies. Free cells of haemolymph called haemocytes are the effectors of these reactions. They are achieved under the control of humoral factors of the plasma or of the serum. Humoral factors are able to enhance or to decrease the cellular defence reactions. As in mammals, potential parasites or pathogens need to avoid or to inhibit the defence reactions before developing inside the insect body. As an example of the depression of immunity induced by a parasite we will study the relationships between an insect and a nematobacterial complex. PMID- 10542966 TI - [Ciguatoxins and brevetoxins: dissection of the neurobiological actions]. AB - This review focuses on the neurobiological actions of ciguatoxins and brevetoxins which are phycotoxins produced respectively by the dinoflagellates Gambierdiscus toxicus and Ptychodiscus brevis. These actions are illustrated in particular by the effects of the toxins on myelinated nerve fibres and on skeletal neuromuscular junctions of vertebrates. Ciguatoxins and brevetoxins, through different vectors, are responsible for human intoxications characterized mainly by neurological disturbances. The molecular target of these families of lipid soluble cyclic polyethers is the voltage-gated sodium channel, a fundamental transmembrane protein involved in cellular excitability. The different toxins share a common binding site (the receptor-site 5) located on the alpha sub-unit of this neuronal transmembrane protein. Electrophysiological studies of the mode of action of ciguatoxins and brevetoxins identify these toxins as specific sodium channel activators. Indeed, during the action of these phycotoxins, sodium channels remain permanently opened, at the resting membrane potential, which produces a continuous entry of sodium ions in most excitable cells. Such a sodium entry has various consequences on sodium-dependent physiological mechanisms, consisting in a membrane depolarization which, in turn, causes spontaneous and/or repetitive action potential discharges and thereby increases membrane excitability. These neuronal discharges may be transient or continuous according to the preparation and the toxin tested. The increase in membrane excitability during the action of ciguatoxins and brevetoxins is responsible for the different effects exerted by these toxins on various chemical synapses and secretory cells. Another consequence of the continuous entry of sodium ions into cells was revealed using confocal laser scanning microscopy and vital staining of plasma membranes with the fluorescent dye FM1-43. These techniques made feasible the dynamic study of morphological alterations produced by ciguatoxins and brevetoxins on various cellular preparations in situ. Thus, it has been possible to bring to the fore that these phycotoxins cause a marked increase in the volume of nodes of Ranvier of myelinated nerve fibres, motor nerve terminals innervating skeletal muscle and perisynaptic non-myelinating Schwann cell somata. This increase could be reversed by hyperosmotic external solutions and completely prevented by the blockade of voltage-gated sodium channels. The mechanisms involved in the increase in cellular volume, during the action of ciguatoxins and brevetoxins, are discussed. PMID- 10542967 TI - An empirical analysis of two forms of extinction to treat aggression. AB - We compared the effectiveness of two extinction interventions, extinction by omission and noncontingent delivery of reinforcement, to treat aggressive behavior with a 10-year-old boy. Before the intervention, a functional analysis revealed that aggression was maintained by positive reinforcement in the form of attention. The extinction by omission intervention consisted of ignoring aggressive behavior. Noncontingent reinforcement involved delivering attention to the boy on a fixed-time schedule. Both treatments were compared using a multielement research design. Noncontingent reinforcement produced a more rapid elimination of aggression. Additionally, the schedule of noncontingent reinforcement was gradually thinned during the intervention. Finally, both parents successfully implemented the noncontingent schedule of reinforcement independently for up to 5 weeks after treatment. PMID- 10542968 TI - A comparison of the effectiveness of brief versus traditional functional analyses. AB - The study compared a brief and an extended (i.e., traditional) functional analyses with three adults with serious developmental disabilities. Two of the subjects exhibited high levels of aggressive behavior, whereas the third engaged in self-injury. Both analyses examined conditions such as tangible reinforcement, attention, demand, alone and play (i.e., baseline). The brief functional analysis also included functional communication training in which the subjects learned a relevant mand. The brief and extended functional analyses revealed the same controlling variables in all cases, but the brief functional analyses took less than 20% of the time in analog conditions as the extended analyses. These results further the case for the utility of brief functional analyses. We caution, however, that behavior analysts should not generalize from a study that involved only three subjects and that brief functional analyses may be particularly sensitive to establishing operations. PMID- 10542969 TI - Correlates of restraint use in an institutional population. AB - Demographic and behavioral correlates of the use of restraint were analyzed in an institutional population of 300 persons with developmental disabilities. Examination of the frequency distributions of restraint frequency and duration indicated that there were 33 consumers who experiences relatively few, short duration restraint and 11 consumers who had daily restraint for many hours. Separate analyses of these two kinds of restraint were performed. Short-duration restraint was predicted by behavioral variables indicating extra-personal maladaptive behaviors. Demographic variables did not predict short-term restraint. Multiple regression analysis indicated that only independent predictor of short-term restraint was the severity rating of Hurts Others. Similar results were found for predictors of the duration of short-term restraint. The only correlates of long-duration restraint was low weight, Hurts Self, and Withdrawn Behavior. Multiple regression analysis indicated that the only variable that independently predicted long-duration restraint was the severity rating of Hurts Self. The implications of these data for the management of restraint are discussed. PMID- 10542970 TI - The Verbal Behavior Assessment Scale (VerBAS): construct validity, reliability, and internal consistency. AB - A questionnaire, the Verbal Behavior Assessment Scale, was developed to assess the communicative functions of individuals with developmental disabilities and severe mental handicap. To assess its psychometric characteristics, the questionnaire, comprising 15 items, was administered to pairs of caregivers of 115 participants. Exploratory factor analysis, involving 11 more participants, revealed satisfactory evidence concerning the distinction of three different communicative functions with the present sample. The questionnaire had good levels of interrater reliability and internal consistency. PMID- 10542971 TI - Acquisition and transfer of sentence construction in autistic students: analysis by computer-based teaching. AB - In this study, we examined the conditions necessary to construct appropriate sentences in three autistic students using computer-based training and testing procedure. In Experiment 1, when a picture was presented on the computer display as a sample stimulus, the student was required to construct an appropriate sentence with five words. After training with three stimuli, each student could construct the correct sentence for 24 untrained stimuli. Appropriate vocal responses also emerged. In Experiment 2, the appropriate use of particles, which specify the subject and the object, was acquired by particle choice or sentence construction training. The rule was transferred to untrained stimuli and writing response. These results are discussed in terms of applicability of computer-based training for establishing appropriate sequential responding and particle usage in autistic students. PMID- 10542972 TI - The development of a reinforcer choice assessment scale for persons with severe and profound mental retardation. AB - Reinforcement is a cornerstone for treating adaptive skills and eliminating problem behaviors for persons evincing mental retardation. To effectively use reinforcement, professionals must find reinforcers that are most effective for each individual. Establishing reinforcers is particularly important for persons functioning in the severe and profound ranges of mental retardation as reinforcers are often difficult to identify in this population. The current study aimed at developing a reinforcer rating scale that-would have applicability on a broad clinical basis, looking specifically at the psychometric properties of the scale. PMID- 10542973 TI - Dose-ranging study of a new steroid for asthma: mometasone furoate dry powder inhaler. AB - A new formulation of mometasone furoate (MF) for administration by dry powder inhaler (DPI) was evaluated for the treatment of asthma. A 12-week, double-blind, placebo-controlled dose-ranging study compared the efficacy and safety of three doses of MF DPI (100, 200 and 400 mcg b.i.d) with beclomethasone dipropionate (BDP) 168 mcg b.i.d. administered by metered dose inhaler in 365 adult or adolescent patients being treated with inhaled glucocorticoids. The mean change from baseline to endpoint (last treatment visit) for forced expiratory volume in 1 sec (FEV1) was the primary efficacy variable. Secondary efficacy variables included other objective measures of pulmonary function [forced vital capacity (FVC), forced expiratory flow 25-75% (FEV25-75%.) and peak expiratory flow rate (PEFR)] as well as subjective measures of therapeutic response (patients' daily evaluation of asthma symptoms and physicians' evaluation). At endpoint, all four active treatments were significantly more effective than placebo (P < 0.01) in improving FEV1 (MF DPI 5 to 7%, BDP 3%, placebo -6.6%) and all other measures of pulmonary function (FVC: MF DPI 4 to 5%, BDP 2%, placebo -4.7%; FEF25-75%: MF DPI 6 to 18%, BDP 7.5%, placebo -9.5%; PEFR (AM): MF DPI 5 to 10%, BDP 5.7%, placebo 7%). A consistent trend was observed for better improvement in patients treated with MF DPI 200 mcg b.i.d. than with MF DPI 100 mcg b.i.d., with no apparent additional benefit of MF DPI 400 mcg b.i.d. Results for the MF DPI 100 mcg b.i.d. and BDP 168 mcg b.i.d. treatment groups were similar. Patients' and physicians' subjective evaluations of symptoms found similar improvement in the MF DPI 200 and 400 mcg b.i.d. treatment groups, which were slightly better than that in the MF DPI 100 mcg b.i.d. group. Symptoms tended to worsen in the placebo group. MF DPI was well tolerated at all dose levels and the most frequently reported treatment-related adverse effects were headache, pharyngitis and oral candidiasis. No evidence of HPA-axis suppression was detected in any treatment group. In summary, all doses of MF DPI were well tolerated and significantly improved lung function and MF DPI 400 mcg (200 mcg b.i.d.) was the optimal dose in this study of patients with moderate persistent asthma. PMID- 10542974 TI - The effect of high-dose fluticasone propionate and budesonide on lung function and asthma exacerbations in patients with severe asthma. AB - The purpose of this study was to investigate the comparative efficacy and safety of equal doses of inhaled fluticasone propionate (FP) and inhaled budesonide (BUD) using their respective dry powder inhalers in a population of severe asthmatics requiring high doses of inhaled corticosteroid. This double-blind double-dummy parallel-group study compared the effects of 24 weeks of treatment with FP (2000 micrograms daily via a Diskhaler inhaler; Glaxo Wellcome, Evreux, France) and BUD (2000 micrograms daily via a Turbuhaler inhaler; Astra Pharmaceuticals, Rijswijka, Netherlands) on lung function and asthma exacerbations in 395 patients with asthma. FP was statistically significantly superior to BUD with respect to the percentage of symptom-free days (P = 0.02), the incidence of days free from rescue bronchodilator usage (P = 0.02) and the distribution of change in peak expiratory flow (PEF) expressed as a percentage of the predicted PEF (P = 0.04). During the treatment period FP was statistically significantly superior to BUD for change in forced expiratory volume in 1 sec (FEV1) at 8, 16 and 24 weeks, change in the median daytime symptom score during weeks 5-16, for incidence of symptom-free days and incidence of days free from rescue bronchodilator usage during weeks 17-24. There was no significant difference between FP and BUD with respect to the number of patients experiencing one or more asthma exacerbation (33.8 and 28.4% of patients, respectively). There was, however, evidence that the exacerbations were clinically less severe in patients treated with FP, in that the time to resolution was quicker (11.0 vs. 14.7 days; P = 0.035), mean duration of all exacerbations (for an individual patient) tended to be shorter (18.5 vs. 23.6 days; P = 0.12), the time off work was reduced (4.2 vs. 7.6 days; P = 0.012) and the lowest PEF recorded during the exacerbation was higher (301 vs. 263 l min-1; P = 0.07). There were no clinically relevant differences in the safety (serum cortisol levels, markers of bone turnover, adverse events) of FP and BUD at these microgram equivalent doses. The patients recruited into this study, in retrospect, probably had no need for such high doses of inhaled corticosteroid but, irrespective of this, FP at microgram equivalent doses showed evidence of superior efficacy to BUD with respect to lung function and severity of asthma exacerbations without producing any greater adverse systemic effect. PMID- 10542975 TI - The role of fibre-optic bronchoscopy in solid organ, transplant patients with pulmonary infections. AB - Organ transplantation is currently the standard therapy for patients with end stage organ dysfunction. The immunosuppression caused by this therapy increases the rate of infection, particularly in the lungs. Early diagnosis is extremely important and fibre-optic bronchoscopy is a helpful tool in reaching diagnosis. Knowing the timing of various pathogens following transplantation, and the radiological picture as well as the prophylactic regimen, is helpful when specific pathogens are suspected. Bronchoscopy with bronchoalveolar lavage and transbronchial biopsies are particularly helpful in diagnosis of bacterial cytomegalovirus (CMV) and pneumocytosis carinii pneumocytosis, and is considered a safe procedure. Open lung biopsy is reserved for those who have negative bronchoscopy with a reasonable prognosis. PMID- 10542976 TI - Evaluation of bronchodilator response in patients with airway obstruction. AB - The aim of this study was to define the most useful index of expressing bronchodilator response and to distinguish between asthma and COPD. A prospective study was carried out of bronchodilator response in 142 asthmatics and 58 COPD patients in a university hospital. Reversibility was expressed as: 1. absolute change (delta abs); 2. % of initial (delta %init); 3. % of predicted (delta %pred) and 4. % of maximum possible response (delta %max). Dependence on forced expirations volume in 1 sec (FEV1) as % of predicted and sensitivity and specificity for diagnosis of asthma were established. A relationship between delta abs and initial FEV1 was not found in asthma (delta abs vs. % initial FEV1. r = 0.07) or COPD (r = 0.02). delta %pred did not show a correlation in asthma (r = 0.10) or COPD (r = 0.06). delta %init was dependent on the baseline value in asthma (r = 0.38, P < or = 0.001) but not in COPD (r = 0.18, P = n.s.). delta max was dependent in both. The combination of best sensitivity and specificity to separate asthma and COPD was obtained with delta abs (70.4 or 70.6%). The worst specificity for asthma diagnosis was obtained with delta %init (50%). The best likelihood ratios were obtained with delta abs and delta %pred and the worst likelihood ratio with delta %init. delta %init is not recommended as an index for differential diagnosis between asthma and COPD; 2) delta %init overscores bronchodilator response in patients with low FEV1. The independence of each bronchodilator response index should be verified in clinical trials for each selected sample. PMID- 10542977 TI - Role of the endogenous prostaglandin E2 in human lung fibroblast interleukin-11 production. AB - Interleukin-11 (IL-11) is known to be a member of the interleukin-6 (IL-6)-type cytokine family. IL-11 is likely to be a major determinant of immune regulation in acute and chronic inflammatory lung diseases, although it is not directly linked with specific disease processes. It has already been shown that although unstimulated human lung fibroblasts did not produce significant amounts of IL-11, the addition of interleukin-1 alpha (IL-1 alpha) and/or transforming growth factor-beta (TGF-beta) stimulated fibroblasts dose-dependently to produce IL-11. Northern blot analysis showed that these stimulators also upregulated IL-11 mRNA expression. As it has been previously reported that IL-1 and TGF-beta stimulate prostaglandin E2 (PGE2) release from lung fibroblasts, we investigate here the role of endogenous PGE2 and the direct effects of the two inhibitors of prostaglandin synthesis, indomethacin and dexamethasone, on IL-11 production by human lung fibroblasts. The addition of indomethacin, a cyclo-oxygenase inhibitor, resulted in significant suppression of IL-11 production and mRNA expression in lung fibroblasts. There was no detectable effect of PGE2 alone on IL-11 levels; however, the suppression of IL-11 production by indomethacin was almost completely reversed by addition of PGE2. In contrast, suppression of IL-11 production by indomenthacin was not reversed by addition of thromboxane B2 and carbocyclic thromboxane A2. In addition, dexamethasone completely suppressed IL 11 production and downregulated IL-11 mRNA. These results suggest that endogenous PGE2 acts as an autocrine stimulus for IL-11 production by human lung fibroblasts activated by IL-1 alpha and TGF-beta. PMID- 10542978 TI - Volume calibration alone may be misleading. AB - The use of spirometry is becoming more and more widespread in non-laboratory situations such as general practice or occupational medicine. In these non laboratory situations, volume calibration with a 3000 ml syringe is often the only feasible method to ensure that the spirometer produces valid and reproducible data. Sophisticated equipment to calibrate forced manoeuvres with standard waveforms are not present. In this study, we assessed whether volumetric calibration is a guarantee for valid and comparable spirometric results. Two portable spirometers were tested. On 8 consecutive test days, both spirometers were calibrated with a 3000 ml syringe in accordance with the American Thoracic Society (ATS) guidelines. The comparability of the spirometric results (forced expiratory volume in 1 S, FEV1) was tested in two ways. Firstly, the spirometers were compared to each other using the results from 43 volunteers on the same 8 test days. The spirometers were presented in a randomized order and volunteers were asked to perform a series of reproducible manoeuvres in both spirometers. Paired observations were analysed, using Bland and Altman plots. Secondly, the spirometers were compared to a 'gold standard', a computer-driven syringe (CDS). Calibration with the 3000 ml syringe showed that both spirometers complied with the ATS criteria for volume calibration for diagnostic spirometry. However, paired FEV1 data obtained in subjects showed a systematic, volume-dependent difference between the two spirometers (mean difference: 289 ml, P < 0.001, systematic difference: 8.6%, P < 0.0001). This systematic difference was confirmed by the comparisons with the CDS. Volume calibration may be misleading. The results from volume calibration may meet the ATS criteria, but this is no guarantee that data from forced manoeuvres are accurate. If CDS equipment to simulate standard wave forms is not available, it is recommended that biological calibration is performed regularly and, if possible, that paired data from two (or more) different spirometers are compared.